PMID- 15650281 TI - Synchronous pancreatic metastases from asymptomatic renal cell carcinoma. AB - CONTEXT: Pancreatic involvement from metastatic renal cell carcinoma accounts for 0.25-3% of the cases. The distinction between primary pancreatic clear cell tumour and metastatic deposits from renal cell carcinoma may lead to diagnostic difficulties. Usually metastasis from renal cell carcinoma presents many years after the primary tumour has been resected. A typical metastatic lesion in pancreas is seen as hyper-vascular lesion on CT scan. Any such lesion should be first biopsied before resection. CASE REPORT: We report herein the case of a patient with a solitary pancreatic metastatic head lesion from renal cell carcinoma was successfully resected before asymptomatic primary could be identified. CONCLUSIONS: Early identification of pancreatic metastasis from renal cell carcinoma is possible with characteristic appearance on radiological imaging and surgical resection of the metastasis provides better results. PMID- 15650282 TI - Pancreatic pseudoaneurysm of the superior mesenteric artery complicated with obstructive jaundice. A case report. AB - CONTEXT: Pancreatic pseudoaneurysm rupture is a rare complication of chronic pancreatitis, with severe prognosis and high mortality. Angiography is usually required for confirmation of the diagnosis, but transabdominal ultrasound and CT angiography are useful noninvasive diagnostic methods. CASE REPORT: We present the case of a 66-year-old patient with a large pancreatic pseudoaneurysm of the superior mesenteric artery complicated with obstructive jaundice. Transabdominal ultrasound with color and power Doppler showed a large pancreatic head pseudoaneurysm that communicated directly to the superior mesenteric artery. Presence of a spinning blood flow inside the pseudoaneurysm was visualized by color Doppler, with evidence of bidirectional flow in the pseudoaneurysm neck that was showed by Doppler spectral analysis. The contrast-enhanced helical computer tomography with multiplanar sagittal and the 3D reconstruction of coronal images confirmed the communication of the pseudoaneurysm with the superior mesenteric artery. The patient was scheduled for selective angiography and embolization. However, clinical evolution was rapidly deteriorating, with collapse, hemorrhagic shock and massive hemorrhage. The patient was operated on and subsequently died despite three days of intensive care, due to disseminated intravascular coagulation and multiorgan failure. CONCLUSION: Non invasive imaging methods consisting of transabdominal ultrasound with color Doppler and contrast-enhanced computer tomography with sagittal reconstruction of coronal images were very useful in the establishment of the diagnosis of pancreatic pseudoaneurysm of the superior mesenteric artery complicated with cholestatic jaundice. However, these imaging methods do not obviate the need for diagnostic and therapeutic angiography, eventually followed by surgical intervention in cases of recurrent bleeding or hemodynamic unstable patients. PMID- 15650283 TI - Fascioliasis: an exceptional cause of acute pancreatitis. AB - CONTEXT: Fasciola hepatica is known to cause bile duct inflammation and biliary obstruction but is rarely reported as responsible for producing acute pancreatitis. CASE REPORT: We report on a patient complaining of acute pancreatitis. Endoscopic retrograde cholangio-pancreatography showed distinct features and sphincterotomy allowed extraction of multiple parasites. CONCLUSIONS: Pancreatitis may occur in some patients with fascioliasis, but the condition may be overlooked in chronic cases. Endoscopic retrograde cholangio pancreatography rule out other possible causes of irregularity and thickening of the common bile duct wall. Parasite removal during endoscopic retrograde cholangio-pancreatography is one therapeutic option. Hepatic involvement must be ruled out and complete pharmacological treatment is advised in this patient. PMID- 15650284 TI - Pancreatic portal cavernoma. PMID- 15650285 TI - Some more news on the metastatic pathway in pancreatic cancer. PMID- 15650286 TI - Proceedings of the 28th National Congress of the Italian Association for the Study of the Pancreas. Verona, Italy. October 28-30, 2004. PMID- 15650287 TI - Genetics of chronic pancreatitis. PMID- 15650288 TI - Genetics of pancreatic cancer: where are we now? Where are we going? PMID- 15650289 TI - Alcohol and chronic pancreatitis: leading or secondary etiopathogenetic role? PMID- 15650290 TI - The various imaging aspects of chronic pancreatitis. PMID- 15650291 TI - Autoimmune pancreatitis: etiology, pathogenesis, clinical findings and treatment. The Japanese experience. AB - A concept of "autoimmune pancreatitis" has recently been proposed. Computed tomography, magnetic resonance imaging or ultrasonography can demonstrate the diffusely enlarged pancreas with its so called "sausage-like" appearance. Hypergammaglobulinemia, increased serum levels of total IgG or IgG4, positive autoantibodies such as antinuclear antibody, anti-lactoferrin antibody, anti-CA II antibody and rheumatoid factor have often been observed in patients with autoimmune pancreatitis. Microscopic findings have shown fibrotic changes involving infiltration of lymphocytes and plasmacytes, and often obliterative phlebitis in the pancreas. The major lymphocytes infiltrating the zone around the pancreatic duct were T cells producing IFN-g. HLA-DR was also expressed on the pancreatic duct and acinar cells as were lymphocytes. It is important to make the diagnosis of a diffusely enlarged pancreas based on clinical laboratory findings and pancreatic imaging such as narrowing pancreatogram. Laboratory data, pancreatic images and diabetes mellitus in most patients improved after steroid treatment. In conclusion, autoimmune pancreatitis appears to be a unique clinical entity. PMID- 15650292 TI - Autoimmune pancreatitis: pathological findings. AB - In recent years, autoimmune pancreatitis has been established as a special type of chronic pancreatitis. It is characterized by its histopathological and immunological features. The morphological hallmarks are periductal infiltration by lymphocytes and plasma cells and granulocytic epithelial lesions with consequent destruction of the duct epithelium and venulitis. Autoimmune pancreatitis has therefore also been called lymphoplasmacytic sclerosing pancreatitis, duct-destructive chronic pancreatitis, or sclerosing pancreatitis. Autoimmune pancreatitis most commonly involves the head of the pancreas and the distal bile duct. Occasionally, masses are formed and it has been described as an inflammatory myofibroblastic tumor. PMID- 15650293 TI - Autoimmune pancreatitis: instrumental diagnosis. PMID- 15650294 TI - Intraductal papillary mucinous tumors of the pancreas: incidence, clinical findings and natural history. PMID- 15650295 TI - Intraductal papillary mucinous tumors of the pancreas. Surgical treatment: at what point should we stop? AB - The intraoperative management of the margins of intraductal papillary mucinous tumors (IPMNs) undergoing pancreatic resection is crucial. The surgeon must discontinue the resection whenever the pancreatic margin is negative and, of course, when a total pancreatectomy is indicated.Nevertheless, a wide gray area exists. The real surgical problems are represented by i) IPMNs involving only a segment of the pancreatic gland, thus necessitating intraoperative histological examination of the resection and the decision as to "when to stop the resection"; ii) the intraoperative management of those margins which are neither clearly negative nor clearly positive; iii) the actual indications either for surgery or for follow-up in those patients affected by peripheral IPMNs. In the literature, negative resection margins have a range of between 49 to 81% with a pancreatic recurrence rate of from 0 to 25% in follow-ups ranging from 6 months to 11 years after the first operation. In general, in this disease which mainly involves the head and uncinate process of the gland, decisions are a "balance" between the patient and the disease. In fact, on the one hand, there is the usual elderly patient with possible comorbidities, symptoms and the presence of diabetes; on the other hand, the disease, which usually involves the head and the uncinate process of the gland, tends to grow along the duct and be potentially malignant if carcinoma is not already present. PMID- 15650296 TI - New frontiers in the neoadjuvant therapy of pancreatic adenocarcinoma: apart from therapeutical protocols. PMID- 15650297 TI - Sketches of otohistory. Part 6: Gustaf Retzius. PMID- 15650298 TI - Distortion product otoacoustic emissions after intratympanic gentamicin therapy for unilateral Meniere's disease. AB - The treatment of patients with Meniere's disease that do not respond to conventional therapy becomes complicated, particularly when taking into account the issue of hearing damage as well as the control of vertigo. Treatment often involves the intratympanic administration of gentamicin, for which different protocols are used. Hence, it is important that we better understand how this treatment influences hearing, beyond mere audiometric assessments. The aim of this prospective study was to evaluate the effect of intratympanic gentamicin treatment for Meniere's disease on cochlear function, as assessed by otoacoustic emissions. The 41 patients included in the study had been diagnosed with unilateral Meniere's disease as defined by the American Academy of Otolaryngology Head and Neck Surgery guidelines (1995), and had been refractory to medical treatment for at least 1 year. Intratympanic injections of gentamicin at a concentration of 27 mg/ml were performed at weekly intervals until indications of vestibular hypofunction appeared in the treated ear. Before beginning the treatment and 3 months after ending it, pure tone and speech audiometry tests were performed and the results are expressed in terms of the pure tone average (0.5, 1, 2, and 3 kHz) and the speech discrimination score, respectively. At the same time, a distortion product otoacoustic emission (DPOAE) study was performed and the results are expressed in terms of its presence or absence, and as the amplitude and threshold of the emission. When analyzed 3 months after the treatment had terminated, hearing loss was seen in 13 patients (31.7%). However, no significant change in the threshold and/or amplitude of otoacoustic emissions was observed in any of the patients. Neither were changes in the audiometric stage, number of injections required or the existence of DPOAE before treatment detected. Hence, the treatment method used here for patients with intractable unilateral Meniere's disease can be considered as having a low risk on auditory function, as assessed both audiometrically and with otoacoustic emissions, and can be considered as subablative for hearing function. PMID- 15650299 TI - Serial audiometry and speech recognition findings in Finnish Usher syndrome type III patients. AB - Audiometric features, evaluated by serial pure tone audiometry and speech recognition tests (n = 31), were analysed in 59 Finnish Usher syndrome type III patients (USH3) with Finmajor/Finmajor (n = 55) and Finmajor/Finminor (n = 4) USH3A mutations. These patients showed a highly variable type and degree of progressive sensorineural hearing impairment: from normal to moderate USH2A-like hearing impairment at young ages to profound or even USH1B-like hearing impairment at more advanced ages. Compound heterozygous patients generally showed a milder phenotype. The highest progression was seen during the first two decades of life, gradually slowing down with further ageing. This type of non-linear progression may be unique amongst the Usher syndromes. Speech recognition started to deteriorate at highly variable ages. In some patients, it jeopardised normal speech and language development, whereas in others it was still remarkably good at advanced ages. PMID- 15650300 TI - Morphological characteristics of the human macula sacculi. AB - The surface morphology of 20 human maculae sacculi is presented. Individual data on the total area, shape and the relative area of the pars superior are given. The mean area of 14 adult maculae was (mean +/- SD) 2.35 +/- 0.31 mm2. The ratio between the length and width of the macula was 2.54 +/- 0.28. The pars superior was significantly larger than the pars inferior. The percentage of pars superior was 56.4 +/- 4.7. As regards the shape of the macula, there was a large interindividual variability. The findings are discussed, taking into consideration comparative anatomy as well as spatial orientation. PMID- 15650301 TI - Aging effects on vestibulo-ocular responses in C57BL/6 mice: comparison with alteration in auditory function. AB - Age-related changes in auditory function are well documented in animal models; however, this is not the case as regards vestibular function. In this study, we evaluated age-related changes in vestibulo-ocular responses in C57BL/6 mice that are considered as a model of presbycusis. The functional data were substantiated by the findings of histological analysis of vestibular and auditory peripherals. The gain in vestibulo-ocular reflex, which reflects functionality of the vestibular system, increased in an age-dependent manner until 12 weeks and exhibited limited functional loss due to aging after 24 weeks. By contrast, no alteration in the thresholds of the auditory brainstem response (ABR) was observed from 3 to 12 weeks of age; however, ABR thresholds were significantly elevated from age 24 weeks and onwards. Histological analysis demonstrated that the degeneration of auditory peripherals was closely related with functional loss due to aging. Vestibular peripherals also exhibited age-related degeneration morphologically, although age-related dysfunction was not apparent. Age-related changes in the vestibular function of C57BL/6 mice followed a different time course when compared to changes in auditory function. These findings indicate that mechanisms for age-related changes in vestibular function differ from those of auditory function. PMID- 15650303 TI - The cognition and behaviour of children with cochlear implants, children with hearing aids and their hearing peers: a comparison. AB - Hearing impairment has been associated with cognitive deficits. It is not yet clear whether cochlear implants enable children to reduce or overcome these deficits. This study examined whether hearing impaired children with cochlear implants differed from hearing impaired children without cochlear implants or from hearing children. The three age-matched groups were compared on a non-verbal measure of cognition, the Leiter International Performance Scale-Revised (LIPS R), and a measure of behaviour, the Child Behaviour Checklist (CBCL). Apart from their scores on the Attention Sustained Subtest of the LIPS-R, the performance of the implanted group and the hearing group closely corresponded, while the hearing impaired children without cochlear implants performed below the level achieved by both these groups. There were no differences between any groups on the CBCL. It was concluded that hearing impaired children with cochlear implants were able to perform at the same non-verbal cognitive level as hearing children. PMID- 15650302 TI - Normative findings of electrically evoked compound action potential measurements using the neural response telemetry of the Nucleus CI24M cochlear implant system. AB - One hundred and forty-seven adult recipients of the Nucleus 24 cochlear implant system, from 13 different European countries, were tested using neural response telemetry to measure the electrically evoked compound action potential (ECAP), according to a standardised postoperative measurement procedure. Recordings were obtained in 96% of these subjects with this standardised procedure. The group results are presented in terms of peak amplitude and latency, slope of the amplitude growth function and ECAP threshold. The effects of aetiological factors and the duration of deafness on the ECAP were also studied. While large intersubject variability and intrasubject variability (across electrodes) were found, results fell within a consistent pattern and a normative range of peak amplitudes and latencies was established. The aetiological factors had little effect on the ECAP characteristics. However, age affected ECAP amplitude and slope of the amplitude growth function significantly; i.e., the amplitude is higher in the lowest age category (15-30 years). Principal component analysis of the ECAP thresholds shows that the thresholds across 5 electrodes can be described by two factors accounting for 92% of the total variance. The two factors represent the overall level of the threshold profiles ('shift') and their slopes across the electrode array ('tilt'). Correlation between these two factors and the same factors describing the T- and C-levels appeared to be moderate, in the range of 0.5-0.6. PMID- 15650304 TI - Laryngeal mask airway used as a delivery conduit for the administration of surfactant to preterm infants with respiratory distress syndrome. AB - BACKGROUND: The laryngeal mask airway (LMA(TM), Laryngeal Mask Co. Ltd, Jersey, UK) is a supraglottic device used to administer positive pressure ventilation (PPV) in adults, pediatric and neonatal patients. OBJECTIVES: To avoid endotracheal intubation, we evaluated the feasibility and practicality of administering surfactant via the LMA(TM) in preterm infants with respiratory distress syndrome (RDS). METHODS: Infants less than 72 h old with a gestational age of < or =35 weeks and a birth weight of >800 g, treated with nasal continuous positive airway pressure (CPAP, 5 cm H2O) for RDS were eligible for inclusion in the study if the arterial-to-alveolar oxygen tension ratio (a/APO2) was <0.20 over a period of >60 min. RESULTS: Eight preterm infants, median gestational age 31 (range 28-35) weeks; birth weight 1,700 (880-2,520) g, treated with nasal CPAP for RDS were enrolled. Three hours after surfactant instillation, the mean a/APO2 was significantly increased (0.13 +/- 0.04 to 0.34 +/- 0.11; p < 0.01) without complications. CONCLUSIONS: The LMA may be a useful and noninvasive conduit for the administration of surfactant therapy. A large randomized comparative clinical trial will be required to confirm the efficacy of this technique. PMID- 15650305 TI - Intravenous immunoglobulin g attenuates pulmonary hypertension but induces local neutrophil influx in meconium aspiration in piglets. AB - BACKGROUND: Pulmonary hypertension and inflammation are well-identified pathogenetic features in meconium aspiration syndrome of newborns, but current approaches to their treatment or prevention are still often unsatisfactory. OBJECTIVES: To investigate the possible protective effects of human intravenous immunoglobulin G (IVIG) on the hypertensive and inflammatory lung injury in severe neonatal meconium aspiration. METHODS: Eleven newborn (10-12 days old) ventilated and catheterized piglets that received an intratracheal bolus (3 ml/kg) of a 65-mg/ml mixture of human meconium were studied for 6 h. IVIG was infused in 5 piglets 30 min before meconium administration, and 6 piglets served as controls and received the vehicle only. RESULTS: Meconium instillation induced a biphasic pulmonary hypertensive response, which was significantly diminished by IVIG pretreatment. Similarly, IVIG improved the oxygenation of the piglets, but the intrapulmonary shunt fraction or systemic hemodynamic parameters did not differ between the study groups, except of a minor decrease in the mean arterial blood pressure caused by IVIG. The blood leukocyte count was comparable in the 2 groups. The lung tissue ultrastructural and histological changes, number of apoptotic cells and phospholipase A2 activity were similar in the 2 groups. The amount of neutrophil accumulation, assessed by myeloperoxidase activity, was however significantly increased in macroscopically damaged lung tissue after IVIG administration. CONCLUSIONS: Our results thus indicate that IVIG treatment of newborns with severe meconium aspiration significantly diminishes the pulmonary hypertensive response and improves oxygenation, but the effects do not extend to protection of lung cellular injury. PMID- 15650307 TI - Evaluation of cross-reactivity between Holoptelea integrifolia and Parietaria judaica. AB - BACKGROUND: Holoptelea integrifolia and Parietaria judaica belong to the family Urticaceae, but are geographically distantly located. H. integrifolia is an important pollen allergen of India and sensitizes almost 10% of the atopic population in Delhi. P. judaica, on the other hand, is a very dominant pollen allergen of the Mediterranean region, sensitizing almost 80% of the allergic population. Since both these important pollen allergens belong to the family Urticaceae, the objective of the present study was to assess cross-reactivity between these two pollen allergens from different geographical regions. METHODS: Cross-reactivity between these two pollen allergens was assessed on the basis of skin prick tests and ELISA, ELISA inhibition and immunoblot inhibition studies. RESULTS: Out of 44 atopic Indian patients skin prick tested with H. integrifolia extract, 34% were found to be sensitized. All the patients sensitized to H. integrifolia also showed varying degrees of skin positivity to P. judaica pollen extract. ELISA and ELISA inhibition studies suggested strong cross-reactivity between H. integrifolia and P. judaica pollen. Immunoblot inhibition studies revealed that 14-, 16-, 28-, 38-, 42- and 46-kDa proteins are the cross-reactive proteins in H. integrifolia and P. judaica. However, Par j 1, the major allergen of P. judaica, is absent in H. integrifolia pollen. CONCLUSION: H. integrifolia and P. judaica pollens share cross-reactive as well as unique epitopes. The major allergen of P. judaica, Par j 1, seems to be absent in H. integrifolia pollen allergen. PMID- 15650306 TI - Botulinum toxin: mechanisms of action. AB - Botulinum toxin (BT) has been perceived as a lethal threat for many centuries. In the early 1980s, this perception completely changed when BT's therapeutic potential suddenly became apparent. We wish to give an overview over BT's mechanisms of action relevant for understanding its therapeutic use. BT's molecular mode of action includes extracellular binding to glycoprotein structures on cholinergic nerve terminals and intracellular blockade of the acetylcholine secretion. BT affects the spinal stretch reflex by blockade of intrafusal muscle fibres with consecutive reduction of Ia/II afferent signals and muscle tone without affecting muscle strength (reflex inhibition). This mechanism allows for antidystonic effects not only caused by target muscle paresis. BT also blocks efferent autonomic fibres to smooth muscles and to exocrine glands. Direct central nervous system effects are not observed, since BT does not cross the blood-brain barrier and since it is inactivated during its retrograde axonal transport. Indirect central nervous system effects include reflex inhibition, normalisation of reciprocal inhibition, intracortical inhibition and somatosensory evoked potentials. Reduction of formalin-induced pain suggests direct analgesic BT effects possibly mediated by blockade of substance P, glutamate and calcitonin gene-related peptide. PMID- 15650308 TI - The major grass pollen group 5 allergen from Dactylis glomerata and its C terminal split product both behave as dimers: implications for allergen standardization. AB - BACKGROUND: On SDS-PAGE grass pollen group-5 allergens migrate as a doublet with an apparent molecular mass (M(r)) of 25 kDa. Immunoblot analysis revealed additional group 5 reactivity at double and half this M(r). The aim of this study was to investigate these group 5 molecular entities and to compare their allergenicity and behavior in quantitative immunoassays. METHODS: Group-5 specific monoclonal antibodies were produced and used for the development of a group-5-specific sandwich ELISA. Affinity-purified Dac g 5 was separated by SDS PAGE/Western blotting; individual bands were analyzed by N-terminal sequencing. Size exclusion chromatography (SEC) in conjunction with group-5-specific ELISA, competitive RIA and RAST inhibition were used to analyze the size distribution of Dac g 5. Basophil histamine release assays were used to assess biological activity. RESULTS: The lower band of the typical group 5 doublet was identified as a truncated form lacking the typical group 5 N-terminus AD(L)/(A)GY, observed in the upper band. The 12-kDa peptide was shown to be the C-terminal half of Dac g 5 (amino acid 127 onwards). SEC in conjunction with competitive RIA revealed that around 45% of Dac g 5 is represented by the 12-kDa peptide. Both the C terminal half and the whole allergen dimerize under nondenaturing conditions. In competitive RIA and RAST inhibition both forms are equally well detected. In contrast, the half molecule is poorly recognized in sandwich ELISA and displays negligible biological activity in basophil histamine release tests with purified IgE. CONCLUSIONS: These observations stress the need to evaluate the performance of allergen standardization protocols in detail, with special attention to allergen size distribution. PMID- 15650309 TI - Biochemical and immunological characterization of pollen-derived beta galactosidase reveals a new cross-reactive class of allergens among Mediterranean trees. AB - BACKGROUND: The most potent allergens in the Spermatophytae family exhibit significant homology with enzymes. Some of these are though to be involved in pectin metabolism, recognition of compatible stigma and delivery of sperm cells to the ovule. OBJECTIVE: To test if glycohydrolase activities from some Mediterranean tree pollens could act as allergens in sensitized hosts. METHODS: Freshly collected Cupressus and Olea pollens were investigated for their glycohydrolase activities by means of synthetic fluorogenic substrates and isoenzymes characterized by DEAE-cellulose ion-exchange chromatography. Binding of specific IgE was investigated by immunoblotting in 30 tree-sensitive subjects, as well as in 20 atopic non-tree-sensitive and 15 healthy controls. The enzymes were also adopted to stimulate proliferation of allergen-specific T cell clones. Finally, they were tested in vivo in a cutaneous immediate wheal and flare reaction. RESULTS: beta-Galactosidase (beta-GAL) is present with different isoenzymatic patterns on both pollen extracts, could be recognized by circulating IgE, as well as immunoprecipitated by sera from allergic subjects. The enzyme could stimulate the proliferation of T cells from allergic subjects, and favor the emergence of CD4+ T cell clones with specific in vitro reactivity to beta GAL. Finally, the enzyme induced in vivo a cutaneous wheal and flare reaction in clinically sensitive subjects. CONCLUSIONS: Despite different isoenzymatic patterns, Olea-derived beta-GAL cross-reacted with that from cypress pollen, suggesting that these enzymatic glycoproteins may represent major native allergens among these Mediterranean trees. PMID- 15650310 TI - Lack of detectable alterations in immune responses during sublingual immunotherapy in children with seasonal allergic rhinoconjunctivitis to grass pollen. AB - BACKGROUND: Recent work indicates that subcutaneous specific immunotherapy induces specific T-cell anergy, a shift in the TH1/TH2 ratio, and antibody production in favor of IgG4. There are few data on sublingual immunotherapy (SLIT), especially in children. METHODS: We assessed the proliferation of peripheral blood mononuclear cells ((3)H-thymidine incorporation) and secretion of interleukin (IL)-4, interferon (IFN)gamma and IL-5 (ELISA) after in vitro stimulation with allergen or phytohemagglutinin (PHA) in 29 children with allergic rhinoconjunctivitis receiving SLIT with grass pollen before, and after 1 and 2 years of treatment in a multicenter placebo-controlled study on the efficacy of the treatment. Further, non-specific intracellular production of IL 4, IL-13, IFNgamma, IL-2, IL-10 and IL-5 (FACS) and serum total and specific IgE and IgG4 (ELISA) were analyzed. RESULTS: Proliferation and IL-4 and IL-5 secretion after stimulation with allergen or PHA did not differ between the groups. In addition, we observed no effect of SLIT on intracellular cytokine production. IFNgamma secretion after allergen coculture was comparable between the groups. Following PHA stimulation, IFNgamma secretion was significantly higher in the SLIT group after 1 year, and a trend was observable already before and after 2 years of treatment, probably due to the inhomogeneity in the groups despite randomization (for age and asthma). No significant changes were observed for sIgE/sIgG4 ratios over time either in or between the groups. CONCLUSION: During 2 years of SLIT in children with a positive effect on rescue medication use, we observed no significant effects on in vitro T-cell immune responses or immunoglobulins. So far, pediatric studies demonstrating stable effects of SLIT on such reactions are missing, probably due to limited effects of SLIT on systemic immunologic reactions. PMID- 15650311 TI - Immunosuppressive effect of restraint stress on the initiation of allergic rhinitis in mice. AB - BACKGROUND: Exposure to acute stressors modulates both innate and acquired immune function. However, little is known about whether stress has the potential to modulate the pathogenesis of allergic rhinitis. OBJECTIVES: To determine the effects of acute restraint stress on the initiation of allergic rhinitis in a murine model. METHODS: CBA/J mice were repeatedly intranasally sensitized with phospholipase A2 (PLA2) from honeybee venom without adjuvant. Restraint stress was applied using uniform cylinders once a week for a continuous 8-hour period, on five occasions in total. Production of PLA2-specific antibodies and degree of nasal and blood eosinophilia were compared between stressed and control mice. RESULTS: Repeated intranasal sensitization with PLA2 induced PLA2-specific IgE and marked eosinophilia in both the nose and blood in CBA/J mice. Exposure to restraint stress significantly inhibited production of PLA2-specific IgE, IgG1 and IgG2a. Conversely, the stress exerted no significant effect on eosinophilia. CONCLUSIONS: Exposure to acute restraint stress inhibits antigen-specific antibody production, but not local or systemic eosinophilia. The results of this study suggest that acute stress has the potential to modulate the initiation of allergic rhinitis. PMID- 15650313 TI - Anaphylactic reaction to inulin: first identification of specific IgEs to an inulin protein compound. AB - BACKGROUND: A woman with a past history of allergy to artichoke presented with two episodes of immediate allergic reactions, one of which was a severe anaphylactic shock after eating two types of health foods containing inulin. RESULTS: Dot blot assay techniques identified specific IgEs to artichoke, to yoghurt F, and to a heated BSA + inulin product. Dot blot inhibition techniques confirmed the anti-inulin specificity of specific IgE. CONCLUSIONS: The absence of a positive reaction to an unheated milk-inulin mixture indicates the probability of protein-inulin binding. There is no cross-reactivity with the carbohydrates of the glycosylated allergens. PMID- 15650312 TI - Fexofenadine improves the quality of life and work productivity in Japanese patients with seasonal allergic rhinitis during the peak cedar pollinosis season. AB - BACKGROUND: Although currently in its infancy, quality of life (QOL) research in Japan is rapidly expanding and is expected to become a standard outcome measure in clinical trials. In Japan, QOL has not previously been assessed in patients with allergic rhinitis (AR); we report the first clinical study applying the recently validated Japanese translations of the Rhinoconjunctivitis Quality of Life Questionnaire (RQLQ) and Work Productivity and Activity Impairment-Allergy Specific (WPAI-AS) Questionnaire to assess the effects of the oral antihistamine, fexofenadine, on QOL and work productivity due to cedar pollinosis. PATIENTS AND METHODS: A randomized, double-blind, placebo-controlled, single-site study was conducted during the peak cedar pollinosis season in Japan. After a 7-day run-in period, subjects were randomized to receive fexofenadine HCl 60 mg twice daily (bid) or placebo for 2 weeks. RESULTS: Overall, 206 Japanese subjects with AR were included in the intention-to- treat population (fexofenadine, n = 104, and placebo, n = 102). Fexofenadine statistically significantly improved overall QOL compared with placebo (p = 0.005) and improvements were reported in the RQLQ domains: activities (p = 0.047), practical problems (p = 0.003), nasal symptoms (p = 0.003) and eye symptoms (p 2)-beta-glucopyranoside], (20(R)-Rg3) and 20-S proto-panaxatriol-3-[O-beta-D-glucopyranosyl (1-->2)-beta-glucopyr-anoside], (20(S)-Rg3) in regulating voltage-dependent Ca2+, K+ or Na+ channel currents and 5-HT3A and a3b4 nicotinic acetylcholine (nACh) receptor channel currents expressed in Xenopus oocytes. 20(S)-Rg3 but not 20(R)-Rg3 inhibited the Ca2+, K+ and Na+ channel currents in a dose- and voltage-dependent manner. The fact that only 20(S)-Rg3 is active indicates that its hydroxyl group may be geometrically better aligned with the hydroxyl acceptor group in the ion channels than that of 20(R)-Rg3. However, both Rg3 stereoisomers inhibited 5-HT3A and a3beta4 nACh receptor channel currents. These results indicate that the selectivity of action of the Rg3 stereoisomers differs between voltage-dependent and ligand-gated ion channels. PMID- 15650338 TI - Enhanced expression of a gene encoding a nucleoside diphosphate kinase 1 (OsNDPK1) in rice plants upon infection with bacterial pathogens. AB - A cDNA library was constructed using mRNA extracted from rice leaves infected with Xanthomonas oryzae pv. oryzae (Xoo), a bacterial leaf blight pathogen, to isolate rice genes induced by Xoo infection. Subtractive hybridization and differential screening of the cDNA library led to the isolation of many induced genes including a nucleotide diphosphate kinase 1 (OsNDPK1) and a pathogenesis related protein 1 (OsPR1) cDNA. Nucleoside diphosphate kinases (NDPKs) are key metabolic enzymes that maintain the balance between cellular ATP and other nucleoside triphosphates (NTPs). Three other OsNDPK genes (NP922751, OsNDPK2 and OsNDPK3) found in databases were obtained by RT-PCR. Three different programs for predicting subcellular targeting indicated that OsNDPK1 and NP922751 were non organellar, OsNDPK2 plastidic, and OsNDPK3 mitochondrial. Only transcripts of OsNDPK1 accumulated strongly after infection with Xoo. When rice plants were infected with Burkholderia glumae, a bacterial grain/seedling rot pathogen, the pattern of expression of the rice NDPK genes was similar to that following infection with Xoo. OsNDPK1 gene expression was also strongly induced in response to exposure to salicylic acid, jasmonic acid, and abscisic acid, although the level of transcripts and their pattern of expression depended on the inducer. PMID- 15650340 TI - Toward optimal health: the experts discuss important issues in hormone replacement and in diabetes. Interview by Jody R. Godfrey. PMID- 15650341 TI - The hypoglycemic effects of soy isoflavones on postmenopausal women. AB - OBJECTIVES: Soy isoflavones have many effects similar to those of estrogen and have become popular among postmenopausal women as an alternative for hormone replacement therapy (HRT). The purpose of this study was to determine the effects of soy isoflavones on glucose, insulin, and lipid profiles in postmenopausal Taiwanese women. METHODS: We conducted a randomized, double-blind, active placebo controlled clinical trial to compare the effects of isoflavones with estrogen replacement therapy (ERT) on blood glucose, insulin, and lipid profiles in postmenopausal Taiwanese women. Thirty postmenopausal Taiwanese women were randomly assigned to two groups, and each received treatment for 6 months. The isoflavone group received 100 mg isoflavone soft capsules, 300 mg calcium, and a blank vitamin capsule per day. The estrogen active control group received 0.625 mg conjugated estrogen, 300 mg calcium, and blank isoflavone soft capsules per day. We measured baseline levels of fasting blood sugar, insulin, lipid profiles, and isoflavone concentrations prior to the study and repeated the same measurements every 3 months for a total duration of 6 months. RESULTS: Two-way ANOVA revealed that fasting glucose (p < 0.001) and insulin (p < 0.005) levels were significantly affected by estrogen and isoflavone treatments after 6 months. For the isoflavone group, the average blood genistein concentration was 6-10 times higher than those of the estrogen group. Within the same periods, the fasting blood glucose was reduced to 83% and 85% of the baseline levels, and insulin was reduced to 56% and 67% of the baseline levels, respectively, in the estrogen and isoflavone groups. CONCLUSIONS: Soy isoflavones (100 mg) and 0.625 mg conjugated estrogen equally lower fasting blood glucose and insulin levels in postmenopausal women. PMID- 15650342 TI - Identification of women's coronary heart disease and risk factors prior to first myocardial infarction. AB - PURPOSE: To understand when women's coronary heart disease (CHD) and CHD risk factors are recognized prior to first myocardial infarction (MI). METHODS: Medical record review of the 10 years prior to incident MI among women with a confirmed incident MI between January 1, 1996, and December 31, 2001, to determine the timing of CHD diagnosis as well as assessment and treatment for risk factors. RESULTS: One hundred fifty women had incident MIs during the study period. They made 8732 ambulatory visits and had 457 hospitalizations during the period of review (mean 9.1 years, range 6.2-10 years). Average age at incident MI was 74.7 years (SD 12.6, range 38.9-99.8 years). A CHD diagnosis prior to first MI was present in 52% (n = 78) of the women but was less common in those <70 years (p = 0.001). All but 3 women had one or more modifiable risk factors identified prior to their first MI. Treatment of recognized risk factors varied from 81% (antihypertension medications) to only 28% (drug therapy for abnormal lipid levels). Having a diagnosis of CHD was associated with an increased likelihood of having identified risk factors and receiving drug treatment for identified risk factors. CONCLUSIONS: Women with undiagnosed CHD (48%) and those with unrecognized or untreated risk factors for CHD, especially younger women, represent missed opportunities for prevention of cardiac events. PMID- 15650343 TI - Consistency in efficacy and safety of ezetimibe coadministered with statins for treatment of hypercholesterolemia in women and men. AB - BACKGROUND: Women are often not treated as aggressively as men to control levels of low-density lipoprotein cholesterol (LDL-C), despite evidence that women and men realize comparable cardiovascular benefit from lipid-lowering therapy. Statins are the most effective drugs currently available for treating hypercholesterolemia. Despite the impressive cholesterol-lowering capacity of statins, however, many patients on statin therapy fail to reach established target levels of LDL-C. The cholesterol absorption inhibitor, ezetimibe, blocks the intestinal absorption of dietary and biliary cholesterol, a mechanism of action complementary to that of statins, which inhibit hepatic cholesterol synthesis. Ezetimibe coadministered with statins produces significant incremental reductions in LDL-C compared with statin monotherapy. METHODS: Four randomized, double-blind, placebo-controlled, balanced-parallel group trials compared the efficacy and safety of statin monotherapy (lovastatin or pravastatin 10, 20, or 40 mg; simvastatin or atorvastatin 10, 20, 40, or 80 mg) vs. ezetimibe 10 mg plus statin (as above). A gender subset analysis (women, n = 1065; men, n = 796) on data pooled across these four trials was carried out to determine whether ezetimibe plus statin for the treatment of hypercholesterolemia is equally efficacious in women and men. RESULTS: Compared with statin monotherapy, ezetimibe plus statin demonstrated greater efficacy in reducing blood levels of LDL-C, apolipoprotein B, and triglycerides and raising high-density lipoprotein cholesterol. The beneficial effects of ezetimibe were comparable in women and men. The safety profile of subjects receiving ezetimibe plus statin was similar to that of patients receiving statin monotherapy and similar between the two sexes. CONCLUSIONS: Ezetimibe plus statin was more effective than statin alone in improving the lipid profile of patients with hypercholesterolemia and was equally efficacious in women and men. Ezetimibe plus statin was well tolerated and had a favorable safety profile in both patient subgroups. Ezetimibe coadministered with statins, a dual inhibition treatment strategy that targets both cholesterol absorption and synthesis, is an effective therapeutic option for women with hypercholesterolemia. PMID- 15650344 TI - Women's employment status and mortality: the atherosclerosis risk in communities study. AB - BACKGROUND: As women's labor force participation in the United States has increased over the past decades, there has been an interest in the potential health effects of employment. To date, however, research findings have been contradictory. METHODS: Thus, the aim of this study was to investigate the association between employment status and mortality among 7361 middle-aged African American and white women who participated in the Atherosclerosis Risk in Communities (ARIC) Study. Women were classified as employed or homemakers at the baseline examination (1987-1989) and were followed for approximately 11 years. Proportional hazards regression was used to estimate unadjusted and adjusted hazard ratios. RESULTS: After adjusting for sociodemographic factors and selected risk factors for mortality, employed women had a lower risk of mortality than homemakers (hazard ratio [HR] = 0.65, 95% confidence interval [CI] = 0.49, 0.86). This decreased risk of mortality persisted in additional analyses that excluded those who died within the first 2 years of follow-up or, alternatively, those with a history of coronary heart disease (CHD), stroke, cancer, hypertension, diabetes, or a perception of fair or poor health at baseline. In cause of death specific analyses, the mortality advantage among employed women persisted for circulatory system-related deaths; however, the association for cancer-related deaths was weaker, and the CI included one. CONCLUSIONS: As the association between employment status and mortality was not explained by known risk factors for mortality, additional research is needed to identify other potential factors that may help to explain this relationship. PMID- 15650345 TI - The influence of natural menopause on postprandial lipemia in heterozygotes for familial hypercholesterolemia. AB - BACKGROUND: Heterozygous familial hypercholesterolemia (hFH) is a genetic disease that leads to premature atherosclerosis. Natural menopause leads to an adverse lipid profile and an enhanced risk of coronary heart disease (CHD). Raised plasma triglyceride (TG) levels also contribute to the risk of vascular events. The aim of this study was to evaluate the postprandial TG levels (after a standardized fatty meal) in premenopausal and postmenopausal women with hFH. METHODS: Thirty three Greek women with hFH were divided into the premenopausal group--n = 16, mean age 34(SD = 7), mean total cholesterol = 330(30) mg/dl--and the postmenopausal group--n = 17, mean age 62(5), mean total cholesterol = 346(63) mg/dl. Plasma TG concentrations were measured before and 2, 4, 6, and 8 hours after a standardized fat load. A value of >219 mg/dl (2.5 mmol/L) was taken as an abnormal response to the fat load, according to our previous studies. RESULTS: Postmenopausal women had higher TG levels at 2 (p = 0.001), 4 (p = 0.003), 6 (p = 0.003), and 8 hours (p = 0.005) after the fatty meal compared to premenopausal women. Forty-one percent of postmenopausal hFH women had abnormal TG response (hFH-A) after a fatty meal, and such women had higher fasting TG levels than postmenopausal hFH women with a normal response to the fatty meal (hFH-N) (p = 0.0014). CONCLUSIONS: Women with hFH tend to have an abnormal TG response to a fatty meal after the menopause. Fasting TG levels may be able to predict the abnormal response to a fatty meal. PMID- 15650346 TI - Adolescent issues associated with knowledge of and access to topical microbicides. AB - PURPOSE: Topical microbicides could be a female-controlled method for prevention of sexually transmitted diseases (STDs). Microbicides could offer girls a potential preventive option, if they are able to access them. The purpose of this investigation was to evaluate girls', mothers', experienced healthcare providers', and medical students' views on adolescent girls' obtaining information about and acquiring microbicides and where adolescent girls would keep or carry a microbicide. METHODS: Focus groups were conducted and videotaped with each group of individuals. The videotapes were transcribed, and the transcriptions were coded for relevant themes. RESULTS: Participants anticipated that adolescents would learn about microbicides from a variety of sources and that specific educational efforts for each of these sources (mothers, healthcare providers, and school-based sexuality education) would be needed. Healthcare providers stressed the importance of enhancing access by having both prescription and over-the-counter (OTC) options. For nonprescription products, the participants felt that the microbicide should be in stores and venues to which adolescents typically go by themselves. A microbicide that was girl-friendly and adolescent-friendly consistently was highlighted by the participants. Participants thought this would increase discretion and decrease embarrassment. The location of keeping and carrying the microbicide was anticipated to be dependent on parents' knowledge of their adolescent's sexuality and the likelihood that parents search their adolescent's belongings. CONCLUSIONS: Consideration of the adolescent-specific issues in the design of microbicides and marketing strategies may be crucial for adolescent adoption of this female controlled method. PMID- 15650347 TI - Recruitment of minority women and their main sexual partners in an HIV/STI prevention trial. AB - Recruiting heterosexual couples into randomized clinical trials (RCTs) to test the efficacy of HIV/sexually transmitted infection (STI) prevention interventions is a challenge that requires innovative strategies and consideration of ethical issues, including participant safety and confidentiality. METHODS: This paper provides a brief review of the literature on minority and couple RCT recruitment and describes the development (preparation phase and protocol development) and implementation (strategies employed and barriers) of a recruitment protocol that safely enrolled 217 predominantly African American and Latino heterosexual couples into a relationship-based, HIV/STI prevention study. RESULTS: The success of this recruitment protocol with no reported adverse events demonstrates the feasibility of engaging urban minority women and men in RCTs. This study builds on a small literature base articulating specific couple recruitment strategies. CONCLUSION: More research delineating and testing specific strategies for recruiting defined populations into clinical trials is needed to advance the science of study recruitment and improve generalizability of research findings. PMID- 15650349 TI - Consortium for the Assessment of Research on Progestins and Estrogens (CARPE) Fort Worth, Texas August 1-3, 2003. PMID- 15650348 TI - Aerobic exercise and lipids and lipoproteins in women: a meta-analysis of randomized controlled trials. AB - BACKGROUND: Cardiovascular disease (CVD) in women is the leading cause of mortality in the United States, and less than optimal lipid and lipoprotein levels are major risk factors for CVD. The purpose of this study was to use the meta-analytic approach to examine the effects of aerobic exercise on lipids and lipoproteins in women. METHODS: Studies were retrieved via computerized literature searches, review of reference lists, hand searching selected journals, and expert review of our reference list. The inclusion of studies was limited to randomized controlled trials published in the English language literature between January 1955 and January 2003 in which aerobic exercise was used as the primary intervention in adult women aged > or =18 years. One or more of the following lipids and lipoproteins were assessed: total cholesterol (TC), high-density lipoprotein cholesterol (HDL-C), low-density lipoprotein cholesterol (LDL-C), and triglycerides (TG). RESULTS: Using a random effects model, statistically significant improvements were observed for all lipids and lipoproteins (TC, +/- SEM, -4.3 +/- 1.3 mg/dl, 95% CI -6.9 to -1.7 mg/dl; HDL-C, +/- SEM, 1.8 +/- 0.9 mg/dl, 95% CI 0.1 to 3.5 mg/dl; LDL-C, +/- SEM, -4.4 +/- 1.1 mg/dl, 95% CI -6.5 to -2.2 mg/dl; TG, +/- SEM, -4.2 +/- 2.1 mg/dl, 95% CI -8.4 to -0.1 mg/dl). Reductions of approximately 2%, 3%, and 5%, respectively, were observed for TC, LDL-C, and TG, whereas an increase of 3% was observed for HDL-C. CONCLUSIONS: Aerobic exercise is efficacious for increasing HDL-C and decreasing TC, LDL-C, and TG in women. PMID- 15650351 TI - Modeling the TSH Receptor. PMID- 15650352 TI - Analysis of the thyrotropin receptor-thyrotropin interaction by comparative modeling. AB - We have used the most advanced programs currently available to construct the first three-domain structure of the human thyrotropin receptor (TSHR) using comparative modeling. The model consists of a leucine-rich domain (LRD; amino acids 36-281; porcine ribonuclease inhibitor used as a template for modeling), a cleavage domain (CD; amino acids 282-409; tissue inhibitor of matrix metalloproteinases 2 as template) and transmembrane domain (TMD amino acids 410 699; bovine rhodopsin as template). Models of human, porcine, and bovine TSH were also constructed (human chorionic gonadotropin [hCG] and human follicle stimulating hormone [hFSH] as templates). The LRD has a characteristic horseshoe shape with 10 tandem homologous repeats. The CD consists of beta-barrel and alpha helix structures (OB-like fold) with two disulfide bridges and the structure around these disulfide bridges remains stable after cleavage. The TMD presents the typical seven membrane-spanning helices. The TSH, LRD, CD, and TMD models were brought together in an extensive series of docking experiments. Known features of the TSH-TSHR interaction were used for selection of appropriate complexes that were then validated using a different set of experimental data. A similar approach was used to build a model of a complex between the TSHR and a monoclonal TSHR antibody with weak thyroid stimulating activity. Human thyrotropin (hTSH) alpha chains were found to make contact with many amino acids on the LRD surface and CD surface whereas no interaction between the beta chains and the CD were found. The higher affinity of bovine thyrotropin (bTSH) and porcine thyrotropin (pTSH) (relative to hTSH) for the TSHR is explained well by the models in terms of charge-charge interactions between their alpha chains and the receptor. Experimental observations showing increased sensitivity of the TSHR to hCG after mutation of TSHR Lys209 to Glu are explained well by our model. Furthermore, several mutations in the TMD that are associated with increased TSHR basal activity are predicted from our model to be caused by the formation of new interactions that stabilize the activated form of the TMD. PMID- 15650353 TI - Relative potencies and additivity of perchlorate, thiocyanate, nitrate, and iodide on the inhibition of radioactive iodide uptake by the human sodium iodide symporter. AB - The presence of perchlorate (ClO(4) (-)) in some U.S. drinking water supplies has raised concern about potential adverse thyroidal health effects, because ClO(4) ( ) is known to competitively inhibit iodide uptake at the sodium iodide symporter (NIS). Humans are nutritionally and environmentally exposed to other competitive inhibitors of iodide uptake, including thiocyanate (SCN(-)) and nitrate (NO(3) ( )). The joint inhibiting effects of these three anions was studied by exposing Chinese hamster ovary cells stably expressing human NIS to varying concentrations of each anion separately, and in combination, and conducting measurements of (125)I(-) uptake. The entire data set was fit to a single Hill equation using maximum likelihood. The relative potency of ClO(4) (-) to inhibit (125)I(-) uptake at the NIS was found to be 15, 30 and 240 times that of SCN(-), I(-), and NO(3) (-) respectively on a molar concentration basis, with no evidence of synergism. These results are consistent with a common mode of action by these anions of simple competitive interaction, in which a concentration of any one of ClO(4) (-) SCN(-), and NO(3) (-), occurring either individually or as part of a mixture of the three anions, is indistinguishable from a concentration or dilution of either one of the remaining two ions in inhibiting iodine uptake at the NIS. PMID- 15650354 TI - Immunohistochemical and sequencing analyses of the Wnt signaling components in Japanese anaplastic thyroid cancers. AB - We investigated the status of the components and target genes of the Wnt signaling pathway in Japanese anaplastic thyroid cancers (ATCs) in the present study. Nuclear and cytoplasmic positive staining of beta-catenin, which might indicate the existence of alterations in the Wnt signaling pathway, were found in 40.9% and 63.6% of the 22 ATC samples, respectively. The beta-catenin, adenomatous polyposis coli (APC) and Axin 1 gene mutations were observed in 4.5%, 9.0%, and 81.8% of the 22 ATC samples, respectively. Overexpression of cyclin D1 and c-myc, which are the target genes of the Wnt signaling pathway, was observed in 27.3% and 59.1% of the ATC samples, respectively. There was no significant correlation between nuclear or cytoplasmic positive staining of beta-catenin and nuclear positive staining of cyclin D1 or c-myc. Taken together, the results of beta-catenin immunohistochemistry suggest that alterations in the Wnt signaling pathway are associated with carcinogenesis of ATC, but the frequency of beta catenin gene mutation in our series is lower than that previously reported. Furthermore, cyclin D1 and c-myc frequently accumulated in ATC, independently of dysfunction in the Wnt signaling pathway. PMID- 15650355 TI - The nature of analogue-based free thyroxine estimates. AB - Clinical laboratories often use analogue-based immunoassays to estimate serum free thyroxine (FT(4)) concentrations. These assays yield FT(4) estimates that correlate closely with thyroxine (T(4)) binding protein concentrations. This correlation implies that either T(4) binding proteins or protein bound T(4) contribute to analogue-based FT(4) values. To study the contributions made by T(4) binding proteins to these FT(4) estimates further, four analogue-based FT(4) assays were applied to: (1) FT(4) solutions without T(4) binding proteins, (2) to T(4) binding protein solutions without T(4), and (3) to total T(4) solutions containing T(4) binding protein, FT(4), and protein-bound T(4). The FT(4) estimates obtained with these solutions ranged from 0.2-8.6 ng/dL, when FT(4) concentrations ranged from less than 0.2-12,000 ng/dL. In the FT(4) solutions, gravimetrically determined FT(4) concentrations were 500-12,000 ng/dL (0.5-12.0 microg/dL) without protein-bound T(4), and the FT(4) estimates obtained were 0.3 6.9 ng/dL. In the total T(4) solutions, dialyzable FT(4) concentrations were less than 0.2-59 ng/dL, retained T(4) concentrations were 499.8-11,441 ng/dL, and the analogue-based FT(4) estimates obtained were 0.2-8.6 ng/dL. Similar FT(4) estimates (0.2-8.6 ng/dL and 0.3-6.9 ng/dL) were obtained with similar concentrations of either protein-bound T(4) or FT(4). Similar test results were associated with similar total T(4) concentrations, not similar FT(4) concentrations. Protein-bound T(4) and T(4) binding protein contributed variably to test results. T(4) quantifications included large analytical losses that are unaccounted for. These assays passed tests of correlation with FT(4) concentrations, but they failed tests of specificity for FT(4) and accuracy in T(4) quantification. PMID- 15650356 TI - Cytoplasmic localization of the paired box gene, Pax-8, is found in pediatric thyroid cancer and may be associated with a greater risk of recurrence. AB - The paired box-8 protein (Pax-8) has been observed in the nucleus of normal adult thyroids, follicular adenomas, follicular thyroid cancers, and papillary thyroid cancers (PTC) but not undifferentiated thyroid cancers. To our knowledge, Pax-8 has not been studied in pediatric thyroid cancer. Because of the more favorable prognosis for PTC in children compared to young patients, we hypothesized that Pax-8 expression might be different in pediatric thyroid cancers. To test this, we stained 47 thyroid lesions from children and young patients for Pax-8. Pax-8 was located in the cytoplasm (cPAX) or nucleus (nPAX) in the majority of samples. There was no significant difference in nPAX between benign and malignant lesions. However, cPAX was more commonly seen in PTC than autoimmune diseases (p = 0.01) and the intensity of cPAX staining correlated with tumor size (p = 0.041), metastasis, age, completeness of resection, local invasion, and tumor size (MACIS) scores (p = 0.045), and the presence of invasion, metastasis, recurrence, or persistence (p = 0.012). Disease-free survival was significantly reduced for cancers with intense cPAX staining (p = 0.0003). These data show that cPAX is common in PTC, and although limited by small sample size, suggest an association with higher MACIS scores, an aggressive clinical course, and an increased risk of clinically evident recurrence for children and young patients. PMID- 15650357 TI - Stress and thyroid autoimmunity. AB - While many studies have shown a connection between stress and autoimmune disease, most of the evidence for stress contributing to the onset and course of autoimmune disease is circumstantial and the mechanisms by which stress affects autoimmune disease are not fully understood. The best circumstantial evidence for an effect of stress on autoimmune thyroid disease is the well-known relationship between the onset of Graves' hyperthyroidism and major stress but even this is debated. However, most of the recent case-control studies have supported stress as a factor that affects the onset and clinical course of Graves' disease. On the other hand, there have been few reports concerning the possible relationship between stress and Hashimoto's thyroiditis. Because the onset and course of Hashimoto's thyroiditis is generally insidious, the effect of stress on Hashimoto's thyroiditis might be overlooked. Numerous human and animal studies have demonstrated that psychological and physiologic stressors induce various immunologic changes. Stress affects the immune system either directly or indirectly through the nervous and endocrine systems. These immune modulations may contribute to the development of autoimmunity as well as the susceptibility to autoimmune disease in genetically predisposed individuals. Stress can be one of the environmental factors for thyroid autoimmunity. PMID- 15650358 TI - Increased incidence of thyroid carcinoma in france: a true epidemic or thyroid nodule management effects? Report from the French Thyroid Cancer Committee. AB - Thyroid cancer (TC) incidence, in France, has dramatically increased over the last two decades. In order to try and account for this observation, the French Department of Health requested the Public Health Agency to coordinate a multidisciplinary Thyroid Cancer Committee (TCC). The TCC analysed the temporal incidence trend in France, evaluated the contribution of changes in diagnostic practices of thyroid diseases to the observed increase of TC, and set up guidelines to improve the national surveillance system of TC. The increased incidence of TC is real (8.1% and 6.2% per year in women and in men, respectively), mainly due to papillary type with an epidemic of microcarcinomas (43% of operated cancers, period 1998-2001) associated to the extensiveness of thyroidectomies. Multicentric studies showed a significant increase, from 1980 to 2000, in ultrasonographic (3 to 84.8%) and cytological procedures (8 to 36% of patients with thyroid nodules) as well as a significant association between the increase in TC prevalence among operated patients (12.5 to 37%) and the spread of fine needle aspiration. Epidemiological evidence does not favour any link with the Chernobyl accident. The TCC recommended a national registry dedicated to thyroid cancer of the youths ( <18 years old). For adults, in addition with the strengthening of the French regional registries, a continuous registration of incident cases through the National Hospital Discharge Survey that covers all the territories is proposed. Such system, matched with pathological data derived from a national standardized collection, will provide a relevant model for epidemiological surveys of TC. PMID- 15650359 TI - Chromosomal imbalances in post-chernobyl thyroid tumors. AB - Tissue samples from 60 post-Chernobyl childhood thyroid tumors have been investigated. We used comparative genomic hybridization (CGH) to detect chromosomal gains and losses within the tumor DNA. This is the first CGH study on childhood thyroid tumors. The post-Chernobyl tumors showed chromosomal imbalances in 30% of tumors. The most frequent DNA copy number changes in post-Chernobyl tumors involved chromosomes 2, 7q11.2-21, 13q21-22, 21 (DNA gains), and chromosomes 16p/q, 20q, 22q (DNA losses). Some of these specific alterations detected in post-Chernobyl thyroid tumors (deletions on chromosomes 16p/q and 22q) have previously been reported in thyroid tumors as associated with an aggressive biologic behavior and may therefore also account for the more aggressive phenotype of papillary thyroid carcinoma (PTC) found in post- Chernobyl tumors. Eighteen percent of post-Chernobyl PTC that exhibit RET rearrangements also showed chromosomal imbalances indicating that either additional genetic events are involved in this subset of tumors, or that intratumoral genetic heterogeneity exists in these tumors, suggesting a oligoclonal pattern to tumor development. PMID- 15650360 TI - Neural network analysis for evaluating cancer risk in thyroid nodules with an indeterminate diagnosis at aspiration cytology: identification of a low-risk subgroup. AB - Thyroid nodules with a predominant follicular structure are often diagnosed as indeterminate at fine-needle aspiration biopsy (FNAB). We studied 453 patients with a thyroid nodule diagnosed as indeterminate at FNAB by using a feed-forward artificial neural network (ANN) analysis to integrate cytologic and clinical data, with the goal of subgrouping patients into a high-risk and in a low-risk category. Three hundred seventy-one patients were used to train the network and 82 patients were used to validate the model. The cytologic smears were blindly reviewed and classified in a high-risk and a low-risk subgroup on the basis of standard criteria. Neural network analysis subdivided the 371 lesions of the first series into a high-risk group (cancer rate of approximately 33% at histology) and a low-risk group (cancer rate of 3%). Only cytologic parameters contributed to this classification. Analysis of the receiver operating characteristic (ROC) curves demonstrated that the ANN model discriminated with higher sensitivity and specificity between benign and malignant nodules compared to standard cytologic criteria (p < 0.001). This value did not show degradation when ANN predictions were applied to the validation series of 82 nodules. In conclusion, neural network analysis of cytologic data may be a useful tool to refine the risk of cancer in patients with lesions diagnosed as indeterminate by FNAB. PMID- 15650361 TI - High-intensity focused ultrasound for localized thyroid-tissue ablation: preliminary experimental animal study. AB - OBJECTIVE: Thyroid surgery is common and complications are not rare. High intensity focused ultrasound (HIFU) could be a possible minimally invasive alternative to surgery. The aim of this study was to assess the feasibility of using HIFU to obtain localized ablation of thyroid tissue without affecting neighboring structures. METHODS: The ewe was chosen as the model because its thyroid is easily accessible, with a size comparable to the human gland. An HIFU device designed to treat human prostate cancer was used. Eight ewes were anesthetised and their thyroids were ablated with ultrasound-guided HIFU. HIFU was generated by a 3-MHz spherical piezocomposite transducer that delivered an average of 24 (range, 9-44) ultrasound pulses per lobe covering a mean volume of 0.7 cm(3) (range, 0.2-1.8). RESULTS: Ewes were sacrificed 6-13 days after HIFU treatment and the anterior part of the neck was fixed in formalin before macroscopic and microscopic examinations. Adverse events occurred mainly for the first treated ewes before the energy delivered to the thyroid tissues was fully mastered: one animal died 3 days after HIFU, most probably because of inhalation pneumonia; the ultrasound beam hit adjacent organs in three animals. As expected, typical histologic lesions of the thyroid were obtained: central coagulative necrosis with ghost vesicular structures, disappearance of the nuclei, and cytoplasmic flocculation. At the periphery of the necrotic zone, a cell reaction was observed with fibroblastic granulation tissue, mononuclear cell infiltrate and regenerating thyroid tissue. CONCLUSION: The results of this preliminary study confirm the possibility of using HIFU in order to destroy a defined area in thyroid tissue. Future experiments in ewes will be focused on the safety of the method by refining the HIFU parameters and by developing a new equipment specially built for the thyroid. PMID- 15650362 TI - Transient hypothyroidism or persistent hyperthyrotropinemia in neonates born to mothers with excessive iodine intake. AB - Perinatal exposure to excess iodine can lead to transient hypothyroidism in the newborn. In Japan, large quantities of iodine-rich seaweed such as kombu (Laminaria japonica) are consumed. However, effects of iodine from food consumed during the perinatal period are unknown. The concentration of iodine in serum, urine, and breast milk in addition to thyrotropin (TSH), free thyroxine (FT(4)), and thyroglobulin was measured in 34 infants who were positive at congenital hypothyroidism screening. Based on the concentration of iodine in the urine, 15 infants were diagnosed with hyperthyrotropinemia caused by the excess ingestion of iodine by their mothers during their pregnancy. According to serum iodine concentrations, these infants were classified into group A (over 17 microg/dL) and group B (under 17 microg/dL) of serum iodine. During their pregnancies these mothers consumed kombu, other seaweeds, and instant kombu soups containing a high level of iodine. It was calculated that the mothers of group A infants ingested approximately 2300-3200 microg of iodine, and the mothers of group B infants approximately 820-1400 microg of iodine per day during their pregnancies. Twelve of 15 infants have required levo-thyroxine (LT(4)) because hypothyroxinemia or persistent hyperthyrotropinemia was present. In addition, consumption of iodine by the postnatal child and susceptibility to the inhibitory effect of iodine may contribute in part to the persistent hyperthyrotropinemia. We propose that hyperthyrotropinemia related to excessive iodine ingestion by the mother during pregnancy in some cases may not be transient. PMID- 15650363 TI - Trimester-specific changes in maternal thyroid hormone, thyrotropin, and thyroglobulin concentrations during gestation: trends and associations across trimesters in iodine sufficiency. AB - OBJECTIVES: To describe the interrelationships of thyroid functions based on trimester-specific concentrations in healthy, iodine-sufficient pregnant women across trimesters, and postpartum. METHODS: Circulating total 3,5,3'- triidothyronine (T(3)) and thyroxine (T(4)) concentrations were determined simultaneously using liquid chromatography tandem mass-spectrometry (LC/MS/MS). Free thyroxine (FT(4)), thyroid-stimulating hormone (TSH), and thyroglobulin (Tg) were measured using immunoassay techniques. Linear mixed effects models and correlations were calculated to determine trends and associations, respectively, in concentrations. RESULTS AND CONCLUSIONS: Trimester-specific T(3), FT(4), TSH, and Tg concentrations were significantly different between the first and third trimesters (all p < 0.05); second and third trimester values were not significantly different for FT(4), TSH, and Tg (all p > 0.25) although T3 was significantly higher in the third, relative to the second trimester. T(4) was not significantly different at any trimester (all p > 0.80). With two exceptions, analyte concentrations tended not to be correlated at each trimester and at 1 year postpartum. One exception was that T(3) and T(4) tended to be associated (all p < 0.05) at all time points except the third trimester (rho = 0.239, p > 0.05). T(4) and FT(4) concentrations tended to correlate positively during pregnancy (rho 0.361-0.382, all p < 0.05) but not postpartum (rho = 0.179, p > 0.05). Trends suggest that trimester-specific measurements of T(3), FT(4), Tg, and possibly TSH are warranted. PMID- 15650364 TI - Papillary thyroid carcinomas with lung metastases. AB - BACKGROUND: Limited information is available on the presentation, treatment outcomes, and prognostic factors of papillary thyroid carcinoma with lung metastases. METHODS: This study retrospectively analyzed the data of 2003 patients with thyroid cancer who were treated and followed up at Chang Gung Memorial Hospital from January 1979 to December 2002. In total, 1516 papillary thyroid carcinomas were enrolled. One hundred two (6.7%) papillary thyroid carcinomas with lung metastases were followed including 57 women with mean age of 41.7 +/- 17.3 years and 45 men with mean age of 49.6 +/- 17.2 years. These patients included 52 patients with papillary thyroid carcinoma who presented with lung metastases at the time of diagnosis. The 102 cases of papillary thyroid carcinomas with lung metastases included 72 cases with lung metastases only, and 30 cases with other organ involvement. RESULTS: After mean follow-up periods of 8.8 +/- 0.6 years, 28 (27.5%) of the patients with lung metastasis died, while 6 improved to clinical stage I. The 5-, 10-, 15-, and 20-year survival rates in patients with papillary thyroid carcinoma without distant metastasis and in the lung metastases groups were 99.0%, 98.5%, 98.0%, 98.0%, and 91.3%, 75.0%, 64.0%, 51.2%, respectively. Comparing the Kaplan-Meier survival curves between the patients with papillary thyroid carcinomas with lung metastases only and those with multiorgan metastases demonstrates no statistically significant difference in mortality rates. However, age, gender, postoperative thyroglobulin (Tg) level and tumor size displayed statistically significant differences between the lung metastases and no distant metastasis groups. Fifty of the 102 patients with papillary thyroid carcinomas with lung metastases developed lung metastases during follow-up. Larger amounts of remnant thyroid tissues with higher Tg levels were noted in these patients compared to those without distant metastasis. CONCLUSIONS: The prognosis of patients with papillary thyroid carcinoma with lung metastases at time of diagnosis is the same as for those whose lung metastases are discovered later. Survival analysis demonstrates no difference between lung metastases and multiorgan metastases. PMID- 15650365 TI - Graves' disease after interleukin-2 therapy in a patient with human immunodeficiency virus infection. AB - Interleukin-2 (IL-2) is a cytokine that regulates the proliferation and differentiation of lymphocytes, and is currently used clinically in the treatment of assorted malignancies. Additionally, IL-2 is being actively investigated in clinical trials for treatment of human immunodeficiency virus (HIV) infection. Patients treated with IL-2 are susceptible to autoimmune thyroid disease (AITD), presenting as thyroiditis, which leads to either thyrotoxicosis or hypothyroidism, if not correctly and promptly identified and treated. IL-2 induced hypothyroidism can also sometimes follow a thyrotoxic phase. However, the development of Graves' disease (GD) in this clinical setting has not been reported to date. Here, we report the case of a 39-year-old HIV-infected man in whom GD developed after IL-2 therapy. We correlated the immunologic parameters pertinent to the patient's HIV infection status with clinical, hormonal, and serologic evidence of GD during its emergence. This revealed an association between peripheral blood cell numbers of specific lymphocyte subpopulations (CD4(+), CD3(+)CD25(+), and naive T-cells) and serum levels of markers for AITD (free thyroxine [T(4)] and thyroid-stimulating immunoglobulin). Interestingly, no association was found between natural killer (NK) cell numbers and AITD markers. The immunopathogenesis of GD in this patient may be similar to that hypothesized for the GD that occurs in immune-reconstituted patients after combination antiretroviral therapy. From a practical standpoint, we propose that patients who have received or are receiving treatment with IL-2 who show signs of hyperthyroidism need to be carefully evaluated for GD. PMID- 15650366 TI - A giant cervical lymph node metastasis from thyroid papillary microcarcinoma considered as a parotid gland tumor in preoperative diagnosis. PMID- 15650367 TI - Thyroglossal duct cyst with papillary carcinoma. PMID- 15650370 TI - Three-dimensional model and molecular mechanism of Mycobacterium tuberculosis catalase-peroxidase (KatG) and isoniazid-resistant KatG mutants. AB - Mycobacterium tuberculosis KatG enzyme functions both as catalase for removing hydrogen peroxide (H(2)O(2)) and as peroxidase for oxidating isoniazid (INH) to active form of anti-tuberculosis drug. Although mutations in M. tuberculosis KatG confer INH resistance in tuberculous patients, structural bases for INH-resistant mutations in the KatG gene remains poorly understood. Here, three M. tuberculosis KatG mutants bearing Arg418--> Gln, Ser315 --> Thr, or Trp321 --> Gly replacement were assessed for changes in catalase-peroxidase activities and possible structure bases relevant to such changes. These three M. tuberculosis KatG mutants exhibited a marked impairment or loss of catalase-peroxidase activities. The possible structural bases for the mutant-induced loss of enzyme activities were then analyzed using a three-dimensional model of M. tuberculosis KatG protein constructed on the basis of the crystal structure of the catalase peroxidase from Burkholderia pseudomallei. The model suggests that three M. tuberculosis KatG mutants bearing Arg418 --> Gln, Ser315 -->Thr, or Trp321--> Gly replacement affect enzyme activities by different mechanisms, although each of them impacts consequently on a heme-associated structure, the putative oxidative site. Moreover, in addition to the widely accepted substrate-binding site, M. tuberculosis KatG may bear another H(2)O(2) binding site. This H(2)O(2) binding site appears to interact with the catalytic site by a possible electron-transfer chain, a Met255-Tyr229-Trp107 triad conserved in many catalase-peroxidases. The Ser315 --> Thr mutant may have direct effect on the catalytic site by interfering with electron transfer in addition to the previously proposed mechanism of steric constraint. PMID- 15650371 TI - Isoniazid resistance and the future of drug-resistant tuberculosis. AB - Bacterial chromosomal mutations that confer antibiotic resistance often have deleterious effects that impose costs on reproductive fitness. This observation has led to the generalization that in the absence of the selection pressure exerted through treatment, the frequency of resistance will decrease. This model implies that the prudent use of antibiotics will eventually result in a decline in the prevalence of drug resistance. Recent work, however, suggests that some resistance-conferring mutations may not significantly impair fitness and that others may be accompanied by compensatory mutations that restore the organisms' reproductive potential. Thus drug resistance, once introduced, may persist unless specific measures are implemented to target prevalent drug-resistant cases. Here we present ecological evidence to support the hypothesis that mutations at the 315 position of katG confer isoniazid resistance for Mycobacterium tuberculosis without diminishing virulence or transmissibility. PMID- 15650372 TI - Genotypic characterization of carbapenem-nonsusceptible Acinetobacter spp. isolated in Latin America. AB - The principal aim of this study was to evaluate the genomic diversity among the imipenem-nonsusceptible Acinetobacter spp. (INSA) collected from the Latin American medical centers within the SENTRY Antimicrobial Surveillance Program. The INSA isolates were collected from patients with bloodstream infections, who were hospitalized in seven Latin American countries between 1997 and 1999. For epidemiologic comparison, 20 carbapenem-susceptible Acinetobacter spp. (CSA) isolates were collected in the same period of time from the respective medical centers. A total of 23 Acinetobacter spp. isolates exhibiting imipenem MIC values of >/=8 microg/ml were typed by ribotyping, an automated molecular method. The isolates showing an identical ribogroup were also typed by pulsed-field gel electrophoresis (PFGE). The antimicrobial susceptibility to various antimicrobial agents was evaluated using a reference broth microdilution technique. Among the INSA isolates, 13 distinct ribogroups were observed, whereas 16 ribogroups were detected among the CSA. Nearly 57% of the INSA belonged to only four ribogroups. Identical ribogroups and PFGE patterns were observed among INSA and CSA isolates collected from medical centers located in different countries (Brazil and Argentina). Our results showed: (1) a higher genomic variability among the CSA; (2) presence of epidemic clones among INSA isolates encountered in Latin American medical centers; and (3) spread of INSA and CSA epidemic clones between Latin American countries. PMID- 15650373 TI - Variable resistance patterns of integron-associated multidrug-resistant Acinetobacter baumannii isolates in a surgical intensive care unit. AB - Between 1998 and 2000, we characterized 91 nosocomial isolates of Acinetobacter baumannii by antibiotyping and genotyping. A total of 25 ribotypes were obtained among these 91 isolates. When the isolates from surgical intensive care units (ICUs) and other wards were compared, multiresistant A. baumannii isolates with the same ribotype 25 (R-25) were significantly more prevalent in nosocomial infections among the surgical patients. Further subtyping of the strains by pulsed-field gel electrophoresis confirmed that strains of the same ribotype in surgical ICUs and a few isolates from other wards were identical or clonally related. Different antibiotic resistance profiles were observed among these R-25 isolates. All R-25 isolates contained intI1 integrase, and two clusters of integron cassettes were found. These clusters of cassettes were encoded by an open reading frame (ORF) of -5'CS-aac(3)Ia-aadA1a-unknown or f-3'CS or 5'CS-aacA4 aadA1-catB8-3'CS, indicating the involvement of different resistant genes. Two isolates contained bla (IMP-1), which was acquired from a conjugatively transferable plasmid and did not involve integron-associated resistance. In conclusion, an epidemic of nosocomial infections associated with A. baumannii strains that have different resistance profiles was identified. Resistance profiles can change by a combination of plasmid- and integron-associated acquisition, especially in a unit with high antibiotic selective pressures. Infectious control personnel should be alert to the change in resistance profiles during routine monitoring. PMID- 15650374 TI - Antibiotic resistance and genotypic characteristics of group a streptococci associated with acute pharyngitis in Korea. AB - Group A streptococci (GAS) is the most common pathogen of bacterial pharyngitis. GAS were isolated from pediatric outpatients with symptomatic acute pharyngitis. T typing and emm genotyping were performed for an epidemiologic study. The phenotypes of erythromycin (EM) resistance were evaluated, and the frequencies of ermB, ermTR(A), and mefA genes were determined by polymerase chain reaction. A total of 125 (50.8%) of 246 children yielded GAS. The two most frequent T types were T12 (35.2%) and T nontypeable (30.4%). The most frequent emm type was emm 12 (28.5%), followed by emm75 (18.7%), emm22 (13.0%), emm2 (12.2%), and emm8 (8.1%). The resistance rates to erythromycin and clindamycin were 44.8% and 19.2%, respectively. Among the EM-resistant strains, constitutive resistance, inducible resistance, and the M phenotype were observed in 42.1%, 0%, and 57.9%, respectively. The ermB, ermA, and mefA genes were present in 39.3%, 0%, and 58.9%, respectively. Most of the emm12 strains showed constitutive resistance, while emm18 and emm75 demonstrated the M phenotype. The organisms with other emm genotypes were susceptible to both EM and clindamycin. These results indicate that some emm genotypes might be associated with a specific mechanism of EM resistance. PMID- 15650375 TI - Polyclonal spread of erythromycin-resistant Streptococcus agalactiae in southern Taiwan. AB - Resistance to erythromycin is common among Streptococcus agalactiae in Taiwan, however the genetic relatedness of erythromycin-resistant isolates has not yet been reported. From 1991 to 2001, 629 clinical isolates of S. agalactiae were collected in a medical center at Tainan in southern Taiwan, of which 189 (30.0%) were resistant to erythromycin. The isolation rate of erythromycin-resistant group B streptococcus (GBS) was stable, irrespective of the clinical sources or study period. Among them, 145 (76.7%) isolates showed the macrolide-lincosamide streptogramin B (MLS)-resistant phenotype, and 44 (23.3%) had the macrolide (M)- resistant phenotype. Of the isolates with MLS phenotype, 141 (97.2%) isolates harbored the ermB gene alone and only three (2.1%) the ermTR gene, whereas 41 (93.2%) of 44 isolates with M phenotype harbored the mefA/E gene. Of 177 typeable isolates, there were 26 unrelated pulsed-field gel electrophoresis (PFGE) patterns. PFGE type 1 accounted for 17.8% (24/135) of MLS phenotype isolates with the ermB gene and 48.7% (18/37) of M phenotype isolates with the mefA/E gene. During the study period, the proportion of PFGE type 6 decreased significantly, whereas that of type 8 increased. Our results suggest that erythromycin resistance is not uncommon among clinical isolates of S. agalactiae and is, at least, partially related to polyclonal spread in southern Taiwan. PMID- 15650376 TI - Occurrence and Characteristics of Erythromycin-Resistant Streptococcus pneumoniae Strains Isolated in Three Major Brazilian States. AB - We investigated the occurrence and phenotypic and genotypic characteristics of erythromycin-resistant Streptococcus pneumoniae strains isolated in three major states in Brazil, from 1990 to 1999. Of the 931 pneumococcal strains evaluated, 40 (4.3%) were erythromycin-resistant (Ery-R). Among the 40 Ery-R strains, 90.0%, 80.0%, 27.5%, 5.0%, and 2.5% were resistant to tetracycline, trimethoprim sulfamethoxazole, penicillin, chloramphenicol, and rifampin, respectively. None of the strains were resistant to ofloxacin or to vancomycin. Most [37 (92.5%)] of the 40 Ery-R isolates presented the MLS(B) phenotype and 3 (7.5%) strains showed the M phenotype. PCR testing indicated that all MLS(B) phenotype isolates harbored the erm(B) gene only, whereas the mef(A/E) gene was present in all isolates presenting the M phenotype. The tet(M) gene was the most frequent (86.1%) among Ery-R isolates that were also resistant to tetracycline. Pulsed field gel electrophoresis (PFGE) analysis after SmaI digestion revealed the occurrence of clonal relationships within groups of strains belonging to serotypes 14, 19A, and 23F. All Ery-R isolates belonging to serotype 14 were susceptible to penicillin and were included in a single clonal group (named Ery(14)-A) related to the England(14-)9 internationally spread clone. PMID- 15650377 TI - Antibiotic resistance patterns of mecA-positive Staphylococcus aureus isolates from clinical specimens and nasal carriage. AB - The present study demonstrates that the nasal carriage rate of Staphylococcus aureus was 40% in Jordanian healthy young adult population, and 19% of nasal S. aureus and 57% of clinical isolates over the same period were resistant to oxacillin (MRSA), respectively. The mecA gene was detected in all MRSA isolates in both groups. Most of MRSA isolates were multiresistant to three antibiotic classes (beta-lactams, aminoglycosides, macrolides-lincosamides). This result suggests a serious problem may be encountered in treatment of staphylococcal infections in Jordan. PMID- 15650378 TI - Sequence analysis of rpoB mutations in rifampin-resistant clinical Mycobacterium tuberculosis isolates from Turkey. AB - Drug-resistant tuberculosis is a serious problem throughout the world. Resistance to Rifampicin (RIF) is mainly caused by the mutations in the rpoB gene coding the beta-subunit of RNA polymerase. In this study, we aimed to detect the distribution of rpoB gene mutations in 80 RIF-resistant clinical Mycobacterium tuberculosis (MTB) isolates from Turkey. The rpoB gene was amplified by PCR and mutations leading to RIF resistance were determined by automated sequence analysis. A total of 72 of the 80 isolates (90%) were found to carry mutations in the amplified region, whereas eight isolates (10%) carried no mutations. Overall, 24 different missense mutations affecting 14 codons, and two deletion mutants were identified. Nine new mutations, six in the hot-spot region and three outside this region, were found. The codon numbers of the most frequently encountered mutations were 531 (51.4%), 526 (18.1%), 516 (13.9%), and 513 (12.5%). As a result, 90% of the RIF-resistant MTB isolates from the Turkish patients were found to carry a mutation in the rpoB gene, Ser531Leu being the most frequent one. Although molecular methods identify mutations leading to RIF resistance very quickly, results of the antimycobacterial susceptibility tests must be taken into consideration for the patients carrying no mutations in this region. PMID- 15650379 TI - Prevalence of beta-lactamases among ampicillin-resistant Escherichia coli and Salmonella isolated from food animals in Denmark. AB - The genetic background for beta-lactamase-mediated resistance to beta-lactam antibiotics was examined by PCR and sequencing in 160 ampicillin-resistant isolates (109 Escherichia coli and 51 Salmonella) obtained from healthy and diseased food animals in Denmark. Sequencing revealed three different variants of bla (TEM-1), of which bla (TEM-1b) was the most frequently detected (80 E. coli and 47 Salmonella), followed by bla (TEM-1a) (eight E. coli, one Salmonella) and bla (TEM-1c) (seven E. coli). A few isolates were found to express OXA, TEM-30, or PSE beta-lactamases. Mutations in the ampC promoter leading to increased production of the AmpC beta-lactamase were demonstrated in 11 cefoxitin-resistant or intermediate E. coli isolates. Nine of these isolates did not contain any bla (TEM) genes, whereas the remaining two did. No genes encoding SHV or extended spectrum beta-lactamases were detected. Two new variants of bla (TEM) were detected, which have been designated bla (TEM-127) and bla (TEM-128). In TEM-127, amino acid 158 is substituted from His to Asn, whereas a substitution from Asp to Glu is seen at amino acid 157 in TEM-128. According to MIC determinations, these novel enzymes do not possess activity against extended-spectrum beta-lactams. PMID- 15650380 TI - Distribution of the erm (B) gene, tetracycline resistance genes, and Tn1545-like transposons in macrolide- and lincosamide-resistant enterococci from pigs and humans. AB - The distribution of the erm (B) and the tetracycline resistance genes tet(K), tet(L), tet(M), tet(O), and tet(S) was investigated among macrolide- and lincosamide-resistant enterococci originating from humans, pigs, and pork carcasses. The presence of transposons of the Tn916/Tn1545 family was also traced in these isolates. Furthermore, the porcine strains were tested for the presence of glycopeptide resistance genes vanA and vanB. The erm(B) gene was found in 85% of the porcine and in all human isolates. Ninety-eight percent of the porcine and 89% of the human erm(B)-positive enterococci carried the tet(M) gene. Seventy seven percent and 70%, respectively, of these strains harbored a Tn1545-like element. Tet(L) was observed in 68% of the porcine and in 65% of the human enterococci. The other tetracycline resistance genes were very rare and the glycopeptide resistance genes vanA and vanB were not detected among the porcine isolates. The similar frequencies of resistance genes and the highly mobile Tn1545-like transposon among porcine and human enterococci might indicate exchange of resistant strains or their resistance genes between humans and pigs or the existence of a common reservoir. PMID- 15650381 TI - A mouse peritonitis model for the study of glycopeptide efficacy in GISA infections. AB - In recent years, the emergence of Staphylococcus aureus strains with reduced susceptibility to glycopeptides has raised considerable concern. We studied the efficacy of vancomycin and teicoplanin, as well as cloxacillin and cefotaxime, against the infection caused by four S. aureus strains with different glycopeptide and beta-lactam susceptibilities (strains A, B, C, and D; MICs for vancomycin of 1, 2, 4, and 8 microg/ml respectively), using a modified model of mouse peritonitis. This optimized model appeared to be straightforward and reproducible, and was able to detect low differences in bacterial killing between antibiotics and also between different S. aureus strains. Bactericidal activities in peritoneal fluid for vancomycin, teicoplanin, cloxacillin, and cefotaxime decreased from -2.98, -2.36, -3.22, and -3.57 log(10) cfu/ml, respectively, in infection by strain A (MICs for vancomycin and cloxacillin of 1 and 0.38 microg/ml, respectively) to -1.22, -0.65, -1.04, and +0.24 in peritonitis due to strain D (MICs for vancomycin and cloxacillin of 8 and 1,024 microg/ml). Our data confirm the superiority of beta-lactams against methicillin-susceptible S. aureus and show that bactericidal activity of glycopeptides decreases significantly with slight increases in MICs; this finding suggests a reduced efficacy of glycopeptides in the treatment of serious glycopeptide-intermediate S. aureus infections. PMID- 15650382 TI - Outbreak of SHV-5 beta-lactamase-producing Klebsiella pneumoniae in a neonatal pediatric intensive care unit in Spain. AB - The objective was to analyze the beta-lactamase genes and the clonal relationship in a series of 12 clinical Klebsiella pneumoniae strains resistant to ceftazidime or cefotaxime (MIC >/=16 microg/ml) recovered in the neonatalpediatric intensive care unit (ICU) ward of a Spanish hospital during a 1-year period. TEM, SHV, CTX M, CMY, or FOX beta-lactamase genes were analyzed by PCR and sequencing. The clonal study was performed by pulsed-field gel electrophoresis (PFGE) using XbaI. All but one of the 12 K. pneumoniae strains harbored the bla (SHV-5) gene, and the bla TEM-1a gene was also detected in one of them. These 11 strains belonged to two different clonal types: A (9 strains) and B (2 strains) and were grouped in the subtypes A(1) (6 strains), A(2), A(3), A(4), B(1), and B(2) (1 strain each). The clonal type A strains were isolated from 9 patients (in five cases from blood) during a 6-month period. The remaining K. pneumoniae strain harbored both the bla (SHV-11) + bla (CTX-M-14) genes and showed the clonal type C. A nosocomial outbreak by a SHV-5-producing multiresistant K. pneumoniae is reported in Spain in a neonatal-pediatric ICU ward. This is the first description of a K. pneumoniae harboring both the bla (SHV-11) and bla (CTX-M-14) genes in Spain. PMID- 15650383 TI - Recurrent Klebsiella pneumoniae mycotic aneurysm in a diabetic patient and emergence of an extended-spectrum beta-lactamase (CTX-M-24)-containing Klebsiella pneumoniae strain after prolonged treatment with first-generation cephalosporins for mycotic aneurysm. AB - A 68-year-old diabetic woman suffered from mycotic aneurysm due to Klebsiella pneumoniae over her abdominal aorta; she received surgical intervention, followed by treatment with first-generation cephalosporins for 6 months. She was hospitalized again 11 months later because of another episode of mycotic aneurysm caused by K. pneumoniae on her thoracic aorta. Fingerprinting generated by pulsed field gel electrophoresis and infrequent-restriction-site polymerase indicated K. pneumoniae isolates of the identical clonal strain were responsible for these two mycotic-aneurysm episodes. Unfortunately, nosocomial pneumonia developed at the second hospitalization; blood and purposefully sampled feces specimen cultures both grew CTX-M-24-producing K. pneumoniae, which were of the same strain and genetically nonrelated to the K. pneumoniae strain causing mycotic aneurysms earlier. This is the first report on infection due to CTX-M-24-producing K. pneumoniae. It is unclear whether the prolonged use of first-generation cephalosporins in this case selected a strain of enteric organism possessing the ESBL in question, which was capable of passing this ESBL plasmid to the K. pneumoniae strain causing the nosocomial infection. This report suggests that further observation is needed before one can draw a conclusion on the possibility of the selection of ESBL enteric organism by extensive exposure to first generation cephalosporins. PMID- 15650384 TI - Tumors, wounds, and lymph. PMID- 15650385 TI - Limb lymph node response to bone fracture. AB - In previous clinical studies, dilation of afferent lymphatics and enlargement of inguinal lymph nodes (LN) were observed in lymphoscintigrams from patients with persistent posttraumatic edema of lower extremities after fractures and trauma of soft tissues. In this study, changes in rat popliteal and iliac lymph nodes draining lymph from the site of tibial fracture and adjacent soft tissue injury were investigated. The observed parameters were lymph node weight, cell number, phenotype frequency, cell cytokine expression, and reactivity to mitogens. The key observations included: a) increase in the weight and total cell number of the lymph nodes; b) increased autotransformation rate and responsiveness of lymph node cells to mitogen; c) decreased frequency of ED1 macrophages and activated OX8 cytotoxic cells in flow cytometry analysis; d) high expression of OX6 class II-positive, OX7 (stem cells), OX62 (migrating dendritic cells), ED1 (macrophages), and OX12 (B cells) on immunohistochemical sections of LNs with some few HIS48 (granulocytes); e) high expression of NOS3 and TGF beta by lymph node lymphocytes and endothelial cells. In summary, local lymph nodes reacted to internal wounds, such as bone fracture and injury to adjacent tissues, through mobilization of cells from the blood circulation, along with activation of cellular subsets. The molecular mechanism that provides the signal for this reaction remains unknown. The absence of major changes in the frequency of lymph node cell subpopulations indicates that lymph nodes are constitutively prepared for influx of antigens from damaged tissues and react only with increase in cell number and cell activation. The nature of the reaction, including lack of immunization against autoantigens, remains unclear. Further elucidation will require studies on the mechanism of cross-tolerance to self-antigens during wound healing. PMID- 15650386 TI - Visualizing function in the tumor-associated lymphatic system. AB - The recent surge of interest in the lymphatic system can be attributed to two factors: the discovery of cytokines that induce the growth of new lymphatic capillaries and the identification of lymphatic endothelial-specific markers. In contrast to the above, there is a paucity of techniques for studying lymphatic function in vivo. This article reviews imaging and other techniques that allow the assessment of lymphatic function, particularly in the tumor microenvironment, and proposes novel solutions for probing the same in vivo. PMID- 15650387 TI - The potential for molecular treatment strategies in lymphatic disease. PMID- 15650388 TI - Literature watch. Peritumor Lymphatics Induced by Vascular Endothelial Growth Factor-C Exhibit Abnormal Function. PMID- 15650389 TI - Research matters from the lymphatic research foundation. PMID- 15650390 TI - Redox signaling in the lungs. PMID- 15650391 TI - H2O2-induced proliferation of primary alveolar epithelial cells is mediated by MAP kinases. AB - Exposure to supraphysiological oxygen concentrations during ventilatory oxygen therapy often causes tissue damage. Alveolar type II (AT II) cells are a major target for oxidant injury, and their ability to proliferate plays a critical role during the repair phase following injury. We hypothesized that reactive oxygen species (ROS), which are produced during hyperoxia, not only cause cellular damage, but may also play a role in the repair process by promoting AT II cell proliferation. We have tested the ability of ROS to induce proliferation in primary cultures of AT II cells by using a wide range of chronic and acute hydrogen peroxide (H2O2) exposures to mimic different types of oxidative stress. We found that chronic exposure to an extracellular flux of 10 microM H2O2/h can significantly increase the intracellular concentration of oxidants, DNA synthesis, and cell proliferation. H2O2-induced AT II cell proliferation was preceded by activation of the mitogen-activated protein kinase ERK (extracellular signal-regulated kinase). Inhibition of ERK and p38 activation prevented H2O2 induced proliferation. These results show that changes in intracellular oxidant concentrations can modulate downstream signaling pathways controlling AT II cell proliferation. This mechanism could be important in the repair process following hyperoxia-induced injury. PMID- 15650392 TI - Redox regulation of p38 MAPK activation and expression of ICAM-1 and heme oxygenase-1 in human alveolar epithelial (A549) cells. AB - We have explored the potential role of redox events in p38 mitogen-activated protein kinase (MAPK) activation and their relevance to the inducible expression of intercellular adhesion molecule-1 (ICAM-1) and heme oxygenase-1 (HO-1) in A549 cells. Tumor necrosis factor-alpha (TNFalpha) and hydrogen peroxide (H2O2) both activated p38, but only TNFalpha activated nuclear factor-kappaB (NF-kappaB). N Acetyl-L-cysteine (20 mM) inhibited both H2O2- and TNFalpha-induced p38 phosphorylation (14 +/- 7 and 37 +/- 4% of control, respectively). The mitochondrial complex I and III inhibitors, rotenone and antimycin A, and allopurinol partially inhibited H2O2- but not TNFalpha-induced p38 activation. However, rotenone and antimycin A augmented intracellular oxidative stress measured by dichlorofluorescein fluorescence. TNFalpha, but not H2O2, induced ICAM-1 in A549 cells, which was attenuated by a proteasome inhibitor, but not by the p38 MAPK inhibitor SB203580. In contrast, hemin and hemoglobin, but neither TNFalpha nor H2O2, caused efficient HO-1 expression. However, hemin had no effect on p38 activation and SB203580 did not influence hemin-induced HO-1 protein expression. Collectively, these data suggest that p38 is a cytokine- and oxidative stress-responsive pathway in A549 cells. Whereas NF-kappaB appears crucial in ICAM-1 induction, p38 activation itself is not sufficient to confer HO 1 expression and may not be involved in HO-1 and ICAM-1 induction in A549 cells. PMID- 15650393 TI - Inhibition of NFkappaB activation and IL-8 expression in human bronchial epithelial cells by acrolein. AB - Lipid oxidation and environmental pollutants are major sources of alpha,beta unsaturated aldehydes such as acrolein and 4-hydroxynonenal. Acrolein (2 propenal), a major product of organic combustion such as tobacco smoke, represents the most reactive alpha,beta-unsaturated aldehyde, with high reactivity toward nucleophilic targets such as sulfhydryl groups. To investigate how acrolein affects respiratory tract cell activation, we exposed either primary (NHBE) or immortalized human bronchial epithelial cells (HBE1) to 0-25 microM acrolein, and determined effects on basal and tumor necrosis factor-alpha (TNFalpha)-induced production of the chemokine interleukin (IL)-8. Cell exposure to acrolein dose-dependently suppressed IL-8 mRNA levels in HBE1 cells (26, 40, and 79% at 5, 10, and 25 microM acrolein concentrations, respectively) and resulted in corresponding decreases in IL-8 production. Studies of nuclear factor kappaB (NFkappaB) activation, an essential event in IL-8 production, showed decreased TNFalpha-induced NFkappaB activation by acrolein, illustrated by inhibition of nuclear translocation of NFkappaB and reduced IkappaBalpha degradation. Immunochemical analysis of IkappaB kinase (IKK), a redox-sensitive regulator of NFkappaB activation, indicated direct modification of the IKK beta subunit by acrolein, suggesting that acrolein may act directly on IKK. In summary, our results demonstrate that acrolein can suppress inflammatory processes in the airways by inhibiting epithelial IL-8 production through direct or indirect inhibitory effects on NFkappaB activation. PMID- 15650394 TI - Curcumin induces glutathione biosynthesis and inhibits NF-kappaB activation and interleukin-8 release in alveolar epithelial cells: mechanism of free radical scavenging activity. AB - Oxidants and tumor necrosis factor-alpha (TNF-alpha) activate transcription factors such as nuclear factor-kappaB (NF-kappaB), which is involved in the transcription of proinflammatory mediators, including interleukin-8 (IL-8). Curcumin (diferuloylmethane) is a naturally occurring flavonoid present in the spice turmeric, which has a long traditional use as a chemotherapeutic agent for many diseases. We hypothesize that curcumin may possess both antioxidant and antiinflammatory properties by increasing the glutathione levels and inhibiting oxidant- and cytokine-induced NF-kappaB activation and IL-8 release from cultured alveolar epithelial cells (A549). Treatment of A549 cells with hydrogen peroxide (H2O2; 100 microM) and TNF-alpha (10 ng/ml) significantly increased NF-kappaB and activator protein-1 (AP-1) activation, as well as IL-8 release. Curcumin inhibited both H2O2- and TNF-alpha-mediated activation of NF-kappaB and AP-1, and IL-8 release. Furthermore, an increased level of GSH and glutamylcysteine ligase catalytic subunit mRNA expression was observed in curcumin-treated cells as compared with untreated cells. Curcumin interacted directly with superoxide anion (O2*-) and hydroxyl radical (*OH) as shown by electron paramagnetic resonance, quenching the interaction of the radicals with the spin trap, Tempone-H. This suggests that curcumin has multiple properties: as an oxygen radical scavenger, antioxidant through modulation of glutathione levels, and antiinflammatory agent through inhibition of IL-8 release in lung cells. PMID- 15650395 TI - Glutathione, stress responses, and redox signaling in lung inflammation. AB - Changes in the ratio of intracellular reduced and disulfide forms of glutathione (GSH/GSSG) can affect signaling pathways that participate in various physiological responses from cell proliferation to gene expression and apoptosis. It is also now known that many proteins have a highly conserved cysteine (sulfhydryl) sequence in their active/regulatory sites, which are primary targets of oxidative modifications and thus important components of redox signaling. However, the mechanism by which oxidants and GSH/protein-cysteine-thiols actually participate in redox signaling still remains to be elucidated. Initial studies involving the role of cysteine in various proteins have revealed that cysteine-SH may mediate redox signaling via reversible or irreversible oxidative modification to Cys-sulfenate or Cys-sulfinate and Cys-sulfonate species, respectively. Oxidative stress possibly via the modification of cysteine residues activates multiple stress kinase pathways and transcription factors nuclear factor-kappaB and activator protein-1, which differentially regulate the genes for proinflammatory cytokines as well as the protective antioxidant genes. Understanding the redox signaling mechanisms for differential gene regulation may allow for the development of novel pharmacological approaches that preferentially up-regulate key antioxidants genes, which, in turn, reduce or resolve inflammation and injury. This forum article features the current knowledge on the role of GSH in redox signaling, particularly the regulation of transcription factors and downstream signaling in lung inflammation. PMID- 15650396 TI - Redox regulation of lung inflammation by thioredoxin. AB - The lungs are the richest in oxygen among the various organs of the body and are always subject to harmful reactive oxygen species. Regulation of the reduction/oxidation (redox) state is critical for cell viability, activation, proliferation, and organ functions. Although the protective importance of various antioxidants has been reported, few antioxidants have established their clinical usefulness. Thioredoxin (TRX), a key redox molecule, plays crucial roles as an antioxidant and a catalyst in protein disulfide/dithiol exchange. TRX also modulates intracellular signal transduction and exerts antiinflammatory effects in tissues. In addition to its beneficial effects in other organs, the protective effect of TRX in the lungs has been shown against ischemia/ reperfusion injury, influenza infection, bleomycin-induced injury, or lethal inflammation caused by interleukin- 2 and interleukin-18. Monitoring of TRX in the plasma, airway, or lung tissue may be useful for the diagnosis and follow-up of pulmonary inflammation. Promotion/modulation of the TRX system by the administration of recombinant TRX protein, induction of endogenous TRX, or gene therapies can be a therapeutic modality for oxidative stress-associated lung disorders. PMID- 15650397 TI - The regulation and role of extracellular glutathione peroxidase. AB - Reactive oxygen species and reactive nitrogen species are mediators of lung tissue damage. To minimize the effect of oxidative stress, the lung is well equipped with an integrated antioxidant system. In some circumstances, antioxidants increase in response to oxidants and reduce tissue injury. The lung is somewhat unique in that it has an extracellular surface, which is often directly exposed to oxidative stresses. In this context, the extracellular antioxidant system, comprised primarily of glutathione and glutathione peroxidase, is especially important in protecting against oxidant injury. Induction of extracellular glutathione peroxidase occurs in airway inflammation and undoubtedly plays an important defense against oxidative injury to the airway surface. PMID- 15650398 TI - Heme oxygenase-1: redox regulation of a stress protein in lung and cell culture models. AB - Reactive oxygen species (ROS) may contribute to tissue damage in many pathophysiological conditions and participate in physiological signaling processes. The mechanisms by which cells sense prooxidant states, and activate signaling pathways leading to adaptive responses, remain incompletely understood. Bacteria contain several transcriptional regulators (e.g., OxyR) and a low molecular-weight heat shock protein (HSP33), whose activity increases upon oxidation of critical sulfhydryl residues. These proteins participate in cellular adaptation to oxidative stress. In higher organisms, heme oxygenase-1 (HO-1) has been widely studied as a model for redox-regulated gene expression. Expression of HO-1 responds to chemical and physical agents that directly or indirectly generate ROS. Depletion of cellular reduced glutathione may act as a signal for HO-1 transcriptional activation. Furthermore, antioxidants and metal-chelating compounds can modulate HO-1 expression. Several signaling molecules (e.g., mitogen-activated protein kinases), transcriptional regulators (activator protein 1, NF-E2-related factor-2, hypoxia-inducible factor-1, Bach-1), as well as two enhancer regions in the ho-1 5' regulatory region, participate in the regulation of the ho-1 gene. HO-1 protein expression can occur in the lung in response to oxidative stress associated with infection, altered oxygen tension, and inflammatory diseases. HO-1 remains widely regarded as a protective mechanism against oxidative tissue injury. PMID- 15650399 TI - Redox regulation of lung development and perinatal lung epithelial function. AB - Throughout gestation, low oxygen tensions are a dominant feature of the fetal environment and so may be important in sustaining a normal pattern of lung morphogenesis until the moment of birth. As breathing begins, the equilibration of the lung lumen to postnatal PO2 evokes a series of physiologic and morphogenic maturation events that are partially reversible by hypoxia. In this review, we discuss the experimental evidence that fetal and perinatal oxygen tensions differently influence lung morphogenesis through oxygen- and redox-responsive signaling pathways and identify five loci at which this regulation may occur: (I) proliferation of undifferentiated lung mesenchyme as governed by hypoxia regulated transcription factors (HIF-1alpha, C/EBPbeta); (II) transient production of reactive oxygen species (ROS) and nuclear oxidation of the perinatal lung epithelium; (III) nuclear transport and oxidation of thioredoxin in hand with the acute activation of nuclear factor- kappaB (NF-kappaB); (IV) ROS evoked chronic rise in intracellular glutathione and thioredoxin redox buffering capacity; and (V) NF-kappaB-dependent increase in transepithelial Na+ transport and lung lumenal fluid clearance. Although not exhaustive, this analysis leads us to the conclusion that redox events that occur in the lung during gestation, parturition, and the early neonatal period may dramatically influence the expression of genes and physiological events that are crucial to the successful transition from fetal to postnatal lung maturation. PMID- 15650400 TI - Redox activation of p21Cip1/WAF1/Sdi1: a multifunctional regulator of cell survival and death. AB - Cell division requires the coordinated assembly of cyclins and cyclin-dependent kinases that promote cell-cycle progression through S phase and mitosis. Two families of cyclin-dependent kinase inhibitors prevent abnormal or premature proliferation by blocking cyclin kinase activity. Expression of the cyclin dependent kinase inhibitor p21, a member of the Cip/Kip family, increases when cells are damaged. In addition to controlling cell-cycle progression, p21 participates in DNA repair and apoptotic processes. The recent appreciation that p21 regulates cell survival and death implies that it is a master regulator of cell fate. This review discusses how p21 can affect the cellular response to oxidative stress. PMID- 15650401 TI - Life and death decisions: ceramide generation and EGF receptor trafficking are modulated by oxidative stress. AB - Reactive oxidants are associated with the pathogenesis of pulmonary diseases and affect various cell functions, from proliferation to apoptosis. We have shown that oxidants exert growth control on airway epithelial cells by modulating upstream receptor function. Additionally, hydrogen peroxide-mediated oxidative stress modulates ceramide levels to induce apoptosis in lung epithelium. Depletion of glutathione in lung epithelial cells results in ceramide accumulation, suggesting that ceramide elevation, coupled to oxidative stress, initiates apoptosis. While it is desirable to prevent cell death and tissue injury induced by oxidants in diseases such as asthma or acute respiratory distress syndrome, the opposite is sought in cancer. But oxidants may also activate growth factor receptors, enhancing cell proliferation and facilitating tumor promotion. Under oxidative stress, phosphorylation of the epidermal growth factor receptor (EGFR) is abrogated at tyrosine 1,045, the docking site for the ubiquitin ligase c-Cbl, rendering EGFR unable to recruit c-Cbl and be ubiquitylated and degraded. We thus proposed that this deficiency, which confers prolonged receptor signaling at the plasma membrane, links oxidative stress, EGFR, and tumorigenesis. Decoding the molecular interactions between oxidative stress and ceramide pathways and characterizing ubiquitylation control of receptor desensitization should provide new strategies for intervention in diverse pulmonary diseases and in diagnosing and eradicating cancer. PMID- 15650403 TI - Redox regulation of histone deacetylases and glucocorticoid-mediated inhibition of the inflammatory response. AB - Gene expression, at least in part, is regulated by changes in histone acetylation status induced by activation of the proinflammatory redox-sensitive transcription factors activator protein-1 (AP-1) and nuclear factor-kappaB (NF-kappaB). Hyperacetylated histone is associated with open actively transcribed DNA and enhanced inflammatory gene expression. In contrast, hypoacetylated histone is linked to a closed repressed DNA state and a lack of gene expression. The degree of inflammatory gene expression is a result of a balance between histone acetylation and histone deacetylation. One of the major mechanisms of glucocorticoid function is to recruit histone deacetylase enzymes to the site of active gene expression, thus reducing inflammation. Oxidative stress can enhance inflammatory gene expression by further stimulating AP-1- and NF-kappaB-mediated gene expression and elevating histone acetylation. In addition, oxidants can reduce glucocorticoid function by attenuating histone deacetylase activity and expression. Thus, oxidant stress, acting through changes in chromatin structure, can enhance inflammation and induce a state of relative glucocorticoid insensitivity. This may account for the lack of glucocorticoid sensitivity in patients with chronic obstructive pulmonary disease. Antioxidants should reduce the inflammation and restore glucocorticoid sensitivity in these subjects. PMID- 15650402 TI - Nitric oxide and redox signaling in allergic airway inflammation. AB - A number of diseases of the respiratory tract, as exemplified in this review by asthma, are associated with increased amounts of nitric oxide (NO) in the expired breath. Asthma is furthermore characterized by increased production of reactive oxygen species that scavenge NO to form more reactive nitrogen species as demonstrated by the enhanced presence of nitrated proteins in the lungs of these patients. This increased oxidative metabolism leaves less bioavailable NO and coincides with lower amounts of S-nitrosothiols. In this review, we speculate on mechanisms responsible for the increased amounts of NO in inflammatory airway disease and discuss the apparent paradox of higher levels of NO as opposed to decreased amounts of S-nitrosothiols. We will furthermore give an overview of the regulation of NO production and biochemical events by which NO transduces signals into cellular responses, with a particular focus on modulation of inflammation by NO. Lastly, difficulties in studying NO signaling and possible therapeutic uses for NO will be highlighted. PMID- 15650404 TI - Introduction for special forum issue on isoprostanes and related compounds. PMID- 15650405 TI - Free radical oxidation of plasmalogen glycerophosphocholine containing esterified docosahexaenoic acid: structure determination by mass spectrometry. AB - Plasmalogen phospholipids have a vinyl ether substituent at the sn-1 position that is susceptible to oxidative reactions that occur at cell membranes. However, the mechanism by which this oxidation occurs and the effect of the polyunsaturated fatty acid at the sn-2 position have not been established. To gain insight into these mechanisms, the oxidized phospholipid products resulting from the exposure of 1-O-hexadec-1'-enyl-2-docosahexaenoyl-sn-glycero-3 phosphocholine (16:0p/22:6-GPCho) to the free radical initiator 2, 2'-azobis (2- amidinopropane) hydrochloride were examined. Electrospray ionization tandem mass spectrometry, UV spectroscopy, and electron ionization-gas chromatography/mass spectrometry were used to structurally characterize the oxidized glycerophosphocholine (GPCho) products. The radical-induced peroxidation of 16:0p/22:6-GPCho revealed two major classes of oxidized phospholipids. The first class of products was formed by oxidation at the sn-1 position and included 1 lyso-2-docosahexaenoyl-GPCho and 1-formyl-2-docosahexaenoyl-GPCho. Additionally, the second class of oxidized products where oxidation occurred at the sn-2 position was classified into three categories that included chain-shortened omega aldehydes, terminal gamma-hydroxy-alpha,beta-unsaturated aldehydes, and the addition of one or two oxygen atoms onto the sn-2 position of 16:0p/22:6-GPCho. These results clearly indicate that free radical-induced oxidation of plasmalogen phospholipids with esterified docosahexaenoic acid at the sn-2 position underwent oxidation at both the sn-1 and sn-2 positions. PMID- 15650406 TI - New insights regarding the autoxidation of polyunsaturated fatty acids. AB - Free radical-initiated autoxidation of polyunsaturated fatty acids (PUFAs) has been implicated in numerous human diseases, including atherosclerosis and cancer. This review covers the free radical mechanisms of lipid oxidation and recent developments of analytical techniques to analyze the lipid oxidation products. Autoxidation of PUFAs generates hydroperoxides as primary oxidation products, and further oxidation leads to cyclic peroxides as secondary oxidation products. Characterization of these oxidation products is accomplished by several mass spectrometric techniques. Ag+ coordination ion spray mass spectrometry has proven to be a powerful tool to analyze the intact lipid peroxides. Monocyclic peroxides, bicyclic endoperoxides, serial cyclic peroxides, and a novel class of endoperoxides (dioxolane-isoprostane peroxides) have been identified from the oxidation of arachidonate. Electron capture atmospheric pressure chemical ionization mass spectrometry has been applied to study lipid oxidation products after derivatization. All eight possible diastereomeric isoprostanes are observed from the oxidation of a single hydroperoxide precursor. 5- and 15-series isoprostanes are more abundant than the 8- and 12-series because the precursors that lead to 8- and 12-series compounds can undergo further oxidation and form dioxolane-isoprostane peroxides. Furthermore, formation of isoprostanes from 15 hydroperoxyeicosatetraenoate occurs from beta-fragmentation of the corresponding peroxyl radical to generate a pentadienyl radical rather than a "dioxetane" intermediate, as previously suggested. PMID- 15650407 TI - Levuglandins and isolevuglandins: stealthy toxins of oxidative injury. AB - Inspired by a reaction discovered through basic research on the chemistry of the bicyclic peroxide nucleus of the prostaglandin endoperoxide PGH2, we postulated that levulinaldehyde derivatives with prostaglandin side chains, levuglandins (LGs), and structurally isomeric analogues, isolevuglandins (iso[n]LGs), would be generated by nonenzymatic rearrangements of prostanoid and isoprostanoid endoperoxides. Two decades of subsequent studies culminated in our discoveries of the LG and isoLG pathways, branches of the cyclooxygenase and isoprostane pathways, respectively. In cells, PGH2 rearranges nonenzymatically to LGs even in the presence of enzymes that use PGH2 as a substrate. IsoLGs, also known as isoketals or neuroketals, are generated in vivo through free radical-induced autoxidation of polyunsaturated phospholipid esters. Hydrolysis occurs after rapid adduction of isoLG phospholipids to proteins. The proclivity of these reactive species to avidly bind covalently with and cross-link proteins and nucleic acids complicated the hunt for LGs and isoLGs in vivo. The extraordinary reactivity of these "stealthy toxins" underlies much, if not all, of the biological consequences of LG and isoLG generation. They interfere with protein function and are among the most potent neurotoxic products of lipid oxidation known. Because they can accumulate over the lifetimes of proteins, iso[n]LG protein adducts represent a convenient dosimeter of oxidative stress. PMID- 15650408 TI - Isofurans: novel products of lipid peroxidation that define the occurrence of oxidant injury in settings of elevated oxygen tension. AB - We recently reported the discovery of isofurans, novel products of free radical induced peroxidation of arachidonic acid that exhibit favored formation with increasing oxygen concentrations. In this review, the biochemistry of isofuran formation is compared with that of isoprostanes, with an emphasis on the mechanistic basis for the favored formation of isofurans at elevated oxygen tensions. In addition, the formation of isofurans in various disease states in vivo is also discussed. Parkinson's disease is presented as a disease model involving mitochondrial dysfunction, a situation in which quantification of isofurans can provide a uniquely sensitive indicator of oxidant injury. Measurement of isofurans has also provided unexpected insights into the earliest events in hyperoxic lung injury, an important clinical problem in which measurement of isofurans might prove to be uniquely valuable in the evaluation of approaches to limit this injury. These two settings are then used as models to suggest a variety of other pathological settings in which measurement of isofurans together with isoprostanes could provide a complete and robust picture of oxidative stress status in ongoing and future investigations. PMID- 15650409 TI - The cyclopentenone (A2/J2) isoprostanes--unique, highly reactive products of arachidonate peroxidation. AB - Cyclopentenone (A2/J2) isoprostanes (IsoPs) are a group of prostaglandin (PG) like compounds generated in vivo from the free radical-induced peroxidation of arachidonic acid. Unlike other classes of IsoPs, cyclopentenone IsoPs contain highly reactive unsaturated carbonyl moieties on the prostane ring analogous to cyclooxygenase-derived PGA2 and PGJ2 that readily adduct relevant biomolecules such as thiols via Michael addition. The purpose of this review is to summarize our knowledge of the A2/J2-IsoPs. As a starting point, we will briefly discuss the formation and biological properties of PGA2 and PGJ2. Next, we will review studies definitively showing that cyclopentenone IsoPs are formed in large amounts in vivo. This is in marked contrast to cyclopentenone PGs, for which little evidence exists that they are endogenously produced. Subsequently, we will discuss studies related to the chemical syntheses of the 15-A2-IsoP series of cyclopentenone IsoPs. The successful synthesis of these compounds provides the recent impetus to explore the metabolism and biological properties of A-ring IsoPs, particularly as modulators of inflammation, and this work will be discussed. Finally, the formation of cyclopentenone IsoP-like compounds from other fatty acids such as linolenic acid and docosahexaenoic acid will be detailed. PMID- 15650410 TI - Factors regulating isoprostane formation in vivo. AB - Discovery of the F2-isoprostanes, a group of prostaglandin F2-like compounds biosynthesized from arachidonic acid nonenzymatically, has uncovered a new and novel facet of free radical biology. Some of these compounds are bioactive and thus may mediate adverse effects associated with oxidant stress. F2-Isoprostanes have also been shown to be reliable biomarkers of lipid peroxidation. Factors influencing their formation and metabolism have been studied to some extent, although much remains to be determined. The purpose of this review is to summarize our current knowledge of conditions that modulate endogenous generation of these compounds. Isoprostanes have a wide daily variation in secretion in humans. Although normal levels can be defined, these compounds are found in increased concentrations in various pathophysiological states, including ischemia reperfusion injury, atherosclerosis, and diabetes, and in experimental conditions of oxidative stress and inflammation. Alterations in isoprostane biosynthesis, secretion, and excretion in normal physiology and in pathophysiological states are due to the various types of endogenous and exogenous regulatory mechanisms that control the availability of precursors required for isoprostane synthesis, such as dietary and tissue arachidonic acid content, oxygen concentration, and the generation of various free radical species. Selected aspects of issues related to isoprostane formation and metabolism in vivo will be examined herein. PMID- 15650411 TI - Isoprostanes and the kidney. AB - Isoprostanes are not mere bystanders of oxidative injury, but possess potent biological activity and may thus contribute to the pathophysiology of various disorders associated with an increase in free radical formation. 15-F2t-IsoP (8 iso-prostaglandin F2) and 15-E2t-IsoP (8-iso-prostaglandin E2), two of the most abundant isoprostanes, are potent vasoconstrictors in various vascular beds, including the kidney. Since their discovery, numerous studies have aimed to define the receptors through which isoprostanes exert their effects. Whether the thromboxane receptor and/or other prostaglandin receptors mediate the actions of isoprostanes, or whether these compounds interact with their own unique receptors, remains to be clarified. Regardless of their exact mode of action, isoprostanes are being implicated in the pathophysiology of a variety of diseases, and their discovery might give rise to novel therapies for these diseases. Here we describe early studies that defined the vasoactive properties of isoprostanes in the kidney, and subsequent discoveries relating to their renal actions and pathophysiologic significance. PMID- 15650412 TI - The pulmonary biology of isoprostanes. AB - Isoprostanes were first recognized as convenient markers of oxidative stress, but their powerful effects on a variety of cell functions are now also being increasingly appreciated. This is particularly true of the lung, which is comprised of a wide variety of different cell types (smooth muscle, innervation, epithelium, lymphatics, etc.), all of which have been shown to respond to exogenously applied isoprostanes. In this review, we summarize these biological responses in the lung, and also consider the roles that isoprostanes might play in a range of pulmonary clinical disorders. PMID- 15650413 TI - Lipid peroxidation in diabetes mellitus. AB - There is considerable evidence that hyperglycemia represents the main cause of complications of diabetes mellitus (DM), and oxidative stress resulting from increased generation of reactive oxygen species plays a crucial role in their pathogenesis. In fact, in the absence of an appropriate response from endogenous antioxidant mechanisms, the redox imbalance causes the activation of stress sensitive intracellular signaling pathways. The latter play a key role in the development of late complications of DM, as well as in mediating insulin resistance (i.e., resistance to insulin-mediated glucose uptake by some cells) and impaired insulin secretion. This review, focused on lipid peroxidation in DM, will examine the mechanisms and clinical readouts of oxidative stress in this setting, the relationship between lipid peroxidation and antioxidant status in type 1 and type 2 DM, the effects of hyperglycemia and metabolic control on in vivo markers of lipid peroxidation (i.e., isoprostanes), and the association between isoprostane formation and platelet activation. Finally, possible targets of antioxidant therapy for diabetic vascular complications will be discussed. PMID- 15650414 TI - F2-isoprostanes in Alzheimer and other neurodegenerative diseases. AB - Increased free radical-mediated injury to brain is proposed to be an integral component of several neurodegenerative diseases, including Alzheimer's disease (AD). Lipid peroxidation is a major outcome of free radical- mediated injury to brain, where it directly damages membranes and generates a number of oxidized products. F2-Isoprostanes (F2-IsoPs), one group of lipid peroxidation products derived from arachidonic acid, are especially useful as in vivo biomarkers of lipid peroxidation. F2-IsoP concentration is selectively increased in diseased regions of brain from patients who died from advanced AD, where pathologic changes include amyloid beta (Abeta) amyloidogenesis, neurofibrillary tangle formation, and extensive neuron death. Interestingly, cerebral F2-IsoPs are not reproducibly elevated in aged mouse models of cerebral Abeta amyloidogenesis only. There is broad agreement that increased cerebrospinal fluid (CSF) levels of F2-IsoPs also are present in patients with early AD. Demonstrated applications of quantifying CSF F2-IsoPs have improved laboratory diagnostic accuracy of AD and objective assessment of antioxidant therapeutics. In contrast, quantification of F2-IsoPs in plasma and urine of AD patients has produced conflicting data. These results indicate that brain lipid peroxidation is a potential therapeutic target early in the course of AD, and that CSF F2-IsoPs may aid in the assessment of antioxidant experimental therapeutics and laboratory diagnosis of AD. PMID- 15650415 TI - Selenium attenuates expression of MnSOD and uncoupling protein 2 in J774.2 macrophages: molecular mechanism for its cell-death and antiinflammatory activity. AB - Selenium can activate cell death. However, the mechanism of action is not yet fully defined. We hypothesized that selenium may impede mitochondrial superoxide dismutation to H2O2 and O2, leading to cell death in macrophages and that this effect may be relevant to antiinflammatory treatment by selenium. In this study, the mechanism of action of selenium was investigated in nonactivated and activated (immune-stimulated) J774.2 macrophages. Sodium selenite treatment decreased dichlorodihydrofluorescein-reacting intracellular reactive oxygen species (ROS) (mainly peroxides and hydroxyl radicals), with no correlation to glutathione peroxidase activity. However, selenite decreased the transcription and expression of manganese superoxide dismutase (MnSOD) and uncoupling protein 2 (UCP2). This cellular effect was due to inhibition of specificity protein-1 (Sp1) binding to its DNA binding site. Following immune stimulation of macrophages using lipopolysaccharides plus interferon-gamma, MnSOD was up-regulated. Activated macrophages showed higher mitochondrial membrane potential, intracellular ROS levels, and cellular resistance to cell death. Selenite treatment attenuated all of these parameters. Selenite prevented nuclear factor kappaB (NF-kappaB) activation as a mechanism of its inhibitory activity on MnSOD expression in the immune-stimulated cells. In addition, overexpression of human MnSOD protected against death induced by selenite treatment. It is therefore concluded that selenium at high nanomolar to low micromolar concentrations shifts the balance between inflammatory response and cell death toward the latter, through a direct effect on the transcription factors Sp1 and NF-kappaB, and down regulation of MnSOD and UCP2. PMID- 15650416 TI - Vitamin C-induced loss of redox-dependent viability in lung microvascular endothelial cells. AB - Recent clinical trials have shown that vitamin C, at pharmacological concentrations (milligram to approximately gram), upon infusion into circulation, modulates vasodilation and vascular tone in humans. This also results in the elevated concentrations of vitamin C in circulation in the millimolar range. Here, it was hypothesized that vitamin C at pharmacological concentrations (millimolar) would induce oxidative stress and cause loss of redox-dependent cell viability in vascular endothelial cells (ECs). To test the hypothesis, bovine lung microvascular ECs (BLMVECs) in monolayer cultures were exposed to vitamin C (0-10 mM) for different time periods (0-2 h). Electron paramagnetic resonance spectroscopy revealed the intracellular formation of ascorbate free radical in a dose- and time-dependent fashion. Vitamin C also induced formation of intracellular reactive oxygen species in a dose-dependent fashion. It was observed that vitamin C induced morphological alterations and loss of cell viability in a dose- and time-dependent fashion, as measured by light microscopy and Alamar Blue redox cell viability assay, respectively. Vitamin C analogues failed to induce such changes. Vitamin C depleted cellular GSH levels in a dose dependent fashion, suggesting that vitamin C altered thiol-redox status in BLMVECs. Antioxidants, intracellular iron chelator, and catalase protected cells against vitamin C-induced loss of redox-dependent cell viability, confirming the role of hydrogen peroxide and iron during redox cycling of vitamin C. These results, for the first time in detail, established that vitamin C at pharmacological doses induced oxidative stress and loss of redox-dependent cell viability in microvascular ECs. PMID- 15650419 TI - Influence of prior structured treatment interruptions on the length of time without antiretroviral treatment in chronically HIV-infected subjects. AB - The influence of previous structured treatment interruptions (STIs) on the length of time off therapy when highly active antiretroviral treatment (HAART) is discontinued in chronically HIV-infected subjects was assessed. A comparative, retrospective clinical cohort study included patients with plasma viral load (VL) <50 copies/ml and CD4 cell count >500 cells/mm(3) who interrupted HAART. Fifteen patients interrupted HAART after six 2-weeks-off-/4-weeks-on therapy cycles (STI group) and 30 subjects discontinued HAART without previous STIs (NSTI group). The criteria for treatment resumption were development of AIDS-defining clinical events, VL >100,000 copies/ml or CD4 <350 cells/mm(3). Median (IQR) time off therapy was 48 (29-56) weeks in the STI group and 31 (8-77) weeks in the NSTI group (p < 0.15). After 48 weeks, 46% of the patients in the STI group and 40% in the NSTI group remained off HAART (p < 0.74). No patient developed AIDS-defining events and all but one achieved virological control after treatment resumption. The CD4 nadir was 341 (298-464) cells/mm(3) among patients who reinitiated HAART and 560 (364-682) cells/mm(3) in those who remained off therapy by week 48 (p <0.01). Likewise, CD4 count prior to treatment interruption was 902 (806-1040) cells/mm(3) and 1123 (924-1234) cells/mm(3) in subjects resuming and remaining off HAART, respectively (p = 0.03). No relationship between treatment resumption and pre-ART VL or with the time with undetectable VL before enrollment was found. CD4 nadir was a significant predictor for treatment reinitiation in a multivariate analysis. Previous STIs do not influence time off therapy when HAART is definitively discontinued in chronically HIV-infected subjects. CD4 nadir is an important factor in the treatment discontinuation decision. PMID- 15650420 TI - Short communication: benefits in the lipid profile after substitution of abacavir for Stavudine: a 48-week prospective study. AB - Stavudine (d4T) has been associated with lipoatrophy and hyperlactatemia. In recent studies, d4T has also been related to both hypercholesterolemia and hypertriglyceridemia. Replacing d4T with another nucleoside analogue such as abacavir (ABC) may reduce lactate levels and improve lipoatrophy in the long term. However, the impact of this strategy on the lipid profile is still unclear. In a prospective and randomized study, fasting lipids were examined over 48 weeks in 112 subjects on d4T regimens, 49 of whom replaced d4T with ABC. The substitution of ABC for d4T was found to be safe and provided a reduction in both LDL cholesterol and the total cholesterol (TC)/HDLc ratio, which might impact favorably on cardiovascular risk. PMID- 15650421 TI - Advanced liver fibrosis in HIV/HCV-coinfected patients on antiretroviral therapy. AB - HIV infection is believed to adversely affect the progression of hepatitis C virus (HCV)-related liver disease. However, information regarding HIV and HCV coinfection in the era of highly active antiretroviral therapy (HAART) is scarce. A cross-sectional study in 75 HCV/HIV-coinfected patients (most of them on HAART) and 75 HCV-monoinfected patients paired by age, sex, and date of liver biopsy analyzed the association of HIV infection with advanced liver fibrosis (Knodell fibrosis stages 3 + 4). The median CD4 cell count in HIV-coinfected patients was 546 cells/microl; 78.7% had an HIV-1 viral load <1000 copies/ml and 88% were on antiretroviral therapy. The percentage of patients harboring genotype 4 and with a higher HCV viral load was greater in the HIV-coinfected group. HCV/HIV coinfected patients had more advanced liver fibrosis (Knodell fibrosis stages 3 + 4) than HCV-monoinfected patients (46.7% vs. 12%, p < 0.0001). In the univariate analysis, the factors associated with advanced liver disease were male sex (OR: 2.7, 95% CI: 1.05-7.1), history of injecting drug use (OR: 4.6, 95% CI: 2.0 10.2), HIV infection (OR: 6.4, 95% CI: 2.7-14.7), and previous exposure to therapy with protease inhibitors (OR: 3.0, 95% CI:1.4-6.3). In the multivariate analysis; only male sex (OR: 3.17, 95% CI: 1.152-8.773) and HIV infection (OR: 6.85, 95% CI: 2.93-16.005) were associated with advanced liver fibrosis. HIV infection is associated with advanced liver fibrosis. HIV/HCV-coinfected individuals on HAART are at risk of developing end-stage liver disease despite virological success and immunological reconstitution. PMID- 15650422 TI - Use of a replication-defective vector to track cells initially infected by SIV in vivo: infected mononuclear cells rapidly appear in the draining lymph node after intradermal inoculation of rhesus monkeys. AB - A better understanding of the mechanisms of HIV dissemination, a key step in pathogenesis, would be possible if the cellular pathways of viral dissemination could be followed in simian immunodeficiency virus (SIV)- inoculated monkeys or HIV-infected people. In an initial attempt to follow this process using a traceable virus infection, we inoculated rhesus monkeys intradermally (ID) or directly into lymph nodes with a replication-defective SIV-based vector expressing the enhanced jellyfish green fluorescent protein (EGFP), V1EGFP. EGFP expression was detected in mononuclear cells isolated from the sites of inoculation (skin and lymph node) at 5 and 16 hr after inoculation and then cultured in vitro for 6 days to allow maximum EGFP expression. Similarly, EGFP expressing, SIV-infected cells could be detected at 16 hr postinfection in the lymph nodes that drained the sites of ID inoculation. Since V1EGFP is a replication-defective vector, the EGFP-expressing cells are the initial target cells infected by the virions in the original inoculum. The results of flow cytometric analysis were confirmed by a nested PCR assay to detect SIV DNA and hence infection of cells and reverse transcription. These experiments indicate that 16 hr after ID inoculation newly infected cells either remain in the skin at the site of inoculation or have migrated to the draining lymph node. The results in this SIV vector model probably reflect the short time (less than 16 hr) required for HIV to move from a site of epithelial penetration to the lymphoid tissues via lymphatic vessels. PMID- 15650423 TI - Genotypic and phenotypic correlates of the HIV Type 1 env gene evolution in infected children with discordant response to antiretroviral therapy. AB - The genotypic-phenotypic correlates of human immunodeficiency virus type 1 (HIV 1) env gene evolution were investigated in samples from eight infected children under antiretroviral therapy (ART) and virological failure. Evolution of the gp120 C2-V5 env sequence was demonstrated in all subjects but one by sequence analysis of the replicating plasma virus collected at baseline and 12-15 months after ART initiation. The analysis of the host's selective pressure showed that in four subjects, the ratio of nonsynonymous to synonymous amino acid substitutions was higher in the V3 sequences than in the C2-V5 region ([K (a)/K (s)](V3)/[K (a)/K (s)](C2V5) >1.0). Interestingly, this feature was observed only in subjects (four of five) showing an increase in T cell receptor rearrangement excision circle (TREC)-bearing cells and in CD4(+) T-lymphocytes despite persistent viremia (discordant therapeutic response) (p = 0.02), thus suggesting that the V3 loop is a target of the immune reconstitution under ART. Using phenotypic analysis of recombinant viruses expressing exogenous V3 sequences, a reverse shift from CXCR4-tropic to CCR5-tropic variants was demonstrated in two of the four subjects, further indicating that the host's selective pressure sharply forces the V3 evolution of replicating variants. The data indicate that a complex HIV-1-host interplay occurs in children receiving antiretroviral treatments and suggest that the recovery of thymic function places a selective constraint on the viral V3 loop. PMID- 15650424 TI - Mutation, fitness, viral diversity, and predictive markers of disease progression in a computational model of HIV type 1 infection. AB - The aim of this study was to develop a computational model of HIV infection able to simulate the natural history of the disease and to test predictive parameters of disease progression. We describe the results of a numerical simulation of the cellular and humoral immune response to HIV-1 infection as an adaptive pathway in a "bit-string" space. A total of 650 simulations of the HIV-1 dynamics were performed with a modified version of the Celada-Seiden immune system model. Statistics are in agreement with epidemiological studies showing a log normal distribution for the time span between infection and the development of AIDS. As predictive parameters of disease progression we found that HIV-1 accumulates "bit" mutations mainly in the peptide sequences recognized by cytotoxic CD8 T cells, indicating that cell-mediated immunity plays a major role in viral control. The viral load set point was closely correlated with the time from infection to development of AIDS. Viral divergence from the viral quasispecies that was present at the beginning of infection in long-term nonprogressors (LTNP) was found to be similar to that found in rapid progressors at the time CD4 T cells drop below the critical value of 200 cells/microl. In contrast, the diversity indicated by the number of HIV strains present at the same time was higher for rapid and normal progressors compared to LTNP, suggesting that the early immune response can make the difference. This computational model may help to define the predictive parameters of HIV dynamics and disease progression, with potential applications in therapeutic and vaccine simulations. PMID- 15650425 TI - Inhibition of HIV type 1 replication using lentiviral-mediated delivery of mutant tRNA(Lys3)A58U. AB - In previous studies, we showed that residue A58 of cellular tRNALys3 is necessary for appropriate termination of viral plus-strand strong-stop DNA (+SS DNA), and therefore plays a critical role in the life cycle of HIV-1. We also performed proof-of-principle studies that established that a mutant form of this tRNA primer (tRNA(Lys3)A58U, which lacks the M1A58 residue necessary for +SS DNA termination) could inhibit HIV-1 replication. In the present work, we examined whether a third generation lentiviral vector (SIN) could be used to deliver tRNA(Lys3)A58U to CEM cells. Using both viral kinetic studies and limiting dilution assays (LDA), we observed significant impairment of HIV-1 replication, up to 3 logs in the LDA, in CEM sublines expressing mutant tRNA(Lys3)A58U. No inhibition occurred in cells that either expressed wild-type tRNA(Lys3) or were transduced with empty SIN vector. Further, we observed impairment of viral replication using primary isolates of both HIV-1 and HIV-2 in sublines containing tRNA(Lys3)A58U. We also detected "breakthrough" HIV-1 replication in some tRNA(Lys3)A58U-expressing cultures. Interestingly, analyzed breakthrough viruses appeared to be both genetically and phenotypically wild type. One possible explanation for virological breakthrough is that it reflects the gradual accumulation of HIV-1 within the infected cell culture, to a level that ultimately exceeds the containment "threshold" conferred by tRNA(Lys3)A58U. The fact that HIV-1 does not appear to acquire heritable resistance to tRNA(Lys3)A58U mediated blockade differentiates this antiviral modality from other therapeutic interventions. It also suggests that tRNA-mediated inhibition of viral replication might be a valuable adjunct to other antiviral approaches. PMID- 15650426 TI - DNA/MVA vaccine for HIV type 1: effects of codon-optimization and the expression of aggregates or virus-like particles on the immunogenicity of the DNA prime. AB - Recently, a vaccine consisting of DNA priming followed by boosting with modified vaccinia Ankara (MVA) has provided long-term protection of rhesus macaques against a virulent challenge with a chimera of simian and human immunodeficiency viruses. Here, we report studies on the development of the DNA component for a DNA/MVA HIV vaccine for humans. Specifically, we assess the ability of a codon optimized Gag-expressing DNA and two noncodon-optimized Gag-Pol-Env-expressing DNAs to prime the MVA booster dose. The codon-optimized DNA expressed virus-like particles (VLPs), whereas one of the noncodon-optimized DNAs expressed VLPs and the other expressed aggregates of HIV proteins. The MVA boost expressed Gag-Pol and Env and produced VLPs. Immunogenicity studies in macaques used one intramuscular prime with 600 microg of DNA and two intramuscular boosts with 1 x 10(8) pfu of MVA at weeks 8 and 30. The codon-optimized and noncodon-optimized DNAs proved similar in their ability to prime anti-Gag T cell responses. The aggregate and VLP-expressing Gag-Pol-Env DNAs also showed no significant differences in their ability to prime anti-Env Ab responses. The second MVA booster dose did not increase the peak CD4 and CD8 T cell responses, but increased anti-Env Ab titers by 40- to 90-fold. MVA-only immunizations elicited 10-100 times lower frequencies of T cells and 2-4 lower titers of anti-Env Ab than the Gag-Pol-Env DNA/MVA immunizations. Based on the breadth of the T cell response and a trend toward higher titers of anti-Env Ab, we are moving forward with human trials of the noncodon-optimized VLP-expressing DNA. PMID- 15650427 TI - A link between SIVsm in sooty mangabeys (SM) in wild-living monkeys in Sierra Leone and SIVsm in an American-based SM colony. AB - We have developed a noninvasive method for SIVsm virion RNA detection in feces of captive sooty mangabeys (SMs) (Cercocebus atys). Employing this method to investigate the natural history of SIVsm in endangered SMs is useful for understanding the diversity and evolution of SIVsm and HIV-2. The fecal samples of 61 wild-living SMs and 14 chimpanzees (Pan troglodytes verus) were studied. Samples were collected in rural Sierra Leone in 1993. One SM sample tested positive by reverse transcriptase-PCR. No viral sequence was detected in the feces of 14 chimpanzees. Phylogenetic analysis of the env sequence obtained from SM#13 showed that it clustered within the SIVsm lineage that includes SIVsmH4, B670, and PBj, confirming a direct connection between SIVsm from West Africa and an American-based colony of SM. The virus, designated as SIVsmSL93g, supports a link between the SIVB670/SIVsmH4/SIVPbj lineage and SMs living in Northern Sierra Leone in 1993. The discovery of this strain in a wild-living SM also indicates that noninvasive methods can be used for SIV detection from monkey feces collected in the field. PMID- 15650428 TI - Genetic diversity of HIV type 1 in Likasi, southeast of the Democratic Republic of Congo. AB - To investigate the prevalence of subtypes A and C, and the existence of recombinants of both subtypes in the southeast of the Democratic Republic of Congo (DRC), blood samples were collected from 27 HIV-infected individuals in Likasi, located in an area bordering close to Zambia, and analyzed phylogenetically. Out of the 24 strains with a positive PCR profile for pol-IN and env-C2V3, 15 (62.5%) had a discordant subtype or CRF designation: one subtype A/G (pol/env), four A/U (unclassified), three G/A, one G/CRF01, three H/A, one J/C, one CRF02 (G)/A, and one U/A. Nine (37.5%) strains had a concordant subtype or CRF designation: five subtype A, two C, one D, and one CRF02/G. The remaining three samples negative for PCR with env-C2V3 primers used in this study were further analyzed with env-gp41 primers and revealed the presence of two profiles: two J/J (pol-IN/env-gp41) and one C/G. These data highlight the presence of a high proportion (16/27, 59.3%) of recombinant strains and a low prevalence (4.1 and 7.4%) of subtype C based on env-C2V3 and pol-IN analyses, respectively, in Likasi. In addition, this is the first report that CRF02_AG exists in DRC, though the epidemiological significance of the existence of CRF02_AG in DRC remains unknown. PMID- 15650429 TI - HIV-1 CRF09_cpx circulates in the North West Province of Cameroon where CRF02_AG infections predominate and recombinant strains are common. AB - This study describes the HIV-1 genetic diversity that currently circulates in Bamenda, the provincial capital of the North West province of Cameroon. Phylogenetic analysis of the protease (pro) gene of 20 HIV-1-seropositive individuals identified 11 (55%) CRF02_AG, one D, one F2, one J, and four (20%) unclassifiable strains. Interestingly, the remaining two (10%) samples, 02CMNYU3072 and 03CMNYU3224, originating from epidemiologically unlinked individuals, were classified as CRF09_cpx, representing the first reported cases of this complex circulating recombinant form (CRF) in Cameroon. Additional analysis of the C2V5 portion of the envelope (env) gene confirmed the CRF09_cpx identity of these isolates and classified the remaining isolates as CRF02_AG (n = 12, 63%), subtype D (n = 2, 11%), subtype F2 (n = 2, 11%), and subtype A1 (n = 1). In combination, the pro and env subtyping results revealed three (16%) isolates with discordant subtypes including J( pro )CRF02_AG( env ), CRF02_AG( pro )D( env ), and CRF02_AG( pro )F2( env ). In conclusion, this study highlights the presence of HIV-1 CRF09_cpx in Cameroon and identifies three possible intersubtype recombinants (ISRs) containing CRF02_AG in a town where CRF02_AG infections predominate, and stresses the commonness of HIV-1 recombinant strains in a region where broad genetic diversity exists. PMID- 15650430 TI - The molecular epidemiology of HIV type 1 among Vietnamese Australian injecting drug users in Melbourne, Australia. AB - The proportion of human immunodeficiency virus type 1 (HIV-1) among Vietnamese injecting drug users (IDUs) in Melbourne, Australia exceeds that of the background population. To investigate the molecular epidemiology of HIV-1 among this group, the C2-V4 region of the HIV-1 envelope was directly sequenced from 11 Vietnamese Australians and 19 non-Vietnamese Australian controls. A significant difference in the distribution of the HIV-1 subtypes was demonstrated, with greater than 50% of Vietnamese Australian IDU shown to be infected with CRF01_AE the predominant subtype in Southeast Asia, rather than subtype B, which dominates the Australian epidemic and which was found in 89.5% of the non-Vietnamese controls. The genetic diversity of the CRF01_AE epidemic in Vietnamese Australian IDUs was substantially lower that that of the background subtype B, consistent with a more recent introduction of a limited number of viral strains from Vietnam. These results support public health policy targeting Australian IDUs of Vietnamese ethnicity as a distinct vulnerable population. PMID- 15650431 TI - Sequence variability of the HIV type 1 protease gene in thai patients experienced with antiretroviral therapy. AB - One hundred sequences of HIV-1 protease with the two flanking cleavage sites from 10 antiretroviral drug-treated Thai patients were examined for residue variability and for mutations at positions associated with the resistance to protease inhibitors. Seven patients were infected with CRF01_AE, two with subtype B, and one with a strain that did not belong to any of the currently identified subtypes or recombinant forms. A total of 46 out of the 99 positions (46%) in HIV 1 protease showed at least one amino acid change as compared with the HXB2 subtype B protease. Interestingly, 35% of these mutations were at positions associated with resistance to the PIs. The observation indicated that the viral protease is flexible in tolerating some degree of amino acid substitutions even in the critical regions under the selective pressure driven by antiretroviral drugs. The PR/RT cleavage site revealed conservation of the HXB2 sequence. Nevertheless, the p6*/PR cleavage site displayed variability, notably in the p6* region. PMID- 15650432 TI - Phylogenetic demonstration of two cases of perinatal human immunodeficiency virus type 2 infection diagnosed in adulthood. AB - HIV-2 infection was diagnosed in two patients 15 and 24 years of age and, thereafter, in their mothers. Epidemiological data suggested that vertical transmission was the most probable mode of infection in both patients (Mota Miranda A, et al.: AIDS 2001;15:2460-2461). Phylogenetic analysis of env C2-C3 sequences from the patients and their mothers was used in an attempt to confirm or exclude the events of perinatal HIV-2 transmission. Sequences from each putative transmission pair formed monophyletic clusters in phylogenetic analysis, a clear indication of common ancestry. Interpatient nucleotide distances increased with the period of infection being consistent with long-term infection. In conclusion, the results are consistent with an epidemiological linkage between the viruses infecting each mother-child pair and support the occurrence of perinatal HIV-2 infection in both cases. Application of similar phylogeny methods to other suspected transmission cases may permit a better understanding of the epidemiology and molecular evolution of HIV-2. PMID- 15650434 TI - Interview with Marcelle C. Layton, MD assistant commissioner, Bureau of Communicable Disease New York City Department of Health and Mental Hygiene. Interview by Madeline Drexler. PMID- 15650433 TI - Complete genome analysis of one of the earliest SIVcpzPtt strains from Gabon (SIVcpzGAB2). AB - Chimpanzees in west central Africa (Pan troglodytes troglodytes) are known to harbor simian immunodeficiency viruses (SIVcpzPtt) that represent the closest relatives of human immunodeficiency virus type 1 (HIV-1); however, the number of SIVcpzPtt strains that have been fully characterized is still limited. Here, we report the complete nucleotide sequence of SIVcpzGAB2, a virus originally identified in 1989 in a chimpanzee (P. t. troglodytes) from Gabon. Analysis of this sequence reveals that SIVcpzGAB2 is a member of the SIVcpzPtt group of viruses, but that it differs from other SIVcpzPtt strains by exhibiting a highly divergent Env V3 loop with an unusual crown (NLSPGTT) containing a canonical N linked glycosylation site, an unpaired cysteine residue in Env V4, and two late (L) domain motifs (PTAP and YPSL) in Gag p6. Moreover, phylogenetic analyses indicate evidence of recombination during the early divergence of SIVcpzPtt strains; in particular, part of the pol gene sequence of SIVcpzGAB2 appears to be derived from a previously unidentified SIVcpz lineage ancestral to HIV-1 group O. These data indicate extensive diversity among naturally occurring SIVcpzPtt strains and provide new insight into the origin of HIV-1 group O. PMID- 15650435 TI - Agroterrorism: betting far more than the farm. PMID- 15650436 TI - Factors influencing compliance with quarantine in Toronto during the 2003 SARS outbreak. AB - The purpose of this study was to cull lessons from Toronto's experiences with large-scale quarantine during the outbreak of Severe Acute Respiratory Syndrome in early 2003. We focused on issues that affected the population's willingness to comply with quarantine. Information was acquired from interviews, telephone polling, and focus groups. Issues of quarantine legitimacy, criteria for quarantine, and the need to allow some quarantined healthcare workers to leave their homes to go to work were identified. Also important was the need to answer questions from people entering quarantine about the continuation of their wages, salaries, and other forms of income while they were not working, and about the means by which they would be supplied with groceries and other services necessary for daily living. The threat of enforcement had less effect on compliance than did the credibility of compliance-monitoring. Fighting boredom and other psychological stresses of quarantine, muting the forces of stigma against those in quarantine, and crafting and delivering effective and believable communications to a population of mixed cultures and languages also were critical. The need for officials to develop consistent quarantine policies, procedures, and public messages across jurisdictional boundaries was paramount. PMID- 15650437 TI - The protective oversight of biotechnology. PMID- 15650438 TI - Risk of occupationally acquired illnesses from biological threat agents in unvaccinated laboratory workers. AB - Many vaccines for bioterrorism agents are investigational and therefore not available (outside of research protocol use) to all at-risk laboratory workers who have begun working with these agents as a result of increased interest in biodefense research. Illness surveillance data archived from the U.S. offensive biological warfare program (from 1943 to 1969) were reviewed to assess the impact of safety measures on disease prevention (including biosafety cabinets [BSCs]) before and after vaccine availability. Most laboratory-acquired infections from agents with higher infective doses (e.g., anthrax, glanders, and plague) were prevented with personal protective measures and safety training alone. Safety measures (including BSCs) without vaccination failed to sufficiently prevent illness from agents with lower infective doses in this high-risk research setting. Infections continued with tularemia (average 15/year), Venezuelan equine encephalitis (1.9/year), and Q fever (3.4/year) but decreased dramatically once vaccinations became available (average of 1, 0.6, and 0 infections per year, respectively). While laboratory-acquired infections are not expected to occur frequently in the current lower-risk biodefense research setting because of further improvements in biosafety equipment and changes in biosafety policies, the data help to define the inherent risks of working with the specific agents of bioterrorism. The data support the idea that research with these agents should be restricted to laboratories with experience in handling highly hazardous agents and where appropriate safety training and precautions can be implemented. PMID- 15650439 TI - Challenges in managing volunteers during bioterrorism response. PMID- 15650440 TI - Pediatric terrorism preparedness national guidelines and recommendations: findings of an evidenced-based consensus process. AB - A cadre of experts and stakeholders from government agencies, professional organizations, emergency medicine and response, pediatrics, mental health, and disaster preparedness were gathered to review and summarize the existing data on the needs of children in the planning, preparation, and response to disasters or terrorism. This review was followed by development of evidence-based consensus guidelines and recommendations on the needs of children in disasters, including chemical, biological, and radiological terrorism. An evidence-based consensus process was used in conjunction with a modified Delphi approach for selection of topic areas and discussion points. These recommendations and guidelines represent the first national evidence-based standards for pediatric disaster and terrorism preparedness. PMID- 15650441 TI - Taking the measure of countermeasures: leaders' views on the nation's capacity to develop biodefense countermeasures. PMID- 15650442 TI - Executive government positions of influence in biodefense: the Bio-Plum book. PMID- 15650448 TI - Absorbed dose distribution in glioma tumors in rat brain after therapeutic intratumoral injection of 201Tl-chloride. AB - Studies on animals with gliomas inoculated in the brain and treated with intratumoral injections of 201Tl-chloride have previously shown very promising results, with a survival several weeks longer than controls. Total regression was found in some animals, and necrosis was found in all the 201Tl-treated brain tumors. This study was undertaken to estimate the absorbed dose and dose distribution to the tumor based on the localization and clearance properties obtained from images with two high-resolution imaging techniques; pinhole single photon emission computed tomography (SPECT) and beta-camera. The images from the beta-camera were used to calculate the absorbed dose rate, using an in-house developed, voxel-based Monte Carlo program, based on the EGS4 package. To evaluate the effects of different beta-particle energies on the absorbed dose rate distribution, simulations of medium- and high-energy electrons were conducted. Dose-volume histograms from these simulations show that the energy absorption is very locally distributed for 201Tl and medium energy, whereas high energy beta emitters show a broader dose-volume distribution. The calculated total absorbed dose of 2-8 Gy in the tumor seems to be relativity low when considering the therapeutic effect that was seen. Further investigations, to determine the cause of the high therapeutic efficacy, are needed. PMID- 15650447 TI - Antilymphoma effects of anti-HLA-DR and CD20 monoclonal antibodies (Lym-1 and Rituximab) on human lymphoma cells. AB - AIM: Anti-HLA-DR and anti-CD20 monoclonal antibodies (MAbs) have been effective for immunotherapy and radioimmunotherapy in non-Hodgkin's lymphoma (NHL). The aim of our study was to compare the antilymphoma effects of Lym-1 and rituximab in human lymphoma cell lines, using assays of viability, apoptosis, antibody dependent cellular cytotoxicity (ADCC), and complement-dependent cytotoxicity (CDC), under conditions relevant to the clinic. METHODS: To characterize response relationships at varied concentrations of Lym-1 and rituximab, growth inhibition and cell death were assayed over 96 hours in four NHL cell lines derived from Burkitt's or large-cell lymphoma patients. Untreated cells and cells treated with an mLym-1 isotype-matched MAb were used as negative controls for direct assays. Western blot was used to detect apoptosis through the activation of caspase-3 and cleavage of poly (ADP-ribase) polymerase (PARP). The indirect cytotoxicity of Lym 1 and rituximab was assayed at varied concentrations, using ADCC activity in the presence of purified peripheral blood leukocytes and CDC activity in the presence of human donor serum. RESULTS: Lym-1 and rituximab showed significant direct and indirect antilymphoma effects. Lym-1 had a substantial, and statistically greater, effect than rituximab over longer intervals of time. In Raji and B35M cells, Lym-1 induced potent growth inhibition reflected by 90% and 94% reductions in viable cells, respectively, whereas rituximab induced 63% and 56% reductions. Concurrently, Lym-1 increased nonviable cells by 372% and 153% in these cells, respectively, whereas rituximab induced 139% and 43% increases. Lym-1-induced apoptosis was greater than that of rituximab in all cell lines tested. Lym-1, both the chimeric form and the mouse parent, mediate ADCC more effectively, in the presence of a total peripheral blood leukocyte (PBL) population, than does rituximab, although the results for CDC activity were mixed. CONCLUSIONS: In conclusion, Lym-1 had more potent direct and indirect cytotoxic effects than rituximab in lymphoma cells under conditions achievable in patients. Because the HLA-DR target antigen of Lym-1 is enriched on most B-cell lymphomas, these results support its complementary use in patients as an alternative to CD20 for monoimmunotherapy and for combination immunotherapy with rituximab, because the HLA-DR and CD20 antigens are physically and functionally coupled on human B cells. PMID- 15650449 TI - Autologous tumor cell line-derived vaccine for patient-specific treatment of advanced renal cell carcinoma. AB - AIM: We previously reported the laboratory methodology for producing patient specific irradiated autologous tumor-cell products derived from short-term cultured tumor cells from resected renal cell carcinoma, and described preliminary clinical results. In this study, we report the final clinical results and efforts to define vaccine potency on the basis of clinical outcome for these 25 patients with advanced renal cell carcinoma. MATERIALS AND METHODS: Approximately 10(8) cells from successful short-term cell lines were irradiated, frozen in aliquots of 10(7) cells, then thawed and administered subcutaneously (s.c.) once a week for 3 weeks, then once a month for 5 months. Patients included 19 men and 6 women, who were 43-82 years of age. Six (6) patients had a large primary lesion, 2 patients had regionally advanced disease, 3 patients had been rendered disease-free by surgical resection of distant metastases, and 14 patients had measurable distant metastatic disease. RESULTS: The vaccines were well tolerated, and no delayed autoimmune effects were documented. Delayed-type hypersensitivity (DTH) tests of irradiated tumor cells were positive in only 1 of 25 patients at week 0, but converted to positive in 6 of 18 patients of DTH negative patients who were retested at week 4. Objective response rate in patients who had measurable metastatic disease was 0 of 14 patients. With a median follow-up of greater than 7 years from the date of the first DTH test, median survival is 33.4 months, 5-year survival is 43%, and 10 patients are alive 3-12 years later. The 7 DTH+ patients survived a median of 2.5 years, and 3 patients are alive after 3, 4, and 7 years. There was no correlation between the number of irradiated cells or viable irradiated cells injected and tumor DTH reactivity or survival. CONCLUSION: This approach is feasible and the therapy is well tolerated, but clinical benefit was not established in this trial. Any further exploration of this product should be limited to the adjuvant setting in a randomized trial. PMID- 15650450 TI - Phase I/II trial of autologous tumor cell line-derived vaccines for recurrent or metastatic sarcomas. AB - AIM: We previously reported the laboratory methodology for producing patient specific irradiated autologous tumor-cell products derived from short-term cultured tumor cells. We attempted to determine the feasibility, safety, and clinical effects of autologous tumor vaccine-derived sarcomas. PATIENTS AND METHODS: Efforts were made to establish tumor cell lines in tissue culture with expansion to 100 million cells for patients who were candidates for therapy. Cells were irradiated and cryopreserved in aliquots of 10 million cells for subcutaneous (s.c.) injections, once a week for 3 weeks, then once a month for 5 months. RESULTS: Efforts were made to establish short-term tumor cell lines from 86 fresh sarcoma specimens (10 primary, 14 recurrent, and 62 metastatic). Initial growth was successful for 48 patients (56%), and cultures were expanded for 36 patients, with 25 patients treated. There were 23 evaluable patients, including 12 women and 11 men, with a median age of 52 years and a range from 16-79 years. Vaccine therapy was well tolerated. Delayed-type hypersensitivity (DTH) tests to irradiated tumor cells were positive in 0 of 20 patients tested at baseline, but converted to positive after 3 weekly vaccinations in 8 of 16 patients who were retested. Median survival for the 8 DTH converters was 16.6 months versus 8.2 months for the 8 responders whose tumor DTH test remained negative, and 6.0 months for the 7 patients who were not tested. No objective responses were recorded among 12 evaluable patients with measurable disease; 10 patients have survived more than 1 year. CONCLUSIONS: This approach is feasible, well tolerated, and the resulting DTH conversion rate is of interest. Patients with minimal tumor burden would be preferred for further future testing. PMID- 15650451 TI - Phase I clinical trial of targeted therapy using 131I-Hepama-1 mAb in patients with hepatocellular carcinoma. AB - OBJECTIVES: The aim of this study was to evaluate the response- and treatment related toxicity for patients with unresectable hepatocellular carcinoma (HCC) treated with 131I-Hepama-1 mAb (DGDK-1). MATERIALS AND METHODS: Seeking approval for the use of 131I-hepama-1 mAb, 32 patients with unresectable HCC were divided into 5 groups to take part in the Phase I clinical trial study. Systemic reactions and acute toxicity were evaluated weekly for at least 45 days following treatment with an intravenous injection of 10 mg of antibody radiolabeled with 740-3700 MBq (20-100 mCi), divided into 5 groups. RESULTS: No patients demonstrated any evidence of fever, severe side effects, or impairments of important organs, such as the lung, liver, and heart. During, and for 45 days after, the treatment, most of their routine examinations of blood, functions of liver and kidneys, and the thyroid hormone were all normal. Hypothyroidism was not observed during the course of monitoring. None of the 32 patients developed human antimurine antibodies (HAMA) within 15 days of therapy; however, 82% of patients were HAMA-positive by 45 days of therapy. According to the pilot study of prognosis, survival from the start of radioimmunotherapy (RIT) was a median of 4 months, and the 1-year survival rate was 31%. The median survival time of 15 patients without metastases was 10 months, and the 1-year survival rate was 60%. Of alpha-fetoprotein (AFP)-positive patients, 75% of AFP levels decreased by at least 50% or became normal. Of all patients, 84% showed an improved Eastern Cooperative Oncology Group (ECOG) performance status. CONCLUSION: This phase I clinical trial demonstrated that it was safe to use 131I-hepama- 1 mAb by peripheral intravenous administration. The recommended dose of DGDK-1 for clinical use is 1480-2960 MBq/10 mg. PMID- 15650452 TI - Effect of mitomycin C and vinblastine on FDG uptake of human nonsmall-cell lung cancer xenografts in nude mice. AB - This study was designed to preliminarily evaluate the use of positron emission tomography (PET) with [18F]-2-fluoro-2-deoxy-D-glucose (FDG) for monitoring chemotherapy effects, using a nude-mouse model of human nonsmall-cell lung cancer (NSCLC), the Lu-99 cell line. Tumor-FDG uptakes and volumes were measured after administrating a single dose of mitomycin (MMC) and vinblastine (VLB) and then compared these for a nontherapy group. A significant reduction in tumor volume after either chemotherapy occurred and associated with significantly lower FDG uptake values than the control group (p < 0.001), as early as day 1. These observations suggest that FDG-PET may be useful for noninvasively monitoring the effects of cancer chemotherapy. PMID- 15650453 TI - 18F-FDG PET early after radiotherapy in lymphoma patients. AB - OBJECTIVE: The aim of this study was to evaluate the rate of postactinic inflammatory alterations that could lead to false-positive results in FDG-PET images, in a group of lymphoma patients studied with positron emission tomography (PET) early after the end of radiation therapy. MATERIALS AND METHODS: Sixteen (16) consecutive patients were referred to our center for malignant lymphoma; 14 of 16 patients had a mediastinal bulky mass at diagnosis. Each patient underwent chemotherapy and then radiotherapy (RT): for clinical reasons, shortly after RT (range, 25-56 days; mean, 38.7 days) a FDG PET scan was required to evaluate the effect of therapy. We intravenously injected 370 MBq of 18F-FDG, and after 60-90 minutes we recorded images. RESULTS: Despite a relatively short time after RT, there was no pathological tracer uptake in 13 of 16 patients. In 3 cases, a mild increase in FDG uptake was observed, but no findings which would lead to a false positive diagnosis. In 2 of 3 cases, postactinic pneumopathy was diagnosed (PET scan performed 51 and 52 days after RT); while in 1 patient, soft-tissue inflammation was present (PET scan performed 42 days after RT). CONCLUSION: Our data indicates that the rate of postactinic PET inflammatory alterations in lymphoma patients is not very high and appear to be not strictly linked to the elapsed time since the end of RT treatment. PMID- 15650454 TI - Efficacy of 99mTc-EDDA/HYNIC-TOC scintigraphy in differential diagnosis of solitary pulmonary nodules. AB - Fifty consecutive patients with solitary pulmonary nodules (SPN) on chest radiographs were studied scintigraphically after the administration of a somatostatin analog 99mTc-EDDA/HYNIC-TOC. The activity amounted to 740-925 MBq and a single photon emission computed tomography (SPECT) technique was applied. Verification of the nodule etiology was based on histology or cytology and bacteriology. As additional criterion for nodule benignity, its stable size in a chest radiograph for at least 3 years was accepted. In 31 patients, malignant etiologies of nodules were found. The diagnoses included: 11 adenocarcinomas, 6 squamous-cell carcinomas, 2 large-cell carcinomas, 6 nonsmall-cell lung cancers (NSCLC) of unspecified, more detailed morphology, 2 small-cell lung cancers (SCLC), 2 typical carcinoids, and 2 metastatic tumors: leiomyosarcoma and malignant melanoma. In 19 patients, the following benign tumors were diagnosed: 6 tuberculomas, 2 other granulomas, 4 hamartomas, 2 nonspecific inflammatory infiltrates, 1 abscess, 1 peripheral carcinoid of morphological characteristics of a benign tumor, 1 ectopic lesion of thyroid tissue, and 2 benign tumors of unspecified etiology, with stable size over 3 and 5 years. Positive scintigraphic results were obtained in 28 of 31 patients (90%) with malignant SPNs; among these there were 26 of 27 (96%) cases of primary pulmonary carcinoma. The remaining 2 false-negative cases included metastatic tumors: liposarcoma and melanoma. Among 19 benign lesions, 15 (79%) did not accumulate the radiopharmaceutical. The remaining 4 tumors visible on scintigrams included: 1 tuberculoma, 1 hamartoma, 1 abscess, and 1 case of nonestablished diagnosis (with stable size over 3 years). In conclusion, scintigraphy with 99mTc-EDDA/HYNIC-TOC appears to be an effective procedure for differentiation between malignant and benign SPNs. PMID- 15650455 TI - Clinical utility of 99mTc-Sestamibi scintimammography in the management of equivocal breast lesions. AB - The aim of this study was to assess the utility of 99mTc-sestamibi scintimammography (SM) in patients with suspected primary or recurrent breast cancer. Forty-four (44) breast lesions (17 with suspected recurrence of disease) in 40 patients were included into the study. In these patients, the results of conventional diagnostic methods were equivocal or inconclusive. Twenty-one (21) lesions were palpable and 23 lesions were not. Histological examinations performed during the follow-up revealed malignancy in 24 specimens. SM correctly identified 21 of them, as well as 12 true negatives. There were 8 false-positive studies; therefore, the sensitivity of SM was 87.5%, specificity was 60%, positive predictive value (PPV) was 72.4%, and the negative predictive value (NPV) was 80%. The sensitivity in palpable lesions was 100%; three (3) false negatives, 1 recurrence, and 2 cancers, all of them nonpalpable. In conclusion, SM is useful in equivocal palpable lesions for resolving diagnostic uncertainty after conventional examination, and can limit the number of surgical interventions for benign disease. However, its use in nonpalpable tumors is not recommended. PMID- 15650456 TI - Preclinical investigations of drug and radionuclide conjugates of bisphosphonates for the treatment of metastatic bone cancer. AB - The potential targeting of therapeutic bisphosphonate conjugates to bone metastatic lesions was evaluated in vivo in mice. A bisphosphonate conjugate with 5-fluorouracil was synthesized as a potential chemotherapy agent, and a bisphosphonate conjugate with diethylenetriaminepentaacetic acid (DTPA) was prepared as a potential carrier of cytotoxic radionuclides. The compounds are hypothesized to be able to deliver either high doses of radiation or a high concentration of chemotherapy agents at sites of increased osteoclastic activity in patients with bony metastases while exhibiting minimal toxicity to normal tissues. Tissue distribution studies with the 99mTc-labeled bisphosphonate conjugates with DTPA and 5-fluorouracil showed rapid blood clearance and excretion of unbound activity, clearance from most tissues, and substantial retention of the bisphosphonates in bone. For the DTPA conjugate, activity in the bone represents 13.6% of the total injected dose at 8 hours following injection, representing 54.3% of the total whole-body activity at this time period. Under the same conditions, the 5-fluorouracil conjugate showed a 17.1% bone uptake at 60.2% of the whole-body activity. This normal bone uptake predicts that high concentrations of conjugates are expected to be achieved at sites of bone metastatic disease. Chemotherapy and radiotherapy studies with these compounds in animal models of metastatic bone cancer are underway. PMID- 15650457 TI - The cytotoxicity of Scytosiphon lomentaria against HL-60 promyelocytic leukemia cells. AB - This study examined the cytotoxicity of Scytosiphon lomentaria, using various cancer cell lines. The ethyl acetate (EtOAc) fraction of this alga showed the cytotoxicity to leukemia cells, including HL-60. When HL-60 cells were treated with its EtOAc fraction, several apoptotic characteristics, such as DNA fragmentation, chromatin condensation, and an increase of the population of sub G1 hypodiploid cells, were observed. Moreover, the EtOAc fraction decreased c-Myc expression in a dose-dependent manner. In order to understand the mechanism of apoptosis induction by S. lomentaria, we examined the changes of Bcl-2 and Bax protein expression levels. The EtOAc fraction reduced Bcl-2, an antiapoptotic protein, but increased Bax, a proapoptotic protein, in a dose-dependent manner. When we examined the activation of caspase-3, an effector of apoptosis, the expression of the active form (19 kDa) of caspase-3 increased, and the increase of their activities was demonstrated by the cleavage of poly (ADP-ribose) polymerase, a substrate of caspase-3, to 85 kDa. The results suggest that the inhibitory effect of S. lomentaria on the growth of HL-60 appears to arise from the induction of apoptosis by way of the down-regulation of Bcl-2 and the activation of caspase. PMID- 15650458 TI - A combination of flk1-based DNA vaccine and an immunomodulatory gene (IL-12) in the treatment of murine cancer. AB - AIM: The aim of this study was to investigate the antivasculature effects and the antitumor effects of combining attenuated Salmonella typhimurium vaccine strain encoding murine vascular endothelial growth factor (VEGF) receptor-2 (flk1) with plasmid DNA vector encoding the murine IL-12 (mIL-12) gene. METHODS: Mouse models of Gl261 glioblastoma were treated with combining orally given attenuated Salmonella typhimurium vaccine strain encoding flk1 with direct intratumoral injection of a nonviral plasmid DNA vector encoding the murine IL-12 (mIL-12) gene. The volumes of tumors were observed. Cytolytic T lymphocyte (CTL) response was measured by a 4-hour 51Cr release assay, vessle density and tumor cell proliferation were observed by immunostaining, and tumor apoptosis was determined by TUNEL staining. RESULTS: Compared to mice receiving single agent therapy, received either oral immunization flk1-based vaccine only or the therapeutic gene IL-12 plasmid DNA only or those in the control group, the combination therapy groups developed a strong CTL response and showed more significantly inhibited tumor growth, apoptosis of tumor cells, and reduced neovascularization and cell proliferation in these mice. CONCLUSIONS: The therapy of attenuated Salmonella typhimurium vaccine strain encoding flk1 combined with the interleukin-12 gene has significant synergistic effect against tumors. PMID- 15650459 TI - Phase I/II trial of melanoma patient-specific vaccine of proliferating autologous tumor cells, dendritic cells, and GM-CSF: planned interim analysis. AB - AIM: The aim of this study was to investigate the feasibility, safety, and clinical efficacy of patient-specific dendritic cell vaccines in patients with metastatic melanoma. A planned interim analysis was conducted on the first 20 patients. METHODS: Tumor cell lines were established from metastatic tumor, expanded to 200 million cells, and then incubated with interferon-gamma for patients who were candidates for therapy. Cells were irradiated and cryopreserved. Patients underwent leukapheresis to obtain mononuclear cells that were cultured in the presence of IL-4 and GM-CSF to produce dendritic cells, which were incubated with cryopreserved, irradiated tumor cells, and then stored in aliquots of about 20 million cells for subcutaneous (s.c.) injections with GM CSF once a week for 3 weeks, then once a month for 5 months. RESULTS: The first 20 eligible patients included 10 men and 10 women, with a median age of 48 years (range, 16-79 years). Three (3) patients had brain metastases, and 13 patients had experienced disease progression after biochemotherapy. At the time of vaccine treatment, 6 patients had evaluable metastatic disease, while 14 patients lacked measurable disease. Vaccine therapy was well tolerated, except for what appeared to be GM-CSF-related allergic reactions in 2 patients. Delayed-type hypersensitivity (DTH) tests to irradiated tumor cells were positive in 0 of 20 patients tested at baseline, but converted to positive in 8 patients (40%). At a median follow-up of 13.8 months, there is a 95% overall survival and a 48% progression-free survival. Four (4) patients have already survived more than 3.0 years since starting the vaccine. CONCLUSION: Based on tolerability, rate of tumor DTH conversion, and encouraging survival, the trial will continue accrual to at least 19 patients with measurable disease and 40 patients who lack measurable disease at the time of treatment. PMID- 15650460 TI - Our place in nature: reconnecting with the Earth for better sleep. PMID- 15650461 TI - Effects of Morinda citrifolia on quality of life and auditory function in postmenopausal women. PMID- 15650462 TI - Tradition and modernity in Mongolian medicine. PMID- 15650463 TI - The healing presence: can it be reliably measured? AB - Religious and spiritual traditions from all cultures and times describe a spiritual or loving presence as a contributor to healing. In addition, there is a common belief that a special "presence" can exude from certain practitioners. Is it possible to measure a healing presence in an objective and reliable way? Most research on healing has focused on trying to prove spiritual and "energy" healing in comparative tests. However, if a measure of healing presence is to be sensitive and reliable, objective and real-time indicators of such a presence will be needed rather than comparative and statistical outcomes. This paper discusses what healers from various traditions have felt are the primary components of a healing presence, summarizes various attempts to measure healing, and describes two recently published approaches that have potential to provide such an objective and realtime indicator of a healing presence. PMID- 15650464 TI - Hemodynamic observations on a yogic breathing technique claimed to help eliminate and prevent heart attacks: a pilot study. AB - OBJECTIVE: This pilot study investigated the hemodynamics of a yogic breathing technique claimed "to help eliminate and prevent heart attacks due to abnormal electrical events to the heart," and to generally "enhance performance of the central nervous system (CNS) and to help eliminate the effects of traumatic shock and stress to the CNS." DESIGN: Parameters for (4) subjects were recorded during a preexercise resting period, a 31-minute exercise period, and a postexercise resting period. SETTINGS/LOCATION: Parameters for subjects were recorded in a laboratory at the University of California, San Diego. SUBJECTS: Parameters for 3 males (ages 44, 45, 67) and 1 female (age 41) were recorded. One (1) subject (male age 45) had extensive training in this technique. INTERVENTIONS: This yogic technique is a 1 breath per minute (BPM) respiratory exercise with slow inspiration for 20 seconds, breath retention for 20 seconds, and slow expiration for 20 seconds, for 31 consecutive minutes. OUTCOME MEASURES: Fourteen beat-to beat parameters were measured noninvasively and calculated for body surface area to yield: stroke index (SI), heart rate (HR), cardiac index, end diastolic index, peak flow, ejection fraction, thoracic fluid index, index of contractility, ejection ratio, systolic time ratio, acceleration index, and systolic, diastolic, and mean arterial pressures (MAPs). Left stroke work index (LSWI) and stroke systemic vascular resistance index (SSVRI) were calculated. RESULTS: We report on SI, HR, MAP, LSWI, and SSVRI and how they can help to describe hemodynamicstate changes. This technique induces dramatic shifts in all hemodynamic variables during the 1 BPM exercise and can produce unique changes in the postexercise resting period after long-term practice that appears to have a unique effect on the brain stem cardiorespiratory center regulating the Mayer wave (0.1-0.01 Hz) patterns of the cardiovascular system. CONCLUSIONS: Preclinical studies are warranted to examine the possible long-term effects of this technique that appear to reset a cardiorespiratory brain-stem pacemaker. We postulate that this effect may be the basis for the purported yogic health claim. PMID- 15650465 TI - The biologic effects of grounding the human body during sleep as measured by cortisol levels and subjective reporting of sleep, pain, and stress. AB - OBJECTIVES: Diurnal cortisol secretion levels were measured and circadian cortisol profiles were evaluated in a pilot study conducted to test the hypothesis that grounding the human body to earth during sleep will result in quantifiable changes in cortisol. It was also hypothesized that grounding the human body would result in changes in sleep, pain, and stress (anxiety, depression, irritability), as measured by subjective reporting. SUBJECTS AND INTERVENTIONS: Twelve (12) subjects with complaints of sleep dysfunction, pain, and stress were grounded to earth during sleep for 8 weeks in their own beds using a conductive mattress pad. Saliva tests were administered to establish pregrounding baseline cortisol levels. Levels were obtained at 4-hour intervals for a 24-hour period to determine the circadian cortisol profile. Cortisol testing was repeated at week 6. Subjective symptoms of sleep dysfunction, pain, and stress were reported daily throughout the 8-week test period. RESULTS: Measurable improvements in diurnal cortisol profiles were observed, with cortisol levels significantly reduced during night-time sleep. Subjects' 24-hour circadian cortisol profiles showed a trend toward normalization. Subjectively reported symptoms, including sleep dysfunction, pain, and stress, were reduced or eliminated in nearly all subjects. CONCLUSIONS: Results indicate that grounding the human body to earth ("earthing") during sleep reduces night-time levels of cortisol and resynchronizes cortisol hormone secretion more in alignment with the natural 24-hour circadian rhythm profile. Changes were most apparent in females. Furthermore, subjective reporting indicates that grounding the human body to earth during sleep improves sleep and reduces pain and stress. PMID- 15650466 TI - Long-term effects of integrated rehabilitation in patients with advanced angina pectoris: a nonrandomized comparative study. AB - OBJECTIVES: An evaluation of Integrative Rehabilitation (IR) of patients with angina pectoris with respect to death rate, the need for invasive treatment, and cost effectiveness. DESIGN: A report from a clinical database. Death rates were compared to those of the general Danish population matched for age, gender, and observation period, as well as with data from the literature concerning medical and invasive treatments. SETTING: The treatment was carried out as an ambulatory treatment in a private clinic. SUBJECTS: One hundred and sixty-eight (168) patients with angina pectoris, of whom 103 were candidates for invasive treatment and 65 for whom this had been rejected. INTERVENTIONS: Integrated rehabilitation consists of acupuncture, a self-care program including acupressure, Chinese health philosophy, stress management techniques, and lifestyle adjustments. OUTCOME MEASURES: Death rate from any cause, the need for invasive treatment, and health care expenses. RESULTS: The 3-year accumulated risk of death was 2.0% (95% confidence limits: 0.0%-4.7%) for the 103 candidates for invasive treatment, 6.4% for the general Danish population, 5.4% (4.7%-6.1%), and 8.4% (7.7%-9.1%) for patients who underwent percutaneous transluminal balloon angioplasty and coronary artery bypass grafting, respectively, in New York. For the 65 inoperable patients the risk of death due to heart disease was 7.7% (3.9%-11.5%), compared to 16% (10%-34%) and 25% (18%-36%) for American patients, who were treated with laser revascularization or medication, respectively. Of the 103 candidates for invasive treatment, only 19 (18%) still required surgery. Cost savings over 3 years were US 36,000 dollars and US 22,000 dollars for surgical and nonsurgical patients, respectively. These were mainly achieved by the reduction in the use of invasive treatment and a 95% reduction in in-hospital days. CONCLUSIONS: Integrated rehabilitation was found to be cost effective, and added years to the lives of patients with severe angina pectoris. The results invite further testing in a randomized trial. PMID- 15650467 TI - The unexpected outcomes of acupuncture: case reports in support of refocusing research designs. AB - Two distinct positive outcomes are possible with Classical Acupuncture; resolution of the "main complaint" for which the patient is seeking treatment and unexpected resolution of health concerns for which the patient is not seeking treatment. Two clinical cases are presented to illustrate this phenomenon clearly. In both cases, the unexpected outcomes follow logically from the central therapeutic imperative of Oriental medicine; treatment of both the patient's root and branch. Research designs focused on single health conditions do little to reflect this central clinical reality. To highlight this problem, the two cases presented here are repositioned within the framework of research designs focused on single health conditions. It becomes clear that any eventual "gold standard" of acupuncture-appropriate research designs would reflect the full health care service provided by Classical Acupuncture. This, in turn, would ensure that health care policies based on research results would be appropriate to the realities of clinical acupuncture. PMID- 15650468 TI - Acupuncture as a complex intervention: a holistic model. AB - OBJECTIVES: Our understanding of acupuncture and Chinese medicine is limited by a lack of inquiry into the dynamics of the process. We used a longitudinal research design to investigate how the experience, and the effects, of a course of acupuncture evolved over time. DESIGN AND OUTCOME MEASURES: This was a longitudinal qualitative study, using a constant comparative method, informed by grounded theory. Each person was interviewed three times over 6 months. Semistructured interviews explored people's experiences of illness and treatment. Across-case and within-case analysis resulted in themes and individual vignettes. SUBJECTS AND SETTINGS: Eight (8) professional acupuncturists in seven different settings informed their patients about the study. We interviewed a consecutive sample of 23 people with chronic illness, who were having acupuncture for the first time. RESULTS: People described their experience of acupuncture in terms of the acupuncturist's diagnostic and needling skills; the therapeutic relationship; and a new understanding of the body and self as a whole being. All three of these components were imbued with holistic ideology. Treatment effects were perceived as changes in symptoms, changes in energy, and changes in personal and social identity. The vignettes showed the complexity and the individuality of the experience of acupuncture treatment. The process and outcome components were distinct but not divisible, because they were linked by complex connections. The paper depicts these results as a diagrammatic model that illustrates the components and their interconnections and the cyclical reinforcement, both positive and negative, that can occur over time. CONCLUSIONS: The holistic model of acupuncture treatment, in which "the whole being greater than the sum of the parts," has implications for service provision and for research trial design. Research trials that evaluate the needling technique, isolated from other aspects of process, will interfere with treatment outcomes. The model requires testing in different service and research settings. PMID- 15650469 TI - The effectiveness of electroacupuncture versus electrical heat acupuncture in the management of chronic low-back pain. AB - DESIGN: Forty-two (42) subjects suffering from chronic low-back pain were matched with the nature of their occupations and then randomly allocated into: (1) an electroacupuncture group (EA); (2) an electrical heat acupuncture (EH) group or; (3) a control group. INTERVENTIONS: Subjects in the EA group and the EH group received treatment for 20 minutes on a total of 6 acupuncture points. Treatment was delivered twice per week for 4 weeks (a total of 8 sessions). Back exercise was taught to all subjects including the control group as a home program. OUTCOMES MEASURES: A numerical rating scale of pain (NPRS), straight leg raise (SLR), and Roland Morris Disability Questionnaire (RMDQ) were recorded. RESULTS: There were significant reduction of NPRS within the EA (p = 0.000), EH (p = 0.000), and control (p = 0.013) groups across sessions. Significant between-group differences were shown in session 4 (p = 0.006), session 8 (p = 0.001), and 1 month follow-up sessions (p = 0.001). Posthoc tests showed that the NPRS of the EH group was significantly lower than that of the EA group and the control group by session 4 (p = 0.004). After session 8, the NPRS of both the EA group (p = 0.003) and the EH group (p = 0.001) were significantly lower than that of the control group. Such a difference was maintained at least up to the 1-month follow up. Only the EA group had significant improvement in the measurement of SLR across sessions (p = 0.000). The between-group difference reached significance level in session 8 (p = 0.001) and at 1-month follow-up (p = 0.002). Posthoc tests showed that EA group had significantly greater gain than the EH group and the control group. For the RMDQ score, the improvement was statistically significant within each of the three groups over time (p = 0.000). However, the between-group difference did not reach statistical significance. CONCLUSIONS: Our findings suggest that 4 sessions of EH treatment over 2 weeks produced significantly greater reduction in the NPRS than that of the EA or the control. However, EA produced greater improvement in SLR and reduction in RMDQ score than that of the EH and the control. PMID- 15650470 TI - Complementary and alternative therapy use at end-of-life in community settings. AB - OBJECTIVE: To investigate the prevalence of complementary and alternative medicine (CAM) use by an end-of- life population. DESIGN: Random selection of death certificates used to locate family caregivers who were interviewed by telephone 2-5 months following decedents' deaths. PARTICIPANTS: Decedent subjects died of natural causes in community settings. Family caregivers were very involved in the care and decision making for decedents during their terminal illness. MEASURE: Family caregivers reported on whether decedents had used CAM, type of modality, and motivation for use. RESULTS: CAM use by decedents was reported by 53.7% of family caregivers. Decedents who had used CAM were more likely to be younger, to have college degrees and higher household incomes, and to have used one or more life-sustaining treatment. The most frequent reason the decedents had used CAM was for symptom relief. CONCLUSIONS: As baby boomers age, bringing their CAM familiarity and previous practices into the end-of-life phase, clinicians will need to be aware that CAM use for symptom control is likely to be prevalent. PMID- 15650471 TI - A pilot study to examine the effects of a mindfulness-based stress-reduction and relaxation program on levels of stress hormones, physical functioning, and submaximal exercise responses. AB - OBJECTIVE: Stress has been cited as a causal factor in heart disease. The objective of this study was to examine the effects of an 8-week mindfulness-based stress-reduction program on the resting levels of stress hormones, physical functioning, and submaximal exercise responses in women with heart disease. SUBJECTS: Random selection with the numbers 1 and 2 were used to assign 18 women (60 +/-6.3 years old) with documented histories of heart disease to a treatment group (n = 9) or a control group (n = 9). Speilberger's state anxiety scores for the treatment (M = 37.88; standard deviation (SD) = 10.91) and control group (M = 43.22; SD = 12.26) were not significantly different prior to the start of the study. However, their scores fell in the upper percentile rank for normal adults in their age category. INTERVENTION: The intervention was provided one night each week for 2 hours over a period of 8 weeks. The intervention included didactic, inductive, and experiential modes of learning regarding stress responses and mindfulness skill-development training. DESIGN: Pre-post test hormonal measurements and physical function were analyzed using a 2 (group) by 2 (time) analysis of variance (ANOVA) with repeated measures following the 8-week program. Submaximal exercise responses were also compared between the treatment group and the control group following the 8-week program. A 2 (group) by 3 (time) ANOVA with repeated measures was used to analyze the data. SETTINGS/LOCATION: Weekly meetings were held on a university medical school campus. Submaximal exercise responses were recorded while participants cycled on a stationary bike in an applied physiology laboratory following the 8-week program. RESULTS: There were no significant main effects or interaction for the resting levels of stress hormones or physical functioning. There were no significant interactions for the submaximal exercise responses, however, there were significant main effects between groups for ventilation [F(2,32) = 7.65, p < .01, f = 0.8], and between group [F(1,16) = 8.84, p < .01, f = 0.8] and time [F(2,32) = 10.42, p < .01, f = 0.9], for breathing frequency. CONCLUSION: While the 8-week stress reduction program for women with heart disease did not show significant interactions between groups for resting levels of stress hormones, physical functioning, or submaximal exercise responses, there was a significant difference in breathing patterns between the 2 groups during exercise following the mindfulness-based stress-reduction program. There was also a trend for change in the intervention group in the resting levels of cortisol and physical function scores that was not seen in the control group. Future studies could use the effect size generated from this pilot study to calculate the number of subjects needed for adequate power to detect significant differences between groups. PMID- 15650472 TI - Does homeopathically potentized antimony stimulate coagulation? A summary of previous findings and results of an in vitro pilot study by means of thrombelastography. AB - BACKGROUND: Potentized antimony is traditionally used in anthroposophic medicine to enhance hemostasis in bleeding disorders, but evidence of its effectiveness is scarce. On the other hand, non-toxic and economic additional therapeutic options for hemostatic disorders are desirable. OBJECTIVES: We examined all available literature on the subject and performed a controlled pilot in vitro study to test the procoagulatory potency of antimony D 5. DESIGN: Freshly drawn citrated whole blood of 12 healthy volunteers and 12 patients with bleeding disorders was equally distributed into 344 portions, after which it was mixed with antimony D 5, or its potentized vehicle (lactose D 5) as control solution and tested with thrombelastography. The paired t-test and the Wilcoxon signed rank test were used for statistical analysis. In 5 of the 12 healthy donors, a second blood sample was drawn to assess individual variability and increase the total number of replicates. Thus three separate calculations were performed: for the 12 patients, the 12 healthy donors, and the 5 later samples from the same donors. The analysis was exploratory, and no Bonferroni correction was applied. RESULTS: In the antimony D5 samples of the 12 healthy subjects, but not the patients, there was a tendency toward a shorter clotting time (CT) (p = 0.074) and a trend for an increased clot firmness, expressed as maximal amplitude (MA) (p = 0.058). The increase of MA was significant (p = 0.011) when the later samples were included. No statistical difference was detected for the clot formation time and the clot lysis index. CONCLUSION: The exploratory results of this pilot study are inconclusive as to whether antimony D5 has a procoagulatory effect in vitro, although the results suggest an effect on MA and possibly CT. More research is warranted. PMID- 15650473 TI - Effects of topical elk velvet antler on cutaneous wound healing in streptozotocin induced diabetic rats. AB - OBJECTIVE: Wound repair is a finely orchestrated process involving cellular, molecular, physiologic, and biochemical interactions that restore the integrity of damaged tissue. Cyclic replacement of deer antlers requires rapid regenerative growth, in many ways analogous to that encountered during tissue repair. Molecular mechanisms regulating these processes are not fully understood, but it is increasingly apparent that growth factors are important mediators. Previous studies have shown that elk velvet antler (EVA) contains various growth factors and that a water-soluble extract stimulates dermal fibroblast growth in vitro. DESIGN: The efficacy of EVA water-soluble extract on wound healing in streptozotocin-induced diabetic rats was EVAluated using a full-thickness cutaneous wound model. Animals were randomly selected to receive topical application of either control or EVA gel. Daily photographs of the wounds served to measure the rate of wound closure. Wound-edge biopsies obtained on postoperative days 2 and 10 allowed histologic evaluation and measurement of transforming growth factor-beta 1 (TGF-beta (1)) concentrations by enzyme-linked immunoabsorbent assay. RESULTS: Wounds treated with the EVA topical gel were significantly smaller by postoperative day 6. TGF- beta (1) protein expression was not different in EVA-treated wounds compared to control wounds. CONCLUSIONS: This study indicates that topical treatment with an EVA water-soluble extract accelerates repair of cutaneous wounds in diabetic rats. Further studies are warranted to reveal the mechanisms involved in EVA enhancement of wound closure and to determine if this compound is an economical pharmacologic agent in the treatment of normal and compromised wounds. PMID- 15650474 TI - A pharmacologic study on the mechanism of action of Kakkon-to: body temperature elevation and phagocytic activation of macrophages in dogs. AB - OBJECTIVE: The phagocytic activity of macrophages as a novel approach to scientific elucidation of the effects of Chinese medicines was studied through administration of a kampo preparation, by measuring the rise in body temperature, which is thought to stimulate innate defensive functions of organisms and enhance the immune systems. DESIGN: Using dogs as experimental models, a rise in body temperature following administration of Kakkon-to was observed, and the average number and average rate of phagocytosis of macrophages in blood using latex micro particles was investigated. RESULTS: The body temperature of the treated animals significantly increased 30 minutes after administration (p<0.01), and remained elevated for more than 5 hours. A comparison of body temperatures before and after administration showed significant increases over controls from 1 to 11 hours, p<0.01; and at 12 hours, p<0.05 after administration. The average number and the average rate of phagocytosis were significantly increased 1 (p<0.05) and 2 (p<0.01) hours after administration. The mean number of phagocytized cells significantly increased (p<0.05) at 1 hour after administration compared with that before administration, and the mean phagocytic rate also increased significantly (p<0.01) 2 hours after administration. Increases (p<0.01) in both the rate of phagocytosis and the number of cells phagocytized were found at every measurement point from 2 to 24 hours after administration. Significant increases (p<0.01) were also observed in both the rate of phagocytosis and the number of cells phagocytized 3 hours after administration, when compared with the control group. CONCLUSION: This paper demonstrates that ingestion of Kakkon-to not only increases the body temperature but also enhances the phagocytic activity of macrophages, an in vivo defense mechanism, suggesting that Kakkon-to contributes to the suppression of multiplication of common cold viruses and influenza viruses, which consequently results in improvement of various symptoms during infection with common cold viruses. PMID- 15650475 TI - Use of alternative medicine among Hispanics. AB - OBJECTIVES: To determine the prevalence and patterns of use of alternative medicine among urban Hispanics. DESIGN: A cross-sectional survey. SETTINGS/LOCATION: Churches located in the northern part of Los Angeles and outpatients and inpatients in a county hospital in southern California. SUBJECTS: 179 Hispanic subjects. OUTCOME MEASURES: The following issues related to alternative medicine were evaluated: frequency and reasons for its use; perceived efficacy; safety; costs; sources of information; overall satisfaction; and subject-physician interaction with respect to alternative medicine. RESULTS: Sixty-three percent (63%) of respondents reported using or had used one or more types of alternative medicine. The majority of users were women (75%, 95% confidence interval [CI], 66%-83%). The most commonly reported alternative therapies were herbs (57%, 95% CI 47%-66%), prayer (43%, 95% CI 34%-53%), and dietary supplements (21%, 95% CI 14-30%). Compared to nonusers, users of alternative medicine had significantly lower yearly income, less proficiency in English language, and lower educational level. The main indications for using alternative medicine were pain (61.4%, 95% CI 51%-71%), lack of energy (38.6%, 95% CI 29%-49%), and overweight (22.8%, 95% CI 16%-33%). Only 5.2% of subjects (95% CI 2%-12%) believed that herbs are more effective than medications prescribed by physician. A majority of users (61%, 95% CI 50%-68%) had more confidence in their physicians than in their alternative medicine provider. Forty five percent (45%) of users (95% CI 34%-56%) believed prescribed medication was safer than alternative medicine, whereas only 7% (95% CI 3%-15%) believed the reverse. Adverse effects related to herbs and dietary supplements were reported by 11 users. The majority of participants (78.9%, 95% CI 69-87%) did not speak to their physicians about alternative therapy. The mean score of overall satisfaction with conventional medicine was significantly higher than with alternative medicine. CONCLUSIONS: Despite the common use of alternative medicine among Hispanics, the majority of users believe that conventional medicine is superior to alternative medicine. There is a significant gap in communication between physicians and patients with respect to alternative medicine. PMID- 15650476 TI - Trends in complementary and alternative medicine in eastern Turkey. AB - OBJECTIVE: To investigate the types and prevalence of complementary and alternative medicine (CAM) use among adults in eastern Turkey and to evaluate the relationship between the use of CAM and sociodemographic characteristics. DESIGN: This study was a random-sampled survey examining patterns of use of complementary therapies in two university hospitals in the Eastern Anatolia region of Turkey. SETTING AND SUBJECTS: Seven-hundred-fourteen (714) adult participants were interviewed at two sites: two university hospitals both located in Erzurum. Subjects were excluded if they did not speak Turkish because this may sometimes cause communication problems between patients and health care professionals. Patients younger than 18 years old and those residing outside of the eastern region of Turkey were not included in this study. MEASUREMENT: Use of CAM within the previous year. RESULTS: Seventy percent (70%) of participants reported the use of CAM. Herbs (41%) were the most frequently cited therapies. Although 87% of the subjects reported that they were satisfied with their use of CAM, the majority (84.8%) did not discuss the use of these therapies with their primary physicians. CONCLUSIONS: This study revealed that there is a high prevalence of use of alternative therapies by individuals living in eastern Turkey. PMID- 15650477 TI - A personal perspective on energies in future energy medicine. AB - This paper deals with the evolution of medicine from chemical medicine to energy medicine to information medicine as, first, standard electric, magnetic, and electromagnetic fields are applied to a patient and, second, as subtle energy fields are applied to the patient. Next, a brief reporting is given on our specific intention imprinting of a simple electrical device, which, when turned on in a given space, lifts the inner symmetry state of that space and tunes that space so that the specific intention, acting on the materials in that space, causes well-designed target experiments running in that space to yield results in full accord with the specific intention. A new theoretical model of nature is provided to explain these seemingly anomalous phenomena. A new perspective on what constitutes the whole person is also provided and one finds that human consciousness will become an important experimental variable in future medicine. PMID- 15650478 TI - A biostatistical approach to ayurveda: quantifying the tridosha. AB - OBJECTIVE: To compute quantitative estimates of the tridosha--the qualitative characterization that constitutes the core of diagnosis and treatment in Ayurveda -to provide a basis for biostatistical analysis of this ancient Indian science, which is a promising field of alternative medicine. SUBJECTS: The data sources were 280 persons from among the residents and visitors/training students at the Brahmvarchas Research Centre and Shantikuj, Hardwar, India. DESIGN/METHODOLOGY: A quantitative measure of the tridosha level (for vata, pitta, and kapha) is obtained by applying an algorithmic heuristic approach to the exhaustive list of qualitative features/factors that are commonly used by Ayurvedic doctors. A knowledge-based concept of worth coefficients and fuzzy multiattribute decision functions are used here for regression modeling. VALIDATION AND APPLICATIONS: Statistical validation on a large sample shows the accuracy of this study's estimates with statistical confidence level above 90%. The estimates are also suited for diagnostic and prognostic applications and systematic drug-response analysis of Ayurvedic (herbal and rasayanam) medicines. An application with regard to the former is elucidated, extensions of which might also be of use in investigating the role of nadis in Ayurvedic healing vis-a-vis acupuncture and acupressure techniques. The importance and scope of this novel approach are discussed. CONCLUSIONS: This pioneering study shows that the concept of tridosha has a sound empirical basis that could be used for the scientific establishment of Ayurveda in a new light. PMID- 15650479 TI - Complementary and alternative medicine in multiple sclerosis: survey of licensed naturopaths. AB - OBJECTIVE: This paper describes the treatments and treatment outcome measures used by licensed naturopathic physicians in the United States who treat people with multiple sclerosis (MS). DESIGN: A cross-sectional mail survey was used. SUBJECTS: The participants were licensed naturopaths who were members of the American Association of Naturopathic Physicians. OUTCOME MEASURES: Outcome measures included practitioner demographics; patient demographics by practitioner report; recommended therapies; perceived effectiveness of treatments for MS; methods for measuring treatment effectiveness. RESULTS: Forty three percent (43%) of the respondents (166/385) had treated at least one patient with MS while 56.9% (291/385) had never treated MS. 63.3% had treated 1-10 patients with MS, 19.9% had treated 11-20 patients with MS, and 16.8% had treated > or =20 patients with MS. Among the naturopaths, 68.1% communicated with an M.D. about their patient(s)' care and the majority of patients with MS were diagnosed by an M.D. (mean % = 96.3). The mean number of therapies recommended for M.S. was 3.91 (standard deviation [SD] =2.01, range 1-10). The most frequently recommended therapies included, diet (52.4%), essential fatty acid supplementation (44.6%), vitamin/mineral supplementation (33.7%), homeopathy (30.7%), botanicals (22.3%), and antioxidants (18.1%). Respondents perceived their treatments as "very effective" for the following stages of MS: early stage (57.2%); middle stage (25.3%); and late stage (3.0%). Respondents perceived their treatments as "very effective" for the following disease-related outcomes: improved quality of life (59.0%); decrease relapse rates (48.2%); decreased symptom severity (45.8%); prevention of disease progression (41.6%). The methods used "most often" for measuring treatment effectiveness included, patient report (88.0%); physical examination (27.1%); medical records/laboratory testing (13.3%). The mean estimated percentage of patients not taking conventional disease-modifying medication was 51.2% (SD = 42.7%). CONCLUSIONS: Naturopaths use both a broad range and multiple complementary and alternative medicine CAM therapies for treating MS and report treatment effectiveness on the following outcomes: quality of life; symptom severity; relapse rates; and disease progression. Further research on single CAM therapies and holistic CAM systems is warranted in MS. PMID- 15650480 TI - An approach to teaching dermatologists about complementary medicine. AB - OBJECTIVES: There is increasing evidence for the extensive use of complementary and alternative medicine (CAM) among patients with dermatologic ailments amid a scarcity of information about teaching health providers on how to engage with patients who use CAM or request information about it. An approach to educating dermatologists and nurses about CAM is described in this paper. DESIGN: A course on CAM and dermatology was offered to 17 dermatologists and nurses at a dermatology department in a university hospital in Israel. The course objective was to expose participants to common methods in CAM, emphasizing the role of an evidence-based approach and patient-doctor communication relating to CAM. RESULTS: Course evaluation revealed that participants acknowledged the existence of evidence-based research as an important consideration before referring their patients to CAM. CONCLUSIONS: Teaching CAM to dermatologists and nurses using evidence-based medicine and a patient-centered approach could enable them to provide informed and balanced advice to their patients. PMID- 15650481 TI - Defining minimum standards of practice for incorporating African traditional medicine into HIV/AIDS prevention, care, and support: a regional initiative in eastern and southern Africa. AB - In many resource-poor settings of Africa, a majority of people living with HIV/AIDS depend on and choose traditional healers for psychosocial counseling and health care. If the current pan-African prevention and care efforts spurred by the HIV pandemic do not actively engage African Traditional Medicine, they will effectively miss 80%, the vast majority of the African people who, according to the World Health Organization, rely on traditional medicine for their primary health care needs. In 2001, the Ugandan nongovernmental organization, Traditional and Modern Health Practitioners Together Against AIDS and Other Diseases, in Kampala, identified the need for a concerted, systematic, and sustained effort at both local and regional levels to support and validate African Traditional Medicine on several fronts. The Eastern & Southern Africa Regional Initiative on Traditional Medicine and AIDS was borne out of this assessment. It convened a regional consultation in May 2003, which produced a series of proposed standards around six main themes related to traditional medicine and HIV/AIDS: the systematic evaluation of traditional remedies; spiritual aspects of healing; HIV prevention and care; processing and packaging of traditional remedies; protection of indigenous knowledge; and intellectual property rights related to traditional health systems. These standards, summarized in this paper, will be incorporated into programs on traditional medicine and HIV/AIDS by various implementers in the region. A number of strategies to test and implement these recommendations are also defined. PMID- 15650484 TI - Special issue on bipolar disorder. PMID- 15650487 TI - Possible acute confusional state after risperidone in a pediatric patient. PMID- 15650488 TI - Occurrence of priapism with risperidone-paroxetine combination in an autistic child. PMID- 15650489 TI - Oxcarbazepine treatment of juvenile-onset bipolar disorder. PMID- 15650490 TI - Psychopharmacology news. PMID- 15650491 TI - Medicare-approved prescription drug discount card program: will this really help our seniors? PMID- 15650492 TI - A prospective study of hyperprolactinemia in children and adolescents treated with atypical antipsychotic agents. AB - OBJECTIVE: Few studies have prospectively compared the change of prolactin levels in children and adolescents associated with the use of atypical antipsychotic agents. In our study, we present preliminary data of an ongoing study, which compares changes in prolactin levels in children and adolescents after treatment to risperidone versus olanzapine versus quetiapine. We hypothesized: (1) risperidone would be associated with hyperprolactinemia most frequently, and (2) postpubertal females may be at higher risk of prolactin elevation and associated adverse effects. METHODS: Prolactin levels were obtained at baseline and after a mean of 11.2 weeks (SD = 2.2; range, 4-15 weeks) from 40 subjects (mean age, 13.4 years; SD = 3.4; range, 5-18 years) who were started on risperidone (n = 21), olanzapine (n = 13), or quetiapine (n = 6). End-point prolactin levels were compared using a Kruskal-Wallis test. RESULTS: End-point prolactin levels were significantly higher with risperidone, compared to olanzapine (p = 0.027) or quetiapine (p = 0.008). With the Bonferroni correction, the latter remains significant. Twenty-five percent of our subjects experienced sexual side effects at end point, independent of prolactin levels and antipsychotic agents. CONCLUSION: Risperidone significantly increased prolactin levels in children and adolescents. The duration of this prolactin elevation, and its long-term effects in children and adolescents, are unknown. PMID- 15650493 TI - The effect of atypical antipsychotic agents on prolactin levels in children and adolescents. AB - This report is a review of the available literature on the effect of atypical antipsychotic agents on prolactin in children and adolescents. Fourteen reports are reviewed. Most reports (79%) have included adolescents. Three reports (21%) consisted of children only, while 7 reports (50%) included only adolescents. A total of 4 reports (29%) included both children and adolescents. The total number of subjects listed in all the reports is 276, while only 49 of the individuals on atypical neuroleptics had prolactin elevations clearly identified as outside of the normal range. The details of the reports are provided by individual atypical antipsychotic agent. Clinical implications, such as the potential impact of hyperprolactinemia on bone density, osteoporosis, gynecomastia, galactorrhea, and weight gain, are presented. Discussion of pertinent medical differential and treatment options are also reported. PMID- 15650494 TI - Second-generation antipsychotic medications in children and adolescents. AB - OBJECTIVE: We reviewed available pediatric literature on second-generation antipsychotic medications to assess current evidence of efficacy and safety. METHOD: An English language MEDLINE search (1974-2003) was conducted using key words-atypical antipsychotics, children and adolescents, toxicity, clozapine, risperidone, olanzapine, quetiapine, ziprasidone, and aripiprazole. Additional efficacy and safety data were obtained from drug manufacturers. RESULTS: We identified 176 reports, including 15 double-blind, controlled trials, 58 openlabel studies, 18 retrospective chart reviews, and 85 case series/reports. The majority of these studies (43%) were of risperidone. Evidence suggests that second-generation antipsychotics are efficacious in the treatment of psychosis, bipolar disorders, pervasive developmental disorders, and Tourette's Disorder, and are potentially useful in mental retardation, conduct disorder, and severe attention deficit hyperactivity disorder (ADHD). The most frequently reported side effects included cardiovascular effects, weight gain, sedation, sialorrhea, extrapyramidal signs, and hyperprolactinemia, although the relative frequencies of these untoward effects vary among medications. CONCLUSION: Although the evidence base for pediatric use of second-generation antipsychotics is expanding, the majority of available studies are anecdotal, or short-term, openlabel trials. Reports suggest that these compounds are effective for a variety of psychiatric disorders in children and adolescents, but additional double-blind, controlled studies are required to establish definitive efficacy. Although these medications appear to be well tolerated in short-term studies, long-term follow-up investigations and ongoing clinical monitoring are necessary to confirm their safety in this age group. PMID- 15650495 TI - Predictors of treatment nonresponse in bipolar children and adolescents with manic or mixed episodes. AB - Even though juvenile bipolar disorder (BD) is reported to be more treatment resistant than adult BD, predictors of nonresponse are not well studied. The aim of this study was to address this issue in a naturalistic sample of bipolar children and adolescents with manic or mixed episodes treated under the condition of routine clinical practice. This study was comprised of 40 patients (19 females and 21 males; mean age, 14.2 years; SD = 3.3; range, 7-18) with a Diagnostic and Statistical Manual of Mental Disorders-fourth edition (DSM-IV) diagnosis of manic (n = 23) or mixed episodes (n = 17). The clinical characteristics of 20 patients considered to be treatment responders, according to the Clinical Global Impression-Improvement (CGI-I) scores, were compared to those of the 20 nonresponders. The effect of predictors on the probability of treatment nonresponse was analyzed using the multiple stepwise logistic regression, backward procedure. Demographic variables (mean age, gender ratio, socioeconomic status), as well as the inpatients-outpatients ratio (75% versus 65%), duration of the follow-up (10.5 +/- 2.5 months versus 9.6 +/- 3.2 months), index episode (manic versus mixed), and rates of pharmacologic hypomania did not differentiate the 2 groups. According to stepwise logistic regression, predictors of nonresponse were the presence of comorbidity with conduct disorder (odd ratio, 3.36; 95% CI, 2.20-4.52), attention deficit hyperactivity disorder (ADHD) (odd ratio, 2.30; CI, 1.24-3.26), and the baseline CGI Severity score (odd ratio, 2.31; CI, 1.33-3.29). It is relevant to point out that patient age at the onset of BD, and at the first visit, and comorbid anxiety disorders did not influence treatment response. Follow-up studies with a larger sample size with BD and/or externalizing disorders appropriately managed with different treatment options and/or combinations are warranted. PMID- 15650496 TI - Quetiapine adjunctive and monotherapy for pediatric bipolar disorder: a retrospective chart review. AB - OBJECTIVE: The aim of this study was to assess the safety and effectiveness of quetiapine for children and adolescents with bipolar disorder when used as monotherapy or in combination with other agents. METHODS: The outpatient medical charts of youths with a Diagnostic and Statistical Manual of Mental Disorders fourth edition (text revision) (DSM-IV-TR) diagnosis of bipolar disorder who were treated with quetiapine were retrospectively reviewed using the Clinical Global Impression (CGI) Scale. RESULTS: Thirty-two patients (mean age, 10.8 +/- 3.9 years) with bipolar disorder, type I (n = 16), type II (n = 10), cyclothymia (n = 2), and bipolar not otherwise specified (NOS) (n = 4) who had been treated (mean duration, 6.1 +/- 5.9 months) openly with quetiapine (mean dose, 397.4 +/- 221.4 mg/day) were identified. Fourteen patients (43.8%) received only quetiapine. No serious adverse events occurred. Response rate (defined by a CGI-Improvement (CGI I) score of < or =2 at end point) was 80.0% for the entire group and 78.6% for the subgroup who received quetiapine monotherapy. CGI-Severity (CGI-S) scores significantly improved from baseline to end point for both groups. CONCLUSIONS: Preliminary results suggest that quetiapine is a safe and effective treatment for both children and adolescents with bipolar disorder when used as monotherapy or in combination with other agents. PMID- 15650497 TI - Safety of subchronic treatment with fluoxetine for major depressive disorder in children and adolescents. AB - OBJECTIVE: The aim of this study was to assess the safety of subchronic fluoxetine treatment for major depressive disorder (MDD) in children and adolescents. METHODS: Patients received up to 19 weeks of treatment with fluoxetine, 10 mg-60 mg daily. Safety was evaluated through the reporting of concomitant medications, vital signs, routine laboratory testing, electrocardiograms (ECGs), and adverse event data. RESULTS: Ninety-six patients, aged 9-17 years, completed 19 weeks of treatment with fluoxetine (n = 49) or placebo (n = 47). There were statistically significant differences between the fluoxetine and placebo groups in mean change from baseline for alkaline phosphatase and total cholesterol levels (p < 0.001, and p < 0.014, respectively), but there were no statistically significant differences between the incidence of abnormal laboratory values for these 2 analytes. Fluoxetine treated patients gained statistically significantly less height (fluoxetine: 1.0 cm +/- 2.4; placebo: 2.1 cm +/- 2.6; p = 0.004) and weight (fluoxetine: 1.2 kg +/ 2.7; placebo: 2.3 kg +/- 2.6; p = 0.008) than placebo-treated patients during the 19 weeks of treatment. There was no difference in the rate of reported suicide-related events between fluoxetine and placebo. CONCLUSION: Fluoxetine was found to be safe and well tolerated in this study of children and adolescents with MDD. Clarification and determination of the clinical significance of the growth-rate reduction seen during fluoxetine treatment requires further investigation. During treatment with fluoxetine, the growth of child and adolescent patients should be monitored. PMID- 15650498 TI - Selegiline in comparison with methylphenidate in attention deficit hyperactivity disorder children and adolescents in a double-blind, randomized clinical trial. AB - OBJECTIVES: The aim of this study was to examine the selegiline treatment compared to methylphenidate (MPH) in children and adolescents with attention deficit hyperactivity disorder (ADHD). METHOD: Forty subjects, aged 6-15 years, boys and girls, who were diagnosed as having ADHD, using the criteria of the Diagnostic and Statistical Manual of Mental Disorders- Fourth Edition (DSM-IV), were randomly assigned to receive either selegiline or MPH for 60 days. Treatment outcomes were assessed using the Attention Deficit Hyperactivity Scale (ADHS) administered at baseline and on days 14, 28, 42, and 60 following the commencement of treatment. Side effects were also rated. RESULTS: There were no significant differences between sex, age, weight, and ethnicity of participants in the 2 groups. Both groups showed a significant improvement over the 60 days of treatment resulting from the teachers' and parents' ADHS scores across the treatment. CONCLUSION: Following the trial, MPH did not effect greater mean improvement as a result of the parents' or teachers' ADHS scores than selegiline. Thus, selegiline appears to be effective and well tolerated for ADHD in children and adolescents. PMID- 15650499 TI - A retrospective assessment of topiramate in children and adolescents with pervasive developmental disorders. AB - An open-label retrospective study was conducted to assess the effectiveness and tolerability of topiramate in children and adolescents with pervasive developmental disorders (PDD). Topiramate is a novel antiepileptic drug approved as an adjunctive treatment for seizure disorders. A retrospective chart review was conducted in an outpatient clinic specialized in treating individuals with developmental disabilities, to identify all children and adolescents with PDD who received topiramate. Patients were included if concomitant medications remained unchanged. Treatment response was assessed using the Global Improvement item of the Clinical Global Impressions scale (CGI-GI), based on a review of medical records and the Conners Parent Scale (CPS), as completed by parents. Fifteen patients were identified (12 male, 3 female; age = 14.7 +/- 3.3 years), including 11 patients with autistic disorder, 2 patients with Asperger's disorder, and 2 patients with PDD not otherwise specified (PDD, NOS). Eight patients (4 patients with autistic disorder, 2 patients with Asperger's disorder, and 2 patients with PDD, NOS) were judged to be responders, as defined by a score of 1 or 2 on the CGI-GI. Treatment duration was 25 +/- 16 weeks, and the mean dose was 235 +/- 88 mg. Differences between baseline and the end-of-trial period were observed in the following CPS subscales: conduct, hyperactivity, and inattention. No differences were noted in the psychosomatic, learning, and anxiety subscales. Three patients discontinued topiramate because of side effects, with 2 patients experiencing cognitive difficulties and 1 patient a skin rash. Topiramate may be beneficial for treating secondary symptoms of PDD, and prospective openlabel studies and double-blind, placebo-controlled studies to assess its efficacy and safety are needed. PMID- 15650500 TI - Psychotropic utilization and psychiatric presentation of hospitalized very young children. PMID- 15650501 TI - Atomoxetine for the treatment of pediatric nocturnal enuresis. AB - Current treatments for pediatric nocturnal enuresis rely upon the use of behavioral interventions, such as enuresis alarms, and pharmacological interventions, such as antidiuretic hormone analogs, tricyclic antidepressants, and anticholinergics. However, a considerable number of patients do not respond fully to any of these interventions, in spite of optimal behavioral management and medication dosing. This report describes 4 children with attention deficit hyperactivity disorder (ADHD) comorbid with nocturnal enuresis. Each child was treated with atomoxetine for ADHD and experienced serendipitous resolution of enuresis. PMID- 15650502 TI - Stimulant drug treatment in childhood-onset schizophrenia with comorbid ADHD: an open-label case series. AB - The administration of psychostimulants to children with psychotic symptoms is controversial. This study reports the stimulant drug response of 5 children, aged 8-15 years, with childhood-onset schizophrenia (COS) and comorbid attention deficit hyperactivity disorder (ADHD). Four COS inpatients were given stimulants for comorbid ADHD after stabilization of psychosis on antipsychotic medication. A fifth COS inpatient received stimulants while still actively psychotic, despite concurrent neuroleptic treatment. Data from the 10-item Brief Conners Teachers Ratings Scale (BCTRS) were examined the week before, and the week after, stimulant addition. A paired t test, conducted using Conners Teachers data from these 4 subjects, indicated significant improvement in ADHD symptoms (p = 0.02). Data obtained from a retrospective chart review indicated no significant worsening of psychosis. The 2 subjects treated with mixed salts of dextroamphetamine sulfate and amphetamine sulfate remained on that medication at 6 months and at the 2-year follow-up. Our results suggest that ADHD comorbid with COS may be safely treated with a stimulant, once the psychosis is stabilized. A systematic investigation of this question may be warranted. PMID- 15650503 TI - Aripiprazole for maladaptive behavior in pervasive developmental disorders. AB - OBJECTIVE: The aim of this study was to obtain preliminary data regarding the effectiveness and tolerability of aripiprazole in the treatment of children and adolescents with a pervasive developmental disorder (PDD). METHOD: Five youths (mean age, 12.2 years; range, 5-18 years) meeting Diagnostic and Statistical Manual of Mental Disorders-Fourth Edition (DSM-IV) criteria for a PDD received a naturalistic, open-label trial of aripiprazole (mean dosage, 12.0 mg/day; range, 10-15 mg/day) for a minimum of 8 weeks (mean duration, 12 weeks; range, 8-16 weeks). RESULTS: All 5 patients were judged to be responders, as determined by a Clinical Global Impressions-Improvement (CGI-I) scale rating of "much improved" or "very much improved." Aripiprazole was well tolerated. No extrapyramidal symptoms or clinically significant changes in heart rate or blood pressure occurred during the short-term trials. Two of 5 patients experienced mild somnolence. Two subjects lost weight, 2 subjects had no change, and 1 subject gained weight (mean change, -8.2 lbs; range, -30 to +1 lb). The weight loss was likely the result of the discontinuation of atypical antipsychotics that had led to significant weight gain. CONCLUSIONS: This case series describes the effectiveness of aripiprazole in the treatment of maladaptive behaviors in 5 patients with a PDD. No significant adverse effects emerged during these short term trials. Additional research is needed to support these preliminary findings. PMID- 15650504 TI - Intramuscular ziprasidone for acute agitation in adolescents. AB - Several neuropsychiatric disorders in children and adolescents often present with aggressive behavior. In fact, aggression is one of the most common reasons for psychiatric admission for inpatient hospitalization. Psychotropic medication can be helpful in reducing the need for more restrictive interventions, such as seclusion or restraint. The use of "as needed " (PRN) medications has been reported to decrease seclusion and restraint in a university-affiliated hospital setting. In our study, we report the cases of 3 youngsters whose escalating aggression responded to intramuscular ziprasidone with an immediate calming effect and good clinical outcome. PMID- 15650505 TI - Ziprasidone monotherapy in pediatric bipolar disorder. AB - Four children, aged 7-16 years, with bipolar disorder were switched to ziprasidone from mood stabilizers, anticonvulsants, or other atypical antipsychotics because of poor response, troubling side effects, breakthrough symptoms, or concern over potential toxicity. Within 3 days, patients experienced a resolution of hypomania, hallucinations, aggression, irritability, depression, and insomnia. One 16-year-old, who was switched from carbamazapine, also required adjunctive lorazepam for situational anxiety; the others either responded to, or were ultimately managed with, ziprasidone monotherapy. Side effects were mostly mild and transitory. Patients experiencing sedation or wakefulness at dose escalation were maintained at the previous 20- or 40-mg dose level until side effects resolved. Ziprasidone's efficacy, fast onset of action, and good safety profile warrant a more systematic study of this agent in pediatric patients with bipolar disorder. PMID- 15650506 TI - Pediatric bupropion-induced serum sicknesslike reaction. AB - This reports the first 2 cases of serum sicknesslike reaction to bupropion in children (age 12 and 14). Serum sicknesslike reactions are an example of immune complex medicated disease. The cardinal symptoms of serum sickness are fever, lymphadenopathy, arthralgias or arthritis, and urticaria. Symptoms usually resolve without long-term sequela following discontinuation of the exogenous antigen. It is likely that serum sicknesslike reactions to bupropion are either relatively rare or underrecognized and underreported. Between May 1998 and May 2001, GlaxoSmith Kline received 172 reports of seizures (a well-known adverse drug reaction) and only 37 reports of serum sicknesslike reactions (Wooltorton 2002). We do not know if children and adolescents are more prone than adults to develop serum sicknesslike reactions to bupropion. Luckily, the reported cases of serum sicknesslike reactions to bupropion have not caused irreversible morbidity or mortality. Nevertheless, the symptoms are painful, temporarily disfiguring and disabling, and warrant prompt medical attention. Parents and patients should be educated about this potential side effect at the onset of treatment, because symptoms are similar to many infectious childhood illnesses, and the treatment of serum sicknesslike reactions to bupropion should include the discontinuation of bupropion. PMID- 15650507 TI - Neuroleptic malignant syndrome in an adolescent after brief exposure to olanzapine. AB - A 17-year-old African-American male developed neuroleptic malignant syndrome (NMS) with hyperthermia, autonomic instability, increased muscle tone, rhabdomyolysis, and obtundation after a maximum of 2 days of treatment with olanzapine and 1 day of treatment with divalproex sodium. Intensive care unit (ICU)-level care was required. Paranoid psychosis with catatonia was present after recovery from the NMS. Because of his continued psychotic symptoms following resolution of the NMS, the alternate atypical antipsychotic, clozapine, was started under close observation. Reports of NMS resulting from atypical antipsychotic agents are generally uncommon, and much more so in the child and adolescent population. However, these agents are frequently prescribed in this population and require due caution. PMID- 15650511 TI - Journal of ocular pharmacology and therapeutics: history and future directions. PMID- 15650512 TI - Intraocular pressure and safety in glaucoma patients switching to latanoprost/timolol maleate fixed combination from mono- and adjunctive therapies. AB - PURPOSE: The aim of this study was to evaluate the efficacy and safety in ocular hypertensive or open-angle glaucoma patients changed to a latanoprost/timolol fixed combination (LTFC) from previous therapy. METHODS: We employed a retrospective, multicenter, active-controlled study evaluating patients who had a previous therapy substituted with LTFC and had a 3-month follow-up. RESULTS: In the 168 patients included in the trial, LTFC reduced the intraocular pressure (IOP) after switching from previous therapies: timolol (22.8+/-3.5 to 19.0+/-3.9 mmHg, N=49, p<0.001), latanoprost (21.2+/-3.8 to 18.3+/-2.5 mmHg, N=54, p<0.001), and a dorzolamide/timolol fixed combination (20.9+/-2.4 to 20.0+/-2.7 mmHg, N=32, p=0.03). In switching from a latanoprost and timolol unfixed combination, the pressure changed from 18.3+/-3.8 to 18.9+/-3.0 mmHg (N=33, p=0.38). LTFC was persistent in 131 (78%) patients within the 3-month treatment period. The most common adverse event with LTFC was conjunctival hyperemia, which occurred in patients not previously treated with latanoprost therapy (N=16, 10%). CONCLUSIONS: LTFC generally provides reduced IOP and limited side effects when substituted for other common glaucoma therapies, while providing similar pressure when switched from its own individual components. PMID- 15650513 TI - A persistency and economic analysis of latanoprost, bimatoprost, or beta-blockers in patients with open-angle glaucoma or ocular hypertension. AB - PURPOSE: The aim of this study was to evaluate differences in the persistency and treatment costs for latanoprost, bimatoprost, or beta-blockers in open-angle glaucoma or ocular hypertensive patients. METHODS: This study was a retrospective, multicenter, parallel, active-controlled comparison of patients who were prescribed with ocular hypotensive monotherapy between September 1996 and August 2002. RESULTS: 1,182 patients were included. The Kaplan Meier life table analysis showed that latanoprost was continued longest among the groups for the first year of therapy (p=0.02). A significant difference existed between groups in the final intraocular pressure for latanoprost (17.3+/-3.9, N=357), for bimatoprost (18.0+/-3.6, N=146), and for the beta-blockers (17.9+/-3.7, N=335) (p=<0.0001). The average number of visits was statistically higher for beta blockers (3.3), compared to latanoprost (2.9) and bimatoprost (3.1) (p=0.01). Further, the mean number of medicine changes was greater for bimatoprost (0.45) and beta-blockers (0.47) than for latanoprost (0.27) (p=0.0008). The cost of visits and medications was lowest for beta-blockers ($119.3+/-$78.9) and highest for bimatoprost ($163.8+/-$51.2) (p<0.0001). CONCLUSIONS: Patients were more persistent with latanoprost and demonstrated lower intraocular pressure, fewer visits, and fewer medicine changes when compared to bimatoprost or beta-blocker therapy. In contrast, the beta-blocker group provided lower overall cost. PMID- 15650514 TI - Short-term effects of bimatoprost in glaucoma patients from an outpatient clinic. AB - PURPOSE: A drug utilization trial was performed to investigate acute versus short term effects after switching or adding bimatoprost in open-angle glaucoma patients over a 3- month observation period. METHODS: One (1) eye was randomly chosen from 47 glaucomatous patients (abnormal visual field and/or abnormal optic nerve and intraocular pressure (IOP) above 21 mmHg without treatment). Only patients who did not reach the target IOP with their ongoing treatment were recruited in this study. IOP was measured at baseline, after 1 hour, and 2 hours from the first instillation and after 1 week, 1, and 3 months of treatment. RESULTS: The IOP before bimatoprost administration was 20.16+/-3.6 mmHg (mean+/ standard deviation). There was no statistically significant decrease of IOP after 1 hour (mean IOP, 19.96+/-4.25 mmHg) and after 2 hours (mean IOP, 17.73+/-3.24 mmHg). Statistically significant (p<0.001) decreases after 1 week (mean IOP, 16.48+/-2.9 mmHg), after 1 month (mean IOP, 16.48+/-2.9 mmHg) and after 3 months (mean IOP, 16.15+/-2.7 mmHg) were found. CONCLUSION: The results suggested that bimatroprost had a significant acute effect on IOP in monotherapy, while no significant effect was found when the therapy was switched or added. The effect for primary open-angle glaucoma was very evident. There was no specific side effect attributable to combining bimatoprost with any of the treatments in use. PMID- 15650515 TI - Intraocular pressure fluctuations in response to the water-drinking provocative test in patients using latanoprost versus unoprostone. AB - Impairment of outflow facility in glaucoma causes large intraocular pressure (IOP) fluctuations that have been shown to be a risk factor for disease progression. The water-drinking provocative test (WDT) has been proposed as an indirect measurement of outflow facility to compare intraocular pressure responses of glaucoma eyes to different drugs. This study was a double-masked, randomized, parallel-group clinical trial comparing the IOP fluctuations in response to the WDT in patients using latanoprost versus unoprostone. After completing a wash-out of ocular hypotensive medications, patients with primary openangle glaucoma or ocular hypertension were randomized to receive either latanoprost (N=40) or unoprostone (N=42). IOP was measured before treatment and at 8 weeks after treatment (baseline IOP for WDT), followed by the WDT. IOP fluctuations and maximum IOP after water ingestion were compared between the two groups. Analysis of covariance was used to adjust for the effects of baseline IOP and treatment efficacy. The mean percentage reduction of IOP was 27% in patients using latanoprost, as compared to 13% in patients using unoprostone (p<0.001). Patients on treatment with latanoprost had significantly less IOP fluctuations in response to the WDT, compared to patients using unoprostone. From an overall baseline IOP of 20.0 mmHg and an overall treatment efficacy of 20%, the mean+/ standard error of the mean (SEM) of the IOP fluctuation during the WDT was 5.3+/ 0.4 mmHg in the unoprostone group, and 3.6+/-0.4 mmHg in the latanoprost group (p=0.005, ANCOVA). This could represent an additional benefit of latanoprost over unoprostone in controlling the intraocular pressure of glaucomatous patients. PMID- 15650516 TI - Effect of different dose schedules of 0.15% unoprostone isopropyl on intraocular pressure and pupil size in the glaucomatous beagle. AB - The changes in intraocular pressure (IOP) and pupil size (PS) after instillations of 0.15% unoprostone isopropyl (Rescula, Novartis Ophthalmics, Duluth, GA) were investigated in the spontaneous glaucoma Beagle model. From the first-day baseline IOP of 27.3+/-3.2 mmHg placebo eye and 32.8+/-5.1 mmHg control eye, the mean+/-standard error of the mean (SEM) diurnal changes after 0.15% unoprostone, at 8 AM once-daily for the next 4 days, were 15.5+/-1.3 mmHg, 14.7+/-1.9 mmHg, 16.1+/-1.1 mmHg, and 17.0+/-1.5 mmHg, respectively, and were significantly different from the control eye. After 0.15% unoprostone was instilled at 8 PM, the mean+/-SEM baseline changes from the baseline IOP (insert drug eye 9 AM) in the drug eyes were 5.9+/-2.5 mmHg, 5.2+/-4.1 mmHg, 9.7+/-2.5 mmHg, and 3.6+/-3.6 mmHg, respectively. When 0.15% unoprostone was instilled twice-daily, the mean+/ SEM baseline IOP (insert drug eye 9 AM) changes were 13.6+/-0.7 mmHg, 13.9+/-1.4 mmHg, 11.3+/-1.0 mmHg, and 9.3+/-1.4 mmHg, respectively, and were significantly different from the control eyes. Miosis occurred within 2 hours and lasted several hours. Unoprostone isopropyl instilled once-daily (AM or PM), as well as twice-daily, produces significant decreases in IOP and PS in the glaucomatous Beagle. PMID- 15650517 TI - Diffusion of metronidazole released from aqueous solution and a gel through human and rabbit corneas. AB - Metronidazole has been suggested as an adjunct in the treatment of Acanthamoeba keratitis. This study involves the permeation of metronidazole from topically applied solutions and gels through human and rabbit corneas. The permeation of metronidazole, from the solutions and gels through the corneas, was determined using a flow-through diffusion apparatus. Human (32) and rabbit (54) corneas were obtained from an eye bank and animal nursery farm, respectively. Experiments were conducted at 20 degrees C and over a time period of 24 hours. High-performance liquid chromatography (HPLC) analysis was used as a detection method. Statistical tests used included an Analysis of Variance (ANOVA) and Duncan's multiple range test, for a determination of the steady state, and a nonlinear regression analysis, using an F test, was used to compare entire curves. Flux rates for 0.5% and 1% metronidazole solutions across human corneas were found to be 14% and 30% higher, respectively, than for rabbit corneas. However, the flux rates of the 0.5% and 0.8% metronidazole gels across rabbit corneas were found to be 7% and 25% higher than those for human corneas. The flux rates measured for at least the first 10 hours were not, statistically, significantly different between the metronidazole released from aqueous solutions and from the gel. Steady-state flux rates of the 0.5% metronidazole aqueous and gel formulations were approximately half those of the 0.8% and 1% preparations. Because of the longer corneal contact times, it may be therapeutically advantageous to use 0.8% metronidazole concentrations in gels for treating Acanthamoeba keratitis infestations. Further in vivo studies on the ocular compatibility of these formulations need to be performed. The use of rabbit corneas as an in vitro permeability model for human corneas is supported by these studies. PMID- 15650518 TI - Vitrectomy with fluconazole infusion: retinal toxicity, pharmacokinetics, and efficacy in the treatment of experimental candidal endophthalmitis. AB - The aim of this study was to determine the retinal toxicity and intraocular pharmacokinetics of vitrectomy with fluconazole infusion in rabbit eyes and to study its efficacy in the treatment of experimental candidal endophthalmitis. The right eyes of 13 New Zealand White (NZW) rabbits were vitrectomized and infused with 20 mL of 0.2 mg/mL, 1 mg/mL, or 2 mg/mL of fluconazole. An electroretinogram (ERG) was performed on both eyes of each rabbit at different time points. The right eyes of 26 different NZW rabbits were vitrectomized and infused with 20 mL of 2 mg/mL of fluconazole. These rabbits were sacrificed, and their right eyes were enucleated at hours 2, 4, 8, and 24 after the operation, and the concentration of fluconazole in the vitreous was measured by highpressure liquid chromatography. Experimental candidal endophthalmitis was induced in the right eye of 42 other NZW rabbits. Twenty-six (26) of the eyes were then vitrectomized and infused with 20 mL of 2 mg/mL of fluconazole, and the other 16 rabbits served as control. Severity of ocular infection was graded from 0-4 at different time intervals, using an indirect ophthalmoscope. In the first group, ERG showed no significant difference between the experimental eyes and the control eyes--in all concentrations of fluconazole--for up to 3 months. In the second group, the intraocular concentration of fluconazole declined so rapidly that, as of 8 hours after operation, there was none in the vitreous cavity. In the third group, significantly less vitreous opacity was found in the treated eyes on days 3 and 6. However, the difference ceased to be apparent on day 15. Our study suggests that there is no retinal toxicity resulting from vitrectomy with a 2 mg/mL fluconazole infusion, and that it is temporally effective in the treatment of experimental candidal endophthalmitis. PMID- 15650519 TI - Intraocular properties of urokinase-derived antiangiogenic A6 peptide in rabbits. AB - To investigate the intraocular properties of an antiangiogenic peptide, A6, a total of 70 New Zealand rabbit eyes were used. For the toxicity study, 0.05 mL of 0.459 M or 0.148 M A6 was injected intravitreally; right eyes received A6, and left eyes received a vehicle. Serial intraocular pressure measurement, slit lamp, and indirect ophthalmoscopy were performed. The rabbit eyes were evaluated by fluorescein angiography, electroretinography, and histology after the scheduled sacrifice. The pharmacokinetics of an intravitreal A6 (0.05 mL of 0.488 M) and a subtenon A6 (0.5 mL of 0.305 M) injection was studied. There was no toxicity observed following the 0.148 M A6 intravitreal injections. In 2 eyes with a 0.459 M A6 intravitreal injection, focal retinal pigmentary change was observed at the injection site, which was contacted by the hyperosmolar drug bolus. Choroidal A6, following the intravitreal injection, remained therapeutic (>or=10 microM) for 72 hours. The vitreous half-life was 19.4 hours. Choroidal concentrations following the subtenon injection were minimal. The low choroidal concentrations observed may relate to the polar nature of A6. More hydrophobic analogs of A6 are likely to cross the retina more efficiently. However, in diseased eyes, in the area of choroidal neovascularization (CNV), the fluid-filled, damaged, edematous retina may permit the drug to enter the choroid in higher concentrations. PMID- 15650520 TI - Retinal function and histologic changes following intravitreal injection of indocyanine green in a rabbit model. AB - The aim of this animal study was to investigate the effects of intravitreal injection of indocyanine green (ICG) applied in macular hole surgery on retinal functional, morphology, and histologic changes. Eighteen (18) New Zealand albino rabbits were divided equally into three groups (6 rabbits in each). In Group A, both eyes of each rabbit were vitrectomized by perflouropropane gas compression. One (1) month later, 0.1 cc of different doses of ICG was injected into the vitreous in the left eyes. In the right eyes, 0.1 cc of balanced salt solution was injected intravitreally, allowing them to serve as control eyes. In Group B, the same doses of ICG were injected intravitreally. ICG was washed out by fluid fluid exchange 3 minutes after injection. In Group C, the same doses of ICG were injected intravitreally in nonvitectomized eyes. Scotopic and photopic electroretinogram (ERG) recordings and indirect ophthalmoscopy examinations were performed to detect any functional and morphologic changes. Rabbit eyes were enucleated 4 months after ICG injections to observe histologic changes. Significant decreased of scotopic and photopic ERG amplitude and marked histologic changes were noted in eyes injected with 0.5 mg/cc and 0.1 mg/cc of ICG in nonvitrectomized eyes (Group C). In vitrectomized eyes (Group A), decreased scotopic and photopic ERGs and mild histologic changes were noted in eyes injected with 0.5 mg/cc, but no histologic changes were noted in eyes injected with 0.1 mg/cc. There was a transient, mild decrease in scotopic and photopic ERGs and no morphologic changes were noted in the eyes with fluid-fluid exchange (Group B). The toxicity of intravitreous ICG is dose- and time dependent. ICG at 0.5 mg/cc, with short exposure time, is recommended in macular hole surgery. PMID- 15650522 TI - Telepsychiatry with children and adolescents: are patients comparable to those evaluated in usual outpatient care? AB - Several studies have described successful applications of telepsychiatry with children and adolescents. However, there has been little examination of the populations served by telepsychiatry and the ability to evaluate youth accurately through this medium. In this article, we examined whether telepsychiatry patients are representative of those in usual outpatient care. Participants included 369 patients 3-19 years old evaluated at two clinics. A new telepsychiatry clinic (TPC) developed to provide services to under-served communities, and a child and adolescent psychiatric outpatient clinic (CAPOC) that served youth from predominantly metropolitan areas were included in the study. The telepsychiatry sites were linked using ISDN lines at 384 KB/sec. We examined these two samples regarding demographics, payor status, and diagnostic profiles. Results indicated that youth evaluated through the TPC were broadly comparable to youth evaluated in the CAPOC. Therefore, telepsychiatry appears to serve youth that are representative of those seeking psychiatric care, and it is not restricted to youth with no medical insurance or with selected diagnoses. The similarity of diagnoses further suggests that telepsychiatry provides adequate technical resolution and doctor-patient rapport to detect psychopathology of youths. These findings suggest the need for further systematic investigation of telepsychiatry as a tool for providing psychiatric care to young people. PMID- 15650523 TI - Willingness to use telemedicine for psychiatric care. AB - The aim of this study was to assess potential users' willingness to use telepsychiatry and its correlates within a theoretical framework guided by the principles of social-cognitive models. The conceptual model proposed willingness to use telepsychiatry to be associated directly to attitudes toward telemedicine, attitudes toward the patient-physician relationship, and satisfaction with current medical care, and indirectly to anxiety to technology. Phone interviews were conducted with a representative sample of 1,204 Jewish, noninstitutionalized adults in Israel (mean age = 58; 51% female). Structural equation modeling was used to examine hypothesized relationships. Willingness to use telepsychiatry was relatively high and was affected by the participants' attitudes toward telemedicine, the patient-physician relationship, and by the level of anxiety toward technology. Efforts aimed at encouraging the use of telemedicine for psychiatric care should target potential users' attitudes, as well as feelings of uneasiness and anxiety regarding technology. PMID- 15650524 TI - Is JPEG compression of videomicroscopic images compatible with telediagnosis? Comparison between diagnostic performance and pattern recognition on uncompressed TIFF images and JPEG compressed ones. AB - Early melanoma diagnosis is an important goal for dermatologists. Polarized light systems are increasingly employed for dermatoscopic diagnosis of melanocytic lesions. For the purpose of teledermoscopy, whose importance is increasingly growing for consultation and teaching purposes, it is of utmost importance to establish whether, after compression, polarized light images maintain their informativeness. The aim of our study was to check the effects of compression on melanocytic lesion images acquired by means of a digital videomicroscope on the identification of morphological details of the image and on diagnostic accuracy. A total of 170 50-fold-magnified pigmented skin lesion images, acquired in Tagged Image File Format (TIFF) by a digital videomicroscope, were compressed using Joint Photographic Experts Group (JPEG) algorithms (compression factor 30). Two experts in videomicroscopy evaluated both original and compressed images twice by describing single lesion features and expressing a diagnosis. Reproducibility in the assessment of dermoscopic parameters and observer performance were studied by kappa statistics and Receiver Operating Characteristic (ROC) analysis. Both intra and interobserver reproducibility in the assessment of morphological details were higher when TIFF images were considered, indicating a better image quality. Nonetheless, there was no significant difference in the diagnostic accuracy between uncompressed images and compressed ones, although the intraobserver reproducibility in the diagnostic judgement was higher for uncompressed images. Despite loss in image details, factor 30 compressed videomicroscopic images enable a good diagnostic accuracy. PMID- 15650525 TI - Use of the internet for health information by African-Americans with modifiable risk factors for cardiovascular disease. AB - The aim of this study was to evaluate the frequency of Internet use among African Americans at risk of cardiovascular disease and to determine the feasibility of developing Webbased, culturally relevant health information programs to reduce cardiovascular disease in the African American faith community. A cross-sectional survey was used to investigate the association of Internet use with the risk of cardiovascular disease (CVD) among members of African Methodist Episcopal churches in South Carolina. A telephone survey including questions on demographic factors, medical history, diet, and exercise was administered to a sample of 208 adult church members. A preventable CVD risk factor was defined as having any of the following: hypertension, obesity, diabetes, high cholesterol, cigarette use, and not participating in moderate physical activity. A logistic regression determined the relationship of having a preventable CVD risk factor with Internet use while controlling for potential confounding variables. Among respondents, 47% used the Internet with most (65%) using it for 1 month or less. Most (87%) have at least one preventable CVD risk factor. Both higher education and younger age were significantly associated with Internet use. Results of the logistic regression indicate that individuals with a cardiovascular risk factor were not significantly less likely to use the Internet when controlling for age and education. Use of the Internet has potential to reach large populations, however, more work needs to be done to increase access to the Internet by older African Americans at greatest risk of CVD before this approach can be effective. PMID- 15650526 TI - Organizing safety: conditions for successful information assurance programs. AB - Organizations must continuously seek safety. When considering computerized health information systems, "safety" includes protecting the integrity, confidentiality, and availability of information assets such as patient information, key components of the technical information system, and critical personnel. "High Reliability Theory" (HRT) argues that organizations with strong leadership support, continuous training, redundant safety mechanisms, and "cultures of high reliability" can deploy and safely manage complex, risky technologies such as nuclear weapons systems or computerized health information systems. In preparation for the Health Insurance Portability and Accountability Act (HIPAA) of 1996, the Office of the Assistant Secretary of Defense (Health Affairs), the Offices of the Surgeons General of the United States Army, Navy and Air Force, and the Telemedicine and Advanced Technology Research Center (TATRC), US Army Medical Research and Materiel Command sponsored organizational, doctrinal, and technical projects that individually and collectively promote conditions for a "culture of information assurance." These efforts include sponsoring the "P3 Working Group" (P3WG), an interdisciplinary, tri-service taskforce that reviewed all relevant Department of Defense (DoD), Miliary Health System (MHS), Army, Navy and Air Force policies for compliance with the HIPAA medical privacy and data security regulations; supporting development, training, and deployment of OCTAVE(sm), a self-directed information security risk assessment process; and sponsoring development of the Risk Information Management Resource (RIMR), a Web enabled enterprise portal about health information assurance. PMID- 15650527 TI - Telemedicine information analysis center. AB - Congress mandated a pilot project to demonstrate the feasibility of establishing a Department of Defense (DoD) telemedicine information analysis center (TIAC). The project developed a medical information support system to show the core capabilities of a TIAC. The productivity and effectiveness of telemedicine researchers and clinical practitioners can be enhanced by the existence of an information analysis center (IACs) devoted to the collection, analysis, synthesis, and dissemination of worldwide scientific and technical information related to the field of telemedicine. The work conducted under the TIAC pilot project establishes the basic IAC functions and assesses the utility of the TIAC to the military medical departments. The pilot project capabilities are Web-based and include: (1) applying the science of classification (taxonomy) to telemedicine to identify key words; (2) creating a relational database of this taxonomy to a bibliographic database using these key words; (3) developing and disseminating information via a public TIAC Web site; (4) performing a specific baseline technical area task for the U.S. Army Medical Command; and (5) providing analyses by subject matter experts. PMID- 15650528 TI - Shadow Bowl 2003: a collaborative exercise in community readiness, agency cooperation, and medical response. AB - This paper describes a model for homeland security, community readiness, and medical response that was applied during an operational exercise around Super Bowl XXXVII. In addition, it describes the products provided by private companies involved in the exercise and how they would have contributed to a medical disaster had one occurred. The purpose of Shadow Bowl was to demonstrate community readiness and medical response to a mass casualty event. The goals of the project were to: (1) provide enhanced public safety using an advanced communication network and sensor grid; (2) develop mass casualty surge capabilities through medical reach-back; and (3) build a collaboration model between civilian, military, public, and private partners. The results of the Shadow Bowl Exercise accentuated the value of new telehealth and disaster medicine tools in treating large numbers of patients when infrastructure overload occurs. PMID- 15650529 TI - Navy telemedicine: a review of current and emerging research models. AB - With extended deployments of military telemedicine to remote, austere, and high risk settings, there is a danger that implementation will outrun the research designed to assess it. A review of existing research evaluates current assessment models and indicates gaps around which future research may be designed. A review of models for evaluating telemedicine was conducted in September, 2001. Seven areas of assessment recurred that could be subsumed under the framework of "interoperability." A follow-up search was conducted during September, 2003, to ascertain the degree to which assessments of interoperability were being advocated. Although extensive deployments of telemedicine systems in high-risk settings make it imperative to know that human, organizational, and technical components are integrating productively (i.e., interoperating), current research does not evaluate telemedicine as a system. The structure and functioning of telemedicine systems should be assessed empirically and systemically to avoid subjective or limited assessments. PMID- 15650530 TI - Telepediatrics in Canada: an overview. AB - Telehealth in Canada is undergoing phenomenal growth owing to recent federal and provincial funding to implement the use of information and communication technologies to improve access to health care for all Canadians. This funding has been a significant factor in establishing telehealth networks in the 10 provinces and 3 territories, thus increasing access to specialty and subspecialty care from urban tertiary care centers. Every province and territory in Canada is now part of a functioning telehealth network. For tertiary pediatric centers, this offers new opportunities for providing support and outreach services to children in remote and northern communities. This article provides a broad overview of development of telepediatrics in Canada. PMID- 15650531 TI - Behavioral health care of isolated military personnel by videoconference. AB - The changing role of the military presents a unique challenge for military behavioral health organizations in dealing with the geographic isolation of personnel. Telemedical solutions should help to meet this challenge. In the behavioral health arena, there are many uses for videoconferencing and other communications technology in maintaining continuity when patients or clinicians travel or deploy; assisting isolated behavioral health-care providers and general practitioners; evaluating and treating isolated military personnel; and addressing family issues. Family interventions by videoconference and other communications technology may be particularly useful in treating military personnel and in promoting morale of the fighting force. Although much remains to be researched and proven empirically, anecdotal experience supports the utility of using communications technology to enhance behavioral health interventions. This article describes the clinical videoconference experience within the Department of Psychiatry at Tripler Army Medical Center (TAMC), during a Multinational force deployment to the Sinai, and during training at the Joint Readiness Training Center, Fort Polk, Louisiana. A brief discussion of potential legal issues involving videoconferencing in behavioral health is also included. PMID- 15650533 TI - Expense comparison of a telemedicine practice versus a traditional clinical practice. AB - This paper compares the expenses of a telemedicine program to those of a traditional clinical practice using data from two fiscal years (FY) 1998/1999 and 2000/2001. As part of that evaluation, we compared expenses of the University of Arizona's clinical practice group, the University Physicians Incorporated (UPI), to those of the Arizona Telemedicine Program (ATP) practice. For this study, we used the reporting categories published in the year-end UPI financial statement. These categories included clinical services, administration, equipment depreciation, and overhead. Results showed that clinical service expenses and administrative expenses for FY 2000/2001 were higher in the traditional UPI practice, whereas equipment depreciation and overhead expenses are higher in the telemedicine practice. This differs somewhat from FY 1998/1999, where clinical expenses and overhead were higher in the UPI practice and administration and equipment depreciation were higher in the telemedicine practice. We will discuss the relevance of these results and the critical factors that contribute to these differences. PMID- 15650534 TI - Clinical encounters costing for telemedicine services. AB - The Arizona Telemedicine Program (ATP) has developed a costing model that includes a number of factors contributing to the cost of providing services via telemedicine and generates a per-encounter cost. We first developed the model in 1999 and in this paper used it to analyze clinical services for the fiscal year 2000/2001. We evaluated the calculated weighted cost for providing those services via telemedicine as compared to the cost of providing those services "face-to face." The cost of providing services via telemedicine was found to be lower when a patient had to travel 127.5 miles or more to receive clinical services. Our study provides analysis of the factors, their contribution to costs, as well as some factors, which are pertinent, but are more problematic to calculate. In this paper, we discuss the relevance of these results as they relate to telemedicine programs and a possible means to reduce costs even further. PMID- 15650535 TI - Use of American Sign Language in telepsychiatry consultation. AB - The aim of this study was to provide a psychiatric consultation to a deaf patient in a rural community via telemedicine. The patient successfully communicated with the telepsychiatrist via an American Sign Language (ASL) interpreter and her psychiatric symptoms improved. This case study demonstrated the feasibility of delivering specialty medical services to a special needs population via telemedicine. PMID- 15650537 TI - Continuing education via telemedicine: analysis of reasons for attending or not attending. AB - This study analyzed factors that influence the decision to attend or not attend tele-education broadcasts via a telemedicine network for continuing education (CE) credits. A questionnaire was developed and sent to all participating sites on our telemedicine network that have attended CE broadcasts. The majority of respondents attend specific types of broadcasts, receiving less than 25% of their CE credits from the tele-education broadcasts. Factors most influencing the decision to attend a broadcast include interest in the topic and perceived utility of the information. The most common reason influencing the decision not to attend is an explicit preference for attending conferences. Tele-education broadcasts may supplement other CE activities but may not replace them. The attraction of attending conferences in person is very high and may be especially so for health-care professionals in rural environments. PMID- 15650536 TI - Effect of camera resolution and bandwidth on facial affect recognition. AB - This preliminary study explored the effect of camera resolution and bandwidth on facial affect recognition, an important process and clinical variable in mental health service delivery. Sixty medical students and mental health-care professionals were recruited and randomized to four different combinations of commonly used teleconferencing camera resolutions and bandwidths: (1) one chip charged coupling device (CCD) camera, commonly used for VHSgrade taping and in teleconferencing systems costing less than $4,000 with a resolution of 280 lines, and 128 kilobytes per second bandwidth (kbps); (2) VHS and 768 kbps; (3) three chip CCD camera, commonly used for Betacam (Beta) grade taping and in teleconferencing systems costing more than $4,000 with a resolution of 480 lines, and 128 kbps; and (4) Betacam and 768 kbps. The subjects were asked to identify four facial affects dynamically presented on videotape by an actor and actress presented via a video monitor at 30 frames per second. Two-way analysis of variance (ANOVA) revealed a significant interaction effect for camera resolution and bandwidth (p = 0.02) and a significant main effect for camera resolution (p = 0.006), but no main effect for bandwidth was detected. Post hoc testing of interaction means, using the Tukey Honestly Significant Difference (HSD) test and the critical difference (CD) at the 0.05 alpha level = 1.71, revealed subjects in the VHS/768 kbps (M = 7.133) and VHS/128 kbps (M = 6.533) were significantly better at recognizing the displayed facial affects than those in the Betacam/768 kbps (M = 4.733) or Betacam/128 kbps (M = 6.333) conditions. Camera resolution and bandwidth combinations differ in their capacity to influence facial affect recognition. For service providers, this study's results support the use of VHS cameras with either 768 kbps or 128 kbps bandwidths for facial affect recognition compared to Betacam cameras. The authors argue that the results of this study are a consequence of the VHS camera resolution/bandwidth combinations' ability to improve signal detection (i.e., facial affect recognition) by subjects in comparison to Betacam camera resolution/bandwidth combinations. PMID- 15650539 TI - The platelet-related effects of tenecteplase versus alteplase versus reteplase. AB - Clinical studies have investigated the combination of glycoprotein (GP) IIb/IIIa inhibitors and thrombolytic agents for acute myocardial infarction. However, thrombolytic agents alone may possess direct antiplatelet properties that could affect reperfusion. Blood from 11 patients with coronary disease and five healthy subjects was incubated for 30 min with tenecteplase (4, 12, and 24 microg/ml), alteplase (1, 4, and 10 microg/ml), reteplase (1, 5, and 10 microg/ml) or control buffer. Platelet aggregation induced by 1, 20 and 50 micromol/l adenosine diphosphate (ADP), the stimulated expression of GP IIb/IIIa and P-selectin, and plasma fibrinogen levels were determined. Platelet aggregation in patients was inhibited by medium and high concentrations of alteplase when induced by 1 micromol/l ADP [1.6 +/- 0.5%, P = 0.001 and 0.9 +/- 0.2%, P = 0.002 versus 8.3 +/ 1.6% (control)] and 20 micromol/l ADP [46.9 +/- 3.9%, P = 0.001 and 46.2 +/- 4.8%, P = 0.001 versus 65.7 +/- 2.7% (control)]. High concentration tenecteplase was associated with lower aggregation by 20 micromol/l ADP (58 +/- 2.1% versus control, P = 0.033). There were no changes in GP IIb/IIIa activation or P selectin expression in patients or healthy subjects. Platelet aggregation (1 micromol/l ADP) in healthy subjects was inhibited only by high doses of alteplase (P = 0.001). Plasma fibrinogen levels were significantly decreased after treatment with reteplase at 1 microg/ml(1.53 +/- 0.21 versus 2.65 +/- 0.31, P = .009) and 5 microg/ml(1.55 +/- 0.16 versus 2.65 +/- 0.31, P = .005). Alteplase inhibits platelet aggregation more than tenecteplase and reteplase. The attenuation of platelet aggregation by alteplase is dissociated from the expression of activated GP IIb/IIIa and P-selectin, and by fibrinogen degradation. These results suggest that alteplase exerts its antiplatelet effect independent of GP IIb/IIIa and P-selectin expressions and fibrinogen degradation. These findings may be directly relevant to the effect of alteplase on reperfusion and to future studies using combined platelet inhibitors and thrombolytic therapy. PMID- 15650540 TI - Genetic analysis of hereditary factor X deficiency in a French patient of Sri Lankan ancestry: in vitro expression study identified Gly366Ser substitution as the molecular basis of the dysfunctional factor X. AB - We investigated a new family with cross-reactive material-positive factor X (FX) deficiency. The proband was an 11-year-old French girl from Sri Lanka with a tendency towards severe bleeding. The FX antigen level was 67%, although the activity with extrinsic pathway was 1 U/dl. The complete nucleotide sequences of all exons and exon/intron junctions of the patient's genomic DNA revealed a homozygous G <-- A substitution in exon 8, which would result in replacement of Gly366 with Ser. The proband is the first reported case of homozygote for the FX Gly366Ser mutation. Heterozygosity for Gly366Ser substitution was previously reported in a Japanese patient (FX Nagoya 2). We studied the functional consequences by expressing mutant FX Gly366Ser protein in HEK293 cells. FX Gly366Ser was secreted into the culture media at levels similar to wild-type FX; however, mutant FX activities were only 0.04, 1.05, and 0.75% of wild-type FX upon activation by the extrinsic system, the intrinsic system, and Russell's viper venom, respectively. Moreover, the activity of FX Gly366Ser was undetectable when analyzed with chromogenic-activated FX and thrombin generation assays. These data suggest that the Gly366Ser substitution would cause a major defect in function of the FX molecule. PMID- 15650541 TI - Thrombomodulin accelerates activated protein C production and inhibits thrombin generation in the plasma of disseminated intravascular coagulation patients. AB - Thrombomodulin (TM) has been under development as a medicine for disseminated intravascular coagulation (DIC), and is expected to exhibit strong anticoagulant activity by inhibiting thrombin generation via the acceleration of protein C activation. In the present study, we examined the pharmacological action of TM in plasma obtained from DIC patients. TM was found to inhibit thrombin generation and accelerate activated protein C (APC) production at 0.3-30 TM units/ml in plasma obtained from DIC patients irrespective of their underlying disorders. In addition, there was a positive correlation between the inhibition of thrombin generation and the amount of APC produced. Thrombin generation was inhibited by over 50% when the plasma level of APC was increased by more than 0.2 microg/ml. These results indicate that TM inhibits thrombin generation in plasma obtained from DIC patients by accelerating APC production. Moreover, the results imply that the thrombin generation test may be a good method to speculate the efficacy of TM on every patient before the administration of TM. PMID- 15650542 TI - Collagen deposition in myocardium after inhibition of fibrinolytic activity. AB - Proper function of the heart directly depends on the protection of the myocardial architecture. The fibrinolytic system plays an important role(s) in the protection of myocardial architecture and aiding the pumping function of the heart. We investigated the effect of fibrinolytic system inhibition by tranexamic acid (TXA) on cat myocardium and collagen, which are important constituents of the extracellular matrix. Twenty-eight cats (seven per group) were used. Isotonic saline was administered to the control group (C) and TXA (200 mg/kg) diluted isotonic saline given to experimental groups 1 and 2 (E1 and E2). Experimental group 3 (E3) animals received 100 mg/kg TXA intravenously daily for 7 days. Bloods were drawn from groups C, E1, and E2, and fibrinolytic activity was determined by the euglobulin lysis time, fibrinogen degradation products and fibrin plate lytic area diameters. The group C and group E1 cats were sacrificed following the infusion. The group E2 and group E3 animals were sacrificed at 24 h and day 7, respectively. Light and electron microscopy, along with the collagen contents of the myocardium, were used to examine the myocardial tissues. Electron microscopic examination in groups E1 and E2 showed inter-myofibrillar edema, glycogen loss, mitochondrial swelling and disorganization of Z-bands, and a decrease of pinocytotic vesicles in capillary endothelial cells. In group E3, increases of collagen fibrils in intercellular areas and perivascular areas were noted. Biochemical analyses revealed a highly significant collagen accumulation in the myocardium in group E3 (P < 0.01) as compared with group C. These findings suggest that accumulation of the extracellular matrix containing collagen in the myocardium by fibrinolytic system inhibition may be responsible for the abnormal myocardial architecture, leading to cardiomyopathy and altered cardiac function . PMID- 15650543 TI - Soluble thrombomodulin levels among women with a history of recurrent pregnancy loss, with or without antiphospholipid antibodies. AB - The endothelial cells produce substances whose elevated plasma levels acquire predictive value for the development of events. For instance, soluble thrombomodulin (sTM) levels evidence endothelial cell injury. Under specific clinical conditions the levels of sTM are raised, such as in patients with certain autoimmune disorders, pre-eclampsia or antiphospholipid syndrome. The levels of sTM, as an endothelial injury marker, were evaluated in 65 women with a history of recurrent pregnancy loss (12 with autoimmune disorders, 19 pregnant women and nine with a history of gestational hypertension or pre-eclampsia or eclampsia); 13 of them had antiphospholipid antibodies. sTM levels could be used as a predictor of pregnancy loss in future prospective studies. We compared those levels with the levels found in control groups without recurrent pregnancy loss (20 healthy women and 14 women with autoimmune disorder). There were no statistically significant differences (P = 0.729) in the levels of sTM between the recurrent pregnancy loss group (31.1 ng/ml) and the healthy control group (31.4 ng/ml) or between the different subgroups with recurrent pregnancy loss (P = 0.873) and the healthy control group or the control group with autoimmune disorder (28.0 ng/ml). There were no statistically significant differences (P = 0.605) in the levels of sTM among the patients with recurrent pregnancy loss, with or without moderate or high antiphospholipid antibodies (32.0 versus 23.3 ng/ml). Consequently, the levels of sTM would not seem to be a useful tool, as an endothelial injury marker, in women with a history of recurrent pregnancy loss with or without antiphospholipid antibodies. PMID- 15650544 TI - Plasma replacement therapy during labor is not mandatory for women with severe factor XI deficiency. AB - Severe factor XI deficiency is an injury-related bleeding disorder. The risk of excessive post-partum hemorrhage in affected women has so far been evaluated in a relatively small number of patients and it is uncertain whether prophylactic treatment with fresh frozen plasma or factor XI concentrate is needed during or after vaginal or cesarean delivery. We retrospectively analyzed bleeding manifestations related to vaginal and/or cesarean deliveries in a cohort of 62 women with factor XI activity < 17 U/dl and evaluated whether replacement therapy is essential. Fifty-one women had 139 vaginal deliveries, six women had 13 cesarean deliveries, and five women had seven vaginal as well as five cesarean deliveries. Forty-three of the 62 women (69.4%) never experienced post-partum hemorrhage during 93 deliveries (85 vaginal, eight cesarean). Hemorrhage occurred in 19 women, which in six women accompanied each one of their 17 vaginal deliveries. Post-partum hemorrhage had no relationship with the abnormal genotype that caused factor XI deficiency nor with factor XI level. These observations suggest that the use of fresh frozen plasma or factor XI concentrate during and/or after vaginal delivery is not mandatory in women with severe factor XI deficiency and can be reserved for patients who develop excessive hemorrhage. For women requiring cesarean section it appears that the same policy can be advocated but more observations are needed. PMID- 15650545 TI - Early cerebral sinovenous thrombosis in a child with acute lymphoblastic leukemia carrying the prothrombin G20210A variant: a case report and review of the literature. AB - This paper describes the case of a male child with acute lymphoblastic leukemia who developed cerebral sinovenous thrombosis on day 6 of induction therapy with intrathecal methotrexate and methylprednisolone. A central venous catheter had been implanted the day before. The heterozygous prothrombin G20210A variant was found together with acquired activated protein C resistance and a reduced activated partial thromboplastin time. Clinical course and magnetic resonance imaging with magnetic resonance venography gradually improved over the following days after starting anticoagulant therapy (heparin and nadroparin). Cerebral sinovenous thrombosis is a serious disease in leukemic children, occurring in up to 6% of these patients. Data from the literature are in favor of anti-coagulant treatment, even though the efficacy and safety of thromboprophylaxis during chemotherapy in leukemic children with inherited thrombophilic conditions remain to be demonstrated. PMID- 15650546 TI - Resolution of Budd-Chiari syndrome due to activation of endogenous fibrinolysis that may be induced by weight reduction. AB - We describe a 45-year-old female with polycythemia vera and Leiden factor V mutation, who suffered the subacute form of Budd-Chiari syndrome and was treated with anticoagulants and diuretics. Surprisingly, after 3 months clinical signs of Budd-Chiari syndrome resolved; venography disclosed the resolution of thrombosis in the vena cava inferior and hepatic veins. This was associated with considerable increase of endogenous fibrinolytic activity, documented by a substantial change in the euglobulin clot lysis time, and a decrease of plasminogen activator inhibitor antigen and activity. During the disease the patient followed a diet and significantly reduced her body weight. Putting all data together it could be speculated that weight reduction (along with anticoagulants) considerably activated endogenous fibrinolysis, resulting in spontaneous resolution of Budd-Chiari syndrome. The validity of this explanation should be explored in a larger clinical study. PMID- 15650547 TI - An increase of blood anti-beta2-glycoprotein I antibody in Japanese encephalitis associated with cerebral ischemia. AB - The role of humoral autoimmunity in virus-induced vascular thrombosis is still not clear. We encountered a patient who experienced cerebral ischemia in his early course of Japanese encephalitis. At the beginning, an increase of blood immunoglobulin G isotype of anti-beta2-glycoprotein I antibody, a prolonged activated partial thromboplastin time and thrombocytopenia resembling antiphospholipid antibody syndrome were found, and these abnormalities disappeared when the patient recovered later. A molecular mimicry between the T(2688)LRVLE in Japanese encephalitis virus and hexapeptide-TLRVYK may contribute for the patient's anti-beta2-glycoprotein I antibody generation. Therefore, an increase of procoagulative antibody, such as anti-beta2-glycoprotein I antibody, may display a crucial role for cerebral thrombosis associated with infectious pathogens such as Japanese encephalitis virus. The interaction between autoimmunity induction by infectious agents and procoagulation in the occurrence of vascular thrombosis may be more important than has been understood in previous studies. PMID- 15650548 TI - Modifications of flow measurement to determine fibrin gel permeability and the preliminary use in research and clinical materials. AB - Our earlier investigations employing a flow measurement have yielded intriguing findings as to what governs the fibrin network porosity. To make the method suitable for use by more groups with various laboratory conditions, sample materials or study purposes, we simplified the essential equipment and thereby minimized the sample volume to 250 microl in comparison with the need for 3000 microl in the previous method. To assess whether the fibrin gel permeability depends on changes in thrombin generation potential and/or fibrinogen clotting property, different concentrations of thrombin with or without frozen-thawed platelets, serving as phospholipids, were used. The platelets and 0.05 IU/ml thrombin were added to plasma samples from patients with previous myocardial infarction. The fibrin gel permeability, expressed as Darcy constant (Ks), was decreased compared with that in controls, supporting findings about high risk of thromboembolism in this disease due to increases of thrombin activity and fibrinogen function. When 0.4 IU/ml thrombin was used in samples provided by 10 healthy individuals treated with acetysalicylic acid, Ks levels were increased during versus before therapy. Since almost no thrombin generation was found in the samples with the higher dose of exogenous thrombin, we considered that modifications in fibrinogen clotting property by acetysalicylic acid rendered the fibrin network more permeable. In summary, as the reproducibility remains satisfactory (coefficient of variation < 10%) despite aforementioned modifications in the equipment and reagents, any interested laboratory ought to be able to repeat the method. Assays of fibrin permeability in such a simple way may help to determine the fibrin clot stability in pathological/pharmacological studies, and probably serve as a tool to estimate thromboembolism risk in clinical materials, such as patients with cardiovascular diseases. PMID- 15650549 TI - Performance and prognostic importance of a new clinical and laboratory scoring system for identifying non-overt disseminated intravascular coagulation. AB - A template for diagnosing the non-overt phase of disseminated intravascular coagulation (DIC) has recently been proposed. However, validation of its performance and the proposal of a defining score are required. The aim was to assess feasibility of the non-overt DIC scoring template and its potential prognostic significance. Consecutive patients admitted to a university hospital intensive care unit were initially assessed over 2 months. Following this, a 12 month study examined the prognostic performance of the derived scores prospectively. Outcome parameters were overt DIC and 28-day mortality. The 2 month study, involving 66 patients and 919 time points, demonstrated practical feasibility and prognostic associations for mortality with scores of 5 and greater. The 12-month study involving 450 patients showed that the mortality rate was 29% (105 of 360) and 78% (70 of 90) for scores below 5 and scores of 5 or above, respectively. The mortality rate for overt DIC was also 78% (38 of 49). The non-overt DIC scoring template is workable and has prognostic relevance. A score of 5 and greater is recommended as diagnostic of non-overt DIC. PMID- 15650550 TI - Differential effect of the ABO blood group on von Willebrand factor collagen binding activity and ristocetin cofactor assay. AB - Diagnosis of von Willebrand disease requires a combination of quantitative and qualitative tests including the von Willebrand factor ristocetin cofactor assay (vWF:RCo), the von Willebrand factor collagen binding assay (vWF:CB) and von Willebrand factor antigen quantification (vWF:Ag). Genetic factors, especially the ABO blood group, contribute significantly to the variation in vWF levels and function. Recent studies suggest that ethnicity may be another important modulator. We investigated the effect of the ABO blood group on these tests in 52 blood group O and 54 non-group O Chinese blood donors who were well-matched for sex and age distribution. Group O donors had significantly lower vWF:RCo, vWF:Ag and vWF:CB (P < 0.0001). Reduction in vWF:CB was greater than vWF:RCo in group O donors (53 versus 27%). This led to a lower vWF:CB/vWF:Ag ratio (P < 0.0001) in group O donors whereas the vWF:RCo/vWF:Ag ratio was unaffected (P = 0.97). These variations should be taken into consideration when interpreting these results in the Chinese. PMID- 15650551 TI - A case of deficiency of plasma plasminogen activator inhibitor-1 related to Ala15Thr mutation in its signal peptide. AB - The patient was a 34-year-old man with life-long bleeding episodes, whose hemorrhage problem was characterized predominantly by prolonged bleeding at surgical or traumatic sites. All routine coagulation parameters were within normal ranges. The patient's bleeding tendency was not caused by factor XIII deficiency, alpha2-antiplasmin deficiency, or tissue type-plasminogen activator increase. His characteristic abnormalities of fibrinolysis included shortened euglobulin clot lysis time, low plasminogen activator inhibitor-1 (PAI-1) activity and antigen in plasma, which were remarkably reduced to only about 10% of control. An operation was performed in order to clear two hematomas in the patient's left leg and hip, and subsequent bleeding episodes were well controlled with adjuvant administration of intravenous aminomethylbenoic acid after surgery. PAI-1 gene analysis by polymerase chain reaction product sequencing revealed that the patient had a heterozygous missense mutation G to A transition at nucleotide position 4497 in exon 2, causing replacement of alanine 15 (GCC) to threonine (ACC) at signal peptide. The restriction endonuclease analysis showed that this gene mutation also existed in the patient's father, but not in his mother and 60 normal subjects. The wild-type and mutant plasmids were constructed and transiently transfected into Chinese hamster ovary cell lines; the levels of PAI 1 activity and antigen in the media of the mutant were approximately 70% of the wild type, and the levels of PAI-1 protein in cell lysates were almost equal in wild-type and mutant plasmids. These results indicate that the mutation in signal peptide may partly impair the secretion of PAI-1. PMID- 15650552 TI - Normal D-dimer levels in patients with latent tuberculosis infection. AB - Studies have demonstrated a link between acute pulmonary tuberculosis and a hypercoagulable state, but there are no data on the coagulation state of patients with latent tuberculosis infection (LTI). The present prospective observational study was designed to help fill this gap. The sample included 84 patients (high school students and adults) with suspected LTI referred for the purified protein derivative (PPD) test. Results were read according to the criteria of the American Thoracic Society. Blood samples were collected at admission and assayed for D-dimer, the marker of the coagulation state, with the quantitative Miniquant test. D-dimer values were correlated with the PPD status and clinical parameters. Fifty-seven patients tested positive for LTI and 27 tested negative. There was no significant difference in D-dimer level between these groups (341 +/- 106 and 360 +/- 60 microg/ml, respectively). No significant correlation was found between D dimer level and PPD status, patient age or occupation (health care worker or not), or clinical indication for the tuberculin test. The normal D-dimer levels in this series suggest that low-level inflammations such as LTI do not lead to a hypercoagulable state. PMID- 15650554 TI - Is complete remission of cyclic thrombocytopenia related to correction of cobalamin deficiency due to H. pylori eradication? PMID- 15650556 TI - Nutrients and cardiovascular disease: no easy answers. PMID- 15650557 TI - Intentional weight loss, blood lipids and coronary morbidity and mortality. AB - PURPOSE OF REVIEW: Although weight reduction has been recommended to reduce cardiovascular risk, studies on the association between weight loss and coronary morbidity and mortality show conflicting results. This review summarizes findings from large studies examining this issue and accentuates the importance of carrying out additional well-designed research. RECENT FINDINGS: Many observational studies report that weight loss in older men and women is associated with increased cardiovascular morbidity and mortality. Recent studies suggest that this association may arise from the confounding effect of preexisting disease. Many studies do not report whether weight loss is intentional or unintentional. Unintentional weight loss may mask beneficial changes in cardiovascular risk due to intentional weight loss. In addition to issues related to the cause of weight loss, use of reported rather than measured weight may bias the results of large studies. However, one recent observational study with a methodology aimed at overcoming these limitations found that individuals who intentionally lost weight experienced a decreased coronary risk. SUMMARY: Weight reduction in overweight individuals is not universally associated with good health. This is true even if the weight loss results in normal body mass index. Reports of increased coronary risk associated with intentional weight loss may be explained by comorbidities that are also associated with weight loss. Individuals who are overweight and at high coronary risk may benefit from professionally supervised dieting and avoiding regain of lost weight. Clinical trials on cardiovascular outcomes in individuals who lose weight under supervised dieting are needed to assess this recommendation definitively. PMID- 15650558 TI - Omega-3 fatty acids: molecular approaches to optimal biological outcomes. AB - PURPOSE OF REVIEW: This review discusses recent advances in delineating basic mechanisms underlying the beneficial effects of omega-3 fatty acids on health and on disease. RECENT FINDINGS: While a substantial number of studies have delineated many differences between the biological effects of saturated versus polyunsaturated fatty acids, less is known about the long-chain omega-3 fatty acids commonly present in certain fish oils. In this review, we focus on recent studies relating to basic mechanisms whereby omega-3 fatty acids modulate cellular pathways to exert beneficial effects on promoting health and decreasing risks of certain diseases. We will use, as examples, conditions of the cardiovascular, neurological, and immunological systems as well as diabetes and cancer, and then discuss basic regulatory pathways. SUMMARY: Omega-3 fatty acids are major regulators of multiple molecular pathways, altering many areas of cellular and organ function, metabolism and gene expression. Generally, these regulatory events lead to "positive" endpoints relating to health and disease. PMID- 15650559 TI - ATP-binding cassette transporter AI and its role in HDL formation. AB - PURPOSE OF REVIEW: ATP-binding cassette transporter AI (ABCA1)-mediated assembly of phospholipid and free cholesterol with apoA-I plays an important role in HDL biogenesis. This review focuses on recent progress in ABCA1-mediated HDL formation and regulation of ABCA1 expression. RECENT FINDINGS: Studies of hepatic ABCA1 overexpression suggest that the liver is a major site for HDL formation. Lipidation of apoA-I by ABCA1 increases its potential for reverse cholesterol transport based on the following findings: (1) apoA-I/lipid complexes formed by ABCA1 are better acceptors of cellular lipid via non-ABCA1-mediated efflux pathways than lipid-free apoA-I in vitro and (2) lipidation of apoA-I prevents it from rapidly associating with plasma HDL in vivo, resulting in more available nascent pre-beta HDL for cellular lipid efflux. Several novel regulatory mechanisms for ABCA1 at the post-transcriptional level have been identified recently. Interaction of apoA-I with ABCA1 prevents phosphorylation of a sequence rich in proline, glutamic acid, serine and threonine in a cytoplasmic domain of ABCA1, resulting in less degradation by calpain proteolysis and increased surface expression of ABCA1. In addition, destabilization and decreased cellular surface expression of ABCA1 protein by unsaturated fatty acids have been identified. SUMMARY: Initial lipidation of apoA-I by hepatic ABCA1 is critical for plasma HDL formation because it enables pre-beta HDL to function more efficiently as a cholesterol acceptor for other pathways of cholesterol efflux in the reverse cholesterol transport pathway and prevents apoA-I from rapidly associating with preexisting plasma HDL particles, resulting in greater availability of pre-beta HDL particles for cholesterol efflux. PMID- 15650560 TI - Metabolic and health effects of isomeric fatty acids. AB - PURPOSE OF REVIEW: Isomeric fatty acids have the same number of carbon and hydrogen atoms, but may have distinct metabolic and health effects. Two well known examples of isomeric fatty acids are cis and trans monounsaturated fatty acids, and conjugated isomers of linoleic acid (CLA). The purpose of this review is to summarize recent findings from human studies on the metabolic and health effects of these two classes of isomeric fatty acids. RECENT FINDINGS: Apart from an unfavorable effect on serum lipoproteins, trans monounsaturated fatty acids from hydrogenated oils may increase plasma markers for a low-grade inflammatory state. From epidemiological studies, however, it is not possible to conclude if effects of ruminant and industrial trans fatty acids on cardiovascular risk are different. In contrast to in-vitro and animal studies, there are no indications that in humans the two most common CLA isomers (cis9,trans11-CLA and trans10,cis12-CLA) affect body composition differently. Longer-term supplementation, however, may slightly decrease body fat mass without apparent effects on plasma markers for glucose and lipid metabolism. Other studies have even reported adverse effects of CLA supplementation on insulin resistance and lipid peroxidation. SUMMARY: Evidence is increasing that trans monounsaturated fatty acids from hydrogenated oils increase plasma markers of low-grade chronic inflammation. From epidemiological studies, however, it is not clear if effects of ruminant and industrial trans fatty acids on cardiovascular risk are different. Effects of CLA on body composition remain controversial and more research is needed before the widely available CLA supplements should be advocated as an adjunct to control body weight. PMID- 15650561 TI - Food components and immune function. AB - PURPOSE OF REVIEW: Enhancing immune function or alternatively dampening inflammatory processes by specific food components has received a lot of interest. The purpose of this review is to summarize recent findings with the emphasis on underlying mechanisms. RECENT FINDINGS: Dietary beta-glucans are relatively new candidates in the field of immune modulation by diet. In-vitro and animal studies suggest that beta-glucans shift inflammatory profiles to a Th1 type, which may enhance resistance against bacterial and parasitic infections. Regarding polyunsaturated fatty acids, there is evidence that n-3 fatty acids from fish oils (eicosapentaenoic acid and docosahexaenoic acid) dampen inflammatory responses. Whether eicosapentaenoic acid or docosahexaenoic acid is responsible for this phenomenon remains controversial. It is also inconclusive whether the plant-derived n-3 fatty acid alpha-linolenic acid has the same antiinflammatory effects as observed for fish oils. Saturated fatty acids may activate toll-like receptors and consequently the inflammatory pathway. The effect of total fat intake is controversial, since high-fat diets have been found to suppress immune function, while also improving intestinal barrier function. Finally, Gingko biloba was found to lower nuclear factor kappaB and activator protein 1 activation, possibly due to its high content of polyphenols. SUMMARY: In this review we discuss the nutritional components able to enhance immune function or show antiinflammatory effects. It can be concluded that diet certainly has the potential to direct immune responses. Apart from studies on fish oils, however, evidence from human studies is limited. PMID- 15650562 TI - The clinical and nutritional implications of lipid-lowering drugs that act in the gastrointestinal tract. AB - PURPOSE OF REVIEW: A new class of cholesterol-lowering therapy that reduces intestinal sterol absorption has recently been introduced. This increases the number of classes of lipid-lowering agents that directly affect gastrointestinal function and raises questions concerning the overall effect of these agents on absorption and nutritional status. RECENT FINDINGS: A recent assessment notes a paucity of information concerning the factors that affect the bioavailability and intestinal absorption of lipophilic nutrients. By contrast, the specificity of the mechanisms of action of new drugs acting on the gastrointestinal tract may circumvent some of the detrimental effects on nutrient and drug bioavailability that have been noted with older forms of treatment. SUMMARY: The clinical imperative for aggressive control of lipid and metabolic risk factors makes widespread use, alone or in combination, of lipid-lowering agents that affect the gastrointestinal tract seem increasingly likely. Whilst the opportunity for therapeutic synergy is attractive, care will be required to avoid interference with intestinal absorptive function. PMID- 15650563 TI - Dietary antioxidants and cardiovascular disease. AB - PURPOSE OF REVIEW: Oxidative damage is involved in cardiovascular diseases. Intervention with alpha-tocopherol, ascorbic acid and beta-carotene does not appear to reduce pathogenesis. The purpose of this review is to describe alternative antioxidant mechanisms that may be involved. RECENT FINDINGS: Antioxidants with different chemical properties may recharge each other in an antioxidant network. The total antioxidant content of dietary plants may therefore be a useful tool for testing the "antioxidant network" hypothesis. Several berries, fruits, nuts, seeds, vegetables, drinks and spices have been found to be high in total antioxidants. Initial studies in animals and humans are supportive as to the beneficial effects of dietary plants rich in total antioxidants. Additionally, antioxidants and other plant compounds may also improve the endogenous antioxidant defence through induction of antioxidant and phase 2 enzymes. Dietary plants rich in such compounds include broccoli, Brussels sprouts, cabbage, kale, cauliflower, carrots, onions, tomatoes, spinach and garlic. SUMMARY: Although initial studies have indicated that antioxidants may reduce oxidative stress, human intervention studies do not support a beneficial effect of antioxidant supplements. Further research is needed to clarify whether other plant antioxidants, plants rich in a combination of antioxidants, or plant compounds that induce the endogenous antioxidant defence can reduce pathogenesis of cardiovascular disease and other oxidative stress-related diseases. PMID- 15650564 TI - Insulin resistance, low-fat diets, and low-carbohydrate diets: time to test new menus. AB - PURPOSE OF REVIEW: Insulin resistance increases the risk of cardiovascular disease and diabetes, and the risk of cardiovascular disease increases further once diabetes has developed. As insulin resistance is a precursor to diabetes, it is critically important to identify cost-effective means, such as dietary changes, by which to reduce insulin resistance. The purpose of this review is to evaluate recent findings concerning dietary composition and insulin resistance, with particular focus on low-fat diets compared with the currently popular low carbohydrate diets. RECENT FINDINGS: Recent findings indicate little support for the value of low-carbohydrate diets as therapies for insulin resistance. In contrast, the limited data available suggest that the higher fat content of typical low-carbohydrate diets may exacerbate insulin resistance in the long term. Preliminary data indicate that proteins from different sources may have differing effects on insulin resistance. Preliminary data also suggest the potential value of whole grains, fruits and vegetables in therapeutic diets to reduce insulin resistance. SUMMARY: Current evidence supports the inclusion of whole grains, fruits and vegetables, and lean sources of animal proteins including low-fat dairy products in dietary therapies for insulin resistance. Those who wish to follow a low-carbohydrate diet should be encouraged to follow a new menu low in fat, and with most of the protein derived from plant sources. PMID- 15650565 TI - The effect of genetic variation on the lipid response to dietary change: recent findings. AB - PURPOSE OF REVIEW: Dyslipidaemia is an important risk factor for cardiovascular disease, and can be modified by diet. However, the lipid response to dietary change may be influenced by genetic variation. This review examines recent research (published since August 2003) on the effect of genetic variation on the lipid response to dietary change. RECENT FINDINGS: In 10 reports describing intervention studies and seven reports describing observational studies, the lipid response to diet was modified by polymorphisms within the genes for apoE, apoB, apoCIII, lipoprotein lipase, hepatic lipase, endothelial lipase, the liver fatty acid-binding protein, the beta3-adrenergic receptor, adipsin and the peroxisome proliferator-activated receptor gamma. The studies varied widely in terms of the number and type of study participants, the composition and duration of the dietary interventions, the nutrients studied and dietary assessment methods used in the observational studies, and the polymorphisms analysed--some of which had not been studied before with regard to the lipid response to diet. SUMMARY: The lipid response to dietary change is highly complex. Future studies will have to be large in order to assess the effects of multiple polymorphisms, and will have to control for many factors other than diet. At present, it is premature to recommend the use of genotyping in the design of therapeutic diets. However, such studies may be useful in identifying the mechanisms by which dietary components influence lipid levels. PMID- 15650566 TI - Glycemic index, postprandial glycemia and cardiovascular disease. AB - PURPOSE OF REVIEW: Several lines of evidence indicate that exaggerated postprandial glycemia puts individuals without diabetes at greater risk of developing cardiovascular disease. In large, prospective observational studies, including meta-analyses, higher 120 min post-load blood glucose and glycated hemoglobin (a measure of average blood glucose level over time) independently predict cardiovascular mortality and morbidity in individuals without diabetes. These findings imply that the glycemic nature of dietary carbohydrates may also be relevant. We aim to provide a clearer perspective on how the glycemic impact of carbohydrates may modulate development of cardiovascular disease. RECENT FINDINGS: In ecological studies, average dietary glycemic index (a measure of the postprandial glycemic potential of carbohydrates) and glycemic load (average glycemic index x amount of carbohydrate) predicts coronary infarct and cardiovascular disease risk factors, including HDL cholesterol, triglycerides and C-reactive protein. In short-term intervention studies of overweight and hyperlipidemic patients, low glycemic index diets lead to improvements in cardiovascular disease risk factors, including reduced LDL cholesterol and improved insulin sensitivity, as well as greater body fat loss on energy restricted diets. Molecular studies indicate that physiological hyperglycemia induces overproduction of superoxide by the mitochondrial electron-transport chain, resulting in inflammatory responses and endothelial dysfunction. SUMMARY: Taken together, the findings suggest that conventional high-carbohydrate diets with their high glycemic index may be suboptimal, particularly in insulin resistant individuals. Because around one in four adults has impairments in postprandial glucose regulation, the glycemic potential of carbohydrates warrants further investigation in cardiovascular disease prevention. PMID- 15650567 TI - Polyphenols and prevention of cardiovascular diseases. AB - PURPOSE OF REVIEW: Polyphenols are the most abundant dietary antioxidants and research on their role in the prevention of degenerative diseases has developed quickly over these last few years. This paper reviews the recent studies on the prevention of cardiovascular diseases by polyphenols, focusing on human studies. RECENT FINDINGS: A large number of recent intervention studies have shown that several biomarkers of cardiovascular risk are influenced by the consumption of polyphenol-rich foods. Effects on biomarkers of oxidative stress, lipemia and inflammation appear so far inconclusive. More consistent effects have been observed on endothelial function and haemostasis and support a reduction of risk by polyphenols in agreement with the few epidemiological studies already published. All clinical studies have used foods or beverages containing a mixture of different polyphenols and the exact nature of the most active compounds remains largely unknown. Absorption, metabolism and elimination vary widely between polyphenols. These data on bioavailability should be taken into account to improve the experimental design and the interpretation of the observed effects. SUMMARY: Future intervention studies should include a detailed assessment of the bioavailability of polyphenols. Beyond clinical trials carried out with polyphenol-rich foods, more studies with pure polyphenols will also be needed to establish their role in the prevention of cardiovascular diseases. PMID- 15650568 TI - Bibliography. Current world literature. Nutrition and metabolism. PMID- 15650569 TI - Nutrition and metabolism: antioxidants, the food matrix and methodological considerations. PMID- 15650570 TI - Genetics and molecular biology: ABCA1 in brain apolipoprotein E metabolism and lipidation. PMID- 15650571 TI - Lipid metabolism: lipoproteins in the metabolic syndrome and subclass sex and age differences. PMID- 15650573 TI - The age of refractive lens surgery. PMID- 15650574 TI - Bimanual microincisional phacoemulsification: the future of cataract surgery? AB - PURPOSE OF REVIEW: Bimanual microincisional cataract surgery has recently become a procedure of interest among cataract surgeons, and a number of trials have shown its potential as a minimally invasive cataract surgery. The purpose of this review is to examine the studies that have been published to date and to evaluate the potential of bimanual phacoemulsification as a method of cataract extraction. RECENT FINDINGS: Recent studies have reinforced the safety of bimanual phacoemulsification. In particular, recently published studies have focused on evaluating various phacoemulsification technologies and their safety when used in bimanual phacoemulsification. Newly developed rollable hydrophilic acrylic ThinOptX lenses have been shown to be implantable in 2.2-mm incisions safely with good visual outcomes. SUMMARY: Bimanual phacoemulsification has been a potential technique for a number of years, but only recently have the technology, software, and technique advanced sufficiently to make bimanual phacoemulsification a feasible method of cataract extraction. Although the main disadvantage to bimanual phacoemulsification remains the lack of intraocular lenses that can fit through microincisions, necessitating the enlargement of corneal wounds for intraocular lens implantation, bimanual phacoemulsification has a number of advantages over traditional small-incision phacoemulsification. Theses advantages have been a source of interest for cataract surgeons and surgical companies who are now developing technologies that will permit the performance of truly microincisional cataract surgery. PMID- 15650575 TI - Accommodative intraocular lenses: current status. AB - PURPOSE OF REVIEW: The possibility of using a monofocal IOL with accommodative ability allows refractive cataract surgery with a clearly decreased potential of photic phenomena. Three IOLs of different designs and materials have demonstrated accommodative ability, but the degree of accommodative amplitude has been reported to different extents and variabilities. The plate-haptic CrystaLens has a hinged design that might permit forward movement of the optic as a result of pressure changes in the vitreous cavity. The 1CU has modified haptics that bend in the bag as the lens capsule contracts, which are supposed to cause anterior displacement of the lens optic. With the dual-optic one-piece Synchrony, springlike haptics separate a high-plus anterior lens from a posterior minus lens. With accommodative effort, the capsular bag expands and the springs express kinetic energy, which might allow the optics to separate as the anterior lens moves forward. RECENT FINDINGS: This article seeks to clarify and distinguish the concepts of true accommodation and pseudo accommodation. Current designs of accommodative IOLs are supposed to work by the focus-shift principle to allow true pseudophakic accommodation. Studies that biometrically assessed optic shift found no or only low amplitudes of forward movement. The amount of forward movement, if present, was highly variable between patients. To date, most studies present psychophysical data for the proof of concept, which alone seems insufficient. Capsule bag performance and posterior capsule opacification with accommodative IOLs seem worse than those with standard intraocular lenses. SUMMARY: The potential clinical benefits of accommodative IOL technology for both cataract patients and refractive patients may place accommodative IOLs in a competitive position with multifocal IOL technology. PMID- 15650576 TI - Teaching phacoemulsification in US ophthalmology residencies: can the quality be maintained? AB - PURPOSE OF REVIEW: To examine the current status of phacoemulsification training and the factors that will influence the future quality of training in phacoemulsification. RECENT FINDINGS: Recent developments include residents performing phaco without previous ECCE experience, residents using topical anesthesia in early surgical cases, virtual surgery, improved techniques for surgical practice with animal eyes, and modular surgical training. SUMMARY: Learning to perform cataract surgery is a universal experience for US ophthalmologists. Training in phacoemulsification during residency is one of the most important components of postgraduate education in ophthalmology. The quality of phacoemulsification training in the US is currently threatened by a confluence of factors including decreasing surgical numbers, rapidly changing technology, and shrinking financial resources. The impending decline in quality of phacoemulsification training in US residencies can be prevented if creative and innovative solutions to these problems are applied. These types of solutions include use of improved animal eye practice, virtual surgical practice, alterations in live surgery teaching techniques, and finding additional sources of financial support for surgical teaching. PMID- 15650577 TI - Multifocal intraocular lenses. AB - PURPOSE OF REVIEW: Multifocal intraocular lenses (IOLs) are growing in popularity among patients and surgeons, and opened the way to refractive lens exchange. Still they are not used routinely in cataract surgery, for reasons probably connected to the frequently observed reduction in contrast sensitivity. Recent papers with clinical study outcomes can help in understanding the advantages and the limits of multifocal IOLs. RECENT FINDINGS: Emerging from every published study, both refractive and diffractive multifocal IOLs usually provide good near visual acuity with distance correction. As many multifocal IOLs are distance dominant, near vision can be improved by correcting for near the distance focus. The near contrast sensitivity thus obtained is similar to that of monofocal IOLs. Multifocal IOLs have been employed with success in complicated cataract surgery and in trauma cases, with the same outcome as in normal cataract cases. Presbyopic lens exchange remains controversial, with a high success rate in original ametropic eyes, but limited success in original emmetropic eyes. Secondary procedures to improve the refractive outcome are usually of little efficacy in improving patient satisfaction. A new anterior chamber phakic multifocal IOL has been designed to correct presbyopia and small refractive errors. The first clinical results indicate high patient satisfaction, with 7.3% explantation rate. SUMMARY: Multifocal IOLs can be more widely used after cataract surgery, but should be used with caution in almost emmetropic eyes with little or no cataract. Refractive lens exchange with multifocal IOL is especially worthwhile in hyperopic eyes. The new phakic multifocal IOL adds the concept of reversibility to presbyopic lens exchange. PMID- 15650578 TI - New phacoemulsification technology. AB - PURPOSE OF REVIEW: Familiarization with each of the manufacturers' latest machines and upgrades assists in understanding the advances made in phacoemulsification technology. RECENT FINDINGS: Improvements in the delivery of energy through power modulations, alternative and adjunctive energy sources, and advanced fluidics allow the safe use of bimanual microincision techniques for lens extraction. SUMMARY: Continual advances in phacoemulsification technology and adjunctive surgical devices allow cataract surgery to be performed more safely and efficiently. PMID- 15650579 TI - Surgical strategies in patients with cataract and glaucoma. AB - PURPOSE OF REVIEW: This review analyzes the most relevant studies on current surgical strategies to treat glaucoma patients with cataracts. RECENT FINDINGS: No clear evidence has confirmed better results with trabeculectomy alone compared with phacotrabeculectomy. Recent studies have reported successful outcomes combining deep sclerectomy and two-site phacoemulsification. The phacoemulsification cataract extraction will not vary the intraocular pressure of patients with previous deep sclerectomy. Mitomycin C proved to be effective in maintaining lower pressure levels with the combined surgery technique; however, 5 fluorouracil did not show any improvement. Minimally invasive cataract surgery reduces surgical trauma, making it possible to obtain better results with combined surgery and previous glaucoma surgery. SUMMARY: The surgical strategy decision must be customized to every patient. Only filtering surgeries are recommended in glaucoma patients with incipient cataract. Combined surgical procedures are recommended for progressive or advanced glaucoma. Two-site phacotrabeculectomy with mitomycin C achieves better stabilized results; however, combined phacoemulsification with deep sclerectomy or viscocanalostomy achieves similar results with a lower rate of complications. These promising findings need more study to be confirmed. PMID- 15650580 TI - Refractive lens exchange for myopia: a new perspective? AB - PURPOSE OF REVIEW: Although lens surgery for high myopia has a long History, it has been extremely controversial. Colin's study of 1999 indicated a much higher than expected rate of retinal detachment. The purpose of this review is to reassess the place of refractive lens exchange in high myopia in the light of more recent reports in the literature and a more critical examination of Colin's data. RECENT FINDINGS: A meta analysis of papers on refractive lens exchange and cataract in high myopia between 1996 and 2004 included some 2036 eyes. The mean follow up was 43.5 months and overall incidence of retinal detachment was 1.85%. This is only slightly different from that expected in the highly myopic population as reported by Burton at 1.5%. The recent reports in 2003 of Ravalico et al. and Guell et al., with 4 year follow up, show only 1 detachment for 422 eyes. Further Colin's study had a group very high in risk factors having a low incidence of posterior detachment and 30% pre-existing retinal lesions. SUMMARY: Modern surgical techniques, IOLs and choosing the appropriate group to whom this procedure should be offered should lead to it becoming an accepted, safe and common refractive operation. PMID- 15650581 TI - Scleral and iris fixation of posterior chamber lenses in the absence of capsular support. AB - PURPOSE OF REVIEW: To review the methods described in the literature in the past year (from August 2003 to date). RECENT FINDINGS: Techniques for scleral and iris suturing have proven safe and effective to fixate posterior chamber intraocular lenses and avoid the sight-threatening complications of anterior chamber lenses. SUMMARY: Scleral and iris fixation have both become well-established effective means for stabilizing posterior chamber lenses in the lack of adequate capsular support. PMID- 15650582 TI - Biometry and intraocular lens power calculation. AB - This article surveys the literature of 1 year, between July 2003 and August 2004, on the topic of biometry and intraocular lens power calculation for cataract surgery. There is an increasing demand for low postoperative refractive error with rising patient expectations, especially with patients who have already undergone refractive surgery, and with developing intraocular lens technologies such as multifocal, accommodating, or toric intraocular lenses. Optical biometry has become an invaluable tool for axial length measurement, especially for a setting with a less experienced biometrist. Introduction of ray tracing for power calculation and new methods of dealing with power calculation in eyes that have undergone previous refractive surgery seem promising. New intraocular lens designs that allow adjusting the axial optic position and therefore the effective refractive power of the intraocular lens have been evaluated in animal studies. PMID- 15650583 TI - Endophthalmitis: incidence and prevention. AB - PURPOSE OF REVIEW: To present current peer-reviewed articles related to the incidence and prevention of postoperative endophthalmitis. RECENT FINDINGS: Recent literature indicates that the incidence of postoperative endophthalmitis may be on the rise. Although the preoperative use of antibiotics as prophylaxis is still controversial, it is becoming more common. SUMMARY: The reports of endophthalmitis analyzed from peer-reviewed ophthalmic journals suggest that the incidence of endophthalmitis has increased, ranging from 0.1 to 0.18% in different countries. This may be related to factors associated with the incision. Although some resistance has been detected, fourth-generation fluoroquinolones seem to be a proper antibiotic for endophthalmitis prophylaxis because of their spectrum, mode of action, and penetration. PMID- 15650584 TI - Bibliography. Current world literature. Cataract surgery and lens implantation. PMID- 15650585 TI - Uranium-munching microbes. PMID- 15650586 TI - A review of the history of U.S. radiation protection regulations, recommendations, and standards. AB - Shortly after the discovery of x rays by Wilhelm Konrad Roentgen in 1895, and the isolation of the element radium by Pierre and Marie Curie three years later, the fascination with and potential for an array of uses of ionizing radiation in medicine, science, and technology was born. As with any new technology, there was a need to balance both the beneficial and potential detrimental effects of uses of these new technologies for the advancement of humankind. In the early days, radiation hazards were not well understood. Over the decades increasing concerns in the scientific community and lay population demanded that standardized guidance and recommendations be developed for the use of ionizing radiation. Today, U.S. radiation protection standards and recommendations to protect the occupational worker, members of the general public, and the environment are numerous and complex. This review summarizes the history of the development and application of radiation protection standards and regulations to assure the safe use of radiation and radioactive materials. The evolution and roles of international and national scientific recommending and regulatory organizations that shape U.S. radiation protection policy are described and discussed. PMID- 15650587 TI - Adaptation of the ICRP publication 66 respiratory tract model to data on plutonium biokinetics for Mayak workers. AB - The biokinetics of inhaled plutonium were analyzed using compartment models representing their behavior within the respiratory tract, the gastrointestinal tract, and in systemic tissues. The processes of aerosol deposition, particle transport, absorption, and formation of a fixed deposit in the respiratory tract were formulated in the framework of the Human Respiratory Tract Model described in ICRP Publication 66. The values of parameters governing absorption and formation of the fixed deposit were established by fitting the model to the observations in 530 autopsy cases. The influence of smoking on mechanical clearance of deposited plutonium activity was considered. The dependence of absorption on the aerosol transportability, as estimated by in vitro methods (dialysis), was demonstrated. The results of this study were compared to those obtained from an earlier model of plutonium behavior in the respiratory tract, which was based on the same set of autopsy data. That model did not address the early phases of respiratory clearance and hence underestimated the committed lung dose by about 25% for plutonium oxides. Little difference in lung dose was found for nitrate forms. PMID- 15650588 TI - Method for estimating ingestion doses to the public near the Savannah River site following an accidental atmospheric release. AB - At the Savannah River Site, emergency response computer models are used to estimate dose following releases of radioactive materials to the environment. Downwind air and ground concentrations and their associated doses from inhalation and ground shine pathways are estimated. The emergency response model (PUFF PLUME) uses real-time data to track either instantaneous (puff) or continuous (plume) releases. A site-specific ingestion dose model was developed for use with PUFF-PLUME that includes the following ingestion dose pathways pertinent to the surrounding Savannah River Site area: milk, beef, water, and fish. The model is simplistic and can be used with existing code output. PMID- 15650589 TI - Electron paramagnetic resonance and fluorescence in situ hybridization-based investigations of individual doses for persons living at Metlino in the upper reaches of the Techa River. AB - Waterborne releases to the Techa River from the Mayak Production Association in Russia during 1949-1956 resulted in significant doses to persons living downstream; the most contaminated village was Metlino, about 7 km from the site of release. Internal and external doses have been estimated for these residents using the Techa River Dosimetry System-2000 (TRDS-2000); the primary purpose is to support epidemiological studies of the members of the Extended Techa River Cohort. Efforts to validate the calculations of external and internal dose are considered essential. One validation study of the TRDS-2000 system has been performed by the comparison of calculated doses to quartz from bricks in old buildings at Metlino with those measured by luminescence dosimetry. Two additional methods of validation considered here are electron paramagnetic resonance (EPR) measurements of teeth and fluorescence in situ hybridization (FISH) measurements of chromosome translocations in circulating lymphocytes. For electron paramagnetic resonance, 36 measurements on 26 teeth from 16 donors from Metlino were made at the GSF-National Research Center for Environment and Health (16 measurements) and the Institute of Metal Physics (20 measurements); the correlation among measurements made at the two laboratories has been found to be 0.99. Background measurements were also made on 218 teeth (63 molars, 128 premolars, and 27 incisors). Fluorescence in situ hybridization measurements were made for 31 residents of Metlino. These measurements were handicapped by the analysis of a limited number of cells; for several individuals no stable translocations were observed. Fluorescence in situ hybridization measurements were also made for 39 individuals believed to be unexposed. The EPR- and FISH based estimates agreed well for permanent residents of Metlino: 0.67 +/- 0.21 Gy and 0.48 +/- 0.18 Gy (mean +/- standard error of the mean), respectively. Results of the two experimental methods also agreed well with the estimates derived from the use of the TRDS-2000. For all persons investigated according to each technique, the EPR-measured dose to enamel was 0.55 +/- 0.17 Gy, and the TRDS 2000 prediction for the dose to enamel for these individuals is 0.55 +/- 0.07 Gy. The fluorescence in situ hybridization-based dose, 0.38 +/- 0.10 Gy, compared well to the TRDS-2000 prediction of external dose, 0.31 +/- 0.03 Gy, to red bone marrow for these persons. Validation of external doses at the remaining villages is an active area of investigation. PMID- 15650590 TI - In situ gamma spectrometry measurements and Monte Carlo computations for the detection of radioactive sources in scrap metal. AB - A very limited number of field experiments have been performed to assess the relative radiation detection sensitivities of commercially available equipment used to detect radioactive sources in recycled metal scrap. Such experiments require the cooperation and commitment of considerable resources on the part of vendors of the radiation detection systems and the cooperation of a steel mill or scrap processing facility. The results will unavoidably be specific to the equipment tested at the time, the characteristics of the scrap metal involved in the tests, and to the specific configurations of the scrap containers. Given these limitations, the use of computer simulation for this purpose would be a desirable alternative. With this in mind, this study sought to determine whether Monte Carlo simulation of photon flux energy distributions resulting from a radiation source in metal scrap would be realistic. In the present work, experimental and simulated photon flux energy distributions in the outer part of a truck due to the presence of embedded radioactive sources in the scrap metal load are compared. The experimental photon fluxes are deduced by in situ gamma spectrometry measurements with portable Ge detector and the calculated ones by Monte Carlo simulations with the MCNP code. The good agreement between simulated and measured photon flux energy distributions indicate that the results obtained by the Monte Carlo simulations are realistic. PMID- 15650591 TI - Output of radiopharmaceutical nuclides of known injected doses from a municipal sewage treatment system. AB - The concentrations of (99m)Tc, (123)I, (67)Ga, and (201)Tl of the discharge water and the sewage sludge from Kurume Central Sewage Treatment Plant were determined once a week for 2 mo. The radiopharmaceutical doses injected into the patients for all of four hospitals that use nuclear medicines in the service area of the plant were also surveyed. Approximately 1.5% of the (99m)Tc, 1.5% of the (123)I, 4.3% of the (67)Ga, and 0.41% of the (201)Tl of the injected doses were detected in the discharged water from the plant. The behavior of these radionuclides in the sewage treatment system was analyzed using a compartment model. The analyses suggest that the average residence times in storage tanks and drainage pipes before entering the plant were 9.5 h for (99m)Tc, 81 h for (123)I, 120 h for (67)Ga, and 480 h for (201)Tl. PMID- 15650592 TI - Measurements of radioactivity in books and calculations of resultant eye doses to readers. AB - Natural and fallout radioactivity in some book samples were measured with an HPGe detector coupled with a 4096 channel analyzer in order to estimate the radiation doses to readers' eyes from books. The radiation doses to a population at large due to the presence of radioactivity in the book are not significant. Thus, no radiation hazard occurs from the radioactivity content in the book. The estimated radiation doses to eyes were found to be below the maximum permissible dose to eyes recommended by ICRP. PMID- 15650596 TI - Tales of two drugs. PMID- 15650597 TI - Toxoplasmosis retinochoroiditis and elevated intraocular pressure: a retrospective study. AB - OBJECTIVES: To report the prevalence of abnormal intraocular pressure (IOP) in patients with toxoplasmosis retinochoroiditis and to determine risk factors for such abnormality. METHODS: In a retrospective clinic-based chart review, the IOP levels of 61 patients with active retinochoroiditis were recorded. Patients were separated into groups with elevated IOP, equal IOP, and lower IOP. The time taken for normalization of IOP was also recorded. Additionally, age, gender, visual acuity, anterior chamber and vitreous inflammatory activity, presence of macular lesions, keratoprecipitates, synechiae, toxoplasmosis antibody titers, and required medical and surgical treatments were noted. The IOP in 61 patients with active retinochoroiditis were also compared with the IOP in 59 age- and gender matched control patients with unilateral anterior uveitis. RESULTS: Thirty-eight percent of patients (23/61) with active retinochoroiditis had elevated IOP > 21 mm Hg, demonstrated IOP difference > 4 mm Hg between involved and uninvolved eyes, or received on IOP-lowering medications. In the equal IOP category, 55.7% (34/61) of patients had an IOP < or = 21 mm Hg in the actively inflamed eye and had an IOP difference of < or = 3 mm Hg between the active and inactive eyes. Only 6.6% (4/61) of patients with active ocular toxoplasmosis had a decreased IOP in the affected eye. The mean IOP in patients with active retinochoroiditis was 21.2 mm Hg (SD 11.5) and 15.6 mm Hg (SD 2.9) in involved and uninvolved eyes, respectively. A statistically significant average IOP difference of 5.8 mm Hg (SD 11.6) was found between the involved and uninvolved eyes (P < 0.001 by two-tailed student T test). Of the patients with abnormal IOP, 20 patients had IOP > 21 mm Hg, 10 of whom had IOP > 30 mm Hg, 6 of whom had IOP > 40 mm Hg, and 2 of whom had IOP > 50 mm Hg. In the elevated IOP group, the average time from onset of symptoms until presentation for their IOP measurement was 13 days (median: 7 days; range: 1 to 100) with resolution of abnormal IOP occurring in 32 days (median: 28 days; range: 1 to 84 days). The average time of onset of symptoms in the normal to low IOP category was 70 days (median: 17.5 days; range: 2 to 330 days). Elevated IOP was more common in active retinochoroiditis, 23/61 (38%), when compared with anterior uveitis control group, 6/59 (10%) (odds ratio of 5.3; P < 0.001). No statistically significant predictor of elevated IOP was identified, though a trend associating anterior chamber cells with elevated IOP (P = 0.08, r = 0.25 Spearman rank correlation coefficient) was seen. CONCLUSION: Abnormal IOP is a feature in almost half of patients with active toxoplasmosis retinochoroiditis. Elevated IOP occurs in 38% and low IOP occurs in 6.6% of affected eyes. Patients with elevated IOP due to active toxoplasmosis present for earlier evaluation than patients with normal or low IOP. The IOP elevation is generally transient and concurs with the uveitic episode. Medical management of IOP is usually sufficient to treat this generally transient eye pressure rise though chronic administration of eye pressure lowering drops or glaucoma surgery may be necessary in a small proportion of patients. PMID- 15650598 TI - Psychophysical investigation of ganglion cell loss in early glaucoma. AB - PURPOSE: To evaluate ganglion cell loss in early glaucoma using a variety of psychophysical tests and to identify optimal perimetric technique(s) for detection of early glaucomatous visual function loss. METHODS: Five perimetric tests, short wavelength automated perimetry (SWAP), temporal modulation perimetry (TMP), frequency doubling technology perimetry (FDT), detection acuity perimetry (DAP), and resolution acuity perimetry (RAP) were compared in their ability to discriminate between normal individuals and patients with early glaucoma or glaucoma suspects. Comparisons were also made by their ability to produce repeatable defects. The tests examined different visual functions that are likely to be mediated by different retinal ganglion cell subpopulations, thereby permitting examination of hypotheses of ganglion cell death in early glaucoma. RESULTS: All visual field tests demonstrated high performance in separating glaucoma patients from normal individuals. SWAP, TMP, FDT, and DAP provided the greatest discrimination between normal individuals and high- and low-risk glaucoma suspects. However, SWAP, TMP, and FDT obtained better consistency across the various analysis approaches (global indices and pointwise) than DAP and RAP. Of all the test types, FDT exhibited the highest proportion of repeatable abnormal test locations, with poor confirmation rates achieved by DAP and RAP. CONCLUSION: The performance of SWAP, FDT, and TMP suggests that these test types may all be suitable for detection of early loss of visual function in glaucoma. Ganglion cell subpopulations with lower levels of redundancy and/or those with larger cell sizes offer the most parsimonious explanation for earliest ganglion cell losses occurring in glaucoma. PMID- 15650599 TI - Predicting subsequent visual field loss in glaucomatous subjects with disc hemorrhage using retinal nerve fiber layer polarimetry. AB - PURPOSE: To predict progression of visual field loss after an episode of disc hemorrhage in glaucoma patients on the basis of retinal nerve fiber layer (RNFL) GDx polarimetry measurements analyzed by wavelet-Fourier analysis (WFA). METHODS: Retrospective GDx data from 16 subjects (10 progressors and 6 non-progressors based on visual fields) obtained near the time of disc hemorrhage were analyzed to predict which patients would have visual field progression. Polarimetry scans throughout a follow-up period (31 months average) were also analyzed to compare field progression to RNFL thickness change after the hemorrhage. Mean RNFL thickness inferred from the polarimetry data at sixteen 22.5 degrees sectors at distances of 1.6, 1.7, and 1.8 disc diameters were used. Data were analyzed by applying to appropriate regions of disc hemorrhage patients a structural analysis (WFA) we had developed previously. A linear discriminant function (Fischer) was produced and a leave-one-out method using separate training and test data was used to assure validity of the results. RESULTS: Patients who subsequently progressed were successfully predicted with moderate success (sensitivity / specificity was 0.77 / 0.88 with ROC area = 0.858). A separate analysis comparing pre- and post-hemorrhage RNFL sector thickness revealed clear evidence of RNFL thinning at the inferior and superior sectors before progression of visual field. The thinning of RNFL thickness was not restricted to regions corresponding to the location of the hemorrhage. CONCLUSION: Wavelet-Fourier analysis can differentiate progressors from non-progressors with moderate accuracy. Comparison to a prior study of this same cohort emphasizes that relatively small regions must be considered (as opposed to larger quadrants) to see these significant changes in RNFL. PMID- 15650600 TI - The prevalence of glaucomatous defects with short-wavelength automated perimetry in patients with elevated intraocular pressures. AB - PURPOSE: To determine the prevalence of abnormal short-wavelength automated perimetry (SWAP) visual fields in subjects with elevated intraocular pressures (IOP) for 7 existing definitions of mild glaucomatous loss, and to explore the agreement between them. PATIENTS AND METHODS: Seven hundred and forty-four eyes of 379 subjects with an IOP > or = 22 and < or = 32 mm Hg and normal visual fields with standard automated perimetry (SAP) were tested with SWAP on 3 separate occasions, of which the second and third visual field were used for analysis. The appearance of the optic disc was not an eligibility criterion. We determined the number of visual fields classified as abnormal on 2 successive occasions by 7 existing definitions. In addition, we explored the agreement between the various definitions. RESULTS: The proportion of eyes with a glaucomatous visual field with SWAP ranged between 0% and 9.9%, depending on the criterion used to define abnormality. A pairwise comparison of the various definitions showed that several definitions classified different eyes as having an abnormal field. CONCLUSIONS: We found a large variation in the proportion of visual fields with SWAP classified as abnormal by the various definitions. More importantly, various definitions identified different individuals to have an abnormal field with SWAP. Therefore, the diagnostic accuracy and clinical significance of all definitions must be determined before SWAP is used in routine clinical care. PMID- 15650601 TI - Carotid artery elasticity and baroreflex sensitivity in patients with glaucoma. AB - PURPOSE: To investigate non-invasively the elasticity of common carotid artery and the arterial baroreflex function in patients with chronic open-angle glaucoma. PATIENTS AND METHODS: Data were obtained from patients with chronic open-angle glaucoma (n = 16) [primary open angle- and juvenile open-angle glaucoma] and from age- and sex-matched healthy control subjects (n = 18). End diastolic diameter and pulsatile distension of the common carotid artery were measured with a high-precision, automated ultrasound wall-tracking system (WTS). Spontaneous fluctuation in cardiac interval and systolic pressure were used to determine baroreflex sensitivity. RESULTS: There was no difference in heart rate and pulse pressure between patients with glaucoma and the control subjects. In the glaucoma patients both distensibility coefficient of common carotid artery and baroreflex sensitivity were reduced (two-way factorial ANOVA; P < 0.002). Stiffness (characterizing the rigidity of vessel wall) was higher in the glaucoma patients than in the controls (P = 0.002). CONCLUSIONS: Our results suggest that in patients with chronic open-angle glaucoma the arterial rigidity is significantly increased and therefore the baroreflex function is significantly decreased. These findings may have relevance in the understanding of the autonomic dysfunction and vascular dysregulation in glaucoma. PMID- 15650602 TI - Reproducibility of self-measured intraocular pressure with the phosphene tonometer in patients with ocular hypertension and early to advanced glaucoma. AB - PURPOSE: To evaluate the reproducibility of self-measured intraocular pressure with the Proview tonometer in ocular hypertensives and early to advanced glaucoma patients. PATIENTS AND METHODS: Thirty-three patients with glaucoma and three with ocular hypertension (72 eyes) were prospectively enrolled. History of cornea surgery was the only exclusion criterion. A trainer took a set of three readings first. Subsequently, the patient (after successful training) took one set of three readings and a physician took two sets of three Goldmann readings. One hour later this was repeated. RESULTS: 72% of patients could see the phosphene in both eyes, 8% could see it in one eye, and 19% could not see it in either eye. Patients who could not see the Phosphene had an average mean deviation of -10.9 compared with -3.4 for those who could see it (P = 0.01). Eyes that could not see the phosphene were excluded from further analysis. The average absolute difference between two Proview measurements one hour apart was 1.76 +/- 1.76 mm Hg (r = 0.71). The measurement error (difference between Goldmann and Proview readings) tended to remain constant between sets of measurements taken one hour apart (average error was 2.4 +/- 2.1 mm Hg, r = 0.85). No adverse event was recorded. CONCLUSIONS: Repeated self-tonometry measurements showed very good reproducibility. The systematic, reproducible error between Proview and Goldmann readings may be clinically acceptable and suggests that a baseline "calibration" should be obtained for each patient before the Proview values are used clinically. Self-administered tonometry can play a role in the clinical care of ocular hypertensive and glaucoma patients with the possible exception of very advanced cases. PMID- 15650603 TI - Incidence and management of elevated intraocular pressure after silicone oil injection. AB - PURPOSE: To determine the incidence and clinical features of chronic elevated intraocular pressure after pars plana vitrectomy and silicone oil injection for complicated retinal detachments, and to evaluate the clinical management of eyes with secondary glaucoma. METHODS: This was an observational consecutive case series of 450 eyes in 447 patients who were treated with pars plana vitrectomy and silicone oil injection. Patients who developed secondary glaucoma were treated medically with antiglaucoma medications and surgically with glaucoma drainage implants placed in an inferior quadrant. Main outcome measures were intraocular pressure, number of glaucoma medications, surgical success, and complications. RESULTS: Fifty-one of 450 eyes (11%) developed elevated intraocular pressure after pars plana vitrectomy and silicone oil injection whereas 399 eyes (89%) did not have a rise in intraocular pressure. Of the 51 eyes that developed elevated intraocular pressure, 40 (78%) were treated only with glaucoma medicines. Medical therapy reduced the intraocular pressure from a mean +/- SD of 26 +/- 13.4 mm Hg before treatment to 18 +/- 9.1 mm Hg after medical treatment (P = 0.002). The 11 of 51 eyes (22%) with elevated intraocular pressure that failed medical therapy were treated surgically with Ahmed Glaucoma Valve implantation within 12 months of silicone oil injection. In the surgical group, intraocular pressure was reduced from a mean +/- SD of 44 +/- 11.8 mm Hg before surgery to 14 +/- 4.2 mm Hg at the most recent follow-up after surgery (P < 0.001). The number of antiglaucoma medications was reduced from 3.5 +/- 0.7 before surgery to 1.2 +/- 0.5 at the most recent follow-up after surgery (P < 0.001). CONCLUSION: Chronic intraocular pressure elevation occurs in a minority (11%) of patients who are treated with silicone oil. Most of these eyes are effectively treated with antiglaucoma medications. Eyes that do not respond to medical therapy may be effectively managed with glaucoma drainage implant placement in an inferior quadrant. PMID- 15650604 TI - Needle revision of failed filtering blebs using 5-Fluorouracil and a combined ab externo and ab-interno approach. AB - PURPOSE: To evaluate a surgical technique to revise a failed filtering bleb using subconjunctival 5-Fluorouracil with a combined ab-externo and ab-interno approach. PATIENTS AND METHODS: This study is a retrospective review of the outcome of 77 consecutive bleb revisions, with greater than 6-month follow-up, performed by a single glaucoma surgeon (MW). All eyes had previously functioning filtering blebs with currently inadequately controlled intraocular pressures (IOP) prior to the bleb revisions. All surgery was performed in the operating room, using a retrobulbar injection and a microscope. Visco-elastic was injected into the anterior chamber. 5-Fluorouracil (0.1 mL; 50 mg/ml) was infiltrated around the bleb. A 30-gauge needle was used to lyse subconjunctival fibrosis and episcleral scar tissue binding down the scleral flap, and elevate the scleral flap. Through an inferior paracentesis, a cyclodialysis spatula was used to confirm and enlarge the communication with the subconjunctival space. The main outcome measurements were IOP and number of glaucoma medications. A successful outcome was defined as a 20% reduction from baseline IOP and a maximum IOP of 18 mm Hg, with or without medications, and a minimal follow-up of 6 months. RESULTS: 52% of patients achieved success after one revision with an average follow-up of 29.6 +/- 14.4 months. In successful cases, the mean IOP decreased from 22.7 +/- 4.5 mm Hg to 11.3 +/- 3.5 mm Hg and medications were reduced from an average of 2.2 +/- 1.1 to 0.4 +/- 0.7. Kaplan-Meier survival analysis calculated a success of 77% at 1 year, 68% at 2 years, and 58% at 3 years. CONCLUSIONS: In failed filtering blebs, needle revision with 5-Fluorouracil and a combined ab-externo and ab-interno approach results in high success and low complication rates. The outcome of this procedure compares favorably with previously reported revision techniques. PMID- 15650605 TI - Slit-lamp needle revision of failed filtering blebs using high-dose mitomycin C. AB - PURPOSE: To investigate the efficacy of bleb needle revision with high-dose mitomycin C in reviving failed filtering blebs after 1 year. PATIENTS AND METHODS: We reviewed the charts of 44 patients with one eye that had undergone bleb needle revision with a mixture of 0.1 mL of mitomycin (0.4 mg/mL) mixed with 0.1 mL of non-preserved 1% lidocaine. At least 12 months of follow-up were required. A successful bleb needle revision was defined as one that did not require a subsequent needling, glaucoma surgery, or medication to reach an intraocular pressure (IOP) greater than 4 mm Hg but less than 22 mm Hg. A qualified success was defined as a successful bleb needle revision that required subsequent needling or medication. RESULTS: The bleb needle revision with high dose mitomycin was a success or qualified success after 12 months in 28 patients or 64% (95% confidence interval, 50% to 78%). The baseline IOP in these patients was 26.7 +/- 8.2 mm Hg (range 15 to 48 mm Hg) using an average of 1.5 +/- 1.5 glaucoma medications. The IOP after 1 year was 13.6 +/- 4.0 (range 6 to 21 mm Hg) with an average of 0.5 +/- 0.8 medications. Of 44 patients, 17 (39%) were successes and 11 (25%) were qualified successes. CONCLUSION: Bleb needle revision with high-dose MMC was effective in reducing the IOP in 64% of eyes with a failed filtering bleb with minimal long-term complications. PMID- 15650606 TI - A cross-sectional study of Krukenberg spindles and pigmented lens striae in a predominately black population: two highly associated clinical signs of anterior segment pigment dispersal. AB - PURPOSE: To investigate the relationship of Krukenberg spindles (KS) and pigmented lens striae (PLS), clinical signs related to iris pigment dispersal and possibly glaucoma. METHODS: During a 31-month period, 5 practitioners in an urban, primary eye care setting examined consecutive patients for KS and PLS. Multiple logistic regression was used to evaluate relationships among KS, PLS, and other variables. RESULTS: Krukenberg spindles were present in 65 patients (52 females), comprised of 57 of 2647 (2.2%) blacks, 5 of 303 (1.7%) whites, 2 of 121 (1.7%) Hispanics, and 1 of 55 (1.8%) Asians. PLS were present in 64 subjects (56 females), comprised of 59 (2.2%) blacks, 3 (1.0%) whites, and 2 (3.6%) Asians. KS and PLS were coexistent in 27 subjects. Mean age +/- SD (range) of the KS and PLS subjects was 63.1 +/- 15.0 years (24-88 years) and 67.0 +/- 10.4 years (33-88 years), respectively. Mean refractive error +/- SD (range) of KS and PLS right eyes was +0.55 +/- 2.32D (-6.50 to +5.50D) and +1.34 +/- 2.18D (-6.50 to +7.25D), respectively. Controlling for other variables, PLS were highly predictive (OR = 30.2, P < 0.0001) of KS, and KS were highly predictive (OR = 29.5, P < 0.0001) of PLS. Ignoring presence or absence of PLS, increasing age (in decades) (OR = 1.60, P < 0.0001) was strongly associated with KS. Ignoring presence or absence of KS, age (OR = 1.74, P < 0.0001), female gender (OR = 2.96, P = 0.009), and increasing hyperopic refractive error (OR = 1.30, P < 0.0001) were strongly associated with PLS. CONCLUSIONS: Krukenberg spindles and PLS were strongly associated in our patient population, and the likelihood of both increased with increasing age. Female gender and increasing hyperopic refractive error were highly significant predictors of PLS. PMID- 15650607 TI - Matrix metalloproteinases and their tissue inhibitors in aqueous humor of patients with primary open-angle glaucoma, exfoliation syndrome, and exfoliation glaucoma. AB - PURPOSE: To study extracellular matrix (ECM) metabolism by matrix metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) in aqueous humor (AH) samples collected from primary open-angle glaucoma (POAG), exfoliation syndrome (EXS), and exfoliation glaucoma (EXG) in relation to samples derived from cataract control patients. MATERIALS AND METHODS: Seventy-one AH samples were collected during cataract extraction and trabeculectomy. The expression and molecular forms of MMP-2, -8, -9, -13, and -14 and tissue inhibitor of metalloproteinases-1 and -2 (TIMPs) were analyzed by Western immunoblotting. Gelatinase and collagenase activities were studied by zymography and type I collagen degradation assays, respectively. MMP-2 and TIMP-2 concentrations were measured by ELISA assays. RESULTS: By Western immunoblotting all the studied MMPs were mainly in their latent form in all diagnostic groups. Zymography demonstrated that MMP-2 represents the major gelatinase in AH. Similarly, type I collagenolytic activity was low and similar in cataract and glaucoma samples. In ELISA measurements the TIMP-2 levels were significantly elevated in glaucoma and EXS samples in comparison to cataract controls (P < 0.05). CONCLUSION: TIMP-2 is elevated in glaucomatous process over MMP-2, which support and further extend the conjuncture that the ECM accumulation rather than degradation predominates in the pathogenesis of POAG and EXG. PMID- 15650608 TI - Effects of travoprost on aqueous humor dynamics in monkeys. AB - PURPOSE: To determine the mechanism by which travoprost, a prodrug of a prostaglandin F2alpha analog, reduces intraocular pressure (IOP) in cynomolgus monkey eyes. METHODS: One eye each of 12 monkeys was treated with laser burns to the trabecular meshwork to elevate IOP. At least 4 months later (Baseline Day), IOP was measured by pneumatonometry (9:00 AM and 11:45 AM), and aqueous flow and outflow facility were determined by a fluorophotometric method. Uveoscleral outflow was calculated. Both eyes were treated with travoprost 0.004% at 9:00 AM and 5:00 PM for two days and at 9:30 AM on the third day (Treatment Day), when measurements were repeated as on Baseline Day. Statistical analyses were performed using two-tailed, paired t tests. RESULTS: On Treatment Day compared with Baseline Day, IOP in hypertensive eyes was reduced at 2.25 hours (25.8 +/- 11.2 vs 33.7 +/- 13.2 mm Hg; mean +/- standard error of the mean [SEM]; P = 0.02) and 16 hours (26.3 +/- 10.2 vs 35.1 +/- 13.6 mm Hg; P = 0.02) after treatment. The increase in uveoscleral outflow was not significant. In normotensive eyes, IOP was reduced at 2.25 hours (19.0 +/- 3.7 vs 23.0 +/- 4.0 mm Hg; P = 0.03) and 16 hours (20.7 +/- 5.4 vs 23.4 +/- 5.3 mm Hg; P = 0.01) after treatment, and uveoscleral outflow was significantly (P = 0.02) increased (1.02 +/- 0.43 vs 0.35 +/- 0.72 microL/min). CONCLUSION: Travoprost reduces IOP in normotensive monkey eyes by increasing uveoscleral outflow. The IOP reduction in hypertensive eyes is probably via the same mechanism, although the increased uveoscleral drainage did not reach statistical significance. Travoprost had no effect on aqueous flow or outflow facility. PMID- 15650609 TI - A trabecular bypass flow hypothesis. AB - PURPOSE: To introduce a hypothesis that theorizes the effect of a trabecular bypass, a channel created through the trabecular meshwork, on the facility of outflow and the intraocular pressure (IOP). METHODS: Equations that govern the pressure and circumferential flow in Schlemm's canal are established, based on the linear relationships between pressure drop and flow, the balance of flows in the canal, and a uniform leaking structure of collector channels. Two types of bypasses permitting either unidirectional or bidirectional flow are incorporated through boundary conditions to solve the equations and to derive the facility of outflow and the reduced IOP. RESULTS: In normal healthy eyes, the facility of outflow increases by 13% and 26% in the presence of a unidirectional and bidirectional bypass, respectively. The circumferential flow is significant only in the immediate quadrant to the bypass. The elevated IOP due to the abnormally high resistance in trabecular meshwork in open angle glaucoma is substantially reduced with a single bypass. Mean physiological level of IOP is attained if the bidirectional bypass is placed; slightly higher IOP is attained if a unidirectional bypass is placed. In either case, the higher the baseline IOP, the greater the reduction. The effectiveness of the bypass on IOP reduction is related to the resistances in the canal and the collector channels. Multiple bypasses can be used to further reduce the IOP. CONCLUSION: It is theoretically demonstrated that a single patent trabecular bypass can enhance the facility of outflow and reduce the IOP to physiological levels. PMID- 15650610 TI - Memantine and progressive glaucoma. PMID- 15650611 TI - Pupillary block? PMID- 15650612 TI - Penetrating trabeculectomy in topical anesthesia. PMID- 15650615 TI - Outcome of eyes undergoing trabeculectomy after intravitreal injections of triamcinolone acetonide. PMID- 15650616 TI - Treatment-treatment interactions. PMID- 15650617 TI - Complexity of medication use in the Veterans Affairs healthcare system: Part I: Outpatient use in relation to age and number of prescribers. AB - CONTEXT: Multiple medication use is associated with an increased incidence of adverse drug-drug interactions (DDIs), medication errors, noncompliance, hospitalization, and healthcare costs. Drugs acting systemically or gastrointestinally ("SG" drugs) are of particular concern because of their potential to interact. A better understanding is needed of the relationship between multiple medication use, particularly of SG drugs, and age, number of prescribers, and common drug regimens. OBJECTIVE: to determine the levels of multiple SG medication use in relation to age, number of prescribers, and common drug regimens in an outpatient population served by U.S. Veterans Integrated Service Network 15 (VISN 15). DESIGN, SETTING, AND PARTICIPANTS: cross-sectional analysis of the subset of 5,003 currently active patients from a stratified random sample of 7,000 potentially active outpatients (1,000 each from 7 sites comprising VISN 15) selected from the prescription database on a single day. MAIN OUTCOME MEASURES: number of SG drugs/patient; number and frequency of SG drug entities and regimens. RESULTS: Most patients (97%) were dispensed at least one SG drug: 80% received > or =2 SG drugs, of which 42% received 2-4 SG drugs, 24%, 5-7 SG drugs, and 14%, > or =8 SG drugs. 394 different SG drugs were dispensed, only 88 of which occurred in 1% or more of patients. A significant increase (p < 0.0001) in level of multiple medication use occurred with increasing age and number of prescribers. Proportions of patients receiving 8 or more SG drugs approximately doubled with each additional prescriber, up to 4 or more prescribers. No drug regimen containing 2 or more drugs occurred in 1% or more of patients; 71% of patients were receiving a unique drug regimen (based on specific SG drugs without regard to dose or administration schedule). CONCLUSIONS: The uniqueness of SG drug regimens suggests no single prescriber could have extensive clinical experience with even a small fraction of the drug regimens patients receive. These findings suggest that potential DDIs cannot be predicted based on occurrence of common drug regimens in a general patient population. A follow-up study (reported separately) investigated whether common drug regimens can be identified by selecting for a specific drug treatment (e.g., an antidepressant). The improved ability to predict DDIs is particularly relevant for psychiatric patients, who are at increased risk for DDIs because of greater frequency of multiple medication use. In addition, DDIs may present in this population in ways that mimic worsening of primary symptoms, which may lead to increased doses of the medication that is actually responsible for the problem, causing still more toxicity. PMID- 15650619 TI - Individual and group cognitive-behavioral therapy for psychotic disorders: a pilot investigation. AB - The present study is an uncontrolled pilot investigation of individual and group cognitive-behavioral therapy (CBT) for patients with positive symptoms of psychosis (n = 6). While previous studies have utilized either individual or group CBT for schizophrenia, the present investigation is the first to include both components for patients in the chronic phase of a psychotic illness. The results of this pilot study suggest that this approach may be useful for both positive and negative symptoms of psychosis. In addition, depression, anxiety, and hopelessness scores all decreased dramatically. The majority of the gains made during treatment were maintained over an 11-month follow-up period. A combined CBT treatment program may offer benefits in terms of delusional thinking, depression, and anxiety when used as an adjunctive treatment to medication. While the present study demonstrated positive effects from treatment, results are limited by the small sample size. PMID- 15650618 TI - Complexity of medication use in the Veterans Affairs healthcare system: Part II. Antidepressant use among younger and older outpatients. AB - CONTEXT: A previous study, described in Part I of this report, found that 71% of a sample of 5,003 general outpatients in the Veterans Affairs healthcare system were receiving a unique drug regimen (i.e., total specific drug entities regardless of dose, formulation, or administration schedule). The simplest regimens contained only one drug, while the most complex regimens exceeded 20 different drugs. The purpose of the present study was to determine if patients receiving a specific therapeutic class of medications (e.g., antidepressants) have more homogeneous drug regimens. OBJECTIVE: to examine the extent and complexity of multiple medication use in younger and older adult outpatients receiving antidepressants compared with those not receiving antidepressants. The study focused on drugs that act systemically or gastrointestinally and hence have the potential to interact. DESIGN, SETTING, AND PARTICIPANTS: Two subsets of stratified random samples of outpatients selected from prescription databases of U.S. Veterans Integrated Service Network 15. The first group involved 1,991 patients deemed to be on antidepressants (AD patients): 891 aged < 60 years and 1,100 aged > or = 60 years. The second group involved 3,732 patients who had received no antidepressants within the previous 365 days but who had a supply of at least one other current prescription (NoAD patients): 1,195 aged < 60 years and 2,535 aged > or = 60 years; 2 missing age information. MAIN OUTCOME MEASURES: number of drugs, frequency of drug regimens, level of multiple medication use including and excluding antidepressants. RESULTS: Younger AD patients received 3 more drugs than younger NoAD patients. 23.6% of younger AD patients, versus 5.9% of younger NoAD patients, received > or = 8 drugs. Older AD patients received 2 more drugs than older NoAD patients. 37.6% of older AD patients, versus 12.8% of older NoAD patients, received > or = 8 drugs. In both the AD and NoAD groups, 62% 96% of patients of all ages were receiving unique drug regimens. Each drug regimen containing 2 or more drugs occurred in fewer than 1% of patients. CONCLUSIONS: AD patients were receiving more complex drug regimens and had a higher frequency of unique drug regimens than NoAD patients, even when the results were adjusted for age group and number of prescribers. The high prevalence of unique drug combinations in all patient groups in this study indicates that clinicians in this system have only limited experience with the total effects of all of the medications their patients are receiving and thus cannot rely on experience to guard against adverse multi-drug interactions. This fact is a particular concern with psychiatric medications because adverse DDIs involving these medications can mimic psychiatric symptoms and may therefore be more difficult to detect. PMID- 15650620 TI - Infanticide in female forensic patients: the view from the evolutionary standpoint. AB - Evolutionary theory predicts that very young mothers would be more likely to kill an infant than older women, given that the younger mother has a much greater ability to "replace" the dead child through subsequent pregnancies and thus to produce offspring for the next generation. Evolutionary theory also predicts that a woman would be more likely to kill a child if the child was obviously defective, the pregnancy was the result of incest or rape, or if the mother's means of supporting the child were severely compromised. The authors hypothesized that mentally ill mothers would behave in a way that differed significantly from evolutionary expectations, i.e., that they would be more likely to kill children who were older than those killed by mothers in the general population and that the mothers themselves would be likely to be older than mothers in the general population when the murders occurred. To test this hypothesis, the authors compared infanticides (both filicides and neonaticides) committed by mentally ill mothers with those committed by mothers in the general population. They examined two samples: 1) all cases of maternal infanticide from the Mid-Hudson Forensic Psychiatric Hospital from 1978 (when the hospital began admitting female patients) through the year 2000 and 2) a general population sample from a 10-year Canadian study reported by Daly and Wilson in 1998. The authors focused on the following variables: ages of the mothers, ages of the child-victims, whether the pregnancy was the result of rape or incest, whether the child had significant behavioral or physical problems, and whether there were problems supporting the child (e.g., having no partner, poverty, mother's lack of education). The results of the analyses supported the authors' hypothesis about ages of mothers and children. The mentally ill mothers in the Mid-Hudson sample were generally older when they killed their children and the children who were killed were generally older than in the Daly and Wilson general population sample (where the majority of the cases involved neonaticide and the mothers were generally younger than 25 years of age). The three factors, poverty, low education level (or low intellectual capacity), and lack of a spouse were common in both samples. Findings concerning cultural factors, motives, and methods used will be presented in separate publications. PMID- 15650621 TI - Multiple medication use in patients seen in the veterans affairs healthcare system: so what? PMID- 15650622 TI - Therapy for the brain and gut. PMID- 15650623 TI - Contracting for safety redux. PMID- 15650624 TI - BluefishRx Prescription Writer. PMID- 15650625 TI - Barriers to initial outpatient treatment engagement following first hospitalization for a first episode of nonaffective psychosis: a descriptive case series. AB - Due to the increasingly recognized importance of adequate treatment early in the course of schizophreniform disorder and schizophrenia, this report addresses the dearth of hypothesis-generating case series describing facilitators and barriers to engagement in initial outpatient care. This case series included six single, African-American first-episode patients. Narratives describing the initial hospitalization and the first outpatient appointments in an urban community mental health setting are presented. Several barriers to outpatient treatment engagement emerged from this relatively homogenous series of first-episode patients. Apparent barriers included inadequate remission of paranoia, impaired insight, and involvement with the criminal justice system between hospital discharge and the first outpatient appointment. Good family support appeared to be an important facilitator of treatment engagement during the first several months of outpatient treatment. A variety of other potential barriers, such as involuntary status at the time of hospital discharge, are considered. Though these are preliminary findings from a small case series, further research, based at least in part on the hypotheses generated here, is warranted. Many factors, at the level of the patient, the family, and the system of care, likely affect treatment engagement early in the course of schizophreniform disorder and schizophrenia. Clinicians should give special attention to this issue when caring for first-episode patients. PMID- 15650626 TI - Detection of hematogenous tumor cell dissemination predicts tumor relapse in patients undergoing surgical resection of colorectal liver metastases. AB - OBJECTIVE: To examine the prognostic significance of disseminated tumor cells in blood and bone marrow of patients undergoing surgical resection of colorectal liver metastases. SUMMARY BACKGROUND DATA: Despite curative hepatic resection of colorectal liver metastases, a high percentage of patients develop tumor recurrence. These recurrences probably originate from disseminated tumor cells released into the circulation before or during surgery. METHODS: Thirty-seven patients with potentially curative (R0) resection of colorectal liver metastases were prospectively enrolled into the study. Preoperative bone marrow samples and preoperative, intraoperative, and postoperative blood samples were examined for disseminated tumor cells by CK20 RT-PCR. RESULTS: Tumor cells were detected in preoperative blood samples in 11 of 37 (30%) patients, in intraoperative blood samples in 17 of 37 (46%) patients, and in postoperative blood samples in 8 of 37 (22%) patients. Four of 25 (16%) patients tested positive for disseminated tumor cells in bone marrow samples. Median follow-up time for all patients was 38 months (range, 10-63 months). Multivariate analysis confirmed tumor cell detection in intraoperative blood (P = 0.009) and in bone marrow samples (P = 0.013) to be independent prognostic factors of tumor relapse. CONCLUSIONS: This is the first study demonstrating that detection of hematogenous tumor cell dissemination during hepatic resection of colorectal cancer metastases predicts tumor relapse. Detection of disseminated tumor cells may help to individualize adjuvant therapy for patients with colorectal liver metastases and to develop surgical strategies to prevent intraoperative hematogenous tumor cell shedding. PMID- 15650627 TI - Detection of hematogenous tumor cell dissemination predicts tumor relapse in patients undergoing surgical resection of colorectal liver metastases. PMID- 15650628 TI - High incidence of thrombosis of the portal venous system after laparoscopic splenectomy: a prospective study with contrast-enhanced CT scan. AB - OBJECTIVE: The aims of this prospective study were to investigate the true incidence of portal or splenic vein thrombosis (PSVT) after elective laparoscopic splenectomy using contrast-enhanced computed tomography (CT) scan, and outcome of anticoagulant therapy for PSVT. SUMMARY BACKGROUND DATA: Although rare, thrombosis of the portal venous system is considered a possible cause of death after splenectomy. The reported incidence of ultrasonographically detected PSVT after elective open splenectomy ranges from 6.3% to 10%. METHODS: Twenty-two patients underwent laparoscopic splenectomy (LS group), and 21 patients underwent open splenectomy (OS group). Preoperative and postoperative helical CT with contrast were obtained in all patients, and the extent of thrombosis was investigated. Prothrombotic disorder was also determined. RESULTS: PSVT occurred in 12 (55%) patients of the LS group, but in only 4 (19%) of the OS group. The difference was significant (P = 0.03). Clinical symptoms appeared in 4 of the 12 LS patients. Thrombosis occurred in the intrahepatic portal vein (n = 9), extrahepatic portal vein (n = 2), mesenteric veins (n = 1), proximal splenic vein (n = 4), and distal splenic vein (n = 8). Prothrombotic disorder was diagnosed in 1 patient. Anticoagulant therapy was initiated once the diagnosis was established, and complete recanalization, except for distal splenic vein, was observed without any adverse event. Patients with splenomegaly were at high risk of PSVT. CONCLUSIONS: PSVT is a more frequent complication of laparoscopic splenectomy than previously reported but can be treated safely following early detection by CT with contrast. PMID- 15650629 TI - Much remains to be learned. PMID- 15650630 TI - The physiologic effects of pneumoperitoneum in the morbidly obese. AB - OBJECTIVE: To review the physiologic effects of carbon dioxide (CO2) pneumoperitoneum in the morbidly obese. SUMMARY BACKGROUND DATA: The number of laparoscopic bariatric operations performed in the United States has increased dramatically over the past several years. Laparoscopic bariatric surgery requires abdominal insufflation with CO2 and an increase in the intraabdominal pressure up to 15 mm Hg. Many studies have demonstrated the adverse consequences of pneumoperitoneum; however, few studies have examined the physiologic effects of pneumoperitoneum in the morbidly obese. METHODS: A MEDLINE search from 1994 to 2003 was performed using the key words morbid obesity, laparoscopy, bariatric surgery, pneumoperitoneum, and gastric bypass. The authors reviewed papers evaluating the physiologic effects of pneumoperitoneum in morbidly obese subjects undergoing laparoscopy. The topics examined included alteration in acid-base balance, hemodynamics, femoral venous flow, and hepatic, renal, and cardiorespiratory function. RESULTS: Physiologically, morbidly obese patients have a higher intraabdominal pressure at 2 to 3 times that of nonobese patients. The adverse consequences of pneumoperitoneum in morbidly obese patients are similar to those observed in nonobese patients. Laparoscopy in the obese can lead to systemic absorption of CO2 and increased requirements for CO2 elimination. The increased intraabdominal pressure enhances venous stasis, reduces intraoperative portal venous blood flow, decreases intraoperative urinary output, lowers respiratory compliance, increases airway pressure, and impairs cardiac function. Intraoperative management to minimize the adverse changes include appropriate ventilatory adjustments to avoid hypercapnia and acidosis, the use of sequential compression devices to minimizes venous stasis, and optimize intravascular volume to minimize the effects of increased intraabdominal pressure on renal and cardiac function. CONCLUSIONS: Morbidly obese patients undergoing laparoscopic bariatric surgery are at risk for intraoperative complications relating to the use of CO2 pneumoperitoneum. Surgeons performing laparoscopic bariatric surgery should understand the physiologic effects of CO2 pneumoperitoneum in the morbidly obese and make appropriate intraoperative adjustments to minimize the adverse changes. PMID- 15650631 TI - Bombesin-induced gastroprotection. AB - Bombesin is an endogenous gut peptide that is prominent in the stomach. In addition to its effects on modulating acid and gut peptide secretion, recent evidence indicates that bombesin is a potent gastroprotective agent. This review article examines the ability of bombesin to prevent gastric injury. Its protective actions appear to be mediated primarily via the release of endogenous gastrin, as gastroprotection is negated by blockade of gastrin receptors. Bombesin-induced gastroprotection and gastrin release are modified by somatostatin. Immunoneutralization of endogenous somatostatin increases the ability of bombesin to prevent gastric injury by increasing gastrin release. In mechanistic studies, ablation of capsaicin-sensitive afferent neurons abolishes bombesin-induced gastroprotection while cyclo-oxygenase inhibition partially reverses this effect. Nitric oxide synthase inhibition also negates bombesin induced gastroprotection as well as the ability of bombesin to increase gastric mucosal blood flow. Taken together, the available evidence indicates that bombesin causes release of endogenous gastrin that activates sensory neurons located in the gastric mucosa. Activation of sensory neurons causes increased production of nitric oxide through activation of constitutive nitric oxide synthase, which leads to a resultant increase in gastric mucosal blood flow and renders the stomach less susceptible to damage from luminal irritants. PMID- 15650632 TI - Laparoscopic versus open subtotal gastrectomy for distal gastric cancer: five year results of a randomized prospective trial. AB - OBJECTIVE: The aim of this study was to compare technical feasibility and both early and 5-year clinical outcomes of laparoscopic-assisted and open radical subtotal gastrectomy for distal gastric cancer. SUMMARY BACKGROUND DATA: The role of laparoscopic surgery in the treatment of gastric cancer has not yet been defined, and many doubts remain about the ability to satisfy all the oncologic criteria met during conventional, open surgery. METHODS: This study was designed as a prospective, randomized clinical trial with a total of 59 patients. Twenty nine (49.1%) patients were randomized to undergo open subtotal gastrectomy (OG), while 30 (50.9%) patients were randomized to the laparoscopic group (LG). Demographics, ASA status, pTNM stage, histologic type of the tumor, number of resected lymph nodes, postoperative complications, and 5-year overall and disease free survival rates were studied to assess outcome differences between the groups. RESULTS: The demographics, preoperative data, and characteristics of the tumor were similar. The mean number of resected lymph nodes was 33.4 +/- 17.4 in the OG group and 30.0 +/- 14.9 in the LG (P = not significant). Operative mortality rates were 6.7% (2 patients) in the OG and 3.3% (1 patient) in the LG (P = not significant); morbidity rates were 27.6% and 26.7%, respectively (P = not significant). Five-year overall and disease-free survival rates were 55.7% and 54.8% and 58.9% and 57.3% in the OG and the LG, respectively (P = not significant). CONCLUSIONS: Laparoscopic radical subtotal gastrectomy for distal gastric cancer is a feasible and safe oncologic procedure with short- and long term results similar to those obtained with an open approach. Additional benefits for the LG were reduced blood loss, shorter time to resumption of oral intake, and earlier discharge from hospital. PMID- 15650633 TI - H-graft portacaval shunts versus TIPS: ten-year follow-up of a randomized trial with comparison to predicted survivals. AB - OBJECTIVE: To report long-term outcome of patients undergoing prosthetic 8-mm H graft portacaval shunts (HGPCS) or TIPS and to compare actual with predicted survival data. METHODS: A randomized trial comparing TIPS to HGPCS for bleeding varices began in 1993. Predicted survival was determined using MELD (Model for End-stage Liver Disease). RESULTS: Patients undergoing TIPS (N = 66) or HGPCS (N = 66) were very similar by Child's class and MELD scores and predicted survival. After TIPS (P = 0.01) and HGPCS (P = 0.001), actual survival was superior to predicted survival. Through 24 months, actual survival after HGPCS was superior to actual survival after TIPS (P = 0.04). Compared with TIPS, survival was superior after HGPCS for patients of Child's class A and B (P = 0.07) and with MELD scores less than 13 (P = 0.04) with follow-up at 5 to 10 years. Shunt failure was less following HGPCS (P < 0.01). CONCLUSIONS: Predicted survival data for patients undergoing TIPS or HGPCS confirms an unbiased randomization. Actual survival following TIPS or HGPCS was superior to predicted survival. Shunt failure favored HGPCS, as did survival after shunting, particularly for the first few years after shunting and for patients of Child's class A or B or with MELD scores less than 13. This trial irrefutably establishes a role for surgical shunting, particularly HGPCS. PMID- 15650634 TI - Prediction of recurrence after radical surgery for gastric cancer: a scoring system obtained from a prospective multicenter study. AB - OBJECTIVE: The aim of this prospective multicenter study was to define a scoring system for the prediction of tumor recurrence after potentially curative surgery for gastric cancer. SUMMARY BACKGROUND DATA: The estimation of the risk of recurrence in individual patient may be relevant in clinical practice, to apply adjuvant therapies after surgery, and plan an adequate follow-up program. Only a few studies, most of which were retrospective or performed on a limited number of patients, have developed a prognostic score in patients with gastric cancer. METHODS: A total of 536 patients who underwent UICC R0 resection between 1988 and 1998 at 3 surgical departments in Italy were considered. All patients were followed up using a standard protocol after discharge from the hospital. The mean follow-up period was 56 +/- 44 months, and 94 +/- 29 months for surviving patients. The scoring system was calculated on the basis of a logistic regression model, where the presence of the recurrence was the dependent variable, and clinicopathologic variables were the covariates. RESULTS: Recurrence occurred in 272 of 536 patients (50.7%). The scoring system for the prediction of the risk in individual cases gave values ranging from 1.4 to 99.9; the model distributed most cases in the extremes of the range. The risk of recurrence increased remarkably with score values; it was only 5% in patients with a score below 10, up to 95.4% in patients with a score of 91 to 100. No recurrence was observed in 43 patients with a score below 4, whereas all of the 56 patients with a score over 97 presented a recurrence. The model correctly predicted recurrence in 227 of 272 patients (sensitivity, 83.5%), whereas the absence of recurrence was correctly predicted in 214 of 264 patients (specificity, 81.1%); the overall accuracy was 82.2%. Prognostic score was clearly superior to UICC tumor stage in predicting recurrence. The high effectiveness of the score was confirmed in preliminary data of a validation study. CONCLUSIONS: The scoring system obtained with a regression model on the basis of our follow-up data is useful for defining subgroups of patients at a very low or very high risk of tumor recurrence after radical surgery for gastric cancer. Final results of the validation study are essential for a clinical application of the model. PMID- 15650635 TI - A method to attenuate pneumoperitoneum-induced reductions in splanchnic blood flow. AB - OBJECTIVE: To determine if increasing nitric oxide bioactivity by inclusion of ethyl nitrite (ENO) in the insufflation admixture would attenuate pneumoperitoneum-induced decreases in splanchnic perfusion. SUMMARY BACKGROUND DATA: Organ blood flow is reduced during pneumoperitoneum and can contribute to laparoscopy-associated morbidity and mortality. Previous attempts to control such decreases in flow have been ineffective. METHODS: Laser-Doppler flow probes were placed on the liver and right kidney of anesthetized pigs. After a baseline recording period, animals were insufflated to a final intraperitoneal pressure of 15 mm Hg. Group one received CO2 (standard practice), whereas group 2 received CO2 plus 100 ppm ENO. Insufflation was maintained for 60 minutes and then the abdomen was manually deflated; monitoring was continued for another 60 minutes. RESULTS: CO2 insufflation (n = 5) cut liver blood flow in half; liver flow remained at this level throughout the postinsufflation period. Inclusion of 100 ppm ENO (n = 6) attenuated both the acute and prolonged blood flow decreases. Statistical modeling of the data showed that, on average, liver blood flow was 14.3 U/min higher in the ENO pigs compared with the CO2 group (P = 0.0454). In contrast, neither treatment significantly altered kidney blood flow (P = 0.6215). CONCLUSION: The data indicate that ENO can effectively attenuate pneumoperitoneum induced blood flow decreases within the peritoneal cavity. The result suggests a novel therapeutic method of regulating hemodynamic changes during laparoscopic procedures. PMID- 15650636 TI - Evaluation of the learning curve in ileal pouch-anal anastomosis surgery. AB - SUMMARY BACKGROUND DATA: We define the learning curve required to attain satisfactory training in ileal pouch-anal anastomosis (IPAA) and identify possible differences in the learning curve for stapled and hand-sewn IPAA surgery. Various studies have addressed the differences in failure rate between stapled and hand-sewn IPAA, but there is no literature that evaluates the differences in attaining satisfactory training in each of these techniques. METHODS: Data were collected from 1965 patients undergoing IPAA surgery by 12 surgeons in a single center between 1983 and 2001. Using ileoanal pouch failure as the primary end point, a parametric survival model was used to adjust for case mix (patient comorbidity, preoperative diagnosis, manometric findings, and prior anal pathology). A risk-adjusted cumulative sum (CUSUM) model was used for monitoring outcomes in IPAA surgery. RESULTS: The 5-year ileal pouch survival was 95.6% (median patient follow-up of 4.2 years; range 0-19 years). Fifty percent of trainee staff demonstrated a learning curve in IPAA surgery. Having adjusted for case mix, trainee staff undertaking stapled IPAA surgery showed an improvement in the pouch failure rate following an initial training period of 23 cases versus 40 cases for senior staff. The learning curve for hand-sewn IPAA surgery was quantified only for senior staff who attained adequate results following an initial period of 31 procedures. CONCLUSIONS: The CUSUM method was a useful tool for objectively measuring performance during the learning phase of IPAA surgery. With adequate training, supervision, and monitoring, the learning curve in IPAA surgery may be reduced even further. PMID- 15650637 TI - Partial hepatectomy for metastases from noncolorectal, nonneuroendocrine carcinoma. AB - OBJECTIVE: To define perioperative and long-term outcome and prognostic factors in patients undergoing hepatectomy for liver metastases arising from noncolorectal and nonneuroendocrine (NCNN) carcinoma. SUMMARY BACKGROUND DATA: Hepatic resection is a well-established therapy for patients with liver metastases from colorectal or neuroendocrine carcinoma. However, for patients with liver metastases from other carcinomas, the value of resection is incompletely defined and still debated. METHODS: Between April 1981 and April 2002, 141 patients underwent hepatic resection for liver metastases from NCNN carcinoma. Patient demographics, tumor characteristics, treatment, and postoperative outcome were analyzed. RESULTS: Thirty-day postoperative mortality was 0% and 46 of 141 (33%) patients developed postoperative complications. The median follow up was 26 months (interquartile range [IQR]) 10-49 months); the median follow up for survivors was 35 months (IQR 11-68 months). There have been 24 actual 5-year survivors so far. The actuarial 3-year relapse-free survival rate was 30% (95% confidence interval [CI], 21-39%) with a median of 17 months. The actuarial 3-year cancer-specific survival rate was 57% (95% CI, 48-67%) with a median of 42 months. Primary tumor type and length of disease-free interval from the primary tumor were significant independent prognostic factors for relapse-free and cancer-specific survival. Margin status was significant for cancer-specific survival and showed a strong trend for relapse-free survival. CONCLUSIONS: Hepatic resection for metastases from NCNN carcinoma is safe and can offer long-term survival in selected patients. Hepatic resection should be considered if all gross disease can be removed, especially in patients with metastases from reproductive tract tumors or a disease-free interval greater than 2 years. PMID- 15650638 TI - In situ hypothermic perfusion of the liver versus standard total vascular exclusion for complex liver resection. AB - SUMMARY BACKGROUND DATA: We compare the results of liver resection performed under in situ hypothermic perfusion versus standard total vascular exclusion (TVE) of the liver <60 minutes and > or =60 minutes in terms of liver tolerance, liver and renal functions, postoperative morbidity, and mortality. The safe duration of TVE is still debated. Promising results have been reported following TVE associated with hypothermic perfusion of the liver with durations of up to several hours. The 2 techniques have not been compared so far. METHODS: The study population includes 69 consecutive liver resections under TVE <60 minutes (group TVE<60', 33 patients), > or =60 minutes (group TVE> or =60', 16 patients), and in situ hypothermic perfusion (group TVEHYOPOTH, 20 patients). Liver tolerance (peaks of transaminases), liver and kidney function (peak of bilirubin, minimum prothrombin time, and peak of creatinine), morbidity, and in-hospital mortality were compared within the 3 groups. RESULTS: The postoperative peaks of aspartate aminotransferase (IU/L) and alanine aminotransferase (IU/L) were significantly lower (P[r] < 0.05) in group TVE HYPOTH (450 +/- 298 IU/L and 390 +/- 391 IU/L) compared with the groups TVE<60' (1000 +/- 808; 853 +/- 743) and TVE> or =60' (1519 +/- 962; 1033 +/- 861). In the group TVEHYPOTH, the peaks of bilirubin (micromol/L) (84 +/- 31), creatinine (micromol/L) (75 +/- 22), and the number of complications per patient (1.2 +/- 0.9) were comparable to those of the group TVE<60' (80 +/- 111; 109 +/- 77; and 0.8 +/- 1.1 respectively) and significantly lower to those of the group TVE> or =60' (196 +/- 173; 176 +/- 176, and 2.6 +/- 1.8). In-hospital mortality rates were 1 in 33, 2 in 16, and 0 in 20 for the groups TVE<60', TVE> or =60', and TVEHYOPOTH, respectively, and were comparable. On multivariate analysis, the size of the tumor, portal vein embolization, and a planned vascular reconstruction were significantly predictive of TVE > or =60 minutes. CONCLUSIONS: Compared with standard TVE of any duration, hypothermic perfusion of the liver is associated with a better tolerance to ischemia. In addition, compared with TVE > or =60 minutes, it is associated with better postoperative liver and renal functions and a lower morbidity. Predictive factors for TVE > or =60 minutes may help to indicate hypothermic perfusion of the liver. PMID- 15650639 TI - Prognostic significance of preoperative [18-F] fluorodeoxyglucose (FDG) positron emission tomography (PET) imaging in patients with resectable soft tissue sarcomas. AB - OBJECTIVE: The objective of this study was to evaluate the prognostic significance of preoperative positron emission tomography (PET) using 2-fluoro-2 deoxy-D-glucose (FDG) by calculating the mean standardized uptake values (SUV) in patients with resectable soft tissue sarcomas (STS). SUMMARY AND BACKGROUND DATA: FDG-PET might be used as an adjunctive tool (in addition to biopsy and radiologic tomography) in the preoperative prognostic assessment of resectable STS. METHODS: A total of 74 adult patients with STS underwent preoperative FDG-PET imaging with calculation of the SUV. Clinicopathologic data and the SUV were analyzed for an association with the clinical outcome. The first and the third quartiles of the SUV distribution function were used as cutoff values (1.59 and 3.6). Survival was estimated by the Kaplan-Meier method. Univariate and multivariate analyses were performed using log-rank test and the Cox proportional hazards regression model. RESULTS: In 55 cases, STS were completely resected (follow up 40 months): 5-year recurrence-free survival rates in patients with SUV <1.59, 1.59 to <3.6, and > or =3.6 were 66%, 24%, and 11%, respectively (P = 0.0034). SUV was a predictor for overall survival (5-year rates: 84% [SUV <1.59], 45% [SUV 1.59 to <3.6], and 38% [SUV > or =3.6]; P = 0.057) and local tumor control (5-year rates: 93% [SUV <1.59], 43% [SUV 1.59 to <3.6], and 15% [SUV > or =3.6]; P = 0.0017). By multivariate analysis, SUV was found to be predictive for recurrence-free survival. The prognostic differences with respect to the SUV were associated with tumor grade (P = 0.002). CONCLUSION: The semiquantitative FDG uptake, as measured by the mean SUV on preoperative PET images in patients with resectable STS, is a useful prognostic parameter. SUV with cutoff values at the first and the third quartiles of the SUV distribution predicted overall survival, recurrence-free survival, and local tumor control. Therefore, FDG-PET can be used to improve the preoperative prognostic assessment in patients with resectable STS. PMID- 15650640 TI - Long-term results of intraoperative electron beam irradiation (IOERT) for patients with unresectable pancreatic cancer. AB - SUMMARY BACKGROUND DATA: To analyze the effects of a treatment program of intraoperative electron beam radiation therapy (IOERT) and external beam radiation therapy and chemotherapy on the outcome of patients with unresectable or locally advanced pancreatic cancer. METHODS: From 1978 to 2001, 150 patients with unresectable and nonmetastatic pancreatic cancer received IOERT combined with external beam radiation therapy and 5-fluorouracil-based chemotherapy for definitive treatment. RESULTS: The 1-, 2-, and 3-year actuarial survival rates of all 150 patients were 54%, 15%, and 7%, respectively. Median and mean survival rates were 13 and 17 months, respectively. Long-term survival has been observed in 8 patients. Five patients have survived beyond 5 years and 3 more between 3 and 4 years. There was a statistically significant correlation of survival to the diameter of treatment applicator (a surrogate for tumor size) used during IOERT. For 26 patients treated with a small-diameter applicator (5 cm or 6 cm), the 2- and 3-year actuarial survival rates were 27% and 17%, respectively. In contrast, none of the 11 patients treated with a 9-cm-diameter applicator survived beyond 18 months. Intermediate survival rates were seen for patients treated with a 7- or 8-cm-diameter applicator. Operative mortality was 0.6%, and postoperative and late complications were 20% and 15%, respectively. CONCLUSIONS: A treatment strategy employing IOERT has resulted in long-term survival in 8 of 150 patients with unresectable pancreatic cancer. Survival benefit was limited to patients with small tumors. Enrollment of selected patients with small tumors into innovative protocols employing this treatment approach is appropriate. PMID- 15650641 TI - Long-term survival following treatment of pseudomyxoma peritonei: an analysis of surgical therapy. AB - SUMMARY BACKGROUND DATA: Pseudomyxoma peritonei (PMP) is a clinical syndrome with a poorly defined natural history. Relative contributions of tumor biology, patient selection, and the extent of treatment on ultimate outcome are not well characterized. METHODS: Patients treated at the Memorial Sloan-Kettering Cancer Center between 1980 and 2002 with a diagnosis of PMP were identified. Patient characteristics, pathologic features, and details of treatment were analyzed retrospectively. RESULTS: The 97 patients included in this study underwent a mean 2.2 +/- 0.1 operations (range, 1-6). Although complete cytoreduction was achieved in 55% (53/97), disease recurred in 91% (48/53) of patients. The median disease free interval after complete cytoreduction was 24 months. The median overall survival was 9.8 years and was independently associated with low-grade pathologic subtype (P < 0.001) and the ability to achieve complete cytoreduction (P < 0.001). Ten-year survival was attained in 21% (20/97) of the patients, of which 90% (18/20) had low-grade pathologic features. At the time of death or completion of follow-up, only 12% (12/97) of the patients were disease free. CONCLUSIONS: Outcome in patients with PMP is strongly associated with tumor biology. Although improved survival is associated with low-grade pathology and tumors amenable to complete cytoreduction, recurrence of PMP is common. Treatment may be beneficial, particularly in controlling symptoms, but absolute cure, defined as a prolonged disease-free state, is uncommon. PMID- 15650642 TI - Modeling the effect of tumor size in early breast cancer. AB - SUMMARY BACKGROUND DATA: The purpose of this study was to determine the type of relationship between tumor size and mortality in early breast carcinoma. METHODS: The data was abstracted from 83,686 cases registered in the Surveillance, Epidemiology, and End Results Program of women diagnosed with primary breast carcinoma between 1988 and 1997 presenting with a T1-T2 lesion and no metastasis in whom axillary node dissection was performed: 58,070 women were node-negative (N0) and 25,616 were node-positive (N+). End point was death from any cause. Tumor size was modeled as a continuous variable by proportional hazards using a generalized additive models procedure. RESULTS: Functionally, a Gompertzian expression exp(-exp(-(size-15)/10)) provided a good fit to the effect of tumor size (in millimeters) on mortality, irrespective of nodal status. Quantitatively, for tumor size between 3 and 50 mm, the increase of crude cumulative death rate (number of observed deaths divided by the number of patients at risk) increased with size from 10% to 25% for N0 and from 20% to 40% for N+. CONCLUSIONS: The functional relationship of tumor size with mortality is concordant with current knowledge of tumor growth. However, its qualitative and quantitative independence of nodal status is in contradiction with the prevailing concept of sequential disease progression from primary tumor to regional nodes. This argues against the perception that nodal metastases are caused by the primary tumor. PMID- 15650643 TI - Predicting the risk for additional axillary metastases in patients with breast carcinoma and positive sentinel lymph node biopsy. AB - OBJECTIVE: To assess whether the risk for nonsentinel node metastases may be predicted, thus sparing a subgroup of patients with breast carcinoma and a positive sentinel lymph node (SLN) biopsy completion axillary lymph node dissection (ALND). SUMMARY BACKGROUND DATA: The SLN is the only involved axillary lymph node in the majority of the patients undergoing ALND for a positive SLN biopsy. A model to predict the status of nonsentinel axillary lymph nodes could help tailor surgical therapy to those patients most likely to benefit from completion ALND. METHODS: All the axillary sentinel and nonsentinel lymph nodes of 1228 patients were reviewed histologically and reclassified according to the current TNM classification of malignant tumors as bearing isolated tumor cells only, micrometastases, or (macro)metastases. The prevalence of metastases in nonsentinel lymph nodes was correlated to the type of SLN involvement and the size of the metastasis, the number of affected SLNs, and the prospectively collected clinicopathologic variables of the primary tumors. RESULTS: In multivariate analysis, further axillary involvement was significantly associated with the type and size of SLN metastases, the number of affected SLNs, and the occurrence of peritumoral vascular invasion in the primary tumor. A predictive model based on the characteristics most strongly associated with nonsentinel node metastases was able to identify subgroups of patients at significantly different risk for further axillary involvement. CONCLUSIONS: Patients with the most favorable combination of predictive factors still have no less than 13% risk for nonsentinel lymph node metastases and should be offered completion ALND outside of clinical trials of SLN biopsy without back-up axillary clearing. PMID- 15650644 TI - Histopathologic excision margin affects local recurrence rate: analysis of 2681 patients with melanomas < or =2 mm thick. AB - OBJECTIVE: Prospective trials have shown that 1-cm and 2-cm margins are safe for melanomas <1 mm thick and > or =1 mm thick, respectively. It is unknown whether narrower margins increase the risk of LR or mortality. SUMMARY BACKGROUND DATA: To determine the relationship between histopathologic excision margin, local recurrence (LR) and survival for patients with melanomas < or =2 mm thick. METHODS: Data were extracted from the Sydney Melanoma Unit database for all patients with cutaneous melanoma < or =2 mm thick, diagnosed up to 1996. Patients with positive excision margins or follow-up <12 months were excluded, leaving 2681 for analysis. Outcome measures were LR (recurrence <5 cm from the excision scar), in-transit recurrence, and disease-specific survival. Factors predicting LR and overall survival were tested with Cox proportional hazards analysis. RESULTS: Median follow-up was 83.8 months. LR was identified in 55 patients (median time to recurrence, 37 months). At 120 months, the actuarial LR rate was 2.9%. Five-year survival after LR was 52.8%. In multivariate analysis, only margin of excision and tumor thickness were predictive of LR (both P = 0.003). When all patients with a margin <0.8 cm in fixed tissue (corresponding to a margin of <1 cm in vivo) were excluded from analysis, margin was no longer significant in predicting LR. Thickness, ulceration, and site were predictive of survival, but margin was not (P = 0.49). CONCLUSIONS: Histopathologic margin affects the risk of LR. However, if the in vivo margin is > or =1 cm, it no longer predicts risk of LR. Patient survival is not affected by margin. PMID- 15650645 TI - Influence of metformin on glucose intolerance and muscle catabolism following severe burn injury. AB - SUMMARY BACKGROUND DATA: Hyperglycemia and accelerated muscle catabolism have been shown to adversely affect immune response and survival. The purpose of this study was to determine the effect of metformin on glucose kinetics and muscle protein metabolism in severely burned patients and assess any potential benefit of metformin in this clinical setting. METHODS: In a double-blind, randomized manner, 8 adult burn patients received metformin (850 mg every 8 hours x 7 days), while 5 burn patients received placebo. Infusions of 6,6d2 glucose, d5 phenylalanine, sequential muscle biopsies, and femoral arterial, venous blood sampling allowed determination of glucose and muscle protein kinetics. Measurements were obtained immediately prior and at the conclusion of 7 days of treatment (metformin versus placebo). All patients received enteral feeds of comparable amounts during study. RESULTS: Patients receiving metformin had a significant decrease in their plasma glucose concentration, the rate of glucose production, and an increase in glucose clearance. Metformin administration was also associated with a significant increase in the fractional synthetic rate of muscle protein and improvement in net muscle protein balance. Glucose kinetics and muscle protein metabolism were not significantly altered in the patients receiving placebo. CONCLUSIONS: Metformin attenuates hyperglycemia and increases muscle protein synthesis in severely burned patients, thereby indicating a metabolic link between hyperglycemia and muscle loss following severe injury. Therefore, therapies that improve glucose tolerance such as metformin may be of clinical value in ameliorating muscle catabolism in critically injured patients. PMID- 15650646 TI - Hepatic resection-related hypophosphatemia is of renal origin as manifested by isolated hyperphosphaturia. AB - OBJECTIVE: The objective of this study was to elucidate and define the pathophysiological mechanism(s) responsible for the clinically relevant phenomenon of posthepatic resection hypophosphatemia. SUMMARY BACKGROUND DATA: Although biochemically significant hypophosphatemia has been described after major hepatic resection, no mechanism or validated scientific explanation exists. The phenomenon is of considerable clinical relevance because numerous patients, after hepatic resection, develop significant hypophosphatemia requiring large doses of phosphate replacement to maintain metabolic homeostasis. This event has previously been empirically ascribed to amplified phosphate utilization of regenerating hepatocytes, although no rigorous data attest to this postulate. Recent data identifying a novel mechanism of phosphaturia in X-linked hypophosphatemic rickets, autosomal-dominant hypophosphatemic rickets, and oncogenic osteomalacia demonstrate that elevated levels of novel circulating phosphaturic factors such as fibroblast growth factor 23 (FGF-23) and PHEX are responsible for phosphate wasting. We hypothesize that posthepatectomy hypophosphatemia reflects a derangement of normal hepatorenal messaging and is the result of a disruption of renal phosphate handling consequent on aberrations in the metabolism of an as yet unrecognized chemical messenger(s) responsible for tubular phosphate homeostasis. This postulate has not previously been proposed or examined. METHODS: Twenty patients undergoing hepatic resection were studied prospectively with respect to serum phosphate, phosphate requirements, as well as renal phosphate handling. Fractional excretion of phosphate was calculated on a daily basis. To confirm the relationship between phosphate loss and a circulating renal-targeted messenger, the plasma levels of the circulating phosphaturic factor FGF-23 were measured using a c-terminal assay both pre- and postoperatively. RESULTS: All patients developed hypophosphatemia with a nadir on postoperative day 2 (average drop of 47% despite phosphate administration). This phenomenon was associated with hyperphosphaturia (mean +/- standard error) with high fractional excretion of phosphate. A consistent change in FGF-23 was not identified. CONCLUSION: Hypophosphatemia after hepatic resection is a frequent occurrence. Transient isolated hyperphosphaturia and not increased phosphate utilization is the predominant cause of this phenomenon, although the identity of the agent involved remains to be identified. PMID- 15650647 TI - Increased susceptibility of glutamine-depleted monocytes to fever-range hyperthermia: the role of 70-kDa heat shock protein. AB - OBJECTIVE: This study investigates the effect of fever-range hyperthermia on Gln starving monocytes and the role of the 70-kDa heat shock protein Hsp70. SUMMARY BACKGROUND DATA: Fever is a protective acute-phase response to infection. However, in critically ill patients, the harmful effects of fever seem to be predominant. Critical illness is frequently associated with reduced plasma glutamine (Gln) levels, which contribute to the immune suppression in these patients due to impaired monocyte function. METHODS: Isolated monocytes were suspended in Gln-depleted medium and exposed to 41 degrees C. Cell survival was determined by an MTT-based assay, and phagocytosis of Escherichia coli was measured by flow cytometry. Expression of Hsp70 was determined by Western blot. RESULTS: Hyperthermia for 300 minutes strongly decreased the viability of Gln depleted monocytes (85%), whereas it had only a moderate effect on Gln-supplied cells (45%, P < 0.05). Shorter treatments (45 minutes) of Gln-starving monocytes had almost no effects on viability but decreased the phagocytosis activity by 30.8%. In addition, the expression of Hsp70 was inhibited almost completely. CONCLUSION: These data show that Gln-starving monocytes have a reduced thermoresistance. This suggests that elevated body temperature damages monocytes in critically ill patients with reduced plasma Gln-levels possibly via an inhibition of the cytoprotective protein Hsp70. PMID- 15650648 TI - Carotid endarterectomy remains the standard of care, even in high-risk surgical patients. AB - OBJECTIVE: This study tested the hypothesis that high-risk patients can undergo carotid endarterectomy without associated increased risk of stroke, transient ischemic attack (TIA), or death. SUMMARY BACKGROUND DATA: Carotid endarterectomy (CEA) has clearly been shown to be effective in reducing the risk of stroke in selected symptomatic and asymptomatic patients with extracranial carotid stenosis. However, recently, carotid angioplasty with stenting (CAS) has been suggested as an alternative treatment in high-risk surgical patients. METHODS: Medical records for consecutive patients who underwent CEA from 1996 to 2001 were reviewed for demographics, medical history, and hospital course. High-risk patients were defined as those experiencing a myocardial infarction (MI) or an exacerbation of congestive heart failure (CHF) within 4 weeks before CEA; unstable angina; steroid-dependent chronic obstructive pulmonary disease (COPD); prior ipsilateral CEA, neck dissection or irradiation; high carotid bifurcation; and those with combined cardiac-carotid procedures. Poor postoperative outcome was defined as stroke, TIA, or death within 30 days. Univariate, multivariate, and Kaplan-Meier analysis were used as appropriate. RESULTS: Four hundred twenty nine patients underwent 499 CEAs, of which 84 (17%) were considered high risk. The overall stroke-death rate among all patients was 2.8%. A total of 11 postoperative strokes (2.2%), 7 TIAs (1.4%), and 3 deaths (0.6%) occurred within 30 days after surgery. There was no difference in 30-day poor outcome between high- and low-risk patients (4.8% vs. 4.1%, P = 0.77). When these risk factors were assessed independently, those with recent MI were at higher risk for poor outcome (odds ratio [OR], 13.3; 95% confidence interval [CI], 2.2-82.0; P = 0.03). Multivariate analysis also revealed that a history of contralateral stroke or TIA conferred an increased risk of poor outcome (OR, 3.0; 95% CI, 1.1-8.4; P = 0.02), whereas use of preoperative angiotensin-converting enzyme inhibitors was associated with reduced risk (OR, 0.36; 95% CI, 0.11-1.0; P = 0.05), as was a history of hyperlipidemia (OR, 0.33; 95% CI, 0.13-0.87; P = 0.03). By log-rank analysis, 12-month survival was significantly worse in the high-risk group as compared with the low-risk (96% vs. 91%, P = 0.03). CONCLUSIONS: Patients considered a surgical high risk can undergo CEA without any worse outcome compared with those patients deemed low risk. The benefit of CAS will likely be marginal, and only controlled clinical trials will be able to determine if certain subgroups demonstrate improved outcome with CAS. Carotid endarterectomy remains the standard of care, even in high-risk surgical patients. PMID- 15650649 TI - Virtual reality simulation for the operating room: proficiency-based training as a paradigm shift in surgical skills training. AB - SUMMARY BACKGROUND DATA: To inform surgeons about the practical issues to be considered for successful integration of virtual reality simulation into a surgical training program. The learning and practice of minimally invasive surgery (MIS) makes unique demands on surgical training programs. A decade ago Satava proposed virtual reality (VR) surgical simulation as a solution for this problem. Only recently have robust scientific studies supported that vision METHODS: A review of the surgical education, human-factor, and psychology literature to identify important factors which will impinge on the successful integration of VR training into a surgical training program. RESULTS: VR is more likely to be successful if it is systematically integrated into a well-thought out education and training program which objectively assesses technical skills improvement proximate to the learning experience. Validated performance metrics should be relevant to the surgical task being trained but in general will require trainees to reach an objectively determined proficiency criterion, based on tightly defined metrics and perform at this level consistently. VR training is more likely to be successful if the training schedule takes place on an interval basis rather than massed into a short period of extensive practice. High-fidelity VR simulations will confer the greatest skills transfer to the in vivo surgical situation, but less expensive VR trainers will also lead to considerably improved skills generalizations. CONCLUSIONS: VR for improved performance of MIS is now a reality. However, VR is only a training tool that must be thoughtfully introduced into a surgical training curriculum for it to successfully improve surgical technical skills. PMID- 15650650 TI - Predictors of surgery resident satisfaction with teaching by attendings: a national survey. AB - OBJECTIVE: To identify factors that predict fourth- and fifth-year surgical resident satisfaction of attending teaching quality. SUMMARY BACKGROUND DATA: With the training of surgical residents undergoing major changes, a key issue facing surgical educators is whether high-quality surgeons can still be produced. Innovative techniques (eg, computer simulation surgery) are being developed to substitute partially for conventional teaching methods. However, an aspect of training that cannot be so easily replaced is the faculty-resident interaction. This study investigates resident perceptions of attending teaching quality and the factors associated with this faculty-resident interaction to identify predictors of resident educational satisfaction. METHODS: A national survey of clinical fourth- and fifth-year surgery residents in 125 academically affiliated general surgery training programs was performed. The survey contained 67 questions and addressed demographics, hospital, and service characteristics, as well as surgery, education, and clinical care-related factors. Univariate analyses were performed to describe the characteristics of the sample; multivariate analyses were performed to evaluate the factors associated with resident educational satisfaction. RESULTS: The response rate was 61.5% (n = 756). Average age was 32 years; most were male (79%), white (72%), and married (69%); 42% had children. Ninety-five percent of respondents graduated from U.S. medical schools, and the average debt was $80,307. Of 20 potentially mutable factors, 6 variables had positive associations with resident education satisfaction and 7 had negative associations. Positive factors included the resident being the operating surgeon in major surgeries, substantial citing of evidence-based literature by the attending, attending physicians giving spontaneous or unplanned presentations, increasing the continuity of care, clinical teaching aimed at the chief resident level, and having clinical decisions made together by both the attending and resident. There were 7 negative factors such as overly supervising in surgery, being interrupted so much that teaching was ineffective, and attending physicians being rushed and/or eager to finish rounds. CONCLUSION: This study identifies several factors that were associated with resident educational satisfaction. It offers the perspective of the learners (ie, residents) and, importantly, highlights mutable factors that surgery faculty (and departments) may consider changing to improve surgery resident education and satisfaction. Improving such satisfaction may help to produce a better product. PMID- 15650651 TI - The NIH criteria for parathyroidectomy in asymptomatic primary hyperparathyroidism: are they too limited? PMID- 15650652 TI - Influence of transfusions on perioperative and long-term outcome in patients following hepatic resection for colorectal metastases. PMID- 15650653 TI - Differences in completion of screening logs between Europe and the United States in an emergency phase III trial resulting from HIPAA requirements. PMID- 15650654 TI - Laparoscopic Roux-en-Y gastric bypass, but not rebanding, should be proposed as a rescue procedure for patients with failed laparoscopic gastric banding. PMID- 15650655 TI - Evaluating obesity before surgery. PMID- 15650657 TI - Arthroscopic findings in the knee in nail-patella syndrome: a case report. AB - Nail-patella syndrome is a rare genetic disorder that can cause significant morbidity in several organs, including the musculoskeletal system. This case report describes our arthroscopic findings in the knees of an adolescent with bilateral hypoplastic and subluxed patellae. Our findings suggest that early intervention should be considered whenever this syndrome is recognized. This may modify some of the long-term joint problems seen in adults with nail-patella syndrome. PMID- 15650660 TI - Vascularity of the hip labrum: a cadaveric investigation. AB - PURPOSE: The purpose of this study was to define the vascularity of the hip labrum and to identify regional differences in vascular penetration that may have implications for the healing potential of this structure. TYPE OF STUDY: Injection study of human cadavers to investigate the vascularity of the hip labrum. METHODS: Twelve hips from 6 human frozen cadavers devoid of severe articular pathology were used. High-resolution surface-coil magnetic resonance imaging (MRI) was performed in both sagittal and coronal planes to better define anatomic planes in 2 dimensions and to correlate Spalteholz sections with the surrounding joint and labral anatomy. Each pelvis was injected with intra arterial India Ink and frozen; 3-mm sagittal or coronal sections were then cut and processed using a modified Spalteholz technique, yielding anatomic zones of the labrum. Six specimens were cut in the sagittal plane and 6 were cut in the coronal plane. Specimens were examined at x10 magnification with transillumination. Anatomic zones with regional variations in vascularity were defined. RESULTS: The anterior and superior aspects of the labrum showed degenerative changes on MRI and under direct manual transillumination in 75% of specimens. Overall, there was a relatively poor vascular supply to the labrum; however, there were regional differences between anatomic zones. Zone I (capsular contribution) had significantly more vascularity than zone II (articular side) (P < .01). Zone IA (the portion of the zone not attached to bone) showed the most consistent source of vessels across all specimens (smallest variation between specimens); however, zone IB (the portion of the zone attached to bony acetabulum) had the greatest overall mean vascularity score. These differences were not statistically significant. Furthermore, vascularity patterns were not significantly different in the anterior, superior, posterior, and inferior labral regions, nor were they different in torn versus intact specimens. CONCLUSIONS: The cadaveric specimens evaluated in this study had a relatively avascular hip labrum. However, the increased vascularity seen in zone I (capsular side) may have implications for treatment, similar to that described in the meniscus of the knee. CLINICAL RELEVANCE: A better understanding of the vascularity of the hip labrum will guide treatment of labral pathology and may have implications for the healing potential of this structure. Labral tears occurring in the vascular zone may be amenable to arthroscopic repair rather than debridement. PMID- 15650662 TI - Variation in anterior cruciate ligament scar pattern: does the scar pattern affect anterior laxity in anterior cruciate ligament-deficient knees? AB - PURPOSE: This prospective study documented patterns of scar formation after anterior cruciate ligament (ACL) rupture in order to test the hypothesis that ACL remnants can contribute to anterior knee passive motion limits tested with a ligament arthrometer. TYPE OF STUDY: Prospective cohort study. METHODS: Forty eight consecutive patients undergoing ACL reconstructions had intra-operative ligament arthrometry testing before and immediately after preparation of the notch and debridement of the ACL remnant. Patterns of scar formation were compared with changes in knee laxity after debridement of the ACL remnant. RESULTS: Eighteen patients (38%) had ligament scarring to the posterior cruciate ligament (group 1). Four patients (8%) had scar tissue that appeared to extend from the ACL fibers to the roof of the notch (group 2). Six ACL remnants (12%) appeared to have healed to the lateral wall of the notch or the medial aspect of the lateral femoral condyle in a position anterior and distal to the ACL anatomic footprint (group 3). In 20 patients (42%), there was no identifiable ligament tissue remaining (group 4). Changes in anterior laxity were associated with the specific pattern of scar formation within the notch. The greatest increase in anterior laxity after debridement was observed in knees in which the injured ACL had an aberrant reattachment to the femur: group 1 (mean, -1.3 mm; P < .01), group 2 (mean, -3.4 mm; P < .05), and group 3 (mean, -4.3 mm; P < .05). In group 4, the change in knee laxity was not significant (mean, 0.2 mm; 95% CI, -0.29 to 0.74 mm). Overall, 14 of 48 knees (29%) loosened more than 2 mm after ACL resection ( P < .01). CONCLUSIONS: Resection of the ACL scar resulted in a measurable increase in passive anterior laxity in a subset of ACL-deficient knees. This increase in anterior laxity occurred in patients whose ligament healed to the femur, effectively crossing the joint. When performing arthroscopy without reconstruction in ACL-injured knees, we recommend caution in resecting the torn ACL or scar tissue because removal of this tissue contributed to increased anterior laxity in some ACL-deficient knees. LEVEL OF EVIDENCE: Level II, diagnostic study of consecutive patients. PMID- 15650661 TI - Arthroscopic treatment of solitary benign intra-articular lesions of the knee that cause mechanical symptoms. AB - PURPOSE: Most cases of knee locking and giving-way are caused by meniscal tears, loose bodies, or chondral lesions. Intra-articular benign tumors or tumor-like lesions can present with symptoms that resemble acute mechanical derangement. From a database of 2,200 arthroscopic interventions, 19 cases of intra-articular masses that presented with catching and locking symptoms in the knee were retrospectively analyzed. TYPE OF STUDY: Case series. METHODS: Our review revealed 33 patients with benign intra-articular masses in the knee joint. Nineteen of these individuals had sought medical attention for mechanical symptoms of catching or locking. The other 14 patients had a variety of symptoms including pain, swelling, and limitation of knee motion, but did not have mechanical symptoms. Age, sex, history of trauma, knee pain and effusion, medical illnesses, physical examination, arthroscopic findings, and pathologic findings were noted. RESULTS: The average age of the 6 male and 13 female patients was 37.5 years (range, 18 to 58 years). Preoperative magnetic resonance imaging confirmed the diagnosis in 7 cases and 12 cases were diagnosed during knee arthroscopy. The mean follow-up time after surgery was 52.5 months (range, 6 to 120 months). Pathologic examination of the lesions revealed 15 cases of localized pigmented villonodular synovitis (79%), 1 lipoma arborescens (5%), 1 pseudocyst (5%), and 2 nonspecific synovial masses (11%). None of the lesions showed malignant transformation. CONCLUSIONS: Solitary benign intra-articular lesions should be considered a rare cause of mechanical knee symptoms. Localized pigmented villonodular synovitis originating from the extensor mechanism or fat pad is the most common solitary intra-articular mass lesion in the knee and usually arises in the patellofemoral compartment. Recurrence has not occurred in our series, which includes 14 of 19 patients with greater than 24 months follow up. LEVEL OF EVIDENCE: Level IV. PMID- 15650663 TI - Cross-pin femoral fixation versus metal interference screw fixation in anterior cruciate ligament reconstruction with hamstring tendons: results of a controlled prospective randomized study with 2-year follow-up. AB - PURPOSE: To determine if there is a different clinical outcome after cross-pin versus interference screw fixation in anterior cruciate ligament (ACL) reconstruction with hamstring tendons. TYPE OF STUDY: Prospective randomized clinical follow-up study. METHODS: Sixty-two patients were randomized into either TransFix cross-pin (Arthrex, Naples, FL) (TransFix group, n = 31) or metal interference screw femoral fixation (screw group, n = 31) in ACL reconstruction with hamstring tendons. The distal fixations were an AO screw with spiked washer post and metal interference screw. The evaluation methods were clinical examination, CA 4000 laxity (OSI, Hayward, CA), and Lido isokinetic muscle torque measurements (Lido MultiJoint II; Loredon, West Sacramento, CA) as well as International Knee Documentation Committee (IKDC), Tegner activity level, Lysholm knee, and Kujala patellofemoral scores. RESULTS: There were no differences between the study groups preoperatively. For the 2-year follow-up, 26 patients of the TransFix group and 30 patients of the screw group were available (90%). The evaluation methods disclosed no statistical differences between the groups at the 1- or 2-year follow-up examinations. At the 2-year follow-up, 22 (85%) of the TransFix and 22 (73%) of the screw group patients were in the IKDC A or B categories. There were significantly more additional procedures postoperatively in the TransFix group owing to the removal of the tibial fixation post hardware in 15 (48%) cases. CONCLUSIONS: There were no statistically or clinically relevant differences in the results 1 or 2 years postoperatively and both techniques seemed to improve patients' performance. LEVEL OF EVIDENCE: Level I. PMID- 15650664 TI - A prospective randomized comparison of bone-patellar tendon-bone and hamstring grafts for anterior cruciate ligament reconstruction. AB - PURPOSE: The aim of this study was to compare the results after arthroscopic anterior cruciate ligament (ACL) reconstruction using central-third, bone patellar tendon-bone (BPTB group), 3-strand semitendinosus (ST group), or 4 strand semitendinosus/gracilis (ST/G group) autografts. TYPE OF STUDY: Prospective randomized trial. METHODS: A randomized series of 134 patients, all with unilateral ACL rupture was included in the study. In all 3 groups, interference screw fixation of the graft was used at both ends and 125 of 134 (93%) of the patients returned for the follow-up examination after 26 months (range, 20 to 43 months). The preoperative assessments in all 3 groups were similar in terms of gender, Tegner activity level, Lysholm score, KT-1000 measurements, 1-leg hop test, and the knee-walking test. RESULTS: At follow-up, the knee-walking test was significantly worse in the BPTB group than in the ST group (P = .0004) and ST/G group (P < .0001). Furthermore, the knee-walking test was significantly worse at follow-up than preoperatively in the BPTB group (P < .0001). The corresponding findings were not made in the other 2 groups. A significant reduction in knee laxity and an increase in activity level compared with the preoperative assessments were found in all 3 groups, without any significant differences between the groups. CONCLUSIONS: Two years after ACL reconstruction, the use of ST and ST/G autografts rendered significantly less discomfort during the knee-walking test than the use of BPTB autografts. However, in terms of functional outcome and knee laxity, the groups displayed no significant differences. LEVEL OF EVIDENCE: Level I. PMID- 15650665 TI - Biomechanical comparison of bioabsorbable sutureless screw anchor versus suture anchor fixation for rotator cuff repair. AB - PURPOSE: Anchors that directly fix the rotator cuff to bone without using sutures may simplify the challenges associated with arthroscopic rotator cuff repair. This study compared a bioabsorbable screw fixation device to a standard suture anchor fixation method. TYPE OF STUDY: Randomized trial. METHODS: A poly-L lactide acid (PLLA), bioabsorbable screw anchor was compared with a metallic suture anchor loaded with No. 2 braided polyester suture in 12-week-old fresh frozen bovine shoulders. Twelve specimens were randomly assigned to the 2 repair groups (6 per group). A 1 x 2-cm defect was created at the insertion site of the infraspinatus tendon. Two anchors were implanted 1 cm apart in the anatomic insertional area of the infraspinatus tendon. After preconditioning to 10 N, each construct was cycled between 10 and 180 N for up to a maximum of 2,500 cycles at a rate of 33 mm/second using a materials testing machine. The number of cycles to 5- and 10-mm gap formation at the repair site and mode of failure were recorded. Clinically, 10-mm gap formation defines complete fixation failure. Data were evaluated using an analysis of variance with significance set at P < .05. RESULTS: Mattress suture fixation had significantly higher number of cycles to 10 mm failure compared with the PLLA device (P = .015). Failure occurred by tissue pullout in half the specimens and by device failure in the other half, without differences in failure modalities between groups. CONCLUSIONS: This anchor provides low-profile fixation of the rotator cuff and eliminates suture placement and knot tying during arthroscopic and mini-open cuff repair. However, these data suggest that this anchor may not perform adequately under cyclic loading conditions in patients during postoperative rehabilitation. CLINICAL RELEVANCE: This bioabsorbable anchor may not provide sufficient fixation for rotator cuff repair in humans. PMID- 15650666 TI - Arthroscopic rotator cuff repair in patients older than 60 years. AB - PURPOSE: The purpose of this study was to analyze results by age, tear size, and type of suture repair (margin convergence by side-to-side suture, direct tendon to-bone repair with suture anchor, or side-to-side plus suture anchor) of arthroscopic rotator cuff repair in patients older than 60 years (average, 67.7 years). TYPE OF STUDY: Retrospective case series. METHODS: From January 1999 to January 2001, 64 patients (64 shoulders) older than 60 years had an arthroscopic rotator cuff repair performed by the same surgical team. Fifty-four patients (84.3%) were available for follow-up (average, 27 months); 54 shoulders were evaluated using the UCLA rating scale (0-35 points). Tears were categorized according to size. The pattern of tears determined the repair technique: crescent shaped tears were repaired in a direct tendon-to-bone fashion and U-shaped tears by margin convergence with or without suture anchor. RESULTS: The mean of postoperative UCLA score improved from 10.4 (range, 8.2-20 points) to 30.5 (range, 21-35 points) (P < .0001). Sixteen patients (29.6%) had excellent results (34-35 points), 28 (51.8%) had good results (28-33 points), 10 (18.5%) had fair results (21-27 points), and none had poor results. There were no statistical differences between postoperative UCLA score and groups by age (P = .53), suture technique (P = .06), and tear size (P = .5). The correlation between technique of repair, age, and postoperative score has not shown statistical differences between margin convergence with or without suture anchor and direct tendon-to bone repair with suture anchor in the 61- to 65-year-old patients. Patients older than 65 years achieved the best results with margin convergence by side-to-side suture. CONCLUSIONS: Arthroscopic rotator cuff repair in the elderly achieves satisfactory results in a large percentage of cases regardless of the patient's age, tear size, and type of suture repair. These results are comparable to those of traditional open repair. We believe that arthroscopic evaluation allows an accurate assessment of rotator cuff tear configuration resulting in a more precise repair. LEVEL OF EVIDENCE: Level IV, case series (no, or historical, control group). PMID- 15650667 TI - Prospective randomized clinical trial comparing the effectiveness of immediate arthroscopic stabilization versus immobilization and rehabilitation in first traumatic anterior dislocations of the shoulder: long-term evaluation. AB - PURPOSE: To report the long-term results of a prospective randomized clinical trial comparing the effectiveness of immediate arthroscopic stabilization versus immobilization and rehabilitation after a first traumatic anterior dislocation of the shoulder. TYPE OF STUDY: Randomized clinical trial. METHODS: Forty subjects younger than 30 years with a first traumatic anterior shoulder dislocation were randomized to receive immediate anterior stabilization plus rehabilitation or immobilization followed by rehabilitation. Patients completed the American Shoulder and Elbow Surgeons (ASES), Disabilities of the Arm, Shoulder and Hand (DASH), and the Western Ontario Shoulder Instability Index (WOSI) questionnaires. RESULTS: At an average follow-up of 75 months, there was a significant difference in the rate of redislocation between the groups but no statistical significant difference in shoulder function with the ASES or the DASH. The mean difference between the 2 groups with the WOSI estimates a small, but clinically significant difference. CONCLUSIONS: It is recommended that immediate arthroscopic stabilization is the treatment of choice in a subset of patients who are younger than 30 years and are higher level athletes, and the timing for surgery is good or their sport is risky, i.e., rugby, football, kayaking, rock climbing. LEVEL OF EVIDENCE: Level II. PMID- 15650668 TI - Assessment of clavicular translation after arthroscopic Mumford procedure: direct versus indirect resection--a cadaveric study. AB - PURPOSE: To compare the horizontal stability of the distal clavicle following arthroscopic resection of its lateral end by direct and indirect techniques. TYPE OF STUDY: Biomechanical test of cadaveric specimens. METHODS: We performed arthroscopic distal clavicle resection on 12 fresh-frozen human cadaveric shoulders using direct (group 1, n = 6) or indirect (group 2, n = 6) approaches. In both groups 5 mm of distal clavicle were resected using an arthroscopic burr. The specimens were mounted on a materials testing device that allowed translation of the clavicle along the anteroposterior axis. The degree of posterior translation was measured from maximum anterior displacement of the clavicle. RESULTS: Mean posterior translation was 19.4 mm (range, 18 to 23 mm; SD, 2.2) and 21.3 mm (range, 18 to 25 mm; SD, 3.1) for groups 1 and 2, respectively. This difference was not statistically significant ( P = .27). CONCLUSIONS: This study suggests that there is no significant difference in anteroposterior stability of the clavicle following arthroscopic distal clavicle resection with either a direct or indirect approach. CLINICAL RELEVANCE: Clinically, this study addresses concerns about increased potential instability associated with the indirect technique of distal clavicle resection. From a biomechanical standpoint, based on this study, there is no concern for increased instability with the indirect technique of distal clavicle resection compared to a direct technique. PMID- 15650669 TI - Arthroscopic knot tying: the role of instrumentation in achieving knot security. AB - PURPOSE: To determine the role of arthroscopic knot-tying instrumentation in achieving knot security. TYPE OF STUDY: Biomechanical study. METHODS: The security achieved by 3 different commercially available knot-tying instruments using No. 2 braided suture and a standardized knot configuration was compared. Instrumentation included (1) a single-hole knot pusher, (2) a cannulated double diameter knot pusher, and (3) a mechanical end-splitting knot tightener. Hand tied knots served as controls. Testing included both cyclic loading and load to failure parameters using the Bionix 858 (MTS Inc, Eden Prairie, MN) and Instron Mini 44 (Instron, Canton, MA) materials testing systems, respectively. An experienced arthroscopic shoulder surgeon and a less experienced junior-level orthopaedic resident surgeon performed tying. RESULTS: Knot displacement after cyclic loading was smallest for the mechanical end-splitting knot tightener (average, 0.054 mm) and did not differ statistically from hand-tied knots (average, 0.058 mm). Knots tied with this device also withstood the greatest load to failure (average, 126.0 N) and were statistically similar to hand-tied knots (average, 134.1 N). The single-hole pusher had an average cyclic loading displacement of 0.095 mm and failed at 108.6 N. The cannulated double-diameter pushers had an average cyclic loading displacement of 0.106 mm and failed at 93.1 N. Both were statistically less secure than hand-tied knots or those tied with the end-splitting tightener. Investigator experience did not affect knot security with those tied by the mechanical end-splitting tightener or hand-tied knots. CONCLUSIONS: The end-splitting tightener provided the most secure arthroscopic knots that were statistically as secure as those tied by hand for both investigators. The knot-tying instrument used in arthroscopic shoulder surgery may be an important consideration with potential clinical implications based on knot security. CLINICAL RELEVANCE: The specific knot tying instrument that is selected for use in arthroscopic shoulder surgery may play an important role in the degree of ultimate knot security achieved in suture repair. PMID- 15650670 TI - Accuracy of anterior intra-articular injection of the glenohumeral joint. AB - PURPOSE: Intra-articular glenohumeral injections are an important part of orthopaedic practices, and the therapeutic benefit and diagnostic information of certain injections is based on the premise of the injection reaching its desired target. This study assessed the accuracy of an anterior intra-articular injection in awake subjects without radiologic assistance. TYPE OF STUDY: Case control study. METHODS: Forty-one patients scheduled for magnetic resonance imaging arthrography underwent anterior placement of a spinal needle using a location just lateral to the coracoid as the anterior landmark for injection, without radiographic assistance. After the needle was placed and clinically estimated to be intra-articular, 1 mL of gadolinium was injected into the joint to determine accuracy of position. The presence of intra-articular contrast was judged as an accurate injection. RESULTS: Only 26.8% (11 of 41) of injections placed anteriorly were actually intra-articular. The remaining were extra-articular, the most common location of error being either too medial or too superficial in the deltoid muscle. CONCLUSIONS: Based on our cadaveric study, we believed that an unassisted anterior injection to the glenohumeral joint would be accurately placed. However, this study shows that without some form of radiologic guidance, it is unlikely that an anteriorly placed intra-articular glenohumeral injection will be accurately placed in awake patients, and we do not recommend this technique. LEVEL OF EVIDENCE: Level IV. PMID- 15650671 TI - Pullout strength of knotless suture anchors. AB - PURPOSE: Suture anchors are used consistently for repairs of soft tissues, especially around the glenohumeral joint. These anchors can be used either arthroscopically or in an open procedure to anatomically restore the labrum and capsular tissues to the glenoid after avulsion injuries (Bankart lesion). The purpose of this study was to analyze the pullout strength of a new knotless suture anchor (Mitek Knotless Suture Anchor; Mitek, Norwood, MA) compared with 2 commercially available suture anchors that require knots to be tied (Mitek Panalok 3.5-mm Anchor and Mitek GII Quick Anchor). TYPE OF STUDY: Randomized cadaveric study. METHODS: Three groups of 10 anchors were tested on 15 fresh frozen cadaveric glenoids. Two anchors were affixed to the anterior glenoid in subchondral bone, 1 each from 2 groups. In this way, the variance of bone density among groups was minimized. The anchors requiring knots were fixed to the glenoid and tied to a ring using a Duncan knot with 3 half-hitches alternating posts. The Knotless Anchor was looped through the ring and anchored into the glenoid as described by the manufacturer. All constructs were then tested for tensile strength on an Instron machine (Canton, MA) using a crosshead speed of 200 mm/min. Ultimate failure was defined as complete failure of the construct (either suture breakage or anchor pullout). Data were then analyzed for statistical significance using analysis of variance analysis among the 3 groups, and a 2 tailed t test for statistical significance among groups. RESULTS: The average failure under tensile load for the GII, Panalok, and Knotless Anchors were 471.5 N, 432.8 N, and 650.0 N, respectively. Statistical analysis showed a statistical difference between the Knotless Anchor and the GII and Panalok sutures ( P = .02). Two-tailed t tests between the Knotless Anchor and the GII or Panalok Anchors were also significant (P = .02 and P = .02, respectively). Observations included a large standard deviation within groups. This is thought to result from the variation in bone density because markedly lower tensile loads were recorded for those anchors that pulled out from the bone before suture failure. CONCLUSIONS: The Knotless Suture Anchor is a statistically stronger construct with respect to tensile loads. It appears to be a viable option for any type of soft-tissue repair around the glenoid. CLINICAL RELEVANCE: Because the knot in suture repair is consistently the weakest point in the construct and because of the difficulty in tying knots arthroscopically, the Knotless Suture Anchor appears to be a stronger and easier method for both arthroscopic and open Bankart repair, with or without capsular shift. PMID- 15650672 TI - Part II: arthroscopic treatment of tibial plateau fractures: intercondylar eminence avulsion fractures. AB - Arthroscopic reduction and internal fixation (ARIF) of tibial intercondylar eminence fractures is the emerging state-of-the-art. ARIF is recommended for displaced type III fractures and should be considered for all cases of displaced type II fractures. Fractures without displacement after closed reduction require careful evaluation to rule out meniscal entrapment. Subjective results of ARIF are uniformly excellent, despite reports of objective anteroposterior laxity. Early range-of-motion exercises are essential to prevent loss of extension. Repair using nonabsorbable suture fixation, when of adequate strength to allow early range-of-motion, has the advantages of eliminating the risks of comminution of the fracture fragment, posterior neurovascular injury, and need for hardware removal, compared with ARIF using screws. PMID- 15650673 TI - Arthroscopic-assisted revision of telescopic rods in osteogenesis imperfecta. AB - Bailey-Dubow extensible nails are intramedullary stabilization devices designed to prevent deformities and recurrent long bone fractures in patients with osteogenesis imperfecta. The rods consist of a hollow outer sleeve and a solid inner obturator, allowing for telescoping of the rods and expansion of the device with longitudinal growth of the bone. Migration of the nail into the joint is a known complication of this procedure, which can lead to pain and loss of motion. This is a case report of the use of an arthroscopic approach to manipulate femoral and tibial Bailey-Dubow rods that migrated into the knee joint. PMID- 15650674 TI - Intra-articular lipoma of the knee joint in a girl. AB - Intra-articular lipoma is a rare benign mass that commonly occurs in the knee joint. We present the case of a 15-year-old girl with a slowly growing mass in her right knee. There was no history of trauma. The patient had no catching, locking, or giving way of the knee. On magnetic resonance imaging, multiple fibrous septa within a hyperintense mass was determined. After the arthroscopic examination, complete resection by open surgery was performed. There was no tumor recurrence at the end of the 1-year follow-up period. True intra-articular lipoma should be distinguished from lipoma arborescens, which is a similar but more common condition. PMID- 15650675 TI - Deep venous thrombosis and pulmonary embolism after knee arthroscopy in athletes carrying the thrombophilic factor lupus anticoagulant. AB - In patients undergoing arthroscopic knee surgery, deep venous thrombosis and pulmonary embolism are rare and there is not clear indication as to the necessity of thromboprophylactic treatment in these patients. However, the role of coagulation disorders in thrombotic events following arthroscopy is unknown. We report 2 cases of massive deep venous thrombosis and pulmonary embolism after knee arthroscopy in athletes carrying the thrombophilic factor lupus anticoagulant, but with no personal or familial history of thrombotic diseases. A few days after the arthroscopic intervention, both patients presented with deep venous thrombosis and 1 developed a severe pulmonary embolism. Blood examination showed that both athletes were lupus anticoagulant-positive. This is the first description of an association between venous thromboembolism, knee arthroscopy, and a prothrombotic condition. This report suggests that screening for hypercoagulability might be useful in athletes undergoing even minimally invasive orthopaedic surgery and that in cases of venous thromboembolism after knee arthroscopy, a prothrombotic disorder should be suspected. PMID- 15650676 TI - One-portal endoscopically assisted fasciotomy for exertional compartment syndrome. AB - A newly described method of single-incision endoscopically assisted fasciotomy for chronic exertional compartment syndrome is outlined. The procedure affords a small 2- to 3-cm incision with visualization of the anterior compartment fascia, lateral compartment fascia, superficial peroneal nerve, perforating vessels, and underlying muscle. The single-portal endoscopically assisted fasciotomy for chronic exertional compartment syndrome in the anterior and lateral compartments of the lower leg is a safe and reliable technique with excellent outcomes and patient satisfaction. Moreover, this technique affords the patient an expeditious recovery because of the small incision and decreased soft tissue trauma throughout the lower leg. PMID- 15650677 TI - New secure suture relay technique for arthroscopic Bankart repair without making an additional working portal. AB - Bankart repair is more frequently performed arthroscopically these days. To do this, an additional working portal is usually necessary, especially for repair of the badly damaged labrum or capsular ligaments using various types of relay techniques. We have devised a suture relay technique to perform arthroscopic Bankart repair using suture anchors without making any additional working portal. This is advantageous not only cosmetically, but also in terms of cost because only a simple device is required. This technique can be applied to almost any type of Bankart lesion repair as well as the plication of the capsule or SLAP lesion repair. PMID- 15650678 TI - Arthroscopic capsular plication-shift. AB - We describe a technique of arthroscopic capsular plication and shift used to address both anterior and posterior capsular laxity. This technique is used when the labrum is intact or if laxity persists after labral repair. PMID- 15650679 TI - Salvage rotator cuff repair using a biotenodesis screw. AB - Rotator cuff tear fixation can be difficult when humeral head bone stock is poor as a result of osteopenia or osteoporosis. This technical note describes biotenodesis screw use to achieve effective fixation after 2 previous attempts at implanting suture anchors failed. PMID- 15650680 TI - Acute pulmonary embolism: an update on diagnosis and management. AB - Pulmonary embolism (PE) is a common problem. Given the significant overlap of symptoms and signs between the presentation of PE and acute coronary syndromes, it becomes clear that cardiologists must be familiar with the diagnosis and treatment of PE. The critical issue is always to consider PE in the diagnosis of chest pain. It is then important to determine the likelihood of the diagnosis. For patients at moderate-to-high risk, helical CT provides a rapid and noninvasive diagnostic tool. Several other imaging studies are also available including ventilation/perfusion (V/Q) scan, magnetic resonance imaging, and pulmonary arteriography. Echocardiography can also provide valuable prognostic information. Several biomarkers including the d-dimers, troponins, and natriuretic peptides may provide additional information. The cornerstone of treatment includes anticoagulation. For patients with massive or submassive PE, thrombolysis and embolectomy should be considered. Finally, both primary and secondary prevention are critical to the long-term health of the patient. PMID- 15650682 TI - Effects of elevated atmospheric CO2 concentrations on soil microorganisms. AB - Effects of elevated CO(2) on soil microorganisms are known to be mediated by various interactions with plants, for which such effects are relatively poorly documented. In this review, we summarize and synthesize results from studies assessing impacts of elevated CO(2) on soil ecosystems, focusing primarily on plants and a variety the of microbial processes. The processes considered include changes in microbial biomass of C and N, microbial number, respiration rates, organic matter decomposition, soil enzyme activities, microbial community composition, and functional groups of bacteria mediating trace gas emission such as methane and nitrous oxide. Elevated CO(2) in atmosphere may enhance certain microbial processes such as CH(4) emission from wetlands due to enhanced carbon supply from plants. However, responses of extracellular enzyme activities and microbial community structure are still controversy, because interferences with other factors such as the types of plants, nutrient availabilitial in soil, soil types, analysis methods, and types of CO(2) fumigation systems are not fully understood. PMID- 15650683 TI - Monitoring of bacterial community in a coniferous forest soil after a wildfire. AB - Changes in the soil bacterial community of a coniferous forest were analyzed to assess microbial responses to wildfire. Soil samples were collected from three different depths in lightly and severely burned areas, as well as a nearby unburned control area. Direct bacterial counts ranged from 3.3-22.6 x 10(8) cells/(g.soil). In surface soil, direct bacterial counts of unburned soil exhibited a great degree of fluctuation. Those in lightly burned soil changed less, but no significant variation was observed in the severely burned soil. The fluctuations of direct bacterial count were less in the middle and deep soil layers. The structure of the bacterial community was analyzed via the fluorescent in situ hybridization method. The number of bacteria detected with the eubacteria targeted probe out of the direct bacterial count varied from 30.3 to 84.7%, and these ratios were generally higher in the burned soils than in the unburned control soils. In the surface unburned soil, the ratios of alpha-, beta- and gamma-proteobacteria, Cytophaga-Flavobacterium group, and other eubacteria groups to total eubacteria were 9.9, 10.6, 15.5, 9.0, and 55.0%, respectively, and these ratios were relatively stable. The ratios of alpha-, beta- and gamma proteobacteria, and Cytophaga-Flavobacterium group to total eubacteria increased immediately after the wildfire, and the other eubacterial proportions decreased in the surface and middle layer soils. By way of contrast, the composition of the 5 groups of eubacteria in the subsurface soil exhibited no significant fluctuations during the entire period. The total bacterial population and bacterial community structure disturbed by wildfire soon began to recover, and original levels seemed to be restored 3 months after the wildfire. PMID- 15650684 TI - Monitoring of soil bacterial community and some inoculated bacteria after prescribed fire in microcosm. AB - The soil bacterial community and some inoculated bacteria were monitored to assess the microbial responses to prescribed fire in their microcosm. An acridine orange direct count of the bacteria in the unburned control soil were maintained at a relatively stable level (2.0 approximately 2.7 x 10(9) cells/g(-1).soil) during the 180 day study period. The number of bacteria in the surface soil was decreased by fire, but was restored after 3 months. Inoculation of some bacteria increased the number of inoculated bacteria several times and these elevated levels lasted several months. The ratios of eubacteria detected by a fluorescent in situ hybridization (FISH) method to direct bacterial count were in the range of 60 approximately 80% during the study period, with the exception of some lower values at the beginning, but there were no definite differences between the burned and unburned soils or the inoculated and uninoculated soils. In the unburned control soil, the ratios of alpha-, beta- and gamma-subgroups of the proteobacteria, Cytophaga-Flavobacterium and other eubacteria groups to that of the entire eubacteria were 13.7, 31.7, 17.1, 16.8 and 20.8%, respectively, at time 0. The overall change on the patterns of the ratios of the 5 subgroups of eubacteria in the uninoculated burned and inoculated soils were similar to those of the unburned control soil, with the exception of some minor variations during the initial period. The proportions of each group of eubacteria became similar in the different microcosms after 6 months, which may indicate the recovery of the original soil microbial community structure after fire or the inoculation of some bacteria. The populations of Azotobacter vinelandii, Bacillus megaterium and Pseudomonas fluorescens, which had been inoculated to enhance the microbial activities, and monitored by FISH method, showed similar changes in the microcosms, and maintained high levels for several months. PMID- 15650685 TI - Introduction of saxicolous lichens distributed in coastal rocks of U-do islet in Jeju, Korea. AB - This study reports, for the first time, the investigation of the distribution of Korean saxicolous lichens in the coastal rocks of U-do islet, which is known as an unpolluted zone in Jeju. More than thirty lichens were obtained and investigated from the coastal rocks frequently contacted by seawater. A molecular analysis using PCR amplification of the rRNA ITS regions revealed the coastal rock lichens could be placed into 8 families and 14 genera, Ramalinaceae (Bacidia, Ramalina), Physciaceae (Buellia, Dirinaria, Phaeophyscia, Physcia, Pyxine), Lecanoraceae (Candelaria, Lecanora), Parmeliaceae (Xanthoparmelia), Graphidaceae (Graphis), Pertusariaceae (Pertusaria), Rhizocarpaceae (Rhizocarpon), and Teloschistaceae (Caloplaca), showing a diversity of lichens, with foliose (flat leaf-like), crustose (crust-like), and fruticose (miniature shrub-like) life forms might be distributed in the coastal rocks. These findings suggested the possibility that the lichens identified in the present work might be resistant to a salty environment. PMID- 15650686 TI - Detection of enterovirus, cytomegalovirus, and Chlamydia pneumoniae in atheromas. AB - To investigate the presence of infectious agents in human atherosclerotic arterial tissues. Atherosclerotic plaques were removed from 128 patients undergoing carotid endarterectomy or other bypass procedures for occlusive disease, and from twenty normal arterial wall samples, obtained from transplant donors with no history of diabetes, hypertension, smoking, or hyperlipidemia. Using the polymerase chain reaction (PCR) or reverse transcription-PCR, these samples were analyzed for the presence of Chlamydia pneumoniae, cytomegalovirus, enterovirus, adenovirus, herpes simplex viruses types 1 and 2, and Epstein-Barr virus. The amplicons were then sequenced, and phylogenetic analyses were performed. Enteroviral RNA was found in 22 of 128 atherosclerotic vascular lesions (17.2%), and C. pneumoniae and cytomegalovirus were each found in 2 samples (1.6%). In contrast, adenovirus, herpes simplex viruses, and Epstein-Barr virus were not identified in any of the atherosclerotic samples. Enterovirus was detected in 6/24 (25.0%) aortas, 7/33 (21.2%) carotid arteries, 6/40 (15.0%) femoral arteries, and 3/31 (9.7%) radial arteries of patients with chronic renal failure. There were no infectious agents detected in any of the control specimens. Using phylogenetic analysis, the enterovirus isolates were clustered into 3 groups, arranged as echovirus 9 and coxsackieviruses B1 and B3. Enteroviral RNA was detected in 17.2% of atherosclerotic plaques, but was not observed in any of the control specimens. This suggests a connection between enteroviral infection and atherosclerosis. These findings differ from those of other studies, which found more frequent incidence of C. pneumoniae and cytomegalovirus infection in atherosclerotic plaques. PMID- 15650687 TI - Axenic culture of Gyrodinium impudicum strain KG03, a marine red-tide microalga that produces exopolysaccharide. AB - An exopolysaccharide-producing microalgal dinoflagellate was isolated from a red tide bloom and designated strain KG03. A bacteria-free culture of strain KG03 was achieved using a modified wash with phototaxis and antibiotic treatment. Combined treatment with neomycin and cephalosporin was the most effective for eliminating the bacteria associated with the microalgae. Strain KG03 was identified as Gyrodinium impudicum by analyzing the ITS regions of the 5.8S rDNA, 18S rDNA, morphological phenotype and fatty acid composition. The exopolysaccharide production and cell growth in a 300-ml photobioreactor were increased 2.7- and 2.4-fold, respectively, compared with that in a flask culture at the first isolation step. PMID- 15650688 TI - Probiotication of tomato juice by lactic acid bacteria. AB - This study was undertaken to determine the suitability of tomato juice as a raw material for production of probiotic juice by four lactic acid bacteria (Latobacillus acidophilus LA39, Lactobacillus plantarum C3, Lactobacillus casei A4, and Lactobacillus delbrueckii D7). Tomato juice was inoculated with a 24-h old culture and incubated at 30 degrees C. Changes in pH, acidity, sugar content, and viable cell counts during fermentation under controlled conditions were measured. The lactic acid cultures reduced the pH to 4.1 or below and increased the acidity to 0.65% or higher, and the viable cell counts (CFU) reached nearly 1.0 to 9.0 x 10(9)/ml after 72 h fermentation. The viable cell counts of the four lactic acid bacteria in the fermented tomato juice ranged from 10(6) to 10(8) CFU/ml after 4 weeks of cold storage at 4 degrees C. Probiotic tomato juice could serve as a health beverage for vegetarians or consumers who are allergic to dairy products. PMID- 15650690 TI - Characteristics of HIV-Tat protein transduction domain. AB - The human immunodeficiency virus type 1 (HIV-1) Tat protein transduction domain (PTD), which contains rich arginine and lysine residues, is responsible for the highly efficient transduction of protein through the plasma membrane. In addition, it can be secreted from infected cells and has the ability to enter neighboring cells. When the PTD of Tat is fused to proteins and exogenously added to cells, the fusion protein can cross plasma membranes. Recent reports indicate that the endogenously expressed Tat fusion protein can demonstrate biodistribution of several proteins. However, intercellular transport and protein transduction have not been observed in some studies. Therefore, this study examined the intercellular transport and protein transduction of the Tat protein. The results showed no evidence of intercellular transport (biodistribution) in a cell culture. Instead, the Tat fusion peptides were found to have a significant effect on the transduction and intercellular localization properties. This suggests that the HIV-1 PTD passes through the plasma membrane in one direction. PMID- 15650689 TI - Expression of the promoter for the maltogenic amylase gene in Bacillus subtilis 168. AB - An additional amylase, besides the typical alpha-amylase, was detected for the first time in the cytoplasm of B. subtilis SUH4-2, an isolate from Korean soil. The corresponding gene (bbmA) encoded a maltogenic amylase (MAase) and its sequence was almost identical to the yvdF gene of B. subtilis 168, whose function was unknown. Southern blot analysis using bbmA as the probe indicated that this gene was ubiquitous among various B. subtilis strains. In an effort to understand the physiological function of the bbmA gene in B. subtilis, the expression pattern of the gene was monitored by measuring the beta-galactosidase activity produced from the bbmA promoter fused to the amino terminus of the lacZ structural gene, which was then integrated into the amyE locus on the B. subtilis 168 chromosome. The promoter was induced during the mid-log phase and fully expressed at the early stationary phase in defined media containing beta cyclodextrin (beta-CD), maltose, or starch. On the other hand, it was kept repressed in the presence of glucose, fructose, sucrose, or glycerol, suggesting that catabolite repression might be involved in the expression of the gene. Production of the beta-CD hydrolyzing activity was impaired by the spo0A mutation in B. subtilis 168, indicating the involvement of an additional regulatory system exerting control on the promoter. Inactivation of yvdF resulted in a significant decrease of the beta-CD hydrolyzing activity, if not all. This result implied the presence of an additional enzyme(s) that is capable of hydrolyzing beta-CD in B. subtilis 168. Based on the results, MAase encoded by bbmA is likely to be involved in maltose and beta-CD utilization when other sugars, which are readily usable as an energy source, are not available during the stationary phase. PMID- 15650691 TI - Hydrogen peroxide produced by two amino acid oxidases mediates antibacterial actions. AB - The antibacterial actions of two amino acid oxidases, a D-amino acid oxidase from hog kidney and a L-amino acid oxidase from the venom of Agkistrodon halys, were investigated, demonstrating that both enzymes were able to inhibit the growth of both Gram-positive and Gram-negative bacteria, and that hydrogen peroxide, a product of their enzymatic reactions, was the antibacterial factor. However, hydrogen peroxide generated in the enzymatic reactions was not sufficient to explain the degree to which bacterial growth was inhibited. A fluorescence labeling assay showed that both of these two enzymes could bind to the surfaces of bacteria. To the best of our knowledge, this is the first report regarding the antibacterial activity of the D-amino acid oxidases. PMID- 15650692 TI - Expression of a recombinant Cry1Ac crystal protein fused with a green fluorescent protein in Bacillus thuringiensis subsp. kurstaki Cry-B. AB - To investigate the co-expression and crystallization of a fusion gene between the Bacillus thuringiensis crystal protein and a foreign protein in B. thuringiensis, the expression of the Cry1Ac fused with green fluorescent protein (GFP) genes in a B. thuringiensis Cry(-)B strain was examined. The cry1Ac gene was cloned in the B. thuringiensis-E. coli shuttle vector, pHT3101, under the control of the native cry1Ac gene promoter, while the GFP gene was inserted into the XhoI site upstream of the proteolytic cleavage site, in the middle region of the cry1Ac gene (pProAc GFP). The B. thuringiensis Cry(-)B strain carrying pProAc-GFP (ProAc-GFP/CB) did not produce any inclusion bodies. However, the transformed strain expressed fusion protein forms although the expression level was relatively low. Furthermore, an immunoblot analysis using GFP and Cry1Ac antibodies showed that the fusion protein was not a single species, but rather multiple forms. In addition, the N-terminal fragment of Cry1Ac and a non-fused GFP were also found in the B. thuringiensis Cry(-)B strain after autolysis. The sporulated cells before autolysis and the spore-crystal mixture after autolysis of ProAc-GFP/CB exhibited insecticidal activities against Plutella xylostella larvae. Accordingly, the current results suggest that a fusion crystal protein produced by the transfomant, ProAc-GFP/CB, can be functionally expressed but easily degraded in B. thuringiensis. PMID- 15650693 TI - Enzymatic and non-enzymatic degradation of poly (3-hydroxybutyrate-co-3 hydroxyvalerate) copolyesters produced by Alcaligenes sp. MT-16. AB - Poly(3-hydroxybutyrate-co-3-hydroxyvalerate), poly(3HB-co-3HV), copolyesters with a variety of 3HV contents (ranging from 17 to 60 mol%) were produced by Alcaligenes sp. MT-16 grown on a medium containing glucose and levulinic acid in various ratios, and the effects of hydrophilicity and crystallinity on the degradability of the copolyesters were evaluated. Measurements of thermo mechanical properties and Fourier-transform infrared spectroscopy in the attenuated total reflectance revealed that the hydrophilicity and crystallinity of poly(3HB-co-3HV) copolyesters decreased as 3HV content in the copolyester increased. When the prepared copolyester film samples were non-enzymatically hydrolysed in 0.01 N NaOH solution, the weights of all samples were found to have undergone no changes over a period of 20 weeks. In contrast, the copolyester film samples were degraded by the action of extracellular polyhydroxybutyrate depolymerase from Emericellopsis minima W2. The overall rate of weight loss was higher in the films containing higher amounts of 3HV, suggesting that the enzymatic degradation of the copolyester is more dependent on the crystallinity of the copolyester than on its hydrophilicity. Our results suggest that the degradability characteristics of poly(3HB-co-3HV) copolyesters, as well as their thermo-mechanical properties, are greatly influenced by the 3HV content in the copolyesters. PMID- 15650694 TI - Transcriptional regulation of the Schizosaccharomyces pombe gene encoding glutathione S-transferase I by a transcription factor Pap1. AB - In a previous study, a gst gene was isolated from the fission yeast Schizosaccharomyces pombe. This gene was dubbed gst I, and was characterized using the gstI-lacZ fusion plasmid pYSH2000. In this work, four additional fusion plasmids, pYSHSD1, pYSHSD2, pYSHSD3 and pYSHSD4, were constructed, in order to carry (respectively) 770, 551, 358 and 151 bp upstream regions from the translational initiation point. The sequence responsible for induction by aluminum, mercury and hydrogen peroxide was located in the range between -1,088 and -770 bp upstream of the S. pombe gst I gene. The same region was identified to contain the nucleotide sequence responsible for regulation by Pap1, and has one putative Pap1 binding site, TTACGTAT, located in the range between -954 approximately -947 bp upstream of the gst I gene. Negatively acting sequences are located between -1,088 and -151 bp. These findings imply that the Pap1 protein is involved in basal and inducible transcription of the gst I gene in the fission yeast S. pombe. PMID- 15650695 TI - Bacterial aggregates formation after addition of glucose in Lake Baikal water. AB - For determining the process of bacterial aggregation, glucose was added into water from Lake Baikal which had been stored for seven months. In the presence of a higher concentration of glucose, the abundance of single bacteria and aggregates were higher, but the biovolumes of both bacteria were similar. Theses results mean that both free-living and aggregated bacteria have similar maximum sizes and that aggregates are forming with available organic materials. With available organic materials, the biovolume of aggregates becomes larger. PMID- 15650696 TI - Comparative analysis of intracellular trans-splicing ribozyme activity against hepatitis C virus internal ribosome entry site. AB - Internal ribosome entry site (IRES) of the hepatitis C virus (HCV) is known to be essential for HCV replication and most conserved among HCV variants. Hence, IRES RNA is a good therapeutic target for RNA-based inhibitors, such as ribozymes. We previously proposed a new anti-HCV modulation strategy based on trans-splicing ribozymes, which can selectively replace HCV transcripts with a new RNA that exerts anti-HCV activity. To explore this procedure, sites which are accessible to ribozymes in HCV IRES were previously determined by employing an RNA mapping method in vitro. In this study, we evaluate the intracellular accessibility of the ribozymes by comparing the trans-splicing activities in cells of several ribozymes targeting different sites of the HCV IRES RNA. We assessed the intracellular activities of the ribozymes by monitoring their target-specific induction degree of both reporter gene activity and cytotoxin expression. The ribozyme capable of targeting the most accessible site identified by the mapping studies then harbored the most active trans-splicing activity in cells. These results suggest that the target sites predicted to be accessible are truly the most accessible in the cells, and thus, could be applied to the development of various RNA-based anti-HCV therapies. PMID- 15650697 TI - Occurrence of the strA-strB streptomycin resistance genes in Pseudomonas species isolated from kiwifruit plants. AB - The occurrence of strA-strB streptomycin-resistance genes within transposon Tn5393 was examined in Pseudomonas syringae pv. actinidiae, P. syringae pv. syringae, and P. marginalis, isolated from kiwifruit plants in Korea and Japan. PCR amplification with primers specific to strA-strB revealed that three of the tested Pseudomonas species harbored these genes for a streptomycin-resistance determinant. Tn5393, containing strA-strB, was also identified with PCR primers designed to amplify parts of tnpA, res, and tnpR. No IS elements were detected within tnpR, nor were they found in the intergenic region between tnpR and strA. Nucleotide sequence analysis indicated that the strA sequence of P. syringae pv. actinidiae contained a single nucleotide alteration at position 593 (CAA-->CGA), as compared to Tn5393a in P. syringae pv. syringae. This resulted in an amino acid change, from Gln to Arg. PMID- 15650698 TI - Strategies against human papillomavirus infection and cervical cancer. AB - Papillomaviruses infect a wide variety of animals, including humans. The human papillomavirus (HPV), in particular, is one of the most common causes of sexually transmitted disease. More than 200 types of HPV have been identified by DNA sequence data, and 85 HPV genotypes have been well characterized to date. HPV can infect the basal epithelial cells of the skin or inner tissue linings, and are, accordingly, categorized as either cutaneous or mucosal type. HPV is associated with a panoply of clinical conditions, ranging from innocuous lesions to cervical cancer. In the early 1980s, studies first reported a link between cervical cancer and genital HPV infection. Genital HPV infections are now recognized to be a major risk factor in at least 95% of cervical cancers. 30 different HPV genotypes have been identified as causative of sexually transmitted diseases, most of which induce lesions in the cervix, vagina, vulva, penis, and anus, as the result of sexual contact. There is also direct evidence demonstrating that at least four of these genotypes are prerequisite factors in cervical cancer. The main aim of this review was to evaluate the current literature regarding the pathovirology, diagnostics, vaccines, therapy, risk groups, and further therapeutic directions for HPV infections. In addition, we reviewed the current status of HPV infections in South Korean women, as evidenced by our data. PMID- 15650700 TI - Arbuscular mycorrhizal fungi: hyphal fusion and multigenomic structure. AB - Arbuscular mycorrhizal (AM) fungi (Glomeromycota) reproduce asexually, are multinucleate, and have high genetic variation within single cells. Pawlowska and Taylor find that genetic variation within AM fungal cells is not lost as a result of segregation, and they interpret this as evidence that the variation is present within each nucleus and that all nuclei within individual spores are genetically identical (that is, homokaryotic). Here we show that their empirical observations are also consistent with a distribution of genetic variation between nuclei within spores (that is, heterokaryotic), given that there is fusion of fungal hyphae. This analysis, together with complementary findings, suggests that AM fungi have an unusual genomic structure in which multiple, genetically diverse nuclei are maintained within cells through remixing by hyphal fusion. PMID- 15650699 TI - Difficulties in achieving vs maintaining erection: organic, psychogenic and relational determinants. AB - Achieving and maintaining a penile erection are two essential components of the male sexual response. It has recently been suggested that distinct molecular mechanism could underlie the two disturbances. The aim of the present study is to verify possible clinical differences on pathogenetic factors underlying difficulties of achieving and maintaining an erection. We studied a consecutive series of 560 patients (aged 51.9+/-12.8 y old) reporting erectile dysfunction (ED), using SIEDY structured interview. Patients were classified into two distinct categories: those with difficulties in maintaining, rather than achieving, an erection (sample A) and those with main problems in achieving an erection (sample B). A complete physical examination and a series of metabolic, biochemical, hormonal, psychometric, penile vascular tests and nocturnal penile tumescence and rigidity evaluations (NPT) were also performed. Sample B patients showed a higher prevalence of organic conditions related to ED, when compared with sample A as confirmed by higher SIEDY scale 1 scores (3[1-5] vs 1[0.1-3] for sample B vs sample A, respectively; P < 0.0001) which explores organic component of ED and higher prevalence of pathological instrumental parameters. No difference among groups was observed for SIEDY scale 2 (relational component) and SIEDY scale 3 (intrapsychic component) of ED. In conclusion, this study shows for the first time that patients with difficulties in maintaining erection are less likely to be affected by organic disturbances interfering with sexual function, when compared with those unable to achieve a valid erection. PMID- 15650702 TI - Rockets in Russia's back yard. PMID- 15650703 TI - Dangerous state of denial. PMID- 15650704 TI - Forces gather behind proposal for a natural-disaster agency. PMID- 15650705 TI - Scientists seek action to fix Asia's ravaged ecosystems. PMID- 15650706 TI - Agencies fear global crises will lose out to tsunami donations. PMID- 15650707 TI - Study links sickness to Russian launch site. PMID- 15650708 TI - Health rules may hamper Japanese import of lab mice. PMID- 15650709 TI - European research framework set to expand. PMID- 15650710 TI - Science's next generation finds its own way. PMID- 15650712 TI - Historical monuments: the film crew. PMID- 15650713 TI - Infectious disease: Vietnam's war on flu. PMID- 15650714 TI - No political interference in US agricultural grants. PMID- 15650715 TI - Oceans: fisheries not to blame for damage. PMID- 15650716 TI - Oceans need protection from scientists too. PMID- 15650721 TI - Body doubles. PMID- 15650722 TI - Medicine: knockout malaria vaccine? PMID- 15650723 TI - Planetary science: construction-site inspection. PMID- 15650724 TI - Palaeoclimate: ripples of stormy weather. PMID- 15650725 TI - Mammalian palaeobiology: living large in the Cretaceous. PMID- 15650727 TI - Astrophysics: the process of carbon creation. PMID- 15650728 TI - Conservation biology: parasite rattles diversity's cage. PMID- 15650729 TI - Behavioural ecology: tool manufacture by naive juvenile crows. AB - New Caledonian crows (Corvus moneduloides) are the most prolific avian tool users. Regional variation in the shape of their tools may be the result of cumulative cultural evolution--a phenomenon considered to be a hallmark of human culture. Here we show that hand-raised juvenile New Caledonian crows spontaneously manufacture and use tools, without any contact with adults of their species or any prior demonstration by humans. Our finding is a crucial step towards producing informed models of cultural transmission in this species, and in animals in general. PMID- 15650730 TI - Extreme winds and waves in the aftermath of a Neoproterozoic glaciation. AB - The most severe excursions in the Earth's climatic history are thought to be associated with Proterozoic glaciations. According to the 'Snowball Earth' hypothesis, the Marinoan glaciation, which ended about 635 million years ago, involved global or nearly global ice cover. At the termination of this glacial period, rapid melting of continental ice sheets must have caused a large rise in sea level. Here we show that sediments deposited during this sea level rise contain remarkable structures that we interpret as giant wave ripples. These structures occur at homologous stratigraphic levels in Australia, Brazil, Canada, Namibia and Svalbard. Our hydrodynamic analysis of these structures suggests maximum wave periods of 21 to 30 seconds, significantly longer than those typical for today's oceans. The reconstructed wave conditions could only have been generated under sustained high wind velocities exceeding 20 metres per second in fetch-unlimited ocean basins. We propose that these extraordinary wind and wave conditions were characteristic of the climatic transit, and provide observational targets for atmospheric circulation models. PMID- 15650731 TI - Simultaneous determination of protein structure and dynamics. AB - We present a protocol for the experimental determination of ensembles of protein conformations that represent simultaneously the native structure and its associated dynamics. The procedure combines the strengths of nuclear magnetic resonance spectroscopy--for obtaining experimental information at the atomic level about the structural and dynamical features of proteins--with the ability of molecular dynamics simulations to explore a wide range of protein conformations. We illustrate the method for human ubiquitin in solution and find that there is considerable conformational heterogeneity throughout the protein structure. The interior atoms of the protein are tightly packed in each individual conformation that contributes to the ensemble but their overall behaviour can be described as having a significant degree of liquid-like character. The protocol is completely general and should lead to significant advances in our ability to understand and utilize the structures of native proteins. PMID- 15650732 TI - Mid-infrared images of beta Pictoris and the possible role of planetesimal collisions in the central disk. AB - When viewed in optical starlight scattered by dust, the nearly edge-on debris disk surrounding the A5V star beta Pictoris (distance 19.3 pc; ref. 1) extends farther than 1,450 au from the star. Its large-scale complexity has been well characterized, but the detailed structure of the disk's central approximately 200 au region has remained elusive. This region is of special interest, because planets may have formed there during the star's 10-20-million-year lifetime, perhaps resulting in both the observed tilt of 4.6 degrees relative to the large scale main disk and the partial clearing of the innermost dust. A peculiarity of the central disk (also possibly related to the presence of planets) is the asymmetry in the brightness of the 'wings', in which the southwestern wing is brighter and more extended at 12 microm than the northeastern wing. Here we present thermal infrared images of the central disk that imply that the brightness asymmetry results from the presence of a bright clump composed of particles that may differ in size from dust elsewhere in the disk. We suggest that this clump results from the collisional grinding of resonantly trapped planetesimals or the cataclysmic break-up of a planetesimal. PMID- 15650733 TI - Revised rates for the stellar triple-alpha process from measurement of 12C nuclear resonances. AB - In the centres of stars where the temperature is high enough, three alpha particles (helium nuclei) are able to combine to form 12C because of a resonant reaction leading to a nuclear excited state. (Stars with masses greater than approximately 0.5 times that of the Sun will at some point in their lives have a central temperature high enough for this reaction to proceed.) Although the reaction rate is of critical significance for determining elemental abundances in the Universe, and for determining the size of the iron core of a star just before it goes supernova, it has hitherto been insufficiently determined. Here we report a measurement of the inverse process, where a 12C nucleus decays to three alpha particles. We find a dominant resonance at an energy of approximately 11 MeV, but do not confirm the presence of a resonance at 9.1 MeV (ref. 3). We show that interference between two resonances has important effects on our measured spectrum. Using these data, we calculate the triple-alpha rate for temperatures from 10(7) K to 10(10) K and find significant deviations from the standard rates. Our rate below approximately 5 x 10(7) K is higher than the previous standard, implying that the critical amounts of carbon that catalysed hydrogen burning in the first stars are produced twice as fast as previously believed. At temperatures above 10(9) K, our rate is much less, which modifies predicted nucleosynthesis in supernovae. PMID- 15650734 TI - Systematic design of chemical oscillators using complexation and precipitation equilibria. AB - Concentration oscillations are ubiquitous in living systems, where they involve a wide range of chemical species. In contrast, early in vitro chemical oscillators were all derived from two accidentally discovered reactions based on oxyhalogen chemistry. Over the past 25 years, the use of a systematic design algorithm, in which a slow feedback reaction periodically drives a bistable system in a flow reactor between its two steady states, has increased the list of oscillating chemical reactions to dozens of systems. But these oscillating reactions are still confined to a handful of elements that possess multiple stable oxidation states: halogens, sulphur and some transition metals. Here we show that linking a 'core' oscillator to a complexation or precipitation equilibrium can induce concentration oscillations in a species participating in the equilibrium. We use this method to design systems that produce periodic pulses of calcium, aluminium or fluoride ions. The ability to generate oscillations in elements possessing only a single stable oxidation state (for example, Na+, F-, Ca2+) may lead to reactions that are useful for coupling to or probing living systems, or that help us to understand new mechanisms by which periodic behaviour may arise. PMID- 15650735 TI - Efficient export of carbon to the deep ocean through dissolved organic matter. AB - Oceanic dissolved organic carbon (DOC) constitutes one of the largest pools of reduced carbon in the biosphere. Estimated DOC export from the surface ocean represents 20% of total organic carbon flux to the deep ocean, which constitutes a primary control on atmospheric carbon dioxide levels. DOC is the carbon component of dissolved organic matter (DOM) and an accurate quantification of DOM pools, fluxes and their controls is therefore critical to understanding oceanic carbon cycling. DOC export is directly coupled with dissolved organic nitrogen and phosphorus export. However, the C:N:P stoichiometry (by atoms) of DOM dynamics is poorly understood. Here we study the stoichiometry of the DOM pool and of DOM decomposition in continental shelf, continental slope and central ocean gyre environments. We find that DOM is remineralized and produced with a C:N:P stoichiometry of 199:20:1 that is substantially lower than for bulk pools (typically >775:54:1), but greater than for particulate organic matter (106:16:1- the Redfield ratio). Thus for a given mass of new N and P introduced into surface water, more DOC can be exported than would occur at the Redfield ratio. This may contribute to the excess respiration estimated to occur in the interior ocean. Our results place an explicit constraint on global carbon export and elemental balance via advective pathways. PMID- 15650736 TI - Magma-assisted rifting in Ethiopia. AB - The rifting of continents and evolution of ocean basins is a fundamental component of plate tectonics, yet the process of continental break-up remains controversial. Plate driving forces have been estimated to be as much as an order of magnitude smaller than those required to rupture thick continental lithosphere. However, Buck has proposed that lithospheric heating by mantle upwelling and related magma production could promote lithospheric rupture at much lower stresses. Such models of mechanical versus magma-assisted extension can be tested, because they predict different temporal and spatial patterns of crustal and upper-mantle structure. Changes in plate deformation produce strain-enhanced crystal alignment and increased melt production within the upper mantle, both of which can cause seismic anisotropy. The Northern Ethiopian Rift is an ideal place to test break-up models because it formed in cratonic lithosphere with minor far field plate stresses. Here we present evidence of seismic anisotropy in the upper mantle of this rift zone using observations of shear-wave splitting. Our observations, together with recent geological data, indicate a strong component of melt-induced anisotropy with only minor crustal stretching, supporting the magma-assisted rifting model in this area of initially cold, thick continental lithosphere. PMID- 15650737 TI - Large Mesozoic mammals fed on young dinosaurs. AB - Mesozoic mammals are commonly portrayed as shrew- or rat-sized animals that were mainly insectivorous, probably nocturnal and lived in the shadow of dinosaurs. The largest known Mesozoic mammal represented by substantially complete remains is Repenomamus robustus, a triconodont mammal from the Lower Cretaceous of Liaoning, China. An adult individual of R. robustus was the size of a Virginia opossum. Here we report a new species of the genus, represented by a skeleton with most of the skull and postcranium preserved in articulation. The new species is 50% larger than R. robustus in skull length. In addition, stomach contents associated with a skeleton of R. robustus reveal remains of a juvenile Psittacosaurus, a ceratopsian dinosaur. Our discoveries constitute the first direct evidence that some triconodont mammals were carnivorous and fed on small vertebrates, including young dinosaurs, and also show that Mesozoic mammals had a much greater range of body sizes than previously known. We suggest that Mesozoic mammals occupied diverse niches and that some large mammals probably competed with dinosaurs for food and territory. PMID- 15650738 TI - The simplicity of metazoan cell lineages. AB - Developmental processes are thought to be highly complex, but there have been few attempts to measure and compare such complexity across different groups of organisms. Here we introduce a measure of biological complexity based on the similarity between developmental and computer programs. We define the algorithmic complexity of a cell lineage as the length of the shortest description of the lineage based on its constituent sublineages. We then use this measure to estimate the complexity of the embryonic lineages of four metazoan species from two different phyla. We find that these cell lineages are significantly simpler than would be expected by chance. Furthermore, evolutionary simulations show that the complexity of the embryonic lineages surveyed is near that of the simplest lineages evolvable, assuming strong developmental constraints on the spatial positions of cells and stabilizing selection on cell number. We propose that selection for decreased complexity has played a major role in moulding metazoan cell lineages. PMID- 15650739 TI - Unexpected complexity of the Wnt gene family in a sea anemone. AB - The Wnt gene family encodes secreted signalling molecules that control cell fate in animal development and human diseases. Despite its significance, the evolution of this metazoan-specific protein family is unclear. In vertebrates, twelve Wnt subfamilies were defined, of which only six have counterparts in Ecdysozoa (for example, Drosophila and Caenorhabditis). Here, we report the isolation of twelve Wnt genes from the sea anemone Nematostella vectensis, a species representing the basal group within cnidarians. Cnidarians are diploblastic animals and the sister group to bilaterian metazoans. Phylogenetic analyses of N. vectensis Wnt genes reveal a thus far unpredicted ancestral diversity within the Wnt family. Cnidarians and bilaterians have at least eleven of the twelve known Wnt gene subfamilies in common; five subfamilies appear to be lost in the protostome lineage. Expression patterns of Wnt genes during N. vectensis embryogenesis indicate distinct roles of Wnts in gastrulation, resulting in serial overlapping expression domains along the primary axis of the planula larva. This unexpectedly complex inventory of Wnt family signalling factors evolved in early multi cellular animals about 650 million years (Myr) ago, predating the Cambrian explosion by at least 100 Myr (refs 5, 8). It emphasizes the crucial function of Wnt genes in the diversification of eumetazoan body plans. PMID- 15650740 TI - Low gene copy number shows that arbuscular mycorrhizal fungi inherit genetically different nuclei. AB - Arbuscular mycorrhizal fungi (AMF) are ancient asexually reproducing organisms that form symbioses with the majority of plant species, improving plant nutrition and promoting plant diversity. Little is known about the evolution or organization of the genomes of any eukaryotic symbiont or ancient asexual organism. Direct evidence shows that one AMF species is heterokaryotic; that is, containing populations of genetically different nuclei. It has been suggested, however, that the genetic variation passed from generation to generation in AMF is simply due to multiple chromosome sets (that is, high ploidy). Here we show that previously documented genetic variation in Pol-like sequences, which are passed from generation to generation, cannot be due to either high ploidy or repeated gene duplications. Our results provide the clearest evidence so far for substantial genetic differences among nuclei in AMF. We also show that even AMF with a very large nuclear DNA content are haploid. An underlying principle of evolutionary theory is that an individual passes on one or half of its genome to each of its progeny. The coexistence of a population of many genomes in AMF and their transfer to subsequent generations, therefore, has far-reaching consequences for understanding genome evolution. PMID- 15650741 TI - Binding of brassinosteroids to the extracellular domain of plant receptor kinase BRI1. AB - Both animals and plants use steroids as signalling molecules during growth and development. Animal steroids are principally recognized by members of the nuclear receptor superfamily of transcription factors. In plants, BRI1, a leucine-rich repeat (LRR) receptor kinase localized to the plasma membrane, is a critical component of a receptor complex for brassinosteroids. Here, we present the first evidence for direct binding of active brassinosteroids to BRI1 using a biotin tagged photoaffinity castasterone (BPCS), a biosynthetic precursor of brassinolide (the most active of the brassinosteroids). Binding studies using BPCS, (3)H-labelled brassinolide and recombinant BRI1 fragments show that the minimal binding domain for brassinosteroids consists of a 70-amino acid island domain (ID) located between LRR21 and LRR22 in the extracellular domain of BRI1, together with the carboxy-terminal flanking LRR (ID-LRR22). Our results demonstrate that brassinosteroids bind directly to the 94 amino acids comprising ID-LRR22 in the extracellular domain of BRI1, and define a new binding domain for steroid hormones. PMID- 15650743 TI - Scientific personalities. PMID- 15650742 TI - Stabilization of microtubule dynamics at anaphase onset promotes chromosome segregation. AB - Microtubules of the mitotic spindle form the structural basis for chromosome segregation. In metaphase, microtubules show high dynamic instability, which is thought to aid the 'search and capture' of chromosomes for bipolar alignment on the spindle. Microtubules suddenly become more stable at the onset of anaphase, but how this change in microtubule behaviour is regulated and how important it is for the ensuing chromosome segregation are unknown. Here we show that in the budding yeast Saccharomyces cerevisiae, activation of the phosphatase Cdc14 at anaphase onset is both necessary and sufficient for silencing microtubule dynamics. Cdc14 is activated by separase, the protease that triggers sister chromatid separation, linking the onset of anaphase to microtubule stabilization. If sister chromatids separate in the absence of Cdc14 activity, microtubules maintain high dynamic instability; this correlates with defects in both the movement of chromosomes to the spindle poles (anaphase A) and the elongation of the anaphase spindle (anaphase B). Cdc14 promotes localization of microtubule stabilizing proteins to the anaphase spindle, and dephosphorylation of the kinetochore component Ask1 contributes to both the silencing of microtubule turnover and successful anaphase A. PMID- 15650745 TI - Scientists and societies. Building a regional postdoc community. PMID- 15650747 TI - XIAP inhibits caspase-3 and -7 using two binding sites: evolutionarily conserved mechanism of IAPs. AB - The X-linked inhibitor of apoptosis protein (XIAP) uses its second baculovirus IAP repeat domain (BIR2) to inhibit the apoptotic executioner caspase-3 and -7. Structural studies have demonstrated that it is not the BIR2 domain itself but a segment N-terminal to it that directly targets the activity of these caspases. These studies failed to demonstrate a role of the BIR2 domain in inhibition. We used site-directed mutagenesis of BIR2 and its linker to determine the mechanism of executioner caspase inhibition by XIAP. We show that the BIR2 domain contributes substantially to inhibition of executioner caspases. A surface groove on BIR2, which also binds to Smac/DIABLO, interacts with a neoepitope generated at the N-terminus of the caspase small subunit following activation. Therefore, BIR2 uses a two-site interaction mechanism to achieve high specificity and potency for inhibition. Moreover, for caspase-7, the precise location of the activating cleavage is critical for subsequent inhibition. Since apical caspases utilize this cleavage site differently, we predict that the origin of the death stimulus should dictate the efficiency of inhibition by XIAP. PMID- 15650748 TI - Elf5 is essential for early embryogenesis and mammary gland development during pregnancy and lactation. AB - Elf5 is an epithelial-specific ETS factor. Embryos with a null mutation in the Elf5 gene died before embryonic day 7.5, indicating that Elf5 is essential during mouse embryogenesis. Elf5 is also required for proliferation and differentiation of mouse mammary alveolar epithelial cells during pregnancy and lactation. The loss of one functional allele led to complete developmental arrest of the mammary gland in pregnant Elf5 heterozygous mice. A quantitative mRNA expression study and Western blot analysis revealed that decreased expression of Elf5 correlated with the downregulation of milk proteins in Elf5(+/-) mammary glands. Mammary gland transplants into Rag(-/-) mice demonstrated that Elf5(+/-) mammary alveolar buds failed to develop in an Elf5(+/+) mammary fat pad during pregnancy, demonstrating an epithelial cell autonomous defect. Elf5 expression was reduced in Prolactin receptor (Prlr) heterozygous mammary glands, which phenocopy Elf5(+/ ) glands, suggesting that Elf5 and Prlr are in the same pathway. Our data demonstrate that Elf5 is essential for developmental processes in the embryo and in the mammary gland during pregnancy. PMID- 15650749 TI - Inducible DNA-loop formation blocks transcriptional activation by an SV40 enhancer. AB - It is well established that gene expression in eukaryotes is controlled by sequence-dependent binding of trans-acting proteins to regulatory elements like promoters, enhancers or silencers. A less well understood level of gene regulation is governed by the various structural and functional states of chromatin, which have been ascribed to changes in covalent modification of core histone proteins. And, much on how topological domains in the genome take part in establishing and maintaining distinct gene expression patterns is still unknown. Here we present a set of regulatory proteins that allow to reversibly alter the DNA structure in vivo and in vitro by adding low molecular weight effectors that control their oligomerization and DNA binding. Using this approach, we completely regulate the activity of an SV40 enhancer in HeLa cells by reversible loop formation to topologically separate it from the promoter. This result establishes a new mechanism for DNA-structure-dependent gene regulation in vivo and provides evidence supporting the structural model of insulator function. PMID- 15650750 TI - Tyrosine phosphatase SHP-2 is a mediator of activity-dependent neuronal excitotoxicity. AB - Calcium influx can promote neuronal differentiation and survival, at least in part by activating Ras and its downstream targets, including the Erk pathway. However, excessive calcium influx can initiate molecular signals leading to neuronal death during excitotoxicity or in neurodegenerative diseases. Here we describe a new signaling pathway associated with calcium influx that contributes to neuronal cell death in cerebellar neurons. Influx of calcium, mediated either by L-type voltage-sensitive calcium channels or glutamate receptors, is associated with the suppression of brain-derived neurotrophic factor (BDNF) activation of Ras and its effectors Erk and Akt. This is the result of enhanced association of the tyrosine phosphatase Shp-2 with TrkB receptors, which inhibits BDNF-induced TrkB autophosphorylation and activation. Deletion of the Shp2 gene in neuronal cultures reverses inhibition of TrkB function and increases neuronal survival after extended depolarization or glutamate treatment. These findings implicate Shp-2 in a feedback system initiated by calcium that negatively regulates neurotrophin signaling and sensitizes neurons to excitotoxicity. PMID- 15650751 TI - Functional analysis of a structural model of the ATP-binding site of the KATP channel Kir6.2 subunit. AB - ATP-sensitive potassium (KATP) channels couple cell metabolism to electrical activity by regulating K+ flux across the plasma membrane. Channel closure is mediated by ATP, which binds to the pore-forming subunit (Kir6.2). Here we use homology modelling and ligand docking to construct a model of the Kir6.2 tetramer and identify the ATP-binding site. The model is consistent with a large amount of functional data and was further tested by mutagenesis. Ligand binding occurs at the interface between two subunits. The phosphate tail of ATP interacts with R201 and K185 in the C-terminus of one subunit, and with R50 in the N-terminus of another; the N6 atom of the adenine ring interacts with E179 and R301 in the same subunit. Mutation of residues lining the binding pocket reduced ATP-dependent channel inhibition. The model also suggests that interactions between the C terminus of one subunit and the 'slide helix' of the adjacent subunit may be involved in ATP-dependent gating. Consistent with a role in gating, mutations in the slide helix bias the intrinsic channel conformation towards the open state. PMID- 15650752 TI - Notch1 antiapoptotic activity is abrogated by caspase cleavage in dying T lymphocytes. AB - Excessive signaling via the Notch1 receptor inhibits apoptosis in T lymphocytes. Since several antiapoptotic proteins are cleaved by caspases during cell death, we investigated whether Notch1 was a caspase substrate. Results demonstrate that the intracellular domain of Notch1 (NICD) is cleaved into six fragments during apoptosis in Jurkat cells or peripheral T lymphocytes. Notch1 cleavage is prevented by the caspase inhibitors DEVD-fmk and VEID-fmk or by Bcl-2 expression. Caspase-3 and caspase-6 cleave the NICD into six fragments using sites located within the NF-kappaB binding domain, the ankyrin repeats and the transactivation domain. Notch1 cleavage correlates with the loss of HES-1 expression in apoptotic T cells. Notch1 fragments cannot inhibit activation-induced cell death in a T cell hybridoma, confirming the abrogation of Notch1 antiapoptotic activity by caspases. The ability of the NICD but not the fragments to antagonize Nur77 activity supports a role for this factor in Notch1 antiapoptotic function. PMID- 15650753 TI - Cardiolipin and phosphatidylglycerol are not required for the in vivo action of Bcl-2 family proteins. PMID- 15650755 TI - Mechanism of mitochondrial stress-induced resistance to apoptosis in mitochondrial DNA-depleted C2C12 myocytes. AB - In this study, we show that partial mitochondrial DNA (mtDNA) depletion (mitochondrial stress) induces resistance to staurosporine (STP)-mediated apoptosis in C2C12 myoblasts. MtDNA-depleted cells show a 3-4-fold increased proapoptotic proteins (Bax, BAD and Bid), markedly increased antiapoptotic Bcl-2, and reduced processing of p21 Bid to active tBid. The protein levels and also the ability to undergo STP-mediated apoptosis were restored in reverted cells containing near-normal mtDNA levels and restored mitochondrial transmembrane potential. Inhibition of apoptosis closely correlated with sequestration of Bax, Bid and BAD in the mitochondrial inner membrane, increased Bcl-2 and Bcl-X(L), and inability to process p21 Bid. These factors, together with the reduced activation of caspases 3, 9 and 8 are possible causes of mitochondrial stress induced resistance to apoptosis. Our results suggest that a highly proliferative and invasive behavior of mtDNA-depleted C2C12 cells is related to their resistance to apoptosis. PMID- 15650754 TI - ATR and GADD45alpha mediate HIV-1 Vpr-induced apoptosis. AB - The human immunodeficiency virus type-1 (HIV-1) accessory gene vpr encodes a conserved 96-amino-acid protein that is necessary and sufficient for the HIV-1 induced block of cellular proliferation. Expression of vpr in CD4+ lymphocytes results in G2 arrest, followed by apoptosis. In a previous study, we identified the ataxia telangiectasia-mutated (ATM) and Rad3-related protein (ATR) as a cellular factor that mediates Vpr-induced cell cycle arrest. In the present study, we report that the breast cancer-associated protein-1 (BRCA1), a known target of ATR, is activated in the presence of Vpr. In addition, the gene encoding the growth arrest and DNA damage-45 protein alpha (GADD45alpha), a known transcriptional target of BRCA1, is upregulated by Vpr in an ATR-dependent manner. We demonstrate that RNAi-mediated silencing of either ATR or GADD45alpha leads to nearly complete suppression of the proapoptotic effect of Vpr. Our results support a model in which Vpr-induced apoptosis is mediated via ATR phosphorylation of BRCA1, and consequent upregulation of GADD45alpha. PMID- 15650756 TI - BI-1 protects cells from oxygen glucose deprivation by reducing the calcium content of the endoplasmic reticulum. PMID- 15650757 TI - Operating conditions for ocular surgery under general anaesthesia: an eccentric problem. AB - PURPOSE: To describe and examine the observation that eccentric eye positions may develop under certain general anaesthetic conditions and negatively impact upon operating conditions during ocular surgery. METHODS: A case series description with historical review followed by a 2-week prospective observational survey examining the general anaesthetic techniques utilized during ocular surgery and the associated development of an eccentric eye position. RESULTS: A total of 52 cases were studied, of which four developed eccentric eye positions. 50% of patients received effective muscle relaxation. Without muscle relaxants, the incidence of eccentric eye positioning was 18%. CONCLUSIONS: Eccentric ocular positions are likely to occur in light planes of general anaesthesia. In the case of ocular surgery, this phenomenon may create surgical difficulty and increase the risk of complication and patient morbidity. Arguments for the planned use of balanced general anaesthesia with non-depolarizing muscle relaxants are presented. PMID- 15650758 TI - Moderate to severe obstructive sleep apnoea patients is associated with a higher incidence of visual field defect. AB - PURPOSE: To compare the visual fields (VFs) and optic nerve head changes between obstructive sleep apnoea (OSA) in normotensive patients and an age-matched non OSA population. DESIGN: Case-control study. PARTICIPANTS: A total of 41 ethnic Chinese patients diagnosed with moderate to severe OSA referred from the Sleep Laboratory, ENT Department, Tuen Mun Hospital. A total of 35 age-matched non-OSA subjects recruited from the Ophthalmology Department, North District Hospital. METHODS: Comprehensive ophthalmological and systemic history, complete ophthalmological examination, including central-30 computerized perimetry for all studied patients. MAIN OUTCOME MEASURES: Polysomnographic data, VF indices, optic disc changes. RESULTS: In the OSA arm, VF indices were significantly subnormal and the incidence of suspicious glaucomatous disc changes was four times higher than that of the control arm. None of the studied patients suffered from any form of anterior segment complications. CONCLUSIONS: Moderate to severe OSA is associated with a higher incidence of VF defect and glaucomatous optic nerve changes. PMID- 15650759 TI - Pointwise linear progression criteria and the detection of visual field change in a glaucoma trial. AB - BACKGROUND: Current pointwise linear regression (PLR) change criteria for visual field analysis are largely empirical. METHODS: Two independent sets of Humphrey Field Analyzer fields were analysed using PLR. Set i, 56 patients, and set ii, 97 patients, were followed over 16 months. Criteria were tested against set i, and then validated using set ii. Each criterion specified a fixed critical slope of 1 dB/year and with a range of significance from P<0.001 to 0.05. The criteria were varied by altering location number, cluster arrangement, and by requiring points to show change over both 12 and 16 months. True glaucomatous change was differentiated from noise by looking for exclusive progression (EP), the detection of progression without detection of improvement. RESULTS: Set i required 1 point to have a slope of 1 dB/year and P<0.05 labelled 64% progressing and 58% improving, whereas several stricter criteria were capable of detecting EP. Two points in a perimetric nerve fibre bundle (PNFB) cluster gave optimal EP detection, labelling 8.9% progressing in set i and 7.2% progressing in set ii with a cutoff P-value of 0.026 inset i and 0.013 inset ii. CONCLUSION: Lax PLR criteria detect large amounts of change. Validating criteria using two data sets allow selection of better criteria, capable of detecting EP. The criterion involving 2 points changing in a PNFB cluster offers the best option for exclusively detecting progression. PMID- 15650760 TI - Cerebral haemodynamics in patients with pseudoexfoliation glaucoma. AB - PURPOSE: To evaluate the cerebral haemodynamics in patients with pseudoexfoliation glaucoma. METHODS: A total of 19 consecutive patients with pseudoexfoliation glaucoma and 19 age- and sex-matched healthy volunteers were recruited in a prospective comparative study. The affected eye in patients with unilateral glaucoma and the eye with more advanced lesions in patients with bilateral glaucoma were included in the study, and the ipsilateral middle cerebral arteries (MCA) were evaluated. While in the controls, the study eye and the ipsilateral MCA were chosen randomly. Measurements included systemic arterial pressure, heart rate, intraocular pressure (IOP), and transcranial colour Doppler parameters, including peak systolic velocity (PSV), end-diastolic velocity (EDV), time-averaged maximum velocity (TAMAX), resistivity index (RI), and pulsatility index (PI). RESULTS: The mean PSV, EDV, and TAMAX were significantly lower in the patients with pseudoexfoliation glaucoma than in healthy controls (P<0.001, <0.001 and <0.001, respectively). Additionally, patients with pseudoexfoliation glaucoma had significantly higher PI and RI than healthy controls (P=0.001 and 0.04, respectively). Systolic and diastolic blood pressures (P=0.19 and 0.91, respectively) and heart rate (P=0.06) were not different between the groups. The mean IOP were significantly higher in the glaucoma patients compared with the controls (P<0.001). CONCLUSION: This study suggests that pseudoexfoliation glaucoma is associated with a reduction in the blood flow velocity and elevation in the resistance of the MCA. PMID- 15650761 TI - Basal cell carcinoma of the eyelid in children: a report of three cases. AB - Basal cell carcinoma is the most common eyelid malignancy, accounting for approximately 90% of malignant eyelid tumours. Despite its prevalence in adults, it is extremely rare in children, and usually occurs in the setting of a known genetic defect or following radiotherapy treatment. We report three cases of de novo basal cell carcinoma in children. These children had no known genetic syndromes and had not undergone radiotherapy. PMID- 15650762 TI - Bee sting-induced ciliochoroidal detachment. PMID- 15650763 TI - Tumor cells as cellular vehicles to deliver gene therapies to metastatic tumors. AB - A long-pursued goal in cancer treatment is to deliver a therapy specifically to metastases. As a result of the disseminated nature of the metastatic disease, carrying the therapeutic agent to the sites of tumor growth represents a major step for success. We hypothesized that tumor cells injected intravenously (i.v.) into an animal with metastases would respond to many of the factors driving the metastatic process, and would target metastases. Using a model of spontaneous metastases, we report here that i.v. injected tumor cells localized on metastatic lesions. Based on this fact, we used genetically transduced tumor cells for tumor targeting of anticancer agents such as a suicide gene or an oncolytic virus, with evident antitumoral effect and negligible systemic toxicity. Therefore, autologous tumor cells may be used as cellular vehicles for systemic delivery of anticancer therapies to metastatic tumors. PMID- 15650764 TI - Restoring DNA repair capacity of cells from three distinct diseases by XPD gene recombinant adenovirus. AB - The nucleotide excision repair (NER) is one of the major human DNA repair pathways. Defects in one of the proteins that act in this system result in three distinct autosomal recessive syndromes: xeroderma pigmentosum (XP), Cockayne syndrome (CS) and trichothiodystrophy (TTD). TFIIH is a nine-protein complex essential for NER activity, initiation of RNA polymerase II transcription and with a possible role in cell cycle regulation. XPD is part of the TFIIH complex and has a helicase function, unwinding the DNA in the 5' --> 3' direction. Mutations in the XPD gene are found in XP, TTD and XP/CS patients, the latter exhibiting both XP and CS symptoms. Correction of DNA repair defects of these cells by transducing the complementing wild-type gene is one potential strategy for helping these patients. Over the last years, adenovirus vectors have been largely used in gene delivering because of their efficient transduction, high titer, and stability. In this work, we present the construction of a recombinant adenovirus carrying the XPD gene, which is coexpressed with the EGFP reporter gene by an IRES sequence, making it easier to follow cell infection. Infection by this recombinant adenovirus grants full correction of SV40-transformed and primary skin fibroblasts obtained from XP-D, TTD and XP/CS patients. PMID- 15650765 TI - Modulation of retrovirally driven therapeutic genes by mutant TP53 in anaplastic thyroid carcinoma. AB - We previously demonstrated that restoration of TP53 activity in anaplastic thyroid carcinoma inhibits cell growth and induces expression of thyroid differentiation markers. Here, we investigated whether TP53 status may condition the expression of therapeutic genes driven by retroviral LTR or tissue-specific enhancer elements. The TP53-defective ARO anaplastic thyroid carcinoma cells were transfected with TP53(Val135), which exhibits wild-type activity at 32 degrees C, and transduced with retroviral vectors, in which therapeutic genes were driven either by wild-type LTR or by a reshuffled LTR containing thyroglobulin (TG) enhancer. Both at 37 and 32 degrees C, expression of transgenes driven by TG enhancer was 10-fold lower than that obtained with wild-type LTR retroviral vector. TP53(Val135) transfer into ARO cells repressed transcription from wild type LTR but increased expression of TG-driven therapeutic genes. This effect was markedly enhanced by cell culture at 32 degrees C and by TSH treatment. Cytotoxic effects shown after ganciclovir treatment paralleled therapeutic gene expression levels. In conclusion, TP53 status in the tumor cell can influence expression of therapeutic genes. When using retroviral-vector-based gene therapy, wild-type LTR vectors should be employed to target TP53-defective tumors, whereas thyroid specific promoters should be used for transcriptional targeting of thyroid carcinomas carrying wild-type TP53. PMID- 15650766 TI - Delta24-hyCD adenovirus suppresses glioma growth in vivo by combining oncolysis and chemosensitization. AB - Replication-competent adenoviruses could provide an efficient method for delivering therapeutic genes to tumors. The most promising strategies among adenovirus-based oncolytic systems are designed to exploit free E2F-1 activity in cancer cells, which in the absence of pRb activates transcription and regulates the expression of genes involved in differentiation, proliferation, and apoptosis. We previously developed Delta24, an E1A-mutant, conditionally replicative oncolytic adenovirus. Here, we examine the ability of a second generation Delta24 (Delta24-hyCD) engineered to express a humanized form of the Saccharomyces cerevisiae cytosine deaminase gene (hyCD). Real-time quantitative PCR, Western blotting, thin-layer chromatography, and radioisotope quantitative enzymatic assays confirmed the production of a catalytically active hyCD enzyme in the setting of an oncolytic infection in vitro; other experiments assessing local production of 5-fluorouracil and a concomitant bystander effect showed improved cytotoxicity. The IC50 dose of 5-fluorocytosine (5-FC) required for a complete cytopathic effect by the Delta24-hyCD virus was fivefold lower than with Delta24 alone in U251MG and U87MG malignant glioma (MG) cell lines. Intratumoral treatment of mice bearing intracranial U87MG xenografts with Delta24-hyCD+5-FC significantly improved survival, confirming that Delta24-hyCD with 5-FC is a more efficient anticancer tool than Delta24 alone. Histopathologically, Delta24-hyCD replication was accompanied by progressively augmented oncolysis and drug-induced necrosis. These findings demonstrate that Delta24-hyCD with concomitant systemic 5-FC is a significant improvement over the earlier Delta24 oncolytic tumor selective strategy for therapy of experimental gliomas. PMID- 15650767 TI - Development of a hypoxia-inducible cytosine deaminase expression vector for gene directed prodrug cancer therapy. AB - One important feature of human solid tumors is the presence of a hypoxic microenvironment. Under hypoxia, genes that contain a hypoxia-response element (HRE) can be activated by the binding of hypoxia-inducible factor-1. To reach the goal of selectively killing tumor cells in a hypoxic microenvironment using a gene therapy approach, we developed a cytosine deaminase (CD) gene construct (pH9YCD2) that contains an HRE gene enhancer. CD is an enzyme that catalyzes the conversion of noncytotoxic 5-fluorocytosine (5-FC) to the cytotoxic and radiosensitizing drug 5-fluorouracil (5-FU). Yeast CD was cloned into an SV40 promoter-based mammalian expression vector, and an HRE enhancer was inserted in front of the promoter. Human glioblastoma U-87 MG cells were transfected with pH9YCD2. Western blots revealed that CD was strongly expressed under hypoxic conditions (0.3-1% O2), whereas only minor CD expression was seen under normoxic conditions. To confirm that the expressed CD enzyme retains catalytic activity, we performed a 5-FC/5-FU-conversion assay in which 5-FC was incubated with the lysates of pH9YCD2-transfected cells. The percentage of conversion from 5-FC to 5 FU was 63% under hypoxia versus 13% under normoxia. In vitro, cell viability and colony-forming efficiency assays demonstrated that the gene construct was able to significantly kill glioblastoma cells in a hypoxia-dependent manner. In addition, 5-FC treatment of hypoxic pH9YCD2-transfected cells produced a marked bystander effect, which could be a distinct advantage for gene therapy. If this construct exhibits antitumor efficacy in vivo, it may have promise as an antitumor agent in humans. PMID- 15650768 TI - Macrophage inflammatory protein-1alpha as a costimulatory signal for mast cell mediated immediate hypersensitivity reactions. AB - Regulation of the immune response requires the cooperation of multiple signals in the activation of effector cells. For example, T cells require signals emanating from both the TCR for antigen (upon recognition of MHC/antigenic peptide) and receptors for costimulatory molecules (e.g., CD80 and CD60) for full activation. Here we show that IgE-mediated reactions in the conjunctiva also require multiple signals. Immediate hypersensitivity reactions in the conjunctiva were inhibited in mice deficient in macrophage inflammatory protein-1alpha (MIP-1alpha) despite normal numbers of tissue mast cells and no decrease in the levels of allergen specific IgE. Treatment of sensitized animals with neutralizing antibodies with specificity for MIP-1alpha also inhibited hypersensitivity in the conjunctiva. In both cases (MIP-1alpha deficiency and antibody treatment), the degranulation of mast cells in situ was affected. In vitro sensitization assays showed that MIP 1alpha is indeed required for optimal mast cell degranulation, along with cross linking of the high-affinity IgE receptor, FcepsilonRI. The data indicate that MIP-1alpha constitutes an important second signal for mast cell degranulation in the conjunctiva in vivo and consequently for acute-phase disease. Antagonizing the interaction of MIP-1alpha with its receptor CC chemokine receptor 1 (CCR1) or signal transduction from CCR1 may therefore prove to be effective as an antiinflammatory therapy on the ocular surface. PMID- 15650769 TI - Dynamic changes in Mcl-1 expression regulate macrophage viability or commitment to apoptosis during bacterial clearance. AB - Macrophages are critical effectors of bacterial clearance and must retain viability, despite exposure to toxic bacterial products, until key antimicrobial functions are performed. Subsequently, host-mediated macrophage apoptosis aids resolution of infection. The ability of macrophages to make this transition from resistance to susceptibility to apoptosis is important for effective host innate immune responses. We investigated the role of Mcl-1, an essential regulator of macrophage lifespan, in this switch from viability to apoptosis, using the model of pneumococcal-associated macrophage apoptosis. Upon exposure to pneumococci, macrophages initially upregulate Mcl-1 protein and maintain viability for up to 14 hours. Subsequently, macrophages reduce expression of full-length Mcl-1 and upregulate a 34-kDa isoform of Mcl-1 corresponding to a novel BH3-only splice variant, Mcl-1(Exon-1). Change in expression of Mcl-1 protein is associated with mitochondrial membrane permeabilization, which is characterized by loss of mitochondrial inner transmembrane potential and translocation of cytochrome c and apoptosis-inducing factor. Following pneumococcal infection, macrophages expressing full-length human Mcl-1 as a transgene exhibit a delay in apoptosis and in bacterial killing. Mcl-1 transgenic mice clear pneumococci from the lung less efficiently than nontransgenic mice. Dynamic changes in Mcl-1 expression determine macrophage viability as well as antibacterial host defense. PMID- 15650770 TI - Role of Gas6 receptors in platelet signaling during thrombus stabilization and implications for antithrombotic therapy. AB - Mechanisms regulating thrombus stabilization remain largely unknown. Here, we report that loss of any 1 of the Gas6 receptors (Gas6-Rs), i.e., Tyro3, Axl, or Mer, or delivery of a soluble extracellular domain of Axl that traps Gas6 protects mice against life-threatening thrombosis. Loss of a Gas6-R does not prevent initial platelet aggregation but impairs subsequent stabilization of platelet aggregates, at least in part by reducing "outside-in" signaling and platelet granule secretion. Gas6, through its receptors, activates PI3K and Akt and stimulates tyrosine phosphorylation of the beta3 integrin, thereby amplifying outside-in signaling via alphaIIbbeta3. Blocking the Gas6-R-alphaIIbbeta3 integrin cross-talk might be a novel approach to the reduction of thrombosis. PMID- 15650771 TI - Chronic lymphocytic leukemia B cells contain anomalous Lyn tyrosine kinase, a putative contribution to defective apoptosis. AB - B cell chronic lymphocytic leukemia (B-CLL) is a neoplastic disorder characterized by accumulation of B lymphocytes due to uncontrolled growth and resistance to apoptosis. Analysis of B cells freshly isolated from 40 patients with chronic lymphocytic leukemia demonstrated that the Src kinase Lyn, the switch molecule that couples the B cell receptor to downstream signaling, displays anomalous properties. Lyn is remarkably overexpressed at the protein level in leukemic cells as compared with normal B lymphocytes, with a substantial aliquot of the kinase anomalously present in the cytosol. Whereas in normal B lymphocytes Lyn activation is dependent on B cell-receptor stimulation, in resting malignant cells, the constitutive activity of the kinase accounts for high basal protein tyrosine phosphorylation and low responsiveness to IgM ligation. Addition of the Lyn inhibitors PP2 and SU6656 to leukemic cell cultures restores cell apoptosis, and treatment of malignant cells with drugs that induce cell apoptosis decreases both activity and amount of the tyrosine kinase. These findings suggest a direct correlation between high basal Lyn activity and defects in the induction of apoptosis in leukemic cells. They also support a critical role for Lyn in B-CLL pathogenesis and identify this tyrosine kinase as a potential therapeutic target. PMID- 15650772 TI - Muscle-specific expression of IGF-1 blocks angiotensin II-induced skeletal muscle wasting. AB - Advanced congestive heart failure is associated with activation of the renin angiotensin system and skeletal muscle wasting. We previously showed that angiotensin II infusion in rats produces cachexia secondarily to increased muscle proteolysis and also decreases levels of circulating and skeletal muscle IGF-1. Here we show that angiotensin II markedly downregulates phospho-Akt and activates caspase-3 in skeletal muscle, leading to actin cleavage, an important component of muscle proteolysis, and to increased apoptosis. These changes are blocked by muscle-specific expression of IGF-1, likely via the Akt/mTOR/p70S6K signaling pathway. We also demonstrate that mRNA levels of the ubiquitin ligases atrogin-1 and muscle ring finger-1 are upregulated in angiotensin II-infused WT, but not in IGF-1-transgenic, mice. These findings strongly suggest that angiotensin II downregulation of IGF-1 in skeletal muscle is causally related to angiotensin II induced wasting. Because the renin-angiotensin system is activated in many catabolic conditions, our findings have broad implications for understanding mechanisms of skeletal muscle wasting and provide a rationale for new therapeutic approaches. PMID- 15650773 TI - MyD88-dependent induction of allergic Th2 responses to intranasal antigen. AB - MyD88 is a common Toll-like receptor (TLR) adaptor molecule found to be essential for induction of adaptive Th1 immunity. Conversely, innate control of adaptive Th2 immunity has been shown to occur in a MyD88-independent manner. In this study, we show that MyD88 is an essential innate component in the induction of TLR4-dependent Th2 responses to intranasal antigen; thus we demonstrate what we believe to be a novel role for MyD88 in pulmonary Th2 immunity. Induction of the MyD88-independent type I IFN response to LPS is defective in the pulmonary environment. Moreover, in the absence of MyD88, LPS-induced upregulation of costimulatory molecule expression on pulmonary DCs is defective, in contrast to what has been observed with bone marrow-derived DCs (BMDCs). Reconstitution of Th2 responses occurs upon adoptive pulmonary transfer of activated BMDCs to MyD88 deficient recipients. Furthermore, the dependence of Th2 responses on MyD88 is governed by the initial route of antigen exposure; this demonstrates what we believe are novel site-specific innate mechanisms for control of adaptive Th2 immunity. PMID- 15650775 TI - [Treatment of AIDS patients with Chinese medicinal herbs qudu zengning capsule]. AB - OBJECTIVE: To evaluate the therapeutic effect of QuDu ZengNing Capsule on AIDS. METHODS: QuDu ZengNing Capsule is a capsule containing extract from 4 Chinese medicinal herbs. Totally 1,000 AIDS patients were treated, among them 60 patients were clinically observed weekly. Blood routine tests, liver, heart and kidney function, X-ray, CD4, CD8 cells were examined before and after treatment at 1, 3, 6 month. The patients were treated with 4 capsules t.i.d for 6 months. RESULTS: The symptoms were improved in most of the patients, the CD4 cells increased from 115.0 to 295.2/ul and the viral load (RNA copies/ml) in most patients reduced markedly or maintained at the same level. CONCLUSION: These data indicated that QuDu ZengNing Capsule was effective for treatment of AIDS patients. PMID- 15650774 TI - Tamm-Horsfall glycoprotein links innate immune cell activation with adaptive immunity via a Toll-like receptor-4-dependent mechanism. AB - Tamm-Horsfall glycoprotein (THP) is expressed exclusively in the kidney and constitutes the most abundant protein in mammalian urine. A critical role for THP in antibacterial host defense and inflammatory disorders of the urogenital tract has been suggested. We demonstrate that THP activates myeloid DCs via Toll-like receptor-4 (TLR4) to acquire a fully mature DC phenotype. THP triggers typical TLR signaling, culminating in activation of NF-kappaB. Bone marrow-derived macrophages from TLR4- and MyD88-deficient mice were nonresponsive to THP in contrast to those from TLR2- and TLR9-deficient mice. In vivo THP-driven TNF alpha production was evident in WT but not in Tlr4-/- mice. Importantly, generation of THP-specific Abs consistently detectable in urinary tract inflammation was completely blunted in Tlr4-/- mice. These data show that THP is a regulatory factor of innate and adaptive immunity and therefore could have significant impact on host immunity in the urinary tract. PMID- 15650776 TI - [Prevalence of drug resistance mutations among antiretroviral drug-naive HIV-1 infected patients in China]. AB - OBJECTIVE: To collect background information on drug-resistant HIV-1 strains in various regions before the start of nation-wide antiretroviral therapy in China. METHODS: Twenty percent of the 2,000 blood samples from antiretroviral therapy naive patients collected for the 2nd national HIV molecular epidemiology survey (NHMES) in 2002 were randomly sampled for this study. The entire protease gene and 20-230 amino acids of the reverse transcriptase gene were amplified by PCR from provirus DNA and sequenced. The results were analyzed with HIV db-Drug Resistance Algorithm and genotypic resistance mutations were determined to particular anti-HIV drugs. RESULTS: Totally 164 protease gene sequences and 138 reverse transcriptase gene sequences were obtained from patients; 0.61% of 164 sequences displayed primary resistance mutations in the protease gene, whereas 99.39% carried 1 or more secondary mutations. Genotypic resistance to at least one nucleoside reverse transcriptase inhibitors (NRTI) was present in 5.80%,and resistance to at least one non-nucleo side reverse transcriptase inhibitors (NNRTI) was present in 1.45% of samples. CONCLUSION: The prevalence of genotypic drug resistance is very low in drug-naive HIV infected patients from 21 provinces of China tested in this study. Laboratories participated in the NHMES have organized a network to provide drug resistance monitoring service in the current nation-wide antiviral treatment program in China. PMID- 15650777 TI - [Immune potency of recombinant adeno-associated virus combined with recombinant adenovirus vaccine containing HIV-1 gp120]. AB - OBJECTIVE: To study the immune effect of recombinant adeno-associated virus (rAAV) combined with recombinant adenovirus (rAdV) vaccine in BALB/c mice. METHODS: The codon-modified HIV-1 gp120 gene was inserted into plasmid of adeno associated virus and adenovirus vector separately. Then the rAAV and rAdV vaccines were constructed. BALB/c mice were immunized with rAAV and rAdV vaccines in different administration scheme. The IgG antibody was detected by ELISA and CTL response was detected by intracellular cytokine stain assay. RESULTS: Both rAAV and rAdV vaccine could express gp120 gene; the mice primed with rAAV at week 0, 2 and boosted with rAdV at week 5, 14 and 20 elicited the strongest gp120 specific CTL and IgG antibody response. CONCLUSION: The mice primed with rAAV and boosted with rAdV could elicit specific CTL response and IgG antibody. PMID- 15650778 TI - [Preparation and characterization of monoclonal antibodies against SARS associated coronavirus nucleocapsid protein]. AB - OBJECTIVE: To obtain monoclonal antibodies (McAbs) against severe acute respiratory syndrome (SARS) associated coronavirus (SARS-CoV) nucleocapsid (N) protein to develop diagnostic test for SARS and study the pathogenesis of the disease. METHODS: BALB/c mice were immunized with purified N protein of SARS-CoV. Hybridoma cell lines secreting monoclonal antibodies against SARS-associated coronavirus nucleocapsid were established after cell fusion with mouse splenic cells and SP2/0 cells. The specificity of the McAbs obtained was examined by Western blot and indirect fluorescence assay. Epitopes reacted with the McAbs were preliminarily located through Western blot by expressing truncated N proteins. RESULTS: After cell fusion and three rounds of cell cloning, six hybridoma cell lines secreting monoclonal antibodies specifically against SARS CoV nucleocapsid were obtained. Western blot and indirect fluorescence assay showed that the McAbs reacted specifically with nucleocapsid protein and SARS CoV. Among the six McAbs, three recognize the epitopes located in the N-terminus of the protein, whereas the others reacted with those located in the C-terminus. CONCLUSION: The anti-SARS-CoV nucleocapsid McAbs were developed and these McAbs may be useful in the development of diagnosis assays and basic research of SARS. PMID- 15650779 TI - [Establishment of the reference panel for HIV RNA]. AB - OBJECTIVE: To establish a national reference panel for HIV RNA diagnostic reagents. METHODS: Sera from patients with HIV infection and healthy blood donors were collected and tested for HIV and HCV antibodies and HBsAg by using ELISA. The HIV antibody positive samples with ELISA were confirmed with HIV Blot 2.2 (Genelabs). The quantitative samples for HIV RNA were calibrated with the WHO HIV RNA standard. The stability of the panel was evaluated with acceleration method. RESULTS: After screening and calibration, 8 negative samples, 8 positive samples, 3 quantitative samples, 6 sensitivity samples and 5 samples for linear analysis were composed of the national reference panel for HIV RNA. The convinced international units (IU) for the quantitative samples were obtained by seven independent calibration and the logarithm of international units for the quantitative samples (b1-b3) were less than x +/- s. The results showed that this panel may stabilize for 4 days at 4 degrees C. CONCLUSION: A national reference panel for HIV RNA reagents has been established. It may provide the basis for evaluating HIV RNA diagnostic reagents. PMID- 15650780 TI - [Sero-epidemiological characteristics of antibody to SARS-associated coronavirus in Gansu Province]. AB - OBJECTIVE: To study the levels of SARS-associated coronavirus IgG antibody of SARS patients, people who closely contacted SARS patients and normal subjects in Gansu province. METHODS: The levels of SARS-associated coronavirus IgG antibody were measured by ELISA. The material included acute and (or) recovery period sera of 9 SARS patients, sera from 1,109 doctors and nurses closely contacted with SARS patients, laboratory workers, personnel for disease control and prevention, persons who contacted SARS patients, and sera from 978 normal subjects. RESULTS: SARS coronavirus IgG antibody was detected positive in 6 of the 9 patients, it was still positive in the sera twelve months after recovery; 1 of the closely contacted persons and 3 normal subjects were found positive. CONCLUSION: The positive rate of SARS coronavirus IgG antibody of patients was consistent with the clinical diagnosis. The low positive rate of the persons who closely contacted SARS patients and normal subjects suggests that SARS probably had no subclinical infection. PMID- 15650781 TI - [Dynamic expression of matrix metalloproteinase-2, membrane type-matrix metalloproteinase-2 in experimental hepatic fibrosis and its reversal in rat]. AB - OBJECTIVE: To investigate the expression dynamics and significance of matrix metalloproteinase-2 (MMP-2) membrane type-matrix metalloproteinase-2 (MT-MMP-2) in hepatic fibrosis and its reversal counterpart. METHODS: An experimental CCl4 induced hepatic fibrosis rat model was established by intraperitoneal administration of carbon tetrachloride for 2, 4, 6, 8, 10 weeks, and normal rats were used as a control group. The immunohistochemical methods and in situ hybridization were used to detect MMP-2,MT-MMP-2 mRNA and related antigens in the liver. RESULTS: MMP-2,MT-MMP-2 mRNA and related antigens were expressed in mesenchymal cells and parts of hepatocytes besides active pathological changes, especially in the fibrous septum and portal area. Expression of MMP-2,MT-MMP-2 mRNA and related antigens were increased in hepatic fibrosis and decreased gradually in its reversal counterpart. CONCLUSION: This study suggested that mesenchymal cells are the main cellular origins of MMPs. The levels of MMP-2 and MT-MMP-2 antigens and gene expression were closely related to hepatic fibrosis. MMP-2 and MT-MMP-2 may play important roles in hepatic fibrosis and its reversal counterpart. PMID- 15650782 TI - [Retroviral vector-mediated HSVtk gene expression and acquisition of high titer recombinant virus]. AB - OBJECTIVE: To explore the HSVtk gene expression mediated by the retroviral vector and to obtain high titer recombinant retroviral virus. METHODS: The recombinant vector pRevTRE/HSVtk was constructed by inserting HSVtk gene into pRevTRE. The recombinant retrovirus, which was produced from cloned PA317 cells screened by hygromycin B after "micro-pingpong" technique transferring with pRevTRE/HSVtk plasmids DNA by using modified calcium phosphate precipitation method. HSVtk gene expression was performed on target cells and virus titers were detected in different cultured temper, time and sodium butyrate concentration. RESULTS: The recombinant retroviral vector pRevTRE/HSVtk was constructed and HSVtk gene expression was detected on target cells after they were infected with the recombinant retrovirus. CONCLUSION: High titer of retroviruses could be obtained in the culture medium of PA317 cell line through "micro-pingpong" technique at 30 hours and 10 mmol/L sodium butyrate concentration followed by frozen ultrafiltration. PMID- 15650783 TI - [Molecular epidemiological study on rubella virus]. AB - OBJECTIVE: To understand the hereditary and variant characteristics of rubella virus(RV), especially the strains isolated from China, investigating the epidemical trend and variation principle of RV. METHODS: The envelope glycoprotein E1 gene was amplified from rubella virus by RT-PCR. After sequencing, the gene sequence was handled by the software DNASTAR and the phylogenetic tree was drawn to analyze the molecular epidemiological characteristics of RV. RESULTS: The sequence of RV strain JR23 was sequenced, the phylogenetic tree was drawn taking 30 strains isolated at different times and locations in GenBank, including three strains from China as reference. The regions that encode the peptides which react with the HI antibody and the neutralization antibody were compared to show if there was any amino acid mutation in the sequence. (1) In general, the nucleotide and amino acid sequences of RV were highly conserved. The four strains isolated from China had relatively large variations. Strain 379 and strain BRD constituted genotype II, which is different from the other 29 strains. Further study is needed to understand their heritable resources and biological characteristics. (2) Strain JR23 showed little difference from the strains that were epidemic during 1960s in UK, USA and Japan, so maybe it is the derivative strain of that in epidemic 1960s. But the accurate epidemic time is not known. CONCLUSION: There are differences among areas and time in epidemics of rubella. The mobility and the region difference seem to be the key factors that affect the epidemic characteristics of RV. PMID- 15650784 TI - [Inhibition of hepatitis C virus gene expression by antisense nucleotide in vitro]. AB - OBJECTIVE: To study the mechanism of hepatitis C virus (HCV) gene regulation and the inhibitory effect of antisense RNA on HCV gene expression in vitro. METHODS: The hepatoblastoma cell line (HepG2) was co-transfected by recombinant plasmid of antisense RNA complementary to HCV 5' untranslated region (5'UTR)and HCV 5' UTR Directed luciferase (luc) gene expression recombinant plasmid. Meanwhile a reversed HCV 5'UTR recombinant plasmid which can not transcribe as antisense RNA in the cell and a recombinant plasmid in which the luc was regulated by simian virus 40 (sv40) 5'UTR were used as controls respectively. The level of luc gene expression was determined by an enzymatic assay. RESULTS: The antisense RNA which directed to HCV 5'UTRcould obviously knock down the level of luc gene expression and the close-dependent inhibition of antisense RNA was observed at the same time. However the above inhibition was not shown in the cells co-transfected by reversed HCV 5'UTR recombinant plasmid and HCV 5'UTR directed luc gene expression recombinant plasmid. No reduction was observed in luc gene expression level in the cell co-transfected by both antisense RNA recombinant plasmid and SV40 5'UTR directed luc gene expression recombinant plasmid. CONCLUSION: HCV 5'UTR plays an important role in regulation of viral gene expression. The antisense RNA complementary to HCV 5'UTR could effectively inhibit the gene expression regulated by HCV 5'UTR in vitro. PMID- 15650785 TI - [Immunoreactivity studies on synthetic peptides deriving from immunodominant region of hepatitis C virus NS5a gene]. AB - OBJECTIVE: To identify the location of major immunodominant antigenic region and study the relationship between the gene heterogeneity and immunoreactivity via detecting antigenic reactivity of synthetic peptides deriving from immunodominant region in different genotypes of hepatitis C virus (HCV) NS5a gene. METHODS: In total, 305 non-identical 30-mer long and overlapping by 15 aa peptides derived from HCV NS5a region from codon 2,182 to 2,343 among 45 unique published HCV sequences in GenBank corresponding to different genotype were designed and synthesized. The amino acid sequences of all peptides were compared with DNA Star software. The antigenic reactivity of those peptides was detected with indirect ELISA with both anti-HCV and anti-NS5 positive serum. RESULTS: The sequences showed highly conserved among HCV genotype in regions 2,272-2,301 and 2,302-2,331 as compared to regions 2,212-2,241 and 2,257-2,286. The peptides basing on amino acid residues among 2,212-2,241, 2,272-2,301 and 2,302-2,331 showed stronger immunoreactivity than any other peptides. Eighteen peptides derived from this region showed a broad immunoreactivity, 3 of them could react with 96% of anti HCV positive sera. Whereas the immunoreactivity of the peptides derived from the region showing highly variable among HCV genotype was found to react more strongly with homologous-genotype sera. CONCLUSION: The major linear antigenic region was located at amino acid residues among 2,212-2,241, 2,272-2,301 and 2,302-2,331; the short synthetic peptide derived from NS5a region at position 2,212-2,241, 2,272-2,301 and 2,302-2,331 can be used for efficient detection of HCV antibody; some peptides showed genotype specific immuunoreactivity. PMID- 15650786 TI - [Biological characteristics of the adapted rabies virus 3aG-V in Vero cell]. AB - OBJECTIVE: To study the biological features of the adapted rabies virus 3aG-V in Vero cell. METHODS: The virus structure, culture condition, pathogenicity, immunocompetence, toxicity and protective efficiency were studied. RESULTS: The rabies virus 3aG-V strain could well adapt to Vero cell, the pathogenicity was very weak and the immunogenicity was very good. CONCLUSION: The Vero cell-adapted rabies virus strain may possibly be used as a vaccine strain. PMID- 15650787 TI - [Relationship between the pathologic changes of human embryo fibroblast cells and expression of late mRNA after human cytomegalovirus infection in vitro]. AB - OBJECTIVE: To study the relationship between late mRNA and the cytopathic effect(CPE) and ultrastructural features after human cytomegalovirus (HCMV) infection in vitro. METHODS: Human embryo fibroblast cells(HEL) were infected with HCMV AD169 strain. The expression of the HCMV late mRNA was measured by semiquantitative RT-PCR, the cytopathic effect (CPE) and the cell ultrastructure were observed by means of light microscopy and electron microscopy, respectively. RESULTS: The HCMV late mRNA could be detected 12 hours postinfection and increased gradually, but the CPE appeared 48 hours postinfection in HEL cells. The HCMV infected cells exhibited significant mitochondrial enlargement and the number of mitochondrial ridge deletion, the cisternae lumen of endoplasmic reticulum (ER) dilation and vacuolization (at the end age). The mature nucleocapsid could be observed 96 hours postinfection. CONCLUSION: The ultrastructural changes have an intimate correlation with the expression of HCMV late mRNA and play an important role in the life circle of the virus. HCMV late mRNA may serve as a indicator of the clinical effect of treatment in active HCMV infection. PMID- 15650788 TI - [Evaluation of anti-HCV detection kits using recombinant antigens derived from various HCV regions]. AB - OBJECTIVE: To evaluate the first and second assay kits currently used in blood centers for screening HCV infected blood, and to provide basis for a better match of the two assay kits. METHODS: Using the newly developed multi-recombinant-HCV antigen supplementary assay kit, the authors evaluated concurrently the specificity and sensitivity of two domestic and one imported anti-HCV detection kits. RESULTS: Discrepancy in specificity and sensitivity existed among the two domestic HCV kits, and overall quality was slightly below that of leading or main stream imported HCV kit. CONCLUSION: The newly developed multi-recombinant-HCV antigen supplementary assay kit is useful in the evaluation of HCV antibody detection kit currently in use. It provides qualified assessing kit to capture antibodies against various HCV antigens. The present paper provided guidance for selecting a better match of the two screening kits and improved screening efficiency. PMID- 15650789 TI - [Cloning and characterization of a full-length HIV-1 genome of a prevalent subtype B-Thai strain in Henan Province]. AB - OBJECTIVE: To clone, identify and phylogenetically characterize a clade B-Thai HIV isolate representing the most prevalent virus in Henan province. METHODS: Peripheral blood mononuclear cells (PBMCs) from an HIV-1 infected patient in Henan Province were separated, and co-cultivated with phytohemagglutinin stimulated healthy donor PBMCs. Proviral DNA was extracted from productively infected PBMCs. The full-length HIV-1 genome was amplified by using the LA Tag long template PCR system. Primers were positioned in conserved regions within the HIV-1 long terminal repeats. Purified PCR products were T-A ligated into a pWSK29 T vector(CNHN 24 clone). Three recombinant clones containing virtually full length HIV-1 genome were identified by PCR. The full-length genome was sequenced by using the primer-walking approach. Nucleotide sequence similarities were calculated by the local-homology algorithm. Phylogenetic trees of gag, pol and env reading frames were constructed using the Phylip software. RESULTS: HIV-1 C3V4 sequences indicate that the epidemic in this area was B-Thai subtype. V3 loop multiple amino acid sequence alignments showed amino acid alterations at nine positions. The 9,010 bp genomic sequence derived from isolate CNHN 24 contained all known structural and regulatory genes of an HIV-1 genome. No major deletions, insertions, or rearrangements were found. The highest homologies of the gag, pol, vpr, and vif reading frames to the corresponding clade B-Thai RL 42 sequences were 95.42%-97.08%. Phylogenetic trees showed the closest relationship of CNHN 24 and RL 42. CONCLUSION: The cloning and characterization of a virtually full-length HIV-1 B-Thai subtype in central China was completed in our laboratory. The data should be helpful to future studies on the genetic diversity of HIV-1. PMID- 15650790 TI - [Genomic sequence of hepatitis A virus L-A-1 vaccine strain]. AB - OBJECTIVE: To study the genome sequence of hepatitis A virus L-A-1 strain which has been applied for live attenuated vaccine production in China, to compare with other HAV strains, to understand some characteristics of L-A-1 strain, and to find the mechanism of attenuation and cell adaptation. METHODS: Genome fragments were prepared by antigen-capture PCR from infected cell (2BS), PCR products were cloned into T vector, sequenced and analyzed by using bioinformatics program. RESULTS: Analysis of the genomic sequences(nt 25-7,418) showed that the open reading frame contains 6,675 nucleotides in length encoding 2,225 amino acids. Sequence homology comparison showed 98.00% and 94.00% homology at nucleotide level, and 98.51% and 98.65% homology at amino acid level with international strains MBB and HM 175, respectively. Through comparison with other attenuated, cell adapted and cytopathic effect (CPE) strains, L-A-1 strain had mutation at nt 152, 591, 646, 687 and insertion at nt 180-181 in 5?NTR and had mutation at nt 3,889 (aa 1 052-Val) in 2B region, these mutations and insertion are molecular basis for cell adaptation; mutation at nt 4,185 (aa 1 152-Lys) in 2C region should be attenuated marker; deletion in 3A region (nt 5,020-5,025) that caused two amino acids deletion is virus fast growth basis. CONCLUSION: Through analyzing L-A-1 strain genomic sequence, certain sites related to cell adaptation and attenuation were found. PMID- 15650791 TI - [Differential clinical diagnostic parameters of acute hepatitis B and flare of chronic HBV infection]. AB - OBJECTIVE: To evaluate useful clinical diagnostic parameters for differentiating acute hepatitis B and flare of chronic HBV infection. METHODS: Using PCR method to detect viral level in the patient's serum, HBV marker was detected by ELISA kit. Liver function was also detected. RESULTS: The patient can be diagnosed as acute hepatitis B if a patient has one of the following parameters: (1)HBV-DNA negative on admission. (2) When the patient's ALT was lower than 400 IU/L, HBV DNA was negative or HBsAg became negative or HBeAg/HBeAb seroconverted. CONCLUSION: The viral DNA level, HBV marker and ALT can help differentiate acute hepatitis B and flare of chronic HBV infection. PMID- 15650792 TI - [Clinical features of liver failure in children]. AB - OBJECTIVE: To analyze the etiology, clinical and laboratory characteristics of hepatic failure in 105 children. METHODS: The clinical data of 105 children with hepatic failure treated in our hospital from January 1986 to June 2003 were retrospectively analyzed by EXCELL 2000 and t test. RESULTS: (1)Of the 105 children with hepatic failure, 9 were cases with fulminant hepatic failure, 38 with subacute hepatic failure and 58 with chronic hepatic failure. (2)Morbidity was the highest in 7-12 years old children (43/105, 41.0%) followed by infants (30/105, 28.6%). (3)CMV infection could be confirmed in 9 infants (30.0%), etiological diagnosis was not possible in 13 infants (43.3%). Etiological diagnosis could be confirmed in children over 1 year of age, which included hepatitis B (n=22, 29.3%), Wilson's disease (n=15, 20.0%), hepatitis A (n=10,13.3%). Etiology in 21 cases (28.0%) could not be confirmed. (4)Seventy-one cases (67.6%) had ascites, 34 of them (47.9%) had spontaneous peritonitis. Thirty five cases were complicated with other infections. The commonest complication was pulmonary infection and sepsis was the next. Fifty-one cases (48.6%) had hydroelectrolyte imbalance. Forty-eight cases (46.2%) had hepatic encephalopathy, which may be subclinical in children under three years of age. (5)The incidence of hypoglycemia was 77.2%(71/92). CONCLUSION: The etiology of liver failure was related to age. CMV infection was the commonest in infants. HBV, HAV infection was the commonest in children over 1 year of age and Wilson?s disease was the next. It is necessary to prevent and manage the associated complications as early as possible such as spontaneous peritonitis, hepatic encephalopathy, hydroelectrolyte imbalance and hypoglycemia etc. PMID- 15650793 TI - [Experimental and clinical research of "Keyouling" on treatment of condyloma acuminata]. AB - OBJECTIVE: To investigate the effects of traditional Chinese medicine "Keyouling" in treatment of condyloma acuminata (CA). METHODS: The effects of "Keyouling" at different concentrations on growth and proliferation of rabbits' prepuce epithelial cells and CA warts tissue were assessed by means of cellular primary culture and subculture, and the effects of "Keyouling" at different concentrations on the expression level of HPV6.11 DNA and expression level of HPV6.11 DNA of different area of CA patient's penis were quantitatively detected by means of FQ-PCR. RESULTS: "Keyouling" had no effect on growth and proliferation of rabbits' prepuce cuticle cells, but it had significant inhibition on CA warts tissue cells and expression of HPV6.11 DNA. Concentrations of "Keyouling" assumed negative correlated with cell proliferation rate of CA warts tissue and expression level of HPV6.11 DNA, and also had significant inhibition on expression of HPV6.11 DNA of negative cutaneous lesion area and cutaneous lesion area of CA patients. CONCLUSION: "Keyouling" had significant anti HPV6.11 effect, could significantly restrain growth and proliferation of CA warts tissue cells, while it had no damaging effect on normal organic cuticle cells. It was suggested that "Keyouling" was safe in clinical use and had good therapeutic effect on CA and subclinical infection. PMID- 15650794 TI - [Gene chip analysis of mutation of HBV-DNA open reading frame]. AB - OBJECTIVE: To detect the mutation of HBV-DNA open reading frame (ORF), provide evidences for clinical diagnosis and treatment. METHODS: By using gene chip technique, HBV-DNA was amplified by PCR, incorporated with fluorescence, hybridized with oligonucleotide to detect the gene sequence of DNA by computer analysis and observe the natural mutation of HBV-DNA ORF. RESULTS: The mutations of HBV-DNA ORF existed widespread. The rates of mutation on Pre-C 1896, PreC 1814, BCP 1762, BCP 1764, P 528, P 552MI, P 552MV were 23.5%, 3.9%, 55.9%, 53.9%,39.2%, 38.2%, 10.8%, respectively. CONCLUSION: The gene chip technique possesses extremely high sensitivity and reliability,it is one of the effective methods to detect gene mutation. The mutation of HBV-DNA has important influence on the stability and progress of the disease, and on the judgement of prognosis. PMID- 15650795 TI - Marlex mesh Prefix plug hernioplasty retrospective analysis of 865 operations. AB - For 5 years (January 1998 to November 2002) our department has applied the Marlex Mesh Perfix Plug hernioplasty. This article demonstrates the experience gained in operative and postoperative aspects, costs, and outcome along with the results of a follow-up analysis. Altogether, 801 patients (749 males, 52 females) were operated on Sixty-four males had bilateral groin hernias, so the total number of hernioplasties amounted to 865. A total of 19 hernias were recurrent, 297 were direct, 545 were indirect or scrotal (or both) 21 were femoral, and 2 were Spigelian. Fifty-three operations were performed on an emergency basis. Preoperative routine use of antibiotics was minimized throughout the years. Operating time fluctuated from 20 to 25 min (30-40 minutes for recurrent hernias), and the postoperative hospital stay was 28 hours (6-72) hours. The complication rate was 5% and the recurrence rate less than 1%. Early patient mobilization and return to everyday activities (1-2 weeks) was encouraged. The follow-up of 95% of the patient population lasted 12 to 60 months and was performed at 1 week, 1 month, 1 year, and yearly thereafter. The technique demonstrates less overall postoperative pain, discomfort, and complications combined with a remarkably low recurrence rate. The rapid rehabilitation with great patient comfort and decreased operating room time, resulting in lower financial costs, have led us nowadays to repair all types of inguinal hernias, femoral and recurrent ones, using this technique. PMID- 15650796 TI - Surgical treatment of Hurthle cell tumors of the thyroid. AB - Hurthle cell tumors are relatively rare thyroid tumors and their management and prognosis is controversial. We retrospectively review 135 Hurthle cell adenomas and 28 Hurthle cell carcinomas of the thyroid surgically treated at our institute. No significant difference was found between the adenoma and carcinoma groups in patient age, gender, and tumor size. The sensitivity, specificity, accuracy, positive predictive value, and negative predictive value of fine-needle aspiration cytology (FNAC) in detecting malignancy were 87.5%, 19.3%, 31.2%, 18.6%, and 88.0%. The sensitivity, specificity, accuracy, positive predictive value, and negative predictive value of frozen section in detecting malignancy of the thyroid gland were 40.0%, 100.0%, 92.9%, 100.0%, and 92.0%. Most of the adenomas were treated with procedures no less invasive than lobectomy/isthmusectomy. Meanwhile, most of the carcinoma patients underwent total thyroidectomy. Operative morbidity occurred in six patients (3.8%), including transient recurrent laryngeal nerve palsy in three patients, damage to the external branch of the superior laryngeal nerve in two patients, and transient hypocalcemia in one patient. No recurrence of Hurthle cell adenomas was noted. Five adenoma patients died of causes unrelated to their Hurthle cell tumors, but no carcinoma patients died during the follow-up period. In conclusion, clinical factors and FNAC are not helpful in the differentiation between adenoma and carcinoma. Unilateral Hurthle cell adenoma can be treated by lobectomy/isthmusectomy, and Hurthle cell carcinoma can be treated by total thyroidectomy with minimal operative morbidity. PMID- 15650797 TI - Heterogeneity of subvesical ducts or the ducts of Luschka: a study using drip infusion cholangiography-computed tomography in patients and cadaver specimens. AB - The objective of this study was to examine the heterogeneity of the subvesical duct or the ducts of Luschka as well as the reliability of drip-infusion cholangiography with computed tomography (DIC-CT) for their identification. DIC CT was used in 277 consecutive patients (135 men, 142 women) and for the dissection and histologic study of 10 cadaver livers. We found 32 subvesical ducts in 28 (10.1%) of 277 patients. Irrespective of whether the terminals were extra- or intraparenchymal, the subvesical ducts often (17/32 ducts, 15/28 patients) drained into a subsegmental duct of S5 or S4. Notably, some displayed an "intermediate" course along the gallbladder fossa with an intraparenchymal origin and terminated at S5. Such ducts tended to be seen in the elderly. Another 15 ducts drained into a thick, centrally located duct, such as the right sectorial or hepatic duct. The anatomic study demonstrated that the subvesical duct likely included the cystic vein-concomitant duct. A major type of subvesical duct, characterized by an intermediate course along its bed, appeared to result from degenerative exposure of a peripheral duct with aging. The subvesical duct draining into the thick duct included both a "daughter branch," such as seen in the lung, and the so-called anomaly. Such ducts can be demonstrated with DIC-CT, but the incidence might be limited. However, the cystic vein/concomitant duct was difficult to identify radiologically. Consequently, although presurgical radiology is useful for avoiding bile leakage after laparoscopic cholecystectomy, surgeons should be aware of its limitations. PMID- 15650798 TI - Chlorhexidine gluconate: an ideal scolicidal agent in the treatment of intraperitoneal hydatidosis? AB - Hydatid disease is a parasitic disease that is treated primarily by surgery. The most important complication of surgical treatment is spillage of the contents of the cyst, leading to secondary dissemination. In this study, the effect of chlorhexidine gluconate (Chx-Glu) was investigated in the treatment of experimental intraperitoneal hydatidosis (IPH). IPH was reproduced in 100 Wistar albino rats by inoculation with 1 ml of a suspension contained approximately 1500 viable protoscolices of Echinococcus granulosus following determination of scolicidal activity of chlorhexidine gluconate in vitro. Five minutes after protoscolex inoculation, 5 ml of the scolicidal solution was instilled into the peritoneal cavity: 0.9% NaCl (control group), 4.0% Chx-Glu, 0.4% Chx-Glu, and 0.04% Chx-Glu. After 6 months of follow-up, the rats were sacrificed, and the number of isolated cysts, peroperative and postoperative deaths, and toxicity were evaluated. Cyst formation did not occur in any of the Chx-Glu groups compared to the control group (p < 0.05), whereas it was detected in all of the control rats. In addition, to 4.0% Chx-Glu was found to be more toxic and to cause a high mortality rate compared to the 0.4% and 0.04% Chx-Glu groups and the control group (p < 0.05). Chx-Glu 0.04% was found to be the most potent, nontoxic agent; it is easily available, inexpensive, and highly potent in a short period of time at the low concentration. Chx-Glu 0.04% can be used safely in the treatment of intraperitoneal hydatidosis and hydatid cyst. PMID- 15650799 TI - New pancreas-preserving total duodenectomy technique. AB - Pancreas-preserving total duodenectomy (PPTD) was first described by Chung et al. in 1994. Since then, several surgeons have used PPTD to treat diseases that involve the duodenum diffusely but not the head of the pancreas, mostly familial adenomatous polyposis (FAP). The PPTD method has been changed in each report and seems to have improved over time. We performed PPTD on three patients with different diseases--one with intestinal hemorrhage due to small intestinal amyloidosis; another with numerous duodenal gastrinomas in a patient with multiple endocrine neoplasia type 1 (MEN-1) and Zollinger-Ellison syndrome (ZES); and the third with numerous duodenal polyposis and FAP--using a new method that is simpler and safer than those previously reported. When resecting the whole duodenum, we performed mucosectomy of the major papillar portion and saved the structure of the major papilla. After an approximately 8 mm long sphincteropapillotomy, the opened major papilla was anastomosed to an incisional opening of the small intestine. The orifice of the main pancreatic duct (MPD) was stented by a catheter, and the MPD was kept intact under direct vision during the operative procedures. The head of the pancreas was fixed with the small intestine by interrupted 4-0 silk sutures. Reconstruction of the alimentary tract was performed after either the Billroth I or the Billroth II method. This is the first report of PPTD in which the entire MPD was preserved to simplify the biliopancreatic-ductal reconstruction. PMID- 15650800 TI - Bacterial translocation: not a clinically relevant phenomenon in colorectal cancer. AB - The aim of this study was to identify the risk factors for bacterial translocation and to determine the clinical significance of bacterial translocation in patients with colorectal cancer. Mesenteric lymph node sampling was performed to identify the presence of bacterial translocation in 75 patients with colorectal cancer undergoing laparotomy. Bacterial translocation was identified in 29 patients (39%), with the most common organism being Escherichia coli (31%). Three factors for bacterial translocation were identified, including a preoperative low peripheral lymphocyte count, metastasis to lymph nodes, and invasion depth (= T3). Stepwise regression analysis, however, selected only = T3 [odds ratio (OR) 4.0, 95% confidence interval (CI) 1.2-13.5]. Altogether, 35% of patients with bacterial translocation developed septic complications, compared with 20% in patients without bacterial translocation. In the multivariate analysis, bacterial translocation was not an independent risk factor for infection, with an OR of 1.8 (95% CI 0.56-5.96). Systemic inflammatory response syndrome developed on the first day in 62% of patients with bacterial translocation, compared with 50% of patients without bacterial translocation. Adjusting for the other factors, bacterial translocation was not a significant risk factor in the occurrence of systemic inflammatory response syndrome after surgery (OR 1.1, 95% CI 0.37-3.29). We concluded that in patients with colorectal cancers bacterial translocation does occur and is increased in patients with deep invasion. However, it appears to be of no clinical significance. PMID- 15650802 TI - A prospective randomized comparison of bilateral subtotal thyroidectomy versus unilateral total and contralateral subtotal thyroidectomy for graves' disease. AB - To reduce the chance of recurrent hyperthyroidism, two methods of subtotal thyroidectomy were performed and compared. From January 1998 to December 2002, 340 patients were operated on with subtotal thyroidectomy. They were prospectively randomized into two groups. Group A included 166 patients and group B 174 patients. Group A patients had a 2.5 x 1 x 1 cm thyroid remnant on each side and group B patients had a 2.5 x 1 x 1 cm remnant on one side plus total lobectomy on the other side. Thyroid function tests including T(3), T(4), TSH, and antimicrosomal antibody (AMA) were checked preoperatively and in the follow up period of 3 months, and later up to 26.4 +/- 1.1 months (mean +/- SE). The age, sex, duration of oral medicine, and blood loss of the two groups were not significantly different during surgery and the follow-up period. The operative time was less in group A (113 +/- 3.3 minutes) than that in group B (131 +/- 3.2 minutes) (p < 0.001). In the long-term follow-up period, recurrent hyperthyroidism was noted in 15 patients in group A and 3 patients in group B. The difference was significant (p = 0.003). Hypothyroidism was noted in 35 of the group A patients and in 46 of the group B patients. The differences between the two groups regarding hypothyroidism was not significant (p = 0.181). Multivariate logistic regression analysis revealed preoperative titers of AMA > or = 6400, which was the only factor affecting the incidence of hypothyroidism in the later follow-up period. In consideration of hypothyroidism, recurrent hyperthyroidism, and postoperative complications, subtotal thyroidectomy with total lobectomy plus subtotal lobectomy provides a better outcome than bilateral subtotal lobectomy. PMID- 15650801 TI - Duodenogastric reflux after esophagectomy and gastric pull-up: the effect of the route of reconstruction. AB - Duodenogastric reflux (DGR) is a common sequel of subtotal esophagectomy and gastric pull-up, and it may contribute to mucosal changes of both the gastric conduit and the esophageal remnant. This study investigated the effect of the route of reconstruction on the DGR. 24-hour ambulatory bilirubin monitoring was performed on patients who underwent transhiatal subtotal esophagectomy and a gastric tube interposition either in the posterior mediastinum (PM group, n = 11), or in the retrosternal space (RS group, n = 8): A Control group of 8 healthy volunteers was also studied. The median percentage of reflux time, the median number of reflux episodes, and the median number of reflux episodes longer than 5 minutes, in PM versus RS groups, were 29.1% versus 0.15% (p < 0.001), 185 versus 8 (p = 0.002) and 10 versus 0 (p = 0.001), respectively. The values of the above variables in PM versus control groups were 29.1% versus 3.95% (p = 0.007), 185 versus 21 (p = 0.02), and 10 versus 2 (p = 0.009), respectively, whereas in RS versus control groups they were 0.15% versus 3.95% (p = 0.01), 8 versus 21 (p = 0.04), and 0 versus 2 (p = 0.05), respectively. Posterior mediastinal gastric interposition is associated with high reflux of duodenal contents, whereas retrosternal interposition minimizes the reflux at levels even lower than those of the healthy individuals. The latter type of reconstruction may be a good alternative from that perspective, especially in patients with long life expectancy. PMID- 15650803 TI - Long-term follow-up after adrenalectomy for primary aldosteronism. AB - The objective of this study was to assess the long-term effects of adrenalectomy on the blood pressure and antihypertensive medication in patients with primary aldosteronism (PA). Twenty-four patients (15 female and 9 male) with a mean age of 48.3 +/- 10.8 years underwent surgery for PA in our institution between 1988 and 2001. All subjects were re-examined with a complete clinical work-up after a mean follow-up period of 86 +/- 48 months, including blood pressure readings (<140/90 mmHg defined as normal), endocrine adrenal function, and specific medication. All patients suffered from hypertension (onset 8.5 +/- 5.5 years prior to surgery). In 92% of the patients, hypokalemia was present (onset 2.0 +/- 2.6 years prior to surgery). The histopathologic examinations revealed unilateral adenomas in 23 patients and a bilateral hyperplasia in one patient. At follow-up, 33% (8) of the patients were completely cured (normal blood pressure and no antihypertensive treatment), with seven of these eight patients being under 50 years of age at the time of surgery. One patient revealed a contralateral aldosterone-secreting adrenal adenoma during the subsequent endocrine and imaging examination 44 months after the first operation. Despite normalized plasma aldosterone concentration (PAC), plasma-renin-activity (PRA) and serum potassium levels, a long-lasting insufficiently treated hypertension due to the delayed diagnosis in patients with PA may explain the persistent blood pressure elevation, indicating the necessity of a life-long, regular control of the blood pressure and antihypertensive medication. PMID- 15650804 TI - Sequential changes in plasma intact and whole parathyroid hormone levels during parathyroidectomy for secondary hyperparathyroidism. AB - Most commercial assays for intact parathyroid hormone (iPTH) cross-react with non PTH1-84 fragments (likely to be PTH7-84). We aimed to evaluate a whole PTH assay that measured only PTH1-84 by comparing it with an assay measuring iPTH levels during parathyroidectomy in secondary hyperparathyroidism (HPT). Twenty-eight patients with secondary HPT who underwent total parathyroidectomy with autotransplantation served as subjects. Blood samples for postoperative assay were drawn after anesthesia; immediately prior to excision of the last parathyroid gland; and at 5, 10, and 15 minutes after excision. The PTH7-84 level was calculated by subtracting the whole PTH value from the iPTH value. Plasma whole PTH decreased more rapidly than iPTH after parathyroidectomy (p < 0.0001). PTH levels that decreased by 50% or more from levels prior to excision to 10 minutes after excision were used to predict successful parathyroidectomy; decreases in whole PTH substantiated curative surgery for all patients without introducing false-positive and false-negative results. iPTH levels decreased by at least 50% in only 16 patients at 10 minutes after excision without false positive results. Out of 11 cases in which iPTH decreased less than 50%, two were true-negatives and nine were false-negatives. Decreases in whole PTH levels more accurately reflect surgical outcome than do decreases in iPTH levels during parathyroidectomy in secondary HPT patients. Even though the quick iPTH assay is used infrequently during surgery for secondary HPT, our results suggest that a quick whole PTH assay may be more useful than the iPTH assay currently used in parathyroidectomy procedures for secondary HPT. PMID- 15650808 TI - Male takeover in Colobus vellerosus at Boabeng-Fiema Monkey Sanctuary, central Ghana. AB - We describe a case of male takeover in the ursine black-and-white colobus (Colobus vellerosus). In April 2001, an all-male group attacked and eventually invaded our uni-male study group. Aggression increased following the takeover and the former resident male, severely wounded, became peripheral. The youngest immature received severe aggression from the new males but survived. The immature's mother intervened in most instances of this aggression. Eventually, the former resident male re-established relationships with some of the females and concurrently intervened to protect the immature. Defeated males that stay in their group can contribute to the protection of infants born during their tenure from infanticidal males. The females mated with the new males. Takeovers may be a means by which males acquire groups of females in C. vellerosus. PMID- 15650805 TI - Major bronchial trauma in the pediatric age group. AB - Tracheobronchial injuries are rare among all age groups and are extremely rare among the pediatric age group. Yet, the incidence has seemed to increase. Most of these patients die before reaching the hospital from severe associated injuries. Isolated bronchial injury is even more rare than tracheal injury, and it is the focus of the present study. A retrospective national survey was conducted among all tertiary referral and trauma centers in Israel regarding the period between the years 1983 and 1998. Only six cases (3 males and 3 females) of isolated bronchial rupture were found the throughout the country. Ages of the patients ranged from 2 to 14 years; all were involved in motor vehicle accidents, four of them as pedestrians. Ruptures occurred in the bronchus intermedicus (2 cases), left and right main bronchus (2 cases each). All but one patient went through primary repair. We give a full description of the procedure and discuss the literature regarding incidence, diagnosis, treatment, and outcome. PMID- 15650809 TI - Molecular identification of the mycorrhizal fungi of the epiparasitic plant Monotropastrum humile var. glaberrimum (Ericaceae). AB - Achlorophyllous monotropoid plants (Monotropoideae, Ericaceae) are epiparasites that obtain all of their carbon from their host plants via connections with mycorrhizal fungi. The mycorrhizal fungi of the epiparasitic monotropoid Monotropastrum humile var. glaberrima were identified based on mitochondrial, large ribosomal DNA sequences, and were compared with those of another variety, M. humile var. humile. The fungi that inhabit M. humile var. glaberrimum belong to the Thelephoraceae, whereas that of M. humile var. humile is a member of the Russulaceae. Two explanations are possible for this phenomenon: a misunderstanding of the taxonomic position of M. humile var. glaberrimum, or a change in the fungal partner within the Monotropastrum. PMID- 15650810 TI - Cough as the presenting symptom of large vessel vasculitis. AB - We report the case of a 57-year-old woman presenting with persistent cough, weight loss, and fever. An extensive work-up revealed laboratory signs of inflammation and a mild thickening of the aortic wall on computed tomographic scan of the thorax. These findings raised the suspicion of large vessel vasculitis that was elegantly confirmed by fluorodeoxyglucose positron emission tomography. Persistent cough as the inaugural symptom and involvement of large vessels in Horton's disease are also discussed. PMID- 15650811 TI - Development and mapping of a codominant SCAR marker linked to the andromonoecious gene of melon. AB - Monoecy is an important goal for melon breeding because of the agronomic advantages it provides to parental lines in that they do not require hand emasculation to develop monoecious F1 hybrids, the latter producing fruits of higher quality. Monoecious phenotype is conferred by the dominant allele of the andromonoecious (a) gene, whereas recessive homozygous plants are andromonoecious. A bulked segregant analysis (BSA) approach performed in a set of 38 double-haploid lines has allowed us to identify an AFLP marker linked to the a gene at 3.3 cM. Following cloning and sequencing of the AFLP fragment, specific PCR primers were designed and used in the amplification of a codominant SCAR marker. Using a backcrossed mapping population of 530 plants, the SCAR marker could be mapped near the a locus (5.5 cM). Size difference between the two allelic SCAR fragments is 42 bp and might be due to a deletion/insertion. The SCAR marker is closest to the a gene identified to date, and can be useful in breeding programs, using marker-assisted selection procedures to screen for sexual types in melon. PMID- 15650812 TI - Construction and characterization of a plant transformation-competent BIBAC library of the black Sigatoka-resistant banana Musa acuminata cv. Tuu Gia (AA). AB - A plant transformation-competent binary bacterial artificial chromosome (BIBAC) library was constructed from Musa acuminata cv. Tuu Gia (AA), a black Sigatoka resistant diploid banana. After digestion of high-molecular-weight banana DNA by HindIII, several methods of DNA size selection were tested, followed by ligation, using a vector/insert molar ratio of 4:1. The library consists of 30,700 clones stored in 80 384-well microtiter plates. The mean insert size was estimated to be 100 kb, and the frequency of inserts with internal NotI sites was 61%. The majority of insert sizes fell into the range of 100+/-20 kb, making them suitable for Agrobacterium-mediated transformation. Only 1% and 0.9% of the clones contain chloroplast and mitochondrial DNA, respectively. This is the first BIBAC library for banana, estimated to represent five times its haploid genome (600 Mbp). It was demonstrated by hybridization that the library contains typical members of resistance gene and defense gene families that can be used for transformation of disease susceptible banana cultivars for banana genetic improvement. PMID- 15650813 TI - Phylogenetic analysis of Oryza species, based on simple sequence repeats and their flanking nucleotide sequences from the mitochondrial and chloroplast genomes. AB - Simple sequence repeats (SSR) and their flanking regions in the mitochondrial and chloroplast genomes were sequenced in order to reveal DNA sequence variation. This information was used to gain new insights into phylogenetic relationships among species in the genus Oryza. Seven mitochondrial and five chloroplast SSR loci equal to or longer than ten mononucleotide repeats were chosen from known rice mitochondrial and chloroplast genome sequences. A total of 50 accessions of Oryza that represented six different diploid genomes and three different allopolyploid genomes of Oryza species were analyzed. Many base substitutions and deletions/insertions were identified in the SSR loci as well as their flanking regions. Of mononucleotide SSR, G (or C) repeats were more variable than A (or T) repeats. Results obtained by chloroplast and mitochondrial SSR analyses showed similar phylogenetic relationships among species, although chloroplast SSR were more informative because of their higher sequence diversity. The CC genome is suggested to be the maternal parent for the two BBCC genome species (O. punctata and O. minuta) and the CCDD species O. latifolia, based on the high level of sequence conservation between the diploid CC genome species and these allotetraploid species. This is the first report of phylogenetic analysis among plant species, based on mitochondrial and chloroplast SSR and their flanking sequences. PMID- 15650814 TI - A genetic linkage map of Lens sp. based on microsatellite and AFLP markers and the localization of fusarium vascular wilt resistance. AB - Microsatellites have currently become the markers of choice for molecular mapping and marker-assisted selection for key traits such as disease resistance in many crop species. We report here on the mapping of microsatellites which had been identified from a genomic library of lentil (Lens culinaris Medik.). The majority of microsatellite-bearing clones contained imperfect di-nucleotide repeats. A total of 41 microsatellite and 45 amplified fragment length polymorphism (AFLP) markers were mapped on 86 recombinant inbred lines derived from the cross ILL 5588 x L 692-16-1(s), which had been previously used for the construction of a random amplified polymorphic DNA and AFLP linkage map. Since ILL 5588 was resistant to fusarium vascular wilt caused by the fungus Fusarium oxysporum Shlecht. Emend. Snyder & Hansen f.sp. lentis Vasud. & Srini., the recombinant inbreds were segregating for this character. The resulting map contained 283 markers covering about 751 cM, with an average marker distance of 2.6 cM. The fusarium vascular wilt resistance was localized on linkage group 6, and this resistance gene was flanked by microsatellite marker SSR59-2B and AFLP marker p17m30710 at distances of 8.0 cM and 3.5 cM, respectively. These markers are the most closely linked ones known to date for this agronomically important Fw gene. Using the information obtained in this investigation, the development and mapping of microsatellite markers in the existing map of lentil could be substantially increased, thereby providing the possibility for the future localization of various loci of agronomic interest. PMID- 15650815 TI - Development of a codominant PCR-based marker for allelic selection of the pink trait in onions (Allium cepa), based on the insertion mutation in the promoter of the anthocyanidin synthase gene. AB - Bulb color in onions (Allium cepa) is an important trait and is inherited in a complex manner. However, the mechanism of color inheritance is poorly understood at the molecular level. A previous study showed that pink bulb color in onions is inherited as a single recessive trait. This trait is attributable to a significantly reduced transcription of the anthocyanidin synthase (ANS) gene. In this study, we developed a PCR-based marker for an allelic selection of the ANS gene to avoid the laborious progeny tests traditionally employed. To identify polymorphisms between pink and red alleles of the ANS gene, promoter sequences of both alleles were isolated. There was 97% nucleotide sequence identity between the promoter sequences of the two alleles. A 390-bp insertion was identified 632 bp upstream from the putative transcription start site in the pink allele. A pair of primers was designed on the flanking sequences of the inserted region and utilized as a PCR-based marker for allelic selection of the ANS gene. The reliability of the marker was tested using parents, F1 hybrids, and F3 lines whose genotypes had been identified by progeny tests. The marker was also used to evaluate the distribution of the pink allele in white and yellow breeding lines. The results indicated that a majority of the breeding lines tested were homozygous recessive. PMID- 15650816 TI - Using cDNA and genomic sequences as tools to develop SNP strategies in cassava (Manihot esculenta Crantz). AB - Single nucleotide polymorphisms (SNP) are the most abundant type of DNA polymorphism found in animal and plant genomes. They provide an important new source of molecular markers that are useful in genetic mapping, map-based positional cloning, quantitative trait locus mapping and the assessment of genetic distances between individuals. Very little is known on the frequency of SNPs in cassava. We have exploited the recently-developed collection of cassava expressed sequence tags (ESTs) to detect SNPs in the five cultivars of cassava used to generate the sequences. The frequency of intra-cultivar and inter cultivar SNPs after analysis of 111 contigs was one polymorphism per 905 and one per 1,032 bp, respectively; totaling 1 each 509 bp. We have obtained further information on the frequency of SNPs in six cassava cultivars by analysis of 33 amplicons obtained from 3' EST and BAC end sequences. Overall, about 11 kb of DNA sequence was obtained for each cultivar. A total of 186 SNPs (136 and 50 from ESTs and BAC ends, respectively) were identified. Among these, 146 were intra cultivar polymorphisms, while 80 were inter-cultivar polymorphisms. Thus the total frequency of SNPs was one per 62 bp. This information will help to develop new strategies for EST mapping as well as their association with phenotypic characteristics. PMID- 15650818 TI - [Differential diagnosis of inguinal hernias in combination with bladder diverticulum]. AB - In patients with clinical findings suspicious for an inguinal hernia in combination with obstructive or neurogenic bladder dysfunction, herniation of bladder diverticula should be taken into consideration. This differential diagnosis is demonstrated by two case reports. In the patient with the hernia caused by obstructive micturition, we performed transurethral resection of the prostate (TUR-P) prior to resection of the diverticulum. Spontaneous micturition was possible after surgery. In the patient with the neurogenic bladder, we did the hernia repair and resection of the diverticulum primarily. PMID- 15650820 TI - Symptoms, signs and complications in newly diagnosed type 2 diabetic patients, and their relationship to glycaemia, blood pressure and weight. AB - AIMS/HYPOTHESIS: To document the prevalence of typical diabetic symptoms, signs and complications in the diagnosis of type 2 diabetes mellitus, examine their pre diagnostic duration, and analyse associations with glycaemic level, blood pressure (BP), and weight. METHODS: An epidemiological population-based study of 1137 Danish patients with type 2 diabetes newly diagnosed by general practitioners (GPs). GPs and patients together filled in a questionnaire about typical symptoms, signs and complications preceding the diagnosis. RESULTS: Abnormal thirst, frequent urination, weight loss, genital itching, stomatitis, visual disturbances, fatigue, confusion and (in men) balanitis were associated with glycaemic level irrespective of age, sex, BMI, BP, complications and antihypertensive treatment. Eighty-nine percent of the patients presented with one or more of these hyperglycaemic symptoms and signs, and the pre-diagnostic duration was typically less than 3 months. Only a few symptoms, signs and complications were associated with weight and BP. CONCLUSIONS/INTERPRETATION: In patients newly diagnosed with type 2 diabetes in family practice, typical diabetic symptoms, signs and complications are common. Typical diabetic symptoms and signs are associated with hyperglycaemia. The pre-diagnostic duration of hyperglycaemic symptoms and signs were typically short, thus questioning the feasibility of early detection relying on increased anticipatory care by GPs. In contrast, elevated levels of cardiovascular risk factors and longer pre diagnostic duration of cardiovascular complications suggest these might have a central role in an early diagnosis of type 2 diabetes. PMID- 15650819 TI - Relationship between HbA(1)c and mortality in a Japanese population. AB - AIM/HYPOTHESIS: HbA(1)c concentrations are known to be associated with all-cause excess mortality risk in Caucasians. However, the relationship has not been clarified well in the Japanese. In addition, studies of the relationship between HbA(1)c and mortality from malignant neoplasms are scarce. METHODS: HbA(1)c was measured for 3,710 people of a cohort composed of A-bomb survivors and controls. At baseline they were divided into five groups: a normal HbA(1)c group of 1,143 individuals with HbA(1)c of <5.5%, a slightly high but normal HbA(1)c group of 1,341 individuals with HbA(1)c > or =5.5% to <6.0%, a slightly high HbA(1)c group of 589 individuals with HbA(1)c > or =6.0% to <6.5%, a high HbA(1)c group of 259 individuals with HbA(1)c > or =6.5%, and a group of 378 individuals known to have type 2 diabetes. Using a Cox proportional hazards model, hazard ratios based on comparisons with the normal HbA(1)c group were obtained. RESULTS: During the observation period there were 754 deaths. For all-cause and cardiovascular disease mortality, a significant increase of the hazard ratio was observed for the slightly high HbA(1)c group. A similar increase in malignant neoplasm-related mortality was observed for both the high HbA(1)c group and the diabetes group. CONCLUSIONS/INTERPRETATION: Our results suggest that individuals in the Japanese population with HbA(1)c levels of 6% or more might have increased mortality risk. The results indicate that HbA(1)c measurements should be sought even for people who have not been diagnosed with diabetes. PMID- 15650821 TI - [Surgery for fractures of the lower extremities in cases of chronic spinal cord injury]. AB - According to the literature, the need for surgical treatment of fractures of the lower extremity in patients with chronic spinal cord injury (SCI) is very limited. Conservative management is usually presented as the treatment of choice. We present the results of a retrospective review of 55 fractures in 44 patients from 1996 to 2000 which were managed surgically. Rating of the outcome was based on basic clinical and radiological criteria. The results of the radiological investigation on bone remodeling and bone positioning after healing were rated as good or excellent in 43 and fair in five limbs. In five cases, the treatment failed. For 53 of the 55 fractures, the patients regained their normal level of independence. Comparing our results to those presented in the literature, we clearly had fewer complications as well as better clinical and radiological results. We therefore advocate surgical treatment as an appropriate choice of treatment for lower extremity fractures of patients with chronic SCI. PMID- 15650822 TI - [Functional rehabilitation of spinal cord injured persons using neuroprostheses]. AB - Recent technological advancements in microelectronics have led to the establishment of systems for restoration of basic functions in spinal cord injured (SCI) persons using functional electrical stimulation (FES). FES systems for the restoration of bladder and diaphragm function are well established in clinical practice. While FES systems in the lower extremities for standing/walking have not yet achieved widespread clinical acceptance, devices which enhance or restore the grasp function in tetraplegic patients with missing control of hand and fingers are demonstrably successful. Especially with the use of implantable systems a reliable, easy to handle application is possible. The most recent developments in micromechanical engineering are aimed at providing minimally invasive, subminiature systems for functional support in incomplete SCI persons. The possibility of direct brain control of FES systems will expand the application of neuroprostheses for patients with injury of the high cervical spinal cord. PMID- 15650823 TI - [Paraplegia in cases of septic diseases of the spine]. AB - BACKGROUND: Paraplegia of non-traumatic origin has an increasing incidence. Infection of the spine is a major contributor to the number of these cases. Besides adequate management of spinal cord injury, the treatment of these patients requires specific therapeutic techniques. PATIENTS AND METHODS: Eighteen patients with acute pyogenic infection of the spine and spinal cord injury were admitted to our facility in the years 1997-2000. All except two patients were treated by surgical debridement and internal stabilisation. RESULTS: The infections healed after surgical treatment in all cases. Neurological function improved and no patient showed neurological deterioration. Both patients who were not fit for surgery died during treatment. CONCLUSION: Surgical instrumentation and replacement of the destroyed vertebra, despite a pyogenic environment, resulted in permanent resolution of the infection. There is potential for neurological recovery but full recovery is rare. PMID- 15650824 TI - Protocatechuate 4,5-dioxygenase from Comamonas testosteroni T-2: biochemical and molecular properties of a new subgroup within class III of extradiol dioxygenases. AB - Comamonas testosteroni T-2 degraded at least eight aromatic compounds via protocatechuate (PCA), whose extradiol ring cleavage to 2-hydroxy-4 carboxymuconate semialdehyde (HCMS) was catalysed by PCA 4,5-dioxygenase (PmdAB). This inducible, heteromultimeric enzyme was purified. It contained two subunits, alpha (PmdA) and beta (PmdB), and the molecular masses of the denatured proteins were 18 kDa and 31 kDa, respectively. PCA was converted stoichiometrically to HCMS with an apparent K(m) of 55 muM and at a maximum velocity of 1.5 mukat. Structure-activity-relationship analysis by testing 16 related compounds as substrate for purified PmdAB revealed an absolute requirement for the vicinal diol and for the carboxylate group of PCA. Besides PCA, only 5'-hydroxy-PCA (gallate) induced oxygen uptake. The N-terminal amino acid sequence of each subunit was identical to the corresponding sequences in C. testosteroni BR6020, which facilitated sequencing of the pmdAB genes in strain T-2. Small differences in the amino acid sequence had significant effects on enzyme stability. Several homologues of pmdAB were found in sequence databases. Residues involved in substrate binding are highly conserved among the homologues. Their sequences grouped within the class III extradiol dioxygenases. Based on our biochemical and genetic analyses, we propose a new branch of the heteromultimeric enzymes within that class. PMID- 15650825 TI - High benzene selectivity of mesoporous silicate for BTX gas sensing microfluidic devices. AB - The gas selectivities of highly ordered mesoporous silicates and commercially obtained porous silicates with respect to benzene, toluene and xylene were studied. After studying the porosities, pore uniformities, and surface silanol structures of the silicates and their relationships to gas selectivity in detail, we found that we could achieve high benzene selectivity by controlling the micropore size (less than 1 nm). Concluding that mesoporous silicate has a suitable micropore size and structure for benzene selectivity, we also observed that mesoporous silicate SBA-16 exhibited a high (>6) benzene selectivity from toluene and xylene even in a pseudo-atmospheric environment. A benzene detection limit of about 100 ppb was achieved by introducing SBA-16 into a microfluidic device originally developed for the separate detection of benzene, toluene, and xylene gases. PMID- 15650826 TI - Unusual cause of small bowel perforation: metastasis of a subcutaneous angiosarcoma of the head. PMID- 15650827 TI - Ileostomy-site adenocarcinoma after proctocolectomy for ulcerative colitis. PMID- 15650829 TI - Transjugular intrahepatic porto-systemic shunt and variceal embolisation in the management of bleeding stomal varices. AB - BACKGROUND AND AIMS: Bleeding from stomal varices is uncommon. Local measures to control the bleeding offer short-lived control. Our experience with transjugular intrahepatic porto-systemic shunt (TIPS) and variceal embolisation is presented and appraised. PATIENT AND METHODS: Three patients presented with bleeding from stomal varices (Child-Pugh class B, n=2 and class C, n=1) in association with primary sclerosing cholangitis, autoimmune hepatitis and alcoholic liver disease. Local treatment measures including suture ligation, sclerotherapy and re-siting of the stoma achieved short-lived control. TIPS were inserted in all 3 patients, with embolisation of the stomal varices in 2. RESULTS/FINDINGS: The radiological interventions were uncomplicated and resulted in cessation of the bleeding in all patients. One of the patients has had no further bleeding at 12 months' follow-up post-TIPS insertion. The other two patients re-bled at 5 and 6 months post-TIPS insertion and were successfully managed by insertion of a second TIPS in one patient and by balloon dilatation of the TIPS in another. The former patient has had no re-bleeding at a further 8 months' follow-up, while the latter had re bleeding at 12 months post-TIPS insertion and underwent liver transplantation. INTERPRETATION/CONCLUSION: Transjugular intrahepatic porto-systemic shunt with variceal embolisation offers an effective, minimally invasive management option in patients with bleeding stomal varices, and may be used as the primary mode of intervention in conjunction with medical therapy, and as the definitive therapy in patients unfit for surgery. TIPS and variceal embolisation do not preclude subsequent liver transplantation, and may be used during the acute situation as a bridge to transplantation. PMID- 15650828 TI - The role of the insulin-like growth factor system in colorectal cancer: review of current knowledge. AB - BACKGROUND: The insulin-like growth factor system, which includes insulin-like growth factors (IGF-I and IGF-II), IGF receptors (IGF-IR and IGF-IIR) and IGF binding proteins (IGFBPs), plays an important role in epithelial growth, anti apoptosis and mitogenesis. There is a growing body of evidence showing that IGFs control growth and proliferation of several types of cancer. This review introduces the latest information on the biology of the IGF system and its pathophysiological role in the development of colorectal cancer. DISCUSSION: The growth promoting effects of IGF-I and IGF-II on cancer cells are mediated through the IGF-IR, which is a tyrosine kinase and cancer cells with a strong tendency to metastasise have a higher expression of the IGF-IR. Most of the IGFs in circulation are bound to the IGFBPs, which regulate the bioavailability of the IGFs. All IGFBPs inhibit IGF action by high affinity binding, while some of them also potentiate the effects of IGFs. Colon cancer cells produce specific proteases that degrade the IGFBP so that the IGF will be free to act on the cancer cell in an autocrine manner. Therefore, the IGFBPs play a crucial role in the development of the cancer. CONCLUSION: The current knowledge about the link between IGFs and colon cancer is mainly based on in vitro investigations. Further in vivo study is needed to understand the exact role of the IGF system, especially its binding proteins, so that they can be manipulated for the prevention and treatment of colorectal cancer. PMID- 15650831 TI - Plasma vascular endothelial growth factor and serum soluble angiopoietin receptor sTIE-2 in patients with dural arteriovenous fistulas: a pilot study. AB - Our aim was to correlate concentrations of circulating vascular endothelial growth factor (VEGF) and serum soluble angiopoietin receptor (sTIE-2) before and after endovascular treatment with the grading in human dural arteriovenous fistulas (DAVFs). In ten patients with DAVFs undergoing diagnostic cerebral angiography and endovascular intervention, pre-treatment and post-treatment levels of plasma VEGF and serum TIE-2 were examined in a prospective study design. A total of 32 plasma samples and 19 serum samples was collected from the cubital vein, the arterial sheath before and--if applicable--after intervention. Plasma VEGF and serum Tie-2 levels were measured by standardized ELISA protocols. In eight of ten patients with DAVF increased circulating VEGF levels (elevation of more than mean + 2 SD of published normal values) were found, whereas two patients showed increased sTIE-2 levels. Six of the seven patients treated by endovascular embolization displayed a post-interventional decrease of VEGF values. The serum TIE-2 levels decreased slightly after intervention. Pre treatment vVEGF levels varied significantly between patients with grades I and II/III fistulas according to the Cognards classification system. Our pilot study suggests that assessment of angiogenesis parameters in patients with DAVFs might correlate with the DAVFs' grade. To support the hypothesis that a change in angiogenic indicators may serve as indicators for a response to therapy, a larger number of patients should be followed for a longer time period. PMID- 15650830 TI - T1 Gd-enhanced compared with CISS sequences in retinoblastoma: superiority of T1 sequences in evaluation of tumour extension. AB - BACKGROUND: As adequate therapy for retinoblastoma in young children depends on infiltration of extra-retinal structures, diagnostic modalities play an essential role. METHODS: In this widely extended study, 80 children with retinoblastoma were studied with MRI (standard fat-suppressed Gd-enhanced T1, T2 thin-slice sequences (additionally with small loop surface coil), constructive interference in steady state (CISS) sequence covering the orbita). The images were analysed by two blinded neuroradiologists. Histology was used as the gold standard. RESULTS: MRI assumed infiltration of extra-retinal structures in 13 of 80 patients of which ten were confirmed by histology. Affected extra-retinal structures were: optic nerve (five, of which two were on CISS and three on T1 with higher image resolution using the surface coil), scleral infiltration (five, of which four on CISS and T1) and ciliary body infiltration (one on CISS and T1). Another 61 enucleated patients did not have any extra-retinal infiltration in histology. The CISS sequence with multiplanar reconstruction was mainly helpful in revealing exact three-dimensional tumour extension with excellent clinical acceptance and pre-surgical planning but T1 fat-suppressed Gd-enhanced images were superior in revealing exact tumour extension. CONCLUSION: CISS sequences allow to produce excellent anatomical images and to perform multiplanar reconstruction to better demonstrate tumour extension. However, T1-weighted sequences after contrast application are more sensitive (60 versus 40%) in detecting infiltration of the optic nerve but equal in detecting scleral infiltration. PMID- 15650832 TI - Carotid bifurcation calcium and correlation with percent stenosis of the internal carotid artery on CT angiography. AB - The aim of this paper was to determine the correlation between calcium burden (expressed as a volume) and extent of stenosis of the origin of the internal carotid artery (ICA) by CT angiography (CTA). Previous studies have shown that calcification in the coronary arteries correlates with significant vessel stenosis, and severe calcification (measured by CT) in the carotid siphon correlates with significant (greater than 50% stenosis) as determined angiographically. Sixty-one patients (age range 50-85 years) underwent CT of the neck with intravenous administration of iodinated contrast for a variety of conditions. Images were obtained with a helical multidetector array CT scanner and reviewed on a three-dimensional workstation. A single observer manipulated window and level to segment calcified plaque from vascular enhancement in order to quantify vascular calcium volume (cc) in the region of the bifurcation of the common carotid artery/ICA origin, and to measure the extent of ICA stenosis near the origin. A total of 117 common carotid artery bifurcations were reviewed. A "significant" stenosis was defined arbitrarily as >40% (to detect lesions before they become hemodynamically significant) of luminal diameter on CTA using NASCET like criteria. All "significant" stenoses (21 out of 117 carotid bifurcations) had measurable calcium. We found a relatively strong correlation between percent stenosis and the calcium volume (Pearson's r = 0.65, P<0.0001). We also found that there was an even stronger correlation between the square root of the calcium volume and the percent stenosis as measured by CTA (r= 0.77, P<0.0001). Calcium volumes of 0.01, 0.03, 0.06, 0.09 and 0.12 cc were used as thresholds to evaluate for a "significant" stenosis. A receiver operating characteristic (ROC) curve demonstrated that thresholds of 0.06 cc (sensitivity 88%, specificity 87%) and 0.03 cc (sensitivity 94%, specificity 76%) generated the best combinations of sensitivity and specificity. Hence, this preliminary study demonstrates a relatively strong relationship between volume of calcium at the carotid bifurcation in the neck (measured by CT) and percent stenosis of the ICA below the skull base (as measured by CTA). Use of calcium volume measurements as a threshold may be both sensitive and specific for the detection of significant ICA stenosis. The significance of the correlation between calcium volume and ICA stenosis is that potentially a "score" can be obtained that will identify those at risk for high grade carotid stenosis. PMID- 15650833 TI - Effects of home-based daily exercise therapy on joint mobility, daily activity, pain, and depression in patients with ankylosing spondylitis. AB - We investigated the effects of home-based daily exercise on joint mobility, functional capacity, pain, and depression in patients with ankylosing spondylitis (AS). The patients were randomly assigned to a wait-list control group or to an exercise-therapy group. The exercise-therapy group performed a 20-min exercise program once per day for 8 consecutive weeks. After 8 weeks, compared with the control group, the exercise group showed improvements in joint mobility (cervical flexion, extension, shoulder flexion, abduction, hip abduction, and knee flexion), finger-floor distance, and functional capacity. Pain and depression scores were significantly lower after the exercise program in the exercise group than in the control group. These findings indicate that exercise therapy increases joint mobility and functional capacity, and decreases pain and depression in patients with AS. Home-based exercise, which is easily accessible to patients, might be an effective intervention for AS. PMID- 15650834 TI - Prevalence of fat redistribution and metabolic disorders in human immunodeficiency virus-infected children. AB - In this study we assessed the prevalence of fat redistribution and metabolic disorders in a population of HIV-infected children on antiretroviral treatment. To make associations with epidemiological parameters, clinical-immune status, viral load and highly active antiretroviral therapy (HAART), we performed a cross sectional study in HIV-infected children. Epidemiological parameters (age, sex, family history), clinical and immune status, viral load, and duration of antiretroviral treatment (ART) and HAART, including protease inhibitors, were recorded. Presence of clinical signs of fat redistribution and lipid, glucose and lactic acid levels were evaluated. A total of 56 HIV-infected children, including 30 boys (54%), aged between 21 months and 18 years (mean 9.5 years) were studied. In all, 49 patients (87.5%) were receiving ART (mean duration 4 years) and 43 (77%) were receiving HAART (mean duration 3.6 years). Fat redistribution or lipodystrophy was present in 14 patients (25%); seven had lipohypertrophy (12.5%), two lipoatrophy (3.5%) and five a mixed pattern (8.9%). Fat redistribution was higher in children older than 11 years (50%). Of the lipodystrophic patients, 71.4% presented hypertriglyceridaemia (> 130 mg/dl) and 57% hypercholesterolaemia (> 180 mg/dl). We found significant associations between lipodystrophy and age, ART and HAART duration and hypertriglyceridaemia ( P < 0.001, 0.002, 0.016 and < 0.001, respectively), but no significant association with sex, family history, clinical or immune status and viral load. CONCLUSION: The prevalence of lipodystrophy was 25% (95% confidence interval 14.8 34.6) with lipohypertrophy being the commonest pattern. Clinical fat redistribution was significantly associated with older age, duration of antiretroviral treatment and highly active antiretroviral therapy and hypertriglyceridaemia. PMID- 15650836 TI - A (bittersweet?) taste of "American pie": foreign medical school graduates in the US postgraduate medical work force. PMID- 15650835 TI - Uncommon complications of sacrospinous fixation for treatment of vaginal vault prolapse. AB - OBJECTIVE: The present study was undertaken to evaluate uncommon complications following transvaginal sacrospinous colpopexy for treatment of vaginal vault prolapse. CASE REPORTS: A series of three patients who developed uncommon complications following sacrospinous fixation are reported. A 64-year-old patient undergoing bilateral sacrospinous colpopexy for the treatment of an ICS stage III vaginal vault prolapse developed a perineal necrotizing infection. Another patient, a 69-year-old woman with total vaginal vault prolapse and anterior vaginal wall defect (ICS stage II), underwent a right transvaginal sacrospinous colpopexy and anterior repair, presenting postoperatively with a perineal hernia. The third case consisted of a 71-year-old woman who underwent a right sacrospinous colpopexy with paravaginal repair, rectocele repair, and perineorrhaphy for treatment of an ICS stage III post-hysterectomy vaginal vault prolapse, stage II cystocele secondary to a bilateral paravaginal defect, and a stage II rectocele. Six months later the patient developed a left lateral enterocele, which was successfully repaired with a left sacrospinous ligament fixation. DISCUSSION: Etiological factors and treatment considerations for these uncommon complications of sacrospinous colpopexy are discussed in detail, and prophylactic measures, when applicable, are emphasized. PMID- 15650837 TI - Labor analgesia for the parturient with respiratory disease: what does an obstetrician need to know? AB - BACKGROUND: Significant alterations occur in the pulmonary system during pregnancy, which primarily serve to meet the increased oxygen consumption by the growing fetus. When pregnancy is complicated by a respiratory tract disorder such as asthma, cystic fibrosis or tobacco-related respiratory complications the peripartum management (both obstetric and anesthetic) may become challenging. TECHNIQUE: Regional anesthesia remains the technique of choice for these parturients because airway manipulation and endotracheal intubation are avoided. CONCLUSION: When providing labor analgesia to parturients with asthma, cystic fibrosis, and tobacco-related respiratory disorders every effort should be made to reduce the likelihood of high (thoracic) levels of analgesia and/or anesthesia and subsequent airway compromise. PMID- 15650838 TI - Molecular analysis of the conserved C4 region of the S11-RNase of Solanum chacoense. AB - The stylar component to gametophytic self-incompatibility in Solanaceae is an S RNase. Its primary structure has a characteristic pattern of two hypervariable regions, involved in pollen recognition, and five constant regions. Two of the latter (C2 and C3) constitute the active site, while the highly hydrophobic C1 and C5 are believed to be involved in protein stability. We analyzed the role of the C4 region by site-directed mutagenesis. A GGGG mutant, in which the four charged residues in the C4 region were replaced with glycine, did not accumulate the protein to detectable levels in styles, suggestive of a role in protein stability. A R115G mutant, in which a charged amino acid was eliminated to reduce the potential binding affinity, had no effect on the pollen rejection phenotype. This suggests the C4 does not interact with partners such as potential pollen tube receptors facilitating S-RNase uptake. Finally, a K113R mutant replaced a potential ubiquitination target with arginine. However, this RNase acted as the wild type in both incompatible and compatible crosses. The latter crosses rule out the role of the conserved C4 lysine in ubiquitination. PMID- 15650839 TI - Mucosa-associated lymphoid tissue lymphoma of the thymus: a case report with no evidence of MALT1 rearrangement. AB - We report a case of thymic mucosa-associated lymphoid tissue (MALT) lymphoma (TML) that presented as an asymptomatic mediastinal mass in a 40-year-old woman with a past history of Sjogren syndrome. This case had the characteristic clinical and pathological features of TML, as found in most of the 24 previously reported cases, i.e., autoimmune context, especially Sjogren syndrome, IgA secretion, large epithelial cysts, lymphoepithelial lesions involving residual Hassal's corpuscles, epithelial cysts, and a marked plasmacytic differentiation with IgA expression. Reverse-transcription polymerase chain reaction for t(11;18)(q21;q21) was negative, in agreement with recently published data. In this case, investigation for t(14;18)(q32;q21) using fluorescent in situ hybridization was also performed and supplied negative results. Neoplastic cells were negative for MAL, a marker of primary mediastinal large B cell lymphoma (PMBL). Altogether, these findings further support that among MALT lymphomas, TML have peculiar clinical and morphological characteristics and appear not to involve MALT1 rearrangement. They also suggest the absence of a relationship between TML and PMBL. PMID- 15650840 TI - Stability of the HER2 gene after primary chemotherapy in advanced breast cancer. AB - We investigated whether alterations of the Her2 gene could be detected in breast cancer samples following primary chemotherapy in advanced breast cancer. The prospective study involved 23 patients with stage-II, -III or -IV breast cancer. All patients were treated with two to six cycles of fluorouracil-epirubicin and/or cyclophosphamid/epi-docetaxel. The Her2 protein and gene were assessed both on core needle biopsies prior to and on surgical specimens after completing chemotherapy using immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) methods. Estrogen and progesterone receptors (ER/PR) were also determined on both samples using IHC. Her2 status was modified in eight patients using IHC (35%) and in three patients using FISH (13%). Changes in ER/PR expression were detected in seven patients (30%). Our data suggest that alterations of the Her2 gene can occur, although not usually after primary or neoadjuvant chemotherapy. However, changes in ER/PR status seem to be a more common event; thus, both can lead to different therapeutic options. Intratumoral heterogeneity as well as sampling variations can contribute to modification of the Her2 status after primary chemotherapy. PMID- 15650841 TI - Long-term evaluation of isolation-rearing induced prepulse inhibition deficits in rats: an update. AB - RATIONALE: Rats reared in social isolation from weaning show prepulse inhibition (PPI) deficits which are thought to model the sensorimotor gating deficits seen in schizophrenia and other psychiatric disorders. We have previously reported that ten cohorts of Lister Hooded rats reared in isolation showed robust and reliable PPI deficits. OBJECTIVE: Our methodology differed from those used by others (Weiss and Feldon in Psychopharmacology 156(2-3):305-326, 2001), most notably in the weaning of pups at postnatal day (PND) 28 compared with PND20-22. Since our initial report, we have studied 18 more cohorts weaned at PND28 and one cohort weaned at PND21. METHOD: At weaning, male Lister Hooded pups were singly (isolates) or group (n=5) housed (grouped). Eight weeks later, startle and PPI responses of isolates and grouped rats were investigated using conditions of fixed inter-stimulus interval (ISI) (pulse=110 dB/50 ms; prepulse (PP)=75-80 dB/30 ms; ISI=100 ms). RESULTS: Isolates from 14 of the subsequent 18 cohorts demonstrated PPI deficits, giving an overall success rate of 86% for all 28 cohorts. %PPI ranged from 12 to 26% in the isolates and from 26 to 47% in the grouped for the successful cohorts, compared to 16-30% (isolates) and 19-35% (grouped) for those that failed. Only five out of the 19 subsequent cohorts demonstrated startle hyperreactivity, which was unrelated to PPI response. The isolates from the cohort weaned at PND21 did not show a significant deficit in PPI, suggesting, in our hands at least, a requirement for weaning at PND28. CONCLUSION: The data presented here reinforce our original findings that isolation-rearing of Lister Hooded rats provides a viable, non-pharmacological model of impaired PPI. PMID- 15650843 TI - The role of 5-HT in the impairment of thermoregulation observed in rats administered MDMA ('ecstasy') when housed at high ambient temperature. AB - RATIONALE: Administration to rats of a neurotoxic dose of 3,4 methylenedioxymethamphetamine (MDMA, ecstasy) produces an impairment in thermoregulation which is reflected in a prolonged hyperthermic response to a subsequent dose of MDMA given to rats housed at high ambient temperature. OBJECTIVE: We wished to examine whether the impaired thermoregulation was associated with decreased cerebral 5-HT content produced by the prior neurotoxic dose of MDMA. METHODS: Rats were injected with drugs decreasing 5-HT function [the tryptophan hydroxlase inhibitor p-chlorophenylalanine (PCPA), and 5-HT receptor antagonists] and rectal temperature was measured after administering MDMA to rats housed at 30 degrees C. RESULTS: PCPA pretreatment decreased 5-HT and 5-HIAA concentrations in cortex, hippocampus and striatum by >80% and prolonged the hyperthermia induced in rats housed at 30 degrees C by administering MDMA (5 mg/kg i.p.). A similar prolongation of the hyperthermic response to MDMA was seen when rats were pretreated with methysergide (10 mg/kg i.p.) or the 5-HT(1A) antagonist WAY100635 (0.5 mg/kg s.c.). CONCLUSIONS: Decreasing 5-HT function in diverse ways enhanced the hyperthermic response to MDMA given to rats housed at high ambient temperature. This suggests that loss of 5-HT acting on 5-HT(1A) receptors leads to impaired thermoregulation in rats and suggests that the impairment seen in MDMA pretreated rats housed at high ambient temperature is due to a loss in 5-HT function. These data could have implications for recreational users of MDMA, who may have damaged serotoninergic neurons because of prior heavy or frequent use of the drug, when taking further doses of MDMA in hot environments such as dance clubs. PMID- 15650844 TI - Effects of extended-access self-administration and deprivation on breakpoints maintained by cocaine in rats. AB - RATIONALE: Animal models that identify the effects of self-administration histories on subsequent patterns, levels of intake, and other aspects of reinforcement will help clarify the controlling variables of human drug use. OBJECTIVE: Identify the effects of extended-access to cocaine and 1 or 7 days of deprivation on cocaine-maintained breakpoints on a progressive ratio (PR) schedule of reinforcement. METHODS: Male, Sprague-Dawley rats were trained to self-administer intravenous cocaine (expt 1: 1.5 mg/kg per infusion; expt 2: 0.75 mg/kg per infusion), and then given various histories of self-administration and deprivation. Breakpoints, the number of infusions self-administered on a PR schedule, were assessed following the deprivation period. RESULTS: Rates of cocaine intake increased when access to cocaine was extended to 6 h/day. From day 1 to day 14, daily intake increased from 92 (+/-2.5) to 101 (+/-2.8) mg/kg in expt 1, and from 55 (+/-4) to 78 (+/-2.2) mg/kg in expt 2. Total intake across this 2-week period was approximately 1260 and 970 mg/kg in expts 1 and 2. Breakpoints were not different following this escalation period. The introduction of a 7-day deprivation period failed to alter breakpoints. CONCLUSIONS: There is dissociation between changes in rate of cocaine intake (or consumption) and breakpoints maintained on a PR schedule. Extended-access to cocaine produced increases in rate of intake without altering breakpoints. Depending on the experimental question, extended-access conditions may prove useful for studying changes in certain aspects of reinforcement, such as consumption, but not others, such as the strength of a drug as a reinforcer. PMID- 15650845 TI - Effect of ecstasy use on neuropsychological function: a study in Hong Kong. AB - RATIONALE: Previous studies on the effects of ecstasy on neuropsychological performance have often recruited small sample sizes. OBJECTIVES: The present study was conducted to validate previous findings regarding the effects of ecstasy consumption on neuropsychological performance. METHOD: A comprehensive neuropsychological investigation was conducted in 100 abstinent ecstasy users and 100 matched non-user counterparts on standardized measures of working memory, verbal and non-verbal memory, verbal and figural fluency, and selective and switching attention. RESULTS: Abstinent ecstasy users were impaired on verbal and non-verbal memory, complex attention, and verbal fluency, but not on working memory, relative to their non-user counterparts. Of particular interest was the fact that abstinent ecstasy users performed better on figural fluency relative to their non-user counterparts. In addition, only cumulative ecstasy consumption correlated with neuropsychological performances among abstinent ecstasy users. Canonical discriminant analysis yielded verbal and visual memory, switching attention, and verbal fluency as potential core neuropsychological variables for differentiating abstinent ecstasy users from non-users. Levels of depression and general non-verbal intelligence, as measured by the Beck Depression Inventory and the test of non-verbal Intelligence, respectively, were not likely to affect these findings, since these measures were matched between ecstasy users and non users. CONCLUSIONS: These findings suggest that previous ecstasy consumption can affect a wide range of neuropsychological performance, though figural fluency may be subsequently enhanced as a result of the phenomenon of "cortical disinhibition." Furthermore, measures of verbal and visual memory, switching attention, and verbal fluency may be particularly useful for differentiating abstinent ecstasy users from non-users. PMID- 15650846 TI - Ziprasidone and haloperidol in the treatment of acute exacerbation of schizophrenia and schizoaffective disorder: comparison of intramuscular and oral formulations in a 6-week, randomized, blinded-assessment study. AB - RATIONALE: Conventional intramuscular (IM) antipsychotics used in managing acute exacerbation of schizophrenia are associated with side effects such as acute dystonia. OBJECTIVES: To compare the efficacy and tolerability of sequential IM/oral ziprasidone with haloperidol in acute exacerbation of schizophrenia or schizoaffective disorder. METHODS: In a 6-week, multicenter, parallel-group, flexibly dosed study, patients were randomized to ziprasidone (IM up to 3 days, then oral 40-80 mg, b.i.d.) or haloperidol (IM up to 3 days, then oral 5-20 mg/day). Assessments were rater-blinded. RESULTS: At the end of IM treatment, patients receiving ziprasidone (n=427) showed significantly improved Brief Psychiatric Rating Scale Total (BPRS total) scores compared with those receiving haloperidol (n=138) [least-squares (LS) mean change -6.14 for ziprasidone versus 4.13 for haloperidol, P<0.0018]. At endpoint, there were no significant between group differences in BPRS total scores. There was a significantly greater improvement in BPRS negative subscale scores in ziprasidone-treated patients, both at the end of IM treatment (LS mean change -1.15 for ziprasidone and -0.28 for haloperidol, P<0.0001) and at study endpoint (LS mean change -2.94 for ziprasidone and -2.24 for haloperidol, P<0.0001). Haloperidol-treated patients exhibited significantly greater increases in Extrapyramidal Symptom Rating Scale at end of IM treatment and at endpoint (P<0.0001). They also had significantly higher ratings on the Barnes Akathisia Scale (P<0.0001) and the Movement Disorder Burden Score (P<0.005), as well as higher incidences of movement disorder-related adverse events. CONCLUSIONS: Sequential IM and oral ziprasidone offers important efficacy and tolerability advantages over haloperidol in acute schizophrenia. PMID- 15650848 TI - Cloning and characterization of arabinoxylan arabinofuranohydrolase-D3 (AXHd3) from Bifidobacterium adolescentis DSM20083. AB - Arabinoxylan arabinofuranohydrolase-D3 (AXHd3) from Bifidobacterium adolescentis releases only C3-linked arabinose residues from double-substituted xylose residues. A genomic library of B. adolescentis DSM20083 was screened for the presence of the axhD3 gene. Two plasmids were identified containing part of the axhD3 gene. The nucleotide sequences were combined and three open reading frames (ORFs) were found. The first ORF showed high homology with xylanases belonging to family 8 of the glycoside hydrolases and this gene was designated xylA. The second ORF was the axhD3 gene belonging to glycoside hydrolase family 43. The third (partial) ORF coded for a putative carboxylesterase. The axhD3 gene was cloned and expressed in Escherichia coli. Several substrates were employed in the biochemical characterization of recombinant AXHd3. The enzyme showed the highest activity toward wheat arabinoxylan oligosaccharides. In addition, beta-xylanase from Trichoderma sp. was able to degrade soluble wheat arabinoxylan polymer to a higher extent, after pretreatment with recombinant AXHd3. Arabinoxylan oligosaccharides incubated with a combination of recombinant AXHd3 and an alpha-L arabinofuranosidase from Aspergillus niger did not result in a higher maximal release of arabinose than incubation with these enzymes separately. PMID- 15650847 TI - Valpha14 NKT cell-mediated anti-tumor responses and their clinical application. AB - A unique lymphocyte population, Valpha14 NKT cells, has recently been revealed to be a key player in the immune responses against tumors. Activation of Valpha14 NKT cells affects various cell types, particularly dendritic cells (DCs), NK cells, CD4 Th1 cells, and CD8 cytotoxic T cells in the innate and acquired immune systems, eventually resulting in the enhanced activation of NKT cell-mediated cellular cascade in the anti-tumor responses. The specific ligand, alpha galactosylceramide (alpha-GalCer), effectively stimulates mouse and human NKT cells, making NKT cells an ideal target for the development of cancer immunotherapy. Clinical trials using alpha-GalCer have actually started in several centers in the world. In this review, we summarize the Valpha14 NKT cell mediated cellular cascade in the anti-tumor response in mice and discuss potential clinical applications of alpha-GalCer-pulsed DC therapy. PMID- 15650849 TI - Synthesis of ethyl phenylacetate by lyophilized mycelium of Aspergillus oryzae. AB - Lyophilized mycelia of Aspergillus oryzae CBS 102.07, Aspergillus oryzae MIM, Rhizopus oryzae CBS 112.07, Rhizopus oryzae CBS 391.34, Rhizopus oryzae CBS 260.28 and Rhizopus oryzae CBS 328.47 were tested in this study to select the best biocatalysts for ethanol acylation with phenylacetic acid. The mycelium bound carboxylesterase activity of A. oryzae MIM, which exhibited the best performances, was initially investigated at 50 degrees C, either in 0.1 M phosphate buffer or in n-heptane to catalyse the hydrolysis or the synthesis, respectively, of ethyl phenylacetate. The results in terms of product and substrate concentrations versus time were used to estimate the maximum molar conversions at equilibrium, the equilibrium constants, and the times needed to reach half maximum conversions, thus providing sufficient information about this biotransformation. The values of the apparent equilibrium constants, estimated at 20 degrees C0.5 in predicting coughing or movement in response to tracheal suction, but they were unable to predict increases in heart rate or systolic blood pressure. CONCLUSIONS: The aepEX was the best predictor, followed by bispectral index. Although in most intensive care patients subjective sedation scales are sufficient to assess levels of sedation, the aepEX and bispectral index were potential alternatives to subjective scales when they do not work well in the setting of neuromuscular blockade or may not be sufficiently sensitive to evaluate very deep sedation. PMID- 15650863 TI - Goal-directed medical therapy and point-of-care testing improve outcomes after congenital heart surgery. AB - OBJECTIVE: A goal-directed therapy algorithm based on serial lactate values obtained from a point-of-care testing device was utilized in an attempt to reduce the mortality of patients after congenital heart surgery. DESIGN: Prospective study of patients undergoing surgery utilizing a goal-directed therapy algorithm in the postoperative period. The results of this group are compared with a historical cohort. Operative risk was determined using the RACHS-1 scoring system. SETTING: A 12-bed cardiac intensive care unit (ICU) in a pediatric hospital. PATIENTS: Patients undergoing surgery from July 2001 through September 2003 (group B, n=710) were compared to cohorts from June 1995 through June 2001 (group A, n=1,656). Group B patients were smaller and younger (median weight 6.2 vs 8 kg, p<0.001; median age 184 vs 327 days, p=0.004). INTERVENTIONS: Beginning in July 2001, blood lactate measurements were performed serially for 24 h after heart surgery. Based on lactate values and trends, therapy was amended. MEASUREMENTS AND RESULTS: Mortality was lower for group B (1.8 vs 3.7%, p=0.02). A reduction in mortality between group B and group A was noted in neonates (3.4 vs 12%, p=0.02), but not in older patients. Group B patients undergoing higher risk operations (Risk Adjustment for Congenital Heart Surgery-1 [RACHS-1] categories 3-6) had a significant reduction in mortality when compared to group A (3 vs 9%, p=0.006), no difference was noted in patients undergoing lower risk operations (RACHS-1 categories 1 and 2). CONCLUSIONS: The combination of goal directed therapy and point-of-care testing was associated with a marked decrease in mortality for patients undergoing congenital heart surgery. Improvement was greatest in the highest risk patients. PMID- 15650864 TI - Exceptional nonfatal metabolic alkalosis (blood base excess +48 mEq/l). PMID- 15650865 TI - Tissue capnometry: does the answer lie under the tongue? AB - Increases in tissue partial pressure of carbon dioxide (PCO(2)) can reflect an abnormal oxygen supply to the cells, so that monitoring tissue PCO(2) may help identify circulatory abnormalities and guide their correction. Gastric tonometry aims at monitoring regional PCO(2) in the stomach, an easily accessible organ that becomes ischemic quite early when the circulatory status is jeopardized. Despite substantial initial enthusiasm, this technique has never been widely implemented due to various technical problems and artifacts during measurement. Experimental studies have suggested that sublingual PCO(2 )(P(sl)CO(2)) is a reliable marker of tissue perfusion. Clinical studies have demonstrated that high P(sl)CO(2) values and, especially, high gradients between P(sl)CO(2) and arterial PCO(2) (DeltaP(sl-a)CO(2)) are associated with impaired microcirculatory blood flow and a worse prognosis in critically ill patients. Although some questions remain to be answered about sublingual capnometry and its utility, this technique could offer new hope for tissue PCO(2) monitoring in clinical practice. PMID- 15650867 TI - Performance of two measures of general health-related quality of life, the EQ-5D and the RAND-36 among critically ill patients. AB - OBJECTIVE: To compare two health-related quality of life measures, the preference based EQ-5D with five questions and the profile-based RAND-36 with 36 questions, in previous critically ill patients. DESIGN: Prospective observational study. SETTING: A ten-bed medical-surgical intensive care unit (ICU) in a tertiary care university hospital. PATIENTS: Of the 2,709 critically ill patients, treated during the years 1995-2000, the 1,099 patients of the 1,443 still alive who returned both mailed measures were included in the study. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: The EQ-5D and the RAND-36 correlated well (P <.001). Ceiling effect was more obvious with the EQ-5D; the values of the RAND-36 varied usually from 0 to 100 in all the three levels of the corresponding EQ-5D question, and the weakest statistically significant differences were between the EQ levels 2 and 3. In particular, the RAND-36 proved to differentiate better the levels of mobility, self-care, and poor outcome. CONCLUSIONS: The EQ-5D and the RAND-36 correlated well, but when more precisely stated information is needed, especially regarding mobility, self-care, or low quality of life levels of previous critically ill patients, the profile-based RAND-36 may discriminate better. PMID- 15650866 TI - A 12-month clinical survey of incidence and outcome of acute respiratory distress syndrome in Shanghai intensive care units. AB - OBJECTIVE: To investigate incidence, causes, and outcome of acute respiratory distress syndrome (ARDS) in adult patients admitted to intensive care units (ICU) in Shanghai. DESIGN: A prospective 12-month survey during 2001-2002 of the predispositions, clinical management strategies, complications, and 90-day survival rates of patients with ARDS. PATIENTS AND SETTING: Fifteen ICU in 12 university hospitals in Shanghai. All ICU admissions >/=15 years of age in the 12 month period were assessed. Patients fulfilling diagnostic criteria of ARDS, as defined by the American-European Consensus Conference, and having a continuous treatment period >/=24 h, were recruited. MEASUREMENTS AND RESULTS: Of 5320 adult patients admitted to ICUs, there were 108 (2%) with ARDS. At inclusion, ARDS patients had a mean PaO(2)/FiO(2) value of 111.3+/-40.3 mmHg and a mean acute physiology and chronic health evaluation score (APACHE II) of 17.3+/-8.0; 33 patients had a lung injury score >2.5. Forty-one and 67 patients had ARDS associated with diseases of pulmonary and extrapulmonary origin, respectively. The most common predisposing factors for ARDS were pneumonia (34.3%) and nonpulmonary sepsis (30.6%). The overall ICU mortality was 10.3%. In-hospital and 90-day mortalities of ARDS patients were 68.5 and 70.4%, respectively, and accounted for 13.5% of the overall ICU mortality. For ARDS patients, multiple organ dysfunction syndrome was a major risk factor associated with death (59.5%). CONCLUSION: The high morbidity and mortality of ARDS in the ICUs in Shanghai require reassessment of respiratory and intensive care management and implementation of effective therapeutic interventions. PMID- 15650868 TI - Genetic regions that interact with loss- and gain-of-function phenotypes of deltex implicate novel genes in Drosophila Notch signaling. AB - The Notch signaling pathway is an evolutionarily conserved mechanism that regulates many cell fate decisions. The deltex (dx) gene encodes an E3-ubiquitin ligase that binds to the intracellular domain of the Notch protein and regulates Notch signaling in a positive manner. However, it is still not clear how Dx does this. We generated a transgenic line, GMR-dx, which overexpresses dx in the developing Drosophila eye disc. The GMR-dx line showed a rough-eye phenotype, specific transformation of a photoreceptor cell (R3 to R4), and a rotation defect in the ommatidia. This phenotype was suppressed in combination with a dx loss-of function mutant, indicating that it was due to a dx gain-of-function. We previously reported that overexpression of Dx results in the stabilization of Notch in late endosomes. Here, we found that three motifs in Dx, a region that binds to Notch, a proline-rich motif and a RING-H2 finger, were required for this stabilization, although the relative activity of these variants in this assay did not always correspond to the severity of the rough-eye phenotype. In an attempt to identify novel genes of the Notch pathway, we tested a large collection of chromosomal deficiencies for the ability to modify the eye phenotypes of the GMR dx line. Twelve genomic segments that enhanced the rough-eye phenotype of GMR-dx were identified. To evaluate the specificity of these interactions, we then determined whether the deletions also interacted with the wing phenotypes associated with a loss-of-function mutation of dx, dx24. Analyses based on whole genome information allowed us to conclude that we have identified two novel loci that probably include uncharacterized genes involved in Dx-mediated Notch signaling. PMID- 15650869 TI - Four distinct classes of proteins as interaction partners of the PABC domain of Arabidopsis thaliana Poly(A)-binding proteins. AB - Poly(A)-binding proteins (PABPs) play an important role in the regulation of translation and the control of mRNA stability in eukaryotes, and their functions are known to be essential in many organisms. PABPs contain a highly conserved C terminal segment termed the PABC domain. The PABC domain from human PABP interacts with the proteins PAIP1, PAIP2 and RF3 via its PAM2 motifs. These interactions are important for modulating translation. Arabidopsis has eight PABPs, an unexpectedly large number in comparison to other eukaryotes whose genomes have been sequenced. Six of the Arabidopsis PABPs contain the conserved PABC domain. In this work, we have identified PABC-interacting proteins in Arabidopsis. Two proteins, which we named CID1 and CID7, were initially isolated in a two-hybrid screen, and eleven more were predicted to be present in the Arabidopsis proteome and eleven in the rice proteome. Among the 24 PAM2 containing proteins in this set, we observed a diversity of modules of intriguing function, ranging from acidic regions similar to the PAM1 motif found in human PAIP1 and PAIP2, to domains such as the small MutS-related domain, the Lsm domains of Ataxin-2, and RNA recognition motifs (RRMs). We suggest that the large number of PABPs and PAM2-containing proteins may have evolved to provide plants with greater flexibility in modulating the metabolism of specific transcripts. We also found that two PABP genes, PAB2 (ubiquitously expressed) and PAB5 (expressed in reproductive tissues), are essential for viability, suggesting that each has a vital and specific function. PMID- 15650870 TI - O-(2-[18F]fluorethyl)-L-tyrosine PET in the clinical evaluation of primary brain tumours. AB - PURPOSE: The aim of this study was to evaluate the differential uptake of O-(2 [18F]fluorethyl)-L-tyrosine (FET) in suspected primary brain tumours. METHODS: Positron emission tomography (PET) was performed in 44 patients referred for the evaluation of a suspected brain tumour. Acquisition consisted of four 10-min frames starting upon i.v. injection of FET. Tumour uptake was calculated as the ratio of maximal tumour intensity to mean activity within a reference region (FETmax). RESULTS: FET uptake above the cortical level was observed in 35/44 lesions. All histologically confirmed gliomas and many other lesions showed FET uptake to a variable extent. No uptake was observed in nine lesions (one inflammatory lesion, one dysembryoplastic neuroepithelial tumour, one mature teratoma, six lesions without histological confirmation). An analysis of uptake dynamics was done in the patients with increased FET uptake (22 gliomas, three lymphomas, three non-neoplastic lesions, three lesions with unknown histology and four other primaries). Upon classification of tumours into low (i.e. WHO I and II) and high grade (i.e. WHO III and IV), a significant difference in FETmax between the two categories was observed only in the first image frame (0-10 min p.i.), with FETmax=2.0 in low-grade and 3.2 in high-grade tumours (p<0.05); no significant differences were found in frame 4 (30-40 min p.i.), with FETmax=2.4 vs 2.7. Similar results were obtained when the analysis was applied only to astrocytic tumours (2.0 vs 3.1 in the first frame; 2.4 vs 2.6 in the fourth frame). CONCLUSION: These initial results indicate that FET PET is a useful method to identify malignant brain lesions. It appears that high- and low-grade brain tumours exhibit a different uptake kinetics of FET. A kinetic analysis of FET PET may provide additional information in the differentiation of suspected brain lesions. PMID- 15650871 TI - Evaluation of Streptomyces sp. strain g10 for suppression of Fusarium wilt and rhizosphere colonization in pot-grown banana plantlets. AB - Streptomyces sp. strain g10 exhibited strong antagonism towards Fusarium oxysporum f.sp. cubense (Foc) races 1, 2 and 4 in plate assays by producing extracellular antifungal metabolites. Treating the planting hole and roots of 4 week-old tissue-culture-derived 'Novaria' banana plantlets with strain g10 suspension (10(8) cfu/ml), significantly (P < 0.05) reduced wilt severity when the plantlets were inoculated with 10(4) spores/ml Foc race 4. The final disease severity index for leaf symptom (LSI) and rhizome discoloration (RDI) was reduced about 47 and 53%, respectively, in strain g10-treated plantlets compared to untreated plantlets. Reduction in disease incidence was not significant (P < 0.05) when plantlets were inoculated with a higher concentration (10(6) spores/ml) of Foc race 4. Rhizosphere population of strain g10 showed significant (P = 0.05) increase of more than 2-fold at the end of the 3rd week compared to the 2nd week after soil amendment with the antagonist. Although the level dropped, the rhizosphere population at the end of the 6th week was still nearly 2 fold higher than the level detected after 2 weeks. In contrast, the root-free population declined significantly (P = 0.05), nearly 4-fold after 6 weeks when compared to the level detected after 2 weeks. Neither growth-inhibiting nor growth-stimulating effects were observed in plantlets grown in strain g10-amended soil. PMID- 15650872 TI - Isoprenoid biosynthesis in chloroplasts via the methylerythritol phosphate pathway: the (E)-4-hydroxy-3-methylbut-2-enyl diphosphate synthase (GcpE) from Arabidopsis thaliana is a [4Fe-4S] protein. AB - The mevalonate-independent methylerythritol phosphate pathway is widespread in bacteria. It is also present in the chloroplasts of all phototrophic organisms. Whereas the first steps, are rather well known, GcpE and LytB, the enzymes catalyzing the last two steps have been much less investigated. 2-C-Methyl-D erythritol 2,4-cyclodiphosphate is transformed by GcpE into 4-hydroxy-3-methylbut 2-enyl diphosphate, which is converted by LytB into isopentenyl diphosphate or dimethylallyl diphosphate. Only the bacterial GcpE and LytB enzymes have been investigated to some extent, but nothing is known about the corresponding plant enzymes. In this contribution, the prosthetic group of GcpE from the plant Arabidopsis thaliana and the bacterium Escherichia coli has been fully characterized by Mossbauer spectroscopy after reconstitution with (57)FeCl(3), Na(2)S and dithiothreitol. It corresponds to a [4Fe-4S] cluster, suggesting that both plant and bacterial enzymes catalyze the reduction of 2-C-methyl-D erythritol 2,4-cyclodiphosphate into (E)-4-hydroxy-3-methylbut-2-enyl diphosphate via two consecutive one-electron transfers. In contrast to the bacterial enzyme, which utilizes NADPH/flavodoxin/flavodoxin reductase as a reducing shuttle system, the plant enzyme could not use this reduction system. Enzymatic activity was only detected in the presence of the 5-deazaflavin semiquinone radical. PMID- 15650873 TI - Effect of complex formation between Zn2+ ions and the anticancer drug mithramycin upon enzymatic activity of zinc(II)-dependent alcohol dehydrogenase. AB - Mithramycin (MTR), a member of the aureolic group of anticancer antibiotics, is a drug that supposedly acts via the inhibition of transcription. It binds to bivalent cations such as Mg(2+) and Zn(2+). In this paper, we report the association of MTR with Zn(2+), a biologically important bivalent cation whose coordination property leads to its important role as a cofactor in different enzymes and nucleosomal DNA-binding proteins. First, we have characterized the complex formation between MTR and Zn(2+) using spectroscopic methods. In the second part, we have examined the effect of the association of Zn(2+) with MTR on the enzymatic activity of a typical zinc(II)-dependent alcohol dehydrogenase (ADH) from yeast. Our data show that MTR forms a single complex with Zn(2+) in a mole ratio of 2:1 in terms of MTR:Zn(2+). We also observe a negative effect for the preincubation of ADH with MTR upon the enzymatic activity. These results indicate that MTR induced structural changes in the enzyme as a sequel to its complex formation with Zn(2+) present in the enzyme, thereby leading to a loss of enzymatic activity. PMID- 15650874 TI - Variable domains and a VpreB-like molecule are present in a jawless vertebrate. AB - Immunoglobulins (Igs) and T cell antigen receptors (TCRs) that undergo somatic diversification have not been identified in the two extant orders of jawless vertebrates, which occupy essential positions in terms of understanding the evolution of the emergence of adaptive immunity. Using a single motif-dependent PCR-based approach coupled with a vector that allows selection of cDNAs encoding secretion signal sequences, four different genes encoding Ig V-type domains were identified in the sea lamprey (Petromyzon marinus). One of the predicted proteins encoded by these genes shares structural characteristics with mammalian VpreB molecules, including the absence of a recognizable transmembrane region, a relatively high proportion of charged amino acids in its C-terminal tail and distinctive features of its secretion signal peptide. This is the first indication of a molecule related to the B cell receptor (BCR) complex in a species that diverged prior to the jawed vertebrates in which RAG-mediated adaptive immunity is first encountered. PMID- 15650875 TI - Identification of peptides associated with chicken major histocompatibility complex class II molecules of B21 and B19 haplotypes. AB - Chicken major histocompatibility complex (MHC) molecules present peptides to T cells to initiate immune response. Some variants of the chicken MHC, such as B19 and B21 haplotypes, are strongly associated with susceptibility and resistance to Marek's disease, respectively. The objective of the present study was to characterize the repertoire and origin of self-peptides presented by chicken MHC class II (B-L) molecules of B19 and B21 haplotypes. Following immunoaffinity purification of B21 and B19 B-L molecules from transformed B cell lines, their associated peptides were eluted, high performance liquid chromatography fractionated, and sequenced by tandem mass spectrometry. Four peptides were identified associated with B21 B-L molecules. These ranged from 16 to 21 residues in length and had originated from membrane-bound, cytosolic, and mitochondrial proteins. Two of these peptides were present in form of an overlapping set, which is a common characteristic of MHC II-associated peptides. The single B19 associated peptide was 17 residues long and had originated from a cytosolic source. Presentation of endogenous peptides, such as those derived from cytosolic and mitochondrial proteins, by B-L molecules is indicative of cross-sampling between MHC class I and II antigen presentation pathways. These findings facilitate future studies aimed at elucidating mechanisms of chicken MHC association with disease resistance. PMID- 15650876 TI - Molecular cloning of KLRI1 and KLRI2, a novel pair of lectin-like natural killer cell receptors with opposing signalling motifs. AB - We here report the molecular cloning of a novel family of killer-cell lectin-like (KLR) receptors in the rat and the mouse, termed KLRI. In both species, there are two members, KLRI1 and KLRI2. While the extracellular lectin-like domains of KLRI1 and KLRI2 are similar [74% (rat) and 83% (mouse) amino acid identity], they differ intracellularly. KLRI1 has two immunoreceptor tyrosine-based inhibition motifs (ITIMs) in the cytoplasmic domain, suggesting an inhibitory function. KLRI2 has no ITIM, but a positively charged lysine residue in the transmembrane region, suggesting association with activating adapter molecules. Klri1 and Klri2 are localized within the natural killer (NK) cell gene complex on rat chromosome 4 and mouse chromosome 6. By RT-PCR and Northern blot analysis KLRI1 and KLRI2 were selectively expressed by NK cells in both rat and mouse. Epitope-tagged expression constructs of rat KLRI1 and rat KLRI2 induced surface expression of a nondisulphide-linked protein of M(r) 36,000/39,000 and M(r) 34,000, respectively. PMID- 15650877 TI - An immunogenetic basis for the high prevalence of urogenital cancer in a free ranging population of California sea lions (Zalophus californianus). AB - In response to an unprecedented prevalence of cancer recently identified in free ranging populations of California sea lions [(CSL) (Zalophus californianus], we examined the role of the immunologically important major histocompatibility (MHC) genes in this disease epidemic. Associations between MHC genes and cancer have been well established in humans, but have never before been investigated in wildlife. Using a previously developed technique employing sequence-specific primer-based PCR with intercalating dye technology, MHC genotypes were examined from 27 cancer-positive and 22 cancer-negative CSL stranded along the California coastline. Analyses elucidated an underlying immunogenetic component to the high prevalence of urogenital cancer in sea lions. Furthermore, these results demonstrate the functional relevance of CSL class II MHC by revealing a non random nature of cancer susceptibility associated with the presence of specific genes. PMID- 15650878 TI - The mutation spectrum of purified AID is similar to the mutability index in Ramos cells and in ung(-/-)msh2(-/-) mice. AB - Somatic hypermutation and class switch recombination are initiated by the enzyme activation-induced cytidine deaminase (AID). Although other models exist for AID function, one model suggests that AID initiates these processes by deaminating cytidines within DNA, thereby initiating mutagenic repair pathways that involve either UNG or Msh2. Recent work shows that GST-hAID prefers to mutate WRC motifs, a motif frequently mutated in vivo. Because this is a strong argument in favor of the DNA deamination model, we sought to extend this analysis by examining the activity of purified AID with a small polyhistidine tag (His-hAID) on all 16 trinucleotide combinations (i.e., NNC). Here we show that purified His-hAID preferentially mutated cytidines within WRC (i.e., A/T, A/G, C) motifs, but poorly mutated cytidines within GYC (G, C/T, C) motifs. We next compared this mutability preference with those in hypermutating Ramos cells and in msh2(-/ )ung(-/-) mice, since both are reduced or deficient in UNG- and/or Msh2-induced mutations and are thus likely to reflect the sequence specificity of the mutator in vivo. Indeed, the mutation spectrums of purified His-hAID and GST-hAID matched the trinucleotide mutability indexes in Ramos cells and in msh2(-/-)ung(-/-) mice. Thus, the activity of AID on single-stranded DNA produces the same mutation pattern as double-stranded DNA in hypermutating cells. These data lend support to the DNA deamination model and indicate that AID does not require co-factors for its WRC specificity. PMID- 15650879 TI - HLA polymorphisms are associated with Helicobacter pylori infected gastric cancer in a high risk population, China. AB - Helicobacter pylori is one of the most common bacterial infections associated with an increased risk of gastric cancer, but its association with host factors, particularly polymorphisms of the immune response genes, such as human leukocyte antigen (HLA) genes, is still unclear. To investigate the role of HLA polymorphisms in the risk of gastric cancer among subjects with H. pylori infection, a case-control study involving 52 gastric cancer patients and 139 non cancer controls was conducted in Linqu County, China, an area with a high incidence of gastric cancer. Polymorphisms of HLA class I and class II alleles were determined by PCR with sequence-specific primers (PCR-SSP). The information about H. pylori infection was obtained from previous records. Among 48 class I and 19 class II HLA alleles detected in this study, two alleles, CW*03 and DRB1*01, were found to be distributed significantly differently between patients and controls [odds ratio(OR)=1.95, 95% confidence interval (CI)=1.13-3.35, P=0.017 and OR=4.39, 95% CI=1.39-13.84, P=0.012, respectively). The OR of gastric cancer risk in individuals carrying CW*03/CW*03 or CW*03/CW*N was 2.06, 95% CI=1.05-4.02, P=0.035, while the OR was 3.49, 95% CI=1.0-12.4, P=0.04 for DRB1*01/DRB1*01 or DRB1*01/DRB1*N carriers. The analysis of the interaction between H. pylori infection and HLA risk genotypes of CW*03 or DRB1*01 revealed that the effect of CW*03 and DRB1*01 genotypes on gastric cancer risk was manifested stronger in H. pylori-positive individuals (OR=5.30, 95% CI=1.73 16.29, P=0.004 and OR=13.38, 95% CI=2.52-70.98, P=0.002, respectively) than in H. pylori-negative ones (OR=1.25, 95% CI=0.25-6.12, P=0.785 and OR=2.26, 95% CI=0.18 28.88, P=0.531, respectively). The combined effect of the two risk HLA genotypes on gastric cancer risk was also analysed. The result showed that the individuals carrying both the CW*03 and DRB1*01 alleles could only be found in cancer patients (5/52), and not in controls (0/139), further suggesting that CW*03 and DRB1*01 are risk alleles advancing the progression of tumorigenesis. These observations demonstrate that host HLA genotypes may play an important role in the risk of gastric cancer, especially among persons with H. pylori infection. PMID- 15650880 TI - Characterization of EST-derived microsatellites in the wheat genome and development of eSSR markers. AB - EST-derived microsatellites or simple sequence repeats (eSSR) occur in expressed sequence tags (EST). Here we report characteristics of eSSRs in the wheat genome, construction of consensus chromosome bin maps of SSR-containing ESTs ((SSR)ESTs), and development of eSSR markers for the 21 wheat chromosomes. A Perl script known as MISA was used to identify eSSRs in wheat ESTs available in the database http://wheat.pw.usda.gov/cgi-bin/ace/search/wEST ). Among 492,832 ESTs from the database, 36,520 (7.41%) contained 43,598 eSSRs. This is equivalent to 1 eSSR per 5.46 kb EST sequence. About 60% of the eSSRs were trinucleotides, 19.7% were mononucleotide, 16.7% were dinucleotides, and the remaining approximately 3% consisted of tetra-, penta-, and hexanucleotides. Among the identified eSSRs, (CCG/CGG)n is the most frequent (20.5%) followed by (A/T)n at 13.6%, (AAC/GTT)n at 11.7%, and (AG/CT)n at 8.7%. Among ESTs previously mapped to wheat chromosome bins, a total of 1,010 eSSR loci were derived from 341 (SSR)ESTs. Consensus chromosome bin maps showing the chromosome locations of (SSR)ESTs, SSR sequence motif, and cDNA library were constructed. A chi(2) test indicated that the distribution pattern of eSSR loci was generally similar to that of the original mapped ESTs in the wheat genome. Forty-eight (SSR)ESTs were converted into PCR based eSSR markers, and 266 eSSR loci were mapped to specific chromosome arms using wheat cytogenetic stocks. The average polymorphism information content (0.45+/-0.16) of eSSR markers was lower than that reported for genomic SSRs (0.54+/-0.19), but higher than RFLPs (0.30+/-0.27). The eSSR markers were transferable among related Triticeae species, Triticum aestivum, T. durum, T. dicoccoides, Hordeum spontaneum, H. vulgare, and Secale cereale. The results confirm the presence of SSRs in expressed genes of wheat and demonstrate another application of ESTs in genomics research. eSSRs will be useful for gene tagging, gene cloning, and comparative genomics studies of cereal crops. PMID- 15650882 TI - Elucidation of a strange gunshot injury. AB - A 50-year-old obese man sought medical treatment for bleeding injuries in the abdomen and thigh. According to his statement he felt a dull blow on the front of the trunk while he was feeding swans at a lake. The forensic wound findings and the examination of the clothing suggested that the injuries were caused by a single close-range gunshot with the bullet path running nearly vertically downwards and having a total length of 38 cm. The bullet primarily entered in the right mesogastrium, left the body in the right inguinal region, re-entered and ultimately lodged in the right thigh. After confronting the man with the results of the forensic examination he admitted that he himself had caused the gunshot injury accidentally when cleaning his small-bore revolver. PMID- 15650881 TI - Ile118Val genetic polymorphism of CYP3A4 and its effects on lipid-lowering efficacy of simvastatin in Chinese hyperlipidemic patients. AB - OBJECTIVES: To determine the frequencies of CYP3A4 alleles (CYP3A4*4,*5 and *6) in Chinese hyperlipidemic patients and to observe the impact of CYP3A4*4 (Ile118Val) genetic polymorphism on the lipid-lowering effects of simvastatin and on the activity of CYP3A4. METHODS: From hospitalized and non-hospitalized patients, 211 unrelated hyperlipidemic patients were recruited for genotyping. CYP3A4 genotypes were determined by means of polymerase chain reaction and restriction fragment length polymorphism analysis. Of the non-hospitalized hyperlipidemic patients, 8 with CYP3A4*1/*1 and 8 with CYP3A4*1/*4 genotypes were selected to be treated with 20 mg simvastatin daily for 4 weeks. Serum triglycerides (TG), cholesterol (CHO) and low-density lipoprotein (LDL) levels were determined using an automated analyzer (Hitachi 747, Boehringer Mannheim, Mannheim, Germany). CYP3A4 activity was determined by the ratio of 6 hydroxycortisol to free cortisol (6-OHC/FC) in the morning spot urine with a high throughput liquid chromatography-tandem mass spectrometry method. RESULTS: Of 211 subjects, 14 (allele frequency 3.32%) were heterozygous for CYP3A4*4 (Ile118Val). Nevertheless, no subjects with a CYP3A4*5 or CYP3A4*6 allele or homozygous for CYP3A4*4 were identified. The ratio of 6beta-OHC/FC was 9.9 +/- 13.7 and 56.6 +/- 35.7 in subjects with the Ile118Val variant (n = 8) and in CYP3A4 wild-type subjects (n = 8), respectively (P = 0.0039). After oral intake of simvastatin 20 mg daily for 4 weeks, the change of serum lipids in CYP3A4*1/*1 and CYP3A4*1/*4 groups showed a significant difference, with a mean decrease in triglycerides and total cholesterol of 38.1 +/- 7.6% versus 25.1 +/- 8.3% (P = 0.034) and of 35.8 +/- 9.6% versus 22.0 +/-20.4% (P = 0.0015) (means +/- SD), respectively. We found no statistically significant difference in the reductions of LDL between subjects carrying the *1 and *4 genotypes (29.0 +/- 7.4% versus 36.8 +/- 8.8%, P = 0.0721). CONCLUSIONS: The allele frequency of CYP3A4*4 was 3.32% among the hyperlipidemic patients from the Chinese mainland. CYP3A4*4 was an allelic variant related to a functional decrease of CYP3A4 activity, and *4 expression seemed to increase the lipid-lowering effects of simvastatin. PMID- 15650883 TI - Steroid withdrawal in pediatric and adult renal transplant recipients. AB - Corticosteroids are still a cornerstone in the immunosuppressive regimen in pediatric renal transplant recipients despite their numerous side effects, such as inhibition of longitudinal growth, body disfigurement, arterial hypertension, cardiovascular complications, osteopathy, and others. Previous attempts to spare steroids in cyclosporine (CsA)-based protocols have been associated with an increased risk for acute rejection episodes. The recent introduction of more potent immunosuppressive medications, such as mycophenolate mofetil (MMF), have, however, renewed interest in steroid-sparing protocols to avoid or ameliorate steroid-associated side effects. Recent studies in Caucasian adult renal transplant recipients receiving CsA and MMF have shown a beneficial effect of late (>/=6 months post transplant) steroid withdrawal on steroid-associated side effects without the burden of an increased rate of acute rejection episodes. These favorable results compared with previous reports in patients on CsA and azathioprine (AZA) can be ascribed to the higher immunosuppressive potency of MMF compared with AZA. We have shown in a retrospective case control study in 40 pediatric renal transplant recipients that late steroid withdrawal is safe and successful in stable patients under an immunosuppressive maintenance therapy with CsA and MMF. The Mid-European Study Group on Pediatric Renal Transplantation and the Arbeitsgemeinschaft fur Padiatrische Nephrologie are currently performing a prospective randomized trial to validate these observations. PMID- 15650884 TI - The therapeutic potential of novel phosphate binders. AB - Hyperphosphatemia and an increased serum calcium-phosphate (CaxP) product are associated with cardiovascular mortality in adult dialysis patients. Target levels for the treatment of elevations in serum phosphorus (P) and the CaxP product have recently been redefined, but are difficult to achieve in clinical practice. This problem may be especially prevalent in children and adolescents who have a higher recommended P intake than adults and also a higher calcium (Ca) load with the intake of comparatively higher doses of Ca-containing phosphate binders. Current treatment practice with Ca-containing P binders carries the risk of hypercalcemic episodes and ectopic calcifications, including vascular calcifications, which are associated with an increased risk of cardiovascular disease. Novel P binders in the development phase include several iron-containing preparations. Clinical experience with lanthanum carbonate is limited to date and further studies are needed to establish its safety. The only available Ca-free P binder with widespread use and large-scale clinical safety data in adults is sevelamer. Preliminary data suggest that this drug can also be safely given to children. PMID- 15650885 TI - What is needed for effective antitumor immunotherapy? Lessons learned using Listeria monocytogenes as a live vector for HPV-associated tumors. AB - As a vaccine vector, Listeria monocytogenes targets the innate immune system, resulting in a cytokine response that enhances antigen-presenting cell function as well as inducing a Th1 profile. It also enhances cell-mediated immunity by targeting antigen delivery in antigen-presenting cells to both the MHC class I pathway of exogenous presentation that activates CD8 T cells and the MHC class II pathway that processes antigen endogenously and presents it to CD4 T cells. In this review, we describe the development of vaccine constructs that target the human papillomavirus 16 (HPV-16) E7 antigen, and we characterize their effects on tumor regression as well as various immune parameters both innate and adaptive. In particular, we describe the effect on tumor angiogenesis, induction of antitumor suppressor factors like CD4+CD25+ T cells and regulatory cytokines TGF beta and IL-10, homing and infiltration of antigen-specific CD8+ T cells to the tumor, and also effects of the vaccines on antigen-presenting cells, especially focusing on dendritic cell maturation and ability to influence tumor regression. We believe that the identification of several immune parameters that correlate with antitumor efficacy, and of some that have a negative correlation, may have wider application for other cancer immunotherapeutic approaches. PMID- 15650886 TI - Effects of 5 alpha reductase inhibitors on androgen-dependent human prostatic carcinoma cells. AB - PURPOSE: To investigate the effects of MK906, a selective 5 alpha reductase (5alphaR) type 2 (5alphaR2) inhibitor, and of MK386, a specific 5alphaR1 inhibitor, on the cellular proliferation of androgen-dependent human prostatic cancer (PCa) cells in cultures of cells derived from bioptic and surgical tissues. METHODS: In this study we tested the effects of MK906 and MK386 in 30 cultures derived from PCa, 6 from PIN and 10 from benign prostatic hyperplasia specimens. RESULTS: Prostate primary cultures under short-term conditions (with <4 subcultures) represent a mixture of epithelial and stromal cells. Epithelial cells require testosterone (T) for optimal growth, but were not able to grow in the presence of T under long-term conditions even if DHT was able to induce cellular proliferation to a similar extent in both conditions, suggesting that 5alphaR can be lost in long-term cultures. Therefore, our studies were performed under short-term conditions. Both 5alphaR inhibitors decreased cell proliferation significantly and dose-dependently in all the samples tested. MK906 was more efficient than MK386 in 7 out of 10 cultures derived from BPH tissues, in 4 out of 6 cultures derived from PIN and in 18 out of 30 cultures derived from PCa. In 3 out of 10 BPH, in 2 out of 6 PIN and in 5 out of 30 PCa-derived cultures, both inhibitors presented similar efficacy, whereas in 1 out of 10 BPH and 7 out of 30 PCa-derived cultures MK386 was more efficient than MK906. In addition, MK386 was more efficient than MK906 in 4 out of 15 non-metastatic PCa and 2 out of 7 metastatic PCa-derived cultures. CONCLUSIONS: Considering that 5alphaR1 (responsible primarily for androgenic catabolism) is mostly expressed in epithelial cells and that 5alphaR2 (responsible for local DHT synthesis and release) is expressed in the stromal cells (which provides several paracrine growth factors and DHT itself to the epithelial cells), our experiments suggest that the inhibition of both 5alphaR1 and 5alphaR2 by MK386 and MK906, respectively, may have therapeutic potential in order to reduce the growth and progression of human prostatic cancers, through the inhibition of autocrine or paracrine mechanisms involving the stromal cell compartment. In addition, some effects of 5alphaR inhibitors could be mediated by estrogens, which are synthesized by the aromatase enzyme present in the epithelial cells. These aspects could be considered in order to improve the therapeutical management of PCa and for future clinical trials. PMID- 15650887 TI - Effects of pretension on work and power output of the muscle-tendon complex in dynamic elbow flexion. AB - The purpose of the present study was to investigate the effects of pretension on work and power output of the muscle-tendon complex during dynamic elbow flexion under several submaximal and maximal conditions. The subjects were 10 healthy female students. Randomized trials from 0% to 100% maximal voluntary contraction (MVC) pretension (PT) at 60 degrees elbow flexion were conducted. After about 3 s of static PT, subjects maximally flexed the elbow joint to 90 degrees using a quick release method. The weight was individually selected for each subject to provide an optimal load for the development of maximal power. A Hill-type model was utilized to analyze the performance of the elbow muscle-tendon complex (MTC). PT 0, 30, 60 and 90% MVC data were used for comparison, and all data were expressed as the mean and standard deviation. Multiple paired comparisons between the value of PT 0% MVC and that of the other PT levels were performed post-hoc using Dunnett's method. The work of the series elastic component (SEC) increased gradually with the PT level because elastic energy was stored in the PT phase. However, the work of the contractile component (CC) decreased gradually with an increase in PT level. Moreover, the work of the MTC also decreased, closely related to the CC work decrement. The phenomenon of CC work decrement was caused by force depression and was not related to either the force-length or force velocity relationships of the CC. EMG activity (agonist and antagonist) showed no significant differences. Muscle geometry changes or intracellular chemical shifts may have occurred in the PT phase. PMID- 15650889 TI - Spinal cord compression as a result of Rosai-Dorfman disease of the upper cervical spine in a child. AB - BACKGROUND: Rosai-Dorfman disease is characterized by massive painless cervical lymphadenopathy, but can also include nasal obstruction, tonsillar enlargement, or hearing abnormalities. The disease occurs most often in the third and fourth decades. Most authors have suggested that it represents either an autoimmune disease or a reaction to an infectious agent that has yet to be discovered. Less than 50 cases of central nervous system involvement have been reported. CASE ILLUSTRATION: We report a child with cervical spinal cord compression due to Rosai-Dorfman disease of the cervical spine. PROGNOSIS: This disease has variable outcomes from relatively benign to insidious over decades. Our case is unusual in that the patient was a child and had involvement of the spine with resultant cord compression, a combination that has been reported rarely. PMID- 15650888 TI - Effect of load and speed on the energetic cost of human walking. AB - It is well established that the energy cost per unit distance traveled is minimal at an intermediate walking speed in humans, defining an energetically optimal walking speed. However, little is known about the optimal walking speed while carrying a load. In this work, we studied the effect of speed and load on the energy expenditure of walking. The O(2) consumption and CO(2) production were measured in ten subjects while standing or walking at different speeds from 0.5 to 1.7 m s(-1) with loads from 0 to 75% of their body mass (M(b)). The loads were carried in typical trekker's backpacks with hip support. Our results show that the mass-specific gross metabolic power increases curvilinearly with speed and is directly proportional to the load at any speed. For all loading conditions, the gross metabolic energy cost (J kg(-1) m(-1)) presents a U-shaped curve with a minimum at around 1.3 m s(-1). At that optimal speed, a load up to 1/4 M(b) seems appropriate for long-distance walks. In addition, the optimal speed for net cost minimization is around 1.06 m s(-1) and is independent of load. PMID- 15650890 TI - Midvastus approach in total knee arthroplasty: a randomized, double-blinded study on early rehabilitation. AB - The medial parapatellar approach has become the standard technique in total knee arthroplasty (TKA). However, recent studies have reported superior results regarding functional recovery when using the midvastus approach. It was the aim of this study to evaluate the early functional outcome of both surgical techniques. In a prospective, double-blinded, randomized study, 50 patients for TKA were consecutively operated on either by the medial parapatellar or the midvastus approach. Exclusion criteria were defined as previous open knee surgery and leg deformity of more than 10 degrees varus or valgus. TKA was performed in all patients by one surgeon using the same type of implants in both groups. Pain scores (VAS) were documented and follow-up data including quadriceps strength and proprioception were obtained 3 weeks and 6 weeks postoperatively. Both groups were comparable in preoperative demographic data. Postoperatively, patients in the midvastus group demonstrated significantly lower pain in rest (VAS: mean 2.25 vs. 3.03) and under movement (VAS: mean 2.92 vs. 3.13). Further, they showed superior isometric quadriceps strength at 3 weeks (41.4 vs. 27.6 Nm) and 6 weeks (47.6 vs. 35.5 Nm). Moreover, this group showed a superior postoperative proprioception, while the range of motion was similar in both groups. The midvastus approach offers advantages over the standard parapatellar arthrotomy, in the early rehabilitation period. No adverse effects were observed associated with this approach. Therefore, the midvastus approach should be considered as a valuable alternative in TKA. PMID- 15650892 TI - The effects of epidermal keratinocytes and dermal fibroblasts on the formation of cutaneous basement membrane in three-dimensional culture systems. AB - The cutaneous basement membrane (BM) plays an important role in normal and pathological conditions. However, few studies have addressed the formation of the cutaneous BM using three-dimensional culture systems. In this study, to elucidate the effects of human epidermal keratinocytes and dermal fibroblasts on the formation of the cutaneous BM, keratinocytes were cultured on several dermal substrates in the presence or absence of fibroblasts at the air-liquid interface. After 2 weeks of culture, immunohistochemical stainings for the components of the BM and electron microscopic studies of the BM zone (BMZ) were performed. In cultures of keratinocytes alone on dead reticular dermis or collagen gel without fibroblasts, beta4 integrin chain, laminin, type IV and VII collagens were all expressed. However, ultrastructurally, BMZ was not formed. In cultures of keratinocytes on fibroblast-populated collagen matrix, laminin, and type IV and VII collagens were expressed more strongly than in the absence of fibroblasts. In addition, elements of the BMZ such as hemidesmosomes, lamina lucida, lamina densa and anchoring fibrils were formed, although it was still incomplete. In the culture of keratinocytes alone on de-epidermized dermis (DED) (surface up), beta4 integrin chain, laminin, and type IV and VII collagens were strongly expressed. Also, the BMZ appeared similar to that in normal skin. In cocultures of keratinocytes and fibroblasts on DED or cultures of keratinocytes on DED combined with fibroblast-populated collagen matrix, type IV collagen was expressed more strongly than in cultures of keratinocytes alone. Ultrastructurally, similar findings to those of cultures of keratinocytes alone on DED were seen. Interestingly, when keratinocytes and fibroblasts were cocultured on DED, some fibroblasts were seen in the upper dermis as a result of migration into the dermis through partial loss of the lamina densa. These results show that keratinocytes produce most of the components of the BM such as laminin, and type IV and VII collagens. In addition, fibroblasts stimulate the expression of the components of the BM and the formation of a BMZ, suggesting that fibroblasts may produce laminin, and type IV and VII collagens or influence the effects of keratinocytes on the formation of the BM through a keratinocyte-fibroblast interaction. PMID- 15650891 TI - Amphetamine injections into the nucleus accumbens affect neither acquisition/expression of conditioned fear nor baseline startle response. AB - The acoustic startle response is enhanced during states of fear and attenuated during pleasant ones. Our question was whether pharmacological stimulation of the reward system disrupts the learning and retrieval of conditioned fear as measured by fear-potentiated startle. We therefore injected the dopamine agonist amphetamine into the nucleus accumbens (NAC) immediately before either acquisition or expression of conditioned fear and measured the effect of these injections on fear-potentiated startle and baseline startle response. This study clearly showed that amphetamine injections into the NAC had no effect on baseline startle amplitude and acquisition/expression of conditioned fear. In contrast, amphetamine injections into the nucleus accumbens clearly enhanced spontaneous motor activity. These results suggest that dopamine within the NAC is not involved in modulation of fear-potentiated startle and baseline startle. PMID- 15650893 TI - Cytokine polymorphisms in patients with pemphigus. AB - The aim of this study was to assess whether tumor necrosis factor alpha (TNF alpha), transforming growth factor beta1 (TGF-beta1) and interleukin-10 (IL-10) polymorphisms are among the factors influencing the development of pemphigus. Whole blood from 20 patients with pemphigus and 24 control subjects was taken. Genomic DNA was obtained and cytokine genotyping for IL-10 (-1082 G/A; -819 C/T), TGFB1 (codon 10 C/T, codon 25 G/C) and TNFA (-308 G/A) was performed using the ARMS-PCR method. The distribution of IL-10 (-819) alleles was significantly different between the pemphigus and control groups (P=0.009). In particular, allele T was associated with the disease (OR 3.291, 95% CI 1.350-8.020). Similar results were observed when only pemphigus vulgaris (PV) patients were analyzed (P=0.012, OR 3.410, 95% CI 1.346-8.639). An increased frequency of the low producer IL-10 haplotype (-1082/-819 A/T) in patients with pemphigus compared with controls was observed (OR 2.714, 95% CI 1.102-6.685) and this association was also significant when only PV patients were considered (OR 2.667, 95% CI 1.043-6.816). There were no differences between patients and controls in the frequency of any other gene polymorphism analyzed. The increased frequency of the low producer IL-10 haplotype (-1082 /-819 A/T) suggest that the carriage of this haplotype might predispose to pemphigus or the high and intermediate producer haplotypes may be protective factors. The prevalence of the allele IL-10 (-819 T) in pemphigus patients cannot be explained by the current hypothesis, according to which a particular allele of the gene is associated with a different level of cytokine production and therefore affects the predisposition to a particular disease. However, this cytokine polymorphism might be linked to an unknown susceptibility factor. PMID- 15650894 TI - Meteorological influence on NaOH irritation varies with body site. AB - BACKGROUND: Dry air and cold weather have been shown to influence skin irritability. There is conflicting evidence concerning seasonal variability of irritant patch test results and there are no data concerning the influence of season on NaOH-induced irritation. PATIENTS AND METHODS: A Swift Modified Alkaline Resistance Test (SMART) was carried out simultaneously on the volar forearm (FA) and the back of the hand (BOH) of 554 consecutive individuals (277 female, median age 36 years; 277 male, median age 42 years) who had previously had an occupational skin disease. The test comprises two challenges with 0.5 M NaOH each for 10 min. The clinical outcome in each patient was related to standardized data on the local ambient temperature (T) and absolute humidity (AH) on the day of examination obtained from the German Meteorological Service. RESULTS: Of the 554 patients, 212 (38.3%) showed a positive reaction on the FA, and 126 (22.7%) a positive reaction on the BOH. An atopic skin disposition significantly increased the odds for a positive clinical reaction on both the FA (OR 4.8, 95% CI 3.0-7.8) and the BOH (OR 3.1, 95% CI 1.8-5.5). In a logistic regression analysis a significant association between low T and AH, respectively, and clinical reactivity to NaOH on the BOH was found. The odds of a positive reaction were increased by 3.9 (95% CI 2.1-7.6) for an ambient T of < or =6 degrees C and by 2.9 (95% CI 1.6-5.7) for an AH of < or =8 mg l(-1). The effects of AH and T on the reactivity on the BOH were even more pronounced in atopic individuals. In contrast, there was no significant association between the test outcome on the FA and climatic parameters. Furthermore, the test outcome showed no significant association with duration of healing of hand eczema or eczema at other sites before the investigation. Moreover, there were no associations detectable between previous wet work load and NaOH reactivity on both the BOH and FA. CONCLUSIONS: Unlike skin challenge with sodium lauryl sulphate, challenge with NaOH on a body area not directly exposed to environmental factors (FA) seems robust against seasonal influences. Furthermore, a NaOH test in this location (FA) may provide supporting evidence to help identify individuals with atopic skin. On the BOH, seasonal effects were demonstrated. Thus, when interpreting NaOH challenges on the BOH it would seem helpful to take ambient meteorological parameters into consideration. PMID- 15650895 TI - Crossreacting IgG antibodies against fox mite antigens in human scabies. AB - Scabies continues to be an important parasitic disease of mammals. There remain, however, major gaps in the understanding of the human host immune response, and a simple diagnostic test is lacking. In contrast to human mites, red fox mites (Sarcoptes scabiei var. vulpis) can be collected easily and have been used, due to crossreactivity, for enzyme-linked immunosorbent assay (ELISA) studies in dogs and pigs. We wanted to investigate the possibility that crossreactivity might also exist for the human mite, and determined titers against fox mite antigens by ELISA in 41 patients with scabies. Specific IgG was significantly higher in patients with scabies than in healthy controls (P=0.01). The sensitivity was, however, only 48%, although it increased slightly during treatment (P=0.86). A positive correlation was also noted between disease duration and severity of infestation (r=0.5), with specific IgG titers increasing in parallel with severity of symptoms (P=0.01). Patients with symptomatic scabies for more than 4 weeks had furthermore significantly higher IgG titers than patients with a shorter duration of disease (P=0.007). In conclusion, these findings demonstrate IgG antibodies in human scabies that crossreact with fox mite antigens, thus encouraging the search for improved ELISAs with more specific mite antigens to produce a more sensitive detection system for scabies in humans. PMID- 15650896 TI - Orally administered ethanol: transepidermal pathways and effects on the human skin barrier. AB - Ethanol intake is associated with a variety of skin diseases. The aim of the present study was (1) to identify the pathways of release of orally administered ethanol through the skin, and (2) to investigate the effects of a single oral dose of ethanol on the penetration of topically applied substances into the skin. Ethanol evaporation via the skin was measured using the new technique of ion mobility spectrometry (IMS). Transepidermal water loss (TEWL) and skin surface temperature were simultaneously measured before and after ethanol consumption. Measurements were performed on skin sites with different stratum corneum (SC) thickness, and density of follicles and sweat glands. These appendages were selectively sealed to investigate their participation in ethanol evaporation. The penetration of a topically applied UV filter substance was studied before and after ethanol consumption after removing the SC with adhesive tape. Ethanol evaporation was measured within 5 min of consumption, while the skin surface temperature remained nearly constant. The sealing of the appendages did not have a significant effect on ethanol evaporation. On the forehead, a higher TEWL value was measured than on the forearm. On both skin sites, an increase in TEWL was observed after ethanol ingestion. No influence of orally administered ethanol on the penetration of the topically applied UV filter substance was observed. The results indicate that ethanol evaporation occurs via the lipid layers without a significant effect on the penetration of the topically applied substance. PMID- 15650897 TI - A model of the saccade-generating system that accounts for trajectory variations produced by competing visual stimuli. AB - Variable saccade trajectories are produced in visual search paradigms in which multiple potential target stimuli are present. These variable trajectories provide a rich source of information that may lead to a deeper understanding of the basic control mechanisms of the saccadic system. We have used published behavioral observations and neural recordings in the superior colliculus (SC), gathered in monkeys performing visual search paradigms, to guide the construction of a new distributed model of the saccadic system. The new model can account for many of the variations in saccade trajectory produced by the appearance of multiple visual stimuli in a search paradigm. The model uses distributed feedback about current eye motion from the brainstem to the SC to reduce activity there at physiologically realistic rates during saccades. The long-range lateral inhibitory connections between SC cells used in previous models have been eliminated to match recent physiological evidence. The model features interactions between visually activated multiple populations of cells in the SC and distributed and topologically organized inhibitory input to the SC from the SNr to produce some of the types of variable saccadic trajectories, including slightly curved and averaging saccades, observed in visual search tasks. The distributed perisaccadic disinhibition of SC from the substantia nigra (SNr) is assumed to have broad spatial tuning. In order to produce the strongly curved saccades occasionally recorded in visual search, the existence of a parallel input to the saccadic burst generators in addition to that provided by the distributed input from the SC is required. The spatiotemporal form of this additional parallel input is computed based on the assumption that the input from the model SC is realistic. In accordance with other recent models, it is assumed that the parallel input comes from the cerebellum, but our model predicts that the parallel input is delayed during highly curved saccadic trajectories. PMID- 15650898 TI - On partial contraction analysis for coupled nonlinear oscillators. AB - We describe a simple yet general method to analyze networks of coupled identical nonlinear oscillators and study applications to fast synchronization, locomotion, and schooling. Specifically, we use nonlinear contraction theory to derive exact and global (rather than linearized) results on synchronization, antisynchronization, and oscillator death. The method can be applied to coupled networks of various structures and arbitrary size. For oscillators with positive definite diffusion coupling, it can be shown that synchronization always occurs globally for strong enough coupling strengths, and an explicit upper bound on the corresponding threshold can be computed through eigenvalue analysis. The discussion also extends to the case when network structure varies abruptly and asynchronously, as in "flocks" of oscillators or dynamic elements. PMID- 15650899 TI - Spatial and temporal structure of phase synchronization of spontaneous alpha EEG activity. AB - Spatiotemporal characteristics of spontaneous alpha EEG activity patterns are analyzed in terms of large-scale phase synchronization. During periods with strong phase synchronization over the entire scalp, phase patterns take either of two forms; one is a gradual phase shift between frontal and occipital regions and the other is a stepwise pattern with a sudden phase shift in the central region. The former is regarded as a traveling wave of electrocortical activity, of which the direction of propagation is predominantly from anterior to posterior in three out of four subjects, and opposite in the remaining one. The other activity pattern observed may correspond to a standing wave composed of two traveling waves propagating in opposite directions. The duration distributions of these patterns have similar forms within a subject, which suggests that they share the same mechanism for their generation. PMID- 15650900 TI - [Mold indoors]. PMID- 15650901 TI - [Molds in air-sampling and detection. Validation of sampling methods to verify molds in air]. AB - The assessment of mold concentrations in air samples requires generally accepted evaluation criteria. It is not possible to derive guideline values based on risk assessment. Background concentrations are therefore used as the basis for the assessment of measured concentrations. The assessment criteria established by the Federal Environmental Agency in Berlin and by the State Health Agency of Baden Wurttemberg are founded on the experience of some laboratories. The general application of these criteria is only meaningful if validated and generally applied methods of analysis are used. Until now, laboratories have been using many different methods for sampling and detection of culturable molds and total spore counts in air. In this study, different steps for sampling by impaction or filtration are validated. The results presented will contribute to standardization of sampling and detection of molds in air. PMID- 15650902 TI - [Background concentrations of molds in air. Determination of mold concentrations in dwellings without known mold infestations in three parts of Germany]. AB - The mold concentration of indoor air and outdoor air in three parts of Germany was investigated in both winter and summer. In total, air samples from 80 dwellings, reportedly free from mold infestations, were analysed with both the cultivation method and the total spore count method. With the cultivation method, 40 mold species and genera were differentiated, while with the total spore count method, 11 spore types were distinguished. The concentration of the genera Alternaria, Cladosporium, Fusarium and yeast increased in summertime. In contrast, the concentrations of Aspergillus and Penicillium measured in summer were nearly equal to those measured in winter. The majority of the various molds were only found in a small number of samples, too small for a reliable comparison of the winter and summer findings. The 95th percentile of the indoor mold concentrations is suggested as the upper limit of the background concentration. The results are discussed comparing the assessment proposals of various authors, and a new assessment proposal is described. PMID- 15650903 TI - [Molds in house dust. Sampling and detection]. AB - Examination of dust samples is widely used to detect sources of mold growth in indoor environments. The advantage of this investigation is that house dust can be regarded as a passive sampler collecting mold spores over a long time period. No generally accepted method exists, however, for sampling and analysis of house dust for the detection of molds. Therefore, results obtained by different laboratories are not comparable. Measurements with standardized validated methods are a prerequisite to assess concentrations of molds in indoor environments. The method presented is based on sampling with a modified filter holder (in accordance with VDI 4300 page 8). Mini-sieves of different sizes are used for sample preparation. The fraction <63 microm is used for cultivation of molds after suspension. Problems of dust sampling and preparation are discussed. Criteria for the detection of sources of mold growth based on dust samples are presented elsewere. PMID- 15650904 TI - [Background concentrations of molds in house dust. Determination of mold concentrations in dwellings without known mold infestations in three parts of Germany]. AB - Mold concentrations of house dust samples from dwellings reportedly free from mold infestations were analysed to obtain background values. Samples from carpet floors were taken from 80 dwellings in three parts of Germany in winter and summer. Samples were analysed with the cultivation method (using suspension). This resulted in the detection of 35 mold species or genera. Concentrations of the genera Alternaria, Cladosporium, Fusarium and yeast increased in summer. In contrast concentrations of Aspergillus and Penicillium in the summer samples were nearly equal to those in the winter samples. The majority of the various molds were only found in a number of samples too small for a reliable comparison of the winter and summer findings to be made. The results were compared with findings of other authors. The authors propose the concentrations of the 95th percentile of each species (when representing over 10,000 KBE/g of dust) as background values, while a threefold higher value is regarded as an indication of mold presence. An exception should be made for the concentrations of molds which increase sharply in the outdoor air in summer. Because of the difficulty in estimating the outdoor influence, no evaluation proposals can be given for these species for the summer season. PMID- 15650905 TI - [Quality assurance in the diagnosis of indoor fungi]. AB - In co-ordination with the Umweltbundesamt Berlin, the Landesgesundheitsamt Baden Wurttemberg (LGA) initiated external quality assurance in the diagnosis of indoor fungi in autumn 2001. Four of six fungal strains commonly found indoors have to be fully identified (on the genus and species level). There are two distributions per year; the six distributions hitherto carried out resulted in correct identification by 46-89% of laboratories (40-71 participants, total 148). It is clear from the results that repeat participants were more successful. In addition to the pure cultures sent out we offered actual samples (two air samples, one wood material, one sample of house dust, hitherto); 43- 69% of participating laboratories also took part in this facultative investigation of actual samples and 29-62% were successful. Results that differed considerably revealed problems while treating and evaluating actual samples. Therefore, activities in this field should be enhanced. In conclusion, external quality assurance in the diagnosis of indoor fungi is a useful management aid in the maintenance and improvement of laboratory performance. PMID- 15650906 TI - [Comparison of methods for quantification of MVOC in indoor environments]. AB - For several years now, MVOC have been regarded as indicators for microbial growth in indoor environments. Until now, a direct correlation between the occurrence of microfungi and MVOC could not be shown in scientific investigations. One reason may be that different analytical methods were applied, and moreover they were not validated sufficiently. The present investigation aimed to test the comparability of both methods (Tenax adsorption/thermal desorption; charcoal adsorption/elution). It turned out that with both methods comparable results can be achieved if the technical handling of the calibration is standardized to a wide extent. Thus, highest demands have to be made on quality assurance. Therefore, it is necessary to formulate technical regulations for the quantification of MVOC. PMID- 15650907 TI - [Global warming and spread of infectious diseases]. AB - At the end of the twentieth century, tropical infectious diseases increased despite earlier successes of eradication campaigns. As a global warming of 1.4 5.8 degrees C is anticipated to occur by 2100, mainly the vector-borne tropical diseases that are particularly sensitive to climate are expected to spread. Although biological reasons seemingly support this hypothesis, ecological and socioeconomic factors have in the past proven to be stronger driving forces for the spread of infectious disease than climate. PMID- 15650908 TI - [Electrosmog, cellular phones, sunbeds etc. -- adverse health effects from radiation? Health aspects of non-ionizing radiation]. AB - This review supplies a survey of the three physical influences, i. e. UV radiation, high-frequency electromagnetic fields of radio telephone systems and other wireless radio applications as well as low-frequency fields of electric power supply. The exposure to UV radiation must be considered to be by far the highest health risk. The annual rate of about 2000 deaths from skin cancer in Germany, mainly caused by extensive exposure to solar UV radiation, demands protective measures. Teaching reasonable behaviour is the supreme issue. Recommended protective measures in the order of their effectiveness are protection by adaptation of behaviour, by clothes, sun hats and sunglasses as well as by sun creams. Children are the most important target group. With regard to UV tanning appliances it is recommended not to use artificial UV radiation for cosmetic purposes because of the related health risks. For the assessment of health impairments caused by exposure to electromagnetic fields, direct field reactions due to induced electric body currents, reactions on the surface of the body or heating effects should be separated from indirect field reactions (e. g. electric shocks and burns) due to contact currents or interference with electronic body aids and implants. Risk assessment has led to recommendations of threshold values which-in agreement with international research results-exclude all impairments of health caused by direct field reactions scientifically proven to date. Contrary to public concerns, which are mostly related to base transmitters of radio telephone systems, exposure due to handheld radio telephones (cellular phones) should rather be considered from the viewpoint of precautionary health protection, since it is more likely that their use can lead to high exposure of the user. Due to the protective measures provided so far and observance of the threshold values based on scientific results, exposures do not lead to health impairments-not even in children. Because of the introduction of new technologies more portable devices are to be expected that may be operated near the body. This will further increase the exposure of people to high frequency electromagnetic fields. PMID- 15650909 TI - [Cosmetic colorants. Toxicology and regulation]. AB - Some recent publications raised concern over a possible link between hair dye use and the incidence of bladder tumours in a Californian population. The Scientific Committee for Cosmetic Products and Non-Food Products intended for Consumers (SCCNFP) demanded the toxicological testing of all hair dyes used in Europe to exclude any risk. The EU commission initiated corresponding measures. Only safe hair dyes will be included on a positive list while all other hair dyes will be banned. The hair dye lawsone--the dyeing ingredient of henna--was evaluated by the SCCNFP as genotoxic but the BfR came to another conclusion. The regulation of both lawsone and henna remains an open question. Furthermore, some cosmetic colorants were critically discussed. The azo dyes CI 12150, CI 26100, CI 27290 and CI 20170 are allowed for use in cosmetics. On cleavage they form the carcinogenic aromatic amines o-anisidine, 4-aminoazobenzene and 2,4-xylidine, respectively. For three of these dyes the cleavage by human skin bacteria in vitro to the respective arylamine was shown experimentally. Further problems may arise from colorants used for tattoos and permanent makeup. These products up to now are not subject to legislation and there are no regulatory stipulations with respect to health safety and purity for colorants used for these purposes. PMID- 15650910 TI - [Estimation of the dietary intake of pesticide residues based on new consumption data for children]. AB - The risk assessment of pesticide residues is based on the estimation of their dietary intake. Models based on a new national consumption survey were developed to estimate the short- and long-term dietary intake of pesticide residues for children from 2 to under 5 years, allowing a realistic risk assessment to be made. The recommended methods are described. At the national level, the new models shall replace the previous methods for evaluating dietary intake. PMID- 15650911 TI - [Report on Notifications pursuant to Section 21 German Transfusion Act for 2001 and 2002]. AB - The report pursuant to Section 21 German Transfusion Act is based on data referring to 4.53 and 4.66 million allogeneous whole blood donations and 1.35 and 1.89 million apheresis donations respectively for the years 2001 and 2002. Since 2000, the quantity of plasma for fractionation from apheresis has doubled to approx. 1 million litres in 2002. At the same time, the number of private apheresis facilities has also increased. The data on plasma for fractionation (both from whole blood and from apheresis) have shown a sharp increase for both report periods, mainly due to the rise in the quantity of plasma from apheresis. In spite of this, the quantity for plasma for fractionation available to the German market declined by 43% during the report period, due to increasing export figures. The data on the manufacture of blood components are reliable, since the notifications are almost complete. In 2001, 4.32 million transfusion units of erythrocyte concentrates, the most important product derived from whole blood donations, were manufactured, compared with 4.45 million in 2002, out of these, slightly less than 77 percent were manufactured in the blood donations services of the Red Cross. The consumption values reported were clearly below the figures for the manufacture, with 3.2 million transfusion units in 2001 and 3.5 million in 2002. However, some facilities of the health service did not meet the requirement to notify their figures. As far as the data on manufacture, imports and exports of plasma derivatives are concerned, further improvements are required because of the complexity of the matter and the various implications to be taken into account. However, a statement on the supply situation cannot be yet made anyway, due to the missing notifications on the consumption figures. PMID- 15650912 TI - ["Textiles" Working Group in the BfR report of the 11th meeting of the "Health Evaluation of Textile Adjuvants and Colorants" Study Circle of the Federal Institute of Risk Assessment (BfR) 16 December 2003 Berlin]. PMID- 15650913 TI - [Meetings of the Preliminary Synthetic Products Committee of BfR. Reports of 2-10 April 2003, 12-13 November 2003 and 28-29 April 2004}]. PMID- 15650914 TI - [4th Ministerial Conference about the Environment and Health from 23-25 June 2004 in Budapest. The future of our children]. PMID- 15650915 TI - Frequency and diversity of nitrate reductase genes among nitrate-dissimilating Pseudomonas in the rhizosphere of perennial grasses grown in field conditions. AB - A total of 1246 Pseudomonas strains were isolated from the rhizosphere of two perennial grasses (Lolium perenne and Molinia coerulea) with different nitrogen requirements. The plants were grown in their native soil under ambient and elevated atmospheric CO2 content (pCO2) at the Swiss FACE (Free Air CO2 Enrichment) facility. Root-, rhizosphere-, and non-rhizospheric soil-associated strains were characterized in terms of their ability to reduce nitrate during an in vitro assay and with respect to the genes encoding the membrane-bound (named NAR) and periplasmic (NAP) nitrate reductases so far described in the genus Pseudomonas. The diversity of corresponding genes was assessed by PCR-RFLP on narG and napA genes, which encode the catalytic subunit of nitrate reductases. The frequency of nitrate-dissimilating strains decreased with root proximity for both plants and was enhanced under elevated pCO2 in the rhizosphere of L. perenne. NAR (54% of strains) as well as NAP (49%) forms were present in nitrate reducing strains, 15.5% of the 439 strains tested harbouring both genes. The relative proportions of narG and napA detected in Pseudomonas strains were different according to root proximity and for both pCO2 treatments: the NAR form was more abundant close to the root surface and for plants grown under elevated pCO2. Putative denitrifiers harbored mainly the membrane-bound (NAR) form of nitrate reductase. Finally, both narG and napA sequences displayed a high level of diversity. Anyway, this diversity was correlated neither with the root proximity nor with the pCO2 treatment. PMID- 15650930 TI - The American Society of Hematology--46th Annual Meeting and Exposition. PMID- 15650931 TI - The American Society of Hematology--46th Annual Meeting and Exposition. HDAC, Flt and farnesyl transferase inhibitors. PMID- 15650932 TI - The American Society of Hematology--46th Annual Meeting and Exposition. Drug highlights I. PMID- 15650933 TI - The American Society of Hematology--46th Annual Meeting and Exposition. Drug highlights II. PMID- 15650934 TI - American Epilepsy Society--58th Annual Meeting. Novel compounds for the treatment of seizures. PMID- 15650935 TI - Drug therapy in HIV infection--Seventh International Congress. PMID- 15650936 TI - San Antonio Breast Cancer Symposium--27th Annual Meeting. PMID- 15650937 TI - Clinical Trials in CNS--SMi Conference. PMID- 15650938 TI - Advances in Antiviral Therapeutics--SMi's Second Annual Conference. PMID- 15650939 TI - Genesis IV--Fourth London Biotechnology Conference. PMID- 15650940 TI - Oncology business development global summit. PMID- 15650941 TI - Interscience Conference on Antimicrobial Agents and Chemotherapy--44th Annual Meeting. PMID- 15650942 TI - Ethnomedicine in malaria treatment. AB - The most pressing problem in malaria is parasite resistance to classical antimalarial drugs such as chloroquine, particularly as no alternative antimalarial agents are currently available that combine the low cost, safety and efficacy that once characterized chloroquine. Indeed, people in these endemic countries must survive on less than US dollars 15 per month, and consequently a maximum cost of US dollars 1 per person is all that can be afforded for antimalarial treatment. Only chloroquine and sulfadoxine-pyrimethamine are widely available at this price but resistance to these drugs is widespread, hence the development of cheap alternative therapies are essential. For these endemic populations, traditional medicine appear indispensable in the fight against this disease. Moreover, with regard to the scientific community, natural products demonstrate the highest chemical structural diversity. PMID- 15650943 TI - Recent advances in the research and development of CB1 antagonists. AB - The worldwide prevalence of obesity has dramatically increased over the last 30 years, yet due to modest efficacy and side effects, current treatments for obesity are inadequate. Antagonists of the cannabinoid receptor type 1 (CB(1)) have emerged as a highly promising therapeutic strategy for obesity, with the 1,5 diaryl-pyrazole rimonabant (Sanofi-Aventis) being the most characterized and the furthest advanced agent in the clinic. This review focuses on recent publications (2002 to 2004) that describe the effects of CB(1) antagonists both on feeding behavior and in models of obesity, and the identification and characterization of new structure classes as CB(1) antagonists. PMID- 15650944 TI - M-40403 Metaphore Pharmaceuticals. AB - Metaphore Pharmaceuticals Inc. is developing M-40403, a superoxide dismutase mimetic, for the potential treatment of pain, dermatological disease and inflammation. The company is also investigating the compound for its potential in the treatment for the side effects associated with chemotherapy and radiation therapy. PMID- 15650956 TI - [(Adaptation to stress: meaning and sense in medicine]. PMID- 15650957 TI - [Expensive modern therapy options in intensive care medicine in Germany-- are they being used? Results of a questionnaire]. AB - BACKGROUND: The use of new, high-priced therapies in intensive care medicine should be assessed by a questionnaire. METHODS: Questionnaires were sent to 849 intensive care units in Germany. The use of three new strategies were asked: treatment of methicilline resistant staphylococcus aureus (MRSA) patients (using linezolid), of severe bleeding (using rFVIIa), and of severe sepsis (using activated protein C [aPC]). RESULTS: Approximately 39 % of the questionnaires were answered and analyzed. All three new strategies were only rarely or very rarely used in Germany even in universities and hospitals with more than 1000 beds. This appears to be very astonishing because all substances were subject to extensive marketing campaigns, received scientific prices (linezolid) or were strongly recommended by some scientific societies (aPC). One major concern to the use of the new approaches was based on the high pricing. Prices were assessed as excessive or very excessive. In spite of a mass of information about the substances, a lot of the intensivists reviewed the scientific basis as weak and not justifying the use of the costly substances. CONCLUSIONS: Modern, costly pharmaceutical approaches in intensive care medicine are widely not accepted in Germany. Especially a tight financial corset hinders most intensivists to use these strategies that may be life-saving in some patients. A solution to this problem is urgently required that can be reached only in intensive exchange with all who are responsible for this dilemma. PMID- 15650958 TI - [Duodenal ileus due to an intramural duodenal haematoma. Conservative therapy using a multiple lumen intestinal probe]. AB - HISTORY AND ADMISSION FINDINGS: A 68-year-old woman was admitted with upper abdominal symptoms, epigastric fullness and nausea, eight days after initiation of heparin anticoagulation for deep-vein thrombosis. There were no other clinical abnormalities. INVESTIGATIONS: Abdominal ultrasound and computed tomography showed an intramural duodenal haematoma. Endoscopy revealed a stenosing lesion in the duodenum. TREATMENT AND COURSE: Instead of conventional parenteral nutrition, a triple-lumen feeding tube was placed, one lumen in the jejunum for feeding, the other for gastric decompression. Six months later the symptoms had completely disappeared and ultrasound, endoscopy and computed tomography no longer detected any lesion. CONCLUSION: Intrajejunal feeding and gastric decompression via a multi-lumen tube provided effective conservative treatment of duodenal ileus together with gastric decompression. PMID- 15650959 TI - [Adult patients with congenital heart abnormalities: present and future]. AB - The incidence of congenital heart disease is 8 to 10 per 1000 newborn. Due to the advances of heart surgery, medical treatment and interventional cardiology the primarily high mortality rate of relevant congenital heart disease has declined from 80 % to 20 %. Nowadays more than 80 % of children with relevant congenital heart disease reach adulthood. Currently approx. 150 000 adults with significant congenital heart anomalies live in Germany presently. Their number is rising continuously. With the exception of an patent Ductus Botalli and a small atrial septal defect, which could be operated in early childhood curatively, all other congenital heart defects need regular control, since even after successful primary interventional or operative treatment significant residue or sequelae have to be expected. Optimal care of these patients begins in the childhood and must be continued down to the high adulthood. Almost all of them need a careful follow-up in order to recognize residue or sequelae and to treat them in time. The complexity of many congenital heart defects makes a multidisciplinary approach necessary. A specialized, interdisciplinary cooperation between cardiologists, pediatric cardiologists, cardiac surgeons and other specialists is mandatory. PMID- 15650960 TI - [Current aspects of psychosomatic and endocrine stress research]. AB - This review covers basic principles of (neural-)endocrine and psychosomatic aspects and its relationship to stress science. The term stress is rather imprecise, recent reviews defined stress in biological systems as any condition that seriously perturbs the physiological and psychological homeostasis of concerning organism. The review summarizes current knowledge of regulation of the HPA axis and other neural-endocrine-systems. The neurobiological data obtained suggest that inappropriate stress control involve dysfunction of hypothalamo pituitary-adrenocortical (HPA) axis, limbic system and other brain regions. Stress induced different functional (increase of catecholamines and glucocorticoids) and structural changes in brain regions such as the hippocampus, with suppression of adult neurogenesis and synaptic plasticity. These stress associated changes can potentially influence among other things learning and memory processes. The plasticity of brain allows to conceptualize a neurobiological perspective on psychotherapy that reflects the dynamic nature of the interaction between genes and the environment, respectively the consequences of stress. These developments point the way towards a new era of psychotherapy research and practice. PMID- 15650961 TI - [Neuroimmunologic aspects of functional-somatic syndromes]. PMID- 15650962 TI - [Revision of the guideline for hospital hygiene and the prevention of infection from the Robert Koch Institute]. PMID- 15650963 TI - [Diagnosis of chronic mesenteric artery ischemia]. PMID- 15650964 TI - [Therapy of bladder weakness]. PMID- 15650966 TI - Diagnostic algorithm for chronic hepatitis C virus infection: role of the new HCV core antigen assay. AB - BACKGROUND: The diagnosis of chronic hepatitis C virus infection is based on nucleic acid testing (NAT) for HCV-RNA. We evaluated whether total HCV core antigen testing could be a substitute for NAT testing. PATIENTS AND METHODS: Samples from 192 untreated chronic HCV positive patients previously tested for HCV-RNA by four different commercially available assays (SuperQuant, Amplicor HCV Monitor v 1.0 and v 2.0, Quantiplex) were tested for total HCV core antigen using the Ortho trak-C assay (Ortho Clinical Diagnostics, Raritan, NJ, USA). Furthermore, 52 HCV-RNA positive paired serum and plasma samples were analysed. Finally, inter-assay coefficients of variation for core antigen were determined by repeated testing of 59 samples. RESULTS: 172/192 (89.6 %) samples from untreated HCV patients showed positive results with the trak-C assay. Importantly, all but two trak-C positive samples were NAT positive. Only four of the twenty trak-C negative samples tested positive by two NAT assays with viral loads below 30,000 copies/mL. Moreover, HCV core antigen levels correlated significantly with HCV-RNA levels (r > 0.72; p > 0.001), gave consistent results in paired serum and plasma samples (r = 0.991), and showed a very low inter-assay variability (r = 0.943) independent of genotype. CONCLUSION: Based on the performance characteristics, easiness of use, and potential lower cost of the core Ag assay, we propose an alternative testing algorithm for establishing the diagnosis of chronic HCV infection in which the trak-C assay could substitute for NAT as the first choice for detection of HCV viraemia in anti-HCV positive individuals. NAT would only be necessary in rare cases with low viral load. PMID- 15650967 TI - [The diagnosis of liver cirrhosis: a comparative evaluation of standard laparoscopy, mini-laparoscopy and histology]. AB - BACKGROUND: The superiority of laparoscopy with guided biopsy over biopsy alone is established. It is still uncertain if this is also true for midi-/ and mini laparoscopy. AIM: The aim of this study was to determine the diagnostic gain of laparoscopic-guided biopsy for standard laparoscopy and the use of midi-/mini laparoscopies. Characteristics of the liver biopsies were investigated with regard to a missed diagnosis of cirrhosis. METHODS: In a consecutive series of 425 patients clinical, laparoscopic and histological findings were investigated. The length and fragmentation of the liver biopsies were investigated with regard to a missed diagnosis of cirrhosis. RESULTS: The sensitivity of laparoscopy in the diagnosis of cirrhosis was 96 %, that of histology 68 %. The sensitivity of histology alone was especially low in macronodular cirrhosis. The sensitivity of both laparoscopy and histology improved from Child A to C. When cirrhosis was apparent, liver biopsies were shorter and more frequently fragmented. However, the diagnosis of cirrhosis was independent of these parameters. CONCLUSION: Mini laparoscopy may replace standard laparoscopy and appears to be superior compared with histology alone. PMID- 15650968 TI - Identification of fibrosis-relevant proteins using DIGE (difference in gel electrophoresis) in different models of hepatic fibrosis. AB - Proteomics became a more and more important technique for the large-scale analysis of proteins during the last years. Two-dimensional (2D) electrophoresis as a major tool of proteomics is a powerful method to compare two different biological stages (e. g. healthy and diseased tissue) and to find differences in their protein pattern. One major problem in proteomics is the gel to gel variation of two-dimensional gel electrophoresis, which could cause artefacts in the detection of expression differences. The "difference in gel electrophoresis" (DIGE) technique allows the separation of two proteomes in the same gel. The protein pools were labelled with different fluorescent dyes and equal amounts of protein were separated in the same gel. Another advantage of DIGE is the possibility to separate an internal standard labelled with a third dye in the same gel to allow quantitative expression analysis. We compared proteomes of three different fibrosis models with the appropriate control (tissue inhibitor of metalloproteinases-1 (TIMP-1) overexpressing HepG2 cells in comparison to a HepG2 control, freshly isolated HSC in comparison to activated HSC and healthy mouse liver in comparison to fibrotic mouse liver). Among the differentially expressed proteins several were already found to be relevant for fibrosis but we also detected some proteins like the selenium binding protein 2 which might be relevant for hepatic fibrosis. PMID- 15650969 TI - [Advances in therapy for ascites and hepatorenal syndrome]. AB - Sequential diuretic treatment of ascites with spironolactone and furosemide is equivalent to initial combination therapy. Orally applicable vasopressin-V2 receptor antagonists are an interesting novel therapeutic approach for the elimination of free water. The therapeutic efficacy for patients with cirrhosis and ascites is currently being investigated in phase II trials. Following paracentesis of up to 6 liters volume, infusion of 3.5 % saline is as effective as 20 % albumin. Another trial confirms the superiority of TIPS for the treatment of massive ascites, also demonstrating survival benefit. Determination of leukocyte esterase activity with a simple stix method may be helpful for the rapid and easy diagnosis of spontaneous bacterial peritonitis. Patients with hepatorenal syndrome seem to benefit from a combination of terlipressin and albumin whereas the effect of albumin dialysis on survival remains to be proven. PMID- 15650970 TI - [Gastrointestinal bleeding in portal hypertension in liver cirrhosis]. AB - There are three major goals in the prophylaxis and treatment of upper gastrointestinal bleeding in portal hypertensive patients: prophylaxis of the first bleeding episode, therapy of active bleeding and prophylaxis of recurrent bleeding. Several therapeutic options are available: non-selective beta-blockers are the treatment of choice in the primary prophylaxis of the first bleeding episode in patients with large esophageal varices. Alternatively, endoscopic band ligation therapy is an option. Acute bleeding varices should be treated by ligation pharmacological and antibiotic therapy. Prophylaxis of recurrent bleeding is patient-dependent: shunt surgery is an option in young patients in a good medical condition (Child-Pugh class A). In patients with refractory ascites and a bilirubin below 3 mg/dl, TIPS is a good option together with recurrent bleeding. At the moment, there are no trials showing that endoscopic ligation therapy is superior to prevent pharmacological therapy. Nevertheless, the first line treatment in most patients in Germany is endoscopic band ligation. Bleeding from ectopic varices and bleeding from hypertensive gastropathy should be treated individually either by endoscopy, TIPS or drug therapy. PMID- 15650971 TI - [Local radiofrequency ablation of liver lesions--possibilities and limitations]. AB - Radiofrequency ablation (RFA) is currently the most established technique for the local destruction of tumors within the liver. It is technically straightforward, can be repeated and combines a high efficiency in tissue destruction with a low to moderate incidence of complications. This review explains the principle of RFA and lists currently available systems. In addition, the rationale behind RFA is discussed, and an overview of clinical indications, results and complications is presented. One of the main problems in assessing the clinical value of RFA is the fact that no large and controlled randomized studies are available, while the number of ablations and treated tumor entities are steadily increasing. From a technical point of view, the major problem is the still limited volume of necrosis that is achievable. The indication for RFA has to be discussed as interdisciplinary. A complete tumor necrosis should be the aim of all ablations, with the exception of symptomatic metastases of neuroendocrine tumors. The value of additional chemotherapy to RFA is still unclear, especially in the treatment of liver metastases. PMID- 15650974 TI - Strict blood glucose control in the ICU: panacea or Pandora's box? PMID- 15650975 TI - Improving glycemic control in the cardiothoracic intensive care unit: clinical experience in two hospital settings. AB - OBJECTIVE: Recent studies suggest that strict perioperative glycemic control improves clinical outcomes after cardiothoracic surgery. However, optimal methods and targets for controlling blood glucose (BG) levels in this setting have not been established. Currently published intensive insulin infusion protocols (IIPs) have important practical limitations, which may affect their utility. In this article, the authors present their experience with a safe, effective, nurse driven IIP, which was implemented simultaneously in 2 cardiothoracic intensive care units (CTICUs). DESIGN: Prospective cohort study. SETTING: Tertiary referral hospital and community teaching hospital. PARTICIPANTS: CTICU patients. INTERVENTIONS: A standardized, intensive IIP was used for all patients admitted to both CTICUs. Hourly BG levels, relevant baseline variables, and clinical interventions were collected prospectively from the active hospital chart and CTICU nursing records. MEASUREMENTS AND MAIN RESULTS: The IIP was used 137 times in 118 patients. The median time required to reach target BG levels (100-139 mg/dL) was 5 hours. Once BG levels decreased below 140 mg/dL, 58% of 2,242 subsequent hourly BG values fell within the narrow target range, 73% within a "clinically desirable" range of 80 to 139 mg/dL, and 94% within a "clinically acceptable" range of 80 to 199 mg/dL. Only 5 (0.2%) BG values were less than 60 mg/dL, with no associated adverse clinical events. CONCLUSIONS: The IIP safely and effectively improved glycemic control in 2 CTICUs, with minimal hypoglycemia. Based on prior studies showing the benefits of strict glycemic control, the implementation of this IIP should help to reduce morbidity and mortality in CTICU patients. PMID- 15650976 TI - Off-pump versus on-pump coronary artery bypass surgery and postoperative pulmonary dysfunction. AB - OBJECTIVE: To investigate how off-pump coronary artery bypass grafting (CABG) affects postoperative pulmonary function when compared with on-pump CABG. DESIGN: Prospective clinical study. SETTING: University-affiliated teaching hospital. PARTICIPANTS: Adult patients (n = 39) undergoing elective coronary artery bypass surgery with or without cardiopulmonary bypass. INTERVENTIONS: Two groups of patients were compared: 19 consecutive patients undergoing off-pump CABG surgery and 20 consecutive patients undergoing conventional CABG surgery. MEASUREMENTS AND MAIN RESULTS: Pulmonary function tests (flow volume loops and lung volumes with plethysmography) were done preoperatively and 72 hours postoperatively. Arterial blood gases and PaO2/FIO2 were measured at various stages. Sequential chest x-rays were obtained and evaluated for pleural changes, pulmonary edema, and atelectasis. In both groups, PaO2/FIO2 ratios decreased progressively throughout the perioperative period, with no significant differences between the groups at any stage during the study. There was a significant decline in postoperative pulmonary function tests in both groups, but there was no difference between groups at 72 hours postoperatively. No differences were found in the time to extubation, atelectasis scores, or postoperative complications. CONCLUSIONS: Off-pump CABG does not confer major protection from postoperative pulmonary dysfunction compared with CABG surgery with CPB. Strategies for minimizing pulmonary impairment after CABG surgery should be directed to factors other than the use of CPB. PMID- 15650977 TI - Predictors of atrial fibrillation after off-pump coronary artery bypass graft surgery. AB - OBJECTIVE: Atrial fibrillation is the most common complication after coronary artery bypass graft surgery. This arrhythmia may lead to hemodynamic compromise, prolonged hospitalization, and increased risk for cerebral thromboembolism. Older age is the only variable consistently associated with the development of postoperative atrial fibrillation; however, no strong predictive model exists. The purpose of this study was to identify perioperative characteristics associated with new-onset atrial fibrillation in patients undergoing off-pump coronary artery bypass grafting. DESIGN: Prospective, observational. SETTING: University tertiary care hospital. PARTICIPANTS: One hundred sixty consecutive patients undergoing off-pump coronary artery bypass grafting. INTERVENTIONS: None. MEASUREMENTS AND MAIN RESULTS: Incidence of postoperative atrial fibrillation was the major outcome. Atrial fibrillation occurred in 33 patients (20.6%). Multivariate analysis identified reintervention (odds ratio 26.8), revascularization of the ramus medianus (odds ratio 3.9), and age (odds ratio 1.069 per year) as the only independent predictors of postoperative atrial fibrillation. All patients were in sinus rhythm at hospital discharge. One hospital death was noted. CONCLUSIONS: Despite the less invasive approach, the incidence of postoperative atrial fibrillation is high after off-pump coronary artery bypass grafting. Older age, grafting of the ramus medianus, and a redo operation were predictors of new-onset postoperative atrial fibrillation. It is possible that left atrial stretching with heart dislocation during revascularization of the lateral wall could lead to postoperative atrial fibrillation. PMID- 15650978 TI - Effects of amiodarone and thoracic epidural analgesia on atrial fibrillation after coronary artery bypass grafting. AB - OBJECTIVE: This study was designed to assess the effects of a perioperative dosing regimen of amiodarone administration, high thoracic epidural anesthesia (TEA), or a combination of the 2 regimens on atrial fibrillation (AF) after coronary artery bypass grafting (CABG). DESIGN AND SETTING: The study was prospective, controlled, and randomized and was performed in a tertiary health care center associated with a university. PARTICIPANTS: One hundred sixty-three patients scheduled for coronary artery bypass graft surgery. INTERVENTIONS: In this 2 x 2 factorial-designed study the patients were randomized to 1 of 4 regimens in which group E had perioperative TEA, group E+A had TEA and amiodarone, group A had amiodarone, and group C served as control. The epidural catheter was inserted at T1-3 the day before surgery. TEA groups received TEA for 96 hours. The amiodarone regimen consisted of a single loading dose of 1,800 mg of amiodarone orally. Intravenous infusion of amiodarone was started after induction of anesthesia and was administered at 900 mg over 24 hours for the subsequent 3 days. MEASUREMENTS AND MAIN RESULTS: AF was documented using Holter monitoring. In group E 22 of 44 (50%), in group E+A 10 of 35 (28.6%), in group A 10 of 36 (27.8%), and in the control group 20 of 48 (41.7%) patients developed AF (odds ratio amiodarone/nonamiodarone 0.47 [0.24-0.90]; P = 0.02). CONCLUSIONS: The perioperative amiodarone regimen used in this study was effective in reducing the incidence of AF after CABG while TEA was not. PMID- 15650979 TI - The use of the SonoSite ultrasound device for intraoperative evaluation of the aorta. AB - OBJECTIVE: Neurologic complications remain a significant cause of morbidity and mortality in cardiac surgery. Risk factors for neurologic injuries include the presence of atheromatous disease in the aorta. Epiaortic ultrasound has been shown to be superior in detecting the extent and location of atheromatous disease. The SonoSite Corporation (Bothell, WA) has recently introduced an affordable, portable, high-resolution ultrasound device. This new device was compared with the Hewlett-Packard Sonos 5550 ultrasound device (currently manufactured by Philips, Andover, MA) to determine suitability for this purpose. DESIGN: Prospective, serial comparison of 2 devices. SETTING: University hospital. PARTICIPANTS: Fifty consecutive cardiac surgery patients. INTERVENTIONS: Intraoperative epiaortic ultrasound images were obtained using a SonoSite 180 Plus ultrasound device and a Hewlett-Packard Sonos 5500 ultrasound device. Three observers graded recorded images based on extent of atheromatous disease. MEASUREMENTS AND MAIN RESULTS: Two patients were excluded because of errors in recording images. For the 48 remaining patients, consensus (median) grades had an observed agreement of 93.6% compared with a chance agreement of 67.7%. This correlates to a kappa value of 0.80 or near-excellent agreement. CONCLUSIONS: The near-excellent agreement of the 2 devices is acceptable, thus providing a unique opportunity to expand the use of epiaortic ultrasound imaging. PMID- 15650980 TI - Assessment of the tricuspid valve: a comparison of four transesophageal echocardiographic windows. AB - OBJECTIVE: Tricuspid valve annular dilation is associated with persistent tricuspid valve regurgitation (TR) after cardiac surgery. The purpose of this study is to compare assessment of the tricuspid valve annulus from 4 different echocardiographic windows. DESIGN: Prospective study. SETTING: Single tertiary care hospital. PATIENTS: Twenty patients undergoing cardiac surgery, including right atriotomy. METHODS: Measurements of the tricuspid valve annular diameter during ventricular diastole (TVADd) and of TR jet dimensions, obtained from 3 different transesophageal and 1 transgastric echocardiographic window, were compared with each other and with that measured by the surgeon during cardiopulmonary bypass. Data were compared using correlation statistics, bias analysis, and analysis of variance for repeated measures. RESULTS: TVAD measured by the surgeon both agreed and correlated best with TVADd measured from the transgastric right ventricular inflow/outflow window (r = 0.98; mean bias 0.04 cm). The proximal TR jet widths and TR jet lengths were larger when measured from midesophageal 4-chamber and midesophageal right ventricular inflow/outflow windows compared with other windows (p < 0.01). CONCLUSION: TVADd was more accurately measured from transgastric windows, whereas the measurements of the TR jet were greater from esophageal windows. These data support imaging from multiple windows to obtain a more comprehensive assessment of the tricuspid valve anatomy and function. PMID- 15650981 TI - Diagnosis of patent foramen ovale with multiplane transesophageal echocardiography in adult cardiac surgical patients. AB - OBJECTIVE: To evaluate multiplane transesophageal echocardiography (TEE) for detection of patent foramen ovale (PFO) and to compare multiplane TEE with visual inspection (VI) for PFO detection. DESIGN: A prospective observational study. SETTING: University hospital (single institution). PARTICIPANTS: Patients presenting for cardiac surgery requiring TEE. INTERVENTIONS: Multiplane TEE including 2 atrial views with color-flow Doppler (CFD) and contrast echocardiography (CE) with a provocative respiratory maneuver (PRM) and comparison of multiplane TEE and VI with respect to PFO detection. MEASUREMENTS AND MAIN RESULTS: The cohort size was 187. PFO prevalence was 27.3%. CFD with serial decrease of the Nyquist limit detected 51% of all PFO: 41.2% in the bicaval view alone, 27.5% in the 4-chamber view alone, and 9.8% in both views. CE detected 78.4% of all PFO: 72.5% with PRM, 45.1% with no PRM, and 27.4% with/without PRM. PFO detection by multiplane TEE and visual inspection were correlated in 41 subjects. TEE diagnosed 11 PFO (26.8% prevalence, 3 missed by VI). VI diagnosed 12 PFO (29.3% prevalence, 4 missed by TEE). CONCLUSIONS: Multiplane TEE is a gold standard for detection of PFO. Despite advances in TEE technology, 2-dimensional imaging does not detect all PFO. To maximize PFO detection, multiple TEE modalities are required in multiple views, despite a low Nyquist limit for CFD or a PRM for CE. Even though multiplane TEE is equivalent to VI for PFO detection, the discrepancy rate may be an important consideration in the individual case. PMID- 15650982 TI - Alterations of mesenteric blood flow after cardiopulmonary bypass: a Doppler sonographic study. AB - OBJECTIVE: Mesenteric ischemia after cardiopulmonary bypass is a serious complication associated with high mortality. It was the aim of this study to investigate mesenteric blood flow with the help of Doppler sonography in asymptomatic patients before and after cardiopulmonary bypass and cardiac surgery. DESIGN: Observational study of consecutive patients. SETTING: Nonuniversity cardiac center. PARTICIPANTS: Twenty-five patients undergoing elective coronary revascularization with normal left ventricular function and stable postoperative circulations with no need for catecholamines. MEASUREMENTS AND MAIN RESULTS: Preoperative and postoperative color and CW-Doppler sonography of the superior mesenteric artery. Diameter of the superior mesenteric artery and the Doppler flow profile were analyzed. Preoperative and postoperative hemodynamic data were measured by using a pulmonary artery thermodilution catheter. Mesenteric systolic flow velocity was 135 +/- 11 cm/s preoperatively and 193 +/- 13 cm/s postoperatively (p < 0.05). The corresponding preoperative and postoperative values of diastolic flow velocity were 14 +/- 4 and 4 +/- 2 cm/s (p < 0.05) and the values of mean flow velocity were 24 +/- 3 and 17 +/- 2 cm/s (p < 0.05), respectively. The preoperative Pourcelot resistive index was 0.87 +/- 0.05, and the preoperative Gosling pulsatility index was 4.6 +/- 0.5. Both indices increased postoperatively to values of 0.98 +/- 0.04 and 9.5 +/- 0.7, respectively (p < 0.05). Preoperative and postoperative hemodynamic data did not differ significantly. CONCLUSION: The postoperative changes in the flow velocities and the increases of the resistive and pulsatility index are indications of rigidity of the mesenteric vascular bed and decreased mesenteric perfusion after cardiopulmonary bypass. PMID- 15650983 TI - Surgery for mesenteric infarction: prognostic factors associated with early death within 72 hours. AB - OBJECTIVE: The aim of this study was to look for preoperative and postoperative prognostic factors for early mortality, likely to be of use to clinicians in decision making. DESIGN: Prospective multicenter study. SETTING: This study was conducted in 4 university hospitals in Northeast France. PARTICIPANTS: One hundred thirty-one patients with mesenteric infarction confirmed by pathologic examination were included. INTERVENTIONS: All patients underwent surgery and were hospitalized in the intensive care unit. MAIN RESULTS: Twenty-eight patients (21.3%) underwent exploratory laparotomy only; 103 patients underwent bowel resection with/or without associated revascularization. The overall mortality rate at discharge was 74.8%. In the first 3 days, 60% of deaths occurred. The prognostic factors for death within 72 hours, obtained by logistic regression, were preoperative heart failure, lactate level over 5 mmol/L, aspartate aminotransferase over 200 IU/L, and total cholesterol level below 80 mg/dL, or procalcitonin level over 40 ng/L. From these results, a mortality prognostic score was derived. Probability of mortality within 72 hours was estimated to be 5% for patients with none of these factors and 97% for those with all 4. For deaths occurring after 72 hours, the only mortality prognostic factor was the existence of necrosed areas at the ends of bowel resections. CONCLUSION: When patients have fewer than 3 of the prognostic factors described in the score, aggressive medical and surgical strategies could be appropriate. If necrosis is recorded at the ends of the resection, renewed surgery should be undertaken as soon as signs of new disturbances appear. PMID- 15650985 TI - Pharmacodynamics and pharmacokinetics of high-dose oxycodone infusion during and after coronary artery bypass grafting. AB - OBJECTIVE: In small to moderate doses, oxycodone has similar analgesic efficacy to morphine with fewer side effects. The present study evaluated the pharmacokinetics and dynamics of high doses of oxycodone during anesthesia for primary coronary artery bypass grafting. DESIGN: A randomized, prospective clinical evaluation. SETTING: A major Scandinavian university clinic. PARTICIPANTS: Two groups with 10 patients each were studied. INTERVENTIONS: Invasive hemodynamics, echocardiograms, and electrocardiograms were monitored. Oxycodone kinetics, histamine liberation, and plasma cortisol levels were measured. Anesthesia was induced with 1.0 mg/kg of oxycodone and, thereafter, in a random order, maintained with a continuous infusion of oxycodone at a rate of either 0.5 mg/kg/h (group OX 0.5, 10 patients) or 1.0 mg/kg/h (group OX 1.0, 10 patients). An additional bolus dose of 0.5 mg/kg (OX 0.5) or 1.0 mg/kg (OX 1.0) of oxycodone was given before the incision. Enflurane was administered according to hemodynamic criteria. MEASUREMENTS AND MAIN RESULTS: The induction of and the course of anesthesia were hemodynamically stable in all patients. Enflurane was given to every patient. The mean total doses of oxycodone were 3.5 mg/kg (OX 0.5) and 6.2 mg/kg (OX 1.0). The median t(1/2) of oxycodone varied from 5.1 to 5.9 hours. No hemodynamic differences were found between the groups. No histamine liberation was detected. During anesthesia, the predominant waves in the EEG were theta;- and delta-waves. The mean times to awakening were 3.8 hours and 7.0 hours in the groups OX 0.5 and 1.0, respectively. All patients were intubated until the first postoperative morning. No recall of awareness was reported. CONCLUSION: A combination of oxycodone and enflurane provides hemodynamically stable anesthesia. No advantages were gained with the higher dose. Elimination of oxycodone was slower than reported previously. PMID- 15650984 TI - Opioid-sparing effects of diclofenac and paracetamol lead to improved outcomes after cardiac surgery. AB - OBJECTIVE: This study assessed the analgesic efficacy, side effects, time to extubation, and oxygenation of 3 analgesic regimens after coronary artery bypass surgery using diclofenac, paracetamol, and placebo suppositories. DESIGN: Prospective, randomized, double-blind, placebo-controlled study. SETTING: Referral center for cardiothoracic surgery at a university hospital. PARTICIPANTS: Sixty consenting adults scheduled for elective coronary artery bypass grafting (CABG). INTERVENTIONS: Patients were divided into 3 groups postoperatively: diclofenac/paracetamol: diclofenac, 100 mg rectally, and paracetamol, 1 g rectally. Diclofenac was repeated after 18 hours and paracetamol every 6 hours for 24 hours; diclofenac: diclofenac as in diclofenac/paracetamol, with placebos replacing paracetamol; and placebo: 2 placebo suppositories at same times as diclofenac/paracetamol. All patients received morphine patient controlled analgesia. RESULTS: Twenty-four hour morphine consumption with diclofenac/paracetamol was 12 +/- 6 mg, diclofenac 22 +/- 13 mg, and placebo 37 +/- 15 mg (diclofenac/paracetamol and diclofenac, p = 0.0003 and p = 0.0159 compared with placebo). Patients in the placebo group had significantly greater pain scores at 12 and 24 hours compared with diclofenac/paracetamol and diclofenac. Extubation time was significantly prolonged in the placebo group compared with the diclofenac/paracetamol and diclofenac groups (mean [SD] minutes diclofenac/paracetamol, diclofenac, and placebo 478 [150], 487 [257], and 710 [326], respectively). Oxygenation following extubation was significantly lower in the placebo group compared with the diclofenac/paracetamol and diclofenac groups (mean [SD] mmHg: diclofenac/paracetamol, diclofenac, and placebo 175 [44], 157 [43], and 117 [22], respectively). Episodes of nausea and vomiting were significantly less in the diclofenac/paracetamol and diclofenac groups than in the placebo group (46% and 51% reduction, respectively). all groups had similar blood loss and change in serum creatinine. CONCLUSION: Diclofenac alone or with paracetamol has a significant opioid-sparing effect after CABG, producing more rapid extubation and better oxygenation. PMID- 15650987 TI - Epidural hematoma complicating high thoracic epidural catheter placement intended for cardiac surgery. PMID- 15650986 TI - Comparison of analgesic effects of morphine, fentanyl, and remifentanil with intravenous patient-controlled analgesia after cardiac surgery. AB - OBJECTIVE: The purpose of this study was to compare the analgesic effects of remifentanil with 2 other opioid agents, morphine and fentanyl, after cardiac surgery. DESIGN: Prospective, randomized, and double-blinded study. SETTINGS: This study was performed at Uludag University hospital. PARTICIPANTS: Seventy five patients undergoing off-pump coronary artery bypass surgery were included in the study. INTERVENTIONS: Anesthesia was standardized. Cases were randomized into 3 groups consisting of 25 patients in each. Groups M, F, and R were given morphine HCl (1 mg/mL) with an infusion rate of 0.3 mg/h and 1-mg bolus doses; fentanyl (50 microg/mL) with an infusion rate of 1 microg/kg/h and 10-microg bolus; and, remifentanil (50 microg/mL) with an infusion rate of 0.05 microg/kg/min and 0.5-microg/kg bolus, respectively. Continuous infusion was started immediately after the completion of the surgery. MEASUREMENTS AND MAIN RESULTS: Pain was assessed by using a visual analog scale (0-10), and sedation was assessed with the Ramsey sedation score (1-6) 30 minutes, 1, 2, 4, 12, and 24 hours after extubation. The number of boluses and demands, time to extubation, and side effects were analyzed. Visual analog scale, sedation scores, and mean extubation times were similar in all groups. Total number of boluses and demands were statistically more in the remifentanil group. Regarding the side effects, nausea and vomiting was higher in group M (p < 0.05), whereas itching was prominent in group F (p < 0.05). CONCLUSIONS: Despite the different durations of these 3 opioid agents, the infusion dose of remifentanil was as effective as morphine and fentanyl after OPCAB surgery with fewer side effects. PMID- 15650988 TI - The use of a pulmonary artery catheter after emergency cardiac surgery to reduce shunt through a patent foramen ovale. PMID- 15650989 TI - Intraoperative transesophageal echocardiography detects a sheared catheter wire and alters surgical approach in a Jehovah's Witness patient undergoing coronary artery bypass graft surgery. PMID- 15650990 TI - Pulseless electrical activity after induction of anesthesia: a witnessed cardiac rupture. PMID- 15650991 TI - Resuscitation of a patient with giant left atrial myxoma after cardiac arrest. PMID- 15650992 TI - Successful treatment of dilative cardiomyopathy in a 12-year-old girl using the calcium sensitizer levosimendan after weaning from mechanical biventricular assist support. PMID- 15650993 TI - Use of sildenafil with inhaled nitric oxide in the management of severe pulmonary hypertension. PMID- 15650994 TI - Pumpless extracorporeal lung assist in severe blunt chest trauma. PMID- 15650995 TI - Brain natriuretic peptide: a diagnostic and treatment hormone for perioperative congestive heart failure. PMID- 15650996 TI - Anomalous right coronary artery originating from the left sinus of Valsalva. PMID- 15650997 TI - Pro: Ischemic preconditioning has a myocardial protective effect during coronary artery bypass surgery. PMID- 15650998 TI - Con: Ischemic preconditioning is not necessary because volatile agents accomplish it. PMID- 15650999 TI - Subcutaneous pulsating neck mass after left internal jugular venipuncture. PMID- 15651000 TI - An unusual transesophageal echocardiographic finding during femoral embolectomy. PMID- 15651001 TI - Anomaly in the descending thoracic aorta. PMID- 15651002 TI - Anesthesia in a patient with nasopharyngeal angiofibroma and hemophilia A. PMID- 15651003 TI - Chronic paravalvular abscess after aortic valve replacement. PMID- 15651005 TI - Anesthetic implications of recurrent laryngeal nerve palsy after esophagectomy. PMID- 15651004 TI - Abdominal wall mucormycosis after heart transplantation. PMID- 15651006 TI - Atrial septal defect or patent foramen ovale. PMID- 15651007 TI - The growing field of anesthesia for liver transplantation. PMID- 15651008 TI - Detecting protein-protein interaction in live yeast by flow cytometry. AB - BACKGROUND: The yeast Saccharomyces cerevisiae is the most commonly used organism for studying protein- protein interactions. In this report we demonstrate the use of flow cytometry in observing fluorescence resonance energy transfer (FRET) between cyan and yellow fluorescent fusion proteins (CFP and YFP, respectively) as a marker for protein interaction in live yeast cells. Probability binning is also employed to provide a statistical confirmation of our observations. METHODS: We coexpressed CFP and YFP fusions containing the N-terminal transmembrane domain (NTM) of Tom70p in yeast and analyzed FRET in live cells with a multilaser flow cytometer. The Tom70p NTM was previously shown to be sufficient for mitochondrial localization and protein-protein interaction (Millar and Shore, 1994, J Biol Chem 269:12229-12232). RESULTS: FRET was observed only in cells that expressed CFP and YFP fusions that each contained the wild-type NTM. The introduction of mutations previously shown to disrupt NTM interaction eliminated FRET. Probability binning confirmed that differences between the FRET channels of experimental and control samples were statistically and physiologically significant. CONCLUSION: Flow cytometric analysis of FRET in yeast is a powerful technique for studying protein protein interactions. The use of flow cytometry allows FRET data to be gathered from a large number of individual cells, thus providing important advantages unavailable to other techniques. Its application to yeast presents a new method to a popular system widely used in proteomic studies. PMID- 15651009 TI - A novel approach to prepare extended DNA fibers in plants. AB - BACKGROUND: The extended DNA fiber preparation procedure is still imperfect in plants due to the existence of a hard cell wall; thus, high quality of extended DNA fibers for fluorescence in situ hybridization (FISH) analysis is often difficult to be obtained rapidly and efficiently. In this study we have developed a fast and widely effective method to prepare DNA fibers from various plant species and the fibers are suitable for fiber FISH mapping. METHODS: Fresh young leaves were chopped with a sharp sterile scalpel in a Petri dish that contained ice-cold nucleus isolation buffer followed by filtration through 33-mum nylon mesh. Nuclei were obtained by centrifuging the filtrates at high speed (16,000g) for 40 s. Nucleus lysis buffer (0.5% sodium dodecylsulfate, 5 mM ethylenediaminetetraacetic acid, 100 mM Tris, pH7.0) was added to nuclei on slides, and DNA fibers were dragged and extended with a clean coverslip. RESULTS: The key of this method is that liquid nitrogen grinding of leaves is replaced by chopping with a blade in ice-cold nucleus isolation buffer. With the liquid nitrogen method, over- or under-grinding of leaves occurs more frequently, and DNA fibers with the desired quality are not obtained easily. In contrast, it is easier to release nuclei from cells in nucleus isolation buffer by chopping, which results in fewer nuclei being destroyed. Highly extended, intact, and long DNA fibers can be generated to a great probability with this method. In addition, this method is very simple and rapid, requiring only 20 min for the entire process, and is also safe because poisonous mercaptoethanol is replaced by dithiothreitol. The results of fiber-FISH with maize genomic DNA and 45S rDNA as probes showed that DNA fiber size as long as 1.96 Mb could be measured. The successful and reliable preparation of maize, wild rice, and barley DNA fibers suitable for FISH mapping proves that this technique is a widely effective approach for obtaining extended DNA fibers in plants. CONCLUSIONS: A simple, rapid, safe, and widely effective method for getting extended DNA fibers has been developed in plants. (c) 2005 Wiley-Liss, Inc. PMID- 15651010 TI - Methyl group rotation driven by proton transfer through a long-range chemical interaction. PMID- 15651012 TI - Using dipoles to control the directionality of functional groups: syn- and anti oriented benzene-1,3-dicarboxamides. PMID- 15651011 TI - Synthesis of the heterocyclic core of the GE 2270 antibiotics and structure elucidation of a major degradation product. PMID- 15651013 TI - Photomagnetic effects in polycyanometallate compounds: an intriguing future chemically based technology? PMID- 15651014 TI - Mass production and high photocatalytic activity of ZnS nanoporous nanoparticles. PMID- 15651015 TI - Direct organocatalytic de novo synthesis of carbohydrates. PMID- 15651016 TI - Determination of chlortetracycline in swine plasma by LC-ESI/MS/MS. AB - A rapid, selective and sensitive method has been developed for the determination of chlortetracycline in swine plasma by LC-ESI/MS/MS. The method consists of a protein precipitation extraction for sample preparation and liquid chromatography ionspray tandem mass spectrometry for analysis. The plasma samples were extracted with acetonitrile and the supernatants were analyzed using an LC-ESI/MS/MS instrument. Separation was achieved using a C(8) analytical column and an isocratic mobile phase composed of 70:30 acetonitrile:0.5% formic acid in water at a flow rate of 500 microL/min. A linear (weighted 1/concentration) relationship was used to perform the calibration over an analytical range 20- 2000 ppb (ng/mL). The intra-batch precision and accuracy at LLOQ, medium and high concentrations were 9.0, 11.3 and 9.9% and 97.7, 100.3 and 98.4%, respectively, and the inter-batch precision and accuracy at LLOQ, medium and high concentrations were 9.1, 8.4 and 7.4% and 95.1, 102.1 and 97.1%, respectively. This LC-ESI/MS/MS method for the determination of chlortetracycline in swine plasma has been proven to be within generally accepted criteria used for bioanalytical assay. PMID- 15651017 TI - Organic intercalation material: reversible change in interlayer distances by guest release and insertion in sandwich-type inclusion crystals of cholic acid. AB - Cholic acid (CA) forms inclusion crystals that have a sandwich-type lamellar structure constructed by the alternative stacking of host bilayers and guest layers. Five disubstituted benzenes, o-toluidine, m-fluoroaniline, o chlorotoluene, o-bromotoluene, and indene, are accommodated in the two dimensional void space between the host bilayers at 1:2 host-guest stoichiometries. Thermal gravimetric analysis of the inclusion crystals revealed that all the guest molecules, except o-toluidine, are released in two separate steps, indicating the formation of intermediate crystals after the first guest release. Adequate heat treatment of the four inclusion crystals induces release of half or three quarters of the guest molecules. X-ray diffraction patterns of the intermediate crystals revealed that the crystals have a bilayer structure the same as those of the common CA inclusion crystals. They have one-dimensional cavities, in which the guest molecules are included at a 1:1 or 2:1 host-guest stoichiometry. These facts indicate that the host bilayers move 1.6-4.5 A perpendicular to the layer direction by desorption of the guest molecules. Furthermore, a reverse structural change is also achieved by absorption of the guest molecules to regenerate the starting sandwich-type inclusion crystals. This reversible change in the host bilayer by the guest sorption and desorption is a novel example of organic intercalation materials. PMID- 15651018 TI - Ruthenium-catalyzed propargylic substitution reactions of propargylic alcohols with oxygen-, nitrogen-, and phosphorus-centered nucleophiles. AB - The scope and limitations of the ruthenium-catalyzed propargylic substitution reaction of propargylic alcohols with heteroatom-centered nucleophiles are presented. Oxygen-, nitrogen-, and phosphorus-centered nucleophiles such as alcohols, amines, amides, and phosphine oxide are available for this catalytic reaction. Only the thiolate-bridged diruthenium complexes can work as catalysts for this reaction. Results of some stoichiometric and catalytic reactions indicate that the catalytic propargylic substitution reaction proceeds via an allenylidene complex formed in situ, whereby the attack of nucleophiles to the allenylidene C(gamma) atom is a key step. Investigation of the relative rate constants for the reaction of propargylic alcohols with several para-substituted anilines reveals that the attack of anilines on the allenylidene C(gamma) atom is not involved in the rate-determining step and rather the acidity of conjugated anilines of an alkynyl complex, which is formed after the attack of aniline on the C(gamma) atom, is considered to be the most important factor to determine the rate of this catalytic reaction. The key point to promote this catalytic reaction by using the thiolate-bridged diruthenium complexes is considered to be the ease of the ligand exchange step between a vinylidene ligand on the diruthenium complexes and another propargylic alcohol in the catalytic cycle. The reason why only the thiolate-bridged diruthenium complexes promote the ligand exchange step more easily with respect to other monoruthenium complexes in this catalytic reaction should be that one Ru moiety, which is not involved in the allenylidene formation, works as an electron pool or a mobile ligand to another Ru site. The catalytic procedure presented here provides a versatile, direct, and one-step method for propargylic substitution of propargylic alcohols in contrast to the so far well-known stoichiometric and stepwise Nicholas reaction. PMID- 15651019 TI - Polymers of intrinsic microporosity (PIMs): bridging the void between microporous and polymeric materials. AB - Novel types of microporous material are required for chemoselective adsorptions, separations and heterogeneous catalysis. This concept article describes recent research directed towards the synthesis of polymeric materials that possess microporosity that is intrinsic to their molecular structures. These polymers (PIMs) can exhibit analogous behaviour to that of conventional microporous materials, but, in addition, may be processed into convenient forms for use as membranes. The excellent performance of these membranes for gas separation and pervaporation illustrates the unique character of PIMs and suggests immediate technological applications. PMID- 15651020 TI - Electron-donating alkoxy-group-driven synthesis of heteroleptic tris(phthalocyaninato) lanthanide(III) triple-deckers with symmetrical molecular structure. AB - Reaction of heteroleptic bis(phthalocyaninato) lanthanide compounds [(Pc)M{Pc(OC8H17)8}] [H2Pc=unsubstituted phthalocyanine; H2Pc(OC8H17)8 = 2,3,9,10,16,17,23,24-octakis(octyloxy)phthalocyanine] with monomeric complexes [(Pc)M(acac)] (Hacac=acetylacetone), both of which generated in situ, led to the isolation of heteroleptic phthalocyaninato-[2,3,9,10,16,17,23,24 octakis(octyloxy)phthalocyaninato] lanthainde(III) triple-decker complexes [(Pc)M{Pc(OC8H17)8}] (M=Gd-Lu) (1-8) as the sole product. Heterodinuclear analogues [(Pc)Lu{Pc(OC8H17)8}M(Pc)] (M=Gd-Yb) (9-15) were obtained in a similar manner from the reaction of [(Pc)M{Pc(OC8H17)8}] (M=Gd-Yb) and [(Pc)Lu(acac)]. The molecular structures of the herterodinuclear compound [(Pc)Lu{Pc(OC8H17)8}Er(Pc)] (13) and its homodinuclear counterparts [(Pc)M{Pc(OC8H17)8}M(Pc)] (M=Er, Lu) (5, 8) have been determined by X-ray diffraction analysis; these structures exhibit a symmetrical molecular structure with one inner planar Pc(OC8H17)8 ligand and two outer domed Pc ligands. In addition to various spectroscopic analyses, the electrochemistry of these compounds has also been studied by cyclic voltammetry (CV) and differential pulse voltammetry (DPV) methods, revealing the gradually enhanced pi-pi interactions among the phthalocyanine rings in the triple-deckers along with the lanthanide contraction. PMID- 15651021 TI - New types of blue, red or near IR luminescent phosphonate-decorated lanthanide oxalates. AB - Hydrothermal reactions of the lanthanide chlorides with MeN(CH2CO2H)(CH2PO3H2), (H3L1) (or Me2NCH2PO3H2, H2L2) and sodium oxalate lead to seven new lanthanide oxalate phosphonate hybrids with three types of 3D network structures, namely, [Ln(C2O4){MeNH(CH2CO2)(CH2PO3H)}]0.5 H2O (Ln=Nd: 1; Eu: 2; Gd: 3), [Ln4(C2O4)5(Me2NHCH2PO3)2(H2O)4]2 H2O (Ln=La: 4, Nd: 5), [Ln3(C2O4)4(Me2NHCH2PO3)(H2O)6]6 H2O (Gd: 6, Er: 7). Their structures have been established by X-ray single-crystal diffraction. Complexes 1-3 are isostructural and feature a 3D network formed by the interconnection of 3D network of {Ln(H2L1)}2+ with 1D chains of {Ln(C2O4)}+. Complexes 4 and 5 are isostructural and feature a complex 3D network built from 3D network of lanthanide oxalate and {Ln4(HL2)2} units. The isostructural 6 and 7 form another type of 3D network composed of porous lanthanide-oxalate network inserted by 1D chains of lanthanide oxalate phosphonate. Compounds 1, 5 and 7 are luminescent materials in the near IR region. Compounds 3 and 6 exhibit a broad blue fluorescent emission band at 451 and 467 nm, respectively. Compound 2 displays very strong and sharp emission bands at 592, 616 and 699 nm with a long luminescent lifetime of 1.13 ms. PMID- 15651022 TI - Generation and reactions of ruthenium phosphido complexes [(eta5 C5H5)Ru(PR'3)2(PR2)]: remarkably high phosphorus basicities and applications as ligands for palladium-catalyzed Suzuki cross-coupling reactions. AB - Reactions of [(eta5-C5H5)Ru(PR'3)2(Cl)] with NaBAr(F) [BAr(F)-=B{3,5 [C6H3(CF3)2]}4-; PR'3=PEt3 or 1/2Et2PCH2CH2PEt2) (depe)] and PR2H (R=Ph, a; tBu, b; Cy, c) in C6H5F, or of related cationic Ru(N2) complexes with PR2H in C6H5F, gave the secondary phosphine complexes [(eta5-C5H5)Ru(PR'3)2(PR2H)]+ BAr(F)- (PR'3=PEt3, 3 a-c; 1/2depe, 4 a,b) in 65-91 % yields. Additions of tBuOK (3 a, 4 a; [D6]acetone) or NaN(SiMe3)2 (3 b,c, 4 b; [D8]THF) gave the title complexes [(eta5-C5H5)Ru(PEt3)2(PR2)] (5 a-c) and [(eta5-C5H5)Ru(depe)(PR2)] (6 a,b) in high spectroscopic yields. These complexes were rapidly oxidized in air; with 5 a, [(eta5-C5H5)Ru(PEt3)2{P(=O)Ph2}] was isolated (>99 %). The reaction of 5 a and elemental selenium yielded [(eta5-C5H5)Ru(PEt3)2{P(=Se)Ph2}] (70 %); selenides from 5 c and 6 a were characterized in situ. Competitive deprotonation reactions showed that 5 a is more basic than the rhenium analog [(eta5 C5H5)Re(NO)(PPh3)(PPh2)], and that 6 b is more basic than PtBu3 and P(iPrNCH2CH2)3N. The latter is one of the most basic trivalent phosphorus compounds [pK(a)(acetonitrile) 33.6]. Complexes 5 a-c and 6 b are effective ligands for Pd(OAc)2-catalyzed Suzuki coupling reactions: 6 b gave a catalyst nearly as active as the benchmark organophosphine PtBu3; 5 a, with a less bulky and electron-rich PR2 moiety, gave a less active catalyst. The reaction of 5 a and [(eta3-C3H5)Pd(NCPh)2]+ BF4- gave the bridging phosphido complex [(eta5 C5H5)Ru(PEt3)2(PPh2)Pd(NCPh)(eta3-C3H5)]+ BAr(F)- in approximately 90 % purity. The crystal structure of 4 a is described, as well as substitution reactions of 3 b and 4 b. PMID- 15651023 TI - Electrochemical antitumor drug sensitivity test for leukemia K562 cells at a carbon-nanotube-modified electrode. AB - The change in electrochemical behavior of tumor cells induced by antitumor drugs was detected by using a multiwall carbon nanotubes (MWNTs)-modified glass carbon electrode (GCE). Based on the changes observed, a simple, in vitro, electrochemical antitumor drug sensitivity test was developed. MWNTs promoted electron transfer between the electroactive centers of cells and the electrode. Leukemia K562 cells exhibited a well-defined anodic peak of guanine at +0.823 V at 50 mV s(-1). HPLC assay with ultraviolet detection was used to elucidate the reactant responsible for the electrochemical response of the tumor cells. The guanine content within the cytoplasm of each K562 cell was detected to be 920 amol. For the drug sensitivity tests, 5-fluorouracil (5-FU) and several clinical antitumor drugs, such as vincristine, adriamycin, and mitomycin C, were added to cell culture medium. As a result, the electrochemical responses of the K562 cells decreased significantly. The cytotoxicity curves and results obtained corresponded well with the results of MTT assays. In comparison to conventional methods, this electrochemical test is highly sensitive, accurate, inexpensive, and simple. The method proposed could be developed as a convenient means to study the sensitivity of tumor cells to antitumor drugs. PMID- 15651024 TI - Steric control over hydrogen bonding in crystalline organic solids: a structural study of N,N'-dialkylthioureas. AB - Hydrogen bonding in crystalline N,N'-dialkylthioureas was examined with the help of single-crystal X-ray diffraction, DFT calculations, and Cambridge Structural Database (CSD) analysis. A CSD survey indicated that unlike the related urea derivatives, which persistently self-assemble into one-dimensional hydrogen bonded chains, the analogous thioureas can form two different hydrogen-bonding motifs in the solid state: chains, structurally similar with those found in ureas, and dimers, that further associate into hydrogen-bonded layers. The formation of one motif or another can be manipulated by the bulkiness of the organic substituents on the thiourea group, which provides a clear example of steric control over the hydrogen bonding arrangement in crystalline organic solids. PMID- 15651025 TI - Intramolecular, reductive cyclization of beta-ketoisothiocyanates promoted by using samarium diiodide. AB - A novel samarium diiodide (SmI2) promoted intramolecular cyclization of beta ketoisothiocyanate, derived from alpha,beta-unsaturated esters and ammonium thiocyanate led to alpha-hydroxythiolactams and/or thiolactams in high yields. Treatment of beta-ketoisothiocyanate with two equivalents of SmI2 gave a mixture of alpha-hydroxythiolactam and thiolactam. Four equivalents of SmI2 afforded only thiolactam in high yields. The intramolecular cyclization took place with high to complete stereoselectivity. A mechanism to explain this transformation is proposed. PMID- 15651026 TI - The key role of atom types, reference states, and interaction cutoff radii in the knowledge-based method: new variational approach. AB - We present a variational method to derive knowledge-based potentials. The method is based on an optimization procedure of objective variables: atom types, reference states, and interaction cutoff radii. We suggest and apply new unsymmetrical reference states. The cutoff radii and atom types are optimized to improve docking accuracy of the corresponding potentials. The atom types are varied along an atom type tree, with 6 root and 49 top atom types, and the set of 18 optimal atom types is obtained. We demonstrate strong dependence between the choice of atom types and the docking accuracy of the potentials derived with these atom types. The averaged root-mean square deviations (RMSDs) of the ligand docked positions relative to the experimentally determined positions decrease when the elements C, N, O are split into the optimal types. PMID- 15651027 TI - Crystal structure of a formiminotetrahydrofolate cyclodeaminase (TM1560) from Thermotoga maritima at 2.80 A resolution reveals a new fold. PMID- 15651028 TI - Modulation of CXCL14 (BRAK) expression in prostate cancer. AB - BACKGROUND: Recent studies suggest inflammatory processes may be involved in the development or progression of prostate cancer. Chemokines are a family of cytokines that can play several roles in cancer progression including angiogenesis, inflammation, cell recruitment, and migration. METHODS: Real-time quantitative RT-PCR, in situ RNA hybridization, laser capture microscopy, immunohistochemistry, and cDNA array based technologies were used to examine CXCL14 (BRAK) expression in paired normal and tumor prostate. To determine the role CXCL14 expression has on cancer progression, LAPC4 cells were engineered to overexpress mouse or human CXCL14, and xenograft studies were performed. RESULTS: CXCL14 RNA expression was observed in normal and tumor prostate epithelium and focally in stromal cells adjacent to cancer. CXCL14 mRNA was significantly upregulated in localized prostate cancer and positively correlated with Gleason score. CXCL14 levels were unchanged in BPH specimens. LAPC4 cells expressing CXCL14 resulted in a 43% tumor growth inhibition (P = 0.019) in vivo compared to vector only xenografts. CONCLUSIONS: CXCL14 mRNA upregulation is a common feature in prostate cancer. The finding that CXCL14 expression inhibits tumor growth suggests this gene has tumor suppressive functions. PMID- 15651029 TI - Bi-directional interactions of prostate cancer cells and bone marrow endothelial cells in three-dimensional culture. AB - BACKGROUND: Prostate cancer preferentially metastasizes to bone, yet little is known about the cellular and molecular factors that support this growth. Endothelial cells are likely the initial contact for circulating prostate cells entering the bone microenvironment. METHODS: Using co-culture and conditioned media experiments, we studied cellular and molecular interactions of prostate cancer cells of varying aggressiveness (PC-3 and LNCaP) with bone marrow endothelial (HBME-1) cells in collagen gels. RESULTS: In co-culture, HBME-1 cells stimulated proliferation ( approximately 90% increase) and migration of the more aggressive PC-3 cell line, while having little effect on LNCaP cell proliferation or migration. Concomitantly, HBME-1 cell growth was inhibited by both PC-3 and LNCaP cells and their conditioned media. Additionally, HBME-1 cells underwent significant morphological changes in co-culture, forming large, branching, cord like structures, which mimic angiogenesis. Prostate cancer cell conditioned media induced a similar effect on HBME-1 cells. In comparison, conditioned media from PC-3 cells also inhibited growth of non-bone marrow-derived endothelial cells, but did not affect their morphology. CONCLUSIONS: Significant bi-directional interactions, including secreted factors and direct cellular interactions, exist between bone marrow endothelial cells and highly metastatic prostate cancer cells, and may underlie the propensity for prostate cancer to metastasize to the bone. PMID- 15651030 TI - Guanidinoacetate methyltransferase deficiency identified in adults and a child with mental retardation. AB - Our study describes the adult clinical and biochemical spectrum of guanidinoacetate methyltransferase (GAMT) deficiency, a recently discovered inborn error of metabolism. The majority of the previous reports dealt with pediatric patients, in contrast to the present study. A total of 180 institutionalized patients with a severe mental handicap were investigated for urine and plasma uric acid and creatinine. Patients with an increased urinary uric acid/creatinine ratio and/or decreased creatinine were subjected to the analysis of guanidinoacetate (GAA). Four patients (three related and one from an unrelated family) were identified with GAMT-deficiency. A fifth patient had died before a biochemical diagnosis could be made. They all had shown a normal psychomotor development for the first year of life, after which they developed a profound mental retardation. Three out of four had convulsions and all four totally lacked the development of speech. Their GAMT activity in lymphoblasts was impaired and two novel mutations were identified: the 59 G > C and 506 G > A missense mutations. Urinary GAA was increased, but highly variable 347-1,624 mmol/mol creat (Controls <150 mmol/mol creat). In plasma and CSF the GAA levels were fairly constant at 17.3-27.0 mumol/L (Controls 1.33-3.33) and 11.0-12.4 mumol/L, respectively (Controls 0.068-0.114). GAMT deficiency in adults is associated with severe mental retardation and absence or limited speech development. Convulsions may be prominent. The nonspecific nature of the clinical findings as well as the limited availability of GAA assays and/or in vivo magnetic resonance spectroscopy of the brain may mean that many more patients remain undiagnosed in institutions for persons with mental handicaps. PMID- 15651031 TI - Vibrational computations beyond the harmonic approximation: performances of the B3LYP density functional for semirigid molecules. AB - The performances of the B3LYP density functional in the computation of harmonic and anharmonic frequencies were tested using 14 standard basis sets of double and triple zeta quality for a set of semirigid molecules containing from 4 to 12 atoms. The quality of the results is assessed by comparison with the most reliable computations available in the literature. The study reveals that the relatively cheap 6-31+G(d,p) basis set performs a very good job for harmonic frequency calculations and that B3LYP anharmonicities are in close agreement with the reference values irrespective of the basis set used. On these grounds "hybrid force fields" are proposed to achieve the best compromise between computer time and quality of the results. PMID- 15651032 TI - Continuous medium theory for nonequilibrium solvation: III. Solvation shift by monopole approximation and multipole expansion in spherical cavity. AB - According to the classical electrodynamics, a new and reasonable method about electrostatic energy decomposition of the solute-solvent system has been proposed in this work by introducing the concept of spring energy. This decomposition in equilibrium solvation gives the clear comprehension for different parts of total electrostatic free energy. Logically extending this cognition to nonequilibrium leads to the new formula of electrostatic free energy of nonequilibrium state. Furthermore, the general solvation shift for light absorption/emission has been reformulated and applied to the ideal sphere case with the monopole approximation and multipole expansion. Solvation shifts in vertical ionizations of atomic ions of some series of main group elements have been investigated with monopole approximation, and the variation tendency of the solvation shift versus atomic number has been discussed. Moreover, the solvation shift in photoionization of nitrate anion in glycol has been investigated by the multipole expansion method. PMID- 15651033 TI - A scoring function for docking ligands to low-resolution protein structures. AB - We present a docking method that uses a scoring function for protein-ligand docking that is designed to maximize the docking success rate for low-resolution protein structures. We find that the resulting scoring function parameters are very different depending on whether they were optimized for high- or low resolution protein structures. We show that this docking method can be successfully applied to predict the ligand-binding site of low-resolution structures. For a set of 25 protein-ligand complexes, in 76% of the cases, more than 50% of ligand-contacting residues are correctly predicted (using receptor crystal structures where the binding site is unspecified). Using decoys of the receptor structures having a 4 A RMSD from the native structure, for the same set of complexes, in 72% of the cases, we obtain at least one correctly predicted ligand-contacting residue. Furthermore, using an 81-protein-ligand set described by Jain, in 76 (93.8%) cases, the algorithm correctly predicts more than 50% of the ligand-contacting residues when native protein structures are used. Using 3 A RMSD from native decoys, in all but two cases (97.5%), the algorithm predicts at least one ligand-binding residue correctly. Finally, compared to the previously published Dolores method, for 298 protein-ligand pairs, the number of cases in which at least half of the specific contacts are correctly predicted is more than four times greater. PMID- 15651034 TI - Quantum vs. classical models of the nitro group for proton chemical shift calculations and conformational analysis. AB - A model based on classical concepts is derived to describe the effect of the nitro group on proton chemical shifts. The calculated chemical shifts are then compared to ab initio (GIAO) calculated chemical shifts. The accuracy of the two models is assessed using proton chemical shifts of a set of rigid organic nitro compounds that are fully assigned in CDCl3 at 700 MHz. The two methods are then used to evaluate the accuracy of different popular post-SCF methods (B3LYP and MP2) and molecular mechanics methods (MMX and MMFF94) in calculating the molecular structure of a set of sterically crowded nitro aromatic compounds. Both models perform well on the rigid molecules used as a test set, although when using the GIAO method a general overestimation of the deshielding of protons near the nitro group is observed. The analysis of the sterically crowded molecules shows that the very popular B3LYP/6-31G(d,p) method produces very poor twist angles for these, and that using a larger basis set [6-311++G(2d,p)] gives much more reasonable results. The MP2 calculations, on the other hand, overestimate the twist angles, which for these compounds compensates for the deshielding effect generally observed for protons near electronegative atoms when using the GIAO method at the B3LYP/6-311++G(2d,p) level. The most accurate results are found when the structures are calculated using B3LYP/6-311++G(2d,p) level of theory, and the chemical shifts are calculated using the CHARGE program based on classical models. PMID- 15651035 TI - Role of trehalose and heat in the structure of the C-terminal activation domain of the heat shock transcription factor. AB - The heat shock transcription factor (HSF) is the primary transcriptional regulator of the heat shock response in eukaryotes. Saccharomyces cerevisiae HSF1 has two functional transcriptional activation domains, located N- and C-terminal to the central core of the protein. These activation domains have a low level of transcriptional activity prior to stress, but they acquire a high level of transcriptional activity in response to stresses such as heat. Previous studies on the N-terminal activation domain have shown that it can be completely disordered. In contrast, we show that the C-terminal activation domain of S. cerevisiae HSF1 does contain a certain amount of secondary structure as measured by circular dichroism (CD) and protease resistance. The alpha-helical content of the domain can be increased by the addition of the disaccharide trehalose but not by sucrose. Trehalose, but not sucrose, causes a blue shift in the fluorescence emission spectra, which is suggestive of an increase in tertiary structure. Trehalose, which is known to be a chemical chaperone, also increases proteases' resistance and promotes heat-induced increases in alpha-helicity. The latter is particularly intriguing because of the physiological role of trehalose in yeast. Trehalose levels are increased dramatically after heat shock, and this is thought to protect protein structure prior to the increase of heat shock protein levels. Our results suggest that the dramatic changes in S. cerevisiae HSF1 transcriptional activity in response to stress might be linked to the combined effects of trehalose and elevated temperatures in modifying the overall structure of HSF1's C-terminal activation domain. PMID- 15651036 TI - Prediction of binding modes for ligands in the cytochromes P450 and other heme containing proteins. AB - The cytochromes P450 (P450s) are a family of heme-containing monooxygenase enzymes involved in a variety of functions, including the metabolism of endogenous and exogenous substances in the human body. During lead optimization, and in drug development, many potential drug candidates are rejected because of the affinity they display for drug-metabolising P450s. Recently, crystal structures of human enzymes involved in drug metabolism have been determined, significantly augmenting the prospect of using structure-based design to modulate the binding and metabolizing properties of compounds against P450 proteins. An important step in the application of structure-based metabolic optimization is the accurate prediction of docking modes in heme binding proteins. In this paper we assess the performance of the docking program GOLD at predicting the binding mode of 45 heme-containing complexes. We achieved success rates of 64% and 57% for Chemscore and Goldscore respectively; these success rates are significantly lower than the value of 79% observed with both scoring functions for the full GOLD validation set. Re-parameterization of metal-acceptor interactions and lipophilicity of planar nitrogen atoms in the scoring functions resulted in a significant increase in the percentage of successful dockings against the heme binding proteins (Chemscore 73%, Goldscore 65%). The modified scoring functions will be useful in docking applications on P450 enzymes and other heme binding proteins. PMID- 15651038 TI - From RNAi to epigenomes: how RNA rules the world. PMID- 15651037 TI - High dose fractionated ionizing radiation inhibits prostate cancer cell adhesion and beta(1) integrin expression. AB - BACKGROUND: The effect of ionizing radiation on extracellular matrix (ECM) mediated cellular functions is an important area of research for translational science. Mechanisms of tumor cell ability to proliferate, migrate, and survive appear dependent on integrin-mediated adhesion to the ECM; however, the exact role therapeutic radiation plays in altering signaling pathways and promoting cell death within remains less well established. METHODS: To examine these effects on prostate carcinoma cell lines, cells were irradiated at sub-lethal doses. We have studied two human prostate cancer cell lines (PC3 and DU-145) irradiated with different fractionated radiation schedules. Three groups were compared to non-irradiated controls. Group A was given a single dose of 5 Gy. Group B was given 5 Gy the first week and then 10 Gy the second week for a total of 15 Gy. Group C was given 5 Gy the first week, and then 10 Gy the second and third week for a total of 25 Gy. Cells were analyzed at their prescribed total dose. At 48 hr post irradiation, cells were detached from culture dishes and were subsequently used for adhesion assays and immunoblotting analysis. RESULTS: Our findings revealed that two prostate carcinoma cell lines, PC3 and DU-145, had a reduced cellular adhesion to fibronectin (FN) compared to the non-irradiated control groups. Both prostate cancer cell lines showed decreased adhesion to FN and reduced beta(1) integrin protein levels at a total dose of 25 Gy, but not at the doses of 15 and 5 Gy. In a parallel analysis, at the maximum total dose of 25 Gy, both PC3 and DU-145 demonstrated a significant decrease in cell proliferation. CONCLUSIONS: High dose radiation treatment of prostate cancer cell lines inhibits integrin expression. Our study suggests that promoting a synergistic decrease in adhesion could bring additional therapeutic benefit to patients treated with radiation therapy. PMID- 15651039 TI - Viral vectors as tools to model and treat neurodegenerative disorders. AB - The identification of disease-causing genes in familial forms of neurodegenerative disorders and the development of genetic models closely replicating human central nervous system (CNS) pathologies have drastically changed our understanding of the molecular events leading to neuronal cell death. If these achievements open new opportunities of therapeutic interventions, including gene-based therapies, the presence of the blood-brain barrier and the post-mitotic and poor regenerative nature of the target cells constitute important challenges. Efficient delivery systems taking into account the specificity of the CNS are required to administer potential therapeutic candidates. In addition, genetic models in large animals that replicate the late stages of the diseases are in most cases not available for pre-clinical studies. The present review summarizes the potential of viral vectors as tools to create new genetic models of CNS disorders in various species including primates and the recent progress toward viral gene therapy clinical trials for the administration of therapeutic candidates into the brain. PMID- 15651040 TI - Structure and biosynthesis of myxochromides S1-3 in Stigmatella aurantiaca: evidence for an iterative bacterial type I polyketide synthase and for module skipping in nonribosomal peptide biosynthesis. AB - The myxobacterium Stigmatella aurantiaca DW4/3-1 harbours an astonishing variety of secondary metabolic gene clusters, at least two of which were found by gene inactivation experiments to be connected to the biosynthesis of previously unknown metabolites. In this study, we elucidate the structures of myxochromides S1-3, novel cyclic pentapeptide natural products possessing unsaturated polyketide side chains, and identify the corresponding biosynthetic gene locus, made up of six nonribosomal peptide synthetase modules. By analyzing the deduced substrate specificities of the adenylation domains, it is shown that module 4 is most probably skipped during the biosynthetic process. The polyketide synthase MchA harbours only one module and is presumably responsible for the formation of the variable complete polyketide side chains. These data indicate that MchA is responsible for an unusual iterative polyketide chain assembly. PMID- 15651041 TI - The biosynthesis of vancomycin-type glycopeptide antibiotics--a model for oxidative side-chain cross-linking by oxygenases coupled to the action of peptide synthetases. PMID- 15651042 TI - The functional role of selenocysteine (Sec) in the catalysis mechanism of large thioredoxin reductases: proposition of a swapping catalytic triad including a Sec His-Glu state. AB - Thioredoxin reductases catalyse the reduction of thioredoxin disulfide and some other oxidised cell constituents. They are homodimeric proteins containing one FAD and accepting one NADPH per subunit as essential cofactors. Some of these reductases contain a selenocysteine at the C terminus. Based on the X-ray structure of rat thioredoxin reductase, homology models of human thioredoxin reductase were created and subsequently docked to thioredoxin to model the active complex. The formation of a new type of a catalytic triad between selenocysteine, histidine and a glutamate could be detected in the protein structure. By means of DFT (B3LYP, lacv3p**) calculations, we could show that the formation of such a triad is essential to support the proton transfer from selenol to a histidine to stabilise a selenolate anion, which is able to interact with the disulfide of thioredoxin and catalyses the reductive disulfide opening. Whereas a simple proton transfer from selenocysteine to histidine is thermodynamically disfavoured by some 18 kcal mol(-1), it becomes favoured when the carboxylic acid group of a glutamate stabilises the formed imidazole cation. An identical process with a cysteine instead of selenocysteine will require 4 kcal mol(-1) more energy, which corresponds to a calculated equilibrium shift of approximately 1000:1 or a 10(3) rate acceleration: a value close to the experimental one of about 10(2) times. These results give new insights into the catalytic mechanism of thioredoxin reductase and, for the first time, explain the advantage of the incorporation of a selenocysteine instead of a cysteine residue in a protein. PMID- 15651043 TI - Kinetics study of Bungarus fasciatus venom acetylcholinesterase immobilised on a Langmuir-Blodgett proteo-glycolipidic bilayer. AB - This study deals with the kinetics properties of an enzyme immobilised in a defined orientation in a biomimetic environment. For this purpose, acetylcholinesterase (AChE) was captured at the surface of a nanostructured proteo-glycolipidic Langmuir-Blodgett film through specific recognition by a noninhibitor monoclonal antibody (IgG) inserted in a neoglycolipid bilayer. Modelling of this molecular assembly provided a plausible interpretation of the functional orientation of the enzyme. The AChE activity being stable for several weeks, the enzyme kinetics were investigated, and fitted perfectly with heterogeneous biocatalytic behaviour representative of cellular enzymatic catalysis. The AChE-IgG-glycolipid nanostructure was directly interfaced with an efficient optical device. Such an association, leading to an intimate contact between the nanostructure and the biochemical signal transducer, gives direct access to the intrinsic AChE behaviour. This study thus demonstrates the potential for direct investigation of the kinetic behaviour of an immobilised enzyme on a lipid bilayer through an efficient transduction system. PMID- 15651044 TI - Small-molecule inhibitors and probes for ubiquitin- and ubiquitin-like-specific proteases. PMID- 15651045 TI - A new type of metalloprotein: The Mo storage protein from azotobacter vinelandii contains a polynuclear molybdenum-oxide cluster. AB - Azotobacter vinelandii is a diazotrophic bacterium characterized by the outstanding capability of storing Mo in a special storage protein, which guarantees Mo-dependent nitrogen fixation even under growth conditions of extreme Mo starvation. The Mo storage protein is constitutively synthesized with respect to the nitrogen source and is regulated by molybdenum at an extremely low concentration level (0-50 nM). This protein was isolated as an alpha4beta4 octamer with a total molecular mass of about 240 kg mol(-1) and its shape was determined by small-angle X-ray scattering. The genes of the alpha and beta subunits were unequivocally identified; the amino acid sequences thereby determined reveal that the Mo storage protein is not related to any other known molybdoprotein. Each protein molecule can store at least 90 Mo atoms. Extended X ray absorption fine-structure spectroscopy identified a metal-oxygen cluster bound to the Mo storage protein. The binding of Mo (biosynthesis and incorporation of the cluster) is dependent on adenosine triphosphate (ATP); Mo release is ATP-independent but pH-regulated, occurring only above pH 7.1. This Mo storage protein is the only known noniron metal storage system in the biosphere containing a metal-oxygen cluster. PMID- 15651046 TI - Transport of surface-modified nanoparticles through cell monolayers. AB - We synthesized three peptides, a D-polyarginyl peptide (r8(FITC)), a Tat peptide (Tat(FITC)), and a control peptide (Cp(FITC)) and attached each to amino-CLIO, a nanoparticle 30 nm in diameter. We then examined the effective permeability, Peff, of all six materials through CaCo-2 monolayers. The transport of peptide nanoparticles was characterized by a lag phase (0-8 h) and a steady-state phase (9-27 h). The steady-state Peff values for peptides were in the order r8(FITC)>Tat(FITC)=Cp(FITC). When r8(FITC) and Tat(FITC) peptides were attached to the nanoparticle, they conferred their propensity to traverse cell monolayers onto the nanoparticle, whereas Cp(FITC) did not. Thus, when the r8(FITC) peptide was attached to the amino-CLIO nanoparticle, the resulting peptide-nanoparticle had a Peff similar to that of this poly-D-arginyl peptide alone. The Peff of r8(FITC)-CLIO (MW approximately 1000 kDa) was similar to that of mannitol (MW=182 Da), a poorly transported reference substance, with a far lower molecular weight. These results are the first to indicate that the modification of nanoparticles by attachment of membrane-translocating sequence-based peptides can alter nanoparticle transport through monolayers. This suggests that the surface modification of nanoparticles might be a general strategy for enhancing the permeability of drugs and that high-permeability nanoparticle-based therapeutics can be useful in selected pharmaceutical applications. PMID- 15651047 TI - Induction of DNA-double-strand breaks by auger electrons from 99mTc complexes with DNA-binding ligands. AB - The potential of certain Auger electron emitting nuclides for systemic radiotherapeutic applications has recently gained much attention. In particular, the ability of several nuclides, including 111In, 125I, and 123I, to induce DNA double-strand breaks (dsb), a good indicator of cytotoxicity, has been extensively studied. However, this ability has never previously been shown experimentally for 99mTc, which, besides the well-known gamma radiation that is used for diagnostic applications, also emits an average of 1.1 conversion electrons and 4 Auger or Coster-Kronig electrons per decay. Owing to the short range of Auger electrons, the radionuclide needs to be located very close to the DNA for dsb to occur. We synthesized two cationic 99mTcI-tricarbonyl complexes with pendant DNA binders, pyrene and anthraquinone. The X-ray crystal structures of the two complexes could be elucidated. Linear dichroism and UV/Vis spectroscopy revealed that the complex with pyrene intercalates DNA with a stability constant, K, of 1.1 x 10(6) M(-1), while the analogous complex with anthraquinone interacts with DNA in a groove-binding mode and has an affinity value of K=8.9 x 10(4) M(-1). We showed with phiX174 double-stranded DNA that the corresponding 99mTc complexes induce a significant amount of dsb, whereas non-DNA binding [TcO4]- and nonradioactive Re compounds did not. These results indicate that the Auger electron emitter 99mTc can induce dsb in DNA when decaying in its direct vicinity and this implies potential for systemic radiotherapy with 99mTc complexes. PMID- 15651048 TI - Oligosaccharide mimics containing galactose and fucose specifically label tumour cell surfaces and inhibit cell adhesion to fibronectin. AB - With the aim of establishing a versatile and easy synthesis of branched saccharides for biological applications, we used molecular-dynamics simulations to model Lewis(y) to two classes of di- or triantennary saccharide mimetics. One set of mimetics was based on 1,3,5-tris(hydroxymethyl)cyclohexane (TMC) as the core, the other on furan, and both were derivatised with galactose and/or fucose. The TMC-based saccharides were biotinylated, while the furan disaccharides were treated with maleimide-activated biotin in a Diels-Alder fashion to yield oxazatricyclodecanes (OTDs). These were then assayed as cell-surface labels in human colon (SW480 and CaCo-2), liver (PLC), Glia (U333 CG 343) and ovary (SKOV 3) tumour cell lines. Discrete staining patterns were observed in all cells, usually at one or two poles of the cells, particularly with the asymmetric 3-beta L-fucopyranosyloxymethyl-4-beta-D-galactopyranosyloxymethyl-OTD. Normal SV40 transformed fibroblasts (SV80) showed no staining. Adhesion of the highly metastatic mouse melanoma line B16 F10 to fibronectin was inhibited by 80 % by the TMC-digalactoside and by 30 % by 3,4-bis-(beta-D galactopyranosyloxymethyl)furan. None of the saccharide mimetics inhibited the adhesion of the less metastatic B16 F1 line. Migration of B16 F10 cells through Matrigel was greatly inhibited by the TMC-digalactoside and weakly inhibited by the TMC-trigalactoside. The saccharide mimetics that had shown the best structural agreements with the terminal saccharides of Lewis(y) in the molecular dynamics simulation were also the most biologically potent compounds; this underlines the predictive nature of molecular dynamics simulations. The use of the non-saccharide cores enabled us to adapt spacer lengths and terminal saccharides to optimise the structures to bind more avidly to cell-surface lectins. PMID- 15651050 TI - A "solvated rotamer" approach to modeling water-mediated hydrogen bonds at protein-protein interfaces. AB - Water-mediated hydrogen bonds play critical roles at protein-protein and protein nucleic acid interfaces, and the interactions formed by discrete water molecules cannot be captured using continuum solvent models. We describe a simple model for the energetics of water-mediated hydrogen bonds, and show that, together with knowledge of the positions of buried water molecules observed in X-ray crystal structures, the model improves the prediction of free-energy changes upon mutation at protein-protein interfaces, and the recovery of native amino acid sequences in protein interface design calculations. We then describe a "solvated rotamer" approach to efficiently predict the positions of water molecules, at protein-protein interfaces and in monomeric proteins, that is compatible with widely used rotamer-based side-chain packing and protein design algorithms. Finally, we examine the extent to which the predicted water molecules can be used to improve prediction of amino acid identities and protein-protein interface stability, and discuss avenues for overcoming current limitations of the approach. PMID- 15651049 TI - 1.70 A X-ray structure of human apo kallikrein 1: structural changes upon peptide inhibitor/substrate binding. AB - Human kallikreins are serine proteases that comprise a recently identified large and closely related 15-member family. The kallikreins include both regulatory- and degradative-type proteases, impacting a variety of physiological processes including regulation of blood pressure, neuronal health, and the inflammatory response. While the function of the majority of the kallikreins remains to be elucidated, two members are useful biomarkers for prostate cancer and several others are potentially useful biomarkers for breast cancer, Alzheimer's, and Parkinson's disease. Human tissue kallikrein (human K1) is the best functionally characterized member of this family, and is known to play an important role in blood pressure regulation. As part of this function, human K1 exhibits unique dual-substrate specificity in hydrolyzing low molecular weight kininogen between both Arg-Ser and Met-Lys sequences. We report the X-ray crystal structure of mature, active recombinant human apo K1 at 1.70 A resolution. The active site exhibits structural features intermediate between that of apo and pro forms of known kallikrein structures. The S2 to S2' pockets demonstrate a variety of conformational changes in comparison to the porcine homolog of K1 in complex with peptide inhibitors, including the displacement of an extensive solvent network. These results indicate that the binding of a peptide substrate contributes to a structural rearrangement of the active-site Ser 195 resulting in a catalytically competent juxtaposition with the active-site His 57. The solvent networks within the S1 and S1' pockets suggest how the Arg-Ser and Met-Lys dual substrate specificity of human K1 is accommodated. PMID- 15651051 TI - Lithium and valproate attenuate dextroamphetamine-induced changes in brain activation. AB - BACKGROUND: Previous studies have suggested that both lithium and valproate may decrease phosphoinositol second messenger system (PI-cycle) activity. There is also evidence that dextroamphetamine may increase PI cycle activity. It was previously demonstrated that dextroamphetamine administration in volunteers causes a region and task dependent decrease in brain activation in healthy volunteers. The current study assessed the effect of 14 days pretreatment with lithium and valproate on these dextroamphetamine-induced changes in regional brain activity in healthy volunteers. METHODS: This was a double-blind, placebo controlled, study in which volunteers received either 1000 mg sodium valproate (n = 12), 900 mg lithium (n = 9) or placebo (n = 12). Functional images were acquired using functional magnetic resonance imaging (fMRI) while subjects performed three cognitive tasks, a word generation paradigm, a spatial attention task and a working memory task. fMRI was carried out both before and after administration of dextroamphetamine (25 mg). Changes in the number of activated pixels and changes in the magnitude of the blood-oxygen-level-dependent (BOLD) signal after dextroamphetamine administration were then determined. RESULTS: In keeping with previous findings dextroamphetamine administration decreased regional brain activation in all three tasks. Pretreatment with lithium attenuated changes in the word generation paradigm and the spatial attention task, while pretreatment with valproate attenuated the changes in the working memory task. CONCLUSIONS: These results suggest that both lithium and valproate can significantly attenuate dextroamphetamine-induced changes in brain activity in a task dependent and region specific manner. This is the first human evidence to suggest that both lithium and valproate may have a similar effect on regional brain activation, conceivably via similar effects on PI-cycle activity. PMID- 15651052 TI - The use of atypical antipsychotics in the treatment of schizophrenia in North Staffordshire. AB - AIMS: To examine the long-term effectiveness of atypical antipsychotics in a naturalistic setting for patients with schizophrenia. METHOD: A retrospective analysis of atypical antipsychotic prescribing in one Health District between 1994 and 2001. Time to discontinuation of the first atypical antipsychotic prescribed was calculated using survival analysis. RESULTS: 253 patients were identified. Clozapine had a significantly lower discontinuation rate compared with olanzapine and risperidone (p = 0.018). Patients taking risperidone were 1.3 times more likely to discontinue than those taking olanzapine (p = 0.23). Older age (p = 0.0001), male sex (p = 0.016) and exposure to antidepressants (p = 0.014) significantly predicted compliance. CONCLUSIONS: Clozapine is an effective long-term schizophrenia treatment. The trend to superior effectiveness of olanzapine over risperidone in the long-term has not been reported before and warrants further investigation. PMID- 15651053 TI - A systematic review of guided imagery as an adjuvant cancer therapy. AB - AIM: The aim of this paper is to summarise and critically evaluate the evidence available from controlled clinical trials regarding the use of guided imagery as a sole adjuvant therapy for cancer patients. METHODS: Electronic searches for controlled clinical trials were carried out in eight databases and two clinical trial registers. Trials that featured guided imagery as a sole adjuvant therapy were included. No language restrictions were imposed. Data were extracted and validated independently by two researchers. RESULTS: Six randomised clinical trials were included. Detailed results were available for four studies only. Poor reporting and heterogeneous populations, interventions and outcome measures across trials precluded statistical pooling of results. The methodological quality was on average low. Three studies reported significant differences in measures of anxiety, comfort or emotional response to chemotherapy for patients who received guided imagery over the control groups. Two studies showed no differences between guided imagery and other interventions in any of the outcome measures. CONCLUSION: Guided imagery, as a sole adjuvant cancer therapy may be psycho-supportive and increase comfort. There is no compelling evidence to suggest positive effects on physical symptoms such as nausea and vomiting. The data seem sufficiently encouraging for the use of guided imagery as an adjuvant cancer therapy to merit further research. PMID- 15651054 TI - Effects of prolonged wakefulness combined with alcohol and hands-free cell phone divided attention tasks on simulated driving. AB - Simulated driving ability was assessed following administration of alcohol, at an estimated blood level of 0.05%, and combined prolonged wakefulness, while participants were undertaking divided attention tasks over a hands-free mobile phone. Divided attention tasks were structured to provide a sustained cognitive workload to the subjects. Twenty three young healthy individuals drove 10 km simulated driving under four conditions in a counterbalanced, within-subject design: alcohol, alcohol and 19 h wakefulness, alcohol and 24 h wakefulness, and while sober. Study measures were: simulated driving, self-reported sleepiness, critical flicker fusion threshold (CFFT), Stroop word-colour interference test (Stroop) and simple visual reaction times (SVRT). As expected, subjective sleepiness was highly correlated with both sleep restriction and alcohol consumption. The combination of alcohol and 24 h sustained wakefulness produced the highest driving impairment, significantly beyond the alcohol effect itself. Concurrent alcohol and 19 h wakefulness significantly affected only driving time to-collision. No significant changes of study measures occurred following alcohol intake in unrestricted sleep conditions. CFFT, SVRT and Stroop results showed a similar trend in the four study conditions. Thus apparently 'safe' blood alcohol levels in combination with prolonged wakefulness resulted in significant driving impairments. In normal sleep conditions alcohol effects on driving were partially counteracted by the concomitant hands-free phone based psychometric tasks. PMID- 15651055 TI - Taking CHARGE: A self-management program for women following breast cancer treatment. AB - The purpose of the study was to develop and evaluate Taking CHARGE, a self management intervention designed to facilitate successful transitions to survivorship after breast cancer treatment. The Taking CHARGE intervention involves a two-pronged approach building on self-regulation principles to (1) equip women with self-management skills to address concerns following breast cancer treatment, and (2) provide information about common survivorship topics. The program involved four intervention contacts, two small group meetings and two individualized telephone sessions, delivered by nurse/health educators. This paper focuses on the process evaluation findings from a preliminary test of the Taking CHARGE intervention conducted with 25 women, aged 34-66 years, completing breast cancer treatment, who were randomly assigned to the intervention group. The process evaluation was conducted to obtain systematic information about the relevance and usefulness of the self-regulation approach, informational aspects, and program delivery. The findings indicated that intervention group participants found the Taking CHARGE program to be timely, relevant, and to have high utility in dealing with concerns that exist following breast cancer treatment. The process evaluation findings provide early evidence of the usefulness of the Taking CHARGE intervention for successful transition to survivorship following breast cancer treatment. PMID- 15651056 TI - Safety and efficacy of adeno-associated viral vector-mediated insulin gene transfer via portal vein to the livers of streptozotocin-induced diabetic Sprague Dawley rats. AB - BACKGROUND: Previous studies demonstrating the efficacy of insulin gene therapy have mostly involved use of adenoviral vectors or naked DNA to deliver the insulin gene. However, this procedure may not guarantee long-term insulin production. To improve the performance, we prepared recombinant adeno-associated viral vectors (rAAV) harboring the gene encoding a furin-modified human insulin under the cytomegalovirus (CMV) promoter [rAAV-hPPI(F12)]. METHODS: Streptozotocin (STZ)-induced diabetic Sprague-Dawley rats were used as a diabetic animal model. The levels of blood glucose, insulin, and HbA1c were measured to test the effect. An intraperitoneal glucose tolerance test was performed to test the capability of blood glucose disposal. Immunohistochemical staining and Northern blot analyses were performed to survey the expression pattern of the therapeutic insulin gene. RESULTS: STZ-induced diabetic Sprague-Dawley rats infused via the portal vein with rAAV-hPPI(F12) produced human insulin and after a 6-h fast were normoglycemic for over 90 days post-treatment, whereas diabetic rats treated with recombinant adenoviral vector harboring the hPPI(F12) gene [rAV hPPI(F12)] were normoglycemic only for days 3 to 13 post-treatment. Insulin mRNA was detected mainly in the liver of the rAAV-hPPI(F12)-treated diabetic rats. The glucose tolerance capability of the rAAV-hPPI(F12)-treated diabetic rats was comparable to that of non-diabetic rats, even without injection of recombinant insulin. Furthermore, blood HbA1c concentrations in rAAV-hPPI(F12)-treated diabetic rats were reduced to almost the normal level. Importantly, studies of rAV or rAAV vector-dependent side effects on the targeted liver strongly suggested that only rAAV treatment caused no side effects. CONCLUSIONS: These results demonstrate that our rAAV-mediated in vivo insulin gene therapy provides safer maintenance of the insulin gene expression required for long-term and thus more effective blood glycemic control. PMID- 15651057 TI - Survival in prostate carcinoma--outcomes from a prospective, population-based cohort of 8887 men with up to 15 years of follow-up: results from three countries in the population-based National Prostate Cancer Registry of Sweden. AB - BACKGROUND: To decide on screening strategies and curative treatments for prostate carcinoma, it is necessary to determine the incidence and survival in a population that is not screened. METHODS: The 15-year projected survival data were analyzed from a prospective, complete, population-based registry of 8887 patients with newly diagnosed prostate carcinoma from 1987 to 1999. RESULTS: The median patient age at diagnosis was 75 years (range, 40-96 years), and 12% of patients were diagnosed before the age 65 years. The median follow-up was 80 months for patients who remained alive. In total, 5873 of 8887 patients (66.1%) had died, and 2595 of those patients (44.2%) died directly due to prostate carcinoma. The overall median age at death was 80 years (range, 41-100 years). The projected 15-year disease-specific survival rate was 44% for the whole population. In total, 18% of patients had metastases at diagnosis (M1), and their median survival was 2.5 years. Patients with nonmetastatic T1-T3 prostate carcinoma (age < 75 years at diagnosis; n=2098 patients) had a 15-year projected disease-specific survival rate of 66%. Patients who underwent radical prostatectomy had a significantly lower risk of dying from prostate carcinoma (relative risk, 0.40) compared with patients who were treated with noncurative therapies or radiotherapy. CONCLUSIONS: The disease-specific mortality was comparatively high, but it took 15 years to reach a disease-specific mortality rate of 56%. These data form a truly population-based baseline on how prostate carcinoma will affect a population when screening is not applied and can be used for comparison with other health care strategies. PMID- 15651058 TI - Incidence of noncutaneous melanomas in the U.S. AB - BACKGROUND: Description of the epidemiology of noncutaneous melanoma has been hampered by its rarity. The current report was the largest in-depth descriptive analysis of incidence of noncutaneous melanoma in the United States, using data from the North American Association of Central Cancer Registries. METHODS: Pooled data from 27 states and one metropolitan area were used to examine the incidence of noncutaneous melanoma by anatomic subsite, gender, age, race, and geography (northern/southern and coastal/noncoastal) for cases diagnosed between 1996 and 2000. Percent distribution by stage of disease at diagnosis and histology were also examined. RESULTS: Between 1996 and 2000, 6691 cases of noncutaneous melanoma (4885 ocular and 1806 mucosal) were diagnosed among 851 million person years at risk. Ocular melanoma was more common among men compared with women (6.8 cases per million men compared with 5.3 cases per million women, age-adjusted to the 2000 U.S. population standard), whereas mucosal melanoma was more common among women (2.8 cases per million women compared with 1.5 cases per million men). Rates of ocular melanoma among whites were greater than eight times higher than among blacks. Rates of mucosal melanoma were approximately two times higher among whites compared with blacks. CONCLUSIONS: In contrast to cutaneous melanoma, there was no apparent pattern of increased noncutaneous melanoma among residents of southern or coastal states, with the exception of melanoma of the ciliary body and iris. Despite their shared cellular origins, both ocular and mucosal melanomas differ from cutaneous melanoma in terms of incidence by gender, race, and geographic area. PMID- 15651059 TI - The treatment of primary central nervous system lymphoma in 122 immunocompetent patients: a population-based study of successively treated cohorts from the British Colombia Cancer Agency. AB - BACKGROUND: The objective of this study was to evaluate the clinical outcome of a population-based cohort of immunocompetent patients with primary central nervous system lymphoma (PCNSL) treated with 3 different strategies over 13 years. METHODS: One hundred twenty-two consecutive patients (median age, 66 years) with PCNSL were identified. Three treatment strategies were employed: 1) whole-brain irradiation with (from January, 1990, to June, 1991) or without (from April, 1995, to December, 1999) cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP)-type chemotherapy (n=50 patients); 2) combined-modality therapy, including 1 g/m2 methotrexate plus whole-brain irradiation (from July, 1991, to March, 1995; n=34 patients); and 3) 8 g/m2 methotrexate alone (from January, 2000, to March, 2003) with whole-brain irradiation reserved for those with progressive disease (n=38 patients). Treatment failure was defined as progressive disease, disease recurrence, death from toxicity or lymphoma, or toxicity that necessitated a change in primary treatment. RESULTS: The median failure-free survival was 7 months, and the median overall survival (OS) was 17 months. The median OS was similar in all 3 eras. In this population-based analysis, one-third of patients did not receive the treatment strategy of the era. Therefore, the data also were analyzed by treatment received. On multivariate analysis (including era of treatment), 3 factors-age > 60 years, lactate dehydrogenase > normal, and omission of methotrexate-were associated significantly with poorer OS (hazard ratio: 2.3, 2.2, and 2.3, respectively). CONCLUSIONS: Outcomes for a general population with PCNSL remained constant despite different treatment strategies over three eras. For the two-thirds of patients who could receive potentially curative treatment, age, lactate dehydrogenase level, and receipt of > or = 1 g/m2 methotrexate appeared to be important determinants of OS. PMID- 15651060 TI - Increase in amphiregulin and epiregulin in prostate cancer xenograft after androgen deprivation-impact of specific HER1 inhibition. AB - BACKGROUND: We investigated the expression of the epidermal growth factor (EGF) network before and after castration in the prostate cancer xenograft CWR22 implanted in nude mice, and examined the effects of gefitinib (Iresssa, ZD1839), a new drug directed towards the EGF tyrosine kinase receptor (HER1) of the EGF network. METHODS: CWR22 prostate cancer xenografts were propagated in immunodeficient male mice. The expression of the growth factors and receptors in the EGF network was examined by real-time PCR analysis and ELISA at 0, 7, 14, and 30 days after castration, and the tumor growth was examined after treatment with gefitinib or placebo. RESULTS: A fraction of growth factors showed a steady increase in the mRNA expression reaching between fourfold and eightfold 30 days after castration, including amphiregulin (P < 0.005) and epiregulin (P < 0.001). ELISA for amphiregulin showed a fivefold increase 30 days after castration. Tumor bearing mice were castrated and treated with or without the HER1 tyrosine kinase inhibitor gefitinib. Tumor involution was significantly increased by castration plus gefitinib as compared to castration alone. CONCLUSIONS: Castration leads to adaptive increase in the concentrations of a subset of growth factors from the EGF network in the androgen sensitive CWR22 prostate cancer xenograft. Specific inhibition of the HER1 tyrosine kinase receptor significantly increases the tumor involution, and suggests that HER1 targeted drugs could be of therapeutic relevance in the treatment of advanced prostate cancer. PMID- 15651061 TI - Role of insulin-like growth factor binding proteins in 1alpha,25-dihydroxyvitamin D(3)-induced growth inhibition of human prostate cancer cells. AB - BACKGROUND: The mechanisms underlying 1alpha,25-dihydroxyvitamin D(3) (1,25D) induced growth inhibition of human prostate cancer cells have not been fully elucidated. To determine whether alterations in the insulin-like growth factor (IGF) signaling axis are associated with 1,25D-induced growth inhibition, we examined the ability of 1,25D to regulate expression of IGF binding proteins (IGFBPs) in human prostate cancer cell lines. METHODS: Northern and Western blot analyses were used to detect 1,25D-induced alterations in IGFBP expression. Additional in vitro studies were performed to determine the role of IGFBP-3 in 1,25D-induced growth inhibition. RESULTS: 1,25D decreased mRNA levels of the growth stimulatory IGFBP-2 and induced IGFBP-3 mRNA in LNCaP and C4-2 cells. 1,25D treatment also increased secreted IGFBP-3 protein levels in prostate cancer cell lines sensitive to 1,25D growth inhibition but had little effect on IGFBP-3 expression in 1,25D-resistant DU145 cells. However, recombinant IGFBP-3 had only a minor effect on LNCaP cell growth in the presence of serum. Furthermore, siRNA duplexes that reduced IGFBP-3 expression did not alter 1,25D growth inhibition in either LNCaP or PC-3 cell lines grown in serum-containing media. CONCLUSIONS: Our studies indicate 1,25D-induced up-regulation of IGFBP-3 is not required for the growth inhibitory effects of 1,25D in prostate cancer cells grown in serum containing media. PMID- 15651063 TI - Personality traits, health behavior, and risk for cancer: a prospective study of Swedish twin court. AB - BACKGROUND: The authors conducted a prospective investigation into the relation between personality traits and the risk for cancer. METHODS: The study cohort consisted of 29,595 Swedish twins from the national Swedish Twin Registry who were ages 15-48 years at time of entry. In 1973, the twins completed a questionnaire eliciting information on personality traits and health behavior. The Eysenck Personality Inventory was used to measure neuroticism and extroversion as two personality dimensions. A Cox proportional hazards model was used to estimate hazard ratios and 95% confidence intervals for extroversion and neuroticism separately as well as for their joint effect, and conditional logistic regression analyses were conducted to estimate the relation between personality traits and risks for cancer in twin pairs who were discordant for cancer. All analyses were conducted for six etiologically different groups of cancers: hormone-related organ cancers, virus-related and immune-related cancers, digestive organ cancers (excluding liver), respiratory organ cancers, cancers in other sites, and all cancer sites. RESULTS: Follow-up in the Swedish Cancer Registry for 1974-1999 revealed 1898 incidents of primary cancer. The authors found no significant association between neuroticism, extroversion, their joint effects and the risk for any cancer group. CONCLUSIONS: The current results did not support the hypothesis that certain personality traits are associated with cancer risk. PMID- 15651062 TI - In vivo imaging of retinoic acid receptor beta2 transcriptional activation by the histone deacetylase inhibitor MS-275 in retinoid-resistant prostate cancer cells. AB - BACKGROUND: In retinoid resistant epithelial tumors, the lack of retinoic acid receptor beta2 (RARbeta2) expression due to epigenetic silencing impairs the activation of retinoid target genes including RARbeta2, and has been associated with the development of cancer. In this study we developed a strategy to monitor the re-activation of RARbeta2 by chromatin remodeling agents combined with retinoids in real time, and to correlate the RARbeta2 re-activation with anti tumor activity. METHODS: We selected the RARbeta2-negative retinoid resistant human prostate carcinoma cell line PC3 and stably transfected it with a luciferase expression vector under the control of a functional segment of RARbeta2 promoter (pGL2-RARbeta2-PC3). Then, we used the bioluminescence technology to monitor the reporter gene expression in real time both in vitro and in vivo following combination treatment with the histone deacetylase inhibitor MS 275 and 13-cis retinoic acid (CRA). Based on the effective dose for the RARbeta2 re-activation, we tested the anti-tumor activity of this drug combination. RESULTS: Following combination treatment with MS-275 and CRA, we observed endogenous RARbeta2 re-expression, acetylation at the RARbeta2 promoter level, and synergistic activation of the luciferase reporter gene by real time imaging both in vitro and in vivo. Combination treatment with MS-275 and CRA restored retinoid sensitivity in human prostate carcinoma cell lines, and had a greater inhibitory effect on tumor cell growth than single agents in vitro and in vivo. CONCLUSIONS: This study provides evidence that HDAC inhibitors restore retinoid sensitivity in prostate cancer cells, and in vivo real time imaging of RARbeta2 activation may represent a useful tool to study the pharmacodynamics of combination therapy with HDAC inhibitors and retinoids. PMID- 15651064 TI - Expression of tumor-associated trypsinogens (TAT-1 and TAT-2) in prostate cancer. AB - BACKGROUND: Trypsinogens are pancreatic serine proteinases and expressed in several cancers as tumor-associated trypsinogens (TAT). Trypsin mediates activation of pro-uPA and pro-MMPs, thus promoting angiogenesis and tumor invasion. Recently, we described expression of TAT in the human male genital tract and now we studied TAT in relation to PSA in PCa. METHODS: TAT expression was studied by immunohistochemistry, in situ hybridization, RT-PCR, DNA sequencing and IFMA. LNCaP cells were used to study secretion of TAT and PSA after androgen stimulation. RESULTS: Immunoreactive TAT was localized in all prostatic tumors (n = 109), lymph node (n = 16), and bone metastases (n = 17). Immunostaining intensity increased with higher Gleason's grade, whereas PSA immunostaining decreased significantly. PSA and TAT were not identically distributed in benign and malignant cells. Androgen stimulation of LNCaP cells decreased secretion of TAT and increased that of PSA. TAT mRNA was demonstrated in tissue sections and identified as TAT-1 and -2 by RT-PCR and DNA-sequencing. CONCLUSIONS: Expression of TAT is better preserved than PSA in high-grade PCa. Expression of TAT and PSA is regulated by different mechanisms as demonstrated in tissue sections and in vitro. Locally produced TAT may act in a paracrine mode to promote angiogenesis and tumor invasion in PCa by both activating and degrading of other proteinases. Further studies on the role of TAT in invasive PCa and on the mechanisms involved in the regulation of TAT expression are warranted. PMID- 15651065 TI - Elevated intraocular pressure is associated with insulin resistance and metabolic syndrome. AB - BACKGROUND AND AIM: Elevated intraocular pressure (IOP), a well-known risk factor for glaucoma, has recently been shown to be associated with some metabolic complications and obesity. We investigated the link between IOP and metabolic disturbances, focusing especially on metabolic syndrome and insulin resistance. METHODS: Eye examinations, including IOP measurement, were conducted on 943 subjects (533 men and 410 women). Body mass index (BMI), percent body fat, waist circumference, systolic and diastolic pressure, fasting insulin, glucose, lipids, and other metabolic parameters were measured. The homeostasis model assessment (HOMA) score and McAuley index were calculated to assess whole-body insulin resistance. RESULTS: Both of these insulin resistance indices showed positive associations with IOP (p < 0.05), even after statistical adjustment for other risk factors. IOP was higher in participants with metabolic syndrome, as compared to those who did not have metabolic syndrome. The mean IOP tended to increase linearly with the presence of increasing numbers of components for metabolic syndrome. CONCLUSIONS: These results suggest that insulin resistance might contribute to an explanation that would account for many previous findings concerning the association between IOP and obesity, hypertension, and diabetes. PMID- 15651066 TI - Catheter-mediated subselective intracoronary gene delivery to the rabbit heart: introduction of a novel method. AB - BACKGROUND: Recent studies suggest that gene therapy using replication-deficient adenoviruses will benefit treatment of cardiovascular diseases including heart failure. A persistent hurdle is the effective and reproducible delivery of a transgene to the myocardium with minimal iatrogenic morbidity. In this study, we sought to design a relatively non-invasive percutaneous gene delivery system that would maximize cardiac transgene expression and minimize mortality after intracoronary adenovirus injection. METHODS: Adult rabbits received a left circumflex coronary artery (LCx) infusion of 5x10(11) total viral particles of an adenovirus containing the marker transgene beta-galactosidase (Adeno-betaGal) via either a continuous infusion method utilizing an oxygenated, normothermic, physiologic pH Krebs solution driven by a Langendorff apparatus (n=12) or a timed bolus and set concentration at a constant infusion rate to the LCx (n=12). Six rabbits underwent global transgene delivery via an invasive method involving intraventricular delivery and aortic root cross-clamping. The efficacy of transgene expression via these three distinct delivery methods was determined in the left ventricle at 5 days by histological staining and colorimetric quantification assay. RESULTS: While the open-chest, aortic cross-clamping method provides the highest level of gene expression throughout the heart, the morbidity of this procedure is clinically prohibitive. Percutaneous LCx delivery of Adeno betaGal using the Langendorff apparatus was associated with the lowest morbidity and mortality while still supporting significant myocardial gene expression. CONCLUSIONS: Percutaneous delivery of an adenovirus solution using a continuous infusion of oxygenated Krebs solution via a Langendorff apparatus appears to be a gene delivery modality offering the best compromise of gene expression and clinical utility to maximize any potential therapeutic outcome. PMID- 15651068 TI - Skin-sparing mastectomy and immediate reconstruction is an acceptable treatment option for patients with high-risk breast carcinoma. AB - BACKGROUND: Skin-sparing mastectomy (SSM) followed by immediate reconstruction is an effective treatment option for patients with early-stage breast carcinoma, but its use in patients with more advanced disease is controversial. METHODS: A retrospective review was performed that included 38 consecutive patients with high-risk breast carcinoma who underwent SSM and immediate reconstruction (between July 1996 and January 2002). Tumor characteristics, type of reconstruction, margin status, timing of adjuvant therapy, postoperative complications, and incidence of recurrence were evaluated. RESULTS: High-risk patients (Stage IIA [n=4 patients] Stage IIB [n=23 patients] Stage IIIA [n=8 patients] and Stage IIIB [n=3 patients]) underwent immediate reconstruction after SSM with the use of a transverse rectus abdominis myocutaneous flap (n=31 patients), a latissimus dorsi myocutaneous flap plus an implant (n=3 patients), or tissue expanders with subsequent implant placement (n=4 patients). The median follow-up was 52.9 months (range, 27.5-92.0 months), and the median time to recurrence has not yet been reached at the time of last follow-up. The median interval from surgery to the initiation of postoperative adjuvant therapy was 38 days (range, 25-238 days). Local recurrence was seen in 1 patient (2.6%), systemic recurrence in was seen in 10 patients (26.3%), and both local and distant metastases in were seen in 2 other patients (5.3%). CONCLUSIONS: SSM with immediate reconstruction appeared to be an oncologically safe treatment option for high-risk patients with advanced stages of breast carcinoma. In addition to the aesthetic and psychological benefits of performing SSM with immediate reconstruction, local recurrence rates and disease-free survival were favorable when combined with the use of radiation therapy and adjuvant chemotherapy, as indicated. PMID- 15651067 TI - Expression of interleukin-8 gene in radical prostatectomy specimens is associated with advanced pathologic stage. AB - BACKGROUND: We previously reported that relative expression of E-cadherin, matrix metalloproteinases (MMPs)-2 and -9, and vascular endothelial growth factor (VEGF)/vascular permeability factor in radical prostatectomy specimens (RP) can distinguish organ-confined cancers from advanced prostate cancers. Here, we evaluate the expression of interleukin-8 (IL-8) and basic fibroblast growth factor (bFGF), two other genes involved in angiogenesis and metastasis, in RP specimens. METHODS: The expression level of IL-8 and bFGF mRNA in the invasive edge of 41 prostate cancers of different stages was determined using a rapid colorimetric in situ hybridization (ISH) technique. Gene expression levels of IL 8 and bFGF were correlated with the Gleason score and pathologic stage to ascertain their relationship to prostate cancer progression. RESULTS: The expression of IL-8 and bFGF genes was detected by ISH in histologically normal prostate gland epithelium as well as in glands with foci of cancer. Increased mRNA expression of IL-8 was associated with both the Gleason score and pathologic stage of tumors and distinguished organ-confined from non-confined tumors (P = 0.002). In contrast, the expression of bFGF mRNA did not correlate with the Gleason score or pathologic stage. CONCLUSIONS: Overexpression of Il-8 mRNA, but not bFGF mRNA, in RP specimens is directly associated with progression of prostate cancer. PMID- 15651069 TI - Physician-patient communication among Southern European cancer physicians: the influence of psychosocial orientation and burnout. AB - Physician-patient communication is a critical factor for comprehensive care in oncology. Although a number of studies have been carried out in Northern Europe and the US on this subject, no data are available in Southern European countries. As a part of a multicenter Southern European Psycho-Oncology study (SEPOS), the present investigation was conducted to examine communication skills and related variables (i.e. psychosocial orientation, and burnout) among 125 physicians from Italy, Portugal, and Spain. The Self-Confidence in Communication Skills (SCCS) scale was given to assess physicians' perception of their communication skills and the Expected Outcome of Communication (EOC) scale was administered to examine the physicians' expectations about the effects of communicating with their patients. Doctors' psychosocial orientation was measured by using the Physician Belief Scale (PBS) and burnout was measured by using the Maslach Burnout Inventory (MBI). Although the physicians reported receiving minimal training in communication during their education, they tended to perceive themselves as skilled in patient communication, apart from some areas (e.g. dealing with denial, managing uncertainty, assessing anxiety and depression, and promoting patient-family openness). Low psychosocial orientation and burnout symptoms (i.e. emotional exhaustion, depersonalization, and poor personal accomplishment in their job) were associated with lower confidence in communication skills and higher expectations of a negative outcome, following physician-patient communication. The results suggest that there is a need for training cancer physicians in communication and for increasing a more definite psychosocially oriented approach in cancer care in Mediterranean countries. PMID- 15651070 TI - Expression of p53, or targeting towards EGFR, enhances the oncolytic potency of conditionally replicative adenovirus against neuroblastoma. AB - BACKGROUND: Advanced stage and relapsing neuroblastoma (NB) has a poor prognosis with frequent treatment failures, warranting new treatment options and enhanced local tumor control. Treatment with conditionally replicative adenoviruses (CRAds) has shown effectiveness in various preclinical cancer models, but has not yet been evaluated for local control of NB. Here, we tested the efficacy of the CRAd AdDelta24 and of two AdDelta24 derivatives against NB. Derivative AdDelta24 425S11 infects cells deficient in coxsackie/adenovirus receptor (CAR) via the epidermal growth factor receptor (EGFR). Derivative AdDelta24-p53 expresses the tumor suppressor protein p53 to promote oncolysis. METHODS: Expression of CAR and EGFR, and p53 pathway and DNA damage responses were analyzed in six NB cell lines and two xenografts derived from primary NB using immunohistochemistry, reporter gene transactivation, Western blot and fluorescence-activated cell sorting (FACS) analysis. Efficacy of AdDelta24, AdDelta24-425S11 and AdDelta24-p53 against NB was evaluated in vitro by cell viability analysis and in vivo by monitoring subcutaneous xenograft tumor growth in mice and by histological analysis of treated tumors. RESULTS: Neuroblastoma cell lines were sensitive to oncolysis by AdDelta24, with a higher susceptibility of those with functional p53 and intact DNA damage responses. Compared to AdDelta24, AdDelta24-p53 exhibited enhanced oncolytic potency on all NB cell lines independent of their p53 status and AdDelta24-425S11 was more effective against CAR-low IGR-NB8 cells. Moreover, five daily intratumoral injections of 10(8) plaque-forming units (pfu) of AdDelta24 p53 or AdDelta24-425S11 into subcutaneous IGR-NB8 and IGR-N91 xenografts at an advanced tumor stage yielded significant tumor growth delays (TGD). In contrast, at this dose, AdDelta24 did not cause significant TGD of neuroblastoma xenografts. Injection of AdDelta24-p53 was associated with extensive cell lysis, apoptotic cell death, and fibrous fascicles in the tumors. CONCLUSION: CRAds expressing p53 and targeted towards EGFR appear promising new agents for local control in the treatment of neuroblastoma. PMID- 15651071 TI - Identification of an immunodominant H-2D(b)-restricted CTL epitope of human PSA. AB - BACKGROUND: Human prostate specific antigen (PSA) is expressed selectively in prostate epithelium and is a potential target for the immunotherapy against prostate cancer. Various PSA-based vaccines have been reported to induce cytotoxic T lymphocyte (CTL) responses in animal models. Here, we present the identification and validation of an immunodominant CTL epitope of PSA in C57Bl/6 mice (H-2(b)). METHODS: PSA-specific CTLs were induced by immunization with a plasmid expressing PSA. Epitope specificity of the CTLs was determined by their reactivity against a panel of C-terminus truncated or mutated PSA proteins and use of bioinformatical prediction with the SYFPEITHI algorithm. RESULTS: The majority of PSA-specific CTLs were directed against a single H-2D(b) restricted epitope corresponding to the amino acid residues 65-74 (HCIRNKSVIL) of the protein. The CTLs had similar functional avidity against two putative H-2D(b) binding peptides: a 9-aa-long psa65-73 (HCIRNKSVI) and a 10-aa-long psa65-74 (HCIRNKSVIL). CONCLUSIONS: We demonstrate that the psa65-73 peptide can be used for reactivation of PSA-specific CTLs in vitro and ex vivo, and H-2D(b) pentamers assembled with this peptide are an efficient tool for monitoring of PSA-specific CTL responses after DNA vaccination. PMID- 15651072 TI - Role of community risk factors and resources on breast carcinoma stage at diagnosis. AB - BACKGROUND: The current study investigated the individual and community determinants of breast carcinoma stage at diagnosis (BCSAD) using multiple data sources merged with cancer registry data. The literature review yielded 5 studies that analyzed cancer registry data merged with community-level variables (1995 2004). METHODS: Community variables constructed for the current study reflected social and economic risk factors, physician supply, and health maintenance organization penetration. Multivariate logistic regression was used to identify the significant predictors of increasingly progressive BCSAD. RESULTS: Disparities remained for black and Hispanic females in California, who were least likely to be diagnosed early compared with their white counterparts. Younger (< 40 years) and middle-aged (40-64 years) females were less likely to be diagnosed with early BCSAD, compared with older females (> or = 65 years). Utilizing services at hospitals serving a lower volume of patients with breast carcinoma was associated with later BCSAD. After controlling for individual-level factors, community-level variables constructed at the census block group and county level were tested. If a woman resided in a neighborhood with greater percentages of female-headed households, persons living below the poverty level, less educated people, and more recent immigrants, then her chances of being diagnosed at an earlier stage were diminished. If, conversely, she resided in a neighborhood with greater percentages of females > or = 65 years (a proxy for Medicare coverage), her access to medical care and the probability of earlier BCSAD increased. County level insurance rates and residing in counties where greater percentages of women ever had a mammogram were associated with in situ and early-stage diagnosis. Similarly, the supply of primary care physicians and radiologists was associated positively with earlier BCSAD. CONCLUSIONS: Results confirmed community-level predictors of socioeconomic and delivery system context matter, although the individual-level predictors showed a stronger effect. Nevertheless, analysis of community variables is promising for guiding and evaluating the effects of health policy and developing community and delivery system interventions for earlier detection and treatment of breast carcinoma. PMID- 15651073 TI - Fetal lung volumetry using two- and three-dimensional ultrasound. AB - OBJECTIVES: To compare methods of measuring fetal pulmonary volume and to establish nomograms of fetal pulmonary volume according to gestational age for the accurate diagnosis of pulmonary hypoplasia. METHODS: Three methods of measuring fetal pulmonary volume in 39 normal fetuses were compared: two dimensional (2D) ultrasound measurement assuming that the lung is a geometrical pyramid, three-dimensional (3D) ultrasound using the VOCAL rotational method, and the conventional multiplanar 3D mode. Linear regression was used to construct an equation for 3D volume calculation from 2D measurements (the re-evaluated pulmonary volume equation (RPVE)). Lung volume measurements were recorded from 622 singleton fetuses in order to construct nomograms. RESULTS: There was no statistically significant difference between the lung volume values obtained using the two 3D modes. However, in comparison with the 2D measurements the volumes obtained were larger (mean difference = 11.99, P < 0.1 x 10(-6)). The relationship between the 2D and 3D volumes was determined using a statistical linear regression method: RPVE (mL) = 4.24 + (1.53 x 2DGPV), where 2DGPV (2D geometric pulmonary volume) = (surface area right lung base (cm2) + surface area left lung base (cm2)) x 1/3 height right lung (cm). Two nomograms were constructed, one for use with 2D and one for 3D technology. CONCLUSION: 2D pulmonary volume assessment can be used in clinical situations where fetal prognosis depends on lung volume and its growth potential. It is routinely available and easy to perform particularly when repeat measurements are required in evaluation of lung growth. We therefore propose this method as an alternative to magnetic resonance imaging or 3D ultrasound. PMID- 15651074 TI - Change in post-traumatic stress symptoms following psychosocial treatment for breast cancer. AB - The diagnosis of cancer is a traumatic experience, which may result in post traumatic stress symptoms, such as arousal, re-experiencing the diagnostic process and avoidance. Changes in post-traumatic symptoms were assessed in 181 women with breast cancer who participated in either a standard support group or complementary/alternative (CAM) oriented intervention. At baseline 26 women were classified as having significant PTSD symptoms. After the 12-week sessions, significant decreases in the number of women with PTSD was seen in both interventions, however it was more evident in the Standard group where there was a 91% reduction in the number of women with PTSD versus an 80% reduction in the CAM group. For women with PTSD both groups showed significant decreases in overall PTSD symptoms and arousal. However, only the women in the Standard support group showed significant decreases in re-experiencing and avoidance symptoms. For the entire sample only women in the Standard group had significant decreases in overall PTSD, re-experiencing, and arousal. These results indicate that PTSD symptoms can be prevalent among women with breast cancer, and that while psychosocial interventions can be effective in reducing this type of distress, a support group might be more effective than a more complementary/alternative oriented intervention. PMID- 15651075 TI - Cytokine/cytokine receptor gene expression in childhood acute lymphoblastic leukemia: correlation of expression and clinical outcome at first disease recurrence. AB - BACKGROUND: Recent studies have shown that cytokines/cytokine receptors (C/CR) affect leukemic cell growth and survival. The goal of the current study was to investigate possible correlations between gene expression patterns of C/CR in leukemic cells, clinical features, and outcome in children with acute lymphoblastic leukemia (ALL) at first disease recurrence. METHODS: Between January 1997 and December 2000, bone marrow (BM) samples were collected from 68 children with first ALL recurrence at diagnosis. These patients were enrolled in the ALL-REZ 95-96 disease recurrence trials of the Berlin-Frankurt-Munster study group. C/CR gene expression (interleukin [IL]-7, IL-10, IL-12p40, IL-15, IL-18, IL-7Ralpha, IL-10R1, IL-15Ralpha, interferon-gamma [IFN-gamma], vascular epithelial growth factor [VEGF], Flt1, and transforming growth factor-beta) was quantified by real-time reverse transcriptase-polymerase chain reaction and correlated with protein expression by immunofluorescence. RESULTS: In comparison with T-lineage ALL specimens, expression of IL-10, IFN-gamma, IL-15Ralpha, and Flt1 was significantly higher in B-cell precursor (BCP) ALL specimens (P <0.01). Among BCP ALL samples, gene expression of IL-7Ralpha and Flt1 was higher in pre-B than in common or pro-B leukemic cells. Moreover, expression levels of VEGF, IL 7Ralpha, IL-10R1, and IL-15Ralpha were lower in lymphoblasts of patients with a combined BM recurrence than in those with an isolated recurrence (P <0.05). Children with IL-15Ralpha expression above the median level had a significantly better probability of event-free survival (0.65 vs. 0.34, P=0.04) and survival (0.71 vs. 0.37, P=0.02) at 5 years. CONCLUSIONS: Expression of distinct C/CR in ALL cells was associated with lineage commitment and differentiation of leukemic cells, as well as with prognosis. It remains to be evaluated whether these prognostic and biologic findings of distinct C/CR expression in leukemic cells also have therapeutical implications for future antileukemic strategies. PMID- 15651076 TI - LIT1 and H19 methylation defects in isolated hemihyperplasia. AB - We performed LIT1 and H19 methylation studies on 27 children with isolated hemihyperplasia (IH). Eight children (29.6%) had a defect in methylation of one or both of these alleles, supporting our hypothesis that these epigenetic changes can result in a phenotype distinct from typical Beckwith-Wiedemann syndrome. PMID- 15651077 TI - The dihydroxyacetone unit--a versatile C(3) building block in organic synthesis. AB - Nature employs dihydroxyacetone phosphate (DHAP) as the donor component in various enzyme-catalyzed aldol reactions. Probably the most significant example in this regard is photosynthesis, in which D-glucose, the most widespread natural product, is formed in just a few steps from DHAP. In recent years a number of synthetic equivalents of DHAP have been reported that deserve particular attention, as their applicability in organic synthesis is not limited to (stereoselective) aldol reactions. The power of these reagents has also been demonstrated convincingly in numerous other asymmetric electrophilic alpha substitution reactions in target-oriented syntheses. Furthermore, the related 1,3 dioxins are useful equivalents of 2-substituted acrolein derivatives. PMID- 15651078 TI - Perovskite membranes in ammonia oxidation: towards process intensification in nitric acid manufacture. PMID- 15651079 TI - Stimulation of Epstein-Barr virus-infected human B cell growth by physiological concentrations of 4-hydroxynonenal. AB - We had previously shown that cyclosporin A (CsA) directly promoted the immortalization of Epstein-Barr virus (EBV)-infected human B cells (EBV-B cells) via an oxidative stress mechanism. 4-Hydroxynonenal (HNE) is a reactive end product of lipid peroxidation. We hypothesized that HNE may mediate a direct oxidative stress-promoting effect of CsA on EBV-B cells. HNE-protein adducts in CsA-treated EBV-B cell extracts were assayed immunochemically using a Slot-Blot method. Cell proliferation was assayed by [(3)H]-thymidine incorporation. EBV oncogene latent membrane protein-1 (LMP1) expression was assayed by using PE conjugated anti-LMP1 antibody in flow cytometry. We found that CsA at 500 ng ml( 1) and 1000 ng ml(-1) significantly increased the level of HNE-protein adducts in EBV-B cells over the control (arbitrary units +/- SE) by 251.3 +/- 7.5 to 361.3 +/- 9.7 and 342.7 +/- 10.7, respectively (p < 0.05, n = 3). EBV-B cells treated with a physiological concentration of HNE (1 microM) for 0.5 and 1 h and cultured for 2 and 4 weeks showed significantly increased [(3)H]-thymidine incorporation. EBV-B cells treated with HNE (1 microM) for 1 h and subsequently cultured for 2 and 4 weeks had a significantly higher ( > 2.0 times) LMP1-positive cell population over the control. In conclusion, in accordance with our previous findings, we show that CsA treatment of EBV-B cells results in increased production of the lipid peroxidation reactive end-product HNE that directly promotes EBV-B cell proliferation and LMP1 expression. This observation provides evidence for further understanding the mechanism of CsA-induced oxidative stress on EBV-related post-transplant lymphoproliferative disorder (PTLD). PMID- 15651080 TI - Investigation of oxidative stress during fracture healing in the rats. AB - One of the most damaging effects of reactive oxygen species (ROS) is lipid peroxidation, the end-product of which is malondialdehyde (MDA). This study was aimed to evaluate erythrocyte MDA levels during fracture healing in rats. Thirty male rats were used and the rats were divided into two groups to serve as controls and tests. Six rats were used as a control group that was not subject to fracture. The remaining 24 rats were divided into four groups and erythrocyte MDA levels were examined on days 5, 10, 20 and 30 post fracture. The right fibulas of rats were broken by manual angulation in the experimental group. The erythrocyte malondialdehyde level was measured in the experimental and control groups. The difference between malondialdehyde levels of control and experimental groups was statistically significant (p<0.05). Oxidative stress clearly increases during fracture healing in rats. PMID- 15651081 TI - Patterns of therapeutic prescription medication category use among community dwelling homebound older adults. AB - PURPOSE: The measurement of prescription medication use is usually through a simple count of medications, which tends to ignore therapeutic categories. This research investigated prescription medication use among homebound older adults, by documenting the therapeutic prescription medication categories used by these individuals and identifying the factors associated with use of multiple therapeutic categories. METHODS: Baseline Nutrition and Function Study (2000 2001) data from 326 homebound older persons who completed the medication review component (visual inspection of medications) of the baseline in-home interview and used > or =1 prescribed medication were included in this analysis. RESULTS: More than 40% (n = 133) regularly took medications from three to four different therapeutic categories and 31.6% (n = 103) used > or =5 different therapeutic categories. The use of respiratory medications declined with increasing age, and more women than men used diuretic and thyroid replacement medications. Independent of other factors, increased use of multiple therapeutic categories was associated with sociodemographic characteristics (gender, age, living arrangement, marital status and medication coverage), medical conditions (diabetes, heart problems and lung disease) and inability to self-manage medications. CONCLUSIONS: Our findings suggest that individual characteristics and medical conditions may help identify homebound elders at high risk for using prescription medications from an increased number of different therapeutic categories. This observation may help clinicians and community-based providers of services to older persons to be aware of differences in therapeutic medication use within an older population, and how patterns of use may alter service needs. PMID- 15651082 TI - Lipophilicity coefficients of potential tumor imaging agents, positron-labeled O(6)-benzylguanine derivatives. AB - Novel radiolabeled O(6)-benzylguanine (O(6)-BG) derivatives, 6-O-[(11)C] [(methoxymethyl)benzyl]guanines ([(11)C]p-O(6)-MMBG, 1a; [(11)C]m-O(6)-MMBG, 1b; [(11)C]o-O(6)-MMBG, 1c), 2-amino-6-O-[(11)C]-[(methoxymethyl)benzyloxy]-9-methyl purines ([(11)C]p-O(6)-AMMP, 2a; [(11)C]m-O(6)-AMMP, 2b; [(11)C]o-O(6)-AMMP, 2c), 2-amino-6-O-[(11)C]-[(methoxymethyl)benzyloxy]-9-benzyl purines ([(11)C]p-O(6) AMBP, 3a; [(11)C]m-O(6)-AMBP, 3b; [(11)C]o-O(6)-AMBP, 3c), 2-amino-6-O-benzyloxy 9-[(11)C]-[(methoxycarbonyl)methyl]purine ([(11)C]ABMMP, 4), and 2-amino-6-O benzyloxy-9-[(11)C]-[(4'-methoxycarbonyl)benzyl]purine ([(11)C]ABMBP, 5), have been synthesized for evaluation as potential novel positron emission tomography tumor imaging agents. The ability of O(6)-BG analogs to penetrate the blood- brain barrier in brain tumor could be due, at least in part, to their lipophilicity. In this paper, we measured lipophilicity coefficients (log P) of compounds 1--5 by the C(18) HPLC method. These log P values were compared, and correlations between lipophilicity and biological activity of selected analogs were made. The results suggest the appropriate level of lipophilic character in this class of compounds as useful imaging agents appears to be in the range of 1.4--2.6. PMID- 15651083 TI - Determination of ephedrine and related compounds in pharmaceutical preparations by ion chromatography with direct conductivity detection. AB - An ion chromatographic method with conductivity detection for the simultaneous determination of ephedrine, pseudoephedrine and norephedrine was developed. A mixture of 2.0 mmol/L HNO3 and 2% (v/v) acetonitrile was used as eluent. The three ephedrine-like compounds were separated and determined within 20 min. The linear ranges were 0.08-50 microg/mL for ephedrine, 0.08-40 microg/mL for pseudoephedrine and 0.06-40 microg/mL for norephedrine. The detection limits were 0.03 microg/mL for ephedrine and pseudoephedrine, and 0.02 microg/mL for norephedrine. The method has been applied successfully to the determination of these sympathomimetics in pharmaceutical preparations and in Ephedra herbs. PMID- 15651084 TI - GC-MS analysis of penta- and tetra-cyclic triterpenes from resins of Pistacia species. Part I. Pistacia lentiscus var. Chia. AB - Pistacia species contain oleoresins with bioactive triterpenes. In this study triterpenes, including minor components, were identified and quantified in both neutral and acidic fraction of Pistacia lentiscus var. Chia resin, grown exclusively in Chios island (Greece), collected traditionally, as well as by the use of stimulating agents (liquid collection). It was proved that these two resin samples were composed of several different minor triterpenes. In the traditional collection of the resin, 36 triterpenes were identified, 23 of which are new minor compounds (five in the acidic and eighteen in the neutral fraction). In the liquid collection resin eight compounds were identified in the acidic and 11 in the neutral fraction, while seven compounds were not contained in resin traditionally collected. The main triterpenes in both resin samples collected traditionally and by use of stimulating agents were in the following order: isomasticadienonic acid (24 and 22.5% w/w of triterpenic fraction respectively), masticadienonic acid (9.3 and 14.7% w/w of triterpenic fraction) and 28-norolean 17-en-3-one (19 and 36% w/w of triterpenic fraction respectively). The aim of this study was to compare the qualitative and quantitative composition of triterpenes in the resin samples collected using the traditional and new liquid techniques, and examine whether the collection technique influences the contained triterpenes in P. lentiscus var. Chia resin samples. Finally, since there is confusion on interpreting mass spectra of triterpenes we present an analytical review on the base peaks, main fragments and fragmentation mechanism/pattern of several skeleton penta- and tetra- cyclic triterpenes reported in P. lentiscus resin. Also, a biosynthetic route for triterpene skeletons contained in P. lentiscus resin was approached. PMID- 15651085 TI - Validation and application of a method for the determination of nicotine and five major metabolites in smokers' urine by solid-phase extraction and liquid chromatography-tandem mass spectrometry. AB - An SPE-LC-MS/MS method was developed, validated and applied to the determination of nicotine and five major metabolites in human urine: cotinine, trans-3' hydroxycotinine, nicotine-N-glucuronide, cotinine-N-glucuronide and trans-3' hydroxycotinine-O-glucuronide. A 500 microL urine sample was pH-adjusted with phosphate buffer (1.5 mL) containing nicotine-methyl-d3, cotinine-methyl-d3 and trans-3'-hydroxycotinine-methyl-d3 internal standards. For the unconjugated metabolites, an aliquot (800 microL) of the buffered solution was applied to a 30 mg Oasis HLB-SPE column, rinsed with 2% NH4OH/H2O (3.0 mL) and H2O (3.0 mL) and eluted with methanol (500 microL). The eluate was analyzed isocratically (100% methanol) by LC-MS/MS on a diol column (50 x 2.1 mm). For the total metabolites, a beta-glucuronidase/buffer preparation (100 microL) was added to the remaining buffered solution and incubated at 37 degrees C (20 h). An aliquot (800 microL) of the enzymatically treated buffered solution was extracted and analyzed in the same manner. The conjugated metabolites were determined indirectly by subtraction. The quantitation range of the method (ng/mL) was 14-10,320 for nicotine, 15-9800 for cotinine and 32-19,220 for trans-3'-hydroxycotinine. The validated method was used to observe diurnal variations from a smoker's spot urine samples, elimination half-lives from a smoker's 24 h urine samples and metabolite distribution profiles in the spot and 24 h urine samples. PMID- 15651086 TI - Hydrophilic interaction liquid chromatographic tandem mass spectrometric determination of atenolol in human plasma. AB - A hydrophilic interaction liquid chromatographic method with tandem mass spectrometry for the determination of atenolol, a beta-blocking agent, in human plasma has been developed and validated over the curve range of 10--2000 ng/mL. The assay was based on protein precipitation followed by evaporation of the extraction solvent, reconstitution with acetonitrile, and chromatography on an Hypersil silica column (50 x 4.6 mm) using a low aqueous--high organic mobile phase. The mobile phase consists of 85% acetonitrile, 15% water, 0.5% acetic acid and 0.04% trifluoroacetic acid and runs isocratically at a flow rate of 2.0 mL/min. The column ef fluent was split so that 50% of it was transferred into the LC-MS/MS interface operated in positive electrospray ionization mode. The chromatographic run time was 2.0 min per injection. Atenolol and the internal standard, atenolol-d(7), showed a retention time of 1.0 min. The inter-day and intra-day precision and accuracy of the quality control samples were <5.3% relative standard deviation and <8.0% relative error, respectively. To explore the application of the current method for the analysis of other beta-blocking agents, propranolol and metoprolol were tested under the same chromatographic conditions with retention times of 0.68 and 0.75 min, respectively. The present method could be used for therapeutic drug monitoring, pharmacokinetic and drug- drug interaction studies of beta-blocking agents. PMID- 15651087 TI - An application of propensity score matching using claims data. AB - BACKGROUND: Confounding by indication is a common problem in pharmacoepidemiology, where predictors of treatment also have prognostic value for the outcome of interest. The tools available to the epidemiologist that can be used to mitigate the effects of confounding by indication often have limits with respect to the number of variables that can be simultaneously incorporated as components of the confounding. This constraint becomes particularly apparent in the context of a rich data source (such as administrative claims data), applied to the study of an outcome that occurs infrequently. In such settings, there will typically be many more variables available for control as potential confounders than traditional epidemiologic techniques will allow. METHODS: One tool that can indirectly permit control of a large number of variables is the propensity score approach. This paper illustrates the application of the propensity score to a study conducted in an administrative database, and raises critical issues to be addressed in such an analysis. In this example, the effect of statin therapy on the occurrence of myocardial infarction was examined, and numerous potential confounders of this association were adjusted simultaneously using a propensity score to form matched cohorts of statin initiators and non initiators. RESULTS: The incidence of myocardial infarction observed in the statin treated cohort was lower than the incidence in the untreated cohort, and the magnitude of this effect was consistent with results from randomized placebo controlled clinical trials of statin therapy. CONCLUSIONS: This example illustrates how confounding by indication can be mitigated by the propensity score matching technique. Concerns remain over the generalizability of estimates obtained from such a study, and how to know when propensity scores are removing bias, since apparent balance between compared groups on measured variables could leave variables not included in the propensity score unbalanced and lead to confounded effect estimates. PMID- 15651088 TI - Drug utilization statistics for individual-level pharmacy dispensing data. AB - The emergence of large, computerized pharmacoepidemiologic databases has enabled us to study drug utilization with the individual user as the unit of analysis. A recurrent problem in such analyses, however, is the overwhelming volume and complexity of data. This paper reviews the methods to estimate basic epidemiologic measures of drug utilization and some methods to screen for aberrant prescribing patterns. It also presents the basis and application of the waiting time distributions that can provide information about the period prevalence, point prevalence, incidence, duration of use, seasonality and rate of prescription renewal or relapse for specific drugs. If analyzed regularly, these measures can disclose subtle trends in clinical drug use that would not be evident from the wholesale figures. In specific situations, pharmacy dispensing data without diagnostic information can be used to assess the association between drug use and adverse effects in a prescription symmetry design. PMID- 15651089 TI - Behavioral features of CHARGE syndrome: parents' perspectives of three children with CHARGE syndrome. AB - As the population first diagnosed with CHARGE Syndrome has aged, information has emerged about specific behavioral features of this syndrome. Once the medical features are managed, it is the behavioral features that interfere with our children's ability to form reciprocal relationships, to gain formal communication skills, to make the educational gains they are cognitively capable of, and to ultimately care for themselves. In telling the informal case studies of our children, we hope their behavioral commonalities, in spite of their medical and educational differences, will be apparent as well as the complexity of the multiple types of behavior observed. We remain hopeful that answers can be found that will help our children and our families cope with this very debilitating feature of CHARGE. PMID- 15651090 TI - Decreasing age at prostate cancer diagnosis over successive generations in prostate cancer families. AB - BACKGROUND: The decline in age at prostate cancer diagnosis over the past decade is partially attributable to prostate specific antigen (PSA) screening. We examined age at diagnosis over successive generations within prostate cancer families. METHODS: Families with at least two affected men were selected from the University of Michigan Prostate Cancer Genetics Project. The 1,345 individuals from 489 families were grouped into three generations. RESULTS: Risk of prostate cancer diagnosis at a given age was estimated to increase 1.31 (95% CI: 1.13 1.51) times from one generation to the next. Among men diagnosed prior to the PSA era, inferences were similar (hazard ratio = 1.28, 95% CI: 0.97-1.68). No maternal versus paternal disease transmission effect was observed. CONCLUSIONS: Age at prostate cancer diagnosis was observed to decrease over successive generations in families from an ongoing familial prostate cancer study. This finding, if confirmed, may have important implications for familial prostate cancer risk assessment. PMID- 15651091 TI - High-performance liquid chromatographic mass spectrometric identification of the photoproducts of cymoxanil. AB - The photochemical degradation of the fungicide cymoxanil {2-cyano-N [(ethylamino)carbonyl]-2-(methoxyimino) acetamide} was studied in aqueous buffer solution (pH 5.9+/- 0.1) under UV light and in laboratory conditions. The degradation followed a pseudo- first-order kinetic with significant correlation coefficient. The main photoproducts were separated and tentatively identified by HPLC/UV and HPLC/MS data, as 3-ethyl-4-(methoxyamino)-2,5-dioxo-4 imidazolidinecarbonitrile, 1-ethyl-5-(methoxyimino)-2,4-imidazolidin-2,4-dione, ethylimidazolidinetrione and {[(ethylamino)carbonyl]-amino}oxoacetic acid. A photolysis pathway of cymoxanil is proposed. PMID- 15651092 TI - Structure determination of oligomeric alkannin and shikonin derivatives. AB - Monomeric alkannin and shikonin (A/S) are potent pharmaceutical substances with a wide spectrum of biological activity and comprise the active ingredients for several pharmaceutical preparations. Therefore, the determination of the impurities, degradation products or byproducts in alkannin and shikonin samples is of great importance. Oligomeric alkannin and shikonin are formed during biosynthesis of these bioactive secondary metabolites in Boraginaceaous root plants, during tissue culture production of A/S, during alkaline hydrolysis of A/S esters and also thermal treatment of A/S. In the present study, a dimeric alkannin/shikonin compound was isolated by size exclusion chromatography from alkannin and shikonin commercial samples and its structure was determined by one- and two-dimensional NMR spectroscopy. The structure of the most abundant oligomeric species in these samples, a dimeric naphthoquinone, was established for the fi rst time, indicating that coupling of the side chain of one naphthoquinone unit with the aromatic ring of a second naphthoquinone leads to dimer formation. This type of coupling allows further oligomerization by leaving one isohexenyl side chain available at the second monomer unit. PMID- 15651093 TI - Investigation of artificial biomembrane systems in biopartitioning micellar chromatography by method of mathematical design. AB - An attempt to create and study an artificial membrane system was realized via biopartitioning micellar liquid chromatography. Towards this end the known formula of membrane permeability (on the basis of Fick's diffusion equation) was modified so that membrane permeability may be estimated in terms of chromatographic characteristics. The two-factoral experiments on the basis of mathematical design of second order were carried out. The regression equations are derived which describe the dependence of membrane permeability on the concentration of polyoxyethylene (23) lauryl ether in the mobile phase and its flow-rate for compounds with biomedical significance. Some regularities were revealed, which characterize the permeability of compounds of the different nature through membranes. The extremal dependence (with passing through minimum) of permeability on the concentration of non-ionic surfactant was observed for anionic compounds. The increasing character of permeability in relation with flow rate of mobile phase was recognized for cationic samples. Both dependences were basically fulfilled for zwitterionic compounds. PMID- 15651094 TI - Measurement of unbound cocaine in blood, brain and bile of anesthetized rats using microdialysis coupled with liquid chromatography and verified by tandem mass spectrometry. AB - To investigate the disposition of unbound cocaine in the rat blood, brain and bile, we demonstrate an in vivo multiple sampling microdialysis system coupled with liquid chromatography for cocaine assay and verified by tandem mass spectrometry. Three microdialysis probes were concurrently inserted into the jugular vein, bile duct and brain striatum of each anesthetized rat. After a period of 2 h post-surgical stabilization, cocaine (10 mg kg(-1)) was administered through the femoral vein. Separation of unbound cocaine from various biological fluids was applied to a reversed-phase C(18) column (250 x 4.6 mm I.D., 5 microm). The mobile phase consisted of acetonitrile--10 mm potassium dihydrogen phosphate buffer (25:75, v/v, pH 4.0) and 0.8% diethylamine at a flow rate of 1 mL min(-1). The UV detector wavelength was set at 235 nm. The results indicate that cocaine penetrates the blood--brain barrier with a rapid distribution. However, unbound cocaine in the bile dialysate was not detectable in the UV detection. We therefore use LC--tandem mass spectrometry to detect the bile fluid after cocaine administration (3 mg kg(-1), i.v.). The results indicate that cocaine goes through hepatobiliary excretion. PMID- 15651095 TI - A chromatographic method for baicalin quantification in rat thalamus. AB - A rapid reversed-phase high-performance liquid chromatographic (rp-HPLC) assay for the determination of baicalin in rat thalamus was developed. This was carried out on a Hypersil -C(18) column using 4-nitro-benzoic acid as the internal standard with a mobile phase of methanol-water-H(3)PO(4) (45:55:0.2, v/v/v). Detection was by UV at 277 nm. The calibration curve for baicalin was linear (r=0.9992) over the concentration range of 0.05--4.0 microg/mL and the limit of detection was 10 ng/mL. The coefficients of variation of intra- and inter-day assays were 2.64, 5.19 and 3.19% and 3.46, 6.21 and 5.58% at concentrations of 0.5, 1.0 and 3.0 microg/mL, respectively. The recoveries of baicalin from rat thalamus were 85.4+/- 5.62, 90.7+/- 2.43 and 89.1+/- 4.75% at concentrations of 0.5, 1.0 and 3.0 microg/mL, respectively. The method was applied to determine the time course of baicalin in rat thalamus, following a single dosage of intravenous administration of Scutellariae radix extract at 90 mg/kg of baicalin to male Wistar rats. PMID- 15651096 TI - Concurrent determination of thalidomide in rat blood, brain and bile using multiple microdialysis coupled to liquid chromatography. AB - A rapid and sensitive system of liquid chromatography coupled with microdialysis was developed for the simultaneous determination of unbound thalidomide in rat blood, brain and bile for pharmacokinetic study. Microdialysis probes were concurrently inserted into the jugular vein toward the right atrium, the brain striatum and the bile duct of the anesthetized Sprague-Dawley rats for biological fluid sampling after the administration of thalidomide (5 mg kg(-1)) through the femoral vein. Thalidomide and dialysates were separated using a Zorbax ODS C(18) column and a mobile phase comprising acetonitrile-methanol-0.1 mm 1 octanesulufonic acid (32:3:65, v/v/v, pH 5.3) at flow rate of 1 mL min(-1). The UV wavelength was set at 220 nm. The concentration-response relationship was linear (r(2)>0.995) over a concentration range of 0.025--25 microg mL(-1). The intra-assay and inter-assay precision and accuracy of thalidomide fell within 7%. The average in vivo recoveries were 0.31+/- 0.02,0.046+/- 0.004 and 0.57+/- 0.02 (n=6), respective to the dialysates of blood, brain and bile, with thalidomide at concentrations 2, 5 and 10 microg mL(-1). The disposition of thalidomide in the blood, brain and bile fluid suggests that there is a rapid thalidomide exchange and equilibration between the blood and brain systems. In addition, thalidomide undergoes hepatobiliary excretion. PMID- 15651097 TI - Determination of picroside II in dog plasma by HPLC and its application in a pharmacokinetics study. AB - A sensitive and simple high-performance liquid chromatography method with UV detection was developed and validated for determining picroside II in dog plasma. Paeoniflorin was employed as internal standard and the sample pre-treatment procedure consists of deproteinization by addition of acetonitrile. Chromatographic separations were performed on a Shimadzu VP-ODS column (250 x 4.6 mm i.d., 5 microm). The mobile phase consisted of acetonitrile-0.1% acetic acid aqueous (v/v), 23:77, v/v, at a rate of 1 mL/min. Detection was carried out at a wavelength of 266 nm. Calibration standards ranged from 0.25 to 500 microg/mL in dog plasma and the mean correlation coefficient of 0.9981 was found for the linear calibration curves (n = 6). The limit of quantification (LOQ) was 0.25 microg/mL. Intra- and inter-assay RSD ranged from 0.70 to 7.5%. Accuracy (%bias) ranged from -6.3 to 6.0%. This method was applied to the pharmacokinetic study of picroside II in dogs. The study demonstrated the plasma picroside II concentration-time curves were fitted to the two-compartment open model and showed linear pharmacokinetics. PMID- 15651099 TI - Research in PhD in nursing programs. PMID- 15651098 TI - Improved liquid chromatographic tandem mass spectrometric determination and pharmacokinetic study of glimepiride in human plasma. AB - An improved liquid chromatographic tandem mass spectrometric method for the determination of glimepiride in human plasma has been developed and fully validated. The article describes in detail the bioanalytical procedure and summarizes the validation results obtained. The samples were extracted using liquid--liquid extraction with a mixture of 1-chlorobutane-isopropanol-ethyl acetate (88:2:10, v/v/v). The chromatographic separation was performed on a reversed-phase Hypersil ODScolumn (250 x 4.6 mm i.d.; 5 microm particle size) using a mobile phase consisting of formic acid 0.05 M-acetonitrile (28:72, v/v), pumped at a flow rate of 0.3 ml min(-1) heated to 25 degrees C. The analytes were detected using an API 3000 triple quadrupole mass spectrometer with positive electrospray ionization in multiple reaction monitoring mode. Tandem mass spectrometric detection enabled the quantitation of glimepiride down to 0.50 ng mL(-1). Calibration graphs were linear (r better than 0.998, n=1), in concentration range 0.50--1000 ng mL(-1), and the intra- and inter- day RSD values were less than 10.37 and 11.55% for glimepiride. The method was successfully applied to a kinetic study in order to assess the main pharmacokinetic parameters of glimepiride. PMID- 15651100 TI - Directions for doctoral research. PMID- 15651101 TI - Chudnovsky replies. PMID- 15651102 TI - Drug reaction. Rev. Jackson calls study 'crime against humanity'. PMID- 15651104 TI - Global HIV/AIDS funding group issues guide. PMID- 15651103 TI - Study identifies traits of effective needle exchange programs. PMID- 15651105 TI - Asylum. Gay persecution not sufficient to prevent deportation. PMID- 15651106 TI - AIDS illness fails to secure a shorter sentence for thief. PMID- 15651107 TI - Needle exchange. Needle programs spread to halt transmission of HIV. PMID- 15651108 TI - Consumers: businesses should get involved in AIDS fight. PMID- 15651109 TI - Limited SSI benefits affirmed for HIV-positive man. PMID- 15651110 TI - Sentencing. Court finds no new evidence in rape case, upholds death penalty. PMID- 15651111 TI - Clock expires on case of an inmate sickened by AIDS drugs. PMID- 15651112 TI - Prescription drugs. Bribes alleged in sales policy for AIDS drug. PMID- 15651113 TI - HIV exposure. Hospital faces class-action suit over sterilization procedures. PMID- 15651114 TI - Freed murderer with HIV gets life for knife attacks. PMID- 15651115 TI - ADAP. Report: Alabama has longest wait list for AIDS drugs. PMID- 15651116 TI - Diagnostic methods for typhoid fever in Nigeria. AB - In this study, 65 patients are screened for Salmonella typhi by conventional culture and the Widal test. In addition, the patients undergo full blood count are screened for malaria parasites. Of the 65 patients, 50 report febrile conditions, while the remaining 15 are used as a control population. In the febrile group, 13 (26%) were positive for S. typhi, while in the control group only one (7%) was positive for S. typhi. Overall, 36 (64.3%) patients had malaria parasites. Patients with a higher O antibody titre (> or = 1 in 80) by Widal test were found to have consumed both tap water and pure water. More females (10/14; mean age: 33) had typhoid fever as a result of S. typhi infection, the majority of which were isolated from stool samples (57%). Nine of the isolates were also positive for malaria parasites, seven of which were in the trophozoite stage. Plasmodium falciparum was the predominant parasite (78%), the remainder being P. malariae. The majority of patients (12/14) with typhoid fever had normal PCV values. In conclusion, it is recommended that tests for the diagnosis of typhoid fever in Nigeria should include malaria parasites, S. typhi culture from faecal samples, and the Widal test. PMID- 15651117 TI - Recombinant human lactoferrin prevents NSAID-induced intestinal bleeding in rodents. AB - Recombinant human lactoferrin (RHLF) was tested for its ability to prevent non steroidal anti-inflammatory drug (NSAID)-induced intestinal injury in rats and mice. Acute and chronic models using indometacin, naproxen and diclofenac were used. Measurements were made of intestinal bleeding and inflammation. Orally administered RHLF was effective at preventing acute NSAID-induced increases in gut bleeding and myeloperoxidase activity. Oral RHLF was also effective at blocking some chronic manifestations of indometacin usage. Protection by RHLF of the intestinal tract from NSAIDs appears to be linked to attenuation of neutrophil migration to the intestine, and is independent of prostaglandins and nitric oxide. RHLF does not bind to the NSAID or interfere with the NSAID biological activity. We conclude that orally administered RHLF is effective at preventing NSAID-induced intestinal injury in rodents and should be investigated for this potential therapeutic use in man. PMID- 15651118 TI - Chronic oral administration of Ocimum sanctum Linn. augments cardiac endogenous antioxidants and prevents isoproterenol-induced myocardial necrosis in rats. AB - Wistar rats (200-250 g) of either sex were fed with fresh leaf homogenate of Ocimum sanctum by oral gavage in two different doses, 50 mg kg-1(Os 50) and 100 mg kg-1 (Os 100), daily for 30 days. This was followed by isoproterenol administration (85 mg kg-1 s.c. two doses at 24h intervals) in both control and 0. sanctum-fed rats to induce myocardial necrosis. Hearts were isolated for estimation of endogenous myocardial antioxidants (superoxide dismutase (SOD), catalase, reduced glutathione (GSH) and glutathione peroxidase (GPx) and myocardial lipid peroxidation) and light microscopic study. Increased basal myocardial antioxidant SOD (9.3 +/- 1.2 vs 3.7 +/- 0.7 units mg-1 protein; P<0.05) and catalase activities (34.3 +/- 5.4 vs 17.9 +/- 5.1 units mg-1 protein; P< 0.05) were observed in the Os 50 group only without any evidence of cellular injury in both the groups. In control rats, isoproterenol administration caused significant depletion of myocardial SOD (1.7 +/- 0.2 units mg-1 protein) and GPx (104 +/- 2mU mg-1 protein) activities and increase in GSH (551.7 +/- 30.9, microg g-1 wet weight of tissue) level, with evidence of myocardial necrosis. Isoproterenol-induced changes in myocardial SOD, GPx and GSH were prevented by both the doses of 0. sanctum, however cellular injury was minimal only with 50mg kg-1. The results indicate that long-term feeding of 0. sanctum offered significant protection against isoproterenol-induced myocardial necrosis through a unique property of enhancement of endogenous antioxidants. PMID- 15651119 TI - Clinical trial registration. PMID- 15651120 TI - Clinical trial registration. PMID- 15651121 TI - Chronic renal disease and cardiovascular risk. PMID- 15651122 TI - The Supreme Court and managed-care liability. PMID- 15651123 TI - Sinusitis. PMID- 15651124 TI - Thiazolidinediones. PMID- 15651125 TI - Thiazolidinediones. PMID- 15651126 TI - Proceedings of Neuropeptides 2004, the XIV European Neuropeptides Club meeting. PMID- 15651128 TI - Hungarian Rhapsody in C major--Hommage to Janos Szolcsanyi. PMID- 15651127 TI - ENC 2004 Manfred Zimmermann Award. PMID- 15651129 TI - [The Medical Association of Puerto Rico]. PMID- 15651130 TI - [Postpartum dyspnea, edema and fever]. AB - BACKGROUND: Cardiomyopathy is a condition with heart failure caused by a reduction in the contractility of the heart. Peripartum cardiomyopathy is a rare, but serious condition, characterised by development of heart failure during the last month of pregnancy or the first five months after delivery. MATERIAL AND METHODS: We present a case in which a woman pregnant with twins developed heart failure a few days after giving birth. We give a short report of symptoms, clinical signs, diagnosis, treatment and prognosis for subsequent pregnancies. RESULTS AND INTERPRETATION: The condition was initially diagnosed and treated as pneumonia, but despite treatment and improvement in her laboratory tests, her condition worsened. The symptoms were dyspnoea, peripheral oedemas and pulmonary oedema. Peripartum cardiomyopathy was diagnosed after echocardiography. Treatment of heart failure with diuretics, nitroglycerine, angiotensin-converting enzyme inhibitor and beta blocker was given with good results. This case is presented in order to draw attention to a rare, but serious condition in pregnancy or the postnatal period which easily can be misjudged or mistreated. PMID- 15651131 TI - [Heart disease as a complication to pregnancy and birth]. PMID- 15651132 TI - Special report: genotyping for cytochrome P450 polymorphisms to determine drug metabolizer status. PMID- 15651133 TI - Special report: the publication of new evidence and effect on physician prescribing behaviors. PMID- 15651134 TI - Percutaneous kyphoplasty for vertebral fractures caused by osteoporosis and malignancy. PMID- 15651135 TI - Percutaneous vertebroplasty for vertebral fractures caused by osteoporosis, malignancy, or hemangioma. PMID- 15651136 TI - Vertebral assessment using dual-energy X-ray absorptiometry for osteoporotic fracture risk assessment. PMID- 15651137 TI - Intrahippocampal nicotine in alcohol drinking rats--effects on lever-press response. AB - We have previously shown differences on learning processes between alcohol drinking and non-alcohol drinking rats. Underlying these effects, functional differences in the septo-hippocampal pathway were hypothesized. We have performed a dose-response study for intrahippocampal nicotine (CA1) on acquisition and extinction of the lever-press response and antagonization test by co administration of mecamylamine. Results show that the administration of nicotine in CAI region has a detrimental dose-dependent effect on acquisition in alcohol drinkers, with a dose of 10 nM being the most disruptive. In the controls, only doses of 10 and 20 nM had detrimental effect. The effect of nicotine (10 nM) was partially (alcoholics) or fully (controls) antagonized by mecamylamine co administration (30 nM). Summarizing, the alcohol groups showed a dose-response curve for nicotine shifted leftwards, and a partial antagonism of these effects by mecamylamine; these effects may be consequence of the functional sensitization of the nicotinic responsivity in the CAI region which were produced by the chronic alcohol. PMID- 15651138 TI - [Glamour and poverty of oncology]. AB - The problematics of oncology is staying at the fore of the health problems in our republic. Every third of us falls ill with malignant tumour and every fourth dies of it. There were 37,000 malignant tumours diagnosed in our republic in 1977, in 1999 it was 59,500 and the amount is expected to increase up to 70,000 in 2004. Our results regarding the relative 5-year survival of the patients with all the oncologic diagnoses from 15 to 44 years are below European average. The most frequent tumour is lung cancer and bronchial cancer in males, breast cancer in females and the most frequent malignant tumour in both sexes is the colorectal cancer, the absolute rate of which has unambiguously the leading position. Czech republic stands at the forefront in Europe as for the incidence of all the tumours except the skin tumours. Among all the European males it is just Czech males who have a large probability of colorectal cancer. Moreover, regarding malignant renal cancer our republic is in the first position in both males and females. What do these epidemiologic dates imply? The risk of the development of malignant disease imminently endangers each of us. For Czech republic the malignant disease constitutes a big load as personal hardship and disability, increased need for hospital bed capacities and consequent increased economic costs. In 2003, the Czech oncologic society of Czech medical company of J. E. Purkyne created the National oncologic programme (NOP) of the Czech republic. Further there are named its aims, strategy and tasks. In the conclusion we mention what is the "glamour" and the "poverty" of oncology. PMID- 15651139 TI - [Heart failure--epidemic of the 21st century]. AB - Number of patients with chronic heart failure is increasing in all developed countries. The reasons are both, the improving prevention and treatment of deadly cardiovascular diseases, like acute myocardial infarction or stroke, and the increasing life expectancy. The cardiovascular mortality has declined by 30% and the average life expectancy has increased by 4 years also in the Czech Republic during the last 15 years. The prevalence of heart failure is about 1.5% in a general population, which means that there is about 150,000 patients in the Czech Republic. The annual incidence is about 0.4%, which means that there is about 40,000 new patients in the Czech Republic every year. The prevalence is increasing with age significantly. With respect to the ageing of the population it is expected that number of heart failure patients will be increasing. Based on the results of big clinical trials the treatment of heart failure has changed significantly. ACE inhibitors and beta-blockers became the first choice treatment improving not only symptoms but also mortality. In spite of all the progress in pharmacotherapy the prognosis of heart failure is still bad. Over 40% of patients will die within 4 years after making the diagnosis of heart failure and one-year mortality of patients with advanced heart failure (NYHA class IV) is over 50%. Therefore, a research of new therapeutic possibilities is still continuing. At present, a clinical significance of different devices, like biventricular cardiac pacemakers (cardiac resynchronization therapy) or implantable cardioverter/defibrillators in heart failure treatment is studied. A gene and cell therapy represents a great hope for heart failure treatment in future. PMID- 15651140 TI - [Echocardiography--a stethoscope of the 21st century]. AB - The goal of this article is to provide a report of a current potential of an ultrasound examination of heart and to outline future development of echocardiography and its use in clinical practice. Since echocardiography appearance in the middle of the 20th century till today, the method has undergone a very dynamic development. From a simple one-dimensional image (M-mode), through two-dimensional image (2D) complemented with Doppler imaging of blood flows, echocardiography developed into currently up to date methods such as contrast echocardiography, tissue Doppler imaging, assessment of myocardial strain and strain rate, methods based on detection of ultrasonic backscatter (acoustic densitometry, automated border detection, colored image of wall motion--"color kinesis") and others. Possibility of an ultrasonic assessment of heart significantly supplemented transesophageal echocardiography and newly emerging three-dimensional imaging of heart structures. As a routine method to prove myocardial ischemia and viability a stress echocardiography is currently used. Its results significantly improved after technical innovations enabling simultaneous monitoring and evaluation of images at various projection displays and at various levels of stress. A big promise for further bigger use of echocardiography has become a miniaturisation of devices enabling one man to carry the device to a patient outside a health care facility. These small, more affordable ultrasound devices can soon become real stethoscopes of the 21st century. PMID- 15651141 TI - [New trends in pharmacotherapy of cardiovascular diseases]. AB - There has not been a year that would not have brought something new, often upheaval in the field of cardiovascular pharmacotherapy during last decades. This overview addresses the perspectives that may be expected in the treatment of cardiovascular diseases in the coming years. As for the field of dyslipidemy treatment there are some new options of blocking cholesterol resorption at the enterocyte level opening up in the field of dyslipidemy treatment (e.g. brush border transport system inhibitors, inhibitors of esterification or bind to apolipoprotein), further big revolution may be foreseen in the field of the stimulation of peroxysomal receptors controlling the lipids and glycides metabolism. It is also the field of antithrombotic drugs where we encounter the series of innovative approaches as the inhibitors of receptors facilitating the thrombocyte adhesion, new direct thrombin inhibitors or tissue factor blockers. There is no significant advance in the field of arrhythmias pharmacology, that field is completely posessed by electro-impulse therapy and ablative methods. On the contrary, great perspectives may be foreseen in the field of heart failure therapy. Along with the new methods moderating hyperactivated regulation mechanisms (e.g. renin or vasopeptidases inhibitors) promising is the field of the new inotropics active without increasing the supply of calcium (calcium sensitizers, the stimulators of sarcoplasmatic calcium ATPase). In the field of diuretics there may be expected the introduction of adiuretin blockers (akvaretics). Finally the last promising field is represented by the drugs intervening the metabolism of non-cellular matrix which are expected primarily to have a positive influence on the ventricle remodellation. PMID- 15651142 TI - [Drug eluting stents--end of restenosis?]. PMID- 15651143 TI - [Defibrillators--end of sudden cardiac death?]. AB - The ventricular fibrillation is still the main cause of a sudden cardiac death, even though it was described 155 years ago in experiment (M. Hoffa 1849) and its therapy--defibrillation--has been known since 1947 (C. Beck). In Europe 2500 inhabitants suffer from cardiac arrest daily and 90% is caused by ventricular fibrillation. A key interval for an effective defibrillation seems to be 3-8 minutes from the begining of a cardiac arrest. Automated (automatized) external defibrillators (AED) have been used for last 15 years, especially in USA. However it is still unclear how many devices will be needed and where to place them. We don't know if they improve the prognosis of patients with out of hospital cardiac arrest during ventricular fibrillation. The individualisation of the risk of a sudden cardiac death has brought a new method to the clinical practise- implantation of cardioverter-defibrillator (ICD). Their efficacy in reduction of total mortality was verified first in the field of secondary prevention--in patients after cardiac arrest (AVID study) and than in the field of primary prevention--in patients with risk markers (left ventricle dysfunction, non sustained ventricular tachycardias) but without sustained malignant arrhythmia in anamnesis (MUSTT, CIDS, MADIT I, MADIT II). Defibrillators (external, automated, implantable) obviously don't mean the end of the sudden cardiac death. The incidence of sudden cardiac death can be reduced significantly with prevention (nutrition, prevention of CAD) and one attention should be drawn to the fact even in the future. PMID- 15651144 TI - [Preventable diseases: an appeal at the beginning of the 21st century]. PMID- 15651145 TI - [Dyslipidemia at the turn of the century: optimism or scepticism?]. AB - Treatment of dyslipidaemia is one of the most important factors that decreases cardiovascular mortality and morbidity. Although we know lipid levels goals, we are not very successful in achiving them. The number of hypolipidemic substances is increasing and we have got guidelines for treatment dyslipidaemic patients. It is very important to implement this guidelines in the clinical practice. Another problem is rather poor patients's compliance both for dyslipidaemia treatment and for lifestyle changes. Pharmacologic treatment of dyslipidaemia may be limited due to its high expense. Pharmacologic treatment thus should be target to the high risk population: patients with coronary heart disease, patients with diabetes mellitus and with familial hypercholesterolaemia. In primary prevention only persons with high risk of cardiovascular event should be treated with hypolipidemic drugs. PMID- 15651146 TI - [New trends in diabetology]. AB - Number of diabetics increases enormously in the whole world. In last years many new approaches and technologies were developed e.g. strategies of diabetes prevention, new findings in metabolic syndrome pathogenesis, new findings in postprandial state, new antidiabetic and antiobesity drugs, new insulin analogues, new technologies in glucose monitoring and insulin delivery, new approaches were developed also in insulin dosing in intensive care. Many changes will influence diabetology in next years: inhalation and bucal application of insulin, development of drugs functioning in all components of metabolic syndrome, automatisation of insulin dosing (closing of feedback from glucose level to insulin dosing), experiments with embryonic and stem cells in diabetology, preventive gene vaccination in both types of diabetes. Extreme incidence of diabetes is stimulating new technologies development in diabetology. PMID- 15651147 TI - [Some important news in clinical nephrology in years 2003-2004]. AB - A new classification of chronic renal diseases enables to stratify risk of complications of chronic renal insufficiency and cardiovascular diseases. Chronic renal diseases are not rare, their incidence increases with age. Main causes of morbidity and mortality of patients with chronic renal diseases are cardiovascular complications. Besides typical risk factors, patients with chronic renal disease are at risk of specific renal diseases (such as anaemia or hyperphosphatemia). A cardiovascular risk of renal patients also increases dislipidemia. Statin treatment is able to decrease not only cardiovascular morbidity and mortality but also to slow down progression of chronic renal insufficiency. Nondiabetic nephropathies progression also significantly decreases a combination of an angiotensin converting enzyme and an angiotensin antagonist. Also in patients with chronic glomerulonephritides treatment is currently indicated according to evidence based medicine. Some monoclonal antibodies (such as infliximab or rituximab) seem to be promising among new drugs. PMID- 15651148 TI - [What has changed in knowledge of etiopathogenensis, diagnostics and therapy of gastroesophageal reflux over last 15 years?]. AB - An incidence of gastroesophageal reflux disease (GERD) in population is continuously increasing. A probable etiology, except anatomical changes (such as hiatus hernia, transient lower esophageal sphincter relaxation etc.), can also include a change in life style For diagnostic purposes upper endoscopy and in unclear cases esophageal pH-metry is used. From an etiopathogenetic point of view, besides reflux of acidic content, the alkaline content and night change in production of hydrochloric acid during treatment with proton pump blockers (PPI) are considered to be important too. Attention should be paid to extraesophageal signs of GERD. In treatment prevail PPI, in resistant cases and in younger patients fundoplications are considered, and, also therapeutic endoscopy becomes more important (application of gel prostheses, electricity to cardiac part followed with scaring or ligatures). PMID- 15651149 TI - [Gastric and duodenal ulcers--etiopathogenesis, diagnosis and therapy at the milestone of the 20th century]. AB - Peptic ulcer disease has been still very actual topic as 10% of western population would fell ill during its life. Substantial changes in understanding of etiopathogenesis were achieved within last hundred years. The first milestone means development of gastric physiology (gastric secretion, histamine, gastrin, cyclo-oxygenase, H2-receptors, proton pump). There is no doubt peptic ulcer is a typically human disease with involvement of the cortico-visceral axis and psychosomatic traits. Others factors play their role in the etiopathogenesis (like genetic, ethnic, environmental and socio-economic factors). However, main etiologic factors are Helicobacter pylori (approximately 90% duodenal and 50% gastric ulcers) and non-steroidal anti-inflammatory drugs (approximately 30% gastric and 10% duodenal ulcers). Diagnostics were determined by technical progress most of all by the construction of flexible digestive endoscopes into clinical practice. Surgery has been step by step replaced by pharmacotherapy (H2 receptor antagonists, proton pump inhibitors, Helicobacter pylori eradication). In near future we can expect further decrease in prevalence of peptic ulcer disease and gastric cancer in developed countries and increase in gastro oesophageal reflux disease. Let us hope that current controversies in indication of eradication therapy will be overcome by vaccination against Helicobacter pylori. PMID- 15651150 TI - [Idiopathic intestinal inflammations--contemporary therapeutic approaches]. AB - Current pharmacotherapy of idiopathic intestinal inflammations can benefit from standard (conventional) therapies based on results of controlled studies, non standard (unconventional) therapies based on pilot and uncontrolled studies, or can be a subject of clinical experiments. Clinical experimental area is a way to a progress in traditional treatment methods. Currently it includes a group of new drugs (immunosuppressives tacrolimus mycophenolate, thalidomide, biologic therapy, probiotics, neuroinflammation blockers), new treatment techniques (cytaphereses, sequential immunosuppression, immunosuppression with high doses), and finally new indications (chemoprophylaxis). PMID- 15651151 TI - [Chronic pancreatitis--a disease we can treat but cannot cure]. AB - Chronic pancreatitis is one of conditions with a significant impact on quality of life of patients and is an important risk factor for carcinoma emergence. Currently we have a lot of fundamental evidence concerning etiopathogenic mechanisms of chronic pancreatitis genesis. Among conclusive etiology factors belong genetic influence, heredity or autoimmune influence. However, in majority of cases the disease is not diagnosed until diagnosis of manifesting stages is made both clinically and morphologically. In spite of it new very sensitive diagnostic methods appear to give precision to diagnostics, such as new generations of NMR appliances making possible not burdening diagnostic NMR cholangio-pancreatographies or the use of intraductar endosonographic probes. Therapy still consist of treatment of symptoms. A causal therapy, with exception of an obstructive form and autoimmune form, is unknown. Above all we treat pancreatic pain and regulate symptoms of exocrine pancreatic insufficiency through an application of a capsule with micropellets enabling controlled release of enzymes in the upper part of duodenum. Undoubtedly, in future a genetic treatment will be introduced and new drugs containing pancreatic enzymes developed, primarily from the viewpoint of use and stability of digestive pancreatic enzymes. PMID- 15651152 TI - [The screening of sporadic colorectal cancer]. AB - World Health Organization (WHO) defined the criteria for screening (i.e. the location of asymptomatic individuals) of the diseases seriously endangering the public health condition. The decisive criterion is the decrease of mortality with the disease the screening is made of. The programme based on the testing the asymptomatic individuals for the presence of the fecal occult bleeding (FOBT), which was introduced also in Czech republic, meets the criterion at the screening of colorectal cancer (CR-CA), which has been proved since the year 1993 by the two case-control studies and further particularly by the three prospective, randomized, controlled studies which independently proved the decrease of mortality for CR-CA in the screened population by 15-33%. This programme has been introduced in Federal Republic of Germany since 1977 as a part of free anticancer examination. In 1995 the programme was recommended by the commission of the independent specialists of US Department of Health and in 1998 by the commission of experts of the European Group for Colorectal Cancer Screening. In 2001 the commission of independent specialists of the US Department of Health analysed the contemporary preventive programmes in USA according to the importance of the disease monitored for the condition of public health and the effectiveness of the preventive programme. Based on these criteria there were defined 8 priority preventive programmes and as for the organ malignities there was only CR-CA screening filed there. All the screening programmes are based on the colonoscopic examinations in various time intervals, depending on familiar anamnesis, polyps size and their histologic structure. An alternative to the colonoscopy is a combination of flexible sigmoideoscopy and irigography, but this technique did not establish itself in a larger scale. On the contrary the developed countries offer--in the frame of the large studies as well as standardly--the screening colonoscopy as an alternative to the occult bleeding examination. The predictive ability of the less invasive imaging methods e.g. virtual CT colography is also assayed. The screening programme in the frame of Czech republic has been standardized and as a reccommendation available to the professional community since 2000. PMID- 15651153 TI - [Renal transplantation--effective or useful?]. AB - In indicated cases the renal transplantations constitute a very effective method of solving the final renal failure. Their medical and economic importance is constantly increasing. They positively influence not only the quality of life, but also a morbidity and mortality of transplanted. It has been a long time since the comparison of annual financial costs per dialysed patient (in average 750,000 CZK) and transplanted one (in average 500,000 CZK in first year after transplantation, consequently 250,000 CZK per year) is known. Annual saving of 250,000 CZK or half a million crowns per patient with a functioning implant is a sufficiently expressive argument reflecting the effectiveness of these operations. If the total amount of patients with functional renal implant in our country is 2889, the possible saving of 1,4445 billion CZK is staggering. The quality of life after a succesful renal transplantation is incomparable with the life at the regular dialysis regime. It is documented not only by medical parametres, but mostly by the spontaneous manifestations and the evaluations made by the patients themselves. The renal transplantations are an effective and also a useful, standard method of the therapy of final renal failure in the contemporary medicine. PMID- 15651154 TI - [The role of liver transplantation in clinical practice]. AB - The first succesful liver transplantation was made in USA in 1967. These operations had been made only in few centres in the world until the 80-ies of the last century and the results were not so good. However, along with the development of surgical techniques, immunosuppression and postoperational care the results of these operations significantly improved and their amount increased. The liver transplantation was declared the clinical therapy in USA in 1983. In the same year there was made a succesful liver transplantation in Czech republic too. Nowadays there is annually made approximately 10,000 such operations worldwide. During the last decade it is also in Czech republic where the liver transplantation has become an available therapeutic method. There are two workplaces (CKTCH Brno and IKEM Prague) where 70 such operations are made every year. The results of one-year survival are more than 90% and the long-term results and the quality of life of the patients are also very good. The liver transplantation has experienced a dynamic development in the 90-ies of the 20th century and at the edge of the millenium. There were made significant changes in the field of indications and contraindications for transplantation and immunosuppressive therapy. There developed a brand new surgical techniques making possible to use the organs of the donors who are too big, so-called reduction of the implants. Another option constitute so-called split operations when one implant may be divided into left and right lobes and thus each of them may be used separately for the two donees. Due to the significant regeneration ability of the liver tissue it is possible to make the collection of the parts of the liver from the living donor. According to the statistics of the european countries the optimal need should be 10 transplantations/1 million of inhabitants per year. The difference between the need and the real amount of operations in our country is not given by the disability of the transplantation centres to make these operations but by the fact that many suitable patients are not offered this therapeutic modality at all. PMID- 15651155 TI - [New horizons in heart transplantation]. AB - Since its beginning in 1967 heart transplantations have become in 80s of the 20th century a routine treatment method of an advanced heart failure. In spite of the successes in transplantations a range of possible topics and research goals still has to be disclosed to improve quality of life and survival of patients. New horizons in heart transplantations can be summarised in following points: 1 Search of new more efficient immunosuppressives with less adverse effects. Classically a combination of Cyclosporin, Azathioprin, and corticoids have been used. Mycophenolate mofetil starts to be used in treatment instead of Azathioprin, Tacrolimus has been introduced as a drug for resistant rejections and also Rapamycin or Baziliximab are planned to be used in future. Use of Rapamycin seems desirable in patients suffering from nephropathy because it makes possible to lower doses of Cyclosporin or Tacrolimus or to stop taking them. 2 Diagnostics of rejection episodes without endomyocardial biopsy. From noninvasive methods echocardiography methods are looked for--tissue doppler imaging, densitometry, contrast echocardiography, strain and strain rate or assessing levels of some of the humoral agents: interleukin, cerebral natriuretic peptide or troponin T or direct assessment of donor DNA release in receptor's blood. 3 Earlier diagnostics of vasculopathy (here intravascular ultrasound is possible to use), particularly its pharmacology prevention. Statin treatment has been preventively introduced. 4 Xenotransplantation as a possible way of a heart replacement with an animal heart. Progresses in gene engineering lead to a development of transgenic animals, particularly pigs, with human proteins inserted in their genome and regulating activation of immune system. This way hyperacute rejection could be prevented. A range of unclear questions still remains in this area, particularly ethical ones and a risk of animal diseases transfer onto a human together with transplanted tissue. 5 Transplantation of cells from skeletal muscles (myoblasts) or marrow stem cells. 6 Development of apparatuses able to substitute heart function--total artificial heart. However, a range of questions is still unanswered in this area: a suitable material or source for long-term operation is needed to be found. Heart transplantation has become in last two decades a well-established treatment method of a serious heart failure. In spite of numerous difficulties it extends life of patients and improves its quality. Future will show whether we are able to replace it with other procedures. PMID- 15651157 TI - Release of dental records and HIPAA. PMID- 15651158 TI - Top 10 ways to avoid embezzlement. PMID- 15651159 TI - Tapping into transitions success. PMID- 15651161 TI - Reduce the stress of litigation. PMID- 15651160 TI - Emergency preparedness: a new role for the dental practitioner. PMID- 15651162 TI - [Recurrent pneumonia--case report and review of the literature]. AB - We report the case of a 70-year old man with recurrent pneumonia due to aspiration from an otherwise asymptomatic small oesophageal diverticula in the mid-oesophageal region. The diagnosis was finally established by videofluoroscopy of the oesophagus after repeated bronchoscopies and CT scans in the proceeding months. After thoracoscopic removal of the diverticula the patient remained free of disease. Oesophageal diverticula as a rare cause of repeated pneumonias should be kept in mind, even though there are no symptoms of gastro-oesophageal disease. PMID- 15651163 TI - [Microwave-induced heating injury]. AB - The case reported herein involves burns at the lower extremities leading to amputation followed by an MRSA infection with subsequent myocardial infarction and death in a patient with diabetes on chronic ambulatory peritoneal dialysis. The burns were produced by the use of a warm-up pillow containing natural stones (grape seeds) heated by microwaves. This represents another of the strange potential hazards of serious microwave related injuries. PMID- 15651164 TI - [Fake pearls... imperative day somnolence--a personal experience]. AB - In this review, narcolepsy is described based on personal experience of the author with this disease, especially with imperative day-somnolence and cataplexy. Narcolepsy is frequently underestimated and repressed. The economical (disability) and ethical (traffic accidents) implications of the disease may be significant. Optimal treatment and changes of behaviour are important. PMID- 15651165 TI - [Septic ovarian vein thrombosis after cesarean section]. AB - Report of a case of septic ovarian vein thrombosis after cesarean section and premature rupture of membranes. The clinical signs were first to suspect the thrombotic complication. Although antibiotics were given immediately after the cesarean section, heparin administration was too late to prevent the right ovarian vein from being occluded by a large thombus. The resection of the occluded ovarian vein led, under the continued administration of heparin per infusion, to a fast rehabilitation of the patient. PMID- 15651166 TI - [An auto-iatrogenic disease]. AB - A 55-year-old practitioner from an island in the northern sea felt an increasing hypersensitivity of his entire body to various ambient and nutritional allergens and toxics. He started to treat himself with increasing doses of glucocorticoids and moved to a southern climate in Lanzarote and later on to the Swiss mountains in the grisons. On admission to our hospital in December he was in a disastrous psychotic condition, trying to cool down his body by laying naked on his bed at ambient temperatures around the freezing point. He had consumed on average 250 mg prednisone daily over weeks. As we found out later his personal assistant travelling with him was giving him glucocorticoids through the infusion during his hospital stay. He developed a necrotizing septic phlebitis at the infusion site followed by a Pseudomonas aeruginosa sepsis with fatal multiorgan failure. This case illustrates the dangers of self-treatment by doctors and the difficulties in treating a physician. PMID- 15651167 TI - [An unsual case of an extensive heat necrosis]. AB - We report on an unusual case of an extensive heat necrosis of the tibia and its surrounding tissues including the skin after closed IM nailing. The first clinical sign appeared shortly after the procedure as circular skin necrosis over the tibial crest. The severe necrosis of a bone segment, the surrounding soft parts as well as the skin was probably the sequella of reaming of a pathologically altered bone with the tourniquet inflated. Only after a free latissimus dorsi transfer and the resection of the dead bone that was then bridged by a segmental bone transport according to the Ilizarov technique the situation could be salvaged. We concluded that reaming with the tourniquet inflated should never be done, as it can cause severe damage due to abnormal heat generation. PMID- 15651168 TI - [Acute renal failure--"a well meant present from a friend"]. AB - Acute allergic interstitial nephritis (AIN) due to non-steroidal anti inflammatory drugs (NSAID) is a well known but rare adverse drug event. Here, we describe the case of a 70 year old woman with recurrent episodes of acute renal failure. A first episode of (AIN) occurred after the intake of mefenaminic acid tablets. A second episode of AIN occurred two years later, this time after transdermal application of diclofenac. Our case illustrates cross-reactivity between NSAIDs and shows that transdermally applied medication can cause systemic adverse events as well. Patients do not mention ointments because they often do not realize that ointments contain active substances, and physicians forget to ask. PMID- 15651169 TI - [Interviewing the patient--an "impossible" task]. AB - A patient with atypical facial pain (conversion symptom) is presented, who had to suffer for more than 10 years, because physicians were unable to conduct a biopsychosocial interview. The characteristics of hard and soft data are mentioned. The significance of empathy for the data collection is stressed. The relationship between empathy and countertransference feelings is briefly discussed. PMID- 15651170 TI - [Sudden incapacity of walking]. AB - Two cases of pain and sudden incapacity of walking are presented. In one case aortic dissection, and in the other a massive cholesterol embolism due to aortography were the causing events. The diagnostic evaluation of the patients and the organisation of the emergency services were insufficient in both cases. There was also a lack of adequate communication with patients and relatives, leading to a loss of trust in the hospitals. Although the diagnoses were incorrect, this had no influence on the clinical courses. Nevertheless, the demonstrated deficiencies should stimulate the reevaluation of one's own emergency concept. PMID- 15651171 TI - Apoptosis, Bcl-2 antisense, and cancer therapy. AB - Apoptosis, or programmed cell death, is a complex process of cell turnover involved in both normal and pathologic processes in the body. Impairments in the apoptotic pathways contribute to tumorigenesis and the development of tumor resistance to chemotherapy. The proto-oncogene bcl-2 appears to serve a critical antiapoptotic function. Its broad expression in tumors coupled with its role in resistance to chemotherapy-induced apoptosis make bcl-2 a rational target for anticancer therapy. The Bcl-2 antisense drug oblimersen sodium (Genasense) enhances apoptosis alone and in combination with cytotoxic chemotherapy in vitro and in numerous xenograft models of solid tumors and hematologic cancers. Results from xenograft models of melanoma were especially encouraging, prompting melanoma to be identified as an initial human trial candidate. In a phase II trial in patients with advanced malignant melanoma resistant to first-line chemotherapy (including dacarbazine [DTIC-Dome]), three objective responses and three minor responses to oblimersen plus dacarbazine were observed among 14 patients. In a large randomized phase III trial, oblimersen plus dacarbazine showed a near doubling of response rate vs dacarbazine alone and a significant prolongation of progression-free survival. For the primary endpoint of overall survival, a significant benefit for the combination was not seen. The ability of oblimersen to modulate apoptosis suggests a new paradigm of anticancer therapy that has clinical potential in a variety of solid tumors and hematologic malignancies. Further, oblimersen is the first antisense molecule studied in clinical trials for its anticancer properties, opening up an entirely new direction for therapy. PMID- 15651172 TI - Apoptosis mechanisms: implications for cancer drug discovery. AB - Defects in the regulation of apoptosis (programmed cell death) make important contributions to the pathogenesis and progression of most cancers and leukemias. Apoptosis defects also figure prominently in resistance to chemotherapy, radiotherapy, hormonal therapy, and immune-based treatments. Apoptosis is caused by activation of intracellular proteases, known as caspases, that are responsible directly or indirectly for the morphologic and biochemical events that characterize the apoptotic cell. Numerous proteins that regulate these cell death proteases have been discovered, including proteins belonging to the Bcl-2, inhibitor of apoptosis, caspase-associated recruitment domain, death domain, and death effector domain families. These caspase-regulating proteins provide mechanisms for linking environmental stimuli to cell death responses or to maintenance of cell survival. Alterations in the expression and function of several apoptosis-regulating genes have been demonstrated in cancer, suggesting targets for drug discovery. Knowledge of the molecular details of apoptosis regulation and the three-dimensional structures of apoptosis proteins has revealed new strategies for identifying small-molecule drugs that may yield more effective treatments for malignancies. Apoptosis-regulating genes are also beginning to find utility as targets for antisense oligonucleotides. PMID- 15651173 TI - Bcl-2 antisense therapy in multiple myeloma. AB - Most malignant plasma cells overexpress Bcl-2, which contributes to resistance against apoptosis induced by dexamethasone and other anticancer agents. Oblimersen sodium (Genasense, previously known as G3139), an antisense oligonucleotide that specifically binds to bcl-2 messenger RNA, decreases production of Bcl-2 protein in both human myeloma cell lines, as well as in ex vivo purified myeloma cells, and enhances the cytotoxicity of dexamethasone and doxorubicin. Combining oblimersen with other anticancer agents represents a therapy-enhancing strategy to reverse the multidrug resistance seen in multiple myeloma (MM). Phase II trials are evaluating the potential role of oblimersen in reversing resistance to standard therapies. Preliminary results from these trials in patients with refractory or relapsed MM indicate that the combination of oblimersen with dexamethasone/thalidomide (Thalomid) or vincristine/doxorubicin/dexamethasone is active and well tolerated and that oblimersen may help overcome chemotherapy resistance and restore sensitivity to MM cells. A randomized phase III clinical trial comparing dexamethasone plus oblimersen with dexamethasone alone in patients with relapsed or refractory myeloma has completed enrollment, with results expected to be available in 2004. Future studies will focus on the role of oblimersen in combination with novel biologic agents such as bortezomib (Velcade). PMID- 15651174 TI - Targeting the proapoptotic factor Bcl-2 in non-Hodgkin's lymphoma. AB - Bcl-2 functions as a key survival factor for lymphocytes and is highly expressed in a majority of non-Hodgkin's lymphomas. The ability of oblimersen sodium (Genasense, previously known as G3139) to target bcl-2 messenger RNA and decrease Bcl-2 protein levels has the potential to enhance the activity of cytotoxic chemotherapy. Pretreatment with oblimersen followed by cyclophosphamide (Cytoxan, Neosar) markedly improved survival relative to single-agent cyclophosphamide in a murine xenograft model. Oblimersen has also enhanced the cytotoxicity of a variety of other agents against non-Hodgkin's lymphoma, including etoposide, rituximab (Rituxan), and alemtuzumab (Campath). An initial phase I study of oblimersen in non-Hodgkin's lymphoma demonstrated modest single-agent activity. Recent reports suggest that oblimersen may add to the activity of R-CHOP (rituximab-cyclophosphamide/doxorubicin/vincristine/prednisone) in previously untreated mantle cell lymphoma and to rituximab alone in a variety of subtypes of relapsed non-Hodgkin's lymphoma. Additional studies in both treatment-naive and relapsed patients will define the role of oblimersen in the treatment of non Hodgkin's lymphoma. PMID- 15651175 TI - Potential therapeutic applications of oblimersen in CLL. AB - Bcl-2 protein is upregulated in a wide variety of lymphoid malignancies, including chronic lymphocytic leukemia (CLL). The protein is thought to be responsible for maintaining the viability of malignant lymphoid cells and may contribute to chemotherapy and radiotherapy resistance. Previous studies have shown that reduction of bcl-2 expression by antisense therapy sensitizes cells to chemotherapy-induced apoptosis. In vitro, the Bcl-2 antisense drug oblimersen sodium (Genasense, previously known as G3139) enhances the apoptotic response in CLL cells to fludarabine (Fludara), corticosteroids, alemtuzumab (Campath), and rituximab (Rituxan). A phase I trial in patients with refractory/relapsed CLL showed that patients with CLL were more sensitive to oblimersen than patients with solid tumors. The maximum tolerated oblimersen dose was 3 mg/kg/d, and the most common dose-limiting reaction was hypotension, frequently in association with high spiking fever. In this study, oblimersen displayed limited single-agent activity, including tumor lysis syndrome, transient decreases in circulating CLL cells, and reduction of splenomegaly and size of lymph nodes. Major responses were observed in 9% of patients. Subsequently, a phase III trial evaluating fludarabine and cyclophosphamide with or without oblimersen (3 mg/kg/d for 7 days) was initiated in patients with relapsed or refractory CLL. This trial recently completed accrual of 241 patients. PMID- 15651176 TI - Apoptotic mechanisms of gallium nitrate: basic and clinical investigations. AB - Gallium nitrate inhibits the growth of various lymphoma cell lines in vitro and exhibits antitumor activity in patients with lymphoma. The mechanism(s) of cytotoxicity is (are) only partly understood but appears to involve a two-step process: (1) targeting of gallium to cells, and (2) acting on multiple, specific intracellular processes. Gallium shares certain chemical properties with iron; therefore, it binds avidly to the iron transport protein transferrin. Transferrin gallium complexes preferentially target cells that express transferrin receptors on their surface. Expression of transferrin receptors is particularly high on lymphoma cells. Cellular uptake of the gallium-transferrin complex leads to inhibition of cellular proliferation primarily via disruption of iron transport and homeostasis and blockade of ribonucleotide reductase. Recent studies have shown that cellular uptake of gallium leads to activation of caspases and induction of apoptosis. In phase II trials in patients with relapsed or refractory lymphoma, the antitumor activity of gallium nitrate is similar to, or better than, that of other commonly used chemotherapeutic agents. Gallium nitrate is not myelosuppressive and may be used in patients with neutropenia or thrombocytopenia. A multicenter trial to evaluate the use of gallium nitrate in patients with relapsed non-Hodgkin's lymphoma is currently ongoing. PMID- 15651177 TI - Management of invasive mycoses in hematology patients: current approaches. AB - Candidiasis and aspergillosis are the most common fungal infections in hematopoietic stem cell transplant recipients and other hematology/oncology patients. Strategies for reducing the morbidity and mortality associated with these infections include antifungal prophylaxis, empiric therapy in patients with persistent fever and neutropenia, and preemptive therapy. Antifungal therapies include amphotericin B deoxycholate, lipid formulations of amphotericin B, the triazoles (fluconazole, itraconazole, and voriconazole), and the echinocandins (caspofungin and the investigational agents micafungin and anidulafungin). Fluconazole is a reasonable choice for the treatment of invasive candidiasis if the patient has not previously received a triazole and the institution has a low incidence of triazole resistance. If resistance is a concern, an echinocandin, such as caspofungin, is an appropriate option. Voriconazole may be the initial choice in most patients with invasive aspergillosis. If patients are intolerant of or refractory to conventional therapy, effective alternatives include a lipid formulation of amphotericin B or an echinocandin. PMID- 15651178 TI - Use of newer antifungal therapies in clinical practice: what do the data tell us? AB - Considering the significant morbidity and mortality associated with invasive fungal infections in immunocompromised patients, it is particularly important to make the diagnosis as early as possible and to make best use of the available antifungal drugs for prophylaxis and treatment. The newer antifungal drugs include the lipid products of amphotericin B, such as amphotericin B lipid complex (ABLC) and liposomal amphotericin B; voriconazole (a triazole); and caspofungin (an echinocandin). ABLC and liposomal amphotericin B are as effective as amphotericin B deoxycholate but are less nephrotoxic; ABLC is probably the drug of choice for zygomycosis. Voriconazole is approved for use in the treatment of invasive aspergillosis and may have a role in preventing breakthrough fungal infections in patients with persistent fever and neutropenia. Caspofungin is effective against both invasive aspergillosis and invasive candidiasis. PMID- 15651179 TI - Combination antifungal therapy: where are we now, and where are we going? AB - Combination therapy with amphotericin B and flucytosine is considered to be the treatment of choice for cryptococcal infections. However, for other infections and combinations of antifungal infections, the data are less clear-cut. The concurrent use of amphotericin B with an azole has elicited controversy, given the potential of antimicrobial antagonism. The results of one recent candidemia study suggest that the potential antagonism may not be an issue; the combination of amphotericin B and fluconazole provided more effective clearance of Candida from the bloodstream than did fluconazole used alone. Several in vitro and animal studies have shown antagonism between the azoles and amphotericin B for aspergillosis. However, introduction of the new class of agents that target beta glucan synthase (echinocandins) has invigorated the prospects of combination therapy. The echinocandins and polyenes are not antagonistic, and there is evidence that the echinocandins may provide additive to synergistic activity in combination with triazoles. For patients whose aspergillosis is progressing despite monotherapy, the addition of a second agent, such as an echinocandin, may be reasonable. PMID- 15651180 TI - Merging medicine with science: the birth of a targeted therapy in cancer. PMID- 15651181 TI - Bone marrow transplantation for leukaemia--current status and strategies for improvement. AB - Thirty-five years ago, bone marrow transplantation was first being explored as a last-ditch effort to treat patients with end stage leukaemia. Through the efforts of a large number of laboratory and clinical scientists, the application of transplantation has broadened and outcomes have dramatically improved. The science of transplantation continues to attract a great deal of research, and with this effort we can expect continued progress and patient benefit. PMID- 15651182 TI - Stem cell and cartilage in orthopaedics. PMID- 15651183 TI - Endocrinological aberrations in survivors of the severe acute respiratory syndrome (SARS) in Singapore. PMID- 15651184 TI - Epidemiology of recurrent abdominal pain among Singaporean adolescents. PMID- 15651185 TI - Comparison between haematological parameters in severe acute respiratory syndrome (SARS) and dengue fever (DF). PMID- 15651186 TI - Prevalence of dementia in Singapore--results of the National Mental Health Survey of the Elderly 2003. PMID- 15651187 TI - Preoperative microalbuminuria, haptoglobin phenotype 2-2, and age are independent predictors for acute renal failure following coronary artery bypass graft. PMID- 15651188 TI - Proteomics of breast cancer: enhanced expression of CK19 in HER-2/neu-positive tumours. PMID- 15651189 TI - Celecoxib enhances brain tumour cell radiosensitivity leading to massive tumour necrosis. PMID- 15651190 TI - In vitro drug sensitivity and expression profiling for disease prognostication in childhood acute lymphoblastic leukaemia (ALL): an exploratory model using cell lines. PMID- 15651191 TI - The effects of exercise on the functional status of the elderly persons living in the long-term care setting of a psychiatric hospital. PMID- 15651193 TI - Between the rod and reason: a study on Asian parental disciplinary methods and child emotional/behavioural outcomes. PMID- 15651192 TI - The impact of microanastomosis of the intramuscular nerve branch on the healing of a completely lacerated skeletal muscle: a histopathological analysis. PMID- 15651194 TI - Anthropometric indices as screening tools for cardiovascular risk factors in Singaporeans: receiver operating characteristic curves analysis. PMID- 15651195 TI - CC3/TIP30 expression was strongly associated with HER-2/NEU status in breast cancer. PMID- 15651196 TI - A KAP survey of evidence-based medicine and clinical practice guidelines among primary care doctors in Singapore. PMID- 15651197 TI - Attitudes and perceptions of physicians to genetic testing and establishment of genetic database. PMID- 15651199 TI - Perinatal depressive disorders in Singaporean women and their partners. PMID- 15651198 TI - Psychometric properties of a new systemic lupus erythematosus-specific quality-of life instrument (SLEQOL). PMID- 15651200 TI - Improved outcome with intensive chemotherapy in paediatric acute myeloid leukaemia. PMID- 15651201 TI - Effects of inhibitors of the tyrosine signalling cascade on antigen challenge of guinea pig airways in vitro. PMID- 15651202 TI - Serum transferrin receptor levels in the normal population and subjects with iron deficiency and thalassaemia trait. PMID- 15651203 TI - Escherichia coli-asparaginase (Elspar) is superior to Erwinia-asparaginase (Erwinase) in childhood acute lymphoblastic leukaemia (ALL) induction--an early response study using minimal residual disease (MRD) markers. PMID- 15651204 TI - Determinants of patient's willingness towards participation in clinical drug trial in a psychiatric setting. PMID- 15651205 TI - Algorithm to achieve prescribed dialysis adequacy targets for non-compliant children on automated peritoneal dialysis (APD). PMID- 15651206 TI - Health screening outcomes that lead to health seeking behaviours. PMID- 15651207 TI - Long-term survival following liver resection for colorectal metastases--an Adelaide experience. PMID- 15651208 TI - Rapid prenatal diagnosis by AmnioPCR and AmnioFISH: routine testing for Down's syndrome (trisomy 21) and sex chromosome trisomies, but targeted testing for Edward's (trisomy 18) and Patau's syndromes (trisomy 13). PMID- 15651210 TI - The expression of insulin-like growth factor-I in periodontal healing following tooth replantation. PMID- 15651209 TI - Reconstruction of the ocular surface by transplantation of a serum free cultivated conjunctival tissue equivalent. PMID- 15651211 TI - Early detection of epithelial ovarian cancer using a proteomics-based protein profiling approach combined with a novel selection strategy. PMID- 15651212 TI - Non-invasive prenatal diagnosis of fetal gender using real-time polymerase chain reaction amplification of SRY in maternal plasma. PMID- 15651213 TI - Neuropsychological correlates of hippocampal volumes in first episode psychosis. PMID- 15651214 TI - Nuclear factor kappa B transcription profiling of genes protect against nitric oxide-induced neuronal apoptosis. PMID- 15651215 TI - Comparison of body mass index and subjective global assessment as indices of nutrition in hospital inpatients. PMID- 15651216 TI - The impact of time-to-balloon on outcomes in patients undergoing modern primary angioplasty for acute myocardial infarction. PMID- 15651217 TI - Signal-averaged electrocardiograms of children with ventricular septal defects before and after surgical repair. PMID- 15651218 TI - Causes of isolated prolonged activated partial thromboplastin time (APTT) in an acute general hospital: a guide to fresh frozen plasma (FFP) usage. PMID- 15651219 TI - MARS liver dialysis in children with acute liver failure. PMID- 15651220 TI - The influence of breast feeding compared to formula feeding on infant adiposity. PMID- 15651221 TI - Use of complementary and alternative medicine in paediatric oncology patients in Singapore. PMID- 15651222 TI - Psychiatric morbidity among emergency department doctors and nurses after the SARS outbreak. PMID- 15651223 TI - Impact of reversibility of no-reflow phenomenon on 30-day mortality following percutaneous revascularisation for acute myocardial infarction--insights from a 1328-patient registry. PMID- 15651224 TI - Cardiovascular abnormalities in children on long-term dialysis: analysis of risk factors. PMID- 15651225 TI - Umibilical cord blood stem cell from unrelated donors is a feasible alternate stem cell source for transplant in patients with genetic diseases. PMID- 15651226 TI - Double deletions of glutathione S-transferase genes (GSTM1 and GSTT1) reduce the risk of early relapse in childhood B-lineage acute lymphoblastic leukaemia (ALL). PMID- 15651227 TI - Evaluation of particulate respirators during the SARS outbreak. PMID- 15651228 TI - Laparoscopic radical nephrectomy: oncologic outcome in 100 cases. PMID- 15651229 TI - Aquaporin-4 is correlated with peri-tumoural oedema in meningiomas. PMID- 15651230 TI - Estrogen receptor alpha gene PvuII polymorphism and polycystic ovary syndrome. PMID- 15651231 TI - Ultrarapid prenatal detection of Down syndrome using real-time multiplex polymerase chain reaction (PCR) in amniotic fluid. PMID- 15651232 TI - Ceramic restorations. PMID- 15651233 TI - Comparing resin-based composites. PMID- 15651234 TI - TMJ lubrication system: its effect on the joint function, dysfunction, and treatment approach. AB - Normal temporomandibular joint (TMJ) movements primarily depend on free sliding of the disc down the slope of the eminence. Therefore, understanding the lubrication system and how aberrations in this system contribute to TMJ is important. Its effect on dysfunctions, such as disc displacement, anchored disc phenomenon, open lock, and osteoarthritis, are discussed in this article. PMID- 15651235 TI - Dental industry and academia unite to foster product development. PMID- 15651236 TI - Ten-year retrospective observations of the impact of a rotary-powered brush vs manual techniques in periodontal maintenance. AB - The purpose of this retrospective study is to determine whether the type of home brushing instrument used by periodontal patients can affect recare compliance and tooth retention. Data from 201 cohorts, half of which used a rotary mechanical brush, were evaluated throughout 10 years. At recall, 25% of the manual and 67% of the rotary users exhibited good to very good oral hygiene, the rotary group lost 0.082 teeth/patient/year compared with 0.185 teeth/patient/year, and 92% of the rotary group maintained acceptable recare intervals compared with 51% of the manual group. The findings for the efficacy of the rotary brush for these 3 parameters are both statistically and clinically relevant. PMID- 15651237 TI - "Don't bleach until you see the white of their eyes". PMID- 15651238 TI - Using a dental operating microscope in a pediatric dental practice. AB - It often is suggested that using a dental operating microscope on children cannot work because of their inability to sit still. Not only is this premise incorrect, but also using the microscope can improve the ergonomics of practicing dentistry and, in some cases, reduce the potential for head and neck strain. PMID- 15651239 TI - Zirconia: simplifying esthetic dentistry. PMID- 15651240 TI - [Anatomy and physiology of the human trapezometacarpal joint]. AB - Biomechanics of the trapeziometacarpal joint (TMJ) are controversial, because of differences between its normal anatomical description and theories of its pathophysiology. The aim of this article is to explain the mechanisms underlying trapeziometacarpal osteoarthritis by means of the anatomy and physiology of the TM joint. Having described the main ligaments and their roles in the physiology of the joint, it is then difficult to understand why trapezectomy usually improves patients with TMO, when the ligaments are destroyed or only partially repaired by tendinoplasty. Different explanations of TMO are detailed but it seems, according to the literature, that TMO can be explained more by genetic considerations than anatomic dysfunction. The theory of Zancolli is not verified by facts, and other concepts such as ligament theory, and dysplasia of the caudal aspect of the trapezial bone are not sufficient to understand this disease. The same discussion is engaged with regard to explaining the automatic rotation of the thumb in the TM joint. The cantilever bending principle is described as proposed by Bettinger and al. PMID- 15651241 TI - [Wrist arthroscopy. Current indications and results]. AB - Arthroscopy of the wrist is a recent technique but already validated for many indications. It rarely gives rise to complications but requires a learning curve just like any other endoscopic procedure. It allows complete visualization of the osseous, articular, synovial and ligamentous structures of the different joints comprising the wrist. There are two main indications for wrist arthroscopy: i) Diagnostic; when other investigations have proved inconclusive (assessment of carpal instabilities, osteochondral fractures and certain synovial pathologies...); ii) therapeutic; these have recently widened in terms of carpal pathology. Lesions of the triangular fibrocartilage complex (traumatic or degenerative) are best diagnosed and treated arthroscopically. Arthroscopic treatment is the least invasive, most effective and safest means of performing procedures such as debridement, TFCC reattachment, and ulna shortening. Other conditions can benefit greatly from wrist arthroscopy such as certain wrist ganglions, rheumatoid synovitis, radiocarpal fractures and some scaphoid fractures. Degenerative pathology of the wrist also presents opportunities for articular debridement ("shaving"), synovectomy or even various "ectomies". PMID- 15651242 TI - First metacarpal lengthening following traumatic amputation of the thumb: long term follow-up. AB - Thumb amputation is a relatively frequent injury in industrial accidents, producing a significant effect on hand function. The long-term results of 18 first metacarpal elongations in patients who had had a non-replantable thumb amputation at the metacarpophalangeal level are reported. Lengthening was performed using a unilateral external fixator. After a learning curve with a high complication rate and prolonged treatment, good functional results were obtained after some technical modifications, with eight patients returning to their previous occupation, and the rest requiring a change in their job. PMID- 15651243 TI - [Technical note: ulnar "pseudo-pollicisation" in congenital hand differences]. AB - In certain rare cases of thumb aplasia or hypoplasia, pollicisation of the second finger is not recommended or impossible. It happens when the most radial fingers are hypoplastic or abnormal or when an ulnar prehension pattern has already developed. In these cases, if functional, pollicisation of the most ulnar finger in not suitable because of the ugly cosmetic result. In order to combine the functional advantages of the ulnar pollicisation with a satisfactory cosmetic appearance, the authors describe pseudo-ulnar pollicisation by supination osteotomy. This operation combines rotation of the fifth ray with the establishment of a larger web space between the two most ulnar fingers. We have used this technique in five preliminary cases with a minimum follow-up of 18 months. PMID- 15651244 TI - [The ulnar longitudinal deficiency: proposition of a descriptive classification]. AB - Ulnar longitudinal deficiency is an extended malformation sometimes involving the whole upper extremity, even including sometimes the opposite side. The clinical and radiological aspects are variable and none of the existing classifications takes into account all the possible deformities. Multiple decisive factors in the surgical indications are missing such as shoulder stability, elbow position (extension or flexion with or without pterygium), orientation of the hand (internal rotation), wrist inclination and number of digits. Based on a review of our 46 clinical cases as well as the published cases in the literature we have developed a simple way to describe each level. PMID- 15651245 TI - [What wrist fracture did Pouteau actually describe?]. AB - The so-called " Pouteau-Colles" fracture was never specifically described by Pouteau. This surgeon from Lyons wrote a paper published in his posthumous works in 1783. The author was allowed access to an original edition of the paper at the "Ancienne Ecole de Medecine Navale de Rochefort". When reading this paper, it seems that Pouteau described a great variety of different kinds of forearm fractures. His original contribution was to define the mechanism of injury as occurring during a fall on the outstretched hand as opposed to direct injury. Pouteau presents a hypothetical muscular theory of fracture pattern etiology, but one that is useful in the understanding of the displacemnt of the bone fragments. He questions the principal role attributed to sprains and luxations at this time in the differential diagnosis of forearm injury. He described four clinical signs to diagnose, at a glance, which bone was injured, the site of the fracture, its displacements and mechanism of inury. Pouteau also presented a precise method of reduction and an original system of immobilisation, able to accomodate secondary displacements and avoid complications that were not fully recognised at that time, namely compartment syndrome and nosocomial infection. Pouteau simply concludes with a question concerning the future methods needed to reduce bony collapse and to fill the bone defects. In order to render special homage to this visionary surgeon, the author propose to attribute to the nosologic entity "fore arm bone fractures" the qualificative of "Pouteau's fractures", or to associate to every fracture of the fore-arm described by any other author the name of the Lyons surgeon! PMID- 15651246 TI - [Tumeral calcinosis: case report of an erosive form affecting the long finger]. AB - Tumoral calcinosis is characterised by deposits of hydroxyapatite in the soft tissues. The authors report an observation of localization at the level the P.I.P. joint of a ring finger. The lesion appeared to be a subcutaneous tumour but had completely eroded the distal epiphysis of the middle phalanx. The diagnosis was made radiographically and was confirmed by histology. Calcinosis presents in two very different forms; either disseminated or localised. The localised form can invade the juxta-articular gliding spaces. It may exhibit one of two clinical courses: one is acute and diffuse. The other is chronic, localised and insidious and gives rise to tumoral masses arising near joints, but without invading them. An erosive tumoral calcinosis is exceptional. It is characterized by bony right up to the articular surfaces. At the level of the wrist and the hand, tumoral forms are rare and we could only find one other case in the literature and it was localized in the middle finger. PMID- 15651247 TI - [Primary Swanson metacarpophalangeal joint arthroplasty in major hand traumas. A report of two cases at 14 years follow-up]. AB - In major hand injuries, destroyed joints may be treated by joint transfer, arthroplasty or fusion with or without temporary external fixation. Four immediate digital metacarpophalangeal joint Swanson implants in two patients with amputation or devascularizated near-amputation were reviewed with a 14 years follow-up. No infection was noted. Prostheses were stable with a 31 degrees active range of motion and grip strength was 42% of the opposite side. Functional results were "useful" with patients satisfaction. In one patient, heavy manual worker, implants were replaced fourteen years after trauma because of recent pain on one joint and implants fragmentation. The other patient was painless. Immediate Swanson arthroplasty in major hand injuries may be an alternative in metacarpophalangeal joints destruction. PMID- 15651248 TI - [Volar distal radio ulnar joint dislocation, case report and literature review]. AB - The authors report the case of a volar distal radio ulnar joint dislocation. The rarity of this lesion seems to be linked to the anatomical relationship of the distal portion of the ulna with the tendons of the extensor carpi ulnaris and the tendon of the extensor digiti minimi. Early diagnosis based on physical examination and radiology gives a better prognosis for closed treatment, reserving surgery for failure or irreducible luxations. PMID- 15651249 TI - Frequent flyer: loneliness is an emergency, when it's yours. PMID- 15651250 TI - Station safety. PMID- 15651251 TI - Osteoporotic hip fractures: trauma calls revisited. PMID- 15651252 TI - Prehospital care of the intoxicated individual. PMID- 15651253 TI - Developing your personal reference library. PMID- 15651254 TI - EMS at 35,000 feet. PMID- 15651255 TI - A day in the life. PMID- 15651256 TI - The basics of new product development for emergency care. PMID- 15651257 TI - Emergency treatment of avulsed teeth. PMID- 15651258 TI - Milton Packer, MD, Director, Center for Biostatistics and Clinical Science at UT Southwestern. PMID- 15651259 TI - Considerations in developing data and safety monitoring plans: a framework. PMID- 15651260 TI - Monitoring participant safety in phase I and II interventional trials: options and controversies. AB - BACKGROUND: The need for phase I and II clinical trials to have a data and safety monitoring plan (DSMP) is widely accepted; however, whether and/or when these trials should also have oversight by a data and safety monitoring board (DSMB) is controversial. Monitoring requirements also depend on whether the study is funded by the National Institutes of Health, funded and/or monitored by the US Food and Drug Administration (FDA), or funded by industry. However, there is little guidance about when and how to constitute a DSMB for phase I and II clinical trials. OBJECTIVES: The objectives of this article are to suggest guidelines for when and how to constitute a DSMB for phase I and II studies and to highlight the similarities and differences between DSMBs monitoring phase I and II versus phase III clinical studies. We highlight the utility of these guidelines in the safety monitoring of a mechanism of disease-based study of inhaled nitric oxide for acute vaso-occlusive crisis in pediatric patients with sickle cell disease that was funded and regulated by the FDA. CONCLUSION: The goal of DSMPs regardless of the phase of the clinical trial is to protect participant safety and ensure the integrity and validity of the data. A DSMB can ensure that risks and data are evaluated in a timely, efficient, and unbiased manner. We describe "risk-based" guidelines to determine situations in which a DSMB may be an appropriate addition to a DSMP for phase I and II clinical trials. We also address the roles and responsibilities of the DSMB for these studies. PMID- 15651261 TI - Forming your phase III trial's data and safety monitoring board: a perspective on safety. AB - A data safety monitoring board (DSMB) established in a phase III trial to monitor the safety of participants in a clinical trial views itself as protecting participants and ensuring the integrity of the study. The DSMB should operate under a clear charter, with expectations understood by all members of the Board, the sponsor, and the investigators. Sponsors must trust their DSMBs. The sponsor must give the DSMB the tools and the data that it needs to operate effectively in protecting the safety of the participants. PMID- 15651262 TI - Evaluating and implementing data and safety monitoring plans. AB - Data and safety monitoring plans are essential for fulfilling assurances that are given to patients enrolling in clinical trials that they will not be exposed to avoidable risks and that any information that may influence their willingness to participate in the clinical trial will be made available to them whether the clinical trial is a phase I, phase II, or phase III effort. In addition to the often explicitly detailed nomination of members of an independent data and safety monitoring board and selection of interim analysis plans for control of type I error and avoidance of conduct of a futile study, data and safety monitoring requires organization with regard to schedules for data collection, active surveillance for adverse events, with special emphasis on serious or unexpected events, and the ability to bring all information within a study and newly accruing information external to the research project together as needed for assessment of patient safety. PMID- 15651263 TI - Role of research subject advocates in the development of data safety and monitoring plans. PMID- 15651264 TI - Clinical experience with drotrecogin alfa in treating gram-positive and -negative pathogens in patients with severe sepsis. AB - BACKGROUND: Newer concepts in the management of severe sepsis and, in particular, in the understanding of the relationship between proinflammatory and procoagulant activities during severe infection have led to the introduction of activated protein C (drotrecogin) into the therapeutic program. The combination of effective antimicrobial therapy, aggressive supportive care, and efforts to antagonize procoagulants and inhibitors of fibrinolysis was used in this study. METHODS: We treated 12 patients with severe sepsis using this combination of antimicrobial agents and drotrecogin. All patients presented with hypotension and organ failure and some with multiple organ failure. Infected patients were separated into those with gram-positive and those with gram-negative infections. RESULTS: In contrast to an expected mortality rate of nearly 40% in this group of patients, only 2 (9%) expired. Both deaths were due to infection by gram-negative organisms in patients with complicated abdominal infections and concurrent cancer. All patients with gram-positive organisms survived. CONCLUSION: Those patients with infections caused by gram-positive organisms seemed to have a better prognosis than those with gram-negative infections, perhaps because their illnesses are less complicated by local disease. Although our study is small, it suggests that activated protein C will have a significant beneficial effect on the future treatment of severe sepsis and can reduce the mortality rate significantly. Further improvement in survival rates will require more effective treatment of local disease and associated noninfectious ailments. PMID- 15651266 TI - The longue duree in the history of science. PMID- 15651265 TI - Ectonucleotidase NTPDase2 is selectively down-regulated in biliary cirrhosis. AB - BACKGROUND: Portal fibroblasts are newly identified, potentially fibrogenic liver cells that are distinct from hepatic stellate cells. The ectonucleotidase* nucleoside triphosphate diphosphohydrolase 2 (NTPDase2) is restricted to portal fibroblasts in the normal liver. However, the fate of NTPDase2 after bile duct ligation (BDL) is unknown. AIMS: The aim of this study was to assess the effect of experimental rat and disease-mediated human biliary cirrhosis on NTPDase2 expression in the liver. METHODS: Cirrhosis was induced in experimental rats via BDL and carbon tetrachloride (CCl4) administration. Archived human liver biopsy specimens from normal liver, primary biliary cirrhosis, or hepatitis C cirrhosis were examined. Changes in expression of NTPDase2 were determined using confocal immunofluorescence, immunoblot, and real-time polymerase chain reaction. RESULTS: Confocal immunofluorescence demonstrated a decrease in NTPDase2 expression after BDL. Immunoblot and real-time polymerase chain reaction demonstrated a decrease in NTPDase2 expression by portal fibroblasts after BDL. No decrease in NTPDase2 protein was noted after CCl4 administration, and NTPDase2 messenger ribonucleic acid was markedly up-regulated after CCl4 administration. Confocal immunofluorescence demonstrated a shift of NTPDase2 expression from portal areas to central areas that colocalized with alpha-smooth muscle actin after CCl4 administration. In human biopsy specimens, NTPDase2 expression was lost in cirrhosis owing to primary biliary cirrhosis, whereas NTPDase2 expression was shifted to bridging fibrous bands in cirrhosis owing to hepatitis C. CONCLUSIONS: Loss of NTPDase2 is a common pathway in both rat and human manifestations of biliary cirrhosis. Conversely, in non-biliary cirrhosis, NTPDase2 is shifted from the portal area to bridging fibrous bands. Elucidations of the mechanisms regulating NTPDase2 expression may lead to new therapeutic approaches to fibrotic liver disease. PMID- 15651267 TI - [Continuity and transformation of body logic]. AB - This article is concerned with two distinct corporeal logics. In the first, corporeality is founded on joints, tendons, and mobility; in the second, the envelope and its apertures are considered primordial. The first logic is extant in very few works. Although these texts (e.g. The Iliad, Beowulf) clearly share the same, very specific, conception of the body, they belong to different histories. The corporeal logic of the 'jointed body' (corps articulaire) cannot, therefore, be appraised in terms of longue duree. The texts represent, instead, a moment of transition between the psychodynamics of orality and literacy. A problem correlated to this fact is that readers (ancient and modern) no longer think using the same logic as that pertaining to the jointed body. They tend to translate information regarding the logic of the jointed body into data meaningful in their own logic. PMID- 15651268 TI - Historians and plagues in pre-industrial Italy over the longue duree. AB - This essay deals with plague and plagues in renaissance and early modern Europe over the longue duree, principally from a methodological perspective. I shall combine an historiographical approach with an historical account of developing reactions to plague and in passing compare measures to cope in the early sixteenth century with reactions to the impact of the Great Pox or the Mal de Naples. I shall concentrate on southern Europe and in particular on Italy and my aim is to re-assess the historiography of plague through the lens of some of the more recent Anglo-Saxon literature in this field. In the process I shall outline some of the debates within the field and end with some general methodological observations drawn from early modern Italy. PMID- 15651269 TI - [The physical body and the political body: analysis of the social history of medicine (16th-17th centuries)]. AB - This paper offers an analysis of the analogies between the physical and the political body during the Sixteenth and the Seventeenth centuries. Focusing on a meaningful pattern of medical, legal and political treatises, this study aims to outline the deep roots of an organic image of the human society widespread until the Englightenment and the crisis of the Old Regime. PMID- 15651270 TI - [Pertinence of the Elias approach in history of medicine]. AB - Norbert Elias has notably influenced the historiography of the Early Modern Period since the 1970s. In the course of the borrowing of his concepts by different disciplines, the uniqueness of these concepts was nevertheless altered. This article aims to show that Elias' 'Sociology of configurations' proposed an original point of view on Longue Duree in history. It thus suggests: 1) to re evaluate the historical dimension of Elias' sociology; 2) to question eventual contributions to present-day research to the history of medicine and health. PMID- 15651271 TI - [Clinical evaluation of transbronchial needle aspiration for histological specimens]. AB - Specimens were obtained from 10 patients with radiological abnormalities involving mediastinal or hilar areas, for histological examination using 19 G transbronchial needle aspiration (TBNA). The tissues examined proved diagnostic in 6 patients (60%) including one with a benign condition (sarcoidosis). Clinical characteristics (sex, age, puncture site, number of specimens, size of radiological abnormalities) in the 6 positive patients were compared with those for the 4 negative patients. No significant differences in the number of specimens or the size of radiological abnormalities were observed between groups. However, a punctured trachea was seen in significantly more cases in the negative group (3/4, 75%) than in the positive group (1/6, 17%; p<0.05). Few minor complications were encountered. The use of a 19 G TBNA appears effective for obtaining tissue for histological examination and diagnosis of benign conditions. Training in this technique should increase the diagnostic utility of the histology needle. PMID- 15651272 TI - [Cyclosporin treatment of interstitial pneumonia and monitoring of serum concentration]. AB - Recently, cyclosporin has been reported to be a promising drug for the treatment of interstitial pneumonia. Monitoring of the serum cyclosporin concentration is important for the safety and efficacy of treatment. We measured the concentrations of this drug just before (C0) and 2 hours after (C2) administration, and the area under the concentration-time curve from the start of administration for 5 hours (AUC 0-5) in 58 patients. We found that C2 has the strongest correlation with AUC 0-5, which indicates the efficacy of cyclosporin. In 11 cases of interstitial pneumonia, 5 showed deterioration despite cyclosporin treatment. Three of those 5 cases had low C2 and AUC 0-5 levels, indicating that they were low absorbers and slow absorbers, which may be associated with a poor response. Therefore, the monitoring of the cyclosporin concentration is important especially in progressive cases of interstitial pneumonia that deteriorate despite cyclosporin treatment. PMID- 15651273 TI - [Clinical predictors for the detection of community-acquired pneumonia in adults as a guide to ordering chest radiographs]. AB - We sought to identify sensitive clinical predictors for the detection of community-acquired pneumonia in adults as a guide to the ordering of chest radiographs. The subjects were 79 outpatients with at least one clinical sign of fever, cough, sputum, chest pain, dyspnea and coarse crackle who underwent radiography of the chest to detect pneumonia. The relationship between these clinical signs and the presence of pneumonia was examined. Twenty-four patients (30.4%) had pneumonia. Twenty-two of these had 4 clinical signs: fever, cough, sputum and coarse crackle. Altogether, twenty-six of the 79 patients had these 4 clinical signs, and of them, 22 had pneumonia. Between the 4 clinical signs and the presence of pneumonia, the sensitivity was 91.7% and the specificity was 92.7%. On the basis of the above, as a diagnostic strategy, the ordering of chest radiographs to detect community-acquired pneumonia in adults was recommended when patients showed the 4 clinical signs of fever, cough, sputum and coarse crackle. PMID- 15651275 TI - [A case of constrictive bronchiolitis]. AB - A 31-year-old woman was admitted to our hospital because of a dry cough and progressive exertional dyspnea after her second delivery. She had almost normal laboratory data except for an elevated value for antibody for nucleic acid, and her chest radiograph was normal. However, pulmonary function tests revealed a mixed pattern of dysfunction, and chest CT revealed a mosaic perfusion pattern. The VATS lung biopsy demonstrated complete occlusion of the membranous bronchiole. These results led to a diagnosis of constrictive bronchiolitis (CB). The patient and her family rejected the option of lung transplantation, and selected immunosuppressive therapy (steroid pulse therapy and cyclophosphamide pulse therapy). Her condition improved temporarily, but her respiratory condition worsened progressively, and finally she died one and a half years after the appearance of the symptoms. Although steroid pulse therapy and cyclophosphamide pulse therapy failed to cure the CB, they did retard its progression. PMID- 15651274 TI - [A case of subacute idiopathic interstitial pneumonia resistant to steroids, successfully treated with cyclophosphamide]. AB - A 47-year-old woman was admitted to our hospital because of dry cough, fever, and subacute, progressive dyspnea. Chest radiography and computed tomography showed ground glass opacities in the lower lung fields. We suspected pneumonia caused by atypical pathogens and administered antibiotics, but they had no effect at all. Histopathologic findings from a transbronchial lung biopsy (TBLB) included intensive infiltration of mononuclear cells and edema on the alveolar wall with no evidence of fibrosis, fibroblasts, hyaline membrane, or granuloma. On the basis of these findings, we suspected interstitial pneumonia, but a surgical lung biopsy was not possible because the patient would not give her consent. After TBLB, corticosteroid was administered repetitively, but dyspnea was deteriorating as the ground glass opacities became more widespread, and tractional bronchiectasis appeared throughout the lung fields. Therefore, we decided to administer cyclophosphamide (CPA). This was very effective: all of her symptoms improved and the ground glass opacities and tractional bronchiectasis disappeared. Though we tapered and then discontinued corticosteroids a few months after CPA, there was no recurrence whatever. No signs suggesting the association of collagen vascular diseases were detected. The effectiveness of CPA in interstitial pneumonia associated with collagen vascular disease is occasionally reported, but the effect on idiopathic interstitial pneumonia, especially in acute and subacute progressive cases, is rarely reported. We think this is an interesting case to consider the availability of CPA in idiopathic interstitial pneumonia with subacute progression. PMID- 15651276 TI - [A case of Legionella pneumonia complicated with acute respiratory distress syndrome treated with methylprednisolone and sivelestat sodium in combination with intravenous erythromycin and ciprofloxacin]. AB - A 64-year-old man was referred to us because of pneumonia refractory to panipenem/betamipron. His chest radiography showed patchy consolidations in the lower lobe of the right lung and in the middle field of the left lung, and severe hypoxia was present. He was diagnosed as having acute respiratory distress syndrome due to severe pneumonia, and was treated with pulse methylprednisolone and sivelestat sodium in combination with intravenous erythromycin and ciprofloxacin. The patient recovered with this treatment. Serological examination using blood samples collected on the 12th and 28th hospital days revealed elevation of anti-L. pneumophila serogroup I antibody. It is suggested that administration of methylprednisolone and sivelestat sodium in combination with intravenous erythromycin and ciprofloxacin in a case of severe Legionella pneumonia complicated with acute respiratory distress syndrome is effective, and may be of use in similar cases. PMID- 15651277 TI - [A case of solitary endobronchial metastasis of prostatic cancer]. AB - We report the case of a 67-year-old man found to have an abnormal chest shadow in February 2003, who was referred to our hospital in April 2003, and was admitted in July 2003 because of the expansion of this shadow. Chest radiography and CT on admission showed a small nodular shadow in the right S6. Bronchoscopy revealed a soft polypoid mass in the lumen of the right B6b ii, which was considered to be an endobronchial neoplasm. A CT scan performed as a routine screening was found to be abnormal and revealed a prostatic tumor that was diagnosed as adenocarcinoma by pathologic examinations of lung and prostate tissue. We confirmed the diagnosis of prostatic cancer and its endobronchial metastasis by immunohistological staining with prostate specific antigen (PSA). An endobronchial metastatic tumor arising from prostatic cancer is a rare phenomenon that appears on the chest radiograph as a solitary mass. PMID- 15651278 TI - [A case of primary adenocarcinoma of the trachea]. AB - Primary adenocarcinoma of the trachea is rare. We present a case that was successfully treated with radiotherapy followed by surgical resection. A 66-year old man was admitted to our hospital because of cough, sputum and dyspnea. A chest CT film showed a tracheal tumor, and transtracheal biopsy specimen showed well-differentiated papillary adenocarcinoma. Other examinations excluded the diagnosis of metastatic adenocarcinoma from other organs. Since stenosis of the trachea was rapidly progressive, a tracheal stent was inserted to alleviate the obstruction. After radiotherapy, the tumor size decreased, after which surgical resection was performed. No recurrence had been observed 26 months after the operation. PMID- 15651279 TI - [Intense 18F-fluorodeoxyglucose uptake caused by pulmonary Mycobacterium intracellulare infection]. AB - A 70-year-old woman was admitted to our hospital for the evaluation of a pulmonary nodule in the left S5 segment. On 18F-fluorodeoxyglucose positron emission tomography (18FDG-PET), the nodule showed substantial uptake of 18F fluorodeoxyglucose. Bronchoscopy was performed, but the cytology was negative. For a pathological diagnosis, a lung biopsy was carried out using video associated thoracoscopy. The biopsy specimen showed granuloma formation with multinuclear giant cells. An acid-fast bacteria culture of the specimen was positive for Mycobacterium intracellulare. An atypical mycobacterium infection should be considered as a possibility when the 18FDG-PET of patients with pulmonary nodules is interpreted. PMID- 15651280 TI - [A case of multiple pulmonary infarctions associated with Sjogren's syndrome and antipsychotic agent administration]. AB - A 56-year-old woman presented with complaints of general malaise and left chest pain. Chest radiography and CT scanning revealed multiple nodules and infiltrations in both lung fields. Her symptoms diminished and the extent of some of the lung shadows decreased spontaneously. However, since new shadows appeared later in other parts of the lung, she was admitted to our hospital on September 3. A transbronchial lung biopsy was not adequate for diagnosing a particular disease. But thoracoscopic lung biopsy specimens revealed necrosis with localized pleural fibrosis, and so a diagnosis of pulmonary infarction was made. The patient did not have any underlying disease or coagulation abnormalities, but Sjogren's syndrome and an antipsychotic agent were suspected to be background factors. PMID- 15651281 TI - [Progress in the basic and clinical aspects of Parkinson's disease]. AB - We report a review on progress in the etiology and pathogenesis of Parkinson's disease (PD). We also report the long-term prognosis of PD patients seen in our clinic. Modern research on the pathogenesis started after the discovery of MPTP. We found inhibition of mitochondrial complex I by MPTP and MPP+. Mitochondrial respiratory failure induces oxidative damage to high molecular weight substances. Both mitochondrial failure and oxidative stress are important triggers of apoptosis. We found TUNEL positive nigral neurons in PD patients suggesting involvement of apoptosis in the pathogenesis. Interaction of genetic risk factors and environmental neurotoxins has been implicated in the etiology of PD. While we were investigating MnSOD gene polymorphism in PD patients, we found a young onset autosomal recessive PD family that was linked to the MnSOD locus. Subsequent linkage analysis on 13 families of young onset autosomal recessive families disclosed the linkage of this disease to the telomeric region of the long arm of chromosome 6 (6q25.2-27). Then we were lucky enough to find a patient who had a deletion of one of the microsatellite markers (D6S305) that we were using in the linkage analysis. We thought this marker might be located within the disease gene and this was the case. We screened the Keio BAC library with this marker, and eventually we cloned a novel gene encompassing 1.4 Mb; we named it parkin. The coding region consisted of 1,395 base pairs. The parkin protein had an unique sequence in that there was a 30% homology in the amino terminal region and two RING-finger motives on the carboxy terminal side. This unique structure suggested that the parkin protein was related to the ubiquitin-proteasome system. Parkin protein turned out to be an ubiquitin-protein ligase. Numbers of parkin interacting proteins were reported in the literature and accumulation of parkin substrates is likely to be the cause for the nigral neuronal death in this familial PD. Regarding the prognosis of PD, we analyzed the patients who visited our clinic from January 1, 1989 to December 31, 2002. The total of patients recruited was 1,772. The average age of onset was 57.2 years. Mean levodopa dose at the final examination was 479 mg/day. The most common initial symptom was tremor which was seen in 51% of the patients. Total percentage of patients who had tremor during the course of the disease was 75%. Long-term prognosis was evaluated on a subgroup of the patients who visited our clinic within 5 years from the onset and Hoehn and Yahr stage III or less when first seen. Analysis was done by the Kaplan-Meier survival curve. Percentages of patients who reached Hoehn and Yahr III 5, 10, and 15 years after the onset were 24%, 46%, and 65%, respectively. Percentages of patients who developed wearing off fluctuations were 5, 10, and 15 years after the start of levodopa were 18%, 46%, and 55%, respectively. Overall mortality on the total investigated patients was 7.9%. When compared to the age at death of Japanese population, mortality of men PD patients became very close to that of the general population in the year 2003. However, that in women PD patients showed significantly shorter survival compared to Japanese female population. Average ages of onset and the death were essentially similar between men and women PD patients. Survival curves to reach stage III and wearing off showed slightly but significantly faster time courses for women compared to those of men. This was an unexpected observation and its mechanism was discussed. It is our conclusion that overall prognosis of PD patients is improving and both patients and treating physicians should take an optimistic attitude to the disease. PMID- 15651282 TI - [Primary prevention of thrombosis for patients with atrial fibrillation]. AB - Clinical importance of antithrombotic prophylaxis strategy for patients with atrial fibrillation has been focused because of the high prevalence of severe cerebral embolism especially in elderly patients. Several multicenter, randomized clinical trials for prevention of thromboemblolism performed in the Western countries have revealed efficacy of the anticoagulation therapy with warfarin over the anti-platelet aggregation therapy with aspirin. Therapeutic guideline for thromboembolism in patients with atrial fibrillation has been established based on these observations. However, special attentions should be paid for applying the guideline for Japanese patients since bleeding risk in antithrombotic treatment seems to be high in Japanese. Recently, therapeutic guideline for thromboembolism in Japanese patients with atrial fibrillation has been established. In this guideline, treatment with warfarin is also recommended for patients with atrial fibrillation. However treatment target of warfarin, ie, PT-INR, is designated rather low in Japanese patients, especially in eldely patients over 75 years old. Usage of antiplatelet treatment with aspirin is also described in this guideline for Japanese, however, the evidence suggesting the usefulness for prophylaxis with aspirin is not sufficiently obtained in Japanese patients. Further clinical studies for antithrombotic treatment are needed for Japanese patients with atrial fibrillation. PMID- 15651283 TI - [Therapeutic application of cell transplantation and increased neurogenesis in cerebral infarction]. AB - Cerebral ischemia often results in neuronal loss, leading to the neurological deficits in stroke patients. To obtain the functional recovery after stroke, cell transplantation and enhancement of endogenous neurogenesis may have potential application. Recent evidence has demonstrated that neural stem cells exist in the adult mammalian brain. After cerebral ischemia, newly born neurons were found not only in hippocampal dentate and olfactory bulb but also in hippocampal CA1 and striatum, where neurons were lost after ischemia. Administration of neurotrophic factors or genes encoding them into the lateral venticule could enhance endogenous neurogenesis in experimental ischemia model. Furthermore, we have recently developed non-invasive gene transfer into macrophages infiltrating an infarct to stimulate proliferation of neural stem cells in cerebral infarction. Several strategies including gene therapy and pharmacological approach will be tried in stroke patients in near future. However, it remains unclear whether the number of new-born neurons from endogenous neural stem cells is sufficient for replacement of damaged neurons. Cell transplantation will have the advantage of preparing the large amount of transplanted cells. Human neural stem cells, embryonic stem cells and bone marrow-derived cells will be donor cells in stroke patients. Surprisingly, neuron-like cells derived from human teratoma cell line were already applied in stroke patients. However, ethical aspect will have to be discussed carefully before cells from other individuals are used as donor cells in stroke patients. PMID- 15651284 TI - [Consensus and controversy in the treatment of ischemic cerebrovascular diseases]. AB - Surgical and endovascular revascularization for ischemic cerebrovascular diseases (CVD) should be strictly indicated based on medical treatment. In this report, we describe current consensus and controversy in the treatment of ischemic CVD, and perspectives. 1) Local intra-arterial fibrinolytic therapy for acute cerebral embolism; intra-venous t-PA can be beneficial when given within 3 hours of stroke onset (NINDS), but many patients present later after stroke onset and alternative treatments are needed. Despite an increased frequency intracranial hemorrhage, treatment with intra-arterial proUK within 6 hours for MCA occlusion significantly improved clinical outcome at 90 days (mRS 40% >25%, PROACT-II). MELT-Japan are going now and waiting for results. 2) Carotid stenting; Carotid angioplasty and stenting (CAS) has been proposed as an alternative to carotid endarterectomy (CEA) in those considered at high risk for CEA. SAPPHIRE study confirmed CAS is an excellent option for patients with coexisting coronary artery disease, congestive heart failure, and other comorbid conditions that make them poor candidates for CEA. Now, CREST in USA and CSSA in Europe are going for randomized trial compared with CEA and CAS in any risk for CEA patients. 3) Stenting for intracranial arteries; Stroke rates in patients with symptomatic intracranial stenosis may be high on medical therapy. Although there is no clinical evidence and appropriate devices for intracranial vessels, it seems to be a potentially effective in the future. PMID- 15651285 TI - [Natural history and management of asymptomatic unruptured cerebral aneurysms]. AB - Natural history of asymptomatic unruptured cerebral aneurysms (AUCAs) still remains unclear. The concern on the natural history is incidence of rupture that was thrown into confusion with the report by International Study of Unruptured Intracranial Aneurysm Investigators (ISUIA) published on the New England Journal of Medicine in 1998. The annual rupture rate of AUCAs of our series was 1.1%. The study of Unruptured Cerebral Aneurysm Study of Japan (UCAS Japan) is now on going. Management modalities for AUCA includes surgery, intravascular treatment, and conservative treatment. Optimal treatment still remains under discussion. We first consider surgery for AUCAs, second intravascular treatment, and third conservative treatment. Our surgical indications are as follows: 1) patient's age is under 75 years old, 2) aneurysm locates in the anterior circulation, and 3) aneurysm size is over 5 mm. For patients with conservative treatment, periodic observation with magnetic resonance angiography is required. When aneurysm size increases with or without bleb, surgical or intravascular treatment should be considered. Our management strategy provides satisfactory results in patients with AUCAs. It is important to select the best treatment among those three modalities. In management of AUCAs, good surgical result (no mortality and low morbidity) are required for neurosurgeon. PMID- 15651286 TI - [Pathogenesis of Alzheimer's disease: implications from amyloid research front]. AB - Deposition of amyloid beta peptides as senile plaques is a hallmark lesion of Alzheimer's disease that is implicated in its pathogenesis. A beta is produced from amyloid precursor protein by sequential cleavages by beta- and gamma secretases. Gamma-secretase is a membrane protease complex harboring presenilin as a catalytic subunit. Recent studies revealed how presenilin is assembled with its cofactor proteins and acquires the gamma-secretase activity: Aph-1 and nicastrin initially form a subcomplex to bind and stabilize presenilin, and then Pen-2 confers the gamma-secretase activity and facilitates endoproteolysis of presenilin. Understanding the mechanism of gamma-secretase cleavage will help to clarify how intercellular cell signaling through transmembrane proteins are regulated by intramembrane proteolysis, and eventually to cure Alzheimer's disease by inhibiting secretases. PMID- 15651287 TI - [Biomarkers of Alzheimer's disease]. AB - Recent researches of CSF biomarkers, such as combinations of tau, A beta40, A beta42, are now establishing clinical evidence. These specific biomarkers should be used in clinical practice. To detect incipient AD form MCI and very early AD, these CSF biomarkers are quite useful. Newly developing biomarkers, such as phosphorylated tau and plasma A beta, must be established by large scale prospective study and autopsy-correlations. Thus, the contribution of these biomarkers is desired for diagnosis and clinical practice for Alzheimer's disease. PMID- 15651288 TI - [Towards presymptomatic diagnosis, prevention and treatment of Alzheimer's disease]. AB - Alzheimer's disease (AD) is a progressive neurodegenerative disorder, the pathology of which becomes irreversible as it proceeds downstream of the disease cascade. Therefore, overcoming AD primarily requires early (presymptomatic) diagnosis, followed by preventive treatment. Since accumulation of amyloid beta peptide (A beta) in brain seems to play a central role AD pathogenesis, we established a new imaging technique to visualize A beta plaques in a mouse model of A beta amyloidosis in a non-invasive manner using a high-power MRI. This will open a new avenue to search for biochemical markers that correlate with the pathological parameters. We also found that a dimeric form of A beta, the quantity of which can be metabolically regulated by neprilysin, impairs in vivo neuronal plasticity, i.e. hippocampal long term potentiation. This suggests that reducing A beta dimers by upregulating neprilysin activity is likely to contribute to alleviation of memory-associated symptoms. Finally, we discovered that a neuropeptide, somatostatin, upregulates neuronal neprilysin activity. Because brain somatostatin expression is known to decline during aging, the finding indicates that the aging-induced downregulation of somatostatin may be a trigger for A beta accumulation leading to late-onset sporadic AD development and that somatostatin receptor(s) now emerge as pharmacological target candidates for the prevention and treatment of AD. PMID- 15651289 TI - [Treatment of Alzheimer disease: A beta vaccine]. AB - Alzheimer disease (AD) is regarded as an amyloidosis of the brain, and, therefore, the prevention/deletion of beta amyloid deposition is one of the most promising target of the treatment. Particularly, immune-mediated strategy is now known as A beta vaccination, which is thought to be a highly feasible way being applicable to AD patients. The A beta vaccination was originated by Schenk et al. in 1999. Since active and passive immunization of amyloid precursor protein (APP) gene- transgenic mice showed significant reduction of beta amyloid deposits in the brain and the immunized mice showed improvement in cognitive functions, clinical trials were performed in US and Europe. However, the trial was suspended, because about 6% of patients who received the vaccine developed meningoencephalitis probably mediated by T cells reactive to A beta. Although the trial was halted, an autopsy case who had had the meningoencephalitis suggested the disappearance of senile plaques (Nicoll et al., 2003). Moreover, patients who developed antibodies that recognize senile plaques by immunohistochemistry showed significantly milder decline of cognitive functions than those who did not (Hock et al., 2003). Here, we report a safe and effective oral vaccine using adeno associated virus vector carrying A beta cDNA tested in tg2576 mice. PMID- 15651290 TI - [Autosomal dominant spinocerebellar degeneration--new forms and pathomechanisms]. AB - In our country, hereditary spinocerebellar degeneration accounted for approximately 30% of the total cases. Most of them are autosomal dominant and include more than 20 diseases. The outlines of some new members, namely autosomal dominant cortical cerebellar atrophy linked to chromosome 16 (16q-ADCCA), SCA14, an ataxia caused by FGF14 mutation and a form of neuroferritinopathy were described. The etiology of many autosomal dominant SCDs has been identified as the abnormal expansion of CAG repeat. The latter three diseases are caused by missense mutations of the causative genes, which clearly shows the presence of other new mechanisms of cerebellar degeneration than repeat expansion. 16q-ADCCA is the most frequent after Machado-Joseph disease and SCA6 according to our genetic diagnosis of 185 SCD patients. The disease is characterized by Purkinje cell degeneration and atrophy with somatic sprouts as well as the halo-like structure surrounding the soma. The halo is positive for synaptophysin. These features are so unique that 16q-ADCCA may be diagnosed by neuropathology alone. PMID- 15651291 TI - [Clinical features and molecular genetics of autosomal recessive spinocerebellar degenerations]. AB - The number of patients with spinocerebellar degeneration (SCD) has recently exceeds 20,000 in Japan. Among them, sporadic form is the most common form (67.2%). Among the hereditary forms of SCD, autosomal dominant (AD) form comprises 27.0%, while autosomal recessive (AR) form is rare (1.8%). Because of the rare occurrence of AR-SCD, the molecular genetic studies have been difficult to conduct. Recent progresses in molecular genetics, however, have enabled identification of causative genes for the majority of AR-SCD. Although Friedreich's ataxia is the most representative form of AR-SCD, patients with molecular diagnosis of Friedreich's ataxia have not been described in the Japanese population. Among the various forms of AR-SCD, early-onset ataxia with ocular motor apraxia and hypoalbuminemia (EAOH) seems to be the most common form in the Japanese population. Aprataxin, the causative gene for EAOH, has been suggested to play a role in the single strand DNA break repair. Interestingly, abnormalities in DNA break repair processes have been implicated in several forms of AR-SCD including AOA2, SCAN1 and ataxia telangiectasia. In this group of AR SCD, cerebellar atrophy is more marked compared to that observed in Friedreich's ataxia. Taken together, abnormalities in DNA break repair processes may play an essential role in cerebellar degeneration in this group of AR-SCD. PMID- 15651292 TI - [Amyotrophic lateral sclerosis: recent insights from transgenic animal models with SOD1 mutations]. AB - Mutations in Cu/Zn superoxide dismutase (SOD1) have been linked to some familial cases of amyotrophic lateral sclerosis (ALS). In order to reproduce the different degree of toxicity to the mutant protein by mutations, we generated new transgenic mice with two mutations from which the progression of the disease in human family is rapid (L84V) or extremely slow (H46R). By comparing the two transgenic mice with different SOD1 mutations, we demonstrate that the time course and the first symptoms in these mice were likely to human SOD1-mediated familial ALS. In addition, we report here that rats that express a human SOD1 transgene with two different ALS-associated mutations (G93A and H46R) develop striking motor neuron degeneration and paralysis. The larger size of this rat model as compared with the ALS mice will facilitate studies involving manipulations of spinal fluid (implantation of intrathecal catheters for chronic therapeutic studies; CSF sampling) and spinal cord (e.g., direct administration of viral- and cell-mediated therapies). Using this rat model we showed that intrathecal administration of the hepatocyte growth factor attenuates motoneuron death and prolongs the duration of the disease of transgenic rats. PMID- 15651293 TI - [Recessive motor neuron diseases: mutations in the ALS2 gene and molecular pathogenesis for the upper motor neurodegeneration]. AB - We have initially identified a mutation in ALS2 as a causative for a juvenile autosomal recessive form of amyotrophic lateral sclerosis (ALS), termed ALS2 (OMIM 205100). ALS2 mutations also are causative for an autosomal recessive juvenile primary lateral sclerosis, and infantile-ascending hereditary spastic paralysis. To date, nine homozygous ALS2 mutaions from nine independent families have been identified. All of these mutations result in predicted premature translation termination caused by the recessive frameshift or nonsense mutation. ALS2 is a 184-kD protein comprising several putative guanine nucleotide exchange factor (GEF) domains [RLD; RCC1 like domain, DH. PH domain, VPS9; Vacuolar protein sorting 9 domain]. In vitro, ALS2 specifically binds to the small GTPase Rab5 and functions as a GEF for Rab5. Ectopic expression of full-length ALS2 has further implied an association with endosomal membranes mediated by the VPS9 domain, consistent with ALS2 involvement in endosomal trafficking and fusion in conjunction with the activation of Rab5. These results combined with our findings suggest that an obstruction of endosomal dynamics might underlie neuronal dysfunction and degeneration in ALS2, PLSJ, and HSP, as well as in a number of other motor neuron diseases. PMID- 15651294 TI - [Multiple sclerosis: treatment and prevention of relapses and progression in multiple sclerosis]. AB - The benefit of interferon beta-1a and 1b treatment on relapsing remitting multiple sclerosis (MS) is now firmly established. IFN beta-1b, the only disease modifying drug approved in Japan, was comparably effective in OS-MS and C-MS in a Japanese treatment trial. Although the effect of Copaxone is less conclusive, superior safety profiles make this treatment an important alternative choice. Mitoxantrone has been approved by FDA for rapidly progressing patients. The usage should be restricted for two or three years to avoid cardiac side effects. Antegren, which is an anti-cell adhesion molecule monoclonal antibody and has been shown to be very effective for relapsing MS patients, now awaits approval by FDA. Many combination therapies are currently under systematic survey and hoped to provide superior efficacy. PMID- 15651295 TI - [Limbic encephalitis: etiology, pathogenesis, diagnosis and therapy]. AB - Limbic encephalitis affects the mesial temporal lobes and is characterized by subacute onset of memory impairment, personal change, temporal seizures and autonomic nervous disorders. It can occur as viral infections, especially caused by Herpes simplex, paraneoplastic syndrome as a remote effect of cancer, CNS complication of well defined autoimmune diseases. Recently acute reversible limbic encephalitis which probably autoimmune mediated disorders has been reported. Anti-immunotherapy including steroids, plasma exchange and intravenous immunoglobulin often improves this condition and anti-voltage gated potassium channel (VGKC) antibody or anti-glutamate receptor epsilon2 antibody has been detected in some patients. Establishing the means of early detection of these antibodies as well as other characteristic paraneoplastic antibodies should now be the aim. Detection of neurotropic viral genomes in CSF by PCR is also important for differential diagnosis. As complete recovery of higher cerebral function is generally difficult, immunotherapy and anti-convulsants in addition to vitamin B, and acyclovir should be considered in an early stage of disease. PMID- 15651296 TI - [Bickerstaff's brainstem encephalitis and Fisher syndrome: their relationship and treatment]. AB - The nosological position of Bickerstaff's brainstem encephalitis (BBE) was not eatablished, and its etiology was not clear until 1993. Because anti-GQ1b IgG antibody frequently occurs in patients with Fisher syndrome (FS) and there are clinical similarities between FS and BBE, we investigated anti-ganglioside antibodies in sera from three patients with BBE. High anti-GQ1b IgG antibody titers were present in their sera, but decreased with the clinical course of the illness. The findings that BBE and FS shared common autoantibody suggest that autoimmune mechanism common to FS is likely in BBE, and that both conditions form a continuous spectrum. To clarify the clinical features of BBE, we reviewed detailed clinical profiles and laboratory findings for 62 cases of BBE diagnosed by the strict criteria of progressive, relatively symmetric external ophthalmoplegia and ataxia by 4 weeks, and disturbance of consciousness or hyperreflexia. A striking feature was the association with flaccid symmetric tetraparesis seen in 60%. Limb weakness in the BBE cases studied was considered the result of overlap by the axonal subtype of Guillain-Barre syndrome. These indicates that the two disorders are closely related and form a continuous spectrum, and that treatment for Guillain-Barre syndrome should be applied for BBE. PMID- 15651297 TI - [Isaacs' syndrome, stiff person syndrome and Satoyoshi disease: pathomechanisms and treatment]. AB - Neurological disorders with characteristic clinical manifestations of painful muscle cramp and stiffness are not infrequent. The immune-mediated mechanism with specific antibodies among these diseases is particularly important for treatment. Isaacs' syndrome (acquired neuromyotonia) is an antibody-mediated potassium channelopathy. The suppression of voltage-gated potassium channel (VGKC) by antibodies induces peripheral nerve hyperexcitability. Antibodies may decrease VGKC density by cross-linking F (ab)2 fractions and increasing the degradation rate of VGKCs. Stiff person syndrome (SPS) and its variants show characteristic symptoms and signs of central nervous system hyperexcitability due to antibodies to the GABA-ergic system such as glutamic acid decarboxylase (GAD), amphiphysin 1 and gephyrin. The role of GAD is the subject of debate, however, recent studies reveal the intrathecal synthesis of GAD which is specific for SPS and appears to impair GABA synthesis. Satoyoshi disease is characterized by painful muscle cramp, baldness, intractable diarrhea, bone and joint deformity, and endocrine disturbances. Muscle cramp may be due to inhibition of the spinal interneuron and hyperexciatability of the anterior horn cell. In patients with Satoyoshi disease, sera reacted with an 85 kDa protein of human brain lysate. In all these disorders, suppression or removal of specific antibodies is critical, however, the effects are short-lived, and supplemental treatment to reduce the hyperexcitability of the peripheral or central nervous system will be needed. PMID- 15651298 TI - [Mechanism based prevention and treatment of migraine]. AB - Triptan therapy results in good relief of headache in 68% of the patients. For many of the patients with migraine, triptan provides complete pain relief in some attacks but not in others. A recent theory proposes that allodynia (pain sensitization) develops in the brain during migraine to increase the intensity of headache significantly. The presence of cutaneous allodynia, an exaggerated painful sensation resulting from a no noxious stimulus to normal skin, is reported in more than 70% of the patients. In our prospective study with 41 patients with migraine, 54.2% presented with skin allodynia. Triptan was effective in 77% of the patients without allodynia which was higher than the base line efficacy rate of 68% in all the patients. Our data support the hypothesis that the development and maintenance of cutaneous allodynia is propelled by sensitization of central trigeminal nucleus. It is suggested that the pain-free outcome increases drastically if triptan therapy is given before the phenomenon of allodynia develops during the attacks of migraine. PMID- 15651299 TI - [Cluster headache and other trigeminal autonomic cephalalgias: diagnosis and treatment]. AB - Cluster headaches are characterized by strictly unilateral paroxysmal attacks of severe pain with associated autonomic sign and symptom. Prevalence is 5 times higher in men than in women in our cases. About 10-15% of patients have chronic symptoms without remissions, but we estimated less frequent in Japanese (6.6% in our series). Pain almost invariably recurs on the same side, but in some patients (16.4%) the affected site switches. Cluster headache may be inherited in about 5% of our cases. Attacks frequently occur at night (60.7%). The patients (64.8%) are restless or agitated during an attack. Recent PET studies elucidated that acute attacks causes activation of the posterior hypothalamic grey matter. The excitement of the area might be responsible for peculiar clinical characteristics of agitation. Some patients (66.0%) have also have symptoms (especially a visual aura) usually attributed to migraine. Treatment of cluster headache includes both acute therapy aimed at aborting individual attacks and prophylactic therapy aimed at preventing recurrent attacks during the cluster period. There are many choices using for both therapies. Based on our clinical experience, we recommended the combination of nasal sumatriptan for acute attacks and verapamil 240 mg/day for prophylaxis. PMID- 15651300 TI - [Chronic daily headache and medication-induced headache]. AB - Chronic daily headache (CDH) and medication-induced headache are refractory headaches to treatment. I evaluate the classification and pharmacological treatment of CDH and medication-induced headache. The subjects of this study are 508 patients with CDH (out of 2,241 patients with chronic headache who were examined and treated in Toyonaka Municipal Hospital for six years from November 1997 to October 2003). CDH was classified into three types of chronic migraine in 232 cases (45.7%), chronic tension-type headache in 223 cases (43.9%) and new daily-persistent headache in 53 cases (10.4%). 245 cases overused medication (analgesic in 230 cases, ergotamine in eight cases and triptan in seven cases). 108 cases abused medication (analgesic in 95 cases, ergotamine in seven cases and triptan in six cases). For treatment, muscle relaxants, calcium-channel blockers, tricyclic antidepressants, selective serotonin reuptake inhibitors, antiepleptic drugs and leukotriene receptor antagonists were administered to 231 patients for two months from September to October 2003. Awareness of CDH and medication induced headache is important to physicians who treat patients with headache. PMID- 15651301 TI - [Headache originating front-orbital area]. AB - Headache originating front-orbital area can be divided to (1) Which has no autonomic symptoms such as lacrimation, rhinorrea, rhinostasis. This include psychogenic headache and epileptic headache. In the case of psychogenic headache, pericranial tenderness is not observed, and headache is medium in intensity. Most often patient complains of a headache originating frontal area. There are more than five various symptoms such as general malaise, numbness, tingling sensation, vertigo, sleeplessness. However, although symptoms are multiple, patients spend a life commonly. In other words, a patient is protected by a headache against his or her stress. No medication is needed in such a case. In epileptic headache, pressing type pain is felt over the forehead for several minutes to a few hours. Tremor or convulsion sometimes follow the headache. EEG shows spike and wave activities. In the case of focal epilepsy, headache occurs contralateral to the focus. Anti-epileptic drugs such as VPA or CBZ is a choice in such case, and headache as well as seizure disappears. (2) Front-orbital headache with autonomic symptoms include various trigeminal autonomic cephalalgias. These include cluster headache, episodic paroxysmal hemicrania, hemicrania continua, among others. Precise history taking is necessary for the treatment, because no drug is 100% effective. PMID- 15651302 TI - [The gene products for familial Parkinson's disease provide us hints to elucidate the mechanisms of nigral degeneration]. AB - In the majority of patients with Parkinson's disease (PD), it is now clear that genetic factors contribute to the pathogenesis of PD, although the contribution of genetic and environmental factors remained to be elucidated. Recently, several genes for familial PD (FPD) based on the single gene defects have been mapped and identified. Alpha-synuclein and UCH-L1 are involved in the dominant form of FPD. In contrast, parkin, DJ-1, and PINK1 are responsible for the recessive form of FPD. The presence of different loci or different causative genes indicates that PD is not a single entity but a highly heterogeneous disorder. However, the functions of causative genes may share common pathways such as an ubiquitin proteasome pathway, oxidative stress, and mitochondrial dysfunction. Thus, the identification and elucidation of the causative genes should enhance our understanding of the pathogenesis of not only FPD, but also sporadic PD. PMID- 15651303 TI - [Parkinson disease with dementia--responsible locus for dementia]. AB - Responsible locus for dementia in Parkinson disease (PD) was investigated. Serial 1,395 autopsy cases were studied for the combined pathology of PD and Alzheimer disease (AD). Following the one-year rule by the first Consensus Guidelines, definite AD pathology was quite rare in PD with dementia (PDD) but common in dementia with Lewy bodies (DLB) . Plaque-dominant senile changes apparently enhanced neocortical Lewy-body pathology in both the conditions. About the hypometabolism in the visual cortex of PDD, a 66-year old man presented with fluctuation in hallucination commensurate with fluctuating hypometabolism. Considering the paucity in pathological changes of the visual cortex, this hypometabolism may represent functional impairment in the fiber connection. Comparative pathological studies with PD and PDD were carried out. Only one case of a 48-year-old woman, who unexpectedly died of heart failure, was free from cognitive decline, and did not show limbic and neocortical involvement. Another case of a 75-year old man with MCI presented with the similar pathology. All other cases showed clinical documentation of cognitive impairment and limbic and neocortical pathological involvement. Thus, the combination of prospective clinical and radiological studies and retrospective pathological studies (dynamic neuropathology) may be essential to investigate a role of the basal-forebrain cholinergic system. PMID- 15651304 TI - [A useful marker for differential diagnosis of Parkinson's disease--MIBG myocardial scintigraphy]. AB - We performed [123I]-meta-iodobenzylguanidine (MIBG) myocardial scintigraphy in 180 patients with PD, 24 patients with DLB and 73 patients with the other related diseases. The heart-to-mediastinum (H/M) ratio in PD and DLB was significantly low compared to that in MSA, PSP, CBD and AD. The H/M ratio tended to decrease with the disease progression in PD. These findings suggest that MIBG myocardial scintigraphy could be a marker for differential diagnosis of PD and DLB. We immunohistochemically examined heart tissues from patients with pathologically confirmed PD, DLB and the other related diseases using monoclonal antibody against tyrosine hydroxylase (TH). The number of TH-immunoreactive nerve fibers in the epicardium was moderate or abundant in most patients with MSA, PSP, CBD and AD as well as in the control subjects. Contrarily, the number of TH immunoreactive nerve fibers in the epicardium had almost completely disappeared in nearly all the patients with PD and DLB. These findings suggest that postganglionic cardiac sympathetic nerve denervation occurs in PD and DLB but not in MSA, PSP, CBD and AD, which accounts for the decreased cardiac uptake of MIBG in PD and DLB, but not in the other diseases. PMID- 15651305 TI - [Comment on guidelines for treatment of Parkinson's disease]. AB - The guidelines for the therapy of Parkinson's disease (PD) based on the concept of evidence-based medicine (EBM) were edited by the Japanese Society of Neurology in 2002. The author described his impression of these guidelines from a user's viewpoint after using these guidelines. In the chapter on the therapeutic efficacy and safety of drugs, compared with other antiparkinsonian drugs, levodopa seems to be only briefly described. Because the therapy of PD highly depends on the effect of levodopa, a more detailed description of levodopa will be helpful for neurologists. In the chapter on general and detailed discussions, the use of a dopamine agonist (DA) for young PD patients is recommended because DA prevents motor complications; this is the most emphasized part based on the concept of EBM in the guidelines. Measures of managing various complications based on various lines of evidence and experiences of members of Ad hoc committee for the treatment of PD are well documented in this chapter. In general, since the guidelines are technical ones, they clearly describe how to treat PD patients. For the effective use of these guidelines, neurologists should have the ability to correctly determine PD symptoms, some knowledge about the pharmacology of antiparkinsonian drugs particularly levodopa and DA, and balanced thinking regarding the management of PD patients. PMID- 15651306 TI - [How to examine a patient with higher cortical dysfunction at bedside]. AB - In practice, a neuropsychological evaluation may serve one or more purpose. The first systematic applications of neuropsychological assessment dealt with diagnosis. Before the days of sophisticated neuroimaging, the localization of a cerebral lesion was neuropsychology's most important function. Recent developments in neuroradiological techniques have greatly reduced the contributions of neuropsychological assessment to diagnosis and lesion localization. Today, neuropsychological assessment is most usually called upon for the detailed behavioral description necessary for intelligent patient care, for rational treatment, and for appropriate rehabilitation training. When neurologists examines patients with higher cortical dysfunction, they should bear the following points in mind. At first, good clinical history often holds the key to diagnosis. This is especially true in the neurologic and the neuropsychologic history. The examiners should not hesitate to ask the patient history before and during the neuropsychological examinations. Secondly sometimes the patients with neuropsychological dysfunction do not recognize their disturbances. For example, the patients with apraxia or optic ataxia usually do not complain their disturbances. Only by storing the range of neuropsychological dysfunction in their mind can neurologists recognize these behavioral disturbances. Thirdly in neurology when we localize lesions of the nervous system, it is helpful to think about the major syndrome supervene with lesions at different anatomical levels, from the muscle to the cortex. Also in neuropsychology, for example, when we localize lesions of with speech disturbance it is useful to think about from the simplest level (the muscle) to the cortex. At last test in neuropsychological assessment can be used to establish a comparison standard--i.e. for estimating premorbid ability. The Wechsler test (WAIS-R) is one of the most frequently used measures in neuropsychological batteries. It is a core instrument, giving information about the overall level of intellectual functioning disability, and providing clues to altered functions. PMID- 15651307 TI - [Symptomatology of aphasia]. AB - Recently, some standard test batteries such as Boston Diagnostic Aphasia Examination and Western Aphasia Battery are frequently used to assess language abnormalities and to identify individual types of aphasia. Aphasia is, however, a neurologic disorder and differential diagnosis of the type should be made naturally in the neurological examination. Characteristics of verbal output are most important clinically for the classification. The table shows core symptoms of classical aphasic types. The figure presents a systematic approach to aphasia diagnosis based on several different patterns of aphasic output. PMID- 15651308 TI - [Cognitive neuropsychological investigation of apraxia]. AB - Apraxia is a higher-order motor disturbance of goal-directed behavior characterized by an inability to perform skilled, previously learned movements in the absence of weakness and sensory defects. Clinical findings associated apraxia are divided into two large domains, production and conceptual. In this study, we reports two severely apraxic patients, case TT and UH, whose performances suggests that production and conceptual domains may be dissociable. Case TT is a 67-year-old, right-handed man who showed content errors on actual tool use, gesture production to command and gesture to sight of tool after a left inferior parietal infarction. He demonstrated severely impaired conceptual knowledge on tool-action and tool-object associative tests with intact recognition of many common tools. On the other hand, case UH, who is 79-year-old, mixed-handed man, had spatial and movement errors on gesture production to command and gesture to sight following a right fronto-parietal infarction. His performances on tests of conceptual praxis knowledge were intact. Gesture recognition was preserved in both cases. The selective impairment of conceptual praxis knowledge and action production in two cases supports the notion that production and conceptual systems are distinct features of the representation of object-oriented action. PMID- 15651309 TI - [Symptoms and lesion localization in visual agnosia]. AB - There are two cortical visual processing streams, the ventral and dorsal stream. The ventral visual stream plays the major role in constructing our perceptual representation of the visual world and the objects within it. Disturbance of visual processing at any stage of the ventral stream could result in impairment of visual recognition. Thus we need systematic investigations to diagnose visual agnosia and its type. Two types of category-selective visual agnosia, prosopagnosia and landmark agnosia, are different from others in that patients could recognize a face as a face and buildings as buildings, but could not identify an individual person or building. Neuronal bases of prosopagnosia and landmark agnosia are distinct. Importance of the right fusiform gyrus for face recognition was confirmed by both clinical and neuroimaging studies. Landmark agnosia is related to lesions in the right parahippocampal gyrus. Enlarged lesions including both the right fusiform and parahippocampal gyri can result in prosopagnosia and landmark agnosia at the same time. Category non-selective visual agnosia is related to bilateral occipito-temporal lesions, which is in agreement with the results of neuroimaging studies that revealed activation of the bilateral occipito-temporal during object recognition tasks. PMID- 15651310 TI - [Bacterial meningitis: determination of pathogens and therapeutic management]. AB - Recent aspects of pathogen determination and therapeutic management are reviewed based on a clinical analysis of 50 of our adult patients with bacterial meningitis (BM). The sensitivity for detecting pathogens using gram-stained smears and cultures of CSF was high in untreated patients, but low in patients previously treated with antibiotics. Latex agglutination for antibodies of pathogens is rapid and has a potential for determining pathogens in partially treated meningitis. The PCR also has a potential for determining pathogens in pre treated, culture-negative cases and for detecting whether the bacterial pathogen is resistant or sensitive to antibiotics. The initial empiric regimen of antibiotics has been modified with elevation in the detection rate of drug resistant bacilli. The proportion of patients undergoing treatment with VCM and/or Carbapenems has recently increased at our department. A combination of dexamethasone under administration of antibiotics has also been established as effective in adult BM. On admission, some patients are difficult to diagnose as BM rather than herpes simplex encephalitis (HSVE). A serum CRP value of >2.0 mg/dl proved useful for such different diagnosis in our patients. The predictors of a poor outcome based on multivariate logistic analysis in our BM patients were level of unconsciousness and sepsis. PMID- 15651311 TI - [Newly discovered human herpesviruses: pathogenesis and treatments for central nervous system infection due to HHV-6]. AB - Human herpesvirus-6 (HHV-6) is the causative agent of the common childhood infectious disease, exanthem subitum. Soon after the virus was isolated from humans it was found to be closely related to human cytomegalovirus, and thus was classified within the beta subgroup of human herpesviruses. HHV-6 possesses neuro tropism in-vitro, and it has been suggested that primary infection can cause complications of the central nervous system (CNS), including febrile seizures and encephalitis/encephalopathy. In addition, this virus has recently been associated with limbic encephalitis, which occurred in mainly immunocompromised adult patients. It is proposed that direct invasion of the virus into the CNS may play an important role in causing these neurological complications. Although pathogenesis of HHV-6 encephalitis/encephalopathy remains unclear, it is thought that two different mechanisms such as direct invasion (primary encephalitis) and immune mediated impairment (secondary encephalitis) might play important role in causing CNS manifestations. It is possible that anti-viral drugs (ganciclovir and foscarnet) might be effective to patients with primary encephalitis due to HHV-6. Therefore, rapid diagnosis of active HHV-6 infection is crucial for patient management. PMID- 15651312 TI - [The neurology of HIV infection: clinical features of HIV encephalopathy and future problems in the HAART era]. AB - Since the introduction of highly active antiretroviral therapy (HAART), which has been available for most patients in developed world, HIV/AIDS has become a chronic disorder with dramatic reduction in mortality and morbidity due to HIV/AIDS and related diseases. However, from the nationwide survey of HIV-related neurological complications and analysis of our series of total 290 AIDS patients before and after HAART introduction, main neurological complications and opportunistic infections of AIDS patients remain unchanged after the introduction of HAART in Japan. It is crucial that we let a people know that the prevention and treatment of HIV infection is impotant. Recent epidemiological studies show that neurological complications still remain in an important cause of disability, although their relative incidence as their course and clinical presentation have been modified. It is expected that the clinical features of HIV encephalopathy will change particularly from severe subacute progressive dementia in the terminal stage of HIV infection into different patterns of dementia, for example slowly progressive neurological deficits occurring without immune deficiency under HAART. PMID- 15651313 TI - [Prion diseases: disease diversity and therapeutics]. AB - More than one hundred victims of iatrogenic CJD with cadaveric dura mater grafting have been recognized in Japan, and the people have been also exposed to a risk of outbreaks of variant CJD. These diseases are distinct from other forms of prion diseases as well as other neuropsychiatric disorders, but on an early clinical stage, their differential diagnoses from other atypical forms of prion diseases are not necessarily easy. Thus, atypical forms of prion diseases were overviewed and discussed here. In addition, data on recent clinical trials of enteral antimalarial drug (quinacrine or quinine) treatment or intracerebroventricular pentosan polysulfate treatment were presented and discussed, because research progress in the therapeutics for prion diseases has been remarkably made on the basis of the prevalence of those acquired forms of prion diseases. PMID- 15651314 TI - [Molecular genetics of epilepsy]. AB - Recent identifications of genes responsible for epilepsies are now contributing to diagnosis and treatment. Mutations of voltage-gated sodium channel genes SCN1A and SCN2A have been reported in epilepsies with a variety of phenotypes including generalized epilepsy with febrile seizures plus (GEFS +), severe myoclonic epilepsy in infancy (SMEI), intractable childhood epilepsy with generalized tonic clonic seizures (ICEGTC), and benign familial neonatal-infantile seizures (BFNIS). We also identified a sporadic nonsense mutation of SCN2A in a patient with intractable epilepsy with severe mental decline. Lafora's disease (LD) is a fatal autosomal recessive epilepsy characterized by stimuli sensitive myoclonus, grand mal seizures, and progressive intellectual and neurological deterioration. The EPM2A gene has been reported to be responsible for LD. We found multiple disease mutations of EPM2A in LD patients, and also identified a subclass of LD who shows an early onset cognitive defect and correlated with EPM2A exon 1 mutations. We reported that the laforin protein encoded by the EPM2A gene has a dual-specificity phosphatase activity, associates with polyribosome, and interacts with the HIRIP5 protein with NifU-like domain. We recently generated and reported the EPM2A KO mice those develop neurodegeneration and other features similar to those of LD patients. PMID- 15651315 TI - [Treatment guidelines for adult epilepsy]. AB - We reviewed the treatment strategies and choices of drugs for adult patients with epilepsy. As evidence from controlled studies for the older drugs is scarcely available from the literature, we added a clinical study performed in our hospital and an expert consensus study. Published randomized clinical studies revealed carbamazepine (CBZ) to be the treatment of choice, but provided little evidence for the differences in efficacy between phenytoin (PHT), valproate (VPA), phenobarbital (PB) and zonisamide (ZNS). The clinical study in our patients with intractable localization-related epilepsy indicated the superior efficacy of CBZ and PHT over VPA or ZNS. Sixty-nine experts converged on the opinion that monotherapy should be started with VPA for seizures of both idiopathic and symptomatic generalized epilepsy, and with CBZ for seizures of localization-related epilepsy. When the trials with 2-3 monotherapy failed, surgery should be considered for localization-related epilepsy. We proposed practical guidelines for treatment of patients with adult epilepsy, including rehabilitation and care for improvement of quality of life. PMID- 15651316 TI - [Social aspects of epilepsy: marriage, pregnancy, driving, antiepileptic drug withdrawal and against social stigma]. AB - Persons with epilepsy need adequate advice and effective counselling about issues such as marriage, pregnancy, risks of inheriting epilepsy, driving, employment and antiepileptic drug withdrawal, because these persons are not receiving important information and education about their condition and possible adverse effects of treatment. Furthermore, women with epilepsy have increased rates of pregnancy complications and poor fetal outcomes including congenital malformations and developmental delay related to both their epilepsy and antiepileptic drugs. However, approximately 90% of all women with epilepsy undergo normal pregnancy and give birth to children free of birth defects. Pregnancy is generally safe in women with epilepsy. The study of long-term prognosis of childhood-onset epilepsy in Japan shows that the majority of these patients have lower levels of educational background as well as employment and marital status compared with the general population (Wakamoto H. et al). Of patients with epilepsy, 60% to 70% achieve control with antiepileptic medication. However, several antiepileptic drug withdrawal studies show variable rates of success, with relapse rates ranging from 12% to 63% (Britton J.W.). Driving is listed as major problem in persons with epilepsy. However, the patients with seizure-free more than two years have been able to get the driver's license since June, 2002. Social attitudes towards epilepsy cause more distress to the patient than the disease itself. We should realize that persons with epilepsy are normal or near-normal. To ameliorate the social stigma against epilepsy, continuous and repetitive educational efforts would be needed. PMID- 15651317 TI - [Surgical treatment for temporal lobe epilepsy with preservation of postoperative memory function]. AB - Temporal lobe epilepsy is frequently drug-resistant in adult epilepsy. However, its surgical treatment is very effective and about 70% of operated patients become seizure-free. As preoperative evaluation, analysis of seizure characteristics, scalp EEG with sphenoidal lead, and neuroimaging are most important. When these noninvasive examinations are concordant in focus localization, surgical treatment without prolonged recording by intracranial electrodes become feasible. In spite of good seizure outcome after temporal lobectomy, postoperative impairment of verbal memory has remained to be a long standing problem. To cope with this ominous complication, we developed a new surgical method of hippocampal transection, based on the principle of multiple subpial transection (MST). Placing a small corticotomy on the anterior part of superior temporal gyrus, the inferior ventricle is opened through suctioning the temporal stem. Intraoperative electrocorticography (ECoG) is recorded over the hippocampus. After the extent of the epileptic area is determined, the alveus is cut with microscissors and the pyramidal layer is transected in parallel with neuronal fibers by a specially designed ring transector. With this procedure, epileptic discharges from the hippocampus can be completely abolished. In all 7 cases undergoing left hippocampal transection, postoperative verbal memory was preserved with excellent seizure outcome. PMID- 15651318 TI - [Frontotemporal dementia and frontotemporal degeneration--how to define?]. AB - The term of the frontotemporal dementia was first proposed by Lund and Manchester group in 1994, but the definition of frontotemporal dementia has been still controversial. Frontotemporal dementia is caused by several diseases which have fronto-temporal atrophy. The diseases are collectedly designated as frontotempoal degeneration. The frontotemporal degeneration encompasses several diseases such as Pick disease (frontotemporal degeneration with Pick bodies) and frontotemporal degeneration with ubiquitin-positive inclusions and frontotemporal degeneration (no inclusion bodies are observed). Pick bodies are consisted of abnormally phosphorylated tau protein. The recent discoveries of tau mutations in frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17) suggest important role of tau abnormalities in the disease mechanism. The frontotemporal degeneration has also another clinical phenotype such as slowly progressive aphasia. Slowly progressive aphasia has subtypes of non-fluent aphasia and semantic aphasia. Some patients with corticobasal degeneration or progressive supranuclear palsy also reveal the clinical pictures of frontotemporal dementia or slowly progressive aphasia and should be considered as differential diagnosis in the patients with frontotemporal dementia or slowy progressive aphasia. PMID- 15651319 TI - [Clinical, genetic and pathological aspects of frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17)]. AB - Frontotemporal dementia and parkinsonism linked to chromosome 17 (FTDP-17) is a group of hereditary, adult-onset, progressive neurodegenerative syndromes, which lead to the accumulation of intracellular deposits of hyperphosphorylated tau protein. Since the original definition of FTDP-17 in the Consensus Conference held in Ann Arbor, Michigan in 1996, it has become apparent that this syndrome has worldwide distribution. More than 80 families have been described in North America, Europe, Australia and Asia. The molecular genetic studies have identified 35 different mutations outside and on exon 10 of tau gene. The symptomatic onset of FTDP-17 is usually insidious. The clinical phenotypes are characterized by behavioral, cognitive and motor disturbances that may occur in various combinations and in varying degrees of severity. Affected individuals develop a constellation of signs, including at least two of the three cardinal manifestations of FTDP-17. It should be noted that there is significant clinical phenotypic heterogeneity in individuals with different mutations. In addition, interfamilial and intrafamilial variability of clinical phenotype is often seen among individuals carrying the same mutation. Macroscopically, the degree of brain atrophy observed varies with a brain weight ranging from approximately 825 to 1,290 grams. In the advanced stages, the degree of atrophy varies and may be present in the frontal and temporal lobes, caudate nucleus, putamen, globus pallidus, amygdala, hippocampus and ventral hypothalamus. Microscopically, the neuropathologic hallmark is the presence of tau protein deposits in neurons or in both neurons and glia. The cellular pathology of the neuron may resemble that of Alzheimer disease (AD) or Pick disease for the presence of neurofibrillary tangles or Pick bodies. The cellular pathology of glial cells may resemble that of progressive supranuclear palsy or corticobasal degeneration for the presence of coiled bodies in oligodendroglial cells, tufted astrocytes or astrocytic plaques. Mutations in exons 1, 10 and intron following exon 10 are associated with neuronal and glial tau deposition. Mutations in exons 9, 11, 12 and 13 lead to deposits of tau filaments predominantly in neurons. PMID- 15651320 TI - [Frontotemporal dementia: tau mutations, deposition, and molecular mechanisms of neuronal cell death]. AB - Exonic and intronic tau mutations have been described in a number of families of frontotemporal dementia and parkinsonism linked to chromosome 17. Most of missense mutations alter the ability of tau to promote microtubule assembly, whereas others influence splicing of exon10 and change the ratio of 3Rtau to 4Rtau isoform. In either case, filamentous hyperphosphorylated tau pathology in neurons and glial cells was observed in affected brains. These observations suggest that the effects of tau mutations may induce its hyperphosphorylation and accumulation, resulting in cell death. In sporadic tauopathies, decreased levels of 3R tau mRNA were detected not only in severely affected cases with progressive supranuclear palsy or corticobasal degeneration but also in cases with Alzheimer's disease or Pick's disease. In addition, levels of 3R tau mRNA were closely correlated with levels of neurofilament mRNA. These results suggest that decreased levels of 3R tau mRNA in sporadic tauopathies may be due to degeneration and loss of neurons that express 3R tau isoforms. In tauopathies, neuronal cell death may occur with multiple defects or abnormalities arisen directly or indirectly from hyperphosphorylation of tau and formation of oligomer or filamentous tau. PMID- 15651321 TI - [Anterior type dementia from the viewpoint of neuropathology]. AB - Anterior type dementia is characterized by primary degeneration in the anterior brain region and represents pick symptom. Anterior type dementia includes several pathological disease entities, almost of them usually exhibit variation of atrophic brain regions in greater or lesser degree. This variation of each disease cause confusion, namely, the same clinical manifestations occur even if it is a different disease and diversely, different clinical manifestations can occur even if it is the same disease entity. This is the reason why discrepancy between clinical diagnosis and pathology diagnosis occurs sometimes in anterior type dementia. For reasonable clinical diagnosis, anterior type dementia, separate from neuropathological diseases entity, should be deal with clinical symptom-complex, which consists of frontotemporal dementia (FTD) progressive nonfluent aphasia (PA) and semantic dementia (SD). FTD, PA and SD have, though broadly, corresponding responsible brain lesion respectively. PMID- 15651322 TI - [Application of multislice CT for the diagnosis of cerebro-vascular disease]. AB - Introduction of multislice CT (MSCT) has revolutionizing the diagnosis of cerebro vascular disease. Newly developed 32-slice MSCT enabled us to acquire isotropic volumetric data of whole brain with the resolution of 0.5-mm. CT perfusion is one of the promising application for the diagnosis of early-staged cerebral ischemia. However, it can be hazardous in terms of ionic radiation because of multiplied exposure to the same level. A new quantum denoising filter was developed in order to solve this problem. It is possible to reduce more than 80% of dose using the filter in combination with lower kv/lower mA technique. This filter can also aids to improve the detection of early CT signs, which is important for the diagnosis of cerebral ischemia. Detection of the penumbra can be made by revealing the absence of early CT sign and low perfusion area in CT perfusion. Isotropic volumetric data provided by MSCA can also be an ideal source data for the high quality 3D-CT angiography. Improved temporal resolution of MSCT in obtaining volumetric data made it possible separated visualization of arteries and veins on single 3D-CTA image. The technique to visualize brain surface using isotropic data and volume rendering algorithm was also reported. PMID- 15651323 TI - [Progress in MR imaging: differential diagnosis of infectious and inflammatory diseases with MR imaging]. AB - MR imaging is useful for diagnosis of infectious and inflammatory diseases. Diffusion weighted images (DWI) and fluid-attenuated inversion recovery (FLAIR) images revealed hyperintense signal changes in the anterior striata and cortices in patients with Creutzfeldt-Jakob disease, which were characteristic. In patients with nonherpetic and non-paraneoplastic acute limbic encephalitis, DWI and FLAIR images showed hyperintense changes in the hippocampi, amygdales, cingulate gyri, and occasionally claustrum, but not in the anterior and lateral temporal cortices, which were often involved in herpes simplex encephalitis. In ventriculitis, DWI and FLAIR images showed high signal intensity changes with niveau formation in the lateral ventricle, which indicated the presence of pus in the ventricle. In Japanese encephalitis, T2-weighted images showed high signal intensity changes in the thalami and substantia nigra, which were most often involved by Japanese encephalitis. In idiopathic cranial hypertrophic pachymeningitis, thallium-201 SPECT showed a remarkable accumulation in the abnormal dura matter. Furthermore, the abnormal uptake of thallium-201 returned to normal after treatment with corticosteroid. Thallium-201 SEPCT was useful for the evaluation of disease activity in patients with hypertrophic cranial pachymeningitis. PMID- 15651324 TI - [Advance of SPECT: differential diagnosis and evaluation of pathophysiology of neurodegenerative disorders]. AB - Development of the new radiopharmaceuticals and the statistical analysis techniques for the brain SPECT were reviewed. Both 123I-beta-CIT and 123I-FPCIT SPECT demonstrate the neuronal degeneration of the presynaptic dopaminergic system. The preclinical trials are on-going for the differentiation between essential tremor and Parkinsonian patients. 123I-iomazenil was approved by the Japanese Minister of Health, Labour and Welfare in Aril 2004, only for the use of epileptic patients considered surgical treatment. Both SPM and 3D-SSP are powerful techniques for statistical analysis of the brain perfusion SPECT. They are helpful for the detection of the functional alteration in neurodegenerative disorders, and useful for the differentiation between the Parkinsonian patients, including PD, MSA, PSP and CBD. Hypoperfusion in the posterior cingulated cortex can be also easily demonstrated by these techniques in Alzheimer's disease. 3D SRT is a technique using the anatomical standardization and ROI template consisting 636 ROIs for the whole brain. Although this technique was originally developed for the evaluation of the acetazolamide test, it can be used for the evaluation of the perfusion pattern in neurodegenerative disorders. PMID- 15651325 TI - [Measurement of brain function by positron emission tomography--clinical application of PET to neurological disorders]. AB - Positron emission tomography has enabled us to measure various fundamental parameters of human brain physiology and chemistry, such as cerebral blood flow, metabolism and synaptic function. Blood flow and oxygen metabolism is important for the understanding of cerebrovascular Disease. Glucose metabolism is useful for the determination of epileptic foci and for the evaluation of tumor malignancy. Measurement of pre- and post-synaptic function is applicable to the differential diagnosis of parkinsonism. Imaging analysis of functional neuroanatomy of these parameters are very promising for the early diagnosis of dementia, such as Alzheimer's disease. Developmennt of new radiotracers is expected for detection of early specific pathological changes and more previous changes underlying the deterioration of neurochemistry, such as genetic abnormalities. PMID- 15651326 TI - [Muscular dystrophy: advances in research works and therapeutic trials]. AB - Muscular dystrophy is defined as "a group of hereditary disorders with the major symptom of progressive muscle weakness due to muscle fiber degeneration and necrosis". After the discovery of the dystrophin gene and the gene product for Duchenne muscular dystrophy in 1986, there has been remarkable progress in the differential diagnosis and in understanding the pathogenetic mechanism of muscle fiber necrosis. With discoveries of genes responsible for many other disorders, the classification of muscular dystrophy has become more complicated; for instance, there are at least 15 diseases in the limb-girdle muscular dystrophy (LGMD) group, including the autosomal dominant forms, LGMD1A-1E and the recessive forms, LGMD2A-2I. Among them, gene defects in the sarcoglycan complex (sarcoglycanopathy) have been added to LGMD2C-2F. Sarcoglycanopathy seems to be rare in Japan since only 6-7% of LGMD patients had this defect. There are two major possible strategies in treating these patients. One is gene therapy, which is recently being investigated in the mdx mouse by using adenovirus-associated virus (AAV) vector inserted with a microdystrophin gene. Dr Takeda has reported favorable results in mdx mouse muscle with this method. Another is regeneration therapy using stem cells. There are many barriers to overcome to treat patients with stem cells isolated from bone marrow. The most difficult problem to solve is how to culture the stem cells to increase their numbers for application and how to introduce the normal dystrophin gene into these cells. PMID- 15651327 TI - [The pathomechanism and the direction of therapy development in view of cDNA microarray]. AB - We recently developed a new cDNA microarray encompassing more than 5,000 genes expressed in human skeletal muscle. We successfully identified the differences at the gene expression profiles among Duchenne muscular dystrophy patients. Using our microarray, we catalogued gene expression during myogenic differentiation. The resultant expression patterns were classified into eight groups by hierarchical cluster analysis. Among them, clusters 6, 7, and 8 contain genes which show high expression level at the later differentiation stage and encode mainly sarocmere and extracellular matrix proteins. We used genes in these clusters as markers for regeneration. We identified that these regeneration associated genes were not necessarily upregulated in Fukuyama congenital muscular dystrophy (FCMD) even though necrosis-associated genes were highly upregulated, suggesting the insufficient regenerating capability in FCMD. We have also characterized genes regulated by IGF-I simulation. We subject cascade specific inhibitors and IGF-I to human myotubes and performed gene expression profiling using our cDNA microarray. We found that PI3K/Akt-1 cascade first activates transcriptional factors such as MyoD, myogenin, and MEF2C, and then genes in clusters 6, 7, and 8, which have E-box and MEF-box where these transcriptional factors associate. We expect to develop a new therapeutic method by elucidating the molecular mechanism of muscular dystrophy and the effect of IGF-I and anti myostatin treatments. PMID- 15651328 TI - [Possible chemotherapy of muscular dystrophy caused by nonsense mutation]. AB - Gentamicin, an aminoglycoside antibiotic which causes read-through of premature termination codon during translation, has been used to rescue genetic diseases caused by nonsense mutation. Its strong side effects, however, has always threaten patients. In order to utilize other antibiotics with less side effects than gentamicin, we have shown that negamycin, a dipeptide antibiotic with read through activity in prokaryotes, restored dystrophin in skeletal and cardiac muscles of mdx mouse, an animal model for Duchenne type muscular dystrophy caused by nonsense mutation. To avoid miscoding and emerging resistant bacteria for these read-through antibiotics, further drug design and high throughput screening of gentamicin- or negamycin-related molecules will be needed. PMID- 15651329 TI - [Current status and perspective of gene therapy on dystrophic animal model]. AB - Duchenne muscular dystrophy (DMD) is an X-linked, lethal muscle disorder caused by mutations in the dystrophin gene. An adeno-associated virus (AAV) vector mediated gene transfer is one of attractive approaches to the treatment of DMD, though it has a limitation in insertion size up to 4.9 kb. Therefore, a full length dystrophin cDNA (14 kb) cannot be incorporated into an AAV vector. We previously generated micro-dystrophin transgenic dystrophin-deficient mdx mice. Micro-dystrophin CS1 transgenic mdx mice showed almost complete amelioration of dystrophic phenotypes. We, therefore, constructed an AAV vector expressing micro dystrophin deltaCS1, a modified version of CS1, driven by skeletal muscle specific MCK promoter, since the MCK promoter in AAV vector drives longer expression of the LacZ gene than the CMV promoter in skeletal muscle. We injected the AAV-MCK deltaCS1 into anterior tibial (TA) muscles of 5-week-old or 10-day old mdx mice. Dystrophic phenotypes were largely improved in both injections. Especially in the latter occasion, less than 20% of muscle fibers were microdystrophin positive at 24 weeks after the injection, but specific tetanic force of the injected muscle was not statistically different from that of control normal muscle. In conclusion, deltaCS1 micro-dystrophin introduced by an AAV vector could be a powerful tool for the gene therapy of DMD. A bigger animal model, canine X-linked muscular dystrophy will contribute to pre-clinical study of gene therapy. PMID- 15651330 TI - [Differential diagnosis of acute ischemic stroke and management on the basis of acute ischemic stroke subtype]. AB - Recent advances in neuro-imaging technology assist early clinical diagnosis for ischemic stroke subtype. The precise early diagnosis for stroke subtype plays an important role for the management in patients with acute cerebral infarction. The differential diagnosis is made according to the algorithm in the modified the Trial of Org 10172 in Acute Stroke Treatment (TOAST) classification, with reference to the results of diffusion-weighted MRI and MRA. Artery-to-artery embolism and branch atheromatus disease are diagnosed as atherothrombotic infarction according to this algorithm. Thrombolytic therapy is recommended in cardioembolic stroke within 3 hours after onset. The anticoagulant therapy with heparin is often used to prevent the recurrence for thrombosis in acute cardioemboic infarction. The selective thrombin inhibitor has recently been used in the treatment of acute atherothrombotic infarction. The antiplatelet therapy with aspirin is recommended in acute atherothrombotic infarction and lacunar infarction. The sodium ozagrel is recommended in the treatment of acute lacunar infarction. The treatment of acute ischemic stroke should be managed according to Japanese Guidelines for the management of stroke (2004). PMID- 15651331 TI - [Neurosonological examinations]. AB - Neurosonological examinations play an important role in the diagnosis of cerebrovascular disease. The target used to be limited to the extracranial carotid artery but is now extended to the intracranial arteries. Carotid artery ultrasound can evaluate not only the degree of stenosis but also the vulnerability of plaque. Transcranial Doppler can detect the micro-embolus in the intracranial artery. Transesophagial echocardiography and cruralvein ultrasound are indispensable to detect embolic source. Transesophagial echocardiography is able to detect thrombus, tumor and vegitation in the heart, patent foramen ovale (PFO) and atheroma of aorta. Because PFO is found rather commonly in healthy subjects, the detection of deep vein thrombosis (DVT) is essential for the diagnosis of paradoxical embolism. In stroke patients with PFO, RI venography can rarely detect DVT, while cruralvein ultrasound commonly detect DVT in small vein. The demand of neurosonological examinations is considered to increase more and more because of their non-invasiveness and convenience. PMID- 15651332 TI - [Natural progression of Alzheimer's disease (AD): a perspective on possible therapies for advanced AD]. AB - Patients with Alzheimer's disease (AD) show progressive deterioration in cognitive function without a spontaneous improvement in their course. Currently, acetylcholine esterase inhibitors (AchEIs) are generally used for treating AD and their beneficial effects have been confirmed for mild to moderate AD. Treatment options for patients with severe or advanced AD are limited; however, previous studies suggested that AchEIs, such as donepezil and galantamine, can provide some benefits for improving cognitive deficits even in patients with advanced AD. Furthermore, evidence of long-term benefits of donepezil treatment was reported by a recent study. Thus, AchEIs may have a wide spectrum of beneficial effects than we previously considered. Antiamyloid therapeutic strategies, including inhibition of the generation of amyloid beta-protein (A beta) and acceleration of A beta clearance, are likely to be highly effective for AD. A recent study of conditional transgenic mouse models of other neurodegenerative diseases suggested that a continuous influx of toxic aggregates of a mutant protein is required to maintain the pathological progression of the disease. Thus, although A beta generation and deposition are upstream of the amyloid cascade, it is likely that antiamyloid therapies can slow the natural progression of AD even in an advanced stage of the disease. PMID- 15651333 TI - [Clinical heterogeneity in mild cognitive impairment--beyond clinical diagnosis- towards imaging amyloid]. AB - Alzheimer's disease (AD) generally begins with mild memory problems in an insidious manner and progresses to develop multiple cognitive and functional impairment within a few years. Currently, the diagnosis of AD requires multiple cognitive deficits including memory disturbance and exclusion of other dementing disorders. However, normal elderly people quite commonly complain of increasing forgetfulness with age, and we have not developed an objective tool that can reliably support the diagnosis of AD. Recent advances in our understanding of neurobiology of AD demonstrate that AD starts with accumulation of amyloid beta protein (A beta) followed by abnormal phosphorylation of tau protein and a massive neuron death in vulnerable brain areas. We have shown that cerebrospinal fluid tau are elevated in subjects with mild cognitive impairment (MCI), the earliest detectable clinical stage of dementia and AD, suggesting that the pathogenic cascade of AD may arrive at the stage that finally leads to an accumulation of abnormally phosphorylated tau in the MCI stage. These results may highlight the need to develop another diagnostic tool that reliably monitors and visualize brain beta-amyloid burden in living subjects who are at increased risk of developing AD. We assume that the detection of asymptomatic stage of AD followed by an early intervention may lead to maximum therapeutic benefits. In an attempt to accomplish this goal, we have generated several novel chemicals that specifically bind to A beta peptide upon entry into rat brain. The "amyloid imaging" seems quite promising if safely and successfully applied in humans because it is a noninvasive technique and applied multiple times in a single subject. PMID- 15651334 TI - [Inherited dystonia update]. AB - Among idiopathic dystonia, inherited dystonia whose causative gene or linkage has been clarified are named as DYT1 to DYT15. The causative genes of DYT1, 5 and 11 were identified as genes of Torsin A, GTP cyclohydrolase I, and epsilon sarcoglycan, respectively. All three are inherited dominantly. DYT1, and DYT5 which is known as Segawa disease, are dystonia with onset in childhood. After identification of the causative gene, each disorder was found to show the various phenotypes. In both DYT1 and Segawa disease, early onset develops generalized dystonia, and later onset focal or segmental dystonia. Deep brain stimulation of globus pallidus internal segment shows remarkable effect on DYT1. Segawa disease responds markedly to L-dopa without any side effect lifelong. The pathophysiology of Segawa disease is that partial deficiency of BH4 resulted from GCH I deficiency, rate limiting enzyme of synthesis of BH4, affects the TH activity at terminal of nigrostriatal dopamine neuron. The role of Torsin A in the pathogenesis of DYT1 is unknown. For a certain neuron or neuronal system to manifest a clinical symptom, it should reach to a certain maturational level. The symptoms of inherited dystonia are influenced by the developmental level of responsible neuron or neuronal circuit. PMID- 15651335 TI - [Hereditary chorea--update]. AB - Understanding molecular genetical background of hereditary chorea has recently been progressed so far. Triplet repeat expansion diseases including, Huntington disease, in which CAG expansion has been identified in the IT-15 or Huntingtin gene, and Huntington disease like-2, in which CTG expansion in junctophilin-3 (JPH3) gene occurs, causes selective degeneration of striatum in the brain. Octapeptide repeat expansion in the prion gene in Huntington disease like-1 has been also identified. Neuroacanthocytosis syndromes including McLeod syndrome and chorea-acanthocytosis cause acanthocytosis in the red blood cells and chorea due to the degeneration of caudate nucleus in the brain. The XK gene on the X chromosome is mutated to lose its function in McLeod syndrome. CHAC gene coding chorein is mutated to lead loss of function in chorea-acanthocytosis. Selective degeneration in the striatum, especially in the caudate nucleus might be associated with the molecular cascade of expanded polyglutamine or polyleucine or octapeptide and the loss of function of the XK protein and of chorein protein. PMID- 15651336 TI - [Diagnostic significance of autoantibodies in neurological diseases]. AB - Significance of autoantibodies in neurological diseases is roughly grouped into three, as 1) disease process is thought to be directly caused by the antibodies because the antibody can reproduce the disease process, like anti-acetylcholine receptor antibody in myasthenia gravis, anti-P/Q type voltage-gated calcium channel antibody in Lambert-Eaton myasthenic syndrome, etc., 2) antibodies are thought to be closely related to the disease because plasma exchange ameliorate the disease promptly, however without success of transfer experiments, like Guillain-Barre syndrome/Miller Fisher syndrome and anti-ganglioside antibodies and 3) antibodies are very specific and become the useful markers for the diagnosis, however without direct evidence of disease transfer, like paraneoplastic neurological syndromes and anti-Yo or Hu antibodies. Overall, the roles of antibodies are different between diseases, but the presence of antibodies support the basis of intervening immunotherapy, antibody titers predicted the activity of the diseases, or they are useful markers for exploring underlying cancers in paraneoplastic neurological syndromes. PMID- 15651337 TI - [Channelopathy update]. AB - Electrophysiological techniques, molecular technology and molecular genetics have been quickly developed in the late 20th century. Now many genetical abnormalities of channel proteins including voltage gated ion channels, ligand gated ion channels and Ca induced Ca channel have been discovered. In this article, I briefly review these genetical abnormality of ion channels ("channelopathies") and the pathophysiological background of these channelopathies. PMID- 15651338 TI - [New international classification of headache disorders (ICHD-II)]. AB - It has been almost 15 years since the first edition of the International Headache Classification appeared in 1988. It was widely accepted and well tolerated. However, rapid progress of the headache research is pushing for a drastic revision of the classification. The new International Classification of Headache Disorders (ICHD-II) was disclosed in September 2003 as a preprinted version and formally published in January 2004. The ICDH-II classifies headache disorders into 3 parts: the primary headache, the secondary headache and cranial neuralgia, central and primary facial pain and other headaches. Basically, the most important diagnostic criteria, those of migraine and tension-type headache, remain unchanged. Several new entities such as chronic migraine, hypnic headache, hemicrania continua, benign thunderclap headache and medication overuse headache have been added. This will encourage intensified headache researches in the future. PMID- 15651339 TI - [An update on the familial headache syndromes]. AB - Migraine is a common form of the chronic headache syndromes. Although the pathogenesis of migraine still remains enigmatic, there have been remarkable progress in headache research. Point mutations of P/Q-type Ca2+ channel alpha 1 subunit (CACNA1A) gene have been identified in familial hemiplegic migraine (FHM), which linked to chromosome 19 (FHM-1, OMIM 141500). Na-K ATPase alpha2 gene has been identified as the causative gene for FHM linked to 1q21-23 (FHM-2, OMIM 602481). Common forms of migraine (i.e. migraine with and without aura) seems to be caused from multifactorial genetic factors and environmental factors. An association study of allelic variation at Codon 23 (Cys or Ser) of 5HT2C-R gene in Japanese samples revealed that the Ser allele frequency in migraine with aura was significantly higher than that in the non-headache controls. However, negative association of this polymorphism have been reported in Caucasian migrainures. The C677T allelic variation of 5,10-methylenetetrahydrofolate reductase (MTHFR) are focused on in association with the coronary heart diseases and the cerebrovascular diseases. The T allelic frequency in migraine sufferers was significantly higher than that in controls. The C677T mutation of MTHFR is one of the genetic risk factors for migraine. These observations are confirmed in Turkish, Australian and Spanish samples. Positive associations of angiotensin converting enzyme (ACE) gene, endotheline receptor-A (ET-A) gene, and insulin receptor gene have been reported. Using the genomewide screen technology, significant linkage between the migraine with aura and a marker on 4q24 has been reported in Finnish families. The genome wide screen analysis will be one of the powerful strategies on exploring migraine gene. Genetic study of migraine headache is a promised and fruitful field and will provide deep understanding to migraine headache. Discovery of new responsible or susceptible genes to migraine will also open an avenue to develop new therapeutic strategy of migraine. PMID- 15651340 TI - [Future of gene therapy for Parkinson's disease]. AB - Parkinson's disease (PD) is a good target for gene therapy because the lesion is localized to the substantia nigra (SN). There are several approaches in gene therapy for PD. For enhancing dopamine production, the candidate genes are tyrosine hydroxylase, AADC and/or GTP cyclohydroxylase I. The second approach is a neuroprotective strategy, which is based on the usage of genes for neurotophic factors or anti-apoptotic agents. We also showed that Apaf-1-dominant negative inhibitor delivery using an AAV vector system could prevent nigrostriatal degeneration in MPTP mice, suggesting that it might be an anti-mitochondrial apoptotic gene therapy for PD. In 2003, the first gene therapy trial for PD performed at New York Weill Cornell Medical Center. The treatment is designed to deliver glutamic acid decarboxylase (GAD), the gene responsible for making GABA, into the subthalamic nucleus to "quiet down" that nucleus and alleviate Parkinson's symptoms. The last approach is replacement of disease for autosomal recessive PD. Because autosomal recessive juvenile parkinsonism (ARJP) involves the loss of function of parkin gene, gene therapy employing the parkin gene may prevent nigral cell death. PMID- 15651341 TI - [Cell therapy in Parkinson's disease]. AB - An approach for symptomatic Parkinson's disease (PD) therapy is fetal dopamine neuron transplantation. This approach remains the technical and ethical difficulties in obtaining sufficient and appropriate donor fetal brain tissue. In developments of stem cell biology, neural stem cells exist in the adult brain as well as embryo and have the capacity to regenerate and to give rise to the three cell lineages in the nervous system. Embryonic stem cells (ES cells) and multipotent adult progenitor cells (MAPCs) are pluripotent cells, which give rise to all cells in the organism. Current findings suggest that stem cells but not fetal brain tissues may be suitable for cell replacement therapies in the treatment of neurodegenerative disorders. We will briefly review the current state of cell therapy, and will critically discuss the potential of stem cells for the treatment of PD. PMID- 15651342 TI - [Principles of diffusion-weighted MR imaging and application to clinical neurology]. AB - Diffusion-weighted Imaging (DWI) is a advantageous method for early detection of cerebral ischemia. DWI with echo-planar sequence (EP-DWI) offers multisectional images sensitive to cytotoxic edema in a very short aquisition time and is almost free from motion artifact. However, the susceptibility artifacts and low spatial resolution of EP-DWI must be improved. In estimation of DWI, influence of T2 must be considered, because DWI is almost always based on T2-weighted imaging. DWI is applied to other cerebral disorders such as degenerative and demyelinating disease, infectious disease, tumors or so. In order to demonstrate water diffusion precisely, diffusion tensor imaging (DTI) must be introduced and applied to anisotropy indices such as fractional anisotropy (FA) and depiction of neurofiber direction, tractography. Measurements of FA in various degenerative diseases may contribute to differentiation in normal appearing white matter. Diffusion tensor tractography may provide more information about relationship of major white matter tract such as corticospinal tract with brain lesion. Furthermore, DWI and DTI are expected to demonstrate diffusion of protons of aminoacids such as choline, creatine, NAA and provide more pertinent information of regional pathologic state of the brain in future. PMID- 15651343 TI - [Brain tumors: the current WHO classification and neuroimagings]. AB - The current World Health Organization Classification of tumors of the Central Nervous System (WHO 2000) lists more than 120 types of brain tumors. Classification of brain tumors is an evolving process, with obsolete entities discarded and newly recognized tumors added with each successive revision. In the past, classification has relied heavily on morphologic pattern recognition and immunohistochemical identification of differentiation antigens. With the discovery of the association between deletions on chromosomes 1 and 19 and the chemosensitivity of anaplasic oligodendrogliomas, a new era of molecular classification of brain tumors began. It is a virtual certainty that current advances in molecular methodologies, particularly in the fields of genomics, transcriptonomics, and proteomics, will revolutionize brain tumor classificaton in the very near future. PMID- 15651344 TI - [Diagnosis and treatment of CNS parasite infection with special reference to parasitic myelitis]. AB - The occurrence of visceral larva migrans due to Ascaris suum (A. suum) and Toxocara canis (T. canis) has occasionally been reported in Japan, although parenchymatous involvement of the CNS is extremely rare in A. suum/T. canis visceral larvae migrans. Recently we experienced 7 cases with myelitis caused by visceral larva migrans due to A. suum/T. canis (parasite myelitis). The characteristics of this myelitis are: (1) sensory disturbances (Lhermitte's sign, paresthesia, and hypesthesia) are predominant symptoms, while severe motor weakness is rare, (2) spinal cord lesions on T2-weighted MRI show more extensive lesions compared with mild symptoms, (3) Gadolinium enhancement of spinal cord lesions are limited as compared with spinal cord lesions on T2-weighted MRI lesions, (4) Some cases show the presence of eosinophils in CSF, while others show Th2 deviation in CSF supernatant, and (5) Tests for anti-A. suum/T. canis IgG antibody are strongly positive in serum and CSF. Moreover, 6 percent of 108 consecutive cases with non-compression myelopathy presenting at the Department of Neurology at Kyushu University Hospital from January, 1998 to December, 2002 had parasitic myelitis. Myelitis from visceral larva migrans due to A. suum/T. canis might be overlooked because of its mild neurologic impairment without systemic symptoms, but should be considered as one of the differential diagnoses in non compression myelopathy. PMID- 15651345 TI - [Influenza-associated encephalopathy]. AB - Infection with influenza viruses produces a spectrum of clinical responses, ranging from upper respiratory illness to central nervous system (CNS) involvement. Recently, the number of reports of influenza-associated encephalopathy in Japan has increased. During the winters of 1997-1998 and 1998 1999, when epidemics of type A influenza (H3N2) occurred, many pediatricians reported cases of influenza-associated encephalitis or encephalopathy in children. The prominent indicators of influenza-associated encephalopathy are the abrupt onset of seizures and coma within a few days of developing a high-grade fever. These patients often develop multi-organ failure and have high morbidity and mortality. The pathogenesis of influenza-associated encephalopathy remains unclear. Because not all patients with influenza develop encephalopathy, infection with the influenza vieus is necessary, but not sufficient, for the development of influenza-associated encephalopathy. Viral RNA is rarely detected in the cerebrospinal fluid (CSF), and the presence of viral antigen in the brain has not been proven. Pathological findings, including the lack of detectable viral antigen and inflammatory cells in brain tissues, suggest that direct viral invasion and inflammation are unlikely to be causes of this disease. In influenza associated encephalopathy, serum and CSF concentrations of several proinflammatory cytokines and cytokine receptors--such as interleukin (IL)-6, IL 1beta, and soluble tumor necrosis factor (TNF) receptor-1 (sTNF-R1)--are elevated and are related to the clinical severity of the disease. Moreover, the damage of vascular endothelial cells has been shown. Using SNPs, molecular analysis of whole genome of the patients are now on going. PMID- 15651346 TI - [International classification of epileptic seizures, epilepsies, and epileptic syndromes]. AB - Epilepsy is one of the common diseases in neurology. Its correct diagnosis and classification are important for choosing the most appropriate treatment. Currently, Classification of Epileptic Seizures in 1981 and Classification of Epilepsies and Epileptic Syndromes in 1989, both were proposed by International League Against Epilepsy (ILAE), are available. As for Classification of Epilepsies and Epileptic Syndromes, two divisions continue to be widely used to shape the major classes: The first separates epilepsies with generalized seizures (generalized epilepsy) from epilepsies with partial or focal seizures (localization-related, partial or focal epilepsies). The other separates epilepsies of known etiology (symptomatic or secondary epilepsies) from those that are idiopathic (primary) and those that are cryptogenic. In 2001, ILAE Task Force proposed a new classification. However, its daily use has yet been limited. Simple and useful classification will lead to better understanding and more appropriate management of epilepsy. PMID- 15651347 TI - [Genetic approach to the pathogeneses of epilepsy]. AB - Recently, genetic causes have been identified in certain epilepsy syndromes in which the phenotypes are similar to common idiopathic epilepsies. Interestingly, almost all such genetic abnormalities were detected in genes encoding ion channels expressed in the brain. Thus such epilepsy syndromes are disorders of ion channels, i.e., "channelopathies". The list of ion channel abnormalities that are associated with childhood epilepsy is expanding and includes the followings. Mutations of the genes encoding two subunits of the neuronal nicotinic acetylcholine receptor, a ligand-gated ion channel, were found in autosomal dominant nocturnal frontal lobe epilepsy. Mutations of two KCNQ K+-channel genes were identified in benign familial neonatal convulsions. Mutations of the genes encoding several subunits of the voltage-gated Na+-channel and GABA(A) receptor, a ligand-gated ion channel, were also identified as underlying causes of various epilepsy syndromes, such as autosomal dominant epilepsy with febrile seizures plus or generalized epilepsy with febrile seizures plus, benign familial neonatal infantile seizures and autosomal dominant juvenile myoclonic epilepsy. Mutations within the same gene can result in different epilepsy phenotypes. Abnormalities of Na+-channel alpha1 subunit were also associated with severe myoclonic epilepsy in infancy. Epilepsy syndromes mentioned above, except for severe myoclonic epilepsy in infancy, were familial epilepsy syndromes showing dominant inheritance with high penetrance while common idiopathic epilepsies do not show obvious inheritance. However, the similarities in symptomatology between such familial epilepsies and common idiopathic epilepsy may provide us with clues to the genetics of common idiopathic epilepsies. PMID- 15651348 TI - [Multiple system atrophy--update]. AB - Multiple system atrophy (MSA) is an adult-onset neurodegenerative disorder, showing various combination of progressive autonomic failure, cerebellar ataxia, and levodopa poorly responsive parkinsonism. MSA accounts for more than 40% of spinocerebellar ataxias in Japan. Pathologically, myelinopathy, neuronal loss and gliosis are the cardinal features in the brain of MSA. In addition, excessive accumulation of alpha-synuclein, mostly in oligodendroglia and partly in neuron, characterizes the cellular pathology of the disorder. However, mechanism causing the disorder is not known. Clinical diagnosis of MSA is based on Quinn's criteria or, more recently, on Consensus Criteria. In Japan, criteria of the Research Committee of Ataxic Diseases, the Ministry of Health and Welfare of Japan, was popular, and it contributed to the research base on MSA. Besides the major manifestations incorporated into those criteria, various dystonic manifestation, rhythmic myoclonus, emotional incontinence, sleep disturbance, sleep-related movement disturbances, and signs of vasomotor dysfunction, provide aids in the differential diagnosis of MSA. In neuroimaging studies, not only MRI but dopamine transporter and D2 receptor imaging by SPECT also contribute to the diagnosis of MSA. These laboratory diagnostic procedures can contribute to improve reliability of the diagnosis, and needs to be taken into account when preventive measures become available. PMID- 15651349 TI - [Corticobasal degeneration and atypical progressive supranuclear palsy: their symptomatology, laboratory examination and differential diagnosis]. AB - Corticobasal degeneration (CBD) and atypical progressive supranuclear palsy (PSP) were reviewed with special reference to their symptomatology, laboratory examination and differential diagnosis. In our survey of the autopsy cases of CBD in Japan, only about 60% of the pathologically confirmed CBD cases were correctly diagnosed clinically, meaning that atypical (non-classical) clinical forms are common in CBD. Concerning the autopsy cases of PSP in Japan, 75% of the PSP cases had correct clinical diagnosis. In literatures, the clinically atypical CBD includes (1) frontotemporal dementia, also with primary progressive aphasia and frontal lobe dementia as subforms, (2) PSP-like form, and (3) others. The clinically atypical PSP comprises (1) pure akinesia, (2) pure easy falling syndrome (Yuasa), (3) no postural instability, (4) no gaze palsy, (5) asymmetric parkinsonism, (6) no or severe dementia, etc.. PSP with cortical manifestations such as primary progressive aphasia and CBD-like features were also reported. The atypical CBD and PSP probably reflect the distribution of tau pathology different from that in typical forms. Except for the report that phosphorylated tau is increased in CSF in CBD, but not in PSP (Urakami et al), no reliable laboratory data have been available on clinical differentiation between atypical CBD and PSP. PMID- 15651350 TI - [Physiological approach to peripheral neuropathy. Conventional nerve conduction studies and magnetic motor root stimulation]. AB - In this communication, I first show some points we should mind in the conventional peripheral nerve conduction studies and later present clinical usefulness of motor root stimulation for peripheral neuropathy. CONVENTIONAL NERVE CONDUCTION STUDIES (NCS): The most important point revealed by the conventional NCSs is whether neuropathy is due to axonal degeneration or demyelinating process. Precise clinical examination with this neurophysiological information leads us to a diagnosis and treatment. Poor clinical examination makes these findings useless. Long standing axonal degeneration sometimes induces secondary demyelination at the most distal part of involved nerves. On the other hand, severe segmental demyelination often provokes secondary axonal degeneration at distal parts to the site of demyelination. These secondary changes show the same abnormal neurophysiological findings as those of the primary involvement. We should be careful of this possibility when interpreting the results of NCS. NCS of sensory nerves is not good at revealing demyelinating process. Mild temporal dispersion of potentials often reduces an amplitude of SNAP or loss of responses, which usually suggests axonal degeneration, because of short duration of sensory nerve potentials. MOTOR ROOT STIMULATION IN PERIPHERAL NEUROPATHY: Magnetic stimulation with a coil placed over the spine activates motor roots and evokes EMG responses from upper and lower limb muscles. The site of activation with this method was determined to be where the motor roots exit from the spinal canal (intervertebral foramina) (J Neurol Neurosurg Psychiatry 52 (9): 1025-1032, 1989) because induced currents are very dense at such a foramen made by electric resistant bones. In several kinds of peripheral neuropathy, this method has been used to detect a lesion at a proximal part of the peripheral nerves which can not be detected by the conventional NCSs. I present a few cases in whom motor root stimulation had a clinical merit. In a patient with neuralgic amyotrophy, motor root stimulation disclosed a conduction block between the cervical intervertebral foramen and brachial plexus which was not detected by conventional NCSs. Motor root stimulation clearly revealed demyelination in a patient with CIDP in whom sural nerve biopsy findings suggested axonal degeneration, that must be secondary to demyelination. In a patient with tomacular neuropathy, magnetic stimulation revealed conduction delay in the spinal nerve within the spinal canal (Clin Neurol (Jap), 28: 447-452, 1988). Based on the above results, combination of NCSs and magnetic motor root stimulation must brush up the neurophysiological approach to peripheral neuropathy. PMID- 15651351 TI - [Update on hereditary neuropathy]. AB - Hereditary neuropathies are classified into several subtypes according to clinical, electrophysiologic and pathologic findings. Recent genetic studies have revealed their phenotypic and genetic diversities. In the primary peripheral demyelinating neuropathies (CMT1), at least 15 genes have been associated with the disorders; altered dosage or point mutation of PMP22, GJB1, MPZ, EGR2, MTMR2, NDRG1, PRX, SOX10, GDAP1 and MTMR13/SBF2. In the primary peripheral axonal neuropathies (CMT2), at least 10 genes have been associated with these disorders; NEFL, KIF1B, MFN2, GAN1, LMNA, RAB7, GARS, TDP1, APTX, and SETX. In addition, some mutations in GJB1, MPZ, GDAP1 and NEFL also present with clinical and electrophysiologic findings of CMT2. Patients with TDP1, APTX or SETX mutations share common clinical findings; autosomal recessive inheritance, cerebellar ataxia, and axonal neuropathy. These genes are suspected to be related to DNA/RNA repair and induce cell death especially in neuronal cells. In addition to the above diseases, we have reported a new type of NMSNP (MIM# * 604484) characterized by proximal dominant neurogenic atrophy, obvious sensory nerve involvement and the gene locus on 3q12.3. Here, we summarize the genetic bases of hereditary neuropathies and attempt to highlight significant genotype-phenotype correlations with a special interest in nonsense-mediated mRNA decay pathway. PMID- 15651352 TI - [Limb-girdle muscular dystrophy; update]. AB - Sixteen different forms of limb-girdle muscular dystrophies (LGMDs) have emerged from recent molecular genetic studies, six forms with a dominant trait and ten forms with a recessive trait. Among 1,420 Japanese patients with muscular dystrophy analyzed at NCNP, LGMD is the secondly largest category (19%) following dystrophinopathy (56%). Within LGMDs, the occurrence of LGMD2A (calpainopathy), LGMD2B (dysferlinopathy), and LGMD2C-F (sarcoglycanopathy) is 26%, 18%, and 6.6%, respectively, however, causative genes have not been specified in about 50% of the LGMD patients. LGMD2A patients show atrophy prominent in shoulder and pelvic girdle muscles without calf muscle hypertrophy, and abundant lobulated fibers in muscle biopsy. Four major mutations unique to the Japanese population, have been identified. Pathogenesis attributes to a loss of proteolytic activity of mutant calpain-3. Dysferlin, the defective protein in LGMD2B, is a ferlin family molecule possessing six C2 domains probably mediating the resealing mechanism of the damaged sarcolemma. Mutations in the dysferlin gene result in Miyoshi distal myopathy and distal anterior compartment myopathy other than LGMD2B. Among four sarcoglyconopathies, LGMD2D is the most common form, whereas LGMD2F has not yet been reported. In sarcoglycan-deficient skeletal muscle, matrix metalloproteinases may be involved in the beta-dystroglycan processing which underlies the pathogenesis of sarcoglycanopathy. PMID- 15651353 TI - [Plasticity of skeletal muscle differentiation generating stem cell-like phenotype: possible application to cell therapy for muscular diseases]. AB - Drug therapy, gene therapy, and cell therapy may be effective to degenerative muscular diseases caused by genetic mutations including muscular dystrophy. Stem cells including embryonic stem (ES) cells and adult stem cells (tissue stem cells) are generally believed to be applicable to cell therapy. However, both types of cells have several problems to be solved for the cell therapy. We have shown that the expression of SV40 large T antigen in terminally differentiated mouse C2 skeletal muscle myotubes induce mitosis and dedifferentiation. This finding has exploded the concept that terminally differentiated cells never proliferate. It remains to be determined, however, whether mature myofibers in vivo are capable of dedifferentiation and proliferation. Here we present the data showing that mouse myofibers dedifferentiate and proliferate to form many mononucleated cells. Eventually, these dedifferentiated cells redifferentiate and regenerate myofibers. The isolated dedifferentiated cells show stem cell-like phenotype. These cells are expected to be applied to the cell therapy for degenerative muscular diseases. PMID- 15651354 TI - [Towards the establishment of a postgraduate training program of neurology--in the case of Tokyo Medical and Dental University]. AB - The training as neurologist in Department of Neurology and Neurological Science, Tokyo Medical and Dental University has mainly been performed through 3rd to 5th year after the graduation. It includes training for neurophysiology, neuroradiology and neuropathology as well as clinical neurology. Neurophysiology training has the separate 3 months course where trainee residents study exclusively neurophysiology but neuroradiology and neuropathology were learned during training for clinical neurology in general. The major problem of our training program is that the quota is so small to prepare enough trainee's positions. This appears also true in many institutes in Japan according to a questionnaire survey by this workshop. The number of neurology staff should be increased to accomplish better training for Neurology in each institute. The Japanese Society of Neurology should support this effort in each institute. PMID- 15651355 TI - [Postgraduate training program for neurology]. AB - In order to establish neurology as a field of medical subspecialty in Japan, it is inevitable to make an authorized postgraduate training program and board system. It should concretely define essential levels required for clinical practice of neurology. Those who participate in the training should register to each of the authorized program. It is of great importance that we neurologists should share the concept of integrated postgraduate training system for neurology. PMID- 15651356 TI - [Postgraduate education program in neurology at Iwate Medical University]. AB - The major goal of the program is to provide residents with the opportunity to gain the knowledge, attitudes, and skills to enable them to provide exemplary health care to people with neurological disease. The program will facilitate learning through an atmosphere of collegiality and mutual respect that fosters active communication between residents and faculty. We will stress the pursuit of excellence in clinical neurology and encourage the development of a deeper interest in the academic aspects of the discipline. We will endeavor to create a level of excitement that will stimulate our residents to seek further education and pursue careers in academic neurology. Our program aims to serve both the present and future requirements of our patients and our discipline. The program consists of 24 months of internal medicine and other clinical rotations; 12 months of clinical and basic neuroscience research including neurophysiology, neurochemistry, neuroradiology, neuro-rehabilitation and neurogenetics; 24 months of clinical neurology including the primary care of inpatients on the neurology ward, the assessment of emergency department, and neurological consultation; and 12 months of residency as a senior neurology resident consultant in inpatient and outpatient settings with responsibility for teaching and supervising junior residents. PMID- 15651357 TI - [Postgraduate neurology training in new clinical training system]. AB - Postgraduate neurology training in new clinical training system in Oita University Hospital was presented. Clinical intern training for first 2 years are performed according to programs proposed by Clinical Training Institute for Interns in Oita University Hospital. This program includes neurology for at least 1 month. Interns will get in clinical management for common and main neurological disorders such as convulsion, unconsciousness, stroke, Parkinson disease and intractable neurological disorders in addition to standard medical skills. Neurology resident training for specialist begins after clinical intern training and is performed in own program proposed by our department. However, the level of educational and training quality is not always high, because of the lack of teaching staff, and insufficient curriculums without rotation system to neuropathology and neuroradiology. To improve this educational program, we need to alter own well-established postgraduate educational programs. The quality of programs must to be approved by Japanese Society of Neurology. PMID- 15651358 TI - [CT and MR imaging of gynecological emergency disease]. AB - We describe the CT and MRI findings of gynecologic emergency diseases: pelvic inflammatory disease, ectopic pregnancy, ovarian hemorrhage, ovarian torsion, rupture of ovarian tumor, eclampsia, and HELLP (hemolysis, elevated liver enzymes, and low platelet count) syndrome. Diagnostic keys to these diseases are presented in this review. CT and MRI play a complementary role to sonography in accurately diagnosing these diseases. In situations that require an exact, immediate diagnosis, radiologists should be familiar with the key imaging findings. PMID- 15651359 TI - [Evaluation of enhancement patterns using three-dimensional dynamic contrast enhanced MR imaging in 209 cases of breast cancer]. AB - PURPOSE: To evaluate contrast-enhanced patterns using three-dimensional (3D) dynamic MR imaging in 209 cases of breast cancer. MATERIALS AND METHODS: Three dimensional dynamic imaging of the breast (1.5-T scanner) was performed in 755 cases. Of 227 breast carcinomas, 209 cases that were histologically confirmed were enrolled in this study. The histological diagnoses included ductal carcinoma in situ (DCIS) (n=12), invasive ductal carcinoma (n=176), mucinous carcinoma (n=10), medullary carcinoma (n=4), invasive lobular carcinoma (n=6), and other (n=1). Tumor size was a mean 24.6 mm in diameter (range, 7-110 mm), including 110 cases of small breast carcinomas (< or = 2 cm). The contrast-enhancement pattern was analyzed from the early phase of 3D-MRI and the post-contrast T1-weighted SE image acquired before the delayed phase of 3D-MRI. The type of peripheral enhancement (PE) was evaluated on the early and delayed phases of 3D-MRI. These enhancement patterns were also compared with the histological findings of small breast carcinomas. RESULTS: The sensitivity of the 209 cases of breast carcinoma was 99% (207/209) on 3D dynamic MRI. Two hundred six cases (98.6%), including all DCIS, showed strong enhancement on the early phase of 3D MRI. An increased washout pattern showing signal intensity lower than that of fat on post-contrast T1-weighted imaging was noted in 201 cases (96.1%), of which 179 cases (85.6%) showed washout patterns. PE was identified in 63 cases (30.4%) and in 35 of 110 cases of small breast carcinoma (31.8%). Delayed PE following central washout was noted in 65% of 63 cases and 71.4% of 35 cases of small breast carcinoma. Delayed PE was well correlated with marginal fibrosis. CONCLUSION: Three-dimensional dynamic MRI of the breast was highly sensitive for breast carcinoma. Delayed PE following central washout was considered a specific finding of breast carcinoma. It is important to understand the enhancement patterns of 3D dynamic MRI for excellent specificity of breast carcinomas. PMID- 15651360 TI - [MR arthrographic findings of SLAP lesion type V]. AB - PURPOSE: The purpose of this study was to determine the findings of MR arthrography of the shoulder and to assess the role of MR arthrography in the diagnosis of superior labrum anterior to posterior (SLAP) lesion type V. METHODS AND MATERIALS: Two radiologists retrospectively reviewed fat-suppressed T1 weighted MR arthrography images of six patients who were diagnosed with SLAP lesion type V by arthroscopy. Each imaging plane, including the transverse, oblique coronal, oblique sagittal, and oblique transverse in abductor external rotation (ABER) position were evaluated for the following three findings: tear of the superior labrum at biceps tendon insertion, Bankart lesion, and continuity of the two former findings. RESULTS: Tear of the superior labrum was shown in all patients on oblique coronal images. Bankart lesion was noted in five patients on the transverse images and in four on the oblique sagittal images. On the oblique transverse images in ABER position, Bankart lesion was shown in all patients. The continuity of the two former findings was noted in three patients on the ABER positioned images. Therefore, three patients could be diagnosed as having SLAP lesion type V by MR arthrography in our series. CONCLUSION: It is difficult to detect all three findings of SLAP lesion type V in one imaging plane, however, a combination of multi-directional images may increase the feasibility of MR arthrography in diagnosing SLAP lesion type V. PMID- 15651361 TI - [Breast multidetector-row CT with histopathologic correlation]. AB - PURPOSE: To evaluate the correlation between multidetector-row CT (MDCT) and histopathologic findings using the same MDCT image as the histopathologic cross section. MATERIALS AND METHODS: MDCT with contrast enhancement was performed in 10 patients with breast cancers (8 invasive ductal carcinomas, one invasive lobular carcinoma, and one non-invasive ductal carcinoma). We tried to reconstruct multiplanar reconstructions (MPR) in the same plane as the histopathologic cross-section, and we evaluated the histopathologic findings of the false-positive lesions. RESULTS: In all cases, we obtained the same MDCT image as the histopathologic cross-section. There were 10 main lesions and 18 other lesions. In the other lesions, we found no false-negative lesions and 11 false-positive lesions. False-positive lesions included periductal fibrosis, cystic change, duct papillomatosis, sclerosing adenosis, fibroadenoma, and others. CONCLUSION: Using MDCT of the breast, it is possible to obtain good correlation between CT images and histopathologic findings. MDCT is thought to be useful in the evaluation CT findings on the basis of histopathologic evidence. PMID- 15651362 TI - [Peripheral venous stenoses of hemodialysis shunts: improved long-term patency rates with stent placement]. AB - PURPOSE: The purpose of this study was to evaluate the effectiveness of endovascular stents placed in the peripheral veins of hemodialysis shunts. MATERIALS AND METHODS: 156 endovascular stents were placed in the peripheral veins of 155 hemodialysis shunts with 220 stenoses. Among these, 106 stenoses of 93 hemodialysis shunts had been treated by percutaneous transluminal angioplasty (PTA) before stent placement. RESULTS: The initial success rate was 97.7%. Primary radiologic patency rates of the stents at 6 months, 1 year, and 2 years were 69.8%, 49.1%, and 45.8%, respectively. Secondary radiologic patency rates at 1 year, 2 years, and 3 years were 94.0%, 91.8%, and 88.0%, respectively. Primary clinical patency rates at 6 months, 1 year, and 2 years were 64.4%, 43.4%, and 27.3%; while secondary clinical patency rates at 1 year, 2 years, and 3 years were 93.5%, 86.5%, and 73.4%, respectively. Radiologic and clinical primary patency rates of 106 stenoses and 93 hemodialysis shunts were significantly higher than those of PTA that had been performed before stents were placed. CONCLUSION: Stent placement for stenoses of the peripheral veins of hemodialysis shunts recurring in three months and treated by PTA alone can improve long-term patency. PMID- 15651363 TI - [Survival of glottic carcinoma patients treated with definitive radiation therapy: value of the Sixth edition of the UICC TNM staging system]. AB - PURPOSE: To evaluate the survival of glottic carcinoma patients treated with radiation therapy (RT). The predictive value of the new sixth edition of the UICC staging system was also evaluated. METHODS AND MATERIALS: 193 patients with T1 2N0 glottic squamous cell carcinoma, classified according to the fifth edition of the UICC staging system, were treated with definitive RT. Of them, 130 patients evaluated with pretreatment radiological examinations (CT and/or MR) were retrospectively classified according to the UICC sixth edition. RESULTS: According to the UICC fifth edition, 132 lesions were staged as T1 and 61 as T2. Thirty lesions were categorized as stage T3 according to the UICC sixth edition. Of all patients, 13 died of glottic carcinoma, 25 of second malignancy, and 20 of intercurrent disease. Second primary sites included lung (n=8), esophagus (n=4), liver (n=4), pancreas (n=3), and others (n=6). The 10-year overall survival rate was 66%, and cause-specific survival rates of glottic carcinoma and second malignancy were 91% and 86%, respectively. Multivariate analysis confirmed T stage in the UICC sixth edition as an independent predictor for death from glottic carcinoma. Although there were no significant factors for second malignancy, there was no death from second malignancy in non-smoking patients. CONCLUSION: Second malignancy was the most frequent cause of death, and an association with smoking was suggested. The UICC sixth edition appears to correctly identify patients with T3 lesions as a high-risk group not only for local failure but also for survival. PMID- 15651364 TI - [Vaccination therapy for Alzheimer's disease]. PMID- 15651365 TI - [Regenerative medicine of bone and cartilage]. AB - Regenerative medicine, which takes advantage of the unique capacity of stem cells, is a novel medical strategy to cure irreversible organ failure. There are three important elements in regenerative medicine: cells, scaffolds and signals. Although substantial progress regarding each element has been made in the past few years, it still falls short of clinical applications. In the geriatric field, osteoporosis, osteoarthritis and periodontitis are the major targets of regenerative medicine. They are usually not life-threatening, but often severely affect QOL of elderly patients. Since elderly people have already reduced number of stem cells in bone and cartilage, we need to know the sufficient conditions for osteogenesis and chondrogenesis by comprehensively screening various conditions. We developed a marker gene system expressing green fluorescence protein (GFP) under the control of osteoblast- and chondrocyte-specific promoters. This system helps us monitor osteoblast and chondrocyte differentiation easily, precisely and non-invasively. Using this system, we are now trying to find the sufficient conditions for osteogenesis and chondrogenesis, and to discover osteogenic and chondrogenic small compounds. PMID- 15651366 TI - [Diseases other than Parkinson's disease presenting with parkinsonism]. AB - Among the diseases causing parkinsonism, drug-induced parkinsonism is important because physicians often use anti-dopaminergic drugs such as benzamides for gastrointestinal disorders. In 28 patients with drug-induced parkinsonism, 13 (47%) showed persistent parkinsonism even 6 months after the cessation of the causative drugs. This means anti-dopaminergic drugs may bring subclinical parkinsonism to the clinical level especially in the elderly. Progressive supranuclear palsy (PSP), corticobasal degeneration (CBD) and multiple system atrophy including striatonigral degeneration were recognized in 2003 as specific diseases by the National Health Insurance of Japan. The diagnostic criteria of PSP (the modification of NINDS-SPSP criteria) made by the study group of the Ministry of Health, Welfare and Labour (MHWL) of Japan has demonstrated that their sensitivity was 83% and specificity 97% in "probable PSP". Patients with CBD were surveyed among the main neurologic institutions in Japan using the provisional diagnostic criteria There were 151 patients with "probable CBD" and 13 with "definite CBD". Based on these data, the number of patients with PSP and CBD in 2001 was estimated as 6,400 (5.03 per 100,000) and 2,500 (1.93 per 100,000), respectively, in Japan. Neuroprotective drugs hold hope for the treatment of neurodegenerative diseases. Although many agents were reported to be neuroprotective for Parkinson's disease from in vitro studies, pramipexole, a dopamine agonist, is the only one with good clinical evidence based on neuroimaging. Other dopamine agonists as well as various nerve trophic factors may also have the same effects, though further confirmatory studies are necessary. PMID- 15651367 TI - [Malignant lymphoma, multiple myeloma and myeloproliferative diseases in the elderly]. AB - Hematopoietic and immune function tend to deteriorate in the elderly. The incidence of hematologic diseases in the elderly is increasing as the percentage of elderly people in the whole population increases. Acute leukemia, myelodysplastic syndrome, malignant lymphoma, multiple myeloma, and myelodysplastic syndromes are commonly seen in the elderly. Malignant lymphomas are frequently seen in the elderly, and many elderly patients have poor performance status, and because they are more likely to suffer from impaired cardiac, respiratory, hepatic and renal function, as well as glucose intolerance, they are also more likely to suffer side effects due to chemotherapy. Particularly in patients aged over 80 years, to avoid side effects it is essential to adjust dosage and route of administration of chemotherapy. Although age is a significant negative prognostic factor for non-Hodgkin's lymphoma, it is possible for patients to enter complete remission with improvement of host-side factors. The clinical application of Rituximab is expected to improve chemotherapy outcomes in elderly B-cell lymphoma. The median age at the time of initial diagnosis of multiple myeloma (MM) is 60-70 years, and age is a negative prognostic factor. Clinically, higher rates of infection and heavy comorbidity are characteristic of this condition in the elderly. Although the incidence of bony lesions in elderly patients with MM is not different from the non-elderly, they do have a higher incidence of bone pain and pathologic fractures compared with the non-elderly patients. As the response to chemotherapy is good in the elderly, it is worth trying chemotherapy for MM. Polycythemia vera must be treated in the elderly, because chemotherapy decreases the incidence of thrombosis. PMID- 15651369 TI - [Prevention and antiaging therapy for skin aging]. PMID- 15651368 TI - [Pathomechanism of androgenetic alopecia and new treatment]. AB - Hair follicles are composed primarily of epithelial and dermal components that develop from embryonic ectoderm and mesoderm respectively. The hair growth cycle is coordinated with complex processes that are dependent on the interactions of epithelial and dermal components. Beard and frontal scalp dermal papilla cells (DPCs) show the characteristics of androgen target cells. These DPCs expressed androgen receptor and type II 5alpha-reductase mRNA. To understand the mode of androgen action in human hair follicles, we developed an in vitro co-culture system using DPCs and follicular keratinocytes. Androgen significantly stimulated the proliferation of keratinocytes co-cultured with beard DPCs, suggesting that these DPCs produce androgen-dependent diffusible growth factors. Insulin-like growth factor-I (IGF-I) was identified as one of the androgen dependent paracrine growth factors from beard DPCs. On the other hand, we identified inhibitory roles of androgen on the growth of keratinocytes co-cultured with DPCs from human balding frontal scalp, when DPCs were transfected with the AR expression vector. This inhibitory effect was mediated by TGF-beta1 from the DPCs. Minoxidil and Finasteride were recently introduced for the treatment of androgenetic alopecia in Japan, and TGF-beta1 is the next target for innovative treatment. PMID- 15651370 TI - [Age-related alteration of visual function and its prevention]. PMID- 15651371 TI - [Mechanisms of age-related hearing loss and strategies for protection and restoration of hearing]. AB - Hearing impairment is the most frequent disability in elderly individuals; however, we have limited options for treatment of age-associated hearing loss. To establish novel strategies for the treatment of age-associated hearing loss, it is crucial to elucidate the mechanisms of age-associated hearing loss. Studies on animal models and human temporal bones have indicated a close relationship between degeneration of the spiral ganglion neurons and stria vascularis and age associated hearing loss. We have developed a drug delivery system using biodegradable gel for the inner ear to protect inner ears against aging. In addition, recent studies on cell therapy for the inner ear have suggested the efficacy of cell transplantation for restoration of hearing. PMID- 15651372 TI - [The mechanism of skeletal muscle aging]. PMID- 15651373 TI - [Molecular mechanism of age-related osteoporosis]. AB - Bone loss by ageing has been investigated from standpoints of systemic abnormality and some deficiency in osteoblastic bone formation. This seminar summarizes the involvements of a key molecule of adipocytic differentiation PPAR gamma, essential IGF-I signaling molecules IRS-1 and IRS-2, and an anti-aging gene klotho in the pathophysiology of age-related osteoporosis. PMID- 15651374 TI - [Longevity signals are conserved between invertebrates and mammals]. PMID- 15651375 TI - [Parkin protein and molecular mechanism of Parkinson's disease]. PMID- 15651376 TI - [How does bone mass decrease with advancing age?]. PMID- 15651377 TI - [Wnt coreceptor low density lipoprotein receptor related protein 5 (LRP5) mediates the bone formation and glucose induced insulin secretion]. PMID- 15651378 TI - [History and perspective of geriatric medicine]. PMID- 15651379 TI - [Measures for preventing the elderly from becoming the condition requiring care]. PMID- 15651380 TI - [Community based intervention study for prevention of falling among the elderly]. PMID- 15651381 TI - [Involvement of public health organizations at prefecture level in services conducted by municipalities for the purpose of preventing the long-term care use of older persons]. PMID- 15651382 TI - [Implication and future issues in disability prevention]. PMID- 15651383 TI - [Comparison of activities of daily living for a convalescent rehabilitation ward and general ward for stroke patients]. AB - We investigated changes in the Activities of Daily Living (ADL) of stroke patients in a convalescent rehabilitation ward and a general ward using a Functional Independent Measure (FIM). The subjects were 109 patients hospitalized for rehabilitation purposes at the Oyamada Memorial Spa Hospital. The change in FIM at the time of hospitalization and that at 1 week later was investigated in 81 patients in the convalescent rehabilitation group (CRG) and 28 patients in the control group (CG). In addition, the CRG was investigated again after one month. Intensive rehabilitation service based on ADL and worksheets was introduced in the CRG. On the other hand, these were not introduced in the CG. The total score of FIM increased significantly (p<0.01) in the first week after hospitalization in both groups. The FIM-gain after one week in the CRG was high. With regard to each item, a significant improvement was observed in patients' motor skills while eating, grooming, bathing, dressing the upper body, dressing the lower body, toilet, bladder management, transfer bed/chair, toilet and tub, and walking/wheelchair (11/13). Multiple regressions were used to assess the relationships between FIM-gain (one week, one month), age, rehabilitation intensity and other predictive variables. Better rehabilitation outcomes were observed in patients with lower level of dementia and high rehabilitation intensity. It was thought that planned rehabilitation based on ADL was effective in the CRG, and it was suggested that the CRG's system is effective in the rehabilitation of stroke patients. PMID- 15651384 TI - [A pitfall in analyses of relevant factors concerning phenomena in the process of initiating dialysis: alterations in the characteristics of subjects diminish the significance of age]. AB - In our previous studies, relevant factors concerning the main phenomena related to the process of initiating dialysis were examined in elderly patients with chronic renal failure. Examined phenomena were as follows: (1) the acceptance of dialysis; (2) the urgency of initiating dialysis; (3) short-term outcome; (4) returning home. Multivariate logistic regression analysis was used to determine relevant factors. Although we speculated that age should be a relevant factor for each phenomenon, the phenomenon on which age had some impact was only the first. We suspected the existence of a pitfall, through which the relation of age was lost in the second, the third, or the fourth phenomenon. The fact that every phenomenon had its own relevant factors was thought to be an important clue to the discovery of pitfalls. Relevant factors were derived from both the number of dropout-patients and their demographic and clinical status. From the viewpoint of nondropout-patients, the progression of the process of initiating dialysis might alter the characteristics of subjects for successive phenomena In this study, we set out to investigate whether alterations in the characteristics of subjects were pitfalls. Alterations were regarded as a fall of the mean age, an increment of the rate of the patients with ability to walk, and an increment of the rate of the patients with normal cognitive function. In addition, the old-old patients tended to have limited numbers of those who had the ability to walk and normal cognitive function. In other words, aging changes in ambulatory and cognitive function were not brought to subjects. These alterations may cause the loss of the relation of age to each phenomenon. Thus, we presumed these alterations to be pitfalls. We must clarify whether aging changes are brought to subjects beforehand in analyses that include the old-old patients as subjects. PMID- 15651385 TI - [Problems and therapeutic strategy for emergency operation of the abdomen in the aged]. AB - AIM: To study the problems and therapeutic strategies for emergency abdominal operations in elderly patients by comparing aged and non-aged who underwent an emergency operations for abdominal diseases. MATERIALS AND METHODS: The patients were divided into two groups: those 70 years old or more (85 cases, elderly) and those under 70 years old (228 cases, non-elderly). The type of disease, preoperative complications, postoperative course and outcome of main diseases such as ileus and perforation peritonitis were investigated. RESULTS: Regarding emergency diseases in the aged group, ileus, peritonitis and acute appendicitis were frequently found, in that order, and accounted for 79% of the cases. Incarceration of hernia was more frequently found than in the non-elderly group. On the contrary, acute appendicitis was more frequently found in the non-elderly group. Preoperative complications were found in 46 cases (54%), and were most prevalent in hypertension, cardiac diseases, and diabetes mellitus in this order. In the non-elderly group, preoperative complications were found in 59 cases (26%). Postoperative complications were found in 52 cases (61%) in the elderly group and in 21 (9%) in the non-elderly group. Respiratory complications were found most frequently, and were seen in 32% of the elderly cases. Respiratory complications were significantly more frequent in the elderly group than in the non-elderly group. Death following the operation was found in 9% in the aged group which was mostly due to ileus and peritonitis. CONCLUSION: With regard to the acute abdominal diseases in the elderly patients, both diagnosis and indications of surgery for ileus and peritonitis should be adequately done. Preparation for postoperative respiratory complications and cardiovascular diseases might help to save lives of the elderly. PMID- 15651386 TI - [Analysis of factors affecting prolonged hospitalizations in elderly patients with community-acquired pneumonia]. AB - To determine the factors involved in prolonged hospital stay of elderly patients (over 60 years old) with community-acquired pneumonia (CAP), we investigated: (1) the age, (2) underlying diseases, (3) severity of pneumonia, and (4) length of hospital stay etc. of such cases at a 19-bed local facility. Of the 30 elderly patients with CAP, 20 had no underlying diseases (Group A). The mean length of hospital stay for these patients was about 10 days (mean age 69.1 years). The other 10 patients examined had some chronic underlying diseases, and these patients were assigned to Group B. The mean length of hospital stay in this group was 17 days (mean age was 71.2 years). The finding that the mean length of hospital stay was significantly shorter in Group A than in Group B indicates that prolonged stay of elderly patients with CAP is associated with the presence of underlying chronic diseases. When the severity of pneumonia of patients with underlying chronic diseases increased by one level according to the Guidelines of the Japanese Respiratory Society (2000), the number of patients with moderate pneumonia was 13, and the mean length of hospital stay was 16 days. The number of patients with mild pneumonia was 17, and the mean length of hospital stay was 10 days. Thus, the length of hospitalization was significantly shorter in the mild group than in the moderate group. PMID- 15651387 TI - [Rupture of a thoracic aortic aneurysm in a patient with hypertension, myocardial infarction, hyperlipidemia and operation for abdominal aortic aneurysm rupture after follow-up for 28 years]. AB - A Japanese man who died at age 86 had been followed since the age of 58, when he presented with hypertension of 150/95 mmHg. The patient remained socially active until he died suddenly of a ruptured thoracic aortic aneurysm, although he experienced angina pectoris in August 1974, and myocardial infarction was identified on electrocardiography in October 1974. He underwent operation for rectal cancer in 1987, and an abdominal aortic aneurysm 38mm in diameter was identified at that time. The patient underwent an operation for rupture of the abdominal aortic aneurysm in 1991. A thoracic aneurysm of 40 mm diameter was identified in 1995, and this expanded to 53 mm by 1997. Autopsy revealed a thoracic aortic aneurysm in the arch (8 x 5 x 5 cm) and descending aorta (7 x 7 x 8 cm). A large volume (2,080 ml) of bloody pleural fluid was present PMID- 15651388 TI - [A case of interstitial pneumonia caused by gosha-jinki-gan]. AB - A 74-year-old woman was admitted to the hospital of Ehime University School of Medicine because of slight fever, dry cough and dyspnea on exertion. Chest X-ray films on admission showed diffuse infiltrations in both lungs. Drug-induced pneumonia due to the herbal medicine Gosha-jinki-gan was suspected, as Gosha jinki-gan had been administered for 5 months for the treatment of a right knee pain. Her symptoms and the X-ray abnormality improved after cessation of administration. The lymphocyte stimulation test against Gosha-jinki-gan was positive. To the best of our knowledge, this is the first case of interstitial pneumonia caused by Gosha-jinki-gan. PMID- 15651389 TI - [New horizon of the treatment of pancreatic cancer]. PMID- 15651390 TI - [Fundamental knowledge for understanding FDG-PET]. PMID- 15651391 TI - [Diagnosis of gastrointestinal tract malignancies using positron emission tomography (PET) with 18F-fluorodeoxyglucose (FDG)]. PMID- 15651393 TI - [A case of early esophageal adenocarcinoma arising from cardiac glands, resected by endoscopic mucosal resection]. PMID- 15651392 TI - [Influence of urease activity of the oral cavity and oropharynx on 13C-urea breath test]. AB - We examined whether 13C-urea breath test with powdered 13C-Urea is affected with urease activity in the oral cavity and oropharynx in H. pylori negative cases. We also examined whether the degree of gargling affects the results of 13C-UBT. The results demonstrated that the urease activity not only in the oral cavity, but in the oropharynx may influence the UBT value. Therefore, it is possible that the UBT value may be affected with relic urease activity in the oropharynx if only the oral cavity is gargled. In conclusion, Gargling of the oropharynx in addition to the oral cavity is important to obtain true negative diagnosis with 13C-UBT in patients without Helicobacter pylori infection. PMID- 15651394 TI - [Protein-losing gastroenteropathy associated with Sjogren syndrome-case report and review of the Japanese literature]. PMID- 15651395 TI - [An autopsy case of hepatocellular carcinoma with pleural metastasis]. PMID- 15651396 TI - [A case report of diffuse hepatic angiosarcoma]. PMID- 15651397 TI - [A case of diffuse type of hepatocellular carcinoma with tumor thrombosis of inferior vena cava (Vv3) and main trunk of portal vein (Vp4), showing marked response to transcatheter arterial chemoembolization with degradable starch microspheres (DSM-TACE)]. PMID- 15651398 TI - [A case of gastric sarcoidosis with remarkable imaging features on multislice CT]. PMID- 15651399 TI - [Hyper-carcinoembryonic antigen with pancreatic cancer]. PMID- 15651400 TI - Changes in left ventricular volume and predictors of cardiac events after endoventricular circular patch plasty. AB - OBJECTIVE: The aim of this study was to identify predictors of cardiac events after endoventricular circular patch plasty (Dor operation) by analyzing our experience with Dor operation. METHODS: Thirty patients with left ventricular aneurysm and/or ischemic cardiomyopathy who underwent Dor operation were included in this study. Hemodynamic and clinical results were analyzed, and the predictors of cardiac events were examined. RESULTS: Hospital mortality was 3.3%. Postoperative clinical status and left ventricular (LV) function in all survivors significantly improved. The survival rates at 1, 3, and 5 years after operation were 93%, 89% and 89%. The corresponding cardiac event-free rates were 75%, 67% and 49%. Pre- and postoperative LV function and volume did not differ significantly between patients with or without cardiac events. However, the proportion of reduced end-diastolic volume index (EDVI) (preoperative EDVI postoperative EDVI) to preoperative EDVI was significantly higher in patients with cardiac events than in cardiac event-free patients. Postoperative LV volume re-increased in the cases with cardiac events during follow-up. Cox regression analysis confirmed that preoperative clinical premature ventricular contraction and end-systolic volume index (ESVI), postoperative EDVI, ESVI, and ejection fraction were independent predictors of late cardiac events. There was a significant positive correlation between preoperative ESVI and postoperative EDVI. CONCLUSION: Though LV function significantly improved after Dor operation, LV reconstruction with excessive reduction can cause restarting LV remodeling and increasing mortality and morbidity. Therefore, LV reconstruction of appropriate sizes and shapes, considering the function of residual myocardium, has a significant effect on prognosis. It is highly reasonable to expect that preoperative ESVI can predict the optimal size of reconstructed left ventricle. PMID- 15651401 TI - Rationale of off-pump coronary artery bypass grafting for left main trunk disease. AB - OBJECTIVE: Off-pump coronary artery bypass grafting (OPCAB) remains controversial in patients with left main trunk (LMT) disease because of a concern about the ability to tolerate hemodynamic instability. This study examined the safety of OPCAB for LMT disease compared with conventional coronary artery bypass grafting (CABG). METHODS: Between April 1997 and December 2002, 257 consecutive patients with LMT stenosis who underwent CABG were enrolled. There were 98 patients who received CABG with the aid of cardiopulmonary bypass (CCAB group), and 159 patients who received OPCAB (OPCAB group). RESULTS: There was no patient who converted to on-pump intraoperatively due to hemodynamic instability. Both intraoperative blood loss and blood transfusion incidence were lower in the OPCAB group. Postoperative course was similar, however, pulmonary complications were less observed postoperatively in the OPCAB group. No hypoperfusion syndrome was seen postoperatively in both groups. The average number of anastomosis was 3.2+/ 1.1 in the CCAB group and 3.2+/-1.0 in the OPCAB group (p=0.645). Total arterial OPCAB with an aorta no-touch technique was achieved in 142 patients (89.3%) in OPCAB group. Postoperative angiography was performed in 95 patients in CCAB (96.9%), and in 141 patients in OPCAB (89.8%). Although graft patency of arterial grafts was good in both groups (100% in CCAB and 98.3% in OPCAB), saphenous vein graft patency was slightly lower in both groups (93.4% in CCAB and 76.5% in OPCAB) compared with arterial grafts. CONCLUSION: OPCAB allows a safe and effective treatment of LMT disease. PMID- 15651402 TI - Cystic lymphangioma confined to mediastinum in an adult. AB - Mediastinal lymphangiomas are very rare tumors among the slow-growing mediastinal masses in the literature. We present the successful resection of a 52-year-old woman who was referred to our hospital. Past medical history consisted of surgical treatment for cervical mass and pathological diagnosis of lymphoma nine years earlier. She underwent postoperative radiotherapy. Preoperative chest roentgenogram and computed tomogram of the chest showed a cystic mass in the anterior mediastinum. The tumor was completely resected. The preoperative diagnosis was never questioned until the histopathological examination confirmed that the lesion was a cystic lymphangioma. Cystic lymphangiomas are benign tumors with the evidence of progression in tumor size and invasion into the vital structures. As our case shows, the tumor involvement with the vital structures causes difficulty in removal. PMID- 15651403 TI - Inverted L-shape sternotomy as a minimally invasive approach: re-do cardiac surgery for papillary muscle rupture. AB - We report a successful treatment of the complete papillary muscle rupture occurring 16 months after coronary artery bypass grafting (CABG). A 57-year-old man was admitted for the sudden onset of chest pain and cardiogenic shock. Emergency cardiac catheterization revealed severe mitral regurgitation and total occlusion in the right coronary artery, which was successfully revascularized by percutaneous coronary intervention under intra-aortic balloon pumping. The right internal thoracic artery grafted to the left anterior descending artery in the previous CABG was functioning well. An echocardiogram distinctly indicated the ruptured head of the papillary muscle. Since an emergency operation revealed complete rupture of the posterior papillary muscle, mitral valve replacement was carried out through an inverted L-shape sternotomy with T-shape left atriotomy. Our case indicates that the inverted L-shape sternotomy was a useful approach to preserve the function of grafts, and that T-shape left atriotomy offered a good exposure of the mitral valve in the limited surgical field. PMID- 15651404 TI - Thymic carcinoma with a large cystic lesion. AB - A 59-year-old man presented with a large anterior mediastinal mass. A computed tomography (CT) and magnetic resonance imaging showed a well-circumscribed cystic mass, 12 cm in size adjacent to the heart border and superior vena cava (SVC). A CT guided needle biopsy was performed, and instead of detecting malignant tissues but finding that gray muddy fluid was suctioned, suggesting cystic teratoma. At surgery, the tumor was confirmed advanced thymic carcinoma with pleural dissemination, then the tumor was extirpated with resection of SVC, followed by 2 cycles of chemotherapy. Histologically, the cystic wall was lined with malignant cells. We herein present a diagnostic pitfall case of thymic carcinoma having a large cystic component with which rare association should be recognized. PMID- 15651405 TI - Rapid occurrence of pulmonary aspergillosis after pulmonary wedge resection. AB - A 61-year-old man with diabetes mellitus undertook wedge resection of pulmonary tuberculoma in the left upper lobe. He was discharged uneventfully. However, he became febrile with productive cough five weeks after the discharge, and was readmitted. Imaging studies showed a cavitary lesion at the formerly resected site and multiple infiltrates in the other lobes. A diagnosis was rapidly deteriorating pulmonary aspergillosis. While medical treatments helped his general condition to improve, pulmonary shadows remained unchanged. Finally, we successfully treated him firstly, by performing left upper lobectomy and secondly, by treating multiple infiltrates with postoperative medical treatments. Multiple infiltrates improved gradually and diminished one month after surgery. He remains well without relapse for ten years after surgery. PMID- 15651406 TI - Omental herniation through the esophageal hiatus mimics mediastinal lipomatous tumor. AB - A 61-year-old man with a mediastinal abnormal mass on computed tomography is presented. Sagittal sections of magnetic resonance imaging (MRI) clearly indicated the continuity of the fatty mass from the abdomen to the thorax. The diagnosis was an omental herniation through the esophageal hiatus during the operation. First, we returned the omentum into the abdominal cavity, and then repaired the hiatus. The patient had an uneventful postoperative recovery. A herniation of the omentum through the esophageal hiatus is rare; this case is the tenth found in both English and Japanese literatures. The coronal and sagittal planes of MRI were useful in distinguishing the herniation of omentum through the esophageal hiatus from lipomatous tumor. It is our intention of raising awareness about the disease. PMID- 15651407 TI - Surgical treatment for abnormal echoes in the left ventricular outflow tract caused by ruptured mitral chordae tendineae. AB - We performed surgical treatment for abnormal echoes in the left ventricular outflow tract caused by ruptured mitral chordae tendineae. An asymptomatic 68 year-old man had a chordal rupture exhibiting a tumor-like lesion in the left ventricular outflow tract on echocardiography. Considering a high level of mobility of the tumor as well as its texture, the risk of embolization was found to be significant. Therefore, it was decided on the surgical resection of the tumor-like lesion under extracorporeal circulation. We report a highly rare case of rupture in redundant mitral chordae tendineae without mitral regurgitation, revealed by uncommon echoes in the left ventricular outflow tract during systole. PMID- 15651408 TI - Surgical repair for chronic traumatic thoracic aneurysm after 12-year follow-up. AB - A 39-year-old man underwent surgical repair of chronic traumatic thoracic aneurysm after 12-year follow-up at our hospital. Eighteen years prior to surgery, he had been involved in a traffic accident, suffering a left hemopneumothorax. Chronic traumatic thoracic aneurysm is extremely rare that few guidelines for surgical intervention exist for this disorder. However, it has been observed that all patients with new symptoms should be operated promptly, and that asymptomatic aneurysm detected over 2 years after the initial trauma can be monitored by careful follow-up pending symptomatic or radiologic change. The present case provides additional support for these strategies. PMID- 15651409 TI - Left ventricular rupture following mitral valve replacement due to oversize prosthesis. AB - Left ventricular rupture following mitral valve replacement is one of the most serious complications. We report our experience in successful treatment of type III left ventricular rupture following mitral valve replacement probably due to an oversize prosthesis. A 67-year-old woman, with the history of percutaneous transluminal mitral commissurotomy 11 years previously, underwent mitral valve replacement for mitral restenosis with a 27 mm CarboMedics mechanical bileaflet valve (Sulzer CarboMedics Inc., Austin, TX, U.S.A.). There were some difficulties in placing the entire prosthesis into the annulus at the posterior because of the oversize prosthesis. After the complete placement of the prosthesis, bulge of the left ventricular muscle was evident around the left lateral region. Following the cessation of cardiopulmonary bypass, type III left ventricular rupture, half a circular rip between the papillary muscles and posterior mitral annulus, occurred. The rip was suture-closed and a 23 mm CarboMedics valve was placed. Postoperative ultrasonic cardiography showed no prosthetic stenosis, periprosthetic leak, left ventricular pseudoaneurysm, nor left ventricular asynergy. Under cardioplegic arrest, we should not select the oversize prosthesis to prevent left ventricular rupture. PMID- 15651410 TI - Stenting for critical airway stenosis under percutaneous cardiopulmonary support. AB - Airway stenting for severe central airway stenosis is inherently a dangerous procedure. There is the risk of critical airway obstruction due to bleeding, tumor debris, and airway perforation during the procedure. Once such situations occur, percutaneous cardiopulmonary support (PCPS) can be one of the most valuable rescue options to prevent critical hypoxic complications. At our institute, four of 49 patients who received stenting or other airway intervention required PCPS support (8%). Two of these cases required PCPS to be performed in an emergency setting during the procedure while the procedure was elective in the other 2. All procedures were performed effectively and safely without any complications caused by PCPS, including massive airway bleeding due to anticoagulant treatment. Patients were able to be weaned off PCPS uneventfully. PCPS is considered to be a valuable procedure in remedying critical hypoxic situations during airway intervention. PMID- 15651411 TI - [Bacterial RND-type multicomponent efflux systems: the efflux systems involved in production of pathogenic factors as well as resistance to antimicrobial agents]. PMID- 15651412 TI - [Atomic structure and function of the efflux-pump machinery]. PMID- 15651413 TI - [Structure and function of the type III secretion system in pathogenic bacteria]. PMID- 15651414 TI - [Type III secretion systems and bacterial pathogenicity]. PMID- 15651415 TI - [Type IV secretion system in Helicobacter pylori: comparison with bacterial type III secretion apparatus]. PMID- 15651416 TI - [Type IVB secretion systems of intracellular pathogenic bacteria]. PMID- 15651417 TI - [Mechanisms for the determination of the cell division plane]. PMID- 15651418 TI - [Life of proteins]. PMID- 15651419 TI - [DNA manipulation using artificial restriction enzymes]. PMID- 15651420 TI - [Sports and cardiorespiratory function]. AB - Regular physical training is generally considered to help prevent numerous diseases. These beneficial effects of aerobic exercise are particularly well established with regard to cardiovascular and respiratory functions. However, exhaustive exercise is clearly a risk in some athletes who have not undergone a correct and complete assessment of cardiovascular and respiratory functions. Exercise training is a crucial tool in integrative rehabilitation programs for patients with chronic heart failure and respiratory diseases. PMID- 15651421 TI - [Metabolic functions and sport]. AB - Current epidemiological studies emphasize the increased of metabolic diseases of the adults, such as obesity, type-2 diabetes and metabolic syndromes. Even more worrying is the rising prevalence of obesity in children. It is due more to sedentariness, caused more by inactivity (television, video, games, etc.) than by overeating. Many studies have shown that regular physical activities benefit various bodily functions including metabolism. After dealing with the major benefits of physical exercise on some adult metabolic disorders, we focus on the prime role played by physical activity in combating the public health problem of childhood obesity. PMID- 15651422 TI - [Overtraining syndrome]. AB - Numerous studies have shown that enhanced physical load can result in an overtraining syndrome, with a decreased capacity for physical exercise and behavioral disturbances. Overtraining is caused by an imbalance between energy intake and output, and is facilitated by chronobiological and psychological stress. These factors are responsible for hormonal changes such as a decrease in gonadal steroids or in the hypothalamo-pituitary-adrenal axis. These metabolic and hormonal influences are lead to change in brain neuromediator activity, such as reduced monamine and increased serotonin levels. Experimental data indicate that these neuromediator changes are responsible more for behavioral changes than for decreased physical performance. PMID- 15651423 TI - [Doping and dietary supplements]. AB - Dietary supplements are widely promoted and used all over the world as a means of losing weight, increasing energy and athletic performance, or for compensating for true or supposed deficits. Traditionally, dietary supplements have been defined as products composed of one or more essential nutrients such as vitamins, minerals, and protein. But, in 1994, the US Congress broadened the definition to include (albeit with some exceptions) any product intended for ingestion as a supplement to the diet. This includes vitamins, minerals, herbs, botanicals, and other plant-derived substances; amino acids and substances such as enzymes, hormones, and metabolites. Some dietary supplements contains substances prohibited by anti-doping rules or may pose a health risk to some persons. Positive cases of doping can be found in three cases: amphetamine-like compounds that have potentially powerful stimulant effects on the nervous system and heart; hormones and pro-hormones; and intentional contamination of dietary supplements with hormones (mainly anabolic steroids). French and EU laws protect French consumers from adverse events related to the use of these products. Nevertheless, lack of controls in some European countries, Internet sales, and poor knowledge among some health care professionals create a persistent risk of use of fake products. PMID- 15651424 TI - [Blood boosting today]. AB - The last Winter Olympics and recent cycling events have once again called attention to the use by endurance sportsmen and women of a variety of approaches designed to increase the blood hemoglobin concentration and oxygen-carrying capacity. An increased hemoglobin concentration and maximal oxygen uptake can be achieved by training at high altitude or by "living high and training low". It can also be procured by "blood boosting" with erythropoietin or darbepoietin, blood infusion, or misuse of hemoglobin-based oxygen carriers. This paper describes methods of increasing oxygen transport, with the accent on blood infusion, which is again relevant today with the development of a test able to detect recombinant human erythropoietins. PMID- 15651425 TI - [The fight against doping: today and tomorrow]. AB - Since Antiquity, sport persons have used doping methods to improve their performance. The methods used today are increasingly specific and professional As a corrolary, doping is becoming extremely difficult to detect: because many prohibited drugs are rapidly metabolized (short half life); it is often impossible to distinguish between the natural and recombinant hormone products; several prohibited substances are not screened for; therapeutic use authorization (TUA)--a recent recommendation from the World antidoping agency (WADA)--and advances in gene therapy are giving cheaters new possibilities. It is necessary to promote a new approach to doping detection by determining potential specific links between changes in human biological profiles (proteom, transcriptom, metabonom), and the use of a given drug. Developments in nanotechnology, and notably" labs on a chip" should prove useful in this respect. PMID- 15651426 TI - [Children of the war]. AB - Children are always concerned by wars. Physical consequences (death, severe wounds, disability) are surely important, but physiological trauma can also be grave. We studies the impact of war on two groups of children aged between 6 and 10 years. The first group consisted of children living in Lebanon during the war, and the second was composed of children born in Paris of Lebanese migrant parents. PMID- 15651427 TI - [Pulmonary hypertension associated with systemic lupus erythematosus]. AB - Pulmonary hypertension (PH) is a rare but potentially life-threatening complication of systemic lupus erythematosus (SLE). We reviewed the literature on this complication, its pathogenesis and clinical presentation, and treatment options. PH is reported in 0.5% to 14% of patients with SLE. The literature describes the cases of 105 patients, 90% of whom were female. Average age was 33 years, and overall mortality was 25 to 50% two years after PH diagnosis. As in patients with primary pulmonary hypertension, dyspnea is the most common presenting symptom of PH in patients with SLE. Up to 58% of patients with both PH and SLE have Raynaud's phenomenon. Echocardiography can show right ventricular hypertrophy and dilation, even before symptom onset. Right-heart catheterization, with an assessment of vasoreactivity, is a necessary part of the work-up and is also needed for treatment decision-making. PH in patients with SLE is associated with intimal hyperplasia, smooth-muscle hypertrophy and medial thickening, similar to the changes seen in primary PH. Several pathological mechanisms have been proposed for PH associated with SLE. They include vasoconstriction, vasculitis, thrombosis, anticardiolipin antibody and lupus anticoagulant. Endothelial dysfunction may be an important factor in the onset of PH, possibly by contributing to vasospasm. Higher serum endothelin levels are found in patients with SLE and pulmonary hypertension than in other SLE patients. Several specific antibody patterns have been reported in patients with PH and SLE. Oral calcium channel blockers are indicated for patients who respond to acute NO challenge. Continuous intravenous prostacyclin represents a therapeutic advance, although it appears less effective than in primary PH. Some patients have been improved by new oral endothelin receptor antagonists, usually combined with intensive immunosuppressive therapy. Patients with SLE have an increased risk of PH. Vigilance is therefore required to detect early signs of PH. Early diagnosis allows treatment to start before irreversible vascular lesions occur. PMID- 15651428 TI - [Depression and aging]. AB - Depression is one of the most common health disorders in elderly people. It is still often considered as a natural consequence of aging, arising in reaction to a medical disease, cognitive or functional decline, or a loss of social fabric. Many studies have highlighted the low rates of diagnosis and treatment of depression, especially in the primary care setting. Major depression in old age is characterized by the same core symptoms as in other periods of life. However, aging may accentuate some symptoms and alleviate others. Somatic concern, marked anxiety, poor subjective memory, psychotic ideation, and recurrent thoughts of death can mask sadness and anhedonia. Organic factors and adverse life events are often intricately linked with the pathogenesis of depressive states in the elderly. The role of cerebrovascular lesions has also been established, particularly in late-onset depressive disorders. The management of depressive disorders in older people, as in younger adults, involves pharmacological and psychological treatments. Electroconvulsive therapy can be beneficial in some cases. Transcranial magnetic stimulation is being evaluated in this setting. PMID- 15651429 TI - [Antibiotic-loaded bone cements: from laboratory studies clinical evaluation]. AB - Antibiotic-loaded bone cements (ALBC) were initially used empirically, both for the treatment of infected prostheses and for antibioprophylaxis. We conducted in vitro and animal studies as a prerequisite for proper clinical evaluation. We measured gentamicin diffusion from methacrylate bone cement, and found that the concentration in surrounding fluid was significantly above the minimal inhibitory concentration (MIC). We then implanted ALBC in the proximal femora of 100 ewes, in conditions close to those of total hip replacement (THR). Antibiotic concentrations in bone were above 4 x MIC for more than 6 months. We subsequently measured antibiotic concentrations in drainage fluid, blood and urine samples from 50 patients undergoing THR. Concentrations were over 20 x MIC in periprosthetic fluid, but were below the detection limit in blood after 24 hours and in urine after one week. The blood concentration was less than 1 mg/l--far below the ototoxic and nephrotoxic threshold (8 mg/l). Having established antibiotic bioavailability, we then examined the clinical efficacy of ALBC. We participated in several studies, including a French multicentric review of 342 THR procedures (direct and two-stage exchanges). ALBC was beneficial in both settings, with an 85% infection control rate. This included patients with very severe infections (multiresistant strains, severe osteolysis) treated with two stage exchanges; and moderate infections treated by direct exchange. (This latter procedure offers fewer complications, more durable functional results, a shorter hospital stay and lower treatment costs.) We also used ALBC for antibioprophylaxis in over 2000 THR procedures, and noted no complications. This method is now being assessed in more than 200 000 cases contained in the Swedish THR Register. The experimental methods that we developed (laboratory studies, animal implantation, clinical pharmacokinetics) set the groundwork for clinical studies of gentamicin ABLC, and also allowed us to develop other ALBC formats, containing vancomycin, for example (restricted to salvage therapy). We also used these methods to evaluate antibiotic-loaded bone substitutes (calcium and carbon phosphates), designed not only to control infectious osteitis but also to replace osteolytic bone. PMID- 15651430 TI - [Recurrent kidney disease and its prevention after renal transplantation. Review of the literature]. AB - Recurrence of the initial renal disease after kidney transplantation is a major cause of graft dysfunction and failure. Glomerulopathies are reported to account for 80% of such cases, but this figure should be interpreted with care, as extensive investigations (immunofluorescence or electron microscopy studies of the explanted kidney) are needed to prove that the initial disease has indeed recurred and to determine prevalence rates. In the following order of frequency, focal segmental glomerulosclerosis, membranous glomerulonephritis, mesangiocapillary glomerulonephritis and IgA nephropathy have a tendency to recur, while, surprisingly, anti-glomerular basement membrane nephritis and systemic lupus erythematosus are less likely to relapse on the donor kidney. There is no evidence that anti-calcineurin therapy has any impact on the risk of recurrence. Options for the prevention and treatment of these recurrences are very limited, calling for very cautious use of living familial donors. PMID- 15651431 TI - [Spiegelian hernias. Report of 51 cases]. AB - We describe 51 cases of Spiegel's hernias, and report a critical review of the relevant literature. The patients presented with an isolated mass in 17 cases, abdominal pain in 17 cases, and a complication in 14 cases (intestinal occlusion in 8, incarceration in 6). The hernia was discovered fortuitously by compute tomography in three cases. The diagnosis was made preoperatively in 31 cases and during surgery in 17 cases. Sonography was contributory in 10 cases and compute tomography in 6 cases. In 47 cases the deep orifice was located at the level of Douglas' arch, in the anterolateral abdomen. The hernial sack remained interstitial in every case, under the aponeurosis of the external oblique. The contents were irreducible or strangulated in 14 cases. Surgery was necessary in all but three cases. The approach was lateral in 43 cases and medial in 5 cases. The indications of laparoscopy in this setting are discussed. PMID- 15651432 TI - The cellular basis of traumatic bleeding. AB - Bleeding is clearly a major cause of morbidity and death after trauma. When bleeding is attributable to transection of major vessels, surgical repair is appropriate. Posttraumatic microvascular bleeding attributable to coagulopathy secondary to metabolic derangements, hypothermia, and depletion or dysfunction of cellular and protein components requires a different approach. Although transfusion of blood products may be necessary to replace the blood loss, it does not always correct the problem of microvascular bleeding. The type of injury, mode of care, and treatment objectives differ significantly for combat-wounded soldiers versus civilian trauma patients. Although hemorrhage is responsible for 50% of combat deaths, published information about coagulation monitoring among combat patients is very limited. These articles summarize the appropriate monitoring of hemostasis among combat trauma patients, review the unique nature of combat casualties and the medical system used to treat them, and discuss information available from civilian studies. Because the development of coagulopathy is relatively infrequent in the young, otherwise healthy, military population, the routine screening measures currently used are adequate to guide initial blood product administration. However, as new intravenous hemostatic agents are used for these patients, better laboratory measures will be required. Although hemorrhage is the leading cause of death for combat casualties, catastrophic hemorrhage is rarely a prehospital combat medical management problem because, when it occurs, it tends to cause death before medical care can be provided. In civilian environments, most seriously injured victims can be reached and transported by emergency medical services personnel within minutes; in combat, it often takes hours simply to transport casualties off the battlefield. In combat situations, even if the transport distances are small, the hazardous nature of the forward combat areas frequently prevents medical personnel from quickly reaching the wounded. Furthermore, whereas civilian blunt trauma victims may have a "golden hour," casualties with penetrating battlefield trauma often have only a "platinum 5 minutes." Because of the challenges of treating hemorrhage during combat, it is important for military medical personnel to understand their options for treating hemorrhage quickly and efficiently. These articles discuss the causes of posttraumatic microvascular bleeding and the potential treatment options for controlling catastrophic hemorrhage in combat areas. PMID- 15651433 TI - Treating traumatic bleeding in a combat setting. AB - Bleeding is clearly a major cause of morbidity and death after trauma. When bleeding is attributable to transection of major vessels, surgical repair is appropriate. Posttraumatic microvascular bleeding attributable to coagulopathy secondary to metabolic derangements, hypothermia, and depletion or dysfunction of cellular and protein components requires a different approach. Although transfusion of blood products may be necessary to replace the blood loss, it does not always correct the problem of microvascular bleeding. The type of injury, mode of care, and treatment objectives differ significantly for combat-wounded soldiers versus civilian trauma patients. Although hemorrhage is responsible for 50% of combat deaths, published information about coagulation monitoring among combat patients is very limited. These articles summarize the appropriate monitoring of hemostasis among combat trauma patients, review the unique nature of combat casualties and the medical system used to treat them, and discuss information available from civilian studies. Because the development of coagulopathy is relatively infrequent in the young, otherwise healthy, military population, the routine screening measures currently used are adequate to guide initial blood product administration. However, as new intravenous hemostatic agents are used for these patients, better laboratory measures will be required. Although hemorrhage is the leading cause of death for combat casualties, catastrophic hemorrhage is rarely a prehospital combat medical management problem because, when it occurs, it tends to cause death before medical care can be provided. In civilian environments, most seriously injured victims can be reached and transported by emergency medical services personnel within minutes; in combat, it often takes hours simply to transport casualties off the battlefield. In combat situations, even if the transport distances are small, the hazardous nature of the forward combat areas frequently prevents medical personnel from quickly reaching the wounded. Furthermore, whereas civilian blunt trauma victims may have a "golden hour," casualties with penetrating battlefield trauma often have only a "platinum 5 minutes." Because of the challenges of treating hemorrhage during combat, it is important for military medical personnel to understand their options for treating hemorrhage quickly and efficiently. These articles discuss the causes of posttraumatic microvascular bleeding and the potential treatment options for controlling catastrophic hemorrhage in combat areas. PMID- 15651434 TI - Monitoring of hemostasis in combat trauma patients. AB - Bleeding is clearly a major cause of morbidity and death after trauma. When bleeding is attributable to transection of major vessels, surgical repair is appropriate. Posttraumatic microvascular bleeding attributable to coagulopathy secondary to metabolic derangements, hypothermia, and depletion or dysfunction of cellular and protein components requires a different approach. Although transfusion of blood products may be necessary to replace the blood loss, it does not always correct the problem of microvascular bleeding. The type of injury, mode of care, and treatment objectives differ significantly for combat-wounded soldiers versus civilian trauma patients. Although hemorrhage is responsible for 50% of combat deaths, published information about coagulation monitoring among combat patients is very limited. These articles summarize the appropriate monitoring of hemostasis among combat trauma patients, review the unique nature of combat casualties and the medical system used to treat them, and discuss information available from civilian studies. Because the development of coagulopathy is relatively infrequent in the young, otherwise healthy, military population, the routine screening measures currently used are adequate to guide initial blood product administration. However, as new intravenous hemostatic agents are used for these patients, better laboratory measures will be required. Although hemorrhage is the leading cause of death for combat casualties, catastrophic hemorrhage is rarely a prehospital combat medical management problem because, when it occurs, it tends to cause death before medical care can be provided. In civilian environments, most seriously injured victims can be reached and transported by emergency medical services personnel within minutes; in combat, it often takes hours simply to transport casualties off the battlefield. In combat situations, even if the transport distances are small, the hazardous nature of the forward combat areas frequently prevents medical personnel from quickly reaching the wounded. Furthermore, whereas civilian blunt trauma victims may have a "golden hour," casualties with penetrating battlefield trauma often have only a "platinum 5 minutes." Because of the challenges of treating hemorrhage during combat, it is important for military medical personnel to understand their options for treating hemorrhage quickly and efficiently. These articles discuss the causes of posttraumatic microvascular bleeding and the potential treatment options for controlling catastrophic hemorrhage in combat areas. PMID- 15651435 TI - Treating traumatic bleeding in a combat setting: possible role of recombinant activated factor VII. AB - Bleeding is clearly a major cause of morbidity and death after trauma. When bleeding is attributable to transection of major vessels, surgical repair is appropriate. Posttraumatic microvascular bleeding attributable to coagulopathy secondary to metabolic derangements, hypothermia, and depletion or dysfunction of cellular and protein components requires a different approach. Although transfusion of blood products may be necessary to replace the blood loss, it does not always correct the problem of microvascular bleeding. The type of injury, mode of care, and treatment objectives differ significantly for combat-wounded soldiers versus civilian trauma patients. Although hemorrhage is responsible for 50% of combat deaths, published information about coagulation monitoring among combat patients is very limited. These articles summarize the appropriate monitoring of hemostasis among combat trauma patients, review the unique nature of combat casualties and the medical system used to treat them, and discuss information available from civilian studies. Because the development of coagulopathy is relatively infrequent in the young, otherwise healthy, military population, the routine screening measures currently used are adequate to guide initial blood product administration. However, as new intravenous hemostatic agents are used for these patients, better laboratory measures will be required. Although hemorrhage is the leading cause of death for combat casualties, catastrophic hemorrhage is rarely a prehospital combat medical management problem because, when it occurs, it tends to cause death before medical care can be provided. In civilian environments, most seriously injured victims can be reached and transported by emergency medical services personnel within minutes; in combat, it often takes hours simply to transport casualties off the battlefield. In combat situations, even if the transport distances are small, the hazardous nature of the forward combat areas frequently prevents medical personnel from quickly reaching the wounded. Furthermore, whereas civilian blunt trauma victims may have a "golden hour," casualties with penetrating battlefield trauma often have only a "platinum 5 minutes." Because of the challenges of treating hemorrhage during combat, it is important for military medical personnel to understand their options for treating hemorrhage quickly and efficiently. These articles discuss the causes of posttraumatic microvascular bleeding and the potential treatment options for controlling catastrophic hemorrhage in combat areas. PMID- 15651436 TI - Circle of Midwives. PMID- 15651437 TI - HIV and midwifery. PMID- 15651438 TI - Midwifery in the media. PMID- 15651439 TI - Another Jewish midwife's perspective on Israel. PMID- 15651440 TI - Do you do anything to prevent preterm and/or postterm pregnancies? If so, what? If not, why not? PMID- 15651441 TI - Marion's message. A time to be born. PMID- 15651442 TI - A timely birth. PMID- 15651443 TI - Prematurity is preventable! PMID- 15651444 TI - The tree and the fruit. Routine versus selective strategies in postmaturity. PMID- 15651445 TI - Antioxidants and preeclampsia. PMID- 15651446 TI - GBS, pregnancy and garlic: be a part of the solution. AB - In 2002 the Centers for Disease Control (CDC) wrote new GBS protocols that call for the 4,000,000 pregnant women in the US each year to have a rectal/vaginal culture for GBS at 36 weeks. The protocols recommend that the 800,000 women per year who will have a GBS-positive result receive IV antibiotics during labor, costing $20,000,000 a year, at a low estimate of $25 per woman. This will prevent the deaths of about 100 infants a year from early onset GBS, while 80 newborns a year will still die of the disease. It will prevent about 4,000 (60% of the total) newborn early onset GBS infections. Garlic kills GBS, but because no profit can be made from its use, no research exists on the use of garlic to prevent GBS in newborns. Women are encouraged to consider following the protocols described in this article at 35 weeks and culturing for GBS at 36 weeks, as a proactive way to research the use of garlic to prevent newborn GBS disease. PMID- 15651447 TI - Does mother nature really make mistakes? PMID- 15651448 TI - The power of words. PMID- 15651449 TI - HIV basics. PMID- 15651450 TI - Just how long is "normal" for labour? PMID- 15651451 TI - The 30-minute third stage. PMID- 15651452 TI - The birth of a midwife (with a long latent phase). PMID- 15651453 TI - Freedom from fear. PMID- 15651454 TI - The business of midwifery coding I. PMID- 15651455 TI - Cultural lack of birth experience empowers media representations, not women. PMID- 15651456 TI - Holistic evaluation of healing after cesarean birth. PMID- 15651457 TI - To vaccinate or not to vaccinate: rubella and the adult woman. PMID- 15651458 TI - South Africa missive. PMID- 15651459 TI - Standard term of pregnancy. AB - What is it? This question arose early this year during a discussion with my doula partner, who lived in Great Britain for 20 years, and one of the midwives I work with, who attends only homebirths, which is very rare in France. The mom whose case we were discussing was late going into labor but not postdated according to the official pregnancy term here in France (37-42 WA--"weeks of amenorrhea"). The midwife expressed her discomfort with waiting. My doula partner and I felt differently, but we knew we were influenced by American and British midwives' practices. I had been shocked the previous December by the position of the chief of the maternity department in a private hospital (a small unit with no residents, in which nurse-midwives attend "normal" births and obstetricians are called in case of complications only). This OB explained to my client and me that if she didn't go into labor naturally, she would be called at 41 + 1 for a vaginal exam to check her cervix and would be induced at 41 + 2. Waiting until 42 weeks requires daily checks, for which he has neither the room nor personnel. He clearly stated that it was a matter of management of time and finances. A month later one of our clients reported the story of her brother and sister-in-law's planned homebirth in London. Their doctors had put a lot of pressure on the mother during her pregnancy with gestational diabetes regarding her length of term. They started to talk about induction. The parents didn't feel comfortable with this, and at that point our client had asked me to refer them to someone who could help them there. We referred them to the National Childbirth Trust and to the sweetest doula we know there. This doula (Liliana Lammers) and her famous doctor partner (Dr. Michel Odent) were a good match. The doctor advised waiting, on the condition that the health of the baby and the amount of fluid be checked daily at the local hospital. The mother had already been waiting several weeks past what was supposed to be her term. Finally, she went naturally into labor at home. The doctor and the doula came and, after some hours of observation, decided it would be wiser for the mother to deliver in the hospital. The doctor and doula were not comfortable with the prolonged prelabor, when, at nearly 44 weeks, the health of the baby and the amount of fluid had not been checked for five days. The mother finally had a vaginal birth without drugs at the hospital. After hearing this story, I suggested it would be interesting to collect the official lengths of term and the different routines in other countries as a learning tool and in order to give us something other than French protocol on which to base our practice. So I sent the question to every midwife for whom I had an e-mail address from the last Midwifery Today conference in Paris. Beyond this motivation was my own curiosity regarding the relationship between the official term in each country and its uses and routines. The most significant (because the most unique) answer, in my opinion, is from The Netherlands, where physiology is a priority. PMID- 15651460 TI - Priya's birth. PMID- 15651461 TI - Berta Juarez Fuentes. PMID- 15651462 TI - I believe it is time to go. PMID- 15651463 TI - Promotion of the use of a thermoplastic material to aid in the identification of unidentified human remains. AB - There has been a recent effort to promote the use of a thermoplastic bite impression material for the identification of children. The American Board of Forensic Odontology is a certification board for forensic dentists in the United States and Canada. It is the position of the ABFO that this technique is of limited value when used for the dental identification of children. A presentation was made to the Odontology Section of the American Academy of Forensic Sciences in Dallas, Texas, in February 2004. The presentation promoted the use of a thermoplastic material to record the morphology of the human dentition for use by qualified personnel to identify missing persons. Currently, the target population for the recording of this information is children. In addition to recording the morphology of the dentition, the presenter claimed that the material, properly stored after impressing the teeth, could also be used as a source of DNA for identification purposes, as well as a source of the individual's scent to be used by tracking dogs. The weeks and months following this presentation generated significant comments and concerns regarding the information presented amongst members of the ABFO and members of the Odontology Section of AAFS. It was suggested that the ABFO look into this tooth impression method and comment on its validity and usefulness in the identification process. The following comments regarding this method have been formulated following significant input from diplomates of the ABFO. The ABFO is a certifying organization for dentists who wish to become board certified in the field of forensic odontology, and as such, requires its diplomates to maintain significant experience in the use of the human dentition for identification purposes. The ABFO also has a mission to promote and enhance the science of forensic odontology. As the ABFO primarily deals with the dentition, the comments in this paper will be confined to the use of the thermoplastic material as it relates to the dental identification process. Experts in DNA analysis and tracking should address the issues of DNA and scent. PMID- 15651464 TI - The impact of mercury on the environment. PMID- 15651465 TI - Dr. Jack F, Conley retires from CDA Journal. PMID- 15651466 TI - Airborne spread of disease--the implications for dentistry. AB - The potential for the airborne spread of disease has been recognized for many years. Recent studies have shown that this mode for disease transmission is capable of spreading a fatal disease such as Severe Acute Respiratory Syndrome over a wide area. Many dental procedures produce extensive aerosols and splatter that are routinely contaminated with bacteria, viruses, and blood. In the past, the potential for these aerosols and splatter to be a vector for disease spread has not been emphasized in dental infection control. Recently published data shows a need to reassess the potential for dental aerosols and splatter to spread disease and the need for their control. Simple and inexpensive methods for the control of dental aerosols and splatter are given. Dental personnel are urged to make the control of aerosols a standard part of their infection control procedures. PMID- 15651467 TI - Infection control compliance issues and questions. AB - This article uses the 2003 CDC infection control guidelines for dentistry as a framework for discussing representative questions and issues that continue to be raised by dental health care workers. Where applicable, additional supporting evidence will be incorporated to provide appropriate, useful information, to assist in understanding and complying with updated recommendations. PMID- 15651468 TI - 2003 CDC guidelines offer more choices for managing operatory surfaces. AB - The objective of this article to help dental professionals understand the changes in surface asepsis recommendations, to be able to classify environmental surfaces in the practice setting and successfully prevent or manage their contamination, as well as become familiar with the terminology used in discussing environmental surfaces and related infection control efforts. PMID- 15651470 TI - Dr. Conley has left the building. PMID- 15651469 TI - Looking inside the 2003 CDC dental infection control guidelines. AB - On Dec. 19, 2003, the Centers for Disease Control and Prevention published updated infection control guidelines for dentistry. The guidelines provide comprehensive information on all aspects of dental infection control. The recommendations are designed to prevent or reduce the potential for disease transmission from patient to dental health care personnel, from dental health care personnel to patient, and from patient to patient. Most recommendations will be familiar and are already practiced routinely. This article highlights major updates and additions in the CDC guidelines and provides additional information to assist readers in applying the latest guidelines. Almost a year ago, the CDC and Prevention published updated dental infection control guidelines in a supplement to the Morbidity and Mortality Weekly Report. The Guidelines for Infection Control in Dental Health Care Settings--2003 represent a collaborative effort between leading experts in infection control from other federal agencies, public health, and hospital epidemiology and infection control. Unlike regulatory agencies such as the Occupational Safety and Health Administration, the U.S. Food and Drug Administration, or the U.S. Environmental Protection Agency, the CDC cannot mandate certain practices; it can only recommend. However, the CDC is recognized as the nation's disease prevention agency and develops a broad range of guidelines intended to improve health care and to inform clinicians and the public. As a result, many dental licensing boards adopt CDC's recommendations, or variations of them, as the infection control standard for dental practice in their states. In contrast to the 1986 and 1993 CDC dental infection control recommendations, the 2003 CDC publication includes more background information and the scientific rationale for the recommendations. Also, readers will notice that each recommendation has a rank assigned to it categorizing the recommendation on the basis of existing scientific data, theoretical rationale, and applicability (Table 1). Most recommendations will be familiar and already are practiced routinely. As with previous CDC recommendations, the guidelines are designed to prevent or reduce the potential for disease transmission from patient to dental health care personnel; from dental health care personnel to patient, and from patient to patient. The following is an overview highlighting major updates and additions in the 2003 CDC guidelines. It is not intended to be a comprehensive review. Readers can access the complete document (Figure 1) by visiting www.cdc.gov/oralhealth/ infectioncontrol. PMID- 15651471 TI - [What should surgeons know about inflammatory pulmonary disease?]. PMID- 15651472 TI - [Pulmonary infection in immunocompromised hosts]. AB - While typical pulmonary infections can be cured with antimicrobial agents, three types require surgical lung resection: those in immunocompromised patients; those with acquired resistance to medication; and those caused by microorganisms against which there are no effective drugs. We discuss these three types from the viewpoint of physicians. With the development of chemotherapy for malignant disease, patients with leukemia can be cured with bone marrow transplantation. During the leukopenia accompanying chemotherapy, Aspergillus sp. can infect the lungs. Aspergillus infections are resistant to antimicrobial agents, and thus surgical resection is necessary. Aspergillus infections may occur in previous sites of pulmonary tuberculosis lesions after the tuberculosis is cured producing massive hemoptysis. In this case, surgical resection is also needed. When patients who are immunocompromised due to various underlying diseases become infected with multidrug-resistant tuberculosis, they require surgical resection. Finally, when lesions of nontubercular mycobacterial infection are found, these patients also require surgical lung resection. PMID- 15651473 TI - [Surgical treatment for pulmonary mycosis]. AB - Although most human mycosis develops in immunocompromised patients as an opportunistic infection, its frequency will be influenced by historical changes in diseases and by progress in medical technology. An example of local occurrence is pulmonary aspergilloma in an open negative cavity, and a systemic example is the onset of invasive pulmonary aspergillosis in neutropenic patients with hematologic malignancies. Surgical candidates for mycotic lung disease are usually patients who do not improve with medical treatment The author also stresses the prophylactic indications for prevention of unexpected impairment due to mycotic lung disease. The usefulness of surgery for pulmonary fungal infection has gradually been recognized Additional efforts should be made by surgeons to achieve better outcomes and to minimize operative complications by choosing the appropriate surgical technique. PMID- 15651474 TI - [Role of surgical treatment in atypical Mycobacterium avium infection of the lung]. AB - Mycobacterium avium complex (MAC) disease has recently become an increasingly significant problem. MAC is frequently difficult to treat because of its drug resistance. Therefore surgical treatment still plays an important role in the treatment of this disease. Lobectomy has mainly been performed, and the postoperative complication rate is relatively high (up to 32%). The rate of relapse after surgical treatment is reported to be satisfactorily low (less than 20%). Early pulmonary resection is beneficial in patients whose disease is still localized and who can tolerate resection surgery. PMID- 15651475 TI - [Surgical treatment of pulmonary tuberculosis]. AB - The indications for the surgical treatment of pulmonary tuberculosis are mainly for multidrug-resistant tuberculosis (MDR-Tb). In Fukujuji Hospital there have been 91 pulmonary resections for 83 MDR-Tb cases during the past 20 years. Of those resections, upper lobectomies with or without partial resection of other lobes comprised 55%, segmentectomies 10%, and pneumonectomies 30%. The following major postoperative complications occurred: prolonged air leakages in 19%, empyema in 9%, and respiratory failure in 6%. However, there were no operative deaths. After pulmonary resection for MDR-Tb, immediate negative conversion rate of expectoration of tuberculous bacilli was 98%, reexpectoration rate of them was 12%, The final cure rate was 92%. Surgical treatment is worth consideration for the treatment of refractory pulmonary tuberculosis. PMID- 15651476 TI - [Pathogenesis and treatment of chronic empyema]. AB - In chronic empyema (CE), thickened pleura, collapsed chest wall, and the accumulation of purulent fluid in the thoracic cavity are typical findings. Patients complaints of symptoms with bronchopleural fistula (BPF). On the other hand, there is another type of CE in which the pleural space expands progressively to shift the neighboring lungs, mediastinum, and diaphragm. This type of CE is considered to be chronic expanding hematoma (Reid et al.) occurring in the thoracic cavity. In the empyemic cavity, mycobacterial infection is found approximately in 20-30% of cases, pyogenic bacillus or fungus in about 40%, but the cavity is aseptic in other 30-40%. Although the fundamental treatment procedures include decortication and pleuropneumonectomy, the method of muscle or omental plombage to manage dead space or BPF are far superior functionally in intractable CE. Recently, the methods of plastic and reconstructive surgery have been used to utilize the muscle or omentum more effectively. The classic thoracoplasty procedure should not be undertaken unnecessarily to avoid additional deterioration of respiratory function. Additionally, it should be remembered that malignant lymphoma occurs frequently in the empyemic chest wall. PMID- 15651477 TI - [Perioperative lung injury: acute exacerbation of idiopathic pulmonary fibrosis and acute interstitial pneumonia after pulmonary resection]. AB - The mortality rate after surgical resection for lung cancer has been reported to range between 1% and 3%, with 30% caused by acute exacerbation of idiopathic pulmonary fibrosis (IPF) or acute interstitial pneumonia (AIP). Approximately 20% of patients with IPF have lung cancer, while 2% to 4% of lung cancer patients have IPF. The incidence of postoperative acute exacerbation of IPF is about 20%. Some investigations in Japan revealed that 10% to 17% of lung cancer patients undergoing lung resection, who have not been diagnosed with IPF preoperatively, have localized-usual interstitial pneumonia (Lo-UIP) lesions. Approximately 20% of patients with Lo-UIP show postoperative acute exacerbation, while about 0.5% of those without Lo-UIP develop AIP after surgery. There is no confirmed treatment or prophylaxis. Most patients who develop postoperative acute exacerbation or AIP are treated with methylpredonisolone (1,000 mg/day x 3 days), but the mortality rate is 50% or greater. We emphasize that more efforts should be made to develop strategies to prevent postoperative acute exacerbation of IPF and AIP. PMID- 15651478 TI - [Robotic surgery in neurosurgical field]. AB - Computer-aided surgery commenced in the late 1980s when computer was clinically used for diagnosis and surgical planning. Since then the computer has been used in a surgical navigation system. In the early 1990s a robotic surgery using intelligent manipulator as surgeon's new hands took place. Nowadays intraoperative diagnostic imaging as surgeon's new eyes has become ubiquitous Diagnosis, surgical planning, and navigation are required to be real-timely performed intraoperatively. The time has really come to concurrently diagnose and treat, in which technology visualizing intraoperative medical information and minimally invasive surgery are fused. For that it is necessary to develop a system that real-timely updates information for decision making, and at the same time to present the timely, optimum treatment to be done according to the results of instant evaluation of ongoing treatment. To realize and support above system it is essential to combine a sensor which can precisely distinguishes a focal area from a normal tissue intraoperatively, and a manipulator which participates the treatment. In addition, the manipulator should be accurately controlled using a computer (computer-aided manipulation) according to the surgical plan made by a method aided by a computer (computer-aided design) based on intraoperatively acquired information. It is about to change quality of life to quality of treatment. PMID- 15651479 TI - [What can we learn from a case of medical malpractice]. PMID- 15651480 TI - [Hydatid cyst of the kidney]. AB - Hydatidosis is an endemic disease caused by the larval form of Echinococcus Granulosus. Renal involvement represents less than 5% of confirmed cases. It remains clinically silent for a long time, and only presents at the stage of complications. Diagnosis is suspected on the basis of epidemiological, clinical, radiological and biological data. There are various clinical presentations. Hydaturia, which is observed in 10 to 30% of the cases, is the only pathognomonic feature. Diagnostic accuracy has been improved since the wide use of ultrasonography. Computed tomography and magnetic resonance imaging are helpful tools to confirm the diagnosis. The treatment is mainly based on surgery. The resection of the prominent dome remains the standard option as it allows preservation of the kidney. Total nephrectomy should be proposed only in case of renal destruction. The percutaneous management, which includes puncture, aspiration, injection, and reaspiration, can be performed in very selected cases. However, the results of this technique are still under debate. PMID- 15651481 TI - [Tissue engineering in urology]. AB - Tissue engineering refers to the techniques that are aimed at regeneration of human tissues and organs. Two elements are necessary for these techniques: matrix and cells. Matrix is the scaffold where tissues may organise. Cells are either autologous cells stimulated to regenerate in vivo, aided by implantation of matrix ("guided tissue regeneration"), or autologous cells cultured outside the body (in vitro) and later returned as auto-transplants. All types of conventional tissue reconstructive surgery need tissue engineering. These techniques have been introduced recently into the clinical practice. One of the main limitations of reconstructive surgery in genitourinary tract is the lack of autologous tissue. Two autotransplants could be distinguished: coherent tissue structure or cell suspensions. The great number of studies published in this area emphasizes the importance of the future clinical implication in urology. PMID- 15651482 TI - [Recent traumatic injuries of the urethra]. AB - Traumatic injuries of the urethra are uncommon. Most Lesions of the anterior (bulbar urethra) are straddle injuries and are initially dealt with by a suprapubic catheter with delayed treatment when urethral stenosis does ensue. Traumatic disruption of the posterior urethra is in most cases related to a pelvic fracture and is often associated with multiple life-threatening injuries, which receive priority treatment. Management of posterior urethral disruption remains a highly controversial issue: alternative treatments include early endoscopic realignment, early open surgical repair and suprapubic catheter and delayed open surgical repair. Management of urethral injuries is described and the different operative techniques are detailed. PMID- 15651483 TI - [Penis tumours: techniques and indications]. AB - Although penis epidermoid carcinoma is a relatively rare tumour, early identification and treatment are necessary to avoid mutilating and sometimes morbid surgery. Awareness of disease epidemiology is mandatory for preventing the evolution of an underlying tumour (absent or insufficient hygiene, congenital or acquired phimosis, preepitheliomathous lesions such as Bowen's disease). Conservative surgical or radio-therapeutic techniques may be considered, provided the lesion is superficial and with a diameter < 30 mm. In all other cases, penis amputation is necessary. Penis epidermoid carcinoma spreads by vascular and lymphatic diffusion. Dynamic scintigraphy for the identification of a sentinel node and screening of subclinical metastasis is currently under evaluation. In case of palpable inguinal adenopathy, inguinal lymphadenectomy should be proposed if technically performable. Although superficial lymphadenectomy is associated with a low morbidity rate (about 3% of mild complications) deep inguinal Lymphadenectomy is far more morbid and disabling. Compliance with pre- and post operative measures, and total patient compliance are necessary to minimize as far as possible such morbidity. PMID- 15651484 TI - [Augmentation enterocystoplasty]. AB - Augmentation enterocystoplasty refers to a technique that consists in removing a bowel segment in order to suture it onto the bladder. This technique is indicated in case of reduced bladder capacity and/or compliance, in case of failure of conservative treatments. The goal is to improve the patient's urination comfort, but above all to ensure long-term protection of the upper urinary tract. All bowel segments may be utilized but the ileum is the segment of choice. The selected digestive segment must be detubulized in order to better decrease its peristaltic contractions and obtain low-pressure urine storage. Bi-valving the bladder while preserving the detrusor usually performs augmentation enterocystoplasty. However, in case of very fibrous and thickened detrusor, a supra-trigonal cystectomy should be considered. The digestive segment is removed and detubulized, then it is sutured on to the bladder as a patch at the incision level. Following such surgery, over 90% of the patients report significantly improved quality of life. Nocturnal bladder competence is obtained in more than 90% of the patients, while 91 to 100% report diurnal bladder competence. Long term complications may be observed, such as chronic infections with asymptomatic bacteruria (70% of the cases) not necessitating any treatment. Within the two first years, there is a 10% to 15% risk for stone formation following intestinal mucus development. Enterocystoplasty perforation may occur at a frequency estimated to range from 5 to 13%, which is a very serious and life-threatening complication. Similarly, a risk of enterocystoplasty cancer may be observed after five to ten years, in 1% of the cases. PMID- 15651485 TI - [Benign prostatic hyperplasia: CPG, PSA and 5-alpha-reductase inhibitors]. PMID- 15651486 TI - [Clinical practice guidelines (CPG): are they useful? Example of the benign prostatic hyperplasia (BPH)]. AB - Clinicians develop routine strategies with multiple and sometimes hardly relevant origins. The logical theoretical solution would be to make available systematically drafted clinical practice guidelines (CPG) with the aim of helping the physician during daily practice. These CPG summarize the best available proofs and relate each recommendation to a level of proof. However, if the CPG is not rigorously drafted, their usefulness is questionable. The AGREE scale, translated into several languages and used by several european countries, including France (ANAES), is useful for the evaluation of the quality of drafting of a CPG but also as a guide when making a CPG. The analysis of different CPG for the management of benign prostatic hyperplasia (HBP) originating from different countries and companies shows important variations which are best explained by large differences in the rigour of drafting. Besides, whatever the quality of the CPG, they too often neglect the key element which is its application in practice. A survey of the modalities of prescription of subsequent examinations in France in HBP demonstrates an important disparity between CPG and routine attitudes in clinical practice. A potential solution may require a deeper involvement of the learned urologic society in the drafting and distribution of the CPG. PMID- 15651487 TI - [Combination therapy in benign prostatic hyperplasia (BPH)]. AB - Because of the constant progress in our understanding of the physiopathology of benign prostatic hyperplasia, it is now possible to propose a more rational use of combination therapy, which is often used empirically, though not recommended, in routine practice. This chronic disorder, which is in fact more complex than it appears, may benefit in theory from a combination of molecules with different complementary action mechanisms. Prostatic obstruction may be treated either by alphablockers with their peripheral muscle-relaxant action on the smooth muscle fibers of the prostate and bladder neck, or by 5-alpha-reductase inhibitors for their reducing effect on gland volume. Irritative bladder symptoms involving the detrusor may be treated by antimuscarinics and to a lesser extent by alphablockers. Currently available data from recent studies suggest that a combination of an alphablocker and a 5-alpha-reductase inhibitor may be useful in patients with symptomatic BPH and a prostate of more than 40 grams with PSA > 1.6 ng/ml. Combinations including an antimuscarinic are effective in patients with BPH with marked irritative symptoms. Finally, the combination of an alphablocker and a phosphodiesterase type 5 inhibitor may be useful in patients with lower urinary tract symptoms (LUTS) associated with erectile dysfunction. PMID- 15651488 TI - [The PCPT study: the viewpoint of an urologist involved]. AB - The PCPT study investigating the possibility of decreasing the incidence of prostate cancer by daily administration of 5 mg of finasteride for 7 years appears to be a major clinical study in the management of this disease. Recently published in the New England Journal of Medicine, it makes difficult reading. The current article aims to demonstrate the main methodological characteristics of this study, to describe the most significant results and to discuss its clinical applications in daily urological practice. PMID- 15651489 TI - [Use of 5-alpha-reductase inhibitors in the prevention of prostate cancer]. AB - 5-alpha-reductase inhibitors are now in widespread use for the treatment of benign prostatic hyperplasia (BHP) and these molecules have recently come under the spotlight in prostate cancer. Their peripheral "hormonal" action inducing reduced intraprostatic DHT synthesis seems to involve them in this hormone dependant disorder. Finasteride evaluated in the treatment of BPH (PLESS study) was found to have a preventive effect on the incidence of cancer and this activity was assessed in a specific trial (PCPT study). Nevertheless, in the latter randomized study with a 7-year follow-up period, a reduction in the global incidence of the number of cases of cancers was associated with an increase in the number of high-grade cancers. A slight reduction in prostate cancer was also noted in the studies with dutasteride in BPH (ARIA3001, ARIA3002 and ARIB3003). An international multicenter study (REDUCE) is currently being conducted to confirm the preventive value of this molecule which has a more complete activity than finasteride with its inhibitory action on the two 5-alpha-reductase iso enzymes, and may therefore have a clearer efficiency and rule out the risk of onset of high-grade cancer. PMID- 15651490 TI - [PSA and benign prostatic hyperplasia]. AB - In the opinion of many urologists, prostate-specific antigen (PSA), which is not specific of any disease, is not considered in the diagnosis and follow-up of benign prostatic hyperplasia (BPH). Nonetheless, prostate cancer diagnosis is based on PSA screening. PSA value is thus available for a majority of men above 50. Recent data suggest that there is a link between PSA value and BPH natural history, turning PSA into a tool for BPH management and prevention of complications such as retention. It hasn't got into practice so far, as recommended criteria for PSA screening in BPH have not been issued. The debate over this topic is only at its beginning. PMID- 15651491 TI - [Potential sexual consequences associated with benign prostatic hyperplasia and its treatments]. AB - In this article, an updated review of the potential consequences of treatment of benign prostatic hyperplasia (BPH) on sexual function is proposed in the light of recent data. Differences exist between the different therapeutic options available for the treatment of lower urinary tract symptoms (LUTS) when we want to analyze the impact on sexuality. Surgical procedures including minimally invasive techniques are associated with ejaculation and also erection disorders. Drug treatments also have repercussions on sexuality though there are significant differences between the various classes of these drugs. In daily practice, the relation between the prostate and sexuality must be assessed during any medical examination for one of these diseases and before any decision on treatment. PMID- 15651492 TI - [Arguments for deciding to change treatment in benign prostatic hyperplasia]. AB - Insufficient efficacy above all, but also the appearance of side effects or the occurrence of complications may motivate a change in the treatment of lower urinary tract symptoms (LUTS) related to benign prostatic hyperplasia (HBP). In terms of efficacy, surgery remains superior to all medical treatments including alphablockers which are the most active. 5-alpha-reductase inhibitors have a slow efficacy which is all the more marked when the prostatic volume is large. Phytotherapy has a slight activity which is higher than that of placebo. Drug combinations are currently being studied and, although a benefit has been shown in certain publications, their clinical efficacy remains to be specified. They may become an alternative in case of failure of single-agent therapy to prevent or delay surgery. The incidence of side effects is low and varies with the treatment used and within a same therapeutic class. With alphablockers for instance, orthostatic hypotension, which is the main side effect, varies from 1% to 8% approximately depending on the study and molecule. The side effects of 5 alpha-reductase inhibitors are mainly sexual disorders, observed in 1 to 4% of cases approximately. Hence, the occurrence of a drug side effect causing treatment discontinuation should incite a change of molecule, even within the same class for alphablockers. On the other hand, if complications of HBP occur, it is then necessary to resort to surgery. PMID- 15651493 TI - [Bladder sphincter disorders in Parkinson's disease]. AB - Parkinson's disease related to degeneration of the extrapyramidal structures is characterized in its typical form by the classic triad of tremors, rigidity and akinesia, constituting Parkinson's syndrome. These are combined with neurovegetative disorders, responsible for sexual and bladder sphincter dysfunction. The Latter occurs in from 30% to 90% of cases, depending on the stage of progression of the disease. The dopamine deficiency in the nigrostriataL tract leads to a lifting of the inhibition which is probably the cause of the bladder hyperreflexia. Irritative signs are most frequently observed. The flowmetry data are contradictory, depending on the publication whereas the bladder overactivity, objectified by cystomanometry, is described in most of the studies although certain authors report, on the contrary, bladder hypoactivity. Conflicting data have also been published on bladder sphincter dyssynergia although in most studies, micturition was described as synergic in Parkinson's disease patients. L-dopa, the main drug for Parkinson's disease, has, according to the publication, either no action, or contradictory effects with bladder hypoactivity or hyperactivity. Anticholinergics are effective on overactive bladder, the alphablockers on the urethral hypertonia at the price of a higher risk of arterial hypotension in this diathesis. The indication for prostatic surgery must be carefully considered and preceded by precise clinical, urodynamic and sometimes electromyography evaluation. There is a high risk of post surgical incontinence. Endourethral prostheses provide an attractive alternative in the case of prostatic obstruction. PMID- 15651494 TI - [Dutasteride: main results in the treatment of benign prostatic hyperplasia]. AB - Benign prostatic hyperplasia (BPH) is a progressive disease characterized by an increase in prostatic volume. The resulting urinary irritative and/or obstructive symptoms, when present, often reduce the patient's quality of life. Prostatic dihydrotestosterone, the androgen produced following transformation of testosterone by the enzyme 5-alpha-reductase, is responsible for these changes. Among various products used for drug therapy, 5-alpha-reductase inhibitors play a specific role in the long-term management of BPH. Between the two currently available products, dutasteride is the only dual 5-alpha-reductase inhibitor (types 1 and 2). The efficacy of dutasteride was tested in several scientific studies and the cumulative results of three controlled, double-blind pivotal studies, showed at 2-year a clear improvement in all parameters modified during this disorder. A significant decrease in the severity and frequency of the urinary symptoms, in prostate volume, the risk of urine retention, the necessity to resort to surgery and an increase in peak urinary flow were observed with dutasteride. These results were confirmed in the 4-year-open label extension of these pivot studies both for tolerability of the product and the maintenance of its efficacy over time. PMID- 15651495 TI - [Intimate partner abuse of women: identification of victims in medical practice]. AB - Intimate partner abuse is world-wide much more prevalent than often suspected. Three female patients consulted their family doctor with a variety of complaints. A 53-year-old woman, mother of three grown-up children, and divorced a year ago, sought help after four days, for a large infected wound of her right hand. A 32 year-old single mother of a 2-year-old girl asked the practice assistant for an iteration of a tranquillizer because of tension headache. A 36-year-old divorced mother of three sons was sent by her company doctor to ask for a referral to a psychiatrist because of chronic fatigue. All three women appeared to have been severely abused by their partners, recently or in the past, and were reluctant to disclose the abuse. Two women were regular visitors of doctor's offices with diverse unspecific complaints. The third woman was a victim of sexual abuse as a child and was severely depressed. Active asking about experiences of intimate partner abuse helped these women to disclose the real nature of their problem. Important effect is that abused women can accept help and experience support to improve their situation. Doctors should suspect intimate partner abuse more often in patients who are heavy medical consumers, depressed or victims of sexual abuse in childhood. Single and divorced mothers can hide a history of abuse. Doctors should ask clear and specific questions about relationships and listen without passing judgement so as to help women to disclose abuse. PMID- 15651496 TI - [The treatment of tobacco dependence in clinical practice is promising but difficult to implement]. AB - According to the Dutch interdisciplinary guideline 'Treatment of tobacco dependence', medical practitioners in The Netherlands should play a more active role in discouraging the use of tobacco. They should try to motivate their patients to stop smoking and offer their support. The various treatments aimed at helping people to stop smoking require large amounts of personnel, time and money. Clear guidelines and follow-up programmes are still missing. Much will depend on a strict anti-smoking policy from the government and the availability of financial support in order to effectively reduce the amount of smoking-related illness. At present, an optimistic view in this respect does not seem realistic. PMID- 15651497 TI - [Transplantation of the islets of Langerhans in patients with diabetes mellitus type 1]. AB - Since 1921 and until recently, insulin by injection has been the only treatment for patients with diabetes mellitus type 1. After pancreas transplantation, which became possible in 1977, the next logical step to cure patients with diabetes mellitus type 1 is the transplantation of the islets of Langerhans. In the last few years, the results of islet transplantation are markedly improved thanks to developments in the isolation technique and better immunosuppressive protocols. Ongoing problems in islet transplantation are allo-immunity, auto-immunity and the growing shortage of donor pancreases. Alternatives to pancreas donation, be it post-mortem or from a living donor, could be: sources for islets are xenotransplantation with the aid of pig islets and beta-cell neogenesis from embryonic stem cells or pancreatic duct cells. PMID- 15651498 TI - [The practice guideline: 'Treatment of tobacco dependence']. AB - The interdisciplinary guideline 'Treatment of tobacco dependence' discusses the approach to smoking patients. The point of departure is the concept that smoking is an addiction and that its treatment should be based on this fact. The effectiveness of treatment depends on the patient's motivation to stop smoking and on the intensity of the intervention. Pharmacotherapy may be of added value here, particularly for persistent smokers. The guideline aims at reaching as many smoking patients as possible, regardless of whether there is a relationship between their symptoms or ill health and smoking. Practitioners should ask about smoking habits regularly and smokers should be advised at least once to stop. Further treatment and guidance aims in particular to increase the motivation to quit and at those who are prepared to stop on the basis of the stages-of-change model. More intensive possibilities for treatment, by both general practitioners and specialists, are needed for persistent smokers. Such facilities are hardly available at all in The Netherlands at present, but will have to be developed to make the cost-effective treatment of tobacco addiction possible on a large scale. PMID- 15651499 TI - [Diagnostic image (221). A female multitrauma patient]. AB - A 39-year-old woman who was involved in a high-energy trauma suffered a haematopneumothorax, subcutaneous emphysema, multiple fractures including a fracture of corpus Tx, and air in the epidural space. PMID- 15651500 TI - [Use of oral contraceptives in the years 1994-2002: different but not less]. AB - OBJECTIVE: To describe the use of contraceptives among women aged 10-59, particularly in relation to the type of oral contraceptives (OCs) among starting users and women already using OCs. DESIGN: Descriptive. METHOD: Data on the use of medication in the period 1994-2002 by women in the age range 10-59 years were selected from the Interaction Database of North and East Netherlands. The study population was 33,795 women in 1994 and 102,894 in 2002. Cross-sectional studies were performed of prevalent and incident OC users per year and the contribution of the various types of OC was measured. RESULTS: The use of contraceptives among women aged 10-59 years has remained fairly constant since 1994, being about 33%. However, Dutch women started using OCs at an increasingly younger age: 47% of the 15-19-year-olds in 2002 compared with 35% in 1994. The use of third-generation OCs among young starting OC-users (10-19 years of age) in 2002 was very low (3.5% of all OCs). Among women who already used OCs, the switch from third- to second generation OCs was less pronounced. In 2002, 23% of all OC-users used third generation preparations compared with 46.5% in 1994. The proportion using cyproteron-ethinylestradiol increased slightly over the years. CONCLUSION: After 1994, mainly second-generation OCs have been prescribed. The switch from third- to second-generation OCs was especially marked among the young starting users. The use of cyproteron-ethinylestradiol increased slightly during the years. PMID- 15651501 TI - [A modified informed-consent procedure in which the complete information is given retrospectively: no objection from participating patients]. AB - OBJECTIVE: To determine whether a modified informed-consent procedure, in which permission is requested to send a questionnaire and to provide all the information about the study only after it is completed, would raise objections from patients who were asked to participate in a study on satisfaction with the follow-up care after a cerebrovascular accident. DESIGN: Descriptive cohort study. METHOD: 123 patients were approached for the randomised study. The patients who were included were interviewed by telephone two weeks after they had been sent the postponed information. RESULTS: During recruitment, 5 patients refused to participate but this was not because of the modified procedure. Of the 112 survivors, 102 (91%) could be interviewed and 79 had read the letter. Nobody (0%; 95% CI: 0-5) said that their trust in doctors had decreased; one patient (1%; 95% CI: 0-7) indicated decreased willingness to participate in future studies; two patients (3%; 95% CI: 0-9) were dissatisfied about the procedure followed. CONCLUSION: The participants in the trial had no major objections to the modified informed-consent procedure in which full information was given only after their participation had ended. Furthermore, patients did not refuse participation because of the modified procedure. PMID- 15651502 TI - [Persistent fever in a patient with Salmonella typhimurium gonarthritis]. AB - A 65-year-old patient with systemic lupus erythematodes (SLE) developed fever and gonarthritis whilst taking prednisone and hydrochloroquine. Salmonella typhimurium, sensitive to amoxicillin, was grown in cultures of synovial fluid, blood and urine. After high dosages of intravenous amoxicillin, blood cultures and knee punctate became negative, but fever seemingly without a clinical focus persisted. By means of a fluoro-18-deoxyglucose positron emission tomography (FDG PET) scan, an abscess was located in the left upper leg and successfully treated. In patients with impaired cell-mediated immunity, an extra-intestinal manifestation of Salmonella should be taken into consideration particularly if there appears to be no clinical focus. This may be detected by FDG-PET. PMID- 15651503 TI - [Surgical removal of debilitating neurogenic heterotopic ossifications of the hip]. AB - Two men, aged 24 and 52 years, developed neurogenic heterotopic ossifications of the hip, the first following a cervical spinal-cord injury and the second after prolonged artificial ventilation following bowel surgery. The stiffness caused problems when sitting and when performing general daily activities; the second patient who was ambulatory, also had problems walking. CT investigation clearly showed the localisation of the ossifications. After surgical removal of the ossifications both patients were able to sit for longer periods without low-back pain and the ambulatory function of the second patient was much improved. Indomethacin was given postoperatively and no recurrence of the ossification was seen. A number of per- and postoperative complications are associated with the surgical removal of neurogenic heterotopic ossifications of the hip. These include damage to surrounding structures and infection. However taking into account the improved range of movement and increased independence of the patient, it is a useful intervention for this debilitating condition. PMID- 15651504 TI - [How should age be documented?]. PMID- 15651505 TI - [Outbreak of amoebiasis in a Dutch family; tropics unexpectedly nearby]. PMID- 15651506 TI - [Outbreak of amoebiasis in a Dutch family; tropics unexpectedly nearby]. PMID- 15651507 TI - [The practice guideline 'Photo(chemo)therapy and systemic therapy in severe chronic plaque-psoriasis']. PMID- 15651508 TI - Pictorial history of anesthesia at the American University of Beirut. PMID- 15651509 TI - Randomized prospective crossover study of biphasic intermittent positive airway pressure ventilation (BIPAP) versus pressure support ventilation (PSV) in surgical intensive care patients. AB - BACKGROUND AND OBJECTIVE: The aim of this prospective, randomized and crossover study was to assess the role of a relatively new mode of mechanical ventilation, biphasic intermittent positive airway pressure (BIPAP) in comparison to another well established one, pressure-support ventilation (PSV) in surgical intensive care patients. METHODS: 24 generally stable patients, breathing on their own after short-term (< 24 hours) postoperative controlled mechanical ventilation (CMV) were randomized to start on either PSV or BIPAP, and indirect calorimetry measurements were performed after 1 hour adaptation period at two time intervals; immediately after the investigated ventilatory mode was started and 1 hour later. Statistics included a two-tailed paired t-test to compare the two sets of different data, p < 0.5 was considered significant. RESULTS: Oxygen consumption (VO2), energy expenditure (EE), Carbon dioxide production (VCO2), and respiratory quotient (RQ) did not differ significantly between the two groups. There were also no significant differences regarding respiratory rate (RR), minute volume (MV) and arterial blood gas analysis (ABGs). Both modes of ventilation were well tolerated by all patients. CONCLUSION: PSV and BIPAP can be used for weaning patients comfortably in surgical intensive care after short-term postoperative ventilation. BIPAP may have the credit of being smoother than PSV where no patient effort is required. PMID- 15651510 TI - Reducing cardiovascular responses to laryngoscopy and tracheal intubation: a comparison of equipotent doses of tramadol, nalbuphine and pethidine, with placebo. AB - The stress response to tracheal intubation may be obtunded by opioids given with induction of anesthesia. Tramadol is an opioid acting on mu-receptors and the monoaminergic pain modulating systems. This study examined vasomotor responses to tracheal intubation after equipotent doses of tramadol, nalbuphine and pethidine (3.0, 0.3 mg/kg(-1), and 1.5 mg/kg(-1), respectively), and placebo, given prior to induction of anesthesia in 118 healthy patients. Premedication and induction of anesthesia were standardized. Recordings of HR and SAP were made prior and subsequent to induction of anesthesia, and at 1, 3, 5 and 7 minutes after tracheal intubation. Prior to laryngoscopy and intubation, HR increased in all groups (p < or = 01, all comparisons), but least so after nalbuphine, whilst SAP remained unchanged after placebo, tramadol and pethidine, but fell after nalbuphine (p < 0.025). Maximum increases in HR (p < or = 0.005, all comparisons) and SAP (p < or = 0.02, all comparisons) occurred one minute after intubation. Maximum HR after placebo (108 SD 15 bpm), tramadol (107 SD 20 bpm), pethidine (113 SD 16 bpm) and nalbuphine (110 SD 26 bpm) was similar; with placebo HR remained faster than baseline until the seventh minute but had returned to baseline by the fifth minute with the opioids. Maximum SAP with tramadol (151 SD 26 mmHg) was similar to that with placebo (157 SD 20 mmHg), but was greater than after pethidine (136 SD 27 mmHg; p < 0.05) and nalbuphine (135 SD 19 mmHg; p < 0.02). With each test drug SAP returned to baseline by the third minute. It is concluded that, in these doses, 1) tramadol does not attenuate the chronotropic nor the inotropic response to tracheal intubation, and 2) pethidine and nalbuphine reduce only the inotropic response to airway instrumentation. PMID- 15651511 TI - Carotid endarterectomy: a study of cerebral oxygen saturation vs stump pressure. AB - BACKGROUND AND AIMS: Carotid endarterectomy (CEA) is an established surgical procedure for treatment of internal carotid artery (ICA) stenosis. To determine whether or not a carotid shunt is necessary to place, some surgeons measure the stump pressure. We conducted the current study in order to identify whether or not cerebral oxygen saturation (rS02%) can serve as another quantitative measurement to determine the need of carotid shunt during CEA. MATERIALS AND METHODS: Ten patients who underwent CEA under general anesthesia were studied. The stump pressure was measured during ICA clamping and rSO2% was measured during three phases: A) prior to ICA clamping, B) during ICA clamping and C) after ICA declamping. The data were subjected to one-way ANOVA and correlation coefficient analysis. The mean age was 62+/-7 yr and the mean body weight was 66+/-7kg. RESULTS: The stump pressure and rSO2% mean values were 45+/-9 mmHg and 57+/-7% respectively during ICA clamping. Correlation coefficient revealed significant positive relationship, r = 0.724(P = 0.009). CONCLUSIONS: rSO2% may serve as another quantitative measurement to determine the need for carotid shunt during CEA surgery. Due to the small number of cases in the current study, the critical rSO2% which warrants carotid shunt placement could not be identified. Therefore, large number of patients are required to define the critical rSO2% during CEA surgery. PMID- 15651512 TI - Critical incident reports in adults: an analytical study in a teaching hospital. AB - Self-auditing is important to improve the standard of care. In a previous study (1991--1997), we reported human error and equipment factor as the two major causes leading to critical incidents. This study analyzes the computerized database and the medical records of 71 critical incidents reported in the Department morbidity and mortality meeting between 1998--2002, in an attempt to identify factors that have led to critical incidents and compare them with our previous report. PATIENTS & METHODS: We use a standard form where factors which lead to critical incidents are identified such as: anesthesia providers, patient's characteristics, type of anesthesia and others. The form includes a section for the reporter to narrate the whole incident. RESULTS: Seventy-one incident reports in adult patients were studied. In 35.3% consultants reported the incidents, while 64.7% were reported by residents. Most of the incidents were reported in patients undergoing emergency surgery (P = 0.0034) and with physical status III-IV (P = 0.008). Incidents due to circulatory events were 33.8%, while respiratory incidents were 35.2%. The majority of incidents were due to human error (38%), lack of team communication (23.9%), patient condition (10%) and technical problems, (19.7%). Suggestions for prevention of incidents were, better training 29.2%, better communication 20.3%, developing of algorithms 12.6%, checklists 10.2%, better supervision 15.2%, improved equipment 6.3%, improved arrangement of drugs 5.2% and for additional monitoring 1%. CONCLUSIONS: Emergency surgery and patient ASA physical status III-IV were significant predictors of critical incident reports. Analysis of anesthesia related problems may contribute to improved preventive strategies. PMID- 15651513 TI - Pressure and volume changes of tracheal tube cuff following inflation with various inflating agents during nitrous oxide anesthesia. AB - The study was designed to investigate the changes in pressure and volume of a tracheal tube-cuff inflated with air, mixture of N2O + O2, saline and 4% lidocaine during nitrous oxide anesthesia. This study was conducted in 80 patients (33 male & 47 female). The pressure and volume of a tracheal tube cuff increased with air, decreased with mixture of N2O + O2 and almost remained the same with saline and 4% lidocaine. The complications were more in the air group. PMID- 15651514 TI - Simultaneous off-pump coronary revascularization with pulmonary resection or vascular surgery. AB - Patients who require coronary artery bypass grafting and who also have vascular disease or lung malignancy constitute a high-risk group, and their management remains controversial. Combining off-pump coronary surgery (OPCAB) with peripheral artery revascularization or lung resection is an attractive proposition, as it avoids the risks associated with cardiopulmonary bypass. This paper presents the results of 26 such combined procedures consisting of simultaneous OPCAB and peripheral revascularization or lung resection, between April 2001 and March 2003. Twenty underwent concomitant carotid endarterectomy and OPCAB, four underwent aortobifemoral bypass and OPCAB, and two underwent pneumonectomy and OPCAB. There was no in-hospital mortality. Prolonged ventilatory support was necessary in one patient who had a lung resection. The median postoperative length of stay in the Intensive Care Unit was 1.3 days (range 1 to 6) and the median length of hospital stay was 5.7 days (range 4 to 16 days). Off-pump coronary surgery clearly has a place for patients with peripheral vascular disease or pulmonary malignancy who additionally require myocardial revascularization. PMID- 15651515 TI - Toxic epidermal necrolysis syndrome--management and case report. AB - Toxic epidermal necrolysis syndrome [known also as Lyell's syndrome] is a severe drug reaction, characterized by fever. systemic toxicity, extensive skin rash with blisters and exfoliation similar to that of major burns. It carries an average mortality rate of 25%. The data concerning most frequent triggering agents and the differential diagnosis is presented; An overview of the management protocol for these patient in the Burn Center of the Lebanese Hospital is exposed with a case presentation of a young woman who sustained a 95% body surface area [BSA] cutaneous loss after the ingestion of only 2 tablets of amoxycilline documented by the complication and evolution. PMID- 15651516 TI - The effect of ketamine and fentanyl in reducing the pain of diazepam injection. AB - Diazepam is an effective drug that is used widely in modern anesthesia. Venous irritation is one of its major side effects attributed to its acqueous insolubility and requisite solvents. There is some evidence that ketamine may acts as a local anesthetic drug, because of its effect on N-methyl-D-aspartate receptors. Fentanyl also may reduce the pain of diazepam injection by blocking the opiate receptors in vessels walls. To determine the effectiveness of ketamine and fentanyl in reducing the pain of diazepam injection, 150 patients (ASA I, II) were randomly assigned to one of three Groups and before intravenous diazepam injection. 2 ml normal saline, 2 ml fentanyl or 10 mg ketamine were administered for Groups 1, 2, 3 respectively. The pain of diazepam injection was then evaluated at 30 minutes intervals. Our results showed that ketamine and fentanyl reduce the pain of diazepam dramaticaly (p < 0.001) in comparison with placebo. Ketamine is more effective than fentanyl in reducing such pain (p < 0.001). PMID- 15651517 TI - Pain relief after arthroscopic knee surgery--intraarticular sufentanil vs morphine. AB - BACKGROUND: Prevention of postoperative pain by injecting opioid into the knee joint is believed to support the hypothesis of peripheral opioid receptor activation in inflammation. Main outcomes were reduction of pain intensity and need for supplementary analgesics. The aim of the present study was to evaluate the analgesic effects of intraarticular sufentanil in comparison with morphine, following arthroscopic procedures of knee joint. METHODS: In this prospective, double-blind study, 45 patients were randomized in three equal number groups to receive either sufentanil 5 ug (group S), morphine 3 mg (group M) or normal saline 20 cc as placebo, (group P) intra-articularly at the end of arthroscopic knee surgery, under general anesthesia. Pain levels at rest and at intervals of 1,2,4,8,12,14 hours and during movement of knee at discharge time, were measured by a Visual Analog Scale (VAS) and Numeric Rating Scale (NRS) postoperatively and the day after surgery. RESULTS: Significant difference between the groups (p < 0.001) were found. Pain scores were statistically lower in the sufentanil (S) and morphine (M) groups in comparison with the normal saline placebo (P) group. Inta articular sufentanil was however more effective than morphine. CONCLUSION: The Intra-articular injection of morphine and sufentanil, reduce both the post arthroscopic knee procedures pain, and the need for supplementary analgesics. However, sufentanil, 5 microg is more effective than morphine. PMID- 15651518 TI - Assessment of post-operative pain management at the American University of Beirut Medical Center. PMID- 15651519 TI - The influence of surgical site on early postoperative hypoxemia. PMID- 15651520 TI - Osler-Weber-Rendu disease--unexpected complication following excision of splenic aneurysm--a case report. AB - Osler-Weber-Rendu disease is an autosomal dominant disease, sometimes known as hereditary hemorrhagic telangiectasia (HHT) with a family history. It is a rare disease and there are no records of families with inheritance lineage in Riyadh. We experienced an anesthetic management of a 28-yr-old woman with Osler-Weber Rendu disease. She was diagnosed to have splenic aneurysm and was scheduled for excision of splenic aneurysm. A chest X-ray revealed pulmonary arterio-venous malformation all over the lungs. The anesthesia and surgery went deliberately smooth with selective period of Intensive Care admission. In the intermediate post operative period, however, she was diagnosed as having pleural effusion. A chest drain was inserted as an emergency, but the patient developed tension pneumothorax, which necessitated resuscitation and readmission to Intensive Care Unit. Lung atelactasis was diagnosed. Attendance by qualified surgical night staff, communication, vigilance and consultations are as important as the proper management of such rare cases. PMID- 15651521 TI - Traumatic aortic pseudoaneurysm. AB - A case report of a 5 years old male child, with a history of crash injury--(blunt trauma)--on the chest. Computed tomography (CT) scan, magnetic resonance imaging (MRI) of the chest revealed an aneurysm involving the arch of the aorta. An aortogram showed its exact extension. Chest X-Ray showed collapsed left lung (due to pressure of aneurysm on left main bronchus). Surgery was done o aortic arch: Resection of the aneurysm, and patch repair of aorta, under profound hypothermic circulatory arrest (PHCA) and CPB. Left bronchial tear was also closed. Post operatively the patient was ventilated for about 36 hours to allow for lung expansion. The patient had a smooth postoperative course. Intensive chest physiotherapy and repeated bronchoscopies helped in recovering the left lung function. PMID- 15651522 TI - Biology, ecology and control of the Penthaleus species complex (Acari: Penthaleidae). AB - Blue oat mites, Penthaleus spp. (Acari: Penthaleidae), are major agricultural pests in southern Australia and other parts of the world, attacking various pasture, vegetable and crop plants. Management of these mites has been complicated by the recent discovery of three cryptic pest species of Penthaleus, whereas prior research had assumed a single species. The taxonomy, population genetics, ecology, biology and control of the Penthaleus spp. complex are reviewed. Adult Penthaleus have a dark blue-black body approximately 1 mm in length, and eight red-orange legs. Within Australia, they are winter pests completing two or three generations a season, depending on conditions. The summer is passed as diapausing eggs, when long-distance dispersal is thought to occur. The Penthaleus spp. reproduce by thelytokous parthenogenesis, with populations comprising clones that differ ecologically. The three pest Penthaleus spp. differ markedly in their distributions, plant hosts, timing of diapause egg production and response to pesticides, highlighting the need to develop control strategies that consider each species separately. Chemicals are the main weapons used in current control programs, however research continues into alternative more sustainable management options. Host plant resistance, crop rotations, conservation of natural enemies, and improved timing of pesticide application would improve the management of these pests. The most cost-effective and environmentally acceptable means of control will result from the integration of these practices combined with the development of a simple field-based kit to distinguish the different mite species. PMID- 15651523 TI - Combining plant- and soil-dwelling predatory mites to optimise biological control of thrips. AB - The efficiency of a natural enemy combination compared to a single species release for the control of western flower thrips (WFT) Frankliniella occidentalis (Pergande) on cucumber plants was investigated. Since a large part of F occidentalis seems to enter the soil passage, a joint release of the plant inhabiting predatory mite Amblyseius cucumeris (Oudemans) that feeds on thrips first-instar larvae and the soil-dwelling predatory mite Hypoaspis aculeifer (Canestrini) that preys on thrips pupae in the ground might offer a promising approach for a holistic control strategy. Therefore, two sets of experiments were conducted in cooperation with a commercial vegetable grower where the plants in plots were infested with a defined number of larval and adult F occidentalis. Two species of natural enemies were released either synchronously or solely, and their efficacy was compared to control plots devoid of antagonists. In both experiments, the predatory mites were released twice with a density of 46 A. cucumeris/m2, and 207 H. aculeifer/m2 (low-density) in the first experiment and 528 H. aculeifer/m2 (high-density) in the second one. Population growth of all arthropod species on the plants and in the soil was quantified at regular intervals and included all soil-dwelling mites and alternative preys present in the substrate. The results showed that H. aculeifer alone had a significant impact on thrips population development only when released at high-densities, but competence was lower compared to the other antagonist treatments. The impact of A. cucumeris alone and A. cucumeris & H. aculeifer combined was similar. Thus, the pooled exploitation of natural enemies did not boost thrips control compared to the single species application of A. cucumeris (non-additive effect), which could be explained by resource competition between both predatory mite species. Species number and population size in the soil of the experimental plots both showed a high variability, a possible consequence of their interaction with released soil-dwelling predatory H. aculeifer mites. The impact of resource competition and presence of alternative preys on thrips biological control is exhaustively discussed. From our study, we can extract the subsequent conclusions: (1) the combined use of H. aculeifer and A. cucumeris cannot increase thrips control on cucumber compared to the release of A. cucumeris alone, but the overall reliability of thrips biological control might be enhanced, (2) the availability of alternative preys seemed to affect the thrips predation rate of H. aculeifer, and (3) the impact of naturally occurring soil predatory mites on the control of WFT seemed to be partial. PMID- 15651524 TI - Prey stage preference and functional response of Euseius hibisci to Tetranychus urticae (Acari: Phytoseiidae, Tetranychidae). AB - The aims of this study were: (a) determine the prey stage preference of female Euseius hibisci (Chant) (Phytoseiidae) at constant densities of different stages of Tetranychus urticae Koch (Tetranychidae), (b) assess the functional response of the predator females to the varying densities of eggs, larvae, or protonymphs of T. urticae, and (c) estimate the functional response of E. hibisci when pollen of Ligustrum ovalifolium was present as well. We conducted experiments on excised pieces of strawberry leaf arenas (Fragaria ananassa) under laboratory conditions of 25+/-2 degrees C, 60+/-5% RH and 12 h photophase. Our results indicated that the predator consumed significantly more prey eggs than other prey stages. Consumption of prey deutonymphs and adults was so low that they were excluded from the non-choice functional response experiments. The functional response on all food items was of type II. The two parameters of the functional response were estimated for each prey type by means of the adjusted non-linear regression model. The highest estimated value a' (instantaneous rate of discovery) and the lowest value of Th (handling time, including digestion) were found for the predator feeding on prey eggs, and a' was lowest and Th highest when fed protonymphs. Using the jack-knife method, the values for the functional response parameters were estimated. The values of a' and Th produced by the model were similar among all prey types except for the eggs, which were different. Using pollen simultaneously with prey larvae decreased the consumption of the latter over the full range of prey densities The suitability of this predator for biological control of T. urticae on strawberry is discussed. PMID- 15651525 TI - Mitochondrial DNA and RAPD polymorphisms in the haploid mite Brevipalpus phoenicis (Acari: Tenuipalpidae). AB - Brevipalpus phoenicis (Geijskes) (Acari: Tenuipalpidae) is recognized as the vector of citrus leprosis virus that is a significant problem in several South American countries. Citrus leprosis has been reported from Florida in the past but no longer occurs on citrus in North America. The disease was recently reported in Central America, suggesting that B. phoenicis constitutes a potential threat to the citrus industries of North America and the Caribbean. Besides B. phoenicis, B. obovatus Donnadieu, and B. californicus (Banks) have been incriminated as vectors of citrus leprosis virus and each species has hundreds of host plants. In this study, Brevipalpus mite specimens were collected from different plants, especially citrus, in the States of Florida (USA) and Sao Paulo (Brazil), and reared on citrus fruit under standard laboratory conditions. Mites were taken from these colonies for DNA extraction and for morphological species identification. One hundred and two Random Amplified Polymorphic DNA (RAPD) markers were scored along with amplification and sequencing of a mitochondrial cytochrome oxidase subunit I gene fragment (374 bp). Variability among the colonies was detected with consistent congruence between both molecular data sets. The mites from the Florida and Brazilian colonies were morphologically identified as belonging to B. phoenicis, and comprise a monophyletic group. These colonies could be further diagnosed and subdivided geographically by mitochondrial DNA analysis. PMID- 15651526 TI - 'Salivary secretions' of eriophyoids (Acari: Eriophyoidea): first results of an experimental model. AB - This paper concerns an approach to direct collection of eriophyoid 'salivary secretions', and reports preliminary results on biological assays providing evidence for the presence of plant growth promoting substances in these secretions. Eleven species belonging to the Phytoptidae, Eriophyidae and Diptilomiopidae, characterized by different host-plant interactions, were studied by immersing mites into the following oils: condensate of cedar oil, oil for immersion lenses, two kinds of olive oil, alpha-terpineol, hystolemon, vaseline oil, and soybean oil. Some species secreted small droplets of lipophobic substances at the tip of their mouthparts when they were immersed in objective lens oil. Mite mortality and percentage of secreting specimens depended on the species and the medium used. Aceria caulobia (Nalepa) was selected as the candidate for subsequent study, because this species displayed a higher percentage of secreting mites than the other species and numerous specimens were easily collected by means of an airflow and filtering device. The induced secretions were studied from January to June of 2000, 2001 and 2002. Rapid salivary bioassays were performed during the period of maximum induced secretion in 2001. They were evaluated using a wheat-coleoptile and an excised-radish cotyledon growth test, respectively, for indole-3-acetic acid-like and cytokinin like activity. The bioassays suggested the presence of chemicals with plant growth regulatory effects. A brief account of eriophyoid mortality in the oils was also given. PMID- 15651527 TI - Predation risk affects diapause induction in the spider mite Tetranychus urticae. AB - Whenever diapause induction triggers movement into another microhabitat or the development of protective morphological structures, this may also alter predation risk. If the risk of being eaten is lower in the diapause phase, then there may be selection favouring diapause induction in response to predators or their cues. In this article, we studied the effect of the predatory mite Typhlodromus pyri on diapause induction in the spider mite Tetranychus urticae. We used a Greek strain because under long-night photoperiods and low temperature only part of the population enters diapause, thereby leaving room for the impact of another factor. In spider mite groups under predation, the percentage diapause induction increased whenever night-lengths were such that diapause was induced (13-16 h of night). Given this diapause induction in response to predation risk, the question arises whether entering diapause helps spider mites to escape from predation and contribute more offspring to the spring generation next year. PMID- 15651528 TI - Chlorfenapyr resistance in two-spotted spider mite (Acari: Tetranychidae) from Australian cotton. AB - The responses of Tetranychus urticae Koch from Australian cotton to chlorfenapyr has been monitored since the 1997--1998 growing season. Resistance was first detected in the 2001--2002 season and then increased quickly in both level and proportion of resistant strains detected. In response, the resistance management strategy for chlorfenapyr use in cotton was altered and now recommends a further restriction of use from two to one spray per season. There was no evidence of negative cross-resistance to the pyrethroid bifenthrin, but chlorfenapyr was associated with an undefined negative cross-resistance. PMID- 15651529 TI - The attachment and stylostome of Trombidium newelli (Acari: Trombidiidae), an ectoparasitic mite on adults of alfalfa weevil, Hypera postica (Coleoptera: Curculionidae). AB - Larvae of the mite Trombidium newelli Welbourn and Flessel are ectoparasitic on adult alfalfa weevil, Hypera postica (Gyllenhall), an insect pest of alfalfa. The mite larvae are found under the elytra, attached to the dorsal surface of weevils' abdomen. T. newelli larvae use their chelicerae to penetrate the pliable and weakly sclerotized areas of the host's integument, and to hold on firmly to the host. The attachment sites associated with larval mites of different degrees of engorgement were examined using both light and scanning electron microscopy. The ventral inside surface of parasitized host tergites revealed a characteristic injury as spots varying in color from white-yellow to golden-brown depending on the engorged size of the mite. In addition, spots associated with fully engorged mite larvae showed an aggressive dendritically radiating mass expanding outward from the site of cheliceral penetration. This mass, known as feeding tube or stylostome was branched, with each branch ending in a cluster of closed bulbs. Stylostomes appear to exist independently in the host's tissue. Stylostome mass corresponded with the engorgement state of the mite, increasing as the mite larva increased in size. The possible nature and mechanism of stylostome formation is discussed. PMID- 15651530 TI - Penetration of light into carpet, and responses of the European house-dust mite (Dermatophagoides pteronyssinus) to varying light intensity and diurnal cycle. AB - This study examined the question of what effect exposure to light might play in determining the vertical distribution of house-dust mites in carpet, and the degree to which light penetrates worn and unworn carpets of different pile conformation (loop- versus cut-pile), height and colour. The effect on population increase of a diurnal lighting cycle versus continual darkness was also investigated. It was found that the penetration of light into carpets was largely unaffected by pile colour or conformation. Pile height was an important factor, however, and for a given height within the pile, light intensity was higher in carpets subjected to a greater degree of wear. This corresponded to the reduction in effective pile height that occurs with carpet use. Whilst the differences observed were sometimes large (up to two-fold for a given height within the pile), Petri dish studies suggested no mite preference for habitation of areas of low light intensity compared to high intensity. Additionally, culturing mites under a diurnal light cycle was shown to be no more efficient than culturing in complete darkness. These results suggest that exposure to light is not an important determinant of house-dust mite behaviour, or their ability to colonise textile substrates. PMID- 15651531 TI - Response of soil mites to organic cultivation in an ultisol in southeast Brazil. AB - Soil-dwelling mites of four plots under organic management were investigated in April and December 1998 and in December 1999. Their populations were compared with mite populations in a pasture and forest in the vicinity. It was observed that there was always an initial reduction in the populations of soil mites and in the activity of the epigeic forms whenever a plot was opened up and disturbed mechanically in preparation for cultivation, irrespective of previous organic inputs. With time, the densities and activities of mites recovered under organic management. The uropodine and oribatid mites in particular benefited more from organic management than gamasine and actinedid mites. Uropodine mites increased tremendously under banana where there was fresh cow dung manure. Oribatid mite species Nothrus seropedicalensis and Archegozetes magnus were dominant in organic plots where the soil was moist and temperatures were lower than the ambient. Protoribates rioensis was dominant in organic plots where the soil was drier and temperatures were higher than the ambient. Galumna was the most active oribatid taxon on the floor of all plots, with the highest activity recorded under maracuja and in pasture plots. The results suggest that while densities and activities of soil mites increased in the organic plots, the community structure and recruitment period of oribatid mites were altered. Oribatid mite diversity was higher in the organic plots than in the pasture but lower than in the forest, where Belba sp. and many Eremobelboid brachypiline genera were present, but absent in the organic plots and pasture. PMID- 15651532 TI - The phenology, oviposition and feeding rate of Anystis baccarum, a predatory mite in Bramley apple orchards in Northern Ireland. PMID- 15651533 TI - Integrated control of Boophilus microplus ticks in Cuba based on vaccination with the anti-tick vaccine Gavac. AB - Boophilus microplus has developed resistance against a range of chemical acaricides which has stimulated the development of alternative methods such as vaccination against ticks. In Cuba, the Bm86-based recombinant vaccine Gavac has been successfully used in a number of controlled laboratory and field trials in cattle against B. microplus. In this paper, we have evaluated Gavac in a large scale field trial wherein 588,573 dairy cattle were vaccinated with the aim to reduce the number of acaricidal treatments. It was found that the number of acaricidal treatments could be reduced by 87% over a period of 8 years (1995- 2003). Prior to the introduction of the vaccine, 54 clinical cases of babesiosis and six fatal cases were reported per 1000 animals. Six years later, the incidence of babesiosis was reduced to 1.9 cases per 1000 cattle and mortality reduced to 0.18 per 1000. The national consumption of acaricides in Cuba could be reduced by 82% after the implementation of the integrated anti-B. microplus control program. PMID- 15651534 TI - Perceived cognitive function is a major determinant of health related quality of life in a non-selected population of patients with coronary artery disease--a principal components analysis. AB - OBJECTIVE: To assess health related quality of life (HRQL) and explore its underlying structure in a non-selected population of patients with coronary artery disease (CAD). DESIGN, SETTING AND SUBJECTS: HRQL was estimated by the disease specific Cardiac Health Profile (CHP) questionnaire and the EuroQol-VAS (EQ) in 253 consecutive unselected CAD patients in Sodertalje, Stockholm County, Sweden. Explorative factor analysis was used to identify independent dimensions of HRQL. Current angina was ranked according to Canadian Cardiovascular Society Classification (CCS). RESULTS: Four independent principal factors representing perceived cognitive, physical, social and emotional functions underlying the patients' HRQL were found. Identical factors were recognized with an alternate technique. The major factor--explaining 43 % of HRQL--was perceived cognitive function reflecting ability to concentrate, activity drive, memory and problem solving. Cognitive function correlated to EQ but not to CCS. Perceived physical function/general health explained 9% of HRQL and was as expected related both to EQ and CCS. Total CHP scores differed significantly to those of healthy controls. CONCLUSIONS: Perceived cognitive function seems to be a major determinant of HRQL in CAD patients. This, in addition to earlier reports of possible prognostic information of reduced cognitive function, would prompt us to propose that HRQL assessments should include questions aimed to assess cognitive function. PMID- 15651535 TI - Quality of life in tuberculosis: a review of the English language literature. AB - INTRODUCTION: Tuberculosis (TB) studies have concentrated on clinical outcomes; few studies have examined the impact of TB on patients' quality of life (QOL). METHODS: A systematic review of published medical literature using specific MESH terms: [Tuberculosis] and 1-[Outcome], 2-[Outcome Assessment], 3-[Quality of Life], 4-[Mood Disorder], 5-[Cost and Cost Analysis], 6-[Religion], 7 [Perception], 8-[Social Support], 9-[Optimism], 10-[Stress], 11-[Signs and Symptoms], and 12-[Cost of Illness]. This yielded 1972 articles; 60 articles met inclusion criteria and were reviewed. RESULTS: TB somatic symptoms have been well studied, but there were no studies of effects on physical functioning or general health perceptions. Patients tend to be worried, frustrated, or disappointed by their diagnosis, but it is unknown how emotional health changes with treatment. Diagnosed patients are less likely to find work, and less able to work and care for their families. TB creates the greatest financial burden on the poor. In developing, countries, patients and their families are ostracized by society, and families sometimes ostracize patients; the extent of TB's social stigma in the developed countries is unknown. CONCLUSION: There has been relatively little research on TB QOL and even less in developed countries. A better understanding may help improve treatment regimens, adherence to treatment, and functioning and well-being of people with TB. PMID- 15651537 TI - Comparing short form 6D, standard gamble, and Health Utilities Index Mark 2 and Mark 3 utility scores: results from total hip arthroplasty patients. AB - OBJECTIVES: The objectives are to compare SF-6D, standard gamble (SG), and Health Utilities Index (HUI) utility scores, compare change scores, and compare responsiveness. METHODS: A cohort of osteoarthritis patients referred for total hip arthroplasty (THA) were evaluated at the time of referral and followed until 3 months after THA. Patients were assessed using the SF-36, HUI2, HUI3, and the SG. Agreement is assessed using the intra-class correlation (ICC). Responsiveness is assessed using effect size, standardized response mean, and paired t-test. RESULTS: Data was available for 86 patients at baseline and for 63 at both pre- and post-surgery. At baseline mean SF-6D (0.61), SG (0.62), and HUI2 (0.62) scores were similar; the mean HUI3 score (0.52) was lower. Standard deviations were 0.10, 0.32, 0.19, and 0.22. At baseline, agreement between SF-6D and SG scores was 0.13, agreement between SF-6D and HUI2 was 0.47, and agreement between SF-6D and HUI3 was 0.28. Agreement at pre- and post-surgery was similar. The change in scores between post- and pre-surgery was 0.10 for SF-6D, 0.16 for SG, 0.22 for HUI2, and 0.23 for HUI3. Effect sizes were 1.10 for HUI2, 1.08 for HUI3, 1.06 for SF-6D, and 0.48 for the SG. CONCLUSIONS: Agreement between SG scores and SF-6D and HUI scores was low. The estimate of change in utility associated with THA was lowest for SF-6D. Additional longitudinal studies to compare utility measures appear to be warranted. PMID- 15651536 TI - Questionnaires to measure sexual quality of life. AB - CONTEXT: Sex is important to quality of life. There are a number of questionnaires to measure sexual-function, but many lack applicability and usefulness to certain groups. OBJECTIVE: To identify questionnaires measuring sexual function, determine the domains most commonly assessed, and examine evidence for their usefulness in different populations. DATA SOURCES: Computerized literature search using Medline, PubMed and PsychLit, reference lists, and unpublished reports, published in English between 1957 and 2001. MESH terms included sexual function, sexual dysfunction, sexual satisfaction, quality of life, and questionnaire. Articles were excluded if the questionnaire did not measure sexual function from the patient perspective. DATA EXTRACTION: Questionnaires were grouped as general questionnaires that include a sexual function domain, and sexual-function-specific questionnaires. Questionnaires were evaluated for domains, applicability to different populations, and evidence for reliability, validity and responsiveness. DATA SYNTHESIS: Literature search yielded 62 questionnaires, 57 which assessed sexual function from the patient perspective; 12 were general and 45 specific. Six domains were commonly represented, including interest and desire, satisfaction/quality of experience, excitement/arousal, performance, attitude/behavior, and relationship. Only 28% could be used in homosexual patients, and 52% were applicable to both genders; 57% were designed for use in chronic disease populations. Only nine questionnaires had evidence for both adequate reliability and validity. CONCLUSIONS: Current measures of sexual functioning often exclude important domains, lack applicability to gender and sexual preference groups, or lack adequate testing of validity and testing in important populations. Future questionnaires should take into account these concerns. PMID- 15651538 TI - Comparison of a single global item and an index of a multi-item health status measure among persons with and without diabetes in the US. AB - This study examined the hypothesis that a single global item can be substituted for an index of a multi-item assessment and lead to equivalent interpretative outcomes. Substitutability would be demonstrated if: (1) the two measures were strongly correlated, and regression analysis showed that the same variables accounted for variation in each measure, and (2) difference scores between multi item and global scores were close to zero and remained so as socio-demographic and co-morbid conditions varied. A multi-item assessment was constructed by mapping items from the NHANES I Epidemiologic Follow-up Study (NHEFS), using available data for persons with and without diabetes, onto the health-status classification system of the Health Utilities Index Mark 1 (HUI), creating the NHEFS-HUI. NHEFS-HUI data, when correlated to the self-assessed health status (SAHS) item, revealed a coefficient of 0.55. Regression analyses identified 9 of 14 variables contributed to the variability of each health status index, but differences existed in which variables were significant for which measure. Five of the possible 14 difference scores for persons with diabetes and non-diabetics approached zero. Persons with diabetes had lower NHEFS-HUI scores than non diabetics. These data were considered insufficient for demonstrating substitutability. Suggestions were made on how optimal substitutability could be achieved. PMID- 15651539 TI - Use of item response theory to develop a shortened version of the EORTC QLQ-C30 emotional functioning scale. AB - BACKGROUND: As part of a larger study whose objective is to develop an abbreviated version of the EORTC QLQ-C30 suitable for research in palliative care, analyses were conducted to determine the feasibility of generating a shorter version of the 4-item emotional functioning (EF) scale that could be scored in the original metric. METHODS: We used data from 24 European cancer studies conducted in 10 different languages (n = 8242). Item selection was based on analyses by item response theory (IRT). Based on the IRT results, a simple scoring algorithm was developed to predict the original 4-item EF sum scale score from a reduced number of items. RESULTS: Both a 3-item and a 2-item version (item 21 'Did you feel tense?' and item 24 'Did you feel depressed?') predicted the total score with excellent agreement and very little bias. In group comparisons, the 2-item scale led to the same conclusions as those based on the original 4 item scale with little or no loss of measurement efficiency. CONCLUSION: Although these results are promising, confirmatory studies are needed based on independent samples. If such additional studies yield comparable results, incorporation of the 2-item EF scale in an abbreviated version of the QLQ-C30 for use in palliative care research settings would be justified. The analyses reported here demonstrate the usefulness of the IRT-based methodology for shortening questionnaire scales. PMID- 15651540 TI - Use of tranquilizers and sleeping pills among cancer patients is associated with a poorer quality of life. AB - PURPOSE: To evaluate the association between sleeping pill/tranquilizer (SP/T) use and quality of life (QOL) among cancer patients. PATIENTS AND METHODS: Oncology patients (n = 909) in three Israeli hospitals were interviewed in clinics, day centers and in-patient departments regarding SP/T use in the previous week. Crude and adjusted QOL scores, measured using the European Organisation for Research and Treatment of Cancer Quality of Life Questionnaire C30 (EORTC QLQ-C30), were compared in users vs. non-users. RESULTS: Sleeping pill/tranquilizer use was self-reported by 234 (25.7%) participants, but rarely documented in medical charts. Factors associated with SP/T use were female gender (adjusted Odds ratio, OR: 1.79; 95% Confidence interval, CI: 1.22-2.62, age (OR: 4.6; 95% CI: 1.66-12.53 for age 70+), place of birth (OR: 1.97; 95% CI: 1.19-3.26 for Eastern Europe compared with Israel), concomitant use of painkillers (OR: 2.88; 95% CI: 1.97-4.20) and presence of cardiovascular disease (OR: 2.41; 95% CI: 1.48-3.91). Controlling these factors as well as disease status, users had a poorer QOL on all functional scales (p < 0.001) as well as global QOL. Furthermore, users reported increased severity of symptoms, especially fatigue, insomnia, pain, dyspnea and constipation (p < 0.01), compared to non-users. CONCLUSIONS: Use of SP/T, reported by one fourth of cancer patients, was associated with substantially poorer QOL and increased severity of symptoms. Causal inference is not possible given the cross-sectional design. Periodic inquiry regarding use of these medications in the Oncology Clinic is recommended since it may identify patients with poor QOL and unmet needs. PMID- 15651541 TI - Inter-observer agreement of a comprehensive health status classification system for pre-school children among patients with Wilms' tumor or advanced neuroblastoma. AB - We assessed inter-observer agreement on a new comprehensive health status classification system for preschool children (CHSCS-PS). Prospective assessments of children aged 2-4.9 years at the time of diagnosis of neuroblastoma (stages 3 4, excluding 4S) or Wilms' tumor (stages II-V) were collected independently from a parent and nurse by self-report during therapy. Responses were used to determine functional status on 10 health domains, as well as an overall disability score. Inter-observer agreement was evaluated by a kappa statistic for agreement about levels within individual domains, and by an intraclass correlation coefficient (ICC) for agreement of overall disability scores. Twenty four parent/nurse pairs of assessments were collected. Agreement was almost perfect for mobility and self-care, substantial for emotion and pain, and slight for speech. There was high percent agreement for vision, hearing, dexterity, learning and remembering, and thinking and problem solving, but insufficient variability in responses to calculate a kappa statistic. The ICC for overall disability scores between observers was 0.86, indicating strong agreement. Given the need for, and paucity of, instruments for the measurement of health-related quality of life in very young children, these results strongly support further evaluation of the CHSCS-PS. PMID- 15651542 TI - Health-related quality of life in a rare disease: hereditary hemorrhagic telangiectasia (HHT) or Rendu-Osler-Weber disease. AB - The levels of the health-related quality of life (HR-QoL) were analyzed in hereditary hemorrhagic telangiectasia (HHT) patients. The Short Form-36 Health Survey (SF-36) was administered to 50 HHT patients and scores were compared to a cohort of 2301 normal subjects. Clinical variables were patient age, illness duration, number of epistaxis episodes in the previous year and hemoglobin levels. Physical functioning, physical role limitations, bodily pain, social functioning, emotional role limitations and the physical component scores were lower among females. In multivariable analyses increasing age was related to lower physical functioning (p < 0.04), physical role limitations (p < 0.008), bodily pain (p < 0.05) and emotional role limitations (p < 0.01), while higher hemoglobin levels improved physical functioning (p < 0.03). The number of epistaxis episodes was negatively associated with physical role limitations (p < 0.009), vitality (p < 0.002), social functioning (p < 0.001), physical component summary (p < 0.001) and bodily pain (p < 0.01). Illness duration was negatively related to the mental component summary (p < 0.004). HHT patients had a lower HR QoL with respect to normal controls in all domains except for bodily pain. Females had lower scores for several domains. Epistaxis was the most important clinical variable. PMID- 15651543 TI - Validation of the 'Vienna List' as a proxy measure of quality of life for geriatric rehabilitation patients. AB - The reason for the study was to investigate the discriminative, content- and criterion-related validity of the 'Vienna List', a newly developed proxy-rating measurement for quality of life in very old and severely demented persons. The total population of patients of a geriatric rehabilitation clinic of one year (from November 2001 to October 2002; n = 687) was evaluated at admission and discharge by means of this proxy rating method together with the mini mental state examination, the timed up and go test, the Tinetti measure, the Barthel Index, and the Katz activities of daily living list. Differences, mainly on factors communication, mobility, aggression, and negative affect, appeared between seven diagnostic groups as well between several care-related categories and destination after discharge. The five factors of the measurement explained a significant amount of variance with a high specificity and more than 50% of the Barthel Index scores. The 'Vienna List', originally developed for the assessment of quality of life in severely demented patients, proved to be a useful, differentiating, less time-consuming and practical tool for the documentation of the outcome of geriatric inpatient rehabilitation, as well. PMID- 15651544 TI - Development and psychometric evaluation of the patient assessment of upper gastrointestinal symptom severity index (PAGI-SYM) in patients with upper gastrointestinal disorders. AB - OBJECTIVE: Describe the development and evaluation of a new self-report instrument, the patient assessment of upper gastrointestinal disorders-symptom severity index (PAGI-SYM) in subjects with gastroesophageal reflux disease (GERD), dyspepsia, or gastroparesis. METHODS: Recruited subjects with GERD (n=810), dyspepsia (n = 767), or gastroparesis (n = 169) from the US, France, Germany, Italy, the Netherlands, and Poland. Subjects completed the PAGI-SYM, SF 36, a disease-specific HRQL measure (PAGI-QOL), and disability day questions. Two week reproducibility was evaluated in 277 stable subjects. We evaluated construct validity by correlating subscale scores with SF-36, PAGI-QOL, disability days, and global symptom severity scores. RESULTS: The final 20-item PAGI-SYM has six subscales: heartburn/regurgitation, fullness/early satiety, nausea/vomiting, bloating, upper abdominal pain, and lower abdominal pain. Internal consistency reliability was good (alpha = 0.79-0.91); test-retest reliability was acceptable (Intraclass correlation coefficients alpha=0.60-0.82). PAGI-SYM subscale scores correlated significantly with SF-36 scores (all p < 0.0001), PAGI-QOL scores (all p < 0.0001), disability days (p < 0.0001), and global symptom severity (p < 0.0001). Mean PAGI-SYM scores varied significantly in groups defined by disability days (all p < 0.0001), where greater symptom severity was associated with more disability days. CONCLUSIONS: Results suggest the PAGI-SYM, a brief symptom severity instrument, has good reliability and evidence supporting construct validity in subjects with GERD, dyspepsia, or gastroparesis. PMID- 15651545 TI - Cross-cultural development and validation of a patient self-administered questionnaire to assess quality of life in upper gastrointestinal disorders: the PAGI-QOL. AB - OBJECTIVE: Summarize the Patient Assessment of Upper GastroIntestinal Disorders Quality of Life (PAGI-QOL) development and provide results on its reliability and validity from the international psychometric validation in dyspepsia, GastroEsophageal Reflux Disease (GERD), and gastroparesis. METHODS: Subjects completed the pilot PAGI-QOL at baseline and 8 weeks; and a subsample also at 2 weeks. Other assessments were: Patient Assessment of Upper Gastrointestinal Disorders-Symptom Severity Index, SF-36, number of disability days. RESULTS: 1736 patients completed the PAGI-QOL at baseline. The questionnaire was reduced, producing a 30-item final version covering five domains: Daily Activities, Clothing, Diet and Food Habits, Relationship (REL), and Psychological Well-Being and Distress. Internal consistency was excellent (Cronbach's alpha range: 0.83 0.96). Test-retest reproducibility was good: intraclass correlations coefficients were over 0.70 except for the REL scale (0.61). Concurrent validity between the PAGI-QOL total score and all SF-36 subscale scores was good with moderate (0.52) to strong (0.72) correlations. PAGI-QOL scores showed excellent discriminant properties: patients who had spent some days in bed, had missed some days at work, and were kept from usual activities had much lower PAGI-QOL scores than those who did not (p < 0.0001). CONCLUSION: The PAGI-QOL is a valid and reliable instrument assessing quality of life in patients with dyspepsia, GERD, or gastroparesis. PMID- 15651546 TI - Entrapment of the small intestine in the epiploic foramen in horses: a retrospective analysis of 71 cases recorded between 1991 and 2001. AB - The clinical features of 71 cases in 70 horses in which part of the small intestine became entrapped in the epiploic foramen are described. The horses' sex, age and breed, the month during which they were affected, and whether they exhibited stereotypic behaviour were compared with the same variables in 1279 horses which suffered other types of surgical colic during the same period. Thoroughbred and thoroughbred cross horses were over-represented among the 70 affected horses. There was no age or sex predilection. Fifty-five (77.5 per cent) of the cases occurred between October and March and 15 (21.1 per cent) occurred in January. The cases were significantly more likely to have a history of crib biting/windsucking than the control group (odds ratio 7.9, 95 per cent confidence interval 4.1 to 15.3). The condition had recurred in two of the horses. Fifty eight (81.7 per cent) recovered from surgery and 49 (69 per cent) survived until they were discharged from hospital. The median survival time of 31 of the affected horses discharged from the hospital was 700 days, whereas 417 horses with other types of surgical colic had a median survival time in excess of 1931 days. PMID- 15651547 TI - Comparison of xylazine, lidocaine and the two drugs combined for modified dorsolumbar epidural anaesthesia in cattle. AB - Twenty-four Holstein cattle scheduled for flank surgery in a standing position were randomly assigned to four groups of six. A 16 G, 120 mm Tuohy needle was inserted into the first interlumbar epidural space and its position was confirmed by the hanging drop technique. After air had been allowed to enter freely for approximately one minute, the epidural needle was slowly inserted 7 to 10 mm deeper to penetrate the epidural fat, and anaesthetic solution containing either 0.05 mg/kg bodyweight xylazine hydrochloride (xylazine), 0.025 mg/kg xylazine, 0.025 mg/kg xylazine and 0.1 mg/kg lidocaine hydrochloride (lidocaine), or 0.2 mg/kg lidocaine alone was administered. Signs of sedation were observed in the three groups treated with xylazine and the number of spinal segments involved in the area of analgesia when the anaesthetic contained xylazine was significantly greater than with 0.2 mg/kg lidocaine alone ( < 0.01). After the treatment with 0 025 mg/kg xylazine and 0.1 mg/kg lidocaine, flank surgery was performed successfully without additional line block or side effects. PMID- 15651548 TI - Properties of a calicivirus isolated from cats dying in an agitated state. AB - In June 1993, two of five pet cats kept in Yokohama city in Japan suddenly became agitated and died. Feline calicivirus (FCV) was isolated from them. One strain (FCV-S) was isolated from the spinal cord, lung and tonsil of cat 1, another (FCV B) from the ileum, medulla oblongata and cervical spinal cord of cat 2, and a third (FCV-SAKURA) from the oral cavity of one of the three surviving cats which showed no clinical signs. These three strains were equally resistant to pH 3.0 and serologically similar to each other, but distinct from strain F9. A genetic analysis, using a 208 base pair fragment from region E of the capsid, showed that FCV-Ari had a 70.4 per cent nucleotide and 77.3 per cent amino acid homology and FCV-F9 had a 68.6 per cent nucleotide and 73.9 per cent amino acid homology with the three strains, indicating that these two strains were genetically distinct from the three new isolates. Unvaccinated cats and cats which had been vaccinated against FCV-F9 developed watery diarrhoea but did not become agitated after the administration of FCV-S. The FCV-S strain did not induce signs of excitability after it was administered orally to specific pathogen-free cats. PMID- 15651549 TI - Neuropathological changes in ovine fetuse caused by tickborne fever. PMID- 15651550 TI - Attaching and effacing lesions in the intestines of an adult goat associated with natural infection with Escherichia coli O145. PMID- 15651551 TI - Intestinal obstruction due to sand in a dog. PMID- 15651552 TI - Regulation of paraprofessionals. PMID- 15651553 TI - Diaphragmatic hernia in a tup lamb. PMID- 15651554 TI - Effect of trace element supplementation on the fertility of dairy herds. PMID- 15651555 TI - International criminal prosecution of physicians: a critique of Professors Annas and Grodin's proposed International Medical Tribunal. PMID- 15651556 TI - Maternal brain death. PMID- 15651557 TI - Medical liability insurance and damage caps: getting beyond band aids to substantive systems treatment to improve quality and safety in healthcare. PMID- 15651558 TI - The obesity agency: centralizing the nation's fight against fat. PMID- 15651559 TI - Insulin kinetics in type-I diabetes: continuous and bolus delivery of rapid acting insulin. AB - We investigated insulin lispro kinetics with bolus and continuous subcutaneous insulin infusion (CSII) modes of insulin delivery. Seven subjects with type-1 diabetes treated by CSII with insulin lispro have been studied during prandial and postprandial conditions over 12 hours. Eleven alternative models of insulin kinetics have been proposed implementing a number of putative characteristics. We assessed 1) the effect of insulin delivery mode, i.e., bolus or basal, on the insulin absorption rate, the effects of 2) insulin association state and 3) insulin dose on the rate of insulin absorption, 4) the remote insulin effect on its volume of distribution, 5) the effect of insulin dose on insulin disappearance, 6) the presence of insulin degradation at the injection site, and finally 7) the existence of two pathways, fast and slow, of insulin absorption. An iterative two-stage parameter estimation technique was used. Models were validated through assessing physiological feasibility of parameter estimates, posterior identifiability, and distribution of residuals. Based on the principle of parsimony, best model to fit our data combined the slow and fast absorption channels and included local insulin degradation. The model estimated that 67(53 82)% [mean (interquartile range)] of delivered insulin passed through the slow absorption channel [absorption rate 0.011(0.004-0.029) min(-1)] with the remaining 33% passed through the fast channel [absorption rate 0.021(0.011-0.040) min(-1)]. Local degradation rate was described as a saturable process with Michaelis-Menten characteristics [VMAX = 1.93(0.62 - 6.03) mU min(-1), KM = 62.6(62.6 - 62.6) mU]. Models representing the dependence of insulin absorption rate on insulin disappearance and the remote insulin effect on its volume of distribution could not be validated suggesting that these effects are not present or cannot be detected during physiological conditions. PMID- 15651560 TI - Numerical simulations of light scattering by red blood cells. AB - Scattering of electromagnetic waves from a red blood cell is simulated using the finite-difference time-domain method (FDTD), the Rytov approximation and the discrete dipole approximation (DDA). Both FDTD and DDA are full wave methods that give accurate results in a wide range of wavelengths. The Rytov approximation is a much simpler method that is limited to scattering angles within 30 degrees from the forward direction. The investigation comprehends different wavelengths and different orientations of the cell. It shows that the shape, volume, and orientation of the cell have a large influence on the forward scattering. PMID- 15651561 TI - Identification of cardiac rhythm features by mathematical analysis of vector fields. AB - Automated techniques for locating cardiac arrhythmia features are limited, and cardiologists generally rely on isochronal maps to infer patterns in the cardiac activation sequence during an ablation procedure. Velocity vector mapping has been proposed as an alternative method to study cardiac activation in both clinical and research environments. In addition to the visual cues that vector maps can provide, vector fields can be analyzed using mathematical operators such as the divergence and curl. In the current study, conduction features were extracted from velocity vector fields computed from cardiac mapping data. The divergence was used to locate ectopic foci and wavefront collisions, and the curl to identify central obstacles in reentrant circuits. Both operators were applied to simulated rhythms created from a two-dimensional cellular automaton model, to measured data from an in situ experimental canine model, and to complex three dimensional human cardiac mapping data sets. Analysis of simulated vector fields indicated that the divergence is useful in identifying ectopic foci, with a relatively small number of vectors and with errors of up to 30 degrees in the angle measurements. The curl was useful for identifying central obstacles in reentrant circuits, and the number of velocity vectors needed increased as the rhythm became more complex. The divergence was able to accurately identify canine in situ pacing sites, areas of breakthrough activation, and wavefront collisions. In data from human arrhythmias, the divergence reliably estimated origins of electrical activity and wavefront collisions, but the curl was less reliable at locating central obstacles in reentrant circuits, possibly due to the retrospective nature of data collection. The results indicate that the curl and divergence operators applied to velocity vector maps have the potential to add valuable information in cardiac mapping and can be used to supplement human pattern recognition. PMID- 15651562 TI - Automatic detection of epileptiform events in EEG by a three-stage procedure based on artificial neural networks. AB - This paper introduces a three-stage procedure based on artificial neural networks for the automatic detection of epileptiform events (EVs) in a multichannel electroencephalogram (EEG) signal. In the first stage, two discrete perceptrons fed by six features are used to classify EEG peaks into three subgroups: 1) definite epileptiform transients (ETs); 2) definite non-ETs; and 3) possible ETs and possible non-ETs. The pre-classification done in the first stage not only reduces the computation time but also increases the overall detection performance of the procedure. In the second stage, the peaks falling into the third group are aimed to be separated from each other by a nonlinear artificial neural network that would function as a postclassifier whose input is a vector of 41 consecutive sample values obtained from each peak. Different networks, i.e., a backpropagation multilayer perceptron and two radial basis function networks trained by a hybrid method and a support vector method, respectively, are constructed as the postclassifier and then compared in terms of their classification performances. In the third stage, multichannel information is integrated into the system for contributing to the process of identifying an EV by the electroencephalographers (EEGers). After the integration of multichannel information, the overall performance of the system is determined with respect to EVs. Visual evaluation, by two EEGers, of 19 channel EEG records of 10 epileptic patients showed that the best performance is obtained with a radial basis support vector machine providing an average sensitivity of 89.1%, an average selectivity of 85.9%, and a false detection rate (per hour) of 7.5. PMID- 15651563 TI - The contribution of blood-flow-induced conductivity changes to measured impedance. AB - The paper considers the contribution of conductivity changes undergone in an anisotropical medium to measured resistance. This was achieved by extending the relationship proposed by Geselowitz to anisotropical materials described, therefore, by a conductivity tensor. It was found that each element of a conductivity change tensor contributed to the measured resistance only if a corresponding component of the electrical field was nonzero. Numerical calculations were performed for blood-flow-associated conductivity changes. A special experiment stand was developed which allowed experiments to be performed proving the theoretical results. It was found that the absolute value of resistance change measured in the direction perpendicular to the vessel axis was much smaller than that measured along the vessel axis. The results obtained may explain the fact that the actual change of measured resistance created by changes of conductivity induced by aortic blood flow is lower than expected from simplified models. PMID- 15651564 TI - Measuring curvature and velocity vector fields for waves of cardiac excitation in 2-D media. AB - Excitable media theory predicts the effect of electrical wavefront morphology on the dynamics of propagation in cardiac tissue. It specifies that a convex wavefront propagates slower and a concave wavefront propagates faster than a planar wavefront. Because of this, wavefront curvature is thought to be an important functional mechanism of cardiac arrhythmias. However, the curvature of wavefronts during an arrhythmia are generally unknown. We introduce a robust, automated method to measure the curvature vector field of discretely characterized, arbitrarily shaped, two-dimensional (2-D) wavefronts. The method relies on generating a smooth, continuous parameterization of the shape of a wave using cubic smoothing splines fitted to an isopotential at a specified level, which we choose to be -30 mV. Twice differentiating the parametric form provides local curvature vectors along the wavefront and waveback. Local conduction velocities are computed as the wave speed along lines normal to the parametric form. In this way, the curvature and velocity vector field for wavefronts and wavebacks can be measured. We applied the method to data sampled from a 2-D numerical model and several examples are provided to illustrate its usefulness for studying the dynamics of cardiac propagation in 2-D media. PMID- 15651565 TI - Encoding frequency modulation to improve cochlear implant performance in noise. AB - Different from traditional Fourier analysis, a signal can be decomposed into amplitude and frequency modulation components. The speech processing strategy in most modern cochlear implants only extracts and encodes amplitude modulation in a limited number of frequency bands. While amplitude modulation encoding has allowed cochlear implant users to achieve good speech recognition in quiet, their performance in noise is severely compromised. Here, we propose a novel speech processing strategy that encodes both amplitude and frequency modulations in order to improve cochlear implant performance in noise. By removing the center frequency from the subband signals and additionally limiting the frequency modulation's range and rate, the present strategy transforms the fast-varying temporal fine structure into a slowly varying frequency modulation signal. As a first step, we evaluated the potential contribution of additional frequency modulation to speech recognition in noise via acoustic simulations of the cochlear implant. We found that while amplitude modulation from a limited number of spectral bands is sufficient to support speech recognition in quiet, frequency modulation is needed to support speech recognition in noise. In particular, improvement by as much as 71 percentage points was observed for sentence recognition in the presence of a competing voice. The present result strongly suggests that frequency modulation be extracted and encoded to improve cochlear implant performance in realistic listening situations. We have proposed several implementation methods to stimulate further investigation. Index Terms-Amplitude modulation, cochlear implant, fine structure, frequency modulation, signal processing, speech recognition, temporal envelope. PMID- 15651566 TI - Spike detection using the continuous wavelet transform. AB - This paper combines wavelet transforms with basic detection theory to develop a new unsupervised method for robustly detecting and localizing spikes in noisy neural recordings. The method does not require the construction of templates, or the supervised setting of thresholds. We present extensive Monte Carlo simulations, based on actual extracellular recordings, to show that this technique surpasses other commonly used methods in a wide variety of recording conditions. We further demonstrate that falsely detected spikes corresponding to our method resemble actual spikes more than the false positives of other techniques such as amplitude thresholding. Moreover, the simplicity of the method allows for nearly real-time execution. PMID- 15651567 TI - Redundant system of passive markers for ultrasound scanhead tracking. AB - Scanhead tracking by opto-electronic (OE) systems allows high accuracy in three dimensional (3-D) freehand ultrasound imaging. In this paper, a new set of methods is proposed and compared with the standard approach [Gram-Schmidt method (GS)]. Three redundancy-based algorithms are introduced to compensate for possible loss of markers during data acquisition: regression plane (RP), multiple Gram-Schmidt (MGS), and center of mass least square (CMLS). When combined with the ultrasound instrument, the root-mean-squared (RMS) uncertainty in locating target points, over a working volume of 420 mm x 490 mm x 100 mm, improved by 7% and 24% using MGS and CMLS method respectively, compared to GS. A lower improvement was obtained with RP methods (5%), using the best marker configuration. In conclusion, CMLS method provides a robust and accurate procedure for 3-D freehand ultrasound scanhead tracking, able to manage possible loss of markers, with interesting perspectives for image fusion and body referenced 3-D ultrasound. PMID- 15651568 TI - A compact large voltage-compliance high output-impedance programmable current source for implantable microstimulators. AB - A new CMOS current source is described for biomedical implantable microstimulator applications, which utilizes MOS transistors in deep triode region as linearized voltage controlled resistors (VCR). The VCR current source achieves large voltage compliance, up to 97% of the supply voltage, while maintaining high output impedance in the 100 MOmega range to keep the stimulus current constant within 1% of the desired value irrespective of the site and tissue impedances. This approach improves stimulation efficiency, extends power supply lifetime, and saves chip area especially when the stimulation current level is high in the milliampere range. A prototype 4-channel microstimulator chip is fabricated in the AMI 1.5-microm, 2-metal, 2-poly, n-well standard CMOS process. With a 5-V supply, each stimulating site driver provides at least 425-V compliance and > 10 MOmega output impedance, while sinking up to 210 microA, and occupies 0.05 mm2 in chip area. A modular 32-site wireless neural stimulation microsystem, utilizing the VCR current source, is under development. PMID- 15651570 TI - A finite-element analysis of the effect of muscle insulation and shielding on the surface EMG signal. AB - We simulate the effect that insulating or shielding a muscle may have on electromyographic signal propagation using the finite element method. The results suggest that the crosstalk between insulated or shielded muscles is small but that it increases with increasing subcutaneous fat. The findings may be useful in the control of multifunctional prostheses. PMID- 15651569 TI - Mapping the human blood-retinal barrier function. AB - The aim of the work herein presented is to map blood-retinal barrier function by measuring retinal fluorescein leakage from the blood stream into the human vitreous using a confocal scanning laser ophthalmoscope (CSLO). Existing methods for the assessment of fluorescein leakage into the human vitreous are based on the qualitative evaluation of fluorescein angiographies (FA) and on volume measurements, as performed by the Fluorotron Master. A new procedure is presented capable of measuring fluorescein leakage into the vitreous while simultaneously imaging the retina. The present methodology computes the fluorescein leakage in a fully automated way, based on the three-dimensional fluorescence distribution in the human eye by using a single data acquisition. The processing includes signal filtering, volume alignment and profile deconvolution. The deconvolved profile obeys the established physical model. Representative cases shown are: a healthy eye; an eye with drusen from a nondiabetic person; a photocoagulated eye; and an eye with nonproliferative diabetic retinopathy. The results are in agreement with previous findings and go a step further by making possible its daily usage in a clinical setup based on currently available instrumentation. PMID- 15651571 TI - Continuous myoelectric control for powered prostheses using hidden Markov models. AB - This paper represents an ongoing investigation of dexterous and natural control of upper extremity prostheses using the myoelectric signal. The scheme described within uses a hidden Markov model (HMM) to process four channels of myoelectric signal, with the task of discriminating six classes of limb movement. The HMM based approach is shown to be capable of higher classification accuracy than previous methods based upon multilayer perceptrons. The method does not require segmentation of the myoelectric signal data, allowing a continuous stream of class decisions to be delivered to a prosthetic device. Due to the fact that the classifier learns the muscle activation patterns for each desired class for each individual, a natural control actuation results. The continuous decision stream allows complex sequences of manipulation involving multiple joints to be performed without interruption. The computational complexity of the HMM in its operational mode is low, making it suitable for a real-time implementation. The low computational overhead associated with training the HMM also enables the possibility of adaptive classifier training while in use. PMID- 15651572 TI - Hematocrit measurement by dielectric spectroscopy. AB - Based on permittivity changes, a new method to measure hematocrit (HCT) in extracorporeal blood systems is presented. Human blood samples were tested at different HCT levels pairing the values of permittivity change, obtained by means of a commercial impedance analyzer, with traditional centrifugation measurements. Data were correlated using both linear and nonlinear regression. When using the lineal model, the comparison yielded a high correlation coefficient (r = 0.99). Theoretical simplifications suggest that the method is independent of changes in the conductivities of the intracellular and extracellular compartments. The influence of osmolarity and conductivity of the extracellular compartment was analyzed. It is shown that HCT can be predicted within an error lower than 5% when those parameters changed as much as 1 mS/cm and 50 mOsm/kg, respectively. Thus, the method appears as valid and viable showing good possibilities in applications such as renal dialysis. PMID- 15651573 TI - Navigator echo motion artifact suppression in synthetic aperture ultrasound imaging. AB - We develop a simple yet effective technique for motion artifact suppression in ultrasound images reconstructed from multiple acquisitions. Assuming a rigid-body motion model, a navigator echo is computed for each acquisition and then registered to estimate the motion in between acquisitions. By detecting this motion, it is possible to compensate for it in the reconstruction step to obtain images that are free of lateral motion artifacts. The theory and practical implementation details are described and the performance is analyzed using computer simulations as well as real data. The results indicate the potential of the new method for real-time implementation in lower cost ultrasound imaging systems. PMID- 15651574 TI - Cole electrical impedance model--a critique and an alternative. AB - The Cole single-dispersion impedance model is based upon a constant phase element (CPE), a conductance parameter as a dependent parameter and a characteristic time constant as an independent parameter. Usually however, the time constant of tissue or cell suspensions is conductance dependent, and so the Cole model is incompatible with general relaxation theory and not a model of first choice. An alternative model with conductance as a free parameter influencing the characteristic time constant of the biomaterial has been analyzed. With this free conductance model it is possible to separately follow CPE and conductive processes, and the nominal time constant no longer corresponds to the apex of the circular arc in the complex plane. PMID- 15651576 TI - Interim 2003-based national population projections for the United Kingdom and constituent countries. AB - The 2003-based national population projections, carried out by the Government Actuary in consultation with the Registrars General, and using essentially the same underlying assumptions as for the previous 2002-based projections, show the population of the United Kingdom rising from 59.6 million in 2003, passing 60 million during 2005, to reach 65.7 million by 2031. Longer-term projections suggest the population will peak around 2050 at nearly 67 million and then very gradually start to fall. The population will become older with the median age expected to rise from 38.4 years in 2003 to 43.3 years by 2031. In 2003, there were around 700 thousand (six per cent) more children aged under 16, than people of state pensionable age. However, from 2007, the population of pensionable age is projected to exceed the number of children. PMID- 15651575 TI - Electrocardiographic motion artifact versus electrode impedance. AB - We degraded electrocardiographic electrodes by exposing them to air for four days and evaluated them on 12 subjects. After application, we recorded the electrocardiogram (including motion artifact), missed QRS detections and electrode impedance during 5 min of arm and body movements. Missed QRS detections increased with electrode impedance but correlation was poor. Increased electrode impedance was not a reliable predictor of a poor electrode and the need to replace it. PMID- 15651577 TI - European wide issues in population statistics. AB - This article discusses the issues facing those in Europe who measure population. It first sets out a high level description of the current demographic position in Europe, focussing on the main demographic changes that are underway and that will influence the population in years to come. Consideration is then given to the issues these raise for those who measure population and how population statistics systems will need to evolve in response. PMID- 15651578 TI - Demographic data needs for an ageing population. AB - A consequence of population ageing is an increased need for demographic information on the older population. This presents new challenges for data collection. This article analyses several key aspects of data collection on older people, including survey design and questions. It also critically examines the current range of UK and international sources for data on older people. Specifically, it addresses the under-representation of the following groups in some UK statistical sources: ) older people in institutions, 2) the oldest old and 3) older people from ethnic minorities. PMID- 15651579 TI - Advancements in the design of endodontic instruments for root canal preparation. PMID- 15651580 TI - Gutta-percha--the end of an era? PMID- 15651582 TI - Use of the operating microscope in endodontics. PMID- 15651581 TI - Clinical applications of mineral trioxide aggregate. PMID- 15651583 TI - Advanced technologies in endodontic practice: computer infrastructure and digital radiography. PMID- 15651584 TI - Advancements in electronic root length measuring. PMID- 15651585 TI - Incorporating laser technology into endodontic treatment. PMID- 15651586 TI - If you are not part of the solution, you are part of the problem. PMID- 15651587 TI - Patients' perceptions of nurse staffing, nursing care, adverse events, and overall satisfaction with the hospital experience. AB - Noticeably absent in the discussions surrounding nurse staffing and adverse events is the patients' perspective, except for their satisfaction with various aspects of their hospitalization experience. The results of a study undertaken to test a theoretical model of patients' perceptions of nurse staffing, the nursing care they receive, self-reported adverse events, and overall satisfaction with the hospital experience are presented. Perception of nurse staffing only weakly predicted the number of adverse events reported by patients, but was a strong predictor of the perception of nursing care received, and the perception of nursing care received was the only significant predictor of overall satisfaction with the hospital experience. PMID- 15651588 TI - One vision of academic nursing centers. AB - Reconciling vision, mission, and financial realities into a successful socially responsive endeavor is a challenge for academic nursing centers. A financially viable faculty practice enterprise is a response to this challenge. Entrepreneurial management and strategy assist in establishing financial sustainability. PMID- 15651589 TI - Social support in the workplace: nurse leader implications. AB - The state of the science as it relates to the stress-mediating role of social support in the health care work environment is explored. The ability to build a nursing leadership workforce is dependent upon a type of social support mechanism that mediates the inherent stress that comes along with managerial positions. Independent of the inevitable work-related stress associated with the nurse manager role, social support provided to current and aspiring nurse leaders is assumed to assist with coping and thriving in the work environment. PMID- 15651590 TI - A review of the effect of an accommodation program to support nurses with functional limitations. AB - The National Institutes of Health Clinical Center, Nursing and Patient Care Services, has developed a program to manage staff with functional restrictions using consistent processes and assignments that support the goals of the department while helping to contain costs. PMID- 15651591 TI - Access to quality health care: links between evidence, nursing language, and informatics. PMID- 15651592 TI - Building confident organizations by filling buckets, building infrastructures, and shining the flashlight. AB - Leaders have the ability to create confidence or fear in the organization. Kanter (2004) writes that confidence is the "...sweet spot between arrogance and despair" (p. 8). Overconfidence causes people to overshoot, and to assume they are invulnerable. Under-confidence is just as harmful because it leads to under investing in people, under-innovating, and eventually leads to the disenfranchisement of staff and poor morale. If we take Kanter's advice, we will build an infrastructure that creates confidence in everyone and the organization. But we will not stop there. We will focus "flashlights" on people and activities that are inspiring to others. PMID- 15651593 TI - The ultimate health care IT consumers: how nurses transform patient data into a powerful narrative of improved care. PMID- 15651594 TI - Publish or perish: the write thing for nursing. PMID- 15651595 TI - Creating safe spaces in organizations to talk about safety. PMID- 15651596 TI - Nurses and collaboration. PMID- 15651597 TI - Noncompete clauses in employment contracts violate a physicist's freedom to practice his/her profession. PMID- 15651598 TI - Noncompete clauses in employment contracts violate a physicist's freedom to practice his/her profession. PMID- 15651599 TI - Predicting energy response of radiographic film in a 6 MV x-ray beam using Monte Carlo calculated fluence spectra and absorbed dose. AB - The advantage of radiographic film is that it allows two-dimensional, high resolution dose measurement. While there is concern over its photon energy dependence, these problems are considered acceptable within small fields, where the scatter component is small. The application of film dosimetry to intensity modulated radiotherapy (IMRT) raises additional concern since the primary fluence may vary significantly within the field. The varying primary fluence in combination with a large scatter fraction, present for large fields and large depths, causes the spectrum at various points within the IMRT field to differ from the spectrum in the uniform fields typically used for calibrating the film. As a result, significant artifacts are introduced in the measured dose distribution. The purpose of this work is to quantify and develop a method to correct for these artifacts. Two approaches based on Monte Carlo (MC) simulations are examined. In the first method, the film artifact, as quantified by film and ion chamber output measurements in uniform square fields, is derived from the MC calculated ratio of absorbed doses to film and to water. In the second method, the measured film artifact is correlated with MC calculated photon spectra, revealing a strong correlation between the measured artifact and the "scatter"-to "primary" ratio, defined by the ratio of the number of photons below to the number of photons above 0.1 MeV, independent of field size and depth. These methods are evaluated in high- and low-dose regions of a large intensity modulated field created with a central block. The spectral approach is also tested with a clinical IMRT field. The absorbed dose method accurately corrects the measured film dose in the open part of the field and in points under the block and outside the field. The dose error is reduced from as much as 16% of the open field dose to less than 1%, as verified with an ion chamber. The spectral method accurately corrects the measured film dose in the open region of the centrally blocked field, but does not fully correct for the film artifact for points under the block and outside the field, where the spectrum is substantially different. Applied to the clinical field, the corrected film measurement shows good agreement with data obtained with a two-dimensional diode array. PMID- 15651600 TI - Four-dimensional (4D) PET/CT imaging of the thorax. AB - We have reported in our previous studies on the methodology, and feasibility of 4D-PET (Gated PET) acquisition, to reduce respiratory motion artifact in PET imaging of the thorax. In this study, we expand our investigation to address the problem of respiration motion in PET/CT imaging. The respiratory motion of four lung cancer patients were monitored by tracking external markers placed on the thorax. A 4D-CT acquisition was performed using a "step-and-shoot" technique, in which computed tomography (CT) projection data were acquired over a complete respiratory cycle at each couch position. The period of each CT acquisition segment was time stamped with an "x-ray ON" signal, which was recorded by the tracking system. 4D-CT data were then sorted into 10 groups, according to their corresponding phase of the breathing cycle. 4D-PET data were acquired in the gated mode, where each breathing cycle was divided into ten 0.5 s bins. For both CT and PET acquisitions, patients received audio prompting to regularize breathing. The 4D-CT and 4D-PET data were then correlated according to respiratory phase. The effect of 4D acquisition on improving the co-registration of PET and CT images, reducing motion smearing, and consequently increase the quantitation of the SUV, were investigated. Also, quantitation of the tumor motions in PET, and CT, were studied and compared. 4D-PET with matching phase 4D CTAC showed an improved accuracy in PET-CT image co-registration of up to 41%, compared to measurements from 4D-PET with clinical-CTAC. Gating PET data in correlation with respiratory motion reduced motion-induced smearing, thereby decreasing the observed tumor volume, by as much as 43%. 4D-PET lesions volumes showed a maximum deviation of 19% between clinical CT and phase- matched 4D-CT attenuation corrected PET images. In CT, 4D acquisition resulted in increasing the tumor volume in two patients by up to 79%, and decreasing it in the other two by up to 35%. Consequently, these corrections have yielded an increase in the measured SUV by up to 16% over the clinical measured SUV, and 36% over SUV's measured in 4D-PET with clinical-CT Attenuation Correction (CTAC) SUV's. Quantitation of the maximum tumor motion amplitude, using 4D-PET and 4D-CT, showed up to 30% discrepancy between the two modalities. We have shown that 4D PET/CT is clinically a feasible method, to correct for respiratory motion artifacts in PET/CT imaging of the thorax. 4D PET/CT acquisition can reduce smearing, improve the accuracy in PET-CT co-registration, and increase the measured SUV. This should result in an improved tumor assessment for patients with lung malignancies. PMID- 15651601 TI - Effect of MLC leaf width on the planning and delivery of SMLC IMRT using the CORVUS inverse treatment planning system. AB - This study investigates the influence of multileaf collimator (MLC) leaf width on intensity modulated radiation therapy (IMRT) plans delivered via the segmented multileaf collimator (SMLC) technique. IMRT plans were calculated using the Corvus treatment planning system for three brain, three prostate, and three pancreas cases using leaf widths of 0.5 and 1 cm. Resulting differences in plan quality and complexity are presented here. Plans calculated using a 1 cm leaf width were chosen over the 0.5 cm leaf width plans in seven out of nine cases based on clinical judgment. Conversely, optimization results revealed a superior objective function result for the 0.5 cm leaf width plans in seven out of the nine comparisons. The 1 cm leaf width objective function result was superior only for very large target volumes, indicating that expanding the solution space for plan optimization by using narrower leaves may result in a decreased probability of finding the global minimum. In the remaining cases, we can conclude that we are often not utilizing the objective function as proficiently as possible to meet our clinical goals. There was often no apparent clinically significant difference between the two plans, and in such cases the issue becomes one of plan complexity. A comparison of plan complexity revealed that the average 1 cm leaf width plan required roughly 60% fewer segments and over 40% fewer monitor units than required by 0.5 cm leaf width plans. This allows a significant decrease in whole body dose and total treatment time. For very complex IMRT plans, the treatment delivery time may affect the biologically effective dose. A clinically significant improvement in plan quality from using narrower leaves was evident only in cases with very small target volumes or those with concavities that are small with respect to the MLC leaf width. For the remaining cases investigated in this study, there was no clinical advantage to reducing the MLC leaf width from 1 to 0.5 cm. In such cases, there is no justification for the increased treatment time and whole body dose associated with the narrower MLC leaf width. PMID- 15651602 TI - Potential third-party radiation exposure from outpatients treated with 131I for hyperthyroidism. AB - Thirty-three hyperthyroid patients treated with radioiodine (mean administered activity 414 MBq, range 163-555) were studied to determine if pretreatment dosimetry could be used to give radiation protection advice that could assure compliance with the effective dose constraints suggested by the European Commission. Effective doses to travelers, co-workers, and sleeping partners were estimated by integrating the effective dose rate-versus-time curve obtained by fitting the dose rates measured several times after radioiodine administration to a biexponential function. The mean estimated effective doses to travelers, co workers, and sleeping partners were 0.11 mSv (0.05-0.24), 0.24 mSv (0.07-0.52), and 1.8 mSv (0.6-4.1), respectively. The best correlation was found between effective dose (D) in mSv and maximum activity (AUmax) in MBq taken up in the thyroid: Dtraveler=0.0005 * (AUmax) +0.04 (r=0.88,p< 0.01); Dco-worker=0.0013 * (A Umax) +0.03 (r=0.89,p < 0.01); Dsleeping partners=0.0105 * (AUmax)+0.16 (r=0.93,p < 0.01). Private/public transports are always allowed. For the co workers the effective dose constraint of 0.3 mSv is met without restrictions and with 3 days off work if AUmax is lower or higher than 185 MBq, respectively. For the sleeping partners the effective dose constraint of 3 mSv is met without restriction and with 4 nights separate sleeping arrangements if AUmax is lower or higher than 185 MBq, respectively. The potential for contamination by the patients was determined from perspiration samples taken from the patient's hands, forehead, and neck and in saliva at 4, 24, and 48 h after radioiodine treatment. The mean highest 131I activity levels for hands, forehead, neck, and saliva were 4.1 Bq/cm2, 1.9 Bq/cm2, 0.9 Bq/cm2, and 796 kBq/g, respectively. The results indicate that there is minimal risk of contamination from these patients. PMID- 15651603 TI - Influence of field size on a PTW type 23342 plane-parallel ionization chamber's response. AB - The response of a PTW type 23342 plane-parallel ionization chamber, both in air and in phantom, was evaluated for x-ray tube potentials between 30 and 100 kV and radiation field diameters ranging from 30 to 70 mm. The experiments were performed with a calibrated Pantak x-ray machine and made use of the same set of x-ray qualities adopted by the PTB primary laboratory for the calibration of such chambers. A Plexiglas phantom (1.18 g cm(-3)) 110 mm long, 110 mm wide, and 80 mm deep was used for phantom measurements. X-ray qualities were characterized by using 99.99% pure aluminum filters. On the basis of the IAEA's TRS 398, the article discusses the dependence of the plane-parallel ionization chamber readings with field size in air and in phantom, its implication with regard to clinical dosimetry, cross-calibration, and dissemination of calibration factors. PMID- 15651604 TI - An error-reduction-based algorithm for cone-beam computed tomography. AB - Image reconstruction from cone-beam projections collected along a single circular source trajectory is commonly done using the Feldkamp algorithm, which performs well only with a small cone angle. In this report, we propose an error-reduction based algorithm to increase the cone angle by several folds to achieve satisfactory image quality at the same radiation dose. In our scheme, we first reconstruct the object using the Feldkamp algorithm. Then, we synthesize cone beam projection data from the reconstructed volume in the same geometry, and reconstruct the volume again from the synthesized projections. Finally, these two reconstruction results are combined to reduce the reconstruction error and produce a superior image volume. The merit of this algorithm is demonstrated in numerical simulation. PMID- 15651606 TI - Adaptive algebraic reconstruction technique. AB - Algebraic reconstruction techniques (ART) are iterative procedures for reconstructing objects from their projections. It is proven that ART can be computationally efficient by carefully arranging the order in which the collected data are accessed during the reconstruction procedure and adaptively adjusting the relaxation parameters. In this paper, an adaptive algebraic reconstruction technique (AART), which adopts the same projection access scheme in multilevel scheme algebraic reconstruction technique (MLS-ART), is proposed. By introducing adaptive adjustment of the relaxation parameters during the reconstruction procedure, one-iteration AART can produce reconstructions with better quality, in comparison with one-iteration MLS-ART. Furthermore, AART outperforms MLS-ART with improved computational efficiency. PMID- 15651605 TI - Transmission and dose perturbations with high-Z materials in clinical electron beams. AB - High density and atomic number (Z) materials used in various prostheses, eye shielding, and beam modifiers produce dose enhancements on the backscatter side in electron beams and is well documented. However, on the transmission side the dose perturbation is given very little clinical importance, which is investigated in this study. A simple and accurate method for dose perturbation at metallic interfaces with soft tissues and transmission through these materials is required for all clinical electron beams. Measurements were taken with thin-window parallel plate ion chambers for various high-Z materials (Al, Ti, Cu, and Pb) on a Varian and a Siemens accelerator in the energy range of 5-20 MeV. The dose enhancement on both sides of the metallic sheet is due to increased electron fluence that is dependent on the beam energy and Z. On the transmission side, the magnitude of dose enhancement depends on the thickness of the high-Z material. With increasing thickness, dose perturbation reduces to the electron transmission. The thickness of material to reduce 100% (range of dose perturbation), 50% and 10% transmission is linear with the beam energy. The slope (mm/MeV) of the transmission curve varies exponentially with Z. A nonlinear regression expression (t=E[alpha+beta exp(-0.1Z)]) is derived to calculate the thickness at a given transmission, namely 100%, 50%, and 10% for electron energy, E, which is simple, accurate and well suited for a quick estimation in clinical use. Caution should be given to clinicians for the selection of thickness of high Z materials when used to shield critical structures as small thickness increases dose significantly at interfaces. PMID- 15651607 TI - Model-based microwave image reconstruction: simulations and experiments. AB - We describe an integrated microwave imaging system that can provide spatial maps of dielectric properties of heterogeneous media with tomographically collected data. The hardware system (800-1200 MHz) was built based on a lock-in amplifier with 16 fixed antennas. The reconstruction algorithm was implemented using a Newton iterative method with combined Marquardt-Tikhonov regularizations. System performance was evaluated using heterogeneous media mimicking human breast tissue. Finite element method coupled with the Bayliss and Turkel radiation boundary conditions were applied to compute the electric field distribution in the heterogeneous media of interest. The results show that inclusions embedded in a 76-diameter background medium can be quantitatively reconstructed from both simulated and experimental data. Quantitative analysis of the microwave images obtained suggests that an inclusion of 14 mm in diameter is the smallest object that can be fully characterized presently using experimental data, while objects as small as 10 mm in diameter can be quantitatively resolved with simulated data. PMID- 15651608 TI - Geometric misalignment and calibration in cone-beam tomography. AB - We present a new high-precision method for the geometric calibration in cone-beam computed tomography. It is based on a Fourier analysis of the projection-orbit data, recorded with a flat-panel area detector, of individual point-like objects. For circular scan trajectories the complete set of misalignment parameters which determine the deviation of the detector alignment from the ideal scan geometry are obtained from explicit analytic expressions. To derive these expressions we show how to disentangle the problems of calculating misalignment parameters and point coordinates. The calculation of the coordinates of the point objects inside the scanned volume, in units of the distance from the focal spot to the center of rotation, is then possible analytically likewise. We simulate point-projection data on a misaligned detector with various amounts of randomness added to mimic measurement uncertainties. This data is then employed in our calibration to validate the method by comparing the resulting misalignment parameters and point coordinates to the known true ones. We also present our implementation and results for the geometric calibration of micro-CT systems. The effectiveness of the corresponding misalignment correction in reducing image artifacts is exemplified by reconstructed micro-CT images. PMID- 15651609 TI - Modifications to the IMFAST leaf sequencing optimization algorithm. AB - The optimizing leaf sequencer IMFAST minimizes intensity modulated treatment times. However, algorithm modifications can yield improved results. Currently, during segment extraction, the largest extract for a given number of levels is chosen. The modification chooses an extract that yields the fewest segments and levels when the rod-pushing algorithm is applied to the difference between the original map and the extract. Also, successive optimization parameter values are now allowed to increase. These modifications reduced the number of segments and the relative fluence from the original algorithm by an average of 7%-11% and 8% 17%, respectively, depending on whether interdigitation and/or tongue-and-groove constraints were considered. The tests were done on two clinical head and neck intensity modulated radiation therapy cases. Compared to the sweeping window algorithm, a reduction of 35%-55% of the number of segments is possible with a change in the relative fluence of -9% - 16%, depending on the constraints. Compared to other previously published algorithms that deal with the constraints tested here, the modified IMFAST algorithm provides the greatest reduction in the number of segments with the minimum increase in the relative fluence. PMID- 15651610 TI - The use of modified single pencil beam dose kernels to improve IMRT dose calculation accuracy. AB - Intensity modulated radiation therapy (IMRT) is used to deliver highly conformal radiation doses to tumors while sparing nearby sensitive tissues. Discrepancies between calculated and measured dose distributions have been reported for regions of high dose gradients corresponding to complex radiation fluence patterns. For the single pencil beam convolution dose calculation algorithm, the ability to resolve areas of high dose structure is partly related to the shape of the pencil beam dose kernel (similar to how a photon detector's point spread function relates to imaging resolution). Improvements in dose calculation accuracy have been reported when the treatment planning system (TPS) is recommissioned using high-resolution measurement data as input. This study proposes to improve the dose calculation accuracy for IMRT planning by modifying clinical dose kernel shapes already present in the TPS, thus avoiding the need to reacquire higher resolution commissioning data. The in-house optimization program minimizes a cost function based on a two-dimensional composite dose subtraction/distance-to agreement (gamma) analysis. The final modified kernel shapes are reintroduced into the treatment planning system and improvements to the dose calcula tion accuracy for complex IMRT dose distributions evaluated. The central kernel value (radius =0 cm) has the largest effect on the dose calculation resolution and is the focus of this study. PMID- 15651611 TI - Beam collimation with polycapillary x-ray optics for high contrast high resolution monochromatic imaging. AB - Monochromatic imaging can provide better contrast and resolution than conventional broadband radiography. In broadband systems, low energy photons do not contribute to the image, but are merely absorbed, while high energy photons produce scattering that degrades the image. By tuning to the optimal energy, one can eliminate undesirable lower and higher energies. Monochromatization is achieved by diffraction from a single crystal. A crystal oriented to diffract at a particular energy, in this case the characteristic line energy, diffracts only those photons within a narrow range of angles. The resultant beam from a divergent source is nearly parallel, but not very intense. To increase the intensity, collimation was performed with polycapillary x-ray optics, which can collect radiation from a divergent source and redirect it into a quasi parallel beam. Contrast and resolution measurements were performed with diffracting crystals with both high and low angular acceptance. Testing was first done at 8 keV with an intense copper rotating anode x-ray source, then 17.5 keV measurements were made with a low power molybdenum source. At 8 keV, subject contrast was a factor of five higher than for the polychromatic case. At 17.5 keV, monochromatic contrast was two times greater than the conventional polychromatic contrast. The subject contrasts measured at both energies were in good agreement with theory. An additional factor of two increase in contrast, for a total gain of four, is expected at 17.5 keV from the removal of scatter. Scatter might be simply removed using an air gap, which does not degrade resolution with a parallel beam. PMID- 15651612 TI - Monte Carlo dosimetric study of best industries and Alpha Omega Ir-192 brachytherapy seeds. AB - Ir-192 seeds are widely used in the USA for low dose rate interstitial brachytherapy. There are two commercially available models: those manufactured by Best Industries filtered with stainless steel, and those manufactured by Alpha Omega seeds filtered with Pt. Newly developed 3D correction algorithms for brachytherapy are based on dosimetry data obtained on unbounded phantom size, allowing corrections for heterogeneities and actual tissue boundaries. Published dosimetric datasets for both seeds have been obtained under bounded conditions. The aim of the present study is to obtain dosimetric datasets for these seeds under full scatter conditions. The Monte Carlo GEANT4 code has been used to estimate air-kerma strength and dose rate in water around the Ir-192 seeds. Functions and parameters following the TG43 formalism are obtained and presented in tabular forms: the dose rate constant, the radial dose function, and the anisotropy function. Tables for the anisotropy factor have been obtained in order to apply punctual approximation. Differences between dose rate distributions for both seeds show that specific dataset must be used for each type of seed in clinical dosimetry. The data in the present study improve on published data in the following aspects: (i) dosimetric data were obtained under full scatter conditions, which affect dose values at distances greater than 4-5 cm from the source; (ii) the dose rate tables are given at greater distances from the source; and (iii) the spatial resolution in high dose gradient areas, such as those near the longitudinal source axis, has been improved. PMID- 15651613 TI - Endoprobe: a system for radionuclide-guided endoscopy. AB - Methods to guide the surgical treatment of cancer utilizing handheld beta sensitive probes in conjunction with tumor-avid radiopharmaceuticals [such as 18F fluorodeoxyglucose (FDG)] have previously been developed. These technologies could also potentially be used to assist in minimally invasive techniques for the diagnosis of cancer. The goal of this project is to develop and test a system for performing radionuclide-guided endoscopies. This system (called Endoprobe) has four major subsystems: beta detector, position tracker, endoscope, and user interface. The beta detection unit utilizes two miniaturized solid state detectors to preferentially detect beta particles. The position tracking system allows real-time monitoring of the unit's location. The beta detector and position tracking system's receiver are mounted on the tip of an endoscope. Information from the beta detector and tracking system, in addition to the video signal from the endoscope, are combined and presented to the user via a computer interface. The system was tested in a simulated search for radiotracer-avid areas of esophageal cancer. The search for esophageal cancer was chosen because this type of cancer is often diagnosed with endoscopic procedures and has been reported to have good affinity for FDG. Accumulations of FDG in the normal organs of the abdomen were simulated by an anthropomorphic torso phantom filled with the appropriate amounts of radioactivity. A 1.5- mm-thick gelatin film containing FDG was used to simulate radiotracer uptake in the lining of normal esophagus. Esophageal lesions (both benign and malignant) were simulated by thin disks of gelatin (diameters=3.5-12 mm) containing appropriate concentrations of FDG embedded in the gelatin film simulating normal esophagus. Endoprobe facilitated visual identification and examination of the simulated lesions. The position tracking system permitted the location of the Endoprobe tip to be monitored and plotted in real time on a previously acquired positron emission tomography computed tomography (PET-CT) image of the phantom. The detection system successfully acquired estimates of the beta flux emitted from areas chosen by the user. Indeed, Endoprobe was able to assist in distinguishing simulated FDG-avid areas as small as 3.5 mm in diameter from normal esophagus (p value <0.025). In addition to FDG, Endoprobe can be used with other positron or electron-emitting radionuclides such as IC or 131I. The next phase of this project will focus on modification of the prototype to make it more suitable for clinical use. PMID- 15651614 TI - Optimal field splitting for large intensity-modulated fields. AB - The multileaf travel range limitations on some linear accelerators require the splitting of a large intensity-modulated field into two or more adjacent abutting intensity-modulated subfields. The abutting subfields are then delivered as separate treatment fields. This workaround not only increases the treatment delivery time but it also increases the total monitor units (MU) delivered to the patient for a given prescribed dose. It is imperative that the cumulative intensity map of the subfields is exactly the same as the intensity map of the large field generated by the dose optimization algorithm, while satisfying hardware constraints of the delivery system. In this work, we describe field splitting algorithms that split a large intensity-modulated field into two or more intensity-modulated subfields with and without feathering, with optimal MU efficiency while satisfying the hardware constraints. Compared to a field splitting technique (without feathering) used in a commercial planning system, our field splitting algorithm (without feathering) shows a decrease in total MU of up to 26% on clinical cases and up to 63% on synthetic cases. PMID- 15651615 TI - Micro-CT with respiratory and cardiac gating. AB - Cardiopulmonary imaging in rodents using micro-computed tomography (CT) is a challenging task due to both cardiac and pulmonary motion and the limited fluence rate available from micro-focus x-ray tubes of most commercial systems. Successful imaging in the mouse requires recognition of both the spatial and temporal scales and their impact on the required fluence rate. Smaller voxels require an increase in the total number of photons (integrated fluence) used in the reconstructed image for constant signal-to-noise ratio. The faster heart rates require shorter exposures to minimize cardiac motion blur imposing even higher demands on the fluence rate. We describe a system with fixed tube/detector and with a rotating specimen. A large focal spot x-ray tube capable of producing high fluence rates with short exposure times was used. The geometry is optimized to match focal spot blur with detector pitch and the resolution limits imposed by the reproducibility of gating. Thus, it is possible to achieve isotropic spatial resolution of 100 microm with a fluence rate at the detector 250 times that of a conventional cone beam micro-CT system with rotating detector and microfocal x ray tube. Motion is minimized for any single projection with 10 ms exposures that are synchronized to both cardiac and breathing motion. System performance was validated in vivo by studies of the cardiopulmonary structures in C57BL/6 mice, demonstrating the value of motion integration with a bright x-ray source. PMID- 15651616 TI - Dosimetric study of the 15 mm ROPES eye plaque. AB - The main aim of this paper is to make a study of dose-rate distributions obtained around the 15 mm, radiation oncology physics and engineering services, Australia (ROPES) eye plaque loaded with 125I model 6711 radioactive seeds. In this study, we have carried out a comparison of the dose-rate distributions obtained by the algorithm used by the Plaque Simulator (PS) (BEBIG GmbH, Berlin, Germany) treatment planning system with those obtained by means of the Monte Carlo method for the ROPES eye plaque. A simple method to obtain the dose-rate distributions in a treatment planning system via the superposition of the dose-rate distributions of a seed placed in the eye plaque has been developed. The method uses eye plaque located in a simplified geometry of the head anatomy and distributions obtained by means of the Monte Carlo code GEANT4. The favorable results obtained in the development of this method suggest that it could be implemented on a treatment planning system to improve dose-rate calculations. We have also found that the dose-rate falls sharply along the eye and that outside the eye the dose-rate is very low. Furthermore, the lack of backscatter photons from the air located outside the eye-head phantom produces a dose reduction negligible for distances from the eye-plaque r<1 cm but reaches up to 20% near the air-eye interface. Results showed that the treatment planning system lacks accuracy around the border of the eye (in the sclera and the surrounding area) due to the simplicity of the algorithm used. The BEBIG treatment planning system uses a global attenuation factor that takes into account the effect of the eye plaque seed carrier and the lack of backscatter photons caused by the metallic cover, which in the case of a ROPES eye plaque has a default value of T= 1 (no correction). In the present study, a global attenuation factor T=0.96 and an air interface correction factor which improve on treatment planning system calculations were obtained. PMID- 15651617 TI - A simple method of independent treatment time verification in gamma knife radiosurgery using integral dose. AB - The purpose of this study is to develop a simple independent dose calculation method to verify treatment plans for Leksell Gamma Knife radiosurgery. Our approach uses the total integral dose within the skull as an end point for comparison. The total integral dose is computed using a spreadsheet and is compared to that obtained from Leksell GammaPlan. It is calculated as the sum of the integral doses of 201 beams, each passing through a cylindrical volume. The average length of the cylinders is estimated from the Skull-Scaler measurement data taken before treatment. Correction factors are applied to the length of the cylinder depending on the location of a shot in the skull. The radius of the cylinder corresponds to the collimator aperture of the helmet, with a correction factor for the beam penumbra and scattering. We have tested our simple spreadsheet program using treatment plans of 40 patients treated with Gamma Knife in our center. These patients differ in geometry, size, lesion locations, collimator helmet, and treatment complexities. Results show that differences between our calculations and treatment planning results are typically within +/ 3%, with a maximum difference of +/-3.8%. We demonstrate that our spreadsheet program is a convenient and effective independent method to verify treatment planning irradiation times prior to implementation of Gamma Knife radiosurgery. PMID- 15651618 TI - Automatic phase determination for retrospectively gated cardiac CT. AB - The recent improvements in CT detector and gantry technology in combination with new heart rate adaptive cone beam reconstruction algorithms enable the visualization of the heart in three dimensions at high spatial resolution. However, the finite temporal resolution still impedes the artifact-free reconstruction of the heart at any arbitrary phase of the cardiac cycle. Cardiac phases must be found during which the heart is quasistationary to obtain outmost image quality. It is challenging to find these phases due to intercycle and patient-to-patient variability. Electrocardiogram (ECG) information does not always represent the heart motion with an adequate accuracy. In this publication, a simple and efficient image-based technique is introduced which is able to deliver stable cardiac phases in an automatic and patient-specific way. From low resolution four-dimensional data sets, the most stable phases are derived by calculating the object similarity between subsequent phases in the cardiac cycle. Patient-specific information about the object motion can be determined and resolved spatially. This information is used to perform optimized high-resolution reconstructions at phases of little motion. Results based on a simulation study and three real patient data sets are presented. The projection data were generated using a 16-slice cone beam CT system in low-pitch helical mode with parallel ECG recording. PMID- 15651619 TI - Adaptive radiation therapy for compensation of errors in patient setup and treatment delivery. AB - In this paper, an adaptive radiation therapy algorithm is derived and evaluated using numerical simulations. Patient setup errors are considered and an off-line adaptive method to compensate for the effect of these is provided. The method consists of two parts, one for correction of patient position to account for the systematic error, and one for modulation of the fluence profiles to account for the random errors. The method is based on standard control theory for linear systems. It is investigated if this adaptive method can replace the use of a planning target volume (PTV) and therefore increase the possibilities to escalate the dose. Numerical simulations of treatments of a prostate patient indicate that this is the case. The simulations show that better organ-at-risk protection can be achieved when using the adaptation algorithm to correct for the geometrical uncertainties than when using a PTV. PMID- 15651620 TI - Dosimetric characteristics of the Leipzig surface applicators used in the high dose rate brachy radiotherapy. AB - The nucletron Leipzig applicator is designed for (HDR) 192Ir brachy radiotherapy of surface lesions. The dosimetric characteristics of this applicator were investigated using simulation method based on Monte Carlo N-particle (MCNP) code and phantom measurements. The simulation method was validated by comparing calculated dose rate distributions of nucletron microSelectron HDR 192Ir source against published data. Radiochromic films and metal-oxide-semiconductor field effect transistor (MOSFET) detectors were used for phantom measurements. The double exposure technique, correcting the nonuniform film sensitivity, was applied in the film dosimetry. The linear fit of multiple readings with different irradiation times performed for each MOSFET detector measurement was used to obtain the dose rate of each measurement and to correct the source transit-time error. The film and MOSFET measurements have uncertainties of 3%-7% and 3%-5%, respectively. The dose rate distributions of the Leipzig applicator with 30 mm opening calculated by the validated MC method were verified by measurements of film and MOSFET detectors. Calculated two-dimensional planar dose rate distributions show similar patterns as the film measurement. MC calculated dose rate at a reference point defined at depth 5 mm on the applicator's central axis is 7% lower than the film and 3% higher than the MOSFET measurements. The dose rate of a Leipzig applicator with 30 mm opening at reference point is 0.241+/-3% cGy h(-1) U(-1). The MC calculated depth dose rates and profiles were tabulated for clinic use. PMID- 15651621 TI - The impact of tumor motion upon CT image integrity and target delineation. AB - Accurate planning target volume delineation is vital to the success of conformal radiation techniques such as standard three-dimensional conformal radiotherapy and intensity modulated radiation therapy. With the exception of breath-hold schemes, all current approaches acquire images while the tumor is nonstationary and, as such, are subject to the presence of motion artifacts. In lung cancer sites where tumor mobility can be significant, the detrimental effect of these motion-induced distortions on image quality and subsequently target volume delineation cannot be ignored in the pursuit of improved treatment outcomes. To investigate the fundamental nature and functional dependence of computed tomography (CT) artifacts associated with lung tumor motion, and the implications for tumor delineation, a filtered backprojection algorithm was developed in MATLAB to generate transverse CT simulation images. In addition, a three dimensional phantom capable of mimicking the essential motions of lung tumors was constructed for experimental verification. Results show that the spatial extent of a mobile object is distorted from its true shape and location and does not accurately reflect the volume occupied during the extent of motion captured. The presence of motion also negatively impacts image intensity (density) integrity rendering accurate volume delineation highly problematic and calling into question the use of such data in CT-based heterogeneity correction algorithms for dosimetric calculation. PMID- 15651622 TI - Distortions induced by radioactive seeds into interstitial brachytherapy dose distributions. AB - In a previous article, we presented development and verification of an integral transport equation-based deterministic algorithm for computing three-dimensional brachytherapy dose distributions. Recently, we have included fluorescence radiation physics and parallel computation to the standing algorithms so that we can compute dose distributions for a large set of seeds without resorting to the superposition methods. The introduction of parallel computing capability provided a means to compute the dose distribution for multiple seeds in a simultaneous manner. This provided a way to study strong heterogeneity and shadow effects induced by the presence of multiple seeds in an interstitial brachytherapy implant. This article presents the algorithm for computing fluorescence radiation, algorithm for parallel computing, and display results for an 81-seed implant that has a perfect and imperfect lattice. The dosimetry data for a single model 6711 seeds is presented for verification and heterogeneity factor computations using simultaneous and superposition techniques are presented. PMID- 15651623 TI - Near-field dosimetry of 125I sources for interstitial brachytherapy implants measured using thermoluminescent sheets. AB - The dosimetric characteristics were measured for two types of 125I low-energy photon-emitting sources by using a wide and highly sensitive thermoluminescent (TL) sheet film, which was developed for two-dimensional dose distribution measurements. The TL film is made of Teflon homogeneously mixed with small powders of thermoluminescence (BaSO4:Eu doped). Various dosimetric parameters (i.e., radial dose function, 2D and 1D anisotropy functions) of model 6711 and 6702 125I sources were obtained at various distances from the source surfaces to 15 mm. These parameters obtained with TL sheet were compared with the data recommended in the updated AAPM TG-43 report. The radial dose functions measured with TL sheet are in agreement with those established data of model 6711 125I seed and model 6702 125I seed at most of the distances within 5% and 7%, respectively. All the measured anisotropy functions showed symmetry about the longitudinal source axis. The anisotropy of dose distributions was clearly present in the immediate vicinity of the source edges. The measured 2D anisotropy function values at 1 cm are in reasonably good agreement with the recommended values. The differences at two points in the 1D anisotropy functions measured with TL sheet and the established data at 1 cm from source center were 0.7% and 1.9% for model 6711 and 6702 125I sources, respectively; the differences at 0.5 cm were 1.5% and 1.7% for model 6711 and 6702 125I sources, respectively. The relative dosimetric characteristics in the vicinity of actual interstitial brachytherapy sources containing 125I have been experimentally determined by using the TL sheet as a 2D dosimeter. PMID- 15651624 TI - Automated matching of temporally sequential CT sections. AB - In the evaluation of patient response to therapy through measurements on thoracic computed tomography (CT) scans, the selection of anatomically equivalent sections in temporally sequential scans is required. We developed an automated method based on normalized mutual information (NMI) to expedite the selection of anatomically equivalent sections. The method requires as input two temporally sequential CT scans from the same patient. A specified section from the baseline scan is then compared with the sections of a follow-up scan. Each section in the follow-up scan is successively translated and rotated relative to the baseline section, and NMI is calculated. The section in the follow-up scan that yields the highest NMI with respect to the baseline section is selected as the matching section. The method was applied to a database of 22 pairs of temporally sequential CT scans from mesothelioma patients. Five observers manually selected their choice of the best anatomically matched section for each of three predetermined sections in the 22 baseline scans, and the range of selected sections was recorded. The automated method was applied to the same baseline sections to determine the computer-based anatomically matched sections in the corresponding follow-up scan. The automated process was performed using both original CT sections and sections automatically segmented so that only intrathoracic pixels contributed to NMI calculations. The accuracy of the automated method was quantified on a section-by-section basis by comparison with the range of sections selected by the observers. The automated method without segmentation selected equivalent sections within the observers' range for 54 of the 66 matching tasks (81.8%). An 11% improvement was achieved when thoracic segmentation was performed as a pre-processing step. PMID- 15651626 TI - Quasimonochromatic x-ray computed tomography by the balanced filter method using a conventional x-ray source. AB - A quasimonochromatic x-ray computed tomography (CT) system utilizing balanced filters has recently been developed for acquiring quantitative CT images. This system consisted of basic components such as a conventional x-ray generator for radiography, a stage for mounting and rotating objects, and an x-ray line sensor camera. Metallic sheets of Er and Yb were used as the balanced filters for obtaining quasimonochromatic incident x rays that include the characteristic lines of the W Kalpha doublet from a tungsten target. The mean energy and energy width of the quasimonochromatic x rays were determined to be 59.0 and 1.9 keV, respectively, from x-ray spectroscopic measurements using a high-purity Ge detector. The usefulness of the present x-ray CT system was demonstrated by obtaining spatial distributions of the linear attenuation coefficients of three selected samples--a 20 cm diameter cylindrical water phantom, a 3.5 cm diameter aluminum rod, and a human head phantom. The results clearly indicate that this apparatus is surprisingly effective for estimating the distribution of the linear attenuation coefficients without any correction of the beam-hardening effect. Thus, implementing the balanced filter method on an x-ray CT scanner has promise in producing highly quantitative CT images. PMID- 15651625 TI - CSnrc: correlated sampling Monte Carlo calculations using EGSnrc. AB - CSnrc, a new user-code for the EGSnrc Monte Carlo system is described. This user code improves the efficiency when calculating ratios of doses from similar geometries. It uses a correlated sampling variance reduction technique. CSnrc is developed from an existing EGSnrc user-code CAVRZnrc and improves upon the correlated sampling algorithm used in an earlier version of the code written for the EGS4 Monte Carlo system. Improvements over the EGS4 version of the algorithm avoid repetition of sections of particle tracks. The new code includes a rectangular phantom geometry not available in other EGSnrc cylindrical codes. Comparison to CAVRZnrc shows gains in efficiency of up to a factor of 64 for a variety of test geometries when computing the ratio of doses to the cavity for two geometries. CSnrc is well suited to in-phantom calculations and is used to calculate the central electrode correction factor Pcel in high-energy photon and electron beams. Current dosimetry protocols base the value of Pcel on earlier Monte Carlo calculations. The current CSnrc calculations achieve 0.02% statistical uncertainties on Pcel, much lower than those previously published. The current values of Pcel compare well with the values used in dosimetry protocols for photon beams. For electrons beams, CSnrc calculations are reported at the reference depth used in recent protocols and show up to a 0.2% correction for a graphite electrode, a correction currently ignored by dosimetry protocols. The calculations show that for a 1 mm diameter aluminum central electrode, the correction factor differs somewhat from the values used in both the IAEA TRS-398 code of practice and the AAPM's TG-51 protocol. PMID- 15651627 TI - Stereotactic arc therapy for small elongated tumors using cones and collimator jaws; dosimetric and planning aspects. AB - Stereotactic arc treatment of small intracranial tumors is usually performed with arcs collimated by circular cones, resulting in treatment volumes which are basically spherical. For nonspherical lesions this results in a suboptimal dose distribution. Multiple isocenters may improve the dose conformity for these lesions, at the cost of large overdosages in the target volume. To achieve improved dose conformity as well as dose homogeneity, the linac jaws (with a minimum distance of 1.0 cm to the central beam axis) can routinely be used to block part of the circular beams. The purpose of this study was to investigate the feasibility of blocking cones with diameters as small as 1.0 cm and a minimum distance between the jaw and the central beam axis of 0.3 cm. First, the reproducibility in jaw positioning and resulting dose delivery on the treatment unit were assessed. Second, the accuracy of the TPS dose calculation for these small fields was established. Finally, clinically applied treatment plans using nonblocked cones were compared with plans using the partially blocked cones for several treatment sites. The reproducibility in dose delivery on our Varian Clinac 2300 C/D machines on the central beam axis is 0.8% (1 SD). The accuracy of the treatment planning system dose calculation algorithm is critically dependent on the used fits for the penumbra and the phantom scatter. The average deviation of calculated from measured dose on the central beam axis is -1.0%+/-1.4% (1 SD), which is clinically acceptable. Partial cone blocking results in improved dose distributions for elongated tumors, such as vestibular schwannoma and uveal melanoma. Multiple isocenters may be avoided. The technique is easy to implement and requires no additional workload. PMID- 15651628 TI - Goniometric and conoscopic measurements of angular display contrast for one-, three-, five-, and nine-million-pixel medical liquid crystal displays. AB - Active-matrix liquid crystal displays (AMLCDs) have become the preferred choice for displaying digital diagnostic images because of their low cost of ownership, high contrast, and small footprint. However, the contrast and luminance of AMLCDs changes significantly with viewing direction. In this paper, we use a goniometric and a conoscopic method to measure angular contrast and luminance in one-, three , five-, and nine-million-pixel medical AMLCDs. The viewing angle characteristics of the displays are compared by evaluating the compliance with the desired grayscale and recommended tolerance limits. Using the measured contrast responses, we determined the angle along each of the orthogonal and diagonal axes for which the luminance ratio drops to 0.5, 0.2, and 0.1 of the maximum ratio, seen at perpendicular viewing. The results show a reduction as large as 54% in available JND levels between the perpendicular and off-normal viewing directions. All of the display systems proved to have better angular response in the horizontal and vertical direction as compared to the diagonal directions, and none were able to satisfy a 20% contrast tolerance limit for any viewing direction lying in a cone at an angle of 45 degrees from the normal. Additionally, we show that the measured contrast can be significantly affected by the measurement method used to record the angular change in luminance. PMID- 15651629 TI - Image reconstruction algorithm for a spinning strip CZT SPECT camera with a parallel slat collimator and small pixels. AB - This paper discusses the use of small pixels in a spinning CdZnTe single photon emission computed tomography (SPECT) camera that is mounted with a parallel slat collimator. In a conventional slat collimation configuration, there is a detector pixel between two adjacent collimator slats. In our design, the pixel size is halved. That is, there are two smaller pixels to replace a regular pixel between two adjacent slats while the collimator remains unchanged. It has an advantage over our older design that uses tilted slats. In order to acquire a complete data set the tilted-slat collimator must spin 360 degrees at each SPECT view while the proposed design requires only 180 degrees at each SPECT view. Computer simulations and phantom experiments have been carried out to investigate the performance of the small-pixel configuration. It is observed that this design has the potential to increase the spatial resolution of the detector while keeping photon counts the same. PMID- 15651630 TI - Segmentation of the prostate from suprapubic ultrasound images. AB - We present a technique for semiautomated segmentation of human prostates using suprapubic ultrasound (US) images. In this approach, a speckle reducing anisotropic diffusion (SRAD) is applied to enhance the images and the instantaneous coefficient of variation (ICOV) is utilized for edge detection. Segmentation is accomplished via a parametric active contour model in a polar coordinate system that is tailored to the application. The algorithm initially approximates the prostate boundary in two stages. First a primary contour is detected using an elliptical model, followed by a primary contour optimization using an area-weighted mean-difference binary flow geometric snake model. The algorithm was assessed by comparing the computer-derived contours with contours produced manually by three sonographers. The proposed method has application in radiation therapy planning and delivery, as well as in automated volume measurements for ultrasonic diagnosis. The average root mean square discrepancy between computed and manual outlines is less than the inter-observer variability. Furthermore, 76% of the computer-outlined contour is less than 1 sigma manual outline variance away from "true" boundary of prostate. We conclude that the methods developed herein possess acceptable agreement with manually contoured prostate boundaries and that they are potentially valuable tools for radiotherapy treatment planning and verification. PMID- 15651631 TI - Determination of the modulation transfer function using the edge method: influence of scattered radiation. AB - The edge method for measuring the modulation transfer function (MTF) has recently gained popularity due to its simplicity and appropriateness particularly for digital imaging systems. Often edge test devices made of rather thin metal sheets are used, which are semitransparent to x rays and may generate scattered radiation. The effect of this scattered radiation on the determined MTF was investigated both theoretically (assuming an ideal detector) and experimentally using a CsI-based digital detector. It was found that the MTF increases due to the scattered radiation for all spatial frequencies larger than 0 mm(-1). The theoretical model developed in this study predicts that the maximum error compared to the true detector MTF is given by S/A, where A is the attenuated fraction and S is the scattered fraction reaching the detector, relative to the incident radiation. Theoretical and experimental results are in good agreement for radiation qualities corresponding to general radiography (RQA3, RQA5, and RQA7), whereas for chest beam quality (RQA9) the experimentally observed MTF error is larger than predicted by the simple model, possibly because the energy response of the CsI-based detector differs from that of an ideal one. The theoretical MTF error reaches a value of 18% for a 0.25 mm thick lead edge of RQA9. Since the MTF enters squared into the determination of the detective quantum efficiency (DQE), an error of at least 36% in DQE may result when using this edge test device. In conclusion, the use of fully absorbing edge material is advised for MTF determination with the edge method. PMID- 15651632 TI - On the use of EPID-based implanted marker tracking for 4D radiotherapy. AB - Four-dimensional (4D) radiotherapy delivery to dynamically moving tumors requires a real-time signal of the tumor position as a function of time so that the radiation beam can continuously track the tumor during the respiration cycle. The aim of this study was to develop and evaluate an electronic portal imaging device (EPID)-based marker-tracking system that can be used for real-time tumor targeting, or 4D radiotherapy. Three gold cylinders, 3 mm in length and 1 mm in diameter, were implanted in a dynamic lung phantom. The phantom range of motion was 4 cm with a 3-s "breathing" period. EPID image acquisition parameters were modified, allowing image acquisition in 0.1 s. Images of the stationary and moving phantom were acquired. Software was developed to segment automatically the marker positions from the EPID images. Images acquired in 0.1 s displayed higher noise and a lower signal-noise ratio than those obtained using regular (> 1 s) acquisition settings. However, the markers were still clearly visible on the 0.1 s images. The motion of the phantom blurred the images of the markers and further reduced the signal-noise ratio, though they could still be successfully segmented from the images in 10-30 ms of computation time. The positions of gold markers placed in the lung phantom were detected successfully, even for phantom velocities substantially higher than those observed for typical lung tumors. This study shows that using EPID-based marker tracking for 4D radiotherapy is feasible, however, changes in linear accelerator technology and EPID-based image acquisition as well as patient studies are required before this method can be implemented clinically. PMID- 15651633 TI - Development of an integral system test for image-guided radiotherapy. AB - An integral system test was developed to determine the precision and accuracy of an image-guided radiotherapy system involving an x-ray volumetric imaging device mounted onto the gantry of a medical linear accelerator. The test was designed to interrogate the system components as a whole without deconstructing the individual sources of error. The integral system test was based on the imaging of an unambiguous stationary object in the treatment position and so took no account of patient related errors. An array of micromosfets interspersed within slices of a tissue equivalent phantom was developed as an imaging test object. It has previously been demonstrated that micromosfets have a very small active volume, are clearly visible on CT images, and produce no significant artifacts. In addition, the active volume of the micromosfets can be accurately inferred radiographically via the use of x-ray volumetric imaging. X-ray volumetric imaging was performed with the object in the treatment position, then reconstructed and transferred to a treatment planning system. With the phantom remaining undisturbed in the treatment position a series of treatment fields were designed to produce a series of fields with the leaf edge sweeping across active volume of the micromosfets. The fields were delivered with a micro-MLC to dosimetrically verify the position of the mosfets and compare with dose values produced by the treatment planning system. It was demonstrated that the systematic gantry flex could be accounted for by the imaging and delivery systems. For the delivery system small changes in leaf positions of the micro-MLC were required to account for gantry flex. The position of the micromosfets determined by the 50% dose position was on average (0.15+/-0.13) mm away from the position determined radiographically for the x and y axes, and (1.0+/-0.14) mm for the z axis. This implies that a margin of approximately 0.2 mm in the axial plane and 1.0 mm in the superior-inferior plane would be required at the delineation stage to ensure coverage of a tumor volume to account purely for imprecision in the image-guided radiotherapy system. The integral system test demonstrated that the image-guided radiotherapy system is capable, in the absence of patient motion, of imaging an object in the treatment position and delivering dose to that object with submillimeter accuracy. PMID- 15651634 TI - The influence of antiscatter grids on soft-tissue detectability in cone-beam computed tomography with flat-panel detectors. AB - The influence of antiscatter x-ray grids on image quality in cone-beam computed tomography (CT) is evaluated through broad experimental investigation for various anatomical sites (head and body), scatter conditions (scatter-to-primary ratio (SPR) ranging from approximately 10% to 150%), patient dose, and spatial resolution in three-dimensional reconstructions. Studies involved linear grids in combination with a flat-panel imager on a system for kilovoltage cone-beam CT imaging and guidance of radiation therapy. Grids were found to be effective in reducing x-ray scatter "cupping" artifacts, with heavier grids providing increased image uniformity. The system was highly robust against ring artifacts that might arise in CT reconstructions as a result of gridline shadows in the projection data. The influence of grids on soft-tissue detectability was evaluated quantitatively in terms of absolute contrast, voxel noise, and contrast to-noise ratio (CNR) in cone-beam CT reconstructions of 16 cm "head" and 32 cm "body" cylindrical phantoms. Imaging performance was investigated qualitatively in observer preference tests based on patient images (pelvis, abdomen, and head and-neck sites) acquired with and without antiscatter grids. The results suggest that although grids reduce scatter artifacts and improve subject contrast, there is little strong motivation for the use of grids in cone-beam CT in terms of CNR and overall image quality under most circumstances. The results highlight the tradeoffs in contrast and noise imparted by grids, showing improved image quality with grids only under specific conditions of high x-ray scatter (SPR> 100%), high imaging dose (Dcenter> 2 cGy), and low spatial resolution (voxel size > or = 1 mm). PMID- 15651635 TI - Automatic real-time surveillance of eye position and gating for stereotactic radiotherapy of uveal melanoma. AB - A new prototype (hardware and software) for monitoring eye movements using a noninvasive technique for gated linac-based stereotactic radiotherapy (SRT) of uveal melanoma was developed. The prototype was tested within the scope of a study for 11 patients. Eye immobilization was achieved by having the patient fixate a light source integrated into the system. The system is used in conjunction with a Head&Neck mask system for immobilization, and uses infrared tracking technology for positioning (both BrainLAB AG Heimstetten/Germany). It was used during CT and MR image acquisition as well as during all of five treatment fractions (6 MeV, 5 x 12 Gy to 80% isodose) to guarantee identical patient setup and eye rotational state during treatment planning and treatment delivery. Maximum temporal and angular deviations tolerated during treatment delivery can be chosen by the physician, the radiation then being interrupted automatically and instantaneously if those criteria are being exceeded during irradiation. A graphical user interface displays life video images of the treated eye and information about the current and previous rotational deviation of the eye from its reference treatment position. The physician thus has online access to data directly linked to the success of the treatment and possible side effects. Mean angular deviations during CT/MR scans and treatment deliveries ranged from 1.61 degrees to 3.64 degrees (standard deviations 0.87 degrees to 2.09 degrees ) which is in accordance with precision requirements for SRT. Typical situations when preset deviation criteria were exceeded are slow drifts (fatigue), sudden large eye movements (irritation), or if patients closed their eyes (fatigue). In these cases radiation was reliably interrupted by the gating system. In our clinical setup the novel system for computer-controlled eye movement gated treatments was well tolerated by all patients. The system yields quantitative real-time information about the eye's rotational state with respect to a reference position (treatment planning situation). Together with the possibility of performing movement-gated treatments of uveal melanoma, the system thus greatly improves the quality of this treatment. PMID- 15651636 TI - Fractional calculus in bioengineering, part 3. AB - Fractional calculus (integral and differential operations of noninteger order) is not often used to model biological systems. Although the basic mathematical ideas were developed long ago by the mathematicians Leibniz (1695), Liouville (1834), Riemann (1892), and others and brought to the attention of the engineering world by Oliver Heaviside in the 1890s, it was not until 1974 that the first book on the topic was published by Oldham and Spanier. Recent monographs and symposia proceedings have highlighted the application of fractional calculus in physics, continuum mechanics, signal processing, and electromagnetics, but with few examples of applications in bioengineering. This is surprising because the methods of fractional calculus, when defined as a Laplace or Fourier convolution product, are suitable for solving many problems in biomedical research. For example, early studies by Cole (1933) and Hodgkin (1946) of the electrical properties of nerve cell membranes and the propagation of electrical signals are well characterized by differential equations of fractional order. The solution involves a generalization of the exponential function to the Mittag-Leffler function, which provides a better fit to the observed cell membrane data. A parallel application of fractional derivatives to viscoelastic materials establishes, in a natural way, hereditary integrals and the power law (Nutting/Scott Blair) stress-strain relationship for modeling biomaterials. In this review, I will introduce the idea of fractional operations by following the original approach of Heaviside, demonstrate the basic operations of fractional calculus on well-behaved functions (step, ramp, pulse, sinusoid) of engineering interest, and give specific examples from electrochemistry, physics, bioengineering, and biophysics. The fractional derivative accurately describes natural phenomena that occur in such common engineering problems as heat transfer, electrode/electrolyte behavior, and sub-threshold nerve propagation. By expanding the range of mathematical operations to include fractional calculus, we can develop new and potentially useful functional relationships for modeling complex biological systems in a direct and rigorous manner. In Part 2 of this review (Crit Rev Biomed Eng 2004; 32(1):105-193), fractional calculus was applied to problems in nerve stimulation, dielectric relaxation, and viscoelastic materials by extending the governing differential equations to include fractional order terms. In this third and final installment, we consider distributed systems that represent shear stress in fluids, heat transfer in uniform one-dimensional media, and subthreshold nerve depolarization. Classic electrochemical analysis and impedance spectroscopy are also reviewed from the perspective of fractional calculus, and selected examples from recent studies in neuroscience, bioelectricity, and tissue biomechanics are analyzed to illustrate the vitality of the field. PMID- 15651637 TI - Compatible and counteracting solutes: protecting cells from the Dead Sea to the deep sea. AB - Cells of many organisms accumulate certain small organic molecules--called compatible and counteracting solutes, compensatory solutes, or chemical chaperones--in response to certain physical stresses. These solutes include certain carbohydrates, amino acids, methylamine and methylsulphonium zwitterions, and urea. In osmotic dehydrating stress, these solutes serve as cellular osmolytes. Unlike common salt ions and urea (which inhibit proteins), some organic osmolytes are compatible; i.e., they do not perturb macromolecules such as proteins. In addition, some may protect cells through metabolic processes such as antioxidation reactions and sulphide detoxification. Other osmolytes, and identical or similar solutes accumulated in anhydrobiotic, heat and pressure stresses, are termed counteracting solutes or chemical chaperones because they stabilise proteins and counteract protein-destabilising factors such as urea, temperature, salt, and hydrostatic pressure. Stabilisation of proteins, not necessarily beneficial in the absence of a perturbant, may result indirectly from effects on water structure. Osmotic shrinkage of cells activates genes for chaperone proteins and osmolytes by mechanisms still being elucidated. These solutes have applications in agriculture, medicine and biotechnology. PMID- 15651638 TI - Near-field optics and spectroscopy for molecular nano-imaging. AB - Application of near-field optical microscopy with a sharp metallic probe to Raman spectroscopy brings microanalysis of materials to their nano-identification and imaging. The local plasmon polariton excitation on the probe tip results in the localization and amplification of the optical field at the vicinity of the tip. The tip-enhanced near-field Raman spectroscopy has analyzed DNA base molecules and single-walled carbon nanotubes (SWNTs) with the nanometric spatial resolution and sufficient sensitivity. Combined with tip pressurization and nonlinear effects, the tip-enhanced near-field Raman spectroscopy gives additional spectral information or improves the spatial resolution and sensitivity. This article introduces the recent progresses on the tip-enhanced near-field Raman spectroscopy and imaging. PMID- 15651639 TI - Superconductivity. AB - Electrons in metals can self-organise. The complex interplay between lattice dynamics, electrostatic interaction and band structure brings forth numerous types of electronic order. Because of its spectacular phenomenology, superconductivity has enjoyed a central place among these, since its discovery nearly 100 years ago. This short introduction into one of the largest fields of condensed matter research focuses on the most fundamental experimental signatures of superconductivity--perfect conductivity and perfect diamagnetism--and their explanation. A conventional broken symmetry argument is presented, which introduces a superconducting order parameter in analogy to the case of superfluid 4He, and discusses its microscopic origin in the framework of the BCS model of superconductivity. New materials have brought to light novel forms of superconductivity. Many cases are now known which fall outside the orthodox BCS model, ranging from the high temperature superconductors, to various organic and d- and f- metal compounds. The article presents key concepts from this intense area of research and touches on the equally puzzling behaviour of many of these materials above their superconducting transition temperature. PMID- 15651640 TI - Guidelines for health research development in Brazil. PMID- 15651641 TI - Trends and priorities in nursing research. AB - This document initially presents an introduction to the global scenario in which nursing research is set today. Next, the main nursing research priorities are described in accordance with the agendas of international health and nursing organizations on different continents, as well as the characteristics of nursing research in Latin America until the end of the XXth century, based on studies and publications in which the main thematic tendencies are identified. Finally, the author reflects on the priority definition process in nursing knowledge production, with a view to attending the need for foundations of the subject area as well as public health needs. PMID- 15651642 TI - [Health aspects in adults over 80 with independent community life: an Australian perspective]. AB - This qualitative study describes the factors that influence the health care practices of 14 elderly persons living independently in Perth, Western Australia. Through in-depth interviews with the participants, the health care practices people were explored. Five distinct themes emerged from the data. These themes were the past time, an active life, nutrition, getting old, economical security and personal satisfaction. This study provides an insight into the life of older adults and the problems associated with ageing. PMID- 15651643 TI - [Mother's difficulties in breastfeeding premature babies at a neonatal ICU]. AB - This study aimed to describe the care and difficulties mothers face when breastfeeding their premature babies at a neonatal intensive care unit (ICU). Therefore, a qualitative approach was adopted. Data were obtained through structured interviews, which were recorded with five mothers of premature babies under 2,000 g who were assisted at the neonatal ICU in Cuiaba-MT, Brazil. The results revealed five subthemes: maintaining milk production by milking, facing discomfort during the stay at the neonatal ICU, living with medical food procedures for premature babies, facing the fragility of the premature baby and the neonatal ICU environment and needing logistic support for breastfeeding. A lack of systematic interventions was observed to stimulate breastfeeding while mothers and their premature babies are in hospital. Hospital professionals need to be trained and intervention protocols have to be implanted, in view of the maternal experiences and needs and the articulation between intrahospital and interinstitutional actions. PMID- 15651644 TI - [Newborn children under phototherapy: the mother's perception]. AB - Since 1958, phototherapy has been used as a method to cure jaundice, which is still an important disease in newborn children. Supported by a phenomenological and qualitative approach, this study aims to investigate the mothers' perception of the phototherapy treatment their children are submitted to. Research subjects were ten mothers of newborns under phototherapy treatment at the Neonatological Hospitalization Unit of a public maternity in Fortaleza-CE, Brazil. Data were collected between May and July 2002. We used group meetings with the mothers as suggested by Carl Rogers. Discourse was organized into categories according to Bardin, which revealed themes that were analyzed in view of Paterson's and Zderad's humanistic nursing theory, as follows: mothers' knowledge on phototherapy and concerns about the treatment. We concluded that the analyzed mothers' major concern is related to the babies' vision. PMID- 15651645 TI - [The qualification of nursing attendants: transformations in work and life]. AB - This study examines the transformations that occurred in the work, life and especially in the way of thinking and acting of nursing attendants after their professional qualification through the Projeto Larga Escala (PLE). Subjects were attendants who qualified as nursing auxiliaries through the PLE, which was conducted by the Regional Health Administration-5 (Sao Paulo city), from 1990 to 1992. The study used life stories as a strategy to apprehend reality, and work and qualification were used a analytical categories. The analysis allowed us to recognize the possibilities and limitations of the training process that guarantee the attendants with: promotion to a professional category and conquest of some rights; knowledge improvement and care humanization. The qualification process seems to have some power to modify the quality of health care, although it does not seem capable to promote changes in the health care model. PMID- 15651646 TI - [The representations of nursing auxiliaries and technicians about team work at an intensive-care unit]. AB - This study aims to explain and analyze the representations of nursing auxiliaries and technicians about team work at an intensive-care unit. The qualitative approach was used and the Theory of Social Representations was adopted as a theoretical-methodological reference framework. The empirical material was obtained from interviews with four technicians and twelve nursing auxiliaries working at the intensive care units of Medical, Surgical and Pediatric Clinics. The information obtained was analyzed by means of the content analysis technique, following the thematic analysis mode. Results indicated conflicting interaction among nursing workers, highlighting the need to implement more efficient staff management models that focus on conflict management actions among different nursing teams. PMID- 15651647 TI - [Freire's contribution to practice and critical education in nursing]. AB - The authors present some biographical data about Paulo Freire, his main works and his most important production, showing the fertility of his legacy. This article aims to discuss the articulation of some concepts used by Freire in nursing practice and education, offering reflections for turning professional practice more critical and creative on the basis of this Brazilian educator's ideas. PMID- 15651648 TI - [Quality of life of nursing students]. AB - This research involved six nursing courses located in the South of Brazil and aimed to find out and evaluate the quality of life of the students. The model WHOQOL Bref model of the World Health Organization was adopted for data collection and analysis and revealed sensitivity for the intended diagnosis. The results indicated that 64% of the students mention satisfaction with their quality of life, but 36% present significant problems, which demand specific needs and justify the implantation of support programs to face the situations of suffering. PMID- 15651649 TI - [Characterization of inpatients at the medical and surgical units of the University of Sao Paulo at Ribeirao Preto Medical School Hospital, according to the level of dependence on nursing care]. AB - This study characterized inpatients' level of dependence on nursing care at the medical and surgical hospitalization units of the University of Sao Paulo at Ribeirao Preto Medical School Hospital, Brazil, from April to June 2002. It also identified the mean bed occupation rate in the units under study. Perroca's Patient Classification System (2000) was used. Nine thousand, seven hundred and nineteen patients/day were classified at the medical units and 8,667 at the surgical units. In both the medical and surgical hospitalization units, there was a predominance of patients classified under the minimum level of nursing care with 70.3% and 66.9%, respectively. The mean bed occupation rate varied from 54.3 to 93.1% at the medical units and from 43.4 to 60.2% at the surgical units. The study allowed for the acquisition of knowledge about the clients with respect to the demand for nursing care with a view to the further development of nursing staff dimensioning. PMID- 15651650 TI - [Nightingale's and Semmelweis' postulates: vital power and contagion prevention as strategies to avoid infections]. AB - This qualitative research used observation as a data collection technique, guided by an instrument elaborated on the basis of Nightingale and Semmelweis' postulates: vital power/life and prevention/contagion. A brief history of infection control is offered from the focus of prevention/contagion. The postulate vital power/life is discussed, presenting the nursing care components deriving from it with a view to care development. This study recommends that both postulates--vital power/life and prevention/contagion--be considered in an articulated way as strategies to avoid infections. PMID- 15651651 TI - [Different approaches to the family in the context of the family health program/strategy]. AB - This study presents the scenario that favored the inclusion of the family as a care focus in public policies. The strategies to interrupt the impoverishment and vulnerability of families in the XXth century occur in a different form, according to different "welfare states" in capitalist societies. However, in view of the welfare state crisis and the increasing costs of public and private services and privates, at least a partial family solution is required in terms of reducing its dependency. The Family Health Program (PSF) put the family on the Brazilian social policy agenda in 1994, reflecting interests from the neoliberal model as well as from solidary social forces. This inclusion generated different approaches, such as: family/individual; family/home; family/individual/home; family/community; family/social risk; family/family. These approaches, due to the lack of a mutual dialogue, end up composing an insufficiently identified picture, thus turning care more difficult. The conditions indicated here should be examined as a way of giving a true chance to the family PMID- 15651652 TI - [Inter-subjectivity on the qualification of health care]. AB - This theoretical production, based on academic debates concerning health care processes, offers elements to support the qualification of individualized care. Arguments in favor of the recovery of inter-subjectivity in care are developed, focusing on sociocultural and psycho-affective aspects of interaction and considering them as an essential dimension in the construction of a new relation with the people who receive care, as well as a broad approach to their health experiences and needs. PMID- 15651653 TI - [Explanatory models for alcohol and drugs use and the nursing practice]. AB - Nursing care practice, research and teaching have led to the accumulation of considerable experience in care for patients with problems related to alcohol abuse. This experience has shown that, among nursing professionals in general, there is a great need to acquire theoretical knowledge, to review the models or informal protocols that support practice in this area. This article aims at examining literature on the main theoretical models to explain alcohol and drugs abuse. It is concluded that, through an understanding of the explanatory models, nurses can seek means of facilitating their practice according to the health demand needs. PMID- 15651654 TI - [Methodological education and care strategies in basic health care]. AB - This paper discusses methodological and care strategies or tools used in basic health care practice. It is based on the dialogue established between what we think and what we carry out at the Life Quality Promotion Outpatient Centers (APQVs). These centers are located at two basic health care centers in Porto Alegre/RS, Brazil. Its users are mostly adult and elderly patients with long-term illnesses. The proposal of this discussion arose from a research project financed by the Brazilian Council for Scientific and Technological Development--CNPq, and integrates a thematic network called Education and Care Methodologies to Promote Life Quality. Starting from this empirical and conceptual base, methodological tools were built to develop nursing consulting services in outpatient health care to individuals and groups. This article aims to present relational and operational concepts used in care at these services. PMID- 15651655 TI - [Reflections on learning the educational role in the nursing formation process]. AB - Reflection in view of the new curricular lines for the undergraduate nursing course, aimed at highlighting the characteristics pointed out in the document on the role of educator in nursing formation and discussing the implications of the teacher-student relation in this process. PMID- 15651656 TI - [Clinical significance of examinations of hemodynamics of mother and fetus in physiological and complicated pregnancy]. AB - Summarized in the paper is the experience of defining the clinical significance of examination of hemodynamics of mother and fetus in physiological and complicated pregnancy. The normative parameters of blood circulation in the vessels of mother and their central hemodynamics are described. An interrelation between the changes of central hemodynamics of mother and fetus are shown in the development of gestosis and placental insufficiency. The orientation, succession and changes of blood circulation in the arteries and venous vessels of fetus are demonstrated in chromosomal anomalies, congenital malformation, premature pregnancy, gestosis, fetoplacental insufficiency and postmaturity. It was proven as prognostically valuable to study the parameters of blood circulation of mother and fetus not only in diagnostically complicated pregnancy but also in evaluating the therapy efficiency, as well as in choosing a method and time for delivery and in prognosticating a perinatal outcome. PMID- 15651657 TI - [Results and outlooks of using cell technologies in the treatment of neurological diseases]. AB - An attempt was undertaken in the last decade of the 20th century to use a principally new approach to the treatment of neurological diseases--cell therapy. Main efforts were focused on developing a method related with replacement of neurons dying in neurodegenerative pathology, primarily, in Parkinson disease (PD). Outlined below are the key elements of the technology:--ensuring, in experiment, of a prolonged therapeutic effect in transplantation, to the affected part, first of embryonic neurons of the animal of the same species (allografting) and then of homologous embryonic neurons of man (heterografting);--obtaining, standardization and preparation (for transplantation) of embryonic nervous tissue of man; transplantation of embryonic nervous tissue of man to the brain of patient and evaluation, in situ, of the functional activity of its neurons; and evaluation of the therapeutic effect of grafting. Cell suspension of meseencephalon of 6-9 week human fetus containing around 10% of differentiating dopaminergic neurons was used for grafting in PD. Embryonic dopaminergic neurons, administered stereotactically into the striatum of patient, established synaptic links with neurons of the recipient, which was accompanied by the onset of synthesis and reverse uptake of dopamine (DA) as well as by the onset of spontaneous and stimulated release of DA. Neurografting ensured a temporary improvement of the condition in a part of PD patients but did not cure them. Moreover, such positive therapeutic effect was registered only in patients with the akineticorigid but not trembling variation of the disease. Hence, although there was a certain progress in clinical neurografting, the approach cannot be now recommended for introduction in neurology and neurosurgery. The limited therapeutic effect of the treatment method is primarily explained by a low rate of survival of transplanted dopaminergic neurons and, consequently, by the persisting DA deficit in patient's body. Therefore, the outlooks for perfecting the cell technology are related with increasing the survival rate of implanted dopaminergic neurons and with stimulating the innervation of target neurons in patient's striatum as well as with using the neural (glia) and non-neural (fibroblasts, myoblasts) cells with modified gene and stem cells. Finally, despite a certain progress of advancing the cell technology in neurology the approach still needs more research, which would enable further clinical trials. PMID- 15651658 TI - [Results of Befnorin clinical trails in healthy volunteers]. AB - Clinical trails of Befnorin based on the human recombinant TNF-beta elaborated at the Research Design and Technology Institute of Biologically Active Substances, "Vector" State Research Center of Virology and Biotechnology, were carried out on healthy volunteers in compliance with a decision passed by the Committee of Medical and Immunobiological Preparations, Russia's Health Ministry. Single Befnorin doses of 5-10(4) U, 10(5) U, 5-10(5) U, and 10(6) U were administered as intramuscular injections. Clinical, biochemical and immunological parameters were registered for 7 days after a single dose. The drug had an impact on the below immunity indices: Fc-phagocytosis of monocytes, migration index and migration inhibition index. The dose of 10(5) U was proven to be most effective and safe. Supposedly, the drug can be effective in the treatment of herpetic diseases. PMID- 15651659 TI - [An experimental study of possibility of treatment of hemorrhagic fever Marburg by Remicade]. AB - The effect of Remicade, administered according to different schemes, produced on the course of experimental Marburg hemorrhagic fever was studied. When the drug was used from the first infection day, the treated animals died reliably earlier versus the controls (infected animals). A reliably lower concentration of TNF alpha in blood serum (versus the controls) was registered on day 3. At the same time, when Remicade was used from day 3 after infection, a 50% survival of animals was registered. It is noteworthy, that the TNF-alpha concentration in blood serum did not differ, on day 3, from that of controls, whereas, beginning from day 5 after infection, the animals displayed a downtrend of TNF-alpha concentration. Hence, it can be an evidence of a dual TNF-alpha role in the immunopathogenesis of Marburg hemorrhagic fever: on day 1 after infection the production of TNF-alpha is needed; whereas, if there is an overproduction of the cytokine, it is necessary to inhibit it. The treatment scheme in hemorrhagic fever must, apparently, comprise both elements of cytokine therapy and drugs affecting other chains of pathogenesis--they must mainly protect the endothelial vascular cells. PMID- 15651660 TI - [Functional polymorphism of p53 and CCR5 genes in the long-lived of the Siberian region]. AB - The P53 protein is a key regulator of modified-cell apoptosis. The functional oligonucleotide polymorphism of the p53 gene causes the substitution of arginine (Arg) for praline (Pro) in the codon 72. A reduced apoptotic activity of p53 and, as a consequence, development of oncology pathology is associated with the above polymorphism. CCR5 is a compound transmembrane receptor-protein, which apart from chemokines, binds with some molecules and is a coreceptor for HIV-1. 32 bp deletion within the CCR5 encoding region results in the loss of the protein's receptor function. It has been demonstrated that the transmission of the "external" (in respect to cell) stimulus, via the CCR5 system, induces expression of the p53 gene and initiates apoptosis. Allele variants and p53 and CCR5 genotypes (separately and in combinations) were investigated, within the present case study, for 131 long-livers from Novosibirsk and Tyumen Regions. A trend was detected towards accumulation of the p53 Pro alleles in association with the CCR5del32 allele in the study group, which, as the authors believe, can enhance the genome resistance to variable factors that cut the life span. PMID- 15651662 TI - [Outlooks for using the interferon inducers in the treatment and prevention of influenza and acute respiratory viral diseases]. AB - The methods of treatment by using the stimulators of the non-specific resistance system have been growing even more topical. Drugs based on interferon (INF) and its inducers belong to the group. IFN is of key importance in the regulation of antiviral immunity. The use of IFN inducers can be referred to as the most promising approach to enhancing and activating the body resistance mechanisms. The advantages of IFN inducers before IFN drugs, a wide spectrum of antiviral effects, compatibility with many pharmaceuticals, good tolerability by patients and effectiveness in different administration modes served as a basis for designing a new class of antiviral drugs--IFN inducers. A lot of experimental medical-and-biological research was made to evaluate the efficiency of IFN inducers in influenza. They were shown to possess a pronounced therapeutic-and preventive action in experiment and clinical trials. Synthetic and natural dsRNA belong to IFN inducers. Multiple research made in cell culture and with experimental animals demonstrated a high interferon-inducing and antiviral activity of double-stranded RNA against influenza virus. A number of dsRNA-based IFN inducers are permitted for clinical use. Data obtained in clinical trials of such inducers are indicative of their pronounced therapeutic-and-clinical effect. Hence, the IFN inducers having different structures and origins can be regarded as promising in the treatment and prevention of influenza and acute respiratory viral diseases. PMID- 15651661 TI - [Experimental evaluation of possible transmission of human alpha-2 interferon artificial gene to the other microorganism species]. AB - Described within the case study are experimental evaluation data on the remote results of introduction of gene-engineered microorganisms (GEM) of the B-7092 B. subtilis strain into the environment with the follow-up ranging from 1 to 7 years. No bacteria of the above GEM were detected (among selected 19 bacterial cultures) in soil samples from an agricultural farm, where the VETOM 1.1 probiotic was used for the treatment and prevention of cattle; primary and precise identification (two-round PCR with specific primers) was applied. The introduction of the B-7092 B. subtilis strain into the environment did not result in its unlimited growth and proliferation in soil. No transfer of plasmid genes (genes of leukocytic human alpha-2 interferon) from the B-7092 B. subtilis strain GEM to microorganism species available in the environment (soil) was registered. No recombinant plasmid DNA from lysed GEM cells was detected in soil samples. PMID- 15651663 TI - [Experimental evaluation of delivery means of potential anti-HIV microbicides]. AB - The purpose was to develop an effective drug of local intravaginal membranetropic microbicide based on polycarboxylate and matrix modified by norbonene and/or peptide (simulator of CCR5 coreceptor). The drug was added to the composition of polyacrykic acid-polyvinylpyrrolidone (interpolymer complexes) in order to prevent the interplay of its non-specific groups with proteins. The antiviral effect and absence of toxicity of the finished microbicide form were demonstrated on the human lymphoid MT-4 culture. PMID- 15651665 TI - [Development of microencapsulating measles live vaccine]. AB - Designing of non-injection methods of immunization against measles has recently turned into a topical issue. Development of mucosal vaccines ensuring the "entry gate" immunity, which is highly effective in airborne infection, is in the focus of attention. The authors developed a method of microencapsulating the viral particles into the matrix of pH-dependent polymers. Microencapsulated live measles vaccine shaped as 0.6-2.0 microm particles was obtained. The specific activity of measles virus in the drug was 3.36-4.31 log TCD50/0.5 ml. In subcutaneous immunization of guinea pigs with capsules, the best results were obtained in a single administration of vaccine based on ethylcrylate, sodium alginate/ chitosan and sodium slaginate/HMDA. In the intranasal administration of vaccine based on sodium alginate/spermin and sodium alginate/HMDA, there was a need in 2 and 3 stages of immunization. PMID- 15651664 TI - [The anti-HIV activity of glycyrrhizic acid penta-O-nicotinate]. AB - The anti-HIV activity of niglizin (penta-O-nicotinate of glycyrrhizic acid) and of its combinations was studied in the culture of infected MT-4 cells and in respect to the recombinant reverse HIV-1 transcriptase. Niglizin was shown to suppress effectively the HIV replication and to be a noncompetitive inhibitor of reverse transcriptase. Research of a combined anti-HIV action of niglizin and of azidothimidine (AZT) demonstrated that the preparations, when used at ratios of 1:20, 1:50, 1:200 and 1:2000, suppressed the synergetic effect both in the cell culture and in the recombinant reverse HIV transcriptase. A study of the antiviral activity of niglizin and of its joint use with AZT in respect to the AZT-resistant HIV-1 mutant showed niglizin to be more effective (ID50 = 0.134 microM) versus the "wild" strain. (ID50 = 9.64 micriM); whereas, its combined use AZT:niglizin = 1:100 displayed synergism (FIC = 0.553). The efficiency of the combined AZT/niglizin anti-HIV effect both in respect to the "wild" strain and to the AZT-resistant mutant confirms that such combinations are promising for the treatment of HIV infection. PMID- 15651667 TI - Expression and cytotoxicity of EGFP-labeled D-amino acid oxidase in HeLa cells. AB - To observe the expression of R. gracilis D-amino acid oxidase (DAAO) gene labeled by EGFP in HeLa cells and how it mediates cytotoxicity, DAAO cDNA and EGFP gene were cloned into Eukaryotic expression vector pIRES to construct DAAO gene expression vector pIRES-DAAO. HeLa cells were transfected with pIRES-DAAO in vitro. Expression of EGFP gene in HeLa-D cells was observed under a fluorescent microscope. The efficiency of transfection was analyzed by flow cytometry (FCM). The fluorescent cells were screened by FCM and were named HeLa-D. The activity of HeLa-D cells was detected by MTT colorimetric assay after they were treated with D-Ala at various concentrations. The results indicted that expression of EGFP gene in HeLa-D cells was seen under fluorescent microscope and HeLa-D cells were screened by FCM. Apparently,the prodrug D-Ala killed HeLa-D cells. These results demonstrate that EGFP gene can be regarded as a reporter gene to screen the cells transduced with DAAO quickly, and DAAO/D-Ala suicide gene system may prove helpful in gene therapy of cancer. PMID- 15651666 TI - [Designing of a candidate edible vaccine against hepatitis B and HIV on the basis of a transgenic tomato]. AB - The synthetic chimeric gene TBI-HBS encoding the synthesis of immunogenic ENV and GAC epitopes of HIV-1 (immunogenes of T- and B-lymphocytes) and of the surface protein (HBsAg) of the hepatitis B virus was introduced into tomato plants var. Ventura by agrobacterial vector pBIN35TBI-HBS; transgenic tomato plants with the integrated gene TBI-HBS were generated. The integration of the TBI-HBS target gene was confirmed by PCR. The synthesis of antigenic proteins of TBI and HBsAg in fruits of transgenic tomato plants was displayed by immunoassay. The fruits of transgenic tomato plants were fed to experimental mice with a 1-week interval. On days 14 and 28, there was discovered a sufficiently high content of antibodies to the antigenic proteins of HBV and HIV-1 in serum of experimental animals. Antibodies were found in feces of experimental mice; no antibodies were found in the control group of mice. Hence, it was established that the TBI (HIV-1) and HBsAg (HBV) antigens were synthesized in transgenic tomato fruits due to the integrated construction of pBINNp35TBI-HBS in an amount that was enough to induce the immunogenic response in mice to the oral delivery of edible vaccine. PMID- 15651668 TI - Expression of human soluble gp190 in yeast Pichia pastoris. AB - The cDNA of N-terminal extracellular domain of human leukemia inhibitory factor receptor a-subunit (gp190) ,which encoded soluble gp190 (sgp190) ,was cloned into the methylotrophic yeast Pichia pastoris secreted expression vector pPIC9K. The linearized plasmid sgp190-pPIC9K was then integrated with GS115 genome by spheroplasts transformation. MD medium and PCR method were used for screening positive transformants. The transformants (His+ Mut+) containing multi-copy gene insertion were selected with increasing concentration of antibiotic G418 and induced by 0.5% methanol in shaking flask to express sgp190. The sgp190 expressed level could reach up to 26% of total proteins in culture supernatant. SDS-PAGE and Western blotting analysis showed that the molecular weight of sgp190 is about 125 kD and could be specifically recognized by polyclonal antibodies against human sgp190,which indicated good antigenicity of this secreted expressed protein. Biological activity assay showed that purified sgp190 could inhibit the effect of LIF on hCG biosynthesis of cytotrophoblast in vitro. PMID- 15651669 TI - [Detection of a novel mutation in COL4A5 gene from a Chinese family with X-linked alport syndrome]. AB - By means of PCR and direct sequencing, all 51 exons and their neighbouring intronic sequences of the COL4A5 gene were analyzed to detect mutations in 17 members from a Chinese family with X-linked Alport syndrome(XLAS). At the position 2240 in exon 26, a single-base deletion(2240deIC) is found in all male patients, and a heterozygous deletion is found in all female patients, whereas no mutation is found in normal and 80 control individuals. Meanwhile, the corresponding PCR products of female patients are cloned and sequenced to confirm the results. It is concluded that the 2240deIC mutation is the underlying cause of XLAS in this family,not a polymorphism. Furthermore,this single-base deletion mutation in COL4A5 gene is first reported in X-linked Alport syndrome. PMID- 15651670 TI - [ACE gene polymorphisms associated with serum ACE activity in the Henan Hans of China]. AB - In order to investigate the relationship between gene polymorphisms and serum activity of the angiotensin-1 converting enzyme(ACE) in Chinese population, eight loci in ACE gene were detected by polymerase chain reaction combined with restriction fragment length polymorphism or electrophoresis directly, and serum ACE activity was measured by spectrophometry. Using maximum likelihood estimation(MLE), the pattern of intragenic linkage disequilibrium and the haplotype structure were estimated. All the eight polymorphisms formed nine haplotypes,in which the two most frequent haplotypes were A (A-T-A-T-G-I-A-3) and B(C-C-T-C-A-D-G-2). Also haplotype A and haplotype B were completely different at all eight sites. Deduced from the maximum-parsimony tree,the haplotypes in Chinese population could be grouped into three clades. Clade III seemed to be generated by an ancestral recombination event between clade I and clade II. Every locus formed, the haplotype B was associated with high ACE activity. This findings suggested that serum ACE levels were partially determined by genetic predisposition (the polymorphism of ACE gene). Haplotype B may be related to the quantitative trait loci of high-level activity,whereas high-resolution genetic mapping linked with ACE was still necessary to characterize definitely the functional variants. PMID- 15651671 TI - Study on polymorphisms of microsatellites DNA of six Chinese indigenous sheep breeds. AB - The polymorphisms of 17 microsatellites loci of six indigenous sheep breeds, including Mongolian sheep, Ujumuqin sheep, Kazakstan sheep, Aletai sheep and Tibetan sheep, were studied using polypropylene gel electrophoresis in order to investigate their genetic diversity,origin, differentiation and relationships. The results indicated that there was a significant difference in genetic diversity between different loci (P < 0.01). There was no significant difference in PIC, Fis and observed heterozygosity (Obs. Het) among populations (P > 0.05), but a significant difference in gene diversity and expected heterozygosity (Exp. Het) (P < 0.05). Chinese indigenous sheep breeds had similar genetic diversity as those from Europe,but with higher inbreeding coefficient. It could be inferred from the cluster of individuals and populations that Chinese indigenous sheep breeds might originate from two ancestors. The cluster of populations showed that Mongolian sheep and Ujumuqin sheep had close relationship and did not differentiate obviously. Mongolian sheep and Tibetan sheep had far relationship and differentiated significantly. Aletai sheep differentiated from Kazakstan sheep but not significantly. Tan sheep,Aletai sheep and Tibetan sheep also had close relationships. The Fst of Chinese indigenous sheep breeds was close to some Spanish sheep breeds,but much smaller than that of other European sheep breeds. More loci and samples should be studied in the future in order to obtain more accurate results. PMID- 15651672 TI - [Sequencing and analyzing of expressed sequence tags (ESTs) of pig fat tissue]. AB - In this paper,the ESTs of pig fat tissue were sequenced and analyzed using the large-scale DNA sequencing method, 7790 high quality ESTs were gained,from which 4354 genes were obtained using cluster analysis with the STACK-PACK software, including 3609 singlet genes and 745 multicopy genes. The 4354 candidate genes were compared with BLASTN to the nr library (e =1e-10), of which 2712 were known genes, containing 1 987 singlet genes and 725 multicopy genes,there are 2 109 unknown genes and new ESTs. Based on the results of BlastN and the index of GenBank Accenssion No., the known gene expression profile of pig fat were constructed. It showed that the genes participating in metabolism held the highest proportion in the 7 sorts, and in some aspects showed the hearty metabolism activity of fat tissue. 257 total reads of the Major Histocompatibility Complex (MHC) genes in the pig fat tissue were found,including 181 singlet genes and 44 multicopy genes. They account for 44.9% of the cell and organism defense, and 5.4% of all known genes,from this we can see that the MHC has a fairly high expression degree. Comparing all ESTs related with MHC (257reads) with known pig BAC we found the partial sequences (approximately 200 bp) of the ESTs distributing to every exon of. We can predict that the ESTs in MHC contain approximately 200 bp highly consistent regions and every exon possesses one of these regions. Although these MHC sequences in different BAC have different protein domains,they are highly consistent and related with immune functions. When the MHC genes transfer to the next generation,they can duplicate repeatly and inherit stably. PMID- 15651673 TI - [Comparative study on lipogenic and lipolytic gene expression in intramuscular fat tissue between growing Erhualian and large white pigs]. AB - To investigate the mechanism of difference in intramuscular fat deposition between Erhualian and Large White pigs,single tube relative-quantitative RT-PCR method was used to investigate the development patterns of lipogenic (ACX, LPL, ME) and lipolytic (HSL) gene expression with 18S internal standard control. Sixteen Large White boars and twenty Erhualian boars were selected and raised according to normal nutrition standard respectively. The animals were selected randomly and slaughtered at 15 kg, 40 kg, 60 kg and 90 kg for Large White pigs and at 18 kg, 40 kg, 60 kg, 80 kg and 90 kg for Erhualian pigs respectively; with four animals of each breed at each time. The supraspinatus and semimembranosus muscles were removed for total RNA extraction and longisimus dorsi muscle for intramuscular fat (IMF) analysis using ether extract method. The results showed: (1) The property of IMF between Erhualian and Large White boars was similar during early growing period (before 40 kg) (P > 0.05) ,thereafter, IMF level of Erhualian boars increased dramatically to 4% at 60 kg and over 5% at 90 kg while Large White boars kept steadily at about 2% (P < 0.05); (2) The pattern of lipogenic and lypolytic gene expression was similar between semimembranosus and supraspinatus muscle in each breed; (3) The tendency for LPL and ME mRNA expression coincided with that of IMF development among 20 approximately 60 kg in Erhualian pigs. The results suggest that the development of IMF between 20 kg and 60 kg in Erhualian pigs may play a meaningful role in deposition of IMF,and that the expression of ME and LPL mRNA may contribute to fast sediment of IMF in Erhualian pigs. PMID- 15651674 TI - Genetic variability and relationships in mitochondrial DNA CO II gene sequence of red common carps in China. AB - Nucleotide sequence of Cytochrome coxidase subunit II (CO II) of mitochondrial DNA ( mtDNA) was analyzed from four populations of the red common carp (C. c. var. singuonensis, C. c. var. wananensis, C. c. var. wuyuanensis and C. c. var. color) as well as one population of wild common carp in China to detect their genetic variability and phylogenetic relationship. Nineteen nucleotide variable positions were detected in analyzed 607 bp length sequence and 15 haplotypes were defined among five populations,but only one shared haplotype was observed in C. c. var. singuonensis and C. c. var. color, respectively. The characteristic intrapopulation single nucleotide polymorphisms (SNP) were observed in 103th, 149th, 239th, 368th, 395th nucleotide position. The highest haplotypic diversity (h) and nucleotide diversity (pi) existed in C. c. var. wuyuanensis and the lowest exited in C. c. var. wananensis. The significant fixation indices (Fst) in most comparisons between populations were observed. The neighbor-joining trees from haplotypes and populations of red common carp indicated C. c. var singuonensis and C. c. var. color might have originated from one monophyletic group, while C. c. var. wananensis and C. c. var. wuyuanenesis might have originated from an independent evolutionary branch. PMID- 15651675 TI - [Population genetic structure of Pelophylax nigromaculata in Chinese mainland based on mtDNA control region sequences]. AB - Molecular genetic data were used to investigate population genetic structure and differentiation of Pelophylax nigromaculata (Anura: Ranidae). Frogs were collected from 12 localities across Chinese mainland. Sample sizes of up to 10 frogs per population were assayed for mitochondrial control region sequence variation. The aligned 685 bp of the 5'CR include 111 variable sites. Sixty-seven haplotypes were defined. Most of the haplotypes are unique to local populations, only 7 of the 67 are shared among a few local populations. The overall species haplotype diversity is quite high (h = 0.98 +/- 0.005), and the nucleotide diversity is also high (pi = 0.0303 +/- 0.0029). This is corresponding to huge population size and extensive distribution of Pelophylax nigromaculata throughout Palaearctic and Oriental Regions. Phylogenetic tree of mtDNA control region haplotypes based on maximum parsimony algorithm and reconstructed phylogenetic relationships among the local populations based on neighbor joining algorithm all suggest that the haplotypes of Jiling and Liaoning populations in northeastern China have a sister relationship with 10 local populations in northern, central, southern and southwestern China. The analysis of molecular variance (AMOVA), the pairwise FST values and nucleotide divergence all support significant population subdivision between the Jilin-Liaoning group and multi-population group. The most possible cause of the significant genetic subdivision between the two groups might be referred to the effect of Quaternary glaciation. The possible reasons for the lack of obvious geographic structure on the whole in the multi-population group and the genetic differentiation in some local populations of the multi population group were also discussed. PMID- 15651676 TI - [Differential acute mortality among the allozyme genotypes of Oxya chinensis by pesticide avermectin]. AB - The rice grasshopper Oxya chinensis exhibits polymorphic loci at Ldh, Gpi, Pgm and Me. The data of the mean number of alleles per locus (A = 2.8), percentage of polymorphic loci (P = 80.0%), the observed mean heterozygosities (Ho = 0.271 approximately 0.279) and the expected mean heterozygosities (He = 0.305 approximately 0.316) of the species suggest that O. chinensis possesses sufficient genetic diversity. It was hypothesized that the high polymorphisms at Ldh, Gpi, Pgm and Me might make it possible for pesticide avermectin to act as a selective agent through differential lethality among the insect individuals with different genotypes. In this study a total of 855 grasshoppers were injected with avermectin (1.3 x 10(-2) g/g) to obtain a mortality of 54% after 24 hours. The allozyme analysis was then employed to determine the genotypes of Ldh, Gpi, Pgm and Me for both dead and surviving individuals. Contingency table chi2 tests showed that avermectin displayed random lethal effects on the genotypes at the loci of Ldh, Pgm and Me, without correlation between the genotype and mortality. In contrast, at Gpi locus, the grasshopper demonstrated a mortality cline of Gpi AA (38%), Gpi-AB (51%), Gpi-BB (58%) and Gpi-BC (74%). The significant mortality differences were found among the following genotype pairs: Gpi-AA vs. Gpi-BB, Gpi AA vs. Gpi-BC and Gpi-AB vs. Gpi-BC. These data implied the Gpi-AA genotype was likely related to the specie's resistance to the pesticide avermectin. It was also noted that the Gpi-A allele was present in the genotypes with low morality,while Gpi-B was present in the genotypes with moderate mortality, and the individuals with Gpi-C allele exhibited the highest mortality. The data obtained in this study suggested that the increasing proportion of Gpi-AA genotype and perhaps Gpi-A allele in a population may be useful as a potential resistant biomarker of O. chinensis to pesticide avermectin. PMID- 15651677 TI - [Cloning and sequence analysis of mrjp1 cDNA from Apis cerana cerana]. AB - A cDNA library was constructed from eight-day-old worker heads of Apis cerana cerana. A probe derived from part of Apis cerana genomic mrjp3 segment was used to screen this library. A total of 120 positive clones of mrjps were screened out from the cDNA library with DIG-probe. The positive clones were amplified and sequenced with T3/T7 primer. 12 sequences were similar to Apis cerana india and Apis mellifera mrjp1 gene by BLAST analysis. The cDNA sequence analysis indicated that the homology was 93.78% between Apis mellifera mrjp1 and Apis cerana mrjp1 while the homology was 99.36% between Apis cerana cerana and Apis cerana india. This result confirmed in molecular level that Apis cerana cerana and Apis cerana india had a common ancestor since the relationship between Apis cerana and Apis mellifera was more far. PMID- 15651678 TI - [Maximum likelihood analysis for mapping dynamic trait QTL in outbred population II. Simulation]. AB - Based on the maximum likelihood analysis principle for dynamic trait QTL mapping, selecting three orders Legendre polynomial as sub-model,the effects of the individual number, the frequency of test day,the marker density and heritability on detecting power are investigated by Monte-Carlo simulation. Each factor is divided into three levels such as the high,middle and low,the combinations of which are arranged by using orthologal design. The simulating result shows that the high heritability needs less number of individuals and frequency of test day than the low one, but the number of individuals of more than 300 and the frequency of test day of 5% above can guarantee sufficient detecting power no matter what the heritability is like. The number of individuals has almost the same effect as the frequency of test day in dynamic traits QTL mapping and the number of individuals and the frequency of test day compensate each other under the same size of samples. As to the QTL detection for a dynamic point,simulation indicates that the presented dynamic trait QTL mapping method that focuses on the whole dynamic process is significantly superior to the classical mapping methods on each individual dynamic point. PMID- 15651679 TI - PCR-based screening BAC library and direct end sequencing of BAC clones. AB - A PCR based strategy was developed which required four steps to identify positive BAC clones from barley BAC (bacterial artificial chromosome) library. In the protocol, two levels of BAC DNA pools (super-pool and pool) were prepared for analysis. One pool is made of one plate DNAs and one super-pool is made of mixing ten consecutive 1/100 diluted pool DNAs (1 approximately 10, 11 approximately 20 ect). First,super-pool DNAs were analysed and then 10 pool DNAs contained in every positive super-pool were analysed. Once positive BAC plates were identified,the bacterial cultures were dipped into PCR mixtures and reaction is made to identify positive BAC clones. The BAC clones identified by each marker were grouped into contigs. BAC-end sequence was obtained from BACs within each contig and primers were designed for the next step chromosome walking. In case of the BAC ends belong to repetitive sequence, the primers were designed based on the subcloned unique band in the contig (identified by Hind III digestion pattern). This method allows us to construct the BAC contig without the costly and time-consuming efforts, and no radioactivity harmful to the body. PMID- 15651680 TI - [Study on chromosomes aberration in wheat-rye disomics addition lines induced by the gametocidal chromosome 2C]. AB - In the present study,Chinese Spring-Imperial (1 R-7R) wheat-rye disomic addition lines were hybridized with Chinese Spring-2C (derived from Aegilops cylindrica) disomic addition lines. The F1 hybrids were examined by mitotic and meiotic analysis. There were observed abnormal chromosome configurations. A total of 430 F2 plants were obtained by self-pollination. Chromosomes aberrations, such as translocation, deletions, isobrachial and dicentromere chromosomes, are identified in F2 individual plants by C-banding combined with fluorescent in situ hybridization (FISH). Additionally, chromosome spontaneous substitutions such as 2C substituting for wheat chromosomes 2A, 2B and 2D were also observed. The rule and frequency of chromosome aberration in F2 are the following: 22 out of 430 F2 plants (5.11%) were found involving aberration rye chromosomes. Among them, 10 plants were identified as wheat-rye chromosome translocation lines comprising 2.3%. Rye chromosome deletions comprised 12 of them (2.79%). 3 isobrachial aberrations were detected (about 0.7%), too. Most of the translocation lines are with wheat centromere, only one of them is with rye centromere. Rye chromosome aberrations occurred unevenly among homoeologous groups. There were 5 in 1R, 3 in 2R, 1 in 3R, 3 in 4R, 6 in 5R and 4 in 6R. The majority of the translocation lines are terminal translocation. 54 out of the total 430 progenies are wheat deletions,and 27 are distributed in the A group, 20 in the B group and 7 in the D group respectively. Finally,we discussed the possible cause for the uneven chromosome aberration among homoeologous groups in wheat and rye as well as the effect characteristics of 2C on wheat and rye chromosome. PMID- 15651681 TI - [Stable expression of QTL for grain shape of milled rice (Oryza sativa L. ) using a CSSLs population]. AB - A set of chromosome segment substitution lines (CSSLs), derived from Asominori/IR24 with Asominori as the recurrent parent,was planted and phenotyped for grain length (GL), grain width (GW) and length-width ratio (LWR) of milled rice in two successive years and four sites. QTL for GL,GW and LWR were characterized and stability of their expression was investigated. The phenotypic values for each trait showed a continuous distribution and some transgressive lines were also observed in the CSSLs population. Additionally, a total of 13 QTL for GL, GW and LWR were identified,and six of them were consistently detected in the eight different environments. Phenotypic values were different significantly (P < 0. 001) between the CSSLs harboring any of the six QTL alleles and the genetic background parent, Asominori. Significant phenotypic correlations (r > or = 0.75, r0.05 = 0.67) were detected among different environments for these CSSLs carrying the same target QTL. Also, the results indicated that the six QTL, namely, qGL-3 for GL, qGW-5a and qGW-5b for GW, qLWR-3, qLWR-5a and qLWR-5b for LWR, were stably expressed in different environments. Since QTL qGL-3 and qLWR-3 were mapped in the R19-C1677 interval, qGW-5a and qLWR-5a in the vicinity of RFLP marker C263, qGW-5b and qLWR-5b near R569,the four RFLP markers, R19, C1677, C263 and R569,would be useful for further marker-assisted selection (MAS) in rice quality improvement. PMID- 15651682 TI - [Comparative phylogenetic analysis of the rice and Arabidopsis PHD-finger proteins]. AB - PHD-finger (plant homeodomain finger), a unique Cys4-His-Cys3 pattern zinc containing domain, is widely found in many transcriptional regulation proteins from plants or animals. This protein has many different family members distributing in plant or animal proteome,and plays key roles in the development. Here, we identified 44 different putative PHD-finger proteins in the rice (Oryza sativa) genome and performed a phylogenetic analysis with 45 Arabidopsis thaliana proteins. The analysis led to a classification of the 89 proteins into two major groups based on the number of PHD-fingers. Phylogentic analysis of these proteins indicated that there are common ancestors in both plant and human genomes, from which two or three PHD-finger containing proteins might duplicate. PMID- 15651683 TI - [Construction and expression of vector with aroA-in gene and its transformation in tobacco]. AB - Inteins are intervening protein sequences that undergo self-excision from precursor protein with concomitant joining of flanking sequences. Here, we demonstrated intein cis-splicing in Nicotiana tabacum nuclear genomes by using artificial cis Ssp DnaB and Rma DnaB intein. We want to test whether protein splicing can occur in higher eucaryotic cell,which would play an important role in transgene containment in transgenic plants. Glyphosate-resistant Salmonella typhimurium aroA gene was divided at position 235/236 aa within EPSPS by inverse PCR from pLEPSPS. Amplified gene products with artificial cis-Ssp DnaB/Rma DnaB intein and split-Ssp DnaB/Rma DnaB intein were inserted at position 235 of EPSPS respectively to construct plasmid pLEBC, pLERC, pLEBT and pLERT. Above four aroA In gene fusions were ligated into pET-32 to obtain E. coli expression vectors termed pETLEBC, pETLEBT, pETLERC and pETLERT. E. coli DE3 cells containing individual recombinant plasmids described above were induced by IPTG to produce corresponding protein products. Detectable spliced EPSPS and unspliced precursor demonstrated that splicing occurred in bacteria. aroA-cis SSp DnaB and aroA-cis Rma DnaB were ligated into Agrobacterium tumefaciens binary vector pLYM. Then A. tumefaciens containing EPSPS-(cis) intein cassettes were used for leaf disk transformation in N. tabacum. Integration of aroA-In gene into plant genome was confirmed by genomic PCR analyses. To verify the expression of fusion genes at transcriptional level, RT-PCR analyses were performed and the expected products were identified. These results suggested that plant cells support expression of S. typhimurium aroA-In fusion gene in nulear genomes. Thus,we speculated the existence of protein-splicing activity in plant cells. This opens the possibility of applying intein trans-splicing technique to reduce/prevent gene transfer by way of pollen in transgenic plants. PMID- 15651684 TI - [Isolation and characterization of BcMF3, a gene expressed only in maintainer line in Chinese cabbage-pak-choi (Brassica campestris L. ssp. chinensis Makino var. communis Tsen et lee)]. AB - Expression profiling was performed using cDNA-AFLP technology on floral buds of a genic male sterile line (A line) and a maintainer line (B line) in Chinese cabbage pak-choi (Brassica campestris L. ssp. chinensis Makino var. communis Tsen et Lee cv. Aijiaohuang). A differentially expressed cDNA fragment, BA18-T16, was obtained via selective amplification with A18/T16 primer pair. BA18-T16 was specifically associated with the maintainer line, and was expressed only in floral buds revealed by half-quantitative RT-PCR. The 3' end and 5' end sequences of BA18-T16 were obtained by 3' and 5' RACE amplification and the full cDNA,designated as BcMF3, is 2082 bp long containing a 1755 bp open reading frame. BcMF3 shares high homology with Bp19, a pectin methylesterase from Brassica rapa L. with 99% identity at nucleotide level and 85% identity at amino acid level. Suggesting that BcMF3 most likely encodes a pectin methylesterase in Chinese cabbage pak-choi. PMID- 15651685 TI - [Linkage map construction and mapping of a dominant genic male sterility gene (Ms) in Brassica napus]. AB - A backcross population derived from a cross between the dominant genic male sterile line Rs1046A and the double-low cultivar 'Samourai' was used for linkage map construction in Brassica napus. A linkage map with a total length of 2 646 cM was developed using 138 AFLP markers, 83 SSR markers and one morphology trait, which were distributed over 18 major linkage groups, two triplets and one linkage pair. 11.7% of the mapped markers distorted from the expected 1:1 ratio. The dominant genic male sterility gene (Ms) was mapped in LG10 and surrounded by five closely linked AFLP markers. Meanwhile,two evident marker segregation distortion regions were observed in both LG8 and LG16. The map constructed in the present study and the mapping of Ms gene are highly valuable in designing marker-assisted breeding program for the genic male sterile two-type line in Brassica napus. They are also important to map-based cloning of the Ms gene and to better understanding of the mechanism of genic male sterility at molecular level. PMID- 15651686 TI - Phylogenetic analysis indicates bacteria-to-apicoplast lateral gene transfer. AB - Apicomplexan protozoa contains a highly reduced plastid-like organelle termed apicoplast. Data from clpC gene in apicoplast and their homologs in other plastids and bacteria were used to reconstruct phylogeny of apicoplast. Trees were reconstructed using neighbor-joining, minimum evolution, maximum parsimony and maximum likelihood. The reconstructions robustly support the monophyly of apicoplast and B. burgdorferi. This result underpins the mixture-genome hypotheses of apicoplast, furthermore, provides a new insight into the origin of this mixture genome. PMID- 15651687 TI - [Progress on cis-acting regulatory elements in nonsense-mediated mRNA decay]. AB - In eukaryotic cells, nonsense-mediated mRNA decay (NMD) is an effective mRNA surveillance mechanism that detects and degrades mRNAs with premature termination codons ( PTC) and protects cells from the potentially deleterious effects of truncated proteins. Some cis-acting regulatory elements have been reported involving in NMD. They are PTC presence, PTC recongnation by downstream elements that termed as downstream sequence element (DSE) in yeast and primarily exon-exon junction ( EEJ) in mammalian cells, stabilizer sequence (STE) inactivation the NMD, and other sequences correlated with NMD such as extended poly (A) in 3' UTRs,upstream open reading frame (uORF) located in 5' UTR and programmed -1 ribosomal frameshift (-1 PRF). Progress on cis-acting regulatory elements in nonsense-mediated mRNA decay was reviewed in this paper. PMID- 15651688 TI - [Hydrodynamic investigations of the current mechanical artificial heart valves]. AB - The work presents results of comparative investigations of the integral hydrodynamic characteristics of the best for today domestic mechanical heart valve (MHV) Medinzh-2 (MI-2) and the turn-disk domestic MHV--LIKS-2, EMIKS (E) and foreign MHV--Medtronic-Hall, Bjork-Shiley (BS), Sorin (SO) and bicuspid valves--St. Jude Medical (SJ), Carbomedics in the stationary and in the pulsatile flows. It was found that according to the main hydrodynamic characteristics including the outlet flow structure, MHV MI-2 was substantially superior to MHV E, BS and SO, and by certain characteristics it approximated to the best foreign MHV SJ. Like all mechanical heart valves, the valve MI-2 disturbs the uniformity of the inlet flow and forms a complex non-stationary flow with detached zones, areas of great shear stresses, with vortex structures of different types, and hence the task of hydrodynamic optimization of its construction is thought to become actual. The comparison of clinical parameters of work of the valve MI-2 with its hydrodynamic characteristics confirms its high hemodynamic effectiveness and thrombo-resistance in patients at long-term periods of follow-up, taking into account the adequate correction of disbalance of hemostasis. PMID- 15651689 TI - [Two approaches to correction of disturbed hemostasis in patients with mechanical artificial heart valves at the long-term follow-up]. AB - The investigation of the hemostasis system performed in 47 patients with mechanical heart valves (MHV) at the terms of (3.50 +/- 0.25) years included 30 men and 17 women. There were 24 cases of aortal MHV and 23 cases of mitral MHV, I II functional class (NYHA). The patients were given oral anticoagulants: phenylin to 35 patients (0.009 mg/day), varfarin to 12 patients (52.6 mg/a week). The primary examination of the patients has revealed insufficient anticoagulation protection and clinically asymptomatic permanent intravascular coagulation of the third degree (PIC-3), with the signs of activation of thrombocytic and/or coagulation link of hemostasis. It has been found that a complex assessment of the hemostasis system state including the vascular-thrombocytic link is an obligatory condition of the right choice and timely correction of the PIC-3 in patients with MHV. The methods of correction of impaired hemostasis depend on the primary involvement of the vascular-thrombocytic and/or coagulation link of hemostasis. The pathogenetically selected antithrombotic therapy for patients with MHV allows timely arrest of PIC-3 and less risk of the development of thrombosis and thrombo-embolic complications at long-term periods of observation. PMID- 15651690 TI - [Biomechanical and flow characteristics of stentless and stented aortic valve bioprostheses]. AB - A comparative investigation of biomechanical and hydrodynamic characteristics of stentless and stented bioprostheses was carried out. It was demonstrated that stentless bioprostheses are functionally superior to stented ones. The differences between biomechanics and hydrodynamics of the examined stentless bioprostheses "BioLab", "AB-Mono-Kemerovo" and DD2 and stented bioprostheses "Wessex", "KemCor" and "Bionox-2" were shown. PMID- 15651691 TI - [The dynamics of morphological alterations in the interatrial septum of the heart in at the occlusion defect of the septum made with the appliance Amplatzer septal occluder]. AB - The experimental work performed in 6 dogs was aimed at studying the dynamics of morphological alterations of the interatrial septum (IAS) tissues in the place of occlusion of the endovascular appliance ("ASO") and around it on the right and left atrium side at different terms after operation. A defect of the animals' interatrial septum in the area of the oval fossa was modeled artificially and then abolished with the "ASO". No substantial differences were revealed between processes of organization of thrombotic masses and formation of the endothelium on the "ASO" from both atria. The endothelium on the micropreparations on the "ASO" surfaces was formed as early as by the 14th day after its implantation in the heart IAS. PMID- 15651692 TI - [Morphological assessment of physical methods of dissection and coagulation in operations on the stomach]. AB - The first results of experimental investigation of the potencies of a new domestic electrosurgical apparatus "ESA-MINI-01" are presented. The investigation of the morphological alterations in the stomach wall after using different physical methods of dissection and coagulation has proved the advantages of the new apparatus which has the principle of work based on an original means of forming the outlet parameters. PMID- 15651693 TI - [Intrathoracic esophagogastroanastomosis: a problem and ways for its solution]. AB - The investigation has shown that the due level of blood supply of the wound surfaces of the organs being sutured, the obligatory absence of a strain, perfect technique of the proper manual putting the end-to-end type anastomosis and the rational placing of the transplant in the posterior mediastinum with a strictly vertical rectilinear orientation, without bends and rotation, allowed to get a smooth course in 98.8% of the operated patients. Incompetent sutures took place in 1.2% of cases. It was also shown that the anastomosis of choice might be an invagination variant of the applying of it having not only high degree of reliability, but also the marked antireflux properties. The dynamic endoscopic control with the correction of forming anastomosis during 2-3 months allowed minimization of possibility of the development of scary stenosis. The results obtained suggest that the problems of intrathoracic esophageal anastomosis in clinic have been solved. PMID- 15651694 TI - [Surgical treatment of the tracheal scarry stenoses]. AB - Surgical policy for scarry tracheal stenosis is analyzed. In the years 1998-2003 in the clinic there were 32 circular resections of the trachea. More than 5 cm of the tracheal segment was ablated in 15 of the cases. The initially performed endoscopic treatment including bougieurage and trachea endoprosthesis was carried out in 19 patients. The immediate and long-term results were satisfactory in all cases that allowed the authors to recommend the one-step circular resection of the trachea as a method of choice for the treatment of lengthy scarry stenoses of the trachea. PMID- 15651695 TI - [The diagnostic and curative strategy for spontaneous pneumothorax]. AB - The article presents the authors' experiences with treatment of 292 patients with spontaneous pneumothorax. Results of the suction drainage (188 patients), thoracoscopy and talc pleurodesis (43 patients), videothoracoscopic surgical treatment and pleurodesis (61 patients) are compared. The proposed algorithm allowed to make recurrences 2.8% rarer, the indications for tracheotomy appeared in 1.7% of the patients. PMID- 15651696 TI - [Direct results of endovideosurgical transabdominal adrenalectomy]. AB - Results of transabdominal adrenalectomy performed in 29 patients with different tumors of the adrenals were estimated. The lower traumaticity of the access due to using a new technology as well as taking prophylactic measures for lower aggressive influence of the pneumoperitoneum allowed performing combined operative interventions in 30% of cases and avoiding serious intra- and postoperative complications. PMID- 15651697 TI - [Prophylactics of postoperative complications in destructive appendicitis and typhlitis]. AB - The method of extraperitonization of the blind gut cupola based on the removable monofilament sutures and drainage of the area was designed, approved in experiments in dogs (n=18) and corpses (n=14) and successfully applied in the clinic (n=25). Unlike the traditional method, the using of this one results in less impaired trophicity, LPO intensification and phospholipase activity of the tissues of regenerating structures that was the grounds of sufficiently rapid reparation process under conditions of inflammation. The proposed complex of surgical measures in patients with destructive forms of acute appendicitis complicated by the inflammatory-necrotic phenomena in the blind gut wall was shown to prevent the development of dangerous postoperative complications, incompetence of the sutures included. PMID- 15651698 TI - [Tumors of the hand bones]. AB - The article is devoted to an actual problem of surgical treatment of hand bone tumors. It presents a classification of hand bone tumors, pathogenesis, clinical course of the most common tumors, methods of surgical treatment. Results of treatment of 108 patients with hand bone tumors are described. PMID- 15651700 TI - [Assessment of the immuological status in patients with nonspecific spinal osteomyelitis]. AB - The authors describe results of the investigation of the immunological status of 54 patients with nonspecific osteomyelitis of the spine (NOS). The immunological status of NOS patients was shown to be substantially different in the acute and chronic phases of the disease. Cases with lower T-lymphocyte activity, with a dysfunction of the phagocytic activity of lymphocytes prevail in the acute phase. Patients with septic forms of NOS have a hyperergic systemic inflammatory response which is accompanied by functional defectiveness of T-lymphocytes and excessive stimulation of the humoral link. Patients with the chronic form of NOS as a rule have hypoergic systemic inflammatory response with the normal quantity of T-cells, non-completed phagocytosis and high activity of B-lymphocytes (CD20, CD25). So, the early immunological diagnosis of NOS allows prognosis of the disease course and a timely decision for the pathogenetical therapy. PMID- 15651699 TI - [The diagnosis and treatment of injuries of the upper respiratory tract in wounds and traumas of the neck]. AB - The authors made an analysis of results of treatment of 339 patients with wounds and closed injuries of the neck. Laryngotracheal injuries were found in 27 (7.9%) patients, in 15 patients the injuries being of combined character. Active surgical strategy including the necessary thorough revision of the penetrating wounds of the neck during operation under general narcosis anesthesia, by instrumental methods of examination. Such strategy allowed timely detection and adequate elimination of injuries of the organs and vessels of the neck and improvement of the results of treatment of such patients. PMID- 15651701 TI - [Compression stenosis of the celiac trunk in children and adolescents]. AB - Operations for compression stenosis of the celiac trunk (CSCT) were made on 97 patients aged 5.5-18 years (56 women and 41 men). Three basic clinical syndromes were observed: pain abdominal, dyspeptic and neurovegetative. The appearance or intensification of abdominal pains observed in 71.3% of the patients were connected with meal, in 59.69% of them it was simultaneous with physical exercise. The diagnosis of OSCT was based mainly on findings of ultrasonic duplex scanning. The indications to decompression of the celiac trunk were: clinical manifestations of CSCT, the stenosis degree at the maximal expiration more than 50%, peak systolic velocity of the blood flow more than 2 m/s and a gradient of arterial pressure more than 15 mm Hg. The decompression of the celiac trunk consisted in the dissection of a median arcuate ligament of the diaphragm, its interior crura and celiac ganglion fibers. Convalescence and recovery was stated in 87 of 97 patients in the nearest and long-term follow-up. PMID- 15651702 TI - [Autodermal-monophylic plasty of postoperative ventral hernias]. AB - In the clinic of general and urgent surgery of Kazan Medical Academy in the period 1998-2002 operations were performed on 160 patients with great and giant postoperative ventral hernias. Four variants of autodermal-monophyl plasty were used: contact suturing of the hernia defect margins with strengthening the line of sutures with a autodermal-monophyl flap, autodermal lacing, skin flap armoured with an monophyl thread, and a skin flap crimped with an monophyl thread. The results of the operative treatment were complicated by lymphorrhea in 8 patients, thromboembolism in 4 patients (with lethal outcome in 2 cases). Recurrences were noted in 11% of the patients. PMID- 15651703 TI - [The influence of liposuction on the blood leptin level]. AB - The level of serum leptin was studied in 20 patients before and after liposuction carried out for excessive subcutaneous fatty tissue. The dynamics of indices depending on the volume of the ablated fatty tissue was analyzed. It was found that in most patients the level of blood serum leptin correlated with the body mass and fatty tissue index. A removal of limited volumes of fatty tissue results in the elevated level of blood serum leptin. The aspiration of great volume of fatty tissue is followed by the primary decrease of the leptin level which further reaches the initial level and even surpasses it. PMID- 15651704 TI - [The influence of hereditary thrombophilic mechanisms on the degree of permanent intravascular coagulation in patients with artificial heart valves]. AB - The genotyping of 40 patients with artificial heart valves (AHV) was performed after prosthesis of the mitral and aotic valves with bicuspid AHV (Medinzh-2 and CarboMedics). The patients took phenylin and varfarin. The patients' genotype was estimated by the thrombophylic genes: factor V Leiden (FVL), prothrombin G20210A, methylene tetrahydrofolate reductase C677T, G/A--455FGB, 4G/5G PAI-1, PI A1/A2 GPIIIa. The genes determining the thrombocytic activity or the vascular wall state substantially influence the third degree of the intensity of the permanent intravascular coagulation (PIC-3) independent of the degree of correction of hemostasis of oral anticoagulants. The addition of anti-aggregants to therapy is the only that can normalize functional activity of thrombocytes in patients with AHV having such defects. The laboratory detection of the genetic defects is of great practical importance for the determination of risk groups of formation of PIC-3 and the strategy of antithrombotic protection of patients with AHV. PMID- 15651706 TI - [A new method of diagnosis of valvular incompetence of the subcutaneous and perforant veins of the lower extremities using the apparatus Telephleboscanner]. AB - The examination of 76 patients with chronic venous insufficiency has shown that the new method allows visualization of subcutaneous and perforant veins at a depth up to 0.5 cm. Special tests can diagnose pathological venous refluxes suggesting valvular incompetence. Telephleboscanning allows marking the perforant veins that makes the operative interventions more radical. PMID- 15651705 TI - [Local neurotoxicity of isobaric 2% solution of lidocaine at epidural administration]. AB - The morphological and histoenzymological changes in the spinal neurons and spinal ganglia of dogs were analyzed after epidural administration of a 2% solution of lidocaine in acute and chronic experiments. In the control group the animals were given epidural infusions of 0.9% saline. The results obtained have shown the absence of pathological structural and metabolic changes in the nervous tissue after lidocaine injection. Certain signs of morpho-functional reorganization were noted in the spinal neurons of animals in both experimental and control groups. The registered changes were found to vary within the physiological fluctuation limits and are rather indicative of the functional reaction of the nervous tissue structures in question to the epidural injections of either saline or lidocaine. PMID- 15651707 TI - [Surgical treatment of perforative gastroduodenal ulcers in elderly and senile patients]. AB - The work is based on experiences with surgical treatment of 206 elderly and senile patients with perforating gastroduodenal ulcers. In 67.5% of cases the perforations took place in duodenal ulcers, in 16%--in the pyloric canal. The gastric body and antral portion ulcers were perforative more rarely--9.8% and 4.8% respectively. Perforations of the cardial and retrobulbar ulcers were found in 1.9% and 1% respectively. At early terms after operation 67 patients (33.5%) died. After suturing the perforative opening 38 out of 98 patients died (38.8%), after dissection and suturing the ulcers died 7 out of 19 patients, after Oppel Polikarpov operation died 7 out of 11, after resection of the stomach died 4 out of 6, after truncal vagotomy with pyloroplasty died 11 out of 72 patients (15.3%). The main cause of lethal outcomes is thought to be complications of the coexistent diseases, totally responsible for 46.2% of deaths. Truncal vagotomy with a dissection of the ulcer and pyloroplasty performed in 60-70 year old patients gave the least indices of lethality and early postoperative complications, so the indications to radical organ-saving operations in patients of this age must be wider. This method of treatment for perforative ulcers in patients of 71-80 years of age should be used with restrictions due to not bad long-term results of suturing the ulcers (good and excellent results took place in 53.8% of cases). In patients older than 80 radical operations are not indicated. In such cases the ulcer should be better dissected and sutured, the posterior wall of the organ being examined for a "mirror" ulcer. PMID- 15651708 TI - [Improvement of curative measures for peritonites of obstetric and gynecological etiology]. AB - The authors describe results of treatment of 117 patients with obstetric and gynecological peritonites. The first group consisted of 43 patients treated by conventional methods. The second group included 74 patients treated by unguent sanitation of the abdominal cavity and prolonged intraarterial catheter (PIAC) against the background of unguent sanitation. The unguent sanitation of the abdominal cavity with water-soluble antibacterial ointments has decreased the number of intra-abdominal and extra-abdominal complications. However in especially severe cases and at high virulence of the abdominal cavity microflora the application of the unguent sanitation alone was not sufficiently effective. So, in 19 patients PIAC was used by the developed scheme. The complex treatment with the unguent sanitation of the abdominal cavity and PIAC allowed to perform the adequate sanitation of the abdominal cavity with lower microbe dissemination of the abdominal cavity up to the critical level on the third day after operation and resulted in less number of intra-abdominal complications from 39.0 to 19.6%, extra-abdominal complications from 15.0 to 7.5% and lethality from 23.2 to 10.8%. PMID- 15651709 TI - [Surgical prophylactics of bronchopleural fistulas after lung resection and pneumonectomies]. PMID- 15651710 TI - [Blood based cardioplegy. The present state of the problem and perspectives of the development]. PMID- 15651711 TI - [Treatment of complications of reconstructive operations in the surgery of atherosclerotic lesions of the lower extermiety arteries]. PMID- 15651712 TI - Cell wall-deficient bacteria as a cause of infections: a review of the clinical significance. AB - Cell wall-deficient bacteria (CWDB) are pleomorphic bacterial forms. These atypical organisms may occur naturally or they can be induced in the laboratory. Their presence has been known about for over a century, but a definite link to clinical disease outcomes has not been demonstrated. A number of case reports and laboratory studies suggest some disease associations, however. Considerable controversy surrounds the true relevance of CWDB to disease; there is a widespread belief that they may represent a response by the walled organism to adverse extracellular conditions like antibiotic pressure. This review looks at studies published between 1934 and 2003, which were identified by Dialog DataStar using the key words 'cell wall deficient bacteria and clinical significance and infections' and by further scanning the reference list at the end of the papers retrieved. We conclude that the evidence for the clinical significance of CWDB in disease is not compelling. PMID- 15651714 TI - Antioxidant and cardioprotective properties of the sulphydryl angiotensin converting enzyme inhibitor zofenopril. AB - Zofenopril, a new potent sulphydryl angiotensin-converting enzyme (ACE) inhibitor, is characterized by high lipophilicity, selective cardiac ACE inhibition, and antioxidant and tissue protective activities. In vitro and in vivo experiments suggest that zofenopril exerts antioxidant properties at clinically achievable tissue concentrations. In endothelial cells, zofenopril enhances nitric oxide production, attenuates atherosclerotic lesion development and inhibits adhesion molecule expression by reducing reactive oxygen species. These peculiar characteristics are reflected in the drug's cardioprotective activity, which has been shown to be greater than that of non-sulphydryl ACE inhibitors. Cardiac hypertrophy was also reduced by chronic zofenopril administration, independently of its blood pressure-reducing effect. ACE inhibitors with a sulphydryl group could have an advantage in improving vascular function and reducing cardiac impairment compared with non-sulphydryl-containing ACE inhibitors. This could explain zofenopril's remarkable clinical efficacy post infarction, and potentially beneficial use in prevention and therapy of cardiovascular diseases, such as atherosclerosis, thrombosis and heart failure. PMID- 15651713 TI - Efficacy of lumiracoxib in osteoarthritis: a review of nine studies. AB - Lumiracoxib is a cyclooxygenase-2 selective inhibitor in development for the treatment of osteoarthritis (OA), rheumatoid arthritis and acute pain. We reviewed nine clinical studies of 1-52 weeks' duration demonstrating the efficacy of lumiracoxib in OA. Male and female patients aged > or = 18 years with primary OA of the hand, hip or knee received lumiracoxib, placebo or active comparators (diclofenac, celecoxib or rofecoxib). Lumiracoxib provided consistent reductions in OA pain intensity and improvements in the patient's global assessment of disease activity and functional status (assessed using the Western Ontario and McMaster Universities Osteoarthritis Index questionnaire or the Australian/Canadian OA Hand Index). These results were superior to placebo and similar to the active comparators tested. In addition, lumiracoxib was consistently superior to placebo and generally similar to active comparators in terms of the new Outcome Measures in Clinical Trials and Osteoarthritis Research Society International criteria. These were used to provide a single measure of response to treatment, taking into account pain, the patient's global assessment of disease activity and functional status. PMID- 15651715 TI - Effects of sevoflurane, isoflurane and propofol infusions on post-operative recovery criteria in geriatric patients. AB - We compared the effects of sevoflurane, isoflurane and propofol infusions on postoperative recovery criteria in geriatric patients. Sixty patients aged > 65 years, classified as American Society of Anesthesiologists (ASA) group 1 or 2 and undergoing gynaecological or urological procedures were randomized equally into three groups. Group 1 received 1 minimum alveolar concentration (MAC) sevoflurane in a 50% O2/N2O mixture and group 2 received 1 MAC isoflurane in a 50% O2/N2O mixture. Group 3 received a 50% O2/N2O mixture plus propofol total intravenous anaesthesia (8 mg/kg for the first 30 min, followed by 6 mg/kg for maintenance). Recovery criteria comprising the times to spontaneous eye opening, extubation, response to verbal stimuli and orientation were recorded following the discontinuation of anaesthesia. Recovery times were significantly shorter in groups 1 and 3 compared with group 2. We conclude that sevoflurane and propofol had similar effects on recovery criteria and were associated with a faster recovery than isoflurane. PMID- 15651716 TI - Clinical features associated with circulating concentration of soluble leptin receptor in patients with diabetes. AB - We aimed to determine if there were any clinical features that were significantly associated with the circulating concentration of soluble leptin receptor (OB-Re) in 67 Japanese subjects with diabetes mellitus. The characteristics evaluated included age, height, body weight, body mass index (BMI), systolic and diastolic blood pressure, duration of diabetes, haemoglobin A1C and blood lipid concentrations, urinary albumin excretion rate, circulating concentrations of leptin, tumour necrosis factor-alpha (TNF-alpha), TNF-alpha receptor 1 and genotypes of the angiotensin-converting enzyme (ACE) gene. We found statistically significant negative correlations between circulating OB-Re concentration and body weight, BMI, diastolic blood pressure, concentrations of leptin and TNF alpha receptor 1. Serum OB-Re concentration was not associated with any of the other clinical characteristics that were measured, or with the different ACE genotypes. Our results suggest that OB-Re might have an important influence on the biological activity of leptin in diabetic subjects. PMID- 15651717 TI - Preparation and evaluation of a hyaluronate-collagen film for preventing post surgical adhesion. AB - Post-surgical adhesion occurs when fibrous strands of scar tissue form, leading to the abnormal joining of anatomical structures. Patients undergoing abdominal surgery are at risk of the complications associated with intraperitoneal adhesions. Hyaluronic acid (HA) is a biocompatible, biodegradable and non-toxic natural polymer, which is gaining popularity as a barrier agent for preventing post-surgical adhesions. As HA is water-soluble and rapidly degraded in vivo, chemical modification is required to produce a non-soluble sheet that might be used to prevent tissue adhesion. We developed a range of biocompatible cross linked HA-collagen composites and then evaluated them in a rat model of post surgical adhesion. The results showed that cross-linked HA-collagen was almost totally resistant to hyaluronidase digestion. HA-collagen membranes induced minimal tissue reactions and were bioresorbed within 14 days post-surgery. These results suggest that cross-linked HA-collagen membrane may be a valuable anti adhesion material to prevent post-surgical intraperitoneal adhesion. PMID- 15651718 TI - An alternative, less invasive approach to median sternotomy for cardiac operations in adults: right infra-axillary minithoracotomy. AB - We compared the use of right infra-axillary minithoracotomy and conventional median sternotomy in direct open-heart surgery in 59 adults undergoing elective surgery for mitral valve stenosis, mitral valve disease, atrial septal defect repair, left atrial myxoma excision or mitral and tricuspid valve disease. Patients were randomized to the infra-axillary minithoracotomy group (Group A; n = 29) or the median sternotomy group (Group B; n = 30). Post-operative outcomes (post-operative bleeding; cross-clamp time; length of hospital and intensive care unit stays; and post-operative blood transfusion and analgesic requirements) were recorded and compared; they were found to be significantly lower in Group A than Group B. We concluded that right infra-axillary minithoracotomy is less invasive and can be used safely in adults as an alternative approach to conventional median sternotomy for some cardiac operations. Further multicentre studies in adults are now needed. PMID- 15651719 TI - Effect of carboxylesterase inhibition on the anti-tumour effects of irinotecan. AB - Irinotecan (CPT-11) is an important anti-cancer agent activated by carboxylesterase (CE). Treatment with CPT-11 may be associated with severe adverse effects, however, so determining the optimal dose would greatly benefit patients. We investigated the relationship between the anti-tumour effects of CPT 11 and CE concentration using bis-p-nitrophenylphosphate (BNPP), a specific inhibitor of CE, in nude mice with xenograft tumours. Initial experiments showed that the optimal dose of CPT-11 was 100 mg/kg. This dose was then used to study the anti-tumour effects of CPT-11 with and without BNPP. A direct correlation was found between the dose of administered BNPP and the growth rate of the tumour, demonstrating that the anti-tumour effects of CPT-11 were related to the CE concentration. Measuring the concentration of CE may allow the optimum dose of CPT-11 to be determined, opening up the possibility of individualized chemotherapy programmes. PMID- 15651720 TI - Therapeutic effect of vitamin D3 in a rat diffuse axonal injury model. AB - We investigated the therapeutic effect of vitamin D3 in a rat diffuse axonal injury model. A total of 60 male Sprague-Dawley rats weighing 175-200 g were anaesthetized and subjected to head trauma using Marmarou's impact-acceleration model. The rats were then separated into two groups; one group was treated with vitamin D3 and the other with saline for up to 4 days after the head trauma. Rats from both groups were killed 1, 3 or 8 days post-injury. The brains were examined histopathologically and scored according to the level of neuronal, vascular and axonal damage. There were no significant differences between the groups after 1 or 3 days, but evaluation after 8 days revealed a significant improvement in the group treated with vitamin D3. Our data indicate that vitamin D3 has a beneficial effect in diffuse axonal injury and may be useful in the management of this condition. PMID- 15651722 TI - Ulcerative colitis activity index: a useful prognostic factor for predicting ulcerative colitis outcome. AB - We evaluated the usefulness of various parameters in predicting the prognosis of ulcerative colitis. The records of 73 patients with ulcerative colitis were examined retrospectively. Patients were divided into two groups according to whether they had received only 5-aminosalicylic acid (5-ASA; n = 26) or glucocorticoids and/or azathioprine with or without 5-ASA (n = 47). The disease extent, endoscopic activity and ulcerative colitis activity index (UCAI) before therapy were recorded, together with the disease outcome. No statistically significant differences in outcome were observed in relation to therapy group, disease extent or endoscopic activity. UCAI had a significant effect on outcome, however: patients with lower UCAI values were more likely to remain in remission and less likely to require urgent surgery or experience a fatal outcome than those with higher UCAI values. This difference was apparent in both treatment groups. Thus a high pre-treatment UCAI may indicate a worse outcome. PMID- 15651721 TI - Incidence of the type and calcification patterns in patients with elongated styloid process. AB - We investigated the incidence of elongated styloid process (Eagle's syndrome) using panoramic radiographs taken of 860 patients referred to our clinic. Any styloid process identified was classified according to its length, type and the pattern of calcification. Fifty-nine elongated styloid processes were identified in 32 patients (3.7%), most being bilateral; 24 patients were female and eight were male (female/male ratio: 3:1). The mean age of these patients was 43 +/- 14 years (range: 18 - 78 years). Type I (elongated) was the most frequent type on both sides (42/59); and the most frequent patterns of calcification were partially calcified on the left side (18/59) and completely calcified on the right side (16/59). Only two patients were symptomatic. A corrected differential diagnosis is important to distinguish elongated styloid process from other pathologies with partially overlapping symptoms. We would recommend that clinicians consider the possibility of Eagle's syndrome when both the clinical and radiographic evidence support this diagnosis. PMID- 15651723 TI - Effect of anti-hypertensive drug dose frequency on the clinic-home blood pressure difference in patients with stage 1 treated hypertension. AB - Clinic blood pressure (CBP) is generally used for diagnosis and treatment monitoring in hypertension, but target organ damage correlates more closely with home blood pressure (HBP). Eliminating the clinic-home blood pressure difference (CHBPD) would make conventional CBP a more accurate alternative to HBP. This prospective, randomized, open trial compared the effect of a once-daily versus a twice-daily regimen of anti-hypertensive therapy on CHBPD. After a 2-week wash out period, 85 confirmed stage 1 hypertensive patients were randomized to receive 2 mg trichlormethiazide daily in one (40 subjects) or two (45 subjects) daily doses for 3 weeks. CBP and HBP measurements were taken during the third week of treatment and the CHBPD calculated. After treatment, the systolic and diastolic CHBPD values were significantly greater in the once-daily regimen than in the twice-daily regimen. Conventional CBP should not be used as an alternative to HBP for evaluating prognosis and monitoring anti-hypertensive therapy when using a once-daily regimen. PMID- 15651724 TI - Enlargement of post-traumatic intracerebral haematoma: incidence and time course. AB - We retrospectively assessed the incidence and time course of enlargement in post traumatic intracerebral haematoma (PTICH). Computed tomography (CT) scans from 165 patients who underwent a scan within 72 h and a repeat scan within 120 h of the onset of trauma were examined. A semi-automated method using region deformation-based segmentation was used to calculate the haematoma volume. The presence of haematoma enlargement was also determined based on a consensus by five observers. Seventy cases (42%) showed enlargement of the haematoma. The frequency of haematoma enlargement decreased as the interval between the onset of trauma and the initial scan increased. The discriminant value of the ratio of the haematoma volume in the second scan to that in the initial scan was ascertained, and the cut-off value for haematoma enlargement was determined to be 1.45. The radiographic criterion for enlargement in PTICH on CT scan was, therefore, defined as a > or = 1.45 times increase in haematoma volume. PMID- 15651725 TI - Fresh homografts obtained through a national organ-sharing programme for repair of congenital heart disease. AB - Homografts have been used in congenital cardiac surgery for over 30 years. We utilized the resources of a national organ-sharing programme to obtain fresh homografts and report their use in correcting cardiac pathologies in 20 children between March 2001 and May 2003. In 16 patients, a valved conduit was used to form a connection between the pulmonary ventricle and the pulmonary artery. In three patients, a non-valved aortic conduit was used to form an extra-cardiac Fontan circulation and in one patient, non-valved pulmonary and aortic conduits were used to repair an infected aortic aneurysm. Three patients died following surgery. Survivors were followed up using echocardiography between 2 and 24 months post-surgery. Results demonstrate that, with the help of a national organ sharing programme, the use of fresh homograft conduits is feasible in a paediatric patient population with reasonable waiting times. PMID- 15651726 TI - Equine ergonomics: a new era? PMID- 15651727 TI - The influence of head and neck position on kinematics of the back in riding horses at the walk and trot. AB - REASONS FOR PERFORMING STUDY: A common opinion among riders and in the literature is that the positioning of the head and neck influences the back of the horse, but this has not yet been measured objectively. OBJECTIVES: To evaluate the effect of head and neck position on the kinematics of the back in riding horses. METHODS: Eight Warmblood riding horses in regular work were studied on a treadmill at walk and trot with the head and neck in 3 different predetermined positions achieved by side reins attached to the bit and to an anticast roller. The 3-dimensional movement of the thoracolumbar spine was measured from the position of skin-fixed markers recorded by infrared videocameras. RESULTS: Head and neck position influenced the movements of the back, especially at the walk. When the head was fixed in a high position at the walk, the flexion-extension movement and lateral bending of the lumbar back, as well as the axial rotation, were significantly reduced when compared to movements with the head free or in a low position. At walk, head and neck position also significantly influenced stride length, which was shortest with the head in a high position. At trot, the stride length was independent of head position. CONCLUSIONS: Restricting and restraining the position and movement of the head and neck alters the movement of the back and stride characteristics. With the head and neck in a high position stride length and flexion and extension of the caudal back were significantly reduced. POTENTIAL RELEVANCE: Use of side reins in training and rehabilitation programmes should be used with an understanding of the possible effects on the horse's back. PMID- 15651728 TI - Three-dimensional kinematics of the equine distal forelimb: effects of a sharp turn at the walk. AB - REASONS FOR PERFORMING STUDY: Sharp turns are suspected to increase expression of several distal forelimb lamenesses even at the walk but the biomechanical consequences of such a movement remain unknown. OBJECTIVE: To quantify the effects of a sharp turn at the walk on the 3-dimensional movements of the distal segments of the forelimb. METHODS: Kinematics of the distal segments were measured in 4 healthy horses invasively with an ultrasonic system. Three dimensional rotations of the digital joints were calculated by use of a joint coordinate system. Data obtained for a turn at the walk were compared to those obtained in a straight line. RESULTS: During the stance phase in a turn, the inside forelimb underwent an adduction that induced lateromotion and medial rotation in the distal interphalangeal joint and medial rotation in the proximal interphalangeal joint. These movements were maximal at heel-off and decreased during breakover as the hoof underwent a sudden lateral rotation. CONCLUSIONS: Walking in a sharp turn affects the kinematics of the digital joints outside the sagittal plane. POTENTIAL RELEVANCE: This knowledge offers the opportunity to derive hypotheses on biomechanical factors that could contribute to the pathogenesis of digital injuries and on consequences for rational shoeing. PMID- 15651729 TI - Relationship between synovial fluid levels of glycosaminoglycans, hydroxyproline and general MMP activity and the presence and severity of articular cartilage change on the proximal articular surface of P1. AB - REASONS FOR PERFORMING STUDY: Osteoarthritis (OA) is one of the most prevalent and disabling chronic conditions affecting horses and leads to degeneration of articular cartilage. Diagnosis is based on clinical signs in combination with radiography, which is relatively insensitive and provides only an indication of accumulated damage. Alternative methods, such as molecular markers, are therefore needed that can quantitatively, reliably and sensitively detect osteoarthritic changes in the joints at an early stage of the disease. If such markers are to be used reliably, it is important to know the relationship between marker concentration and cartilage composition. OBJECTIVES: To study the relationship between cartilage composition, synovial fluid levels of glycosaminoglycans (GAGs), hydroxyproline (Hyp) and general matrix metalloproteinase (MMP) activity, and the presence and severity of articular cartilage damage on the articular surface of P1. METHODS: Synovial fluid (SF) was collected from the metacarpophalangeal joints of 60 mature horses, and levels of GAGs, Hyp and general MMP activity were determined. Further, GAG and denatured collagen content of the articular cartilage were determined at the dorsal articular margin of P1 (site 1) and central cavity (site 2). The presence and severity of cartilage change was quantified using the cartilage degeneration index (CDI), measured at the same 2 sites. Correlations between SF parameters, cartilage composition and degree of cartilage degeneration were sought using correlation analysis. RESULTS: There was no correlation between GAG or Hyp content of SF and the amount of GAGs or denatured collagen, respectively, in cartilage. In joints with moderate to severe cartilage damage, the GAG content of site 1 was significantly lower than in joints with no to minimal cartilage change (P = 0.005) and there was a negative correlation between the amount of denatured collagen and GAG content at site 1 in all joints (r = -039, P = 0.002). Further, in joints with moderate to severe cartilage damage, there was a significant positive correlation between MMP activity in SF and Hyp levels in SF (r = 0.72, P < 0.001) and CDI at sites 1 (r = 0.46, P = 0.03) and 2 (r = 0.43, P = 0.04). CONCLUSIONS: General MMP activity in joints with moderate to severe cartilage damage is related to the severity of those cartilage changes and to Hyp levels in SF. Glycosaminoglycan levels in SF are not directly related to MMP activity, GAG content of articular cartilage or severity of cartilage change. POTENTIAL RELEVANCE: Glycosaminoglycan levels in SF are not helpful for the early detection of cartilage lesions. In damaged joints, Hyp levels may give an indication of the severity of cartilage change as they are strongly related to MMP activity, but do not qualify as markers for the presence or absence of cartilage lesions. PMID- 15651730 TI - Ultrastructural immunolocalisation of bone sialoprotein in the osteocartilagenous interface of the equine third carpal bone. AB - REASONS FOR PERFORMING STUDY: One of the most common causes of lameness in racehorses is osteoarthritis (OA). Pathogenesis is not clear and pathological processes of the different joint tissues interact in often progressive events. The interface between cartilage and newly synthesised bone has been shown to be particularly enriched in bone sialoprotein (BSP), a cell-binding matrix protein. OBJECTIVES: To establish whether changes in the concentration of BSP may serve as a marker for early biochemical changes of the subchondral bone. METHODS: Articular cartilage, cartilage/bone interface and subchondral bone of the proximal third carpal bone from 3 Standardbred trotters were analysed ultrastructurally for the presence of BSP in normal and degenerative areas. RESULTS: A marked increase of BSP in the cartilage/bone interface with degenerative changes of the bone and cartilage compared to the morphologically intact cartilage/bone interface was noted, but levels of the protein were distinctly lower in the distal bone. CONCLUSIONS: The results indicate that BSP has the potential to be used as a marker for changes in bone metabolism in the subchondral bone. POTENTIAL RELEVANCE: Tools to monitor early biochemical changes within the connective tissues of the joint in vivo are essential in studies of the pathogenesis of OA. These could be used to monitor and understand such changes in relation to load, exercise, training programmes, inflammation and the development of OA. PMID- 15651731 TI - Analysis of cartilage oligomeric matrix protein (COMP) degradation and synthesis in equine joint disease. AB - REASONS FOR PERFORMING STUDY: Cartilage oligomeric matrix protein (COMP) is abundant within cartilage; its turnover and/or degradation have been investigated in various equine joint diseases and it has been suggested that COMP fragmentation might be useful for monitoring such conditions. OBJECTIVES: To determine whether COMP metabolism is compromised in equine osteoarthritis (OA) and whether COMP degradation is a useful joint marker representing cartilage destruction. HYPOTHESIS: A monoclonal antibody (mAb) with a higher affinity for degraded COMP allows discrimination of diseased joints by quantifying COMP levels and fragmentation. METHODS: A mAb (clone14G4) was generated against equine cartilage COMP. The NH2-terminal sequence of enzyme-cut COMP fragments recognised by 14G4 was determined, as was the efficiency of binding to COMP (using a generated COMP peptide). COMP concentration and fragmentation were analysed in synovial fluid (SF) from normal horses and those with OA. RESULTS: The mAb 14G4 had a higher affinity for the smaller fragments of equine COMP, compared with a mAb (clone 12C4) generated against human COMP. The 14G4 epitope was identified as between C134 and F147. The COMP values in OA (mean +/- s.d. 205.8 +/- 90.9 microg/ml) were significantly higher than in the normal (133.1 +/- 31.5 microg/ml) SF. On the immunoblots of OA sample, the proportions of intact COMP were significantly lower, while smaller fragments ranging from 75 to 290 kDa were higher compared with the normal SF. CONCLUSIONS AND POTENTIAL RELEVANCE: The mAb 14G4 reliably detects COMP degradation as well as synthesis, and fragmentation analysis combined with quantification in SF could be useful to study equine OA. PMID- 15651732 TI - Age-related changes in serum insulin-like growth factor-I, insulin-like growth factor-I binding protein-3 and articular cartilage structure in Thoroughbred horses. AB - REASONS FOR PERFORMING STUDY: Structural changes in articular cartilage associated with the ageing process require definition for investigators performing developmental and age-related studies, for which information is lacking. OBJECTIVES: To 1) determine the onset and end of puberty as defined by serum insulin like growth factor (IGF-I) and IGF-binding protein-3 (IGFBP-3) concentrations and 2) correlate articular-epiphyseal cartilage complex structural changes with the onset and end of puberty. METHODS: IGF-I and IGFBP-3 were measured in serum samples from normal female and male horses age 9-715 days to determine peak and steady-state values for horses transitioning through puberty. Osteochondral tissue sections were obtained from horses age 120-840 days (4-28 months) and examined histologically for cartilage canals and tidemark formation. RESULTS: In male and female horses, serum IGF-I/IGFBP-3 concentrations peaked at approximately 225 days, defining the onset of puberty. Cartilage canals were absent from articular cartilage just prior to this time point. IGF-I/IGFBP-3 concentrations declined to steady-state levels at approximately age 450 days, signalling exit from puberty and therefore the beginning of ageing. This time point correlated to initial formation of a tidemark in the osteochondral tissue sections. CONCLUSIONS: Horses may be considered pubescent at age 225-450 days, and post pubescent and ageing after age 450 days. POTENTIAL RELEVANCE: Defining the normal post natal to post pubescent concentrations for serum IGF-I and serum IGFBP-3 establishes subsets of animals for age-related studies and may be used to monitor horses for abnormally high IGF-I concentrations due to natural disease or subsequent to systemic growth hormone administration. PMID- 15651733 TI - Freezability of equine semen after glass beads column separation. AB - REASONS FOR PERFORMING STUDY: The success rate of artificial insemination following the freezing of stallion semen is limited; therefore, improving the stallion semen quality after the freezing and thawing process is a necessary objective. OBJECTIVES: To investigate the influence of glass bead column separation on the freezability of stallion semen. HYPOTHESIS: Glass beads in a column separator remove damaged and dead spermatozoa in the ejaculate during centrifugation. METHODS: In total, 50 ejaculates from 6 Lipizzaner stallions were studied. Each ejaculate was divided into 2 parts, one half processed following standard procedure and the second half used for the column separation procedure. After freezing, semen quality was evaluated using standard tests for motility, morphology and viability of semen. RESULTS: Motility and progressive motility of the column-separated (CS) semen were significantly higher (P < 0.001) before freezing and immediately, 24 and 48 h after thawing. A significant increase (P < 0.001) in the percentage of hypoosmotic positive spermatozoa was observed in CS samples. The percentage of total morphological changes in the separated samples before and after freezing was significantly lower (P < 0.001) compared with samples prepared using the standard procedure. A substantial decrease (P < 0.001) was found in the percentage of spermatozoa with damaged acrosomes. However, the percentage of spermatozoa with coiled tails was increased in the separated samples (P < 0.001). CONCLUSIONS: Column separation before freezing has a positive effect on the quality of thawed equine semen. POTENTIAL RELEVANCE: The quality of CS frozen/thawed samples indicates their potential use for increasing insemination success in mares. PMID- 15651734 TI - Immunoglobulin and peripheral B-lymphocyte concentrations in Fell pony foal syndrome. AB - REASONS FOR PERFORMING STUDY: Fell pony foals are affected by a congenital fatal disease that leads to profound anaemia and immunodeficiency. Previous studies comparing healthy and affected foals have shown normal T-cell populations, but a severe B-lymphopenia. OBJECTIVES: To measure the levels of individual immunoglobulin subisotypes in normal and affected Fell ponies and correlate these levels with the number of peripheral B-lymphocytes. METHODS: Serum levels of individual immunoglobulin subisotypes were measured by ELISA and correlated with the number of peripheral B-lymphocytes (measured by flow cytometry). RESULTS: Affected foals had significantly reduced serum levels of IgM, IgGa, IgGb and IgG(T) that coincided with the normal reduction in maternally derived immunoglobulin in foals and, with the exception of IgGb, correlated strongly with the B-lymphopenia. CONCLUSIONS: These data suggest that affected foals are unable to produce their own immunoglobulins. Therefore, once maternal immunity has waned, it may be the lack of specific foal-derived immunoglobulin that gives rise to the clinical signs of immunodeficiency. POTENTIAL RELEVANCE: Measurement of IgM after age 4 weeks may provide a more accessible means of confirming the status of future affected Fell pony foals than the measurement of B-lymphocytes (a technique limited to a few specialist laboratories). PMID- 15651735 TI - Arterial lactate concentration, hospital survival, sepsis and SIRS in critically ill neonatal foals. AB - REASONS FOR PERFORMING STUDY: Blood lactate concentration has been shown to be a useful clinical indicator in human patients, but has not been formally investigated in critically ill foals. OBJECTIVE: To investigate the association of blood lactate with hospital survival, markers of cardiovascular status, metabolic acid base status, sepsis and systemic inflammatory response syndrome (SIRS). METHODS: A database containing clinical, haematological, plasma biochemical and hospital outcome data on neonatal foals referred to an intensive care unit in 2000-2001 was analysed. Seventy-two foals for which arterial lactate was measured at admission were included in the study. RESULTS: Sixty-one foals had an admission lactate concentration > 2.5 mmol/l. Admission lactate was statistically associated with hospital survival, mean arterial pressure, blood creatinine concentration, bacteraemia, anion gap, lactate concentration at 18-36 h after admission and evidence of SIRS, but not with packed cell volume or heart rate. Lactate at 18-36 h was also associated with survival and evidence of SIRS. Anion gap, base excess, base excess due to unidentified anions (BEua), simplified strong ion gap or bicarbonate correctly classified foals for presence of hyperlactaemia (> 5 mmol/l) in < or = 80% of animals. CONCLUSIONS: Admission blood lactate gives important prognostic information. Lactate should be measured rather than assumed from the anion gap, base excess, BEua, simplified strong ion gap or bicarbonate. POTENTIAL RELEVANCE: Blood lactate concentrations at admission are clinically relevant in neonatal foals and warrant further investigation. This should include the clinical value of measuring changes in lactate in response to treatment. PMID- 15651736 TI - Cardiopulmonary effects and pharmacokinetics of i.v. dexmedetomidine in ponies. AB - REASONS FOR PERFORMING STUDY: Currently available sedatives depress cardiopulmonary function considerably; therefore, it is important to search for new, less depressive sedatives. The study was performed to assess duration and intensity of cardiopulmonary side effects of a new sedative, dexmedetomidine (DEX), in horses. OBJECTIVES: To study pharmacokinetics and cardiopulmonary effects of i.v. DEX. METHODS: Pharmacokinetics of 3.5 microg/kg bwt i.v. DEX were studied in a group of 8 mature (mean age 4.4 years) and 6 old ponies (mean age 20 years). Cardiopulmonary data were recorded in mature ponies before and 5, 10, 20, 30, 45 and 60 mins after administration of DEX 3.5 microg/kg bwt i.v. Data were analysed using ANOVA for repeated measures, and where appropriate Dunnett's t test was used to detect differences from resting values (P < 0.05). RESULTS: Within 2 h after DEX administration, plasma levels were beyond limits of quantification (0.05 ng/ml). Mean values for kinetic parameters for mature and old ponies were: Cmax (ng/ml) 4.6 and 3.8, t 1/2 (min) 19.8 and 28.9 and AUC (ng.min/ml) 34.5 and 44.3, respectively. Heart rate, central venous pressure, pulmonary artery pressure and pulmonary capillary wedge pressure did not change significantly compared to presedation values throughout the 60 min observation period. Compared to presedation values, stroke volume and mixed venous PO2 were reduced for the first 5 mins, paralleled by an increase in systemic and pulmonary vascular resistance. Cardiac index was reduced for the first 10 mins, arterial blood pressures at 20, 30 and 45 mins and respiratory rate throughout the 60 min observation period, but no change in arterial PO2 or PCO2 occurred. CONCLUSIONS: DEX administration i.v. causes similar cardiopulmonary changes to those caused by other alpha-2 adrenoceptor agonists, but of very short duration. DEX is redistributed particularly rapidly. POTENTIAL RELEVANCE: DEX might be safer for sedation of horses because of its very short-lasting cardiopulmonary side effects. For long duration sedation, its kinetics favour its use as a continuous infusion. PMID- 15651737 TI - The effect of bilateral glossopharyngeal nerve anaesthesia on swallowing in horses. AB - REASONS FOR PERFORMING STUDY: Dysfunction of the glossopharyngeal nerve has been implicated as a cause of dysphagia in horses. However, recent studies have indicated that this is not the case. OBJECTIVES: To determine whether bilateral glossopharyngeal nerve anaesthesia would cause dysphagia in horses or result in measurable alterations in the timing, function, or sequence of swallowing. METHODS: Swallowing was evaluated in 6 normal horses with and without bilateral glossopharyngeal nerve anaesthesia. Swallowing dynamics were assessed subjectively and objectively based on time from prehension of food until swallowing, number of tongue movements until initiation of swallowing, depth of bolus at the base of the tongue prior to initiation of swallow and evidence of tracheal aspiration using fluoroscopy and endoscopy. RESULTS: There was no evidence of aspiration or dysphagia in horses before or after bilateral glossopharyngeal nerve block. No observed or measured differences in swallowing sequence or function could be detected in blocked compared to unblocked horses. However, there was a trend in blocked horses for the number of tongue pushes and the time to swallowing to be increased. CONCLUSIONS: Glossopharyngeal nerve function may not be essential for normal swallowing function in otherwise healthy horses. POTENTIAL RELEVANCE: Clinically, normal swallowing is not an appropriate test of glossopharyngeal nerve function and dysphagic horses should not be assumed to have glossopharyngeal nerve dysfunction. PMID- 15651738 TI - Colorectal distention in the horse: visceral sensitivity, rectal compliance and effect of i.v. xylazine or intrarectal lidocaine. AB - REASONS FOR PERFORMING STUDY: Most current models of visceral sensitivity testing in the horse have required visceral cannulation. Colorectal distention (CRD) is a widely used, noninvasive method for testing in other species and could be adapted for use in horses. OBJECTIVES: To develop a protocol of controlled CRD in the conscious horse and to evaluate the effect of i.v. xylazine or intrarectal lidocaine on CRD threshold and rectal compliance. METHODS: Eight horses were used for baseline studies (3 trials each) and 6 horses to evaluate treatments (4 trials, 2 per treatment). A 45 cm diameter polyvinyl balloon attached to plastic tubing was used for rectal distention following a stepwise barostat-controlled inflation pattern. RESULTS: The procedure was well tolerated by all horses. Mean baseline threshold pressure was 14.17 mmHg. Xylazine i.v. resulted in significantly (P < 0.05) higher mean threshold pressures compared to baseline or rectal lidocaine. Rectal compliance increased following lidocaine treatment relative to baseline or xylazine. CONCLUSIONS: CRD offers a noninvasive method for visceral sensitivity testing in the horse. Xylazine raises CRD threshold, while lidocaine increases rectal compliance. POTENTIAL RELEVANCE: The increased rectal compliance following intrarectal lidocaine administration may explain the benefit of its use to facilitate rectal examination. PMID- 15651739 TI - Effects of cyclooxygenase inhibitors flunixin and deracoxib on permeability of ischaemic-injured equine jejunum. AB - REASONS FOR PERFORMING STUDY: Recent studies have shown that flunixin prevented recovery of equine jejunum post ischaemia. However, the use of a purported cyclooxygenase (COX)-2 preferential inhibitor, etodolac, also prevented recovery. These findings may have implications for the use of nonsteroidal anti inflammatory drugs in colic patients. OBJECTIVE: To compare the effects of deracoxib, a highly selective canine COX-2 inhibitor, with flunixin on in vitro recovery of ischaemic-injured equine jejunum. METHODS: Six horses underwent 2 h jejunal ischaemia, after which mucosa was mounted in Ussing chambers and recovered for 240 mins. Transepithelial electrical resistance (TER) and mucosal to-serosal fluxes of 3H-mannitol were monitored as indices of barrier function in the presence of flunixin or deracoxib. RESULTS: The TER of ischaemic-injured tissue recovered significantly over 240 mins in the presence of no treatment, but not in the presence of flunixin or deracoxib. In addition, flunixin-treated ischaemic jejunum was significantly more permeable to mannitol when compared with untreated tissue by the end of the recovery period, whereas deracoxib treatment did not increase permeability. Addition of the PGE1 analogue misoprostol to flunixin-treated tissue restored recovery of TER. CONCLUSIONS AND POTENTIAL RELEVANCE: Treatment of horses with ischaemic jejunal disease with flunixin may result in a prolonged permeability defect in recovering mucosa. Addition of misoprostol or replacement of flunixin with deracoxib may ameliorate effects of COX inhibitors on recovering mucosa. PMID- 15651740 TI - Comparison of intra-articular drilling and diode laser treatment for arthrodesis of the distal tarsal joints in normal horses. AB - REASONS FOR PERFORMING STUDY: Anecdotal reports suggest that laser-facilitated arthrodesis of the distal tarsal joints improves the prognosis compared with intra-articular drilling but no objective comparisons have been performed. OBJECTIVES: To evaluate intra-articular drilling and laser-facilitated arthrodesis using in situ and in vivo techniques. METHODS: Fourteen cadaver limbs were evaluated in situ for chondrocyte viability after both surgical techniques. In vivo, one randomly selected limb was subjected to laser-facilitated arthrodesis and the other underwent intra-articular drilling in 6 normal horses. Clinical examinations were performed at 1, 3 and 5 months. Two horses were subjected to euthanasia at 1, 3 and 5 months. RESULTS: Significantly more chondrocyte death was observed with laser-facilitated arthrodesis compared to drilling, but the overall degree suggested only a focal effect. In vivo, both groups demonstrated minimal post operative morbidity. There was more evidence of arthrodesis seen at all time points with intra-articular drilling. CONCLUSIONS: This study demonstrated that changes associated with ultimate arthrodesis occur earlier after intra-articular drilling of the distal tarsal joints than laser facilitated arthrodesis, although clinically affected horses may respond differently. POTENTIAL RELEVANCE: Intra-articular drilling may provide earlier arthrodesis of the distal tarsal joints, but not necessarily a better long-term result. PMID- 15651741 TI - Biochemical markers of bone metabolism and risk of dorsal metacarpal disease in 2 year-old Thoroughbreds. AB - REASONS FOR PERFORMING STUDY: Dorsal metacarpal disease (DMD) is a common problem in 2-year-old racehorses and results in loss of a significant number of days from training. Biochemical markers of bone cell activity measured early in the training season could have value for identifying 2-year-old Thoroughbred racehorses that develop DMD. OBJECTIVES: To determine the association between serum concentrations of osteocalcin, the carboxyterminal propeptide of type I collagen (PICP) and the carboxyterminal cross-linked telopeptide of type I collagen (ICTP) measured early in the training season and the risk of DMD. METHODS: Blood samples were collected from 165 two-year-old Thoroughbreds during late November/early December. Osteocalcin and PICP were measured as markers of bone formation, and ICTP as a marker of bone resorption. Training and veterinary records for each horse were monitored over the following training/racing season (10 months). Cases were defined as an episode where signs of DMD were sufficiently severe for a horse to miss at least 5 consecutive days of training. Classification tree and logistic regression analysis were used to identify the most important factors suitable for prediction of DMD risk. RESULTS: There were 24 cases of DMD during the season (14.6% cumulative incidence), with an average time to recognition of approximately 6 months (May). The earliest recognised case was in February and the latest in September. Osteocalcin and ICTP concentrations in the early stages of the training season were significantly higher in horses that subsequently developed DMD (P = 0.017 and 0.019, respectively). DMD cases were also significantly older compared to noncases (21.04 vs. 20.44 months, P = 0.023). Using a multivariable logistic regression model, it was possible to postulate a set of diagnostic rules to predict the likelihood of DMD injury during the season. This suggested that horses with ICTP concentrations above 12365 ug/l and older than 20.5 months are 2.6 times more likely to develop DMD. CONCLUSIONS: The measurement of the bone resorption marker ICTP could be useful for identification of 2-year-olds at increased risk of developing DMD. POTENTIAL RELEVANCE: These findings, together with other strategies such as modification of training regimens, e.g. early introduction of short distances of high-speed exercise into the training programme, could help reduce the days lost to training as a result of DMD. PMID- 15651742 TI - Faecal bile loss in horses following small intestinal resection. PMID- 15651744 TI - Complications associated with NRTI therapy: update on clinical features and possible pathogenic mechanisms. AB - The availability of durable, effective antiretroviral therapy for HIV-infected patients has fundamentally altered the prognosis of this disease and has also increased awareness that long-term drug toxicities have the potential to cause significant morbidity and even mortality in this patient population. The long term use of nucleoside analogue reverse transcriptase inhibitor (NRTI) drugs has been associated with a number of clinically relevant toxicities including hyperlactataemia and lactic acidosis, neuropathy, pancreatitis and, more recently, a syndrome of pathological loss of subcutaneous fat tissue (lipoatrophy). Importantly, the toxicity profile of each NRTI drug within this class is unique in terms of the overall risk of long-term complications, as well as the tissue specificity of its toxic effects. In this review, the clinical manifestations, risk factors and pathological basis for NRTI-associated toxicity syndromes are explored, with an emphasis on clinical assessment and management. PMID- 15651743 TI - Updated European recommendations for the clinical use of HIV drug resistance testing. AB - In most European countries, HIV drug resistance testing has become a routine clinical tool. However, its practical implementation in a clinical context is demanding. The European HIV Drug Resistance Panel was established to make recommendations to clinicians and virologists on this topic and to propose quality control measures. The panel recommends resistance testing for the following indications: i) drug-naive patients with acute or recent infection; ii) therapy failure, including suboptimal treatment response, when treatment change is considered; iii) pregnant HIV-1-infected women and paediatric patients with detectable viral load when treatment initiation or change is considered; and iv) genotype source patient when post-exposure prophylaxis is considered. In addition, for drug-naive patients with chronic infection in whom treatment is to be started, the panel suggests that resistance testing should be strongly considered and recommends testing the earliest sample for drug resistance if suspicion of resistance is high or prevalence of resistance in this population exceeds 10%. The panel does not favour genotyping over phenotype, however it is anticipated that genotyping will be used more often because of its greater accessibility, lower cost and faster turnaround time. For the interpretation of resistance data, clinically validated systems should be used to the greatest extent possible. It is mandatory that laboratories performing HIV resistance tests take regular part in quality assurance programs. Similarly, it is necessary that HIV clinicians and virologists take part in continuous education and meet regularly to discuss problematic clinical cases. Indeed, resistance test results should be used in the context of all other clinically relevant information for predicting therapy response. The panel also encourages the timely collection of epidemiological information to estimate the impact of transmission of resistant HIV and the prevalence of HIV-1 non-B subtypes in the different European countries. PMID- 15651745 TI - The problems faced when assessing the prevalence and incidence of antiretroviral related toxicities. AB - Although the dramatic effect of highly active antiretroviral therapy (HAART) in reducing morbidity and mortality must not be underestimated, it is also important to consider the incidence and prevalence of HAART-related toxicities. Although several studies have investigated HAART-related toxicities, there has been great variety between them in the reported incidence and prevalence rates of these toxicities. Various factors, including whether the study type was a clinical trial or an observational study, the definition of the toxicity endpoints, the demographic characteristics of the study populations and the effect of calendar year on analyses, may all influence the rates observed. We investigated the possible explanations for the differences in the incidence and prevalence rates of HAART-related toxicities between studies, focussing on metabolic and hepatotoxic disorders. PMID- 15651746 TI - Lipodystrophy in patients with HIV-1 infection: effect of stopping protease inhibitors on TNF-alpha and TNF-receptor levels, and on metabolic parameters. AB - OBJECTIVE: To evaluate the effects of stopping treatment with protease inhibitors (PIs) on tumour necrosis factor (TNF)-alpha and TNF-receptor levels, and on the metabolic and morphological abnormalities seen in patients with lipodystrophy. DESIGN: Longitudinal study. METHODS: Ten HIV-positive patients with lipodystrophy (LD) were studied whilst on PIs (LD1) and 3 months after stopping PIs (LD2) together with 10 HIV-positive subjects on PIs without LD (controls). TNF-alpha and TNF-receptor levels, insulin resistance parameters, lipid and hormonal profiles, body composition and fat distribution were measured in all subjects. RESULTS: TNF-alpha, TNF-receptor I (-RI) and TNF-RII levels were significantly lower in controls (P=0.02) than in subjects with LD, and there was a significant decrease in TNF-RI and TNF-RII levels (P=0.01 and 0.03, respectively) on stopping PIs. Insulin levels and the homeostasis model assessment for insulin resistance (HOMA-IR) index were significantly higher in LD1 subjects (P=0.02) than in controls but did not alter when PIs were stopped. Bioelectrical impedance analysis showed a significant decrease on stopping PIs but CT scans showed no significant difference in fat distribution. Apart from high-density lipoprotein, there was no change in lipid parameters on stopping PIs. There was no difference in the level of testosterone, sex hormone binding globulin and cortisol between the three groups. CONCLUSION: Our results show that TNF-alpha activity in patients with LD is modulated by PIs. This was not accompanied by significant changes in body habitus or insulin resistance, although this may have been a consequence of the short follow-up in this study. PMID- 15651747 TI - Alternation of antiretroviral drug regimens for HIV infection. Efficacy, safety and tolerability at week 96 of the Swatch Study. AB - INTRODUCTION: Alternation of antiretroviral drug regimens has been proposed as a novel treatment strategy for HIV infection. However, some concerns persist regarding antiviral efficacy, adherence, toxicity and resistance evolution in the long term. METHODS: A total of 161 antiretroviral-naive HIV-1-infected patients were randomized to receive stavudine/didanosine/efavirenz (group A) or zidovudine/lamivudine/ nelfinavir (group B) or to alternate between the two regimens every 3 months starting with regimen A (group C). Antiviral efficacy, adherence, safety and tolerability were analysed every 12 weeks. RESULTS: After 96 weeks, time to virological failure was significantly delayed in the alternating regimen compared with the standards of care regimens. Virological suppression was seen in 46%, 48% and 58% of patients in groups A, B and C, respectively, in the intention-to-treat analysis and in 75%, 76% and 97% in the on-treatment analysis (A vs C: P=0.014; B vs C: P=0.016; A vs B: P=0.849). At the end of the study, 94% of patients in group A and 92% in groups B and C reported an adherence greater than 95%. Alternating therapy was associated with a similar impact on CD4+ counts in comparison with the standards of care regimens, as well as a lower mitochondrial DNA/nuclear DNA (mtDNA/nDNA) ratio decrease in the mitochondrial substudy performed on 37 patients. The frequency and intensity of adverse events in the alternating group decreased during subsequent cycles. DISCUSSION: Our results favour the hypothesis that proactive therapy switching may delay the accumulation of resistance mutations. Moreover, the alternating regimen was well tolerated and adherence remained comparably high in all treatment groups. The lower mtDNA/nDNA ratio decrease observed in this group may imply a lower impact on mitochondrial toxicity than in standard regimens. PMID- 15651748 TI - Lack of interferon sensitivity-determining region in the genome of hepatitis B virus genotype Ba. AB - BACKGROUND/AIMS: In chronic hepatitis B, both host and viral factors may predict the response to interferon (IFN) treatment. Whether IFN sensitivity-determining regions exist within the hepatitis B virus (HBV) genomic background remains largely unknown. We therefore performed full-length viral genomic comparison between HBVs obtained from IFN responders and non-responders. METHODS: We enrolled 18 HBV genotype Ba patients who had received 24-week IFN 5 MU three times weekly and were followed monthly for 12 months post-treatment. There were 10 responders and eight non-responders. Pretreatment full-length viral nucleotide consensus sequence was obtained. In six non-responders and four responders, post treatment viral nucleotide sequence was further compared with their corresponding pre-treatment specimens. In addition, the average number of nucleotide substitutions of the HBV quasispecies was compared between three responders and three non-responders. RESULTS: HBV nucleotide consensus sequence was identical between responders and non-responders. We found 0-15 (mean 7.7) nucleotide substitutions in the post-treatment HBV genome in the six non-responders and 0-14 (mean 3.8) nucleotide substitutions in the four responders, respectively. Genetic complexity of HBV quasispecies was comparable between responders and non responders. CONCLUSIONS: Our results suggest that an IFN sensitivity-determining region might not exist within the genome of HBV genotype Ba. Host factors and virus-host interactions may be more important in determining the response to IFN treatment. PMID- 15651749 TI - Depressive symptoms after initiation of interferon therapy in human immunodeficiency virus-infected patients with chronic hepatitis C. AB - BACKGROUND: Interferon alpha (IFN-alpha) therapy for chronic hepatitis C (CHC) infection is commonly associated with neuropsychiatric side effects including depressive symptomatology. In this study, we evaluated the incidence and management of depressive symptoms during IFN-alpha therapy in HIV-infected patients with CHC. METHODS: HIV-infected patients with CHC who began IFN-alpha and ribavirin therapy during the recruitment period April 2001 to April 2003 were included in the study. Patients with a history of major depressive disorder were excluded. RESULTS: Of 113 co-infected patients who started IFN-alpha therapy during the recruitment period, 45 (40%) developed symptoms of depression (sadness, tiredness and apathy). Twenty of them (44%) were treated with citalopram, a selective serotonin re-uptake inhibitor, resulting in a significant improvement in their symptoms. Most of the patients (60%) showed depressive side effects in the first 3 months after initiation of IFN-alpha. In addition, during the study, three patients developed psychotic symptoms and one committed suicide. CONCLUSIONS: The incidence of depressive symptoms in patients with HIV/HCV co infection treated with IFN-alpha is high. Most of the depressive symptoms were not severe and improved with antidepressant therapy, without reduction or cessation of IFN-alpha therapy. During the first weeks after initiating IFN-alpha therapy for HIV/HCV co-infection, close assessment of psychiatric symptoms is recommended. Early treatment of these side effects with antidepressants would help avoid early dropouts from interferon therapy. PMID- 15651750 TI - Antiretroviral drugs with adverse effects on adipocyte lipid metabolism and survival alter the expression and secretion of proinflammatory cytokines and adiponectin in vitro. AB - OBJECTIVE: The lipodystrophy syndrome is a major adverse effect of highly active antiretroviral therapy (HAART), associated with altered circulating levels and adipose tissue mRNA expression of proinflammatory cytokines interleukin-6 (IL-6) and tumour necrosis factor (TNF)alpha, and adiponectin. Proinflammatory cytokines and adiponectin, which are secreted by adipose tissue, regulate fat metabolism, insulin sensitivity and adipose cell apoptosis. We examined the direct effects of individual antiretrovirals on lipid metabolism and cytokine and adiponectin production by cultured adipocytes. METHODS: Differentiating 3T3-F442A cells and differentiated 3T3-L1 adipocytes were treated for 12 or 4 days, respectively, with protease inhibitors (PIs) indinavir, nelfinavir, amprenavir, lopinavir and ritonavir, or nucleoside reverse transcriptase inhibitors (NRTIs) stavudine and zidovudine, at near-Cmax concentrations. Lipid metabolism was estimated by Oil Red O staining of intracellular lipids, mRNA expression of fatty acid synthase and adipocyte lipid binding protein 2, and insulin activation of lipogenesis. Apoptosis was estimated by flow cytometry. The expression and secretion of proinflammatory cytokines (IL-6, TNFalpha and IL-1beta) and adiponectin were evaluated by real-time reverse transcription PCR and ELISA. RESULTS: Chronic treatment of 3T3-F442A differentiating adipocytes and differentiated 3T3-L1 adipocytes with PIs and NRTIs reduced lipid accumulation, mRNA expression of lipid markers and insulin-induced lipogenesis. IL-6, TNFalpha, IL-1beta and adiponectin expression and secretion were markedly altered in differentiating 3T3 F442A adipocytes. PIs had either no effect on differentiated 3T3-L1 adipocytes (TNFalpha expression and sucretion) or their effect was less marked than in 3T3 F442A cells. Indinavir and amprenavir did not alter cytokine secretion and expression by mature adipocytes. The effects of stavudine and zidovudine on differentiating and mature adipocytes were similar, despite the difference in treatment procedure. The drugs with the strongest effect on TNFalpha expression also increased adipocyte apoptosis, in contrast to the drugs that only moderately increased TNFalpha expression. CONCLUSIONS: These results suggest that increased cytokine and decreased adiponectin secretion and expression induced by some PIs and NRTIs may contribute to the adipose tissue loss (via apoptosis and lipid leakage) and insulin resistance associated with the lipodystrophy syndrome. PMID- 15651751 TI - Low-dose hepatitis B immune globulin and higher-dose lamivudine combination to prevent hepatitis B virus recurrence after liver transplantation. AB - Post-transplant prevention of hepatitis B virus (HBV) infection is based on treatment with lamivudine and/or hepatitis B immune globulin (HBIG). However, optimum doses and duration for these drugs are not yet clear. We tested high doses of lamivudine (300 mg/day) in combination with low doses of HBIG (200-400 IU/2-4 weeks). Eighty patients who had post-transplant prophylaxis of lamivudine and HBIG were included in the study. Of those, 20 had hepatitis D virus co infection and eight were HBV DNA-positive at the time of transplantation. Ten HBV DNA-positive patients were treated with lamivudine (150 mg/day) before transplantation; all were HBV DNA-negative after lamivudine treatment. All patients in the anhepatic phase were given 4000 IU of HBIG. Following this, 400 or 800 IU HBIG was administered intramuscularly daily for 5-10 days post transplantation and 2-4 times weekly thereafter, according to serum titre of antibodies to hepatitis B surface antigen (anti-HBs). Lamivudine was maintained or initiated at the time of transplantation and was continued indefinitely. Median follow-up was 21 months (range 3-73 months). Recurrence of hepatitis B surface antigen (HBsAg)-positivity occurred in only three out of 78 (4%) patients; two of these three were HBV DNA-positive. Median anti-HBs titre at the final follow-up was 68 IU. Patient and graft survival was 85% at 1 year. In conclusion, a combination of lamivudine 300 mg/day and low-dose HBIG prevents post-transplantation recurrence of hepatitis B, even in the presence of viral replication in the pre-transplant period. PMID- 15651752 TI - Influence of single-nucleotide polymorphisms in the multidrug resistance-1 gene on the cellular export of nelfinavir and its clinical implication for highly active antiretroviral therapy. AB - Protease inhibitors (PIs) such as nelfinavir (NFV) suppress HIV replication. PIs are substrates of P-glycoprotein (P-gp), the product of the multidrug-resistance 1 (MDR1) gene. Three single-nucleotide polymorphisms (SNPs) are present in exons of the MDR1 gene: MDR1 1236, MDR1 2677 and MDR1 3435. We speculated that these genetic polymorphisms affected PI concentration in the cell. To verify this hypothesis, we first genotyped these SNPs in 79 Japanese patients by the SNaPshot method and found incomplete linkage disequilibrium between the SNPs. Because the SNP at MDR1 3435 has been reported to be associated with P-gp expression, we evaluated the effect of that SNP on the export of NFV from HIV-positive patients' lymphoblastoid cell lines by measuring time-dependent decrease in the amount of intracellular NFV by high-performance liquid chromatography. We found the intracellular concentration of NFV in lymphoblastoid cell lines (LCLs) with the homozygous T/T genotype at MDR1 3435 were higher than that with C/C genotype with statistical significance. This suggests that the activity of P-gp in patients' LCL cells with the MDR1 3435 T/T genotype was lower. In a retrospective study we evaluated the effect of the SNPs on CD4 cell count recovery in response to antiretroviral treatment with PIs, and obtained statistically significant evidence that suggested marginal association of the SNP at MDR1 1236 but not at MDR1 2677 or MDR1 3435. As in vitro results were not consistent with the clinical evaluation, clinical importance of MDR1 genotyping for antiretroviral therapy remains to be investigated in a larger, case-controlled study. PMID- 15651754 TI - Virological effects of ISIS 14803, an antisense oligonucleotide inhibitor of hepatitis C virus (HCV) internal ribosome entry site (IRES), on HCV IRES in chronic hepatitis C patients and examination of the potential role of primary and secondary HCV resistance in the outcome of treatment. AB - Antisense oligonucleotides represent a promising class of antiviral agents. ISIS 14803 is a 20-unit phosphorothioate oligodeoxynucleotide that inhibited hepatitis C virus (HCV) replication and protein expression in cell culture and mouse models. A Phase I dose-escalation clinical study of ISIS 14803 was performed in 24 patients with HCV genotype 1 chronic hepatitis C. The patients received 0.5, 1.0, 2.0 or 3.0 mg/kg of ISIS 14803 for 4 weeks. Two of them receiving 2.0 mg/kg, experienced a significant (>1.0 log10) viral load reduction and nine other patients experienced minor (<1.0 log10) viral load reductions that were difficult to definitively distinguish from assay or patient variations. The aims of this study were to examine the effect of ISIS 14803 on its target site and neighbouring region quasispecies evolution, and to determine whether primary and secondary HCV resistance contributed to the observed virological response rate. The HCV internal ribosome entry site (IRES), including the ISIS 14803 target site in virus specimens collected from patients at baseline and end-of-treatment, was sequenced. An extensive IRES quasispecies analysis was performed in 10 of the patients at various time points before, during and after ISIS 14803 treatment. A significant IRES genetic evolution was found in three out of 10 patients through quasispecies analysis suggesting that treatment with ISIS 14803, a drug designed to bind to HCV RNA, exerted a selective pressure on HCV IRES. However, no mutations in the ISIS 14803 target site, which would inhibit binding of the oligonucleotide to HCV RNA, were detected before (primary resistance) or after treatment (secondary resistance) with the oligonucleotide. Furthermore, no obvious nucleotide changes in the surrounding IRES region that might possibly affect oligonucleotide binding were detected. PMID- 15651753 TI - Clevudine therapy with vaccine inhibits progression of chronic hepatitis and delays onset of hepatocellular carcinoma in chronic woodchuck hepatitis virus infection. AB - We examined a rational approach to therapy of chronic hepatitis B virus (HBV) infection that utilized the reduction of viral load combined with appropriately timed immune modulation/stimulation. In a placebo-controlled study, chronic woodchuck hepatitis virus (WHV) carrier woodchucks received clevudine (L-FMAU), previously shown to have especially potent and sustained antiviral activity in woodchucks, for 32 weeks followed by WHV surface antigen (WHsAg) alum-adjuvanted vaccine at 32, 36, 40 and 48 weeks. Clevudine induced significant reductions in viraemia, surface antigenaemia, hepatic WHV nucleic acids, and hepatic core and surface antigens. Viral replication markers remained markedly suppressed in 75% of the clevudine-treated woodchucks following drug withdrawal, but remained at high levels in the vaccine monotherapy and placebo groups. Combination drug and vaccine therapy had benefits based on sustained reduction of viraemia, antigenaemia, and hepatic WHV DNA and RNA; inhibition of progression of chronic hepatitis; reduced frequency of chronic liver injury; and delayed onset of hepatocellular carcinoma (HCC). Combination therapy contributed to prevention of HCC in up to 38% of treated carriers, although the growth rate of established HCC was not affected. This study demonstrates enhanced benefits of combination chemo immunotherapy against viral load and disease progression in chronic hepadnaviral infection, and provides a platform for further development of such treatment regimens. PMID- 15651755 TI - Mitochondrial effects of a 24-week course of pegylated-interferon plus ribavirin in asymptomatic HCV/HIV co-infected patients on long-term treatment with didanosine, stavudine or both. AB - BACKGROUND: It has been suggested that the addition of ribavirin (RBV) as a part of the treatment for chronic hepatitis C virus (HCV) in HIV co-infected patients on didanosine (ddI) or stavudine (d4T) might increase the nucleoside-induced impairment of mitochondrial function. DESIGN: Comparative study to investigate the impact on mitochondrial function of adding RBV to a long-term treatment with ddI, d4T or both in HCV/HIV non-cirrhotic, asymptomatic patients. We included 26 patients: 16 continued with their current antiretroviral therapy (control group) and 10 patients received a concomitant 24-week course of RBV plus pegylated interferon (PEG-IFN) alpha-2b therapy (HCV-treated group). METHODS: We assessed peripheral blood mononuclear cells mitochondrial DNA (mtDNA) content and mitochondrial respiratory chain (MRC) function at baseline and at 24 weeks of follow-up. In the HCV-treated group we performed additional determinations at 12 weeks during anti-HCV therapy and 24 weeks after finishing anti-HCV therapy. RESULTS: Times on ddI or d4T exposure were 194 +/- 54.9 and 131 +/- 66.5 weeks in the HCV-treated and control groups, respectively. There were no differences either in mtDNA content, the enzyme activity of MRC complexes or clinical parameters at baseline. Throughout the study, mitochondrial measurements remained stable within groups and without differences when we compared HCV-treated and control groups. CONCLUSIONS: In our study, the addition of RBV and PEG-IFN during a 24-week period in HCV/HIV non-cirrhotic, asymptomatic patients on long-term ddI, d4T or both had no impact on mitochondrial function. These findings could suggest that additional triggers are required to achieve a critical threshold in the degree of mitochondrial damage needed for symptoms to develop. PMID- 15651756 TI - Health-related quality of life in HIV patients switching to twice-daily indinavir/ritonavir regimen or continuing with three-times-daily indinavir-based therapy. AB - OBJECTIVE: To evaluate health-related quality of life (HRQoL) changes in patients treated with indinavir three-times daily after switching to a twice-daily indinavir/ritonavir regimen or continuing with the same regimen. METHODS: Patients on HAART including indinavir three-times-daily with undetectable viral load were randomly assigned to continue with this therapy or to change to a twice daily indinavir/ritonavir (800/100 mg) regimen. The Medical Outcomes Study HIV Health Survey (MOS-HIV) questionnaire was used as the HRQoL measure. RESULTS: A total of 118 patients participated in the study, of which 59 (50%) were randomly assigned to continue with the three-times-daily regimen. Patients had a mean age of 39 years and 80% of them were male. At baseline, subjects included in the three-times-daily group presented a significantly greater number of symptoms than subjects in the twice-daily group, but no statistically significant differences were observed in MOS-HIV scores between the groups. In the intention-to-treat (ITT) analysis, a reduction in HRQoL scores was observed in both groups, which was greater in the twice-daily group. In the per protocol analysis, reduction of HRQoL was minimal. CONCLUSIONS: A HRQoL deterioration, greater in the twice-daily group, was observed in this study in the ITT analysis, while HRQoL remained stable in both groups in patients who continued with and tolerated the allocated regimen. PMID- 15651757 TI - Long-term follow-up of HIV-infected patients with chronic hepatitis C virus infection treated with interferon-based therapies. AB - BACKGROUND: Chronic hepatitis C virus (HCV) infection is frequent among HIV infected patients. Clearance of serum HCV RNA 6 months after discontinuing HCV therapy is generally interpreted as a cure of HCV infection in HIV-negative subjects. However, the occurrence of liver complications (including hepatocellular carcinoma) and/or HCV relapses in coinfected patients when followed for long periods of time after HCV therapy is not well known. METHODS: All HIV-infected patients who had been treated for chronic hepatitis C at our institution and had a minimum follow-up of 6 months after discontinuing therapy were retrospectively analysed. They had received one of three HCV treatment modalities: IFN monotherapy, IFN plus ribavirin (RBV) or pegylated interferon (PEG-IFN) plus RBV. RESULTS: A total of 351 patients were retrospectively analysed. Sustained virological response (SVR) to HCV therapy had been reached by 77 (22%) of them: 22/119 (18.5%) with IFN monotherapy, 17/106 (16%) with IFN plus RBV and 38/126 (30.2%) with PEG-IFN plus RBV. Considering the HCV genotypes, SVR had been reached by 19/184 (10.3%) of patients with genotype 1, 54/138 (39.1%) with genotypes 2 or 3, and 4/29 (13.8%) of those with genotype 4. Within a total of 4466 patient-months follow-up (mean of 58 months), none of the 77 patients with SVR showed HCV-RNA rebounds, elevations in liver enzymes potentially linked to HCV, development of hepatocellular carcinoma or episodes of decompensated cirrhosis. In contrast, all 274 patients who did not reach SVR with HCV therapy showed evidence of persistent serum HCV RNA and 90% of them showed liver enzyme elevations during a total of 15344 patient-months follow-up (mean of 56 months). Moreover, 11 (4%) developed clinical complications of liver cirrhosis and two of them died of end-stage liver disease. CONCLUSIONS: HCV replication and HCV related liver disease seem to be permanently halted in HIV/HCV-coinfected patients showing HCV-RNA clearance 6 months after completing any kind of IFN based therapy. In contrast, complications of liver disease due to persistent HCV infection continue to occur in non-responders. The role of maintenance HCV therapy should be explored in HIV/HCV-coinfected patients. PMID- 15651758 TI - Interaction of nucleoside inhibitors of HIV-1 reverse transcriptase with the concentrative nucleoside transporter-1 (SLC28A1). AB - Human concentrative nucleoside transporter-1 (hCNT1) (SLC28A1) is a widely expressed, high-affinity, pyrimidine-preferring, nucleoside transporter implicated in the uptake of naturally occurring pyrimidine nucleosides as well as a variety of derivatives used in anticancer treatment. Its putative role in the uptake of other pyrimidine nucleoside analogues with antiviral properties has not been studied in detail to date. Here, using a hCNT1 stably transfected cell line and the two-electrode voltage-clamp technique, we have assessed the interaction of selected pyrimidine-based antiviral drugs, inhibitors of HIV-1 reverse transcriptase such as zidovudine (AZT), stavudine (d4T), lamivudine (3TC) and zalcitabine (ddC), with hCNT1. hCNT1 transports AZT and d4T with low affinity, whereas 3TC and ddC are not translocated, the latter being able to bind the transporter protein. Selectivity appears to rely mostly upon the presence of a hydroxyl group in the 3'-position of the ribose ring. Thus, hCNT1 cannot be considered a broad-selectivity pyrimidine nucleoside carrier; in fact, very slight changes in substrate structure provoke a dramatic shift in selectivity. PMID- 15651759 TI - Increased sensitivity of SARS-coronavirus to a combination of human type I and type II interferons. AB - There is currently an urgent need to identify effective antiviral agents that will prevent and treat severe acute respiratory syndrome coronavirus (SARS-CoV) infection. In this study, we have investigated and compared the antiviral effect of different interferons (IFNs) on SARS-CoV replication in the epithelial kidney monkey Vero cell line. The results showed that SARS-CoV grown in Vero cells is moderately sensitive to IFN-beta and only weakly sensitive to IFN-alpha and IFN gamma, in comparison to other IFN-sensitive viruses, such as those for encephalomyocarditis, vesicular stomatitis and Newcastle disease. Simultaneous incubation of Vero cells with IFN-beta and IFN-gamma indicated that they may act synergistically against SARS-CoV replication. The IFN-induced MxA protein was detected in the IFN-treated Vero cells. The data, however, suggest that the antiviral activity of IFN against SARS-CoV virus is independent of MxA expression. PMID- 15651760 TI - Management of hepatitis B patients with antiviral resistance. AB - Drug resistance is an expected consequence of antiviral therapy for chronic hepatitis B because of the high rate of hepatitis B virus (HBV) replication, the lack of proof-reading during reverse transcription of the pregenomic RNA and the low efficacy of available therapies in eliminating covalently closed circular HBV DNA. Mutations involving the YMDD motif of the catalytic domain of HBV reverse transcriptase have been reported in patients who have received lamivudine, emtricitabine and telbivudine. Drug-resistant mutations affecting other regions of HBV polymerase have also been reported, but at much lower rates in patients who have received adefovir dipivoxil or entecavir. Antiviral resistance is initially manifested as virological breakthrough infection. In most patients, this is followed by biochemical breakthrough and, in some patients, hepatitis flares and hepatic decompensation. Monitoring drug resistance may improve the management of patients with antiviral-resistant HBV and can guide the selection of salvage therapy. The optimal management of patients with antiviral-resistant HBV continues to evolve. The ideal approach is to prevent antiviral resistance through judicious use of antiviral therapy and the use of more potent antiviral agents, possibly in combination. PMID- 15651761 TI - Partial splenic embolization for the treatment of hypersplenism in cirrhotic HIV/HCV patients prior to pegylated interferon and ribavirin. AB - Partial splenic embolization (PSE), a non-surgical treatment for hypersplenism, has also been reported to improve hepatic function. As severe thrombocytopaenia or leukopaenia contraindicate the use of combined therapy with pegylated interferons (PEG-IFNs) and ribavirin (RBV) in HCV-related cirrhosis, we evaluated, from July 2002 to October 2003, the safety and effectiveness of PSE as a procedure to allow therapy for HCV in three Child-Pugh class B cirrhotic patients with hypersplenism and HIV co-infection. HCV genotypes were 1b (n=2) and 3a (n=1). Severe thrombocytopaenia (in all) and leukopaenia (in two) precluded therapy for HCV. PSE was successfully performed in all with a mean infarcted area of 80%, leading to a significant increase in platelet and leukocyte counts that allowed therapy with weight-adjusted RBV and PEG-IFN-alpha-2b (patients 1 and 3) or 180 microg of PEG-IFN-alpha-2a (patient 2) 8 weeks after the procedure. Moderate pain, well controlled with conservative measures, followed PSE in 100% of cases, but during follow-up (mean 422 days) there were no infectious complications or liver decompensation episodes. Although preliminary, these results suggest the potential role of PSE in HIV/HCV-cirrhotic subjects with hypersplenism as a procedure to allow the use of combined PEG-IFN and RBV. PMID- 15651762 TI - The use of saquinavir/ritonavir 1000/100 mg twice daily in patients with tuberculosis receiving rifampin. PMID- 15651763 TI - Saquinavir trough concentration before and after switching NRTI to tenofovir in patients treated with once-daily saquinavir hard gel capsule/ritonavir 1600 mg/100 mg. PMID- 15651764 TI - Clinically validated genotype analysis. PMID- 15651765 TI - Thrombolytic therapy for myocardial infarction facilitated by mobile coronary care. AB - BACKGROUND: The benefit of Thrombolytic Therapy (TT) for acute myocardial infarction is time sensitive. In Northern Ireland widespread availability of mobile coronary care units facilitates delivery of TT to heart attack victims. This region-wide prospective observational study assessed the efficacy of various methods of delivery of TT. METHODS: All 15 acute hospitals providing acute coronary care in Northern Ireland participated and data were collected prospectively over six months on all patients admitted with acute myocardial infarction or who received TT. The information was analysed regarding appropriateness of TT, methods and timeliness of delivery of TT and mortality rates. Performance was measured against National Service Framework standards. FINDINGS: Of 1638 patients with acute myocardial infarction 584 were considered eligible for TT and 494 (85 %) received it, in addition to 18 patients without infarction. Of the 512 thrombolysed patients 282 (55%) were treated in hospital coronary care units, 131 (26%) were treated pre-hospital, 97 (19%) in accident and emergency departments, and two in general medical wards. Overall median call to-needle time was 87 (7-1110) mins and this was shortest for pre-hospital treatment when 55% of call-to-needle times were < or = 60 mins. For patients treated in hospital median door-to-needle time was 46 (0-1065) mins and this was shortest when TT was administered by accident and emergency staff, when 65% of door-to-needle times were < or = 30 mins. In patients with ST elevation myocardial infarction TT was associated with lower mortality, especially when administered pre-hospital. INTERPRETATION: NSF targets for TT are unlikely to be met in Northern Ireland without increasing pre-hospital delivery of TT and by improving collaboration between coronary care and accident and emergency staff with TT availability in accident and emergency departments. PMID- 15651766 TI - Aggressive treatment of metastasis to the parotid. AB - OBJECTIVES: Assess the value of aggressively treating metastatic lesions in the parotid, taking into account the histology of the disease. STUDY DESIGN: Retrospective analysis of 13 patients diagnosed with metastasis to the parotid treated by one surgeon in a tertiary referral head and neck unit in the United Kingdom. METHODS: The following variables were reviewed and tabulated: age, sex, histology, latent period to secondary tumour, treatment instituted, postoperative facial nerve outcome, follow-up and survival. RESULTS: Twelve patients were treated aggressively with at least total parotidectomy and adjunctive therapy, whilst one patient required only a superficial parotidectomy. Ten patients had metastatic cutaneous tumours, and three had metastatic adenocarcinoma. Seven of these 13 patients (53.8 %) are alive and well (six had metastatic cutaneous tumours, one had metastatic adenocarcinoma). Four patients succumbed to tumour (two had metastatic cutaneous tumours and two had metastatic adenocarcinoma), and two patients succumbed from unrelated medical causes (both had metastatic cutaneous tumours). The mean follow-up for those alive is 65.9 months and mean follow-up for those deceased is 15.3 months. CONCLUSIONS: In the absence of systemic spread, parotid metastases from primary cutaneous squamous cell carcinoma should be treated aggressively, while metastases from non-cutaneous primary tumours should be approached with caution. PMID- 15651767 TI - Unrecognised spinal cord compression as a cause of morbidity. AB - Predicting outcomes is important in planning patient management and rehabilitation. Two cases, one with illustrative radiology, are described. Each presented with potentially preventable morbidity, secondary to unrecognised compression of the spinal cord. Detailed history and examination may have revealed the underlying problem: a condition with potential associated long-term morbidity. PMID- 15651768 TI - Antenatal HIV testing: evaluation of uptake and women's attitudes in a low risk population. AB - The number of HIV-positive heterosexuals in the UK is increasing, with a resultant increase in the number of pregnant women who are HIV-positive. The benefits of diagnosing an HIV-positive woman antenatally are well established. The Department of Health of England issued guidelines recommending named voluntary antenatal testing, with a view to achieving a maternal diagnosis rate of 90% by December 2002. In Northern Ireland the policy was distributed in 2003. The screening programme in our hospital had an uptake rate of 98.7 %. Responses to a questionnaire to evaluate the process indicate that HIV testing was associated with low levels of anxiety and that patients were well satisfied with the counselling they received. PMID- 15651769 TI - Knowledge, attitudes and behaviour in the sun: the barriers to behavioural change in Northern Ireland. AB - To inform future health promotion programmes, we studied the knowledge, attitudes and behaviour of the Northern Ireland population to sun care. An interviewer administered questionnaire was applied to one adult per household from a random sample of 1242 addresses. Lower levels of knowledge were found among respondents who were male, aged under 25 years or over 65 years, in a manual occupation or living in the west where health promotion activity on this topic was less active than in the east. Younger adults, females and professional groups were more likely to indicate that a suntan was important, healthy or attractive. Use of high factor sunscreen was inversely proportional to perceived importance of a suntan. Sunburn was more common in younger adults but more men reported multiple episodes of burning. Regular skin checks were uncommon and self-assessment of skin type was unrealistic indicating that sun care advice based on self assessment should be avoided in this population. Future campaigns should target appropriate messages at specific population subgroups. The study highlights the importance of collecting baseline information before implementing health promotion programmes and suggests that repeat monitoring is essential to ensure that key messages remain relevant. This study also indicates that Care in the Sun campaigns here impacted on general awareness in the population even with limited resources. There is, therefore, potential for greater impact with high funding levels. PMID- 15651770 TI - All-rounders and 'equanimity'--Terence John Millin (1903--1980), Irish urological surgeon. A lecture to commemorate professor Gary Love (1934--2001). PMID- 15651771 TI - A new RVH for a new century: maintaining clinical excellence. Annual oration: Royal Victoria Hospital, Belfast, October 2003. PMID- 15651772 TI - Lemierre syndrome: remember the forgotten disease. PMID- 15651773 TI - Gastrobronchial fistula--a complication of splenectomy. PMID- 15651774 TI - Acute presentation of lymphangioma of the retroperitoneum. PMID- 15651775 TI - Solitary extramedullary plasmacytoma of the tongue. PMID- 15651776 TI - "The lucky penny"--an incidental finding of hip dysplasia in a child with foreign body ingestion. PMID- 15651777 TI - Merkel cell carcinoma of the breast: report of a case and review of the literature. PMID- 15651778 TI - Secondary haemorrhage after rubber band ligation of haemorrhoids in patients taking clopidogrel--a cautionary note. PMID- 15651779 TI - Oligomers of "extended viologen", p-phenylene-bis-4,4'-(1-aryl-2,6 diphenylpyridinium), as candidates for electron-dopable molecular wires. AB - We report the synthesis and spectral characterization of the first five members of an oligomeric series built from alternating p-connected 1,4-benzene and 1,4 pyridinium rings, 1[n]-4[n], n=1-5, with p-phenylene-bis-4,4'-(1-aryl-2,6 diphenylpyridinium) ("extended viologen") as the repeating unit. The lengths of these rodlike molecules range from 2 to 9 nm. The monomer was obtained from p phenylene-bis-4,4'-(2,6-diphenylpyrylium) (5) and p-phenylenediamine (6) or p aminoacetanilide (9). Higher oligomers were synthesized by stepwise elaboration of the monomer by reactions with the appropriate bis-pyrylium (5) or pyrylium phenylene-pyridinium (8) salts. Eight different counterions were used, and dodecamethylcarba-closo-dodecaborate was found to offer particularly favorable solubility characteristics. Ultraviolet absorption spectra of the oligomers suggest that the individual extended viologen segments interact only weakly, as a result of the strongly twisted orientation of the benzene rings that separate them. The UV spectrum of the monomer was interpreted by comparison with semiempirical INDO/S calculations performed at a DFT optimized geometry. PMID- 15651780 TI - Double N-arylation of primary amines: carbazole synthesis from 2,2' biphenyldiols. AB - The double N-arylation of primary amines with 2,2'-biphenylylene ditriflates was investigated for the synthesis of multisubstituted carbazoles. Palladium complexes supported by 2-dicyclohexylphosphino-2'-methylbiphenyl or Xantphos [4,5 bis(diphenylphosphino)-9,9-dimethylxanthene] were found to be efficient catalysts for the reaction. The catalysts allow the use of anilines with an electron donating or electron-withdrawing substituent and multisubstituted 2,2' biphenylylene ditriflates as substrates. Ammonia equivalents, such as O-tert butyl carbamate, are also employable as a nitrogen source to give the N-protected carbazoles which can easily give the corresponding N-unsubstituted carbazoles after deprotection. By using this methodology, a carbazole alkaloid, mukonine, is synthesized in 40% yield for five steps, in comparable efficiency to the recent precedents. PMID- 15651781 TI - Reactivity of lactones and GHB formation. AB - The behavior of lactones in their hydrolysis reactions is a good indicator of their reactivity as electrophilic molecules. The hydrolysis of four- to six membered lactones was investigated in neutral (water) and slightly acid media and in water/dioxane media. The following conclusions were drawn: (i) The reactivity of beta-propiolactone in neutral water is more than four times greater than that of beta-butyrolactone, due to the flow of charge caused by the latter's methyl substituent. Reactivity is enthalpy-controlled. (ii) The reactivity of beta lactones diminishes in water/dioxane media when the percentage of dioxane increases. The increase in the dioxane percentage relaxing the intermolecular hydrogen bonds in the ordered structure of the water reduces DeltaH# and simultaneously increases the -DeltaS# value. (iii) An inverse solvent kinetic isotope effect in the acid-catalyzed hydrolysis of gamma-butyrolactone and delta valerolactone was observed, this being indicative of acyl cleavage. (iv) The DeltaH# and DeltaS# values permit discrimination between alkyl and acyl cleavage. (v) A correlation was found between the chemical reactivity of lactones and their carcinogenic activity. (vi) The results suggest that orally ingested gamma butyrolactone remains largely in its nonhydrolyzed form in the stomach before passing into the blood. (vii) The concentration equilibrium constant of GHB formation at human body temperature is Keq (37 degrees C)=0.40. (viii) Study of GHB formation shows that, contrary to earlier results, this is an endothermic process, with DeltarH=3.6 kJ mol(-1). PMID- 15651782 TI - Synthesis, conformation, and complexation of novel 25,26,27,28- tetrahydroxy 5,11:17,23-bis[[2,2'-thioxydi(o-phenylene)dithioxy]- diphenylthio]calix[4]arene. AB - Calix[4]arenes 1a,b having an electron-donating group, i.e., OH and OMe, at the lower rim reacted with thianthrene cation radical perchlorate in CH3CN at room temperature to give the corresponding thianthrenium perchlorates 3a,b in excellent yields. Treatment of 3a,b comprising a mixture of conformational isomers with NaSH.xH2O in DMF at reflux afforded the sulfur-containing cyclized compounds 4a,b, respectively. Compound 4a having a cone conformation consisted of two conformational isomers in a ratio of 0.077:1. Temperature-dependent 1H NMR study in DMF showed that conformational isomerization between the two isomers occurred with energy barriers of 14.97 and 14.10 kcal/mol at 100 and 110 degrees C, respectively. The Jobs plot of 4a against Ag+ picrate indicated that compound 4a strongly produced a 1:2 complex with Ag+ ions. Molecular mechanics calculations indicated that the conformation of the energy-minimized 4a-2Ag+ picrate complex had two crushed pyramidal geometries made up of Ag+ ion and four sulfur atoms, i.e., S1, S2, S4, S5 and S6, S7, S9, S10, respectively. PMID- 15651783 TI - Polystyrene-supported (R)-2-piperazino-1,1,2-triphenylethanol: a readily available supported ligand with unparalleled catalytic activity and enantioselectivity. AB - A very active and highly enantioselective catalytic resin, designed for minimal perturbation of the catalytic center by the polymer matrix, has been assembled in two steps from (S)-triphenylethylene oxide, piperazine, and Merrifield resin and tested in the enantioselective ethylation of aldehydes. 1-Arylpropanols of 94-95% ee are obtained in high yield by the use of only 2 mol % of catalytic resin at 0 degrees C for 4 h. PMID- 15651784 TI - A new synthetic route to (North)-methanocarba nucleosides designed as A3 adenosine receptor agonists. AB - Activation of the A3 adenosine receptor (AR) is associated with cerebroprotective, cardioprotective, and anticancer effects. Among potent and selective A3 AR agonists are novel methanocarba adenosine analogues in which the conformation of a pseudo-ribose moiety is locked in the North (N) hemisphere of the pseudorotational cycle. 5'-Uronamide (N)-methanocarba nucleosides, such as MRS1898 and MRS2346, are examples of full agonists of the human A3 AR. An improved convergent approach from easily accessible 2,3-O-isopropylidene-d erythrose (2b), and the combination of a strategic intramolecular cyclopropanation step plus the acid-catalyzed isomerization of an isopropylidene group, provided a suitable pseudosugar precursor (23) for the synthesis of MRS1898, MRS2346, and related analogues. This new synthetic route uses readily available building blocks and opens the way for the preparation of a variety of targets on a reasonable scale. PMID- 15651785 TI - Catalytic asymmetric addition of alkylzinc and functionalized alkylzinc reagents to ketones. AB - We describe our full report of the catalytic asymmetric addition of simple and functionalized dialkylzinc reagents to a broad range of saturated ketones and enones. The functionalized organozinc reagents contain esters, silyl ethers, alkyl chlorides, and alkyl bromides. In general, the resulting tertiary alcohol products are isolated with high ee's. With some substrates, yields are low as a result of the formation of aldol byproducts. Most substrates undergo additions with good yields reaching as high as 91%. PMID- 15651786 TI - Stereomutations of two-bladed propeller derivatives: ortho-substituted diaryl ethylene and diaryl ketone. AB - Stereochemical analysis, supported by ab initio computations, predicts the existence of three possible stable helical conformers for o,o'-diisopropyl-1,1' diphenylethylene (1) and o,o'-diisopropylbenzophenone (2). At low temperature the NMR spectra of 1 showed distinct sets of signals for these conformers, thus allowing the measurement of the three barriers involved in the related stereomutation processes to be obtained (DeltaG=6.45, 4.65, and < or =4.0 kcal mol(-1)). The NMR spectra also indicate that the asymmetric conformer (C1 point group) is the most stable one in solution, as anticipated by calculations. X-ray diffraction confirmed that this structure is that adopted in the crystalline state. On the other hand, o,o'-diisopropylbenzophenone (2) is predicted by calculations to exist essentially as a C2-type conformer, a result that was confirmed by the low-temperature NMR spectra. The interconversion barrier for the enantiomeric forms of this conformer was also measured (DeltaG=6.3(5) kcal mol( 1)). PMID- 15651787 TI - New synthesis of chiral 1,3-oxazinan-2-ones from carbohydrate derivatives. AB - Chiral 1,3-oxazinan-2-ones are useful intermediates in synthesizing pharmaceutical compounds and amino alcohols. In this paper, we report a new synthetic method to chiral 6-hydroxymethyl 1,3-oxazinan-2-ones and their analogues from carbohydrate derivatives. The synthesis was accomplished by the reaction of optically pure 3-hydroxy-gamma-butyrolactone with a primary amine to give an amide, which was reduced and carbonylated to give the desired compound class. PMID- 15651788 TI - Ring-opening of unsymmetrical 1,2-dioxines using cobalt(II) salen complexes. AB - The regioselectivity of the metal-catalyzed ring opening of unsymmetrical 1,2 dioxines to cis-gamma-hydroxyenones was investigated using two different Co(II) salen complexes. Regioselectivity was determined by direct examination of the enone ratios and by derivitization with a stabilized phosphorus ylide. The steric influence of the substituents on the 1,2-dioxine was the primary influence on regioselectivity. Temperature played little role; however, solvent and selection of Co(II) complex could be used to mildly influence the outcome of the rearrangement for selected substrates. The origins of the selectivity for the reaction are discussed. PMID- 15651789 TI - General and selective synthesis of (Z)-3-haloacrylates via palladium-catalyzed carbonylation of terminal alkynes. AB - A general and selective palladium-catalyzed carbonylation of terminal alkynes method for the synthesis of (Z)-3-haloacrylates is presented. In the presence of a catalytic amount of PdX2 and 5 equiv of CuX2 (X=Cl and Br), terminal alkynes were carbonylated to afford the corresponding (Z)-3-haloacrylates exclusively in moderate to good yields. The results showed that the effect of solvent had a fundamental influence on the chemoselectivity and stereoselectivity of the palladium-catalyzed carbonylation reaction. PMID- 15651790 TI - Michael addition-elimination mechanism for nucleophilic substitution reaction of cycloalkenyl iodonium salts and selectivity of 1,2-hydrogen shift in cycloalkylidene intermediate. AB - Reactions of cyclohexenyl and cyclopentenyl iodonium salts with cyanide ion in chloroform give cyanide substitution products of allylic and vinylic forms. Deuterium-labeling experiments show that the allylic product is formed via the Michael addition of cyanide to the vinylic iodonium salt, followed by elimination of the iodonio group and 1,2-hydrogen shift in the 2-cyanocycloalkylidene intermediate. The hydrogen shift preferentially occurs from the methylene rather than the methine beta-position of the carbene, and the selectivity is rationalized by the DFT calculations. The Michael reaction was also observed in the reaction of cyclopentenyliodonium salt with acetate ion in chloroform. The vinylic substitution products are ascribed to the ligand-coupling (via lambda3 iodane) and elimination-addition (via cyclohexyne) pathways. PMID- 15651791 TI - New approaches to bicyclic vinyl heterocycles from propargylic acetals. AB - The paper describes further studies on the intramolecular carbolithiation of propargylic acetals with aryllithiums leading to 2-vinylbenzofurans and 3 vinylfuropyridines. Attempts to extend the cascade to [4.4.0] binuclear heterocycles met with limited success. An alternative, two-step entry to such ring systems has been developed using the palladium-induced cyclization/hydride capture methodology. A new route to isoquinilinones from simple benzamides is also disclosed. PMID- 15651793 TI - Exploratory studies aimed at a synthesis of vinigrol. 2. Attempts to exploit ring closing metathesis for construction of the central cyclooctane belt. AB - A program directed to the possible adaptation of ring closing metathesis to a total synthesis of vinigrol is described. With a convenient route to intermediates of general type 3 available from a prior investigation, several candidate substrates were prepared. These included the epoxy dienes 10 and 22, the diacetoxy triene 42, and the heavily functionalized cyclohexane 48. The central issue of this approach was to convey a maximum degree of conformational flexibility to these functionalized intermediates, such that the olefinic termini of the side chains could enter into intramolecular carbon-carbon bond formation. In no example was ring closure observed to operate. Instead, the strategically placed pi-bonds were seen to migrate internally to the chain in select examples. Although the pivotal transformations failed, the deployment of a number of useful stereo-controlled reactions has ultimately resulted in the preparation of heavily substituted cis-decalins. PMID- 15651792 TI - Syntheses of (+)-cytisine, (-)-kuraramine, (-)-isokuraramine, and (-)-jussiaeiine A. AB - Total syntheses of (+)-cytisine, (-)-kuraramine, (-)-isokuraramine, and (-) jussiaeiine A were achieved via a samarium diiodide-promoted reductive deamination reaction, followed by simultaneous recyclization of a proline derivative to give the corresponding delta-lactam derivative, as a key step. PMID- 15651794 TI - Exploratory studies aimed at a synthesis of vinigrol. 3. Evaluation of a lactone bridge as a conformational lock. AB - Evaluated in the present investigation are possible synthetic approaches to vinigrol based on the involvement of lactone rings as tools for the conformational rigidification of functionalized cis-octalins. Emphasis was placed on the structural arrangements resident in 3 and 5. The first of these systems proved to be highly strained and inaccessible. Especially notable was the finding that hydroxy ketenes 14 and 21 could be isolated and shown not to be amenable to cyclization when heated. The stereo-controlled assembly of 5 was successfully accomplished through exploitation of a related synthetic pathway. However, neither this attractive intermediate nor its close relative 33 could be processed in a manner that delivered the vinigrol framework. Nonetheless, several features of the routes deployed offer the prospect of wider application in other contexts. PMID- 15651795 TI - Exploratory studies aimed at a synthesis of vinigrol. 4. Probe of possible means for direct connection of the side arms and of ring-contraction alternatives. AB - Attempts have been made to gain access to the vinigrol structural framework by way of three routes. These include reductive transannular cyclization, adaptation of the Ramberg-Backlund rearrangement, and deployment of the lactam-sulfoxide ring contraction protocol. While the first of these options involves direct transannular C-C bond formation, the other two embody the concept of larger ring construction as a prelude to ring contraction. The initial installation of a sulfur atom involves prior thiacyclononane formation, a process believed to be potentially easier to accomplish. However, arrival at 13, 14, or 17 was not achieved. Installation of the heterocyclic ring contained in 31 proved to be equally problematic. Increased disassembly of the molecular structure as featured in dibromide 20 did allow for direct conversion to sulfone 22. This advanced building block proved not be conducive to in situ alpha-chlorination and extrusion of the sulfur atom. PMID- 15651796 TI - A study of vinyl radical cyclization using N-alkenyl-7-bromo-substituted hexahydroindolinones. AB - A new method for the synthesis of the octahydropyrrolo[3,2,1-ij]quinoline ring system that possesses the characteristic skeleton of the aspidosperma family of alkaloids has been developed. The method utilizes an intramolecular Diels-Alder reaction of an amido-substituted furan across a tethered indole pi-bond. To apply this strategy to the synthesis of the indole alkaloid spegazzinidine, it was necessary to address the problem of assembling the final D-ring of the pentacyclic skeleton. Radical cyclization of a model N-allyl-7-bromo-3a methylhexahydroindolinone system was found to preferentially lead to the 6-endo trig cyclization product, with the best yield being obtained under high dilution conditions. The six-membered cyclized product is generated through two reaction pathways: (a) 6-endo-trig ring closure and (b) rearrangement of an intermediate methylene-cyclopentyl radical obtained by 5-exo-trig cyclization. A number of related 7-bromo-substituted hexahydroindolinones containing tethered olefinic groups were prepared and found to undergo efficient cyclization under both radical and palladium-mediated reaction conditions. Vinyl radical cyclization with several N-butenyl-substituted systems afforded a mixture of 6-exo and 7-endo cyclization products. A protocol to introduce an ethyl substituent into the C20 position of the aspidospermidine skeleton was also developed. PMID- 15651797 TI - A new entry to asymmetric synthesis of optically active alpha,gamma-substituted gamma-butyrolactones, using a carbohydrate derived amide as both a chiral auxiliary and a proton source. AB - A new entry for the asymmetric synthesis of optically active alpha,gamma substituted gamma-butyrolactones was developed by using a carbohydrate-derived amide as both a chiral auxiliary and a proton source. Unlike the previously reported examples, the chiral auxiliary employed in this reaction also successfully functioned as a protonating agent. Excellent asymmetric induction could be achieved by this dual stereoselective control in the reaction process. PMID- 15651798 TI - Diastereoselectivities realized in the amino acid catalyzed aldol cyclizations of triketo acetonides of differing ring size. AB - A study designed to assess the diastereoselectivity of the intramolecular aldol reaction of two differently sized monocyclic 1,3-diketones bearing a chiral, oxygenated side chain has been undertaken. The cyclizations were brought about under catalysis by pyrrolidine, a series of D- and L-amino acids including proline, and several proline derivatives. The levels of selectivity were found to be consistently higher with the six-membered ring system than its cycloheptane counterpart. PMID- 15651799 TI - A theoretical study of the competitive homolytic/heterolytic aniomesolytic cleavages of C-O alkyl ether bonds. AB - Density functional theory electronic structure calculations of the homolytic/heterolytic aniomesolytic C-O fragmentations in the gas phase of a series of radical anions of substituted-phenyl benzyl ethers and substituted benzyl phenyl ethers have been carried out. Along the series, the electron withdrawing strength of the substituents increases. An intramolecular electron transfer from the pi system to the sigma molecular orbital of the scissile C-O bond is required to produce the fragmentation. As the electron-withdrawing strength of the substituents increases, the transition-state structures appear later with higher potential energy and Gibbs free energy barriers. The homolytic mesolytic cleavages are always thermodynamically favored versus the corresponding heterolytic mesolytic ones. The heterolytic mesolytic fragmentations in radical anions containing only weak electron-withdrawing groups are faster than the corresponding homolytic mesolytic ones. Conversely, in radical anions supporting strong electron-withdrawing groups the homolytic mesolytic fragmentations are faster in terms of potential energy barriers. However, the entropic contribution makes it comparable the homolytic and the heterolytic Gibbs free energy barriers in this case. The main factors that determine the relative rates of those kind of aniomesolytic cleavages are discussed. PMID- 15651800 TI - Scope and limitations of the nitro-Mannich reaction for the stereoselective synthesis of 1,2-diamines. AB - The acetic acid-promoted addition of lithium nitropropanate and the Lewis acid catalyzed [Sc(OTf)3, Cu(OTf)2, or Ti(OiPr)4] addition of trimethylsilyl nitropropanate to a range of heteroaromatic and simple aliphatic aldimines gave anti-rich (approximately 3-19:1) beta-nitroamines in >95% yields as the kinetic products. It was found that a nonpolar N-imine protecting group was essential for reactivity with the o-methoxybenzyl (OMB) group giving better selectivities and yields than p-methoxybenzyl (PMB) or p-methoxyphenyl (PMP) in the Lewis acid catalyzed addition reactions. Reduction with SmI2, treatment with COCl2, followed by OMB deprotection gave diastereomerically pure cis-imidazolidinones in 55-79% overall yield from imine. Preliminary results have shown that acetic acid can catalyze the reaction of N-OMB-benzylideneamine with nitropropane, used as solvent, to give the thermodynamically more stable syn-beta-nitroamine product. PMID- 15651801 TI - Regioselective hydrosilylations of propiolate esters with tris(trimethylsilyl)silane. AB - Lewis acid and substituent dependency on the regioselectivity of hydrosilylation of propiolate esters 1a-c with tris(trimethylsilyl)silane (2a) was found. The reaction of methyl and ethyl propiolate esters and 2a without Lewis acid and in the presence of EtAlCl2 and Et2AlCl gave beta-silicon-substituted Z-alkenes 3 selectively. On the other hand, reaction in the presence of AlCl3 in dichloromethane gave alpha-silicon-substituted alkenes 4. In the case of trifluoroethyl propiolate ester 1c, reaction with aluminum chloride-based Lewis acids gave alpha-silicon-substituted alkenes 4 exclusively. Two competitive mechanisms, free-radical and ionic, are proposed as the source of the complementary regioselectivity displayed in these reactions. A transition state of the radical-forming step was obtained computationally. The reaction of various reactive acetylene substrates and 2a without Lewis acid and without solvent at room temperature gave beta-silicon-substituted Z-alkenes 3 selectively. PMID- 15651802 TI - The intermediacy of metathiophosphonate PhPSO in the reaction of N-tert-butyl-P phenylphosphonamidothioic acid with alcohols. AB - Kinetic studies of the reaction of N-tert-butyl-P-phenylphosphonamidothioic acid (1) with alcohols were carried out in CH2Cl2 by means of 31P NMR spectrometry. The reaction is of the first order with respect to thio acid 1. The first-order rate constant at 30 degrees C increases with increasing methanol concentration below 0.25 M, but otherwise the rate constants are either independent of alcohol concentration (MeOH above 0.25 M, BuOH) or decrease with increasing alcohol concentration (i-PrOH, t-BuOH). The effect of alcohols on the order of the reaction and parameters of activation, as well as results of competition experiments, lead us to the conclusion that reaction of 1 with alcohols occurs by an elimination-addition mechanism involving the association of the thio acid 1 and the alcohol and then formation in the rate-determining step of an encounter complex 2' ' involving metathiophosphonate 4, amine, and alcohol. Metathiophosphonate 4 reacts preferentially with the alcohol as the encounter complex (primary alcohols) or after diffusion apart as a "free" intermediate (hindered alcohols). PMID- 15651803 TI - Investigations of the halogen dance reaction on N-substituted 2-thiazolamines. AB - The halogen dance (HD) reaction on various 2-thiazolamine systems was investigated providing an easy access to a series of 5-substituted 4-bromo-2 thiazolamine derivatives. We could show that HD is a very favored process for the investigated systems and that prevention of HD is only possible when optimized reaction conditions and selected electrophiles are applied. PMID- 15651804 TI - A novel highly selective chiral auxiliary for the asymmetric synthesis of L- and D-alpha-amino acid derivatives via a multicomponent Ugi reaction. AB - This paper describes the synthesis of a bicyclic beta-amino acid scaffold in both pure enantiomeric forms and its application as chiral auxiliary in an intramolecular version of the Ugi multicomponent reaction (U-5C-4CR) to prepare alpha-amino acid derivatives of both D- and L-series in a straightforward and very stereoselective manner. The mild conditions required for the Ugi condensation and for the removal of the chiral auxiliary make this method very attractive to prepare a wide range of differently structured N-alkylated and unalkylated amino acid derivatives. PMID- 15651805 TI - Conjugate additions of a simple monosilylcopper reagent with use of the CuI.DMS complex: stereoselectivities and a dramatic impact by DMS. AB - Conjugate additions utilizing the simple monosilylcuprate reagent Li[PhMe2SiCuI] to alpha,beta-unsaturated carbonyl compounds are described. The presence of dimethyl sulfide (DMS), either as a component originating from the (CuI)4(DMS)3 complex or as a solvent added, has an amazing influence on both chemical yield and the level of diastereomeric ratio (dr) of the products. Gilman-type silylcyanocuprates {Li(Ph2MeSi)2Cu/LiCN} have previously been used to guarantee good results in conjugate addition reactions. External additives such as HMPA, tributylphosphine, or dialkylzinc are not necessary in conjunction with the simple Li[PhMe2SiCuI] reagent. It is demonstrated that the monosilylcuprate reagent with DMS as the solvent is very useful with sterically hindered (beta,beta-disubstituted) enones, and provides very high yields of the beta silylated 1,4-addition products. Since there is no oligomerization problem associated with the simple monosilylcuprate reagent, this reagent should be considered as a very useful 1,4-silyl donor to enals, enones, and enoates in conjugate addition reactions. PMID- 15651806 TI - The rearrangement route to 3-carboxy- and 3-hydroxymethyl-2 azabicyclo[2.1.1]hexanes: 3,5-methanoprolines. AB - Improved stereocontrolled syntheses of 5-anti-hydroxy-3-exo-methoxycarbonyl-2 azabicyclo[2.1.1]hexanes have been effected from pyridine. The key step in the electrophilic addition-rearrangement of 2-azabicyclo[2.2.0]hex-5-ene precursors incorporates either a 3-endo-phenyl group, as an acid precursor, or a 3-endo phenyldimethylsilylmethyl group, as a potential hydroxymethyl and acid precursor. PMID- 15651807 TI - Synthesis of 1-aryl-1H-indazoles via palladium-catalyzed intramolecular amination of aryl halides. AB - Palladium-catalyzed cyclization of arylhydrazones of 2-bromoaldehydes and 2 bromoacetophenones to give 1-aryl-1H-indazoles has been studied in detail. The cyclization of arylhydrazone of 2-bromobenzaldehydes can be performed with good to high yields using Pd(dba)2 and chelating phosphines, of which the most effective are rac-BINAP, DPEphos, and dppf, in the presence of Cs2CO3 or K3PO4 as a base. Electron-rich, bulky ligands commonly employed for intermolecular amination such as PtBu3 and o-PhC6H4PtBu2 were shown to be ineffective for cyclization and to lead instead to extensive oligomerization and tarring. The method developed is applicable for preparation of a wide scope of indazoles bearing electron-donating or electron-withdrawing substituents, among them, unprotected carboxyl, as well as various indazole heteroanalogues. The cyclization of arylhydrazones of less reactive halides such as 2 chlorobenzaldehyde, as well as 2-bromoacetophenone and bromotetralone, has been achieved. The purity of the starting hydrazone has been shown to be a critical parameter, as various impurities inhibit the cyclization. PMID- 15651808 TI - (Sensitized) photolysis of diazonium salts as a mild general method for the generation of aryl cations. Chemoselectivity of the singlet and triplet 4 substituted phenyl cations. AB - The photolysis of a series of 4-X-benzenediazonium tetrafluoroborates is studied in MeCN. Loss of nitrogen occurs from the singlet excited state with X=H, t-Bu, and NMe2 and leads to the singlet aryl cation. This adds to the solvent yielding the corresponding acetanilides. With other substituents, ISC competes with (X=Br, CN) or overcomes (X=COMe, NO2) fragmentation and the aryl cation is formed in part or completely in the triplet state. In neat MeCN, this either abstracts hydrogen from the solvent (in most cases inefficiently) or undergoes intersystem crossing to the more stable singlet that reacts as above. In the presence of pi nucleophiles (allyltrimethylsilane or benzene), the triplet aryl cation is efficiently trapped giving substituted allylbenzenes and biphenyls, respectively. By triplet sensitization by xanthone, the triplet cation and the products from it are obtained from the whole series considered. The direct or sensitized photodecomposition of diazonium fluoroborates, substituted with both electron donating and -withdrawing substituents, in the presence of alkenes and arenes offers an access to an alternative arylation procedure. PMID- 15651809 TI - Regioselectively nucleus and/or side-chain fluorinated 2-(Phenanthryl)propionic acids by an effective combination of radical and organometallic chemistry. AB - Regioselectively nucleus and/or side-chain fluorinated 2-(phenanthr-1-yl)- and 2 (phenanthr-2-yl)propionic acids 1-5 were prepared using phenanthren-1(2H)-ones 6a c as key intermediates. Thus, ethyl 2-(fluorophenanthryl)propionates 11 were obtained in good yields by Reformatsky reaction of 6a-c with ethyl 2 bromopropionate followed by dehydratation and DDQ-promoted aromatization of the resulting beta-hydroxyesters. Side-chain alkyl 2-hydroxy-2-(phenanthr-1 yl)propionates 14 were obtained by bromine/lithium permutation of dihydrophenanthryl bromides 12a-c with butyllithium followed by quenching of the lithiated intermediates with methyl pyruvate or ethyl 3,3,3-trifluoropyruvate and subsequent DDQ-promoted aromatization. The alkyl 2-hydroxy-2-(phenanthr-1 yl)propionates 25 were prepared by reacting 8-bromo-1,3-difluorophenanthrene 24 with butyllithium for 10 seconds at -110 degrees C and subsequent addition of the suitable pyruvate to the lithiated intermediates. Alkyl 2-hydroxy-2-(phenanthr-2 yl)propionates 26 and 29 were suitably obtained by site-selective metalation of 1,3-difluorophenanthrene 28 and the bromophenanthrene 24, respectively, with LDA followed by quenching of the metalated intermediates with the suitable alkyl pyruvate. Fluorination of the above alpha-hydroxypropionates with DAST, followed by the alkaline hydrolysis, allowed the expected 2-(phenanthryl)propionic acids 1 5 to be obtained in satisfactory overall yields. PMID- 15651810 TI - Di- and trinuclear Zn2+ complexes of calix[4]arene based ligands as catalysts of acyl and phosphoryl transfer reactions. AB - The calix[4]arene scaffold, blocked in the cone conformation by proper alkylation of the lower rim hydroxyls, was used as a convenient molecular platform for the design of bi- and trimetallic Zn2+ catalysts. The catalytic activity of the Zn2+ complexes of calix[4]arenes decorated at the 1,2-, 1,3-, and 1,2,3-positions of the upper rim with 2,6-bis[(dimethylamino)methyl]pyridine units were investigated in the cleavage of ester 6 and of the RNA model compound HPNP. High rate enhancements, up to 4 orders of magnitude, were observed in a number of catalyst substrate combinations. Interestingly the order of catalytic efficiency among regioisomeric dinuclear complexes in the cleavage of ester 6 is 1,2-vicinal >> 1,3-distal, but it is reversed in the reaction of HPNP. The higher efficiency of trinuclear compared to dinuclear complexes provides an indication of the cooperation of three Zn2+ ions in the catalytic mechanism. PMID- 15651811 TI - Identification of the function of gene lndM2 encoding a bifunctional oxygenase reductase involved in the biosynthesis of the antitumor antibiotic landomycin E by Streptomyces globisporus 1912 supports the originally assigned structure for landomycinone. AB - The angucycline antibiotic family of the landomycins displays potent antitumor activity. To elucidate early post polyketide synthase (PKS) tailoring steps of the landomycin E biosynthetic pathway in Streptomyces globisporus 1912, the mutant S. globisporus M12 was prepared through gene replacement experiment of lndM2. It encodes an enzyme with putative oxygenase and reductase domains, according to sequencing of the gene and its counterpart lanM2 from S. cyanogenus S136 landomycin A biosynthetic gene cluster. The isolation of the novel shunt products 11-hydroxytetrangomycin and 4-hydroxytetrangomycin along with the well known angucyclines tetrangomycin and tetrangulol from the culture of S. globisporus M12 provides evidence for the involvement of lndM2 in the early biosynthetic pathway of the landomycins, in particular in the formation of the alicyclic 6-hydroxy function of the landomycin aglycon. We therefore propose LndM2 to be responsible for both hydroxylation of the 6-position and its subsequent reduction. These reactions are necessary before the glycosylation reactions can occur. The results are in agreement with the originally published structure of landomycin but do not support the recently suggested revised structure. PMID- 15651812 TI - Size-expanded analogues of dG and dC: synthesis and pairing properties in DNA. AB - We describe the completion of the set of four benzo-fused expanded DNA (xDNA) nucleoside analogues. We previously reported the development of benzo-fused analogues of dA and dT and their inclusion in an exceptionally stable new four base genetic system, termed xDNA, in which the base pairs were expanded in size. Here we describe the preparation and properties of the second half of this nucleotide set: namely, the previously unknown dxC and dxG nucleosides. The dxC analogue was prepared from the previously reported dxT nucleoside in three steps and 57% yield. The large-sized deoxyguanosine analogue was prepared from an intermediate in the synthesis of dxA, yielding dxG in 14 steps overall (2.4%). Suitably protected versions of the deoxynucleosides were prepared for oligonucleotide synthesis following standard procedures, and they were readily incorporated into DNA by automated synthesizer. "Dangling-end" measurements revealed that the benzo-fused homologues stack considerably more strongly on neighboring DNA sequences than do their natural counterparts. Base pairing experiments with xC or xG bases showed that they pair selectively with their Watson-Crick partners, but with mild destabilization, due apparently to their larger size. Overall, the data suggest that the fluorescent xG and xC bases may be useful probes of steric effects in the study of biological nucleotide recognition mechanisms. In addition, the completion of the xDNA nucleoside set makes it possible in the future to construct full four-base xDNA strands that can target any sequence of natural DNA and RNA. PMID- 15651813 TI - Catalytic asymmetric synthesis of chiral allylic amines. Evaluation of ferrocenyloxazoline palladacycle catalysts and imidate motifs. AB - Palladium(II) catalysts based on a ferrocenyloxazoline palladacyclic (FOP) scaffold were synthesized and evaluated for the rearrangement of prochiral allylic N-(4-methoxyphenyl)benzimidates. When iodide-bridged dimer FOP precatalysts are activated by reaction with excess silver trifluoroacetate, the allylic rearrangement of both E and Z prochiral primary allylic N-(4 methoxyphenyl)benzimidates takes place at room temperature to give the corresponding chiral allylic N-(4-methoxyphenyl)benzamides in high yield and good ee (typically 81-95%). Several allylic imidate motifs were evaluated also. Because the corresponding enantioenriched allylic amide products can be deprotected in good yield to give enantioenriched allylic amines, allylic N aryltrifluoroacetimidates were identified as promising substrates. PMID- 15651814 TI - Structure elucidation and synthesis of lycoposerramine-B, a novel oxime containing Lycopodium alkaloid from Lycopodium serratum Thunb. AB - A new alkaloid, lycoposerramine-B (1), containing an oxime function, was isolated from the club moss Lycopodium serratum Thunb. The structure of 1 was elucidated by spectroscopic analysis, including J-resolved HMBC spectroscopy, and confirmed by its synthesis from the known alkaloid, serratinine (3). PMID- 15651815 TI - Synthesis, chromatographic separation, vibrational circular dichroism spectroscopy, and ab initio DFT studies of chiral thiepane tetraol derivatives. AB - Optically pure enantiomers of the chiral tetrahydroxythiepane derivative 3,6 dihydroxy-4,5-O-isopropylidene-thiepane (3) are obtained using a novel protocol in which a library of all possible stereoisomers of 3 is synthesized, followed by two-step stereoselective chromatography, using, first, conventional achiral and, then, chiral stationary phases. Configurational and conformational analysis of 3 are carried out using Vibrational Circular Dichroism (VCD) spectroscopy in conjunction with ab initio DFT calculations. The absolute configuration of 3 is shown to be 3R,4S,5R,6R-(+)/3S,4R,5S,6S-(-). PMID- 15651816 TI - Luminescent crown ether amino acids: selective binding to N-terminal lysine in peptides. AB - Crown ether amino acids (CEAAs) with a luminescent phthalic ester or phthalimide moiety have been prepared. Simple peptide chemistry covalently tethers the macrocycles to give ditopic ammonium-ion binders. The binding events of both crown ether groups are monitored independently by changes of their specific emission properties. The affinity of the bis-CEAA to bis-ammonium ions is distance dependent, which allows distinguishing between isomeric small peptides containing a lysine residue in different positions. PMID- 15651817 TI - 13-methyl-2,6-dithia[7]metacyclophane: a useful molecule to connect VT NMR results and structure with calculations. AB - Ground state energies (DFT) and 1H and 13C NMR chemical shifts are calculated for the conformers of 13-methyl-2,6-dithia[7]metacyclophane (1), and the results are compared with X-ray structural data and variable-temperature NMR data, including the determination of the activation barrier. Calculations predict the correct low energy conformer with good agreement with chemical shifts, bond distances, and angles. VT NMR data for the 10-tert-butyl-substituted derivative 2 indicate that it undergoes the same conformational equilibria as 1. This paper should enhance the confidence that organic chemists have in calculations to satisfactorily predict conformer energies. PMID- 15651818 TI - Thiol-catalyzed acyl radical cyclization of alkenals. AB - Thiol-catalyzed direct generation of acyl radicals and their intramolecular addition to olefins of alkenals gave 2-substituted five- and six-membered cyclic ketones in reasonably good yields. The combination of odorless tert-dodecanthiol and AIBN or V-40 was the initiator of choice among surveyed radical generators for the cyclization of alkenals. Aldehydes having electron-deficient olefins cyclized more easily than those having unactivated olefins. PMID- 15651819 TI - Competing reactions of secondary alcohols with sodium hypochlorite promoted by phase-transfer catalysis. AB - With aqueous hypochlorite and a phase transfer catalyst, secondary alcohols undergo hitherto unreported free radical reactions that compete with and effectively limit traditional ketone syntheses. Product mixture profiles are determined by reactant ratios, organic cosolvent, and availability of oxygen to the system. Under argon, over half of substrate alcohols, PhCH(OH)R, are converted to benzaldehyde and free radical products through beta-scission of intermediate alkyl hypochlorites. Secondary alcohols with R containing three or more carbons also may undergo delta chlorination. PMID- 15651820 TI - Readily synthesized novel fluorescent dipyrrinones. AB - A new, highly fluorescent (phiF up to 0.85) rigid anti-Z-dipyrrinone chromophore has been synthesized in high yield in a one-pot reaction by condensing two monopyrroles in the presence of DBU to form the pyrrolo[3,2-f]indolizine-4,6 dione nucleus. PMID- 15651821 TI - Investigation of the diastereoselectivity of tricyclo(5.2.1.0(2,6))decan-10-ones: controversies and agreements. AB - The diastereoselectivities of tricyclo(5.2.1.0(2,6))decan-10-one and its derivatives are controlled by antiperiplanar and vicinal sigma --> piC=O interactions rather than the hyperconjugation effects as reported previously. PMID- 15651822 TI - Palladium(II) acetate in pyridine as an effective catalyst for highly regioselective hydroselenation of alkynes. AB - A highly regioselective hydroselenation of terminal alkynes with benzeneselenol can be achieved by the combination of palladium acetate and pyridine, providing the corresponding terminal alkenes, (i.e., 2-phenylseleno-1-alkenes) as a sole product. In this hydroselenation, pyridine may act as a suitable ligand for active palladium intermediates. PMID- 15651823 TI - Analogues of key precursors of aspartyl protease inhibitors: synthesis of trifluoromethyl amino epoxides. AB - The synthesis of the title compound is described through original and tailored synthetic protocols. The addition of vinylmagnesium bromide to CF(3)-N-aryl and N alkyl aldimines was efficient and did not require an activating N-substituent. The resultant CF3-allylamines were converted in an efficient and completely stereoselective route to syn CF3-epoxides 3 via formation of bromhydrins 8. The same sequence performed from the aldimine substituted with the methyl ether of the (R)-phenylglycinol provided the homochiral (R,R)-amino epoxide (de >98%). This study has allowed access to the novel racemic and homochiral trifluoromethyl beta-amino epoxides, analogues of key precursors of various HIV protease inhibitors. PMID- 15651824 TI - Requirement for an oxidant in Pd/Cu co-catalyzed terminal alkyne homocoupling to give symmetrical 1,4-disubstituted 1,3-diynes. AB - Palladium-catalyzed terminal alkyne dimerization, through oxidative homocoupling, is a useful approach to the synthesis of symmetrical 1,4-diynes. Recent investigations have suggested that this reaction might be accomplished in the absence of intentionally added stoichiometric oxidants (to reoxidize Pd(0) to Pd(II)). In this paper, we have fully addressed the question of whether oxygen (or added oxidant) is required to facilitate this process. The presence of a stoichiometric quantity of air (or added oxidant such as I2) is essential for alkyne dimerization. Excess PPh3 inhibits alkyne dimerization to enyne, which only occurs to a significant extent when the reaction is starved of oxidant. Theoretical studies shed more light on the requirement for an oxidant in the homocoupling reaction in order for the process to be theromodynamically favorable. The employment of I2 as the stoichiometric oxidant appears to be the method of choice. The dual role of Cu both in transmetalation of alkynyl units to Pd(II) and in assisting reoxidation of Pd(0) to Pd(II) is suggested. PMID- 15651825 TI - Ruthenium(III) chloride catalyzed oxidation of pyrene and 2,7-disubstitued pyrenes: an efficient, one-step synthesis of pyrene-4,5-diones and pyrene 4,5,9,10-tetraones. AB - Pyrene and 2,7-disubstituted pyrenes have been oxidized with ruthenium(III) chloride (RuCl3) and sodium periodate (NaIO4) under very mild conditions to 4,5 diones or 4,5,9,10-tetraones. Thus, the oxidation has been controlled by varying the amount of oxidant and reaction temperature to proceed exclusively at the pyrene 4- and 5-positions or at the 4-, 5-, 9-, and 10-positions. PMID- 15651826 TI - Highly regioselective coupling reactions of allylic and propargylic alcohol derivatives with gamma,gamma-dialkoxyallylic zirconium species via Zr-to-Cu transmetalation. AB - In the presence of CuCN, reaction of gamma,gamma-dialkoxyallylic zirconium species 4, generated in situ by treating triethyl orthoacrylate with zirconocene butene complex, with allylic and propargylic phosphates proceeded at the alpha position of 4 in a highly SN2'-selective manner to give the corresponding 5 alkenoates and 4,5-alkadienoates, respectively. In the present Cu(I)-mediated coupling reaction, the gamma,gamma-dialkoxyallylic zirconium species 4 serves as a synthetically useful homoenolate anion equivalent of propionate. PMID- 15651827 TI - Stereoselective synthesis of the naturally occurring styryllactones (+) goniofufurone and (+)-cardiobutanolide. AB - The naturally occurring gamma-lactones (+)-goniofufurone 1 and (+) cardiobutanolide 2, two pharmacologically active products from Goniothalamus species (Annonaceae), have been synthesized in enantiopure form using l erythrulose as the chiral starting material. Key steps of these syntheses were a stereoselective anti boron aldol reaction and an asymmetric allylboration. PMID- 15651829 TI - Tetrahydropyran formation by rearrangement of an epoxy ester: a model for the biosynthesis of marine polyether toxins. AB - Acid-catalyzed rearrangement of the (S)-epoxide derived from 2alpha-allyl cholestanyl acetate resulted in a 1:1 mixture of a steroidal tetrahydrofuran and a steroidal tetrahydropyran. Formation of a six-membered ring supports the hypothesis that epoxy ester-orthester-cyclic ether rearrangement may be involved in the biosynthesis of ladder-type marine polyether toxins. This reaction represents a new biomimetic preparation of medium ring cyclic ethers. PMID- 15651828 TI - Preparation of 8-(arylsulfanyl)adenines with diazonium salts under mild, aerobic conditions. AB - 8-(Arylsulfanyl)adenines 11 were prepared in up to 75% yield by reacting the 8 thionoadenine 6 (acetic acid 3-(6-amino-8-thioxo-7,8-dihydropurin-9-yl)propyl ester) with benzenediazonium tetrafluoroborates in DMSO. Benzenediazonium ions carrying an electron-withdrawing substituent gave the highest yields. The reaction proceeded smoothly at room temperature without any base and could be performed under air atmosphere. The extremely mild conditions are compatible with a wide range of functional groups. PMID- 15651830 TI - The addition of sp2- and sp-hybridized nucleophiles to a bridgehead bromoketone. AB - The reaction of various sp2- and sp-hybridized organolithium species with bromoketone 1 is presented. In most cases, control over the course of the process is possible and products from simple addition, addition followed by a quasi Favorskii rearrangement or addition, rearrangement, and addition can be selectively prepared. PMID- 15651831 TI - Highly efficient catalytic aerobic oxidations of benzylic alcohols in water. AB - A highly efficient catalytic system without transition metals in water has been developed for aerobic oxidations of benzylic alcohols. The newly developed catalyst system could oxidize benzylic alcohols and heteroaromatic analogues with 1 mol % TEMPO as a catalyst and with a catalytic amount of 1,3-dibromo-5,5 dimethylhydantoin and NaNO2 as cocatalysts. Under the optimal conditions, various alcohols could be converted into their corresponding aldehydes or ketones in high yields. PMID- 15651833 TI - Total stereochemical control in the addition of thiols to p toluenesulfonylacetylene. synthesis of Z- and E-2-sulfanylvinylsulfonyl derivatives. AB - Alkyl and aryl sulfides react with equimolecular amounts of p toluenesulfonylacetylene in CH3CN at 0 degrees C or rt without the use of any catalytic reagent to give good yields of Z-2-sulfanylvinylsulfonyl derivatives with total diastereoselectivity. On the other hand, in the presence of 1.1 equiv of NaH in THF, the same reaction affords the corresponding E-diastereomer also with total diastereoselectivity. PMID- 15651832 TI - Second-generation, highly abbreviated route for elaboration of the oxetane D-ring in a fully functionalized taxane. AB - A six-step conversion of oxirane 3 to oxetane 9 is reported. The synthetic route takes particular advantage of the acid-catalyzed ring opening of 3 to allyl alcohol 4 in a polar reaction medium and of the heightened capability of the OsO4.TMEDA complex to effect the efficient stereocontrolled dihydroxylation of this intermediate. The overall yield of the new sequence is 33%. PMID- 15651834 TI - A succinct method for preparing the Stork-Jung vinylsilane robinson annulation reagent. AB - The Stork-Jung vinylsilane reagent (1) is prepared in two steps and in good overall yield. This provides rapid and efficient access to a useful methyl vinyl ketone surrogate. PMID- 15651835 TI - Enantiodivergent formal synthesis of (+)- and (-)-cyclophellitol from D-xylose based on the latent symmetry concept. AB - Formal synthesis of (+)- and (-)-cyclophellitol from d-xylose has been accomplished through utilization of the latent plane of chirality present in the starting carbohydrate. The synthetic pathway is suitable for preparation and biological evaluation of cyclophellitol analogues in both enantiomeric series. PMID- 15651836 TI - The diastereoselective synthesis of quaternary substituted thioindolines from sulfur ylide intermediates. AB - The Rh(II)-catalyzed coupling of chiral 2-thiopyranylindoles with vinyl diazoacetates results in the generation of indolines having quaternary substitution at C3 in high diastereoselectivity. PMID- 15651837 TI - Syntheses of hexakis(4-functionalized-phenyl)benzenes and hexakis[4-(4' functionalized- phenylethynyl)phenyl]benzenes directed to host molecules for guest-inclusion networks. AB - The syntheses of various types of hexakis(4-functionalized-phenyl)benzenes 1 and hexakis[4-(4'-functionalized-phenylethynyl)phenyl]benzenes 2 by the cobalt catalyzed cyclotrimerization of diarylacetylenes and by the Sonogashira coupling reaction of 1e with arylacetylenes, respectively, are described. X-ray crystallographic analysis showed that host 1e or 2f forms a 2-D network by unique I...I or CH...O=C interactions, respectively. PMID- 15651838 TI - Efficient synthesis of the glucosidase inhibitor blintol, the selenium analogue of the naturally occurring glycosidase inhibitor salacinol. AB - An efficient synthesis of blintol, the selenium congener of the naturally occurring glycosidase inhibitor salacinol, and a potent glucosidase inhibitor itself, is described. Unlike our previously reported synthesis, this improved route makes use of p-methoxybenzyl ether protecting groups in the synthesis of one of the two key intermediates, 2,3,5-tri-O-p-methoxybenzyl-1,4-anhydro-4 seleno-D-arabinitol, from L-xylose. The other key intermediate, 2,4-O-benzylidene L-erythritol-1,3-cyclic sulfate, was successfully prepared from D-glucose instead of the expensive L-glucose. All protecting groups in the resulting adducts were removed with trifluoroacetic acid to yield a mixture of stereoisomers, thereby obviating the problematic deprotection of benzyl ethers by hydrogenolysis. The major stereoisomer, blintol, was then obtained by fractional crystallization. PMID- 15651839 TI - Copper-catalyzed allylations of zirconocene intermediates generated from o alkenyl or o-alkynylbenzyl ether derivatives and zirconocene-butene complex. AB - o-Alkenyl or alkynyl benzylzirconocene intermediate, which was readily generated by the reaction of 2-alkoxymethylstyrene or 2-alkoxymethyl-1 (trimethylsilylethynyl)-benzene derivative with zirconocene-butene complex (Negishi reagent, "Cp2Zr"), reacted with allyl or propargyl halides in the presence of a catalytic amount of CuCl to give allylation or allenylation products. Conversion to Dane's diene, which is a key intermediate for estrone synthesis, was efficiently carried out by enyne olefin metathesis of the allylation products. PMID- 15651842 TI - Importance of chromium-DNA adducts in mutagenicity and toxicity of chromium(VI). PMID- 15651843 TI - Urea lesion formation in DNA as a consequence of 7,8-dihydro-8-oxoguanine oxidation and hydrolysis provides a potent source of point mutations. AB - The DNA oxidation product 7,8-dihydro-8-oxoguanine (8-oxoG) forms several mutagenic oxidation products, including a metastable oxaluric acid (Oa) derivative. We report here that a synthetic oligonucleotide containing Oa hydrolyzes under simulated "in vivo" conditions to form a mutagenic urea (Ua) lesion. Using the Oa 2'-deoxyribonucleoside as a model, the hydrolysis rate depended strongly upon the concentrations of bicarbonate and divalent magnesium. In buffered solutions containing physiologically relevant levels of these species, the half-life of Oa nucleoside was approximately 40 h at 37 degrees C. The mutagenic properties of Ua in DNA were investigated using a M13mp7L2 bacteriophage genome containing Ua at a specific site. Transfection of the lesion containing genome into wild-type AB1157 Escherichia coli allowed determination of the mutation frequency and DNA polymerase bypass efficiency from the resulting progeny phage. Ua was bypassed with an efficiency of 11% as compared to a guanine control and caused a 99% G-->T mutation frequency, assuming the lesion originated from G, which is at least an order of magnitude higher than the mutation frequency of 8-oxoG under the same conditions. SOS induction of bypass DNA polymerase(s) in the bacteria prior to transfection caused the mutation frequency and type to shift to 43% G-->T, 46% G-->C, and 10% G-->A mutations. We suggest that Ua is instructional, meaning that the shape of the lesion and its interactions with DNA polymerases influence which nucleotide is inserted opposite the lesion during replication and that the instructional nature of the lesion is modulated by the size of the binding pocket of the DNA polymerase. Replication past Ua, when formed by hydrolysis of the 8-oxoG oxidation product Oa, denotes a pathway that nearly quantitatively generates point mutations in vivo. PMID- 15651844 TI - Subway particles are more genotoxic than street particles and induce oxidative stress in cultured human lung cells. AB - Epidemiological studies have shown an association between airborne particles and a wide range of adverse health effects. The mechanisms behind these effects include oxidative stress and inflammation. Even though traffic gives rise to high levels of particles in the urban air, people are exposed to even higher levels in the subway. However, there is a lack of knowledge regarding how particles from different urban subenvironments differ in toxicity. The main aim of the present study was to compare the ability of particles from a subway station and a nearby very busy urban street, respectively, to damage DNA and to induce oxidative stress. Cultured human lung cells (A549) were exposed to particles, DNA damage was analyzed using single cell gel electrophoresis (the comet assay), and the ability to induce oxidative stress was measured as 8-oxo-7,8-dihydro-2' deoxyguanosine (8-oxodG) formation in lung cell DNA. We found that the subway particles were approximately eight times more genotoxic and four times more likely to cause oxidative stress in the lung cells. When the particles, water extracts from the particles, or particles treated with the metal chelator deferoxamine mesylate were incubated with 2'-deoxyguanosine (dG) and 8-oxodG was analyzed, we found that the oxidative capacity of the subway particles was due to redox active solid metals. Furthermore, analysis of the atomic composition showed that the subway particles to a dominating degree (atomic %) consisted of iron, mainly in the form of magnetite (Fe3O4). By using electron microscopy, the interaction between the particles and the lung cells was shown. The in vitro reactivity of the subway particles in combination with the high particle levels in subway systems give cause of concern due to the high number of people that are exposed to subway particles on a daily basis. To what extent the subway particles cause health effects in humans needs to be further evaluated. PMID- 15651845 TI - Glutathione depletion enhances the formation of endogenous cyclic DNA adducts derived from t-4-hydroxy-2-nonenal in rat liver. AB - Earlier, we detected the cyclic adducts of deoxyguanosine (dG) derived from t-4 hydroxy-2-nonenal (HNE), a long chain alpha,beta-unsaturated aldehyde (enal) product from oxidation of omega-6 polyunsaturated fatty acids, in tissue DNA of rats and humans as endogenous DNA damage. Recent evidence implicates the cyclic HNE adducts in human liver carcinogenesis. Because glutathione (GSH) protects against oxidative stress, we undertook a study to examine the effect of GSH depletion on the HNE-derived cyclic adducts in vivo. Four F344 rats were administered L-buthionine-(S,R)-sulfoximine (BSO), a potent inhibitor of GSH biosynthesis, at 10 mM in drinking water for 2 weeks. Rats in the control group were given water only. Livers were harvested, and each liver was divided into portions for GSH and DNA adduct analyses. The BSO treatment depleted hepatic GSH by 77%; the GSH levels were reduced from 6.3 +/- 0.3 in the control rats to 1.5 +/- 0.1 micromol/g tissues in the treated group. The formation of HNE-dG adducts, analyzed by an HPLC-based 32P-postlabeling assay, was increased by 4-fold, from 6.2 +/- 2.2 nmol/mol dG in liver DNA of control rats to 28.5 +/- 16.1 nmol/mol dG in the rats treated with BSO (p <0.05). The formation of 8-oxodG in liver DNA was also increased as a result of BSO treatment, although the increase was not statistically significant. These results further support the endogenous origin of HNE-dG adducts and, more importantly, indicate a critical role that GSH plays in protecting against in vivo formation of the promutagenic cyclic DNA adducts derived from HNE. PMID- 15651846 TI - Genomic analysis of rodent pulmonary tissue following bis-(2-chloroethyl) sulfide exposure. AB - Bis-(2-chloroethyl) sulfide (sulfur mustard, SM) is a carcinogenic alkylating agent that has been utilized as a chemical warfare agent. To understand the mechanism of SM-induced lung injury, we analyzed global changes in gene expression in a rat lung SM exposure model. Rats were injected in the femoral vein with liquid SM, which circulates directly to the pulmonary vein and then to the lung. Rats were exposed to 1, 3, or 6 mg/kg of SM, and lungs were harvested at 0.5, 1, 3, 6, and 24 h postinjection. Three biological replicates were used for each time point and dose tested. RNA was extracted from the lungs and used as the starting material for the probing of replicate oligonucleotide microarrays. The gene expression data were analyzed using principal component analysis and two way analysis of variance to identify the genes most significantly changed across time and dose. These genes were ranked by p value and categorized based on molecular function and biological process. Computer-based data mining algorithms revealed several biological processes affected by SM exposure, including protein catabolism, apoptosis, and glycolysis. Several genes that are significantly upregulated in a dose-dependent fashion have been reported as p53 responsive genes, suggesting that cell cycle regulation and p53 activation are involved in the response to SM exposure in the lung. Thus, SM exposure induces transcriptional changes that reveal the cellular response to this potent alkylating agent. PMID- 15651847 TI - Dissecting reactivity of clerocidin toward common buffer systems by means of selected drug analogues. AB - The model drug clerocidin (CL) can form covalent adducts with both Tris and phosphate buffers with negative effects on biological activity, even though the latter is considered a largely inert physiological buffer. With the ultimate goal of learning how to control such reactivity and reduce undesired side reactions, we have investigated the influence of the different functionalities of CL on the formation of buffer adducts. For this reason, selected drug analogues were tested for their ability to react with the two buffers and comprehensive information was gained on both thermodynamics and kinetics aspects of these reactions. Two distinctive reactivity modes were readily observed. The first proved to be under kinetic control and involved the reaction of drug carbonyls (especially the aldehyde in C15) with the Tris amino group to form a Schiff base. The second was found to proceed under thermodynamic control through the attack at the oxirane ring of CL by the buffer's nucleophilic groups (amino nitrogen in Tris and oxygen in phosphate). Important relationships between the two modes were noted, thus providing further demonstration that drug reactivity toward buffers cannot be directly predicted from the functionalities that are potentially involved in the initial reaction. On the contrary, as true for almost any structure bearing potentially reactive functionalities, any solid prediction should be based on a deeper understanding of the mutual influence of vicinal groups. PMID- 15651848 TI - Alkylation of cytochrome c by (glutathion-S-yl)-1,4-benzoquinone and iodoacetamide demonstrates compound-dependent site specificity. AB - The reaction of cytochrome c with the electrophilic compounds (glutathion-S-yl) 1,4-benzoquinone (GSBQ) and iodoacetamide was studied using mass spectrometry. GSBQ is a nephrotoxic quinol-thioether metabolite of benzoquinone, while iodoacetamide is an alkylating agent targeting cysteine thiols. Both chemicals formed covalent adducts with cytochrome c. GSBQ formed adducts with cytochrome c at pH 6 on several histidine and lysine residues. At a pH >7, the initial product rearranged to a disubstituted cyclic quinone species preferentially found at two sites on the protein, Lys25-Lys27 and Lys86-Lys87, via quinol amine linkages. These two sites were previously determined to be the targets of benzoquinone adduct formation [Person et al. (2003) Chem. Res. Toxicol. 16, 598-608]. Cyclic reaction products are preferentially formed at two sites on the protein because of the presence of multiple basic residues in a conformationally flexible region whereas noncyclic products bind to a broad spectrum of available lysine and histidine nucleophiles. Iodoacetamide was a less selective alkylating agent able to form adducts on the majority of the nucleophilic sites of the protein. MS/MS spectra were used to identify signature ions for GSBQ-adducted peptides from the characteristic fragmentation patterns. Neutral losses of the 129 Da gamma glutamate residue and of the 273 Da glutathione moiety were found in both cysteine thiol- and lysine amine-linked GSBQ adduct MS/MS. Characteristic fragment ions were used in conjunction with the scoring algorithm for spectral analysis to search for adducted species present at low levels in the sample, and the analysis is applicable generally to detection of glutathione conjugates by MS/MS. Parallel analysis using matrix-assisted laser desorption/ionization-MS to compare spectra of control and treated samples allowed identification of peptide adducts formed by direct addition of GSBQ and by the subsequent loss of the glutathione moiety in a pH-dependent cyclization reaction. PMID- 15651849 TI - Analysis of M1G-dR in DNA by aldehyde reactive probe labeling and liquid chromatography tandem mass spectrometry. AB - A novel method for the measurement of pyrimido[1,2-a]purin-10(3H)one (M1G) has been developed. Previous methods for analysis of M1G have been confounded by the fact that this lesion exists in equilibrium between a ring-closed form and a ring opened aldehyde form. Poor detection sensitivity of the aldehydic form can result from loss of the adduct during analysis by its reaction with amines or proteins. We utilized the aldehyde reactive probe (ARP) to produce a stable ARP-M1G deoxyribose (ARP-M1G-dR) conjugate to minimize adduct loss. This conjugate has increased the hydrophobicity that enhances separation of ARP-M1G-dR from unmodified DNA nucleosides by using solid phase extraction. In addition, measuring ARP-M1G-dR by selective reaction monitoring (SRM) of the [ARP-M1G-dR + H]+ (635) --> [M1G + H]+ (188) transition increases the detection sensitivity by nearly an order of magnitude relative to the measurement of M1G-dR by SRM. For accurate measurement, analytical standard (AS) DNA and internal standard (IS) DNA were used. High purity 15N-labeled DNA was isolated from Escherichia coli that had been grown in minimum salt medium containing (15NH4)2SO4. The 15N-DNA and calf thymus DNA were treated with malondialdehyde to induce a high number of M1G adducts to prepare the IS and AS DNA, respectively. A consistent calibration curve was established from the analysis of 200 microg of blank DNA, 23 ng of IS DNA (400 fmol of 15N5-M1G-dR), and AS DNA containing 0-810 fmol of M1G-dR. With the use of this novel IS DNA and selective labeling, this assay is a useful tool for monitoring oxidative stress-induced DNA damage from small amounts of DNA without the need of a specific antibody or laborious procedures. By this assay, two M1G adducts/10(8) guanines can readily be detected. Furthermore, this approach should be applicable to the analysis of other aldehydic DNA adducts as well as the measurement of an array of DNA lesions. PMID- 15651850 TI - Cytochrome P450 2A-catalyzed metabolic activation of structurally similar carcinogenic nitrosamines: N'-nitrosonornicotine enantiomers, N nitrosopiperidine, and N-nitrosopyrrolidine. AB - N'-Nitrosonornicotine (NNN) and N-nitrosopiperidine (NPIP) are potent esophageal and nasal cavity carcinogens in rats and pulmonary carcinogens in mice. N Nitrosopyrrolidine (NPYR) induces mainly liver tumors in rats and is a weak pulmonary carcinogen in mice. These nitrosamines may be causative agents in human cancer. alpha-Hydroxylation is believed to be the key activation pathway in their carcinogenesis. P450 2As are important enzymes of nitrosamine alpha hydroxylation. Therefore, a structure-activity relationship study of rat P450 2A3, mouse P450 2A4 and 2A5, and human P450 2A6 and 2A13 was undertaken to compare the catalytic activities of these enzymes for alpha-hydroxylation of (R) NNN, (S)-NNN, NPIP, and NPYR. Kinetic parameters differed significantly among the P450 2As although their amino acid sequence identities were 83% or greater. For NNN, alpha-hydroxylation can occur at the 2'- or 5'-carbon. P450 2As catalyzed 5' hydroxylation of (R)- or (S)-NNN with Km values of 0.74-69 microM. All of the P450 2As except P450 2A6 catalyzed (R)-NNN 2'-hydroxylation with Km values of 0.73-66 microM. (S)-NNN 2'-hydroxylation was not observed. Although P450 2A4 and 2A5 differ by only 11 amino acids, they were the least and most efficient catalysts of NNN 5'-hydroxylation, respectively. The catalytic efficiencies (kcat/Km) for (R)-NNN differed by 170-fold whereas there was a 46-fold difference for (S)-NNN. In general, P450 2As catalyzed (R)- and (S)-NNN 5'-hydroxylation with significantly lower Km and higher kcat/Km values than NPIP or NPYR alpha hydroxylation (p <0.05). Furthermore, P450 2As were better catalysts of NPIP alpha-hydroxylation than NPYR. P450 2A4, 2A5, 2A6, and 2A13 exhibited significantly lower Km and higher kcat/Km values for NPIP than NPYR alpha hydroxylation (p <0.05), similar to previous reports with P450 2A3. Taken together, these data indicate that critical P450 2A residues determine the catalytic activities of NNN, NPIP, and NPYR alpha-hydroxylation. PMID- 15651851 TI - Measurement of N7-(2'-hydroxyethyl)guanine in human DNA by gas chromatography electron capture mass spectrometry. AB - An improved method is presented, based on gas chromatography-electron capture mass spectrometry (GC-EC-MS), for measuring N7-(2'-hydroxyethyl)guanine (N7-HEG) in DNA from an in vivo sample. The method was used to detect this adduct in amounts of human DNA ranging from 0.07 to 11.5 microg isolated from granulocytes. In this method, the DNA is spiked with a stable isotope internal standard (N7-HEG d4) and heated in water to release the adduct in a nucleobase form. After the adduct is extracted into 1-butanol, it is purified by reverse phase HPLC and derivatized with HONO, pentafluorobenzyl bromide, and pivalic anhydride. Further purification by silica solid phase extraction and reverse phase HPLC is done prior to injection into a GC-EC-MS. Relatively clean GC-EC-MS chromatograms result, contributing to the high sensitivity that is observed. In the samples tested, from 1.6 to 240 N7-HEG adducts in 10(7) nucleotides were observed, a 150 fold range. PMID- 15651852 TI - Effects of peroxynitrite dose and dose rate on DNA damage and mutation in the supF shuttle vector. AB - Peroxynitrite (ONOO-) induces oxidative and nitrosative DNA damage, and previous studies by our group have shown that it is strongly mutagenic in the supF shuttle vector pSP189 replicated in Escherichia coli MBL50 cells. In those experiments, however, the pSP189 plasmid was exposed under unphysiological conditions to large single bolus doses of ONOO-, which limits extrapolation of the data to in vivo pathological states in which ONOO- may play a role. We have thus sought to define the effects of ONOO- dose and dose rate on the DNA damage and mutations induced in the supF gene by three different dosage mechanisms: (i) by infusion of ONOO- solution into suspensions of pSP189 at rates approximating those estimated to occur in inflamed tissues; (ii) by exposure to 3-morpholinosydnonimine (SIN-1), which generates ONOO- spontaneously during decomposition; and (iii) by bolus doses of ONOO- solution. In all cases, plasmid DNA was exposed in the presence of 25 mM bicarbonate, since the reaction of CO2 with ONOO- (to form nitrosoperoxycarbonate) has a major impact on mutagenic potency of ONOO- in this system. Nucleobase and deoxyribose damage were evaluated by a plasmid nicking assay immediately after ONOO- and SIN-1 exposures. Mutation frequency (MF) and mutational spectra in the supF gene were determined after plasmid pSP189 replicated in host E. coli cells. Bolus ONOO- addition caused the highest amount of DNA damage, including base and deoxyribose lesions, while infusion caused the least. SIN-1 was found to induce almost exclusively deoxyribose oxidation, while bolus addition generated a high percentage of base damage. MF increased in a dose dependent manner following all treatments, but infused ONOO- and SIN-1 exposures were less mutagenic than bolus ONOO- exposure. MFs induced by infusion and by SIN 1 incubated for 100 min at the highest level (4 mM) were 63 and 43% less, respectively, than that induced by bolus. All mutational hot spots were located at G:C sites except for A121 and A177 induced by SIN-1 exposure. Hot spots at C108 and C168 were common to all exposures; G113, G115, and G116 were common to bolus and infused ONOO- exposures; and G129 was common to infused ONOO- and SIN-1 exposures. Almost all mutations were single base pair substitutions under all exposure conditions. Whereas those induced by infused or bolus ONOO- and SIN-1 consisted predominantly of G:C to T:A transversions (66, 65, and 51%, respectively), G:C to C:G mutations were much less frequent following infusion and SIN-1 (8 and 19%, respectively) than those induced by bolus exposure (29%). A:T to T:A mutations induced were detected only after ONOO- infusion and SIN-1 exposure (9 and 11%, respectively). In conclusion, both dose and dose rate at which a genetic target is exposed to ONOO- substantially influence the damage and mutational response, indicating that these parameters will need to be taken into account in assessing the potential effects of ONOO- in vivo. Furthermore, the results indicate that the chemistry of SIN-1-induced DNA damage differs substantially from native ONOO-, which suggests the need for caution in interpreting the biological relevance of SIN-1 as a surrogate for ONOO-. PMID- 15651854 TI - Neutral metallomacrocycles with four or ten (PEt3)Pd(II) centers. AB - The reaction of [(Et(3)P)PdCl(2)](2) with 2,3-dihydroxypyridine, 5-chloro-2,3, dihydroxypyridine, or 2-hydroxynicotinic acid in the presence of base gives neutral, metallomacrocyclic compounds in which the heterocycles act as dianionic, bridging ligands. The macrocycles contain four or ten palladium atoms as evidenced by single crystal X-ray analyses. PMID- 15651853 TI - Effects of substrate specificity on initiating the base excision repair of N methylpurines by variant human 3-methyladenine DNA glycosylases. AB - The human 3-methyladenine (AAG, ANPG, MPG) DNA glycosylase excises alkylated purines from DNA. In previous studies, we determined the importance of an active site amino acid (asparagine 169) in the recognition of substrates by AAG. In this study, we characterize the consequences of expressing the AAG variants bearing amino acid substitutions at position 169 in Saccharomyces cerevisiae that lack endogenous 3-methyladenine DNA glycosylase. Survival, mutation induction, and DNA double strand break formation were determined in response to methyl methanesulfonate. The ability of purified wild-type and AAG variants to remove 3 methyladenine and 7-methylguanine, the two most abundant adducts produced by methyl methanesulfonate, was also determined. The N169D AAG variant displayed a approximately 100-fold lower activity for 3-methyladenine as compared to wild type and did not detectably remove 7-methylguanine. When expressed in S. cerevisiae, the N169D variant provided better protection against methyl methanesulfonate toxicity than wild-type. Fewer strand breaks in vivo were also seen in the presence of the N169D variant following MMS exposure. In contrast, the N169A and N169S AAG variants displayed approximately 30-fold lower activity for 3-methyladenine and 7-methylguanine. Expression of the N169A and N169S AAG variants in S. cerevisiae during methyl methanesulfonate exposure resulted in greater sensitivity, greater mutation induction following MMS exposure, and more strand breaks in vivo. Strand breaks seen in S. cerevisiae that express wild-type AAG or the N169 variants were resolved to varying extents during recovery. In contrast, strand breaks formed in S. cerevisiae that expressed a catalytically inactive AAG variant were not resolved during the recovery times examined. Taken together, the results provide evidence that 3-methyladenine adducts not repaired by base excision repair cause double strand breaks that are not rapidly resolved. Evidence is also provided that the BER intermediates resulting from excision of 7 methylguanine by wild-type AAG contributes to the mutagenicity and cytotoxicity of alkylating agents. PMID- 15651855 TI - Insertion of Ga(DDP) into the Au-Cl bond of (PPh3)AuCl: a first structurally characterized Au-Ga bond. AB - The reaction of (PPh(3))AuCl with the low valent gallium compound Ga(DDP) (DDP = 2-((2,6-diisopropylphenyl)amino-4-((2,6-diiso-propylphenyl)imino)-2-pentene) yields the insertion products [{Ga(DDP)}Au{Ga(DDP)Cl}] (1) and [(PPh(3))Au{Ga(DDP)Cl}] (2), the first examples of molecular compounds with Au-Ga bonds. PMID- 15651856 TI - Synthesis and X-ray structures of dinuclear and trinuclear gold(I) and dinuclear gold(II) amidinate complexes. AB - The structures of the trinuclear gold(I), [Au(3)(2,6-Me(2)-form)(2)-(THT)Cl], the dinuclear [Au(2)(2,6-Me(2)-form)(2)], and the oxidative-addition product [Au(2)(2,6-Me(2)-form)(2)Cl(2)] formamidinate complexes are reported. The trinuclear complex is stable with gold-gold distances 3.01 and 3.55 A. The gold gold distance in the dinuclear complex decreases upon oxidative-addition with halogens from 2.7 to 2.5 A, similar to observations made with the dithiolates and ylides. PMID- 15651857 TI - Confirmation of the semivacant Wells-Dawson polyoxotungstate skeleton. The structures of [Ce{X(H4)W17O61}2]19- (X = P, As) indicate the probable location of internal protons. AB - Reaction of Ce(III) with lacunary versions of [H(4)XW(18)O(62)](7-) (X = P, As) yields the 1:2 complexes [Ce(H(4)XW(17)O(61)](19-) (X = As, 1; P, 2) in good yield, characterized in solution and the solid state by NMR spectroscopy and X ray crystallographic analysis, respectively. The structures confirm a syn C(2) conformation that is analogous to that observed for [Ln(alpha(2) P(2)W(17)O(61))(2)](17-) but with "empty" O(4) tetrahedra that are in positions remote from the cerium atom. Bond valence sum calculations for these structures show that the four protons that are required for charge balance in all salts of the XW(18) anions and their lacunary derivatives are almost certainly bound to the oxygen atoms of the empty tetrahedra. PMID- 15651858 TI - Bis(1,2,3-trimethylindenyl)iron(III) 2,3-dicyanonaphtho-1,4-quinonide, a non metallocene, charge-transfer salt metamagnet with complementary donor-acceptor geometries. AB - Bis(1,2,3-trimethylindenyl)iron(III) can be paired with 2,3-dicyanonaphtho-1,4 quinone to give an air-stable pi-stacked metamagnet with T(c) = 4.1 K, the first example of a non-metallocene CT salt magnet in this class. A single crystal X-ray structure of the Fe(1,2,3-Me(3)C(9)H(4))(2)[DCNQ] salt indicates that it consists of stacks of alternating donors and acceptors, in which the DCNQ ring systems are aligned with indenyl groups above and below, and the long axes of the DCNQ and the indenyl ligand are parallel to each other. This arrangement suggests that the magnetic properties arise from favorable interactions due to geometric similarities between the donor and acceptor, and not from a unique property of the donor itself. PMID- 15651859 TI - Synthesis of a vanadium(III) tris(arylthiolato) complex and its reactions with azide and azo compounds: formation of a sulfenamide complex via cleavage of an azo N=N bond. AB - The tris(arylthiolate) vanadium(III) complex (1) has been synthesized in good yield. This complex is found to undergo CH activation across a V-S bond in the presence of TMEDA to give a cyclometalated species along with free arylthiol. Complex 1 behaves as a two-electron reductant toward Ad-N(3), yielding an imide complex. Treatment of 1 with azobenzene produces an imide-sulfenamide compound, in which an azo N=N bond cleavage takes place concomitant with formation of a V=N and an S-N bond. PMID- 15651860 TI - Engineering of a novel ruthenium sensitizer and its application in dye-sensitized solar cells for conversion of sunlight into electricity. AB - A novel ligand 4,4'-bis(carboxyvinyl)-2,2'-bipyridine (L) and its ruthenium(II) complex [Ru(II)L(2)(NCS)(2)] (K8) were synthesized and characterized by analytical, spectroscopic, and electrochemical techniques. The performance of the K8 complex as a charge transfer photosensitizer in nanocrystalline TiO(2) based solar cells was studied. When the K8 complex anchored onto a nanocrystalline TiO(2) film, we achieved very efficient sensitization yielding 77 +/-5% incident photon-to-current efficiencies (IPCE) in the visible region using an electrolyte consisting of 0.6 M methyl-N-butyl imidiazolium iodide, 0.05 M iodine, 0.05 M LiI, and 0.5 M 4-tert-butylpyridine in a 50/50 (v/v) mixture of valeronitrile and acetonitrile. Under standard AM 1.5 sunlight, the complex K8 gave a short circuit photocurrent density of 18 +/- 0.5 mA/cm(2), and the open circuit voltage was 640 +/- 50 mV with fill factor of 0.75 +/- 0.05, corresponding to an overall conversion efficiency of 8.64 +/- 0.5%. PMID- 15651861 TI - Transformation of a metal-organic framework from the NbO to PtS net. AB - Two metal-organic frameworks (MOFs), MOF-501 and MOF-502, respectively, formulated as Co(2)(BPTC)(H(2)O)(5).G(x) and Co(2)(BPTC)(H(2)O)(DMF)(2).G(x) (BPTC = 3,3',5,5'-biphenyltetracarboxylate; G = guest molecules), have been synthesized and structurally characterized, and their topologies were found to be based on the NbO (MOF-501) and PtS (MOF-502) nets. Heating MOF-501 in solution results in the more thermodynamically favored MOF-502. PMID- 15651862 TI - Incorporating transition metal complexes into tetrathioarsenates(V): syntheses, structures, and properties of two unprecedented [Mn(dien)2]n[Mn(dien)AsS4]2n.4nH2O and [Mn(en)3]2[Mn(en)2AsS4][As3S6]. AB - Two transition-metal tetrathioarsenate complexes, [Mn(dien)(2)](n)[Mn(dien)AsS(4)](2n).4nH(2)O (1) with one-dimensional water chain and [Mn(en)(3)](2)[Mn(en)(2)AsS(4)][As(3)S(6)] (2) with mixed-valence As(3+)/As(5+) character, have been synthesized and structurally characterized. The tetrathioarsenate(V) anion acts as a novel mu(2)-eta(1),eta(2) ligand in 1 and as a chelating ligand in 2. The two compounds exhibit intriguing semiconducting properties (E(g) = 2.18 eV (1), 2.48 eV (2)) and strong photoluminescence with the emission maximum occurring around 440 nm. PMID- 15651863 TI - Synthesis, crystal structure, and high-precision high-frequency and -field electron paramagnetic resonance investigation of a manganese(III) complex: [Mn(dbm)2(py)2](ClO4). AB - The complex [Mn(dbm)(2)(py)(2)](ClO(4)) (dbm = anion of 1,3-diphenyl-1,3 propanedione (dibenzoylmethane), py = pyridine) was synthesized and characterized by X-ray crystallography. It has tetragonally distorted geometry with the axial positions occupied by the py ligands and the equatorial positions by the dbm ligands. This mononuclear complex of high-spin Mn(III) (3d(4), S = 2) was studied by high-frequency and -field electron paramagnetic resonance (HFEPR) both as a solid powder and in frozen dichloromethane solution. Very high quality HFEPR spectra were recorded over a wide range of frequencies. The complete dataset of resonant magnetic fields versus transition energies was analyzed using automated fitting software. This analysis yielded the following spin Hamiltonian parameters (energies in cm(-1)): D = -4.504(2), E = -0.425(1), B(4)(0) = -1.8(4) x 10(-4), B(4)(2) = 7(3) x 10(-4), B(4)(4) = 48(4) x 10(-4), g(x) = 1.993(1), g(y) = 1.994(1), and g(z) = 1.983(1), where the B(4)(n) values represent fourth-order zero-field splitting terms that are generally very difficult to extract, even from single-crystal measurements. The results here demonstrate the applicability of HFEPR at high-precision measurements, even for powder samples. The zero-field splitting parameters determined here for [Mn(dbm)(2)(py)(2)](+) are placed into the context of those determined for other mononuclear complexes of Mn(III). PMID- 15651864 TI - Structural chemistry of A3CuBO6 (A = Ca, Sr; B = Mn, Ru, Ir) as a function of temperature. AB - Variable temperature X-ray and neutron powder diffraction techniques have been used to identify structural phase transitions in Cu-rich A(3)A'BO(6) phases. A transition from monoclinic to rhombohedral symmetry was observed by X-ray diffraction between 700 and 500 K in Sr(3)Cu(1-x)M(x)IrO(6) (M = Ni, Zn; 0 < or = x < or = 0.5). The temperature of the phase change decreased in a linear manner with Cu-content and was essentially independent of the nature of M. Ca(3.1)Cu(0.9)MnO(6) was shown to pass from a rhombohedral phase to a triclinic phase on cooling below 290 K; the structure of the triclinic phase was refined against neutron diffraction data collected at 2 K. Ca(3.1)Cu(0.9)RuO(6) undergoes a transition between a disordered rhombohedral phase and an ordered monoclinic phase when cooled below 623 K. Neutron diffraction has been used to determine the structure as a function of temperature in the range 523 < or =T/K < or = 723 and hence to determine an order parameter for the low temperature phase; the second order transition is shown to be incomplete 100 K below the critical temperature. PMID- 15651865 TI - High-frequency, high-field EPR; magnetic susceptibility; and X-ray studies on a ferromagnetic heterometallic complex of diethanolamine (H2L), [Cu4(NH3)4(HL)4][CdBr4]Br2.3dmf.H2O. AB - The novel heterometallic complex [Cu(4)(NH(3))(4)(HL)(4)][CdBr(4)]Br(2).3dmf.H(2)O has been prepared in the reaction of zerovalent copper with cadmium oxide in the air-exposed solution of ammonium bromide and diethanolamine (H(2)L) in dimethylformamide (dmf). The compound is monoclinic, with space group P2(1)/c, a = 14.876(3) A, b = 33.018(6) A, c = 11.437(2) A, beta = 108.182(3)(o), and Z = 4. The crystal lattice consists of [Cu(4)(NH(3))(4)(HL)(4)](4+) cations, [CdBr(4)](2)(-), Br(-) anions, and uncoordinated dmf and water molecules. In the cation, four independent Cu atoms occupy vertexes of a distorted tetrahedron with bridged Cu...Cu distances in the range 3.127(2)-3.333(3) A and other Cu...Cu separations being 3.445(3)-3.503(2) A. The magnetic susceptibility and the EPR spectra were measured over the temperature ranges 1.8-300 and 3-300 K, respectively. The magnetic moment was found to increase with decreasing temperature to reach a maximum of 2.60 muB per one copper atom at ca. 10 K and was found, subsequently, to diminish slightly at lower temperatures owing to zero-field and Zeeman splitting of the S = 2 ground state. The temperature dependence of the magnetic susceptibility was fitted to the spin Hamiltonian H = J(ab)S(a)S(b) + J(bc)S(b)S(c) + J(cd)S(c)S(d) + J(ad)S(a)S(d) + J(ac)S(a)S(c) + J(bd)S(b)S(d) with the exchange integrals J(ab) = J(bc) = J(cd) = J(ad) = -65(3) cm(-1) and J(ac) = J(bd) = +1(3) cm(-1). High field, high-frequency (95-380 GHz) EPR spectra due to an S = 2 ground state were simulated with g(x) = 2.138(1), g(y)) = 2.142(1), g(z) = 2.067(1), D = -0.3529(3) cm(-1), and E = -0.0469(8) cm(-1). Calculations based on the X-ray structure indicate a negligible contribution of the magnetic dipole-dipole interactions to the zfs parameters D and E. A discussion of the isotropic and anisotropic exchange interactions and their effect on the zfs parameters is also given. PMID- 15651866 TI - Knotted network consisting of 3-threads and a zwitterionic one-dimensional polymorphs of trans-3-(3-pyridyl)acrylate of cobalt and nickel, MII(C8H6NO2)2(H2O)2. AB - We present the hydrothermal synthesis, characterization (IR, DT-TGA), single crystal structures, and magnetic properties of two polymorphs of trans-3(3 pyridyl)acrylate of cobalt(II) and of nickel(II), M(II)(C(8)H(6)NO(2))(2)(H(2)O)(2). Hydrothermal reaction at 120 or 170 degrees C results exclusively in the different polymorphs. The infrared spectra and thermogravimetric analyses of the complexes are almost similar for the two polymorphs but show a difference between cobalt and nickel in energies of the vibrational modes and in the decomposition temperatures. The crystal structures of the two polymorphs are quite different; one crystallizes in a monoclinic space group and the other in a triclinic. This major difference is due to the different stereochemistry, cis or trans, of the coordination at the metal sites. When it is trans-MN(2)O(4), it results in the monoclinic cell consisting of a 3D-network of metals bridged by the ligands through single bonds (M-N and M-O). There is threading of three sublattices up to 2a x 4b x 2c, at which point the three sublattices are knotted into one infinite framework. When it is cis-MN(2)O(4), it results in the triclinic cell and consists of Zwitterionic linear chains of metals bridged by one single ligand via the pyridine and a bidentate carboxylate group and the other ligand is bonded only via the pyridine while its carboxylate end is free. All four compounds are paramagnetic with Weiss constants suggesting weak interactions. PMID- 15651867 TI - Laser photoinitiated nitrosylation of 3-electron reduced Nm europaea hydroxylamine oxidoreductase: kinetic and thermodynamic properties of the nitrosylated enzyme. AB - Hydroxylamine-cytochrome c554 oxidoreductase (HAO) catalyzes the 4-e(-) oxidation of NH(2)OH to NO(2)(-) by cytochrome c554. The electrons are transferred from NH(2)OH to a 5-coordinate heme known as P(460), the active site of HAO. From P(460), c-type hemes transport the electrons through the enzyme to a remote solvent-exposed c-heme, where cyt c554 reduction occurs. When 3-60 microM NO* are photogenerated by laser flash photolysis of N,N'-bis-(carboxymethyl)-N,N' dinitroso-1,4-phenylenediamine, in a solution containing approximately 1 microM HAO prereduced by 3 e(-)/subunit, the HAO c-heme pool is subsequently oxidized by up to 1 e(-)/HAO subunit. The reaction rate for HAO oxidation shows first-order dependence on [HAO], and zero-order dependence on [NO*] (k(obs) = 1250 +/- 150 s( )(1)). However, the total HAO oxidized shows hyperbolic dependence on [NO*]. We suggest that NO* first binds reversibly to P(460) giving a {Fe(NO)}(6) moiety. Intramolecular electron transfer (IET) from the c-heme pool then reduces P(460) to {Fe(NO)}.(7) The overall binding constant (K) for formation of {Fe(NO)}(7) from free NO* and 3-e(-) reduced HAO was measured at (7.7 +/- 0.6) x10(4) M(-1). This value is larger than that for typical ferriheme proteins ( approximately 10(4) M(-1)), but much smaller than that for the corresponding ferroheme proteins ( approximately 10(11) M(-1)). The final product generated by nitrosylating 3-e( ) reduced HAO is believed to be the same species obtained by adding NH(2)OH to the fully oxidized enzyme. The experiments described herein suggest that when NH(2)OH and HAO first react, only two of the NH(2)OH electrons end up in the c heme pool. The other two remain at P(460) as part of an {Fe(NO)}(7) moiety. These results are discussed in relation to earlier studies that investigated the effect of putting fully oxidized and fully reduced HAO under 1 atm of NO*. PMID- 15651868 TI - Novel chiral "calixsalen" macrocycle and chiral robson-type macrocyclic complexes. AB - A family of novel chiral "calixsalen" Schiff base macrocycles R,R-H(3)L4, R,R H(3)L5, containing three chiral diamino moieties were synthesized by an efficient self-assembly and characterized by (1)H and (13)C NMR, mass spectrometry, and X ray diffraction. The systematic synthesis, structure, and coordination properties of the [2 + 2] and [3 + 3] Robson-type Schiff base macrocyclic mono-, di-, tri-, and tetranuclear metal complexes were explored. PMID- 15651869 TI - Synthesis, structure, and properties of [Pt(II)(diimine)(dithiolate)] dyes with 3,3'-, 4,4'-, and 5,5'-disubstituted bipyridyl: applications in dye-sensitized solar cells. AB - A family of [Pt(II)(diimine)(dithiolate)] complexes of general formula [Pt{X,X'(CO(2)R)(2)-2,2'-bipyridyl}(maleonitriledithiolate)] (where X = 3, 4, or 5 and R = H or Et) have been synthesized, spectroscopically and electrochemically characterized, and attached to a TiO(2) substrate to be tested as solar cell sensitizers. A single-crystal X-ray structure showing a large torsion angle between the bipyridyl rings was determined for [Pt{3,3'(CO(2)Et)(2)-2,2' bipyridyl}(maleonitriledithiolate)].MeCN. The effect of changing the position of the bipyridyl substituents from 3,3' to 4,4' and 5,5' is discussed with reference to structural and electronic changes seen within the different members of the family of molecules. The first UV/vis/NIR spectroelectrochemical study of complexes of this general formula is discussed. All three complexes (where R = H) were tested as solar cell sensitizers, with the 3,3'-disubstituted bipyridyl complex giving an intermediate dye loading value but superior photovoltaic performance to those of the other two. The performance of this sensitizer is then compared with that of a well-known Ru polypyridyl sensitizer, the ditetrabutylammonium salt of [RuL(2)(NCS)(2)] (L = 2,2'-bipyridyl-4,4' dicarboxylato), commonly called N719. PMID- 15651870 TI - New titanium and nickel gallophosphates with layered structures. AB - Two new transition-metal gallophosphates, (H(2)C(4)H(10)N(2))(3)[(Ti(2.5)(H(2)O)(4)Ga(5.5)(PO(4))(10)].2H(2)O (TGP-1) and [H(3.5)(C(4)H(13)N(3))(2)][(Ni(0.5)(OH)(4)Ga(5.5)(PO(4))(3)(HPO(4))(4)].2H(2)O (NGP-1), have been synthesized under mild hydrothermal conditions and characterized by single-crystal X-ray diffraction, thermogravimetric analysis, electron paramagnetic resonance, electron probe microanalysis, and magnetic susceptibility data. TGP-1 exhibits a unique two-dimensional structure consisting of tetrahedral and octahedral metals centers and is the foremost paramagnetic TiGaPO material ever prepared. NGP-1 as well represents the first NiGaPO compound and adopts a layer structure that is constructed from hexameric M-O clusters of trigonal bipyramids and octahedra. In both compounds, the transition metals incorporate with gallium into octahedral sites only, while the four- and five coordinated metals centers are only Ga(3+) ions. The unique sites for Ti(3+) and Ni(2+) ions have been initially elucidated from single-crystal structure refinements and further confirmed by bond-valence-sum calculations, EPR, and magnetic susceptibility studies. Crystal data: TGP-1, monoclinic, P2(1)/c; a = 25.692(2) A, b = 9.6552(8) A, c = 9.8418(8) A, beta = 96.737(2)(o) , V = 2424.5(3) A(3), and Z = 2; NGP-1, monoclinic, C2/c, a = 20.8363(12) A, b = 11.9546(7) A, c = 16.4577(9) A, beta = 117.285(1)(o) , V = 3643.3(1) A(3), Z = 4. PMID- 15651871 TI - Toward templated metal-organic frameworks: synthesis, structures, thermal properties, and luminescence of three novel lanthanide-adipate frameworks. AB - Three novel praseodymium-adipate frameworks were synthesized hydrothermally. GWMOF-3 ([Pr(2)(adipic acid)(3)(H(2)O)(4)].adipic acid.4H(2)O) and GWMOF-6 ([Pr(2)(adipic acid)(3)(H(2)O)(2)].4,4'-dipyridyl) formed three-dimensional structures, whereas GWMOF-4 ([Pr(2)(adipic acid)(3)(H(2)O)(2)].H(2)O) produced a more dense, two-dimensional topology. Single-crystal X-ray and powder diffraction, IR spectroscopy, fluorescence spectroscopy, thermogravimetric analysis, and elemental analysis were employed to characterize all samples. GWMOF 6 represents an innovative step forward in metal-organic framework synthesis where a neutral molecular species not used in the construction of the framework is utilized as a structure-directing agent, or template. Furthermore, this template molecule (4,4'-dipyridyl) is shown to sensitize the fluorescence of lanthanide metal centers in a europium analogue of GWMOF-6. PMID- 15651872 TI - Hydrogen bonding and covalent effects in binding of cisplatin to purine bases: ab initio and atoms in molecules studies. AB - Ab initio and density functional calculations are employed to investigate the role of hydrogen bonding in the binding of cisplatin to the purine bases guanine and adenine. Through the use of the theory of atoms in molecules (AIM), it is shown that hydrogen bonds are ubiquitous in such systems, with N-H...N and N H...Cl interactions present in addition to the expected N-H...O. This in turn means that the known stability of cisplatin-guanine complexes cannot be ascribed solely to hydrogen bonding and allows decomposition of total binding energy into contributions from covalent and hydrogen bonds. To do so, a new method for predicting hydrogen bond energies from bond critical point properties is proposed, employing partial least-squares analysis to remove the family dependence of simple models. Still more hydrogen bond motifs are found in bifunctional complexes of the general type purine-[Pt(NH(3))(2)](2+)-purine, including purine...purine contacts, though again the energetics of these are insufficient to explain the observed trends in stability. Finally, the effect of platination on the pairing of guanine with cytosine is studied in a similar manner, revealing large redistributions of hydrogen bonding but surprisingly small overall changes in pairing energy. PMID- 15651873 TI - A spirocyclic system comprising both phosphazane and phosphazene rings. AB - The reaction of hexakis(cyclohexylamino) cyclotriphosphazene [NP(CyNH)(2)](3), 1, with phosphorus trichloride yields [NP(CyN)(2)PCl](3), 2, which contains three four-membered phosphazane rings fused in spirocyclic fashion to a central six membered phosphazene ring and constitutes the first structurally characterized compound that comprises both phosphazene and phosphazane rings. The peripheral P atoms feature stereoactive lone pairs, and, thus, 2 exists in isomeric C(3h) and C(s) forms. The spirocyclic phosphazene-phosphazane derivative 2 carries three reactive PCl functions in peripheral positions, promising an interesting precursor molecule for the synthesis of extended phosphorus nitrogen structures of high rigidity. Extension of the PN moiety can be achieved by reaction of 2 with a primary amine yielding [NP(CyN)(2)PN(H)(t)Bu](3), 3, which features a central scaffold of 6 phosphorus and 12 nitrogen centers and aggregates via N H...P hydrogen bonds in the solid state. On the contrary, the reaction of 1 with SbCl(3) undergoes incomplete proton abstraction, resulting in the formation of the tricyclic compound N(3)P(3)(CyNH)(4)(CyNSbCl(2))(2), 4, which contains two four-coordinate Sb centers chelated by N(exo)-N(ring) sites of the phosphazene. PMID- 15651874 TI - Two distinct Ln(III)-Cu(I) cyanide extended arrays: structures and synthetic methodology for inclusion and layer complexes. AB - Encapsulation complexes formulated as {[La(DMF)(9)](2)[Cu(12)(CN)(18)].2DMF}(infinity), 1, and {[Ln(DMF)(8)][Cu(6)(CN)(9)].2DMF}(infinity) (Ln = Eu, 2; Gd, 3; Er, 4) were obtained from the one step reaction of LnCl(3) (Ln = La, Eu, Gd, Er) with CuCN and KCN in DMF. They consist of a three-dimensional Cu-CN anionic array with pockets occupied by the cation, [Ln(DMF)(x)](3+) (x = 8, 9). These complexes are believed to be the first examples of encapsulated Ln(3+) cations, and the zeolite like anionic network is unique. A two step procedure that employs the same components generates the layer structure {Ln(DMF)(4)Cu(2)(CN)(5)}(infinity) (Ln = La, 5; Gd, 6; Er, 7) in which the five-membered ring repeating unit has Cu-CN-Ln and Cu-CN-Cu linkages which are also without precedent. Encapsulation complexes can also be prepared from CuCl, reacting with LnCl(3) and KCN. The crystal structure of {K(DMF)(2)Cu(CN)(2)}(infinity) (8) provides insight into the proposed reaction pathways for forming these two different structural types. PMID- 15651875 TI - Reaction of thiols with N-bonded sulfenamide complexes of cobalt(III): steric effect and reaction pathway. AB - The products and kinetics of the reaction of several thiols (RSH = 2 aminoethanethiol, cysteine, penicillamine, cysteine ethyl ester) with N-bonded sulfenamide complexes ([Co(en)(2)(NH(2)S(CH(2))(2)NH(2)](3+) (IA), [Co(en)(2)(NH(2)SCH(2)CH(CO(2)H)NH(2)](3+) (IC), [Co(en)(2)(NH(2)SC(CH(3))(2)CH(CO(2)H)NH(2)](3+) (IP)) have been studied. The reaction proceeds by nucleophilic attack at sulfur with cleavage of the N-S bond to form a disulfide and leave a coordinated NH(3) ligand. The kinetics (pH 4-10) reveal that the deprotonated thiol, RS(-), is the reactive nucleophile and that the N-deprotonated sulfenamide complex is unreactive. The reactions of IP are approximately 10(4) times slower than those of IA or IC, and the reasons and consequences of this large steric effect are discussed. It is concluded, on the basis of these and other observations from the literature, that there will be substantial steric retardation to nucleophilic attack at two-coordinate sulfur in a R-C(CH(3))(2)-S-X-R' unit because of the regiospecificity of the reaction. The acid dissociation constants of IP and the X-ray structure of its bromide salt also are reported. PMID- 15651876 TI - Syntheses, structures, and ion-exchange properties of the three-dimensional framework uranyl gallium phosphates, Cs4[(UO2)2(GaOH)2(PO4)4].H2O and Cs[UO2Ga(PO4)2]. AB - The reaction of UO(2)(NO(3))(2).6H(2)O with Cs(2)CO(3) or CsCl, H(3)PO(4), and Ga(2)O(3) under mild hydrothermal conditions results in the formation of Cs(4)[(UO(2))(2)(GaOH)(2)(PO(4))(4)].H(2)O (UGaP-1) or Cs[UO(2)Ga(PO(4))(2)] (UGaP-2). The structure of UGaP-1 was solved from a twinned crystal revealing a three-dimensional framework structure consisting of one-dimensional (1)(infinity)[Ga(OH)(PO(4))(2)](4-) chains composed of corner-sharing GaO(6) octahedra and bridging PO(4) tetrahedra that extend along the c axis. The phosphate anions bind the UO(2)(2+) cations to form UO(7) pentagonal bipyramids. The UO(7) moieties edge-share to create dimers that link the gallium phosphate substructure into a three-dimensional (3)(infinity)[(UO(2))(2)(GaOH)(2)(PO(4))(4)](4-) anionic lattice that has intersecting channels running down the b and c axes. Cs(+) cations and water molecules occupy these channels. The structure of UGaP-2 is also three dimensional and contains one-dimensional (1)(infinity)[Ga(PO(4))(2)](3-) gallium phosphate chains that extend down the a axis. These chains are formed from fused eight-membered rings of corner-sharing GaO(4) and PO(4) tetrahedra. The chains are in turn linked together into a three-dimensional (3)(infinity)[UO(2)Ga(PO(4))(2)](1-) framework by edge-sharing UO(7) dimers as occurs in UGaP-1. There are channels that run down the a and b axes through the framework. These channels contain the Cs(+) cations. Ion-exchange studies indicate that the Cs(+) cations in UGaP-1 and UGaP-2 can be exchanged for Ca(2+) and Ba(2+). Crystallographic data: UGaP-1, monoclinic, space group P2(1)/c, a = 18.872(1), b = 9.5105(7), c = 14.007(1) A, beta = 109.65(3)(o) , Z = 4 (T = 295 K); UGaP-2, triclinic, space group P, a = 7.7765(6), b = 8.5043(7), c = 8.9115(7) A, alpha = 66.642(1)(o), beta = 70.563(1)(o), gamma = 84.003(2)(o), Z = 2 (T = 193 K). PMID- 15651877 TI - Cooperative pull and push effects on the O-O bond cleavage in acylperoxo complexes of [(Salen)MnIIIL]: ensuring formation of manganese(V) oxo species. AB - The acidity (pull) and the axial ligand (push) effects on the O-O bond cleavage in the [(Salen)Mn(III)(RCO(3))L] acylperoxo complexes, with model L = none, NH(3), and HCO(2)(-) (1), have been studied with B3LYP density functional calculations. The acidic conditions have been mimicked by explicit protonation of 1 to afford a variety of [(Salen)Mn(III)(RCO(3)H)L] (2) and [(SalenH)Mn(III)(RCO(3))L] (3) complexes in ground quintet states. The protonation assists the O-O bond heterolysis, thus primarily forming highly reactive Mn(V)(O) species, and consequently suppresses formation of the less reactive Mn(IV)(O) species through homolytic channel described earlier in 1 [Khavrutskii, I. V.; Rahim, R. R.; Musaev, D. G.; Morokuma, K. J. Phys. Chem. B 2004, 108, 3845-3854]. In addition to the qualitative change of the O-O bond cleavage mode, the protonation affects the rate of the O-O bond cleavage. Therefore, varying the acidity of the reaction media helps control the O-O bond cleavage mode and rate. PMID- 15651878 TI - Oxidative dissolution of copper and zinc metal in carbon dioxide with tert-butyl peracetate and a beta-diketone chelating agent. AB - A series of beta-diketone ligands, R(1)COCH(2)COR(2) [tmhdH (R(1) = R(2) = C(CH(3))(3)); tfacH (R(1) = CF(3); R(2) = CH(3)); hfacH (R(1) = R(2) = CF(3))], in combination with tert-butyl peracetate (t-BuPA), have been investigated as etchant solutions for dissolution of copper metal into carbon dioxide solvent. Copper removal in CO(2) increases in the order tfacH < tmhdH < hfacH. A study of the reactions of the hfacH/t-BuPA etchant solution with metallic copper and zinc was conducted in three solvents: scCO(2) (supercrical CO(2)); hexanes; CD(2)Cl(2). The etchant solution/metallic zinc reaction produced a diamagnetic Zn(II) complex, which allowed NMR identification of the t-BuPA decomposition products as tert-butyl alcohol and acetic acid. Gravimetric analysis of the amount of zinc consumed, together with NMR studies, confirmed the 1:1:2 Zn:t BuPA:hfacH reaction stoichiometry, showing t-BuPA to be an overall two-electron oxidant for Zn(0). The metal-containing products of the copper and zinc reactions were characterized by elemental analysis, IR spectroscopy, and, as appropriate, NMR spectroscopy and single-crystal X-ray diffraction [trans M(hfac)(2)(H(2)O)(CH(3)CO(2)H) (1, M = Cu; 2, M = Zn)]. On the basis of the experimental results, a working model of the oxidative dissolution reaction is proposed, which delineates the key chemical variables in the etching reaction. These t-BuPA/hfacH etchant solutions may find application in a CO(2)-based chemical mechanical planarization (CMP) process. PMID- 15651879 TI - Structural and spectroscopic characterization of first-row transition metal(II) substituted blue copper model complexes with hydrotris(pyrazolyl)borate. AB - [CuL(SC(6)F(5))] (1) (L = hydrotris(3,5-diisopropyl-1-pyrazolyl)borate anion) has been reported as a good model for blue copper proteins [Kitajima, N.; Fujisawa, K.; Tanaka, M.; Moro-oka, Y. J. Am. Chem. Soc. 1992, 114, 9232-9233]. To obtain more structural and spectroscopic insight, the first-row transition metal(II) substituted complexes of Cu(II) (1) to Mn(II) (2), Fe(II) (3), Co(II) (4), Ni(II) (5), and Zn(II) (6) were synthesized and their crystal structures were determined. These model complexes have a distorted tetrahedral geometry arising from the tripodal ligand L. The d value, which is defined by the distance from the N(2)S basal plane to the metal(II) ion, and the bond angles such as N-M-N and S-M-N are good indicators of these structural distortions. The obtained complexes were characterized by UV-vis absorption, EPR, NMR, far-IR, and FT-Raman spectroscopies and electrochemical and magnetic properties. In UV-vis absorption spectra, the sulfur-to-metal(II) CT bands and the d-d transition bands are observed for 1 and 3-5. For 1, the strong sulfur to Cu(II) CT band at 663 nm, which is one of the unique properties of blue copper proteins, is observed. The CT energies of the Fe(II) (3), Co(II) (4), and Ni(II) (5) complexes are shifted to higher energy (308 and 355 nm for 3, 311 and 340 nm for 4, 357 and 434 nm for 5) and are almost the same as the corresponding Co(II)- and Ni(II)-substituted blue copper proteins. In the far-IR spectra, three far-IR absorption bands for 2 6 at ca. 400, ca. 350, and ca. 310 cm(-1) are also observed similar to those for 1. Other properties are consistent with their distorted tetrahedral geometries. PMID- 15651880 TI - Electrochemical synthesis and structural characterization of silver(I) complexes of N-2-pyridyl sulfonamide ligands with different nuclearity: influence of the steric hindrance at the pyridine ring and the sulfonamide group on the structure of the complexes. AB - A new series of silver complexes, [AgL], of the anionic forms of potentially bidentate N-2-pyridyl sulfonamide ligands [N-(3-methyl-2-pyridyl)-p toluenenesulfonamide (HTs3mepy), N-(3-methyl-2-pyridyl)mesitylenesulfonamide (HMs3mepy), N-(4-methyl-2-pyridyl)-p-toluenesulfonamide (HTs4mepy), and N-(6 methyl-2-pyridyl)mesitylenesulfonamide (HMs6mepy)] have been prepared by an electrochemical procedure. In addition, heteroleptic complexes of composition [AgLL'] (L' = 1,10-phenanthroline and 2,2'-bipyridine) were obtained when the coligand L' was added to the electrolytic phase. The complexes were characterized by microanalysis, IR and (1)H NMR spectroscopy, and LSI mass spectrometry. In the cases of the compounds [Ag(Ts3mepy)](n)() (1), [Ag(4)(Ms3mepy)(4)] (2a), [Ag(Ms3mepy)](n)() (2b), [Ag(4)(Ms6mepy)(4)] (3a), [Ag(2)(Ms6mepy)(2)](n)() (3b), [Ag(2)(Ms3mepy)(2)(phen)(2)] (5), [Ag(2)(Ms6mepy)(2)phen] (7), and [Ag(2)(Ts4mepy)(2)(bipy)(2)] (8), characterization was also carried out by single crystal X-ray diffraction. Compounds 1 and 2b present a polymer structure formed by an {AgN(2)} digonal core. Compounds 2a and 3a are tetranuclear and also have a distorted {AgN(2)} digonal core. Compound 3b is based on binuclear distorted {AgN(2)} digonal units joined by an intermolecular sulfonyl oxygen atom to produce a stairlike polymer structure. The heteroleptic complexes 5 and 8 are dimeric with a distorted {AgN(4)} tetrahedral geometry, while compound 7 shows two different geometries around the metal, distorted {AgN(2)} digonal and {AgN(4)} tetrahedral. The supramolecular structures of all species are organized by pi,pi-stacking, C-H...pi, or C-H...O interactions. PMID- 15651881 TI - Synthesis and structural characterization of bi- and trimetallic octacyanometalate(IV) complexes: [Delta,Lambda-MII(en)3][cis MII(en)2(OH2)][MIV(CN)8].2H2O and [cis-MII(en)2(OH2)]2[(mu-NC)2MIV(CN)6].4H2O (MII = Mn, Co, Ni; MIV = Mo, W). AB - Treatment of [M(II)(en)(3)][OTs](2) or methanolic ethylenediamine solutions containing transition metal p-toluenesulfonates (M(II) = Mn, Co) with aqueous K(4)M(IV)(CN)(8).2H(2)O or Cs(3)M(V)(CN)(8) (M(IV) = Mo, W; M(V) = Mo) affords crystalline clusters of [M(II)(en)(3)][cis-M(II)(en)(2)(OH(2))(mu NC)M(IV)(CN)(7)].2H(2)O (M(IV) = Mo; M(II) = Mn, 1; Ni, 5; M(IV) = W; M(II) = Mn, 2; Ni, 6) and [cis-M(II)(en)(2)(OH(2))](2)[(mu-NC)(2)M(IV)(CN)(6)].4H(2)O (M(IV) = Mo; M(II) = Co, 3; Ni, 7; M(IV) = W; M(II) = Co, 4) stoichiometry. Each cluster contains cis-M(II)(en)(2)(OH(2))(mu-NC)(2+) units that likely result from dissociative loss of en from [M(II)(en)(3)](2+), affording cis M(II)(en)(2)(OH(2))(2)(2+) intermediates that are trapped by M(IV)(CN)(8)(4-). PMID- 15651882 TI - Insertion reactions of carbon dioxide into Zn-N bonds: syntheses and structures of tetrameric and dimeric alkylzinc carbamato complexes. AB - The formal insertions of carbon dioxide into a series of methylzinc dialkylamide complexes (MeZnNR(2)) initially form solvent-free, tetrameric zinc carbamato complexes [Me(4)Zn(4)(O(2)CNR(2))(4)] (NR(2) = N(i-Pr)(2) (1), N(i-Bu)(2) (2), and piperidinyl (3)). These compounds have been characterized by traditional techniques as well as by single-crystal X-ray diffraction analyses. The tetrameric backbones seen in the solid state for 1-3 were structurally similar to each other. Addition of excess pyridine (py) to 1-3 breaks apart the tetramers and converts them into solvated dimeric species [Me(2)Zn(2)(O(2)CNR(2))(2)(py)(2)] (NR(2) = N(i-Pr)(2) (4), N(i-Bu)(2) (5), and piperidinyl (6)). X-ray crystallographic analyses of 4 and 5 confirmed the dimeric structure in the solid state. This study significantly increases the number of well-characterized zinc carbamates prepared via CO(2) insertion into zinc amides. PMID- 15651883 TI - Studies of bicarbonate binding by dinuclear and mononuclear Ni(II) complexes. AB - Bicarbonate ion reacts with the dinuclear nickel(II) complex containing the taec ligand (taec = N,N',N' ',N' ''-tetrakis(2-aminoethyl)-1,4,8,11 tetraazacyclotetradecane) in buffered aqueous solution to form the mu eta(2),eta(2)-carbonate complex with a large effective binding constant for bicarbonate ion, log K(B) = 4.39 at pH = 7.4. In contrast, the dinuclear nickel(II) complex containing the o-xyl-DMC(2) ligand (o-xyl-DMC(2) = alpha,alpha'-bis(5,7-dimethyl-1,4,8,11-tetraazacyclotetradecan-6-yl)-o-xylene) does not react with bicarbonate or carbonate ion in aqueous solution. In propylene carbonate, the reaction of [Ni(2)(o-xyl-DMC(2))](4+) with bicarbonate proceeds rapidly to form the mu-eta(1),eta(1)-carbonate complex. The structure of this carbonate complex has been determined by an X-ray diffraction study that confirms the mu-eta(1),eta(1)-carbonate binding mode. A mononuclear analogue of [Ni(2)(taec)](4+), [Ni(2,3,2-tetraamine)](2+) does not form a detectable mononuclear or dinuclear product with bicarbonate ion in aqueous solution, but [NiDMC](2+) (DMC = 5,7-dimethyl-1,4,8,11-tetraazacyclotetradecane) reacts slowly with carbonate ion in aqueous solution to form a 2:1 complex. PMID- 15651884 TI - Electronic structure of Li2Ga and Li9Al4, two solids containing infinite and uniform zigzag chains. AB - The electronic structure of inorganic solids such as Li(2)Ga and Li(9)Al(4) containing infinite zigzag homoatomic chains is discussed. It is shown that Li(2)Ga, a solid for which a Zintl-type electron-counting approach would suggest that a half-filled pi-type band occurs as in trans-polyacetylene, is really a three-dimensional solid with strong covalent interchain connections and small effective charge transfer. The zigzag chains do not play a dominant role as far as the electronic structure near the Fermi level is concerned, and there is no reason for the occurrence of a Peierls distortion despite the possible analogy with trans-polyacetylene. It is suggested that even assuming that a Zintl-type approach is appropriate for electron counting purposes, the infinite zigzag chains in this compound and those in trans-polyacetylene are not isolobal. The bonding in Li(9)Al(4) and Li(2)Ga is very similar, and both phases are predicted to be stable three-dimensional metals. PMID- 15651885 TI - Synthesis and characterization of the novel monoamide derivatives of Gd-TTDA. AB - For this study, the N'-monoamide derivatives of TTDA (3,6,10-tri(carboxymethyl) 3,6,10-triazadodecanedioic acid), N'-methylamide (TTDA-MA), N'-benzylamide (TTDA BA), and N'-2-methoxybenzylamide (TTDA-MOBA), were synthesized. Their protonation constants and stability constants (log K(ML)'s) formed with Ca(2+), Zn(2+), Cu(2+), and Gd(3+) were determined by potentiometric titration in 0.10 M Me(4)NCl at 25.0 +/- 0.1 degrees C. The relaxivity values of [Gd(TTDA-MA)](-), [Gd(TTDA BA)](-), and [Gd(TTDA-MOBA)](-) remained constant with respect to pH changes over the range 4.5-12.0. The (17)O NMR chemical shift of H(2)O induced by [Dy(TTDA MA)(H(2)O)](-) at pH 6.80 showed 0.9 inner-sphere water molecules. Water proton relaxivity values for [Gd(TTDA-MA)(H(2)O)](-), [Gd(TTDA-BA)(H(2)O)](-), and [Gd(TTDA-MOBA)(H(2)O)](-) at 37.0 +/- 0.1 degrees C and 20 MHz are 3.89, 4.21, and 4.25, respectively. The water-exchange lifetime (tau(M)) and rotational correlation time (tau(R)) of [Gd(TTDA-MA)(H(2)O)](-), [Gd(TTDA-BA)(H(2)O)](-), and [Gd(TTDA-MOBA)(H(2)O)](-) are obtained from reduced the (17)O relaxation rate and chemical shifts of H(2)(17)O. The (2)H NMR longitudinal relaxation rates of the deuterated diamagnetic lanthanum complexes for the rotational correlation time were also thoroughly investigated. The water-exchange rates (K(298)(ex) for [Gd(TTDA-MA)(H(2)O)](-), [Gd(TTDA-BA)(H(2)O)](-), and [Gd(TTDA-MOBA)(H(2)O)](-) are lower than that of [Gd(TTDA)(H(2)O)](2)(-) but significantly higher than those of [Gd(DTPA)(H(2)O)](2)(-) and [Gd(DTPA-BMA)(H(2)O)]. The rotational correlation times for [Gd(TTDA-BA)(H(2)O)](-) and [Gd(TTDA-MOBA)(H(2)O)](-) are significantly longer than those of [Gd(TTDA)(H(2)O)](2)(-) and [Gd(DTPA)(H(2)O)](2)(-) complexes. The marked increase of the relaxivity of [Gd(TTDA-BA)(H(2)O)](-) and [Gd(TTDA-MOBA)(H(2)O)](-) results mainly from their longer rotational correlation time. The noncovalent interaction between human serum albumin (HSA) and [Gd(TTDA-BA)(H(2)O)](-) and [Gd(TTDA-MOBA)(H(2)O)](-) complexes containing a hydrophobic substituent was investigated by measuring the water proton relaxation rate of the aqueous solutions. The binding association constant (K(A)) values are 1.0 +/- 0.2 x 10(3) and 1.3 +/- 0.2 x 10(3) M(-1) for [Gd(TTDA-BA)(H(2)O)](-) and [Gd(TTDA-MOBA)(H(2)O)](-), which indicates a stronger interaction of [Gd(TTDA-BA)(H(2)O)](-) and [Gd(TTDA-MOBA)(H(2)O)](-) with HSA. PMID- 15651886 TI - Dendritic tetranuclear Ru(II) complexes based on the nonsymmetrical PHEHAT bridging ligand and their building blocks: synthesis, characterization, and electrochemical and photophysical properties. AB - Dinuclear and tetranuclear Ru(II) compounds 1, 2, 3, and 4 based on the PHEHAT ligand (PHEHAT = 1,10-phenanthrolino[5,6-b]-1,4,5,8,9,12-hexaazatriphenylene) are prepared and characterized on the basis of the data for other related mononuclear species. Their electrochemical and spectroscopic behaviors are discussed. The nonspectroelectrochemical correlation obtained for 1, 2, 3, and 4 is explained on the basis of these data. From the behavior in emission, it is concluded that the internal energy transfer takes place from the core to the peripheral metallic units in 3 and 4. PMID- 15651887 TI - Synthesis, structure, and kinetics and stereochemistry of base-catalyzed hydrolysis of meso- and rac-[Co2(tmpdtne)Cl2]4+, bis(pentaamine) complexes devoid of deprotonatable NH centers. AB - The bis(pentadentate) ligand tmpdtne binds two Co(II) centers, and the entity is readily oxidized to the dicobalt(III) derivative [Co(2)(tmpdtne)Cl(2)](4+) which has been separated into two isomeric forms. NMR studies establish these as meso and rac isomers arising from the different or same absolute configurations for the asym configuration about each Co(III) center. Each dinuclear ion base hydrolyses to the dihydroxo derivative [Co(2)(tmpdtne)(OH)(2)](4+) with retained asym configurations about each metal ion and also retained rac or meso configurations. The kinetics for the stepwise loss of the two Cl(-) ligands is uniphasic, and data are presented to show that the loss of the first chloride is rate determining and is followed by very rapid intramolecular and loss of the second Cl(-) via a hydroxo-bridged species to yield the observed dihydroxo derivative. Meso and rac forms of the latter have been crystallized. The X-ray crystal structure of the rac-dihydroxo complex is reported, and it establishes the configurations of all the complexes reported. The (1)H NMR spectra for the hydroxo ions show very high field Co-OH resonances (ca. delta-0.5 ppm) not observed previously for such ions, and this result is discussed in the context of published (1)H NMR data for bridged Co-OH-Co species. The base hydrolysis kinetics for the dichloro ions are first order in [OH(-)], and deprotonation at an alpha-CH(2) center (alpha to a pyridyl) is identified as the source of the catalysis, since there is no NH center available for deprotonation on the ligand. These data further support the new pseudoaminate base hydrolysis mechanism first reported in 2003. The values of k(OH) for the second-order base-catalyzed reaction are ca. 4.0 M(-1) s(-1) for both the rac and meso isomers, and these results are discussed in terms of the increased acidities of these 4+ cations compared to their 2+ ion counterparts. PMID- 15651888 TI - Carbon-halogen bond activation mechanism by copper(I) complexes of (2 pyridyl)alkylamine ligands. AB - The reaction of p-substituted benzyl halides ((Y)BnX; X = Cl, Br, and I; Y = p substituent, OMe, t-Bu, Me, H, F, Cl, and NO(2)) and copper(I) complexes supported by a series of (2-pyridyl)alkylamine ligands has been investigated to shed light on the mechanism of copper(I) complex mediated carbon-halogen bond activation, including ligand effects on the redox reactivity of copper(I) complexes which are relevant to the chemistry. For both the tridentate ligand (Phe)L(Pym2) [N,N-bis(2-pyridylmethyl)-2-phenylethylamine] and tetradentate ligand TMPA [tris(2-pyridylmethyl)amine] complexes, the C-C coupling reaction of benzyl halides proceeded smoothly to give corresponding 1,2-diphenylethane derivatives and copper(II)-halide complex products. Kinetic analysis revealed that the reaction obeys second-order kinetics both on the copper complex and the substrate; rate = k[Cu](2)[(Y)BnX](2). A reaction mechanism involving a dinuclear copper(III)-halide organometallic intermediate is proposed, on the basis of the kinetic results, including observed electronic effects of p-substituents (Hammett plot) and the rate dependence on the BDE (bond dissociation energy) of the C-X bond, as well as the ligand effects. PMID- 15651889 TI - Trigonal-bipyramidal tin(IV) complexes containing tetradentate tripodal tristhiolatophosphine ligands: synthesis, characterization, crystal structure, and transmetalation reactions. AB - The reactions of the lithium salts of the proligands P(C(6)H(4)-2-SH)(3) (P((H)SH)(3)), P(C(6)H(3)-3-SiMe(3)-2-SH)(3) (P((TMS)SH)(3)), and P(C(6)H(3)-5-Me 2-SH)(3) (P((Me)SH)(3)) with RSnCl(3) (R = Ph, Me, n-Bu), in THF at 0 degrees C, produced a series of trigonal-bipyramidal complexes of the type RSn(PS(3)). The crystal structures of PhSn(P(H)S(3)), PhSn(P(TMS)S(3)), and PhSn(P(Me)S(3)) reveal considerable distortion from local C(3v) symmetry for the Sn(PS(3)) group. Unique to PhSn(P(Me)S(3)) is the presence of intramolecular hydrogen bonding between one sulfur atom and an ortho H atom of the Ph group, creating a plane that includes this S atom and the corresponding C(6)H(3) ring, a phosphorus atom, and the PhSn group. An analysis of the (1)H, (13)C, and (31)P NMR data from a combination of HMQC, HMBC, 2-D COSY, and (1)H{(31)P} NMR studies reveals that in solution the Sn(PS(3)) groups exhibit local C(3v) symmetry, even at low temperature. Byproducts frequently found in the synthesis of the proligands and tin complexes, and subsequent reactions, result from the oxidation of the trianionic tristhiolatophosphine ligand. The crystal structure of one of these, [OP((H)S(3))](2), shows that the molecule contains two ligands joined by a S-S bond. Within each original ligand the remaining two sulfur atoms form a S-S bond, and each phosphorus atom is oxidized. PhSn(P(TMS)S(3)) reacted with 2 equiv of FeCl(3) in CH(2)Cl(2) to produce the iron(IV) complex FeCl(P(TMS)S(3)). FeCl(P(TMS)S(3)) decomposed in the presence of excess FeCl(3). Similar transmetalation reactions with FeCl(2) or [Fe(2)OCl(6)](2)(-) required the addition of ferrocenium ion to complete the oxidation of iron to 4+. RuCl(P(TMS)S(3)) was prepared by the reaction between PhSn(P(TMS)S(3)) and RuCl(2)(DMSO)(4) without the addition of an external oxidizing agent. PMID- 15651890 TI - Short-range structure and cation bonding in calcium-aluminum metaphosphate glasses. AB - Comprehension of short- and medium-range order of phosphate glasses is a topic of interest, due to the close relation between network structure and mechanical, thermal, and optical properties. In this work, the short-range structure of glasses (1 - x)Ca(PO(3))(2).xAl(PO(3))(3) with 0 < or = x < or = 0.47 was studied using solid-state nuclear magnetic resonance spectroscopy, Raman spectroscopy, density measurements, and differential scanning calorimetry. The bonding between a network modifier species, Al, and the network forming phosphate groups was probed using high-resolution nuclear magnetic resonance spectroscopy of (27)Al and (31)P. Changes in the compositional behavior of the density, glass transition temperature, PO(2) symmetric vibrations, and Al coordination number were verified at around x = 0.30. (31)P NMR spectra show the presence of phosphorus in Q(2) sites with nonbridging oxygens (NBOs) coordinated by Ca ions and also Q(2) sites with one NBO coordinated by Al (namely, Q(2)(1Al)). The changes in the properties as a function of x can be understood by considering the mean coordination number measured for Al and the formation of only Q(2) and Q(2)(1Al) species. It is possible to calculate that a network formed only by Q(2)(1Al) phosphates can just exist up to the upper limit of x = 0.48. Above this value, Q(2)(2Al) species should appear, imposing a major reorganization of the network. Above x = 0.30 the network undergoes a progressive reorganization to incorporate Al ions, maintaining the condition that only Q(2)(1Al) species are formed. These observations support the idea that bonding principles for cationic species inferred originally in binary phosphate glasses can also be extended to ternary systems. PMID- 15651891 TI - "Venetian blinds" mechanism of solvation/desolvation in palladium(II) wheel-and axle organic-inorganic diols. AB - A family of organic-inorganic wheel-and-axle diols (Pd(LOH)(2)Cl(2), Pd(LOH)(2)(CH(3))Cl, Pd(LOH)(2)(CH(3)COO)(2), LOH = alpha-(4-pyridyl)benzhydrol) and several corresponding solvates are synthesized and characterized by single crystal X-ray diffraction analysis. Their structures are compared to investigate the factors governing the modes of solid state association, the propensity to clathration, and the structural basis of guest inclusion. In all the complexes, the palladium coordination is a slightly distorted square. The LOH ligands coordinate Pd(2+) by means of the 4-pyridyl ring. In the chloride complexes solvation occurs with a 1:2 host/guest ratio by hydrogen bonding between the terminal -OH groups of the complex diol and one acceptor atom on the guest, and it is further assisted by guest stacking between host aryl rings. All solvates are organized in layers with practically invariant metrics, while the layers may be assembled in different arrangements. The structures of the nonsolvate compounds are related to the metrics of the solvate forms by rotation of the complex molecules within the layer plane. In all cases the nonsolvates are completely converted into the corresponding crystalline solvate forms by exposure to the vapor of the guest, and conversely they are quantitatively recovered from the solvate upon removal of the guest by mild conditions. On the basis of the structural data, it is proposed that the solvation/desolvation process proceeds by a concerted rotation of the complex molecules in the layer plane. The structural analysis of Pd(LOH)(2)(CH(3)COO)(2) and of its tetrahydrofuran monosolvate form suggests that the first step of the solid/gas solvation process may imply the clathration of 1 mol of guest between the aryl rings, which successively triggers the collective reorientation of the host molecules. PMID- 15651892 TI - Experimental and theoretical investigations of tellurium(IV) diimides and imidotelluroxanes: X-ray structures of B(C6F5)3 adducts of OTe(mu-NtBu)2TeNtBu, [OTe(mu-NtBu)2Te(mu-O)]2 and tBuNH2. AB - The hydrolysis of (t)BuNTe(mu-N(t)Bu)(2)TeN(t)Bu (1) with 1 or 2 equiv of (C(6)F(5))(3)B.H(2)O results in the successive replacement of terminal imido groups by oxo ligands to give the telluroxane-Lewis acid adducts (C(6)F(5))(3)B.OTe(mu-N(t)Bu)(2)TeN(t)Bu (2) and [(C(6)F(5))(3)B.OTe(mu N(t)Bu)(2)Te(mu-O)](2) (3), which were characterized by multinuclear NMR spectroscopy and X-ray crystallography. The Te=O distance in 2 is 1.870(2) A. The di-adduct 3 involves the association of four (t)()BuNTeO monomers to give a tetramer in which both terminal Te=O groups [d(TeO) = 1.866(3) A] are coordinated to B(C(6)F(5))(3). The central Te(2)O(2) ring in 3 is distinctly unsymmetrical [d(TeO) = 1.912(3) and 2.088(2) A]. The X-ray structure of (C(6)F(5))(3)B.NH(2)(t)()Bu (4), the byproduct of these hydrolysis reactions, is also reported. The geometries and energies of tellurium(IV) diimides and imido telluroxanes were determined using quantum chemical calculations. The calculated energies for the reactions E(NR)(2) + Te(NR)(2) (E = S, Se, Te; R = H, Me, (t)Bu, SiMe(3)) confirm that cyclodimerization of tellurium(IV) diimides is strongly exothermic. In the mixed-chalcogen systems, the cycloaddition is energetically favorable for the Se/Te combination. The calculated energies for the further oligomerization of the dimers XE(mu-NMe)(2)EX (E = Se, Te; X = NMe, O) indicate that the formation of tetramers is strongly exothermic for the tellurium systems but endothermic (X = NMe) or thermoneutral (X = O) for the selenium systems, consistent with experimental observations. PMID- 15651893 TI - Facile C-H bond activation: synthesis of the N4C donor set pentadentate ligand 1,4-bis(2-pyridylmethyl)-1,4-diazacyclononane (dmpdacn) and a structural study of its alkyl-cobalt(III) complex [Co(dmpdacn-C)(OH2)](ClO4)2.H2O and its hydroxylated derivative [Co(dmpdacnOH-O)Cl](ClO4)2.C3H6O. AB - The 1,4-bis(2-pyridylmethyl)-1,4-diazacyclononane (dmpdacn) ligand with a N(4)C donor set deprotonates at a CH(2) gamma to an amine under extraordinarily mild conditions (pH 7) and binds as a pentadentate ligand to Co(III) as the [Co(dmpdacn-C)(OH(2))](2+) complex. This complex was characterized by 1D and 2D NMR techniques, and a single-crystal X-ray structure is reported. In an alternative synthesis from Co(II), dmpdacn, and air, the same C-bonded complex is obtained along with a novel hydroxylated Co(III) complex [Co(dmpdacnOH-O)Cl](2+) which has been similarly characterized. Here the carbanion has been oxidized, a C to O-bonded rearrangement has taken place, and the bound aqua group is replaced by Cl(-). The base hydrolysis kinetics of the hydroxylated Co(III) complex are reported, and mechanisms for this and the unusually facile C-H cleavage and CH(2) oxidation reactions are discussed. PMID- 15651894 TI - Synthesis and molecular structure of the first rhodium(I) complex containing a tetra-tert-butylcyclopentaphosphanide ligand. AB - Na[cyclo-(P(5)(t)Bu(4))] (1) reacts with [RhCl(PPh(3))(3)] (1:1) to give the first rhodium(I) complex with a tetra-tert-butylcyclopentaphosphanide ligand, [Rh{cyclo-(P(5)(t)Bu(4))}(PPh(3))(2)] (2). 2 was characterized by NMR ((1)H, (13)C, (31)P), MS, IR, and X-ray structure determination. PMID- 15651896 TI - Use of DNA microarrays to monitor host response to virus and virus-derived gene therapy vectors. AB - Given the biological complexity of viral infections, the variability of the host response, and the safety concerns related to viral-mediated gene transfer, recent studies have made use of DNA mircoarrays to integrate multi-layered experimental approaches aimed at completely clarifying virus-host interactions. Particular attention has been given to those viruses that are implicated in clinical use and/or in life-threatening diseases. Examples of such use can be divided into three main categories, including: (i) the use of microarrays to study viral expression; (ii) the use of microarrays to analyze the host response to viral infection; and (iii) the use of microarrays to characterize the host response to viral vector-mediated transduction. Significant information on virus- and viral vector-host interactions can be obtained with the microarray approach, including the recognition of master pathways of virally-induced responses, the identification of new target genes for specific viruses, and indications on the molecular toxicity of specific gene transfer vectors currently used for gene therapy trials (in particular, adeno-associated viruses and adenovirus-derived vectors). We predict that the development of accessible repositories containing most of the DNA microarray data on viral infections will certainly help to elucidate the puzzling pictures of different viral infections. This will be crucially important for the correct handling of viral diseases and the intelligent amelioration of viral vectors for gene therapy. PMID- 15651897 TI - Sequence tag catalogs of dust mite-expressed genomes: utility in allergen and acarologic studies. AB - Dust mites are a major source of indoor allergens. They contain a large number of components that react with immunoglobulin (Ig) E in individuals with allergies and are capable of inducing sensitization, and allergic respiratory and cutaneous diseases. With a significant proportion of the population affected in some way by mite allergies, it is essential that we improve our understanding of these organisms so that control strategies could be defined and its allergens better understood. Thus, we have initiated a project using the expressed sequence tagging (EST) strategy to study the major species of dust mites associated with allergic diseases, in particular, the American house dust mite, Dermatophagoides farinae, as well as Blomia tropicalis, the most prevalent mite in domestic tropical dwellings. The work has recently been expanded to include 'storage' mites such as Tyrophagus putrescentiae, Acarus siro, Lepidoglyphus destructor, Glycyphagus domesticus, Suidasia medanensis, and Aleuroglyphus ovatus. More than 50% of the initial 3000 ESTs from the D. farinae and B. tropicalis dust mites showed significant matches to known genes and were categorized into eight functional groups (such as proteins involved in metabolism, gene expression, protein synthesis, cell signaling, etc.). Of specific interest, however, were the homologs to known mite allergens, in addition to a number of sequences bearing significant homology to allergens from non-mite sources previously not known to exist in mites. The availability of these allergen sequences has facilitated their expression and subsequent characterization in our laboratory in terms of their IgE-binding reactivity. The wealth of sequence information, generated via the EST project, has also facilitated the identification of polymorphic forms of allergens, the investigation of differential gene expression under various environmental conditions via DNA microarrays, as well as the analysis of protein level expression profiling via the proteomics approach. Additionally, ESTs have also ameliorated the understanding of the phylogenetic relationships between mites, and enabled the isolation of gene products crucial for life processes so that mite control strategies can be more effectively devised. Taken together, the utilization of the EST strategy has opened up numerous new avenues by which the allergist can engage more effectively in the study of dust mites with the ultimate aim of developing appropriate treatment regimens for mite-induced allergy. PMID- 15651898 TI - Activity-based protein profiling: applications to biomarker discovery, in vivo imaging and drug discovery. AB - The genomic revolution has created a wealth of information regarding the fundamental genetic code that defines the inner workings of a cell. However, it has become clear that analyzing genome sequences alone will not lead to new therapies to fight human disease. Rather, an understanding of protein function within the context of complex cellular networks will be required to facilitate the discovery of novel drug targets and, subsequently, new therapies directed against them. The past ten years has seen a dramatic increase in technologies that allow large-scale, systems-based methods for analysis of global biological processes and disease states. In the field of proteomics, several well established methods persist as a means to resolve and analyze complex mixtures of proteins derived from cells and tissues. However, the resolving power of these methods is often challenged by the diverse and dynamic nature of the proteome. The field of activity-based proteomics, or chemical proteomics, has been established in an attempt to focus proteomic efforts on subsets of physiologically important protein targets. This new approach to proteomics is centered around the use of small molecules termed activity-based probes (ABPs) as a means to tag, enrich, and isolate, distinct sets of proteins based on their enzymatic activity. Chemical probes can be 'tuned' to react with defined enzymatic targets through the use of chemically reactive warhead groups, fused to selective binding elements that control their overall specificity. As a result, ABPs function as highly specific, mechanism-based reagents that provide a direct readout of enzymatic activity within complex proteomes. Modification of protein targets by an ABP facilitates their purification and isolation, thereby eliminating many of the confounding issues of dynamic range in protein abundance. In this review, we outline recent advances in the field of chemical proteomics. Specifically, we highlight how this technology can be applied to advance the fields of biomarker discovery, in vivo imaging, and small molecule screening and drug target discovery. PMID- 15651899 TI - Biomedical literature mining: challenges and solutions in the 'omics' era. AB - It is now obvious that the rate-limiting step in high throughput experimentation is neither data acquisition nor analysis, but rather our ability to interpret data on a genome-wide scale. Indeed, the explosion of data sampling capacity combined with increasing publication rates greatly impairs our ability to find meaning in vast collections of data. In order to support data interpretation, bioinformatic tools are needed to identify critical information contained in large bodies of literature. However, extracting knowledge embedded in free text is an arduous task, compounded in the biomedical field by an inconsistent gene nomenclature, domain-specific language and restricted access to full text articles. This paper presents a selection of currently available biomedical literature mining software. These tools rely on statistic and, more recently, semantic analyses (Natural Language Processing) to automatically extract information from the literature. In addition, a literature mining strategy has been developed to explore patterns of term occurrences in abstracts. This method automatically identifies relevant keywords in collections of abstracts, and uses a pattern discovery algorithm to generate a visual interface for exploring functional associations among genes. Term occurrence heatmaps can also be combined with gene expression profiles to provide valuable functional annotations. Furthermore, as demonstrated with tumor cell line literature profiling results, this approach can be applied to a variety of themes beyond genomic data analysis. Altogether, these examples illustrate how literature analysis can be employed to support knowledge discovery in biomedical research. PMID- 15651900 TI - Polymorphisms of CYP2C19 and CYP2D6 in Israeli ethnic groups. AB - BACKGROUND: The cytochrome P450 isoenzymes CYP2C19 and CYP2D6 catalyze reactions involved in the metabolism of many widely used drugs. Their polymorphisms give rise to important interindividual and interethnic variability in the metabolism and disposition of several therapeutic agents and may cause differences in clinical response to some drugs. Individuals who carry two null alleles of either gene are known as poor metabolizers (PMs), while those who carry more than two copies of the functional CYP2D6 gene are ultrarapid metabolizers (UMs). AIM: The aim of the current study was to genotype Israelis from four different ethnic backgrounds with respect to CYP2C19 and CYP2D6. STUDY DESIGN: Polymorphisms of the CYP2C19 and CYP2D6 genes were determined by genotyping the four ethnic groups using PCR and/or restriction fragment length polymorphism (RFLP) analysis. The groups consisted of three Jewish communities, Yemenite Jews (n = 36), Sephardic Jews (n = 47), Ethiopian Jews (n = 28), and one Arabian population, Bedouins (n = 50). RESULTS: CYP2C19*2 allele frequencies ranged from 12.0 to 19.6% among the four ethnic groups. Within the study population, the CYP2C19*3 gene was only found in one Bedouin individual, in the heterozygous state (CYP2C19*1/*3). In each group, one individual was homozygous for CYP2C19*2, and were predicted to be PMs. The data revealed a high prevalence of CYP2D6*2, *4, *10, *41, and gene duplication, followed by *5 and *17, while *3 was very rare. The frequencies of the CYP2D6*4, *10, and *17 alleles and CYP2D6 gene duplication were significantly different among the four groups. However, the CYP2D6*2, *3, and *5 and *41 alleles showed similar frequencies in the four groups. Four (8.5%) Sephardic Jews and one (2.0%) Bedouin were found with the genotype CYP2D6*4/*4 (two null alleles), and were thus presumably PMs. A total of 15 individuals, distributed in all groups, were found with functional CYP2D6 gene duplications. The frequencies of predicted UMs (duplication of CYP2D6) were 17.8% (5/28) and 12.8% (6/47) in Ethiopian Jews and Sephardic Jews, respectively, which were higher than that of Yemenite Jews (5.6%, 2/36) and Bedouins (4.0%, 2/50). CONCLUSIONS: This is the first study of the CYP2D6 gene polymorphism in Israeli ethnic groups, either Jewish or Arab. Furthermore, this is also the first study of the CYP2C19 gene polymorphism in Jewish or Arab subgroups living in Israel. The frequencies of various alleles for the CYP2D6 gene are significantly different among the ethnic groups in Israel. These new findings may have important clinical implications in administrating drugs metabolized by CYP2D6 and for CYP2D6-related adverse drug reactions in the Israeli population. PMID- 15651901 TI - Arterial stiffness and stroke in hypertension: therapeutic implications for stroke prevention. AB - Stroke is the second leading cause of mortality worldwide, and the leading cause of death in China and Japan. Its prevention represents a major goal. Identification of primary stroke risk, particularly through newly individualised risk factors including biomarkers of large artery damage such as arterial stiffening, is necessary for determining the appropriate level of intervention. The purpose of this review is to focus on the pathophysiology of arterial stiffness, its predictive value for stroke and the therapeutic implications of this risk factor for stroke prevention. The predictive value of arterial stiffness for stroke was demonstrated in a longitudinal study that included 1715 patients with essential hypertension and measurements of carotid-femoral pulse wave velocity (PWV) [an indicator of arterial stiffness] at entry. Over a mean follow-up period of 7.9 years, during which 25 fatal strokes occurred, PWV significantly predicted stroke (relative risk = 1.39 [(95% CI 1.08, 1.72]; p = 0.02 for each 4 m/sec increase) independently of classical cardiovascular risk factors, including age, cholesterol level, diabetes mellitus, smoking and mean blood pressure. Additional longitudinal studies are needed to confirm the predictive value of aortic stiffness on primary and secondary events, in low- and high-risk populations, in various countries, and using different methodologies of arterial stiffness measurement. Drug treatment could prevent stroke through a reduction in arterial stiffness in parallel with correction of cardiovascular risk factors such as hypertension, dyslipidaemia, diabetes mellitus and smoking, all of which are associated with arterial stiffening. In view of the important local actions of angiotensin II on arterial stiffening, drugs interfering with the renin-angiotensin-aldosterone system should be particularly effective. Promising therapeutic strategies to reduce arterial stiffness include taking advantage of the non-lipid-lowering effects of statins and directly targeting the molecular events leading to arterial stiffening, such as formation of advanced glycation end products. PMID- 15651904 TI - Efficacy and tolerability of reboxetine in depressive patients treated in routine clinical practice. AB - OBJECTIVES: Reboxetine, a potent and selective noradrenaline reuptake inhibitor, has been approved for treatment of major depression. The aim of this study was to investigate the efficacy and tolerability of reboxetine in depressive outpatients undergoing treatment in routine clinical practice. STUDY DESIGN AND METHODS: This post-marketing surveillance study was conducted to evaluate the therapeutic efficacy and tolerability of standard therapeutic doses of reboxetine in patients with depressive symptoms, particularly when administered in routine clinical practice. The 1835 patients (mean 54 years of age) evaluated showed demographic characteristics representative of the general depressive population. The majority of patients received the recommended dose of reboxetine 8 mg/day. RESULTS: Measures of efficacy showed improvement in depressive symptoms with reboxetine therapy over the mean observational period of 9.6 weeks. Response to therapy, defined as Hamilton depression scale 21-item version score reduction of > or =50%, was reported in 83% of patients. The effects of reboxetine were rated by physicians as 'good' or 'very good' in 86% of patients at the last visit. The tolerability of reboxetine was rated by physicians as 'good' or 'very good' in 92% of patients at all evaluations. No adverse events that were possibly related to reboxetine therapy occurred in >1% of patients. CONCLUSION: The results of this study suggest that reboxetine is safe and well tolerated and may improve symptoms in depressive patients treated in routine clinical practice. PMID- 15651903 TI - Switching between second-generation antipsychotics: why and how? AB - The introduction of second-generation antipsychotics represents an important advance in the treatment of schizophrenia. Although these drugs are generally very effective, not all patients respond in the same way. Partial response with persistent positive and negative symptoms and residual symptoms may force physicians to change antipsychotic medication. As more and more second-generation antipsychotics are introduced, the need for practical guidelines on switching these medications becomes increasingly important. In this article we provide a short summary of the second-generation antipsychotics, focusing on efficacy, adverse effect profile and safety. Indications for switching antipsychotic medication are outlined, as well as recommendations when switching is disadvantageous. Three basic switching strategies (abrupt, gradual and overlapping switching) and their potential risks and benefits are described. We review the available evidence concerning techniques, problems and consequences when switching from one second-generation antipsychotic agent to another and discuss potential difficulties. PMID- 15651902 TI - Novel approaches to the treatment of cocaine addiction. AB - Cocaine addiction continues to be an important public health problem with over 1.7 million users in the US alone. Although there are no approved pharmacotherapies for cocaine addiction, a number of medications have been tested with some promising results. In this review, we summarise some of the emerging targets for cocaine pharmacotherapy including dopaminergic and GABA medications, adrenoceptor antagonists, vasodilators and immunotherapies. The brain dopamine system plays a significant role in mediating the rewarding effects of cocaine. Among dopaminergic agents tested for cocaine pharmacotherapy, disulfiram has decreased cocaine use in a number of studies. Amantadine, another medication with dopaminergic effects, may also be effective in cocaine users with high withdrawal severity. GABA is the main inhibitory neurotransmitter in the brain, and accumulating evidence suggests that the GABA system modulates the dopaminergic system and cocaine effects. Two anticonvulsant medications with GABAergic effects, tiagabine and topiramate, have yielded positive findings in clinical trials. Baclofen, a GABA(B) receptor agonist, is also promising, especially in those with more severe cocaine use. Some of the physiological and behavioural effects of cocaine are mediated by activation of the adrenergic system. In cocaine users, propranolol, a beta-adrenoceptor antagonist, had promising effects in individuals with more severe cocaine withdrawal symptoms. Cerebral vasodilators are another potential target for cocaine pharmacotherapy. Cocaine users have reduced cerebral blood flow and cortical perfusion deficits. Treatment with the vasodilators amiloride or isradipine has reduced perfusion abnormalities found in cocaine users. The functional significance of these improvements needs to be further investigated. All these proposed pharmacotherapies for cocaine addiction act on neural pathways. In contrast, immunotherapies for cocaine addiction are based on the blockade of cocaine effects peripherally, and as a result, prevent or at least slow the entry of cocaine into the brain. A cocaine vaccine is another promising treatment for cocaine addiction. The efficacy of this vaccine for relapse prevention is under investigation. Many initial promising findings need to be replicated in larger, controlled clinical trials. PMID- 15651905 TI - Intramuscular interferon-beta-1a: in patients at high risk of developing clinically definite multiple sclerosis. AB - Intramuscular interferon-beta-1a, a recombinant interferon-beta approved in the US for the treatment of relapsing forms of multiple sclerosis (MS), has also been evaluated in the treatment of patients with a first clinical demyelinating episode and brain lesions consistent with MS confirmed by magnetic resonance imaging (MRI). In a randomised, double-blind trial, patients at high risk of developing clinically definite MS who received intramuscular interferon-beta-1a 30 microg once weekly had a 44% reduction in the cumulative probability of developing MS, compared with placebo recipients (rate ratio 0.56; p = 0.002), over a 3-year period after a first, MRI-confirmed demyelinating event. These results were supported by MRI findings that showed significantly smaller increases in the volume of brain lesions and the number of new/enlarging and gadolinium-enhancing lesions in interferon-beta-1a recipients than in those receiving placebo. A nonblind extension of this trial demonstrated that early treatment with interferon-beta-1a significantly reduced the probability of developing MS by 35% (p = 0.03), compared with delayed treatment, over a 5-year period. Intramuscular interferon-beta-1a was generally well tolerated; however, influenza-like syndrome was documented in >50% of patients at high risk of developing clinically definite MS who received the drug. PMID- 15651908 TI - Zolpidem: a review of its use in the management of insomnia. AB - Zolpidem (Ambien, Stilnox, Myslee, an imidazopyridine, is a nonbenzodiazepine hypnotic indicated for the short-term treatment of insomnia. Zolpidem improves sleep in patients with insomnia. Its overall tolerability is favourable when administered according to the manufacturer's prescribing information, with a low propensity to cause clinical residual effects, withdrawal, dependence or tolerance. In addition, most evidence suggests that the drug is associated with minimal rebound insomnia. In the only clinical trials that investigated the use of a hypnosedative drug in an 'as-needed' regimen, zolpidem produced a global improvement in sleep. Thus, zolpidem continues to be a useful therapeutic option in the pharmacological treatment of patients with insomnia. PMID- 15651909 TI - Diet and exercise for weight loss: a review of current issues. AB - Obesity is a fast growing epidemic that is primarily due to environmental influences. Nutrition and exercise represent modifiable factors with a major impact on energy balance. Despite considerable research, there remains continued debate regarding the energy content and the optimal macronutrient distribution for promoting healthy and effective weight loss. Low-fat diets have been advised for many years to reduce obesity. However, their effectiveness has been recently challenged, partly because the prevalence of obesity continues to rise despite reductions in fat intake. There are also concerns regarding the methodology of clinical trials showing benefits of fat reduction on weight loss. Although often viewed as a fad diet, very low-carbohydrate (ketogenic) diets are very popular and several recent clinical trials indicate they are more effective at promoting short-term weight loss and improving characteristics of the metabolic syndrome than low-fat diets. However, there is a need to obtain long-term safety and efficacy data. Clearly, weight loss can be achieved with a variety of diet interventions but the effects on other health-related aspects also need to be considered and studied in more detail. Exercise can have positive effects on weight loss, weight control and overall general health, although debate exists concerning the most effective mode, duration and intensity of exercise required to achieve these effects. Importantly, any effective weight control treatment must consider a life-long plan or there will likely be weight regain. Perhaps the most challenging, but rewarding, question that faces researchers is how to predict individual responses to diet and exercise interventions. PMID- 15651911 TI - The use of vibration training to enhance muscle strength and power. AB - Vibration has been combined with conventional resistance training in an attempt to attain greater gains in neuromuscular performance than from conventional resistance training alone. Although there is a lack of strictly controlled studies on the vibration training effect, current findings in this area suggest that vibration may have a beneficiary acute and/or chronic training effect on strength and power enhancement. However, the effect of vibration on strength and power development appears dependent upon the vibration characteristics (method of application, amplitude and frequency) and exercise protocols (training type, intensity and volume) employed. Vibration amplitude and frequency determine the load that vibration imposes on the neuromuscular system. This vibration load should be in an optimal range to elicit strength and power enhancement. To activate the muscle most effectively, vibration frequency should be in the range of 30-50 Hz. It is less clear to what the optimal amplitude should be, but smaller amplitudes may be insufficient to elicit an enhancement. It should also be noted that the method of vibration application (i.e. vibration applied directly or indirectly to a targeted muscle) may have an influence on the magnitude of amplitude and frequency that are delivered to the muscle and, therefore, may have an influence on vibration training effect. The employment of a greater exercise intensity and volume within a vibration training programme may facilitate a larger enhancement in strength and power. In addition, benefits from vibration training may be greater in elite athletes than non-elite athletes. Further studies are required to examine these inter-dependencies, especially in relation to chronic adaptation to dynamic exercises, which are the most relevant response to practitioners, but where the least amount of research has been undertaken. PMID- 15651910 TI - Aggregation and activation of blood platelets in exercise and training. AB - This article presents an overview of the progress that has been made in recent years in our understanding of the interaction between exercise and platelets in health and disease. Although platelets are important in normal haemostasis, recent evidence emphasises the pivotal role of abnormal platelet function in acute coronary artery diseases, myocardial infarction, unstable angina and stroke. In light of the positive health benefits of exercise, interest has been heightened on the association between exercise and platelet aggregation and function, not only in normal healthy subjects but also in patients. However, the study of exercise effects on blood platelets are highly contentious because of the fact that the analytical methods employed to study platelets are bedevilled by numerous methodological problems. While exercise effects on platelet aggregation and function in healthy individuals have been extensively examined, the evidence reported has been conflicting. Somewhat less contradictory are the results generated from studies in patients with coronary heart disease, as the preponderance of evidence available would strongly suggest that platelet aggregation and function are increased with exercise. Several drugs are known to influence platelet aggregation and function, the most examined among these medications is aspirin (acetylsalicylic acid). However, aspirin appears to be ineffective to attenuate exercise-induced increases in platelet aggregation and activation. Few studies are available on the effect of training on blood platelets and the exact effects of exercise training on platelet activation and function is not as yet known. This lack of information makes further studies particularly important, in order to clarify whether there are favourable effects of exercise training on platelet aggregation and function in health and disease. PMID- 15651912 TI - The impact of instability resistance training on balance and stability. AB - The most predominant literature regarding balance has emphasised the physiological mechanisms controlling stability. Topics range from extrinsic factors (environment) to intrinsic factors (i.e. muscle coordination, vestibular response). Balance is achieved through an interaction of central anticipatory and reflexive actions as well as the active and passive restraints imposed by the muscular system. However, less research has attempted to document the effects of balance on performance measures (i.e. force, power). Furthermore, short- and long term adaptations to unstable environments need more substantial research. While force and other performance measures can be adversely affected by a lack of balance, the transferability of instability training to activities of daily living and sport is not precisely known. The applicability of instability and resistance training using unstable platforms or implements may have strong relevance in a rehabilitative or athletic setting. Therefore, a comprehensive review of the literature in this area may possibly be of benefit to practitioners who deal with the general population, athletes or persons debilitated by balance and/or stability disabilities. PMID- 15651913 TI - Physiological aspects of surfboard riding performance. AB - Surfboard riding (surfing) has experienced a 'boom' in participants and media attention over the last decade at both the recreational and the competitive level. However, despite its increasing global audience, little is known about physiological and other factors related to surfing performance. Time-motion analyses have demonstrated that surfing is an intermittent sport, with arm paddling and remaining stationary representing approximately 50% and 40% of the total time, respectively. Wave riding only accounts for 4-5% of the total time when surfing. It has been suggested that these percentages are influenced mainly by environmental factors. Competitive surfers display specific size attributes. Particularly, a mesomorphic somatotype and lower height and body mass compared with other matched-level aquatic athletes. Data available suggest that surfers possess a high level of aerobic fitness. Upper-body ergometry reveals that peak oxygen uptake (VO2peak) values obtained in surfers are consistently higher than values reported for untrained subjects and comparable with those reported for other upper-body endurance-based athletes. Heart rate (HR) measurements during surfing practice have shown an average intensity between 75% and 85% of the mean HR values measured during a laboratory incremental arm paddling VO2peak test. Moreover, HR values, together with time-motion analysis, suggest that bouts of high-intensity exercise demanding both aerobic and anaerobic metabolism are intercalated with periods of moderate- and low-intensity activity soliciting aerobic metabolism. Minor injuries such as lacerations are the most common injuries in surfing. Overuse injuries in the shoulder, lower back and neck area are becoming more common and have been suggested to be associated with the repetitive arm stroke action during board paddling. Further research is needed in all areas of surfing performance in order to gain an understanding of the sport and eventually to bring surfing to the next level of performance. PMID- 15651915 TI - Neurotoxicology: principles and considerations of in vitro assessment. AB - Neurotoxicology is an exciting area of science, not only because of the importance of toxic injury to the nervous system in human disease, but also because specific toxicants have served as invaluable tools for the advancement of our knowledge of "normal" neurobiological processes. In fact, much of our understanding of the organisation and function of the nervous system is based on observations derived from the actions of neurotoxicants. This paper addresses various physiological aspects behind the exquisite sensitivity of the nervous system to toxic agents, including the privileged status of the nervous system vis a-vis blood-brain barrier function, the extensions of the nervous system over space and the requirements of cells with such a complex geometry, and the transmission of information across extracellular space. In addition, in vitro models and their utility in the assessment of neurotoxicological outcome are discussed, with reference to both their advantages and disadvantages. PMID- 15651916 TI - Organotypic brain slice cultures: an efficient and reliable method for neurotoxicological screening and mechanistic studies. AB - This paper reviews the current state of the use of organotypic brain slice cultures for neurotoxicological and neuropharmacological screening and mechanistic studies, as exemplified by excitotoxin application. At present, no in vitro systems have been approved by the regulatory authorities for neurotoxicity testing. For the evaluation of the slice culture method, organotypic hippocampal slice cultures were exposed to toxic doses of the excitotoxins, glutamate, N methyl-D-aspartate (NMDA), kainic acid and 2-amino-3-hydroxy-5-methyl-4-isoxazole propionate (AMPA), and the glial toxin, DL-alpha-aminoadipic acid (DLAAA). Neuronal cell death was quantified by propidium iodide (PI) uptake, and visualised by Fluoro-Jade (FJ) staining. General cell death was monitored by lactate dehydrogenase (LDH) release into the culture medium. EC50 values for the different compounds, based on PI uptake after exposure for 48 hours in entire cultures, were: glutamate, 3.5 mM; DL-AAA, 2.3 mM; kainic acid, 13 microM; NMDA, 11 microM; and AMPA, 3.7 microM. In the slice cultures, the hippocampal subfields displayed the same differences in vulnerability as those observed in vivo. When subfield analysis was performed on the cultures, the CA1 subfield was most susceptible to glutamate, NMDA and AMPA, while CA3 was most susceptible to kainic acid. The amount of LDH release for DL-AAA was about four times that of L glutamate, in accordance with the additional toxic effect on glial cells, which was also found by confocal microscopy to stain for FJ. In conclusion, it was found that organotypic brain slice culture, combined with standardised protocols and quantifiable markers, such as PI and FJ staining, is a relevant and feasible in vitro system for neurotoxicity testing. Considering the amount and quality of the available published data, it is recommended that the brain slice culture method could be subjected to pre-validation and formal validation for inclusion in a tiered in vitro neurotoxicity testing scheme to supplement and replace conventional animal tests. PMID- 15651917 TI - Development of a sensory neuronal cell model for the estimation of mild eye irritation. AB - In an attempt to improve the in vitro test strategy for the estimation of eye irritation, a neuronal cell model has been developed, with cells expressing vanilloid receptor type 1 (VR1) nociceptors. The currently accepted method for measuring eye irritancy is the ethically and scientifically criticised Draize rabbit eye test, despite the fact that alternative in vitro methods are available which have proved to be reliable and reproducible for predicting severe ocular toxicity. However, no alternative tests for measuring neuronal stimulation have yet been developed, and the prediction of eye irritation in the mild range is therefore insufficient. VR1 is a nociceptor localised in C-fibre neurons innervating the cornea and the surrounding tissue, and it responds to potentially damaging stimuli by releasing Ca2+ into the cytoplasm. As a sensory endpoint, [Ca2+]i was measured in VR1 transfected cells, as well as in control cells. Short term cell cytotoxicity studies (cell membrane rupture and morphological divergence) were used to determine the non-corrosive concentrations of the test chemicals. Preliminary results indicated that hygiene products used daily may induce eye irritation via VR1 nociceptors. The lowest toxic concentration (0.025%) of liquid hand soap, as determined by morphologic divergences of cells, generated an 80% increase in [Ca2+]i over the basal [Ca2+]i in VR1 transfected cells, whereas the non-specific [Ca2+]i increased by 33%. Furthermore, all the endpoints studied indicated that shampoo for children was less active than shampoo for adults. If this method is successfully validated with standardised reference chemicals, the model could complete the test battery of in vitro alternatives, resulting in the saving of thousands of laboratory animals. PMID- 15651918 TI - Assessment of ocular irritation by image processed quantification of cell injury in human corneal cell cultures and in corneal constructs. AB - Currently, there are no accepted alternative tests for the replacement of animals in ocular irritation testing. This study focused on the quantification of cellular viability as a measure of toxic events in immortalised human corneal cell cultures and a three-dimensional corneal construct. Simultaneous vital dye staining by calcein AM and ethidium homodimer-1 was used to provide "live" and "dead" probes, respectively. For further quantification, we have developed image processing tools to evaluate digital images obtained from confocal fluorescence scanning microscopy measurements. Based on the finding that ocular irritation can be related to the extent of cell injury at the various cell layers of the cornea, we extended our studies from corneal cell cultures to an in vitro human corneal equivalent system comprising epithelial, stromal keratocyte and endothelial layers. Our results showed that the microscopic measurement of cellular injury by using either cell cultures or in vitro corneal constructs, combined with image processed quantification, can provide insight into the extent of the toxic effects. PMID- 15651914 TI - Patellar tendinopathy in athletes: current diagnostic and therapeutic recommendations. AB - Formerly known as 'jumper's knee', patellar tendinopathy gives rise to considerable functional deficit and disability in recreational as well as professional athletes. It can interfere with their performance, often perseveres throughout the sporting career and may be the primary cause to end it. The diagnosis of patellar tendinopathy is primarily a clinical one but new imaging techniques, such as Doppler ultrasonography, may provide additional diagnostic value. Current therapeutic protocols are characterised by wide variability ensuing from anecdotal experience rather than evidence. Moreover, numerous reports in recent years have shattered previous doctrines and dogmatic belief on tendon overuse. Histopathological and biochemical evidence has indicated that the underlying pathology of tendinopathy is not an inflammatory tendinitis but a degenerative tendinosis. Consequently, pain in chronic patellar tendinopathy is not inflammatory in nature, but its exact origin remains unexplained. In pursuit of pathology- and evidence-based management, conservative therapy should be shifted from anti-inflammatory strategies towards a complete rehabilitation with eccentric tendon strengthening as a key element. If conservative management fails, surgery is opted for. However, considering the heterogeneity of surgical procedures and the absence of randomised studies, no conclusive evidence can be drawn from the literature regarding the effectiveness of surgical treatment for patellar tendinopathy. Parallel with the improved knowledge on the pathophysiology and pain mechanisms in patellar tendinopathy, new treatment strategies are expected to emerge in the near future. PMID- 15651919 TI - Cellular effects of electromagnetic fields. AB - Studies at the cellular level are needed to reveal the cellular and molecular biological mechanisms underlying the biological effects and possible health implications of non-ionising radiation, such as extremely low frequency (ELF) magnetic fields (MFs) and radiofrequency (RF) fields. Our research group has studied the effects of 50 Hz ELF MFs (caused by power lines and electric devices) and 872 MHz or 900 MHz RFs (emitted by mobile phones and their base stations) on cellular ornithine decarboxylase activity, cell cycle kinetics, cell proliferation, and necrotic or apoptotic cell death. For RFs, pulse-modulated (217 Hz modulation frequency corresponding a global system for mobile communication-type signal) or continuous wave (unmodulated) signals were used. To expose the cell cultures to MFs or RFs, specially developed exposure systems were used, where levels of electromagnetic field exposure and the conditions of cell culture could be precisely controlled. A coexposure approach was used in many studies, i.e. the cell cultures were exposed to other stressors in addition to MFs or RFs. Ultraviolet radiation, serum deprivation, or fresh medium addition, were used as co-exposures. The results presented in this short review show that the effects of mere MFs or RF on cell culture models are quite minor, but that various co-exposure approaches warrant additional study. PMID- 15651920 TI - Effects of aluminium and lead on ATPase activity of knockout +/- mouse cerebral synaptosomes in vitro. AB - In this in vitro study, changes in the activity of the neural membrane integral protein, ATPase, were recorded after the exposure of isolated synaptosomes to different concentrations of aluminium and lead. Both total ATPase activity and Mg(2+)-ATPase activity were studied. A specific mouse strain, heterozygous for a glial cell line-derived neurotrophic factor (GDNF), and the corresponding wildtype mouse cerebral tissue, were used for the synaptosome isolations. The ATPase activities of the +/- mouse synaptosomes were compared with those of wild type synaptosomes. The decrease in total ATPase activity was similar in both types of synaptosomes, but after exposure to aluminium, the decrease of Mg(2+) ATPase activity in the GDNF+/- synaptosomes was smaller than that in the wild type synaptosomes. After exposure to lead, the protective effect of GDNF was not so clear. The synaptosomal effects of lead were already found at concentrations lower than those where cell toxicity appeared in SH-SY5Y cell cultures. Thus, synaptosomal ATPase activity was considered to be a sensitive marker for the detection of lead-induced neurotoxicity. PMID- 15651921 TI - Cytotoxicity of the dicarboximide fungicides, vinclozolin and iprodione, in rat hepatoma-derived Fa32 cells. AB - Dicarboximide fungicides are widely used to control various fungal species. Their primary action is not known, due to a lack of knowledge concerning the mechanism of action of the dicarboximide group. The cytotoxicities of vinclozolin and iprodione in rat hepatoma-derived Fa32 cells were investigated. Cytotoxicity was measured by neutral red uptake inhibition after treatment for 24 hours. Iprodione was more toxic than vinclozolin. Vinclozolin was less toxic in glutathione depleted cells than in control cells. This was also true for iprodione at lower concentrations, but iprodione became more toxic at higher concentrations. Both the fungicides increased the endogenous glutathione content by 20% after 1 hour. After 24 hours, the glutathione content was doubled by vinclozolin, but was not affected by iprodione. No effect on glutathione S-transferase activity or reactive oxygen species formation could be observed. Cytochrome P450-dependent ethoxyresorufin-O-deethylase and pentoxyresorufin-O-depentylase activities were moderately activated by iprodione and strongly activated by vinclozolin. A glutathione-related cytochrome P450-dependent metabolic attack of vinclozolin and iprodione could be responsible for their cytotoxicity in Fa32 cells. Further research is needed to fully elucidate these (or other) mechanisms. PMID- 15651922 TI - Cytotoxicity assays with fish cells as an alternative to the acute lethality test with fish. AB - In ecotoxicology, in vitro assays with fish cells are currently applied for mechanistic studies, bioanalytical purposes and toxicity screening. This paper discusses the potential of cytotoxicity assays with fish cells to reduce, refine or replace acute lethality tests using fish. Basal cytotoxicity data obtained with fish cell lines or fish primary cell cultures show a reasonable to good correlation with lethality data from acute toxicity tests, with the exception of compounds that exert a specific mode of toxic action. Basal cytotoxicity data from fish cell lines also correlate well with cytotoxicity data from mammalian cell lines. However, both the piscine and mammalian in vitro assays are clearly less sensitive than the fish test. Therefore, in vivo LC50 values (concentrations of the test compounds that are lethal to 50% of the fish in the experiment within 96 hours) currently cannot be predicted from in vitro values. This in vitro-in vivo difference in sensitivity appears to be true for both fish cell lines and mammalian cell lines. Given the good in vitro-in vivo correlation in toxicity ranking, together with the clear-cut difference in sensitivity, the role of cytotoxicity assays in a tiered alternative testing strategy could be in priority setting in relation to toxic hazard and in the toxicity classification of chemicals and environmental samples. PMID- 15651923 TI - Oestradiol potentiates the effects of certain pyrethroid compounds in the MCF7 human breast carcinoma cell line. AB - Pyrethroids are the most widely used insecticides for indoor pest control, so human exposure to them is common. The main target of pyrethroids is the nervous system, but their endocrine disrupting capabilities may also be of toxicological concern. In the present study, the proliferation of the breast cancer cell line, MCF7, was studied after a 7-day exposure to various concentrations of pyrethrin, permethrin and cypermethrin. The effects of oestradiol and the combined effects of oestradiol (0.10 nM) and pyrethroids (0.1-100 microM) on MCF7 cell proliferation were also evaluated. Proliferation and cell toxicity were studied by measuring the ATP content with a luminescence method, and mitochondrial metabolic enzyme activity with the WST-1 test. In the ATP test, low concentrations (0.1-1 microM) of pyrethroids in co-exposure with oestradiol caused a clear statistically significant increase in the proliferation of MCF7 cells. This was evident when compared to the proliferative effect caused by 0.1 nM oestradiol alone. High concentrations were cytotoxic, and the greatest cell toxicity was that of cypermethrin, which has a cyano group in its molecular structure. PMID- 15651924 TI - A new human breast carcinoma cell line resistant to DNA-damaging drugs. AB - To investigate the phenomenon of active dissociation of the vital dye, Hoechst 33342 (Ho342), from DNA (DNA clearing), a new MCF7HoeR-7 human breast carcinoma cell line was isolated from parent MCF7 cells by step-wise selection with increasing concentrations of Ho342. This cell line possesses an enhanced ability for DNA clearing. The MCF7HoeR-7 line is characterised in detail and compared with the parental MCF7 line and a typical P-glycoprotein-mediated multidrug resistant (MDR) cell line, MCF7/Adr. MCF7HoeR-7 cells have an increased population growth rate, a lower DNA content and a reduced number of chromosomes. Enhanced DNA clearing in MCF7HoeR-7 cells is associated with the high resistance of the cells to the toxic effects of Ho342 and cross-resistance to etoposide, a topoisomerase II inhibitor in clinical use. The MCF7HoeR-7 and parent MCF7 cell lines have similar expression levels of transport proteins. The results obtained confirm that DNA clearing is an atypical MDR mechanism in tumour cells. PMID- 15651925 TI - Modelling of normal and premalignant oral tissue by using the immortalised cell line, SVpgC2a: a review of the value of the model. AB - Normal oral keratinocytes (NOKs), and a Simian virus 40 T-antigen-immortalised oral keratinocyte line termed SVpgC2a, were cultured in an effort to model the human oral epithelium in vitro, including normal and dysplastic tissue. Monolayer and organotypic cultures of NOKs and SVpgC2a were successfully established in a standardised serum-free medium with high levels of amino acids, by using regular tissue culture plastic for monolayers and collagen gels containing oral fibroblasts as the base for generating tissue equivalents. NOKs express many characteristics of normal tissue, including those associated with terminal squamous differentiation. After > 150 passages, SVpgC2a cells retained an immortal, nontumourigenic phenotype that, relative to NOKs, was associated with aberrant morphology, enhanced proliferation, deficiency in terminal differentiation, proneness to apoptosis, and variably altered expression of structural epithelial markers. Transcript and protein profiling, as well as activity assays, demonstrated the expression of multiple xenobiotic-metabolising enzymes in SVpgC2a cells, some of which were higher in comparison to NOKs. A generally preserved, or even activated, ability for xenobiotic metabolism in long term cultures of SVpgC2a cells indicated that this cell line could be useful in safety testing protocols--for example, in the development of consumer products in the oral health care field. However, SVpgC2a cells displayed some features reminiscent of a severe oral dysplasia, implying that this cell line could also to some extent serve as a model of a premalignant oral epithelium. PMID- 15651926 TI - Mitochondrial toxicity detected in a health product with a boar spermatozoan bioassay. AB - Seaweed and organic alfalfa capsules sold as "health promoting" products had repeatedly caused emesis in a consumer. Using the boar spermatozoan bioassay, the capsule contents were found to contain a toxic substance that inhibited boar sperm motility and depolarised mitochondria at low exposure concentrations of 10 microg/ml. The capsule also contained high amounts (10(5)-10(7) cfu/g), of endospore-forming bacteria and Streptomyces-like bacteria. Bacteria from the capsule produced toxic substances when cultured in the laboratory. Three different toxic responses were provoked in the spermatozoa exposed to extracts from the Streptomyces-like isolates: a) hyperpolarisation of the plasma membrane and depolarisation of the mitochondria; b) depolarisation of mitochondria similar to that caused by the capsule content extract; and c) motility inhibition, with no observed change of any cytosolic transmembrane potential. Membrane potential changes in the sperm cells exposed to the bacterial extracts were similar to those provoked by exposure to valinomycin and bafilomycin A1, to nigericin, and to oligomycin and ionomycin, respectively. Extracts prepared from Bacillus isolated from the capsule non-specifically depolarised all the cellular transmembrane potentials. The results demonstrate the potential value of a cell toxicity assay with boar spermatozoa for detecting hazardous substances in products intended for human consumption, without whole-animal exposure or using fetal calf serum for cell cultures. PMID- 15651927 TI - Microinjection of living adherent cells by using a semi-automatic microinjection system. AB - Testing in vitro is an alternative to animal experimentation. The capillary pressure microinjection technique is a supporting technology for efficient in vitro testing. The main benefit of the technique is the possibility of injecting large molecules into a single living cell. The ultimate goal of the research discussed in this paper is to increase the cell survival rate in capillary pressure microinjection. A method to reliably evaluate cell survival rate is therefore needed. A three-phase evaluation process is presented in this paper. The first phase determines the success rate of the injection capillary to penetrate the cell membrane. The second phase studies the success rate of delivering the injection substance inside the cell, while the third phase studies cell survival after the microinjection. In addition to the three-phase evaluation process, this paper describes the initial results of penetration and injection tests performed by using a semi-automatic capillary pressure microinjection system developed by the research group. Three adherent cell lines, namely, retinal pigment epithelial cells, MCF-7 human breast cancer cells and SH-SY5Y neuroblastoma cells, were used in the experiments. The results of the penetration tests show that the average success rate of penetrating the cell membrane using the micromanipulator was 87%. The goal of the injection tests was to demonstrate the successful microinjection of living cells and to study the injection success rate. Fluorescein dextran was injected into MCF-7 cells, and preliminary results showed an injection success rate of 49%. In the survival tests, the neuronal cells were microinjected with KCl. During long-term observation after the microinjection, the microinjected cells first decreased their adhesion to the plate, but later adhered to the bottom of the plate and even grew some dendrites. In the next phase of the study, more tests will be performed in order to obtain a statistically reliable value for the survival rate. PMID- 15651929 TI - Status and prospects of in vitro tests in risk assessment. AB - According to the new chemicals policy of the European Union (EU), most chemicals, i.e. the 20,000 chemicals manufactured or imported at 1-10 tons annually, should be tested primarily by using in vitro methods. Also, for other chemicals, the use of in vitro methods is encouraged in the testing strategies given in the draft EU legislation. However, the validation and international acceptance of in vitro tests has been slow. Only recently has the OECD approved four new in vitro test methods, validated by the European Centre for the Validation of Alternative Methods. An analysis of ten randomly selected risk assessment reports of the EU Existing Chemicals Risk Assessment Programme showed that in vitro studies, for example, on cytotoxicity to different cell cultures, cell transformation, metabolism and skin penetration (a total of 115 studies) were used for the assessments. Key metabolic pathways and mechanisms of toxicity have been elucidated, for some chemicals, by using in vitro methods. On the other hand, the results of in vitro studies were regarded as secondary or unreliable in some cases. For several toxic endpoints, in vitro methods will probably serve as screening tools and for mechanistic studies, while target organ toxicity or physiologically regulated adverse effects caused by long-term exposure are difficult to observe without the use of animal models. PMID- 15651928 TI - In vitro methods in the prediction of kinetics of drugs: focus on drug metabolism. AB - The absorption, distribution, metabolism, excretion and toxicity (ADMET) properties of a candidate drug influence its final clinical success. These properties have traditionally been evaluated by using various in vivo animal approaches, but recently, a number of in vitro and in silico methods have been introduced to determine key ADMET features. Basic events, such as absorption through the gut wall, binding to plasma proteins, active and passive transfer through the blood-brain barrier, and various metabolic parameters, can now be screened with rapid in vitro and computer modelling methods. The focus in this short review is on the basic in vitro and in silico methods that are used for studying the metabolism properties of new drug molecules. PMID- 15651930 TI - Strategies to replace in vivo acute systemic toxicity testing. The report and recommendations of ECVAM Workshop 50. PMID- 15651931 TI - Genetics of brain structure and intelligence. AB - Genetic influences on brain morphology and IQ are well studied. A variety of sophisticated brain-mapping approaches relating genetic influences on brain structure and intelligence establishes a regional distribution for this relationship that is consistent with behavioral studies. We highlight those studies that illustrate the complex cortical patterns associated with measures of cognitive ability. A measure of cognitive ability, known as g, has been shown highly heritable across many studies. We argue that these genetic links are partly mediated by brain structure that is likewise under strong genetic control. Other factors, such as the environment, obviously play a role, but the predominant determinant appears to be genetic. PMID- 15651936 TI - Is medicine a "cultural good"? PMID- 15651937 TI - Medical humanities: to cure sometimes, to relieve often, to comfort always. AB - The medical humanities are concerned with "the science of the human", and bring the perspectives of disciplines such as history, philosophy, literature, art and music to understanding health, illness and medicine. The medical humanities are designed to overcome the separation of clinical care from the "human sciences" and to foster interdisciplinary teaching and research to optimise patient care. Medical humanities have become part of the mainstream in medical education in North America and the United Kingdom, and are now integrated into many medical curricula in Australia. The Australasian Association for Medical Humanities was inaugurated in November 2004; a postgraduate program in the medical humanities began at the University of Sydney in 2003. PMID- 15651938 TI - Narrative and illness: the death of a doctor's friend. PMID- 15651939 TI - Safety of emergency medical service helicopters. AB - Robust safety specifications and funding arrangements are needed. PMID- 15651940 TI - Rural inequalities in cancer care and outcome. AB - We need improved primary care, access to expert multidisciplinary services, and appropriate coordination of the two. PMID- 15651941 TI - Immunisation at the crossroads: 9th National Immunisation/1st Asia-Pacific Vaccine Preventable Diseases Conference. PMID- 15651942 TI - Safety of helicopter aeromedical transport in Australia: a retrospective study. AB - OBJECTIVES: To determine the accident rate for Australian helicopter emergency medical services (HEMS) per 100,000 flying hours and to determine the patient mortality risk per mission from a HEMS accident. METHOD: Retrospective observational study of Australian HEMS flying hours and accidents from 1992-2002. RESULTS: The calculated accident rate for Australian HEMS is 4.38 per 100,000 flying hours. One patient died as a direct result of helicopter accident in 50,164 missions. Overall, one accident occurred every 16,721 missions. CONCLUSIONS: The overall Australian HEMS accident rate is similar to that reported from other countries, with all accidents occurring in Queensland community HEMS. Helicopters flown at night under Visual Flight Rules (VFR) appear to represent a high-risk subgroup. HEMS flights do not appear to present significant mortality risk to patients being transported. PMID- 15651943 TI - The effect of a reduction in heroin supply on fatal and non-fatal drug overdoses in New South Wales, Australia. AB - OBJECTIVE: To examine the impact of a sudden and dramatic decrease in heroin availability, concomitant with increases in price and decreases in purity, on fatal and non-fatal drug overdoses in New South Wales, Australia. DESIGN AND SETTING: Time-series analysis was conducted where possible on data on overdoses collected from NSW hospital emergency departments, the NSW Ambulance Service, and all suspected drug-related deaths referred to the NSW Coroner's court. MAIN OUTCOME MEASURES: The number of suspected drug-related deaths where heroin and other drugs were mentioned; ambulance calls to suspected opioid overdoses; and emergency department admissions for overdoses on heroin and other drugs. RESULTS: Both fatal and non-fatal heroin overdoses decreased significantly after heroin supply reduced; the reductions were greater among younger age groups than older age groups. There were no clear increases in non-fatal overdoses with cocaine, methamphetamines or benzodiazepines recorded at hospital emergency departments after the reduction in heroin supply. Data on drug-related deaths suggested that heroin use was the predominant driver of drug-related deaths in NSW, and that when heroin supply was reduced overdose deaths were more likely to involve a wider combination of drugs. CONCLUSION: A reduction in heroin supply reduced heroin-related deaths, and did not result in a concomitant increase, to the same degree, in deaths relating to other drugs. Younger people were more affected by the reduction in supply. PMID- 15651944 TI - Randomised trial of intranasal versus intramuscular naloxone in prehospital treatment for suspected opioid overdose. AB - OBJECTIVE: To determine the effectiveness of intranasal (IN) naloxone compared with intramuscular (IM) naloxone for treatment of respiratory depression due to suspected opiate overdose in the prehospital setting. DESIGN: Prospective, randomised, unblinded trial of either 2 mg naloxone injected intramuscularly or 2 mg naloxone delivered intranasally with a mucosal atomiser. PARTICIPANTS AND SETTING: 155 patients (71 IM and 84 IN) requiring treatment for suspected opiate overdose and attended by paramedics of the Metropolitan Ambulance Service (MAS) and Rural Ambulance Victoria (RAV) in Victoria. MAIN OUTCOME MEASURES: Response time to regain a respiratory rate greater than 10 per minute. Secondary outcome measures were proportion of patients with respiratory rate greater than 10 per minute at 8 minutes and/or a GCS score over 11 at 8 minutes; proportion requiring rescue naloxone; rate of adverse events; proportion of the IN group for whom IN naloxone alone was sufficient treatment. RESULTS: The IM group had more rapid response than the IN group, and were more likely to have more than 10 spontaneous respirations per minute within 8 minutes (82% v 63%; P = 0.0173). There was no statistically significant difference between the IM and IN groups for needing rescue naloxone (13% [IM group] v 26% [IN group]; P = 0.0558). There were no major adverse events. For patients treated with IN naloxone, this was sufficient to reverse opiate toxicity in 74%. CONCLUSION: IN naloxone is effective in treating opiate-induced respiratory depression, but is not as effective as IM naloxone. IN delivery of naxolone could reduce the risk of needlestick injury to ambulance officers and, being relatively safe to make more widely available, could increase access to life-saving treatment in the community. PMID- 15651945 TI - Optimising the management of allergic rhinitis: an Australian perspective. AB - Allergic rhinitis (AR) is one of the most prevalent medical conditions. It has significant effects on quality of life and can have considerable socioeconomic effects. The traditional classification of perennial and seasonal rhinitis does not distinguish between provoking factors, nor does it indicate the most appropriate treatment. A more useful classification is based on symptoms, which may be intermittent or persistent, and vary widely in severity. The goal of management is to achieve optimal symptom control. Therapeutic options include allergen avoidance, pharmacotherapy and immunotherapy. Antihistamines and intranasal corticosteroids (INCS) have become the cornerstones of therapy. A variety of effective treatments are available for consumers to self-select, without the advice of a doctor or pharmacist. INCS are widely recognised as the most effective pharmacotherapy for AR, in both adults and children. The efficacy of various preparations is similar, but those with low systemic bioavailability are preferred for children and for patients who are also receiving inhaled, topical or systemic corticosteroids. PMID- 15651946 TI - From "silos" to seamless healthcare: bringing hospitals and GPs back together again. AB - Linda Mann, a Sydney GP, proposes ways of potentially enhancing patient care and reducing hospital stays, without increasing overall healthcare costs. Her suggested solutions include making it simpler for all healthcare practitioners involved in caring for the same patient to share (rather than not share) all patient information by applying the information technology of today (rather than tomorrow). Also, pooling some funding from state and federal sources could allow for further "seamless care" at areas of reasonable overlap between the current "silos" of hospital-based and community-based care. PMID- 15651947 TI - Hepatitis C-associated cryoglobulinaemia presenting with refractory hypertensive crisis and acute pulmonary oedema. AB - We report two elderly women who presented with hypertensive crisis and acute pulmonary oedema, which responded poorly to antihypertensive therapy. The patients were later diagnosed as having hepatitis C virus-related cryoglobulinaemia. PMID- 15651948 TI - Tramadol and new-onset seizures. PMID- 15651949 TI - Varicella seroprevalence and vaccine uptake in preschool children. PMID- 15651950 TI - Audit of prescribed nicotine replacement therapy to hospital inpatients who smoke. PMID- 15651951 TI - Tissue plasminogen activator (tPA) in acute ischaemic stroke: time for collegiate communication and consensus. PMID- 15651952 TI - Smoking cessation and elective surgery: the cleanest cut. PMID- 15651953 TI - Web and telecounselling in Australia. PMID- 15651954 TI - Are current playground safety standards adequate for preventing arm fractures? PMID- 15651955 TI - Subsidised access to TNF-alpha inhibitors: is the rationale for exclusion of rheumatoid-factor-negative patients defensible? PMID- 15651959 TI - Screening for colorectal cancer: virtually there. AB - A national rollout of faecal occult-blood screening, federally funded, is the best approach. PMID- 15651960 TI - A new integrated vision of how to prevent harmful drug use. AB - The medical community has important roles in reducing harm from alcohol and other drugs. PMID- 15651961 TI - Biobank: who'd bank on it? PMID- 15651962 TI - Barriers to Australian physicians' and paediatricians' involvement in randomised controlled trials. AB - OBJECTIVE: To compare attitudes of Australian physicians and paediatricians about treatment and randomised controlled trial (RCT) participation. DESIGN AND PARTICIPANTS: A cross-sectional survey using the validated "Physician Orientation Profile" (POP), with 250 physicians and 250 paediatricians surveyed. OUTCOME MEASURES: Five indices - primary allegiance, decision making under uncertainty, professional activities, perceived rewards, and peer-group influence - with scores for each participant ranging along a continuum from clinician-oriented to research-oriented and expressed as a number between 0 and 1. RESULTS: Overall response rate was 60%, with 135 physicians (54%) and 165 paediatricians (66%) responding. Paediatricians and physicians were similar in their attitudes to RCT participation, being generally clinician-oriented rather than research-oriented and less inclined to participate in RCTs when there is uncertainty about the best treatment. Most assign limited time to research, with 26.9% not currently involved in research and 31.5% having no experience of RCT participation. Doctors perceive few rewards and little peer-group influence regarding trial participation. Independent predictors of favourable attitudes to trial participation (based on POP scores) were the presence of allocated research time (0.37 for no allocated research time v 0.61 for > 70% research time; P < 0.0001), previous experience enrolling a patient in an RCT (0.40 for no experience v 0.46 for experience; P < 0.0001), and articles published in the past 12 months (0.40 for no publications v 0.55 for > 3 publications; P < 0.0001). CONCLUSIONS: This study highlights the minor importance of research for most Australian physicians. Research plays only a small role in their professional activities, and the importance of research participation is not recognised. They are clinician oriented in their attitudes to RCT participation. To encourage greater involvement in trials among physicians in Australia, clinical research needs to be restructured in a primarily clinically oriented setting with dedicated research time. PMID- 15651963 TI - Is the Framingham coronary heart disease absolute risk function applicable to Aboriginal people? AB - OBJECTIVE: To determine the extent to which the Framingham function predicts the risk of coronary heart disease (CHD) in Aboriginal people. DESIGN AND SETTING: Cohort study in an Aboriginal community in the Northern Territory. PARTICIPANTS: 687 Aboriginal people aged 20-74 years were followed up from a baseline examination in 1992-1995 through to 31 December 2003. MAIN OUTCOME MEASURE: First CHD events were identified through hospital and death records during the follow up period. METHODS: An original Framingham function was used to predict CHD risk according to the duration of follow-up and the values of traditional risk factors, which included age, sex, total cholesterol level, high-density lipoprotein (HDL) cholesterol level, blood pressure, the presence of diabetes, and smoking status. The predicted CHD incidence using the Framingham function was 4.4 per 1000 person-years, while the observed incidence was 11.0 (95% CI, 8.7 13.9) per 1000 person-years. The observed number of CHD events (68) was 2.5 times the number predicted (27) using the Framingham function. The observed incidence was about four and three times the predicted incidence for age groups < 35 and 35 44 years, respectively, and about twice the predicted incidence for those over 45 years of age. The Framingham function was a particularly unreliable predictor for women, especially younger women, in whom the observed CHD rate was 30 times the predicted rate. CONCLUSIONS: The Framingham function substantially underestimates the actual risk of CHD observed in Aboriginal people in a remote community, especially for women and younger adults. This implies that traditional risk factors have different degrees of impact and/or that other factors are contributing to risk. A population-specific risk function is needed. PMID- 15651964 TI - Working with registrars: a qualitative study of interns' perceptions and experiences. AB - OBJECTIVE: To identify and explore behavioural characteristics of registrars that interns find helpful in their working relationships and workplace learning. DESIGN, SETTING AND PARTICIPANTS: Semistructured interviews with 18 interns at Nepean Hospital, Penrith, NSW, at the end of their first working year as doctors. The survey was conducted between December 2003 and February 2004. MAIN OUTCOME MEASURE: Desirable and undesirable behavioural characteristics in registrars, as reported by interns. RESULTS: Overall, interns' opinions of registrars were positive. Desirable characteristics in registrars included approachability, availability, good communication skills, and a willingness to teach. Undesirable characteristics included an unwillingness to listen, unreasonably high expectations, a condescending attitude, apathy and rudeness. CONCLUSION: The behavioural characteristics of registrars that interns find helpful are identifiable, and there is significant room for improvement in the quality of clinical mentoring by registrars. The next step is to facilitate regular feedback from interns on registrars' performance, and to develop ways to encourage desirable behaviours in registrars while actively discouraging undesirable behaviours. PMID- 15651965 TI - A syndromic rash in patients attending methadone clinics in New South Wales. AB - We report an outbreak of a "rash" syndrome in patients attending methadone clinics in New South Wales. It presents with a pruritic, exanthematous or purpuric rash involving the trunk, limbs, palms and soles, which develops over a week and proceeds in most patients to desquamation (mainly of palms and soles) persisting for 3-4 weeks. Mucosae are not involved, and patients are generally systemically well. To date, the rash has affected 22% of 316 patients attending one methadone clinic in western Sydney, as well as patients in clinics elsewhere in Sydney and rural NSW. The aetiology is as yet unknown. PMID- 15651966 TI - A case of desquamating rash associated with methadone use. AB - A man who had been taking prescribed methadone for many years presented with a desquamating rash (predominantly affecting the hands and feet) complicated by cellulitis of the right leg. There have now been multiple reports of a similar rash among methadone users in Sydney. The cause remains unknown. PMID- 15651967 TI - Advances in childhood leukaemia: successful clinical-trials research leads to individualised therapy. AB - In most cases, childhood leukaemia has a fetal origin, but multiple molecular events are required after birth for pre-leukaemic cells to progress to leukaemia. Cure rates for acute lymphoblastic leukaemia (ALL) now approach 80%. A high level of minimal residual disease detected by polymerase chain reaction in patients with ALL in remission has profound prognostic importance and is the focus of a major Australian study attempting to prevent relapse in these children. Greater awareness of the late effects of chemotherapy has led to changes in the treatment protocols for ALL, with improvement in neurocognitive outcomes and reduced rates of second malignancies. Pharmacogenetics is a new field of research that aims to enhance treatment efficacy by assessing the individual's metabolism of and response to chemotherapeutic agents. Targeted therapies currently being developed show some promise of being able to further improve cure rates. Adolescents with ALL have a better prognosis if treated with paediatric rather than adult protocols. PMID- 15651968 TI - Hypertension guidelines, meta-analyses and clinical trials: do we assume too much? AB - Given fundamental differences in the recommendations in guidelines from major national and international committees, we cannot rely on them unquestioningly. Different antihypertensive agents are known to have differing effects according to age and race. Exchanging (rather than following guideline recommendations of adding to) an ineffective first-line antihypertensive drug can result in control of hypertension with monotherapy. Conclusions about a preferable first-line antihypertensive agent are limited by trial protocols with varying drug doses and questionable drug combinations. Guidelines are often based on meta-analyses of drugs of a particular class, which could ignore important differences between drugs within a class. Trials of 3-5 years cannot determine the long-term effects of drugs which patients often take for decades. PMID- 15651969 TI - Does chewing sucrose-free chewing gum after meals reduce the development of carious lesions? PMID- 15651970 TI - Allocation concealment and blinding: when ignorance is bliss. PMID- 15651972 TI - Transoesophageal echocardiography in routine cardiac surgery. PMID- 15651971 TI - Antidepressant use in children: a less depressing story. PMID- 15651973 TI - Postpartum toxic shock syndrome associated with multiple splenic infarcts. PMID- 15651974 TI - Abortion: time to clarify Australia's confusing laws. PMID- 15651975 TI - Throwing the baby out with the spa water? PMID- 15651977 TI - Growth-factor-dependent phosphorylation of Bim in mitosis. AB - The regulation of survival and cell death is a key determinant of cell fate. Recent evidence shows that survival and death machineries are regulated along the cell cycle. In the present paper, we show that BimEL [a BH3 (Bcl-2 homology 3) only member of the Bcl-2 family of proteins; Bim is Bcl-2-interacting mediator of cell death; EL is the extra-long form] is phosphorylated in mitosis. This post translational modification is dependent on MEK (mitogen-activated protein kinase/extracellular-signal-regulated kinase kinase) and growth factor signalling. Interestingly, FGF (fibroblast growth factor) signalling seems to play an essential role in this process, since, in the presence of serum, inhibition of FGF receptors abrogated phosphorylation of Bim in mitosis. Moreover, we have shown bFGF (basic FGF) to be sufficient to induce phosphorylation of Bim in serum-free conditions in any phase of the cell cycle, and also to significantly rescue cells from serum-deprivation-induced apoptosis. Our results show that, in mitosis, Bim is phosphorylated downstream of growth factor signalling in a MEK-dependent manner, with FGF signalling playing an important role. We suggest that phosphorylation of Bim is a decisive step for the survival of proliferating cells. PMID- 15651978 TI - Evidence for co-operativity in coenzyme binding to tetrameric Sulfolobus solfataricus alcohol dehydrogenase and its structural basis: fluorescence, kinetic and structural studies of the wild-type enzyme and non-co-operative N249Y mutant. AB - The interaction of coenzyme with thermostable homotetrameric NAD(H)-dependent alcohol dehydrogenase from the thermoacidophilic sulphur-dependent crenarchaeon Sulfolobus solfataricus (SsADH) and its N249Y (Asn-249-->Tyr) mutant was studied using the high fluorescence sensitivity of its tryptophan residues Trp-95 and Trp 117 to the binding of coenzyme moieties. Fluorescence quenching studies performed at 25 degrees C show that SsADH exhibits linearity in the NAD(H) binding [the Hill coefficient (h) approximately 1) at pH 9.8 and at moderate ionic strength, in addition to positive co-operativity (h=2.0-2.4) at pH 7.8 and 6.8, and at pH 9.8 in the presence of salt. Furthermore, NADH binding is positively co-operative below 20 degrees C (h approximately 3) and negatively co-operative at 40-50 degrees C (h approximately 0.7), as determined at moderate ionic strength and pH 9.8. Steady-state kinetic measurements show that SsADH displays standard Michaelis-Menten kinetics between 35 and 45 degrees C, but exhibits positive and negative co-operativity for NADH oxidation below (h=3.3 at 20 degrees C) and above (h=0.7 at 70-80 degrees C) this range of temperatures respectively. However, N249Y SsADH displays non-co-operative behaviour in coenzyme binding under the same experimental conditions used for the wild-type enzyme. In loop 270 275 of the coenzyme domain and segments at the interface of dimer A-B, analyses of the wild-type and mutant SsADH structures identified the structural elements involved in the intersubunit communication and suggested a possible structural basis for co-operativity. This is the first report of co-operativity in a tetrameric ADH and of temperature-induced co-operativity in a thermophilic enzyme. PMID- 15651979 TI - Experimental optimization of enzymic kinetic resolution of racemic flurbiprofen. AB - Immobilized lipase from Candida antarctica (Novozym 435) was employed for the kinetic resolution of racemic flurbiprofen by the method of enantioselective esterification with alcohols. However, the presence of accumulated water from the esterification influenced the enantiomeric ratio and reaction rate, due to increased rate of hydrolysis of the esterified enantiomer. In the present study, the procedure for optimizing the experimental resolution of the racemate was tested, with a focus on solvent and alcohol types, inhibition of alcohol substrates and the nature of the reversible reaction. The optimal concentration of feed flurbiprofen (180 mM or 44 mg/ml) was determined on basis of the maximum water content favourable for esterification, in single resolution, with the use of methanol in the solvent of cyclohexane. PMID- 15651980 TI - Role of the ERC motif in the proximal part of the second intracellular loop and the C-terminal domain of the human prostaglandin F2alpha receptor (hFP-R) in G protein coupling control. AB - The human FP-R (F2alpha prostaglandin receptor) is a Gq-coupled heptahelical ectoreceptor, which is of significant medical interest, since it is a potential target for the treatment of glaucoma and preterm labour. On agonist exposure, it mediates an increase in intracellular inositol phosphate formation. Little is known about the structures that govern the agonist-dependent receptor activation. In other prostanoid receptors, the C-terminal domain has been inferred in the control of agonist-dependent receptor activation. A DRY motif at the beginning of the second intracellular loop is highly conserved throughout the G-protein coupled receptor family and appears to be crucial for controlling agonist dependent receptor activation. It is replaced by an ERC motif in the FP-R and no evidence for the relevance of this motif in ligand-dependent activation of prostanoid receptors has been provided so far. The aim of the present study was to elucidate the potential role of the C-terminal domain and the ERC motif in agonist-controlled intracellular signalling in FP-R mutants generated by site directed mutagenesis. It was found that substitution of the acidic Glu(132) in the ERC motif by a threonine residue led to full constitutive activation, whereas truncation of the receptor's C-terminal domain led to partial constitutive activation of all three intracellular signal pathways that had previously been shown to be activated by the FP-R, i.e. inositol trisphosphate formation, focal adhesion kinase activation and T-cell factor signalling. Inositol trisphosphate formation and focal adhesion kinase phosphorylation were further enhanced by ligand binding in cells expressing the truncation mutant but not the E132T (Glu132-->Thr) mutant. Thus C-terminal truncation appeared to result in a receptor with partial constitutive activation, whereas substitution of Glu132 by threonine apparently resulted in a receptor with full constitutive activity. PMID- 15651981 TI - Negative autoregulation of HNF-4alpha gene expression by HNF-4alpha1. AB - HNF-4alpha (hepatocyte nuclear factor-4alpha) is required for tissue-specific expression of many of the hepatic, pancreatic, enteric and renal traits. Heterozygous HNF-4alpha mutants are inflicted by MODY-1 (maturity onset diabetes of the young type-1). HNF-4alpha expression is reported here to be negatively autoregulated by HNF-4alpha1 and to be activated by dominant-negative HNF 4alpha1. Deletion and chromatin immunoprecipitation analysis indicated that negative autoregulation by HNF-4alpha1 was mediated by its association with the TATA-less HNF-4alpha core promoter enriched in Sp1, but lacking DR-1 response elements. Also, negative autoregulation by HNF-4alpha1 was independent of its transactivation function, being similarly exerted by transcriptional-defective MODY-1 missense mutants of HNF-4alpha1, or under conditions of suppressing or enhancing HNF-4alpha activity by small heterodimer partner or by inhibiting histone deacetylase respectively. Negative autoregulation by HNF-4alpha1 was abrogated by overexpressed Sp1. Transcriptional suppression by HNF-4alpha1 independently of its transactivation function may extend the scope of its transcriptional activity to interference with docking of the pre-transcriptional initiation complex to TATA-less promoters. PMID- 15651982 TI - Sodium-coupled and electrogenic transport of B-complex vitamin nicotinic acid by slc5a8, a member of the Na/glucose co-transporter gene family. AB - SMCT (sodium-coupled monocarboxylate transporter; slc5a8) is a Na+-coupled transporter for lactate, pyruvate and short-chain fatty acids. Similar to these already known substrates of SMCT, the water-soluble B-complex vitamin nicotinic acid also exists as a monocarboxylate anion (nicotinate) under physiological conditions. Therefore we evaluated the ability of SMCT to mediate the uptake of nicotinate. In mammalian cells, the cloned mouse SMCT (slc5a8) induced the uptake of nicotinate. The SMCT-induced uptake was Na+-dependent. The Michaelis constant for the uptake process was 296+/-88 microM. The Na+-activation kinetics indicated that at least two Na+ ions are involved in the process. Among the various structural analogues tested, nicotinate was the most effective substrate. Nicotinamide and methylnicotinate were not recognized by the transporter. 2 pyrazine carboxylate and isonicotinate interacted with the transporter to a moderate extent. SMCT-mediated uptake of nicotinate was inhibited by lactate and pyruvate. In the Xenopus laevis oocyte expression system, SMCT-mediated nicotinate transport was electrogenic, as evident from the nicotinate-induced inward currents under voltage-clamp conditions. Substrate-induced currents in this expression system corroborated the substrate specificity determined in the mammalian cell expression system. The kinetic parameters with regard to the affinity of the transporter for nicotinate and the Hill coefficient for Na+ activation, determined by using the oocyte expression system, were also similar to those obtained from the mammalian cell expression system. We conclude that SMCT functions not only as a Na+-coupled transporter for short-chain fatty acids and lactate but also as a Na+-coupled transporter for the water-soluble vitamin nicotinic acid. PMID- 15651984 TI - Sister Joseph's nodule in a liver transplant recipient: Case report and mini review of literature. AB - BACKGROUND: Umbilical metastasis is one of the main characteristic signs of extensive neoplastic disease and is universally referred to as Sister Mary Joseph's nodule. CASE PRESENTATION: A 59-years-old Caucasian female underwent liver transplant for end stage liver disease due to hepatitis C with whole graft from cadaveric donor in 2003. After transplantation the patient developed multiple subcutaneous nodules in the umbilical region and bilateral inguinal lymphadenopathy. The excision biopsy of the umbilical mass showed the features of a poorly differentiated papillary serous cystadenocarcinoma. Computed tomographic scan and transvaginal ultrasonography were unable to demonstrate any primary lesion. Chemotherapy was start and the dosage of the immunosuppressive drugs was reduced. To date the patient is doing well and liver function is normal. CONCLUSIONS: The umbilical metastasis can arise from many sites. In some cases, primary tumor may be not identified; nonetheless chemotherapy must be administrated based on patient's history, anatomical and histological findings. PMID- 15651983 TI - Lsb5p interacts with actin regulators Sla1p and Las17p, ubiquitin and Arf3p to couple actin dynamics to membrane trafficking processes. AB - The importance of coupling the process of endocytosis to factors that regulate actin dynamics has been clearly demonstrated in yeast, and many proteins involved in these mechanisms have been identified. Sla1p is a well-characterized yeast protein that binds both to activators of actin dynamics, Las17p and Pan1p, and to cargo proteins, such as the pheromone receptor Ste2p. Previously, we reported that the Lsb5 protein plays a role in endocytosis in yeast and that it localizes to the plasma membrane. Lsb5p has a similar structure to the GGA [Golgi localized, gamma-ear-containing, Arf (ADP-ribosylation factor)-binding] family of proteins with an N-terminal VHS [Vps27p (vacuolar protein sorting protein 27), Hrs, Stam] domain and a GAT (GGA and Tom1) domain. It does not, however, contain either a gamma-adaptin ear or a clathrin-binding motif. In the present study, we have further defined its interaction site with both Sla1p and with Las17p, two regulators of actin dynamics. The site of interaction with Sla1p involves the Sla1 HD1 (homology domain 1), which also was shown previously to interact with the pheromone receptor Ste2p. We also demonstrate hitherto unknown interactions between Lsb5p and the active form of the yeast Arf3 protein, and with ubiquitin. Finally, we demonstrate a requirement for Arf3p expression in order to localize Lsb5p to the correct cortical site in cells. Taken together, our data provide further evidence for the role of Lsb5p in membrane-trafficking events at the plasma membrane and also demonstrate for the first time an interaction of Arf3 with the endocytic machinery in yeast. PMID- 15651985 TI - Patient involvement in medical decision-making and pain among elders: physician or patient-driven? AB - BACKGROUND: Pain is highly prevalent among older adults, but little is known about how patient involvement in medical decision-making may play a role in limiting its occurrence or severity. The purpose of this study was to evaluate whether physician-driven and patient-driven participation in decision-making were associated with the odds of frequent and severe pain. METHODS: A cross-sectional population-based survey of 3,135 persons age 65 and older was conducted in the 108-county region comprising West Texas. The survey included self-reports of frequent pain and, among those with frequent pain, the severity of pain. RESULTS: Findings from multivariate logistic regression analyses showed that higher patient-driven participation in decision-making was associated with lower odds (OR, 0.82; 95% CI, 0.75-0.89) of frequent pain, but was not significantly associated with severe pain. Physician-driven participation was not significantly associated with frequent or severe pain. CONCLUSIONS: The findings suggest that patients may need to initiate involvement in medical decision-making to reduce their chances of experiencing frequent pain. Changes to other modifiable health care characteristics, including access to a personal doctor and health insurance coverage, may be more conducive to limiting the risk of severe pain. PMID- 15651987 TI - The burden of non communicable diseases in developing countries. AB - BACKGROUND: By the dawn of the third millennium, non communicable diseases are sweeping the entire globe, with an increasing trend in developing countries where, the transition imposes more constraints to deal with the double burden of infective and non-infective diseases in a poor environment characterised by ill health systems. By 2020, it is predicted that these diseases will be causing seven out of every 10 deaths in developing countries. Many of the non communicable diseases can be prevented by tackling associated risk factors. METHODS: Data from national registries and international organisms are collected, compared and analyzed. The focus is made on the growing burden of non communicable diseases in developing countries. RESULTS: Among non communicable diseases, special attention is devoted to cardiovascular diseases, diabetes, cancer and chronic pulmonary diseases. Their burden is affecting countries worldwide but with a growing trend in developing countries. Preventive strategies must take into account the growing trend of risk factors correlated to these diseases. CONCLUSION: Non communicable diseases are more and more prevalent in developing countries where they double the burden of infective diseases. If the present trend is maintained, the health systems in low-and middle-income countries will be unable to support the burden of disease. Prominent causes for heart disease, diabetes, cancer and pulmonary diseases can be prevented but urgent (preventive) actions are needed and efficient strategies should deal seriously with risk factors like smoking, alcohol, physical inactivity and western diet. PMID- 15651986 TI - Irradiation specifically sensitises solid tumour cell lines to TRAIL mediated apoptosis. AB - BACKGROUND: TRAIL (tumor necrosis factor related apoptosis inducing ligand) is an apoptosis inducing ligand with high specificity for malignant cell systems. Combined treatment modalities using TRAIL and cytotoxic drugs revealed highly additive effects in different tumour cell lines. Little is known about the efficacy and underlying mechanistic effects of a combined therapy using TRAIL and ionising radiation in solid tumour cell systems. Additionally, little is known about the effect of TRAIL combined with radiation on normal tissues. METHODS: Tumour cell systems derived from breast- (MDA MB231), lung--(NCI H460) colorectal -(Colo 205, HCT-15) and head and neck cancer (FaDu, SCC-4) were treated with a combination of TRAIL and irradiation using two different time schedules. Normal tissue cultures from breast, prostate, renal and bronchial epithelia, small muscle cells, endothelial cells, hepatocytes and fibroblasts were tested accordingly. Apoptosis was determined by fluorescence microscopy and western blot determination of PARP processing. Upregulation of death receptors was quantified by flow cytometry. RESULTS: The combined treatment of TRAIL with irradiation strongly increased apoptosis induction in all treated tumour cell lines compared to treatment with TRAIL or irradiation alone. The synergistic effect was most prominent after sequential application of TRAIL after irradiation. Upregulation of TRAIL receptor DR5 after irradiation was observed in four of six tumour cell lines but did not correlate to tumour cell sensitisation to TRAIL. TRAIL did not show toxicity in normal tissue cell systems. In addition, pre-irradiation did not sensitise all nine tested human normal tissue cell cultures to TRAIL. CONCLUSIONS: Based on the in vitro data, TRAIL represents a very promising candidate for combination with radiotherapy. Sequential application of ionising radiation followed by TRAIL is associated with an synergistic induction of cell death in a large panel of solid tumour cell lines. However, TRAIL receptor upregulation may not be the sole mechanism by which sensitation to TRAIL after irradiation is induced. PMID- 15651989 TI - Selection and characterization of a promoter for expression of single-copy recombinant genes in Gram-positive bacteria. AB - BACKGROUND: In the past ten years there has been a growing interest in engineering Gram-positive bacteria for biotechnological applications, including vaccine delivery and production of recombinant proteins. Usually, bacteria are manipulated using plasmid expression vectors. The major limitation of this approach is due to the fact that recombinant plasmids are often lost from the bacterial culture upon removal of antibiotic selection. We have developed a genetic system based on suicide vectors on conjugative transposons allowing stable integration of recombinant DNA into the chromosome of transformable and non-transformable Gram-positive bacteria. RESULTS: The aim of this work was to select a strong chromosomal promoter from Streptococcus gordonii to improve this genetic system making it suitable for expression of single-copy recombinant genes. To achieve this task, a promoterless gene encoding a chloramphenicol acetyltransferase (cat), was randomly integrated into the S. gordonii chromosome and transformants were selected for chloramphenicol resistance. Three out of eighteen chloramphenicol resistant transformants selected exhibited 100% stability of the phenotype and only one of them, GP215, carried the cat gene integrated as a single copy. A DNA fragment of 600 base pairs exhibiting promoter activity was isolated from GP215 and sequenced. The 5' end of its corresponding mRNA was determined by primer extention analysis and the putative -10 and a -35 regions were identified. To study the possibility of using this promoter (PP) for single copy heterologous gene expression, we created transcriptional fusions of PP with genes encoding surface recombinant proteins in a vector capable of integrating into the conjugative transposon Tn916. Surface recombinant proteins whose expression was controlled by the PP promoter were detected in Tn916 containing strains of S. gordonii and Bacillus subtilis after single copy chromosomal integration of the recombinant insertion vectors into the resident Tn916. The surface recombinant protein synthesized under the control of PP was also detected in Enterococcus faecalis after conjugal transfer of a recombinant Tn916 containing the transcriptional fusion. CONCLUSION: We isolated and characterized a S. gordonii chromosomal promoter. We demonstrated that this promoter can be used to direct expression of heterologous genes in different Gram positive bacteria, when integrated in a single copy into the chromosome. PMID- 15651991 TI - Simultaneous development of the Pediatric GERD Caregiver Impact Questionnaire (PGCIQ) in American English and American Spanish. AB - BACKGROUND: The objective of this study was to develop simultaneously a new questionnaire, the Pediatric GERD Caregiver Impact Questionnaire (PGCIQ), in American English and American Spanish in order to elucidate the impact of caring for a child with GERD. METHODS: Two focus group discussions were conducted in American English and American Spanish to develop a relevant conceptual model. Focus group participants were the primary caregivers of children with GERD (newborn through 12 years of age). Participant responses were qualitatively analyzed to identify potential differences in caregiver perspectives by the caregiver's language, socio-economic status and demographic profile as well as the child's age and disease severity level. Items in the PGCIQ were generated simultaneously in English and Spanish by reviewing results of qualitative analysis from focus groups in each language. The PGCIQ was finalized in both languages after testing content validity and conducting an in-depth translatability assessment. RESULTS: Analysis of focus group comments resulted in the development of a first draft questionnaire consisting of 58 items in 10 domains. Content validity testing and an in-depth translatability assessment resulted in wording modification of 37 items, deletion of 14 items and the addition of a domain with five items. Feedback from the content validity testing interviews indicated that the instrument is conceptually relevant in both American English and American Spanish, clear, comprehensive and easy to complete within 10 minutes. The final version of the PGCIQ contains 49 items assessing ten domains. An optional module with nine items is available for investigative research purposes and for use only at baseline. CONCLUSION: The PGCIQ was developed using simultaneous item generation, a process that allows for consideration of concept relevance in all stages of development and in all languages being developed. The PGCIQ is the first questionnaire to document the multidimensional impact of caring for an infant or young child with GERD. Linguistic adaptation of the PGCIQ in multiple languages is ongoing. A validation study of the PGCIQ is needed to examine its psychometric properties, further refine the items and develop an appropriate scoring model. PMID- 15651988 TI - Gene expression patterns associated with blood-feeding in the malaria mosquito Anopheles gambiae. AB - BACKGROUND: Blood feeding, or hematophagy, is a behavior exhibited by female mosquitoes required both for reproduction and for transmission of pathogens. We determined the expression patterns of 3,068 ESTs, representing ~2,000 unique gene transcripts using cDNA microarrays in adult female Anopheles gambiae at selected times during the first two days following blood ingestion, at 5 and 30 min during a 40 minute blood meal and at 0, 1, 3, 5, 12, 16, 24 and 48 hours after completion of the blood meal and compared their expression to transcript levels in mosquitoes with access only to a sugar solution. RESULTS: In blood-fed mosquitoes, 413 unique transcripts, approximately 25% of the total, were expressed at least two-fold above or below their levels in the sugar-fed mosquitoes, at one or more time points. These differentially expressed gene products were clustered using k-means clustering into Early Genes, Middle Genes, and Late Genes, containing 144, 130, and 139 unique transcripts, respectively. Several genes from each group were analyzed by quantitative real-time PCR in order to validate the microarray results. CONCLUSION: The expression patterns and annotation of the genes in these three groups (Early, Middle, and Late genes) are discussed in the context of female mosquitoes' physiological responses to blood feeding, including blood digestion, peritrophic matrix formation, egg development, and immunity. PMID- 15651990 TI - Effectiveness and cost-effectiveness of a multidisciplinary intervention programme to prevent new falls and functional decline among elderly persons at risk: design of a replicated randomised controlled trial [ISRCTN64716113]. AB - BACKGROUND: Falls are common among community-dwelling elderly people and can have a considerable impact on quality of life and healthcare costs. People who have sustained a fall are at greater risk of falling again.We replicated a British randomised controlled trial which demonstrated the effectiveness of a multidisciplinary intervention programme to prevent falls. The objective is to describe the design of a replication study evaluating a multidisciplinary intervention programme on recurrent falls and functional decline among elderly persons at risk. The study consists of an effect evaluation, an economic evaluation and a process evaluation. METHODS/DESIGN: The programme is aimed at community-dwelling elderly people aged 65 years or over who have visited an accident and emergency department (A&E department) or a general practitioners' cooperative (GP cooperative) because of a fall. The design involves a two-group randomised controlled trial. Participants are followed for twelve months after baseline. The intervention programme consists of a detailed medical and occupational therapy assessment with referral to relevant services if indicated. People in the control group receive usual care. The main outcome measures of the effect evaluation are number of falls and daily functioning. The economic evaluation will be performed from a societal perspective. A process evaluation will be carried out to evaluate the feasibility of the intervention programme. PMID- 15651992 TI - Variants in the vitamin D receptor gene and asthma. AB - BACKGROUND: Early lifetime exposure to dietary or supplementary vitamin D has been predicted to be a risk factor for later allergy. Twin studies suggest that response to vitamin D exposure might be influenced by genetic factors. As these effects are primarily mediated through the vitamin D receptor (VDR), single base variants in this gene may be risk factors for asthma or allergy. RESULTS: 951 individuals from 224 pedigrees with at least 2 asthmatic children were analyzed for 13 SNPs in the VDR. There was no preferential transmission to children with asthma. In their unaffected sibs, however, one allele in the 5' region was 0.5 fold undertransmitted (p = 0.049), while two other alleles in the 3' terminal region were 2-fold over-transmitted (p = 0.013 and 0.018). An association was also seen with bronchial hyperreactivity against methacholine and with specific immunoglobulin E serum levels. CONCLUSION: The transmission disequilibrium in unaffected sibs of otherwise multiple-affected families seem to be a powerful statistical test. A preferential transmission of vitamin D receptor variants to children with asthma could not be confirmed but raises the possibility of a protective effect for unaffected children. PMID- 15651994 TI - Explaining inconsistencies between data on condom use and condom sales. AB - BACKGROUND: Several HIV prevention programs use data on condom sales and survey based data on condom prevalence to monitor progress. However, such indicators are not always consistent. This paper aims to explain these inconsistencies and to assess whether the number of sex acts and the number of condoms used can be estimated from survey data. This would be useful for program managers, as it would enable estimation of the number of condoms needed for different target groups. METHODS: We use data from six Demographic and Health Surveys to estimate the total annual number of sex acts and number of condoms used. Estimates of the number of sex acts are based on self-reported coital frequency, the proportion reporting intercourse the previous day, and survival methods. Estimates of the number of condoms used are based on self-reported frequency of use, the proportion reporting condom use the previous day and in last intercourse. The estimated number of condoms used is then compared with reported data on condom sales and distribution. RESULTS: Analysis of data on the annual number of condoms sold and distributed to the trade reveals very erratic patterns, which reflect stock-ups at various levels in the distribution chain. Consequently, condom sales data are a very poor indicator of the level of condom use. Estimates of both the number of sexual acts and the number of condoms used vary enormously based on the estimation method used. For several surveys, the highest estimate of the annual number of condoms used is tenfold that of the lowest estimate. CONCLUSIONS: Condom sales to the trade are a poor indicator of levels of condom use, and are therefore insufficient to monitor HIV prevention programs. While survey data on condom prevalence allow more detailed monitoring, converting such data to an estimated number of sex acts and condoms used is not straightforward. The estimation methods yield widely different results, and it is impossible to determine which method is most accurate. Until the reliability of these various estimation methods can be established, estimating the annual number of condoms used from survey data will not be feasible. Collecting survey data on the number of sex acts and the number of condoms used in a fixed time period may enable the calculation of more reliable estimates of the number of sex acts and condoms used. PMID- 15651995 TI - Validating viral quasispecies with digital organisms: a re-examination of the critical mutation rate. AB - BACKGROUND: In this report we re-examine some recent experiments with digital organisms to test some predictions of quasispecies theory. These experiments revealed that under high mutation rates populations of less fit organisms previously adapted to such high mutation rates were able to outcompete organisms with higher average fitness but adapted to low mutation rates. RESULTS: We have verified that these results do hold in the original conditions and, by extending the set of initial parameters, we have also detected that the critical mutation rate was independent of population size, a result that we have found to be dependent on a different, contingent factor, the initial fitness vector. Furthermore, in all but one case, the critical mutation rate is higher than the error threshold, a key parameter in quasispecies theory, which prevents its extrapolation to natural viral populations. CONCLUSION: From these results we conclude that digital organisms are useful tools for investigating evolutionary patterns and processes including some predictions from the quasispecies theory. PMID- 15651996 TI - General practitioners' reasoning when considering the diagnosis heart failure: a think-aloud study. AB - BACKGROUND: Diagnosing chronic heart failure is difficult, especially in mild cases or early in the course of the disease, and guidelines are not easily implemented in everyday practice. The aim of this study was to investigate general practitioners' diagnostic reasoning about patients with suspected chronic heart failure in comparison with recommendations in European guidelines. METHODS: Think-aloud technique was used. Fifteen general practitioners reasoned about six case vignettes, representing authentic patients with suspected chronic heart failure. Information about each case was added successively in five steps. The general practitioners said their thoughts aloud while reasoning about the probability of the patient having chronic heart failure, and tried to decide about the diagnosis. Arguments for and against chronic heart failure were analysed and compared to recommendations in guidelines. RESULTS: Information about ejection fraction was the most frequent diagnostic argument, followed by information about cardiac enlargement or pulmonary congestion on chest X-ray. However, in a third of the judgement situations, no information about echocardiography was utilized in the general practitioners' diagnostic reasoning. Only three of the 15 doctors used information about a normal electrocardiography as an argument against chronic heart failure. Information about other cardio vascular diseases was frequently used as a diagnostic argument. CONCLUSIONS: The clinical information was not utilized to the extent recommended in guidelines. Some implications of our study are that 1) general practitioners need more information about how to utilize echocardiography when diagnosing chronic heart failure, 2) guidelines ought to give more importance to information about other cardio-vascular diseases in the diagnostic reasoning, and 3) guidelines ought to treat the topic of diastolic heart failure in a clearer way. PMID- 15651993 TI - Estrogen, mitochondria, and growth of cancer and non-cancer cells. AB - In this review, we discuss estrogen actions on mitochondrial function and the possible implications on cell growth. Mitochondria are important targets of estrogen action. Therefore, an in-depth analysis of interaction between estrogen and mitochondria; and mitochondrial signaling to nucleus are pertinent to the development of new therapy strategies for the treatment of estrogen-dependent diseases related to mitochondrial disorders, including cancer. PMID- 15651997 TI - Patient satisfaction with out-of-hours primary care in the Netherlands. AB - BACKGROUND: In recent years out-of-hours primary care in the Netherlands has changed from practice-based to large-scale cooperatives. The purpose of this study is to determine patient satisfaction with current out-of-hours care organised in general practitioner (GP) cooperatives, and gain insight in factors associated with this satisfaction. METHODS: From March to June 2003, 2805 questionnaires were sent to patients within three weeks after they had contacted the GP cooperative in their region. The study was conducted in the province of Limburg in the South of the Netherlands. One-third of these questionnaires was sent to patients who had only received telephone advice, one-third to patients who attended the GP cooperative for consultation, and one-third to patients who received a home visit. Four weeks after the first reminder, a non-respondents telephone interview was performed among a random sample of 100 patients. Analyses were performed with respect to the type of consultation. RESULTS: The total response was 42.4% (1160/2733). Sixty-seven percent of patients who received telephone advice only reported to be satisfied with out-of-hours care. About 80% of patients who went to the GP cooperative for consultation or those receiving a home visit, reported to be satisfied. Factors that were strongly associated with overall satisfaction included, the doctor's assistant's attitude on the phone, opinion on GP's treatment, and waiting time. CONCLUSION: Patients seem generally satisfied with out-of-hours primary care as organised in GP cooperatives. However, patients who received telephone advice only are less satisfied compared to those who attended the GP cooperative or those who received a home visit. PMID- 15651998 TI - Gene expression profiling of tumours derived from rasV12/E1A-transformed mouse embryonic fibroblasts to identify genes required for tumour development. AB - BACKGROUND: In cancer, cellular transformation is followed by tumour development. Knowledge on the mechanisms of transformation, involving activation of proto oncogenes and inactivation of tumour-suppressor genes has considerably improved whereas tumour development remains poorly understood. An interesting way of gaining information on tumour progression mechanisms would be to identify genes whose expression is altered during tumour formation. We used the Affymetrix-based DNA microarray technology to analyze gene expression profiles of tumours derived from rasV12/E1A-transformed mouse embryo fibroblasts in order to identify the genes that could be involved in tumour development. RESULTS: Among the 12,000 genes analyzed in this study, only 489 showed altered expression during tumour development, 213 being up-regulated and 276 down-regulated. The genes differentially expressed are involved in a variety of cellular functions, including control of transcription, regulation of mRNA maturation and processing, regulation of protein translation, activation of interferon-induced genes, intracellular signalling, apoptosis, cell growth, angiogenesis, cytoskeleton, cell-to-cell interaction, extracellular matrix formation, metabolism and production of secretory factors. CONCLUSIONS: Some of the genes identified in this work, whose expression is altered upon rasV12/E1A transformation of MEFs, could be new cancer therapeutic targets. PMID- 15651999 TI - Epimorphin expression in interstitial pneumonia. AB - Epimorphin modulates epithelial morphogenesis in embryonic mouse organs. We previously suggested that epimorphin contributes to repair of bleomycin-induced pulmonary fibrosis in mice via epithelium-mesenchyme interactions. To clarify the role of epimorphin in human lungs, we evaluated epimorphin expression and localization in normal lungs, lungs with nonspecific interstitial pneumonia (NSIP), and lungs with usual interstitial pneumonia (UIP); we also studied the effect of recombinant epimorphin on cultured human alveolar epithelial cells in vitro. Northern and Western blotting analyses revealed that epimorphin expression in NSIP samples were significantly higher than those in control lungs and lungs with UIP. Immunohistochemistry showed strong epimorphin expression in mesenchymal cells of early fibrotic lesions and localization of epimorphin protein on mesenchymal cells and extracellular matrix of early fibrotic lesions in the nonspecific interstitial pneumonia group. Double-labeled fluorescent images revealed expression of matrix metalloproteinase 2 in re-epithelialized cells overlying epimorphin-positive early fibrotic lesions. Immunohistochemistry and metalloproteinase activity assay demonstrated augmented expression of metalloproteinase induced by recombinant epimorphin in human alveolar epithelial cells. These findings suggest that epimorphin contributes to repair of pulmonary fibrosis in nonspecific interstitial pneumonia, perhaps partly by inducing expression of matrix metalloproteinase 2, which is an important proteolytic factor in lung remodeling. PMID- 15652000 TI - Single nucleotide polymorphisms (SNPs) are inherited from parents and they measure heritable events. AB - Single nucleotide polymorphisms (SNPs) are extensively used in case-control studies of practically all cancer types. They are used for the identification of inherited cancer susceptibility genes and those that may interact with environmental factors. However, being genetic markers, they are applicable only on heritable conditions, which is often a neglected fact. Based on the data in the nationwide Swedish Family-Cancer Database, we review familial risks for all main cancers and discuss the evidence for a heritable component in cancer. The available evidence is not conclusive but it is consistent in pointing to a minor heritable etiology in cancer, which will hamper the success of SNP-based association studies. Empirical familial risks should be used as guidance for the planning of SNP studies. We provide calculations for the assessment of familial risks for assumed allele frequencies and gene effects (odds ratios) for different modes of inheritance. Based on these data, we discuss the gene effects that could account for the unexplained proportion of familial breast and lung cancer. As a conclusion, we are concerned about the indiscriminate use of a genetic tool to cancers, which are mainly environmental in origin. We consider the likelihood of a successful application of SNPs in gene-environment studies small, unless established environmental risk factors are tested on proven candidate genes. PMID- 15652001 TI - Atrial fibrillation (acute). PMID- 15652002 TI - Heart failure. PMID- 15652003 TI - Peripheral arterial disease. PMID- 15652004 TI - Stroke management. PMID- 15652005 TI - Varicose veins. PMID- 15652006 TI - Absence seizures in children. PMID- 15652007 TI - Acute otitis media. PMID- 15652008 TI - Asthma and other wheezing disorders in infants and children. PMID- 15652009 TI - Cardiorespiratory arrest in children. PMID- 15652010 TI - Constipation in children. PMID- 15652011 TI - Depression in children and adolescents. PMID- 15652012 TI - Gastro-oesophageal reflux in children. PMID- 15652013 TI - Infantile colic. PMID- 15652014 TI - Measles (prevention). PMID- 15652015 TI - Migraine headache in children. PMID- 15652016 TI - Neonatal jaundice. PMID- 15652017 TI - Nocturnal enuresis. PMID- 15652018 TI - Urinary tract infection in children. PMID- 15652019 TI - Acute cholecystitis. PMID- 15652020 TI - Anal fissure. PMID- 15652021 TI - Appendicitis. PMID- 15652022 TI - Colonic diverticular disease. PMID- 15652023 TI - Constipation in adults. PMID- 15652024 TI - Gastro-oesophageal reflux disease. PMID- 15652025 TI - Irritable bowel syndrome. PMID- 15652026 TI - Pancreatic cancer. PMID- 15652027 TI - Meniere's disease. PMID- 15652028 TI - Middle ear pain and trauma during air travel. PMID- 15652029 TI - Otitis externa. PMID- 15652030 TI - Otitis media with effusion. PMID- 15652031 TI - Seasonal allergic rhinitis. PMID- 15652032 TI - Sinusitis (acute). PMID- 15652033 TI - Tinnitus. PMID- 15652034 TI - Tonsillitis. PMID- 15652035 TI - Foot ulcers and amputations in diabetes. PMID- 15652036 TI - Obesity. PMID- 15652037 TI - Prevention of cardiovascular events in diabetes. PMID- 15652039 TI - Cataract. PMID- 15652038 TI - Age related macular degeneration. PMID- 15652040 TI - Diabetic retinopathy. PMID- 15652041 TI - Glaucoma. PMID- 15652042 TI - Ocular herpes simplex. PMID- 15652043 TI - Trachoma. PMID- 15652044 TI - HIV infection. PMID- 15652045 TI - HIV: mother to child transmission. PMID- 15652046 TI - Pneumocystis carinii pneumonia treatment in people infected with HIV. PMID- 15652047 TI - Chickenpox. PMID- 15652048 TI - Dengue fever. PMID- 15652049 TI - Diarrhoea in adults (acute). PMID- 15652050 TI - Hepatitis B (prevention). PMID- 15652051 TI - Malaria: prevention in travellers. PMID- 15652052 TI - Malaria: severe, life threatening. PMID- 15652053 TI - Tuberculosis. PMID- 15652054 TI - Acute renal failure. PMID- 15652055 TI - Benign prostatic hyperplasia. PMID- 15652056 TI - Erectile dysfunction. PMID- 15652057 TI - Varicocele. PMID- 15652058 TI - Anorexia nervosa. PMID- 15652059 TI - Dementia. PMID- 15652060 TI - Generalised anxiety disorder. PMID- 15652061 TI - Obsessive compulsive disorder. PMID- 15652062 TI - Panic disorder. PMID- 15652063 TI - Post-traumatic stress disorder. PMID- 15652064 TI - Ankle sprain. PMID- 15652065 TI - Bunions. PMID- 15652066 TI - Gout. PMID- 15652067 TI - Herniated lumbar disc. PMID- 15652068 TI - Leg cramps. PMID- 15652070 TI - Musculoskeletal disorders. Non-steroidal anti-inflammatory drugs. PMID- 15652069 TI - Neck pain. PMID- 15652071 TI - Plantar heel pain and fasciitis. PMID- 15652072 TI - Shoulder pain. PMID- 15652073 TI - Tennis elbow. PMID- 15652074 TI - Altitude sickness. PMID- 15652075 TI - Essential tremor. PMID- 15652076 TI - Migraine headache. PMID- 15652077 TI - Parkinson's disease. PMID- 15652078 TI - Aphthous ulcers (recurrent). PMID- 15652079 TI - Burning mouth syndrome. PMID- 15652080 TI - Impacted wisdom teeth. PMID- 15652081 TI - Postoperative pulmonary infections. PMID- 15652082 TI - Paracetamol (acetaminophen) poisoning. PMID- 15652083 TI - Ectopic pregnancy. PMID- 15652084 TI - Nausea and vomiting in early pregnancy. PMID- 15652085 TI - Perineal care. PMID- 15652086 TI - Postnatal depression. PMID- 15652087 TI - Pre-eclampsia and hypertension. PMID- 15652088 TI - Preterm birth. PMID- 15652089 TI - Bronchitis (acute). PMID- 15652090 TI - Community acquired pneumonia. PMID- 15652091 TI - Spontaneous pneumothorax. PMID- 15652092 TI - Asthma. PMID- 15652093 TI - Bronchiectasis. PMID- 15652094 TI - Chronic obstructive pulmonary disease. PMID- 15652095 TI - Lung cancer. PMID- 15652096 TI - Bacterial vaginosis. PMID- 15652097 TI - Chlamydia (uncomplicated, genital). PMID- 15652098 TI - Genital herpes. PMID- 15652099 TI - Genital warts. PMID- 15652100 TI - Gonorrhoea. PMID- 15652101 TI - Partner notification. PMID- 15652102 TI - Pelvic inflammatory disease. PMID- 15652103 TI - Athlete's foot. PMID- 15652104 TI - Cellulitis and erysipelas. PMID- 15652105 TI - Chronic plaque psoriasis. PMID- 15652106 TI - Head lice. PMID- 15652107 TI - Herpes labialis. PMID- 15652108 TI - Warts. PMID- 15652109 TI - Insomnia. PMID- 15652110 TI - Jet lag. PMID- 15652111 TI - Breast cancer (metastatic). PMID- 15652112 TI - Breast cancer (non-metastatic). PMID- 15652113 TI - Breast pain. PMID- 15652114 TI - Domestic violence towards women. PMID- 15652115 TI - Endometriosis. PMID- 15652116 TI - Fibroids (uterine myomatosis, leiomyomas). PMID- 15652117 TI - Infertility and subfertility. PMID- 15652118 TI - Menopausal symptoms. PMID- 15652119 TI - Ovarian cancer. PMID- 15652120 TI - Pyelonephritis in non-pregnant women. PMID- 15652121 TI - Stress incontinence. PMID- 15652122 TI - Bites (mammalian). PMID- 15652123 TI - Pressure sores. PMID- 15652140 TI - GENE impedance: a natural process for control of gene expression and the origin of RNA interference. AB - Gene expression is controlled by coordinated transcriptional and post transcriptional mechanisms. Normally, expression of a gene switches on and off in response to specific physiological signals that are triggered by cellular demand for the gene products at a given time. Based on our previous studies and the scientific literature, we hypothesize that when a gene promoter switches to transcriptional repression mode, transcription of the gene ceases, and a small amount of double-stranded RNA (dsRNA) is synthesized by the RNA polymerase switching to the opposite DNA strand at the termination region of the gene. These dsRNA structures, which result from normal transcriptional repression, can then be processed into short interfering RNAs (siRNAs) within the nucleus. These molecules subsequently direct specific cleavage of the cognate mRNAs and interfere with their translation through sequence complementarily. We further hypothesize that cellular defense mechanisms invoked by invading genetic elements could be rooted in this fundamental regulatory pathway that we call "GENE impedance", or simply, GENEi. Here, we present a working model that illustrates how transcription-termination and transcription-arrest can contribute to the regulation of gene expression via GENEi. In our model RNAi is only one component of GENEi, which is a more generalized mechanism of gene regulation. PMID- 15652141 TI - Modification patterns in germinating barley--malting II. AB - Modification refers to breaking down of cell walls and the conversion of starch to-sugars in the endosperm of grains by the action of enzymes released from the aleurone layer and possibly the scutellum during germination. Experimentalists have observed two completely different modification patterns in germinating barley. Based on an enzyme reaction, strongly nonlinear diffusion model developed in Part I of this series of papers on malting we examine simple models which predict modifications patterns similar to both observed patterns. We show that one of the observed patterns represents a natural propagation mode that acts as an attractor for the system. The speed of approach to this mode is strongly effected by initial conditions, a consequence of the enzymic nature of the reaction and the dramatic change of diffusivity brought about by the reaction. PMID- 15652142 TI - Random biochemical networks: the probability of self-sustaining autocatalysis. AB - We determine conditions under which a random biochemical system is likely to contain a subsystem that is both autocatalytic and able to survive on some ambient 'food' source. Such systems have previously been investigated for their relevance to origin-of-life models. In this paper we extend earlier work, by finding precisely the order of catalysation required for the emergence of such self-sustaining autocatalytic networks. This answers questions raised in earlier papers, yet also allows for a more general class of models. We also show that a recently described polynomial-time algorithm for determining whether a catalytic reaction system contains an autocatalytic, self-sustaining subsystem is unlikely to adapt to allow inhibitory catalysation--in this case we show that the associated decision problem is NP-complete. PMID- 15652143 TI - A possible role for phosphate in complexing the arginines of S4 in voltage gated channels. AB - Phosphate ions are known to complex guanidinium groups, which are the side chains of arginine. Voltage gated channels that allow passage of ions through cell membranes, producing, for example the nerve impulse, are in many cases composed of four domains, each with six transmembrane segments. The S4 transmembrane segments of these channels have arginines placed in such a way that they would be expected to complex phosphate. Known phosphate-arginine complexes are reasonably strong. Here, an ab initio calculation reinforces the expectation that a strong complex could form. As a consequence, if the S4 moved, it would carry either no charge, or at most half of what is expected from fully charged arginines. This suggests that it may be necessary to rethink voltage gating models in which the gating current is produced by physical motion of the S4 transmembrane segments. PMID- 15652144 TI - On the validity of the quasi-steady state approximation of bimolecular reactions in solution. AB - Two-step binding kinetics are extensively used to study the relative importance of diffusion in biochemical reactions. Classical analysis of this problem assumes ad hoc that the encounter complex is at quasi-steady state (QSS). Using scaling arguments we derive a criterion for the validity of this assumption in the limit of irreversible product formation. We find that the QSS approximation (QSSA) of two-step binding is only valid if the total ligand and receptor concentrations are much smaller than (k2+k-1)/k1, where k1 and k-1 are, respectively, the forward and reverse diffusion encounter rate constants and k2 is the chemical association rate constant. This criterion can be shown to imply that the average time between encounters is much longer than the half-life of the encounter complex and also guarantees that the concentration of the encounter complex is negligible compared to the reactant and product concentrations. Numerical examples of irreversible and reversible cases corroborate our analysis and illustrate that the QSS may be invalid even if k-2<4)-O-alpha-D-glucopyranosyl (1-->2)-beta-D-fructofuranoside] as an acceptor product with maltose. The optimum temperature and pH of this enzyme for levan formation were 30 degrees C and pH 6.2, respectively. M1FT levansucrase activity was completely abolished by 1 mM Hg2+ or Ag2+. The Km and Vmax values for levansucrase were calculated to be 26.6 mM and 126.6 micromol min-1 mg-1. PMID- 15652154 TI - Identification and characterization of a family of Caenorhabditis elegans genes that is homologous to the cadmium-responsive gene cdr-1. AB - Six Caenorhabditis elegans genes that are homologous to the novel, cadmium responsive gene cdr-1 have been identified and characterized. Nucleotide and amino acid sequence comparisons among the CDR family, which includes cdr-1, cdr 2, cdr-3, cdr-4, cdr-5, cdr-6, and cdr-7, reveals a high degree of identity among the seven members in this family. There are high levels of amino acid and nucleotide sequence similarity in the lengths of the open reading frames, predicted sizes, and protein characteristics. The seven proteins are predicted to be extremely hydrophobic, and are classified as integral membrane proteins. Structural analysis of the predicted proteins suggests that they may have similar biological functions. In response to cadmium exposure, cdr-1, cdr-2, cdr-3, and cdr-4 transcription significantly increases. In contrast, the levels of cdr-5, cdr-6, and cdr-7 transcription are not significantly affected or inhibited by cadmium exposure. Further, in non-exposed C. elegans, cdr-2, cdr-4, cdr-6, and cdr-7 are constitutively expressed. When CDR-1 expression was inhibited using RNAi, numerous fluid droplets were observed throughout the nematode body cavity. This phenotype became more pronounced in the presence of hypotonic stress. This suggests that CDR-1 may function in osmoregulation to maintain salt balance in C. elegans. PMID- 15652155 TI - Organization and functional analysis of the Schistosoma mansoni cathepsin D-like aspartic protease gene promoter. AB - We have cloned a 969-bp fragment of genomic DNA that spans 821 bp of the 5' untranslated region, exon 1, a short intron, and part of exon 2 of the Schistosoma mansoni cathepsin D gene by inverse PCR. Inspection of this sequence revealed the presence of two TATA-box motifs, two inverted CCAAT-box (inverted NF Y) motifs and sequences with homology to binding sites for the transcription factors, AP-1 and NF-Y. This sequence and deletion variants were cloned into reporter gene constructs, in order to examine the ability of these putative regulatory sequences to drive heterologous reporter gene activity. PCR products were cloned into the luciferase reporter vector pXP2. These reporter gene constructs were used to transform HeLa cells which were cultured and examined for luciferase activity. Additionally, HeLa cells transiently transfected with an EGFP reporter plasmid driven by the putative promoter from the S. mansoni cathepsin D gene were examined for EGFP transcripts and fluorescence. The 5' untranslated region of the S. mansoni cathepsin D gene, from position -772 to +40 (translation start ATG), included functional regulatory sequences capable of driving luciferase and EGFP expression, whereas shorter fragments from position 264 or -185 to +40 were insufficient to drive reporter activities. PMID- 15652156 TI - Two tyrosine hydroxylase genes in teleosts. AB - We report the finding of two non-allelic genes encoding tyrosine hydroxylase (TH) from the diploid teleost barramundi Lates calcarifer. Barramundi TH1 is the homologue of the higher vertebrate TH genes and encodes a protein of 489 amino acids that shares 90% sequence identity to the THs of other teleost species. A second non-allelic tyrosine hydroxylase gene (TH2) encodes a protein of 472 amino acids and shares 62% identity with TH1 and the vertebrate THs. TH1 mRNA is found in the brain and kidney of barramundi while TH2 mRNA is found only in brain. The TH2 gene is also present in the genomes of the pufferfish Takifugu and zebrafish Danio. Estimates of the rates of nucleotide substitution suggest the teleost TH2 genes are selectively constrained although not to the degree seen in the TH1 genes. Differential regulation of the two TH genes is, however, indicated by differences in transcript distribution, the nature of the Ca2+-responsive elements found in the proximal promoter region and the lack of recognised phosphorylation sites in TH2. Preservation of two apparently functional TH genes in phylogenetically distant teleost species is consistent with the notion of partitioning of function between duplicate genes. PMID- 15652157 TI - Transcriptional regulation of the human reduced folate carrier promoter C: synergistic transactivation by Sp1 and C/EBP beta and identification of a downstream repressor. AB - The human reduced folate carrier (hRFC) is ubiquitously but differentially expressed in human tissues and its levels are regulated by up to six alternatively spliced non-coding regions (designated A1/A2, A, B, C, D, and E) and by at least four promoters. By transient transfections of HepG2 human hepatoma cells with 5' and 3' deletion constructs spanning 2883 bp of upstream sequence, a transcriptionally important region was localized to within 177 bp flanking the transcriptional start sites for exon C. By gel shift and chromatin immunoprecipitation assays, Sp1 and C/EBP beta transcription factors were found to bind consensus elements (GC-box, CCAAT-box) within this region. The functional importance of these elements was confirmed by transient tranfections of HepG2 cells with hRFC-C reporter constructs in which these elements were mutated, and by co-transfections of Drosophila SL-2 cells with wild-type hRFC-C promoter and expression constructs for Sp1 and C/EBP beta. Whereas both Sp1 and C/EBP beta transactivated hRFC-C promoter activity, C/EBP alpha and gamma were transcriptionally inert. Sp1 combined with C/EBP beta resulted in a synergistic transactivation. In HepG2 cells, transfections with Sp1 and C/EBP beta both increased endogenous levels of hRFC-C transcripts. By 3' deletion analysis, a repressor sequence was localized to within 71 bp flanking the minimal promoter. On gel shifts, a novel transcriptional repressor was localized to within 30 bp. Collectively, these results identify transcriptionally important regions in the hRFC-C minimal promoter that include a GC-box and CCAAT-box, and suggest that cooperative interactions between Sp1 and C/EBP beta are essential for hRFC-C transactivation. Another possible factor in the tissue-specific regulation of the hRFC-C region involves the downstream repressor flanking the minimal promoter. PMID- 15652158 TI - Cloning and characterization of 5' upstream promoter region of rat WAP gene. AB - Regulatory regions of genes encoding milk proteins are frequently used to produce in the mammary gland of transgenic animals a variety of pharmaceutically and medically important human proteins. One such example is the whey acidic protein (WAP) promoter region, identified so far in the genome of mouse, rat, rabbit, camel, pig, brushtail possum and Tammar wallaby. The aim of the present study was cloning and characterization of the 5' upstream promoter region of rat WAP gene. Using Genome Walking procedure, we cloned the region extending from -849 to -3671 bp. We have shown that there are two conserved regions highly similar to hypersensitive sites present in mouse and rabbit upstream region of WAP gene with binding sites for STAT5 transcription factor, essential for expression of WAP gene in mammary glands during lactation. We characterized dispersed and tandem repeats in the upstream region of rat WAP gen localized not far away from the translation initiation site. PMID- 15652159 TI - Fish possess multiple copies of fgfrl1, the gene for a novel FGF receptor. AB - FGFRL1 is a novel FGF receptor that lacks the intracellular tyrosine kinase domain. While mammals, including man and mouse, possess a single copy of the FGFRL1 gene, fish have at least two copies, fgfrl1a and fgfrl1b. In zebrafish, both genes are located on chromosome 14, separated by about 10 cM. The two genes show a similar expression pattern in several zebrafish tissues, although the expression of fgfrl1b appears to be weaker than that of fgfrl1a. A clear difference is observed in the ovary of Fugu rubripes, which expresses fgfrl1a but not fgfrl1b. It is therefore possible that subfunctionalization has played a role in maintaining the two fgfrl1 genes during the evolution of fish. In human beings, the FGFRL1 gene is located on chromosome 4, adjacent to the SPON2, CTBP1 and MEAEA genes. These genes are also found adjacent to the fgfrl1a gene of Fugu, suggesting that FGFRL1, SPON2, CTBP1 and MEAEA were preserved as a coherent block during the evolution of Fugu and man. PMID- 15652160 TI - Molecular cloning and expression of cDNA coding for four spliced isoforms of casein kinase Ialpha in goldfish oocytes. AB - Casein kinase I (CKI) is a member of the serine/threonine protein kinases and located in a separate group within the superfamily of eukaryotic protein kinases. CKI isoforms regulate several checkpoints of the cell cycle and meiosis. In higher eukaryotes, CKIalpha has four isoforms produced through the alternative splicing of two short inserts. Here, we report the cloning, sequencing and expression of four alternatively spliced isoforms of CKIalpha from goldfish ovary. The cloned cDNAs were 2099-3002-bp long and classified as CKIalpha, CKIalphaS, CKIalphaL and CKIalphaLS. It was revealed that two major (3.0 and 2.0 kb) messages were strongly expressed in the ovary. Four isoforms are expressed in previtellogenic to vitellogenic oocytes. In the huge nucleus of the oocyte, referred to as the germinal vesicle, CKIalphaS is dominant and CKIalphaL is expressed at a detectable level. Immunoblot analysis revealed that CKIalpha and CKIalphaS are major products in both immature and mature oocytes. These two isoforms were expressed in a tissue-dependent manner. PMID- 15652162 TI - Clinical evaluation of 546 tetracycline-stained teeth treated with porcelain laminate veneers. AB - OBJECTIVE: The purpose of this study was to evaluate the clinical result of 546 tetracycline-stained teeth restored with a porcelain laminate veneer system (Cerinate, Den-Mat, USA) for aesthetic reasons. METHODS: Tetracycline-stained teeth (546) were restored with a porcelain veneer system, and bonded with Ultra Bond resin cement. The restorations were recalled after 0.5, 1.5 and 2.5 years, respectively. Modified Ryge criteria were used to evaluate the veneers marginal adaptation, interfacial staining, secondary caries, postoperative sensitivity and the patients' satisfaction of the colour of the restorations. RESULTS: This study found that 99% veneers had excellent marginal adaptations; and less than 1% veneers required rebonding in the first 6 months; the colour of the veneers was stable and no evident staining was found. Almost all patients were satisfied with the colour match of their restorations 1 year after placement. CONCLUSIONS: The research indicated that the porcelain veneer restoration system under investigation provided a reliable and highly satisfactory choice for the aesthetic restoration of tetracycline-stained teeth. PMID- 15652163 TI - Influence of surface and heat treatments on the flexural strength of Y-TZP dental ceramic. AB - OBJECTIVE: The aim of the investigation was to assess the influence of sandblasting, grinding, grinding orientation, polishing and heat treatment on the flexural strength of a yittria stabilized tetragonal zirconia polycrystals ceramic (Y-TPZ). METHODS: The specimens (160 beams) were equally divided into four groups according to the surface treatment (sandblasted, polished, ground parallel to the tensile axis, ground perpendicular). Twenty specimens from each group underwent heat treatment under the firing conditions used to fire a layer of porcelain and glaze. After treatment, the three-point flexure test was used to calculate the flexural strength and X-ray diffraction analysis was used to estimate the relative amount of monoclinic phase. The reliability of strength was assessed through the Weibull distribution. Statistical analysis was conducted with multiple regression analysis, one-way ANOVA and Tukey's pairwise multiple comparisons. Treated and fractured surfaces were observed with SEM. RESULTS: The following values of strength and relative content of monoclinic phase of zirconia were measured for each group: sandblasted (1540MPa; 9.5%); ground parallel (1330MPa; 8.3%); ground perpendicular (1525MPa; 8.3%); ground parallel and heated (1225MPa; monoclinic content not detectable); ground perpendicular and heated (1185MPa; monoclinic content not detectable); polished and heated (1165MPa; monoclinic content not detectable); polished (1095MPa; 0.8%); sandblasted and heated (955MPa; 0.3%). CONCLUSIONS: The present study suggests that sandblasting and grinding may be recommended to increase the strength of dental Y-TZP, provided they are not followed by heat treatment. Fine polishing may remove the layer of compressive stresses and therefore, lower the mean flexural strength. PMID- 15652164 TI - A comparison of a reformulated potassium citrate desensitising toothpaste with the original proprietary product. AB - OBJECTIVES: The recent tendency for toothpaste manufacturers to combine active ingredients for benefits to a number of oral problems requires that such combinations do not vitiate the action of any one agent. The aim of this study was to compare the desensitising effects of a modified formulation with the established product. MATERIALS AND METHODS: The study was a randomised, double blind, two treatment parallel design comparing an existing potassium citrate toothpaste product (Positive Control) with the same formulation to which triclosan and zinc citrate had been added (Test). A total of 56 subjects with dentine hypersensitivity (DH) completed the 8 week study period. At baseline, days 14 and 56 visual analogue scale scores were recorded by the subjects in response to air evaporative and cold water stimuli. RESULTS: Both groups showed improvements, mostly statistically significant, from baseline to day 14 and through to day 56. Most data favoured the Test paste but differences were, for the most part, not significant. CONCLUSIONS: It is concluded that the addition of known anti-plaque agents to an established potassium salt based desensitising toothpaste did not adversely affect the activity of the product in the treatment of DH. PMID- 15652165 TI - The failure rate of adhesively retained composite core build-ups in comparison with metal-added glass ionomer core build-ups. AB - OBJECTIVES: The purpose of this study was to evaluate the clinical performance of two adhesively retained composite core materials and compare them with a metal added glass ionomer. The main objective evaluated was total or partial loss of build-ups during the treatment prior to crown cementation. METHODS: In 187 patients, 315 vital and non-vital teeth were built up after randomisation with either Rebilda D (RD), Rebilda SC (RSC) or Ketac Silver Aplicap (KSA). The composites were applied in the total-etch-technique with the corresponding dentin bonding agent. The metal-added glass ionomer was used with a conditioner. One group of patients was treated by experienced dentists, the other by dental students, in order to evaluate the effects of different levels of experience. Data were analysed using Mann-Whitney-U-Test and binomial logistic regression. RESULTS: The early failure rate (partial or total loss) of core build-ups before crown cementation was significantly higher for KSA (28.8%), as compared to RSC (15.3%, p=0.037) and RD (15%, p=0.025). Most failures were observed during the removal of the temporary crowns. The rate of replacements was between 3.0 (RD/dentists) and 20.4% (KSA/students). Furthermore, we found that build-ups made by students had a significantly higher risk of loss than those made by dentists (p=0.028). CONCLUSIONS: Adhesively retained self-curing composites show a better clinical short-term performance and can be recommended as core build-up materials. PMID- 15652166 TI - The bleaching depth of a 35% hydrogen peroxide based in-office product: a study in vitro. AB - OBJECTIVES: The aim of the present study was to quantify the penetration of 35% hydrogen peroxide into enamel and dentine and to relate this to the resultant shade change of the tooth. METHOD: The crowns of 24 caries and developmental defect free human maxillary incisors were stained internally with a standardised tea solution. Twelve specimens were power bleached with light activated 35% hydrogen peroxide and 12 placed in water; both exposure times were 30min. Three different shade assessment methods (Vita shade guide [SG], shade vision system [SVS] and a chromometer) were employed prior to, after tea staining and after power bleaching/water treatments. Twelve specimens each from the bleach group and the water control water group were sectioned mesio-distally. An additional 12 specimens from the bleach and the control group were sectioned labio-palatally. The stain area for each specimen was measured using image analysis software. RESULTS: With tea staining, the mean changes in Vita shade guide units (SGU) ranged from 3.66 to 8.33. With the SVS system changes of 3.66-9 units were seen. Chromometer readings showed that following bleaching the L* values moved in the direction of black (3.8-6.7) and a* and b* values were in the red (0.3) and yellow (1.5) direction, respectively. Samples bleached and sectioned mesio distally showed stain coverage of 28.6-39.4%, while palatal sections showed stain coverage of 58-72%. Control samples, whether sectioned mesio-distally or labio palatally, showed staining throughout the dentine (97-100% coverage). CONCLUSION: A 35% hydrogen peroxide in-office bleaching gel demonstrated bleaching into dentine of uniform depth. PMID- 15652167 TI - Success rate of root canal treatment in primary molars. AB - OBJECTIVES: The purpose of the present retrospective large-scale study is to evaluate rate of success of root canal treatments in primary infected molars using Endoflas F.S as a filling material. METHODS: The study material consisted of 382 records of root canal treated primary molars of which 174 teeth with high quality radiographs and sufficient data comprised the 'study group'. The technique for root canal treatment in one visit is described. Treatment was considered success if clinically the tooth was asymptomatic and radiographs showed decrease or no change in pre-existing pathologic radiolucent defects. RESULTS: In the total pulpectomy group there was a statistically significant difference between mandibular teeth presented with a periapical lesion prior to the treatment compared to maxillary teeth. This difference was not significant in the study group. Decrease in post treatment radicular radiographic defects was statistically significant more in the maxilla compared to the mandible. The success rate of teeth restored with a permanent restoration (stainless steel crown or amalgam filling) was statistically significant higher than those teeth left with a temporary filling. There was no statistically significant difference between success rate and the extent of root canal filling material. Rate of success was not statistically different depending on pre-existing radiolucent area prior to RCT. CONCLUSIONS: Endoflas F.S. can be used as an alternative root canal filling material for primary teeth. One condition for success is the prevention of microleakage. A permanent restoration should be placed as soon as possible after clinical signs and symptoms of inflammation are eliminated. PMID- 15652168 TI - In vitro embryotoxicity assessment with dental restorative materials. AB - OBJECTIVES: Resin (co)monomers may be released from restorative dental materials and can diffuse into the tooth pulp or the gingiva, and can reach the saliva and the circulating blood. Genotoxic potential of some dental composite components has been clearly documented. The genotoxic effects of xenobiotics can represent a possible step in tumor initiation and/or embryotoxicity/teratogenesis. A modified fluorescent mouse embryonic stem cell test (R.E.Tox) was used to test the embryotoxic potential of following dental restorative materials: Bisphenol A glycidylmethacrylate (BisGMA), urethanedimethacrylate (UDMA), hydroxyethylmethacrylate (HEMA), and triethyleneglycoldimethacrylate (TEGDMA), as well as some of their metabolic intermediates 2,3-epoxy-2-methyl-propionicacid methylester (EMPME), methacrylic acid (MA), and 2,3-epoxy-2-methylpropionic acid (EMPA). METHODS: Mouse embryonic stem (ES) cells stably transfected with a vector containing the gene for the green fluorescent protein under control of the cardiac alpha-myosin heavy chain promoter were differentiated in the presence of various concentrations of the test compounds for 12 days. Fluorescence was measured using the TECAN Safire and values were expressed as percent of control values. To distinguish between cytotoxic and embryotoxic effects, all compounds were tested in a standard MTT assay. RESULTS: HEMA, TEGDMA and EMPME did not influence the differentiation process of ES cells towards cardiac myocytes. No cytotoxic effects were observed at any of the concentration levels tested. Exposure to BisGMA resulted in a 50% decrease in cell survival and a very strong inhibition of cell differentiation at 10(-5)M (p<0.01). Embryotoxic effects were also present at 10(-6) and 10(-7)M (p<0.05). EMPA induced a decrease in ES cell differentiation at 10(-5)M (p<0.01) without cytotoxic effects. No embryotoxic effects were induced at lower concentrations. Exposure to UDMA resulted in a slight decrease of cell differentiation at 10(-5)M (p<0.05). Exposure of cells to MA resulted in an increase of cardiac differentiation up to 150% (p<0.05) at 10( 5)M without cytotoxic effects. CONCLUSIONS: BisGMA induced a significant high embryotoxic/teratogenic effect over a large range of concentration. Therefore attention should be focused on this dental monomer, which should be investigated further by in vivo experiments. PMID- 15652170 TI - Grey-scale reversed radiographic display in the detection of approximal caries. AB - OBJECTIVES: Purpose of this study was to determine the influence of grey-scale reversal of digital radiographic images on the detection of proximal caries. METHODS: Five observers assessed digital bitewing radiographs (Sidexis((R)), Sirona) of 320 unrestored surfaces of extracted posterior teeth embedded in 20 models, simulating pairing of maxillary and mandibular arches, using a six category caries rating scale. Images were displayed in normal [N] and inverse [IN] mode at different image sizes (display ratio: 1:1, 1:2, 1:7) on a cathode ray tube monitor (Nokia 446 XS) and a TFT display (Panasonic LC 50 S). Validation standard was defined as histological lesion depth. Validity was expressed as areas under receiver operating characteristic curves (AUC) calculated for two levels of histological caries penetration: presence of caries and presence of a dentine lesion. The factors 'inverse display', 'image size on-screen' 'display type' and 'disease cut-off' were analysed by repeated measures ANOVA. RESULTS: Inverse image display significantly influenced the diagnostic validity (P=0.014), but a reduced accuracy was only seen at the lowest image size (AUC (SE): [N] 0.64 (0.02); [IN] 0.62 (0.02)). At the validation threshold 'dentine caries' approximal caries detection deteriorated when using grey-scale reversal (AUC (SE): [N] 0.71 (0.02); [IN] 0.69 (0.02)). CONCLUSIONS: In conclusion, grey-scale reversal of digital radiographs did not optimise approximal caries detection and aggravated the detectability of dentinal lesions. PMID- 15652169 TI - Effect of bench cooling on the dimensional accuracy of heat-cured acrylic denture base material. AB - OBJECTIVES: In order to obtain an insight on the internal stress caused by both polymerization and thermal shrinkage during the fabrication process of heat-cured denture resin, the effect of bench cooling on the dimensional accuracy of a heat cured denture base resin was examined. METHODS: A dimensional change of a dumbbell-shaped specimen during the fabrication process was measured directly by using the strain gauge method. After polymerization, the specimens were treated in one of the following two processing methods: (1) rapid cooling: the specimen was removed from a stone mold within a container of boiling water at 100 degrees C and then left to cool in a thermo-stabilized room of 20+/-1 degrees C; (2) bench cooling: the flask was left to cool in a thermo-stabilized room of 20+/-1 degrees C for 140min, after which, the specimen was removed from the stone mold. The strain from deflasking was derived from the difference in the strain, before and after the removal of the specimen from the stone mold. The strain differential, before and after cooling, was determined as the total strain. RESULTS: The bench cooling for the heat-cured denture base resin reduced the strain caused by thermal shrinkage during the fabrication process. The observed reduction in the strain was 26% for the C(L) (direction of center's length), 11% for the E(L) (direction of left-edge's length), and 12% for the E(W) (direction of left-edge's width), when compared with the results obtained from the rapid cooling method. CONCLUSIONS: The flask should be slowly cooled to room temperature, since the internal stress developed by thermal shrinkage will be relaxed during the cooling process. PMID- 15652171 TI - A preliminary investigation of a spectroscopic technique for the diagnosis of natural caries lesions. AB - OBJECTIVE: To report the use of spectroscopic analysis of dental fluorescence excited with a blue InGaN laser diode operating at 405 nm. METHOD: The spectra resulting from three classifications of smooth surface non-cavitated caries lesions (dull, shiny, brown) with 20 samples in each group were examined using the ratio of integrated fluorescence intensity in two spectral bands. RESULTS: All lesions demonstrated spectra which were significantly different from sound tooth structure. As expected, the 'brown' lesions demonstrated a significantly different spectral profile from the two white spot lesion classifications. Dull and shiny lesions had significantly different spectral measurements when examining the ratio of the integrated fluorescence in spectral bands between 480 500 and 620-640 nm. CONCLUSION: This method has application for detection of dental caries as well as demonstrating potential application to evaluate lesions which may represent different degrees of caries activity. PMID- 15652172 TI - Performance of DIAGNOdent for detection and quantification of smooth-surface caries in primary teeth. AB - OBJECTIVES: To evaluate the performance of the laser fluorescence device (DIAGNOdent) for detection and quantification of smooth-surface caries lesions in primary teeth. METHODS: Seventy-seven approximal surfaces with no caries or with white spot caries lesions without cavitation was selected from 51 primary molars. Laser fluorescence readings were performed and the site of higher value in each surface was indicated. After the readings, the surfaces were cut and the samples were destined for histopathological and cross-sectional microhardness evaluation. ROC analysis was conducted and the area under the ROC curve and the best cut-off points were obtained at D1, D2 and D3 thresholds. With these cut-off values, the sensitivity, specificity and accuracy were calculated at these thresholds. The regression lines of laser fluorescence readings as function of lesion extension, lesion depth and cross-sectional microhardness values were plotted and the Pearson correlation coefficients were obtained. RESULTS: The best DIAGNOdent performance was obtained at D3 threshold (A(z)=0.90) and the worst performance was achieved at D1 threshold (A(z)=0.82). The correlation was better between the laser fluorescence readings and lesion depth obtained in polarized light microscope examination (r=0.78), than that obtained with the microhardness values (r=-0.57). CONCLUSION: The laser fluorescence shows a good performance to predict caries lesions extension. Device presents a better correlation with the lesion depth than with mineral loss in smooth-surface caries lesions. PMID- 15652173 TI - Structural characterization of human liver heparan sulfate. AB - The isolation, purification and structural characterization of human liver heparan sulfate are described. 1H-NMR spectroscopy demonstrates the purity of this glycosaminoglycan (GAG) and two-dimensional 1H-NMR confirmed that it was heparan sulfate. Enzymatic depolymerization of the isolated heparan sulfate, followed by gradient polyacrylamide gel, confirmed its heparin lyase sensitivity. The concentration of resulting unsaturated disaccharides was determined using reverse phase ion-pairing (RPIP) HPLC with post column derivatization and fluorescence detection. The results of this analysis clearly demonstrate that the isolated GAG was heparan sulfate, not heparin. Human liver heparan sulfate was similar to heparin in that it has a reduced content of unsulfated disaccharide and an elevated average sulfation level. The antithrombin-mediated anti-factor Xa activity of human liver heparan sulfate, however, was much lower than porcine intestinal (pharmaceutical) heparin but was comparable to standard porcine intestinal heparan sulfate. Moreover, human liver heparan sulfate shows higher degree of sulfation than heparan sulfate isolated from porcine liver or from the human hepatoma Hep 2G cell line. PMID- 15652174 TI - Characterization of NTPDase (NTPDase1; ecto-apyrase; ecto-diphosphohydrolase; CD39; EC 3.6.1.5) activity in human lymphocytes. AB - Human lymphocytes contain NTPDase (NTPDase-1; ecto-apyrase; ecto diphosphohydrolase; CD39; EC 3.6.1.5), a cation-dependent enzyme that hydrolyzes ATP and ADP and also other di- and triphosphate nucleosides, acting at an optimum pH of 8.0. A significant inhibition of ATP and ADP hydrolysis (P<0.05) was observed in the presence of 20 mM sodium azide. NTPDase inhibitors, 20 mM sodium fluoride, 0.2 mM trifluoperazine and 0.3 mM suramin, significantly decreased ATP and ADP hydrolysis (P<0.05) and ADP hydrolysis was only inhibited by 0.5 mM orthovanadate (P<0.05). ATP and ADP hydrolysis was not inhibited in the presence of 0.01 mM Ap5A (P1,P5-di(adenosine-5')pentaphosphate), 0.1 mM ouabain, 1 mM levamisole, 2 microg/mL oligomycin, 0.1 mM N-ethylmaleimide (NEM), or 5 mM sodium azide. With respect to kinetic behavior, apparent K(m) values of 77.6+/-10.2 and 106.8+/-21.0 microM, and V(max) values of 68.9+/-8.1 and 99.4+/-8.5 (mean+/-S.E., n=3) nmol Pi/min/mg protein were obtained for ATP and ADP, respectively. A Chevilard plot demonstrated that only one enzymatic site is responsible for the hydrolysis of ATP and ADP. The presence of CD39 was determined by flow cytometry, showing a low density of 2.72+/-0.24% (mean+/-S.E.; n=30) in human peripheral lymphocytes. The study of NTPDase activity in human lymphocytes may be important to determine the immune response status against infectious agents related to ATP and ADP hydrolysis. PMID- 15652175 TI - Phosphorylation modulates the alpha-helical structure and polymerization of a peptide from the third tau microtubule-binding repeat. AB - Paired helical filaments (PHFs) isolated from patients with Alzheimer's disease (AD) mainly consist of the microtubule-associated protein tau in a hyperphosphorylated form. It has been found that PHFs are the first example of pathological protein aggregation associated with formation of alpha-helices [Biochemistry (2002) 41, 7150-5]. In an effort to investigate the interplay between phosphorylation and the putative role of short regions of alpha-helix in the polymerization of tau, we have focused on the region of tau encompassing residues 317 to 335. This region is able to form protein fibrils in vitro and has two serines that are often found phosphorylated in PHFs. Using trifluoroethanol as an indicator of the alpha-helix, we find that the stability of the alpha-helix conformation is enhanced by phosphorylation. Circular dichroism data show that the phosphorylated peptide in water presents a content in alpha-helix similar to the unphosphorylated peptide at 40% of trifluoroethanol. Phosphorylation also stimulates the effect of juglone in promoting the in vitro polymerization. Furthermore, Fourier transformed infrared spectroscopy of samples of phosphorylated peptide polymerized with juglone renders a spectrum with maxima at approximately 1665 and approximately 1675 cm(-1), which are suggestive of a mixture of turns and alpha-helix conformations. Our results provide a direct mechanistic connection between phosphorylation and polymerization in tau. The connection between phosphorylation and polymerization appears to involve formation of alpha-helix structure. PMID- 15652176 TI - Biosynthesis of lysine in plants: evidence for a variant of the known bacterial pathways. AB - With the aim of elucidating how plants synthesize lysine, extracts prepared from corn, tobacco, Chlamydomonas and soybean were tested and found to lack detectable amounts of N-alpha-acyl-L,L-diaminopimelate deacylase or N-succinyl-alpha-amino epsilon-ketopimelate-glutamate aminotransaminase, two key enzymes in the central part of the bacterial pathway for lysine biosynthesis. Corn extracts missing two key enzymes still carried out the overall synthesis of lysine when provided with dihydrodipicolinate. An analysis of available plant DNA sequences was performed to test the veracity of the negative biochemical findings. Orthologs of dihydrodipicolinate reductase and diaminopimelate epimerase (enzymes on each side of the central pathway) were readily found in the Arabidopsis thaliana genome. Orthologs of the known enzymes needed to convert tetrahydrodipicolinate to diaminopimelic acid (DAP) were not detected in Arabidopsis or in the plant DNA sequence databases. The biochemical and reinforcing bioinformatics results provide evidence that plants may use a novel variant of the bacterial pathways for lysine biosynthesis. PMID- 15652177 TI - Fructose metabolizing enzymes from mouse liver: influence of age and caloric restriction. AB - The influence of caloric restriction (CR) on the activities of liver fructose metabolizing enzymes and metabolite levels were studied in young (3 months) and old (30 months) mice. Fructokinase activity was increased (P<0.05) in both young and old CR mice when compared to controls while triokinase activity was increased (P<0.05) only in old CR versus control mice. Aldolase was not altered by CR in either old or young mice. No age-related differences in activities were observed in controls although a trend towards an increase was observed for triokinase, while significant age-related increases were observed for fructokinase and triokinase, but not aldolase, in CR mice. Both young and old mice on CR showed significant decreases in fructose and fructose-1-phosphate, however, no age related changes in metabolite levels were observed for either control or CR mice. A fructose-1-phosphate kinase activity was also measured and found to be unchanged in both young and old mice on CR, but the activity was significantly lower in the old mice compared with young. We show here that the enzymes involved in fructose metabolism are influenced by CR and that this could contribute to alterations in gluconeogenesis and glycolysis observed with CR. PMID- 15652178 TI - Glycosaminoglycan destabilization of DNA-chitosan polyplexes for gene delivery depends on chitosan chain length and GAG properties. AB - Chitosan-based gene delivery systems are promising candidates for non-viral gene therapy. A wide range of chitosans has been studied to optimize the properties of the DNA-chitosan complexes to yield high transfection efficiencies. An important parameter to control is the polyplex stability to allow transport towards the cells, subsequent internalization and release of DNA intracellularly. The stability of the DNA-chitosan complexes was here studied after exposure to heparin and hyaluronic acid (HA) using atomic force microscopy (AFM) and ethidium bromide (EtBr) fluorescence assay. To study the effect of polycation chain length on the polyplex stability, chitosans with a degree of polymerization (DP) varying from approximately 10 to approximately 1000 were employed for DNA compaction. Whereas HA was unable to dissociate the complexes, the degree of dissociation caused by heparin depended on both the chitosan chain length and the amount of chitosan used for complexation. When increasing the chitosan concentration, larger heparin concentrations were required for polyplex dissociation. Furthermore, increasing the chitosan chain length yielded more stable complexes. Varying the chitosan chain length thus provides a tool for controlling the ability of the polyplex to deliver therapeutic gene vectors to cells. PMID- 15652179 TI - Induction of apoptosis by electrotransfer of positively charged proteins as Cytochrome C and Histone H1 into cells. AB - Cytochrome C (Cyt. C) is a mitochondrial protein inducing apoptosis when it is accumulated in the cytosol by a currently unknown mechanism, but regulated by the bcl-2 family of proteins. The linker Histone H1 is another basic protein with highly conservative structure, composition, and equal molecular weight, not changed during the evolution. An attempt was made to understand better the apoptotic processes by electroloading of leukemic cells, such as K562, HL-60, and SKW3, and human lymphocytes with positively charged proteins, such as Cyt. C, Histone H1, and methylated BSA albumin (mBSA). The triggering apoptotic processes followed by MTT test, FACS analysis, and DNA fragmentation after the electrotransfer of these proteins into the cells were observed. Histone H1 and mBSA induce the release of Cyt. C from rat liver mitochondria. Cytochrome C release was higher when mitochondria were in "high-energy" state. It is supposed that release of Cyt. C from mitochondria is due to the mechanical rupture of the outer mitochondrial membrane, rich in negatively charged groups, predominately due to cardiolipin. The reason for the morphological rupture of the outer mitochondial membrane could be the rigidification and segregation of the membrane and the destroyed membrane asymmetries of both monolayers in the presence of positively charged proteins at higher linear charges such as Histone H1. We suggested that Histone H1, at a given moment of activated signaling for apoptosis, could be not transported to the nucleus and could lead to the release of Cyt. C from the mitochondria in the cytoplasm. It is temping to speculate that Histone H1 has other physiological extranuclear functions involved in apoptosis. PMID- 15652180 TI - Novel nonmatrix-metalloproteinase-mediated collagen degradation. AB - Matrix metalloproteinases (MMPs) and their inhibitors have long been believed to play a major role in the collagen loss seen in destructive temporomandibular joint (TMJ) disorders. This project was originally designed to examine the expression of MMPs and the tissue inhibitors of MMPs (TIMPs) by diseased human TMJ synovial fibroblasts and to determine their ability to degrade Type I collagen. Reverse transcriptase-PCR indicated that these TMJ synovial fibroblasts expressed mRNA for multiple MMPs and TIMPs. The collagen degradation assay showed that these TMJ synovial fibroblasts at passage 3 to 8 were capable of digesting the collagen underneath them on collagen-coated plates. This degradation was inhibited by GM6001, a synthetic MMP inhibitor. During passage 8 to 13, these TMJ fibroblasts were able to digest all the collagen in the wells. This degradation was completely inhibited by combining GM6001 and soybean trypsin inhibitor (STI), a serine proteinase inhibitor. The collagen cleavage activity of collected conditioned media was dramatically inhibited by STI but not by 1,10 phenanthroline, an MMP inhibitor. The data suggest that these TMJ cells utilize a MMP-dependent pathway and a novel MMP-independent pathway to digest Type I collagen. PMID- 15652181 TI - cDNA microarray analysis of lactoferrin expression in non-neoplastic human hepatocyte PH5CH8 cells. AB - Lactoferrin (LF), a milk protein belonging to the iron transporter transferrin family, is known as a primary defense protein against pathogenic microorganisms. Previously, we found that bovine and human LFs prevented hepatitis C virus infection in cultured human hepatocytes by a direct interaction with the virus. Since LF is proposed to have transcriptional regulatory activity in addition to its antimicrobial function, we sought to identify the target genes that these two types of LF have in common. To this end, we were the first to perform microarray analysis (9970 genes) using human hepatocytes that expressed bovine or human LF by retrovirus-mediated gene transfer. In the results, LF could give a variety of expression profiles in the human hepatocytes, and showed that 9 and 19 genes were commonly up-regulated (more than 2.0-fold) and down-regulated (less than 0.50 fold), respectively, in both bovine and human LF-expressing cells compared with control cells. Among these genes, we found that gamma-aminobutyric acid (GABA)-B receptor 2 was transcriptionally down-regulated by bovine and human LFs, but not by human transferrin. Furthermore, we obtained the suggestive result that LF may modulate the level of intracellular cAMP. This modulation is one of the cellular responses that the GABA-B receptor modifies. This is the first report of microarray analysis applied to search inclusively for the target genes of LF. PMID- 15652182 TI - The extractability of extracellular matrix components as a marker of cartilage remodeling in laryngeal squamous cell carcinoma. AB - Sequential extraction was applied to investigate the proteoglycan (PG) organization in healthy laryngeal cartilage (HLC) and laryngeal cartilage squamous cell carcinoma (LCSCC). Highly stable aggrecan aggregates, extracted from both HLC and LCSCC with strong dissociative reagents, i.e., 4 M guanidine HCl (GdnHCl), represented 53% and 7%, respectively, of total extracted macromolecules. Less stable complexes/aggregates, extracted with mild dissociative reagents (1 and 2 M GdnHCl), represented 40% and 61% of total extracted PGs from healthy and cancerous cartilage, respectively. Interestingly, a relative high proportion (32%) of uronic acid (UA)-containing macromolecules were removed from the cancerous cartilage using associative extracting solutions (PBS and 0.5 M GdnHCl), which obviously represented molecules freely extractable from the tissue. In contrast, the corresponding proportion in HLC was impressively low (about 7%). The major proportion of these molecules was chondroitin sulfate-containing PGs (CSPGs), which identified mainly as aggrecan. Differential digestion of the sequential extracts with chondroitinase ABC and chondroitinase AC II demonstrated the presence of dermatan sulfate-containing PGs (DSPGs) in both HLC and LCSCC, being mainly present in the 1 M GdnHCl extract, and identified as decorin. All cancerous extracts were found to be rich in 4 sulfated disaccharides, mostly participating in DS structures. In conclusion, the applied procedure permitted the elucidation of the changes in the cartilage status, regarding the stability and identity of its proteoglycan aggregates/complexes, in both HLC and LCSCC. PMID- 15652183 TI - Peptides and hydrolysates from casein and soy protein modulate the release of vasoactive substances from human aortic endothelial cells. AB - Food proteins were shown to affect atherogenic risk factors, which is supposed to be related to specific peptide sequences encrypted within their primary sequence. The aim of this study was to evaluate the effects of peptides and hydrolysates from two food proteins, casein and soy protein, on endothelial cell functions (cell proliferation and release of vasoactive substances). Cell proliferation was not influenced by dipeptides and most of the tripeptides, whereas several total hydrolysates from casein and soy protein inhibited cell proliferation at higher concentrations (>0.25 mg/mL; P<0.05). The release of one or more of the vasoactive substances, thromboxan B2 (stable marker of thromboxan A2), 6-keto prostaglandin F1alpha (stable marker of prostaglandin I2), endothelin-1, and nitric oxide, was significantly influenced by the incubation with various peptides compared with control cells (P<0.05). Various hydrolysate fractions from casein and soy protein influenced the release of 6-keto-prostaglandin F1alpha and nitric oxide (P<0.05) but did not influence the release of thromboxan B2 and endothelin-1. In conclusion, the present study demonstrates that peptides and hydrolysate fractions from casein and soy protein influence endothelial cell function as evidenced by the modulation of endothelial cell proliferation and alterations in the release of vasoactive substances. PMID- 15652184 TI - Low tumor cell density environment yields survival advantage of tumor cells exposed to MTX in vitro. AB - Stable resistance to methotrexate has been well characterized after prolonged treatment of the HT-29 colon cancer cell line, but the mechanism of cell survival at the early stages of the drug resistance process still remains unclear. Here, we demonstrate that human cancer cells in vitro are sensitive to methotrexate only above a critical cell culture density, which specifically coincides with their ability to deplete the extracellular nucleosides from a fully supplemented culture medium. At lower cell densities, extracellular nucleosides remain intact and allow salvage nucleotide synthesis that renders cells insensitive to the drug. Consistently, medium conditioned by cells seeded at standard cell densities sensitizes low cell density cultures. Extracellular nucleosides are the determinants of sensitivity because the latter effect can be mimicked with the use of inhibitors of nucleoside cellular import and reversed by supplying exogenous thymidine and hypoxanthine. Interestingly, treatment at a sensitizing cell density does not preclude the survival of less than 1% of the cells--which have no intrinsic resistance--owing to the inability of the dying cell population to condition the culture medium; this population thus survives indefinitely to continuous treatment by keeping adapted to a low cell number. This cell density dependent adaptive process accounts for the initial steps of in vitro resistance to methotrexate (MTX) and provides a novel mechanistic insight into the cell population dynamics of cell survival and cell death during drug treatment. PMID- 15652185 TI - Detection of glycosyltransferases in the golden hamster (Mesocricetus auratus) oviduct and evidence for the regulation of O-glycan biosynthesis during the estrous cycle. AB - Recently, we provided evidence that the glycosylation of hamster oviductin, a member of the mucin family of glycoproteins, is regulated during the estrous cycle. In order to further elucidate the glycosylation process of oviductal glycoproteins, we identified biosynthetic pathways involved in the assembly of mucin-type O-linked oligosaccharide (O-glycan) chains in the hamster oviduct. Our results demonstrated that the hamster oviduct has high activities of glycosyltransferases that synthesize O-glycans with core 1, 2, 3 and 4 structures as well as elongated structures. Oviduct therefore represents a typical mucin secreting tissue. Our results also showed that specific glycosyltransferase activities are regulated during the estrous cycle. Mucin-type core 2 beta6-GlcNAc transferase (C2GnT2) is responsible for synthesizing core 2 and core 4 structures in the oviduct. Specific assays for C2GnT2 revealed a cyclical pattern throughout the estrous cycle with high activity at the stages of proestrus and estrus and low activity at diestrus 1. Using semiquantitative RT-PCR, the mRNA levels for C2GnT2 in the estrous cycle stages could be correlated with the enzyme activities. An increase in glycosyltransferase activity in the hamster oviduct at the time of ovulation suggests that glycosylation of oviductal glycoproteins may be necessary for these proteins to exert their functions during the process of fertilization. PMID- 15652186 TI - An extracellular endodeoxyribonuclease from Streptomyces aureofaciens. AB - Several extracellular DNases were detected after cultivation of Streptomyces aureofaciens B96 under submerged conditions. These DNases are nutritionally regulated and high content of amino acid nitrogen in cultivation medium repress their production. By varying cultivation conditions, there remained only two extracellular nuclease activities. The major one, extracellular endodeoxyribonuclease SaD I, was purified to homogeneity by ammonium sulfate precipitation, adsorption on Spheron, chromatography on Superose-12P followed by FPLC on MonoQ and final purification on HiTrapQ. The molecular weight of the purified SaD I determined by SDS-PAGE was 31 kDa. The DNase hydrolyses endonucleolytically both double-stranded and single-stranded circular and linear DNA. It does not cleave RNA and does not exhibit phosphodiesterase nor phosphomonoesterase activity. It requires a divalent cation (Zn2+, Co2+, Mn2+, Mg2+) and its activity optimum is at neutral pH (pH 7.2). The optimal temperature for DNA cleavage was 40 degrees C. Activity was strongly inhibited in the presence of phosphate, Hg2+, chelating agents or iodoacetate, but it was stimulated by addition of dimethyl sulphoxide. PMID- 15652188 TI - Boundary-element calculations for dielectric behavior of doublet-shaped cells. AB - In order to simulate dielectric relaxation spectra (DRS) of budding yeast cells (Saccharomyces cerevisiae) in suspension, the complex polarization factor (Clausius-Mossotti factor) beta for a single cell and the complex permittivity of a cell suspension epsilon(sus)* were calculated with a doublet-shaped model (model RD), in which two spheres were connected with a part of a ring torus, using the boundary element method. The beta values were represented by a diagonal tensor consisting of components beta(z) parallel to the rotation axis (z axis) and beta(h) in a plane (h plane) perpendicular to the axis. The epsilon(sus)* values were calculated from the complex permittivity of the suspending medium epsilon(a)* and the components of beta. The calculation was compared with that of a conventional prolate spheroid model (model CP). It was found that model CP could be used as a first approximation to model RD. However, differences existed in beta(z) between models RD and CP; beta(z) showed three relaxation terms in the case of model RD in contrast with two terms in model CP. Narrowing the junction between the two spheres in model RD markedly decreased the characteristic frequency of one of the relaxation terms in beta(z). This suggests that the structure of the junction can be estimated from DRS. Effects of the shape change from model RD to a two-sphere model (model RD without the junction) were also examined. The behavior of beta(z) in the two-sphere model, the relaxation intensity of which was much lower than model RD, was quite similar to that in a single-sphere model. These simulations were consistent with the experimental observations of the dielectric behavior of the yeast cells during cell cycle progression. PMID- 15652187 TI - Inhibition of firefly luciferase by alkane analogues. AB - We reported that anesthetics increased the partial molal volume of firefly luciferase (FFL), while long-chain fatty acids (LCFA) decreased it. The present study measured the actions of dodecanol (neutral), dodecanoic acid (negatively charged), and dodecylamine (positively charged) hydrophobic molecules on FFL. The interaction modes are measured by (1) ATP-induced bioluminescence of FFL and (2) fluorescence of 2-(p-toluidino)naphthalene-6-sulfonate (TNS). TNS fluoresces brightly in hydrophobic media. It competes with the substrate luciferin on the FFL binding. From the Scatchard plot of TNS titration, the maximum binding number of TNS was 0.83, and its binding constant was 8.27 x 10(5) M(-1). Job's plot also showed that the binding number is 0.89. From the TNS titration of FFL, the binding constant was estimated to be 8.8 x 10(5) M(-1). Dodecanoic acid quenched the TNS fluorescence entirely. Dodecanol quenched about 25% of the fluorescence, whereas dodecylamine increased it. By comparing the fluorescence of TNS and bioluminescence of FFL, the binding modes and the inhibition mechanisms of these dodecane analogues are classified in three different modes: competitive (dodecanoic acid), noncompetitive (dodecylamine), and mixed (dodecanol). PMID- 15652189 TI - Do clustered beta-propeller domains within the N-terminus of LRP1 play a functional role? AB - The six beta-propellers located within the N-terminus of low density lipoprotein receptor-related protein 1 (LRP1) are arranged in two clusters that contain two and four beta-propellers, respectively. Working with LRP1 deletion mutants, we found that randomly removing large segments of amino acid sequences did not affect the intracellular trafficking of LRP1 as long as the clustered beta propeller domains were retained. However, deletion mutants with crippled beta propeller clusters invariably exhibited retarded exit from the endoplasmic reticulum (ER). To determine potential functions of the clustered beta propellers, we generated a series of deletion mutants in which the beta propellers were systematically removed from the C-terminal end of the second cluster. The resulting minireceptors, designated LRPbeta1-6, beta1-5, beta1-4, beta1-3, and beta1-2 containing decreasing numbers of the beta-propellers, were stably expressed in LRP1-null CHO cells. Binding/degradation assays with receptor associated protein or alpha2-macroglobulin showed that removing one or more beta propellers had little effect on binding or degradation of these ligands. However, minireceptors containing odd number of beta-propellers (i.e., LRPbeta1-3 and beta1-5) showed prolonged retention within the ER and remained endoglycosidase H sensitive, whereas minireceptors containing even number of beta-propellers (i.e., LRPbeta1-2, beta1-4 and beta1-6) exited ER at variable rates. Cell surface biotinylation experiments showed that LRPbeta1-3 was absent from the cell surface. Prolonged retention of LRPbeta1-3 within the ER was accompanied by increased association with molecular chaperone Grp78/Bip. These results suggest that the clustered beta-propellers may play a role in folding and intracellular trafficking of LRP1. PMID- 15652190 TI - Lectin KM+-induced neutrophil haptotaxis involves binding to laminin. AB - The lectin KM+ from Artocarpus integrifolia, also known as artocarpin, induces neutrophil migration by haptotaxis. The interactions of KM+ with both the extracellular matrix (ECM) and neutrophils depend on the lectin ability to recognize mannose-containing glycans. Here, we report the binding of KM+ to laminin and demonstrate that this interaction potentiates the KM+-induced neutrophil migration. Labeling of lung tissue by KM+ located its ligands on the endothelial cells, in the basement membrane, in the alveolus, and in the interstitial connective tissue. Such labeling was inhibited by 400 mM D-mannose, 10 mM Manalpha1-3[Manalpha1-6]Man or 10 microM peroxidase (a glycoprotein containing mannosyl heptasaccharide). Laminin is a tissue ligand for KM+, since both KM+ and anti-laminin antibodies not only reacted with the same high molecular mass components of a lung extract, but also determined colocalized labeling in basement membranes of the lung tissue. The relevance of the KM+ laminin interaction to the KM+ property of inducing neutrophil migration was evaluated. The inability of low concentrations of soluble KM+ to induce human neutrophil migration was reversed by coating the microchamber filter with laminin. So, the interaction of KM+ with laminin promotes the formation of a substrate-bound KM+ gradient that is able to induce neutrophil haptotaxis. PMID- 15652191 TI - Flavonoid-serum albumin complexation: determination of binding constants and binding sites by fluorescence spectroscopy. AB - After a meal rich in plant products, dietary flavonols can be detected in plasma as serum albumin-bound conjugates. Flavonol-albumin binding is expected to modulate the bioavailability of flavonols. In this work, the binding of structurally different flavonoids to human and bovine serum albumins is investigated by fluorescence spectroscopy using three methods: the quenching of the albumin fluorescence, the enhancement of the flavonoid fluorescence, the quenching of the fluorescence of the quercetin-albumin complex by a second flavonoid. The latter method is extended to probes whose high-affinity binding sites are known to be located in one of the two major subdomains (warfarin and dansyl-L-asparagine for subdomain IIA, ibuprofen and diazepam for subdomain IIIA). Overall, flavonoids display moderate affinities for albumins (binding constants in the range 1-15 x 10(4) M(-1)), flavones and flavonols being most tightly bound. Glycosidation and sulfation could lower the affinity to albumin by one order of magnitude depending on the conjugation site. Despite multiple binding of both quercetin and site probes, it can be proposed that the binding of flavonols primarily takes place in subdomain IIA. Significant differences in affinity and binding location are observed for the highly homologous HSA and BSA. PMID- 15652192 TI - Evaluation of the antioxidant activity of flavonoids by "ferric reducing antioxidant power" assay and cyclic voltammetry. AB - Flavonoids, naturally occurring phenolic compounds, have recently been studied extensively for their antioxidant properties. The structure-antioxidant activity relationships (SAR) of flavonoids have been evaluated against different free radicals, but "ferric reducing antioxidant power" (FRAP) assay, which determines directly the reducing capacity of a compound, has not been used for this purpose. In this study, the antioxidant activities of 18 structurally different flavonoids were evaluated by FRAP assay modified to be used in 96-well microplates. Furthermore, their oxidation potentials were also measured, which were in the range of +0.3 V (myricetin) to +1.2 V (5-hydroxy flavone) and were in good agreement with FRAP assay results. Quercetin, fisetin and myricetin had the lowest oxidation potentials and appeared the most active compounds in FRAP assay and were 3.02, 2.52 and 2.28 times more active than Trolox, respectively. Indications were found that the o-dihydroxy structure in the B ring and the 3 hydroxy group and 2,3-double bond in the C ring give the highest contribution to the antioxidant activity. PMID- 15652193 TI - Regulation of acetylated tubulin/Na+,K+-ATPase interaction by L-glutamate in non neural cells: involvement of microtubules. AB - A subpopulation of membrane tubulin consisting mainly of the acetylated isotype is associated with Na+,K+-ATPase and inhibits the enzyme activity. We found recently that treatment of cultured astrocytes with L-glutamate induces dissociation of the acetylated tubulin/Na+,K+-ATPase complex, resulting in increased enzyme activity. We now report occurrence of this phenomenon in non neural cells. As in the case of astrocytes, the effect of L-glutamate is mediated by its transporters and not by specific receptors. In COS cells, the effect of L glutamate was reversed by its elimination from culture medium, provided that d glucose was present. The effect of L-glutamate was not observed when Na+ was replaced by K+ in the incubation medium. The ionophore monensin, in the presence of Na+, had the same effect as L-glutamate. Treatment of cells with taxol prevented the dissociating effect of L-glutamate or monensin. Nocodazole treatment of intact cells or isolated membranes dissociated the acetylated tubulin/Na+,K+-ATPase complex. The dissociating effect of nocodazol does not require Na+. These results indicate a close functional relationship among Na+,K+ ATPase, microtubules, and L-glutamate transporters, and a possible role in cell signaling pathways. PMID- 15652194 TI - Purification and partial characterization of marinocine, a new broad-spectrum antibacterial protein produced by Marinomonas mediterranea. AB - This work describes the purification and partial characterization of a novel antibacterial compound, here named marinocine, produced by Marinomonas mediterranea, a melanogenic marine bacterium with rich secondary metabolism. The antibacterial compound is a protein detected in the medium at death phase of growth. It has been purified to apparent homogeneity from the supernatants of cultures by means of ethanol precipitation followed by column chromatographies on DEAE-Sephadex and Sephacryl HR-200. The protein has an apparent molecular mass of 140-170 kDa according to gel permeation chromatography and non-denaturing SDS PAGE, although in denaturing SDS-PAGE two mayor bands of 97 and 185 kDa appear. Marinocine is relatively heat-stable and shows a great resistance against many hydrolytic enzymes such as glycosidases, lipase, and proteases. The antibacterial range of the molecule includes Gram-positive and Gram-negative microorganisms, as well as some nosocomial isolates, Staphylococcus aureus and Pseudomonas sp., highly resistant to classical antibiotics. By contrast, marinocine did not show any effect on the eukaryotic microorganisms tested. Regarding eukaryotic CHO cells, the decrease on viability was much lower than the one observed on bacterial cells. PMID- 15652195 TI - Protein aggregation and its inhibition in biopharmaceutics. AB - Protein aggregation is arguably the most common and troubling manifestation of protein instability, encountered in almost all stages of protein drug development. Protein aggregation, along with other physical and/or chemical instabilities of proteins, remains to be one of the major road barriers hindering rapid commercialization of potential protein drug candidates. Although a variety of methods have been used/designed to prevent/inhibit protein aggregation, the end results are often unsatisfactory for many proteins. The limited success is partly due to our lack of a clear understanding of the protein aggregation process. This article intends to discuss protein aggregation and its related mechanisms, methods characterizing protein aggregation, factors affecting protein aggregation, and possible venues in aggregation prevention/inhibition in various stages of protein drug development. PMID- 15652197 TI - Determinations of ephedrine in mixtures of ephedrine and pseudoephedrine using diffuse reflectance infrared spectroscopy. AB - There are a number of situations where there is a need to determine the concentrations of components in solid-state mixtures without dissolving the sample. Diffuse reflectance infrared Fourier transform spectroscopy (DRIFTS) coupled with partial-least-squares (PLS) data analysis has been used to determine the minor component in a mixture of structurally similar solid-state compounds, in this case mixtures of ephedrine and pseudoephedrine. Factors that limit the precision and accuracy of the determinations are discussed. It is shown that when care is taken to produce homogeneous calibration samples very good results can be obtained, in this case cross-validated standard error of predictions of 0.74 wt% when the minor component spanned the concentration range of 0-50 wt%, and 0.11 wt% when the minor component spanned the concentration range of 0-5 wt%. Results are presented that indicate that the amount of data available to the PLS calibration routine relative to the range over which the calibration is performed can limit the precision and accuracy of the determinations. PMID- 15652196 TI - A study of the differences between two amorphous spray-dried samples of cefditoren pivoxil which exhibited different physical stabilities. AB - The objective of this study was to investigate the reasons for the difference in physical stability of two amorphous cefditoren pivoxil samples that had been prepared using spray drying at inlet-air temperatures of 40 degrees C (SD-A) and 100 degrees C (SD-B). The two samples appeared amorphous by powder X-ray diffraction and had indistinguishable glass transition temperatures. Despite the fact that glass transition is often regarded as an indicator of the stability of amorphous forms, crystallisation was observed for SD-A, but not for SD-B, during storage at 60 degrees C and 81% relative humidity (RH). Gravimetric water sorption data demonstrated very similar water sorption until high RH values, at which point SD-A sorbed more water than did SD-B. The values of the dispersive, acidic (K(A)) and basic (K(D)) components of surface energy of the spray-dried samples were obtained using inverse gas chromatography (IGC), in the dry state and after equilibration with different RH environments. The data showed that the two amorphous samples had different surface properties and that the effect of sorbed water on these samples was also different. It is concluded that the two samples did not have long-range order, but had differences in the orientation of molecules at the surface, which were significant enough to alter the stability when the samples were stressed with water vapour and high temperature storage. IGC proved a valuable tool with which to study changes in the surface properties of amorphous materials. PMID- 15652198 TI - Lyophilised wafers as a drug delivery system for wound healing containing methylcellulose as a viscosity modifier. AB - Lyophilised wafers have potential as drug delivery systems for suppurating wounds. A dual series of wafers made from low molecular weight sodium alginate (SA) and xanthan gum (XG) respectively, modified with high molecular weight methylcellulose (MC) were produced. The swelling and flow properties of these wafers on model suppurating surfaces were both qualitatively and quantitatively investigated. The wafers instantaneously adhered to the surfaces, absorbing water and transforming from glassy, porous solids to highly viscous gels. The rate at which this occurred varied for the series studied with clear distinctions between the behaviour of SA and XG systems. For SA wafers there was a distinct relationship between the flow-rate and MC content. Increased amounts of MC decreased the rate at which the SA wafers flowed across a model gelatine surface. Flow rheometry was used to quantify the effect of increased MC content on both series of wafers and for the SA series, highlighted a substantial increase in apparent viscosity as a function of incremental increases in MC content. These results reflected those from the gelatine model. Observations of the reluctance of a swollen, unmodified XG wafer to flow compared with the relative ease of unmodified, low molecular weight SA was attributed to the yield stress characteristic of xanthan gels. XG is known to exhibit complex, loosely bound network structures in solution via the association of helical backbone structures. The inclusion of sodium fluorescein as a visible model for a soluble drug highlighted the potential of lyophilised wafers as useful drug delivery systems for suppurating wounds. PMID- 15652199 TI - Studies on the poly(lactic-co-glycolic) acid microspheres of cisplatin for lung targeting. AB - Lung-targeting cisplatin-loaded poly(lactic-co-glycolic) acid microspheres (CDDP PLGA-MS) were prepared by a solvent evaporation method. The uniform design was used to optimize the technology of preparation, the appearance and size distribution were examined by scanning electron microscope, and the aspects such as in vitro release characteristics, stability, drug loading, loading efficiency, pharmacokinetics and tissue distribution in rabbit were studied. The experimental results showed that the microspheres were globular in appearance and dispersed well. The average particle size was 12.8 microm with 98% of the microspheres being in the range of 5-30 microm. The drug loading and loading efficiency were 17.68 and 53.2%, respectively. The in vitro release behavior could be expressed by the following equation: 1-Q=0.424e(-0.360t)+0.474e(-0.001t). After i.v. administration (15 min), the drug concentration of microspheres group in lung in rabbits was 212 microg/g, while that of controlled group was 1.37 microg/g. CDDP PLGA-MS showed a combination of lung-targeting and sustained drug release in experiments on rabbits. PMID- 15652201 TI - In vivo evaluation of time and site of disintegration of polysaccharide tablet prepared for colon-specific drug delivery. AB - Compression coating has been found to be useful for colonic drug delivery. The aim of the present investigation was to evaluate a formulation with a considerably reduced coat weight and gum concentration for colonic drug delivery in vivo using gamma scintigraphy. In vitro studies have found this formulation to be useful for delivery of 5-fluorouracil to the colon. Rapidly disintegrating core tablets containing (99m)Tc-DTPA were prepared and compression coating with 150 mg of granules containing a mixture of xanthan (XG), guar gum (GG) and starch. The ratios of the two gums XG:GG in the coat was kept 10:20. In vitro dissolution studies on XG:GG::10:20 tablets containing (99m)Tc-DTPA were carried out in simulated upper GIT conditions and also in presence of colonic contents. Cumulative percent release of technetium in the upper GIT conditions and transit time amounted to 4%. The total amount of technetium released in the 24 h of the dissolution study was 53+/-3.23%. Upon introduction of cecal content into the dissolution medium (4%), the release of technetium from the compression-coated tablet increased to 78.34+/-5.34%. Gamma scintigraphy studies carried out in six healthy human volunteers showed that the tablet remained intact during its transit through the upper GIT. The anatomical site of disintegration was found to be the ascending colon/hepatic flexure and the disintegration of the tablet started between 4 and 6 h post-dose in all the volunteers with a further spread of tracer into the ascending, transverse, descending and sigmoidal colon. PMID- 15652200 TI - Effect of adsorbents on the absorption of lansoprazole with surfactant. AB - Lansoprazole (LPZ) is a representative drug that shows a high inter-subject variation of bioavailability (BA). Solid preparation composed of surfactant, adsorbent and LPZ were prepared to improve the dissolution and absorption of LPZ, and the BA of LPZ was measured in rats and dogs. As surfactant, Tween 80, polyoxy 60 hydrogenated caster oil derivative (HCO-60) and PEG-8 caprylic/capric glycerides (Labrasol) were used. As adsorbant, porous silicon dioxide (Sylysia 550, 320), magnesium aluminometa silicate (Neusilin S2, NS2N, US2) and porous calcium silicate (Florite RE) were used. After small intestinal administration of LPZ, 5.0 mg/kg, solution with HCO-60 showed the highest plasma LPZ concentration versus time curve of which C(max) and AUC was 0.46+/-0.01 microg/mL and 0.73+/ 0.03 microgh/mL. By comparing to that after i.v. injection of LPZ solution, 2.0 mg/kg, the BA of LPZ from HCO-60 solution was 39.0%, which was about seven times higher than that of LPZ powder. To solidify the LPZ solution with HCO-60, adsorbents were used and the obtained solid preparations were used for in vitro release experiment. Sylysia 320, Neusilin S2 and Neusilin NS2 showed the T50% of about 1h. To evaluate the BA of these solid preparations, absorption study was performed in rats. Sylysia 550 system showed the higher AUC than other systems, showing the BA of 28.1%. Sylysia 550 system was filled in an enteric capsule and was orally administered to dogs and BA was compared with enteric tablet. The AUC of Sylysia 550 system was 2.16+/-0.26 microgh/mL and was greater than enteric tablet and the BA of 71.7% was obtained. Solid system composed of LPZ, surfactant and adsorbent has suggested the possibility as a good tool to improve the BA of LPZ. PMID- 15652202 TI - Optimization of pH-independent release of nicardipine hydrochloride extended release matrix tablets using response surface methodology. AB - The purpose of this study was to optimize the pH-dependent release of nicardipine hydrochloride extended release formulations by using simultaneously combination two hydrophilic polymers: hydroxypropylmethylcellulose (HPMC) and sodium alginate as retardant and avicel as additive. The constrained mixture experimental design was used to prepare systematic model formulations which were composed of three formulation variables: the content of HPMC (X1), avicel (X2), and sodium alginate (X3). The response surface methodology (RSM) and multiple response optimization utilizing the polynomial equation were used to search for the optimal formulation with specific release rate at different time intervals and to quantify the effect of each formulation variables. The drug release percent at 3, 6 and 12 h were the target responses and were restricted to 10-30% (Y3h), 40-65% (Y6h) and not less than 80% (Y12h), respectively. The results showed that the effect of combination of HPMC and sodium alginate was the most influence factor on the drug release from extended-release matrix tablets. The observed results of Y3h, Y6h and Y12h coincided well with the predictions in the RSM optimization technique, indicating it was quite useful for optimizing pharmaceutical formulation. The mechanism of drug release from extended-release matrix tablets was dependent on the added amount of alginate. The release kinetic of drug from HPMC matrix tablets with alginate was followed the zero-order release pattern. PMID- 15652203 TI - A new scleroglucan/borax hydrogel: swelling and drug release studies. AB - The aim of the work was the characterization of a new polysaccharidic physical hydrogel, obtained from Scleroglucan (Sclg) and borax, following water uptake and dimension variations during the swelling process. Furthermore, the release of molecules of different size (Theophylline (TPH), Vitamin B12 (Vit. B12) and Myoglobin (MGB)) from the gel and from the dried system used as a matrix for tablets was studied. The increase of weight of the tablets with and without the loaded drugs was followed together with the relative variation of the dimensions. The dry matrix, in the form of tablets was capable, during the swelling process, to incorporate a relevant amount of solvent (ca. 20 g water/g dried matrix), without dissolving in the medium, leading to a surprisingly noticeable anisotropic swelling that can be correlated with a peculiar supramolecular structure of the system induced by compression. Obtained results indicate that the new hydrogel can be suitable for sustained drug release formulations. The delivery from the matrix is deeply dependent on the size of the tested model drugs. The experimental release data obtained from the gel were satisfactorily fitted by an appropriate theoretical approach and the relative drug diffusion coefficients in the hydrogel were estimated. The release profiles of TPH, Vit. B12 and MGB from the tablets have been analyzed in terms of a new mathematical approach that allows calculating of permeability values of the loaded drugs. PMID- 15652204 TI - Influence of the morphogranulometry and hydrophobicity of talc on its antisticking power in the production of tablets. AB - Antisticking power varies according to the talc considered. It is difficult to define the physical properties of talc implicated in its antisticking power. In this work, different talcs were characterized and an evaluation made of their performance in reducing sticking in tablet manufacturing. Determination of the specific surface area was made by permeametry, morphogranulometric analysis by laser diffractometry using a method, which made it possible to assess the mean thickness of talc particles, and measurement of water absorption kinetics was taken to assess hydrophobicity. The relationship between the characteristics of talcs and their antisticking power was then considered. There is a correlation between the particle size of talc and surface hydrophobicity. The detaching force of tablets appears to be dependent on the basal dimension of talc. PMID- 15652206 TI - Studies on the reactions between daptomycin and glyceraldehyde. AB - The objectives of this project were to determine the reaction pathways of daptomycin in the presence of glyceraldehyde in acidic solutions, and to quantitate the kinetics of the major pathways. In the presence of glyceraldehyde (pH range 1-7 at 25 to 60 degrees C), daptomycin formed two major products separable by RP-HPLC. The products were identified using UV spectroscopy, fluorimetry, mass spectrometry, and 2D-1H NMR. The reaction scheme involved the reversible formation of imine and anilide derivatives. Carbinolamine was believed to be a common intermediate in formation pathways of both products. The carbinolamine intermediate underwent either acid catalyzed dehydration resulting in imine formation or intramolecular hydrogen bonding and bond cleavage giving rise to anilide formation. In mild acid conditions, both products reversed to daptomycin. The reaction between daptomycin and glyceraldehyde was first-order with respect to both reactants. In a pH range of 1-7, the imine formation rate was pH dependent with a maximum rate at approximate pH values of 3-4. The observed pH dependence was consistent with the pH dependence of typical amine aldehyde reactions. PMID- 15652205 TI - The evaluation of cosmetic and pharmaceutical emulsions aging process using classical techniques and a new method: FTIR. AB - The purpose of this paper is to show how the utilization of Fourier Transform Infrared (FTIR) spectroscopy can be interesting in stability studying of cosmetic or pharmaceutical "oil in water" (O/W) emulsions. In this study temperature storage tests were performed to accelerate the aging process and evaluate the stability of five emulsions. Emulsions were analyzed by FTIR and classical methods (conductivity, viscosity, pH, texture analysis) in order to determine a method that would enable predicting the emulsion's stability. During the aging process, modifications of chemical functions are measured by FTIR (using spectrometric indices), such modifications included: a decrease of unsaturation index, an increase of carbonyl index and a broadening of the carbonyl band. This band was deconvoluted to evaluate the contribution of different species in the broadening phenomenon, which seems to be caused by the appearance of free fatty acids. Conductimetry seems to be the most sensitive technique to assess physical modifications during emulsion's aging. Concerning the most unstable emulsions, a progressive increasing of conductivity was observed several months before the emulsion destabilizes. Consequently, FTIR and conductimetry are two complementary techniques. Conductimetry is a useful technique to predict emulsion destabilization while FTIR allows the measurement of chemical modifications and helps to understand the chemical mechanisms which occur during the oxidation. PMID- 15652207 TI - In vivo distribution of arsenic after i.p. injection of arsonoliposomes in balb-c mice. AB - We recently showed that arsonoliposomes (novel arsenic containg liposomes) demonstrate differential toxicity towards various types of cancer and normal cells, in cell culture studies, as well as anti-parasitic activity. In this study, the in-vivo distribution of the active moiety of these vesicles, As, is evaluated. Sonicated arsonoliposomes were prepared using the arsonolipid with palmitic acid acyl chain (C16) mixed with egg-phosphatidyl choline (PC) and cholesterol (Chol) [C16/PC/Chol at 8:12:10 mol/mol/mol]. A dose of arsonoliposomes, corresponding to 5 mg arsenate/kg was administered by intraperitoneal injection in balb-c mice. At various time points post-injection the mice were sacrificed and the distribution of As in the organs was measured, by atomic absorption spectroscopy. Results demonstrate that a high portion of the dose administered is rapidly excreted; since 1-h post-injection only about 30% of the dose administered was detected cumulatively in the animal tissues. After this the elimination of arsenic was a slow process with a total body elimination rate constant of 0.023 h(-1), corresponding to a half-life of 30 h. Tissues with the highest arsenic concentration during the study period are: spleen-kidneys stomach, followed by lung, liver, intestines-heart, carcass+skin and finally blood. No acute toxicity, or effect on the body or organ weight of the mice was observed. PMID- 15652208 TI - A novel method to calculate the extent and amount of drug transported into CSF after intranasal administration. AB - The aim of this paper is to establish a novel method to calculate the extent and amount of drug transported to brain after administration. The cerebrospinal fluid (CSF) was chosen as the target region. The intranasal administration of meptazinol hydrochloride (MEP) was chosen as the model administration and intravenous administration was selected as reference. According to formula transform, the extent was measured by the equation of X(A)CSF, infinity/X0 = Cl(CSF) AUC(0-->infinity)CSF/X0 and the drug amount was calculated by multiplying the dose with the extent. The drug clearance in CSF (Cl(CSF)) was calculated by a method, in which a certain volume of MEP solution was injected directly into rat cistern magna and then clearance was assessed as the reciprocal of the zeroth moment of a CSF level-time curve normalized for dose. In order to testify the accurateness of the method, 14C-sucrose was chosen as reference because of its impermeable characteristic across blood-brain barrier (BBB). It was found out that the MEP concentrations in plasma and CSF after intranasal administration did not show significant difference with those after intravenous administration. However, the extent and amount of MEP transported to CSF was significantly lower compared with those to plasma after these two administrations. In conclusion, the method can be applied to measure the extent and amount of drug transported to CSF, which would be useful to evaluate brain-targeting drug delivery. PMID- 15652209 TI - Skin permeation of propranolol from polymeric film containing terpene enhancers for transdermal use. AB - To develop the suitable film formulations of propranolol hydrochloride (PPL) containing enhancers for transdermal use, polymeric film formulations were prepared by employing ethyl cellulose (EC) and polyvinyl pyrrolidone (PVP) as a film former, and dibutyl phthalate (DBP) as a plasticizer. Terpenes such as menthol and cineole, and propylene glycol (PG) were also employed as a chemical enhancer to improve the skin penetration of PPL. The film preparations were characterized in physical properties such as uniformity of drug content, thickness and moisture uptake capacity. Release and skin permeation kinetics of PPL from film preparations were examined in the in vitro studies using a Franz type diffusion cell. The uniformity of drug content was evidenced by the low S.D. values for each film preparation. The moisture uptake capacity and drug release rate increased with the increase of PVP in each preparation. Enhancers examined in the present study also increased the moisture uptake capacity and release rate of PPL from the film preparations. Increasing the concentration of PPL from 1 to 2 mg/cm2 in the film enhanced the release rate of PPL, while no effect of enhancer concentrations on the release rate from the film preparations was observed. In vitro skin permeation study showed that cineole was the most promising enhancer among the enhancers examined in the present study and suggested that the suitable compositions of film preparation would be EC:PVP:PPL=6:3:4 with 10% (w/w) cineole and 7:2:4 with 10% (w/w) PG and cineole, which provided high skin permeation rates at 93.81+/-11.56 and 54.51+/-0.52 microg/cm2/h, respectively. PMID- 15652210 TI - Physicochemical studies on Ciclopirox olamine complexes with divalent metal ions. AB - Ciclopirox olamine (CPO) metal complexes have been prepared and characterized using elemental analysis, infra red (IR), melting point and differential scanning calorimetry (DSC). Spectroscopic titration using molar ratio method indicated the occurrence of 1:1 complexes for CPO with almost all the examined metals. Physicochemical properties were also studied including aqueous solubility and apparent partition coefficient. Results showed that generally complex formation dramatically decreased the solubility and increased apparent partition coefficient. However, some metal complexes exhibited opposite effect. It could be concluded that complex formation can modify the solubility and apparent partition coefficient, which may suggest the use of complexes to manipulate the physicochemical properties of the drug. PMID- 15652211 TI - Labeling efficiency and biodistribution of Technetium-99m labeled nanoparticles: interference by colloidal tin oxide particles. AB - The interference of colloidal tin oxides on the biodistribution of (99m)Technetium radiolabeled chitosan nanoparticles has been overcome by using sodium borohydride instead of commonly used stannous salts as reducing agent for the reduction of (99m)Tc (VII) to lower valency states. Biodistribution of radiolabeled chitosan nanoparticles prepared by using stannous chloride method revealed localization of the radioactivity mainly in the liver and spleen while that of radiolabeled chitosan nanoparticles prepared by using sodium borohydride method manifested the presence of radioactivity in blood up to an extent of 10% even after 2 h. Interestingly, the reduction of radioactivity in the latter case with the progress of time was not manifested through an increase in activity in the liver. Rather, a time dependent increased accumulation of radioactive materials was observed in the stomach. From the results it has been concluded that the biodistribution is strongly influenced by the presence of colloidal particles of tin oxides and (99m)Tc labeled chitosan nanoparticles are RES evading and long circulating in blood when Tc (VII) is reduced by sodium borohydride and not by stannous chloride during radiolabeling process. PMID- 15652212 TI - Physicochemical characterization and gene transfection efficiency of lipid emulsions with various co-emulsifiers. AB - Transfection systems based on complexes of DNA and lipid emulsions were evaluated with respect to their effectiveness, toxicity, physicochemical characteristics, and cell-type dependence. The potential of a series of co-emulsifiers to serve as vectors was investigated. The co-emulsifiers examined included 1,2-dioleoyl-sn glycero-3-phosphoethanolamine (DOPE), Tween, cholesterol, stearylamine, and polyethylenimine (PEI). Squalane and 1,2-dioleoyl-sn-glycero-3-trimethylammonium propane (DOTAP), respectively, were the main oil phase and cationic lipid added to the lipid emulsions. Cell viability was reduced after inclusion of either of the two cationic components of stearylamine and PEI. DOPE and cholesterol showed both higher transfection activity and cell viability as compared to the other co emulsifiers. The incorporation of DOPE and cholesterol also prevented droplet aggregation of the emulsions after long-term storage. Results of the transfection of COS-1, A549, or HaCat cell lines with lipid emulsions indicated differences in transfection activities of each formulation for the different cell lines. It is concluded that DOPE and cholesterol as co-emulsifiers for DOTAP were preferable for stability and DNA transfection of emulsions. PMID- 15652213 TI - Protective properties of melatonin-loaded nanoparticles against lipid peroxidation. AB - The aim of this study was to prepare melatonin-loaded nanoparticles (nanocapsules and nanospheres) by nanoprecipitation, using Eudragit S100 as polymer. The potential of these systems to protect lipids against peroxidation was evaluated in comparison to melatonin in aqueous solution and nanoemulsion. Liposomes and microsomes were used as model of a lipid membrane and lipid peroxidation was induced by free radical ascorbyl. Nanocapsule and nanosphere suspensions presented total recoveries of melatonin near 100% and associated drug around 55%. The zeta potential values were negative and the hydrodynamic diameter of particles were lower than 255 nm. The results demonstrate that the lipids were protected against peroxidation from 8 to 51% due to the presence of the melatonin and that this effect depended on the drug dose, the type of the lipid substrate and the type of colloid, in which melatonin was incorporated. Nanocapsules and nanospheres provided an important increase in the antioxidant effect of melatonin against lipid peroxidation. PMID- 15652214 TI - Multiplex PCR detection and species differentiation of orthopoxviruses pathogenic to humans. AB - A method for one-stage rapid identification of four orthopoxvirus species pathogenic to humans based on multiplex polymerase chain reaction (MPCR) was developed. Five pairs of oligonucleotide primers--one, genus-specific; and the rest, species-specific for variola, monkeypox, cowpox, and vaccinia viruses, respectively--were used concurrently for MPCR assay of orthopoxvirus DNAs. Specificity and sensitivity of the method developed were evaluated using DNAs of 57 orthopoxvirus strains, including the DNAs isolated from human case clinical materials. PMID- 15652215 TI - Rapid presumptive identification of Burkholderia pseudomallei with real-time PCR assays using fluorescent hybridization probes. AB - Burkholderia pseudomallei (the etiologic agent of melioidosis) can cause pyogenic or granulomatous lesions in almost any organ. Septicemia has a case fatality rate of >40%. Early diagnosis and appropriate antibiotic therapy are crucial for survival, but cultivation, biochemical identification, and conventional PCR of B. pseudomallei are time consuming. We established real-time PCR assays using fluorescent hybridization probes targeting the 16S rDNA, the flagellin C (fliC) and the ribosomal protein subunit S21 (rpsU) genes. The test sensitivity and specificity were assessed with a representative panel of 39 B. pseudomallei, 9 B. mallei, 126 other Burkholderia strains of 29 species, and 45 clinically relevant non-Burkholderia organisms. The detection limit for the 16S rDNA, fliC, and rpsU assay was 40, 40, and 400 genome equivalents per reaction, however, in spiked blood samples it was 300, 300, and 3000, respectively. Specificity, positive and negative predictive value of the assays was 100%. In conclusion, we recommend the use of the 16S rDNA and/or fliC real-time PCR assays for the rapid identification of B. mallei and B. pseudomallei in positive blood cultures or from suspicious bacterial colonies. PMID- 15652216 TI - Specific identification of Egyptian Biomphalaria species and possible hybrids using the polymerase chain reaction based on nuclear and mitochondrial loci. AB - The snail historically implicated in the transmission of Schistosoma mansoni in Egypt is Biomphalaria alexandrina. The problem of schistosomiasis in Egypt has been complicated in recent years by the introduction of Biomphalaria glabrata, which has been reported to hybridize with B. alexandrina. Both introduced and hybrid snails also pose a threat with respect to S. mansoni transmission. As morphological differentiation of these snails is difficult, using three DNA loci, nuclear ITS1 and ITS2, and mitochondrial ND1, PCR-based assays were developed to identify these species and possible hybrids. The assays are rapid, reproducible, sensitive and specific. This technique may be used in field surveys to study the distribution of the two species of intermediate host and their putative hybrids in Egypt. PMID- 15652217 TI - Molecular characterization of multidrug-resistant Enterococcus spp. from poultry and dairy farms: detection of virulence and vancomycin resistance gene markers by PCR. AB - Thirty multidrug-resistant Enterococcus spp. strains, including two from the milk of cows with mastitis, nine from chicken litter and 19 from turkey litter, were isolated. Twenty-five were identified by biochemical methods as E. gallinarum and five as E. faecalis. Most of the isolates were resistant to vancomycin, gentamicin, streptomycin, tetracycline, erythromycin, bacitracin, kanamycin and nalidixic acid but sensitive to ciprofloxacin, sulfamethoxazole, chloramphenicol, ampicillin and ofloxacin. Attempts were made by partial amplification of the gene sequences to detect the vancomycin resistance markers vanA (734-bp), vanB (420 bp), vanC1 (531-bp), and vanC2-C3 (673-bp); virulence markers cylA (427-bp) and cylB (225-bp) for enterococcal cytolysin and a biofilm-forming surface protein (Esp). Individual and multiplex-PCR assays for vancomycin resistance markers revealed the vanC1 gene in 22 E. gallinarum strains. None of the remaining isolates including five E. faecalis strains (MIC=2 microg ml(-1)) and three E. gallinarum strains (MIC=8 microg ml(-1)) had any of the van genes tested. Analysis by pulsed-field gel electrophoresis (PFGE) and a comparison of smaI banding profiles showed 11 different patterns. Probing with a DIG-labeled vanC1 PCR product indicated a common 38.0 kb SmaI DNA fragment in all the E. gallinarum strains harboring the vanC1 gene. The genes cylA and cylB were detected only in one clinical E. gallinarum isolate and two quality control clinical strains of E. faecalis (ATCC 51299 and 29212). None of the virulence factors were found in milk or poultry isolates. Intermediate level resistance to vancomycin in enterococci from the US animal farms was predominantly due to the presence of vanC1 gene. PMID- 15652218 TI - PCR-SSCP as a molecular tool for the identification of Benedeniinae (Monogenea: Capsalidae) from marine fish. AB - PCR-based single strand conformation polymorphism (SSCP) analysis was used to characterize monogenean specimens of the subfamily Benedeniinae of morphologically uncertain specific status from different marine fish species, using Neobenedenia melleni, N. girellae and Entobdella corona for comparison. The first internal transcribed spacer (ITS-1) and the 5' terminal variable region (D1 D3 domains) of the large subunit ribosomal DNA (lsrDNA) were amplified separately from individual monogeneans, and the amplicons were subjected to PCR-SSCP analyses, followed by direct sequencing. Both SSCP patterns and the ITS-1 sequences data allowed specimens representing Entobdella spp. from the host Taeniura melanospilos to be unequivocally distinguished from those representing Neobenedenia spp. and those representing Benedenia spp. Neobenedenia girellae, a morphologically controversial species, had identical SSCP banding pattern and ITS 1 sequence to that of N. melleni, supporting the proposal that N. girellae is a synonym of N. melleni. Neobenedenia spp. and Benedenia spp. had identical SSCP patterns and ITS-1 sequences. These findings and the PCR-SSCP approach taken should have implications for the accurate identification and assessment of taxonomic validity of other important monogenean groups of the marine fish. PMID- 15652219 TI - DNA microarray-based detection and identification of Chlamydia and Chlamydophila spp. AB - A microarray hybridization assay for identification of chlamydiae was developed using the ArrayTube platform. The technology is comparatively inexpensive and involves plastic tube-integrated microchips and signal amplification by enzyme catalyzed silver precipitation. Hybridization probes were designed on the basis of the most variable window approach, which identified species-specific nucleotide polymorphisms in a region of generally high sequence similarity. The selected 26-nt probe sequences were used on two different series of customized microarrays, i.e. combinatorial high-density in situ synthesized arrays and low density spotted arrays. Target DNA was prepared by consensus PCR amplifying a 1 kbp segment of the ribosomal RNA operon. Unique species-specific hybridization patterns were obtained for all nine species of the family Chlamydiaceae on both microarray types. The present assay proved suitable for unambiguous species identification of chlamydial cell cultures and showed a potential for direct detection of these bacteria from clinical tissue. PMID- 15652221 TI - A collection of bovine cDNA probes for gene expression profiling in muscle. AB - Array technology has been increasingly used to monitor global gene expression patterns in various tissues and cell types. However, applications to muscle development and pathology as well as meat production in livestock species have been hampered by the lack of appropriate cDNA collections. To overcome this problem, a directed cDNA library was constructed starting from 23 muscles of meat producing bovines to derive a collection of 3573 clones. A preliminary sequence characterization of this collection indicated that the most abundant transcripts correspond to genes encoding proteins involved in energy metabolism (COX and NADH dehydrogenase subunits) and belonging to the contractile apparatus (myosin chains and troponin isoforms). From this cDNA library, we selected a set of 435 clones representing 340 unique genes, of which 24 were novel. This collection was subsequently completed with 75 specific cDNA probes for genes of interest already studied in our laboratory. The bovine 'muscle' cDNA repertoire thus designed was spotted onto a nylon membrane (macroarray) in order to test its utility to further investigate the transcriptome of bovine muscles in relation to meat quality traits. It is also anticipated that this type of collection might be useful for the study of chronic myologic diseases in other mammalian species, including humans. PMID- 15652220 TI - Development of a novel internal positive control for Taqman based assays. AB - Development of rapid amplification assays for the detection and identification of biological threat agents has become a focus of diagnostic efforts in recent years. The use of real-time PCR assays as diagnostic tools depends upon two critical processes. First, nucleic acid purification must provide template that is both amplifiable and free of PCR inhibitors. Second, the assays themselves must be sensitive and specific for their nucleic acid targets. A differentiation must be made between results achieved due to the lack of target nucleic acid (true negatives) and those produced due to the inability to amplify target DNA (false negatives) so confidence in negative reactions is possible. False negatives can occur when inhibitors are present in the sample being tested, especially if clinical samples such as blood are analyzed. To address the problem of detecting inhibition in purified nucleic acids, an exogenous internal positive control (IPC) based on Taqman chemistry was developed. A previously optimized assay was cloned and the primer and probe sites were mutated to produce novel sequences with no known homology to published sequence data. The IPC was sensitive to a variety of inhibitors, including hemoglobin, heparin, EDTA, humic acids, and fulvic acid. It was also equally sensitive to inhibition when labeled with either 6FAM or ROX dyes. In addition, the IPC was successfully multiplexed with agent specific assays without any loss in their sensitivity. The designed IPC assay has proven to be an effective tool for monitoring inhibitors of PCR and builds confidence in negative results obtained with agent specific assays. PMID- 15652222 TI - Characterization of a single nucleotide polymorphism in the ZNF9 gene and analysis of association with myotonic dystrophy type II (DM2) in the Italian population. AB - Myotonic dystrophy type 2 (DM2) is a dominant inherited disorder clinically similar to myotonic dystrophy type 1 (DM1) with a peculiar pattern of multisystemic phenotypic features. The mutation responsible for DM1 is a CTG repeat in the 3' UTR of the dystrophia myotonica protein kinase gene (DMPK) on chromosome 19q13.3, while DM2 is caused by an unstable CCTG expansion in intron 1 of the zinc finger protein 9 gene (ZNF9) on chromosome 3q21.3. Southern blotting analysis is the conventional test used to determinate the size of the repeats in the molecular diagnosis of DM2. However, the large number of CCTG repeats and their somatic instability complicates this diagnostic protocol. In order to improve the DM2 test, we have recently characterised a single nucleotide polymorphism located in the first intron of the ZNF9 gene. This SNP consists in a C to A nucleotide change, which creates or disrupts and ApaI enzyme restriction site, easily detectable by PCR amplification followed by restriction analysis. We genotyped this SNP in 30 unrelated DM2 patients and 70 unrelated Italians healthy individuals. Our results show that this polymorphism is in linkage disequilibrium with the DM2 mutation. PMID- 15652223 TI - The importance of mutation, then and now: studies with yeast cytochrome c. AB - The development of a genetic system based on the CYC1 gene was initiated over 40 years ago, primarily because of the anticipated ease of sequencing of the corresponding encoded protein, iso-1-cytochrome c from Saccharomyces cerevisiae. The success of the iso-cytochrome c system was dependent on the early development of methods for detecting and selecting cyc1 defective mutants and CYC1 functional revertants, and of methods for fine-structure genetic mapping using deletions and single-site mutations. The nonsense codons TAA and TAG, and the initiation codon ATG, were determined from the amino acid alterations of iso-1-cytochromes c from intragenic revertants; this represented the first assignments of such codons in a eukaryotic organism. The types of desired sequences were expanded by selecting recombinants from cyc1 x cyc1 nonfunctional mutants or CYC1 x CYC1 functional mutants, permitting the early determination of the rules of translation, which differed from those of prokaryotes by use of the most 5' AUG codon for initiation of translation. The sequence of 44 base pairs of CYC1 was determined with altered iso-1-cytochromes c from revertants of frameshift and initiation mutants, allowing the early cloning of the gene. A method was developed for transforming yeast directly with synthetic oligonucleotides, resulting in the convenient production of CYC1 mutants with defined sequences. At this point in time, Sherman and colleagues have published approximately 240 papers on or using the iso cytochrome c system, dealing with such diverse topics as translation, informational suppressors, transcription and transcription termination, recombination, ectopic recombination, mutagen specificity, regulation by Ty1 elements, evolution of duplicated chromosomal segments, structure-function relationships of cytochrome c, protein stability and degradation, biosynthesis and mitochondrial import of cytochrome c, mitochondrial proteases, co- and post translational modifications, and mRNA degradation. Current work on degradation of proteins in mitochondria, on degradation of mRNA in the nucleus, and on N terminal acetylation stems from properties of CYC1 mutants isolated in early screens more than a decade ago. PMID- 15652224 TI - Scientific authorship. Part 1. A window into scientific fraud? AB - The examination of a single scientific manuscript seldom alerts scientists, reviewers, editors, and scientific administrators to the fabrication and falsification of data and information. This review shows that most documented cases of scientific fraud involve falsification (altering truthful information) and fabrication (inventing information where none previously existed). Plagiarism is much less frequent. The review of published accounts also shows that the publication of scientific papers containing recognizable fraudulent material is very low, probably less than 0.02% and extremely difficult to detect. Because most reported cases of fraud have involved research done at prestigious organizations with distinguished co-authors, and that is published in journals with exacting review processes, it becomes evident that some unscrupulous scientists are adept at fabricating and falsifying data. However, "significant" scientific fraud is detected when scientists repeatedly report results that cannot be independently verified, when colleagues report suspicious behavior, or scientific audits are performed. This review documents and compares many of the better-known cases of scientific fraud. Fraudulent behavior has served as the impetus for the scientific community to develop publication procedures and guidelines that help to guard against not only fraudulent behavior but also against other types of unethical or undesirable behaviors. A companion paper reviews the non-fraudulent issues associated with scientific publication. PMID- 15652225 TI - Scientific authorship. Part 2. History, recurring issues, practices, and guidelines. AB - One challenge for most scientists is avoiding and resolving issues that center around authorship and the publishing of scientific manuscripts. While trying to place the research in proper context, impart new knowledge, follow proper guidelines, and publish in the most appropriate journal, the scientist often must deal with multi-collaborator issues like authorship allocation, trust and dependence, and resolution of publication conflicts. Most guidelines regarding publications, commentaries, and editorials have evolved from the ranks of editors in an effort to diminish the issues that faced them as editors. For example, the Ingelfinger rule attempts to prevent duplicate publications of the same study. This paper provides a historical overview of commonly encountered scientific authorship issues, a comparison of opinions on these issues, and the influence of various organizations and guidelines in regards to these issues. For example, a number of organizations provide guidelines for author allocation; however, a comparison shows that these guidelines differ on who should be an author, rules for ordering authors, and the level of responsibility for coauthors. Needs that emerge from this review are (a) a need for more controlled studies on authorship issues, (b) an increased awareness and a buy-in to consensus views by non-editor groups, e.g., managers, authors, reviewers, and scientific societies, and (c) a need for editors to express a greater understanding of authors' dilemmas and to exhibit greater flexibility. Also needed are occasions (e.g., an international congress) when editors and others (managers, authors, etc.) can directly exchange views, develop consensus approaches and solutions, and seek agreement on how to resolve authorship issues. Open dialogue is healthy, and it is essential for scientific integrity to be protected so that younger scientists can confidently follow the lead of their predecessors. PMID- 15652226 TI - Bile acids as carcinogens in human gastrointestinal cancers. AB - Bile acids were first proposed to be carcinogens in 1939 and 1940. On the basis of later work with rodent models, bile acids came to be regarded as cancer promoters rather than carcinogens. However, considerable indirect evidence, obtained more recently, supports the view that bile acids are carcinogens in humans. At least 15 reports, from 1980 through 2003, indicate that bile acids cause DNA damage. The mechanism is probably indirect, involving induction of oxidative stress and production of reactive oxygen species that then damage DNA. Repeated DNA damage likely increases the mutation rate, including the mutation rate of tumor suppressor genes and oncogenes. Additional reports, from 1994 through 2002, indicate that bile acids, at the increased concentrations accompanying a high fat diet, induce frequent apoptosis. Those cells within the exposed population with reduced apoptosis capability tend to survive and selectively proliferate. That bile acids cause DNA damage and may select for apoptosis-resistant cells (both leading to increased mutation), indicates that bile acids are likely carcinogens. In humans, an increased incidence of cancer of the laryngopharyngeal tract, esophagus, stomach, pancreas, the small intestine (near the Ampulla of Vater) and the colon are associated with high levels of bile acids. The much larger number of cell generations in the colonic (and, likely, other gastrointestinal) epithelia of humans compared to rodents may allow time for induction and selection of mutations leading to cancer in humans, although not in rodents. PMID- 15652227 TI - Review of the in vivo genotoxicity tests performed with styrene. AB - Results from new genotoxicity tests in laboratory animals have necessitated a comprehensive re-evaluation of the mutagenic potential of styrene in vivo. Available data suggest that styrene, after being metabolized to styrene oxide, is weakly positive in indicator tests detecting DNA adducts, DNA strand breaks and sister chromatid exchanges (SCEs). There is no convincing evidence of styrene clastogenicity in experimental animals when the quality of the studies and the plausibility of the test results are considered. Equivocal results were obtained after exposure to high doses causing lethality. A recently published in vivo micronucleus test (MNT) in bone marrow cells of mice conforming to the current OECD guideline was clearly negative. Consequently, our evaluation of the published genotoxicity data comes to the conclusion that styrene at high doses can induce genotoxic effects in indicator tests. These DNA effects depend upon the exposure levels of the target cells, the metabolic activation to styrene oxide and the efficiency of detoxification. Mutagenic effects of styrene can only be expected under extreme exposure conditions if styrene oxide is not efficiently detoxified and primary DNA lesions are not completely repaired. However, there is no clear evidence that styrene induces mutagenic/clastogenic effects in vivo when tested under appropriate test conditions. PMID- 15652228 TI - Modulating the cytokine response to treat Helicobacter gastritis. AB - The conventional view of gastric acid secretion is that a negative feedback mechanism arises in response to high acidity, such that somatostatin keeps G cells and parietal cells from producing more gastrin and acid, respectively. When the stomach becomes infected, for example with Helicobacter pylori (H. pylori), the feedback mechanism is impaired. In animal models, our laboratory has demonstrated that other types of bacteria besides H. pylori can cause gastritis. For example, under conditions of low acidity, gastritis is secondary to bacterial overgrowth, not production of excessive acid, thus suggesting a new paradigm for the regulation of gastric acid secretion under inflammatory conditions. Cytokines, released during the gastric inflammatory response, including IFN gamma, TNF alpha and IL-1 beta stimulate the G-cell to produce gastrin. Gastrin in turn triggers the release of acid, and hypergastrinemia suppresses somatostatin, the inhibitor of acid. The overall response results in maximal gastric acid output that acts as the stomach's most important anti-microbial agent. The increased acid secretion by the stomach in the presence of H. pylori seems to be part of the innate immune response, in that gastrin and somatostatin are reciprocally regulated by Th1 or Th2 cytokines, respectively. In a mouse model, we showed that octreotide, a somatostatin, analog, is an efficacious treatment for Helicobacter gastritis. In humans, octreotide might accelerate recovery from H. pylori infection, reducing the duration of antibiotic therapy. PMID- 15652229 TI - High effectiveness of platinum(IV) complex with adamantylamine in overcoming resistance to cisplatin and suppressing proliferation of ovarian cancer cells in vitro. AB - [(OC-6-43)-bis(acetato)(1-adamantylamine)amminedichloroplatinum(IV)], coded as LA 12, is an octahedral platinum(IV) complex containing a bulky hydrophobic ligand - adamantylamine. The use of bulky hydrophobic amines as non-leaving ligands, may increase uptake of the compound by the cancer cells. Therefore, the effects of LA 12 on sensitive (A2780) and cisplatin resistant (A2780cis) ovarian cancer cell lines were investigated and compared to those of cisplatin. IC(50) and IC(90) concentrations of LA-12 were 6- (A2780) or 18-fold (A2780cis) lower than those for cisplatin (MTT assay). Equitoxic concentrations (IC(50) or IC(90)) of both compounds caused a significant and similar time- and dose-dependent inhibition of cell proliferation and an increase in the number of floating cells which corresponded to the decrease of total cell viability. A different type and dynamics of cell cycle perturbation after cisplatin and LA-12 treatment were detected. Exposure to LA-12 resulted in transient accumulation of A2780 and A2780cis cells in S phase, while cisplatin caused G(2)/M arrest in sensitive and S phase arrest in resistant cells. A relatively low rate of apoptosis after exposure to IC(50) or IC(90) of both complexes was observed, markedly higher in resistant A2780cis cells. Western blot analysis indicated a concentration dependent p53 level increase in both lines (higher after cisplatin treatment). PARP cleavage was observed only in A2780cis cells. In conclusion, LA-12 was found to be significantly more efficient than cisplatin, and it was able to overcome the acquired cisplatin resistance (showing resistance factor 2.84-fold lower than those for cisplatin). In spite of the low rate of apoptosis, LA-12 caused increase of p53 level and cell cycle perturbations in the ovarian cancer cell lines studied. PMID- 15652230 TI - Inhibition of interleukin-8 (CXCL8/IL-8) responses by repertaxin, a new inhibitor of the chemokine receptors CXCR1 and CXCR2. AB - Repertaxin is a new non-competitive allosteric blocker of interleukin-8 (CXCL8/IL 8) receptors (CXCR1/R2), which by locking CXCR1/R2 in an inactive conformation prevents receptor signaling and human polymorphonuclear leukocyte (PMN) chemotaxis. Given the unique mode of action of repertaxin it was important to examine the ability of repertaxin to inhibit a wide range of biological activities induced by CXCL8 in human leukocytes. Our results show that repertaxin potently and selectively blocked PMN adhesion to fibrinogen and CD11b up regulation induced by CXCL8. Reduction of CXCL8-mediated PMN adhesion by repertaxin was paralleled by inhibition of PMN activation including secondary and tertiary granule release and pro-inflammatory cytokine production, whereas PMN phagocytosis of Escherichia coli bacteria was unaffected. Repertaxin also selectively blocked CXCL8-induced T lymphocyte and natural killer (NK) cell migration. These data suggest that repertaxin is a potent and specific inhibitor of a wide range of CXCL8-mediated activities related to leukocyte recruitment and functional activation in inflammatory sites. PMID- 15652231 TI - Dietary rutin, but not its aglycone quercetin, ameliorates dextran sulfate sodium induced experimental colitis in mice: attenuation of pro-inflammatory gene expression. AB - Oxidative stress has been shown to play a pivotal role in the onset of inflammatory bowel disease (IBD) and carcinogenesis. We evaluated the effects of two dietary anti-oxidants, rutin and its aglycone quercetin, on dextran sulfate sodium (DSS)-induced experimental colitis in mice. Female ICR mice were fed a diet containing 0.1% rutin or 0.1% quercetin for 2 weeks, and given 5% DSS in drinking water during the second week to induce colitis. We also examined the dose-dependency of rutin and quercetin (0.01% and 0.001% each) as well as their therapeutic efficacy, which was evaluated following DSS administration, on DSS induced colitis. The protein level of interleukin (IL)-1 beta in both colonic mucosa and peritoneal macrophages was quantified by enzyme-linked immunosorbent assay. Further, mRNA expression levels of IL-1 beta, tumor necrosis factor-alpha, IL-6, granulocyte macrophage-colony stimulating factor, inducible nitric oxide synthase, and cyclooxygenase (COX)-1 and COX-2 in colonic mucosa were determined by reverse transcription-polymerase chain reaction. A diet containing 0.1% rutin, but not quercetin, attenuated DSS-induced body weight loss and shortening of the colorectum (P<0.01 and <0.05, respectively), and dramatically improved colitis histological scores. Further, DSS-induced increases in colonic mucosal IL-1 beta levels were blunted significantly in rutin-, but not quercetin-, fed mice (P<0.01), while dietary rutin attenuated the expressions of IL-1 beta and IL-6 mRNA in colonic mucosa (each, P<0.01). As for dose dependency, 0.01%, but not 0.001%, dietary rutin significantly reduced mucosal IL-1 beta levels (P<0.01). Notably, a 0.1% rutin diet given 3 days after DSS treatment significantly suppressed both colorectal shortening and IL-1 beta production (P<0.05 and <0.01, respectively). Dietary rutin ameliorates DSS-induced colitis, presumably by suppressing the induction of pro-inflammatory cytokines. Our results suggest that rutin may be useful for the prevention and treatment of IBD and colorectal carcinogenesis via attenuation of pro-inflammatory cytokine production. PMID- 15652232 TI - Antimetastatic effect of prodigiosin through inhibition of tumor invasion. AB - Prodigiosin, a bacterial metabolite, was reported to have immunosuppressive and anticancer activities. In this study, we investigated novel functions of prodigiosin about anti-metastasis and anti-invasion. Prodigiosin dose-dependently inhibited 95-D cells' migration and invasion according to wound healing assay and the Transwell assay. The inhibitive effect could reach about 50% when cells were treated with 5 microM prodigiosin for 12 h. In animal experiment, intraperitoneal administration of 5 mg kg(-1) prodigiosin decreased the number of metastatic nodules by 53% and elevated the survival rate of mice about one-fold comparing with control group. Results of cell aggregation and adhesion assay showed that prodigiosin could promote cell aggregation and simultaneously inhibit cell from adhering to extracellular matrix (ECM). In addition, prodigiosin suppressed RhoA gene expression, hence, decreased protein level of RhoA in 95-D cells, according to RT-PCR assay and Western blot assay. Gel zymogram assay revealed that prodigiosin could suppress the activity of matrix metalloproteinase-2 (MMP-2). These results demonstrate that prodigiosin effectively inhibit tumor metastasis in vitro and in vivo. The action mechanisms of prodigiosin are associated with the promotion of cell aggregation and the inhibition of various steps in cell invasive process, which include the inhibition of cell adhesion and mobility in a RhoA-dependent way and the suppression of MMP-2 ability. PMID- 15652234 TI - Inhibition of interleukin-12 expression in diltiazem-treated dendritic cells through the reduction of nuclear factor-kappa B transcriptional activity. AB - Diltiazem is a calcium channel blocker that suppresses the activation of a variety of immune cells, such as T and B cells, NK cells, monocytes and dendritic cells (DCs). It has been used in the treatment of cardiovascular disorders and has been widely included in clinical protocols to prevent rejection after kidney transplantation. In line with these data, we previously showed that diltiazem directly affects maturation of human DCs and the production of IL-12. Here, we extended our analysis studying the effect of diltiazem on the transcription of IL 12 p35 and p40 subunits focusing on the activity of nuclear factor-kappa B (NF kappa B). A marked reduction of NF-kappa B binding to the kappa B sequences present within the p35 and p40 subunit promoters was observed in diltiazem treated DCs following the stimulation with lipopolysaccharide (LPS) or CD40L. In order to examine the mechanisms by which NF-kappa B binding activity is reduced by diltiazem, we analyzed the NF-kappa B inhibitor, I kappa B alpha. No significant differences were observed in the phosphorylation and/or the degradation of I kappa B alpha. On the other hand, the subcellular distribution of NF-kappa B subunits was clearly affected in diltiazem-treated DCs following LPS stimulation, with a reduced nuclear translocation of p65, and RelB, and a nuclear accumulation of p50 subunit. Thus, all together, our data provided evidence that in addition to the inhibition of p65/p50 nuclear translocation, the selective induction and translocation of p50/p50 homodimers is an important mechanism by which diltiazem inhibits NF-kappa B activity, and in turn, IL-12 expression. PMID- 15652233 TI - Directional trans-epithelial transport of organic anions in porcine LLC-PK1 cells that co-express human OATP1B1 (OATP-C) and MRP2. AB - The transcellular transport of many compounds, which cannot readily cross the lipid bilayer, is mediated by drug uptake and efflux transporters. Human OATP1B1 and MRP2 have been implicated in the hepato-biliary transport of many endogenous and exogenous compounds. Here, we have established epithelial porcine kidney LLC PK1 derived cell lines, that express both transporters in a polarized fashion, as a model to predict hepato-biliary transport. Immunological identification of OATP1B1 in the recombinant cell lines was greatly facilitated by its C-terminal tagging with a peptide sequence derived from hemagglutinin (HA) avoiding the generation of OATP1B1 specific antibodies. Importantly, the tag did not interfere with the functionality of the transporter. Compared to LLC-PK1 cells and cells which expressed only OATP1B1, the cell line that co-expressed MRP2 and OATP1B1 displayed high directional basolateral-to-apical transport of 17 beta-estradiol 17 beta-glucuronide and estrone-3-sulfate. Dehydroepiandrosterone sulfate already displayed a significant basolateral-to-apical transport in the parental cell line, which was further stimulated upon expression of both transporters. Transcellular flux of all steroid conjugates in the opposite direction (apical-to basolateral) was much lower. By employing this cellular model we were able to demonstrate for the first time that OATP1B1 together with MRP2 mediates the trans cellular transport of rifampicin. It is anticipated that the models established herein will greatly facilitate the identification of transporters involved in the disposition of novel drug candidates. PMID- 15652235 TI - Histamine H1 receptor antagonist blocks histamine-induced proinflammatory cytokine production through inhibition of Ca2+-dependent protein kinase C, Raf/MEK/ERK and IKK/I kappa B/NF-kappa B signal cascades. AB - Histamine H1 receptor (H1R), a therapeutic target for alleviation of acute allergic reaction, may be also involved in mediating inflammatory responses via effects on cytokine production. However, the mechanisms whereby histamine induces cytokine production are poorly defined. In this study, we comprehensively investigated the signaling pathway involved in cytokine expression caused by histamine, using native human epidermal keratinocytes. We confirmed the expression of functional H1R by reverse transcription-polymerase chain reaction (RT-PCR), Western blotting and histamine-induced Ca(2+) elevation. Histamine induced concentration- and time-dependent production of granulocyte-macrophage colony stimulating factor (GM-CSF), interleukin (IL)-8 and IL-6, which was completely blocked by olopatadine, an H1 antagonist. Histamine activated the phosphorylation of protein kinase C (PKC), c-Raf, mitogen-activated protein/extracellular signal-regulated kinase kinase (MEK), extracellular signal regulated kinase (ERK), I kappa B kinase (IKK), inhibitory kappa B (I kappa B) alpha and nuclear factor-KB (NF-kappa B) p65, which was inhibited by Ro-31-8220, a PKC inhibitor. Also, Ro-31-8220 significantly suppressed the expression of these cytokines. BAPTA-AM, an intracellular Ca(2+) chelator, also reduced PKC phosphorylation and cytokine expression. PD98059, a MEK inhibitor, and BAY 11 8702, an I kappa B-alpha inhibitor, reduced ERK and NF-kappa B cascade activation, respectively, with little effect on PKC phosphorylation. PD98059 preferentially inhibited GM-CSF production whereas BAY 11-8702 prevented IL-8 and IL-6 production. Furthermore, in addition to the above cytokines, histamine stimulated the biosynthesis and/or release of numerous keratinocyte-derived mediators, which are probably regulated by the ERK or NF-kappa B cascades. Our study suggests that histamine activates Ca(2+)-dependent PKC isoforms that play crucial roles in the activation of Raf/MEK/ERK and IKK/I kappa B/NF-kappa B cascades, leading to up-regulation of cytokine expression. Thus, the anti inflammatory benefit of H1 antagonists may be in part due to prevention of cytokine production. PMID- 15652236 TI - Role of two adjacent cytoplasmic tyrosine residues in MRP1 (ABCC1) transport activity and sensitivity to sulfonylureas. AB - The human ATP-binding cassette (ABC) protein MRP1 causes resistance to many anticancer drugs and is also a primary active transporter of conjugated metabolites and endogenous organic anions, including leukotriene C(4) (LTC(4)) and glutathione (GSH). The sulfonylurea receptors SUR1 and SUR2 are related ABC proteins with the same domain structure as MRP1, but serve as regulators of the K(+) channel Kir6.2. Despite their functional differences, the activity of both SUR1/2 and MRP1 can be blocked by glibenclamide, a sulfonylurea used to treat diabetes. Residues in the cytoplasmic loop connecting transmembrane helices 15 and 16 of the SUR proteins have been implicated as molecular determinants of their sensitivity to glibenclamide and other sulfonylureas. We have now investigated the effect of mutating Tyr(1189) and Tyr(1190) in the comparable region of MRP1 on its transport activity and sulfonylurea sensitivity. Ala and Ser substitutions of Tyr(1189) and Tyr(1190) caused a > or =50% decrease in the ability of MRP1 to transport different organic anions, and a decrease in LTC(4) photolabeling. Kinetic analyses showed the decrease in GSH transport was attributable primarily to a 10-fold increase in K(m). In contrast, mutations of these Tyr residues had no major effect on the catalytic activity of MRP1. Furthermore, the mutant proteins showed no substantial differences in their sensitivity to glibenclamide and tolbutamide. We conclude that MRP1 Tyr(1189) and Tyr(1190), unlike the corresponding residues in SUR1, are not involved in its differential sensitivity to sulfonylureas, but nevertheless, may be involved in the transport activity of MRP1, especially with respect to GSH. PMID- 15652237 TI - Impaired activation of caspases and prevention of mitochondrial dysfunction in the metastatic colon carcinoma CC531s-m2 cell line. AB - In a previous paper we described the properties of a rapidly metastasizing cell line CC531s-m2 derived from the poorly metastasizing CC531s cell. The m2-cell line was relatively resistant to killing by NK cells. Both CD95L and TRAIL mediated apoptosis was decreased in the m2-cell line. Now, by flow cytometrical analysis of intra- and extra-cellular expressed receptors, we show that the localization of the receptors for CD95L and TRAIL was not altered in the CC531s m2 cells as compared to the parental cell line. Subsequently caspase-activation and mitochondrial function were studied by enzymatic cleavage of fluorescent caspase-substrates and retention of the mitochondrial dye rhodamine-123, respectively. The activation of caspases as well as the loss of the mitochondrial membrane potential (MMP) was less in the CC531s-m2 cell line upon CD95L- and TRAIL-signalling. Furthermore, the sensitivity of the CC531-m2 towards cisplatin induced apoptosis was strongly decreased. This was consistent with less mitochondrial damage, delayed caspase cleavage and decreased caspase activity. Altogether, we conclude that an Natural Killer-cell insensitive cell is less sensitive to CD95L- and TRAIL-induced apoptosis as well as anti-cancer drug induced apoptosis by prevention of mitochondrial damage and activation of caspases. PMID- 15652239 TI - 3,5-di-t-butylcatechol (DTCAT) as an activator of rat skeletal muscle ryanodine receptor Ca2+ channel (RyRC). AB - In the present study, the effects of 3,5-di-t-butylcatechol (DTCAT) on ryanodine receptor Ca(2+) channel (RyRC) of skeletal muscle sarcoplasmic reticulum (SR) vesicles were investigated, both by monitoring extravesicular Ca(2+) concentration directly with the Ca(2+) indicator dye arsenazo III and by studying the high-affinity [(3)H]ryanodine binding. DTCAT stimulated Ca(2+) release from junctional (terminal cisternae) vesicles in a concentration-dependent manner, with a threshold activating concentration of 30 microM and a pEC(50) value of 3.43+/-0.03 M. The release of Ca(2+) induced by DTCAT was antagonized in a concentration-dependent manner by ruthenium red, thus indicating that RyRC is involved in the mechanism of stimulation. A structure-activity relationship analysis carried out on a limited number of compounds suggested that both hydroxy and t-butyl groups in DTCAT were important for the activation of RyRC. DTCAT inhibited [(3)H]ryanodine binding to SR vesicles with a K(i) of 232.5 microM, thus indicating that it acted directly at the skeletal muscle ryanodine receptor binding site to stimulate Ca(2+) release. In conclusion, the ability of DTCAT to release Ca(2+) from TC vesicles of skeletal muscle is noteworthy in view of its possible use as an alternative compound to either caffeine or halothane for performing the "In vitro contracture test" to diagnose the susceptibility of some patients to develop malignant hyperthermia under particular pharmacological treatments. PMID- 15652238 TI - Type 4 cAMP phosphodiesterase (PDE4) inhibitors augment glucocorticoid-mediated apoptosis in B cell chronic lymphocytic leukemia (B-CLL) in the absence of exogenous adenylyl cyclase stimulation. AB - cAMP-mediated signaling potentiates glucocorticoid-mediated apoptosis in lymphoid cells, but an effective means by which to take advantage of this observation in the treatment of lymphoid malignancies has not been identified. The primary objective of the current study was to determine whether PDE4 inhibitors, a class of compounds in late clinical development that raise intracellular cAMP levels by inhibiting type 4 cyclic nucleotide phosphodiesterases (PDE4), increase the efficacy of glucocorticoid-mediated apoptosis in leukemic cells from patients with B cell chronic lymphocytic leukemia (B-CLL). Rolipram, a prototypic PDE4 inhibitor, synergized with glucocorticoids in inducing B-CLL but not T cell apoptosis. Rolipram also augmented glucocorticoid receptor element (GRE) transactivation in B-CLL cells. In contrast, inhibition of protein kinase A (PKA) with the cAMP antagonist Rp-8Br-cAMPS reversed both glucocorticoid-induced apoptosis and GRE transactivation. CCRF-CEM cells, a well-studied model of glucocorticoid and cAMP-induced apoptosis, differed from B-CLL cells in that stimulation of adenylyl cyclase with the diterpene forskolin was required to increase both glucocorticoid-mediated apoptosis and GRE activation, while PDE4 inhibition had no effect. Consistent with these results, inhibition of PDE4 induced cAMP elevation in B-CLL but not CCRF-CEM cells, while forskolin augmented cAMP levels in CCRF-CEM but not B-CLL cells. While rolipram treatment up regulated PDE4B in B-CLL, forskolin treatment up-regulated PDE4D in CCRF-CEM cells. These studies suggest that PKA is required for and enhances glucocorticoid induced apoptosis in B-CLL by modulating glucocorticoid receptor signal transduction. Clinical trials that examine whether PDE4 inhibitors enhance the efficacy of glucocorticoid-containing chemotherapy regimens in B-CLL are indicated. PMID- 15652240 TI - Attenuation of cyclosporine A toxicity by sublethal heat shock. Role of catalase. AB - Cyclosporine A (CsA) is the immunosuppressor most frequently used in transplant surgery and in the treatment of autoimmune diseases because of its specific inhibiting effect on signal transduction pathways of cell T receptor. It has been shown that CsA is able to generate reactive oxygen species and lipid peroxidation, which are directly involved in the CsA hepatotoxicity. In the present study, we investigated the effect of a sublethal heat pre-treatment (43 degrees C for 30 min) on the hepatoma cell line HepG2 exposed to cytotoxic concentrations of CsA (10 and 25 microM) for 3 and 24 h. Parameters of cytotoxicity were assayed by measuring LDH (lactate dehydrogenase) leakage into the medium. Peroxide concentration was tested by flow cytometry by measuring the fluorescence intensity of DCF (dichlorofluorescein). Gene expression of catalase was detected by measuring the respective mRNA and proteins, as well as protein level of HSP70. The enzymatic activity of catalase was also determined. Heat pre treatment significantly reduced CsA cytotoxicity as well as the level of peroxide generation. The protective effect of the previous heat treatment (corroborated by the irreversible catalase inhibitor 3-aminotriazole) against the CsA cytotoxicity was due to an increased expression and activity of catalase that was significantly reduced by the effect of CsA. We conclude that heat pre-treatment strongly protects against CsA injury, and the mechanism of this protection is by means of inducing not only the expression of HSP70 but also the expression and activity of catalase, the main enzyme system involved in H(2)O(2) elimination. PMID- 15652242 TI - Inhibition of human CYP2B6 by N,N',N''-triethylenethiophosphoramide is irreversible and mechanism-based. AB - The chemotherapeutic agent N,N',N''-triethylenethiophosphoramide (thioTEPA) is frequently used in high-dose chemotherapy regimens including cyclophosphamide. Previous studies demonstrated partial inhibition by thioTEPA of the cytochrome P4502B6 (CYP2B6)-catalyzed 4-hydroxylation of cyclophosphamide, which is required for its bioactivation. The aim of our study was to investigate the detailed mechanism of CYP2B6 inhibition by thioTEPA. Using human liver microsomes and recombinant P450 enzymes we confirmed potent inhibition of CYP2B6 enzyme activity determined with bupropion as substrate. ThioTEPA was found to inhibit CYP2B6 activity in a time- and concentration-dependent manner. The loss of CYP2B6 activity was NADPH-dependent and could not be restored by extensive dialysis. The maximal rates of inactivation (K(inact)) were 0.16 min(-1) in human liver microsomes and 0.17 min(-1) in membrane preparations expressing recombinant CYP2B6. The half-maximal inactivator concentrations (K(I)) were 3.8 microM in human liver microsomes and 2.2 microM in recombinant CYP2B6. Inhibition was attenuated by the presence of alternative active site ligands but not by nucleophilic trapping agents or reactive oxygen scavengers, further supporting mechanism-based action. Inactivated CYP2B6 did not lose its ability to form a CO reduced complex suggesting a modification of the apoprotein, which is common for sulfur-containing compounds. Pharmacokinetic consequences of irreversible inactivation are more complicated than those of reversible inactivation, because the drug's own metabolism can be affected and drug interactions will not only depend on dose but also on duration and frequency of application. These findings contribute to better understanding of drug interactions with thioTEPA. PMID- 15652241 TI - Some flavonoids and DHEA-S prevent the cis-effect of expanded CTG repeats in a stable PC12 cell transformant. AB - Expanded CUG triplet repeats carrying mRNA seem to be responsible for myotonic dystrophy type 1 (DM1). To study the pathogenesis of DM1, we constructed a DM1 cell culture model using a PC12 neuronal cell line and screened flavonoids that ameliorate this mRNA gain of function. The expanded 250 CTG repeat was subcloned into the 3'-untranslated region of the luciferase gene yielding a stable transformant of PC12 (CTG-250). The cytotoxicity of CTG-250 was evaluated by intracellular LDH activity, and the cis-effect by luciferase activity. To find agents that alter CTG-250 toxic effects, 235 bioflavonoids were screened. An increased cis-effect and cytotoxicity were found when CTG-250 was treated with nerve growth factor to induce differentiation. Western blotting with anti-caspase 3 antibody suggested that cell death was caused by apoptosis. Screening analysis confirmed that a flavone (toringin), an isoflavones (genistein and formononetin), a flavanone (isosakuranetin), and DHEA-S prevent both the cytotoxicity and cis effect of CTG-250 and that a flavanone (naringenin), isoflavone (ononin), and xanthylatin strongly inhibit the cis-effect of CTG repeats. In conclusion, we found that this neuronal cell line, which expresses the CUG repeat-bearing mRNA, showed cis-effects through the reporter gene and neuronal death after cell differentiation in vitro. However, some flavonoids and DHEA-S inhibit both the cis-effect and cytotoxicity, indicating that their chemical structures work to ameliorate both these toxic effects. This system makes it easy to evaluate the toxic effects of expanded CTG repeats and therefore should be useful for screening other DM1 treatments for their efficacies. PMID- 15652243 TI - Identification and functional analysis of two rare allelic variants of the thiopurine S-methyltransferase gene, TPMT*16 and TPMT*19. AB - Human thiopurine S-methyltransferase (TPMT) catalyses the S-methylation of thiopurine drugs. TPMT is genetically polymorphic and is associated with large interindividual variations in thiopurine drug toxicity and therapeutic efficacy. During routine genotyping of patients with Crohn's disease, one novel missense mutation, 365A>C (TPMT*19, Lys(122)Thr), and a recently described missense mutation, 488G>A (TPMT*16, Arg(163)His), were identified in a Caucasian and a Moroccan patient, respectively. Using a heterologous yeast expression system, kinetic parameters (K(m) and V(max)) of the two variants with respect to 6 thioguanine S-methylation were determined and compared with those obtained with the wild-type enzyme. The Lys(122)Thr exchange did not significantly decrease the intrinsic clearance value (V(max)/K(m)) of the variant enzyme. In contrast, the Arg(163)His substitution significantly decreased the intrinsic clearance value by three-fold. The Arg(163) is located in a highly conserved region of the human TPMT protein and, as such, the Arg(163)His substitution is expected to result in a marked reduction of enzyme activity, as confirmed by the in vitro data. Phenotyping by measurement of red blood cell TPMT activity indicated that the patient heterozygous for the Lys(122)Thr mutation had normal TPMT activity, whereas the patient heterozygous for the Arg(163)His mutation was an intermediate methylator, which demonstrated a positive correlation between TPMT phenotyping and the in vitro data. The identification of a novel non-functional allele of the TPMT gene improves our knowledge of the genetic basis of interindividual variability in TPMT activity. These data further enhance the efficiency of genotyping methods to predict patients at risk of an inadequate response to thiopurine therapy. PMID- 15652244 TI - Mechanisms involved in spironolactone-induced choleresis in the rat. Role of multidrug resistance-associated protein 2. AB - The mechanisms involved in spironolactone (SL, 200 micromol/kg body weight, 3 days i.p.)-induced choleresis were explored in vivo by evaluating bile salt export pump (Bsep)-, multidrug resistance-associated protein 2 (Mrp2)-, and anion exchanger 2 (AE2)-mediated secretory processes in rat liver. Hepatic bile salt metabolism was also analyzed. Total bile flow was significantly increased by SL, primarily due to an increase in bile salt-independent bile flow, whereas bile salt secretion was decreased. SL did not produce any choleresis in TR(-) rats. SL decreased the de novo bile salt synthesis rate in concordance with impaired microsomal cholesterol 7 alpha-hydroxylase activity, thus leading to a decrease in endogenous bile salt pool size. In contrast, the maximum secretory rate of tauroursodeoxycholate as well as expression of Bsep protein detected by Western blotting were not affected. Thus, decreased bile salt availability for canalicular transport rather than transport capability itself likely explains reduced biliary secretion of bile salts. Biliary secretion of glutathione, an endogenous substrate of Mrp2, and HCO(3)(-), the AE2 substrate, were increased by SL, as a main factor explaining enhanced bile salt-independent bile flow. Western blot studies revealed increased expression of Mrp2 in response to SL whereas AE2 content remained unchanged. Enhanced activity and expression of Mrp2 was confirmed by analyzing the excretion rate of dinitrophenyl S-glutathione, an exogenous substrate of Mrp2, in isolated hepatocytes and by immunofluorescence microscopy, respectively. We conclude that SL increased bile flow mainly by increasing the biliary secretion of glutathione species and HCO(3)(-); increased expression of Mrp2 is also involved. PMID- 15652245 TI - The developing art of protein crystallisation: new advances from improved knowledge automation and miniaturisation. PMID- 15652246 TI - Protein crystallization: virtual screening and optimization. AB - Advances in genomics have yielded entire genetic sequences for a variety of prokaryotic and eukaryotic organisms. This accumulating information has escalated the demands for three-dimensional protein structure determinations. As a result, high-throughput structural genomics has become a major international research focus. This effort has already led to several significant improvements in X-ray crystallographic and nuclear magnetic resonance methodologies. Crystallography is currently the major contributor to three-dimensional protein structure information. However, the production of soluble, purified protein and diffraction quality crystals are clearly the major roadblocks preventing the realization of high-throughput structure determination. This paper discusses a novel approach that may improve the efficiency and success rate for protein crystallization. An automated nanodispensing system is used to rapidly prepare crystallization conditions using minimal sample. Proteins are subjected to an incomplete factorial screen (balanced parameter screen), thereby efficiently searching the entire "crystallization space" for suitable conditions. The screen conditions and scored experimental results are subsequently analyzed using a neural network algorithm to predict new conditions likely to yield improved crystals. Results based on a small number of proteins suggest that the combination of a balanced incomplete factorial screen and neural network analysis may provide an efficient method for producing diffraction-quality protein crystals. PMID- 15652247 TI - Robotic nanolitre protein crystallisation at the MRC Laboratory of Molecular Biology. AB - We have set up high-throughput robotic systems to screen and optimise crystallisation conditions of biological macromolecules with the aim to make difficult structural biology projects easier. The initial screening involves two robots. A Tecan Genesis liquid handler is used to transfer commercially available crystallisation reagents from 15 ml test tubes into the reservoirs of 96-well crystallisation plates. This step is fully automated and includes a carousel for intermediate plate storage, a Beckman plate sealer and a robotic arm, which transfers plates in between steps. For adding the sample, we use a second robot, a 17-tip Cartesian Technologies PixSys 4200 SynQuad liquid handler, which uses a syringe/solenoid valve combination to dispense small quantities of liquid (typically 100 nl) without touching the surface of the plate. Sixteen of the tips are used to transfer the reservoir solution to the crystallisation wells, while the 17th tip is used to dispense the protein. The screening of our standard set of 1440 conditions takes about 3 h and requires 300 microl of protein solution. Once crystallisation conditions have been found, they are optimised using a second Tecan Genesis liquid handler, which is programmed to pipette gradients from four different corner solutions into a wide range of crystallisation plate formats. For 96-well plates, the Cartesian robot can be used to add the sample. The methods described are now used almost exclusively for obtaining diffraction quality crystals in our laboratory with a throughput of several thousand plates per year. Our set-up has been copied in many institutions worldwide. PMID- 15652248 TI - Methods for separating nucleation and growth in protein crystallisation. AB - The availability of high-quality crystals is crucial to the structure determination of proteins by X-ray diffraction. With the advent of structural genomics the pressure to produce crystals is greater than ever before. Finding favourable conditions for crystallisation is usually achieved by screening of the protein solution with numerous crystallising agents. Optimisation of the crystallisation conditions involves the manipulation of the crystallisation phase diagram with the aim of leading crystal growth in the direction that will produce the desired results. This article highlights recent advances in experimental methods for improving crystal size and quality by separating the nucleation and growth phases of crystallisation using the vapour diffusion and microbatch techniques. PMID- 15652250 TI - Life in the fast lane for protein crystallization and X-ray crystallography. AB - The common goal for structural genomic centers and consortiums is to decipher as quickly as possible the three-dimensional structures for a multitude of recombinant proteins derived from known genomic sequences. Since X-ray crystallography is the foremost method to acquire atomic resolution for macromolecules, the limiting step is obtaining protein crystals that can be useful of structure determination. High-throughput methods have been developed in recent years to clone, express, purify, crystallize and determine the three dimensional structure of a protein gene product rapidly using automated devices, commercialized kits and consolidated protocols. However, the average number of protein structures obtained for most structural genomic groups has been very low compared to the total number of proteins purified. As more entire genomic sequences are obtained for different organisms from the three kingdoms of life, only the proteins that can be crystallized and whose structures can be obtained easily are studied. Consequently, an astonishing number of genomic proteins remain unexamined. In the era of high-throughput processes, traditional methods in molecular biology, protein chemistry and crystallization are eclipsed by automation and pipeline practices. The necessity for high-rate production of protein crystals and structures has prevented the usage of more intellectual strategies and creative approaches in experimental executions. Fundamental principles and personal experiences in protein chemistry and crystallization are minimally exploited only to obtain "low-hanging fruit" protein structures. We review the practical aspects of today's high-throughput manipulations and discuss the challenges in fast pace protein crystallization and tools for crystallography. Structural genomic pipelines can be improved with information gained from low-throughput tactics that may help us reach the higher-bearing fruits. Examples of recent developments in this area are reported from the efforts of the Southeast Collaboratory for Structural Genomics (SECSG). PMID- 15652249 TI - Membrane protein crystallization in amphiphile phases: practical and theoretical considerations. AB - Integral membrane proteins are amphiphilic molecules. In order to enable chromatographic purification and crystallization, a complementary amphiphilic microenvironment must be created and maintained. Various types of amphiphilic phases have been employed in crystallizations and intricate amphiphilic microenvironmental structures have resulted from these and are found inside membrane protein crystals. In this review the process of crystallization is put into the context of amphiphile phase transitions. Finally, practical factors are considered and a pragmatic way is suggested to pursue membrane protein crystallization trials. PMID- 15652251 TI - Individual differences in behavior and physiology: causes and consequences. PMID- 15652252 TI - The Darwinian concept of stress: benefits of allostasis and costs of allostatic load and the trade-offs in health and disease. AB - Why do we get the stress-related diseases we do? Why do some people have flare ups of autoimmune disease, whereas others suffer from melancholic depression during a stressful period in their life? In the present review possible explanations will be given by using different levels of analysis. First, we explain in evolutionary terms why different organisms adopt different behavioral strategies to cope with stress. It has become clear that natural selection maintains a balance of different traits preserving genes for high aggression (Hawks) and low aggression (Doves) within a population. The existence of these personality types (Hawks-Doves) is widespread in the animal kingdom, not only between males and females but also within the same gender across species. Second, proximate (causal) explanations are given for the different stress responses and how they work. Hawks and Doves differ in underlying physiology and these differences are associated with their respective behavioral strategies; for example, bold Hawks preferentially adopt the fight-flight response when establishing a new territory or defending an existing territory, while cautious Doves show the freeze-hide response to adapt to threats in their environment. Thus, adaptive processes that actively maintain stability through change (allostasis) depend on the personality type and the associated stress responses. Third, we describe how the expression of the various stress responses can result in specific benefits to the organism. Fourth, we discuss how the benefits of allostasis and the costs of adaptation (allostatic load) lead to different trade offs in health and disease, thereby reinforcing a Darwinian concept of stress. Collectively, this provides some explanation of why individuals may differ in their vulnerability to different stress-related diseases and how this relates to the range of personality types, especially aggressive Hawks and non-aggressive Doves in a population. A conceptual framework is presented showing that Hawks, due to inefficient management of mediators of allostasis, are more likely to be violent, to develop impulse control disorders, hypertension, cardiac arrhythmias, sudden death, atypical depression, chronic fatigue states and inflammation. In contrast, Doves, due to the greater release of mediators of allostasis (surplus), are more susceptible to anxiety disorders, metabolic syndromes, melancholic depression, psychotic states and infection. PMID- 15652253 TI - Prenatal origins of individual variation in behavior and immunity. AB - The in utero environment plays a critical role in initiating the normal ontogeny of many physiological systems. As a consequence, disturbances during prenatal life can affect the baby's maturational trajectory and sometimes cause chronic alterations that influence health postpartum. Our review summarizes a series of studies in rhesus monkeys supporting these conclusions. Psychological disturbance or pharmacological stimulation of the gravid female's pituitary-adrenal axis affected the infant's neurological development: monkeys evinced immature neuromotor reflexes at birth, greater emotionality during the first year of life, and a smaller hippocampus as juveniles. Immune responses of the infants were also affected: lymphocyte proliferation, natural killer activity and cytokine production were reduced. Several mediating pathways were implicated, including the placental transfer of hormones and nutrients, and a differential response of the infant monkey to the rearing environment. For example, the establishment of beneficial types of microflora in the gastrointestinal tract was significantly reduced, which was associated with a greater risk for enteric infection. These findings indicate that events during fetal life can persistently influence physiology after birth and tilt the balance away from health and toward illness. PMID- 15652254 TI - Neurobiological characteristics of rhesus macaque abusive mothers and their relation to social and maternal behavior. AB - Previous studies have reported hyperactivation of catecholaminergic systems and elevated concentrations of corticotropin-releasing-hormone (CRH) in the cerebrospinal fluid (CSF) of child maltreatment victims or combat veterans with post-traumatic stress disorder (PTSD). This study investigated the CSF concentrations of CRH and monoamine metabolites in rhesus macaque mothers that physically abused their infants and had themselves been abused as infants. Ten abusive mothers and 10 controls served as study subjects. All animals were sampled for CSF during pregnancy and the postpartum period. Focal observations of social and maternal behavior were also made. Abusive mothers had significantly higher CSF concentrations of CRH and 5-HIAA than controls. Across both subjects and controls, higher CRH, 5-HIAA and MHPG concentrations were associated with anti-social behavior patterns including a high frequency of maternal aggression, infant rejection, and a low frequency of contacts received from other individuals. These findings are consistent with those of previous primate and human studies and suggest that the neurobiological alterations associated with infant abuse may play an important role in the occurrence of maladaptive behavior in adulthood, including the perpetuation of infant abuse across generations. PMID- 15652255 TI - Individual differences in cardiovascular response to social challenge. AB - An important determinant of cardiovascular stress reactivity and morbidity is the individual behavioral strategy of coping with social challenge. This review summarizes the results of a number of studies that we performed in rats, aimed at investigating the relationship between aggression and cardiovascular responsivity under social stress conditions. We show that rats belonging to the 'aggressive tail' of a population are characterized by a higher sympathetic-adrenomedullary activation during social and non-social stress episodes. Wild-type rats are characterized by a larger sympathetic dominance and a higher susceptibility to cardiac arrhythmias during defeat as compared to Wistars. Cardiovascular habituation takes place when social challenge is an intermittent victory experience, whereas no habituation is observed across repeated defeat episodes. Dominant rats whose social dominance is challenged by the aggression of another subject display long-term alterations of heart rate circadian rhythmicity. Such changes are linked to individual proness to defend social dominance: the more the animal counterattacks the aggressor, the smaller the subsequent rhythm disturbance. These data underline how important it is to carefully consider individual differences in aggression and the context in which aggression is expressed, when studying cardiovascular effects of social interactions. PMID- 15652256 TI - Social factors and individual vulnerability to chronic stress exposure. AB - The stress-response is adaptive in the short-term, but it can be maladaptive if sustained levels of its mediators are chronically maintained. Furthermore, not all individuals exposed to chronic stress will progress to disease. Thus, understanding the causes of individual differences and the consequences of variation in vulnerability is of major importance. The aim of this review is to shed light on this issue by presenting a new naturalistic model of chronic psychosocial stress in male mice. Resident/intruder pairs of mice lived in continuous sensory contact and physically interacted daily. Four categories were identified: Resident Dominant, Resident Subordinate (RS), Intruder Dominant, and Intruder Subordinate. Behavior, autonomic and immune functions, hypothalamic pituitary-adrenocortical responses, brain cytokine expression and cardiac histology were investigated in stress-exposed mice. Certain stress-induced alterations were present in all mice independent of their social status, while others clearly differentiated dominants from subordinates. RS mice showed a unique profile of alterations suggesting that the loss of relevant resources, such as the territory, is the key factor determining why only certain stress exposed individuals ultimately show malignancy and psychopathologies. PMID- 15652257 TI - Long-term effects of social stress on brain and behavior: a focus on hippocampal functioning. AB - In order to study mechanisms involved in the etiology of human affective disorders, there is an abundant use of various animal models. Next to genetic factors that predispose for psychopathologies, environmental stress is playing an important role in the etiology of these mental diseases. Since the majority of stress stimuli in humans that lead to psychopathology are of social nature, the study of consequences of social stress in experimental animal models is very valuable. The present review focuses on one of these models that uses the resident-intruder paradigm. In particular the long-lasting effects of social defeat in rats will be evaluated. Data from our laboratory on the consequences of social defeat on emotional behavior, stress responsivity and serotonergic functionality are presented. Furthermore, we will go into detail on hippocampal functioning in socially stressed rats. Very recent results show that there is a differential effect of a brief double social defeat and repetitive social defeat stress on dendritic remodeling in hippocampal CA3 neurons and that this has repercussions on hippocampal LTP and LTD. Both the structural and electrophysiological changes of principal neurons in the hippocampal formation after defeat are discussed as to their relationship with the maintenance in cognitive performance that was observed in socially stressed rats. The results are indicative of a large dynamic range in the adaptive plasticity of the brain, allowing the animals to adapt behaviorally to the previously occurred stressful situation with the progression of time. PMID- 15652258 TI - Inter-individual vs line/strain differences in psychogenetically selected Roman High-(RHA) and Low-(RLA) Avoidance rats: neuroendocrine and behavioural aspects. AB - Inter-individual differences in neuroendocrine and behavioural responses to environmental challenges will be considered within the context of psychogenetic selection, using the Roman High-(RHA) and Low-(RLA) Avoidance rat lines as an example. We assume that the selected genotypes, by interacting with environmental factors, determine specific 'biobehavioural profiles'. Practical and theoretical problems regarding the measurement of inter-individual vs line/strain differences, the definition of 'traits' vs experimental variables, and possible correlations between physiological and behavioural parameters will be discussed. We will argue that environmental influences are the main cause of inter individual variability, and that the genotype only constitutes a 'blueprint' from which typical biobehavioural profiles are established, notably under the influence of early environmental factors. These biobehavioural profiles may correspond in part to human categories known as 'types', 'temperaments' or 'personality traits'. Within each category (including those which can be obtained by psychogenetic selection), more individual personality traits can evolve, notably as a result of social interactions and particular life events. PMID- 15652259 TI - Behavioural, neural and cardiovascular adaptations in mice lacking the NPY Y1 receptor. AB - Neuropeptide Y (NPY) is primarily synthesised and released by neurones, it is co localised with noradrenaline and is involved in the regulation of cardiovascular function. In a mouse model lacking NPY Y1 receptor (KO), the ability of NPY to potentiate noradrenaline-induced vasoconstriction is abolished during stress but normal in baseline conditions, locomotor activity and metabolic rate are lowered, blood insulin levels and glucose storage activity are increased. The present study was aimed at further characterising NPY Y1 mutants, with special emphasis on: behavioural responses to novelty seeking and open-field with objects tests, heart rate responsiveness during acute social defeat, alpha2-adrenoceptor (alpha2 ARs) function in brain areas involved in cardiovascular regulation, and cardiac structure. As compared to wild-type controls (n=9), NPY Y1 KOs (n=9) showed: reduced somatomotor activation during non-social challenges, lower heart rate in baseline conditions, larger heart rate responsiveness during social defeat, increased number of alpha2-ARs in the dorsal motor nucleus of the vagus (nX) and the locus coeruleus (LC), moderately larger volume fraction of myocardial fibrosis. The remarkable increment of alpha2-adrenoceptor density in the nX and LC allows to view KO mice behavioural and anatomo-physiological peripheral characteristics as 'adaptations' to central adrenergic rearrangement due to NPY Y1 receptor deletion. PMID- 15652260 TI - Individual variation by self-organisation. AB - In this paper, we show that differences in dominance and spatial centrality of individuals in a group may arise through self-organisation. Our instrument is a model, called DomWorld, that represents two traits that are often found in animals, namely grouping and competing. In this model individual differences grow under the following conditions: (1) when the intensity of aggression increases and grouping becomes denser, (2) when the degree of sexual dimorphism in fighting power increases. In this case the differences among females compared to males grow too, (3) when, upon encountering another individual, the tendency to attack is 'obligate' and not conditional, namely 'sensitive to risks'. Results resemble phenomena described for societies of primates, mice, birds and pigs. PMID- 15652261 TI - Avian personalities: characterization and epigenesis. AB - The work presented here aims at understanding the nature, epigenesis and function of personality types (here called behavioral profiles) in birds, focusing on a wild bird species, the great tit (Parus major). Lines bidirectionally selected for exploration show a wide array of social and non-social behavioral differences, and also some differences in physiological parameters. Line differences in these characteristics and their relationships show significant temporal consistency. The results show a surprising similarity between the great tit and a rodent model, suggesting a fundamental principle in the organization of behavioral profiles. The nature of this principle and whether or not it is multi dimensional is discussed. However, the similarity with a chicken model is less clear, which points to some caution for generalization. The epigenesis of great tit behavioral profiles is discussed. Selection experiments with replication and backcrosses reveal a strong genetic basis, and suggest an influence of maternal effects. Ontogenetic manipulations indicate strong developmental plasticity, suggesting adaptive adjustment to prevailing environmental circumstances. They also show that behavioral characteristics belonging to the same profile can become uncoupled. Finally, field data on several fitness parameters of the different personalities in wild great tits are summarized. These data suggest that variation in selection pressure in time and space and assortative mating are plausible mechanisms accounting for the maintenance of different behavioral profiles within the same population. PMID- 15652263 TI - The concept of alternative strategies and its relevance to psychiatry and clinical psychology. AB - The intent of this article is to introduce the evolutionary concept of alternative strategies into the fields of psychiatry and clinical psychology. In behavioral ecology, the term alternative strategies refers to the presence of two or more discrete behavioral variants among adults of one sex and one population when those variants serve the same functional end. Often discrete behavioral variants are associated with specific morphological, physiological, and life history characters. The concept of alternative strategies has been applied to human behavior to explain the origin of some behavioral syndromes that are currently classified as mental disorders or emotional dysfunctions. Antisocial personality could represent a high-risk strategy of social defection associated with resource acquisition and reproduction. Insecure attachment could represent an evolved psychological mechanism that used the quality of parental care received during childhood as a cue for optimizing adult reproductive strategies. Since a major contribution of evolutionary theory is the insight that individual differences are core biological features of any animal species, including Homo sapiens, the application of the concept of alternative strategies to psychiatry and clinical psychology can be a powerful antidote to the growing tendency to medicalize human diversity. PMID- 15652262 TI - Bold attitude makes male urban feral domestic cats more vulnerable to Feline Immunodeficiency Virus. AB - Individual differences in behaviour are a phenomenon that is more and more attracting the attention of scientists. Among the other reasons, behavioural individuality occurs because selection favours the adoption of different tactics by individuals. It is now widely recognized that within many vertebrate species, individuals vary along an axis the extremes of which are represented by individuals 'bold' and 'shy', sometimes called 'proactive' and 'reactive'. Here we present the case of feral domestic cats (Felis catus L.) living in group in the urban environment where showing bold attitudes is linked to the benefit of a high annual reproductive success but, on the other hand, to a high probability to be infected by the Feline Immunodeficiency Virus (FIV), a lethal disease caused by a retrovirus. In this species, natural selection has probably favoured proactive temperament in spite of the cost represented by getting the disease. In fact, proactive individuals, even if FIV positive, reproduce more than reactive individuals before the last stage of FIV-infection (AIDS) characterized by a loss of immunological defences and subsequent opportunistic infections. Evolutionary implications are discussed. PMID- 15652264 TI - Anxiety and panic: from human studies to animal research and back. AB - The role of learning and conditioning varies across human anxiety disorders, and distinguishing between fear and panic is important to guide investigation in panic disorder. By reminding that some psychological and psychobiological theories view panic attacks as false alarms of unconditioned biological origin, we suggest that employing endophenotypes of biological and evolutionary relevance -such as the respiratory responses to suffocative stimuli--can be fruitful for both human research and animal models of panic, and can help keeping unconditioned components of the clinical picture separate from the conditioned components in the experimental setting. We present a review of a model of panic disorder by which idiosyncratic environmental adverse events can promote unconditioned and unexpected spells of physical alarm. Along the proposed causal pathway the alternative splicing expression of polymorphic genes of the cholinergic system play an important role. The overproduction of the Acetylcholinesterase readthrough splice variant after minor stress can promote passive avoidance and learning through action at the level of the corticolimbic circuitries, as well as heightened sensitivity to suffocative stimuli by action upon the cholinergic components of chemoception. When a component of anticipatory anxiety complicates the clinical picture of recurrent panic attacks, and the HPA becomes activated, the glucocorticoid response element 17 kb upstream of the Acetylcholinesterase gene transcription initiation site may sustain sensitivity to suffocative stimuli for prolonged time. Finally, we review how animal models of human panic based on unconditioned provocation of alarm reactions by the same respiratory panicogens that are employed in man are viable and promising. PMID- 15652265 TI - Psychological intimate partner violence: the major predictor of posttraumatic stress disorder in abused women. AB - Intimate partner violence (IPV) significantly impacts women mental and physical wellbeing and therefore represents a worldwide public health problem. A clear association between IPV and increased risk to develop posttraumatic stress disorder (PTSD) has been documented. However, few studies examined how different features of IPV (physical, psychological, sexual) interact with other traumatic stress experiences (physical, psychological and sexual childhood abuse and adulthood victimization by other/s than the partner) in determining PTSD. Women abused by the partner (n=75) were compared with non-abused control women (n=52). Information about sociodemographic profile and relevant personal characteristics was obtained through structured interviews. A comprehensive questionnaire was designed for a face-to-face interview in order to obtain detailed information about duration and frequency of the different types of violent acts above mentioned. The incidence and severity of symptoms of current PTSD were assessed with Echeburua's Severity of Symptom Scale of Posttraumatic Stress Disorder, a structured interview based on DSM-IV criteria. Women suffering from IPV had a significantly higher rate of PTSD symptomatology as compared to control women, whereas childhood abuse variables did not explain PTSD score variance. In addition, the severity of IPV was significantly and positively correlated with the intensity of PTSD symptoms. Women involved in an abusive relationship were more frequently exposed to other experiences of adulthood victimization, suggesting that their higher PTSD vulnerability could be a result of cumulative traumatic experiences. A relevant result of the correlation analysis was the strong, positive association between PTSD and each different type of IPV. In particular, the psychological component of intimate partner violence was the strongest predictor of posttraumatic stress disorder. This study underlines the importance of separating the effects of the different types of intimate partner abuse when taking into account its effects on women mental health. PMID- 15652266 TI - Coping with competitive situations in humans. AB - The analysis of effects of competitive situations in our species may contribute to acquiring deeper knowledge about the effects of social stress and its relationship with different pathologies. The latest studies indicate that the neuroendocrine response to competition depends more on subjective factors related to the cognitive evaluation of the situation than on the outcome itself. Findings suggest that when subjects cope with a competition, they assess it in such a way that it activates a psychobiological coping response. The pattern of this response may correspond to a predominant active or passive coping strategy, the choice ultimately depending on factors such as the importance of the competition for the subject, the subject's involvement or perceived possibilities of control of outcome or success (e.g. past experience in similar competitions, judge or rank of the opponent), among others. More important than winning or losing is the coping pattern displayed by the subject, which determines the hormonal changes experienced when facing competition and its outcome. PMID- 15652268 TI - The role of TGF-beta 1 and cytokines in the modulation of liver fibrosis by Sho saiko-to in rat's bile duct ligated model. AB - Liver fibrosis is an over-accumulation of extra-cellular matrix (ECM) and the hepatic stellate cell (Ito cell) play a central role in the pathogenesis of liver fibrosis. There are a lot of growth factors and cytokines involved in the activation of hepatic stellate cell, including of transforming growth factor (TGF alpha, TGF-beta1), platelet-derived growth factor (PDGF), interleukin (IL 1alpha,beta, IL-6) and tumor necrosis factor (TNF-alpha). Sho-saiko-to (TJ-9; Xiao-Chai-Hu-Tang in Chinese) was the most popular herbal medicine for the treatment of chronic liver disease in Chinese and Japanese. Our aim of the current study was to examine whether TJ-9 regulated the growth factors and cytokines in the fibrogenesis of bile duct ligated model. Therefore, we assessed the TJ-9's potential in regulating TGF-beta1, PDGF mRNA expression, the amount of IL-1alpha, IL-1beta, IL-6, TNF-alpha and the fibrotic marker "PIII NP" in the serum. Then, using the immunohistochemical stain to observe the TGF-beta1 expression in the tissue. Our results showed that TJ-9 at a dose of 0.5 g/(kgday) significantly reduced the serum level of PIII NP, the mRNA expression of TGF beta1 and PDGF. For the cytokines involved in the activation of Ito cell, TJ-9 at a dose of 0.5 g/(kgday) significantly suppressed the increasing tendency of IL 1beta and enhanced the production of TNF-alpha. Finally, we concluded that: (1) TJ-9 at a dose of 0.5g/(kgday) significantly reduced the serum fibrotic marker PIII NP in the bile duct ligated model, and its mechanism was partly by means of downregulating the mRNA of TGF-beta1 and PDGF. These results also confirmed by the immunohistochemical staining of TGF-beta1. (2) TJ-9 at a dose of 0.5 g/(kgday) suppressed the increasing tendency of IL-1beta and stimulated the production of TNF-alpha to inhibit Ito cell proliferation and collagen formation. PMID- 15652267 TI - Flavonoids and antiulcerogenic activity from Byrsonima crassa leaves extracts. AB - Byrsonima crassa Niedenzu (IK) (Malpighiaceae) is used in Brazilian folk medicine for the treatment of diseases related mainly to gastric ulcers. In this study, we evaluated the potential antiulcerogenic effect of three different extracts obtained from the leaves of Byrsonima crassa namely hydromethanolic (80% MeOH), methanolic (MeOH) and chloroformic extracts (CHCl(3)). The oral administration (250, 500 and 1000 mg/kg) of all the extracts reduced the formation of lesions associated with HCl/ethanol administration in mice. The 80% MeOH extract significantly reduced the incidence of gastric lesions by 74, 78 and 92% at doses of 250, 500 and 1000 mg/kg, respectively (P<0.01). The MeOH extract reduced the ulceration by 93 and 99% only at the doses of 500 and 1000 mg/kg (P<0.01). The lower gastroprotective action (69%) was observed when animals were treated with CHCl(3) extract at the dose of 1000 mg/kg (P<0.01). Phytochemical investigation of Byrsonima crassa afforded five known substances: quercetin-3-O-beta-d galactopyranoside, quercetin-3-O-alpha-l-arabinopyranoside, the biflavonoid amentoflavone, (+)-catechin and (-)-epicatechin. The presence of these phenolic compounds may probably explain the antiulcerogenic effect of the extracts of Byrsonima crassa leaves. PMID- 15652269 TI - The in vitro cytotoxic and apoptotic activity of Triphala--an Indian herbal drug. AB - A study on cytotoxic effect of acetone extract of "Triphala" whose antimutagenicity has already been tested. The in vitro antimutagenic activity of Triphala--an Indian herbal drug. Food Chemistry and Toxicology 40, 47-54) was extended to test its cytotoxic effects on cancer cell-lines using Shionogi 115 (S115) and MCF-7 breast cancer cells and PC-3 and DU-145 prostate cancer cells as models. The results revealed that acetone extract of "Triphala" showed a significant cytotoxic effect on these cancer cell-lines and the effect was similar on all cancer cell lines used in this study. The major phenolic compounds in the most potent acetone extracts were isolated and purified. Structural analysis was conducted using spectroscopic techniques including mass spectroscopy, nuclear magnetic resonance (NMR) and infrared (IR) which showed gallic acid as the major component. The suppression of the growth of cancer cells in cytotoxic assays may be due to the gallic acid-a major polyphenol observed in "Triphala". PMID- 15652270 TI - Reactive oxygen species mediation of baizhu-induced apoptosis in human leukemia cells. AB - Baizhu (Atractylodes macrocephala Koidz) has traditionally been used as an important ingredient of several Chinese herbal medicines, which have been used for abdominal pain and gastroenterology diseases for thousands of years. Despite its popularity in herbal therapies, little is known about the anticancer effect of Baizhu. In this study, the anticancer potential of Baizhu on human hepatoma and leukemia cell lines was evaluated. Baizhu methanol extract induced apoptosis in human lymphoma Jurkat T cells, leukemia U937, and HL-60 cells. This was confirmed by several methods, including hypodiploid cells detection using flow cytometry, the examination of apoptotic bodies containing cells using confocal laser scanning microscopy, and hypodiploid cell population inhibition using the broad spectrum caspase inhibitor z-VAD. Finally, the intracellular reactive oxygen species (ROS), especially hydrogen peroxide (H(2)O(2)) and superoxide anion (O(2)(-)), were found to be elevated after treatment of these cells with Baizhu extracts. Antioxidant N-acetyl cysteine (NAC) pretreatment almost completely inhibited Baizhu-induced apoptosis, suggesting that ROS are the key mediators for Baizhu-induced apoptosis. All these data indicate that Baizhu is a possible anti-tumor agent that induces apoptosis of human leukemia cells through ROS generation. PMID- 15652272 TI - Antioxidant activity of Cassia fistula (Linn.) flowers in alloxan induced diabetic rats. AB - Aqueous extract of Cassia fistula (Linn.) flowers (ACF) was screened for its antioxidant effect in alloxan induced diabetic rats. An appreciable decrease in peroxidation products viz thiobarbituric acid reactive substances, conjugated dienes, hydroperoxides was observed in heart tissues of ACF treated diabetic rats. The decreased activities of key antioxidant enzymes such as superoxide dismutase, catalase, glutathione peroxidase, glutathione reductase and glutathione in diabetic rats were brought back to near normal range upon ACF treatment. These results suggest that ACF has got promising antioxidative activity in alloxan diabetic rats. PMID- 15652271 TI - The antidiabetic activity of the herbal preparation ADD-199 in mice: a comparative study with two oral hypoglycaemic drugs. AB - The antidiabetic and antioxidant effects of the herbal preparation ADD-199 were investigated in STZ-induced diabetic C(3)H mice and results were compared with two allopathic hypoglycaemic drugs, glibenclamide and metformin. Plasma glucose, insulin and lipids as well as liver glycogen, lipids and lipid peroxidation were measured following treatment for 8 weeks. The results indicated that plasma insulin levels in normal controls at termination were about 76 micromol/L compared to trace levels in untreated diabetic mice. Glibenclamide and ADD-199 increased insulin levels in diabetic mice up to 70% of levels in untreated non diabetic mice whilst metformin had no effect. Basal plasma glucose levels in diabetic controls (18.8 mM) were reduced to 14.0 mM by 100 mg/kg ADD-199 in <2 weeks compared to 4 and 6 weeks for glibenclamide and metformin, respectively. This hypoglycaemic effect of ADD-199 appeared to be associated with the alkaloidal content of the extract. Treatment with ADD-199 or the hypoglycaemic agents reversed the observed elevation in plasma lipids but increased hepatic glycogen, triacylglycerol and cholesterol levels. Treatment also increased glucose uptake by isolated diaphragms and attenuated hepatic lipid peroxidation. These antihyperglycaemic and antioxidant actions of ADD-199 at a dose of 100mg/kg/day are comparable to those of the maximum daily therapeutic doses of glibenclamide (0.25 mg/kg) and metformin (50 mg/kg). These could explain the basis for use of this plant extract to manage diabetes mellitus (DM). PMID- 15652273 TI - Some pharmacological properties of extracts of Terminalia sericea roots. AB - Terminalia sericea Burch. Ex. DC (Combretaceae) extracts are used to treat bacterial infections, diarrhea, and diabetes. Intermediate and polar extracts of the roots exhibited antibacterial activity against Staphylococcus aureus, Escherichia coli, Bacillus anthracis, and Pseudomonas aeruginosa, while the petroleum ether extract was inactive. The extracts were mildly active against Bacillus anthracis and Pseudomonas aeruginosa but exhibited the highest activity against Staphylococcus aureus. They also exhibited antifungal activity against Candida albicans and Aspergillus niger. An 80% aqueous ethanol extract of the roots did not have any effect on blood glucose levels during an oral glucose tolerance test (OGTT), in mice (P>0.05). With the exception of the dichloromethane and petroleum ether extracts, all the intermediate and polar extracts were toxic to brine shrimps giving LC(50) (95% confidence intervals) values ranging from 5.4 (3.5-8.4) to 17.4 (11.4-26.5) microg/ml, while that of cyclophosphamide, a standard anticancer drug, was 16.3 (10.6-25.2) microg/ml. Further work is in progress to isolate and identify active compounds in the extracts. PMID- 15652274 TI - Antifungal sesquiterpene from the root of Vernonanthura tweedieana. AB - The bioassay-guided fractionation of the antifungal dichloromethane extract from the roots of Vernonanthura tweedieana (Baker) H. Rob. (Asteraceae), using an agar overlay bioautographic method, allowed the isolation of one active sesquiterpene (1), identified as 6-cinnamoyloxy-1-hydroxyeudesm-4-en-3-one. Minimal inhibitory concentrations (MIC) and minimal fungicidal concentrations (MFC) of 1 showed Trichophyton mentagrophytes as the most sensitive strain, with the same MIC and MFC values (4 microg/ml). PMID- 15652275 TI - Neuropharmacological effects of the aqueous extract of Nauclea latifolia root bark in rats and mice. AB - The present study evaluated the neuropharmacological effects of the aqueous extract of Nauclea latifolia root bark in rodents. Effects on the spontaneous motor activity (SMA), exploratory behaviour, pentobarbital sleeping time, apomorphine-induced stereotypic behaviour and motor coordination (rota-rod performance) were investigated. The extract (50-200 mg/kg p.o.) significantly (P<0.05) decreased the SMA and exploratory behaviour in mice and prolonged pentobarbital sleeping time in rats dose-dependently. The extract also remarkably attenuated the intensity of apomorphine-induced stereotypy dose-dependently in mice, but had no effect on motor coordination as determined by the performance on rota-rod. These results indicate the presence of psychoactive substances in the aqueous extract of the root bark of Nauclea latifolia. PMID- 15652276 TI - Phenylethanoid glycosides from Cistanches salsa inhibit apoptosis induced by 1 methyl-4-phenylpyridinium ion in neurons. AB - In our study we investigated the neuroprotective effects of phenylethanoid glycosides (PhGs) from Cistanches salsa on 1-methyl-4-phenylpyridinium ion (MPP(+))-induced apoptosis in cerebellar granule neurons (CGNs). CGNs were treated with 100 microM MPP(+) for 24h to induce apoptosis, simultaneously CGNs were incubated with PhGs at 10, 20 and 40 microg/ml, respectively. In addition CGNs were pretreated with PhGs at 20 microg/ml for 6, 12, 24 h, respectively, and then treated with 100 microM MPP(+) for 24 h. 3-(4,5-Dimethylthiazol-2-ylthiazol 2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay revealed that the treatment of CGNs with PhGs inhibited the decrease of cell viability induced by MPP(+). The activation of caspase-3 and caspase-8 was induced by MPP(+) in apoptosis. The caspase-3 and caspase-8 fluorogenic assays showed that the treatments of CGNs with PhGs efficiently suppressed the activation of caspase-3 and caspase-8 induced by MPP(+). It is concluded that PhGs can prevent the MPP(+)-induced apoptosis in CGNs and exert its anti-apoptosis effect by inhibiting caspase-3 and caspase-8 activities. PMID- 15652277 TI - Suppressive effects of a water extract of Ulmus davidiana Planch (Ulmaceae) on collagen-induced arthritis in mice. AB - Ulmus davidiana Planch (Ulmaceae) has long been known to have anti-inflammatory and protective effects on damaged tissue, inflammation and bone resorption. Therefore, this study was undertaken to address (1) whether the water extract of the bark of Ulmus davidiana Planch (Ulmaceae) (UD) can modulate the expression of inducible inflammatory cytokines in mice; (2) in order to assess the therapeutic effects of UD in collagen-induced arthritis (CIA) in mice. DBA/1 mice were immunized with bovine type II collagen. After a second collagen immunization, mice were treated with UD orally at 100mg/kg once a day for 3 weeks. Paws were evaluated macroscopically for redness, swelling and deformities. The levels of TNF-alpha and IL-1beta in the ankle were examined. The severity of arthritis within the knee joints was evaluated by histological assessment of cartilage destruction and pannus formation. Administration of UD significantly suppressed the progression of CIA and inhibited the production of TNF-alpha and IL-1beta in the paws. The erosion of cartilage was dramatically reduced in mouse knees after treatment with UD. In the serum of UD-treated mice, the levels of IL-4 and IL-10, anti-inflammatory cytokines, were increased. From the results, it was concluded that administration of UD has therapeutic effects on CIA including protection of cartilage and RA for a potential therapy. PMID- 15652278 TI - Aqueous extract of Ilex paraguariensis decreases nucleotide hydrolysis in rat blood serum. AB - Mate is a xanthine-containing beverage, which is prepared as an infusion of the dried and ground leaves of Ilex paraguariensis St. Hil. (Aquifoliacea). Previous reports have shown that Ilex paraguariensis has the highest levels of caffeine and theobromine when compared to other Ilex species. Furthermore, mate is able to interfere in the circulatory system, acting as a diuretic and hypotensive agent. Many processes of vascular injury result in the release of adenine nucleotides, which exert a variety of effects. Nucleoside 5' tri- and diphosphates may be hydrolyzed by members of the ecto-nucleoside triphosphate diphosphohydrolase (E NTPDase) family. The synchronic action of a NTPDase and a 5'-nucleotidase promotes the catabolism of ATP to adenosine, which is able to control the extracellular nucleotides/nucleosides ratio. The chronic ingestion of aqueous extract of Ilex paraguariensis by rats during 15 days significantly decreased ATP (55%), ADP (50%) and AMP (40%) hydrolysis in blood serum. These results suggest changes in the balance of purine levels induced by Ilex paraguariensis ingestion. Considering the potential effects of Ilex paraguariensis in the circulatory system, these results may be relevant since NTPDases are a novel drug target for the treatment of cardiovascular diseases. PMID- 15652279 TI - Antitussive effect of Carum copticum in guinea pigs. AB - Several therapeutic effects including anti-asthma and dyspnea have been described for the seeds of Carum copticum In previous studies the relaxant and anticholinergic (functional antagonism) effects, histamine (H(1)) inhibitory effect of Carum copticum have been demonstrated on guinea pig tracheal chains. In the present study the antitussive effect of this plant was evaluated. The antitussive effects of aerosols of two different concentrations of aqueous and macerated extracts and carvacrol, codeine, and saline were tested by counting the number of coughs produced due to aerosol of citric acid 10 min after exposing animals to aerosols of different solutions (for carvacrol n=5 and for other solutions n=6). The results showed significant reduction of cough number obtained in the presence of both concentrations of aqueous and macerated extracts and codeine (p<0.001 for extracts and p<0.01 for codeine). The cough number obtained in the presence of higher concentration of aqueous and macerated extracts was significantly less than those of lower concentrations (p<0.05 for both extracts). In addition the cough number obtained in the presence of both concentrations of aqueous and macerated extracts was significantly lower than that of codeine (p<0.05 to 0.001). However, carvacrol did not show any antitussive effect. These results indicated an antitussive effect of Carum copticum which was even greater than that of codeine at concentrations used. In addition the antitussive effect of Carum copticum was not due to its main constituent, carvacrol. PMID- 15652280 TI - Cheongyeolsaseuptang inhibits production of TNF-alpha, IL-6 and IL-8 as well as NF-kappa B activation in human mast cells. AB - Traditional Korean medicine, Cheongyeolsaseuptang (CYSST) has been widely applied as a treatment of rheumatoid arthritis (RA) in Korea. However, its effect in experimental models remains unknown. Recent reports suggest that in patients with RA, synovial mast cells increase in number and show signs of activation and production of cytokines. In this study, we investigated the effect of CYSST on production of cytokines by activated human mast cell line, HMC-1. When CYSST (1mg/ml) was added, the production of tumor necrosis factor-alpha, interleukin (IL)-6, and IL-8 was significantly inhibited about 37, 33.6, and 48%, respectively on phorbol 12-myristate 13-acetate (PMA) plus calcium ionophore A23187-stimulated HMC-1 cells. In addition, CYSST inhibited PMA plus A23187 induced activation of nuclear factor-kappaB. These findings may help understanding the mechanism of action of this medicine leading to control activated mast cells on inflammatory condition like RA. PMID- 15652281 TI - Pharmacological properties of citrus and their ancient and medieval uses in the Mediterranean region. AB - This paper reviews the pharmacological properties of Mediterranean-grown citrus species (Citrus L., Rutaceae), including citron (Citrus medica L.), lime (Citrus xauantiifolia [Christm.] Swingle), lemon (Citrus xlimon [L.] Osbeck), bitter orange (Citrus xaurantium L.) and pomelo (Citrus maxima [Burm.] Merr.), as referred to in ancient, medieval and 16th century sources. The virtues of the species reported in these texts were compared to those known to modern science. A much broader spectrum of pharmacological properties was recorded by these early writers than one might expect. The use of the citron and lemon as antidotes for 'poison and venom' is recorded in the very earliest material. According to modern scientific literature the citron and the bitter orange may possess anti-cancer activity, lime may have an immunomodulatory effect in humans, and the pomelo may be useful for treating circulatory problems. Lemons might even ease hangover symptoms. Research is required to confirm these properties. PMID- 15652282 TI - Antioxidant properties and TLC characterization of four Chilean Haplopappus species known as bailahuen. AB - "Bailahuen" is the common name of a medicinal shrub native to Chile where this resinous herb is widely used for its liver stimulating properties. Although the official species is Haplopappus baylahuen Remy (Asteraceae), other species of the same genus are also used in different regions as "bailahuen". A thin layer chromatography (TLC) method for rapid identification of different species and detection of adulterations is described for four of the species: Haplopappus baylahuen, Haplopappus taeda Reiche, Haplopappus multifolius Phil., and Haplopappus remyanus Wedd. To confirm efficiency in all species antioxidant properties were screened in resins, infusions, and methanolic extracts by tests of lipid peroxidation in erythrocytes and free radical scavenging activity by DPPH. In both studies Haplopappus baylahuen showed the lowest antioxidant capacity. In DPPH analyses, infusion and resins of Haplopappus baylahuen also showed the lowest and Haplopappus remyanus the major inhibiting activity of free radicals, while Haplopappus multifolius proved to have the highest result when the methanolic extracts were used. The chemical characterization of the studied species showed important levels of flavonoids and coumarins, with flavonoids predominating in Haplopappus taeda, coumarins in Haplopappus multifolius and both of them in Haplopappus baylahuen and Haplopappus remyanus. PMID- 15652283 TI - Large-scale and effective screening of Korean medicinal plants for inhibitory activity on matrix metalloproteinase-9. AB - Matrix metalloproteinase-9 (MMP-9) degrades type IV collagen constituting the major structural component of the basement membrane and extra cellular membrane. The enzymatic activity is found to be elevated in tumor tissues. With the aim of finding novel MMP-9 inhibitors from natural products, 87 extracts of oriental medicinal herbs, which are used as prescriptions for cancer treatment in traditional Korean medicine, were screened for their inhibitory activities towards MMP-9. It was found that most of the hexane and chloroform fractions as well as water extracts showed a weak inhibitory effect on MMP-9 activity at a concentration of 100mug/ml. However, a strong inhibition was found in the butanol fractions of Cinnamomum cassia PRESL, Magnolia obovata THUEB., Magnolia officinalis REHD. et WILS., Magnolia officinalis REHD. et WILS. var. biloba REHD. et WILS., and Euonymus alatus (THUNB.) SIEB. with inhibitory activity (>90%) at a concentration of 100 microg/ml. PMID- 15652284 TI - Effect of storage on the chemical composition and biological activity of several popular South African medicinal plants. AB - The in vitro biological activity of nine frequently used medicinal plants in South Africa was assessed and re-assessed after various lengths of storage. The plants investigated were Alepidea amatymbica, Leonotis leonurus, Drimia robusta, Vernonia colorata, Merwilla natalensis, Eucomis autumnalis, Bowiea volubilis, Helichrysum cymosum and Siphonochilus aethiopicus. Water, ethanol and hexane extracts of fresh, 90-day-old and 1-year-old material were assayed for antibacterial activity against four strains of bacteria and for COX-1 inhibition activity. TLC-fingerprints of the fresh and stored extracts were produce to document chemical changes. Alepidea amatymbica, Eucomis autumnalis, Helichrysum cymosum, Leonotis leonurus, Siphonochilus aethiopicus and Vernonia colorata were investigated further as to the effect of 1 year's storage. Elevated temperature and humidity (55 degrees C and 100% relative humidity) were used to accelerate the ageing process of Alepidea amatymbica, Leonotis leonurus and Vernonia colorata plant material for further investigation. The TLC-fingerprints indicated that there was chemical breakdown during storage in certain species. The degree of changes in biological activity and chemistry due to storage were species specific. In general, antibacterial activity was retained in most species while COX-1 inhibition activity was lost rapidly. PMID- 15652285 TI - Antidiabetic activity of Caesalpinia bonducella F. in chronic type 2 diabetic model in Long-Evans rats and evaluation of insulin secretagogue property of its fractions on isolated islets. AB - Caesalpinia bonducella F., is a shrub widely distributed throughout the coastal region of India and is ethnically used by the tribal people of Andaman and Nicober Island as a remedy of symptoms of diabetes mellitus. This ethnic report prompted the detail investigation of hypoglycemic activity of Caesalpinia bonducella seeds, initially on physiological hyperglycemic model and then on type 1 and type 2 sub-acute diabetic animal models which has already been reported. Evaluation of different extracts from Caesalpinia bonducella in chronic type 2 diabetic model alongwith insulin secretagogue activity of five fractions isolated from the Caesalpinia bonducella seed kernel are presented in this paper. Both the aqueous and ethanolic extracts showed potent hypoglycemic activity in chronic type 2 diabetic model. Two fractions BM 169 and BM 170 B could increase secretion of insulin from isolated islets. PMID- 15652286 TI - Leaf extract of Crinum bulbispermum has antinociceptive activity in rats. AB - In Sri Lankan folk medicine, the leaves of Crinum bulbispermum Burm. (Family: Amaryllidaceae) are claimed to be useful for earache, which is suggestive of antinociceptive activity. The aim of this study was to examine scientifically the antinociceptive activity of Crinum bulbispermum leaves. One of three doses (1000, 1500 and 3000 mg/kg, N=6/group) of aqueous leaf extract (ALE) or 1 ml of distilled water (DW) (vehicle) (N=6) was orally administered to male rats and antinociceptive activity was determined using three models of nociception (tail flick, hot plate and formalin tests). In the hot plate test, slight (18-42%) but significant (P<0.05) overall prolongation of reaction time was evident up to 4h. In the hot plate test, reaction time was markedly (84-121%) and significantly increased by the high dose up to 3h. In contrast, in the formalin test, the high and mid doses significantly impaired the number of lickings and time spent on licking at both phases, whilst the lower dose impaired only the time spent on licking at the late phase. Collectively these results suggest that the antinociception is mediated both spinally and supraspinally and is effective against phasic and continuous non-inflammatory/inflammatory pain. Additionally, the ALE had antihyperalgesic activity. The antinociception of the ALE appear to results from opioid mechanisms, sedation and antioxidant activities. Unfortunately, ALE is mild to moderately toxic. PMID- 15652287 TI - Traditional plant use in the areas of Monte Vesole and Ascea, Cilento National Park (Campania, Southern Italy). AB - The present ethnobotanical field study conducted during summer 2003 in two distinct regions of the Cilento National Park (Mt Vesole and Ascea) documents the local use of 90 different plant species for medicinal, food and domestic purposes. Overall, 59 people native to the area were interviewed, and 883 use reports have been recorded. The scientific names, local names, plant parts used, preparation and administration processes are given and compared with practices in other Italian regions. In total, 63 species are documented as medicinal, 49 as food, and 22 as craft plants. Over 40% of all species are used in more than one category and over half of the food plants are also used for medicinal purposes. In general the recorded species are well known in the traditional phytotherapy of Campania and Italy. However, some uses are unusual and are discussed in detail. PMID- 15652288 TI - Screening of traditionally used Lebanese herbs for neurological activities. AB - In the recent decades the use of traditional medicine in Lebanon has increased. Aqueous, ethanol and ethyl acetate extracts of seven Lebanese plants that are used traditionally for neurological disorders as Alzheimer's disease, epilepsy and affective disorders as depression were tested for inhibition of acetylcholinesterase and affinity to the GABA(A)-benzodiazepine site and to the serotonin transporter. Ethyl acetate extracts of Salvia triloba, Lavandula officinalis, Origanum syriacum and Artemisia herba-alba exhibited weak activity in the acetylcholinesterase assay. None of the plants were active in the serotonin transporter assay. An ethanolic extract of Artemisia herba-alba had good affinity to the GABA(A)-benzodiazepine receptor site; ethanolic extracts of Melissa officinalis and Salvia triloba had moderate activity. PMID- 15652289 TI - Ayurveda and gynecological disorders. AB - The science of life--Ayurveda is practiced in India since time immemorial. Besides being cheap and easily available Ayurvedic drugs are considered safe. Moreover, there is surge in the interest in Ayurveda due to quest of alternative medicines. Many of the gynecological disorders being not reported to the physicians, are treated with household remedies in India. The science of Ayurveda deals with these issues in a systematic manner as evident from the classification of diseases available and the number of plant drugs or the combinations thereof available for the treatment. In the present article, Ayurvedic herbal formulations and single plant drugs used traditionally in treatment of gynecological disorders are described. PMID- 15652290 TI - Construction of a rhamnose mutation in Bacillus anthracis affects adherence to macrophages but not virulence in guinea pigs. AB - Carbohydrate analyses of whole-spore extracts have confirmed the presence of rhamnose in the spore of the fully virulent Ames strain of Bacillus anthracis. A gene cluster containing loci with high homology to the rhamnose biosynthetic genes, rmlACBD, was identified within the B. anthracis chromosome. The first gene of this cluster, rmlA, was inactivated by forming a merodiploid cointegrate using an internal fragment of the gene within the Ames strain of B. anthracis to construct the mutant strain Ames-JAB1. Carbohydrate analysis of spores from this mutant demonstrated the loss of rhamnose. When assaying for spore infection of macrophages, we detected a significant decrease in the recovery with the Ames JAB1 strain compared to the recovery with the Ames wild-type strain. When pre treating macrophages with cytochalasin-D, spores of the mutant were further hindered in recovery, indicating that the spores were not able to bind as well to the macrophages. However, in guinea pigs challenge experiments, no difference in virulence was observed between the mutant and wild-type strains. These results suggest that the incorporation of rhamnose into the spore coat of B. anthracis is required for optimal interaction with macrophages but is not required for full virulence in this animal model. PMID- 15652291 TI - Interleukin-10 (IL-10) ameliorates corneal disease in a mouse model of recurrent herpetic keratitis. AB - Interleukin 10 (IL-10), a moderator of Delayed Type Hypersensitivity (DTH) responses, has been demonstrated to be present late in acute HSV corneal infection and may help limit blinding inflammatory lesions there. In contrast, IL 10 is present early in the development of recurrent herpetic stromal keratitis (HSK) lesions in mice. To determine the role of IL-10 and DTH responses in recurrent HSK, we examined DTH responses and disease parameters in latently infected IL-10 knock out (KO) mice, and latently infected normal mice that were untreated or received anti-IL-10 antibodies or recombinant IL-10 following ultraviolet-B stimulated ocular HSV recurrence. Low DTH responses were associated with less severe corneal disease while high DTH responses were associated with greater corneal disease. In IL-10 KO mice, and in normal mice given anti-IL-10 antibodies, corneal opacification was increased and DTH responses were significantly prolonged. Normal mice receiving rIL-10 by ocular and intra peritoneal routes had less severe corneal lesions. Our results indicate that IL 10 and DTH responses play an important role in the pathogenesis of recurrent HSK in mice. PMID- 15652292 TI - Bovine platelets activated by Haemophilus somnus and its LOS induce apoptosis in bovine endothelial cells. AB - Haemophilus somnus is a bacterial pathogen that causes respiratory disease and vasculitis in cattle. Thrombotic meningoencephalitis (TME) and other severe forms of H. somnus-mediated vascular disease are characterized histopathologically by vasculitis, thrombosis, and infiltration of polymorphonuclear cells. It has been reported previously that activated human platelets express CD40L, FasL and P selectin (CD62P). We hypothesized that if these surface markers are up-regulated on bovine platelets after in vitro exposure to H. somnus and its lipooligosaccharide (LOS), they might contribute to endothelial cell damage. Using flow cytometry, we demonstrated low baseline expression of these molecules by bovine platelets and increased expression following in vitro stimulation with ADP, H. somnus or H. somnus LOS. H. somnus stimulated platelets were capable of causing apoptosis in endothelial cells as measured by Hoechst-33342 staining and caspase-3 activity. If these events occur in vivo, they might promote vascular damage and endothelial cell apoptosis, leading to the development of vasculitis and thrombosis that characterize bovine H. somnus infection. PMID- 15652293 TI - Mouse model characterisation for anthrax vaccine development: comparison of one inbred and one outbred mouse strain. AB - In order to evaluate the immunogenicity and protective efficacy of anthrax vaccine candidates a suitable small animal model is required. The inbred A/J strain of mouse has been selected as a potential model, and its immune response to immunisation with recombinant protective antigen (rPA) vaccine characterised, by assessment of rPA specific antibody production, and protection against injected challenge, with the unencapsulated STI strain of Bacillus anthracis. Studies were conducted to determine the time required post immunisation to develop a protective immune response, to define the minimum protective dose of vaccine required and to assess the long-term immune response to immunisation. From the results of these studies it was possible to establish that the A/J mouse is a consistent and robust small animal model for rPA vaccine testing. A comparison of the immune response to rPA vaccine immunisation in the Turner Outbred (TO) mouse strain was also conducted. Both inbred and outbred mouse strains displayed a predominantly Th2 biased immune response and showed a comparable antibody response to rPA immunisation. An assessment of protection in the TO mouse against aerosol challenge with the fully virulent strain of B. anthracis, Ames, was also made. PMID- 15652294 TI - Ultrastructural analysis of the interactions between Bordetella pertussis, Bordetella parapertussis and Bordetella bronchiseptica and human tracheal epithelial cells. AB - Bordetella pertussis, Bordetella parapertussis and Bordetella bronchiseptica are respiratory pathogens that colonize the respiratory tract of their host after adhesion to the respiratory epithelium. Presently, the intracellular fate of these bacteria in human tracheal epithelial cells was compared by use of transmission electron microscopy. The three species, even when cytotoxic, were taken-up by epithelial cells. Although, some intracellular bacteria appeared morphologically intact and survived a few days inside epithelial cells, most of them appeared quickly degraded, phenomenon which was associated with an intense cell metabolic activity. Even cytotoxic Bordetella species is ultimately killed by human epithelial cells. PMID- 15652295 TI - Molecular detection of Bartonella spp. in the dental pulp of stray cats buried for a year. AB - Bartonella henselae causes chronic bacteremia in cats. To test if B. henselae DNA can be recovered from the dental pulp of cats buried a year previously, we used PCR with primers for a sequence of the conserved groEL to test 104 teeth from 11 cats. Seven of the cats were found positive; canine teeth were more frequently positive than molar teeth. Where PCR sequences could be determined, they were identical to those of B. henselae Marseille (four cats), B. henselae Houston (one cat) or similar to those of B. grahamii (one cat). Our study indicates that dental pulp from the teeth of cats, especially the canine teeth, may be used for the PCR detection of Bartonella in animals buried for a year. PMID- 15652296 TI - Combined MEG/EEG analysis of the interictal spike complex in mesial temporal lobe epilepsy. AB - We studied the functional organization of the interictal spike complex in 30 patients with mesial temporal lobe epilepsy (MTLE) using combined magnetoencephalography (MEG)/electroencephalography (EEG) recordings. Spikes could be recorded in 14 patients (47%) during the 2- to 3-h MEG/EEG recording session. The MEG and EEG spikes were subjected to separate dipole analyses; the MEG spike dipole localizations were superimposed on MRI scans. All spike dipoles could be localized to the temporal lobe with a clear preponderance in the medial region. Based on dipole orientations in MEG, patients could be classified into two groups: patients with anterior medial vertical (AMV) dipoles, suggesting epileptic activity in the mediobasal temporal lobe and patients with anterior medial horizontal (AMH) dipoles, indicating involvement of the temporal pole and the anterior parts of the lateral temporal lobe. Whereas patients with AMV dipoles had strictly unitemporal interictal and ictal EEG changes during prolonged video-EEG monitoring, 50% of patients with AMH dipoles showed evidence of bitemporal affection on interictal and ictal EEG. Nine patients underwent epilepsy surgery so far. Whereas all five patients with AMV dipoles became completely seizure-free postoperatively (Class Ia), two out of four patients with AMH dipoles experienced persistent auras (Class Ib). This difference, however, was not statistically significant. We therefore conclude that combined MEG/EEG dipole modeling can identify subcompartments of the temporal lobe involved in epileptic activity and may be helpful to differentiate between subtypes of mesial temporal lobe epilepsy noninvasively. PMID- 15652297 TI - Cortical response to subjectively unconscious danger. AB - Cortical involvement in the evolution-favored automatic reaction to danger was studied. Electrical neural activity was recorded from 31 subjects, reporting fear of spiders, at 60 scalp locations. Visual stimuli containing spiders (negative elements) or, alternatively, nonnegative elements were presented to subjects, though they were unaware of their presence: a concurrent visual detection task using consciously perceived targets was administered. Spatial and temporal principal component analyses were employed to define and quantify, in a reliable manner, the main components of the neuroelectrical response to unconscious stimuli, and a source localization algorithm provided information on their neural origin. Results indicated that around 150 ms after stimulus onset, ventromedial prefrontal areas previously reported as responding rapidly to danger-related (conscious) stimuli were activated by unconsciously perceived spiders more markedly than by nonnegative unconscious stimuli. Subsequently, around 500 ms after stimulus onset, activation of the posterior cingulate and visual association cortices increased in this same direction. These data support previous results indicating that the ventromedial prefrontal cortex is involved in the top-down regulation of attention (through its capability to modulate the activity of posterior cortices in charge of visual processing) and that it automatically facilitates danger processing. PMID- 15652298 TI - Linking semantic priming effect in functional MRI and event-related potentials. AB - The aim of this study is to examine the neural substrates involved in semantic priming using a combined event-related functional magnetic resonance imaging (fMRI) and event-related potentials (ERP) study. Twelve subjects were instructed to judge whether the presented target word was a real word or a nonword. Under the related condition, target words were preceded by a semantically related prime word. On the other hand, under the unrelated condition, prime words did not have semantic relatedness with the target word. The reaction time for reaching a judgment was longer under the unrelated condition than under the related condition, indicating that the recognition of target words is promoted by semantic priming under the related condition. In the fMRI results, we found reduced activity in the dorsal and ventral left inferior frontal gyrus, the anterior cingulate, and left superior temporal cortex for related versus unrelated conditions (i.e., the repetition suppression effect). ERP analysis revealed that the amplitude of the N400 component was reduced under the related condition compared with the unrelated condition (i.e., the N400 priming effect). Correlation analysis between the BOLD repetition suppression effect and the N400 priming effect decomposed by independent component analysis (ICA) across subjects showed significant correlation in the left superior temporal gyrus. This finding is consistent with the recent MEG data suggesting that the source of N400 is judged to be the bilateral superior temporal lobe. We discussed this finding herein in relation to the modulation of access to the phonological representation caused by semantic priming. PMID- 15652299 TI - In vivo detection limits of magnetically labeled embryonic stem cells in the rat brain using high-field (17.6 T) magnetic resonance imaging. AB - Stem cell transplantation is a promising therapeutic approach for several neurological disorders. However, it has yet to fulfill its high expectations, partially due to the lack of a reliable noninvasive method for monitoring the biodistribution of the grafted stem cells in vivo. We have used high-resolution magnetic resonance imaging (MRI) at 17.6 T, combined with efficient magnetic labeling of the stem cells with iron oxide nanoparticles, in order to assess the in vivo detection limit in small animal models. Injection of different concentrations of magnetically labeled stem cells in gel phantoms led to significant reductions in image intensity from small cellular clusters of less than 10 cells. To determine the detection limit in vivo, various numbers of both labeled and unlabeled cells were injected stereotactically into the striatum of rats. Significant hypointense signal changes were observed for 100 labeled cells. After injection of approximately 20 labeled cells, signal reduction at the injection site was observed but could not be assigned unambiguously to the cells. Our results show that high-field MRI allows tracking of a minimal number of cells in vivo, well below the number used in previous studies, opening the possibility of gaining new insights into cell migration and differentiation. PMID- 15652300 TI - The cerebral oscillatory network associated with auditorily paced finger movements. AB - Motor tasks involve neural activity in a spatially distributed network. It is assumed that coherent activity between these brain structures reflects functional connectivity. The aim of the present study was to investigate brain areas associated with a unimanual auditorily paced finger-tapping task and to characterize their dynamic interplay. We examined cerebromuscular and cerebrocerebral coupling in 10 right-handed subjects using recordings of continuous brain activity with a 122-channel whole-head neuromagnetometer while subjects performed the task with both hands consecutively. Additionally, surface EMG of the first dorsal interosseus was measured. Our data demonstrate that an oscillatory network composed of primary sensorimotor cortex, lateral as well as mesial premotor areas, the posterior parietal cortex and thalamus contralateral, and cerebellum and primary auditory cortex ipsilateral to the tapping hand subserves task execution. Connectivity between these areas and direction of coupling agree well with anatomical findings. During the right-hand condition, additional oscillatory activity in the primary sensorimotor cortex ipsilateral to the tapping hand was evident. This result suggests an asymmetric motor control in right-handers. Cerebrocerebral coupling predominantly occurs at 8-12 Hz. Therefore, our data support the hypothesis that coupling at 8-12 Hz in a cerebello-thalamic-cortical network represents a fundamental characteristic of the motor system and provides evidence for the significance of 8-12 Hz oscillations in a large scale network during the execution of simple motor tasks. PMID- 15652301 TI - Lateralization of ventral and dorsal auditory-language pathways in the human brain. AB - Recent electrophysiological investigations of the auditory system in primates along with functional neuroimaging studies of auditory perception in humans have suggested there are two pathways arising from the primary auditory cortex. In the primate brain, a 'ventral' pathway is thought to project anteriorly from the primary auditory cortex to prefrontal areas along the superior temporal gyrus while a separate 'dorsal' route connects these areas posteriorly via the inferior parietal lobe. We use diffusion MRI tractography, a noninvasive technique based on diffusion-weighted MRI, to investigate the possibility of a similar pattern of connectivity in the human brain for the first time. The dorsal pathway from Wernicke's area to Broca's area is shown to include the arcuate fasciculus and connectivity to Brodmann area 40, lateral superior temporal gyrus (LSTG), and lateral middle temporal gyrus. A ventral route between Wernicke's area and Broca's area is demonstrated that connects via the external capsule/uncinate fasciculus and the medial superior temporal gyrus. Ventral connections are also observed in the lateral superior and middle temporal gyri. The connections are stronger in the dominant hemisphere, in agreement with previous studies of functional lateralization of auditory-language processing. PMID- 15652302 TI - Temporal dynamics of the BOLD fMRI impulse response. AB - Using computer simulations and high-resolution fMRI experiments in humans (n=6) and rats (n=8), we investigated to what extent BOLD fMRI temporal resolution is limited by dispersion in the venous vasculature. For this purpose, time-to-peak (TTP) and full-width at half-maximum (FWHM) of the BOLD impulse response (IR) function were determined. In fMRI experiments, a binary m-sequence probe method was used to obtain high-sensitivity model-free single-pixel estimates of IR. Simulations of postcapillary flow suggested that flow-related dispersion leads to a TTP and FWHM increase, which can amount to several seconds in larger pial veins. fMRI experiments showed substantial spatial variation in IR timing within human visual cortex, together with a correlation between TTP and FWHM. Averaged across the activated regions and across subjects, TTP and FWHM were 4.51+/-0.52 and 4.04+/-0.42 s, respectively. In regions of interest (ROI) weighted toward the larger venous structures, TTP and FWHM increased to 5.07+/-0.64 and 4.32+/-0.48 s, respectively. In rat somatosensory cortex, TTP and FWHM were substantially shorter than in humans (2.73+/-0.60 and 2.28+/-0.63 s, respectively). These results are consistent with a substantial macrovascular dispersive contribution to BOLD IR in humans, and furthermore suggest that neurovascular coupling is a relatively rapid process, with a resolution below 2.3 s FWHM. PMID- 15652303 TI - Automatic analysis of cerebral asymmetry: an exploratory study of the relationship between brain torque and planum temporale asymmetry. AB - Leftward occipital and rightward frontal lobe asymmetry (brain torque) and leftward planum temporale asymmetry have been consistently reported in postmortem and in vivo neuroimaging studies of the human brain. Here automatic image analysis techniques are applied to quantify global and local asymmetries, and investigate the relationship between brain torque and planum temporale asymmetries on T1-weighted magnetic resonance (MR) images of 30 right-handed young healthy subjects (15 male, 15 female). Previously described automatic cerebral hemisphere extraction and 3D interhemispheric reflection-based methods for studying brain asymmetry are applied with a new technique, LowD (Low Dimension), which enables automatic quantification of brain torque. LowD integrates extracted left and right cerebral hemispheres in columns orthogonal to the midsagittal plane (2D column maps), and subsequently integrates slices along the brain's anterior-posterior axis (1D slice profiles). A torque index defined as the magnitude of occipital and frontal lobe asymmetry is computed allowing exploratory investigation of relationships between this global asymmetry and local asymmetries found in the planum temporale. LowD detected significant torque in the 30 subjects with occipital and frontal components found to be highly correlated (P<0.02). Significant leftward planum temporale asymmetry was detected (P<0.05), and the torque index correlated with planum temporale asymmetry (P<0.001). However, torque and total brain volume were not correlated. Therefore, although components of cerebral asymmetry may be related, their magnitude is not influenced by total hemisphere volume. LowD provides increased sensitivity for detection and quantification of brain torque on an individual subject basis, and future studies will apply these techniques to investigate the relationship between cerebral asymmetry and functional laterality. PMID- 15652304 TI - Coherent theta-band EEG activity predicts item-context binding during encoding. AB - Episodic memories consist of semantic information coupled with a rich array of contextual detail. Here, we investigate the neural processes by which information about the sensory context of a learning event is "bound" to the semantic representation of the to-be-encoded item. We present evidence that item-context binding during encoding is mediated by frontoposterior electroencephalographic (EEG) phase locking within and between hemispheres in the theta (4-8 Hz) band. During a task in which subjects encoded words in different font colors, later memory for the word was associated with sustained frontal theta activity and frontoposterior theta-band coherence, primarily within the left hemisphere. When the word-color association was later successfully retrieved, however, neurons synchronized their theta-band responses bilaterally in a more sustained fashion, particularly during the latter part of the stimulus epoch (>800 ms). Our results confirm the importance of functional coupling between frontal and posterior regions for successful encoding. One interpretation of these data is hemispheric contributions to item and context encoding may be asymmetric, with left hemisphere coherence facilitating semantic processing of an item and right hemisphere coherence facilitating processing of sensory context. Theta-band coherence may be an important mechanism by which brain networks exchange information during learning. PMID- 15652305 TI - Neural correlates of conscious perception in the attentional blink. AB - If attending to a target in a rapid stream of visual stimuli within the next 400 ms or so, a second target in the stream is frequently not detected by an observer. This so-called attentional blink can provide a comparison of neural signals elicited by identical stimuli that, in one condition, reach conscious awareness and, in the other, fail to be selected for awareness. In the present study, using event-related functional magnetic resonance imaging (fMRI), differences of neural activation were studied in an attentional blink experiment in order to identify brain regions putatively involved in controlling the access of information to consciousness. Subjects viewed a rapid stream of black letters in which the second target (T2) was either presented within or outside the attentional blink period, or not at all. We observed an increase in activation for detected as compared to missed T2 presented during the attentional blink in frontal and parietal cortices. In contrast, in occipitotemporal regions activation was increased for missed as compared to detected T2. Furthermore, in several frontal and parietal areas, missed targets were associated with increased activity if compared to the condition in which no second target was presented. Finally, a selective decrease in activation for detected T2 presented during the attentional blink was observed in areas associated with emotional and predominantly automatic processing. While activations in occipitotemporal regions might mainly reflect duration of attentive search, the frontoparietal areas seem to be involved in a highly distributed network controlling visual awareness. PMID- 15652306 TI - Comparing extent of activation: a robust permutation approach. AB - The number of contiguous voxels activated in a brain image can differ between groups or conditions even though the amplitude of activation does not markedly differ. Existing techniques test for differences in amplitude given that extent (number of contiguous voxels) exceeds some threshold. We present a technique that tests for differences in extent of activation given that amplitude of activation exceeds some threshold. The technique was motivated by apparent differences in extent of regional cerebral blood flow (rCBF) between hypertensive and normotensive participants performing cognitive tasks. These data are used to illustrate our test for extent of activation. We threshold the estimated parameter map for each subject, count the number of voxels exceeding the threshold over a defined region enclosing activated cortical area, and test the hypothesis of difference in the number of activated voxels between the two groups. Due to the large number of zeros resulting from the thresholding and the occurrence of extreme observations, we use a Robust permutation test [Lambert, D., 1985. Robust two-sample permutation tests. Ann. Stat., 13, 606-625], which is based on the sum of censored log-likelihood ratios. This statistic has desirable properties relative to the usual permutation test in contaminated distributions, i.e., idealized histogram with outliers, and provides an appropriate and robust test of extent of activation between conditions or groups. PMID- 15652307 TI - Cerebral activation using a MR-compatible piezoelectric actuator with adjustable vibration frequencies and in vivo wave propagation control. AB - Functional magnetic resonance imaging (fMRI) studies are increasingly used in patients with brain tumors near the sensory motor cortex for planning of therapy. Passive stimuli can be helpful for reproducible results. The purpose of our study was to investigate frequency and amplitude dependencies of cerebral activation patterns using a vibratory stimulus that involves sensory and motor function and allows exact adjustment of vibratory frequencies and direct control of penetration depth into the tissue. Fifteen volunteers were studied with fMRI during vibratory stimulation of the right biceps muscle utilizing a block design (frequencies: 150 and 300 Hz, amplitudes: 400, 600, and 800 microm). In addition, visualization of the wave propagation into the biceps tissue itself was performed with a modified phase contrast sequence. A specially developed MR-compatible mechanical oscillator was used to apply the vibrotactile sensations. fMRI revealed activation of the left primary somatosensory cortex during application of both vibratory frequencies. Additionally, activity of the primary and supplementary motor cortex was revealed using 150-Hz stimuli, while only minimal at 300 Hz. The activity strength correlated with increasing stimulus amplitudes and the visualized penetration depth. Activation of motor areas was more pronounced at the beginning of the rest period. In conclusion, sensory motor areas can be activated using a piezoelectric actuator, with less pronounced motor area activation at higher frequencies. Our setup allowed local control of stimulus penetration through the tissue correlated to central activation, providing objective stimulus control. The pronounced activation of the motor cortex during the rest condition may reflect the subjective feeling of arm movement after the end of the stimulus. PMID- 15652308 TI - Viewing speech modulates activity in the left SI mouth cortex. AB - The ability to internally simulate other persons' actions is important for social interaction. In monkeys, neurons in the premotor cortex are activated both when the monkey performs mouth or hand actions and when it views or listens to actions made by others. Neuronal circuits with similar "mirror-neuron" properties probably exist in the human Broca's area and primary motor cortex. Viewing other person's hand actions also modulates activity in the primary somatosensory cortex SI, suggesting that the SI cortex is related to the human mirror-neuron system. To study the selectivity of the SI activation during action viewing, we stimulated the lower lip (with tactile pulses) and the median nerves (with electric pulses) in eight subjects to activate their SI mouth and hand cortices while the subjects either rested, listened to other person's speech, viewed her articulatory gestures, or executed mouth movements. The 55-ms SI responses to lip stimuli were enhanced by 16% (P<0.01) in the left hemisphere during speech viewing whereas listening to speech did not modulate these responses. The 35-ms responses to median-nerve stimulation remained stable during speech viewing and listening. Own mouth movements suppressed responses to lip stimuli bilaterally by 74% (P<0.001), without any effect on responses to median-nerve stimuli. Our findings show that viewing another person's articulatory gestures activates the left SI cortex in a somatotopic manner. The results provide further evidence for the view that SI is involved in "mirroring" of other persons' actions. PMID- 15652309 TI - Signal and noise characteristics of Hahn SE and GE BOLD fMRI at 7 T in humans. AB - At very high magnetic fields, GE BOLD fMRI is expected to contain nonspecific contributions and behave differently than HSE fMRI data. Similarly, the two approaches can conceivably suffer from different contributions to temporal instabilities in a times series that ultimately determine the contrast-to-noise ratio (CNR). We investigate the signal and signal fluctuation characteristics in GE and HSE fMRI data with the imaging parameters separately optimized for each contrast at 7 T. In HSE fMRI, activation-induced fractional signal change (DeltaS/S) decreased rapidly, and the ratio of standard deviations of image-to image fluctuations due to physiological processes (sigmaPhys) to thermal noise (sigmaTherm) remained constant with increasing voxel volume. In contrast, DeltaS/S as well as volume of activated voxels was virtually independent of voxel size for GE BOLD, and sigma(Phys)/sigmaTherm increased with increasing voxel size. The ratio of BOLD signal changes (GE/HSE) was much closer to 1 in tissue areas compared to vessel areas. These observations led to the conclusions that the spatial extent of the activation-induced DeltaS/S was much broader in the GE data, and that the physiological processes that give rise to the temporal fluctuations lost coherence over millimeter distances in HSE compared to GE fMRI data. While further studies are needed to characterize it fully, sigmaPhys in HSE data was clearly different than in GE data. It was concluded that HSE imaging yields a significantly reduced amount of nonspecific signals compared to GE imaging, and, would be the method of choice (over GE) for high-resolution applications in humans. PMID- 15652310 TI - Activity in the human brain predicting differential heart rate responses to emotional facial expressions. AB - The James-Lange theory of emotion proposes that automatically generated bodily reactions not only color subjective emotional experience of stimuli, but also necessitate a mechanism by which these bodily reactions are differentially generated to reflect stimulus quality. To examine this putative mechanism, we simultaneously measured brain activity and heart rate to identify regions where neural activity predicted the magnitude of heart rate responses to emotional facial expressions. Using a forewarned reaction time task, we showed that orienting heart rate acceleration to emotional face stimuli was modulated as a function of the emotion depicted. The magnitude of evoked heart rate increase, both across the stimulus set and within each emotion category, was predicted by level of activity within a matrix of interconnected brain regions, including amygdala, insula, anterior cingulate, and brainstem. We suggest that these regions provide a substrate for translating visual perception of emotional facial expression into differential cardiac responses and thereby represent an interface for selective generation of visceral reactions that contribute to the embodied component of emotional reaction. PMID- 15652311 TI - ROC evaluation of statistical wavelet-based analysis of brain activation in [15O] H2O PET scans. AB - This paper presents and evaluates a wavelet-based statistical analysis of PET images for the detection of brain activation areas. Brain regions showing significant activations were obtained by performing Student's t tests in the wavelet domain, reconstructing the final image from only those wavelet coefficients that passed the statistical test at a given significance level, and discarding artifacts introduced during the reconstruction process. Using Receiver Operating Characteristic (ROC) curves, we have compared this statistical analysis in the wavelet domain to the conventional image-domain Statistical Parametric Mapping (SPM) method. For obtaining an accurate assessment of sensitivity and specificity, we have simulated realistic single subject [15O]-H2O PET studies with different hyperactivation levels of the thalamic region. The results obtained from an ROC analysis show that the wavelet approach outperforms conventional SPM in identifying brain activation patterns. Using the wavelet method, activation areas detected were closer in size and shape to the region actually activated in the reference image. PMID- 15652312 TI - How do we perceive the pain of others? A window into the neural processes involved in empathy. AB - To what extent do we share feelings with others? Neuroimaging investigations of the neural mechanisms involved in the perception of pain in others may cast light on one basic component of human empathy, the interpersonal sharing of affect. In this fMRI study, participants were shown a series of still photographs of hands and feet in situations that are likely to cause pain, and a matched set of control photographs without any painful events. They were asked to assess on-line the level of pain experienced by the person in the photographs. The results demonstrated that perceiving and assessing painful situations in others was associated with significant bilateral changes in activity in several regions notably, the anterior cingulate, the anterior insula, the cerebellum, and to a lesser extent the thalamus. These regions are known to play a significant role in pain processing. Finally, the activity in the anterior cingulate was strongly correlated with the participants' ratings of the others' pain, suggesting that the activity of this brain region is modulated according to subjects' reactivity to the pain of others. Our findings suggest that there is a partial cerebral commonality between perceiving pain in another individual and experiencing it oneself. This study adds to our understanding of the neurological mechanisms implicated in intersubjectivity and human empathy. PMID- 15652313 TI - Language lateralization in young children assessed by functional transcranial Doppler sonography. AB - Compared to adults, children show superior recovery of language function after damage to the dominant brain hemisphere. Possible explanations are that children have different patterns of language representation or display different patterns of reorganization. Information about language lateralization in children could provide insights into the repair mechanisms of the young brain. While functional magnetic resonance imaging (fMRI) is usually difficult to perform in children younger than 5 years, functional transcranial Doppler sonography (fTCD) is nonfrightening and readily applicable in young and very young children. However, for serial examinations, sufficient validity and reliability are required. To this end, we designed a picture-description language task (PDLT) for fTCD examinations in children, compared the outcome to established protocols and determined the 1 month retest-reliability of the measurement in 16 children aged 2-9 years. The dependent variable was the task-related hemispheric perfusion difference based on averaged relative cerebral blood flow velocity (CBFV) increases in the middle cerebral arteries. This picture-description language lateralization index was compared to language lateralization by a phonetic word generation task (PWGT) in adults revealing good intermethod validity (r=0.70; P 5% and to define an appropriate time period for image acquisition, three untreated groups of animals received 29.6, 37, and 44.4 MBq of [123I]IBZM and underwent five consecutive SPECT acquisitions lasting 64 min each. Ratio calculations between specific striatal radioligand uptake and nondisplaceable cerebellar uptake revealed a secular equilibrium between 75 and 355 min post-tracer application in all three animal groups. Consequently, since the highest regional uptake values were obtained in the animal group receiving 44.4 MBq [123I]IBZM, this injection dose was considered to be appropriate. Finally, the capacity of the imaging method to detect distinct severity levels of striatal dopamine D2/D3 receptor loss was tested in a low, medium, and high dose quinolinic acid (QA) animal model of Huntington's disease. Motor impairment indicative of striatal dysfunction was monitored using amphetamine-induced rotational behavior and locomotor activity. Loss of striatal D2/D3R bearing medium-sized spiny neurons was assessed by DARPP-32 immunohistochemistry and compared to [123I]IBZM binding. Optical density measures of DARPP-32 immunohistochemistry demonstrated QA dose dependent mild to subtotal unilateral striatal lesions ranging from 29.4% to 96.9% when compared to the nonlesioned side. Linear regression analysis showed that measurements of striatal DARPP-32 optical density and striatal [123I]IBZM uptake of the lesioned side were highly correlated (r2=0.83; P<0.001) whereas correlation with locomotor activity was less tight (r2=0.23; P<0.05; amphetamine induced rotational behavior was not significantly correlated). This is the first study to demonstrate that in vivo [123I]IBZM SPECT and MRI coregistration are highly sensitive and, in contrast to behavioral measures, accurately detect mild to subtotal striatal lesions by measuring loss of D2/D3R availability. SPECT-MRI based estimation of regional [123I]IBZM uptake provides a cost effective and widely available in vivo imaging technique for assessing striatal integrity in animal studies. PMID- 15652318 TI - The neural correlates of spatial language in English and American Sign Language: a PET study with hearing bilinguals. AB - Rather than specifying spatial relations with a closed-class set of prepositions, American Sign Language (ASL) encodes spatial relations using space itself via classifier constructions. In these constructions, handshape morphemes specify object type, and the position of the hands in signing space schematically represents the spatial relation between objects. A [15O]water PET study was conducted to investigate the neural regions engaged during the production of English prepositions and ASL locative classifier constructions in hearing subjects with deaf parents (ASL-English bilinguals). Ten subjects viewed line drawings depicting a spatial relation between two objects and were asked to produce either an ASL locative classifier construction or an English preposition that described the spatial relation. The comparison task was to name the figure object (colored red) in either ASL or in English. Describing spatial relations in either ASL or English engaged parietal cortex bilaterally. However, an interaction analysis revealed that right superior parietal cortex was engaged to a greater extent for ASL than for English. We propose that right parietal cortex is involved in the visual-motoric transformation required for ASL. The production of both English prepositions and ASL nouns engaged Broca's area to a greater extent than ASL classifier constructions. We suggest that Broca's area is not engaged because these constructions do not involve retrieval of the name of an object or the name of a spatial relation. Finally, under the same task conditions, only left parietal activation was observed for monolingual English speakers producing spatial prepositions (H. Damasio et al., 2001, NeuroImage, 13). We conclude that the right hemisphere activation observed for ASL-English bilinguals was due to their life-long experience with spatial language in ASL. PMID- 15652319 TI - Productive and perceptive language reorganization in temporal lobe epilepsy. AB - The aim of this work was to determine whether productive and perceptive language functions are differentially affected in homogeneous groups of epilepsy patients with right and left temporal lobe epilepsy (TLE). Eighteen patients with left TLE, 18 with right TLE, and 17 healthy volunteers were studied using fMRI during performance of three tasks assessing the productive and perceptive aspects of language (covert semantic verbal fluency, covert sentence repetition, and story listening). Hemispheric dominance for language was calculated in the frontal and temporal regions using laterality indices (LI). Atypical lateralization was defined as a right-sided LI (LI<-0.20) in the frontal lobes during the verbal fluency task or in the temporal lobes during the story listening task. Control subjects and right TLE patients demonstrated a strong left lateralization for language in the frontal lobes during the fluency task, whereas activation was less lateralized to the left hemisphere in left TLE patients, although the difference did not reach significance. In the story listening and the repetition tasks, activation was significantly more right sided in the temporal lobes of patients with left TLE. Atypical language representation was found in 19% of TLE patients (five left and two right TLE). The shift toward the right hemisphere was significantly larger in the temporal than the frontal lobes in patients with atypical language lateralization compared to TLE patients with a typical language lateralization. Neuropsychological performances of patients with atypical language patterns were better than those of patients with typical patterns, suggesting that this reorganization may represent a compensatory mechanism. PMID- 15652320 TI - Separate neural systems for processing action- or non-action-related sounds. AB - The finding of a multisensory representation of actions in a premotor area of the monkey brain suggests that similar multimodal action-matching mechanisms may also be present in humans. Based on the existence of an audiovisual mirror system, we investigated whether sounds referring to actions that can be performed by the perceiver underlie different processing in the human brain. We recorded multichannel ERPs in a visuoauditory version of the repetition suppression paradigm to study the time course and the locus of the semantic processing of action-related sounds. Results show that the left posterior superior temporal and premotor areas are selectively modulated by action-related sounds; in contrast, the temporal pole is bilaterally modulated by non-action-related sounds. The present data, which support the hypothesis of distinctive action sound processing, may contribute to recent theories about the evolution of human language from a mirror system precursor. PMID- 15652321 TI - An fMRI study of reward-related probability learning. AB - The human striatum has been implicated in processing reward-related information. More recently, activity in the striatum, particularly the caudate nucleus, has been observed when a contingency between behavior and reward exists, suggesting a role for the caudate in reinforcement-based learning. Using a gambling paradigm, in which affective feedback (reward and punishment) followed simple, random guesses on a trial by trial basis, we sought to investigate the role of the caudate nucleus as reward-related learning progressed. Participants were instructed to make a guess regarding the value of a presented card (if the value of the card was higher or lower than 5). They were told that five different cues would be presented prior to making a guess, and that each cue indicated the probability that the card would be high or low. The goal was to learn the contingencies and maximize the reward attained. Accuracy, as measured by participant's choices, improved throughout the experiment for cues that strongly predicted reward, while no change was observed for unpredictable cues. Event related fMRI revealed that activity in the caudate nucleus was more robust during the early phases of learning, irrespective of contingencies, suggesting involvement of this region during the initial stages of trial and error learning. Further, the reward feedback signal in the caudate nucleus for well-learned cues decreased as learning progressed, suggesting an evolving adaptation of reward feedback expectancy as a behavior-outcome contingency becomes more predictable. PMID- 15652322 TI - Identification of the neural sources of the pattern-reversal VEP. AB - This study aimed to characterize the neural generators of the early components of the visual-evoked potential (VEP) to pattern-reversal gratings. Multichannel scalp recordings of VEPs and dipole modeling techniques were combined with functional magnetic resonance imaging (fMRI) and retinotopic mapping in order to estimate the locations of the cortical sources giving rise to VEP components in the first 200 ms poststimulus. Dipole locations were seeded to visual cortical areas in which fMRI activations were elicited by the same stimuli. The results provide strong evidence that the first major component of the VEP elicited by a pattern-reversal stimulus (N75/P85) arises from surface-negative activity in the primary visual cortex (area V1). Subsequent waveform components could be accounted for by dipoles that were in close proximity to fMRI activations in the following cortical areas: P95 (area MT/V5), P125/N135 (area V1), N150 (transverse parietal sulcus, TPS), N160 (ventral occipital areas VP, V4v, and V4/V8), and N180 (dorsal occipital areas V3A/V7). These results provide a detailed spatiotemporal profile of the cortical origins of the pattern-reversal VEP, which should enhance its utility in both clinical and basic studies of visual perceptual processing. PMID- 15652323 TI - Influence of bodily harm on neural correlates of semantic and moral decision making. AB - Moral decision-making is central to everyday social life because the evaluation of the actions of another agent or our own actions made with respect to the norms and values guides our behavior in a community. There is previous evidence that the presence of bodily harm--even if irrelevant for a decision--may affect the decision-making process. While recent neuroimaging studies found a common neural substrate of moral decision-making, the role of bodily harm has not been systematically studied so far. Here we used event-related functional magnetic resonance imaging (fMRI) to investigate how behavioral and neural correlates of semantic and moral decision-making processes are modulated by the presence of direct bodily harm or violence in the stimuli. Twelve participants made moral and semantic decisions about sentences describing actions of agents that either contained bodily harm or not and that could easily be judged as being good or bad or correct/incorrect, respectively. During moral and semantic decision-making, the presence of bodily harm resulted in faster response times (RT) and weaker activity in the temporal poles relative to trials devoid of bodily harm/violence, indicating a processing advantage and reduced processing depth for violence related linguistic stimuli. Notably, there was no increase in activity in the amygdala and the posterior cingulate cortex (PCC) in response to trials containing bodily harm. These findings might be a correlate of limited generation of the semantic and emotional context in the anterior temporal poles during the evaluation of actions of another agent related to violence that is made with respect to the norms and values guiding our behavior in a community. PMID- 15652324 TI - Noradrenaline mediates amygdala activation in men and women during encoding of emotional material. AB - The amygdala is a pivotal structure in humans for encoding of emotional information, as shown by recent imaging studies. It is unknown which neurotransmitters are specifically involved in the human amygdala, although in animal studies noradrenaline was shown to be essential. In our study, participants received the betablocker propranolol (which blocks the noradrenergic response) or placebo when watching neutral to highly negative arousing pictures. Amygdala activation, monitored with functional magnetic resonance imaging (fMRI), increased with emotional intensity of the pictures under placebo condition. Betablockade selectively decreased amygdala activation for emotional pictures of the second highest category, but not for the highest or lower (neutral) category pictures. Two findings add to the existing knowledge in this area. First, the activation pattern in the amygdala under placebo condition shows a nonlinearity related to the emotional categories of the pictures. Second, propranolol disturbs this activation pattern in the amygdala. Explorations with respect to gender show a similar effect of betablockade on amygdala activation in both men and women, but a difference in its effect on long-term memory for emotional pictures. This study supports the hypothesis that the neurotransmitter noradrenaline also mediates amygdala activity in humans when processing emotional stimuli and that betablockers can disrupt the normal activation pattern in the amygdala. PMID- 15652325 TI - Brain structural mapping using a novel hybrid implicit/explicit framework based on the level-set method. AB - This paper presents a novel approach to feature-based brain image warping, by using a hybrid implicit/explicit framework, which unifies many prior approaches in a common framework. In the first step, we develop links between image warping and the level-set method, and we formulate the fundamental mathematics required for this hybrid implicit/explicit approach. In the second step, we incorporate the large-deformation models into these formulations, leading to a complete and elegant treatment of anatomical structure matching. In this latest approach, exact matching of anatomy is achieved by comparing the target to the warped source structure under the forward mapping and the source to the warped target structure under the backward mapping. Because anatomy is represented nonparametrically, a path is constructed linking the source to the target structure without prior knowledge of their point correspondence. The final point correspondence is constructed based on the linking path with the minimal energy. Intensity-similarity measures can be naturally incorporated in the same framework as landmark constraints by combining them in the gradient descent body forces. We illustrate the approach with two applications: (1) tensor-based morphometry of the corpus callosum in autistic children; and (2) matching cortical surfaces to measure the profile of cortical anatomic variation. In summary, the new mathematical techniques introduced here contribute fundamentally to the mapping of brain structure and its variation and provide a framework that unites feature and intensity-based image registration techniques. PMID- 15652326 TI - Distinct neural substrates for the perception of real and virtual visual worlds. AB - Virtual environments have been frequently used for training and skill improvement. However, do real and virtual worlds engage the same brain states in human perceivers? We measured brain activity using functional magnetic resonance imaging (fMRI) while adults watched movie and cartoon clips, simulating real and virtual visual worlds, respectively. Relative to baselines using random static images, the medial prefrontal cortex (MPFC) and the cerebellum were activated only by movie clips of other humans. In contrast, cartoon clips of human and non human agents activated the superior parietal lobes, while movie clips of animals also activated the superior parietal lobes. Our fMRI findings suggest that the perception of real-world humans is characterised by the involvement of MPFC and the cerebellum, most likely for on-line representation of the mental states of others, whereas the perception of virtual-world agents engages the parietal cortex in attention to actions. PMID- 15652328 TI - Eco-epidemiological survey of Leishmania (Viannia) braziliensis American cutaneous and mucocutaneous leishmaniasis in Ribeira Valley River, Parana State, Brazil. AB - Leishmaniasis is endemic since last century in Adrianopolis Municipality, Ribeira Valley and is a serious public health. A study carried out during 1993-2003 on epidemiological surveys conducted in rural communities showed 339 new cases of cutaneous leishmaniasis (CL) detected from four municipalities (Adrianopolis, Cerro Azul, Doutor Ulysses and Rio Branco do Sul). A larger prevalence of cutaneous lesions was observed in rural workers (36%), women with domestic activities (18%), and younger students (31%). Multiple lesions were noticed in 53% of patients, but only one case of mucocutaneous leishmaniasis was reported. Twenty stocks were isolated from patients with characteristics lesions and were identified as Leishmania (Viannia) braziliensis using multi-locus enzyme electrophoresis (MLEE) and Random Amplified DNA (RAPD). In Phlebotominae survey, five species were obtained. Lutzomyia intermedia sl. represented 97.5% in peridomiciliar area and 100% in domicile. A canine serological survey made (Indirect Immunofluorescence Antibody Test, IFAT and Enzyme Linked Immunosorbent Assay, ELISA) in six rural county of Adrianopolis Municipality during 1998-1999 showed that 15.1% (24/159) of dogs were sera reactive. No lesions were observed in dogs and no parasite was isolated from lymph node aspirates and biopsies. In wild reservoirs study, only seven animals (Cricetidae, Desmodus sp. and edentates) were captured, but no parasites were found in culture from deep organs. The paper presents results of our 10 years study on cutaneous leishmaniasis survey in the Ribeira River Valley, East Region of Parana State, Brazil. Environment changes in this region are also discussed. PMID- 15652327 TI - A six-year follow-up of schoolchildren for urinary and intestinal schistosomiasis and soil-transmitted helminthiasis in Northern Tanzania. AB - Two cross-sectional studies with a 6-year interval were undertaken in two primary schools in neighbouring villages in Mwanga District, Tanzania, to determine the prevalence of schistosome and soil-transmitted helminth infections before and 6 years after treatment. Within this interval, health-related interventions such as one mass treatment of the villagers, health education, improvement of sanitation and access to safe water were undertaken in the villages. In 1996, urinary schistosomiasis occurred in Kileo (n=284) and Kivulini (n=350) in 37.0 and 86.3% of the schoolchildren. Intestinal schistosomiasis was found in 22.9 and 43.5% of the children. The infection with soil-transmitted helminths ranged between 2.7 and 18.1% in both villages. After 6 years, the prevalence of urinary schistosomiasis in schoolchildren remained nearly constant in Kileo (33.5%; n=544), but dropped from 86.3% to 70.0% in Kivulini (n=514). In the latter village the proportion of children heavily infected with Schistosoma haematobium (> or = 50 eggs/10 ml urine) decreased from 53.8% to 34.4%. With the exception of hookworm infection, soil-transmitted helminthiasis and intestinal schistosomiasis were significantly less seen in 2002 compared to the baseline. The previous chemotherapy of schoolchildren and villagers in 1996 and 1998, respectively, might have led to a reduced transmission of schistosomiasis in the following years. However, the reduction of prevalence of soil-transmitted helminthiasis is more likely to be the result of health-related interventions. PMID- 15652329 TI - Development of membrane-based tests for the detection of urinary antigens and antibodies in human toxoplasmosis: preliminary studies in Ghanaian patients. AB - Two membrane-based ELISA systems were used in detecting Toxoplasma antigens and anti-Toxoplasma antibodies in urine samples collected from 54 ophthalmology (22 suggestive active and 32 suggestive past infection) patients and 26 pregnant women attending obstetrics/gynaecology clinic (OGP), suspected of toxoplasmosis by eye examination, past medical records and questionnaire, respectively, in Ghana from mid-February to April 2002. The antigen detecting ELISA was able to demonstrate antigen in 100% (22/22) ophthalmology (active infection) and 62.5% (20/32) ophthalmology (past infection) patients, and 42% (11/26) of OGP which included 3 that were sero-negative prior to and during this study, giving an overall prevalence of 66.3% (53/80). The urinary antigen positive samples also included 6 that were negative for both the Dye Test (DT) and latex agglutination test (LAT). Antigen was not detected in the urine of 22 normal (sero-negative for antibodies to Toxoplasma) individuals. The membrane-based urinary antibody detecting sandwich ELISA also detected anti-Toxoplasma antibodies in 100% (22/22) of ophthalmology (active infection) and 81.3% (26/32) of ophthalmology (past infection) patients, a total of 89% (48/54); and 80.8% (21/26) of OGP with an overall prevalence of 86.3% (69/80), including 7 ophthalmology patients' samples that were sero-negative for both DT and LAT. Antibody sero-positivity of the samples was determined by DT as 87% (47/54) in ophthalmology patients and 73.1% (19/26) in pregnant women, LAT as 85.2% (46/54) and 65.4% (17/26), and an overall prevalence as 82.5% (66/80) and 78.8% (63/80), respectively. The membrane-based ELISA systems appear promising but need to be investigated further for its efficacy as reliable diagnostic tests. PMID- 15652330 TI - Activity of a paromomycin hydrophilic formulation for topical treatment of infections by Leishmania (Leishmania) amazonensis and Leishmania (Viannia) braziliensis. AB - Studies on in vitro skin permeation and in vivo anti-leishmanial activity in mice experimentally infected with Leishmania (Leishmania) major pointed out to the potential of a new paromomycin (PA) formulation (hydrophilic gel) for treatment of cutaneous leishmaniasis (CL). In this study, the activity of this formulation was evaluated in animals experimentally infected by Leishmania species that prevail in the New World. PA gel activity was compared to antimony treatment, since it is still the first choice treatment to the different clinical forms of leishmaniasis. The topical treatment activity with 10% PA gel in BALB/c mice infected by Leishmania (Leishmania) amazonensis was higher than that observed for parenteral antimony treatment, while the efficacy of these two regimes in hamsters infected by Leishmania (Viannia) braziliensis was similar. These results suggest that this formulation could be suitable for clinical studies and may represent an alternative novel formulation for topical treatment of CL. PMID- 15652331 TI - Serodiagnosis of Schistosoma mansoni infections in an endemic area of Burkina Faso: performance of several immunological tests with different parasite antigens. AB - The performance of indirect haemagglutination assays (IHA), enzyme-linked immunosorbent assays (ELISA) and indirect immunofluorescent antibody tests (IFAT) were compared with 450 sera from a Schistosoma mansoni-endemic area in Burkina Faso. All participants in this survey provided at least one sample each of stool, urine and serum. From those with an egg-negative Kato-Katz thick smear, a second stool sample was examined. IHA was based on either extracts of adult S. mansoni worms (SmIHA) or S. japonicum egg antigen (SjIHA). For ELISA, three antigen preparations were used, namely: (i) soluble S. mansoni adult worm antigens (SWAP); (ii) soluble S. mansoni egg antigens (SEA); and (iii) a cationic exchange fraction of S. mansoni eggs (CEF6). IFAT was performed with S. mansoni male worm sections. Among the egg-excretors, the sensitivity of ELISA was high and egg antigens performed slightly better (SEA, 96%; CEF6, 97%) than worm antigen (94%). Sensitivity of IHA was satisfactory with homologous (Sm, >85%), but not heterologous (Sj, 56%) parasite antigen. In IFAT, the parenchyma-associated fluorescence showed high sensitivity (95%), but gut-associated fluorescence, which is known to be a sensitive diagnostic marker for schistosome-infected European travelers, was observed only in 76% of a sub-sample of 100 of the endemic sera. Among sera from egg-negative individuals, many gave positive reactions in several or all of the tests employed. These reactions (formally "false positive") are considered to represent true infections, since chemotherapy had not yet been delivered to this population. For the purpose of further surveys in Burkina Faso or other resource-poor settings, we suggest IHA as an accurate diagnostic test and propose to further improve its performance by including egg rather than worm antigens. PMID- 15652332 TI - Effectiveness of different approaches to mass delivery of praziquantel among school-aged children in rural communities in Nigeria. AB - Schistosomiasis is hyperendemic in Nigeria. The national policy on schistosomiasis control has adopted praziquantel as the main drug of use in the control strategy aimed at reducing morbidity. However, the best delivery channel for praziquantel in schistosomiasis control in Nigeria is yet to be determined. This study assessed different channels for praziquantel delivery in a mass treatment effort. Three groups, each of two communities, were assigned to one of the following three channels: health facility (HF), school, and community for mass delivery of praziquantel to school-aged children. Distributors were trained on all aspects of praziquantel distribution and sufficient praziquantel tablets were provided for the drug distribution within 3 weeks following which an evaluation of the drug distribution by the different treatment channels was conducted. Overall, the community channel of delivery achieved the best coverage of 72.2% (range: 69-73%) compared to 44.3% (range: 39.5-62%) and 28.5% (range: 26.3-74.5%) for the HF and school channels, respectively. Community channel of praziquantel delivery as shown in this study ensures good coverage of both in and out-of-school children. PMID- 15652333 TI - A neotropical snail host of Schistosoma mansoni introduced into Africa and consequences for the schistosomiasis transmission Biomphalaria tenagophila in Kinshasa (Democratic Republic of Congo). AB - Malacological surveys carried out in the early 1970s in water bodies of the Kinshasa area, Lower Zaire (Democratic Republic of Congo), showed the appearance of a Biomphalaria species which was identified as Biomphalaria camerunensis. In 1976, other surveys confirmed the presence of the species in several sites and showed numerous infected snails with Schistosoma mansoni, demonstrating for the first time an active transmission of the parasite responsible of the intestinal schistosomiasis in this area. The most recent malacological sampling was carried out by one of us in 1994 in Mangungu River and revealed the presence of apparently the same snail species. However, conchological, anatomical and molecular studies showed that this snail may be considered as an introduced neotropical species, B. tenagophila. To our knowledge, this is the second example of the introduction of a neotropical snail host of schistosomes into Africa. PMID- 15652334 TI - Rice irrigation and schistosomiasis in savannah and forest areas of Cote d'Ivoire. AB - Prevalence and intensity of infection of Schistosoma haematobium and Schistosoma mansoni were studied in relation to irrigated rice cultivation in Cote d'Ivoire. Urine and stool samples were collected from 4 to 15-year-old children in 24 villages in the savannah zone and 21 villages in the forest zone. Villages were classified according to surrounding inland valleys into three agro-ecosystems: (R2) full or partial water control allowing two rice cycles per year; (R1) no or partial water control allowing one harvest per year and (R0) absence of rice growing. In the savannah zone, S. haematobium prevalence was 4.8%, 2.3% and 0.7% and S. mansoni prevalence was 16.1%, 11.9% and 2.1% in R2, R1 and R0, respectively. In the forest zone, S. haematobium prevalence was 0.9%, 4.4% and 1.7% and S. mansoni prevalence was 61.3%, 46.6% and 17.5% in R2, in R1 and R0, respectively. Prevalences of S. mansoni adjusted for village effects were significantly different between agro-ecosystems in both zones. Significance of differences between agro-ecosystems of S. haematobium infection were strongly influenced by outlying villages. In savannah rice growing villages, negative binomial regression on infection intensity of each species showed significant positive relations to the surface of rice cultivated inland valleys, whereas uncultivated inland valleys showed no significant relation. However, in forest rice growing villages, S. mansoni infection intensity showed significant positive relations to the surface of uncultivated inland valleys, whereas surface water on rice cultivated land showed significant negative relations with infection intensity of each schistosomiasis species. PMID- 15652335 TI - A discrepancy between cystic echinococcosis confirmed by ultrasound and seropositivity in Turkish children. AB - In three randomly selected villages of Manisa, Turkey, 630 primary school children were examined for cystic echinococcosis (CE) by a portable ultrasound scanner (US) and chest microfilm. Sera samples of 483 (76.7%) children were tested for anti-E. granulosus antibodies by ELISA and indirect hemagglutination (IHA) tests. Hepatic CE were detected in two cases (0.3%) by US, while 43 (8.9%) and 49 (10.1%) cases were found to be positive for CE by ELISA and IHA, respectively. The high seropositivity levels could have been attributed to extra abdominal or abortive Echinococcus infections, but probably most of them were false-positives. Due to the discrepancy in results, US should be selected as the primary test in field studies and serologic tests should be performed in all cases with suspected lesions. We could not find any reported community based study on CE in Turkey, in which US was applied; but our results suggest that extensive epidemiological studies are required. PMID- 15652336 TI - Real-time measurements of membrane surface dynamics on macrophages and the phagocytosis of Leishmania parasites. AB - Defocusing microscopy was used for real-time observation and quantification of membrane surface dynamics in murine bone marrow macrophages. Small random membrane fluctuations (SRMF), possibly metabolic driven, were detected uniformly over all membrane surface. Morphological and dynamical parameters of ruffles, such as shape, dimensions, and velocity of propagation, were analyzed. Optical tweezers were used to promote phagocytosis of single Leishmania amazonensis amastigotes by selected macrophages. Analysis of ruffling activity on the macrophages before and during phagocytosis of the parasites indicated that increased ruffling response near forming phagosomes, most likely induced by the parasite, accelerates phagocytosis. The effects of temperature decrease on the dynamics of membrane surface fluctuations and on the phagocytosis of parasites were used to determine the overall activation energies involved in these processes. The values obtained support the existence of strong correlation between membrane motility and phagocytic capacity. PMID- 15652338 TI - Nuclear actin is partially associated with Cajal bodies in human cells in culture and relocates to the nuclear periphery after infection of cells by adenovirus 5. AB - Cajal bodies are intra-nuclear structures enriched in proteins involved in transcription and mRNA processing. In this study, immunofluorescence microscopy experiments using a highly specific antibody to actin revealed nuclear actin spots that colocalized in part with p80 coilin-positive Cajal bodies. Actin remained associated with Cajal bodies in cells extracted to reveal the nuclear matrix. Adenovirus infection, which is known to disassemble Cajal bodies, resulted in loss of actin from these structures late in infection. In infected cells, nuclear actin was observed to relocate to structures at the periphery of the nucleus, inside the nuclear envelope. Based on these findings, it is suggested that actin may play an important role in the organization or function of the Cajal body. PMID- 15652337 TI - Myocilin binding to Hep II domain of fibronectin inhibits cell spreading and incorporation of paxillin into focal adhesions. AB - Myocilin, a novel matricellular protein found in the human eye, can modify signaling events mediated by the Heparin II domain of fibronectin. Using myocilin produced in sf9 insect cells, myocilin inhibited spreading of cycloheximide treated human skin fibroblasts plated on substrates co-coated with myocilin and either fibronectin or its Heparin II domain. Cell spreading could be rescued by adding back either substrate adsorbed or soluble Heparin II domains. Myocilin did not inhibit cell attachment to fibronectin even in the presence of a 2400 M excess of myocilin. Myocilin impaired focal adhesion formation and specifically blocked the incorporation of paxillin, but not vinculin, into focal adhesions. The Heparin II domain mediated the incorporation of paxillin into focal adhesions, since paxillin was not assembled into focal adhesions unless the Heparin II domain was present. The effect of myocilin on focal adhesions could be overcome by treating cells with either phorbol 12-myristate (PMA) or oleoyl-L alpha-lysophosphatidic acid (LPA). Myocilin bound to the fibroblast cell surface, but its binding could not be competed with excess fibronectin, suggesting that myocilin does not compete for cell surface binding sites of fibronectin. Myocilin therefore appears to specifically block functions mediated by the Heparin II domain possibly through direct interactions with it. PMID- 15652340 TI - Growth of intestinal epithelium in organ culture is dependent on EGF signalling. AB - Differentiation of endoderm into intestinal epithelium is initiated at E13.5 of mouse development when there are significant changes in morphology resulting in the conversion of undifferentiated stratified epithelium into a mature epithelial monolayer. Here we demonstrate that monolayer formation is associated with the selective apoptosis of superficial cells lining the lumen while cell proliferation is progressively restricted to cells adjacent to the basement membrane. We describe an innovative embryonic gut culture system that maintains the three-dimensional architecture of gut and in which these processes are recapitulated in vitro. Explants taken from specific regions of the gut and placed into organ culture develop and express molecular markers (Cdx1, Cdx2 and A33 antigen) in the same spatial and temporal pattern observed in vivo indicating that regional specification is maintained. Inhibition of the epidermal growth factor receptor (EGFR) tyrosine kinase using the specific inhibitor AG1478 significantly reduced the proliferation and survival of cells within the epithelial cell layer of cultured gut explants. This demonstrates an essential role for the EGF signalling pathway during the early stages of intestinal development. PMID- 15652339 TI - Nuclear accumulation of epidermal growth factor receptor and acceleration of G1/S stage by Epstein-Barr-encoded oncoprotein latent membrane protein 1. AB - Epstein-Barr virus (EBV)-encoded latent membrane protein 1 (LMP1) is considered to be the major oncogenic protein of EBV-encoded proteins and has always been the core of the oncogenic mechanism of EBV. Advanced studies on nuclear translocation of the epidermal growth factor receptor (EGFR) family have greatly improved our knowledge of the biological function of cell surface receptors. In this study, we used the Tet-on LMP1 HNE2 cell line as a cell model, which is a dual-stable LMP1 integrated nasopharyngeal carcinoma (NPC) cell line and the expression of LMP1 which could be regulated by the Tet system. We found that LMP1 could regulate the nuclear accumulation of EGFR in a dose-dependent manner quantitatively and qualitatively. We also demonstrated that the nuclear localization sequence of EGFR played some roles in the location of the protein within the nucleus under LMP1 regulation and EGFR in the nucleus could bind to the promoters of cyclinD1 and cyclinE, respectively. We further demonstrated that EGFR is involved in the acceleration of the G1/S phase transition by LMP1 through binding to cyclinD1 and cyclinE directly. These findings provided a novel view that the acceleration of LMP1 on the G1/S transition via the nuclear accumulation of EGFR was critical in the process of nasopharyngeal carcinoma. PMID- 15652341 TI - BNIP-2 induces cell elongation and membrane protrusions by interacting with Cdc42 via a unique Cdc42-binding motif within its BNIP-2 and Cdc42GAP homology domain. AB - The Cdc42 small GTPase regulates cytoskeletal reorganization and cell morphological changes that result in cellular extensions, migration, or cytokinesis. We previously showed that BNIP-2 interacted with Cdc42 and its cognate inactivator, p50RhoGAP/Cdc42GAP via its BNIP-2 and Cdc42GAP homology (BCH) domain, but its cellular and physiological roles still remain unclear. We report here that following transient expression of BNIP-2 in various cells, the expressed protein was located in irregular spots throughout the cytoplasm and concentrated at the leading edge of cellular extensions. The induced cell elongation and membrane protrusions required an intact BCH domain and were variously inhibited by coexpression of dominant negative mutants of Cdc42 (completely inhibited), Rac1 (partially inhibited), and RhoA (least inhibited). Presence of the Cdc42/Rac1 interactive binding (CRIB) motif alone as the dominant negative mutant of p21-activated kinase also inhibited the BNIP-2 effect. Bioinformatic analyses together with progressive deletional mutagenesis and binding studies revealed that a distal part of the BNIP-2 BCH domain contained a sequence with low homology to CRIB motif. However, in contrary to most effectors, BNIP-2 binding to Cdc42 was mediated exclusively via the unique sequence motif 285VPMEYVGI292. Cells expressing the BNIP-2 mutants devoid of this motif or/and the 34-amino acids immediately upstream to this sequence failed to elicit cell elongation and membrane protrusions despite that the protein still remained in the cytoplasm and interacted with Cdc42GAP. Evidence is presented where BNIP-2 in vivo induces cell dynamics by recruiting Cdc42 via its BCH domain, thus providing a novel mechanism for regulating Cdc42 signaling pathway. PMID- 15652342 TI - Endogenous N-cadherin in a subpopulation of MDCK cells: distribution and catenin complex composition. AB - Epithelial (E)-cadherin plays a critical role in developing a normal epithelial phenotype but neural (N)-cadherin can disrupt epithelial shape, at least in carcinoma-derived cells. Here the normal epithelial cell line MDCK was used to select for a trypsin-sensitive (TS-MDCK) subpopulation that expresses low levels of endogenous N-cadherin. Similar amounts of E-cadherin and all catenins are found in both TS-MDCK and trypsin-resistant cells (TR-MDCK), but TS-MDCK are less phenotypically epithelioid and more motile, and junctional proteins are more detergent soluble. In TS-MDCK, N-cadherin is largely nonjunctional; a similar N cadherin distribution and mesenchymal phenotype are found in TR-MDCK transfected to express low levels of exogenous N-cadherin. Little N-cadherin was attracted to junctions between TS-MDCK and hTERT-RPE1 cells, a retinal pigment epithelium derived line that expresses dominantly N-cadherin. No differences were seen in E cadherin-catenin complexes in TS- and TR-MDCK, but N-cadherin-catenin complexes in TS-MDCK have more abundant p120 catenin. Overall, the results indicate that E- and N-cadherin assemble stoichiometrically different complexes with p120 in the same cells. Further, N-cadherin does not participate with E-cadherin in a zonular epithelial junction in normal MDCK epithelial cells. Rather, even low levels of endogenous N-cadherin contribute to a disrupted epithelial phenotype, resembling the effect of N-cadherin on carcinoma-derived epithelial cells. PMID- 15652343 TI - Recruitment of mRNA-destabilizing protein TIS11 to stress granules is mediated by its zinc finger domain. AB - TIS11, a member of the CCCH zinc finger protein family, was found to be distributed throughout cells with a preferential cytoplasmic localization when transiently expressed in COS-7 cells. Upon treatment with heat shock, TIS11 became localized in discrete particles in the cytoplasm of the transfectants. We showed the TIS11-positive particles to be stress granules (SGs), which are known to be formed in the cytoplasm of eukaryotic cells in response to environmental stresses. By deletion studies using the green fluorescent protein fusion system, we mapped a functional stress granule (SG) localization signal to a region containing two tandem repeats of the zinc finger motif of TIS11. Site-directed mutations of Tyr105/Tyr113, Gly109/Gly 114, and Phe119 in the first zinc finger motif diminished the ability of this TIS11 domain to direct SG localization. Importantly, when the zinc-chelating Cys residues in either the first or second zinc finger were mutated to Ala residues, the recruitment of the TIS11 zinc finger region to SG was significantly inhibited by the mutation and was completely abolished by the mutation in both zinc fingers. These results suggest that recruitment of TIS11 to heat shock-induced SG is governed by the tandem zinc finger domains of this protein. PMID- 15652344 TI - Saposin C-LBPA interaction in late-endosomes/lysosomes. AB - Acidic phospholipids and saposins associations are involved in the degradation process of glycosphingolipids/sphingolipids in late endosomes/lysosomes. In this report, we showed the colocalization of saposin C and lysobisphosphatidic acid (LBPA) in human fibroblasts by using cytoimmunofluorescence analysis. This colocalization pattern was not seen with other saposins. Large numbers of saposins A, B, and D illustrated the staining patterns that differ from LBPA. In addition, ingested anti-LBPA antibody altered the location of saposin C in human wild-type fibroblasts. In vitro assays demonstrated that saposin C at nM concentrations induced membrane fusion of LBPA containing phospholipid vesicles. Under the same condition, other saposins had no fusion induction on these vesicles. These results suggested a specific interaction between saposin C and LBPA. Total saposin-deficient fibroblasts showed a massive accumulation of multivesicular bodies (MVBs) by electron microscopic analysis. No significant increase of MVBs was found in saposins A and B deficient cells. Interestingly, the accumulated MVBs were significantly reduced by loading saposin C alone into the total saposin-deficient cells. Therefore, we propose that saposin C-LBPA interaction plays a role in the regulation of MVB formation in cells. PMID- 15652345 TI - Proliferative effects of apical, but not basal, matrix metalloproteinase-7 activity in polarized MDCK cells. AB - Matrix metalloproteinase-7 (MMP-7) is primarily expressed in glandular epithelium. Therefore, its mechanism of action may be influenced by its regulated vectorial release to either the apical and/or basolateral compartments, where it would act on its various substrates. To gain a better understanding of where MMP 7 is released in polarized epithelium, we have analyzed its pattern of secretion in polarized MDCK cells expressing stably transfected human MMP-7 (MDCK-MMP-7), and HCA-7 and Caco2 human colon cancer cell lines. In all cell lines, latent MMP 7 was secreted to both cellular compartments, but was 1.5- to 3-fold more abundant in the basolateral compartment as compared to the apical. However, studies in the MDCK system demonstrated that MMP-7 activity was 2-fold greater in the apical compartment of MDCK-MMP-7(HIGH)-polarized monolayers, which suggests the apical co-release of an MMP-7 activator. In functional assays, MMP-7 over expression increased cell saturation density as a result of increased cell proliferation with no effect on apoptosis. Apical MMP-7 activity was shown to be responsible for the proliferative effect, which occurred, as demonstrated by media transfer experiments, through cleavage of an apical substrate and not through the generation of a soluble factor. Taken together, our findings demonstrate the importance of MMP-7 secretion in relation to its mechanism of action when expressed in a polarized epithelium. PMID- 15652346 TI - The p38 SAPK pathway is required for Ha-ras induced in vitro invasion of NIH3T3 cells. AB - Constitutive activation of the ras oncoprotein plays a critical role in cancer invasion and metastasis. Particularly, ras-related protease expression such as the serine protease urokinase plasminogen activator (u-PA) has been implicated in mediating cancer cell invasion. Previous studies have shown that ras-mediated u PA expression is regulated through the mitogen- (MAPK) and stress-activated protein kinase (SAPK) signal transduction pathways extracellular signal-regulated kinase (ERK) and c-Jun-activating kinase (JNK). We therefore asked the question, if ras-related cell invasion might additionally require the third MAPK/SAPK signal transduction cascade, p38. Indeed, we found that ras induces invasion based on the activation of certain p38 protein kinase isoforms, in particular, p38alpha. Moreover, ras activation through transient or stable expression of a Ha rasEJ mutant induced the expression of u-PA. This was found to be a consequence of an increase of u-PA m-RNA, which was paralleled by only a modest activation of the u-PA promoter. In conclusion, we provide evidence for the requirement of a novel ras-p38alpha-u-PA pathway for ras-dependent cellular invasion. PMID- 15652347 TI - GCP-2/CXCL6 synergizes with other endothelial cell-derived chemokines in neutrophil mobilization and is associated with angiogenesis in gastrointestinal tumors. AB - The precise role of chemokines in neovascularization during inflammation or tumor growth is not yet fully understood. We show here that the chemokines granulocyte chemotactic protein-2 (GCP-2/CXCL6), interleukin-8 (IL-8/CXCL8), and monocyte chemotactic protein-1 (MCP-1/CCL2) are co-induced in microvascular endothelial cells after stimulation with pro-inflammatory stimuli. In contrast with its weak proliferative effect on endothelial cells, GCP-2 synergized with MCP-1 in neutrophil chemotaxis. This synergy may represent a mechanism for tumor development and metastasis by providing efficient leukocyte infiltration in the absence of exogenous immune modulators. To mimic endothelial cell-derived GCP-2 in vivo, GCP-2 was intravenously injected and shown to provoke a dose-dependent systemic response, composed of an immediate granulopenia, followed by a profound granulocytosis. By immunohistochemistry, GCP-2 was further shown to be expressed by endothelial cells from human patients with gastrointestinal (GI) malignancies. GCP-2 staining correlated with leukocyte infiltration into the tumor and with the expression of the matrix metalloproteinase-9 (MMP-9/gelatinase B). Together with previous findings, these data suggest that the production of GCP-2 by endothelial cells within the tumor can contribute to tumor development through neovascularization due to endothelial cell chemotaxis and to tumor cell invasion and metastasis by attracting and activating neutrophils loaded with proteases that promote matrix degradation. PMID- 15652348 TI - dlk acts as a negative regulator of Notch1 activation through interactions with specific EGF-like repeats. AB - The protein dlk, encoded by the Dlk1 gene, belongs to the Notch epidermal growth factor (EGF)-like family of receptors and ligands, which participate in cell fate decisions during development. The molecular mechanisms by which dlk regulates cell differentiation remain unknown. By using the yeast two-hybrid system, we found that dlk interacts with Notch1 in a specific manner. Moreover, by using luciferase as a reporter gene under the control of a CSL/RBP-Jk/CBF-1-dependent promoter in the dlk-negative, Notch1-positive Balb/c 14 cell line, we found that addition of synthetic dlk EGF-like peptides to the culture medium or forced expression of dlk decreases endogenous Notch activity. Furthermore, the expression of the gene Hes-1, a target for Notch1 activation, diminishes in confluent Balb/c14 cells transfected with an expression construct encoding for the extracellular EGF-like region of dlk. The expression of Dlk1 and Notch1 increases in 3T3-L1 cells maintained in a confluent state for several days, which is associated with a concomitant decrease in Hes-1 expression. On the other hand, the decrease of Dlk1 expression in 3T3-L1 cells by antisense cDNA transfection is associated with an increase in Hes-1 expression. These results suggest that dlk functionally interacts in vivo with Notch1, which may lead to the regulation of differentiation processes modulated by Notch1 activation and signaling, including adipogenesis. PMID- 15652349 TI - Side population cells isolated from different tissues share transcriptome signatures and express tissue-specific markers. AB - Side Population (SP) cells, isolated from murine adult bone marrow (BM) based on the exclusion of the DNA dye Hoechst 33342, exhibit potent hematopoietic stem cell (HSC) activity when compared to Main Population (MP) cells. Furthermore, SP cells derived from murine skeletal muscle exhibit both hematopoietic and myogenic potential in vivo. The multipotential capacity of SP cells isolated from variable tissues is supported by an increasing number of studies. To investigate whether the SP phenotype is associated with a unique transcriptional profile, we characterized gene expression of SP cells isolated from two biologically distinct tissues, bone marrow and muscle. Comparison of SP cells with differentiated MP cells within a tissue revealed that SP cells are in an active transcriptional and translational status and underexpress genes reflecting tissue-specific functions. Direct comparison of gene expression of SP cells isolated from different tissues identified genes common to SP cells as well as genes specific to SP cells within a particular tissue and further define a muscle and bone marrow environment. This study reports gene expression of muscle SP cells, common features and differences between SP cells isolated from muscle and bone marrow, and further identifies common signaling pathways that might regulate SP cell functions. PMID- 15652350 TI - CIR, a corepressor of CBF1, binds to PAP-1 and effects alternative splicing. AB - We have reported that PAP-1, a product of a causative gene for autosomal retinitis pigmentosa, plays a role in splicing. In this study, CIR, a protein originally identified as a CBF1-interacting protein and reported to act as a transcriptional corepressor, was identified as a PAP-1 binding protein and its function as a splicing factor was investigated. In addition to a basic lysine and acidic serine-rich (BA) domain and a zinc knuckle-like motif, CIR has an arginine/serine dipeptide repeat (RS) domain in its C terminal region. The RS domain has been reported to be present in the superfamily of SR proteins, which are involved in splicing reactions. We generated CIR mutants with deletions of each BA and RS domain and studied their subcellular localizations and interactions with PAP-1 and other SR proteins, including SC35, SF2/ASF, and U2AF35. CIR was found to interact with U2AF35 through the BA domain, with SC35 and SF2/ASF through the RS domain, and with PAP-1 outside the BA domain in vivo and in vitro. CIR was found to be colocalized with SC35 and PAP-1 in nuclear speckles. Then the effect of CIR on splicing was investigated using the E1a minigene as a reporter in HeLa cells. Ectopic expression of CIR with the E1a minigene changed the ratio of spliced isoforms of E1a that were produced by alternative selection of 5'-splice sites. These results indicate that CIR is a member of the family of SR-related proteins and that CIR plays a role in splicing regulation. PMID- 15652351 TI - Nesprin-1alpha contributes to the targeting of mAKAP to the cardiac myocyte nuclear envelope. AB - Muscle A-kinase anchoring protein (mAKAP) is a scaffold protein found principally at the nuclear envelope of striated myocytes. mAKAP maintains a complex consisting of multiple signal transduction molecules including the cAMP-dependent protein kinase A, the ryanodine receptor calcium release channel, phosphodiesterase type 4D3, and protein phosphatase 2A. By an unknown mechanism, a domain containing spectrin repeats is responsible for targeting mAKAP to the nuclear envelope. We now demonstrate that the integral membrane protein nesprin 1alpha serves as a receptor for mAKAP on the nuclear envelope in cardiac myocytes. Nesprin-1alpha is inserted into the nuclear envelope by a conserved, C terminal, klarsicht-related transmembrane domain and forms homodimers by the binding of an amino-terminal spectrin repeat domain. Through the direct binding of the nesprin-1alpha amino-terminal dimerization domain to the third mAKAP spectrin repeat, nesprin-1alpha targets mAKAP to the nuclear envelope. In turn, overexpression of these spectrin repeat domains in myocytes can displace mAKAP from nesprin-1alpha. PMID- 15652353 TI - Changes in spatial and temporal localization of Dictyostelium homologues of TRAP1 and GRP94 revealed by immunoelectron microscopy. AB - TRAP1 (tumor necrosis factor receptor-associated protein 1) is a member of the molecular chaperone HSP90 (90-kDa heat shock protein) family. In this study, we mainly examined the behavior of Dictyostelium TRAP1 homologue, Dd-TRAP1, during Dictyostelium development by immunoelectron microscopy. In vegetatively growing D. discoideum Ax-2 cells, Dd-TRAP1 locates in nucleolus and vesicles in addition to the cell cortex including cell membrane. Many of Dd-TRAP1 molecules moved to the mitochondrial matrix in response to differentiation, although Dd-TRAP1 on the cell membrane seems to be retained. Some Dd-TRAP1 was also found to be secreted to locate outside the cell membrane in Ax-2 cells starved for 6 h. At the multicellular slug stage, Dd-TRAP1 was primarily located in mitochondria and cell membrane in both prestalk and prespore cells. More importantly, in differentiating prespore cells, a significant number of Dd-TRAP1 locates in the PSV (prespore-specific vacuole) that is a sole cell type-specific organelle and essential for spore wall formation, whereas some Dd-TRAP1 in the cell cortical region of prestalk cells. These findings strongly suggest the importance of Dd TRAP1 regulated temporally and spatially during Dictyostelium development. Incidentally, we also have certified that the glucose-regulated protein 94 (Dd GRP94) is predominantly located in Golgi vesicles and cisternae, followed by its colocalization with Dd-TRAP1 in the PSV. PMID- 15652352 TI - Regulation of hyaluronan binding by F-actin and colocalization of CD44 and phosphorylated ezrin/radixin/moesin (ERM) proteins in myeloid cells. AB - Proinflammatory cytokines such as TNF-alpha up-regulate the expression of the cell adhesion molecule, CD44, and induce hyaluronan (HA) binding in peripheral blood monocytes (PBM). Here we show that in PBM, TNF-alpha induced cytoskeletal rearrangement, increased threonine phosphorylation of ERM proteins, and induced the redistribution and colocalization of phospho-ERM proteins (P-ERM) with CD44. In the myeloid progenitor cell line, KG1a, hyaluronan binding occurred in the pseudopod where CD44, P-ERM, and F-actin were highly localized. Hyaluronan binding correlated with high expression of both CD44 and P-ERM clustered in a single pseudopod. Disruption of polymerized actin reduced hyaluronan binding in both PBM and KG1a cells and abolished CD44 clustering and the pseudopod in KG1a cells. The pseudopod was not required for the clustering of CD44, the colocalization with P-ERM, or hyaluronan binding. However, treatment with a kinase inhibitor abolished ERM phosphorylation and reduced hyaluronan binding. Furthermore, expression of CD44 lacking the putative ERM binding site resulted in reduced hyaluronan binding. Taken together, these data suggest that CD44-mediated hyaluronan binding in human myeloid cells is regulated by P-ERM and the actin cytoskeleton. PMID- 15652354 TI - Involvement of the TRAP-1 homologue, Dd-TRAP1, in spore differentiation during Dictyostelium development. AB - Tumor necrosis factor receptor-associated protein 1 (TRAP1) is a member of the molecular chaperone HSP90 (90-kDa heat shock protein) family. We have previously demonstrated that Dictyostelium discoideum TRAP1 (Dd-TRAP1) synthesized at the vegetative growth phase is retained during the whole course of D. discoideum development, and that at the multicellular slug stage, it is located in prespore specific vacuoles (PSVs) of prespore cells as well as in the cell membrane and mitochondria. Thereupon, we examined the function of Dd-TRAP1 in prepore and spore differentiation, using Dd-TRAP1-knockdown cells (TRAP1-RNAi cells) produced by the RNA interference method. As was expected, Dd-TRAP1 contained in the PSV was found to be exocytosed during sporulation to constitute the outer-most layer of the spore cell wall. In the TRAP1-RNAi cells, PSV formation and therefore prespore differentiation were significantly impaired, particularly under a heat stress condition. Although the TRAP1-RNAi cells formed apparently normal-shaped spores with a cellulosic wall, the spores were less resistant to heat and detergent treatments, as compared with those of parental MB35 cells derived from Ax-2 cells. These findings strongly suggest that Dd-TRAP1 may be closely involved in late development including spore differentiation, as well as in early development as realized by its induction of prestarvation response. PMID- 15652355 TI - Detection of mitochondrial DNA depletion in living human cells using PicoGreen staining. AB - Human mitochondria DNA (mtDNA) is arranged within the mitochondria into discrete DNA-protein complexes, termed nucleoids. The size of the human mitochondrial genome is less than that of yeast and is more difficult to visualise by fluorescent DNA stains such as DAPI and Hoescht. We have developed a simple yet effective method to visualise mtDNA in situ within living cells using the fluorescent stain PicoGreen. Quantitative analysis shows that PicoGreen can be used to estimate the degree of mtDNA depletion within living cells. We have used this approach to study the arrangement and fluorescence of nucleoids in cells depleted of mtDNA by treatment with the anti-viral nucleoside analogue, 2',3' dideoxycytidine. We also studied the distribution of mtDNA in fibroblasts cultured from patients with mitochondrial disease. Combining PicoGreen staining with histochemical and immunocytochemical approaches enabled us to examine the effects of mtDNA depletion on mtDNA-related components at the level of single cells. This method is able to detect an intermediate degree of mtDNA depletion in living cells, and can be used to detect mtDNA free cells (rho0 cells) in culture even at very low numbers. We have also adapted the technique to efficiently sort rho0 cells from populations of normal cells by fluorescent-assisted cell sorting (FACS), without the need for selection of respiratory competence. This should be useful for the construction of new trans-mitochondrial 'cybrid' cell lines. PMID- 15652356 TI - HIF-2alpha expression in human fetal paraganglia and neuroblastoma: relation to sympathetic differentiation, glucose deficiency, and hypoxia. AB - Solid tumors are frequently necrotic and hypoxic due to poor vascularization. Tumor cells adapt to hypoxia by modulating their phenotype. Key players in this process are the hypoxia-inducible factors (HIF-1alpha to 3alpha). HIFs are also expressed during normal development; for example, HIF-2alpha is specifically expressed and appears to be involved in the development of the murine sympathetic nervous system (SNS). Here, we demonstrate that HIF-2alpha protein is selectively present in human fetal week 8.5 SNS paraganglia. Neuroblastoma is derived from SNS precursors. In a subset of neuroblastomas, a spontaneous neuronal to neuroendocrine differentiation occurs in areas adjacent to necrotic zones. As HIF 2alpha activity has been associated not only with hypoxic but also with hypoglycemic conditions, we have investigated putative effects of hypoxia, glucose depletion, and HIF-2alpha on the neuroblastoma phenotype. HIF-2alpha was detected in hypoxic and in well-oxygenized neuroblastoma cells and tissue, presumably reflecting their embryonic features. With regard to differentiation, hypoxic cells lost their neuronal/neuroendocrine features and gained marker gene expression associated with an immature, neural crest-like phenotype. Low glucose potentiated the effect of hypoxia. These findings suggest that poorly vascularized neuroblastomas become immature and maintain a more aggressive phenotype, which possibly could involve a sustained stabilization and activation of HIF-2alpha. PMID- 15652357 TI - Identification of MAGI-3 as a transforming growth factor-alpha tail binding protein. AB - The cytoplasmic domain of the transforming growth factor-alpha precursor (proTGFalpha) contains a C-terminal PSD-95/SAP90, Discs Large, and Zona Occludens 1 (PDZ) recognition motif (TVV). By yeast two-hybrid screening of a mouse embryo library, we have found that a third member of a family of PDZ-containing proteins, membrane associated guanylate kinase inverted-3 (MAGI-3), binds to TGFalpha's TVV. MAGI-3 is widely expressed in multiple mouse tissues, including brain. Immunolocalization showed that MAGI-3 and TGFalpha were colocalized in neurons in the cortex and dentate gyrus, as well as in ependymal cells and some astrocytes. In vitro, proTGFalpha bound the PDZ-1 domain of MAGI-3 and MAGI-2, but not MAGI-1. MAGI-3 and the 17-kDa cell surface form of proTGFalpha interact transiently in MDCK cells stably transfected with both MAGI-3 and human proTGFalpha cDNAs. MAGI-3 and wild-type proTGFalpha colocalize at the cell surface. In contrast, MAGI-3 forms a stable complex with membrane-fixed TGFalpha early in the secretory pathway and interacts with immature and cell surface forms of membrane-fixed TGFalpha. Overexpression of MAGI-3 resulted in increased levels of TGFalpha in the basolateral medium of polarized MDCK cells, suggesting that MAGI-3 has a role in efficient trafficking of TGFalpha to the cell surface in polarized epithelial cells. PMID- 15652358 TI - S100A6 is a negative regulator of the induction of cardiac genes by trophic stimuli in cultured rat myocytes. AB - S100A6 (calcyclin), a member of the S100 family of EF-hand Ca2+ binding proteins, has been implicated in the regulation of cell growth and proliferation. We have previously shown that S100B, another member of the S100 family, is induced postinfarction and limits the hypertrophic response of surviving cardiac myocytes. We presently report that S100A6 expression is also increased in the periinfarct zone of rat heart postinfarction and in cultured neonatal rat myocytes by treatment with several trophic agents, including platelet-derived growth factor (PDGF), the alpha1-adrenergic agonist phenylephrine (PE), and angiotensin II (AII). Cotransfection of S100A6 in cultured neonatal rat cardiac myocytes inhibits induction of the cardiac fetal gene promoters skeletal alpha actin (skACT) and beta-myosin heavy chain (beta-MHC) by PDGF, PE, AII, and the prostaglandin F2alpha (PGF2alpha), induction of the S100B promoter by PE, and induction of the alpha-MHC promoter by triiodothyronine (T3). By contrast, S100B cotransfection selectively inhibited only PE induction of skACT and beta-MHC promoters. Fluorescence microscopy demonstrated overlapping intracellular distribution of S100B and S100A6 in transfected myocytes and in postinfarct myocardium but heterodimerization of the two proteins could not be detected by co immunoprecipitation. We conclude that S100A6 may function as a global negative modulator of differentiated cardiac gene expression comparable to its putative role in cell cycle progression of dividing cells. PMID- 15652359 TI - The human ubiquitin-conjugating enzyme Cdc34 controls cellular proliferation through regulation of p27Kip1 protein levels. AB - Ubiquitin-mediated degradation of the cyclin-dependent kinase inhibitor p27Kip1 was shown to be required for the activation of key cyclin-dependent kinases, thereby triggering the onset of DNA replication and cell cycle progression. Although the SCFSkp2 ubiquitin ligase has been reported to mediate p27Kip1 degradation, the nature of the human ubiquitin-conjugating enzyme involved in this process has not yet been determined at the cellular level. Here, we show that antisense oligonucleotides targeting the human ubiquitin-conjugating enzyme Cdc34 downregulate its expression, inhibit the degradation of p27Kip1, and prevent cellular proliferation. Elevation of p27Kip1 protein level is found to be the sole requirement for the inhibition of cellular proliferation induced upon downregulation of Cdc34. Indeed, reducing the expression of p27Kip1 with a specific antisense oligonucleotide is sufficient to reverse the anti proliferative phenotype elicited by the Cdc34 antisense. Furthermore, downregulation of Cdc34 is found to specifically increase the abundance of the SCFSkp2) ubiquitin ligase substrate p27Kip1, but has no concomitant effect on the level of IkBalpha and beta-catenin, which are known substrates of a closely related SCF ligase. PMID- 15652360 TI - Laminin isoforms: biological roles and effects on the intracellular distribution of nuclear proteins in intestinal epithelial cells. AB - Laminins are structurally and functionally major components of the extracellular matrix. Four isoforms of laminins (laminin-1, -2, -5 and -10) are expressed in a specific pattern along the crypt-villus axis of the intestine. Previous works indicated that expression of these isoforms is developmentally regulated and that laminins could modulate the behaviour of intestinal cells, but the exact role of each isoform remained unclear. Here, we report the first systematic analysis of the cellular functions of the four isoforms using the human colon adenocarcinoma Caco2/TC7 cell line as a model. We compared the respective abilities of each isoform to modulate adhesion, proliferation and differentiation of intestinal epithelial cells. We found that the isoforms were functionally distinct, with laminin-10 being the most adhesive substratum, laminin-2, laminin-5 and laminin 10 enhancing cellular proliferation and at the opposite, laminin-1 stimulating intestinal cell differentiation. To begin to characterise the molecular events induced by the different isoforms, we examined by immunofluorescence the intracellular distribution of several nuclear proteins, recently highlighted by a nuclear proteomic approach. We observed clear nucleocytoplasmic redistribution of these proteins, which depended on the laminin isoform. These results provide evidence for a distinct functional role of laminins in intestinal cell functions characterised by specific localisation of nuclear proteins. PMID- 15652361 TI - Anticonvulsant and neurotoxicity evaluation of some 6-substituted benzothiazolyl 2-thiosemicarbazones. AB - Various 6-substituted benzothiazolyl-2-thiosemicarbazones were synthesized and screened for anticonvulsant activity in maximal electroshock induced seizure (MES) and subcutaneous pentylenetetrazole (scPTZ) induced seizure models in mice. The neurotoxicity was assessed using the rotorod method. The 6-methyl benzothiazolyl-2-thiosemicarbazones showed anticonvulsant activity in both mice i.p. and rat oral MES screen. The 6-nitro benzothiazolyl thiosemicarbazone derivative 1a emerged as the most promising one with anti-MES activity in mice i.p., rat i.p. and rat p.o. evaluations. All the compounds exhibited lesser or no neurotoxicity compared to phenytoin. The isatinimino derivatives had shown better activity when compared to the benzylidene or acetophenone derivatives. PMID- 15652362 TI - Synthesis and biological evaluation of new phenidone analogues as potential dual cyclooxygenase (COX-1 and COX-2) and human lipoxygenase (5-LOX) inhibitors. AB - A new series of potential human 5-LOX inhibitors structurally related to the 1 phenyl-3-pyrazolidinone (phenidone, 2) has been synthesized and the activity against COX-1, COX-2, and human 5-LOX enzymes has been evaluated. In contrast with literature data, we observed that phenidone resulted to be inactive against human 5-LOX, while retains its activity against cyclooxygenases in a micromolar range. The present results suggest that the substitution of the amino function at the 4-position is detrimental in terms of activity toward COX-1 and COX-2, while the presence of a double bond at the 4,5-position does not alter the biological profile against COX. The absence of activity vs. human 5-LOX strongly suggests a re-consideration of phenidone and its analogs as 5-LOX inhibitors in humans. PMID- 15652363 TI - Synthesis and analgesic action of N-(substituted-ethyl)pyrrole-3,4 dicarboximides. AB - We prepared a series of new N-[2-(4-substitutedpiperazin-1-yl)ethyl]-1-(n-butyl or phenyl)-2,5-dimethyl-3,4-pyrroledicarboximides 3 and the related products 4 and 5, nine representatives of which were evaluated as potential analgesic agents in an animal model (mice). The new pyrroledicarboximides were not toxic (LD50 > or = 1466 mg/kg) and eight of them displayed analgesic activity approximately 1.5 5 times superior to that of ASA in the writhing test. However, the compounds were found to be unstable in methanol solution and in dilute bases (methanol/NaOMe). The S-A relationship is discussed. PMID- 15652364 TI - Synthesis and antibacterial activity of 4-benzoyl-1-methyl-5-phenyl-1H-pyrazole-3 carboxylic acid and derivatives. AB - Some new 1H-pyrazole-3-carboxylic acid and pyridazinone derivatives were synthesized and evaluated for their antibacterial activities against Bacillus cereus ATCC 7064, Staphylococcus aureus ATCC 6538, Escherichia coli ATCC 4230 and Pseudomonas putida using tube dilution method. The minimal inhibitory concentrations (MICs) experiments revealed that all chemical compounds showed inhibitor effects on the growth of the test microorganisms. Moreover, the results of this research showed that the compound named as 5c was the best compound in the series, exhibiting antibacterial activity against both Gram-positive and Gram negative bacteria. PMID- 15652365 TI - A new approach for preparing a controlled release ketoprofen tablets by using beeswax. AB - Solid lipid ketoprofen micropellets (SLKM) at different drug/beeswax ratios [(1:1) and (1:2)] were prepared by emulsion congealing technique and then compressed into tablets. Ketoprofen in solid state was incorporated into the melted beeswax at 90 degrees C and the mixture was emulsified in the hot aqueous Tween 80 solution by stirring at a constant rate. The SLKM were obtained by cooling the coarse emulsion down to room temperature and filtering. Drug entrapment efficiency and particle size analysis by laser diffractometry (LD) were determined, and existence of a drug-lipid interaction was investigated by differential scanning calorimetry (DSC) on the SLKM, before being compressed into the tablets by direct compression method. Finally, in vitro release studies were performed and the release kinetics of the waxy tablets were calculated. A commercial ketoprofen retard tablet (reference: Profenid Retard 200 mg) was also examined to compare the release properties. While the data obtained from DSC were indicating absence of drug-lipid interaction in the SLKM, it was determined that 28.62% (+/-2.08), 38.60% (+/-1.91) and 47.00% (+/-1.82) of ketoprofen was released from the tablets containing (1:2) and (1:1) SLKM and Profenid Retard 200 mg in pH 7.4 phosphate buffer solution after 8 h, respectively. PMID- 15652366 TI - FT-IR spectrophotometric analysis of dehydroepiandrosterone and its pharmaceutical formulation. AB - A Fourier transform infrared (FT-IR) spectrometric method was developed for the rapid, direct measurement of dehydropeiandrosterone. Conventional KBr spectra and KBr + 2.0 mg microcrystalline cellulose (MCC) spectra were compared for best determination of active substance in drug formulation. Two chemometric approaches, partial least-squares (PLS) and principal component regression (PCR+) methods were used in data processing. The best results were obtained with PCR+ method. PMID- 15652367 TI - Simultaneous determination of Fe(II) and Fe(III) in pharmaceutical formulations with chromogenic mixed reagent by using principal component artificial neural network and multivariate calibration. AB - The use of chemometric approaches for the simultaneous determination of Fe(II) and Fe(III) ions has been explored by means of a two component reagent. Mixed reagents of 1,10-phenanthroline and thiocyanate were used as a selective chromogenic system for speciation of Fe(II) and Fe(III). Although the complexes of Fe(II) and Fe(III) with mixed reagent show a spectral overlap, they have been simultaneously determined with chemometric approaches, such as principal component artificial neural network (PC-ANN), principal component regression (PCR) and partial least squares (PLS). A set of synthetic mixtures of Fe(II) and Fe(III) was evaluated and the results obtained by the applications of these chemometric approaches were discussed and compared. It was found that the PC-ANN and PLS methods afforded better precision relatively than its of PCR. PC-ANN and PLS methods were also applied satisfactorily in determination of Fe(II) and Fe(III) in pharmaceutical samples. PMID- 15652368 TI - Differential pulse voltammetric determination of carvedilol in tablets dosage form using glassy carbon electrode. AB - The electrochemical oxidation of antihypertensive drug carvedilol has been studied in pH range 2.0-11.0 at a glassy carbon electrode by cyclic and differential pulse voltammetry. Two oxidation processes were produced in different supporting electrolyte media. Both oxidation processes were irreversible and diffusion-controlled. The first oxidation process was chosen for the analysis of carvedilol. A very resolved voltammetric peak was obtained in Britton-Robinson buffer at pH 8.0 using differential pulse mode. The linear response was obtained in the range of 0.25-10.00 microg ml(-1). The limit of detection was found to be 0.10 microg ml(-1). The developed method was used for the determination of carvedilol in tablet dosage form. PMID- 15652369 TI - 2,2'-Bipyridine as a new and sensitive spectrophotometric reagent for the determination of nanoamounts of certain dibenzazepine class of tricyclic antidepressant drugs. AB - 2,2'-bipyridine is proposed as new and sensitive spectrophotometric reagent for the determination of certain dibenzazepine class of tricyclic antidepressants. The spectrophotometric method is based on the reaction of imipramine hydrochloride (IPH), desipramine hydrochloride (DPH), clomipramine hydrochloride (CPH), trimipramine maleate (TPM) and opipramol (OPP) with iron (III) and subsequent reaction with 2,2'-bipyridine in an acetic acid medium to yield a pink color with maximum absorption at 530 nm. The color developed was stable over 3-4 h at room temperature (approximately 27 degrees C). The commonly encountered excipients and additives did not interfere with the determination. Results from the analysis of pure drugs and commercial tablets agreed well with those of the official method (United States Pharmacopoeia, 24, USP Convention, Rockville 2000, pp. 505-506, 865-867.). PMID- 15652370 TI - Investigations on the synthesis and pharmacological properties of N-substituted derivatives of 4-alkoxy-6-methyl-1H-pyrrolo[3,4-c]pyridine-1,3(2H)-diones. AB - Synthesis of N-substituted derivatives of 4-alkoxy-6-methyl-1H-pyrrolo[3,4 c]pyridine-1,3(2H)-diones (10-14, 18-21) is described. In the "writhing syndrome" test all compounds studied exhibited potent analgesic activity which was superior than that of acetylsalicylic acid. In the "hot plate" test imides 10, 12, 13, 18 20 acted also stronger than aspirin. Furthermore all compounds tested significantly suppressed the spontaneous locomotor activity of mice and prolonged barbiturate sleep of these animals. PMID- 15652371 TI - Extractive spectrophotometric methods for the determination of doxepin hydrochloride in pharmaceutical preparations using titanium (IV) and iron (III) thiocyanate complexes. AB - Two simple, precise, and accurate extractive spectrophotometric methods have been developed for the determination of doxepin hydrochloride in pharmaceutical preparations. The methods are based on the formation of ion association complexes of doxepin with titanium (IV) and iron (III) thiocyanate complexes in acidic medium. The produced compounds are insoluble in water but well soluble in some organic solvents. They are extracted with mixtures of butyl alcohol-chloroform (2:3, v/v) and (1:4, v/v) and measured spectrophotometrically at 400 and 490 nm for DOX-Ti-SCN and DOX-Fe(III)-SCN methods, respectively. Beer's law was obeyed in the concentration ranges of 5-50 and 3-30 microg/ml with molar absorptivity of 7.12 x 10(3) and 1.36 x 10(4) l mol(-1) cm(-1) for DOX-Ti-SCN and DOX-Fe-SCN systems, respectively. The proposed methods have been successfully applied for the analysis of the drug in dosage forms. No interference was observed from common pharmaceutical adjuvants. The methods have been also used for the determination of the drug in the presence of its degradation product. Statistical comparison of the obtained results with the reference methods shows excellent agreement and indicates no significant difference in accuracy and precision. PMID- 15652372 TI - The properties of slightly soluble Zn(II)-protein complexes in the form of a suspension. AB - The properties of slightly soluble Zn(II)-protein complexes in the form of a suspension. Slightly soluble Zn(II) complexes with selected hormones were obtained in the form of a suspension. Zn(II)-protein complexes were obtained at different ligand concentrations. The amount of Zn(II) in microM bound to a protein increases along with an increase in the added ligand concentration. This amount is statistically significant as far as its binding to INS is concerned. Proteins bind to Zn(II) through two classes of binding sites (n). Most Zn(II) binds to albumin and least to LH and FSH. The protein molecular weight and number of its amino acid residues have an influence on binding Zn(II) to a selected protein. Zn(II) has the highest affinity for PRL and INS, and the lowest for albumin and FSH. The kinetics analysis of Zn(II) binding to proteins has shown that Zn(II) creates the most stable complexes with PRL, INS, and LH. Relatively, weak complexes were obtained with albumin and FSH. The protein molecular weight and the number of its amino acid residues have a significant influence on the stability of Zn(II)-protein complex. The lower protein molecular weight and number of its amino acid residues, the more stable Zn(II)-protein complex is. The stability of complexes is also associated with the amount of Zn(II) bound to a protein and its affinity for a protein. PMID- 15652373 TI - Micellar electrokinetic chromatography for determination of drug partition in phospholipids. AB - The lipophilicity of pipemidic, nalidixic and oxolinic acids was determined by forming phospholipidic micelles directly in an electrophoretic capillary. Phosphatidylcholine derivatives, namely L-alpha-dilauroyl phosphatidylcholine (DLPC) or L-alpha-dimiristoyl phosphatidylcholine (DMPC), were added in the run buffer (50 mM phosphate buffer at pH 7.4). To obtain a mixed micelle, phospholipidic derivatives and sodium cholate were together added in the run buffer. Considering the increasing of migration time when phosphatidylcholine derivative is added in the run buffer, Ks can be determined and then quinolones lipophilicity. PMID- 15652374 TI - Development and validation of a capillary electrophoresis method for the determination of codeine, diphenhydramine, ephedrine and noscapine in pharmaceuticals. AB - The present work describes a simple, accurate and rapid method for the separation and simultaneous determination of codeine, diphenhydramine, ephedrine and noscapine present in cough-cold syrup formulations by capillary zone electrophoresis. Factors affecting the separation were the buffer pH and concentration, applied voltage, and presence of additives. Separations were carried out in less than 10 min with a 20 mM sodium tetraborate buffer, pH 8.50. The carrier electrolyte gave baseline separation with good resolution, great reproducibility and accuracy. Calibration plots were linear over at least three orders of magnitude of analyte concentrations, the lower limits of detection being within the range 0.42-1.33 microg ml(-1). Detection was performed by UV absorbance at wavelengths of 205 and 250 nm. Quantification of the components in actual syrup formulations was calculated against the responses of freshly prepared external standard solutions. The method was validated and met all analysis requirements of quality assurance and quality control. The procedure was fast and reliable and commercial pharmaceuticals could be analyzed without prior sample clean-up procedure. PMID- 15652375 TI - Estradiol valerate and intake of sweetened water. AB - Recently, it has been shown that female rats receiving very large doses (e.g., 2 mg) of estradiol valerate (EV) take considerably more alcoholic beverage than placebo controls. The question asked, with these procedures, is whether the enhanced appetite for alcoholic beverages was specific to those beverages or was a reflection of a general increase in appetite. Female rats were provided with various sweetened beverages. In one experiment, they were provided a palatable saccharin solution (0.25% solution) and a less palatable one (2% saccharin solution). EV treatment led to more intake of the palatable saccharin solution and reduced intake of the less palatable solution. EV induces changes leading to enhanced appetite for some ingesta (including palatable saccharin solutions and alcoholic beverages), but surely not all ingesta. PMID- 15652376 TI - Recovery from experimental Parkinson's disease in the 1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine hydrochloride treated marmoset with the melatonin analogue ML 23. AB - A new mechanism has been recently proposed, whereby melatonin may participate in the ongoing process of neuronal degeneration in models of neurodegenerative disorders, such as Parkinson's disease (PD). Antagonism of the melatonin receptor in rats using constant light or pinealectomy induced recovery and reduced the mortality typically associated with dopamine (DA) degeneration. In additional studies, employing ML-23 in the 6-OHDA-treated rat, remission from experimental PD was achieved using this drug in a post 6-OHDA treatment regime. To permit the further assessment of ML-23 as a potential clinical candidate for the treatment of PD, the present study was undertaken to determine the efficacy of ML-23 in the 1-methyl-4-phenyl, 1,2,3,6 tetrahydropyridine (MPTP) model in the common marmoset. ML-23 was administered orally in a dose of 3 mg/kg twice daily to half of the animals, while the other half received vehicle only, in a blinded protocol, for 56 days. The effects of the treatment on positive and negative features of MPTP-induced PD were assessed, including horizontal and vertical movement, head checking, general behaviour and Parkinsonian condition, raisin board performance, the ability to remove a foot label, palatable and dry food intake, water consumption, bradykinaesia, and the positive symptoms of tremor, obstinate progression, and agitation. On all parameters, ML-23 produced a significant remission from MPTP-induced Parkinsonism, and this effect did not abate when ML-23 treatment was withdrawn. In a further pilot study involving a crossover of two animals, one animal treated previously with MPTP plus vehicle showed some remission of negative and positive features, although ML-23 treatment was not commenced until 8 weeks post-MPTP. Conversely, a recurrence of Parkinsonian signs was not observed when ML-23 treatment was withdrawn and substituted with oral vehicle. Dopamine transporter was severely impaired in all marmosets treated with ML-23 or vehicle for the duration of the study. These results suggest that a novel mechanism involving melatonin is involved in the primary aetiology of the chronic aspects of PD, and such a mechanism is not related to the antioxidative function of this hormone. From these preliminary results, it is concluded that ML-23 and other melatonin analogues have an important role to play in the treatment and clinical management of Parkinson's disease. PMID- 15652377 TI - Gender differences in modulatory effects of tamoxifen upon the nigrostriatal dopaminergic system. AB - It has been demonstrated that the gonadal steroid hormone estrogen can exert neuroprotective effects upon the nigrostriatal dopaminergic (NSDA) system against methamphetamine (MA)-induced neurotoxicity in female, but not male, mice. In contrast, the anti-estrogen, tamoxifen (TMX) can function as a NSDA neuroprotectant within both female and male mice. In an attempt to understand these effects of TMX, the effects of this anti-estrogen upon both behavioral and neurochemical indices of NSDA function were examined within female and male mice following treatment with MA. In general, TMX exerted markedly different (bi directional) effects upon NSDA function between female and male mice. Notably, treatment with TMX resulted in a relative decrease in striatal dopamine (DA) and 3,4-dihydroxyphenylacetic acid (DOPAC) concentrations within male mice and a relative increase in female mice when treated with MA to produce a significant gender difference. Similar effects were obtained for locomotor behaviors related with NSDA function. That is, TMX produced increases in horizontal activity, number of movements and total distance traveled within MA-treated female mice resulting in statistically significant gender differences for the two former parameters. For non-locomotor behaviors, like time occupying the center and margin of the cage, TMX-treated male mice showed statistically significant increases and decreases compared within TMX-treated female mice, respectively. These results show that in contrast to the similar neuroprotective effects of TMX upon MA-induced NSDA neurotoxicity, a number of other NSDA indices induced by MA show markedly different response profiles between TMX-treated female and male mice. PMID- 15652378 TI - Delta-9-tetrahydrocannabinol differentially suppresses emesis versus enhanced locomotor activity produced by chemically diverse dopamine D2/D3 receptor agonists in the least shrew (Cryptotis parva). AB - The principal psychoactive component of marijuana, delta-9-tetrahydrocannabinol (Delta9-THC), suppresses nausea and vomiting in cancer patients caused by chemotherapeutics such as cisplatin. Cisplatin induces vomiting via a number of emetic stimuli, including dopamine. Currently, there is controversy as to whether Delta9-THC can prevent emesis produced by dopaminergic agonists such as apomorphine. The present investigation utilizes the least shrew to evaluate the antiemetic potential and the cannabinoid receptor by which Delta9-THC may prevent emesis produced by four dopamine receptor agonists with differing selectivity for D2 and D3 receptors, i.e., a nonselective dopamine receptor agonist (apomorphine), a D2-preferring receptor agonist (quinpirole), and two D3 preferring receptor agonists (quinelorane and 7-OH DPAT). In addition, relative to its antiemetic doses, the motor suppressive doses of Delta9-THC in dopamine D2/D3-receptor-agonist-treated shrews were also evaluated. Thus, different groups of shrews were injected with either vehicle (V) or varying doses of Delta9-THC [0.5, 1, 2.5, 5, or 10 mg/kg, intraperitoneal (i.p.)] 10 min prior to administration of a 2 mg/kg dose of one of the four cited D2/D3 agonists. Immediately after the last injection, the frequency of vomiting for each shrew was recorded for the next 30 min. To investigate which cannabinoid receptor is involved in the antiemetic action of Delta9-THC, various doses of the CB1 receptor antagonist SR 141716A [0, 5, 10, and 20 mg/kg, subcutaneous (s.c.)] were administered to shrews 10 min prior to an injection of a fully effective antiemetic dose of Delta9-THC (5 mg/kg, i.p.). Ten minutes later, each treated shrew was administered with a 2 mg/kg dose of apomorphine. The emesis frequency was recorded for the next 30 min. For locomotor studies, different groups of shrews received either vehicle or various doses of Delta9-THC (0, 5, 10, 20, or 30 mg/kg) 10 min prior to an injection of vehicle or a 2 mg/kg dose of one of the four D2/D3 receptor agonists. The triad of motor behaviors (spontaneous locomotor activity, total duration of movement, and rearing frequency) were recorded for the next 30 min by a computerized video tracking system. Delta9-THC dose dependently attenuated the frequency of emesis as well as fully protecting shrews from vomiting produced by each one of the four cited dopamine D2/D3 receptor agonists with ID50s ranging from 1 to 4 mg/kg. SR 141716A reversed the antiemetic activity of Delta9-THC against apomorphine-induced emesis. Delta9-THC also differentially suppressed the triad of motor activities in dopamine D2/D3 receptor-agonist-treated shrews with ID50s ranging from 7 to 21 mg/kg. The results suggest that Delta9-THC prevents emesis via cannabinoid CB1 receptors in a potent and dose-dependent manner in D2/D3-receptor-agonist-treated shrews at doses well below those which cause significant motor depression. PMID- 15652379 TI - Thiolic antioxidant supplementation of the diet reverses age-related behavioural dysfunction in prematurely ageing mice. AB - We have studied in a model of premature ageing in mice based on their impaired behavioural response in a simple T-maze test the effect of the ingestion of thioproline (TP) plus N-acetylcysteine (NAC) (0.1% w/w of each antioxidant) by female and male mice of Swiss and BALB/c strains on performance in two behaviour tests. The antioxidant treatment (4 weeks in two different periods of life, i.e., adult and old age) protected all animals against early-age-associated behavioural impairment, but this improvement was more evident in the prematurely ageing mice (PAM) in comparison to the control group or non-prematurely ageing mice (NPAM). An improvement of the exploratory activity and neuromuscular coordination after the thiolic antioxidant treatment was found in the PAM, bringing the behavioural parameters to the NPAM levels. These effects could be due to the glutathione precursor role of NAC and TP that replenish the intracellular reduced glutathione (GSH) levels despite advancing age. In conclusion, diet supplementation with thiolic compounds appears to be an effective therapy for protection against early behavioural decline in prematurely ageing mice. PMID- 15652380 TI - Effects of acetylcholinesterase and butyrylcholinesterase inhibition on breathing in mice adapted or not to reduced acetylcholinesterase. AB - We investigated the contributions of acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) inhibition to the respiratory dysfunction produced by organophosphates in mice which were adapted or not to low AChE activity. Effects of acute selective inhibition of AChE and BChE on ventilation measured by whole-body plethysmography were compared in mice with either normal AChE activity (wild-type), or mice adapted to a null AChE activity (homozygotes for AChE gene deletion) or adapted to an intermediate level of activity (heterozygotes). In wild-type mice acute reduction of AChE by Huperzine A (1 mg/kg) to the level found in asymptomatic heterozygotes, induced tremors but no respiratory depression, whereas the same dose of Huperzine in heterozygote animals further reduced AChE activity, increased tidal volume (V(T)) and decreased breathing frequency (f(R)). A lethal dose of Huperzine in wild-type mice augmented these respiratory effects, but was ineffective in homozygotes. BChE inhibition by bambuterol was ineffective in wild-type mice and heterozygotes, decreased V(T) in homozygotes adapted to null AChE activity but increased V(T) in wild-type mice acutely treated with Huperzine, also aggravating the cholinergic syndrome. We conclude that: (1) Huperzine does not perturb respiration at a dose inhibiting 40% of AChE, and at a lethal dose does not affect any other enzyme important for respiration; (2) Respiratory function is more sensitive to anticholinesterases in heterozygotes than in wild-type mice; (3) BChE may play distinct roles in respiratory function, because its inhibition has opposite effects on tidal volume depending on whether the mouse has adapted to null AChE or whether AChE has been lowered acutely; (4) BChE inhibition may contribute to the respiratory toxicity of organophosphates. PMID- 15652381 TI - Inhibition of PKC in basolateral amygdala and posterior parietal cortex impairs consolidation of inhibitory avoidance memory. AB - Hippocampal alpha- and betaI/betaII protein kinase C (PKC) are crucial for the formation of different types of memory in several species, including that for a one trial inhibitory avoidance (IA) task in rats. Many studies, however, have shown that other brain structures besides the hippocampus, notably the basolateral amygdala (BLA) and posterior parietal cortex (PC) are also necessary for memory consolidation. Here, we examine the role of alpha- and betaI/betaII PKC in the BLA and PC on the consolidation of the memory for IA in rats. The selective inhibitor of alpha- and betaI/betaII-PKC Go 6976 and the nonselective PKC inhibitor Go 7874 were administered into these structures at different times after training at concentrations known to inhibit PKC and to produce retrograde amnesia when given into the hippocampus. Go 7874 blocked consolidation of IA memory when infused into BLA immediately and 30 min or into PC 180 to 360 min posttraining. Go 6976 caused amnesia when given into the BLA also immediately or 30 min posttraining but in the PC hindered memory retention only when infused 270 and 360 min after the training session. Our data indicate that alpha- and betaI/betaII-PKC are critical for consolidation of IA memory shortly after training in BLA and that, first other isoforms and subsequently the alpha- and betaI/betaII PKC are required 3 or more hours after training in the PC. The findings on BLA are similar to those previously reported in the hippocampus, but those on PC suggest an entirely different molecular dynamics for memory formation in that area. PMID- 15652382 TI - Involvement of potassium channels and nitric oxide in tramadol antinociception. AB - It has been considered that tramadol, a centrally acting analgesic, shows its effect via opiatergic, noradrenergic, and serotonergic systems. It has a low affinity for opioid receptors, and its effect can be partly blocked by naloxone. Since the noradrenergic and serotonergic mechanisms are still unknown, other systems which are associated with pain and analgesia may have a role on the antinociceptive effect of tramadol. The aim of this study was to evaluate the effects of K+ channels and nitrergic systems on the antinociceptive action of tramadol. The antinociceptive effects of tramadol were determined in mice by the hot plate test. To examine the effects of K+ channels and the nitrergic system nonspecific voltage-dependent K+ channel blockers 4-aminopyridine (4-AP) and tetraethylammonium (TEA), nitric oxide (NO) precursor L-arginine, and the NO synthase (NOS) inhibitor N(G)-nitro-L-arginine methyl ester (L-NAME) were used. Our results indicated that 4-AP, TEA, and L-arginine reduced the antinociceptive effect of tramadol. However, L-NAME augmented the antinociceptive effect of tramadol. The reduction of the effects of tramadol by L-arginine was reversed by L-NAME. The results of our study suggest that nonspecific voltage-dependent K+ channels and nitrergic system have a role on the antinociceptive effect of tramadol in mice hot plate test. PMID- 15652384 TI - Differential behavioral responses to nicotine in Lewis and Fischer-344 rats. AB - Individual and strain variability in the effects of nicotine suggests the involvement of a genetic component in nicotinic cholinergic receptor (nAChR) function, which may help explain nicotine's variable behavioral and pharmacological effects in different individuals. The present study evaluated differential responses to the discriminative stimulus (DS) and rewarding properties of nicotine in Lewis (LEW) and Fischer-344 (F-344) rats. Drug discrimination (DD) data suggest that the LEW rat is more sensitive to nicotine as LEW rats acquired the nicotine discrimination at a dose of 0.4 mg/kg, whereas F-344 rats acquired the dose of 0.9 mg/kg (all nicotine doses expressed as free base). Similarly, LEW rats exhibited nicotine-conditioned place preference (CPP) at 0.6 mg/kg, whereas the F-344 rats did not. Subsequent testing with a higher dose (0.9 mg/kg) failed to maintain the nicotine-CPP in the LEW rats. Conversely, nicotine-place preference in the F-344 rats was not changed at the higher dose. Taken together, these results suggest potential differences of sensitivities in LEW and F-344 rats to the rewarding and discriminative stimulus (DS) properties of nicotine. These findings support previous research by demonstrating that the F 344 rat is less sensitive to nicotine compared to the LEW rat. PMID- 15652383 TI - Chronic morphine increases the pituitary-adrenocortical response of juvenile rats to mild stress. AB - We previously reported that chronic exposure of adult male rats to morphine by pellet implantation has no effect on corticosterone secretion but causes a marked testosterone-dependent increase in CBG. In the studies reported here, we examined the effects of chronic morphine on the pituitary-adrenocortical axis of male rats prior to the developmental rise in testosterone. In contrast to adults, morphine had little effect on CBG in peripubertal males. We found nothing remarkable with regard to basal hormone levels; morphine caused only a transient increase in ACTH and corticosterone in juveniles. However, while the pituitary-adrenocortical response to mild stress was normal in adults exposed to morphine, it was markedly increased in juveniles. After 7 days of morphine exposure, the stress response was as much as 2.5 times greater than normal in morphine-treated juveniles. This exaggerated response to stress did not appear to be due to the passive withdrawal of morphine or to an additive effect of stress plus morphine. Instead, morphine may either increase the perceived severity of stressors or decrease sensitivity to the negative feedback effects of stress levels of corticosterone in juvenile males. Either way, there is a striking shift in morphine's effects on the pituitary-adrenocortical axis across development. PMID- 15652385 TI - Comparison of five different rat models of peripheral nerve injury. AB - Described here is a comparison of five peripheral sciatic nerve injury models in rats which all result in various degrees of neuropathic pain symptoms. They are the chronic constriction injury (CCI), the spinal nerve ligation (SNL), the partial sciatic ligation (PSL), the tibial and sural transection (TST), and the complete sciatic transection (CST) model. Behavioral testing was performed on these models over a 56-day period under strict experimental conditions to minimise variability between the surgical models and to allow for an accurate evaluation of the sensory deficits produced by each model. Overall, all five models of neuropathic pain produced signs of allodynia and hyperalgesia to various stimuli. However, the duration and magnitude of the evoked responses varied considerably between the different models. PMID- 15652386 TI - Accurate caloric compensation in rats for electively consumed ethanol-beer or ethanol-polycose mixtures. AB - High elective intake of ethanol was achieved in rats by presenting ethanol in palatable vehicles. We simultaneously measured intake of food (chow) to assess the accuracy of caloric compensation for the energy in the alcoholic commodity. In the first study, we used beer; nonalcoholic beer was consumed in large amounts, and when 5% or 10% ethanol was added, intake amounted to approximately 10% of daily calories. In the second study, Polycose in either solution or a gel matrix was used as the palatable vehicle for ethanol. The intake of ethanol was even higher than in the beer study, particularly in the gel preparation. In all cases, both male and female rats showed accurate caloric compensation by a reduction in chow intake. In a final study, we showed that restricted time access to the Polycose-alcohol gel produced high elective intakes and substantial blood alcohol levels. Over 24 h, caloric compensation was again accurate. Thus, unlike some reports in humans, rats seem able to compensate accurately for alcohol calories and in particular when, as with most alcohol consumption by humans, these are presented in palatable vehicles. PMID- 15652387 TI - The behavioral thermoregulatory response of febrile female rats is not attenuated by vagotomy. AB - An oft-overlooked consequence of fever is the occurrence of thermoregulatory heat seeking/producing behaviors. Subdiaphragmatic vagotomy attenuates fever resulting from low dose, peripherally administered pyrogens, suggesting that the vagus is involved in generating the pathogen-induced rise in core body temperature (T(c)). This study was designed to confirm that rats utilize behavioral thermoregulation to augment fever following systemic administration of lipopolysaccharide (LPS), and to test the hypothesis that, in febrile animals, vagotomy would block the preference for a higher ambient temperature (T(a)) as T(c) is rising. First, female Sprague-Dawley rats received IP injections of either saline or LPS (50 microg/kg), prior to placement inside a thermal gradient that offered subjects T(a) values between 7 and 45 degrees C. LPS injection caused significant increases in T(c) and selection of a higher T(a) as compared to saline administration. Second, groups of rats were vagotomized, sham-vagotomized or received no surgery, and then underwent the same gradient testing procedure. Vagotomy attenuated LPS-induced fever, but did not influence the concomitant behavioral thermoregulatory response. All groups selected comparable, higher T(a) values following LPS vs. saline. These data suggest that the reduction in the febrile response to LPS administration following vagotomy is not due to inhibition of the behavioral thermoregulatory response to the pyrogen. Rather, this behavioral response to LPS appears to be mediated by a nonvagal mechanism. PMID- 15652389 TI - Knock down of the alpha 5 nicotinic acetylcholine receptor in spinal nerve ligated rats alleviates mechanical allodynia. AB - Nicotinic acetylcholine receptor (nAChR) agonists are known to alleviate neuropathic and inflammatory pain via activation of a heterogeneous population of receptors. However, the function of nAChRs in the maintenance of neuropathic pain is not known. Spinal nerve ligation (SNL) increases the spinal expression of the alpha5 nAChR subunit ipsilateral to injury. The alpha5 subunit is unique because it modifies numerous characteristics of existing functional nAChRs, but it does not form functional nAChRs when expressed alone or with beta nicotinic subunits. Because there are no alpha5 subunit selective ligands, we used antisense oligonucleotides (ODNs) to assess the contribution of the alpha5 subunit to the maintenance of mechanical allodynia following SNL. Intrathecal antisense oligonucleotides were administered to SNL rats after the development of mechanical allodynia (10-12 days post-SNL). I.t. antisense specifically reduced alpha5 immunoreactivity (alpha5-IR) by 50-70% in the outer laminae of the dorsal horn and moderately alleviated mechanical allodynia. Furthermore, using the phosphorylation of cAMP response element-binding protein (pCREB) as a general marker of neuronal activation, a significant increase in pCREB immunoreactivity was observed in SNL rats. Treatment of SNL rats with alpha5-antisense significantly reduced pCREB immunoreactivity. These results suggest that the increased expression of the alpha5 nAChR subunit following SNL contributes to spinal CREB phosphorylation and the maintenance of mechanical allodynia. PMID- 15652388 TI - Evidence for Pavlovian conditioning of cocaine-induced responses linked to emotional behavioral effects. AB - The pairing of cocaine treatments with a specific test environment typically leads to cocaine-conditioned drug effects. In this study, we first pre-exposed rats 10 times to an open-field environment to establish an habituation asymptote in locomotor activity prior to the initiation of cocaine treatments. Two groups (N=10) equated for locomotion, grooming, central zone penetrations and rearing behavior were used. One group received five pairings of cocaine (10.0 mg/kg) and the second group five pairings of saline injections with placements in the open field environment. Subsequently, both groups received a saline test to detect possible cocaine-conditioned behavioral effects. During the cocaine treatment phase, cocaine enhanced locomotion and central zone penetrations but decreased rearing and grooming. On the conditioning test, the cocaine group exhibited enhanced central zone penetrations and decreased grooming as compared to the saline group. There were no group differences in locomotion or rearing. When within group comparisons were performed between behavioral responses on the pre conditioning test vs. the conditioning test, the saline group scores were essentially unchanged. In contrast, the cocaine group exhibited higher central zone penetrations and decreased grooming without changes in locomotion or rearing. In that a cocaine conditioning test can also be viewed as a cocaine withdrawal test, two additional experiments were conducted using an unpaired conditioning protocol to test for withdrawal effects without conditioning. These results indicated that the central zone and grooming effects observed in the conditioning protocol were not withdrawal effects. Altogether, these findings provide support for Pavlovian conditioning of cocaine-induced changes in emotion related behavioral responses. PMID- 15652390 TI - Effects of the cannabinoid receptor agonist WIN 55,212-2 on operant behavior and locomotor activity in rats. AB - Cannabinoids influence the motivational state of a subject and affect motor behavior. In the present study, we examined the acute effects of the cannabinoid (CB) receptor agonist WIN 55,212-2 (WIN) in three different doses (0.6, 1.2 and 1.8 mg/kg) on the performance of rats in a progressive ratio operant behavior task and on locomotor activity. WIN dose-dependently reduced the break point and the total number of lever-presses under a progressive ratio schedule. A food preference test revealed a preference for freely available casein pellets over lab chow in all treatment groups, indicating no WIN-effects on primary motivation. There was a significant reduction in the amount of casein pellets consumed by animals treated with 1.8 mg/kg WIN. Locomotor activity in the open field was increased by 0.6 mg/kg, but not by higher doses of WIN. These data show that administration of the synthetic cannabinoid receptor agonist WIN leads to dose-dependent alterations of the performance in an operant behavior task and of motor behavior. We confirm previous findings of dose-dependent motor stimulating and inhibiting effects of cannabinoids, and show an impairment of a complex operant behavior at higher doses of WIN. PMID- 15652391 TI - Effects of chlordiazepoxide on single-unit activity in the septal region of the freely moving rat: aversive vs. non-aversive contexts. AB - Evidence suggests that stimuli that have the property of inhibiting fear in a Pavlovian fear conditioning paradigm increase cellular activity in the lateral septum, a result consistent with the idea that the lateral septum is actively involved in the inhibition of fear. The experiments reported here were designed to determine if an anxiolytic drug with fear-inhibiting properties would also increase neuronal activity in the lateral septum in a manner that might relate to its mechanism of action as an anxiolytic. An experiment was performed to compare the effects of the benzodiazepine anxiolytic chlordiazepoxide (CDP) upon single unit activity in the septal region of the rat brain during Pavlovian aversive conditioning with the effects of CDP in a non-aversive context. During Pavlovian conditioning there was a decrease in unit activity in the more lateral regions of the septum, the dorsolateral and ventrolateral nuclei, when a stimulus signaling footshock (CS+) was presented. This conditioned suppression of unit activity was blocked by an intraperitoneal injection of CDP. Additionally, CDP increased baseline unit activity in these regions in the absence of conditioned stimuli. In the more medial regions of the septum, the intermediate lateral septum, we observed few consistent changes either to the conditioned stimuli or to the drug. In a non-aversive context CDP had either no effect at low to moderate doses, or a suppressant effect at a higher dose. The results support a fear-relief hypothesis of lateral septal functioning and suggest the lateral septum as a possible site for the anxiolytic action of benzodiazepines. PMID- 15652392 TI - Clonidine pre-treatment fails to block acute smoking-induced EEG arousal/mood in cigarette smokers. AB - Given the arousal eliciting actions of smoking and nicotine, and the contributing role of noradrenaline in brain arousal systems, this study examined the neuroelectric and affective correlates of cigarette smoking following acute pre treatment with the alpha 2-noradrenergic auto-receptor agonist, clonidine. In a double-blind placebo-controlled crossover design, quantitative electroencephalography (EEG), mood, and smoking withdrawal symptoms were assessed in 12 overnight smoking abstinence smokers, before and after sham and cigarette smoking. Orally administered clonidine (0.1 mg) failed to alter overnight smoking abstinence symptoms or the EEG arousal and mood-elevating response seen with the smoking of a single cigarette. The results are discussed in relation to neural mechanisms underlying the acute reinforcement maintaining nicotine use. PMID- 15652393 TI - Patterns of waterpipe use and dependence: implications for intervention development. AB - Despite the dramatic increase of tobacco smoking via waterpipe in Arab societies, and the apparent potential of waterpipe use to produce tobacco-related disease, little is known about the pharmacological effects of this method of tobacco smoking, particularly its ability to support dependence. This review focuses on recent waterpipe research and current theories of dependence in an attempt to identify patterns of waterpipe use and features likely to reveal dependence. Recent work indicates that, relative to cigarette smoking, this form of tobacco use is characterized by more intermittent use, later age of onset, greater spread among women and lower interest in quitting or appreciation of addictive properties. Waterpipe use is associated with classic features of tobacco/nicotine dependence, as well as features unique to this tobacco use method. However, even shared features of dependence, such as craving and addiction-induced socio cognitive behavioral changes, can be displayed differently in waterpipe users, indicating the need for waterpipe-specific research approaches. Preliminary evidence suggests that an important step toward dependence involves a transition from social to individual patterns of waterpipe use. Surveillance and research into factors affecting use and cessation of this tobacco use method should pave the way for the development of effective prevention and intervention strategies to curb the burgeoning waterpipe use epidemic. PMID- 15652394 TI - Water soluble fraction (<10 kDa) from bee venom reduces visceral pain behavior through spinal alpha 2-adrenergic activity in mice. AB - We have previously shown that subcutaneous bee venom (BV) injection reduces visceral pain behavior in mice, but it is not clear which constituent of BV is responsible for its antinociceptive effect. In the present study, we now demonstrate that a water-soluble subfraction of BV (BVA) reproduces the antinociceptive effect of BV in acetic acid-induced visceral pain model. We further evaluated three different BVA subfractions that were separated by molecular weight, and found that only the BVAF3 subfraction (a molecular weight of <10 kDa) produced a significant antinociceptive effect on abdominal stretches and suppressed visceral pain-induced spinal cord Fos expression. Injection of melittin (MEL), a major constituent of BVAF3, also produced a visceral antinociception. However, melittin's antinociception was completely blocked by boiling for 10 min at 100 degrees C, while boiling either whole BV or BVAF3 did not prevent their antinociception. The antinociceptive effect of BVAF3 was completely blocked by intrathecal pretreatment with the alpha2-adrenoceptor antagonist, yohimbine (YOH), while intrathecal pretreatment with the opioid antagonist, naloxone (NAL) or the serotonin antagonist, methysergide, had no effect. These data demonstrate that BVAF3 is responsible for the visceral antinociception of whole BV and further suggest that this effect is mediated in part by spinal alpha2-adrenergic activity. PMID- 15652395 TI - AVP V1b selective antagonist SSR149415 blocks aggressive behaviors in hamsters. AB - Arginine vasopressin (AVP) has been implicated in a variety of physiological and behavioral responses to stress. Synthesis of receptor-selective AVP agonist and antagonist compounds allows differential analysis of the specific roles of particular receptor subtypes with respect to these responses. Here, effects of the recently synthesized AVP V1b selective antagonist, SSR149415, were examined for offensive aggression in male Syrian hamsters, using a resident-intruder paradigm. Oral administration of vehicle or 1, 10, or 30 mg/kg of SSR149415 to resident hamsters was followed by evaluation of a range of aggression-related measures of residents confronted by intruders. The 10 and 30 mg/kg doses significantly reduced the duration of offensive sideways and chase behaviors, and the 30 mg/kg dose also reduced chase frequency. The 10 and 30 mg/kg dose also significantly reduced frequency and duration of olfactory investigation and duration of flank marking. These findings suggest a link between activity of the V1b receptor and the modulation of offensive aggression. These findings agree with previous research on V1b receptor effects in suggesting that antagonism of this receptor may be useful in modulating a range of emotional responses to highly stressful or threatening conditions. PMID- 15652396 TI - Differential effects of morphine and LiCl on schedule-induced polydipsia. AB - Lithium chloride (LiCl) and morphine both produce a conditioned taste avoidance response, while only LiCl is able to elicit a conditioned rejection response (taste reactivity), indicating that the effects of conditioning are drug and preparation dependent. The present experiments extend this assessment to another behavioral preparation, schedule-induced polydipsia (SIP), by examining the ability of LiCl and morphine to produce conditioned suppression of nonregulatory drinking. In Experiment 1, schedule-induced saccharin consumption was followed by LiCl or morphine (at doses comparably effective in conditioning taste avoidance under water deprivation) or by the distilled water vehicle. Although both LiCl and morphine suppressed SIP, morphine produced a significantly weaker suppression than did LiCl. Using a massed feeding design in which animals received all their food pellets in a single meal, Experiment 2 determined that LiCl and morphine were equally effective in suppressing consumption, indicating that the differential effects seen under SIP were due to the schedule of spaced food pellet deliveries. The basis for the differential effects of LiCl and morphine on SIP may be a function of an increase in the reinforcing properties of drugs of abuse (such as morphine) within this procedure that mask the acquisition and/or display of the conditioned suppression. If so, then this procedure may be useful in assessing the reinforcing properties of such drugs. PMID- 15652397 TI - Toll-like receptors on astrocytes: patterning for immunity. PMID- 15652398 TI - Preferential expression and function of Toll-like receptor 3 in human astrocytes. AB - In contrast to other tissues, the central nervous system (CNS) is essentially devoid of MHC expression and shielded from antibodies by the blood-brain barrier. Therefore, a rapid local innate immune response by resident brain cells is required to effectively fight infectious agents. This study analyzed the expression and function of Toll-like receptors (TLRs) in cultured human astrocytes. Quantitative PCR for TLRs 1 to 10 showed a basal expression of TLR3 that could be enhanced by IFN-gamma, IL-1beta, and IFN-beta. The other TLRs were barely detectable and not inducible by the same cytokines. IFN-gamma-activated astrocytes responded to TLR3 ligand poly (I:C) engagement with IL-6 production, while ligands of other TLRs, like LPS, lipoteichoic acid, peptidoglycan, flagellin, and CpG, had no effect. Poly (I:C) also triggered astrocyte production of TNF-alpha and the chemokines CCL2/MCP-1, CCL5/RANTES, CCL20/MIP-3alpha, and CXCL10/IP-10. The adapter molecules MyD88 (full length and short isoform), TIRAP/Mal, and TICAM-1/TRIF, which are required for TLR signaling, were all expressed by astrocytes. Thus, resting and activated human astrocytes express preferentially TLR3 and, upon TLR3 engagement, produce IL-6 and chemokines active on T cells, B cells, monocytes, and dendritic cells. These data indicate that astrocytes function as sentinels for viral infections in the CNS. PMID- 15652399 TI - Evidence for serotonin influencing the thalamic infiltration of mast cells in rat. AB - Serotonin (5-HT) is involved in neuroimmunomodulation. We analyzed the effects of sumatriptan, a 5-HT(1B/1D) receptor agonist, and ondansetron, a 5-HT(3) receptor antagonist, on thalamic mast cell (TMC) population, the only immunocytes known to infiltrate the brain in physiological conditions. Only sumatriptan was effective, significantly increasing TMC numbers versus controls, and especially those containing 5-HT. 5-HT(1B) receptors are concentrated in the median eminence on non-serotonergic axonal endings, probably hypothalamic terminal fibers, involved in hypothalamic-pituitary neuroendocrine modulating processes. TMC variations could reflect serotonergic actions on these fibers. TMCs would thus be cellular interfaces mediating immune action in the nervous system in relation with the hormonal status of the organism. PMID- 15652401 TI - Immunohistochemical detection of the human cytochrome P4507B1: production of a monoclonal antibody after cDNA immunization. AB - The cytochrome P4507B1 (P4507B1) is responsible for the 7alpha-hydroxylation of dehydroepiandrosterone (DHEA) and other 3beta-hydroxysteroids in the brain and other organs. The cDNA of human P4507B1 was used for DNA immunization of mice. The best responding mouse led to the production of monoclonal antibodies (mAbs). The clone D16-37 produced an IgM specific for P4507B1 with no cross-reaction with other human P450s. This antibody permitted the immunohistochemical detection of P4507B1 in slices of human hippocampus. P4507B1 was expressed in neurons only. This new tool will be used for the extensive examination of the P4507B1 presence and determination of its levels in slices of human normal and diseased brain and in other human tissues. PMID- 15652400 TI - Characterization of serotonin transporter in blood lymphocytes of rats. Modulation by in vivo administration of mitogens. AB - Serotonin transporter sites were characterized in blood lymphocytes of rats. Pharmacological characteristics of drug interactions were in concordance with recent studies in nervous and human immune cells. The potency order of inhibition of [(3)H]paroxetine binding was imipramine>citalopram>alaproclate>serotonin. Selective inhibitors of dopamine or noradrenaline transporters did not inhibit it. The specific binding of [(3)H]paroxetine was higher at intermediate than at low concentrations, and the plot of free vs. specific binding had a sigmoid shape. The affinity constant or K(d), 1.77 nM, was in close agreement with data obtained from kinetic studies (K(d)=1.33 nM), which evidences that the equilibrium was reached. In addition, serotonin transporter was evaluated by lipopolysaccharide or concanavalin A administration in vivo (0.1 mg/kg, i.p., 18 h). After the treatment with lipopolysaccharide, no changes were observed in the numbers of sites or B(max) or in the affinity, K(d). The treatment with concanavalin A showed a significant reduction in B(max) and reduction in K(d). Additionally, serotonin and 5-hydroxyindoleacetic acid levels were determined in plasma and lymphocytes by high-performance liquid chromatography. Treatment with lipopolysaccharide produced a significant increased of serotonin levels in lymphocytes without changes in 5-hydroxyindoleacetic acid level; in plasma, it produced an increase in serotonin and 5-hydroxyindolacetic acid levels. In addition, serotonin synthesis was evaluated by adding 300 microM of tryptophan in the medium, which significantly increased serotonin levels in control lymphocytes. Moreover, the concentrations of 5-hydroxyindoleacetic acid was enhanced significantly, both in plasma and lymphocytes in the presence of tryptophan after treatment with lipopolysaccharide. The administration of concanavalin A significantly decreased plasma levels of serotonin, as well as the concentrations of serotonin and 5-hydroxyindoleacetic acid in lymphocytes. These results demonstrate the presence of serotonin transporter in lymphocytes of rat blood, the capacity for serotonin synthesis in lymphocytes, and the modulation of these parameters by systemic administration of mitogens. The findings of this work contribute to understanding the immunological role of serotonin and the communication of immune and nervous systems. PMID- 15652402 TI - Effects of interferon-beta on the cytokine production of astrocytes. AB - The effects of interferon (IFN)-beta used for treatment of multiple sclerosis (MS) on the central nervous system (CNS) have not yet been elucidated. The effect of IFN-beta and IFN-gamma on astrocytes as the major component of the CNS was investigated using murine primary astrocytes and human astrocytoma U251 cell line. IL-1beta, IL-6, TNF-alpha and iNOS were analyzed by RT-PCR. Both protein and mRNA levels of IL-6 were increased by IFN-beta. IFN-gamma augmented the effect of IFN-beta on IL-6 production. These results suggest that IFN-beta has an activity to modulate inflammatory and immune responses by up-regulating IL-6 in the CNS of MS. PMID- 15652403 TI - Targeting fibroblast growth factor-inducible-14 signaling protects from chronic relapsing experimental autoimmune encephalomyelitis. AB - The TNF-related weak inducer of apoptosis (TWEAK) is a TNF family member mediating proinflammatory effects by its receptor fibroblast growth factor inducible-14 (Fn14). We studied the role of TWEAK/Fn14 in experimental autoimmune encephalomyelitis (EAE) by protein vaccination with TWEAK and Fn14 and recombinant TWEAK-DNA, respectively. TWEAK-DNA vaccination worsened the clinical course of EAE and increased central nervous system (CNS) inflammation. TWEAK increased the secretion of CCL2 [monocyte chemotactic protein-1 (MCP-1)] by CNS endothelial cells and astrocytes in vitro, suggesting CCL2 as a critical mediator of TWEAKs proinflammatory effects. Vaccination with the extracellular domain of TWEAK or with Fn14 resulted in the induction of specific inhibitory antibodies and an amelioration of EAE signs in two different models in rats and mice. Spinal cord inflammatory infiltrates were significantly diminished. Purified IgG from TWEAK- or Fn14-vaccinated rats prevented TWEAK-induced production of CCL2 by endothelial cells. Blocking Fn14 signaling represents a novel approach with potential for the treatment of CNS autoimmunity. PMID- 15652404 TI - IL-8 enhancement of amyloid-beta (Abeta 1-42)-induced expression and production of pro-inflammatory cytokines and COX-2 in cultured human microglia. AB - The effects of the chemokine IL-8 on amyloid beta peptide (Abeta(1-42))-induced responses in cultured human microglia were investigated using RT-PCR, ELISA and immunocytochemistry. Abeta(1-42) (5 microM) applied for 8 h induced the expression and increased the production of the pro-inflammatory cytokines IL-6, IL-1beta, TNF-alpha, the inducible enzyme COX-2 and chemokine IL-8. Microglial treatment with IL-8 added (at 100 ng/mL) with Abeta(1-42) led to enhancement in both expression and production of all of these pro-inflammatory factors compared with peptide alone. Stimulation with IL-8 itself was effective in increasing microglial expression of pro-inflammatory cytokines, IL-8 and COX-2, however, had no effect on protein levels of all these factors. The expression of the anti inflammatory cytokines IL-10 and TGFbeta(1) remained unchanged from basal levels with stimulation using either Abeta(1-42), IL-8 or the peptide together with IL 8. The actions of IL-8 to potentiate Abeta(1-42)-induced inflammatory mediators may have particular relevance to Alzheimer disease brain which exhibits elevated levels of the chemokine. PMID- 15652405 TI - Cuff electrode implantation around the sciatic nerve is associated with an upregulation of TNF-alpha and TGF-beta 1. AB - Epineurial fibrosis, fiber loss, limited reproducibility of recordings and variability of stimulation conditions have been documented after extraneural cuff electrode implantation. These morphological and electrophysiological modifications could be due to the local release of cytokines. We report the expression of two cytokines, tumor necrosis factor-alpha (TNF-alpha) and transforming growth factor-beta1 (TGF-beta1) in the rat sciatic nerve after 'cuff' implantation for 18 h, 7 days and 1 month. Immunohistochemical and Western blot analyses showed a transient upregulation of TNF-alpha, during the first week, and a prolonged increase of TGF-beta1, over the 1-month period duration of this study. Considering the known pro-inflammatory roles of TNF-alpha and the pro fibrotic action of TGF-beta, our results strongly suggest that these cytokines may contribute to nerve alterations occurring within the acute and sub-acute phases after cuff electrode implantation. PMID- 15652406 TI - Dose-dependent activation of microglial cells by Toll-like receptor agonists alone and in combination. AB - Microglial cells express Toll-like receptors (TLRs) recognising exogenous and endogenous ligands. Upon stimulation with agonists of TLR2, TLR4, and TLR9, nitric oxide (NO) and tumor necrosis factor-alpha (TNF-alpha) were released by primary mouse microglial cell cultures. Endotoxin was most potent in stimulating microglia followed by pneumolysin, cytosine-guanosine (CpG) oligodesoxynucleotide (ODN), and Tripalmitoyl-S-glyceryl-cysteine. Maximum stimulation of TLR2, TLR4, and TLR9 resulted in approximately equal amounts of nitric oxide release. Pneumolysin was a potent activator of microglial cells; at high concentrations, it reduced cell viability. No cytotoxicity was noted with the other TLR agonists. Costimulation with maximum concentrations of two TLR agonists did not further increase nitric oxide release. Costimulation with submaximum concentrations was additive or supraadditive, suggesting that even low concentrations of products of infectious agents can lead to microglial activation via TLRs. PMID- 15652407 TI - The nonpsychoactive component of marijuana cannabidiol modulates chemotaxis and IL-10 and IL-12 production of murine macrophages both in vivo and in vitro. AB - Cannabidiol is the main nonpsychoactive component of marijuana. We examined the ability of in vivo and in vitro cannabidiol to interfere with the production of interleukin (IL)-12 and IL-10 by murine macrophages and to modulate macrophage chemotaxis. Cannabidiol added in vitro to peritoneal macrophages significantly increased IL-12 and decreased IL-10 production. The CB1 and CB2 receptor antagonists prevented this modulation. Macrophages from animals treated with cannabidiol at the dose of 30 mg kg(-1) either orally or i.p. produced higher levels of IL-12 and lower levels of IL-10 in comparison to controls, and the CB receptor antagonists did not prevent these effects. Cannabidiol dose-dependently decreased fMLP-induced chemotaxis of macrophages, and the CB2 receptor antagonist prevented this decrease. PMID- 15652408 TI - Maternal poly I:C exposure during pregnancy regulates TNF alpha, BDNF, and NGF expression in neonatal brain and the maternal-fetal unit of the rat. AB - Maternal infection during pregnancy is associated with increased risk for neurodevelopmental disorders. Polyriboinosinic-polyribocytidilic acid (poly I:C) or saline was administered to rats to model maternal infection; levels of TNFalpha, brain-derived neurotrophic factor (BDNF), and nerve growth factor (NGF) were determined by ELISA. TNFalpha was significantly increased in maternal plasma, placenta, and amniotic fluid, while it was significantly decreased in fetal liver/spleen and neonatal brain. NGF and BDNF were significantly decreased in the placenta and fetal liver/spleen. There was no change in BDNF or NGF in the fetal or neonatal brain. Changes in TNFalpha, BDNF, and NGF after maternal exposure to poly I:C represent a potential mechanism through which maternal infection increases risk for neurodevelopmental disorders. PMID- 15652409 TI - Peritoneal macrophages express the serotonin transporter. AB - Although it is known that macrophages take up serotonin, a specific monoamine transporter has not been identified in macrophages. In this study, mRNA coding for the serotonin transporter (SERT) was detected with the reverse transcription polymerase chain reaction (RT-PCR) in recruited mouse peritoneal macrophages. Sequencing confirmed the identity of the RT-PCR product to mouse SERT mRNA. SERT protein was detected by Western blotting. Macrophage activation with lipopolysaccharide had no effect on expression of SERT mRNA or protein. Consistent with expression of a functional SERT, specific uptake of (3)H serotonin in macrophages was sodium dependent and inhibited by fluoxetine (IC(50) 6.9 nM) and desipramine (IC(50) 32 nM) but not by nisoxetine or reserpine. PMID- 15652410 TI - Acute stress suppresses pro-inflammatory cytokines TNF-alpha and IL-1 beta independent of a catecholamine-driven increase in IL-10 production. AB - Interleukin (IL)-10 is an anti-inflammatory cytokine that can down-regulate various aspects of the immune response. In this study we demonstrate that exposure to a psychophysiological stressor (swim stress) increases IL-10 production in female rats in response to an in vivo challenge with bacterial lipopolysaccharide (LPS). This increase in LPS-induced IL-10 was associated with suppression of the pro-inflammatory cytokines IL-1beta and TNF-alpha, indicating that overall, swim stress promotes an immunosuppressive cytokine phenotype. Despite the well-documented ability of IL-10 to suppress pro-inflammatory cytokine production, neutralisation of IL-10 failed to block the stress-induced suppression of IL-1beta and TNF-alpha. These data indicate that the suppressive effect of swim stress on these pro-inflammatory cytokines occurs independently of increased IL-10 production. To determine if swim stress-induced immunosuppression was mediated by increased sympathetic nervous system activity, and subsequent beta-adrenoceptor activation, we assessed the ability of the beta-adrenoceptor antagonist nadolol to block stressor-induced changes in cytokine production. Whilst pre-treatment with nadolol completely blocked the stress-induced increase in IL-10, it failed to alter the suppression of TNF-alpha or IL-1beta. Similarly, pre-treatment with the glucocorticoid receptor antagonist mifepristone also failed to attenuate the suppressive effect of swim stress on IL-1beta and TNF alpha production. These data indicate that neither increased glucocorticoid secretion, nor catecholamine-induced beta-adrenoceptor activation, mediates the suppressive effect of swim stress on pro-inflammatory cytokine production. Taken together, these data demonstrate a role for beta-adrenoceptor activation in the ability of acute swim stress to increase LPS-induced IL-10 production, and also highlight a mechanistic dissociation between the ability of swim stress to increase IL-10 and suppress pro-inflammatory cytokine production. PMID- 15652411 TI - Effects of phospholipids on antiganglioside antibody reactivity in GBS. AB - Serum antibody activities to mixtures of a ganglioside and various phospholipids were compared with those to a ganglioside alone in 30 anti-GM1 IgG-positive GBS patients and 30 anti-GQ1b IgG-positive Miller Fisher syndrome (MFS) patients. Anti-GM1-positive sera had higher antibody reactivities against a mixture of GM1 and several phospholipids including PA, PI and PS, than against GM1 alone. In contrast, in case of anti-GQ1b antibody, no phospholipid provided significant enhancement. Sphingomyelin provided decrease of the activity for both anti-GM1 and anti-GQ1b IgG. The effects of phospholipids must be considered to determine the pathogenetic role of antiganglioside antibodies in GBS and MFS. PMID- 15652412 TI - Oligoclonal IgG bands in Japanese patients with multiple sclerosis. A comparative study between isoelectric focusing with IgG immunofixation and high-resolution agarose gel electrophoresis. AB - A low prevalence of oligoclonal IgG bands (OB) has been reported as a unique feature of Japanese multiple sclerosis (MS) as compared with Western MS. We compared the frequency of OB between isoelectric focusing (IEF) and agarose gel electrophoresis (AGE) in 59 Japanese patients with clinically definite MS [39 with conventional form of MS (CMS) and 20 with optic-spinal form of MS (OSMS)]. The frequency of IEF-OB in total MS was 54%, whereas that of AGE-OB was only 17%. When OSMS was excluded, the frequency increased to 77% by IEF, whereas it remained 23% by AGE. Our study strongly suggests that IEF is highly effective for detecting OB in Japanese MS patients. PMID- 15652413 TI - Myasthenia gravis patients, but not healthy subjects, recognize epitopes that are unique to the epsilon-subunit of the acetylcholine receptor. AB - Myasthenia gravis (MG) is an autoimmune disease characterized by deficits in neuromuscular transmission due to antibody-mediated damage of the acetylcholine receptor (AChR). We examined the in vitro immune response of peripheral blood mononuclear cells isolated from MG patients (n=38) and healthy nonmyasthenic subjects (n=31) to epitopes on the alpha-, epsilon-, and gamma-chains of the AChR. The epsilon- and gamma-epitopes tested represent regions with little sequence homology to the alpha-chain, and little sequence homology between the epsilon- and gamma-chains. No differences were observed in the immune response of MG patients and healthy subjects to any of the alpha-chain epitopes tested. Serial studies of the immune response to the alpha-peptides suggest that epitope spread does occur over time. Cells from MG patients were stimulated by the epsilon- and gamma-chain peptides, although the response was weaker than that to the alpha-peptides. Cells from healthy subjects showed reactivity to gamma-chain peptides only; none of the healthy subjects responded to the epsilon-chain peptides tested. Differences between the epsilon- and gamma-chains may be important in the development of MG, because only MG patients respond to epitopes that are unique to the epsilon-subunit. PMID- 15652414 TI - The pro and the active form of matrix metalloproteinase-9 is increased in serum of patients with amyotrophic lateral sclerosis. AB - Pro and active-matrix metalloproteinase-9 (MMP-9) was measured in sera from patients with amyotrophic lateral sclerosis (ALS), Guillain-Barre syndome (GBS), and healthy subjects. Both forms of MMP-9 were elevated in sera of ALS and GBS patients, compared with healthy controls. It has been postulated that elevated MMP-9 reflects damage to peripheral nerve and muscle. This possibility was investigated in sera, and tissue extracts of sciatic nerves and muscle from mice 5 and 12 days after axotomy of the sciatic nerve. Pro-MMP-9 was elevated in sera and extracts of damaged nerve and muscle, suggesting such damage may be followed by elevated pro-MM9-9 in sera. Active MMP-9 was only elevated in the sera. However, in situ activation of MMP-9 is tightly regulated and localised, and probably difficult to demonstrate by ELISA, resulting in a short half-life active MMP-9, implying any active MMP-9 in the serum may have a more immediate origin than injured muscle or nerve, for example circulating blood cells. PMID- 15652415 TI - Monocyte-derived HLA-G acts as a strong inhibitor of autologous CD4 T cell activation and is upregulated by interferon-beta in vitro and in vivo: rationale for the therapy of multiple sclerosis. AB - Peripheral antigen presenting cells (APCs) contribute to the maintenance of immune tolerance and are considered to play a critical role in promoting the (re)activation of autoreactive T cells in multiple sclerosis (MS). Interferon beta (IFN-beta) is the principle immune-modulatory agent used in the treatment of MS, but its mechanism of action remains elusive. HLA-G is a non-classical MHC molecule (MHC class Ib) attributed chiefly immune-regulatory functions. We here investigated the role of monocyte-derived HLA-G in the immune-regulatory processes of MS and its implications for current immune-modulatory therapies. Monocytes constitutively express cell surface HLA-G1 and soluble HLA-G5. Comparison of monocytic HLA-G expression between patients with relapsing remitting MS (n=17) and healthy donors (n=20) revealed significantly lower levels of HLA-G1 protein in MS patients. However, both groups showed a significant upregulation of HLA-G in response to IFN-beta in vitro. Serial measurements of HLA-G mRNA levels in MS patients before and during IFN-beta therapy corroborated the relevance of these results in vivo: 1 month after initiation of IFN-beta1b therapy (n=9), HLA-G1 and HLA-G5 were significantly increased compared to baseline levels and remained elevated during treatment for 6 months (n=3). Importantly, functional experiments demonstrated that monocyte-derived HLA-G inhibits both Th1 (IFN-gamma, IL-2) and Th2 (IL-10) cytokine production by antigen-stimulated autologous CD4 T cells. Soluble HLA-G added to antigen specific T cell lines (TCLs) has similar effects on the release of cytokines and reduces T cell proliferation. Although both IFN-beta and IFN-gamma strongly enhance HLA-G1 and HLA-G5 expression by monocytes in vitro, IFN-beta leads to a stronger relative upregulation of HLA-G compared to classical MHC class I molecules than stimulation with IFN-gamma. Taken together, monocyte-derived HLA-G mediates the inhibition of autologous CD4 T cell activation and might be involved in immune-regulatory pathways in the pathogenesis of MS. We conclude that some desirable immune-modulatory effects of INF-beta might be accomplished via the upregulation of the immune-tolerogenic molecule HLA-G. PMID- 15652417 TI - CSF-chemokines in HTLV-I-associated myelopathy: CXCL10 up-regulation and therapeutic effect of interferon-alpha. AB - We measured four chemokines in the cerebrospinal fluid (CSF) in human T lymphotropic virus type I (HTLV-I)-associated myelopathy/tropical spastic paraparesis (HAM/TSP) with ELISA. CXCL10/IP-10, a T cell type 1 (Th1)-associated chemokine, was significantly elevated in HAM/TSP compared with controls, and the values were even significantly higher in HAM/TSP than in multiple sclerosis (MS) in which CXCL10/IP-10 up-regulation was previously reported. Among Th2-associated chemokines, CCL17/TARC and CCL11/Eotaxin in HAM/TSP were not different from those in controls. As shown in MS, CCL2/MCP-1 was significantly lower in HAM/TSP than in control. Following interferon (IFN)-alpha therapy in HAM/TSP, CCL2/MCP-1 became significantly higher than that before therapy, which may reflect a Th2 induction, while CXCL10/IP-10 remained elevated. PMID- 15652416 TI - Dendritic cells in the cerebrospinal fluid and peripheral nerves in Guillain Barre syndrome and chronic inflammatory demyelinating polyradiculoneuropathy. AB - The role of antigen-presenting cells (APC) involved in induction of T and B cell mediated autoaggressive immunity in Guillain-Barre syndrome (GBS) and chronic inflammatory demyelinating polyradiculoneuropathy (CIDP) is poorly understood. We studied the numbers and phenotype of dendritic cells (DC) in blood and cerebrospinal fluid (CSF) over the course of GBS and CIDP before and after immunomodulatory treatment. Four out of seven GBS patients examined prior to treatment with high-dose intravenous immunoglobulins (IvIg) had elevated numbers of CD123(+) plasmacytoid DC in the CSF, while both GBS and CIDP patients examined prior to treatment had elevated numbers of CD11c(+) myeloid DC in the CSF, as compared to patients with noninflammatory neurological diseases (OND). The percentages of blood DC expressing the cell surface marker CD1a, co-stimulatory molecules CD80 and CD86, adhesion molecule CD54, and chemokine receptors CCR1, CCR2, CCR5, and CXCR4 were not affected in GBS or CIDP. The immunohistochemistry of sural nerve biopsies revealed CD11c(+)CD83(-)CD14(-)CD16(-) immature myeloid DC at low numbers, mostly in the perineurium, without difference between CIDP patients and controls. In contrast, the numbers of CD11c(+)CD14(+)/CD16(+) macrophages were higher within the endoneurium in CIDP patients compared with the controls. The recruitment of DC to CSF in GBS and CIDP may be important in capturing antigens released from inflamed spinal nerve roots into CSF and in transferring these antigens from CSF to local lymph nodes, where naive T and B cells may be activated. PMID- 15652418 TI - Specific immunoadsorption of the autoantibodies from myasthenic patients using the extracellular domain of the human muscle acetylcholine receptor alpha subunit. Development of an antigen-specific therapeutic strategy. AB - Antibodies against the acetylcholine receptor (AChR) are the main pathogenic factor in myasthenia gravis (MG). Clinical improvement correlates well with a reduction in levels of circulating anti-AChR antibodies, and plasmapheresis is an efficient short-term MG treatment. The Sepharose-immobilized N-terminal extracellular domain of human muscle AChR alpha-subunit was used to immunoadsorb anti-AChR autoantibodies from 50 MG patients sera. The immunoadsorbents removed 60-94% of the anti-AChR antibodies in 10 sera and a mean of 35% from all samples combined. Immunoadsorption was fast, efficient, and the columns could be used repeatedly without any release or proteolysis of the polypeptide, suggesting the feasibility of antigen-specific MG immunoadsorption therapy. PMID- 15652419 TI - Antibodies against OspA epitopes of Borrelia burgdorferi cross-react with neural tissue. AB - Neurological sequela of chronic Lyme disease include encephalopathy, myelopathy and peripheral neuropathy. These have generally been attributed to either persistent infection or pathogen-induced autoimmunity. In this study, we investigated the presence of cross-reactive human neural epitopes that share amino acid sequences with Borrelia burgdorferi OspA protein. Sequence similarity analysis was carried out by searching known cDNA sequences from brain tissue. The cDNA database search yielded three sequences that were identical to sequences in OspA. Corresponding peptides were synthesized and antibodies were generated against them in rabbits. Antibodies against two of the homologous OspA peptides were found to react with neurons in human brain, spinal cord and dorsal root ganglia by immunohistochemistry. PMID- 15652420 TI - Involvement of p38 MAPK signaling pathway in IFN-gamma and HTLV-I expression in patients with HTLV-I-associated myelopathy/tropical spastic paraparesis. AB - We analyzed the relationship between the expression of interferon (IFN)-gamma and HTLV-I p19 antigen and activation of p38 mitogen-activated protein kinase (p38 MAPK) in two HTLV-I-infected T cell lines derived from two patients (HCT-1 and HCT-4) with HTLV-I-associated myelopathy/tropical spastic paraparesis (HAM/TSP), and three HTLV-I-infected T cell lines derived from three patients with adult T cell leukemia (ATL). Expression of phosphorylated (activated)-p38 MAPK was markedly increased concomitant with high levels of both IFN-gamma and HTLV-I p19 antigen expression in both HCT-1 and HCT-4 compared with cell lines derived from ATL patients. Treatment with SB203580, a specific inhibitor of p38 MAPK, suppressed IFN-gamma and HTLV-I p19 antigen expression levels in HCT-1, HCT-4 and peripheral blood CD4(+) T cells of HAM/TSP patients. These findings strongly suggest that activation of p38 MAPK signaling pathway is involved in the up regulation of IFN-gamma expression with high HTLV-I proviral load in HAM/TSP patients. PMID- 15652421 TI - Gene expression profiling in chronic inflammatory demyelinating polyneuropathy. AB - Gene expression in archived frozen sural nerve biopsies of patients with chronic inflammatory demyelinating polyneuropathy (CIDP) was compared to that in vasculitic nerve biopsies (VAS) and to normal nerve (NN) by DNA microarray technology. Hierarchical clustering analysis demonstrated distinct gene expression patterns distinguishing these disease groups. Of particular interest were: (1) Tachykinin precursor 1, which may be involved in pain mediation; (2) Stearoyl-CoA-desaturase, which may be a marker for remyelination and (3) the Allograft Inflammatory Factor 1 (AIF-1), a modulator of immune response during macrophage activation. Differential gene expression may help distinguish between CIDP, VAS and NN in sural nerve biopsies and identify genes that may be involved in disease pathogenesis. PMID- 15652422 TI - Evidence for systemic immune system alterations in sporadic amyotrophic lateral sclerosis (sALS). AB - Sporadic amyotrophic lateral sclerosis (sALS) is a progressive neuroinflammatory disease of spinal cord motor neurons of unclear etiology. Blood from 38 patients with sALS, 28 aged-match controls, and 25 Alzheimer's disease (AD) patients were evaluated and activated monocyte/macrophages were observed in all patients with sALS and AD; the degree of activation was directly related to the rate of sALS disease progression. Other parameters of T-cell activation and immune globulin levels showed similar disease associated changes. These data are consistent with a disease model previously suggested for AD, wherein systemic immunologic activation plays an active role in sALS. PMID- 15652423 TI - CTLA-4 gene polymorphism is not associated with conventional multiple sclerosis in Japanese. AB - We investigated the polymorphisms of exon 1 (+49A/G) and promoter (-318C/T and 651C/T) regions of the CTLA-4 gene in 133 Japanese patients with conventional/classical multiple sclerosis (MS) and 156 healthy controls. Patients with optico-spinal MS (OSMS) or atypical clinical attacks were excluded from the study. There was no significant difference in the distribution of polymorphisms between patients and controls. Furthermore, there were no associations between polymorphisms and clinical characteristics, such as age at onset, disease prognosis, and HLA profiles. Our results suggest that CTLA-4 gene polymorphisms are neither conclusively related to susceptibility nor to the clinical characteristics of MS, especially in Japanese patients with conventional/classical form and clinical features identical to those of their counterparts in Western countries. PMID- 15652424 TI - HLA and smoking in prediction and prognosis of small cell lung cancer in autoimmune Lambert-Eaton myasthenic syndrome. AB - Patients with small cell lung cancer (SCLC) survive longer if they have the antibody-mediated Lambert-Eaton myasthenic syndrome (LEMS), making this autoimmune disorder a prototype disease for studying cancer immunosurveillance. Patients with nontumor LEMS (NT-LEMS) never develop SCLC but are otherwise indistinguishable clinically. Therefore, we have compared immunogenetic factors in SCLC-LEMS and NT-LEMS and studied their role in the pathogenesis of LEMS and survival from SCLC. In 48 British and 29 Dutch Caucasian LEMS patients, we studied clinical symptoms, antibody titers, HLA types and alleles at six nearby located microsatellite loci. Highly significant associations were found in NT LEMS, which appeared strongest with HLA-B8, but also involved HLA-DQ2, -DR3 and six flanking microsatellite alleles. SCLC-LEMS patients were not different from controls. Smoking was a strong predictor of SCLC. In contrast, HLA-B8 positivity correlated with a decreased risk of SCLC even among the smokers. Moreover, in SCLC-LEMS patients, HLA-B8 positivity correlated with prolonged survival after LEMS onset. We propose that two distinct immunopathogenetic routes can lead to one clinically and serologically indistinguishable autoimmune myasthenic syndrome. HLA-DR3-B8 is strongly associated with LEMS in nontumor patients only. In other LEMS patients, SCLC apparently provides a powerful autoimmunogenic stimulus that overrides HLA restrictions in breaking tolerance to calcium channels. Moreover, negativity for HLA-B8 combined with smoking behavior points more strongly to an underlying SCLC and predicts a worse prognosis in SCLC-LEMS patients. PMID- 15652425 TI - Improved stabilization of chemically aminated enzymes via multipoint covalent attachment on glyoxyl supports. AB - The surface carboxylic groups of penicillin G acylase and glutaryl acylase were chemically aminated in a controlled way by reaction with ethylenediamine via the 1-ethyl-3-(dimethylamino-propyl) carbodiimide coupling method. Then, both proteins were immobilized on glyoxyl agarose. In both cases, the immobilization of the chemically modified enzymes improved the enzyme stability compared to the stability of the immobilized but non-modified enzyme (by a four-fold factor in the case of PGA and a 20-fold factor in the case of GA). The chemical modification presented a deleterious effect on soluble enzyme stability. Therefore, the improved stability should be related to a higher multipoint covalent attachment, involving both the lysine amino groups and also the new amino groups chemically introduced on the enzyme. Moreover, the lower pK(a) of the new amino groups permitted to immobilize the enzyme under milder conditions. In fact, the aminated proteins could be immobilized even at pH 9, while the non modified enzymes could only be immobilized at pH over 10. PMID- 15652426 TI - Modulation of gene expression by promoter mutants of the lambdacI857/pRM/pR system. AB - Gene expression driven by the p(R) promoter of the lambdacI857/p(RM)/p(R) system results from inactivation of the temperature-sensitive CI857 repressor. The CI857 repressor, whose gene is transcribed by the divergently orientated p(RM) promoter, is destabilised at temperatures above 30 degrees C. In this study, the lambdacI857/p(RM)/p(R) system was modified by the introduction of a single (A 32G) and a double mutation (A-32G and T-41C). The mutated lambdap(R) expression modules, 32G and 32G/41C, tightly repressed the highly lethal phage PhiX174 lysis gene E at temperatures up to 37 and 39 degrees C, respectively. Expression of protein E and subsequent lysis of Escherichia coli was still induced by a temperature up-shift to 42 degrees C. The impact of the mutations on gene expression levels driven by the lambdap(R) and p(RM) promoters was evaluated at various temperatures using the lacZ reporter gene. Results indicate that the A 32G mutation confers a lambdap(R) promoter-down phenotype. The additional increase in the temperature stability of the 32G/41C expression system is due to the T-41C mutation leading to a higher p(RM) activity. The described lambdap(R) expression modules can be used to obtain a defined expression level at a given temperature and to tightly repress in particular highly lethal genes at different bacterial growth temperatures. PMID- 15652427 TI - Enzymatic grafting of chitosan onto Bombyx mori silk fibroin: kinetic and IR vibrational studies. AB - The potential for using tyrosinase to graft the polysaccharide chitosan (Ch) onto Bombyx mori silk fibroin (SF) was examined. FT-IR spectroscopy coupled to HPLC amino acid analysis showed that mushroom tyrosinase (MT) catalyses the oxidation of tyrosine (Tyr) of SF to electrophilic o-quinones. Kinetic studies showed that only a fraction of the Tyr residues available on the SF chain were oxidized. This result was interpreted in the light of the structure assumed by SF in aqueous solution: Tyr aromatic side chain groups buried into the folded hydrophobic portions of the chain were probably less accessible to MT for steric reasons. Using slightly acidic conditions (pH 6), it was possible to modify SF under homogeneous conditions. FT-IR spectroscopy provided evidence that Ch was grafted onto MT-oxidized SF: the o-quinones were found to undergo a subsequent non enzymatic reaction with nucleophilic amino groups of Ch via Schiff-base and Michael addition mechanisms. Various factors, i.e. reaction time, pH, MT/SF ratio, were found to influence the grafting yield. The highest grafting yield was achieved at pH 7, i.e. more favorable to MT activity rather than to Ch solubility, suggesting that the determining step of the grafting reaction is the formation of o-quinones. The FT-IR spectroscopy revealed that grafting induced a beta-sheet --> random coil conformational transition. PMID- 15652428 TI - Adaptive dissolved oxygen control through the glycerol feeding in a recombinant Pichia pastoris cultivation in conditions of oxygen transfer limitation. AB - In high cell density cultivation processes the productivity is frequently constrained by the bioreactor maximum oxygen transfer capacity. The productivity can often be increased by operating the process at low dissolved oxygen concentrations close to the limitation level. This may be accomplished with a closed-loop controller that regulates the dissolved oxygen concentration by manipulating the dominant carbon source feeding rate. In this work we study this control problem in a pilot 50l bioreactor with a high cell density recombinant P. pastoris cultivation in complex media. The study focuses on the design of accurate stable adaptive controllers, with guaranteed exponential convergence and its relation with the calibration of controller parameters. Two adaptive control strategies were tested in the pilot bioreactor: a model reference adaptive controller with a linear reference model and an integral feedback controller with adaptive gain. The latter alternative proved to be more robust to errors in the measurements of the off-gas composition. Concerning the instrumentation, algorithms were derived assuming that both the dissolved oxygen tension and off gas composition are measured on-line, but also the case of only dissolved oxygen being measured is addressed. It was verified that the measurement of off-gas composition might not improve the controller performance due to measurement and process time delays. PMID- 15652430 TI - Pellet morphology, culture rheology and lovastatin production in cultures of Aspergillus terreus. AB - Pellet growth of Aspergillus terreus ATCC 20542 in submerged batch fermentations in stirred bioreactors was used to examine the effects of agitation (impeller tip speed u(t) of 1.01-2.71 ms(-1)) and aeration regimens (air or an oxygen-enriched mixture containing 80% oxygen and 20% nitrogen by volume) on the fungal pellet morphology, broth rheology and lovastatin production. The agitation speed and aeration methods used did not affect the biomass production profiles, but significantly influenced pellet morphology, broth rheology and the lovastatin titers. Pellets of approximately 1200 microm initial diameter were reduced to a final stable size of approximately 900 microm when the agitation intensity was >/=600 rpm (u(t)>/=2.03 ms(-1)). A stable pellet diameter of approximately 2500 microm could be attained in less intensely agitated cultures. These large fluffy pellets produced high lovastatin titers when aerated with oxygen-enriched gas but not with air. Much smaller pellets obtained under highly agitated conditions did not attain high lovastatin productivity even in an oxygen-enriched atmosphere. This suggests that both an upper limit on agitation intensity and a high level of dissolved oxygen are essential for attaining high titers of lovastatin. Pellet size in the bioreactor correlated equally well with the specific energy dissipation rate and the energy dissipation circulation function. The latter took into account the frequency of passage of the pellets through the high shear regions of the impellers. Pellets that gave high lovastatin titers produced highly shear thinning cultivation broths. PMID- 15652429 TI - Chemo-enzymatic synthesis of tripeptide RGD diamide in organic solvents. AB - The tripeptide BzArgGlyAsp(NH(2))(2) was synthesized by a combination of chemical and enzymatic methods in this study. First of all, GlyAsp(NH(2))(2) was synthesized by a novel chemical method in three steps including chloroacetylation of L-aspartic acid, esterification of chloroacetyl L-aspartic acid and ammonolysis of chloroacetyl L-aspartic acid diethyl ester. Secondly, kinetically controlled synthesis of BzArgGlyAsp(NH(2))(2) catalyzed by trypsin in organic solvent was conducted. The optimum conditions are pH 8.0, 30 degrees C in ethanol/Tris-HCl buffer system (85:15, v/v) for 80 min in the maximum yield of 74.4%. PMID- 15652431 TI - Separation and purification of benzylpenicillin produced by fermentation using coupled ultrafiltration and nanofiltration technologies. AB - The purpose of this study was to evaluate the capacity of using coupled ultrafiltration-nanofiltration technologies for separation and purification of benzylpenicillin (BP). More specifically, we verified the efficiency of three ultrafiltration (UF) membranes (cut-off of 5000, 30,000 and 100,000 Da) to remove impurities that cause stable emulsion during the chemical extraction of the antibiotic. We also tested the effectiveness of a nanofiltration (NF) membrane (cut-off of 300 Da) to concentrate the benzylpenicillin recovered from permeates and to decrease the osmotic pressure by reducing the ionic charge of the broth. Results have shown that high recovery (89.0-91.0%) can be obtained in permeate generated by the 30,000 and 100,000 UF membranes, but a slight emulsion will be formed during phase separation. With the 5000 UF membrane, lower recovery is obtained (81.0%) but no emulsion is produced, leading to a high solvent extraction yield (94.6%). The nanofiltration of 30,000 and 100,000 UF permeates leads to very high recovery (98.0%), but stable emulsions are formed, reducing the chemical extraction yield (80.0-82.6%). For the nanofiltration of 5000 UF permeate, excellent recovery of the antibiotic is noted (97.4%) leading to high extraction yield (92.4%) with no emulsion formed. Diafiltration step should be applied during UF procedure in order to increase the antibiotic recovery in the generated permeates. PMID- 15652432 TI - One-step chromatography method for efficient separation and purification of R phycoerythrin from Polysiphonia urceolata. AB - Phycoerythrins have been widely used in food, cosmetics, immunodiagnostics and analytical reagents. An efficient one-step chromatography method for purification of R-phycoerythrins from Polysiphonia urceolata was described in this paper. Pure R-phycoerythrin was obtained with an absorbance ratio A(565)/A(280) of 5.6 and a high recovery yield of 67.33% using a DEAE-Sepharose Fast Flow chromatography with a gradient elution of pH, alternative to common gradient elution of ionic strength. The absorption spectrum of R-phycoerythrin was characterized with three absorbance maxima at 565, 539 and 498 nm, respectively and the fluorescence emission spectrum at room temperature was measured to be 580 nm. The results of native-PAGE, and SDS-PAGE showed no contamination by other proteins in the phycoerythrin solution, which suggests an efficient method for the separation and purification of R-phycoerythrins from Polysiphonia urceolata. PMID- 15652434 TI - Pain and palliative care: the emergence of new specialties. AB - In the second half of the twentieth century, the clinical management of patients suffering pain from advanced cancer was transformed. This paper describes cancer pain management during this period, identifying three key elements that promoted innovation: First, the development of a patient-centered approach to analgesic evaluation, which resulted from the search for an alternative analgesic to morphine in studies led by Raymond Houde. Second, the re-introduction by John Bonica of the idea that pain is what the individual feels and thinks it is. Third, the work of Cicely Saunders in establishing the foundations of the modern hospice and palliative care movement. The work of these three clinicians must be set in the context of a time when new hopes emerged that cancer could be cured and, at the same time, the cancer patient began to be remolded from a passive participant in treatment and care to an active collaborator. PMID- 15652435 TI - The measurement of pain, 1945-2000. AB - Three strands of activity can be identified in the history of pain measurement. The first, psychophysics, dates back to the nineteenth century and measures the effect of analgesia by quantifying the noxious stimulation required to elicit pain, as well as the maximum stimulation tolerated. The second uses standardized questionnaires for patients, developed to categorize pain according to its emotional impact, distribution, character, and other dimensions. The third asks patients to report on pain intensity using rating scales, and is used in clinical trials where analgesics are evaluated and results can be combined to influence clinical guidelines and protocols. Although all three strands have found a place in modern clinical practice or drug development, it is the reporting of pain by patients undergoing treatment using simple scales of intensity which has emerged as the crucial method by which analgesic therapies can now be evaluated and compared. PMID- 15652436 TI - Stories of cancer pain: a historical perspective. AB - This article considers published accounts by people with personal experience of cancer, and cancer pain, from 1945 to the present. These firsthand stories communicate deeply personal truths and understandings--and this is their value. Narratives of cancer can inform and support others with the disease and contribute meaningful knowledge to health professionals about the subjective experience of living with cancer and cancer pain. They also highlight where improvements can be made in health care and serve as a platform from which patients may criticize, expose, petition, support, share, challenge, and call for autonomy. A cancer diagnosis can rupture the life of the person who receives it, but as this article illustrates, inscribing the words of a narrative and revealing experiences to a public audience can bring meaning to a life event which might otherwise be viewed as meaningless. PMID- 15652437 TI - The syringe driver and the subcutaneous route in palliative care: the inventor, the history and the implications. AB - Since the early 1980s, the syringe driver has become a commonly used technology in British palliative care, used to administer continuous subcutaneous infusions (CSCI) for symptom management. Although the device itself has not been adopted universally, it has stimulated interest in the use of CSCI in palliative care and played a significant role in the modern history of this approach. This historical case study of the syringe driver examines the life and work of its inventor, explores its development for use in childhood thalassemia, and analyzes the circumstances surrounding its adoption in palliative care. We conclude by considering the reasons for the continued popularity of the syringe driver, despite problems in its use, and reflect on the lessons which can be learned about the use of CSCI in palliative care internationally. PMID- 15652438 TI - The ladder and the clock: cancer pain and public policy at the end of the twentieth century. AB - The origins of the WHO Cancer Pain Relief Program (the Analgesic Ladder) and its research basis in two very different research traditions, one at Memorial Sloan Kettering Cancer Center in New York, the other at St. Christopher's Hospice in London, are discussed. The Sloan-Kettering group emphasized precise relative differences in analgesic effects of various drugs, whereas Twycross at St. Christopher's used patient well-being as the crucial benchmark. Despite these differences, both traditions presented evidence of the safe and effective use of strong opioids for cancer pain relief, in a setting of individualized attention and close physician monitoring. The success and limitations of the Ladder as a global health policy are briefly addressed. PMID- 15652439 TI - Contracts, covenants and advance care planning: an empirical study of the moral obligations of patient and proxy. AB - Previously we had speculated that the patient-proxy relationship existed on a contractual to covenantal continuum. In order to assess this hypothesis, and to better understand the moral obligations of the patient-proxy relationship, we surveyed 50 patient-proxy pairs as well as 52 individuals who had acted as proxies for someone who had died. Using structured vignettes representative of three distinct disease trajectories (cancer, acute stroke, and congestive heart failure), we assessed whether respondents believed that proxies should follow explicit instructions regarding life-sustaining therapy and act contractually or whether more discretionary or covenantal judgments were ethically permissible. Additional variables included the valence of initial patient instructions--for example, "to do nothing" or "to do everything"--as well as the quality of information available to the proxy. Responses were graded on a contractual to covenantal continuum using a modified Likert scale employing a prospectively scored survey instrument. Our data indicate that the patient-proxy relationship exists on a contractual to covenantal continuum and that variables such as disease trajectory, the clarity of prognosis, instructional valence, and the quality of patient instructions result in statistically significant differences in response. The use of interpretative or covenantal judgment was desired by patients and proxies when the prognosis was grim, even if initial instructions were to pursue more aggressive care. Nonetheless, there was a valence effect: patients and proxies intended that negative instructions to be left alone be heeded. These data suggest that the delegation of patient self-determination is morally complex. Advance care planning should take into account both the exercise of autonomy and the interpretative burdens assumed by the proxy. Patients and proxies think inductively and contextually. Neither group viewed deviation from patient instructions as a violation of the principal's autonomy. Instead of adhering to narrow notions of patient self-determination, respondents made nuanced and contextually informed moral judgments. These findings have implications for patient education as well as the legal norms that guide advance care planning. PMID- 15652440 TI - Barriers to effective symptom management in hospice. AB - The barriers to effective symptom management in hospice are not well described. We surveyed nurses of hospices affiliated with the Population-based Palliative Care Research Network (PoPCRN) to identify barriers to the effective management of common symptoms in terminally ill patients. 867/1710 (51%) nurses from 67 hospices in 25 U.S. States returned surveys. Of 32 symptoms, nurses reported agitation (45%), pain (40%), and dyspnea (34%) as the 'most difficult to manage.' The most common perceived barriers to effective symptom management were inability of family care providers to implement or maintain recommended treatments (38%), patients or families not wanting recommended treatments (38%), and competing demands from other distressing symptoms (37%). Patterns of barriers varied by symptom. These nurses endorsed multiple barriers contributing to unrelieved symptom distress in patients receiving hospice care. Interventions to improve symptom management in hospice may need to account for these differing barrier patterns. PMID- 15652441 TI - Efficacy, safety, and steady-state pharmacokinetics of once-a-day controlled release morphine (MS Contin XL) in cancer pain. AB - The efficacy, safety, and pharmacokinetics of a novel once-daily morphine formulation (OAD morphine) and a 12-hourly formulation (twice-daily CR morphine) were compared in a double-blind, multi-centered crossover study. Chronic cancer pain patients (n=25) were randomized to OAD morphine (mean 238 +/- 319 mg q24h) or twice-daily CR morphine (mean 119 +/- 159 mg q12h) for one week. They then crossed over to the alternate drug, which also was taken for one week. There was no difference between treatments for evaluations of overall pain intensity, analgesic efficacy, or adverse events. However, whereas pain scores increased during the day on twice-daily CR morphine (P=0.0108), they remained stable on OAD morphine. Most patients (68%) chose once-daily dosing for continuing pain management (P=0.015). The AUC ratio was 100.3%, indicating equivalent absorption. Fluctuation indices were 93.5 +/- 28.8% and 179.3 +/- 41.3% (P=0.0001) for OAD morphine and twice-daily CR morphine, respectively. OAD morphine provides analgesia similar to twice-daily CR morphine with reduced fluctuation in plasma morphine concentration and more stable pain control. PMID- 15652442 TI - Development of an Impact Thermometer for use in combination with the Distress Thermometer as a brief screening tool for adjustment disorders and/or major depression in cancer patients. AB - Screening cancer patients for adjustment disorders and major depression is important, because both are prevalent and often underrecognized. The purpose of this study was to validate the Distress and Impact Thermometer, a 2-item questionnaire, which we newly developed as a brief screening tool for detection of adjustment disorders and/or major depression. Two hundred ninety-five cancer patients completed the Distress and Impact Thermometer and the Hospital Anxiety and Depression Scale (HADS), and were examined by psychiatrists based on DSM-IV criteria. Using cutoff points for detection of adjustment disorders and major depression of "3/4" on "distress" score and "2/3" on "impact," the sensitivity and specificity were 0.82 and 0.82, respectively. Screening performance of the Distress and Impact Thermometer was comparable to that of the Hospital Anxiety and Depression Scale. Its brevity and good performance suggest that the Distress and Impact Thermometer is an effective tool for routine screening in clinical oncology settings. PMID- 15652443 TI - Severe pruritus of cholestasis in disseminated cancer: developing a rational treatment strategy. A case report. AB - Severe pruritus is a frequent complication of cholestasis. Both serotonin and opioids play an important role in the development of this symptom. Guidelines to provide rational management of pruritus of cholestasis do not exist. We describe a patient with complex and malignant course of pruritus. She responded to several measures proposed (among other naltrexone), but rapidly became tolerant to them. Buprenorphine with an ultra low dose of naloxone was able to control her symptoms without development of tolerance until her death. PMID- 15652444 TI - Alcohol withdrawal as an underrated cause of agitated delirium and terminal restlessness in patients with advanced malignancy. AB - A significant number of patients with terminal cancer experience terminal restlessness or an agitated delirium in the final days of life. Multifactorial etiologies may contribute to agitation and restlessness for any one patient; alcohol withdrawal may be underrated as a contributing factor. The symptoms and signs of alcohol withdrawal--autonomic dysfunction, tremor, anxiety, sleep disturbances, insomnia, and abnormal vital signs--may continue for 6 to 12 months after the cessation of alcohol. We report four patients with terminal restlessness in whom we believe alcohol withdrawal to be a significant causal factor and a fifth patient who subsequently benefited from our team's increased awareness of this clinical problem. Formal assessment of alcohol withdrawal may be of more value in the palliative setting than using the currently accepted assessment instruments. Many of the medications utilized for the treatment of agitated delirium and terminal restlessness in the palliative care setting are effective therapies for alcohol withdrawal. PMID- 15652445 TI - Palliative care management services in a Medicare Social HMO. PMID- 15652446 TI - Pro- and anti-inflammatory cytokines gene polymorphisms and Helicobacter pylori infection: interactions influence outcome. AB - The aim of this study was to evaluate whether there was any correlation between Helicobacter pylori-associated diseases and (1) H. pylori virulence genes or (2) IL-1B, IL-1RN, IFN-G, TNF-A, IL-10 genetic polymorphisms. Patients with non cardia gastric cancer (NCGC, n=129) or benign gastroduodenal diseases (n=792) were studied. IL-1RN intron 2 VNTR polymorphism (PCR), IL-1B -31 C/T (RFLP), the SNPs of IFN-G (+874 A/T), TNF-A (-1031 C/T, -857 C/T, -376 A/G, -308 A/G, -238 A/G), IL-10 (-1082 A/G, -819 C/T, -592 A/C) (Taqman chemistry) were studied. cagA, s1 and m1 vacA, were PCR amplified. Duodenal ulcer was more frequent in TNF A -857 TT and in IL-1RN 1,2 subjects. TNF-A -857 TT genotype was also correlated with gastric ulcer. IL-10 -819 TT genotype was associated with intestinal metaplasia and NCGC. Antral inflammation was associated with TNF-A -1031 TT, while corpus activity with IL-10 -819 CC. H. pylori infection was associated with TNF-A -308 AG genotype, while IFN-G +874 AA genotype was associated with cagA. In conclusion, among host genetic factors contributing to H. pylori disease outcome, IFN-G +874 AA genotype favors cagA positive infections, TNF-A -857 TT duodenal ulcer while IL-10 -819 TT intestinal metaplasia and NCGC. PMID- 15652447 TI - Circulating levels of the interleukin (IL)-4 receptor and of IL-18 in patients with Plasmodium falciparum malaria. AB - Circulating levels of sIL-4R, IL-18 and IFN-gamma were studied by ELISA in 36 Gabonese patients with Plasmodium falciparum malaria (29 children, 7 adults). Drug induced clearance of parasitemia, studied in 22 patients with mild disease, was accompanied by a rapid decrease of sIL-4R and IFN-gamma to normal values and an increase of circulating IL-18, suggesting the downregulation of a type 2 biased immune response and a dissociated type 1 responsiveness while resolving parasitemia. Comparing subgroups with hyperparasitemia/severe anemia and mild malaria, children with severe malaria had significant higher levels of sIL-4R and IFN-gamma, whereas IL-18 levels were not statistically different. Furthermore, among those children, higher levels of circulating IL-18 correlated with a lower degree of parasitemia. PMID- 15652448 TI - Interleukin-1 stimulates cytokines, prostaglandin E2 and matrix metalloproteinase 1 production via activation of MAPK/AP-1 and NF-kappaB in human gingival fibroblasts. AB - Interleukin-1 (IL-1) plays a crucial role in the immunopathological responses involved with tissue destruction in chronic inflammatory diseases, such as periodontal disease, as it stimulates host cells including fibroblasts to produce various inflammatory mediators and catabolic factors. We comprehensively investigated the involvement of mitogen-activated protein kinases (MAPKs)/activator protein-1 (AP-1) and IkappaB kinases (IKKs)/IkappaBs/nuclear factor-kappaB (NF-kappaB) in IL-1beta-stimulated IL-6, IL-8, prostaglandin E(2) (PGE(2)) and matrix metalloproteinase-1 (MMP-1) production by human gingival fibroblasts (HGF). Three MAPKs, extracellular signal-regulated kinase (ERK), p38 MAPK and c-Jun N-terminal kinase (JNK), which were simultaneously activated by IL 1beta, mediated subsequent c-fos and c-jun mRNA expression and DNA binding of AP 1 at different magnitudes. IKKalpha/beta/IkappaB-alpha/NF-kappaB was also involved in the IL-1 signaling cascade. Further, IL-1beta stimulated HGF to produce IL-6, IL-8, PGE(2) and MMP-1 via activation of the 3 MAPKs and NF-kappaB, as inhibitors of each MAPK and NF-kappaB significantly suppressed the production of IL-1beta-stimulated factors, though these pathways might also play distinct roles in IL-1beta activities. Our results strongly suggest that the MAPKs/AP-1 and IKK/IkappaB/NF-kappaB cascades cooperatively mediate the IL-1beta-stimulated synthesis of IL-6, IL-8, PGE(2) and MMP-1 in HGF. PMID- 15652449 TI - Sequential measurement of IL-6 blood levels in patients with systemic inflammatory response syndrome (SIRS)/sepsis. AB - This study was undertaken to investigate whether sequential measurement of blood interleukin (IL)-6 levels using chemiluminescent enzyme immunoassay (CLEIA) would be useful for the management of patients with systemic inflammatory response syndrome (SIRS)/sepsis. Forty consecutive patients with SIRS/sepsis admitted to ICU were involved in the study. Blood IL-6 level was measured everyday throughout their ICU stay at the clinical laboratory by CLEIA method. The platelet count and the sequential organ failure assessment (SOFA) score were measured consecutively. The blood IL-6 levels were elevated in SIRS/sepsis patients and were extremely high in patients with septic shock. There was no significant difference in the blood IL-6 level on admission between survivors (n=27) and non-survivors (n=13). However, the mean blood IL-6 level during ICU stay was significantly higher in the non-survivors (p<0.05). There were significant correlation between the peak IL-6 blood level and the lowest platelet count, and between the peak IL-6 blood level and the maximum SOFA score, respectively. The platelet count became lowest 2.0+/-2.0 days later on average, and the SOFA score became maximal 2.5+/-1.4 days later on average following the day when IL-6 reached its peak value. Sequential measurement of blood IL-6 levels by CLEIA is useful in evaluating the severity and in predicting the outcome of the patients with SIRS/sepsis. PMID- 15652452 TI - Tissue homeostasis and the control of the neoplastic phenotype in epithelial cancers. AB - Neoplastic cells at various stages of tumor progression may remain dormant for many years. The suppression of the neoplastic phenotype and tumor outgrowth depends on close contact of neoplastic cells with surrounding normal cells. This review examines the nature of these contacts primarily in models for skin cancer induction. Junctional complexes, membrane associated growth factors and their receptors, and paracrine mechanisms likely contribute to this state of tumor cell dormancy. Understanding these mechanisms will be important in primary cancer prevention and for counteracting recurrences in cancer survivors. PMID- 15652450 TI - Interleukin-10 regulates arterial pressure in early primate pregnancy. AB - OBJECTIVES: In pregnancy, the placental contribution of cytokines to maternal immunosuppression has been established, however their role in normal maternal blood pressure regulation has not been identified. We investigate the contribution of interleukin-10 (IL-10) and tumor necrosis factor-alpha (TNF alpha) to the vasodilation of early pregnancy in non-human primates. We also sequenced the IL-10 baboon gene and compared it with humans. METHODS: The effect of four different treatments, administered sequentially (semi-random-design) on resting 18h, night time, or hourly mean arterial pressure (MAP) and heart rate (HR) were measured using telemetry. An anti-human IL-10 monoclonal antibody (MAb, 1mg, n=7), anti-TNF-alpha antibody (n=3), a combination of anti-IL-10 and anti TNF-alpha antibodies (n=5) or saline (n=3) control were administered intravenously to baboons in early pregnancy. Plasma and placental IL-10 concentration was measured before and after injection in all animals. RESULTS: Anti-human IL-10 MAb caused a significant increase in MAP of 2.6+/-0.5mmHg over the 18-h period (p<0.05). Administration of TNF-alpha alone or in combination with IL-10 did not alter MAP. There was 97% sequence homology of IL-10 cDNA between humans and baboons. CONCLUSIONS: IL-10 was shown to regulate the vasodilation of early pregnancy in Papio hamadryas. This partial role of IL-10 in the early BP response of primate pregnancy may be relevant to pathophysiological states of human pregnancy such as preeclampsia. PMID- 15652453 TI - Microenvironmental control of premalignant disease: the role of intercellular adhesion in the progression of squamous cell carcinoma. AB - Interactions between potentially neoplastic cells and their normal neighbors act as an important microenvironmental control during the premalignant, intraepithelial (IE) stage of squamous cell cancer development. Using human three dimensional (3D) tissue models, it has been shown that such interactions induce a state of IE dormancy in which cancer progression is suppressed. This dormant state can be overcome by altering tissue dynamics in response to the tumor promoter TPA, UV irradiation, decreasing adhesive interactions between tumor cells and adjacent epithelia or enabling tumor cells to interact with basement membrane proteins. This chapter will provide an overview of investigations that have characterized how the tissue microenvironment can regulate the incipient development of squamous cell carcinoma. PMID- 15652454 TI - Colonization of adjacent stem cell compartments by mutant keratinocytes. AB - A key step in cancer development is clonal expansion. The increased number of mutant cells allows a clinical phenotype and increases the probability that one of the cells will be mutated in an additional gene. For skin cancer, observations on p53-mutant keratinocyte clones in epidermal sheets of UVB-irradiated mice reveal that mutant stem cells are normally restrained within their stem cell compartment. Chronic UVB exposure drives clonal expansion by a non-mutational mechanism, and this mechanism allows mutant stem cells to escape from their own stem cell compartment and colonize adjacent compartments. In the absence of escape, cells proliferate without occupying additional territory. The non mutational clonal expansion mechanism appears to be UVB-induced apoptosis, which deletes DNA-damaged cells in unmutated stem cell compartments but will preferentially spare death-resistant p53-mutant cells. An additional source of apoptotic selection pressure may come from UV-irradiated melanin. PMID- 15652455 TI - Origins and selection of p53 mutations in lung carcinogenesis. AB - Molecular epidemiologists usually consider the spectrum of p53 mutations found in human tumors to be a signature of the corresponding environmental carcinogen(s). In lung cancer, this signature is the spectrum of G --> T transversions, presumably induced by polycyclic aromatic hydrocarbons (PAH) from cigarette smoke. What complicates the situation, however, is that in the p53 gene the same codons are preferential targets for not only mutagenesis but also tumorigenic selection. In this review, we compare the G --> T spectra induced by PAH o quinones and diol epoxides with those in lung cancer and show that the main "shaper" of the latter is selection, not mutagenesis. In addition, we propose the approach that allows to distinguish selection and mutagenesis components of the p53 spectra and, therefore, to test the suspect carcinogens for their "in vivo" mutagenic involvement. Collectively, the reviewed basic premises, concepts and data are consistent with the increasing recognition of environmental cancer risk conditions as selecting rather than inducing tumorigenic mutations. PMID- 15652456 TI - Expanding fields of genetically altered cells in head and neck squamous carcinogenesis. AB - Recent molecular pathology studies in head and neck cancer support a carcinogenesis model in which the development of a field with genetically altered cells plays a central role. This preneoplastic field is of monoclonal origin and expands non-invasively superseding normal epithelium. Clonal divergence and selection within the field ultimately leads to the development of cancer. These fields can be large (over 7 cm diameter) and are often not visible for the surgeon explaining that they may remain after resection of the primary tumor. When not removed, a field is an important risk factor for secondary cancer. PMID- 15652457 TI - Membrane stabilization by intimate contact between cells reduces their Mg2+ activity and suppresses the neoplastic phenotype. AB - The mechanism of normalizing transformed cells by contact with normal cells is considered in relation to the mechanism of stimulating the growth of normal contact-inhibited cells. Stimulation is based on perturbation of the plasma membrane throughout the G1 period by growth factors. A primary effect of the membrane perturbation is a long-sustained increase of intracellular free Mg(2+) which increases the initiation frequency of polypeptide synthesis and accelerates the onset of DNA synthesis. Because the plasma membrane of transformed cells is kept in a continuously perturbed state and forms poor intercellular adhesions, it loses control of intracellular Mg(2+) activity. However, maximizing contact between transformed cells at extremely high density stabilizes their plasma membrane, lowers their Mg(2+) content and normalizes their phenotype. Normalization may also be obtained in contact with lower densities of quiescent, confluent normal cells. Evidence against a need for junctional communication supports membrane stabilization with restoration of Mg(2+) regulation as the mechanism of normalization. PMID- 15652458 TI - The role of the stromal microenvironment in prostate cancer. AB - It is becoming increasingly clear that a prostate cancer cell's microenvironment is crucial to its survival, progression, and metastasis. However, the mechanisms by which the tumor cell interacts with its surroundings are largely unidentified. Experiments have shown that the growth of prostate carcinoma cells can be either inhibited or stimulated by specific stromal environments. Angiogenesis, a critical factor in cancer progression, is likewise stromally-mediated. As many of the cellular and humoral factors involved in angiogenesis are also significant to the process of wound healing, the "reactive stroma" of a malignant tumor may parallel the granulation tissue of a healing wound in many ways, thereby facilitating the development of valuable experimental models. Metastasis to bone, perhaps the most clinically significant aspect of prostate cancer, is also dependent on stromal-epithelial crosstalk, as prostate carcinoma cells must induce the hospitality of bone cells in order to take up residence in an osseous microenvironment. This article outlines several notions regarding these interplays, and addresses their role in prostate carcinogenesis. PMID- 15652461 TI - Hen and egg, you mitochondria! PBC revisits mitochondria again. PMID- 15652459 TI - Integrative radiation carcinogenesis: interactions between cell and tissue responses to DNA damage. AB - Tissue function requires coordinated multicellular behavior as a consequence of diverse signals integrated through the tissue microenvironment; importantly, these cell-cell and cell-microenvironment interactions also actively suppress cancer. Ionizing radiation (IR) elicits a well-defined cellular response to DNA damage that mediates the fate of the individual cell, concomitantly with a less well-characterized overarching tissue stress response that coordinates the response of multiple cell types via microenvironment signaling. We have now shown that these programs to reestablish homeostasis intersect via mutual regulation by transforming growth factor beta1 (TGF beta 1), which acts as an extracellular sensor and signal of stress. In this review, the concept that this type of functional integration of cell and tissue stress response programs is essential to cancer suppression will be discussed. Our experiments using IR, and several recent studies that experimentally manipulate stromal TGF beta, show that disruption of microenvironment signaling actively promotes malignant progression. Understanding the dynamic interactions between tissue and cell stress responses will be necessary for an accurate assessment of cancer risk and may also provide targets for prevention. PMID- 15652460 TI - Cathepsin B and tumor proteolysis: contribution of the tumor microenvironment. AB - Tumor-stromal interactions induce expression of matrix metalloproteinases and serine proteases and, as shown recently, the cysteine protease cathepsin B. We speculate that such interactions upregulate the transcription factor Ets1, resulting in increased cathepsin B expression. This would be consistent with the observed concomitant upregulation of matrix metalloproteinases and serine proteases as well as with the ability of extracellular matrices and their binding partners to alter cathepsin B expression and secretion. Using a confocal assay to analyze the contribution of tumor-stromal interactions to proteolysis, we have been able to confirm enhanced degradation of extracellular matrices by all three classes of proteases. PMID- 15652462 TI - Complications of fine needle biopsy for focal liver lesions. PMID- 15652463 TI - Virulent Klebsiella pneumoniae strains and liver abscess with metastatic lesions. PMID- 15652464 TI - Expression of Notch signalling markers in bone marrow cells that differentiate into a liver cell lineage in a rat transplant model. AB - Notch signalling pathway plays an important role in cell differentiation. To investigate the implications of Notch signalling in the differentiation of rat bone marrow (BM) cells into a liver cell lineage, we used cultured BM cells to examine the mRNA expression of Musashi-1, which positively regulates Notch signalling, and made a transplant model to examine the protein expression of Notch signalling markers. For the in vivo experiment, BM cells were collected from transgenic rats expressing green fluorescence protein (GFP) and transplanted into the spleens of recipient rats, in which liver damage had been induced with carbon tetrachloride. The expression of Notch receptor 1 (Notch-1), Jagged-1 and Musashi-1, in the transplanted GFP-positive BM cells was investigated by immunohistochemistry. The expression of the liver-specific proteins, alpha fetoprotein and cytokeratin19 was also investigated. Musashi-1 mRNA became detectable in the BM cells on culture day 7 in vitro. After transplantation, GFP positive BM cells were observed in the portal areas of the recipient's livers. Notch-1, Jagged-1, Musashi-1, alpha-fetoprotein and cytokeratin19 were all expressed in the transplanted BM cells. These results suggest that the Notch signalling pathway plays a role in the differentiation of BM cells into a liver cell lineage. PMID- 15652466 TI - Spontaneous resolution of chronic hepatitis C virus infection after antiviral treatment and relapse. AB - Clearance of acute hepatitis C virus (HCV) infection is associated with strong and multi-specific cellular immune responses which are often weak in chronic hepatitis C. We here report a case of spontaneous and sustained resolution of chronic hepatitis C virus infection in the absence of apparent HCV-specific immunity. The patient received standard antiviral therapy for chronic HCV infection and was HCV-RNA negative at the end of treatment but relapsed between follow-up week 4 and 12. Surprisingly, from follow-up week 28 on, he persistently was HCV-RNA negative in serum, even when being tested with the highly sensitive TMA-assay (cut-off 5-10IU/ml). ALT levels were within the normal range throughout follow-up. Virus-specific CD4+ T cell responses were prospectively analysed during the relapse period and during spontaneous resolution by interferon-gamma ELISPOT assays. Importantly, no HCV-specific cellular immune responses were detectable at any time-point. The patient suffered from an acute respiratory tract infection before HCV clearance and serum IL-8 levels were significantly increased during this period. Thus, spontaneous resolution of hepatitis C after antiviral treatment and relapse may occur even in the absence of hepatitis flares and apparent HCV-specific immune responses in single cases. The role of heterologous infections for HCV clearance requires further investigation. PMID- 15652465 TI - Body surface area is an independent factor contributing to the effects of lamivudine treatment. AB - BACKGROUND:: It has been suggested that lamivudine therapy may be even more effective if administered at higher doses than is dictated by the current standard regimen. We analyzed the correlation between the effects of lamivudine and body surface area (BSA). METHOD:: We evaluated 134 patients with chronic hepatitis B who had been treated with lamivudine for more than 12 months. The effect of the treatment was evaluated from the levels of serum alanine aminotransferase (ALT) and HBV-DNA. Several variables that could influence the response to treatment, including ALT, albumin, and bilirubin levels, platelet counts, BSA, HBV-DNA, and HBeAg were analyzed. RESULTS:: Univariate logistic analysis selected platelet counts, BSA, HBV-DNA and HBeAg in the biological evaluation, and bilirubin, BSA, HBV-DNA and HBeAg in the virological evaluation (chi(2)>1.0). Using these factors, multivariate analysis revealed that BSA (chi(2)=12.8, p=0.0004) was the only factor that could contribute significantly to the improvement of ALT levels, and that BSA (chi(2)=4.4, p=0.0354) and HBeAg (chi(2)=8.1, p=0.0044) were independent factors that could influence the suppression of HBV-DNA. CONCLUSION:: We revealed that BSA is a significantly predictor of the effect of lamivudine therapy, suggesting that lamivudine dosage should be based on the individual BSA. PMID- 15652467 TI - Enhancement of mitochondrial gene expression in the liver of primary biliary cirrhosis. AB - Primary biliary cirrhosis (PBC) is one of the most important autoimmune liver diseases but the etiology and pathogenesis remain unknown. In this study, we analyzed differential mRNA expression in the liver of a patient with PBC using suppression subtractive hybridization to identify overexpressed genes. Overexpression of mRNA transcripts from mitochondrial DNA was observed in the PBC liver, compared to normal liver. To explore the mechanism of increased mitochondrial transcription, we investigated the mRNA levels of nuclear DNA encoded regulator molecules of mitochondrial gene expression in 60 liver biopsy samples from various diseases, including PBC, using competitive RT-PCR. Increased expression of mitochondrial transcriptional factor A (mtTFA) and mitochondrial nuclear respiratory factor 1 (NRF-1) mRNA was demonstrated in PBC liver compared to other liver diseases, while NRF-1 coactivator 1, PGC-1 was suppressed. Mitochondrial DNA-encoded mRNA molecules are overexpressed in the PBC liver, and this is associated with up-regulation of mitochondrial transcription factor mtTFA and its transactivator NRF-1. Further studies are needed to focus on the relevance of this perturbation of mitochondrial gene expression in the pathogenesis of PBC. PMID- 15652468 TI - Safety of fine-needle aspiration in patients with small hepatocellular carcinoma. AB - OBJECTIVE.: This study was to clarify the safety of fine-needle aspiration of small hepatocellular carcinoma (HCC). METHODS.: Ninety consecutive patients with small hepatocellular carcinoma (46 with single nodule, 44 with multiple nodules, all 0.05) whereas the residual strain of ileum at the mucosal surface decreased with age (p < 0.001). The circumferential and longitudinal stress-strain curves fitted well to a mono-exponential function. At a given circumferential stress, the corresponding strain values increased during the first 8 weeks of age (p < 0.05) where after no further change was observed. Hence, the small intestine became more compliant during early life. At a given longitudinal stress, the corresponding strains of ileum and duodenum became larger during the first 2-4 weeks of age (p < 0.05) where after no further change was observed. The small intestine was stiffer in longitudinal direction compared to the circumferential direction. In conclusion, pronounced morphometric and biomechanical changes were observed in the rat small intestine during physiological growth. Such data may prove useful in the understanding of the functional changes of the digestive tract during early life. PMID- 15652540 TI - Regional load bearing of the feline acetabulum. AB - OBJECTIVE: To test if the caudal acetabulum of the cat is unloaded at mid-stance as has been asserted. MATERIALS AND METHODS: A kinematic study of five healthy cats was performed to ascertain the mid-stance angle of the hip joint, and the orientation of the pelvis and femur. Femora and pelves from 10 feline cadavers were loaded at physiological load at the mid-stance angle. Impression material placed within the acetabulum was extruded from areas of load bearing. Digital images before and after loading were used to assess whether three areas of the acetabulum, cranial, central and caudal, were fully, partially or non-load bearing. RESULTS: There was a significant difference in load bearing in the three regions (p < 0.001, Kruskal-Wallis test), with a significant difference in load bearing between the cranial and central thirds (p < 0.001) and the cranial and caudal thirds (p < 0.001) but no difference between the central and caudal thirds of the acetabulum (Mann-Whitney-U test). CONCLUSIONS: The load bearing areas of the feline acetabula are the caudal and central thirds. CLINICAL RELEVANCE: The caudal acetabulum of the cat is loaded, therefore the recommendations for simple fractures being treated conservatively needs to be reconsidered. PMID- 15652541 TI - Estimation of the binding force of the collagen molecule-decorin core protein complex in collagen fibril. AB - Decorin belongs to the small leucine proteoglycans family and is considered to play an important role in extracellular matrix organization. Experimental studies suggest that decorin is required for the assembly of collagen fibrils, as well as for the development of proper tissue mechanical properties. In tendons, decorins tie adjoining collagen fibrils together and probably guarantee the mechanical coupling of fibrils. The decorin molecule consists of one core protein and one glycosaminoglycan chain covalently linked to a serine residue of the core protein. Several studies have indicated that each core protein binds to the surface of collagen fibrils every 67 nm, by interacting non-covalently to one collagen molecule of the fibril surface, while the decorin glycosaminoglycans extend from the core protein to connect to another decorin core protein laying on adjacent fibril surface. The present paper investigates the complex composed of one decorin core protein and one collagen molecule in order to obtain their binding force. For this purpose, molecular models of collagen molecules type I and decorin core protein were developed and their interaction energies were evaluated by means of the molecular mechanics approach. Results show that the complex is characterized by a maximum binding force of about 12.4 x 10(3) nN and a binding stiffness of 8.33 x 10(-8) N/nm; the attained binding force is greater than the glycosaminoglycan chain's ultimate strength, thus indicating that overloads are likely to damage the collagen fibre's mechanical integrity by disrupting the glycosaminoglycan chains rather than by causing decorin core protein detachment from the collagen fibril. PMID- 15652542 TI - Ground reaction forces during downhill and uphill running. AB - We investigated the normal and parallel ground reaction forces during downhill and uphill running. Our rationale was that these force data would aid in the understanding of hill running injuries and energetics. Based on a simple spring mass model, we hypothesized that the normal force peaks, both impact and active, would increase during downhill running and decrease during uphill running. We anticipated that the parallel braking force peaks would increase during downhill running and the parallel propulsive force peaks would increase during uphill running. But, we could not predict the magnitude of these changes. Five male and five female subjects ran at 3m/s on a force treadmill mounted on the level and on 3 degrees, 6 degrees, and 9 degrees wedges. During downhill running, normal impact force peaks and parallel braking force peaks were larger compared to the level. At -9 degrees, the normal impact force peaks increased by 54%, and the parallel braking force peaks increased by 73%. During uphill running, normal impact force peaks were smaller and parallel propulsive force peaks were larger compared to the level. At +9 degrees, normal impact force peaks were absent, and parallel propulsive peaks increased by 75%. Neither downhill nor uphill running affected normal active force peaks. Combined with previous biomechanics studies, our normal impact force data suggest that downhill running substantially increases the probability of overuse running injury. Our parallel force data provide insight into past energetic studies, which show that the metabolic cost increases during downhill running at steep angles. PMID- 15652543 TI - Morphological age-dependent development of the human carotid bifurcation. AB - The unique morphology of the adult human carotid bifurcation and its sinus has been investigated extensively, but its long-term, age-dependent development has not. It is important fundamentally and clinically to understand the hemodynamics and developmental forces that play a role in remodeling of the carotid bifurcation and maturation of the sinus in association with brain maturation. This understanding can lead to better prognostication and therapy of carotid disease. We analyzed the change of sinus morphology and the angle of the carotid bifurcation in four postnatal developmental stages (Group I: 0-2 years, Group II: 3-9 years, Group III: 10-19 years, and Group IV: 20-36 years, respectively) using multiprojection digital subtraction angiograms and image post-processing techniques. The most significant findings are the substantial growth of the internal carotid artery (ICA) with age and the development of a carotid sinus at the root of the ICA during late adolescence. The bifurcation angle remains virtually unchanged from infancy to adulthood. However, the angle split between the ICA and external carotid artery (ECA) relative to the common carotid artery (CCA) undergoes significant changes. Initially, the ICA appears to emanate as a side branch. Later in life, to reduce hydraulic resistance in response to increased flow demand by the brain, the bifurcation is remodeled to a construct in which both daughter vessels are a skewed continuation of the parent artery. This study provides a new analysis method to examine the development of the human carotid bifurcation over the developmental years, despite the small and sparse database. A larger database will enable in the future a more extensive analysis such as gender or racial differences. PMID- 15652544 TI - A comparison of mechanical properties derived from multiple skeletal sites in mice. AB - Laboratory mice provide a versatile experimental model for studies of skeletal biomechanics. In order to determine the strength of the mouse skeleton, mechanical testing has been performed on a variety of bones using several procedures. Because of differences in testing methods, the data from previous studies are not comparable. The purpose of this study was to determine which long bone provides the values closest to the published material properties of bone, while also providing reliable and reproducible results. To do this, the femur, humerus, third metatarsal, radius, and tibia of both the low bone mass C57BL/6H (B6) and high bone mass C3H/HeJ (C3H) mice were mechanically tested under three point bending. The biomechanical tests showed significant differences between the bones and between mouse strains for the five bones tested (p < 0.05). Computational models of the femur, metatarsal, and radius were developed to visualize the types of measurement error inherent in the three-point bending tests. The models demonstrated that measurement error arose from local deformation at the loading point, shear deformation and ring-type deformation of the cylindrical cross-section. Increasing the aspect ratio (bone length/width) improved the measurement of Young's modulus of the bone for both mouse strains (p < 0.01). Bones with the highest aspect ratio and largest cortical thickness to radius ratio were better for bending tests since less measurement error was observed in the computational models. Of the bones tested, the radius was preferred for mechanical testing because of its high aspect ratio, minimal measurement error, and low variability. PMID- 15652545 TI - Effect of foot orthotics on rearfoot and tibia joint coupling patterns and variability. AB - The purpose of this study was to compare joint coupling patterns and variability of the rearfoot and tibia during running in subjects who were treated with two types of orthotic devices to that of controls. Eleven subjects with various lower extremity injuries were treated unsuccessfully with a standard orthotic, and then successfully with an inverted orthotic. Three-dimensional kinematic data were collected while subjects ran without orthoses and then in standard and inverted orthoses. Eleven healthy subjects ran without orthoses for comparison. The rearfoot inversion/eversion and tibial internal/external rotation joint coupling pattern and variability relationship was assessed using a vector coding technique. It was hypothesized that when the treated runners ran without orthotic devices, they would exhibit lower joint coupling angles and lower joint coupling variability compared to the controls. In addition, it was hypothesized that there would be no difference in the coupling angle or coupling variability between the standard and no orthotic conditions of the treated runners. Finally, it was hypothesized that coupling angle would decrease and variability would increase in the inverted versus the standard and non-orthotic conditions. No significant differences in joint coupling pattern or variability were observed between the treated and control subjects. In addition, no significant differences were noted between the orthotic conditions in the treated group. These results suggest that foot orthotic devices do not produce significant changes in rearfoot-tibial coupling. Therefore, the relief experienced with the inverted orthotic is likely due to factors other than alterations in this coupling. PMID- 15652546 TI - Expression of matrix metalloproteinase genes in the rat intramembranous bone during postnatal growth and upon mechanical stresses. AB - A cranial suture consists of neural-crest derived cells and matrices between mineralized skull bones. Little is known regarding the involvement of matrix metalloproteinases (MMPs) in the degradation of extracellular matrix of cranial sutures. In the postnatal rat model, the posterior frontal suture (PFS) undergoes complete ossification between P12-P22, whereas the sagittal suture (SS) remains patent. The present study utilized reverse transcriptase-polymerase chain reaction (RT-PCR) to explore the expression of MMP-1 and MMP-2 genes in the PFS and SS in P8 and P32 rats, and also to determine whether these MMP genes are modulated by exogenous mechanical forces. RNA was isolated from P8 and P32 normal PFS and SS each by pooling sutural specimens from 14 to 20 rats. RT-PCR analysis and semi-quantitative luminosity demonstrated the expression of MMP-1 and MMP-2 genes in the patent P8 PFS, P8 SS, and P32 SS, but no apparent MMP-2 expression in the physiologically ossified P32 PFS. Exogenous cyclic forces applied to the maxilla at 1000 mN and 4 Hz elicited corresponding cyclic bone strain waveforms with peak strain of 134.14+/-38.15 muepsilon (mean+/-S.D.) for the PFS, and 28.35+/-10.86 muepsilon for the SS in P32 rats. These cyclic forces delivered for 20 min/d over 2 consecutive days induced the expression of MMP-2 gene in the physiologically fused P32 PFS that was not expressed without mechanical stresses. Taken together, these data suggest potentially important roles of MMP genes in the postnatal development of cranial sutures, and their susceptibility to mechanical stresses. PMID- 15652547 TI - Influence of stress rate on water loss, matrix deformation and chondrocyte viability in impacted articular cartilage. AB - The biomechanical response of articular cartilage to a wide range of impact loading rates was investigated for stress magnitudes that exist during joint trauma. Viable, intact bovine cartilage explants were impacted in confined compression with stress rates of 25, 50, 130 and 1000 MPa/s and stress magnitudes of 10, 20, 30 and 40 MPa. Water loss, cell viability, dynamic impact modulus (DIM) and matrix deformation were measured. Under all loading conditions the water loss was small (approximately 15%); water loss increased linearly with increasing peak stress and decreased exponentially with increasing stress rate. Cell death was localized within the superficial zone (< or =12% of total tissue thickness); the depth of cell death from the articular surface increased with peak stress and decreased with increasing stress rate. The DIM increased (200-700 MPa) and matrix deformation decreased with increasing stress rate. Initial water and proteoglycan (PG) content had a weak, yet significant influence on water loss, cell death and DIM. However, the significance of the inhomogeneous structure and composition of the cartilage matrix was accentuated when explants impacted on the deep zone had less water loss and matrix deformation, higher DIM, and no cell death compared to explants impacted on the articular surface. The mechano-biological response of articular cartilage depended on magnitude and rate of impact loading. PMID- 15652548 TI - Hyaluronic acid diminishes the resistance to excursion after flexor tendon repair: an in vitro biomechanical study. AB - Adhesion between the tendon and tendon sheath after primary flexor tendon repair is seen frequently, and postoperative finger function is occasionally unsatisfactory. A reduction of the friction may facilitate tendon mobilization, which in turn may reduce the risk of the adhesion and restriction of range of motion. We considered the possibility of utilizing the hyaluronic acid (HA) as a lubricant. To evaluate the effect of HA, the gliding resistance between the canine flexor digitorum profundus tendon repaired by a modified Kessler suture technique with running epitendinous suture and the annular pulley located on the proximal phalanx (corresponding to the A2 pulley in humans) was evaluated and compared before and after administration of HA. The HA solution measurement groups were identified as follows; intact tendon as a control; repaired tendon; tendon soaked in 0.1, 1, and 10 mg/ml HA. The resistance increased after repairing, then it decreased after soaking in 10 mg/ml HA solution. The results of this study revealed that HA diminishes the excursion resistance after flexor tendon repair. We believe that some style of administration of the HA might reduce the excursion resistance and prevent adhesion until the synovial surface is fully developed. PMID- 15652550 TI - Representing and identifying alternative movement techniques for goal-directed manual tasks. AB - Differences in motion patterns subserving the same movement goal can be identified qualitatively. These alternatives, which may characterize 'movement techniques' (e.g., the stoop and the squat lifting technique), may be associated with significantly different biomechanical constraints and physiological responses. Despite the widely shared understanding of the significance of alternative movement techniques, quantitative representation and identification of movement techniques have received little attention, especially for three dimensional whole-body motions. In an attempt to systematically differentiate movement techniques, this study introduces a quantitative index termed joint contribution vector (JCV) representing a motion in terms of contributions of individual joint degrees-of-freedom to the achievement of the task goal. Given a set of uncharacterized (unlabeled) motions represented by joint angle trajectories (motion capture data), the JCV and statistical clustering methods enable automated motion classification to uncover a taxonomy of alternative movement techniques. The results of our motion data analyses show that the JCV was able to characterize and discern stoop and squat lifting motions, and also to identify movement techniques for a three-dimensional, whole-body, one-handed load transfer task. The JCV index would facilitate consideration of alternative movement techniques in a variety of applications, including work method comparison and selection, and human motion modeling and simulation. PMID- 15652549 TI - Osteoarthritic changes in the biphasic mechanical properties of the chondrocyte pericellular matrix in articular cartilage. AB - The pericellular matrix (PCM) is a narrow region of cartilaginous tissue that surrounds chondrocytes in articular cartilage. Previous modeling studies indicate that the mechanical properties of the PCM relative to those of the extracellular matrix (ECM) can significantly affect the stress-strain, fluid flow, and physicochemical environments of the chondrocyte, suggesting that the PCM plays a biomechanical role in articular cartilage. The goals of this study were to measure the mechanical properties of the PCM using micropipette aspiration coupled with a linear biphasic finite element model, and to determine the alterations in the mechanical properties of the PCM with osteoarthritis (OA). Using a recently developed isolation technique, chondrons (the chondrocyte and its PCM) were mechanically extracted from non-degenerate and osteoarthritic human cartilage. The transient mechanical behavior of the PCM was well-described by a biphasic model, suggesting that the viscoelastic response of the PCM is attributable to flow-dependent effects, similar to that of the ECM. With OA, the mean Young's modulus of the PCM was significantly decreased (38.7+/-16.2 kPa vs. 23.5+/-12.9 kPa, p < 0.001), and the permeability was significantly elevated (4.19+/-3.78 x10(-17) m(4)/Ns vs. 10.2+/-9.38 x 10(-17) m(4)/Ns, p < 0.01). The Poisson's ratio was similar for both non-degenerate and OA PCM (0.044+/-0.063 vs. 0.030+/-0.068, p > 0.6). These findings suggest that the PCM may undergo degenerative processes with OA, similar to those occurring in the ECM. In combination with previous theoretical models of cell-matrix interactions in cartilage, our findings suggest that changes in the properties of the PCM with OA may have an important influence on the biomechanical environment of the chondrocyte. PMID- 15652551 TI - Cell relaxation after electrodeformation: effect of latrunculin A on cytoskeletal actin. AB - Precise measurement of the mechanical properties of a cell provides useful information about its structural organization and physiological state. It is interesting to understand the effect of individual components on the mechanical properties of the entire cell. In this study, we investigate the influence of the cytoskeletal actin on the viscoelastic properties of a cell. Actin-specific agents, including latrunculin A and jasplakinolide, are used to alter the organization of the cytoskeletal actin. Brassica oleracea protoplasts are treated with the drugs and deformed under an external electric potential. The relaxation processes of single protoplasts after electrodeformation are measured. The data are analyzed by a model-independent spectrum recovery algorithm. Two distinct characteristic time constants are obtained from the relaxation spectra. Treatment with latrunculin A increases both of the relaxation time constants. The longest relaxation times for control, latrunculin A treated, and jasplakinolide treated cells are determined to be 0.28, 1.0, and 0.21 s, respectively. PMID- 15652552 TI - The effect of wire plane tilt and olive wires on proximal tibia fracture fragment stability and fracture site motion. AB - To analyze the effect of the tilt angle relationship between the crossed wire plane and the bone axis on the stiffness of fine wire external fixation, load deformation behavior was compared across different tilt angles (0 degree, 10 degrees, and 20 degrees) of the plane containing crossed smooth or olive wires under identical conditions of central axial compression, medial compression bending, posterior compression-bending, posteromedial compression-bending, and torsion. Stiffness values were calculated from the load-deformation and torque angle curves. A tilt angle of 20 degrees with olive wires provided significantly greater stiffness compared to smooth wires at any angle in any loading condition (p < 0.05). A tilt angle of 20 degrees with olive wires was also significantly more stiff than a tilt angle of 0 degree with olive wires in any loading condition. In torsion, olive wires with 10 degrees and 20 degrees tilt were not significantly different, while in posterior bending olive wires with 10 degrees tilt were significantly stiffer than olive wires with 0 degree or 20 degrees tilt. With smooth wires, tilting the wire plane caused a decrease in stiffness in posterior bending, posteromedial bending, and torsion. Overall, the use of olive wires in conjunction with tilting the wire plane enhances the fixation stiffness for proximal tibia fractures while allowing more options for wire configurations that avoid neurovascular and musculotendinous structures, and wounds. PMID- 15652553 TI - 3-D computational modeling of media flow through scaffolds in a perfusion bioreactor. AB - Media perfusion bioreactor systems have been developed to improve mass transport throughout three-dimensional (3-D) tissue-engineered constructs cultured in vitro. In addition to enhancing the exchange of nutrients and wastes, these systems simultaneously deliver flow-mediated shear stresses to cells seeded within the constructs. Local shear stresses are a function of media flow rate and dynamic viscosity, bioreactor configuration, and porous scaffold microarchitecture. We have used the Lattice-Boltzmann method to simulate the flow conditions within perfused cell-seeded cylindrical scaffolds. Microcomputed tomography imaging was used to define the scaffold microarchitecture for the simulations, which produce a 3-D fluid velocity field throughout the scaffold porosity. Shear stresses were estimated at various media flow rates by multiplying the symmetric part of the gradient of the velocity field by the dynamic viscosity of the cell culture media. The shear stress algorithm was validated by modeling flow between infinite parallel plates and comparing the calculated shear stress distribution to the analytical solution. Relating the simulation results to perfusion experiments, an average surface shear stress of 5x10(-5)Pa was found to correspond to increased cell proliferation, while higher shear stresses were associated with upregulation of bone marker genes. This modeling approach can be used to compare results obtained for different perfusion bioreactor systems or different scaffold microarchitectures and may allow specific shear stresses to be determined that optimize the amount, type, or distribution of in vitro tissue growth. PMID- 15652554 TI - An alternative method of anthropometry of anterior cruciate ligament through 3-D digital image reconstruction. AB - Accurate and flexible measurements of length, area, and volume are important in evaluation of the mechanical properties of soft tissue. Although a number of contact-based and non-contact techniques have been reported in the literature, due to a variety of reasons such as cost, complexity, and low accuracy, the research community has not adopted a standardized technique. In this paper, an alternative method of measuring the geometric parameters of cadaver anterior cruciate ligament (ACL) is presented. In this method, a 3-D scan of the ACL is constructed using a simple, commercially available, scanning system. The 3-D scan is then analyzed using the 3-D Doctor Software to extract important information regarding the length, cross-sectional area, and volume of the ACL. The accuracy and repeatability of measurements obtained by this method are acceptable and comparable to existing non-contact methods. The limitation of the method is that surface concavities cannot be detected. However, the non-contact optical method, described here, has inherent advantages over the existing methods: (1) it is inexpensive; (2) it allows the determination of area at any distance along the length of the tissue of interest; (3) all relevant information including minimum area is extracted from one single application of the method; (4) the volume can be calculated with a simple additional step of length measurement although, for accurate results, condylar blockage must be minimized by coring the ACL out. The entire process of scanning takes less than 30 min. This technique has the potential to become a standard method in anthropometry of soft tissue. PMID- 15652555 TI - Mechanical damage to the intervertebral disc annulus fibrosus subjected to tensile loading. AB - Damage of the annulus fibrosus is implicated in common spinal pathologies. The objective of this study was to obtain a quantitative relationship between both the number of cycles and the magnitude of tensile strain resulting in damage to the annulus fibrosus. Four rectangular tensile specimens oriented in the circumferential direction were harvested from the outer annulus of 8 bovine caudal discs (n = 32) and subjected to one of four tensile testing protocols: (i) ultimate tensile strain (UTS) test; (ii) baseline cyclic test with 4 series of 400 cycles of baseline cyclic loading (peak strain = 20% UTS); (iii & iv) acute and fatigue damage cyclic tests consisting of 4 x 400 cycles of baseline cyclic loading with intermittent loading to 1 and 100 cycles, respectively, with peak tensile strain of 40%, 60%, and 80% UTS. Normalized peak stress for all mechanically loaded specimens was reduced from 0.89 to 0.11 of the baseline control levels, and depended on the magnitude of damaging strain and number of cycles at that damaging strain. Baseline, acute, and fatigue protocols resulted in permanent deformation of 3.5%, 6.7% and 9.6% elongation, respectively. Damage to the laminate structure of the annulus in the absence of biochemical activity in this study was assessed using histology, transmission electron microscopy, and biochemical measurements and was most likely a result of separation of annulus layers (i.e., delamination). Permanent elongation and stress reduction in the annulus may manifest in the motion segment as sub-catastrophic damage including increased neutral zone, disc bulging, and loss of nucleus pulposus pressure. The preparation of rectangular tensile strip specimens required cutting of collagen fibers and may influence absolute values of results, however, it is not expected to affect the comparisons between loading groups or dose-response reported. PMID- 15652556 TI - Mechanics of the ankle and subtalar joints revealed through a 3D quasi-static stress MRI technique. AB - A technique to study the three-dimensional (3D) mechanical characteristics of the ankle and of the subtalar joints in vivo and in vitro is described. The technique uses an MR scanner compatible 3D positioning and loading linkage to load the hindfoot with precise loads while the foot is being scanned. 3D image processing algorithms are used to derive from the acquired MR images bone morphology, hindfoot architecture, and joint kinematics. The technique was employed to study these properties both in vitro and in vivo. The ankle and subtler joint motion and the changes in architecture produced in response to an inversion load and an anterior drawer load were evaluated. The technique was shown to provide reliable measures of bone morphology. The left-to-right variations in bone morphology were less than 5%. The left-to-right variations in unloaded hindfoot architecture parameters were less than 10%, and these properties were only slightly affected by inversion and anterior drawer loads. Inversion and anterior drawer loads produced motion both at the ankle and at the subtalar joint. In addition, high degree of coupling, primarily of internal rotation with inversion, was observed both at the ankle and at the subtalar joint. The in vitro motion produced in response to inversion and anterior drawer load was greater than the in vivo motion. Finally, external motion, measured directly across the ankle complex, produced in response to load was much greater than the bone movements measured through the 3D stress MRI technique indicating the significant effect of soft tissue and skin interference. PMID- 15652557 TI - Passive and active mechanical properties of the superficial and deep digital flexor muscles in the forelimbs of anesthetized Thoroughbred horses. AB - The superficial (SDF) and deep digital flexor (DDF) muscles are critical for equine forelimb locomotion. Knowledge of their mechanical properties will enhance our understanding of limb biomechanics. Muscle contractile properties derived from architectural-based algorithms may overestimate real forces and underestimate shortening capacity because of simplistic assumptions regarding muscle architecture. Therefore, passive and active (=total - passive) force length properties of the SDF and DDF muscles were measured directly in vivo. Muscles from the right forelimbs of four Thoroughbred horses were evaluated during general anesthesia. Limbs were fixed to an external frame with the muscle attached to a linear actuator and load cell. Each muscle was stretched from an unloaded state to a range of prefixed lengths, then stimulated while held at that length. The total force did not exceed 4000 N, the limit for the clamping device. The SDF and DDF muscles produced 716+/-192 and 1577+/-203 N maximum active isometric force (F(max)), had ascending force-length ranges (R(asc)) of 5.1+/-0.2 and 9.1+/-0.4 cm, and had passive stiffnesses of 1186+/-104 and 1132+/-51 N/cm, respectively. The values measured for F(max) were much smaller than predicted based on conservative estimates of muscle specific tension and muscle physiological cross-sectional area. R(asc) were much larger than predicted based on muscle fiber length estimates. These data suggest that accurate prediction of the active mechanical behavior of architecturally complex muscles such as the equine DDF and SDF requires more sophisticated algorithms. PMID- 15652558 TI - An inhomogeneous and anisotropic constitutive model of human dentin. AB - Dentin constitutes the major part of human tooth. It is composed of a large number of tubules with both variational radii and radially parallel pattern. In addition, peritubular dentin surrounds each tubule lumen and has a higher elastic modulus than the matrix of dentin, i.e. intertubular dentin. Considering the above microstructural characteristics, a micromechanics model is used in this paper to evaluate the overall elastic properties of dentin. Five independent effective elastic parameters in transverse isotropic elasticity matrix can be expressed analytically by the material parameters of peri- and intertubular dentin and the volume fraction of tubules. To determine the effectivity of this theoretical model, a finite element (FE) model simulating a longitudinal tooth slice in moire fringe testing of Wang and Weiner (J. Biomech. 31 (1998) 135) was performed. Furthermore, the FE model was developed incorporate modeling of variation of tubule's diameter and softer characteristic of intertubular dentin near the dentin-enamel junction and around the pulp chamber. It turned out that the isoline figure of longitudinal displacement by FE calculation has very similar patterns to the moire fringe results. However, the FE results of displacement by traditional stress-strain models which regard dentin as a homogeneous and isotropic material show an obviously different strain distributions as compared to published moire fringes results. Thus the inhomogeneous and anisotropic model developed in this paper more accurately reflects the true physical nature of human dentin. PMID- 15652559 TI - Load-shift--numerical evaluation of a new design philosophy for uncemented hip prostheses. AB - All hip replacement prostheses alter the load transfer from the hip joint into the femur by changing the mechanical loading of the proximal femur from an externally to an internally loaded system. This alteration of the load transfer causes stress shielding and might lead to severe bone loss, especially with uncemented prostheses. To minimize these effects, load transfer to the femur should occur as proximal as possible. In order to support sufficient primary stability, however, directly post operative (PO) distal stabilization is reasonable. Consequently, the prostheses have to alter its mechanical characteristics after implantation. This concept is referred to as load-shift concept. Primary stability during the early PO state is provided by a prosthesis shaft, which is widened at the tip by a biodegradable pin. After resorption of the pin load transfer occurs no longer distally. The objective of this study was the numerical evaluation of the load-shift concept. The analysis was performed with a finite element model. Three-dimensional non-linear dynamic gait analyses data were used to evaluate whether the load transfer during walking can be altered effectively by insertion and resorption of a distal pin. Directly PO 38% of the transverse load is transferred through the prosthesis shaft and micromotion of the proximal prostheses tip is below 55 microm. After resorption of the pin, no transverse loads are transferred through the prosthesis shaft. Therefore, the loading of the proximal bone tissue is far more pronounced than in the case of a standard prosthesis, demonstrating the feasibility of the load shift concept. A balanced degradation of the pin simultaneously with the ingrowth of the prosthesis is expected to reduce hip replacement complications. PMID- 15652560 TI - Cross sectional area measurement of tendon and ligament in vitro: a simple, rapid, non-destructive technique. AB - The structural properties of tendons and ligaments are integral to their ability to function effectively in vivo and are determined from both the geometrical form and material properties. Many studies of tendon and ligament include a mechanical assessment of the structure in vitro. However, to determine the material properties of the constituent tissues it is necessary to measure cross-sectional area (CSA). Problems associated with this include damage to the structure, inaccurate values for non-uniform shapes, labour intensive techniques or requirement of expensive equipment. We describe a non-destructive technique for measurement of tendon and ligament CSA based on that of Race and Amis (J. Biomech. 29 (1996) 1207). The modified technique uses aqueous rapid curing alginate dental impression paste, digital photography and computerised image analysis. This technique is quick and simple to carry out and provides accurate values (within 0.8%) for CSA which are reproducible (coefficient of variation = 1.42%). The technique is non-destructive and can be used on specimens with an irregular shape. PMID- 15652561 TI - Frequency and length-dependent effects of Botulinum toxin-induced muscle weakness. AB - While the pathogenesis of Botulinum toxin type A (BTX-A)-induced muscle weakness has been systematically researched, little is known about the effects of motor fibre paralysis on the mechanical properties of skeletal muscle. Here, the long term effect of BTX-A injection on the force-length and force-frequency properties of rabbit knee extensors is investigated. BTX-A-induced muscle weakness was greater at short compared to long muscle lengths and at low compared to high stimulation frequencies four weeks following intervention. Therefore, we conclude that the paralysing effects of BTX-A are not uniform, and must be considered in biomechanical models of musculoskeletal rehabilitation in which BTX-A is used therapeutically, as for example in patients with cerebral palsy. Although the present results could be explained through a variety of mechanisms, this study does not allow for drawing firm conclusions about the length and frequency dependent effects of BTX-A. PMID- 15652562 TI - Mechanics of coracoacromial ligament transfer augmentation for acromioclavicular joint injuries. AB - The objective of this study was to determine how effectively the Weaver-Dunn repair (both unaugmented and augmented with a suture and suture anchor) restores joint translation in response to applied loads to normal. Translation of a reference point on the clavicle relative to a reference point on the acromion was assessed in five cadaver shoulders by applying anterior, posterior and superior loads of 50 N to the clavicle using a specially designed test rig while measuring movement of the acromion and clavicle with an optical measurement system. Translation was determined for the intact joint, after simulated injury and Weaver-Dunn repair, and after augmentation of the Weaver-Dunn repair with a suture fixed to a suture anchor in the coracoid process. Joints were significantly more mobile after Weaver-Dunn repair (16.1 mm anterior, 15.7 mm posterior, 11.1 mm superior) than when intact (4.1 mm anterior, 3.2 mm posterior, 4.0 mm superior) (p < 0.005). Augmentation with a suture and suture anchor reduced separation of the Weaver-Dunn reconstruction significantly (to 5.3 mm anterior, 4.1 mm posterior, 2.0 mm superior) (p < 0.005). Joints reconstructed using an augmented Weaver-Dunn repair were not significantly more (or less) mobile than normal joints (p > 0.005), although the power of the test to detect this difference was low (power = 0.107). We anticipate that, when surgery is indicated for treating acromioclavicular joint injury, an augmented Weaver-Dunn reconstruction will yield a joint that is less painful, more functional and less likely to require revision. PMID- 15652564 TI - How early reactions in the support limb contribute to balance recovery after tripping. AB - Tripping causes a forward angular momentum that has to be arrested to prevent a fall. The support limb, contralateral to the obstructed swing limb, can contribute to an adequate recovery by providing time and clearance for proper positioning of the recovery limb, and by restraining the angular momentum of the body during push-off. The present study investigated how such a contribution is achieved by the support limb in terms of response times and muscle moment generation, in order to provide more insight in the requirements for successful recovery after tripping. Twelve young adults repeatedly walked over a platform in which 21 obstacles were hidden. Each subject was tripped over one of these obstacles during mid-swing in at least five trials. Kinematics, dynamics and muscle activity were measured. Very rapid responses were seen in the muscles of the support limb (approximately 65 ms), causing fast increases in muscle moments in the joints during the primary phase of recovery. Especially a large ankle plantar flexion moment (204 Nm), a knee flexion moment (-54 Nm) and a hip extension moment (52 Nm), generated by triceps surae and hamstring muscle activity, brought about the necessary push-off reaction and simultaneously caused a restraining of the forward angular momentum of the body. These required joint moments could be a problem for the elderly, who might not be able to generate such powerful moments. Strength training in these muscle groups may be indicated in elderly subjects to reduce the risk of falling after a trip. PMID- 15652563 TI - Determination of patient-specific multi-joint kinematic models through two-level optimization. AB - Dynamic patient-specific musculoskeletal models have great potential for addressing clinical problems in orthopedics and rehabilitation. However, their predictive capability is limited by how well the underlying kinematic model matches the patient's structure. This study presents a general two-level optimization procedure for tuning any multi-joint kinematic model to a patient's experimental movement data. An outer level optimization modifies the model's parameters (joint position and orientations) while repeated inner level optimizations modify the model's degrees of freedom given the current parameters, with the goal of minimizing errors between model and experimental marker trajectories. The approach is demonstrated by fitting a 27 parameter, three dimensional, 12 degree-of-freedom lower-extremity kinematic model to synthetic and experimental movement data for isolated joint (hip, knee, and ankle) and gait (full leg) motions. For noiseless synthetic data, the approach successfully recovered the known joint parameters to within an arbitrarily tight tolerance. When noise was added to the synthetic data, root-mean-square (RMS) errors between known and recovered joint parameters were within 10.4 degrees and 10 mm. For experimental data, RMS marker distance errors were reduced by up to 62% compared to methods that estimate joint parameters from anatomical landmarks. Optimized joint parameters found using a loaded full-leg gait motion differed significantly from those found using unloaded individual joint motions. In the future, this approach may facilitate the creation of dynamic patient-specific musculoskeletal models for predictive clinical applications. PMID- 15652565 TI - Simulation of a knee joint replacement during a gait cycle using explicit finite element analysis. PMID- 15652567 TI - Letter to the editor. Hip simulator wear testing according to the newly introduced standard ISO 14242. PMID- 15652569 TI - Biochemical characterization of elongase activity in corn (Zea mays L.) roots. AB - Chemical analysis of 4-day-old corn (Zea mays L.) root cell walls revealed that the lipophilic biopolymer suberin forms an important constituent of rhizodermal and hypodermal cell walls. Identified aliphatic monomers had chain lengths ranging from C16 to C26 and they belonged to 5 substance classes (omega hydroxycarboxylic acids, 1,omega-dicarboxylic acids, 2-hydroxycarboxylic acids, carboxylic acids and alcohols) by which suberin is characterized. Biochemical experiments proved the occurrence of elongase activities in corn roots. Highest enzymatic activities were found in corn root microsomes, and major products synthesized by root elongases were elongated fatty acids with chain lengths ranging from C20 to C24. Preferred substrates of root elongases were acyl-CoAs of the chain length C18 and C20, whereas monounsaturated acyl-CoAs (C16:1 and C18:1) and acyl-CoAs of lower (C12-C16) and higher chain lengths (C22-C24) were rarely elongated. Elongase activities significantly decreased over the length (40 cm) of 10-day-old corn roots going from the young tip to the older base of the root. Thus, results presented here show the presence and activity of elongases in roots of plants. PMID- 15652570 TI - Biosynthesis of the hyperforin skeleton in Hypericum calycinum cell cultures. AB - Hyperforin is an important antidepressant constituent of Hypericum perforatum (St. John's wort). Cell cultures of the related species H. calycinum were found to contain the homologue adhyperforin and to a low extent hyperforin, when grown in BDS medium in the dark. Adhyperforin formation paralleled cell culture growth. Cell-free extracts from the cell cultures contained isobutyrophenone synthase activity catalyzing the condensation of isobutyryl-CoA with three molecules of malonyl-CoA to give phlorisobutyrophenone, i.e. the hyperforin skeleton. The formation of the hyperforins during cell culture growth was preceded by an increase in isobutyrophenone synthase activity. The cell cultures also contained benzophenone synthase and chalcone synthase activities which are involved in xanthone and flavonoid biosyntheses, respectively. The three type III polyketide synthases were separated by anion exchange chromatography. PMID- 15652571 TI - Purification and characterisation of adenosine nucleosidase from Coffea arabica young leaves. AB - An adenosine nucleosidase (ANase) (EC 3.2.2.7) was purified from young leaves of Coffea arabica L. cv. Catimor. A sequence of fractionating steps was used starting with ammonium sulphate salting-out, followed by anion exchange, hydrophobic interaction and gel filtration chromatography. The enzyme was purified 5804-fold and a specific activity of 8333 nkat mg-1 protein was measured. The native enzyme is a homodimer with an apparent molecular weight of 72 kDa estimated by gel filtration and each monomer has a molecular weight of 34.6 kDa, estimated by SDS-PAGE. The enzyme showed maximum activity at pH 6.0 in citrate-phosphate buffer (50 mM). The calculated Km is 6.3 microM and Vmax 9.8 nKat. PMID- 15652572 TI - Purification and properties of a beta-galactosidase from carambola fruit with significant activity towards cell wall polysaccharides. AB - beta-Galactosidase (EC. 3.2.1.23) from ripe carambola (Averrhoa carambola L. cv. B10) fruit was fractionated through a combination of ion exchange and gel filtration chromatography into four isoforms, viz. beta-galactosidase I, II, III and IV. This beta-galactosidases had apparent native molecular masses of 84, 77, 58 and 130 kDa, respectively. beta-Galactosidase I, the predominant isoform, was purified to electrophoretic homogeneity; analysis of the protein by SDS-PAGE revealed two subunits with molecular masses of 48 and 36 kDa. N-terminal amino acid sequence of the respective polypeptides shared high similarities albeit at different domains, with the deduced amino acid sequence of certain plant beta galactosidases, thus, explaining the observed low similarity between the two subunits. beta-Galactosidase I was probably a heterodimer that have glycoprotein properties and a pI value of 7.2, with one of the potential glycosylation sites appeared to reside within the 48-kDa-polypeptide. The purified beta-galactosidase I was substantially active in hydrolyzing (1-->4)beta-linked spruce and a mixture of (1-->3)beta- and (1-->6)beta-linked gum arabic galactans. This isoform also had the capability to solubilize and depolymerize structurally intact pectins as well as to modify alkaline-soluble hemicelluloses, reflecting in part changes that occur during ripening. PMID- 15652573 TI - Cyanogenic glucosides in grapevine: polymorphism, identification and developmental patterns. AB - Twelve grapevine (Vitis vinifera L.) cultivars were surveyed for 'cyanide potential' (i.e. the total cyanide measured in beta-glucosidase-treated crude, boiled tissue extract) in mature leaves. Two related cultivars (Carignan and Ruby Cabernet) had mean cyanide potential (equivalent to 110 mgHCNkg-1fr.wt) ca. 25 fold greater than that of the other 10 cultivars, and so the trait is polymorphic in the species. In boiled leaf extracts of Carignan and Ruby Cabernet, free cyanide constituted a negligible fraction of the total cyanide potential because beta-glucosidase treatment was required to liberate the major cyanide fraction - which is therefore bound in glucosylated cyanogenic compound(s) (or cyanogenic glucosides). In addition, cyanide was liberated from ground leaf tissue of Ruby Cabernet but not Sultana (a cultivar with low cyanide potential). Hence, the high cyanide potential in Ruby Cabernet leaves is coupled with endogenous beta glucosidase(s) activity and this cultivar may be considered 'cyanogenic'. A method was developed to detect and identify cyanogenic glucosides using liquid chromatography combined with tandem mass spectrometry (LC-MS/MS). Two putative cyanogenic glucosides were found in extracts from leaves of Carignan and Ruby Cabernet and were identified as the epimers prunasin and sambunigrin. Cyanide potential measured at three times over the growing season in young and mature leaves, petioles, tendrils, flowers, berries, seeds and roots of Ruby Cabernet was substantially higher in the leaves compared with all other tissues. This characterisation of cyanogenic glucoside accumulation in grapevine provides a basis for gauging the involvement of the trait in interactions of the species with its pests and pathogens. PMID- 15652574 TI - Quantification of glucosinolates in leaves of leaf rape (Brassica napus ssp. pabularia) by near-infrared spectroscopy. AB - The potential of near-infrared spectroscopy (NIRS) for screening the total glucosinolate (t-GSL) content, and also, the aliphatic glucosinolates gluconapin (GNA), glucobrassicanapin (GBN), progoitrin (PRO), glucoalyssin (GAL), and the indole glucosinolate glucobrassicin (GBS) in the leaf rape (Brassica napus L. ssp. pabularia DC), was assessed. This crop is grown for edible leaves for both fodder and human consumption. In Galicia (northwestern Spain) it is highly appreciated for human nutrition and have the common name of "nabicol". A collection of 36 local populations of nabicol was analysed by NIRS for glucosinolate composition. The reference values for glucosinolates, as they were obtained by high performance liquid chromatography on the leaf samples, were regressed against different spectral transformations by modified partial least squares (MPLS) regression. The coefficients of determination in cross-validation (r2) shown by the equations for t-GSL, GNA, GBN, PRO, GAL and GBS were, respectively, 0.88, 0.73, 0.81, 0.78, 0.37 and 0.41. The standard deviation to standard error of cross-validation ratio, were for these constituents, as follows: t-GSL, 2.96; GNA, 1.94; GBN, 2.31; PRO, 2.11; GAL, 1.27, and GBS, 1.29. These results show that the equations developed for total glucosinolates, as well as those for gluconapin, glucobrassicanapin and progoitrin, can be used for screening these compounds in the leaves of this species. In addition, the glucoalyssin and glucobrassicin equations obtained, can be used to identify those samples with low and high contents. From the study of the MPLS loadings of the first three terms of the different equations, it can be concluded that some major cell components as protein and cellulose, highly participated in modelling the equations for glucosinolates. PMID- 15652575 TI - Synthesis of the flavour precursor, alliin, in garlic tissue cultures. AB - The path of synthesis of alkyl cysteine sulphoxides, or flavour precursors, in the Alliums is still speculative. There are two proposed routes for alliin biosynthesis, one is from serine and allyl thiol while the other is from glutathione and an allyl source via gamma glutamyl peptides. The routes have been investigated by exposing undifferentiated callus cultures of garlic and onion to potential pathway intermediates. After a period of incubation of 2 days the callus was extracted, and analysed for flavour precursors and related compounds by HPLC. Standards of alliin, isoallin and propiin were synthesised and their identity confirmed by HPLC and NMR. Putative intermediates selected included the amino acids serine and cysteine, as well as more complex intermediates such as allylthiol, allyl cysteine and glutathione. Both garlic and onion tissue cultures were able to synthesize alliin following incubation with allylthiol, and cysteine conjugates such as allyl cysteine. The ability of the tissue cultures to form alliin from intermediates was compatible with the proposed routes of synthesis of alliin. PMID- 15652576 TI - Distribution of the fungal endophyte Neotyphodium lolii is not a major determinant of the distribution of fungal alkaloids in Lolium perenne plants. AB - The relationships of the distributions of the insect and mammalian mycotoxins, lolitrem B and ergovaline, and the insect-feeding deterrent, peramine, with the distribution of fungal mycelium were investigated in three genotypes of perennial ryegrass (Lolium perenne L.) infected with the endophyte Neotyphodium lolii. In planta levels and distribution of the endophyte and of the three alkaloids were assessed in parallel, and different spatial or temporal concentration gradients were observed for each. Variation in the tissue distribution of the endophyte accounted only for 20%, 6%, and 31% of the variation in ergovaline, lolitrem B, and peramine, respectively. Alkaloid-endophyte ratios, determined in individual grass tissues, showed distinct in planta distribution patterns for each alkaloid and differed in magnitude among genotypes. The ergovaline-endophyte ratio was higher in the very basal plant tissues than in the apical tissues, while the lolitrem B and peramine ratios tended to be higher in apical tissues. The lolitrem B-endophyte ratio increased with leaf age, while no consistent temporal trends were detected for the other alkaloids. The results indicate that endophyte colonisation is a minor determinant of alkaloid levels, and that accumulation of the alkaloids relative to the endophyte mycelium is affected by plant genotype and tissue in a manner specific to each alkaloid. Possible factors in the regulation of alkaloid levels in the grass plant are discussed. PMID- 15652577 TI - Qualitative and quantitative analyses of flower scent in Silene latifolia. AB - The quantitative and qualitative variability in floral scent of 98 specimens of the dioecious species Silene latifolia belonging to 15 European and 19 North American populations was determined. Floral scent was collected from single flowers using dynamic headspace methods, and analysed by Micro-SPE and GC-MS methods. The flowers showed a nocturnal rhythm, and scent was emitted only at night. The amount of emitted volatiles varied greatly during the season, from 400 ng/flower/2 min in June to 50 ng/flower/2 min in August and September. The qualitative variability in the floral scent was high and different chemotypes, characterised by specific scent compounds, were found. Female and male flowers emitted the same type and amount of volatiles. The differences in floral scent composition between European and North American populations were small. Typical compounds were isoprenoids like lilac aldehyde isomers, or trans-beta-ocimene, and benzenoids like benzaldehyde, phenyl acetaldehyde, or veratrole. Some of these compounds are known to attract nocturnal Lepidoptera species. The high qualitative variability is discussed in relation to the pollination biology of S. latifolia, and the results are compared with other studies investigating intraspecific variability of flower scent. PMID- 15652578 TI - Differential distribution of leaf chemistry in eucalypt seedlings due to variation in whole-plant nutrient availability. AB - We investigated the effects of whole-plant nutrient-availability on the degree of distribution of some plant primary and secondary (nitrogen, fibre, flavonols, gallotannins and cineole) chemicals across young, mature and old leaves of seedlings of Eucalyptus nitens. Four treatments that ranged from low to high nutrient-application rates resulted in mean whole-plant foliar concentrations of 0.63%, 0.85%, 1.11% and 1.82% nitrogen dry matter (N%DM) for treatments A, B, C and D, respectively. Within-plant distribution (across the leaf age profile of young, mature and old leaves within a eucalypt seedling) of N%DM ranged from zero in treatment A to a wide range of distribution in treatment D (low N%DM concentrations in old leaves to high N%DM concentrations in young leaves). Similarly, the distribution of fibre ranged from zero in treatment A to a wide range of distribution in treatment D, but with high concentrations in old leaves and low concentrations in young leaves. In contrast, flavonols (weakly) and gallotannins had a wide range of distribution in treatment A (low concentrations in old leaves to high concentrations in young leaves) but were little or not distributed in the other treatments. Again in contrast, cineole was strongly distributed between old and young leaves (low concentrations in old leaves to high concentrations in young leaves) across all treatments while concentrations in mature leaves reflected one or other leaf age depending on treatment. Protein precipitable phenols in treatments A, B and C were high in young, and low in old leaves; whereas in treatment D they were low and similar between leaves of different ages. PMID- 15652579 TI - Ipangulines and minalobines, chemotaxonomic markers of the infrageneric Ipomoea taxon subgenus Quamoclit, section Mina. AB - A comprehensive GC-MS analysis of 8 Ipomoea species belonging to the subgenus Quamoclit, section Mina revealed that the members of this taxon form combinations of two necine bases with rare necic acids resulting in unique pyrrolizidine alkaloids. The occurrence and diversity of these metabolites show remarkable variations: Some species, especially Ipomoea hederifolia and Ipomoea lobata, are able to synthesize a large number of alkaloids whereas others, especially Ipomoea coccinea and Ipomoea quamoclit, are poor synthesizers with only a few compounds. However, these metabolites are apparently chemotaxonomic markers of this infrageneric taxon in general. They represent either esters of (-)-platynecine (altogether 48 ipangulines and 4 further esters including results of a previous study) or esters of (-)-trachelanthamidine, an additional novel structural type called minalobines (altogether 21 alkaloids). Both types are characterized by section-specific rare necic acids, e.g., ipangulinic/isoipangulinic acid, phenylacetic acid. The alkaloids of Ipomoea cholulensis, I. coccinea, I. hederifolia, Ipomoea neei, and Ipomoea quamoclit were mono and diesters of platynecine. Minalobines turned out to be metabolites of I. lobata (Cerv.) Thell. (syn.: Mina lobata Cerv.) lacking ipangulines. The major alkaloid of this species, minalobine R, has been isolated and identified as 9-O-(threo-2-hydroxy-2 methyl-3-phenylacetoxy-butyryl)-(-)-trachelanthamidine on the basis of spectral data. Apparently only two of the species included in this study, Ipomoea cristulata and Ipomoea sloteri, are able to synthesize both, ipangulines as well as minalobines. Minalobine O could be isolated as a major alkaloid of I. cristulata, its structure has been established as 9-O-(erythro-2-hydroxy-2-methyl 3-tigloyloxy-butyryl)-(-)-trachelanthamidine on the basis of spectral data. PMID- 15652580 TI - Bioactive constituents of Artemisia monosperma. AB - During a study on the chemistry and biological activity of Kuwaiti plants, new metabolites including 4,6-dihydroxy-3-[3'-methyl-2'-butenyl]-5-[4''-hydroxy-3'' methyl-2''-butenyl]-cinnamic acid (1), the 3R,8R stereoisomer of the C17 polyacetylene dehydrofalcarindiol (2) and a C10 polyacetylene glucoside (3) were characterised by spectroscopic means. Additionally, the previously characterised natural products 1,3R,8R-trihydroxydec-9-en-4,6-yne (4), spathulenol (5) and eriodyctiol-7-methyl ether (6) were also isolated. Compounds 2, 3, and 4 were evaluated for their ability to inhibit the enzyme 12-lipoxygenase and 3 and 4 showed moderate activity at 30 microg/ml. Compound 2 was evaluated against a panel of colorectal and breast cancer cell lines and IC50 values ranged from 5.8 to 37.6 microg/ml. Against a panel of fast-growing mycobacteria and a standard ATCC strain of Staphylococcus aureus, compound 6 exhibited minimum inhibitory concentrations in the range of 64-128 microg/ml. PMID- 15652581 TI - Bioactive ellagitannins from Cunonia macrophylla, an endemic Cunoniaceae from New Caledonia. AB - Chemical study of Cunonia macrophylla, a New Caledonian Cunoniaceae, based on bioactive effects of a crude methanol extract of the leaves, detected bioactive tannins for the first time in this plant family. These ellagitannins have been identified as ellagic acid-4-O-beta-D-xylopyranoside (6), mallorepanin (3), mallotinic acid (1) along with corilagin (2), chebulagic acid (4), ellagic acid (5) and gallic acid (7) and have been shown to possess antimicrobial activity and to inhibit xanthine oxidase. Antimicrobial effects on bacterial human pathogens (Staphylococcus aureus, Corynebacterium accolans) and on a plant pathogen (Erwinia carotovora) as well as on a human pathogenic yeast (Candida albicans) were investigated. Activity is reported here for the first time for compounds 1, 3, 4 and 6. The inhibitory effects of all molecules against xanthine oxidase in relation to their structure was evaluated and compared. Compound 6 presented the best activity and seems to be of considerable interest for further studies. PMID- 15652582 TI - Sesquiterpenes from Cupressus macrocarpa foliage. AB - Ten sesquiterpenes, many with unusual carbon skeletons, were identified in foliage of Cupressus macrocarpa. These are (-)-10-epi-beta-acoradiene; ent-widdra 2,4(14)-diene; (E)-iso-gamma-bisabolene, i.e., (4E)-4-(1,5-dimethylhex-5 enylidene)-1-methylcyclohexene; (-)-cumacrene, i.e., (4S)-4-[(1R,2S)-2 isopropenyl-1-methylcyclobutyl]-1-methylcyclohexene; (-)-alpha-chamipinene, i.e., (1S,6S,7S)-2,2,6,8-tetramethyltricyclo[5.3.1.01,6]undec-8-ene; and five sesquiterpenes with a 3,3,4'-trimethyl-1,1'-bi(cyclohexyl) skeleton for which the trivial name macrocarpane is proposed. The possible single-enzyme biogenesis of these sesquiterpenes is discussed. PMID- 15652583 TI - What does the EU do for its citizens' health? PMID- 15652584 TI - Hopes and fears for rotavirus vaccines. PMID- 15652585 TI - Threats to human survival: a WIRE to warn the world. PMID- 15652586 TI - Homocysteine and stroke. PMID- 15652587 TI - Effect of malaria on HIV-1 progression and transmission. PMID- 15652588 TI - NOTCH1 pathway: a molecular target in T-cell cancers? PMID- 15652589 TI - Gene-expression profiling: time for clinical application? PMID- 15652590 TI - Preventing hospitalisations for children. PMID- 15652594 TI - Links with the tobacco industry. PMID- 15652595 TI - Possible explanations for the results of CRASH. PMID- 15652596 TI - Possible explanations for the results of CRASH. PMID- 15652598 TI - Possible explanations for the results of CRASH. PMID- 15652599 TI - Clinical predictors of bioterrorism-related inhalational anthrax. PMID- 15652600 TI - Clinical predictors of bioterrorism-related inhalational anthrax. PMID- 15652601 TI - China's infamous one-child policy. PMID- 15652603 TI - Greek kleroteria: the first randomisation technique? PMID- 15652604 TI - Global burden of hypertension: analysis of worldwide data. AB - BACKGROUND: Reliable information about the prevalence of hypertension in different world regions is essential to the development of national and international health policies for prevention and control of this condition. We aimed to pool data from different regions of the world to estimate the overall prevalence and absolute burden of hypertension in 2000, and to estimate the global burden in 2025. METHODS: We searched the published literature from Jan 1, 1980, to Dec 31, 2002, using MEDLINE, supplemented by a manual search of bibliographies of retrieved articles. We included studies that reported sex specific and age-specific prevalence of hypertension in representative population samples. All data were obtained independently by two investigators with a standardised protocol and data-collection form. RESULTS: Overall, 26.4% (95% CI 26.0-26.8%) of the adult population in 2000 had hypertension (26.6% of men [26.0 27.2%] and 26.1% of women [25.5-26.6%]), and 29.2% (28.8-29.7%) were projected to have this condition by 2025 (29.0% of men [28.6-29.4%] and 29.5% of women [29.1 29.9%]). The estimated total number of adults with hypertension in 2000 was 972 million (957-987 million); 333 million (329-336 million) in economically developed countries and 639 million (625-654 million) in economically developing countries. The number of adults with hypertension in 2025 was predicted to increase by about 60% to a total of 1.56 billion (1.54-1.58 billion). INTERPRETATION: Hypertension is an important public-health challenge worldwide. Prevention, detection, treatment, and control of this condition should receive high priority. PMID- 15652605 TI - Homocysteine and stroke: evidence on a causal link from mendelian randomisation. AB - BACKGROUND: Individuals homozygous for the T allele of the MTHFR C677T polymorphism have higher plasma homocysteine concentrations (the phenotype) than those with the CC genotype, which, if pathogenetic, should put them at increased risk of stroke. Since this polymorphism is distributed randomly during gamete formation, its association with stroke should not be biased or confounded. We investigated consistency between the expected odds ratio for stroke among TT homozygotes, extrapolated from genotype-phenotype and phenotype-disease studies, and the observed odds ratio from a meta-analysis of genotype-disease association studies. METHODS: We searched MEDLINE and EMBASE up to June, 2003, for all relevant studies on the association between homocysteine concentration and the MTHFR polymorphism, and until December, 2003, for those on the association between the polymorphism and the risk of stroke. Pooled odds ratios and 95% CI were calculated by random-effects and fixed-effects models. Consistency between expected and observed odds ratios was assessed by interaction test. FINDINGS: 111 studies met the selection criteria. Among 15635 people without cardiovascular disease, the weighted mean difference in homocysteine concentration between TT and CC homozygotes was 1.93 micromol/L (95% CI 1.38 to 2.47). The expected odds ratio for stroke corresponding to this difference based on previous observational studies was 1.20 (1.10 to 1.31). In our genetic meta-analysis (n=13928) the odds ratio for stroke was 1.26 (1.14 to 1.40) for TT versus CC homozygotes, similar to the expected odds ratio (p=0.29). Consistency between the odds ratios was preserved in analyses by age-group, ethnic background, and geographical location. INTERPRETATION: The observed increase in risk of stroke among individuals homozygous for the MTHFR T allele is close to that predicted from the differences in homocysteine concentration conferred by this variant. This concordance is consistent with a causal relation between homocysteine concentration and stroke. PMID- 15652606 TI - Effect of Plasmodium falciparum malaria on concentration of HIV-1-RNA in the blood of adults in rural Malawi: a prospective cohort study. AB - BACKGROUND: Raised HIV viral load in blood has been associated with accelerated disease progression and increased transmission of infection. To assess the effect of Plasmodium falciparum malaria on concentrations of HIV in blood, we did a prospective cohort study in Malawi. METHODS: We recruited 367 HIV-1-infected adults. Among 334 people aparasitaemic at baseline, 148 had at least one malaria episode during follow-up and received antimalarial treatment. Of these, 77 had HIV-1-RNA measurements at baseline, during malaria, and post-malaria. We used linear regression with generalised estimating equations to assess effect of four definitions of malaria (any parasitaemia, parasite density > or =2000/microL, febrile parasitaemia, and febrile parasitaemia with parasite density > or =2000/microl) on changes in log HIV-1 RNA, overall and by baseline CD4 count. FINDINGS: With malaria defined as any parasitaemia, HIV-1-RNA concentration almost doubled between baseline (median 96215 copies per mL) and malaria (168901 copies per mL), a 0.25 (95% CI 0.11-0.39) log increase in mean RNA concentration. HIV-1-RNA concentration fell to median 82058 copies per mL by about 8-9 weeks post-malaria. Increases in HIV-1-RNA were greatest for people with fever, parasite density 2000/microL or greater, and CD4 count more than 300 cells per muL, in whom concentrations rose from median 38483 copies per mL at baseline to 196098 copies per mL during malaria, a mean log increase of 0.82 (95% CI 0.55 1.10, p<0.0001), and fell to median 75331 copies per mL post-malaria. People who remained aparasitaemic showed no changes in HIV-1-RNA concentration. INTERPRETATION: HIV-infected individuals with malaria have a significantly increased viral load, which might enhance HIV transmission and accelerate disease progression. PMID- 15652607 TI - Porphyrias. AB - Seven different porphyrias form a group of inherited metabolic disorders, each resulting from a partial deficiency of a specific enzyme in the haem biosynthesis pathway. Clinically, the three most important entities are an acute porphyric attack and acute and chronic skin symptoms. Porphyrias are rare and sometimes misdiagnosed, because various symptoms and signs mimic other diseases. Once porphyria is suspected, biochemical analyses easily detect porphyrins and their precursors from blood, urine, or faeces. Mutation screening can be done at the quiescent phase of the disease. Pathogenetic mechanisms and clinical manifestations differ in individual porphyrias and most of them require a specific treatment. Early diagnosis and information about precipitating factors can diminish mortality and prevent subsequent attacks among patients with acute porphyrias, so mutation screening is recommended for family members. PMID- 15652608 TI - Human polymicrobial infections. AB - CONTEXT: Polymicrobial diseases, caused by combinations of viruses, bacteria, fungi, and parasites, are being recognised with increasing frequency. In these infections, the presence of one micro-organism generates a niche for other pathogenic micro-organisms to colonise, one micro-organism predisposes the host to colonisation by other micro-organisms, or two or more non-pathogenic micro organisms together cause disease. STARTING POINT: Recently, Gili Regev-Yochay (JAMA 2004; 292: 716-20) and Debby Bogaert (Lancet 2004; 363: 1871-72), and their colleagues, suggested another interaction: microbial interference-the ability of Streptococcus pneumoniae carriage to protect against Staphylococcus aureus carriage, and the inverse effect of pneumococcal conjugate vaccination on the increased carriage of Staph aureus and Staph-aureus-related disease. Strep pneumoniae carriage protected against Staph aureus carriage, and the bacterial interference could be disrupted by vaccinating children with pneumococcal conjugate vaccines that reduced nasopharyngeal carriage of vaccine-type Strep pneumoniae. WHERE NEXT: The medical community is recognising the significance of polymicrobial diseases and the major types of microbial community interactions associated with human health and disease. Many traditional therapies are just starting to take into account the polymicrobial cause of diseases and the repercussions of treatment and prevention. PMID- 15652609 TI - Treating individuals 3: from subgroups to individuals: general principles and the example of carotid endarterectomy. AB - Clinicians often have to make treatment decisions based on the likelihood that an individual patient will benefit. In this article we consider the relevance of relative and absolute risk reductions, and draw attention to the importance of expressing the results of trials and subgroup analyses in terms of absolute risk. We describe the limitations of univariate subgroup analysis in situations in which there are several determinants of treatment effect, and review the potential for targeting treatments with risk models, especially when benefit is probably going to be dependent on the absolute risk of adverse outcomes with or without treatment. The ability to systematically take into account the characteristics of an individual patient and their interactions, to consider the risks and benefits of interventions separately if needed, and to provide patients with personalised estimates of their likelihood of benefit is shown using the example of endarterectomy for symptomatic carotid stenosis. PMID- 15652610 TI - Lymphadenopathy, pneumonia, and HIV--a common trio, an uncommon outcome. PMID- 15652611 TI - Comparative analysis of inhibitory effects of caged ligands for the NMDA receptor. AB - Photolytic release of neurotransmitters from caged precursors is a useful method to study synaptic processes with high temporal and spatial resolution. At present, the two most widely used classes of caged precursors for studies on glutamate receptors are based on derivatives of the 2-nitrobenzyl caging group (alpha-carboxy-2-nitrobenzyl, CNB) and the nitroindoline caging group (7 nitroindoline, NI, and 4-methoxy-7-nitroindoline, MNI). Besides NI- and MNI-caged amino acids being thermally more stable than the CNB-caged amino acids, there have been no other major advantages reported of using compounds from either of these two classes. Here, we show inhibitory effects of CNB-glutamate and a number of other CNB-caged agonists on N-methyl-D-aspartate (NMDA) receptors at non saturating concentrations of the co-agonist glycine. In contrast, NI- and MNI glutamate and most other NI-/MNI-caged agonists that we tested were inert under these conditions. Furthermore, we demonstrate that carboxynitroindoline-caged glycine (CNI-glycine), which was previously found to inhibit glycine receptors, has no such effect on NMDA receptors. Together, these findings underline the usefulness of NI- and MNI-caged ligands and show that CNB-caged compounds should be avoided in studies involving NMDA receptors. PMID- 15652612 TI - Chronic implantation of deep brain stimulation leads in animal models of neurological disorders. AB - Deep brain stimulation (DBS) has routinely been used as a treatment option in Parkinson's disease (PD), tremor disorders and, more recently, dystonia. Here, we describe a method of implantation of DBS leads in the monkey model of PD. By adapting procedures used in human patients, we have devised implantation techniques that can be readily applied to any animal model in which stimulation of subcortical structures is desired. The procedure for implantation consists of microelectrode mapping of the target structure, DBS lead preparation and implantation, and verification of lead placement. The stimulation system described in this paper allows for simultaneous recording of neuronal activity (during stimulation) and observation of animal behavior without restriction of the subject's head or body. In addition, we detail techniques for stimulation and recording from distant structures (utilizing either a one or two chamber system) to facilitate examination of the effects of DBS on neural activity. Thus, the correlation of changes in neuronal activity with behavior during stimulation of subcortical structures can be accomplished. In addition, the use of leads in primates which are analogous in size to human devices allows for close reproduction of the effects of stimulation as observed in humans. PMID- 15652613 TI - Trimmed estimators for robust averaging of event-related potentials. AB - Averaging (in statistical terms, estimation of the location of data) is one of the most commonly used procedures in neuroscience and the basic procedure for obtaining event-related potentials (ERP). Only the arithmetic mean is routinely used in the current practice of ERP research, though its sensitivity to outliers is well-known. Weighted averaging is sometimes used as a more robust procedure, however, it can be not sufficiently appropriate when the signal is nonstationary within a trial. Trimmed estimators provide an alternative way to average data. In this paper, a number of such location estimators (trimmed mean, Winsorized mean and recently introduced trimmed L-mean) are reviewed, as well as arithmetic mean and median. A new robust location estimator tanh, which allows the data-dependent optimization, is proposed for averaging of small number of trials. The possibilities to improve signal-to-noise ratio (SNR) of averaged waveforms using trimmed location estimators are demonstrated for epochs randomly drawn from a set of real auditory evoked potential data. PMID- 15652614 TI - A software solution for the control of visual behavioral experimentation. AB - Psychophysical and neurophysiological research requires precise control of experimental devices for the purpose of delivering stimuli and monitoring behavioral and neural responses. This has previously been accomplished by complex, often proprietary, programmable systems, interfacing with a limited range of hardware. We have developed a software solution entirely within the Matlab environment that can achieve high-speed control of experimental and behavioral variables. We make this Wake-Forest Visual Experimentation (WaVE) software freely available under the GNU public license, and demonstrate how to customize it to individual laboratory needs. WaVE takes advantage of existing Matlab libraries and toolboxes to present visual stimuli, collect experimental data, update behavioral variables, and communicate with other computers. Although we have developed it for use in a Windows-based Personal Computer, the portability of the Matlab code makes possible its customization for use in a variety of other systems. We present simulation results showing sub-millisecond sampling rate and updating precision, running on single-processor, desktop PCs. The WaVE software offers a simple, flexible and powerful solution that compares favorably with many of its costly alternatives. PMID- 15652616 TI - Single neuronal recordings using surface micromachined polysilicon microelectrodes. AB - Bulk micromachining techniques of silicon have been used successfully in the past several years to microfabricate microelectrodes for monitoring single neurons in acute and chronic experiments. In this study we report for the first time a novel surface micromachining technique to microfabricate a very thin polysilicon microelectrode that can be used for monitoring single-unit activity in the central nervous system. The microelectrodes are 3 mm long and 50 microm x 3.75 microm in cross-section. Excellent signal to noise ratios in the order of 25-35 dB were obtained while recording neuronal action potentials. The microelectrodes successfully penetrated the brains after a microincision of the dura mater. Chronic implantation of the microprobe for up to 33 days produced only minor gliosis. Since the polysilicon shank acts as a conductor, additional processing steps involved in laying conductor lines on silicon substrates are avoided. Further, surface micromachining allows for fabricating extremely thin microelectrodes which could result in decreased inflammatory responses. We conclude that the polysilicon microelectrode reported here could be a complementary approach to bulk-micromachined silicon microelectrodes for chronic monitoring of single neurons in the central nervous system. PMID- 15652615 TI - A stimulation method using odors suitable for PET and fMRI studies with recording of physiological and behavioral signals. AB - A design for a semi-automatic olfactometric system is described for PET and fMRI experiments. The olfactometer presents several advantages because it enables the use of an 'infinite' number of odorants and the synchronization of stimuli with breathing. These advantages mean that the subject is recorded while breathing normally during olfactory judgment tasks. In addition, the design includes a system for recording the behavioral (rating scale) and physiological (breathing, electrodermal reaction (ED), plethysmography (PL)) signals given by the subject. Both systems present the advantage of being compatible with fMRI magnetic fields since no ferrous material is used in the Faraday cage and signals are transmitted via an optical transmission interface to an acquisition system. PMID- 15652617 TI - Applying small quantities of multiple compounds to defined locations of in vitro brain slices. AB - When studying in vitro brain slices, rapidly applying multiple agonists, antagonists, drugs, or modulatory compounds is a significant technical problem. There are three major ways that multiple compounds are applied to slices: by bath, via a U-tube device, or by pressure application using a "puffer" pipette. Each of these methods has advantages and disadvantages, making each more appropriate for particular purposes. Because puffer pipettes have a small, sharp tip, they are best suited to apply a small quantity of a compound to a well defined location within the slice. When used in this way, puffer pipettes have two shortcomings. Solution leaking from the tip of the pipette can contaminate the signal, and it is difficult to apply more than one test solution to exactly the same local area of the slice. We describe methods and newly designed devices aimed at overcoming those limitations. Relatively inexpensive approaches are described to apply eight different solutions to the same exact location deep within a brain slice. The validity of the approach is verified by measuring ligand-gated channel currents activated by glutamate (Glu), acetylcholine (ACh), and gamma-amino butyric acid (GABA). PMID- 15652618 TI - Quantitative comparison of algorithms for inter-subject registration of 3D volumetric brain MRI scans. AB - The objective of inter-subject registration of three-dimensional volumetric brain scans is to reduce the anatomical variability between the images scanned from different individuals. This is a necessary step in many different applications such as voxelwise group analysis of imaging data obtained from different individuals. In this paper, the ability of three different image registration algorithms in reducing inter-subject anatomical variability is quantitatively compared using a set of common high-resolution volumetric magnetic resonance imaging scans from 17 subjects. The algorithms are from the automatic image registration (AIR; version 5), the statistical parametric mapping (SPM99), and the automatic registration toolbox (ART) packages. The latter includes the implementation of a non-linear image registration algorithm, details of which are presented in this paper. The accuracy of registration is quantified in terms of two independent measures: (1) post-registration spatial dispersion of sets of homologous landmarks manually identified on images before or after registration; and (2) voxelwise image standard deviation maps computed within the set of images registered by each algorithm. Both measures showed that the ART algorithm is clearly superior to both AIR and SPM99 in reducing inter-subject anatomical variability. The spatial dispersion measure was found to be more sensitive when the landmarks were placed after image registration. The standard deviation measure was found sensitive to intensity normalization or the method of image interpolation. PMID- 15652619 TI - Audiophile hardware in vision science; the soundcard as a digital to analog converter. AB - The design objective was to develop an inexpensive digital to analog (D/A) converter for use in vision science. Soundcards are hardware units that can be integral or can be added to a computer to add sound capability. A soundcard contains D/A converters designed to work in the audio frequency range, typically 20-20,000 Hz. Soundcard outputs are high-pass filtered and thus do not convey sub audio frequency or dc information. It is possible to circumvent this design feature by programming the desired output waveform as an amplitude modulation of a high frequency carrier, and then demodulating the soundcard output. The circuit, using a 20 kHz carrier, provides precise D/A conversion for the frequency range relevant for vision experiments, dc to 100 Hz, using inexpensive readily available components. The specific application was for 8 channels of D/A conversion using a Macintosh computer running under OS X. The software needed to program stimuli was created using CoreAudio, a library for programming sounds in OS X. Using soundcards on other platforms would not be a problem, as long as there exists a low level library that would enable the wave table to be filled. PMID- 15652620 TI - Sensitivity of surface EMG-based conduction velocity estimates to local tissue in homogeneities--influence of the number of channels and inter-channel distance. AB - The aim of this simulation study was to investigate the influence of local tissue in-homogeneities on the estimates of muscle fiber conduction velocity (CV) from surface EMG signals. A recently developed analytical surface EMG model was used to generate simulated surface EMG signals from a planar layered volume conductor, comprised of the muscle tissue and fat layer, with spheres (1 mm radius) in the fat layer of conductivity different from the surrounding tissue. CV was estimated with a maximum likelihood multi-channel approach, varying the number of channels and the inter-channel distance used for the estimate. The action potentials detected along the muscle fiber direction changed shape due to the presence of the in-homogeneities, thus affecting CV estimates. CV estimates were influenced by the location of the in-homogeneities with respect to the fiber and detection electrodes. The maximum percent variation of CV estimates due to the presence of in-homogeneities decreased with increasing number of channels and inter-channel distance: 19.6% (2 channels), 12.1% (3 channels), 6.4% (4 channels), for 5 mm inter-channel distance, and 12.0% (2 channels), 5.2% (3 channels), 2.4% (4 channels), for 10 mm inter-channel distance (for double differential detection). The results were in agreement and explained previous experimental findings. It was concluded that multi-channel methods for CV estimation significantly reduce the sensitivity of CV estimates to tissue in-homogeneities. PMID- 15652621 TI - A new method for measuring blood-brain barrier permeability demonstrated with Europium-bound albumin during experimental lipopolysaccharide (LPS) induced meningitis in the rat. AB - The blood-brain barrier (BBB) is a critical extrameningeal site of injury during bacterial meningitis, manifested by enhanced BBB permeability (BBBP). Previous methods to measure altered BBBP during meningitis involve radioactive materials, or are poorly quantified. Europium (EU) is a fluorescent, non-radioactive metal that is a sensitive and stable marker. Europium fluorescence can be measured with a spectrophotometer capable of time-resolved fluorescence (TRF). We used EU albumin (EU-A) to examine BBBP in experimental lipopolysaccharide (LPS) induced meningitis. The results presented here introduce a simple and accurate method for measuring BBB permeability. PMID- 15652622 TI - An economical multi-channel cortical electrode array for extended periods of recording during behavior. AB - We report the development of a low-cost chronic multi-channel microwire electrode array for recording multi-unit cortical responses in behaving rodents. The design was motivated by three issues. First, standard connector systems tended to disconnect from the head-stage during extended periods of behavior. Disconnections resulted in a loss of data and an interruption of the animals' behavior. Second, the use of low insertion force connectors with locking mechanisms was cost prohibitive. Finally, connecting the head-stage to a skull mounted connector on an unrestrained animal was highly stressful for both the researcher and animal. The design developed uses a high insertion force DIP socket separated from the skullcap that prevents inadvertent disconnects, is inexpensive, and simplifies connecting unrestrained rodents. Electrodes were implanted in layer IV of primary auditory cortex in 11 Sprague-Dawley rats. Performance of the electrodes was monitored for 6 weeks. None of the behaving animals became disconnected from the recording system during recording sessions lasting 6 h. The mean signal-to-noise ratio on all channels (154) following surgery was 3.9+/-0.2. Of the 154 channels implanted, 130 exhibited driven activity following surgery. Forty percent of the arrays continued to exhibit driven neural activity at 6 weeks. PMID- 15652623 TI - A repetitive movement detector used for automatic monitoring and quantification of scratching in mice. AB - We have designed an economical non-invasive movement detector for small animal studies and used it for monitoring and quantifying itch in mice. The system is based on a sensitive force transducer positioned below a recording platform holding a lightweight polystyrene recording box in which an animal is placed. A programmed micro-controller is used to discriminate between non-specific movement, grooming behaviour, and scratching movements made by the animal's hind limb. Following sub-dermal injection of histamine receptor agonists into the neck of a mouse, dose-related scratching occurred which was detected and quantified. There was 91% correlation between bouts of scratching as counted manually from playback of the video recording and recorded by the detector. The detector was also able rapidly to count the individual scratch movements of the hind limb that comprise a bout, with 95% accuracy in comparison with manual counting during slow motion playback of video tape, something that is impossible for an unaided observer to achieve because individual scratch movements are too fast to discriminate by eye. Separate detectors were used for the efficient non-invasive study of four animals simultaneously, and this number could easily be increased by adding more platforms. The system could also be modified to record the animal's position within the box, which would be of value in studies involving exploratory behaviour. In summary, the non-invasive multichannel repetitive movement detector will be very useful for accurate measurement of scratching during pruritus studies in small animals, with considerable savings in staff time and effort. It should therefore be a valuable tool for helping to investigate pruritus and in the evaluation of anti-pruritic drugs. PMID- 15652624 TI - Functional magnetic resonance mapping of intracerebroventricular infusion of a neuroactive peptide in the anaesthetised rat. AB - Pharmacological magnetic resonance imaging (phMRI) methods map the cerebral haemodynamic response to challenge with psychotropic agents as a surrogate for drug-induced changes in brain activity. However, many neuroactive compounds present low blood-brain barrier penetration and thus systemic administration may result in insufficient brain concentration. Intracerebroventricular (ICV) administration has been long used as an effective way of bypassing the blood brain barrier in studies with non-brain-penetrant compounds, such as neuropeptides. In order to extend the range of pharmacological substances accessible to phMRI, we have developed methods to map relative cerebral blood volume (rCBV) changes induced by in situ ICV administration of neuroactive agents in the anaesthetised rat. We have applied this method to study for the first time the phMRI response to central administration of a neuropeptide, the metabolically stable and potent NK1 receptor agonist GR-73632. ICV administration of 4.2 pmol of GR-73632 produced a rapid onset and sustained rCBV increase in several brain structures, such as the amygdala, the caudate putamen and the cortex. These results demonstrate the feasibility of phMRI as a tool to study the functional correlates of brain activity induced by central administration of neuroactive agents. PMID- 15652625 TI - Maintenance of eupnea and gasping following alterations in potassium ion concentration of perfusates of in situ rat preparation. AB - Levels of extracellular potassium ion, above those in vivo, are required for the generation of rhythmic activities of in vitro preparations. Our purpose was to define whether hyperkalemia in the perfusate of an in situ preparation was likewise necessary for the expression of eupnea and gasping. Studies were performed using the in situ preparation of the juvenile rat, in which activity of the phrenic nerve was recorded as an index of the respiratory rhythm. Eupnea and gasping were impervious to modifications of potassium ion of the perfusate. Eupnea was maintained uninterrupted for more than 60 min whether the total concentration of potassium was hypokalemic (2.75 mM), normokalemic (4.0 mM), or hyperkalemic (6.25 and 7.75 mM). Gasping, with identical characteristics, was elicited at any concentration of potassium ion. We conclude that both eupnea and gasping are unaffected by modest changes in the concentration of potassium ion in the perfusate with which the in situ rat preparation is maintained. These results add further support to the conclusion that the in situ preparation represents a model that accurately reproduces mechanisms of rhythm generation of both eupnea and gasping in vivo. PMID- 15652626 TI - A flexible auditory research platform using acoustic or electric stimuli for adults and young children. AB - A user-friendly and versatile research platform for use in auditory experiments, referred to as APEX (Application for PsychoElectrical eXperiments), is described. The platform takes care of automatic stimulus presentation and collection of the subject's responses. Acoustical auditory, as well as electrical auditory experiments with CI recipients can be conducted. The platform currently supports LAURA, Nucleus CI22 and Nucleus CI24 cochlear implants. The graphical user interface for the subjects has been extended to allow for testing very young children, by embedding the psychophysical procedures in a computer game. The research platform is available free of charge. PMID- 15652627 TI - Effective gene delivery to adult neurons by a modified form of electroporation. AB - Non-viral methods of transfection of cDNAs into adult neurons and other post mitotic cells are generally very inefficient. However, the recent development of Nucleofector technology developed by Amaxa Biosystems allows direct delivery of cDNAs into the nucleus, enabling transfection of non-dividing cells. In this study, we describe a reliable method for culturing large numbers of retinal cells from adult rats and using Nucleofection, we were able to transfect cDNA-encoding GFP (jellyfish green fluorescent protein) into retinal ganglion cells (RGCs) with relatively high efficiency (up to 28%). Neuronal GFP expression was observed within 18 h and continued for up to 14 days. This compares with values up to 60% of RGCs expressing GFP following infection with an HSV-1 vector. Adult rat dorsal root ganglion (DRG) neurons were also successfully transfected. Thus, in summary, Nucleofection provides the possibility for a fast and efficient method for cDNA delivery and study of gene function in adult mammalian neurons. PMID- 15652628 TI - Basophilia, acidophilia and argyrophilia of "dark" (compacted) neurons during their formation, recovery or death in an otherwise undamaged environment. AB - By means of a condenser discharge electric shock paradigm, "dark" (compacted) granule neurons were momentarily produced in the hippocampal dentate gyri of rats, which were sacrificed either immediately or following survival periods ranging from 1h to 30 days. Except for the morphological changes related to the formation, recovery or death of the "dark" neurons, the affected brain areas remained undamaged. Vibratome, frozen, cryostat and paraffin sections were stained with Mayer's hematoxylin, acid fuchsin, or Fluoro-Jade and by three silver methods widely used for the demonstration of damaged neurons; with or without previous removal of nucleic acids, partial digestion of proteins or blockade of the negatively charged side-groups of nucleic acids and proteins. The results allowed the following conclusions: (i) "Dark" neurons acquire argyrophilia and excess basophilia simultaneously with their momentary formation. (ii) Negatively charged protein molecules are responsible for these processes. (iii) From the recovering "dark" neurons, the acquired basophilia and argyrophilia disappear within a few hours post-insult. (iv) From the moribund or dead "dark" neurons, the acquired basophilia disappears in the same period of time while the acquired argyrophilia in a few days. (vi) Freshly-produced or recovering "dark" neurons are slightly acidophilic, whereas the moribund or dead ones display intense acidophilia. PMID- 15652631 TI - Explicit solutions for a class of indirect pharmacodynamic response models. AB - Explicit solutions for four, ordinary differential equation (ODE)-based, types of indirect response models are presented. These response models were introduced by Dayneka et al. in 1993 [J. Pharmacokinet. Biopharm. 21 (1993) 457] to describe pharmacodynamic responses utilizing inhibitory or stimulatory E(max) type functions. The explicit solutions are expressed in terms of hypergeometric 2F1 functions and their analytical continuations. A practical application is demonstrated for modeling the kinetics of drug action for ibandronate, a potent bisphosphonate that suppresses bone turnover resulting in a reduction in the markers of bone turnover. Ten times shorter model evaluation times, with the explicit solution compared with the differential equation implementation, may enhance situations where a large number of model evaluations are needed, such as clinical trial simulations and parameter estimation. PMID- 15652629 TI - HPLC with electrochemical and fluorescence detection procedures for the diagnosis of inborn errors of biogenic amines and pterins. AB - The analysis of biogenic amines (BA) and pterins in cerebrospinal fluid (CSF) is essential for the early diagnosis of neurotransmission defects in the paediatric age. Our aim was to standardize previously reported HPLC procedures for the analysis of BA and pterins in CSF and to establish reference values for a paediatric population. Samples from 127 subjects (age range 11 days to 16 years; average 3.8) were analyzed by HPLC with electrochemical and fluorescence detection. Both BA (homovanilic and 5-hydroxyindoleacetic acid) and pterins (neopterin and biopterin) concentrations in CSF showed a negative correlation with age. This finding led us to stratify reference values into six groups according to age. In conclusion, analysis of BA and pterins in CSF by HPLC procedures is a useful set of tools for the diagnosis of inborn errors of metabolism of these compounds. The establishment of reference intervals may be difficult, since there is a strong correlation between BA concentrations and the age of controls and, as a result, a large number of CSF samples from control populations would be necessary for this purpose. PMID- 15652630 TI - Enhancing protein expression in single HEK 293 cells. AB - Recombinant proteins are routinely expressed in heterologous expression systems such as human embryonic kidney 293 (HEK 293) cells. The efficiency of the expression is critical when the expressed protein must be characterized at the single-cell level. Here we describe a simple method by which the protein expression efficiency in single HEK 293 cells is enhanced by coexpressing simian virus 40 large T antigen (TAg), a powerful oncoprotein. Using the GluR2 ionotropic glutamate receptor as an example, we found that the receptor expression in single HEK 293S cells increased approximately seven-fold. The ratio of the plasmid amount of TAg to that of the receptor was optimized at 1:10, while the receptor function was unaffected in the presence of TAg. We further used fluorescence imaging from a population of cells as an independent detection method and found a similar increase in expression of green fluorescent protein (GFP) by TAg coexpression. This method is thus applicable for enhancing the expression of both membrane and soluble proteins at the single-cell level. More importantly, the function of a protein can be studied directly in intact cells, a feature particularly useful for studying membrane proteins. PMID- 15652632 TI - A Java application for tissue section image analysis. AB - The medical industry has taken advantage of Java and Java technologies over the past few years, in large part due to the language's platform-independence and object-oriented structure. As such, Java provides powerful and effective tools for developing tissue section analysis software. The background and execution of this development are discussed in this publication. Object-oriented structure allows for the creation of "Slide", "Unit", and "Cell" objects to simulate the corresponding real-world objects. Different functions may then be created to perform various tasks on these objects, thus facilitating the development of the software package as a whole. At the current time, substantial parts of the initially planned functionality have been implemented. Getafics 1.0 is fully operational and currently supports a variety of research projects; however, there are certain features of the software that currently introduce unnecessary complexity and inefficiency. In the future, we hope to include features that obviate these problems. PMID- 15652634 TI - Digital image analysis of haematopoietic clusters. AB - Counting and differentiating cell clusters is a tedious task when performed with a light microscope. Moreover, biased counts and interpretation are difficult to avoid because of the difficulties to evaluate the limits between different types of clusters. Presented here, is a computer-based application able to solve these problems. The image analysis system is entirely automatic, from the stage screening, to the statistical analysis of the results of each experimental plate. Good correlations are found with measurements made by a specialised technician. PMID- 15652633 TI - Introducing DoT-U2--an XML-based knowledge supported checklist software for documentation of a newborn clinical screening examination. AB - In a project concerning the German newborn screening examination "U2" we developed a software system called DoT-U2 for concurrent documentation at the point of care. Physicians can enter findings in(to) a tree structured protocol with management of logical dependencies. Additionally, all findings except free text annotations can be entered by speech recognition. The software system program is written in Java and uses separate XML-based modules both for knowledge and language representation. It can, therefore, easily be adapted to other languages and further documentation scenarios. We showed the high flexibility of the software system by integrating it in a completely new setting in Salt Lake City without major problems. We found that modular software development with platform independent Java and XML leads to highly flexible software which can be adapted to very different scenarios without knowing their requirements ahead of time. PMID- 15652635 TI - Robust tests for the multivariate Behrens-Fisher problem. AB - Hotelling's T2 procedure is used to test the equality of means in two-group multivariate designs when covariances are homogeneous. A number of alternatives to T2, which are robust to covariance heterogeneity, have been proposed in the literature. However, all are sensitive to departures from multivariate normality. We demonstrate how to obtain multivariate tests that are robust to covariance heterogeneity and non-normality with estimators of location and scale based on trimming and Winsorizing. The performance of six alternatives to T2 was examined via Monte Carlo methods when characteristics of the research design, degree of covariance heterogeneity, and degree of non-normality were manipulated. We have recently developed a program written in the SAS/IML language that can be used to implement these robust multivariate tests. Recommendations are provided on the specific data-analytic conditions under which these tests should be adopted. PMID- 15652636 TI - Bifurcation software in Matlab with applications in neuronal modeling. AB - Many biological phenomena, notably in neuroscience, can be modeled by dynamical systems. We describe a recent improvement of a Matlab software package for dynamical systems with applications to modeling single neurons and all-to-all connected networks of neurons. The new software features consist of an object oriented approach to bifurcation computations and the partial inclusion of C-code to speed up the computation. As an application, we study the origin of the spiking behaviour of neurons when the equilibrium state is destabilized by an incoming current. We show that Class II behaviour, i.e. firing with a finite frequency, is possible even if the destabilization occurs through a saddle-node bifurcation. Furthermore, we show that synchronization of an all-to-all connected network of such neurons with only excitatory connections is also possible in this case. PMID- 15652637 TI - Anaesthesia record system on handheld computers--pilot experience and uses for quality control and clinical guidelines. AB - This paper describes a mobile information system to collect patient information for anesthesia quality control. In this system, a mobile database program was designed for use on handheld computers (Pocket PC). This program is used to collect patient data at the bedside on the handhelds, with a daily synchronization of the data between the anaesthesiologists' handhelds with the anaesthesia database. All collected data are later used for quality control analysis. Furthermore, clinical guidelines will be included on these same handhelds. During the pilot phase, data from a sample set of about 300 patients were incorporated. The processes and interfaces of the system are presented in the paper. The current mobile database system has been designed to replace the original paper-based data collection system. The individual anaesthesiologist's handheld synchronizes patient data daily with anaesthesia database center. This information database is analyzed and used not only to give feedback to the individual doctor or center, but also to review the use of the guidelines provided and the results of their utilization. PMID- 15652638 TI - Power calculation for the likelihood ratio-test when comparing two dependent intraclass correlation coefficients. AB - Comparing the reproducibility level of two devices with continuous outcome on a unique sample of subjects (each subject being assessed several times with both devices) comes down to compare two dependent intraclass correlation coefficients (ICCs). When planning such a reproducibility study, one has to specify both the number of subjects to be included and the number of replicates per subject associated to each device. We propose SAS and S-plus macros, which allow power calculations by implementing a simulation study where dependent ICCs are compared by means of a likelihood ratio-test. PMID- 15652639 TI - A computer program to estimate the parameters of covariate dependent higher order Markov model. AB - This paper presents a computer program developed in S-plus to estimate the parameters of covariate dependent higher order Markov Chain and related tests. The program can be applied for two states Markov Chain with any order and any number of covariates depending on the PC capabilities. The program provides the maximum likelihood estimates of the parameters, together with their estimated standard error, t-value and significance level. It also produces the test results for likelihood ratio and model chi-square. To illustrate the program we have used a longitudinal data set on maternal morbidity of rural women in Bangladesh. The occurrences of haemorrhage, convulsion, or fits at different follow-ups were used as outcome variable. Economic status, wanted pregnancy, ages at marriage, and education of women were used as covariates. PMID- 15652640 TI - Aplysia seductin is a water-borne protein pheromone that acts in concert with attractin to stimulate mate attraction. AB - Mate attraction in Aplysia involves the long-distance water-borne protein pheromones attractin, enticin, and temptin which are released during egg-laying. Other water-borne pheromones are predicted to act in concert with attractin, enticin, and temptin, but their identities were unknown. We recently identified a highly expressed Aplysia californica albumen gland gene (Alb-23) that encoded a novel protein by differential library screening of an albumen gland cDNA library. To determine whether Alb-23 ('seductin') was a water-borne pheromone, we employed Western blot analysis, purification and expression of albumen gland proteins, immunolocalization studies, pheromone secretion assays, comparative genomics, and behavioral bioassays. Immunoreactive seductin was detected in eluates of egg cordons, indicating that seductin was secreted onto the cordon during egg laying. Aplysia brasiliana seductin was 94% identical to its A.californica homolog. In T maze attraction assays, the combination of attractin and seductin was significantly attractive to potential mates, whereas either protein alone was not. Data from this and previous studies support the hypothesis that seductin is a water-borne protein pheromone that acts in concert with attractin, enticin, and temptin to attract Aplysia to form and maintain mating aggregations. PMID- 15652641 TI - Direct cDNA cloning of novel conopeptide precursors of the O-superfamily. AB - Conotoxins from the venom of marine cone snails (genus Conus) represent large families of proteins exhibiting a similar precursor organization, but highly diverse pharmacological activities. A directed PCR-based approach using primers according to the conserved signal sequence was applied to investigate the diversity of conotoxins from the O-superfamily. Using 3' RACE, cDNA sequences encoding precursor peptides were identified in five Conus species (Conus capitaneus, Conus imperialis, Conusstriatus, Conus vexillum and Conus virgo). In all cases, the sequence of the signal region exhibited high conservancy, whereas the sequence of the mature peptides was either almost identical or highly divergent among the five species. These findings demonstrate that beside a common genetic pattern divergent evolution of toxins occurred in a highly mutating peptide family. PMID- 15652642 TI - Deletion of two C-terminal Gln residues of 12-26-residue fragment of melittin improves its antimicrobial activity. AB - In our previous paper it was shown that the two C-terminal Gln residues of a C terminal 15-residue fragment, Mel(12-26) (GLPALISWIKRKRQQ-NH2), of melittin and a series of individual substituted analogues might not involved in the interaction with bacterial membranes. In this paper, peptides with one and two Gln residues deletion, respectively, Mel(12-25) and Mel(12-24), were synthesized and characterized. Both of the deletion peptides showed higher antimicrobial activities than the parent peptide, Mel(12-26). If both of the Gln residues of Mel(12-26) were respectively replaced by a hydrophilic amino acid Gly, the antimicrobial activity increased slightly. If the Gln residue of Mel(12-25) was replaced by a hydrophobic amino acid Leu, the antimicrobial activity changed little, although the substituted peptide possessed much higher hydrophobicity and higher alpha-helical conformation percentage in 1,1,1,3,3,3-hexafluoro-2 propanol/water determined by circular dichroism spectroscopy (CD) than the parent peptide. These results indicated that the two C-terminal residues might be indeed not involved in the binding to bacterial membranes. The antimicrobial activity increasing with the residue deletion may be caused by the decrease of the translational and rotational entropic cost of the binding of the peptides to bacterial membranes because of the lower molecular weights of the deletion peptides. PMID- 15652643 TI - Molecular cloning of mRNA from toad granular gland secretion and lyophilized skin: identification of Bo8--a novel prokineticin from Bombina orientalis. AB - Prokineticins are small (approximately 8 kDa), biologically active secretory proteins whose primary structures have been highly conserved throughout the Animal Kingdom. Representatives have been identified in the defensive skin secretions of several amphibians reflecting the immense structural/functional diversity of polypeptides in such. Here we describe the identification of a prokineticin homolog (designated Bo8) from the skin secretion of the Oriental fire-bellied toad (Bombina orientalis). Full primary structural characterization was achieved using a combination of direct Edman microsequencing, mass spectrometry and cloning of encoding skin cDNA. The latter approach employed a recently described technique that we developed for the cloning of secretory peptide cDNAs from lyophilized skin secretion, and this was further extended to employ lyophilized skin as the starting material for cDNA library construction. The Bo8 precursor was found to consist of an open-reading frame of 96 amino acid residues consisting of a putative 19-residue signal peptide followed by a single 77-residue prokineticin (Mr=7990 Da). Amino acid substitutions in skin prokineticins from the skin secretions of bombinid toads are confined to discrete sites affording the necessary information for structure/activity studies and analog design. PMID- 15652644 TI - Cholecystokinin mRNA in Atlantic herring, Clupea harengus--molecular cloning, characterization, and distribution in the digestive tract during the early life stages. AB - The mRNA of the peptide hormone cholecystokinin (CCK) was isolated from juvenile Atlantic herring, Clupea harengus, by RT-PCR. The open reading frame encodes a 137 amino acid-long precursor protein. The peptide sequence of herring CCK-8, DYMGWMDF, is identical to that of higher vertebrates and elasmobranchs, and contains methionine in the sixth position from the C-terminus, which has not been reported previously in teleosts. Expression analysis by in situ hybridization shows that positive endocrine-like cells were mainly located in the pyloric caeca and to a less extent in the rectum of the juvenile. A few positive cells were also found in the pyloric portion of the stomach and the intestine. CCK cells were present in all the larvae examined from the day of hatching onwards. Although the CCK cells were scattered throughout the whole midgut, no signals were detected in either the foregut or the hindgut. Since herring larvae have a straight gut, the distribution pattern of CCK cells seems to be reflected in the anatomy of the gut. PMID- 15652645 TI - Identification of immunodominant regions of Brassica juncea glyoxalase I as potential antitumor immunomodulation targets. AB - Glyoxalase I activity has been shown to be directly related to cancer and its inhibitors have been used as anti-cancer drugs. Immunochemical studies have shown immunochemical relatedness among animal and plant glyoxalase I, but its potential application for biomedical research has not been investigated. In order to understand the conserved immunochemical regions of the protein and to determine probable immunomodulation targets, a cellulose-bound scanning peptide library for Brassica juncea glyoxalase I was made using the spot synthesis method. Immuno probing of the library, using B. juncea anti-glyoxalase I monospecific polyclonal antibodies, revealed three immunodominant regions, epitope I, II, and III. In the homology model of B. juncea glyoxalase I generated by threading its sequence onto the human glyoxalase I, the high accessible surface area and the hydrophilic nature of the epitopes confirmed their surface localization and hence their accessibility for antigen-antibody interaction. Epitopes I and II were specific to B. juncea glyoxalase I. Localizing the epitopes on available glyoxalase I sequences showed that epitope III containing the active site region was conserved across phyla. Therefore, this could be used as a potential immunomodulation target for cancer therapy. Moreover, as the most immunogenic epitopes were mapped on the surface of the protein, this method could be used to discover potential therapeutic targets. It is a simple and fast approach for such investigations. This study, to our knowledge, is the first in epitope mapping of glyoxalase I and has great biomedical potential. PMID- 15652646 TI - The inhibitory effects of synthetic short peptides, mimicking MICA and targeting at NKG2D receptors, on function of NK cells. AB - NKG2D is an activating receptor expressed on most of human NK cells, one of whose ligands is MICA. Based on the crystal structure of NKG2D-MICA complex, we synthesized three short peptides (P1, P2 and P3), mimicking functional alpha1 and alpha2 domain of MICA. The inhibitory effects of three peptides on NK-92 cells, a human NK cell line against Hela cells were observed and the inhibitory percentage was 38% at maximum for P1+P2+P3 in concentration of 1nM. The same peptides had no effect on NK-92 cell against target cells lacking MICA (K562 cells line). The unrelated peptides as controls had no effect on the system. Two peptides (P2 and P3) were prolonged at one or both ends, and the longer forms of peptides exerted stronger inhibitory effects than their shorter forms. Each combination of two peptides exerted a stronger function than single peptide (P1, P2, P3), indicating that shedding of longer amino acid sequence of alpha1 domain or more domain sites of MICA are better than shorter sequence and fewer sites. P1+P2+P3 revealed the almost same inhibitory rate as the soluble MICA (sMICA). P1+P2+P3 were also able to alleviate the concanavalin A-induced murine autoimmune hepatitis in vivo, conforming the similarity of NKG2D between human and mice. The results demonstrate that MICA-mimicking peptides will be useful to search the specific functional sites for NKG2D-MICA interaction, but also promising in explaining NKG2D-related autoimmunity. PMID- 15652647 TI - Radioprotection by N-palmitoylated nonapeptide of human interleukin-1beta. AB - Interleukin-1beta (IL-1beta) is a cytokine involved in homeostatic processes of the immune system and specifically in inflammatory reactions. The nonapeptide of human IL-1beta (VQGEESNDK, position 163-171) has been shown to retain adjuvant and immunostimulatory activities of the native molecule without any inflammatory and pyrogenic properties. A lipophilic derivative of IL-1beta nonapeptide having a palmitoyl residue at the amino terminus was synthesized in order to determine the effects of such structural modification on its bioactivities. The structurally modified peptide derivative, palmitoylated peptide, significantly protected C3H/HeN mice against potentially lethal doses of ionizing radiation. The dose reduction factor was found to be 1.07. Hematological studies show improved recovery of red blood cells and platelets in irradiated and palmitoylated peptide treated mice as compared with the untreated and irradiated group. These results suggest the importance of the derivatization of small peptides of radioprotective, but toxic cytokines in order to enhance radioprotective activity while reducing unwanted toxic side effects. PMID- 15652649 TI - Pharmacodynamics and pharmacokinetics of recombinant hirudin via four non parenteral routes. AB - One of recombinant hirudin variants, rHV2, a polypeptide used as an anticoagulant agent in clinic, was administered to anesthetized rats via intratracheal, buccal, nasal and rectal routes. Prolongation in clotting time and thrombin time was measured to calculate pharmacological bioavailability. Plasma concentration of rHV2 was determined using a chromogenic thrombin substrate assay and pharmacokinetic parameters were obtained on the basis of a non-compartmental model. Intravenous administration was also performed as the gold standard by which the other routes were compared. Difference in pharmacological bioavailability (P.A.), bioavailability (F) and absorption rate of rHV2 was found for the four non-parenteral routes. The rank order for both P.A. and F was intratracheal>nasal>buccal>rectal. Absorption was more rapid after both intratracheal and rectal administration (tmax approximately 20-40 min), compared with that after nasal and rectal administration. It is evident that the pulmonary route is preferable to other three routes for successful systemic delivery of rHV2. PMID- 15652648 TI - Importance of the central region of lamprey gonadotropin-releasing hormone III in the inhibition of breast cancer cell growth. AB - Naturally occurring isoforms of the decapeptide gonadotropin-releasing hormone (GnRH) share residues 1-4 and 9-10. lGnRH-III, the third isoform isolated in the sea lamprey has no endocrine effect in mammals but shows a direct antiproliferative effect on human breast, prostate and endometrial cancer cell lines. To investigate these features, residues 5-8 of lGnRH-III were systematically replaced with Ala. The ability of the synthetic analogs to interact with receptors on MDA-MB 231 human breast cancer cells and their effect on the growth of the same cell line were investigated. [Ala6]lGnRH-III and [Ala7]lGnRH-III have neither receptor binding nor antiproliferative activity. Replacement of His5 with Ala resulted in an analog that binds to the receptor but does not have antiproliferative activity. The results are in agreement with previous reports that modifications of Lys at position 8 are well tolerated. PMID- 15652652 TI - The effect of lipopolysaccharide on cholecystokinin in murine plasma and tissue. AB - Several mechanisms have been proposed for neuroimmune communication supporting sickness behavior (fever, anorexia, inactivity, and cachexia) following infection. We examined the role of cholecystokinin as a neurochemical intermediary of sickness behavior by determining plasma, duodenum, hypothalamus, and brainstem cholecystokinin concentrations 30 and 60 min and 12 h following intraperitoneal lipopolysaccharide (LPS) (0.25 and 2.5 mg/kg). Hypothalamic cholecystokinin was significantly lower in LPS- versus saline-treated mice 30 min (0.25 and 2.5 mg/kg) and 12 h (2.5 mg/kg) post-injection. Plasma cholecystokinin of LPS-treated mice was significantly lower than that of controls 1 and 12 h post injection, a finding consistent with a non-endocrine action of peripheral cholecystokinin. PMID- 15652651 TI - Effect of CCK-8 on insulin-induced hyperphagia and hypothalamic orexigenic neuropeptide expression in the rat. AB - The influence of cholecystokinin octapeptide (CCK-8) on normal and insulin induced feeding and expression of orexigenic hypothalamic neuropeptides was investigated in male rats. CCK-8, administered during meals (4 microg/kg) or continuously (32 microg/kg over 60 min), blunted the stimulating effect of insulin (50 IU/kg) on feeding by reducing meal size (-60%; P<0.05 or -86%; P<0.0001, respectively). Rats without access to food and injected with IP insulin (50 IU/kg) showed increased hypothalamic mRNA levels of orexin (+30%; P<0.05) and melanin-concentrating hormone (+52%; P<0.05), as compared with ad libitum-fed and saline-injected control rats. Continuous IP infusion of CCK-8 (32 microg/kg) blunted these increases. Our results suggest that both orexin and melanin concentrating hormone participate in the response to insulin hypoglycemia without food being present; these neurons may be involved in mechanisms related to insulin-induced hyperphagia. Signals triggered by peripheral CCK-8 act to decrease the expression of orexin and melanin-concentrating hormone. This may be associated with a reduction in hyperphagia. PMID- 15652650 TI - Antinociceptive action of hemopressin in experimental hyperalgesia. AB - Endogenous hemorphins, derived from degradation of the beta-chain of hemoglobin, lower arterial blood pressure and exert an antinociceptive action in experimental models of nociception. Hemopressin, derived from the alpha-chain of hemoglobin, also decreases blood pressure, but its effects on pain have not been studied. In this work, we examined the influence of hemopressin on inflammatory pain. Hemopressin reverted the hyperalgesia induced by either carrageenin or bradykinin when injected concomitantly or 2.5 h after the phlogistic agents. Hemopressin administered systemically also reverted the hyperalgesia induced by carrageenin. Naloxone did not prevent the antinociceptive action of this peptide. These data suggest that hemopressin inhibits peripheral hyperalgesic responses by mechanisms independent of opioid receptor activation. PMID- 15652653 TI - Peptide ligand binding properties of the corticotropin-releasing factor (CRF) type 2 receptor: pharmacology of endogenously expressed receptors, G-protein coupling sensitivity and determinants of CRF2 receptor selectivity. AB - The CRF2 receptor is involved in stress responses, cardiovascular function and gastric motility. Endogenous agonists (urocortin (UCN) 2, UCN 3) and synthetic antagonists (astressin2-B, antisauvagine-30) are selective for CRF2 over the CRF1 receptor. Peptide ligand binding properties of the CRF2 receptor require further investigation, including ligand affinity for endogenously expressed receptors, the effect of receptor-G-protein coupling on ligand affinity, and the molecular basis of ligand selectivity. Ligand affinity for rat CRF(2a) in olfactory bulb and CRF(2b) in A7r5 cells was similar to that for the cloned human CRF(2a) receptor (within three-fold), except for oCRF (9.4- and 5.4-fold higher affinity in olfactory bulb and A7r5 cells, respectively). Receptor-G-protein uncoupling reduced agonist affinity only 1.2- to 6.5-fold (compared with 92-1300-fold for the CRF1 receptor). Ligand selectivity mechanisms were investigated using chimeric CRF2/CRF1 receptors. The juxtamembrane receptor domain determined selectivity of antisauvagine-30, the N-terminal-extracellular domain contributed to selectivity of UCN 3, and both domains contributed to selectivity of UCN 2 and astressin2-B. Therefore ligands differ in the contribution of receptor domains to their selectivity, and CRF2-selective antagonists bind the juxtamembrane domain. These findings will be important for identifying the CRF2 receptor in tissues and for developing ligands targeting the receptor, both of which will be useful in identifying the emerging physiological functions of the CRF2 receptor. PMID- 15652654 TI - Effects of orexins/hypocretins on neuronal activity in the paraventricular nucleus of the thalamus in rats in vitro. AB - Orexin-A (ORX-A) and orexin-B (ORX-B), also called hypocretin-1 and hypocretin-2, respectively, act upon orexin 1 (OX1R) and orexin 2 (OX2R) receptors, and are involved in the regulation of sleep-wakefulness and energy homeostasis. Orexin neurons in the lateral hypothalamic perifornical region project heavily to the paraventricular nucleus of the thalamus (PVT), which is deeply involved in the control of motivated behaviors. In the present study, electrophysiological and cytosolic Ca2+ concentration ([Ca2+]i) imaging studies on the effects of ORX-A and ORX-B on neurons in the PVT were carried out in rat brain slice preparations. ORX-A and/or ORX-B were applied extracellularly in the perfusate. Extracellular recordings showed that about 80% of the PVT neurons were excited dose-dependently by both ORX-A and ORX-B at concentrations of 10(-8) to 10(-6)M, and the increase in firing rate was about three times larger for ORX-B than for ORX-A at 10(-7)M. When both ORX-A and ORX-B were applied simultaneously at 10(-7)M, the increase in firing rate was almost equal to that of ORX-B at 10(-7)M, suggesting that the PVT neurons do not show a high affinity to ORX-A which is expected if they have OX1R receptors. The excitatory effect of ORX-B was seen in low Ca2+ and high Mg2+ ACSF as well as in normal ACSF, and the increase in firing rate was greater in low Ca2+ and high Mg2+ ACSF than in normal ACSF. [Ca2+]i imaging studies demonstrated that [Ca2+]i was increased in about 50% of the PVT neurons by both 10(-7)M ORX-A and ORX-B with a stronger effect for ORX-B, and the increase in [Ca2+]i induced by ORX-B was abolished in Ca2+-free ACSF, suggesting that ORX-B does not release Ca2+ from intracellular Ca2+ stores. Subsequent whole cell patch clamp recordings revealed that an after hyperpolarization seen following each action potential in normal ACSF disappeared in Ca2+-free ACSF, and the mean magnitude of the depolarization induced by ORX-B was same in normal, Ca2+-free and TTX-containing Ca2+-free ACSFs. Furthermore, ORX-B-induced depolarization was reversed to hyperpolarization when membrane potential was lowered to about -97 mV, and an increase of extracellular K+ concentration from 4.25 to 13.25 mM abolished the ORX-B-induced depolarization, indicating that the ORX-B-induced depolarization is associated with an increase in the membrane resistance resulting from a closure of K+ channels. These results suggest that orexins depolarize and excite post synaptically PVT neurons via OX2R receptors, and that orexin-activated PVT neurons play a role in the integration of sleep-wakefulness and energy homeostasis, and in the control of motivated behaviors. PMID- 15652655 TI - Oxytocin ameliorates oxidative colonic inflammation by a neutrophil-dependent mechanism. AB - Oxytocin (OT), a nonapeptide produced in the paraventricular and the supraoptical nuclei in the hypothalamus has a wide range of effects in the body. However, the role of OT on the gastrointestinal (GI) tract has to be settled. OT may participate in the regulation of motility, secretion, blood flow, cell turnover and release of neurotransmitters and/or peptides in the GI tract, possesses antisecretory and antiulcer effects, facilitates wound healing and is involved in the modulation of immune and inflammatory processes. The present work was conducted to assess the possible therapeutic effects of OT against the acetic acid-induced colonic injury in the rat. METHODS: Colitis was induced by intracolonic administration of acetic acid (5%) in Sprague-Dawley rats (200-250 g). Either saline or OT (0.5 mg/kg) was injected subcutaneously, immediately after the induction of colitis and repeated two times a day for 4 days. On the 4th day, rats were decapitated and distal 8 cm of the colon were removed for the macroscopic and microscopic damage scoring, determination of tissue wet weight index (WI), malondialdehyde (MDA) levels, an end product of lipid peroxidation; glutathione (GSH) levels, a key antioxidant; and myeloperoxidase (MPO) activity, as an indirect index of neutrophil infiltration. Colonic collagen content, as a fibrosis marker was also determined. Lactate dehydrogenase (LDH) and tumor necrosis factor-alpha (TNF-alpha) levels were assayed in serum samples. In the acetic acid-induced colitis, macroscopic and microscopic damage scores, WI, MDA and MPO levels were significantly increased, while GSH levels were decreased when compared to control group (p <0.05-<0.001). Treatment with OT abolished the colitis-induced elevations in damage scores, WI, MDA and MPO levels and restored the GSH levels (p <0.05-0.001). Similarly, acetic acid increased the collagen content of colonic tissues and OT-treatment reduced this value to the level of the control group. Serum LDH and TNF-alpha levels were also elevated in the acetic acid-induced colitis group as compared to control group, while this increase was significantly decreased by OT treatment. The results suggest that OT, which improves the antioxidative state of the colonic tissue and ameliorates oxidative colonic injury via a neutrophil-dependent mechanism, requires further investigation as a potential therapeutic agent in colonic inflammation. PMID- 15652657 TI - Protective effects of intermedin/adrenomedullin2 on ischemia/reperfusion injury in isolated rat hearts. AB - Intermedin (IMD) is a novel member of the calcitonin/calcitonin gene-related peptide (CT/CGRP) family identified from human and other vertebrate tissues. Preprointermedin can generate a 47-amino acid mature peptide (IMD(1-47)) and a shorter 40-amino acid one (IMD(8-47)) by proteolytic cleavage. The present study was designed to determine the protective effect of IMD on cardiac ischemia/reperfusion (I/R) injury and its possible mechanism. Isolated rat hearts were perfused on a Langendorff apparatus and subjected to 45-min global ischemia and 30-min reperfusion. Cardiac function was measured. The release of myocardial protein and lactate dehydrogenase (LDH) and the formation of malondialdehyde (MDA) were assayed. Myocardial cAMP content was determined by radioimmunoassay (RIA). Cardiac I/R induced a marked inhibition of cardiac function and myocardial injury. Reperfusion with IMD significantly attenuated the I/R injury. Compared with I/R alone, perfusion with 10(-8)mol/L IMD(1-47) and IMD(8-47) induced a 36% and 33% increase in Delta left ventricular pressure (DeltaLVP), 30% and 28% in maximal rate of increase of LV pressure (+LVdP/dt max), and 34% and 31% in maximal rate of decrease of LV pressure (-LVdP/dt max), respectively (all P<0.01) but an approximately 58% and 51% decrease in LV diastolic pressure, respectively (P<0.01). In addition, perfusion with IMD markedly attenuated the leakage of LDH, total protein and myoglobin from myocardia compared with I/R alone. The contents of ventricular myocardia cAMP after reperfusion with 10(-8)mol/L IMD(1-47) and IMD(8-47) were 130% and 91% higher, respectively, than that with I/R alone (all P<0.01). However, formations of myocardial MDA were 52% and 50% lower than that with I/R alone (all P<0.01), respectively. Interestingly, the above IMD effects were similar to those of adrenomedullin (10(-8)mol/L). These results suggest that IMD, like adrenomedullin, exerts cardio-protective effects against myocardial I/R injury. PMID- 15652656 TI - Octreotide ameliorates sepsis-induced pelvic inflammation in female rats by a neutrophil-dependent mechanism. AB - Sepsis is a generalized inflammatory response, which involves organ systems remote from the locus of the initial infectious insult, accompanied by the release of cytokines and the subsequent formation of reactive oxygen and nitrogen species. The aim of this study was to investigate the possible protective effect of octreotide (OCT), a synthetic somatostatin analogue, against sepsis-induced oxidative damage in the uterine and ovarian tissues of rats. Sepsis was induced by caecal ligation and puncture method in female Wistar albino rats. Sepsis and sham operated (control) groups received either saline or OCT (50 microg/kg, i.p.; Novartis) immediately after the operation and at 12 h. Twenty-four hours after the surgery, rats were decapitated and serum TNF-alpha levels and tissue malondialdehyde (MDA) content, glutathione (GSH) levels and myeloperoxidase (MPO) activity were determined in the uterus and ovaries. Oxidant-induced tissue fibrosis was determined by tissue collagen contents, while the extent of tissue injuries was analyzed microscopically. Sepsis increased serum TNF-alpha levels and resulted in decreased GSH levels and increased MDA levels, MPO activity and collagen contents in both the uterus and the ovaries (p<0.05-0.001) indicating the presence of the oxidative damage, as also confirmed by histological analysis. On the other hand, OCT administration reversed these oxidant responses and reduced the severity of microscopic damage (p<0.001). In conclusion, OCT protects against sepsis-induced oxidative injury of the uterine and ovarian tissues by diminishing neutrophil infiltration, an important source of oxygen free radicals. Our results suggest that OCT may be of therapeutic value in ameliorating sepsis associated pelvic inflammation. PMID- 15652658 TI - All D-VIP mitigates vasodilation elicited by L-VIP, micellar L-VIP and micellar PACAP1-38, but not PACAP1-38, in vivo. AB - The purpose of this study was to determine whether all D-vasoactive intestinal peptide (VIP), an inactive optical isomer of L-VIP, modulates the vasorelaxant effects of human L-VIP and pituitary adenylate cyclase activating peptide (PACAP)1-38, two ubiquitous and pleiotropic neuropeptides that activate VPAC1 and VPAC2, two VIP subtype receptors, in the intact peripheral microcirculation. Using intravital microscopy, we found that suffusion of all D-VIP had no significant effects on arteriolar diameter in the intact hamster cheek pouch. However, all D-VIP significantly attenuated L-VIP-induced vasodilation in a concentration-dependent fashion (P<0.05). likewise, all D-VIP significantly attenuated the vasorelaxant effects of L-VIP associated with sterically stabilized phospholipid micelles (SSM; P<0.05). Although all D-VIP had no significant effects on L-PACAP1-38-induced vasodilation, it abrogated PACAP1-38 in SSM-induced responses (P<0.05). The effects of all D-VIP were specific because it had no significant effects on acetylcholine-, nitroglycerin- and bradykinin induced vasodilation. Taken together, these data indicate that all D-VIP attenuates the vasorelaxant effects of random coil and alpha-helix L-VIP as well as those of alpha-helix but not random coil PACAP in the intact peripheral microcirculation in a specific fashion. These effects are mediated, most likely, through interactions with VPAC1/VPAC2 receptors. We suggest that all D-VIP could be exploited as a novel, safe and active targeting moiety of VPAC1/VPAC2 receptors in vivo. PMID- 15652659 TI - Development of a selective peptide antagonist for the human natriuretic peptide receptor-B. AB - Activation by C-type natriuretic peptide (CNP) of its receptor NPRB results in venodilation and inhibition of cellular proliferation. NPRB-selective antagonists should be useful to understand their physiological implications. We previously observed that [Thr9,Ser11,Arg16](N,C-ANP)pBNP (P12) is an antagonist for bNPRB and a potent agonist for bNPRA. The antagonist [Ser11](N-CNP,C-ANP)pBNP(2-26) (P18) displays six-fold selectivity towards hNPRB versus hNPRA. Deletion of the C terminus in [Ser11](N-CNP,C-ANP)pBNP(2-25) (P19) decreases its affinity for hNPRA but improves its selectivity 35-fold. Peptide libraries based on P19 using phage display methodology yielded two positive clones P20 and P21. P19 behaves as the most potent antagonist, but P20 is the most selective. PMID- 15652661 TI - Effect of corticosterone on CART peptide levels in rat blood. AB - We have recently demonstrated that CART peptides display a diurnal rhythm in blood that depends partly on glucocorticoids levels. This study extends previous findings by directly testing the effects of acute administration of corticosterone and metyrapone on CART peptide levels in blood. Acute treatment with corticosterone augmented CART levels, while metyrapone administration prevented the increase in CART in the evening hours. These results further support the hypothesis that glucocorticoids play a role in the regulation of CART levels in blood. PMID- 15652660 TI - The effects of CGRP on calcium transients of dedifferentiating cultured adult rat cardiomyocytes compared to non-cultured adult cardiomyocytes: possible protective and deleterious results in cardiac function. AB - CGRP has potent cardiovascular effects but its role in heart failure is unclear. Effects of CGRP on calcium concentrations in fresh adult rat cardiomyocytes, cultured adult cardiomyocytes and neonatal cardiomyocytes were determined by real time fluorescence spectrophotometry. Treatment of cultured adult cardiomyocytes with CGRP resulted in a rapid cessation of beating and a reduction in intracellular calcium. Similar results were obtained in cultured neonatal myocytes. However, rod-shaped adult cardiomyocytes revealed a number of responses; (a) non-beating cells began to beat with increased intracellular calcium; (b) spontaneously beating cells exhibited increased intracellular calcium content and a faster beating rate or (c), myocytes increased their beating rate and became arrhythmic, suggesting that CGRP action on cultured dedifferentiated adult and neonatal myocytes depletes intracellular calcium, whereas in the rod-shaped mature myocytes calcium is retained, pointing to a different mode of action for CGRP on developing and dedifferentiating cardiomyocytes, compared to fully developed cardiomyocytes. PMID- 15652663 TI - Factors influencing the immune response to foreign antigen expressed in recombinant BCG vaccines. AB - A wide range of recombinant BCG vaccine candidates containing foreign viral, bacterial, parasite or immunomodulatory genetic material have been developed and evaluated, primarily in animal models, for immune response to the foreign antigen. This review considers some of the factors that may influence the immunogenicity of these vaccines. The influence of levels and timing of expression of the foreign antigen and the use of targeting sequences are considered in the first section. Genetic and functional stability of rBCG is reviewed in the second section. In the last section, the influence of dose and route of immunization, strain of BCG and the animal model used are discussed. PMID- 15652662 TI - Safety and immunogenicity of gelatin-free varicella vaccine in epidemiological and serological studies in Japan. AB - Following gelatin-containing varicella vaccine (1994-1999: 1,410,000 distributed doses), 28 serious anaphylactic reactions and 139 non-serious allergic reactions were reported, with no serious and only five non-serious reactions following gelatin-free vaccine (1999-2000: 1,300,000 distributed doses). All nine sera available from children with serious reactions tested positive for gelatin specific IgE, whereas 55 of the 70 available from those with non-serious reactions were positive, with one false positive. There was no correlation between gelatin-specific IgE antibody titers and severity of allergic reaction. Post-immunization anti-varicella antibody titers were comparable for both gelatin free and gelatin-containing vaccine groups. The new gelatin-free varicella vaccine is thought to be safe, with similar immunogenicity to the earlier gelatin containing vaccine. PMID- 15652664 TI - DNA vaccination with the Aleutian mink disease virus NS1 gene confers partial protection against disease. AB - Aleutian disease virus (ADV) causes severe losses in mink. This happens in nature as well as in farms. In spite of several attempts to provide an efficient protective protein based vaccine, experiments have failed so far. Only partial protection has been obtained. The aim of this work was to construct and test a protective DNA vaccine based on the gene encoding for the ADV non-structural protein 1 (NS1) and to test this construct as a potential vaccine candidate against ADV infection or disease. First, the vaccine construct was tested by in vitro transfection studies. NS1 protein expression was found by immunofluorescent studies and the expected size of translated protein confirmed by Western blot. Then, 18 female mink were divided into three groups: a control group, a DNA vaccinated group, and a group which received DNA vaccine plus a boost with recombinant NS1 protein in the last immunization. After virus challenge, the two DNA vaccinated groups induced higher antibody levels in the first 23 weeks of the 32 week observation period. One month after virus challenge, the most interesting finding was, that the "DNA+protein" group exhibited a significantly higher percentage of CD8+ cells, when compared to the levels in the two other groups. This, we believe, indicate a memory CTL response created by the vaccination. Most CD8+ cells were found to contain interferon gamma as measured by FACS intracellular staining. Severity of Aleutian disease was judged by quantification of plasma gammaglobulin levels and mink death statistics. The findings let us to conclude, that the two DNA vaccinated groups of mink did show milder disease characteristics, but that the vaccine effect also in this trial could only be characterized as partial. PMID- 15652665 TI - Virosome-mediated delivery of protein antigens in vivo: efficient induction of class I MHC-restricted cytotoxic T lymphocyte activity. AB - Induction of CTL responses against protein antigens is an important aim in vaccine development. In this paper we present fusion-active virosomes as a vaccine delivery system capable of efficient induction of CTL responses in vivo. Virosomes are reconstituted viral membranes, which do not contain the genetic material of the virus they are derived from. Foreign macromolecules, including protein antigens, can be encapsulated in virosomes during the reconstitution process. Functionally reconstituted virosomes retain the cell binding and fusion characteristics of the native virus. Thus, upon uptake by cells through receptor mediated endocytosis, virosomes will deliver their content to the cell cytosol. In a previous study, we demonstrated that protein antigens delivered in this manner to dendritic cells are efficiently processed for both MHC class I and class II presentation. Here, we studied in vivo induction of cellular immune responses against virosome-encapsulated ovalbumin (OVA) in mice. As little as 0.75 microg OVA delivered by fusion-active virosomes was sufficient to induce a powerful class I MHC-restricted CTL response. All immunization routes that were used (i.m., i.p. and s.c.) resulted in efficient induction of CTL activity. The CTLs induced were cytotoxic in a standard 51Cr-release assay and produced IFNgamma in response to OVA peptide. Thus, virosomes represent an ideal antigen delivery system for induction of cellular immunity against encapsulated protein antigens. PMID- 15652666 TI - DNA vaccination against respiratory syncytial virus in young calves. AB - A DNA vaccine encoding the fusion (F) gene (DNA-F) of bovine respiratory syncytial virus (BRSV) induced significant protection against BRSV infection in young calves. However, serum antibody to RSV developed more slowly in animals vaccinated with DNA-F when compared with those previously infected with BRSV. Furthermore, protection against BRSV infection was not as great as that induced by prior BRSV infection. Although there was little difference in the level of protection induced in calves vaccinated with DNA-F by either the intramuscular (i.m.) or intradermal (i.d.) routes, only the i.m. route primed for a rapid BRSV specific IgA response after BRSV challenge. These results indicate that a DNA vaccination may be effective against RSV infection even in very young infants and calves. PMID- 15652667 TI - Acute lower respiratory tract infections and respiratory syncytial virus in infants in Guinea-Bissau: a beneficial effect of BCG vaccination for girls community based case-control study. AB - Among measles unvaccinated infants in Guinea-Bissau, we tested whether case infants with acute lower respiratory tract infection (ALRI), especially ALRI caused by respiratory syncytial virus (RSV), were more likely to be Bacille Calmette Guerin (BCG)-unvaccinated and to have no scar after BCG vaccination than were control infants without symptoms of ALRI. Three hundred and eighty-six case infants with ALRI were identified at a paediatric clinic (N=84), a health centre (N=82), and in a community morbidity surveillance system (N=220). Control infants were matched on sex, age, and district and were also measles unvaccinated. In ALRI case infants, the adjusted OR of being BCG unvaccinated was 2.87 (1.31 6.32), 1.72 (0.48-6.19) in boys and 4.45 (1.48-13.4) in girls. Among BCG vaccinated ALRI case infants, the adjusted OR of having no BCG scar was 1.54 (0.86-2.75), 0.93 (0.45-1.91) in boys and 2.70 (1.21-6.02) in girls. In ALRI case infants with RSV infection, similar trends were observed. BCG vaccination may have a non-targeted protective effect against ALRI, the effect being most marked in girls. PMID- 15652668 TI - TLR9 pathway is involved in adjuvant effects of plasmid DNA-based vaccines. AB - The presence of unmethylated CpG motifs in bacterial plasmids is thought to provide necessary immunoadjuvant signals to DNA vaccination. We took advantage of CpG-unresponsive toll-like receptor 9 (TLR9) knock-out mice to study whether this pathway was required to generate immune responses to DNA vaccination. We compared two vectors, one encoding the surface glycoprotein C of pseudorabies virus shown to protect target animals against challenge, and the other encoding the cytoplasmic enzyme beta-galactosidase. In the absence of TLR9, bone marrow derived dendritic cells lost their ability to secrete IL-12 and type I IFN in response not only to CpG as expected but also to the plasmids used for vaccination. In contrast, DNA vaccination experiments showed that TLR9-deficient mice were able to mount Th1-biased antigen-specific antibody and IFN-gamma responses, albeit at lower levels than normal mice. Thus, TLR9 signaling is not needed for eliciting T- and B-cell responses to DNA encoded antigens. However, TLR9 signaling tended to enhance plasmid-adjuvant effects on antigen-specific immune responses. PMID- 15652669 TI - Defining putative T cell epitopes from PE and PPE families of proteins of Mycobacterium tuberculosis with vaccine potential. AB - The identification of T cell epitopes from immune relevant antigens of Mycobacterium tuberculosis is a critical step in the development of a vaccine covering diverse populations. Two multigene families, PE-PGRS and PPE make up about 10% of the M. tuberculosis genome. However, the functions of the proteins coded by these large numbers of genes are unknown. All possible nonameric peptide sequences from PE and PPE proteins were analysed in silico for their ability to bind to 33 alleles of class I HLA. These results reveal that of all PE and PPE proteins, a significant number of these peptides are predicted to be high affinity HLA binders, irrespective of the length of the protein. The pathogen peptides that could behave as self or partially self-peptides in the host were eliminated using a comparative study with human proteome, thus reducing the number of peptides for analysis. The structural basis for recognition of the nonamers by the respective HLA molecules thus predicted was analyzed by molecular modeling. The structural analysis showed good correlation with the binding prediction. The analysis also led to an understanding of the binding profile of the peptides with respect to different alleles of class I HLA. The predicted epitopes can be tested experimentally for their inclusion in a potential vaccine against tuberculosis that is HLA haplotype-specific. PMID- 15652670 TI - A DNA vaccine encoding genetic fusions of carcinoembryonic antigen (CEA) and granulocyte/macrophage colony-stimulating factor (GM-CSF). AB - The anti-tumor immunologic effects of plasmid DNA vaccines encoding human carcinoembryonic antigen (CEA) fused to mouse granulocyte/macrophage colony stimulating factor (GM-CSF) were examined. Immunization of C57BL/6 mice with the CEA-GMCSF fusion plasmids in a three injection, high-dose immunization schedule led to T cell and antibody responses specific for CEA. Mice injected with CEA GMCSF fusion plasmids also developed IgG autoantibodies to GM-CSF. Tumor challenge with the CEA-expressing syngeneic mouse adenocarcinoma line, MC38-CEA 2, showed delayed tumor growth in mice immunized with the CEA-GMCSF fusion plasmids but complete protection in mice immunized with plasmid encoding CEA alone. In contrast, a single low-dose immunization with CEA-GMCSF fusion plasmids provided better tumor protection than low-dose CEA plasmid alone and resulted in lower titers of GM-CSF antibodies. PMID- 15652671 TI - Population-wide benefits of routine vaccination of children against influenza. AB - Using a stochastic simulation model of influenza transmission, clinical illness, and economic costs, we estimated the population-wide benefits of routinely vaccinating US children (ages 6 months to 18 years) against influenza. Disease burden was projected to decline as a result of both reduced susceptibility to infection among vaccinees and reductions in disease transmission to others in the community. Vaccination of 20% of children would reduce the total number of influenza cases in the US by 46%; 80% coverage would reduce the total number of cases by 91%. Similar reductions were estimated to occur in influenza-related mortality and economic costs. PMID- 15652673 TI - A serine protease inhibitor (serpin) from Haemaphysalis longicornis as an anti tick vaccine. AB - The application of anti-tick vaccine has been shown to be the most promising alternative tick control strategy compared to the current use of acaricides that suffer from a number of limitations. The success of this strategy is dependent on the cloning, and characterization of tick molecules involved in the mediation of tick central physiological roles. Rapid amplification of the cDNA ends (RACE) and primers designed based on a conserved serpin amino acid motif (NAVYFKG) were used to clone a cDNA with high similarity in the reactive center loop (RCL) to representative serpin, heparin cofactor II. We have named this novel gene as Haemaphysalis longicornis serpin-2 (HLS2). RT-PCR analysis showed that HLS2 mRNA transcripts are not expressed in salivary glands but in hemolymph by feeding ticks. HLS2 was not introduced into the bite site as measured by Western blot analysis. The activated partial thromboplastin time (APTT) and the thrombin inhibitory assay using recombinant HLS2 (rHLS2) demonstrated prolonged coagulation time and inhibition of thrombin activity. These results suggested that HLS2 is present only in hemolymph of the feeding ticks and the function of HLS2 is homeostasis in tick physiological compartment. Vaccination of rabbits with rHLS2 conferred protective immunity against ticks, resulting in 44.6 and 43.0% mortality in nymphal and adult ticks, respectively. These results show that rHLS2 could be an important candidate as a component of a cocktail anti-tick vaccine. PMID- 15652672 TI - A recombinant P4 protein of Haemophilus influenzae induces specific immune responses biologically active against nasopharyngeal colonization in mice after intranasal immunization. AB - Outer membrane protein P4, together with P6, is highly conserved among all typeable and nontypeable strains of Haemophilus influenzae (H. influenzae). Thus, the protein is an attractive antigen for the inclusion in a vaccine against nontypeable H. influenzae (NTHi). However, the ability of P4 to induce antibodies protective against NTHi infections is still controversial. In this study, we investigated the specific mucosal immune responses against NTHi induced by intranasal immunization with the lipidated form of recombinant P4 protein (rP4) and non-fatty acylated recombinant P6 protein (rP6) with or without cholera toxin (CT) in BALB/c mice model. Intranasal immunization with either rP4+CT, a mixture of rP4 and rP6+CT, or rP4 and rP6 without CT elicited anti-rP4 specific IgG antibody in serum of mice. Intranasal immunization with either rP4+CT or a mixture of rP4, rP6+CT elicited anti-rP4 specific IgA antibody in nasopharyngeal washing (NPW), while intranasal immunization with rP4 and rP6 without CT did not induced anti-rP4 specific IgA antibody responses in NPWs. Sera from mice intranasally immunized with rP4+CT and a mixture of rP4, rP6+CT also showed bactericidal activity. Significant clearance of NTHi in nasopharynx was seen 3 days after the inoculation of live NTHi in mice intranasally immunized with rP4+CT. The current findings suggested that P4 would be a useful antigen as the component of the vaccine to induce protective immune responses against NTHi. The use of an intranasal vaccine composed of the different surface protein antigens is an attractive strategy for the development of a vaccine against NTHi. PMID- 15652674 TI - Interactions of proteoliposomes from serogroup B Neisseria meningitidis with bone marrow-derived dendritic cells and macrophages: adjuvant effects and antigen delivery. AB - Exposure to proteoliposomes from serogroup B Neisseria meningitidis (PL) induced up-regulation of MHC-II, MHC-I, CD40, CD80 and CD86 expression on the surface of murine bone marrow-derived dendritic cells (DC). CD40, CD80 and CD86 were up regulated on bone marrow-derived macrophages (MPhi) upon stimulation with PL. Both DC and MPhi released TNFalpha, but only DC produced IL12(p70) in response to PL. A small increase in the expression of MHC-II, CD40 and CD86, as well as production of IL12(p70), was observed on the cell surface of DC, but not MPhi from LPS-non-responder C3H/HeJ after exposure to PL. DC, but not MPhi, incubated with PL containing ovalbumin (PL-OVA) presented OVA-specific peptides to CD4+ and CD8+ OVA-specific T-cell hybridomas. These data clearly indicate that PL exert an immunomodulatory effect on DC and MPhi, with some contribution of non-LPS components besides the main role of LPS. The work also shows the potential of PL as a general system to deliver antigens to DC for presentation to CD4+ and CD8+ T cells. PMID- 15652675 TI - The purpose of attributing cause: beliefs about the causes of myocardial infarction. AB - Patients' beliefs concerning the causes of their illnesses are core to a number of theoretical models of illness behaviour. The current study explored the themes that emerged from patients' accounts of the causes of their first myocardial infarctions (MIs). Semi-structured interviews were conducted with 12 patients within a week of their MI, at a UK district hospital. Transcripts were analysed using interpretative phenomenological analysis. Three researchers noted themes that emerged from each interview, then common themes were selected and refined on the basis of an aggregation of interviews. Three dominant themes emerged: (1) single versus multiple causation, (2) causes as triggers versus underlying dispositions, and (3) the potentially conflicting motives of avoiding blame whilst at the same time, seeking control. Whilst many participants had complex ideas concerning what caused their MI, and could often name several causes, they tended to emphasise the importance of a single cause, which often related to their symptoms. Further, several participants interpreted "cause" in terms of an acute trigger of MI, rather than as a chronic causal factor. Participants were apparently attempting to answer the question about why they had an MI now, leading to talk about single causes that trigger MI. By contrast, much previous research has been concerned with patients' knowledge of "risk factors", considering the production of only a few causes to reflect ignorance. A key process in participants attributing cause appeared to be attempting to avoid blaming themselves or others for their MI, whilst simultaneously seeking to assert control over future recurrence. Analysis of the functions and purposes of causal attribution suggests that patients' focus on blame and control may be both emotionally and behaviourally adaptive, if not necessarily epidemiologically precise. These findings suggest that interventions to change causal attributions may be misguided, and may even be harmful. PMID- 15652676 TI - Lay perceptions of type 2 diabetes in Scotland: bringing health services back in. AB - The growing prevalence of type 2 diabetes is placing Scottish health services under considerable strain. Consequently, diabetes services are undergoing a major process of reorganisation, including the devolvement of routine diabetes care/diabetic review from secondary to primary healthcare settings. This qualitative study was devised to explore newly diagnosed type 2 diabetes patients' perceptions of their disease and the health services they receive at a time when this restructuring of services is taking place. The sample comprised 40 patients resident in Lothian, Scotland, who had diverse experiences of services, some receiving GP-based care only, others having varying contact with hospital diabetes clinics. In-depth interviews were undertaken with patients, three times at six monthly intervals over 1 year, enabling their experiences to be tracked at critical junctures during the post-diagnostic period. Disease perceptions and health service delivery were found to be mutually informing and effecting. Not only did (different types of) health service delivery influence the ways in which patients thought about and self-managed their disease, over time patients' disease perceptions also informed their expectations of, and preferences for, diabetes services. We thus argue that there is a need for a reconceptualisation within the medical social sciences to take into account the context of healthcare and the economic/policy factors that inform health service delivery when looking at patients' disease perceptions. We also discuss the logistical and ethical challenges of drawing upon patients' perspectives, preferences and views in the design and delivery of future health services. PMID- 15652677 TI - Moral hazard and prescription medicine use in Australia--the patient perspective. AB - All Australian citizens are provided affordable access to prescription medicines through the nation's system of universal pharmaceutical subsidies--the pharmaceutical benefits scheme. The rapid increase in pharmaceutical related expenditure has generated the concern that Australians are taking advantage of prescription subsidies and are using more medicines than are necessary, thereby creating a 'moral hazard'. This concern is predicated on a number of assumptions about patient behaviour rather than on empirical observation. These assumptions amount to a view that patients are consumers who treat prescription medicines as common goods subject to informed and rational calculation of the cost and benefits of their use. This paper reports the findings of an in-depth interview study undertaken to explore how prescription cost influences Australians' medicine use. Qualitative data were analysed to compare medicine users' descriptions of the role of prescription cost in medicine use against the assumptions that underlie the belief in moral hazard. Moral hazard did not appear to be significantly operating in the accounts of medicine use collected for this study. Interviewees' accounts of medicine use revealed an act characterised by ambivalence, a mix of desire and antipathy, faith and suspicion. Medicines appeared in interviewees' accounts as both pharmacologically and symbolically potent substances, which despite their familiarity as objects, are often mysterious to non-expert patients. Cost appeared as a secondary factor in patients' decision to access a prescription medicine. Using a prescription was predicated on the medicine being necessary, with necessity typically established by an expert doctor prescribing the medicine. Prescription medicines did not appear as 'common goods' where subsidised access motivates a 'consumer' to demand more or make the prospect of prescription use more attractive or necessary. PMID- 15652678 TI - Fee-for-service or donation? Hungarian perspectives on informal payment for health care. AB - Informal payments are known to be widespread in the post-communist health care systems of Central and Eastern Europe. However, their role and nature remains contentious with the debate characterized by much polemic. This paper aims to make sense of this debate by reviewing and summarizing the main arguments of the theoretical debate in Hungary. The review examines the possible causes of informal payment, the motivation of the actors involved and the impact of informal payment on system performance, focusing on efficiency and equity. The lines of arguments are summarized in two contrasting hypotheses, which envisage informal payment as either a donation or a fee-for-service. Evidence pertaining to the scale of informal payments and the motivation of patients are reviewed, but found to be inconclusive to verify the hypotheses. Although focused on Hungary, accounts from other countries facing informal payments show similar threads of discussion and dilemmas. These theories should be tested further using evidence from existing studies and new empirical research, since the validity of the gratitude payment concept is a central dilemma of effective policy making in the area. To orient future research, a possible agenda is outlined, which links evidence to be obtained to the defining features of gratitude payments. PMID- 15652679 TI - Whose informational needs are considered? A comparison between cancer patients and their spouses' perceptions of their own and their partners' knowledge and informational needs. AB - The present study examines information exchange patterns between 98 married couples in Israel where one is a cancer patient and the other is the main caregiver. Specifically, the accuracy of each spouse's perception of the extent of knowledge and the need to receive more disease-related information is examined as a function of the role (patient-caregiver) and gender of the participants. The results showed that women, regardless of their role, were inaccurate in their perception of their husbands' knowledge and motivation to know more. For men, a difference between the roles was found for the perception of their wives' knowledge. As caregivers, they were inaccurate while as patients they were accurate in this estimation. Their perception of their wives' needs to know more was accurate. Moreover, female patients, more than male, relied on their perception of themselves when assessing their spouse's knowledge and informational needs. Thus, it is concluded that female patients were more egocentric and their perception of their spouse's preferences was influenced by their own needs. The results demonstrated that in the context of cancer patients and their spouse as caregivers, neither partner considered the informational needs of his or her spouse. PMID- 15652680 TI - Health information-seeking behaviour in adolescence: the place of the internet. AB - The internet is one of a range of health information sources available to adolescents. It is recognised that young people have difficulties accessing traditional health services; in theory, the internet offers them confidential and convenient access to an unprecedented level of information about a diverse range of subjects. This could redress adolescents' state of relative health 'information poverty', compared to adults. This paper seeks to explore United Kingdom (UK) and United States (US) adolescents' perceptions and experiences of using the internet to find information about health and medicines, in the context of the other health information sources that are available to them. The study involved a series of 26 single-gender focus groups with 157 English-speaking students aged 11-19 years from the UK and the US. Many students reported that the internet was their primary general information source. Information sources were defined during analysis in terms of previous experience of the source, saliency of the available information, and credibility of the source (defined in terms of expertise, trustworthiness and empathy). Most focus group participants had extensive personal experience with the internet and some information providers therein (notably search engines). Internet health information was regarded generally as salient. Its saliency was increased through active searching and personalisation. Perceived credibility of the internet varied because expertise and trustworthiness were sometimes difficult to determine, and empathy could be facilitated through online communities but the individual could control disclosure. The internet combines positive features of traditional lay and professional, personal and impersonal sources. Although it is unlikely to supplant the role of trusted peers and adults, the internet has found an important place among adolescents' repertory of health information sources. PMID- 15652681 TI - The impact of an organ donation registration information program for high school students in the Netherlands. AB - In 1998, a new organ donor registration system was implemented in the Netherlands to increase the number of potential donors. A high school education program was developed to prepare adolescents to make an informed decision about organ donation. A post-test only randomised controlled trial was conducted in 39 high schools including 2868 students. Students within schools were randomly allocated to either attend the organ donation education program or not. The impact of the program on students' intention to register their organ donation preference (and determinants thereof) were analysed using multivariate multileveling modelling (MlwiN). The results show that students who were exposed to the education program had more favourable registration intentions (B = .40), were more often willing to be donors (OR = 1.45), and had greater knowledge about (B = 3.84) and more positive social outcome expectations (B = .09) and self-efficacy regarding organ donation registration (B=.22). Lastly, they experienced significantly less negative outcome expectations related to organ donation registration (B = -.15). Students' evaluation of the school-based education program was favourable. The present organ donation registration program proved to be effective in changing determinants of organ donation registration, and a large-scale implementation in the Dutch high school curriculum is planned. PMID- 15652682 TI - Exploring the social and cultural context of sexual health for young people in Mongolia: implications for health promotion. AB - Recent political, economic, and cultural changes in Mongolia make its large proportion of young people vulnerable to HIV infection. While there had been only two clinical cases of HIV in Mongolia by the year 2000, the incidence of sexually transmitted infections (STIs) is on the rise, especially among people aged 15-24. Little is known about the social and cultural context in which the sexual knowledge, attitudes, and behaviour of Mongolian young people are created and negotiated. This context must be better understood in order to promote safer sex practices. This study employed qualitative research methods to explore and describe the social and cultural context in which sexual behaviour is negotiated among secondary school students in Ulaanbaatar, Mongolia. Students and teachers from two schools in Ulaanbaatar and health professionals were selected by purposeful sampling to participate in six semi-structured focus group interviews in autumn 2000. Thematic content analysis was conducted on the focus group transcripts. Seven themes were extracted including embarrassment, lack of knowledge, concepts of sex, perceptions of condoms, gender roles, peer norms, and the influence of drinking on sexual activity. Results presented are the first description of the social and cultural context of sexual health and highlight the combined impact of these themes on safer sex practices in Mongolia. These findings are not generalizable, but their agreement with the Mongolian and the international literature indicates that they may be transferable. Implications for STI and HIV/AIDS prevention efforts and further research in Mongolia are discussed. PMID- 15652683 TI - Association of sociodemographic characteristics of children with intellectual disability in Western Australia. AB - The social determinants of intellectual disability (ID) are poorly understood, particularly in Australia. This study has investigated sociodemographic correlates of ID of unknown cause in Western Australian born children. Using record linkage to the Western Australian Maternal & Child Health Research Database, maternal sociodemographic characteristics of children with ID (of unknown cause) born between 1983 and 1992 (n = 2871) were compared with those of children without ID (n = 236,964). Socioeconomic indices for areas based on the census district of mother's residence were also included in the analysis. Aboriginal mothers (OR = 2.83 [CI: 2.52, 3.18]), teenagers (OR = 2.09 [CI: 1.82, 2.40]) and single mothers (OR = 2.18 [CI: 1.97, 2.42]) were all at increased risk of having a child with mild or moderate ID. Children of mothers in the most socioeconomically disadvantaged 10% had more than five times the risk of mild and moderate ID compared with those in the least disadvantaged 10% (OR = 5.61 [CI: 4.42, 7.12]). Fourth or later born children were also at increased risk (OR = 1.82 [CI: 1.63, 2.02]). The results of the study have implications both for further aetiological investigation as well as service provision for children with ID. Furthermore, many of the sociodemographic correlates identified in this study, particularly in the mild/moderate category of ID, are potentially modifiable, opening up opportunities for primary prevention. PMID- 15652684 TI - Does social inequality matter? Changing ethnic socio-economic disparities and Maori smoking in New Zealand, 1981-1996. AB - This paper builds on an earlier study of the effects of inequality on smoking by explicitly incorporating a temporal dimension. We examine the effects of changing levels of inequality upon ethnic variations in smoking rates in New Zealand for the period 1981 to 1996. This was a period of rapid structural change in New Zealand's economy and welfare state, changes which had a disproportionate effect on Maori. While Maori smoking rates declined during this period, the gap in smoking levels between Maori and Pakeha (persons of European descent) increased. The results suggest that levels of social inequality between Maori and Pakeha have an independent effect on Maori smoking rates and that communities which experienced increased social inequality during both the 1980s and 1990s were more likely to have higher Maori smoking rates. Controlling for confounders, the effect of increased ethnic inequality on smoking was particularly evident for Maori women (net R(2) = 0.150) compared to Maori men (net R(2) = 0.079). Nevertheless, absolute rather than relative socio-economic deprivation remains the most important predictor of smoking. PMID- 15652685 TI - Ethnic differences in social participation and social capital in Malmo, Sweden: a population-based study. AB - The aim of this study was to investigate ethnic differences in different aspects of social participation in Malmo, Sweden. The public health survey in Malmo 1994 is a cross-sectional study. A total of 5600 randomly chosen individuals aged 20 80 years were asked to complete a postal questionnaire. The participation rate was 71%. The population was divided into categories born in Sweden, Denmark/Norway, other Western countries, former Yugoslavia, Poland, Arabic speaking countries and all other countries. The age-adjusted and multivariate analyses were performed using a logistic regression model in order to investigate the importance of possible confounders (age, education, economic stress and unemployment) on the differences by country of origin in different aspects of social participation. Men and women born in Arabic speaking countries and other countries (Iran, Turkey, Vietnam, Chile and subsaharan Africa) participate to a significantly lower extent in a variety of civic and social activities when compared to the reference population born in Sweden. The differences in participation in these groups compared to the group born in Sweden are observed both for social participation items at the core of the definition of social capital and cultural and other activities unrelated to social capital. This pattern is particularly pronounced for women born in Arabic speaking countries. These women even sharply differ from the participation rates of men born in Arabic speaking countries. The ethnic differences in most cases do not seem to be explained satisfactorily by education, economic stress or possibly unemployment. PMID- 15652686 TI - Residential mobility in the California Teachers Study: implications for geographic differences in disease rates. AB - BACKGROUND: Especially for cancers with long latency periods, such as breast cancer, the issue of residential mobility hinders ecologic analyses seeking to examine the role of environmental contaminants in chronic disease etiology. This study describes and evaluates characteristics associated with residential mobility in a sub-sample of the California Teachers Study (CTS) cohort. METHODS: In 2000, lifetime residential histories were collected for a sub-sample of 328 women enrolled in the CTS; women's degree of residential mobility and associated factors were analyzed. RESULTS: While most women moved many times during their lives (average = 8.9), the average number of years at their residence when they enrolled in the study was reasonably long (15.1 years). Age strongly predicted duration at current residence but was not related to the number of lifetime residences. After adjusting for age, California-born women and women living in high socioeconomic status (SES) neighborhoods were significantly more residentially stable. Agreement between self-reported urbanization of recent residences and that based on census data of the geocoded residences was very good (80% concordant). Among women currently living in urban areas, an average of 43.3 years, or 77%, of their lifetimes were spent in urban residences; among women currently living in a rural area, an average of 37.3 years, or 67% of their lifetimes were spent in rural residences. CONCLUSIONS: This suggests that analyses of incidence rates based on current residence, while not capturing a woman's full exposure history, may reasonably reflect some aspect of longer term chronic exposures, especially those related to urbanization, at least in professional women. PMID- 15652687 TI - Neighbourhood inequality, neighbourhood affluence and population health. AB - While there is now considerable evidence that the neighbourhood income levels (poverty/affluence) exert an independent effect on health, there is little evidence that neighbourhood income inequality is consequential, net of individual level socio-economic resources. We show that the usual explanation for the absence of an independent effect of neighbourhood inequality--the assumption of economic homogeneity at the neighbourhood level--cannot account for this result. The authors use hierarchical models that combine individual micro-data from Statistics Canada's 1996/97 National Population Health Survey (NPHS) with neighbourhood and city-level socio-economic characteristics from the 1996 Census of Canada to estimate the effects of neighbourhood affluence and income inequality on self-reported health status. The findings indicate that the negative "ecological" correlation between average neighbourhood health and neighbourhood income inequality is the result not only of compositional differences among individuals but also of contextual neighbourhood effects associated with low and high inequality neighbourhoods. PMID- 15652688 TI - A comparison of generic, indirect utility measures (the HUI2, HUI3, SF-6D, and the EQ-5D) and disease-specific instruments (the RAQoL and the HAQ) in rheumatoid arthritis. AB - Rheumatoid arthritis (RA) is a common, chronic disease where health-related quality of life (HRQL) is one of the main goals of therapy. As such, instruments used to measure HRQL in RA must be able to discriminate across RA severity. The two basic categories of instruments used to measure HRQL are generic instruments and disease-specific instruments. Generic instruments can be further subdivided into preference-based measures which yield both single and multi-attribute utility values anchored at zero (death) and 1.00 (perfect health) as a measure of HRQL. The scores from these types of instruments can be integrated into cost utility analyses as the weightings for quality adjusted life years. We assessed the construct validity of utility scores from four generic preference-based measures (the Health Utilities Index 2 and 3 (HUI2, HUI3), the EuroQol 5D (EQ 5D), and the Short Form 6-D (SF-6D) and disease specific measures (the Rheumatoid Arthritis Quality of Life Questionnaire (RAQoL) and the Health Assessment Questionnaire (HAQ)) in a sample of 313 RA patients in British Columbia, Canada. We also estimated the minimally important differences (MID) for each of the measures. Generally, as anticipated, the disease-specific measures were better able to discriminate across groups with higher RA severity; however, utility scores from each of the scales also appeared to discriminate well across RA severity categories. The MID values agreed with those previously reported in the literature for the HUI2, SF-6D and the HAQ and provided new information for the HUI3, EQ-5D and the RAQoL. We conclude that the all of the preference-based utility measures that were evaluated appear to adequately discriminate across levels of RA severity. PMID- 15652689 TI - Autonomy-related behaviors of patient companions and their effect on decision making activity in geriatric primary care visits. AB - The objective of this study, undertaken in the USA, was to investigate the consequences of autonomy-related companion behaviors on patient decision-making activity during geriatric primary care visits. Videotapes were analyzed to characterize patient and companion decision-making activity and related companion behaviors. These behaviors were coded throughout the visit using an autonomy based framework that included both autonomy enhancing (i.e. facilitating patient understanding, patient involvement, and doctor understanding) and detracting behaviors, (i.e. controlling the patient and building alliances with the physician). Patients (N = 93) in this cross-sectional sample range in age from 65 to 95 years and are mostly white (n = 73, 79%) and female (n = 67, 72%). Companions are spouses (n = 42, 46%), adult children (n = 33, 36%), or other relatives and friends (n = 15, 16%) of patients. Companions are active participants in medical visits and engage in more autonomy enhancing than detracting behaviors. Companions of sicker (compared with less sick) patients were more likely to facilitate patient understanding, p < .05; doctor understanding, p < .01; and patient involvement, p = .06, in care. Patients whose companions facilitated their involvement in the medical visit by asking the patient questions, prompting the patient to talk, and asking for the patient's opinion were more than four times as likely to be active in decision-making as patients whose companions did not assist in this manner (unadjusted OR 3.5, CI 1.4-8.7, p < .01; adjusted OR 4.5, CI 1.6-12.4, p < .01). Companions can play an important role in the visits of geriatric patients by facilitating communication throughout the visit as well as patient activity in decision-making. PMID- 15652690 TI - Self-reported job insecurity and health in the Whitehall II study: potential explanations of the relationship. AB - This paper examines the potential of demographic, personal, material and behavioural characteristics, other psychosocial features of the work environment and job satisfaction to explain associations between self-reported job insecurity and health in a longitudinal study of British white-collar civil servants. Strong associations were found between self-reported job insecurity and both poor self rated health and minor psychiatric morbidity. After adjustment for age, employment grade and health during a prior phase of secure employment, pessimism, heightened vigilance, primary deprivation, financial security, social support and job satisfaction explained 68% of the association between job insecurity and self rated health in women, and 36% in men. With the addition of job control, these factors explained 60% of the association between job insecurity and minor psychiatric morbidity, and just over 80% of the association with depression in both sexes. PMID- 15652691 TI - Psychosocial and cultural factors affecting the perceived risk of genetically modified food: an overview of the literature. AB - The rapid globalization of the world economy has increased the need for an astute understanding of cultural differences in perceptions, values, and ways of thinking about new food technologies. In this paper, we describe how socio psychological and cultural factors may affect public perceptions of the risk of genetically modified (GM) food. We present psychological, sociological, and anthropological research on risk perception as a framework for understanding cross-national differences in reactions to GM food. Differences in the cultural values and circumstances of people in the US, European countries, and the developing world are examined. The implications of cultural theory for risk communication and decision making about GM food are discussed and directions for future research highlighted. PMID- 15652692 TI - AIDS and "shared sovereignty" in Tanzania from 1987 to 2000: a case study. AB - After more than 15 years of foreign assistance to support HIV/AIDS prevention in Sub-Saharan Africa, HIV rates in the sub-continent remain high with only a few examples of reduced HIV incidence. This case study used the frame of "shared sovereignty" between nation-states and official development assistance agencies to analyze 13 years of technical assistance for HIV/AIDS programs in Tanzania from 1987 to 2000. The study draws on 21 key informant interviews and a systematic review of key program documents from the National AIDS Control Programme (NACP) and 14 other international agencies. Applying Jamison et al.'s (Lancet 351 (1998) 514) shared sovereignty framework, the analysis focused on fulfilled shared functions in moving Tanzania's NACP from dependence to independence. The analysis revealed an uneven and inconsistent level of technical assistance to the NACP with a rotation of multilateral and bilateral donors over the period of study. The Tanzanian government was often ambivalent toward agencies providing assistance towards its HIV/AIDS programs and toward its own NACP. Results are discussed in terms of implications for future strategic planning to mitigate the effects of HIV/AIDS. Determining roles, shared accountability and responsibility in a shared sovereignty framework remain a challenge in the governance of HIV/AIDS programs in Tanzania. PMID- 15652694 TI - Identity change and the human dissection experience over the first year of medical training. AB - The aim of this study is to explore identity change in medical students over their first year of medical training, particularly in relation to their experience of human dissection. Each of our four participants completed two repertory grids at the end of term one and, again, towards the end of term three. One grid tapped their identity construction, and the other, their experience of human dissection. Our participants were optimistic about becoming similar to a doctor they admired and, towards the end of term three, began to develop a stable identity as a medical student. Their identity constructs involved three common themes: dedication, competence, and responsibility. However, the data also revealed negative reactions to the demands of training, such as feeling driven and stressed. Three major themes were apparent in their experience of human dissection: involvement, emotional coping, and ability. Our participants' dedication to their studies was reflected in their appreciation of the need to become involved actively in the process of dissection but some experienced an erosion of their self-confidence and perceived some of their colleagues to have lost much of their enthusiasm for learning. Emotional coping could be an additional challenge within this context and their reaction tended to reflect distancing processes previously identified in the literature. In all, we see a development of a vulnerable sense of professionalism alongside a frustration of losing out potentially on wider aspects of personal development due to the high work demands. PMID- 15652693 TI - Cultural differences in conceptual models of depression. AB - Members of ethnic minority groups are less likely than white middle class people to seek professional treatment for depression and other mental health problems. One explanation is that the former conceptualize depressive symptoms as social problems or emotional reactions to situations, while the latter are more apt to view depression as a disease requiring professional treatment. Though considerable evidence supports this hypothesis, it is rarely explored directly through cross-cultural comparisons. The present study compares conceptual models of depressive symptoms in two diverse cultural groups in New York City (USA): 36 South Asian (SA) immigrants and 37 European Americans (EA) were presented with a vignette describing depressive symptoms and participated in a semi-structured interview designed to elicit representational models of the symptoms. Results indicate pervasive differences in representational models across the two groups. SA participants identified the "problem" in the vignette in largely social and moral terms. Suggestions for management and health seeking in this group emphasized self-management and lay referral strategies. EAs, by contrast, often proposed alternate, sometimes contradictory, explanatory models for the depressive symptoms. One model emphasized biological explanations ranging from "hormonal imbalance" to "neurological problem." The second model resembled the "situational stress" or "life problem" model described by SAs. The implications of these findings, and directions for future research, are discussed. PMID- 15652695 TI - Between formal and enacted policy: changing the contours of boundaries. AB - This study examines the strategies of the biomedical discourse vis-a-vis the growing public demand for alternative medicine by comparing formal and informal claims for jurisdiction. The analysis is based on two main sources of data from Israel: (a) two formal position statements, and (b) a series of participant observations and interviews with practitioners in clinical settings where biomedical and alternative practitioners collaborate. At the formal level, the biomedical discourse seeks to secure its dominant position by drawing strict cognitive and moral lines differentiating "proper biomedicine" from "improper alternative medicine." At this level alternative medicine appears morally "contaminated" and its knowledge-base delegitimized by extreme forms of boundary work. At the informal level, the contour of boundaries change. In the hospital field where alternative and biomedical practitioners are collaborating, mutual respect was expressed even as social and symbolic boundaries were being demarcated. Modifying the forms of boundary-work appears to be biomedicine's reactive strategy in the field to changing environmental and market demands. It is a strategy that allows biomedical discourse to absorb its competitor within its professional jurisdiction with no battle, while retaining absolute epistemological hegemony and Institutional Control. PMID- 15652696 TI - Expression of serotonin 5-HT2C receptors in GABAergic cells of the anterior raphe nuclei. AB - We have used double in situ hybridization to examine the cellular localization of 5-HT2C receptor mRNA in relation to serotonergic and GABAergic neurons in the anterior raphe nuclei of the rat. In the dorsal and median raphe nuclei 5-HT2C receptor mRNA was not detected in serotonergic cells identified as those expressing serotonin (5-HT) transporter mRNA. In contrast, 5-HT2C receptor mRNA was found in most GABAergic cells, recognized by the presence of glutamic acid decarboxylase mRNA. Such 5-HT2C receptor-positive GABAergic neurons were mainly located in the intermediolateral and lateral portions of the dorsal raphe and lateral part of the median raphe. The present data give anatomical support to a previous hypothesis that proposed a negative-feedback loop involving reciprocal connections between GABAergic interneurons bearing 5-HT2A/2C receptors and 5-HT neurons in the dorsal raphe and surrounding areas. According to this model, the excitation of GABAergic interneurons through these 5-HT2C (and also 5-HT2A) receptors would result in the suppression of 5-HT cell firing. PMID- 15652697 TI - 5-HT1A receptor mRNA and immunoreactivity in the rat medial septum/diagonal band of Broca-relationships to GABAergic and cholinergic neurons. AB - Activation of 5-HT1A receptors results in a variety of physiological responses, depending on their localization on neurons with different phenotypes in the brain. This study investigated the localization of 5-HT1A receptor mRNA and 5 HT1A receptor immunoreactivity in cell bodies of the rat septal complex using in situ hybridization and immunohistochemistry. In adjacent sections of the medial septum/diagonal band of Broca (MSDB), the distribution of cell bodies expressing 5-HT1A receptor mRNA was closely related to cells labeled with oligonucleotide probes to GAD (glutamic acid decarboxylase), VAChT (vesicular acetylcholine transporter) or parvalbumin mRNA. Using antiserum to GAD and antibodies to GABA, 5-HT1A receptor immunoreactivity was demonstrated in a majority of GABAergic cells in the MSDB. 5-HT1A receptor-immunoreactive GABAergic cells in the MSDB were also demonstrated to contain the calcium-binding protein parvalbumin, a marker for septohippocampal projecting GABAergic neurons. In the lateral septum, 5-HT1A receptor immunoreactivity was colocalized with the calcium-binding protein calbindin D-28k, a marker for septal GABAergic somatospiny neurons. 5-HT1A receptor immunoreactivity was also detected in a subpopulation of VAChT containing cholinergic neurons of the MSDB. In MSDB neurons, colocalization of 5 HT1A and 5-HT2A receptor immunoreactivities was demonstrated. These observations suggest that serotonin via 5-HT1A receptors may represent an important modulator of hippocampal transmission important for cognitive and emotional functions through actions on both GABAergic and cholinergic neurons of the rat septal complex. In addition, 5-HT may exert its effects in the MSDB via cells expressing both 5-HT1A and 5-HT2A receptors. PMID- 15652698 TI - Glial overexpression of heme oxygenase-1: a histochemical marker for early stages of striatal damage. AB - The level of heme oxygenase-1 (HO-1) in the normal striatum is below the limit of immunodetection. However, HO-1 is overexpressed in both neural and non-neural cells in response to a wide range of lesions. We induced different types of lesions affecting the striatal cells or the main striatal afferent systems in rats to investigate if overexpression of HO-1 could be a useful histochemical marker of striatal damage. Thirty-six hours after intrastriatal or intraventricular injection of excitotoxins that affect striatal neurons (ibotenic acid) or of neurotoxins that affect striatal dopaminergic (6-hydroxydopamine) or serotonergic (5,7-dihydroxytriptamine) afferent terminals, or after surgical lesioning of cortico-striatal projections, there was intense induction of striatal HO-1 immunoreactivity (HO-1-ir). Double immunolabeling revealed that the HO-1-ir was located in glial cells. After intrastriatal injection of ibotenic acid, a central zone of neuronal degeneration contained numerous round and pseudopodic HO-1-ir cells, and was surrounded by a ring of HO-1-ir cells, most of which were immunoreactive for astroglial markers. Intraventricular injection of neurotoxins induced astroglial HO-1-ir cells which were more evenly distributed throughout the lesioned or denervated areas. HO-1-ir microglial cells were also observed in areas subjected to mechanical damage. The HO-1-ir was markedly lower or absent 1 week after lesion, and even more so 3 weeks after, although some HO-1 ir cells were still observed after intrastriatal injection of ibotenic acid or surgical corticostriatal deafferentation. The results indicate that determination of glial HO-1-ir is a useful histochemical marker for early stages of striatal damage. PMID- 15652699 TI - Soluble guanylate cyclase and neuronal nitric oxide synthase colocalize in rat nucleus tractus solitarii. AB - Nitric oxide has been implicated in transmission of cardiovascular signals in the nucleus tractus solitarii (NTS). Pharmacological studies suggest that activation of neurons by nitric oxide in the NTS may involve soluble guanylate cyclase (sGC). However, anatomical data supporting this suggestion have not been available. In this study, we tested the hypothesis that neurons and fibers containing neuronal nitric oxide synthase (nNOS) lie in close proximity to those containing sGC and the two enzymes colocalize in some neurons and fibers in the NTS. We perfused six rats and obtained brain stem sections for double immunofluorescent staining utilizing antibodies selective for sGC and for nNOS combined with confocal microscopy. The distribution and staining intensity of nNOS-immunoreactivity (IR) was similar to our earlier reports. IR of sGC was present in cell bodies, proximal dendrites and fibers of many brain stem regions. Strong sGC-IR was noted in the hypoglossal, dorsal motor nucleus of vagus and gracilis nuclei. The NTS exhibited moderate sGC-IR. Superimposed images showed that many NTS neurons contained both nNOS-IR and sGC-IR. The percentage of sGC-IR positive cells that were also nNOS-IR positive differed among NTS subnuclei. Similarly, the percentage of nNOS-IR positive cells that were also sGC positive differed among NTS subnuclei. Fibers stained for both nNOS-IR and sGC-IR were also present in NTS subnuclei. In addition, we identified fibers that were stained for nNOS-IR or sGC-IR alone and often found such singly labeled fibers apposed to each other. These data support our hypothesis and provide anatomical support for the suggestion that nitroxidergic activation of the NTS involves sGC. PMID- 15652700 TI - Neuronal distribution of melanin-concentrating hormone, cocaine- and amphetamine regulated transcript and orexin B in the brain of the Djungarian hamster (Phodopus sungorus). AB - The distribution of melanin-concentrating hormone-, cocaine- and amphetamine regulated transcript- and orexin B-immunoreactive elements as well as their morphological relationships in selected brain structures harbouring the neuroendocrine pathways controlling energy balance and circadian rhythmicity in the Djungarian hamster (Phodopus sungorus) were studied. Cocaine- and amphetamine regulated transcript-(55-102)-immunoreactive perikarya co-expressed melanin concentrating hormone-immunoreactivity in the lateral hypothalamic area, dorsomedial hypothalamic nucleus, zona incerta and posterior hypothalamic area. In addition, arcuate nucleus, hypothalamic periventricular nucleus, Edinger Westphal nucleus, and the rostral aspect of the dorsal raphe nucleus contained cocaine- and amphetamine-regulated transcript-immunoreactive cell profiles. Orexin B-immunoreactive perikarya were distributed in the lateral hypothalamic area, dorsomedial hypothalamic nucleus and retrochiasmatic area. Cells immunoreactive for orexin B did not co-express melanin-concentrating hormone immunoreactivity, but orexin B-immunoreactive fibers had close apposition to many melanin-concentrating hormone-immunoreactive cells. Whereas immunoreactivity for all examined peptides was absent in the suprachiasmatic nucleus, dense and large orexin B-immunoreactive fibers and to a lesser extent melanin-concentrating hormone- and cocaine- and amphetamine-regulated transcript-immunoreactive fibers of smaller size were present in the intergeniculate leaflet and raphe nucleus. These observations in Djungarian hamsters indicate that the neuronal distribution of the examined peptides is strongly conserved between species. In addition, the presence of fibers within the neuronal components of the circadian timing system suggests that they may indirectly influence circadian rhythms. PMID- 15652701 TI - Neuronal NADPH-d/NOS expression in the nodose ganglion of severe hypoxic rats with or without mild hypoxic preconditioning. AB - This study aimed to test the hypothesis that mild hypoxic preconditioning (MHPC) induced NOS expression would attenuate the neuropathological changes in the nodose ganglion (NG) of severe hypoxic exposure (SHE) rats. Thus, the young adult rats were caged in the altitude chamber for 4 weeks prior to SHE for 4 h to gain hypoxic preconditioning. The altitude chamber was used to set the height at the level from 5500 m (0.50 atm; pO2=79 Torr) to 10,000 m (0.27 atm; pO2=43 Torr) for MHPC and SHE, respectively. The experimental animals were allowed to survive for 0, 7, 14, 30 and 60 successive days, respectively. Nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) histochemistry and neuronal nitric oxide synthase (nNOS) immunohistochemistry were used to detect NADPH-d/nNOS reactivity in the NG at various time points following hypoxic exposure. The present results showed that about 38% of the neurons in the NG displayed NADPH d/nNOS positive [NADPH-d/nNOS(+)] in normoxic rats. In SHE rats, a peak in the percentage (71%) and staining intensity (230%) of NADPH-d/nNOS(+) nodose neurons at 0 day, which then gradually decreased at 7-60 days. About 25% of the nodose neurons died 60 days after SHE. However, in MHPC rats subjected to SHE, NADPH d/nNOS(+) neurons peaked in the percentage (51%) and staining intensity (171%) at 0 day, which then decreased at 7-60 days. In addition, neuronal survival was markedly increased by MHPC. These results suggested that MHPC might have a neuroprotective effect that reduces the susceptibility of the nodose neurons to NOS mediated neuropathy subsequent to SHE. PMID- 15652702 TI - The T-box transcription factor Tbx15 is required for skeletal development. AB - During early limb development several signaling centers coordinate limb bud outgrowth as well as patterning. Members of the T-box gene family of transcriptional regulators are crucial players in these processes by activating and interpreting these signaling pathways. Here, we show that Tbx15, a member of this gene family, is expressed during limb development, first in the mesenchyme of the early limb bud, then during early endochondral bone development in prehypertrophic chondrocytes of cartilaginous templates. Expression is also found in mesenchymal precursor cells and prehypertrophic chondrocytes, respectively, during development of skeletal elements of the vertebral column and the head. Analysis of Tbx15 null mutant mice indicates a role of Tbx15 in the development of skeletal elements throughout the body. Mutants display a general reduction of bone size and changes of bone shape. In the forelimb skeleton, the scapula lacks the central region of the blade. Cartilaginous templates are already reduced in size and show a transient delay in ossification in mutant embryos. Mutants show a significantly reduced proliferation of prehypertrophic chondrocytes as well as of mesenchymal precursor cells. These data suggest that Tbx15 plays an important role in the development of the skeleton of the limb, vertebral column and head by controlling the number of mesenchymal precursor cells and chondrocytes. PMID- 15652703 TI - Requirement of mesodermal retinoic acid generated by Raldh2 for posterior neural transformation. AB - Studies in amphibian embryos have suggested that retinoic acid (RA) may function as a signal that stimulates posterior differentiation of the nervous system as postulated by the activation-transformation model for anteroposterior patterning of the nervous system. We have tested this hypothesis in retinaldehyde dehydrogenase-2 (Raldh2) null mutant mice lacking RA synthesis in the somitic mesoderm. Raldh2(-/-) embryos exhibited neural induction (activation) as evidenced by expression of Sox1 and Sox2 along the neural plate, but differentiation of spinal cord neuroectodermal progenitor cells (posterior transformation) did not occur as demonstrated by a loss of Pax6 and Olig2 expression along the posterior neural plate. Spinal cord differentiation in Raldh2(-/-) embryos was rescued by maternal RA administration, and during the rescue RA was found to act directly in the neuroectoderm but not the somitic mesoderm. RA generated by Raldh2 in the somitic mesoderm was found to normally travel as a signal throughout the mesoderm and neuroectoderm of the trunk and into tailbud neuroectoderm, but not into tailbud mesoderm. Raldh2(-/-) embryos also exhibited increased Fgf8 expression in the tailbud, and decreased cell proliferation in tailbud neuroectoderm. Our findings demonstrate that RA synthesized in the somitic mesoderm is necessary for posterior neural transformation in the mouse and that Raldh2 provides the only source of RA for posterior development. An important concept to emerge from our studies is that the somitic mesodermal RA signal acts in the neuroectoderm but not mesoderm to generate a spinal cord fate. PMID- 15652704 TI - Intestinal growth and differentiation in zebrafish. AB - Intestinal development in amniotes is driven by interactions between progenitor cells derived from the three primary germ layers. Genetic analyses and gene targeting experiments in zebrafish offer a novel approach to dissect such interactions at a molecular level. Here we show that intestinal anatomy and architecture in zebrafish closely resembles the anatomy and architecture of the mammalian small intestine. The zebrafish intestine is regionalized and the various segments can be identified by epithelial markers whose expression is already segregated at the onset of intestinal differentiation. Differentiation of cells derived from the three primary germ layers begins more or less contemporaneously, and is preceded by a stage in which there is rapid cell proliferation and maturation of epithelial cell polarization. Analysis of zebrafish mutants with altered epithelial survival reveals that seemingly related single gene defects have different effects on epithelial differentiation and smooth muscle and enteric nervous system development. PMID- 15652705 TI - The tumor suppressor gene fat modulates the EGFR-mediated proliferation control in the imaginal tissues of Drosophila melanogaster. AB - Molecules involved in cell adhesion can regulate both early signal transduction events, triggered by soluble factors, and downstream events involved in cell cycle progression. Correct integration of these signals allows appropriate cellular growth, differentiation and ultimately tissue morphogenesis, but incorrect interpretation contributes to pathologies such as tumor growth. The Fat cadherin is a tumor suppressor protein required in Drosophila for epithelial morphogenesis, proliferation control and epithelial planar polarization, and its loss results in a hyperplastic growth of imaginal tissues. While several molecular events have been characterized through which fat participates in the establishment of the epithelial planar polarity, little is known about mechanisms underlying fat-mediated control of cell proliferation. Here we provide evidence that fat specifically cooperates with the epidermal growth factor receptor (EGFR) pathway in controlling cell proliferation in developing imaginal epithelia. Hyperplastic larval and adult fat structures indeed undergo an amazing, synergistic enlargement following to EGFR oversignalling. We further show that such a strong functional interaction occurs downstream of MAPK activation through the transcriptional regulation of genes involved in the EGFR nuclear signalling. Considering that fat mutation shows di per se a hyperplastic phenotype, we suggest a model in which fat acts in parallel to EGFR pathway in transducing different cell communication signals; furthermore its function is requested downstream of MAPK for a correct rendering of the growth signals converging to the epidermal growth factor receptor. PMID- 15652706 TI - A cytokine secreted from the suboesophageal body is essential for morphogenesis of the insect head. AB - The suboesophageal body of insects was identified over a century ago in the silkworm embryo, but its biological function is still unknown. We discovered that this tissue is differentiated in the earliest embryonic stages of the cabbage armyworm and secretes the insect cytokine, growth-blocking peptide (GBP), transiently from 24 to 60 h after oviposition when gastrulation is in progress. Over-expression of GBP, achieved by microinjection of the GBP gene driven by a cytomegalovirus (CMV) constitutive promoter, resulted in complex deformities of the procephalon (embryonic head). Severe abnormal phenotypes of the head structure were produced by silencing the GBP expression in the embryo by treating with GBP double-stranded RNA: the procephalon-containing optic lobes diminished and completely separated into bilateral halves. This indicates that GBP secreted from the suboesophageal body plays an essential role in the formation of the procephalic domain during early embryogenesis. The cytokine-induced fusion of bilateral procephalic lobes is thought to be evolutionarily conserved at least in insects, because of the widespread occurrence of the suboesophageal body in insect embryos. PMID- 15652707 TI - Tbx1 is required for proper neural crest migration and to stabilize spatial patterns during middle and inner ear development. AB - Tbx1 belongs to the family of T-box containing transcription factors. In humans, TBX1 is implicated in the etiology of the DiGeorge syndrome. Inactivation of the Tbx1 gene in mice produces a variety of malformations including abnormal branching of the heart outflow tract, deficiencies in the branchial arch derivatives, agenesis of pharyngeal glands and abnormal development of the auditory system. We analyze here the middle and inner ear phenotypes of the Tbx1 null mice. The middle ear is strongly affected. Its skeletal components are malformed to varying degrees, some being slightly hypoplastic and others completely absent. However, a seemingly normal-looking tympanic membrane can still be recognized. Middle ear anomalies are associated with other skeletal deficiencies in the branchial arch-derived skeleton. These phenotypes derive from a combination of the failure of the posterior branchial arches to develop and the misrouting of neural crest cells. The inner ears of Tbx1(-/-) animals are hypoplastic. No vestibular or cochlear structures are detectable, but the endolymphatic duct, the cochleovestibular ganglia and residual sensory patches are still identifiable. Molecular analyses revealed a seemingly normal spatial distribution of a variety of patterning markers in the otic vesicles of Tbx1 null mutants at E9.0. However, 1 day later, several of these markers presented altered domains of expression in the otocysts of these mutant embryos, suggesting that Tbx1 is not required for the establishment of spatial patterns in the otocyst, but rather for their maintenance. The inability of the Tbx1(-/-) embryos to keep properly segregated functional domains in the otocyst is likely the cause of the strong inner ear phenotypes observed in these mutants. PMID- 15652708 TI - Genomic imprinting of IGF2, p57(KIP2) and PEG1/MEST in a marsupial, the tammar wallaby. AB - Genomic imprinting is widespread amongst mammals, but has not yet been found in birds. To gain a broader understanding of the origin and significance of imprinting, we have characterized three genes, from three separate imprinted clusters in eutherian mammals in the developing fetus and placenta of an Australian marsupial, the tammar wallaby Macropus eugenii. Imprinted gene orthologues of human and mouse p57(KIP2), IGF2 and PEG1/MEST genes were isolated. p57(KIP2) did not show stable monoallelic expression suggesting that it is not imprinted in marsupials. In contrast, there was paternal-specific expression of IGF2 in almost all tissues, but the biased paternal expression of IGF2 in the fetal head and placenta, demonstrates the occurrence of tissue-specific imprinting, as occurs in mice and humans. There was also paternal-biased expression of PEG1/MESTalpha. The differentially methylated region (DMR) of the human and mouse PEG1/MEST promoter is absent in the wallaby. These data confirm the existence of common imprinted regions in eutherians and marsupials during development, but suggest that the regulatory mechanisms that control imprinted gene expression differ between these two groups of mammals. PMID- 15652709 TI - In vivo inhibition of endogenous brain tumors through systemic interference of Hedgehog signaling in mice. AB - The full spectrum of developmental potential includes normal as well as abnormal and disease states. We therefore subscribe to the idea that tumors derive from the operation of paradevelopmental programs that yield consistent and recognizable morphologies. Work in frogs and mice shows that Hedgehog (Hh)-Gli signaling controls stem cell lineages and that its deregulation leads to tumor formation. Moreover, human tumor cells require sustained Hh-Gli signaling for proliferation as cyclopamine, an alkaloid of the lily Veratrum californicum that blocks the Hh pathway, inhibits the growth of different tumor cells in vitro as well as in subcutaneous xenografts. However, the evidence that systemic treatment is an effective anti-cancer therapy is missing. Here we have used Ptc1(+/-); p53( /-) mice which develop medulloblastoma to test the ability of cyclopamine to inhibit endogenous tumor growth in vivo after tumor initiation through intraperitoneal delivery, which avoids the brain damage associated with direct injection. We find that systemic cyclopamine administration improves the health of Ptc1(+/-);p53(-/-) animals. Analyses of the cerebella of cyclopamine-treated animals show a severe reduction in tumor size and a large decrease in the number of Ptc1-expressing cells, as a readout of cells with an active Hu-Gli pathway, as well as an impairment of their proliferative capacity, always in comparison with vehicle treated mice. Our data demonstrate that systemic treatment with cyclopamine inhibits tumor growth in the brain supporting its therapeutical value for human HH-dependent tumors. They also demonstrate that even the complete loss of the well-known tumor suppressor p53 does not render the tumor independent of Hh pathway function. PMID- 15652710 TI - caudal is required for gnathal and thoracic patterning and for posterior elongation in the intermediate-germband cricket Gryllus bimaculatus. AB - Although the molecular mechanisms directing anteroposterior patterning of the Drosophila embryo (long-germband mode) are well understood, how these mechanisms were evolved from an ancestral mode of insect embryogenesis remains largely unknown. In order to gain insight into mechanisms of evolution in insect embryogenesis, we have examined the expression and function of the orthologue of Drosophila caudal (cad) in the intermediate-germband cricket Gryllus bimaculatus. We observed that a posterior (high) to anterior (low) gradient in the levels of Gryllus bimaculatus cad (Gb' cad) transcript was formed in the early-stage embryo, and then Gb' cad was expressed in the posterior growth zone until the posterior segmentation was completed. Reduction of Gb' cad expression level by RNA interference resulted in deletion of the gnathum, thorax, and abdomen in embryos, remaining only anterior head. We found that the gnathal and thoracic segments are formed by Gb' cad probably through the transcriptional regulation of gap genes including Gb' hunchback and Gb' Kruppel. Furthermore, Gb' cad was found to be involved in the posterior elongation, acting as a downstream gene in the Wingless/Armadillo signalling pathways. These findings indicate that Gb' cad does not function as it does in Drosophila, suggesting that regulatory and functional changes of cad occurred during insect evolution. Since Wnt/Cdx pathways are involved in the posterior patterning of vertebrates, such mechanisms may be conserved in animals that undergo sequential segmentation from the posterior growth zone. PMID- 15652711 TI - Juxtaposition between two cell types is necessary for dorsal appendage tube formation. AB - The Drosophila egg chamber provides an excellent model for studying the link between patterning and morphogenesis. Late in oogenesis, a portion of the flat follicular epithelium remodels to form two tubes; secretion of eggshell proteins into the tube lumens creates the dorsal appendages. Two distinct cell types contribute to dorsal appendage formation: cells expressing the rhomboid-lacZ (rho lacZ) marker form the ventral floor of the tube and cells expressing high levels of the transcription factor Broad form a roof over the rho-lacZ cells. In mutants that produce defective dorsal appendages (K10, Ras and ectopic decapentaplegic) both cell types are specified and reorganize to occupy their stereotypical locations within the otherwise defective tubes. Although the rho-lacZ and Broad cells rearrange to form a tube in wild type and mutant egg chambers, they never intermingle, suggesting that a boundary exists that prevents mixing between these two cell types. Consistent with this hypothesis, the Broad and rho-lacZ cells express different levels of the homophilic adhesion molecule Fasciclin 3. Furthermore, in the anterior of the egg, ectopic rhomboid is sufficient to induce both cell types, which reorganize appropriately to form an ectopic tube. We propose that signaling across a boundary separating the rho-lacZ and Broad cells choreographs the cell shape-changes and rearrangements necessary to transform an initially flat epithelium into a tube. PMID- 15652712 TI - A year is a terrible thing to waste: early experience with HIPAA. PMID- 15652713 TI - Body mass index and mortality: a meta-analysis based on person-level data from twenty-six observational studies. AB - PURPOSE: For this report, we examined the relationships between the conditions of being overweight and obese and mortality from all causes, heart disease, cardiovascular disease, and cancer. METHODS: We defined the categories of body weight according to level of body mass index, BMI=wt(kg)/ht(m)2, using classifications suggested by the National Institutes of Health and the World Health Organization. These classifications are as follows: "normal weight" is defined as BMI > or = 18.5, but less than 25; "overweight" equals BMI > or = 25, but less than 30; and "obese" individuals have BMIs > or = 30. Our investigation is based on person-level data from 26 observational studies that include both genders, several racial and ethnic groups, and samples from the US and other countries. The database consists of 74 analytic cohorts, arranged according to natural strata including gender, race, and area of residence. It includes 388,622 individuals, with 60,374 deaths during follow-up. We use proportional hazards models to examine the relationships between the BMI categories and mortality, controlling for age and smoking status. We use random-effects models to assess summary relative risks associated with the overweight and obesity conditions across cohorts. RESULTS: The relative risks among the heaviest individuals for overall death, death caused by coronary heart disease (CHD), and death caused by cardiovascular disease (CVD) are 1.22, 1.57, and 1.48, respectively, when compared with the those within the lowest BMI category. The summary relative risk among the heaviest participants for death from cancer is 1.07. CONCLUSIONS: We document once again, excess mortality associated with obesity. Our results do, however, question whether the current classification of individuals as "overweight" is optimal in the sense, since there is little evidence of increased risk of mortality in this group. PMID- 15652714 TI - Attrition in a population-based cohort eight years after baseline interview: The Cornella Health Interview Survey Follow-up (CHIS.FU) Study. AB - PURPOSE: To examine how response at follow-up varied from baseline sociodemographic data in a Spanish population-based cohort after 8 years of follow-up. METHODS: The Cornella Health Interview Survey Follow-up (CHIS.FU) Study is a population-based cohort study on lifestyle risk factors and their consequences on health status with 2500 participants at baseline. We have compared the distribution of baseline characteristics according to the results at follow-up (interview, decease, migration, or refusal). RESULTS: Almost two-thirds of the subjects who did not respond to the follow-up interview had died or moved to another town. Sex was a determinant of attrition in deceased and non-traced participants. Refusal appeared to be associated with working status and place of birth. Self-perceived health was one of the characteristics associated with mortality; subjects who perceived their health as poor were 2.6 times more likely to die than those who felt they were in good health. Disabled and retired subjects together with housewives showed a higher risk of dying than individuals still working. The determinants of attrition among emigrated subjects were civil status, age, level of studies, working status, and birth place. CONCLUSION: Although the attrition was non-random, there was no serious bias in estimates of change and in determinants of change due to attrition. PMID- 15652715 TI - Participation and retention in a study of female condom use among women at high STD risk. AB - PURPOSE: Differential participation and retention can bias the findings of a follow-up study. This problem was evaluated in a study of barrier contraception among women at high STD risk. The goal of this study was to identify predictors of participation and retention and determine whether they could influence study results. METHODS: Six-month follow-up study of women attending STD clinics. Determinants of participation and retention were evaluated using logistic and proportional hazards models. RESULTS: Agreement to participate was associated with young age, black race, low education and income, older age at first intercourse, the number of lifetime partners, and STD history. Early attrition was associated with young age, non-black race, higher income, lack of interest/commitment to using the female condom, high coital frequency, no STD history, not using a birth control method at baseline, and with inconsistent condom use, high coital frequency, and pregnancy during follow up. CONCLUSIONS: There was little evidence that differential participation influenced the validity of the study. Differential attrition may have biased behavioral measures of intervention effectiveness, but not necessarily measures of condom use effectiveness. PMID- 15652716 TI - Mortality patterns among workers in a US pharmaceutical production plant. AB - PURPOSE: To examine mortality among workers in a pharmaceutical production plant and to address community concerns about 1980 to 1990 increases in local county cancer mortality rates. METHODS: Subjects were 1999 workers with some full-time employment during the period between 1970 and 1996. We identified deaths through the year 2000 and reconstructed exposures to nine chemical agents with available exposure measurements. Data analyses included standardized mortality ratios (SMRs) and time trends in local cancer mortality rates. RESULTS: We observed deficits in deaths from all causes combined, all cancers combined, and most cause of death categories examined. Male workers with potential plant exposure had excesses in deaths from all lymphatic-hematopoietic tissue cancers (LHTC), in particular non-Hodgkin's lymphoma (NHL), and respiratory system cancers (RSC) that were larger among long-term workers, but the pattern of findings suggested the excesses were probably not related to occupational factors at the plant. The increase in local county cancer mortality rates was simply the upward cycle of a periodic trend that peaked in 1990 and returned to 1980 levels in 2000. CONCLUSIONS: With the possible exceptions of LHTC, in particular NHL, and RSC, this study provided no evidence of elevated total or cause-specific cohort mortality risks. It does not appear that plant factors played a role in the 1980 to 1990 increases in local county cancer mortality rates. PMID- 15652717 TI - Body mass index and breast cancer risk in African American women. AB - PURPOSE: Studies of the relationship between body mass index (BMI) and breast cancer in African American women have been few. We conducted a case-control study to examine whether BMI is associated with risk of breast cancer in this population. METHODS: Cases were 304 women diagnosed with breast cancer at the ages of 20 to 64 years. Controls were 305 women without a history of breast cancer. Telephone interviews were conducted to collect data on history of exposure to various factors at or before the date of diagnosis in cases or equivalent date in controls (reference date). Using logistic regression, we compared cases and controls in BMI at age 18, BMI at the reference date, and change in BMI between the two dates. RESULTS: Using BMI at reference date, we found an odds ratio (OR) of 1.75 (95% confidence interval [CI], 1.02-3.02) and 2.32 (95% CI, 1.33-4.03) for women with BMI of 25 to 29.9 and 30 or higher, respectively, compared with women having BMI lower than 25. The corresponding OR estimates for BMI at age 18 were not significantly different from the unity. The average annual change in BMI between age 18 and date of diagnosis or reference date was associated with breast cancer risk, as shown that more BMI change tended to increase breast cancer risk compared with the baseline quartile of change. When data were analyzed by menopausal status, the association was found for both post-menopausal and pre-menopausal tumors for BMI at reference date but not for BMI at age 18. There was a higher risk for more annual BMI change compared with the baseline for both pre-menopausal and post-menopausal women. CONCLUSION: Our results suggest that BMI at reference date and change in BMI were associated with increased risk of breast cancer in African American women, and the association might be found for both post-menopausal and pre-menopausal tumors. PMID- 15652718 TI - An application of a weighting method to adjust for nonresponse in standardized incidence ratio analysis of cohort studies. AB - PURPOSE: Cohort studies often conduct periodic follow-up interviews (or waves) to determine disease incidence since the previous follow-up and to update measures of exposure and confounders. The common practice of excluding nonrespondents from standardized incidence ratio (SIR) analyses of these cohorts can bias the estimates of interest if nonrespondents and respondents differ on important characteristics related to outcomes of interest. We propose an analytic approach to reduce the impact of nonresponse in the analyses of SIRs. METHODS: Logistic regression models controlling baseline information are used to estimate the propensity, or the probability of response; the reciprocals of these propensities are used as weights in the analysis of risk. This is illustrated in the analysis of 15 years of follow-up of a cohort of US radiologic technologists after an initial interview to assess the risk at several cancer sites from occupational radiation exposure. We use information from the baseline survey and certification records to compute the propensity of responding to the second survey. SIRs are computed using Surveillance, Epidemiology, and End Results (SEER) cancer incidence rates. Variances of the SIRs are estimated by a jackknife method that accounts for additional variability resulting from estimation of the weights. RESULTS: We find that, in this application, weighting alters point estimates and confidence limits only to a small degree, thus providing reassurance that the results are robust to nonresponse. This indicates that results from the analyses excluding the missing data may be slightly biased and weighting helps in reducing the nonresponse bias. CONCLUSION: This method is flexible, practical, easy to use with existing software, and is applicable to missing data from cohorts with baseline information on all subjects. PMID- 15652720 TI - A survey of depressive symptoms among South Korean adults after the Korean financial crisis of late 1997: prevalence and correlates. AB - PURPOSE: We examined the prevalence of high levels of depressive symptoms and their association with sociodemographic and health characteristics in an urban Korean population following the financial crisis in late 1997. METHODS: Using data from a cross-sectional survey conducted in Ansan-city, Korea, from June 1999 to June 2000, we examined the prevalence and correlates of depressive symptoms in a randomly selected sample of 4897 (2531 male and 2366 female) subjects aged 18 to 92 years. The prevalence and odds ratios of "probable" and "definite" depressive symptoms, defined as a Center for Epidemiologic Studies Depression Scale (CES-D) score of > or = 16 and > or = 24, respectively, were calculated. RESULTS: The mean CES-D score and prevalence of "probable" and "definite" depression were significantly higher in women (15.63, 41.67%, and 12.05%, respectively) than in men (14.43, 35.05%, and 8.10%, respectively) (p < 0.0001). In multiple logistic regression analysis, being female, unemployed, unmarried, self-assessed as unhealthy, and having a low income were found to be significant predictors of "probable" and "definite" depressive symptoms in Korean adults. CONCLUSIONS: Although current data cannot be directly compared with those reported in the most recently published Korean data from 1994, considerably high mean CES-D score and prevalence of depression speculate that the Korean financial crisis of 1997 had an effect on the development of depressive symptoms in Korean adults. PMID- 15652719 TI - Application of case-crossover and case-time-control study designs in analyses of time-varying predictors of T-cell homeostasis failure. AB - PURPOSE: To evaluate the association of sexual behavior and recreational drug exposures with T-cell homeostasis failure (TCHF), which corresponds to the onset of a rapid decline in an individual's T lymphocyte count, which occurs on average approximately 1.75 years prior to an initial diagnosis of acquired immunodeficiency syndrome (AIDS). METHODS: A case-crossover design and a case time-control design, both nested within the Multicenter AIDS Cohort Study of 4954 homosexual and bisexual men initiated in 1983. RESULTS: In the case-crossover analysis, use of both recreational drugs and hashish were found to be protective against TCHF (odds ratios < or = 0.41), based on comparisons with four earlier control periods. However, a significant decreasing trend in the prevalence of these exposures was observed over time, thus motivating the implementation of the case-time-control design. Using the latter approach, the associations of drug use (odds ratio=0.53; 95% confidence interval (CI): 0.22, 1.28) and hashish use (odds ratio=0.46; 95% CI: 0.20, 1.05) with TCHF were no longer statistically significant. CONCLUSIONS: The difference in inferences between these approaches demonstrates the importance of evaluating temporal trends in exposures when using a case-crossover design. PMID- 15652722 TI - Evaluating the utility and accuracy of a reverse telephone directory to identify the location of survey respondents. AB - PURPOSE: To investigate the utility and positional accuracy of a reverse telephone directory to enhance geocoding using self-reported street addresses. METHODS: This cross-sectional study used 2001 self-reported survey data from 2636 participants in three Missouri areas. When available, street addresses were appended to participant telephone numbers using a reverse telephone directory. The odds of finding a telephone number in the reverse directory and the positional accuracy between self-reported addresses and those obtained from the reverse directory were calculated. We also determined the quality of self reported address information and that obtained by means of the reverse telephone directory. RESULTS: Rural respondents, younger respondents, women, African Americans, and respondents with less than a high school education were less likely to have their telephone number present in the reverse directory. Using the reverse directory increased the overall percentage of respondents whose addresses were geocoded from 51.5% to 72.0%. Eighty-one percent of addresses were geocoded to the same US Census Block Group and 89% were geocoded to the same Census Tract as the self-reported addresses. CONCLUSIONS: The street address of survey participants obtained through the reverse directory can be used to augment the unknown location of telephone survey respondents but specific groups of people are less likely to be found in the directory. PMID- 15652721 TI - Calpain-10 gene polymorphisms and type 2 diabetes in West Africans: the Africa America Diabetes Mellitus (AADM) Study. AB - PURPOSE: To investigate whether the three single nucleotide polymorphisms (SNPs), SNP-43, -56, and -63 of CAPN10 were associated with type 2 diabetes in a West African cohort. METHODS: A total of 347 diabetic subjects and 148 unaffected controls from four ethnic groups in two West African countries were enrolled in this study. After genotyping three SNPs of CAPN10 and one SNP from CYP19, the allele, genotype, and haplotype frequencies as well as the odds ratios were calculated to test their association with type 2 diabetes. RESULTS: None of the alleles or genotypes was associated with type 2 diabetes. Although statistical analysis indicated that haplotype 221 was associated with type 2 diabetes (OR, 3.765; 95% CI, 1.577-8.989) in the two ethnic groups of Nigeria, the same haplotype did not show any association with type 2 diabetes in the two ethnic groups in Ghana (OR, 0.906; 95% CI, 0.322-2.552). CONCLUSION: Considering the relatively low frequency of haplotype 221 and that none of the haplotypes including 221 was associated with any of the diabetes-related quantitative traits tested, it is concluded that SNP-43, -56, and -63 of the CAPN10 gene variants may play a limited role in the risk of type 2 diabetes risks in this cohort of West Africans. PMID- 15652723 TI - Gender differences in risk factors for attempted suicide among young adults: findings from the Third National Health and Nutrition Examination Survey. AB - PURPOSE: To identify the significant factors associated with attempted suicide among men and women, and determine whether socioeconomic status (SES) and social support indictors, health risk factors, and lifetime history of medical and psychiatric illnesses can explain gender differences in attempted suicide. METHODS: We used data from 3357 men and 4004 women aged 17 to 39 years, who completed a mental disorder diagnostic interview as a part of the Third National Health and Nutrition Examination Survey, 1988-1994. Adjusted odds ratios (ORs) were calculated for the association between risk factors and attempted suicide. RESULTS: The prevalence of lifetime attempted suicides was 7.58% (SE, 0.66) in women and 3.69% (SE, 0.49) in men. In men, low income and smoking were associated with attempted suicide, while attempted suicide in women was associated with poor self-evaluated health, low educational attainment, and drug use. A history of medical and psychiatric illnesses was associated with attempted suicide in both genders, for cancer/pulmonary disease, OR=2.89 (95% CI, 1.25-6.67) in men and 1.94 (1.09-3.45) in women; for major depressive disorder, OR=9.86 (5.08-19.14) in men and 5.00 (3.19-7.83) in women. The significant gender difference of attempted suicide prevalence remained after being adjusted for risk factors selected. CONCLUSION: There were significant gender differences in the risk factors for attempted suicide among young adults, and the gender difference in the prevalence of lifetime attempted suicides could not be explained by differential exposure to risk factors selected. PMID- 15652724 TI - Quality of life in the elderly with epilepsy. PMID- 15652725 TI - The use of diet in the treatment of epilepsy. AB - Fasting and other dietary regimens have been used to treat epilepsy since biblical times. The ketogenic diet, which mimics the metabolism of fasting, was used by modern physicians to treat intractable epilepsy beginning in the 1920s. With the rising popularity of drug treatments however, the ketogenic diet lost its previous status and was used in only a handful of clinics for most of the 20th century. The diet regained widespread recognition as a viable treatment option beginning in 1992 due to the efforts of parent advocate groups. Despite challenges to implementation of the treatment, the ketogenic diet has significant potential as a powerful tool for fighting epilepsy. PMID- 15652726 TI - The problem of psychogenic symptoms: is the psychiatric community in denial? AB - Psychogenic symptoms are common and pose an uncomfortable challenge. Among psychogenic symptoms, psychogenic nonepileptic seizures (PNES) are common and have been extensively studied. They are unique in that, unlike most other psychogenic symptoms, they can be diagnosed with near certainty. PNES can be used as a model, as almost everything that applies to PNES applies to other psychogenic symptoms. According to DSM-IV, somatic symptoms are the main manifestation of three groups of disorders: somatoform disorders, factitious disorder, and malingering. Treatment is challenging. Unfortunately, psychogenic symptoms tend to be neglected. For example, the American Psychiatric Association has abundant written patient education material available on diverse topics, but none on somatoform disorders. Psychogenic symptoms are also not the subject of much clinical research. A search of the journal Neurology for 1994-2003 for the word psychogenic in the title found 21 articles, only 4 of which on topics other than psychogenic seizures. A similar search for original articles in the New England Journal of Medicine found no articles with psychogenic in the title and two with psychogenic in the abstract. Thus, there seems to be a severe disconnect between the frequency of the problem and the amount of attention devoted to it. PMID- 15652727 TI - The electroencephalogram in aviation medical screening: introduction. PMID- 15652728 TI - The electroencephalogram as a screening tool in pilot applicants. AB - The electroencephalogram (EEG) is used to screen pilot candidates for professional licensure irrespective of medical history in many European countries; applicants with paroxysmal abnormalities are excluded. The aim is to reduce the risk of later seizures in flight, which may cause accident or death, but there is no clear evidence that EEG screening can lead to any significant risk reduction. This is partly due to the low predictive value of the EEG, the low risk of seizure in healthy young adults, and the low risk of commercial aircraft accidents. Later-onset seizures, unrelated to the results of earlier screening, cannot be predicted or prevented by screening. Some benefit cannot be excluded, but may be difficult to demonstrate in prospective studies. Specialized screening of targeted populations, such as for photosensitivity in helicopter pilot candidates, may be justifiable; prospective studies with standardized recording and interpretation protocols are needed to assess this. PMID- 15652729 TI - Intermittent photic stimulation as an activation method for electroencephalographic screening of aircrew applicants. AB - Disqualifying criteria for aircrew in Europe (JAR-FCL 3) are, besides a diagnosis of epilepsy after the age of 5 and a history of episode(s) of disturbance of consciousness, epileptiform paroxysmal electroencephalographic abnormalities and focal slow waves. Intermittent photic stimulation (IPS) provokes in about 0.5% of healthy subjects (range 0-2%) a photoparoxysmal response and is most often the only abnormality (70-90%). The literature is scarce and shows great diversity in methodology. Standardized IPS with simultaneous video will not only allow collection of sufficient data for proper epidemiological studies, but can also reveal clinical and often unnoticed or misinterpreted signs and symptoms like myoclonia, loss of consciousness, and occipital seizures with visual auras. The pilot (sleep deprivation, strong sunlight) and the traffic controller (stress, monitors) are more prone to visually induced seizures. Furthermore, the increasing exposure to potentially seizure-triggering visual stimuli might have its impact in a more indirect or cumulative way. PMID- 15652730 TI - Proposal for electroencephalogram standardization in aircrew selection. AB - Current diagnostic electroencephalogram (EEG) investigations in aircrew selection and certification lack both standardization and reference to universally applicable criteria for their effective use. Extrapolation from clinical EEG studies may not be appropriate. Recent studies on serial EEGs in aircrew are lacking, whereas follow-up of individuals who failed certification is nonexistent. Population-based EEG studies in healthy subjects are generally underpowered to establish the significance of pathological EEG findings. Advanced digital video/EEG recording, in combination with standardization of data exchange formats and automated detection of pathological grapho-elements, is cost effective when carried out for extended periods, e.g., during flight simulator sessions. Extensive databases of serial video/EEG records in aircrew may thus be easily obtained and validated over time. Prognostic inferences on the significance of pathological EEG discharges may subsequently be derived from these databases. PMID- 15652731 TI - The impact of subclinical epileptiform discharges on complex tasks and cognition: relevance for aircrew and air traffic controllers. AB - Subtle seizures consisting of brief alteration of consciousness with or without automatisms may go unnoticed in daily life, but can be detected more easily with electroencephalographic (EEG)/video recordings. Generalized and partial epileptiform EEG discharges can nevertheless be subclinical (subclinical epileptiform discharges, SEDs). When appropriate complex tasks are presented, it has been shown that even very short SEDs of 0.5 second disrupt cognition. In daily life this has been shown during automobile driving: half of the subjects showed significant deviations in lateral position of the car during SEDs and made more errors in an attention task while driving. Individual differences in the cognitive effects of SEDs are, however, striking and may be partly due to interaction between level of performance and frequency of spontaneous EEG discharges, as has been shown in another driving study: about 75% of subjects showed suppression of SEDs by driving, which is a combination of sensory, mental, and motor activity. Not only can SEDs negatively influence performance, but in some cases mental activities can provoke epileptiform discharges. It is important to realize that these mechanisms exist and that only detailed EEG studies can clarify these issues. In air traffic controllers, brief alterations of consciousness and cognitive impairment have occurred but cannot be accepted for safety reasons; therefore, Eurocontrol has used the EEG as a screening tool since 1995. PMID- 15652732 TI - Phenotyping the untouchables: environmental enhancement of behavioral and physiological activation in seizure-prone El mice. AB - The onset and frequency of spontaneous and tail suspension-induced seizures in El mice appear to be influenced strongly by developmental and experiential factors over the first 3 months of life. To assess the impact of social factors on behavioral characteristics of El mice prior to the age of seizure susceptibility, locomotor activity and exploratory measures of arousal were recorded in 40-day old El and control DDY mice exposed to group and isolation housing conditions. Once mice reached maturity, physiological reactivity to a tail suspension stressor was evaluated. The locomotor activity measure revealed circadian entrainment in both strains, nocturnal hyperactivity in El mice, and a locomotor activity-attenuating effect of group housing in El mice. In the two-compartment model of exploration, latency to enter, transitions to and from, and rearing in a brightly lit compartment were 50% higher in El relative to DDY mice, again suggesting a hyperactive phenotype. Finally, an acute 2-minute tail suspension stressor applied to 75-day-old mice implanted with radiotelemetry transmitters revealed a reactive tachycardia in El, but not DDY, mice. No seizures were observed during any of the experimental manipulations. Taken together, these results suggest that spontaneously occurring deviations in behavioral and cardiovascular measures of arousal characterize preseizure El mice and that motor features of hyperarousal can be exaggerated by the environmental manipulation of isolation housing. PMID- 15652733 TI - On the psychopathology of unilateral temporal lobe epilepsy. AB - Personality adjustment of patients with unilateral temporal lobe epilepsy (TLE) was investigated in the light of special characteristics of the epilepsy process, psychosocial stressors, and the cognitive status of the patients. Thirty-seven patients with medically intractable unilateral temporal lobe epilepsy (16-55 years of age; 20 right temporal and 17 left temporal foci) were examined with standardized personality inventories (FPI, STAI, IPC, TSK) supplemented by a rating scale evaluated by the neuropsychologist (GEWLE). Patients with left temporal lobe epilepsy were characterized by increased emotional dependency, less externally judged composedness, increased depressive drive and mood, increased nervousness, increased search for information and exchange of disease experience, and greater tendency to persevere (P < 0.05). Cognitive status and psychosocial status did not significantly differ. The evaluation of personality adjustment contributes to the lateralization of the epileptogenic focus and reveals interesting patterns in the preoperative diagnostic puzzle, and in addition provides a strategy to individualize psychotherapeutic strategies. PMID- 15652734 TI - Written language skills in children with benign childhood epilepsy with centrotemporal spikes. AB - PURPOSE: The goal of this work was to study written language skills in children with benign childhood epilepsy with centrotemporal spikes (BCECTS) in the absence of atypical clinical or electroencephalographic (EEG) features (n = 32), as compared with controls (n = 36). METHODS: BCECTS patients (7-16 years), attending regular school, without cognitive or behavioral regression, or atypical EEG patterns, completed four tests assessing written language skills and one nonverbal cognitive test. School performance information was recorded. Seizure types, duration, and frequency; awake and sleep interictal EEG findings; and medication status were documented. Epilepsy and educational outcome was recorded for a period of 1-5 years after diagnosis. RESULTS: As a group, BCECTS patients performed significantly worse than controls in spelling, reading aloud, and reading comprehension; presented dyslexic-type errors; and frequently had below average school performance. Among 11 with poor written language performance, 4 had developmental dysfunctions before school and seizure onset and, as a group, demonstrated low performance on a nonverbal cognitive test. Even though 65.6% were on anticonvulsants due to frequent seizures and/or seizures while awake, none belonged to the atypical BCECTS spectrum with respect to clinical or EEG findings. Increased epilepsy duration and seizure frequency were less represented in patients with no or mild written language problems than in those with poor performance. Otherwise, clinical course and EEG findings in this group were no different than those for the 11 children with severe written language problems. Clinical follow-up indicated that learning problems appear persistent and several children require remedial classes and/or tutoring after the epilepsy has resolved. CONCLUSIONS: Children with severe but not atypical BCECTS performed, as a group, significantly worse than controls in written language skills, specifically in spelling, reading, aloud and reading comprehension; they also presented many difficulties also found in children with dyslexia. However, poor written language performance was not considered specific to BCECTS because it occurred in children with generally low cognitive capacity and/or preexisting developmental dysfunctions. Moreover, a dissociation between epilepsy outcome and learning problem outcome, in the subgroup with poor performance, casts doubt on the existence of a possible causal link between BCECTS and educational performance. PMID- 15652735 TI - The use of complementary medicines and alternative practitioners in a cohort of patients with epilepsy. AB - Complementary and alternative medicines (CAMs) are increasingly used by patients in the Western world. Some of the most popular herbal remedies are known to act on the cytochrome P450 system, with potential effects on antiepileptic drug (AED) levels. Few studies have explored their use in people with epilepsy. We surveyed 400 patients attending epilepsy clinics in Greater Manchester. Thirty-four percent of our patients had used or were using CAMs; the majority had not told their doctor. Use of CAMs was not predicted by age, sex, seizure frequency, number of AEDs, or dissatisfaction with conventional medicine. Patients who had gone onto higher education were significantly (P < 0.05) more likely to have used or be using CAMs. The majority of patients did not use CAMs for their epilepsy but for general health purposes. Most patients stated that CAMs had little or no effect on seizure frequency or severity. PMID- 15652736 TI - A dosing algorithm for converting from valproate monotherapy to lamotrigine monotherapy in patients with epilepsy. AB - A dosing algorithm was used to guide the conversion of 77 patients with epilepsy (16 years of age) from valproate monotherapy through lamotrigine adjunctive therapy at 200mg daily to lamotrigine monotherapy at 500 mg daily in an open label study comprising a lamotrigine escalation phase (8 weeks), a valproate withdrawal phase (6 weeks), and a lamotrigine monotherapy phase (4 weeks). The algorithm was designed to maintain stable trough concentrations of lamotrigine during valproate withdrawal to minimize seizure risk. Of the 77 patients, 11 prematurely withdrew for administrative reasons (noncompliance, consent withdrawn, loss to follow-up, protocol violation). Of the remaining 66 patients, 18 withdrew because of adverse events, and 48 completed the study. Among the 67 patients with at least one lamotrigine trough serum concentration after initiation of therapy (pharmacokinetic population), mean trough serum concentrations at the end of the lamotrigine escalation phase at a lamotrigine daily dose of 200 mg (7.9 microg/mL, SD = 3.3) did not differ significantly from values during the valproate withdrawal phase (8.7 microg/mL, SD = 3.5) and the lamotrigine monotherapy phase at a lamotrigine dose of 500 mg daily (7.2 microg/mL, SD = 3.3). A similar pattern of results was observed among the 34 patients who completed the study on lamotrigine (completer population). No serious rashes were reported, and the incidence of withdrawals because of decreased seizure control was low (n = 2, or 3%; both incidents were judged to be related to noncompliance). By employment of a four-step dosing algorithm, lamotrigine serum concentrations can be maintained at a stable level with favorable tolerability during a transition from lamotrigine 200mg daily as adjunctive therapy with valproate to lamotrigine monotherapy 500 mg daily. PMID- 15652737 TI - Changes in verbal and nonverbal memory following anterior temporal lobe surgery for refractory seizures: effects of sex and laterality. AB - We studied the effects on verbal and nonverbal memory of anterior temporal lobe (ATL) surgery for epilepsy in 91 patients (46 men, 45 women), all of whom had left-hemisphere dominance for speech. Patients were divided into four groups according to sex and laterality of the excision. The memory tasks were administered shortly before surgery, 6 months postoperatively, and at a 2-year follow-up. Test scores were submitted to repeated-measures analyses of variance. We found that men treated with left temporal resection declined significantly in long-delay verbal memory after surgery, whereas no clear pre- to postoperative sex differences were found with respect to other verbal memory scores. Only the results on long-delay verbal memory confirm previous findings, showing a greater vulnerability of verbal memory to left ATL surgery in men than in women. Women with left temporal excisions obtained particularly poor scores on a long-delay nonverbal memory test preoperatively, but improved their performance on this test significantly after surgery. The seemingly gradual improvement during the 2-year follow-up suggests a plastic process. PMID- 15652738 TI - Which seizure-precipitating factors do patients with epilepsy most frequently report? AB - When treating patients with epilepsy, dealing with seizure-precipitating factors is a partly neglected and underestimated supplement to more traditional therapies. The aim of this study was to investigate the incidence of seizure precipitants in a large epilepsy population and to determine which precipitants patients most often reported. Study participants included twins and their family members ascertained from the Norwegian Twin Panel (NTP), the Danish Twin Registry (DTR), and the Mid-Atlantic Twin Registry (MATR). One thousand six hundred seventy-seven patients with epilepsy were identified and were asked about seizure precipitants using a closed-ended questionnaire. Fifty-three percent reported at least one seizure-precipitating factor, while 30% claimed to have experienced two or more such factors. Emotional stress, sleep deprivation, and tiredness were the three most frequently reported precipitants. Patients with generalized seizures seemed to be more sensitive to sleep deprivation and flickering light than those with partial seizures, while women with partial seizures appeared to be more prone to seizures during menstruation than women with generalized seizures. Knowledge of seizure precipitants has practical implications, not only in patient treatment and counseling, but also for diagnosis, in that it may be helpful in facilitating the appearance of interictal epileptiform discharges in EEG and ictal EEG recordings. PMID- 15652739 TI - Knowledge of women's issues and epilepsy (KOWIE-II): a survey of health care professionals. AB - Attendees of the American College of Physicians 2003 annual meeting were invited to complete a computerized version of the Knowledge of Women's Issues and Epilepsy (KOWIE-II) questionnaire. This 10-item survey includes items specific to issues that affect women with epilepsy (WWE), including hormone sensitive seizures, effects of antiepileptic drugs (AEDs) on oral contraception, bone health, sexual function, pregnancy, and breast-feeding. A total of 202 healthcare providers (HCP) responded to the survey, 92% of which identified themselves as physicians. Few understood the effects of endogenous steroid hormones on seizure threshold (24%) and that epilepsy is associated with an increased incidence of female sexual dysfunction (37%). Most knew that enzyme-inducing AEDs may reduce the efficacy of oral contraceptives (71%) and that certain AEDs are associated with bone disease (77%). The majority were aware that most WWE have healthy children (86%), that women do not need to discontinue AEDs during pregnancy (75%), and that the most appropriate AED during pregnancy is one that best addresses the patient's seizures. Fewer than half (47%) of participants knew that women taking AEDs could breast-feed safely. This sample of HCPs was not adequately informed about the unique issues affecting WWE. An aggressive educational effort is necessary to close the gaps in knowledge. PMID- 15652740 TI - A patient with a 44-year history of epilepsia partialis continua caused by a perirolandic cortical dysplasia. AB - Epilepsia partialis continua (EPC), or Kojevnikov's syndrome, is a rare epileptic syndrome arising from a variety of lesions in the perirolandic area. We report herein a 46-year-old woman with drug-resistant EPC due to a cortical dysplasia in the left frontoparietal region. For 44 years she has suffered continuous right sided jerks, particularly in the right arm and hand, with an average frequency of 10-20 jerks per minute. During EEG recordings her jerks were associated with spikes and sharp waves over the left frontocentroparietal region, sometimes also with bursts of high-voltage generalized spike-wave complexes with a maximum bicentrally, followed by an electrodecrement. Despite the continuous jerks she is independent in daily life activities, and she considers the jerks not severe enough to justify surgery, i.e., multiple subpial transections. PMID- 15652741 TI - Idiopathic generalized tonic-clonic seizures limited to exercise in a young adult. AB - Loss of consciousness limited to exercise should suggest a cardiovascular etiology even when clonic activity is reported. Infrequently, epilepsy patients report a preponderance of their seizures related to exertion. In these patients, seizures also occur independent of exercise. This is a report of a normal young adult woman with episodes of loss of consciousness limited to exercise. During cardiac stress testing with simultaneous electroencephalography, she developed generalized paroxysms of epileptiform discharges preceding a generalized tonic clonic seizure. This case demonstrates the utility of simultaneous cardiac stress testing with electroencephalography in confirming exercise-related seizures and distinguishing epilepsy from syncope. PMID- 15652742 TI - Postictal mania associated with frontal lobe epilepsy. AB - A 38-year-old man, possibly with frontal lobe epilepsy, developed postictal mania. The changes in psychiatric symptoms and laboratory examinations over time were investigated in two episodes of postictal mania, using long-term electroencephalography with closed circuit television (EEG/CCTV) monitoring, magnetoencephalography (MEG), and single-photon emission computed tomography (SPECT) to elucidate the underlying mechanism of postictal mania. According to the clinical symptoms, the postictal manic episodes of this case had four phases: a lucid interval, manic phase, hypomanic phase, and recovery phase. EEG showed forced normalization during the florid manic phase. The serial findings of EEG, MEG, and SPECT during the postictal manic episodes suggest that functional changes in bilateral frontal lobes, especially the right frontal lobe, right temporal lobe, and right paralimbic area, are crucial in the development of postictal mania, and that these functional changes are dynamic. PMID- 15652743 TI - Mechanism-based combination telomerase inhibition therapy. AB - Inhibition of telomerase is an exciting therapeutic target, since it is required for the long-term proliferation of most cancer cells but not present in most somatic cells. However, effective telomerase inhibitors have yet to be tested in clinical trials. In this issue of Cancer Cell, Seimiya and coworkers explore inhibiting tankyrase, an enzyme involved in making telomeres accessible to telomerase. Adding a partial inhibition of tankyrase to a partial inhibition of telomerase drove cancer cells into crisis and death. The combination of tankyrase and telomerase inhibitors may offer new opportunities for realizing the promise of telomerase inhibition therapy. PMID- 15652744 TI - A surrogate marker to monitor angiogenesis at last. AB - The first angiogenesis inhibitor has recently been approved for cancer treatment. Nonetheless, optimizing the dose of novel angiogenesis inhibitors remains a formidable challenge--primarily because we lack reliable surrogate markers of tumor angiogenesis. In this issue, Shaked et al. provide evidence that the levels of circulating endothelial precursor cells (CEPs), which contribute to the formation of tumor vessels, are genetically predetermined and regulated by angiogenic factors, and reflect the antitumor efficacy of angiogenesis inhibitors. These findings highlight the potential usefulness of CEPs as a surrogate marker to monitor and adjust antiangiogenic therapy. PMID- 15652745 TI - Spying on cancer: molecular imaging in vivo with genetically encoded reporters. AB - Genetically encoded imaging reporters introduced into cells and transgenic animals enable noninvasive, longitudinal studies of dynamic biological processes in vivo. The most common reporters include firefly luciferase (bioluminescence imaging), green fluorescence protein (fluorescence imaging), herpes simplex virus 1 thymidine kinase (positron emission tomography), and variants with enhanced spectral and kinetic properties. When cloned into promoter/enhancer sequences or engineered into fusion proteins, imaging reporters allow transcriptional regulation, signal transduction, protein-protein interactions, oncogenic transformation, cell trafficking, and targeted drug action to be spatiotemporally resolved in vivo. Spying on cancer with genetically encoded imaging reporters provides insight into cancer-specific molecular machinery within the context of the whole animal. PMID- 15652746 TI - Context, tissue plasticity, and cancer: are tumor stem cells also regulated by the microenvironment? PMID- 15652747 TI - Tankyrase 1 as a target for telomere-directed molecular cancer therapeutics. AB - Telomere elongation by telomerase is repressed in cis by the telomeric protein TRF1. Tankyrase 1 poly(ADP-ribosyl)ates TRF1 and releases it from telomeres, allowing access of telomerase to telomeres. Here we demonstrate that tankyrase 1 inhibition in human cancer cells enhances telomere shortening by a telomerase inhibitor and hastens cell death. Conversely, either tankyrase 1 upregulation or telomere shortening, each of which decreased TRF1 loading on a chromosome end, attenuated the impact of telomerase inhibition. These results are consistent with the idea that telomeres having fewer TRF1s increase the efficiency of their elongation by telomerase. This study implies that both enzyme activity and accessibility to telomeres can be targets for telomerase inhibition. PMID- 15652748 TI - Ectopic expression of VAV1 reveals an unexpected role in pancreatic cancer tumorigenesis. AB - Herein, we show that the hematopoietic-specific GEF VAV1 is ectopically expressed in primary pancreatic adenocarcinomas due to demethylation of the gene promoter. Interestingly, VAV1-positive tumors had a worse survival rate compared to VAV1 negative tumors. Surprisingly, even in the presence of oncogenic KRAS, VAV1 RNAi abrogates neoplastic cellular proliferation in vitro and in vivo, thus identifying Vav1 as a growth-stimulatory protein in this disease. Vav1 acts synergistically with the EGF receptor to stimulate pancreatic tumor cell proliferation. Mechanistically, the effects of Vav1 require its GEF activity and the activation of Rac1, PAK1, and NF-kappaB and involve cyclin D1 upregulation. Thus, the discovery of prooncogenic pathways regulated by Vav1 makes it an attractive target for therapeutic intervention. PMID- 15652749 TI - p27(Kip1)-stathmin interaction influences sarcoma cell migration and invasion. AB - Emerging evidences suggest that cyclin-dependent kinase inhibitors (CKIs) can regulate cellular functions other than cell cycle progression, such as differentiation and migration. Here, we report that cytoplasmic expression of p27(kip1) affects microtubule (MT) stability following cell adhesion on extracellular matrix (ECM) constituents. This p27(kip1) activity is due to its ability to bind and impair the function of the MT-destabilizing protein stathmin. Accordingly, upregulation of p27(kip1) or downregulation of stathmin expression results in the inhibition of mesenchymal cell motility. Moreover, high stathmin and low cytoplasmic p27(kip1) expression correlate with the metastatic phenotype of human sarcomas in vivo. This study provides a functional link between proliferation and invasion of tumor cells based on diverse activities of p27(kip1) in different subcellular compartments. PMID- 15652751 TI - Succinate links TCA cycle dysfunction to oncogenesis by inhibiting HIF-alpha prolyl hydroxylase. AB - Several mitochondrial proteins are tumor suppressors. These include succinate dehydrogenase (SDH) and fumarate hydratase, both enzymes of the tricarboxylic acid (TCA) cycle. However, to date, the mechanisms by which defects in the TCA cycle contribute to tumor formation have not been elucidated. Here we describe a mitochondrion-to-cytosol signaling pathway that links mitochondrial dysfunction to oncogenic events: succinate, which accumulates as a result of SDH inhibition, inhibits HIF-alpha prolyl hydroxylases in the cytosol, leading to stabilization and activation of HIF-1alpha. These results suggest a mechanistic link between SDH mutations and HIF-1alpha induction, providing an explanation for the highly vascular tumors that develop in the absence of VHL mutations. PMID- 15652750 TI - Role for the epidermal growth factor receptor in neurofibromatosis-related peripheral nerve tumorigenesis. AB - Benign neurofibromas and malignant peripheral nerve sheath tumors are serious complications of neurofibromatosis type 1. The epidermal growth factor receptor is not expressed by normal Schwann cells, yet is overexpressed in subpopulations of Nf1 mutant Schwann cells. We evaluated the role of EGFR in Schwann cell tumorigenesis. Expression of EGFR in transgenic mouse Schwann cells elicited features of neurofibromas: Schwann cell hyperplasia, excess collagen, mast cell accumulation, and progressive dissociation of non-myelin-forming Schwann cells from axons. Mating EGFR transgenic mice to Nf1 hemizygotes did not enhance this phenotype. Genetic reduction of EGFR in Nf1(+/-);p53(+/-) mice that develop sarcomas significantly improved survival. Thus, gain- and loss-of-function experiments support the relevance of EGFR to peripheral nerve tumor formation. PMID- 15652752 TI - Stat5 tetramer formation is associated with leukemogenesis. AB - Activation of Stat5 is frequently found in leukemias. To study the mechanism and role of Stat5 activation, we introduced a constitutively activated Stat5a mutant, cS5F, into murine bone marrow (BM) cells. BM transplantation with cS5F transfected cells caused development of multilineage leukemias in lethally irradiated wild-type or nonirradiated Rag2(-/-) mice. The leukemic cells showed strongly enhanced levels of cS5F tetramers but unchanged cS5F dimer levels in a DNA binding assay. Moreover, Stat5a mutants engineered to form only dimers, but not tetramers, failed to induce leukemias. In addition, Stat5 tetramers were found to accumulate in excess compared to dimers in various human leukemias. These data suggest that Stat5 tetramers are associated with leukemogenesis. PMID- 15652753 TI - Genetic heterogeneity of the vasculogenic phenotype parallels angiogenesis; Implications for cellular surrogate marker analysis of antiangiogenesis. AB - Development of antiangiogenic therapies would be significantly facilitated by quantitative surrogate pharmacodynamic markers. Circulating peripheral blood endothelial cells (CECs) and/or their putative progenitor subset (CEPs) have been proposed but not yet fully validated for this purpose. Herein, we provide such validation by showing a striking correlation between highly genetically heterogeneous bFGF- or VEGF-induced angiogenesis and intrinsic CEC or CEP levels measured by flow cytometry, among eight different inbred mouse strains. Moreover, studies using genetically altered mice showed that levels of these cells are affected by regulators of angiogenesis, including VEGF, Tie-2, and thrombospondin 1. Finally, treatment with a targeted VEGFR-2 antibody caused a dose-dependent reduction in viable CEPs that precisely paralleled its previously and empirically determined antitumor activity. PMID- 15652754 TI - Effect of Linomide on adhesion molecules, TNF-alpha, nitrogen oxide, and cell adhesion. AB - Linomide (quinoline-3-carboxamide) is an immunomodulator with anti-inflammatory effects in rodents with autoimmune diseases. Its mode of action still remains to be elucidated. We hypothesized that an investigation of T cell interactions with the extracellular matrix (ECM), composed of glycoproteins such as fibronectin (FN) and laminin (LN), might provide better understanding of their in vivo mode of action in extravascular inflammatory sites. We examined the effect of Linomide on T cell adhesion to intact ECM, and separately to LN, and FN, and on the release and production of tumor necrosis factor (TNFalpha) and nitrogen oxide (NO) in relation to adhesive molecules in non-obese diabetic (NOD) female spleen cells, focusing on intracellular adhesion molecule-1 (ICAM-1) and CD44. NOD female mice that developed spontaneous autoimmune insulitis, which destroys pancreatic islets and subsequently leads to insulin-deficient diabetes mellitus, were studied. Linomide, given in the drinking water or added to tissue cultures in vitro, inhibited the beta1 integrin-mediated adhesion of T cells to ECM, FN and LN, as well as the production and release of TNFalpha and NO, which play a major role in the induction and propagation of T cell-mediated insulitis. In addition, exposure of T cells to Linomide resulted in increased expression of CD44 and ICAM-1 molecules on spleen cells of Linomide-treated mice; such an increase in adhesion molecule expression may lead to more effective arrest of T cell migration in vivo. The regulation of T-cell adhesion, adhesion receptor expression, and inhibition of TNFalpha and NO secretion by Linomide may explain its beneficial role and provide a new tool for suppressing self-reactive T cell dependent autoimmune diseases. PMID- 15652755 TI - Biological activity of conjugates of muramyl dipeptides with batracylin derivatives. AB - Antitumour activity of batracylin (BAT), muramyldipeptide (MDP) and four immunomodulatory conjugates of BAT with MDP were evaluated in the study. The activity was assessed using viability tests performed in the cultures of tumour cell lines of different tissue origin such as WEHI 164 (fibrosarcoma), K562 (leukaemia), and Ab (melanoma), populations of immune cells isolated from peripheral blood, and the tumour cells mixed with immune cells. An intensity of cell death caused by the analogues was measured using flow cytometry analysis as subG1 peak and the distinction between necrotic and apoptotic DNA cleavage during cell death was performed using DNA fragmentation assay. The compounds 11c, 11e and 11h managed to kill WEHI 164 cells in the presence of immune cells in apoptotic manner while BAT and conjugate 11a caused necrosis at the same time. Necrotic pattern of DNA cleavage was also noted in all cultures containing K562 and Ab cells. BAT and MDP caused necrosis in the cultures of pure immune cells, while the conjugates did not affect these cultures at all. Surprisingly, some analogues increased viability of K562 and Ab cells. Low toxicity and ability to induce apoptosis suggested usefulness of some analogues, mainly 11c, as antitumour drugs in limited range of tumours of certain tissue origin, such as WEHI 164. PMID- 15652756 TI - Effects of fish-derived lipoprotein extracts on activation markers, Fas expression and apoptosis in peripheral blood lymphocytes. AB - Several factors may influence numbers and function of peripheral blood lymphocytes (PBLs) by different processes. We conducted this study to evaluate the effect of E-CAB-94011 and E-JUR-94013, two marine fish extracts from S. scombrus and T. trachurus, respectively, on in vitro PBLs activation and on the expression and functionality of Fas, a cell surface molecule that plays a central role in immune homeostasis and cytotoxic activity. PBLs from 24 healthy volunteers were isolated and flow cytometry was performed to measure the state of activation, Fas expression and apoptosis of PBLs. Functionality of Fas was tested by assessing apoptosis after incubation of isolated lymphocytes with agonistic anti-Fas antibodies in blood samples treated with both E-CAB-94011 and E-JUR 94013. Studies on the lymphocyte cell marker suggest a clear immune activation as measured by the increased levels of CD25, CD8, CD38, CD19 and HLA-DR in vitro expression on lymphocytes treated with both extracts. In addition, a significant reduction in the percentages of apoptotic CD19(+)CD38(+) double positive lymphocytes could be demonstrated in the treated samples with respect to controls (p<0.05). Therefore the present results indicate that both E-CAB-94011 and E-JUR 94013 in vitro are powerful immunoregulatory, increasing immune surveillance. PMID- 15652757 TI - Influence of pyrethroids and piperonyl butoxide on the Ca(2+)-ATPase activity of rat brain synaptosomes and leukocyte membranes. AB - Pyrethroids are widely used insecticides of low acute toxicity in mammals but the consequences of long-term exposure are of concern. Their insecticidal action is related to neurotoxicity and, in addition, there are indications of mammalian immunotoxicity. In order to clarify structure-activity relationships of the membrane interactions of pyrethroids, the present study compared the influence of selected pyrethroids, i.e. permethrin and the more water soluble esbiol (S bioallethrin), both type I, and cyfluthrin, type II, on the Ca(2+)-ATPase activity of rat brain synaptosomes and peritoneal leukocyte membranes. The pyrethroids were tested alone as well as mixed with the enhancing substance piperonyl butoxide (PBO) at concentration ratios of 1:5 and 1:10. At the highest concentration tested, permethrin (10 microM) alone inhibited the ATPase activity of leukocyte membranes by 20%, whereas the synaptosomes were affected less. Esbiol and cyfluthrin alone did not affect either membrane preparation significantly, whereas PBO (50 microM) alone caused 10-15% inhibition. Mixtures of either pyrethroid with PBO inhibited the ATPase activity of both types of membranes (up to 40% inhibition) in a synergistic manner, which always tended to be supra-additive. With esbiol a true potentiation took place. The synergistic interaction between pyrethroid and PBO was most apparent with mixtures of a concentration ratio of 1:5. The ATPase activity of leukocyte membranes tended to be more susceptible to inhibition than that of synaptosomes. The results are in accordance with the assumption that the mammalian toxicity of pyrethroids can be ascribed to a general disturbance of cell membrane function in neuronal tissue. The results indicate that it may also be the case in the immune apparatus. PMID- 15652758 TI - Identification of optimal molecular size of modified Aloe polysaccharides with maximum immunomodulatory activity. AB - Polysaccharides isolated from the gel of Aloe species have been known to have diverse biological activities, including immunomodulatory and antitumor activities. The molecular size-immunomodulatory activity relationship of modified Aloe polysaccharide (MAP) was examined in this study. Crude MAP (G2E1) was prepared from the gel of Aloe vera that was partially digested with cellulase. Proteins in crude MAP were removed by passage through a DEAE-Sephacel column, and then the protein-free MAP (G2E1D) was further separated into three fractions, G2E1DS3 molecular weight (MW > or = 400 KDa), G2E1DS2 (5 KDa < or = MW < or = 400 KDa), G2E1DS1 (MW < or = 5 KDa), by Sephacryl column chromatography and ultrafiltration. Immunomodulatory activities of MAP preparations were examined on a mouse macrophage cell line, RAW 264.7 cells, and in ICR strain of mouse implanted with sarcoma 180 cells. We found that polysaccharides between 400 and 5 KDa exhibit the most potent macrophage-activating activity as determined by increased cytokine production, nitric oxide release, expression of surface molecules, and phagocytic activity. In accordance with the in vitro activity, polysaccharides between 400 and 5 KDa also exhibited the most potent antitumor activity in vivo. PMID- 15652759 TI - Involvement of doxorubicin-induced Fas expression in the antitumor effect of doxorubicin on Lewis lung carcinoma in vivo. AB - Deficiency of Fas expression is one of mechanisms involved in the immune evasion by tumors. Several antitumor drugs, such as doxorubicin (DOX) increase Fas expression in tumor cells and sensitize the cells to Fas-mediated apoptosis in vitro. However, the significance of the Fas expression in vivo is still unclear. Therefore, we examined a role of Fas expression on antitumor effect of DOX using a syngeneic tumor model of Lewis lung carcinoma (3LL) cells in C57BL/6-gld mice that lack functional Fas ligand (FasL). In vitro, anti-Fas agonistic antibody, Jo2, did not decrease a viable cell number of 3LL cells in the absence of DOX, whereas it significantly reduced the cell viability in the presence of DOX. The treatment with DOX alone at the same dose did not induce cell death. Flowcytometric analysis of Fas expression revealed that 3LL cells expressed only a marginal amount of Fas, but the treatment of the cells with DOX increased the expression of Fas in the cell surface. When splenic T cells were prepared from 3LL-bearing C57BL/6 mice, the splenic T cells significantly killed DOX-pretreated 3LL cells more than untreated 3LL cells. In the syngeneic models, DOX inhibited growth of 3LL solid tumor both in wild-type C57BL/6 mice and in Fas-deficient C57BL/6-lpr mice, but it failed in C57BL/6-gld mice, suggesting that the interaction between host FasL and tumor Fas is involved in the antitumor effect of DOX. Furthermore, Fas expression was increased in the solid tumor by the treatment of DOX. These results suggest that the antitumor effect of DOX is partly exerted by the Fas expression and host immune defense. PMID- 15652760 TI - Liposomal methylprednisolone differentially regulates the expression of TNF and IL-10 in human alveolar macrophages. AB - Glucocorticoids (GC) are frequently used for therapy of various inflammatory lung diseases by either systemic or inhalative application. Because the oral application often has various side effects and because the inhalative application is not as potent, new formulations of GCs are required. We evaluated the effect of a liposomal (Lip) formulation of methylprednisolone (MP) on the expression of lipopolysaccharide (LPS)-induced proinflammatory tumor necrosis factor (TNF) and antiinflammatory interleukin-10 (IL-10) in human alveolar macrophages (AM). AM were obtained from bronchoalveolar lavage fluids of patients with various inflammatory lung diseases and precultured 20 h+/-MP, either liposomal or free, and then stimulated with LPS. Cells were harvested for analysis of mRNA levels by real-time reverse transcriptase polymerase chain reaction (RT-PCR); supernatants were used to measure protein concentrations by ELISA. We confirm the suppression of LPS-induced TNF production by an average of factor 7 at the mRNA level and factor 3 at the protein level. On the other hand, we detected a strong increase of the IL-10 production by MP. At the mRNA level, liposomal MP alone led to an 18 fold increase, and the LPS-induced IL-10 mRNA was enhanced by factor 2. At the protein level, MP alone had no effect, but LPS-induced IL-10 was increased by factor 2.5. Our data show that liposomal MP can consistently induce IL-10 and reduce TNF when macrophages are exposed for a prolonged period of time. In all respects, liposomal MP had similar activities as free MP, but liposomes were selectively taken up by monocytes and macrophages and not by lymphocytes in blood and in the lung. This suggests that liposomal glucocorticoids when applied locally in the lung may act efficiently but with less side effects. PMID- 15652761 TI - Rescue of in vivo FAS-induced apoptosis of hepatocytes by corticosteroids either associated with alcohol consumption by mice or provided exogenously. AB - The pathogenic effects of many hepatic viral infections are mediated, at least in part, by the immune response to the infected hepatocyte. The immune response in the infected liver involves the interaction of cytotoxic T cells (CTL) with the hepatocytes through the interaction of FAS-ligand on the CTL and FAS on the hepatocyte. The initial hypothesis for this study was that alcohol consumption by mice would sensitize the liver to apoptosis induced by ligation of FAS. C57Bl/6 mice fed ethanol in a liquid diet did show an increased percentage of apoptotic cells 2 h after injection with anti-FAS as compared with the percentage in the control mice. However, 4 and 6 h after anti-FAS injection, control mice showed high percentages of apoptotic cells (20% to 41%) compared with 5% and 4% apoptotic cells in the ethanol-fed mice. The decreased apoptosis of ethanol-fed mice correlated closely with corticosterone levels in the sera. This was confirmed by the finding that adrenalectomized (ADX) mice provided a high level of corticosterone in drinking water were protected against FAS-induced hepatocyte apoptosis. Ethanol-fed mice showed a significant elevation of serum alanine aminotransferase (ALT) levels indicating the development of hepatitis in spite of the relatively low proportion of apoptotic cells in the liver. In conclusion, high levels of corticosterone protect hepatocytes from FAS-mediated apoptosis, but do not prevent the ultimate development of liver damage. In experiments where mice were provided ethanol chronically in drinking water, where stress is minimal, higher levels of ALT were noted in animals in the ethanol group as compared with animals in the control group. These data support the suggestion that ethanol increases hepatocyte sensitivity to FAS-mediated damage. PMID- 15652763 TI - Search for the target genes involved in the suppression of antibody production by TCDD in C57BL/6 mice. AB - 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) suppresses antibody production through activation of a transcription factor, the aryl hydrocarbon receptor (AhR). To explore the genes that are involved in the suppression of antibody production by TCDD, we investigated TCDD-induced changes in gene expression in the CD4 T cells and B cells of C57BL/6 mice immunized with ovalbumin (OVA) plus alum as an adjuvant. Changes in gene expression were analyzed with Affymetrix oligonucleotide microarrays. The results showed that OVA-immunization alone up regulated expression levels of many genes in the CD4 T cells as early as 3 h after immunization, with 55 up-regulated and 5 down-regulated. At 24 h, 42 genes were found to be up-regulated and 30 down-regulated. Fewer genes were affected in the B cells than in the CD4 T cells. In contrast to the up-regulation of genes induced by immunization in the CD4 T cells, administration of TCDD to mice 3 h prior to the immunization mainly caused down-regulation of genes in the CD4 T cells when compared with immunization alone, with 1 being up-regulated and 4 down regulated at 3 h after immunization and 3 up-regulated and 34 down-regulated at 24 h. In particular, at 3 and 24 h, TCDD suppressed expression of three and seven genes, respectively, that were up-regulated by immunization. Another characteristic of the TCDD-induced changes in gene expression was the suppression of many genes encoding proteins that are involved in GTP-binding protein-linked signaling in CD4 T cells. These results suggest that the inhibition of immunization-induced gene expression and modulation of G-protein-linked signaling in CD4 T cells are responsible for the TCDD-induced suppression of antibody production. PMID- 15652762 TI - T cells recognize PD(N/T)R motif common in a variable number of tandem repeat and degenerate repeat sequences of MUC1. AB - The tumor-associated antigen MUC1 is a transmembrane glycoprotein, which is overexpressed in human carcinomas. Peptide epitopes, containing the PDTR fragment from the variable number of tandem repeat (VNTR) domains of MUC1 have been found to be immunodominant in T-cell and B-cell responses. However, little is known about the immunogenicity and specificity of T-cell epitopes from other regions of MUC1 that may also participate in immune responses against tumors. In this study, the combination of immunoinformatics, molecular modeling and a vaccine adjuvant strategy were used to predict and describe a novel T-cell epitope, SAPDNRPAL, located within the degenerate tandem repeat of MUC1. This peptide possesses structural similarity to both VNTR-derived SAPDTRPAP and Sendai virus peptide FAPGNYPAL, which are known to induce cytotoxic T lymphocytes (CTL). We found that SAPDNRPAL had a higher affinity for mouse H-D(b), H-2K(b) and human HLA-A2 molecules than SAPDTRPAP. A chimeric peptide (CP) containing SAPDNRPAL and an adjuvant C5a-derived decapeptide induced epitope-specific type 1 T cells in human MUC1 transgenic mice (ELISPOT). Mice that received dendritic cells (DC) pulsed with the CP or a 25-mer peptide containing the SAPDNRPAL sequence showed increased frequencies of SAPDNRPAL- and SAPDTRPAP-specific interferon-gamma producing T cells. PDTR-specific antibody 214D4 reacted with both SAPDNRPAL and SAPDTRPAP (ELISA). Altogether, our data suggest that the degenerate MUC1 repeat sequence contains the immunogenic T-cell epitope SAPDNRPAL, which is cross reactive with the VNTR-derived peptide SAPDTRPAP. We suggest that the use of immunogenic PDNR-containing epitope(s) in vaccine strategies could be beneficial for developing increased, PD(N/T)R motif-specific T-cell responses against tumors expressing MUC1. PMID- 15652765 TI - Effects of Brazilian and Bulgarian propolis on bactericidal activity of macrophages against Salmonella Typhimurium. AB - Propolis has been used in folk medicine since ancient times due to its many biological properties, such as antimicrobial, antiinflammatory, antioxidant, immunomodulatory activities, among others. Macrophages play an important role in the early phase of Salmonella infection. In this work, macrophages were prestimulated with Brazilian or Bulgarian propolis and subsequently challenged with Salmonella Typhimurium at different macrophage/bacteria ratio. After 60 min of incubation, cells were harvested with Triton-X to lyse the macrophages. To assess the bactericidal activity, the number of colony-forming units (CFU) of S. typhimurium was determined by plating 0.1 mL in Mueller Hinton agar. After 24 h, CFU were counted, and the percentage of bactericidal activity was obtained. Propolis from Brazil and Bulgaria enhanced the bactericidal activity of macrophages, depending on its concentration. Brazilian propolis seemed to be more efficient than that from Bulgaria, because of their different chemical composition. In Bulgaria, bees collect the material mainly from the bud exudate of poplar trees, while in Brazil, Baccharis dracunculifolia DC. was shown to be the main propolis source. Our data also showed that the increased bactericidal activity of macrophages involved the participation of oxygen (H(2)O(2)) and nitrogen (NO) intermediate metabolites. PMID- 15652764 TI - Treatment with Flt3 ligand plasmid reverses allergic airway inflammation in ovalbumin-sensitized and -challenged mice. AB - We have previously reported that fms-like tyrosine kinase 3 ligand (Flt3-L) prevents and reverses established allergic airway inflammation in an ovalbumin (OVA) induced mouse model of asthma. In this study, we investigated the effect of pUMVC3-hFLex, a plasmid, mammalian expression vector for the secretion of Flt3-L on the same mouse model as well as the duration of the effect of the treatment. Allergic airway inflammation to OVA was established in BALB/c mice. OVA sensitized mice received three intramuscular (i.m.) injections of 200 mug pUMVC3 hFLex over 10 days. The response to pUMVC3-hFLex therapy was assessed based on airway hyperresponsiveness (AHR) to methacholine and inflammation, measured as serum cytokine and immunoglobulins (Ig) levels, and the total and differential cells in bronchoalveolar lavage fluid (BALF). pUMVC3-hFLex treatment completely reversed established AHR (P<0.01) and this effect lasted for at least 24 days after the last treatment injection (P<0.001). pUMVC3-hFLex treatment significantly increased BALF interferon-gamma (IFN-gamma) (P<0.01), serum interleukin (IL)-10 (P<0.01) and anti-OVA IgG2a levels (P<0.01). In contrast, serum IL-4 and IgE levels were significantly reduced (P<0.05). Total BALF cellularity, eosinophiles counts and BALF IL-5 levels were also reduced (P<0.01). pUMVC3-hFLex treatment can reverse established experimental asthma and might provide a novel approach for treating asthma. PMID- 15652766 TI - COX expression and PGE(2) and PGD(2) production in experimental acute and chronic gastric lesions. AB - Prostaglandin E(2) and D(2) (PGE(2) and PGD(2)) production and cyclooxygenase-2 (COX-2) expression during the resolution of acute and chronic gastric inflammatory lesions in Wistar rats have been investigated. Differences between ibuprofen, nonselective COX inhibitor, and rofecoxib, specific COX-2 inhibitor, on the development of the induced responses were also analysed. In an acute model, by instillation of HCL, the greatest injury was observed early with a rapid and progressive restoration. Maximal up-regulation of COX-2 protein was detected at 6 h and was accompanied by increase of PGE(2) synthesis but not PGD(2). Both drugs stimulated COX-2 expression in accordance to their capacity of inhibiting this enzymatic activity, driving to delay in the healing. In a chronic model, by acetic acid-induced gastric ulcers, COX-2 was expressed at 7 days and was also associated with PGE(2) increase. Ibuprofen and rofecoxib also augmented COX-2 protein and inhibited PGE(2) levels. However, PGD(2) production was augmented when none signal of COX-2 protein could be detected. Together, this study confirms the role played by COX-2 enzyme in the resolution of acute and chronic gastric inflammatory process, PGE(2) being the principal product. The antiinflammatory effect of nonsteroidal antiinflammatory drugs (NSAIDs) could be mediated not only through the inhibition of COX activity but also through the induction of antiinflammatory PGs production-such as PGD(2)-although further studies would be needed to clarify the mechanisms of this activity and the possible implicated processes. PMID- 15652767 TI - Dermatan disulfate (Intimatan) prevents complement-mediated myocardial injury in the human-plasma-perfused rabbit heart. AB - Intimatan (dermatan 4,6-O-disulfate), a heparin cofactor II agonist, is a highly sulfated negatively charged semisynthetic polysaccharide. The present study examined the hypothesis that Intimatan reduces complement-mediated myocardial injury. The rabbit isolated heart was perfused with 4% normal human plasma (NHP) as a source of complement in the absence or presence of Intimatan (5 microM). Heat-inactivated human plasma (HIHP) was used as a negative control. Previous studies demonstrated that contact of rabbit tissue with human plasma results in activation of the alternative pathway of the human complement system, leading to irreversible myocardial injury. In the presence of NHP, left ventricular end diastolic pressure (LVEDP) was increased significantly to 61.8+/-11.7 mm Hg compared to a value of 17.2+/-6.1 mm Hg in hearts perfused in the presence of HIHP. Left ventricular developed pressure (LVDP) was reduced significantly upon exposure to NHP, 19.3+/-10.2 (NHP) vs. 54.0+/-8.0 mm Hg (HIHP). Functional impairment in the presence of NHP was accompanied by a significant release of cardiac troponin I (cTnI; 131.8+/-20.3 ng/ml) as compared to hearts exposed to HIHP (0.8+/-0.8). Intimatan treatment improved cardiac function and maintained viability of cardiac myocytes (LVEDP 14.6+/-5.6, LVDP 58.0+/-8.1 mm Hg and cTnI 6.7+/-5.2 ng/ml). Immunohistochemical staining demonstrated that Intimatan pretreatment prevented deposition of the human membrane attack complex (MAC) in hearts exposed to NHP. The results indicate that Intimatan, a glycosaminoglycan (GAG), can reduce tissue injury and preserve organ function that otherwise would be compromised during activation of the human complement cascade. PMID- 15652768 TI - Effect of cis-resveratrol on genes involved in nuclear factor kappa B signaling. AB - This study investigated for the first time the effects of the cis isomer of RESV (c-RESV), a polyphenol present in red wine, on an array of genes whose expression is controlled by nuclear factor kappa B (NF-kappaB) and whose transcriptional activation is critical in a number of pathologies (including some cardiovascular diseases). In inflammatory peritoneal macrophages stimulated with lipopolysaccharide (LPS) and gamma interferon (IFN-gamma), c-RESV significantly blocked the expression of genes related to the REL/NF-kappaB/IkappaB family, adhesion molecules and acute-phase proteins; however, the greatest modulatory effect was obtained on the expression of genes related to the pro-inflammatory cytokines. c-RESV down-regulated the nuclear factor of kappa light chain gene enhancer in B-cells 1 (NFkappaBL1) gene product p105 and up-regulated the nuclear factor of kappa light chain gene enhancer in B-cells inhibitor alpha (IkappaBalpha) gene. c-RESV also significantly inhibited intercellular adhesion molecule-1 (ICAM-1) gene expression and the transmembrane receptors RIP (receptor TNFRSF) and TLR3 (toll-like receptor 7). At 100 muM, c-RESV significantly inhibited transcription of Scya2 (chemokine MCP-1), the chemokine RANTES (regulated on activation, normal T cell expressed and secreted), pro-inflammatory cytokines that attract monocyte-granulocyte cells such as M-CSF (colony stimulating factor 1), GM-CSF (colony-stimulating factor 2) and G-CSF (colony stimulating factor 3), the cytokine tumor growth factor beta (TGF-beta) and the extracellular ligand IL-1alpha. In contrast, c-RESV stimulated transcription of the pro-inflammatory cytokines IL-6 and tumor necrosis factor alpha (TNF-alpha), the extracellular ligand IL-1beta, and the IFN regulatory factor (IRF)-1. In conclusion, c-RESV has a significant modulatory effect on the NF-kappaB signaling pathway and, consequently, an important antioxidant role that may partially explain the cardioprotective effects attributed to long-term moderate red wine consumption. PMID- 15652769 TI - Aqueous extract from aerial parts of Artemisia vestita, a traditional Tibetan medicine, reduces contact sensitivity in mice by down-regulating the activation, adhesion and metalloproteinase production of T lymphocytes. AB - In the present paper, the effect of the aqueous extract from aerial parts of Artemisia vestita (AV-ext), a traditional Tibetan medicine, on ear contact sensitivity was examined. AV-ext significantly reduced the ear swelling when administered during the induction phase of picryl-chloride (PCl)-induced ear contact sensitivity in mice. The extract also showed a dose-dependent inhibition on lymphocyte proliferation and IL-2 production in Con A-activated spleen cells. The proliferation inhibition was confirmed in the mixed lymphocytes reaction. Furthermore, the adhesion of the isolated spleen cells from PCl-sensitized mice to type IV collagen was significantly decreased in a dose-dependent manner by AV ext. Such decrease was also seen in AV-ext-treated Jurkat T cells and the T cells purified from above spleen cells. The purified spleen T cells from PCl-sensitized mice produced more matrix metalloproteinase-9 (MMP-9) than naive T cells, and AV ext remarkably reduced MMP-9 production both in vivo and in vitro. These results suggest that AV-ext may alleviate contact sensitivity through blocking the activation of T lymphocytes and decreasing their localization to the inflammatory sites via down-regulating the potential of cell adhesion and metalloproteinase production. PMID- 15652770 TI - Antitumor effects of recombinant human prolactin in human adenocarcinoma-bearing SCID mice with human NK cell xenograft. AB - To survey the immune regulatory function of recombinant human prolactin (rhPRL) and its potential application in adoptive immunotherapy, CB17-SCID mice were loaded with human colon adenocarcinoma HT-29 cells (5 x 10(5) cells/mouse, i.p.) 24 h before adoptive transfer with the purified human NK cells followed by rhPRL injection (10 mug/mouse, every other day for a total of 10 injections). Upon analysis, rhPRL did not exert any direct inhibitory effects on HT-29 cells but slightly improved the tumor cell growth both in vitro and in vivo. After SCID mice were reconstituted with human NK cells, rhPRL improved the antitumor effects of human NK cells in HT-29-bearing SCID mice, showing a prolonged survival from 70.4 to 112.1 days, and the increased survival rate from all died to 40% survival for more than 160 days. rhPRL improved the proliferation of human NK cells with or without PHA stimulation. rhPRL also directly enhanced the cytotoxicity of human NK cells against HT-29 tumor cells in 4-h coculture. The supernatant of rhPRL-stimulating NK cells inhibited the proliferation of HT-29 cells through, at least partly, the interferon-gamma (IFN-gamma) and tumor necrosis factor-alpha (TNF-alpha) in the supernatant. Thus, rhPRL administration in HT-29 tumor-bearing SCID mice promotes the antitumor effects of adoptively transferred NK cells. PMID- 15652772 TI - Autoantibody-associated congenital heart block: TGFbeta and the road to scar. AB - Few diseases exemplify the integration of research from bench to bedside as well as neonatal lupus (NL), often referred to as a model of passively acquired autoimmunity. The signature histologic lesion of autoimmune congenital heart block (CHB) is fibrosis of the conducting tissue and, in some cases, the surrounding myocardium. It is astounding how rapid, and in most cases, irreversible, the fibrotic response to injury is. The mechanism by which maternal anti-SSA/Ro-SSB/La antibodies initiate and finally eventuate in atrioventricular (AV) nodal scarring is not yet defined. In vitro and in vivo studies suggest that one pathologic cascade leading to scarring may be initiated via apoptosis, resulting in the translocation of SSA/Ro-SSB/La antigens and surface binding by maternal autoantibodies. Subsequently, the Fc portion of the bound immunoglobulin engages Fcgamma receptors on tissue macrophages, resulting in the release of TGFbeta at a threshold that favors a pro-fibrotic milieu and irreversible scarring. This cascade also involves a tissue-specific activation of TGFbeta, which promotes the transdifferentiation of fibroblasts into myofibroblasts, a scarring phenotype. Phagocytosis of opsonized apoptotic cardiocytes is distinct from macrophage pathways engaged in physiologic clearance of dying tissue, which also results in the release of TGFbeta but in the latter case appropriately serves to dampen inflammation. Downregulation of TGFbeta (activation/secretion pathway) may provide the basis of a novel approach to treatment of CHB in the future. PMID- 15652771 TI - Efficacy of a traditional Korean medicine, Chung-Sang-Bo-Ha-Tang, in a murine model of chronic asthma. AB - Traditional herbal medicines may be viable alternatives to corticosteroid therapy for treatment of asthma. However, the therapeutic mechanisms of herbal compounds remain a matter of considerable debate. This study was performed to evaluate the effects of Chung-Sang-Bo-Ha-Tang (CSBHT), a herbal compound administrated therapeutically to asthma patients for centuries, on airway inflammation and remodeling in a murine model of chronic asthma. BALB/c mice sensitized to ovalbumin (OVA) were chronically challenged with aerosolized OVA for 6 weeks. During the last 2 weeks, some mice were treated daily with CSBHT by intragastric feeding. Dexamethasone (Dex)-treated, phosphate-buffered saline (PBS)-treated, and naive mice served as controls. The effects of CSBHT on airway inflammation, lung pathology, and cytokine production were evaluated. Mice exposed to recurrent airway challenge with OVA had chronic inflammation and characteristics of airway remodeling, including subepithelial fibrosis, epithelial hypertrophy, and goblet cell hyperplasia. CSBHT was as effective as Dex at moderately reducing these changes compared to the PBS-treated mice. In addition, IL-5 and IFN-gamma levels in supernatants of Concanavalin A (Con A)-activated splenocyte cultures were reduced in mice treated with CSBHT. Treatment with CSBHT during the last 2 weeks of challenge modulated airway inflammation and remodeling in a murine model of chronic asthma. Thus, CSBHT may effectively delay the progression of airway inflammation and remodeling. PMID- 15652773 TI - Hepatitis C virus, Sjogren's syndrome and B-cell lymphoma: linking infection, autoimmunity and cancer. AB - An increased prevalence of hematologic malignancies is often described in patients with Sjogren's syndrome (SS). Viruses have been proposed as possible etiologic or triggering agents of systemic autoimmune diseases (SADs), with hepatitis C virus (HCV) being one of the viruses most frequently associated with autoimmune features and with systemic autoimmune diseases such as mixed cryoglobulinemia or SS. Moreover, the association between HCV infection and hematologic malignancies, mainly non-Hodgkin's lymphoma (NHL), is supported by several studies. For these reasons, the recognized association of specific systemic autoimmune diseases (mainly SS and mixed cryoglobulinemia) with HCV infection, added to the possible evolution of any one of these entities into a B cell NHL, suggests the possibility of a close relationship among SS, HCV and B cell lymphoproliferative disorders, especially in patients with type II mixed cryoglobulinemia. PMID- 15652775 TI - Mast cells and autoimmunity. AB - Mast cells (MCs) are major effector cells in allergic diseases. Recently, it has become evident that the contribution of MCs extends far beyond their accepted role in allergic disease, and that they play a more extensive role in a variety of non-allergic immune processes such as the innate immunity response. These cells have a key role in both the induction and elicitation of several autoimmune conditions. Targeting MC development, maturation or activation may be of value in future prevention and treatment of autoimmune conditions. PMID- 15652774 TI - The immunogenetics of pemphigus vulgaris. AB - Pemphigus vulgaris (PV) is an autoimmune blistering disease of the skin of unknown etiology. While various environmental factors have been implicated as triggering agents, HLA association is probably the most important predisposing factor. The aim of this review is to highlight the association of HLA with pemphigus vulgaris. In addition, we present recent results showing a possible association with the nonclassical HLA-G antigen. PMID- 15652776 TI - TNF-alpha inhibitors in systemic vasculitides and connective tissue diseases. AB - The introduction of TNF-alpha inhibitors in the treatment of rheumatoid arthritis and several other diseases meant a major progress in the management and to the understanding of these chronic inflammatory diseases. In this article, the evidence of the role of TNF-alpha and for TNF-alpha inhibitors in systemic vasculitides and connective tissue diseases is reviewed. TNF-alpha is expressed in inflammatory lesions. TNF-alpha acts as a proinflammatory cytokine in most disease processes analyzed so far, but it might have anti-inflammatory properties under certain conditions as well, e.g. with respect to B-cell regulation in systemic lupus erythematosus. It is not clear to what extent such aspects will be important in the treatment of connective tissue diseases and systemic vasculitides with TNF-alpha inhibitors. So far, most case reports and case series have suggested favourable results with TNF-alpha inhibitor therapy in systemic lupus erythematosus, dermato- and polymyositis, giant cell arteritis, Churg Strauss syndrome, Wegener's granulomatosis and microscopic polyangiitis. Results of randomized, placebo-controlled trials are awaited for several connective tissue diseases and systemic vasculitides. One randomized, placebo-controlled trial has found no efficacy of infliximab treatment in primary Sjogren's syndrome recently. PMID- 15652777 TI - Unique and shared features of Golgi complex autoantigens. AB - Here we summarize recent advances in the characterization of autoimmune antigens associated with the Golgi complex. All Golgi autoantigens identified to date are high molecular weight proteins rich in coiled-coil domains and localized to the cytoplasmic face of the Golgi cisternae. The characteristic features of these Golgi autoantigens are interestingly similar to selected human autoantigens reported in other intracellular compartments such as endosome, centrosome, and centromere. The implication of this class of autoantigens in autoimmunity is discussed. PMID- 15652778 TI - Complex genetics of Wegener granulomatosis. AB - Wegener granulomatosis (WG) belongs to a heterogeneous group of systemic anti neutrophil cytoplasmatic antibody (ANCA) associated vasculitides (AASV). WG is characterized by necrotizing granulomatous inflammation of the upper and lower respiratory tract, glomerulonephritis and vasculitis. As a multifactorial model disease, WG is hallmarked by the presence of specific ANCA-subtypes directed against a defined antigen. WG is more predominant among Caucasians and the genetic predisposition appears quite complex. Here, we provide a brief overview concerning genetic factors in the pathogenesis of WG and discuss intricacies of molecular genetic approaches. PMID- 15652780 TI - Anti-annexin V antibodies: are they prothrombotic? AB - Annexin V inhibits prothrombin activation and is able to prevent thrombus formation under normal venous and arterial blood flow conditions. Antibodies to annexin V have been identified in association with several pathological conditions, including systemic lupus erythematosus (SLE) with or without anti phospholipid syndrome, recurrent spontaneous abortions and systemic sclerosis (SSc). These antibodies are suspected to exert a detrimental role and interfere with annexin V function. Thus, they have been associated with the occurrence of foetal loss and venous and/or arterial thrombosis in SLE patients, as well as digital ischemia in SSc patients. However, their true pathogenic role remains to be proven. PMID- 15652779 TI - The catastrophic antiphospholipid (Asherson's) syndrome in 2004--a review. AB - An unusual variant of the antiphospholipid syndrome (APS) termed the Catastrophic Antiphospholipid Syndrome (CAPS) in 1992 by Asherson is described. The condition may arise "de-novo" in a patient previously not suspected of having an APS or during the course of a "Primary" APS or Secondary APS (most commonly SLE). The patient may already be on therapy. "Trigger" factors (infections most commonly) have been identified in 45% of patients but in the majority, they remain unidentified. Clinically, the patients present with small vessel occlusions involving organs (e.g. bowel, brain, heart, kidney) but large vessels occlusions do occur. Unusual organs are involved and the clinical features depend on which organs are affected. Because of tissue necrosis, the Systemic Inflammatory Response ensues ("SIRS") and many patients develop ARDS. Despite seemingly adequate therapy (parenteral heparin, steroids, antibiotics), the mortality remains high (approximately 50%). PMID- 15652781 TI - Activating auto-antibodies against the AT1 receptor in preeclampsia. AB - Immune mechanisms have been shown to be important in the pathogenesis of preeclampsia. Here, we review our studies of agonistic antibodies against the AT1 receptor in the pathogenesis as well as a pathologic phenotype of this disorder, focusing on observations in our laboratory. We have demonstrated their specificity of the binding by Western blotting, co-localization, and co immunoprecipitation experiments. AT1-AA induce signaling in vascular cells and trophoblasts including AP-1 and nuclear factor-kappaB (NF-kappaB) activation. The signaling results in tissue factor production and reactive oxygen species (ROS) generation, both of which have been implicated in preeclampsia. The role of AT1 AA in preeclampsia and other severe hypertensive conditions has not yet been proven with certainty. However, we believe the findings are compelling and warrant further study. PMID- 15652782 TI - 1st International Conference on Cutaneous Lupus Erythematosus Dusseldorf, Germany, September 1-5, 2004. PMID- 15652783 TI - Carotid chemoafferent plasticity in adult rats following developmental hyperoxia. AB - Developmental hyperoxia impairs carotid chemoreceptor development and induces long-lasting reduction in carotid sinus nerve (CSN) responses to hypoxia in adult rats. Studies were carried out to determine if CSN responses to acute hypoxia would exhibit hypoxia-induced plasticity in adult 3-5-months-old rats previously treated with postnatal hyperoxia (60% O2, PNH) of 1, 2, or 4 weeks duration. CSN responses to acute hypoxia were assessed in adult rats exposed to 1 week of sustained hypoxia (12% O2, SH). In normal adult rats and adult rats treated with 1 week of PNH, CSN responses to acute hypoxia were significantly increased in urethane-anesthetized rats when studied 3-5 h after SH. Apparent increases in CSN responses to hypoxia were not significant in rats treated with 2 weeks of PNH and were clearly absent after 4 weeks of PNH, but exponential analysis suggests a PNH duration-dependent plasticity of the CSN response to acute hypoxia after SH. In a second study rats exposed to 2 weeks of PNH were treated with SH for 1 week as adults and acute hypoxic responses were tested 4-5 months later. CSN responses in these rats were unaffected by SH suggesting a lack of persistent SH-induced functional plasticity. We conclude that rats treated with 1 week of PNH retain the capacity for hypoxia-induced plasticity of carotid chemoafferent function and some potential for plasticity may be present after 2 weeks of PNH, whereas 4 weeks of PNH impairs the capability of rats to exhibit plasticity following 1 week of SH. PMID- 15652784 TI - D1-dopamine receptor blockade slows respiratory rhythm and enhances opioid mediated depression. AB - Previous studies indicate that dopamine modulates the excitability of the respiratory network and its susceptibility to depression by exogenous opioids, but the roles of different subtypes of dopamine receptor in these processes are still uncertain. In this study, D1-dopamine receptor (D1R) involvement in dopaminergic modulation of respiratory rhythm and mu-opioid receptor mediated depression were investigated in pentobarbital-anesthetized cats. Intravenous administration of the D1R blocker SCH-23390 (100-200 microg/kg) slowed phrenic nerve and expiratory neuron respiratory rhythms by prolonging the inspiratory and expiratory phases. Phrenic nerve discharge intensity also increased more gradually during the inspiratory phase. SCH-23390 (150 microg/kg) also enhanced dose-dependent depression of phrenic nerve and expiratory neuron excitability, as well as rhythm disturbances, produced by the mu-opioid receptor agonist fentanyl (2-20 microg/kg, i.v.). The results suggest an important role for the D1-subtype of receptor in respiratory rhythm modulation, and indicate that this type of receptor participates in dopaminergic compensatory mechanisms directed against opioid-mediated network depression. PMID- 15652785 TI - Impaired ventilatory acclimatization to hypoxia in mice lacking the immediate early gene fos B. AB - Earlier studies on cell culture models suggested that immediate early genes (IEGs) play an important role in cellular adaptations to hypoxia. Whether IEGs are also necessary for hypoxic adaptations in intact animals is not known. In the present study we examined the potential importance of fos B, an IEG in ventilatory acclimatization to hypoxia. Experiments were performed on wild type and mutant mice lacking the fos B gene. Ventilation was monitored by whole body plethysmography in awake animals. Baseline ventilation under normoxia, and ventilatory response to acute hypoxia and hypercapnia were comparable between wild type and mutant mice. Hypobaric hypoxia (0.4 atm; 3 days) resulted in a significant elevation of baseline ventilation in wild type but not in mutant mice. Wild type mice exposed to hypobaric hypoxia manifested an enhanced hypoxic ventilatory response compared to pre-hypobaric hypoxia. In contrast, hypobaric hypoxia had no effect on the hypoxic ventilatory response in mutant mice. Hypercapnic ventilatory responses, however, were unaffected by hypobaric hypoxia in both groups of mice. These results suggest that the fos B, an immediate early gene, plays an important role in ventilatory acclimatization to hypoxia in mice. PMID- 15652786 TI - Repeated measurement of hypoxic ventilatory response as an intermittent hypoxic stimulus. AB - Measurement of hypoxic ventilatory response (HVR) involves an exposure to hypoxia which, if repeated over several days might act as an intermittent hypoxic stimulus. The purpose of this study was to measure HVR repeatedly over 5 days to determine whether it was affected by repeated measurement. Nine healthy male subjects completed an isocapnic HVR test, on one occasion, followed 5 days later by one measurement each day for 5 days. Each test lasted approximately 5-8 min with inspired oxygen concentration declining to as a low as 5-6%. No systematic trend was observed in HVR over the 5-day period (p>0.05). There were no significant differences in HVR between any of the test days. Regression failed to show any trend in HVR over the five sequential days. The calculated mean coefficient of variation for HVR for each subject was 27%. There is no evidence that the short exposure to hypoxia as part of HVR measurement is a co intervention when measured repeatedly over 5 days in physiological studies. PMID- 15652787 TI - Changes in respiratory control after 5 days at altitude. AB - These experiments examined changes in the chemoreflex control of breathing and acid-base balance after 5 days at altitude (3480 m) in six healthy males. The partial pressures of carbon dioxide (P(CO2)) at which ventilation increased during isoxic hypoxic and hyperoxic modified rebreathing tests at sea level fell significantly at altitude by mean+/-S.E.M. of 12.8+/-2.51 mmHg and 9.5+/-1.77 mmHg, respectively, but response slopes above threshold were unchanged. Altitude exposure produced a respiratory alkalosis evidenced by a decrease in mean resting end-tidal P(CO2) from 41+/-0.84 mmHg at sea level to 32+/-2.04 mmHg at altitude, but pH did not increase significantly from its sea level value. Blood samples were analyzed to discover acid-base changes, using a modification of the equations for acid-base balance proposed by [Stewart, P.A., 1983. Modern quantitative acid-base chemistry. Can. J. Physiol. Pharmacol. 61, 1444-1461]. While strong ion difference at altitude was not significantly different from its sea level value, albumin concentration was increased significantly from 38.6+/ 0.30 g L(-1) to 49.8+/-0.76 g L(-1). We suggest that the respiratory alkalosis was produced by a fall in the chemoreflex threshold and pH was corrected by an elevation in the concentration of weakly dissociated protein anions. PMID- 15652788 TI - On the interaction between respiratory compartments during passive expiration in ARDS patients. AB - Relaxed expiratory volume-time profile has been frequently analysed by fitting exponential functions of time to one- or two-compartment models. In the latter case, the two exponential constants are assumed as representing the time constants of both compartments. Least-square fittings on the experimental data of five consecutive mechanically ventilated supine patients with acute respiratory distress syndrome (ARDS) were performed using rate-constants (flow/volume ratio) as parameters in order to obtain the model matching. Passive expiratory volume time curves were recorded under PEEP = 0 and 13.6 +/- 3.3 S.D. cmH2O conditions. Model matching was optimal with significant, reliable parameter values. As a result, the use of a PEEP in ARDS patients: (a) delayed expiration; (b) decreased the percentage initial volume contribution of the slow-emptying compartment; and, (c) modified the interaction between compartments. The volume-time profile of the second compartment was found to increase at the beginning of expiration, and, then, progressively decayed towards zero, showing a maximum, although the overall curve decreased throughout expiration. PMID- 15652789 TI - Effect of antitubercular drugs on dipalmitoylphosphatidylcholine monolayers: implications for drug loaded surfactants. AB - The year long chemotherapy in pulmonary tuberculosis results in dose related side effects and may not reach atelectatic areas. On account of its spreading properties and the ability to re-expand atelectatic areas, exogenous surfactant may act as a pulmonary drug delivery agent. We investigated the interactions between antitubercular drugs and the main surfactant component, dipalmitoylphosphatidylcholine (DPPC) with the aim of developing more effective antitubercular drug loaded surfactants. The surface properties were evaluated using a Langmuir-Blodgett trough and Wilhelmy balance at 37 degrees C. Lung surfactant was modeled as DPPC monolayers. The isoniazid (INH)-DPPC combination in 1:1 ratio by weight significantly improved the adsorption of DPPC, reached a minimum surface tension of zero, formed a low compressibility film and required 32.7% area change to decrease surface tension from 30 to 10 mN/m. The triple drug (INH-rifampicin-ethambutol in 1:2:3 ratio by weight) DPPC combination when used in 1:1 or 1:2 ratios by weight also achieved surface properties superior to those of DPPC alone. A significant improvement in the adsorption was observed (surface tensions of 34.7 mN/m for 1:1 and 32.0 mN/m for 1:2 triple drug: DPPC combinations in the first second), and the films had low compressibility reaching a minimum surface tension of zero on compression. Thus, we observed statistically significant improvements in all the surface parameters and we feel encouraged to continue developing a tuberculosis therapy consisting of surfactant liposomes carrying antitubercular drugs. PMID- 15652790 TI - How does lobeline injected intravenously produce a cough? AB - In order to examine, whether the lobeline-induced cough is a true reflex or a voluntary effort to get rid of its irritating sensations in the upper respiratory tract, we systematically studied the cough response to lobeline, of subjects who were unable to make conscious discriminations i.e. were either comatose (n=4) or anaesthetized (n=5). 8 microg/kg lobeline injected into the right atrium of one and 29 microg/kg intravenously (i.v.) into another evenly and spontaneously breathing comatose subject produced a cough after 4s and 12s, respectively. Cough was repeatable and showed a dose response relationship i.e., its latency decreasing and its duration/intensity increasing with the dose. In a third subject, capable only of weak spontaneous respiration, a relatively high dose injected into the right atrium (44 microg/kg) generated a pronounced cough-like respiratory movement superimposed on the artificial ventilation and also during the apnoea after disconnecting the pump. No respiratory response was evoked in a fourth subject who had no evidence of brainstem reflexes. In five normals, cough was elicited with a mean dose of 35+/-5 microg/kg i.v. (latency 14+/-2 s; duration 10+/-3 s). After thiopental anaesthesia, injecting 41+/-7 microg/kg produced a cough within 13+/-2 s that lasted for 12+/-2 s. It may be noted that neither the later dose nor the latency or duration of cough that it produced were significantly different from the pre anaesthesia values (P>0.05). These two sets of results show unequivocally that the lobeline-induced cough is evoked reflexly; its magnitude in the conscious state could vary by subjective influences. We discuss the likelihood of its origin from juxtapulmonary capillary receptors. PMID- 15652792 TI - Sputum induction as a research tool for the study of human respiratory mucus. AB - The study objectives were to compare in vitro transportability and physical properties of respiratory mucus, obtained invasively by direct collection (DC) right after endotracheal intubation and non-invasively by sputum induction with 3% hypertonic saline solution inhalation (SI) 24 h before the anesthesia. Twenty two patients with no pulmonary disease scheduled for elective abdominal surgical procedures were studied. The parameters analyzed and the main results are as follows. (1) Transportability by cilia (MCT), SI was higher than DC (0.94+/-0.25 and 0.62+/-0.25; P<0.001). There was a significant correlation between the two methods and DC could be estimated by: DC=0.21+(0.44 SI) (r=0.44; P<0.001). (2) Transportability by cough (CC), SI was higher than DC (68.23+/-32.1 and 33.58+/ 19.04 mm; P=0.002). (3) Contact angle (CA), SI was lower than DC (10+/-3 degrees and 22+/-14 degrees ; P=0.025). (4) Rheological properties (no significant difference obtained between SI and DC). These results indicated that SI changes mucus physical properties and transportability in non-expectorators. PMID- 15652791 TI - Dielectric behavior of pulmonary edema induced in the rat lung. AB - The dielectric properties (conductivity, kappa and relative permittivity, epsilon) of excised rat lung are modified by lung air and water content. The measurements of these quantities were made over the frequency range of 10 kHz to 100 MHz with an open-ended coaxial probe. The following relationships were analyzed in an oleic acid-induced pulmonary edema model using 18 animals: the spectra of kappa, epsilon and the loss tangent as a function of lung air and water content. Secondly, an isolated-perfused lung system was produced to induce a gradual increase in lung water. The time course of kappa, epsilon and the loss tangent for one excised lung was analyzed. The principal findings were: (i) a decrease in kappa and epsilon with increasing air content, (ii) an increase in kappa and epsilon with increasing water content, and (iii) a good correlation between lung water content and maximum loss tangent that was insensitive to changes in air content. We conclude that this technique could provide a quantitative assessment of lung water during pulmonary edema formation. PMID- 15652794 TI - Combination of two-dimensional electrophoresis and shotgun peptide sequencing in comparative proteomics. AB - Two-dimensional electrophoresis (2-DE) and shotgun peptide sequencing are the two major technologies to compare the expression profile of proteins, which is also referred to as comparative proteomics or quantitative proteomics. Although the methodologies, such as difference gel electrophoresis for 2-DE and isotope-coded affinity tags for shotgun peptide sequencing, have made rapid progress, these two approaches have their own strengths and weaknesses. Therefore, the combination of the two methodologies is beneficial for the purpose of better comparative proteomics, especially in comprehensive coverage of the proteome and protein information such as post-translational modifications. PMID- 15652796 TI - Geometrical distortions in two-dimensional gels: applicable correction methods. AB - Two-dimensional electrophoresis (2-DE) provides a rapid means for separating thousands of proteins from cell and tissue samples in one run. Although this powerful research tool has been enthusiastically applied in many fields of biomedical research, accurate analysis and interpretation of the data have provided many challenges. Several analysis steps are needed to convert the large amount of noisy data obtained with 2-DE into reliable and interpretable biological information. The goals of such analysis steps include accurate protein detection and quantification, as well as the identification of differentially expressed proteins between samples run on different gels. To achieve these goals, systematic errors such as geometric distortions between the gels must be corrected by using computer-assisted methods. A wide range of computer software has been developed, but no general consensus exists as standard for 2-DE data analysis protocol. The choice of analysis approach is an important element depending both on the data and on the goals of the experiment. Therefore, basic understanding of the algorithms behind the software is required for optimal results. This review highlights some of the common themes in 2-DE data analysis, including protein spot detection and geometric image warping using both spot- and pixel-based approaches. Several computational strategies are overviewed and their relative merits and potential pitfalls discussed. Finally, we offer our own personal view of future trends and developments in large-scale proteome research. PMID- 15652795 TI - Peptidomics, current status. AB - Characterisation of the complement of expressed proteins from a single genome is a central focus of the evolving field of proteomics. Traditional proteomics technologies were developed in the 20th century and are based on two-dimensional electrophoresis or multidimensional liquid chromatography. These facilitated functional genomics analysis, but they currently represent a significant bottleneck to progress in this area. We are now witnessing the development of novel alternative technologies for use in expression proteomics research. This review aims to familiarise the reader with the principles underlying the peptidomics approaches to proteomics research and provide examples of their applications. PMID- 15652797 TI - Strategies for revealing lower abundance proteins in two-dimensional protein maps. AB - One of the most challenging contemporary research endeavors is the mapping of proteins and establishing their linkages to normal and pathological conditions. The availability of current proteomics technologies has greatly facilitated the separation and identification of proteins in a complex protein mixture by standard two-dimensional gel electrophoresis and subsequent MALDI-TOF mass spectrometry. Due to the huge differences in the distribution of proteins in complex proteomes of humans, the detection and identification of proteins expressed in low copy number is a major challenge. The low abundance of important physiologically relevant proteins has rendered their analyses almost impossible without some means of prior purification and enrichment from tissue lysates or biological fluids. It is the current limits of detection of the methods that are used that prevents the detection of these proteins not the proteins themselves. More importantly, considering the frequency at which post-translational modifications of proteins occur, the separation of protein isoforms is essential to understand biological changes, and two-dimensional gel electrophoresis remains the only technique that can offer sufficient resolution to address this issue at a functional level. Cellular fractionation techniques followed by specific affinity probes for tracking target proteins have been developed to deplete the proteome of high abundance proteins in order to increase the sample loading for achieving greater sensitivity for proteins present in low abundance. Those applications can entail the removal of one protein or a class of proteins that interferes with the resolution of proteins in a 2-DE map. Moreover, the use of better solubilizing detergents in combination with an overlapping narrow immobilized pH gradients, results in higher resolution by stretching the protein pattern in the first dimension. In this review we will discuss strategies to remove high abundance proteins that can result in the visualization and detection of low abundance proteins in biological samples. The potential use of these strategies, as a means of developing diagnostic tools for early screening of diseases and identification of drug targets for therapeutic intervention, will also be discussed. PMID- 15652798 TI - Role of chromatographic techniques in proteomic analysis. AB - Proteomics, the characterization of the proteome, is conceptually simple but technically challenging. Development of such technologies as mass spectrometry, multidimensional protein separation, and DNA sequencing has allowed the new field of proteomics to flourish. Proteomic analysis relies on a set of techniques chosen on the basis of the biological question. In any proteomic analysis, the first and most important task is the separation of a complex protein mixture, i.e. the proteome. Chromatography, one of the most powerful methods of separation, employs one or more inherent characteristics of a protein-its mass, isoelectric point, hydrophobicity or biospecificity. This review emphasizes high performance liquid chromatography as an integrated part of technologies used to study the proteome, discusses the capabilities and limitations of current instruments, and highlights the potential of multidimensional liquid chromatography in proteomic analysis. PMID- 15652799 TI - Immunoproteomics. AB - A novel immunoproteomic assay, combining specificity of antibody with precision of mass spectral analysis is described, and a number of practical applications are presented. The assay is carried out in three steps. The first step of the assay involves antibody immobilization, using a bacterial Fc binding support. The second step is antigen capture and washing to remove non-specific binding. The third step involves analysis of the captured antigens by SELDI-TOF. The assay has many advantages in sensitivity, speed, and economy of reagents in detection of specific antigens or antibodies. In addition, under appropriate experimental conditions, semi-quantitative data may be obtained. By combining the increasing range of selective specific antibody reagents available, in part due to advances in antibody engineering technology, and the resolving power available, using mass spectrometry, immunoproteomics is a valuable technique in proteomic analysis. A number of examples of the application of this technique to analysis of biological systems are presented. PMID- 15652800 TI - High-throughput and multiplexed protein array technology: protein-DNA and protein protein interactions. AB - Miniaturized protein arrays address protein interactions with various types of molecules in a high-throughput and multiplexed fashion. This review focuses on achievements in the analysis of protein-DNA and protein-protein interactions. The technological feasibility of protein arrays depends on the different factors that enable the arrayed proteins to recognize molecular partners and on the specificity of the interactions involved. Proteome-scale studies of molecular interactions require high-throughput approaches for both the production and purification of functionally active proteins. Various solutions have been proposed to avoid non-specific protein interactions on array supports and to monitor low-abundance molecules. The data accumulated indicate that this emerging technology is perfectly suited to resolve networks of protein interactions involved in complex physiological and pathological phenomena in different organisms and to develop sensitive tools for biomedical applications. PMID- 15652801 TI - Survey of current protein family databases and their application in comparative, structural and functional genomics. AB - The last two decades have witnessed significant expansions in the databases storing information on the sequences and structures of proteins. This has led to the creation of many excellent protein family resources, which classify proteins according to their evolutionary relationship. These have allowed extensive insights into evolution and particularly how protein function mutates and evolves over time. Such analyses have greatly assisted the inheritance of functional annotations between experimentally characterised and uncharacterised genes. Moreover, the development of bioinformatics tools acts as a companion to the new technologies emerging in biology, such as transcriptomics and proteomics. The latter enable researchers to analyse gene expression profiles and interactions on a genome-wide scale, generating vast datasets of proteins, many of which include experimentally uncharacterised proteins. Protein family/function databases can be used to help interpret this data and allow us to benefit more fully from these technologies. This review aims to summarise the most popular sequence- and structure-based protein family databases. We also cover their application to comparative genomics and the functional annotation of the genomes. PMID- 15652802 TI - System, trends and perspectives of proteomics in dicot plants Part I: Technologies in proteome establishment. AB - The first 3 years of the 21st century have seen the impact of plant proteomics on functional genomics that has enhanced our understanding, not only on the plant genome(s), but also more importantly, on the functional aspect of proteins. This is mainly due to availability of the complete genome sequence of the Arabidopsis thaliana-a dicotyledoneous (dicot) model plant-and technological advancements in proteomics. Proteomic analyses of a variety of dicot plants, including both Arabidopsis and the model legume species, barrel medic (Medicago truncatula), have greatly helped in an efficient separation, identification and cataloguing of a large number of proteins, and thereby defining their proteomes. Therefore, we have composed an inclusive review on dicot plant materials, as of February 2004, that provides system, trends and perspectives of proteomics in growth and development and the environment. The review is summarized and discussed as three individual, but interlinked, entities: Part I, technologies in proteome establishment (this review), Part II, proteomes of the complex developmental stages [G.K. Agrawal, M. Yonekura, Y. Iwahashi, H. Iwahashi, R. Rakwal, J. Chromatogr. B (2004)], and Part III, unraveling the proteomes influenced by the environment, and at the levels of function and genetic relationships [G.K. Agrawal, M. Yonekura, Y. Iwahashi, H. Iwahashi, R. Rakwal, J. Chromatogr. B (2004)]. This review deals with the diverse proteomic technologies being used in proteome development of different dicot plants. PMID- 15652803 TI - System, trends and perspectives of proteomics in dicot plants Part II: Proteomes of the complex developmental stages. AB - This review is devoted to the proteomes of the complex developmental stages of dicotyledoneous (dicot) plant materials. The two core technologies, two dimensional gel electrophoresis (2-DGE) and mass spectrometry (MS), independently or in combination with each other, are propelling dicot plant proteomics to new discoveries and functions, with the establishment of tissue-specific and organelle proteomes, mostly in Arabidopsis thaliana and Medicago truncatula, revealing their complexity and specificity. These experimental proteomes have provided a good start towards the establishment of high-density 2-DGE reference maps and peptide mass fingerprint databases, for not only the model dicot plants, A. thaliana and M. truncatula, but also other important dicot plants, which will serve as a basis for proteomes of many other dicot plants and plant materials. PMID- 15652804 TI - System, trends and perspectives of proteomics in dicot plants. Part III: Unraveling the proteomes influenced by the environment, and at the levels of function and genetic relationships. AB - This review is devoted to the proteomics studies in dicotyledoneous (dicot) plants, such as Arabidopsis, Medicago, potato, soybean, and tomato, under the influence of the environment and at the functional and genetic relationship levels, where the two core technologies, two-dimensional gel electrophoresis (2 DGE) and mass spectrometry (MS) have been instrumental in unraveling the proteomes affected therein. Abiotic and biotic stress responses, including the affect of allergens, the symbiotic interaction between the members of the Leguminoseae family and genera of nitrogen fixing bacteria, phosphoproteomics, and proteomics in revealing the genetic relationships between species and genera have been the subject of many proteomics studies, and these are discussed in this review. In all, these studies have complemented and extended the studies of developmental proteomics [G.K. Agrawal, M. Yonekura, Y. Iwahashi, H. Iwahashi, R. Rakwal, J. Chromatogr. B (2004)]. PMID- 15652805 TI - Proteomic analysis of Korean ginseng (Panax ginseng C.A. Meyer). AB - Although many reports have been published regarding the pharmacological effects of ginseng, little is known about the biochemical pathways operant in ginsenoside biosynthesis, or the genes involved therein. Proteomics analysis is an approach to elucidate the physiological characteristics and biosynthetic pathways of ginsenosides, main components of ginseng. In this review, we introduced the recent progress in proteomics studies of ginseng (Panax ginseng C.A. Meyer). We briefly reference the genomic analyses of P. ginseng, without which proteomics approaches would have been impossible. Functional genomics studies regarding secondary metabolism in P. ginseng are also introduced here, in order to introduce possible future prospects for further study. PMID- 15652806 TI - Comparative proteomic analysis of mammalian animal tissues and body fluids: bovine proteome database. AB - Characterizing the complete proteome of multicellular organisms is a challenging task using the currently available technologies. With the increasing degree of genetic complexity, animals acquire a broader repertoire of options to meet environmental challenges. Mammalian cells from different tissues/body fluids express different thousands of proteins with a predicted dynamic range of up to five to six orders of magnitude, thus necessitating the whole arsenal of dedicated analytical strategies for a detailed proteome characterization. Nevertheless, 2D-E analysis of whole cellular lysates still remains the most used initial approach for the proteomic description of specialized cells. It enables to obtain an overview of the main soluble protein components of a specific tissue/body fluid, allowing comparison between different cellular types and molecular description of organ specialization. Massive proteomic investigations have been reported mainly in the case of human, mouse and rat, allowing comparative analysis. For this reason, a research project focused on the 2D-E characterization of tissues and biological fluids from other domestic mammals has been undertaken in our laboratory. A number of high-resolution reference electrophoretic maps have been established for liver, kidney, muscle, plasma and red blood cells samples from Holstein Friesian bovine female individuals. Among the 1863 distinct protein features detected, 534 species were identified and associated to 209 different genes by a combination of MALDI-TOF mass fingerprint, capillary LC-ESI-IT-MS-MS and image gel matching procedures. Identified polypeptide species and differences in expression profiles between various tissues/fluids clearly reflected organ biochemical specialization. This experimental output allowed establishing a 2D-E bovine database accessible at the URL address for image comparison. PMID- 15652807 TI - Proteomics of bronchoalveolar lavage fluid. AB - Lung diseases are essentially multi-factorial diseases that require a global analysis, and thus, cannot be understood through the sole analysis of individual or small numbers of genes. Proteome analysis has rapidly developed in the post genome era and is now widely accepted as the obligated complementary technology for genetic profiling. It has been shown to be a powerful tool for the study of human diseases and for identifying novel prognostic, diagnostic and therapeutic markers. During last years, proteomic approaches applied to lung diseases are centred on the analysis of proteins in samples, such as cell cultures, biopsies and physiological fluids like serum and, especially, bronchoalveolar lavage fluid (BALF). BALF is presently the most common way of sampling the components of the epithelial lining fluid (ELF) and the most faithful reflect of the protein composition of the pulmonary airways. This review focuses on the state of the investigations of BALF proteome and its powerful contribution both to a better knowledge of the lung structure at the molecular level and to the study of lung disorders at the clinical level. PMID- 15652808 TI - Proteomics analysis of human cerebrospinal fluid. AB - Cerebrospinal fluid (CSF) is secreted from several different central nervous system (CNS) structures, and any changes in the CSF composition will accurately reflect pathological processes. Proteomics offers a comprehensive bird's eye view to analyze CSF proteins at a systems level. This paper reviews the variety of analytical methods that have been used for proteomics analysis of CSF, including sample preparation, two-dimensional liquid and gel electrophoresis, mass spectrometry, bioinformatics, and non-gel methods. The differentially expressed CSF proteins that have been identified by proteomics methods are discussed. PMID- 15652809 TI - Extraction and proteomic analysis of proteins from normal and multiple sclerosis postmortem brain. AB - In this study, a reproducible fractionation procedure was developed to reduce levels of the abundant cytoskeletal proteins that are present in normal and pathological central nervous system (CNS) tissues. The fractionation and proteomic analysis techniques employed greatly facilitated comparison of the spectrum of proteins in normal postmortem brain with proteins in samples from patients with multiple sclerosis, an inflammatory demyelinating disease in which complex changes in protein expression occur as lesions develop. This approach may be of value for the proteomic identification and quantitation of proteins which undergo disease-related changes in CNS disorders, and also for protein expression studies on normal adult and developing CNS tissues. PMID- 15652811 TI - Proteomics of breast carcinoma. AB - Beast cancer is the most diagnosed cancer in women, accounting for approximately 40,000 deaths annually in the USA. Significant advances have been made in the areas of detection and treatment, but a significant number of breast cancers are detected late. The advent of proteomics provides the hope of discovering novel biological markers that can be used for early detection, disease diagnosis, prognostication and prediction of response to therapy. Several proteomics technologies including 2D-PAGE, 2D-DIGE, ICAT, SELDI-TOF, MudPIT and protein arrays have been used to uncover molecular mechanisms associated with breast carcinoma at the global level, and a number of these technologies, particularly the SELDI-TOF hold promise as a proteomic approach that can be applied at the bedside for discovering protein patterns that distinguish disease and disease free states with high sensitivity and specificity. Laser microdissection, a method for selection of homogenous cell populations, coupled to 2D-DIGE or MudPIT constitute a new proteomics-based paradigm for detecting disease in pathology specimens and monitoring disease response to therapy. This review describes proteomics technologies, and their application in the proteomic analysis of breast carcinoma. PMID- 15652810 TI - Revealing urologic diseases by proteomic techniques. AB - Proteomics, as the study of the proteomes of tissues and body fluids, has recently been introduced as a tool for revealing urologic diseases. Two dimensional polyacrylamide gel electrophoresis (2D-PAGE) and surface-enhanced laser desorption/ionzation (SELDI) are two techniques used in proteomic studies. Among the many urologic diseases, the malignancies including prostate cancer, bladder cancer, and renal cancer are the subjects most often selected for proteomic analysis. Poor reproducibility is one of the difficulties that must be overcome in order for proteomic technology to be a robust tool. PMID- 15652812 TI - Effectiveness and limitation of two-dimensional gel electrophoresis in bacterial membrane protein proteomics and perspectives. AB - Two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) using isoelectric focusing and SDS-PAGE in the first and second dimensions, respectively, is an established means of simultaneously separating over 1000 proteins and two new types have recently been developed. These procedures have significant shortcomings such as low load ability and poor separation of hydrophobic, acidic and alkaline proteins. We therefore modified the protocols to analyze the Bacillus subtilis membrane proteome. The 2D-PAGE techniques effectively separated membrane proteins having one and two transmembrane segments but not those with more than four. Compared with new LC/MS/MS procedures that are independent of electrophoretic separation, 2D-PAGE can globally analyze and quantify proteins at various stages of the cell cycle when labeled with isotopes such as 35S methionine or the stable isotope, 15N. PMID- 15652813 TI - Proteomics of Halophilic archaea. AB - Halophilic archaea is a member of the Halobacteriacea family, the only family in the Halobacteriales order. Most Halophilic archaea require 1.5M NaCl both to grow and retain the structural integrity of the cells. The proteins of these organisms have thus been adapted to be active and stable in the hypersaline condition. Consequently, the unique properties of these biocatalysts have resulted in several novel applications in industrial processes. Halophilic archaea are also to be useful for bioremediation of hypersaline environment. Proteome data have expended enormously with the significant advance recently achieved in two dimensional gel electrophoresis (2-DE) and mass spectrometry (MS). The whole genome sequencing of Halobacterium species NRC-1 was completed and this would also provide tremendous help to analyze the protein mass data from the similar strain Halobacterium salinarum. Proteomics coupled with genomic databases now has become a basic tool to understand or identify the function of genes and proteins. In addition, the bioinformatics approach will facilitate to predict the function of novel proteins of Halophilic archaea. This review will discuss current proteome study of Halophilic archaea and introduce the efficient procedures for screening, predicting, and confirming the function of novel halophilic enzymes. PMID- 15652814 TI - Proteomics and physiology of erythritol-producing strains. AB - In-depth knowledge bases on physiological properties of microbes are required to design a better microbial system at a gene level and to develop an industrially viable process in an optimized scheme. Proteomic analyses of industrially useful microorganisms are particularly important for achieving such objectives. In this review, industrial application of erythritol in food and pharmaceutical areas and proteomic techniques for erythritol-producing microbes were presented. Proteomic technologies for erythritol-producing strains such as Candida magnoliae contained protein or peptide sample preparation for two-dimensional electrophoresis and mass spectrometry, analysis of proteome with matrix assisted laser desorption ionization/time-of-flight mass spectrometry, liquid chromatography/electrospray ionization/tandem mass spectrometry and similarity searching algorithms. The proteomic information was applied to predict the carbon metabolism of erythritol synthesizing microorganisms. PMID- 15652815 TI - Role of proteomics in taxonomy: the Mytilus complex as a model of study. AB - Several strategies carried out for the application of proteomic methodologies to species and populations are summarized. Species of the genus Mytilus are used as a model organism to explain these strategies. The proteomics needed to differentiate populations, species, following some different approaches are provided. Moreover, there is an explanation of when it is most critical to carry out a rigorous analysis of this type. Sample preparation, selection of the most appropriate tissue of this organism for a comparative analysis, two-dimensional gels, computer analysis, mass spectrometry (MS) combined with two-dimensional electrophoresis (2-DE) are described. Then, the inconveniences of working with species like those of the genus Mytilus that are poorly represented in databases will be presented along with a description of how to approach this problem. Likewise, the review will include the strategy to follow when dealing with species, like mussels, that present a high degree of genetic polymorphism. The different protein-expression-based strategies used to approach the problem of differentiating Mytilus species will also be shown. Examples are presented to illustrate the use of 2-DE and MS to differenciate populations, species in taxonomic analysis. PMID- 15652817 TI - Asthma, the ugly duckling of lung disease proteomics? AB - The human respiratory system represents a vital but vulnerable system. It is a major target for many diseases such as cancer and asthma. The incidence of these diseases has increased dramatically in the last 40-50 years. In the search for possible new therapies, many experimental tools and methods have been developed to study these diseases, ranging from animal models to in vitro studies. In the last decades, genomic and proteomic approaches have gained a lot of attention. After the major scientific breakthroughs in the field of genomics, it is now widely accepted that to understand biological processes, large-scale protein studies through proteomics techniques are required. In the battle against lung cancer, the proteomics approach has already been successfully implemented. Surprisingly, only a few proteomics studies on the ever-increasing global asthma problem have been published so far. And although proteomics also has its limitations and experimental difficulties, in our opinion, proteomics can definitely contribute to the understanding of a complex disease such as asthma. Therefore, the additional values and possibilities of proteomics in asthma research should be thoroughly investigated. A close collaboration between the different scientific disciplines may eventually lead to the development of new therapeutic strategies against asthma. PMID- 15652816 TI - Methods of comparative proteomic profiling for disease diagnostics. AB - The recent development of numerous technologies for proteome analysis holds the promise of new and more precise methods for disease diagnosis. In this review, we provide an overview of some of these technologies including two-dimensional gel electrophoresis (2DE), historically the workhorse of proteomic analysis, as well as some newer approaches such as liquid phase separations combined with mass spectrometry, and protein microarrays. It is evident that each method has its own strengths and weaknesses and no single method will be optimal in all applications. However, the continuing development of innovative strategies for protein separation and analysis is providing a wealth of new tools for multi dimensional protein profiling that will advance our capabilities in disease diagnostics and our understanding of disease pathology. PMID- 15652818 TI - Proteomic changes during the B cell development. AB - An antibody-secreting B cell is derived from a lymphoid stem cell through a series of developmental stages: progenitor B cell (pro-B cell), precursor B cell (pre-B) cell, immature B cell and mature B cell stage. The gene rearrangement of antigen receptor genes and their expression on the cell surface are crucial regulation steps cells to develop to the next stage. This control mechanism occurs at the sequential manner during the B cell development. Proteomic approach using two-dimensional (2-D) gel electrophoresis and mass spectrometry makes possible to analyze the expression pattern of total proteins at each developmental stage and isolate proteins differentially expressed in B cells. Some transcriptional factors such E2A and Pax5 are expressed at the earlier stages of B cell development and repressed at later developmental stages. RAD52 related protein and chromatin assembly factor 1 are dominantly expressed at early B cells undergoing DNA rearrangement. These comparative analyses of total proteins in each B cell can provide some crucial information to understand the molecular basis of B cell ontogeny. PMID- 15652819 TI - Application of proteomics for comparison of proteome of Neospora caninum and Toxoplasma gondii tachyzoites. AB - Protein profiles of two isolates of Neospora caninum (KBA-2 and JPA1) and Toxoplasma gondii RH strain were investigated by proteomic approach. Approximately, 78% of protein spots on two-dimensional gel electrophoresis (2-DE) profiles and 80% of antigen spots on 2-DE immunoblotting profiles were exhibited to share the same pI and M(r) between KBA-2 and JPA1 of N. caninum. On the other hand, a total of 30 antigen spots of T. gondii were recognized on 2-DE immunoblotting profile using rabbit antiserum against N. caninum KBA-2. A number of homologue proteins, such as heat shock protein 70, tubulin alpha- and beta chain, putative protein disulfide isomerase, actin, enolase and 14-3-3 protein homologue are believed as the conserved proteins in both N. caninum and T. gondii. On the contrary, NcSUB1, NcGRA2 and NCDG1 (NcGRA7) might be the species specific proteins for N. caninum tachyzoites. The present study showed that the high degree of similarity between N. caninum isolates (KBA-2 and JPA1), whereas large differences between N. caninum and T. gondii were noticed by proteome comparisons. PMID- 15652820 TI - Non-enzymatic posttranslational modifications of bovine serum albumin by oxo compounds investigated by chromatographic and electrophoretic methods. AB - Non-enzymatic posttranslational modifications of bovine serum albumin (BSA) by oxo-compounds, particularly glucose, ribose, glyoxal and glutardialdehyde, have been investigated using a set of modern chromatographic and electrophoretic separation methods. High-performance liquid chromatography (HPLC) alternatively with UV spectrophotometric (diode array) or mass spectrometric (MS) detection, polyacrylamide gel electrophoresis (PAGE) with Coomassie brilliant blue staining detection, and capillary zone electrophoresis (CZE) with UV spectrophotometric detection have been employed for the investigation of the chemical and structural changes of BSA caused by its reaction with the above oxo-compounds exhibiting different degree of reactivity. The extent of modifications was found to be dependent on the nature of the oxo-compound used and progressed in the glucose1 year. The preserved OPL had an even profile in 7 eyes and a granulated profile in 7 eyes. Mean VA was 0.19 logMAR with a preserved OPL and 0.72 logMAR with an atrophic OPL (P <.001). Cases seen after the detachment resolved included 6 eyes with preserved even OPL, 5 eyes with preserved granulated OPL, and 5 eyes with atrophic OPL. Mean final VA was 0.06 logMAR in eyes with preserved OPL and 0.90 logMAR in eyes with atrophic OPL (P <.001). The VA improved in 73% of eyes with preserved OPL and no eyes with atrophic OPL (P = .025). The VA recovered completely in 83% of eyes with preserved even OPL and no eyes with preserved granulated OPL (P = .015). CONCLUSION: High-resolution OCT demonstrates changes in the foveal photoreceptor layer in CSC that highly correlate with VA loss and may predict visual recovery after macular reattachment. PMID- 15652831 TI - Evisceration via superior temporal sclerotomy. AB - PURPOSE: To describe evisceration via superior temporal equatorial sclerotomy with preservation of the cornea as an alternative technique. DESIGN: Retrospective case series. METHODS: Records of 31 consecutive patients who underwent evisceration via superior temporal equatorial sclerotomy between November 1996 and November 2002 were reviewed. Patient age ranged from 1 to 79 years (mean, 27.93 years). Posterior sclerotomies were performed in 14 eyes (45%). The surgical indications, prior ocular surgeries, complications, cornea alterations, cornea sensitivities, size and material of the implants, and the motility of the globes with and without prosthesis were evaluated. RESULTS: Average follow-up was 31.68 months (range, 5-77 months). No intraoperative complications occurred. One patient had late-term pain due to partial implant extrusion from the melted cornea. All corneas remained clear until the 1st month after surgery. After the 2nd month some corneas became semiopaque (61%) and totally opaque (16%). Seven corneas remained clear (23%). Cornea vascularization started in the second week postsurgery in all patients. It was limited peripherally in six patients and moved forward to the central part in 25 patients. No patient had corneal sensation. In one case, keratitis occurred at month 13 postsurgery; in another, conjunctival dehiscence in the superior temporal region was noted, 26 months after surgery, due to esotropia resulting from pressure exerted on the region by the prosthesis. Globe motility was satisfactory in all patients. CONCLUSIONS: Implant motility is satisfactory and the implant exposure rate is low with this technique. PMID- 15652833 TI - Glaucoma following penetrating ocular trauma: a cohort study of the United States Eye Injury Registry. AB - PURPOSE: To evaluate associations between baseline structural and functional ocular characteristics and the risk of developing posttraumatic glaucoma after penetrating ocular injury. DESIGN: Prospective cohort study. METHODS: Data from the United States Eye Injury Registry (USEIR) were obtained from a total of 3,627 patients who experienced penetrating ocular injury. The risk of posttraumatic glaucoma and associated structural and functional ocular risk factors was estimated. RESULTS: The risk of developing posttraumatic glaucoma was 2.67%. The development of glaucoma was independently associated with several baseline characteristics including advancing age (relative risk 1.02/yr 95% confidence interval [1.00, 1.03]), lens injury (1.56 [1.03, 2.35]), poor baseline visual acuity (2.59 [1.62, 4.14]), and inflammation (3.02 [1.52, 6.02]). CONCLUSIONS: This study provides an estimate for the risk of developing glaucoma after penetrating ocular injury in a large cohort of patients and determined several factors that are significantly associated with the development of post-traumatic glaucoma, including advancing age, lens injury, poor visual acuity, and intraocular inflammation. PMID- 15652834 TI - Psoriatic uveitis: a distinct clinical entity? AB - PURPOSE: To describe the clinical characteristics of uveitis occurring in patients with psoriasis and to compare them with patients with idiopathic anterior uveitis and HLA-B27-associated anterior uveitis. DESIGN: Case-control study. METHODS: The charts of 36 patients with a diagnosis of uveitis and psoriasis visiting the Immunology and Uveitis Service at the Massachusetts Eye and Ear Infirmary between January 2000 and December 2001 were reviewed. Clinical characteristics, therapy, and outcomes of uveitis were compared with 30 randomly selected patients with either idiopathic anterior uveitis or with HLA-B27 associated anterior uveitis. RESULTS: The mean age at presentation for uveitis was significantly higher in patients with psoriasis compared with nonpsoriatic groups (44.6 years in HLA-B27-psoriatic patients vs 35.9 years in HLA-B27- nonpsoriatic patients, P = .034; 52.7 years in HLA-B27+ psoriatic vs 35.7 years in HLA-B27+ nonpsoriatic patients, P = .001). Uveitis in patients with psoriasis also tended to be bilateral (62%) and prolonged (11.2 weeks). HLA-B27+ patients with psoriasis required supplemental therapy with oral nonsteroidal anti inflammatory drugs (95% confidence interval [CI] 1.41 to 5.36, P = .003) for anterior uveitis more often than did the HLA-B27+ nonpsoriatic group. Patients with psoriasis and uveitis who were HLA-B27+ required oral NSAIDs (95% CI 1.51 to 9.79, P = .001) for anterior uveitis more often than did HLA-B27- psoriatic patients. CONCLUSION: Uveitis in patients with psoriasis may have distinguishing clinical features. Further epidemiologic studies are required to determine the strength of association between psoriasis without arthritis but with uveitis. PMID- 15652835 TI - Tomographic features and surgical outcomes of vitreomacular traction syndrome. AB - PURPOSE: To study the relation between preoperative macular changes and surgical outcomes in vitreomacular traction syndrome. DESIGN: Prospective study. METHODS: We prospectively examined 14 eyes of 13 patients (aged 48 to 82 years; mean 66.1) with vitreomacular traction syndrome using optical coherence tomography (OCT) before and after vitreous surgery. RESULTS: OCT demonstrated two types of partial posterior vitreous detachment: incomplete V-shaped detachment in 10 eyes (group 1) and partial detachment temporal to the fovea but attached nasally in 4 eyes (group 2). Preoperative OCT showed foveal retinal detachment in all eyes in group 1; the detached retina was intact in 2 eyes and edematous with (6 eyes) or without (2 eyes) cystic changes. After surgery, these 10 eyes had a normal foveal configuration accompanied by visual improvement. In group 2, 3 of the 4 eyes had prominent cystoid macular edema (CME) without foveal retinal detachment before surgery. After surgery, 2 eyes developed a full-thickness macular hole, 1 had persistent CME, and 1 developed macular atrophy. The visual acuity decreased in 2 eyes and remained the same in 2 eyes. CONCLUSIONS: Two types of vitreous traction develop in vitreomacular traction syndrome: an incomplete V-shaped posterior vitreous detachment that leads to foveal retinal detachment, the surgical outcome of which is favorable, and partial posterior vitreous detachment temporal to the fovea in which prominent CME developed, which may result in a macular hole or macular atrophy postoperatively. PMID- 15652836 TI - The effect of hinge position and depth plate on the rate of recovery of corneal sensation following LASIK. AB - PURPOSE: To evaluate and compare the effect of hinge position and flap thickness on recovery rate of corneal sensation after (LASIK). DESIGN: Prospective, randomized clinical study. METHODS: Forty eyes of 40 patients underwent LASIK to correct myopia ranging from -1.00 to -2.50 diopters. Corneal sensitivity was measured preoperatively and each month after surgery until recovery to preoperative level. The nasal-hinged ACS microkeratome (NH) with depth plates of 130 and 160 microm and the superior-hinged Hansatome microkeratome (SH) with depth plates of 160 and 180 microm were used for corneal flap creation. The eyes were divided into four groups according to the hinge position and depth plate (DP): group 1, NH and 130 microm DP; group 2, NH and 160 microm DP; group 3, SH and 160 microm DP; group 4, SH and 180 microm DP. RESULTS: Corneal sensitivity returned to preoperative level after 3.7 (+/- 0.4), 4.4 (+/- 0.6), 5.4 (+/- 0.8), and 5.8 (+/- 0.9) months, respectively, in groups 1, 2, 3, and 4. Statistically significant differences were found between groups 1 and 2, but not between groups 3 and 4. Corneal sensitivity gradually returned to preoperative levels in all groups. Sensitivity of the hinge area was higher than other areas at every time point. CONCLUSIONS: The results suggest that after LASIK, hinge position and flap thickness seem to be important factors in the rate of return of corneal sensitivity. A thin flap with a nasally placed hinge is associated with the most rapid recovery of corneal sensitivity. PMID- 15652837 TI - Management of endogenous fungal endophthalmitis with voriconazole and caspofungin. AB - PURPOSE: Voriconazole, a new generation triazole, has been shown to achieve therapeutic intraocular levels after oral administration. Caspofungin is the first approved agent from a new class of antifungals, the echinocandins. This series describes experience at two centers using these novel antifungals to treat endogenous fungal endophthalmitis. DESIGN: Retrospective review. METHODS: Treatment of five patients with Candida endophthalmitis are reviewed. Postmortem intraocular voriconazole concentrations on a sixth patient are presented as well. RESULTS: All patients had systemic cultures positive for Candida species. Three patients had prompt resolution of intraocular mycosis with intravenous and oral voriconazole, caspofungin, or both. The fourth patient with bilateral disease responded well to i.v. voriconazole and caspofungin but had a recurrence when discharged on oral voriconazole and i.v. caspofungin. This patient had a bowel resection with an ileostomy; therefore, absorption of oral voriconazole may have been inadequate. Bilateral amphotericin B intravitreal injection ultimately treated this patient. The fifth patient received 100 microg/0.1 ml of intravitreal voriconazole (final vitreous concentration approximately 25 microg/ml) followed by oral voriconazole and responded favorably. Our sixth patient had multisystem failure and passed away 1 week after initiating intravenous voriconazole for non-ocular candidemia. Postmortem HPLC analysis of the aqueous and vitreous revealed voriconazole concentrations of 1.52 microg/ml and 1.12 microg/ml, respectively (MIC90 of Candida albicans is 0.06 microg/ml). CONCLUSIONS: Voriconazole and caspofungin appear to be powerful weapons to add to the existing armamentarium against fungal endophthalmitis. Further studies are warranted to define precisely the role of these new agents alone or in combination with other antifungals. PMID- 15652839 TI - Macular hole hypotheses. PMID- 15652838 TI - A retrospective study of endophtalmitis rates comparing quinolone antibiotics. AB - PURPOSE: To compare endophthalmitis rates between topical quinolone antibiotics over a four-year period in a university setting. DESIGN: Retrospective, cross sectional (prevalence) study. METHODS: STUDY POPULATION: Nine thousand seventy nine patients who underwent a phacoemulsification procedure at a University Eye Center. INTERVENTIONS: The following interventions were conducted: use of a nosocomial infectious reporting database retrospective analysis of prospectively collected data to establish endophthalmitis rates; prevalence analysis of postoperative quinolone antibiotics; analysis of endophthalmitis rate vs postoperative quinolone prescribed. MAIN OUTCOME MEASURES: Occurrence of endophthalmitis after cataract surgery. RESULTS: During a four-year period involving 9079 phacoemulsification procedures, 26 cases of bacterial postoperative endophthalmitis were reported (rate = 0.286%). Eight of the 26 cases were considered complicated; 18 cases were uncomplicated. Among cases performed during this period, postoperative prescription volumes for ciprofloxacin (Ciloxan, Alcon Laboratories, Inc., Fortworth, Texas) and ofloxacin (Ocuflox, Allergan, Inc., Irvine, California) antibiotic drops were nearly identical, with 4538 patients receiving ciprofloxacin (49.98%) and 4541 patients receiving ofloxacin (50.02%). There were 22 endophthalmitis patients (85%) who had used topical ciprofloxacin and 4 patients (15%) who had used topical ofloxacin postoperatively. The difference in infectious rates between antibiotics was highly significant (P < .00026). For uncomplicated cases, 14 patients received ciprofloxacin and 4 patients received ofloxacin. This difference was also significant (P < .015). CONCLUSIONS: Differences in pharmacokinetic and pharmacodynamic properties exist among quinolone antibiotics, which may affect endophthalmitis incidence following cataract surgery. PMID- 15652840 TI - The esthetics of orbital exenteration. PMID- 15652841 TI - Artifacts in optical coherence tomography topographic maps. PMID- 15652842 TI - The Open Access initiative in scientific and biomedical publishing: fourth in the series on editorship. AB - PURPOSE: To provide basic information about the Open Access concept and its historical development, define the benefits and challenges inherent in this new model, and identify the value of the traditional print model and its movement towards more open access. DESIGN: Review of current information on the subject from numerous sources. METHODS: Medline search and Internet search engines on the topic of Open Access Publishing. RESULTS: The Open Access initiative derives from several premises: medical libraries can no longer afford journal subscriptions; society benefits from the open exchange of ideas; society has in large part already paid for this research; the Internet provides an available venue. The traditional journal publishers model, however, has functioned well over many years with a robust peer review system and increasing Internet digital components permitting search and cross referencing, including elements of the Open Access model. CONCLUSIONS: It will be difficult to maintain the costs of both the traditional journal system and the fully implemented Open Access model. Any decisions that are made must ensure that the archive of prior medical knowledge is not lost, that financial barriers do not restrict publication, and that research continues to be available to those who need it, in the media that they prefer. PMID- 15652843 TI - Long-term histopathologic findings in human corneal wounds after refractive surgical procedures. AB - OBJECTIVE: To describe the human corneal histopathology after laser in situ keratomileusis, photorefractive keratectomy, astigmatic keratoplasty, clear corneal refractive lens surgery, conductive keratoplasty, and intrastromal corneal ring segment placement. DESIGN: Retrospective histopathologic study and selected literature review. METHODS: Ninety-four postmortem corneas from 49 patients and 10 corneal button specimens from 6 patients with previous refractive surgery were collected from eye banks and refractive surgeons in North America. This observational case series evaluated the pathologic findings in the wounds of these corneas in the context of previous studies. RESULTS: Three histopathologic types of corneal wound-healing responses were present in corneas after refractive surgery. The first was an epithelial cell modification that reestablished a smooth anterior corneal surface through basal epithelial cell hypertrophy, by epithelial hyperplasia, or both. The second was the production of a hypercellular fibrotic stromal scar, and the third was the production of a hypocellular primitive stromal scar. Although only the hypercellular fibrotic stromal scar could be identified on gross examination, all three wound responses were identified with light, electron, and immunofluorescence microscopy. Confocal microscopy also identified these three processes but with less detail and resolution. CONCLUSION: Understanding the locations, clinical appearance, and functional attributes of these wound-healing responses will help guide evolving techniques or adjunctive treatments. PMID- 15652844 TI - Symptoms of musculoskeletal disorders in ophthalmologists. AB - OBJECTIVE: To describe the presence of musculoskeletal disorders (MSDs) of the neck, low back, and upper extremity among ophthalmologists. DESIGN: Survey. METHODS: Mail survey using a pretested instrument to 2,529 ophthalmologists in the Northeastern United States; 697 ophthalmologists (28%) returned a completed survey. RESULTS: Self-reported prevalence of neck, upper body, or lower back symptoms in the prior month was 51.8%. Low back pain was present in 39% of respondents, followed by upper extremity symptoms (32.9%) and neck pain (32.6%). Approximately 15% of ophthalmologists were slightly to moderately limited in their work as a result of these symptoms. CONCLUSION: MSD symptoms appear to be common among ophthalmologists. With a relatively low response rate, however, it is plausible that asymptomatic subjects represent a large percentage of the nonrespondents. Additional investigation is warranted to evaluate ergonomic risk factor exposure and establish injury prevention guidelines. PMID- 15652845 TI - Homocysteine and retinal vein occlusion: a population-based study. AB - PURPOSE: To report population data on the association between homocysteinemia and retinal vein occlusion (RVO). DESIGN: Cross-sectional, population-based study. METHODS: Total serum homocysteine (tHcy) was estimated in subjects attending the second Blue Mountains Eye Study (BMES) (n = 3509, aged >49 years). Its association with RVO (graded from retinal photographs) was examined. RESULTS: Homocysteine was more strongly associated with RVO in younger than older participants. In those aged <70 years, homocysteine levels >15 micromol/l were associated with increased odds of RVO, odds ratio (OR) 3.76; 95% confidence interval (CI) 1.06-13.40, in multivariable-adjusted analyses. Each 1 micromol/l increase in homocysteine was associated with a 7% increased odds of RVO, OR 1.07; CI 0.99-1.17 (P = .08). CONCLUSION: This population-based study showed that elevated serum homocysteine is associated with the presence of RVO, independent of other risk factors. PMID- 15652846 TI - Persistent outer retinal defect after successful macular hole repair. AB - PURPOSE: To report two cases of a persistent outer retinal defect evident by optical coherence tomography (OCT) after clinically successful macular hole surgery. DESIGN: Retrospective case series. METHODS: A retrospective case series of two patients who had a persistent outer retinal defect by OCT and excellent visual acuity after clinically successful macular hole surgery. RESULTS: Two patients with stage 2 macular holes and preoperative visual acuity of 20/70 and 20/60, respectively, underwent macular hole surgery. Although the macular holes were closed on clinical evaluation, a persistent outer retinal defect was identified by OCT in both patients. At 5 to 6 months postoperatively, the outer retinal defect became less prominent and the visual acuity remained at 20/20 in both patients. CONCLUSIONS: An outer retinal defect by OCT may occur in the early postoperative period after macular hole surgery. Despite this defect, good visual acuity is possible. PMID- 15652847 TI - Surgical management for premacular membranes in children. AB - PURPOSE: To access the efficacy of surgical treatment for premacular membranes in four children. DESIGN: Interventional case series. METHODS: Premacular membranes in four children were observed. Initial visual acuity ranged from 0.08 to 0.4. The premacular membranes were approximately 1/3 PD to 4 PD in size, and the center of them was located at the fovea in all the patients. Vitreous surgeries were performed in all four patients. The membranes were removed successfully without complications. Occlusion therapy was performed one month after the operation. Follow-up was from 6 to 16 months (means 8.3 months). RESULTS: Vision eventually improved in all patients. Final vision acuity ranged from 0.2 to 1.0. CONCLUSIONS: Vitreous surgeries may be safe and effective in children with premacular membrances. PMID- 15652848 TI - A novel missense mutation in a Japanese patient with gelatinous droplike corneal dystrophy. AB - PURPOSE: To report a novel missense mutation in TACSTD2 gene, L186P, responsible for gelatinous droplike dystrophy (GDLD). DESIGN: Case report and experimental study. METHOD: A 10-year-old Japanese boy suffering from typical GDLD was studied. A 1.1-kb DNA fragment of the TACSTD2 gene was amplified and analyzed using a molecular biological method. cDNA from the patient's cornea was also analyzed to determine which allele was expressed in the patient's corneal epithelium. RESULTS: Sequence analysis revealed that the patient is a compound heterozygote for the Q118X mutation and the L186P, the first missense mutation found in Japanese GDLD. Polymerase chain reaction-restriction fragment length polymorphism analysis from cDNA of patient's cornea revealed that the L186P missense mutation allele is expressed in the patient's corneal epithelium. CONCLUSION: We describe a novel mutation in one case of Japanese GDLD. The results confirm that the missense mutation L186P in the TACSTD2 gene is also responsible for the GDLD phenotype. PMID- 15652849 TI - Adenoma arising from nonpigmented ciliary epithelium concomitant with neovascularization of the optic disk and cystoid macular edema. AB - PURPOSE: To report a case of adenoma of the nonpigmented ciliary epithelium concomitant with marked neovascularization of the optic disk and cystoid macular edema (CME). DESIGN: Observational case report. METHODS: A 34-year-old woman presented with a nonpigmented, vascularized tumor behind the iris and neovascularization of the optic disk (NVD). Visual acuity decreased gradually as a result of the development of CME. RESULTS: Iridocyclectomy to excise the tumor was performed 8 months after initial examination. Histopathologic study revealed an adenoma arising from the NPCE. The levels of vascular endothelial growth factor were significantly elevated in both aqueous and vitreous humor obtained at surgery. The NVD and CME regressed after surgery, and the postoperative visual acuity improved to 20/40. CONCLUSIONS: NVD and CME may develop secondary to adenoma of the nonpigmented ciliary epithelium. Increased levels of vascular endothelial growth factor in intraocular fluids may play a role in NVD and CME development. PMID- 15652850 TI - Blinding keratoconjunctivitis and child abuse. AB - PURPOSE: To report an unusual, blinding inflicted eye injury in young children. DESIGN: Observational case report. METHODS: Retrospective study in an institutional clinical practice of two families in whom the probands had inferior half keratoconjunctivitis and additional signs of child abuse. RESULTS: Two unrelated infants presented with bilateral, asymmetrical, external eye disease affecting the lower half of the cornea and conjunctiva. One eye had perforated. All eyes recovered quickly while the patients were in the hospital with no specific treatment. There were other signs of child abuse detected by further studies on the patients, and in one case, the younger sibling was the subject of severely damaging physical abuse. CONCLUSIONS: Inflicted corneal injuries are nonspecific, and unexplained keratoconjunctivitis, especially in the lower half of the conjunctiva and cornea in infants should alert the clinician to the possibility child abuse, but, by itself cannot be taken as being pathognomonic of abuse. PMID- 15652851 TI - Novel CHST6 nonsense and missense mutations responsible for macular corneal dystrophy. AB - PURPOSE: To identify the underlying mutations in two unrelated British families with macular corneal dystrophy (MCD) by screening the carbohydrate sulfotransferase (CHST6) gene. DESIGN: Case reports and results of DNA analysis. METHODS: Two subjects from two British families with MCD were studied. The genetic status of CHST6 was determined for all members of these MCD families. In addition, sulfated keratan sulfate (KS) assay from the probands was also undertaken. CHST6 gene was amplified by polymerase chain reaction (PCR). The PCR products were analyzed by sequencing and restriction digestion. Enzyme-linked immunosorbent assay (ELISA) was performed to assess KS presence in serum. RESULTS: Four compound heterozygous mutations were identified, three of which are novel. The ELISA showed that the probands were of MCD type I. CONCLUSIONS: These novel mutations are expected to result in loss of CHST6 function, which would account for the MCD phenotype. PMID- 15652852 TI - Testing distance stereoacuity with the Frisby-Davis 2 (FD2) test. AB - PURPOSE: To develop a presentation protocol for the new Frisby-Davis 2 (FD2) distance stereoacuity test. DESIGN: Prospective data collection. METHODS: Stereoacuity was tested monocularly and binocularly in 95 patients with a variety of strabismic and nonstrabismic conditions, using the FD2, employing a modified staircase procedure. The Preschool Randot Stereoacuity test and the near Frisby test were used to determine whether a patient was stereoblind. RESULTS: Under monocular conditions, 35 (37%) of 95 patients passed at least the largest disparity of the FD2 indicating a problem with monocular cues. The binocular protocol was then modified to include a monocular test phase. Using the new protocol, if a patient could achieve the same stereoacuity under monocular and binocular conditions, they were deemed to have no stereopsis. Testing 28 additional stereoblind patients using the new modified protocol revealed no false positives. CONCLUSION: The FD2 stereotest is a useful measure of distance stereoacuity, provided the presentation protocol accounts for monocular cues. PMID- 15652853 TI - Delayed suprachoroidal hemorrhage and factor VIII deficiency. AB - PURPOSE: To describe the clinical course of a 52-year-old white man with previously undiagnosed hemophilia who suffered blunt trauma to the right eye with consequent delayed, spontaneous suprachoroidal hemorrhage. DESIGN: Observational case report. METHODS: The patient's clinical course from the time of trauma to postenucleation is discussed and correlated with B-scan ultrasonography and histopathology. RESULTS: The patient developed recurrent hyphemas, delayed choroidal hemorrhage with detachment and eventually loss of the eye. Hematologic evaluation established a diagnosis of mild factor VIII deficiency. CONCLUSION: Despite mild hemophilia, this patient had not experienced any clinically important morbidity until his eye injury. Ophthalmologists should be aware that bleeding disorders such as hemophilia predispose patients to spontaneous intraocular hemorrhage. PMID- 15652854 TI - Ultrastructure of internal limiting membrane in myopic foveoschisis. AB - PURPOSE: To reveal the pathogenesis of myopic foveoschisis (MF). DESIGN: Clinicopathological report. METHODS: Internal limiting membranes (ILMs) were collected from ten patients with MF and five patients with idiopathic macular hole (IMH) as a control. Samples were subjected to transmission electron microscopic study. Characteristics of the ILM were compared between the two groups. RESULTS: Collagen fiber and cell debris were identified on the inner surface of ILM in seven eyes (70%) with MF, significantly more (P < .05) than found in IMH subjects (0%). More fibrous glial cells were likely to be found on the inner surface of ILM. No significant difference in fibroblast-like cell adhesion was observed. CONCLUSIONS: Collagen fiber and cellular component are suggested to play an important role in developing MF. ILM peeling may be essential for vitrectomy for MF. PMID- 15652855 TI - Calotropis procera (ushaar) keratitis. AB - PURPOSE: To report a case of permanent endothelial cell injury after intracorneal penetration of milky latex from Calotropis procera (ushaar). DESIGN: Interventional case report. METHODS: A 40-year-old patient developed painless corneal edema despite minimal epithelial injury after exposure to ushaar latex. RESULTS: Confocal and specular microscopy confirmed permanent endothelial cell loss with morphologic alteration after intracorneal penetration of ushaar latex. Corneal edema resolved completely after 2 weeks, although reduced endothelial cell count and abnormal morphology persisted. CONCLUSION: Ushaar latex is capable of penetrating the corneal stroma and inducing permanent loss of endothelial cells. Corneal edema resolves if sufficient endothelial cell viability is still present after resolution of ushaar keratitis. PMID- 15652856 TI - Abdominal cramp as an adverse effect of travoprost. AB - PURPOSE: To describe the adverse effect of abdominal cramp caused by travoprost ophthalmic solution in a glaucoma patient. DESIGN: Case report. METHODS: A 34 year-old woman reported abdominal cramp on topical application of travoprost ophthalmic solution. Satisfactory dechallenge and rechallenge procedures were performed. RESULTS: Abdominal cramp occurred approximately 30 minutes after topical application of travoprost on 3 consecutive days. No abdominal cramp developed after substitution of travoprost with latanoprost or normal saline (in the Travatan bottle). Abdominal cramps recurred on rechallenging with travoprost, either in the original Travatan bottle, a latanoprost bottle, or in a Rescula bottle. CONCLUSIONS: Flu-like symptoms (abdominal cramps, malaise and URI) were reported in 4% of patients participating in clinical trials of travoprost but seldom reported postmarketing. Based on the findings of dechallenge and rechallenge procedures, abdominal cramp should be considered a certain adverse effect of travoprost according to the WHO classification. PMID- 15652857 TI - Macular abnormalities and optic disk anomaly associated with a new PAX2 missense mutation. AB - PURPOSE: To report a family with macular abnormalities accompanied by anomalies of the optic disk and kidney associated with a new PAX2 missense mutation. DESIGN: Observational case report. METHODS: A 34-year-old female presented with horizontal nystagmus, poor visual acuity, and chronic renal failure. She had bilateral colobomatous disk anomaly and foveal hypoplasia. Her mother also had renal dysfunction and bilaterally impaired vision. Besides the optic disk dysplasia, the fovea was hypoplastic in the right eye, whereas pigmented macular atrophy was observed in the left eye. The entire coding regions of PAX2 and PAX6 were screened for mutations. RESULTS: A heterozygous mutation G755C in exon 2 of PAX2 that results in a missense mutation, R71T, was identified in the proband and her mother. No mutations were detected in PAX6. CONCLUSIONS: A new PAX2 missense mutation, R71T, may cause macular abnormalities in addition to anomalies of the optic disk and the kidney. PMID- 15652858 TI - Central retinal artery occlusion after radial optic neurotomy in a patient with central retinal vein occlusion. AB - PURPOSE: To describe a patient with a central retinal vein occlusion (CRVO) who developed central retinal artery occlusion after radial optic neurotomy. DESIGN: Interventional case report. METHODS: A 70-year-old woman with CRVO underwent a radial optic neurotomy on her right eye. Her preoperative visual acuity in the affected eye was 20/400. RESULTS: Radial optic neurotomy was performed after phacoemulsification and aspiration for a cataract with intraocular lens implantation. At the insertion of a CRVO knife, pulsating bleeding occurred from the cup of the optic disk; the bleeding was stopped within 2 minutes by elevating the intraocular pressure to 80 mm Hg. On the following day, the patient noticed that she had lost light perception. Fluorescein angiography showed a marked delay of arterial filling, indicating a central retinal artery occlusion. Retinal circulation returned to normal 2 months later; however, her vision was still no light perception. CONCLUSIONS: Ophthalmologists should be aware that severe complications such as central retinal artery occlusion can be associated with radial optic neurotomy, which is an unproven surgical procedure with a questionable pathophysiologic mechanism. PMID- 15652859 TI - Ocular surface squamous neoplasia in Papillon-Lefevre syndrome. AB - PURPOSE: To report the occurrence of ocular surface squamous neoplasia (OSSN) in a patient with Papillon-Lefevre syndrome. DESIGN: Interventional case report. METHODS: A 14-year-old boy presented with a progressively growing limbal lesion in the right eye for 2 months. He had associated erythematous scaly lesions on the elbows, knees, and feet with periodontosis suggestive of Papillon-Lefevre syndrome. An excision biopsy of the limbal tumor and a skin biopsy was performed. RESULTS: The limbal tumor showed features of carcinoma in situ with clear margins. The skin biopsy showed epidermal hyperplasia and hyperkeratosis with perivascular infiltrates, consistent with Papillon-Lefevre syndrome. CONCLUSIONS: Ocular surface squamous neoplasia may occur in patients with Papillon-Lefevre syndrome, an association that to the best of our knowledge has not been previously reported. PMID- 15652860 TI - Removal of the intruding Miragel's scleral buckle by pars plana ultrasonic fragmentation. AB - PURPOSE: To introduce a surgical approach for removing intruded scleral buckle resulting from Miragel's implant. DESIGN: Interventional case report. METHODS: After cutting and releasing scleral band externally, a pars plana Fragmatome was used to break up and aspirate pieces of the Miragel's implant that had grown into the eye, filling the vitreous cavity. RESULTS: The intruding Miragel's buckle was shaved to the plane of the retina without any complication. The retina was completely attached. CONCLUSION: Use of ultrasonic fragmentation by a pars plana approach was effective in removing the intavitreally expanded buckling element without harming the retina. PMID- 15652861 TI - Retinal neovascularization associated with retinoblastoma. AB - PURPOSE: To report retinal neovascularization associated with retinoblastoma in a 14-month-old infant. DESIGN: Observational case report. METHODS: Review of clinical and pathologic findings. RESULTS: A large frond of retinal neovascularization was present posterior to the lens in the right eye, which also contained a retinoblastoma. CONCLUSIONS: Retinal neovascularization is an unusual association with retinoblastoma. PMID- 15652862 TI - A device to facilitate limbal stem cell procurement from eye bank donor tissue for keratolimbal allograft procedures. AB - PURPOSE: To develop a device that facilitates the procurement of corneal limbal stem cell grafts for keratolimbal allograft procedures used in the treatment of ocular surface disease associated with stem cell deficiency. DESIGN: Description of device design and technique for use. METHODS: The device is composed of a pedestal with a convex surface mounted to a flat platform. A corneoscleral button placed endothelial side down and centrally upon the convexity is secured by suction conveyed through a hollowed core in the pedestal that connects to fenestrated openings on the convex surface. A donut-shaped stainless steel ring placed on tension by springs braces the peripheral tissue. A circular corneal incision is created of a desired thickness by a suction trephine, and a crescent blade is utilized to peripherally dissect a donut-shaped keratolimbal allograft. RESULTS: This device facilitated the harvesting of the keratolimbal allograft tissue from four eye bank donor practice corneoscleral buttons and was then used to successfully procure grafts from six corneoscleral buttons used in three keratolimbal allograft procedures in three patients, one each with aniridia, alkali burn, and drug-induced limbal stem cell deficiency. CONCLUSIONS: The described device effectively facilitates procurement of corneoscleral buttons for keratolimbal allograft procedures. It appears to offer advantages over freehanded techniques and previously described devices used for the same purpose. PMID- 15652863 TI - Functional changes after photodynamic therapy with verteporfin. AB - OBJECTIVE: To investigate image preferences after photodynamic therapy (PDT). DESIGN: Prospective, cross-sectional study. METHODS: Seventeen patients with neovascular age-related macular degeneration were tested by color vision, contrast sensitivity, and near visual acuity (VA) with positive images and negative images before, 1 week after, and 3 months after PDT. RESULTS: Before PDT, 13 patients (76%) preferred positive images; the difference was not significant (P < .04). One week after PDT, 16 patients (94%) preferred negative images. The average near VA improved to 27 letters on positive images and to 32 letters on negative images (P < .00001). Three months after PDT, near VA declined to 19 letters on positive images and to 26 letters on negative images (P < .000001). Color vision and contrast sensitivity remained constant during all examinations. CONCLUSION: Although PDT may not alter the neurosensory retina, it may affect intraretinal function by changing image preference. PMID- 15652864 TI - Valganciclovir therapy for immune recovery uveitis complicated by macular edema. PMID- 15652866 TI - EDTA chelation for calcific band keratopathy. PMID- 15652868 TI - Postsynaptic signaling networks: cellular cogwheels underlying long-term plasticity. AB - Learning depends on positive or negative changes in synaptic transmission that are synapse-specific and sustained. Synaptic signals can be directly measured and respond to certain kinds of stimulation by becoming persistently enhanced (long term potentiation, LTP) or decreased (long-term depression, LTD). Studying LTP and LTD opens a window on to the molecular mechanisms of memory. Although changes in both pre- and postsynaptic strength have been implicated in LTP and LTD, most attention has been focused on changes in postsynaptic glutamate receptor density. This is controlled by intracellular Ca(2+) ions via a network of signaling molecules. Changes in postsynaptic Ca(2+) concentration depend on the coincidence of appropriate synaptic signals, as is found in learning situations. The long term persistence of LTP and LTD requires gene transcription and translation. It is posited that local translation at the synapse, in a self-sustaining manner, mediates the persistence of long-term changes despite constant turnover of the synaptic components. PMID- 15652869 TI - Early developmental exposure to methylphenidate reduces cocaine-induced potentiation of brain stimulation reward in rats. AB - BACKGROUND: Methylphenidate (MPH) is prescribed for the treatment of attention and hyperactivity disorders. We showed previously that early developmental exposure to MPH in rats causes behavioral alterations during adulthood, including reduced cocaine reward in place conditioning studies. Here we examined if early MPH exposure alters the ability of cocaine to potentiate the rewarding effects of electrical stimulation of the medial forebrain bundle (MFB) using intracranial self-stimulation (ICSS). METHODS: Rats received MPH or saline during pre adolescence (P20-35) and were implanted with MFB stimulating electrodes at adulthood (P60). Rats then were tested with cocaine in the ICSS paradigm. RESULTS: Cocaine dose-dependently decreased ICSS thresholds in all rats, but the threshold-lowering effects of cocaine were smaller in rats exposed to MPH during pre-adolescence. There were no differences between groups in sensitivity to the rewarding effects of MFB stimulation itself. CONCLUSIONS: Early developmental exposure to MPH reduces the reward-related effects of cocaine in the ICSS paradigm. These results are consistent with previous studies in which early exposure to MPH reduced the ability of cocaine to establish conditioned place preferences, as well as the rewarding effects of sucrose and sexual behavior. Reduced sensitivity to these various types of reward may reflect general dysfunctions of brain reward systems. PMID- 15652870 TI - Localized enlargement of the frontal cortex in early autism. AB - BACKGROUND: Evidence from behavioral, imaging, and postmortem studies indicates that the frontal lobe, as well as other brain regions such as the cerebellum and limbic system, develops abnormally in children with autism. It is not yet clear to what extent the frontal lobe is affected; that is, whether all regions of frontal cortex show the same signs of structural maldevelopment. METHODS: In the present study, we measured cortical volume in four subregions of the frontal cortex in 2-year-old to 9-year-old boys with autism and normal control boys. RESULTS: The dorsolateral region showed a reduced age effect in patients when compared with control subjects, with a predicted 10% increase in volume from 2 years of age to 9 years of age compared with a predicted 48% increase for control subjects. In a separate analysis, dorsolateral and medial frontal regions were significantly enlarged in patients aged 2 to 5 years compared with control subjects of the same age, but the precentral gyrus and orbital cortex were not. CONCLUSIONS: These data indicate regional variation in the degree of frontocortical overgrowth with a possible bias toward later developing or association areas. Possible mechanisms for these regional differences are discussed. PMID- 15652871 TI - Nocturnal excretion of 6-sulphatoxymelatonin in children and adolescents with autistic disorder. AB - BACKGROUND: Many studies in autistic disorder report sleep problems and altered circadian rhythms, suggesting abnormalities in melatonin physiology. Additionally, melatonin, a pineal gland hormone produced from serotonin, is of special interest in autistic disorder given reported alterations in central and peripheral serotonin neurobiology. METHODS: Nocturnal urinary excretion of 6 sulphatoxymelatonin was measured by radioimmunoassay in groups of children and adolescents with autistic disorder (n = 49) and normal control individuals (n = 88) matched on age, sex, and Tanner stage of puberty. RESULTS: Nocturnal 6 sulphatoxymelatonin excretion rate was significantly and substantially lower in patients with autism than in normal controls (mean +/- SEM, .75 +/- .11 vs. 1.80 +/- .17 microg/hr, p =.0001), and was significantly negatively correlated with severity of autistic impairments in verbal communication and play (p < .05). CONCLUSIONS: These findings indicate clearly that nocturnal production of melatonin is reduced in autism. Further research is warranted in order to understand the mechanisms underlying the lower melatonin production, to assess the impact of altered melatonin on the pathophysiology and behavioral expression of autistic disorder, and to determine the utility of melatonin administration in individuals with autism. PMID- 15652872 TI - Catechol-O-methyltransferase gene Val/Met functional polymorphism and risk of schizophrenia: a large-scale association study plus meta-analysis. AB - BACKGROUND: A common functional polymorphism (Val/Met) in the catechol-O methyltransferase gene (COMT) that markedly affects enzyme activity has been shown to affect executive cognition and the physiology of the prefrontal cortex in humans. It is hypothesized that the high activity Val allele slightly increases risk for schizophrenia through its effect on dopamine-mediated prefrontal information processing. METHODS: We compared the allele/genotype frequencies of the Val/Met polymorphism in a large independent patient-control sample (862 patient and 928 healthy control subjects) from Han Chinese population, and an update meta-analysis was performed to assess the collective evidence across individual studies. RESULTS: No statistically significant differences were found in allele or genotype frequencies between patient and normal control subjects, although a nonsignificant overrepresentation of the Val allele in schizophrenia patients (odds ratio [OR] = 1.09, 95% confidence interval [CI] = .94-1.26) was suggested. Comparatively, the meta-analysis of all published population-based association studies showed statistically significant evidence for heterogeneity among the group of studies. Stratification of the studies by ethnicity of the samples yielded no significant evidence for an association with the Val allele in Asian population (OR = .96, 95% CI = .85-1.09), nor in European population (OR = 1.06, 95% CI = .95-1.19). CONCLUSIONS: Our data provide minimal evidence that the Val allele is a susceptibility factor for schizophrenia in either European or Asian populations. PMID- 15652873 TI - Abnormalities in visually guided saccades suggest corticofugal dysregulation in never-treated schizophrenia. AB - BACKGROUND: Previous studies have reported intact visually guided saccades in schizophrenia, but these are limited by potential acute and long-term pharmacological treatment effects, small sample sizes, and a failure to follow patients over time. METHODS: Visually guided saccades were examined in 44 antipsychotic-naive patients experiencing their first episode of schizophrenia prior to treatment and again after 6, 26, and 52 weeks of antipsychotic treatment. Thirty-nine matched healthy individuals were followed over the same period. RESULTS: Before treatment, patients showed faster saccade latencies to unpredictable visual targets, suggesting reduced inhibitory regulation of brainstem saccade generators by neocortical attentional systems. Risperidone treatment reduced this deficit, suggesting a facilitation of attentional function, but haloperidol treatment did not. However, there was also a modest decline in saccade accuracy after risperidone treatment. The ability to sustain fixation of static central and peripheral targets was unimpaired before and after treatment. CONCLUSIONS: These findings provide evidence for impairments in neocortical attentional systems that cause reduced corticofugal regulation of brainstem systems in schizophrenia. This dysfunction appears to be minimized by the atypical antipsychotic risperidone but at the cost of a subtle reduction in saccade accuracy, possibly mediated via adverse effects on cerebellar vermis function. PMID- 15652874 TI - Effects of adjunctive mifepristone (RU-486) administration on neurocognitive function and symptoms in schizophrenia. AB - BACKGROUND: It has been suggested that hypercortisolemia may cause or exacerbate both neurocognitive impairment and symptoms in schizophrenia. We hypothesized that antiglucocorticoid treatments, particularly glucocorticoid receptor (GR) antagonists, would improve neurocognitive functioning and clinical symptoms in this disorder. METHOD: Twenty patients with schizophrenia were treated with 600 mg/day of the GR-antagonist mifepristone (RU-486) or placebo for 1 week in a double-blind, crossover design. Neurocognitive function was evaluated at baseline and 2 weeks after each treatment. Neuroendocrine profiling was performed at these times and also immediately after each treatment. Symptoms were evaluated weekly. RESULTS: Mifepristone administration resulted in a temporary two- to threefold increase in plasma cortisol levels (p < .0001). No significant effects were observed on any measure of neurocognitive function, including the primary outcome measures of spatial working memory and declarative memory. Minor changes in symptoms occurred in both arms of the study and were indicative of a general improvement over time, irrespective of treatment. CONCLUSIONS: In contrast to our earlier report of positive effects in bipolar disorder, these data suggest that the GR-antagonist mifepristone has no effect on neurocognitive function or symptoms in this group of patients with schizophrenia. Future studies in schizophrenia should examine patients with demonstrable hypothalmic-pituitary adrenal axis dysfunction. PMID- 15652875 TI - Transcranial magnetic stimulation accelerates the antidepressant effect of amitriptyline in severe depression: a double-blind placebo-controlled study. AB - BACKGROUND: Transcranial magnetic stimulation (TMS) is a noninvasive method to stimulate the cortex, and the treatment of depression is one of its potential therapeutic applications. Three recent meta analyses strongly suggest its benefits in the treatment of depression. The present study investigates whether repetitive TMS (rTMS) accelerates the onset of action and increases the therapeutic effects of amitriptyline. METHODS: Forty-six outpatients meeting DSM IV criteria for nonpsychotic depressive episode were randomly assigned to receive rTMS (n = 22) or sham repetitive TMS (sham) (n = 24) during 4 weeks over dorsolateral prefrontal cortex (DLPFC) in this double-blind controlled trial. All patients were concomitantly taking amitriptyline (mean dose 110 mg/d). The rTMS group received 20 sessions (5 sections per week) of 5 Hz rTMS (120% of motor threshold and 1250 pulses per session). Sham stimulation followed the same schedule, however, using a sham coil. The efficacy variables were the Hamilton Depression Rating Scale-17 items (HAM-D/17), the Montgomery-Asberg Depression Rating Scale (MADRS), a Visual Analogue Scale (VAS), and the Clinical Global Impression (CGI). Tolerability was assessed by clinical examination and a safety screening of TMS side effects. RESULTS: Repetitive TMS had a significantly faster response to amitriptyline. There was a significant decrease in HAM-D/17 scores, already after the first week of treatment (p < .001 compared with baseline and p < .001 compared with sham). The decrease in HAM-D/17 scores in the rTMS group was significantly superior compared with the sham group throughout the study (p < .001 at fourth week). CONCLUSIONS: Repetitive TMS at 5 Hz accelerated the onset of action and augmented the response to amitriptyline. PMID- 15652876 TI - Monoamine oxidase A gene promoter variation and rearing experience influences aggressive behavior in rhesus monkeys. AB - BACKGROUND: Allelic variation of the monoamine oxidase A (MAOA) gene has been implicated in conduct disorder and antisocial, aggressive behavior in humans when associated with early adverse experiences. We tested the hypothesis that a repeat polymorphism in the rhesus macaque MAOA gene promoter region influences aggressive behavior in male subjects. METHODS: Forty-five unrelated male monkeys raised with or without their mothers were tested for competitive and social group aggression. Functional activity of the MAOA gene promoter polymorphism was determined and genotypes scored for assessing genetic and environmental influences on aggression. RESULTS: Transcription of the MAOA gene in rhesus monkeys is modulated by an orthologous polymorphism (rhMAOA-LPR) in its upstream regulatory region. High- and low-activity alleles of the rhMAOA-LPR show a genotype x environment interaction effect on aggressive behavior, such that mother-reared male monkeys with the low-activity-associated allele had higher aggression scores. CONCLUSIONS: These results suggest that the behavioral expression of allelic variation in MAOA activity is sensitive to social experiences early in development and that its functional outcome might depend on social context. PMID- 15652877 TI - Reduced size and abnormal asymmetry of parietal cortex in women with borderline personality disorder. AB - BACKGROUND: Evidence is accumulating that suggests borderline personality disorder (BPD) and posttraumatic stress disorder (PTSD) are related to small hippocampal size. Psychotic symptoms are frequent in both disorders. Psychotic spectrum disorders are known to be related to abnormalities of temporoparietal cortices. METHODS: Using structural magnetic resonance imaging (3D-MRI), parietal cortex and hippocampal volumes were assessed in 30 young women with BPD who had been exposed to severe childhood sexual and physical abuse and in 25 healthy control subjects. RESULTS: Compared with control subjects, BPD subjects had significantly smaller right parietal cortex (-11%) and hippocampal (-17%) volumes. The parietal cortex of borderline subjects showed a significantly stronger leftward asymmetry when compared with control subjects. Stronger psychotic symptoms and schizoid personality traits in borderline subjects were significantly related to reduced leftward asymmetry. Stronger trauma-related clinical symptoms and neuropsychologic deficits were significantly related to smaller hippocampal size. CONCLUSIONS: Our results are consistent with previous findings of small hippocampal size in BPD and PTSD. Reduced right parietal cortex size in individuals with BPD may reflect a neurodevelopmental deficit of the right hemisphere. PMID- 15652878 TI - Sustained attention-deficit confirmed in euthymic bipolar disorder but not in first-degree relatives of bipolar patients or euthymic unipolar depression. AB - BACKGROUND: Cognitive dysfunction persists in the euthymic phase of bipolar disorder and may provide a marker of underlying neuropathology and disease vulnerability. This study aimed to replicate a deficit in sustained attention in euthymic bipolar patients and investigate sustained attention in first-degree relatives of bipolar probands and in remitted patients with major depressive disorder. METHODS: The rapid visual information processing (RVIP) task was used to measure sustained attention in 15 euthymic patients with bipolar disorder and 15 control subjects in experiment 1 and in 27 first-degree relatives of bipolar probands, 15 remitted patients with major depressive disorder, and 46 control subjects in experiment 2. RESULTS: Sustained attention deficit was confirmed in the euthymic bipolar patients in experiment 1, but the deficit was not statistically significant in remitted major depressed patients or in the relatives of bipolar probands. CONCLUSIONS: A deficit of sustained attention is not present in patients with recurrent major depression tested during remission nor is it discriminable in the first-degree relatives of bipolar probands. Thus, the confirmed abnormality in euthymic bipolar patients may be acquired as a consequence of bipolar illness. However, future studies of relatives will require larger sample sizes to exclude or utilize small genetic effects. PMID- 15652879 TI - Slow transcranial magnetic stimulation can rapidly reduce resistant auditory hallucinations in schizophrenia. AB - BACKGROUND: Almost a quarter of patients with schizophrenia present with resistant auditory verbal hallucinations (AVH), a phenomenon that may relate to activation of brain areas underlying speech perception. Repetitive transcranial magnetic stimulation (rTMS) at 1 Hz reduces cortical activation, and recent results have shown that 1-Hz left temporoparietal rTMS may reduce AVH. The aim of this study was to replicate recent data and investigate whether low-frequency rTMS with a high total stimulation number delivered in a shorter 5-day block produces similar benefit. METHODS: Ten right-handed schizophrenia patients with resistant AVH received 5 days of active rTMS and 5 days of sham rTMS (2,000 stimulations per day at 90% of motor threshold) over the left temporoparietal cortex in a double-blind crossover design. The two weeks of stimulation were separated by a 1-week washout period. RESULTS: AVH were robustly improved (56%) by 5 days active rTMS, whereas no variation was observed after sham. Seven patients were responders to active treatment, five of whom maintained improvement for at least 2 months. CONCLUSIONS: These data confirm the efficiency of low frequency rTMS applied to the left temporoparietal cortex, compared with sham stimulation, in reducing resistant AVH. This improvement can be obtained in only 5 days without serious initial adverse events. PMID- 15652880 TI - Locomotor hyperactivity induced by blockade of prefrontal cortical alpha2 adrenoceptors in monkeys. AB - BACKGROUND: Stimulation of alpha(2)-adrenergic receptors (alpha(2)-ARs) in the prefrontal cortex (PFC) has a beneficial effect on working memory and attentional regulation in monkeys. alpha(2)-adrenergic agonists like clonidine and guanfacine have been used experimentally and clinically for the treatment of attention deficit and hyperactivity disorder (ADHD). However, it is unknown if alpha(2)-ARs in the PFC are involved in the neural mechanisms underlying regulation of locomotor activity. METHODS: The alpha(2)-adrenergic antagonist yohimbine was infused bilaterally and chronically into the dorsolateral PFC (dlPFC) in two monkeys, using mini-osmotic pumps. Spontaneous locomotor activity was measured continuously before, during and after drug administration, using an activity monitor. RESULTS: The monkeys exhibited a dramatic increase in motoric activity during infusion of yohimbine into the dlPFC. Similar treatment with saline was without effect. Thus, the locomotor hyperactivity was due to blockade of alpha(2) ARs, not because of nonspecific factors such as cortical damage by drug administration. CONCLUSIONS: The present study suggests that alpha(2)-ARs in the dlPFC are involved in inhibitory control of locomotor activity. PMID- 15652881 TI - Laparoscopic myomectomy for symptomatic uterine myomas. AB - OBJECTIVE: To evaluate the safety, efficacy, and techniques of laparoscopic myomectomy as treatment for symptomatic uterine myomas. DESIGN: Medline literature review and cross-reference of published data. RESULTS: Results from randomized trials and clinical series have shown that laparoscopic myomectomy provides the advantages of shorter hospitalization, faster recovery, fewer adhesions, and less blood loss than abdominal myomectomy when performed by skilled surgeons. Improvements in surgical instruments and techniques allows for safe removal and multilayer myometrial repair of multiple large intramural myomas. Randomized trials support the use of absorbable adhesion barriers to reduce adhesions, but there is no apparent benefit of presurgical use of GnRH agonists. Pregnancy outcomes have been good, and the risk of uterine rupture is very low when the myometrium is repaired appropriately. CONCLUSION(S): Advances in surgical instruments and techniques are expanding the role of laparoscopic myomectomy in well-selected individuals. Meticulous repair of the myometrium is essential for women considering pregnancy after laparoscopic myomectomy to minimize the risk of uterine rupture. Laparoscopic myomectomy is an appropriate alternative to abdominal myomectomy, hysterectomy, and uterine artery embolization for some women. PMID- 15652882 TI - Egg production predicts a doubling of in vitro fertilization pregnancy rates even within defined age and ovarian reserve categories. AB - OBJECTIVE: To examine the age-independent association of ovarian response and IVF outcome in women with normal and abnormal ovarian reserve. DESIGN: Retrospective analysis. SETTING: Academic IVF center. PATIENT(S): Four thousand eight hundred sixty-two consecutive IVF cycles. INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Outcome of IVF was analyzed as a function of ovarian response to controlled ovarian hyperstimulation and ovarian reserve. RESULT(S): The mean patient age was 36.2 +/- 4.5 years. Younger patients and patients with normal ovarian reserve were found to have better implantation and clinical pregnancy rates. Patients with normal ovarian reserve had a higher number of oocytes retrieved, mature oocytes, two-pronuclei embryos, and embryos transferred. A greater number of embryos were transferred for patients with higher ovarian response. Higher clinical pregnancy rates were seen in those patients who had more oocytes retrieved for all patients, regardless of age and ovarian reserve. In fact, clinical pregnancy rates more than doubled for specific patient groups. CONCLUSION(S): In an age-independent fashion, ovarian response is highly predictive of IVF outcome in women with normal and abnormal ovarian reserve. These findings highlight the importance of not solely relying on age when presenting and discussing IVF outcome data and are useful information when helping patients interpret their IVF cycle response. PMID- 15652883 TI - Secretion of human leukocyte antigen-G by human embryos is associated with a higher in vitro fertilization pregnancy rate. AB - OBJECTIVE: To investigate whether secretion of soluble human leukocyte antigen-G (HLA-G) by human embryos is associated with embryo development and IVF pregnancy outcome. DESIGN: Retrospective study. SETTING: In vitro fertilization program affiliated with a university research center. PATIENT(S): Infertile couples attending an IVF program were selected. INTERVENTION(S): Embryo culture conditioned medium (72 hours) from cases in which intracytoplasmic sperm injection was used for fertilization. MAIN OUTCOME MEASURE(S): Soluble HLA-G in embryo culture medium samples from IVF patients was assayed and associations between soluble HLA-G secretion and outcome measures were analyzed. RESULT(S): Two hundred seventy of 386 samples had detectable soluble HLA-G. Soluble HLA-G secretion was independent of embryo grade or patients' age. The cleavage rate of embryos secreting soluble HLA-G was significantly higher than that of those lacking it (blastomere number 6.71 +/- 0.09 vs. 5.86 +/- 0.22). The live birth rate from embryos with soluble HLA-G was significantly higher than that of those without (48.4% vs. 17.1%, chi(2) = 9.09). Combining soluble HLA-G detection and cleavage rate was most predictive of pregnancy. CONCLUSION(S): Our five conclusions are as follows: [1] embryonic secretion of soluble HLA-G protein is variable, [2] secretion of HLA-G is correlated with a higher cleavage rate, [3] secretion of HLA-G is associated with a higher pregnancy rate, [4] HLA-G secretion is a better independent predictor than cleavage rate alone, and [5] the combination of soluble HLA-G detection and high cleavage rate was the best predictor of outcome. PMID- 15652884 TI - Taking a basal follicle-stimulating hormone history is essential before initiating in vitro fertilization. AB - OBJECTIVE: To analyze IVF outcomes in patients with a history of one or more elevations in basal FSH who have a normal basal FSH at the start of their IVF cycle, compared with the general IVF population. DESIGN: Retrospective clinical study. SETTING: University hospital. PATIENT(S): General IVF patient population. INTERVENTION(S): Patients received standard IVF gonadotropin protocols, oocyte retrieval, and embryo transfer. MAIN OUTCOME MEASURE(S): Oocyte yield, fertilization, implantation, clinical pregnancy, and cancellation rate. RESULT(S): Oocyte yields were lower in patients with a history of elevated basal FSH, for all age groups, and showed an age-dependent decline in all patients. Over the age of 40 years, both implantation and clinical pregnancy rates were lower in these patients, with no significant difference observed in patients under the age of 40 years. No pregnancies were observed in patients with a history of three or more elevated FSH levels, regardless of age. CONCLUSION(S): A history of elevated basal FSH levels in patients under the age of 40 years predicts a lower oocyte yield in IVF cycles with normal basal FSH levels but does not translate to either lower pregnancy or implantation rates. Patients aged >40 years with prior elevations in basal FSH levels have both compromised ovarian response and compromised embryo quality relative to those with normal FSH levels, as illustrated by lower oocyte yield, higher cancellation rates, and lower implantation and pregnancy rates. PMID- 15652885 TI - Follicular phase serum levels of luteinizing hormone do not influence delivery rates in in vitro fertilization cycles down-regulated with a gonadotropin releasing hormone agonist and stimulated with recombinant follicle-stimulating hormone. AB - OBJECTIVE: To assess the value of serum LH measurements in early and late follicular phase as predictors of ovarian response and IVF outcome in patients treated with recombinant FSH with GnRH agonist (GnRH-a) pituitary down regulation. DESIGN: Retrospective cohort analysis. SETTING: Institutional. PATIENT(S): Women undergoing 157 consecutive IVF cycles suppressed with leuprolide acetate (LA) started in the midluteal phase and stimulated with recombinant FSH. Only women <40 years of age and with a basal cycle day 3 serum FSH 98% E(2) decline; however, the difference did not reach statistical significance. CONCLUSION(S): Multifactorial analysis refutes the negative role of supraphysiologic levels of E(2) on the day of hCG administration or its dramatic decline at the midluteal phase on the success rate after embryo transfer. A possibly increased rate of early spontaneous abortion in the high-response group warrants further verification. PMID- 15652888 TI - Screening practices and beliefs of assisted reproductive technology programs. AB - OBJECTIVE: To explore assisted reproductive technology (ART) programs' beliefs about and practices for screening program candidates for the use of ART services. DESIGN: An anonymous, self-administered, mailed questionnaire. SETTING: U.S. ART programs. PARTICIPANT(S): Directors of U.S. ART programs. MAIN OUTCOME MEASURE(S): Screening practices and beliefs, agreement with statements about screening rights and responsibility, information collected about candidates, and likelihood of turning away hypothetical candidates. RESULT(S): The majority of programs do not have a formal policy for screening candidates. The majority of program directors agree that they have a right and responsibility to screen candidates. On average, programs turn away 4% of candidates each year. The majority of programs report being very to extremely likely to deny treatment to the couples described in various scenarios, such as physical abuse, positive HIV status, and single parenthood. Significant variation was seen across programs in their likelihood of turning away various hypothetical candidates. CONCLUSION(S): There is substantial variation in ART programs' screening practices. These results highlight the need for increased debate over what constitutes inappropriate denial of access to services, and what are prudent, social, ethical, and medical judgments. PMID- 15652889 TI - Stress reduction in male infertility patients: a randomized, controlled trial. AB - OBJECTIVE: To assess the impact of preparatory information about the fertility workup on the patients' well-being. DESIGN: Two-group, randomized controlled study. SETTING: An andrology clinic. PATIENT(S): Two hundred fifty men enrolled for fertility workup. INTERVENTION(S): A two-page leaflet with preparatory information about the fertility workup, which was mailed to half of the participants after they had made an appointment. MAIN OUTCOME MEASURE(S): Questionnaire score for infertility-related distress at clinic attendance, proportion of participants that still had not attended 6 months after the scheduled appointment. RESULT(S): Distress scores and the proportion of nonattendees were significantly reduced in the group receiving the leaflet. An additional analysis revealed that 55% of the receivers did not know that the andrology clinic has its own web site, which was mentioned in the leaflet several times. CONCLUSION(S): Although it is uncertain how many patients actually read a routinely sent leaflet, preparatory information in written form is beneficial, at least for a significant subgroup of men who consider undergoing fertility workup. Fertility services might reduce the number of nonattendees by sending out leaflets. PMID- 15652890 TI - Semen profile, testicular volume, and hormonal levels in infertile patients with varicoceles compared with fertile men with and without varicoceles. AB - OBJECTIVE: To assess semen analysis, testicular volume, and hormone levels in fertile and infertile patients with varicoceles and fertile men without varicoceles. DESIGN: Retrospective study. SETTING: Academic medical center. PATIENT(S): Patients were divided into three groups: fertile men with varicoceles (n = 79), infertile men with varicoceles (n = 71), and fertile men without varicoceles (n = 217). INTERVENTION(S): None. MAIN OUTCOME MEASURE(S): Levels of LH, FSH, and total T and testicular volume in fertile and infertile men with varicoceles and fertile controls without varicoceles. RESULT(S): LH (IU/L) and T (ng/dl) levels were not statistically different across the three groups. FSH levels were significantly higher in infertile men with varicoceles (7.8 +/- 7.6 IU/L) than in the fertile men with varicoceles (3.5 +/- 2.1 IU/L) or in fertile men without varicoceles (3.5 +/- 1.9 IU/L). The right testicle was smaller in infertile patients with varicoceles (18.7 +/- 8.3 cm(3)) than in fertile men with varicoceles (25.2 +/- 13 cm(3)) or in fertile men without varicoceles (24.9 +/- 10.7 cm(3)). In addition, the left testicle was smaller in infertile men with varioceles (17.6 +/- 8.9 cm(3)) than in fertile men with varicoceles (21.6 +/- 7.8 cm(3)) or in fertile men without varicoceles (23.4 +/- 8.3 cm(3)). Sperm concentration was lower in infertile men with varicoceles (33.7 +/- 23.3 x 10(6)/mL) than in fertile men with varicoceles (101.8 +/- 76.6 x 10(6)/mL) or in fertile men without varicoceles (111.8 +/- 74.2 x 10(6)/mL). In addition, sperm motility was lower in infertile men with varicoceles (37.2% +/- 23.9%) than in fertile men with varicoceles (53.9% +/- 17.4%) or fertile men without varicoceles (58.9% +/- 15.8%). CONCLUSION(S): Infertile patients with varicoceles have higher levels of FSH, smaller testes, and lower sperm concentration and motility compared with controls with or without varicoceles. No statistical differences were seen in the variables evaluated among the fertile men with incidental varicoceles detected at physical examination and those without varicoceles. PMID- 15652891 TI - Tissue immunoexpression and messenger ribonucleic acid localization of inhibin/activin subunit in human epididymis. AB - OBJECTIVE: To determine the expression of inhibin betaA and betaB subunits and follistatin and the ability of human epididymal epithelium to synthesize these molecules. DESIGN: The main aim of this study was to investigate the expression of inhibin alpha, betaA, and betaB-subunits and follistatin in human epididymis with immunohistochemistry, in situ hybridization, and Western blotting in adult life. SETTING: Academic university hospital. PATIENT(S): Epididymes were obtained from 10 men undergoing routine vasectomy or surgery for benign disease at the Royal Hallamshire Hospital, Sheffield, United Kingdom. MAIN OUTCOME MEASURE(S): Immunoexpression of activin betaA and betaB subunits and follistatin proteins and mRNA in human caput and cauda epididymis. RESULT(S): Positive immunoexpression for activin betaA and betaB subunits and follistatin were detected in different parts of the epididymis epithelium. Western blotting under a reducing condition detected a 28-kd band (possibly corresponding to the activin dimer). In situ hybridization indicated positive mRNA localization signal in both caput and cauda epididymal epithelium. CONCLUSION(S): Activins betaA and betaB subunits, but not inhibin alpha subunit, were detected in epididymal epithelium. These finding suggest that activins might have a role in the processes of sperm maturation and sperm fertilizing capability during transit and storage. PMID- 15652892 TI - Effect of experimental varicocele on the expressions of Notch 1, 2, and 3 in rat testes: an immunohistochemical study. AB - OBJECTIVE: To study expressions of Notch receptor isoforms (Notch 1, 2, and 3) in normal and varicocele-induced rat testes to examine their possible functions in cell fate. DESIGN: Comparative and controlled study. SETTING: Animal Care and Operation Unit, Akdeniz University. ANIMAL(S): Wistar male rats for experimental and control groups. INTERVENTION(S): The control group underwent a sham operation (n = 6). The experimental groups underwent partial ligation of the renal vein to induce an experimental varicocele and then were killed 9 (n = 6), 11 (n = 6), and 13 (n = 6) weeks after the induction of varicocele. MAIN OUTCOME MEASURE(S): All tissues were fixed and routinely processed for paraffin embedding. Subsequent immunohistochemical studies were performed. RESULT(S): In the sham-operation rat testes, Leydig cells and elongated spermatids were immunopositive for Notch 1. Notch-2 expression was present in Leydig cells, spermatogonia, and primary spermatocytes. Notch-3 expression was limited to Leydig cells. Varicocele formation diminished the expression of both Notch-1 and Notch-2 receptors as the varicocele formation progressed over time. CONCLUSION(S): The present study suggests that Notch 1 is related to the maturation of spermatids. Notch 2 is related to both proliferation and maturation of spermatogenic cells, whereas Notch 3 seems to be related to Leydig cell functions. The decrease of both Notch 1 and Notch-2 expression depended on the degree of varicocele development over time, indicating a potential role in varicocele-associated testicular dysfunction. PMID- 15652893 TI - Impact of sperm morphology on DNA damage caused by oxidative stress induced by beta-nicotinamide adenine dinucleotide phosphate. AB - OBJECTIVE: To investigate the role of DNA damage induced by beta-nicotinamide adenine dinucleotide phosphate (NADPH) in human spermatozoa. DESIGN: Prospective controlled study. SETTING: Male infertility clinic at the Glickman Urological Institute, Cleveland Clinic Foundation, Cleveland, Ohio. PATIENT(S): Twenty-eight men undergoing infertility screening. INTERVENTION(S): Chemiluminescence assay and terminal deoxynucleotidyl transferase-mediated digoxigenin-dUTP nick-end labeling (TUNEL) assay coupled flow cytometry after incubating mature and immature sperm separated by density gradient with 5 mM NADPH for 0, 3, and 24 hours. MAIN OUTCOME MEASURE(S): Reactive oxygen species (ROS) generation (10(6) counted photons per minute/10(6) sperm) and percentage of spermatozoa with fragmented DNA. RESULT(S): Immature sperm from teratozoospermic semen samples were characterized by a statistically significant presence of cytoplasmic residues in the mid-piece when compared with mature normozoospermic samples. Increased ROS production was observed in spermatozoa rich in cytoplasmic residues that showed a statistically significant positive correlation with sperm DNA damage in a time-dependent manner. CONCLUSION(S): Immature sperm contain high nicotinamide adenine dinucleotide phosphate (NADPH) in cytoplasmic droplets, but it has not yet been clear whether abnormal sperm morphology plays any role in sperm DNA damage induced by oxidative stress. Our data support the role of NAPDH in ROS-mediated sperm DNA damage and suggest that abnormal sperm morphology combined with elevated ROS production may serve as a useful indicator of potential damage to sperm DNA. PMID- 15652894 TI - Acrosin activity as a potential marker for sperm membrane characteristics in unexplained male infertility. AB - OBJECTIVE: To evaluate sperm membrane system integrity in unexplained infertile male subjects with three consecutive conception failures on IUI even though semen clinical parameters were normal. DESIGN: Prospective study. SETTING: Medical biotechnology laboratory, School of Medical Science and Technology IIT Kharagpur, India. PATIENT(S): Twenty-nine patients with unexplained infertility, 17 normal proven-fertile healthy donors, and 21 infertile males with low motility but with other semen parameters remaining normal. INTERVENTION(S): Semen samples were collected from unexplained infertile patients as well as from healthy fertile donors after abstinence of 3-5 days and were analyzed according to World Health Organization guidelines. MAIN OUTCOME MEASURE(S): Release of 5'-nucleotidase (plasma membrane marker), lactate dehydrogenase (mitochondrial marker) and free acrosin, proacrosin, and total acrosin (acrosomal membrane marker). RESULT(S): Plasma membrane integrity and respiratory activity of sperm cells were comparable in all three groups. The proacrosin-acrosin system was adversely affected in unexplained infertile subjects despite high sperm motility. CONCLUSION(S): Total acrosin activity may be considered as a sensitive biochemical marker for clinical evaluation of unexplained infertility in males. PMID- 15652895 TI - Hormonal profile and endometrial morphology in letrozole-controlled ovarian hyperstimulation in ovulatory infertile patients. AB - OBJECTIVE: To evaluate the clinical response and endometrial morphology during the implantation window on ovarian hyperstimulation with the aromatase inhibitor letrozole in infertile ovulatory women. DESIGN: Prospective trial in infertile patients. SETTING: Tertiary care hospital. PATIENT(S): Eight ovulatory infertile patient candidates for ovarian superovulation. INTERVENTION(S): Subjects were monitored in one control cycle. In the next cycle, they received letrozole 5.0 mg daily on days 3 through 7 after menses. MAIN OUTCOME MEASURE(S): Number of ovulatory follicles; dominant follicle diameter; endometrial thickness; hormonal profile of FSH, LH, E(2), A, T, and P; endometrial histological dating; and pinopode formation assessed by scanning electron microscopy. RESULT(S): Cycles stimulated with letrozole resulted in more ovulatory follicles than did natural cycles (mean +/- SD 2.0 +/- 0.9 vs. 1.0 +/- 0.0), which attained a greater preovulatory diameter (mean +/- SD 23.8 +/- 2.7 vs. 19.3 +/- 2.1 mm), with similar endometrial thickness at midcycle compared with spontaneous cycles. Endocrine profile of medicated cycles was characterized on day 7 by increased levels of LH (5.9 +/- 0.8 vs. 3.5 +/- 0.4 IU/mL), reduced E(2) (98.4 +/- 11.4 vs. 161.5 +/- 14.7 pmol/L), and elevated androgens. Preovulatory and midsecretory E(2) were similar to spontaneous cycle, and P levels during midluteal phase were significantly elevated (44.2 +/- 4.6 vs. 27.7 +/- 4.6 pmol/L). Endometrial morphology during the implantation window in letrozole-stimulated cycles was characterized by in-phase histological dating and pinopode expression on scanning electron microscopy. CONCLUSION(S): Letrozole induces moderate ovarian hyperstimulation in ovulatory infertile patients with E(2) levels similar to spontaneous cycles and higher midluteal P, leading to both a normal endometrial histology and development of pinopodes, considered to be relevant markers of endometrial receptivity. PMID- 15652896 TI - Use of a prediction model for high-order multiple implantation after ovarian stimulation with gonadotropins. AB - OBJECTIVE: To determine prospectively the effectiveness in clinical practice of a prediction model for high-order multiple pregnancies (HOMP) (triplets or more). DESIGN: Prospective study. SETTING: University teaching hospital. PATIENT(S): Eight hundred forty-nine consecutive infertile patients undergoing a total of 1,542 treatment cycles. INTERVENTION(S): Gonadotropin ovarian stimulation or induction of ovulation without IVF MAIN OUTCOME MEASURE(S): Observed and predicted overall pregnancy rates and the incidence of HOMP. RESULT(S): The use of the prediction model (implying cancellation of all cycles at high risk for HOMP) would result in an 8% (95% confidence interval, 6.8%-9.2%) reduction of overall pregnancy rate but also in a 285% (95% CI, 279%-291%) reduction of HOMP. CONCLUSION(S): By using our prediction model, it was possible to maintain a low risk of HOMP with a good pregnancy rate in patients receiving gonadotropin ovarian stimulation or induction of ovulation without IVF. PMID- 15652897 TI - A role for the fibrinolytic system in postsurgical adhesion formation. AB - OBJECTIVE: To look for evidence of a fibrinolytic insufficiency as a cause of adhesion formation. DESIGN: Retrospective and prospective study. SETTING: University medical center. PATIENT(S): Retrospective study: 50 patients undergoing laparoscopy, divided into patients with and without endometriosis. Prospective study: 18 patients undergoing infertility surgery involving a second look laparoscopy. INTERVENTION(S): During all surgical procedures, adhesions were scored, and peritoneal fluid and plasma were collected. MAIN OUTCOME MEASURE(S): Parameters of the fibrinolytic system were measured to establish a possible relation with the presence and formation of adhesions. RESULT(S): In patients with endometriosis and adhesions, significantly higher peritoneal fluid concentrations were found for plasminogen activator inhibitor-1 (PAI-1), tissue plasminogen activator (tPA), and plasminogen, compared with patients with endometriosis but without adhesions. In the prospective study, initial peritoneal PAI-1 concentrations correlated significantly with the extent of adhesion formation (r(s) = 0.49) and adhesion-improvement scores (r(s) = -0.52). Also, the change in concentration of tPA and fibrinogen from the initial surgical procedure to the second-look laparoscopy correlated significantly with adhesion-improvement scores (DeltatPA: r(s)= 0.50; Deltafibrinogen: r(s) = -0.64). CONCLUSION(S): This first prospective study in humans adds further weight to the hypothesis that adhesions are caused by an insufficiency in peritoneal fibrinolytic activity. Plasminogen activator inhibitor-1 is a potential marker for the identification of patients at risk for developing adhesions. PMID- 15652898 TI - Metformin reduces serum mullerian-inhibiting substance levels in women with polycystic ovary syndrome after protracted treatment. AB - OBJECTIVE: Assessment of ovarian responses to metformin treatment in obese women with polycystic ovary syndrome (PCOS). DESIGN: Prospective treatment with randomization to two doses of metformin. SETTING: University teaching hospital. PATIENT(S): Obese women (n = 82) with PCOS. INTERVENTION(S): Markers of ovarian function were assessed after 4 and 8 months. MAIN OUTCOME MEASURE(S): Hormone (androgens and mullerian-inhibiting substance [MIS]) changes over time. RESULT(S): There was no difference in the reproductive hormone changes between the doses of metformin, and data were combined for further analyses. Significant responses to treatment were recorded for menstrual frequency and androstenedione (A) (reduction) within the first 4 months of treatment. However, suppression of the elevated circulating MIS concentrations required protracted treatment, because no change was observed in the first 4 months-only in the second 4-month assessment period. CONCLUSION(S): Metformin treatment of PCOS leads to rapid suppression of A and improved menstrual frequency. Suppression of MIS is a delayed response that may be secondary to the development of a cohort of follicles that underwent initial recruitment in an environment of reduced insulin stimulation. PMID- 15652899 TI - Lack of effect of isoflavonoids on the vagina and endometrium in postmenopausal women. AB - OBJECTIVE: To determine the effects of soy-derived isoflavones on vaginal epithelium and the endometrium. DESIGN: Double-blind, randomized, placebo controlled crossover trial. SETTING: Outpatient clinic of a university hospital. PATIENT(S): Sixty-four postmenopausal women with a history of breast cancer. INTERVENTION(S): The women took (in a randomized order) 114 mg of isolated isoflavonoids or placebo in tablets daily for 3 months; the treatment regimens were crossed over after a 2-month washout period. The subjects were studied before and on the last day of each treatment period. MAIN OUTCOME MEASURE(S): Vaginal dryness, maturation index (MI) of vaginal epithelium, endometrial thickness, histology, and expression of estrogen (E) and progesterone (P) receptors and the proliferation marker Ki-67 in the endometrium. RESULT(S): Isolated isoflavones did not relieve vaginal dryness. Maturation index values remained unchanged during the isoflavone regimen, but decreased during the placebo regimen. No changes were found in any of the variables measured in the endometrium. CONCLUSION(S): Daily administration of 114 mg of isolated isoflavones for 3 months had no effect on the subjective perception of vaginal dryness or on objective findings in the vagina or endometrium. This implies safety with regard to the endometrium. PMID- 15652900 TI - "Kissing ovaries": a sonographic sign of moderate to severe endometriosis. AB - OBJECTIVE: To evaluate whether the presence of kissing ovaries at ultrasound is a marker for endometriosis and whether it correlates with the severity of the disease. DESIGN: Prospective observational study. SETTING: Gynecologic departments of two university hospitals. PATIENT(S): A total of 722 consecutive premenopausal women who had laparoscopic surgery for an adnexal mass or suspected pelvic endometriosis. INTERVENTION(S): Preoperative ultrasound evaluation and laparoscopic surgery. MAIN OUTCOME MEASURE(S): Diagnostic and predictive value of ultrasound identification of kissing ovaries in the detection of endometriosis. RESULT(S): Kissing ovaries were diagnosed at ultrasound and confirmed laparoscopically in 32 patients. Of these, 27 had moderate to severe endometriosis and five others had benign adnexal masses. Bowel (18.5% vs. 2.5%) and fallopian tube (92.6% vs. 33%) endometriosis were significantly more frequent in patients with kissing ovaries than in patients without kissing ovaries. In infertile patients (n = 145), kissing ovaries were associated with a higher proportion of women with fallopian tube obstruction (80% vs. 8.6%). Considering patients with moderate to severe endometriosis (n = 189), the median (range) revised American Fertility Society score (74 [32-148] vs. 35 [16-146]) and the operative time (115 minutes [65-245 minutes] vs. 50 [15-180 minutes]) were significantly higher in patients with than in those without kissing ovaries. CONCLUSION(S): The detection of kissing ovaries at ultrasound is strongly associated with the presence of endometriosis and is a marker of the most severe form of this disease. PMID- 15652901 TI - Sensitivity to stress-induced reproductive dysfunction linked to activity of the serotonin system. AB - OBJECTIVE: To use a nonhuman primate model and determine whether individuals sensitive to stress-induced reproductive dysfunction have lower activity of central serotonergic neurons under nonstressed conditions. DESIGN: The activity of the central serotonergic system was assessed by measuring responsiveness to a fenfluramine challenge (5 mg/kg, IV) in sedated monkeys previously categorized as highly stress resistant (HSR; n = 4; normal menstrual cyclicity through two stress cycles), medium stress resistant (MSR; n = 5; ovulatory in the first stress cycle but anovulatory in the second stress cycle), or low stress resistant (i.e., stress sensitive, SS; n = 4; anovulatory as soon as stress is initiated). To control for differences in pituitary stores of prolactin or ACTH, the animals were subsequently administered a bolus of thyrotropin-releasing hormone (3 microg/kg) plus corticotropin releasing factor (CRF), (3 microg/kg). SETTING: Oregon National Primate Research Center, Animal Services Building. PATIENT(S): Female cynomolgus macaques exhibiting normal menstrual cycles. INTERVENTION(S): Administration of fenfluramine, a serotonin-releasing drug. MAIN OUTCOMES MEASURE(S): Serum concentrations of prolactin (PRL) and cortisol (F). RESULT(S): Prolactin release in response to fenfluramine was significantly greater in the HSR group compared with the MSR or SS groups. In contrast, cortisol was higher in the SS group compared with the other two groups. Similar responses were not evident after thyrotropin-releasing hormone + CRF stimulation. CONCLUSION(S): The lower PRL response to fenfluramine in the stress-sensitive animals suggests that stress-sensitive individuals have decreased activity in central serotonergic neurons. However, the F data suggest that the hypothalamic-pituitary-adrenal axis in stress-sensitive individuals is highly responsive to even small increases in serotonin. PMID- 15652902 TI - Distribution of cyclooxygenase-1, cyclooxygenase-2, and cytosolic phospholipase A2 in the luteal phase human endometrium and ovary. AB - OBJECTIVE: To identify the distribution of the enzymes cyclooxygenase-1 (COX-1), cyclooxygenase-2 (COX-2), and cytosolic phospholipase A(2) (cPLA(2)) in the human ovary and endometrium. DESIGN: Prospective clinical study. SETTING: Hospital based unit for reproductive health and research laboratories. PATIENT(S): Twenty nine healthy fertile women with normal menstrual cycles. INTERVENTION(S): Endometrial and ovarian biopsy samples were obtained from healthy, fertile women in the luteal phase of the menstrual cycle or during caesarean section. MAIN OUTCOME MEASURE(S): Pinopode formation and immunohistochemical staining of cPLA(2), COX-1, and COX-2. RESULT(S): In the endometrium, the immunostaining of cPLA(2) was most intense in the luminal epithelium when pinopodes were present. The staining of both COX-1 and COX-2 was most intense in the epithelial cells, with the stroma staining positive only for COX-2. The endometrial vessels expressed COX-2 but not COX-1. The staining of COX-1 and COX-2 was intense on the surface epithelial cells on the outer lining of the ovary. CONCLUSION(S): This study details the distribution of these prostaglandin synthase enzymes and emphasizes their importance for the functions of both the endometrium and the ovary. PMID- 15652903 TI - Expression of the type I interferon receptor and the interferon-induced Mx protein in human endometrium during the menstrual cycle. AB - OBJECTIVE: To investigate the expression of the type I interferon receptor (IFNAR) and interferon-induced Mx protein (Mx) in normal human endometrium throughout the menstrual cycle. DESIGN: Prospective study. SETTING: Medical university in Japan. PATIENT(S): Thirty-seven normal endometrial tissues from fertile women who had undergone hysterectomies for reasons other than endometrial disease. INTERVENTION(S): IFNAR-1, IFNAR-2, MxA, and MxB gene expression was analyzed by reverse transcription-real-time quantitative polymerase chain reaction. Moreover, localization of IFNAR-1 and IFNAR-2 were studied by immunohistochemistry. MAIN OUTCOME MEASURE(S): Expression of IFNAR-1, IFNAR-2, MxA, and MxB. RESULT(S): Expression of IFNAR-2 gene was significantly increased in the menstrual and midsecretory phase as compared with in the proliferative phase. Immunohistochemistry for IFNAR-1 and IFNAR-2 revealed weak staining of glandular epithelium and weak staining of stromal cells during the proliferative phase. However, an intense immunohistochemical staining of IFNAR-2 was observed on the surface and basement membrane of glands in the secretory phase. There was no statistical difference between MxA and MxB gene expression throughout the menstrual cycle. CONCLUSION(S): Our results suggest that IFNAR and Mx are expressed in the human endometrium and that the expression of IFNAR is cyclically changed during the menstrual cycle. PMID- 15652904 TI - Nonsteroidal antiinflammatory drugs differentially suppress endometriosis in a murine model. AB - OBJECTIVE: To determine whether nonsteroidal antiinflammatory drugs (NSAIDs) affect the establishment and progression of endometriotic lesions in a murine model. DESIGN: Pharmacologic intervention in a surgically induced murine model of abdominal/peritoneal endometriosis. SETTING: Animal research facility. PATIENT(S): Eight-week-old, female C57BL/6 mice. INTERVENTION(S): After implantation of autologous endometrium, mice were randomized into groups and treated with one of several NSAIDs or the vehicle-matched control for 4 weeks. MAIN OUTCOME MEASURE(S): Establishment, growth, and total burden of endometriotic lesions. RESULT(S): The NSAIDs differentially inhibited lesion establishment and growth, resulting in significantly reduced disease burden. Compared with controls (5.7 +/- 2.3 mm(2)), lesion burden was reduced by celecoxib (1.3 +/- 1.2 mm(2)), indomethacin (1.4 +/- 1.4 mm(2)), naproxen (2.7 +/- 1.2 mm(2)), sulindac (3.1 +/- 1.5 mm(2)), rofecoxib (3.4 +/- 3.0 mm(2)), and ibuprofen (4.1 +/- 1.4 mm(2)). In contrast, aspirin (5.9 +/- 1.2 mm(2)) had no statistically significant effect. Uninterrupted estrus cycling was confirmed by vaginal exams and smears in celecoxib-treated mice. CONCLUSION(S): Chronic administration of certain NSAIDs limits the progression of endometriosis in this murine model. The data suggest that NSAID selection in the treatment of endometriosis should be extended beyond pain management to maximize the inhibitory effect on disease burden. PMID- 15652905 TI - Changes in the freezing protocol for human zygotes alter embryonic development and pregnancy rates. AB - OBJECTIVE: To determine the effect of various freezing protocols on postthaw development and pregnancy rates resulting from transfer of human zygotes. DESIGN: Prospective study. SETTING: Tertiary care center. PATIENT(S): Couples undergoing assisted reproductive technology (ART) procedures who wished to have their excess zygotes cryopreserved. INTERVENTION(S): We cryopreserved zygotes with one of three protocols. MAIN OUTCOME MEASURE(S): Post-thaw survival and development of the zygotes as well as pregnancy rate after transfer of these zygotes. RESULT(S): A 3-minute hold time after seeding, followed by a final preplunging temperature of -180 degrees C, resulted in a clinical pregnancy rate of 28.6%. In contrast, a 15-minute postseed hold time and a -30 degrees C final chamber temperature resulted in a 37.3% clinical pregnancy rate. When we combined the protocols to provide a 15-minute postseed holding time and a -180 degrees C before plunging into liquid nitrogen, we achieved a 69.6% clinical pregnancy rate. CONCLUSION(S): By increasing the postseeding hold time and decreasing the temperature of the freezing chamber before plunging the zygotes into liquid nitrogen, significant improvements can be made in postthaw development and pregnancy rates. PMID- 15652906 TI - Intrauterine instillation of trichloroacetic acid is effective for the treatment of dysfunctional uterine bleeding. AB - OBJECTIVE: To evaluate the effectiveness of trichloroacetic acid (TCA) instillation into uterine cavity for the treatment of dysfunctional uterine bleeding (DUB). DESIGN: Prospective clinical study. SETTING: A university research hospital. PATIENT(S): Ninety women participated who had dysfunctional uterine bleeding. INTERVENTION(S): Ninety-five percent of TCA was instilled into uterine cavity for endometrial ablation in women with dysfunctional uterine bleeding who want conservative treatment. Participants in group 1 received only TCA; participants in group 2 received a single dose of gonadotropin-releasing hormone analogue 1 month before the procedure. MAIN OUTCOME MEASURE(S): All participants underwent an evaluation that included cycle history, body mass index measurement, and transvaginal ultrasonography of pelvis, diagnostic hysteroscopy and endometrial biopsy. RESULT(S): At the end of 12 months of the treatment, amenorrhea rates in group 1 and group 2 were 26.7% vs. 31.1%, with pooled amenorrhea, hypomenorrhea, and eumenorrhea rates of 95.6% vs. 97.8%, respectively. There was no significant difference between the groups vis-a-vis postprocedure results. More than 90% of women who have this procedure are satisfied with the results. There were no observed negative effects or related complications with this treatment. CONCLUSION(S): An instillation of TCA into uterine cavity produces acceptable results and provides conservative management of DUB. PMID- 15652907 TI - Ascites and pelvic masses: an unusual case of endometriosis. AB - We describe an unusual case of ascites caused by the rupture of a large endometrioma. PMID- 15652908 TI - Live birth of a normal healthy baby after a frozen embryo transfer with blastocysts that were frozen and thawed twice. AB - OBJECTIVE: To report the birth of a normal healthy baby after a frozen ET (FET) with blastocysts that were frozen and thawed twice. DESIGN: Case report. SETTING: Private infertility practice. PATIENT(S): A 26-year-old female who presented with male factor infertility and polycystic ovarian disease. INTERVENTION(S): One cycle of IVF-ET and FET no. 1 followed by the refreezing of blastocysts and FET no. 2. MAIN OUTCOME MEASURE(S): Clinical pregnancy and live birth. RESULT(S): Thirty-six months after a successful IVF-ET cycle, FET no. 1 was performed with eight frozen blastocysts that were thawed. One blastocyst was transferred to the uterus and four were refrozen. The first FET resulted in a singleton pregnancy that ended in a spontaneous abortion after 7 weeks' gestation. Six months later, FET no. 2 was performed. All four refrozen blastocysts were thawed and transferred to the uterus. The second FET resulted in a singleton pregnancy and the birth of a normal healthy baby (male) weighing 3,005 g after 38 weeks' gestation. CONCLUSION(S): Human blastocysts can be refrozen/thawed and produce a normal healthy baby after an FET. Further studies will be required to determine survival, implantation, and live birth rates with refrozen/thawed human blastocysts. PMID- 15652909 TI - A rare case of gonadal agenesis with paramesonephric derivatives in a patient with a normal female karyotype. AB - OBJECTIVE: To report a rare case of gonadal agenesis with rudimentary paramesonephric ducts derivatives in a female with a 46,XX normal karyotype. DESIGN: Case study. SETTING: National Institute of Health. PATIENT(S): An 18-year old female with primary amenorrhea and lack of secondary sexual development. INTERVENTION(S): Clinical, gynecological, endocrine, and genetic evaluation. Laboratory studies conducted included measurement of pituitary, ovary, and thyroid hormones; analyses of G-banded chromosomes in peripheral blood and fibroblast cultures; search for genomic Y-chromosome DNA by fluorescence in situ hybridization and molecular biology techniques; X-ray, ultrasonography, echocardiographic and laparoscopic studies for the assessment of bone age, and genitourinary and other associated malformations. MAIN OUTCOME MEASURE(S): Clinical, hormonal, anatomical, and genetic characteristics of the patient. RESULT(S): The studies performed confirmed a prepubertal female with hypergonadotrophic hypogonadism, bilateral gonadal agenesis, a rudimentary uterus and fallopian tubes, a normal vagina, kidney, and urinary tract structures, and a 46,XX normal karyotype. The search for centromeric Y-chromosome DNA and SRY and ZFY genes was negative. CONCLUSION(S): A primary deficiency confined to the gonadal blastema and the nearby coelomic epithelium is proposed as an alternative embryologic mechanism to explain the occurrence of this singular sexual developmental defect. PMID- 15652910 TI - Ovarian pregnancy resulting from cornual fistulae in a woman who had undergone bilateral salpingectomy. AB - OBJECTIVE: To report a case of ovarian pregnancy following in vitro fertilization embryo transfer (IVF-ET) treatment for which the cornual fistulae was the most probable explanation of the cause. DESIGN: Case report. SETTING: University department and assisted reproduction unit. PATIENT(S): A 29-year-old woman with primary infertility. INTERVENTION(S): Hydrosalpinx of the bilateral fallopian tubes has been noted in patients who have decreased pregnancy rates in the IVF-ET treatment cycles. Salpingectomy before IVF cycles has been suggested to increase the pregnancy rate. We report a patient who presented with bilateral hydrosalpinx for whom bilateral salpingectomy was performed before IVF treatment. MAIN OUTCOME MEASURE(S): Clinical pregnancy. RESULT(S): The treatment cycle resulted in a primary ovarian pregnancy and required laparoscopic operation. A cornual fistulae was found that have might led to ectopic implantation of the embryos. CONCLUSION(S): Ovarian pregnancy may be an unexpected complication of those receiving bilateral salpingectomy before IVF treatment. The presence of cornual fistulae after salpingectomy was probably the cause of the ectopic pregnancy. PMID- 15652911 TI - Spontaneous ovarian hyperstimulation caused by a follicle-stimulating hormone secreting pituitary adenoma. AB - OBJECTIVE: To report a case of spontaneous ovarian hyperstimulation resulting from an FSH-secreting pituitary adenoma. DESIGN: Case report. SETTING: University hospital. PATIENT(S): A 29-year-old previously healthy, nulligravid woman. INTERVENTION(S): Transphenoidal resection of the adenoma. MAIN OUTCOME MEASURE(S): Clinical remission. RESULT(S): After transphenoidal resection of the adenoma, the patient had an uneventful postoperative recovery with complete resolution of ovarian hyperstimulation and associated symptoms. Postoperative magnetic resonance imaging demonstrated residual tumor within the cavernous sinus. CONCLUSION(S): Follicle-stimulating hormone-secreting pituitary adenoma may present as acute spontaneous ovarian hyperstimulation. PMID- 15652912 TI - Ongoing pregnancy in a woman who inadvertently underwent office hysteroscopy during early pregnancy. AB - OBJECTIVE: To report a case of ongoing pregnancy in a woman who underwent hysteroscopy during the implantation phase. DESIGN: Case report. SETTING: A university hospital. PATIENT(S): A 34-year-old woman with unexplained infertility who was scheduled for IVF. INTERVENTION(S): Office hysteroscopy. MAIN OUTCOME MEASURE(S): Hysteroscopy during early pregnancy. RESULT(S): An ongoing pregnancy after hysteroscopy during the implantation phase. CONCLUSION(S): The risk of a properly performed hysteroscopy in the implantation phase of an unrecognized pregnancy may be less than expected. PMID- 15652913 TI - Radiographic and hysteroscopic findings of a placental site nodule. AB - OBJECTIVE: To report a case of placental site nodule discovered on hysterosalpingogram and evaluated by sonohysterography and hysteroscopy in a patient presenting with infertility. DESIGN: Case report. SETTING: Academic reproductive endocrinology center. PATIENT(S): A 30-year-old gravida 1 para 1 receiving evaluation for secondary infertility. INTERVENTION(S): Hysterosalpingogram, sonohysterography, and operative hysteroscopy with excision. MAIN OUTCOME MEASURE(S): Diagnosis and treatment of intrauterine lesion. RESULT(S): Pathology of the excisional biopsy from operative hysteroscopy revealed a placental site nodule. CONCLUSION(S): We report one of the first radiographic and hysteroscopic findings of a placental site nodule. This case demonstrates that in addition to the most common causes of uterine filling defects seen on hysterosalpingogram and sonohysterography, rarer lesions like placental site nodule also need to be considered in the differential diagnosis. Placental site nodule is a benign lesion that requires correct diagnosis to distinguish it from other more serious placental findings, but long-term follow up is unnecessary. Therapy consists of diagnosis and expectant management. PMID- 15652914 TI - A 46,XX SRY-negative man with complete virilization and infertility as the main anomaly. AB - OBJECTIVE: To report a case of a 46,XX SRY-negative man with a male phenotype and azoospermia. DESIGN: Case report. SETTING: Molecular and Cytogenetic Unit in a University Hospital. PATIENT(S): A 35-year-old man with complete masculinization who referred to our institution because of a history of several years of infertility. INTERVENTION(S): Lymphocytic karyotype and genetic counseling. MAIN OUTCOME MEASURE(S): Peripheral blood metaphases were analyzed by standard G banding and Q-banding. Fluorescent in situ hybridization (FISH) and polymerase chain reaction (PCR) analyses were performed. RESULT(S): Semen analysis showed azoospermia. Chromosome analysis revealed a 46,XX karyotype; molecular and cytogenetic analyses excluded the presence of SRY (the sex-determining region of the Y chromosome) gene. CONCLUSION(S): This case is one of the rare patients reported in the literature in whom testicular differentiation and a complete virilization in a 46,XX chromosomal constitution does not account for a translocation of the SRY gene to the X chromosome or to the autosomes. This finding suggests that other genes downstream from SRY, not yet identified, play an important role in sex determination. PMID- 15652915 TI - A uterine cavity-myoma communication after uterine artery embolization: two case reports. AB - OBJECTIVE: To report two cases of uterine cavity-myoma fistula as a medical complication after uterine artery embolization. DESIGN: Case report. SETTING: Women's Medical Center/Viscomi Institute, Sao Paulo, Brazil. PATIENT(S): Two patients presenting with symptomatic myomatous uterus: a 40-year-old woman with increased abdominal volume and hypermenorrhea, and a 42-year-old woman with pelvic pain and infertility. INTERVENTION(S): Uterine artery embolization, hysteroscopic follow-up, and laparoscopic myomectomy. MAIN OUTCOME MEASURE(S): Hysteroscopic follow-up after uterine artery embolization. RESULT(S): The patients underwent uterine artery embolization. In the months that followed, a subsequent surgical hysteroscopic follow-up was performed to eliminate the necrotic material of the degenerated myomas. The two patients presented the same outcome 1 year after the uterine artery embolization had been performed: a communication between the uterine cavity and a degenerated myoma. Laparoscopic correction of the uterine wall defect was performed afterward. CONCLUSION(S): Because embolization is a growing option for the treatment of leiomyoma, it is important that potential complications be reported, especially if the patients want to become pregnant. The natural history of the fistula and its consequences are unknown, and physicians should be aware of these complications. Hysteroscopic follow-up was important in the diagnosis of the two cases. PMID- 15652916 TI - Reversible infertility in a man with 21-hydroxylase deficiency congenital adrenal hyperplasia. AB - OBJECTIVE: To report a case of an azoospermic man diagnosed with 21-hydroxylase deficiency congenital adrenal hyperplasia who successfully conceived with intrauterine insemination (IUI) after hormonal and clomiphene citrate (CC) treatment. DESIGN: Case report. SETTING: Outpatient practice and academic hospital. PATIENT(S): A 32-year-old azoospermic man who presented for evaluation of male factor infertility. INTERVENTION(S): Semen analysis, ultrasonography, endocrinologic assays, hormonal treatment, CC, and IUI. MAIN OUTCOME MEASURE(S): Semen analysis demonstrating increased sperm count and motility, ultrasonography demonstrating persistent nodules in bilateral testes, endocrinologic assays demonstrating elevated FSH and LH after CC treatment. RESULT(S): Successful intrauterine pregnancy after IUI using the patient's sperm. CONCLUSION(S): Clomiphene citrate treatment in addition to hormonal manipulation in azoospermic patients with congenital adrenal hyperplasia can be successful in improving sperm count and motility to allow for successful conception using IUI. The presumed adrenal rests in the testes may not involute after adrenal suppression. PMID- 15652917 TI - Hyperbaric oxygen and ovarian follicular stimulation for in vitro fertilization: a pilot study. AB - Our objective was to assess the safety and tolerability of hyperbaric oxygen therapy (HBO) as an adjunct to IVF therapy in women with a poor prognosis for pregnancy in a prospective observational pilot study. We conclude that HBO is well tolerated by women undergoing IVF treatment and that further study is required to determine whether this is an efficacious adjuvant therapy for women being treated by IVF. PMID- 15652918 TI - Single-dose administration of an aromatase inhibitor for ovarian stimulation. AB - Recently, the success of a 5-day administration of an aromatase inhibitor for ovarian stimulation has been shown. In this report, a convenient simple regimen of administering an aromatase inhibitor as a single dose has been found to have comparable success to a 5-day regimen for ovarian stimulation. PMID- 15652919 TI - Decrease in adiponectin levels in women with polycystic ovary syndrome after an oral glucose tolerance test. AB - Adiponectin levels are decreased after an oral glucose tolerance test. At t = 2 hours, they are increased in obese and overweight women with polycystic ovary syndrome, compared with controls matched for body mass index. PMID- 15652920 TI - A suboptimal endometrial pattern is associated with a reduced likelihood of pregnancy after a day 5 embryo transfer. AB - This retrospective study examined the association of endometrial pattern and pregnancy after a day 5 ET. The pregnancy rate of women with a triple-line ultrasound endometrial pattern on the day of hCG administration was significantly higher than the pregnancy rate of women with the other ultrasound patterns. This observation suggests that, in a subset of patients, a suboptimal endometrial lining may interfere with assisted reproductive technology success. PMID- 15652921 TI - Effects of retinoic acid on the inner cell mass in mouse blastocysts. AB - This is the first evidence that excess retinoic acid has a direct cellular response from proliferation to cell death (apoptosis) and affects in vitro development in mouse inner cell mass. PMID- 15652922 TI - Impact of the presence of one or more multinucleated blastomeres on the developmental potential of the embryo to the blastocyst stage. AB - A retrospective study of 5,982 embryos in 619 blastocyst-stage embryo transfer cycles revealed that detection of multinucleated blastomeres either on day 2 or 3 signifies a poor prognosis for blastocyst formation and that no good-quality blastocyst can be expected from an embryo with more than one multinucleated blastomere. Patients should be counseled regarding a poor prognosis when multiple embryos with multinucleated blastomeres are present. PMID- 15652923 TI - First derivation in Spain of human embryonic stem cell lines: use of long-term cryopreserved embryos and animal-free conditions. AB - The first two human embryonic stem cell lines (VAL-1 and VAL-2) have been derived in Spain with long-term cryopreserved embryos under animal-free conditions. In the first series, 40 human embryos that had been cryopreserved at day 2 of development were thawed after >5 years. A derivation efficiency of 5% per frozen embryo or 12.5% per blastocyst was obtained. PMID- 15652925 TI - Uterine anomalies ascertained before pregnancy-a more frequent and challenging scenario. PMID- 15652927 TI - Endometrial "natural killer cell" lymphocyte population and implantation failure? PMID- 15652928 TI - Effect of phytoestrogens on the endometrium? PMID- 15652930 TI - Fracture treatment in childhood. PMID- 15652931 TI - The choice of treatment according to the type and location of the fracture and the age of the child. AB - Children injure themselves more today then they have in the past.Injuries that were sustained by older children are now occurring in younger children as a result of the new variety of sports ad equipment. In addition, children are traveling and running at higher speeds and the fractures sustained are becoming more complex. This requires a new "code of practice" to ensure children are correctly treated in every hospital. These new types of fractures are not always treated correctly; people tend to think of children as only small adults and that children's fractures always heal well. However, specific injuries to the growth plate and the joints as well as special fractures like the green stick and bowing (plastic deformation) fractures maybe assessed incorrectly in their healing behavior. In the following article, the basis of adequate fracture treatment will be explained. Factors that particularly influence the treatment include: age; weight; type of fracture, bone segment; available equipment; own experience; postoperative management; and cost effectiveness. PMID- 15652932 TI - The principles of elastic stable intramedullary nailing in children. AB - Elastic nailing is the foremost method of surgically stabilizing children's long bone fractures. This article traces the history of the method, summarizes the scientific basis, both biological and biomechanical, and assesses the broad indications. PMID- 15652933 TI - State-of-the-art treatment of forearm shaft fractures. AB - As the treatment of pediatric forearm shaft fractures has changed extensively over the past 30 years, it is worth discussing the current treatment modalities in these fractures. The recommendations are based on an ongoing evaluation of treatment procedures, problems, complications, and final results going back to 1976. Between 1976 and 1985, the 95.9% of fractures that were treated conservatively were tainted with bad functional results because a significant proportion healed with malalignment. The introduction of elastic-stable intramedullary nailing (ESIN) initiated a definitive change and the opportunity to stabilize unstable fractures with a less invasive method. Apart from some technical problems, the functional results of ESIN are very satisfactory. The external fixator is a good treatment for open, comminuted, or special distal dia metaphyseal fractures in older children and adolescents. Therefore, we currently have a variety of methods to treat forearm shaft fractures in children using primary definitive fracture care (PDFC). The differentiated use of conservative and surgical measures should minimize final functional sequelae. A well-tried algorithm for this fracture region is outlined. PMID- 15652934 TI - Principles of fracture remodeling in children. AB - In treating fractures in children, the surgeon must have a good knowledge of the three phases of bone healing, ie, inflammatory, reparative, and remodeling and understand how they contribute to the final recovery of the fracture healing process. By and large, the ability to remodel depends on the bone involved, the patient's age, the proximity to the joint, and its orientation to the joint axis. In the typical long bone, 75% of the remodeling occurs by reorientation of the physis while appositional remodeling of the diaphysis can only be expected to contribute 25% to the remodeling process. The various values of acceptable alignment for each of the major fracture patterns are outlined. These serve only as guidelines. The patient's functional capacity and the surgeon's experience should also be factors in determining whether to depend on the remodeling capacity of the specific fracture or to consider performing a more aggressive, invasive technique to achieve a satisfactory result. There are two advantages in treating children's fractures. First, the healing process is very rapid. Nonunion is a rare event in the pediatric age group. The second perk is that there is a very good remodeling capacity should there be less than anatomical alignment of the affected bone once the fracture has healed. Any individual treating fractures in the pediatric age group should fully understand how pediatric fractures heal and how the remodeling process occurs. PMID- 15652935 TI - What must we respect in articular fractures in childhood? AB - Articular fractures in children are rare and occur mainly in the elbow and ankle joint. Condylar fractures of the humerus, olecranon fractures, medial malleolar, and transitional fractures of the distal tibia are the commonest, though there is an increasing incidence of avulsion fractures of the intercondylar tibial eminence. Conservative treatment is recommended in articular displacements of less than 2 mm, but dislocation of more than 2 mm is a commonly accepted indication for surgery. Although this widely accepted guideline is based on clinical results, it is not scientifically proven. In addition, it is unknown whether there is any difference in the outcome for articular fractures if there is a step or gap, if the joint is weight-bearing or not, or the joint is immobilized or mobilized post-operatively. In experiments with rats, rabbits, and sheep, some clues have suggested that exact reconstruction of the chondral surface, fixation with compression of the fragments, and early continuous passive motion are advantageous. A prospective clinical study with a pre- and postoperative MRI scan is needed, in weight-bearing and non-weight bearing joints to show the factors influencing the outcome of these injuries. PMID- 15652936 TI - Problems of operative and non-operative treatment and healing in tibial fractures. AB - Based on the experience of the department of pediatric surgery at the Strasbourg University Hospital, the authors present indications and limitations of various treatment techniques for tibial shaft fractures in children. Two retrospective series are reported on: one group of patients treated with the Sarmiento technique and one group of patients treated with Elastic Stable Intramedullary Nailing (ESIN or Metaizeau technique). The main problem in treatment is restoration of the mechanical axis of the tibia, especially in isolated fractures treated conservatively. Residual varus deformity is frequent, although it is usually within an acceptable range. ESIN has given better results than conservative treatment in this specific case, but valgus malunion shave been noticed when both bones were fractured, due to insufficient bending of the medial nail. ESIN is perfectly suited for treating polytrauma patients,especially those with multi level or open fractures (Gustilo 1 or 2), as it makes monitoring and nursing easier. Treatment of tibia fractures in children remains principally conservative. ESIN is indicated in cases of failure of conservative treatment, especially in isolated tibia fractures or polytrauma patients. For high grade open fractures (Gustilo 3)or extended comminuted fractures, external fixation remains the gold standard. PMID- 15652937 TI - Concept of treatment in supracondylar humeral fractures. AB - This opinion of treatment for children's supracondylar humeral fractures discusses techniques of fracture reduction and stabilization. A useful method of external visualization of the fracture is reviewed. The article discusses the treatment of 95 fractures in children using crossed and two lateral pin techniques. PMID- 15652938 TI - Pelvic and acetabular fractures in childhood. AB - In pediatric pelvic fractures and acetabular fractures, the treating physician is confronted with a unique pattern of injuries. Throughout the literature, pediatric pelvic fractures are rare, but acetabular fractures are even rarer. The lack of experience with this type of fracture, due to their infrequency, leads to unavailability of standardized protocols for specific diagnosis and therapy. Anatomical differences and various stages of skeletal maturation between children and adults lead to different causes, patterns, and associated injuries. In this article, we review the epidemiology and pathophysiology of this entity of fractures and provide current algorithms for diagnosis and therapy. Both operative and nonoperative treatments have been advocated in the literature, and in this article we define indications for both approaches and review the current literature. PMID- 15652939 TI - Modern treatment of pathological fractures in children. AB - Pathological fractures in children occur from a variety of causes. The clinician has to develop an individual strategy that weighs up the pros and cons of treatment, considering the bone biology and the biomechanics at the fracture site. Determining the lesional pathology is paramount, whether it is in the form of a neoplastic or non-neoplastic entity, so that bone healing can be maximized. Time-efficient protection and reconstruction of the child's musculoskeletal system is the aim of the surgeon's intervention. The whole strategy must be individualized to every situation. Combinations of diseases, fracture sites, and children are many, whereas treatment options are fewer. Options include non operative management with observation, and intralesional resection with or without bone grafting, and with or without internal fixation. A thoughtful, disciplined, and systematic approach will be successful most often. The goals of treating a child's pathological fracture are all based on establishing a diagnosis. Only after establishing the diagnosis with certainty can a proper treatment strategy be formulated. Fracture management is then based on five points: (a) pain relief and comfort of the child; (b) achieve Local control or containment of the pathological lesion; (c) skeletal stabilization, preservation of growth, and maintain anatomical alignment; (d) fracture union; and (e) restoration of function. Maintaining perspective will facilitate optimal patient care and minimize the confounding variables that seem to plague the treatment of pathological fractures. PMID- 15652940 TI - Reduction and osteosynthesis of radial neck fractures in children by centromedullary pinning. AB - Radial neck fractures are infrequent but very difficult to treat and have many possible complications. Conservative treatment is unable to reduce severe displacements, resulting in malunion. Open reduction frequently leads to intraarticular calcification, avascular necrosis, and joint stiffness. In most cases, intramedullary nailing allows complete reduction without open joint and a very low complication rate. The pin is left in place, stabilizes the epiphysis, and prevents secondary displacement. PMID- 15652941 TI - Complications and failures of the ESIN technique. AB - Only knowledge of the possible failures and complications of a method allows its correct application. This also applies to ESIN (Elastic Stable Intramedullary Nailing), which has been promoted as a simple procedure. Most failures occur through neglect of the important biomechanical principles, and by obvious technical errors. The combination of both causes of failure can lead to serious complications in this method, which is generally regarded as "benign". These complications include serious rotational deformity, leg shortening, and growth disorders from growth plate injuries. A fundamental distinction must be made between intraoperative and postoperative problems and true complications. The present paper is intended to contribute towards avoiding such errors and detecting errors as early as possible during surgery in order to correct them. PMID- 15652942 TI - Femoral shaft fractures in children. AB - Femoral shaft fractures are the commonest diaphyseal fractures of childhood after those of the radial and ulnar shaft and the tibial shaft. Common mechanisms include falls, particularly from playground equipment, motor vehicle accidents and sporting injuries. Unlike in adults, femoral shaft fractures are commonly isolated injuries in children. In infants, they may be due to non-accidental injury. Treatment of femoral shaft fractures varies with the age and size of children, associated injuries and local practice. Current modalities used for the treatment of femoral shaft fractures include various forms of traction, immediate and late spica casting, elastic nailing, external fixation, plate fixation, and conventional intramedullary nailing for older children and adolescents. All these forms of treatment have been reported as being successful in cohort studies. Very few comparative studies exist. The main current controversies are the age at which elastic nailing becomes appropriate instead of conservative management, and secondly, the treatment of the older, heavier child for whom elastic nails may not be appropriate. Familiarity with several methods of femoral shaft stabilisation is appropriate. The major determinant in the choice of treatment is cultural. PMID- 15652943 TI - Basic science faculty in surgical departments: advantages, disadvantages, and opportunities. PMID- 15652944 TI - Effect of melatonin on wound healing in normal and pinealectomized rats. AB - Melatonin usage is increasing gradually, but reports of its effects on wound healing are inconsistent. It has been shown that the hormone is synthesized in and secreted from the gastrointestinal system independently of the pineal gland. We have investigated, by means of a comparative study on the healing of incision and anastomotic wounds, whether melatonin has an effect on wound healing independent of the pineal gland. Rats were divided in five groups (n = 10), all of which were subjected to small intestine anastomosis. The first group (control) was otherwise untreated. Exogenous melatonin was given to the rats in second group. The calvaria was opened then closed in the third group (sham operated), whereas the fourth group was pinealectomized and the fifth group were pinealectomized and then treated with melatonin. After anastomosis bursting pressures and incision wound breaking strength were measured on the 7th postoperative day, tissue hydroxyproline levels were determined, and histopathological investigation was performed. It was found that while collagen deposition and epithelization increased concurrently in incision wounds after pinealectomy, only collagen deposition increased at the anastomosis line. Exogenous melatonin decreased collagen synthesis and epithelium proliferation and had negative effects on wound healing in both normal and pinealectomized rats. PMID- 15652945 TI - Lack of enteral nutrition--effects on the intestinal immune system. AB - BACKGROUND: Total parenteral nutrition (TPN) results in a loss of mucosal immune function by alterations in both phenotype and function of intraepithelial lymphocytes (IEL). We hypothesized that the observed changes with TPN administration are caused by the lack of enteral feeding, and not to the TPN solution itself. METHODS: Mice received oral feeding (Control), TPN alone (TPN), or TPN plus oral feeding (TPN+Food). Mice were killed after 7 days, and bacteriological cultures from spleen, liver, and mesenteric lymph nodes obtained, with bacterial translocation (BT) being defined as a positive culture. IEL phenotype was analyzed by flow cytometry. IEL messenger RNA (mRNA) cytokine expression used reverse transcriptase polymerase chain reaction (RT-PCR). Apoptosis was detected by terminal deoxynucleotidyl transferase biotin-dUTP nick end labeling (TUNEL) staining. RESULTS: BT significantly (P < 0.05, with analysis of variance [ANOVA]) increased in the TPN group (53%) compared with Control (9%) and TPN+Food (14%) groups. TPN also resulted in a significant (P < 0.01) increase in epithelial cell apoptosis: TPN 7.6 +/- 1.1% versus Control 2.9 +/- 1.1% and TPN+Food 2.1 +/- 0.3% (mean +/- SD). Height of the villus-crypt complex was significantly decreased in TPN mice (315 +/- 16 microm) compared with Control (431 +/- 27 microm) and TPN+Food (421 +/- 26 microm) groups. IEL phenotypes significantly changed with TPN administration: CD4+ CD8- as well as CD4+ CD8+ subpopulations were reduced compared with Control or TPN+Food mice; as were the CD8alphabeta+ thymus-dependent, and CD8+ CD44+ mature IEL. IEL cytokine mRNA expression was also significantly altered with TPN: IL-2 and IL-10 expression declined, and IL-4 IL-6, interferon gamma (IFN-gamma), transforming growth factor beta-1 (TGF-beta1), and tumor necrosis factor-alpha (TNF-alpha) were increased, when compared with Control or TPN+Food mice. CONCLUSIONS: This study demonstrates that the major factor responsible for TPN-induced BT and IEL-changes is the lack of enteral feeding and not the administration of the TPN solution itself. PMID- 15652946 TI - Supplemental enteral arginine is metabolized to ornithine in injured patients. AB - BACKGROUND: Arginine has been added to immune enhancing diets that may improve patient outcomes, but little is known about the metabolic fate of supplemental arginine. We hypothesize that supplemental enteral arginine in injured patients is metabolized to ornithine by increased activity of the enzyme arginase. MATERIALS AND METHODS: Twenty-five adult patients with injury severity scores > or =20 received up to 14 days of enteral nutrition supplemented with arginine (30 g/day) or placebo in a prospective, randomized, blinded study. Plasma arginine, citrulline, and ornithine concentrations and peripheral blood mononuclear cell (PBMC) arginase activity were measured at baseline and on days 1, 3, 5, 7, 10, and 14. Clinical data collected included demographics, injury patterns, lengths of stay, and infectious complications. Data were analyzed using ANOVA and t test. RESULTS: PBMC arginase activity was elevated in all patients. In the supplemented group, plasma arginine concentrations increased at days 7, 10, and 14 when compared to baseline (P < 0.05) and were higher at day 14 when compared to those of controls (P < 0.05). Citrulline concentrations in both groups were unchanged over time. Ornithine concentration increased within 24 h of arginine supplementation and remained elevated when compared to baseline (P < 0.01). Ornithine concentration in the supplemented group was higher at days 1, 3, 5, and 7 when compared to that of controls (P < 0.05). There were no differences in clinical outcomes. CONCLUSIONS: Supplemental enteral arginine is absorbed in injured patients and increases arginine levels. Supplemental arginine appears to be metabolized to ornithine. Increased arginase enzyme activity in peripheral blood mononuclear cells may be a contributor. PMID- 15652947 TI - Plasma neurohormone levels correlate with left ventricular functional and morphological improvement in LVAD patients. AB - OBJECTIVE: Brain natriuretic peptide (BNP) and endothelin-1 (ET-1) have been shown to be markers of left ventricular (LV) function. To determine the feasibility of using serial assays of these neurohormones in the assessment of cardiac status in the left ventricular assist device (LVAD) setting, we examined the relationship between LV function, myocardial morphology, and plasma levels of these hormones in LVAD recipients. METHODS: Plasma BNP and ET-1 levels were serially assayed in 19 end-stage congestive heart failure (CHF) patients before and after LVAD implantation with various devices (i.e., MicroMed DeBakeyVAD/DVAD, Novacor/NVAD, TCI Heartmate/TCI, Thoratec/TVAD). Echocardiography performed correspondingly at the time points of the hormonal assays and immunohistochemical collagen staining of left ventricular tissue samples, derived from six non failing hearts as well as from LVAD patients at the time of device insertion and removal, were then contrasted. Patients were grouped according to device used and etiology of heart disease (ischemic or dilated cardiomyopathy, ICM/DCM). RESULTS: LVAD therapy significantly improved LV ejection fraction (EF%: 21 +/- 3.8% to 28.11 +/- 3.57%), cardiac output (CO: 3.49 +/- 1.3 to 7.3 +/- 0.2 l/m), and left ventricular end-diastolic diameter (LVEDD: 6.68 +/- 0.92 versus 4.79 +/- 1.54 cm, P < 0.0001) in all patients. Absolute BNP and ET-1 plasma levels remained significantly lower in all patients after LVAD implantation (both P < 0.001). The NVAD group exhibited the most BNP reduction and EF% increase (P < 0.0004 and P < 0.038, respectively). Average collagen levels were reduced in all patients (P < 0.0005). Among the devices, the NVAD group demonstrated the most evident change (P < 0.0036), while there was comparable reduction in the DCM and ICM groups (both P < 0.03). In general, postoperative BNP and ET-1 trends exhibited a notable parallelism with both manifesting bi-phasic tendencies and an inverse proportionality to corresponding EF% measurements. CONCLUSIONS: Device selection appears to influence the cardiac morphological and neurohormonal expressive tendencies exhibited by recipients. Plasma BNP and ET-1 levels correlate with both LV function and myocardial morphological improvement. Alterations in the levels of these hormones during LVAD support may be real-time indicators of prevailing myocardial autocrine/paracrine activity and as such may be of potential use in future algorithms of cardiac assessment and therapeutic decision making with regard to transplant urgency and/or possible device explantation. PMID- 15652948 TI - Angiotensin II type 1 receptor activation increases microvascular permeability via a calcium dependent process. AB - BACKGROUND: Elevated serum angiotensin II (Ang II) has been implicated in the endothelial barrier dysfunction associated with shock. We hypothesized that the increase in microvascular permeability seen with activation of the type 1 (AT1) receptor is a calcium dependent process. MATERIALS AND METHODS: Microvascular hydraulic permeability (Lp) was measured in rat mesenteric venules using the Landis micro-occlusion model. A 100 mm KCl (HK) solution was used to negate the electrochemical potential of calcium influx, and measures of Lp were obtained before and after 20 ng/ml Ang II plus HK solution (n = 5). Intracellular calcium dependence on AT1 activation was evaluated two ways: 1) Lp changes were measured in response to 10 microm of the type 1 receptor agonist [SAR] [1]-angiotensin II in HK solution (n = 6), and 2) Lp changes were measured in response to 25 microg/ml of the type 2 (AT2) receptor blocker PD-123319 (PD) plus 20 ng/ml Ang II in HK solution (n = 6). RESULTS: As expected, HK perfusion (P < 0.08) and Ang II plus HK solution (P < 0.42) did not affect Lp. Although perfusion of [SAR] [1] angiotensin II in HK solution (P < 0.001) and PD plus Ang II in HK solution (P < 0.003) both significantly increased Lp, the magnitude of this response was less than that observed with Ang II alone. CONCLUSIONS: Abrogation of intracellular calcium influx during AT1 activation blunted the known Ang II induced increase in microvascular permeability. Although the effect observed during AT1 activation was blunted by the HK solution, a significant elevation of Lp was still observed. This suggests that Ang II activation of the AT1 receptor increases microvascular permeability primarily, but not exclusively, via modulation of endothelial intracellular calcium ion levels. PMID- 15652949 TI - Matrix metalloproteinases in ascending aortic aneurysms: bicuspid versus trileaflet aortic valves. AB - BACKGROUND: Abnormal matrix metalloproteinase (MMP) and tissue inhibitor of metalloproteinase (TIMP) expression contributes to the development of abdominal aortic aneurysms. Recent data suggest that MMP-2 and MMP-9 may also play a role in thoracic aortic disease. We sought to determine (1) whether ascending aortic aneurysms are associated with increased MMP expression and (2) whether aortic inflammation and MMP expression differ between patients with congenital bicuspid aortic valves (BAVs) and those with trileaflet aortic valves (TAVs). MATERIALS AND METHODS: Samples of ascending aortic aneurysms were obtained from 29 patients; 14 patients had BAVs and 15 had TAVs. Control ascending aorta was obtained from 14 organ donors or heart transplant recipients. Aortic histology and immunohistochemistry were performed to evaluate elastin degradation, inflammatory changes, and MMP-2 and MMP-9 expression. Aortic levels of MMP-2, MMP 9, TIMP-1, and TIMP-2 were measured using ELISA. RESULTS: Aneurysms in the TAV patients exhibited marked inflammation, high CD68 expression, diminished elastin content, increased MMP-9 expression, and normal MMP-2 levels. In contrast, BAV aneurysms were characterized by a relative lack of inflammation, preservation of elastin content, normal MMP-9 levels, and elevated MMP-2 expression. TIMP-1 and TIMP-2 levels were not significantly different among the three groups. CONCLUSIONS: Ascending aortic aneurysms exhibited increased MMP expression. The pattern of MMP expression and the degree of inflammation, however, differed between aneurysms associated with BAVs and those with TAVs. Variations in the molecular mechanisms underlying different types of thoracic aortic aneurysms warrant further investigation. PMID- 15652950 TI - Targeting of vaccinia virus using biotin-avidin viral coating and biotinylated antibodies. AB - INTRODUCTION: To test a general method for altering the tropism of viral vectors, we conjugated targeting antibody to the surface of recombinant vaccinia virus with a biotin-avidin-biotin linker and assessed the resulting infectivity in target cells and controls. MATERIALS AND METHODS: We biotinylated a vaccinia viral vector and used avidin to crosslink the biotinylated viral surface to a biotinylated antibody specific for a molecule on the surface of a target cell. In an in vitro model system, we coated a recombinant vaccinia construct containing the E. coli beta-galactosidase gene with antibody to the murine class I MHC molecule Db. Target cells were B78H1 murine melanoma cells transduced with either the Db gene or, as a control, the Kb gene. Infectivity was assessed by staining target cells with x-gal to demonstrate expression of virally delivered beta galactosidase. This technique was also assessed in a second system with vaccinia/beta-gal targeted to the murine B7.2 molecule. The infectivity of the resulting construct was assessed for murine SA1 fibrosarcoma cells transfected with the B7.2 gene and for wild-type, B7.2-negative SA1. Experiments were repeated in each system with similar results. RESULTS: This strategy demonstrated antibody-mediated viral targeting in both the B78H1 and the SA1 models. Importantly, addition of the targeting coat diminished the infectivity of the modified vaccinia for control cells but preserved infectivity for targeted cells. In the B78H1 system, Db-targeted vaccinia consistently had 2- to 3-fold greater infectivity for B78H1Db than B78H1Kb. Increasing the number of avidin molecules used per virion in the synthesis of the viral coat led to greater selectivity but decreased overall infectivity. In the SA1 system, B7.2-targeted vaccinia demonstrated completely ablated infectivity for control SA1 cells, but maintained infectivity for target SA1/B7.2 cells. CONCLUSIONS: Recombinant viral vectors such as vaccinia may be coated with biotin/avidin and linked to biotinylated antibodies to preferentially target specific cell types in vitro. Such an approach may be useful in targeting recombinant lytic viruses to tumors for destruction and in immune up-regulation in vivo. Similarly, this approach may enhance nonlytic viruses for gene therapy applications. PMID- 15652951 TI - HIV-based vectors and angiogenesis following rabbit hindlimb ischemia. AB - BACKGROUND: Numerous medical and surgical options exist for the treatment of vessel ischemia, which some patients fail or cannot tolerate. These investigations were designed to determine the effects of lentiviral-delivered vascular endothelial-derived growth factor (VEGF) and angiopoietin-2 (Ang-2) on collateralization in a rabbit model of hindlimb ischemia. MATERIALS AND METHODS: Self-inactivating human immunodeficiency virus (HIV)-based vectors were constructed encoding VEGF or Ang-2, co-transfected with vesicular stomatitis virus glycoprotein (VSV G) into 293T cells, and vector supernatants (1 x 10(8) IU/ml after concentration) were harvested. New Zealand white rabbits had ligation of either the right or left external iliac artery and excision of the ipsilateral femoral artery. Ten days later, empty, VEGF, or VEGF+Ang-2 vector supernatant was injected intramuscularly (IM) into the ipsilateral thigh. Ankle systolic blood pressure (SBP) ratios were recorded and venous blood samples collected on postoperative days (POD) 10, 25, and 40. On POD 40, run-off angiography was performed to measure vessel collateralization. Capillary density was determined by thin sectioning of muscle. RESULTS: A significant increase was noted in SBP in the VEGF-treated animals over time. Capillary density was not elevated despite significantly increased large vessel collateralization in rabbits receiving VEGF, which was counteracted by Ang-2. Antibodies against vector components were detected in exposed serum. CONCLUSIONS: Arterial collateralization and SBP increased significantly following VEGF vector administration, which was reversed by the Ang-2 vector. Development of antibody against VSV G can limit repeated injections of vector. Future experiments will involve the addition of other pro angiogenic factors, repeated vector administration, and alternative routes of vector delivery. PMID- 15652952 TI - Photodynamic therapy induced esophageal stricture--an animal model: from mouse to pig. AB - INTRODUCTION: A major limitation of photodynamic therapy (PDT) for Barrett's esophagus is the development of esophageal stricture. We developed an animal model of PDT-induced esophageal stricture to elucidate the mechanism of stricture development. Our studies began in a mouse but, due to its limitations, we advanced to a porcine model. MATERIALS AND METHODS: In the mouse model, 62 mice were injected with Photofrin (2-10 mg/kg) 48 h prior to photoactivation. Light energy (20-400 Joules/cm (J)) was delivered with a laser probe as a single dose, or fractionated doses (20-150 J). Animals were sacrificed when showing signs of distress or 6 to 18 weeks post-illumination. Esophagus was removed, with gross and microscopic examination performed on frozen specimens. To develop a pig model, six pigs were injected with Photofrin (2 mg/kg) 48 h prior to photoactivation. Light energy (400 J) was delivered via an endoscope using a laser probe as a single dose or repeated at 48 h. Animals were sacrificed if they could not eat soft food or lost more than 10% of their original weight according to the University of Pittsburgh Institutional Animal Care and Use Committee. RESULTS: Exposure of mice to doses of 400 J x 1, 125 J x 3, or 150 J x 3 fractions resulted in severe lung damage and death in 90% of the mice without any evidence of esophageal stricture. Lower energy levels caused minor lung damage and no change in the endothelial layer or a stricture. In pigs, exposure of 400 J as one or two fractions resulted in weight loss of 10% within 3 weeks. Endoscopy, upper GI, contrast swallow, and pathological and histological examination showed evidence of esophageal stricture at the exposed area. CONCLUSIONS: In the mouse model, pulmonary toxicity is the limiting factor following esophageal PDT exposure. In the pig model we induced esophageal stricture following PDT. This is the first animal model created to study esophageal strictures resulting from PDT. PMID- 15652953 TI - Glutamine stimulates amino acid transport during ischemia-reperfusion in human intestinal epithelial cells. AB - BACKGROUND: The potential mechanism of intestinal ischemia-reperfusion (I/R) injury includes oxygen-derived toxic free radicals. We tested the hypothesis that glutamine increases intracellular glutathione, a protective substrate against oxidative stress, by stimulating membrane amino acid transport during I/R using human intestinal epithelial cell line Caco-2. METHODS: Ischemic conditions were obtained by combining both hypoxic (1%O2-5%CO2-94% N2) and nutrient-deprived (Phosphate-Buffered Saline; PBS) conditions. After 2 h of ischemia, re oxygenation (5%CO2-95% air) was initiated and the culture medium was changed to PBS, PBS supplemented with amino acids (A.A.), and PBS supplemented with 2 mm glutamine plus amino acids (Gln) (reperfusion). After 4 h of reperfusion, the transport of 3H-glutamine, 3H-glutamate, and 3H-leucine was assayed and intracellular glutathione was measured. 3H-thymidine incorporation was measured for the determination of DNA synthesis. Data (mean +/- SD) were analyzed by ANOVA. RESULTS: Ischemia decreased Na+-dependent glutamine, Na+-dependent glutamate, and Na+-independent leucine transport compared with control (P < 0.01). After reperfusion, glutamine and glutamate transport in the PBS and A.A. groups decreased significantly compared with control (P < 0.01), whereas glutamine supplementation increased glutamine transport to the levels in control (P < 0.01) and partially increased glutamate transport (P < 0.01). Leucine transport significantly increased in the A.A. and Gln groups compared with the PBS group. Glutamine significantly increased intracellular glutathione and DNA synthesis compared with the PBS and A.A. groups (P < 0.01). CONCLUSIONS: This study demonstrated that glutamine up-regulates amino acid transport during I/R in human intestinal epithelial cells, possibly resulting in increased intracellular glutathione and DNA synthesis. PMID- 15652954 TI - Prolonged exposure of canine coronary arteries to a nitric oxide donor desensitizes soluble guanylate cyclase. AB - BACKGROUND: This study investigated the role of soluble guanylate cyclase desensitization in the development of tolerance to organic nitrates. MATERIALS AND METHODS: In organ baths, canine coronary arteries were exposed to either sodium nitroprusside (SNP) (experimental group) or papaverine (control group) at various concentrations (10(-9), 10(-7), or 10(-5) M) for 3 h. Arteries were then compared for response to vascular agonists and for inducible cyclic guanine monophosphate (cGMP) formation. RESULTS: KCl (5 to 50 mM) and prostaglandin F(2alpha) (10(-9) to 10(-5) M) induced similar vascular contractions (n = 7, P > 0.05). Vascular relaxation in response to calcium ionophore A23187 (10(-9) to 10( 6) M) and to authentic nitric oxide (NO) (3 x 10(-9) to 10(-5) M) was attenuated in arteries exposed to SNP at 10(-7) and 10(-5) M concentrations but not at a 10( 9) M concentration (n = 7 each, P < 0.05 versus the respective papaverine group). Pretreatment of arteries with methylene blue (10(-5) M) abolished the responses to authentic NO (n = 4). In cGMP determinations, control arteries demonstrated an increase in cGMP from 364 +/- 89 to 778 +/- 175 pg/mg of protein with A23187 (3 x 10(-5) M) stimulation (n = 5). Conversely, arteries exposed to SNP (10(-5) M) demonstrated similar levels of cGMP before (562 +/- 126 pg/mg of protein) and after (641 +/- 98 pg/mg of protein) A23187 stimulation. CONCLUSIONS: Prolonged exposure of coronary arteries to SNP did not alter vascular smooth muscle function, but it markedly attenuated the relaxation in response to both A23187 and authentic NO at concentrations above 10(-9) M in a concentration-dependent fashion. The constant levels of cGMP in response to an NO donor suggest that the attenuation of relaxation is due to desensitization of soluble guanylate cyclase. Thus, this study supports the role of soluble guanylate cyclase desensitization in the development of tolerance to organic nitrates. PMID- 15652955 TI - The functional and histological effects of clenbuterol on the canine skeletal muscle ventricle. AB - BACKGROUND: We investigated the anabolic effects of the sympatho-mimetic drug clenbuterol upon pumping chambers constructed from latissimus dorsi muscle (LDM). METHODS AND RESULTS: In control and treatment groups (n = 4 dogs each), skeletal muscle ventricles (SMVs) were constructed followed by a 3-week recuperative delay and 6-7 weeks of electrical conditioning at 2 Hz to induce phenotypic expression of fatigue resistant slow muscle fibers. The treatment group received oral administration of clenbuterol (8 microg/kg, 2x/day) during this period. The clenbuterol group increased significantly in body weight as compared with the control group (P < 0.05). In a terminal experiment, the SMVs were assessed with a mock circulation device to determine pumping performance and also were examined with regard to fiber type distribution and area in the SMVs and their contralateral in situ LDMs. Initially the clenbuterol group performed better than the control group, but by the end of a 60-min fatigue test, there were no significant differences. With regard to fiber type distribution and areas, the SMVs of the clenbuterol group exhibited a fast fiber distribution similar to unconditioned muscles (28% +/- 4%), whereas the control group showed complete transformation (100%) to slow fibers. The fast fibers of the clenbuterol group were larger than control (P < 0.05), but the slow fibers were not significantly different. CONCLUSIONS: At the dose given, clenbuterol does induce hypertrophy and preserves the normal percentages of fiber types, possibly by hyperplasia, but it does not affect chronic pumping performance of skeletal muscle ventricles in the canine model. PMID- 15652956 TI - Macrophage inflammatory protein-3alpha promotes pancreatic cancer cell invasion. AB - BACKGROUND: Human CC chemokine Macrophage Inflammatory Protein-3alpha (MIP 3alpha) directs inflammatory cell migration through its binding to the transmembrane receptor CCR6. MIP-3alpha has recently been shown to promote tumor cell migration in pancreatic adenocarcinoma by up-regulation of matrix metalloproteinases (MMPs). We hypothesized that MIP-3alpha promotes pancreatic cancer invasion through the up-regulation of MMP-9, a Type 4 collagenase. MATERIALS, METHODS, AND RESULTS: Immunohistochemistry and RT-PCR confirmed the presence of MIP-3alpha in PANC-1 cells, a human pancreatic adenocarcinoma cell line. MIP-3alpha stimulated the production of both latent and active forms of MMP 9 in PANC-1 by Western analysis. Tumor cell invasion was then evaluated using a modified Boyden chamber invasion assay. MIP-3alpha promoted a dose-dependent increase in pancreatic cancer cell invasion (P < 0.05) at 100 ng/ml. The activity at the putative MIP-3alpha receptor, CCR6, was demonstrated by receptor blockade. Anti-CCR6 antibody and anti-MMP-9 antibody inhibited MIP-3alpha-stimulated PANC-1 cell invasion of collagen to 37% and 35% of control, respectively (P < 0.05). CONCLUSIONS: MIP-3alpha, through its CCR6 receptor, promotes tumor cell invasion by the up-regulation of MMP-9. Molecular based therapy aimed at the inhibition of MIP-3alpha activity through the CCR6 receptor may serve as a future target to prevent tumor cell invasion in pancreatic adenocarcinoma. PMID- 15652957 TI - Improvement of nutritive perfusion after free tissue transfer by local heat shock priming-induced preservation of capillary flowmotion. AB - BACKGROUND: Capillary flowmotion protects pedicled flaps during critical perfusion conditions. However, free tissue transfer, causing ischemia-reperfusion and surgical trauma, have been shown to blunt these protective blood flow fluctuations. Because heat shock priming protects tissue after transfer, we herein studied whether heat shock protein expression is capable to preserve critical perfusion-induced capillary flowmotion in transferred composite flaps. METHODS: In Sprague Dawley rats (n = 16), osteomyocutaneous flaps were subjected to critical perfusion after harvest and 1 h and 4 h after free transfer. In eight animals additional heat shock priming was induced 24 h before flap harvest. Microcirculation including capillary flowmotion was analyzed using intravital fluorescence microscopy. RESULTS: After harvest, critical perfusion induced capillary flowmotion in skeletal muscle tissue of all flaps. By this, functional capillary density (FCD), an indicator of nutritive perfusion, was maintained not only in muscle but also in periosteum, subcutis, and skin. In contrast, 1 h after flap transfer muscle capillary flowmotion was completely abrogated, resulting in a significant decrease of FCD in all tissues. Heat shock-priming completely restored capillary flowmotion, and, by this, maintained tissue FCD. CONCLUSIONS: The loss of muscle capillary flowmotion after free tissue transfer-associated ischemia-reperfusion can be prevented by heat shock-priming. This may represent the mechanism of protection by local heat application. PMID- 15652958 TI - Protamine sulfate reduces the susceptibility of thermally injured mice to Pseudomonas aeruginosa infection. AB - BACKGROUND: In this study, we investigated the ability of protamine sulfate, at sub-bactericidal dosing, to interfere with the in vivo virulence of Pseudomonas aeruginosa (PAO1) during burn wound infection. MATERIALS AND METHODS: The study was conducted using the murine model of thermal injury. Preliminary experiments determined a protocol for administration of protamine sulfate that had no in vivo bactericidal effects. Based on this, the effect of local injection of protamine sulfate on the in vivo virulence of PAO1 was assessed using these parameters: (1) the percent mortality among PAO1-infected, thermally injured mice; (2) the local proliferation and spread of PAO1 within the infected burned tissue; (3) the systemic spread of PAO1 within thermally injured/infected mice; and (4) the local cytokine response elicited by PAO1 thermally injured/infected mice. RESULTS: Injection of protamine sulfate into the thermally injured tissue of PAO1 infected/thermally injured mice significantly decreased the percent mortality and inhibited the systemic dissemination of PAO1 microorganisms to the liver and spleen. It had no effect, however, on the ability of the bacteria to proliferate and spread within the thermally injured tissue. It also was determined that protamine sulfate was ineffective at preventing mouse death at the dose administered if injected intramuscularly instead of directly into burned tissue. Protamine sulfate reduced the expression of the proinflammatory cytokines IL-6 and LIF in the injured/infected tissue. Heparan sulfate given in conjunction with protamine sulfate returned mortality levels to those of untreated mice. CONCLUSIONS: Our results suggest that: (1) local injection of sub-bactericidal doses of protamine sulfate reduces the virulence of P. aeruginosa; (2) this effect is due to interference with the systemic rather than local spread of P. aeruginosa; and (3) local application of protamine sulfate may have potential as supportive therapy for prevention of systemic P. aeruginosa infection in severely burned patients. PMID- 15652959 TI - Organ-specific regulation of pro-inflammatory molecules in heart, lung, and kidney following brain death. AB - BACKGROUND: Nonspecific inflammatory events following brain death may increase the intensity of the immunological host response. The present study investigated the course of pro-inflammatory molecules in heart, lung, kidney, and plasma after brain death induction. MATERIALS AND METHODS: Brain death was induced in five pigs by inflation of an intracranial Foley catheter and five pigs were sham operated as controls. Each experiment was terminated 6 h after brain death/sham operation and the organs were harvested. We measured the mRNA and protein levels for TNF-alpha, IL-1beta, and IL-6 in heart, lung, kidney, and plasma. Additionally, the mRNA expression for IL-6R, ICAM-1, MCP-1, and TGF-beta was determined in each organ. RESULTS: After 6 h, the plasma cytokine levels were higher in the brain-dead animals than in the sham-operated. In heart, lung, and kidney there was an increase in IL-6 and IL-1beta following brain death, while TNF-alpha was up-regulated in lung only (P < 0.05). MCP-1 and TGF-beta were significantly higher in heart and lung and IL-6R increased in heart after brain death (P < 0.05). CONCLUSIONS: Brain death was associated with non-uniform cytokine expression patterns in the investigated organs. These expression patterns may cause variable pro-inflammatory priming resulting in different degrees of damage and explain the organ-specific variation in outcomes after transplantations. PMID- 15652960 TI - Adenosine attenuates C-terminal but not N-terminal proteolysis of cTnI during cardioplegic arrest. AB - BACKGROUND: Specific site proteolysis and loss of troponin I (TnI) during myocardial ischemic events can contribute to myocardial dysfunction. Adenosine supplementation of cardioplegic solutions results in improved functional preservation of the heart. We investigated the effect of adenosine on N-terminal and C-terminal proteolysis of TnI in the heart. MATERIALS AND METHODS: Hearts from male Sprague-Dawley rats were isolated and perfused at a constant pressure. Cardioplegic arrest (St. Thomas #2 +/- 100 microm adenosine) was induced and hearts frozen at various times during the arrest. Antibodies directed against specific regions of TnI were used to visualize TnI in whole heart homogentates, as well as from cellular fractions, using western blot analysis. RESULTS: Cardioplegic arrest alone resulted in early N-terminal proteolysis of TnI, followed by later loss of sequences from the C-terminal end of the molecule. In addition, secondary protein bands that were immunoreactive to amino acid sequences centrally located on the TnI molecule were observed. There was also evidence of dissociation of TnI from the other myofibrillar proteins. The supplementation of cardioplegic solution with adenosine significantly attenuated the late C-terminal proteolytic degradation of TnI and its apparent dissociation from myofibrils proteins but had no effect on the early N-terminal proteolysis associated with cardioplegic arrest. CONCLUSIONS: These data may provide an explanation for partial protection against postarrest myocardial dysfunction provided by adenosine. PMID- 15652961 TI - Hyperinflation during lung preservation and increased reperfusion injury. AB - BACKGROUND: Reperfusion injury after lung transplantation remains a perplexing and unpredictable problem. Most surgeons preserve the lung inflated, but the amount of inflation that should be used is not well documented. Therefore, we studied the effect of high inflation during organ preservation on lung function during reperfusion. Our hypothesis is that donor lung hyperinflation during storage contributes to early allograft dysfunction during reperfusion. METHODS: To test our hypothesis we used an isolated, blood-perfused, ventilated rabbit lung model. Group I lungs (control) underwent immediate reperfusion after harvest. Group II lungs (low-inflation, maintained at 6 mmHg airway pressure) and group III lungs (high-inflation, maintained at 20 mmHg airway pressure) were stored for 4 h in 4 degrees C Euro-Collins solution after harvest. All lungs were then reperfused with whole blood for 1 h, and measurements of arterial oxygenation (PO2, mmHg), pulmonary artery pressure (PAP, mmHg), peak inspiratory pressure (PIP, cm H2O), and wet-to-dry weight ratio (WTD) were obtained. RESULTS: Throughout the 1 h reperfusion period group III lungs had significantly lower oxygenation compared to groups I and II. In addition, throughout reperfusion, group III lungs showed significantly higher PAP and PIP compared to group II. WTD did not differ significantly between groups, however, there was a trend toward increased edema in group III. CONCLUSIONS: These results indicate that high inflation during cold storage results in acute pulmonary dysfunction. Careful monitoring of airway inflation pressure during storage, especially to prevent hyperinflation, should be maintained in the current practice for lung transplantation. PMID- 15652962 TI - Controlled study of inline radiofrequency ablation (ILRFA) assisted transection of ovine liver. AB - BACKGROUND: Liver resection is now a standard treatment for primary and secondary hepatic tumors around the world. Intra-operative blood loss during liver resection is a major factor associated with morbidity and mortality. We have developed a new instrument using radiofrequency energy (ILRFA), which is intended to achieve coagulative ablation in a plane. This plane can then be cut through with a scalpel, ultrasonic dissector, or diathermy with minimal blood loss. MATERIALS AND METHODS: Five sheep were used in this non-recovery experiment. In these sheep we performed five liver resections with the ILRFA and five similar resections using diathermy and suturing as control. Blood loss was measured by determining the difference in the weights of dry sponges and blood stained sponges after resection. RESULTS: ILRFA was successful in achieving coagulative ablation in all cases to a width of 1 cm. The mean blood loss in ILRFA was 43.2 g (SD36) and 221.8 g (SD147) in the control group. The bleeding was significantly reduced in ILRFA group with a P value of 0.005. CONCLUSIONS: Bleeding remains an important complication of liver resection. To reduce bleeding during liver surgery, different techniques have been used. In this study, we have demonstrated that by using ILRFA we can perform liver resections in sheep with minimal blood loss. PMID- 15652963 TI - Gene profiling in muscle of severely burned children: age- and sex-dependent changes. AB - BACKGROUND: Thermal injury is associated with a pronounced catabolic response in skeletal muscle. This study identifies gene expression changes in skeletal muscle of thermally injured girls and boys using high-density oligonucleotide arrays. METHODS: Six burned children with a mean age of 8.3 +/- 1.3 years and TBSA burn size covering 51 +/- 6% admitted to our hospital with in 48 h of injury and six cleft lip and cleft palate patients were studied. Total RNA was isolated, in vitro transcribed, and hybridized to HG-U95 Av.2 Affymetrix arrays. Messenger RNA expression patterns of controls and burn patients were compared using Affymetrix GeneChip Analysis Suite 5.2 and dChip. RESULTS: Statistical analysis of the 12,625 genes on each array showed a significant increase in the expression of 77 genes in burn children and a decrease in 21 genes when compared to controls (P < 0.05). We found three genes in burned males and two genes in burned females with decreased expression in muscle compared to controls. Chromosomes 1, 2, 7, 12, and 16 showed genes with increased expression in muscle from burned children, while chromosomes 3, 7, 8, 19, and 22 had genes with decreased expression. Categories of genes affected were related to metabolism, proliferation, transcription/translation, immune response, stress response, angiogenesis, and signal transduction. CONCLUSIONS: Genes that are differentially expressed in skeletal muscle of burned children, but whose function in muscle is unknown, include those related to various transcription factors and those known to encode proteins involved in signaling pathways. Further analysis is required to achieve the ultimate goal of making functionally relevant conclusions about the molecular pathology of burn injury. PMID- 15652964 TI - Basic science faculty in surgical departments: advantages, disadvantages and opportunities. AB - The number of Ph.D. faculty in clinical departments now exceeds the number of Ph.D. faculty in basic science departments. Given the escalating pressures on academic surgeons to produce in the clinical arena, the recruitment and retention of high-quality Ph.D.s will become critical to the success of an academic surgical department. This success will be as dependent on the surgical faculty understanding the importance of the partnership as the success of the Ph.D. investigator. Tighter alignment among the various clinical and research programs and between surgeons and basic scientists will facilitate the generation of new knowledge that can be translated into useful products and services (thus improving care). To capitalize on what Ph.D.s bring to the table, surgery departments may need to establish a more formal research infrastructure that encourages the ongoing exchange of ideas and resources. Physically removing barriers between the research groups, encouraging the open exchange of techniques and observations and sharing core laboratories is characteristic of successful research teams. These strategies can meaningfully contribute to developing successful training program grants, program projects and bringing greater research recognition to the department of surgery. PMID- 15652965 TI - The NF-kappaB/IkappaB signaling system: a molecular target in breast cancer therapy. AB - The nuclear factor kappaB (NFkappaB) superfamily of eukaryotic transcription factors plays an important role in carcinogenesis. NF-kappaB and its regulators are linked to various signal transduction pathways as well as transcriptional activation events that mediate critical stages of cell proliferation. These intracellular signaling processes are thought to regulate chromatin structure to accommodate transcription, apoptosis, cell-cycle control, and cell transformation. In this capacity, uncontrolled or aberrant NF-kappaB activity may, in part, be responsible for breast cancer progression. Constitutive NF kappaB expression may predict the metastatic potential of breast tumors, indicating early use of adjuvant therapy and suggesting NF-kappaB inhibition as a novel treatment. In this review, we discuss the regulatory mechanisms and physiological significance of NF-kappaB activation, and highlight recent advances in the development of NF-kappaB as an integral mediator of mammary carcinogenesis. PMID- 15652966 TI - Inhibition of human cytomegalovirus signaling and replication by the immunosuppressant FK778. AB - FK778 (Fujisawa Healthcare Inc.) is an immunosuppressant structurally similar to A771726, the active metabolite of leflunomide (Aventis Pharmaceuticals), but with a clinically relevant shorter serum half-life. Leflunomide, a tolerated and efficacious immunosuppressive agent in patients receiving allograft transplantations, was reported to be active against HCMV and HSV-1. Here we report that FK778 is a potent and effective inhibitor of HCMV, and that its mode of antiviral action appears to mirror the biochemical mechanisms elsewhere described to be responsible for its immunosuppressive properties: inhibition of protein tyrosine phosphorylation and inhibition of cellular de novo pyrimidine biosynthesis. Initial HCMV-mediated activation of the EGF receptor/phosphatidylinositol 3-kinase (PI3-K) pathways and Sp1 and NF-kappaB were partially inhibited by FK778. The second tier (phase) of PI3-K, Sp1, and NF kappaB induction by HCMV was more sensitive to FK778. Treatment of HCMV-infected cells with FK778 prevented the appearance of HCMV proteins some 12-24h post infection, and inhibited viral DNA synthesis. In our assays, leflunomide also reduced HCMV DNA levels. The antiviral activity of FK778 was reversed in cell culture by treatment with uridine, consistent with specific inhibition of dihydroorotate dehydrogenase (DHODH), a required enzyme in the de novo biosynthesis of pyrimidines. This report substantiates the clinical possibility of a single drug treatment to achieve immunosuppression and inhibit opportunistic herpesvirus infections. Our results differ from descriptions of leflunomide acting as an inhibitor of HCMV cytoplasmic capsid formation. Additionally, this study indicates that DHODH may be an effective cellular antiviral target. PMID- 15652967 TI - Combination of inhibitors of lymphocyte activation (hydroxyurea, trimidox, and didox) and reverse transcriptase (didanosine) suppresses development of murine retrovirus-induced lymphoproliferative disease. AB - The ribonucleotide reductase inhibitor hydroxyurea (HU) has demonstrated some benefit as a component of drug cocktails for the treatment of HIV-1 infection. However, HU is notoriously myelosuppressive and often administered only as salvage therapy to patients with late-stage disease, potentially exacerbating the bone marrow toxicity of HU. In this report we have compared the antiviral effects of HU and two novel RR inhibitors trimidox (3,4,5-trihydroxybenzamidoxime) and didox (3,4-dihydroxybenzohydroxamic acid) in combination with didanosine (2,3 didoxyinosine; ddI) in the LPBM5 MuLV retrovirus model (murine AIDS). We also evaluated the effects of these drug combinations on the hematopoietic tissues of LPBM5 MuLV-infected animals. The combination of RR inhibitors and ddI was extremely effective (DX>TX>HU) in inhibiting development of retrovirus-induced disease (splenomegaly, hypergammaglobulinemia, activated B-splenocytes and loss of splenic architecture). In addition, relative levels of proviral DNA were significantly lower in combination drug-treated animals compared to infected controls. Evaluation of femur cellularity, numbers of marrow-derived myeloid progenitor cells (CFU-GM and BFU-E) and peripheral blood indices revealed that TX and DX in combination with ddI were well-tolerated. However, treatment with HU and ddI induced moderate myelosuppression. These data demonstrate that RR inhibitors in combination with ddI provide significant protection against retroviral disease in murine AIDS. Moreover, the novel RR inhibitors TX and DX appear to be more effective and less myelosuppressive than HU when administered with ddI in this model. PMID- 15652968 TI - Enhancement of antiviral activity against hepatitis C virus in vitro by interferon combination therapy. AB - Alpha, beta, and gamma interferons (IFN-alpha, IFN-beta, IFN-gamma) have been shown to be effective inhibitors of HCV replication in human cell lines carrying HCV replicons. To help define the divergent cellular processes involved in the control of intracellular HCV replication by these agents, we have characterized the activity of monotherapies and combination therapies with the major types of human interferons against HCV replication in the HCV replicon-containing cell line, AVA5. IFN-alpha, IFN-beta, and omega interferon (IFN-omega) were equally effective at inhibiting HCV replication, while IFN-gamma was approximately 10 fold more potent. In kinetic experiments, IFN-beta and IFN-gamma inhibited HCV replication more rapidly, and for a more prolonged period following the removal of treatment, than IFN-alpha. Combination interferon therapies produced enhanced anti-HCV activity in most cases, and displayed a diverse range of interactions. Mixtures of IFN-alpha and IFN-beta exhibited generally additive to slightly antagonistic interactions, IFN-alpha or IFN-beta combined with IFN-omega were strongly antagonistic, while IFN-alpha/IFN-gamma and IFN-beta/IFN-gamma combinations displayed the most enhanced and strongly synergistic antiviral effects. Simultaneous administration of interferons in the combination treatments was found to be superior to sequential administration. Ribavirin did not exhibit any selective anti-HCV activity in cell culture, consistent with in vivo monotherapies, and did not influence the effectiveness of IFN-alpha in combination treatments. A panel of human cytokines and immune response modifiers induced by interferon and ribavirin therapies in vivo did not demonstrate anti HCV activity in HCV replicon-containing cultures. Combination therapy can be effectively modeled using HCV replicon technology yielding potentially more effective treatment regimens. HCV replicon technology has potential utility in designing combination therapies to significantly enhance the anti-HCV activity of IFN-alpha. PMID- 15652969 TI - The non-nucleoside antiviral, BAY 38-4766, protects against cytomegalovirus (CMV) disease and mortality in immunocompromised guinea pigs. AB - New antiviral drugs are needed for the treatment of cytomegalovirus (CMV) infections, particularly in immunocompromised patients. These studies evaluated the in vitro and in vivo activity of the non-nucleosidic CMV inhibitor, BAY 38 4766, against guinea pig cytomegalovirus (GPCMV). Plaque reduction assays indicated that BAY 38-4766 was active against GPCMV, with an IC(50) of 0.5muM. Yield reduction assays demonstrated an ED(90) and ED(99) of 0.4 and 0.6muM, respectively, of BAY 38-4766 against GPCMV. Guinea pigs tolerated oral administration of 50mg/kg/day of BAY 38-4766 without evidence of biochemical or hematologic toxicity. Plasma concentrations of BAY 38-4766 were high following oral dosing, with a mean peak level at 1-h post-dose of 26.7mg/ml (n=6; range, 17.8-35.4). Treatment with BAY 38-4766 reduced both viremia and DNAemia, as determined by a real-time PCR assay, following GPCMV infection of cyclophosphamide-immunosuppressed strain 2 guinea pigs (p<0.05, Mann-Whitney test). BAY 38-4766 also reduced mortality following lethal GPCMV challenge in immunosuppressed Hartley guinea pigs, from 83% (20/24) in placebo-treated guinea pigs, to 17% (4/24) in BAY 38-4766-treated animals (p<0.0001, Fisher's exact test). Mortality differences were accompanied by reduction in DNAemia in Hartley guinea pigs. Based upon its favorable safety, pharmacokinetic, and therapeutic profiles, BAY 38-4766 warrants further investigation in the GPCMV model. PMID- 15652970 TI - Inhibition of SARS-CoV replication by siRNA. AB - Serious outbreaks of severe acute respiratory syndrome (SARS), caused by the newly discovered coronavirus SARS-CoV, occurred between late 2002 and early 2003 and there is an urgent need for effective antiviral agents. RNA interference in animals and post-transcriptional gene silencing plants is mediated by small double-stranded RNA molecules named small interfering RNA (siRNA). Recently, siRNA-induced RNA interference(RNAi) may provide a new approach to therapy for pathogenic viruses, e.g. HIV and HCV. In this study, the silencing potential of seven synthetic siRNAs against SARS-CoV leader, TRS, 3'-UTR and Spike coding sequence have been applied to explore the possibility for prevention of SARS-CoV infection. We demonstrate that siRNAs directed against Spike sequences and the 3' UTR can inhibit the replication of SARS-CoV in Vero-E6 cells, and holds out promise for the development of an effective antiviral agent against SARS-CoV. PMID- 15652971 TI - Comparative efficacy of acyclovir and vidarabine on the replication of varicella zoster virus. AB - Acyclovir and less frequently, vidarabine are (or have been) used in the treatment of varicella-zoster virus (VZV) infection and are administered either intravenously (vidarabine) or orally (acyclovir, up to five times per day). The pharmacological bases of the administration interval were modeled in vitro in this study. Incubation of VZV-infected cultures with acyclovir or vidarabine for 24, 48, 72 and 96h showed similar duration-dependent anti-viral activities as assessed by a plaque-reduction assay. Treatment with vidarabine for only 8h/day for 4 days (intermittent treatment) showed anti-VZV activity equivalent to that of continuous treatment for 4 days in terms of the inhibitory dose that reduced plaque formation by 50% (IC(50)). In contrast, intermittent treatment with acyclovir exhibited a 7.9 times higher IC(50) value than that of continuous treatment. The mode of inhibition of expression of most of viral protein was similar in both drugs, but the degree of inhibition was different for each protein. Thus, vidarabine with a limited period of treatment showed anti-VZV activity comparable to continuous treatment with acyclovir, indicating the longer duration of anti-viral activity of vidarabine. PMID- 15652973 TI - Xylanases, xylanase families and extremophilic xylanases. AB - Xylanases are hydrolytic enzymes which randomly cleave the beta 1,4 backbone of the complex plant cell wall polysaccharide xylan. Diverse forms of these enzymes exist, displaying varying folds, mechanisms of action, substrate specificities, hydrolytic activities (yields, rates and products) and physicochemical characteristics. Research has mainly focused on only two of the xylanase containing glycoside hydrolase families, namely families 10 and 11, yet enzymes with xylanase activity belonging to families 5, 7, 8 and 43 have also been identified and studied, albeit to a lesser extent. Driven by industrial demands for enzymes that can operate under process conditions, a number of extremophilic xylanases have been isolated, in particular those from thermophiles, alkaliphiles and acidiphiles, while little attention has been paid to cold-adapted xylanases. Here, the diverse physicochemical and functional characteristics, as well as the folds and mechanisms of action of all six xylanase containing families will be discussed. The adaptation strategies of the extremophilic xylanases isolated to date and the potential industrial applications of these enzymes will also be presented. PMID- 15652974 TI - Compartmentalization of prokaryotic DNA replication. AB - It becomes now apparent that prokaryotic DNA replication takes place at specific intracellular locations. Early studies indicated that chromosomal DNA replication, as well as plasmid and viral DNA replication, occurs in close association with the bacterial membrane. Moreover, over the last several years, it has been shown that some replication proteins and specific DNA sequences are localized to particular subcellular regions in bacteria, supporting the existence of replication compartments. Although the mechanisms underlying compartmentalization of prokaryotic DNA replication are largely unknown, the docking of replication factors to large organizing structures may be important for the assembly of active replication complexes. In this article, we review the current state of this subject in two bacterial species, Escherichia coli and Bacillus subtilis, focusing our attention in both chromosomal and extrachromosomal DNA replication. A comparison with eukaryotic systems is also presented. PMID- 15652975 TI - "Microthrix parvicella", a filamentous bacterium causing bulking and foaming in activated sludge systems: a review of current knowledge. AB - This review summarizes the microbiology and physiology of "Microthrix parvicella" and the methods of its growth control in activated sludge wastewater treatment plants. This filamentous bacterium is of high interest because of its worldwide involvement in severe bulking and foaming at wastewater treatment plants. We present a critical analysis of physiological and kinetic data on "M. parvicella" and discuss its growth and storage abilities in various environments with the aim of understanding the strategies of this organism to successfully compete with other bacteria in activated sludge. Additionally, this review elaborates on research needs for defining reliable control strategies of bulking and foaming based on key features of "M. parvicella". PMID- 15652976 TI - The Icm/Dot type-IV secretion systems of Legionella pneumophila and Coxiella burnetii. AB - Type-IV secretion systems are devices present in a wide range of bacteria (including bacterial pathogens) that deliver macromolecules (proteins and single strand-DNA) across kingdom barriers (as well as between bacteria and into the surroundings). The type-IV secretion systems were divided into two subgroups and Legionella pneumophila and Coxiella burnetii are the only two bacteria known today to utilize a type-IVB secretion system for pathogenesis. In this review we summarized the available information concerning the icm/dot type-IVB secretion systems by comparing the two bacteria that possess this system, the proteins components of their systems as well as the homology of proteins from type-IVB secretion systems to proteins from type-IVA secretion systems. In addition, the phenotypes associated with mutants in the L. pneumophila icm/dot genes, their relations to properties of specific Icm/Dot proteins as well as the protein substrates delivered by this system are described. PMID- 15652977 TI - Enteropathogenic Escherichia coli: unravelling pathogenesis. AB - Enteropathogenic Escherichia coli (EPEC) is a gram-negative bacterial pathogen that adheres to intestinal epithelial cells, causing diarrhoea. It constitutes a significant risk to human health and remains an important cause of infant mortality in developing countries. Although EPEC was the first E. coli strain to be implicated in human disease in the 1940s and 1950s, the mechanisms by which this pathogen induced diarrhoea remained a complete mystery throughout most of the 40 years since its description. It was only during the late 1980s that major advances were made in unravelling the mechanisms behind EPEC pathogenesis. Ever since, progress has been made at a stunning pace and there have been major breakthroughs in identifying the bacterial factors involved in attaching and effacing (A/E) lesion formation, host signal transduction pathways in response to EPEC infection and the genetic basis of EPEC pathogenesis. The rapid pace of discovery is a result of intensive research by investigators in this field and portends that EPEC will soon be among one of the most understood diarrhoea causing infectious agents. This review aims to trace the progress of EPEC research since its existence was first reported by John Bray in 1945, highlighting the major findings that have revolutionised our understanding of EPEC pathogenesis. PMID- 15652978 TI - Some lessons from Rickettsia genomics. AB - Sequencing of the Rickettsia conorii genome and its comparison with its closest sequenced pathogenic relative, i.e., Rickettsia prowazekii, provided powerful insights into the evolution of these microbial pathogens. However, advances in our knowledge of rickettsial diseases are still hindered by the difficulty of working with strict intracellular bacteria and their hosts. Information gained from comparing the genomes of closely related organisms will shed new light on proteins susceptible to be targeted in specific diagnostic assays, by new antimicrobial drugs, and that could be employed in the generation of future rickettsial vaccines. In this review we present a detailed comparison of the metabolic pathways of these bacteria as well as the polymorphisms of their membrane proteins, transporters and putative virulence factors. Environmental adaptation of Rickettsia is also discussed. PMID- 15652979 TI - Iron and heme utilization in Porphyromonas gingivalis. AB - Porphyromonas gingivalis is a Gram-negative anaerobic bacterium associated with the initiation and progression of adult periodontal disease. Iron is utilized by this pathogen in the form of heme and has been shown to play an essential role in its growth and virulence. Recently, considerable attention has been given to the characterization of various secreted and surface-associated proteins of P. gingivalis and their contribution to virulence. In particular, the properties of proteins involved in the uptake of iron and heme have been extensively studied. Unlike other Gram-negative bacteria, P. gingivalis does not produce siderophores. Instead it employs specific outer membrane receptors, proteases (particularly gingipains), and lipoproteins to acquire iron/heme. In this review, we will focus on the diverse mechanisms of iron and heme acquisition in P. gingivalis. Specific proteins involved in iron and heme capture will be described. In addition, we will discuss new genes for iron/heme utilization identified by nucleotide sequencing of the P. gingivalis W83 genome. Putative iron- and heme-responsive gene regulation in P. gingivalis will be discussed. We will also examine the significance of heme/hemoglobin acquisition for the virulence of this pathogen. PMID- 15652980 TI - Opposite effects of noradrenaline and acetylcholine upon hypocretin/orexin versus melanin concentrating hormone neurons in rat hypothalamic slices. AB - Hypocretin/orexin (Hcrt/Orx) and melanin concentrating hormone (MCH) are peptides contained in overlapping cell groups of the lateral hypothalamus and commonly involved in regulating sleep-wake states and energy balance, though likely in different ways. To see if these neurons are similarly or differentially modulated by neurotransmitters of the major brainstem arousal systems, the effects of noradrenaline (NA) and carbachol, a cholinergic agonist, were examined on identified Hcrt/Orx and MCH neurons in rat hypothalamic slices. Whereas both agonists depolarized and excited Hcrt/Orx neurons, they both hyperpolarized MCH neurons by direct postsynaptic actions. According to the activity profiles of the noradrenergic locus coeruleus and cholinergic pontomesencephalic neurons across the sleep-waking cycle, the Hcrt/Orx neurons would be excited by NA and acetylcholine (ACh) and thus active during arousal, whereas the MCH neurons would be inhibited by NA and ACh and thus inactive during arousal while disinhibited and possibly active during slow wave sleep. According to the present pharmacological results, Hcrt/Orx neurons may thus stimulate arousal in tandem with other arousal systems, whereas MCH neurons may function in opposition with other arousal systems and thus potentially dampen arousal to promote sleep. PMID- 15652981 TI - Design-based stereology in neuroscience. AB - Quantitative morphology of the CNS has recently undergone major developments. In particular, several new approaches, known as design-based stereologic methods, have become available and have been successfully applied to neuromorphological research. However, much confusion and uncertainty remains about the meaning, implications, and advantages of these design-based stereologic methods. The objective of this review is to provide some clarification. It does not comprise a full description of all stereologic methods available. Rather, it is written by users for users, provides the reader with a guided tour through the relevant literature. It has been the experience of the authors that most neuroscientists potentially interested in design-based stereology need to analyze volumes of brain regions, numbers of cells (neurons, glial cells) within these brain regions, mean volumes (nuclear, perikaryal) of these cells, length densities of linear biological structures such as vessels and nerve fibers within brain regions, and the cytoarchitecture of brain regions (i.e. the spatial distribution of cells within a region of interest). Therefore, a comprehensive introduction to design-based stereologic methods for estimating these parameters is provided. It is demonstrated that results obtained with design-based stereology are representative for the entire brain region of interest, and are independent of the size, shape, spatial orientation, and spatial distribution of the cells to be investigated. Also, it is shown that bias (i.e. systematic error) in results obtained with design-based stereology can be limited to a minimum, and that it is possible to assess the variability of these results. These characteristics establish the advantages of design-based stereologic methods in quantitative neuromorphology. PMID- 15652982 TI - Dissociation of the effect of spatial behaviors on the phosphorylation of cAMP response element binding protein (CREB) within the nucleus accumbens. AB - Several studies have reported a role for the nucleus accumbens (NAcc) in learning and synaptic plasticity. Many of them suggest that the NAcc is involved in translating cortico-limbic information to the motor system mediating spatial learning and memory processes. Previous studies from our laboratory have shown that protein kinase C is activated following training in a food search spatial learning task. The present study further characterizes the molecular substrates associated with NAcc-dependent spatial behavior. The cyclic AMP-response element binding protein (CREB), a transcription factor implicated in the formation of long-term memory, was studied in the NAcc following spatial training in a food search spatial learning task. Western blots were performed to detect phosphorylated (activated) and total CREB protein levels. Our results show that CREB is significantly phosphorylated in the NAcc 48 h after habituation and at 5 min and 1 h after the first spatial training session in comparison with the naive animals that remained in their home cages. Since published data show that NAcc plays a role in novelty detection and reactivity, we conducted further experiments in order to dissociate the effect on CREB phosphorylation and expression of spatial novelty (single exposure), exploration, and spatial learning in the food search apparatus. Results show that CREB phosphorylation is significantly increased 48 h after exposure to a novel environment. The present study suggests that CREB phosphorylation observed in the NAcc during habituation and spatial training may be mainly triggered by detection of spatial novelty. PMID- 15652983 TI - A role for adult neurogenesis in spatial long-term memory. AB - Adult hippocampal neurogenesis has been linked to learning but details of the relationship between neuronal production and memory formation remain unknown. Using low dose irradiation to inhibit adult hippocampal neurogenesis we show that new neurons aged 4-28 days old at the time of training are required for long-term memory in a spatial version of the water maze. This effect of irradiation was specific since long-term memory for a visibly cued platform remained intact. Furthermore, irradiation just before or after water maze training had no effect on learning or long-term memory. Relationships between learning and new neuron survival, as well as proliferation, were investigated but found non-significant. These results suggest a new role for adult neurogenesis in the formation and/or consolidation of long-term, hippocampus-dependent, spatial memories. PMID- 15652984 TI - The differential response of astrocytes within the vestibular and cochlear nuclei following unilateral labyrinthectomy or vestibular afferent activity blockade by transtympanic tetrodotoxin injection in the rat. AB - In this study, we investigated whether changes in the vestibular neuronal activity per se influence the pattern of astrocytes morphology, glial fibrillary acidic protein (GFAP) expression and ultimately their activation within the vestibular nuclei after unilateral transtympanic tetrodotoxin (TTX) injections and after unilateral inner ear lesion. The rationale was that, theoretically the noninvasive pharmacological functional blockade of peripheral vestibular inputs with TTX, allowed us to dissociate the signals exclusively related to the shutdown of the resting activity of the first-order vestibular neurons and from neuronal signals associated with trans-ganglionic changes in first order vestibular neurons induced by unilateral labyrinthectomy (UL). Since the cochlea was removed during the surgical procedure, we also studied the astrocytic reaction within the deafferented cochlear nuclei. No significant changes in the distribution or relative levels of GFAP mRNA expression, relative levels of GFAP protein or immunoreactivity for GFAP were found in the ipsilateral vestibular nuclei at any post-TTX injection times studied. In addition, no sign of microglia activation was observed. In contrast, a robust increase of the distribution and relative levels of GFAP mRNA expression, protein levels and immunoreactivity was observed in the deafferented vestibular and cochlear nuclei beginning at 1 day after inner ear lesion. GFAP mRNA expression and immunoreactivity in the cochlear nucleus was qualitatively stronger than in the ipsilateral vestibular nuclei. The results suggest that astrocyte activation in the vestibular nuclei is not related to drastic changes of vestibular nuclei neuronal activity per se. Early trans ganglionic changes due to vestibular nerve dendrites lesion provoked by the mechanical destruction of vestibular receptors, most probably induced the glial reaction. Its functional role in the vestibular compensation process remains to be elucidated. PMID- 15652985 TI - Pathological cholesterol metabolism fails to modify electrophysiological properties of afflicted neurones in Niemann-Pick disease type C. AB - Niemann-Pick disease type C (NPC) is a recessive inherited neurovisceral lipid storage disease characterized by progressive motor impairment and a loss of neurones including those integrated into the motor system. One of the key neuropathological findings is the intracellular accumulation of lysosomes enriched with free cholesterol. This accumulation is due to impaired transport proteins named NPC1 (approx. 95% of the cases) or NPC2 (approx. 5%) responsible for the transport of endocytosed cholesterol from lysomes to plasma membranes. The perturbed lipid-transport in NPC cells leads to an altered lipid composition of the plasma membrane. Available evidence suggests that the lipid matrix influences the electrophysical properties of ion channels in membranes. We therefore evaluated whether electrophysiological properties of NPC neurones differ from healthy neurones. Both, acute brain slices and primary neuronal cell cultures from wildtype and NPC mice, a well-established mouse model for the Niemann-Pick type C disease, were used for a comparison of electrophysiological properties like resting membrane potential, input resistance, action potential amplitudes and synaptic properties of the neurones. In addition we optically recorded the changes of intraneuronal calcium levels elicited by depolarization. Our results show that the characteristics of ion channels in NPC neurones do not differ significantly from wildtype neurones. We therefore conclude that gross alterations of the electrophysiological properties of neurones will probably not initiate or substantially contribute to the development of the motor impairment or other neurological signs of NPC. PMID- 15652986 TI - Prolonged local neurotrophin-3 infusion reduces ipsilateral collateral sprouting of spared corticospinal axons in adult rats. AB - The corticospinal tract is widely used to study regeneration and is essential for voluntary movements in humans. In young rats, corticospinal axons on the uninjured side sprout and grow into the denervated side. Neurotrophin-3 (NT-3) induces such crossed collateral sprouting in adults. We investigated whether local intraspinal NT-3 infusions would promote collateral sprouting of spared corticospinal terminals from within a partially denervated side, as this would be more appropriate for enhancing function of unilateral and specific movements. Adult rats received a partial bilateral transection of the pyramids, leaving approximately 40% of each tract intact. Vehicle or vehicle plus NT-3 (3 or 10 microg/day) was infused for 14 days into the left side of the cervical (C5/6) or lumbar (L2) cord. The corticospinal processes on the left side were anterogradely traced with cholera toxin B (CTB; which labeled gray matter processes more robustly than biotinylated dextran amine) injected into the front or hind limb area of the right sensorimotor cortex, respectively, 3 days before analysis. Unexpectedly, approximately 40% fewer CTB-labeled corticospinal processes were detectable in the cervical or lumbar gray matter of NT-3-treated rats than in vehicle-infused ones. Vehicle-infused injured rats had more corticospinal processes in the center of the cord than normal rats, evidence for lesion-induced collateral sprouting. NT-3 caused sprouting of local calcitonin gene-related peptide-positive fibers. These results suggest that NT-3 reduces collateral sprouting of spared corticospinal axons from within the denervated regions, possibly because of the injury environment or by increasing sprouting of local afferents. They identify an unexpected context-dependent outgrowth inhibitory effect of NT-3. PMID- 15652987 TI - Modulation of available vesicles and release kinetics at the inhibitor of the crayfish neuromuscular junction. AB - We have investigated the effect of serotonin (5-HT) and okadaic acid (OA) on presynaptic processes at the crayfish inhibitory neuromuscular junction. Two different physiological parameters of transmitter release were examined: release kinetics and the size of the readily releasable pool of vesicles (RRP). Using a paired pulse stimulus and high frequency trains, we established that a single broad action potential, recorded in 20 mM tetraethylammonium and 1 mM 4-amino pyridine, released the RRP in its entirety. Thus, by measuring the amplitude of inhibitory postsynaptic potential (IPSC) we were able to directly assess the effects of 5-HT and OA on the RRP. Serotonin at 200 nM and OA at 2.5 microM each significantly increased IPSC above control levels and the effects of these two modulators were comparable. Both modulators also induced a leftward shift in the rising phase of IPSC, i.e. an apparent acceleration in release kinetics. The shift caused by OA was significantly more pronounced than that induced by 5-HT. This apparent acceleration in release was not associated with a corresponding change in the presynaptic Ca2+ transient measured at a 2 kHz resolution, suggesting that modulation was not due to an acceleration in Ca2+ channel kinetics. In view of the comparable increase in the size of the RRP by the modulators, the differential modulation of release kinetics suggests that these two parameters may be modulated by separate biochemical processes. PMID- 15652988 TI - The effect of peripheral nerve injury on disease progression in the SOD1(G93A) mouse model of amyotrophic lateral sclerosis. AB - Around 20% of familial cases of amyotrophic lateral sclerosis have been shown to carry mutations in Cu/Zn superoxide dismutase 1 (Cu/Zn SOD1). Transgenic mice over-expressing human mutant SOD1 genes have been developed and in this study we examined the effect of nerve injury on disease progression in these mice. Firstly, disease progression in uninjured mice was characterised using physiological methods. Muscle force, contractile characteristics and motor unit survival was established at 90 days, an early symptomatic stage and also at the end-stage of the disease, at 130 days. In addition, muscle histochemistry was examined and the extent of motoneuron survival established morphologically. By 90 days of age, there is a significant reduction in muscle force, and nearly 40% of motoneurons within the sciatic motor pool have already died. By 130 days, the muscles are significantly weaker, and there is a dramatic change in the phenotype of extensor digitorum longus (EDL), which changes from a fast fatigable muscle, to a fatigue resistant muscle with a high oxidative capacity. By this stage of the disease, only 40% of motor units in EDL survive, with only 29% of motoneurons surviving within the sciatic motor pool. Following injury to the sciatic nerve in SOD1(G93A) mice, there is an acceleration in disease progression so that 90 day old mice show deficits that are only seen at the end stage in uninjured SOD1(G93A) mice. It is therefore possible that mutant SOD1 toxicity increases the vulnerability of motoneurons and muscles to stressful stimuli such as nerve injury. PMID- 15652989 TI - Hemispheric asymmetry, modular variability and age-related changes in the human entorhinal cortex. AB - The verrucae areae entorhinalis (VAE) are a characteristic feature of the human brain that occupy the anterior and posterolateral parts of the parahippocampal gyri and correspond to the islands of layer II neurons. We analyzed VAE in 60 neurologically normal subjects ranging from 23 to 85 years of age using a casting method. In 10 of these subjects the total number of neurons in the entorhinal islands was estimated stereologically using the optical fractionator. The number and surface area of VAE were higher in the left hemisphere compared with the right, and this leftward asymmetry was highly significant. Regression analysis showed a negative correlation between average VAE area and age in both hemispheres, representing a rate loss of about 800 microm2 per year. The estimated number of neurons obtained with the optical fractionator showed no significant difference between the left and the right hemisphere (468,000+/ 144,000 vs. 405,000+/-117,000). There was a highly significant negative correlation between neuron numbers and age in both sides. In addition, clusters of small, undifferentiated layer II neurons ('heterotopias') were frequently observed in the rostral part of the entorhinal cortex in young and elderly adults. Layer II entorhinal neurons are among the first to show neurofibrillary changes during normal aging. The present data confirm the occurrence of age related neuron loss in the entorhinal cortex. Considering the consistent projections from ipsilateral auditory association areas that, together with Broca's motor-speech area (Brodmann areas 44 and 45), show leftward asymmetry from early infancy (such as Brodmann area 22, planum temporale, and area 52 in the long insular gyrus), we speculate that functional lateralization of the human entorhinal cortex may be associated with specialization for memory processing related to language. Due to the dependence of hippocampal formation on entorhinal projections, this finding is also consistent with the greater capacity of the left hippocampus for verbal episodic memory. PMID- 15652990 TI - Activation of metabotropic glutamate receptor 3 enhances interleukin (IL)-1beta stimulated release of IL-6 in cultured human astrocytes. AB - Previous studies have demonstrated that human astrocytes express mRNA and receptor protein for group I and II metabotropic glutamate receptors (mGluRs). Whether these receptors can influence the inflammatory and immune response and can modulate the capacity of astrocytes to produce inflammatory cytokines is still unclear. Inflammatory cytokines can be produced by activated glial cells and play a critical role in several neurological disorders. Astrocyte-enriched human cell cultures growing in a serum-free chemically defined medium were used to study the regulation of IL (interleukin)-1beta and IL-6 in response to mGluR activation. Astrocytes cultured in the absence or in the presence of epidermal growth factor (EGF), did not secrete significant IL-1beta and IL-6, as determined by specific enzyme-linked immunosorbent assay (ELISA). Activation of mGluRs using (S)-3,5-dihydroxyphenylglycine (DHPG; selective group I agonist) or DCG-IV (selective group II agonist) did not affect the production of interleukins under both growth conditions. On exposure to IL-1beta high levels of IL-6 were detected. Activation of mGluR3 with DCG-IV (but not of mGluR5 with DHPG) enhanced, in the presence of IL-1beta, the release of IL-6 in a dose dependent manner in astrocytes cultured under conditions (+EGF) in which the mGluR expression is known to be upregulated. The effect of mGluR3 activation on IL 1beta stimulated release of IL-6 was prevented by selective group II mGluR antagonists. The capacity of mGluR3 to modulate the release of IL-6 in the presence of IL-1beta supports the possible involvement of this receptor subtype in the regulation of the inflammatory and immune response under pathological conditions associated with glial cell activation. PMID- 15652991 TI - Prenatal protracted irradiation at very low dose rate induces severe neuronal loss in rat hippocampus and cerebellum. AB - Prenatal irradiation is known to damage the developing brain. However, little is known about the consequences of very low dose rate prenatal protracted irradiation over several days on neuron numbers in the offspring brain, and on volumes of the corresponding brain regions. Pregnant Wistar rats were exposed either to a protracted gamma irradiation from embryonic day (E) 13 to E16 (0.7 mGy/min; total cumulative dose approximately 3 Gy) or were sham-irradiated. Thirty months old male and female offspring were then analyzed for alterations in hippocampal and cerebellar morphology. Using design-based stereology and the analysis of sets of sections systematically and randomly sampled to span the entire brain region of interest, a statistically significant decrease in numbers of hippocampal pyramidal and granule cells as well as of cerebellar Purkinje and granule cells (approximately 50%) was found in male and female irradiated offspring. The volumes of these brain regions were comparably altered. The analysis of only a "representative" section per animal yielded mostly non significant trends. Evaluation of neuron densities showed no differences between prenatally irradiated and sham-irradiated offspring. Most importantly, very low dose rate prenatal protracted gamma irradiation did not result in the same morphologic alterations in the offspring brain as previously observed after prenatal single irradiation such as derangement of the laminar structure of pyramidal cells within the hippocampus or malformation of cerebellar lobules. PMID- 15652992 TI - Hypoxia modifies nuclear calcium uptake pathways in the cerebral cortex of the guinea-pig fetus. AB - Nuclear Ca2+ signals are thought to play a critical role in the initiation and progression of programmed cell death. The present study tests the hypothesis that hypoxia alters nuclear Ca2+ transport pathways and leads to an increase in nuclear Ca(2+)-influx in cerebral cortical neuronal nuclei. To test this hypothesis the effect of tissue hypoxia on high affinity Ca(2+)-ATPase activity and the binding characteristics of inositol 1,4,5-triphosphate (IP3) and inositol 1,3,4,5-tetrakisphosphate (IP4) receptors were studied in neuronal nuclei from the cerebral cortex of guinea-pig fetuses. Results show increased high-affinity Ca(2+)-ATPase activity (nmol/mg protein/h) in the hypoxic group 969.7+/-79 as compared with 602.4+/-90.9 in the normoxic group, P<0.05. The number of IP3 receptors (Bmax, fmol/mg protein) increased from 61+/-21 in the normoxic group to 164+/-49 in the hypoxic group, P<0.05. K(d) values did not change following hypoxia. In contrast, IP4 receptor Bmax (fmol/mg protein) and K(d) (nM) values increased from 360+/-32 in the normoxic group to 626+/-136 in the hypoxic group (P<0.001) and, from 26+/-1 in the normoxic group to 61+/-9 in the hypoxic group (P<0.001), respectively. 45Ca(2+)-influx (pmol/mg protein) significantly increased from 6.3+/-1.9 in the normoxic group to 10.9+/-1.1 the hypoxic group (P<0.001). The data show that hypoxia modifies nuclear Ca2+ transport pathways and results in increased nuclear Ca(2+)-influx. We speculate that hypoxia increases nuclear Ca2+ uptake from the cytoplasm to the nucleoplasm, resulting in increased transcription of proapoptotic genes and subsequent activation of programmed cell death pathways. PMID- 15652993 TI - Recurrent insulin-induced hypoglycemia causes site-specific patterns of habituation or amplification of CNS neuronal genomic activation. AB - Antecedent hypoglycemia is a primary factor in hypoglycemia-associated autonomic failure, a pathophysiological condition characterized by impaired glucose counterregulatory function. Conventional therapeutic strategies involving administration of intermediate dosage-release formulations of insulin in the management of insulin-dependent diabetes mellitus result in frequent iatrogenic hypoglycemia. This study investigated the neuroanatomical location, direction, and magnitude of CNS neuronal genomic activation by singular versus repeated induction of hypoglycemic bouts of greater than 6 h duration achieved by administration of the intermediate-acting insulin, humulin neutral protamine Hagedorn (NPH). Adult male rats injected subcutaneously with Humulin NPH exhibited robust immunolabeling for the nuclear transcription factor, Fos, in discrete telencephalic, diencephalic, midbrain, and caudal hindbrain loci in a pattern that was not identical to that described for regular insulin. Administration of four doses of insulin on as many days significantly diminished or extinguished Fos immunostaining within the parvocellular hypothalamic paraventricular nucleus, lateral hypothalamic area, dorsomedial hypothalamic nucleus, thalamic paraventricular nucleus, nucleus tractus solitarius, and area postrema, but did not modify labeling of other metabolic loci. However, numbers of Fos-immunoreactivity-positive magnocellular neurons in the hypothalamic paraventricular and supraoptic nuclei were significantly increased after the second and fourth insulin doses, relative to the single-dose group. Concurrent observations of exacerbated hypoglycemia and modified patterns of glucoregulatory hormone secretion after serial injections of intermediate-acting insulin suggest that central mechanisms governing compensatory endocrine responses, specifically glucagon, become habituated to repetitive hypoglycemia of extended duration. Resultant alterations in CNS-islet and -adrenomedullary output and hypothalamic pituitary-adrenal activity may reflect diminished neuronal activation within one or more of the brain loci characterized here by nonuniform transcriptional activation. The current studies provide a neuroanatomical foundation for further investigation of the neurochemical phenotypes and interconnectivity of functionally adaptive neurons, underlying cellular and molecular mechanisms of diminished or enhanced activation, as well as the impact of these modified cellular responses on glucose counterregulation during administration of intermediate-acting insulin. PMID- 15652994 TI - Androgen dependence in hamsters: overdose, tolerance, and potential opioidergic mechanisms. AB - Anabolic steroids are drugs of abuse. However, the potential for steroid reward and addiction remains largely unexplored. This study used i.c.v. testosterone self-administration and controlled infusions of testosterone or vehicle in hamsters to explore central mechanisms of androgen overdose. Forty-two hamsters used nose-pokes to self-administer 1 microg/microl testosterone i.c.v. 4 h/day in an operant chamber. During 1-56 days of androgen self-administration, 10 (24%) hamsters died. Deaths correlated with peak daily intake of testosterone. Of the hamsters that self-administered a peak intake of <20 microg/day, there was 100% survival (10/10). Survival decreased to 86% (19/22) when daily testosterone intake peaked at 20-60 microg/day. Only 30% (three of 10) survived when daily testosterone intake exceeded 60 microg/day. Deaths are not due to volume or vehicle because i.c.v. infusions of 80 mul vehicle had no effect. Testosterone overdose resembles opiate intoxication. When male hamsters received infusions of 40 microg testosterone, locomotion (25.1+/-18.8 grid-crossings/10 min), respiration (72.7+/-5.4 breaths/min) and body temperature (33.5+/-0.4 degrees C) were significantly reduced, compared with males receiving vehicle infusions (186.1+/-8.1 crossings/10 min, 117.6+/-1.0 breaths/min, 35.9+/-0.1 degrees C, P<0.05). However, males developed tolerance to continued daily testosterone infusion. After 15 days, locomotion (170.2+/-6.3 crossings), respiration (118.4+/ 1.3 breaths/min), and body temperature (35.3+/-0.3 degrees C) in testosterone infused males were equivalent to that in vehicle controls (P>0.05). The depressive effects of testosterone infusion are blocked by the opioid antagonist, naltrexone. With naltrexone pre-treatment (10 mg/kg s.c.), locomotion (183.7+/ 1.8 crossings/10 min), respiration (116.9+/-0.3 breaths/min), and body temperature (36.1+/-0.4 degrees C) during testosterone infusion were equivalent to vehicle controls. Likewise, naltrexone prevents the reinforcing effects of i.c.v. testosterone self-administration. These results indicate that testosterone at high doses causes central autonomic depression, which may be a factor in deaths during self-administration. As well, the depressive effects of large quantities of testosterone may be mediated, at least in part, by an opioidergic mechanism. PMID- 15652995 TI - Modafinil more effectively induces wakefulness in orexin-null mice than in wild type littermates. AB - Narcolepsy-cataplexy, a disorder of excessive sleepiness and abnormalities of rapid eye movement (REM) sleep, results from deficiency of the hypothalamic orexin (hypocretin) neuropeptides. Modafinil, an atypical wakefulness-promoting agent with an unknown mechanism of action, is used to treat hypersomnolence in these patients. Fos protein immunohistochemistry has previously demonstrated that orexin neurons are activated after modafinil administration, and it has been hypothesized that the wakefulness-promoting properties of modafinil might therefore be mediated by the neuropeptide. Here we tested this hypothesis by immunohistochemical, electroencephalographic, and behavioral methods using modafinil at doses of 0, 10, 30 and 100 mg/kg i.p. in orexin-/- mice and their wild-type littermates. We found that modafinil produced similar patterns of neuronal activation, as indicated by Fos immunohistochemistry, in both genotypes. Surprisingly, modafinil more effectively increased wakefulness time in orexin-/- mice than in the wild-type mice. This may reflect compensatory facilitation of components of central arousal in the absence of orexin in the null mice. In contrast, the compound did not suppress direct transitions from wakefulness to REM sleep, a sign of narcolepsy-cataplexy in mice. Spectral analysis of the electroencephalogram in awake orexin-/- mice under baseline conditions revealed reduced power in the theta; band frequencies (8-9 Hz), an index of alertness or attention during wakefulness in the rodent. Modafinil administration only partly compensated for this attention deficit in the orexin null mice. We conclude that the presence of orexin is not required for the wakefulness-prolonging action of modafinil, but orexin may mediate some of the alerting effects of the compound. PMID- 15652996 TI - Repeated nicotine exposure in rats: effects on memory function, cholinergic markers and nerve growth factor. AB - A decrease in the number of nicotinic-acetylcholine receptors (nAChRs) in the brain is thought to contribute to the cognitive dysfunction associated with diseases as diverse as Alzheimer's disease and schizophrenia. Interestingly, nicotine and similar compounds have been shown to enhance memory function and increase the expression of nAChRs and therefore, could have a therapeutic role in the aforementioned diseases. Nicotine has also been shown to exert positive effects on certain neurotrophins such as nerve growth factor (NGF), and therefore could play a role beyond mere symptomatic therapy. However, to date, comprehensive studies of nicotine's effects on the expression of specific acetylcholine (ACh) receptor subtypes, key cholinergic proteins (that are regulated by NGF) such as choline acetyltransferase (ChAT) and the vesicular ACh transporter (VAChT) are lacking. Studies to further investigate the effects of nicotine on NGF especially its high- and low-affinity receptors are also needed. In the present study, male Wistar rats exposed a relatively low dosage of nicotine (0.35 mg/kg every 12 h) for 14 days demonstrated improved memory performance (assessed in two separate water maze testing methods) when compared with controls. Autoradiographic experiments indicated that nicotine increased [3H]-epibatidine, [125I]-alpha-bungarotoxin and [3H]-AFDX384, but not [3H] pirenzepine binding sites in several learning- and memory-related brain areas. The expression of ChAT, VAChT, as well as tropomyosin-receptor kinase A (TrkA) NGF receptors and phospho-TrK receptors was increased by nicotine in the hippocampus. No changes were observed in the levels of the NGF peptide or low affinity p75 neurotrophin receptors (p75NTR), however. These results suggest that repeated exposure to nicotine results in positive effects on central cholinergic markers and memory function, which may be mediated via effects on high-affinity NGF receptors. PMID- 15652997 TI - GABA(A) receptor facilitation of neurokinin release from primary afferent terminals in the rat spinal cord. AB - Our goal was to test the following hypotheses: 1) GABA(A) receptors facilitate neurokinin release from primary afferent terminals; 2) they do this by suppressing an inhibitory effect of GABA(B) receptors; 3) the activation of these two receptors is controlled by the firing frequency of primary afferents. We evoked neurokinin release by stimulating the dorsal root attached to spinal cord slices, and measured it using neurokinin 1 receptor (NK1R) internalization. Internalization evoked by root stimulation at 1 Hz (but not at 100 Hz) was increased by the GABA(A) receptor agonists muscimol (effective concentration of drug for 50% of the increase [EC50] 3 microM) and isoguvacine (EC50 4.5 microM). Internalization evoked by root stimulation at 100 Hz was inhibited by the GABA(A) receptor antagonists bicuculline (effective concentration of drug for 50% of the inhibition [IC50] 2 microM) and picrotoxin (IC50 243 nM). Internalization evoked by incubating the root with capsaicin (to selectively recruit nociceptive fibers) was increased by isoguvacine and abolished by picrotoxin. Therefore, GABA(A) receptors facilitate neurokinin release. Isoguvacine-facilitated neurokinin release was inhibited by picrotoxin, low Cl-, low Ca2+, Ca2+ channel blockers and N-methyl-D-aspartate receptor antagonists. Bumetanide, an inhibitor of the Na(+) K(+)-2Cl- cotransporter, inhibited isoguvacine-facilitated neurokinin release, but this could be attributed to a direct inhibition of GABA(A) receptors. The GABA(B) agonist baclofen inhibited NK1R internalization evoked by 100 Hz root stimulation (IC50 1.5 microM), whereas the GABA(B) receptor antagonist (2S)-3 [[(1S)-1-(3,4-dichlorophenyl)ethyl]amino-2-hydroxypropyl](phenylmethyl) phosphinic acid (CGP-55845) increased NK1R internalization evoked by 1 Hz root stimulation (EC50 21 nM). Importantly, baclofen inhibited isoguvacine-facilitated neurokinin release, and CGP-55845 reversed the inhibition of neurokinin release by bicuculline. In conclusion, 1) GABA(B) receptors located presynaptically in primary afferent terminals inhibit neurokinin release; 2) GABA(A) receptors located in GABAergic interneurons facilitate neurokinin release by suppressing GABA release onto these GABA(B) receptors; 3) high frequency firing of C-fibers stimulates neurokinin release by activating GABA(A) receptors and inhibiting GABA(B) receptors, whereas low frequency firing inhibits neurokinin release by the converse mechanisms. PMID- 15652998 TI - Brain oxidation is an initial process in sleep induction. AB - CNS activity is generally coupled to the vigilance state, being primarily active during wakefulness and primarily inactive during deep sleep. During periods of high neuronal activity, a significant volume of oxygen is used to maintain neuronal membrane potentials, which subsequently produces cytotoxic reactive oxygen species (ROS). Glutathione, a major endogenous antioxidant, is an important factor protecting against ROS-mediated neuronal degeneration. Glutathione has also been proposed to be a sleep-promoting substance, yet the relationship between sleep and cerebral oxidation remains unclear. Here we report that i.c.v. infusion of the organic peroxide t-butyl-hydroperoxide at a concentration below that triggering neurodegeneration (0.1 micromol/100 microl/10 h) promotes sleep in rats. Also, microinjection (2 nmol, 2 microl) or microdialysis (100 microM, 20 min) of t-butyl-hydroperoxide into the preoptic/anterior hypothalamus (POAH) induces the release of the sleep-inducing neuromodulators, nitric oxide and adenosine, without causing neurodegeneration. Nitric oxide and adenosine release was inhibited by co-dialysis of the N-methyl-D aspartate receptor antagonist, d(-)-2-amino-5-phosphonopentanoic acid (D-AP5; 1 mM), suggesting that glutamate-induced neuronal excitation mediates the peroxide induced release of nitric oxide and adenosine. Indeed, Ca2+ release from mitochondria and delayed-onset Ca2+ influx via N-methyl-D-aspartate receptors was visualized during peroxide exposure using Ca2+ indicator proteins (YC-2.1 and mitochondrial-targeted Pericam) expressed in organotypic cultures of the POAH. In the in vitro models, t-butyl-hydroperoxide (50 microM) causes dendritic swelling followed by the intracellular Ca2+ mobilization, and D-AP5 (100 microM) or glutathione (500 microM) inhibited t-butyl-hydroperoxide-induced intracellular Ca2+ mobilization and protected POAH neurons from oxidative stress. These data suggest that low-level subcortical oxidation under the control of an antioxidant system may trigger sleep via the Ca(2+)-dependent release of sleep-inducing neuromodulators in the POAH, and thus we propose that a moderate increase of ROS during wakefulness in the neuronal circuits regulating sleep may be an initial trigger in sleep induction. PMID- 15652999 TI - Periodic oscillatory activity in parahippocampal slices maintained in vitro. AB - Brain slices maintained in vitro have been extensively used for studying neuronal synchronization. However, the validity of this approach may be questioned since pharmacological procedures are usually required to elicit spontaneous events similar to the EEG activity recorded in vivo. Here, we report that when superfused with control medium, rat brain slices comprising the entorhinal and perirhinal cortices along with a portion of the basolateral/lateral nuclei of the amygdala can synchronously generate periodic oscillatory activity at 5-11 Hz every 5-30 s. The periodic events: (i) correspond intracellularly to synaptic depolarizations in regularly firing neurons analyzed in the three areas; (ii) have no fixed site of onset; (iii) spread with time lags of 8-20 ms; and (iv) continue to occur asynchronously after their surgical isolation. NMDA receptor antagonism reduced the duration of the oscillatory events, while glutamatergic non-NMDA receptor antagonism abolished them. Activation of mu-opioid receptors, a procedure that hyperpolarizes interneurons thus decreasing GABA release, reversibly decreased the rate of occurrence of periodic oscillatory activity (POA). However, periodic events continued to occur during application of GABA(A) or GABA(B) receptor antagonists as well as in the presence of the cholinergic agent carbachol. We also found that POA was abolished by baclofen and irreversibly reduced by the gap junction decoupler carbenoxolone. These findings demonstrate that parahippocampal networks in a brain slice preparation can generate periodic, synchronous activity under quasi-physiological conditions. These network oscillations (i) reflect the activation of ionotropic glutamatergic and GABAergic receptors, (ii) are contributed by gap-junction interactions, and (iii) are controlled by GABA(B) receptors that are presumably located presynaptically. PMID- 15653000 TI - Functional mapping of the prosencephalic systems involved in organizing predatory behavior in rats. AB - The study of the neural basis of predatory behavior has been largely neglected over the recent years. Using an ethologically based approach, we presently delineate the prosencephalic systems mobilized during predation by examining Fos immunoreactivity in rats performing insect hunting. These results were further compared with those obtained from animals killed after the early nocturnal surge of food ingestion. First, predatory behavior was associated with a distinct Fos up-regulation in the ventrolateral caudoputamen at intermediate rostro-caudal levels, suggesting a possible candidate to organize the stereotyped sequence of actions seen during insect hunting. Insect predation also presented conspicuous mobilization of a neural network formed by a distinct amygdalar circuit (i.e. the postpiriform-transition area, the anterior part of cortical nucleus, anterior part of basomedial nucleus, posterior part of basolateral nucleus, and medial part of central nucleus) and affiliated sites in the bed nuclei of the stria terminalis (i.e. the rhomboid nucleus) and in the hypothalamus (i.e. the parasubthalamic nucleus). Accordingly, this network is likely to encode prey related motivational values, such as prey's odor and taste, and to influence autonomic and motor control accompanying predatory eating. Notably, regular food intake was also associated with a relatively weak Fos up-regulation in this network. However, during regular surge of food intake, we observed a much larger mobilization in hypothalamic sites related to the homeostatic control of eating, namely, the arcuate nucleus and autonomic parts of the paraventricular nucleus. Overall, the present findings suggest potential neural systems involved in integrating prey-related motivational values and in organizing the stereotyped sequences of action seen during predation. Moreover, the comparison with regular food intake contrasts putative neural mechanisms controlling predatory related eating vs. regular food intake. PMID- 15653002 TI - What affects the occurrence of nocturia more: menopause or age? AB - OBJECTIVES: Prevalence studies show that nocturia is a very common condition, affecting older age groups in particular. In this study, we evaluate (1) the community-based prevalence of nocturia and its associated overactive bladder according to the patient's perception and disturbance to sleep and (2) whether age or menopausal (MP) transition has a greater effect on the occurrence of nocturia. METHODS: The questions regarding nocturia were part of the Bristol female lower urinary tract symptoms questionnaire (translated into Mandarin, the correlation coefficient r = 0.91). A total of 1253 (79.1% response rate) women were randomly sampled from a community-based female population, aged 20 years or older. Samples were subdivided into reproductive age, perimenopausal stage and elderly. If the participants reported having nocturia, they were further questioned with, "Does it interfere with your sleep or bother your daily life?" The chi-square test and the Cochran-Armitage trend test were used to analyze the data. RESULTS: Nocturia in older age groups was significantly higher than that of the younger age groups. The occurrence of nocturia was significantly associated with the elderly (compared with the reproductive age, P < 0.05). There were no significant differences in the prevalence of nocturia between the perimenopausal stage and the reproductive age (P > 0.05). CONCLUSIONS: Our data implies that the increasing occurrence of nocturia is age-related and is not affected by the transition from pre- to postmenopause. Large-scale prospective longitudinal studies need to be conducted to clarify the effects of the aging process and estrogen deficiency. PMID- 15653001 TI - Cholinergic neurotransmission in the preBotzinger Complex modulates excitability of inspiratory neurons and regulates respiratory rhythm. AB - We investigated whether there is endogenous acetylcholine (ACh) release in the preBotzinger Complex (preBotC), a medullary region hypothesized to contain neurons generating respiratory rhythm, and how endogenous ACh modulates preBotCneuronal function and regulates respiratory pattern. Using a medullary slice preparation from neonatal rat, we recorded spontaneous respiratory-related rhythm from the hypoglossal nerve roots (XIIn) and patch-clamped preBotC inspiratory neurons. Unilateral microinjection of physostigmine, an acetylcholinesterase inhibitor, into the preBotC increased the frequency of respiratory-related rhythmic activity from XIIn to 116+/-13% (mean+/-S.D.) of control. Ipsilateral physostigmine injection into the hypoglossal nucleus (XII nucleus) induced tonic activity, increased the amplitude and duration of the integrated inspiratory bursts of XIIn to 122+/-17% and 117+/-22% of control respectively; but did not alter frequency. In preBotC inspiratory neurons, bath application of physostigmine (10 microM) induced an inward current of 6.3+/-10.6 pA, increased the membrane noise, decreased the amplitude of phasic inspiratory drive current to 79+/-16% of control, increased the frequency of spontaneous excitatory postsynaptic currents to 163+/-103% and decreased the whole cell input resistance to 73+/-22% of control without affecting the threshold for generation of action potentials. Bath application of physostigmine concurrently induced tonic activity, increased the frequency, amplitude and duration of inspiratory bursts of XIIn motor output. Bath application of 4-diphenylacetoxy-N methylpiperidine methiodide (4-DAMP, 2 microM), a M3 muscarinic acetylcholine receptor (mAChR) selective antagonist, increased the input resistance of preBotC inspiratory neurons to 116+/-9% of control and blocked all of the effects of physostigmine except for the increase in respiratory frequency. Dihydro-beta erythroidine (DH-beta-E; 0.2 microM), an alpha4beta2 nicotinic receptor (nAChR) selective antagonist, blocked all the effects of physostigmine except for the increase in inspiratory burst amplitude. In the presence of both 4-DAMP and DH beta-E, physostigmine induced opposite effects, i.e. a decrease in frequency and amplitude of XIIn rhythmic activity. These results suggest that there is cholinergic neurotransmission in the preBotC which regulates respiratory frequency, and in XII nucleus which regulates tonic activity, and the amplitude and duration of inspiratory bursts of XIIn in neonatal rats. Physiologically relevant levels of ACh release, via mAChRs antagonized by 4-DAMP and nAChRs antagonized by DH-beta-E, modulate the excitability of inspiratory neurons and excitatory neurotransmission in the preBotC, consequently regulating respiratory rhythm. PMID- 15653003 TI - Levonorgestrel and 17beta-estradiol given transdermally for the prevention of postmenopausal osteoporosis. AB - AIM: To evaluate the efficacy and safety of a new transdermal continuous combined hormone replacement therapy (HRT) for the prevention of postmenopausal osteoporosis. METHODS: 212 osteopenic (lumbar spine and/or hip (femoral neck) bone mineral density (BMD) between -1.0 and -2.5 S.D. of the premenopausal mean value) postmenopausal women aged 45-65 years participated in a 2-year prospective study. Treatments were 45 microg 17beta-estradiol combined with 30 (n = 69) or 40 microg (n = 72) levonorgestrel daily or placebo (n = 71) given as a 7-day patch. All received a daily supplement of 500 mg calcium. BMD at lumbar spine (L2-L4), hip and total body, as well as blood and urinary biochemical markers of bone turnover (serum osteocalcin (sOC), serum bone-specific alkaline phosphatase (sBSAP), urinary calcium (uCa) and urinary CrossLaps (uCTX)) were measured regularly. RESULTS: BMD at the lumbar spine, hip and total body increased by 8, 6 and 3% (P < 0.001), respectively, in the hormone groups versus placebo. The bone markers all decreased accordingly (sOC: 37%, sBSAP: 34% and uCTX: 65% from baseline (all P < 0.001)), except for uCa that did not change significantly. No significant dose-related effect of levonorgestrel was found. Vaginal bleeding/spotting decreased from 48 to 25% of the HRT-treated women during the study period. Skin tolerance was good in 84% of the women with no difference between the study groups. No incidences of endometrial hyperplasia, uterine or mammary cancer occurred. CONCLUSION: The transdermal combination of 17beta estradiol and levonorgestrel has a positive effect on BMD in an osteopenic postmenopausal population. Furthermore, a high safety profile was observed. PMID- 15653004 TI - Oral dryness, nocturia and the menopause. AB - OBJECTIVE: To analyse the relationship between oral dryness (OD), nocturia, the menopausal transition and some medical conditions and medications in women. METHODS: A questionnaire study was carried out in 3669 randomly selected women (out of 6000 invited) in the County of Jamtland, Sweden. Questions were asked about occurrence of somatic diseases and symptoms, nocturia, nocturnal thirst and drinking, and the use of drugs. RESULTS: The occurrence of oral dryness was 17.8% in women before the menopause, 23.3% during the first 5 years after, 29.2% 5-9 years after and 34.5% 10 or more years after the menopause (P < 0.0001). In a multiple logistic regression analysis independent correlates for oral dryness were: nocturnal micturition, one episode versus none 1.7 (1.4-2.1); two versus none 2.4 (1.7-3.2); > or = 3 versus none 3.3 (2.1-5.2); menopausal state: <5 years after the menopause versus before 1.2 (0.9-1.6); 5-9 years after the menopause versus before 1.6 (1.2-2.1); > or = 10 years after the menopause versus before 1.7 (1.3-2.2); musculo-skeletal pain 1.6 (1.3-2.1) being on one or more prescribed drugs 1.8 (1.5-2.1). CONCLUSION: The present data show that OR is a common and disregarded symptom in women with nocturia independent of menopausal state, musculo-skeletal symptoms and medication with analgesics, diuretics, hypnotics and hormonal replacement therapy (HRT). PMID- 15653005 TI - Effects of low-dose, continuous combined hormone replacement therapy on sleep in symptomatic postmenopausal women. AB - Sleep disturbances in peri- and postmenopausal women may result from hormonal changes, vasomotor symptoms, and possibly psychological factors. Hormone replacement therapy (HRT) seems to diminish the disruption of sleep in climacteric women. The aim of this study was to determine the effects of a low dose of conjugated equine estrogens (CE) in combination with different progestins (LD-HRT) and evaluate differences between regimens on sleep in symptomatic postmenopausal women. Postmenopausal women were recruited and assigned to calcium vitamin (control group) or to LD-HRT with 0.3mg of CE associated with a daily administration at bedtime of a progestin (2.5 mg MPA, CE + MPA, n = 20), or 100 mg natural micronized progesterone (CE + P, n = 20). Subjective symptoms were evaluated by the Greene climacteric scale, and by a visuanalogic graduated scale (0-10) at baseline and after 4, 8, and 12 weeks of study. Greene's scores for the control group were similar to those in LD-HRT group at baseline, and showed no significant modification at all subsequent measurements. Conversely, in LD-HRT group, a significant (P < 0.05) reduction in the scores of all Greene's domains was evident versus corresponding baseline and control group values. Conversely, in LD-HRT group, a significant (P < 0.05) reduction in the scores of all Greene's domains was evident with no difference in the scores of the two treated group. Both CE + MPA and CE + P significantly (P = 0.05) reduced the HF and sleep visuanalogic score in comparison to the control group. The score of sleep was significantly (P = 0.05) lower in the CE + P group in comparison to that measured in the CE + MPA group. No significant correlation between sleep and vasomotor score was found. In conclusion, low estrogen dose may have a value in the treatment of menopausal women in which sleep disturbances may be a symptom of estrogen deprivation. Low-dose estrogen associated with low-dose micronized progesterone may especially benefit women who complain of disturbed sleep. PMID- 15653006 TI - Low-dose, vaginally administered estrogens may enhance local benefits of systemic therapy in the treatment of urogenital atrophy in postmenopausal women on hormone therapy. AB - BACKGROUND: When genital atrophy exists, systemic hormone therapy (HT) has a timing until to induce vaginal proliferation and symptomatic relieve. Thus, in order to obtain a prompt improvement, the association of local therapy acting on the genital epithelium to the systemic treatment should be considered. OBJECTIVE: To evaluate the effects of a combined therapy consisting of vaginal estriol with transdermal 17-beta-estradiol (50 microg/day) plus medroxyprogesterone acetate (5 mg/day) per os in shortening the period of uro-genital symptoms. SUBJECTS AND METHODS: In a randomized, double blind, controlled with placebo study, 27 women with climacteric symptoms and atrophic vaginitis were treated for 4 months with HT plus vaginal estriol 0.5 mg/day (group E) or placebo (group P). Patients use the local medication daily for the first 3 weeks and twice-weekly thereafter. Before entering in the study, patients were asked about HT and selected for inclusion. In the first visit, electible patients after written informed consent were randomized to receive HT plus local estriol or placebo. All the subjects had baseline studies, including medical history, physical examination, blood and urine analysis. In order to evaluate the effect of local treatment on urinary and genital symptoms, a score for genital, urinary and colposcopic complaints (0 minimum-100 maximum) was developed. This score and Blatt-Kuperman were recorded and performed in every control. RESULTS: There were no differences on climacteric symptoms relief between the two groups. Additionally, the improvement in urinary symptoms at the end of the study was similar for both groups (from 16.5 +/- 6.1 to 8.5 +/- 2.4 for E group and from 15.8 +/- 7.8 to 8.8 +/- 2.7 for P group; P < 0.01 versus basal); however, those women in group E reached significant improvement on urinary complaints since the first month of treatment. Additionally, a significant difference between E and P was observed at months 2 and 3, although no differences were detected at the end of the study. Papanicolaou smear showed reactive or reparative changes and karyopyknotic index exhibited a significant increase in superficial cells in both groups and at the end of the study. CONCLUSIONS: Adding vaginal estriol to HRT may shorten the latency period for urinary symptoms. PMID- 15653007 TI - Influence of different HRT regimens on mammographic density. AB - OBJECTIVES: A prospective, randomized, open-label study was conducted to evaluate effects on mammographic density in postmenopausal and late perimenopausal women receiving continuous combined or sequential combined hormone replacement therapy (HRT). METHODS: The subjects were randomized to treatment with low-dose continuous combined HRT containing 1 mg 17beta-estradiol plus 0.5 mg norethisterone acetate (Activelle) or a sequential combined HRT regimen consisting of 0.625 mg conjugated equine estrogens for 28 days plus 5 mg medrogestone for 14 days (Presomen). Mammograms were obtained at baseline and after 9 cycles (each 28 days) of treatment. RESULTS: The majority of women (approximately two-thirds in each treatment group) had no changes in mammographic breast density between baseline and the final study visit. There were no marked differences between treatment groups. Approximately 20% of women in both groups had a slight increase in mammographic density. Only 10-14% of women in both groups had a pronounced increase in mammographic density. The analyses of the degree of change showed no remarkable differences between treatments. CONCLUSION: These results indicate that the increase in mammographic density with a low-dose continuous combined HRT regimen is no greater than that with a sequential combined HRT regimen. The type of progestogen does not have an impact on the extent of mammographic density changes. PMID- 15653008 TI - Transvaginal Doppler sonography: is there a role for this modality in the evaluation of women with postmenopausal bleeding? AB - OBJECTIVE: The aim of this study was to determine whether transvaginal sonography (TVS) and transvaginal Doppler sonography (TDS) can discriminate between normal and abnormal endometrium. MATERIALS AND METHODS: Patients who had vaginal bleeding an year after menopause and were not on HRT or tamoxifen were preoperatively examined by TVS and TDS on the same day of curettage. The endometrial thickness as well as the pulsatility index (PI) and resistance index (RI) of uterine arteries were recorded. RESULTS: Final pathology analysis revealed that 55/81 (67.9%) had normal endometrial tissue and 26/81 (32.1%) had an abnormality (endometrial hyperplasia, polyp or cancer). The mean endometrial thickness was greater in the abnormal group (9.4 mm versus 3.8 mm, P < 0.05). The mean PI of normal and abnormal endometrium were 2.85 and 1.53. The mean RI of normal and abnormal endometrium were 1.04 and 0.68. Univariate analysis found that PI < 2 and RI < 0.9 were correlated with abnormal endometrium (P < 0.05). Multivariate analysis revealed significance only for the endometrial thickness. CONCLUSIONS: TDS cannot distinguish between normal and abnormal endometrium. Using TVS only would result in a significant reduction of endometrial biopsy or curettage. PMID- 15653009 TI - Role of hysteroscopy with endometrial biopsy to rule out endometrial cancer in postmenopausal women with abnormal uterine bleeding. AB - OBJECTIVE: To compare the diagnostic accuracy of ultrasonographic endometrial thickness and outpatient hysteroscopy, to establish the most appropriate exam for the diagnosis of endometrial cancer in postmenopausal women with abnormal uterine bleeding (AUB). The secondary aim was to develop a multivariable approach considering clinical history as an added value for these diagnostic procedures. METHODS: This prospective study was conducted on 220 consecutive postmenopausal patients with AUB, who underwent ultrasonographic evaluation of endometrial thickness, outpatient hysteroscopy and endometrial biopsy. Evaluation of sensitivity, specificity, positive and negative predictive value was performed. Receiver operator characteristic curve (ROC) was calculated to assess the global performance of ultrasonographic measurement of endometrial thickness and diagnostic hysteroscopy as tests for detecting endometrial cancer and atrophy. RESULTS: Histological findings for <4 mm level revealed that atrophy was present in 48 (65%) and in 2 cases (2.7%) endometrial cancer was found; for > or = 4 mm values polyps and myomas were present in 86 (59%) and there were 11 (7.5%) endometrial cancer. Sensibility and specificity for trans-vaginal ultrasound, with a cut-off value > or = 4 mm, was 55.6% and 49.7% while positive predictive value was 83.3% and negative predictive value 98.1% (ROC curve 0.597). Hysteroscopy revealed sensitivity 100%, specificity 49.6%, positive predictive value 81.3% and negative predictive value 100% (ROC curve 0.993). CONCLUSIONS: In conclusion, endometrial thickness <4 mm can miss malignancies but trans-vaginal ultrasound remains the first line diagnostic procedure in postmenopausal women without AUB, because it is not invasive and has high sensitivity for detecting endometrial cancer and other endometrial disease; according to our experience, outpatient hysteroscopy with biopsy is mandatory in all postmenopausal women with AUB. PMID- 15653010 TI - Effect of oral testosterone undecanoate on visuospatial cognition, mood and quality of life in elderly men with low-normal gonadal status. AB - OBJECTIVES: The effects of supplemental testosterone on cognition, mood and wellbeing in ageing men are unclear. This study aimed to assess the effect of 12 months of oral testosterone supplementation on cognitive function, mood and quality of life in elderly men with low-normal gonadal status, not specifically selected for cognitive or mood defects. METHODS: A standard oral dose (80 mg twice daily) of testosterone undecanoate (TU) or placebo was administered for one year to 76 healthy men 60 years or older. All men had a free testosterone index (FTI) of 0.3-0.5, which represents a value below the normal lower limit for young men (19-30 years), but remains within the overall normal male range. A neuropsychological assessment including the trail making test (part B), visuospatial (VSP) block design test, mini mental state exam (MMSE), Geriatric Depression Scale (GDS), a 5-point Likert and a 10-point visual analogue quality of life (QoL) scale, along with serum hormone measurements were obtained at baseline, 6, and 12 months. RESULTS: Although calculated bioavailable testosterone (cBT) and FTI were higher, and muscle mass increased after 12 months, there was no difference in scores on the trail making or VSP block tests, the MMSE, GDS or either of the QoL scales between the testosterone and placebo group. There was no relationship between baseline cBT or FTI and treatment effect for any of the outcome measures. CONCLUSIONS: 12-month supplementation with oral TU does not affect scores on visuospatial tests or mood and quality of life scales in older men with low-normal gonadal status. PMID- 15653011 TI - Psychometric properties of the Persian version of the osteoporosis knowledge and health belief questionnaires. PMID- 15653012 TI - Oro-vaginal-vulvar lichen planus: report of two new cases. AB - Atrophic-erosive lichen planus with oral and genital involvement is a rare condition and is often difficult to diagnose. Patients seldom report genital symptoms to the dentist and dentists do not generally investigate about genital lesions. Delays in diagnosis may cause complications and affect the quality of life. We report the clinical and histopathological features of two new cases of oro-vaginal-vulvar lichen planus and review current treatment options for this condition. Genital lichen planus should be suspected in case of atrophic-erosive oral lichen planus. A thorough multidisciplinary medical management and active early treatment are necessary to improve symptoms and prevent genital sequelae. This might also be a relevant prevention strategy for the risk of squamous cell carcinoma, although data to fully support this statement still need investigation. PMID- 15653014 TI - Efficacy and safety of high-density lipoprotein cholesterol-increasing compounds: a meta-analysis of randomized controlled trials. AB - OBJECTIVES: The aim of this research was to estimate the efficacy and safety of current high-density lipoprotein cholesterol (HDL-C)-increasing drugs. BACKGROUND: Epidemiologic evidence has shown that HDL-C is inversely related to coronary heart disease (CHD) risk. However, the evidence for reducing CHD risk by raising HDL-C is thin, predominantly due to the paucity of effective and safe HDL increasing drugs. METHODS: Randomized controlled trials with fibrates and niacin, published between 1966 through February 2004 (MEDLINE), were retrieved. Information on treatment, baseline characteristics, serum lipids, end points, and side-effects were independently abstracted by two authors using a standardized protocol. RESULTS: Data from 53 trials (16,802 subjects) using fibrates and 30 trials (4,749 subjects) using niacin were included. Random-effects model showed 11% versus 10% reduction in total cholesterol, 36% versus 20% reduction in triglycerides, 8% versus 14% reduction in low-density lipoprotein cholesterol, and 10% versus 16% increase in HDL-C for fibrates and niacin, respectively. Apart from flushes in the niacin group, both fibrates and niacin were shown to be well tolerated and safe. Fibrates reduced the risk for major coronary events by 25% (95% confidence interval 10% to 38%), whereas current available data for niacin indicate a 27% reduction. CONCLUSIONS: Fibrates reduce major coronary events and increase HDL-C levels without significant toxicity. Niacin has a more potent effect on HDL-C levels, whereas data on cardiovascular event rate reduction are limited. Future studies need to evaluate whether additional HDL increase by fibrates or particularly newer niacin formulations on top of statin therapy translates into further event reduction in high-risk subjects, without significant toxicity. PMID- 15653013 TI - Cardiac medical therapy in patients after undergoing coronary artery bypass graft surgery: a review of randomized controlled trials. AB - The purpose of this paper is to review the randomized controlled trial (RCT) data investigating cardiac medical therapy for patients after coronary artery bypass grafting (CABG). We identified RCTs with > or =100 enrolled patients that examined the impact of cardiac medical therapy on outcomes > or =1 year after CABG. The MEDLINE database was searched for trials conducted between 1966 and 2004 on the following medications: aspirin, antilipid agents, beta-blockers, calcium channel blockers (CCBs), nitrates, and angiotensin-converting enzyme (ACE) inhibitors. Both aspirin and antilipid agents were found to reduce the progression of atherosclerosis and the occurrence of graft occlusion. Cardiovascular events were decreased with antilipid agents. In small trials, beta blockers and CCBs failed to decrease the incidence of cardiovascular events. No RCTs examined nitrates, and one small RCT documented a reduction in cardiovascular events among patients treated with ACE inhibitors. We conclude that few RCTs have examined the efficacy of cardiac medical therapy in post-CABG patients. Based on current RCT evidence, aspirin and antilipid agents should be used routinely after CABG. However, current data do not support the use of beta blockers, CCBs, and nitrates, and more evidence is needed regarding the use of ACE inhibitors. PMID- 15653015 TI - Predictive factors for ischemic target vessel revascularization in the Prevention of Restenosis with Tranilast and its Outcomes (PRESTO) trial. AB - OBJECTIVES: The aim of the present study was to determine the rates of target vessel revascularization (TVR) and to determine predictors of TVR from clinical and angiographic variables available in the Prevention of Restenosis With Tranilast and its Outcomes (PRESTO) database. BACKGROUND: The rates of TVR after percutaneous revascularization procedures, and its prediction with available clinical and angiographic variables, is less well known. METHODS: We studied nine month TVR in 11,484 patients enrolled in the PRESTO trial. Clinical, lesion related, and procedural characteristics were analyzed in a logistic regression model. Study data were divided at random into an 80% training set on which the models were developed and a 20% hold-out set on which the model properties were evaluated. RESULTS: A total of 14% (n = 1,609) had ischemic TVR. Clinical variables with increased risk for TVR included younger age; hypertension; diabetes mellitus; nonsmokers; unstable angina; previous coronary artery bypass grafting; peripheral vascular disease; procedure- and lesion-related such as ostial location, multilesion angioplasty, location in the left anterior descending artery, length > or =20 mm, in-stent restenosis at baseline, and use of rotablator. There was significant increase in the risk of ischemic TVR at U.S. treatment sites. Smoking and stent placement were associated with lower risk of ischemic TVR. The mean area (+/- SD) under the receiver-operating characteristic curve of the bootstrap samples was 0.66, indicating a modest ability of the model to discriminate patients who needed TVR on follow-up. CONCLUSIONS: Despite being the largest prospective trial designed to test restenosis, the discriminatory ability of the clinical and angiographic variables to predict TVR is modest. PMID- 15653016 TI - Intravascular ultrasound-guided treatment for angiographically indeterminate left main coronary artery disease: a long-term follow-up study. AB - OBJECTIVES: The purpose of this study was to evaluate the efficacy of an intravascular ultrasound (IVUS)-guided strategy for patients with angiographically indeterminate left main coronary artery (LMCA) disease. BACKGROUND: The assessment of LMCA lesions using coronary angiography is often challenging; IVUS provides useful information for assessment of coronary disease. METHODS: Intravascular ultrasound was performed on 121 patients with angiographically normal LMCAs to determine the lower range of normal minimum lumen area (MLA), defined as the mean - 2 SD. We conducted IVUS studies on 214 patients with angiographically indeterminate LMCA lesions, and deferral of revascularization was recommended when the MLA was larger than this predetermined value. RESULTS: The lower range of normal LMCA MLA was 7.5 mm(2). Of the patients with angiographically indeterminate LMCAs, 83 (38.8%) had an MLA <7.5 mm(2), and 131 (61.2%) an MLA > or =7.5 mm(2). Left main coronary artery revascularization was performed in 85.5% (71 of 83) of patients with an MLA <7.5 mm(2) and deferred in 86.9% (114 of 131) of patients with an MLA > or =7.5 mm(2). Long-term follow up (mean 3.3 +/- 2.0 years) showed no significant difference in major adverse cardiac events (target vessel revascularization, acute myocardial infarction, and death) between patients with an MLA <7.5 mm(2) who underwent revascularization and those with an MLA > or =7.5 mm(2) deferred for revascularization (p = 0.28). Based on outcome, the best cut-off MLA by receiver operating characteristic was 9.6 mm(2). Multivariate predictors of cardiac events were age, smoking, and number of non-LMCA vessels diseased. CONCLUSIONS: Intravascular ultrasound is an accurate method to assess angiographically indeterminate lesions of the LMCA. Furthermore, deferring revascularization for patients with a minimum lumen area > or =7.5 mm(2) appears to be safe. PMID- 15653017 TI - Detection of embolic particles with the Doppler guide wire during coronary intervention in patients with acute myocardial infarction: efficacy of distal protection device. AB - OBJECTIVES: We investigated whether embolic particles could be detected as high intensity transient signals (HITS) with a Doppler guide wire during percutaneous coronary intervention (PCI) in patients with acute myocardial infarction (AMI) We also assessed whether these signals could be reduced using a distal protection (DP) device. BACKGROUND: Embolization of thrombi and plaque components to the microcirculation is a major complication of PCI in patients with AMI. Embolic particles running in the cerebral artery are detected as HITS by transcranial Doppler ultrasound. METHODS: We prospectively studied 16 consecutive patients with AMI who underwent direct PCI within 24 h after the onset of symptoms. A PercuSurge GuardWire (MedtronicAVE, Santa Rosa, California) was used as the DP device. Eight patients were randomly assigned to the non-DP group, and the remaining eight were assigned to the DP group. Coronary flow velocity was recorded continuously from before the first balloon inflation to after balloon deflation. RESULTS: All patients in the non-DP group had HITS detected (12 +/- 9 counts) within five consecutive beats (4 +/- 1 beat) after balloon deflation, but none were detected in any of the patients in the DP group. CONCLUSIONS: The Doppler guide wire can be used to visually detect and count emboli as HITS, and the DP device is effective for prevention of distal embolization. PMID- 15653018 TI - Pressure-derived measurement of coronary flow reserve. AB - OBJECTIVES: We aimed to validate the technique of measuring the coronary flow reserve (CFR) with coronary pressure measurements against an established thermodilution technique. BACKGROUND: The CFR has traditionally required measurement of coronary blood flow velocity with the Doppler wire and, more recently, using a thermodilution technique with the coronary pressure wire. However, recent work has suggested that the CFR may be derived from pressure measurements alone (the ratio of the square root of the pressure drop across an epicardial stenosis during hyperemia to that value at rest). This depends on the assumption that friction losses across a coronary stenosis are negligible. METHODS: We compared pressure-derived CFR values with those obtained by the thermodilution technique using the intracoronary pressure wire in 38 stenoses in 34 patients with significant coronary stenoses undergoing percutaneous intervention. We also compared these two techniques of measuring CFR in 25 stenoses (6 vessels) artificially created by inflating small balloons within a stented coronary artery after percutaneous intervention. RESULTS: There is a close linear relationship between pressure-derived and thermodilution CFR in native (r(2) = 0.52; p < 0.001) and artificial stenoses (r(2) = 0.54; p < 0.05), although the pressure-derived technique appears to systematically underestimate CFR values in both situations. This applies to native and artificial stenoses. CONCLUSIONS: Coronary flow reserve cannot be measured merely with pressure alone, and it cannot be safely assumed that friction losses are negligible across a native coronary stenosis. These data suggest that friction loss is an important determinant of the pressure gradient along an atherosclerotic coronary artery. PMID- 15653019 TI - Health-related quality of life after interventional or conservative strategy in patients with unstable angina or non-ST-segment elevation myocardial infarction: one-year results of the third Randomized Intervention Trial of unstable Angina (RITA-3). AB - OBJECTIVES: We sought to compare the effects of an early interventional strategy (IS) versus a conservative strategy (CS) on health-related quality of life (HRQOL) in patients with non-ST-segment elevation acute coronary syndromes (ACS). BACKGROUND: The third Randomized Intervention Trial of unstable Angina (RITA-3) evaluated early IS (n = 895) versus CS (n = 915). We report one-year results of the RITA-3 trial concerning HRQOL. METHODS: The patients' HRQOL was assessed with the Short Form-36 (SF-36) and Seattle Angina Questionnaire (SAQ) at four-month and one-year follow-up, and the EuroQOL Visual Analogue Scale (EQ-VAS) and EuroQOL 5-Dimensional Classification (EQ-5D) also measured at baseline. Analysis was performed using the two-sample t test and analysis of co-variance. RESULTS: Mean changes from baseline EQ-VAS scores were better for IS than for CS at four months (treatment difference of 3.0, p < 0.001) and one year (2.3, p < 0.01). The EQ-5D utility scores were also higher for IS at four months (treatment difference: 0.036, p < 0.01) and at one year (0.016, p = 0.20). For SF-36, IS scored significantly better at four months for physical function, physical role function, emotional role function, social function, vitality, and general health. The SAQ scores for exertional capacity, anginal stability and frequency, treatment satisfaction, and disease perception were better for IS at four months. These treatment differences were present but attenuated by one-year follow-up. Improvements in HRQOL for IS could be attributed to improvements in anginal symptoms. CONCLUSIONS: In patients with non-ST-segment elevation ACS, an early IS provides greater gains in HRQOL, as compared with CS, mainly due to improvements in angina grade. PMID- 15653020 TI - Pregnancy-associated plasma protein-A levels in patients with acute coronary syndromes: comparison with markers of systemic inflammation, platelet activation, and myocardial necrosis. AB - OBJECTIVES: The goal of this study was to determine the predictive value of pregnancy-associated plasma protein-A (PAPP-A) in patients with acute coronary syndromes (ACS). BACKGROUND: Pregnancy-associated plasma protein-A is a zinc binding matrix metalloproteinase abundantly expressed in eroded and ruptured plaques and may serve as a marker of plaque destabilization. METHODS: In 547 patients with angiographically validated ACS and in a heterogeneous emergency room population of 644 patients with acute chest pain, respectively, PAPP-A as well as markers of myocardial necrosis (troponin T [TnT]), ischemia (vascular endothelial growth factor [VEGF]), inflammation (high-sensitivity C-reactive protein [hsCRP]), anti-inflammatory activity (interleukin [IL]-10), and platelet activation (soluble CD40 ligand [sCD40L]) were determined. Patients were followed for the occurrence of death or myocardial infarction. RESULTS: In patients with ACS, elevated PAPP-A levels (>12.6 mIU/l) indicated an increased risk (odds ratio 2.44 [95% confidence interval (CI) 1.43 to 4.15]; p = 0.001). When the analysis was restricted to TnT-negative patients, PAPP-A still identified a subgroup of high-risk patients (odds ratio [OR] 2.72 [95% confidence interval (CI) 1.25 to 5.89]; p = 0.009). In a multivariable model, PAPP-A (OR 2.01; p = 0.015), sCD40L (OR 2.37; p = 0.003), IL-10 (OR 0.43; p = 0.003), and VEGF (OR 2.19; p = 0.018) were independent predictors. Prospective validation in patients with chest pain confirmed that PAPP-A levels reliably identify high-risk patients (adjusted OR 2.32 [95% CI 1.32 to 4.26]; p = 0.008). Patients negative for all three markers (TnT, sCD40L, and PAPP-A) were at very low cardiac risk (30 days: 3.0% event rate; no death). CONCLUSIONS: The PAPP-A level as a marker of plaque instability is a strong independent predictor of cardiovascular events in patients with ACS. Simultaneous determination of biomarkers with distinct pathophysiological profiles appears to remarkably improve risk stratification in patients with ACS. PMID- 15653021 TI - Persistent systemic inflammation in unstable angina is largely unrelated to the atherothrombotic burden. AB - OBJECTIVES: The aim of this study was to assess the relationship between systemic inflammation, atherosclerosis, and thrombosis in two distinct clinical models of atherothrombosis. BACKGROUND: Persistent unstable angina (UA) is commonly associated with coronary thrombosis and persistent systemic inflammation. METHODS: We assessed circulating markers of activation of the thrombotic and fibrinolytic cascades and systemic soluble and cellular markers of inflammation on admission in 40 patients with persisting UA (Braunwald class IIIB; group 1) and 30 patients with Leriche-Fontaine stage IIB-III peripheral artery disease awaiting revascularization (group 2). RESULTS: The extent of atherosclerosis (p < 0.01) and activation of the coagulation system were greater in group 2, which had higher thrombin-antithrombin III complexes and D-dimer levels (2.7 and 24.4 microg/l, respectively), than in group 1 (2.0 microg/l and 12.9 microg/l, p = 0.02 and p = 0.0001, respectively). In contrast, C-reactive protein and interleukin-6 levels were higher in group 1 (7.6 pg/ml and 7.8 pg/ml, respectively) than in group 2 (4.5 pg/ml and 3.0 pg/ml, p < 0.01 and p = 0.03, respectively). Moreover, neutrophil activation was only found in group 1 (neutrophil myeloperoxidase content -4.0 arbitrary units vs. +3.4 arbitrary units in group 2, p < 0.0001). These differences persisted during the initial three days of hospitalization. CONCLUSIONS: Such a large, consistent discrepancy between atherothrombotic burden and systemic inflammation suggests that atherothrombosis, by itself, is an unlikely cause of persisting, recurring UA. An understanding of the primary inflammatory mechanisms of persistent and recurrent coronary instability could open the way to novel therapeutic strategies. PMID- 15653022 TI - Another look at the age-old question: which came first, the elevated c-reactive protein or the atherothrombosis? PMID- 15653023 TI - Variability in platelet responsiveness to clopidogrel among 544 individuals. AB - OBJECTIVES: We sought to describe the responses of patients to clopidogrel using ex vivo measures of platelet aggregation and activation in a large, heterogeneous population. BACKGROUND: Recently, a number of reports, using various definitions, have dichotomized patients who are treated with clopidogrel into a minority of "non-responders" and a majority of "responders." Such classifications imply that treatment leads to an all-or-none response, with potentially important clinical implications. METHODS: We conducted secondary post-hoc analyses of a dataset consisting of volunteers (n = 94) and patients after coronary stenting (n = 405), with heart failure (n = 25), and after stroke (n = 20). RESULTS: The response of subjects to clopidogrel followed a normal, bell-shaped distribution, with a mean and standard deviation of 41.9 +/- 20.8% when aggregation was induced by 5 mumol/l of adenosine diphosphate. When hyporesponsiveness and hyper responsiveness to clopidogrel were considered to be two standard deviations less than and greater than the mean, respectively, the prevalence of hyporesponsiveness and hyper-responsiveness in these patients was 4.2% and 4.8%, respectively. Pretreatment platelet activity and clinical characteristics were not associated with responsiveness to clopidogrel. CONCLUSIONS: Individuals receiving clopidogrel exhibit a wide variability in response that follows a normal distribution. The clinical implications of this variability are unknown but potentially are important. Clinical trials are needed to define whether hyporesponders to clopidogrel are at increased risk for thrombotic events and whether hyper-responders are at increased risk for bleeding. If so, the individualization of antiplatelet therapy, including clopidogrel dosing, may be possible in the future but will require the ability to easily and reproducibly measure responsiveness by a method that has been proven to be predictive of clinical events. PMID- 15653024 TI - What resting heart rate should one aim for when treating patients with heart failure with a beta-blocker? Experiences from the Metoprolol Controlled Release/Extended Release Randomized Intervention Trial in Chronic Heart Failure (MERIT-HF). AB - OBJECTIVES: The goal of this study was to explore the question: what resting heart rate (HR) should one aim for when treating patients with heart failure with a beta-blocker? BACKGROUND: The interaction of pretreatment and achieved resting HR with the risk-reducing effect of beta-blocker treatment needs further evaluation. METHODS: Cardiovascular risk and risk reduction were analyzed in five subgroups defined by quintiles (Q) of pretreatment resting HR in the Metoprolol Controlled Release/Extended Release Randomized Intervention Trial in Chronic Heart Failure (MERIT-HF). RESULTS: Mean baseline HR in the 5 Qs were 71, 76, 81, 87, and 98 beats/min; achieved HR 63, 66, 68, 72, and 75 beats/min; and net change -8, -10, -11, -13, and -14 beats/min, respectively. Baseline HR was related to a number of baseline characteristics. Cardiovascular risk was no different in Q1 to Q4 (placebo groups) but increased in Q5 (HR above 90 beats/min). No relationship was observed between the risk-reducing effect of metoprolol controlled release/extended release (CR/XL) and baseline HR in the five Qs of baseline HR, or achieved HR, or change in HR during follow-up, respectively. CONCLUSIONS: Metoprolol CR/XL significantly reduced mortality and hospitalizations independent of resting baseline HR, achieved HR, and change in HR. Achieved HR and change in HR during follow-up were closely related to baseline HR; therefore, it was not possible to answer the question posed. Instead, one has to apply a very simple rule: aim for the target beta-blocker dose used in clinical trials, and strive for the highest tolerated dose in all patients with heart failure, regardless of baseline and achieved HR. PMID- 15653025 TI - Contribution of ischemic mitral regurgitation to congestive heart failure after myocardial infarction. AB - OBJECTIVES: The purpose of this study was to define the contribution of ischemic mitral regurgitation (IMR) to the occurrence of congestive heart failure (CHF) after myocardial infarction (MI). BACKGROUND: After MI, CHF is a frequent and serious complication, but its determinants and, particularly, the role of IMR are poorly defined. METHODS: We analyzed 173 asymptomatic patients with previous Q wave MI (>16 days) with echocardiographic quantitation of IMR (measuring effective regurgitant orifice [ERO] and regurgitant volume). The 102 patients with IMR were matched to 71 patients without IMR for age (71 +/- 11 years vs. 68 +/- 9 years; p = 0.11), gender (76% vs. 82% males; p = 0.41), and left ventricular ejection fraction (EF) (37 +/- 14% vs. 36 +/- 11%; p = 0.92). RESULTS: Five-year rates of CHF and of CHF or cardiac death (CD) were 36 +/- 5% and 52 +/- 5%, respectively. Independent determinants of CHF were EF, sodium plasma level, and presence and degree of IMR (p < 0.0001). Five-year CHF rates were 18 +/- 5% without mitral regurgitation (MR), 53 +/- 7% with IMR, 46 +/- 9% with ERO 1 to 19 mm(2) and 68 +/- 12% with ERO > or =20 mm(2) (all p < 0.0001). The adjusted relative risk of CHF was 3.65 (95% confidence interval [CI] 1.86 to 7.75) for IMR presence and 4.42 (95% CI 1.9 to 10.5) for ERO > or =20 mm(2). The adjusted relative risk of CHF/CD was 2.97 (95% CI 1.77 to 5.16) for IMR presence and 4.4 (95% CI 2.4 to 8.2) for ERO > or =20 mm(2). CONCLUSIONS: After MI, incidence of CHF and of CHF/CD are high even in patients with no or minimal symptoms at baseline and are higher in patients with IMR. Congestive heart failure is independently determined by larger ERO of IMR. These data suggest that detecting and quantifying IMR is essential for risk stratification after MI. Value of IMR treatment in improving post-MI outcome should be investigated. PMID- 15653026 TI - Cystatin-C and mortality in elderly persons with heart failure. AB - OBJECTIVES: We sought to evaluate cystatin-C, a novel measure of renal function, as a predictor of mortality in elderly persons with heart failure (HF) and to compare it with creatinine. BACKGROUND: Renal function is an important prognostic factor in patients with HF, but creatinine levels, which partly reflect muscle mass, may be insensitive for detecting renal insufficiency. METHODS: A total of 279 Cardiovascular Health Study participants with prevalent HF and measures of serum cystatin-C and creatinine were followed for mortality outcomes over a median of 6.5 years. RESULTS: Median creatinine and cystatin-C levels were 1.05 mg/dl and 1.26 mg/l. Each standard deviation increase in cystatin-C (0.35 mg/l) was associated with a 31% greater adjusted mortality risk (95% confidence interval [CI] 20% to 43%, p < 0.001), whereas each standard deviation increase in creatinine (0.39 mg/dl) was associated with a 17% greater adjusted mortality risk (95% CI 1% to 36%, p = 0.04). When both measures were combined in a single adjusted model, cystatin-C remained associated with elevated mortality risk (hazard ratio 1.60, 95% CI 1.32 to 1.94), whereas creatinine levels appeared associated with lower risk (hazard ratio 0.73, 95% CI 0.57 to 0.95). CONCLUSIONS: Cystatin-C is a stronger predictor of mortality than creatinine in elderly persons with HF. If confirmed in future studies, this new marker of renal function could improve risk stratification in patients with HF. PMID- 15653027 TI - Independent and incremental prognostic value of early mitral annulus velocity in patients with impaired left ventricular systolic function. AB - OBJECTIVES: This study sought to investigate the incremental prognostic value of non-invasive measures of early myocardial relaxation and left ventricular diastolic pressure (LVDP) in patients with impaired left ventricular (LV) systolic function. BACKGROUND: The early diastolic mitral annulus velocity (Em) reflects myocardial relaxation, and the combined ratio of the early transmitral flow velocity (E) to Em (E/Em) >15 correlates well with elevated mean LVDP. It is unknown if these new indexes will predict poorer survival in patients with LV systolic dysfunction. METHODS: Echocardiograms were prospectively obtained in 182 patients with impaired LV systolic function, defined as an LV ejection fraction <0.50. The end point was cardiac mortality. The majority of this patient sample (80%) has been reported on in a previous publication. RESULTS: After a median 48 months' follow-up, Em emerged as an independent predictor of survival (hazard ratio 0.61, 95% confidence interval 0.45 to 0.82). An Em <3 cm/s was associated with a significantly excess mortality (log-rank statistic 9.36, p = 0.002), and this measurement added incremental prognostic value to standard indexes of systolic or diastolic function, including a deceleration time <140 ms and an E/Em >15 (p = 0.038). CONCLUSIONS: Early diastolic mitral annulus velocity is a powerful predictor of cardiac mortality in patients with LV systolic impairment; Em <3 cm/s emerged as the best prognosticator in long-term follow-up, incremental to other clinical or echocardiographic variables, including the ratio E/Em. PMID- 15653028 TI - Heart rate turbulence after atrial premature beats before spontaneous onset of atrial fibrillation. AB - OBJECTIVES: This study was designed to assess the temporal changes in vagal responses to atrial premature beats before spontaneous onset of atrial fibrillation (AF). BACKGROUND: Enhanced vagal activity plays a major role in the onset and perpetuation of experimental AF, but the role of vagal activation in the onset of clinical AF episodes is not so well established. METHODS: We calculated heart rate turbulence after atrial premature impulses occurring 0 to 60 min before the onset of AF ("prior to AF") and compared it with the hourly means of the other hours of the 24-h electrocardiogram recordings ("non-AF hours") in 39 patients with structural heart disease and 29 patients with lone AF. Traditional heart rate variability measurements and approximate entropy (ApEn) were also analyzed. RESULTS: Turbulence onset (TO) was significantly less negative during the 1 h preceding AF than during the non-AF hours (0.71 +/- 1.76 vs. -0.35 +/- 1.46, p < 0.00001). Less negative TO before AF was observed among both the patients with structural heart disease (1.16 +/- 1.73 vs. 0.07 +/- 1.23; p < 0.0001) and those with lone AF (0.17 +/- 1.67 vs. -0.85 +/- 1.56; p < 0.0001). No significant difference was seen in the turbulence slope between the two periods, and none of the traditional frequency and time domain measurements differentiated between the periods; ApEn was significantly lower before AF than during the non-AF hours (p < 0.01). CONCLUSIONS: Altered heart rate dynamics, suggesting transient enhancement of vagal outflow after premature atrial excitation, are temporally related to spontaneous onset of clinical AF. PMID- 15653029 TI - Pre-existent left atrial scarring in patients undergoing pulmonary vein antrum isolation: an independent predictor of procedural failure. AB - OBJECTIVES: The goal of this study was to assess the impact of left atrial scarring (LAS) on the outcome of patients undergoing pulmonary vein antrum isolation (PVAI) for atrial fibrillation (AF). BACKGROUND: Left atrial scarring may be responsible for both the perpetuation and genesis of AF. METHODS: A total of 700 consecutive patients undergoing first-time PVAI were studied. Before ablation, extensive voltage mapping of the left atrium (LA) was performed using a multipolar Lasso catheter guided by intracardiac echocardiography (ICE). Patients with LAS were defined by a complete absence of electrographic recording by a circular mapping catheter in multiple LA locations, and this was validated by electroanatomic mapping. All four pulmonary vein antra and the superior vena cava were isolated using an ICE-guided technique. Patients were followed at least nine months for late AF recurrence. Univariate and multivariate analyses were performed to assess the predictive value of LAS and other variables on outcome. RESULTS: Of 700 patients, 42 had LAS, which represented 21 +/- 11% of the LA surface area by electroanatomic mapping. Patients with LAS had a significantly higher AF recurrence (57%) compared with non-LAS patients (19%, p = 0.003). Also, LAS was associated with a significantly larger LA size, lower ejection fraction, and higher C-reactive protein levels. Univariate analysis revealed age, nonparoxysmal AF, and LAS as predictors of recurrence. Multivariate analysis showed LAS as the only independent predictor of recurrence (hazard ratio 3.4, 95% confidence interval 1.3 to 9.4; p = 0.01). CONCLUSIONS: Pre-existent LAS in patients undergoing PVAI for AF is a powerful, independent predictor of procedural failure. Left atrial scarring is associated with a lower EF, larger LA size, and increased inflammatory markers. PMID- 15653030 TI - Antagonism of selectin function attenuates microvascular platelet deposition and platelet-mediated myocardial injury after transient ischemia. AB - OBJECTIVES: The goal of this study was to assess whether selectin blockade reduces myocardial platelet deposition and platelet-mediated injury after transient ischemia. BACKGROUND: Selectins participate in platelet adhesion to reperfused endothelium. METHODS: Thiopental-anesthetized, open-chest pigs were subjected to mechanical injury of the left anterior descending coronary artery followed by a 48-min occlusion and 2 (n = 20) or 4 (n = 16) h of reperfusion. Fifteen minutes before occlusion, animals were blindly allocated to receive a continuous intravenous infusion of the selectin blocker fucoidan (30 microg/kg/min, plus a 1-mg/kg bolus in the latter group) or saline. In isolated rat hearts infused with thrombin-activated platelets, the effects of fucoidan (30 microg/ml) administered during reperfusion after 40 min of global ischemia were also analyzed. RESULTS: Fucoidan did not prevent the development of cyclic reductions in coronary flow, but reduced the content of (99m)Tc-labeled platelets in reperfused myocardium after 2 h of reperfusion (23.4 +/- 3.3 vs. 42.1 +/- 8.3 x 10(6) platelets/g in treated and untreated animals, p = 0.03) and attenuated the impairment in the coronary flow reserve and reduced infarct size after 4 h (53 +/- 2% vs. 73 +/- 5% of the ischemic region, respectively, p = 0.003). Treated animals showed a trend toward less neutrophil infiltration early after reperfusion, but not after 4 h. In isolated hearts, fucoidan improved functional recovery and reduced coronary resistance and lactate dehydrogenase release, lacking any beneficial effects if given in the absence of platelets. CONCLUSIONS: The results suggest that selectin-dependent adhesion is a prominent mechanism of platelet deposition in reperfused cardiac microvessels and highlight its potential as a therapeutic target in patients with acute myocardial infarction. PMID- 15653031 TI - Cellular and ionic mechanism for drug-induced long QT syndrome and effectiveness of verapamil. AB - OBJECTIVES: We examined the cellular and ionic mechanism for QT prolongation and subsequent Torsade de Pointes (TdP) and the effect of verapamil under conditions mimicking KCNQ1 (I(Ks) gene) defect linked to acquired long QT syndrome (LQTS). BACKGROUND: Agents with an I(Kr)-blocking effect often induce marked QT prolongation in patients with acquired LQTS. Previous reports demonstrated a relationship between subclinical mutations in cardiac K+ channel genes and a risk of drug-induced TdP. METHODS: Transmembrane action potentials from epicardial (EPI), midmyocardial (M), and endocardial (ENDO) cells were simultaneously recorded, together with a transmural electrocardiogram, at a basic cycle length of 2,000 ms in arterially perfused feline left ventricular preparations. RESULTS: The I(Kr) block (E-4031: 1 micromol/l) under control conditions (n = 5) prolonged the QT interval but neither increased transmural dispersion of repolarization (TDR) nor induced arrhythmias. However, the I(Kr) blocker under conditions with I(Ks) suppression by chromanol 293B 10 micromol/l mimicking the KCNQ1 defect (n = 10) preferentially prolonged action potential duration (APD) in EPI rather than M or ENDO, thereby dramatically increasing the QT interval and TDR. Spontaneous or epinephrine-induced early afterdepolarizations (EADs) were observed in EPI, and subsequent TdP occurred only under both I(Ks) and I(Kr) suppression. Verapamil (0.1 to 5.0 micromol/l) dose-dependently abbreviated APD in EPI more than in M and ENDO, thereby significantly decreasing the QT interval, TDR, and suppressing EADs and TdP. CONCLUSIONS: Subclinical I(Ks) dysfunction could be a risk of drug induced TdP. Verapamil is effective in decreasing the QT interval and TDR and in suppressing EADs, thus preventing TdP in the model of acquired LQTS. PMID- 15653032 TI - A prospective randomized comparison between paclitaxel and sirolimus stents in the real world of interventional cardiology: the TAXi trial. AB - OBJECTIVES: We conducted this trial to assess whether a sirolimus-eluting stent (SES) produces similar results to a paclitaxel-eluting stent (PES) when used in the real world of interventional cardiology. BACKGROUND: Several drug-eluting stents have been shown to exert a beneficial effect on restenosis when used in the treatment of coronary artery disease. Any potential superiority of one drug over the others, however, is still unknown. METHODS: To evaluate whether a PES or an SES is superior in daily practice, we randomized all patients suitable to receive a drug-eluting stent in our institution. Clinical follow-up was obtained after at least six months. RESULTS: A total of 202 patients were included in this trial. One hundred patients received a PES and 102 received an SES. Procedural success was 99% in both groups. Incidence of major adverse cardiac events at follow-up (mean 7 +/- 2 months) was 4% with the PES and 6% with the SES (p = 0.8). The need for target lesion revascularization was very low in both groups (1% with the PES and 3% with the SES). CONCLUSIONS: Our results confirm that the high success rate obtained with both stents in randomized trials can be replicated in routine clinical practice. In this small group of patients we were unable to show any advantage of one stent over the other. PMID- 15653033 TI - Prevalence and clinical impact of stent fractures after femoropopliteal stenting. AB - OBJECTIVES: The aim of this study was to investigate the occurrence and the clinical impact of stent fractures after femoropopliteal stenting. BACKGROUND: The development of femoral stent fractures has recently been described; however, there are no data about the frequency and the clinical relevance. METHODS: A systematic X-ray screening for stent fractures was performed in 93 patients. In total, 121 legs treated by implantation of self-expanding nitinol stents were investigated after a mean follow-up time of 10.7 months. The mean length of the stented segment was 15.7 cm. RESULTS: Overall, stent fractures were detected in 45 of 121 treated legs (37.2%). In a stent-based analysis, 64 of 261 stents (24.5%) showed fractures, which were classified as minor (single strut fracture) in 31 cases (48.4%), moderate (fracture of >1 strut) in 17 cases (26.6%), and severe (complete separation of stent segments) in 16 cases (25.0%). Fracture rates were 13.2% for stented length < or =8 cm, 42.4% for stented length >8 to 16 cm, and 52.0% for stented length >16 cm. In 21 cases (32.8%) there was a restenosis of >50% diameter reduction at the site of stent fracture. In 22 cases (34.4%) with stent fracture there was a total stent reocclusion. According to Kaplan-Meier estimates, the primary patency rate at 12 months was significantly lower for patients with stent fractures (41.1% vs. 84.3%, p < 0.0001). CONCLUSIONS: There is a considerable risk of stent fractures after long segment femoral artery stenting, which is associated with a higher in-stent restenosis and reocclusion rate. PMID- 15653034 TI - President's page: The future is now: ACC goes to Washington. PMID- 15653035 TI - Early recovery of impaired coronary flow reserve by carvedilol therapy in patients with idiopathic dilated cardiomyopathy: a serial transthoracic Doppler echocardiographic study. PMID- 15653036 TI - Efficacy of a patient-activated pharmacologic pump using phenylephrine as active drug and prodromal symptoms as a marker of imminent loss of consciousness to abort tilt-induced syncope. PMID- 15653037 TI - Association of smoking with improved myocardial perfusion and the angiographic characterization of myocardial tissue perfusion after fibrinolytic therapy for ST segment elevation myocardial infarction. PMID- 15653038 TI - Coronary stenting versus balloon angioplasty in small vessels. PMID- 15653040 TI - Thrombolysis should be regarded as first-line therapy for prosthetic valve thrombosis in the absence of contraindications. PMID- 15653041 TI - Cardiologist in the carotids. PMID- 15653043 TI - Advantage of percutaneous coronary intervention over medical therapy in angina relief and the placebo effect. PMID- 15653046 TI - [HIV mother to child transmission: improvement and future]. PMID- 15653047 TI - [Treatment strategies for neonatal pulmonary morbidity in Europe]. AB - OBJECTIVES: To survey practices in 14 European countries and to describe strategies for the prevention and treatment of pulmonary morbidity in very preterm newborns. METHODS: Questionnaires covering the use of prenatal steroids, surfactant and postnatal steroids were sent in 1999-2000 to every neonatal unit taking very preterm newborns in charge in population-based areas covering at least 20,000 births annually. One questionnaire was sent by surveyed unit. RESULTS: Results are given concerning these three treatments and compared to evidence based recommendations. CONCLUSION: Antenatal steroids were given at recommended terms. Surfactant was prescribed with respect of best practices. Postnatal steroids utilisation was not well described. PMID- 15653048 TI - [The pediatrician's waiting room: a place for health education?]. AB - OBJECTIVES: To assess the presence and impact of health education messages in pediatricians' waiting rooms. METHODS: In September 2001, 81 pediatricians completed a questionnaire about the furnishings and equipment in their waiting rooms. They also distributed a questionnaire about waiting room health education messages to parents, to be completed at home. RESULTS: The analysis considered 1830 questionnaires. Health education messages were posted in 91% of the waiting rooms and most frequently concerned children's accidents, vaccines, hygiene and nutrition. The best topics that the parents remembered involved child neglect, antibiotic therapy, AIDS and other sexually transmitted diseases, and nutrition. Although memorization of the messages was not influenced by duration of the wait, it was higher among parents who had previously visited the pediatrician. These messages led 14% of the parents to discuss them spontaneously with their pediatrician. CONCLUSION: Health education messages are posted in nearly all the waiting rooms studied; parents remember them in a variable and rather inexplicable order. Posting these messages demonstrates the pediatricians' willingness to be involved - beyond the simple consultation - in the health education of children and families. Identifying the real impact of these messages would require further study. PMID- 15653049 TI - [Acute osteomyelitis and septic arthritis in children: one year experience]. AB - To describe bacteriologic epidemiology of bone and joint infections, a total of 52 osteomyelitis, 52 arthritis and 20 osteoarthritis of children aged one month to 15 years during a one-year period (2001) were included in a retrospective unicentric review. The mean age was 3,9 +/-3,6 years. Fever and pain were the most common clinical symptoms. The site of infection was single in 95%, involving lower extremities in 80%. Bone scintigraphy was abnormal in 71% of osteomyelitis. Positive cultures was obtained in 29% of all cases (blood cultures: 20%, aspiration cultures: 29%), but in 42% of cases which have both blood and aspiration cultures. Thirty-six bacteria were identified: 19 Staphylococcus (14 aureus), ten Streptococcus (four pneumoniae), three Salmonella, three Kingella kingae, one Moraxella. All the isolates were susceptible to the empiric antibiotic therapy. Outcome was good in 100% of osteomyelitis and in 96% of arthritis. PMID- 15653050 TI - [Salmonella infections in children: a retrospective study over a four-year period]. AB - OBJECTIVES: Epidemiological state of salmonella infections in Languedoc Roussillon (France) and discussion of therapeutic indications. MATERIAL AND METHOD: Retrospective study over a four-year period of paediatric salmonella infections requiring hospitalisation in Montpellier University Hospital. Analysis of clinical, bacteriological, laboratory and therapeutic data. RESULTS: One hundred and sixty nine cases of paediatric salmonella infections have been investigated. The clinical triad consisted of diarrhea, abdominal pain and fever. Six severe cases have been observed and seemed to have been independent of patient's background. Laboratory findings showed inconstantly hyperleucocytosis and increase in CRP levels. Stool analysis remained the reference laboratory test to detect salmonella. Two major strains of salmonella have been identified: Salmonella enterica subsp. enterica serovar Typhimurium and Enteriditis. Treatments have been essentially symptomatic but 20% of the children (n=34) required antibiotic therapy. A favourable outcome was observed in all the children. DISCUSSION: Incidence of salmonella infections is increasing in France, particularly in the Languedoc-Roussillon area. Diagnosis of mild forms is generally easy but certain patient background seems to expose to severe forms. Treatment of salmonella infections is mainly symptomatic, only severe or specific cases (young age, immunodepression...) requiring antibiotic treatment. The growing antibiotic use and the increasing of resistance are currently the main problems in the management of salmonella infections. Those infections remain a public health problem in Languedoc-Roussillon. PMID- 15653051 TI - [Caries free smile: a dental health educational programme]. AB - A multidisciplinary team associating members from the hospital, national education and local council sectors prepared, and led, a carie-free smile educational campaign in Toulouse. OBJECTIVES: The aim was two-fold: to teach children how to manage their routine oral hygiene while giving a positive image to health care-hence the carie-free smile theme chosen by the team; secondly, to promote an awareness campaign targeting professionals in the education sector (public health sector, independent paediatricians and odontologists) who together constitute the transmitters of our initiative. MATERIAL AND METHODS: The action involved 12,000 children in the 5-8-year age group from 76 public and private schools, 36 kindergarten and primary school leisure centres, six holiday centres together with infant patients from the children's hospital, all of whom were volunteers in the scheme. The team produced back-up material in the form of posters, booklets and stickers; there was also a website dealing with oral hygiene themes, caries and their treatment. Those taking part included practising dentists and students of dentistry. Before the presentation, posters were sent to teachers and other educational partners so as to prepare the children; this included an interactive phase in the presence of the teacher, and a brushing session. Each child was given a booklet, a sticker and a toothbrush. RESULTS: By means of a poll organised among the partners the impact of the campaign could be assessed: firstly, on teachers and children by evaluating their motivation in the setting up of the toothbrushing session in 17 classes following the midday meal, thus appraising their appreciation of the visual material, and secondly, to the dentistry students: the future dentists had noted disparities in oral hygiene practices according to residential area, and thus could appreciate the importance of early provision in the school curriculum, as well as the value of accomplishing this health education task in the daily exercise of their profession. PMID- 15653052 TI - [Severe acute pancreatitis in children receiving asparaginase: multicenter retrospective study]. AB - Asparaginase is frequently used in the treatment of lymphoblastic malignancies in children and is a major cause of drug-induced acute pancreatitis. Severe cases of iatrogenic pancreatitis are uncommon but potentially lethal, and represent a diagnostic and therapeutic challenge. PATIENTS AND METHOD: We have retrospectively collected pediatric cases of severe acute pancreatitis induced by asparaginase, having occurred since January 1996 in participating centers from France and Belgium. RESULTS: Eleven patients, between four and 15 years old, have been included. Pancreatitis has been observed in all treatment phases, after 6 to 21 doses of asparaginase, 2 to 16 days after the last injection. Circulatory collapse (5/11), insulin-dependent diabetes (6/11) and pancreatic pseudokysts (7/11) were the major complications. Non-surgical treatment mainly included digestive rest, broad-spectrum antibiotic therapy and prolonged use of morphine. Asparaginase has been eventually reintroduced in three cases, and has caused a recurrence of pancreatitis in two of them. CONCLUSION: Intensive supportive management should enable a favourable outcome in most cases of acute pancreatitis induced by asparaginase in children. There is no way to predict the occurrence of this adverse event. Re-use of asparaginase should probably be ruled out. PMID- 15653053 TI - [Neonatal renal venous thrombosis following an electrical shock in pregnancy]. AB - Neonatal renal venous thrombosis may result in severe morbidity. Predisposing conditions are well known. We report the case of an unusual and early neonatal renal venous thrombosis. The mother received an electrical shock at 34 weeks gestation. This case demonstrates that maternal electrical shock effect on the fetus should be early investigated. PMID- 15653054 TI - [Maxillary and sphenoid sinusitis complicated by acute inflammatory optic neuritis in a 12-year-old patient]. AB - Acute inflammatory optic neuritis (retrobulbar optic neuritis) is an uncommon but described complication of acute sinusitis. It is due to a close anatomic relationship between the sinus and the optic nerve. Inflammatory mechanism is dominant with a possible bilateral neuritis, especially among children. In the reported case, as usual in childhood, visual acuity loss has been particularly severe, and recovery slow (it takes usually 6 weeks) but almost complete. Primary treatment is medical, with antibiotics and corticosteroids that must be prescribed for a prolonged time. PMID- 15653055 TI - [MURCS association: a challenging diagnosis]. AB - MURCS association includes Mullerrian duct aplasia-hypoplasia (MU), renal malformations (R) and cervicothoracic somite dysplasia (CS). This rare disease (1/50 000 females) is sporadic and of unknown aetiology. The reported case is the first one with additional esophageal atresia and ovarian mature teratoma. Esophageal atresia first led to the diagnosis of VACTERL association, which is more frequent and well known, showing that the identification of such malformative association may be challenging. The presence of mullerrian abnormality has allowed the diagnosis of MURCS association, as there is no mullerrian hypoplasia in VACTERL association. Therefore the association of isolated or combined renal and cervical malformation with VACTERL features should lead to the search for mullerrian abnormalities. PMID- 15653056 TI - [Malaria chemoprophylaxis in traveling children]. AB - In France, 4,000 imported malaria cases are reported each year (7,000 to 8,000 estimated). Chemoprophylaxis is essential for prevention in travelers. When malaria is susceptible to chloroquine, this drug (Nivaquine) has to be used. It is given daily in France (1.5 mg/kg per day), from departure to four weeks after return. When low levels of chloroquino-resistance are reported, French authorities recommend the use of chloroquine + proguanil (Savarine) if the body weight is >50 kg or Nivaquine) + Paludrine), if <50 kg), or atovaquone + proguanil (Malarone). Nivaquine) (1.5 mg/kg per day) and Paludrine) (3 mg/kg per day) have to be pursued for one month after return, although Malarone) (1 pediatric tablet/10 kg per day, in children >10 kg weight) may be disrupted after one single week. Adverse events are rarer with atovaquone + proguanil, than with chloroquine + proguanil. When chloroquino-resistance is high, Malarone) or mefloquine (Lariam) are used. Weekly drug regimen is recommended with mefloquine (5 mg/kg per weight) for the travel duration and four weeks after return and the drug tolerance is good in pediatric prophylaxis. Doxycycline is used under conditions in children >8 years of age. New drugs as for tafenoquine, an amino-8 quinoleine, might enhance patients compliance if given monthly. PMID- 15653057 TI - [Congenital nephrogenic diabetes insipidus]. AB - Nephrogenic diabetes insipidus is a rare hereditary disease, characterized by a resistance of the renal collecting duct to the action of the antidiuretic hormone, arginine vasopressin, responsible for the inability of the kidney to concentrate urine. More than 90% of the patients are males and have the X-linked recessive form of the disease usually presenting with polyuria and polydipsia in infancy. This mode of inheritance is related to mutations in the V(2) receptor gene, located in the Xq28 chromosomal region. Less than 10% of the patients have an autosomal-recessive or an autosomal-dominant mode of inheritance with clinical manifestations occurring in males and females, related to mutations in the aquaporin-2 gene, located in chromosome region 12q13. The aim of the treatment is to avoid chronic and acute dehydration episodes. It remains symptomatic, mainly based on an hypoosmotic diet and the use of hydrochlorothiazide and indomethacin. Recent findings showed that pharmacological chaperones, such as V(2) nonpeptide antagonists, are able to rescue some of the V(2) receptor mutants and could be useful tools for treatment in the future. PMID- 15653059 TI - [Acute renal failure in childhood malaria]. PMID- 15653060 TI - [Cancer incidence in children from the Cancer Registry in Brazzaville]. PMID- 15653061 TI - [Pneumopericardium: a rare complication of meconium aspiration syndrome]. PMID- 15653062 TI - [Metabolic diseases are the first indication of liver transplantation in children in the Middle East]. PMID- 15653063 TI - [Evaluation of a non-invasive test for detection of Helicobacter pylori antigen in stool from children]. PMID- 15653064 TI - [Lameness in children: diagnostic process]. AB - A complete examination including the lower limb, the spine as well as a neurological exam must be done for every limp in children to eliminate an infectious process in particular. The complementary exams are guided by this clinical examination but standard assessment necessitate at least plain X-rays with an AP view of the pelvis and a lateral view of the hips, a blood count with the ESR and the CRP to eliminate rapidly a tumoral or an infectious process which requires quickly an appropriate treatment. In particular cases, this work-up must be repeated in association with other complementary exams to find the etiologic diagnosis. Bone pains in children are often secondary to a true pathologic process. All these examination and work-up must be done before classified them as "growing pain". PMID- 15653065 TI - [Vaccine adjuvants and macrophagic myofasciitis]. AB - Aluminium-based adjuvants have been used throughout the world since 1926, and their safety profile is such that they have long been the sole adjuvants registered for clinical use. Their safety has nevertheless been questioned in France over the last few years following the demonstration that aluminium could persist for prolonged periods at the injection site, within macrophages gathered around the muscular fibres and forming a microscopic histological lesion called "macrophagic myofasciitis (MMF)". This image has been observed in patients undergoing a deltoid muscular biopsy for diagnostic purposes of various symptoms essentially including muscular pain and fatigue, in association with a large panel of various symptoms and diseases, including those of an autoimmune nature. Studies of the clinical, biological and epidemiological characteristics undertaken to identify a possible association between the MMF histological image and a systematic disease have remained negative. As of today, available evidence indicates that although vaccine aluminium may persist at the site of injection for years ("vaccine tattoo"), this does not reflect the existence of a diffuse inflammatory muscular disease and is not associated with a specific clinical disease. The existence of sampling bias inherent to the complexity of the clinical and pathological diagnoses remains the most likely hypothesis. PMID- 15653066 TI - [The voluminous primitive psoas abscess in children: report of three cases]. AB - The primitive psoas abscess is a pathology extremely rare in the child. The diagnosis is difficult and often delayed. Their treatment requires a local and general sterilization of the infection. We report three observations of primary abscess of the psoas in the child. The clinic presentation is uncommon. Ultrasonography keeps an important place. The biology objective an infectious and inflammatory syndrome. Histology permits to put in evidence all shapes association of lesions. The bacteriological survey of the pus after drainage is only positive in one case. Surgical drainage with bactericidal antibiotherapy permit a definitive recovery. PMID- 15653067 TI - [Practical advice for exercise-induced asthma in children: experience of the exercise training centre of Necker-Enfants malades hospital]. AB - Now, to care exercise-induced asthma is not only to prescribe drugs. It is a global and interdisciplinary approach: the pulmonary rehabilitation, matching a therapeutic education and a physical training, with the goal of promoting a regular physical activity in the asthmatic child, achieving physiological benefits and improvement of quality of life. Getting from the experience of Necker-Enfants Malades Hospital's Training Centre, a few advises encourage the physical practice of the asthmatic child, and decrease risks of exercise-induced asthma: optimisation of treatments;progressive beginning and end of exercises; use of the diaphragmatic breathing, keeping up with the exercise; use of the ventilatory threshold (or dysponea threshold) as intensity of the aerobic training; practice of different activities promoting play and conviviality in sports and allowing the integration of sports in the daily life of the asthmatic child. PMID- 15653068 TI - [Nonpharmacologic management of pain in neonates]. AB - It has taken a staggering amount of time for the medical community to realize that new-borns are able to feel pain. The treatment of neonatal pain during procedures has become mandatory, not only for humanitarian reasons which could alone justify the soothing of pain in these infants but also because repeated and prolonged pain may have long-term consequences in neonates. Nonpharmacological interventions which comprise environmental and behavioral interventions have a wide applicability for neonatal pain management alone or in combination with pharmacological treatments. These interventions are not necessarily substitutes or alternatives for pharmacological interventions but are complimentary. Nonpharmacological interventions can reduce neonatal pain indirectly by reducing the total amount of noxious stimuli to which infants are exposed and directly, by blocking nociceptive transduction or transmission or activation of descending inhibitory pathways or by activating attention and arousal systems that modulate pain. This article describes prevention, environmental interventions, and behavioral strategies. Within the behavioral strategies, sweet solutions, especially sucrose and glucose, with or without non-nutritive sucking, skin to skin contact, and breastfeeding during procedures have been studied and their analgesic efficacy has been shown. A practical approach is described hereafter. Give 1-2 ml of oral sucrose or glucose 30% at 2 min before a minor painful procedures in term neonates or neonates weighing more than 2500 g. The analgesic efficacy of sucrose and glucose seems similar. For preterm neonates weighing less than 2500 g, give 0.5 ml of oral sucrose or glucose 30%, and for those weighing less than 1500 g, 0.3 ml of oral sucrose or glucose 30%. Since a synergistic effect has been shown for the association of sweet solutions and pacifiers, give a pacifier together with sweet solutions. For term neonates that are breastfed, consider breastfeeding during procedures. These nonpharmacological interventions are suitable for minor procedures. They should not constitute the sole analgesic when performing more invasive procedures. PMID- 15653070 TI - Cytokinesis: the great divide. PMID- 15653069 TI - [Radiological case study]. PMID- 15653071 TI - Ubiquitination of intracellular bacteria: a new bacteria-sensing system? AB - Ubiquitination is a protein modification generally used by cells to tag proteins that are destined for proteasomal degradation. In a recent article, Perrin et al. reported that the ubiquitination system has a role in the recognition of bacterial pathogens in the cytosol of mammalian cells. They showed that polyubiquitinated proteins accumulate on the surface of cytosolic Salmonella typhimurium. In macrophages, but not epithelial cells, proteasomes become associated with the surface of cytosolic bacteria. The authors proposed that the ubiquitin-proteasome machinery might be implicated indirectly in bacterial clearance. PMID- 15653072 TI - A passionate kiss, then run: exocytosis and recycling of IgG by FcRn. AB - The MHC-class-I-like Fcgamma receptor FcRn recycles immunoglobulin (Ig)G from most cells and transports it bi-directionally across epithelial barriers to affect systemic and mucosal immunity. Recent studies have shown that FcRn rescues IgG from intracellular lysosomal degradation by recycling it from the sorting endosome to the cell surface. Most recycling vesicles fuse completely with the plasma membrane in a classical pattern of exocytosis. Similar to the process seen for neurotransmitter release at synaptic junctions, other vesicles fuse only partially, releasing FcRn-IgG complexes to mix into the plasma membrane in cycles of 3-4s over prolonged periods of time. PMID- 15653073 TI - Split decisions: coordinating cytokinesis in yeast. AB - Cytokinesis in eukaryotes involves the regulated assembly and contraction of a ring comprising filamentous (F)-actin and myosin II. Assembly of the contractile ring occurs through the accumulation of cortical cues at the specified division plane, followed by recruitment of F-actin, myosin II and accessory proteins involved in generating the mature ring. Ring contraction is temporally regulated to occur only after chromosome segregation and, in yeast, it is controlled by a conserved signaling cascade that becomes active only after Cdk1-Cyclin-B inactivation. In this article (which is part of the Cytokinesis series), we discuss recent studies that have begun to clarify both the spatial and the temporal order of ring assembly and that have illuminated the signals that trigger ring contraction in yeast. These studies add to the growing knowledge of the processes that control eukaryotic cell division. PMID- 15653074 TI - Disassembling adherens junctions: breaking up is hard to do. AB - Epithelial cells regulate their contacts with neighboring cells during embryonic development and in disease states such as tumor metastasis. The intercellular adherens junctions (AJs) are specialized subapical structures that function as principle mediators of cell-cell adhesion. Their disassembly correlates with a loss of cell-cell contact and an acquisition of migratory potential. Regulation of the expression of AJ components by gene transcription can dictate the stability of intercellular adhesive contacts, and accruing evidence indicates that the coordinated regulation of cellular processes such as membrane trafficking and cytoskeletal remodeling can also result in the effective dissolution of AJs. Studies of the posttranscriptional mechanisms by which adhesive interactions are regulated in response to growth factors and/or developmental cues have opened new avenues for investigating cell-cell adhesion during development and in disease. PMID- 15653075 TI - Mobilizing the proteolytic machine: cell biological roles of proteasome activators and inhibitors. AB - Proteasomes perform the majority of proteolysis that occurs in the cytosol and nucleus of eukaryotic cells and, thereby, perform crucial roles in cellular regulation and homeostasis. Isolated proteasomes are inactive because substrates cannot access the proteolytic sites. PA28 and PA200 are activators that bind to proteasomes and stimulate the hydrolysis of peptides. Several protein inhibitors of the proteasome have also been identified, and the properties of these activators and inhibitors have been characterized biochemically. By contrast, their physiological roles--which have been reported to include production of antigenic peptides, proteasome assembly and DNA repair--are controversial. In this article, we briefly review the biochemical data and discuss the possible biological roles of PA28, PA200 and proteasome inhibitors. PMID- 15653076 TI - Adhesion remodeling underlying tissue morphogenesis. AB - Cell-adhesion molecules localized at adherens junctions (AJs) maintain the polarized architecture of epithelial cells but limit their movements. The morphogenesis of a developing epithelium is associated with the control of both cell shape and cell contacts. Epithelial cells remodel their contacts, and intercellular adhesion controlled by cadherin molecules is spatially and temporally regulated. Cell shape depends, in part, on the regulation of cell adhesion between different groups of cells. Patterned epithelial cell movements such as those that occur during cell intercalation--a universal process whereby cells exchange neighbors--rely on the polarized remodeling of AJs. Recent studies show that the understanding of adhesion will benefit from studies of developing organisms in which adhesion is regulated. PMID- 15653077 TI - New views of cells in 3D: an introduction to electron tomography. AB - The most important goal of structural cell biology is to elucidate the mechanisms of the processes of life. The structure of a membrane system or fibrous array, the changes in such structures over time or the localizations of enzymes relative to organelle boundaries can often illuminate associated cellular functions. To be of maximal value, structural studies should provide isotropic, 3D information about well-preserved samples at the highest possible resolution. Electron tomography can provide such information about many kinds of cells and organelles. PMID- 15653078 TI - Matrix revolutions: "tails" of basement-membrane components with angiostatic functions. AB - Angiogenesis, the creation of neovasculature from native blood vessels, is a prerequisite for many physiological and pathological processes. Recently, C terminal tail fragments of several basement-membrane proteins such as endostatin, tumstatin and endorepellin have been shown to inhibit angiogenesis. Although there seems to be little or no homology among them, a common theme is that these fragments modulate endothelial cells by distinct interactions with integrins and activate distinct intracellular signaling cascades that often lead to disruption of the actin cytoskeleton. In this article, we focus on recent advances regarding the mechanism of action of these angiostatic fragments and the emerging concept of similarities among them, with the underlying premise that appreciating these similarities might lead to improved therapeutics. PMID- 15653080 TI - Syntegration: a "more complete" knowledge-based approach to the practice of medicine--North American Spine Society Presidential Address, Chicago, IL. PMID- 15653081 TI - Quantitative analysis of gene expression in a rabbit model of intervertebral disc degeneration by real-time polymerase chain reaction. AB - BACKGROUND CONTEXT: Serial analysis of gene expression during the course of intervertebral disc degeneration (IDD) could elucidate valuable insight into pathophysiology and provide a basis for identification of potential targets for the development of novel cellular- and gene-based therapies. However, very few previous studies described the changes in gene expression through the process of IDD using a suitable animal model. PURPOSE: To use a recently developed rabbit annular stab model and the technique of real-time reverse transcriptase polymerase chain reaction (RT-PCR) to quantify the change in expression of key rabbit-specific mRNA sequences encoding for selected extracellular matrix (ECM) products, catabolic, anabolic, and anti-catabolic factors in normal and stabbed discs. STUDY DESIGN: Gene expression analyses were performed to characterize a slowly progressive and reproducible animal model of IDD using real-time RT-PCR. METHODS: Twelve rabbits underwent an annular stab with a 16-gauge needle to the L2-L3, L3-L4, and L4-L5 discs, and three rabbits served as sham controls. Nucleus pulposus tissues were harvested from the stabbed discs at 3, 6, 12 and 24 weeks after confirmation of degenerative changes by magnetic resonance imaging (MRI) scan. Real-time RT-PCR was performed with the use of rabbit-specific primers for 1) extracellular matrix (ECM) component genes: collagen type Ia and IIa, and aggrecan; 2) catabolic genes: matrix metalloprotease-3 (MMP-3), inducible nitric oxide synthase (iNOS), and interleukin-1beta (IL-1beta); 3) anabolic growth genes: bone morphogenic protein-2, and -7 (BMP-2, -7), transforming growth factor beta1 (TGF-beta1), and insulin-like growth factor-1 (IGF-1); and 4) anti catabolic gene: tissue inhibitor of metalloprotease-1 (TIMP-1). These data were normalized to mRNA levels of glyceraldehyde phosphate dehydrogenase (GAPDH), a constitutively expressed gene. RESULTS: The MRI images confirmed progressive decline in the nucleus pulposus area of high T2 signal and in the signal intensity of the stabbed discs over the 24-week study period consistent with IDD. The ECM components, aggrecan and collagen type IIa mRNA levels had decreased markedly by week 3 and never recovered, whereas type Ia collagen mRNA gradually increased throughout course of degeneration. BMP-2, BMP-7 and IGF-1 mRNA were relatively decreased from weeks 3 to 6 but then increased at weeks 12 and 24 to end at a level near the preoperative level. The TIMP-1 expression fell dramatically to approximately one tenth of the preoperative level by week 3 and remained low throughout the degenerative process. The remaining results, including those from TGF-beta1 and the catabolic genes (MMP-3, IL-1beta, iNOS) demonstrated a double peak characteristic. The gene expression increased by week 3, decreased to a low level at weeks 6 and 12 and then had a second, late peak at 24 weeks. CONCLUSIONS: The gene expression profiles of ECM components and anabolic, catabolic, and anti-catabolic factors demonstrate many characteristics similar to the findings in human disc degeneration and suggest an inability of the intervertebral disc (IVD) to mount an early anabolic response to injury, thereby offering a possible explanation for the disc's lack of reparative capabilities. Catabolic genes are strongly up-regulated both early and late in degeneration, lending strong support to the hypothesis that an anabolic or catabolic imbalance plays a primary role in IDD. According to the resultant patterns, augmenting early production of BMP-2, BMP-7, IGF-1 or TIMP-1 by gene transfer techniques might possibly alter the progressive course of degeneration as seen in the stab model. The next step will be to transfer these therapeutic genes to regulate the biologic processes and ideally alter the progressive course of disc degeneration. PMID- 15653082 TI - Discographic, MRI and psychosocial determinants of low back pain disability and remission: a prospective study in subjects with benign persistent back pain. AB - BACKGROUND CONTEXT: A range of morphologic and psychosocial variables has been suggested as risk factors for serious low back pain (LBP) illness. Although the relative contributions of structural and psychosocial variables are intensely debated, the validity of differing hypotheses has proven difficult to test because the incidence of serious disabling LBP illness is low in healthy subjects. These factors dictate the requirement for large sample sizes, extensive structural imaging and extended longitudinal study. Previous studies included either small cohorts with intensive imaging testing or large population studies that do not establish a detailed morphologic baseline. PURPOSE: To establish, using a strict patient sample design, the relative contribution of structural and psychosocial determinants of serious LBP illness among subjects with no previous LBP disability or clinical LBP illness. STUDY DESIGN/SETTING: A prospective, longitudinal study of subjects with high risk factors for serious LBP as determined by structural and psychosocial characteristics. PATIENT SAMPLE: One hundred subjects with known mild persistent low back pain and a 2:1 ratio of chronic (non-lumbar) pain syndrome were recruited from a study population with a predisposition to disc degenerative disease, to undergo baseline examination, testing and 5-year follow-up. OUTCOME MEASURES: Observations were made at 6-month intervals over 4 to 6 years (mean, 5.3) for the after primary outcomes measures: episodes of serious back pain (visual analogue scale [VAS] > or =6), episodes of occupational disability less than 1 week, episodes of occupational disability for 1 week, remission episodes of all back pain symptoms at least 6 months and medical visits primarily for LBP evaluation and treatment. METHODS: Lumbar magnetic resonance imaging (MRI), lumbar provocative discography (in psychometrically normal subjects), physical examinations, medical and work histories and psychometric testing were performed at baseline. Imaging and psychometric testing were graded by blinded examiners. A scripted interview was conducted every 6 months during follow-up by independent research assistants who also were blinded to patient baseline data. The interview covered interval medical, occupational and accident or injury histories. RESULTS: Psychosocial variables strongly predicted both long- and short-term disability events, duration and health-care visits for LBP problems (p<0.0001-0.004). The likelihood of a sustained remission from the baseline persistent (subclinical) LBP appeared to be linked to occupation factors (leaving a heavy labor occupation; p=0.0001), neurophysiologic variables (chronic nonlumbar pain; p=0.0002) and psychometric profiles at baseline (DRAM and FABQ-PA; p=0.003-0.002). Of the structural findings measured only moderate or severe Modic changes of the vertebral end plate were weakly associated with an adverse outcome. A positive provocative discogram at baseline did not predict any future adverse event. CONCLUSION: The development of serious LBP disability in a cohort of subjects with both structural and psychosocial risk factors was strongly predicted by baseline psychosocial variables. Structural variables on both MRI and discography testing at baseline had only weak association with back pain episodes and no association with disability or future medical care. PMID- 15653083 TI - The long-term effect of posterolateral fusion in adult isthmic spondylolisthesis: a randomized controlled study. AB - BACKGROUND: Today there is some evidence-based medicine support for a positive short-term treatment effect of fusion in chronic low back pain in spondylolisthesis and in nonspecific degenerative lumbar spine disorders. The long-term effect is, however, unknown. PURPOSE: To determine the long-term outcome of lumbar fusion in adult isthmic spondylolisthesis. STUDY DESIGN: Prospective, randomized controlled study comparing a 1-year exercise program with instrumented and non-instrumented posterolateral fusion with average long-term follow-up of 9 years (range, 5-13). PATIENT SAMPLE: 111 patients aged 18 to 55 years with adult lumbar isthmic spondylolisthesis at L5 or L4 level of all degrees, and at least 1-year's duration of severe lumbar pain with or without sciatica. OUTCOME MEASURES: Pain and functional disability was quantified by pain (VAS), the Disability Rating Index (DRI), the Oswestry Disability Index (ODI) work status, and global assessment of outcome by the patient into much better, better, unchanged or worse. Quality of life was assessed by the SF-36. METHODS: The patients were randomly allocated to treatment with 1) a 1-year exercise program (n=34), 2) posterolateral fusion without pedicle screw instrumentation (n=37), or 3) posterolateral fusion with pedicle screw instrumentation (n=40). Long-term follow-up was obtained in 101 (91%) patients. Nine patients in the exercise group were eventually operated on. RESULTS: Longitudinal analysis: At long-term follow-up pain and functional disability were significantly better than before treatment in both surgical groups. No significant differences were observed between instrumented and non-instrumented patients in any variable studied. In the exercise group the pain was significantly reduced but not the functional disability. Compared with the 2-year follow-up a significant increase in functional disability was observed, as measured by the DRI, but not the ODI, in the surgical group at long term. In the exercise group no significant changes were observed between the 2-year and the long-term follow-up. Cross-sectional analysis: Between the surgical and conservative group no significant differences were observed in any outcome measurement at long-term follow-up except for global assessment, which was significantly better for surgical patients. Of surgical patients 76% classified the overall outcome as much better or better compared with 50% of conservatively treated patients (p=0.015). Quality of life as estimated by the SF-36 at long term was not different between treatment groups in any of the eight domains studied but was considerably lower than for the normal population. CONCLUSIONS: Posterolateral fusion in adult lumbar isthmic spondylolisthesis results ina modestly improved long-term outcome compared with a 1-year exercise program. Although the results show that some of the previously reported short-term improvement is lost at long term, patients with fusion still classify their global outcome as clearly better than conservatively treated patients. Furthermore, because the long-term outcome of the patients conservatively treated most likely reflects the natural course, one can also conclude that no considerable spontaneous improvement should be expected over time in adult patients with symptomatic isthmic spondylolisthesis. Substantial pain, functional disability and a reduced quality of life will in most patients most likely remain unaltered over many years. PMID- 15653084 TI - Restoring geometric and loading alignment of the thoracic spine with a vertebral compression fracture: effects of balloon (bone tamp) inflation and spinal extension. AB - BACKGROUND CONTEXT: In patients with osteoporosis, changes in spinal alignment after a vertebral compression fracture (VCF) are believed to increase the risk of fracture of the adjacent vertebrae. The alterations in spinal biomechanics as a result of osteoporotic VCF and the effects of deformity correction on the loads in the adjacent vertebral bodies are not fully understood. PURPOSE: To measure 1) the effect of thoracic VCFs on kyphosis (geometric alignment) and the shift of the physiologic compressive load path (loading alignment), 2) the effect of fracture reduction by balloon (bone tamp) inflation in restoring normal geometric and loading alignment and 3) the effect of spinal extension alone on fracture reduction and restoration of normal geometric and loading alignment. STUDY DESIGN/SETTING: A biomechanical study using six fresh human thoracic specimens, each consisting of three adjacent vertebrae with all soft tissues and bony structures intact. METHODS: In order to reliably create fracture, cancellous bone in the middle vertebral body was disrupted by inflation of bone tamps. After removal of the bone tamps, the specimen was compressed using bilateral loading cables until a fracture was observed with anterior vertebral body height loss of >/=25%. Fracture reduction was performed under a compressive preload of 250 N first under the application of extension moments, and then using inflatable bone tamps. The vertebral body heights, kyphotic deformity of the fractured vertebra and adjacent segments and location of compressive load (cable) path in the fractured and adjacent vertebral bodies were measured on video-fluoroscopic images. RESULTS: The VCF caused anterior wall height loss of 37+/-15%, middle height loss of 34+/-16%, segmental kyphosis increase of 14+/-7.0 degrees and vertebral kyphosis increase of 13+/-5.5 degrees (p<.05). The compressive load path shifted anteriorly by about 20% of anteroposterior end plate width in the fractured and adjacent vertebrae (p=.008). Bone tamp inflation restored the anterior wall height to 91+/-8.9%, middle-height to 91+/-14% and segmental kyphosis to within 5.6+/-5.9 degrees of prefracture values. The compressive load path returned posteriorly relative to the postfracture location in all three vertebrae (p=.004): the load path remained anterior to the prefracture location by about 9% to 11% of the anteroposterior end plate width. With application of extension moment (6.3+/-2.2 Nm) until segmental kyphosis and compressive load path were fully restored, anterior vertebral body heights were improved to 85+/ 8.6% of prefracture values. However, the middle vertebral body height was not restored and vertebral kyphotic deformity remained significantly larger than the prefracture values (p<.05). CONCLUSIONS: The anterior shift of the compressive load path in vertebral bodies adjacent to VCF can induce additional flexion moments on these vertebrae. This eccentric loading may contribute to the increased risk of new fractures in osteoporotic vertebrae adjacent to an uncorrected VCF deformity. Bone tamp inflation under a physiologic preload significantly reduced the VCF deformity (anterior and middle vertebral body heights, segmental and vertebral kyphosis) and returned the compressive load path posteriorly, approaching the prefracture alignment. Application of extension moments also was effective in restoring the prefracture geometric and loading alignment of adjacent segments, but the middle height of the fractured vertebra and vertebral kyphotic deformity were not restored with spinal extension alone. PMID- 15653085 TI - Biomechanical changes after the augmentation of experimental osteoporotic vertebral compression fractures in the cadaveric thoracic spine. AB - BACKGROUND CONTEXT: Osteoporotic compression fractures are an important public health concern, leading to significant morbidity, mortality and economic burden. Cement augmentation procedures used to treat these fractures alter the biomechanics of the fractured segment, which could promote adjacent failure. However, if alignment is improved or restored, there will be less risk of adjacent failure. PURPOSE: To determine the effects of load (compression/flexion), adjacent vertebral location (superior/inferior) and augmentation on vertebral segment stiffness and adjacent vertebral strain in the upper and lower thoracic spine. STUDY DESIGN: Human cadaveric thoracic spine segments were tested under load control before and after the creation of experimentally augmented vertebral compression fractures. METHODS: Six T1-T5 and six T8-T12 segments were obtained from eight thoracic spines with known bone mineral density (BMD). Rosette strain gauges were applied to T2, T4, T9 and T11 to measure strain adjacent to the experimental fracture sites T3 and T10. Two compression fractures were created in succession, the first in flexion preceded by a weakening defect in T3 and T10 and the second created in an adjacent vertebra in compression without prior weakening. The first fracture was reduced with the inflatable bone tamp (IBT) and augmented with cement. Compression and flexion tests were performed before and after the first fracture while measuring vertebral cortical shear strain on T2, T4, T9 and T11 and stiffness of the entire segment. Strain and stiffness were compared by using a repeated measures analysis using adjacent vertebral location (superior/inferior), augmentation and load (compression/flexion) as factors. RESULTS: The mean BMD was 0.61+/-0.11 g/cm(2) (T1-T5) and 0.78+/-0.07 g/cm(2) (T8-T12). Stiffness in compression and flexion increased with load (p<.05, and p>.27, respectively). Augmentation reduced compressive and bending stiffness (p=.23, and p=.19, respectively), whereas the adjacent vertebral strain increased (p>.11). The adjacent strain in flexion was much greater than in compression (p<.03). Cement augmentation caused greater amounts of inferior than superior adjacent strain (p>.19). The applied moment at first fracture was 2.98+/-1.28 Nm (T1-T5) and 8.44+/-1.02 Nm (T8-T12). The compressive load at second fracture was 1122+/-993 N (T1-T5) and 2906+/-1008 N (T8-T12). Adjacent vertebral strain during the second compression and flexion tests exceeded that during the first compression and flexion tests (p=.11). Adjacent vertebral strain at second fracture exceeded that at first fracture (p=.007) and was greater on the superior adjacent vertebra than the inferior (p=.47). CONCLUSION: With axial compressive loads, the addition of flexion increases fracture risk. Cement augmentation of a fractured vertebral segment reduces stiffness while increasing both the superior and inferior adjacent cortical strain. This increment in strain that is greatest on the inferior adjacent vertebra effectively redistributes loads from the superior adjacent vertebra to the inferior adjacent vertebra, sparing the superior adjacent vertebra from failure. PMID- 15653086 TI - The effect of uniform heating on the biomechanical properties of the intervertebral disc in a porcine model. AB - BACKGROUND CONTEXT: The use of minimally invasive lumbar intradiscal heating techniques, including intradiscal electro-thermal therapy (IDET), endoscopic radio-frequency annuloplasty, nucleoplasty and laser discectomy, for chronic lumbar discogenic pain and contained disc herniation has recently gained popularity. The purported therapeutic mechanisms of these interventions include subtotal nuclectomy, annular nociceptor ablation, and stabilization of the annular fibers. Basic science data elucidating the biomechanical and histomorphologic alterations of heat treatments on disc remain sparse. PURPOSE: The purpose of this study is to examine the effects of uniform heating on biomechanical properties and histomorphology of intervertebral disc tissues using a porcine model. STUDY DESIGN/SETTING: In a laboratory setting, porcine functional spinal units consisting of vertebra-nucleus pulposus-vertebra core and porcine hamstring tendons were harvested. Studies were performed on these tissue samples by uniformly heating the specimens in a constant temperature water bath. Ten porcine lumbar disc core and twenty-five porcine hamstring tendons were utilized as the subjects for this study. The effects of uniform heat treatments on disc core and hamstring tendon were measured for shrinkage, stiffness, and load to failure strength. Histomorphological study was also carried on the same specimen. METHODS: The porcine vertebra-nucleus pulposus-vertebra segments were cored to a uniform 1-cm diameter. The hamstring tendons were cut to uniform 1.2 inch lengths. The tendon specimens were divided into groups of five each and heated in constant temperature water baths of 60 degrees C, 65 degrees C, 70 degrees C, or 75 degrees C for 10 min. Unheated specimens served as controls. The disc core specimens were divided in two group of five each, and tested at room temperature or after immersion in a 70 degrees C bath. The shrinkage was monitored during immersion in the water bath. Biomechanical testing to failure was carried out using mechanical loading on an MTS servohydraulic testing machine operating under stroke control. Strength and stiffness of the tissue was determined. Histomorphology was studied by staining the specimen with hematoxylin and eosin (H&E), and examined under 200 times magnification. Non-heated controls were used for comparisons. RESULTS: The porcine hamstring tendons had no measurable shrinkage in specimens heated up to 65 degrees C. At temperatures above 65 degrees C, the shrinkage was concluded within 2 min of immersion and 70 degrees C appeared to be the optimal temperature, as temperatures higher than this did not demonstrate incremental effects. The disc core samples were heated to 70 degrees C (optimum temperature), and there appeared to be gross contraction of the disc core circumference to visual inspection, but no measurable lengthwise shrinkage could be appreciated. Histologically, the specimens demonstrated progressive loss of individual collagen fiber outline as the temperature increased. In the tendons, at 75 degrees C all of the fibers appear to be fused together, and the voids between individual collagen fibers were no longer present. Biomechanical testing revealed that the tendons undergo a substantial reduction in stiffness after heating. The mean tendon stiffness for the unheated specimens was 19,356 psi, while the corresponding value for the heated tendons was 1023 psi. These were significantly different using the paired t-test at p=0.0043. For the disc core samples, there was no significant difference in either stiffness (p=0.182) or failure strength (p=0.998) after heating. All failures occurred in mid-substance of the specimen. CONCLUSIONS: The application of uniform heating to nucleus pulposus disc core caused visible contraction of its circumference but not lengthwise shrinkage. The same heating shrinks the hamstring tendon and reduces its stiffness. Ultimate failure strength of the disc core specimen remains unchanged. The failure data was not obtainable for the tendon due to premature slippage from the fixation apparatus before failure. The results of this study fail to support a biomechanical justification for the application of uniform heat treatment to the whole intervertebral disc. Heating annulus fibrosus and nucleus pulposus separately to specific temperatures may have potential clinical benefits. PMID- 15653087 TI - Presurgical biopsychosocial factors predict multidimensional patient: outcomes of interbody cage lumbar fusion. AB - BACKGROUND CONTEXT: Interbody cage lumbar fusion (ICLF) has been advanced to improve arthrodesis; however, little attention has been given to quality of life and functional outcomes. Studies suggest that psychosocial factors may be important modifiers of low back surgical outcomes. PURPOSE: To depict outcomes of ICLF surgery across multiple dimensions and to investigate presurgical biopsychosocial predictors of these outcomes. STUDY DESIGN/SETTING: A retrospective-cohort research design was used that involved completion of presurgical medical record reviews and postsurgical telephone outcome surveys at least 18 months after surgery. Presurgical variables included in a regression model were age at the time of surgery, spinal pathophysiology rating, smoking tobacco, depression, and pursuing litigation. PATIENT SAMPLE: Seventy-three patients received ICLF, and of those 56 patients completed the outcome survey an average of 2.6 years after surgery. OUTCOME MEASURES: Outcome measures consisted of arthrodesis status, patient satisfaction, back-specific functioning, disability status, and quality of life. RESULTS: Although arthrodesis occurred in 84% of the patients, nearly half were dissatisfied with their current back condition. Functional status was worse than expected, and 38% were totally disabled at follow-up. Regression analyses revealed tobacco use, depression, and litigation were the most consistent presurgical predictors of poorer patient outcomes. CONCLUSIONS: Overall, despite a high rate of arthrodesis, ICLF was not associated with substantial improvements in patient functioning. Presurgical biopsychosocial variables predicted patient outcomes, which may help improve patient selection and possible targeted interventions. PMID- 15653088 TI - Results of nonsurgical treatment of thoracic spinal tuberculosis in adults. AB - BACKGROUND: The indications for surgery in spinal tuberculosis have been controversial, and more so recently, in the era of renewed understanding of the concept of multi-drug-resistant tuberculosis along with newer modalities of spinal instrumentation. Indications for surgery need to be redefined in this context. PURPOSE: To assess the efficacy and results of nonsurgical treatment in thoracic spinal tuberculosis in adult patients, and redefine indications for surgery. STUDY DESIGN: We present a retrospective analysis of 70 adults with thoracic spinal tuberculosis, with varying presentations, including abscesses and neurological deficits, seen at our spine clinic, in a period between August 1998 and August 2000, treated largely nonsurgically, with rewarding results. METHODS: A retrospective study was made of 70 adult patients with thoracic spinal tuberculosis presenting at our spine clinic, between August 1998 and August 2000. All patients were subjected to medical management, unless there were specific indications for surgery, as per our protocol, wherein absolute indications of surgery in adults included advanced neurological deficit (less than Grade 3 by 5, by the 5-point grading system of the Medical Research Council), neurology worsening while on antituberculous chemotherapy, diagnosis in doubt on clinicoradiological evaluation and significant kyphosis (greater than 40 degrees) on presentation. Clinical and radiological assessment of results was made by an independent observer, at a mean follow up of 40 months. RESULTS: Forty-four patients presented with abscesses, 21 of which were epidural. Seven had neurological signs of cord compression on clinical examination at presentation. Over 98% of our patients (69 of 70) were successfully treated conservatively, and none of these had any residual instability, radiculopathy or neurological compromise. Seventy-four percent had excellent to good results, with no mechanical residues of the disease, and 23% had residual kyphosis, which was clinically obvious, but biomechanically irrelevant. CONCLUSIONS: We think that tuberculous spondylodiscitis in adults can be well managed conservatively in a vast majority of cases, and indications for surgery are few and specific. PMID- 15653089 TI - Direct real-time measurement of in vivo forces in the lumbar spine. AB - BACKGROUND CONTEXT: Accurate knowledge of the mechanical loads in the lumbar spine is critical to understanding the causes of degenerative disc disease and to developing suitable treatment options and functional disc replacements. To date, only indirect methods have been used to measure the forces developed in the spine in vivo. These methods are fraught with error, and results have never been validated using direct experimental measurements. PURPOSE: The major aims of this study were to develop a methodology to directly measure, in real time, the in vivo loading in the lumbar spine, to determine if the forces developed in the lumbar spine are dependent on activity and/or posture and to assess the baboon as an animal model for human lumbar spine research based on in vivo mechanical loading. STUDY DESIGN: Real-time telemetered data were collected from sensor imbedded implants that were placed in the interbody space of the lumbar spines of two baboons. METHODS: An interbody spinal implant was designed and instrumented with strain gauges to be used as a load cell. The implant was placed anteriorly in the lumbar spine of the baboon. Strain data were collected in vivo during normal activities and transmitted by means of a telemetry system to a receiver. The forces transmitted through the implant were calculated from the measured strain based on precalibration of the load cell. Measured forces were correlated to videotaped activities to elucidate trends in force level as a function of activity and posture over a 6-week period. The procedure was repeated in a second baboon, and data were recorded for similar activities. RESULTS: Implants measured in vivo forces developed in the lumbar spine with less than 10% error. Loads in the lumbar spine are dependent on activity and posture. The maximum loads developed in the lumbar spine during normal (baboon) activities exceeded four times body weight and were recorded while animals were sitting flexed. Force data indicate similar trends between the human lumbar spine and the baboon lumbar spine. CONCLUSIONS: It is possible to monitor the real-time forces present in the lumbar spine. Force data correlate well to trends previously reported for in vivo pressure data. Results also indicate that the baboon may be an appropriate animal model for study of the human lumbar spine. PMID- 15653090 TI - Lumbar disc arthroplasty. AB - BACKGROUND CONTEXT: Painful lumbar disc degeneration is one of the most common ailments treated by spine surgeons. Currently, early disc disease and herniation are often treated with microdiscectomy. Late disc degeneration is usually treated with arthrodesis. With the advent of new technology and techniques in lumbar disc arthroplasty, interest in preserving spinal motion at degenerated motion segments has increased. The goals of lumbar disc arthroplasty are to provide long-term pain relief at the degenerated disc level, to restore disc height to protect neural elements and to preserve motion to prevent posterior facet arthropathy and adjacent segment disease. PURPOSE: The purpose of this review is to examine the anatomy and biomechanics of the lumbar motion segment to determine the features that successful disc arthroplasty prosthesis must possess. In addition, the early clinical results of three prostheses currently being used in humans are reviewed. STUDY DESIGN/SETTING: Review of the literature. METHODS: A systematic review of Medline for articles related to lumbar disc arthroplasty was conducted up to and including journal articles published in August 2003. In addition, the abstracts from the annual meetings of the North American Spine Society and Scoliosis Research Society from 1998 to 2003 were searched. The literature was then reviewed and summarized. RESULTS/CONCLUSIONS: Short-term results of lumbar disc arthroplasty as measured by pain relief and disability are good in some studies. Implants are relatively safe in the short term, and with newer designs complications are usually related to the surgical approach rather than early implant failure. Recovery times appear to be shorter than arthrodesis. Despite the relatively good early clinical results of these devices, questions remain about the long-term efficacy in pain relief and maintenance of motion, the results of randomized comparative trials with fusion and the life span of the devices. In addition, late sequelae and revision options are unknown. Current indications for lumbar disc arthroplasty are in the setting of a Food and Drug Administration trial in young, nonosteoporotic patients with one or two level symptomatic disc degeneration without severe facet arthropathy, segmental instability or neural element compression requiring a posterior decompression. PMID- 15653092 TI - Vascular injury during anterior lumbar spine surgery. PMID- 15653093 TI - bFGF activates endothelial Ca2+-activated K+ channels involving G-proteins and tyrosine kinases. AB - Activation of Ca2+-activated K+ channels (BK(Ca)) has been shown to be an important step in the basic fibroblast growth factor (bFGF)-induced proliferation of endothelial cells. In this study, we investigate the signaling cascades of BK(Ca) modulation by bFGF. Using the patch-clamp technique, bFGF (50 ng/ml) significantly increased the BK(Ca) open-state probability in cultured endothelial cells derived from human coronary arteries after 6 min (n=26, p<0.01), which lasted up the whole recording time of 60 min. After preincubation with pertussis toxin (100 ng/ml), bFGF superfusion did not cause a significant increase of BK(Ca) activity until 25 min had passed. When genistein was supplemented to the bath solution, a significant activation of BK(Ca) by bFGF was observed during a time interval of 6-20 min (n=17, p<0.01). In contrast, the addition of the inactive analogue daidzein did not change bFGF-induced activation of the BK(Ca). In conclusion, the results of the present study indicate that the early activation of the BK(Ca) by bFGF is mediated by G-protein-dependent mechanisms, whereas the later effect is due to a tyrosine kinase-dependent signaling pathway. PMID- 15653094 TI - Peroxisome proliferator-activated receptors gamma ligands and ischemia and reperfusion injury. AB - Peroxisome proliferator-activated receptors (PPARs) are members of the nuclear hormone receptor superfamily of ligand-activated transcription factors that are related to retinoid, steroid and thyroid hormone receptors. The PPAR subfamily comprises of three members, PPAR-alpha, PPAR-beta and PPAR-gamma. PPAR-gamma has recently been implicated as a regulator of cellular proliferation and inflammatory responses. There is good evidence that ligands of PPAR-gamma, including certain thiazolinediones, reduce tissue injury associated with ischemia and reperfusion. The potential utility of PPAR-gamma ligands in ischemia and reperfusion will be discussed in this review. PMID- 15653095 TI - Regulation of the spontaneous contractile activity of the portal vein by the sarcoplasmic reticulum: evidence from the phospholamban gene-ablated mouse. AB - The rapid contraction/relaxation cycles of phasic smooth muscles necessitates intracellular calcium cycling at a more rapid rate than that of tonic smooth muscles. Recent studies suggest that sarcoplasmic reticulum calcium handling is an important determinant of portal vein phasic contractions. We evaluated the importance and role of phospholamban, a protein which inhibits the sarcoplasmic reticulum (SR) calcium ATPase (SERCA), in regulating the contractility of the phasic mouse portal vein. PLB gene ablation significantly reduced the basal frequency of spontaneous mechanical activity and increased force development of the portal vein. Cyclopiazonic acid (CPA), an inhibitor of SERCA, did not significantly affect the spontaneous activity of the wild-type (WT) portal vein. CPA (1 microM) eliminated the differences in frequency and force between the PLB KO and WT, localizing the effects to the SR. The PLB-KO portal vein had a lower resting membrane potential than WT controls. There were no significant differences between WT and KO responses to charybdotoxin (250 nM), indicating that calcium-activated potassium channels do not contribute to altered KO portal vein contractility. While contractile sensitivity to acetylcholine was not different between WT and PLB-KO portal veins, force generated in response to a given concentration of acetylcholine was significantly greater in the PLB-KO portal vein, both in the absence and presence of CPA. Our results confirm that SR activity can play a major role in modulating the frequency of the spontaneous mechanical activity of portal veins and removal of PLB inhibition of the SR calcium ATPase has significant effects on the spontaneous activity of the portal vein. PMID- 15653096 TI - Apoptosis of human umbilical vein endothelial cells induced by artesunate. AB - Artesunate (ART), a semi-synthetic derivative of artemisinin isolated from the traditional Chinese herb Artemisia annua, is an effective novel antimalarial drug. The present study investigated the apoptotic activity of artesunate in cultured human umbilical vein endothelial cell (HUVEC) by means of nuclear staining, DNA agarose gel electrophoresis, and flow cytometry. The observations also indicated that artesunate induced apoptosis of HUVEC in a concentration dependent and time-dependent manner. A Western immunoblot analysis showed down regulation of the bcl-2 protein and up-regulation of the bax protein in the artesunate-treated HUVEC. Ca2+ in cells was evaluated by fluorescent spectrophotometer using Fura 2-AM as probe. These results suggest that artesunate may be a potential apoptosis-inducing agent for endothelial cells. PMID- 15653097 TI - AAT as a diagnostic tool. AB - Serum alpha-1-antitrypsin (AAT) concentration can be affected by both inflammatory and non-inflammatory conditions. This paper characterizes the nature of AAT in physiology and pathologic deficiency and increasing states. The relationships between the AAT concentration in different clinical materials (serum, urine, faeces) and various diseases connected with different organs were analyzed. PMID- 15653098 TI - Fatty acid-binding proteins as plasma markers of tissue injury. AB - BACKGROUND: One of the novel and promising plasma markers for detection of tissue injury is the family of 15 kDa cytoplasmic fatty acid-binding proteins of which various tissue-specific types occur. AIMS AND OBJECTIVES: The present status of heart-type fatty acid-binding protein (H-FABP) as a diagnostic and prognostic marker for acute and chronic cardiac injury, as well as the preliminary diagnostic use of other types of FABP for detecting injury in other organs, is reviewed. METHODS: This review is based on an overview of the literature on clinical diagnostics of various forms of organ injury, and uses additional literature on physiological aspects relevant for the interpretation of plasma marker concentrations. RESULTS: H-FABP not only proves to be an excellent early marker for cardiac injury in acute coronary syndromes, but also allows detection of minor myocardial injury in heart failure and unstable angina. Preliminary results indicate that sensitivity, rule-out power and prognostic value of H-FABP in cardiac injury surpass the performance of the standard early marker myoglobin. The liver only contains liver-type FABP (L-FABP), but co-expression of H-FABP and L-FABP occurs in the kidney. Similarly, intestinal-type FABP (I-FABP) and L-FABP are found in intestines, and brain-type FABP (B-FABP) and H-FABP occur in the brain. Preliminary but promising applications of these proteins have been demonstrated for liver rejection, viability selection of kidneys from non-heart beating donors (NHBD), inflammatory and ischemic bowel disease, traumatic brain injury and in the prevention of muscle injury in trained athletes. CONCLUSIONS: Further study of the diagnostic and prognostic use of various FABP types is warranted, but their clinical application will require further commercialization of automated and rapid assays. PMID- 15653099 TI - Measurement of paraoxonase (PON1) status as a potential biomarker of susceptibility to organophosphate toxicity. AB - Organophosphorus (OP) compounds are still among the most widely used insecticides, and their main mechanism of acute toxicity is associated with inhibition of acetylcholinesterase. Measurements of urine metabolites and of blood cholinesterase activity are established biomarkers of exposure to OPs and of early biological effects. In recent years, increasing attention has been given to biomarkers of susceptibility to OP toxicity. Here we discuss the polymorphisms of paraoxonase (PON1), a liver and serum enzyme that hydrolyzes a number of OP compounds, and its role in modulating the toxicity of OPs. We stress the importance of determining PON1 status, which encompasses the PON1192Q/R polymorphism (that affects catalytic ability toward different substrates) and PON1 levels (which are modulated in part by a C-108T polymorphism) over straight genotyping. Epidemiological studies on OP-exposed workers that include assessment of PON1 status to validate in human populations the role of PON1 as a determinant of susceptibility to OPs, as indicated by animal studies, are needed. Documentation of exposure and of early health effects would be most relevant to increase the predictive value of the test. PMID- 15653100 TI - Neurotrophins in clinical diagnostics: pathophysiology and laboratory investigation. AB - There is now growing evidence that a number of multifunctional signaling molecules, originally discovered as signal molecules in specific cells, exert their effects in various other tissue compartments. Neurotrophins, a class of homologues growth factors initially discovered to promote neuronal growth and survival, display such a dual activity and contribute to the development of a variety of non-neuronal tissues. Nowadays, several examples of essential non neuronal functions played by neurotrophins and of variations of neurotrophin expression that accompany these processes can be presented. As will be shown, neurotrophins are found in many body tissues produced by a variety of non neuronal cell types such as immune cells, adipocytes, endothelia, epithelia, fibroblasts, keratinocytes and endocrine cells. Assuming a general role as growth and survival factors, changes in neurotrophin expression may reflect physiological or pathological processes, such as activation, proliferation or repair followed by injury in the tissues. Neurotrophins were also present in the systemic blood circulation and variations in blood concentrations indicate vascular as well as peripheral production. In this review, we will discuss changes in local and systemic neurotrophin concentrations as well as their known pathophysiological relationship in various inflammatory and non-inflammatory disorders. Beside the nervous system, these will include diseases of the airways, skin and joints as well as systemic autoimmune diseases. Furthermore, new aspects of neurotrophin actions in maintenance of body energy balance and in reproductive endocrinology will be presented. PMID- 15653101 TI - Plasma adenosine concentrations increase in women with hyperemesis gravidarum. AB - BACKGROUND: Adenosine is an important metabolic modulator and its concentrations are affected by sympathetic nerve stimulation and cytokine production. Since hyperemesis gravidarum is characterized by overactivation of sympathetic nerves and enhanced production of tumor necrosis factor (TNF)-alpha, plasma adenosine concentrations may be altered. The present study evaluated plasma adenosine concentrations and their relation with norepinephrine and TNF-alpha concentrations in hyperemesis gravidarum. METHODS: Plasma concentrations of adenosine, norepinephrine, and TNF-alpha were measured in 34 healthy nonpregnant women, and 34 women with hyperemesis gravidarum and normal pregnancies, matched for age, parity and gestational week. The relationships between plasma adenosine, and norepinephrine and TNF-alpha concentrations in hyperemesis gravidarum were evaluated. RESULTS: Mean plasma adenosine, norepinephrine, and TNF-alpha concentrations were significantly increased in women with hyperemesis gravidarum compared to those in nonpregnant and normal pregnant women (p<0.05). The increase in plasma adenosine concentrations correlated with the increase of norepinephrine and TNF-alpha in hyperemesis gravidarum (r=0.50, p<0.05 and r=0.43, p<0.05, respectively). CONCLUSIONS: Adenosine is an established suppressor of excessive sympathetic nerves activation and cytokine production, so the increase in plasma adenosine in hyperemesis gravidarum might serve to counteract further progression of hyperemesis gravidarum. PMID- 15653102 TI - Quantitation of long-chain acylcarnitines by HPLC/fluorescence detection: application to plasma and tissue specimens from patients with carnitine palmitoyltransferase-II deficiency. AB - BACKGROUND: Carnitine palmitoyltransferase-II deficiency (CPT-II deficiency) is a rare disorder of lipid metabolism, in which the accumulation of long-chain acylcarnitines is a diagnostic marker. HPLC with fluorescence detection is an attractive analysis method due to its favorable combination of sensitivity, specificity, ease of analysis and minimal capital equipment costs. METHODS: Long chain acylcarnitines were isolated from tissue homogenates (0.5-2 mg wet weight) or plasma (50 microl) using silica gel columns and derivatized with 2-(2,3 naphthalimino)ethyl trifluoromethanesulfonate. Quantitation was by HPLC and fluorescence detection with standard curves (0.0-5.0 nmol/ml) for myristoyl-, palmitoleoyl-, palmitoyl-, oleoyl- and stearoylcarnitine using heptadecanoylcarnitine as the internal standard. RESULTS: Significantly greater amounts of long-chain acylcarnitines were quantified in patients with CPT-II deficiency when compared to controls; e.g. (nmol/ml in patient plasma, controls mean+/-standard deviation): myristoylcarnitine (0.3, not detectable), palmitoleoylcarnitine (0.5, 0.1+/-0.1), palmitoylcarnitine (0.9, 0.1+/-0.0), oleoylcarnitine (3.0, 0.2+/-0.1), stearoylcarnitine (0.4, not detectable). CONCLUSIONS: This method can be used to quantitate long-chain acylcarnitines, illustrating their accumulation in CPT-II deficiency. The analysis was accomplished using inexpensive and widely available instrumentation and is appropriate for research investigators who require precise quantitation of long chain acylcarnitines in complex biological samples. PMID- 15653103 TI - Protein interference in thyroid assays: an in vitro study with in vivo consequences. AB - BACKGROUND: Pathological concentration of plasma proteins may cause problems in immunoanalytics. The low triiodotyronine (T3) and thyroxine (T4) levels, frequently found in seriously ill patients, may be ascribed either to laboratory artifact due to the lower thyroid hormone binding capacity or to a compensatory response of the organism to the disease. METHODS: The authors performed an in vitro experiment, in which sera of seriously ill patients with either low immunoglobulin G (IgG), and/or low albumin levels were investigated for free thyroid hormones (fT3, fT4) following stepwise adjustment of the serum IgG and/or albumin. All two hormones were measured with two different automated immunoassays: the microparticle enzyme immunoassay (MEIA) with two steps (AxSym, Abbott, USA) and the electrochemiluminescence immunoassay (ECLIA). RESULTS: The bias of fT3 and fT4 exhibited positive correlations with serum IgG and albumin. The bias of fT3 was more pronounced than that of fT4 following the addition of albumin (40-150% and 10-40%, respectively) as well as following the addition of IgG (8-30% and 0-8%, respectively). The MEIA method was more sensitively affected in case of fT4, whereas the bias of fT3 was more influenced in the ECLIA assay. In MEIA assay, the influence of albumin on the bias of fT3 and fT4 was stronger if serum IgG levels were low. CONCLUSION: The results confirm that pathological thyroid findings in seriously ill patients may largely be ascribed to some laboratory artifacts. PMID- 15653104 TI - Prevalence of the metabolic syndrome using the Third Report of the National Cholesterol Educational Program Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults (ATP III) and the modified ATP III definitions for Japanese and Mongolians. AB - BACKGROUND: A clustering of insulin resistance, hypertension, and dyslipidemia has been labeled as metabolic syndrome. Asians have a lower frequency of obesity than do Caucasians but have an increasing tendency toward metabolic syndrome. METHODS: We conducted a cross-sectional study of individuals aged 30-60 years. We analyzed the health data of 596 Japanese and Mongolians for metabolic syndrome based on the Third Report of the National Cholesterol Educational Program Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults (ATP III) definition and the three modified ATP III definitions. RESULTS: The prevalence of metabolic syndrome using ATP III criteria was 6% for the Japanese and 12% for the Mongolians, a remarkable lower prevalence relative to the reported prevalence in the United States. With the exception of visceral obesity, the prevalences of individual metabolic abnormalities within each of the two Asian groups were similar to each other and to reported rates of prevalence in the United States. CONCLUSIONS: A universal metabolic syndrome definition is inappropriate for comparisons of metabolic syndrome among Asian ethnic groups. We believe that the ATP III index for visceral obesity should be adjusted for Asian populations. PMID- 15653105 TI - Antioxidants and methimazole in the treatment of Graves' disease: effect on urinary malondialdehyde levels. AB - BACKGROUND: We have postulated that metabolic oxidation could be the source of signs and symptoms of hyperthyroidism. The present study was designed to evaluate urinary malondialdehyde levels in Graves' disease and compare this oxidative stress biomarker with the clinical evolution of patients suffering this illness. METHODS: We evaluated the concentration of urinary and serum malondialdehyde (MDA) in 36 patients with Graves' disease. Patients were treated with the antithyroid drug methimazole (MMI; Group A) or antioxidant mixture (200 mg vitamin E, 3 mg beta-carotene, 250 mg vitamin C, 1 mg Cu, 7.5 mg Zn, 1.5 mg Mn, and 15 microg Se; Group B). RESULTS: MDA concentrations were higher in hyperthyroid patients compared to euthyroid controls, and a positive correlation was observed between serum and urinary MDA levels. Group A decreased urinary MDA to control values. There was a positive correlation between the clinical score and the heart rate of patients with urinary MDA before and during the treatment with MMI (Group A). Similar results were observed after treatment with the antioxidant mixture. CONCLUSIONS: Urinary MDA might be a good parameter in the follow-up of patients during MMI treatment. We proposed that oxidative stress correlates with signs and symptoms of hyperthyroidism. PMID- 15653106 TI - Plasma homocysteine: an independent or an interactive risk factor for coronary artery disease. AB - BACKGROUND: Coronary artery disease (CAD) is emerging as a major public health concern in most developing countries. Further conventional risk factors for CAD do not solely account for the increased mortality, particularly in Asians. Recently, increased plasma homocysteine is being considered as a risk factor, but the strength of relationship and interaction of plasma homocysteine with other risk factors is yet obscure. In this study, the association of plasma homocysteine with CAD and other risk factors was estimated. METHODS: In the present study, 100 patients of CAD and 50 controls of both sexes were included. Plasma total homocysteine (tHcy) concentrations were measured using reverse phase high-performance liquid chromatography. RESULTS: Plasma homocysteine concentrations were significantly raised in cases as compared to age-matched controls (16.57+/-6.86 and 11.47+/-5.19 micromol/l, p<0.001). On calculating relative risk (RR) of each factor by univariate analysis smoking, hypertension, plasma cholesterol and homocysteine appeared to be significant risk factors. However, on applying multiple logistic regression only the latter three emerged as independent risk factors (p<0.005). Further, strong interactive effects were observed between homocysteine levels and increasing age, hypertension and smoking. CONCLUSION: An increase in plasma homocysteine concentration confers an independent risk for CAD. It further increases the risk associated with increasing age, smoking and hypertension. Thus, increased homocysteine concentrations are a significant medical problem and effective strategies are urgently required to counter this challenge. PMID- 15653107 TI - Enhanced inflammatory response of blood monocytes to C-reactive protein in patients with unstable angina. AB - BACKGROUND: Previous studies showed that monocytes from patients with unstable coronary disease exhibit a greatly enhanced production of interleukin-6 (IL-6) in response to lipopolysaccharide and artery injury. Moreover, accumulating evidence suggest that C-reactive protein (CRP) may have direct proinflammatory effects on the cells of vascular wall. Whether this enhanced inflammatory response also exists, however, in cultured monocytes from patients with unstable angina, in response to CRP, has not been investigated. METHODS: Monocytes were isolated from blood of 15 healthy volunteers (normal control), 15 patients with stable angina, and 15 patients with unstable angina by Ficoll density gradient and were stimulated by 20 microg/ml of CRP for 24 h. Measurements of IL-6 and tumor necrosis factor-a (tnf-alpha) were performed from supernatants of cultured medium in duplicate, using a commercial assay kit. RESULTS: the data showed that IL-6 and tnf-alpha concentrations of spontaneous secretion (baseline) were higher in patients with unstable angina than in patients with stable angina and normal control (IL-6: 179+/-19 vs. 87+/-6 and 89+/-8 pg/ml, p<0.05, respectively; tnf alpha: 69+/-13 vs. 30+/-4 and 27+/-3 pg/ml, p<0.05, respectively). CRP induced the enhanced release of IL-6 and tnf-alpha by 17-fold and 23-fold increase, respectively, in patients with unstable angina, while it did about 11-fold and 19 fold increase in patients with stable angina and normal group (IL-6: 3129+/-333 vs. 991+/-134 and 987+/-102 pg/ml, p<0.01, respectively; tnf-alpha: 1554+/-784 vs. 560+/-135 and 558+/-152 pg/ml, p<0.01, respectively). CONCLUSIONS: Increased baseline concentrations of IL-6 and tnf-alpha can be a marker of the hyperresponsiveness of the inflammatory system in patients with unstable coronary disease. CRP could enhance even further this response, suggesting that a persisted and enhanced inflammatory responsiveness to CRP may be involved in the pathogenesis of unstable coronary disease. PMID- 15653108 TI - Effects of phenolic acid esters and amides on stimulus-induced reactive oxygen species production in human neutrophils. AB - BACKGROUND: Phenolic acids and their derivatives are widely distributed in plants. A series of phenolic acid esters and amides have been synthesized. METHODS: We determined the effects of phenolic acid derivatives on antiinflammatory activity against phorbol 12-myristate 13-acetate (PMA) and N formyl-methionyl-leucyl-phenylalanine (fMLP)-induced production of superoxide anion, an inflammatory mediator produced by neutrophils. RESULTS: When the cells were preincubated with phenolic acids and their derivatives, the superoxide generation induced by fMLP (1.0 micromol/l) and PMA (0.16 micromol/l) was inhibited to various degrees with compounds 1, 2 and 4 significantly suppressing such generation in a concentration-dependent manner. CONCLUSION: Phenolic acid derivatives may exert their antiinflammatory action through inhibiting superoxide generation. PMID- 15653109 TI - Prognostic value of cardiac troponin T in patients with both acute and chronic stable congestive heart failure: comparison with atrial natriuretic peptide, brain natriuretic peptide and plasma norepinephrine. AB - BACKGROUND: The prognostic value of cardiac troponin T (cTn-T) in a mixture of patients with both acute and chronic congestive heart failure (CHF), simultaneously assessed and compared with neurohormonal factors, has not yet been thoroughly evaluated. Thus, we focused on the prognostic value of cTn-T in comparison with atrial natriuretic peptide (ANP), brain natriuretic peptide (BNP) and plasma norepinephrine (PNE) in this population. METHODS: Prognostic correlates of elevation of cTn-T, ANP, BNP, PNE were analyzed in 63 acute and chronic CHF patients followed up to record worsening CHF and cardiac death. RESULTS: cTn-T (> or =0.03 microg/L) was found in 17.4% (11 of 63) of patients. cTn-T correlated with ANP, BNP, PNE. Acute CHF patients were more positive for cTn-T and BNP. In our cohort, neither cTn-T (> or =0.03 microg/L) nor PNE were associated with increased mortality and worsening HF in CHF patients. After adjustment, BNP was the only independent predictor of cardiac events (RR, 3.23; p=0.01). CONCLUSIONS: BNP emerged as the only independent predictor of cardiac events in a mixture of patients with both acute and chronic CHF, suggesting that it is the analyte that best reflects long-term prognosis in a diverse population enrolled to mirror the "real world" situation. PMID- 15653110 TI - A modified, economic, sensitive method for measuring total antioxidant capacities of human plasma and natural compounds using Indian saffron (Crocus sativus). AB - BACKGROUND: Free radicals are involved in various human diseases that can possibly be prevented by antioxidants. There are many but rather expensive methods to determine total antioxidant capacity of human plasma (for endogenous antioxidant levels) or plant extracts/natural compounds (for antioxidant potential in terms of radical inhibiting or scavenging properties). We describe a simple, fast and economical 'crocin assay' using the Indian spice saffron. METHODS: In crocin assay, the extent of bleaching of crocin, a carotenoid from saffron, by peroxyl radicals generated by thermal decomposition of azo-initiator was measured. We examined its applicability to clinical samples and plant extracts. RESULTS: The cost of Indian saffron is almost 38 times less per unit dry weight compared to the 'Sigma' saffron. Yet, it gives 26 times better yield of crocin than that from 'Sigma' saffron. It was also shown that Indian saffron is rich in crocin. The total antioxidant capacity (TAC) values of human plasma from normal, healthy individuals, using Sigma as well as Indian crocin, expressed in terms of 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox) equivalent antioxidant capacity (TEAC), were comparable. We have also demonstrated that crocin assay can be used for clinical samples such as plasmas from healthy and diabetic individuals. The antioxidant potentials, TEAC, of plant extracts and pure natural compounds by Indian and Sigma crocin assays were similar. Addition of uric acid to plasma induced a concentration-dependent response. The assay was compared to standard radical scavenging 1,1'-diphenyl-2 picrylhydrazyl (DPPH) assay and was found to match well, showing better sensitivity and hence validates this assay for natural compounds and clinical samples. CONCLUSIONS: Development of crocin assay using the Indian saffron is economical and sensitive method for measurement of total antioxidant capacities from human plasma as well as natural compounds and plant extracts. PMID- 15653111 TI - A new enzymatic cycling method for ammonia assay using NAD synthetase. AB - BACKGROUND: Ammonia is an important marker for liver diseases such as hepatitis and hepatic cirrhosis. Several methods have been developed for ammonia analysis. In particular, the enzymatic assay using glutamate dehydrogenase has been widely used. However, this method is not necessarily high in sensitivity and accuracy due to inhibition by interferences in plasma and instability over long-term storage. METHODS: We developed an ammonia assay using a system consisting of three enzymes, NAD synthetase (NADS; EC 6.3.1.5), glucose dehydrogenase (GlcDH; EC 1.1.1.47), and diaphorase (DI; EC 1.6.99.2). RESULTS: The calibration curve for ammonia with the cycling method was linear (r=0.999) up to 300 micromol/l. The within-run CVs of 10 and 20 micromol/l NH4Cl solutions and 24.1 micromol/l ammonia in human plasma were 2.3%, 1.5%, and 2.8%, respectively. The between-run CVs of them were 4.5%, 3.1%, and 2.8%, respectively. The recovery was between 96.3% and 105%, and the limit of detection was 2.4 micromol/l. No significant interference was observed with addition of the following components: hemoglobin, bilirubin, chyle, EDTA, heparin, and sodium citrate. Due to the high degree of specificity of NAD synthetase to ammonia, no amino compounds exhibited any effect on the ammonia assay. A high correlation was obtained between results of the present method (y) and a conventional glutamate dehydrogenase method in regression analysis; y=0.944x-6.160 with r=0.993 (n=125). However, an addition error was observed from Bland-Altman analysis (the 95% limits of agreement between the two methods; 9.51+/-5.92 micromol/l). CONCLUSION: This new enzymatic method is more sensitive, precise, and accurate than the conventional method. In particular, accurate assay for ammonia can be performed without interference in the presence of various compounds. PMID- 15653112 TI - Effect of pituitary surgery in patients with acromegaly on adiponectin serum concentrations and alanine aminotransferase activity. AB - BACKGROUND: Thiazolidinediones increase adiponectin concentrations, improve insulin sensitivity and fatty liver disease (reflected by decreased alanine aminotransferase [ALT] activity) in type 2 diabetes. This study was performed to test the effect of neurosurgery in acromegaly (sharing at baseline insulin resistance but not increased visceral fat with type 2 diabetes) on insulin sensitivity, adiponectin concentrations and ALT activity. METHODS: Sixteen patients with acromegaly undergoing pituitary surgery (and 16 patients with type 2 diabetes treated with pioglitazone) were included. Insulin sensitivity, adiponectin concentrations and ALT activity were determined at baseline and after 4 months. RESULTS: Pituitary surgery in acromegalic patients increased adiponectin concentrations from mean (+/-S.D.) 9.3+/-3.8 to 10.2+/-4.4 mg/L (p<0.05). HOMA scores fell from 6.8+/-4 at baseline to 3.5+/-0.9 following neurosurgery (p<0.005) and ALT activity decreased from median (range) 21 (13-30) to 13 (10-42) U/L (p<0.05). In type 2 diabetics, pioglitazone treatment increased adiponectin concentrations; HOMA scores and ALT activity fell significantly. CONCLUSION: Pituitary surgery in patients with acromegaly led to a marked increase in insulin sensitivity and a slight increase in adiponectin serum concentrations, whereas ALT activity significantly decreased. PMID- 15653113 TI - Beta-thalassemia major and its effect on amino acid metabolism and growth in patients in the United Arab Emirates. AB - BACKGROUND: There may be a marked reduction in essential amino acids in the serum of children with thalassemia major and this is related to decreased growth in affected children. METHODS: One hundred patients with beta-thalassemia and 50 control children selected from among those who had presented with minor disorders unrelated to hematological disease were recruited. Urine and heparinized blood were collected from fasting thalassemic patients. After deproteinization and dilution, amino acid concentrations were measured using ion-exchange chromatography. RESULTS: Isoleucine (p<0.0001), phenylalanine (p<0.05), tyrosine (p<0.0001), taurine (p<0.0001) and glutamine (p<0.01) were significantly decreased in the plasma of thalassemic patients compared to the control group. Whereas glutamate (p<0.0001), serine (p<0.05) and proline (p<0.05) were significantly higher in thalassemic patients, threonine, glycine, alanine, valine, methionine, leucine, ornithine, lysine, histidine and arginine values were not different. The essential amino acids taurine (p<0.0001), methionine (p<0.01), valine (p<0.01), phenylalanine (p<0.01) and leucine (p<0.05) were significantly decreased in urine of thalassemic patients vs. controls, but threonine and ornithine were not different. The mean urinary excretion rate of beta-aminoisobutyric acid was not different (69+/-96 in thalassemics vs. 41+/-52 in controls). However, most plasma and urinary essential amino acids were found to be lower in thalassemics. Thalassemic patients were also found to be significantly growth impaired for age, both in height and weight compared to controls. CONCLUSION: Lower plasma values of essential amino acids and a decrease in urinary amino acids occur in thalassemic patients. Growth impairment both in height and weight also occurs in thalassemic patients compared to a control population. PMID- 15653114 TI - Dual sugar gut-permeability testing on blood drop in animal models. AB - BACKGROUND: Intestinal permeability is determined by measuring nonmetabolized sugars. In animals, intestinal permeability is determined in urine, using cumbersome and expensive metabolic cages. We developed an HPLC method for determining concentrations of lactulose (L) and L-rhamnose (R) in blood-drop of rabbits and mice, and we compared these results with the procedure based on sugars excreted in urine. We measured the intestinal permeability induced by a fragment (DeltaG) of the zonula occludens toxin which opens the paracellular pathway. METHODS: The animals received sugar solution and later received the same solution+DeltaG. Five-hour urine collection and timed blood tests were performed after ingestion of sugars. Sugars were measured with HPLC, and the percentage of recovered sugars was expressed as L/R ratio. RESULTS: At 60 min after administration of sugars, the mean L/R ratio for rabbits and mice was 0.026 and 0.052, respectively. At 60 min after administration of sugars+DeltaG, the mean L/R ratio for rabbits and mice was 0.22 and 0.53. The mean L/R ratio in the urine was 0.023 at basal condition and 0.25 after DeltaG ingestion. CONCLUSIONS: Testing small serum samples for sugar permeability is effective for monitoring changes in permeability of the gut in animals. This cheap simple method allows us to measure in vivo the biological activity of other molecules which modulate the paracellular pathway. PMID- 15653115 TI - Present status of serum lipid levels in Beijing professional populations and its trend of changes over 15 years--a collaborative study of seven research and clinical laboratories in Beijing. AB - BACKGROUND: China's economy has been growing rapidly since 1980, which may have affected blood lipid levels. We carried out a study on serum lipid levels and prevalence of lipid abnormalities in Beijing professional populations in 2001 2002 and assessed the changing trends of lipid levels by comparing the results with that of a similar study in 1984-1986. METHODS: The study population included 31,068 government employees, medical and educational workers and scientific research personnel (male/female 6:4). All participants had physical examination and blood chemistry tests. Lipid parameters analyzed included total cholesterol, low- and high-density lipoprotein (LDL and HDL) cholesterol and triglyceride. RESULTS: Total cholesterol, LDL cholesterol and triglyceride concentrations increased significantly as compared with the 1984-1986 study, but the variations of lipid levels with age and sex remained unchanged. Age-adjusted prevalence of dyslipidemia and its distribution in different sexes and age groups were statistically analyzed. Comparing the results with the data of the US in the 1990s, total cholesterol concentration was lower by 16 mg/dl in men and 18 mg/dl in women, whereas LDL cholesterol concentration was lower by 20 mg/dl in men and 15 mg/dl in women. HDL cholesterol was significantly higher than the US in both genders. CONCLUSIONS: The mean levels of total cholesterol (LDL cholesterol ) increased rapidly in the 1980s, stabilized and descended slightly in 1990s. Coronary lipid risk level in Beijing professional populations is significantly lower than in the US. PMID- 15653116 TI - Excessive matrix metalloproteinase-9 in the plasma of community-acquired pneumonia. AB - BACKGROUND: It has been shown that matrix metalloproteinase-9 (MMP-9) is involved in the pathogenesis of various pulmonary inflammatory diseases. We determined the MMP-9 concentration in the plasma of community-acquired pneumonia (CAP) patients before and after antibiotic treatment. METHODS: Gelatin zymography and ELISA analysis were used to measure MMP-9 activity and MMP-9 level, respectively, in 35 control subjects and 46 CAP patients. RESULTS: WBC counts, neutrophils, MMP-9 activity and MMP-9 level were significantly higher in CAP patients compared with that of control subjects (P<0.001), while MMP-9 activity and MMP-9 level were returned to normal after the antibiotic treatment (P<0.001). In addition, MMP-9 level correlated positively with WBC counts and neutrophils number both before and after the antibiotic treatment. CONCLUSIONS: MMP-9 may play an important role in the pathogenesis of CAP with a positive correlation with the number of neutrophils. PMID- 15653117 TI - Effects of xuezhikang, an extract of cholestin, on lipid profile and C-reactive protein: a short-term time course study in patients with stable angina. AB - BACKGROUND: Reduction of cholesterol and inflammation can be achieved by administration of a statin. Xuezhikang, an extract of cholestin, available from Chinese red yeast rice, could effectively modify the lipid profile. However, limited information is available regarding rapid effects of Xuezhikang on plasma C-reactive protein (CRP) and the lipid profile in patients with stable angina. We evaluated the short-term time course effects of lipid profile and CRP by Xuezhikang in patients with stable angina. METHODS: Forty-eight consecutive patients with stable angina were randomly assigned to 1200 or 2400 mg/day of Xuezhikang. Blood samples were drawn at days 0, 1, 7 and 14 for lipid profile and CRP levels in all patients, and hepatic enzymes were also evaluated at days 0 and 14. RESULTS: Both doses of Xuezhikang induced significant reductions in median CRP levels and in mean CRP levels at day 1 (13.0% with 1200 and 16.6% with 2400 mg/day; 14.7% with 1200 and 18.4% with 2400 mg/day), and at day 7 (18.3% with 1200 and 20.2% with 2400 mg/day; 18.5% with 1200 and 22.6% with 2400 mg/day) as well as at day 14 (28.6% with 1200 and 30.4% with 2400 mg/day; 21.7% with 1200 and 24.8% with 2400 mg/day) compared with baseline without a dose-dependent effect but a time-dependent manner. In addition, no changes were found at days 1 and 7 regarding lipid profile. However, both doses of Xuezhikang induced significant reductions in total cholesterol (TC, 13% and 22%), and low-density lipoprotein (LDL) cholesterol (23% and 32%) compared with baseline at day 14. The higher dose of Xuezhikang (2400 mg/day) resulted in significantly greater reductions in TC and LDL cholesterol compared with 1200 mg/day group (p<0.05, p<0.01, respectively). A less significant reduction was observed in triglycerides (TG) level (13% and 23%) compared with TC and LDL cholesterol. There was no significant difference in mean high-density lipoprotein (HDL) cholesterol levels compared with baseline in both groups. CONCLUSIONS: Xuezhikang resulted in rapid reduction of CRP within 24 h and lipid profile within 2 weeks, which may be clinically important for patients with coronary artery disease. PMID- 15653118 TI - High mobility group box 1 (HMGB1) protein: possible amplification signal in the pathogenesis of falciparum malaria. AB - High mobility group box 1 (HMGB1) protein, a DNA-binding protein that can also act as a pro-inflammatory cytokine if released from cells, is an important amplification signal in various forms of inflammation. The concentration of HMGB1 in serum taken at admission was increased in falciparum malaria in sixteen African children, more so in fatal cases than in those who subsequently recovered (P<0.001). Serum from both non-fatal (P=0.0048) and fatal (P<0.001) cases contained significantly more circulating HMGB1 than did serum from healthy Caucasian adults. These data provide an additional argument that malaria is fundamentally a systemic inflammatory state. In keeping with its developing role in sepsis, HMGB1 may enhance and prolong the inflammatory processes, and thus illness, in malaria. PMID- 15653119 TI - An index of community ocular Chlamydia trachomatis load for control of trachoma. AB - Quantitative PCR (Q-PCR) technology has recently been applied to the measurement of ocular loads of Chlamydia trachomatis. We present an index called the community ocular C. trachomatis load (COCTL) which is similar to the community microfilarial load (CMFL) of onchocerciasis. Our index has the advantage of being scale-independent so that, for example, percentage changes are the same whether calculated per eye swab or per Q-PCR capillary. The COCTL for a population or subgroup is formed by adding the arbitrary concentration of 1 organism per ml to each individual Q-PCR quantification, calculating the geometric mean, and finally subtracting 1 per ml again. The use of the COCTL is illustrated in a study of trachoma in northern Tanzania. The COCTL is higher in people with clinical trachoma than those without (5.8 organisms per swab vs. 0.1), and in children aged six months to ten years than in the overall population (1.1 vs. 0.4). The COCTL index is potentially useful for sentinel sites, operational research and calibration of clinical measures of trachoma. PMID- 15653120 TI - Spatial and temporal variation in biting rates and parasite transmission potentials of onchocerciasis vectors in Ecuador. AB - The influence of spatial and temporal factors on onchocerciasis transmission by Simulium exiguum s.l. and S. quadrivittatum in Ecuador was investigated to help develop sampling protocols for entomological surveillance of ivermectin programmes. Flies were collected in alternate months (November 1995-November 1996) at four sites each in the hyperendemic communities of San Miguel and El Tigre. A fixed-effects analysis of variance was used to explore the influence on vector abundance of locality, site, month and hour. Infectivity rates detected by dissection and PCR assays were compared. Simulium exiguum s.l. predominated at El Tigre (75%) whereas S. quadrivittatum prevailed at San Miguel (62%). Vector abundance was highest on river banks and outside houses. Biting and infection rates peaked from March to July. Hourly activity patterns were bimodal in S. exiguum but unimodal in S. quadrivittatum. Annual transmission potentials (ATP) for both species combined were 385 and 733 third stage larvae/person in San Miguel and El Tigre respectively, with S. exiguum accounting for 80% of the combined ATP at both localities. We recommend protocols that may maximize detection of parasite transmission. Infection rates thus obtained must be linked with vector density estimates to assess meaningfully host exposure as treatment progresses. PMID- 15653121 TI - Distribution of Buruli ulcer lesions over body surface area in a large case series in Ghana: uncovering clues for mode of transmission. AB - We studied hospital records of 750 consecutive Buruli ulcer patients in a highly endemic area in Amansie West, Ghana. Although more Buruli ulcer lesions were found on the right side of the body, comparison of lesions on arms and legs showed a bilaterally symmetrical distribution. Upper and lower extremities were affected equally by Buruli ulcers, if correction was made for differences in body surface area. Patients from outside the Amansie West district presented significantly more often with ulcerated lesions, which were more often located on a joint, than patients who lived in Amansie West, suggesting that longer travel distance might have caused delay. Our observations of a bilaterally symmetrical distribution of lesions on extremities and equal upper and lower extremity involvement are compatible with a mode of transmission that involves passive exposure of exposed body parts. An asymmetrical distribution of lesions was found in an earlier study, suggesting transmission by vegetation near the ground, through activities like farming or play. Perhaps, transmission in or near water, e.g. by bites of infected aquatic insects, might favour the pattern of distribution of lesions that we found. PMID- 15653122 TI - Isolates of Pneumocystis jirovecii from Harare show high genotypic similarity to isolates from London at the superoxide dismutase locus. AB - Pneumocystis jirovecii is the cause of Pneumocystis pneumonia (PCP) in humans. Isolates of P. jirovecii obtained from patients in Harare, Zimbabwe were genotyped at the superoxide dismutase locus. High genotypic similarity to isolates of P. jirovecii obtained from patients in London, UK was observed. These data provide additional support for the hypothesis that P. jirovecii is genetically indistinguishable in isolates from geographically diverse locations. PMID- 15653123 TI - Campylobacter species and drug resistance in a north Indian rural community. AB - Campylobacter species are known as agents of enteritis worldwide. However, rural community-based studies on Campylobacter infections are lacking. We carried out a prospective case-control study from July to December 2002 to investigate the prevalence and socio-epidemiological determinants of Campylobacter infection in a rural community in north India and the drug resistance of Campylobacter strains isolated from the community. Stool specimens from 348 subjects with diarrhoea and 351 age- and gender-matched asymptomatic controls were cultured for Campylobacter, Salmonella and Shigella. All Campylobacter strains were identified and tested for antibiotic susceptibility. Campylobacter species were isolated from 47 (13.5%) subjects with diarrhoea and 2 (0.6%) asymptomatic controls respectively (P<0.001). Campylobacter infection was significantly higher in children aged less than 5 years, families engaged in agriculture and persons who did not wash their hands with soap after peri-anal washing following defaecation. Campylobacter infections were more frequent than combined Salmonella and Shigella infections (47/348 vs. 15/351; P<0.001) in subjects with diarrhoea. Only two Campylobacter-infected individuals with diarrhoea had bloody stools. Antibiotic resistance of Campylobacter species was as follows: ampicillin 81.6%, ciprofloxacin 71.4%, tetracycline 26.5%, furazolidine 14.3%, gentamicin 10.2% and erythromycin 6.1%; 30.6% of strains were multidrug resistant. Increased quinolone resistance and multidrug resistance pose major risks for treatment failure. PMID- 15653124 TI - Strongyloidiasis: a mistaken diagnosis and a fatal outcome in a patient with diarrhoea. AB - A patient who died in the UK from Strongyloides infection, which he had contracted in the West Indies, is described. The diagnosis was not suspected initially because he had not been forthcoming about his origins. The infection was more severe because the patient was also infected with the human T cell leukaemia/lymphoma virus type 1 (HTLV-1) and this may explain why the infection with Strongyloides was fatal. The features of the case are outlined to help other clinicians faced with such a patient. PMID- 15653125 TI - The household distribution of trachoma in a Tanzanian village: an application of GIS to the study of trachoma. AB - The distribution of active trachoma in Kahe Mpya, Tanzania, an endemic village of approximately 1000 people, was mapped spatially and analysed for associated risk factors and evidence of clustering. An association between distance to water source and active disease was demonstrated, although this was reduced after accounting for the lack of independence between cases in the same household. Significant clustering of active trachoma within households was demonstrated, adding support to the hypothesized importance of intra-familial transmission. The spatial distribution of trachoma was analysed using the spatial scan statistic, and evidence of clustering of active trachoma cases detected. Understanding the distribution of the disease has implications for understanding the dynamics of transmission and therefore appropriate control activities. The demonstrated spatial clustering suggests inter-familial as well as intra-familial transmission of infection may be common in this setting. The association between active trachoma and geographical information system (GIS) measured distance to water may be relevant for planning control measures. PMID- 15653126 TI - The disease profile of poverty: morbidity and mortality in northern Uganda in the context of war, population displacement and HIV/AIDS. AB - The population of Gulu District (northern Uganda) has been severely incapacitated by war, epidemics and social disruption. This study is aimed at describing disease patterns and trends in this area through a retrospective analysis of discharge records for 155205 in-patients of Lacor Hospital in the period 1992 2002. The burden of infectious diseases in childhood is overwhelming, with malaria accounting for the steepest increase in admissions. Admissions for war related injuries and malnutrition fluctuated with the intensity of the war and the severity of famine. Emerging and re-emerging infections, such as HIV/AIDS, tuberculosis and Ebola, accounted for a heavy disease burden; however, there has been a trend for admissions related to HIV/AIDS and tuberculosis to decrease since the implementation of community-based services. Vulnerable groups (infants, children and women) accounted for 79.8% of admissions. Long-term war, population displacement, the collapse of social structures and the breakdown of the health system place people at a much greater risk of persistent, emerging and re emerging infectious diseases, malnutrition and war-related injuries, shaping the 'disease profile of poverty'. Most of the disease burden results from infectious diseases of childhood, whose occurrence could be dramatically reduced by low-cost and effective preventive and curative interventions. PMID- 15653127 TI - Absence of nucleotide polymorphism in a Plasmodium vivax multidrug resistance gene after failure of mefloquine prophylaxis in French Guyana. AB - Vivax malaria is widespread and resistance has been described for chloroquine and sulfadoxine-pyrimethamine. We report on evidence of failure of mefloquine prophylaxis in a French soldier who contracted Plasmodium vivax in French Guyana, South America. Despite regular weekly mefloquine prophylaxis (250 mg/d), the patient presented with a first episode of vivax malaria, which was treated by chloroquine alone, then experienced a second crisis in France. The reappearance of the parasites occurred one day after the end of prophylaxis, confirming parasitological and clinical resistance in a non-immune patient. Mefloquine was detected by a liquid chromatography assay in plasma at a level of 1062 ng/ml, which was higher than the expected concentration after five months of weekly prophylaxis. This isolate had no single nucleotide polymorphisms of the pvmdr1 gene at seven allele positions: pvmdr1 N91, Y189, Y976, S1071, F1076, N1079 and D1291, corresponding to codons 86, 184, 939, 1034, 1039, 1042 and 1246 in P. falciparum. This observation of failure of mefloquine prophylaxis against P. vivax, when added to previously reported chloroquine and atovaquone-proguanil failure, strengthens the case for re-evaluating drug policies for vivax malaria and the need for continuous research on molecular markers of drug resistance. PMID- 15653129 TI - Advances in superporous hydrogels. AB - Superporous hydrogels (SPHs) are different from superabsorbent polymers (SAPs) in that SPHs swell fast, within minutes, to the equilibrium swollen state regardless of their size. The fast swelling property is based on water absorption through open porous structure by capillary force. The poor mechanical strength of SPHs was overcome by developing the second-generation SPH composites (SPHCs) and the third-generation SPH hybrids (SPHHs). This review examines the differences between SAPs and SPHs and describes three different generations of SPHs. PMID- 15653130 TI - Hyaluronic acid hydrogel as Nogo-66 receptor antibody delivery system for the repairing of injured rat brain: in vitro. AB - Nogo-66 and NgR are important receptors inhibiting neuronal regeneration and therefore are targets for treating CNS injury. Antagonists of this receptor including blocking antibodies are potential therapeutic agents for CNS axonal injuries such as spinal cord and brain trauma. A new antibody (IgG) releasing system has been developed by covalently attaching IgG to the biodegradable hyaluronic acid (HA) hydrogel via the hydrolytically unstable hydrazone linkage, aiming to deliver the antibody of CNS regeneration inhibitors to the injured brain. In this paper we describe the synthesis, physico-chemical characteristics and test results of biological activity of antibody released from hyluronic acid hydrogel. To form the conjugates the antibody is attached to the polymer backbone using a condensation reaction between aldehyde group of the antibody and hydrazide group of the HA hydrogel. Furthermore, pH sensitive linkage-hydrozone has been formed between hydrogel and antibody. The amount of conjugated antibodies can reach 135 microg antibody/mg hydrogel in the dry state. At low pH, the antibodies released quite fast. However, the antibodies released much slower in neutral and alkaline environment. The bioactivity of antibody released from hydrogel was retained as demonstrated by indirect immunofluorescence technique. PMID- 15653131 TI - Poly(amidoamine) dendrimers as ophthalmic vehicles for ocular delivery of pilocarpine nitrate and tropicamide. AB - The purpose of this study was to determine the influence of a controlled incremental increase in size, molecular weight and number of amine, carboxylate and hydroxyl surface groups in several series of poly(amidoamine) (PAMAM) dendrimers for controlled ocular drug delivery. The duration of residence time was evaluated after solubilization of several series of PAMAM dendrimers (generations 1.5 and 2-3.5 and 4) in buffered phosphate solutions containing 2 per thousand (w/v) of fluorescein. The New Zealand albino rabbit was used as an in vivo model for qualitative and quantitative assessment of ocular tolerance and retention time after a single application of 25 microl of dendrimer solution to the eye. The same model was also used to determine the prolonged miotic or mydriatic activities of dendrimer solutions, some containing pilocarpine nitrate and some tropicamide, respectively. Residence time was longer for the solutions containing dendrimers with carboxylic and hydroxyl surface groups. No prolongation of remanence time was observed when dendrimer concentration (0.25 2%) increased. The remanence time of PAMAM dendrimer solutions on the cornea showed size and molecular weight dependency. This study allowed novel macromolecular carriers to be designed with prolonged drug residence time for the ophthalmic route. PMID- 15653132 TI - Release of indomethacin from ultrasound dry granules containing lactose-based excipients. AB - Physical mixtures were prepared containing indomethacin and beta-lactose and alpha-lactose-based excipients (Ludipress and Cellactose). The mixtures were compacted with the aid of ultrasound, obtaining tablets, which were milled and sieved. Granules thus obtained were examined by optical microscopy and differential scanning calorimetry. The intense yellow color of the granules and the absence of indomethacin peak in thermograms suggest important modifications of indomethacin physical state; the drug thus modified appears to be spread on the excipient particle surface as a thin film, giving a lustrous appearance. No influence of ultrasound was observed on phase transition concerning lactose; only loss of water was important under high energy ultrasound. Dissolution profiles suggest an increased release of the drug from the systems treated with ultrasound at high energy, with respect to a traditional compaction; while no difference could be evidenced among the three excipients that, however, appear all suitable for this ultrasound-aided direct compression process. PMID- 15653133 TI - Application of Eudragit RS to thermo-sensitive drug delivery systems: II. Effect of temperature on drug permeability through membrane consisting of Eudragit RS/PEG 400 blend polymers. AB - The Eudragit RS and polyethylene glycol 400 (PEG 400) blend polymer (EPG) membranes were prepared by the solvent casting method to pioneer a novel application of Eudragit RS to a thermo-sensitive material. The EPG membranes containing 2.5-10% PEG 400 (2.5-10% EPG) showed the glass transition temperatures (Tgs) around the body temperature (32-42 degrees C). Drug permeation studies through the EPG membranes were carried out using acetaminophen (AAP) and aminopyrine (AMP) as the model drugs. The permeability of AAP and AMP through the EPG membranes has been shown to be a discontinuous function of temperature, that is, their permeability increased steeply above the Tg of the membranes. The amount of AMP permeated at 42 degrees C was nearly eight times as much as that at 36 degrees C. Arrhenius plots of the steady-state permeability coefficient (P) of AAP indicated two straight lines that intersect at the Tg of the 10% EPG membrane. In the water uptake study for the 10% EPG membrane, the degree of the swelling for the membrane tended to increase with increasing temperature above the Tg of the membrane. The thermo-sensitive permeation mechanism for the EPG membranes might be based on the structure change of the membranes caused by the glass transition, so that the membranes could absorb more water. Considering the high biological safety of Eudragit RS and PEG 400, the EPG membranes might be used to develop a novel thermo-sensitive drug delivery system. PMID- 15653134 TI - Effect of L-menthol and 1,8-cineole on phase behavior and molecular organization of SC lipids and skin permeation of zidovudine. AB - The aim of this investigation was to study the effect of 1,8-cineole and L menthol on phase behavior and molecular organization of Stratum corneum (SC) lipids and permeation of zidovudine (AZT) across human cadaver skin (HCS). Permeation studies were conducted across HCS using Franz diffusion cells at 37 degrees C. Differential scanning calorimetry (DSC) and attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) were employed to understand the effect of terpenes on phase behavior and molecular organization of a model SC lipid system consisting of an equimolar mixture of ceramide, palmitic acid and cholesterol. Both 1,8-cineole and L-menthol applied at 5% w/v in 66.6% ethanol as a vehicle significantly enhanced the pseudosteady state flux of AZT across HCS. The vehicle reduced the number of endothermic transitions observed in the DSC thermogram of a hydrated model SC lipid system from three to two with a lowered midtransition temperature (Tm), while the inclusion of terpenes resulted in a single but very broad endothermic transition for the model SC lipid system. Correspondingly, ATR-FTIR studies revealed that both 1,8-cineole and L-menthol increased CH2 stretching frequencies on either side of lipid transition in addition to lowering the Tm of model SC lipid system by approximately 2-8 degrees C. The alterations observed in the amide-I frequencies of model SC lipid system after the inclusion of terpenes suggest that they disrupt the interlamellar hydrogen-bonding network at the polar head group region. Further, terpenes also increased the hydration levels of the lipid system probably by forming new aqueous channels. These results indicate that terpenes enhance transdermal permeation of AZT and other drugs by transforming SC lipids from a highly ordered orthorhombic perpendicular subcellular packing to a less ordered hexagonal subcell packing. PMID- 15653135 TI - How cyclodextrin incorporation affects the properties of protein-loaded PLGA based microspheres: the case of insulin/hydroxypropyl-beta-cyclodextrin system. AB - The aim of this work was to study the influence of cyclodextrin (CD) incorporation on the properties of protein-loaded poly(lactide-co-glycolide) (PLGA) microspheres, with particular regards to protein release kinetics. To this purpose, insulin-loaded microspheres were prepared by spray-drying emulsion or solution formulations, with or without hydroxypropyl-beta-cyclodextrin (HPbetaCD), and fully characterized for encapsulation efficiency and release kinetics of both insulin and cyclodextrin. Homogeneous populations of spherical microparticles entrapping both insulin and HPbetaCD were obtained. In order to get an insight into insulin/HPbetaCD interactions occurring inside microspheres, Fourier transform infrared (FTIR) analysis in the Amide I region was performed. FTIR spectra of dried microspheres containing HPbetaCD showed a change in insulin secondary structure, attributed to the presence of insulin/HPbetaCD complexes within microspheres. Insulin release was affected by the presence of HPbetaCD depending on the initial formulation conditions. In the case of microspheres prepared from emulsion, cyclodextrin reduced only insulin burst, whereas in the case of microspheres obtained from solution, the overall insulin release rate was slowed down. Combining the release kinetics of HPbetaCD with the FTIR results on hydrated microspheres, it was concluded that the formation of insulin/HPbetaCD complexes inside microspheres is critical to decrease protein diffusivity in the polymer matrix and achieve an effective modulation of protein release rate. PMID- 15653136 TI - Enhancement of T helper type 1 immune responses against hepatitis B virus core antigen by PLGA nanoparticle vaccine delivery. AB - Currently, there is a need for therapeutic vaccines that are effective in inducing robust T helper type 1 (Th1) immune responses capable of mediating viral clearance in chronic hepatitis B infection. Hepatitis B therapeutic vaccines were designed and formulated by loading the hepatitis B core antigen (HBcAg) into poly(D,L-lactic-acid-co-glycolic acid) (PLGA) nanoparticles with or without monophospholipid A (MPLA), a Th1-favoring immunomodulator. These particles were around 300 nm in diameter, spherical in shape and had approximately 50% HBcAg encapsulation efficiency. A single immunization with a vaccine formulation containing (MPLA+HBcAg) coformulated in PLGA nanoparticles induced a stronger Th1 cellular immune response with a predominant interferon-gamma (IFN-gamma) profile than those induced by HBcAg alone, free (HBcAg+MPLA) simple mixture or HBcAg loaded nanoparticles in a murine model. More importantly, the level of HBcAg specific IFN-gamma production could be increased further significantly by a booster immunization with the (HBcAg+MPLA)-loaded nanoparticles. In summary, these results demonstrated that codelivery of HBcAg and MPLA in PLGA nanoparticles promoted HBcAg-specific Th1 immune responses with IFN-gamma production. These findings suggest that appropriate design of the vaccine formulation and careful planning of the immunization schedule are important in the successful development of effective HBV therapeutic vaccines. PMID- 15653137 TI - A cell encapsulation device for studying soluble factor release from cells transplanted in the rat brain. AB - The transplantation of a variety of naturally occurring and genetically modified cell types has been shown to be an effective experimental method to achieve sustained delivery of therapeutic molecules to specific target areas in the brain. To acquire a better understanding of dosing, implant mechanism of action, and how certain cell types affect remodeling of central nervous system (CNS) tissue, a refillable cell encapsulation device was developed for introducing cells into the brain while keeping them physically isolated from contact with brain tissue with a semipermeable membrane. The stereotactically placed device consists of a hollow fiber membrane (HFM), a polyurethane grommet with watertight cap that snaps into a precisely drilled hole in the rat skull, and a removable cell-containing insert. The cell-containing insert can be introduced or removed in a time-dependent manner to study the influence of soluble factors released from transplanted cells. The study describes the device design and validates its utility using a well-established cell transplantation model of Parkinson's disease. PMID- 15653138 TI - Self-assembled particles of an elastin-like polymer as vehicles for controlled drug release. AB - Elastin-like polymers (ELPs) are a new kind of protein-based polymers showing interesting properties in the biomaterial field. This work explored the use of self-assembled poly(VPAVG) micro- and nanoparticles as vehicles for the controlled release of the model drug dexamethasone phosphate (DMP). Poly(VPAVG) has shown to form stable particles with a size below 3 mum as a water or PBS polymer solution was warmed above its transition temperature ( approximately 30 degrees C). Due to the peculiar composition of the monomer, the formation and redissolution of the self-assembled microparticles shows an interesting hysteresis behaviour by which the particles are formed at this temperature but do not redissolve until a strong undercooling of approximately 12-15 degrees C is achieved. Therefore, the particles, once formed, are stable either at room or body temperature. These self-assembled particles are able to encapsulate significant amounts of the model drug when self-assembling was carried out in a co-solution polymer-DMP. The release profiles showed a sustained DMP release for about 30 days. Being the potential of this new polymeric carrier high, further research is being conducted to functionalise the poly(VPAVG) base as a way to induce a stronger polymer-drug binding and, accordingly, a more sustained release. PMID- 15653139 TI - Tailoring width of microfabricated nanochannels to solute size can be used to control diffusion kinetics. AB - Top-down microfabrication techniques were used to create silicon-based membranes consisting of arrays of uniform channels having a width as small as 7 nm. The measurement of diffusion kinetics of solutes across these membranes under sink conditions reveals non-Fickian behavior as the nanopore width approaches the hydrodynamic diameter of the solute. Zero-order diffusion of interferon is observed at channel width of 20 nm, and the same phenomenon occurs with albumin and 13-nm-wide channels, whereas Fickian diffusion kinetics is seen at 26 nm and larger pore sizes. A prototypical drug delivery device is described that is fitted with a 13-nm nanopore membrane and loaded with radio-labeled BSA. Following subcutaneous implantation in rats, diffusion from the device provided prolonged levels of BSA in the blood. Such a nonmechanical device offers important advantages in drug delivery applications, including zero-order release and high loading capacity. PMID- 15653140 TI - In vitro and in vivo studies on mucoadhesive microspheres of amoxicillin. AB - Amoxicillin mucoadhesive microspheres (Amo-ad-ms) were prepared using ethylcellulose (Ec) as matrix and carbopol 934P as mucoadhesive polymer for the potential use of treating gastric and duodenal ulcers, which were associated with Helicobacter pylori. The morphological characteristics of the mucoadhesive microspheres were studied under scanning electron microscope. In vitro release test showed that amoxicillin released faster in pH 1.0 hydrochloric acid (HCl) than in pH 7.8 phosphate buffer. Yet, it would be degraded to some extent in a pH 1.0 HCl medium at 37 degrees C, which indicated that amoxicillin was not stable in an acidic surrounding. It was also found that amoxicillin entrapped within the microspheres could keep stable. In vitro and in vivo mucoadhesive tests showed that Amo-ad-ms adhered more strongly to gastric mucous layer than nonadhesive amoxicillin microspheres (Amo-Ec-ms) did and could retain in gastrointestinal tract for an extended period of time. Amo-ad-ms and amoxicillin powder were orally administered to rats. The amoxicillin concentration in gastric tissue was higher in the Amo-ad-ms group. In vivo H. pylori clearance tests were also carried out by administering, respectively, Amo-ad-ms or amoxicillin powder, to H. pylori infectious BALB/c mice under fed conditions at single or multiple dose(s) in oral administration. The results showed that Amo-ad-ms had a better clearance effect than amoxicillin powder did. In conclusion, the prolonged gastrointestinal residence time and enhanced amoxicillin stability resulting from the mucoadhesive microspheres of amoxicillin might make contribution to H. pylori clearance. PMID- 15653141 TI - A model for drug release from fast phase inverting injectable solutions. AB - A model is developed to describe protein release kinetics from injectable, polymer solution depots which undergo rapid phase inversion on injection. The model consists of a polymer-rich phase and a solvent-rich phase, consistent with experimentally observed phase inversion morphology. Equations in the polymer-rich phase are based on diffusion-reaction mass balances for solvent, water and dissolved drug, and the rate of dissolution of dispersed drug particles. Equations in the water-rich phase are also of the diffusion-reaction type. Transport parameters in the polymer-rich phase are coupled to the ternary thermodynamics through friction formalism, and remaining parameters are estimated from literature data, leaving two free parameters: volume fraction of water-rich phase (epsilon) and k, the mass-transfer coefficient for bath-side transfer of the protein. Variations of these parameters lead to predictions of release profiles that vary from a rapid, burst-like behavior followed by a locking-in of the polymer-rich phase, to a uniform, zero-order profile. Comparisons are made to lysozyme release data for three systems: PLGA solutions in N-methlypyrollidinone (NMP), PLA solutions in NMP, and the latter with added Pluronic. Good agreement between model predictions and data is shown; in particular, the transition from rapid release to zero-order kinetics that occurs on addition of Pluronic is illustrated. PMID- 15653142 TI - Preparation and evaluation of a chitosan salt-poloxamer 407 based matrix for buccal drug delivery. AB - The aim of this work was to prepare and evaluate a matrix for buccal drug delivery composed of a chitosan salt and poloxamer 407. Different chitosan salts were formed by reacting chitosan with acetic, citric, and lactic acid. Various proportions of poloxamer 407 were added to the aqueous solution of chitosan salt, and the residue obtained by lyophilisation was compressed into discs, using a 30 kN compression force. An experimental design (3(2)) was used to study the influence of the type of chitosan salt and of the relative amount of poloxamer on drug release capacity, swelling, erosion, and mucoadhesiveness of matrices. The results showed that matrix properties depended significantly on both relative amount of poloxamer and chitosan salt type. The rank orders of chitosan salts for the four processes evaluated were as follows: drug release: chitosan acetate>chitosan citrate>chitosan lactate; swelling: chitosan lactate>chitosan acetate=chitosan citrate; erosion: chitosan citrate>chitosan lactate>chitosan acetate; mucoadhesion: chitosan lactate>chitosan acetate=chitosan citrate. Mucoadhesion was particularly favoured when poloxamer 407 was present at about 30% (w/w). The matrix composed of chitosan lactate and poloxamer 407 showed the best characteristics for buccal administration. PMID- 15653143 TI - Encapsulation of therapeutic nucleoside hydrolase in functionalised nanocapsules. AB - Liposomes are introduced as encapsulating carrier for prodrug activating enzymes. Inosinea-adenosinea-guanosine preferring nucleoside hydrolase of Trypanosoma vivax, a potential prodrug activating enzyme, was encapsulated in porin functionalized dioleyl-phosphatidylglycerol/egg-phosphatidylglycerol (DOPC/EPG) liposomes. Reactors had radiuses in the nanometer scale. First, transport of nucleosides through general diffusion porins OmpF and PhoE was measured in swelling assays, after which fully functional nanoreactors were developed. Enzyme catalysis of p-nitrophenylriboside, a substrate analogue for nucleoside hydrolases, was significantly higher in permeabilized vesicles than in control vesicles without porins. Residual activity of control vesicles possibly resides in an interaction between the enzyme and the liposomes. This interaction was not of electrostatic nature, since it remained unaffected after the addition of high salt or after perturbation of liposome surface charge and charge density. With these vesicles, we have introduced a new strategy for prodrug therapy, combining the benefits of ADEPT and liposome targeting strategies. PMID- 15653144 TI - Intracellular delivery of liposome-encapsulated prolidase in cultured fibroblasts from prolidase-deficient patients. AB - Prolidase is a cytosolic exopeptidase whose deficiency causes the development of a rare autosomal recessive disorder known as Prolidase Deficiency (PD). The main manifestations of PD are intractable ulcerations of the skin, recurrent infections and mental retardation. At this time only a hazardous and expensive chronic therapy based on blood transfusions is the suggested treatment for PD. The aim of this work was to investigate the capability of utilizing liposomes as enzyme carriers: these vesicular systems have been recently evaluated as protein carriers for their potential in terms of "in vivo" localization, drug release and for protein stabilization in biological fluids. Liposomes were prepared, with a 1:1 PC:Col molar ratio with or without DSPE-PEG, by a thin-film hydration. Ex vivo experiments were performed, incubating prolidase loaded liposomes with cultured fibroblasts from PD patients and from controls, to determine the amount of active enzyme delivered to cells. Evaluation of liposomes toxicity on cultured skin fibroblasts showed that liposomes did not interfere with cellular growth. Results showed that all the active prolidase encapsulated in the liposomes was completely vehiculated inside fibroblasts after 6 days incubation. SEM analysis suggests that prolidase is vehiculated inside the cell through liposome endocytosis. PMID- 15653145 TI - Targeting tumor angiogenic vasculature using polymer-RGD conjugates. AB - Sites of neovascular angiogenesis are important chemotherapy targets. In this study, the synthesis, characterization, in-vivo imaging and biodistribution of a technetium-99m labeled, water-soluble, N-(2-hydroxypropyl) methacrylamide (HPMA) copolymer carrying doubly cyclized Arg-Gly-Asp motifs (HPMA copolymer-RGD4C conjugate) are reported. In vitro endothelial cell adhesion assays indicated that HPMA copolymer-RGD4C conjugates inhibited alphaVbeta3-mediated endothelial cell adhesion while HPMA copolymer Arg-Gly-Glu control conjugates (HPMA copolymer RGE4C conjugate) and hydrolyzed HPMA copolymer precursor (HPMA copolymer) showed no activity. The scintigraphic images of prostate tumor bearing SCID mice obtained 24 h post-i.v. injection indicated greater tumor localization of HPMA copolymer-RGD4C conjugate than the control, HPMA copolymer-RGE4C conjugate. The 24-h necropsy radioactivity data showed that HPMA copolymer-RGD4C conjugate had significantly higher (p<0.001) tumor localization compared to HPMA copolymer RGE4C conjugate and HPMA copolymer. Also, HPMA copolymer-RGD4C conjugates had sustained tumor retention over 72 h and reasonably efficient clearance from the background organs. These results suggest that specific tumor angiogenesis targeting is possible with HPMA copolymer-RGD4C conjugates. This construct provides a foundation that should support targeted delivery of radionuclides and drugs to solid tumors for diagnostic and therapeutic applications. PMID- 15653146 TI - Controlled and targeted tumor chemotherapy by micellar-encapsulated drug and ultrasound. AB - The results of a comprehensive in vivo study of a novel tumor-targeting modality are reported. The technique utilized in this study is based on the encapsulation of the chemotherapeutic agent within polymeric micelles in combination with a local ultrasonic irradiation of the tumor. A doxorubicin (DOX) biodistribution, a yield of the internal tumors and a growth rate of the subcutaneous (s.c.) tumors was compared for molecularly dissolved and micellar-encapsulated DOX. This was done with and without tumor sonication, using an ovarian carcinoma tumor model in nu/nu mice. Pure and mixed Pluronic P-105, PEG2000-diacylphospholipid, and poly(ethylene glycol)-co-poly(beta-benzyl-L-aspartate) micelles were used as drug carriers. DOX intracellular uptake was characterized by flow cytometry. A local ultrasonic irradiation of the tumor resulted in a substantially increased drug accumulation in the tumor cells. The effect of the ultrasound was dependent on the time between ultrasound application and drug injection. Ultrasound did not enhance micelle extravasation; the ultrasonic enhancement of drug internalization by the tumor cells required a preliminary passive drug accumulation in the tumor interstitium. Due to the ultrasound-enhanced drug intracellular uptake and cell killing, the yield of intraperitoneal (i.p.) ovarian carcinoma tumors decreased from 70% for DOX dissolved in PBS (positive control) to 36% for the same concentration of DOX encapsulated in Pluronic micelles combined with a 30-s sonication of the abdominal region of a mouse (3 mg/kg DOX, i.p. injection 1 day after inoculation, n>or=10). For s.c. tumors, micellar delivery combined with localized ultrasonic tumor irradiation resulted in a substantial decrease of the tumor growth rates compared to a positive control (3 mg/kg DOX, i.v. injections, n=7, p<0.05). Possible mechanisms of the ultrasound bioeffects on in vivo drug targeting are discussed. PMID- 15653147 TI - Incorporation of low molecular weight biocides into polystyrene-divinyl benzene beads with controlled release characteristics. AB - Triclosan and phosphonium salt biocides have been separately incorporated into polystyrene-divinylbenzene (PS-DVB) beads by suspension polymerization. Ultraviolet (UV) absorption measurements have been used to monitor the release of these low molecular weight biocides out of the PS-DVB beads immersed in water ethanol mixtures and in physiological saline. The release of the biocide agents is strongly dependent on either the DVB or/and the antimicrobial composition ratio in the beads. An increase of biocide incorporation in the PS/DVB beads was accompanied by a corresponding enhancement of its concentration in liquid mixtures. On the contrary, higher cross-linking densities hindered the biocide migration out of the beads by diminishing its release rate into either the aqueous ethanol solutions or the natural serum. Moreover, Fourier transform Raman (FT-Raman) spectra and Attenuated Total Reflectance Infrared (ATR-FTIR) measurements of the PS-DVB-Triclosan and PS-DVB-phosphonium salt beads, before and after their immersion in water-ethanol solutions, gave a similar qualitative evidence of the biocide release. PMID- 15653148 TI - Interactions of carboplatin with fibrin(ogen), implications for local slow release chemotherapy. AB - The effect of carboplatin (CPt) on fibrin(ogen) clot formation and the possible use of this combination for local slow release chemotherapy were examined. CPt significantly reduced thrombin-induced fibrin clotting time (CT) and increased clot turbidity in a concentration-dependent manner. When CPt was mixed with physiological levels of fibrinogen (>1 mg/ml), electron-dense nanoparticles (3 nm) were formed, as demonstrated by both optical particle counter and transmission electron microscopy (TEM). Upon thrombin-induced coagulation, the CPt nanoparticles were trapped within the fibrin mesh. At higher fibrinogen levels (>5 mg/ml), the 3-nm CPt nanoparticles aggregated, so that approximately 2% and approximately 0.5% of the CPt on the fibrinogen appeared as larger particles of 10 and 50 nm, respectively. Dialysis experiments showed that 60-70% of the CPt was released from the fibrin clot within one hour as a non-particulate soluble form, while approximately 30% of particulate CPt were retained. Up to 5 mg/ml this portion of firmly attached CPt was dependent of the initial drug level. CPt released from the fibrin by either diffusion or by fibrinolysis exhibited cytotoxic activity towards retinoblastoma (RB) cell lines (Y-79 and Weri RB1) equivalent to free drug. Our study indicates that CPt enhances fibrin clot formation and suggests the use of fibrin with high dose CPt for slow release chemotherapy against localized tumors such as retinoblastoma. PMID- 15653149 TI - Cationic TAT peptide transduction domain enters cells by macropinocytosis. AB - Naturally occurring and synthetic short arginine containing protein transduction domains (PTDs), including HIV1 TAT, poly-Arg and Antp, have been used to deliver a wide variety of macromolecular, biologically active therapeutic cargo into cells, including peptides, proteins, antisense oligonucleotides and liposomes, in vitro and to treat pre-clinical models of cancer and stroke. PTDs enter cells in a rapid, receptor-independent fashion. Recently, large TAT-fusion proteins (in excess of 30,000 Da) were shown to transduce into cells by fluid-phase macropinocytosis, a specialized form of endocytosis that is independent of caveolae, clathrin and dynamin. However, it remains controversial as to whether or not PTD peptides (1000-5000 Da) enter cells via macropinocytosis and/or through an unknown alternative mechanism. Due to strong ionic interactions with the cell surface, previous measurements of PTD peptide internalization were inaccurate. Cationic PTD peptides containing variable numbers of arginine residues and conditions entered cells exclusively through macropinocytosis. In addition, no PTD peptide was found to enter cells at 4 degrees C, a long held assumption of transduction. Taken together, these observations provide a solid scientific basis for the development of novel biologically active transducible anticancer PTD peptide therapeutics. PMID- 15653150 TI - Cyclodextrin formulation of dorzolamide and its distribution in the eye after topical administration. AB - Due to limited aqueous solubility of dorzolamide at physiologic pH, the pH of Trusopt eye drops (cont. 2% dorzolamide) has to be kept at about 5.65, and to increase the topical bioavailability of the drug from Trusopt the contact time of the drug with the eye surface is increased by increasing the viscosity of the eye drops to 100 cps. This low pH and high viscosity can lead to local irritation. In this study, dorzolamide hydrochloride was formulated as 2% and 4% low viscosity solutions (viscosity 3 to 5 cps) containing randomly methylated beta-cyclodextrin at pH 7.45. These formulations were evaluated in rabbits. The animals were sacrificed at various time points after topical administration of the drug and the dorzolamide concentration determined in the different parts of the eye. Trusopt was used as a reference standard. The topical availability of dorzolamide from the cyclodextrin-containing eye drops appeared to be comparable to that from Trusopt and the drug reached retina and optic nerve to give measurable concentrations for at least 8 h after administration of the eye drops. PMID- 15653151 TI - Non-viral gene delivery for local and controlled DNA release. AB - Non-viral DNA delivery systems show important advantages vs. viral systems that are usually associated with an immunological response and safety risks. In this study, disulfide cross-linked peptide-DNA condensates were investigated for local gene delivery. Two different 21 amino acid peptides were designed to have a DNA binding sequence in combination with a transglutaminase substrate site or a nuclear localization site. The peptides were used in different ratios to each other to form stable cross-linked DNA-peptide condensates with a mean diameter of 164 nm and a size distribution from 43 to 204 nm. Such aggregates showed similar stability compared to condensates formed between DNA and high molecular weight poly-L-lysine (PLL). Peptide-DNA condensates were covalently immobilized into fibrin matrices by the activity of factor XIII and were used for gene delivery in vitro. After internalization, reduction of the cross-linked peptide-DNA condensates yielded increased transfection efficiencies into different cell types cultured in 2D sandwich assays, and comparable values for HUVECs cultured in a 3D fibrin matrix, as compared to PLL-DNA condensates. Cell viability 24 h after transfection remained above 95%. The target was to develop a transfection system based on small peptides that can be covalently cross-linked into fibrin-matrices where DNA-release takes place upon cellular degradation of the matrix. This approach provides an interesting tool in non-viral gene delivery. PMID- 15653152 TI - Copolymers of poly-L-lysine with serine and tryptophan form stable DNA vectors: implications for receptor-mediated gene transfer. AB - Inefficient gene transfer and poor stability in physiological medium are important shortcomings for receptor-mediated gene transfer vectors. Here, we evaluate vectors formulated with random copolymers of L-lysine/L-serine (3:1) and L-lysine/L-tryptophan (4:1), focusing on both their biophysical and functional characterization. By means of dynamic light scattering (DLS) and transmission electron microscopy (TEM), we demonstrate that poly-L-lysine (pK), poly-L-lysine L-tryptophan (pKW) and poly-L-lysine-L-serine (pKS) are able to form compacted, small particles when mixed with plasmid DNA in the absence of salt. Upon dilution in physiological medium, copolymers of both lys/ser and lys/trp do not aggregate, in contrast with poly-L-lysine DNA complexes as determined by scattering, DLS and TEM measurements. Tight packing, as demonstrated by resistance to heparin, SDS and trypsin treatments, is also featured in tryptophan-containing complexes. Successful receptor-mediated endocytosis gene transfer using galactosylated copolymers into cells expressing the asiagloglycoprotein receptor correlated with lack of aggregation. Particles obtained using galactosylated poly-L-lysine-L tryptophan (Gal-pKW) copolymer demonstrated specific receptor-mediated gene transfer since reporter gene activity dropped in the presence of an excess ligand in the culture medium during transfection. Although copolymers of galactosylated poly-L-lysine-L-serine (Gal-pKS) do not aggregate in the presence of salt, they are not able to internalize in a specific receptor-mediated endocytosis fashion. The introduction of bulky aromatic/hydrophobic (tryptophan) or hydrophillic (serine) moieties into the positively charged vectors allows the compacted particles to disperse into salt-containing medium avoiding salt-induced aggregation. Moreover, tryptophan-containing particles are able to mediate specific gene transfer via receptor-mediated endocytosis. PMID- 15653153 TI - Influence of TAT-peptide polymerization on properties and transfection activity of TAT/DNA polyplexes. AB - Use of bioactive cationic peptides as gene carriers is limited by instability of their DNA complexes in vivo and by the loss of their biological activity due to undesired interactions of their bioactive parts with the DNA. To overcome the two major limitations, biodegradable high-molecular-weight form of TAT peptide (POLYTAT) sensitive to cellular redox-potential gradients was synthesized in this study by oxidative polycondensation. Physicochemical and transfection properties of DNA polyplexes based on POLYTAT were investigated and compared with polyplexes based on TAT polymer prepared by in situ template-assisted polymerization. Physicochemical properties of TAT-based polyplexes were affected by the molecular weight and method of polymerization of the TAT peptide. All TAT-based DNA polyplexes exhibited reduced cytotoxicity when compared with control polyethylenimine (PEI) polyplexes. Polyplexes based on both high-molecular-weight TAT polypeptides exhibited increased transfection efficiency compared to control TAT peptide but lower than that of PEI polyplexes. The evidence shows that transfection activity of TAT-based polyplexes is strongly dependent on the presence of chloroquine and therefore suggests that TAT polyplexes are internalized by an endocytosis. Overall, high-molecular-weight reducible polycations based on bioactive peptides has the potential as versatile carriers of nucleic acids that display low cytotoxicity and can prove to be especially beneficial in cases that require surface presentation of membrane-active or cell specific targeting peptides. PMID- 15653154 TI - Microencapsulation by solvent extraction/evaporation: reviewing the state of the art of microsphere preparation process technology. AB - The therapeutic benefit of microencapsulated drugs and vaccines brought forth the need to prepare such particles in larger quantities and in sufficient quality suitable for clinical trials and commercialisation. Very commonly, microencapsulation processes are based on the principle of so-called "solvent extraction/evaporation". While initial lab-scale experiments are frequently performed in simple beaker/stirrer setups, clinical trials and market introduction require more sophisticated technologies, allowing for economic, robust, well-controllable and aseptic production of microspheres. To this aim, various technologies have been examined for microsphere preparation, among them are static mixing, extrusion through needles, membranes and microfabricated microchannel devices, dripping using electrostatic forces and ultrasonic jet excitation. This article reviews the current state of the art in solvent extraction/evaporation-based microencapsulation technologies. Its focus is on process-related aspects, as described in the scientific and patent literature. Our findings will be outlined according to the four major substeps of microsphere preparation by solvent extraction/evaporation, namely, (i) incorporation of the bioactive compound, (ii) formation of the microdroplets, (iii) solvent removal and (iv) harvesting and drying the particles. Both, well-established and more advanced technologies will be reviewed. PMID- 15653155 TI - Controlled release from bioerodible polymers: effect of drug type and polymer composition. AB - The effect of the chemical nature of the drug on matrix degradation and drug release behavior of degradable polymers was studied, using lidocaine as a model drug in base and salt forms. We show in this study that the drug in the base form has a substantial effect on the release characteristics, through an accelerating effect on matrix degradation. Study of drug release from PdlLGA shows that lidocaine salt follows a three-phase release pattern, in contrast to the biphasic release of the lidobase. However, PlLA shows a different drug release pattern, with only a single diffusion phase exhibited for both lidobase and lidosalt. We also demonstrate that the crystallinity of matrix plays an important role on drug release profiles: a crystalline matrix (PlLA IV=2.04) releases the drug at a much slower rate compared to its amorphous counterpart of similar molecular weight (PdlLA IV=2.4). The details of the study of different factors influencing the drug release may have important implications for the control of delivery of potent drugs in various therapeutic windows. PMID- 15653156 TI - Gentamicin-loaded discs and microspheres and their modifications: characterization and in vitro release. AB - Osteomyelitis is an infection of the bone, and successful treatment involves local administration for about 6 weeks. Gentamicin is a very hydrophilic drug and tends to come out into the water phase when microspheres are fabricated using solvent evaporation method. Hence, spray drying is an option, and it was observed that the release rate tends to be fast when the particle size is small and large particles cannot be prepared by spray drying. In an effort to get better encapsulation efficiency and release rate, we have worked on the possibility of compressing the microspheres into discs and modifying the porosity of the discs by using biocompatible materials like polyethylene glycol (PEG) and calcium phosphates and also on the fabrication of double-walled and composite microspheres. In the case of microspheres, two methods of fabrication both based on solvent evaporation method were employed. The two polymers used are poly-L lactide (PLLA) and copolymers of poly-DL-lactic-co-glycolic acid (PLGA). One method is based on the spreading coefficient theory for the formation of double walled microspheres by using single solvent, while the other is based on the property of PLLA not being soluble in ethyl acetate (EA). Characterization to check if the microspheres formed are double-walled was performed. The fabrication method where two solvents, dichloromethane (DCM) and ethyl acetate, were used gave double-walled microspheres, while the other where only dichloromethane was used gave composites. The double-walled microspheres were smaller in size compared to the composites, which were in the range of 100-600 microm. This can be attributed to the difference in the fabrication procedure. We were able to achieve better encapsulation efficiencies of more than 50% and slower release rates, which lasted for about 15 days. It was observed that size played a major role in the encapsulation efficiency and release rates. The possibility of achieving better results by studying the effect of concentration of polymer in solvent and the effect of using different polymers was investigated. PMID- 15653157 TI - Evaluating proteins release from, and their interactions with, thermosensitive poly (N-isopropylacrylamide) hydrogels. AB - Poly (N-isopropylacrylamide) (PNIPAAm) hydrogels possess a lower critical solution temperature (LCST) at around 32 degrees C. When the external temperature is raised above the LCST, the hydrogels experience abrupt and drastic shrinkage. This unique property makes them very useful for biomedical applications such as on-off switches for modulated drug delivery and tissue engineering. The aim of this work was to study the potential of using PNIPAAm hydrogels for protein delivery, and to obtain basic understandings of the protein-gel interactions as well as their effect on protein loading and release. PNIPAAm gels were synthesized with different crosslinker contents. The effects of crosslinker content, in vitro release temperature, protein loading level and molecular size as well as temperature cycling on protein release from PNIPAAm gels were examined. Greater amount of BSA was loaded using gels fabricated with lower crosslinker contents and loading solution with higher concentrations of BSA. An incomplete release of encapsulated BSA from the gels was observed in all cases. Enhanced mass transfer created by oscillating swelling-deswelling in response to temperature cycling across the LCST and lowering in vitro release temperature did not promote BSA release because of strong BSA-gel interactions. Evidence for the residual BSA in the gels after in vitro release was provided by dyeing the gels with protein determination reagent and shift in the LCST of the gels. In contrast, insulin release was much faster and more complete when compared to BSA because of its smaller size. The protein-gel interactions were analysed by investigating the LCST of, and state of water in, the blank and protein-loaded hydrogels. PMID- 15653158 TI - Surface-modified PLGA nanosphere with chitosan improved pulmonary delivery of calcitonin by mucoadhesion and opening of the intercellular tight junctions. AB - Surface-modified DL-lactide/glycolide copolymer (PLGA) nanospheres with chitosan (CS) were prepared by the emulsion solvent diffusion method for pulmonary delivery of peptide, i.e., elcatonin. The nanosphere suspension was successfully aerosolized with a nebulizer similar to the drug solution, whereas the microsphere suspensions could not be aerosolized. After pulmonary administration, CS-modified PLGA nanospheres were more slowly eliminated from the lungs than unmodified PLGA nanospheres. CS-modified PLGA nanospheres loaded with elcatonin reduced blood calcium levels to 80% of the initial calcium concentration and prolonged the pharmacological action to 24 h, which was a significantly longer duration of action than that by CS-unmodified nanospheres. These results were attributed to the retention of nanospheres adhered to the bronchial mucus and lung tissue and sustained drug release at the adherence site. In addition, CS and CS on the surface of the nanospheres enhanced the absorption of drug. The rank order of the absorption of the model drugs with CS solution was carboxyfluorescein>FITC-dextran-4 (FD-4; Mw. 4000)>FD-21 (Mw. 21,000)>FD70 (Mw. 70,000), which corresponded to the molecular weights ([Mw.] given in parentheses). The absorption-enhancing effect may have been caused by opening the intercellular tight junctions. PMID- 15653159 TI - Influence of molecular weight on oral absorption of water soluble chitosans. AB - Chitosan, a cationic polysaccharide, has been widely employed as dietary supplement and in pharmacological and biomedical applications. Although numerous studies have focused on its applications as pharmaceutical excipients or bioactive reagents, molecular weight (MW)-dependent pharmaceutical and bioactive properties remain unclear. As a preliminary study, we investigated the MW dependent Caco-2 cell layer transport phenomena (in vitro) and intestinal absorption patterns after oral administration (in vivo) of water-soluble chitosans (WSCs). The absorption of chitosan was significantly influenced by its MW. As the MW increases, the absorption decreases. Compared to high-MW chitosan (WSC 230K, MW=230 kDa), absorption profiles were observed to increase more than 23 and 25 times with WSC 3.8K (MW=3.8 kDa) in both in vitro and in vivo transport experiments, respectively. Furthermore, the chitosans showed concentration- and MW-dependent cytotoxic effects, and the chitosan oligosaccharides (MW<10 kDa) showed negligible cytotoxic effect on the Caco-2 cells. In consideration with safety and absorption profiles, chitosan oligosaccharides may be considered as safe and potential candidates for pharmaceutical and biomedical applications. PMID- 15653160 TI - Fabrication of monodispersed Taxol-loaded particles using electrohydrodynamic atomization. AB - In the fabrication of controlled drug release matrix, monodispersed particle sizes are usually preferred as they give a more uniform and precisely controlled release profile. Electrohydrodynamic atomization (EHDA) [J.C. Ijsebaert, K.B. Geerse, J.C.M. Marijnissen, J.W.J Lammers, P. Zannen, Electro-Hydrodynamic atomization of drug solutions for inhalation purposes, J. Appl. Physiol. 91(2001), 2735-2741.] is a method that can potentially produce particles with very low polydispersity. In the present work, Taxol-loaded poly-caprolactone (PCL) particles were fabricated using EHDA. Effort was undertaken to investigate the cause of the low yield of EHDA and to improve it to around 80%. This was achieved by increasing the ventilation and properly discharging the residual charges on the particulate cake at the filter paper. A phase Doppler particle analyzer (PDPA) was used to detect the spray modes of EHDA and the optimum operating conditions were determined. With differential scanning calorimetry (DSC), the uniformity of drug and polymer matrix is investigated. Moderate zeta potential values together with laser confocal micrographs give a possible explanation for the in vitro cell uptake data. These results are substantiated by a reasonably high encapsulation efficiency (EE) and sustained release profiles over 1-month period. The EHDA method is shown to be a potentially suitable technique to prepare close to monodispersed drug release particles. PMID- 15653161 TI - Stable paclitaxel formulations in oily contrast medium. AB - Stable paclitaxel/Lipiodol solutions as well as emulsions were developed for the treatment of solid tumors including hepatocellular carcinoma. Paclitaxel could be dissolved in Lipiodol, an oily contrast medium, but precipitated out and formed aggregates with time. Paclitaxel precipitation was due to the inter- and intra molecular hydrogen bonding of paclitaxel molecules. Time-dependent paclitaxel aggregation was completely prevented by adding small amounts of additional solvents, which are miscible with Lipiodol. It was also notable that paclitaxel helped in stabilizing the water-in-oil (w/o) type emulsion of Lipiodol and Iopamiro. The stability, physical properties and in vitro drug release profiles of the stable paclitaxel solutions and emulsions were characterized. When the stable oily paclitaxel solution was used for the treatment of B16F10 melanoma in C57BL/6 mice, the malignant cells were eradicated completely in 2 weeks, whereas the solid tumor grew rapidly and metastasized to the thigh and to other organs in the control group. Also, the mice survived for more than 1 year after the paclitaxel treatment, whereas all of those in the control group died in 40 days. PMID- 15653162 TI - Polymer nanocarriers protecting active enzyme cargo against proteolysis. AB - Polymeric nanocarriers (PNCs), proposed as an attractive vehicle for vascular drug delivery, remain an orphan technology for enzyme therapies due to poor loading and inactivation of protein cargoes. To unite enzyme delivery by PNC with a clinically relevant goal of containment of vascular oxidative stress, a novel freeze-thaw encapsulation strategy was designed and provides approximately 20% efficiency loading of an active large antioxidant enzyme, catalase, into PNC (200 300 nm) composed of biodegradable block copolymers poly(ethylene glycol)-b poly(lactic-glycolic acid). Catalase's substrate, H(2)O(2), was freely diffusible in the PNC polymer. Furthermore, PNC-loaded catalase stably retained 25-30% of H(2)O(2)-degrading activity for at least 18 h in a proteolytic environment, while free catalase lost activity within 1 h. Delivery and protection of catalase from lysosomal degradation afforded by PNC nanotechnology may advance effectiveness and duration of treatment of diverse disease conditions associated with vascular oxidative stress. PMID- 15653163 TI - Controlling release from the lipidic cubic phase by selective alkylation. AB - The lipidic cubic phase can be viewed as a molecular sponge consisting of interpenetrating nanochannels filled with water and coated by lipid bilayers. It has been used as a delivery matrix for low-molecular-weight drugs. For those that are water-soluble, release is fast and unregulated. This study seeks to exploit the lipid bilayer compartment as a location within the cubic phase in which to 'hydrophobically' anchor the water-soluble drug. This was accomplished by controlling partitioning into, and thus release from, the aqueous compartment of the cubic phase. Tryptophan was used as a surrogate water-soluble drug and alkylation was implemented to regulate release. By adjusting alkyl chain length, exquisite control was realized. Without alkylation, 20% of the tryptophan was released under standard conditions (infinite sink with a 30-mg cubic phase source at pH 7 and 20 degrees C) over a period of 30 min (t(20)). In the case of derivatives with alkyl chains two and eight carbon atoms long, t(20) values of 3 and 13 days, respectively, were observed. Eliminating the charge on tryptophan completely by alkylation produced a derivative that became irreversibly lodged in the lipid bilayer. The release behavior of the short-chain derivatives was mathematically modeled and parameters describing transport have been obtained. Cubic phase partition coefficients for tryptophan and its derivatives were measured to facilitate modeling. The implications of these findings with regard to the cubic phase and related delivery systems, and to vaccine efficacy are discussed. PMID- 15653164 TI - Brain-derived neurotrophic factor bound with lecithin derivative showed a markedly enhanced pharmacological potency due to its potent cell membrane affinity followed by prolonged MAPK activation. AB - We synthesized lecithinized brain-derived neurotrophic factor (lecithinized BDNF), in which an average of three molecules of a lecithin derivative were bound to recombinant human BDNF. We evaluated its pharmacological activity in C57BL/KsJ db/db mice, and assessed its targetability and affinity for the nervous system. Subcutaneously administered lecithinized-BDNF markedly reduced the plasma glucose level, food intake, and body weight in C57BL/KsJ-db/db diabetic mice. Its potency was more than 20 times greater than that of unmodified BDNF. We then studied the mechanism for the markedly enhanced pharmacological activity. In vitro cell growth activity of lecithinized-BDNF using the MTT assay was lower than unmodified BDNF, probably due to steric hindrance of the lecithin moieties. While the plasma BDNF level after subcutaneous administration of lecithinized-BDNF was not higher compared with unmodified BDNF. However, higher amount of lecithinized BDNF accumulated in the spinal cord was observed. Lastly, we found that in vitro binding capacity of lecithinized-BDNF for PC-pAB1 neural cells was much higher than unmodified BDNF. Moreover, lecithinized-BDNF bound to PC-pAB1 cells did not exchange with an excessive amount of unmodified BDNF or an excess of lecithinized BDNF. PC-pAB1 cells treated with lecithinized-BDNF showed sustained mitogen activated protein kinase (MAPK, ERK1/2) activation. These data would indicate that the high affinity of lecithinized-BDNF for the target cells, followed by prolonged MAPK activation, would play an important role in its potent pharmacological activity. PMID- 15653165 TI - Cell targeting by a generic receptor-targeted polymer nanocontainer platform. AB - Nanotechnology promises new avenues to medical diagnosis and treatment. Of special interest are injectable nanovehicles that are programmable towards specific targets, are able to evade the immune defense, and are versatile enough to be suited as carriers of complex functionality. Biotin-functionalized (poly(2 methyloxazoline)-b-poly(dimethylsiloxane)-b-poly(2-methyloxazoline) triblock copolymers were self-assembled to form nanocontainers, and biotinylated targeting ligands were attached by using streptavidin as a coupling agent. Specifically, fluorescence-labeled nanocontainers were targeted against the scavenger receptor A1 from macrophages, an important cell in human disease. In human and transgenic cell lines and in mixed cultures, receptor-specific binding of these generic carriers was followed by vesicular uptake. Low nonspecific binding supported the "stealth" properties of the carrier while cytotoxicity was absent. This versatile carrier appears promising for diagnostic or therapeutic medical use. PMID- 15653166 TI - Physical characterizations of microemulsion systems using tocopheryl polyethylene glycol 1000 succinate (TPGS) as a surfactant for the oral delivery of protein drugs. AB - Attempts were to develop microemulsion systems using medium chain triglyceride, deionized water, and TPGS as surfactant for the oral delivery of protein drugs or poorly water-soluble drugs. Phase diagrams were constructed to elucidate the phase behavior of systems composed of Captex 300 and water with D-alpha tocopheryl polyethylene glycol 1000 succinate (TPGS) as main surfactant, polysorbates (Tween 20, Tween 40, Tween 60 and Tween 80) as adjuvant surfactants, and polyethylene glycols (PEG 400 and PEG 600) and polyols (ethanediol, 1,2 propanediol, 1,3-propanediol, 1,3-butanediol, 1,4-butanediol and glycerin) as cosurfactants. The ratios of TPGS to Tweens, PEGs or polyols (K(m)) were set at 4/1, 2/1, 1/1, 1/2, and 1/4. The phase diagram for H(2)O/Captex 300/TPGS system reveals that when TPGS was used as a sole surfactant, it is not capable of producing isotropic solutions of water and oil over a wide range of the compositions. H(2)O/Captex 300/TPGS/Tweens systems with various K(m), regardless of the adjuvant surfactant used were capable of producing an isotropic phase. The extension of microemulsion phase and the presence and extension of the gel phase were found to be dependent on the surfactant mixture. The phase diagrams of H(2)O/Captex 300/TPGS systems using polyols as cosurfactants demonstrate that the types of polyols have a slight effect on the region of existence of the microemulsions. Comparison between the isotropic regions for the polyols system reveals that as the relative concentration of polyols increase, the isotropic region decrease in size. This decrease is towards the S(mix)-water axis indicating that as the relative concentration of polyols increases the maximum amount of oil solubilized decreases. The gel region decreased in size with the increase of polyols weight ratio. All polyols do not solubilized Captex 300 without using TPGS as surfactant. PMID- 15653167 TI - In vivo evaluation of 3-O-alkyl ester transdermal prodrugs of naltrexone in hairless guinea pigs. AB - Naltrexone (NTX) is a potent competitive antagonist with high affinity for the mu opioid receptor. Therapeutically, NTX is used for the treatment of alcohol dependence and opioid addiction; however, it does not have the ideal physicochemical properties necessary to achieve therapeutic plasma concentrations via the transdermal route. The aim of the present investigation was to evaluate the in vivo transdermal delivery of three 3-O-alkyl ester prodrugs of NTX, including NTX-3-O-acetate (ACE-NTX), NTX-3-O-propionate (PROP-NTX), and NTX-3-O hexanoate (HEX-NTX) in hairless guinea pigs. The pharmacokinetic parameters for NTX and the 3-O-alkyl ester prodrugs of NTX were determined after intravenous drug administration and topical drug application of transdermal therapeutic systems (TTS) in guinea pigs. The results of the in vivo studies showed mean steady-state plasma concentrations of NTX from NTX, ACE-NTX, PROP-NTX and HEX-NTX at 4.2, 25.2, 16.0, and 8.3 ng/mL, respectively. These NTX plasma concentrations were maintained for 48 h. The results of these in vivo studies demonstrated that ACE-NTX and PROP-NTX prodrugs of NTX were the most promising drug candidates for transdermal delivery. PMID- 15653169 TI - Part of membrane-bound Abeta exists in rafts within senile plaques in Tg2576 mouse brain. AB - To clarify whether rafts are the site of abnormal amyloid beta protein (Abeta) deposition, we examined the ultrastructural localization of both flotillin-1 (pre embedding) and Abeta (post-embedding) in Tg2576 mouse brains. After observing the exact areas of senile plaques by reflection contrast microscopy, we observed these same plaques under an electron microscope. Membrane-bound Abeta was predominantly observed on plasma membranes of small processes in diffuse plaques. Non-fibrillar and fibrillar Abeta was increased in primitive plaques, and the fibrillar form was predominant in mature plaques. The number of flotillin-1 positive rafts per field in mature plaques was prominently less than those outside of the plaques, in diffuse plaques and in primitive plaques. The colocalization of flotillin-1 with Abeta42 appeared approximately 10% of flotillin-1-positive rafts within senile plaques, while there was no colocalization found outside of the plaques. This study ultrastructurally demonstrated that part of membrane-bound Abeta exists in lipid rafts within senile plaques, and suggests that rafts could be one of the sites for initial Abeta deposition. PMID- 15653168 TI - HSV amplicon-mediated Abeta vaccination in Tg2576 mice: differential antigen specific immune responses. AB - Given the participation of amyloid beta (Abeta) in Alzheimer's disease (AD) pathogenesis the derivation of experimental therapeutics to prevent Abeta fibrillogenesis and/or enhance removal of parenchymal amyloid deposits represent viable disease-modifying approaches. Active Abeta-based immunotherapies have shown promise in mouse AD models, but application in human trials was accompanied by moderate brain inflammation in a subset of patients. Immune-shaping vaccine platforms may mitigate adverse effects. Herein, we describe the use of herpes simplex virus (HSV)-derived amplicons to elicit distinctive immune responses against Abeta. Two vaccine vectors were constructed: one expressing Abeta1-42 alone (HSVAbeta), and a second expressing Abeta1-42 fused with the molecular adjuvant tetanus toxin Fragment C (HSVAbeta/TtxFC). Peripheral administration of these vaccines augmented humoral responses to Abeta and reduced CNS Abeta deposition in Tg2576 AD mice. Interestingly and unexpectedly, HSVAbeta vaccination was uniquely toxic and incited the expression of pro-inflammatory molecule transcripts within the hippocampi of Tg2576 mice, suggesting that this paradigm may serve as a relevant model to study Abeta vaccine-elicited CNS inflammatory syndromes. PMID- 15653170 TI - Beta-Amyloid peptide25-35 depresses excitatory synaptic transmission in the rat basolateral amygdala "in vitro". AB - The effects of beta-amyloid peptide25-35 on resting membrane potential, spontaneous and evoked action potential and synaptic activity have been studied in basolateral amygdaloid complex on slices obtained from adult rats. Intracellular recordings reveal that perfusion with beta-amyloid peptide25-35 at concentrations of 400 nM and less did not generate any effect on resting membrane potential. However, concentrations in the range of 800-1200 nM produced an unpredictable effect, depolarization and/or hyperpolarization, which were blocked by tetrodotoxin or 6-cyano-7-nitroquinoxaline-2,3-dione+D-(-)-2-amino-5 phosphonopentanoic acid together with bicuculline. Excitatory and inhibitory evoked responses mediated by glutamic acid or gamma-aminobutyric acid decreased in amplitude after beta-amyloid peptide25-35 perfusion. Additionally, results obtained using the paired-pulse protocol offer support for a presynaptic mode of action. To determine which type of receptors and/or channels are involved in the presynaptic mechanism of action, a specific blocker of alpha-7 nicotinic receptors (methyllycaconitine citrate) or L-type calcium channel blockers (calcicludine or nifedipine) were used. beta-amyloid petide25-35 decreased excitatory postsynaptic potentials amplitude in control conditions and also in slices permanently perfused with methyllycaconitine citrate. However, this effect was blocked in slices perfused with calcicludine or nifedipine suggesting the involvement of the L-type calcium channels. On the whole, these experiments provide evidence that beta-amyloid peptide25-35 affects neurotransmission in basolateral amygdala and its action is mediated through L-type calcium channels. PMID- 15653171 TI - Oxidative stress related markers in the "VITA" and the centenarian projects. AB - Oxidative stress seems to play an important role in the pathophysiology of Alzheimer's disease (AD). At present there are no easily accessible biochemical markers for AD. We performed activity assays for platelet MAO-B and erythrocyte Cu/Zn-SOD as well as Western blotting for these two proteins. Moreover, we assessed plasma lactoferrin and performed RFLP-analysis for the MAO-B-intron-13 polymorphism in patients from the Vienna-Transdanube Aging (VITA) and from the so called centenarian project. The first one, VITA, is a community-based cohort study of all 75 years old inhabitants of a geographical region of Vienna. The centenarian project investigates chronic care in-old patients suffering from AD. In both sexes platelet MAO-B activity increased significantly in the AD group, and Cu/Zn-SOD activity decreased, but the latter effect was significant only in females. No significant difference was found regarding plasma lactoferrin. No correlation was found between MAO-Bi13 and MAO-B platelet activity or allele MAO Bi13 and disease frequency. These results point to the possibility that a combination of MAO-B and SOD activity levels might be useful tools for an early diagnosis of AD. PMID- 15653172 TI - Complete ascertainment of dementia in the Swedish Twin Registry: the HARMONY study. AB - The purpose of this report is to describe the Study of Dementia in Swedish Twins (known as HARMONY), including procedures for complete ascertainment of all cases of Alzheimer's disease (AD) and other dementias in 14,435 individuals aged 65 and older from the national Swedish twin registry. Telephone cognitive screening identified 11.5% as positive for cognitive dysfunction. Clinical diagnoses were completed for 1557 individuals, including individuals who screened positive, their twin partners, and a sample of normal controls. Estimated prevalence of dementia ranged from 1.4% for age 65-69 to 29.2% for age 90 and older. Concordance rates for Alzheimer's disease were 59% for monozygotic twins, 32% for like-sexed, and 24% for unlike-sexed dizygotic twins. Among monozygotic twins where both twins had Alzheimer's disease, the within pair difference in age of onset ranged from both becoming demented in the same year to 7 years difference in onset. PMID- 15653173 TI - A new promoter polymorphism in the alpha-1-antichymotrypsin gene is a disease modifier of Alzheimer's disease. AB - Increased levels of alpha-1-antichymotrypsin (ACT), a protease inhibitor and an acute phase protein, have been found in the brain and peripheral blood of patients with Alzheimer's disease (AD). Patients from northern Italy with a clinical diagnosis of probable AD, and patients with early onset AD (EOAD) from UK with AD neuropathological diagnosis were genotyped for a new polymorphism in the promoter region of the ACT gene which has been shown to affect ACT expression. A subset of patients with clinical AD from northern Italy was also followed up for 2 years and monitored for cognitive decline. The ACT TT promoter genotype was associated with an increased risk of EOAD independently from the presence of the apolipoprotein E (APOE) epsilon 4 allele. After manifestation of the disease the ACT TT genotype was also associated with faster cognitive decline in patients with the APOE allele epsilon 4. The ACT gene appears to influence the early clinical development of the disease, and the interaction of the ACT and APOE genes affects clinical progression of AD. PMID- 15653174 TI - Genotypes and haplotypes in the IL-1 gene cluster: analysis of two genetically and diagnostically distinct groups of Alzheimer patients. AB - Increased risk of Alzheimer's disease (AD) has been associated with polymorphisms in the IL-1 gene cluster, and in particular with the IL-1alpha-889 T/T genotype. However, this association is still unclear, and needs further investigation. In order to clarify the role of these polymorphisms in the complex pathogenesis of AD we examined genotype and haplotype frequencies of the two C-to-T SNPs at position -889 and -551 in the IL-1alpha and IL-1beta genes, respectively, and of the 86 bp VNTR intron-2 polymorphisms in the IL-1Ra gene. The analysis was performed in two genetically and diagnostically distinct groups of sporadic AD from Italy and the USA. In the Italian group a significant association between the IL-1alpha-889 T/T genotype and AD (OR=3.022, 95% CI: 1.001-9.119) was found, whereas no difference was found in the group from the USA. Results were also compared with previously published studies that analyzed the same IL-1 polymorphisms in AD. In both groups, the analysis of the estimated haplotypes shows that AD patients and controls who carry the IL-1beta-511 C allele, were also more frequently carriers of the IL-1Ra 1 allele (haplotypes -C-1). The total frequency of the two -C-1 haplotypes (C-C-1 plus T-C-1) was about one half of the total frequency of the eight estimated haplotypes. This was confirmed by significant linkage disequilibrium between these two loci in both the Italian and USA groups. In the Italian group a weak association of the T-C-2 haplotype with the disease (OR=1.648, 95% CI: 1.519-1.788) was also found, whereas in the USA group no difference was found. Although ours and other published data on different samples of Caucasian and non-Caucasian AD show a great heterogeneity in the frequencies of the IL-1alpha-889, the IL-1beta-511 and the IL-1Ra VNTR gene polymorphisms, we confirm the role of the IL-1alpha-889 T/T genotype as a risk factor for sporadic AD, and show the presence of an allelic association between IL-1beta C and IL-1Ra 1 alleles in both the Italian and the USA groups, confirmed by the presence of significant levels of linkage disequilibrium between these two loci. PMID- 15653175 TI - Immunochemical crossreactivity of antibodies specific for "advanced glycation endproducts" with "advanced lipoxidation endproducts". AB - Antibodies against advanced glycation endproducts (AGEs) are used for their immunohistological localization in tissues, for example in Alzheimer's disease (AD) or diabetes. Many monoclonal and polyclonal antibodies have been used, and their specificity is unknown in most cases. Increased radical production, leading to the formation of lipid-derived reactive carbonyl species, such as malondialdehyde (MDA), acrolein, and glyoxal, is a characteristic aspect of age related diseases like Alzheimer's disease or diabetic polyneuropathy. These reactive carbonyl species are able to modify proteins, resulting in AGE related structures, termed "advanced lipoxidation products" (ALEs). In this study, the monoclonal carboxymethyllysine-specific antibody 4G9 and the polyclonal AGE antibody K2189 were tested for their immunoreactivity towards these carbonyl derived protein modifications. To investigate which carbonyl-modified amino acid side chains are specifically recognized by these antibodies, peptide membranes were incubated with glyoxal, MDA and acrolein. As model proteins, microtubuli associated protein tau (MAP-tau), beta-amyloid, human serum albumin and chicken egg albumin were incubated likewise. It was found that both antibodies detected reaction products of these carbonyl compounds on lysine- and arginine residues and for the protein modification, it was found that some epitopes might not be detected. In conclusion, AGE-antibodies might not only detect sugar-derived AGEs but also structures derived from lipid peroxidation products (serving as markers of oxidative stress). PMID- 15653176 TI - Apolipoprotein E, cardiovascular disease and cognitive function in aging women. AB - The apolipoprotein E (APOE) e4 allele increases risk of Alzheimer disease and cardiovascular diseases. We examined APOE genotypes alone or combined with cardiovascular conditions in relation to cognitive function in 4227 Nurses' Health Study participants, 70-80 years old. From 1995 to 2001, and again 2 years later, participants received telephone cognitive assessments of general cognition, category fluency, verbal memory and working memory. In biennial questionnaires since 1976, participants have provided extensive health information including cardiovascular conditions. Compared with women with the e3/3 genotype, e4 carriers performed worse at baseline across all tests (mean global composite score was lower by 0.10 points (95% confidence interval=-0.15, 0.05)) and declined more (mean change in global score was -0.07 points (95% CI= 0.12, -0.03)), with a strong allele dose-response trend (P-trend=0.0003). Among participants 75+ years, e2 carriers performed best. Women with an e4 allele and cardiovascular conditions such as transient ischemic attack or untreated hypertension had the worst cognition. Thus, APOE genotypes strongly influenced cognitive function and decline; prevention of cardiovascular disease may limit these effects. PMID- 15653177 TI - Vascular risk factors and cognitive function in a sample of independently living men. AB - Decline of cognitive function with age may be due, in part, to atherosclerotic changes. The aim of the present study was to determine the relative contribution of vascular risk factors to cognitive functioning in a non-clinical sample of men. Cognitive tests were administered to 400 independently living men aged 40-80 years. The measures included short-term memory, speed of information processing, verbal and visual long-term memory, word fluency, cognitive flexibility, an estimate of verbal intelligence, and general cognitive status. Systolic blood pressure, serum cholesterol, glucose levels, smoking, alcohol intake, body mass index, homocysteine and peak expiratory flow rate were entered as independent variables into a multiple regression model, after adjustment for age and education. Hierarchical regression analyses revealed independent contributions of the combination of vascular risk factors in explaining the observed variance in performance on tests of cognitive functioning targeted at information processing capacity and speed and general cognitive status. Of the individual predictor variables, alcohol intake and homocysteine levels were significantly associated with processing capacity and speed, and peak expiratory flow rate was significantly associated with general cognitive status. Our results indicate that the combination of several independent vascular risk factors predicts performance on cognitive tests of information processing capacity and speed in a population based sample of middle-aged and elderly men. PMID- 15653178 TI - Measures of brain morphology and infarction in the framingham heart study: establishing what is normal. AB - Numerous anatomical and brain imaging studies find substantial differences in brain structure between men and women across the span of human aging. The ability to extend the results of many of these studies to the general population is limited, however, due to the generally small sample size and restrictive health criteria of these studies. Moreover, little attention has been paid to the possible impact of brain infarction on age-related differences in regional brain volumes. Given the current lack of normative data on gender and aging related differences in regional brain morphology, particularly with regard to the impact of brain infarctions, we chose to quantify brain MRIs from more than 2200 male and female participants of the Framingham Heart Study who ranged in age from 34 to 97 years. We believe that MRI analysis of the Framingham Heart Study more closely represents the general population enabling more accurate estimates of regional brain changes that occur as the consequence of normal aging. As predicted, men had significantly larger brain volumes than women, but these differences were generally not significant after correcting for gender related differences in head size. Age explained approximately 50% of total cerebral brain volume differences, but age-related differences were generally small prior to age 50, declining substantially thereafter. Frontal lobe volumes showed the greatest decline with age (approximately 12%), whereas smaller differences were found for the temporal lobes (approximately 9%). Age-related differences in occipital and parietal lobe were modest. Age-related gender differences were generally small, except for the frontal lobe where men had significantly smaller lobar brain volumes throughout the age range studied. The prevalence of MRI infarction was common after age 50, increased linearly with age and was associated with significantly larger white matter hyperintensity (WMH) volumes beyond that associated with age-related differences in these measures. Amongst men, the presence of MRI infarction was associated with significant age-related reductions in total brain volume. Finally, statistically significant associations were found between the volume of MRI infarcts in cubic centimeters and all brain measures with the exception of parietal lobe volume for individuals where the volume of MRI infarctions was measured. These data serve to define age and gender differences in brain morphology for the Framingham Heart Study. To the degree participants of the Framingham Heart Study are representative the general population, these data can serve as norms for comparison with morphological brain changes associated with aging and disease. In this regard, these cross-sectional quantitative estimates suggest that age-related tissue loss differs quantitatively and qualitatively across brain regions with only minor differences between men and women. In addition, MRI evidence of cerebrovascular disease is common to the aging process and associated with smaller regional brain volumes for a given age, particularly for men. We believe quantitative MRI studies of the Framingham community enables exploration of numerous issues ranging from understanding normal neurobiology of brain aging to assessing the impact of various health factors, particularly those related to cerebrovascular disease, that appear important to maintaining brain health for the general population. PMID- 15653179 TI - The exercise-induced expression of BDNF within the hippocampus varies across life span. AB - Voluntary exercise increases hippocampal brain-derived neurotrophic factor (BDNF) expression in young animals. In this investigation we examined the induction of BDNF protein in the hippocampus of young (2 months), late middle-aged (15 months) and old (24 months) animals over 4 weeks of exercise. Average running distances decreased with age, with the old animals also maintaining a constant level of activity over time, whereas the other groups tended to increase their average running distance. All animals demonstrated a biphasic profile of BDNF protein induction, with a significant (P<0.05) increase after 1 week of exercise followed by a decrease to near sedentary levels at 2 weeks. After this, BDNF protein levels increased significantly (P<0.05), as compared to baseline, primarily only in the young animals. In whole hippocampal homogenates, only particular BDNF mRNA exons were significantly (P<0.05) changed as a result of exercise, with the largest induction occurring in young animals. BDNF protein induction may, therefore, not be directly correlated with significant mRNA changes. Exercise may represent a therapeutic tool for disorders which involve a decrease in BDNF. PMID- 15653180 TI - Effects of age and dopamine D2L receptor-deficiency on motor and learning functions. AB - Decreases in the activity or density of dopamine D2 receptor (D2R) have been associated with age-related changes and neurodegenerative diseases such as Parkinson's disease. There are two isoforms of the D2R, termed the D2 long receptor (D2LR) and D2 short receptor (D2SR). To study the function of these two isoforms and their role in aging, we generated mice selectively lacking D2LR (D2L /-). Here, we showed that middle-aged (12 months) to aged wild-type (WT) mice (22 24 months) displayed significantly lower levels of motor and learning functions than young WT mice (3 months). Interestingly, young D2L-/- mice (which still express D2SR) showed behavioral deficits similar to aged WT mice. It is possible that deletion of the D2LR might facilitate the aging process in mice. Our results also suggest that a deterioration of the D2LR (but not D2SR) system during aging may account, at least in part, for the motor and learning deficits exhibited in aged WT mice. We also showed that the critical age at which significant reduction in behavior occurred varied among different behaviors. Defining the age-related critical periods and understanding the role of the two D2R isoforms in aging may facilitate the development of new strategies for delaying or ameliorating age related motor and learning impairments. PMID- 15653181 TI - Event-related potentials accompanying motor preparation and stimulus expectancy in the young, young-old and oldest-old. AB - Although aging is accompanied by neurobiological changes and increased susceptibility to many neurological disorders, little is known about neurophysiological changes that start in old age. Here, neurophysiological changes during old age were assessed by recording brain potentials associated with motor preparation and stimulus expectancy (contingent negative variation, CNV) in young-old (60-69), oldest-old (85-98), and young (17-23) subjects. Individual trials began by a button press, followed 2.5 s later by either a low or high pitch tone. In the "motor" condition subjects responded following high pitch tones (P=0.20); in the "non-motor" condition subjects did not respond. Motor condition CNV amplitudes in the oldest old were more positive than the young and young-old groups, which were similar. In the non-motor condition, the young-old and oldest-old had similar CNV amplitudes that were positive in polarity, and were significantly different from young subjects. Motor potentials before button presses that started the trials were comparable among groups. Results show that neural activity associated with motor preparation and stimulus expectancy changes during advanced age, and that group differences can be modulated by task requirements. PMID- 15653182 TI - Sex differences in age-related motor slowing in the rhesus monkey: behavioral and neuroimaging data. AB - The nigrostriatal system is critical for fine motor function and its deterioration during aging is thought to underlie the decline in fine manual ability of old persons. Because estrogen has a neuroprotective effect on this system, one might expect women's motor function to be less vulnerable to the detrimental effects of aging than that of men. We examined this hypothesis in the rhesus monkey, which has been established as an excellent model of human age related motor impairment. We tested 28 young and old rhesus monkeys of both sexes in a task involving the retrieval of a Life Saver candy from rods of different complexity to determine whether fine motor ability (1) is sexually dimorphic, (2) declines with age and (3) declines differently in males and females. In addition, we measured the whole brain volume, the volumes of the caudate, putamen, hippocampal formation and the area of the corpus callosum in a subset of the monkeys (n=15) for which magnetic resonance images of the brain were available. All monkeys performed similarly in the test with the simplest rod. In the test with complex rods; however, age-related slowing of motor function was evident in males, but not in females. Age-related decreases in the normalized caudate and putamen volumes were similar in males and in females. In addition, motor speed was not significantly correlated to any of the neuroanatomical measures under study. Further studies will be necessary to uncover the neurohormonal bases of the differential age-related motor decline between males and females. PMID- 15653184 TI - Comparative radiation impact on biota and man in the area affected by the accident at the Chernobyl nuclear power plant. AB - A methodological approach for a comparative assessment of ionising radiation effects on man and non-human species, based on the use of Radiation Impact Factor (RIF) - ratios of actual exposure doses to biota species and man to critical dose is described. As such doses, radiation safety standards limiting radiation exposure of man and doses at which radiobiological effects in non-human species were not observed after the Chernobyl accident, were employed. For the study area within the 30km ChNPP zone dose burdens to 10 reference biota groups and the population (with and without evacuation) and the corresponding RIFs were calculated. It has been found that in 1986 (early period after the accident) the emergency radiation standards for man do not guarantee adequate protection of the environment, some species of which could be affected more than man. In 1991 RIFs for man were considerably (by factor of 20.0-1.1 x 10(5)) higher compared with those for selected non-human species. Thus, for the long term after the accident radiation safety standards for man are shown to ensure radiation safety for biota as well. PMID- 15653183 TI - White matter lesions are associated with cortical atrophy more than entorhinal and hippocampal atrophy. AB - The goal of this study was to examine the relationship between subcortical vascular disease and brain atrophy in patients with Alzheimer's disease (AD) and mixed dementia (i.e., AD and subcortical vascular disease together). MRI was performed on 77 cognitively normal (CN) subjects, 50 AD and 13 mixed dementia patients. Subcortical vascular disease was determined by white matter hyperintensities (WMH) volume and presence of subcortical lacunes. Brain atrophy was measured using total brain cortical gray matter (CGM), entorhinal cortex (ERC) and hippocampal volumes. CGM volume, but not ERC or hippocampal volume was inversely related to WMH volume in patients and controls. In contrast, no relationship was detected between CGM, ERC, or hippocampal volumes and subcortical lacunes. Furthermore, no interaction was found between WMH and diagnosis on cortical atrophy, implying that WMH affect cortical atrophy indifferently of group. These results suggest that subcortical vascular disease, manifested as WMH, may affect cortical atrophy more than ERC and hippocampal atrophy. Further, AD pathology and subcortical vascular disease may independently affect cortical atrophy. PMID- 15653185 TI - Potential of largemouth bass as vectors of 137Cs dispersal. AB - We conducted a radio telemetry study on the movements of potentially contaminated largemouth bass between Steel Creek, a restricted access (137)Cs contaminated stream on the Savannah River Site (located in South Carolina, USA), and the publicly accessible Savannah River. Largemouth bass were relatively mobile in lower Steel Creek and the portion of the Savannah River near Steel Creek, and there was considerable movement between these two habitats. Largemouth bass had home ranges of about 500 linear meters of shoreline in the Savannah River but sometimes moved long distances. Such movements occurred primarily during the spawning season, largely upstream, and increased when water levels were changing or elevated. However, approximately 90% of the largemouth bass observations were within 10 km of Steel Creek. The total quantity of (137)Cs transported into the Savannah River by largemouth bass was much less than transported by water and suspended sediments discharged from Steel Creek. We conclude that largemouth bass from the Savannah River Site are unlikely to be responsible for long distance dispersal of substantial radiological contamination in the Savannah River. PMID- 15653186 TI - Transfer of 137Cs to rice plants from various paddy soils contaminated under flooded conditions at different growth stages. AB - Soil blocks from 18 paddy fields around three Korean nuclear power plant sites were put into lysimeters. Greenhouse experiments were carried out to investigate the (137)Cs transfer from these paddy soils to rice plants for its deposition at different growth stages. A solution of (137)Cs was applied to the flooded lysimeters at 2-3 different stages. The applied (137)Cs was mixed with the topsoil only at the pre-transplanting application. The transfer was quantified with a transfer factor based on the unit-area deposition (TF(a), m(2)kg(-1)-dry). The TF(a) in the pre-transplanting application showed a remarkable variation with the soils. However, the differences in the mean values among the study sites were not statistically significant. The straw TF(a) was 2-3 times higher than the corresponding seed value. The early-tillering stage and booting stage applications resulted in a higher transfer than the pre-transplanting application by factors of, on an average, 2 and 16 for the straws, and 3 and 25 for the hulled seeds, respectively. The (137)Cs transfer was found to correlate negatively with the soil pH and positively with the organic matter content. Based on the present results, the representative (137)Cs TF(a) values for the rice are proposed for use in the whole of Korea for the deposition at three different growth stages. PMID- 15653187 TI - Genotoxicity and cytotoxicity assay of water sampled from the underground nuclear explosion site in the north of the Perm region (Russia). AB - The results of our study revealed a local biologically relevant surface water contamination in the radionuclide anomaly in the north of Russia (Perm region) by means of Allium schoenoprasum L. anaphase-telophase chromosome aberration assay. This radionuclide anomaly was formed in 1971 as a result of an underground nuclear explosion with soil excavation. Specific activities of main dose-forming radionuclides in all examined reservoirs are below intervention levels officially adopted in Russia for drinking water. We found that (90)Sr significantly contributes to induction of cytogenetic disturbances. Our previous data and the data described here suggest that metal and radionuclide combined exposure (with the dose below permissible exposure limits for human) may cause substantial biological effects. These effects are in part due to synergic response. The findings described here indicated that development of a new concept of radiation protection for humans and biota should be based on the clear understanding of biological effects of low doses of radiation in chronic exposure to multi pollutant mixtures. PMID- 15653188 TI - Levels of 137Cs in muddy sediments of the seabed of the Bay of Cadiz, Spain. Part I. Vertical and spatial distribution of activities. AB - This is the first of a two-part paper. This first part describes a detailed study conducted of the spatial and vertical distribution of the concentration of the radionuclide (137)Cs, utilising columns of sediment extracted from the seabed of the Bay of Cadiz (Spain) as specimens. The activity concentrations have been measured by means of gamma spectrometry using a HPGe detector and (137)Cs is used as an indicator of the sedimentation and contamination processes in the zone. At the same time the relationship between the levels of activity found and other hydrodynamic and sedimentological variables characteristic of the zone have been studied. The experimental results obtained will also serve to develop a model of the distribution of (137)Cs, which will be presented in the second part of this paper. PMID- 15653189 TI - Levels of 137Cs in muddy sediments on the seabed in the Bay of Cadiz (Spain). Part II. Model of vertical migration of (137)Cs. AB - This second part of the study reports the development of a model to describe the vertical migration of the artificial radioisotope (137)Cs in the sediment column on the seabed of the Bay of Cadiz. The application of the model provides an overall picture of the process of sedimentation in the Inner Bay of Cadiz. The spatial distribution of the rate of sedimentation enables us to study the sources of sediments and the means by which the sediments have been transported. A method has been derived from the rate of sedimentation to perform the dating of the layers of sediment. The model describes the behaviour of (137)Cs in the area under study, taking into account the time of residence in the zones that are the source of accumulation, the origin of the sedimentary material, together with the diffusion of the radionuclide in the sediment of the seabed. PMID- 15653190 TI - Modelling the long-term behaviour of radiocaesium and radiostrontium in two Italian lakes. AB - The present paper describes the application of a state-of-the-art model to two lakes in central Italy (Trasimeno and Monterosi) for the assessment of migration parameters of (137)Cs and (90)Sr (migration velocity to sediment, transfer rates from sediment to water and to bottom sediment). Applications of a compartment model and a model based on the diffusion equation to predict the behaviour of pollutants in water and through bottom sediment are presented and discussed. The application of the diffusion equation shows some difficulties of a general nature and typical of such a modelling approach. Moreover, there is no evidence of significant improvements of the model performances when the diffusion equation is applied. Very low levels of sedimentation rate of suspended matter in Lake Monterosi were evaluated by the quantitative assessment of radiocaesium migrating to bottom sediment. This suggests that, in this lake, the removal of radionuclide from the water column is mainly due to the turbulent mixing of bottom sediment causing radionuclide burial. PMID- 15653191 TI - Concentrations of 137Cs in lynx (Lynx lynx) in relation to prey choice. AB - Concentrations of (137)Cs were determined in 747 lynxes killed in Norway during the period 1986-2001. Highly variable (137)Cs concentrations and aggregated transfer coefficient values were observed, probably caused by variable (137)Cs concentrations in prey and the lynx's extensive home ranges and roaming distances. Adult lynxes had higher (137)Cs concentrations than sub-adults, and lynxes killed in regions with extensive reindeer grazing areas were more contaminated than others. A model with (137)Cs deposition density, the year lynxes were killed, age, and extent of reindeer grazing area accounted for 50% of the variability in observed (137)Cs concentrations. The analyses were equivocal regarding the influence of stomach content on (137)Cs concentrations in lynx muscle, i.e., on the lynx's specialization in prey species. Gender was not significant. Information on caesium retention in lynx and better estimates of deposition densities in lynxes' home ranges are important for further elucidation of factors influencing (137)Cs contamination in lynxes. PMID- 15653193 TI - Human pheromones and sexual attraction. AB - Olfactory communication is very common amongst animals, and since the discovery of an accessory olfactory system in humans, possible human olfactory communication has gained considerable scientific interest. The importance of the human sense of smell has by far been underestimated in the past. Humans and other primates have been regarded as primarily 'optical animals' with highly developed powers of vision but a relatively undeveloped sense of smell. In recent years this assumption has undergone major revision. Several studies indicate that humans indeed seem to use olfactory communication and are even able to produce and perceive certain pheromones; recent studies have found that pheromones may play an important role in the behavioural and reproduction biology of humans. In this article we review the present evidence of the effect of human pheromones and discuss the role of olfactory cues in human sexual behaviour. PMID- 15653194 TI - Thrombophilias and recurrent pregnancy loss: a critical appraisal of the literature. AB - Thrombophilias are suggested to play a role in recurrent miscarriage. The aim of this study was to evaluate the literature of the past 10 years regarding the association between thrombophilias and recurrent miscarriage. We concluded that there is a large variety in applied study methodology. Therefore, we defined criteria for an adequate study on the relationship of thrombophilias on recurrent pregnancy loss: (i) no exclusion criteria for patients or at least the same criteria for patients and controls; (ii) a clear definition of the gestational age at previous losses; (iii) a well-described control group; (iv) clear description of the test methods and moment of testing; and (v) a clear description of the (non) significant differences or odds ratio between cases and controls. Eleven out of 69 studies fulfilled these criteria. Their results show significant higher serum homocysteine levels among women with a history of recurrent miscarriage. No relation was found between recurrent miscarriage and the methylenetetrahydrofolate reductase C667T mutation. No relation was observed for the levels of antithrombin, protein C and protein S. Seven studies on the association of factor V Leiden (FVL) and/or pathologic activated protein C ratio (pAPCR) showed that FVL may play a role in second trimester losses, as do antiphospholipid antibodies. Studies on the prothrombin gene mutation yielded conflicting results. Consequently, large prospective studies according to the aforementioned criteria are needed to establish if there is a relationship between thrombophilias and recurrent miscarriage at all. At present, there is only justification for testing for homocysteine levels, antiphospholipid antibodies and FVL in women with a history of recurrent miscarriage. PMID- 15653195 TI - Prenatal diagnosis and management of mild fetal pyelectasis: implications for neonatal outcome and follow-up. AB - OBJECTIVES: To describe the natural history of pyelectasis from its detection in the second trimester to delivery, its capability to predict renal pathology and whether prenatal development of pelvic dilatation is correlated to its postnatal evolution. STUDY DESIGN: A retrospective analysis involving 375 fetuses with a complete urological follow-up. Prenatal ultrasound was correlated with the results of postnatal investigation and frequency of postnatal surgery was established. RESULTS: Two-hundred and eighty cases underwent at least two examinations before birth. 73.1% were male fetuses. 57.4% had a bilateral pyelectasis. Prenatal evolution of pelvic dilatation was the following:18.6% of the cases normalized, in 34.6% of the cases the dilatation reduced but did not disappear, in 30.7% it remained unchanged, while it worsened in 16.4%. One case from the first group, three cases from the second, seven cases from the third and 11 cases from the fourth needed surgical treatment. 1.9, 7.2, 18.6, 23.9% of cases respectively worsened after birth in the four groups (trend: P=0.001). CONCLUSIONS: Prenatal diagnosis of pyelectasis improves the outcome of these children due to a surgical approach that avoids renal damage. There is a good correlation between prenatal evolution and postnatal outcome, although a postnatal follow-up is opportune in those cases that normalized before birth. PMID- 15653196 TI - Endothelium-derived nitric oxide metabolites and soluble intercellular adhesion molecule-1 in diabetic and normal pregnancies. AB - OBJECTIVE: Endothelial dysfunction has been demonstrated in adult subjects with diabetes. We studied if maternal diabetes is associated with altered endothelial function in the fetus, as this might shed light on mechanisms by which adult diseases are programmed in utero. STUDY DESIGN: Total nitrate/nitrite (NOx) concentration was measured spectrophotometrically with the Griess reagent method. Soluble intercellular adhesion molecule-1 (sICAM-1) concentration was measured by enzyme-linked immunoassay. RESULTS: Venous cord serum NOx concentration at birth was highest in pregnancies complicated by type 1 diabetes (29.5+/-1.8 micromol/l, n=63) (P<0.0001 versus controls) and lowest in normal pregnancies (19.0+/-1.0 micromol/l, n=56). The concentration was intermediate in pregnancies complicated by gestational diabetes (23.9+/-2.7 micromol/l, n=24), but not significantly higher than in normal pregnancies (P=0.172). Venous cord serum sICAM-1 concentration did not differ between the three groups (P=0.191). Maternal serum NOx concentration in the third trimester was higher in pregnancies complicated by type 1 diabetes (22.9+/-3.4 micromol/l, n=22) than in normal pregnancies (15.4+/ 1.4 micromol/l, n=21) (P=0.049). CONCLUSIONS: : Increased cord serum NOx but unaltered sICAM-1 concentration in diabetic pregnancies indicates that maternal diabetes does not cause a general alteration in fetal endothelial function. The increase in cord serum and maternal serum NOx concentration in diabetic pregnancies may be due to abnormalities in insulin-induced nitric oxide release or to a diminished reactivity of the vasculature to the effects of nitric oxide. PMID- 15653197 TI - Insulin autoantibodies in cord blood from neonates with and without a sibling with type 1 diabetes--prevalence and characterization. AB - OBJECTIVE: Insulin autoantibodies (INSAAbs) are associated with the onset and progression of type 1 diabetes. Their presence in siblings of patients with type 1 diabetes precedes the development of overt diabetes. The precise time of INSAAbs onset and their relationship to diabetes triggering factors remains unknown. We sought to determine the prevalence and nature of INSAAbs in neonates with and without a sibling affected by type 1 diabetes. METHODS: One hundred and twenty four healthy non-diabetic neonates were studied; 12 had a sibling with type 1 diabetes and 112 had no first-degree relatives with the disease. Cord blood samples were collected from the neonates and their mothers at birth and again from the neonates whose initial sample was INSAAbs-positive at 3 months of age. INSAAbs titer was tested using a competitive radiobinding assay. To further characterize the nature of this binding capacity, the assay was repeated twice, before and after incubation with protein A or insulin-like growth factor (IGF-1). RESULTS: Specific insulin-binding activity was noted at birth in 8/12 neonates with a diabetic sibling (66%) and in 79/112 (70%) control newborns. None of the maternal samples from either group were INSAAb-positive. By age 3 months, INSAAb levels had normalized in 90% of the neonates in both groups. Protein A did not induce precipitation of bound insulin complexes, and IGF-1 failed to displace bound insulin from the cord blood. CONCLUSIONS: Although more than 70% of cord blood samples specifically bound insulin, this action is probably unrelated to the passive transfer of IgG antibodies during pregnancy or to cross-reactivity with the IGF-binding proteins. Furthermore, INSAAb cannot be a predictive sign for early development of type 1 diabetes in the first 3 months of age. PMID- 15653198 TI - In utero transplantation of human hematopoetic stem cells into fetal goats under B-type ultrasonographic scan: an experimental model for the study of potential prenatal therapy. AB - OBJECTIVE: Using fetal goats as animal models, to establish the methodology of in utero transplantation of human hematopoeitic stem cell (HSC) under B-scan ultrasonographic guidance for prenatal therapy. STUDY DESIGN: Human HSC were directly injected into the peritoneal cavities of the recipient fetal goats at 45 55 days of gestation (term: 145 days) under the guidance of B-type ultrasound scan. After birth, the peripheral blood was collected for fluorescence assisted cell sorting (FACS), quantitative real-time PCR and fluorescence in situ hybridization (FISH) to detect and analyze the presence of human cells in the recipients. RESULTS: The 32 recipients were born alive except one miscarriage. To test for the presence of human-goat chimeras, cells from 13 randomly selected transplanted goats were collected. FACS analyses showed the presence of human cells in all the transplanted goats tested. The average proportion of CD34+ cells and GPA+(glycophorin A) cells in the peripheral blood were 1.34 +/- 1.10% and 2.80 +/- 2.10%, respectively. No CD34+ or GPA+ cells were found in the non transplanted goats tested. The results of the quantitative real-time PCR in three engraftment goats were 1.2 x 10(4), 2.9 x 10(4), and 3.2 x 10(4) copies of human GPA DNA per mug of genomic DNA. FISH experiments showed that cells containing human specific alpha-satellite DNA sequence were present in the peripheral blood of the transplanted goats. CONCLUSIONS: The method described herein is safe and reliable, with low miscarriage risk and high chimerism rate. This approach may provide a promising animal model for potential prenatal treatment. PMID- 15653199 TI - Growth factor receptor-protein bound 2 (GRB2) upregulation in the placenta in preeclampsia implies a possible role for ras signalling. AB - OBJECTIVES: To screen for genes with altered expression in placentas from pregnancies complicated by preeclampsia. STUDY DESIGN: To corroborate gene expression profile of preeclamptic and normal placentas (ATLAS Clontech), by dot blot, Northern blot analysis and RT-PCR for growth factor receptor bound-protein 2 (GRB2), using immunohistochemistry to localize its expression in the placenta. RESULTS: Increased expression of GRB2 upregulated in the microarrays was found in preeclampsia by Dot blot and Northern blot analysis. RT-PCR performed with primers specific for GRB2 and its alternatively spliced isoform GRB3-3 showed that most of the cDNA represented in the array was GRB2. The protein was localized to the smooth muscle wall of stem vessels by immunohistochemistry. CONCLUSION: The ras signalling activated by placental receptor tyrosine kinases may play a role in the segmental thickening of the stem vascular wall in preeclamptic placentas, resulting in reduced blood flow to the developing fetus. PMID- 15653200 TI - Placental barrier breakage in preeclampsia: ultrastructural evidence. AB - OBJECTIVE: It is known that the placenta acts as an immunological barrier between the mother and fetal "graft" allowing two antigenically different organisms to tolerate one another. Preeclampsia may be considered as a fetal rejection consequent to severe damage at placental endothelial and syncytiotrophoblast level. In order to verify this placental barrier damage we undertook the present study by electron microscopy. STUDY DESIGN: 14 placentae from preeclaptic women, and the same number of placentae from healthy controls were examined. RESULTS: The results showed that endothelial cells from preeclamptic placentae express various and severe alterations, consisting of swollen and bulbous cytoplasm, degenerated inter-endothelial junctions with consequent crossing of fetal blood cells outside the vessels. CONCLUSIONS: These lesions could be the ultrastructural evidence of the placental barrier breakage leading to rejective reaction we presumed to be basis of preeclampsia. PMID- 15653201 TI - Effect of quercetine and glutathione on the level of superoxide dismutase, catalase, malonyldialdehyde, blood pressure and neonatal outcome in a rat model of pre-eclampsia induced by NG-nitro-L-arginine-methyl ester. AB - OBJECTIVE: To anticipate the impact of antioxidant use on lipid peroxidation products, free oxygen radical scavengers, blood pressure (BP), proteinuria and neonatal outcome (as seen in percentage survival, litter birth weight) in a rat model of pre-eclampsia induced by NG-nitro-L-arginine-methyl-ester (L-NAME), an inhibitor of nitric oxide synthase (NOS). MATERIAL AND METHODS: Female adult non pregnant Sprague-Dawley rats (n=40) with timed pregnancies were allocated into four groups according to medication they received on day 17 to term. Rats were randomised into a sham-treated group (group I, n=10) and groups treated with L NAME, 50 mg/day i.p., only (group II, n=10), L-NAME + quercetine, 10mg/kg i.p. (group III, n=10) and L-NAME + glutathione, 60 mg/kg i.p. (group IV, n=10). Blood levels of superoxide dismutase (SOD), catalase (CAT) and malonyldialdehyde (MDA) were assessed on day 22 of gestation. Intracardiac blood sampling and hysterotomy were performed on day 22 of gestation. Mean systolic BP (measured with a tail cuff device), level of proteinuria, total urine output, pups birth weight and percentages of live and of dead pups were recorded. RESULTS: Mean systolic BP and SOD, CAT and MDA levels were higher in rats infused with L-NAME than in the sham treated group. In group IV, SOD levels were lower than in group II (P <0.001). A linear positive correlation between BPs on day 20 and SOD levels (rp=0.39) was recorded, as were negative correlations between level of proteinuria and SOD levels (rp=-0.39) and between CAT and MDA levels (rp=-0.39). Birth weights were higher in the sham-treated group than in the other groups (P <0.001). Pups of hypertensive gravid rats treated with antioxidants had better survival rates than those of rats in group II and the sham-treated group (Chi-square=15.9, d.f.: 3, P <0.01).However, no correlation was detected between higher pup mortality rate and birth weight of pups. CONCLUSION: In this rat model of pre-eclampsia, adverse outcomes, such as proteinuria and high neonatal death rate, are reversed by exogenous antioxidant use, even though no significant improvement is detected in terms of BP and birth weight of pups. PMID- 15653202 TI - Association of nausea and vomiting in pregnancy with lower body mass index. AB - OBJECTIVE: To assess the effect of body mass index (BMI) on the tendency of pregnant women to vomit and on their general condition during pregnancy. STUDY DESIGN: We included patients in this study who presented to our gynecological emergency room and clinic during their first trimester of pregnancy. All women completed a questionnaire assessing obstetrical and physical characteristics, including gravidity, parity, gestational age, height and weight for BMI calculation. Women were requested to report the number of vomiting episodes per day and their general condition using a 1 to 10 scale (1-good, 10-bad). Patients were allocated to either a low-frequency group (0-1 vomiting episodes per day) or to a high-frequency group (2 and more vomiting episodes per day). RESULTS: Sixty one consecutive women were included in the study. The low frequency group consisted of 35 women and the high frequency group included the remaining 26 women. The BMI was significantly lower in the high frequency group as compared to the low frequency group (21.8 +/- 3.5 versus 24.4 +/- 4.7, respectively; P <0.05). Patients in the high frequency group also reported a worse general condition than those in the low frequency group (7.6 +/- 2.2 versus 3.5 +/- 2.1, respectively; P <0.05). CONCLUSION: Patients with higher frequency of vomiting episodes during the first trimester of pregnancy tend to have a lower BMI score and a worse general condition than patients with low frequency of vomiting episodes. PMID- 15653203 TI - Ethnicity and mode of delivery in 'low-risk' first-time mothers, East London, 1988-1997. AB - OBJECTIVE: To investigate the relationship between 'ethnicity' and mode of delivery in first-time mothers. DESIGN: Retrospective analysis of a computerised obstetric data set. SETTING: Three hospitals in London, 1988-1997. SUBJECTS: 27667 'low-risk' primiparous women. METHODS: Logistic regression of computerised clinical data. MAIN OUTCOMES: Odds of Caesarean section (CS) compared to other deliveries, and odds of forceps or ventouse delivery, compared to normal delivery, by assigned 'ethnic' group. RESULTS: 'African' (OR: 2.8; 95% confidence interval 2.4-3.1), 'West Indian' (OR: 1.7; 1.4-2.0), 'Bangladeshi' (OR: 2.1; 1.7 2.5), 'Indian' (OR: 1.6; 1.4-2.0) and 'Pakistani' (OR: 1.5; 1.1-2.0) women were at elevated risk of delivery by CS compared with 'Caucasian' women, after adjustment for maternal age, antenatal class attendance, late-booking, year of birth, hospital, intra-uterine growth retardation, birthweight, foetal sex, induction, and augmentation. 'Bangladeshi' (OR: 1.3; 1.1-1.5) and 'Indian' (OR: 1.3; 1.1-1.5) women had higher risks of delivery by forceps and ventouse compared with 'Caucasian' women after adjustment for maternal age, birthweight, hospital of birth, foetal sex, induction, augmentation and year of birth. CONCLUSIONS: This large study demonstrates strongly elevated risks of operative delivery in 'ethnic minority' mothers, after adjusting for important confounders. The study should be repeated in different populations, and more research is required to account for the differential risk of operative delivery. PMID- 15653204 TI - Hyperprolactinaemia does not always mean 'hyperprolactinaemia'! AB - The presence of macroprolactinaemia was investigated in the symptom-free hyperprolactinaemia cases to reveal its incidence. The serum prolactin (PRL) fractions in 21 female patients with hyperprolactinaemia without any clinical symptoms were analyzed with PEG (polyethylene glycol precipitation) procedure. In 14 of these 21 cases, hyperprolactinaemia was detected with a high fraction of macroprolactin. In cases with asymptomatic hyperprolactinaemia, it is more appropriate to employ the PEG precipitation test to detect the disorder. High levels of serum prolactin, do not essentially indicate the presence of a prolactinoma but may only indicate macroprolactinaemia. PMID- 15653205 TI - Combined use of metformin and ethinyl estradiol-cyproterone acetate in polycystic ovary syndrome. AB - OBJECTIVE: The aim of our study was to compare the effect of metformin applied independently to the effect of metformin used in combination with oral contraceptive containing ethinyl estradiol (EE) and cyproterone acetate (CA). STUDY DESIGN: This prospective, open clinical study lasted 6 months and included 30 women with PCOS, divided in two groups of 15 women each. Group 1 received 850 mg metformin twice a day and group 2 in which Diane35 was added to the same treatment only during the first 2 months of the investigation. Serum levels of testosterone, immune reactive insulin (IRI), sex hormone binding globulin (SHBG), dehydroepiandrosterone sulfate (DHEAS) and lipid metabolism parameters were measured before the treatment, on the third and sixth month. Free androgen index (FAI) and HOMA-IR were calculated. Body mass index (BMI) and waist-to-hip ratio (WHR) were assessed at baseline and at the end of therapy. RESULTS: Much better and faster decrease in the level of testosterone and free androgen index in group with combined use of metformin and Diane35 was established, without deterioration of the anthropometric and biochemical indices and insulin sensitivity. CONCLUSION: The combination of metformin with intermittent application of Diane35 is an appropriate alternative for the pathogenic influence and clinical improvement of the symptoms of androgen excess in cases with PCOS. PMID- 15653206 TI - The relation of serum and follicular fluid leptin and ovarian steroid levels in response to induction of ovulation in in vitro fertilization cycles. AB - OBJECTIVE: Leptin restores energy homeostasis and regulates appetite and body weight by communicating the energy status to the central nervous system. Although there is strong evidence that leptin affects reproduction, its role in the control of reproductive physiology is little understood. STUDY DESIGN: We studied leptin concentrations in the serum and follicular fluid of 65 women undergoing ovarian hyperstimulation for in vitro fertilization (IVF). Fasting serum samples were collected (1) on the 3rd day of the cycle before IVF and (2) at the time of oocyte retrieval. Serum concentrations of leptin, estradiol (E2), progesterone, FSH, LH, prolactin, total testosterone, DHEA-SO4, and TSH and follicular fluid concentrations of leptin, E2, and progesterone were measured. RESULTS: Serum leptin values increased on average by 66.4% over basal leptin levels on the day of oocyte pick-up (OPU). A positive correlation between leptin increase and body mass index was observed. The serum leptin level was similar to that in follicular fluid o the day of OPU. E2 levels increased 34.5-fold with controlled ovarian hyperstimulation. There was a negative correlation between the increase in leptin levels and in E2 levels (P <0.05) and in the number of oocytes harvested (P <0.05). CONCLUSION: The significant increase in serum leptin levels during controlled ovarian hyperstimulation indicates a possible role of leptin in reproductive function. The increase in leptin levels is negatively correlated with ovarian response evaluated by E2 production and number of oocytes retrieved. This might be due to the reduced ovarian response through negative feedback of leptin to the ovaries at high levels. PMID- 15653207 TI - Vascular endothelial growth factor and insulin-like growth factor-1 in polycystic ovary syndrome and their relation to ovarian blood flow. AB - OBJECTIVES: (1) To determine the serum levels of vascular endothelial growth factor (VEGF) and insulin-like growth factor-1 (IGF-1) in women with polycystic ovary syndrome (PCOS). (2) To study Doppler blood flow changes within the ovarian stroma of women with PCOS. (3) To evaluate the relationship between VEGF and IGF 1 and Doppler indices as well as hormonal profile. SETTING: Department of Obstetrics and Gynecology, and Department of Biochemistry, Faculty of Medicine, Assiut University, Egypt. DESIGN: Cross-sectional study. PATIENTS AND METHODS: Fifty infertile women with PCOS diagnosed by ultrasound examination and a history of oligomenorrhea, hirsutism and obesity were studied. Serum levels of vascular endothelial growth factor (VEGF), insulin-like growth factor-1 (IGF-1) and hormonal profile were measured. Doppler blood flow velocity waveforms analysis in both right and left intraovarian arteries was done. Twenty healthy and fertile women with regular menstrual cycles served as a comparison group were similarly studied at the third day of the cycle. RESULTS: The serum levels of VEGF, IGF-1 (4.79 +/- 0.91, 253.15 +/- 70.07 versus 2.39 +/- 0.42, 186.65 +/- 42.7) were significantly elevated (P <0.001 and P <0.01, respectively) in women with PCOS compared with control. Doppler indices, PI (2.01 +/- 0.77, 2.66 +/- 1.00 versus 2.98 +/- 0.77, 3.75 +/- 0.98) and RI (0.77 +/- 0.12, 0.82 +/- 0.09 versus 0.87 +/ 0.09, 0.89 +/- 0.09) in both right and left intraovarian vessels were significantly lower in the patients than controls. The VEGF and IGF-1 levels were negatively correlated with RI and PI in the uterine and intraovarian arteries. VEGF level was positively correlated with IGF-1 (r=0.41, P <0.05) in women with PCOS. CONCLUSIONS: Higher serum levels of VEGF and IGF-1 in PCOS women may be related to the increased vascularity that underlies the increased blood flow demonstrated by Doppler blood flow measurements in these women. PMID- 15653208 TI - Compliance with hormone replacement therapy in postmenopausal Sicilian women. AB - OBJECTIVE: To verify the compliance with hormone replacement therapy (HRT) over 2 years in a population of postmenopausal women in East Sicily. STUDY DESIGN: Patients starting hormonal therapy for the first time were enrolled in this study. A telephone survey was then conducted after 3, 6, 12 and 24 months and the reasons for any discontinuation were recorded. RESULTS: Of a total of 138 women who agreed to be enrolled in this prospective longitudinal study 72 were still taking the treatment after 1 year and only 56 at the end of the study, although only three patients reported that they had experienced no benefit. CONCLUSIONS: Type of work, surgical menopause and previous use of oral contraceptives were significantly statistically associated with better HRT compliance. Side effects and fear of breast cancer, which we maintain is exaggerated by the women and their family doctors, were the commonest reasons for early discontinuation of the hormonal treatment. PMID- 15653209 TI - Role of human papillomavirus (HPV) testing in the follow-up of patients after treatment for cervical precancerous lesions. AB - OBJECTIVE: To evaluate the role of human papillomavirus (HPV) testing in post treatment follow-up of patients after therapeutic excision of the cervix due to positive screening tests. STUDY DESIGN: A hospital-based retrospective analysis was performed with prospective collection of patient data of women screened for cervical cancer at a Gynecologic Outpatient Clinic. Patients after therapeutic excision due to positive screening results were identified and followed up with HPV testing and serial cytology. RESULTS: After 61 treatment for cervicalis intraepithelialis neoplasia (CIN), high-risk HPV infection was detected during the post-treatment follow-up at 18 cases (29.5%), 10 of them had persisting cytological atypia (positive predictive value (PPV): 56%), 5 developed CIN (PPV: 28%). When the HPV test was negative (43 patients) in the post-treatment period, neither CIN nor persisting cytological atypia developed (negative predictive value (NPV): 100%) during 1201 patient months (median 26 months). CONCLUSIONS: A negative HPV test eliminates the risk of recurrent disease after treatment for CIN. PMID- 15653210 TI - Oxytocin during myomectomy: a randomized study. AB - OBJECTIVE: To evaluate the influence of oxytocin on peroperative blood loss during myomectomy. STUDY DESIGN: From October 1998 to May 2002, 94 patients requiring surgical myomectomy by laparotomy or by the vaginal approach were enrolled in a randomized double blind study. Patients were randomized to two groups. In the first group (47 patients) oxytocin was administered during myomectomy and in the second group (47 patients) a placebo was used. The main outcome measures were peroperative blood loss and rates of blood transfusion and autotransfusion. RESULTS: Peroperative blood loss was no different between the oxytocin group and the placebo group (508 +/- 558 ml versus 451 +/- 336 ml; P=0.55). Rates of autotransfusion and blood transfusion were also similar in both groups. CONCLUSION: Administration of oxytocin during myomectomy did not reduce peroperative blood loss in our study. The benefits of using oxytocin to prevent hemorrhage during myomectomy seem to be limited. PMID- 15653211 TI - Aromatase inhibitor anastrozole for treating endometrial hyperplasia in obese postmenopausal women. AB - This is a preliminary study investigating the efficacy of aromatase inhibitor anastrozole in treating endometrial hyperplasia in obese postmenopausal women. We administered anastrozole for 12 months to 11 obese postmenopausal women with high operative risk, in order to treat endometrial hyperplasia (four simple, five complex and two atypical). Endometrial thickness in all cases and histology too in the two cases with atypia, revealed atrophical endometrium during treatment and additional mean follow-up of 10.2 months. The safety and tolerance profile was satisfactory. Anastrozole appears to be an interesting new modality for the treatment of endometrial hyperplasia in obese postmenopausal women. PMID- 15653212 TI - Magnetic resonance spectroscopy of premalignant and malignant endometrial disorders: a feasibility of in vivo study. AB - OBJECTIVE: To assess the potential clinical utility of in vivo proton magnetic resonance spectroscopy (MRS) in patients with various endometrial lesions. METHODS: Twelve patients with untreated uterine bleeding were included in this study. In-vivo proton MRS was performed using a 1.5 T MR scanner. The metabolite levels were classified into three classes in comparison with the noise level by visual examination. All the patients have endometrial biopsy. For each type of lesions, chemical compound were described. RESULTS: Pathological examination resulted in three endometrial cancer, two simple hyperplasias, one complex hyperplasia, two partial hydatiform mole, two proliferative endometrium and two secretory endometrium. In women with endometrial carcinoma, high choline and lipid signals were detected, whereas no creatine and no lactate signals were found. In women with endometrial hyperplasia, choline signal was detectable in all cases but one case showed lactate signal in addition to choline. In women with partial hydatidiform mole, the only detectable signal was choline. Lipid signals were detected in none of the cases with endometrial hyperplasia and partial hidatidiform mole. In women with either secretory or proliferative endometrium, choline and lactate signals were detectable in all cases but one case showed solely choline. Lipid signals were not detected in any of subjects with secretory or proliferative endometrium. CONCLUSION: The observed difference is the presence of lipid signal only in endometrial carcinoma. PMID- 15653213 TI - CYP1A1 alleles in female genital cancers in the Polish population. AB - OBJECTIVE: Cytochrome P4501A1 (CYP1A1) plays an important role in the bioactivation processes that transform aromatic hydrocarbons into ultimate carcinogens. Genetically determined differences in activity of this enzyme could modulate individual susceptibility to develop cancers. The role of CYP1A1 in metabolic pathway of estrogens suggests an influence on carcinogenic events in genital tissues. The aim of our study was to elucidate the possible role of CYP1A1 alleles in the pathogenesis of endometrial and ovarian cancers. STUDY DESIGN: We have compared CYP1A1 genotype frequency between genital (endometrial and ovarian) cancer groups and 212 healthy women. Cancer patients were stratified using FIGO classification and diagnoses were confirmed histopathologically. The analysis of CYP1A1 genotypes was performed using polymerase chain reaction/restriction fragment length polymorphism (PCR/RFLP) assay. RESULTS: We have observed higher frequency of heterozygotic genotypes containing mutation m4 (CYP1A1*1/*4) in cancer groups (5.1% in ovarian and 5.6% in endometrial cancer versus 1.9% in controls). CONCLUSION: The higher frequency of mutated CYP1A1*4 allele connected with lower frequency of CYP1A1*2A and CYP1A1*2B in endometrial and ovarian cancer groups indicates that differences in the metabolic activity of CYP1A1 could play a significant role in the pathogenesis of genital cancers. PMID- 15653214 TI - Transient diabetes insipidus during pregnancy: a clinical case and a review of the syndrome. PMID- 15653215 TI - Large carcinoid tumor of the appendix during pregnancy. AB - Diagnosis of acute appendicitis is considered more difficult in pregnant than in non-pregnant women. The appendicial carcinoid tumor is a lesion that most frequently is discovered incidentally in the removed organ. We report a rare case of an unruptured acute appendicitis during pregnancy, which proved to be a large carcinoid tumor. PMID- 15653217 TI - Compression sutures instead of emergency peripartum hysterectomy. PMID- 15653218 TI - Presentation with superior vena cava syndrome secondary to solitary lung metastasis mimicking primary lung cancer following resection of a uterine malignant mixed mullerian tumor: brief case report. PMID- 15653219 TI - Vesical and vaginal recurrent endometriosis in postmenopause following estrogen replacement therapy. PMID- 15653220 TI - Relationship between maternal pelvic parameters and idiopathic term breech presentation. PMID- 15653221 TI - Uterine artery ligation for the management of ruptured cornual ectopic pregnancy. PMID- 15653222 TI - Spontaneous pregnancy following multiple laparoscopic myomectomy in a sterile patient with unilateral ovarian agenesis. PMID- 15653223 TI - Specialist life--Lilo Mettler. PMID- 15653224 TI - Fetal arrhythmias: natural history and management. AB - The aim was to delineate the significance and natural history of fetal arrhythmias and provide information about their management. A cohort of 114 infants with fetal arrhythmias detected during prenatal ultrasound (US) screening were studied. All subjects underwent echocardiography and were treated as clinically indicated. Postnatal outcome was obtained in 100% of infants until 1 year of age. The incidence of fetal arrhythmias was 0.3%. Among the 87 fetuses with atrial extrasystoles, 2.3% developed supraventricular tachycardia (SVT) in utero. Of the 10 SVT cases, only five required antiarrhythmic therapy in utero with digoxin and propafenone, which successfully restored sinus rhythm in 100% of fetuses, both nonhydropic and hydropic. Sinus bradycardia was associated with structural anomalies in 5 of 6 patients and only 2 of 4 fetuses with atrioventricular block survived. It is concluded that prognosis is good for most fetal tachyarrhythmias, whereas it is less favorable for bradyarrhythmias. PMID- 15653225 TI - 3-D ultrasonographic imaging of the cerebral ventricular system in very low birth weight infants. AB - The purpose of the study was to assess reference ranges for lateral ventricular volume of very low birth weight (VLBW) infants using 3-D ultrasound (US). A total of 108 patients with birth weights < or =1500 g or mother's postmenstrual age < or =32 weeks were examined prospectively in a longitudinal study. Infants in conditions considered being potential confounders such as intraventricular haemorrhage (IVH) and periventricular leukomalacia (PVL) were not included in the calculations. Hence, 77 subjects remained for final statistical analysis. Mean postmenstrual age at birth was 194.5 (27 weeks and 5.5 days) +/- 14 SD days, mean birth weight was 972.5 +/- 236.3 SD g. Reference ranges for lateral ventricle volume were established from serial images. The exponential regression analyses revealed a weekly increase in volume of 6.3% (95% CI 4.4%-8.3%) and 6.6% (95% CI 4.7%-8.6%) in respect to the left and the right ventricle (p < 0.001). Postmenstrual age correlated significantly (p < or = 0.015) with ventricle volume. No significant association to head circumference could be determined. Establishment of reference values for the lateral ventricle volume of VLBW infants should facilitate application of 3-D US in routine diagnostics in neonatal intensive care units and detection of ventricular enlargement as a prediction of risk for poor neurodevelopmental outcome in high-risk cohorts. PMID- 15653226 TI - Dynamic three-dimensional visualization of the left ventricle by intracardiac echocardiography. AB - Cardiac function and hemodynamics are routinely evaluated during catheterization in patients with heart disease. Although intracardiac echocardiography (ICE) has been employed in guiding electrophysiology procedures, it has not been effectively used in assessing hemodynamics. We tested the utility of ICE in measuring left ventricular (LV) volume throughout the cardiac cycle. In four normal dogs (weight = 26 to 37 kg), a 10-F sheath was inserted through the femoral artery and placed inside the LV along its major axis. An ICE catheter (9 F, 9 MHz) was then inserted through the sheath into the LV. The ICE catheter was pulled back inside the sheath in 1-mm intervals starting from the apex, and 2-D tomographic images were continuously acquired while gating to respiration. Subsequently, the ICE catheter was replaced by a conductance catheter to measure single-beat volume signals. Stroke volume was determined by thermodilution for validation. All measurements were made in each dog while pacing the atrium at two different cycle lengths (range = 300 to 500 ms). The endocardial boundary was digitized from the ICE images throughout the cardiac cycle and LV volume was computed by integrating multiple segments along the major axis (range = 55 to 70 mm). We found that ICE accurately reconstructed LV 3-D anatomy. Stroke volume by ICE was in excellent agreement with thermodilution (error = 3.8 +/- 3.0%, r = 0.99, n = 8) and was highly reproducible. Morphology of LV volume signals correlated well with corresponding instantaneous volume signals derived by conductance (r = 0.93, n = 8). In conclusion, ICE accurately reconstructs LV anatomy and volume throughout the cardiac cycle in the normal heart. This approach could facilitate interventional diagnostic and therapeutic procedures. PMID- 15653227 TI - Evaluation of cardiac function by tissue Doppler echocardiography: hemodynamic determinants and clinical application. AB - A total of 32 patients without regional wall motion abnormality of the left ventricle underwent sequential tissue Doppler echocardiography and cardiac catheterization. Peak velocities of systolic (Sa), early diastolic (Ea), and late diastolic (Aa) motion of the mitral annulus were measured. Normal references for Sa, Ea and Aa were obtained from 138 volunteers. Indices of left ventricular (LV) systolic and diastolic function were evaluated using high-fidelity LV pressure and volume signals. By multivariate analysis, Sa, Ea and As were significantly and independently related to the maximum of the first derivative of pressure over time (dP/dt(max)), LV relaxation time constant (tau), and LV ejection fraction (EF), respectively. Using the fifth percentiles of the age-stratified normal references as cut-offs, low Sa, low Ea and low Aa identified declined dP/dt(max), prolonged tau and reduced EF, respectively, with good sensitivities and specificities. In conclusion, mitral annulus velocities by tissue Doppler echocardiography can be used to identify patients with declined dP/dt(max), prolonged tau and reduced EF. PMID- 15653228 TI - Contrast media induces hypoperfusion in kidneys with ureteral stone: Doppler US study. AB - We aimed to assess the hemodynamic effects of nonionic low-osmolarity contrast media (CM) on kidneys with ureteral stone (KUS). A total of 16 patients with KUS (group A) and 25 control patients (group B) were evaluated with Doppler ultrasound (US) bilaterally for main (MRA), interlobar (ILA) and arcuate renal arteries (ARA), before and 15 min after CM for IV pyelography. Peak systolic (PSV) and end diastolic velocities (EDV) and resistance index (RI) were measured. Data were analyzed by ANOVA and t-test. In group A, CM induced increase in PSV of MRA of contralateral kidney (p = 0.021) and decrease in PSV of ILA (p = 0.024), decrease in PSV and EDV and increase in RI of ARA of KUS (p = 0.010, 0.005, 0.027, respectively). CM induced hypoperfusion in KUS and compensatory changes in contralateral kidneys, similar to the effects of diuresis or mannitol. We conclude that the effect of CM on KUS is related to its osmolar load. PMID- 15653229 TI - Low back pain, the stiffness of the sacroiliac joint: a new method using ultrasound. AB - Abnormal biomechanical properties of the sacroiliac joints are believed to be related to low back and pelvic pain. Presently, physiotherapists judge the condition of the sacroiliac joints by function and provocation tests, and palpation. No objective measuring device is available. Research is ongoing to identify the biomechanical properties of the sacroiliac joints from the dynamic behaviour of the pelvic bones. A new concept based on ultrasound (US) for the measurement of bone vibration is under investigation. The objective of this study was to validate this concept on a physical model and to assess the applicability in vivo. A model consisting of a piezo shaker covered by a layer of US transmission gel (representing bone and soft tissue) has been used. A packet of US detection signals is directed onto the shaker and correlation-based processing is used to estimate the difference in time-of-flight of their echoes. These variations of time are used to compute the displacement of the shaker at each pulse reflection. To assess the validity of our US technique, we compared the obtained measurements with the readings of the built-in strain gauge sensor. The experimental procedure has been tested on a volunteer where low-frequency excitation was provided through the ilium and vibration detected on the sacrum and ilia. The results demonstrated that the correlation-based approach is capable of reproducing the piezo shaker displacements with high accuracy (+/- 7%). Vibration amplitudes from 0.25 microm to 3 microm could be measured. The US technique was able to detect bone vibration in vivo. In conclusion, the principle based on US waves can be used to develop a new measurement tool, instrumental in studying the relation between the biomechanical properties of the sacroiliac joints and low back pain. PMID- 15653230 TI - Strain processing of intraoperative ultrasound images of brain tumours: initial results. AB - The purpose of the study was to investigate a method for strain calculation and its ability to discriminate between brain tumour and normal brain. During surgery of a low-grade astrocytoma and a metastasis, we acquired ultrasound (US) radiofrequency (RF) data with a hand-held probe at the dura mater. Using cross correlation and phase-sensitive processing, we quantified the tissue displacements between consecutive US images and, subsequently, the local strain. In the elastograms, the tumour lesions were associated with lower strain levels than those found in the surrounding normal tissue. For both investigated cases, the strain images showed good agreement with the B-mode images. However, the results also indicated that the tumour interpretation might be different in the two modalities. An important finding was that the tissue motion caused by arterial pulsation is sufficient for generating elastograms. Requiring no specialised equipment or changes to acquisition procedures, strain data can be obtained as easily as conventional US imaging. PMID- 15653231 TI - Image-based cardiac gating for three-dimensional intravascular ultrasound imaging. AB - Three-dimensional (3-D) intravascular ultrasound (US), or IVUS, provides valuable insight into the tissue characteristics of the coronary wall and plaque composition. However, artefacts due to cardiac motion and vessel wall pulsation limit the accuracy and variability of coronary lumen and plaque volume measurement in 3-D IVUS images. ECG-gated image acquisition can reduce these artefacts but it requires recording the ECG signal and may increase image acquisition time. The goal of our study was to reconstruct a 3-D IVUS image with negligible cardiac motion and vessel pulsation artefacts, by developing an image based gating method to track 2-D IVUS images over the cardiac cycle. Our approach involved selecting 2-D IVUS images belonging to the same cardiac phase from an asynchronously-acquired series, by tracking the changing lumen contour over the cardiac cycle. The algorithm was tested with IVUS images of a custom-built coronary vessel phantom and with patient images. The artefact reduction achieved using the image-gating approach was > 86% in the in vitro images and > 80% in the in vivo images in our study. Our study shows that image-based gating of IVUS images provides a useful method for accurate reconstruction of 3-D IVUS images with reduced cardiac motion artefact. PMID- 15653232 TI - Curvature affects Doppler investigation of vessels: implications for clinical practice. AB - In clinical practice, blood velocity estimations from Doppler examination of curved vascular segments are normally different from those of nearby straight segments. The observed "accelerations," sometimes considered as a sort of stochastic disturbances, can actually be related to very specific physical effects due to vessel curvature (i.e., the development of nonaxial velocity [NAV] components) and the spreading of the axial velocity direction in the Doppler sample volume with respect to the insonation axis. The relevant phenomena and their dependence on the radius of curvature of the vessels and on the insonation angle are investigated with a beam-vessel geometry as close as possible to clinical setting, with the simplifying assumptions of steady flow, mild vessel curvature, uniform ultrasonic beam and complete vessel insonation. The insonation angles that minimize the errors are provided on the basis of the study results. PMID- 15653233 TI - Comparison of extended field of view and dual image ultrasound techniques: accuracy and reliability of distance measurements in phantom study. AB - This study was undertaken to investigate and compare the accuracy and reliability of dual image and extended field-of-view (EFOV) ultrasound (US) techniques in distance measurements using acoustic phantoms. Ten tissue phantoms were constructed and were scanned twice with an interval of 3 days by two operators. Measurements of various known distance (ranging from 4.6 to 7.2 cm) in the phantoms were made with dual image and EFOV US. Results showed that both dual image and EFOV US have a high accuracy and reliability in distance measurements, with the EFOV US (r = 0.997 to 0.998, reproducibility = 99.8%, repeatability = 98.2 to 99.8%) being slightly more accurate and reliable than dual image US (r = 0.948 to 0.981, reproducibility = 94.6%, repeatability = 89.6 to 97.9%). EFOV US has a higher accuracy and reliability than dual image US in distance measurements. However, the dual image US is a useful alternative with a high accuracy and reliability when EFOV US is not available. PMID- 15653234 TI - On the potential of the Lagrangian speckle model estimator to characterize atherosclerotic plaques in endovascular elastography: in vitro experiments using an excised human carotid artery. AB - Endovascular ultrasound (US) elastography (EVE) was introduced to supplement endovascular US echograms in the assessment of vessel lesions and for endovascular therapy planning. Indeed, changes in the vascular tissue stiffness are characteristic of vessel wall pathologies and EVE appears as a very appropriate imaging technique to outline the elastic properties of vessel walls. Recently, a model-based approach was proposed to assess tissue motion in EVE. It specifically consists of a nonlinear minimization algorithm that was adapted to speckle motion estimation. Regarding the theoretical framework, such an approach considers the speckle as a material property; this assumption then led to the derivation of the optical flow equations, which were suitably combined with the Lagrangian speckle model estimator to provide the full 2-D polar strain tensor. In this study, the proposed algorithm was validated in vitro using a fresh excised human carotid artery. The experimental setup consisted of a cardiovascular imaging system (CVIS) US scanner, working with a 30-MHz mechanical rotating single-element transducer, a digital oscilloscope and a pressuring system. A sequence of radiofrequency (RF) images was collected while incrementally adjusting the intraluminal static pressure steps. The results showed the potential of this 2-D algorithm to characterize and to distinguish an atherosclerotic plaque from the normal vascular tissue. Namely, the geometry as well as some mechanical characteristics of the detected plaque were in good agreement with histology. The results also suggested that there might exist a range of intraluminal pressures for which plaque detectability is optimal. PMID- 15653235 TI - Comparison of different mathematical models to analyze diminution kinetics of ultrasound contrast enhancement in a flow phantom. AB - Ultrasound (US) energy leads to intensity- and frequency-dependent destruction of US contrast agent (UCA) microbubbles. When applying repeated US pulses, this phenomenon can be detected as contrast diminution over time. Contrast diminution kinetics depend on the replenishment of UCA into the sample volume. Thus, it is related to organ perfusion. To analyze the contrast diminution kinetics following pulsed harmonic US application (SONOS 5500, 1.8-3.6 MHz, MI: 1.6, frame rates: 2, 4, and 6.67 Hz), we performed an in vitro study using SonoVue continuous infusion. Seven flow rates (4.5, 9, 13.5, 18, 22.5, 27 and 36 mL/min) were tested. Based on our results, three mathematical models (linear intensity decrease, exponential decay, and an exponential destruction/reperfusion model) describing diminution kinetics were compared. In 113 (89.7%) of 126 trials, a signal decrease was observed after US application. At higher flow rates (18 to 36 mL/min), curve fitting was not possible for the exponential models. For the linear model, intensity decrease depended significantly on the flow rate (p < or = 0.005, n = 7). A logistic model was fitted to the data, defining the slope in the dynamic range of quasilinear dependence for the different frame rates, as well as the inflection point: The higher the frame rate, the higher the flow rate at the point of inflection. For the exponential model, the contrast half-life was dependent on the flow rate (r = 0.95, p = 0.03, n = 6) only at the highest frame rate (6.67 Hz). The perfusion coefficient derived from the destruction/reperfusion model was not significantly related to the flow rate. In conclusion, the linear intensity decrease correlates well with the flow rate (i.e., flow velocity) and defines optimum frame rates for diminution imaging at different flow velocities. The exponential models, which required curve-fitting procedures, were determined to be inappropriate to describe flow in our phantom. PMID- 15653236 TI - Advantages in using multifrequency excitation of contrast microbubbles for enhancing echo particle image velocimetry techniques: initial numerical studies using rectangular and triangular waves. AB - Accurate measurement of velocity profiles, multiple velocity vectors and local shear stress in arteries is very important for a variety of cardiovascular diseases. We have recently developed an ultrasound based velocimetry technique, termed echo particle image velocimetry (echo PIV). This method takes advantage of the nonlinear backscatter characteristics of ultrasound contrast microbubbles when exposed to certain ultrasonic forcing conditions. Preliminary in vitro, animal and clinical studies have shown significant promise of this method for measuring multiple velocity components with good temporal (up to 2 ms) and spatial (<1 mm) resolution. However, there is still difficulty in maximizing the nonlinearity of bubble backscatter using conventional Gaussian-pulse excitation techniques because: (1) significant harmonic components may not be produced at modest pressure amplitudes; and (2) the higher incident pressure amplitudes required to induce nonlinear behavior may cause bubble destruction. We present here a potential solution to this problem through the use of multifrequency excitation, where rectangular and triangular pulses with four harmonics are used to drive the bubble. The nonlinear behavior of the microbubble, as well as fragility and backscatter, were studied through numerical modeling via a modified Rayleigh-Plesset equation. Results show that the rectangular wave is effective in improving the visibility of microbubbles, with effective scattering cross-section area significantly higher (up to 35 times) than the widely-used Gaussian waveform. However, velocity and acceleration analysis of the bubble wall shows that the rectangular wave may threaten bubble stability. Due to lower wall velocity and acceleration, the triangular wave should decrease the potential for bubble destruction yet maintain relatively high second harmonic backscatter components. The impact of higher harmonics was studied by examining backscatter differences from incident rectangular and triangular pulses with four and two harmonics. Results indicate that a two-frequency excitation (which may be easier to implement practically) may be sufficient to induce nonlinear behavior of the microbubbles at modest incident pressures. These predictions provide support for the use of multifrequency driving to enhance echo PIV applications. PMID- 15653237 TI - In vitro ablation of cardiac valves using high-intensity focused ultrasound. AB - The purpose of this study was to evaluate the possibility of using high-intensity focused ultrasound (US), or HIFU, to create lesions in cardiac valves in vitro. Calf mitral valves and aortic valves were examined. Focused US energy was applied with an operating frequency of 4.67 MHz at a nominal acoustic power of 58 W for 0.2, 0.3 and 0.4 s at 4-s intervals. Mitral valve perforation was achieved with 20.8 +/- 3.7 exposures of 0.2 s, 15.4 +/- 2.1 exposures of 0.3 s or 11.2 +/- 2.3 exposures of 0.4 s. Aortic valve perforation was achieved with 13.3 +/- 2.4 exposures of 0.2 s, 10.3 +/- 2.2 exposures of 0.3 s or 8.4 +/- 1.8 exposures of 0.4 s. The mean diameter of the perforated area was 1.09 +/- 0.11 mm. The lesions were slightly discolored and coagulation of tissue around the perforation was observed. HIFU was successful in perforating cardiac valves. With further refinement, HIFU may prove useful for valvulotomy or valvuloplasty. PMID- 15653238 TI - Antibacterial effects of extracorporeal shock waves. AB - Despite considerable knowledge about effects of extracorporeal shock-wave therapy (ESWT) on eukaryotic tissues, only little data are available concerning their effect on prokaryotic microorganisms. The objective of the present study was to determine the bactericidal activity as a function of energy flux density and shock-wave impulse number. Standardised suspensions of Staphylococcus aureus ATCC 25923 were exposed to different impulse numbers of shock waves with an energy flux density (ED) up to 0.96 mJ mm(-2) (2 Hz). Subsequently, viable bacteria were quantified by culture and compared with an untreated control. After applying 4000 impulses, a significant bactericidal effect was observed with a threshold ED of 0.59 mJ mm(-2) (p < 0.05). A threshold impulse number of more than 1000 impulses was necessary to reduce bacterial growth (p < 0.05). Further elevation of energy and impulse number exponentially increased bacterial killing. ESWT proved to exert significant antibacterial effect in an energy-dependent manner. Certain types of difficult-to-treat infections could offer new applications for ESWT. PMID- 15653239 TI - The alteration of protein profile of Walker 256 carinosarcoma cells during the apoptotic process induced by ultrasound. AB - The objective of this study was to investigate the alteration of the protein profile in cells after sonication and to identify the key proteins involved in the process of cell apoptosis. Walker 256 carinosarcoma cells were exposed to focused ultrasound (US) at the intensity of 2.0, 7.0, 10.2, 14.2 and 17.0 W/cm2 (I(spta)) for 10 min in vitro and the morphologic and functional changes of the cells were detected by hematoxylin & eosin staining and flow cytometry, with double staining of fluorescein isothiocyanate (FITC)-labeled Annexin V/propidium iodide (PI). The protein compositions in the cells after sonication were detected by 2-D SDS polyacrylamide gel electrophoresis. Our results showed that apoptosis of Walker 256 carinosarcoma cells could be induced by US. The percentage of early apoptosis and secondary necrosis increased with increasing intensity of US irradiation. Comparing with the protein patterns of cells before sonication, it was found that around 420 new protein spots were present in the gel after sonication. Among them, Hsp60 and Bcl-2 like protein 13 were found to be involved in the process of cell apoptosis and US-induced apoptosis of the cells was probably performed through the pathway of promoting the activation of caspase-3. PMID- 15653240 TI - Effect of ultrasound on nucleated erythrocytes. AB - In this experimental study, carp erythrocytes were used as nucleated cell models to test the hypothesis that ultrasound (US) exposure can cause the change in plasma membranes and hemoglobin. To identify target cell damage, we studied hemolysis, osmotic fragility, lipid peroxidation of red blood cells and oxidation of hemoglobin in the erythrocytes. The erythrocytes were exposed to 1 MHz continuous-wave US at the intensities of 0.61 to 2.44 W/cm2 for 5 min. These results showed that US sonication at the intensities of 1.90 and 2.44 W/cm2 led to an increase in the degree of hemolysis and, at the highest used intensity (2.44 W/cm2), an increase in osmotic fragility. Ultrasound in all the used intensities induced an increase in lipid peroxidation. The results obtained showed that the level of methemoglobin has a tendency to increase after US exposure. Our results suggest that the changes in biomembranes can be due to inertial cavitation induced by US, but we cannot say which stage of the inertial event causes the damage. PMID- 15653241 TI - Assessing socio-economic differences in patient-provider communication. PMID- 15653242 TI - Socio-economic status of the patient and doctor-patient communication: does it make a difference? AB - This systematic review, in which 12 original research papers and meta-analyses were included, explored whether patients' socio-economic status influences doctor patient communication. Results show that patients from lower social classes receive less positive socio-emotional utterances and a more directive and less participatory consulting style, characterised by significantly less information giving, less directions and less socio-emotional and partnership building utterances from their doctor. Doctors' communicative style is influenced by the way patients communicate: patients from higher social classes communicate more actively and show more affective expressiveness, eliciting more information from their doctor. Patients from lower social classes are often disadvantaged because of the doctor's misperception of their desire and need for information and their ability to take part in the care process. A more effective communication could be established by both doctors and patients through doctors' awareness of the contextual communicative differences and empowering patients to express concerns and preferences. PMID- 15653243 TI - University hospital physicians' attitudes and practices for smoking cessation counseling in Malatya, Turkey. AB - The aim of the study is to determine attitudes and self-reported practices performance for smoking cessation counseling of the physicians working at a university hospital in Malatya, Turkey. All physicians who were providing health care to adult patients in 19 outpatient clinics at the hospital were administered a self-reported questionnaire. Of the physicians, 26.5% reported that they were always asking about their patients' smoking history and 22.6% were always advising their smoker patients to quit. Men non-smoker physicians more often practiced counseling than men smoker physicians. Negative attitudes of physicians about smoking cessation counseling negatively effected their practices. Negative attitudes were significantly higher among men smoker physicians than non-smokers but attitudes did not differ among female smoker and non-smoker physicians. The findings showed that smoking cessation counseling was rarely practiced by physicians and physicians' practices differed by their smoking behavior, departments and attitudes towards smoking cessation. PMID- 15653244 TI - The general public's information needs and perceptions regarding hereditary cancer: an application of the Integrated Change Model. AB - The Integrated Change Model (the I-Change Model) was used to analyse the general public's need and perceptions concerning receiving information on the role of hereditary factors with regard to cancer. The results from a study in 457 Dutch adults showed that 25% correctly indicated the types of cancer where hereditary factors can play a role. Respondents, however, overestimated the role of hereditary factors causing breast cancer. Recognition of warning signs was low, as was the recognition of inheritance patterns. Participants wanted to know the types of cancer with hereditary aspects, how to recognise hereditary cancer in the family, personal risks and the steps to be taken when hereditary predisposition is suspected. The most popular information channels mentioned were leaflets, the general practitioner, and the Internet. Respondents interested in receiving information on heredity and cancer were more often female, had had experiences with hereditary diseases, had more knowledge, perceived more advantages, encountered more social support in seeking information, and had higher levels of self-efficacy. Education should outline the most important facts about hereditary cancer, how to get support, and create realistic expectations of the impact of genetic factors. PMID- 15653245 TI - Measuring adaptations of motivational interviewing: the development and validation of the behavior change counseling index (BECCI). AB - One of the most common challenges faced by health professionals is encouraging patients to change their behavior to improve their health. This paper reports the development of a checklist, the behavior change counseling index (BECCI). This aims to measure practitioner competence in behavior change counseling (BCC), an adaptation of motivational interviewing suitable for brief consultations in healthcare settings. The checklist has demonstrated acceptable levels of validity, reliability and responsiveness, and aims to assist trainers and researchers in assessing change in practitioner behavior before, during and after training in BCC. BECCI will also provide valuable information about the standard of BCC that practitioners were trained to deliver in studies of BCC as an intervention. PMID- 15653246 TI - Information priorities of Italian early-stage prostate cancer patients and of their health-care professionals. AB - The study was designed to compare the information priorities of Italian early stage prostate cancer patients to those of their health-care professionals. Doctors (urologists and oncologists), nurses, radiation therapy technologist (RTs), and patients treated with radiotherapy in Northern Italy were surveyed. Respondents rated the importance of addressing each of 78 questions as: essential, important, or superfluous. We compared priorities between pairs of groups by correlating the percentage of each group that rated each question (a) essential and (b) superfluous. We assessed within-group agreement by identifying questions that >50% of one response. Fifty-nine doctors, 53 nurses, and 45 RTs (overall 70% response rate) and 50 patients (91% response rate) participated. All correlations between professional groups were high (all >0.74). All professional group-patient correlations were significant but much lower than those among the professionals. More importantly, there was considerable variation within each group: within each group, almost all questions were rated essential by some members but less than seven of the 78 questions were rated so by half (or more) of its members. In addition, almost all questions were also rated superfluous by some members of each profession. We concluded that the professional groups appear to agree with one another on questions essential and those superfluous to address more than they agree with a group of their patients. More importantly, there is considerable disagreement within each group. The results imply that the professionals cannot assume that their own information priorities are the same as those of their patients. PMID- 15653247 TI - Development and validation of a Breast Cancer Genetic Counseling Knowledge Questionnaire. AB - Women who undergo genetic counseling concerning their increased risk of developing breast cancer confront large quantities of complex information in a short period of time. Clinical reports have suggested that many women may not retain what they learned during counseling. A validated questionnaire to measure their knowledge, however, is lacking. In this study, we describe the development and validation of a questionnaire to assess knowledge of information typically included in genetic counseling for breast cancer. Items were empirically derived from detailed content analyses of actual genetic counseling sessions. The instrument's content validity was high, as evidenced by high levels of independent interrater agreement (0.93) on items. Subsequent data reduction and confirmatory factor analytic techniques yielded a highly reliable (alpha = 0.92) 27-item Breast Cancer Genetic Counseling Knowledge Questionnaire (BGKQ). Direct comparison of this questionnaire to a scale previously developed in the literature (BCHK; [Breast Cancer Res. Treat. 53 (1999) 69]) supported the utility of the new questionnaire for evaluation of knowledge after counseling. Compared to non-counseled groups (n = 45), women who had undergone genetic counseling (n = 28) scored significantly higher (P < 0.0001) on the BGKQ, but not on the other questionnaire, establishing the BGKQ's criterion validity. The BGKQ may, thus, provide a useful clinical and research tool for assessing knowledge of information provided during genetic counseling and exploring the potential impact of distress on knowledge, as well as the impact of knowledge on screening behaviors. PMID- 15653248 TI - Patient education materials for mental health problems in family practice: does location matter? AB - INTRODUCTION: This study examines how frequently family physicians display printed educational materials about mental health problems in their practices and where these materials should be located for optimal effect. METHODS: A cross sectional observational study of pamphlet display practices in 13 family physicians' offices in Hamilton, Ontario, Canada was followed by an intervention which placed selected mental health educational materials in waiting rooms and examining rooms, and monitored the pick up rate from each location by patients. MAIN RESULTS: The study found that few mental health pamphlets were displayed by the participating physicians, that when a range of these pamphlets was made available, pamphlets on mood disorders were the most popular, and that significantly more pamphlets were picked up from examining rooms than from waiting rooms. CONCLUSIONS: We conclude that patients are interested in having access to printed materials about mental health problems, and that the optimal location is in display racks in examining rooms. PMID- 15653249 TI - Perceptions about breast cancer among African American women: do selected educational materials challenge them? AB - Despite the availability of factual information about breast cancer, there continues to be an abundance of misperceptions about the disease. This study, guided by the Patient/Provider/System Model for cancer screening, describes perceptions about breast cancer among African American women (N = 179) at primary care centers. Data were collected using the Breast Cancer Perceptions and Knowledge Survey and a demographic questionnaire. Breast cancer pamphlets available at the centers were evaluated (readability, extent they challenged misperceptions). The average age of the women was 34 years with an average educational level of 12 years. A number of misperceptions were prevalent. The majority viewed breast self-examination as a form of early detection and some viewed pain as an indicator of cancer. Pamphlets did not explicitly challenge the misperceptions and the SMOG reading level was high. Intervention studies are needed to identify the effective methods to challenge and correct misperceptions about breast cancer for these women. PMID- 15653250 TI - Understanding the narratives of people who live with medically unexplained illness. AB - This paper reports on a qualitative study, which explores the narratives of patients, who live with medically unexplained symptoms (MUS) and who have not secured a diagnostic label. Interviews were undertaken with 18 participants (5 men and 13 women) who attended a neurology outpatients department in the UK. Three features of the patients' narratives identified are: the 'chaotic' structure of their illness narratives; concern that symptoms may be 'all in the mind'; and their status as 'medical orphans'. All the patients acknowledge that diagnosis is difficult and accept that a medical explanation will invariably be possible. However, they are more concerned to secure some form of ongoing medical and social support. An understanding of both the structure as well as the content of patients' narratives of undiagnosed illness may contribute to the development of more effective and sensitive patient centred care. PMID- 15653251 TI - Actual and desired information provision after a stroke. AB - Stroke patients and caregivers have a substantial information need. The study investigates how information was actually provided to stroke patients and caregivers and how they prefer to be informed. The GP, neurologist and physiotherapist are both the actual and desired information providers. The actual and desired information correspond in terms of content, frequency, and method of presentation. However, patients and caregivers prefer to receive information within 24 h and to be informed about, and be given, relevant written information. The information given by the various professional stroke care-providers could be better co-ordinated. The role of the GP as an information provider lagged quite a long way behind. Recommendations for the provision of an improved information system is given. Most of the subjects are relatively young male patients with few disabilities and healthy caregivers. More attention should be paid to encouraging patients and caregivers to actively seek information to supplement the information given by professional stroke care-providers. PMID- 15653252 TI - Client feedback on physiotherapy counselling in primary health care. AB - The extent of rehabilitation services is adequate, but little feedback is available on these services and service users. This paper is based on a client feedback survey of physiotherapy services conducted in Spring 2000. The purpose of the paper is to compare patients' perceptions of the physiotherapy instruction they received in a private outpatient facility and in a public physiotherapy facility. The results of the study showed that physiotherapy clients were satisfied with the therapy and instruction they received. They felt they received an adequate amount of instructions and advice. The rehabilitee's strong commitment to care, high motivation and a support network are important to independent rehabilitation in the future. As for the point of contact, the majority of clients in the private physiotherapy facility were gainfully employed while the health centre had almost as many retired clients. Client instruction was not dependent on the point of contact. PMID- 15653253 TI - Women's experiences of information, psychological distress and worry after treatment for gynaecological cancer. AB - Sensitive, appropriate patient information is considered to be an important element in the psychological support of patients. Specialist nurses are seen to have a key responsibility for this work. With regard to gynaecological cancer, evidence suggests that women do not get optimum psychological care. This study set out to explore women's experiences of information, psychological distress and worry after treatment for gynaecological cancer. The study was a survey (not an RCT) and 70 patients from two specialist gynaecological oncology centres were interviewed at the time of diagnosis/initial treatment and again at 6 months. The semi-structured schedule included recognised instruments to assess; sources of information, concerns, and psychological distress. Both initially and at 6 months there was evidence of a considerable burden of worry; over half the women had four or more significant concerns related to their illness experience. However, women who had initial support from a clinical nurse specialist at the time of diagnosis experienced a clinically significant reduction in their level of psychological distress 6 months from diagnosis. Hospital linked professional sources of information were well used at the time of diagnosis, but by 6 months many patients were using non-professional sources such as television, magazines and newspapers. This study suggests that support from a clinical nurse specialist may be able to assist psychological recovery. However, to be effective in this area nurses should be skilled and willing to assess the individual's need for help with information, and managing their worry. PMID- 15653254 TI - Evaluation of a counseling training program for nursing staff. AB - One of the essential issues in nurses' daily work is interaction with patients, patients' families, and co-workers. However, in the Spanish academic programs for Nursing Schools, social interaction skills do not receive adequate attention and nurses often report communication problems. In order to diminish these difficulties and to train nursing staff to better manage interaction, an intensive counseling training program was designed and implemented in a General University Hospital. The main aim of this study was to evaluate the effects of a counseling training program and assessing the evolution of difficulties that professionals perceived in their work in three different periods: before the training, after the training, and at follow-up, 2 months after the program was delivered. According to the results, we can maintain the hypothesis that the counseling training program reduces perceived interaction difficulties in nursing staff. Consequently, we can expect a further improvement in the interaction performance with patients and their families after the training. These findings suggest that counseling training has to be taken into account to improve quality of care in health care providers, and it may also help to prevent professional burnout by increasing competence level at minimum personal cost. PMID- 15653255 TI - Long-term efficacy of a checklist to improve patient education in cardiology. AB - In a randomised controlled trial a Frequently Asked Questions (FAQ) checklist intended to prepare coronary artery disease (CAD) outpatients for a medical check up visit at the cardiologist was evaluated. The checklist was mailed to patients in preparation to their visits after 1, 4 and 10 months following patients' discharge from hospitalisation for CAD. It was hypothesised that the intervention would result in lower state anxiety, better patient-doctor communication, more knowledge of CAD and greater patient satisfaction, while it would not result in longer visits. Repeated measurements analyses of covariance showed that experimental patients (N = 46) were less anxious before the first visit. This visit was shorter than in the controls, though the third visit was longer. Control patients (N = 59) showed more CAD knowledge than experimental patients at the first and third visit. Experimental patients found the checklist useful, though its value diminished at subsequent visits. Using the checklist thus decreased anxiety prior to the first visit and the duration of that visit, while negatively affecting knowledge. No conclusions about long-term effects could be drawn, due to the likelihood of type II and type III errors. Process evaluation indicated that the approach used is not sufficiently stimulating for patients to use as a preparation to every visit. PMID- 15653256 TI - Suicidal risk during treatment with clozapine: a meta-analysis. AB - OBJECTIVE: Suicide remains a major cause of premature mortality among patients with schizophrenia. Evidence of reduced suicidal risk with available psychiatric treatments is limited, but emerging data suggest such an effect of clozapine in chronically psychotic patients, leading us to compile the reported evidence. METHOD: We searched for published studies with contrasting rates of suicides or attempts by psychotic patients treated with clozapine vs. other agents. RESULTS: Among six such studies, random-effects meta-analysis indicated a substantially lower overall risk of suicidal behaviors with clozapine vs. other treatments (risk-ratio 3.3; 95% confidence interval [CI] 1.7-6.3; p<0.0001). For completed suicides, the risk ratio (RR) was 2.9 ([CI 1.5-5.7]; p=0.002). CONCLUSION: Long term treatment with clozapine was associated with three-fold overall reduction of risk of suicidal behaviors. However, available findings are quantitatively inconsistent, well-designed studies remain rare, and the only randomized trial did not find reduced risk of completed suicide. Additional randomized comparisons among modern treatments for psychotic disorders are required to clarify their impact on mortality. PMID- 15653257 TI - Chronic phencyclidine administration induces schizophrenia-like changes in N acetylaspartate and N-acetylaspartylglutamate in rat brain. AB - Administration of phencyclidine (PCP) to both humans and animals models the symptoms of schizophrenia. Brain concentrations of N-acetylaspartate (NAA) are reduced in this disease, reflecting neuronal dysfunction. This study investigates the effects in rats of a chronic intermittent regime of PCP on NAA and its precursor N-acetylaspartylglutamate (NAAG) in rat frontal and temporal cortex, hippocampus and striatum, determined by HPLC. We found significant PCP-induced deficits of NAA and NAAG only in the temporal cortex; NAAG was significantly elevated in the hippocampus. These changes closely reflect postmortem findings reported in schizophrenia. PMID- 15653258 TI - Evidence for glutamatergic neuronal dysfunction in the prefrontal cortex in chronic but not in first-episode patients with schizophrenia: a proton magnetic resonance spectroscopy study. AB - Based upon pharmacological challenge and postmortem studies, schizophrenia has been hypothesized to be caused by decreased glutamatergic neurotransmission. We investigated the glutamatergic neuronal metabolism of the dorsolateral prefrontal cortex with localized 1H magnetic resonance spectroscopy in 18 first-episode patients, 21 chronic patients with schizophrenia, and 21 age-matched controls. Chronic patients had significantly lower levels of glutamate/glutamine (Glx) and N-acetylaspartate (NAA) compared to healthy controls and first-episode patients. Reduced metabolite levels were not correlated with duration of illness or medication. Our results indicate glutamatergic dysfunction in chronic schizophrenia that could be evidence of a progressive brain disorder. PMID- 15653259 TI - Decreased expression of vesicular glutamate transporter 1 and complexin II mRNAs in schizophrenia: further evidence for a synaptic pathology affecting glutamate neurons. AB - Synaptic protein gene expression is altered in schizophrenia. In the hippocampal formation there may be particular involvement of glutamatergic neurons and their synapses, but overall the profile remains unclear. In this in situ hybridization histochemistry (ISHH) study, we examined four informative synaptic protein transcripts: vesicular glutamate transporter (VGLUT) 1, VGLUT2, complexin I, and complexin II, in dorsolateral prefrontal cortex (DPFC), superior temporal cortex (STC), and hippocampal formation, in 13 subjects with schizophrenia and 18 controls. In these areas, VGLUT1 and complexin II are expressed primarily by excitatory neurons, whereas complexin I is mainly expressed by inhibitory neurons. In schizophrenia, VGLUT1 mRNA was decreased in hippocampal formation and DPFC, complexin II mRNA was reduced in DPFC and STC, and complexin I mRNA decreased in STC. Hippocampal VGLUT1 mRNA declined with age selectively in the schizophrenia group. VGLUT2 mRNA was not quantifiable due to its low level. The data provide additional evidence for a synaptic pathology in schizophrenia, in terms of a reduced expression of three synaptic protein genes. In the hippocampus, the loss of VGLUT1 mRNA supports data indicating that glutamatergic presynaptic deficits are prominent, whereas the pattern of results in temporal and frontal cortex suggests broadly similar changes may affect inhibitory and excitatory neurons. The impairment of synaptic transmission implied by the synaptic protein reductions may contribute to the dysfunction of cortical neural circuits that characterises the disorder. PMID- 15653261 TI - Association between functioning in adolescence prior to first admission for schizophrenia and affective disorders and patterns of hospitalizations thereafter. AB - BACKGROUND: Kraepelin and Blueler suggested that subtle manifestations of schizophrenia are present in some persons for many years before formal diagnosis and that the severity of these is associated with outcomes in schizophrenia. Empirical support for this hypothesis comes primarily from small samples using retrospectively collected data. AIMS: We tested this hypothesis, for the first time, using a population-based cohort. METHOD: The Israeli Draft Board Registry, which contains measures of intellectual and behavioral functioning for the unselected population of 17-year-olds, was merged with the National Psychiatric Hospitalization Case Registry that contains data on all psychiatric hospitalizations. The database was used to identify adolescents assessed by the draft board at least 1 year prior to their first hospitalization for schizophrenia (n=996) or affective disorder (n=335). RESULTS: Poorer social functioning and organizational ability prior to first admission were associated with more days per year in the hospital for the male schizophrenia group. There were no significant correlations between days per year in the hospital and any of the behavioral functioning measures for the affective group. Among females the higher the previous level of intellectual functioning the fewer the days per year in the hospital in both the schizophrenia group and affective groups. For males no such correlations were evident. The comparisons between patients who had one as opposed to more than one admission found that in both diagnostic groups female patients with one admission had higher pre-first hospitalization intellectual functioning. CONCLUSIONS: Gender and disease specific premorbid deficits have may have differential prognostic value for outcomes in schizophrenia and affective disorders. PMID- 15653260 TI - Prodromal interventions for schizophrenia vulnerability: the risks of being "at risk". AB - Given the morbidity and difficulty of treating psychotic disorders, including schizophrenia, there has been a move toward identifying and treating adolescents and young adults who appear to be clinically at risk or "prodromal" to psychosis. The field now has greater specificity in identification, with rates of 40-50% conversion to frank psychosis within 1-2 years. There is further evidence that medications and other treatments may have some efficacy for "prodromal" patients, though with variable side effects. However, controversy remains about some of the inherent risks in prodromal research, such as medication exposure and stigma among false-positives. In this paper, we add to this discussion through an analysis of ethics in prodromal research from the more established field of predictive genetic testing. Issues are raised about the effects of information on patients, families, and institutions, as well as future insurability, the limits of confidentiality (as it relies on discretion of patients and families), the autonomy of minors with psychiatric symptoms, and even the risks for the true positive patient. PMID- 15653262 TI - Targeting synapses and myelin in the prevention of schizophrenia. AB - Many of the functions that are mediated by the prefrontal cortex (PFC) are severely impaired in schizophrenia. The maturation of these functions takes place during late adolescence and early adulthood, which coincides with the period of time when overt symptomatology of schizophrenia most commonly emerges. Two developmental processes occurring during the periadolescence period appear to mediate the functional maturation of the PFC: pruning of exuberant synapses and myelination of axons. It has long been speculated in the literature that disturbances of these processes may result in dysfunction of the PFC and thereby trigger the emergence of symptoms and deficits of schizophrenia. Alternatively, but not mutually exclusively, it has also been suggested that these late developmental processes may not be aberrant but they "unmask" preexisting deficits in the PFC, resulting in the onset of symptoms. The important implication of both of these scenarios is that in either case the emergence of PFC functional disturbances and the onset of symptoms and deficits of schizophrenia would in theory be preventable by pharmacologic manipulation of the synaptic pruning and/or axonal myelination processes. Thus, better understanding of the cellular and molecular mechanisms that mediate these processes will provide truly novel insight into the therapeutics and prevention of schizophrenia. PMID- 15653263 TI - N-acetyl-aspartate levels in the dorsolateral prefrontal cortex in the early years of schizophrenia are inversely related to disease duration. AB - Magnetic resonance spectroscopy studies in schizophrenia have revealed consistently reduced N-acetyl aspartate (NAA) levels in chronic patients, but not in recent-onset patients. Studies on the relationship between this marker and disease duration have commonly been negative, although it is also true that they have been conducted in patients with long-standing disease. We compared NAA levels in the dorsolateral prefrontal cortex in 16 recent-onset patients (duration: 1.8+/-0.6 years), 19 chronic patients (duration: 9.7+/-6.1 years), and 20 healthy controls. We studied the NAA/creatine and choline/creatine ratios in the dorsolateral prefrontal cortex in both hemispheres, controlling for the effect of age. Chronic patients had significantly lower NAA/Cr ratios in the left hemisphere compared to recent-onset patients and healthy controls, with no difference in Cho/Cr ratio. There were no differences between controls and recent onset patients. There was a significant inverse relationship between left-side NAA/Cr and disease duration, suggesting that prefrontal NAA levels may progressively decrease in schizophrenia. Taken within the context of the existing literature, these results indicate that this process may be limited to the early years following the onset of the disease. Therefore, reduced prefrontal levels of NAA may be limited to chronic schizophrenia patients. PMID- 15653264 TI - Meta-analysis of the time-course of brain volume reduction in schizophrenia: implications for pathogenesis and early treatment. AB - BACKGROUND: Whole brain volume (BV) is significantly reduced in groups of schizophrenic patients, but there is disagreement as to when in relation to onset of illness these losses occur. Given what is known about the normal development and lifetime course of BV, intraventricular volume (IVV), extracerebral volume (ECV), and intracranial volume (ICV) changes, it is possible to allocate (within a narrow range of uncertainty) excessive brain volume loss (EBVL) to either the time before or the time after attainment of maximum brain volume (BVmax). METHOD: Decreases in patient ICV relative to control ICV reflect early reductions in brain growth, while increases in ECV reflect later BV losses. There is however uncertainty as whether any relative increases in patient IVV occur early, later, or both. IVV differences were first assumed to be of early origin, so that early EBVL was measured by ICV-IVV differences and later EBVL by ECV differences alone. The IVV differences were then assumed to be of later origin, so that early EBVL was measured by ICV differences alone, and later EBVL by ECV+IVV differences. The results taken together delineate the maximum and minimum values for early and later losses. RESULTS: Patient-control volume differences in BV and ICV for 20 published magnetic resonance imaging (MRI) studies of schizophrenic patients (n=982) and controls (n=1049), and differences in ECV for 17 of the same 20 studies, comprising 889 patients and 942 controls, showed a significant patient BV decrease of 34 cc's (t=-4.94, df=19, p<0.0001), ICV decrease of 20.1 cc's (t= 2.64, df=19, p<0.02) and ECV Increase of 14.1 cc's (t=3.65, df=16, p<0.001). In the 17 studies which included ECV and IVV, as well as ICV differences, the patient ICV-IVV decrease was 20.2 cc's (t=-2.56, df=16, p<0.05) and the ECV+IVV increase was 17.1 cc's (t=-4.11, df=16, p<0.001). CONCLUSION: There is a small but significant whole brain EBVL in schizophrenia patients both before and after BV(max), regardless of when excessive IVV enlargement is assumed to occur. PMID- 15653265 TI - Globus pallidus volume is related to symptom severity in neuroleptic naive patients with schizophrenia. AB - This study compares globus pallidus (GP) volume between neuroleptic naive patients with schizophrenia and healthy controls using structural MRI. The volume of the external segment of the GP (GPe) was positively correlated with the severity of global symptoms, as measured by the Scale for the Assessment of Negative Symptoms and Scale for the Assessment of Positive Symptoms (SANS/SAPS, Andreasen and Olsen, 1982). The volume for the GP, GPe, and internal segment (GPi) did not differ between groups. PMID- 15653266 TI - Superior temporal gyrus differences in childhood-onset schizophrenia. AB - The posterior superior temporal gyrus (STG) is the approximate site of Wernicke's area, a language region, which in previous studies has been reported to be abnormal in adults with schizophrenia. The present study assesses volumetric differences in the superior temporal gyrus of subjects with childhood-onset schizophrenia (COS). MRI scans of 18 subjects diagnosed with childhood-onset schizophrenia and 16 age- and sex-matched normals were analyzed to assess possible volume differences. The COS subjects displayed significant enlargement of the right posterior superior temporal gyrus, showing white matter increases bilaterally in this region. Our findings are consistent with studies that have found increased volumes in temporal lobe regions in COS and may provide a possible neural correlate for the language impairment observed in COS patients. PMID- 15653267 TI - Childhood-onset schizophrenia: smooth pursuit eye-tracking dysfunction in family members. AB - BACKGROUND: Childhood-onset schizophrenia (COS), a severe form of the disorder, is of interest for etiologic studies. Smooth pursuit eye-tracking dysfunction (ETD) is a biological marker for schizophrenia. AIMS: To compare familial eye tracking abnormalities for COS and adult-onset schizophrenia (AOS). METHOD: Eye tracking performance for 70 COS parents, 64 AOS parents and 20 COS siblings was compared to their respective age-matched control groups. RESULTS: COS and AOS parents had higher rate of dichotomously rated eye-tracking dysfunction than their respective controls (16% vs. 1% and 22% vs. 4%, respectively). COS parents and siblings also differed from controls on several continuous measures. However, scores for COS, AOS and control groups overlapped extensively. CONCLUSIONS: Genetic factors underlying eye-tracking dysfunction appear more salient for COS. However, eye-tracking measures have to be used with caution for endophenotypic definition due to low predictive power. DECLARATION OF INTEREST: The study was done at the National Institutes of Health. PMID- 15653268 TI - FOXP2 polymorphisms in patients with schizophrenia. AB - BACKGROUND: FOXP2 was described as the first gene involved in our ability to acquire spoken language. The main objective of this study was to compare the distribution of FOXP2 gene polymorphisms between patients with schizophrenia and healthy controls. METHODS: Two FOXP2 polymorphisms, Intron3a and SNP 923875, and the G-->A transition in exon 14 were analysed in 149 patients with schizophrenia and schizoaffective disorders according to DSM-IV, as well as in 137 controls. All the patients showed a history of auditory hallucinations. RESULTS: The transition G-->A at exon 14, detected in all the affected members in KE family, was not found in any of the analyzed samples from patients or controls. No significant differences were found between individual controls and patients for the two analysed polymorphisms. CONCLUSIONS: This study would not support a possible role of the two FOXP2 analyzed polymorphisms in the vulnerability to schizophrenia. PMID- 15653269 TI - A family-based study of the association between the G72/G30 genes and schizophrenia in the Chinese population. AB - Studies have shown a strong positive association between schizophrenia and G72/G30, demonstrated by both individual markers and haplotypes. A further functional study also supports the role of G72 in the etiology of schizophrenia. In this study, we have replicated these results of transmission/disequilibrium testing (TDT) and haplotype analysis in the Han Chinese population, showing P values of 0.0018 and 0.00007 for individual markers and haplotypes, respectively. Hence, our data supports the hypothesis that G72/G30 are important candidate genes for explaining schizophrenia in the Han Chinese population. PMID- 15653270 TI - A survey of the 22q11 microdeletion in a large cohort of schizophrenia patients. AB - The occurrence of a microdeletion at 22q11 has long been considered to constitute a risk factor for schizophrenia. Higher rates of 22q11 deletions have been reported in cohorts of patients with schizophrenia. In order to estimate the prevalence of the 22q11 deletion in schizophrenia patients more accurately, a screening for 22q11 deletions was conducted on a cohort of 634 schizophrenia patients, the largest sample size screened to date. Seven microsatellites and three SNPs were used to assess the deletion genotype. In cases where all markers were found to be homozygous (hemizygous), the individual was assumed to carry the deletion. The method used here is simple and efficient in comparison with hybridization technologies. Moreover, the rate of false positives is very low (P value in the range of 10(-4) to 10(-3)). Approximately 1% of the patient cohort was found to carry 22q11 deletions. PMID- 15653271 TI - Case-control association study of the close homologue of L1 (CHL1) gene and schizophrenia in the Chinese population. AB - The close homolog of L1 (CHL1), located on human chromosome 3p26.1, is a newly identified member of the L1 family of cell adhesion molecules which play important roles in cell migration, axonal growth, and synaptic remodeling. A positive association has been reported between a missense polymorphism in CHL1 gene and schizophrenia in the Japanese population [Sakurai, K., Migita, O., Toru, M., Arinami, T., 2002. An association between a missense polymorphism in the close homologue of L1 (CHL1, CALL) gene and schizophrenia. Mol. Psychiatry 7, 412 415]. An association between a missense polymorphism in the close homologue of L1 (CHL1, CALL) gene and schizophrenia. In order to test this finding, we genotyped four SNPs in the gene in the Han Chinese population using a sample of 560 cases and 576 controls. Analysis of allele frequencies in both samples also showed strong association between SNP rs2272522 (the same marker studied by K. Sakurai) and the disease (X2=31.591, P<0.000001, OR=1.745, 95% CI=1.435-2.121). Our results confirm the positive association between CHL1 gene and schizophrenia and indicate that CHL1 may be involved in the etiology of schizophrenia. PMID- 15653272 TI - A family-based association study of the MOG gene with schizophrenia in the Chinese population. AB - Recently the expression of human myelin/oligodendrocyte glycoprotein (MOG) has been found to be significantly downregulated in the brain tissue of subjects with schizophrenia, suggesting that the MOG gene resides within a high-susceptibility locus for schizophrenia. In order to test this prediction, we analyzed three microsatellites from MOG in the Han Chinese population using a sample of 532 trios. Analysis of allele, genotype and haplotype frequencies showed weak positive association between the markers and the disease (p=0.01982). Our results would indicate that the MOG gene may play a significant role in schizophrenia in the Han Chinese. However, further study is required using other methods and involving other populations. PMID- 15653273 TI - Five NOTCH4 polymorphisms show weak evidence for association with schizophrenia: evidence from meta-analyses. AB - NOTCH4 initially received consideration as a risk gene for schizophrenia based on its location within a region on chromosome 6p that had previously shown strong evidence for genetic linkage with the illness. The initial published test for allelic association found strong evidence for involvement of this gene in schizophrenia, but subsequent studies failed to confirm this finding. Presently, we have used meta-analysis to derive a best estimate of the nature and magnitude of the associations between schizophrenia and five polymorphisms in and around the NOTCH4 gene. No significant association was detected between schizophrenia and repeat length of alleles at the (TAA)n, (CTG)n, or (TTAT)n polymorphisms, or between the disease and specific risk alleles at these polymorphisms or at the SNP1 or SNP2 polymorphisms. Heterogeneity and stronger evidence of association with the putative risk alleles of the (TAA)n, (CTG)n, SNP1, and SNP2 polymorphisms was observed in family-based studies than in case-control studies, suggesting that these polymorphisms may reliably influence risk for schizophrenia under certain circumstances. Since more consistent and robust associations with schizophrenia risk have been observed for haplotypes of these polymorphisms [especially those containing SNP2 and (CTG)n], additional large family-based or genomic-controlled studies would be helpful for definitively specifying the role of NOTCH4 haplotypes in risk for schizophrenia. PMID- 15653274 TI - Processing of environmental sounds in schizophrenic patients: disordered recognition and lack of semantic specificity. AB - The recognition of environmental sounds is an important feature of higher auditory processing and essential for everyday life. The present study aimed to investigate the potential impairment of this mental function in schizophrenia. This work on immediate sound recognition is complementary to recent studies on auditory linguistic processing. Fifteen schizophrenic patients and 30 control subjects were asked to identify 43 complex environmental sounds from different categories and rate their familiarity when naive to the sounds. In consecutive experiments, patients and control subjects rated the sounds according to emotional valence and arousal, as well as imageability. In both groups, correct identification of non-verbal sounds was highly associated with familiarity. Statistical analysis by group demonstrated a significantly higher error rate in identifying sounds in patients suffering from schizophrenia compared to healthy control subjects. In contrast, the affective recognition of the complex sounds was preserved in the schizophrenic patients. These results suggest a disturbance of higher-order, auditory mnemonic processing in schizophrenic patients in the non-linguistic domain. We discuss their abnormal responses in the context of recent theories of auditory physiological and semantic processing deficits in schizophrenia. PMID- 15653275 TI - Impaired mismatch negativity generation in prodromal subjects and patients with schizophrenia. AB - BACKGROUND: Mismatch negativity (MMN) specifically the response to tone duration deviants has consistently been shown to be reduced in schizophrenia suggesting dysfunction in auditory sensory memory. As part of a multidimensional approach to the early recognition of psychosis, MMN was investigated as a possible risk factor for later development of psychosis in subjects with a prodromal syndrome. Forty-three prodromal subjects, 31 neuroleptic-free inpatients with schizophrenia and 33 healthy controls were studied. A prodromal state was defined by a cluster 'Cognitive Disturbances' as defined by the 'Bonn Scale for the Assessment of Basic Symptoms' (BSABS), which was found highly predictive of first-episode schizophrenia. To elicit MMN, a three-tone auditory oddball paradigm with 10% 'duration deviants' and 10% 'frequency deviants' was used. RESULTS: MMN amplitudes to tone duration deviants were significantly reduced in the patients with schizophrenia compared to controls. The putatively prodromal subjects also showed a slight, though non-significant reduction of the MMN amplitude that was intermediate between normal controls and patients with schizophrenia, and with a larger within-group variance. CONCLUSION: These results support the view that abnormalities in temporal processing are particularly pronounced in patients with schizophrenia. Prodromal subjects are a heterogeneous group with regard to outcome and time until transition to a first psychotic episode. Follow-up of these putatively prodromal subjects will show whether MMN amplitudes further reduce over time in those developing psychosis and if a reduction is state dependent. PMID- 15653276 TI - M50 sensory gating predicts negative symptoms in schizophrenia. AB - Impaired auditory sensory gating is considered characteristic of schizophrenia and a marker of the information processing deficit inherent to that disorder. Predominance of negative symptoms also reflects the degree of deficit in schizophrenia and is associated with poorer pre-morbid functioning, lower IQ, and poorer outcomes. However, a consistent relationship between auditory sensory gating and negative symptoms in schizophrenia has yet to be demonstrated. The absence of such a finding is surprising, since both impaired auditory gating and negative symptoms have been linked with impaired fronto-temporal cortical function. The present study measured auditory gating using the P50 event related potential (ERP) in a paired-click paradigm and capitalized on the relative localization advantage of magnetoencephalography (MEG) to assess auditory sensory gating in terms of the event related field (ERF) M50 source dipoles on bilateral superior temporal gyrus (STG). The primary hypothesis was that there would be a positive correlation between lateralized M50 auditory sensory gating measures and negative symptoms in patients with schizophrenia. A standard paired-click paradigm was used during simultaneous EEG and MEG data collection to determine S2/S1 sensory gating ratios in a group of 20 patients for both neuroimaging techniques. Participants were administered the Schedule for the Assessment of Negative Symptoms (SANS), the Positive and Negative Symptom Scale (PANSS), and the Calgary Depression Scale for Schizophrenia. Consistent with previous reports, there was no relationship between ERP P50 sensory gating and negative symptoms. However, right (not left) hemisphere ERF M50 sensory gating ratio was significantly and positively correlated with negative symptoms. This finding is compatible with information processing theories of negative symptoms and with more recent findings of fronto-temporal abnormality in patients with predominantly negative symptoms. PMID- 15653277 TI - Sensorimotor gating, orienting and social perception in schizophrenia. AB - Basic neurocognition and social cognition appear to influence the social impairments of persons with schizophrenia. This study examined relationships between two very basic automatic processes (i.e., sensorimotor gating and orienting) and social perception in schizophrenic patients. Thirty outpatients with schizophrenia completed psychophysiological measures of sensorimotor gating (prepulse inhibition, PPI), orienting (prepulse facilitation, PPF), and social perception (the Half Profile of Nonverbal Sensitivity, Half PONS). A median split was used to divide patients into poor and good gaters and poor and good orienters. Analyses revealed that patients with good PPI scored significantly higher on the Half PONS than patients with poor PPI. PPI showed a significant correlation (r=-0.54) with Half PONS performance, indicating that schizophrenia patients who were better able to gate out competing stimuli (i.e., less startle) were also better at detecting relevant social cues. Orienting (PPF) and social perception were not related. This study is the first to our knowledge to demonstrate an association between sensorimotor gating and social perception. The findings are consistent with other studies that have demonstrated relationships between basic neurocognition and social cognition. By showing a link between sensorimotor gating and social perception, this study supports social cognition's potential role as a mediator of the relationship between neurocognition and social functioning in schizophrenia. PMID- 15653278 TI - Auditory gating in schizophrenia: a pilot study of the precedence effect. AB - Individuals with schizophrenia are known to exhibit diminished auditory gating, as manifested by prepulse inhibition of the startle response and suppression of the P50 evoked potential. Those observations indicate that schizophrenics have deficits of inhibitory auditory processing. The precedence effect is a phenomenon in which inhibitory processing in the auditory system aids the localization of sound sources and segregation of speech in complex auditory environments. We tested the hypothesis that schizophrenic subjects would exhibit deficits in psychophysical tests of the precedence effect. The performance of 12 male medicated schizophrenic subjects was compared with that of 12 male comparison subjects. Schizophrenics and comparison subjects clearly demonstrated all three aspects of the precedence effect that were evaluated: echo suppression, buildup of echo suppression, and breakdown of echo suppression. In none of these aspects did the performance of schizophrenic subjects differ significantly from that of the comparison subjects. These findings suggest that abnormalities of auditory spatial processing and speech segregation are unlikely to contribute to the clinical pathology of schizophrenia. PMID- 15653279 TI - Cancer in schizophrenia: is the risk higher or lower? AB - Studies exploring the relationship between schizophrenia and cancer have shown conflicting results. Our study explores this association in three Jewish-Israeli population groups defined by their continent/place of birth (Israel, Europe America, and Africa-Asia). The identification of the patients was made through the linkage of the nationwide psychiatric and cancer registries. The incidence of cancer in patients diagnosed with schizophrenia was compared with the incidence in the general population. The results showed that the cancer standardized incidence ratios (SIRs) for all sites were significantly lower among men and women with schizophrenia, 0.86 [95% confidence interval (CI) 0.80-0.93] and 0.91 (95% CI 0.85-0.97), respectively. This reduced overall risk was clearest for those born in Europe-America, both men (SIR 0.85, 95% CI 0.74-0.97) and women (SIR 0.86, 95% CI 0.77-0.94). Among women diagnosed with schizophrenia, the SIR was statistically higher for cancer in the breast among those born in Asia-Africa (1.37, 95% CI 1.12-1.63) and in the corpus uteri among the Israel-born (2.75, 95% CI 1.69-3.81) than among their counterparts in the general population. Lung cancer was significantly higher in men born in Asia-Africa diagnosed with schizophrenia than in the respective comparison population group (1.58, 95% CI 1.13-2.2). Our findings, and those of the literature, justify conducting a multinational study that includes identification of cancer-related risk factors among patients with schizophrenia and their families, and information on the use of psychotropic medications. This effort may clarify an epidemiological puzzle that remains outstanding. PMID- 15653280 TI - Febrile seizures and risk of schizophrenia. AB - BACKGROUND: Febrile seizure is a benign condition for most children, but experiments in animals and neuroimaging studies in humans suggest that some febrile seizures may damage the hippocampus, a brain area of possible importance in schizophrenia. METHODS: A population-based cohort of all children born in Denmark between January 1977 and December 1986 was followed until December 2001 by using data from nationwide registries. RESULTS: We followed 558,958 persons including 16,429 with a history of febrile seizures for 2.8 million person-years and identified 952 persons who were diagnosed with schizophrenia. A history of febrile seizures was associated with a 44% increased risk of schizophrenia [relative risk (RR)=1.44; 95% confidence interval (CI), 1.07-1.95] after adjusting for confounding factors. The association between febrile seizures and schizophrenia remained virtually unchanged when restricting the analyses to people with no history of epilepsy. A history of both febrile seizures and epilepsy was associated with a 204% increased risk of schizophrenia (RR=3.04; 95% CI, 1.36-6.79) as compared with people with no such history. CONCLUSIONS: We found a slightly increased risk of schizophrenia among persons with a history of febrile seizures. The association may be due to a damaging effect of prolonged febrile seizures on the developing brain, shared etiological factors, or confounding by unmeasured factors. PMID- 15653281 TI - Cumulative exposure to estrogen and psychosis: a peak bone mass, case-control study in first-episode psychosis. AB - The organizational structural effects of estrogen may be cumulative and permanent by impacting on neurodevelopment, giving rise to "neuroprotective" effects and eventually reduction of psychosis risk. Reduction in bone mineral density (BMD, in g/cm2), as a biological marker of reduced cumulative exposure to estrogen, may be a marker of increased psychosis risk. A sample of 19 first-episode female psychosis patients with minimal previous antipsychotic exposure (mean 10 weeks) and 20 female controls underwent advanced fan-beam dual X-ray absorptiometry (DXA) to assess lumbal spine BMD of the region of L1-L4. Mean BMD was around one standard deviation lower in patients (1.13, S.D.=0.10) than in controls (1.25, S.D.=0.12; p=0.0021), and 84% of patients scored below the median value of the controls (OR=5.3, 95% CI: 1.2, 24.2). The results are compatible with the hypothesis that psychosis in women may be associated causally with a reduced protective effect of estrogen over the course of development. PMID- 15653282 TI - Plasma concentrations of estradiol in women suffering from schizophrenia treated with conventional versus atypical antipsychotics. AB - BACKGROUND: Low estrogen levels leading to an elevated rate of menstrual dysfunctions such as amenorrhea and irregular menstruation have been described in women with schizophrenia and have often been attributed to antipsychotic-induced hyperprolactinemia. However, there is some evidence that "hypoestrogenism" in schizophrenic women does not occur exclusively under medication with hyperprolactinemia-inducing antipsychotics. While the precise mechanism of low estrogen levels in schizophrenic women has not been elucidated yet, "hypoestrogenism" is of clinical relevance because estrogen seems to endow an antipsychotic-like effect in schizophrenia and thus positively affect the course of illness in schizophrenic women. In addition, low levels of estrogen might have a negative effect on bone mineral density and on the cardiovascular system. METHODS: To test the "hypoestrogenism hypothesis", hormone levels in 75 women with schizophrenia diagnosed according to DSM-IV and ICD-10 were determined in the follicular, periovulatory, and luteal phases of the menstrual cycle. Levels of estradiol, prolactin, luteinizing hormone (LH), follicle-stimulating hormone (FSH), progesterone, and testosterone were assessed. RESULTS: The serum levels of estradiol were generally reduced during the entire menstrual cycle compared to normal reference values. With low levels of LH over the entire cycle and of progesterone in the luteal phase, anovulatory cycles were assumed. Hypoestrogenism was found in about 60% of the patients in accordance with a strict definition (estradiol serum level below 30 pg/ml in the follicular phase and below 100 pg/ml in the periovulatory phase). To rule out a possible effect of hyperprolactinemia on the gonadal axis and a subsequent effect on estradiol levels from treatment with conventional ("typical") antipsychotics, serum estradiol levels of patients treated with certain atypical antipsychotics known to induce only a mild increase in prolactin, or no increase at all, were compared with those from patients treated with conventional antipsychotics. The data clearly indicate high prolactin levels in the latter, but low levels in the group treated with atypical antipsychotics. In both groups, however, low levels of estradiol compared to normal reference values were measured. CONCLUSIONS: The present findings provide evidence that hypoestrogenism in schizophrenia occurs in women with and without antipsychotic-induced hyperprolactinemia. Further research should be conducted to clarify the cause of hypoestrogenism in schizophrenic women and focus on possible clinical implications. PMID- 15653283 TI - Should allosteric positive modulators of GABA(A) receptors be tested in the treatment of schizophrenia? PMID- 15653284 TI - Correlates of substance abuse in adolescents with treatment-refractory schizophrenia and schizoaffective disorder. PMID- 15653285 TI - High-dose quetiapine in treatment refractory schizophrenia. PMID- 15653286 TI - Impairing effects of chronic haloperidol and clozapine treatment on recognition memory: possible relation to oxidative stress. PMID- 15653287 TI - Serum glutamic acid decarboxylase 65 antibody levels in people with schizophrenia and their families. PMID- 15653288 TI - A missense polymorphism (H204R) of a Rho GTPase-activating protein, the chimerin 2 gene, is associated with schizophrenia in men. PMID- 15653289 TI - Mental state inference using visual control parameters. AB - Although we can often infer the mental states of others by observing their actions, there are currently no computational models of this remarkable ability. Here we develop a computational model of mental state inference that builds upon a generic visuomanual feedback controller, and implements mental simulation and mental state inference functions using circuitry that subserves sensorimotor control. Our goal is (1) to show that control mechanisms developed for manual manipulation are readily endowed with visual and predictive processing capabilities and thus allows a natural extension to the understanding of movements performed by others; and (2) to give an explanation on how cortical regions, in particular the parietal and premotor cortices, may be involved in such dual mechanism. To analyze the model, we simulate tasks in which an observer watches an actor performing either a reaching or a grasping movement. The observer's goal is to estimate the 'mental state' of the actor: the goal of the reaching movement or the intention of the agent performing the grasping movement. We show that the motor modules of the observer can be used in a 'simulation mode' to infer the mental state of the actor. The simulations with different grasping and non-straight line reaching strategies show that the mental state inference model is applicable to complex movements. Moreover, we simulate deceptive reaching, where an actor imposes false beliefs about his own mental state on an observer. The simulations show that computational elements developed for sensorimotor control are effective in inferring the mental states of others. The parallels between the model and cortical organization of movement suggest that primates might have developed a similar resource utilization strategy for action understanding, and thus lead to testable predictions about the brain mechanisms of mental state inference. PMID- 15653290 TI - Electrophysiological correlates of forming memories for faces, names, and face name associations. AB - The ability to put a name to a face is a vital aspect of human interaction, but many people find this extremely difficult, especially after being introduced to someone for the first time. Creating enduring associations between arbitrary stimuli in this manner is also a prime example of what patients with amnesia find most difficult. To help develop a better understanding of this type of memory, we sought to obtain measures of the neural events responsible for successfully forming a new face-name association. We used event-related potentials (ERPs) extracted from high-density scalp EEG recordings in order to compare (1) memory for faces, (2) memory for names, and (3) memory for face-name associations. Each visual face appeared simultaneously with a unique spoken name. Signals observed 200-800 ms after the onset of face-name pairs predicted subsequent memory for faces, names, or face-name associations. Difference potentials observed as a function of subsequent memory performance were not identical for these three memory tests, nor were potentials predicting associative memory equivalent to the sum of potentials predicting item memory, suggesting that different neural events at the time of encoding are relevant for these distinct aspects of remembering people. PMID- 15653291 TI - The nature of switch cost: task set configuration or carry-over effect? AB - The primary goal of the present study was to differentiate variants of each class of hypothetical models of task switching. Two event-related potentials, the lateralized readiness potential (LRP) and P300 were analyzed to infer the roles of advance reconfiguration and carry-over effect upon task switching. Participants performed two tasks in a random order, so that on each trial, they either repeated the task from the previous trial or switched to another task. Pre cues that preceded each stimulus were either informative (task-cueing conditions), specifying which of the two tasks to perform, or uninformative (no task-cueing conditions). The results showed that the mean reaction time (RT) and the interval between stimulus onset and LRP onset were longer for switch than for repeat trials. This suggested that task switching affected processes before LRP onset (including stimulus identification and response selection). A further analysis of P300 confined task switching to the process after the stimulus identification stage. Task cueing, meanwhile, influenced both the latency and peak amplitude of P300, suggesting that the cueing effect occurred prior to the response selection stage. The model of configuration-execution with moderate modifications is a candidate to explain the present results. PMID- 15653292 TI - The influence of increased working memory load on semantic neural systems: a high resolution event-related brain potential study. AB - The effects of working memory (WM) on the semantic N400 response were studied using high-resolution event-related brain potentials (ERPs). Participants were presented with semantically related sentence pairs and the terminal word congruence was varied in the second sentence. WM load was varied for the sentence pairs using a modified fan procedure [J.R. Anderson, Retrieval of propositional information from long-term memory. Cogn. Psychol., 6 (1974) 451-474; J.R. Anderson, A spreading activation theory of memory. J. Verbal Learn. Verbal Behav., 22 (1983) 261-295]. ERPs were recorded to the onset of the terminal word (Congruent and Incongruent) for sentence pairs at two levels of WM load (WML1 and WML2). Behavioral data analysis revealed that reaction times (RTs) increased as a function of WM load (i.e., the fan effect). Semantically incongruent words elicited an N400 response. Increased WM load reduced the congruency effect and, importantly, resulted in a significant delay in N400 peak latency (approximately 50 ms). Moreover, the latency delay was correlated with a neuropsychological measure of individual WM capacity. WM load effects on the N400 were interpreted as a result of competing activation in WM, both modulating semantic expectancies and delaying semantic integration. Exploratory source analysis revealed activation in occipital, temporal, and parietal regions. Consistent with prior reports on the N400 and semantic processing, equivalent current dipoles were modeled in the left perisylvian region. Increased WM load led to novel source activation in the left inferior parietal region as well as increased activation levels in anterior temporal sources. The source modeling results were in agreement with the functional roles typically ascribed to these areas and confirmed that the scalp-recorded WM load effects on the N400 were present within the intracranial generators. PMID- 15653293 TI - Neural organization for recognition of grammatical and emotional facial expressions in deaf ASL signers and hearing nonsigners. AB - Recognition of emotional facial expressions is universal for all humans, but signed language users must also recognize certain non-affective facial expressions as linguistic markers. fMRI was used to investigate the neural systems underlying recognition of these functionally distinct expressions, comparing deaf ASL signers and hearing nonsigners. Within the superior temporal sulcus (STS), activation for emotional expressions was right lateralized for the hearing group and bilateral for the deaf group. In contrast, activation within STS for linguistic facial expressions was left lateralized only for signers and only when linguistic facial expressions co-occurred with verbs. Within the fusiform gyrus (FG), activation was left lateralized for ASL signers for both expression types, whereas activation was bilateral for both expression types for nonsigners. We propose that left lateralization in FG may be due to continuous analysis of local facial features during on-line sign language processing. The results indicate that function in part drives the lateralization of neural systems that process human facial expressions. PMID- 15653294 TI - Processing semantic anomalies in two languages: an electrophysiological exploration in both languages of Spanish-English bilinguals. AB - The latency of the brain response to semantic anomalies (N400 effect) has been found to be longer in a bilingual's second language (L2) than in their first language (L1) and/or to that seen in monolinguals. This has been explained in terms of late exposure to L2, although age of exposure and language proficiency are often highly correlated. We thus examined the relative contributions of these factors not only in L2 but also in L1 in a group of Spanish-English bilinguals for whom age of exposure and language proficiency were not highly correlated by recording event-related brain potentials (ERPs) to semantically congruous/incongruous words completing written sentences. We also divided our bilinguals into a Spanish-dominant subset who had late exposure and reduced vocabulary proficiency [as measured by Boston Naming Test (BNT) and Verbal Fluency Scores] in L2 (English) relative to L1 (Spanish) and an English-dominant group who had early exposure to both their languages although greater proficiency in English than in Spanish. In both groups, the N400 effect was significantly later in the nondominant than the dominant language. Although this slowing could be due to late exposure to English in the Spanish-dominant group, late exposure cannot explain the slowing in Spanish in the English-dominant group. Overall, we found that vocabulary proficiency and age of exposure are both important in determining the timing of semantic integration effects during written sentence processing--with vocabulary proficiency predicting the timing of semantic analysis in L1 and both age of exposure and language proficiency, although highly correlated, making additional small but uncorrelated contributions to the speed of semantic analysis/integration in L2. PMID- 15653295 TI - Source analysis of the N2 in a cued Go/NoGo task. AB - Previous source analyses of event-related potential (ERP) data elicited in Go/NoGo tasks have suggested that the anterior cingulate cortex (ACC) plays an important role in response inhibition. So far, however, source models were derived for the difference wave Go stimulus minus NoGo stimulus. This difference wave is confounded with motor- and attention-related activity. To avoid these confounds, we alternatively derived source models for NoGo stimuli only. The problem of the NoGo-N2 being superimposed on a positive deflection was addressed in two ways. First, a baseline correction was applied using the time points just preceding and succeeding the NoGo-N2. Second, a separate source model was derived at the maximum amplitude of this positive deflection. Subjects were presented with a cued version of the continuous performance task (CPT; ABX). In a second study, the probability of the Go stimulus was gradually increased to heighten subjects' tendency to respond and, as a consequence, to enhance the amplitude of the NoGo-N2. The source models of the NoGo-N2 consistently indicated bilateral dipole pairs in medial frontal regions. This is in accordance with a generator in the anterior cingulate cortex. PMID- 15653296 TI - Experience modulates automatic imitation. AB - Action observation gives rise to activation in corresponding areas of the premotor and primary motor cortices. We tested the hypothesis that this activation depends on visual-motor connections established through correlated experience of observing and executing the same action. Previous work has shown that hand opening and hand closing gestures are facilitated when subjects observe the movement they are performing, relative to a condition in which they observe a different movement from the one they are performing. Experiment 1 replicated this finding in a simple reaction time (RT) procedure using stimulus-response (SR) movements in orthogonal planes. This implies that the effect is an example of automatic imitation, an instruction-independent tendency to execute movements that are topologically similar to those observed, and not merely an example of spatially compatible responding. In Experiment 2, the automatic imitation effect found in Experiment 1 was abolished by a brief period of training in which subjects responded to hand opening by closing their hands, and to hand closing by opening their hands. This outcome is consistent with the hypothesis that, rather than being innate, the cortical connections mediating motor activation by action observation are formed through experience. PMID- 15653297 TI - An ERP study of P600 effects elicited by semantic anomalies. AB - Syntactic anomalies reliably elicit P600 effects. Recent studies, however, reported P600 effects to semantic anomalies. These findings are difficult to reconcile with the common view on the P600 as a purely syntactic component. The present study--carried out in Dutch--tested the possibility that a P600 to semantic anomalies would nevertheless reflect syntactic processing. We presented semantic reversal anomalies in syntactically correct and unambiguous sentences, like #The cat that fled from the mice.... If participants would use a plausibility strategy and combine the lexical items in the most plausible way, they would--in the case of the example--assume that the mice were fleeing from the cat. Furthermore, this interpretation could lead them to expect a particular inflection of the verb (here: plural inflection). The violation of this expectation could have elicited the P600 effect. Such a syntactic mismatch can occur only in sentences in which the number of theme and agent are different. Therefore, in the present study, the number of theme and agent was either different or the same. A centroparietal P600 effect was present not only in different number sentences but also in same number sentences. Consequently, the P600 effect was not due to a syntactic mismatch, thereby challenging a purely syntactic account of the P600. An alternative view concerning the functional significance of the P600 is discussed, i.e., that it reflects a monitoring component that checks upon the veridicality of ones sentence perception. PMID- 15653298 TI - Intensive baseball practice improves the Go/Nogo reaction time, but not the simple reaction time. AB - Baseball hitters are required to make decisions whether to swing or not as quickly as possible. Therefore, we can assume that skilled baseball players have a quicker response. To verify this hypothesis, we assessed the effect of baseball experience or skill levels on simple reaction times and Go/Nogo reaction times in 82 university students (22 baseball players, 22 tennis players, and 38 nonathletes) and 17 professional baseball players. Also, to clarify whether this ability was innate or acquired, we examined the effects of long-term practice for baseball hitting in 94 senior high school students (26 baseball players and 68 non-baseball players). There were no differences in simple reaction time either for sports experience or for skill levels. On the contrary, the Go/Nogo reaction time for baseball players was significantly shorter than that of the tennis players and nonathletes. The Go/Nogo reaction time of higher-skill baseball players was significantly shorter than that of lower-skill players, while that of the professional baseball players was the shortest. The professional players showed the highest (almost linear) correlation between the Go/Nogo reaction time and simple reaction time. A longitudinal study showed that 2 years of hitting practice improved the Go/Nogo reaction time, while the simple reaction time remained constant. A cross-sectional study of high school non-baseball players showed no differences either in the simple or Go/Nogo reaction times in relation to school year. In conclusion, intensive practice, including Go or Nogo decision making, improved the Go/Nogo reaction time, but not the simple reaction time. PMID- 15653299 TI - Perceptual fluency, semantic familiarity and recognition-related familiarity: an electrophysiological exploration. AB - Scalp recorded event-related potentials (ERPs) were used to examine the neuronal activity associated with perceptual fluency, semantic familiarity and recognition related familiarity. We assume that ERP differences between first and second presentations of non-famous faces in an implicit memory condition reflect perceptual fluency, ERP differences between first presentations of famous and non famous faces reflect semantic familiarity (i.e., familiarity arising from semantic memory retrieval), and early ERP differences between first and second presentations of non-famous and famous faces in an explicit recognition memory task reflect recognition-related familiarity. Semantic familiarity elicited a broadly distributed effect between 200 and 300 ms after stimulus onset, possibly representing the activation of face recognition units. Between 300 and 450 ms, frontal effects were observed for semantic familiarity and recognition-related familiarity, while perceptual fluency was associated with a centro-parietally focused effect. Thus, familiarity arising from the retrieval of semantic information and recognition-related familiarity depend at least partly on the same neuronal circuits, while these are dissociable from those mediating perceptual fluency. PMID- 15653300 TI - The effect of numerical distance and stimulus probability on ERP components elicited by numerical incongruencies in mental addition. AB - In two experiments, we investigated the effect of the numerical distance of incongruous results from correct results and stimulus probability on the N270/N400 event-related brain potential components. In Experiment 1, 12 subjects saw two one-digit addends and a possible solution and signaled if the proposed result (falling in the range of 3-17) was true or false. Incorrect results could deviate by +/-2 or by +/-9 from the correct answer. The probability of correct results was 50%. Twelve subjects carried out a similar task in Experiment 2 without giving behavioral responses. The probability of incorrect results was 20%, 50% or 80% in different conditions. Both raw potentials and incorrect minus correct difference potentials were analyzed. A fronto-central N3 and a centro parietal dN3 (incorrect-correct difference) were present for incongruous results in both experiments. The amplitude of the dN3 was not sensitive to numerical distance, but the latency of the dN3 was longer when numerical distance was larger. The overall amplitude of the N3 and of the dN3 was not sensitive to the probability manipulation. However, there was a parietally localized effect of probability on N3 amplitude. The dN3 in mental addition is most probably identical to the arithmetic N400 effect reported earlier in mental multiplication. The distance effect in latency may be a correlate of the discrimination of correct vs. incorrect results. A parietally localized probability effect (right greater than left) was found in the N3 amplitude. The dN3 was insensitive to the probability manipulation. In accord with its insensitivity to stimulus probability, the dN3 seems to be more related to the N400 than to the N2b. Posterior attentional processes sensitive to the allocation of attentional resources may have contributed to the topography of the dN3. The N3 is more related to the detection of expectation violation, while the P3 reflects the ease of identifying stimulus categories. PMID- 15653301 TI - An event-related potentials study of biological motion perception in human infants. AB - To clarify the dynamical processing aspect of biological motion (BM) perception from a developmental point of view, we measured event-related potentials (ERPs) in 8-month-old infants during the perception of BM or a scrambled motion (SM; randomization of BM's spatial structure). We found that activation of the right hemisphere in 8-month-old infants was similar to that of adults, suggesting that the neural substrates for processing BM perception begin to mature at around 8 months of age. PMID- 15653303 TI - Evolutionary perspectives on innate immunity from the study of Caenorhabditis elegans. AB - Genetic and functional genomic approaches have begun to define the molecular determinants of pathogen resistance in Caenorhabditis elegans. Conserved signal transduction components are required for pathogen resistance, including a Toll/IL 1 receptor domain adaptor protein that functions upstream of a conserved p38 MAP kinase pathway. We suggest that this pathway is an ancestral innate immune signaling pathway present in the common ancestor of nematodes, arthropods and vertebrates, which is likely to predate the involvement of canonical Toll signaling pathways in innate immunity. We anticipate that the study of pathogen resistance in C. elegans will continue to provide evolutionary and mechanistic insights into the signal transduction and physiology of innate immunity. PMID- 15653304 TI - Sensing and signaling during infection in Drosophila. AB - Most of the progress in dissecting the Drosophila antimicrobial response over the past decade has centered around intracellular signaling pathways in immune response tissues and expression of genes encoding antimicrobial peptide genes. The past few years, however, have witnessed significant advances in our understanding of the recognition of microbial invaders and subsequent activation of signaling cascades. In particular, the roles of peptidoglycan recognition proteins, which have known homologues in mammals, have been recognized and examined at the structural and functional levels. PMID- 15653305 TI - Ligand recognition by antigen-presenting cell C-type lectin receptors. AB - It is now appreciated that the range of ligands interacting with C-type lectin type receptors on antigen presenting cells includes endogenous self-molecules as well as pathogens and pathogen-derived ligands. Interestingly, not all interactions between these receptors and pathogenic ligands have beneficial outcomes, and it appears that some pathogens have evolved immunoevasive or immunosuppressive activities through receptors such as DC-SIGN. In addition to this, recent data indicate that the well-characterised macrophage mannose receptor is not essential to host defence against fungal pathogens, as previously thought, but has an important role in regulating endogenous glycoprotein clearance. New studies have also demonstrated that different ligand binding and/or sensing receptors collaborate for full and effective immune responses. PMID- 15653306 TI - Manipulation of innate immunity by bacterial pathogens. AB - The past decade has witnessed tremendous growth in two related fields: innate immunity and microbial pathogenesis. Many pathogens have evolved mechanisms to infect their hosts in the face of a fully functional innate immune system, and there are numerous examples by which pathogens avoid recognition and/or suppress inflammation. In this review, I suggest that pathogens not only survive the innate immune response, but use it to promote their pathogenesis. PMID- 15653307 TI - NK cells in innate immunity. AB - NK cells have an important role in innate immune responses, particularly in anti viral immunity. Recent studies have revealed a molecular basis for NK cell recognition of virus-infected cells, implicating the activating KIR and Ly49 receptors and NKG2D in this process. Additionally, mutual cooperation between NK cells and dendritic cells suggests that these innate cells can shape the nature of an adaptive immune response. These findings, as well as advances in understanding NK cell development and homeostasis, indicate that NK cell biology is more sophisticated than previously appreciated. PMID- 15653308 TI - Genetic dissection of innate immunity to infection: the mouse cytomegalovirus model. AB - Resistance to infection is largely inherited rather than acquired, and is encoded by a definable set of host genes designated the 'resistome'. Logically speaking, piecemeal disruption of the resistome gives us the best chance to define it, and the most spectacular advances in understanding innate immunity have grown from spontaneous or induced germline mutations of the resistome. Mutations induced by random germline mutagenesis have now become so numerous that we are nearly in a position to define the size of the resistome, and both random and targeted mutations give us a fairly nice sketch of its components and how they interact. Our own N-ethyl-N-nitrosourea mutagenesis effort, which recently showed that components of Toll-like receptor signaling are essential constituents of the arsenal against MCMV infections, validated the forward genetic approach as a powerful tool to define the resistome. PMID- 15653309 TI - Genetic dissection of immunity in leprosy. AB - Leprosy is a chronic infectious disease caused by Mycobacterium leprae that affects an estimated 700,000 new individuals each year. A strong contribution of host genetics to susceptibility to leprosy has long been suggested to account for the considerable variability observed between individuals exposed to M. leprae. As there is no relevant animal model for human leprosy, forward genetics is the main strategy used to identify the genes and, consequently, the immunological pathways involved in protective immunity to M. leprae. With respect to genome wide screens, a major breakthrough has been reported this year; variants in the regulatory region shared by PARK2 and PACRG have been identified as being common risk factors for leprosy. PMID- 15653310 TI - Dissecting innate immune responses with the tools of systems biology. AB - Systems biology strives to derive accurate predictive descriptions of complex systems such as innate immunity. The innate immune system is essential for host defense, yet the resulting inflammatory response must be tightly regulated. Current understanding indicates that this system is controlled by complex regulatory networks, which maintain homoeostasis while accurately distinguishing pathogenic infections from harmless exposures. Recent studies have used high throughput technologies and computational techniques that presage predictive models and will be the foundation of a systems level understanding of innate immunity. PMID- 15653311 TI - Antigen processing and recognition. PMID- 15653312 TI - Epigenetic control of MHC-II: interplay between CIITA and histone-modifying enzymes. AB - Recent advances have shown the crucial role of histone-modifying enzymes in controlling gene activation and repression. This led to the 'histone code' hypothesis, which proposes that combinations of histone modifications work in concert to affect specific gene expression. Mounting evidence suggests that the class II transactivator modulates promoter accessibility by coordinating the recruitment of chromatin modifiers in a time-dependent fashion. MHC-II expression is exquisitely controlled by these highly specific, coordinated and dynamic interactions at the promoter. PMID- 15653313 TI - DM and DO shape the repertoire of peptide-MHC-class-II complexes. AB - The presentation of antigenic peptides by MHC class II molecules is essential for activation of CD4+ T cells. The formation of most peptide-MHC-class-II complexes is influenced by the actions of two specialized accessory proteins--DM and DO- located in the endosomal/lysosomal system where peptide loading occurs. DM removes class-II-associated invariant-chain peptide (CLIP) from newly synthesized class II molecules, but by now it is clearly established that this is only a special case of the general peptide-editing function of DM. Recent data have begun to explain the molecular basis for the editing activity. The other accessory protein, DO, modulates DM activity in vitro, but the physiological importance of DO is unclear. New evidence from several laboratories has provided clues that may soon change this. PMID- 15653314 TI - Viral immune evasion molecules attack the ER peptide-loading complex and exploit ER-associated degradation pathways. AB - The CD8+ cytotoxic-T-cell response is a potent mechanism that controls intracellular pathogens, including many viruses. To facilitate transmission, viruses often counter this response by inhibiting the cell surface display of virus-derived peptides on MHC class I molecules. More specifically, recent studies have demonstrated that viruses have evolved remarkable mechanisms to inhibit MHC class I expression by interfering with the function of the MHC class I assembly machinery (the peptide-loading complex) in the endoplasmic reticulum and/or by exploiting endoplasmic-reticulum-associated degradation pathways. These viral molecules are proving invaluable as research tools to illuminate the novel features of physiological pathways that are central to normal cell biology. Furthermore, the detailed characterization of such pathways has yielded significant new insights into host-pathogen interplay. PMID- 15653315 TI - Antigen presentation and recognition in bacterial infections. AB - Antigen processing and recognition is a key feature of antibacterial immune responses to intracellular bacteria. In contrast to viruses, which are primarily controlled by conventional MHC II- and MHC I-restricted CD4+ or CD8+ T cells, respectively, unconventional T cells participate additionally in antibacterial protection. These unconventional T cells include glycolipid-specific CD1 restricted T cells and phospholigand-specific gammadelta T cells. We are just beginning to understand the broad spectrum of antigen recognition and stimulation of distinct T-cell populations by bacterial pathogens. From the host perspective, a broad spectrum of different T-cell populations that recognize proteins, lipids and carbohydrates strengthens protective immunity. From the perspective of the pathogen, antigen presentation represents a bottleneck that should be exploited for evasion from, or devastation of, acquired immunity. Although several such mechanisms have been described in viral systems, few have thus far been elucidated in bacterial infections. PMID- 15653316 TI - CD1 assembly and the formation of CD1-antigen complexes. AB - The CD1 antigen presentation system presents lipid antigens to effector T cells, which have diverse roles in antimicrobial responses, antitumor immunity and in regulating the balance between tolerance and autoimmunity. The trafficking of CD1 molecules and lipid antigens facilitates their intersection and binding in specific intracellular compartments. Recent studies have now identified unexpected accessory molecules that are critical to CD1 assembly and lipid loading. The atomic structures of CD1-antigen complexes have defined both the orientation of polar headgroups between the alpha1 and alpha2 helices of CD1 and the manner in which distinct CD1 isoforms bind a range of lipids that have different lengths and numbers of hydrocarbon chains. PMID- 15653317 TI - A20 inhibits NF-kappaB activation by dual ubiquitin-editing functions. AB - Deregulation of the transcription factor NF-kappaB can mediate several inflammatory diseases in addition to cancer. Therefore, several proteins, including the zinc finger protein A20, tightly control its activation. Recently, the underlying mechanism by which A20 downregulates NF-kappaB activation in response to the pro-inflammatory cytokine tumor necrosis factor (TNF) has been described. A20 was shown to exert two opposing activities: sequential de ubiquitination and ubiquitination of the TNF receptor-interacting protein (RIP), thereby targeting RIP to proteasomal degradation. PMID- 15653318 TI - Structures of cytochrome P450 3A4. AB - Cytochrome P450 3A4 (CYP3A4) catalyzes the initial step in the clearance of many pharmaceuticals and foreign chemicals. The structurally diverse nature of CYP3A4 substrates complicates rational prediction of their metabolism and identification of potential drug interactions. The first molecular structures of human CYP3A4 were recently determined, revealing an active site of sufficient size and topography to accommodate either large ligands or multiple smaller ligands, as suggested by the heterotropic and homotropic cooperativity of the enzyme. PMID- 15653319 TI - SAGA unveiled. AB - Transcriptional regulation in eukaryotes is intimately coupled to chromatin dynamics. The SAGA (Spt-Ada-Gcn5) histone acetyltransferase (HAT) complex of Saccharomyces cerevisiae is a multi-subunit co-factor for RNA polymerase II transcription. However, not all gene activation events require its intrinsic HAT activity. In addition, SAGA subunits can also restrict gene transcription. The recently published structural model from the laboratories of Fred Winston and Patrick Schultz of the SAGA complex provides a framework to rationalize these findings and to direct further investigation of this crucial transcriptional co factor. PMID- 15653320 TI - The importance of uniformity in reporting protein-function data. PMID- 15653321 TI - Protein folding and the organization of the protein topology universe. AB - The mechanism by which proteins fold to their native states has been the focus of intense research in recent years. The rate-limiting event in the folding reaction is the formation of a conformation in a set known as the transition-state ensemble. The structural features present within such ensembles have now been analysed for a series of proteins using data from a combination of biochemical and biophysical experiments together with computer-simulation methods. These studies show that the topology of the transition state is determined by a set of interactions involving a small number of key residues and, in addition, that the topology of the transition state is closer to that of the native state than to that of any other fold in the protein universe. Here, we review the evidence for these conclusions and suggest a molecular mechanism that rationalizes these findings by presenting a view of protein folds that is based on the topological features of the polypeptide backbone, rather than the conventional view that depends on the arrangement of different types of secondary-structure elements. By linking the folding process to the organization of the protein structure universe, we propose an explanation for the overwhelming importance of topology in the transition states for protein folding. PMID- 15653322 TI - Game-theoretical approaches to studying the evolution of biochemical systems. AB - Evolutionary optimization has been successfully used to increase our understanding of key properties of biochemical systems. Traditional optimization is, however, often insufficient for gaining deeper insights into the evolution of such systems because usually there is a mutual relationship between the properties optimized by evolution and the properties of the environment. Thus, by evolving towards optimal properties, organisms change their environment, which in turn alters the optimum. Evolutionary game theory provides an appropriate framework for analyzing evolution in such 'dynamic fitness landscapes'. We therefore argue that it is a promising approach to studying the evolution of biochemical systems. Indeed, recent studies have applied evolutionary game theory to key issues in the evolution of energy metabolism. PMID- 15653323 TI - Manipulating proteins with chemistry: a cross-section of chemical biology. AB - Chemistry-driven strategies for modifying, controlling and monitoring protein function in vitro and in vivo have attracted widespread interest among chemists in recent years. Several strategies have now emerged that complement standard genetics-based approaches, and they are being increasingly adopted by biologists to address issues in relevant contexts from cells to animals. With the development of these chemical biology tools, we might be approaching a time when detailed quantitative analysis of protein function, to a degree previously available only in reconstituted systems, is attainable in an in vivo setting. PMID- 15653324 TI - Restraining PI3K: mTOR signalling goes back to the membrane. AB - The lipid kinase phosphoinositide 3-kinase (PI3K) is activated in response to various extracellular signals including peptide growth factors such as insulin and insulin-like growth factors (IGFs). Phosphatidylinositol (3,4,5) trisphosphate [PtdIns(3,4,5)P(3)] generated by PI3K is central to the diverse responses elicited by insulin, including glucose homeostasis, proliferation, survival and cell growth. The actions of lipid phosphatases have been considered to be the main means of attenuating PI3K signalling, whereby the principal 3 phosphatase - phosphatase and tensin homologue deleted on chromosome 10 (PTEN) - dephosphorylates PtdIns(3,4,5)P(3), reversing the action of PI3K. Recently, however, another pathway of regulation of PI3K has been identified in which activation of PI3K itself is prevented. This finding, together with earlier work, strongly suggests that a major form of negative feedback inhibition of PI3K results from activated growth signalling via mammalian target of rapamycin (mTOR) and the p70 S6 kinase (S6K) - a pathway that could have consequences for the development of type 2 diabetes and tuberous sclerosis complex. PMID- 15653325 TI - Phosphorylation of NF-kappaB and IkappaB proteins: implications in cancer and inflammation. AB - Nuclear factor-kappaB (NF-kappaB) is a transcription factor that has crucial roles in inflammation, immunity, cell proliferation and apoptosis. Activation of NF-kappaB mainly occurs via IkappaB kinase (IKK)-mediated phosphorylation of inhibitory molecules, including IkappaBalpha. Optimal induction of NF-kappaB target genes also requires phosphorylation of NF-kappaB proteins, such as p65, within their transactivation domain by a variety of kinases in response to distinct stimuli. Whether, and how, phosphorylation modulates the function of other NF-kappaB and IkappaB proteins, such as B-cell lymphoma 3, remains unclear. The identification and characterization of all the kinases known to phosphorylate NF-kappaB and IkappaB proteins are described here. Because deregulation of NF kappaB and IkappaB phosphorylations is a hallmark of chronic inflammatory diseases and cancer, newly designed drugs targeting these constitutively activated signalling pathways represent promising therapeutic tools. PMID- 15653326 TI - Substrate-induced conformational changes in glycosyltransferases. AB - Oligosaccharide chains of glycoproteins, glycolipids and glycosaminoglycans are synthesized by glycosyltransferases by the transfer of specific glycosyl moieties from activated sugar-nucleotide donors to specific acceptors. Structural studies on several of these enzymes have shown that one or two flexible loops at the substrate-binding site of the enzymes undergo a marked conformational change from an open to a closed conformation on binding the donor substrate. This conformational change, in which the loop acts as a lid covering the bound donor substrate, creates an acceptor-binding site. After the glycosyl unit is transferred from the donor to the acceptor, the saccharide product is ejected and the loop reverts to its native conformation, thereby releasing the remaining nucleotide moiety. The specificity of the sugar donor is determined by a few residues in the sugar-nucleotide-binding pocket of the enzyme that are conserved among the family members from different species. PMID- 15653327 TI - N-acyl-N-alkyl-sulfonamide anchors derived from Kenner's safety-catch linker: powerful tools in bioorganic and medicinal chemistry. AB - In 1971 Kenner et al. introduced the safety-catch principle into solid phase peptide synthesis. Thus two contradicting needs were addressed. On the one hand, sufficient stability of the linker substrate bond to impede hydrolysis or similar side reactions, on the other hand mild chemical conditions allowing for unscathed liberation of the precious products. Over the years this linker type emerged in several different chemical disciplines and nowadays it presents a useful and broadly applicable tool. Recent advancements and applications based on Kenner's safety-catch linker are reviewed. PMID- 15653328 TI - A radial distribution function approach to predict A(2B) agonist effect of adenosine analogues. AB - The radial distribution function (RDF) approach has been applied to the study of the A(2B) agonist effect of a set of 89 adenosine analogues reported with this activity. A model able to describe more than 70% of the variance in the experimental activity was developed with the use of the mentioned approach. In contrast, none of the eleven different approaches including the use of Constitutional, Topological, Molecular walk count, BCUT, Galvez topological charge indices, 2D autocorrelations, Randic molecular profiles, Geometrical, 3D Morse, WHIM and GETAWAY descriptors was able to explain more than 47% of the variance in the mentioned property with the same number of descriptors. PMID- 15653329 TI - Synthesis and evaluation of cis-hexahydropyrrolo[3,2-b]pyrrol-3-one peptidomimetic inhibitors of CAC1 cysteinyl proteinases. AB - A stereoselective synthesis of functionalised cis-hexahydropyrrolo[3,2-b]pyrrol-3 ones has been developed through Fmoc and Cbz-protected intermediates 5 and 6. Building blocks 5 and 6 were prepared via the intramolecular cyclisation of anti epoxide 17. The intramolecular reaction occurred exclusively through the anti epoxide to provide the 5,5-cis-fused bicycle, whereas the syn-epoxide, which theoretically would provide the 5,5-trans-fused bicycle, remained unchanged. These experimental observations are consistent with a key design element that we have introduced within this novel bicyclic ketone scaffold. Our bicyclic design strategy provides chiral stability to the bridgehead stereocentre that is situated alpha to the ketone because the cis-fused geometry is both thermodynamically and kinetically stable. Building blocks 5 and 6 have been utilised in both solid phase and solution phase syntheses of peptidomimetics 22, 36-40, which exhibit potent in vitro inhibition against a range of CAC1 cysteinyl proteinases. Compound 22, a potent and selective inhibitor of human cathepsin K exhibited good primary DMPK properties along with promising activity in an in vitro cell-based human osteoclast assay of bone resorption. PMID- 15653330 TI - An efficient asymmetric synthesis of (S)-atenolol: using hydrolytic kinetic resolution. AB - Enantiomerically pure (S)-atenolol was prepared by using (R,R) salen Co(III) complex for the resolution of terminal epoxide. This process was carried out at room temperature in excellent enantio selectivity. The method can be applied for large-scale preparation of (S)-atenolol without any problem. PMID- 15653331 TI - Novel diterpenoids with potent inhibitory activity against endothelium cell HMEC and cytotoxic activities from a well-known TCM plant Daphne genkwa. AB - Twelve highly oxygenated novel daphnane-type diterpenoids genkwanines A-L (1-12), together with four known diterpenes (13-16), were isolated from the bud of Daphne genkwa, a well-known traditional Chinese medicine (TCM). The new structures were elucidated on the basis of spectral methods, especially 2D NMR spectra (HMQC, HMBC, NOESY). Inhibitory activity against endothelium cell HMEC proliferation and cytotoxic activities against two tumor cell lines were assessed for all the compounds 1-16, and found to be significant and structure related. A number of compounds showed very potent cytotoxic activities against two tumor cell lines at the IC50 levels of 0.15-8.40 microM, and most interestingly, five of the compounds 4, 8, 10, 13, and 14 exhibited strong activity to inhibit the endothelium cell HMEC at the IC50 levels of 2.90-15.0 microM. PMID- 15653332 TI - Polybrominated diphenyl ethers from a sponge of the Dysidea genus that inhibit Tie2 kinase. AB - Tie2 kinase, an enzyme that supports angiogenesis essential for tumor growth and survival, was selected as a target in a search for naturally occurring inhibitors of potential utility for antitumor therapy. Two polybrominated diphenyl ethers, 3,5-dibromo-2-(2',4'-dibromophenoxy)phenol (1) and 4,6-dibromo-2-(2',4' dibromophenoxy)phenol (2) were isolated from an extract prepared from Dysidea sp. after bioassay-guided fractionation. PMID- 15653333 TI - Antileishmanial activity, cytotoxicity and QSAR analysis of synthetic dihydrobenzofuran lignans and related benzofurans. AB - A series of synthetic dihydrobenzofuran lignans and related benzofurans were evaluated for their cytotoxicity in a screening panel consisting of various human tumour cell lines, and for their antiprotozoal activity against L. donovani (axenic amastigotes), chloroquine resistant Plasmodium falciparum (strain K1), Trypanosoma brucei rhodesiense and T. cruzi, and for cytotoxicity on L6 cells. No promising cytotoxicities against human tumour cell lines were observed for newly synthesised compounds, but the dimerisation product of some lipophylic esters of caffeic acid, such as compound 2g, showed a high activity against chloroquine resistant P. falciparum (strain K1) (IC50 0.43 microg/mL) and L. donovani (axenic amastigotes) (IC50 0.12 microg/mL), which was confirmed in an infected macrophage assay (IC50 0.19 microg/mL). QSAR models for the cytotoxic and antileishmanial activity were generated using Quasar receptor surface modelling. PMID- 15653334 TI - A structure-taste study of arylsulfonyl(cyclo)alkanecarboxylic acids. AB - A number of sweeteners contain a sulfonyl group. In our current search for new glucophores several new compounds containing such group were obtained. A series of novel 1-phenylsulfonylcyklohexanecarboxylic acids and 2 arylsulfonylalkanecarboxylic acids was obtained and evaluated for their sweet taste quality. It has been found that methyl substituents are of the key importance for the activity of these compounds. PMID- 15653335 TI - Design, synthesis, and evaluation of potent and selective benzoyleneurea-based inhibitors of protein geranylgeranyltransferase-I. AB - A series of novel protein geranylgeranyltransferase-I (PGGTase-I) inhibitors based on a benzoyleneurea scaffold has been synthesized. Using a benzoyleneurea scaffold as a mimetic for the central dipeptide (AA), we have developed CAAX peptidomimetic inhibitors that selectively block the activity of PGGTase-I over the closely related enzyme protein farnesyltransferase. In this new class of PGGTase-I inhibitors, compound (6c) with X=L-phenylalanine, displayed the highest inhibition activity against PGGTase-I with an IC50 value of 170 nM. The inhibitors described in this study represent novel and promising leads for the development of potent and selective inhibitors of mammalian PGGTase-I for potential application as antitumor agents. PMID- 15653336 TI - Synthesis, biological evaluation, and molecular modeling of novel thioacridone derivatives related to the anticancer alkaloid acronycine. AB - The well-reported, but moderate antitumor activity of the acronycine alkaloid led us to synthesize a novel series of thioacridone compounds related to acronycine, as potential anticancer agents. Compounds were designed either as DNA intercalating agents, or as DNA intercalating agents with covalent bond forming potential. Bathochromic shifts of the compounds upon complexation with salmon testis DNA suggested intercalation as the mode of DNA binding. The binding interaction of the compounds was found to be approximately 10(2) M(-1), with that of the most potent compound 1-(2-dimethylaminoethylamino)-9(10H)-thioacridone, 10(4) M(-1). In vitro cytotoxic activity (IC50) against HL-60 cells was found to range between 3.5 and 22 microg/mL. QSAR analyses yielded a multiple linear regression equation with an r2 of 0.847 for DNA binding and an r2 of 0.575 for cytotoxicity. The physicochemical parameters used in the QSAR analyses were logP, polar surface area, and calculated molar refractivity. Docking studies were also performed to compare the binding of the most potent and least potent compounds in the study in order to predict desirable chemical characteristics for further exploitation in drug design efforts. The thioacridone compounds in this series demonstrate cytotoxic activity in vitro that merit future in vivo evaluation. PMID- 15653337 TI - Structure identification and prophylactic antimalarial efficacy of 2 guanidinoimidazolidinedione derivatives. AB - The reported synthetic procedure of WR182393, a 2-guanidinoimidazolidinedione derivative with high prophylactic antimalarial activity, was found to be a mixture of three closely related products. Poor solubility of WR182393 in both water and organic solvents and its impractical synthetic method have made the purification and structure identification of the reaction mixture a highly challenging task. The problems were circumvented by prodrug approach involving carbamate formation of the mixture, which enhances the solubility of the mixture in common organic solvents and facilitates the separation and structure determination of the two products. The structures of the two components were determined by X-ray crystallography and NMR of their corresponding carbamates 3a and 4a. Additional alkyl carbamates were prepared according to the same approach and two new carbamates 3b and 4d were found to possess higher intramuscular (im) efficacy than the parent compound WR182393 against Plasmodium cynomolgi in Rhesus monkey. PMID- 15653338 TI - 2-aryl-8-chloro-1,2,4-triazolo[1,5-a]quinoxalin-4-amines as highly potent A1 and A3 adenosine receptor antagonists. AB - Some 2-aryl-8-chloro-1,2,4-triazolo[1,5-a]quinoxaline derivatives 2-18, obtained by introducing different substituents on either the 4-amino moiety (acyl or carbamoyl groups) or the 2-phenyl ring (4-OCH3) of previously reported 8-chloro-2 phenyl-1,2,4-triazolo[1,5-a]quinoxalin-4-amine (1), have been synthesized and tested in radioligand binding assays at bovine A1 and A(2A) and at cloned human A1 and A3 adenosine receptors. The rationally designed 8-chloro-2-(4-methoxy phenyl)-1,2,4-triazolo[1,5-a]quinoxalin-4-acetylamine (14) can be considered one of the most potent and hA3 versus hA1 selective AR antagonists reported till now. The structure-activity relationships of compounds 2-18 are in agreement with those of previously reported 2-aryl-1,2,4-triazolo[4,3-a]quinoxalines (series A) and 2-arylpyrazolo[3,4-c]quinolines (series B), thus suggesting a similar AR binding mode. In fact, the importance for the A3 receptor-ligand interaction of both a strong acidic NH proton donor and a C=O proton acceptor at position-4, able to engage hydrogen-bonding interactions with specific sites on the A3 AR, has been confirmed. Using our recently published hA3 receptor model, to better elucidate our experimental results, we decided to theoretically depict the putative TM binding motif of the herein reported 1,2,4-triazolo[1,5-a]quinoxaline derivatives on human A3 receptor. Structure-activity relationships have been explained analyzing the three-dimensional structure of the antagonist-receptor models obtained by molecular docking simulation. PMID- 15653339 TI - Synthesis and structure-activity relationships of 6-{4-[(3 fluorobenzyl)oxy]phenoxy}nicotinamide derivatives as a novel class of NCX inhibitors: a QSAR study. AB - The sodium-calcium exchanger (NCX) transports Na+ and Ca2+ ions, and controls the Ca2+ concentration in myocytes. Calcium overload is induced via activation of reverse NCX, and is responsible for reperfusion injury in heart failure. Hence, NCX is an attractive target for prevention and treatment of reperfusion arrhythmias, myocardial contracture, and necrosis. We have synthesized a series of 6-{4-[(3-fluorobenzyl)oxy]phenoxy}nicotinamide derivatives, and evaluated their inhibitory activity against the reverse and forward modes of NCX. N-(3 Aminobenzyl)-6-{4-[(3-fluorobenzyl)oxy]phenoxy}nicotinamide (8) was shown to be a potent inhibitor of reverse NCX activity, with an IC50 value of 0.24 microM. A QSAR study showed that inhibition of reverse NCX activity by 6-{4-[(3 fluorobenzyl)oxy]phenoxy}nicotinamide derivatives is multiply dependent on the hydrophobicity (pi) and the shape (B(iv)) of the substituent at the 3-position of the phenyl ring. PMID- 15653340 TI - Synthesis and structure-activity relationships of benzyloxyphenyl derivatives as a novel class of NCX inhibitors: effects on heart failure. AB - In the context of heart failure and myocardial ischemia reperfusion, the activity of the sodium-calcium exchanger can lead to calcium overload, which in turn can lead to contractile dysfunction and arrhythmia. Therefore, NCX is an attractive target for treatment of heart failure and myocardial ischemia reperfusion. We have designed and synthesized a series of benzyloxyphenyl derivatives as potential NCX inhibitors, based on compound 4. These derivatives have been evaluated for their inhibitory activity against both the reverse and forward modes of NCX, and two novel potent NCX inhibitors (7i, 10a) were discovered. Compound 7i was evaluated for its efficacy on ouabain-induced tonotropy and arrhythmia in a heart-failure model. PMID- 15653341 TI - Synthesis and in vivo biodistribution of BPA-Gd-DTPA complex as a potential MRI contrast carrier for neutron capture therapy. AB - p-boronophenylalanine (BPA) conjugated Gd-DTPA complex (3) was synthesized from the active methyne compound 6, the allylic carbonate 7, and BPA by the palladium catalyzed allylation reaction followed by the DCC coupling reaction. The in vivo biodistribution of complex 3 was evaluated by prompt gamma-ray analysis and alpha autoradiography using the tumor-bearing rats. High accumulation of gadolinium was observed in the kidney and the %ID values were 0.17 and 0.088 at 20 and 60 min after injection of 3, respectively. The accumulation was also observed in the tumor and the %ID values were 0.010 and 0.0025 at 20 and 60 min after injection, respectively. The visualization experiment of boron distribution in the tumor bearing rat by alpha-autoradiography indicates that boron was accumulated in the tumor and the intestines at 20 min after injection. PMID- 15653342 TI - Synthesis and in vitro evaluation of new benzovesamicol analogues as potential imaging probes for the vesicular acetylcholine transporter. AB - Our goal was to synthesize new stereospecific benzovesamicol analogues, which could potentially be used as SPECT or PET radioligands for the vesicular acetylcholine transporter (VAChT). This paper describes the chemical synthesis, resolution and determination of binding affinity for four enantiomeric pairs of derivatives. Their intrinsic affinities were determined by competition against binding of [3H]vesamicol to human VAChT. Of the eight enantiomers, (E)-(R,R)-5 AOIBV [(R,R)-3], and (R,R)-5-FPOBV [(R,R)-4] displayed the highest binding affinities for VAChT (Kd=0.45 and 0.77 nM, respectively), which indicated that an elongation of the chain from 5-idodo as in the case of 5-iodobenzovesamicol (5 IBVM), to a 5-(E)-3-iodoallyloxy or 5-fluoropropoxy substituent, as in 5-AOIBV and 5-FPOBV, respectively, was very well tolerated at the vesamicol binding site. The enantiomer (R,R)-4-MAIBV [(R,R)-16], which retains the basic structure of (-) 5-IBVM but possess an additional aminomethyl substituent in the 4-position of the piperidine ring, displayed lower binding affinity (Kd=8.8 nM). Nevertheless, the result suggests that substitution at this position may be an interesting alternative to investigate for development of new benzovesamicol analogues. As expected, the corresponding (S,S) enantiomers displayed lower Kd values, they were approximately 10-fold lower in the case of (S,S)-5-FPOBV (Kd=8.4 nM) and (E) (S,S)-5-AOIBV (Kd=4.3 nM). (R,R)-3, and (R,R)-4 showed the same high affinity for VAChT as (-)-5-IBVM and may be suitable as imaging agents of cholinergic nerve terminals. PMID- 15653343 TI - Cyclic sulfamide HIV-1 protease inhibitors, with sidechains spanning from P2/P2' to P1/P1'. AB - Previous studies of HIV protease inhibitors have shown that it is possible to elongate the P1/P1' sidechains to reach the S3/S3' binding sites. By analogy, we expected that it would be possible to design inhibitors reaching between the S1/S1' and S2/S2' binding sites. Molecular modeling suggested that this could be achieved with appropriate ortho-substitution of the P2/P2' benzyl groups in our cyclic sulfamide inhibitors. Four different spacer groups were investigated. The compounds were smoothly prepared from tartaric acid in five steps and exhibit low to moderate activity, the most potent inhibitor possessing a Ki value of 0.53 microM. PMID- 15653344 TI - Binding of beta-D-glucopyranosyl bismethoxyphosphoramidate to glycogen phosphorylase b: kinetic and crystallographic studies. AB - In an attempt to identify a new lead molecule that would enable the design of inhibitors with enhanced affinity for glycogen phosphorylase (GP), beta-D glucopyranosyl bismethoxyphosphoramidate (phosphoramidate), a glucosyl phosphate analogue, was tested for inhibition of the enzyme. Kinetic experiments showed that the compound was a weak competitive inhibitor of rabbit muscle GPb (with respect to alpha-D-glucose-1-phosphate (Glc-1-P)) with a Ki value of 5.9 (+/-0.1) mM. In order to elucidate the structural basis of inhibition, we determined the structure of GPb complexed with the phosphoramidate at 1.83 A resolution. The complex structure reveals that the inhibitor binds at the catalytic site and induces significant conformational changes in the vicinity of this site. In particular, the 280s loop (residues 282-287) shifts 0.4-4.3 A (main-chain atoms) to accommodate the phosphoramidate, but these conformational changes do not lead to increased contacts between the inhibitor and the protein that would improve ligand binding. PMID- 15653345 TI - Design, synthesis, and evaluation of novel bifunctional iron-chelators as potential agents for neuroprotection in Alzheimer's, Parkinson's, and other neurodegenerative diseases. AB - Several novel antioxidant-iron chelators bearing 8-hydroxyoxyquinoline moiety were synthesized, and various properties related to their iron chelation, and neuroprotective action were investigated. All the chelators exhibited strong iron(III) chelating and high antioxidant properties. Chelator 9 (HLA20), having good permeability into K562 cells and moderate selective MAO-B inhibitory activity (IC50 110 microM), displayed the hightest protective effects against differentiated P19 cell death induced by 6-hydroxydopamine. EPR studies suggested that Chelator 9 also act as radical scavenger to directly scavenge hydroxyl radical. PMID- 15653346 TI - Synthesis and biological activity of sulphostin analogues, novel dipeptidyl peptidase IV inhibitors. AB - The structure of sulphostin (1), a novel dipeptidyl peptidase IV (DPP-IV) inhibitor, is consisted of three key functional groups, including a characteristic amino(sulfoamino)phosphinyl group, on a piperidine ring. To examine the relationship between its structure and the inhibitory activity against DPP-IV, various analogues were synthesized and their activities were investigated. These results indicated that all of the functional groups on the piperidine ring were crucial to the DPP-IV inhibitory activity of sulphostin, and that the sulfonic acid group, which constructed the amino(sulfoamino)phosphinyl group, contributed to the stability of the compound. Moreover, those functional groups should be adjoined on the piperidine ring for the inhibitory activity. The size of the nitrogen-containing heterocyclic ring including piperidine appeared to scarcely affect the activity. In the present study, the sulfonic acid deficient five-membered ring analogue 27a showed the strongest inhibitory activity (IC50=11 nM). PMID- 15653347 TI - Design of novel nicotinic ligands through 3D database searching. AB - A series of quinoline derivatives have been designed on the basis of results from a 3D search of the Cambridge Structural Database using the nicotinic pharmacophore as a query and further modified using molecular modeling. Some of the synthesized compounds show nanomolar affinity for the central nicotinic receptor on rat cerebral cortex. PMID- 15653348 TI - Design, synthesis and photobiological properties of 3,4-cyclopentenepsoralens. AB - The QSAR directed synthesis of tetracyclic psoralen derivatives (3-5) characterised by the condensation of a cyclopentane ring at the level of the 3,4 double bond of the tricyclic psoralen moiety is reported. The new compounds present a methoxy (3), a hydroxy (4) or a dimethylaminopropoxy (5) side chain inserted in position 8 of the lead chromophore. The evaluation of photoantiproliferative activity on human tumour cell lines reveals for 5 an ability to inhibit cell growth significantly higher with respect to that of the reference drug, 8-MOP. Interestingly, the enhancement in antiproliferative activity is accompanied by the disappearance of skin phototoxicity. On the other hand, no significant photobiological activity was scored for 3 and 4. The ability to photoreact with DNA, evaluated by isolating the 4',5' monoadduct and by estimating the ability to form interstrand cross-links, appeared to be significant for 5, practically negligible for 3 and 4. Furthermore, a back projection of the more active compound identifies structural features suitable for further synthetic modifications. PMID- 15653349 TI - Synthesis, antitumour activity and structure-activity relationships of 5H benzo[b]carbazoles. AB - A series of novel 5H-benzo[b]carbazoles related to the ellipticines was obtained from the reactions of p-benzoquinones with 2-aminomethylene-1-indanones. Most of the compounds were evaluated for their antitumour activity in the National Cancer Institute's in vitro human tumour cell line screening panel. Among them, particularly derivative 15c bearing a p-quinone methide moiety in ring C of the heterocycle was found to show in vitro activity comparable to clinically well established anticancer agents such as amsacrine or mitomycin C. Compounds 9d, 9e and 12k showed increased potency to distinct cell lines like the leukemia or melanoma subpanel of cell lines. Based on the test results, structure-activity relationships for this series of compounds were developed. For instance, it was found that a quinonoid substructure in ring C leads to a noticeable increase in activity. The same observation was made for a 2-hydroxyl substituent at the ring system. 2-Acetoxy and 2-methoxy derivatives as well as 2-unsubstituted 5H benzo[b]carbazoles either had a decreased potency or were found to be inactive. A COMPARE analysis with some of these compounds showed poor or no correlation with anticancer drugs of the NCI's standard agents database indicating a novel mechanism of action. Additionally, UV-vis titrations in the series of 5H benzo[b]carbazoles indicated interactions with calf thymus DNA only for the highly active quinone methide 15c. PMID- 15653350 TI - Concise syntheses of arabinogalactans with beta-(1-->6)-linked galactopyranose backbones and alpha-(1-->3)- and alpha-(1-->2)-linked arabinofuranose side chains. AB - 4-methoxyphenyl glycosides of 2,3''-bis-alpha-L-arabinofuranosyl branched beta-D (1-->6)-linked galactopyranosyl tetraose (16), 3',2''''-bis-alpha-L arabinofuranosyl branched beta-D-(1-->6)-linked galactopyranosyl hexaose (27), and a twentyose (42) consisting of beta-(1-->6)-linked D-galactopyranosyl pentadecaoligosaccharide backbone with alpha-L-arabinofuranosyl side chains alternately attached at C-2 and C-3 of the middle galactose residue of each consecutive beta-(1-->6)-linked galactotriose unit of the backbone, were synthesized with isopropyl 3-O-allyl-2,4-di-O-benzoyl-1-thio-beta-D galactopyranoside (6), 2,3,4,6-tetra-O-benzoyl-alpha-D-galactopyranosyl trichloroacetimidate (7), 2,3,5-tri-O-benzoyl-alpha-L-arabinofuranosyl trichloroacetimidate (12), 6-O-acetyl-2,3,4-tri-O-benzoyl-alpha-D galactopyranosyl trichloroacetimidate (17), 4-methoxyphenyl 2,3,4-tri-O-benzoyl beta-D-galactopyranoside (19), and 2,6-di-O-acetyl-3,4-di-O-benzoyl-alpha-D galactopyranosyl trichloroacetimidate (28) as the key synthons. Condensation of 6 with 7 gave the disaccharide donor 8, and subsequent condensation of 8 with 4 methoxyphenyl 2,3,4-tri-O-benzoyl-beta-D-galactopyranosyl-(1-->6)-2-O-acetyl-3,4 di-O-benzoyl-beta-D-galactopyranoside (9) followed by selective deacetylation afforded the tetrasaccharide acceptor 11. Coupling of 11 with 12 gave the pentasaccharide 13, its deallylation followed by coupling with 12, and debenzoylation gave the hexasaccharide 16 with beta-(1-->6)-linked galactopyranose backbone and 2- and 3''-linked alpha-L-arabinofuranose side chains. The octasaccharide 27 was similarly synthesized, while the twentyoside 42 was synthesized with tetrasaccharides 33 or 24 as the donors and 23, 36, 38, and 40 as the acceptors by consecutive couplings followed by deacylation. PMID- 15653351 TI - Discovery of 1,7-cyclized indoles as a new class of potent and highly selective human beta3-adrenergic receptor agonists with high cell permeability. AB - The synthesis and evaluation of a novel series of 1,7-cyclized indole-based human adrenergic receptor (beta3-AR) agonists are reported. The synthesis of a variety of 1,7-cyclized indole part was accomplished by the Mitsunobu reaction or a ring closing metathesis (RCM) reaction. SAR studies revealed that expansion of the ring size resulted in considerable selectivity against the beta1- and beta2-ARs. Compound 26, an eight-membered ring analogue with a double bond on its 1,7-linker portion, was found to be a potent beta3-AR agonist (EC50 = 0.75 nM, IA = 90%) with extremely high selectivity for the beta3-AR over the beta1- and beta2-ARs. PMID- 15653352 TI - Evaluation of 18F-labeled acetylcholinesterase substrates as PET radiotracers. AB - Four 18F-labeled acetylcholinesterase (AChE) substrates, (S)-N-[18F]fluoroethyl-2 piperidinemethyl acetate (1), (R)-N-[18F]fluoroethyl-3-pyrrolidinyl acetate (2), N-[18F]fluoroethyl-4-piperidinyl acetate (3), and (R)-N-[18F]fluoroethyl-3 piperidinyl acetate (4), were evaluated for in vivo blood and brain metabolism in mice, brain pharmacokinetics in rats monkeys (M. nemistrina) using PET imaging. All 18F-labeled compounds were compared to N-[11C]methyl-4-piperidinyl propionate (PMP). Compound 1 was completely metabolized within 1 min in mouse blood and brain. This compound had relatively fast regional brain pharmacokinetics and poor discrimination between brain regions with different AChE concentration. Compound 4 showed relatively slower blood metabolism and slower pharmacokinetics than compound 1 but again poor discrimination between brain regions. Both compounds 1 and 4 showed different kinetic profiles than PMP in PET studies. Compound 3 had the slowest blood metabolism and slower pharmacokinetics than PMP. Compound 2 showed highly encouraging characteristics with an in vivo metabolism rate, primate brain uptake, and regional brain pharmacokinetics similar to [11C]PMP. The apparent hydrolysis rate constant k3 in primate cortex was very close to that of [11C]PMP. This compound has potential to be a good PET radiotracer for measuring brain AChE activity. The longer lifetime of 18F would permit longer imaging times and allows preparation of radiotracer batches for multiple patients and delivery of the tracer to other facilities, making the technique more widely available to clinical investigators. PMID- 15653353 TI - Amino substituted derivatives of 5'-amino-5'-deoxy-5'-noraristeromycin. AB - The potent antiviral potential of 5'-amino-5'-deoxy-5'-noraristeromycin (2) is limited by associated toxicity. To seek derivatives of 2 that circumvent this undesirable property, three amino substituted derivatives (acetyl, 3; formyl, 4; and methyl, 5) of 2 have been prepared in 4-7 steps from the same intermediate, (1S,4R)-4-(6-chloropurin-9-yl)cyclopent-2-en-1-ol (6). Key steps involved an improved Pd(0)-catalyzed allylic azidation and a novel Pd(0)-catalyzed allylic amidation. The three target compounds were evaluated against a large number of viruses and found to be inactive except for a very weak effect of 5 on human cytomegalovirus, varicella zoster virus, and Epstein-Barr virus. There was also no noteworthy cytotoxicity associated with the new derivatives. Thus, these results indicate variation of the cyclopentyl amine of 2 does not offer a means to improve upon its antiviral potential. PMID- 15653354 TI - N-oxide analogs of WAY-100635: new high affinity 5-HT(1A) receptor antagonists. AB - WAY-100635 [N-(2-(1-(4-(2-methoxyphenyl)piperazinyl)ethyl))-N-(2 pyridinyl)cyclohexanecarboxamide] 1 and its O-desmethyl derivative DWAY 2 are well-known high affinity 5-HT(1A) receptor antagonists, which when labeled with carbon-11 (beta+; t(1/2) = 20.4 min) in the carbonyl group are effective radioligands for imaging brain 5-HT(1A) receptors with positron emission tomography (PET). In a search for new 5-HT(1A) antagonists with different pharmacokinetic and metabolic properties, the pyridinyl N-oxide moiety was incorporated into analogs of 1 and 2. NOWAY 3, in which the pyridinyl ring of 1 was oxidized to the pyridinyl N-oxide, was prepared via nucleophilic substitution of 2-[4-(2-methoxyphenyl)piperazin-1-yl]ethylamine on 2-chloropyridine-N-oxide followed by acylation with cyclohexanecarbonyl chloride. 6Cl-NOWAY 4, a more lipophilic (pyridinyl-6)-chloro derivative of 3, was prepared by treating 1-(2 methoxyphenyl)-4-(2-(2-(6-bromo)aminopyridinyl-N-oxide)ethyl)piperazine with cyclohexanecarbonyl chloride for acylation and concomitant chloro for bromo substitution. NEWWAY 5, in which the 2-hydroxy-phenyl group of 2 is replaced with a 2-pyridinyl N-oxide group with the intention of mimicking the topology of 2, was prepared in five steps from 2-(chloroacetylamino)pyridine. N-Oxides 3-5 were found to be high affinity antagonists at 5-HT(1A) receptors, with 3 having the highest affinity and a Ki value (0.22 nM) comparable to that of 1 (0.17 nM). By calculation the lipophilicity of 3 (LogP = 1.87) is lower than that of 1 by 1.25 LogP units while TLC and reverse phase HPLC indicate that 3 has slightly lower lipophilicity than 1. On the basis of these encouraging findings, the N-oxide 3 was selected for labeling with carbon-11 in its carbonyl group and for evaluation as a radioligand with PET. After intravenous injection of [carbonyl-11C]3 into cynomolgus monkey there was very low uptake of radioactivity into brain and no PET image of brain 5-HT(1A) receptors was obtained. Either 3 inadequately penetrates the blood-brain barrier or it is excluded from brain by an active efflux mechanism. Rapid deacylation of 3 was not apparent in vivo; in cynomolgus monkey plasma radioactive metabolites of [carbonyl-11C]3 appeared less rapidly than from the radioligands [carbonyl-11C]1 and [carbonyl-11C]2, which are known to be primarily metabolized by deacylation. Ligand 3 may have value as a new pharmacological tool, but not as a radioligand for brain imaging. PMID- 15653355 TI - Selective inhibitors of GABA uptake: synthesis and molecular pharmacology of 4-N methylamino-4,5,6,7-tetrahydrobenzo[d]isoxazol-3-ol analogues. AB - A series of lipophilic diaromatic derivatives of the glia-selective GABA uptake inhibitor (R)-4-amino-4,5,6,7-tetrahydrobenzo[d]isoxazol-3-ol [(R)-exo-THPO, 4] were synthesized via reductive amination of 3-ethoxy-4,5,6,7 tetrahydrobenzo[d]isoxazol-4-one (9) or via N-alkylation of O-alkylatedracemic 4. The effects of the target compounds on GABA uptake mechanisms in vitro were measured using a rat brain synaptosomal preparation or primary cultures of mouse cortical neurons and glia cells (astrocytes), as well as HEK cells transfected with cloned mouse GABA transporter subtypes (GAT1-4). The activity against isoniazid-induced convulsions in mice after subcutaneous administration of the compounds was determined. All of the compounds were potent inhibitors of synaptosomal uptake the most potent compound being (RS)-4-[N-(1,1-diphenylbut-1 en-4-yl)amino]-4,5,6,7-tetrahydrobenzo[d]isoxazol-3-ol (17a, IC50 = 0.14 microM). The majority of the compounds showed a weak preference for glial, as compared to neuronal, GABA uptake. The highest degree of selectivity was 10-fold corresponding to the glia selectivity of (R)-N-methyl-exo-THPO (5). All derivatives showed a preference for the GAT1 transporter, as compared with GAT2 4, with the exception of (RS)-4-[N-[1,1-bis(3-methyl-2-thienyl)but-1-en-4-yl]-N methylamino]-4,5,6,7-tetrahydrobenzo[d]isoxazol-3-ol (28d), which quite surprisingly turned out to be more potent than GABA at both GAT1 and GAT2 subtypes. The GAT1 activity was shown to reside in (R)-28d whereas (R)-28d and (S)-28d contributed equally to GAT2 activity. This makes (S)-28d a GAT2 selective compound, and (R)-28d equally effective in inhibition of GAT1 and GAT2 mediated GABA transport. All compounds tested were effective as anticonvulsant reflecting that these compounds have blood-brain barrier permeating ability. PMID- 15653356 TI - A new strategy for glycoprotein synthesis: ligation of synthetic glycopeptides with truncated proteins expressed in E. coli as TEV protease cleavable fusion protein. AB - We report here the use of TEV protease cleavable fusion proteins to produce glycosylated bioactive peptides and proteins. Bacterial expression was utilized to produce two fusion proteins, GPRT-C37-H6 and His-tagged interleukin-2 (amino acids 6-133), which when cleaved by the tobacco etch virus NIa protease (TEV protease) to generate HIV entry inhibitor peptide C37-H6 and a truncated version of the cytokine interleukin-2, both containing N-terminal cysteines. The N terminal cysteine containing C37-H6 and truncated interleukin-2 were then joined to a synthetic glycopeptide thioester utilizing native chemical ligation under nondenaturing and denaturing conditions, respectively. The ligations of the glycopeptide to the C37-H6 peptide and the truncated interleukin-2 protein both proceeded in high yield, though the size, and physical properties of the two polypeptides differ greatly. PMID- 15653358 TI - Optimization of the design and operation of FAIMS analyzers. AB - Field asymmetric waveform ion mobility spectrometry (FAIMS) holds significant promise for post-ionization separations in conjunction with mass-spectrometric analyses. However, a limited understanding of fundamentals of FAIMS analyzers has made their design and operation largely an empirical exercise. Recently, we developed an a priori simulation of FAIMS that accounts for both ion diffusion (including anisotropic components) and Coulomb repulsion, and validated it by extensive comparisons with FAIMS/MS data. Here it is corroborated further by FAIMS-only measurements, and applied to explore how key instrumental parameters (analytical gap width and length, waveform frequency and profile, the identity and flow speed of buffer gas) affect FAIMS response. We find that the trade-off between resolution and sensitivity can be managed by varying gap width, RF frequency, and (in certain cases) buffer gas, with equivalent outcome. In particular, the resolving power can be approximately doubled compared to "typical" conditions. Throughput may be increased by either accelerating the gas flow (preferable) or shortening the device, but below certain minimum residence times performance deteriorates. Bisinusoidal and clipped-sinusoidal waveforms have comparable merit, but switching to rectangular waveforms would improve resolution and/or sensitivity. For any waveform profile, the ratio of two between voltages in high and low portions of the cycle produces the best performance. PMID- 15653359 TI - Algorithm for accurate similarity measurements of peptide mass fingerprints and its application. AB - We present a simple algorithm which allows accurate estimates of the similarity between peptide fingerprint mass spectra from matrix assisted laser desorption/ionization (MALDI) spectrometers. The algorithm, which is a combination of mass correlation and intensity rank correlation, was used to cluster similar spectra and to generate consensus spectra from a data store of more than 100,000 spectra. The resulting first spectra library of 1248 unambiguously identified different protein digests was used to search for missed cleavage patterns that have not been reported so far and to shed light on some peptide ionization characteristics. The findings of this study could be directly implemented in peptide mass fingerprint search algorithms to decrease the false positive error rate to <0.25%. Furthermore, the results contribute to the understanding of the peptide ionization process in MALDI experiments. PMID- 15653360 TI - Electron capture dissociation of polypeptides in a three-dimensional quadrupole ion trap: Implementation and first results. AB - The adverse influence of the radio frequency (RF) voltage on electrons has been the main obstacle for the implementation of electron capture dissociation (ECD) in three-dimensional quadrupole ion traps (3D QITs). Here we demonstrate that the use of axial magnetic field, together with the injection of low-energy (<5 eV) electrons, in the beginning of the positive RF semi-period achieves trapping of electrons for a period of time comparable with the semi-period duration. Importantly, the energy of the electrons remains low during most of the trapping period. With this technique, which we call "magnetized electrons, in-phase injection" (MEPhI), ECD and other ion-electron reactions have become possible in a 3D QIT. Initial ECD results, including single-scan data, were obtained with dications of Substance P. The observed secondary fragmentation of ECD fragments indicates that the trapped electrons are still somewhat hotter than desired. PMID- 15653361 TI - Charge state-dependent fragmentation of oligonucleotide/metal complexes. AB - Collision-activated dissociation (CAD) has been employed to assess the gas-phase fragmentation behavior of a series of 1:1 oligodeoxynucleotide (ODN):metal complexes over a range of charge states, using several ten-residue ODNs and a wide array of alkali, alkaline earth, and transition metals. For parent species in low to intermediate charge states, complexation with Ca(+2), Sr(+2), or Ba(+2) altered the relative intensity of M-B species, promoting loss of cytosine over loss of guanine. The relative intensities of sequence ions were largely unaffected. This behavior was most prevalent for isomeric sequences with complementary residues at the 5'- and 3'-termini, suggesting that metal complexation may change the gas-phase conformation and/or conformational dynamics for some sequences. In higher charge states, some ODN/Ba(+2) complexes produced abundant fragment ions corresponding to metallated a(n)(-m) species, which are not commonly observed in CAD mass spectra for deprotonated ODNs. The formation of these ions was most favored for complexes between Ba(+2) and ODN sequences with a thymine residue at Position 6. Literature precedent exists for the formation of a(n)(-m) ions from sequences in which covalent modification generates one or more neutral sites along the phosphate backbone. ODN/metal adducts in high charge states possess only a few acidic protons, and the juxtaposition of these neutral phosphate groups near thymine residues and the bound Ba(+2) ion may direct formation of the metallated a(n)(-m) species. PMID- 15653362 TI - LC-mass spectrometry analysis of N- and C-terminal boundary sequences of polypeptide fragments by limited proteolysis. AB - Limited proteolysis is an important and widely used method for analyzing the tertiary structure and determining the domain boundaries of proteins. Here we describe a novel method for determining the N- and C-terminal boundary amino acid sequences of products derived from limited proteolysis using semi-specific and/or non-specific enzymes, with mass spectrometry as the only analytical tool. The core of this method is founded on the recognition that cleavage of proteins with non-specific proteases is not random, but patterned. Based on this recognition, we have the ability to determine the sequence of each proteolytic fragment by extracting a common association between data sets containing multiple potential sequences derived from two or more different mass spectral molecular weight measurements. Proteolytic product sequences derived from specific and non specific enzymes can be accurately determined without resorting to the conventional time-consuming and laborious methods of SDS-PAGE and N-terminal sequencing analysis. Because of the sensitivity of mass spectrometry, multiple transient proteolysis intermediates can also be identified and analyzed by this method, which allows the ability to monitor the progression of proteolysis and thereby gain insight into protein structures. PMID- 15653363 TI - Total structure characterization of unsaturated acidic phospholipids provided by vicinal di-hydroxylation of fatty acid double bonds and negative electrospray ionization mass spectrometry. AB - In the present work, the unsaturated fatty acid substituents of some phosphatidic acid, phosphatidylserine, phosphatidylinositol, and phosphatidylglycerol species were converted to their 1,2-di-hydroxy derivatives by OsO(4). The subsequent electrospray ionization tandem low-energy mass spectrometry analysis of the deprotonated species allowed positional determination of the double bonds by the production of specific product-ions. The product-ions are formed by charge-remote and charge-proximate homolytic cleavages as well as charge-directed heterolytic cleavages and rearrangements. The commercial availability of pure species of the phospholipids in question was limited, and a number of species were therefore synthesized. The developed method was used to fully characterize the two isobaric phosphatidylglycerol species 16:0/16:1Delta(9) and 16:0/16:1Delta(10) extracted from the bacteria Methylococcus capsulatus. The presence of these fatty acids was supported by a gas-chromatography mass spectrometry investigation of the dimethyloxazoline derivatives of the species of the lipid extract. The present work demonstrates that a total structure characterization of acidic unsaturated phospholipids in isolate or in mixtures is accomplished by vicinal di hydroxylation of olefinic sites and subsequent electrospray ionization mass spectrometry of the derivatized phospholipids. PMID- 15653364 TI - Oligosaccharide analysis using anion attachment in negative mode electrospray mass spectrometry. AB - Eleven different anionic species were able to form adducts with neutral oligosaccharides at low cone voltage in negative ion mode electrospray mass spectrometry. Among them, fluoride and acetate have the ability to significantly enhance the absolute abundance of [M - H](-) for neutral oliogosaccharides, which otherwise have low tendencies to deprotonate due to the lack of a highly acidic group. Evidence shows that the source of high abundances of [M - H](-) for neutral oligosaccharides arises from the decomposition of [M + F](-) and [M + Ac](-) with neutral losses of HF and HAc, respectively. The chloride adducts have the best stability among all the adduct species investigated, and chloride adducts consistently appeared in higher abundances relative to [M - H](-). In tandem mass spectrometry (ES-MS/MS) experiments, upon collision induced dissociation (CID), F(-) and Ac(-) adducts gave purely analyte-related product ions, i.e., no detection of the attaching anion and no incorporation of these anions into decomposition products. Cl(-) adducts produced both Cl(-) and analyte related product ions. For the above three anions, CID of adduct species may be used for structural determination of neutral oligosaccharides because, in each case, structurally-informative fragment ions were produced. In the presence of F( ) and Ac(-), simultaneous detection of acidic and neutral oligosaccharides was achieved, because the problem of the presence of an acidic group that can impede the deprotonation of a neutral oligosaccharide was minimized. The ratio of Cl( ):non-Cl-containing product ions obtained in CID spectra of chloride adducts of disaccharides was used to differentiate anomeric configurations of disaccharides. Density functional theory (DFT) was employed to evaluate the optimized structures of chloride adducts of disaccharides, and it was found that chloride anions favor close contact with the hydrogen from the anomeric hydroxyl group. Multiple hydrogen bonding further stabilizes the chloride adduct. PMID- 15653365 TI - Mutual storage mode ion/ion reactions in a hybrid linear ion trap. AB - Ion/ion proton transfer reactions involving mutual storage of both ion polarities in a linear ion trap (LIT) that comprises part of a hybrid triple quadrupole/linear ion trap mass spectrometer have been effected. Mutual ion storage in the x- and y-dimensions arises from the normal operation of the oscillating quadrupole field of the quadrupole array, while storage in the z dimension is enabled by applying unbalanced radio-frequency amplitudes to opposing sets of rods of the array. Efficient trapping (>90%) is achieved for thermalized ions over periods of several seconds. Reactions were demonstrated for multiply charged protein/peptide cations formed by electrospray with anions derived from glow discharge ionization of perfluoro(methyldecalin) (PMD) introduced from the side of the LIT rod array. Doubly and singly charged protein ions are readily formed via ion/ion reactions. The parameters that affect ion/ion reactions are discussed, including the degree of RF unbalance on the LIT rods, vacuum pressure, nature of the buffer gas, reaction time, anion abundance, and the low mass cutoff for ion/ion reaction. The present system has a demonstrated upper mass-to-charge ratio limit of at least 33,000. The system also has high flexibility with respect to defining MS(n) experiments involving both collision induced dissociation (CID) and ion/ion reactions. Experiments are demonstrated involving beam-type CID in the pressurized collision quadrupole (Q2) followed by ion/ion reactions involving the product ions in the LIT. Ion parking experiments are also demonstrated using the mutual storage ion/ion reaction mode in the LIT, with a parking efficiency over 60%. PMID- 15653366 TI - Volumetric intensity dependence on the formation of molecular and atomic ions within a high intensity laser focus. AB - The mechanism of atomic and molecular ionization in intense, ultra-short laser fields is a subject which continues to receive considerable attention. An inherent difficulty with techniques involving the tight focus of a laser beam is the continuous distribution of intensities contained within the focus, which can vary over several orders of magnitude. The present study adopts time of flight mass spectrometry coupled with a high intensity (8 x 10(15) Wcm(-2)), ultra-short (20 fs) pulse laser in order to investigate the ionization and dissociation of the aromatic molecule benzene-d1 (C(6)H(5)D) as a function of intensity within a focused laser beam, by scanning the laser focus in the direction of propagation, while detecting ions produced only in a "thin" slice (400 and 800 microm) of the focus. The resultant TOF mass spectra varies significantly, highlighting the dependence on the range of specific intensities accessed and their volumetric weightings on the ionization/dissociation pathways accessed. PMID- 15653367 TI - Extending the solvent-free MALDI sample preparation method. AB - Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry is an important technique to characterize many different materials, including synthetic polymers. MALDI mass spectral data can be used to determine the polymer average molecular weights, repeat units, and end groups. One of the key issues in traditional MALDI sample preparation is making good solutions of the analyte and the matrix. Solvent-free sample preparation methods have been developed to address these issues. Previous results of solvent-free or dry prepared samples show some advantages over traditional wet sample preparation methods. Although the results of the published solvent-free sample preparation methods produced excellent mass spectra, we found the method to be very time-consuming, with significant tool cleaning, which presents a significant possibility of cross contamination. To address these issues, we developed an extension of the solvent free method that replaces the mortar and pestle grinding with ball milling the sample in a glass vial with two small steel balls. This new method generates mass spectra with equal quality of the previous methods, but has significant advantages in productivity, eliminates cross contamination, and is applicable to liquid and soft or waxy analytes. PMID- 15653368 TI - Mass spectral behavior of some homoleptic and mixed aryldichalcogenide bis(diphenylphosphino)ferrocenenickel(II), palladium(II), and platinum(II), and bis(diisopropylphosphino)ferrocenepalladium(II) complexes. AB - The mass spectral behavior of a number of organometallic complexes containing the Group 10 metals Ni, Pd, and Pt, together with various thiolate ligands were studied. For Pd, two main types of complexes, differing by the substituents on the phosphorus atom were studied. Types I and II were substituted with bis(diphenylphosphino)ferrocene and bis(diisopropylphosphino)ferrocene ligands, respectively. The Ni complexes, except for one, and the Pd Type I complexes had no molecular radical cations (M(+.)) in their EI spectra. On the other hand, all the Pt complexes showed intense M(+.) ions in their EI spectra indicating that these complexes were more stable as radical cations than those of Ni and Pd. The FAB and MALDI spectra of all the complexes displayed intense quasi-molecular ions (MH(+)) and the fragmentations in both modes were similar. The MALDI spectra of several complexes displayed only M(+.) ions while one gave evidence of both MH(+) and M(+.) ions. Several Pd Type II complexes yielded intense M(+.) in their EI spectra. PMID- 15653369 TI - Quantitation of synthetic polymers using an internal standard by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. AB - MALDI-TOF MS is utilized to perform quantitative analysis on synthetic polymers. Despite the inherent limitations of MALDI, good quantitative results have been obtained in the three sets of experiments described here. An internal standard with similar molecular properties as the analytes is introduced. Plots of relative integrated intensity ratios as a function of theoretical ratios of stoichiometry are drawn based on the results. The satisfactory slopes and correlation coefficients illustrated the practicality of quantitative measurement by MALDI-TOF MS. PMID- 15653370 TI - In-Situ probing of the biotic-abiotic boundary of plants by laser desorption/ionization time-of-flight mass spectrometry. AB - Laser desorption/ionization time-of-flight (LDI-TOF) mass spectrometry was applied for the direct analysis of cuticular waxes on intact plant tissues. Cuticular wax compounds were ionized by laser desorption in the presence of colloidal silver. Silver-adduct ions were detected on samples from Arabidopsis thaliana and from maize. Good spot-to-spot reproducibility indicated homogeneous coverage of the sample by the fine colloidal material. The results were consistent with GC-MS analyses of cuticular extracts, thus confirming the feasibility of direct analysis based on this protocol. Molecular masses of the adduct ions correspond well with the known composition of cuticular waxes. Moreover, LDI-TOF gave good estimates of the relative local abundances of a given compound. However, bias was found in cases where compounds with different ionization efficiencies were analyzed. PMID- 15653371 TI - Evaluation of protein-DNA binding affinity by electrospray ionization mass spectrometry. AB - Binding affinity of complexes between a DNA-binding domain (DBD) of a transcription factor, c-Myb, and several double-stranded DNA (dsDNA) were evaluated by collision-induced dissociation (CID) of the multiply protonated molecules generated by electrospray ionization mass spectrometry (ESI-MS). Complexes of the c-Myb DBD and dsDNA were prepared in solution and analyzed by ESI-MS. Multiply protonated molecules of a high-affinity complex, the c-Myb DBD and dsDNA with a specific sequence, were clearly observed in ESI mass spectrum. Protonated molecules of the complex were quite stable in the gas-phase, and not easily dissociated even if high cone voltage was applied in the first vacuum chamber source when the sample was prepared in 10 mM ammonium acetate. As for the sample prepared in buffer with higher concentration of ammonium acetate, such as 500 mM ammonium acetate, protein-dsDNA complexes could easily be dissociated with an increase in the cone voltage, giving multiply protonated molecules of free c Myb DBD and some DNA fragments. Systematic CID experiments were carried out on seven complexes between the c-Myb DBD and 22-mer dsDNA with different solution-Kd values in the range of 10(-9) M to 10(-7) M. For each complex dissociation curve as a function of cone voltage was plotted, and the cone voltage where 50% of the complex was dissociated (V(50%)) was calculated. Consequently, positive correlation was obtained between V(50%) and relative binding free energy change (DeltaDeltaG) in complex formation in solution. This suggests that ESI-CID experiments can provide quantitative evaluation of the stability of protein-DNA complexes based on proper calibration. PMID- 15653372 TI - Elemental distribution in fluorinated amorphous carbon thin films. AB - Focused ion beam-secondary ion mass spectrometry (FIB-SIMS) with 20 nm spatial resolution has been used to analyze amorphous fluorinated carbon thin films, deposited by plasma assisted chemical vapor deposition (PACVD), at micro- to nano scale. Mass spectra and ion imaging of film surface were acquired and the presence and distribution of contaminants were investigated. Surface images show the secondary ion distribution for F(-), CH(-), CF(-). A change in size and topology of fluorine-rich areas is correlated with film hardness and with microstructure transition from diamond-like to polymer-like, as indicated by infrared and Raman spectroscopies. Based on the surface distributions of CF(-) and CH(-) and on the vibrational spectroscopy results, a mechanism of fluorine substitution for hydrogen and an attempt to explain the film structure and microstructure is proposed. PMID- 15653373 TI - Obliterative bronchiolitis or chronic lung allograft rejection: a basic science review. PMID- 15653374 TI - IL-1beta in bronchial lavage fluid is a non-invasive marker that predicts the viability of the pulmonary graft from the non-heart-beating donor. AB - BACKGROUND: Viability testing of the pulmonary graft retrieved from the non-heart beating donor (NHBD) is mandatory for successful outcome after lung transplantation. Functional assessment by ex vivo reperfusion, however, remains a cumbersome procedure. In this study, therefore, we wanted to investigate the possible value of the proinflammatory cytokines interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) measured in bronchial lavage fluid (BLF) in predicting functional outcome of the pulmonary graft after reperfusion. METHODS: Domestic pigs (29.9 +/- 0.56 kg) were sacrificed and divided in 5 groups (n = 5/group). In the non-ischemic group (NHBD-0), the heart-lung block was explanted immediately. In the other groups the animals were left untouched with increasing time intervals (1 hour = NHBD-1; 2 hours = NHBD-2; 3 hours = NHBD-3). Thereafter both lungs were cooled topically via chest drains up to a total ischemic interval of 4 hours. Finally, in the heart-beating donor group lungs were flushed and stored for 4 hours (4 degrees C) [HBD]. BLF samples were taken from the right lung in all groups after explantation for measurement of IL-1beta and TNF-alpha and the left lung was prepared for evaluation in an isolated reperfusion circuit. Haemodynamic, aerodynamic and oxygenation parameters were measured. Wet-to-dry weight ratio (W/D) was calculated after reperfusion. RESULTS: Graft function deteriorated with increasing time intervals after death. A strong correlation was found between the increase of IL-1beta concentration measured in BLF and the increase in pulmonary vascular resistance (r = 0.80), mean airway pressure (r = 0.74) and wet-to dry weight ratio (r = 0.78); (p < 0.0001, for all parameters). No significant differences in TNF-alpha levels in BLF were observed amongst groups (p = 0.933). CONCLUSIONS: IL-1beta in BLF prior to reperfusion correlated well with graft function and may therefore be a useful, non-invasive marker that can predict the viability of the pulmonary graft from the NHBD. PMID- 15653375 TI - Decreased recipient survival following orthotopic heart transplantation with use of hearts from donors with projectile brain injury. AB - BACKGROUND: Fatal gunshot injury to the brain can cause significant alterations in the neuroendocrine state and myocardial dysfunction. Therefore heart allografts from these donors may result in graft failure following orthotopic heart transplantation (OHTx). We evaluated whether receiving a heart from a donor who died from fatal gunshot wound to the brain independently affected the outcome of transplantation. METHODS: A retrospective review of 113 consecutive patients undergoing OHTx at a university hospital from 1996 to 2002 was performed. Group 1 received hearts from donors with fatal gun shot brain injury (n = 17), and Group 2 received hearts from donors who died from other causes (n = 96). RESULTS: Recipient age, gender, United Network for Organ Sharing (UNOS) status, indication for transplantation, and other co-morbid conditions were similar in both groups. Young male donors pre-dominated in Group 1, but other donor characteristics were not significantly different. The incidence of Grade 3A rejection was higher in Group 1 than Group 2 (35% vs 6.3%, p = 0.003), as was the incidence of post operative infection (35% vs 7.2%, p = 0.004). Actuarial survival at 1 and 5 years was significantly lower in Group 1 than in Group 2 (81% and 74% vs 97% and 94%, respectively, p = 0.005). Multivariate logistic regression analysis also demonstrated that fatal gunshot brain injury, as cause of donor death, was a risk factor for recipient mortality (p = 0.01). CONCLUSION: Receiving a heart from a donor with fatal gunshot brain injury is a significant risk factor for recipient mortality following OHTx. Cautious use of heart allograft from these donors, especially in low-risk recipients, may lead to improved outcome following heart transplantation. PMID- 15653376 TI - Acceptable recipient outcomes with the use of hearts from donors with hepatitis-B core antibodies. AB - BACKGROUND: The shortage of available donors limits cardiac transplantation. Use of hearts from patients with hepatitis-B core antibodies could expand the donor pool but are usually avoided because of concern about virus transmission. We conducted a retrospective review to determine the safety of transplanting hearts from donors with hepatitis-B core antibodies. METHODS: We reviewed donor and recipient charts for patients who underwent transplantation at our center between January 1, 1997, and December 1, 2002. RESULTS: A total of 541 heart transplantations were performed in this time period. Thirty-three patients (aged 47.5 +/- 18.8 years) received hearts from core-antibody-positive donors (aged 37.7 +/- 10.8 years). Of these, 5 patients received prophylactic antibiotic treatment with lamivudine after transplantation. Only 1 patient (baseline surface antigen-negative and without prophylaxis) experienced donor-transmitted hepatitis B infection 10 months after transplantation that was treated with lamivudine. Two patients (baseline surface-antibody-negative) had hepatitis B seroconversion, becoming surface-antibody positive without evidence of infection. None of the 5 patients who received prophylaxis with lamivudine had donor-transmitted hepatitis, and only 1 lamivudine-treated patient had surface antibodies. Post transplant survival in this small cohort was similar to that for all patients who underwent transplantation at our center during this time period. CONCLUSIONS: Transplantation of hearts from donors with hepatitis-B core antibodies is associated with a small viral-transmission risk, with or without post-transplant, anti-viral prophylaxis. Use of these donor hearts should be considered safe and may help to augment the available donor pool. PMID- 15653377 TI - Depletion of alveolar macrophages by clodronate-liposomes aggravates ischemia reperfusion injury of the lung. AB - BACKGROUND: Macrophages play an important role in ischemia-reperfusion injury of various organs. Liposome-encapsulated dichloromethylene diphosphonate (clodronate liposome) depletes local macrophages in vivo. However, the effect of this approach on alveolar macrophages in pulmonary ischemia-reperfusion injury has not yet been evaluated. METHODS: Clodronate-liposomes in Hanks' balanced salt solution (HBSS) or HBSS alone were given intratracheally to anesthetized male Lewis rats in the clodronate or the control group (n = 6/each group). After 3 days, we subjected the lungs to ischemia (37 degrees C, 60 minutes) and reperfusion (60 minutes) in an isolated blood-perfused rat lung model. Analysis during reperfusion included gas exchange, hemodynamics, and airway mechanics. At the end of reperfusion, we determined leukocyte recruitment and macrophage inflammatory protein-2 (MIP-2) in bronchoalveolar lavage fluid. RESULTS: In the clodronate group, 4 experiments had to be terminated within 10 minutes of reperfusion because of severe lung injury, whereas all lungs of the controls could be studied during the 60-minute reperfusion period (p < 0.05). Clodronate significantly decreased dynamic airway compliance (p < 0.05) and increased airway resistance. Besides a tendency toward greater pulmonary vascular resistance, this was associated with recruitment of polymorphonuclear neutrophils (p < 0.05) and increased MIP-2 concentrations in the bronchoalveolar lavage fluid (p < 0.05). CONCLUSIONS: Intratracheal administration of liposome-encapsulated clodronate does not benefit, but aggravates, warm ischemia-reperfusion injury of the lung, increasing MIP-2-associated alveolar neutrophil recruitment and airway mechanical dysfunction. PMID- 15653378 TI - Simvastatin decreases myocardial tumor necrosis factor alpha content in heart transplant recipients. AB - BACKGROUND: Statins improve patient survival and decrease rejection episodes in heart transplant recipients. We studied the effects of simvastatin treatment on myocardial tumor necrosis factor alpha (TNF-alpha) expression; TNF-alpha is a potent pro-inflammatory cytokine associated with hypertrophy and fibrosis in heart transplant recipients. METHODS: We randomized 10 consecutive heart transplant recipients to receive either 20 mg/day simvastatin (n = 5) or placebo (n = 5) for 6 months after cardiac transplantation. Routine surveillance endomyocardial biopsy specimens were obtained from all patients. We analyzed tissues for myocardial TNF-alpha content, total collagen content, and myocyte size using semiquantitative immunohistochemistry. RESULTS: Myocyte size and total collagen content of placebo and simvastatin groups did not show a statistically significant difference at any biopsy time point. Myocardium TNF-alpha content (% tissue area stained) at 1 week after transplantation was similar in the simvastatin and placebo groups. At the 24(th) week after transplantation, when compared with Week 1 values, we found a significant decrease in myocardium TNF alpha content in the simvastatin group (15.0% +/- 2.3% vs 5.8% +/- 2.4%, p = 0.02) that was not observed in the placebo group (15.0% +/- 1.5% vs 12.0% +/- 2.6%, p = not significant). CONCLUSION: Simvastatin treatment in heart transplant recipients decreased myocardium TNF-alpha expression. This decrease did not translate into a difference in the markers of hypertrophy. However, decreased myocardial TNF-alpha may be a marker of a general statin-mediated decrease in inflammation in the transplanted heart that leads to improved graft and patient survival. PMID- 15653379 TI - Brain natriuretic peptide predicts serious cardiac allograft rejection independent of hemodynamic measurements. AB - BACKGROUND: Serum brain natriuretic peptide (BNP) has been reported to be elevated in heart transplant recipients with both cellular and vascular rejection. Whether BNP can be used to help predict the severity of rejection is not well established. METHODS: We analyzed serial BNP measurements obtained during endomyocardial biopsy procedures in consecutive heart transplant patients occurring >45 days after transplantation. To eliminate potential confounding from prior rejection episodes, we included only observations in which the previous biopsy grade was 0 or 1A. Multivariable linear regression was performed examining the outcome of increasing seriousness of rejection, defined as grade 0 < 1A < 2 < 1B < 3A < vascular rejection. A univariable logistic regression model was performed using log-transformed BNP as a predictor of vascular rejection. RESULTS: There were 77 patients, with 161 separate observations. Median time between transplantation and first assessment was 6.0 months (interquartile range, 2.1, 31.6). Using multivariable linear regression, 3 factors were significantly associated with biopsy score: pulmonary capillary wedge pressure (p < 0.0001), BNP (p = 0.003), and heart rate (p = 0.01). Even after other significant univariable predictors (including pulmonary capillary wedge pressure) were forced into the model, BNP remained a significant predictor of biopsy score (p = 0.02). Log BNP was a significant univariable predictor of vascular rejection, with an odds ratio of 12.55 (per 1 unit increase, 95% confidence interval 3.43-45.84; p = 0.0001) and a model c-statistic of 0.91. CONCLUSIONS: BNP predicts new episodes of serious cardiac allograft rejection, particularly vascular rejection, independent of hemodynamic measurements, and may be a useful part of rejection surveillance. PMID- 15653380 TI - Successful outcome with extended allograft ischemic time in pediatric heart transplantation. AB - BACKGROUND: Many cardiac transplant programs have liberalized donor eligibility criteria in an attempt to maximize donor supply and to accommodate increasing demand. Although many studies have evaluated the potential adverse effects of prolonged donor ischemic time (DIT) in adults undergoing cardiac transplantation, relatively few have focused specifically on pediatric recipients that include a substantial number of patients and long-term follow-up. The focus of this study was to examine the effect of extended DIT on mortality after pediatric heart transplantation. METHODS: We conducted a retrospective review of our pediatric cardiac transplant experience in the past 11 years, comparing patients who received allografts and had ischemic times >240 minutes with those who had ischemic times <240 minutes. RESULTS: A total of 129 pediatric patients (<19 years) underwent orthotopic heart transplantation, of whom 78 (60.5%) had DIT <240 minutes and 51 (39.5%) had DIT >240 minutes. We found no statistically significant difference in age, sex, race, height, weight, or donor age between the groups (p = not significant). Post-transplant survival at 1, 5, and 10 years was similar for both groups: 91.2%, 88.0%, and 85.2%, respectively, for patients with DIT <240 minutes vs 89.6%, 87.2%, and 79.8%, respectively, for patients with DIT >240 minutes (p = 0.433). Additionally, using Cox proportional hazard models, extended DIT >240 minutes was not a statistically significant independent predictor of post-transplant mortality (odds ratio, 0.655; 95% confidence interval, 0.518-0.972; p = 0.684; standard error = 0.468). CONCLUSION: Procurement of hearts from distant locations with associated extended DIT is justified in the setting of increased demand and a fixed donor population. PMID- 15653381 TI - Pravastatin therapy is associated with reduction in coronary allograft vasculopathy in pediatric heart transplantation. AB - BACKGROUND: Hydroxymethylglutaryl CoA reductase inhibitors (statins) have been demonstrated to reduce the risk of developing coronary allograft vasculopathy (CAV) following heart transplantation in adults and are used routinely in many centers. CAV and lipid abnormalities have been reported to be less prevalent in pediatric heart transplant recipients. It is not known whether statins reduce the risk of CAV in this population METHODS: A retrospective review was performed to analyze the risk factors for developing CAV following pediatric heart transplantation with particular attention to the impact of pravastatin therapy. The study population was comprised of 129 pediatric patients who underwent 142 heart transplants at our institution from 1988 to 2003. The outcome variable was freedom from CAV, CAV being determined by coronary angiography or autopsy. RESULTS: CAV was identified in 25 recipients at a median of 3.7 years after transplantation. There were 331 patient-years of pravastatin therapy. Pravastatin therapy resulted in a reduction in total cholesterol levels, 162 +/- 29 to 137 +/ 20 mg/dl, p = 0.01. In multivariate analysis the use of pravastatin was associated with a lower incidence of CAV (p = 0.03), whereas an increased frequency of late rejection (p = 0.003) and earlier year of transplantation (p = 0.04) were associated with increased risk of CAV. CONCLUSIONS: The routine use of pravastatin was associated with a lower risk following pediatric heart transplantation. Further studies into the relationship between lipid abnormalities, inflammation and rejection, and the development of CAV in children are warranted. PMID- 15653382 TI - Endogenous bone-marrow-derived stem cells contribute only a small proportion of regenerated myocardium in the acute infarction model. AB - BACKGROUND: Our recent study showed that granulocyte-colony stimulating factor (G CSF) promoted bone-marrow cells (BMC) to migrate into the infarcted heart and that they differentiated into cardiomyocytes. However, we still do not know to what degree bone-marrow-derived cardiomyocytes contribute to myocardial regeneration after injury. In this study, we verified the proportional contribution of cells from bone marrow (BM) and from non-bone marrow (n-BM) in regenerating neomyocardium after myocardial infarction. METHODS: Eight C57BL/6 mice were irradiated (900 cGy), and green fluorescent protein (GFP) mouse-derived BMCs (GFP-BMC, 1 x 10(6) cells) were injected. Four weeks later, the left descending coronary artery was ligated. Recombinant human G-CSF (200 microg/kg/day, 8 days) was injected. At 4 weeks after ligation, hearts were fixed for histology. We calculated the proportions of cardiomyocytes derived from BM and n-BM after taking the chimeric rate into consideration. RESULTS: The chimeric rate was 54.6% +/- 5.9%. At the infarcted border area, the total cell number was 1000.3 +/- 56.5/mm(2), and mobilized BM-derived GFP-BMC was 103.3 +/- 13.1/mm(2). After compensation with the chimeric rate, we found BM-derived troponin I positive cells at 23.9 +/- 4.1/mm(2), nestin-positive cells at 12.9 +/- 2.6/mm(2), and Ki67-positive cells at 18.3 +/- 2.6/mm(2), respectively. We found significant differences in the contribution of troponin I-(6.7% +/- 1.7% vs 93.3% +/- 1.7%), nestin- (2.4 +/- 0.5 vs 97.6 +/- 0.5), and Ki67-positive (3.9 +/- 1.0 vs 96.1 +/- 1.0) cells derived from BM and n-BM. CONCLUSIONS: Bone marrow was one of the origins of regenerated cardiomyocytes; however, the contribution of cells from BM was very small compared with those of n-BM origin in the infarction model. PMID- 15653383 TI - Sarcomeric genes involved in reverse remodeling of the heart during left ventricular assist device support. AB - BACKGROUND: Left ventricular assist devices (LVADs) implanted in patients with severe congestive heart failure (CHF) as a bridge to transplantation have been shown to reverse chamber enlargement, regress cellular hypertrophy, and increase contractility. The purpose of this study was to gain a better understanding of the molecular changes associated with increased contractility after LVAD support. METHODS: We took tissue sections from the left ventricular apex of 12 patients with CHF who were undergoing LVAD insertion (pre-LVAD) and from the LV free wall of those same patients before transplantation (post-LVAD). To control for sample site differences, we obtained samples from the same regions in 7 patients with CHF who were undergoing transplantation without LVAD support and in 4 non- failing donor hearts. Gene expression was then probed on a custom DNA array containing 2,700 cardiac-enriched cDNA clones. RESULTS: Calcium-handling genes were up-regulated by LVAD support, as previously reported. Sarcomeric genes were the other principle class of genes up-regulated by LVAD support, consistent with a possible restoration of sarcomere structure in reverse ventricular remodeling. However, a decrease in the fibrous component of the myocardium, also potentially involved in reverse remodeling, was not evident at the level of gene transcription because fibroblast markers were either unchanged or up-regulated. The remaining regulated genes did not fall into any defined functional class. CONCLUSIONS: Changes in the regulation of sarcomeric, calcium-handling, and fibroblast genes during LVAD support indicate a cardiac molecular adaptation to mechanical unloading. These molecular changes may play a role in the observed increase in contractile function during reverse remodeling. PMID- 15653384 TI - Experience and result of extracorporeal membrane oxygenation in treating fulminant myocarditis with shock: what mechanical support should be considered first? AB - BACKGROUND: Extracorporeal membrane oxygenation (ECMO), instead of ventricular assist device (VAD), could work as the first-line treatment of choice for fulminant myocarditis (FM) with profound shock if intraaortic balloon pumping was inadequate. We reviewed our experience in treating FM with ECMO and compared it with the literature that described the use of VAD. METHODS: Fifteen consecutive patients (age 27.1 +/- 19.3 years) who had FM with profound shock were rescued with ECMO emergently. Hypotension, depressed left ventricular ejection fraction (19.1% +/- 6.1%), and oliguria occurred in all patients with high-dose catecholamine (inotropic equivalents: 69.0 +/- 37.7 microg/kg/min) and ventilator support. Before ECMO support, 6 patients received intraaortic balloon pumping support, 5 received external cardiac massage, 5 needed a temporary pacemaker, and 4 needed continuous hemofiltration. The pre-ECMO cardiac enzyme and liver enzyme levels were abnormally high. RESULTS: Fourteen patients (93.3%) could be weaned off mechanical support. Three of 14 successfully weaned patients died later as a result of complications. Survival to discharge was 73.3%, and none of survivors needed heart transplantation. The ECMO duration was 137.7 +/- 74.5 hours. The ECMO-related neurological complication (6.7%) and the reexploration rate for hemostasis (8.9%) were lower than the myocarditis group supported by VAD from the literature review. The 11 survivors exhibited no cardiac dysfunction during the follow-up period. CONCLUSIONS: Owning to advantages of fewer complications, easier application, and biventricular support, ECMO can be considered as the first-line treatment of mechanical support for FM with profound shock when intraaortic balloon pumping is inadequate or infeasible. PMID- 15653385 TI - Changes in induced sputum in the presence of bronchiolitis obliterans syndrome and correlation with spirometry in single and bilateral lung transplant recipients. AB - Bronchiolitis obliterans syndrome is a clinical diagnosis based on lung function parameters. Using induced sputum, taken from lung transplant recipients, this paper reports on the correlation between the neutrophil count and the percentage change from postoperative baseline for FEV(1), FEF(50), and FEF(25-75). In double lung transplant recipients the correlations were significant for FEV(1) (r = 0.68, p = 0.002), FEF(50) (r = -0.65, p = 0.016), and FEF(25-75) (r = -0.56, p = 0.016). In single lung transplant recipients, no significant correlations were seen. PMID- 15653386 TI - Troponin T levels in baboons with pig heterotopic heart transplants. AB - Troponin T levels have been monitored in baboons (n = 8) undergoing pig heterotopic heart transplantation, and correlated with a decrease in graft contractions and graft survival. Pig heart graft survival was from 12 to 139 days (mean 45, median 33), and graft failure was associated with predominant thrombotic microangiopathy and ischemia, with focal hemorrhage, and edema. An increase in troponin T levels 5 to 6 days before graft failure correlated closely with diminished graft contractions. An increase in troponin T was a reliable indicator that graft dysfunction was occurring. PMID- 15653387 TI - Noninfectious constrictive pericarditis in a heart transplant recipient. AB - Acute rejection, infection, and allograft coronary artery disease have been recognized as the major causes of postoperative morbidity and mortality in cardiac transplant patients. More recently, pericardial and mediastinal complications have been recognized as a more common complication than previously believed. We describe a case of a heart transplant recipient admitted for apparent congestive heart failure exacerbation who was unresponsive to standard medical management of congestive heart failure and rejection. After further invasive evaluation, it was discovered the patient's condition was attributable to posttransplantation constrictive pericarditis. It is appropriate to consider this diagnosis in any postcardiac surgery (especially heart transplant recipients) in patients presenting with congestive heart failure exacerbations refractory to usual medical management. PMID- 15653388 TI - Successful repair of tracheal dehiscence after heart-lung transplantation. AB - Tracheal dehiscence remains one of the most feared complications after heart-lung transplantation because of its nearly universal fatality rate. Drawing on experiences from the tracheal reconstruction and the lung transplantation literature, we report the successful repair of tracheal dehiscence, after heart lung transplantation, with an intercostal muscle flap. PMID- 15653389 TI - Superiority of voriconazole over amphotericin B in the treatment of invasive aspergillosis after heart transplantation. AB - Invasive aspergillosis in immuncompromised patients occurs frequently. Surgical therapy in combination with chemotherapy with amphotericin B and itraconazole is standard therapy. We describe a heart transplant recipient with invasive Aspergillus fumigatus infection in the lung, which was treated successfully with voriconazole after systemically high-dose intravenous therapy with amphotericin B failed. PMID- 15653390 TI - Gastrointestinal bleeding from arteriovenous malformations in patients supported by the Jarvik 2000 axial-flow left ventricular assist device. AB - The long-term effects of axial-flow mechanical circulatory support in humans are unclear. We report 3 cases of chronic gastrointestinal bleeding after implantation of a Jarvik 2000 axial-flow left ventricular assist device. The bleeding was refractory to aggressive management and in 2 cases resolved only after orthotopic cardiac transplantation. PMID- 15653391 TI - The role of growth hormone replacement in a growth hormone deficient patient with underlying cardiomyopathy and severe congestive heart failure. AB - It has been reported that growth hormone (GH) deficiency induced cardiomyopathy responds to growth hormone replacement therapy. We describe the case of a middle aged male with cardiomyopathic heart failure and growth hormone deficiency of the adult secondary to surgical panhypopituitarism. We demonstrate clinical and hemodynamic improvement of cardiac function with growth hormone replacement therapy despite underlying structural heart disease. PMID- 15653393 TI - When Earth started blooming: insights from the fossil record. AB - Recent palaeobotanical studies have greatly increased the quantity and quality of information available about the structure and relationships of Cretaceous angiosperms. Discoveries of extremely well preserved Cretaceous flowers have been especially informative and, combined with results from phylogenetic analyses of extant angiosperms (based mainly on molecular sequence data), have greatly clarified important aspects of early angiosperm diversification. Nevertheless, many questions still persist. The phylogenetic origin of the group itself remains as enigmatic as ever and, in some cases, newly introduced techniques from molecular biology have given confusing results. In particular, relationships between the five groups of extant seed plants remain uncertain, and it has sometimes proved difficult to reconcile estimates of the time of divergence between extant lineages made using a 'molecular clock' with the fossil record. One result, however, is becoming increasingly clear: a great deal of angiosperm diversity is extinct. Some groups of angiosperms were evidently more diverse in the past than they are today. In other cases, fossils defy assignment to extant groups at the family level or below. This raises the possibility that evolutionary conclusions based solely upon extant taxa that are merely relics of groups that were once much more diverse might be misled by the effects of extinction. It also introduces the possibility that some early enigmatic fossils might represent lineages that diverged from the main line of angiosperm evolution below the most recent common ancestor of all extant taxa. These, and other questions, are among those that need to be addressed by future palaeobotanical research. PMID- 15653394 TI - Evolutionary dynamics of genes controlling floral development. AB - Advances in the understanding of floral developmental genetics in model species such as Arabidopsis continue to provide an important foundation for comparative studies in other flowering plants. In particular, floral organ identity genes are the focus of many projects that are addressing both ancient and recent evolutionary questions. Expanded analyses of the evolution of these gene lineages have highlighted the dynamic nature of the gene birth-and-death process, and may have significant implications for the evolution of genetic pathways. Crucial functional studies of floral organ identity genes in diverse taxa are allowing the first real insight into the conservation of gene function, while findings on the genetic control of organ elaboration offer to open up new avenues for investigation. Taken together, these trends show that the field of floral developmental evolution continues to make significant progress towards elucidating the processes that have shaped the evolution of flower development and morphology. PMID- 15653395 TI - Genomic imprinting in plants: the epigenetic version of an Oedipus complex. AB - Genomic imprinting is a mitotically stable epigenetic modification that results in the functional non-equivalency of both parental genomes following fertilization. In flowering plants, studies of parent-of-origin effects have mostly identified genes that are only transcribed from a maternally inherited allele. In Arabidopsis, the Polycomb group protein MEDEA regulates seed development through the expression of the MADS-box gene PHERES1. Activation of the maternal MEDEA allele requires the function of DEMETER, a plant DNA glycosylase that also controls the transcriptional activity of the maternally inherited allele of the late-flowering gene FWA. Current studies of parent-of origin effects have mostly identified genes that are only transcribed from a maternally inherited allele. Our current understanding of parent-of-origin effects could represent a new form of an Oedipus complex in which flowering plants prefer to rely transcriptionally on their maternal rather than their paternal chromosomes to ensure normal initiation of seed development. PMID- 15653396 TI - Intercellular signalling in the transition from stem cells to organogenesis in meristems. AB - Meristems continuously produce new cells to sustain plant growth. Stem cells are maintained in the centre of the meristem and provide the precursor cells for the initiation of new organs and tissues in the periphery. The structure of the meristem is maintained while cells are constantly displaced by new divisions. Recent advances have been made in understanding the intercellular signals that maintain meristem structure by adjusting gene expression according to cell position. In addition to refinements in our understanding of how the position and size of the stem-cell population is regulated, there have been advances in understanding how the location of new organ primordia is controlled and how the meristem influences organ polarity. PMID- 15653397 TI - Auxin and embryo axis formation: the ends in sight? AB - The major axis of polarity of the plant embryo serves as a reference for the formation of meristems and, thus, for all subsequent development. Mechanisms underlying the establishment of the embryo axis itself have remained elusive. This is now changing with recent reports documenting a role for auxin in embryo axis formation. Auxin accumulates dynamically at specific positions that correlate with developmental decisions in early embryogenesis, and this ties developmental decisions to both transport regulators and components of the response machinery. A major challenge for the future is to determine how auxin dependent processes interact with other as yet unknown factors to mediate differential gene expression patterns in early embryogenesis. PMID- 15653398 TI - The developmental role of microRNA in plants. AB - MicroRNAs (miRNAs) are single-stranded RNA molecules of around 22 nucleotides (nt) in length that are associated with the RNA-induced silencing complex (RISC). They play an important role in plant development, either by targeting mRNA for cleavage or by inhibiting translation. Over the past year, the list of known miRNAs, confirmed targets and developmental effects has expanded, as has the realization that they are conserved during evolution and that small RNAs can play a direct role in cell-cell signaling. PMID- 15653399 TI - Regulation of short-distance transport of RNA and protein. AB - The intercellular trafficking of proteins and RNAs has emerged as a novel mechanism of cell-cell communication in plant development. Plasmodesmata (PD), intercellular cytoplasmic channels, have a central role in cell-cell trafficking of regulatory proteins and RNAs. Recent studies have demonstrated that plants use either a selective or a non-selective PD trafficking pathway for regulatory proteins. Moreover, plants have developed strategies to regulate both selective and non-selective movement. Recent work has focused especially on integrating the recent understanding of the function and mechanisms of intercellular macromolecule movement through PD. PMID- 15653400 TI - Formation of primordia and phyllotaxy. AB - Leaves are made in an iterative pattern by the shoot apical meristem. The mechanism of this pattern formation has fascinated biologists, mathematicians and poets for centuries. Over the past year, fundamental insights into the molecular basis of this process have been gained. Patterns of auxin polar transport dictate when and where new leaf primordia are formed on the surface of the apical meristem. Subsequent events are still obscure but appear to involve both alteration of cell wall characteristics (to facilitate a new vector of growth) and a cascade of spatially co-ordinated transcription factor activity (to determine the fate of cells that are incorporated into new lateral organs). The co-ordinated signalling events involved in these processes are beginning to be elucidated. PMID- 15653401 TI - Networks in leaf development. AB - Shoots are characterized by indeterminate growth resulting from divisions of undifferentiated cells in the central region of the shoot apical meristem. These cells give rise to peripheral derivatives from which lateral organ initials are recruited. During initial stages of cell recruitment, the three-dimensional form of lateral organs is specified. Lateral organs such as leaves develop and differentiate along proximodistal (base-to-tip), dorsoventral (top-to bottom) and mediolateral (middle-to-margin) planes. Current findings are refining our knowledge of the genes and genetic interactions that regulate these early processes and are providing a picture of how these pathways may contribute to variation in leaf form. PMID- 15653402 TI - Regulation of developmental transitions. AB - Plants undergo a series of profound developmental changes throughout their lifetimes in response to both external environmental factors and internal intrinsic ones. When these changes are abrupt and dramatic, the process is referred to as phase change. Recently, several genes have been discovered that play a role in these developmental transitions. Their sequence and expression patterns shed new light on the mechanisms of phase change, and provide a link between the external and internal factors that control them. Examples of these transitions include changes from juvenile to adult leaf formation, vegetative to inflorescence meristem development, and inflorescence to floral meristem initiation. PMID- 15653403 TI - Stepwise understanding of root development. AB - Recent studies using Arabidopsis propose a framework of root development and pattern formation that can be divided to three processes. First, a positional signal that is delivered from neighboring cells controls the fate of undifferentiated cells. Then, cell fate is fixed through a protein-network that includes various transcription factors. Finally, the expression of a particular gene-set leads to fate-dependent cell differentiation, resulting in oriented cell division, cell specification and cell elongation. In addition, these processes could be modified by chromatin stabilization and protein degradation. We focus on three fundamental patterns of root development, circumferential pattern, radial pattern and proximo-distal pattern, and on novel approaches to identify genes that are responsible for the spatiotemporal regulation of root development. PMID- 15653404 TI - A DELLAcate balance: the role of gibberellin in plant morphogenesis. AB - The importance of gibberellin (GA) in vegetative and reproductive development has been known for some time. Recent studies have uncovered new roles of GA in leaf differentiation, photomorphogenesis and pollen-tube growth. Significant contributions to our understanding of GA-regulated morphogenesis include the identification of upstream regulators of GA biosynthesis, the elucidation of the function of GA signaling components, and the isolation of downstream targets. In addition, the mechanisms of interactions between GA and other hormone pathways are beginning to be revealed at the molecular level. PMID- 15653405 TI - The HSP90 chaperone complex, an emerging force in plant development and phenotypic plasticity. AB - The essential cellular functions of the molecular chaperone HSP90 have been intensively investigated in fungal and mammalian model systems. Several recent publications have highlighted the importance of this chaperone complex in plant development and responsiveness to external stimuli. In particular, HSP90 is crucial for R-protein-mediated defense against pathogens. Other facets of HSP90 function in plants include its involvement in phenotypic plasticity, developmental stability, and buffering of genetic variation. Plants have emerged as powerful tools that complement other model systems in attempts to extend our knowledge of the myriad impacts of protein folding and chaperone function. PMID- 15653406 TI - Control of plant development and gene expression by sugar signaling. AB - Coordination of development with the availability of nutrients, such as soluble sugars, may help ensure an adequate supply of building materials and energy with which to carry out specific developmental programs. For example, in-vivo and in vitro experiments suggest that increasing sugar levels delay seed germination and stimulate the induction of flowering and senescence in at least some plant species. Higher sugar concentrations can also increase the number of tubers formed by potatoes and can stimulate the formation of adventitious roots by Arabidopsis. New insights into the mechanisms by which sugar-response pathways interact with other response pathways have been provided by microarray experiments examining sugar-regulated gene expression under different light and nitrogen conditions. PMID- 15653407 TI - Breaking the WAVE complex: the point of Arabidopsis trichomes. AB - Actin filaments comprise an essential cytoskeletal array that organizes the cytoplasm during growth and cell division. In growing cells, actin filaments carry out many functions. Actin filaments position the endomembrane system and act as a substrate on which organelle motility occurs. Other actin-filament arrays appear to be more dynamic and to reorganize in response to growth signals and external cues. The diverse cellular functions of the actin cytoskeleton are mediated by actin-binding proteins that nucleate, destabilize, and bundle actin filaments. The distorted trichome morphology mutants provide a simple genetic system in which to study mechanisms of actin-dependent morphogenesis. Recent results from several groups indicate that 'distorted group' genes encode subunits of the actin-related protein (Arp)2/3 and WAVE complexes, and function in a cell morphogenesis pathway. PMID- 15653408 TI - Practical applications of research into the regulation of plant volatile emission. AB - Throughout their life cycles, plants release diverse blends of volatile compounds that play crucial roles in pollinator attraction, defense and communication. The importance of plant volatiles, in addition to the general appeal of fragrances and flavors to humans, have made these secondary metabolites a target for metabolic engineering. In the past decade, significant discoveries in the plant volatile biosynthetic pathways have provided a starting point for their modification. Pioneering attempts to alter plant volatile profiles have uncovered the complexity of networks and their regulation, and have built new avenues for future successful metabolic engineering. PMID- 15653409 TI - T cell response in hepatitis C virus infection. AB - Hepatitis C virus (HCV) is a hepatotropic RNA virus that causes acute and chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma. It is widely accepted that cellular immune responses play an important role in viral clearance and disease pathogenesis. However, HCV often evades effective immune recognition and has a propensity to persist in the majority of acutely infected individuals (ca. 80%). The immunological and virological basis for the inefficiency of the cellular immune response to clear or control the virus is not known. Recent studies, however, have provided new insights into the mechanisms of viral clearance and persistence that will be discussed in detail. PMID- 15653410 TI - Hepatitis C virus-related resistance mechanisms to interferon alpha-based antiviral therapy. AB - Only 50-60% of the patients chronically infected with the hepatitis C virus (HCV) achieve a sustained virologic response to the current standard antiviral therapy consisting of pegylated interferon alpha in combination with ribavirin. The definite reasons for virologic response or non-response to interferon alpha-based therapy are unknown. Besides host and treatment efficacy factors, it is presumable that HCV is able to antagonize the antiviral activity of interferon alpha. So far, among the different HCV proteins, the envelope (E)2 protein, the non-structural (NS)3/4A protein, and the NS5A protein have been associated with interferon alpha resistance mechanisms in vitro. The clinical significance of amino acid mutations within these HCV proteins in HCV isolates from patients who did or did not respond to interferon alpha-based therapy was investigated in multiple studies. Within the E2 (HVR2, CD81 binding sites, PePHD) and the NS3/4A proteins no specific mutations in correlation with virologic response to interferon alpha-based therapy were observed. For the NS5A protein, mutations within the interferon sensitivity determining region (ISDR) and the complete NS5A protein may be of importance for response to interferon alpha-based treatment in patients infected with HCV subtype 1a/b. PMID- 15653411 TI - The role of core antigen detection in management of hepatitis C: a critical review. AB - Several assays in research format and two commercial assays for the detection of hepatitis C virus (HCV) core protein or HCV core antigen have been developed in recent years. In order to elucidate the role and significance of HCV core antigen detection in the diagnosis and management of hepatitis C, we reviewed 56 studies published in peer-reviewed journals until September 2004. Evaluations in transfusion settings showed that the HCV core antigen assay detects HCV infection, similarly as nucleic acid techniques (NAT), between 40 and 50 days earlier than the current third generation HCV antibody screening assays. HCV core antigen levels closely track HCV RNA dynamics, and allow clinical monitoring of a patient's therapy, independently of HCV genotype, however, mainly in the samples with HCV RNA levels above 20,000 IU/ml. Considering the lower sensitivity of HCV core antigen detection in comparison to NAT, the HCV core antigen assay is not practical for the determination of the end of treatment response and sustained viral response, but could be useful for the determination of early viral response in the pegylated interferon-alpha and ribavirin treated patients infected with HCV genotype 1. The HCV core antigen detection is a viable tool for study of hepatitis C pathogenesis. The HCV core antigen can be used as a marker of HCV replication in anti-HCV positive individuals in the areas of the world that cannot afford NAT and/or in the settings that are not equipped or competent to perform HCV RNA testing. Because the manufacturer of HCV core antigen assays recently stopped an active marketing of these assays in several countries, it will, unfortunately and probably, never be possible to determine the actual potential and usefulness of HCV core antigen testing in the management of hepatitis C. PMID- 15653412 TI - Genetic variability of the S gene of hepatitis B virus: clinical and diagnostic impact. AB - The genetic variability of hepatitis B virus (HBV) represents a challenge for the sensitivity of immunologic and molecular based assays. Based on sequence divergence in the entire genome of >8%, HBV genomes have been classified into eight groups designated A to H. The genotypes of HBV have distinct geographical distributions. Although preliminary clinical studies seem to indicate that there is an association between HBV genotype and natural history of infection and response to antiviral therapy, further evaluations on larger collectives of patients are necessary to give a clearer picture of the subject. The analytical sensitivity of HBsAg and anti-HBs assays may be dependent on HBV genotype or subtype. The influence of genotypic variability on the sensitivity of nucleic acid amplification tests (NAT) has so far been poorly investigated. Preliminary results show that new real-time NAT detect genotypes A to G with an equal sensitivity. Different mechanisms intervening at the translational or post translational level, including conformational changes, hydrophobic changes, insertion of basic residues and reduced synthesis or secretion of HBsAg may account solely or in conjunction for escape mutations to the immune response and to detection in HBsAg immunassays. The clinical significance of S-gene mutants, needs in analogy to that of HBV genotypes, to be further investigated. HBV mutants are stable over time and can be transmitted horizontally or vertically. The sensitivity of HBsAg assays for mutant detection is continuously improved. Immunoassays based on polyclonal capture antibody show the highest sensitivity for the recognition of recombinant mutants or serum samples harboring mutant forms of HBsAg. However, they do not guarantee full sensitivity. Detection of HBsAg needs to be improved by the introduction of new HBsAg assays able to recognize so far described S-gene mutants and with a lower detection threshold than current immunoassays in order to detect smallest amounts of HBsAg in low level carriers. There is also a need for more complete epidemiological data on the prevalence of HBsAg mutants and strategies for the (differential) screening of mutants need to be developed and evaluated. PMID- 15653413 TI - Cellular response to conditional expression of the hepatitis B virus precore and core proteins in cultured hepatoma (Huh-7) cells. AB - BACKGROUND: The expression of the hepatitis Be antigen (HBeAg) is one of several strategies used by hepatitis B virus (HBV) to ensure persistence. The HBeAg may function as a toleragen in utero and has been shown to regulate the host's immune response. AIM: The aim of this study was to examine the effect of the HBV precore and core protein on cellular gene expression in the hepatoma cell line Huh-7. STUDY DESIGN: Huh-7 cells with tight regulated expression of the HBV core or precore protein were produced using the Tet-Off tetracycline gene expression system. Changes in cellular gene expression in response to core/precore expression compared to Huh-7 cells not expressing the proteins were determined using a commercial high-density oligonucleotide array (Affymetrix Hu95A GeneChip) containing probes for 12,626 full-length human genes. RESULTS: Analysis of differential mRNA gene expression profiles at 7 days post precore and core expression revealed 45 and 5 genes, respectively, with mRNA changes greater than three-fold. The most striking feature was in Huh-7 cells expressing the precore protein in which 43/45 genes were downregulated 3-11-fold. These included genes that encoded products that regulate transcription/DNA binding proteins, cell surface receptors, cell-cycle/nucleic acid biosynthesis and intracellular signalling and trafficking. The only known gene, which was upregulated encoded a cytoskeletal protein. For the core cell line, 4/5 genes were downregulated 3-15 fold upon core induction and included genes that encoded products that affect intermediary metabolism, cell surface receptors and intracellular signalling. The one gene, which was upregulated was a cytokine gene. CONCLUSION: The results of this study show that HBV precore protein has a much greater effect on cellular gene expression in comparison to the core protein, suggesting that core and precore proteins may have diverse effects on cellular functions and equally different roles in modulating HBV pathogenesis. PMID- 15653414 TI - Chronic hepatitis B: recommendations for therapy based on the natural history of disease in Australian patients. AB - BACKGROUND: Chronic hepatitis B infection (CHB) is a major health problem in Australia and worldwide. CHB is associated with significant long-term morbidity and mortality. Well tolerated treatment is now available, however the development of resistance is common and the optimal timing of treatment is yet to be determined. Identifying the factors that influence the natural history of CHB may help determine which patients need treatment and when to start it. OBJECTIVE: To determine the demographics, clinical features and virological profile of Australian patients infected with CHB and the influence of these factors on disease activity and severity. STUDY DESIGN: Review of prospectively collected demographic, clinical and virological features of all patients positive for hepatitis B surface antigen (HBsAg) for more than 6 months who were referred to St. Vincent's Hospital liver clinics. Age, sex and ethnicity were correlated with hepatitis B e antigen status (HBeAg), HBV replication status (ALT and HBV DNA), genotype and liver histology. RESULTS: 703 chronic hepatitis B surface antigen positive patients were identified. The patients were predominantly male with an average age of 44. Eighty two percent of patients were born overseas, primarily from Asian (65%) and Mediterranean countries (14%). Two thirds (426) had an elevated ALT (median 79) at presentation. HBeAg was positive in 37%. Active viral replication, defined as abnormal ALT or positive HBVDNA, was present in 74%, 48% of whom were HBeAg negative. In a subset of 103 patients genotyped, 8% had genotype A, 29% B, 41% C and 22% D. Genotype correlated with ethnicity; patients infected with genotypes A were predominantly Caucasian, B and C were Asian, and D were Mediterranean. Of 296 (42%) patients who underwent liver biopsy, 76 (27%) had advanced fibrosis. Advanced fibrosis was associated with increasing age and Mediterranean ethnicity. CONCLUSION AND RECOMMENDATIONS: Perinatal or early childhood transmission is predominant mode of infection in Australia. Two thirds of this cohort had active replication and were at increased risk of developing cirrhosis and/or hepatoma. Advanced disease was associated with age and ethnicity. HBeAg negative CHB accounts for almost half of all those with active viral replication. This parallels the rise in this form of CHB in Asia and the Mediterranean basin. Screening should be offered to people born in, or with parents born in areas of high endemnicity. To detect the development of active disease, patients with positive HBsAg but normal ALT should have liver function tests done 6 monthly and those with elevated ALT should be referred for consideration of therapy, irrespective of HBeAg status. PMID- 15653415 TI - Molecular epidemiology of hepatitis A in Noord-Brabant, The Netherlands. AB - BACKGROUND: Previous studies on the molecular epidemiology of hepatitis A virus (HAV) in Amsterdam, The Netherlands, show that subgenotype 1A is mainly seen among homosexual men practising anonymous oral-anal sex in saunas and darkrooms, while subgenotype 1B is usually detected among children originating from Morocco, and subgenotype 3A is mostly found among travellers to Pakistan. OBJECTIVE: We studied the genotype distribution in a more rural area of The Netherlands, Noord Brabant, and compared it with Amsterdam. STUDY DESIGN: We collected blood and feces samples from 34 HAV IgM(+) individuals who were reported from August 2001 March 2003 at the Municipal Health Service (MHS) Heart for Brabant (Brabant). We also collected feces samples from nine household contacts of whom the HAV IgM status was not known. HAV RNA was isolated and subsequently amplified by reverse transcriptase polymerase chain reaction (RT-PCR) at the VP1-P2a and the VP3-VP1 region, sequenced and analysed. RESULTS AND CONCLUSIONS: In most cases, relations between risk groups and HAV subgenotypes in Noord-Brabant were similar to those in Amsterdam. Next to genotypes 1 and 3 we also detected a genotype 2/7 strain in a Noord-Brabant case. Also, in contrast to the Amsterdam study, sporadic transmission occurred among various risk groups. Children involved in a school related outbreak were infected with strains identical to one that was previously isolated from a man who has sex with men (MSM). Also, Dutch patients having no epidemiological link with Turkish or Moroccan children harboured strains imported from high-endemic countries. Furthermore, we report a special case in which HAV may be causally involved in meningitis. The results of this study show that the molecular epidemiology of HAV in The Netherlands can be more complicated than previously anticipated and that HAV phylogenetic studies can provide important information for the design of appropriate public health measures. PMID- 15653416 TI - A novel I-TAC promoter polymorphic variant is functional in the presence of replicating HCV in vitro. AB - BACKGROUND: Chemokines are strong candidate genes for outcome of HCV infection. I TAC is a chemokine known to be involved in the inflammatory process of HCV infection, and its expression is upregulated in chronic hepatitis C (CHC). OBJECTIVES: The aim of this study was to investigate genetic variability in the I TAC promoter and to determine the correlation of these variants with HCV disease progression. STUDY DESIGN: I-TAC genotyping was performed in 60 chronic HCV patients and 60 controls using GeneScan analysis. Functional analysis of the I TAC promoter was performed with the aid of luciferase reporter constructs transfected into Huh-7 cells or Huh-7 cells harbouring HCV genomic and sub genomic replicons. Cytokine induced production of I-TAC from whole blood cultures was measured using enzyme-linked immunosorbent assay (ELISA). RESULTS: Sequencing of approximately 1 kb upstream of the I-TAC gene start codon revealed the presence of a novel 5 bp deletion mutant (-599del5) in a number of chronic HCV patients. Analysis of the functional potential of this deletion revealed no transcriptional change in Huh-7 cells transfected with luciferase reporter constructs, and this was confirmed in cytokine stimulated whole blood cultures where similar levels of I-TAC were liberated regardless of -599del5 genotype. Conversely, the -599del5 deletion variant significantly reduced transcriptional activity of the I-TAC promoter in the presence of replicating HCV. The distribution frequency of the allele was found to be significantly increased in a chronically HCV infected population compared to healthy controls. CONCLUSIONS: The novel I-TAC -599del5 promoter polymorphism is a functional variant in the presence of replicating HCV. Furthermore, this deletion mutant is significantly increased in a chronic HCV cohort and may predispose to HCV disease susceptibility. PMID- 15653417 TI - Human leukocyte antigen phenotypes and hepatitis C viral load. AB - BACKGROUND: After hepatitis C virus (HCV) infection, 55% to 85% of patients become chronic carriers. HCV-RNA could be detected in the sera of these patients though the viral load varies. Various factors may be involved in determining the viral load. OBJECTIVES: In this study, we want to investigate the relationship between human leukocyte antigen phenotypes and hepatitis C viral load. STUDY DESIGN: One hundred and sixty HCV-RNA positive subjects were investigated in this study. RESULTS: We have analyzed 160 HCV-RNA positive subjects and found that lower HCV viral load is significantly associated with HBsAg-positivity (P = 0.017) but not age, gender, or mixed infection (infection with different HCV genotypes). One hundred and fifty-four HBsAg-negative subjects were further analyzed to explore the relationship between human leukocyte antigen (HLA) phenotypes and HCV viral load. Subjects with certain HLA alleles (A*34, B*56, DRB1*1502) have significantly lower viral load than those without these alleles (P = 0.0074, 0.0039 and 0.016, respectively) while those with HLA-B*4001 have significantly higher viral load (P = 0.0026). Furthermore, lower viral load was significantly associated with HLA-DRB1 heterozygosity in subjects with HLA-B heterozygosity (P = 0.048). CONCLUSIONS: Our data suggests a role for host immunogenetic factors in determining viral load during HCV infection. PMID- 15653418 TI - Longitudinal study in HIV/HCV-coinfected HAART-naive patients and role of HCV genotype. AB - To evaluate the impact of highly active antiretroviral therapy (HAART) on the course of hepatitis C (HCV) infection, we studied the biological and virological characteristics of 23 HCV/HIV-coinfected HAART-naive patients. The HCV genotype, HCV and HIV viral loads, serum alanine aminotransferase, CD4+ and CD8+ cell/mm3 were determined at baseline, 1 month, 6 months and 12 months after initiation of HAART. Results were analyzed both in terms of total population and of HCV genotype. The study of the total population suggests that this therapy did not determine a significant alteration of HCV viremia and levels of ALT, while a significant decrease in HIV viremia (-1.7log10 at one year from the start of HAART) and increase in CD4+ counts was observed (P < 0.005). The biological and virological parameters of HCV/HIV coinfection differed according to the HCV genotype. In particular, only genotype 4 showed a significant inverse correlation between HCV and HIV viral loads. PMID- 15653419 TI - In vivo immunization by vaccine therapy following virus suppression by lamivudine: a novel approach for treating patients with chronic hepatitis B. AB - BACKGROUND: Both the hepatitis B virus (HBV) and the immune response of the hosts to HBV play important roles in the pathogenesis of chronic hepatitis B (CHB). Lamivudine is a potent antiviral agent with minimal immune modulator capacity. Moreover, lamivudine causes severe side effects like breakthrough of HBV DNA and breakthrough hepatitis in patients with CHB. On the other hand, vaccine therapy, a recently-developed immune therapy, exhibits potent immune modulatory potentials and almost no side effects, but possesses little antiviral capacity in patients with CHB. OBJECTIVES: The aim of this clinical trial is to evaluate the efficacy of a combination therapy of lamivudine and vaccine in patients with CHB. STUDY DESIGN: Seventy-two patients with CHB (hepatitis B e antigen (HBeAg)-positive, 40; antibody to HBeAg (anti-HBe)-positive, 32). All patients received lamivudine at a dose of 100 mg daily for 12 months. Fifteen patients (HBeAg+, 9; anti-HBe+, 6) receiving oral lamivudine were also given a vaccine containing 20 microg of hepatitis B surface antigen, intradermally, once every 2 weeks for 12 times (combination therapy). RESULTS: Twelve months after the start of therapy, serum HBV DNA became negative in 9 of 9 (100%) HBeAg+ CHB patients receiving combination therapy and in 15 of 31 (48%) HBeAg+ CHB patients receiving lamivudine monotherapy (P < 0.05). The rate of seroconversion from HBeAg to anti HBe was also significantly higher in patients receiving combination therapy (56% versus lamivudine monotherapy, 16%, P < 0.05). Of the 57 patients receiving lamivudine monotherapy, breakthrough of HBV DNA was found in 10 and breakthrough hepatitis was found in 4; however, these were not seen in any patient receiving combination therapy. CONCLUSIONS: Combination therapy represents a better therapeutic regimen with few complications in patients with CHB. PMID- 15653420 TI - HBV reactivation after kidney transplantation. AB - Recent studies suggest that reappearance of hepatitis B surface antigen (HBsAg) and loss of anti-HBs antibodies may be common events in bone marrow recipients and patients with chemotherapy. In this study, we reviewed the virologic laboratory records from kidney recipients. Out of 1512 patients, 228 had been diagnosed with resolved HBV infection (anti-HBc positive, HBsAg negative) but normal liver enzyme levels prior to kidney transplantation. Reappearance of HBsAg after kidney transplantation was observed in two (0.9%) of those patients, which may be attributed to reactivation of a latent infection or to a new HBV infection. In both of the patients, HBV infection may have been reactivated although immunosuppression was just on a low level over the whole period. We conclude that natural immunity to HBV may not protect against reactivation in patients with suppression of the immune system. Periodic follow-up of HBV serology for early diagnosis of reactivation is highly recommended in transplant recipients. PMID- 15653421 TI - Quantification of the newly detected lamivudine resistant YSDD variants of Hepatitis B virus using molecular beacons. AB - A real-time based amplification assay with molecular beacons was used to detect and quantify PCR amplicons to discriminate between the newly described Lamivudine resistant YSDD variant, a known YIDD variant and wild-type Hepatitis B virus (HBV) DNA in the YMDD region of the polymerase gene. Using this assay, we retrospectively analysed samples from two HBV chronically infected Asian twin sisters, starting 9 weeks before therapy, during and between two periods of treatment with Lamivudine. In order to analyse more accurately the dynamics of variant DNA loads during and after therapy, this real time assay was compared to three other mutation analysis techniques, restriction fragment length polymorphism (RFLP), InnoLipa HBV-DR assay and direct sequence analysis. With this technique, new information on the dynamics of variants during and after therapy was obtained. PMID- 15653422 TI - Sexual transmission of hepatitis B infection despite the presence of hepatitis B virus immunity in recipients of allogeneic bone marrow transplantation. AB - BACKGROUND: After hematopoietic cell transplantation (HCT), hepatitis due to hepatitis B virus (HBV) rarely occurred beyond the initial 12 months after transplantation. OBJECTIVES: We investigated the cause of "late" hepatitis due to HBV infection in two recipients after allogeneic HCT. STUDY DESIGN: Two male patients with acute myeloid leukemia and light chain myeloma, respectively, developed HBV-related hepatitis more than 2 years after HCT. All serum samples collected from the recipients, donors and their respective spouses were tested for HBV DNA by nested PCR, and if positive further quantified by Digene Hybrid Capture assay II. The HBV genotype was determined by PCR and sequencing. RESULTS: Genotypic analysis suggested that the cause of "late" hepatitis was due to acute HBV infection transmitted from their respective spouse. CONCLUSION: Our findings suggested that sexual precautions should be taken in these patients after HCT. Alternatively, or even additionally, active vaccination should be delivered to these patients once they have lost their HBV immunity. PMID- 15653423 TI - Prevalence of chronic HBV infection among 13,581 women at reproductive age in Greece. A prospective single center study. PMID- 15653424 TI - Polyarteritis nodosa after interferon treatment for chronic hepatitis C. PMID- 15653425 TI - Structural flexibility of small GTPases. Can it explain their functional versatility? AB - Multiple interactions with many different partners are responsible for the amazing functional versatility of proteins, especially those participating in cellular regulation. The structural properties that could facilitate multiple interactions are examined for small GTPases. The role of cellular constraints, compartmentation and scaffolds on protein-protein interactions is considered. PMID- 15653426 TI - Neuroendocrine regulation of salivary IgA synthesis and secretion: implications for oral health. AB - Secretory immunoglobulin A (S-IgA) represents the main adaptive immune mechanism in the oral cavity. The regulation of secretion and synthesis of S-IgA is not only dependent on prior antigenic stimulation, but is also under strong neuroendocrine control. Thus, alterations in neuroendocrine functioning (such as induced by stress, exercise, pregnancy, menstrual cycle, and pharmacological interventions) may affect salivary IgA levels. This review deals with the neuroendocrine regulation of synthesis and secretion of salivary IgA and its potential role in the maintenance of oral health. PMID- 15653427 TI - Structural characterization of extracellular lipase from Streptomyces rimosus: assignment of disulfide bridge pattern by mass spectrometry. AB - The cloning, sequencing and high-level expression of the gene encoding extracellular lipase from Streptomyces rimosus R6-554W have been recently described, and the primary structure of this gene product was deduced using a bioinformatic approach. In this study, capillary electrophoresis-on-the-chip and mass spectrometry were used to characterize native and overexpressed extracellular lipase protein from S. rimosus . The exact molecular mass of the wild-type and the overexpressed lipase, determined by matrix-assisted laser desorption/ionization (MALDI) mass spectrometry, were in excellent agreement (Deltam=0.11 Da and Deltam=0.26 Da, respectively) with a value of 24165.76 Da calculated from the structure deduced from the nucleotide sequence, considering the mature enzyme with all six cysteines forming disulfide bridges. The primary structure derived from the nucleotide sequence was completely verified using a combination of tryptic digestion and formic acid cleavage of the protein, followed by peptide mass fingerprinting. Selected peptides were further investigated by MALDI low-energy collision-induced dissociation hybrid tandem mass spectrometry, allowing the unambiguous determination of their predicted amino acid sequence. No post-translational modifications of mature S. rimosus lipase were detected. Comparison of the peptide mass fingerprints from the reduced and non-reduced overexpressed enzyme unequivocally revealed three intramolecular disulfide bonds with the following linkages: C27-C52, C93-C101 and C151-C198. PMID- 15653428 TI - Thermal unfolding of ribonuclease T1 studied by multi-dimensional NMR spectroscopy. AB - Thermal unfolding of ribonculease (RNase) T1 was studied by 1H nuclear Overhauser enhancement spectroscopy (NOESY) and 1H- 15N heteronuclear single-quantum coherence (HSQC) NMR spectroscopy at various temperatures. Native RNase T1 is a single-chain molecule of 104 amino acid residues, and has a single alpha-helix and two beta-sheets, A and B, which consist of two and five strands, respectively. Singular value decomposition analysis based on temperature dependent HSQC spectra revealed that the thermal unfolding of RNase T1 can be described by a two-state transition model. The midpoint temperature and the change in enthalpy were determined as 54.0 degrees C and 696 kJ/mol, respectively, which are consistent with results obtained by other methods. To analyze the transition profile in more detail, we investigated local structural changes using temperature-dependent NOE intensities. The results indicate that the helical region starts to unfold at lower temperature than some beta-strands (B3, B4, and B5 in beta-sheet B). These beta-strands correspond to the hydrophobic cluster region, which had been expected to be a folding core. This was confirmed by structure calculations using the residual NOEs observed at 56 degrees C. Thus, the two-state transition of RNase T1 appears to involve locally different conformational changes. PMID- 15653429 TI - Conserved asparagine residue 54 of alpha-sarcin plays a role in protein stability and enzyme activity. AB - Asparagine 54 of alpha-sarcin is a conserved residue within the proteins of the ribotoxin family of microbial ribonucleases. It is located in loop 2 of the protein, which lacks repetitive secondary structure elements but exhibits a well defined conformation. Five mutant variants at this residue have been produced and characterized. The spectroscopic characterization of these proteins indicates that the overall conformation is not changed upon mutation. Activity and denaturation assays show that Asn-54 largely contributes to protein stability, and its presence is a requirement for the highly specific inhibitory activity of these ribotoxins on ribosomes. PMID- 15653430 TI - Replacement of the interchain disulfide bridge-forming amino acids A7 and B7 by glutamate impairs the structure and activity of insulin. AB - Insulin contains three disulfide bonds, one intrachain bond, A6-A11, and two interchain bonds, A7-B7 and A20-B19. Site-directed mutagenesis results (the two cysteine residues of disulfide A7-B7 were replaced by serine) showed that disulfide A7-B7 is crucial to both the structure and activity of insulin. However, chemical modification results showed that the insulin analogs still retained relatively high biological activity when A7Cys and B7Cys were modified by chemical groups with a negative charge. Did the negative charge of the modification groups restore the loss of activity and/or the disturbance of structure of these insulin analogs caused by deletion of disulfide A7-B7? To answer this question, an insulin analog with both A7Cys and B7Cys replaced by Glu, which has a long side-chain and a negative charge, was prepared by protein engineering, and its structure and activity were analyzed. Both the structure and activity of the present analog are very similar to that of the mutant with disulfide A7-B7 replaced by Ser, but significantly different from that of wild type insulin. The present results suggest that removal of disulfide A7-B7 will result in serious loss of biological activity and the native conformation of insulin, even if the disulfide is replaced by residues with a negative charge. PMID- 15653431 TI - Stereospecificity of horseradish peroxidase. AB - We report here on the stereospecificity observed in the action of horseradish peroxidase (HRPC) on monophenol and diphenol substrates. Several enantiomers of monophenols and o-diphenols were assayed: L-tyrosinol, D-tyrosinol, L-tyrosine, DL-tyrosine, D-tyrosine, L-dopa, DL-dopa, D-dopa, L-alpha-methyldopa, DL-alpha methyldopa, DL-adrenaline, D-adrenaline, L-isoproterenol, DL-isoproterenol and D isoproterenol. The electronic density at the carbon atoms in the C-1 and C-2 positions of the benzene ring were determined by NMR assays (delta1 and delta2). This value is related to the nucleophilic power of the oxygen atom of the hydroxyl groups and to its oxidation-reduction capacity. The spatial orientation of the ring substituents resulted in lower Km values for L- than for D-isomers. The kcat values for substrates capable of saturating the enzyme were lower for D- than for L-isomers, although both have the same delta1 and delta2 NMR values for carbons C-1 and C-2, and therefore the same oxidation-reduction potential. In the case of substrates that cannot saturate the enzyme, the values of the binding constant for compound II (an intermediate in the catalytic cycle) followed the order: L-isomer>DL-isomer>D-isomer. Therefore, horseradish peroxidase showed stereospecificity in its affinity toward its substrates (K m) and in their transformation reaction rates (k cat). PMID- 15653432 TI - Novel thioredoxin targets in Dictyostelium discoideum identified by two-hybrid analysis: interactions of thioredoxin with elongation factor 1alpha and yeast alcohol dehydrogenase. AB - Thioredoxins (Trx) are ubiquitous dicysteine proteins capable of modulating enzymes and other cellular targets through specific disulfide-dithiol redox changes. They are unique in that a large number of very diverse metabolic systems are addressed and redox-regulated in bacteria, animal, and plant cells, but the finite number of thioredoxin interaction partners is still unknown. Two-hybrid methodology should provide a rational way to establish thioredoxin functions in a given organism. We report a search for physiological target proteins of thioredoxin1 in the social amoeba Dictyostelium discoideum , which possesses three developmentally regulated thioredoxin genes, all of which lack functional characterisation. A two-hybrid approach identified at least seven bona fide thioredoxin partners, including oxidoreductases, proteins of the ribosomal translation apparatus, and the cytoskeletal protein filopodin. With the exception of ribonucleotide reductase, none of these systems had previously been linked to specific redox modulation. Molecular interactions in two of the new thioredoxin/target protein couples were verified by biochemical studies: (1) thioredoxin1 and the abundant elongation factor 1alpha from D. discoideum form the mixed heterodisulfide characteristic of the thioredoxin mechanism of action; and (2) reduced thioredoxin, but not glutathione, strongly inhibits yeast alcohol dehydrogenase catalysis of ethanol oxidation. PMID- 15653433 TI - Functional characterization of the postulated intramolecular sphingolipid activator protein domain of human acid sphingomyelinase. AB - Degradation of membrane-bound sphingomyelin to phosphorylcholine and ceramide is catalyzed by the water-soluble lysosomal acid sphingomyelinase (A-SMase). The presence of sphingolipid activator proteins (Saps: saposins A-D; GM2 activator) is not essential to mediate this reaction at the water-lipid interface in vivo . A hypothesis based on amino acid sequence alignments suggests that the enzyme possesses an N-terminal saposin-homologous domain, which may facilitate the enzymatic reaction at the interface. We mutated one homologous and three conserved amino acid residues of this domain and studied the activity of the variant enzymes using different sphingomyelin degradation assays. A variant with an exchange of a conserved amino acid residue, Pro153Ala, still exhibited enzyme activity of approximately 52% of normal in a detergent-containing micellar assay, but only 13% of normal in a detergent-free liposomal assay system, which suggests that the Sap-homologous domain fulfills membrane-disturbing functions. Addition of saposin C to the liposomal assay mixtures increased the Pro153Ala variant sphingomyelinase activity to 46% of normal, indicating that the variant saposin like domain can be substituted by the presence of the sphingolipid activator protein. On the other hand, the addition of saposin C did not result in complete restoration of the variant activity. Thus, the Sap-like domain may also have another role, e.g., to stabilize the fold of acid sphingomyelinase, which cannot be compensated by the presence of saposin C or a detergent. Such an essential second function of the saposin-like domain as an integral part of acid sphingomyelinase is confirmed by our observation that the Lys118Glu, Cys120Ser and Cys131Ser variants were almost completely devoid of activity in the detergent containing micellar assay system as well as in the liposomal assay system in the presence of saposin C. PMID- 15653434 TI - St. John's wort (Hypericum perforatum) counteracts cytokine-induced tryptophan catabolism in vitro. AB - St. John's wort (Hypericum perforatum) is an ancient folk remedy that has antiviral and antibacterial properties. Anti-inflammatory effects of the plant have been described and the application of H. perforatum extract as an effective antidepressant is well established. In this study we assayed the effect of H. perforatum extract on cytokine-induced tryptophan degradation in human peripheral blood mononuclear cells. Simultaneously, changes in the production of the immune activation marker neopterin were monitored. Both these biochemical pathways are triggered by interferon-gamma. Our results show that extracts of H. perforatum strongly down-regulate mitogen-mediated tryptophan degradation in a dose dependent manner. This effect seems to be based on a suppressive activity of H. perforatum on activated immunocompetent cells, resulting in a diminished production of interferon-gamma. In line with this finding, neopterin synthesis was strongly down-regulated by the plant extract. Our results suggest that the reduction of tryptophan degradation by H. perforatum might be important in the action of the plant as an antidepressant. PMID- 15653435 TI - Nuclear fibroblast growth factor-2 interacts specifically with splicing factor SF3a66. AB - Fibroblast growth factor 2 (FGF-2) has a dual role as a classical extracellular signaling protein and as an intracellular factor. Isoforms of FGF-2, resulting from alternatively used start codons on one mRNA species, locate differentially to nuclear compartments. In this study we aimed to analyze functions of intracellular FGF-2 by identification of interacting proteins. We identified the 66-kDa subunit of splicing factor 3a (SF3a66) as a binding partner in a yeast two hybrid screen and confirmed this interaction by pull-down assays. The splicing factor interacted with the 18-kDa (FGF-2(18)) and with the 23-kDa (FGF-2(23)) isoforms, indicating an interaction with a domain common to both isoforms. Moreover, FGF-2 interacted with the C-terminus of SF3a66, a sequence that has not previously been assigned a functional role. In a functional neurite outgrowth assay, SF3a66 enhanced neurite lengths similar to FGF-2(18). We have previously identified the spliceosomal assembly factor survival of motoneuron (SMN) protein as a protein interacting specifically with the FGF-2(23) isoform [Claus et al., J. Biol. Chem. 278 (2003), 479-485]. The identification of two FGF-2 interacting proteins from the same biochemical pathway suggests a novel intranuclear role of FGF-2. PMID- 15653436 TI - Pathophysiology and diagnostic value of urinary trypsin inhibitors. AB - Inflammation is an important indicator of tissue injury. In the acute form, there is usually accumulation of fluids and plasma components in the affected tissues. Platelet activation and the appearance in blood of abnormally increased numbers of polymorphonucleocytes, lymphocytes, plasma cells and macrophages usually occur. Infectious disorders such as sepsis, meningitis, respiratory infection, urinary tract infection, viral infection, and bacterial infection usually induce an inflammatory response. Chronic inflammation is often associated with diabetes mellitus, acute myocardial infarction, coronary artery disease, kidney diseases, and certain auto-immune disorders, such as rheumatoid arthritis, organ failures and other disorders with an inflammatory component or etiology. The disorder may occur before inflammation is apparent. Markers of inflammation such as C-reactive protein (CRP) and urinary trypsin inhibitors have changed our appraisal of acute events such as myocardial infarction; the infarct may be a response to acute infection and (or) inflammation. We describe here the pathophysiology of an anti inflammatory agent termed urinary trypsin inhibitor (uTi). It is an important anti-inflammatory substance that is present in urine, blood and all organs. We also describe the anti-inflammatory agent bikunin, a selective inhibitor of serine proteases. The latter are important in modulating inflammatory events and even shutting them down. PMID- 15653437 TI - Apolipoprotein H (apoH)-dependent autoantibodies and apoH protein polymorphism in selected patients showing lupus anticoagulant activity. AB - Apolipoprotein H (apoH) is considered to be a necessary cofactor for the binding of certain antiphospholipid antibodies to anionic phospholipids. Some apoH dependent antiphospholipid antibodies also exert lupus anticoagulant (LA) activity, which seems to depend on antiphospholipid antibody epitope specificity. The aim of this study was to evaluate whether the presence of less frequent apoH alleles may induce structural or conformational changes in these "LA-dependent" regions that may initiate more frequent autoimmune responses in subjects. We selected patients with confirmed LA activity and none or low titers of anticardiolipin antibodies that had been sent to the laboratory for routine antiphospholipid antibody determination. Many of them had some clinical manifestation of antiphospholipid syndrome. Antibodies to apoH were determined with a commercially available anticardiolipin/apoH ELISA kit. ApoH protein polymorphism (apoH phenotype) was demonstrated by isoelectric focusing and immunoblotting. Our results showed that 47/74 (63.5%) of our selected LA-positive patients also had elevated apoH-dependent antiphospholipid antibody titers. These results point to two subgroups of patients according to the LA potency of apoH dependent antibodies. A strong positive correlation (non-linear or linear) for apoH-dependent antibody titers and LA activity was observed in both subgroups of patients. In this study, we did not find significant differences in the distribution of apoH phenotypes among control subjects and patients with apoH dependent/LA-positive auto- antibodies. PMID- 15653438 TI - Differential production of immunoglobulin classes and subclasses by mucosal-type human B-lymphocytes exposed in vitro to CpG oligodeoxynucleotides. AB - As B-lymphocytes play an important role in innate and adaptive immunity, we aimed to examine the effects of CpG oligodeoxynucleotides (ODNs) on purified tonsil originating CD19+ B-cells, representing mucosal B-cells. We screened various K type ODNs, reactive with human B-cells, and tested for the production of immunoglobulins in vitro. Using one CpG-ODN, DSP30, we observed that it could upregulate not only Toll-like receptor 9 (TLR9) mRNA expression in activated B cells, but also the early expression of CD69 followed by the sequential expression of CD80, CD86 and the nuclear factor (NF)-kappaB pathway. Furthermore, mRNA expression of certain B-cell-derived cytokines was influenced by exposure to DSP30, with a strong upregulation of interleukin 6 (IL-6) and downregulation of IL1-beta. Stimulation of B-cells, co-stimulated with IL-2, IL-10 and soluble CD40 ligand (sCD40L) with different CpG-ODNs, had differing effects on the terminal differentiation in vitro of B-cells into immunoglobulin-secreting cells. TLR9 is involved in innate immunity and the recognition of bound CpG DNA from invading bacterial pathogens. As tonsillar B-cells are mucosal-type B-lymphocytes, this study suggests that CpG-ODNs show promise as mucosal adjuvants in modulating the local production of immunoglobulins of certain classes and subclasses, a crucial issue in vaccine perspectives. PMID- 15653439 TI - The predictive power of serum kappa/lambda ratios for discrimination between monoclonal gammopathy of undetermined significance and multiple myeloma. AB - The predictive power of serum kappa/lambda ratios on initial presentation of immunoglobulin G (IgG) or IgA monoclonal component was studied to differentiate between monoclonal gammopathy of undetermined significance (MGUS) and multiple myeloma (MM) patients. The retrospective study involved 145 patients clinically diagnosed with monoclonal gammopathy of undetermined significance or multiple myeloma, who had serum M-protein IgG <35 g/L or IgA <20 g/L at M-protein detection. Serum light chains kappa and lambda were measured by fixed-time nephelometry. Test performance indices, predictive values and likelihood ratios were calculated according to the Weissler recommendation. MM patients were considered as diseased and MGUS patients as non-diseased in order to estimate the performance characteristics of serum kappa/lambda ratios. There was a statistically significant difference in kappa/lambda ratios distribution between both groups of patients, in both M-protein kappa-type (Mann-Whitney U=168, p<0.001) and in M-protein lambda-type (Mann-Whitney U=143, p<0.001). Negative likelihood ratios at threshold levels of 0.6 and 4.2 were 2.17- and 3.32-fold greater, respectively, than positive likelihood ratios, so that the predictive power of a serum kappa/lambda ratio within these limits is better in ruling out (negative predictive power) than ruling in disease (positive predictive power). The post-test characteristics of a serum kappa/lambda ratio interval between 0.6 and 4.2 in discriminating MGUS from MM in our geographic population were: sensitivity 0.96 (0.93-0.99 95% CI); specificity 0.70 (0.63-0.77); positive predictive value 0.68 (0.64-0.73); negative predictive value 0.96 (0.94-0.99); likelihood ratios (+)LR 3.23 (2.68-4.04); and (-)LR 17.16 (11.00-63.00). Thus, serum M-protein with a kappa/lambda ratio between 0.6 and 4.2 increases the posterior probability of MGUS from 0.60 to 0.96 in asymptomatic patients, for whom only monitoring may be suggested when the serum kappa/lambda ratio is within these limits. PMID- 15653440 TI - The soluble transferrin receptor (sTfR)-ferritin index is a potential predictor of celiac disease in children with refractory iron deficiency anemia. AB - The soluble transferrin receptor (sTfR) distinguishes iron deficiency anemia from other types of anemia. Refractory iron deficiency anemia is often the onset symptom in malabsorption-induced celiac disease. We evaluated whether sTfR levels distinguish celiac disease-associated iron deficiency anemia from iron deficiency anemia of other origin. To this aim we measured sTfR and ferritin levels and their ratio (the sTfR/ferritin index) and other hematological parameters in 42 anemic children (20 with and 22 without celiac disease) vs. 22 non-anemic children with celiac disease and 31 healthy controls (age range 4-12 years). Hemoglobin parameters, mean cell volume, and serum iron and ferritin levels were decreased to a similar extent in the anemic patients (celiac and non-celiac). The sTfR level in non-anemic celiac patients was similar to that of normal controls (1.7+/-0.35 mg/L), whereas it was significantly increased in non-celiac and celiac anemic patients (2.2+/-0.5 mg/L, p<0.05 and 2.7+/-1.2 mg/L, p<0.001, respectively). The sTfR/ferritin index was also increased more in the anemic celiac patients (mean 4.4, range 1.5-12.0) than in anemic non-celiac children (mean 2.6, range 1.4-4.0) compared with non-anemic children (mean 1.2, range 0.7 2.0). Differences were more pronounced when ferritin was <5 ng/mL. Thus, the sTfR/ferritin index may be a predictive measure in discriminating anemic patients with celiac disease from those without celiac disease. PMID- 15653441 TI - Monitoring of fibrinolysis parameters during myocardial revascularization according to type of procedure. AB - The aim of the study was to assess the effect of cardiothoracic surgery on the dynamics of plasminogen, D-dimers and plasminogen activator inhibitor (PAI-I) during the first 24 h after surgery. The study included 14 patients operated with (on-pump) and 14 without (off-pump) the use of extracorporeal circulation (ECC). Blood sampling was carried out on induction of anesthesia (timepoint 1), on introduction of heparin (point 2) and protamine (point 3), at the end of surgery (point 4), and the next morning (point 5). Relative to point 1, the utilization of plasminogen at point 2 was 24% and 17% in the on-pump and off-pump groups, respectively (p=0.001 both). Increased D-dimer concentration from the baseline was more pronounced in the on-pump group (p=0.001). At point 5, D-dimer concentrations were comparable in both groups and different from baseline levels. PAI-I activity showed within-group differences from baseline at point 5 in the off-pump group (p=0.001), and at points 3 and 5 in the on-pump group (p=0.002 and 0.001, respectively). At point 5, the activity of PAI-I was comparable in both groups, yielding p=0.001 vs. baseline. Fibrinolysis was more pronounced and more dynamic in the on-pump group due to activation of the systemic inflammatory response induced by the use of ECC. In the off-pump group, fibrinolysis was a normal physiological response to the surgical procedure. PMID- 15653442 TI - Serum amyloid A protein levels as a possible aid in the diagnosis of acute appendicitis in children. AB - Hematological and biochemical tests, including white blood cell count (WBC), C reactive protein (CRP) and other acute-phase reactants, have been used in the diagnosis of acute appendicitis. However, there is controversy among physicians about the value of this practice in children. The objective of our study was to evaluate serum amyloid A protein (SAA) levels in children with confirmed acute appendicitis and to compare the sensitivity and specificity of this marker of inflammation with those for WBC and CRP. A prospective cohort study of 60 children admitted with abdominal pain to rule out appendicitis was used in the study. Of these, 42 underwent surgery, while 18 children who had spontaneous amelioration within 24 h of admission were not operated on and served as controls. WBC and serum SAA and CRP levels were obtained preoperatively. Serum concentrations of the analytes were determined with particle-enhanced immunonephelometric methods. Patients with acute appendicitis had WBC, SAA and CRP levels higher than those of the control group (p<0.001). There was no appendicitis patient with a normal SAA value, while 21.4% of the patients had CRP values within the normal range. The performance of each test was measured by receiver-operating characteristic curves. Area under the curve (AUC) values were 0.849 for WBC, 0.868 for CRP and 0.964 for SAA. The sensitivity and specificity of these methods were 76% and 75% for WBC>10.0 x 10(9) /L, 62% and 94% for CRP>10 mg/L and 86% and 83% for SAA >45.0 mg/L, respectively. Circulating SAA levels have better discriminatory value than WBC or CRP in the assessment of acute appendicitis in children. Thus, this test appears to be of higher value than the current standards of care in the diagnosis of this condition. PMID- 15653443 TI - External failures as descriptor of quality in clinical microbiology laboratory services. AB - This study employed customer feedback surveys to assess the quality of the laboratory services provided by an accredited clinical microbiology laboratory. The purpose of the study was to classify external failures and determine the quantity of such errors, and to establish how much time was spent processing and correcting these errors, together with the costs arising. The study indicates that the quality of laboratory test results can be regarded as good based on the accuracy of the result. Customer needs were also well taken into account in practice because of the good quality of the service. However, improvements are required in providing customers with test results without delay. The cost of processing and correcting such errors was negligible. The use of a customer feedback system allows systematic monitoring of external failures. In the accredited laboratory under study, it is unnecessary to monitor continually the costs and working time involved in processing and correcting external failures. The most essential point is to monitor the effects of the quality measures taken to reduce the number of failures. PMID- 15653444 TI - The effects of extracorporeal cardiopulmonary circulation on serum total and free prostate specific antigen levels. AB - The aim of this study was to investigate whether or not there is an increase in serum total and free prostate specific antigen levels (t-PSA, f-PSA) in patients with and without benign prostatic hyperplasia (BPH) undergoing extracorporeal circulation during cardiovascular bypass. The study included a total of 50 men. Of these, 35 patients underwent elective coronary artery bypass grafting with extracorporeal circulation [with (n=20, group I) and without (n=15, group II) BPH]. Another 15 patients underwent renal or ureteral surgery (group III) and served as a control group. Serum t-PSA and f-PSA levels were measured before surgery and 3 h and 3 days after surgery. All patients underwent urethral catheterization 24 h before surgery. In groups I and II, patients had an increase in t-PSA 3 h after surgery compared to baseline values (p=0.0001 and p=0.011, respectively). Also, 3 days after surgery, mean t-PSA levels were higher than baseline levels (p=0.004) in group I. Serum t-PSA levels were higher at 3 h than at day 3 in groups I and II (p=0.003 and p=0.02, respectively). Mean serum f-PSA levels obtained 3 h after surgery were increased in both groups I and II when compared to baseline values (p=0.0001 and p=0.001, respectively). There was no significant difference between f-PSA values before and 3 days after surgery in all groups. In the control group, there was no significant increase in either serum t-PSA or f-PSA levels obtained at different times. There was a modest increase in the percentage of free prostate specific antigen (% f-PSA) 3 h after the operation in group II (p=0.025); the values returned to baseline within 3 days. It was suggested that t-PSA and f-PSA serum levels increase due to extracorporeal cardiopulmonary circulation as a consequence of ischemic damage to the prostate. In patients with BPH, this rise may be higher than in those without BPH, because BPH patients generally have larger prostate volumes that may be more vulnerable to ischemic damage. Because f-PSA has a shorter half-life, postsurgical levels of f-PSA may not show this ischemic damage to the prostate during the late postoperative period and f-PSA appears to be cleared more rapidly than t-PSA. PMID- 15653445 TI - Quantitative automated human chorionic gonadotropin measurement in urine using the Modular Analytics E170 module (Roche). AB - Ongoing demands on laboratory performance require optimization of processes. An obvious way to achieve this is to reduce manual labor in favor of automated methods. We describe the validation of an automated quantitative urine human chorionic gonadotropin (hCG) analysis on the Roche Modular E170 analyzer to replace the manual qualitative pregnancy test in urine. At urine hCG concentrations of 476, 45 and 11 U/L, we found inter-assay variation of 4.3%, 4.3% and 6.8% and average intra-assay variation of 3.0%, 2.6% and 3.0%, respectively. The analytical detection limit was 0.7 U/L. We did not detect any loss (due to degradation or adsorption) during a storage period of 5 days at 4 degrees C or at -20 degrees C. Recoveries of hCG in urine of a pregnant woman diluted with urine of a pre-menopausal non-pregnant woman (concentration range between 6 and 800 mU/L) were between 93% and 112% (y=0.997x-3.843, r 2 =0.999). Diluting a serum sample (hCG 42,000 U/L) with urine (negative for hCG) up to 8000 fold yielded a completely linear hCG response, indicating that the assay was not affected by the urine matrix. In a correlation study with 60 urine samples (of which 10 were of male origin), we did not find any discrepancies between results for the manual pregnancy test and the hCG test on the Roche Modular E170 (using a cutoff value of 50 U/L). PMID- 15653446 TI - Interference of dextran in biuret-type assays of serum proteins. AB - Dextran interference in biuret-type assays of total serum proteins was investigated in a Belgian National External Quality Assurance Survey with 256 participants. In vitro supplementation of therapeutic (10% Gentran 70) dextran concentrations showed a broadly varying (from 0 to 20%) negative interference. The analytical interference was found to depend on both the sodium hydroxide and tartrate concentrations in the reagent formulation. The dry chemistry biuret method was not affected by the dextran interference. In a number of cases, the effects observed may be of clinical importance. Both clinicians and laboratory staff should be aware of the persistence of this analytical problem. PMID- 15653447 TI - Evaluation of immunoassays for the measurement of insulin-like growth factor-I and procollagen type III peptide, indirect biomarkers of recombinant human growth hormone misuse in sport. AB - Insulin-like growth factor-I (IGF-I) and procollagen type III peptide (P-III-P) have been proposed as indirect biomarkers for the detection of the misuse of recombinant human growth hormone in sport. An extended intra- and inter laboratory validation of commercially available immunoassays was carried out. For total IGF-I, two radioimmunoassay (RIA) kits (IGF-I/RIA1, Nichols Institute Diagnostics and IGF-I/RIA2, Mediagnost) and one enzyme-linked immunosorbent assay (ELISA) (R&D) were evaluated. For P-III-P, two RIA kits (P-III-P/RIA3, Cis bioInternational and P-III-P/RIA4, Orion Diagnostica) were studied. The intra laboratory precision and accuracy values for all IGF-I assays were better than 15%. The IGF-I/ELISA showed the lowest limit of quantification (LOQ) and its calibration curve covered the range of concentrations found in human serum samples. Higher agreement between laboratory results was obtained for IGF-I/ELISA and IGF-I/RIA1. Low inter-technique correlation was obtained for the three assays; the only comparable results were obtained between IGF-I/ELISA and IGF I/RIA1. For P-III-P, intra-laboratory precision and accuracy values better than 15% were obtained for both assays in almost all cases. The calibration curve for P-III-P/RIA4 covered the range of concentrations of serum samples, while 30% of the values for P-III-P/RIA3 were below the calibration sample with the lowest concentration. Inter-laboratory correlation was also higher for P-III-P/RIA4. In summary, ELISA and RIA4 were the most suitable assays for measurement of IGF-I and P-III-P, respectively, in serum samples. However, the validation studies carried out show the need for harmonization of immunoassay parameters to improve the reproducibility and comparability of results between different laboratories and in different studies. PMID- 15653448 TI - Evaluation of a new automated electrochemiluminescent sex hormone-binding globulin (SHBG) immunoassay. AB - Serum sex hormone-binding globulin (SHBG) regulates the cellular bioavailability of SHBG-bound steroid hormones. Since variations in SHBG levels may affect the concentration of free, i.e., biologically active testosterone in serum, SHBG levels are commonly measured as a supplement to total testosterone determination. The recently developed electrochemiluminescence Elecsys SHBG immunoassay was evaluated analytically on a Modular E170 (Roche Diagnostics, Mannheim, Germany) immunoanalyzer. Major differences in SHBG concentrations have been described among the commercially available methods; we therefore compared the new method with an established SHBG immunoradiometric assay (IRMA) in 99 routine serum samples. To provide reference values to clinicians, SHBG concentration was measured by Elecsys in 304 serum samples from healthy volunteers and several relevant clinical subgroups. The within-run and total imprecision coefficients of variation were 0.982. Accuracy and precision, evaluated by recovery and replicate assays, are also very acceptable. This method is suitable as a fast, cost effective alternative screening method to estimate milk fat content in small samples without prior lipid extraction. PMID- 15653513 TI - Analysis of phospho- and sphingolipids in dairy products by a new HPLC method. AB - Dairy phospho- and sphingolipids are gaining interest due to their nutritional and technological properties. A new HPLC method, using an evaporative laser light scattering detector, was developed, which enabled excellent separation of glucosylceramide, lactosylceramide, phosphatidic acid, phosphatidylethanolamine, phosphatidylinositol, phosphatidylserine, phosphatidylcholine, sphingomyelin, and lysophosphatidylcholine in less than 21 min, including the regeneration of the column. No loss of column performance was observed after 1500 runs because an acid buffer was used. The output signal of the evaporative laser light scattering detector was highly dependent of the flow of the carrier gas and the temperature of the nebulizer, and was maximized by means of a response surface experimental design. Finally, raw milk, cream, butter, buttermilk, Cheddar whey, quarg, and Cheddar cheese were analyzed for their polar lipid content. The absolute values varied substantially (0.018 to 0.181 g/100 g of product). Significant differences were found in the relative content of each polar lipid class among the analyzed products. PMID- 15653514 TI - Flavor and stability of pasteurized milk with elevated levels of conjugated linoleic acid and vaccenic acid. AB - The objectives of this study were to determine if flavor differences between 2% fat pasteurized milks with and without naturally enhanced vaccenic acid (VA) and cis-9, trans-11 conjugated linoleic acids (CLA) levels could be detected over the commercial shelf life of the product and to determine if milk with elevated VA and cis-9, trans-11 CLA levels was more susceptible to development of light induced oxidative flavor defects. Cows were fed a control diet or the same ration supplemented with 2% soybean oil and 1% fish oil (CLA diet). The milk, standardized to 2% fat, was pasteurized, homogenized, and stored in plastic containers at 4 degrees C. Oxidation was induced by exposing half of the containers to light. Testing was conducted at 1, 7, and 14 d postpasteurization. Average cis-9, trans-11 CLA content of the milks from the control and CLA diet groups was 0.52 and 4.74 g/100 g of fatty acids, respectively (8-fold increase). Average VA content of the milk from the control and CLA diet groups was 1.43 and 12.06 g/100 g of fatty acids, respectively (7.5-fold increase). Together, VA plus CLA represented almost 17% of the total milk fatty acids. There was no effect of light exposure on fatty acid composition initially or over the 14-d storage period. Although VA, cis-9, trans-11 CLA, and degree of unsaturation were significantly elevated in the milk from the CLA diet group, untrained panelists were unable to detect flavor differences initially or over time in 15 of 16 triangle test evaluations. Similarly, sensory results indicated no difference in susceptibility to the development of oxidized off-flavors between the milk from the control and CLA diet groups, even when oxidation was induced by light exposure. PMID- 15653515 TI - Protective influence of several packaging materials on light oxidation of milk. AB - Light-induced degradation reactions in milk create a serious problem for the dairy industry because of the development of off-flavors, the decrease in nutritional quality, and the severity and speed by which these phenomena develop. Packaging materials are essential to avoid this particular deterioration of milk. Therefore, efforts are being made to design protective polyethylene terephthalate (PET) packages. In the present study, a number of PET bottles were compared for their ability to avoid photo-oxidation in UHT semi-skimmed milk. The milk was packed in 3 types of PET bottles: one transparent bottle provided with an active oxygen-binding inner layer, one bottle with perfect light barrier, and one transparent bottle provided with a UV-absorbing additive. During 2 storage experiments, running parallel to each other for 2 mo, chemical milk quality parameters such as fat oxidation, vitamin and protein degradation, oxygen consumption, and color change were monitored. A trained taste panel compared the sensory quality of the illuminated milk stored in these bottles, with milk perfectly protected against light and oxygen. In the first study, milk was continuously illuminated at room temperature. A comparison was made for milk under storage conditions that simulated those expected during display in retail and supermarkets. The results of the 2 shelf-life studies showed that an adequate light barrier was apparently sufficient to avoid the light-induced oxidation of milk during extended storage. Oxygen barriers, on the other hand, did not provide a significant protection, nor did bottles with UV filter. If wavelengths detrimental to riboflavin were not completely excluded by the packaging material, incoming light could still give rise to photo degradation of milk. Accordingly, riboflavin and vitamin A were gradually degraded, milk fat was photo-oxidized, oxygen dissolved in the milk was consumed, and the sensorial quality decreased significantly. PMID- 15653516 TI - Thermal and structural behavior of anhydrous milk fat. 3. Influence of cooling rate. AB - The crystallization behavior of anhydrous milk fat has been examined with a new instrument coupling time-resolved synchrotron x-ray diffraction as a function of temperature (XRDT) at both small and wide angles and high-sensitivity differential scanning calorimetry. Crystallizations were monitored at cooling rates of 3 and 1 degrees C/ min from 60 to -10 degrees C to determine the triacylglycerol organizations formed. Simultaneous thermal analysis permitted the correlation of the formation/melting of the different crystalline species monitored by XRDT to the thermal events recorded by differential scanning calorimetry. At intermediate cooling rates, milk fat triacylglycerols sequentially crystallize in 3 different lamellar structures with double-chain length of 46 and 38.5 A and a triple-chain length of 72 A stackings of alpha type, which are correlated to 2 exothermic peaks at 17.2 and 13.7 degrees C, respectively. A time-dependent slow sub-alpha <--> alpha reversible transition is observed at -10 degrees C. Subsequent heating at 2 degrees C/min has shown numerous structural rearrangements of the alpha varieties into a single beta' form before final melting. This polymorphic evolution on heating, as well as the final melting point observed (approximately 39 degrees C), confirmed that cooling at 3 degrees C/min leads to the formation of crystalline varieties that are not at equilibrium. An overall comparison of the thermal and structural properties of the crystalline species formed as a function of the cooling rate and stabilization time is presented. The influence on crystal size of the cooling rates applied in situ using temperature-controlled polarized microscopy is also determined for comparison. PMID- 15653517 TI - Clinical effects of Lactobacillus acidophilus strain L-92 on perennial allergic rhinitis: a double-blind, placebo-controlled study. AB - Studies in animals have suggested that lactic acid bacteria alleviate allergic diseases, however, little information is available on their clinical effect on allergy in humans. Thus, we examined the efficacy of orally administered Lactobacillus acidophilus strain L-92 (L-92) on perennial allergic rhinitis. In a randomized, double-blind, placebo-controlled clinical trial, 49 patients with perennial allergic rhinitis were randomized to receive either 100 mL of heat treated fermented milk containing L-92 (n = 25) or acidified milk without lactic acid bacteria (placebo; n = 24) for 8 wk. The severity of symptoms was evaluated based on the changes in the scores of clinical symptoms. Oral administration of milk fermented with L-92 resulted in a statistically significant improvement of nasal symptom-medication scores. Ocular symptom-medication scores of patients in the L-92 intervention group tended to improve compared with those in the placebo group. In addition, clear decreases of the scores of swelling and color of the nasal mucosa were observed in the L-92 intervention group at 6 and 8 wk after the start of ingestion of fermented milk. There were no significant differences in serum antihouse dust mite immunoglobulin E levels nor in T helper type 1/T helper type 2 ratio between the 2 groups. These results suggest that oral administration of L-92 can alleviate the symptoms of perennial allergic rhinitis, however, statistically significant changes were not shown in blood parameters. PMID- 15653518 TI - Effects of adjuvants on safety and efficacy of an Escherichia coli J5 bacterin. AB - The effects of using a water-soluble adjuvant or an emulsified oil-based adjuvant on the safety, antibody titer, and clinical responses of an Escherichia coli J5 bacterin were tested in an experimental infection trial. Fifty-one cows were assigned to 17 blocks of 3. Two cows within each block of 3 were vaccinated with a commercially prepared E. coli J5 bacterin containing either a water-soluble adjuvant or the same bacterin preparation emulsified in oil. One cow in each block was an unvaccinated control. Cows were immunized at drying off and 42 d later. The right or left front mammary quarter of each experimental cow was challenged by intramammary infusion of E. coli 727 between 14 and 35 DIM. Areas of inflammation at the primary injection site were greater 1, 2, and 3 d following primary vaccination for bacterin containing oil-in-water adjuvant compared with bacterin containing water-soluble adjuvant. Whey anti-E. coli J5 IgG titers were higher at calving for cows vaccinated with bacterin containing oil-in-water adjuvant than for cows either vaccinated with bacterin containing water-soluble adjuvant or unvaccinated controls. Serum x-E. coli J5 IgG titers were higher at calving for vaccinated cows than for unvaccinated controls. Peak bacterial counts in milk from challenged quarters were greater for unvaccinated controls than for cows vaccinated with bacterin containing water-in-oil adjuvant. Bacterial counts in milk from challenged quarters and clinical score both were greater in unvaccinated controls than cows vaccinated with bacterin containing water-in-oil adjuvant between 12 and 24 h postchallenge. Clinical responses were similar between unvaccinated controls and cows vaccinated with bacterin containing water-soluble adjuvant. PMID- 15653519 TI - Effects of bovine somatotropin on uterine genes related to the prostaglandin cascade in lactating dairy cows. AB - Multiparous Holstein cows, averaging 80 d in milk, were used to examine the effect of exogenous bovine somatotropin (bST) on uterine expression of estrogen receptor alpha (ERalpha), prostaglandin endoperoxide synthase-2 (PGHS-2), and peroxisome proliferator-activated receptor delta (PPARdelta). About 12 h before expected ovulation in a synchronization protocol, cows were assigned to receive bST (500 mg, n = 11) or serve as untreated controls (n = 10). Cows that ovulated (n = 9 bST, 8 control) were divided within treatment to be killed on d 3 or 7 postovulation. Samples of intercaruncular endometrial tissue from uterine horns ipsilateral to the corpus luteum were collected and stored at -80 degrees C for subsequent mRNA analyses. Endometrial concentrations of ERalpha and PGHS-2 mRNA transcripts were greater on d 7 than on d 3 of the estrous cycle, but did not differ between treatments. Compared with untreated cows, short-term bST treatment decreased PGHS-2 protein expression at d 7 of the estrous cycle. Concentration of PPARdelta mRNA transcript in the uterus decreased between d 3 and 7 of the estrous cycle and was negatively correlated with ERalpha and PGHS-2 mRNA concentrations. Short-term administration of bST to lactating dairy cows had minimal effects on uterine genes encoding ERalpha, PGHS-2, and PPARdelta at d 3 and 7 of the estrous cycle but there may be an inverse relationship between PPARdelta and uterine expression of ERalpha and PGHS-2 genes. PMID- 15653520 TI - Interlaboratory proficiency testing as a tool for improving performance in laboratories diagnosing bovine mastitis. AB - The National Veterinary and Food Research Institute (Finland) and the Veterinary Laboratories Agency of the Quality Assurance Unit, Department for Environment, Food and Rural Affairs, United Kingdom (previously the Ministry of Agriculture, Fisheries and Food) organized a proficiency testing program for laboratories analyzing veterinary mastitis samples. Three test samples with lyophilized strains of common aerobic bacteria were sent to the participating laboratories 7 times between 2000 and 2003. The participants returned 98% of the requested data. The overall performance of the laboratories varied from 63 to 93% in different testing rounds. All laboratories diagnosed Staphylococcus aureus and Escherichia coli correctly at every round. Improvement in diagnosing individual bacteria was observed for Staphylococcus epidermidis, Streptococcus dysgalactiae, Enterococcus spp. and Klebsiella spp. The overall performance of the laboratories improved with increased participation. The educational role of the program was important. Laboratories working in the veterinary field should implement a documented quality system covering all functions of the laboratory, as well as a planned quality assurance system. PMID- 15653521 TI - Association between somatic cell count in early lactation and culling of dairy heifers using cox frailty models. AB - The association between somatic cell count (SCC) of dairy heifers in early lactation [SCCel; measured between 5 and 14 d in milk (DIM)] and the culling hazard during the first lactation was studied using Cox frailty models. Udder health problems were the culling reason for 10% of the culled heifers in this study. For each unit increase in the log-transformed SCCel (LnSCCel), the culling hazard increased by 11% [Hazard ratio (HR) = 1.11]. The strength of the association depended on 5 factors. Firstly, the association was stronger when SCCel was recorded after 10 DIM than at an earlier DIM. Secondly, the association was stronger if only culling events for udder disorders were considered (HR = 1.32) instead of all culling events (HR = 1.11). Furthermore, for each unit increase of test-day LnSCC after 14 DIM, modeled as a time-varying covariate, the culling hazard in the first lactation increased by 26% (HR = 1.26). Including LnSCC in the model already containing LnSCCel, reduced the estimate of LnSCCel slightly. Fourth, a higher test-day milk yield, modeled as a time-varying covariate, protected against culling and reduced the magnitude of the effect of LnSCCel as well when taken into account. Finally, the association between LnSCCel and culling was still present, although smaller, in the group of heifers with a second test-day SCC /=300,000 cells/mL, the cow was enrolled and randomly allocated to a treatment group. Enrolled cows were monitored for clinical mastitis and other disease for 4 wk after treatment initiation. At 3 and 4 wk after treatment initiation, milk samples were taken from each enrolled quarter to determine the SCC and conduct a bacteriological culture. Bacteriological culture results were interpreted such that quarters where the same bacterial species was cultured before treatment and found in at least 1 of the 2 posttreatment samples were considered a failure. The analysis of SCC used a mixed linear model (SAS proc mixed) and the analysis of bacteriological cure used a mixed logistic model (SAS glimmix macro). Bacteriological cure rate was significantly higher for lower parity, lower number of colonies in the pretreatment culture, longer treatment duration, and for streptococci compared with Staphylococcus aureus. However, treatment regimen affected bacteriological cure differently in major than in minor pathogens and there was a significant interaction of treatment regimen with stage of lactation. Posttreatment SCC was significantly higher with increasing parity, in rear quarters, and with shorter duration of treatment. In the group of second and third parity animals, post treatment SCC was more reduced in front quarters than in rear quarters. Also, the difference in posttreatment SCC between younger and older cows increased with higher pretreatment SCC. In conclusion, when predicting bacteriological cure following treatment of subclinical mastitis during lactation both treatment regimen and other risk factors need to be considered. The other risk factors may vary with treatment regimen. Posttreatment SCC was associated with treatment regimen, other risk factors, and interactions among the other risk factors; but these other risk factors did not vary significantly with treatment regimen. PMID- 15653528 TI - Frictional forces required for unrestrained locomotion in dairy cattle. AB - Most free-stall housing systems in the Netherlands are equipped with slatted or solid concrete floors with manure scrapers. A slipping incident occurs when the required coefficient of friction (RCOF) exceeds the coefficient of friction (COF) at the claw-floor interface. An experiment was conducted to measure ground reaction forces (GRF) of dairy cows (n = 9) performing various locomotory behaviors on a nonslippery rubber-covered concrete floor. The RCOF was determined as the ratio of the horizontal and vertical components of the GRF. It was shown that during straight walking and walking-a-curve, the RCOF reached values up to the COF, whereas for sudden stop-and-start responses, the RCOF reached values beyond the maximum COF that concrete floors can provide. Our results indicate that concrete floors do not provide enough friction to allow natural locomotory behavior and suggest that tractional properties of floors should be main design criteria in the development of better flooring surfaces for cattle. PMID- 15653529 TI - Time of feed delivery affects the feeding and lying patterns of dairy cows. AB - The objective of this experiment was to determine whether it is the return from milking or delivery of fresh feed that has the greater effect on the daily patterns of feeding and lying behavior of dairy cattle. Forty-eight lactating Holstein cows were subjected to each of 2 treatments in a 2 x 2 cross-over design replicated over time. The treatments were 1) milking and feed delivery times coinciding and 2) feed delivery 6 h after milking. Cows were milked twice daily at 0500 and 1700 h. An electronic monitoring system was used to measure the time spent at the feed alley. Time-lapse video was used to quantify the lying time and incidence of aggressive displacements of the cows at the feed alley. Cows increased their total daily feeding time by 12.5% when fed 6 h after milking. This change was driven by an 82% increase in feeding time during the first hour immediately following the delivery of fresh feed and a 26% decrease in feeding time during the first hour after milking. The delivery of feed 6 h after milking did not change the daily lying time of the cows, but did decrease the latency to lie down after milking by 20 min. The reduction in feeding time after milking and decreased latency to lie down resulted in a tendency for less aggressive interactions at the feed alley after the cows returned from milking. These results indicate that the delivery of fresh feed has a greater impact on stimulating feeding behavior than does the return from milking and that changes in feeding management can affect both the feeding and lying behavior of dairy cows. PMID- 15653530 TI - Dry matter intake is decreased more by abomasal infusion of unsaturated free fatty acids than by unsaturated triglycerides. AB - Previous experiments from our group have demonstrated that abomasal infusion of unsaturated free fatty acids (FFA) markedly decreases dry matter intake (DMI) in dairy cows. In contrast, experiments from other groups have noted smaller decreases in DMI when unsaturated triglycerides (TG) were infused postruminally. Our hypothesis was that unsaturated FFA would be more potent inhibitors of DMI than an equivalent amount of unsaturated TG. Four Holstein cows in late lactation were used in a single reversal design. Cows were fed a total mixed ration containing (DM basis) 23% alfalfa silage, 23% corn silage, 40.3% ground shelled corn, and 10.5% soybean meal. Two cows received soy FFA (UFA; 0, 200, 400, 600 g/d) and 2 received soy oil (TG) in the same amounts; cows then were switched to the other lipid source. Cows were abomasally infused with each amount for 5-d periods. The daily amount of lipid was pulse-dosed in 4 equal portions at 0600, 1000, 1700, and 2200 h; no emulsifiers were used and there was no sign of digestive disturbance. Both lipid sources linearly decreased DMI, with a significant interaction between lipid source and amount. Slope-ratio analysis indicated that UFA were about 2 times more potent in decreasing DMI than were TG. Decreased DMI led to decreased milk production. Milk fat content was increased linearly by lipid infusion. Milk fat yield decreased markedly for UFA infusion but was relatively unaffected by infusion of TG. Contents of short- and medium chain fatty acids in milk fat decreased as the amount of either infusate increased. Contents of C(18:2) and C(18:3) in milk fat were increased linearly by abomasal infusion of either fat source; cis-9 C(18:1) was unaffected. Transfer of infused C(18:2) to milk fat was 35.6, 42.5, and 27.8% for 200, 400, and 600 g/d of UFA, and 34.3, 39.6, and 34.0% for respective amounts of TG. Glucagon-like peptide-1 (7-36) amide (GLP-1) concentration in plasma significantly increased as DMI decreased with increasing infusion amount of UFA or TG. Plasma concentration of cholecystokinin-octapeptide (CCK-8) was unaffected by lipid infusion. These results indicate that unsaturated FFA reaching the duodenum are more potent inhibitors of DMI than are unsaturated TG; the effect may be at least partially mediated by GLP-1. PMID- 15653531 TI - Monensin for lactating dairy cows grazing mixed-alfalfa pasture and supplemented with partial mixed ration. AB - The effect of monensin on milk production was evaluated in 58 lactating Holstein cows (48 multiparous; 10 primiparous) grazing a mixed-alfalfa pasture and supplemented with a partial mixed ration in a completely randomized design with repeated measurements. Cows were paired by calving date, lactation number, previous lactation milk production, body weight, and body condition score and were assigned to one of 2 treatments: control or monensin. Cows on the monensin treatment received 2 monensin controlled-release capsules (335 mg/d for 90 d), one 30 d before the expecting calving date and the other 60 d after calving. Short-term (0 to 150 d in milk) and long-term (305-d adjusted lactation) effects of monensin were evaluated. Pasture (measured by difference between pre- and postgrazing pasture mass), supplements, and total dry matter intake did not differ between treatments and averaged 8.7, 14.1, and 22.9 kg/d, respectively. In the short-term, monensin increased milk production (27.7 vs. 26.6 kg/d) and milk protein yield (0.890 vs. 0.860 kg/d); milk fat yield was not affected (0.959 kg/d). Monensin decreased milk fat content (3.51 vs. 3.60%) with no changes in milk protein content (3.25%). In the long term, milk production and milk protein yield were also increased by monensin: 214 and 7 kg, respectively. Monensin reduced the loss of body condition score and increased percentage of pregnancy at first service (44.8 vs. 20.7%). Monensin improves production and reproduction performance of dairy cows grazing a mixed-alfalfa pasture and supplemented with a partial mixed ration. PMID- 15653532 TI - Plasma lipid concentrations in preruminant calves fed whole milk with whey protein isolate. AB - The objective was to investigate the acute effects of retinol acetate added to whey protein isolate (WPI) on postprandial changes in plasma retinol (experiment 1) and the acute effects of milk fat added to WPI on triglyceride (TG), chylomicrons and very low density lipoprotein (VLDL), and fatty acid concentrations (experiment 2) in suckling calves at 1 and 6 wk of age. In experiment 1, 16 Holstein male calves were alloted to 2 equal groups. On the days of measurement, the calves were fed at 0900 h whole milk [4% of body weight (BW)] mixed with vitamin A acetate (500,000 IU) with or without WPI (0.04% of BW). At 1 wk of age, significantly higher postfeeding concentrations of plasma retinol were observed in the calves fed milk with WPI. At 6 wk of age, no differences in the plasma retinol concentrations were observed between 2 groups. On the days of measurement in experiment 2, 16 male calves were fed at 0900 h whole milk (4% of BW) with added milk fat prepared by centrifugation from whole milk (2% of BW) with or without WPI (0.04% of BW). The milk supplemented with fat was prepared on the day before the measurement. At 1 wk of age, significant higher postfeeding concentrations of plasma TG concentrations were obtained in the calves fed WPI than in the control calves, immediately after the meal or from 7 h later onward. Plasma chylomicrons and VLDL concentrations at 1 wk of age were significantly higher in the WPI-fed group than in the control group at 8 h postfeeding. In the calves with the WPI diet, plasma concentrations of myristic, palmitic, stearic, oleic, and linoleic acids at 1 wk of age were significantly higher than those in the control calves at 8 h after feeding. However, chylomicrons and VLDL, and fatty acid concentrations did not differ between the 2 groups after feeding at 6 wk of age. Results indicate that WPI increases plasma lipid concentration of preruminant calves only at 1 wk of age. These data are interpreted to indicate that WPI enhances mainly lipid uptake in the intestines of neonatal calves. PMID- 15653533 TI - Effects of dietary supplements of folic acid and rumen-protected methionine on lactational performance and folate metabolism of dairy cows. AB - The present experiment was undertaken to determine the interactions between dietary supplements of folic acid and rumen-protected methionine on lactational performance and on indicators of folate metabolism during one lactation. Fifty four multiparous Holstein cows were assigned to 9 blocks of 6 cows each according to their previous milk production. Within each block, 3 cows were fed a diet calculated to supply methionine as 1.75% metabolizable protein, equivalent to 70% of methionine requirement, whereas the 3 other cows were fed the same diet supplemented with 18 g of a rumen-protected methionine supplement. Within each diet, the cows received 0, 3, or 6 mg/d of folic acid per kg of body weight. Rumen-protected methionine increased milk total solid concentration but not yield. Supplementary folic acid increased crude protein and casein concentrations in milk of cows fed no supplementary methionine and the effect increased as lactation progressed; it also decreased milk lactose concentration. Folic acid supplements had the opposite effects on milk crude protein, casein, and lactose concentrations in cows fed rumen-protected methionine. Milk and milk component yields and dry matter intake were unchanged. Folic acid supplementation increased serum folates and this response was greater at 8 wk of lactation. It decreased serum cysteine in cows fed rumen-protected methionine, whereas it had no effect in cows fed no supplementary methionine. The highest serum concentrations of cysteine but the lowest of vitamin B(12) were observed at 8 wk of lactation. Serum clearance of folic acid following an i.v. injection of folic acid was slower at 8 wk of lactation. During this period, the high concentrations of serum folates and cysteine, the low serum concentrations of vitamin B(12) and methionine, and the slow serum clearance of folates strongly suggest that the vitamin B(12) supply was inadequate and interfered with folate use. It could explain the limited lactational response to supplementary folic acid observed in the present experiment. PMID- 15653534 TI - Effects of intramuscular injections of vitamin B12 on lactation performance of dairy cows fed dietary supplements of folic acid and rumen-protected methionine. AB - The experiment was undertaken to determine the effects of i.m. injections of vitamin B(12) on lactational performance of primiparous dairy cows fed dietary supplements of folic acid and rumen-protected methionine from 4 to 18 wk of lactation. Fourteen primiparous Holstein cows were assigned to 7 blocks of 2 cows each, according to milk production during the third week of lactation. All cows were fed a basal diet supplemented daily with rumen-protected methionine (18 g of supplement, to bring the estimated supply of methionine to 2.2% of metabolizable protein) plus folic acid (4 mg per kg of BW). Within each block, the cows received a weekly i.m. injection (2 mL) of saline or 10 mg of vitamin B(12). Milk production was recorded daily. Milk and blood were sampled every 2 wk. Supplementary vitamin B(12) increased energy-corrected milk from 25.8 to 29.0 (SE 1.6) kg/d, as well as milk yields of solids [3.52 to 3.90 (SE 0.22) kg/d], fat [0.87 to 1.01 (SE 0.06) kg/d], and lactose [1.48 to 1.64 (SE 0.11) kg/d]. Supplementation also increased concentrations and amounts of vitamin B(12) secreted in milk but had no significant effect on dry matter intake and concentrations and amounts of folates in milk. Packed cell volume, blood hemoglobin, and serum vitamin B(12) were increased by supplementary vitamin B(12), whereas serum methylmalonic acid was decreased. Serum concentrations of sulfur amino acids were unchanged by treatment. These findings support the hypothesis that, in early lactation, supply of vitamin B(12) was not optimal and limited the lactation performance of the cows. PMID- 15653535 TI - Influence of precalving feed allowance on periparturient metabolic and hormonal responses and milk production in grazing dairy cows. AB - Fifty-two multiparous dairy cows were allocated to 4 treatments consuming 5.4, 8.2, 10.0, or 11.0 kg/d of pasture dry matter per cow for 27 +/- 9.6 d precalving. This equated to 1.3, 1.9, 2.4, and 2.6% of body weight (BW; not including the conceptus weight). Following calving, all cows were fed ad libitum on pasture. Blood was sampled 17 d precalving, on day of calving, and on d 1, 2, 3, 4, 7, 14, 28, and 35 postcalving. Results suggest that the near-term grazing dairy cow requires 1.05 MJ of ME/kg of BW(0.75) and that previous estimates of energy requirements were underestimated. Precalving plasma concentrations of glucose, insulin-like growth factor-1, and leptin increased quadratically with increasing pasture intake. This was associated with precalving plasma concentrations of growth hormone that declined linearly, and concentrations of nonesterified fatty acids and beta-hydroxybutyrate that declined quadratically with increasing dry matter intake (DMI). Postcalving plasma concentrations of these metabolites showed no lasting effect of precalving feeding. The effect of precalving nutrition on milk production was small, and other than milk fat, was confined to wk 1 postcalving. Milk fat yield increased with increasing precalving DMI and calving body condition score until wk 3 post-calving, after which treatment effects were not evident. These results indicate that the level of feeding in grazing dairy cows during the last month before calving has only small effects on cow metabolic and hormonal status, and on milk production in the first 5 wk of lactation. PMID- 15653536 TI - Ruminal fermentation and amino acid flow in Holstein steers fed whole cottonseed with elevated concentrations of free fatty acids in the oil. AB - The influence of feeding whole cottonseed (WCS) containing elevated concentrations of free fatty acids (FFA) in the oil on ruminal fermentation and amino acid (AA) flow to the abomasum was evaluated in a 4 x 4 Latin square trial. Four ruminally and abomasally cannulated Holstein steers were fed diets containing 12.5% of dry matter as WCS with concentrations of 8.0, 11.3, 14.7, or 18.0% FFA in the oil. Intake, ruminal digestibility, and flow to the abomasum of dry matter, organic matter, and acid detergent fiber were not affected by FFA level of WCS. Intake of neutral detergent fiber and total kilograms of neutral detergent fiber digested in the rumen were similar for all treatments. Ruminal neutral detergent fiber digestibility was lower for 8 and 14.7% FFA, resulting in a cubic effect on flow to the abomasum. Ruminal pH, molar proportions of isobutyrate, and total branched-chain volatile fatty acids (VFA) decreased linearly, whereas molar proportions of acetate and acetate:propionate ratio increased linearly as FFA in WCS increased. Total VFA were lower, and molar proportions of propionate were higher, for 8 and 14.7% FFA, resulting in a cubic effect. Intake of N, total N flow, and nonmicrobial N flow to the abomasum were similar among treatments. Flow of microbial N was lower for the 11.3% FFA treatment, resulting in a quadratic response. Only nonsignificant differences were observed in AA flow to the abomasum. Results of this trial indicate that WCS with FFA up to 18% may result in small changes in rumen fermentation. PMID- 15653537 TI - Effect of different dietary geometric mean particle length and particle size distribution of oat silage on feeding behavior and productive performance of dairy cattle. AB - Twenty lactating Holstein cows (5 primiparous and 15 multiparous) were used in a 5 x 5 Latin Square design, with 5 treatments and 3 periods of 21 d each. Diets contained 25% corn silage, 25% oat silage, and 50% concentrate (dry matter basis). The 5 treatments tested in the experiment were long oat silage (LOS), medium oat silage (MOS), fine from long oat silage (FLOS), fine from medium oat silage (FMOS), and half LOS plus half FLOS (LFLOS). The geometric mean particle length (GMPL) of the diets was 6.68, 5.19, 4.46, 4.35, and 5.39 mm for LOS, MOS, FLOS, FMOS, and LFLOS, respectively. The LFLOS was designed to provide dietary GMPL similar to MOS, but with a more bimodal particle size distribution (PSD). Linear and quadratic effects of GMPL were tested, based on the mean GMPL of the feed actually consumed (cGMPL). Contrasts were used to test for the effect of different PSD (MOS vs. LFLOS) and to test for differences between FMOS and FLOS, which would indicate unequal fermentations in the MOS and LOS silos. No differences were detected between FMOS and FLOS in most of the variables measured. Increasing cGMPL linearly decreased dry matter intake, milk production, and milk protein percentage and yield without affecting milk fat percentage, milk fat yield, ruminal pH, and ruminal volatile fatty acid concentration. Although cows fed diets with increasing cGMPL spent more time eating and chewing per day and per kilogram of dry matter intake, there was no effect of cGMPL on rumen pH. Feeding medium oat silage increased milk fat percentage and yield compared with feeding a mixture of long and fine oat silage. PMID- 15653538 TI - Effects of supplemental hay and corn silage versus full-time grazing on ruminal pH and chewing activity of dairy cows. AB - Grazing young, highly digestible swards with and without supplemental hay or corn silage (5.5 kg of DM/d) offered overnight was tested for its effects on ruminal pH and chewing activity. A double 3 x 3 Latin square arrangement with 6 rumen cannulated Brown Swiss cows (29 kg/d of milk) was applied. Herbage intake was quantified by controlled-release alkane capsules. Chewing activity was determined using an automatic microcomputer-based system for digital recording of the jaw movements. Except during milking, ruminal pH was measured continuously over 7 d by applying a device consisting of an indwelling pH electrode and a data recording unit integrated in the cannula's cover. The grazing system had no significant effect on body weight, milk yield or composition (except milk urea), or total DM intake (13.5, 13.8, and 15.7 kg/d with full-time grazing, hay, and corn silage supplementation). No differences occurred for ruminating time per day and time per kilogram of DM intake. Full-time grazing cows spent more time eating per day (+26%) and time per kilogram of DM intake (+31%) than the other cows. Ruminal pH and time with pH <5.8 at night did not differ. Throughout the day, hay supplemented cows had a significantly lower pH (-0.23) than full-time grazing cows, and the period of pH <5.8 was longer compared with corn-silage fed cows (77 vs. 11 min). Nocturnal supplement feeding gave no advantage over full-time grazing, and supplemental hay led to lower daytime pH. PMID- 15653539 TI - Relationship among trans and conjugated fatty acids and bovine milk fat yield due to dietary concentrate and linseed oil. AB - Effects on fatty acid profiles and milk fat yield due to dietary concentrate and supplemental 18:3n-3 were evaluated in 4 lactating Holstein cows fed a low- (35:65 concentrate:forage; L) or high- (65:35; H) concentrate diet without (LC, HC) added oil or with linseed oil (LCO, HCO) at 3% of DM. A 4 x 4 Latin square with four 4-wk periods was used. Milk yield and dry matter intake averaged 26.7 and 20.2 kg/d, respectively, across treatments. Plasma acetate and beta hydroxybutyrate decreased, whereas glucose, nonesterified fatty acids, and leptin increased with high-concentrate diets. Milk fat percentage was lower in cows fed high-concentrate diets (2.31 vs. 3.38), resulting in decreases in yield of 11 (HC) and 42% (HCO). Reduced yields of 8:0-16:0 and cis9-18:1 fatty acids accounted for 69 and 17%, respectively, of the decrease in milk fat yield with HC vs. LC (-90 g/d), and for 26 and 33%, respectively, of the decrease with HCO vs. LCO (-400 g/d). Total trans-18:1 yield increased by 25 (HCO) and 59 (LCO) g/d with oil addition. Trans10-18:1 yield was 5-fold greater with high-concentrate diets. Trans11-18:1 increased by 13 (HCO) and 19 (LCO) g/d with oil addition. Trans13+14-18:1 yield increased by 9 (HCO) and 18 (LCO) g/d with linseed oil. Yield of total conjugated linoleic acids (CLA) in milk averaged 6 g/d with LC or HC compared with 14 g/d with LCO or HCO. Cis9,trans11-CLA yield was not affected by concentrate level but increased by 147% in response to oil. Feeding oil increased yields of trans11,cis13-, trans11,trans13-, and trans,trans-CLA, primarily with LCO. Trans10,cis12-CLA yield (average of 0.08 g/d) was not affected by treatments. Yield of trans11,cis15-18:2 was 1 g/d in cows fed LC or HC and 10 g/d with LCO or HCO. Yields of cis9,trans11-18:2, cis9,trans12-18:2, and cis9,trans13-18:2 were positively correlated (r = 0.74 to 0.94) with yields of trans11-18:1, trans12-18:1, and trans13+14-18:1, respectively. Plasma concentrations of biohydrogenation intermediates with concentrate or linseed oil level followed similar changes as those in milk fat. Milk fat depression was observed when HC induced an increase in trans10-18:1 yield. A correlation of 0.84 across 31 comparisons from 13 published studies, including the present one, was found among the increase in percentage of trans10-18:1 in milk fat and decreased milk fat yield. We observed, however, more drastic milk fat depression when HCO increased yields of total trans-18:1, trans11,cis15-18:2, trans isomers of 18:3, and reduced yields of 18:0 plus cis9-18:1. PMID- 15653540 TI - Bypassing the rumen in dairy ewes: the reticular groove reflex vs. calcium soap of olive fatty acids. AB - A 3 x 3 Latin Square experiment was designed to compare 2 ways of bypassing the effects of the rumen with olive oil fatty acids in 'Manchega' dairy ewes. Treatments were a control diet, CaOFA (control diet plus 45 g of olive fatty acids as calcium soap), and OO (control plus 45 g/d of olive fatty acids as olive oil emulsified in skim milk) and bottle-fed to animals trained to maintain the reticular groove reflex). No differences were found in milk, protein, and lactose yields, but fat yield and milk fat content were greater in treatments with added fat (CaOFA and OO). Content of short- and medium-chain fatty acids in milk fat was greater for control treatment than for the other 2 groups, the yield of these fatty acids being similar for all 3 diets, except that of C12:0, which was greater for the control treatment. Content and yield of C18:0 and isomers of C18:1 others than oleic acid were greater in milk from the CaOFA diet than from the other 2 diets. Oleic acid content and yield were greater in milk after OO treatment (23.9% and 16.8 g/d, respectively), intermediate after CaOFA treatment (19.2% and 13.8 g/d, respectively), and lower after control diet (10.7% and 6.52 g/d, respectively). Linoleic acid yield and content were greater in ewes fed the OO diet than in those on the other 2 diets, both of which showed similar data. All these changes indicated that the "protected" olive fatty acids (as calcium soap) were severely affected by the rumen environment and that the use of the reticular groove reflex seems to be a more effective way of bypassing the rumen in adult lactating dairy ewes. PMID- 15653541 TI - Feeding micronized and extruded flaxseed to dairy cows: effects on blood parameters and milk fatty acid composition. AB - Four lactating Holstein cows fitted with ruminal and duodenal cannulas were used in a 4 x 4 Latin square design to determine the effects of feeding micronized and extruded flaxseed on milk composition and blood profile in late lactation. Four diets were formulated: a control (C) diet with no flaxseed, a raw flaxseed (RF) diet, a micronized flaxseed (MF) diet, and an extruded flaxseed (EF) diet. Flaxseed diets contained 12.6% flax-seed (dry matter basis). Experimental periods consisted of 21 d of diet adaptation and 7 d of data collection. Feeding flaxseed reduced milk yield and energy-corrected milk by 1.8 and 1.4 kg/d, respectively. Yields of milk protein and casein were also lower for cows fed flaxseed diets than for those fed the C diet. Milk yield (1.6 kg/d) and milk fat percentage (0.4 percentage unit) were lower for cows fed EF than those fed MF. Plasma cholesterol and nonesterified fatty acid concentrations were higher for cows fed flaxseed diets relative to those fed the C diet. Flaxseed supplementation decreased plasma concentrations of medium-chain (MCFA) and saturated (SFA) fatty acids and increased concentrations of long-chain (LCFA) and monounsaturated fatty acids. Feeding flaxseed reduced the concentrations of short-chain fatty acids (SCFA), MCFA, and SFA in milk fat. Consequently, concentrations of LCFA and unsaturated fatty acids were higher for cows fed flaxseed diets than for those fed the C diet. Flaxseed supplementation increased average concentrations of C(18:3) and conjugated linoleic acid by 152 and 68%, respectively. Micronization increased C(18:3) level, and extrusion reduced concentrations of SCFA and SFA in milk. It was concluded that feeding raw or heated flaxseed to dairy cows alters blood and milk fatty acid composition. Feeding extruded flaxseed relative to raw or micronized flaxseed had negative effects on milk yield and milk composition. PMID- 15653542 TI - Influence of extruded soybeans with or without bicarbonate on milk performance and fatty acid composition of goat milk. AB - The effects of extruded soybeans (ESB) included at 0, 10, or 20% of dry matter (DM) of the diet in combination with sodium bicarbonate (0 vs. 1% bicarbonate added to DM) on rumen fermentation characteristics, production parameters, and fatty acid (FA) profiles of milk fat were examined in 30 midlactation goats and 6 rumen-cannulated goats fed high-concentrate diets (30:70 forage-to-concentrate ratio) ad libitum in a 3 x 2 factorial design. Diets were fed as total mixed rations. The trial lasted 13 wk with the final 9 wk as the test period. Milk yield and composition were recorded each week throughout the trial. Individual samples of milk were taken in wk 4, 7, 10, 11, and 13 to determine FA profile of milk fat. Dry matter intake and intake of net energy for lactation were not affected by dietary treatments. Feeding ESB did not modify ruminal pH or volatile fatty acids concentration in the rumen fluid, but it increased the molar proportion of propionate. Feeding ESB increased fat-corrected milk, milk fat content, and fat yield compared with the control diets. There was no change in milk protein content when ESB were fed. Feeding ESB increased the proportions of oleic, linoleic, and linolenic acids in milk fat at the expense of most of the saturated FA. It also increased the n-6 to n-3 FA ratio of milk. The largest changes in milk yield and milk composition were generally obtained with ESB included at 20% of DM. The addition of sodium bicarbonate tended to increase ruminal pH, VFA concentrations in the rumen fluid, and the molar proportions of acetate. The addition of sodium bicarbonate increased milk fat content and fat yield, with no change in milk FA composition. It is concluded that during midlactation, the inclusion of ESB to 20% of DM prevented low milk fat content for goats fed high-concentrate diets, with no decrease in milk protein content. The addition of sodium bicarbonate may enhance the effects of ESB on milk fat content and fat yield. PMID- 15653543 TI - Immune parameters of dry cows fed mannan oligosaccharide and subsequent transfer of immunity to calves. AB - The objective of this study was to evaluate the benefits of supplementation of mannan oligosaccharide (MOS) to cows during the last 3 wk of the dry period on immune function of the cows and subsequent transfer of passive immunity to their calves. Indicators of nonspecific and specific immunity were evaluated. Cows were vaccinated against rotavirus at 4 and 2 wk before expected parturition. Blood samples were obtained from cows before vaccination and at weekly intervals until calving and from calves at birth and 24 h for analysis of serum protein concentrations, packed cell volume, white blood cell counts, white blood cell differentials, and serum rotavirus neutralization titers. Colostrum quantity and quality were measured at calving, and immunoglobulin isotype concentrations in colostrum were determined. Specific immunity was enhanced by MOS supplementation as evidenced by greater serum rotavirus neutralization titers at calving in cows supplemented with MOS compared with control cows. Colostral rotavirus neutralization titers were not affected by treatment. Although numerical differences appeared large, there was a high degree of variability in the colostral rotavirus neutralization titers. Calves from cows fed MOS tended to have greater serum rotavirus neutralization titers compared with calves from cows fed the control diet. There was a tendency for greater increases in serum protein concentrations from birth to 24 h in calves from cows fed MOS compared with calves from cows fed the control diet. Results indicate that supplementation of MOS to cows during the dry period enhanced their immune response to rotavirus and tended to enhance the subsequent transfer of rotavirus antibodies to calves. PMID- 15653544 TI - In vitro production of Holstein embryos using sex-sorted sperm and oocytes from selected cull cows. AB - The objective of this study was to explore potential synergies between sex-sorted sperm and in vitro embryo production for generating replacement heifers on commercial dairy farms. Selected involuntary cull cows (i.e., genetically suitable cows that were culled due to injury, illness, or infertility) from 7 Wisconsin farms were used as donors, and ovaries were collected via colpotomy or at the time of slaughter. Oocytes were aspirated, fertilized in vitro with sex sorted sperm 22 +/- 0.2 h later, cultured, matured for 7 to 8 d, and transferred into recipient cows and heifers on the farms from which the cull cows originated. From August 2002 to June 2003, ovaries were recovered from 104 Holstein donors. Sex-sorted sperm from 3 Holstein sires (obtained via fluorescence-activated cell sorting) were used. A total of 365 transferable embryos were produced, an average of 3.6 +/- 0.3 per donor. However, due to limited availability of recipient animals, only 272 (fresh) embryos were transferred, an average of 2.6 +/- 0.3 per donor. A random subset of recipients received an injection (i.m.) of GnRH (100 microg) at the time of embryo transfer. When lactating cows were used as recipients, mean conception rates were 16.3% for recipients identified based on standing estrus and 20.0% for recipients synchronized using a timed breeding program (Ovsynch). Conception rates for in vitro-produced embryos were lower than corresponding conception rates for control cows inseminated using unsorted semen. When virgin heifers were used as embryo recipients (all standing estrus), the mean conception rate was 34.2%. The following effects significantly impacted conception rate: farm, season, recipient group (cow vs. heifer), sire of embryo, and GnRH injection. Of 40 full-term calves generated using sex-sorted semen, 37 were female. These results suggest that "low-cost" in vitro embryo production using cull cows as donors, in conjunction with sex-sorted sperm, could be an effective tool in dairy cattle breeding programs, but only if conception rates can be improved. PMID- 15653545 TI - Genetic analysis of an artificial insemination progeny test program. AB - The success of the progeny test (PT) program from one Spanish artificial insemination (AI) organization was evaluated. The annual genetic trend for the organization was compared with PT programs from other countries. The relationships among parents' estimated breeding values (EBV) and PT results for sons were also studied. Estimated breeding values for type and production traits were obtained from international genetic evaluations from February 2004. The annual genetic gain of the Spanish PT program was similar to that of other international programs. The Spanish AI organization graduated 13% of its sampled bulls, and 52% of primiparous cows were daughters of Spanish bulls (32% from proven bulls and 20% from sampling bulls). Correlations between EBV for PT bulls and their pedigree indices (0.52 to 0.70) were slightly lower than correlations between EBV for PT bulls and their parent averages (0.63 to 0.73). Both young and mature cows contributed to genetic progress. Success of PT bulls (defined by number of second-crop daughters) depended mainly on their EBV for final score, protein yield, and the type-production index. Significant correlations of sire EBV were found for final score and type-production index with the number of second-crop daughters (0.22 and 0.17). Likewise, significant correlations of dam EBV for final score and type-production index with the number of second crop daughters were found (0.25 and 0.18). Final score and protein yield were the main factors in success of a PT bull. The type-production index for PT bulls was not important for success unless it was 2.5 standard deviations above average. The PT bulls with low EBV for type-production index were used as proven bulls when they had higher EBV either for protein or final score. PMID- 15653546 TI - Comparisons of Holsteins with Brown Swiss and Jersey cows on the same farm for age at first calving and first calving interval. AB - Our objective was to evaluate breed differences for heat-stress resistance as reflected by age at first calving and first calving interval. We examined the effect of geographic location and birth season on age at first calving, and geographic location and first calving season on first calving interval on Holsteins and Jerseys, and Holsteins and Brown Swiss located on the same farm. We defined 7 regions within the United States: Northwest, Central north, Northeast, Central, Central south, Southwest, and Southeast, and analyzed 7 individual states: Ohio, Wisconsin, Oregon, California, Arizona, Texas, and Florida. Brown Swiss were older than Holsteins at first calving (833 +/- 2.4 vs. 806 +/- 2.0 d in regions, and 830 +/- 3.1 vs. 803 +/- 2.4 d in states), but Holsteins and Brown Swiss did not differ for first calving interval. Jerseys were younger than Holsteins at first calving and had shorter first calving intervals. In data from individual states, Holsteins housed with Brown Swiss were older at first calving than were Holsteins housed with Jerseys (800 +/- 2.7 vs. 780 +/- 2.5 d). Holsteins housed with one breed or the other were analyzed as a separate data set, and referred to as "type of Holstein." The interaction of "type of Holstein" with first calving season was highly significant for first calving interval. Geographic location and season effects were smaller for Jerseys than for Holsteins; thus, Jerseys showed evidence of heat-stress resistance with respect to Holsteins. Management modified age at first calving in Holsteins to more nearly match that of the other breed. Longer calving intervals might be partly due to voluntary waiting period to breed the cows. PMID- 15653547 TI - Comparison between linear models and survival analysis for genetic evaluation of clinical mastitis in dairy cattle. AB - Clinical mastitis was analyzed with mixed linear models (LM) and survival analysis (SA) using data from the first 3 lactations of >200,000 Swedish Holstein cows having their first calving between 1995 and 2000. The model for both methods included fixed effects of year-month and age at calving, fixed regressions of proportions of heterosis and North American Holstein genes, and random effects of herd-year at calving and sire. For the LM, clinical mastitis was defined as a binary trait measured from 10 d before to 150 d after calving. For the SA, clinical mastitis was defined either as the time period from 10 d before calving to the day of first treatment or culling because of mastitis (uncensored record) or from 10 d before to the day of next calving, culling for reasons other than mastitis, movement to a new herd, or to lactation d 240 (censored record). The heritability estimates from SA (0.03 to 0.04) were higher than those obtained with the LM (0.01 to 0.03). Consequently, the accuracies of estimated transmitting abilities were also higher for the trait analyzed with SA. The difference between estimates from the 2 methods was greater for later lactations. This study reveals the potential of analyzing clinical mastitis data with SA. PMID- 15653548 TI - Assessment of the impact of somatic cell count on functional longevity in Holstein and Jersey cattle using survival analysis methodology. AB - Survival analysis in a Weibull proportional hazards model was used to evaluate the impact of somatic cell count (SCC) on the involuntary culling rate of US Holstein and Jersey cows with first calvings from 1990 to 2000. The full data set, consisting of records from 978,043 Holstein and 250,835 Jersey cows, was divided into subsets (5 for Holsteins and 3 for Jerseys) based on herd average lactation SCC values. Functional longevity (also known as herd life or length of productive life) was defined as days from first calving until culling or censoring, after correcting for milk production. Our model included the time dependent effects of herd-year-season, parity by stage of lactation interaction, within-herd-year quintile ranking for mature equivalent production, and lactation average SCC (rounded to the nearest 50,000 cells/mL), as well as the time independent effect of age at first calving. Parameters of the Weibull distribution, as well as variance components for herd-year-season effects, were estimated within each group of herds. Mean failure and censoring times decreased as herd average SCC increased, and a nonlinear relationship was observed between SCC and longevity in all groups. The risk of culling for Holstein cows with lactation average SCC > 700,000 cells/mL was 3.4, 2.7, or 2.3 times greater, respectively, than that of Holstein cows with SCC of 200,000 to 250,000 cells/mL in herds with low, medium, or high average SCC. Likewise, the risk of culling for Jersey cows with lactation average SCC > 700,000 cells/mL was 4.0, 2.9, or 2.2 times greater, respectively, than that of Jersey cows with SCC of 200,000 to 250,000 cells/mL in low, medium, or high SCC herds. These trends may reflect more stringent culling of high SCC cows in herds with few mastitis problems. In addition, cows with lactation average SCC <100,000 cells/mL had a slightly higher risk of culling than cows with SCC of 100,000 to 200,000 cells/mL in both breeds, particularly in herds with high average SCC, where exposure to mastitis pathogens was likely. PMID- 15653549 TI - Effectiveness of national and regional sire evaluations in predicting future daughter milk yield. AB - National and regional bull evaluations were compared for ability to predict standardized milk yield of future daughters. Correlations between evaluations and first-, second-, and third-parity yields of future daughters were calculated within herd-year-month group. Mean correlations with predicted yield of future daughters across the United States were higher for national (0.109, 0.111, and 0.082 for first, second, and third parities, respectively) than for Northeast (0.098, 0.085, and 0.061) Holstein evaluations; corresponding correlations for future Northeast daughters were similar. Bull evaluations based on the first 5 parities of daughters that first calved through 1991 from either California, North Central, Northeast, or Southeast regions as well as from the entire United States were compared with standardized milk yields of daughters that calved later. Correlations with first-, second-, and third-parity yields of future daughters were higher (from 0.001 to 0.011) for national than for regional evaluations. National evaluations were better predictors of future-daughter yield, especially for California and the Southeast. Evaluations based on only first parity were slightly better than those based on the first 5 parities in predicting first-parity yield for 3 of 4 regions but were far less useful in predicting second-or third-parity yield regardless of region. Regional evaluations included fewer bulls because of limited numbers of daughters in each region. The top 100 bulls for genetic merit for milk yield based on regional rankings were inferior to the top 100 bulls based on national ranking by 25 to 173 kg. Reliance on regional rather than national evaluations would reduce current US genetic gains. PMID- 15653550 TI - Genetic and phenotypic relationships among milk yield and somatic cell count before and after clinical mastitis. AB - This paper studies whether cows with originally lower somatic cell count (SCC) are more susceptible to clinical mastitis (CM) than cows with higher somatic cell count, and evaluates the correlations between CM, SCC, and milk yield. Data were extracted from the Finnish national milk-recording database and from the health recording system. First and second lactation records of 87,861 Ayrshire cows calving between January 1998 and December 2000 were included. Traits studied were incidence of CM, test-day SCC, and test-day milk yield before and following CM. Genetic parameters were estimated using multitrait REML with a sire model. Results did not indicate that cows with genetically low SCC would be more susceptible to CM. The genetic correlation between CM in the first and second lactation was reasonably high (0.73), suggesting that susceptibility to mastitis remains similar across lactations. The genetic correlation between CM and milk yield traits was positive (from 0.38 to 0.56), confirming the genetic antagonism between production and udder health traits. The genetic correlation between SCC and milk was positive in the first lactation, but negative, or near zero in the second lactation. This indicates that breeding for lower SCC might not affect milk production in later lactations. The results of this study support the use of SCC as an indicator of mastitis and a tool for selection for mastitis resistance. PMID- 15653551 TI - Genetic evaluation of mastitis resistance using a first-passage time model for wiener processes for analysis of time to first treatment. AB - A Wiener process is a Brownian-motion process initiated in a certain state in a state space, and the first passage time is defined as the time of the process to reach a predefined absorbing state where the process stops. Time from 31 d prepartum to first treatment of clinical mastitis (CM) was modeled as first passage times of such Wiener processes. Two processes were used to allow for several risk factors, and for each process, initiation was at some arbitrary time point, in a certain health state with drift toward or away from absorption (disease). The drift parameter of each process was expressed as linear functions of covariates (year of calving and sire). First passage time was defined as the time from process initiation until the first health status process reached zero (absorption). The model was fitted to records for 36,178 first-lactation daughters of 245 Norwegian cattle sires using a Bayesian approach and Markov chain Monte Carlo methods. Genetic evaluation of sires was carried out by calculating the posterior probability of no CM (the value of the survival function) by d 331, i.e., 300 d after first calving. Alternatively, sire evaluation was based on the integrated area under the survival curve. These measures were highly correlated (0.999), which indicates a small degree of crossings of the sire-dependent survival curves. Hence, sire-specific hazards were close to proportional, resulting in a higher rank-correlation to sire evaluations from a survival model with proportional hazards than to the results from a multivariate threshold model. PMID- 15653552 TI - Peroxisome proliferator-activated receptor gamma-independent ablation of cyclin D1 by thiazolidinediones and their derivatives in breast cancer cells. AB - In light of the clinical relevance of targeting cyclin D1 in breast cancer, we have investigated the mechanism underlying the effect of the peroxisome proliferator-activated receptor-gamma (PPARgamma) agonists troglitazone and ciglitazone on cyclin D1 repression. We obtain evidence that the ability of high doses of troglitazone and ciglitazone to repress cyclin D1 is independent of PPARgamma activation. PPARgamma-inactive troglitazone and ciglitazone analogs 5 [4-(6-hydroxy-2,5,7,8-tetramethyl-chroman-2-yl-methoxy)-benzylidene]-2,4 thiazolidinedione (Delta2-TG) and 5-[4-(1-methyl-cyclohexylmethoxy)-benzylidene] thiazolidine-2,4-dione are able to facilitate cyclin D1 ablation with potency similar to that of troglitazone and ciglitazone in MCF-7 cells. Reverse transcription-polymerase chain reaction shows that the mRNA level of cyclin D1 remains unaltered in drug-treated cells, indicating the repression is mediated at the post-transcriptional level. Moreover, the ablative effect of these agents is specific to cyclin D1, in that the expression levels of many other cyclins and cyclin-dependent kinases examined remain unchanged after drug treatment. Our data indicate that troglitazone- and Delta2-TG-induced cyclin D1 repression is mediated via proteasome-facilitated proteolysis because it is inhibited by different proteasome inhibitors, including N-carbobenzoxy-l-leucinyl-l-leucinyl-l norleucinal (MG132), lactacystin, and epoxomicin, and is preceded by increased ubiquitination. The dissociation of these two pharmacological activities (i.e., PPARgamma activation and cyclin D1 ablation) provides a molecular basis to use Delta2-TG as a scaffold to develop a novel class of cyclin D1-ablative agents. Therefore, a series of Delta2-TG derivatives have been synthesized. Among them, 5 [4-(6-allyoxy-2,5,7,8-tetramethyl-chroman-2-yl-methoxy)-benzylidene]-2,4 thiazolidinedione represents a structurally optimized agent with potency that is an order of magnitude higher than that of Delta2-TG in cyclin D1 repression and MCF-7 cell growth inhibition. PMID- 15653553 TI - WITHDRAWN: Up-Regulation of Insulin Receptor Substrate-1 and -2 by Neuronal Nicotinic Receptor-Induced Sequential Activation of Protein Kinase C-{alpha} and Extracellular Signal-Regulated Kinase (ERK): Enhancement of Insulin-Induced Phosphoinositide 3-Kinase and ERK Signaling Pathways in Adrenal Chromaffin Cells. AB - Ahead of Print article withdrawn by publisher PMID- 15653554 TI - Elastin peptides activate extracellular signal-regulated kinase 1/2 via a Ras independent mechanism requiring both p110gamma/Raf-1 and protein kinase A/B-Raf signaling in human skin fibroblasts. AB - Elastin peptides (EPs) produced during cancer progression bind to the elastin binding protein (EBP) found at the surface of dermal fibroblasts, leading to the expression of collagenase-1 gene. The production of this enzyme involved in stromal reaction is caused by the sustained activation of the extracellular signal-regulated kinases 1/2 (ERK1/2) pathway via cAMP/protein kinase A (PKA) and phosphatidylinositol 3-kinase (PI3K). However, the mechanism of these signaling events remains unknown. We show that kappa-elastin (kappaE), a commonly used EP, induces maximum phosphorylation of mitogen-activated protein kinase/extracellular signal-regulated kinase (MEK)1/2 and ERK1/2 after 30 min. The simultaneous inhibition of PKA and PI3K, by N-(2-(p-bromocinnamylamino)ethyl)-5 isoquinolinesulfonamide (H89) and 2-(4-morpholynil)-8-phenyl-4H-1-bemzopyran-4 one (LY294002), respectively, blocked MEK1/2 and ERK1/2 phosphorylation, as did lactose, an EBP antagonist. kappaE induced Raf-1 phosphorylation and activation in a PI3K-dependent manner. In our system, the PI3K p110gamma is expressed and activated by betagamma-derived subunits from a pertussis toxin-sensitive G protein after fibroblast stimulation. Pertussis toxin also blocks the Raf 1/MEK1/2/ERK1/2 phosphorylation cascade. In addition, we found that B-Raf is expressed in dermal fibroblasts and activated in a PKA-dependent manner after kappaE treatment, thereby integrating PKA signals to MEK1/2. It is noteworthy that Ras involvement was excluded because ERK1/2 activation by kappaE was not blocked in RasN17-transfected fibroblasts. Together, our results identify a novel Ras-independent ERK1/2 activation system in which p110gamma/Raf-1/MEK1/2 and PKA/B-Raf/MEK1/2 cooperate to activate ERK1/2. Thus, p110gamma and B-Raf seem to be important modulators of dermal fibroblasts physiology and should now qualify as therapeutic targets in strategies aiming at limiting elastin degradation contribution to cancer progression. PMID- 15653555 TI - Interaction of rearing environment and reproductive tactic on gene expression profiles in Atlantic salmon. AB - Organisms that share the same genotype can develop into divergent phenotypes, depending on environmental conditions. In Atlantic salmon, young males of the same age can be found either as sneakers or immature males that are future anadromous fish. Just as the organism-level phenotype varies between divergent male developmental trajectories, brain gene expression is expected to vary as well. We hypothesized that rearing environment can also have an important effect on gene expression in the brain and possibly interact with the reproductive tactic adopted. We tested this hypothesis by comparing brain gene expression profiles of the two male tactics in fish from the same population that were reared in either a natural stream or under laboratory conditions. We found that expression of certain genes was affected by rearing environment only, while others varied between male reproductive tactics independent of rearing environment. Finally, more than half of all genes that showed variable expression varied between the two male tactics only in one environment. Thus, in these fish, very different molecular pathways can give rise to similar macro-phenotypes depending on rearing environment. This result gives important insights into the molecular underpinnings of developmental plasticity in relationship to the environment. PMID- 15653556 TI - Retention of latent centromeres in the Mammalian genome. AB - The centromere is a cytologically defined entity that possesses a conserved and restricted function in the cell: it is the site of kinetochore assembly and spindle attachment. Despite its conserved function, the centromere is a highly mutable portion of the chromosome, carrying little sequence conservation across taxa. This divergence has made studying the movement of a centromere, either within a single karyotype or between species, a challenging endeavor. Several hypotheses have been proposed to explain the permutability of centromere location within a chromosome. This permutability is termed "centromere repositioning" when described in an evolutionary context and "neocentromerization" when abnormalities within an individual karyotype are considered. Both are characterized by a shift in location of the functional centromere within a chromosome without a concomitant change in linear gene order. Evolutionary studies across lineages clearly indicate that centromere repositioning is not a rare event in karyotypic evolution and must be considered when examining the evolution of chromosome structure and syntenic order. This paper examines the theories proposed to explain centromere repositioning in mammals. These theories are interpreted in light of evidence gained in human studies and in our presented data from the marsupial model species Macropus eugenii, the tammar wallaby. PMID- 15653557 TI - Adaptive evolution of eye degeneration in the Mexican blind cavefish. AB - The evolutionary mechanisms responsible for eye degeneration in cave-adapted animals have not been resolved. Opposing hypotheses invoking neural mutation or natural selection, each with certain genetic and developmental expectations, have been advanced to explain eye regression, although little or no experimental evidence has been presented to support or reject either theory. Here we review recent developmental and molecular studies in the teleost Astyanax mexicanus, a single species consisting of a sighted surface-dwelling form (surface fish) and many blind cave-dwelling forms (cavefish), which shed new light on this problem. The manner of eye development and degeneration, the ability to experimentally restore eyes, gene expression patterns, and comparisons between different cavefish populations all provide important clues for understanding the evolutionary forces responsible for eye degeneration. A key discovery is that Hedgehog midline signaling is expanded and inhibits eye formation by inducing lens apoptosis in cavefish embryos. Accordingly, eyes could have been lost by default as a consequence of natural selection for constructive traits, such as feeding structures, which are positively regulated by Hh signaling. We conclude from these studies that eye degeneration in cavefish may be caused by adaptive evolution and pleiotropy. PMID- 15653558 TI - New perspectives on eye development and the evolution of eyes and photoreceptors. AB - Recent experiments on the genetic control of eye development have opened up a completely new perspective on eye evolution. The demonstration that targeted expression of one and the same master control gene, that is, Pax6 can induce the formation of ectopic eyes in both insects and vertebrates, necessitates a reconsideration of the dogma of a polyphyletic origin of the various eye types in all the animal phyla. The involvement of Pax6 and six1 and six3 genes, which encode highly conserved transcription factors, in the genetic control of eye development in organisms ranging from planarians to humans argues strongly for a monophyletic origin of the eye. Because transcription factors can control the expression of any target gene provided it contains the appropriate gene regulatory elements, the conservation of the genetic control of eye development by Pax6 among all bilaterian animals is not due to functional constraints but a consequence of its evolutionary history. The prototypic eyes postulated by Darwin to consist of two cells only, a photoreceptor and a pigment cell, were accidentally controlled by Pax6 and the subsequent evolution of the various eye types occurred by building onto this original genetic program. A hypothesis of intercalary evolution is proposed that assumes that the eye morphogenetic pathway is progressively modified by intercalation of genes between the master control genes on the top of the hierarchy and the structural genes like rhodopsin at the bottom. The recruitment of novel genes into the eye morphogenetic pathway can be due to at least two different genetic mechanisms, gene duplication and enhancer fusion.In tracing back the evolution of eyes beyond bilaterians, we find highly developed eyes in some box-jellyfish as well as in some Hydrozoans. In Hydrozoans the same orthologous six genes (six1 and six3) are required for eye regeneration as in planarians, and in the box jellyfish Tripedalia a pax B gene, which may be a precursor of Pax6, was found to be expressed in the eyes. In contrast to the adults, which have highly evolved eyes, the Planula larva of Tripedalia has single- celled photoreceptors similar to some unicellular protists. For the origin of photoreceptor cells in metazoa, I propose two hypotheses, one based on cellular differentiation and a more speculative one based on symbiosis. The former assumes that photoreceptor cells originated from a colonial protist in which all the cells were photosensitive and subsequent cellular differentiation to give rise to photoreceptor cells. The symbiont hypothesis, which I call the Russian doll model, assumes that photosensitivity arose first in photosynthetic cyanobacteria that were subsequently taken up into red algae as primary chloroplasts. The red algae in turn were taken up by dinoflagellates as secondary chloroplasts and in some species evolved into the most sophisticated eye organelles, as found, for example, in some dinoflagellates like Erythropsis and Warnovia, which lack chloroplasts. Because dinoflagellates are commonly found as symbionts in cnidarians, the dinoflagellates may have transferred their photoreceptor genes to cnidarians. In cnidarians such as Tripedalia the step from photoreceptor organelles to multicellular eyes has occurred. These two hypotheses, the cellular differentiation and the symbiont hypothesis, are not mutually exclusive and are the subject of further investigations. PMID- 15653559 TI - Evolution of new hormone function: loss and gain of a receptor. AB - The vertebrate proglucagon gene encodes three glucagon-like sequences (glucagon, glucagon-like peptide-1 [GLP-1], and glucagon-like peptide 2 [GLP-2]) that have distinct functions in regulating metabolism in mammals. In contrast, glucagon and GLP-1 have similar physiological actions in fish, that of mammalian glucagon. We have identified sequences similar to receptors for proglucagon-derived peptides from the genomes of two fish (pufferfish and zebrafish), a frog (Xenopus tropicalis), and a bird (chicken). Phylogenetic analysis of the receptor sequences suggested an explanation for the divergent function of GLP-1 in fish and mammals. The phylogeny of our predicted and characterized receptors for proglucagon-derived peptides demonstrate that receptors for glucagon, GLP-1, and GLP-2 have an origin before the divergence of fish and mammals; however, fish have lost the gene encoding the GLP-1 class of receptors, and likely the incretin action of GLP-1. Receptors that bind GLP-1, but yield glucagon-like action, have been characterized in goldfish and zebrafish, and these sequences are most closely related to glucagon receptors. Both pufferfish and zebrafish have a second glucagon receptor-like gene that is most closely related to the characterized goldfish glucagon receptor. The phylogeny of glucagon receptor-like genes in fish indicates that a duplication of the glucagon receptor gene occurred on the ancestral fish lineage, and could explain the shared action of glucagon and GLP-1. We suggest that the binding specificity of one of the duplicated glucagon receptors has diverged, yielding receptors for GLP-1 and glucagon, but that ancestral downstream signaling has been maintained, resulting in both receptors retaining glucagon-stimulated downstream effects. PMID- 15653560 TI - A nucleotide substitution responsible for the tawny coat color mutation carried by the MSKR inbred strain of mice. AB - "Tawny" is an autosomal recessive coat color mutation found in a wild population of Mus musculus molossinus. The inbred strain MSKR carries the mutation. The causative gene Mc1r(taw) of the tawny phenotype is the second recessive allele at the melanocortin 1 receptor locus and is dominant to the first recessive allele, "recessive yellow" (Mc1r(e)). The Mc1r(taw) gene has six nucleotide substitutions, and its forecasted transcript has three amino acid substitutions (i.e., V101A, V216A, W252C). Though the nucleotide substitutions leading to V101A and V216A exist in various mouse strains, the nucleotide substitution leading to W252C exists in only tawny-colored mice. Thus this substitution is considered to be responsible for the expression of the tawny coat color. The frequency of the allele having this nucleotide substitution was 9.21% in the wild M. m. molossinus population inhabiting Sakai City, Osaka Prefecture, Japan, where the ancestral mice of the MSKR strain were captured. PMID- 15653561 TI - European mink-polecat hybridization events: hazards from natural process? AB - Determining the significance of hybridization events raises essential issues both in conservation and in evolutionary biology. Here, we report a genetic investigation of sympatric polecat and endangered European mink populations. Although the two species were morphologically very similar, the European mink and the polecat were easily discriminated from allozymes and microsatellites and showed a high level of private alleles (effective number of alleles: mink=1.45 and polecat=3.09). Nevertheless, the allozymic polymorphism remained lower in the European mink (4 loci, 10.5%) than in polecat (9 loci, 23.7%). Similarly, from microsatellite data, the polymorphism only reached 36% at 0.99 in the European mink; whereas in the polecat, the polymorphism reached 82% at 0.99. Natural hybridization events between two native species were detected. Because of the low fertility of hybrids, interbreeding could be regarded as producing "hybrid sink" that leads to a progressive assimilation of mink by polecat. Nonetheless, pure mink populations inhabited streams in western France, and hybridization events were only detected in areas where mink were rare and now presumed disappeared. Rather than revealing the poor efficiency of the specific recognition system, our results suggest that hybridization is associated with the scarcity of mating partners. PMID- 15653562 TI - Sex-linked quantitative trait loci for thermotolerance and length in the rainbow trout. AB - We hypothesized that correlation between growth traits and upper thermal tolerance (UTT) in rainbow trout (Oncorhynchus mykiss) might be explained by quantitative trait loci (QTL) localized to the same linkage groups. Microsatellites on three autosomal linkage groups carrying UTT QTL in rainbow trout were tested for associations with fork length (FL) and condition factor (K) in half-sib families of outbred rainbow trout and in backcrosses of trout lines selected on UTT. Additionally, we used a sex-linked microsatellite (OmyFGT19TUF) to test for marker-trait associations at the sex chromosomes. The sex-linked marker OmyFGT19TUF was significantly associated with FL and UTT, accounting for up to 9.6% and 9.7% of variance in these traits, respectively. Male advantages in FL (and, to a lesser extent, UTT) relative to their female sibs were dependent on the origin of the Y chromosome and thus varied among grandsire lines. However, males had higher K in a manner unrelated to Y chromosomal origin, suggesting a partially sex-limited expression of this trait. Omy325UoG was significantly associated with K in one of the outbred half-sib families, but no other significant autosomal marker-trait associations were detected. Our findings illustrate minor evidence that correlation between UTT and FL is partially determined by one or more sex-chromosomal QTL. PMID- 15653564 TI - Platelet heterogeneity: variation in coagulation complexes on platelet subpopulations. AB - OBJECTIVE: Previous work has shown that platelets stimulated with the combination of thrombin and convulxin, a glycoprotein VI agonist, develop 2 populations with different levels of alpha-granule factor V bound to the platelet surface. To evaluate whether this phenomenon is restricted to alpha-granule components or is a feature that can be generalized to other coagulation factors, we studied the binding of factors V, VIII, IX, and X on the surface of platelets stimulated by convulxin and thrombin. METHODS AND RESULTS: The relative amount of factors V, VIII, IX, and X on the surface of platelets stimulated with thrombin or convulxin plus thrombin was measured using flow cytometry. Simultaneous measurements of factor Xa and thrombin generation were obtained and correlated with the binding of coagulation factors on the platelet surface. The binding of factors V, VIII, IX, and X all increased on a subpopulation of platelets when stimulated with the combined agonists. The development of this platelet subpopulation is dependent on convulxin concentration and correlates with increases in factor Xa and thrombin generation. CONCLUSIONS: The development of increased coagulation factor binding to a subpopulation of platelets is not limited to alpha-granule components. Convulxin-induced increases in thrombin generation are regulated by the proportion of platelets with increased coagulation factor binding. PMID- 15653565 TI - Carotid plaque composition differs between ethno-racial groups: an MRI pilot study comparing mainland Chinese and American Caucasian patients. AB - OBJECTIVE: Ethnicity-based research may identify new clues to the pathogenesis of atherosclerotic disease. Therefore, we sought to determine whether carotid lesions differ between 20 Chinese and 20 Caucasian Americans by MRI. METHODS AND RESULTS: Inclusion criteria were >50% stenosis as measured by duplex ultrasound and recent symptoms attributed to carotid artery disease. The patients were imaged in 2 centers (Beijing, China and Seattle, Wash) using a standardized protocol. Both carotid arteries were reviewed quantitatively (lumen, wall, outer wall, tissue components) and morphologically (lesion types, fibrous cap status). Significant differences between the Chinese and Americans were found for the mean size of the lipid/necrotic core (13.6 versus 7.8 mm2; P=0.002), percentage of slices with calcified type VII lesions (1.6 versus 12.4%; P=0.03), and percentage of slices with early type III lesions (19.3 versus 9.3%; P=0.02). Furthermore, the mean outer wall area in the common carotid artery was larger in the Chinese population (P=0.007). CONCLUSIONS: This pilot study suggests that composition and morphology of atherosclerotic lesions in symptomatic carotid disease differ between ethno-racial groups. Quantitative MRI-based review of carotid atherosclerosis comparing plaque morphology and composition between ethno-racial groups is feasible, and future MRI studies may improve our understanding of the pathophysiology of this disease. PMID- 15653566 TI - High glucose induces human endothelial cell apoptosis through a phosphoinositide 3-kinase-regulated cyclooxygenase-2 pathway. AB - OBJECTIVE: Diabetes mellitus causes endothelial dysfunction. The precise molecular mechanisms by which hyperglycemia causes apoptosis in endothelial cells are not yet well understood. The aim of this study was to explore the role of cyclooxygenase-2 (COX-2) and the possible involvement of phosphoinositide 3 kinase (PI3K) signaling in high glucose (HG)-induced apoptosis in human umbilical vein endothelial cells (HUVECs). METHODS AND RESULTS: For detection of apoptosis, the morphological Hoechst staining and Annexin V/propidium iodide staining were used. Glucose upregulated COX-2 protein expression, which was associated with the induction of prostaglandin (PG) E2 (PGE2), caspase-3 activity, and apoptosis. Unexpectedly, we found that PI3K inhibitors could suppress COX-2 expression, PGE2 production, caspase-3 activity, and the subsequent apoptosis under HG condition. Glucose-induced activation of PI3K resulted in the downstream effector Akt phosphorylation. PI3K inhibitors effectively attenuated the intracellular reactive oxygen species (ROS) generation and nuclear factor kappaB (NF-kappaB) activation. Blocking the PI3K and Akt activities with the dominant-negative vectors greatly diminished the HG-triggered NF-kappaB activation and COX-2 expression and apoptosis. CONCLUSIONS: These results suggest that HG, via PI3K/Akt signaling, induces NF-kappaB-related upregulation of COX-2, which in turn triggers the caspase-3 activity that facilitates HUVEC apoptosis. Also, HG may cause ROS generation in HUVECs through a PI3K/Akt-dependent pathway. PMID- 15653567 TI - Regulation of vascular smooth muscle cell proliferation and migration by human sprouty 2. AB - OBJECTIVE: To determine whether the human sprouty 2 (hSPRY2) protein, an inhibitor of receptor tyrosine kinase actions, regulates vascular smooth muscle cell (VSMC) proliferation, migration, and neointima formation in injured carotid artery. METHODS AND RESULTS: The hSPRY2 protein or green fluorescent protein (GFP; control) was transduced into VSMCs by placing an N-terminal TAT epitope on the proteins. The transduction of TAT-tagged hSPRY2 (TAT-hSPRY2) but not TAT-GFP inhibited the ability of serum and different growth factors to stimulate migration of VSMCs. Likewise, TAT-hSPRY2 also inhibited VSMC proliferation in response to serum. The hSPRY2 microtubule association (amino acids 123-177) and membrane translocation (amino acids 178-194) domains were necessary for the biological actions of hSPRY2. In the rat carotid artery injury model, exposure of the injured vessel for 1 hour to TAT-hSPRY2, but not TAT-GFP, markedly inhibited growth of the neointima over the 28-day postangioplasty period as well as VSMC proliferation. The exogenously applied TAT-hSPRY2 was retained in the carotid arteries for at least 3 days after injury, and endogenous SPRY2 expression was maximized around day 14 after injury. The latter is perhaps a compensatory mechanism to regulate neointima formation. CONCLUSIONS: We conclude that TAT tagged proteins are efficiently transduced into VSMCs in vitro and in vivo, that hSPRY2 inhibits growth and migration of VSMCs, and that this protein can decrease neointimal growth after blood vessel injury. PMID- 15653568 TI - Nod1-mediated endothelial cell activation by Chlamydophila pneumoniae. AB - Seroepidemiological and animal studies, as well as demonstration of viable bacteria in atherosclerotic plaques, have linked Chlamydophila pneumoniae infection to development of chronic vascular lesions and coronary heart disease. Inflammation and immune responses are dependent on host recognition of invading pathogens. The recently identified cytosolic Nod proteins are candidates for intracellular recognition of bacteria, such as the obligate intracellular chlamydia. In the present study, mechanisms of endothelial cell activation by C. pneumoniae via Nod proteins were examined. Viable, but not heat-inactivated, chlamydia activated human endothelial cells, suggesting that invasion of these cells is necessary for their profound activation. Endothelial cells express Nod1. Nod1 gene silencing by small interfering RNA reduced C pneumoniae-induced IL-8 release markedly. Moreover, in HEK293 cells, overexpressed Nod1 or Nod2 amplified the capacity of C pneumoniae to induce nuclear factor kappaB (NF-kappaB) activation. Interestingly, heat-inactivated bacteria were still able to induced a NF-kappaB reporter gene activity via Nod proteins when transfected intracellularly, but not when provided from the extracellular side. In contrast, TLR2 sensed extracellular heat-inactivated chlamydia. In conclusion, we demonstrated that C pneumoniae induced a Nod1-mediated and Nod2-mediated NF kappaB activation in HEK293 cells. In endothelial cells, Nod1 played a dominant role in triggering a chlamydia-mediated inflammatory process. PMID- 15653569 TI - Genetic deletion of the A1 adenosine receptor limits myocardial ischemic tolerance. AB - Adenosine receptors may be important determinants of intrinsic ischemic tolerance. Genetically modified mice were used to examine effects of global A1 adenosine receptor (A1AR) knockout (KO) on function and ischemic tolerance in perfused mouse hearts. Baseline contractile function and heart rate were unaltered by A1AR KO, which was shown to abolish the negative chronotropic effects of 2-chloroadenosine (A1AR-mediated) without altering A2 adenosine receptor-mediated coronary dilation. Tolerance to 25 minutes global normothermic ischemia (followed by 45 minutes reperfusion) was significantly limited by A1AR KO, with impaired contractile recovery (reduced by 25%) and enhanced lactate dehydrogenase (LDH) efflux (increased by 100%). Functional effects of A1AR KO involved worsened systolic pressure development with little to no change in diastolic dysfunction. In contrast, cardiac specific A1AR overexpression enhanced ischemic tolerance with a primary action on diastolic dysfunction. Nonselective receptor agonism (10 micromol/L 2-chloroadenosine) protected wild-type and also A1AR KO hearts (albeit to a lesser extent), implicating protection via subtypes additional to A1ARs. However, A1AR KO abrogated effects of 2-chloroadenosine on ischemic contracture and diastolic dysfunction. These data are the first demonstrating global deletion of the A1AR limits intrinsic myocardial resistance to ischemia. Data indicate the function of intrinsically activated A1ARs appears primarily to be enhancement of postischemic contractility and limitation of cell death. PMID- 15653570 TI - Cystatin C deficiency increases elastic lamina degradation and aortic dilatation in apolipoprotein E-null mice. AB - The pathogenesis of atherosclerosis and abdominal aortic aneurysm involves substantial proteolysis of the arterial extracellular matrix. The lysosomal cysteine proteases can exert potent elastolytic and collagenolytic activity. Human atherosclerotic plaques have increased cysteine protease content and decreased levels of the endogenous inhibitor cystatin C, suggesting an imbalance that would favor matrix degradation in the arterial wall. This study tested directly the hypothesis that impaired expression of cystatin C alters arterial structure. Cystatin C-deficient mice (Cyst C-/-) were crossbred with apolipoprotein E-deficient mice (ApoE-/-) to generate cystatin C and apolipoprotein E-double deficient mice (Cyst C-/-ApoE-/-). After 12 weeks on an atherogenic diet, cystatin C deficiency yielded significantly increased tunica media elastic lamina fragmentation, decreased medial size, and increased smooth muscle cell and collagen content in aortic lesions of ApoE-/- mice. Cyst C-/-ApoE /- mice also showed dilated thoracic and abdominal aortae compared with control ApoE-/- mice, although atheroma lesion size, intimal macrophage accumulation, and lipid core size did not differ between these mice. These findings demonstrate directly the importance of cysteine protease/protease inhibitor balance in dysregulated arterial integrity and remodeling during experimental atherogenesis. PMID- 15653571 TI - Dual mechanisms regulating AMPK kinase action in the ischemic heart. AB - AMP-activated protein kinase (AMPK) is emerging as an important signaling protein during myocardial ischemia. AMPK is a heterotrimeric complex containing an alpha catalytic subunit and beta and gamma regulatory subunits. Phosphorylation of Thr172 in the activation loop of the alpha subunit by upstream AMPK kinase(s) (AMPKK) is a critical determinant of AMPK activity. However, the mechanisms regulating AMPK phosphorylation in the ischemic heart remain uncertain and were therefore investigated. In the isolated working rat heart, low-flow ischemia rapidly activated AMPKK activity when measured using recombinant AMPK (rAMPK) as substrate. The addition of AMP (10 to 200 micromol/L) augmented the ability of heterotrimeric alpha1beta1gamma1 or alpha2beta1gamma1 rAMPK to be phosphorylated by heart AMPKK in vitro, whereas physiologic concentrations of ATP inhibited rAMPK phosphorylation. However, neither AMP nor ATP directly influenced AMPKK activity: they had no effect on AMPKK-mediated phosphorylation of rAMPK substrates lacking normal AMP-binding gamma subunits (isolated truncated alpha1(1 312) or alpha1beta1gamma1 rAMPK containing an R70Q mutation in the gamma1 AMP binding site). Regional ischemia in vivo also increased AMPKK activity and AMPK phosphorylation in the rat heart. AMPK phosphorylation could also be induced in vivo without activating AMPKK: AICAR infusion increased AMPK phosphorylation without activating AMPKK; however, the AMP-mimetic AICAR metabolite ZMP enhanced the ability of heterotrimeric rAMPK to be phosphorylated by AMPKK. Thus, heart AMPKK activity is increased by ischemia and its ability to phosphorylate AMPK is highly modulated by the interaction of AMP and ATP with the heterotrimeric AMPK complex, indicating that dual mechanisms regulate AMPKK action in the ischemic heart. PMID- 15653572 TI - Detection of vascular adhesion molecule-1 expression using a novel multimodal nanoparticle. AB - Endothelial vascular adhesion molecule-1 (VCAM-1) is a critical component of the leukocyte-endothelial adhesion cascade, and its strict temporal and spatial regulation make it an ideal target for imaging and therapy. The goal of this study was to develop novel VCAM-1-targeted imaging agents detectable by MRI and fluorescence imaging using phage display-derived peptide sequences and multimodal nanoparticles (NPs). We hypothesized that VCAM-1-mediated cell internalization of phage display-selected peptides could be harnessed as an amplification strategy to chaperone and trap imaging agents inside VCAM-1-expressing cells, thus improving target-to-background ratios. To accomplish our goal, iterative phage display was performed on murine endothelium under physiological flow conditions to identify a family of VCAM-1-mediated cell-internalizing peptides. One specific sequence, containing the VHSPNKK motif that has homology to the alpha-chain of very late antigen (a known ligand for VCAM-1), was shown to bind VCAM-1 and block leukocyte-endothelial interactions. Compared with VCAM-1 monoclonal antibody, the peptide showed 12-fold higher target-to-background ratios. A VHSPNKK-modified magnetofluorescent NP (VNP) showed high affinity for endothelial cells expressing VCAM-1 but surprisingly low affinity for macrophages. In contrast, a control NP without VCAM-1-targeting sequences showed no affinity for endothelial cells. In vivo, VNP successfully identified VCAM-1-expressing endothelial cells in a murine tumor necrosis factor-alpha-induced inflammatory model and colocalized with VCAM 1-expressing cells in atherosclerotic lesions present in cholesterol-fed apolipoprotein E apoE-/- mice. These results indicate that: (1) small peptide sequences can significantly alter targeting of NPs, (2) the used amplification strategy of internalization results in high target-to-background ratios, and (3) this technology is useful for in vivo imaging of endothelial markers. PMID- 15653573 TI - Critical care and emergency medicine neurology in stroke. PMID- 15653574 TI - Advances in stroke. Introduction. PMID- 15653575 TI - Oxidative stress: the curse that underlies cerebral vascular dysfunction? PMID- 15653576 TI - Approval of the MERCI clot retriever: a critical view. PMID- 15653577 TI - Editorial comment--plaque pathology and patient selection for carotid endarterectomy. PMID- 15653578 TI - Systemic inflammatory response depends on initial stroke severity but is attenuated by successful thrombolysis. PMID- 15653579 TI - Preference-based quality of life in patients with cerebral aneurysms. AB - BACKGROUND AND PURPOSE: Functional outcome scales are typically used to measure quality of life (QOL) and outcomes in patients with cerebral aneurysms; however, these instruments only examine a limited number of domains that contribute to QOL. An alternative are preference-based QOL methods, which integrate all factors contributing to QOL and provide a comprehensive individualized measure of how patients value their current health state. An additional advantage of preference based QOL values is that they can be incorporated into decision analyses and cost effectiveness analyses. METHODS: We used 4 preference-based QOL methods to measure QOL in 176 outpatients with cerebral aneurysms: (1) standard gamble; (2) time trade-off; (3) visual analogue scale; and (4) willingness to pay. We measured functional status with the Glasgow Outcome Scale (GOS), Rankin Scale, and Barthel Index. We then built multivariate linear regression models to examine the relationships between preference-based QOL, functional status, and patient characteristics. RESULTS: Preference-based QOL was moderately diminished in the aneurysm patients. Mean values were: standard gamble, 0.78; time trade-off, 0.79; visual analogue scale, 0.67; and willingness to pay, 121,000 dollars. Preference based QOL was not well-explained by functional status or patient characteristics, as shown by regression models that accounted for <15% of the variation in preference-based QOL (R2<0.15). CONCLUSIONS: Preference-based QOL instruments capture components of QOL in patients with cerebral aneurysms not assessed by functional status measures or patient characteristics. Studies of patients with cerebral aneurysms should consider incorporating preference-based QOL measures for a fuller evaluation of the impact of aneurysmal disease and its treatment on QOL. PMID- 15653581 TI - Carotid plaque pathology: thrombosis, ulceration, and stroke pathogenesis. AB - BACKGROUND AND PURPOSE: To determine the relationship between ulceration, thrombus, and calcification of carotid artery atherosclerotic plaques and symptoms of ipsilateral or contralateral stroke. METHODS: We compared microscopic plaque morphology from patients with and without stroke symptoms ipsilateral or contralateral to the plaque. Plaques were characterized for ulceration, thrombus, and calcification. We analyzed plaques from 241 subjects: 170 patients enrolled in the Asymptomatic Carotid Atherosclerosis Study (ACAS) and 71 patients enrolled in the North American Symptomatic Carotid Endarterectomy Trial (NASCET); 128 subjects had no history of stroke symptoms, 80 subjects had ipsilateral symptoms, and 33 had contralateral symptoms. RESULTS: Plaque ulceration was more common in plaques taken from symptomatic patients than those without symptoms (36% versus 14%; P<0.001); frequency of ulceration was similar for plaques associated with ipsilateral (34%) and contralateral (42%) symptoms. Thrombus was most common in plaques taken from patients with both ipsilateral symptoms and ulceration. The extent of calcification was unassociated with stroke symptoms. CONCLUSIONS: Carotid plaque ulceration and thrombosis are more prevalent in symptomatic patients. Ulceration is more common in symptomatic patients regardless of side of carotid symptoms, whereas thrombus is associated with ipsilateral symptoms and plaque ulceration. Preoperative identification of carotid ulceration and thrombus should lead to greater efficacy of stroke prevention by carotid endarterectomy. PMID- 15653582 TI - Restenosis after carotid angioplasty, stenting, or endarterectomy in the Carotid and Vertebral Artery Transluminal Angioplasty Study (CAVATAS). AB - BACKGROUND AND PURPOSE: Carotid and Vertebral Artery Transluminal Angioplasty Study (CAVATAS) patients with carotid stenosis were randomized between endovascular treatment and endarterectomy. The rates of residual severe stenosis and restenosis and their contribution to recurrent symptoms was unclear. METHODS: Endovascular patients were treated by balloon angioplasty alone (88%) or stenting (22%). Patches were used in 63% of endarterectomy patients. Carotid stenosis was categorized as mild (0% to 49%), moderate (50% to 69%), severe (70% to 99%), or occluded, using standardized Doppler ultrasound criteria at the examination closest to 1 month (n=283) and 1 year (n=347) after treatment. Recurrent cerebrovascular symptoms during follow-up were analyzed. RESULTS: More patients had > or =70% stenosis of the ipsilateral carotid artery 1 year after endovascular treatment than after endarterectomy (18.5% versus 5.2%, P=0.0001). Residual severe stenosis was present in 6.5% of patients at 1 month after endovascular treatment. Between 1 month and 1 year, restenosis to > or =70% stenosis occurred in 10.5% of the endovascular group. After endarterectomy, 1.7% had residual severe stenosis at 1 month, and 2.5% developed severe restenosis. The results were significantly better after stenting compared with angioplasty alone at 1 month (P<0.001) but not at 1 year. Recurrent ipsilateral symptoms were more common in endovascular patients with severe stenosis (5/32 [15.6%]) compared with lesser degrees of stenosis at 1 year (11/141 [7.8%], P=0.02), but most were transient ischemic attacks and none were disabling or fatal strokes. There were no recurrent symptoms in the 9 endarterectomy patients with > or =70% stenosis at 1 year. CONCLUSIONS: Carotid stenosis 1 year after endovascular treatment is partly explained by poor initial anatomical results and partly by restenosis. The majority of patients were treated by angioplasty without stenting. Further randomized studies are required to determine whether newer carotid stenting techniques are associated with a lower risk of restenosis. The low rate of recurrent stroke in both endovascular and endarterectomy patients suggests that treatment of restenosis should be limited to patients with recurrent symptoms, but long term follow up data are required. PMID- 15653583 TI - Pelvic pain: overlooked and underdiagnosed gynecologic conditions. AB - Chronic pelvic pain is a common, disabling problem among women. Although chronic pelvic pain can be produced by many conditions, some gynecologic causes are frequently overlooked and underdiagnosed, resulting in inappropriate referral and inadequate treatment. The gynecologic conditions most often unrecognized are endometriosis, adenomyosis, pelvic congestion, and less common congenital and acquired abnormalities. Transvaginal ultrasonography (US) is helpful for assessing endometriotic cysts but has a limited role in the diagnosis of adhesions or peritoneal implants. The classic magnetic resonance (MR) imaging features diagnostic of endometrioma are a cystic mass with high signal intensity on T1-weighted images and loss of signal intensity on T2-weighted images. When transvaginal US findings are suggestive of adenomyosis, MR imaging is used as the definitive imaging modality for diagnosis. High-resolution transvaginal US and MR imaging can help establish the diagnosis of adenomyosis with a high degree of accuracy, since the imaging appearance closely correlates with the histopathologic characteristics. Pelvic varices can be identified by using transvaginal US with color Doppler and Doppler spectral analysis. Three dimensional T1 gradient-echo sequences performed after the intravenous administration of gadolinium are the most effective MR imaging sequence for demonstrating pelvic varices. Blood flow in pelvic varices appears with high signal intensity. Recent advances in radiologic imaging and therapeutic procedures make it possible to diagnose accurately the conditions producing chronic pelvic pain in most women and to guide effective treatment. PMID- 15653584 TI - MR imaging findings of adenomyosis: correlation with histopathologic features and diagnostic pitfalls. AB - Adenomyosis is a nonneoplastic condition, characterized by benign invasion of ectopic endometrium into the myometrium with hyperplasia of adjacent smooth muscle. The common symptoms include dysmenorrhea, menorrhagia, and abnormal uterine bleeding, but these do not allow diagnosis. Therefore, imaging plays an important role because establishment of the correct preoperative diagnosis is critical to avoid unnecessary intervention. Magnetic resonance (MR) imaging is a highly accurate noninvasive modality for diagnosis of adenomyosis, differentiation of adenomyosis from other gynecologic disorders, and planning of appropriate treatment. Although the typical MR imaging findings are well established, adenomyosis actually varies widely in terms of histopathologic features (adenomyosis with sparse glands), growth patterns (polypoid adenomyoma, adenomyotic cyst, and miniature uterus), responses to hormonal activity (tamoxifen, decidual changes), and responses to treatment (gonadotropin-releasing hormone agonist). The MR imaging findings of adenomyosis occasionally mimic those of uterine malignancy or ovarian cancer. Furthermore, malignancy occasionally develops in otherwise benign adenomyosis. Pitfalls in diagnosis of adenomyosis include myometrial contractions, leiomyoma, adenomatoid tumor, metastases, endometrial carcinoma, and endometrial stromal sarcoma. Knowledge of the various appearances of adenomyosis and the possible pitfalls in differential diagnosis help guide the determination of appropriate treatment options. PMID- 15653585 TI - MR imaging appearances of the female pelvis after trachelectomy. AB - Carcinoma of the cervix has a predilection for affecting young women. In recent years, surgical procedures that combine local radical surgery with maintenance of fertility potential in young women have been investigated. One such procedure is radical trachelectomy with pelvic lymphadenectomy, in which the corpus uteri is preserved, thus maintaining fertility potential. Magnetic resonance (MR) imaging is useful in the selection of suitable patients for trachelectomy and in postsurgical follow-up. The MR imaging findings in 45 patients who underwent trachelectomy at one institution between 1996 and 2004 were retrospectively reviewed. The uterovaginal anastomosis has variable appearances, with an end-to end anastomotic appearance or a vaginal neofornix seen after surgery; suture artifacts can also occur. Possible changes affecting the vagina include diffuse wall thickening and hematomas, whereas lymphoceles and exaggeration of the pelvic venous plexuses can occur in the pelvis. Adenomyosis and endometriosis may be found incidentally in this patient group. Successful pregnancies after trachelectomy have been reported and should be treated as high-risk pregnancies. Knowledge of these MR imaging appearances will help radiologists distinguish between normal postsurgical variations, benign postsurgical changes, and recurrent disease in patients who have undergone trachelectomy. PMID- 15653586 TI - Diffusion-tensor MR imaging and fiber tractography: a new method of describing aberrant fiber connections in developmental CNS anomalies. AB - Congenital anomalies of the central nervous system (CNS) often demonstrate aberrant white matter connections, which may be better characterized with diffusion-tensor imaging (DTI) and fiber tractography (FT) than with conventional magnetic resonance (MR) imaging. DTI-FT demonstrates abnormal hemispheric fiber connections in callosal agenesis or acquired disease of the corpus callosum. Decreased anisotropy of white matter adjacent to the malformed cortex and an aberrant course of major fiber pathways due to dysplastic white matter are common findings in cortical dysplasia. Increased anisotropy of dysplastic gray matter in heterotopia supports the hypothesis that developing neurons migrate from the ependyma to the cortex with a radial growth pattern. In periventricular leukomalacia, DTI-FT demonstrates an intact corticospinal tract and decreased thalamocortical sensory connections, which are responsible for the spasticity of cerebral palsy owing to impairment of inhibitory function. Joubert syndrome comprises malformation of the cerebellar vermis and an aberrant connection between the cerebellum and the cerebral cortex via an elongated and abnormally shaped superior cerebellar peduncle, which are well visualized with DTI-FT. In developmental CNS disease, DTI-FT demonstrates additional findings beyond those seen with conventional MR imaging. Future studies will focus on determining the significance of the aberrant fiber connections and their relationships to the clinical manifestations of CNS anomalies. PMID- 15653588 TI - CT and MR imaging of extrahepatic fatty masses of the abdomen and pelvis: techniques, diagnosis, differential diagnosis, and pitfalls. AB - The differential diagnosis of extrahepatic abdominopelvic masses is wide. Demonstration of fat within a lesion at noninvasive imaging is an important clue for narrowing the differential diagnosis. Macroscopic fat is readily identified with both computed tomography (CT) and magnetic resonance (MR) imaging. Demonstration of microscopic fat is more difficult and may require special techniques. Identification of fat with CT is based on x-ray resorption and therefore on the attenuation (typically less than -20 HU). Several MR imaging techniques have been developed for fat suppression. Two of the most widely available are spectroscopic fat saturation and chemical shift (in-phase/opposed phase) imaging. Entities with predominantly macroscopic fat include myelolipoma, angiomyolipoma, teratoma, liposarcoma, lipoma, epiploic appendagitis, omental infarction, and mesenteric panniculitis. Lesions with predominantly microscopic fat include adrenal adenoma and some teratomas. Other fat-containing entities involve the mesentery and bowel wall; these include fibrofatty mesenteric proliferation and submucosal fat deposition. PMID- 15653589 TI - CT in blunt liver trauma. AB - Nonsurgical treatment has become the standard of care in hemodynamically stable patients with blunt liver trauma. The use of helical computed tomography (CT) in the diagnosis and management of blunt liver trauma is mainly responsible for the notable shift during the past decade from routine surgical to nonsurgical management of blunt liver injuries. CT is the diagnostic modality of choice for the evaluation of blunt liver trauma in hemodynamically stable patients and can accurately help identify hepatic parenchymal injuries, help quantify the degree of hemoperitoneum, and reveal associated injuries in other abdominal organs, retroperitoneal structures, and the gastrointestinal tract. The CT features of blunt liver trauma include lacerations, subcapsular or parenchymal hematomas, active hemorrhage, juxtahepatic venous injuries, periportal low attenuation, and a flat inferior vena cava. It is important that radiologists be familiar with the liver injury grading system based on these CT features that was established by the American Association for the Surgery of Trauma. CT is also useful in the assessment of delayed complications in blunt liver trauma, including delayed hemorrhage, hepatic or perihepatic abscess, posttraumatic pseudoaneurysm and hemobilia, and biliary complications such as biloma and bile peritonitis. Follow up CT is needed in patients with high-grade liver injuries to identify potential complications that require early intervention. PMID- 15653590 TI - Imaging of intraarticular masses. AB - Intraarticular masses can be classified as noninfectious synovial proliferative processes (lipoma arborescens, synovial osteochondromatosis, pigmented villonodular synovitis, rheumatoid arthritis), infectious granulomatous diseases (tuberculous arthritis, coccidioidomycosis arthritis), deposition diseases (gout, amyloid arthropathy), vascular malformations (synovial hemangioma, arteriovenous malformations), malignancies (synovial chondrosarcoma, synovial sarcoma, synovial metastases), and miscellaneous (cyclops lesion). Knowledge of articular anatomy aids the radiologist in localizing masses to the joint space. Some joints have complex anatomy with contiguous or adjacent bursae, recesses, and tendinous connections from which masses may originate or into which masses may extend. Many of the diseases causing intraarticular masses have specific imaging characteristics, especially on magnetic resonance images, and knowledge of these characteristics will allow for a more confident diagnosis. PMID- 15653591 TI - Adult ureteropelvic junction obstruction: insights with three-dimensional multi detector row CT. AB - Ureteropelvic junction obstruction (UPJO) is a benign, congenital condition that remains an enigma in terms of both diagnosis and therapy. On the basis of a series of cases that were referred to the authors in their clinical practice, they found that the unprecedented quality and novel perspectives of multi detector row computed tomography (CT) with two- and three-dimensional postprocessing allow a comprehensive, single-study assessment of the ureterovascular relationships in UPJO. This topic is important because the causative role of crossing vessels may be questioned on the basis of such studies, and the therapeutic approach may be altered by using precise anatomic images customized to the pathologic features of the individual patient. Although CT images can provide only circumstantial or supportive evidence of the pathophysiology of UPJO, they greatly facilitate therapeutic intervention when it is clinically indicated and may eliminate the need for it in select cases. Careful attention to the postprocessing of CT images may show that the crossing vasculature has no direct relationship to the transition point of the UPJO in many cases. PMID- 15653592 TI - Thoracic manifestations of tropical parasitic infections: a pictorial review. AB - Parasitic infections are distributed worldwide and affect hundreds of millions of individuals-primarily those living in endemic areas or in regions with a high rate of immigration from endemic areas-causing significant morbidity and mortality. A broad spectrum of parasitic infections (eg, amebiasis, malaria, trypanosomiasis, ascariasis, strongyloidiasis, dirofilariasis, cystic echinococcosis, schistosomiasis, paragonimiasis) frequently affect the lungs, mediastinum, and thoracic wall, manifesting with abnormal imaging findings that often make diagnosis challenging. Although most of these infections result in nonspecific abnormalities, familiarity with their imaging features as well as their epidemiologic, clinical, and physiopathologic characteristics may be helpful to the radiologist in formulating an adequate differential diagnosis. PMID- 15653593 TI - Stent-graft placement for the treatment of thoracic aortic diseases. AB - The recent development of aortic stent-grafts has brought the management of thoracic aortic diseases into the realm of interventional radiology. Stent-graft placement is now an alternative to surgery for the treatment of descending thoracic aortic aneurysms, ulcers, and fistulas and is sometimes indicated in cases of mycotic aneurysm, posttraumatic aortic rupture, or thoracic descending aortic dissection. Pretreatment imaging is crucial for evaluating patient eligibility, selecting the appropriate stent-graft, and planning the intervention. Stent-graft treatment of long atherosclerotic aneurysms, lesions close to aortic branch vessels, and aortic dissections is subject to technical pitfalls, and adverse events such as endoleaks, stent migration or misplacement, aortic perforation, and vascular trauma will require specific interventions, although they occur in only a minority of patients. Thoracic stent-graft placement in good surgical candidates remains controversial because long-term results are unknown. However, short-term morbidity and mortality rates from endovascular treatment compare favorably with those from surgery, and stent-graft placement is proving to be a safe, minimally invasive, and effective treatment for thoracic aortic diseases and is already the best option in many affected patients who are poor surgical candidates. PMID- 15653594 TI - Aortic fenestration: a why, when, and how-to guide. AB - The management of aortic dissection can be challenging. Most cases of acute type A dissection are managed surgically. Most cases of acute type B dissection are managed medically, although open surgery or stent-graft placement is sometimes performed. Patients with type B or surgically treated type A dissection may develop vascular complications such as mesenteric or peripheral ischemia, which cannot be managed medically. Aortic fenestration is a method for decompressing the hypertensive false lumen by creating a hole in the distal part of the dissection flap. This procedure allows outflow from the false lumen, thereby reducing intraluminal pressure, relieving branch vessel obstruction, and reducing the risk of extension of the dissection. Urgent revascularization is required to correct mesenteric and renal ischemia and to reestablish distal perfusion if there is resting ischemia. Few operators will acquire extensive personal experience with percutaneous aortic fenestration. Nevertheless, with a good understanding of the pathologic condition, careful demonstration of the anatomy, good technical skills, and access to high-quality imaging (including intravascular ultrasonography) and the requisite equipment, most interventional radiologists skilled in arterial interventions should be capable of performing this procedure. However, because further interventions are frequently required, the radiologist needs to maintain contact with the patient to ensure timely treatment of any subsequent complications. PMID- 15653595 TI - FDG PET in the evaluation of treatment for lymphoma: clinical usefulness and pitfalls. AB - Positron emission tomography (PET) with 2-[fluorine-18] fluoro-2-deoxy-d-glucose (FDG) may play an important role in the evaluation and management of malignant lymphoma. FDG uptake is predictive of therapeutic response during the course of treatment. After completion of chemotherapy, residual abnormalities representing either residual tumor or necrotic or fibrotic tissue are not uncommon, and FDG PET may be more accurate than computed tomography (CT) or magnetic resonance imaging in assessing residual disease and identifying patients who require more intense treatment. However, posttreatment FDG PET does not help exclude the presence of minimal residual disease, which may lead to disease relapse. Furthermore, FDG is not a tumor-specific substance, and increased accumulation may be seen in a variety of benign entities and scenarios (eg, infection, drug toxicity, granulocyte colony-stimulating factor therapy, radiation therapy, physiologic activity, postoperative or postbiopsy changes, fracture, degenerative change, injection leakage), which may yield false-positive findings. Nevertheless, recognition of these entities and correlation of FDG PET findings with clinical and other radiologic findings-especially those at combined PET and CT or PET-CT fusion imaging-allows improved diagnostic accuracy. If the interpretation of positive findings is exceptionally difficult, short-term follow up may be helpful. PMID- 15653596 TI - Scenes from the past: carthaginian pair of tweezers. PMID- 15653597 TI - From the archives of the AFIP: a comprehensive review of fetal tumors with pathologic correlation. AB - Fetal tumors are a diverse group of neoplasms, which are unique in their histologic characteristics, anatomic distribution, and pathophysiology. The biologic behavior of tumors in the fetus may differ dramatically compared with that of the same tumor detected later in life. Teratomas are the dominant histologic type and constitute the majority of both extracranial and intracranial neoplasms. Although often histologically mature, they may prove lethal because of their location and metabolic demands on the fetus. Large solid tumors may lead to cardiovascular compromise and hydrops fetalis. Extracranial teratomas are most commonly located in the sacrococcygeal area, followed by the head and neck, chest, and retroperitoneum. Fetuses with intracranial tumors have a poor prognosis regardless of histologic type. There are, however, two notable exceptions: lipomas and choroid plexus papillomas, both of which have a more favorable outcome. Neuroblastoma is the most common fetal malignancy. It may be either solid or cystic and is more often located on the right side. It typically has favorable biologic markers and stage at presentation. The prognosis for prenatally diagnosed cases is excellent. Other fetal neoplasms include soft tissue tumors (both benign and malignant), leukemia, mesenchymal hamartoma of the kidney, and liver tumors (hemangioendothelioma, mesenchymal hamartoma, and hepatoblastoma). PMID- 15653598 TI - Best cases from the AFIP: primary diffuse large B-cell lymphoma of the epididymis and testis. PMID- 15653599 TI - Cardiac metastases from melanoma. PMID- 15653600 TI - Informatics in Radiology (infoRAD): album of radiologic signs: a useful tool for training in radiologic semiology. AB - The importance of radiologic semiology can be seen from the large number of articles, books, and Web pages dealing with radiologic signs. Recognition of these signs forms an important part of the training process for radiologists. Computer-assisted instruction complements traditional techniques for teaching radiology and provides an appropriate method of collecting, organizing, and presenting radiologic signs. A computer application that includes more than 350 radiologic signs has been created with Microsoft PowerPoint (version 2002 for Windows). The user can navigate freely to see the signs from a menu that provides access according to anatomic region or from an interactive dictionary that includes 779 terms. From the presentation of each sign, the user can access one or more representative images, information about the examination wherein the sign appears, an explanation of the meaning of the sign and the corresponding diagnosis, additional comparative images (pathologic or normal), and relevant references. This album of radiologic signs is a useful tool for training in radiologic semiology and also allows a quick consultation based on the name of the sign. Although designed for interns and residents, it will also be useful in pregraduate teaching of radiology, as a reference, and in continuing medical education. PMID- 15653603 TI - American Board of Radiology: maintenance of certification. PMID- 15653602 TI - Informatics in Radiology (infoRAD): mobile wireless DICOM server system and PDA with high-resolution display: feasibility of group work for radiologists. AB - A novel mobile system has been developed for use by radiologists in managing Digital Imaging and Communications in Medicine (DICOM) image data. The system consists of a mobile DICOM server (MDS) and personal digital assistants (PDAs), including a Linux PDA with a video graphics array (VGA) display (307,200 pixels, 3.7 inches). The MDS weighs 410 g, has a 60-GB hard disk drive and a built-in wireless local area network (LAN) access point, and supports a DICOM server (Central Test Node). The Linux-based MDS can be accessed with personal computers (PCs) and PDAs by means of a wireless or wired LAN, and client-server communications can be established at any time. DICOM images can be displayed by using any PDA or PC by means of a Web browser. Simultaneous access to the MDS is possible for multiple authenticated users. With most PDAs, image compression is necessary for complete display of DICOM images; however, the VGA screen can display a 512 x 512-pixel DICOM image almost in its entirety. This wireless system allows efficient management of heavy loads of lossless DICOM image data and will be useful for collaborative work by radiologists in education, conferences, and research. PMID- 15653601 TI - Informatics in Radiology (infoRAD): three-dimensional atlas of the brain anatomy and vasculature. AB - Of the existing atlases of the brain anatomy and cerebrovasculature, none integrates the anatomy and vasculature by providing for direct manipulation of three-dimensional (3D) cerebral models. An atlas-based application was developed in four steps: (a) construction of 3D anatomic models, (b) construction of 3D vascular models, (c) interactive spatial coregistration of the anatomic and vascular models, and (d) development of functionality and a user interface for the application. Three-dimensional anatomic models were imported from an electronic brain atlas database derived from classic print atlases. A novel vascular modeling technique was developed and applied to create a vascular atlas from magnetic resonance angiographic data. The use of 3D polygonal models allows smooth navigation (rotation, zooming, panning) and interactive labeling of anatomic structures and vascular segments. This application enables the user to examine 3D anatomic structures and 3D cerebral vasculature and to gain a better understanding of the relationships between the two. The combined anatomic vascular atlas is a user-friendly neuroeducational tool that is useful for medical students and neuroscience researchers as well as for educators in preparing teaching materials. PMID- 15653604 TI - Abstracts of the American Dental Education Association (ADEA) 82nd Annual Meeting, March 5-9, 2005, Baltimore, Maryland, USA. PMID- 15653626 TI - Covalent antibody display--an in vitro antibody-DNA library selection system. AB - The endonuclease P2A initiates the DNA replication of the bacteriophage P2 by making a covalent bond with its own phosphate backbone. This enzyme has now been exploited as a new in vitro display tool for antibody fragments. We have constructed genetic fusions of P2A with single-chain antibodies (scFvs). Linear DNA of these fusion proteins were processed in an in vitro coupled transcription translation mixture of Escherichia coli S30 lysate. Complexes of scFv-P2A fusion proteins covalently bound to their own DNA were isolated after panning on immobilized antigen, and the enriched DNAs were recovered by PCR and prepared for the subsequent cycles of panning. We have demonstrated the enrichment of scFvs from spiked libraries and the specific selection of different anti-tetanus toxoid scFvs from a V-gene library with 50 million different members prepared from human lymphocytes. This covalent antibody display technology offers a complete in vitro selection system based exclusively on DNA-protein complexes. PMID- 15653627 TI - Detection and characterization of horizontal transfers in prokaryotes using genomic signature. AB - Horizontal DNA transfer is an important factor of evolution and participates in biological diversity. Unfortunately, the location and length of horizontal transfers (HTs) are known for very few species. The usage of short oligonucleotides in a sequence (the so-called genomic signature) has been shown to be species-specific even in DNA fragments as short as 1 kb. The genomic signature is therefore proposed as a tool to detect HTs. Since DNA transfers originate from species with a signature different from those of the recipient species, the analysis of local variations of signature along recipient genome may allow for detecting exogenous DNA. The strategy consists in (i) scanning the genome with a sliding window, and calculating the corresponding local signature (ii) evaluating its deviation from the signature of the whole genome and (iii) looking for similar signatures in a database of genomic signatures. A total of 22 prokaryote genomes are analyzed in this way. It has been observed that atypical regions make up approximately 6% of each genome on the average. Most of the claimed HTs as well as new ones are detected. The origin of putative DNA transfers is looked for among approximately 12 000 species. Donor species are proposed and sometimes strongly suggested, considering similarity of signatures. Among the species studied, Bacillus subtilis, Haemophilus Influenzae and Escherichia coli are investigated by many authors and give the opportunity to perform a thorough comparison of most of the bioinformatics methods used to detect HTs. PMID- 15653628 TI - Conditional UAS-targeted repression in Drosophila. AB - The Gal4-UAS enhancer trap system is useful for driving gene expression in various tissues. A new tool that extends Gal4 technology is described here. A fusion protein containing the Gal4 binding domain and the repression domain of the isolator suppressor of hairy wing was placed under the control of a heat shock-inducible promoter. The construct mediates the conditional repression of genes located downstream of a UAS sequence. The repressive effects of the chimeric protein on fasII gene expression were tested by western-blot analysis and in brain sections of adult Drosophila. Owing to the increasing number of Gal4 and UAS transgenic lines, this versatile system will facilitate the study of gene function. PMID- 15653629 TI - Use of a restriction enzyme-digested PCR product as substrate for helicase assays. AB - DNA helicases play essential roles in many cellular processes. The currently available techniques to generate substrates for helicase assays are fairly complicated and need some expertise not available in all laboratories. Here, a PCR-based method to generate a substrate for a helicase assay is described, and its application for several archaeal, bacterial and viral enzymes is demonstrated. PMID- 15653630 TI - BiSearch: primer-design and search tool for PCR on bisulfite-treated genomes. AB - Bisulfite genomic sequencing is the most widely used technique to analyze the 5 methylation of cytosines, the prevalent covalent DNA modification in mammals. The process is based on the selective transformation of unmethylated cytosines to uridines. Then, the investigated genomic regions are PCR amplified, subcloned and sequenced. During sequencing, the initially unmethylated cytosines are detected as thymines. The efficacy of bisulfite PCR is generally low; mispriming and non specific amplification often occurs due to the T richness of the target sequences. In order to ameliorate the efficiency of PCR, we developed a new primer-design software called BiSearch, available on the World Wide Web. It has the unique property of analyzing the primer pairs for mispriming sites on the bisulfite-treated genome and determines potential non-specific amplification products with a new search algorithm. The options of primer-design and analysis for mispriming sites can be used sequentially or separately, both on bisulfite treated and untreated sequences. In silico and in vitro tests of the software suggest that new PCR strategies may increase the efficiency of the amplification. PMID- 15653631 TI - Probing a rate-limiting step by mutational perturbation of AdoMet binding in the HhaI methyltransferase. AB - DNA methylation plays important roles via regulation of numerous cellular mechanisms in diverse organisms, including humans. The paradigm bacterial methyltransferase (MTase) HhaI (M.HhaI) catalyzes the transfer of a methyl group from the cofactor S-adenosyl-L-methionine (AdoMet) onto the target cytosine in DNA, yielding 5-methylcytosine and S-adenosyl-L-homocysteine (AdoHcy). The turnover rate (k cat) of M.HhaI, and the other two cytosine-5 MTases examined, is limited by a step subsequent to methyl transfer; however, no such step has so far been identified. To elucidate the role of cofactor interactions during catalysis, eight mutants of Trp41, which is located in the cofactor binding pocket, were constructed and characterized. The mutants show full proficiency in DNA binding and base-flipping, and little variation is observed in the apparent methyl transfer rate k chem as determined by rapid-quench experiments using immobilized fluorescent-labeled DNA. However, the Trp41 replacements with short side chains substantially perturb cofactor binding (100-fold higher K(AdoMet)D and K(AdoMet)M) leading to a faster turnover of the enzyme (10-fold higher k cat). Our analysis indicates that the rate-limiting breakdown of a long-lived ternary product complex is initiated by the dissociation of AdoHcy or the opening of the catalytic loop in the enzyme. PMID- 15653632 TI - Functional domains of the Xenopus replication licensing factor Cdt1. AB - During late mitosis and early G1, replication origins are licensed for subsequent replication by loading heterohexamers of the mini-chromosome maintenance proteins (Mcm2-7). To prevent re-replication of DNA, the licensing system is down regulated at other cell cycle stages. A small protein called geminin plays an important role in this down-regulation by binding and inhibiting the Cdt1 component of the licensing system. We examine here the organization of Xenopus Cdt1, delimiting regions of Cdt1 required for licensing and regions required for geminin interaction. The C-terminal 377 residues of Cdt1 are required for licensing and the extreme C-terminus contains a domain that interacts with an Mcm(2,4,6,7) complex. Two regions of Cdt1 interact with geminin: one at the N terminus, and one in the centre of the protein. Only the central region binds geminin tightly enough to successfully compete with full-length Cdt1 for geminin binding. This interaction requires a predicted coiled-coil domain that is conserved amongst metazoan Cdt1 homologues. Geminin forms a homodimer, with each dimer binding one molecule of Cdt1. Separation of the domains necessary for licensing activity from domains required for a strong interaction with geminin generated a construct, whose licensing activity was partially insensitive to geminin inhibition. PMID- 15653634 TI - Multiple promoters are a common feature of mitochondrial genes in Arabidopsis. AB - Mitochondrial genes in the plant Arabidopsis thaliana are transcribed by two phage-type RNA polymerases encoded in the nucleus. Little is known about cis elements that are recognized by these enzymes and mediate the transcription of the Arabidopsis mitochondrial genome. Here, 30 transcription initiation sites of 12 mitochondrial genes and gene clusters have been determined using 5'-RACE and ribonuclease protection analysis of primary transcripts labelled in vitro by guanylyltransferase. A total of 9 out of 12 genes were found to possess multiple promoters, revealing for the first time that multiple promoters are a common feature of mitochondrial genes in a dicotyledonous plant. No differences in promoter utilization were observed between leaves and flowers, suggesting that promoter multiplicity reflects a relaxed promoter specificity rather than a regulatory role of promoter selection. Nearly half the identified transcription initiation sites displayed immediately upstream a CRTA core sequence, which was mostly seen within the previously described CRTAAGAGA promoter motif or a novel CGTATATAA promoter element. About as many promoters possessed an ATTA or RGTA core. Our data indicate that the majority of mitochondrial promoters in Arabidopsis deviate significantly from the nonanucleotide consensus derived earlier for dicot mitochondrial promoters. PMID- 15653633 TI - The histone deacetylase inhibitor trichostatin A alters the pattern of DNA replication origin activity in human cells. AB - Eukaryotic chromatin structure limits the initiation of DNA replication spatially to chromosomal origin zones and temporally to the ordered firing of origins during S phase. Here, we show that the level of histone H4 acetylation correlates with the frequency of replication initiation as measured by the abundance of short nascent DNA strands within the human c-myc and lamin B2 origins, but less well with the frequency of initiation across the beta-globin locus. Treatment of HeLa cells with trichostatin A (TSA) reversibly increased the acetylation level of histone H4 globally and at these initiation sites. At all three origins, TSA treatment transiently promoted a more dispersive pattern of initiations, decreasing the abundance of nascent DNA at previously preferred initiation sites while increasing the nascent strand abundance at lower frequency genomic initiation sites. When cells arrested in late G1 were released into TSA, they completed S phase more rapidly than untreated cells, possibly due to the earlier initiation from late-firing origins, as exemplified by the beta-globin origin. Thus, TSA may modulate replication origin activity through its effects on chromatin structure, by changing the selection of initiation sites, and by advancing the time at which DNA synthesis can begin at some initiation sites. PMID- 15653635 TI - RDE-2 interacts with MUT-7 to mediate RNA interference in Caenorhabditis elegans. AB - In Caenorhabditis elegans, the activity of transposable elements is repressed in the germline. One of the mechanisms involved in this repression is RNA interference (RNAi), a process in which dsRNA targets cleavage of mRNAs in a sequence-specific manner. The first gene found to be involved in RNAi and transposon silencing in C.elegans is mut-7, a gene encoding a putative exoribonuclease. Here, we show that the MUT-7 protein resides in complexes of approximately 250 kDa in the nucleus and in the cytosol. In addition, we find that upon triggering of RNAi the cytosolic MUT-7 complex increases in size. This increase is independent of the presence of target RNA, but does depend on the presence of RDE-1 and RDE-4, two proteins involved in small interfering RNA (siRNA) production. Finally, using a yeast two-hybrid screen, we identified RDE 2/MUT-8 as one of the other components of this complex. This protein is encoded by the rde-2/mut-8 locus, previously implicated in RNAi and transposon silencing. Using genetic complementation analysis, we show that the interaction between these two proteins is required for efficient RNAi in vivo. Together these data support a role for the MUT-7/RDE-2 complex downstream of siRNA formation, but upstream of siRNA mediated target RNA recognition, possibly indicating a role in the siRNA amplification step. PMID- 15653637 TI - Comparative study of sequence-dependent hybridization kinetics in solution and on microspheres. AB - Hybridization kinetics of DNA sequences with known secondary structures and random sequences designed with similar melting temperatures were studied in solution and when one strand was bound to 5 mum silica microspheres. The rates of hybridization followed second-order kinetics and were measured spectrophotometrically in solution and fluorometrically in the solid phase. In solution, the rate constants for the model sequences varied by almost two orders of magnitude, with a decrease in the rate constant with increasing amounts of secondary structure in the target sequence. The random sequences also showed over an order of magnitude difference in the rate constant. In contrast, the hybridization experiments in the solid phase with the same model sequences showed almost no change in the rate constant. Solid phase rate constants were approximately three orders of magnitude lower compared with the solution phase constants for sequences with little or no single-stranded structure. Sequences with a known secondary structure yielded solution phase rate constants as low as 3 x 10(3) M(-1) s(-1) with solid phase rate constants for the same sequences measured at 2.5 x 10(2) M(-1) s(-1). The results from these experiments indicate that (i) solid phase hybridization occurs three orders of magnitude slower than solution phase, (ii) trends observed in structure-dependent kinetics of solution phase hybridization may not be applicable to solid phase hybridization and (iii) model probes with known secondary structure decrease reaction rates; however, even random sequences with no known internal single-stranded structure can yield a broad range of reaction rates. PMID- 15653636 TI - The BRCT domain of mammalian Rev1 is involved in regulating DNA translesion synthesis. AB - Rev1 is a deoxycytidyl transferase associated with DNA translesion synthesis (TLS). In addition to its catalytic domain, Rev1 possesses a so-called BRCA1 C terminal (BRCT) domain. Here, we describe cells and mice containing a targeted deletion of this domain. Rev1(B/B) mice are healthy, fertile and display normal somatic hypermutation. Rev1(B/B) cells display an elevated spontaneous frequency of intragenic deletions at Hprt. In addition, these cells were sensitized to exogenous DNA damages. Ultraviolet-C (UV-C) light induced a delayed progression through late S and G2 phases of the cell cycle and many chromatid aberrations, specifically in a subset of mutant cells, but not enhanced sister chromatid exchanges (SCE). UV-C-induced mutagenesis was reduced and mutations at thymidine thymidine dimers were absent in Rev1(B/B) cells, the opposite phenotype of UV-C exposed cells from XP-V patients, lacking TLS polymerase eta. This suggests that the enhanced UV-induced mutagenesis in XP-V patients may depend on error-prone Rev1-dependent TLS. Together, these data indicate a regulatory role of the Rev1 BRCT domain in TLS of a limited spectrum of endogenous and exogenous nucleotide damages during a defined phase of the cell cycle. PMID- 15653638 TI - Serum-deprivation stimulates cap-binding by PARN at the expense of eIF4E, consistent with the observed decrease in mRNA stability. AB - PARN, a poly(A)-specific ribonuclease, binds the 5' cap-structure of mRNA and initiates deadenylation-dependent decay. Eukaryotic initiation factor 4E (eIF4E) also binds to the cap structure, an interaction that is critical for initiating cap-dependent translation. The stability of various mRNA transcripts in human cell lines is reduced under conditions of serum starvation as determined by both functional and chemical half-lives. Serum starvation also leads to enhanced cap association by PARN. In contrast, the 5' cap occupancy by eIF4E decreases under serum-deprivation, as does the translation of reporter transcripts. Further, we show that PARN is a phosphoprotein and that this modification can be modulated by serum status. Taken together, these data are consistent with a natural competition existing at the 5' cap structure between PARN and eIF4E that may be regulated by changes in post-translational modifications. These phosphorylation induced changes in the interplay of PARN and eIF4E may determine whether the mRNA is translated or decayed. PMID- 15653639 TI - Plant tRNA ligases are multifunctional enzymes that have diverged in sequence and substrate specificity from RNA ligases of other phylogenetic origins. AB - Pre-tRNA splicing is an essential process in all eukaryotes. It requires the concerted action of an endonuclease to remove the intron and a ligase for joining the resulting tRNA halves as studied best in the yeast Saccharomyces cerevisiae. Here, we report the first characterization of an RNA ligase protein and its gene from a higher eukaryotic organism that is an essential component of the pre-tRNA splicing process. Purification of tRNA ligase from wheat germ by successive column chromatographic steps has identified a protein of 125 kDa by its potentiality to covalently bind AMP, and by its ability to catalyse the ligation of tRNA halves and the circularization of linear introns. Peptide sequences obtained from the purified protein led to the elucidation of the corresponding proteins and their genes in Arabidopsis and Oryza databases. The plant tRNA ligases exhibit no overall sequence homologies to any known RNA ligases, however, they harbour a number of conserved motifs that indicate the presence of three intrinsic enzyme activities: an adenylyltransferase/ligase domain in the N terminal region, a polynucleotide kinase in the centre and a cyclic phosphodiesterase domain at the C-terminal end. In vitro expression of the recombinant Arabidopsis tRNA ligase and functional analyses revealed all expected individual activities. Plant RNA ligases are active on a variety of substrates in vitro and are capable of inter- and intramolecular RNA joining. Hence, we conclude that their role in vivo might comprise yet unknown essential functions besides their involvement in pre-tRNA splicing. PMID- 15653640 TI - Destabilization of tetranucleotide repeats in Haemophilus influenzae mutants lacking RnaseHI or the Klenow domain of PolI. AB - A feature of Haemophilus influenzae genomes is the presence of several loci containing tracts of six or more identical tetranucleotide repeat units. These repeat tracts are unstable and mediate high frequency, reversible alterations in the expression of surface antigens. This process, termed phase variation (PV), enables H.influenzae to rapidly adapt to fluctuations in the host environment. Perturbation of lagging strand DNA synthesis is known to destabilize simple sequence repeats in yeast and Escherichia coli. By using a chromosomally located reporter construct, we demonstrated that the mutation of an H.influenzae rnhA (encoding RnaseHI) homologue increases the mutation rates of tetranucleotide repeats approximately 3-fold. Additionally, deletion of the Klenow domain of DNA polymerase I (PolI) resulted in a approximately 35-fold increase in tetranucleotide repeat-mediated PV rates. Deletion of the PolI 5'>3' exonuclease domain appears to be lethal. The phenotypes of these mutants suggest that delayed or mutagenic Okazaki fragment processing destabilizes H.influenzae tetranucleotide repeat tracts. PMID- 15653642 TI - Translocation of XRCC1 and DNA ligase IIIalpha from centrosomes to chromosomes in response to DNA damage in mitotic human cells. AB - DNA single-strand breaks (SSBs) are the most frequent lesions caused by oxidative DNA damage. They disrupt DNA replication, give rise to double-strand breaks and lead to cell death and genomic instability. It has been shown that the XRCC1 protein plays a key role in SSBs repair. We have recently shown in living human cells that XRCC1 accumulates at SSBs in a fully poly(ADP-ribose) (PAR) synthesis dependent manner and that the accumulation of XRCC1 at SSBs is essential for further repair processes. Here, we show that XRCC1 and its partner protein, DNA ligase IIIalpha, localize at the centrosomes and their vicinity in metaphase cells and disappear during anaphase. Although the function of these proteins in centrosomes during metaphase is unknown, this centrosomal localization is PAR dependent, because neither of the proteins is observed in the centrosomes in the presence of PAR polymerase inhibitors. On treatment of metaphase cells with H2O2, XRCC1 and DNA ligase IIIalpha translocate immediately from the centrosomes to mitotic chromosomes. These results show for the first time that the repair of SSBs is present in the early mitotic chromosomes and that there is a dynamic response of XRCC1 and DNA ligase IIIalpha to SSBs, in which these proteins are recruited from the centrosomes, where metaphase-dependent activation of PAR polymerase occurs, to mitotic chromosomes, by SSBs-dependent activation of PAR polymerase. PMID- 15653641 TI - Conserved transcription factor binding sites of cancer markers derived from primary lung adenocarcinoma microarrays. AB - Gene transcription in a set of 49 human primary lung adenocarcinomas and 9 normal lung tissue samples was examined using Affymetrix GeneChip technology. A total of 3442 genes, called the set M AD, were found to be either up- or down-regulated by at least 2-fold between the two phenotypes. Genes assigned to a particular gene ontology term were found, in many cases, to be significantly unevenly distributed between the genes in and outside M AD. Terms that were overrepresented in M AD included functions directly implicated in the cancer cell metabolism. Based on their functional roles and expression profiles, genes in M AD were grouped into likely co-regulated gene sets. Highly conserved sequences in the 5 kb region upstream of the genes in these sets were identified with the motif discovery tool, MoDEL. Potential oncogenic transcription factors and their corresponding binding sites were identified in these conserved regions using the TRANSFAC 8.3 database. Several of the transcription factors identified in this study have been shown elsewhere to be involved in oncogenic processes. This study searched beyond phenotypic gene expression profiles in cancer cells, in order to identify the more important regulatory transcription factors that caused these aberrations in gene expression. PMID- 15653643 TI - Probing the DNA kink structure induced by the hyperthermophilic chromosomal protein Sac7d. AB - Sac7d, a small, abundant, sequence-general DNA-binding protein from the hyperthermophilic archaeon Sulfolobus acidocaldarius, causes a single-step sharp kink in DNA (approximately 60 degrees) via the intercalation of both Val26 and Met29. These two amino acids were systematically changed in size to probe their effects on DNA kinking. Eight crystal structures of five Sac7d mutant-DNA complexes have been analyzed. The DNA-binding pattern of the V26A and M29A single mutants is similar to that of the wild-type, whereas the V26A/M29A protein binds DNA without side chain intercalation, resulting in a smaller overall bending (approximately 50 degrees). The M29F mutant inserts the Phe29 side chain orthogonally to the C2pG3 step without stacking with base pairs, inducing a sharp kink (approximately 80 degrees). In the V26F/M29F-GCGATCGC complex, Phe26 intercalates deeply into DNA bases by stacking with the G3 base, whereas Phe29 is stacked on the G15 deoxyribose, in a way similar to those used by the TATA box binding proteins. All mutants have reduced DNA-stabilizing ability, as indicated by their lower T m values. The DNA kink patterns caused by different combinations of hydrophobic side chains may be relevant in understanding the manner by which other minor groove-binding proteins interact with DNA. PMID- 15653644 TI - Locked nucleic acid (LNA) mediated improvements in siRNA stability and functionality. AB - Therapeutic application of the recently discovered small interfering RNA (siRNA) gene silencing phenomenon will be dependent on improvements in molecule bio stability, specificity and delivery. To address these issues, we have systematically modified siRNA with the synthetic RNA-like high affinity nucleotide analogue, Locked Nucleic Acid (LNA). Here, we show that incorporation of LNA substantially enhances serum half-life of siRNA's, which is a key requirement for therapeutic use. Moreover, we provide evidence that LNA is compatible with the intracellular siRNA machinery and can be used to reduce undesired, sequence-related off-target effects. LNA-modified siRNAs targeting the emerging disease SARS, show improved efficiency over unmodified siRNA on certain RNA motifs. The results from this study emphasize LNA's promise in converting siRNA from a functional genomics technology to a therapeutic platform. PMID- 15653645 TI - A pragmatic perspective on molecular targeted radionuclide therapy. PMID- 15653646 TI - Radiobiologic principles in radionuclide therapy. AB - Although the general radiobiologic principles underlying external beam therapy and radionuclide therapy are the same, there are significant differences in the radiobiologic effects observed in mammalian cells. External beam and brachytherapy emissions are composed of photons, whereas radiations of interest in radionuclide therapy are particulate. The special features that characterize the biologic effects consequent to the traversal of charged particles through mammalian cells are explored with respect to DNA lesions and cellular responses. Information about the ways in which these radionuclides are used to treat cancers in experimental models are highlighted. PMID- 15653647 TI - Combination radionuclide therapy using 177Lu- and 90Y-labeled somatostatin analogs. AB - Peptide receptor-targeted radionuclide therapy of somatostatin receptor expressing tumors is a promising application of radiolabeled somatostatin analogs. Suitable radionuclides are (90)Y, a pure, high-energy beta-emitter (2.27 MeV), and (177)Lu, a medium-energy beta-emitter (0.5 MeV) with a low-abundance gamma. METHODS: Lewis rats, each bearing both a small (approximately 0.5 cm(2)) and a large (7-9 cm(2)) somatostatin receptor-positive rat pancreatic CA20948 tumor in their flanks, were used. We investigated the radiotherapeutic effects of [(90)Y-tetraazacyclododecanetetraacetic acid (DOTA),Tyr(3)]octreotide, [(90)Y DOTA,Tyr(3)]octreotate, [(177)Lu-DOTA,Tyr(3)]octreotate, and the combination of (90)Y- and (177)Lu-labeled analogs at the same tumor radiation dose (60 Gy). RESULTS: Radiotherapeutic effects of the (90)Y- and (177)Lu-labeled analogs were found in the rat tumor model. In these animals bearing tumors of different sizes, the antitumor effects of the combination of 50% (177)Lu- plus 50% (90)Y-analogs were superior to those in animals treated with either (90)Y- or (177)Lu- analog alone. In smaller tumors, the (90)Y radiation energy was not completely absorbed in the tumor, whereas in larger tumors the increased number of clonogenic tumor cells at the fixed level of absorbed dose may account for the failure of (177)Lu alone to go completely into remission. CONCLUSION: This study shows the superior antitumor effects of the combination of (177)Lu- and (90)Y-somatostatin analogs when compared with either (90)Y- or (177)Lu-analog alone in animals bearing tumors of various sizes. PMID- 15653648 TI - Dosimetry of internal emitters. AB - Response and toxicity prediction is essential to rational implementation of cancer therapy. The biologic effects of radionuclide therapy are mediated via a well-defined physical quantity, the absorbed dose, which is defined as the energy absorbed per unit mass of tissue. The concepts, basic definitions, and different approaches to the clinical implementation of absorbed dose estimation are reviewed in this article. Ongoing efforts to improve the accuracy of dosimetry calculations are discussed, as well as studies examining the relationship between absorbed dose and response. Particular attention is placed on the marrow and kidney as dose-limiting organs. Finally, the potential role of radiobiologic modeling in helping to account for differences in dose rate and spatial distribution are reviewed. A treatment planning approach to radionuclide therapy will eventually require incorporation of biologic and radiobiologic considerations. Until such methods are developed and validated, absorbed dose remains an important variable--but still one of several--likely to predict response in an individual patient. PMID- 15653649 TI - The evolving role of (131)I for the treatment of differentiated thyroid carcinoma. AB - The use of radioactive iodine ((131)I) for the treatment of thyroid carcinoma has changed over the past 50 y. These changes are based on increasing awareness of the biophysical properties of (131)I and new discoveries concerning the biology of iodine handling by thyroid cells. The therapeutic administration of (131)I for thyroid remnant ablation and for metastases requires an appreciation of iodine clearance kinetics, of factors that can alter the occupancy time of (131)I within lesions, and of the role of thyroid-stimulating hormone in stimulating the sodium iodide symporter. The potential complications and adverse events associated with (131)I are discussed. (131)I will continue to be a major weapon in the fight against metastatic thyroid carcinoma. Its future role will be modified by expanding knowledge of its relative risks and benefits. PMID- 15653650 TI - Bone-seeking radionuclides for therapy. AB - Systemic therapy using bone-seeking radiopharmaceuticals has clear advantages for the treatment of multisite metastatic pain. Evidence supporting the use of beta particle, electron, and alpha-particle-emitting radiopharmaceuticals is reviewed here. Appropriate patient selection relies on correlating clinical symptoms with focal abnormalities on conventional bone scintigraphy. Time to symptom relief and response duration vary with the physical half-life and dose rate of the radionuclide used, offering the opportunity to tailor radiopharmaceutical choice to individual patient circumstances. Toxicity is limited to temporary myelosuppression, governed by the administered activity and underlying bone marrow reserve. Optimal responses are achieved in patients with a modest skeletal tumor burden, suggesting that targeted therapy should be considered early in the management of bone metastases. The development of reliable dosimetric models will facilitate patient-specific prescribing to deliver enhanced symptom response within acceptable toxicity limits. It is likely that targeted therapy will be most effective in the context of multimodality tumor management. Further research is required to examine the potential of radionuclides in combination with external-beam irradiation, bisphosphonates, or chemotherapy. This approach might allow targeted therapy to progress beyond symptom palliation to early intervention for survival gain. PMID- 15653651 TI - Radiosynovectomy in rheumatology, orthopedics, and hemophilia. AB - Radiosynovectomy (RSV) is a local intraarticular injection of radionuclides in colloidal form for radiotherapy. First used by Fellinger et al. in 1952, the technique has now been applied for more than 50 y for treatment of resistant synovitis of individual joints after failure of long-term systemic pharmacotherapy and intraarticular steroid injections. RSV relieves pain and inflammation from rheumatoid arthritis (RA), for which it initially was used, and is accepted as an alternative to surgical synovectomy in cases of RA or other inflammatory arthropathies such as osteoarthritis and hemophiliac arthropathy. A good understanding of the clinical pathophysiology of the disease processes is mandatory, and close interdisciplinary collaboration with other clinicians who diagnose and treat patients is strongly suggested. Reported success rates range from 40% to 90% for the different joints and underlying diseases. A few well designed prospective double-blind trials have evaluated the efficacy of RSV and justified the procedure as a viable option for treating chronic synovitis in RA or secondary to inflammatory arthropathies. In comparison with surgical synovectomy, RSV produces equivalent results, costs less, allows the patient to remain ambulatory, and is repeatable. RSV has to be considered the initial procedure of choice for the treatment of patients with hemarthrosis in hemophilia. In addition, local instillation of radiopharmaceuticals can effectively reduce effusions after implantation of a prosthesis. PMID- 15653652 TI - MIBG and somatostatin receptor analogs in children: current concepts on diagnostic and therapeutic use. AB - Nuclear imaging techniques such as bone scans, metaiodobenzylguanidine (MIBG) scans, and (111)In-diethylenetriaminepentaacetic acid-octreotide scans have greatly increased the sensitivity and specificity of both diagnostic and follow up protocols for pediatric solid tumors. Molecular targets that are specific for certain pediatric tumors are now being developed. Targets include cell membrane receptors targeted by specific ligands, subcellular organelles targeted by false transmitters, and cellular proteins targeted by antibodies. This review focuses on the use of MIBG (which is a false transmitter) and octreotide (which is a ligand for G protein receptor) in the diagnosis and treatment of solid tumors that affect children and young adults. PMID- 15653653 TI - Overview of results of peptide receptor radionuclide therapy with 3 radiolabeled somatostatin analogs. AB - A new treatment modality for inoperable or metastasized gastroenteropancreatic tumors is the use of radiolabeled somatostatin analogs. Initial studies with high doses of [(111)In-diethylenetriaminepentaacetic acid (DTPA)(0)]octreotide in patients with metastasized neuroendocrine tumors were encouraging, although partial remissions were uncommon. Another radiolabeled somatostatin analog that is used for peptide receptor radionuclide therapy (PRRT) is [(90)Y-1,4,7,10 tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid (DOTA)(0),Tyr(3)]octreotide. Various phase 1 and phase 2 PRRT trials have been performed with this compound. Despite differences in the protocols used, complete and partial remissions in most of the studies with [(90)Y-DOTA(0),Tyr(3)]octreotide were in the same ranges, 10%-30%; these ranges were higher than those obtained with [(111)In DTPA(0)]octreotide. Treatment with the newest radiolabeled somatostatin analog, [(177)Lu-DOTA(0),Tyr(3)]octreotate, which has a higher affinity for the subtype 2 somatostatin receptor, resulted in complete or partial remissions in 30% of 76 patients. Tumor regression was positively correlated with a high level of uptake on OctreoScan imaging, a limited hepatic tumor mass, and a high Karnofsky performance score. Treatment with radiolabeled somatostatin analogs is a promising new tool in the management of patients with inoperable or metastasized neuroendocrine tumors. Symptomatic improvement may occur with all (111)In-, (90)Y , or (177)Lu-labeled somatostatin analogs that have been used for PRRT. The results obtained with [(90)Y-DOTA(0),Tyr(3)]octreotide and [(177)Lu DOTA(0),Tyr(3)]octreotate are very encouraging in terms of tumor regression. Also, if kidney protective agents are used, the side effects of this therapy are few and mild, and the duration of the therapy response for both radiopharmaceuticals is more than 2 y. These data compare favorably with those for the limited number of alternative treatment approaches. PMID- 15653654 TI - Candidates for peptide receptor radiotherapy today and in the future. AB - Regulatory peptide receptors are overexpressed in numerous human cancers. These receptors have been used as molecular targets by which radiolabeled peptides can localize cancers in vivo and, more recently, to treat cancers with peptide receptor radiation therapy (PRRT). This review describes the candidate tumors eligible for such radiotherapy on the basis of their peptide receptor content and discusses factors in PRRT eligibility. At the present time, PRRT is performed primarily with somatostatin receptor- and cholecystokinin-2 (CCK2)-receptor expressing neuroendocrine tumors with radiolabeled octreotide analogs or with radiolabeled CCK2-selective analogs. In the future, PRRT may be extended to many other tumor types, including breast, prostate, gut, pancreas, and brain tumors, that have recently been shown to overexpress several other peptide receptors, such as gastrin-releasing peptide-, neurotensin-, substance P-, glucagon-like peptide 1-, neuropeptide Y-, or corticotropin-releasing factor-receptors. A wide range of radiolabeled peptides is being developed for clinical use. Improved somatostatin or CCK(2) analogs as well as newly designed bombesin, neurotensin, substance P, neuropeptide Y, and glucagon-like peptide-1 analogs offer promise for future PRRT. PMID- 15653655 TI - Molecular imaging as in vivo molecular pathology for gastroenteropancreatic neuroendocrine tumors: implications for follow-up after therapy. AB - Peptide receptor scintigraphy in combination with anatomic imaging methods such as CT can be regarded as molecular imaging. It offers insight into the variability of somatostatin receptor expression in neuroendocrine tumor lesions within a patient. The somatostatin receptor status of all tumors in a patient is an important issue, because receptor-negative lesions may be poorly differentiated and characterized by aggressive growth and poor prognosis, with consequences for the choice of therapy. METHODS: Follow-up studies of 3 patients with gastroenteropancreatic neuroendocrine tumors who had been previously treated with peptide receptor radionuclide therapy (PRRT) are presented. RESULTS: Patient 1 had a mixed response after treatment with (90)Y-1,4,7,10-tetraazacyclododecane N,N',N'',N'''-tetraacetic acid(0) (DOTA),Tyr(3)-octreotide. The response included fibrosis and a low rate of mitosis in receptor-positive lesions that had decreased in volume after treatment and vital tumor cells and a high rate of mitosis in receptor-negative lesions that had grown since the start of treatment. Patient 2 had a good clinical and biochemical response after PRRT with [(177)Lu DOTA(0),Tyr(3)]octreotate ((177)Lu-DOTATATE), with disappearance of (111)In pentetreotide uptake on follow-up scans, whereas on CT the size of the lesions remained unchanged, possibly indicating tumor necrosis. Patient 3 had a complete remission after PRRT with (177)Lu-DOTATATE but subsequently relapsed with many receptor-negative metastases, requiring intensive chemotherapy. CONCLUSION: Although biopsy is required for initial diagnosis and treatment planning, noninvasive molecular imaging may evolve into in vivo molecular pathology in selected groups of patients, especially in treatment follow-up. PMID- 15653656 TI - Long-term follow-up of renal function after peptide receptor radiation therapy with (90)Y-DOTA(0),Tyr(3)-octreotide and (177)Lu-DOTA(0), Tyr(3)-octreotate. AB - The kidneys are critical organs in peptide receptor radiation therapy (PRRT). Renal function loss may become apparent many years after PRRT. We analyzed the time course of decline in creatinine clearance (CLR) in patients during a follow up of at least 18 mo after the start of PRRT with (90)Y-1,4,7,10 tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid (DOTA),Tyr(3)-octreotide ((90)Y-DOTATOC) or (177)Lu-DOTA(0),Tyr(3)-octreotate ((177)Lu-DOTATATE). METHODS: Twenty-eight patients with metastasized neuroendocrine tumors received 1-5 cycles of (90)Y-DOTATOC, leading to renal radiation doses of 5.9-26.9 Gy per cycle and a total of 18.3-38.7 Gy. Median follow-up was 2.9 y (range, 1.5-5.4 y), with a median of 16 measurements (range, 5-53) per patient. Thirty-seven patients with metastasized neuroendocrine tumors received 3-7 cycles of (177)Lu-DOTATATE, leading to renal radiation doses of 1.8-7.8 Gy per cycle and a total of 7.3-26.7 Gy. Median follow-up was 2.4 y (range, 1.7-4.0 y), with a median of 10 (range, 6 27) measurements per patient. All renal dose estimates were calculated with the MIRDOSE3 model. All patients were infused with renoprotective amino acids during the administration of the radioactive peptides. The time trend of CLR was determined by fitting a monoexponential function through the data of individual patients, yielding the decline in CLR in terms of percentage change per year. RESULTS: The median decline in CLR was 7.3% per y in patients treated with (90)Y DOTATOC and 3.8% per y in patients treated with (177)Lu-DOTATATE (P = 0.06). The time trend of decline in CLR was sustained during the follow-up period. Eleven patients had a >15% per y decline in CLR. Cumulative renal radiation dose, per cycle renal radiation dose, age, hypertension, and diabetes are probable contributing factors to the rate of decline in CLR after PRRT. CONCLUSION: This study showed that the time course of CLR after PRRT was compatible with the pattern of sustained CLR loss in progressive chronic kidney disease. PMID- 15653657 TI - Practical dosimetry of peptide receptor radionuclide therapy with (90)Y-labeled somatostatin analogs. AB - The challenge for internal therapy is to deliver the highest possible dose to the tumor while sparing normal organs from damage. Currently, the potential risk of kidney and red marrow toxicity limits the amount of radioactivity that may be administered. An accurate dosimetry method that would provide reliable dose estimates to these critical organs and to tumors before therapy would allow the clinician to plan a specific therapeutic regimen and also select those patients who would benefit the most from treatment. The dosimetry for (90)Y-1,4,7,10 tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid-d-Phe(1)-Tyr(3)-octreotide is usually based on quantitative imaging at different time points that provides information on activity retention in organs over time and on stylized models representing average individuals. Because the therapeutic agent labeled with (90)Y is not suitable for quantitative imaging, the peptide surrogate labeled with the positron emitter (86)Y can be considered the most appropriate tracer for measuring distribution and retention of the radiopharmaceutical over time. Dose calculations in target organs are generally performed using the MIRDOSE program, in which S values from source to target are integrated. Significant improvement of dose estimates may be achieved by introducing patient-specific adjustments to the standard models. The use of individual kidney volumes assessed by CT instead of the use of a fixed volume for males and females may significantly improve the determination of kidney radiation doses. The use of actual CT-derived tumor volumes has also shown a dose-efficacy relationship. Additional improvements in this field include the validation and use of an (111)In surrogate to avoid the complexity of (86)Y use and the consideration of radiobiologic parameters, such as fractionation effects and the specific biologic efficacy of internally deposited radiation, which are probably underestimated using currently available methods. PMID- 15653658 TI - Patient-specific dosimetry in predicting renal toxicity with (90)Y-DOTATOC: relevance of kidney volume and dose rate in finding a dose-effect relationship. AB - Nephrotoxicity is the major limiting factor during therapy with the radiolabeled somatostatin analog (90)Y-1,4,7,10-tetraazacyclododecane-N,N',N'',N''' tetraacetic acid (DOTA)-d-Phe(1)-Tyr(3)-octreotide (DOTATOC). Pretherapeutic assessment of kidney absorbed dose could help to minimize the risk of renal toxicity. The aim of this study was to evaluate the contribution of patient specific adjustments to the standard dosimetric models, such as the renal volume and dose rate, for estimating renal absorbed dose during therapy with (90)Y DOTATOC. In particular, we investigated the correlation between dose estimates and effect on renal function after therapy. METHODS: Eighteen patients with neuroendocrine tumors (9 men and 9 women; median age, 59 y) underwent treatment with (90)Y-DOTATOC (8.1-22.9 GBq) after pretherapeutic biodistribution study with (86)Y-DOTATOC. Kidney uptake and residence times were measured and the absorbed dose (KAD) was computed using either the MIRDOSE3.1 software assuming a standard kidney volume (KAD(StdVol)) or the MIRD Pamphlet 19 values and the actual kidney cortex volume determined by pretherapeutic CT (KAD(CTVol)). For each patient, the biologic effective dose (BED) was calculated according to the linear quadratic model to take into account the effect of dose rate and fractionation. Renal function was evaluated every 6 mo by serum creatinine and creatinine clearance (CLR) during a median follow-up of 35.5 mo (range, 18-65 mo). The individual rate of decline of renal function was expressed as CLR loss per year. RESULTS: KAD(CTVol) ranged between 19.4 and 39.6 Gy (mean, 28.9 +/- 5.34 Gy). BED, obtained from KAD(CTVol), ranged between 27.7 and 59.3 Gy (mean, 40.4 +/- 10.6 Gy). The CLR loss per year ranged from 0% to 56.4%. In 12 of 18 patients, CLR loss per year was <20%. No correlation was observed between CLR loss per year and the KAD(StdVol) or the KAD(CTVol). In contrast, BED strongly correlated with CLR loss per year (r = 0.93; P < 0.0001). All 5 patients with CLR loss per year >20% received a BED >45 Gy. Patients who were treated with low fractionation were those with the highest rate of renal function impairment. CONCLUSION: Radiation nephrotoxicity after (90)Y-DOTATOC therapy is dose dependent. Individual renal volume, dose rate, and fractionation play important roles in an accurate dosimetry estimation that enables prediction of risk of renal function impairment. PMID- 15653659 TI - Peptide receptor radionuclide therapy for non-radioiodine-avid differentiated thyroid carcinoma. AB - In patients with progressive metastatic (or recurrent) differentiated thyroid carcinoma (DTC) who do not respond to radioiodine therapy or do not show uptake on radioiodine scintigraphy, treatment options are few. Because these tumors may express somatostatin receptors, peptide receptor radionuclide therapy might be effective. We evaluated the therapeutic efficacy of the radiolabeled somatostatin analog (177)Lu-1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid(0) (DOTA), Tyr(3)-octreotate ((177)Lu-DOTATATE) in patients with DTC. The uptake of radioactivity in tumors was also studied in relation to treatment outcome. METHODS: Five patients with DTC (3 with Hurthle cell thyroid carcinoma [HCTC], 1 with papillary thyroid carcinoma [PTC], and 1 with follicular thyroid carcinoma [FTC]) were treated with 22.4-30.1 GBq of (177)Lu-DOTATATE. Response to therapy was evaluated with CT. Uptake on (177)Lu-DOTATATE scintigraphy (24 h after treatment), expressed as percentage of injected dose, was compared with uptake on pretherapy (111)In-octreotide scintigraphy (24 h after injection). RESULTS: After the last treatment with (177)Lu-DOTATATE, 1 patient with HCTC had stable disease as a maximum response, 1 patient with HCTC had minor remission (tumor shrinkage between 25% and 50%), and 1 patient with HCTC had partial remission (shrinkage > or =50%). The responses in PTC and FTC were stable disease and progressive disease, respectively. A decrease in serum thyroglobulin level was found in patients with HCTC. Patients with minor and partial remissions had the highest (177)Lu-DOTATATE-to-(111)In-diethylenetriamine pentaacetic acid(0)-octreotide ((111)In-octreotide) uptake ratios (3.2 and 2.4, respectively) whereas the other patients had uptake ratios smaller than 1.5. CONCLUSION: (177)Lu-DOTATATE therapy can be effective in patients with progressive DTC who have no therapeutic options and sufficient uptake of (111)In-octreotide in tumor lesions as shown on (111)In octreotide scintigraphy. This finding is especially important in patients with HCTC, because they cannot benefit from radioiodine therapy because of non-iodine avid lesions at diagnosis. PMID- 15653660 TI - Perspectives on cancer therapy with radiolabeled monoclonal antibodies. AB - With the approval of 2 radiolabeled antibody products for the treatment of non Hodgkin's lymphoma (NHL), radioimmunotherapy (RIT) has finally come of age as a new therapeutic modality, exemplifying the collaboration of multiple disciplines, including immunology, radiochemistry, radiation medicine, medical oncology, and nuclear medicine. Despite the many challenges that this new therapy discipline has encountered, there is growing evidence that RIT can have a significant impact on the treatment of cancer. Although follicular NHL is currently the only indication in which RIT has been proven to be effective, clinical trials are showing usefulness in other forms of NHL as well as in other hematologic neoplasms. However, the treatment of solid tumors remains a formidable challenge, because the doses shown to be effective in hematologic tumors are insufficient in the more common epithelial cancers. Nevertheless, there has been progress in locoregional applications and in the treatment of minimal residual disease. There is also optimism that pretargeting procedures, including new molecular constructs and targets, will improve the delivery of radioactivity to tumors, do so with less hematologic toxicity, and become the next generation of RIT. PMID- 15653661 TI - Tositumomab and (131)I therapy in non-Hodgkin's lymphoma. AB - Tositumomab and (131)I-tositumomab constitute a relatively new radioimmunotherapeutic regimen for patients with CD20+ follicular non-Hodgkin's lymphoma (NHL). Currently, it is approved for use in patients whose disease has relapsed after chemotherapy and is refactory to rituximab, including patients whose tumors have transformed to a higher histologic grade. This review outlines the current and evolving status of this therapeutic regimen at nonmyeloablative doses. METHODS: Clinical data from multiple published studies and preliminary communications encompassing more than 1,000 patients were reviewed to describe the current status of tositumomab and (131)I-tositumomab therapy. The therapy is delivered in 2 parts, a dosimetric dose and a therapeutic dose. The therapeutic radioactivity millicurie dose is calculated on a patient-individualized ("tailored") basis. A series of 3 total-body gamma-camera scans are used to determine the patient-specific pharmacokinetics (total-body residence time) of the radiolabeled antibody conjugate required to deliver the desired total-body radiation dose, typically 75 cGy. RESULTS: In clinical trials, objective response rates in patients who had been extensively pretreated with chemotherapy ranged from 47% to 68%. Tositumomab and (131)I-tositumomab therapy also was effective in patients who had failed to respond to or who had relapsed after rituximab therapy, with a 68% overall response rate. Thirty percent of such patients achieved complete responses that were generally of several years duration. Single center trials using tositumomab and (131)I-tositumomab therapy alone or after chemotherapy in previously untreated patients have shown response rates in excess of 90%, with most responses complete. Retreatment with tositumomab and (131)I tositumomab and use of lower total-body radiation doses of tositumomab and (131)I tositumomab to treat patients who have relapsed after stem cell transplantation have been shown feasible in limited clinical studies. Toxicity is predominately hematologic; however, human antimouse antibodies, hypothyroidism, and myelodysplastic syndrome have been reported in a small fraction of patients. CONCLUSION: Tositumomab and (131)I-tositumomab therapy at patient-specific, nonmyeloablative doses is safe and effective in treatment of relapsed and refractory follicular NHL. Toxicity is mainly hematologic and reversible. Tositumomab and (131)I-tositumomab therapy is assuming a growing role in this common malignancy. PMID- 15653662 TI - Current status of therapy of solid tumors. AB - The recent approval of 2 radiolabeled antibodies against cluster designation 20 positive lymphoma has led to a resurgence of interest in radioimmunotherapy. As was the case with chemotherapy development, progress has been most marked in the hematologic neoplasms, both in myeloablative and in nonmyeloablative therapeutic strategies. Success in the radioimmunotherapy of solid tumors has lagged because of the immunogenicity of murine proteins and the relatively slow clearance of humanized intact immunoglobulins. Genetic engineering has enabled the development of a variety of antigen-binding constructs of various sizes and immunobiologic characteristics. Developments in radiochemistry as well as production of an increasing number of radionuclides with therapeutic potential or optimal imaging characteristics have spurred tailored therapeutic strategies that include dosimetry and considerations of tumor burden. Such progress has generated pivotal studies that will establish the radiobiologic paradigms for successful radioimmunotherapy of solid tumors. This review will describe seminal studies that have paved the way to an understanding of radioimmunotherapy in solid tumors. Finally, the authors' views of the future of this promising cancer therapy will be presented. PMID- 15653663 TI - Current status of therapy of solid tumors: brain tumor therapy. AB - Treatment of malignant brain tumors with conventional approaches is largely unsuccessful because curative doses generally cannot be delivered without excessive toxicity to normal brain. Radioimmunotherapy is emerging as an attractive alternative for glioma therapy because of the potential for more selectively irradiating tumor cells while sparing normal tissues. Several institutions are engaged in phase I and phase II trials investigating the therapeutic potential of monoclonal antibodies (mAbs) labeled with the beta emitters (131)I and (90)Y and the alpha-emitter (211)At in patients with recurrent and newly diagnosed brain tumors. The current status of these trials will be discussed with regard to efficacy, toxicity, and future directions. PMID- 15653664 TI - Antibody-based targeted radiation to pediatric tumors. AB - Radioimmunotherapy (RIT) for pediatric tumors remains in its infancy despite its potential as an attractive therapeutic modality. Most childhood tumors are radiation sensitive, but the side effects of external beam radiation are well recognized. Despite achieving complete remissions with sophisticated combination therapies, treatment failure primarily results from the inability to eradicate minimal residual disease, which is typically distant and occult. RIT can conceivably target such disease and improve cancer treatment. Because intensive chemotherapy regimens used in most childhood cancers are highly immunosuppressive, repeated administration of radiolabeled monoclonal antibodies is possible without the immediate induction of human antimouse or human antichimeric antibody responses. Despite the differences in biology between childhood and adult hematologic malignancies, they share several tumor antigens for which RIT agents are now available. However, safety and efficacy profiles in children remain to be defined. On the other hand, the antigen repertoire of pediatric solid tumors differs substantially from that in adults, partly because of differing lineages: pediatric solid tumors are typically of embryonal origin, whereas adult tumors are usually carcinomas of epithelial origin. Hence, RIT agents licensed for adult tumors are generally not applicable to pediatric solid tumors. Tumor-selective radioimmunoconjugates specific for embryonal tumors of childhood are currently being actively investigated. Without substantial policy changes in drug development for orphan indications, however, these agents are not likely to be widely available in the near future. PMID- 15653665 TI - The promise of immuno-PET in radioimmunotherapy. AB - Immuno-PET as a quantitative imaging procedure before or concomitant with radioimmunotherapy is an attractive option to improve confirmation of tumor targeting and especially assessment of radiation dose delivery to both tumor and normal tissues. General information about PET, PET systems, and quantification is provided in this review. The requirements for an appropriate positron emitter and characteristics of the most attractive candidate emitters for immuno-PET are discussed. An overview of preclinical and clinical immuno-PET studies reported in the literature is provided. PMID- 15653666 TI - (68)Ga-labeled peptides in tumor imaging. AB - Radiolabeled peptides are of increasing interest in nuclear oncology. Special emphasis has been given to the development of peptides labeled with positron emitters. Among these, (68)Ga deserves special attention, because it is available from an inhouse generator rendering (68)Ga radiopharmacy independent of an onsite cyclotron. (68)Ga has a half-life of 68 min and decays by 89% through positron emission. The parent, (68)Ge, is accelerator produced and decays with a half-life of 270.8 d by electron capture. Currently, at least 1 commercial and several in house generators are available. (68)Ge is strongly absorbed on metal oxides or organic material, making a (68)Ge-breakthrough highly unlikely. Several groups continue to further develop generators to remove cationic impurities from the eluate. Several bifunctional chelators based on 1,4,7-triazacyclononane-N,N',N'' triacetic acid and 1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid (DOTA) macrocycles are available for coupling to peptides and other biomolecules. In addition to these hydrophilic chelators, a lipophilic tetradentate S(3)N ligand was developed. Radiopeptides for (68)Ga labeling have been developed and tested preclinically for the targeting of somatostatin receptors, the melanocortin 1 receptor, and the bombesin receptor. Clinical studies were performed with (68)Ga-DOTA,Tyr(3)-octreotide, localizing neuroendocrine tumors with higher sensitivity than (111)In-diethylenetriaminepentaacetic acid octreotide. In addition, (68)Ga-DOTA-based bombesin derivatives are being investigated with some success in patients with prostate cancer. CONCLUSION: Generator-produced (68)Ga and the development of small chelator-coupled peptides (and other small biomolecules) may open a new generation of freeze-dried, good manufacturing practice-produced, kit-formulated PET radiopharmaceuticals similar to (99)Mo-/(99m)Tc-generator-based, (99m)Tc-labeled radiopharmaceuticals. PMID- 15653667 TI - Radiotargeted gene therapy. AB - The radiotargeted gene therapy approach to localizing radionuclides at tumor sites involves inducing tumor cells to synthesize a membrane-expressed receptor with a high affinity for injected radiolabeled ligands. A second strategy involves transduction of the sodium iodide symporter (NIS) and free radionuclide therapy. Using the first strategy, induction of high levels of human somatostatin receptor subtype 2 expression and selective tumor uptake, imaging, or growth inhibition with radiolabeled somatostatin analogs has been achieved in human tumor xenograft models. Therapy studies have been performed on several tumor xenograft models with various radionuclides using the NIS radiotargeted gene therapy approach. The use of gene transfer technology to induce expression of high-affinity membrane receptors or transporters can enhance the specificity and extent of radioligand or radionuclide localization in tumors, and the use of radionuclides with appropriate emissions can deliver radiation-absorbed cytotoxic doses across several cell diameters to compensate for limited transduction efficiency. Clinical studies are needed to determine the most promising of these new therapeutic approaches. PMID- 15653668 TI - Radiation sensitizers: a selective review of molecules targeting DNA and non-DNA targets. AB - The ideal radiation sensitizer would reach the tumor in adequate concentrations and act selectively in the tumor compared with normal tissue. It would have predictable pharmacokinetics for timing with radiation treatment and could be administered with every radiation treatment. The ideal radiation sensitizer would have minimal toxicity itself and minimal or manageable enhancement of radiation toxicity. The ideal radiation sensitizer does not exist today. This review outlines the concept of combining 2 modalities of cancer treatment, radiation and drug therapy, to provide enhanced tumor cell kill in the treatment of human malignancies and discusses molecules that target DNA and non-DNA targets. Combining drugs that have unique mechanisms of action and absence of overlapping toxicities with systemically administered radiotherapy should be exploited in future clinical trials. This is an exciting time in clinical oncology research, because we have a plethora of new molecules to evaluate. PMID- 15653669 TI - Induction of apoptosis with hybrids of Arg-Gly-Asp molecules and peptides and antimitotic effects of hybrids of cytostatic drugs and peptides. AB - The presence of a high density of somatostatin receptors (SSRs) on human tumors forms the basis for the successful visualization of primary tumors and their metastases using radiolabeled somatostatin analogs. In recent years somatostatin analogs, coupled to beta-emitting radioisotopes, have been successfully applied in the treatment of patients with metastatic SSR-positive neuroendocrine tumors. This concept of targeting SSR-expressing tumors using peptide receptor radionuclide therapy may also apply to the use of somatostatin analogs coupled to chemotherapeutic compounds. Evidence for the effectiveness of such cytotoxic somatostatin analogs as antitumor agents has been provided in a significant number of studies in experimental tumor models. In addition to cytotoxic somatostatin analogs, somatostatin analogs coupled to peptides containing arginine, glycine, and aspartate and coupled to paclitaxel have been synthesized. Here we discuss the development of the different cytotoxic somatostatin analogs and their antitumor effects in vitro and in vivo in experimental models. PMID- 15653670 TI - The promise of targeted {alpha}-particle therapy. AB - The use of monoclonal antibodies to deliver radioisotopes directly to tumor cells has become a promising strategy to enhance the antitumor effects of native antibodies. Since the alpha- and beta-particles emitted during the decay of radioisotopes differ in significant ways, proper selection of isotope and antibody combinations is crucial to making radioimmunotherapy a standard therapeutic modality. Because of the short pathlength (50-80 microm) and high linear energy transfer ( approximately 100 keV/microm) of alpha-emitting radioisotopes, targeted alpha-particle therapy offers the potential for more specific tumor cell killing with less damage to surrounding normal tissues than beta-emitters. These properties make targeted alpha-particle therapy ideal for the elimination of minimal residual or micrometastatic disease. Radioimmunotherapy using alpha-emitters such as (213)Bi, (211)At, and (225)Ac has shown activity in several in vitro and in vivo experimental models. Clinical trials have demonstrated the safety, feasibility, and activity of targeted alpha particle therapy in the treatment of small-volume and cytoreduced disease. Further advances will require investigation of more potent isotopes, new sources and methods of isotope production, improved chelation techniques, better methods for pharmacokinetic and dosimetric modeling, and new methods of isotope delivery such as pretargeting. Treatment of patients with less-advanced disease and, ultimately, randomized trials comparing targeted alpha-particle therapy with standard approaches will be required to determine the clinical utility of this approach. PMID- 15653671 TI - O-fucosylation of notch occurs in the endoplasmic reticulum. AB - LADII (leukocyte adhesion deficiency type II)/CDGIIc (congenital disorder of glycosylation type IIc) is a rare autosomal recessive disease characterized by leukocyte adhesion deficiency as well as severe neurological and developmental abnormalities. It is caused by mutations in the Golgi GDP-fucose transporter, resulting in a reduction of fucosylated antigens on the cell surface. A recent study using fibroblasts from LADII/CDGIIc patients suggested that although terminal fucosylation of N-glycans is reduced severely, protein O-fucosylation is generally unaffected (Sturla, L., Rampal, R., Haltiwanger, R. S., Fruscione, F., Etzioni, A., and Tonetti, M. (2003) J. Biol. Chem. 278, 26727-26733). A potential explanation for this phenomenon is that enzymes adding O-fucose to proteins localize to cell organelles other than the Golgi apparatus. In this study, we investigated the subcellular localization of protein O-fucosyltransferase 1 (O FucT-1), which is responsible for adding O-fucose to epidermal growth factor-like repeats. Our analysis reveals that, unlike all other known fucosyltransferases, O FucT-1 is a soluble protein that localizes to the endoplasmic reticulum (ER). In addition, it appears that O-FucT-1 is retained in the ER by a KDEL-like sequence at its C terminus. Our results also suggest that enzymatic addition of O-fucose to proteins occurs in the ER, suggesting that a novel, ER-localized GDP-fucose transporter may exist. The fact that O-FucT-1 recognizes properly folded epidermal growth factor-like repeats, together with this unique localization, suggests that it may play a role in quality control. PMID- 15653672 TI - Enzymatic properties of purified murine fatty acid transport protein 4 and analysis of acyl-CoA synthetase activities in tissues from FATP4 null mice. AB - Fatty acid transport protein 4 (FATP4) is an integral membrane protein expressed in the plasma and internal membranes of the small intestine and adipocyte as well as in the brain, kidney, liver, skin, and heart. FATP4 has been hypothesized to be bifunctional, exhibiting both fatty acid transport and acyl-CoA synthetase activities that work in concert to mediate fatty acid influx across biological membranes. To determine whether FATP4 is an acyl-CoA synthetase, the murine protein was engineered to contain a C-terminal FLAG epitope tag, expressed in COS1 cells via adenovirus-mediated infection and purified to near homogeneity using alpha-FLAG affinity chromatography. Kinetic analysis of the enzyme was carried out for long chain (palmitic acid, C16:0) and very long chain (lignoceric acid, C24:0) fatty acids as well as for ATP and CoA. FATP4 exhibited substrate specificity for C16:0 and C24:0 fatty acids with a V(max)/K(m) (C16:0)/V(max)/K(m) (C24:0) of 1.5. Like purified FATP1, FATP4 was insensitive to inhibition by triacsin C but was sensitive to feedback inhibition by acyl-CoA. Although purified FATP4 exhibited high levels of palmitoyl-CoA and lignoceroyl CoA synthetase activity, extracts from the skin and intestine of FATP4 null mice exhibited reduced esterification for C24:0, but not C16:0 or C18:1, suggesting that in vivo, defects in very long chain fatty acid uptake may underlie the skin disorder phenotype of null mice. PMID- 15653673 TI - Counteracting osmolyte trimethylamine N-oxide destabilizes proteins at pH below its pKa. Measurements of thermodynamic parameters of proteins in the presence and absence of trimethylamine N-oxide. AB - Earlier studies have reported that trimethylamine N-oxide (TMAO), a naturally occurring osmolyte, is a universal stabilizer of proteins because it folds unstructured proteins and counteracts the deleterious effects of urea and salts on the structure and function of proteins. This conclusion has been reached from the studies of the effect of TMAO on proteins in the pH range 6.0-8.0. In this pH range TMAO is almost neutral (zwitterionic form), for it has a pK(a) of 4.66 +/- 0.10. We have asked the question of whether the effect of TMAO on protein stability is pH-dependent. To answer this question we have carried out thermal denaturation studies of lysozyme, ribonuclease-A, and apo-alpha-lactalbumin in the presence of various TMAO concentrations at different pH values above and below the pK(a) of TMAO. The main conclusion of this study is that near room temperature TMAO destabilizes proteins at pH values below its pK(a), whereas it stabilizes proteins at pH values above its pK(a). This conclusion was reached by determining the T(m) (midpoint of denaturation), delta H(m) (denaturational enthalpy change at T(m)), delta C(p) (constant pressure heat capacity change), and delta G(D) degrees (denaturational Gibbs energy change at 25 degrees C) of proteins in the presence of different TMAO concentrations. Other conclusions of this study are that T(m) and delta G(D) degrees depend on TMAO concentration at each pH value and that delta H(m) and the delta C(p) are not significantly changed in presence of TMAO. PMID- 15653674 TI - SOCS-3 induces myoblast differentiation. AB - Myoblast differentiation is characterized by a sequence of events that includes an increase in insulin-like growth factor (IGF)-I and contractile gene expression. The increase in IGF-I expression activates cell signaling mechanisms that participate in the differentiation process. One potential contributor is the SOCS-3 (suppressor of cytokine signaling-3) gene, which regulates signaling mechanisms and may be sensitive to changes in IGF-I concentrations. For the first time, the role of SOCS-3 is investigated in myoblast differentiation. SOCS-3 mRNA levels and SOCS-3 transcriptional activity increase during myoblast differentiation. SOCS-3 gene expression is induced, at least in part, by activation of the IGF-I receptor during myoblast differentiation. Overexpression of SOCS-3 cDNA significantly increased transcriptional activation of the 2.0-kb skeletal alpha-actin promoter in differentiating C2C12 myoblasts. In addition, overexpression of SOCS-3 specifically increased serum response factor-driven transcriptional activity but had no effect on nuclear-factor of activated T cell driven transcriptional activity. SOCS-3 overexpression induced skeletal alpha actin transcription in a myoblast cell line that cannot respond to endogenous IGF I, indicating that SOCS-3 can contribute to the myoblast differentiation process in the absence of IGF-I. These data suggest that IGF-I induces myoblast differentiation, in part, by increasing SOCS-3 expression. PMID- 15653675 TI - CSX/Nkx2.5 modulates differentiation of skeletal myoblasts and promotes differentiation into neuronal cells in vitro. AB - CSX/Nkx2.5 transcription factor plays a pivotal role in cardiac development; however, its role in development and differentiation of other organs has not been investigated. In this study, we used C2C12 myoblasts and human fetal primary myoblasts to investigate the function of Nkx2.5 in skeletal myogenesis. The expression levels of Nkx2.5 decreased as C2C12 myoblasts elongated and fused to form myotubes. The expression of human NKX2.5 in C2C12 myoblasts inhibited myocyte differentiation and myotube formation, and up-regulated Gata4 and Tbx5 expression. The expression of NKX2.5 in terminally differentiated C2C12 myotubes resulted in a change in morphology and breakdown into smaller myotubes. Furthermore, overexpression of NKX2.5 in C2C12 cells and primary cultures of human fetal myoblasts led to differentiation of myoblasts into neuron-like cells and expression of neuronal markers. This study sheds light on the previously unknown non-cardiac functions of Nkx2.5 transcription factor. PMID- 15653676 TI - Mitotic kinesin inhibitors induce mitotic arrest and cell death in Taxol resistant and -sensitive cancer cells. AB - Taxanes are powerful chemotherapy agents that target the microtubule cytoskeleton, leading to mitotic arrest and cell death; however, their clinical efficacy has been hampered due to the development of drug resistance. Therefore, other proteins involved in spindle assembly are being examined as potential targets for anticancer therapy. The mitotic kinesin, Eg5 is critical for proper spindle assembly; as such, inhibition of Eg5 leads to mitotic arrest making it a potential anticancer target. We wanted to validate Eg5 as a therapeutic target and determine if Eg5 inhibitors retain activity in Taxol-resistant cells. Using affinity chromatography we first show that the compound HR22C16 is an Eg5 inhibitor and does not interact with other microtubule motor proteins tested. Furthermore, HR22C16 along with its analogs, inhibit cell survival in both Taxol sensitive and -resistant ovarian cancer cells with at least 15-fold greater efficacy than monastrol, the first generation Eg5 inhibitor. Further analysis with HR22C16-A1, the most potent HR22C16 analog, showed that it retains efficacy in PgP-overexpressing cells, suggesting that it is not a PgP substrate. We further show that HR22C16-A1 induces cell death following mitotic arrest via the intrinsic apoptotic pathway. Interestingly, the combination of HR22C16-A1 with Taxol results in an antagonistic antiproliferative and antimitotic effect, possibly due to the abrogation of Taxol-induced mitotic spindles by HR22C16-A1. Taken together, our results show that Eg5 inhibitors have promising anticancer activity and can be potentially used to overcome Taxol resistance in the clinical setting. PMID- 15653677 TI - Crystal structures of apo-form and binary/ternary complexes of Podophyllum secoisolariciresinol dehydrogenase, an enzyme involved in formation of health protecting and plant defense lignans. AB - (-)-Matairesinol is a central biosynthetic intermediate to numerous 8-8'-lignans, including the antiviral agent podophyllotoxin in Podophyllum species and its semi synthetic anticancer derivatives teniposide, etoposide, and Etopophos. It is formed by action of an enantiospecific secoisolariciresinol dehydrogenase, an NAD(H)-dependent oxidoreductase that catalyzes the conversion of (-) secoisolariciresinol. Matairesinol is also a plant-derived precursor of the cancer-preventative "mammalian" lignan or "phytoestrogen" enterolactone, formed in the gut following ingestion of high fiber dietary foodstuffs, for example. Additionally, secoisolariciresinol dehydrogenase is involved in pathways to important plant defense molecules, such as plicatic acid in the western red cedar (Thuja plicata) heartwood. To understand the molecular and enantiospecific basis of Podophyllum secoisolariciresinol dehydrogenase, crystal structures of the apo form and binary/ternary complexes were determined at 1.6, 2.8, and 2.0 angstrom resolution, respectively. The enzyme is a homotetramer, consisting of an alpha/beta single domain monomer containing seven parallel beta-strands flanked by eight alpha-helices on both sides. Its overall monomeric structure is similar to that of NAD(H)-dependent short-chain dehydrogenases/reductases, with a conserved Asp47 forming a hydrogen bond with both hydroxyl groups of the adenine ribose of NAD(H), and thus specificity toward NAD(H) instead of NADP(H). The highly conserved catalytic triad (Ser153, Tyr167, and Lys171) is adjacent to both NAD(+) and substrate molecules, where Tyr167 functions as a general base. Following analysis of high resolution structures of the apo-form and two complex forms, the molecular basis for both the enantio-specificity and the reaction mechanism of secoisolariciresinol dehydrogenase is discussed and compared with that of pinoresinol-lariciresinol reductase. PMID- 15653678 TI - Activity of hypoxia-inducible factor 2alpha is regulated by association with the NF-kappaB essential modulator. AB - The hypoxia-inducible factors 1alpha (HIF-1alpha) and 2alpha (HIF-2alpha) are key regulators of the transcriptional response to low oxygen and are closely related in domain architecture, DNA binding, and activation mechanisms. Despite these similarities, targeted disruption of the HIF-alpha genes in mice results in distinctly different phenotypes demonstrating nonredundancy of function, although the underlying mechanisms remain unclear. Here we report on the novel and specific interaction of HIF-2alpha, but not HIF-1alpha, with the NF-kappaB essential modulator (NEMO) using immunoprecipitation, mammalian two-hybrid, and in vitro protein interaction assays. Reporter gene assays demonstrate that this interaction specifically enhances normoxic HIF-2alpha transcriptional activity, independently of the HIF-2alpha transactivation domain, consistent with a model by which NEMO aids CBP/p300 recruitment to HIF-2alpha. In contrast, HIF-2alpha overexpression does not alter NF-kappaB signaling, suggesting that the functional consequence of the HIF-2alpha/NEMO interaction is limited to the HIF pathway. The specificity of NEMO for HIF-2alpha represents one of the few known differential protein-protein interactions between the HIF-alpha proteins, which has important implications for the activity of HIF-2alpha and is also the first postulated NF kappaB-independent role for NEMO. PMID- 15653679 TI - Probing the mechanism of the Archaeoglobus fulgidus inositol-1-phosphate synthase. AB - myo-Inositol-1-phosphate synthase (mIPS) catalyzes the conversion of glucose-6 phosphate (G-6-P) to inositol-1-phosphate. In the sulfate-reducing archaeon Archaeoglobus fulgidus it is a metal-dependent thermozyme that catalyzes the first step in the biosynthetic pathway of the unusual osmolyte di-myo-inositol 1,1'-phosphate. Several site-specific mutants of the archaeal mIPS were prepared and characterized to probe the details of the catalytic mechanism that was suggested by the recently solved crystal structure and by the comparison to the yeast mIPS. Six charged residues in the active site (Asp225, Lys274, Lys278, Lys306, Asp332, and Lys367) and two noncharged residues (Asn255 and Leu257) have been changed to alanine. The charged residues are located at the active site and were proposed to play binding and/or direct catalytic roles, whereas noncharged residues are likely to be involved in proper binding of the substrate. Kinetic studies showed that only N255A retains any measurable activity, whereas two other mutants, K306A and D332A, can carry out the initial oxidation of G-6-P and reduction of NAD+ to NADH. The rest of the mutant enzymes show major changes in binding of G-6-P (monitored by the 31P line width of inorganic phosphate when G-6 P is added in the presence of EDTA) or NAD+ (detected via changes in the protein intrinsic fluorescence). Characterization of these mutants provides new twists on the catalytic mechanism previously proposed for this enzyme. PMID- 15653680 TI - SIRT3, a mitochondrial sirtuin deacetylase, regulates mitochondrial function and thermogenesis in brown adipocytes. AB - SIRT3 is one of the seven mammalian sirtuin homologs of the yeast Sir2 gene, which mediates the effect of caloric restriction on life span extension in yeast and Caenorhabditis elegans. Because adipose tissue is essential in energy homeostasis and also plays a role in life span determination, we decided to investigate the function of sirtuin members in fat. We report here that murine SIRT3 is expressed in brown adipose tissue and is localized on the mitochondria inner membrane. Caloric restriction activates SIRT3 expression in both white and brown adipose. Additionally, cold exposure up-regulates SIRT3 expression in brown fat, whereas elevated climate temperature reduces the expression. Enforced expression of SIRT3 in the HIB1B brown adipocytes enhances the expression of the uncoupling protein PGC-1alpha, UCP1, and a series of mitochondria-related genes. Both ADP-ribosyltransferase and deacetylase activities of SIRT3 are required for this action. Furthermore, the SIRT3 deacetylase mutant exhibits a dominant negative effect by inhibiting UCP1 expression. This inhibitive effect can be abolished by the coexpression of PGC-1alpha, indicating a major role of PGC 1alpha in the SIRT3 action. In addition, SIRT3 stimulates CREB phosphorylation, which reportedly activates PGC-1alpha promoter directly. Functionally, sustained expression of SIRT3 decreases membrane potential and reactive oxygen species production while increasing cellular respiration. Finally, SIRT3, along with genes related to mitochondrial function, is down-regulated in the brown adipose tissue of several genetically obese mice. In summary, our results demonstrate that SIRT3 activates mitochondria functions and plays an important role in adaptive thermogenesis in brown adipose. PMID- 15653681 TI - Androgen receptor activation by G(s) signaling in prostate cancer cells. AB - The androgen receptor (AR) is activated in prostate cancer patients undergoing androgen ablative therapy and mediates growth of androgen-insensitive prostate cancer cells, suggesting it is activated by nonandrogenic factors. We demonstrate that activated alpha subunit of heterotrimeric guanine nucleotide-binding G(s) protein activates the AR in prostate cancer cells and also synergizes with low concentration of androgen to more fully activate the AR. The G alpha(s) activates protein kinase A, which is required for the nuclear partition and activation of AR. These data suggest a role for G alpha(s) and PKA in the transactivation of AR in prostate cancer cells under the environment of reduced androgen levels. PMID- 15653683 TI - ATP-dependent and ATP-independent roles for the Rad54 chromatin remodeling enzyme during recombinational repair of a DNA double strand break. AB - The efficient and accurate repair of DNA double strand breaks (DSBs) is critical to cell survival, and defects in this process can lead to genome instability and cancers. In eukaryotes, the Rad52 group of proteins dictates the repair of DSBs by the error-free process of homologous recombination (HR). A critical step in eukaryotic HR is the formation of the initial Rad51-single-stranded DNA presynaptic nucleoprotein filament. This presynaptic filament participates in a homology search process that leads to the formation of a DNA joint molecule and recombinational repair of the DSB. Recently, we showed that the Rad54 protein functions as a mediator of Rad51 binding to single-stranded DNA, and here, we find that this activity does not require ATP hydrolysis. We also identify a novel Rad54-dependent chromatin remodeling event that occurs in vivo during the DNA strand invasion step of HR. This ATP-dependent remodeling activity of Rad54 appears to control subsequent steps in the HR process. PMID- 15653682 TI - DNA lesion-specific co-localization of the Mre11/Rad50/Nbs1 (MRN) complex and replication protein A (RPA) to repair foci. AB - The DNA damage response, triggered by DNA replication stress or DNA damage, involves the activation of DNA repair and cell cycle regulatory proteins including the MRN (Mre11, Rad50, and Nbs1) complex and replication protein A (RPA). The induction of replication stress by hydroxyurea (HU) or DNA damage by camptothecin (CAMPT), etoposide (ETOP), or mitomycin C (MMC) led to the formation of nuclear foci containing phosphorylated Nbs1. HU and CAMPT treatment also led to the formation of RPA foci that co-localized with phospho-Nbs1 foci. After ETOP treatment, phospho-Nbs1 and RPA foci were detected but not within the same cell. MMC treatment resulted in phospho-Nbs1 foci formation in the absence of RPA foci. Consistent with the presence or absence of RPA foci, RPA hyperphosphorylation was present following HU, CAMPT, and ETOP treatment but absent following MMC treatment. The lack of co-localization of phospho-Nbs1 and RPA foci may be due to relatively shorter stretches of single-stranded DNA generated following ETOP and MMC treatment. These data suggest that, even though the MRN complex and RPA can interact, their interaction may be limited to responses to specific types of lesions, particularly those that have longer stretches of single-stranded DNA. In addition, the consistent formation of phospho-Nbs1 foci in all of the treatment groups suggests that the MRN complex may play a more universal role in the recognition and response to DNA lesions of all types, whereas the role of RPA may be limited to certain subsets of lesions. PMID- 15653684 TI - Novel poly-GalNAcbeta1-4GlcNAc (LacdiNAc) and fucosylated poly-LacdiNAc N-glycans from mammalian cells expressing beta1,4-N-acetylgalactosaminyltransferase and alpha1,3-fucosyltransferase. AB - Glycans containing the GalNAcbeta1-4GlcNAc (LacdiNAc or LDN) motif are expressed by many invertebrates, but this motif also occurs in vertebrates and is found on several mammalian glycoprotein hormones. This motif contrasts with the more commonly occurring Galbeta1-4GlcNAc (LacNAc or LN) motif. To better understand LDN biosynthesis and regulation, we stably expressed the cDNA encoding the Caenorhabditis elegans beta1,4-N-acetylgalactosaminyltransferase (GalNAcT), which generates LDN in vitro, in Chinese hamster ovary (CHO) Lec8 cells, to establish L8-GalNAcT CHO cells. The glycan structures from these cells were determined by mass spectrometry and linkage analysis. The L8-GalNAcT cell line produces complex type N-glycans quantitatively bearing LDN structures on their antennae. Unexpectedly, most of these complex-type N-glycans contain novel "poly-LDN" structures consisting of repeating LDN motifs (-3GalNAcbeta1-4GlcNAcbeta1-)n. These novel structures are in contrast to the well known poly-LN structures consisting of repeating LN motifs (-3Galbeta1-4GlcNAcbeta1-)n. We also stably expressed human alpha1,3-fucosyltransferase IX in the L8-GalNAcT cells to establish a new cell line, L8-GalNAcT-FucT. These cells produce complex-type N glycans with alpha1,3-fucosylated LDN (LDNF) GalNAcbeta1-4(Fucalpha1 3)GlcNAcbeta1-R as well as novel "poly-LDNF" structures (-3GalNAcbeta1-4(Fucalpha 1-3)GlcNAcbeta1-)n. The ability of these cell lines to generate glycoprotein hormones with LDN-containing N-glycans was studied by expressing a recombinant form of the common alpha-subunit in L8-GalNAcT cells. The alpha-subunit N-glycans carried LDN structures, which were further modified by co-expression of the human GalNAc 4-sulfotransferase I, which generates SO4-4GalNAcbeta1-4GlcNAc-R. Thus, the generation of these stable mammalian cells will facilitate future studies on the biological activities and properties of LDN-related structures in glycoproteins. PMID- 15653685 TI - HIV-1 Nef disrupts antigen presentation early in the secretory pathway. AB - Human immunodeficiency virus, type 1 Nef disrupts viral antigen presentation and promotes viral immune evasion from cytotoxic T lymphocytes. There is evidence that Nef acts early in the secretory pathway to redirect major histocompatibility complex class I (MHC-I) from the trans-Golgi network to the endolysosomal pathway. However, a competing model suggests that Nef acts much later by accelerating MHC-I turnover at the cell surface. Here we demonstrate that Nef targets early forms of MHC-I molecules in the endoplasmic reticulum by preferentially binding hypophosphorylated cytoplasmic tails. The Nef-MHC-I complex migrates normally into the Golgi apparatus but subsequently fails to arrive at the cell surface and become phosphorylated. Cell type-specific differences in the rate of MHC-I transport through the secretory pathway correlate with responsiveness to Nef and co-precipitation of adaptor protein 1 with the Nef.MHC-I complex. We propose that the assembly of a Nef.MHC-I.adaptor protein 1 complex early in the secretory pathway is important for Nef activity. PMID- 15653686 TI - The small heat shock protein alpha B-crystallin is a novel inhibitor of TRAIL induced apoptosis that suppresses the activation of caspase-3. AB - Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a member of the tumor necrosis factor alpha family of cytokines that preferentially induces apoptosis in transformed cells, making it a promising cancer therapy. However, many neoplasms are resistant to TRAIL-induced apoptosis by mechanisms that are poorly understood. We demonstrate that the expression of the small heat shock protein alpha B-crystallin (but not other heat shock proteins or apoptosis regulating proteins) correlates with TRAIL resistance in a panel of human cancer cell lines. Stable expression of wild-type alpha B-crystallin, but not a pseudophosphorylation mutant impaired in its assembly and chaperone function, protects cancer cells from TRAIL-induced caspase-3 activation and apoptosis in vitro. Furthermore, selective inhibition of alpha B-crystallin expression by RNA interference sensitizes cancer cells to TRAIL. In addition, wild-type alpha B crystallin promotes xenograft tumor growth and inhibits TRAIL-induced apoptosis in vivo in nude mice, whereas a pseudophosphorylation alpha B-crystallin mutant impaired in its anti-apoptotic function inhibits xenograft tumor growth. Collectively, these findings indicate that alpha B-crystallin is a novel regulator of TRAIL-induced apoptosis and tumor growth. Moreover, these results demonstrate that targeted inhibition of alpha B-crystallin promotes TRAIL-induced apoptosis, thereby suggesting a novel strategy to overcome TRAIL resistance in cancer. PMID- 15653687 TI - Structure/function analysis of alpha2A-adrenergic receptor interaction with G protein-coupled receptor kinase 2. AB - G protein-coupled receptors (GPCRs) mediate the ability of a diverse array of extracellular stimuli to control intracellular signaling. Many GPCRs are phosphorylated by G protein-coupled receptor kinases (GRKs), a process that mediates agonist-specific desensitization in many cells. Although GRK binding to activated GPCRs results in kinase activation and receptor phosphorylation, relatively little is known about the mechanism of GRK/GPCR interaction or how this interaction results in kinase activation. Here, we used the alpha2A adrenergic receptor (alpha(2A)AR) as a model to study GRK/receptor interaction because GRK2 phosphorylation of four adjacent serines within the large third intracellular loop of this receptor is known to mediate desensitization. Various domains of the alpha(2A)AR were expressed as glutathione S-transferase fusion proteins and tested for the ability to bind purified GRK2. The second and third intracellular loops of the alpha(2A)AR directly interacted with GRK2, whereas the first intracellular loop and C-terminal domain did not. Truncation mutagenesis identified three discrete regions within the third loop that contributed to GRK2 binding, the membrane proximal N- and C-terminal regions as well as a central region adjacent to the phosphorylation sites. Site-directed mutagenesis revealed a critical role for specific basic residues within these regions in mediating GRK2 interaction with the alpha(2A)AR. Mutation of these residues within the holo alpha(2A)AR diminished GRK2-promoted phosphorylation of the receptor as well as the ability of the kinase to be activated by receptor binding. These studies provide new insight into the mechanism of interaction and activation of GRK2 by GPCRs and suggest that GRK2 binding is critical not only for receptor phosphorylation but also for full activity of the kinase. PMID- 15653688 TI - Distinct conformations of the corticotropin releasing factor type 1 receptor adopted following agonist and antagonist binding are differentially regulated. AB - The corticotropin releasing factor (CRF) type 1 receptor (CRF1) is a class B family G protein-coupled receptor that regulates the hypothalamic-pituitary adrenal stress axis. Astressin is an amino-terminal truncated analog of CRF that retains high affinity binding to the extracellular domain of the receptor and is believed to act as a neutral competitive antagonist of receptor activation. Here we show that despite being unable to activate the CRF1 receptor, astressin binding results in the internalization of the receptor. Furthermore, entirely different pathways of internalization of CRF1 receptors are utilized following CRF and astressin binding. CRF causes the receptor to be phosphorylated, recruit beta-arrestin2, and to be internalized rapidly, likely through clathrin-coated pits. Astressin, however, fails to induce receptor phosphorylation or beta arrestin2 recruitment, and internalization is slow and occurs through a pathway that is insensitive to inhibitors of clathrin-coated pits and caveolae. The fate of the internalized receptors also differs because only CRF-induced internalization results in receptor down-regulation. Furthermore, we present evidence that for astressin to induce internalization it must interact with both the extracellular amino terminus and the juxtamembrane domain of the receptor. Astressin binds with 6-fold higher affinity to full-length CRF1 receptors than to a chimeric protein containing only the extracellular domain attached to the transmembrane domain of the activin IIB receptor, yet two 12-residue analogs of astressin have similar affinities for both proteins but are unable to induce receptor internalization. These data demonstrate that agonists and antagonists for CRF1 receptors promote distinct conformations, which are then differentially regulated. PMID- 15653690 TI - Characterization of the oligomer structure of recombinant human mannan-binding lectin. AB - Mannan-binding lectin (MBL) belongs to a family of proteins called the collectins, which show large differences in their ultrastructures. These differences are believed to be determined by different N-terminal disulfide bonding patterns. So far only the bonding pattern of two of the simple forms (recombinant rat MBL-C and bovine CL-43) have been determined. Recombinant MBL expressed in human cells was purified, and the structure of the N-terminal region was determined. Preliminary results on human plasma-derived MBL revealed high similarity to the recombinant protein. Here we report the structure of the N terminal part of recombinant human MBL and present a model to explain the oligomerization pattern. Using a strategy of consecutive enzymatic digestions and matrix-assisted laser desorption ionization mass spectrometry, we succeeded in identifying a number of disulfide-linked peptides from the N-terminal cysteine rich region. Based on these building blocks, we propose a model that can explain the various oligomeric forms found in purified MBL preparations. Furthermore, the model was challenged by the production of cysteine to serine mutants of the three N-terminally situated cysteines. The oligomerization patterns of these mutants support the proposed model. The model indicates that the polypeptide dimer is the basic unit in the oligomerization. PMID- 15653689 TI - Transduction of the scorpion toxin maurocalcine into cells. Evidence that the toxin crosses the plasma membrane. AB - Maurocalcine (MCa) is a 33-amino-acid residue peptide toxin isolated from the scorpion Scorpio maurus palmatus. External application of MCa to cultured myotubes is known to produce Ca2+ release from intracellular stores. MCa binds directly to the skeletal muscle isoform of the ryanodine receptor, an intracellular channel target of the endoplasmic reticulum, and induces long lasting channel openings in a mode of smaller conductance. Here we investigated the way MCa proceeds to cross biological membranes to reach its target. A biotinylated derivative of MCa was produced (MCa(b)) and complexed with a fluorescent indicator (streptavidine-cyanine 3) to follow the cell penetration of the toxin. The toxin complex efficiently penetrated into various cell types without requiring metabolic energy (low temperature) or implicating an endocytosis mechanism. MCa appeared to share the same features as the so-called cell-penetrating peptides. Our results provide evidence that MCa has the ability to act as a molecular carrier and to cross cell membranes in a rapid manner (1-2 min), making this toxin the first demonstrated example of a scorpion toxin that translocates into cells. PMID- 15653691 TI - The 14-3-3 proteins of Trypanosoma brucei function in motility, cytokinesis, and cell cycle. AB - The cDNAs for two isoforms (I and II) of the 14-3-3 proteins have been cloned and functionally characterized in Trypanosoma brucei. The amino acid sequences of isoforms I and II have 47 and 50% identity to the human tau isoform, respectively, with important conserved features including a potential amphipathic groove for the binding of phosphoserine/phosphothreonine-containing motifs and a nuclear export signal-like domain. Both isoforms are abundantly expressed at approximately equal levels (1-2 x 10(6) molecules/cell) and localized mainly in the cytoplasm. Knockdown by induction of double-stranded RNA of isoform I and/or II in both bloodstream and procyclic forms resulted first in a reduction of cell motility and then significant reduction in cell growth rates and morphological changes; the changes include aberrant numbers of organelles and abnormal shapes and sizes that mimic phenotypes produced by various cytokinesis inhibitors. Morphological and fluorescence-activated cell sorting analysis of the cell cycle suggested that isoforms I and II might play important roles in nuclear (G2-M transition) and cell (M-G1 transition) division. These findings indicate that the 14-3-3 proteins play important roles in cell motility, cytokinesis, and the cell cycle. PMID- 15653692 TI - Signaling mechanisms responsible for lysophosphatidic acid-induced urokinase plasminogen activator expression in ovarian cancer cells. AB - Lysophosphatidic acid (LPA) enhances urokinase plasminogen activator (uPA) expression in ovarian cancer cells; however, the molecular mechanisms responsible for this event have not been investigated. In this study, we used the invasive ovarian cancer SK-OV-3 cell line to explore the signaling molecules and pathways essential for LPA-induced uPA up-regulation. With the aid of specific inhibitors and dominant negative forms of signaling molecules, we determined that the G(i) associated pathway mediates this LPA-induced event. Moreover, constitutively active H-Ras and Raf-1-activating H-Ras mutant enhance uPA expression, whereas dominant negative H-Ras and Raf-1 block LPA-induced uPA up-regulation, suggesting that the Ras-Raf pathway works downstream of G(i) to mediate this LPA-induced process. Surprisingly, dominant negative MEK1 or Erk2 displays only marginal inhibitory effect on LPA-induced uPA up-regulation, suggesting that a signaling pathway distinct from Raf-MEK1/2-Erk is the prominent pathway responsible for this process. In this report, we demonstrate that LPA activates NF-kappaB in a Ras-Raf-dependent manner and that blocking NF-kappaB activation with either non phosphorylable IkappaB or dominant negative IkappaB kinase abolished LPA-induced uPA up-regulation and uPA promoter activation. Furthermore, introducing mutations to knock out the NF-kappaB binding site of the uPA promoter results in over 80% reduction in LPA-induced uPA promoter activation, whereas this activity is largely intact with the promoter containing mutations in the AP1 binding sites. Thus these results suggest that the G(i)-Ras-Raf-NF-kappaB signaling cascade is responsible for LPA-induced uPA up-regulation in ovarian cancer cells. PMID- 15653693 TI - Regulation of mitochondrial NADP+-dependent isocitrate dehydrogenase activity by glutathionylation. AB - Recently, we demonstrated that the control of mitochondrial redox balance and oxidative damage is one of the primary functions of mitochondrial NADP(+) dependent isocitrate dehydrogenase (IDPm). Because cysteine residue(s) in IDPm are susceptible to inactivation by a number of thiol-modifying reagents, we hypothesized that IDPm is likely a target for regulation by an oxidative mechanism, specifically glutathionylation. Oxidized glutathione led to enzyme inactivation with simultaneous formation of a mixed disulfide between glutathione and the cysteine residue(s) in IDPm, which was detected by immunoblotting with anti-GSH IgG. The inactivated IDPm was reactivated enzymatically by glutaredoxin2 in the presence of GSH, indicating that the inactivated form of IDPm is a glutathionyl mixed disulfide. Mass spectrometry and site-directed mutagenesis further confirmed that glutathionylation occurs to a Cys(269) of IDPm. The glutathionylated IDPm appeared to be significantly less susceptible than native protein to peptide fragmentation by reactive oxygen species and proteolytic digestion, suggesting that glutathionylation plays a protective role presumably through the structural alterations. HEK293 cells and intact respiring mitochondria treated with oxidants inducing GSH oxidation such as H(2)O(2) or diamide showed a decrease in IDPm activity and the accumulation of glutathionylated enzyme. Using immunoprecipitation with anti-IDPm IgG and immunoblotting with anti-GSH IgG, we were also able to purify and positively identify glutathionylated IDPm from 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine treated mice, a model for Parkinson's disease. The results of the current study indicate that IDPm activity appears to be modulated through enzymatic glutathionylation and deglutathionylation during oxidative stress. PMID- 15653694 TI - Interleukin-3 and granulocyte-macrophage colony-stimulating factor inhibits tumor necrosis factor (TNF)-alpha-induced osteoclast differentiation by down-regulation of expression of TNF receptors 1 and 2. AB - Osteoclasts, the multinucleated cells that resorb bone, differentiate from hemopoietic precursors of monocyte/macrophage lineage, which also give rise to macrophages or dendritic cells. In this study we investigated the mechanism by which interleukin-3 (IL-3) and granulocyte-macrophage colony-stimulating factor (GM-CSF) inhibit tumor necrosis factor (TNF)-alpha-induced osteoclast differentiation in mouse osteoclast precursors. We show here that both IL-3 and GM-CSF potently inhibits TNF-alpha-induced osteoclast differentiation by direct action on osteoclast precursors. The inhibitory effect of IL-3 and GM-CSF on osteoclast differentiation was completely neutralized by anti-IL-3 and anti-GM CSF antibodies, respectively. In addition, the inhibitory effect of IL-3 and GM CSF on TNF-alpha-induced osteoclast differentiation was irreversible. In osteoclast precursors, IL-3 and GM-CSF inhibited c-Fms expression post transcriptionally. Interestingly, IL-3 and GM-CSF down-regulated both mRNA and surface expression of TNF receptor 1 (TNFR1) and TNFR2. Furthermore, cells in the presence of IL-3 and GM-CSF showed high expression of macrophage antigen CD11b, and low expression of dendritic cells antigen CD11c and prolong exposure of osteoclast precursors to GM-CSF increased the CD11c expression compare with IL-3. In summary, we provide the first evidence that IL-3 and GM-CSF block TNF-alpha induced osteoclast differentiation by down-regulation of mRNA and surface expression of TNFR1 and TNFR2. PMID- 15653696 TI - The TSG-6 and I alpha I interaction promotes a transesterification cleaving the protein-glycosaminoglycan-protein (PGP) cross-link. AB - During co-incubation of human inter-alpha-inhibitor (IalphaI) and human tumor necrosis factor-stimulated gene 6 protein (TSG-6) SDS-stable interactions are formed between the two proteins. We have analyzed the products of this reaction and characterized the mechanism of complex formation. Following the incubation seven new bands not previously identified were apparent in SDS-PAGE. Three of these bands did not contain TSG-6, including heavy chain (HC)1.bikunin, HC2.bikunin, and free bikunin. In addition high molecular weight complexes composed of the same components as I alpha I, including HC1, HC2, and bikunin, were formed. The formation of these complexes was prevented by the addition of hyaluronan. The cross-links stabilizing these complexes displaying properties similar to the protein-glycosaminoglycan-protein (PGP) cross-link. The TSG-6 containing SDS-stable complexes were composed of HC1.TSG-6 or HC2.TSG-6 exclusively. Both glycosylated and non-glycosylated TSG-6 participated in the complex formation. The HC.TSG-6 cross-links were different from the PGP cross link and were determined to be ester bonds between the alpha-carbonyl of the C terminal Asp of the heavy chain and most likely a hydroxyl group containing the TSG-6 residue. The mechanism involved cleaving the PGP cross-link of I alpha I during a transesterification reaction. A TSG-6 hydroxyl group reacts with the ester bond between the alpha-carbonyl of the C-terminal Asp residues of HC1 or HC2 and carbon-6 of an internal N-acetylgalactosamine of the chondroitin-4 sulfate chain. An intermediate is formed resulting in a partitioning of the reaction between HC(1 or 2).TSG-6 complexes and transfer of HC(1 or 2) to the chondroitin via competing pathways. PMID- 15653695 TI - Granulin and granulin repeats interact with the Tat.P-TEFb complex and inhibit Tat transactivation. AB - The cellular positive transcription elongation factor b (P-TEFb), containing cyclin T1 and cyclin-dependent kinase 9 (CDK9), interacts with the human immunodeficiency virus, type 1 (HIV-1) regulatory protein Tat to enable viral transcription and replication. Cyclin T1 is an unusually long cyclin and is engaged by cellular regulatory proteins. Previous studies showed that the granulin/epithelin precursor (GEP) binds the histidine-rich region of cyclin T1 and inhibits P-TEFb function. GEP is composed of repeats that vary in sequence and properties. GEP also binds directly to Tat. Here we show that GEP and some of its constituent granulin repeats can inhibit HIV-1 transcription via Tat without directly binding to cyclin T1. The interactions of granulins with Tat and cyclin T1 differ with respect to their binding sites and divalent cation requirements, and we identified granulin repeats that bind differentially to Tat and cyclin T1. Granulins DE and E bind Tat but do not interact directly with cyclin T1. These granulins are present in complexes with Tat and P-TEFb in which Tat forms a bridge between the cellular proteins. Granulins DE and E repress transcription from the HIV-1 LTR and gene expression from the viral genome, raising the possibility of developing granulin-based inhibitors of viral infection. PMID- 15653697 TI - The c-myc DNA-unwinding element-binding protein modulates the assembly of DNA replication complexes in vitro. AB - The presence of DNA-unwinding elements (DUEs) at eukaryotic replicators has raised the question of whether these elements contribute to origin activity by their intrinsic helical instability, as protein-binding sites, or both. We used the human c-myc DUE as bait in a yeast one-hybrid screen and identified a DUE binding protein, designated DUE-B, with a predicted mass of 23.4 kDa. Based on homology to yeast proteins, DUE-B was previously classified as an aminoacyl-tRNA synthetase; however, the human protein is approximately 60 amino acids longer than its orthologs in yeast and worms and is primarily nuclear. In vivo, chromatin-bound DUE-B localized to the c-myc DUE region. DUE-B levels were constant during the cell cycle, although the protein was preferentially phosphorylated in cells arrested early in S phase. Inhibition of DUE-B protein expression slowed HeLa cell cycle progression from G1 to S phase and induced cell death. DUE-B extracted from HeLa cells or expressed from baculovirus migrated as a dimer during gel filtration and co-purified with ATPase activity. In contrast to endogenous DUE-B, baculovirus-expressed DUE-B efficiently formed high molecular mass complexes in Xenopus egg and HeLa extracts. In Xenopus extracts, baculovirus-expressed DUE-B inhibited chromatin replication and replication protein A loading in the presence of endogenous DUE-B, suggesting that differential covalent modification of these proteins can alter their effect on replication. Recombinant DUE-B expressed in HeLa cells restored replication activity to egg extracts immunodepleted with anti-DUE-B antibody, suggesting that DUE-B plays an important role in replication in vivo. PMID- 15653699 TI - Effectiveness of speed cameras in preventing road traffic collisions and related casualties: systematic review. AB - OBJECTIVES: To assess whether speed cameras reduce road traffic collisions and related casualties. DESIGN: Systematic review. DATA SOURCES: Cochrane Injuries Group Specialised Register, Cochrane Central Register of Controlled Trials, Medline, Embase, Social Science Citation Index, TRANSPORT database, ZETOC, the internet (including websites of road safety and motoring organisations), and contact with key individuals and organisations. MAIN OUTCOME MEASURES: Road traffic collisions, injuries, and deaths. INCLUSION CRITERIA: Controlled trials and observational studies assessing the impact of fixed or mobile speed cameras on any of the selected outcomes. RESULTS: 14 observational studies met the inclusion criteria; no randomised controlled trials were found. Most studies were before-after studies without controls (n = 8). All but one of the studies showed effectiveness of cameras up to three years or less after their introduction; one study showed sustained longer term effects (4.6 years after introduction). Reductions in outcomes across studies ranged from 5% to 69% for collisions, 12% to 65% for injuries, and 17% to 71% for deaths in the immediate vicinity of camera sites. The reductions over wider geographical areas were of a similar order of magnitude. CONCLUSIONS: Existing research consistently shows that speed cameras are an effective intervention in reducing road traffic collisions and related casualties. The level of evidence is relatively poor, however, as most studies did not have satisfactory comparison groups or adequate control for potential confounders. Controlled introduction of speed cameras with careful data collection may offer improved evidence of their effectiveness in the future. PMID- 15653700 TI - Laparoscopic management of borderline ovarian tumors: results of a French multicenter study. AB - BACKGROUND: Laparoscopy in the management of women with borderline ovarian tumors remains controversial. We therefore evaluated the adequacy of initial laparoscopic staging according to FIGO guidelines, by comparison with laparotomy. PATIENTS AND METHODS: In a French retrospective multicenter study of 358 women with borderline ovarian tumors, we compared epidemiological characteristics, sonographic findings, serum tumor marker levels, and surgical and histological parameters between women undergoing laparoscopy and women undergoing laparotomy. RESULTS: One hundred and forty-nine (41.6%) of the 358 women underwent laparoscopy. Mean age, mean gestity and parity, and mean tumor size were higher in the laparotomy group. Forty-two women (28.2%) underwent laparoconversion, mainly for suspected ovarian cancer or large tumor volume. Conservative treatment and cyst rupture were more frequent in the laparoscopy group than in the laparoconversion and laparotomy groups (P < 0.0001). The rate of complete staging was lower in the laparoscopy group than in the laparoconversion and laparotomy groups (P < 0.0001), with no difference between these latter two groups. No difference in the recurrence rate was noted between the groups, but a higher recurrence rate was observed after conservative treatment (P < 0.001). CONCLUSIONS: Laparoscopic management of borderline ovarian tumors is associated with a higher rate of cyst rupture and incomplete staging. Recurrence was more frequent after conservative treatment. Whatever the surgical route, the rate of complete initial staging was low, emphasizing the need to respect treatment guidelines for borderline ovarian tumors. PMID- 15653701 TI - Prognostic factors in advanced synovial sarcoma: an analysis of 104 patients treated at the Royal Marsden Hospital. AB - AIMS: Key prognostic factors at diagnosis of synovial sarcoma are well defined from the literature. There are few data regarding prognostic parameters in the setting of advanced disease. Our aim was to look specifically at a cohort of patients with advanced synovial sarcoma and to identify potential prognostic factors. PATIENTS AND METHODS: One hundred and four patients with advanced synovial sarcoma were identified from the Royal Marsden Hospital's sarcoma database between 1978 and 2003. Patient data were analysed retrospectively. Most patients were aged between 20 and 50 years at diagnosis. Seventy-one patients were deceased at the time of analysis. Ninety-two patients received chemotherapy for management of advanced disease (most commonly doxorubicin + ifosfamide). RESULTS: Median survival following development of advanced disease was 22 months. Predictors of survival with advanced disease were age <35 years (P = 0.03) and response to first-line chemotherapy (P = 0.05). The response rate to doxorubicin plus ifosfamide was 58.6%, and this was superior to either agent when given singly. Metastasectomy was not associated with improved prognosis in this series. CONCLUSIONS: Synovial sarcoma is a chemosensitive soft tissue sarcoma. Compared with historical controls, survival with advanced disease seems to have improved over the years, possibly as a result of better use of chemotherapy. Age <35 years and response to first-line chemotherapy predict for improved survival with advanced disease. PMID- 15653702 TI - Phase I-II study of amrubicin and cisplatin in previously untreated patients with extensive-stage small-cell lung cancer. AB - BACKGROUND: Amrubicin, a totally synthetic 9-amino-anthracycline, demonstrated excellent single-agent activity for extensive-stage small-cell lung cancer (ED SCLC). The aims of this trial were to determine the maximum-tolerated doses (MTD) of combination therapy with amrubicin and cisplatin, and to assess the efficacy and safety at their recommended doses (RD). PATIENTS AND METHODS: Eligibility criteria were patients having histologically or cytologically proven measurable ED-SCLC, no previous systemic therapy, an Eastern Cooperative Oncology Group performance status of 0-2 and adequate organ function. Amrubicin was administered on days 1-3 and cisplatin on day 1, every 3 weeks. RESULTS: Four patients were enrolled at dose level 1 (amrubicin 40 mg/m(2)/day and cisplatin 60 mg/m(2)) and three patients at level 2 (amrubicin 45 mg/m(2)/day and cisplatin 60 mg/m(2)). Consequently, the MTD and RD were determined to be at level 2 and level 1, respectively. The response rate at the RD was 87.8% (36/41). The median survival time (MST) was 13.6 months and the 1-year survival rate was 56.1%. Grade 3/4 neutropenia and leukopenia occurred in 95.1% and 65.9% of patients, respectively. CONCLUSIONS: The combination of amrubicin and cisplatin has demonstrated an impressive response rate and MST in patients with previously untreated ED-SCLC. PMID- 15653703 TI - High-dose methotrexate toxicity in elderly patients with primary central nervous system lymphoma. AB - BACKGROUND: The dose of high-dose methotrexate (HDMTX) in elderly patients often has to be reduced, resulting in a loss of treatment efficacy. We evaluated HDMTX related toxicity with special regard to age distribution in patients with primary central nervous system lymphoma (PCNSL) in a phase IV multicenter trial. PATIENTS AND METHODS: One hundred and fifty-four patients (median age 61 years; 89 patients >60 years old, 21 patients >70 years old) received 619 HDMTX cycles. Toxicity was evaluated prospectively using the WHO classification. Unless a reduced dose was required after calculating a decreased glomerular filtration rate (GFR), the patients received 4 g/m(2) HDMTX followed by leucovorin rescue. RESULTS: Toxicity was generally mild with toxicities of WHO grade > or =3 usually <10%. The differences in the incidence and severity of toxicity were not statistically significant between patients >60 years and < or =60 years old. The same was true for therapy termination owing to MTX toxicity and for delayed serum MTX clearance. Dose reduction significantly differed between patients < or =60 years and those >60 years old (18% versus 44%; P = 0.001). CONCLUSIONS: HDMTX is a safe treatment for PCNSL patients regardless of age, with adherence to dose reduction determined by calculating the GFR before each treatment cycle. PMID- 15653704 TI - Dopexamine and its role in the protection of hepatosplanchnic and renal perfusion in high-risk surgical and critically ill patients. AB - BACKGROUND: Dopexamine is increasingly being used in high-risk surgical and critically ill patients to preserve hepatosplanchnic and renal perfusion. This systematic review of randomized controlled trials was undertaken to investigate the clinical evidence for using dopexamine in this role. METHODS: Data sources included Medline, Cochrane Library, EMBASE and CINAHL and reference lists of relevant articles. Randomized controlled trials which compared dopexamine treatment with a control group, in high-risk surgical and critically ill adult patients and with primary outcome measures designed to assess hepatosplanchnic and renal perfusion were included. Articles not published in English were excluded. RESULTS: Twenty-one trials were selected from the literature search. The results suggest that dopexamine may protect against colonic mucosal damage in patients undergoing abdominal aortic surgery and may improve gastric mucosal pHi in general surgical patients, especially those with preoperative gastric mucosal pHi measurements <7.35 and those undergoing pancreatico-duodenectomy surgery. Dopexamine may have beneficial effects on renal perfusion in patients undergoing cardiac surgery but appears to have little or no benefit on gastric mucosal pHi in the same patient population. In critically ill patients none of the studies demonstrated a beneficial effect of dopexamine on either hepatosplanchnic or renal perfusion. CONCLUSION: The evidence provided by the existing studies is both inadequate and inconsistent. There is insufficient evidence to offer reliable recommendations on the clinical use of dopexamine for the protection of either hepatosplanchnic or renal perfusion in high-risk surgical patients. Furthermore, there is no current evidence to support a role for dopexamine in protecting either hepatosplanchnic or renal perfusion in critically ill patients. PMID- 15653705 TI - Effects of magnesium sulphate and clonidine on propofol consumption, haemodynamics and postoperative recovery. AB - BACKGROUND: This placebo-controlled, double-blind study was designed to assess the effects of magnesium sulphate and clonidine on peroperative haemodynamics, propofol consumption and postoperative recovery. METHODS: Sixty ASA I-II patients undergoing spinal surgery were randomized into three groups. Group M received magnesium sulphate 30 mg kg(-1) as a bolus before induction and 10 mg kg(-1) h( 1) by infusion. Group CL received clonidine 3 microg kg(-1) as a bolus before induction and 2 microg kg(-1) h(-1) by infusion during the operation period. The same volume of isotonic solution was administered to the control group (group CT). Anaesthesia was induced with propofol and was maintained with propofol infusion [dose according to the bispectral index (BIS)], fentanyl and cisatracurium. Analysis of variance and the Bonferroni test were used for statistical analysis. RESULTS: Induction of anaesthesia with propofol was rapid in the presence of magnesium sulphate and clonidine. The time for BIS to reach 60 was significantly shorter in group M and group CL (P<0.0001) but postoperative recovery was slower with magnesium sulphate compared with the clonidine and control groups (P<0.0001). There was no statistical difference in heart rate and arterial blood pressure between the groups. Propofol requirements for induction and maintenance of anaesthesia were significantly lower with magnesium and clonidine (P<0.0001). CONCLUSION: Clonidine caused bradycardia and hypotension and magnesium sulphate caused delayed recovery, but can be used as adjuvant agents with careful management. PMID- 15653706 TI - Life-threatening postoperative blood loss in a Jehovah's Witness, treated with high-dose erythropoietin. AB - Six hours after an uncomplicated extended resection of ovarian cancer, postoperative arterial bleeding led to life-threatening blood loss in a 44-yr-old Jehovah's Witness who refused blood transfusion. Haemoglobin (Hb) decreased from 2.5 g dl(-1) directly after the emergency laparotomy, followed by a 10 h immeasurable period (below detectable minimum value of the analyser), to a measurable minimum of 1.5 g dl(-1) after 20 h. Haematopoiesis was induced by high dose i.v. erythropoietin therapy (600 IU kg(-1)) and continued on days 3, 6, 8, 10 and 13. Iron, folic acid and vitamins were given as supplements. The patient needed ventilatory assistance for 18 days and some inotropic support. Complications included increases in pancreatic enzymes and liver enzymes, jaundice and skin necrosis at the fingertips and toes. Myopathy led to transient tetraparesis. Haemoglobin rose from 1.5 to 3.4 g dl(-1) (day 10) and the patient was discharged from the intensive care unit with haemoglobin 6.5 g dl(-1) on day 24. She made a full recovery and is still free of cancer in remission. PMID- 15653707 TI - Impact of peripheral elimination on the concentration-effect relationship of remifentanil in anaesthetized dogs. AB - BACKGROUND: This study elucidates the impact of sampling site when estimating pharmacokinetic-pharmacodynamic (PK-PD) parameters of drugs such as remifentanil that undergo tissue extraction in the biophase. The interrelationship between the concentrations of remifentanil predicted for the effect compartment and those measured in arterial, venous, and cerebrospinal fluid were investigated under steady-state conditions. METHODS: Following induction of anaesthesia with pentobarbital, an arterial cannula (femoral) and two venous catheters (jugular and femoral) were inserted. Electrodes were placed for EEG recording of theta wave activity. Each dog received two consecutive 5-min infusions for the PK-PD study and a bolus followed by a 60-min infusion was started for the steady-state study. Cerebrospinal fluid, arterial and venous blood samples were drawn simultaneously after 30, 40, and 50 min. At the end of the infusion, arterial blood samples were collected for pharmacokinetic analysis. RESULTS: Remifentanil PK-PD parameters based on theta wave activity were as follows: apparent volume of distribution at steady-state (V(ss)) (231+/-37 ml kg(-1)), total body clearance (Cl) (63+/-16 ml min(-1) kg(-1)), terminal elimination half-life (t(1/2 beta)) (7.71 min), effect compartment concentration at 50% of maximal observed effect (EC(50)) (21+/-13 ng ml(-1)), and equilibration rate constant between plasma and effect compartment (k(e0)) (0.48+/-0.24 min). The mean steady-state cerebrospinal fluid concentration of 236 ng ml(-1) represented 52 and 74% of that in arterial and venous blood, respectively. CONCLUSIONS: Our study re-emphasizes the importance of a sampling site when performing PK-PD modelling for drugs undergoing elimination from the effect compartment. For a drug undergoing tissue elimination such as remifentanil, venous rather than arterial concentrations will reflect more exactly the effect compartment concentrations, under steady-state conditions. PMID- 15653708 TI - ECG recording of central venous catheter misplaced in inferior thyroid artery. AB - A 71-year-old male patient with liver metastases secondary to rectal carcinoma was scheduled for hemi-hepatectomy. Two months earlier he had undergone subtotal resection of the thyroid gland. Prior to surgery, a triple-lumen catheter and an introducer sheath were introduced into the right internal jugular vein using a landmark technique. No problems occurred during insertion of the triple-lumen catheter, but resistance was noticed during insertion of the 8.5 FG introducer sheath. After placement of the introducer sheath, intra-arterial misplacement was confirmed using a pressure transducer. The opportunity was taken to record and compare intravascular ECG by the arterial and venous catheters before removal. No difference was noticed in the P-wave patterns; both showed a marked increase. Surgical exploration of the neck, recommended by the vascular surgeon consulted, showed that the carotid artery was not injured. The introducer sheath had completely punctured the right internal jugular vein and entered the inferior thyroid artery. A thrill was felt. The management of this case is discussed, with suggestions for best practice. Intravascular ECG was unhelpful in differentiating between venous and arterial placement of the catheter. PMID- 15653709 TI - Clinical evaluation of USCOM ultrasonic cardiac output monitor in cardiac surgical patients in intensive care unit. AB - BACKGROUND: The USCOM ultrasonic cardiac output monitor (USCOM Pty Ltd, Coffs Harbour, NSW, Australia) is a non-invasive device that determines cardiac output by continuous-wave Doppler ultrasound. The aim of this study was to evaluate the accuracy of the USCOM device compared with the thermodilution technique in intensive care patients who had just undergone cardiac surgery. METHODS: We conducted a prospective study in the 18-bed intensive care unit of a 600-bed tertiary referral hospital. Twenty-four mechanically ventilated patients were studied immediately following cardiac surgery. We evaluated the USCOM monitor by comparing its output with paired measurements obtained by the standard thermodilution technique using a pulmonary artery catheter. RESULTS: Forty paired measurements were obtained in 22 patients. We were unable to obtain an acceptable signal in the remaining two patients. Comparison of the two techniques showed a bias of 0.18 and limits of agreement of -1.43 to 1.78. The agreement may not be as good between techniques at higher cardiac output values. CONCLUSIONS: The USCOM monitor has a place in intensive care monitoring. It is accurate, rapid, safe, well-tolerated, non-invasive and cost-effective. The learning curve for skill acquisition is very short. However, during the learning phase the USCOM monitor measurements are rather 'operator dependent'. Its suitability for use in high and low cardiac output states requires further validation. PMID- 15653711 TI - 3-nitrotyrosine attenuates respiratory syncytial virus infection in human bronchial epithelial cell line. AB - 3-nitrotyrosine (NO2Tyr), an L-tyrosine derivative during nitrative stress, can substitute the COOH-terminal tyrosine of alpha-tubulin, posttranslationally altering microtubular functions. Because infection of the cells by respiratory syncytial virus (RSV) may require intact microtubules, we tested the hypothesis that NO2Tyr would inhibit RSV infection and intracellular signaling via nitrotyrosination of alpha-tubulin. A human bronchial epithelial cell line (BEAS 2B) was incubated with RSV with or without NO2Tyr. The release of chemokines and viral particles and activation of interferon regulatory factor-3 (IRF-3) were measured. Incubation with NO2Tyr increased nitrotyrosinated alpha-tubulin, and NO2Tyr colocalized with microtubules. RSV-infected cells released viral particles, RANTES, and IL-8 in a time- and dose-dependent manner, and intracellular RSV proteins coprecipitated with alpha-tubulin. NO2Tyr attenuated the RSV-induced release of RANTES, IL-8, and viral particles by 50-90% and decreased alpha-tubulin-associated RSV proteins. 3-chlorotyrosine, another L tyrosine derivative, had no effects. NO2Tyr also inhibited the RSV-induced shift of the unphosphorylated form I of IRF-3 to the phosphorylated form II. Pre exposure of the cells to NO(2) (0.15 ppm, 4 h), which produced diffuse protein tyrosine nitration, did not affect RSV-induced release of RANTES, IL-8, or viral particles. NO2Tyr did not affect the potential of viral spreading to the neighboring cells since the RSV titers were not decreased when the uninfected cells were cocultured with the preinfected cells in NO2Tyr-containing medium. These results indicate that NO2Tyr, by replacing the COOH-terminal tyrosine of alpha-tubulin, attenuated RSV infection, and the inhibition appeared to occur at the early stages of RSV infection. PMID- 15653712 TI - Loss of Gadd45a does not modify the pulmonary response to oxidative stress. AB - It is well established that exposure to high levels of oxygen (hyperoxia) injures and kills microvascular endothelial and alveolar type I epithelial cells. In contrast, significant death of airway and type II epithelial cells is not observed at mortality, suggesting that these cell types may express genes that protect against oxidative stress and damage. During a search for genes induced by hyperoxia, we previously reported that airway and alveolar type II epithelial cells uniquely express the growth arrest and DNA damage (Gadd)45a gene. Because Gadd45a has been implicated in protection against genotoxic stress, adult Gadd45a (+/+) and Gadd45a (-/-) mice were exposed to hyperoxia to investigate whether it protected epithelial cells against oxidative stress. During hyperoxia, Gadd45a deficiency did not affect loss of airway epithelial expression of Clara cell secretory protein or type II epithelial cell expression of pro-surfactant protein C. Likewise, Gadd45a deficiency did not alter recruitment of inflammatory cells, edema, or overall mortality. Consistent with Gadd45a not affecting the oxidative stress response, p21(Cip1/WAF1) and heme oxygenase-1 were comparably induced in Gadd45a (+/+) and Gadd45a (-/-) mice. Additionally, Gadd45a deficiency did not affect oxidative DNA damage or apoptosis as assessed by oxidized guanine and terminal deoxyneucleotidyl transferase-mediated dUTP nick-end labeling staining. Overexpression of Gadd45a in human lung adenocarcinoma cells did not affect viability or survival during exposure, whereas it was protective against UV radiation. We conclude that increased tolerance of airway and type II epithelial cells to hyperoxia is not attributed solely to expression of Gadd45a. PMID- 15653710 TI - 2-aminoethoxydiphenyl borate stimulates pulmonary C neurons via the activation of TRPV channels. AB - This study was carried out to determine the effect of 2-aminoethoxydiphenyl borate (2-APB), a common activator of transient receptor potential vanilloid (TRPV) type 1, 2, and 3 channels, on cardiorespiratory reflexes, pulmonary C fiber afferents, and isolated pulmonary capsaicin-sensitive neurons. In anesthetized, spontaneously breathing rats, intravenous bolus injection of 2-APB elicited the pulmonary chemoreflex responses, characterized by apnea, bradycardia, and hypotension. After perineural treatment of both cervical vagi with capsaicin to block the conduction of C fibers, 2-APB no longer evoked any of these reflex responses. In open-chest and artificially ventilated rats, 2-APB evoked an abrupt and intense discharge in vagal pulmonary C fibers in a dose dependent manner. The stimulation of C fibers by 2-APB was attenuated but not abolished by capsazepine, a selective antagonist of the TRPV1, which completely blocked the response to capsaicin in these C fiber afferents. In isolated pulmonary capsaicin-sensitive neurons, 2-APB concentration dependently evoked an inward current that was partially inhibited by capsazepine but almost completely abolished by ruthenium red, an effective blocker of all TRPV channels. In conclusion, 2-APB evokes a consistent and distinct stimulatory effect on pulmonary C fibers in vivo and on isolated pulmonary capsaicin-sensitive neurons in vitro. These results establish the functional evidence demonstrating that TRPV1, V2, and V3 channels are expressed on these sensory neurons and their terminals. PMID- 15653713 TI - Ethanol stimulates the expression of fibronectin in lung fibroblasts via kinase dependent signals that activate CREB. AB - Ethanol renders the lung susceptible to acute lung injury in the setting of insults such as sepsis. The mechanisms mediating this effect are unknown, but activation of tissue remodeling is considered key to this process. We found that chronic ethanol ingestion in rats increased the expression of fibronectin, a matrix glycoprotein implicated in acute lung injury. In cultured NIH/3T3 cells and in primary rat and mouse lung fibroblasts, ethanol induced fibronectin mRNA and protein expression in a dose- and time-dependent fashion. The effect of ethanol was prevented by inhibitors of protein kinase C and mitogen-activated protein kinases and was associated with the phosphorylation and increased DNA binding of the transcription factor cAMP response element binding protein, followed by increased transcription of the fibronectin gene. Fibroblasts were found to express alpha(7) nicotinic acetylcholine receptor (nAChR), and ethanol induction of fibronectin was abolished by alpha-bungarotoxin and methyllcaconitine, inhibitors of alpha(7) nAChRs. However, ethanol was able to induce fibronectin mRNA and protein in primary lung fibroblasts isolated from alpha(7) nAChR knockout mice. The ethanol-induced fibronectin response was dependent on ethanol metabolism since 4-methylpyrazole, an inhibitor of alcohol dehydrogenase, abolished the effect and acetaldehyde induced it. These observations suggest that ethanol or ethanol metabolites stimulate lung fibroblasts to produce fibronectin by inducing specific signals transmitted via nAChRs independent of the alpha(7-)subunit, and this might represent a mechanism by which ethanol renders the lung susceptible to acute lung injury. PMID- 15653714 TI - Laser Doppler flowmetry is valid for measurement of cerebral blood flow autoregulation lower limit in rats. AB - Laser Doppler flowmetry (LDF) is a recent technique that is increasingly being used to monitor relative changes in cerebral blood flow whereas the intra arterial 133xenon injection technique is a well-established method for repeated absolute measurements of cerebral blood flow. The aim of this study was to validate LDF for assessment of cerebral autoregulation and CO2 reactivity with the 133xenon injection technique as the gold standard. Simultaneous measurements of cerebral blood flow (CBF) were collected by LDF (CBF(LDF)) and the 133xenon method (CBF(Xe)) while (1) cerebral autoregulation was challenged by controlled systemic haemorrhage, or (2) cerebral blood flow was varied by manipulating the arterial partial pressure of CO2 (P(a,CO2)). LDF slightly overestimated CBF under conditions of haemorrhagic shock and haemodilution caused by controlled haemorrhage (paired t test, P < 0.05). However for pooled data, the autoregulation lower limit was similar when determined with the 133xenon and the LDF techniques: 65 +/- 3.9 mmHg and 60 +/- 5.6 mmHg, respectively. Linear regression analysis yielded CBF(Xe) = (1.02 x CBF(LDF)) + 9.1 and r = 0.90. Even for substantial changes in P(a,CO2), the two methods resulted in similar results. We conclude that even though LDF overestimated CBF during haemorrhagic shock caused by controlled haemorrhage, the lower limit autoregulation was correctly identified. The laser Doppler technique provides a reliable method for detection of a wide range of cerebral blood flow changes under CO2 challenge. Haemodilution influences the two methods differently causing relative overestimation of blood flow by the laser Doppler technique compared to the 1(33)xenon method. PMID- 15653715 TI - Role of adrenergic receptors in the reflex diuresis in rabbits during pulmonary lymphatic obstruction. AB - The role of adrenergic receptors in the reflex diuresis in response to pulmonary lymphatic drainage was examined in anaesthetized, artificially ventilated New Zealand White rabbits. Pulmonary lymphatic drainage was obstructed by raising the pressure in a pouch created from the right external jugular vein. This pulmonary lymphatic obstruction results in a reflex increase in urine flow and sodium excretion. This reflex is abolished by renal denervation and by administration of L-NAME, a non-selective inhibitor of nitric oxide synthase. Also, infusion of the relatively selective neuronal nitric oxide synthase blocker, 7-nitroindazole sodium salt, into the renal medulla abolished the reflex diuresis. In this study the effects of adrenergic receptor antagonists on the reflex increase in urine were observed. Both ureters were cannulated in order to determine urine flow from both kidneys separately. Prazosin, an alpha1 adrenergic receptor antagonist, was infused into the renal medulla of the right kidney, while the left kidney acted as control. Administration of prazosin in this manner did not block the reflex diuresis in response to pulmonary lymphatic obstruction in either kidney. However, rauwolscine, an alpha2 adrenergic receptor antagonist, abolished the reflex increase in urine and sodium excretion in the ipsilateral kidney while preserving it in the contralateral kidney. These findings suggest that the increase in urine flow in rabbits caused by pulmonary lymphatic obstruction is dependent upon activation of alpha2 adrenergic receptors within the renal medulla. PMID- 15653716 TI - Kallikrein-kinin in stroke, cardiovascular and renal disease. AB - Tissue kallikrein, a serine proteinase, produces the potent vasodilator kinin peptide from kininogen substrate. The levels of tissue kallikrein are reduced in humans and animal models with hypertension, cardiovascular and renal disease. Using transgenic and somatic gene transfer approaches, we investigated the role of the tissue kallikrein-kinin system in cardiovascular, renal and central nervous systems. A single injection of the human tissue kallikrein gene in plasmid DNA or an adenoviral vector resulted in a prolonged reduction of blood pressure and attenuation of hypertrophy and fibrosis in the heart and kidney of several hypertensive animal models. Furthermore, enhanced kallikrein-kinin levels after gene transfer exerted beneficial effects, with protection against cardiac remodelling, renal injuries, restenosis, cerebral infarction and neurological deficits in normotensive animal models without haemodynamic effects, indicating direct actions of kallikrein independent of its ability to lower blood pressure. The effects of kallikrein were mediated by the kinin B2 receptor, as the specific B2 receptor antagonist icatibant abolished the actions of kallikrein. Moreover, kallikrein-kinin exhibited pleiotropic effects by inhibiting apoptosis, inflammation, hypertrophy and fibrosis, and promoting angiogenesis and neurogenesis in the heart, kidney, brain and blood vessel. Exogenous administration of kallikrein also led to increased nitric oxide (NO)/cGMP and cAMP levels, and reduced NAD(P)H oxidase activities, superoxide formation and pro inflammatory cytokine levels. These results indicate a novel role of kallikrein kinin through the kinin B2 receptor as an antioxidant and anti-inflammatory agent in protection against stroke, cardiovascular and renal disease, and may uncover new drug targets for the prevention and treatment of heart failure, vascular injury, end-stage renal disease and stroke in humans. PMID- 15653717 TI - Disruption of protein-mediated DNA looping by tension in the substrate DNA. AB - Protein-mediated DNA looping is important in a variety of biological processes, including gene regulation and genetic transformation. Although the biochemistry of loop formation is well established, the mechanics of loop closure in a constrained cellular environment has received less attention. Recent single molecule measurements show that mechanical constraints have a significant impact on DNA looping and motivate the need for a more comprehensive characterization of the effects of tension. By modeling DNA as a wormlike chain, we calculate how continuous stretching of the substrate DNA affects the loop formation probability. We find that when the loop size is >100 bp, a tension of 500 fN can increase the time required for loop closure by two orders of magnitude. This force is small compared to the piconewton forces that are associated with RNA polymerases and other molecular motors, indicating that intracellular mechanical forces might affect transcriptional regulation. In contrast to existing theory, we find that for loops <200 bp, the effect of tension is partly dependent on the relative orientation of the DNA-binding domains in the linker protein. Our results provide perspective on recent DNA looping experiments and suggestions for future micromechanical studies. PMID- 15653718 TI - Nano- to microscale dynamics of P-selectin detachment from leukocyte interfaces. I. Membrane separation from the cytoskeleton. AB - We have used a biomembrane force probe decorated with P-selectin to form point attachments with PSGL-1 receptors on a human neutrophil (PMN) in a calcium containing medium and then to quantify the forces experienced by the attachment during retraction of the PMN at fixed speed. From first touch to final detachment, the typical force history exhibited the following sequence of events: i), an initial linear-elastic displacement of the PMN surface, ii), an abrupt crossover to viscoplastic flow that signaled membrane separation from the interior cytoskeleton and the beginning of a membrane tether, and iii), the final detachment from the probe tip by usually one precipitous step of P-selectin:PSGL 1 dissociation. In this first article I, we focus on the initial elastic response and its termination by membrane separation from the cytoskeleton, initiating tether formation. Quantifying membrane unbinding forces for rates of loading (force/time) in the elastic regime from 240 pN/s to 38,000 pN/s, we discovered that the force distributions agreed well with the theory for kinetically limited failure of a weak bond. The kinetic rate for membrane unbinding was found to increase as an exponential function of the pulling force, characterized by an e fold scale in force of approximately 17 pN and a preexponential factor, or apparent unstressed off rate, of approximately 1/s. The rheological properties of tether growth subsequent to the membrane unbinding events are presented in a companion article II. PMID- 15653719 TI - Ligand accumulation in autocrine cell cultures. AB - Cell-culture assays are routinely used to analyze autocrine signaling systems, but quantitative experiments are rarely possible. To enable the quantitative design and analysis of experiments with autocrine cells, we develop a biophysical theory of ligand accumulation in cell-culture assays. Our theory predicts the ligand concentration as a function of time and measurable parameters of autocrine cells and cell-culture experiments. The key step of our analysis is the derivation of the survival probability of a single ligand released from the surface of an autocrine cell. An expression for this probability is derived using the boundary homogenization approach and tested by stochastic simulations. We use this expression in the integral balance equations, from which we find the Laplace transform of the ligand concentration. We demonstrate how the theory works by analyzing the autocrine epidermal growth factor receptor system and discuss the extension of our methods to other experiments with cultured autocrine cells. PMID- 15653720 TI - Experimental generation and computational modeling of intracellular pH gradients in cardiac myocytes. AB - It is often assumed that pH(i) is spatially uniform within cells. A double barreled microperfusion system was used to apply solutions of weak acid (acetic acid, CO(2)) or base (ammonia) to localized regions of an isolated ventricular myocyte (guinea pig). A stable, longitudinal pH(i) gradient (up to 1 pH(i) unit) was observed (using confocal imaging of SNARF-1 fluorescence). Changing the fractional exposure of the cell to weak acid/base altered the gradient, as did changing the concentration and type of weak acid/base applied. A diffusion reaction computational model accurately simulated this behavior of pH(i). The model assumes that H(i)(+) movement occurs via diffusive shuttling on mobile buffers, with little free H(+) diffusion. The average diffusion constant for mobile buffer was estimated as 33 x 10(-7) cm(2)/s, consistent with an apparent H(i)(+) diffusion coefficient, D(H)(app), of 14.4 x 10(-7) cm(2)/s (at pH(i) 7.07), a value two orders of magnitude lower than for H(+) ions in water but similar to that estimated recently from local acid injection via a cell-attached glass micropipette. We conclude that, because H(i)(+) mobility is so low, an extracellular concentration gradient of permeant weak acid readily induces pH(i) nonuniformity. Similar concentration gradients for weak acid (e.g., CO(2)) occur across border zones during regional myocardial ischemia, raising the possibility of steep pH(i) gradients within the heart under some pathophysiological conditions. PMID- 15653721 TI - Multilayer structures in lipid monolayer films containing surfactant protein C: effects of cholesterol and POPE. AB - The influence of cholesterol and POPE on lung surfactant model systems consisting of DPPC/DPPG (80:20) and DPPC/DPPG/surfactant protein C (80:20:0.4) has been investigated. Cholesterol leads to a condensation of the monolayers, whereas the isotherms of model lung surfactant films containing POPE exhibit a slight expansion combined with an increased compressibility at medium surface pressure (10-30 mN/m). An increasing amount of liquid-expanded domains can be visualized by means of fluorescence light microscopy in lung surfactant monolayers after addition of either cholesterol or POPE. At surface pressures of 50 mN/m, protrusions are formed which differ in size and shape as a function of the content of cholesterol or POPE, but only if SP-C is present. Low amounts of cholesterol (10 mol %) lead to an increasing number of protrusions, which also grow in size. This is interpreted as a stabilizing effect of cholesterol on bilayers formed underneath the monolayer. Extreme amounts of cholesterol (30 mol %), however, cause an increased monolayer rigidity, thus preventing reversible multilayer formation. In contrast, POPE, as a nonbilayer lipid thought to stabilize the edges of protrusions, leads to more narrow protrusions. The lateral extension of the protrusions is thereby more influenced than their height. PMID- 15653722 TI - Charge recombination and thermoluminescence in photosystem II. AB - In the recombination process of Photosystem II (S(2)Q(A)(-)-->S(1)Q(A)) the limiting step is the electron transfer from the reduced primary acceptor pheophytin Ph(-) to the oxidized primary donor P(+) and the rate depends on the equilibrium constant between states S(2)PPhQ(A)(-) and S(1)P(+)Ph(-)Q(A). Accordingly, mutations that affect the midpoint potential of Ph or of P result in a modified recombination rate. A strong correlation is observed between the effects on the recombination rate and on thermoluminescence (TL, the light emission from S(2)Q(A)(-) during a warming ramp): a slower recombination corresponds to a large enhancement and higher temperature of the TL peak. The current theory of TL does not account for these effects, because it is based on the assumption that the rate-limiting step coincides with the radiative process. When implementing the known fact that the radiative pathway represents a minor leak, the modified TL theory readily accounts qualitatively for the observed behavior. However, the peak temperature is still lower than predicted from the temperature-dependence of recombination. We argue that this reflects the heterogeneity of the recombination process combined with the enhanced sensitivity of TL to slower components. The recombination kinetics are accurately fitted as a sum of two exponentials and we show that this is not due to a progressive stabilization of the charge-separated state, but to a pre-existing conformational heterogeneity. PMID- 15653723 TI - The role of Phe in the formation of well-ordered oligomers of amyloidogenic hexapeptide (NFGAIL) observed in molecular dynamics simulations with explicit solvent. AB - We observed fast aggregation of partially ordered oligomers in an earlier simulation study of an amyloidogenic hexapeptide NFGAIL. In this work, the nucleation of highly ordered oligomers was further investigated by a combined total of 960 ns molecular dynamics simulations with explicit solvent on NFGAIL and its nonamyloidogenic mutant NAGAIL. In these simulations, four dimer subunits that each was constrained by harmonic forces as a two-strand beta-sheet were used to enhance the rate of formation. It was found that a critical role played by the aromatic residue Phe was to direct the stacking of beta-sheets to form ordered multilayer aggregates. We also found that many molecular arrangements of the peptide satisfied the "cross-beta-structure", a hallmark of amyloid fibrils. The tendency for the peptide to form either parallel or antiparallel beta-sheet was comparable, as was the tendency for the beta-sheets to stack either in parallel or antiparallel orientation. Overall, approximately 85% of the native hexapeptide formed octamers. The fact that only 8% of the octamers were well-ordered species suggests that the dissociation of the disordered oligomers be the rate-limiting step in the formation of highly ordered oligomers. Among the well-ordered subunit pairs, about half was formed by the beta-sheet extension along the main-chain hydrogen-bond direction, whereas the other half was formed by the beta-sheet stacking. Hence, a delicate balance between intersheet and intrasheet interactions appeared to be crucial in the formation of a highly ordered nucleus of amyloid fibrils. The disordered oligomers were mainly stabilized by nonspecific hydrophobic interactions, whereas the well-ordered oligomers were further stabilized by cross-strand hydrogen bonds and favorable side-chain stacking. PMID- 15653724 TI - Light scattering from intact cells reports oxidative-stress-induced mitochondrial swelling. AB - Angularly resolved light scattering measurements were performed on suspensions of EMT6 cells and on mitochondria isolated from rabbit liver. Mie theory analysis of the scattering from intact cells indicated that mitochondrial-sized organelles dominated scattering in the range 5-90 degrees . This interpretation was supported by the analysis of scattering from isolated mitochondria. Intact cells were subjected to oxidative stress by photodynamic insult. After 3 h of incubation in the heme precursor aminolevulinic acid hexylester, EMT6 cells accumulated abundant protoporphyrin IX, an endogenous photosensitizer formed in mitochondria. Irradiation of aminolevulinic acid/protoporphyrin IX-sensitized cells with 10 J cm(-2) of 514 nm light led to pronounced changes in angularly resolved light scattering consistent with mitochondrial swelling. Electron microscopy of similarly treated EMT6 cell monolayers showed significant changes in mitochondrial morphology, which included distension of the outer unit membrane and bloating of the internal mitochondrial compartment. Informed by these electron microscopy results, we implemented a coated sphere model to interpret the scattering from intact cells subjected to oxidative stress. The coated sphere interpretation was compatible with the scattering measurements from these cells, whereas simpler Mie theory models based on homogenous swelling were dramatically unsuccessful. Thus, in this system, angularly resolved light scattering reports oxidative-stress-induced changes in mitochondrial morphology. PMID- 15653725 TI - Accurate FRET measurements within single diffusing biomolecules using alternating laser excitation. AB - Fluorescence resonance energy transfer (FRET) between a donor (D) and an acceptor (A) at the single-molecule level currently provides qualitative information about distance, and quantitative information about kinetics of distance changes. Here, we used the sorting ability of confocal microscopy equipped with alternating laser excitation (ALEX) to measure accurate FRET efficiencies and distances from single molecules, using corrections that account for cross-talk terms that contaminate the FRET-induced signal, and for differences in the detection efficiency and quantum yield of the probes. ALEX yields accurate FRET independent of instrumental factors, such as excitation intensity or detector alignment. Using DNA fragments, we showed that ALEX-based distances agree well with predictions from a cylindrical model of DNA; ALEX-based distances fit better to theory than distances obtained at the ensemble level. Distance measurements within transcription complexes agreed well with ensemble-FRET measurements, and with structural models based on ensemble-FRET and x-ray crystallography. ALEX can benefit structural analysis of biomolecules, especially when such molecules are inaccessible to conventional structural methods due to heterogeneity or transient nature. PMID- 15653726 TI - A model for the Neurospora circadian clock. AB - Circadian clocks are important biological oscillators that generally involve two feedback loops. Here, we propose a new model for the Neurospora crassa circadian clock. First, we model its main negative feedback loop, including only experimentally well-documented reactions, the transcriptional activation of frequency (frq) by the white-collar complex (WCC), and the post-transcriptional dimerization of FRQ with WCC. This main loop is sufficient for oscillations and a similar one lies at the core of almost all known circadian clocks. Second, the model is refined to include the less characterized enhancement of white-collar 1 (WC-1) protein synthesis by FRQ, the positive second feedback loop. Numerical testing of different hypotheses led us to propose that the synthesis of WC-1 is enhanced by FRQ monomers and repressed by FRQ dimers. We demonstrate that this second loop contributes significantly to the robustness of the oscillator period against parameter variation. A phase response curve to light pulses is also computed and agrees well with experiments. On a general level, our results show that explicit time delays are not required for sustained oscillations but that it is crucial to take into account mRNA dynamics and protein-protein interactions. PMID- 15653727 TI - Single-molecule measurements of the persistence length of double-stranded RNA. AB - Over the past few years, it has become increasingly apparent that double-stranded RNA (dsRNA) plays a far greater role in the life cycle of a cell than previously expected. Numerous proteins, including helicases, polymerases, and nucleases interact specifically with the double helix of dsRNA. To understand the detailed nature of these dsRNA-protein interactions, the (bio)chemical, electrostatic, and mechanical properties of dsRNA need to be fully characterized. We present measurements of the persistence length of dsRNA using two different single molecule techniques: magnetic tweezers and atomic force microscopy. We deduce a mean persistence length for long dsRNA molecules of 63.8 +/- 0.7 nm from force extension measurements with the magnetic tweezers. We present atomic force microscopy images of dsRNA and demonstrate a new method for analyzing these, which yields an independent, yet consistent value of 62 +/- 2 nm for the persistence length. The introduction of these single-molecule techniques for dsRNA analysis opens the way for real-time, quantitative analysis of dsRNA protein interactions. PMID- 15653728 TI - Evidence for a highly elastic shell-core organization of cochlear outer hair cells by local membrane indentation. AB - Cochlear outer hair cells (OHCs) are thought to play an essential role in the high sensitivity and sharp frequency selectivity of the hearing organ by generating forces that amplify the vibrations of this organ at frequencies up to several tens of kHz. This tuning process depends on the mechanical properties of the cochlear partition, which OHC activity has been proposed to modulate on a cycle-by-cycle basis. OHCs have a specialized shell-core ultrastructure believed to be important for the mechanics of these cells and for their unique electromotility properties. Here we use atomic force microscopy to investigate the mechanical properties of isolated living OHCs and to show that indentation mechanics of their membrane is consistent with a shell-core organization. Indentations of OHCs are also found to be highly nonhysteretic at deformation rates of more than 40 microm/s, which suggests the OHC lateral wall is a highly elastic structure, with little viscous dissipation, as would appear to be required in view of the very rapid changes in shape and mechanics OHCs are believed to undergo in vivo. PMID- 15653729 TI - On the importance of the phosphocholine methyl groups for sphingomyelin/cholesterol interactions in membranes: a study with ceramide phosphoethanolamine. AB - In this study, we have examined how the headgroup size and properties affect the membrane properties of sphingomyelin and interactions with cholesterol. We prepared N-palmitoyl ceramide phosphoethanolamine (PCPE) and compared its membrane behavior with D-erythro-N-palmitoyl-sphingomyelin (PSM), both in monolayers and bilayers. The pure PCPE monolayer did not show a phase transition at 22 degrees C (in contrast to PSM), but displayed a much higher inverse isothermal compressibility as compared to the PSM monolayer, indicating stronger intermolecular interactions between PCPEs than between PSMs. At 37 degrees C the PCPE monolayer was more expanded (than at 22 degrees C) and displayed a rather poorly defined phase transition. When cholesterol was comixed into the monolayer, a condensing effect of cholesterol on the lateral packing of the lipids in the monolayer could be observed. The phase transition from an ordered to a disordered state in bilayer membranes was determined by diphenylhexatriene steady-state anisotropy. Whereas the PSM bilayer became disordered at 41 degrees C, the PCPE bilayer main transition occurred around 64 degrees C. The diphenylhexatriene steady-state anisotropy values were similar in both PCPE and PSM bilayers before and after the phase transition, suggesting that the order in the hydrophobic core in both bilayer types was rather similar. The emission from Laurdan was blue shifted in PCPE bilayers in the gel phase when compared to the emission spectra from PSM bilayers, and the blue-shifted component in PCPE bilayers was retained also after the phase transition, suggesting that Laurdan molecules sensed a more hydrophobic environment at the PCPE interface compared to the PSM interface both below and above the bilayer melting temperature. Whereas PSM was able to form sterol-enriched domains in dominantly fluid bilayers (as determined from cholestatrienol dequenching experiments), PCPE failed to form such domains, suggesting that the size and/or properties of the headgroup was important for stabilizing sphingolipid/sterol interaction. In conclusion, our study has highlighted how the headgroup in sphingomyelin affect its membrane properties and interactions with cholesterol. PMID- 15653730 TI - The 3-hydroxy group and 4,5-trans double bond of sphingomyelin are essential for modulation of galactosylceramide transmembrane asymmetry. AB - The structural features of SPM that control the transbilayer distribution of beta GalCer in POPC vesicles were investigated by (13)C- and (31)P-NMR spectroscopy using lipid analogs that share physical similarities with GalCer or SPM. The SPM analogs included N-palmitoyl-4,5-dihydro-SPM, 3-deoxy-SPM, 1-alkyl-2 amidophosphatidylcholine, and dipalmitoylphosphatidylcholine, a popular model "raft lipid". The transbilayer distributions of the SPM analogs and SPM in POPC vesicles were similar by (31)P-NMR. To observe the dramatic change in GalCer transbilayer distribution that occurs when SPM is included in POPC vesicles, the 3-OH group, 4,5-trans double bond, and amide linkage all were required in SPM. However, inclusion of 2 and 10 mol % dihydroSPM in SPM/POPC (1:1) vesicles mitigated and completely abrogated the effect of SPM on the transbilayer distribution of GalCer. Despite sharing some structural features with GalCer and localizing preferentially to the inner leaflet of POPC vesicles, dimyristoylphosphatidylethanolamine did not undergo a change in transbilayer distribution when SPM was incorporated into the vesicles. The results support the hypothesis that specific interactions may be favored among select sphingolipids in curvature-stressed membranes and emphasize the potential importance of the SPM dihydroSPM ratio in membrane fission and fusion processes associated with vesicle biogenesis and trafficking. PMID- 15653732 TI - Electrostatic interactions between the syntaxin membrane anchor and neurotransmitter passing through the fusion pore. AB - Recent experiments have shown that flux through the fusion pore is sensitive to manipulations of the side-chain size of certain residues in the syntaxin (syx) membrane anchor. These residues were proposed to line the wall of the fusion pore of Ca(2+)-triggered exocytosis. Here we continued this line of experimentation to examine possible electrostatic interactions between the pore lining residues and the neurotransmitter norepinephrine (NE). Replacing syx pore-lining residues with aspartate enhanced NE flux above that expected for the size of the aspartate side chain. In contrast, substitution with arginine reduced NE flux below that expected for the size of its side chain. Substituting aspartate and arginine into the nonpore-lining residues did not alter the fusion pore flux. Other amino acids with ionizable side chains had variable effects. These results indicate an electrostatic interaction between the pore-lining residues of syx and NE, and provide additional evidence that the syx membrane anchor is a structural component of the fusion pore. PMID- 15653731 TI - Evidence for a second binding/transport site for chloride in erythrocyte anion transporter AE1 modified at glutamate 681. AB - Transport kinetics have been examined in erythrocyte anion transporter AE1 that has been chemically modified to convert glutamate 681 to an alcohol (E681OH AE1). Outward conductive Cl(-) flux in E681OH AE1 is inhibited by removal of extracellular Cl(-); this effect is the opposite of that in native AE1 and is consistent with coupled electrogenic 2:1 Cl(-)/Cl(-) exchange. A second Cl(-) binding/transport site is also suggested by the characteristics of (35)SO(4)(2-) flux in E681OH AE1: bilateral and cis Cl(-), which are normally inhibitory, accelerate (35)SO(4)(2-) flux. These effects would be expected if Cl(-) binds to a second transport site on SO(4)(2-)-loaded E681OH AE1, thereby allowing Cl( )/SO(4)(2-) cotransport. Alternatively, the data can be explained without proposing Cl(-)/SO(4)(2-) cotransport if the rate-limiting event for (35)SO(4)(2 )/SO(4)(2-) exchange is external SO(4)(2-) release, and the binding of external Cl(-) accelerates SO(4)(2-) release. With either interpretation, these data indicate that E681OH AE1 has a binding/transport site for Cl(-) that is distinct from the main transport site. The effects of graded modification of E681 or inhibition by H(2)DIDS are consistent with the idea that the new Cl(-) binding site is on the same E681OH-modified subunit of the AE1 dimer as the normal transport site. PMID- 15653733 TI - The conversion of active to latent plasminogen activator inhibitor-1 is an energetically silent event. AB - PAI-1 is a proteinase inhibitor, which plays a key role in the regulation of fibrinolysis. It belongs to the serpins, a family of proteins that behave either as proteinase inhibitors or proteinase substrates, both reactions involving limited proteolysis of the reactive center loop and insertion of part of this loop into beta-sheet A. Titration calorimetry shows that the inhibition of tissue type plasminogen and pancreatic trypsin are exothermic reactions with DeltaH = 20.3, and -22.5 kcal.mol(-1), respectively. The Pseudomonas aeruginosa elastase catalyzed reactive center loop cleavage and inactivation of the inhibitor is also exothermic (DeltaH = -38.9 kcal.mol(-1)). The bacterial elastase also hydrolyses peptide-bound PAI-1 in which acetyl-TVASSSTA, the octapeptide corresponding to the P(14)-P(7) sequence of the reactive center loop is inserted into beta-sheet A of the serpin with DeltaH = -4.0 kcal.mol(-1). In contrast, DeltaH = 0 for the spontaneous conversion of the metastable active PAI-1 molecule into its thermodynamically stable inactive (latent) conformer although this conversion also involves loop/sheet insertion. We conclude that the active to latent transition of PAI-1 is an entirely entropy-driven phenomenon. PMID- 15653734 TI - Contribution of hydrophobicity to thermodynamics of ligand-DNA binding and DNA collapse. AB - The importance of understanding the dynamics of DNA condensation is inherent in the biological significance of DNA packaging in cell nuclei, as well as for gene therapy applications. Specifically, the role of ligand hydrophobicity in DNA condensation has received little attention. Considering that only multivalent cations can induce true DNA condensation, previous studies exploring monovalent lipids have been unable to address this question. In this study we have elucidated the contribution of the hydrophobic effect to multivalent cation- and cationic lipid-DNA binding and DNA collapse by studying the thermodynamics of cobalt hexammine-, spermine-, and lipospermine-plasmid DNA binding at different temperatures. Comparable molar heat capacity changes (DeltaC(p)) associated with cobalt hexammine- and spermine-DNA binding (-23.39 cal/mol K and -17.98 cal/mol K, respectively) suggest that upon binding to DNA, there are insignificant changes in the hydration state of the methylene groups in spermine. In contrast, the acyl chain contribution to the DeltaC(p) of lipospermine-DNA binding (DeltaC(p ) = DeltaC(p lipospermine) - DeltaC(p spermine)) is significant ( 220.94 cal/mol K). Although lipopermine induces DNA ordering into "tubular" suprastructures, such structures do not assume toroidal dimensions as observed for spermine-DNA complexes. We postulate that a steric barrier posed by the acyl chains in lipospermine precludes packaging of DNA into dimensions comparable to those found in nature. PMID- 15653735 TI - Nano- to microscale dynamics of P-selectin detachment from leukocyte interfaces. II. Tether flow terminated by P-selectin dissociation from PSGL-1. AB - We have used a biomembrane force probe decorated with P-selectin to form point attachments with PSGL-1 receptors on a human neutrophil (PMN) in a calcium containing medium and then to quantify the forces experienced by the attachment during retraction of the PMN at fixed speed. From first touch to final detachment, the typical force history exhibited the following sequence of events: i), an initial linear-elastic displacement of the PMN surface, ii), an abrupt crossover to viscoplastic flow that signaled membrane separation from the interior cytoskeleton and the beginning of a membrane tether, and iii), the final detachment from the probe tip most often by one precipitous step of P selectin:PSGL-1 dissociation. Analyzing the initial elastic response and membrane unbinding from the cytoskeleton in our companion article I, we focus in this article on the regime of tether extrusion that nearly always occurred before release of the extracellular adhesion bond at pulling speeds > or =1 microm/s. The force during tether growth appeared to approach a plateau at long times. Examined over a large range of pulling speeds up to 150 microm/s, the plateau force exhibited a significant shear thinning as indicated by a weak power-law dependence on pulling speed, f(infinity) = 60 pN(nu(pull)/microm/s)(0.25). Using this shear-thinning response to describe the viscous element in a nonlinear Maxwell-like fluid model, we show that a weak serial-elastic component with a stiffness of approximately 0.07 pN/nm provides good agreement with the time course of the tether force approach to the plateau under constant pulling speed. PMID- 15653736 TI - The effects of erythrocyte membranes on the nucleation of sickle hemoglobin. AB - Pathology in sickle cell disease begins with nucleation-dependent polymerization of deoxyhemoglobin S into stiff, rodlike fibers that deform and rigidify red cells. We have measured the effect of erythrocyte membranes on the rate of homogeneous nucleation in sickle hemoglobin, using preparations of open ghosts (OGs) with intact cytoskeletons from sickle (SS) and normal adult (AA) red cells. Nucleation rates were measured by inducing polymerization by laser photolysis of carboxy sickle hemoglobin and observing stochastic variation of replicate experiments of the time for the scattering signals to reach 10% of their respective maxima. By optical imaging of membrane fragments added to a hemoglobin solution we contrast the rate of nucleation immediately adjacent to membrane fragments with nucleation in a region of the same solution but devoid of membranes. From analysis of 29,272 kinetic curves obtained, we conclude that the effect of AA OGs is negligible (10% enhancement of nucleation rates +/-20%), whereas SS OGs caused 80% enhancement (+/-20%). In red cells, where more membrane surface is available to Hb, this implies enhancement of nucleation by a factor of 6. These experiments represent a 10-fold improvement in precision over previous approaches and are the first direct, quantitative measure of the impact of erythrocyte membranes on the homogeneous nucleation process that is responsible for polymer initiation in sickle cell disease. PMID- 15653738 TI - Position and orientation of phalloidin in F-actin determined by X-ray fiber diffraction analysis. AB - Knowledge of the phalloidin binding position in F-actin and the relevant understanding of the mechanism of F-actin stabilization would help to define the structural characteristics of the F-actin filament. To determine the position of bound phalloidin experimentally, x-ray fiber diffraction data were obtained from well-oriented sols of F-actin and the phalloidin-F-actin complex. The differences in the layer-line intensity distributions, which were clearly observed even at low resolution (8 A), produced well-resolved peaks corresponding to interphalloidin vectors in the cylindrically averaged difference-Patterson map, from which the radial binding position was determined to be approximately 10 A from the filament axis. Then, the azimuthal and axial positions were determined by single isomorphous replacement phasing and a cross-Patterson map in radial projection to be approximately 84 degrees and 0.5 A relative to the actin mass center. The refined position was close to the position found by prior researchers. The position of rhodamine attached to phalloidin in the rhodamine phalloidin-F-actin complex was also determined, in which the conjugated Leu(OH)(7) residue was found to face the outside of the filament. The position and orientation of the bound phalloidin so determined explain the increase in the interactions between long-pitch strands of F-actin and would also account for the inhibition of phosphate release, which might also contribute to the F-actin stabilization. The method of analysis developed in this study is applicable for the determination of binding positions of other drugs, such as jasplakinolide and dolastatin 11. PMID- 15653737 TI - Voltage-sensitive equilibrium between two states within a ryanoid-modified conductance state of the ryanodine receptor channel. AB - We have investigated the influence of transmembrane holding potential on the kinetics of interaction of a cationic ryanoid, 8beta-amino-9alpha hydroxyryanodine, with individual ryanodine receptor (RyR) channels and on the functional consequences of this interaction. In agreement with previous studies involving cationic, neutral, and anionic ryanoids, both rates of association and dissociation of the ligand are sensitive to transmembrane potential. A voltage sensitive equilibrium between high- and low-affinity forms of the receptor underlies alterations in rates of association and dissociation of the ryanoid. The interaction of 8beta-amino-9alpha-hydroxyryanodine with RyR influences the rate of cation translocation through the channel. With this ryanoid bound, the channel fluctuates between two clearly resolved subconductance states (alpha and beta). We interpret this observation as indicating that with 8beta-amino-9alpha hydroxyryanodine bound, the pore of the RyR channel exists in two essentially isoenergetic conformations with differing ion-handling properties. The equilibrium between the alpha- and beta-states of the RyR-8beta-amino-9alpha hydroxyryanodine complex is sensitive to transmembrane potential. However, the mechanisms determining this equilibrium differ from those responsible for the voltage-sensitive equilibrium between high- and low-affinity forms of the receptor. PMID- 15653740 TI - Structural locus of the pH gate in the Kir1.1 inward rectifier channel. AB - The closed-state crystal structure of prokaryotic inward rectifier, KirBac1.1, has implicated four inner helical phenylalanines near the cytoplasmic side as a possible locus of the channel gate. In the present study, we investigate whether this structural feature corresponds to the physiological pH gate of the renal inward rectifier, Kir1.1 (ROMK, KCNJ1). Kir1.1 is endogenous to the mammalian renal collecting duct and the thick ascending limb of Henle and is strongly gated by internal pH in the physiological range. It has four leucines (L160-Kir1.1b), homologous to the phenylalanines of KirBac1.1, which could function as steric gates near the convergence of the inner (M2) helices. Replacing these Leu-160 residues of Kir1.1b by smaller glycines abolished pH gating; however, replacement with alanines, whose side chains are intermediate in size between leucine and glycine, did not eliminate normal pH gating. Furthermore, a double mutant, constructed by adding the I163M-Kir1.1b mutation to the L160G mutation, also lacked normal pH gating, although the I163M mutation by itself enhanced the pH sensitivity of the channel. In addition to size, side-chain hydrophobicity at 160 Kir1.1b was also important for normal pH gating. Mutants with polar side chains (L160S, L160T) did not gate normally and were as insensitive to internal pH as the L160G mutant. Hence, either small or highly polar side chains at 160-Kir1.1b stabilize the open state of the channel. A homology model of the Kir1.1 closed state, based on the crystal structure of KirBac1.1, was consistent with our electrophysiological data and implies that closure of the Kir1.1 pH gate results from steric occlusion of the permeation path by the convergence of four leucines at the cytoplasmic apex of the inner transmembrane helices. In the open state, K crosses the pH gate together with its hydration shell. PMID- 15653741 TI - Monte Carlo studies of folding, dynamics, and stability in alpha-helices. AB - Folding simulations of polyalanine peptides were carried out using an off-lattice Monte Carlo simulation technique. The peptide was represented as a chain of residues, each of which contains two interaction sites: one corresponding to the C(alpha) atom and the other to the side chain. A statistical potential was used to describe the interaction between these sites. The preferred conformations of the peptide chain on the energy surface, starting from several initial conditions, were searched by perturbations on its generalized coordinates with the Metropolis criterion. We observed that, at low temperatures, the effective energy was low and the helix content high. The calculated helix propagation (s) and nucleation (sigma) parameters of the Zimm-Bragg model were in reasonable agreement with the empirical data. Exploration of the energy surface of the alanine-based peptides (AAQAA)(3) and AAAAA(AAARA)(3)A demonstrated that their behavior is similar to that of polyalanine, in regard to their effective energy, helix content, and the temperature-dependence of their helicity. In contrast, stable secondary structures were not observed for (Gly)(20) at similar temperatures, which is consistent with the nonfolder nature of this peptide. The fluctuations in the slowest dynamics mode, which describe the elastic behavior of the chain, showed that as the temperature decreases, the polyalanine peptides become stiffer and retain conformations with higher helix content. Clustering of conformations during the folding phase implied that polyalanine folds into a helix through fewer numbers of intermediate conformations as the temperature decreases. PMID- 15653739 TI - A vibrational spectral maker for probing the hydrogen-bonding status of protonated Asp and Glu residues. AB - Hydrogen bonding is a fundamental element in protein structure and function. Breaking a single hydrogen bond may impair the stability of a protein. We report an infrared vibrational spectral marker for probing the hydrogen-bond number for buried, protonated Asp or Glu residues in proteins. Ab initio computational studies were performed on hydrogen-bonding interactions of a COOH group with a variety of side-chain model compounds of polar and charged amino acids in vacuum using density function theory. For hydrogen-bonding interactions with polar side chain groups, our results show a strong correlation between the C=O stretching frequency and the hydrogen bond number of a COOH group: approximately 1759-1776 cm(-1) for zero, approximately 1733-1749 cm(-1) for one, and 1703-1710 cm(-1) for two hydrogen bonds. Experimental evidence for this correlation will be discussed. In addition, we show an approximate linear correlation between the C=O stretching frequency and the hydrogen-bond strength. We propose that a two-dimensional infrared spectroscopy, C=O stretching versus O-H stretching, may be employed to identify the specific type of hydrogen-bonding interaction. This vibrational spectral marker for hydrogen-bonding interaction is expected to enhance the power of time-resolved Fourier transform infrared spectroscopy for structural characterization of functionally important intermediates of proteins. PMID- 15653743 TI - Investigations into the membrane interactions of m-calpain domain V. AB - m-calpain is a calcium-dependent heterodimeric protease implicated in a number of pathological conditions. The activation of m-calpain appears to be modulated by membrane interaction, which has been predicted to involve oblique-orientated alpha-helix formation by a GTAMRILGGVI segment located in domain V of the protein's small subunit. Here, we have investigated this prediction. Fourier transform infrared conformational analysis showed that VP1, a peptide homolog of this segment, exhibited alpha-helicity of approximately 45% in the presence of dimyristoylphosphatidylcholine/dimyristoylphosphatidylserine (DMPS) vesicles. The level of helicity was unaffected over a 1- to 8-mM concentration range and did not alter when the anionic lipid composition of these vesicles was varied between 1% and 10% DMPS. Similar levels of alpha-helicity were observed in trifluoroethanol and the peptide appeared to adopt alpha-helical structure at an air/water interface with a molecular area of 164 A(2) at the monolayer collapse pressure. VP1 was found to penetrate dimyristoylphosphatidylcholine/DMPS monolayers, and at an initial surface pressure of 30 mN m(-1), the peptide induced surface pressure changes in these monolayers that correlated strongly with their anionic lipid content (maximal at 4 mN m(-1) in the presence of 10% DMPS). Neutron diffraction studies showed VP1 to be localized at the hydrophobic core of model palmitoyloleylphosphatidylcholine/palmitoyloleylphosphatidylserine (10:1 molar ratio) bilayer structures and, in combination, these results are consistent with the oblique membrane penetration predicted for the peptide. It would also appear that although not needed for structural stabilization anionic lipid was required for membrane penetration. PMID- 15653742 TI - Protein disorder: conformational distribution of the flexible linker in a chimeric double cellulase. AB - The structural properties of the linker peptide connecting the cellulose-binding module to the catalytic module in bimodular cellulases have been investigated by small-angle x-ray scattering. Since the linker and the cellulose-binding module are relatively small and cannot be readily detected separately, the conformation of the linker was studied by means of an artificial fusion protein, Cel6BA, in which an 88-residue linker connects the large catalytic modules of the cellulases Cel6A and Cel6B from Humicola insolens. Our data showed that Cel6BA is very elongated with a maximum dimension of 178 A, but could not be described by a single conformation. Modeling of a series of Cel6BA conformers with interdomain separations ranging between 10 A and 130 A showed that good Guinier and P(r) profile fits were obtained by a weighted average of the scattering curves of all the models where the linker follows a nonrandom distribution, with a preference for the more compact conformers. These structural properties are likely to be essential for the function of the linker as a molecular spring between the two functional modules. Small-angle x-ray scattering therefore provides a unique tool to quantitatively analyze the conformational disorder typical of proteins described as natively unfolded. PMID- 15653744 TI - Excitation energy transfer pathways in Lhca4. AB - EET in reconstituted Lhca4, a peripheral light-harvesting complex from Photosystem I of Arabidopsis thaliana, containing 10 chlorophylls and 2 carotenoids, was studied at room temperature by femtosecond transient absorption spectroscopy. Two spectral forms of Lut were observed in the sites L1 and L2, characterized by significantly different interactions with nearby chlorophyll a molecules. A favorable interpretation of these differences is that the efficiency of EET to Chls is about two times lower from the "blue" Lut in the site L1 than from the "red" Lut in the site L2 due to fast IC in the former case. A major part of the energy absorbed by the "red" Lut, approximately 60%-70%, is transferred to Chls on a sub-100-fs timescale from the state S(2) but, in addition, minor EET from the hot S(1) state within 400-500 fs is also observed. EET from the S(1) state to chlorophylls occurs also within 2-3 ps and is ascribed to Vio and/or "blue" Lut. EET from Chl b to Chl a is biphasic and characterized by time constants of approximately 300 fs and 3.0 ps. These rates are ascribed to EET from Chl b spectral forms absorbing at approximately 644 nm and approximately 650 nm, respectively. About 25% of the excited Chls a decays very fast-within approximately 15 ps. This decay is proposed to be related to the presence of the interacting Chls A5 and B5 located next to the carotenoid in the site L2 and may imply some photoprotective role for Lhca4 in the photosystem I super-complex. PMID- 15653745 TI - Proapoptotic triterpene electrophiles (avicins) form channels in membranes: cholesterol dependence. AB - Avicins, a family of triterpenoid saponins from Acacia victoriae, can regulate the innate stress response in human cells. Their ability to induce apoptosis in transformed cells makes them potential anticancer agents. We report that avicins can form channels in membranes. The conductance reached a steady state after each addition, indicating a dynamic equilibrium between avicin in solution and in the membrane. The high power dependence (up to 10) of the membrane conductance on the avicin concentration indicates the formation of multimeric channels, consistent with the estimated pore radius of 1.1 nm. This radius is too small to allow protein flux across the mitochondrial outer membrane, a process known to initiate apoptosis. Channel formation is lost when avicin's amphipathic side chain is removed, implicating this as the channel-forming region. A small difference in this side chain results in strong cholesterol dependence of channel formation in avicin G that is not found in avicin D. In neutral membranes, avicin channels are nonselective, but negatively-charged lipids confer cation selectivity (5:1, K(+):Cl(-)), indicating that phospholipids form part of the permeation pathway. Avicin channels in the mitochondrial outer membrane may favor apoptosis by altering the potential across this membrane and the intermembrane space pH. PMID- 15653747 TI - Assembly of lipoprotein particles containing apolipoprotein-B: structural model for the nascent lipoprotein particle. AB - Apolipoprotein B (apoB) is the major protein component of large lipoprotein particles that transport lipids and cholesterol. We have developed a detailed model of the first 1000 residues of apoB using standard sequence alignment programs (ClustalW and MACAW) and the MODELLER6 package for three-dimensional homology modeling. The validity of the apoB model was supported by conservation of disulfide bonds, location of all proline residues in turns and loops, and conservation of the hydrophobic faces of the two C-terminal amphipathic beta sheets, betaA (residues 600-763) and betaB (residues 780-1000). This model suggests a lipid-pocket mechanism for initiation of lipoprotein particle assembly. In a previous model we suggested that microsomal triglyceride transfer protein might play a structural role in completion of the lipid pocket. We no longer think this likely, but instead propose a hairpin-bridge mechanism for lipid pocket completion. Salt-bridges between four tandem charged residues (717 720) in the turn of the hairpin-bridge and four tandem complementary residues (997-1000) at the C-terminus of the model lock the bridge in the closed position, enabling the deposition of an asymmetric bilayer within the lipid pocket. PMID- 15653748 TI - 3-D particle tracking in a two-photon microscope: application to the study of molecular dynamics in cells. AB - We developed a method for tracking particles in three dimensions designed for a two-photon microscope, which holds great promise to study cellular processes because of low photodamage, efficient background rejection, and improved depth discrimination. During a standard cycle of the tracking routine (32 ms), the laser beam traces four circular orbits surrounding the particle in two z planes above and below the particle. The radius of the orbits is half of the x,y-width of the point spread function, and the distance between the z planes is the z width of the point spread function. The z-position is adjusted by moving the objective with a piezoelectric-nanopositioner. The particle position is calculated on the fly from the intensity profile obtained during the cycle, and these coordinates are used to set the scanning center for the next cycle. Applying this method, we were able to follow the motion of 500-nm diameter fluorescent polystyrene microspheres moved by a nanometric stage in either steps of 20-100 nm or sine waves of 0.1-10 microm amplitude with 20 nm precision. We also measured the diffusion coefficient of fluorospheres in glycerol solutions and recovered the values expected according to the Stokes-Einstein relationship for viscosities higher than 3.7 cP. The feasibility of this method for live cell measurements is demonstrated studying the phagocytosis of protein-coated fluorospheres by fibroblasts. PMID- 15653746 TI - Biophysical and kinetic characterization of HemAT, an aerotaxis receptor from Bacillus subtilis. AB - HemAT from Bacillus subtilis is a new type of heme protein responsible for sensing oxygen. The structural and functional properties of the full-length HemAT protein, the sensor domain (1-178), and Tyr-70 mutants have been characterized. Kinetic and equilibrium measurements reveal that both full-length HemAT and the sensor domain show two distinct O(2) binding components. The high-affinity component has a K(dissociation) approximately 1-2 microM and a normal O(2) dissociation rate constant, k(O2) = 50-80 s(-1). The low-affinity component has a K(dissociation) approximately 50-100 microM and a large O(2) dissociation rate constant equal to approximately 2000 s(-1). The low n-value and biphasic character of the equilibrium curve indicate that O(2) binding to HemAT involves either independent binding to high- and low-affinity subunits in the dimer or negative cooperativity. Replacement of Tyr-70(B10) with Phe, Leu, or Trp in the sensor domain causes dramatic increases in k(O2) for both the high- and low affinity components. In contrast, the rates and affinity for CO binding are little affected by loss of the Tyr-70 hydroxyl group. These results suggest highly dynamic behavior for the Tyr-70 side chain and the fraction of the "up" versus "down" conformation is strongly influenced by the nature of the iron ligand complex. As a result of having both high- and low-affinity components, HemAT can respond to oxygen concentration gradients under both hypoxic (0-10 microM) and aerobic (50-250 microM) conditions, a property which could, in principle, be important for a robust sensing system. The unusual ligand-binding properties of HemAT suggest that asymmetry and apparent negative cooperativity play an important role in the signal transduction pathway. PMID- 15653750 TI - Single gene effects in mouse models of host: pathogen interactions. AB - Inbred mouse strains have been known for many years to vary in their degree of susceptibility to different types of infectious diseases. The genetic basis of these interstrain differences is sometimes simple but often complex. In a few cases, positional cloning has been used successfully to identify single gene effects. The natural resistance-associated macrophage protein 1 (Nramp1) gene (Slc11a1) codes for a metal transporter active at the phagosomal membrane of macrophages, and Nramp1 mutations cause susceptibility to Mycobacterium, Salmonella, and Leishmania. Furthermore, recent advances in gene transfer technologies in transgenic mice have enabled the functional dissection of gene effects mapping to complex, repeated parts of the genome, such as the Lgn1 locus, causing susceptibility to Legionella pneumophila in macrophages. Finally, complex traits such as the genetically determined susceptibility to malaria can sometimes be broken down into multiple single gene effects. One such example is the case of pyruvate kinase, where a loss-of-function mutation was recently shown by our group to be protective against blood-stage infection with Plasmodium chabaudi. In all three cases reviewed, the characterization of the noted gene effect(s) has shed considerable light on the pathophysiology of the infection, including host response mechanisms. PMID- 15653749 TI - Repetitive transient depolarizations of the inner mitochondrial membrane induced by proton pumping. AB - Single mitochondria show the spontaneous fluctuations of DeltaPsim. In this study, to examine the mechanism of the fluctuations, we observed DeltaPsim in single isolated heart mitochondria using time-resolved fluorescence microscopy. Addition of malate, succinate, or ascorbate plus TMPD to mitochondria induced polarization of the inner membrane followed by repeated cycles of rapid depolarizations and immediate repolarizations. ADP significantly decreased the frequency of the rapid depolarizations, but the ADP effect was counteracted by oligomycin. On the other hand, the rapid depolarizations did not occur when mitochondria were polarized by the efflux of K(+) from the matrix. The rapid depolarizations became frequent with the increase in the substrate concentration or pH of the buffer. These results suggest that the rapid depolarizations depend on the net translocation of protons from the matrix. The frequency of the rapid depolarizations was not affected by ROS scavengers, Ca(2+), CsA, or BA. In addition, the obvious increase in the permeability of the inner membrane to calcein (MW 623) that was entrapped in the matrix was not observed upon the transient depolarization. The mechanisms of the spontaneous oscillations of DeltaPsim are discussed in relation to the matrix pH and the permeability transitions. PMID- 15653752 TI - K+ currents regulate the resting membrane potential, proliferation, and contractile responses in ventricular fibroblasts and myofibroblasts. AB - Despite the important roles played by ventricular fibroblasts and myofibroblasts in the formation and maintenance of the extracellular matrix, neither the ionic basis for membrane potential nor the effect of modulating membrane potential on function has been analyzed in detail. In this study, whole cell patch-clamp experiments were done using ventricular fibroblasts and myofibroblasts. Time- and voltage-dependent outward K(+) currents were recorded at depolarized potentials, and an inwardly rectifying K(+) (Kir) current was recorded near the resting membrane potential (RMP) and at more hyperpolarized potentials. The apparent reversal potential of Kir currents shifted to more positive potentials as the external K(+) concentration ([K(+)](o)) was raised, and this Kir current was blocked by 100-300 muM Ba(2+). RT-PCR measurements showed that mRNA for Kir2.1 was expressed. Accordingly, we conclude that Kir current is a primary determinant of RMP in both fibroblasts and myofibroblasts. Changes in [K(+)](o) influenced fibroblast membrane potential as well as proliferation and contractile functions. Recordings made with a voltage-sensitive dye, DiBAC(3)(4), showed that 1.5 mM [K(+)](o) resulted in a hyperpolarization, whereas 20 mM [K(+)](o) produced a depolarization. Low [K(+)](o) (1.5 mM) enhanced myofibroblast number relative to control (5.4 mM [K(+)](o)). In contrast, 20 mM [K(+)](o) resulted in a significant reduction in myofibroblast number. In separate assays, 20 mM [K(+)](o) significantly enhanced contraction of collagen I gels seeded with myofibroblasts compared with control mechanical activity in 5.4 mM [K(+)](o). In combination, these results show that ventricular fibroblasts and myofibroblasts express a variety of K(+) channel alpha-subunits and demonstrate that Kir current can modulate RMP and alter essential physiological functions. PMID- 15653751 TI - Down-regulation of normal human T cell blast activation: roles of APO2L/TRAIL, FasL, and c- FLIP, Bim, or Bcl-x isoform expression. AB - A systematic study was undertaken to characterize the role of APO 2 ligand/tumor necrosis factor-related apoptosis-inducing ligand (APO2L/TRAIL) and Fas ligand (FasL) together with the expression of several anti- or proapoptotic proteins in the down-regulation of normal human T cell responses. We have observed for the first time that the higher sensitivity of normal human T cell blasts to apoptosis and activation-induced cell death (AICD) as compared with naive T cells correlates with the increased expression of Bcl-x short (Bcl-xS) and Bim. T cell blasts die in the absence of interleukin 2 (IL-2) with no additional effect of death receptor ligation. In the presence of IL-2, recombinant APO2L/TRAIL or cytotoxic anti-Fas monoclonal antibodies induce rather inhibition of IL-2 dependent growth and not cell death on normal human T cell blasts. This observation is of physiological relevance, as supernatants from T cell blasts, pulse-stimulated with phytohemagglutinin (PHA) or through CD3 or CD59 ligation and containing bioactive APO2L/TRAIL and/or FasL expressed on microvesicles or direct CD3 or CD59 ligation, had the same effect. Cell death was only observed in the presence of cycloheximide or after a pulse through CD3 or CD59, correlating with a net reduction in cellular Fas-associated death domain-like IL-1beta converting enzyme-inhibitory protein long (c-FLIPL) and c-FLIPS expression. We also show that death receptor and free radical generation contribute, at least partially, to AICD induced by PHA and also to the inhibition of IL-2-dependent cell growth by CD3 or CD59 ligation. Finally, we have also shown that T cell blasts surviving PHA-induced AICD are memory CD44high cells with increased c FLIPS and Bcl-xL expression. PMID- 15653753 TI - Ephedrine plus caffeine causes age-dependent cardiovascular responses in Fischer 344 rats. AB - Human consumption of ephedrine and caffeine in dietary supplements has been associated with a number of adverse effects including changes in the ECG, myocardial infarction, hyperthermia, and, in rare instances, death. The purpose of this study was to investigate the potential mechanisms associated with the cardiotoxicity of combined ephedrine and caffeine ingestion. Seven- and fourteen week-old Fischer 344 rats treated with ephedrine in combination with caffeine exhibited increases in heart rate (HR), temperature, and corrected QT interval. Of the 14-wk-old rats treated with 25 mg/kg ephedrine plus 30 mg/kg caffeine, 57% died within 3-5 h of treatment, whereas none of the similarly treated 7-wk-old rats nor any of the rats treated with vehicle died. One hour after treatment with this dose of ephedrine plus caffeine, 14-wk-old rats exhibited a larger increase in HR (as % increase over baseline) than 7-wk-old rats. Furthermore, the 14-wk old rats that died had a higher HR and temperature than the 14-wk-old rats that lived. Histopathological studies suggested interstitial hemorrhage and myofiber necrosis in the 14-wk-old rats treated with the highest concentration of ephedrine and caffeine. This study showed enhanced susceptibility to ephedrine plus caffeine in 14-wk-old rats compared with 7-wk-old rats. The greater mortality in the 14-wk-old rats was associated with increases in body temperature, HR, and myocardial necrosis. PMID- 15653754 TI - Defective calcium handling in cardiomyocytes isolated from hearts subjected to ischemia-reperfusion. AB - Although ischemia-reperfusion (I/R) has been shown to affect subcellular organelles that regulate the intracellular Ca2+ concentration ([Ca2+]i), very little information regarding the Ca2+ handling ability of cardiomyocytes obtained from I/R hearts is available. To investigate changes in [Ca2+]i due to I/R, rat hearts in vitro were subjected to 10-30 min of ischemia followed by 5-30 min of reperfusion. Cardiomyocytes from these hearts were isolated and purified; [Ca2+]i was measured by employing fura-2 microfluorometry. Reperfusion for 30 min of the 20-min ischemic hearts showed attenuated cardiac performance, whereas basal [Ca2+]i as well as the KCl-induced increase in [Ca2+]i and isoproterenol (Iso) induced increase in [Ca2+]i in cardiomyocytes remained unaltered. On the other hand, reperfusion of the 30-min ischemic hearts for different periods revealed marked changes in cardiac function, basal [Ca2+]i, and Iso-induced increase in [Ca2+]i without any alterations in the KCl-induced increase in [Ca2+]i or S(-) BAY K 8644-induced increase in [Ca2+]i. The I/R-induced alterations in cardiac function, basal [Ca2+]i, and Iso-induced increase in [Ca2+]i in cardiomyocytes were attenuated by an antioxidant mixture containing superoxide dismutase and catalase as well as by ischemic preconditioning. The observed changes due to I/R were simulated in hearts perfused with H2O2 for 30 min. These results suggest that abnormalities in basal [Ca2+]i as well as mobilization of [Ca2+]i upon beta adrenoceptor stimulation in cardiomyocytes are dependent on the duration of ischemic injury to the myocardium. Furthermore, Ca2+ handling defects in cardiomyocytes appear to be mediated through oxidative stress in I/R hearts. PMID- 15653755 TI - Studies on rabbit natural and recombinant tissue factors: intracellular retention and regulation of surface expression in cultured cells. AB - Tissue factor (TF) is the most important trigger of blood coagulation in vascular pathology. Rabbit TF, with or without (delta C) its COOH-terminal intracellular tail, has been conjugated to green fluorescent protein (GFP) to study subcellular localization and other functions of TF. TF-GFP and TF delta C-GFP are associated with Na2CO3-resistant buoyant fractions in HEK-293 cells (lipid rafts); there is no morphological difference in the surface distribution of these or other GFP labeled membrane proteins present in or excluded from rafts (confocal microscopy, HEK-293 cells). Endogenous TF expressed by rabbit aortic smooth muscle cells (SMCs) is also raft associated. Membranes from HEK-293 cells expressing recombinant TF-GFP or wild-type TF were equipotent to clot human plasma; however, TF delta C-GFP was approximately 20-fold more active (per membrane weight). Immunoblot confirmed that the deletion mutant is more abundantly expressed, and confocal microscopy showed that it has preferential membrane localization, whereas TF-GFP is mainly intracellular (nuclear lining and multiple granules). With a similar half-life (<4 h), the two constructions differ by their intracellular retention, lower for TF delta C-GFP. In serum-starved SMCs, the expression of endogenous TF was upregulated by interleukin-1 beta and/or FBS treatment (immunoblot, immunofluorescence, clotting assay). However, TF secretion or surface expression was not regulated by stimuli of physiological intensity (such as stimulation of the coexpressed kinin B1 receptors), although a calcium ionophore was highly active in this respect. TF is a raft-associated molecule whose surface expression (secretion) is apparently retarded or impaired by structural determinant(s) located in its COOH-terminal tail. PMID- 15653756 TI - Serine 68 of phospholemman is critical in modulation of contractility, [Ca2+]i transients, and Na+/Ca2+ exchange in adult rat cardiac myocytes. AB - Overexpression of phospholemman (PLM) in normal adult rat cardiac myocytes altered contractile function and cytosolic Ca2+ concentration ([Ca2+]i) homeostasis and inhibited Na+/Ca2+ exchanger (NCX1). In addition, PLM coimmunoprecipitated and colocalized with NCX1 in cardiac myocyte lysates. In this study, we evaluated whether the cytoplasmic domain of PLM is crucial in mediating its effects on contractility, [Ca2+]i transients, and NCX1 activity. Canine PLM or its derived mutants were overexpressed in adult rat myocytes by adenovirus-mediated gene transfer. Confocal immunofluorescence images using canine-specific PLM antibodies demonstrated that the exogenous PLM or its mutants were correctly targeted to sarcolemma, t-tubules, and intercalated discs, with little to none detected in intracellular compartments. Overexpression of canine PLM or its mutants did not affect expression of NCX1, sarco(endo)plasmic reticulum Ca(2+)-ATPase, Na(+)-K(+)-ATPase, and calsequestrin in adult rat myocytes. A COOH-terminal deletion mutant in which all four potential phosphorylation sites (Ser62, Ser63, Ser68, and Thr69) were deleted, a partial COOH-terminal deletion mutant in which Ser68 and Thr69 were deleted, and a mutant in which all four potential phosphorylation sites were changed to alanine all lost wild-type PLM's ability to modulate cardiac myocyte contractility. These observations suggest the importance of Ser68 or Thr69 in mediating PLM's effect on cardiac contractility. Focusing on Ser68, the Ser68 to Glu mutant was fully effective, the Ser63 to Ala (leaving Ser68 intact) mutant was partially effective, and the Ser68 to Ala mutant was completely ineffective in modulating cardiac contractility, [Ca2+]i transients, and NCX1 currents. Both the Ser63 to Ala and Ser68 to Ala mutants, as well as PLM, were able to coimmunoprecipitate NCX1. It is known that Ser68 in PLM is phosphorylated by both protein kinases A and C. We conclude that regulation of cardiac contractility, [Ca2+]i transients, and NCX1 activity by PLM is critically dependent on Ser68. We suggest that PLM phosphorylation at Ser68 may be involved in cAMP- and/or protein kinase C dependent regulation of cardiac contractility. PMID- 15653757 TI - Warm ischemic preconditioning improves mitochondrial redox balance during and after mild hypothermic ischemia in guinea pig isolated hearts. AB - Ischemic preconditioning (IPC) induces distinctive changes in mitochondrial bioenergetics during warm (37 degrees C) ischemia and improves function and tissue viability on reperfusion. We examined whether IPC before 2 h of hypothermic (27 degrees C) ischemia affords additive cardioprotection and improves mitochondrial redox balance assessed by mitochondrial NADH and flavin adenine dinucleotide (FAD) autofluorescence in intact hearts. A mediating role of ATP-sensitive K(+) (K(ATP)) channel opening was investigated. NADH and FAD fluorescence was measured in the left ventricular wall of guinea pig isolated hearts assigned to five groups of eight animals each: hypothermia alone, hypothermia with ischemia, IPC with cold ischemia, 5-hydroxydecanoic acid (5-HD) alone, and 5-HD with IPC and cold ischemia. IPC consisted of two 5-min periods of warm global ischemia spaced 5 min apart and 15 min of reperfusion before 2 h of ischemia at 27 degrees C and 2 h of warm reperfusion. The K(ATP) channel inhibitor 5-HD was perfused from 5 min before until 5 min after IPC. IPC before 2 h of ischemia at 27 degrees C led to better recovery of function and less tissue damage on reperfusion than did 27 degrees C ischemia alone. These improvements were preceded by attenuated increases in NADH and decreases in FAD during cold ischemia and the reverse changes during warm reperfusion. 5-HD blocked each of these changes induced by IPC. This study indicates that IPC induces additive cardioprotection with mild hypothermic ischemia by improving mitochondrial bioenergetics during and after ischemia. Because effects of IPC on subsequent changes in NADH and FAD were inhibited by 5-HD, this suggests that mitochondrial K(ATP) channel opening plays a substantial role in improving mitochondrial bioenergetics throughout mild hypothermic ischemia and reperfusion. PMID- 15653759 TI - Alterations in wall shear stress predict sites of neointimal hyperplasia after stent implantation in rabbit iliac arteries. AB - Restenosis resulting from neointimal hyperplasia (NH) limits the effectiveness of intravascular stents. Rates of restenosis vary with stent geometry, but whether stents affect spatial and temporal distributions of wall shear stress (WSS) in vivo is unknown. We tested the hypothesis that alterations in spatial WSS after stent implantation predict sites of NH in rabbit iliac arteries. Antegrade iliac artery stent implantation was performed under angiography, and blood flow was measured before casting 14 or 21 days after implantation. Iliac artery blood flow domains were obtained from three-dimensional microfocal X-ray computed tomography imaging and reconstruction of the arterial casts. Indexes of WSS were determined using three-dimensional computational fluid dynamics. Vascular histology was unchanged proximal and distal to the stent. Time-dependent NH was localized within the stented region and was greatest in regions exposed to low WSS and acute elevations in spatial WSS gradients. The lowest values of WSS spatially localized to the stented area of a theoretical artery progressively increased after 14 and 21 days as NH occurred within these regions. This NH abolished spatial disparity in distributions of WSS. The results suggest that stents may introduce spatial alterations in WSS that modulate NH in vivo. PMID- 15653758 TI - Differential effects of 17beta-estradiol, conjugated equine estrogen, and raloxifene on mRNA expression, aggregation, and secretion in platelets. AB - Changes in platelet functions could contribute to thrombotic risk associated with estrogen treatments. This study was designed to test the hypothesis that three clinically relevant estrogenic treatments affect platelet function comparably. Adult female pigs were ovariectomized and randomized to either no treatment or treatment with oral 17 beta-estradiol (2 mg/day), conjugated equine estrogen (0.625 mg/day), or raloxifene (60 mg/day) for 4 wk. Platelet turnover, aggregation, and secretion were assessed before and after treatment. Platelet turnover and mRNA increased significantly only in pigs treated with 17 beta estradiol. Expression of estrogen receptors increased with ovariectomy and decreased with all treatments. Platelet aggregation and secretion of ATP, platelet-derived growth factor, and matrix metalloproteinase-2 increased with ovariectomy. All treatments reduced both aggregation and secretion. Expression of mRNA for constitutive endothelial nitric oxide synthase (eNOS), but not eNOS protein, increased with ovariectomy. Only eNOS mRNA decreased with all treatments, but only treatment with 17 beta-estradiol increased secretion of nitric oxide from intact platelets. Platelets from 17 beta-estradiol-treated animals caused relaxation of coronary arteries, which was sensitive to inhibition of nitric oxide. Although three different estrogenic treatments reversed increases in platelet aggregation caused by ovariectomy, only 17 beta-estradiol increased platelet RNA and release of platelet-derived nitric oxide. These differences reflect transcriptional and posttranscriptional regulation of protein synthesis in bone marrow megakaryocytes and circulating platelets. PMID- 15653760 TI - Low-pressure reperfusion alters mitochondrial permeability transition. AB - We hypothesized that low-pressure reperfusion may limit myocardial necrosis and attenuate postischemic contractile dysfunction by inhibiting mitochondrial permeability transition pore (mPTP) opening. Male Wistar rat hearts (n = 36) were perfused according to the Langendorff technique, exposed to 40 min of ischemia, and assigned to one of the following groups: 1) reperfusion with normal pressure (NP = 100 cmH(2)O) or 2) reperfusion with low pressure (LP = 70 cmH(2)O). Creatine kinase release and tetraphenyltetrazolium chloride staining were used to evaluate infarct size. Modifications of cardiac function were assessed by changes in coronary flow, heart rate (HR), left ventricular developed pressure (LVDP), the first derivate of the pressure curve (dP/dt), and the rate-pressure product (RPP = LVDP x HR). Mitochondria were isolated from the reperfused myocardium, and the Ca(2+)-induced mPTP opening was measured using a potentiometric approach. Lipid peroxidation was assessed by measuring malondialdehyde production. Infarct size was significantly reduced in the LP group, averaging 17 +/- 3 vs. 33 +/- 3% of the left ventricular weight in NP hearts. At the end of reperfusion, functional recovery was significantly improved in LP hearts, with RPP averaging 10,392 +/- 876 vs. 3,969 +/- 534 mmHg/min in NP hearts (P < 0.001). The Ca(2+) load required to induce mPTP opening averaged 232 +/- 10 and 128 +/- 16 microM in LP and NP hearts, respectively (P < 0.001). Myocardial malondialdehyde was significantly lower in LP than in NP hearts (P < 0.05). These results suggest that the protection afforded by low-pressure reperfusion involves an inhibition of the opening of the mPTP, possibly via reduction of reactive oxygen species production. PMID- 15653761 TI - Adipokines: molecular links between obesity and atheroslcerosis. AB - Atherosclerotic disease remains the leading cause of death in industrialized nations despite major advances in its diagnosis, treatment, and prevention. The increasing epidemic of obesity, insulin resistance, and diabetes will likely add to this burden. Increasingly, it is becoming apparent that adipose tissue is an active endocrine and paracrine organ that releases several bioactive mediators that influence not only body weight homeostasis but also inflammation, coagulation, fibrinolysis, insulin resistance, diabetes, and atherosclerosis. The cellular mechanisms linking obesity and atherosclerosis are complex and have not been fully elucidated. This review summarizes the experimental and clinical evidence on how excess body fat influences cardiovascular health through multiple yet converging pathways. The role of adipose tissue in the development of obesity linked insulin resistance, metabolic syndrome, and diabetes will be reviewed, including an examination of the molecular links between obesity and atherosclerosis, namely, the effects of fat-derived adipokines. Finally, we will discuss how these new insights may provide us with innovative therapeutic strategies to improve cardiovascular health. PMID- 15653762 TI - In vivo adenosine receptor preconditioning reduces myocardial infarct size via subcellular ERK signaling. AB - The protective effects of adenosine receptor acute preconditioning (PC) are well known; however, the signaling mechanism mediating this effect has not been determined in in vivo models. The purpose of this study was to determine the role of the extracellular signal-regulated kinase (ERK) pathway in mediating adenosine PC in in vivo rat myocardium. Open-chest rats were submitted to 25 min of coronary artery occlusion and 2 h of reperfusion. ERK activation was assessed by measuring total and dually phosphorylated p44/42 ERK isoforms in nuclear and/or myofilament, mitochondrial, cytosolic, and membrane fractions. Adenosine receptor PC with the A1/A2a agonist 1S-[1a,2b,3b,4a(S*)]-4-[7-[[2-(3-chloro-2-thienyl)-1 methylpropyl]amino]-3H-imidazo[4,5-b]pyridyl-3-yl]cyclopentane carboxamide (AMP 579) reduced infarct size from 49 +/- 3% to 29 +/- 3%, an effect that was blocked by the mitogen-activated protein kinase-ERK inhibitor U-0126. ERK isoforms were present in all fractions, with the greatest expression in the cytosolic fraction and the least in the mitochondrial fraction. AMP-579 treatment increased preischemic p44/42 ERK phosphorylation in all fractions 2.7- to 6.9-fold. Reperfusion increased ERK isoform activation in all fractions, but there were no differences between control and AMP-579 hearts. Preischemic increases in phospo p44/p42 ERK with AMP-579 were blunted by U-0126, although only in mitochondrial and membrane compartments. The PC effects of AMP-579 on infarct size and ERK were blunted by both the A1 antagonist 8-cyclopentyl-1,3-dipropylxanthine and, surprisingly, the A2a antagonist ZM-241385. These results indicate that the unique adenosine receptor agonist AMP-579 exerts its beneficial effects in vivo via both A1 and A2a receptor modulation of subcellular ERK isoform signaling. PMID- 15653763 TI - Analysis of nitric oxide donor effectiveness in resistance vessels. AB - Decreased nitric oxide (NO) bioavailability is associated with a number of pathological conditions. Administration of a supplemental source of NO can counter the pathological effects arising from decreased NO bioavailability. A class of NO-nucleophile adducts that spontaneously release NO (NONOates) has been developed, and its members show promise as therapeutic sources of NO. Because the NONOates release NO spontaneously, a significant portion of the NO may be consumed by the myriad of NO reactive species present in the body. Here we develop a model to analyze the efficacy of NO delivery, by membrane-impermeable NONOates, in the resistance arterioles. Our model identifies three features of blood vessels that will enhance NONOate efficacy: 1) the amount of NO delivered to the abluminal region increases with lumen radius; 2) the presence of a flow induced red blood cell-free zone will augment NO delivery; and 3) extravasation of the NONOate into the interstitial space will increase abluminal NO delivery. These results suggest that NONOates may be more effective in larger vessels and that NONOate efficacy can be altered by modifying permeability to the interstitial space. PMID- 15653764 TI - Coronary arteriolar vasoconstriction to angiotensin II is augmented in prediabetic metabolic syndrome via activation of AT1 receptors. AB - The metabolic syndrome is associated with activation of the renin-angiotensin system. However, whether the coronary vascular response to ANG II is altered under this condition is unknown. Experiments were conducted in control and chronically high-fat-fed dogs with the prediabetic metabolic syndrome both in vitro (isolated coronary arterioles, 60-110 microm) and in vivo (anesthetized and conscious). We found that plasma renin activity and ANG II levels are elevated in high-fat-fed dogs and that this increase in ANG II is associated with a significant increase in ANG II-mediated coronary vasoconstriction in isolated coronary arterioles and in anesthetized open-chest dogs. The vasoconstriction to ANG II is abolished by ANG II type 1 (AT1) receptor blockade. In conscious chronically instrumented dogs, AT1 receptor blockade with telmisartan improved the balance between coronary blood flow and myocardial oxygen consumption in the high-fat-fed dogs but not in normal control dogs, i.e., the relationship between coronary venous Po2 and myocardial oxygen consumption was shifted upward, toward normal control values. Quantitative assessment of coronary arteriolar AT1 and ANG II type 2 (AT2) receptor mRNA levels by real-time PCR revealed no significant difference between normal control and high-fat-fed dogs; however, Western blot analysis showed a significant increase in AT1 receptor protein level with no change in AT2 receptor protein density. These findings indicate that AT1 receptor mediated coronary constriction is augmented in the prediabetic metabolic syndrome and contributes to impaired control of coronary blood flow via increases in circulating ANG II and/or coronary arteriolar AT1 receptor density. PMID- 15653765 TI - Extending the cardioprotective window using a novel delta-opioid agonist fentanyl isothiocyanate via the PI3-kinase pathway. AB - Selective delta-opioid agonists produce delayed cardioprotection that lasts for 24-48 h in rats; however, the maximum length of the cardioprotective window is unclear. In this study, we attempted to prolong the cardioprotective window using a unique delta-opioid agonist, fentanyl isothiocyanate (FIT), which binds irreversibly to the delta-receptor, and determined the role of the phosphatidylinositol 3-kinase (PI3K) pathway as a trigger or end effector of FIT induced cardioprotection. Initially, male rats were administered FIT (10 microg/kg) 10 min before hearts were subjected to 30 min of ischemia and 2 h of reperfusion followed by infarct size (IS) assessment. Acute FIT administration reduced IS when given before ischemia, 5 min before reperfusion, or 10 s after reperfusion compared with control. IS reduction also occurred following a single dose of FIT at 48, 72, 96, and 120 h after administration vs. control, with the maximum effect observed at 96 h. FIT-induced IS reduction at 96 h was completely abolished when the irreversible PI3K inhibitor wortmannin (15 microg/kg) was given before FIT during the trigger phase; however, the effect was only partially abrogated when wortmannin was given 96 h later. These data suggest that FIT has a prolonged cardioprotective window greater than that of any previously described cardioprotective agent that requires PI3K primarily in the trigger phase but also partially, as a mediator or end effector. PMID- 15653766 TI - Involvement of p38-mitogen-activated protein kinase in adenosine receptor mediated relaxation of coronary artery. AB - The purpose of this study was to explore the involvement of adenosine receptor(s) in porcine coronary artery (PCA) relaxation and to define the role of MAPK signaling pathways. Isometric tensions were recorded in denuded PCA rings. 5'-(N ethylcarboxamido)adenosine (NECA), a nonselective adenosine receptor agonist, induced a concentration-dependent relaxation (EC(50) = 16.8 nM) of PGF(2alpha) (10 microM)-preconstricted arterial rings. NECA-induced relaxation was completely blocked by 0.1 microM SCH-58261 (A(2A) antagonist) at lower doses (1-40 nM) but not at higher doses (80-1,000 nM). MRS-1706 (1 microM, A(2B) antagonist) was able to shift the NECA concentration-response curve to the right. CGS-21680 (selective A(2A) agonist) induced responses similarly to NECA, whereas N(6) cyclopentyladenosine (A(1) agonist) and Cl-IB-MECA (A(3) agonist) did not. Furthermore, the effect of NECA was attenuated by the addition of SB-203580 (10 microM, p38 MAPK inhibitor) but not by PD-98059 (10 microM, MEK inhibitor). Interestingly, SB-203580 had no effect on CGS-21680-induced relaxation. Western blot analysis demonstrated that PGF(2alpha) and adenosine agonists stimulated p38 MAPK at a concentration of 40 nM in PCA smooth muscle cells. MRS-1706 (1 microM) significantly reduced NECA-induced p38 MAPK phosphorylation. Addition of NECA and SB-203580 alone or in combination inhibited PGF(2alpha)-induced p38 MAPK. Western blot data were further confirmed by p38 MAPK activity measurement using activating transcription factor-2 assay. Our results suggest that the adenosine receptor subtype involved in causing relaxation of porcine coronary smooth muscle is mainly A(2A) subtype, although A(2B) also may play a role, possibly through p38 MAPK pathway. PMID- 15653767 TI - Antiatherogenic potential of red wine: clinician update. AB - Complications of atherosclerosis remain the leading cause of morbidity and mortality in industrialized countries. Epidemiological studies have repeatedly demonstrated that moderate alcohol intake has a beneficial effect on cardiovascular disease. The purpose of this review is to examine the epidemiological and biological evidence supporting the intake of red wine as a means of reducing atherosclerosis. On the basis of epidemiological studies, moderate intake of alcoholic beverages, including red wine, reduces the risk of cardiovascular, cerebrovascular, and peripheral vascular disease in populations. In addition to the favorable biological effects of alcohol on the lipid profile, on hemostatic factors, and in reducing insulin resistance, the phenolic compounds in red wine appear to interfere with the molecular processes underlying the initiation, progression, and rupture of atherosclerotic plaques. Whether red wine is more beneficial than other types of alcohol remains unclear. Definitive data from a large-scale, randomized clinical end-point trial of red wine intake would be required before physicians can advise patients to use wine as part of preventative or medical therapies. PMID- 15653768 TI - Exercise training improves endogenous nitric oxide mechanisms within the paraventricular nucleus in rats with heart failure. AB - Previously, we have demonstrated that an altered endogenous nitric oxide (NO) mechanism within the paraventricular nucleus (PVN) contributes to increased renal sympathetic nerve activity (RSNA) in heart failure (HF) rats. The goal of this study was to examine the effect of exercise training (ExT) in improving the endogenous NO mechanism within the PVN involved in the regulation of RSNA in rats with HF. ExT significantly restored the decreased number of neuronal NO synthase (nNOS)-positive neurons in the PVN (129 +/- 17 vs. 99 +/- 6). nNOS mRNA expression and protein levels in the PVN were also significantly increased in HF ExT rats compared with HF-sedentary rats. To examine the functional role of NO within the PVN, an inhibitor of NOS, N(G)-monomethyl-L-arginine, was microinjected into the PVN. Dose-dependent increases in RSNA, arterial blood pressure (BP), and heart rate (HR) were produced in all rats. There was a blunted increase in these parameters in HF rats compared with the sham-operated rats. ExT significantly augmented RSNA responses in rats with HF (33% vs. 20% at the highest dose), thus normalizing the responses. The NO donor sodium nitroprusside, microinjected into the PVN, produced dose-dependent decreases in RSNA, BP, and HR in both sham and HF rats. ExT significantly improved the blunted decrease in RSNA in HF rats (36% vs. 17% at the highest dose). In conclusion, our data indicate that ExT improves the altered NO mechanism within the PVN and restores NO mediated changes in RSNA in rats with HF. PMID- 15653769 TI - Rhabdomeric phototransduction initiated by the vertebrate photopigment melanopsin. AB - Melanopsin is the photopigment that confers light sensitivity on intrinsically photosensitive retinal ganglion cells. Mammalian intrinsically photosensitive retinal ganglion cells are involved in the photic synchronization of circadian rhythms to the day-night cycle. Here, we report molecular components of melanopsin signaling using the cultured Xenopus dermal melanophore system. Photo activated melanopsin is shown to initiate a phosphoinositide signaling pathway similar to that found in invertebrate photo-transduction. In melanophores, light increases the intracellular level of inositol trisphosphate and causes the dispersion of melanosomes. Inhibition of phospholipase C and protein kinase C and chelation of intracellular calcium block the effect of light on melanophores. At least four proteins, 43, 74, 90, and 134 kDa, are phosphorylated by protein kinase C upon light stimulation. This provides evidence of an invertebrate-like light-activated signaling cascade within vertebrate cells. PMID- 15653770 TI - Characterization of potential selenium-binding proteins in the selenophosphate synthetase system. AB - Selenophosphate, an activated form of selenium that can serve as a selenium donor, is generated by the selD gene product, selenophosphate synthetase (SPS). Selenophosphate is required by several bacteria and by mammals for the specific synthesis of Secys-tRNA, the precursor of selenocysteine in selenoenzymes. Although free selenide can be used in vitro for synthesis of selenophosphate, the physiological system that donates selenium to SPS is incompletely characterized. To detect potential selenium-delivery proteins, two known sulfurtransferases and glyceraldehyde-3-phosphate dehydrogenase (GAPDH; EC 1.2.1.12) were analyzed for ability to bind and transfer selenium. Rhodanese (EC 2.8.1.1) was shown to bind selenium tightly, with only part of the selenium being available as substrate for SPS in the presence of added reductant. 3-Mercaptopyruvate sulfurtransferase (3 MST; EC 2.8.1.2) and GAPDH also bound selenium supplied as selenodiglutathione formed from SeO3(2-) and glutathione. Selenium bound to 3-MST and GAPDH was released more readily than that from rhodanese and also was more available as a substrate for SPS. Although rhodanese retained tightly bound selenium under aerobic conditions, the protein gradually became insoluble, whereas GAPDH containing bound selenium was stable at neutral pH for a long period. These results indicate that 3-MST and GAPDH have more suitable potentials as a physiological selenium-delivery protein than rhodanese. In the presence of a selenium-binding protein, a low level of selenodiglutathione formed from SeO3(2-) and glutathione could effectively replace the high concentrations of selenide routinely used as substrate in the SPS in vitro assays. PMID- 15653771 TI - Seasonal epidemics of cholera inversely correlate with the prevalence of environmental cholera phages. AB - The relationship among (i) the local incidence of cholera, (ii) the prevalence in the aquatic environment of Vibrio cholerae, and (iii) bacterial viruses that attack potentially virulent O1 and O139 serogroup strains of this organism (cholera phages) was studied in Dhaka, Bangladesh. Over nearly a 3-year period, we found that significantly more environmental water samples contained either a phage or a phage-susceptible V. cholerae strain than both (P < 0.00001). The number of cholera patients varied seasonally during this period and frequently coincided with the presence of pathogenic V. cholerae strains in water samples that otherwise lacked detectable cholera phages. Interepidemic periods were characterized by water samples containing cholera phages but no viable bacteria. Our data support the conclusion that cholera phages can influence cholera seasonality and may also play a role in emergence of new V. cholerae pandemic serogroups or clones. PMID- 15653772 TI - Directing cell migration with asymmetric micropatterns. AB - This report shows that the direction of polarization of attached mammalian cells determines the direction in which they move. Surfaces micropatterned with appropriately functionalized self-assembled monolayers constrain individual cells to asymmetric geometries (for example, a teardrop); these geometries polarize the morphology of the cell. After electrochemical desorption of the self-assembled monolayers removes these constraints and allows the cells to move across the surface, they move toward their blunt ends. PMID- 15653774 TI - The protein structure prediction problem could be solved using the current PDB library. AB - For single-domain proteins, we examine the completeness of the structures in the current Protein Data Bank (PDB) library for use in full-length model construction of unknown sequences. To address this issue, we employ a comprehensive benchmark set of 1,489 medium-size proteins that cover the PDB at the level of 35% sequence identity and identify templates by structure alignment. With homologous proteins excluded, we can always find similar folds to native with an average rms deviation (RMSD) from native of 2.5 A with approximately 82% alignment coverage. These template structures often contain a significant number of insertions/deletions. The tasser algorithm was applied to build full-length models, where continuous fragments are excised from the top-scoring templates and reassembled under the guide of an optimized force field, which includes consensus restraints taken from the templates and knowledge-based statistical potentials. For almost all targets (except for 2/1,489), the resultant full-length models have an RMSD to native below 6 A (97% of them below 4 A). On average, the RMSD of full-length models is 2.25 A, with aligned regions improved from 2.5 A to 1.88 A, comparable with the accuracy of low-resolution experimental structures. Furthermore, starting from state-of-the-art structural alignments, we demonstrate a methodology that can consistently bring template-based alignments closer to native. These results are highly suggestive that the protein-folding problem can in principle be solved based on the current PDB library by developing efficient fold recognition algorithms that can recover such initial alignments. PMID- 15653773 TI - Imaging bacterial infections with radiolabeled 1-(2'-deoxy-2'-fluoro-beta-D arabinofuranosyl)-5-iodouracil. AB - Bacterial infections provide diagnostic dilemmas that could be enlightened by modern imaging technologies. We have developed a simple method for imaging bacterial infections in mice that relies on the phosphorylation and trapping of the thymidine kinase (TK) substrate 1-(2'-deoxy-2'-fluoro-beta-D arabinofuranosyl)-5-[125I] iodouracil ([125I]FIAU) within bacteria. FIAU was found to inhibit the growth of WT Escherichia coli but not a TK- strain, indicating that WT E. coli could metabolize this compound. In silico analyses demonstrated that all pathogenic strains of bacteria whose genomes have been sequenced contain a TK gene highly homologous to the E. coli TK. Accordingly, we demonstrated that localized infections caused by representatives of five genera of bacteria could be readily imaged with [125I]FIAU. Such imaging provides a general method for the diagnosis of localized bacterial infections that could be translatable to the clinic. PMID- 15653777 TI - DO comments on bipolar disorder supplement. PMID- 15653775 TI - Elicitation of allergic asthma by immunoglobulin free light chains. AB - The observation that only 50% of patients with adult asthma manifest atopy indicates that other inflammatory mechanisms are likely involved in producing the characteristic features of this disorder; namely reversible airway obstruction, hyperresponsiveness, and pulmonary inflammation. Our recent discovery that antigen-specific Ig free light chains (LCs) mediate hypersensitivity-like responses suggests that these molecules may be of import in the pathophysiology of asthma. Using a murine experimental model of nonatopic asthma, we now have shown that an LC antagonist, the 9-mer peptide F991, can abrogate the development of airway obstruction, hyperresponsiveness, and pulmonary inflammation. Further, passive immunization with antigen-specific LCs and subsequent airway challenge can elicit a mast cell-dependent reaction leading to acute bronchoconstriction. These findings, and the demonstration that the concentration of free kappa LCs in the sera of patients with adult asthma were significantly increased (as compared with age-matched nonasthmatic individuals), provide previously undescribed insight into the pathogenesis of asthma. In addition, the ability to inhibit pharmacologically LC-induced mast cell activation provides a therapeutic means to prevent or ameliorate the adverse bronchopulmonary manifestations of this incapacitating disorder. PMID- 15653779 TI - The newer guidelines for the management of community-acquired pneumonia. AB - Community-acquired pneumonia (CAP) is a leading cause of death in the world and the sixth most common cause of death in the United States. It is the number one cause of death from infectious diseases in the United States. This article reviews the latest available guidelines from two leading organizations-the Infectious Diseases Society of America (IDSA) and the American Thoracic Society (ATS). The IDSA stratifies patients into three categories and recommends antibiotic management based on assigned categories: outpatients, patients admitted to a general medical floor (GMF), and patients requiring intensive care unit (ICU) admission. The ATS, in contrast, stratifies patients into four major groups based on the presence of two cardiopulmonary diseases, certain modifying risk factors that increase the likelihood of acquiring specific infections (such as with drug-resistant Streptococcus pneumoniae, enteric gram-negative organisms, or Pseudomonas aeruginosa), and also based on the site of treatment (such as outpatient setting, GMF, and ICU). PMID- 15653780 TI - Sexually transmitted infections and increased risk of co-infection with human immunodeficiency virus. AB - The incidence of trichomoniasis (Trichomonas vaginalis) in the United States is estimated at 5 million cases annually; chlamydia (Chlamydia trachomatis) at 3 million; gonorrhea (Neisseria gonorrhoeae), 650,000; and syphilis (Treponema pallidum), 70,000. However, most sexually transmitted infections (STIs) are asymptomatic-contributing to underdiagnosis estimated at 50% or more. Diagnosis of an STI signals sexual health risk because an STI facilitates the transmission and acquisition of other STIs, including human immunodeficiency virus (HIV). In fact, comorbid STIs increase patients' susceptibility of acquiring and transmitting HIV by two- to fivefold. Several studies have shown that aggressive STI prevention, testing, and treatment reduces the transmission of HIV. The authors discuss common clinical presentations, screening, diagnosis, and treatment for trichomoniasis, chlamydia, gonorrhea, syphilis, and herpes simplex virus. PMID- 15653781 TI - Maternal and fetal outcomes of spontaneous preterm premature rupture of membranes. AB - The authors retrospectively evaluated maternal and fetal outcomes of 73 consecutive singleton pregnancies complicated by preterm premature rupture of amniotic membranes. When preterm labor occurred and fetuses were at a viable gestational age, pregnant patients were managed aggressively with tocolytic therapy, antenatal corticosteroid injections, and antenatal fetal testing. The mean gestational age at the onset of membrane rupture and delivery was 22.1 weeks and 23.8 weeks, respectively. The latency from membrane rupture to delivery ranged from 0 to 83 days with a mean of 8.6 days. Among the 73 pregnant patients, there were 22 (30.1%) stillbirths and 13 (17.8%) neonatal deaths, resulting in a perinatal death rate of 47.9%. The perinatal survival rate based on gestational age at the onset of fetal membrane rupture was 12.1% at less than 23 weeks of gestation, 60% at 23 weeks, and 100% at 24 to 26 weeks. Maternal morbidity was minimal with puerperal endomyometritis in 5 (6.8%) cases, one of which became septic; however, there was no long-term sequela. Eight (15.7%) liveborn infants had pulmonary hypoplasia, 5 (62.5%) of which resulted in neonatal death. In 33 (45.2%) patients, amniotic membranes ruptured before 23 weeks of gestation. At previable gestational age, the risk of neonatal pulmonary hypoplasia appears to be primarily dependent on gestational age at the onset of premature rupture of membrane rather than gestational age at delivery. Pregnancy outcomes remain dismal when the fetal membrane ruptures before 23 weeks of gestation. PMID- 15653782 TI - So much to see, so little time: how the superior colliculus (SC) suppresses unwanted saccades. Focus on "Physiological characterization of synaptic inputs to inhibitory burst neurons from the rostral and caudal superior colliculus". PMID- 15653783 TI - Biophysical and morphological properties of parasympathetic neurons controlling the parotid and von Ebner salivary glands in rats. AB - The inferior salivatory nucleus (ISN) contains parasympathetic neurons controlling the parotid and von Ebner salivary glands. To characterize the neurophysiological and morphological properties of these neurons, intracellular recordings were made from anatomically identified ISN neurons in rat brain slices. Neurons were also filled with Lucifer yellow and morphometrically analyzed. Based on responses to membrane hyperpolarization followed by depolarization, three types of repetitive discharge patterns were defined for neurons innervating the parotid gland. The regular, repetitive discharge response to membrane depolarization was changed by hyperpolarization resulting either in a delay in the occurrence of the first spike or to an increase in the length of the first interspike interval in the action potential train. Membrane hyperpolarization had little effect on the discharge pattern of some neurons. Similar response discharge patterns were found for neurons innervating the von Ebner salivary gland, which also included a further group of neurons that responded with a short burst of action potentials. Neurons innervating the parotid salivary glands differed morphologically from the von Ebner salivary glands having significantly larger soma and more and longer dendrites than von Ebner gland neurons. In addition, the mean membrane input resistance, time constant, and spike half-width of parotid gland neurons was significantly lower than in von Ebner gland neurons. These differences in intrinsic membrane properties and morphology may relate to the functions of the von Ebner and parotid glands. von Ebner glands are involved in taste stimulus delivery and removal from posterior tongue papillae while the parotid glands contribute saliva to the entire mouth. PMID- 15653784 TI - Physiological characterization of synaptic inputs to inhibitory burst neurons from the rostral and caudal superior colliculus. AB - The caudal superior colliculus (SC) contains movement neurons that fire during saccades and the rostral SC contains fixation neurons that fire during visual fixation, suggesting potentially different functions for these 2 regions. To study whether these areas might have different projections, we characterized synaptic inputs from the rostral and caudal SC to inhibitory burst neurons (IBNs) in anesthetized cats. We recorded intracellular potentials from neurons in the IBN region and identified them as IBNs based on their antidromic activation from the contralateral abducens nucleus and short-latency excitation from the contralateral caudal SC and/or single-cell morphology. IBNs received disynaptic inhibition from the ipsilateral caudal SC and disynaptic inhibition from the rostral SC on both sides. Stimulation of the contralateral IBN region evoked monosynaptic inhibition in IBNs, which was enhanced by preconditioning stimulation of the ipsilateral caudal SC. A midline section between the IBN regions eliminated inhibition from the ipsilateral caudal SC, but inhibition from the rostral SC remained unaffected, indicating that the latter inhibition was mediated by inhibitory interneurons other than IBNs. A transverse section of the brain stem rostral to the pause neuron (PN) region eliminated inhibition from the rostral SC, suggesting that this inhibition is mediated by PNs. These results indicate that the most rostral SC inhibits bilateral IBNs, most likely via PNs, and the more caudal SC exerts monosynaptic excitation on contralateral IBNs and antagonistic inhibition on ipsilateral IBNs via contralateral IBNs. The most rostral SC may play roles in maintaining fixation by inhibition of burst neurons and facilitating saccadic initiation by releasing their inhibition. PMID- 15653785 TI - Histamine contributes to ischemia-related activation of cardiac spinal afferents: role of H1 receptors and PKC. AB - Myocardial ischemia activates cardiac spinal afferents that transmit the nociceptive information leading to chest pain and elicit excitatory cardiovascular reflexes. Previous studies have shown that histamine is increased in coronary sinus blood during myocardial ischemia and that this autacoid stimulates abdominal visceral afferents. The present investigation evaluated the role of endogenous histamine in stimulation of ischemically sensitive cardiac spinal afferents. Nerve activity of single-unit cardiac afferents was recorded from the left sympathetic chain or rami communicans (T2-T5) in anesthetized cats. Sixty-four cardiac afferents were identified. Injection (5-30 microg/kg) of histamine into the left atrium (LA) stimulated 7 ischemically sensitive cardiac afferents resulting in a significant increase in their activity in a dose dependent manner. Also, LA injection of histamine (10 microg/kg) stimulated 7 of 8 ischemically insensitive cardiac spinal afferents. Administrations of 2-(3 chlorophenyl)histamine (250 microg/kg, LA), a specific H1 receptor agonist and histamine (10 microg/kg, LA), stimulated 9 other ischemically sensitive cardiac afferents (0.48 +/- 0.10 to 1.40 +/- 0.20 imp/s). In contrast, dimaprit (500 microg/kg, LA), an H2 receptor agonist, stimulated only one of the 9 afferents and thus did not alter their overall activity (0.40 +/- 0.09 to 0.54 +/- 0.09 imp/s). (R)alpha-Methyl-histamine (500 microg/kg, LA), an H3 receptor agonist, did not stimulate any of the 9 afferents. Pyrilamine (300 microg/kg, i.v.), a selective H1 receptor antagonist, attenuated the activity of 8 afferents during 5 min of ischemia from 3.32 +/- 0.38 to 1.87 +/- 0.28 imp/s and abolished the response of 9 other cardiac afferents to histamine. Finally, administration of PKC-(19-36) (30 microg/kg, i.v.), a selective inhibitor of protein kinase C, attenuated the response of 8 cardiac afferents to histamine by 32%. These data indicate that endogenous histamine contributes to activation of cardiac sympathetic afferents during myocardial ischemia through H1 receptors and that the action of histamine on these cardiac afferents is partially dependent on the intracellular PKC pathway. PMID- 15653786 TI - Ionic current model of a hypoglossal motoneuron. AB - We have developed a single-compartment, electrophysiological, hypoglossal motoneuron (HM) model based primarily on experimental data from neonatal rat HMs. The model is able to reproduce the fine features of the HM action potential: the fast afterhyperpolarization, the afterdepolarization, and the medium-duration afterhyperpolarization (mAHP). The model also reproduces the repetitive firing properties seen in neonatal HMs and replicates the neuron's response to pharmacological experiments. The model was used to study the role of specific ionic currents in HM firing and how variations in the densities of these currents may account for age-dependent changes in excitability seen in HMs. By varying the density of a fast inactivating calcium current, the model alternates between accelerating and adapting firing patterns. Modeling the age-dependent increase in H current density accounts for the decrease in mAHP duration observed experimentally, but does not fully account for the decrease in input resistance. An increase in the density of the voltage-dependent potassium currents and the H current is required to account for the decrease in input resistance. These changes also account for the age-dependent decrease in action potential duration. PMID- 15653787 TI - Morphological correlates of triadic circuitry in the lateral geniculate nucleus of cats and rats. AB - We used an in vitro slice preparation of the lateral geniculate nucleus in cats and rats to study morphological correlates of triadic circuitry in relay cells. The three triadic elements involve a retinal synapse onto a GABAergic dendritic terminal of an interneuron, a synapse from the same retinal terminal onto a relay cell dendrite, and a synapse from the same interneuron terminal onto the same relay cell dendrite. We made whole cell recordings and labeled cells with biocytin. Previous methods were used to identify triadic circuitry based on evidence that the retinal terminal activates a metabotropic glutamate receptor on the interneuronal terminal. Thus application of (+/-)-1-aminocyclopentane-trans 1,3-dicarboxylic acid (an agonist to that receptor) increases the rate of spontaneous inhibitory postsynaptic currents (sIPSCs) recorded in the relay cell, and if some of this increase remains with further addition of TTX (a TTX insensitive response), a triad is indicated. We quantified the extent of the TTX insensitive response and sought morphological correlates. In both rats and cats, this response correlated (negatively) with the number of primary dendrites and (positively) with polarity of the dendritic arbor. There was no correlation with cell size. Curiously, in cats, this response correlated with the presence of appendages at primary dendritic branches, but there was no such correlation in rats. These observations in cats map onto the X/Y classification, with X cells having triads, but it is not clear from our results if a comparable classification exists for rats. PMID- 15653788 TI - Endogenous PGE2 regulates membrane excitability and synaptic transmission in hippocampal CA1 pyramidal neurons. AB - The significance of cyclooxygenases (COXs), the rate-limiting enzymes that convert arachidonic acid (AA) to prostaglandins (PGs) in the brain, is unclear, although they have been implicated in inflammatory responses and in some neurological disorders such as epilepsy and Alzheimer's disease. Recent evidence that COX-2, which is expressed in postsynaptic dendritic spines, regulates PGE2 signaling in activity-dependent long-term synaptic plasticity at hippocampal perforant path-dentate granule cell synapses, suggests an important role of the COX-2-generated PGE2 in synaptic signaling. However, little is known of how endogenous PGE2 regulates neuronal signaling. Here we showed that endogenous PGE2 selectively regulates fundamental membrane and synaptic properties in the hippocampus. Somatic and dendritic membrane excitability was significantly reduced when endogenous PGE2 was eliminated with a selective COX-2 inhibitor in hippocampal CA1 pyramidal neurons in slices. Exogenous application of PGE2 produced significant increases in frequency of firing, excitatory postsynaptic potentials (EPSP) amplitude, and temporal summation in slices treated with the COX-2 inhibitor. The PGE2-induced increase in membrane excitability seemed to result from its inhibition of the potassium currents, which in turn, boosted dendritic Ca2+ influx during dendritic-depolarizing current injections. In addition, the PGE2-induced enhancement of EPSPs was blocked by eliminating both PKA and PKC activities. These findings indicate that endogenous PGE2 dynamically regulates membrane excitability, synaptic transmission, and plasticity and that the PGE2-induced synaptic modulation is mediated via cAMP-PKA and PKC pathways in rat hippocampal CA1 pyramidal neurons. PMID- 15653790 TI - Nocturnal arterial oxygen saturation and academic performance in a community sample of children. AB - OBJECTIVE: Hypoxemia, often assessed via pulse oximetry, is associated with neurocognitive deficits in children. The best way to qualify hypoxemia, or which level of hypoxemia already affects cognition, is unknown. METHODS: We assessed the association of pulse oximetry-derived variables that qualify hypoxemia with impaired academic performance in mathematics in a population-based cross-section of 995 primary school children who underwent overnight home recordings of motion resistant new-generation pulse oximeter saturation (SpO2). Impaired academic performance in mathematics was based on the last school report and defined as grade 4 to 6 on a 6-point scale (ie, approximately the lowest quintile grades). RESULTS: Of 10 variables under study, only the nadir of the SpO2 values was significantly associated with impaired performance. Categories of this variable representing mild (ie, 91%-93% SpO2; odds ratio: 1.65; 95% confidence interval: 1.06-2.56) and moderate hypoxemia (ie, < or =90% SpO2 ; odds ratio: 2.28; 95% confidence interval: 1.30-4.01) both were associated with impaired performance in mathematics. CONCLUSIONS: We suggest using the nadir of the SpO2 values in an overnight study to qualify hypoxemia in future studies. This variable may predict neurocognitive deficits in school children. Mild hypoxemia, as yet widely considered benign, may already affect cognition in childhood. SpO2 PMID- 15653789 TI - Relationship of lead, mercury, mirex, dichlorodiphenyldichloroethylene, hexachlorobenzene, and polychlorinated biphenyls to timing of menarche among Akwesasne Mohawk girls. AB - BACKGROUND: Children are commonly exposed at background levels to several ubiquitous environmental pollutants, such as lead and persistent organic pollutants, that have been linked to neurologic and endocrine effects. These effects have prompted concern about alterations in human reproductive development. Few studies have examined the effects of these toxicants on human sexual maturation at levels commonly found in the general population, and none has been able to examine multiple toxicant exposures. The aim of the current investigation was to examine the relationship between attainment of menarche and levels of 6 environmental pollutants to which children are commonly exposed at low levels, ie, dichlorodiphenyldichloroethylene (p,p'-DDE), hexachlorobenzene (HCB), polychlorinated biphenyls (PCBs), mirex, lead, and mercury. METHODS: This study was conducted with residents of the Akwesasne Mohawk Nation, a sovereign territory that spans the St Lawrence River and the boundaries of New York State and Ontario and Quebec, Canada. Since the 1950s, the St Lawrence River has been a site of substantial industrial development, and the Nation is currently adjacent to a US National Priority Superfund site. PCB, p,p'-DDE, HCB, and mirex levels exceeding the US Food and Drug Administration recommended tolerance limits for human consumption have been found in local animal species. The present analysis included 138 Akwesasne Mohawk Nation girls 10 to 16.9 years of age. Blood samples and sociodemographic data were collected by Akwesasne community members, without prior knowledge of participants' exposure status. Attainment of menses (menarche) was assessed as present or absent at the time of the interview. Congener-specific PCB analysis was available, and all 16 PCB congeners detected in >50% of the sample were included in analyses (International Union of Pure and Applied Chemistry numbers 52, 70, 74, 84, 87, 95, 99, 101 [+90], 105, 110, 118, 138 [+163 and 164], 149 [+123], 153, 180, and 187). Probit analysis was used to determine the median age at menarche for the sample. Binary logistic regression analysis was used to determine predictors of menarcheal status. Six toxicants (p,p'-DDE, HCB, PCBs, mirex, lead, and mercury) were entered into the logistic regression model. Age, socioeconomic status (SES), and BMI were tested as potential cofounders and were included in the model at P < .05. Interactions among toxicants were also evaluated. RESULTS: Toxicant levels were measured in blood for this sample and were consistent with long-term exposure to a variety of toxicants in multiple media. Mercury levels were at or below background levels, all lead levels were well below the Centers for Disease Control and Prevention action limit of 10 microg/dL, and PCB levels were consistent with a cumulative, continuing exposure pattern. The median age at menarche for the total sample was 12.2 years. The predicted age at menarche for girls with lead levels above the median (1.2 microg/dL) was 10.5 months later than that for girls with lead levels below the median. In the logistic regression analysis, age was the strongest predictor of menarcheal status and SES was also a significant predictor but BMI was not. The logistic regression analysis that corrected for age, SES, and other pollutants (p,p'-DDE, HCB, mirex, and mercury) indicated that, at their respective geometric means, lead (geometric mean: 0.49 microg/dL) was associated with a significantly lower probability of having reached menarche (beta = -1.29) and a group of 4 potentially estrogenic PCB congeners (E-PCB) (geometric mean: 0.12 ppb; International Union of Pure and Applied Chemistry numbers 52, 70, 101 [+90], and 187) was associated with a significantly greater probability of having reached menarche (beta = 2.13). Predicted probabilities at different levels of lead and PCBs were calculated on the basis of the logistic regression model. At the respective means of all toxicants and SES, 69% of 12-year-old girls were predicted to have reached menarche. However, at the 75th percentile of lead levels, only 10% of 12-year-old Mohawk girls were predicted to have reached menarche; at the 75th percentile of E-PCB levels, 86% of 12-year-old Mohawk girls were predicted to have reached menarche. No association was observed between mirex, p,p'-DDE, or HCB and menarcheal status. Although BMI was not a significant predictor, we tested BMI in the logistic regression model; it had little effect on the relationships between menarcheal status and either lead or E-PCB. In models testing toxicant interactions, age, SES, lead levels, and PCB levels continued to be significant predictors of menarcheal status. When each toxicant was tested in a logistic regression model correcting only for age and SES, we observed little change in the effects of lead or E-PCB on menarcheal status. CONCLUSIONS: The analysis of multichemical exposure among Akwesasne Mohawk Nation adolescent girls suggests that the attainment of menarche may be sensitive to relatively low levels of lead and certain PCB congeners. This study is distinguished by the ability to test many toxicants simultaneously and thus to exclude effects from unmeasured but coexisting exposures. By testing several PCB congener groupings, we were able to determine that specifically a group of potentially estrogenic PCB congeners affected the odds of reaching menarche. The lead and PCB findings are consistent with the literature and are biologically plausible. The sample size, cross-sectional study design, and possible occurrence of confounders beyond those tested suggest that results should be interpreted cautiously. Additional investigation to determine whether such low toxicant levels may affect reproduction and disorders of the reproductive system is warranted. PMID- 15653791 TI - Letrozole significantly improves growth potential in a pubertal boy with growth hormone deficiency. AB - Clinical experience with using an aromatase inhibitor to suppress estrogen production during puberty for improvement of growth potential in adolescents with short stature is limited. This report documents treatment of such a patient with a combination of growth hormone and letrozole, a third-generation aromatase inhibitor. Our case demonstrates a favorable outcome on a short-term basis. PMID- 15653792 TI - Cyanobacterial phytochrome-like PixJ1 holoprotein shows novel reversible photoconversion between blue- and green-absorbing forms. AB - The gene, pixJ1 (formerly pisJ1), is predicted to encode a phytochrome-like photoreceptor that is essential for positive phototaxis in the unicellular cyanobacterium Synechocystis sp. PCC 6803 [Yoshihara et al. (2000) Plant Cell Physiol. 41: 1299]. The PixJ1 protein was overexpressed as a fusion with a poly histidine tag (His-PixJ1) and isolated from Synechocystis cells. A zinc fluorescence assay suggested that a linear tetrapyrrole was covalently attached to the His-PixJ1 protein as a chromophore. His-PixJ1 showed novel photoreversible conversion between a blue light-absorbing form (Pb, lambdaAmax=425-435 nm) and a green light-absorbing form (Pg, lambdaAmax=535 nm). Dark incubation led Pg to revert to Pb, indicative of stability of the Pb form in darkness. Red or far-red light irradiation, which is effective for photochemical conversion of the known phytochromes, produced no change in the spectra of Pb and Pg forms. Site-directed mutagenesis revealed that a Cys-His motif in the second GAF domain of PixJ1 is responsible for binding of the chromophore. Possible chromophore species are discussed with regard to the novel photoconversion spectrum. PMID- 15653793 TI - Characterization of full-length enriched expressed sequence tags of stress treated poplar leaves. AB - Poplar, whose genome is the first to be sequenced among woody plants, is a favorable model for plant biologists to enable them to understand molecular processes of growth, development and responses to environmental stimuli in trees. The sequence will allow the development of a strategy for improving environmental stress tolerance in forest trees. In this study, we have generated a full-length enriched cDNA library from leaves of axenically grown poplar (Populus nigra var. italica) subjected to environmental stress treatments by dehydration, high salinity, chilling, heat, abscisic acid (ABA) and H2O2. We sequenced >30,000 expressed sequence tags (ESTs) from the cDNA library and consequently collected approximately 4,500 non-redundant clones. We further analyzed cDNAs encoding an ERF/AP2-domain transcription factor which is specific in plants and plays an important role under stress. Thirteen candidates containing the ERF/AP2 domain were found within our EST resource. Some of them showed stress-responsive gene expression. We report here the first collection of full-length enriched stress related ESTs of poplar and discuss environmental stress responses of forest trees in the light of comparative genomics. PMID- 15653794 TI - Zinc transporter of Arabidopsis thaliana AtMTP1 is localized to vacuolar membranes and implicated in zinc homeostasis. AB - Cation diffusion facilitator (CDF) proteins belong to a family of heavy metal efflux transporters that might play an essential role in homeostasis and tolerance to metal ions. We investigated the subcellular localization of Arabidopsis thaliana AtMTP1, a member of the CDF family, and its physiological role in the tolerance to Zn using MTP1-deficient mutant plants. AtMTP1 was immunochemically detected as a 43 kDa protein in the vacuolar membrane fractioned by sucrose density gradient centrifugation. The expression level of AtMTP1 in suspension-cultured cells was not affected by the Zn concentration in the medium. When AtMTP1 fused with green fluorescent protein was transiently expressed in protoplasts prepared from Arabidopsis suspension-cultured cells, green fluorescence was clearly observed in the vacuolar membrane. A T-DNA insertion mutant line for AtMTP1 displays enhanced sensitivity to high Zn concentrations ranging from 200 to 500 microM, but not to Zn-deficient conditions. Mesophyll cells of the mtp1-1 mutant plants grown in the presence of 500 microM Zn were degraded, suggesting that Zn at high concentrations causes serious damage to leaves and that AtMTP1 plays a crucial role in preventing this damage in plants. Thus we propose that AtMTP1 is localized in the vacuolar membrane and is involved in sequestration of excess Zn in the cytoplasm into vacuoles to maintain Zn homeostasis. PMID- 15653795 TI - Interaction of N-acetylglutamate kinase with a PII-like protein in rice. AB - PII protein in bacteria is a sensor for 2-oxoglutarate and a transmitter for glutamine signaling. We identified an OsGlnB gene that encoded a bacterial PII like protein in rice. Yeast two-hybrid analysis showed that an OsGlnB gene product interacted with N-acetylglutamate kinase 1 (OsNAGK1) and PII-like protein (OsGlnB) itself in rice. In cyanobacteria, NAGK is a key enzyme in arginine biosynthesis. Transient expression of OsGlnB cDNA or OsNAGK1 cDNA fused with sGFP in rice leaf blades strongly suggested that the PII-like protein as well as OsNAGK1 protein is located in chloroplasts. Both OsGlnB and OsNAGK1 genes were expressed in roots, leaf blades, leaf sheaths and spikelets of rice, and these two genes were coordinately expressed in leaf blades during the life span. Thus, PII-like protein in rice plants is potentially able to interact with OsNAGK1 protein in vivo. This finding will provide a clue to the precise physiological function of PII-like protein in rice. PMID- 15653796 TI - Sorting signals in the cytosolic tail of plant p24 proteins involved in the interaction with the COPII coat. AB - The ability of the cytosolic tail of a plant p24 protein to bind COPI and COPII subunits from plant and animal sources in vitro has been examined. We have found that a dihydrophobic motif in the -7,-8 position (relative to the cytosolic carboxy-terminus), which strongly cooperates with a dilysine motif in the -3,-4 position for COPI binding, is required for COPII binding. In addition, we show that COPI and COPII coat proteins from plant cytosol compete for binding to the sorting motifs in these tails. Only in the absence of the dilysine motif in the 3,-4 position or after COPI depletion could we observe COPII binding to the p24 tail. This competition is not observed when using rat liver cytosol. PMID- 15653797 TI - Overexpression of phytochelatin synthase in Arabidopsis leads to enhanced arsenic tolerance and cadmium hypersensitivity. AB - Phytochelatin synthase (PCS) catalyzes the final step in the biosynthesis of phytochelatins, which are a family of cysteine-rich thiol-reactive peptides believed to play important roles in processing many thiol-reactive toxicants. A modified Arabidopsis thaliana PCS sequence (AtPCS1) was active in Escherichia coli. When AtPCS1 was overexpressed in Arabidopsis from a strong constitutive Arabidopsis actin regulatory sequence (A2), the A2::AtPCS1 plants were highly resistant to arsenic, accumulating 20-100 times more biomass on 250 and 300 microM arsenate than wild type (WT); however, they were hypersensitive to Cd(II). After exposure to cadmium and arsenic, the overall accumulation of thiol-peptides increased to 10-fold higher levels in the A2::AtPCS1 plants compared with WT, as determined by fluorescent HPLC. Whereas cadmium induced greater increases in traditional PCs (PC2, PC3, PC4), arsenic exposure resulted in the expression of many unknown thiol products. Unexpectedly, after arsenate or cadmium exposure, levels of the dipeptide substrate for PC synthesis, gamma-glutamyl cysteine (gamma-EC), were also dramatically increased. Despite these high thiol-peptide concentrations, there were no significant increases in concentrations of arsenic and cadmium in above-ground tissues in the AtPCS1 plants relative to WT plants. The potential for AtPCS1 overexpression to be useful in strategies for phytoremediating arsenic and to compound the negative effects of cadmium are discussed. PMID- 15653798 TI - Cryptochromes and phytochromes synergistically regulate Arabidopsis root greening under blue light. AB - To increase their fitness, plants sense ambient light conditions and modulate their developmental processes by utilizing multiple photoreceptors such as phytochrome, cryptochrome and phototropin. Even roots, which are normally not exposed to light, express photoreceptors and can respond to light by developing chloroplasts. In the present study, root greening was observed in Arabidopsis thaliana. Seedlings were grown under monochromatic light and chlorophyll levels in the roots were determined. It was found that blue light was far more effective at inducing chloroplast development in Arabidopsis roots than was red light, and this response was under the control of a strong synergistic interaction between phytochromes and cryptochromes. As expected, the cry1 mutant was deficient in this response. Interestingly, the phyAphyB double mutant failed to respond to blue light under these conditions. This strongly suggests that either phytochrome A or phytochrome B, in addition to cryptochrome, was required for this blue light response. It was further demonstrated that the expression of photosynthetic genes was regulated in the same way. Dichromatic irradiation experiments indicated that this interaction depends on the level of phyB P(FR). Analysis of the cop1, det1 and hy5 mutants indicated that the corresponding factors were involved in the response. PMID- 15653799 TI - PSII-Tc protein plays an important role in dimerization of photosystem II. AB - We cloned and determined the nucleotide sequence of PSII genes, psbB and psbTc, from the thermophilic cyanobacterium, Thermosynechococcus elongatus strain BP-1. PSII-Tc, encoded by psbTc, is a small membrane-spanning subunit of the PSII core complex of cyanobacteria and plants. However, its role has not been fully elucidated. We generated an insertional disruptant of psbTc and studied the role of the PSII-Tc protein in cyanobacterial PSII. The following observations were made: (i) The psbTc disruptant could grow photoautotrophically at a rate similar to that of wild-type T. elongatus under a wide range of light conditions. (ii) Thylakoids and oxygen-evolving PSII complexes were successfully isolated from the psbTc disruptant as well as the wild type. There was no significant difference in the oxygen evolution activities of cells, thylakoids or PSII complexes between the psbTc disruptant and the wild type. This is in contrast to the lower activities in the other PSII mutants of T. elongatus. (iii) Chromatographic separation of monomeric and dimeric PSII revealed that recovery of dimeric PSII was dramatically reduced in the psbTc disruptant. (iv) SDS-urea-PAGE showed a complete loss of the 4.7-kDa band in the mutant PSII. Since this band in wild type PSII consists of PSII-M and PSII-Tc, we assume that PSII-Tc is critical for the binding of PSII-M in the PSII complex and is involved directly and indirectly in the dimerization of PSII. These results appear to be in good agreement with the recent structural model of the dimeric PSII complex. PMID- 15653800 TI - Isolation and identification of glycosylphosphatidylinositol-anchored arabinogalactan proteins and novel beta-glucosyl Yariv-reactive proteins from seeds of rice (Oryza sativa). AB - Arabinogalactan proteins (AGPs) are highly glycosylated extracellular glycoproteins playing important roles in plant growth and development. We have previously reported the possibility that AGPs are involved in the induction of alpha-amylase by gibberellin (GA) in barley aleurone layers by using the beta glucosyl Yariv reagent (beta-GlcY), which has been presumed to specifically bind AGPs. In this present study, we isolated beta-GlcY-reactive proteins from rice bran rich in aleurone cells. The N-terminal sequences of classical AGP and AG peptides were determined from hydrophilic fractions obtained by reversed phase HPLC. Interestingly, a novel non-specific lipid transfer protein-like protein (OsLTPL1) and a novel early nodulin-like protein (OsENODL1) were also identified in the more hydrophobic fractions from HPLC as beta-GlcY-reactive proteins. Expression analysis of the genes coding for these proteins was performed. While classical AGP, AG peptides and OsLTPL1 were expressed in various parts of rice, OsENODL1 showed temporally and spatially specific expression in the aleurone layers. This new beta-GlcY-reactive protein is a promising candidate for the extracellular signaling factors of GA action in cereal seeds. Furthermore, the possibility that proteins with the AG glycomodule might react with beta-GlcY may broaden the definition of AGPs. PMID- 15653801 TI - Asymmetric localization of seed storage protein RNAs to distinct subdomains of the endoplasmic reticulum in developing maize endosperm cells. AB - Plant storage proteins are synthesized and stored in different compartments of the plant endomembrane system. Developing maize seeds synthesize and accumulate prolamin (zein) and 11S globulin (legumin-1) type proteins, which are sequestered in the endoplasmic reticulum (ER) lumen and storage vacuoles, respectively. Immunofluorescence studies showed that the lumenal chaperone BiP was not randomly distributed within the ER in developing maize endosperm but concentrated within the zein-containing protein bodies. Analysis of the spatial distribution of RNAs in maize endosperm sections by in situ RT-PCR showed that, contrary to the conclusions made in an earlier study [Kim et al. (2002) Plant Cell 14: 655-672], the zein and legumin-1 RNAs are not symmetrically distributed on the ER but, instead, targeted to specific ER subdomains. RNAs coding for 22 kDa alpha-zein, 15 kDa beta-zein, 27 kDa gamma-zein and 10 kDa delta-zein were localized to ER bounded zein protein bodies, whereas 51 kDa legumin-1 RNAs were distributed on adjacent cisternal ER proximal to the zein protein bodies. These results indicate that the maize storage protein RNAs are targeted to specific ER subdomains in developing maize endosperm and that RNA localization may be a prevalent mechanism to sort proteins within plant cells. PMID- 15653802 TI - Isolation and expression of two aquaporin-encoding genes from the marine phanerogam Posidonia oceanica. AB - Seagrasses such as Posidonia oceanica (L.) Delile are marine phanerogams, widespread in various seas, where they form large prairies representing dynamic substrates exceeding the area of the sediment surface several times over and allowing settlement of epiphyte organisms. Studying mechanisms involved in water transport in marine plants, we isolated two aquaporin-encoding genes, PoPIP1;1 and PoTIP1;1, showing high similarity to plasma membrane- and tonoplast-intrinsic protein-encoding genes, respectively. PoPIP1;1 is unique in the genome of P. oceanica, while PoTIP1;1 belongs to an aquaporin subfamily of at least four members. PoPIP1;1 and PoTIP1;1 encode functional proteins, as indicated by expression experiments in Xenopus oocytes. Both genes are constitutively expressed in the leaves, with higher levels of transcripts in young than in differentiated leaf tissues. Variations of salt concentration in aquarium determined different PoPIP1;1 and PoTIP1;1 transcript accumulation, indicating the existence of adaptation mechanisms related to gene expression also in marine plants, i.e. adapted to very high salt concentrations. Hyposalinity induced lower levels of PIP1 transcripts, while hypersalinity determined more PIP1 transcripts than normal salinity. TIP1 transcripts increased in response to both hypo- and hypersalinity after 2 days of treatment and went back to control levels after 5 d. PMID- 15653803 TI - Ultraviolet-B sensitivities in Japanese lowland rice cultivars: cyclobutane pyrimidine dimer photolyase activity and gene mutation. AB - There is a cultivar difference in the response to ultraviolet-B (UVB: 280-320 nm) in rice (Oryza sativa L.). Among Japanese lowland rice cultivars, Sasanishiki, a leading Japanese rice cultivar, is resistant to the damaging effects of UVB while Norin 1, a close relative, is less resistant. We found previously that Norin 1 was deficient in cyclobutane pyrimidine dimer (CPD) photorepair ability and suggested that the UVB sensitivity in rice depends largely on CPD photorepair ability. In order to verify that suggestion, we examined the correlation between UVB sensitivity and CPD photolyase activity in 17 rice cultivars of progenitors and relatives in breeding of UV-resistant Sasanishiki and UV-sensitive Norin 1. The amino acid at position 126 of the deduced amino acid sequence of CPD photolyase in cultivars including such as Norin 1 was found to be arginine, the CPD photolyase activities of which were lower. The amino acid at that position in cultivars including such as Sasanishiki was glutamine. Furthermore, cultivars more resistant to UVB were found to exhibit higher photolyase activities than less resistant cultivars. These results emphasize that single amino acid alteration from glutamine to arginine leads to a deficit of CPD photolyase activity and that CPD photolyase activity is one of the main factors determining UVB sensitivity in rice. PMID- 15653804 TI - Effects of chloramphenicol on photosynthesis, protein profiles and transketolase activity under extremely high CO2 concentration in an extremely-high-CO2-tolerant green microalga, Chlorococcum littorale. AB - An extremely-high-CO2-tolerant alga, Chlorococcum littorale, showed high quantum efficiency of PSII (PhiII) in the light at 40% CO2, as well as at 5% CO2. However, PhiII decreased greatly when chloramphenicol (CAP) was added at 40% CO2, while no such decrease was observed at 5% CO2. Cycloheximide showed no effect on PhiII at either 5% or 40% CO2. The amount of a 76 kDa polypeptide (p76) on SDS PAGE decreased markedly in the presence of CAP at 40% CO2 but not at 5% CO2. A partial amino acid sequence of p76 was 71-100% identical (10-14 identical residues out of 14 amino acids determined) to those of transketolases (TKLs) reported in higher plants and a cyanobacterium. In agreement with these observations, the TKL activity in C. littorale was decreased by CAP at 40% CO2, but not at 5% CO2. The transient decrease in TKL activity caused by CAP under 40% CO2 was well correlated with that in PhiII. These results indicate that the addition of CAP directly or indirectly influences the stability of TKL in C. littorale at 40% CO2, but not at 5% CO2, and that photosynthetic activity was reduced by a decrease in TKL activity. PMID- 15653805 TI - Development of Q-chromosome-specific DNA markers in tobacco and their use for identification of a tobacco monosomic line. AB - We developed seven Q-chromosome-specific DNA markers in Nicotiana tabacum by random amplified polymorphic DNA (RAPD) and inter-simple sequence repeat (ISSR) analysis using two hybrid lines, and we were able to identify tobacco monosomic plants among F1 progeny derived from the cross N. tabacum Haplo-QxN. tabacum cv. Samsun NN using Q-chromosome-specific DNA markers. Based on the results, we discuss the roles of the Q chromosome in embryo sac development and embryogenesis. Here, we propose a new method for identifying DNA markers for a particular chromosome in the genus Nicotiana. PMID- 15653806 TI - Photosynthetic characteristics of rice leaves grown under red light with or without supplemental blue light. AB - In rice plants grown under red light supplemented with blue light (red/blue-light PPFD ratio was 4/1), photosynthetic rates per unit leaf area measured under white light at 1,600 and 250 micromol m-2) s-1 were higher than those in the plants grown under red light alone. The higher photosynthetic rates were associated with higher total N content of leaves, which was accompanied by larger amounts of key components of photosynthesis-limiting processes, including Rubisco, Cyt f, Chl and LHCII. These results suggested that the increase in total N content of leaves induced by supplemental blue light enhanced both light-saturated and light limited photosynthesis. PMID- 15653807 TI - A comprehensive expression analysis of the Arabidopsis proline-rich extensin-like receptor kinase gene family using bioinformatic and experimental approaches. AB - The Arabidopsis proline-rich extensin-like receptor kinase (PERK) family consists of 15 predicted receptor kinases. A comprehensive expression analysis was undertaken to identify overlapping and unique expression patterns within this family relative to their phylogeny. Three different approaches were used to study AtPERK gene family expression, and included analyses of the EST, MPSS and NASCArrays databases as well as experimental RNA blot analyses. Some of the AtPERK members were identified as tissue-specific genes while others were more broadly expressed. While in some cases there was a good association between these different expression patterns and the position of the AtPERK members in the kinase phylogeny, in other cases divergence of expression patterns was seen. The PERK expression data identified by the bioinformatics and experimental approaches were found generally to show similar trends and supported the use of data from large-scale expression studies for obtaining preliminary expression data. Thus, the bioinformatics survey for ESTs and microarrays is a powerful comprehensive approach for obtaining a genome-wide view of genes in a multigene family. PMID- 15653808 TI - Identification and subcellular localization of two solanesyl diphosphate synthases from Arabidopsis thaliana. AB - Two solanesyl diphosphate synthases, designated SPS1 and SPS2, which are responsible for the synthesis of the isoprenoid side chain of either plastoquinone or ubiquinone in Arabidopsis thaliana, were identified. Heterologous expression of either SPS1 or SPS2 allowed the generation of UQ-9 in a decaprenyl diphosphate synthase-defective strain of fission yeast and also in wild-type Escherichia coli. SPS1-GFP was found to localize in the ER while SPS2 GFP localized in the plastid of tobacco BY-2 cells. These two different subcellular localizations are thought to be the reflection of their roles in solanesyl diphosphate synthesis in two different parts: presumably SPS1 and SPS2 for the side chains of ubiquinone and plastoquinone, respectively. PMID- 15653809 TI - Peroxisomal localization of sulfite oxidase separates it from chloroplast-based sulfur assimilation. AB - Recently, we isolated the sulfite oxidase (SO) gene from Arabidopsis thaliana and characterized the purified SO protein. The purpose of the present study was to determine the subcellular localization of this novel plant enzyme. Immunogold electron-microscopic analysis showed the gold labels nearly exclusively in the peroxisomes. To verify this finding, green fluorescent protein was fused to full length plant SO including the putative peroxisomal targeting signal 1 (PTS1) 'SNL' and expressed in tobacco leaves. Our results showed a punctate fluorescence pattern resembling that of peroxisomes. Co-labelling with MitoTracker-Red excluded that the observed fluorescence was due to mitochondrial sorting. By investigation of deleted or mutated PTS1, no functional peroxisomal targeting signal 2 (PTS2) could be detected in plant SO. This conclusion is supported by expression studies in Pichia pastoris mutants with defined defects either in PTS1 or PTS2-mediated peroxisomal import. PMID- 15653810 TI - Functional genomics of eukaryotic photosynthesis using insertional mutagenesis of Chlamydomonas reinhardtii. AB - The unicellular green alga Chlamydomonas reinhardtii is a widely used model organism for studies of oxygenic photosynthesis in eukaryotes. Here we describe the development of a resource for functional genomics of photosynthesis using insertional mutagenesis of the Chlamydomonas nuclear genome. Chlamydomonas cells were transformed with either of two plasmids conferring zeocin resistance, and insertional mutants were selected in the dark on acetate-containing medium to recover light-sensitive and nonphotosynthetic mutants. The population of insertional mutants was subjected to a battery of primary and secondary phenotypic screens to identify photosynthesis-related mutants that were pigment deficient, light sensitive, nonphotosynthetic, or hypersensitive to reactive oxygen species. Approximately 9% of the insertional mutants exhibited 1 or more of these phenotypes. Molecular analysis showed that each mutant line contains an average of 1.4 insertions, and genetic analysis indicated that approximately 50% of the mutations are tagged by the transforming DNA. Flanking DNA was isolated from the mutants, and sequence data for the insertion sites in 50 mutants are presented and discussed. PMID- 15653812 TI - Extending the role of associative learning processes in nicotine addiction. AB - Compulsive smoking is a worldwide public health problem. Although research has confirmed the importance of associative learning processes in nicotine addiction, therapies targeting nicotine-associated cues still have a high relapse rate. Most theories conceptualize nicotine as an 'outcome' that reinforces behaviors and/or changes the affective value of stimuli. Albeit important, this view does not capture the complexity of associative processes involved in nicotine addiction. For example, nicotine serves as a conditional stimulus acquiring new appetitive/affective properties when paired with a non-drug reward. Also, nicotine functions as an occasion setter that participates in higher-order associative processes that likely permit a more pervasive influence of conditioned cues that are resistant to typically cue-exposure therapy techniques. Finally, nicotine appears to amplify the salience of other stimuli that have some incentive value resulting in enhanced nicotine self-administration and conditioned reinforcement processes. Future smoking intervention strategies should take into consideration these additional associative learning processes. PMID- 15653813 TI - The cognitive neuroscience of human decision making: a review and conceptual framework. AB - Decision making, the process of choosing between options, is a fundamental human behavior that has been studied intensively by disciplines ranging from cognitive psychology to economics. Despite the importance of this behavior, the neural substrates of decision making are only beginning to be understood. Impaired decision making is recognized in neuropsychiatric conditions such as dementia and drug addiction, and the inconsistencies and biases of healthy decision makers have been intensively studied. However, the tools of cognitive neuroscience have only recently been applied to understanding the brain basis of this complex behavior. This article reviews the literature on the cognitive neuroscience of human decision making, focusing on the roles of the frontal lobes, and provides a conceptual framework for organizing this disparate body of work. PMID- 15653811 TI - Evolution of the isoprene biosynthetic pathway in kudzu. AB - Isoprene synthase converts dimethylallyl diphosphate, derived from the methylerythritol 4-phosphate (MEP) pathway, to isoprene. Isoprene is made by some plants in substantial amounts, which affects atmospheric chemistry, while other plants make no isoprene. As part of our long-term study of isoprene synthesis, the genetics of the isoprene biosynthetic pathway of the isoprene emitter, kudzu (Pueraria montana), was compared with similar genes in Arabidopsis (Arabidopsis thaliana), which does not make isoprene. The MEP pathway genes in kudzu were similar to the corresponding Arabidopsis genes. Isoprene synthase genes of kudzu and aspen (Populus tremuloides) were cloned to compare their divergence with the divergence seen in MEP pathway genes. Phylogenetic analysis of the terpene synthase gene family indicated that isoprene synthases are either within the monoterpene synthase clade or sister to it. In Arabidopsis, the gene most similar to isoprene synthase is a myrcene/ocimene (acyclic monoterpenes) synthase. Two phenylalanine residues found exclusively in isoprene synthases make the active site smaller than other terpene synthase enzymes, possibly conferring specificity for the five-carbon substrate rather than precursors of the larger isoprenoids. Expression of the kudzu isoprene synthase gene in Arabidopsis caused Arabidopsis to emit isoprene, indicating that whether or not a plant emits isoprene depends on whether or not it has a terpene synthase capable of using dimethylallyl diphosphate. PMID- 15653814 TI - Theta rhythm of the hippocampus: subcortical control and functional significance. AB - The theta rhythm is the largest extracellular synchronous signal that can be recorded from the mammalian brain and has been strongly implicated in mnemonic processes of the hippocampus. We describe (a) ascending brain stem-forebrain systems involved in controlling theta and nontheta (desynchronization) states of the hippocampal electroencephalogram; (b) theta rhythmically discharging cells in several structures of Papez's circuit and their possible functional significance, specifically with respect to head direction cells in this same circuit; and (c) the role of nucleus reuniens of the thalamus as a major interface between the medial prefrontal cortex and hippocampus and as a prominent source of afferent limbic information to the hippocampus. We suggest that the hippocampus receives two main types of input: theta rhythm from ascending brain stem- diencephaloseptal systems and information bearing mainly from thalamocortical/cortical systems. The temporal convergence of activity of these two systems results in the encoding of information in the hippocampus, primarily reaching it from the entorhinal cortex and nucleus reuniens. PMID- 15653816 TI - Safety of the blood supply in Latin America. AB - Appropriate selection of donors, use of sensitive screening tests, and the application of a mandatory quality assurance system are essential to maintain the safety of the blood supply. Laws, decrees, norms, and/or regulations covering most of these aspects of blood transfusion exist in 16 of the 17 countries in Latin America that are the subject of this review. In 17 countries, there is an information system that, although still incomplete (there are no official reports on adverse events and incidents), allows us to establish progress made on the status of the blood supply since 1993. Most advances originated in increased screening coverage for infectious diseases and better quality assurance. However, in 2001 to 2002, tainted blood may have caused infections in 12 of the 17 countries; no country reached the number of donors considered adequate, i.e., 5% of the population, to avoid blood shortages, or decreased significantly the number of blood banks, although larger blood banks are more efficient and take advantage of economies of scale. In those years, paid donors still existed in four countries and replacement donors made up >75% of the blood donors in another eight countries. In addition, countries did not report the number of voluntary donors who were repeat donors, i.e., the healthiest category. In spite of progress made, more improvements are needed. PMID- 15653817 TI - Allergy and dermatophytes. AB - Tinea pedis (athlete's foot) and onychomycosis (infection of the toenails) caused by the dermatophyte fungus Trichophyton are highly prevalent in adults. Several Trichophyton allergens have been identified based on elicitation of immunoglobulin E antibody-mediated immediate-hypersensitivity (IH) responses. Evidence of an etiologic role for Trichophyton in asthma in some subjects with IH and chronic dermatophytosis is provided by bronchial reactivity to Trichophyton. Improvement of asthma after systemic antifungal treatment corroborates this link. A unique feature of Trichophyton allergens is the ability of the same antigen to elicit delayed-type hypersensitivity (DTH) in individuals who lack IH reactivity. Delayed responses appear to confer protection, while IH responses do not, based on the association with acute versus chronic skin infection. The amino acid sequence identity of Trichophyton allergens with diverse enzyme families supports a dual role for these proteins in fungal pathogenesis and allergic disease. Characterizing the immunologic properties of Trichophyton allergens and defining immune mechanisms which drive dichotomous responses are pivotal to understanding the dermatophyte-allergy relationship. Recent studies have identified DTH associated major T-cell epitopes which could facilitate the development of peptide vaccines. Characterization of additional molecular targets by using new techniques may aid not only in the eradication of infection but also in the resolution of allergic symptoms. PMID- 15653819 TI - Preventing varicella-zoster disease. AB - Varicella-zoster virus (VZV), the cause of chickenpox and shingles, is a pathogen in retreat following the introduction of mass vaccination in the United States in 1995. The live attenuated Oka vaccine, which is safe and immunogenic, gives good protection against both varicella and zoster in the short to medium term. It has undoubtedly been highly effective to date in reducing all forms of varicella, especially severe disease. However, the huge pool of latent wild-type virus in the population represents a continuing threat. Both the biology and the epidemiology of VZV disease suggest that new vaccination strategies will be required over time. PMID- 15653820 TI - Pathway to synthesis and processing of mycolic acids in Mycobacterium tuberculosis. AB - Mycobacterium tuberculosis is known to synthesize alpha-, methoxy-, and keto mycolic acids. We propose a detailed pathway to the biosynthesis of all mycolic acids in M. tuberculosis. Fatty acid synthetase I provides C(20)-S-coenzyme A to the fatty acid synthetase II system (FAS-IIA). Modules of FAS-IIA and FAS-IIB introduce cis unsaturation at two locations on a growing meroacid chain to yield three different forms of cis,cis-diunsaturated fatty acids (intermediates to alpha-, methoxy-, and keto-meroacids). These are methylated, and the mature meroacids and carboxylated C(26)-S-acyl carrier protein enter into the final Claisen-type condensation with polyketide synthase-13 (Pks13) to yield mycolyl-S Pks13. We list candidate genes in the genome encoding the proposed dehydrase and isomerase in the FAS-IIA and FAS-IIB modules. We propose that the processing of mycolic acids begins by transfer of mycolic acids from mycolyl-S-Pks13 to d mannopyranosyl-1-phosphoheptaprenol to yield 6-O-mycolyl-beta-d-mannopyranosyl-1 phosphoheptaprenol and then to trehalose 6-phosphate to yield phosphorylated trehalose monomycolate (TMM-P). Phosphatase releases the phosphate group to yield TMM, which is immediately transported outside the cell by the ABC transporter. Antigen 85 then catalyzes the transfer of a mycolyl group from TMM to the cell wall arabinogalactan and to other TMMs to produce arabinogalactan-mycolate and trehalose dimycolate, respectively. We list candidate genes in the genome that encode the proposed mycolyltransferases I and II, phosphatase, and ABC transporter. The enzymes within this total pathway are targets for new drug discovery. PMID- 15653822 TI - Mechanical transmission of human protozoan parasites by insects. AB - The filthy breeding habits, feeding mechanisms, and indiscriminate travel between filth and food make some groups of synanthropic insects such as nonbiting flies and cockroaches efficient vectors of human enteric protozoan parasites. Twenty one species of filth flies have been listed by regulatory agencies concerned with sanitation and public health as causative agents of gastrointestinal diseases based on synanthropy, endophily, communicative behavior, and strong attraction to filth and human food. Outbreaks and cases of food-borne diarrheal diseases in urban and rural areas are closely related to the seasonal increase in abundance of filth flies, and enforced fly control is closely related to reductions in the occurrence of such diseases. Mechanical transmission of human parasites by nonbiting flies and epidemiological involvement of other synanthropic insects in human food-borne diseases have not received adequate scientific attention. PMID- 15653818 TI - Aspergillus infections in transplant recipients. AB - Aspergillus infections are occurring with an increasing frequency in transplant recipients. Notable changes in the epidemiologic characteristics of this infection have occurred; these include a change in risk factors and later onset of infection. Management of invasive aspergillosis continues to be challenging, and the mortality rate, despite the use of newer antifungal agents, remains unacceptably high. Performing molecular studies to discern new targets for antifungal activity, identifying signaling pathways that may be amenable to immunologic interventions, assessing combination regimens of antifungal agents or combining antifungal agents with modulation of the host defense mechanisms, and devising diagnostic assays that can rapidly and reliably diagnose infections represent areas for future investigations that may lead to further improvement in outcomes. PMID- 15653823 TI - Options for field diagnosis of human african trypanosomiasis. AB - Human African trypanosomiasis (HAT) due to Trypanosoma brucei gambiense or T. b. rhodesiense remains highly prevalent in several rural areas of sub-Saharan Africa and is lethal if left untreated. Therefore, accurate tools are absolutely required for field diagnosis. For T. b. gambiense HAT, highly sensitive tests are available for serological screening but the sensitivity of parasitological confirmatory tests remains insufficient and needs to be improved. Screening for T. b. rhodesiense infection still relies on clinical features in the absence of serological tests available for field use. Ongoing research is opening perspectives for a new generation of field diagnostics. Also essential for both forms of HAT is accurate determination of the disease stage because of the high toxicity of melarsoprol, the drug most widely used during the neurological stage of the illness. Recent studies have confirmed the high accuracy of raised immunoglobulin M levels in the cerebrospinal fluid for the staging of T. b. gambiense HAT, and a promising simple assay (LATEX/IgM) is being tested in the field. Apart from the urgent need for better tools for the field diagnosis of this neglected disease, improved access to diagnosis and treatment for the population at risk remains the greatest challenge for the coming years. PMID- 15653824 TI - Manual and automated instrumentation for identification of Enterobacteriaceae and other aerobic gram-negative bacilli. AB - Identification of gram-negative bacilli, both enteric and nonenteric, by conventional methods is not realistic for clinical microbiology laboratories performing routine cultures in today's world. The use of commercial kits, either manual or automated, to identify these organisms is a common practice. The advent of rapid or "spot" testing has eliminated the need for some commonly isolated organisms to be identified with the systems approach. Commercially available systems provide more in-depth identification to the species level as well as detect new and unusual strains. The answers obtained from these systems may not always be correct and must be interpreted with caution. The patient demographics, laboratory workload and work flow, and technologist's skill levels should dictate the system of choice. Cost considerations introduce another variable into the equation affecting choice. Each system has its own strengths and weaknesses, and each laboratory must decide on the level of sophistication that fulfills its particular needs. PMID- 15653821 TI - Surface proteins of Streptococcus agalactiae and related proteins in other bacterial pathogens. AB - Streptococcus agalactiae (group B Streptococcus) is the major cause of invasive bacterial disease, including meningitis, in the neonatal period. Although prophylactic measures have contributed to a substantial reduction in the number of infections, development of a vaccine remains an important goal. While much work in this field has focused on the S. agalactiae polysaccharide capsule, which is an important virulence factor that elicits protective immunity, surface proteins have received increasing attention as potential virulence factors and vaccine components. Here, we summarize current knowledge about S. agalactiae surface proteins, with emphasis on proteins that have been characterized immunochemically and/or elicit protective immunity in animal models. These surface proteins have been implicated in interactions with human epithelial cells, binding to extracellular matrix components, and/or evasion of host immunity. Of note, several S. agalactiae surface proteins are related to surface proteins identified in other bacterial pathogens, emphasizing the general interest of the S. agalactiae proteins. Because some S. agalactiae surface proteins elicit protective immunity, they hold promise as components in a vaccine based only on proteins or as carriers in polysaccharide conjugate vaccines. PMID- 15653826 TI - Bacterial contamination of blood components. AB - Blood for transfusion is a potential source of infection by a variety of known and unknown transmissible agents. Over the last 20 years, astounding reductions in the risk of viral infection via allogeneic blood have been achieved. As a result of this success, bacterial contamination of blood products has emerged as the greatest residual source of transfusion-transmitted disease. This paper summarizes the current status of detection, prevention, and elimination of bacteria in blood products for transfusion. PMID- 15653827 TI - Secondary syphilitic lesions. AB - An important theme that emerges from all early historical accounts is that in addition to the decreased virulence of Treponema pallidum, the incidence of secondary syphilis has decreased drastically over the past three centuries. Even in the early 20th century, most syphilologists were of the opinion that the disease had undergone changes in its manifestations and that they were dealing with an attenuated form of the spirochete. Such opinions were based primarily on the observations that violent cutaneous reactions and fatalities associated with the secondary stage had become extremely rare. The rate of primary and secondary syphilis in the United States increased in 2002 for the second consecutive year. After a decade-long decline that led to an all-time low in 2000, the recent trend is attributable, to a large extent, by a increase in reported syphilis cases among men, particularly homosexual and bisexual men having sex with men. The present review addresses the clinical and diagnostic criteria for the recognition of secondary syphilis, the clinical course and manifestations of the disease if allowed to proceed past the primary stage of disease in untreated individuals, and the treatment for this stage of the disease. PMID- 15653825 TI - Combination treatment of invasive fungal infections. AB - The persistence of high morbidity and mortality from systemic fungal infections despite the availability of novel antifungals points to the need for effective treatment strategies. Treatment of invasive fungal infections is often hampered by drug toxicity, tolerability, and specificity issues, and added complications often arise due to the lack of diagnostic tests and to treatment complexities. Combination therapy has been suggested as a possible approach to improve treatment outcome. In this article, we undertake a historical review of studies of combination therapy and also focus on recent studies involving newly approved antifungal agents. The limitations surrounding antifungal combinations include nonuniform interpretation criteria, inability to predict the likelihood of clinical success, strain variability, and variations in pharmacodynamic/pharmacokinetic properties of antifungals used in combination. The issue of antagonism between polyenes and azoles is beginning to be addressed, but data regarding other drug combinations are not adequate for us to draw definite conclusions. However, recent data have identified potentially useful combinations. Standardization of assay methods and adoption of common interpretive criteria are essential to avoid discrepancies between different in vitro studies. Larger clinical trials are needed to assess whether combination therapy improves survival and treatment outcome in the most seriously debilitated patients afflicted with life-threatening fungal infections. PMID- 15653829 TI - Making connections between novel transcription factors and their DNA motifs. AB - The key components of a transcriptional regulatory network are the connections between trans-acting transcription factors and cis-acting DNA-binding sites. In spite of several decades of intense research, only a fraction of the estimated approximately 300 transcription factors in Escherichia coli have been linked to some of their binding sites in the genome. In this paper, we present a computational method to connect novel transcription factors and DNA motifs in E. coli. Our method uses three types of mutually independent information, two of which are gleaned by comparative analysis of multiple genomes and the third one derived from similarities of transcription-factor-DNA-binding-site interactions. The different types of information are combined to calculate the probability of a given transcription-factor-DNA-motif pair being a true pair. Tested on a study set of transcription factors and their DNA motifs, our method has a prediction accuracy of 59% for the top predictions and 85% for the top three predictions. When applied to 99 novel transcription factors and 70 novel DNA motifs, our method predicted 64 transcription-factor-DNA-motif pairs. Supporting evidence for some of the predicted pairs is presented. Functional annotations are made for 23 novel transcription factors based on the predicted transcription-factor-DNA-motif connections. PMID- 15653830 TI - Genome-wide regulatory complexity in yeast promoters: separation of functionally conserved and neutral sequence. AB - To gauge the complexity of gene regulation in yeast, it is essential to know how much promoter sequence is functional. Conservation across species can be a sensitive means of detecting functional sequences, provided that the significance of conservation can be accurately calibrated with the local neutral mutation rate. By analyzing yeast coding and promoter sequences, we find that neutral mutation rates in yeast are uniform genome-wide, in contrast to mammals, where neutral mutation rates vary along chromosomes. We develop an approach that uses this uniform rate to estimate the amount of promoter sequence under purifying selection. This amount is approximately 30%, corresponding to roughly 90 bp for a typical promoter. Furthermore, using a hidden Markov model, we are able to separate each promoter into distinct high and low conservation regions. Known regulatory motifs are strongly biased toward high conservation regions, while low conservation regions have mutation rates similar to that of the neutral background. Certain Gene Ontology groupings of genes (e.g., Carbohydrate Metabolism) have large amounts of high conservation sequence, suggesting complexity in their transcriptional regulation. Others (e.g., RNA Processing) have little high conservation sequence and are likely to be simply regulated. The separation of functionally conserved sequence from the neutral background allows us to estimate the complexity of cis-regulation on a genomic scale. PMID- 15653831 TI - Interchromosomal segmental duplications of the pericentromeric region on the human Y chromosome. AB - Basic medical research critically depends on the finished human genome sequence. Two types of gaps are known to exist in the human genome: those associated with heterochromatic sequences and those embedded within euchromatin. We identified and analyzed a euchromatic island within the pericentromeric repeats of the human Y chromosome. This 450-kb island, although not recalcitrant to subcloning and present in 100 tested males from different ethnic origins, was not detected and is not contained within the published Y chromosomal sequence. The entire 450-kb interval is almost completely duplicated and consists predominantly of interchromosomal rather than intrachromosomal duplication events that are usually prevalent on the Y chromosome. We defined the modular structure of this interval and detected a total of 128 underlying pairwise alignments (>/=90% and >/=1 kb in length) to various autosomal pericentromeric and ancestral pericentromeric regions. We also analyzed the putative gene content of this region by a combination of in silico gene prediction and paralogy analysis. We can show that even in this exceptionally duplicated region of the Y chromosome, eight putative genes with open reading frames reside, including fusion transcripts formed by the splicing of exons from two different duplication modules as well as members of the homeobox gene family DUX. PMID- 15653834 TI - Single-molecule fluorescence spectroscopy: new probes of protein function and dynamics. AB - Single-molecule fluorescence methods provide new tools for the study of biological systems. Single-pair fluorescence resonance energy transfer has provided detailed information about dynamics and structure of the Ca2+-signaling protein calmodulin. Single-molecule polarization modulation spectroscopy has probed the mechanism by which calmodulin activates the plasma membrane Ca2+ pump. PMID- 15653832 TI - Divergent V1R repertoires in five species: Amplification in rodents, decimation in primates, and a surprisingly small repertoire in dogs. AB - The V1R gene family comprises one of two types of putative pheromone receptors expressed in the mammalian vomeronasal organ (VNO). We searched the most recent mouse, rat, dog, chimpanzee, and human genome sequence assemblies to compile a near-complete repertoire of V1R genes for each species. Dog, human, and chimpanzee have very few intact V1Rs (8, 2, and 0, respectively) compared to more than a hundred intact V1Rs in each of the rat (106) and mouse (165) genomes. We also provide the first description of the diversity of V1R pseudogenes in these species. We identify at least 165 pseudogenes in mouse, 110 in rat, 102 in chimpanzee, 115 in human, and 54 in dog. Primate and dog pseudogenes are distributed among almost all V1R subfamilies seen in rodents, indicating that the common ancestor of these species had a diverse V1R repertoire. We find that V1R genes were subject to strikingly different fates in different species and in different subfamilies. In rodents, some subfamilies remained relatively stable or underwent roughly equivalent expansion in mouse and rat; other subfamilies expanded in one species but not the other. The small number of intact V1Rs in the dog genome is unexpected given the presumption that dogs, like rodents, have a functional VNO, and a complex system of pheromone-based behaviors. We identify an intact transient receptor potential channel 2beta in the dog genome, consistent with a functional VNO in dogs. The diminished V1R repertoire in dogs raises questions about the relative contributions of V1Rs versus other candidate pheromone receptor genes in the establishment of complex pheromone systems in mammals. PMID- 15653835 TI - Dynamic multiphoton imaging: a live view from cells to systems. AB - Leaps in scientific technology often occur at the interface of seemingly disparate disciplines. This holds true with the recent application of multiphoton microscopy to the biological sciences, leading to a new generation of imaging based studies extending from the tracking of individual molecules within living cells to the observation of whole organisms. PMID- 15653836 TI - Membrane microparticles: two sides of the coin. AB - Microparticles are plasma membrane-derived vesicles shed from stimulated cells, in the broad sense of the term. Their presence is interpreted by proximal or remote cells in fundamental physiological processes including intercellular communication, hemostasis, and immunity. On the other hand, variations of their number or characteristics are frequently observed in pathophysiological situations. PMID- 15653837 TI - Airway remodeling in asthma: therapeutic implications of mechanisms. AB - Asthma is currently recognized as a chronic inflammatory disorder of the airways that leads to tissue injury and subsequent structural changes collectively called airway remodeling. Transgenic modeling of inflammatory mediators allows for the discovery of unexpected effects, dissection of downstream signaling events, and clues to future therapies. PMID- 15653838 TI - Endothelial progenitor cells for vasculogenesis. AB - Postnatal vasculogenesis is considered to be involved in neovascularization of adult tissues, because bone marrow-derived endothelial progenitor cells (EPCs) were isolated from circulating mononuclear cells in peripheral blood and were shown to incorporate into sites of physiological and pathological neovascularization and to differentiate into mature endothelial cells. EPCs might have an attractive potential therapeutic application for cardiovascular ischemic diseases as a novel cell-based strategy mainly via a vasculogenesis mechanism. PMID- 15653839 TI - Molecular control of cytoskeletal mechanics by hemodynamic forces. AB - The endothelium at the interface between blood and tissue acts as a primary transducer of local hemodynamic forces into signals that maintain physiological function or initiate pathological processes in vessel walls. Rapid intracellular spatial gradients of structural dynamics and signaling molecule activity suggest that mechanical cues at the molecular level guide cellular mechanotransduction and adaptation to shear stress profiles. PMID- 15653840 TI - A role for the Wilms' tumor protein WT1 in organ development. AB - Wilms' tumor (nephroblastoma) represents a unique example of an aberrant kidney formation that can result from mutations in a tumor suppressor gene, Wilms' tumor 1 (WT1). Targeted gene inactivation in mice testifies that WT1 is a master switch for the development of the genitourinary system and other organs. PMID- 15653828 TI - Update on human herpesvirus 6 biology, clinical features, and therapy. AB - Human herpesvirus 6 (HHV-6) is a betaherpesvirus that is closely related to human cytomegalovirus. It was discovered in 1986, and HHV-6 literature has expanded considerably in the past 10 years. We here present an up-to-date and complete overview of the recent developments concerning HHV-6 biological features, clinical associations, and therapeutic approaches. HHV-6 gene expression regulation and gene products have been systematically characterized, and the multiple interactions between HHV-6 and the host immune system have been explored. Moreover, the discovery of the cellular receptor for HHV-6, CD46, has shed a new light on HHV-6 cell tropism. Furthermore, the in vitro interactions between HHV-6 and other viruses, particularly human immunodeficiency virus, and their relevance for the in vivo situation are discussed, as well as the transactivating capacities of several HHV-6 proteins. The insight into the clinical spectrum of HHV-6 is still evolving and, apart from being recognized as a major pathogen in transplant recipients (as exemplified by the rising number of prospective clinical studies), its role in central nervous system disease has become increasingly apparent. Finally, we present an overview of therapeutic options for HHV-6 therapy (including modes of action and resistance mechanisms). PMID- 15653841 TI - Broca's region: from action to language. AB - Broca's region, classically considered a motor speech-production area, is involved in action understanding and imitation. It also seems to help in sequencing of actions. Broca's region might have evolved for interindividual communication, both by gestures and speech. PMID- 15653842 TI - Neurotrophin-mediated rapid signaling in the central nervous system: mechanisms and functions. AB - Neurotrophins regulate growth, survival, and differentiation of central neurons. In addition to the "classical" effects that are relatively slow neurotrophins also elicit rapid signaling that modulates a variety of cellular functions such as membrane excitability, synaptic transmission, and activity-dependent synaptic plasticity. These rapid actions are mediated mainly through the interaction of Trk receptors with ion channels and ionotropic receptors in the plasma membrane. PMID- 15653843 TI - Validation of publication of new names and new combinations previously effectively published outside the IJSEM. AB - The purpose of this announcement is to effect the valid publication of the following new names and new combinations under the procedure described in the Bacteriological Code (1990 Revision). Authors and other individuals wishing to have new names and/or combinations included in future lists should send three copies of the pertinent reprint or photocopies thereof to the IJSEM Editorial Office for confirmation that all of the other requirements for valid publication have been met. It is also a requirement of IJSEM and the ICSP that authors of new species, new subspecies and new combinations provide evidence that types are deposited in two recognized culture collections in two different countries (i.e. documents certifying deposition and availability of type strains). It should be noted that the date of valid publication of these new names and combinations is the date of publication of this list, not the date of the original publication of the names and combinations. The authors of the new names and combinations are as given below, and these authors' names will be included in the author index of the present issue and in the volume author index. Inclusion of a name on these lists validates the publication of the name and thereby makes it available in bacteriological nomenclature. The inclusion of a name on this list is not to be construed as taxonomic acceptance of the taxon to which the name is applied. Indeed, some of these names may, in time, be shown to be synonyms, or the organisms may be transferred to another genus, thus necessitating the creation of a new combination. PMID- 15653844 TI - Notification that new names and new combinations have appeared in volume 54, part 5, of the IJSEM. AB - This listing of names published in a previous issue of the IJSEM is provided as a service to bacteriology to assist in the recognition of new names and new combinations. This procedure was proposed by the Judicial Commission [Minute 11(ii), Int J Syst Bacteriol 41 (1991), p. 185]. The names given herein are listed according to the Rules of priority (i.e. page number and order of valid publication of names in the original articles). Taxonomic opinions included in this List (i.e. the creation of synonyms or the emendation of circumscriptions) cannot be considered as validly published nor, in any other way, approved by the International Committee on Systematics of Prokaryotes and its Judicial Commission. PMID- 15653845 TI - Notification of changes in taxonomic opinion previously published outside the IJSEM. AB - The Bacteriological Code deals with the nomenclature of prokaryotes. This may include existing names (the Approved Lists of Bacterial Names) as well as new names and new combinations. In this sense the Code is also dealing indirectly with taxonomic opinions. However, as with most codes of nomenclature there are no mechanisms for formally recording taxonomic opinions that do not involve the creation of new names or new combinations. In particular, it would be desirable for taxonomic opinions resulting from the creation of synonyms or emended descriptions to be made widely available to the public. In 2004, the Editorial Board of the International Journal of Systematic and Evolutionary Microbiology (IJSEM) agreed unanimously that it was desirable to cover such changes in taxonomic opinions (i.e. the creation of synonyms or the emendation of circumscriptions) previously published outside the IJSEM, and to introduce a List of Changes in Taxonomic Opinion [Notification of changes in taxonomic opinion previously published outside the IJSEM; Euzeby et al. (2004). Int J Syst Evol Microbiol 54, 1429-1430]. Scientists wishing to have changes in taxonomic opinion included in future lists should send one copy of the pertinent reprint or a photocopy or a PDF file thereof to the IJSEM Editorial Office or to the Lists Editor. It must be stressed that the date of proposed taxonomic changes is the date of the original publication not the date of publication of the list. Taxonomic opinions included in the List of Changes in Taxonomic Opinion cannot be considered as validly published nor, in any other way, approved by the International Committee on Systematics of Prokaryotes and its Judicial Commission. The names that are to be used are those that are the 'correct names' (in the sense of Principle 6) in the opinion of the bacteriologist, with a given circumscription, position and rank. A particular name, circumscription, position and rank does not have to be adopted in all circumstances. Consequently, the List of Changes in Taxonomic Opinion must be considered as a service to bacteriology and it has no 'official character', other than providing a centralized point for registering/indexing such changes in a way that makes them easily accessible to the scientific community. PMID- 15653848 TI - 'Candidatus Mycoplasma haematoparvum', a novel small haemotropic mycoplasma from a dog. AB - A novel small haemoplasma was detected following cytological examination of blood smears from a splenectomized dog with haemic neoplasia. The 16S rRNA and rnpB genes of the organism were partially sequenced and a phylogenetic tree constructed. The organism was most closely related to the small feline haemoplasma, 'Candidatus Mycoplasma haemominutum' (94 % 16S rRNA gene nucleotide sequence identity; 75 % rnpB) and was only distantly related to Mycoplasma haemocanis (78 % 16S rRNA gene nucleotide sequence identity; 65 % rnpB). As this organism has not been cultured in vitro, the candidate species name 'Candidatus Mycoplasma haematoparvum' is proposed. PMID- 15653849 TI - Lactobacillus suntoryeus sp. nov., isolated from malt whisky distilleries. AB - Eight strains of Lactobacillus with identical partial 16S rRNA gene sequences and similar randomly amplified polymorphic DNA patterns were isolated from fermentation samples from Japanese and Scottish malt whisky distilleries. Phylogenetic analysis of almost complete 16S rRNA gene sequences from three representative strains (two from Japan, one from Scotland) placed them in the genus Lactobacillus as members of the Lactobacillus acidophilus group. Lactobacillus helveticus and Lactobacillus gallinarum were the most closely related species, with 16S rRNA gene similarities of 99.3 and 98.1 %, respectively. A similar phylogeny was derived from partial sequences of elongation factor Tu (tuf) genes in which the alleles from the three distillery isolates were identical and shared 99.0 % similarity with L. helveticus and L. gallinarum tuf genes. S-layer (slp) gene sequences suggested different relationships among the strains and the distillery isolates no longer formed a monophyletic group. The alleles from the Japanese and Scottish strains shared only 54 % similarity. Chromosomal DNA from the distillery strains gave DNA-DNA hybridization values between 79 and 100 % but showed less than 43 and 22 % reassociation with L. helveticus and L. gallinarum DNA, respectively. The name Lactobacillus suntoryeus sp. nov. is proposed for this novel taxon; the type strain is strain SAT (=LMG 22464T=NCIMB 14005T). PMID- 15653847 TI - Phylogenetic and morphological evaluation of the genera Anabaena, Aphanizomenon, Trichormus and Nostoc (Nostocales, Cyanobacteria). AB - The heterocytous cyanobacteria form a monophyletic group according to 16S rRNA gene sequence data. Within this group, phylogenetic and morphological studies have shown that genera such as Anabaena and Aphanizomenon are intermixed. Moreover, the phylogeny of the genus Trichormus, which was recently separated from Anabaena, has not been investigated. The aim was to study the taxonomy of the genera Anabaena, Aphanizomenon, Nostoc and Trichormus belonging to the family Nostocaceae (subsection IV.I) by morphological and phylogenetic analyses of 16S rRNA gene, rpoB and rbcLX sequences. New strains were isolated to avoid identification problems caused by morphological changes of strains during cultivation. Morphological and phylogenetic data showed that benthic and planktic Anabaena strains were intermixed. In addition, the present study confirmed that Anabaena and Aphanizomenon strains were not monophyletic, as previously demonstrated. The evolutionary distances between the strains indicated that the planktic Anabaena and Aphanizomenon strains as well as five benthic Anabaena strains in cluster 1 could be assigned to a single genus. On the basis of the 16S rRNA, rpoB and rbcLX gene sequences, the Anabaena/Aphanizomenon strains (cluster 1) were divided into nine supported subclusters which could also be separated morphologically, and which therefore might represent different species. Trichormus strains were morphologically and phylogenetically heterogeneous and did not form a monophyletic cluster. These Trichormus strains, which were representatives of three distinct species, might actually belong to three genera according to the evolutionary distances. Nostoc strains were also heterogeneous and seemed to form a monophyletic cluster, which may contain more than one genus. It was found that certain morphological features were stable and could be used to separate different phylogenetic clusters. For example, the width and the length of akinetes were useful features for classification of the Anabaena/Aphanizomenon strains in cluster 1. This morphological and phylogenetic study with fresh isolates showed that the current classification of these anabaenoid genera needs to be revised. PMID- 15653850 TI - Lactobacillus rossii sp. nov., isolated from wheat sourdough. AB - Screening of sourdough lactic acid bacteria for bacteriocin production resulted in the isolation of a Gram-positive, catalase-negative, non-spore-forming, non motile rod bacterium (strain CS1T) that could not be associated with any previously described species. Comparative 16S rRNA gene sequence analysis recognized strain CS1T as a distinct member of the genus Lactobacillus. By a species-specific PCR strategy, five additional strains previously isolated from sourdoughs were found to belong to the same species as strain CS1T, as confirmed by 16S rRNA gene sequence analysis. The closest related species were Lactobacillus durianis, Lactobacillus malefermentans and Lactobacillus suebicus, with which strain CS1T shared 93 % sequence similarity. For a further characterization of strain CS1T, physiological (growth temperature, CO2 production, hydrolysis of arginine, isomeric type of lactate, sugar fermentation) and chemotaxonomic (G+C content and peptidoglycan structure) properties were determined. Phenotypic characterization showed that strain CS1T was a member of the obligately heterofermentative group of the genus Lactobacillus. The DNA G+C content was 44.6 mol%. The peptidoglycan was of the A3alpha (L-Lys-L-Ser-L-Ala2) type. Physiological, biochemical and genotypic data, as well as results of DNA DNA hybridization of genomic DNA with one of the closest phylogenetic relatives, L. durianis (34.3 %), indicated that strain CS1T represents a novel species of the genus Lactobacillus for which the name Lactobacillus rossii sp. nov. is proposed. The type strain of this species is CS1T (=ATCC BAA-822T=DSM 15814T). PMID- 15653851 TI - Roseisalinus antarcticus gen. nov., sp. nov., a novel aerobic bacteriochlorophyll a-producing alpha-proteobacterium isolated from hypersaline Ekho Lake, Antarctica. AB - A Gram-negative, aerobic to microaerophilic rod was isolated from 10 m depths of the hypersaline, heliothermal and meromictic Ekho Lake (East Antarctica). The strain was oxidase- and catalase-positive, metabolized a variety of carboxylic acids and sugars and produced lipase. Cells had an absolute requirement for artificial sea water, which could not be replaced by NaCl. A large in vivo absorption band at 870 nm indicated production of bacteriochlorophyll a. The predominant fatty acids of this organism were 16 : 0 and 18 : 1omega7c, with 3-OH 10 : 0, 16 : 1omega7c and 18 : 0 in lower amounts. The main polar lipids were diphosphatidylglycerol, phosphatidylglycerol and phosphatidylcholine. Ubiquinone 10 was produced. The DNA G+C content was 67 mol%. 16S rRNA gene sequence comparisons indicated that the isolate represents a member of the Roseobacter clade within the alpha-Proteobacteria. The organism showed no particular relationship to any members of this clade but clustered on the periphery of the genera Jannaschia, Octadecabacter and 'Marinosulfonomonas' and the species Ruegeria gelatinovorans. Distinct morphological, physiological and genotypic differences to these previously described taxa supported the description of a new genus and a novel species, for which the name Roseisalinus antarcticus gen. nov., sp. nov. is proposed. The type strain is EL-88T (=DSM 11466T=CECT 7023T). PMID- 15653852 TI - Winogradskyella thalassocola gen. nov., sp. nov., Winogradskyella epiphytica sp. nov. and Winogradskyella eximia sp. nov., marine bacteria of the family Flavobacteriaceae. AB - Three novel heterotrophic, Gram-negative, yellow-pigmented, aerobic, gliding, oxidase- and catalase-positive bacteria were isolated from algae collected in the Gulf of Peter the Great, Sea of Japan. 16S rRNA gene sequence analysis revealed that the strains studied represented members of the family Flavobacteriaceae and showed 93.5-93.8 % similarity with their closest relative, Psychroserpens burtonensis. The DNA G+C content of the strains was 34-37 mol%. The major respiratory quinone was MK-6. The predominant fatty acids were iso-C(15 : 0), anteiso-C(15 : 0), iso-C(15 : 1), iso-C(16 : 0)-3OH and iso-C(17 : 0)-3OH. On the basis of their phenotypic, chemotaxonomic, genotypic and phylogenetic characteristics, the newly described bacteria have been assigned to the new genus Winogradskyella gen. nov., as Winogradskyella thalassocola sp. nov. (type strain, KMM 3907T=KCTC 12221T=LMG 22492T=DSM 15363T), Winogradskyella epiphytica sp. nov. (type strain, KMM 3906T=KCTC 12220T=LMG 22491T=CCUG 47091T) and Winogradskyella eximia sp. nov. (type strain, KMM 3944T (=KCTC 12219T=LMG 22474T). PMID- 15653853 TI - Application of sliding-window discretization and minimization of stochastic complexity for the analysis of fAFLP genotyping fingerprint patterns of Vibrionaceae. AB - Minimization of stochastic complexity (SC) was used as a method for classification of genotypic fingerprints. The method was applied to fluorescent amplified fragment length polymorphism (fAFLP) fingerprint patterns of 507 Vibrionaceae representatives. As the current BinClass implementation of the optimization algorithm for classification only works on binary vectors, the original fingerprints were discretized in a preliminary step using the sliding window band-matching method, in order to maximally preserve the information content of the original band patterns. The novel classification generated using the BinClass software package was subjected to an in-depth comparison with a hierarchical classification of the same dataset, in order to acknowledge the applicability of the new classification method as a more objective algorithm for the classification of genotyping fingerprint patterns. Recent DNA-DNA hybridization and 16S rRNA gene sequence experiments proved that the classification based on SC-minimization forms separate clusters that contain the fAFLP patterns for all representatives of the species Enterovibrio norvegicus, Vibrio fortis, Vibrio diazotrophicus or Vibrio campbellii, while previous hierarchical cluster analysis had suggested more heterogeneity within the fAFLP patterns by splitting the representatives of the above-mentioned species into multiple distant clusters. As a result, the new classification methodology has highlighted some previously unseen relationships within the biodiversity of the family Vibrionaceae. PMID- 15653854 TI - Microbacterium halotolerans sp. nov., isolated from a saline soil in the west of China. AB - A Gram-positive short rod isolated from a saline soil in China was characterized using a polyphasic approach. This actinobacterium grew over a wide salinity range [0-15 % NaCl, 0-20 % KCl and 0-30 % (MgCl2).6H2O (w/v); optimum concentrations for growth were 5 % NaCl, 5-10 % KCl, 10 % (MgCl2).6H2O]. The optimum growth temperature and pH were 28-30 degrees C and 7.0-8.0, respectively. Chemotaxonomic features (peptidoglycan-type B2beta with glycolyl residues; major menaquinones MK 10 and MK-11; predominating iso- and anteiso-branched cellular fatty acids; DNA G+C content 66.5 mol%) placed this organism within the genus Microbacterium. 16S rRNA gene sequence analysis confirmed this classification of the strain, but showed that it was distinct from its nearest neighbours. It formed a separate branch with type strains of the genus Microbacterium, and also shared low sequence similarity with them (<96 %). Based on phenotypic and molecular taxonomic results, it is proposed that the unknown isolate should be classified as a novel species in the genus Microbacterium, for which the name Microbacterium halotolerans sp. nov. is proposed. The type strain is YIM 70130T (=KCTC 19017T=CIP 108071T). PMID- 15653855 TI - Erythrobacter seohaensis sp. nov. and Erythrobacter gaetbuli sp. nov., isolated from a tidal flat of the Yellow Sea in Korea. AB - Two Gram-negative, non-spore-forming, slightly halophilic strains, SW-135T and SW 161T, which were isolated from a tidal flat of the Yellow Sea in Korea, were subjected to a polyphasic taxonomic study. The two isolates lacked bacteriochlorophyll a and contained ubiquinone-10 (Q-10) as the predominant respiratory lipoquinone and C(18 : 1)omega7c and C(17 : 1)omega6c as the major fatty acids. The DNA G+C contents of strains SW-135T and SW-161T were 62.2 and 64.5 mol%, respectively. Phylogenetic analyses based on 16S rRNA gene sequences showed that the two strains fall within the radiation of the cluster comprising Erythrobacter species. Strains SW-135T and SW-161T exhibited a 16S rRNA gene sequence similarity value of 96.9 % and a mean DNA-DNA relatedness level of 12.3 %. Sequence similarities between strains SW-135T and SW-161T and the type strains of recognized Erythrobacter species ranged from 96.7 to 98.5 %. Levels of DNA-DNA relatedness were low enough to indicate that strains SW-135T and SW-161T represent members of two species separate from all recognized Erythrobacter species. On the basis of polyphasic taxonomic data, strains SW-135T (=KCTC 12228T=DSM 16221T) and SW-161T (=KCTC 12227T=DSM 16225T) were classified as two novel Erythrobacter species, for which the names Erythrobacter seohaensis sp. nov. and Erythrobacter gaetbuli sp. nov. are proposed, respectively. PMID- 15653857 TI - Lactobacillus satsumensis sp. nov., isolated from mashes of shochu, a traditional Japanese distilled spirit made from fermented rice and other starchy materials. AB - Gram-positive, rod-shaped, motile lactic acid bacteria (strains NRIC 0603, NRIC 0604T, NRIC 0605 and NRIC 0606) were isolated from shochu mashes using an enrichment culture approach. These strains clustered in the Lactobacillus casei Pediococcus group and were closely related to Lactobacillus nagelii and Lactobacillus mali on the basis of 16S rRNA gene sequence similarity. Levels of DNA-DNA relatedness revealed genotypic separation of the four isolates from the above two species. The isolates are therefore considered to represent a novel species, for which the name Lactobacillus satsumensis is proposed. The type strain is NRIC 0604T (=JCM 12392T=DSM 16230T). PMID- 15653856 TI - Lactobacillus gastricus sp. nov., Lactobacillus antri sp. nov., Lactobacillus kalixensis sp. nov. and Lactobacillus ultunensis sp. nov., isolated from human stomach mucosa. AB - In an attempt to study the composition of the Lactobacillus flora from mucosa of human stomach, 16S rRNA gene sequences of 129 isolates were analysed. For 15 of these, the results differed significantly from known sequences, and additional tests were performed to determine whether these isolates represented as yet unrecognized species. Phenotypic and genetic characteristics revealed that these isolates represented four novel Lactobacillus species. Two belong to the Lactobacillus reuteri and the other two to the Lactobacillus delbrueckii subgroup of Lactobacillus. The names Lactobacillus gastricus sp. nov., Lactobacillus antri sp. nov., Lactobacillus kalixensis sp. nov. and Lactobacillus ultunensis sp. nov. are proposed, with the respective type strains Kx156A7T (=LMG 22113T=DSM 16045T=CCUG 48454T), Kx146A4T (=LMG 22111T=DSM 16041T=CCUG 48456T), Kx127A2T (=LMG 22115T=DSM 16043T=CCUG 48459T) and Kx146C1T (=LMG 22117T=DSM 16047T=CCUG 48460T). PMID- 15653858 TI - Citricoccus alkalitolerans sp. nov., a novel actinobacterium isolated from a desert soil in Egypt. AB - An actinobacterium, strain YIM 70010T, which was isolated from a desert soil sample collected in Egypt, was subjected to a polyphasic taxonomy study. The organism was alkalitolerant and its optimum growth occurred at pH 8.0-9.0. The isolate contained chemotaxonomic markers that were characteristic of the genus Citricoccus, i.e. the peptidoglycan type Lys-Gly-Glu (variation A4alpha), the predominant menaquinone MK-9(H2) and a polar lipid profile consisting of diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol and two unknown glycolipids. The major fatty acids were anteiso-C(15 : 0) and iso-C(15 : 0). The G+C content of the genomic DNA was 63.8 mol%. Strain YIM 70010T exhibited a 16S rRNA gene sequence similarity of 99.6 % and DNA-DNA relatedness value of 56 % with Citricoccus muralis DSM 14442T. The phenotypic characteristics and DNA-DNA relatedness data indicate that strain YIM 70010T can be distinguished from C. muralis (DSM 14442T). Therefore, on the basis of the polyphasic taxonomic data presented, a novel species of the genus Citricoccus, Citricoccus alkalitolerans sp. nov. (type strain, YIM 70010)=CCTCC AA 203008T=DSM 15665T=KCTC 19012T) is proposed. PMID- 15653859 TI - Planococcus stackebrandtii sp. nov., isolated from a cold desert of the Himalayas, India. AB - The taxonomic position of a bacterium isolated from a cold desert of the Himalayas, India, was analysed by using a polyphasic approach. The isolated strain, designated K22-03T, had phenotypic characteristics that matched those of the genus Planococcus and it represents a novel species. The almost-complete 16S rRNA gene sequence (1464 bases) of the novel strain was compared with those of previously studied Planococcus type strains and confirmed that the strain belongs to the genus Planococcus. 16S rRNA gene sequence analysis indicated that strain K22-03T differs from all other species of Planococcus by at least 2.5 %. DNA-DNA hybridization showed that it had low genomic relatedness with Planomicrobium mcmeekinii (MTCC 3704T, 23 %), Planococcus psychrophilus (MTCC 3812T, 61 %), Planococcus antarcticus (MTCC 3854T, 45 %) and Planomicrobium okeanokoites (MTCC 3703T, 51 %), the four species with which it was most closely related based on 16S rRNA gene sequence analysis (97-97.5 % similarity). Therefore, strain K22-03T should be recognized as a novel species, for which the name Planococcus stackebrandtii sp. nov. is proposed. The type strain is K22-03T (=MTCC 6226T=DSM 16419T=JCM 12481T). PMID- 15653860 TI - Tenuibacillus multivorans gen. nov., sp. nov., a moderately halophilic bacterium isolated from saline soil in Xin-Jiang, China. AB - Two Gram-positive, rod-shaped, spore-forming and moderately halophilic bacteria (strains 28-1T, 28-4), isolated from a soil sample from a neutral salt lake in Xin-Jiang, China, were characterized polyphasically. On the basis of fasta (ungapped) analyses of 16S rRNA gene sequences, strains 28-1T and 28-4 were shown to belong to the Bacillaceae and to be closely related to Filobacillus milensis DSM 13259T (97.0 %) and Bacillus haloalkaliphilus DSM 5271T (95.7 %). 16S rRNA gene sequence similarity with other recognized species was not more than 94.1 %. Phylogenetic, chemotaxonomic, physiological and biochemical data supported the differentiation of these novel strains from F. milensis and B. haloalkaliphilus. Therefore these two previously unidentified strains are considered to represent a new genus and species, for which the name Tenuibacillus multivorans gen. nov., sp. nov. is proposed. The type strain is 28-1T (=AS 1.3442T=NBRC 100370T). PMID- 15653861 TI - Desulfovibrio putealis sp. nov., a novel sulfate-reducing bacterium isolated from a deep subsurface aquifer. AB - A novel sulfate-reducing bacterium was isolated from a well that collected water from a deep aquifer at a depth of 430 m in the Paris Basin, France. The strain, designated B7-43T, was made up of vibrioid cells that were motile by means of a single polar flagellum. Cells contained desulfoviridin. In the presence of sulfate, the following substrates were used as energy and carbon sources: lactate, pyruvate, malate, fumarate, ethanol, butanol, acetate/H2 and glycine. Sulfite and thiosulfate were also used as electron acceptors in the presence of lactate. In the absence of electron acceptors, pyruvate, malate and fumarate were fermented. Optimal growth was obtained in 1 g NaCl l(-1) and at pH 7. On the basis of 16S rRNA gene sequence analysis, the isolate was most closely related to members of the genus Desulfovibrio (90 % similarity). It is thus proposed that strain B7-43T (=DSM 16056T=ATCC BAA-905T) represents a novel species within this genus, Desulfovibrio putealis sp. nov. PMID- 15653862 TI - Thalassobacter stenotrophicus gen. nov., sp. nov., a novel marine alpha proteobacterium isolated from Mediterranean sea water. AB - A Gram-negative, slightly halophilic, strictly aerobic, chemo-organotrophic bacterium was isolated from Mediterranean sea water near Valencia (Spain). 16S rRNA gene sequence comparisons showed that the isolate represented a separate branch within the alpha-3 subclass of the Proteobacteria, now included within the order 'Rhodobacterales'. Jannaschia helgolandensis was the closest relative, but their low sequence similarity and other features indicated that they were not related at the genus level. Isolate 5SM22T produced bacteriochlorophyll a and grew on solid media as regular salmon-pink colonies. Cells are motile rods, with polar flagella. The DNA G+C content is 59.1 mol%. Morphological, physiological and genotypic differences from related, thus far known genera support the description of Thalassobacter stenotrophicus gen. nov., sp. nov. with strain 5SM22T (=CECT 5294T=DSM 16310T) as the type strain. PMID- 15653863 TI - Bacillus arenosi sp. nov., Bacillus arvi sp. nov. and Bacillus humi sp. nov., isolated from soil. AB - A group of nine Gram-positive endospore-forming bacteria was isolated from soil of the Drentse A agricultural research area in The Netherlands. Using (GTG)5-PCR genomic fingerprinting and fatty acid analysis, the nine isolates were divided into three consistent groups. On the basis of 16S rRNA gene sequence similarity of representative strains, the nine isolates were shown to belong to the genus Bacillus. The first group of four isolates was most closely related to Bacillus carboniphilus (95.5 %) and Bacillus sporothermodurans (95.5 %). The second and third groups of three and two isolates, respectively, showed highest sequence similarity to Bacillus neidei (97.0 and 97.1 %, respectively) and Bacillus pycnus (both 96.7 %). A DNA-DNA relatedness study confirmed the consistency of the three groups delineated by (GTG)5-PCR and fatty acid analysis. A small number of phenotypic characters allowed differentiation of the three groups of isolates. The three groups therefore represent novel species, for which the names Bacillus humi, Bacillus arenosi and Bacillus arvi are proposed, with LMG 22167T (=DSM 16318T), LMG 22166T (=DSM 16319T) and LMG 22165T (=DSM 16317T) as the respective type strains. PMID- 15653865 TI - Chryseobacterium daecheongense sp. nov., isolated from freshwater lake sediment. AB - A novel nitrate-reducing bacterium, CPW406T, was isolated from the sediment of a shallow, freshwater lake. The strain was a Gram-negative, non-motile, non-spore forming rod, which formed yellow-pigmented colonies on nutrient agar and contained a polyamine pattern with sym-homospermidine as the major compound, MK-6 as the predominant menaquinone, 15 : 0 iso and 17 : 0 iso 3-OH as the major fatty acids and phosphatidylethanolamine and several unknown lipids in the polar lipid profile. The 16S rRNA gene sequence of strain CPW406T was found to be most similar to that of the type strain of Chryseobacterium defluvii (DSM 14219T; 97.9 % similarity). However, DNA-DNA relatedness data and its phenotypic properties showed that strain CPW406T could be distinguished from all known Chryseobacterium species and thus represented a novel species, for which the name Chryseobacterium daecheongense sp. nov. is proposed; the type strain is CPW406T (=DSM 15235T=KCTC 12088T). PMID- 15653864 TI - Study of mural painting isolates, leading to the transfer of 'Bacillus maroccanus' and 'Bacillus carotarum' to Bacillus simplex, emended description of Bacillus simplex, re-examination of the strains previously attributed to 'Bacillus macroides' and description of Bacillus muralis sp. nov. AB - A group of 24 strains was isolated from deteriorated mural paintings situated in Spain (necropolis of Carmona) and Germany (church of Greene-Kreiensen). (GTG)5 PCR genomic fingerprinting was performed on these strains to assess their genomic variability and the strains were delineated into four groups. Representatives were studied by 16S rRNA gene sequencing and were found to be closely related to Bacillus simplex and the species 'Bacillus macroides' (strain NCIMB 8796) and 'Bacillus maroccanus' (names not validly published) according to a fasta search. The close similarity between B. simplex, 'B. macroides' NCIMB 8796, 'B. maroccanus' and the mural painting isolates was confirmed by additional (GTG)5 PCR, ARDRA, FAME and SDS-PAGE analyses. Furthermore, these techniques revealed that strains of 'Bacillus carotarum', another name that has not been validly published, also showed high similarity to this group of organisms. On the other hand, it was shown that the strains labelled 'B. macroides' in different collections do not all belong to the same species. Strain NCIMB 8796 can be allocated to B. simplex, while strain DSM 54 (=ATCC 12905) shares the highest 16S rRNA gene sequence similarity with Bacillus sphaericus and Bacillus fusiformis (both around 98.6 %). On the basis of further DNA-DNA hybridization data and the study of phenotypic characteristics, one group of five mural painting strains was attributed to a novel species in the genus Bacillus, for which the name Bacillus muralis sp. nov. is proposed. Finally, the remaining mural painting strains, one (LMG 18508=NCIMB 8796) of two strains belonging to 'B. macroides' and strains belonging to 'B. maroccanus' and 'B. carotarum' are allocated to the species B. simplex and an emended description of B. simplex is given. PMID- 15653866 TI - Reclassification of strain CCM 132, previously classified as Kocuria varians, as Kocuria carniphila sp. nov. AB - A Gram-positive actinobacterium, previously classified as Kocuria varians, was subjected to a polyphasic taxonomic study. The bacterium showed the peptidoglycan type Lys-Ala3 (variation A3alpha), MK-7(H2) was the major menaquinone and anteiso C(15 : 0) and anteiso-C(17 : 0) were the major fatty acids. On the basis of the phylogenetic and phenotypic characteristics of the actinobacterium, a novel species, Kocuria carniphila sp. nov. (type strain, CCM 132T=DSM 16004T), is proposed. PMID- 15653867 TI - Marinobacter bryozoorum sp. nov. and Marinobacter sediminum sp. nov., novel bacteria from the marine environment. AB - Two marine, Gram-negative, aerobic, halophilic strains, designated KMM 3657T and KMM 3840T, were isolated and found to be phylogenetically closely related to each other, showing 96.6 % 16S rRNA gene sequence similarity. Both strains are members of the genus Marinobacter in the gamma-Proteobacteria (94.7-98.0 % 16S rRNA gene sequence similarity). Strain KMM 3657T and Marinobacter lipolyticus SM19T were closely related, with 98.0 % sequence similarity. The novel strains shared generic physiological and chemotaxonomic properties with Marinobacter species, but differed in their temperature range for growth, inability to grow in 20 % NaCl and at >43 degrees C, metabolic properties and fatty acid composition. On the basis of phenotypic and phylogenetic analysis data, it is proposed that the strains represent two novel species, Marinobacter bryozoorum sp. nov., with the type strain KMM 3840T (=50-11T=DSM 15401T), and Marinobacter sediminum sp. nov., with the type strain KMM 3657T (=R65T=DSM 15400T). PMID- 15653868 TI - Halorubrum alkaliphilum sp. nov., a novel haloalkaliphile isolated from a soda lake in Xinjiang, China. AB - A novel haloalkaliphilic archaeon, strain DZ-1T, was isolated from a soda lake in Xinjiang, China. The taxonomy of strain DZ-1T was studied by polyphasic methods. According to 16S rRNA gene sequence similarity, strain DZ-1T was phylogenetically related to Halorubrum tibetense (97.5 %), Halorubrum vacuolatum (95.7 %) and Halorubrum saccharovorum (95.9 %). Strain DZ-1T was able to grow at 20-44 degrees C and was also physiologically different from the above-mentioned species with respect to assimilation of sugars and utilization of organic acids. The DNA G+C content of strain DZ-1T was 62.1 mol% (Tm). The DNA-DNA relatedness of strain DZ 1T to H. tibetense and H. vacuolatum was 22 and 13 %, respectively. It was concluded that strain DZ-1T represents a novel species of the genus Halorubrum, for which the name Halorubrum alkaliphilum (type strain, DZ-1T=AS 1.3528T=JCM 12358T) is proposed. PMID- 15653869 TI - Agromyces salentinus sp. nov. and Agromyces neolithicus sp. nov. AB - A polyphasic study was carried out to clarify the taxonomic position of two Gram positive bacteria isolated from soil samples of the Grotta dei Cervi (Italy), a relatively unexplored hypogean environment. The strains, 20-5T and 23-23T, showed phenotypic and phylogenetic characteristics that were consistent with their classification in the genus Agromyces. 16S rRNA gene sequence comparisons revealed that the two strains formed distinct phyletic lines within the genus Agromyces. Based on 16S rRNA gene sequence similarity, chemotaxonomic data and the results of DNA-DNA relatedness studies, it is proposed that the two isolates represent two novel species of the genus Agromyces. Pronounced differences in a broad range of phenotypic characteristics and DNA G+C content distinguished the two strains from each other and from previously described species of the genus Agromyces. Two novel species are proposed: Agromyces salentinus sp. nov. (type strain, 20-5T=HKI 0320T=DSM 16198T=NCIMB 13990T) and Agromyces neolithicus sp. nov. (type strain, 23-23T=HKI 0321T=DSM 16197T=NCIMB 13989T). PMID- 15653870 TI - Bacillus saliphilus sp. nov., isolated from a mineral pool in Campania, Italy. AB - A haloalkaliphilic Gram-positive bacterium, strain 6AGT, that grew aerobically at an optimum temperature of 37 degrees C and at pH 7-10 (optimum 9.0), was isolated from algal mat from a mineral pool located in Malvizza in the Campania region (southern Italy). The isolate tolerated high concentrations of NaCl, up to 25 %. On the basis of 16S rRNA gene sequence similarity, the strain was shown to belong to the genus Bacillus. Analysis of the 16S rRNA gene sequence revealed high similarity between strain 6AGT and an unidentified isolate from Hailaer soda lake (China) (99.9 % identity) and two Kenyan isolates, 3E1 and WE4 (98.3 and 97.8 % identity, respectively). The G+C content of the DNA was 48.4 mol%. The predominant respiratory quinones were MK-7(H2), MK-7(H4) and DMK-7(H2); phosphatidylglycerol and diphosphatidylglycerol were the predominant polar lipids. iC15 : 0 and aiC15 : 0 were the major fatty acids. Strain 6AGT accumulated osmolytes. The phylogenetic distance of strain 6AGT (=DSM 15402T=ATCC BAA-957T) from any recognized species within the genus Bacillus allowed it to be classified as the type strain of Bacillus saliphilus sp. nov. PMID- 15653871 TI - Pontibacillus chungwhensis gen. nov., sp. nov., a moderately halophilic Gram positive bacterium from a solar saltern in Korea. AB - Three moderately halophilic, spore-forming strains, designated BH030062T, BH030049 and BH030080, were isolated from a solar saltern in Korea. Phylogenetic analyses and comparative 16S rRNA gene sequence studies revealed that the isolates represent a novel distinct monophyletic lineage within the phyletic group classically defined as the genus Bacillus and are most closely related to members of the genera Gracilibacillus (93.7-95.1 % similarity), Virgibacillus (93.5-94.8 %), Halobacillus (94.8-95.9 %), Filobacillus (94.4-94.8 %) and Lentibacillus (93.3-93.7 %). Strain BH030062T was strictly aerobic, rod-shaped, Gram-positive and motile by means of peritrichous flagella. It grew in the presence of 1-15 % (w/v) NaCl and at temperatures of 15-45 degrees C. The cell wall peptidoglycan contained A1gamma-meso-diaminopimelic acid as the diagnostic diamino acid. The predominant cellular fatty acids were iso-C(15 : 0), anteiso C(15 : 0) and iso-C(16 : 0). DNA G+C content was about 41 mol% and the major isoprenoid quinone was MK-7. On the basis of their physiological and molecular properties, the isolates represent a new genus, Pontibacillus gen. nov., and novel species, Pontibacillus chungwhensis sp. nov. The type strain is BH030062T (=KCTC 3890T=DSM 16287T). PMID- 15653872 TI - Psychrobacter alimentarius sp. nov., isolated from squid jeotgal, a traditional Korean fermented seafood. AB - Two Gram-negative, non-motile, non-spore-forming, moderately halophilic strains, JG-100T and JG-102, were isolated from squid jeotgal, a traditional Korean fermented seafood. The two strains grew optimally at 30 degrees C and in the presence of 2-3 % (v/w) NaCl. Strains JG-100T and JG-102 were characterized chemotaxonomically; they both had ubiquinone-8 as the predominant respiratory lipoquinone and C(18 : 1)omega9c as the major fatty acid. Their DNA G+C content was 44 mol%. Strains JG-100T and JG-102 showed 1 bp difference in their 16S rRNA gene sequences and a mean DNA-DNA relatedness level of 88 %. Phylogenetic analysis based on 16S rRNA gene sequences showed that strains JG-100T and JG-102 form a distinct phylogenetic lineage within the cluster comprising Psychrobacter species. The 16S rRNA gene sequences of strains JG-100T and JG-102 had similarity levels of 95.2-98.4 % to sequences of the type strains of recognized Psychrobacter species. Levels of DNA-DNA relatedness between strains JG-100T and JG-102 and the type strains of some phylogenetically related Psychrobacter species were 6-24 %. On the basis of phenotypic and phylogenetic data and genomic distinctiveness, strains JG-100T and JG-102 should be placed in the genus Psychrobacter as a novel species, for which the name Psychrobacter alimentarius sp. nov. (type strain, JG-100T=KCTC 12186T=DSM 16065T) is proposed. PMID- 15653873 TI - Pibocella ponti gen. nov., sp. nov., a novel marine bacterium of the family Flavobacteriaceae isolated from the green alga Acrosiphonia sonderi. AB - A marine, heterotrophic, Gram-negative, aerobic, yellow-pigmented, bacterium that was motile by gliding, isolated from the green alga Acrosiphonia sonderi, was studied by polyphasic taxonomic methods. 16S rRNA gene sequence analysis indicated that strain KMM 6031T formed a distinct lineage within the family Flavobacteriaceae. On the basis of phenotypic, chemotaxonomic, genotypic and phylogenetic analyses, the novel bacterium was classified as Pibocella ponti gen. nov., sp. nov. The type strain is KMM 6031T (=KCTC 12262T=NBRC 100591T=LMG 22573T). PMID- 15653875 TI - Bacillus velezensis sp. nov., a surfactant-producing bacterium isolated from the river Velez in Malaga, southern Spain. AB - Two Gram-positive, endospore-forming bacterial strains, CR-502T and CR-14b, which produce surfactant molecules are described. Phenotypic tests and phylogenetic analyses showed these strains to be members of the genus Bacillus and related to the species Bacillus atrophaeus, Bacillus mojavensis, Bacillus subtilis, Bacillus vallismortis and Bacillus amyloliquefaciens, although they differ from these species in a number of phenotypic characteristics. DNA-DNA hybridization confirmed that they show less than 20 % hybridization with the above-mentioned species and therefore represent a novel species of Bacillus. The DNA G+C content is 46.4 mol% in strain CR-502T and 46.1 mol% in strain CR-14b. The main fatty acids in strain CR-502T are 15 : 0 anteiso (32.70 %), 15 : 0 iso (29.86 %) and 16 : 0 (13.41 %). The main quinone in strain CR-502T is MK-7 (96.6 %). In the light of the polyphasic evidence gathered in this study, it is proposed that these strains be classified as a novel species of the genus Bacillus, with the name Bacillus velezensis sp. nov. The type strain (CR-502T=CECT 5686T=LMG 22478T) was isolated from a brackish water sample taken from the river Velez at Torredelmar in Malaga, southern Spain. PMID- 15653874 TI - Lebetimonas acidiphila gen. nov., sp. nov., a novel thermophilic, acidophilic, hydrogen-oxidizing chemolithoautotroph within the 'Epsilonproteobacteria', isolated from a deep-sea hydrothermal fumarole in the Mariana Arc. AB - A novel thermophilic, acidophilic bacterium, designated strain Pd55T, was isolated from a self-temperature-recording in situ colonization system deployed in a hydrothermal diffusing flow (maximum temperature of 78 degrees C) at the TOTO caldera in the Mariana Arc. Cells of strain Pd55T were motile, short rods with a single polar flagellum. Growth was observed between 30 and 68 degrees C (optimum growth at 50 degrees C; 120 min doubling time) and between (initial) pH 4.2 and 7.0 (optimum at pH 5.2). The isolate was a strictly anaerobic chemolithoautotroph capable of using molecular hydrogen as sole energy source and carbon dioxide as sole carbon source. Elemental sulfur served as the sole electron acceptor to support growth. The G+C content of the genomic DNA was 34.0 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that the isolate was related to members of the genera Nautilia and Caminibacter, although it appeared to be a novel lineage prior to the divergence between Nautilia and Caminibacter. Strain Pd55T could also be differentiated from Nautilia and Caminibacter species on the basis of its physiological properties. It is, therefore, proposed that strain Pd55T (=JCM 12420T=DSM 16356T) represents the type strain of a novel species in a new genus, Lebetimonas acidiphila gen. nov., sp. nov. PMID- 15653876 TI - Polyphasic study of wine Lactobacillus strains: taxonomic implications. AB - One hundred and seventy-eight lactobacilli isolated from wine were characterized by a polyphasic approach. Strains were phenotypically identified at genus and species level by classical tests including the analysis of cell morphology, homo/heterofermentative character, sugar fermentation patterns, growth at different temperatures and the optical nature of the isomer of lactic acid produced from glucose. Molecular techniques such as random amplification of polymorphic DNA (RAPD), amplified 16S rDNA restriction analysis (16S-ARDRA), PFGE RFLP and ribotyping were used to characterize strains, and their potential for identification and/or typing was evaluated. The information obtained with these techniques was processed with the BioNumerics software in order to analyse relationships existing between isolated strains and various reference species of the genus. Then, taxonomic dendrograms were obtained, and this information allowed the proposal of molecular procedures suitable for the identification and typing of these wine micro-organisms. The techniques useful for both identification and typing were RAPD and ribotyping, while 16S-ARDRA was only useful for identification and PFGE-RFLP only for typing purposes. The wine strains were identified as Lactobacillus brevis (19 strains), Lactobacillus collinoides (2 strains), Lactobacillus hilgardii (71 strains), Lactobacillus paracasei (13 strains), Lactobacillus pentosus (2 strains), Lactobacillus plantarum (34 strains) and Lactobacillus mali (10 strains). PMID- 15653877 TI - Emended description of porcine [Pasteurella] aerogenes, [Pasteurella] mairii and [Actinobacillus] rossii. AB - The aim of this study was to improve the definition and identification of a group of veterinarily important bacteria referred to as the [Pasteurella] aerogenes [Pasteurella] mairii-[Actinobacillus] rossii complex. These organisms have mainly been isolated from the reproductive and intestinal tracts of pigs and in most cases have been considered as opportunistic pathogens. A collection of 87 strains were characterized by phenotypic analysis from which 41 strains were selected for 16S rRNA gene sequence comparison, out of which 23 have been sequenced in the present study. One group of 21 strains phenotyped as biovars 1, 3-5, 9-11, 19 and 25-27, including the type strain of [P.] aerogenes, showed 16S rRNA gene sequence similarities of 99.6 % or higher; another group of 18 strains including biovars 2, 6-8, 12-15, 21, 23, 24 and 26A and the type strain of [A.] rossii showed 97.8 % or higher 16S rRNA gene sequence similarity. Between the two groups, 93.8-95.7 % 16S rRNA gene sequence similarity was observed. Strains of [P.] mairii showed 99.5 % similarity, with 95.5-97.2 and 93.8-95.5 % similarity to strains of [P.] aerogenes and [A.] rossii, respectively. Four strains could not be classified with any of these groups and belonged to other members of Pasteurellaceae. Comparisons were also made to DNA-DNA hybridization data. Biovars 1, 9, 10, 11 and 19, including the type strain of [P.] aerogenes, linked at 70 % DNA reassociation, whereas strains identified as biovars 2, 6, 7, 8, 12, 15 and 21 of [P.] aerogenes linked at 81 %. The latter group most likely represents [A.] rossii based on the 16S rRNA gene sequence comparisons. DNA reassociation between the [P.] aerogenes and [A.] rossii groups was at most 37 %, whereas 47 % was the highest DNA reassociation found between [P.] aerogenes and [P.] mairii. The study showed that [P.] aerogenes, [P.] mairii and [A.] rossii can not be easily separated and may consequently be misidentified based on current knowledge of their phenotypic characteristics. In addition, these taxa are difficult to separate from other taxa of the Pasteurellaceae. A revised scheme for separation based upon phenotypic characters is suggested for the three species [P.] aerogenes emend., [P.] mairii emend. and [A.] rossii emend., with the respective type strains ATCC 27883T, NCTC 10699T and ATCC 27072T. PMID- 15653878 TI - Description of Aquimarina muelleri gen. nov., sp. nov., and proposal of the reclassification of [Cytophaga] latercula Lewin 1969 as Stanierella latercula gen. nov., comb. nov. AB - The taxonomic position of three novel sea-water isolates was determined. The strains studied were strictly aerobic, heterotrophic, pigmented, motile by gliding, Gram-negative and oxidase-, catalase-, beta-galactosidase- and alkaline phosphatase-positive. 16S rRNA gene sequence phylogenetic analysis indicated that the strains KMM 6020T, KMM 6021 and KMM 6028 occupied a distinct lineage within the family Flavobacteriaceae. The major respiratory quinone was MK-6. The predominant fatty acids were i15 : 0, i15 : 1, i15 : 0 3-OH, i17 : 1omega9c and i17 : 0 3-OH. On the basis of phenotypic, chemotaxonomic, genotypic and phylogenetic characteristics, the novel bacteria were assigned to the genus Aquimarina gen. nov., as Aquimarina muelleri gen. nov., sp. nov. The type strain is KMM 6020T (=KCTC 12285T=LMG 22569T). From the results of the 16S rRNA gene sequence analysis and phenotypic features, the species [Cytophaga] latercula Lewin 1969 is proposed to be reclassified in the new genus Stanierella as Stanierella latercula gen. nov., comb. nov., with type strain CIP 104806T (=ATCC 23177T=NCIMB 1399T=LMG 1343T). PMID- 15653880 TI - Salegentibacter mishustinae sp. nov., isolated from the sea urchin Strongylocentrotus intermedius. AB - A bacterial strain, designated KMM 6049T, was isolated from the sea urchin Strongylocentrotus intermedius inhabiting the Sea of Japan. The bacterium studied was strictly aerobic, heterotrophic, yellow-pigmented, non-motile, Gram-negative and oxidase-, catalase-, beta-galactosidase- and alkaline phosphatase-positive. 16S rRNA gene sequence analysis indicated that strain KMM 3524T was closely related to Salegentibacter holothuriorum and Salegentibacter salegens (sharing 97.7 and 98 % sequence similarity, respectively). DNA-DNA relatedness levels between strains KMM 6049T and S. holothuriorum KMM 3524T and S. salegens DSM 5424T were 24 and 45 %, respectively, indicating that KMM 6049T belongs to a novel species of the genus Salegentibacter, for which the name Salegentibacter mishustinae sp. nov. is proposed. The type strain is KMM 6049T (=KCTC 12263T=LMG 22584T=NBRC 100592T). PMID- 15653879 TI - Roseivirga ehrenbergii gen. nov., sp. nov., a novel marine bacterium of the phylum 'Bacteroidetes', isolated from the green alga Ulva fenestrata. AB - The taxonomic position of a novel marine bacterium isolated from the green alga Ulva fenestrata collected in the Sea of Japan was established. Cells of the strain studied, designated KMM 6017T, were strictly aerobic, heterotrophic, pink pigmented, non-motile by gliding, Gram-negative and oxidase-, catalase-, beta galactosidase- and alkaline phosphatase-positive. 16S rRNA gene sequence analysis indicated that the strain occupied a distinct lineage within the phylum 'Bacteroidetes' and formed a cluster with [Flexibacter] tractuosus and Reichenbachia agariperforans. The G+C content of the DNA of KMM 6017T was 40.2 mol%. The major respiratory quinone was MK-7. The predominant fatty acids were i15 : 1, i15 : 0 and i17 : 0 3-OH (34.2, 24 and 7.7 %, respectively). On the basis of phenotypic, chemotaxonomic, genotypic and phylogenetic characteristics, the novel bacterium was assigned to the genus Roseivirga gen. nov., as Roseivirga ehrenbergii gen. nov., sp. nov. The type strain is KMM 6017T (=KCTC 12282T=LMG 22567T). PMID- 15653881 TI - Salinimonas chungwhensis gen. nov., sp. nov., a moderately halophilic bacterium from a solar saltern in Korea. AB - A halophilic non-spore-forming bacterium of the gamma-Proteobacteria, designated strain BH030046T, was isolated from a solar saltern in Korea. Cells were Gram negative, chemoheterotrophic, short rod-shaped and motile with a polar flagellum. Comparative 16S rRNA gene sequence analysis revealed that strain BH030046T formed a distinct lineage in the family Alteromonadaceae and was distinguished from its closest related genera Alteromonas (91.4-94.8 %), Aestuariibacter (92.1-93.5 %) and Glaciecola (92.1-93.5 %) on the basis of low 16S rRNA gene sequence similarities. Physiological and biochemical data also showed that the isolate was different from members of these three genera. The predominant cellular fatty acids were C(16 : 0) and C(18 : 1)omega7c. DNA G+C content was 48 mol% and the major isoprenoid quinone was Q-8. The strain grew optimally at 30-35 degrees C, pH 7.0-8.0 and 2-5 % NaCl. On the basis of physiological and molecular properties, strain BH030046T (=KCTC 12239T=DSM 16280T) represents a novel genus and species in the family Alteromonadaceae, for which the name Salinimonas chungwhensis gen. nov., sp. nov. is proposed. PMID- 15653882 TI - Cryptanaerobacter phenolicus gen. nov., sp. nov., an anaerobe that transforms phenol into benzoate via 4-hydroxybenzoate. AB - An anaerobic bacterium that transforms phenol and 4-hydroxybenzoate (4-OHB) into benzoate, strain LR7.2T, was isolated from a culture originating from a mixture of swamp water, sewage sludge, swine waste and soil. Cells of strain LR7.2T are Gram-positive short rods (1 x 2 microm) that are electron-dense when observed by electron microscopy. The optimum pH and temperature for growth and transformation activity of 4-OHB are 7.5-8.0 and 30-37 degrees C, respectively. The bacterium does not use sulphate, thiosulphate, nitrate, nitrite, FeCl3, fumarate or arsenate as an electron acceptor. It does not normally use sulphite, although stimulation of growth and 4-OHB transformation activity at a low concentration (up to 2 mM) has been reported previously under different culture conditions. The presence of 4-OHB or phenol is essential for growth; transformation of 4-OHB or phenol into benzoate is used to produce energy for growth. Using [6D]-phenol, 4 OHB was shown to be an intermediate in the transformation of phenol into benzoate. No spore was observed. The bacterium has a DNA G+C content of 51 mol% and its major membrane fatty acid is anteiso-C(15 : 0). The 16S rRNA gene sequence of strain LR7.2T shows only 90 % similarity to its closest relative (Pelotomaculum thermopropionicum). From these results, a new taxon is proposed: Cryptanaerobacter phenolicus gen. nov., sp. nov. The type strain is LR7.2T (=ATCC BAA-820T=DSM 15808T). PMID- 15653883 TI - Advenella incenata gen. nov., sp. nov., a novel member of the Alcaligenaceae, isolated from various clinical samples. AB - A polyphasic taxonomic study of 14 isolates recovered from various human and veterinary clinical samples was performed. Phenotypically these isolates shared several characteristics with members of the Alcaligenaceae and related genera. Random amplified polymorphic DNA fingerprinting and whole-cell protein analysis suggested the presence of multiple genomic groups, which was confirmed by DNA-DNA hybridization experiments. 16S rRNA gene sequence analysis indicated that these isolates were related to the genera Pelistega, Taylorella, Oligella, Pigmentiphaga, Alcaligenes, Kerstersia, Achromobacter and Bordetella and belonged to the family Alcaligenaceae. Based on the results of the present study the organisms were classified in a novel genus, Advenella gen. nov. This genus comprises one named species, Advenella incenata sp. nov. (type strain LMG 22250T=CCUG 45225T) and five currently unnamed genomic species. The DNA G+C content of members of the novel genus Advenella is between 54.0 and 57.7 mol%. PMID- 15653884 TI - Streptomyces koyangensis sp. nov., a novel actinomycete that produces 4-phenyl-3 butenoic acid. AB - A 4-phenyl-3-butenoic acid-producing actinomycete, designated strain VK-A60T, was isolated from a soil sample collected from Koyang, Korea. Morphological and chemical characteristics of the strain were consistent with those of the genus Streptomyces. The cell wall of the strain contains LL-diaminopimelic acid. The predominant fatty acids are anteiso-C(15 : 0), iso-C(16 : 0) and C(16 : 0). The strain formed a distinct monophyletic line within the 16S rRNA gene sequence phylogenetic tree. Analyses of its morphological, physiological and biochemical characteristics, together with random amplified polymorphic DNA and DNA-DNA relatedness data, confirmed that strain VK-A60T represents a novel Streptomyces taxon that is distinguishable from closely related reference strains. Strain VK A60T (=KCCM 10555T=NBRC 100598T) is proposed as the type strain of a novel species, for which the name Streptomyces koyangensis sp. nov. is proposed. PMID- 15653886 TI - Streptococcus marimammalium sp. nov., isolated from seals. AB - Two strains of an unidentified, Gram-positive, catalase-negative, chain-forming, coccus-shaped organism recovered from seals were characterized using phenotypic and molecular taxonomic methods. Based on morphological and biochemical criteria the strains were tentatively identified as streptococci but they did not appear to correspond to any recognized species of the genus Streptococcus. Comparative 16S rRNA gene sequencing studies showed that the strains were closely related to each other and confirmed their placement in the genus Streptococcus. Sequence divergence values of >5 % with reference streptococcal species demonstrated the organisms from seals represent a novel species. SDS-PAGE analysis of whole-cell proteins confirmed that the two organisms were closely related to each other but were different from all currently defined streptococcal species. Based on biochemical criteria, molecular chemical and molecular genetic evidence, it is proposed that the unknown isolates from seals be assigned to a novel species of the genus Streptococcus, Streptococcus marimammalium sp. nov. The type strain is M54/01/1T (=CCUG 48494T=CIP 108309T). PMID- 15653885 TI - Phylogenetic analysis of the genera Bradyrhizobium, Mesorhizobium, Rhizobium and Sinorhizobium on the basis of 16S rRNA gene and internally transcribed spacer region sequences. AB - A total of 128 strains was isolated from more than 23 legume hosts in Korea. Phylogenetic relationships between these Korean isolates and reference strains of the genera Bradyrhizobium, Mesorhizobium, Rhizobium and Sinorhizobium were analysed using their 16S rRNA gene and internally transcribed spacer (ITS) region sequences. Among the Bradyrhizobium strains, dendrograms based on both the 16S rRNA gene and ITS region sequences produced two main groups. The ITS tree yielded at least two new clusters that were discernable from the seven previously delineated genospecies. Large discrepancies were revealed between phylogenetic dendrograms based on 16S rRNA gene and ITS region sequences for members of the genus Rhizobium, reflecting their taxonomic heterogeneity. The amalgamation of Rhizobium and former members of Agrobacterium was confirmed using the 16S rRNA tree. Phylogenetic analysis of ITS region sequences showed that the Rhizobium giardinii clade (group II) and the Rhizobium radiobacter/Rhizobium rubi clade (group III) could be tentatively recognized as groups that are separable from the core group (group I), which includes Rhizobium leguminosarum. Dendrograms based on the 16S rRNA gene and ITS region sequences of Mesorhizobium strains were highly conflicting due to the poor taxonomic resolution of the 16S rRNA gene sequences and the low confidence in the ITS dendrogram. Several Korean isolates within the genus Mesorhizobium are thought to represent novel taxa when considering their relatively low ITS region sequence similarities (<80 %) to the reference strains. PMID- 15653887 TI - Marinomonas pontica sp. nov., isolated from the Black Sea. AB - A Gram-negative, polarly flagellated bacterium was isolated from a sea-water sample collected from the Karadag Natural Reserve of the Eastern Crimea and characterized to clarify its taxonomic position. 16S rRNA gene sequence-based phylogenetic analysis of this novel organism revealed Marinomonas vaga, Marinomonas communis, Marinomonas mediterranea, Marinomonas primoryensis and 'Marinomonas protea' as its closest relatives (similarity 95-97 %). The G+C content of the DNA was 46.5 mol%. The organism grew between 4 and 33 degrees C, tolerated 10 % NaCl, was slightly alkaliphilic and was not able to degrade starch, gelatin, agar or Tween 80. Phosphatidylethanolamine (53.4 %) and phosphatidylglycerol (46.6 %) were the predominant phospholipids. The major fatty acids were 16 : 0 (15.5 %), 16 : 1omega7 (26.7 %) and 18 : 1omega7 (47.1 %). The phylogenetic, genetic and physiological properties of the organism placed it within a novel species, proposed as Marinomonas pontica sp. nov., the type strain of which is 46-16T (=LMG 22531T=KMM 3492T=UCM 11075T). PMID- 15653888 TI - Methylobacterium hispanicum sp. nov. and Methylobacterium aquaticum sp. nov., isolated from drinking water. AB - Members of the genus Methylobacterium are ubiquitous in nature and can be isolated from almost any freshwater environment where dissolved oxygen exists. This genus is composed of a variety of pink-pigmented, facultatively methylotrophic (PPFM) bacteria. During a screening programme to monitor the bacterial population present in the drinking water of a municipal water supply in Seville (Spain) during the year 2003, five strains of PPFM bacteria were isolated and characterized. Analysis of their complete 16S rRNA gene sequences revealed that they constituted two separate phylogenetic groups (strains GP34T and GR18, and strains GR16T, GP22 and GP32, respectively) showing highest similarity to members of the genus Methylobacterium. The highest 16S rRNA sequence similarities of strain GP34T were found with respect to the type strains of Methylobacterium radiotolerans (96.6 %) and Methylobacterium fujisawaense (96.4 %) and the highest 16S rRNA sequence similarities of strain GR16T were to the type strains of Methylobacterium extorquens (96.0 %) and Methylobacterium rhodesianum (95.8 %). The G+C content of their DNA ranged from 66.5 to 67.8 mol%. DNA-DNA hybridization studies confirmed that they constituted two separate genospecies. On the basis of this phenotypic, phylogenetic and genotypic study, two novel species of the genus Methylobacterium are proposed: Methylobacterium hispanicum sp. nov., with type strain GP34T (CECT 5997T=CCM 7219T=DSM 16372T=CIP 108332T), and Methylobacterium aquaticum sp. nov., with type strain GR16T (CECT 5998T=CCM 7218T=DSM 16371T=CIP 108333T). PMID- 15653889 TI - Spiroplasma atrichopogonis sp. nov., from a ceratopogonid biting midge. AB - Spiroplasma sp. strain GNAT3597T was isolated from the biting midge genus Atrichopogon (Diptera: Ceratopogonidae). It was serologically distinct from other Spiroplasma species, groups or subgroups. Dark-field microscopy of the cells revealed the classical helical shape and subsequent transmission electron microscopy revealed cells surrounded by only a cell membrane (i.e. lacking a cell wall). Growth of strain GNAT3597T occurred in M1D medium at 30 degrees C. Strain GNAT3597T catabolized both glucose and arginine, but did not hydrolyse urea. The DNA G+C content of strain GNAT3597T was 29+/-1 mol%. Only one strain, SMCAT (Spiroplasma mirum), is serologically related to strain GNAT3597T, although the relationship is weak (positive reaction to only a 1 : 80 dilution). It is therefore proposed that strain GNAT3597T (=ATCC BAA-520T=NBRC 100390T) represents a novel species, Spiroplasma atrichopogonis sp. nov. (class Mollicutes: order Entomoplasmatales: family Spiroplasmataceae). PMID- 15653890 TI - A multigene approach to phylogenetic analysis using the genus Mycobacterium as a model. AB - Advances in DNA sequencing and the increasing number of sequences available in databases have greatly enhanced the bacterial identification process. Several species within the genus Mycobacterium cause serious human and animal diseases. In order to assess their relative positions in the evolutionary process, four gene fragments, from the 16S rRNA (564 bp), hsp65 (420 bp), rpoB (396 bp) and sod (408 bp) genes, were sequenced from 97 strains, including all available type strains of the genus Mycobacterium. The results demonstrate that, in this case, the concatenation of different genes allows significant increases in the power of discrimination and the robustness of the phylogenetic tree. The sequential and/or combined use of sequences of several genes makes it possible to refine the phylogenetic approach and provides a molecular basis for accurate species identification. PMID- 15653891 TI - 'Candidatus Phytoplasma pini', a novel taxon from Pinus silvestris and Pinus halepensis. AB - Pinus silvestris and Pinus halepensis trees grown in Germany and Spain, respectively, showing abnormal shoot branching, dwarfed needles and other symptoms were examined for the presence of plant-pathogenic mollicutes (phytoplasmas). While phytoplasmas could not be detected unambiguously with microscopical methods, PCR amplification using universal phytoplasma primers yielded positive results. Samples collected from symptomatic and non-symptomatic plant parts of both symptomatic Pinus silvestris and Pinus halepensis trees tested positive. Also, surrounding non-symptomatic trees proved to be phytoplasma infected. Comparisons revealed that the 16S rRNA gene sequences of the phytoplasmas identified in Pinus silvestris and Pinus halepensis were nearly identical. However, the pine phytoplasma is only distantly related to other phytoplasmas. The closest relatives are members of the palm lethal yellowing and rice yellow dwarf groups and 'Candidatus Phytoplasma castaneae', which share between 94.5 and 96.6 % 16S rRNA gene sequence similarity. From these data it can be concluded that the phytoplasmas identified in the two Pinus species represent a coherent but discrete taxon; it is proposed that this taxon be distinguished at putative species level under the name 'Candidatus Phytoplasma pini'. PMID- 15653892 TI - Marinomonas ushuaiensis sp. nov., isolated from coastal sea water in Ushuaia, Argentina, sub-Antarctica. AB - A Gram-negative, rod-shaped, psychrophilic, motile, non-spore-forming bacterium, strain U1T, was isolated from Ushuaia located at the southernmost tip of Argentina. On the basis of 16S rRNA gene sequence similarity, strain U1T was found to be closely related to Marinomonas communis (DSM 5604T) and Marinomonas primoryensis (IAM 15010T). At the DNA-DNA level, however, the values for similarity were 41 and 25 %, respectively. The major fatty acids present were iso C(16 : 0), C(16 : 1)omega7c, iso-C(17 : 1) and C(18 : 1)omega7c and the G+C content of the DNA was 43.6 mol%. All of the above characteristics support the affiliation of strain U1T to the genus Marinomonas. Furthermore, on the basis of phenotypic features, chemotaxonomic characteristics and phylogenetic analysis of the 16S rRNA gene sequence, it appears that strain U1T is distinct from the four Marinomonas species with validly published names. Strain U1T, therefore, represents a novel species, for which the name Marinomonas ushuaiensis sp. nov. is proposed. The type strain of M. ushuaiensis is U1T (=MTCC 6143T=DSM 15871T=JCM 12170T). PMID- 15653894 TI - Gillisia mitskevichiae sp. nov., a novel bacterium of the family Flavobacteriaceae, isolated from sea water. AB - The taxonomic position of a novel marine, heterotrophic, aerobic, pigmented bacterium, non-motile by gliding, that was isolated from a sea-water sample collected in the Sea of Japan, was determined. 16S rRNA gene sequence analysis revealed that strain KMM 6034T is a member of the genus Gillisia. The phenotypic and chemotaxonomic data showed that the isolate represents a novel species of the genus Gillisia, for which the name Gillisia mitskevichiae sp. nov. is proposed. The type strain is KMM 6034T (=KCTC 12261T=NBRC 100590T=LMG 22575T). PMID- 15653893 TI - Isolation of Lentibacillus salicampi strains and Lentibacillus juripiscarius sp. nov. from fish sauce in Thailand. AB - Eight strains of aerobic, spore-forming, Gram-positive, moderately halophilic bacteria were isolated from sauce (nam-pla and bu-du) produced in Thailand by the fermentation of fish. They grew optimally in the presence of 10 % NaCl, at 37 degrees C and pH 7.0. A diagnostic diamino acid, meso-diaminopimelic acid, was present in the cell-wall peptidoglycan. The predominant menaquinone was MK-7. The major cellular fatty acids were anteiso-C(15 : 0) and iso-C(16 : 0). Phosphatidylglycerol, diphosphatidylglycerol and an unidentified glycolipid were found to be the major polar lipid components. The DNA G+C content was 42-43 mol%. These bacteria were further divided into two groups based on phenotypic characteristics and DNA-DNA similarities. Three strains of Group I were highly affiliated to the type strain of Lentibacillus salicampi in terms of phenotypic characterization and DNA-DNA similarities (96-102 %); accordingly, they were identified as strains of L. salicampi. A representative strain of Group II, strain IS40-3T, was most closely related to L. salicampi in terms of 16S rRNA based phylogenetic analysis, although five strains of Group II could be distinguished from L. salicampi by means of several phenotypic properties, low 16S rRNA gene sequence similarity (95.2 %) and low DNA-DNA similarities (12-32 %). Therefore, the Group II strains should be included in a novel species of the genus Lentibacillus, for which the name Lentibacillus juripiscarius sp. nov. is proposed. The type strain is IS40-3T (=JCM 12147T=PCU 229T=TISTR 1535T). PMID- 15653895 TI - Methanobacterium beijingense sp. nov., a novel methanogen isolated from anaerobic digesters. AB - Two methanogenic strains, 8-2T and 4-1, with rod-shaped (0.4-0.5 x 3-5 microm), non-motile cells, sometimes observed in chains, were isolated from two anaerobic digesters in Beijing, China. The two strains used H2/CO2 and formate for growth and produced methane. The temperature range for growth was 25-50 degrees C, with fastest growth at 37 degrees C. The pH ranges for growth and methane production were 6.5-8.0 for strain 8-2T and 6.8-8.6 for strain 4-1, with the fastest growth at pH 7.2 for strain 8-2T and pH 7.5-7.7 for strain 4-1. The G+C content of genomic DNA for strain 8-2T was 38.9 mol%. The similarity levels of the 16S rRNA sequence of strain 8-2T with other species of the genus Methanobacterium ranged from 93.8 to 96.0 %. Based on the phylogenetic analysis and phenotypic characteristics, the novel species Methanobacterium beijingense sp. nov. is proposed, with the type strain 8-2T (=DSM 15999T=CGMCC 1.5011T). PMID- 15653896 TI - Transfer of the misnamed [Alysiella] sp. IAM 14971 (=ATCC 29468) to the genus Moraxella as Moraxella oblonga sp. nov. AB - Phylogenetic analysis based on 16S rRNA gene sequences revealed that [Alysiella] sp. IAM 14971 (=ATCC 29468) is closely related to the genus Moraxella of the gamma-Proteobacteria (96-97 % similarity). The newly obtained phenotypic data also indicate that [Alysiella] sp. IAM 14971 is distinct from the genus Alysiella and similar to the genus Moraxella. On the basis of these results, the strain should be classified in the genus Moraxella, as Moraxella oblonga sp. nov. The type strain is IAM 14971T (=ATCC 29468T). PMID- 15653897 TI - Photobacterium lipolyticum sp. nov., a bacterium with lipolytic activity isolated from the Yellow Sea in Korea. AB - A Gram-negative, motile, non-spore-forming, pleomorphic and lipolytic bacterial strain, M37T, was isolated from an intertidal sediment of the Yellow Sea in Korea. This organism grew optimally at 25-28 degrees C and in the presence of 1-2 % NaCl. It did not grow without NaCl or in the presence of more than 6 % NaCl. Strain M37T was characterized chemotaxonomically by having Q-8 as the predominant respiratory lipoquinone and C(16 : 1)omega7c and/or iso-C(15 : 0) 2-OH and C(16 : 0) as the major fatty acids. The DNA G+C content was 47 mol%. Phylogenetic analyses based on 16S rRNA gene sequences placed strain M37T within the clade comprising Photobacterium species, forming a coherent cluster with the type strains of Photobacterium profundum and Photobacterium indicum (16S rRNA gene similarity levels of 97.5-98.0 %). The mean DNA-DNA relatedness levels between strain M37T and P. profundum JCM 10084T and P. indicum DSM 5151T were in the range 12-15 %. Similarities between 16S rRNA gene sequences of strain M37T and those of the type strains of the other Photobacterium species ranged from 93.9 % (with Photobacterium fischeri) to 96.2 % (with Photobacterium phosphoreum). On the basis of phenotypic properties and phylogenetic and genomic distinctiveness, strain M37T (=KCTC 10562BPT=DSM 16190T) should be placed in the genus Photobacterium as a novel species, for which the name Photobacterium lipolyticum sp. nov. is proposed. PMID- 15653898 TI - Hydrogenophaga defluvii sp. nov. and Hydrogenophaga atypica sp. nov., isolated from activated sludge. AB - Two Gram-negative, oxidase-positive rods (strains BSB 9.5T and BSB 41.8T) isolated from wastewater were studied using a polyphasic approach. 16S rRNA gene sequence comparisons demonstrated that both strains cluster phylogenetically within the family Comamonadaceae: the two strains shared 99.9 % 16S rRNA gene sequence similarity and were most closely related to the type strains of Hydrogenophaga palleronii (98.5 %) and Hydrogenophaga taeniospiralis (98.0 %). The fatty acid patterns and substrate-utilization profiles displayed similarity to the those of the five Hydrogenophaga species with validly published names, although clear differentiating characteristics were also observed. The two strains showed DNA-DNA hybridization values of 51 % with respect to each other. No close similarities to any other Hydrogenophaga species were detected in hybridization experiments with the genomic DNAs. On the basis of these results, two novel Hydrogenophaga species, Hydrogenophaga defluvii sp. nov. and Hydrogenophaga atypica sp. nov. are proposed, with BSB 9.5T (=DSM 15341T=CIP 108119T) and BSB 41.8T (=DSM 15342T=CIP 108118T) as the respective type strains. PMID- 15653899 TI - Marinilactibacillus piezotolerans sp. nov., a novel marine lactic acid bacterium isolated from deep sub-seafloor sediment of the Nankai Trough. AB - A piezotolerant, mesophilic, marine lactic acid bacterium (strain LT20T) was isolated from a deep sub-seafloor sediment core collected at Nankai Trough, off the coast of Japan. Cells were Gram-positive, rod-shaped, non-sporulating and non motile. The NaCl concentration range for growth was 0-120 g l(-1), with the optimum at 10-20 g l(-1). The temperature range for growth at pH 7.0 was 4-50 degrees C, with the optimum at 37-40 degrees C. The optimum pH for growth was 7.0 8.0. The optimum pressure for growth was 0.1 MPa with tolerance up to 30 MPa. The main cellular phospholipids were phosphatidylglycerols (25 %), diphosphatidylglycerols (34 %) and a group of compounds tentatively identified as ammonium-containing phosphatidylserines (32 %); phosphatidylethanolamines (9 %) were minor components. The fatty acid composition was dominated by side chains of 16 : 0, 14 : 0 and 16 : 1. The G+C content of the genomic DNA was 42 mol%. On the basis of 16S rRNA gene sequence analysis and the secondary structure of the V6 region, this organism was found to belong to the genus Marinilactibacillus and was closely related to Marinilactibacillus psychrotolerans M13-2(T) (99 %), Marinilactibacillus sp. strain MJYP.25.24 (99 %) and Alkalibacterium olivapovliticus strain ww2-SN4C (97 %). Despite the high similarity between their 16S rRNA gene sequences (99 %), the DNA-DNA hybridization levels were less than 20 %. On the basis of physiological and genetic characteristics, it is proposed that this organism be classified as a novel species, Marinilactibacillus piezotolerans sp. nov. The type strain is LT20T (=DSM 16108T=JCM 12337T). PMID- 15653900 TI - Reclassification of Pasteurella gallinarum, [Haemophilus] paragallinarum, Pasteurella avium and Pasteurella volantium as Avibacterium gallinarum gen. nov., comb. nov., Avibacterium paragallinarum comb. nov., Avibacterium avium comb. nov. and Avibacterium volantium comb. nov. AB - This paper describes a phenotypic and genotypic investigation of the taxonomy of [Haemophilus] paragallinarum, Pasteurella gallinarum, Pasteurella avium and Pasteurella volantium, a major subcluster within the avian 16S rRNA cluster 18 of the family Pasteurellaceae. An extended phenotypic characterization was performed of the type strain of [Haemophilus] paragallinarum, which is NAD-dependent, and eight NAD-independent strains of [Haemophilus] paragallinarum. Complete 16S rRNA gene sequences were obtained for one NAD-independent and four NAD-dependent [Haemophilus] paragallinarum strains. These five sequences along with existing 16S rRNA gene sequences for 11 other taxa within avian 16S rRNA cluster 18 as well as seven other taxa from the Pasteurellaceae were subjected to phylogenetic analysis. The analysis demonstrated that [Haemophilus] paragallinarum, Pasteurella gallinarum, Pasteurella avium and Pasteurella volantium formed a monophyletic group with a minimum of 96.8 % sequence similarity. This group can also be separated by phenotypic testing from all other recognized and named taxa within the Pasteurellaceae. As both genotypic and phenotypic testing support the separate and distinct nature of this subcluster, the transfer is proposed of Pasteurella gallinarum, [Haemophilus] paragallinarum, Pasteurella avium and Pasteurella volantium to a new genus Avibacterium as Avibacterium gallinarum gen. nov., comb. nov., Avibacterium paragallinarum comb. nov., Avibacterium avium comb. nov. and Avibacterium volantium comb. nov. The type strains are NCTC 1118T (Avibacterium gallinarum), NCTC 11296T (Avibacterium paragallinarum), NCTC 11297T (Avibacterium avium) and NCTC 3438T (Avibacterium volantium). Key characteristics that separate these four species are catalase activity (absent only in Avibacterium paragallinarum) and production of acid from galactose (negative only in Avibacterium paragallinarum), maltose (negative only in Avibacterium avium) and mannitol (negative in Avibacterium gallinarum and Avibacterium avium). PMID- 15653901 TI - Shewanella decolorationis sp. nov., a dye-decolorizing bacterium isolated from activated sludge of a waste-water treatment plant. AB - A highly efficient dye-decolorizing bacterium, strain S12T, was isolated from activated sludge of a textile-printing waste-water treatment plant in Guangzhou, China. The cells were Gram-negative and motile by means of a single polar flagellum. The strain was capable of anaerobic growth either by fermentation of glucose or by anaerobic respiration and utilized a variety of electron acceptors, including nitrate, iron oxide and thiosulfate. The physiological properties, tested by using the Biolog GN2 system, were similar to those of the genus of Shewanella. Analysis of the nearly complete 16S rRNA gene sequence of strain S12T showed the highest similarity (98 and 97 %, respectively) to Shewanella baltica and Shewanella putrefaciens. However, the level of gyrB similarity between strain S12T and S. putrefaciens was 87 %. DNA from strain S12T showed 41.8 and 41.9 % DNA relatedness, respectively, to the DNA of S. baltica DSM 9439T and S. putrefaciens DSM 6067T. The DNA G+C content of strain S12T was 49.3 mol%. The predominant menaquinone was MK-7 and the predominant ubiquinones were Q-7 and Q 8. The dominant fatty acids were 15 : 0, 16 : 0, iso-15 : 0 and 16 : 1omega7c, similar to the profiles of other Shewanella species. On the basis of its physiological and molecular properties, strain S12T appears to represent a novel species of the genus Shewanella, for which the name Shewanella decolorationis sp. nov. is proposed. The type strain is S12T (=CCTCC M 203093T=IAM 15094T). PMID- 15653902 TI - Sphingopyxis flavimaris sp. nov., isolated from sea water of the Yellow Sea in Korea. AB - A Gram-negative, motile, yellow-pigmented, slightly halophilic bacterial strain, SW-151T, was isolated from sea water of the Yellow Sea in Korea, and subjected to a polyphasic taxonomic study. The isolate grew optimally at 30 degrees C and in the presence of 2-3 % NaCl. Strain SW-151T was characterized chemotaxonomically as having Q-10 as the predominant respiratory lipoquinone and C(18 : 1)omega7c, C(16 : 1)omega7c and/or iso-C(15 : 0) 2-OH and C(17 : 1)omega6c as the major fatty acids. Sphingoglycolipid, diphosphatidylglycerol, phosphatidylglycerol and phosphatidylethanolamine were the major polar lipids. The DNA G+C content was 58 mol%. 16S rRNA gene sequence analysis showed that strain SW-151T joins the evolutionary radiation enclosed by the genus Sphingopyxis. Similarities between the 16S rRNA gene sequences of strain SW-151T and the type strains of Sphingopyxis species ranged from 92.3 to 94.3 %, which is low enough to categorize strain SW-151T as a species distinct from previously described Sphingopyxis species. On the basis of phenotypic properties and phylogenetic distinctiveness, strain SW-151T (=KCTC 12232T=DSM 16223T) should be classified as a novel Sphingopyxis species, for which the name Sphingopyxis flavimaris sp. nov. is proposed. PMID- 15653903 TI - Bizionia paragorgiae gen. nov., sp. nov., a novel member of the family Flavobacteriaceae isolated from the soft coral Paragorgia arborea. AB - A novel marine bacterium isolated from the soft coral Paragorgia arborea in the Sea of Okhotsk was studied by using a polyphasic taxonomic approach. The strain, KMM 6029T, was strictly aerobic, heterotrophic, yellow-pigmented, non-motile by gliding, Gram-negative and oxidase-, catalase- and alkaline phosphatase-positive. From results of 16S rRNA gene sequence analysis, strain KMM 6029T occupied a distinct lineage within the family Flavobacteriaceae and showed 95.5 % similarity to its closest relative, Formosa algae. The DNA G+C content was 37.6 mol%. Major respiratory quinone was MK-6. The predominant fatty acids were i15 : 0, a15 : 0, i15 : 1, a15 : 1, i16 : 1, i16 : 0, i16 : 0 3-OH and summed feature 3 (i15 : 0 2 OH and/or 16 : 1omega7c). On the basis of phenotypic, chemotaxonomic, genotypic and phylogenetic characteristics the novel bacterium has been assigned to Bizionia gen. nov., as Bizionia paragorgiae gen. nov., sp. nov. The type strain is KMM 6029T (=KCTC 12304T=LMG 22571T). PMID- 15653904 TI - Idiomarina seosinensis sp. nov., isolated from hypersaline water of a solar saltern in Korea. AB - A halophilic gamma-proteobacterium, designated CL-SP19T, was isolated from hypersaline water from a solar saltern located in Seosin, Korea. Analysis of the 16S rRNA gene sequence revealed an affiliation with the genus Idiomarina. The sequence similarities between CL-SP19T and type strains of the genus Idiomarina ranged from 95.9 to 96.9 %. Cells were straight or slightly curved rods and were motile by means of a single polar flagellum. The major fatty acids were C(15 : 0) iso (17.1 %) and C(17 : 0) iso (15.2 %). Three fatty acids, C(19 : 0)omega8c cyclo (3.5 %), C(14 : 1)omega5c (1.4 %) and C(18 : 3)omega6c (1.2 %), were found in minor quantities, but uniquely in CL-SP19T among Idiomarina species. The DNA G+C content was 45.0 mol%. On the basis of its physiology, fatty acid composition and 16S rRNA gene sequence, strain CL-SP19T could be assigned to the genus Idiomarina but distinguished from the recognized species of the genus. Strain CL SP19T, therefore, represents a novel species, for which the name Idiomarina seosinensis sp. nov. is proposed, with CL-SP19T (=KCTC 12296T=JCM 12526T) as the type strain. PMID- 15653905 TI - Silanimonas lenta gen. nov., sp. nov., a slightly thermophilic and alkaliphilic gammaproteobacterium isolated from a hot spring. AB - A moderately thermophilic aerobic bacterium, strain 25-4T, was isolated from a hot spring at Baekdoo Mountain in Korea. The cells were Gram-negative, motile rods each having a polar flagellum. Analysis of the 16S rRNA gene sequence indicated that the strain represented a new lineage within the family 'Xanthomonadaceae' of the 'Gammaproteobacteria', being most closely related to the genera Thermomonas, Xanthomonas, Luteimonas, Pseudoxanthomonas, Stenotrophomonas and Xylella and having 16S rRNA gene sequence similarities to the most related species of the genera of between 92.9 and 94.4 %. The strain contained Q-8 as the major isoprenoid quinone and had a fatty acid profile with predominant iso-branched fatty acids. Growth occurred at pH 6.0-10, with an optimum at pH 9.0, and at 25-53 degrees C, with an optimum at 47 degrees C. The G+C content of the genomic DNA was 50.7 mol%. On the basis of phylogenetic analyses and its phenotypic characteristics, strain 25-4T belongs to a new genus, Silanimonas gen. nov., within the 'Gammaproteobacteria'. The sole species of this genus is Silanimonas lenta sp. nov. (type strain, 25-4T=DSM 16282T=KCTC 12236T). PMID- 15653906 TI - Gramella echinicola gen. nov., sp. nov., a novel halophilic bacterium of the family Flavobacteriaceae isolated from the sea urchin Strongylocentrotus intermedius. AB - A novel marine bacterium, strain KMM 6050T, was isolated from the sea urchin Strongylocentrotus intermedius, which inhabits the Sea of Japan. The strain studied was strictly aerobic, heterotrophic, yellow-orange-pigmented, motile by gliding, Gram-negative and oxidase-, catalase-, beta-galactosidase- and alkaline phosphatase-positive. The results of 16S rRNA gene sequence analysis showed that strain KMM 6050T occupies a distinct lineage within the family Flavobacteriaceae and is most closely related to the species Mesonia algae and Salegentibacter salegens (sequence similarity of 92.5-92.6 %). The DNA G+C content of KMM 6050T was 39.6 mol%. The major respiratory quinone was MK-6. The predominant fatty acids were i15 : 0, a15 : 0, 15 : 0, i16 : 1, i16 : 0, i16 : 0 3-OH and i17 : 0 3 OH. On the basis of phenotypic, chemotaxonomic, genotypic and phylogenetic characteristics, the novel bacterium has been assigned to the genus Gramella gen. nov., as Gramella echinicola sp. nov. The type strain is KMM 6050T (=KCTC 12278T=NBRC 100593T=LMG 22585T). PMID- 15653907 TI - Proposal of the genus Thermoactinomyces sensu stricto and three new genera, Laceyella, Thermoflavimicrobium and Seinonella, on the basis of phenotypic, phylogenetic and chemotaxonomic analyses. AB - Phylogenetic analysis based on 16S rRNA gene sequences revealed that Thermoactinomyces species with validly published names can be assigned to four clusters or lineages. The type strains of Thermoactinomyces sacchari and Thermoactinomyces putidus were differentiated from the type strains of Thermoactinomyces vulgaris and Thermoactinomyces intermedius by the predominant menaquinone and fatty acid profiles. The type strains of Thermoactinomyces dichotomicus and Thermoactinomyces peptonophilus formed lines of descent distinct from other Thermoactinomyces species. Thermoactinomyces dichotomicus KCTC 3667T was distinguishable from the type strains of Thermoactinomyces vulgaris and Thermoactinomyces intermedius by the contents of two fatty acids, iso-C(16 : 0) and iso-C(17 : 0). Thermoactinomyces dichotomicus could be distinguished from other Thermoactinomyces species by DNA G+C content and some phenotypic properties, particularly its property of forming a yellow colour. The type strain of Thermoactinomyces peptonophilus was distinguishable from other Thermoactinomyces species by differences in menaquinone profile, major fatty acids, DNA G+C content and some physiological properties including optimal growth temperature. On the basis of these data, the creation of three new genera, Laceyella, Thermoflavimicrobium and Seinonella, is proposed in addition to the genus Thermoactinomyces sensu stricto. The genus Laceyella gen. nov. is proposed to accommodate Thermoactinomyces sacchari and Thermoactinomyces putidus as Laceyella sacchari comb. nov. and Laceyella putida comb. nov., the genus Thermoflavimicrobium gen. nov. is proposed for Thermoactinomyces dichotomicus as Thermoflavimicrobium dichotomicum comb. nov. and the genus Seinonella gen. nov. is proposed for Thermoactinomyces peptonophilus as Seinonella peptonophila comb. nov. PMID- 15653908 TI - Lactobacillus delbrueckii subsp. indicus subsp. nov., isolated from Indian dairy products. AB - Four strains isolated from Indian dairy products and initially identified as Lactobacillus delbrueckii could not be assigned to a definite subspecies because molecular identification and phenotypic traits did not agree with those of recognized subspecies of L. delbrueckii. Hybridization of total DNA (78-86 % against type strains of the other three subspecies), AFLP and RAPD-PCR fingerprints, phylogenetic analysis based on 16S rRNA gene sequences and sequence analysis of two coding genes (recA and hsp60), together with phenotypic profiles, indicated that the four strains form a coherent cluster and represent a novel subspecies, for which the name Lactobacillus delbrueckii subsp. indicus subsp. nov. is proposed. The type strain is NCC725T (=LMG 22083T=DSM 15996T). PMID- 15653909 TI - Paenibacillus xylanilyticus sp. nov., an airborne xylanolytic bacterium. AB - During a search for xylan-degrading micro-organisms, a sporulating bacterium was recovered from xylan-containing agar plates exposed to air in a research laboratory (Salamanca University, Spain). The airborne isolate (designated strain XIL14T) was identified by 16S rRNA gene sequencing as representing a Paenibacillus species most closely related to Paenibacillus illinoisensis JCM 9907T (99.3 % sequence similarity) and Paenibacillus pabuli DSM 3036T (98 % sequence similarity). Phenotypic, chemotaxonomic and DNA-DNA hybridization data indicated that the isolate belongs to a novel species of the genus Paenibacillus. Cells of strain XIL14T were motile, sporulating, rod-shaped, Gram-positive and facultatively anaerobic. The predominant cellular fatty acids were anteiso-C(15 : 0) and C(16 : 0). The DNA G+C content of strain XIL14T was 50.5 mol%. Growth was observed with many carbohydrates, including xylan, as the only carbon source and gas production was not observed from glucose. Catalase was positive and oxidase was negative. The airborne isolate produced a variety of hydrolytic enzymes, including xylanases, amylases, gelatinase and beta-galactosidase. DNA-DNA hybridization levels between strain XIL14T and P. illinoisensis DSM 11733T and P. pabuli DSM 3036T were 43.3 and 36.3 %, respectively. According to the data obtained, strain XIL14T is considered to represent a novel species for which the name Paenibacillus xylanilyticus sp. nov. is proposed (=LMG 21957T=CECT 5839T). PMID- 15653910 TI - Sejongia antarctica gen. nov., sp. nov. and Sejongia jeonii sp. nov., isolated from the Antarctic. AB - Two yellow-pigmented, Gram-negative and aerobic bacterial strains, designated AT1013T and AT1047T, were isolated from terrestrial samples of the Antarctic. On the basis of 16S rRNA gene sequence analyses, the two Antarctic strains shared 97.7 % sequence similarity and showed moderate relationships to the genera Chryseobacterium (92.5-95.3 %), Riemerella (92.3-93.5 %), Bergeyella (92.5-92.6 %) and Kaistella (92.5-93.3 %). In phylogenetic analyses, the two isolates formed a robust monophyletic clade and represented a distinct phyletic line that equated to novel generic status. Cells were non-motile, non-gliding and psychrotolerant with an optimum growth temperature of about 20 degrees C. Flexirubins were absent. The major isoprenoid quinone was MK-6. The predominant cellular fatty acids were 15 : 0 iso, 15 : 0 anteiso and 17 : 1 iso omega9c. DNA G+C contents were 34-36 mol%. The two isolates shared low genomic relatedness (27 %) and were differentiated from each other by several phenotypic characteristics. The polyphasic data presented in this study indicated that these isolates should be recognized as two separate novel species in a novel genus within the family Flavobacteriaceae. The name Sejongia gen. nov. is therefore proposed for the Antarctic isolates, with the type species Sejongia antarctica sp. nov. (type strain AT1013T=IMSNU 14040T=KCTC 12225T=JCM 12381T) and Sejongia jeonii sp. nov. (type strain AT1047T=IMSNU 14049T=KCTC 12226T=JCM 12382T). PMID- 15653911 TI - Micromonospora eburnea sp. nov., isolated from a Thai peat swamp forest. AB - Two actinomycete strains, LK2-10T and LK2-5, which produced single, non-motile spores, were isolated from peat swamp forest soil in Yala Province, Thailand. A polyphasic study was carried out to establish the taxonomic position of these strains. Morphological and chemotaxonomic characteristics of these strains coincided with those of the genus Micromonospora. Phylogenetic analysis using 16S rRNA gene sequences also indicated that these strains should be classified in the genus Micromonospora and clearly separated from their closest relative, Micromonospora nigra DSM 43818T. Furthermore, a combination of DNA-DNA hybridization results and physiological and biochemical properties indicated that these strains were distinguished from all recognized Micromonospora species. These strains therefore represent a novel species, for which the name Micromonospora eburnea sp. nov. is proposed. The type strain is LK2-10T (=JCM 12345T=PCU 238T=DSM 44814T=TISTR 1531T). PMID- 15653912 TI - Chryseobacterium formosense sp. nov., isolated from the rhizosphere of Lactuca sativa L. (garden lettuce). AB - A yellow-pigmented bacterial strain (CC-H3-2T), isolated from the rhizosphere of Lactuca sativa L. (garden lettuce) in Taiwan, was investigated using a polyphasic taxonomic approach. The cells were Gram-negative, rod-shaped and non-spore forming. Phylogenetic analyses using the 16S rRNA gene sequence of the isolate indicated that the organism belongs to the genus Chryseobacterium, with the highest sequence similarity to the type strains of Chryseobacterium indoltheticum (97.7 %), Chryseobacterium scophthalmum (97.5 %), Chryseobacterium joostei (97.2 %) and Chryseobacterium defluvii (97.2 %). The major whole-cell fatty acids were iso-C(15 : 0) (52.2 %) and iso-C(17 : 0) 3-OH. DNA-DNA hybridization experiments revealed levels of only 27.4 % to C. scophthalmum, 27.1 % to C. indoltheticum, 14.1 % to C. joostei and 7.8 % to C. defluvii. DNA-DNA relatedness and biochemical and chemotaxonomic properties demonstrate that strain CC-H3-2T represents a novel species, for which the name Chryseobacterium formosense sp. nov. is proposed. The type strain is CC-H3-2T (=CCUG 49271T=CIP 108367T). PMID- 15653913 TI - Actinomyces dentalis sp. nov., from a human dental abscess. AB - A previously undescribed filamentous, beaded, Gram-positive, rod-shaped bacterium was isolated from pus of a human dental abscess. Based on its cellular morphology and the results of biochemical testing the organism was tentatively identified as a member of the genus Actinomyces, but it did not correspond to any currently recognized species of this genus. Comparative 16S rRNA gene sequencing studies showed the bacterium represents a distinct subline within the genus Actinomyces, clustering within a group of species that includes Actinomyces bovis, the type species of the genus. Sequence divergence values of >8 % with other recognized species within this phylogenetic group clearly demonstrated that the organism represents a hitherto unknown species. Based on biochemical and molecular phylogenetic evidence, it is proposed that the unidentified organism recovered from a dental abscess be classified as a novel species, Actinomyces dentalis sp. nov. The type strain is R18165T (=CCUG 48064T=CIP 108337T). PMID- 15653914 TI - Nocardia takedensis sp. nov., isolated from moat sediment and scumming activated sludge. AB - Chemotaxonomic and morphological characterization of two actinomycete strains, MS1-3T and AS4-2, respectively isolated from moat sediment and scumming activated sludge, was carried out. This characterization clearly demonstrated that strains MS1-3T and AS4-2 belong to the genus Nocardia. 16S rRNA gene sequencing studies showed that these isolates are most closely related to Nocardia beijingensis (98.1-98.3 % similarity), Nocardia brasiliensis (97.9-98.0 %) and Nocardia tenerifensis (97.8-97.9 %). However, the results of DNA-DNA hybridizations and physiological and biochemical tests showed that strains MS1-3T and AS4-2 could be differentiated from their closest phylogenetic relatives both genotypically and phenotypically. It is proposed that the two isolates be classified as representatives of a novel species of Nocardia, Nocardia takedensis sp. nov. The type strain is MS1-3T (=NBRC 100417T=DSM 44801T); AS4-2 (=NBRC 100418=DSM 44802) is a reference strain. PMID- 15653915 TI - Phylogenetic relationships of the genus Kluyvera: transfer of Enterobacter intermedius Izard et al. 1980 to the genus Kluyvera as Kluyvera intermedia comb. nov. and reclassification of Kluyvera cochleae as a later synonym of K. intermedia. AB - In order to assess the relationship between the genus Kluyvera and other members of the family Enterobacteriaceae, the 16S rRNA genes of type strains of the recognized Kluyvera species, Kluyvera georgiana, Kluyvera cochleae, Kluyvera ascorbata and Kluyvera cryocrescens, were sequenced. A comparative phylogenetic analysis based on these 16S rRNA gene sequences and those available for strains belonging to several genera of the family Enterobacteriaceae showed that members of the genus Kluyvera form a cluster that contains all the known Kluyvera species. However, the type strain of Enterobacter intermedius (ATCC 33110T) was included within this cluster in a very close relationship with the type strain of K. cochleae (ATCC 51609T). In addition to the phylogenetic evidence, biochemical and DNA-DNA hybridization analyses of species within this cluster indicated that the type strain of E. intermedius is in fact a member of the genus Kluyvera and, within it, of the species Kluyvera cochleae. Therefore, following the current rules for bacterial nomenclature and classification, the transfer of E. intermedius to the genus Kluyvera as Kluyvera intermedia comb. nov. is proposed (type strain, ATCC 33110T=CIP 79.27T=LMG 2785T=CCUG 14183T). Biochemical analysis of four E. intermedius strains and one K. cochleae strain independent of the respective type strains further indicated that E. intermedius and K. cochleae represent the same species and are therefore heterotypic synonyms. Nomenclatural priority goes to the oldest legitimate epithet. Consequently, Kluyvera cochleae Muller et al. 1996 is a later synonym of Kluyvera intermedia (Izard et al. 1980) Pavan et al. 2005. PMID- 15653916 TI - Bacillus patagoniensis sp. nov., a novel alkalitolerant bacterium from the rhizosphere of Atriplex lampa in Patagonia, Argentina. AB - A Gram-positive, rod-shaped, spore-forming bacterium (PAT 05T) was isolated from the rhizosphere of the perennial shrub Atriplex lampa in north-eastern Patagonia, Argentina. Its overall biochemical and physiological characteristics indicated that this strain should be placed in the alkaliphilic Bacillus group. Strain PAT 05T grew at pH 7-10 (optimum pH 8), but not at pH 6. Its DNA G+C content was 39.7 mol%. Sequence analysis of the 16S rRNA gene of PAT 05T revealed the closest match (99.6 % similarity) with Bacillus sp. DSM 8714. The highest level of DNA DNA relatedness (88.6 %) was also found with this strain. On the basis of 16S rRNA gene sequence similarity and phylogenetic analysis, G+C content and DNA-DNA hybridization data, strain PAT 05T is related at the species level to Bacillus sp. DSM 8714, a member of a group referred as phenon 4a by Nielsen et al. [Nielsen, P., Fritze, D. & Priest, F. G. (1995). Microbiology 141, 1745-1761], which still lacks taxonomic standing. These results support the proposal of strain PAT 05T (=DSM 16117T=ATCC BAA-965T) as the type strain of Bacillus patagoniensis sp. nov. PMID- 15653917 TI - Multilocus sequence phylogenetic study of the genus Haemophilus with description of Haemophilus pittmaniae sp. nov. AB - The phylogeny of human isolates of Haemophilus species was estimated based on partial sequences of four separate housekeeping genes. The clustering of each set of sequences was in accordance with speciation of the strains with few exceptions: of 108 gene fragments examined, only three appeared to have been subject to recombination events across the species barrier. Housekeeping gene similarity supported previous DNA-DNA hybridization data for the genus rather than the phylogeny inferred from 16S rRNA gene sequence comparison. The similarity of sequences of Haemophilus parainfluenzae with those of Haemophilus influenzae suggested preservation of the former species in the genus Haemophilus. Three strains representing a novel taxon were unique with respect to the four investigated gene loci. 16S rRNA gene sequence analysis suggested that this taxon belonged to the Parainfluenzae cluster. DNA-DNA hybridization data supported this generic placement. Nine strains of the novel taxon were available for analysis. They were distinct from representatives of all current species of the genus Haemophilus by conventional phenotypic characterization. Genotypic and phenotypic data show that the strains merit recognition as a novel species of Haemophilus. The name Haemophilus pittmaniae sp. nov. is proposed, with HK 85T (=CCUG 48703T=NCTC 13334T) as the type strain. PMID- 15653918 TI - Arthrobacter bergerei sp. nov. and Arthrobacter arilaitensis sp. nov., novel coryneform species isolated from the surfaces of cheeses. AB - Fourteen isolates of two different bacterial species isolated from the surface of smear-ripened cheeses were found to exhibit many characteristics of the genus Arthrobacter. The isolates were aerobic, Gram-positive, catalase-positive, non spore-forming and non-motile. The cell-wall peptidoglycan contained lysine, alanine and glutamic acid. rrs sequence analysis indicated that the new isolates Re117T and Ca106T are closely related to the Arthrobacter nicotianae group and showed highest sequence similarity (>98 %) to Arthrobacter nicotianae and Arthrobacter protophormiae. However, DNA-DNA hybridization studies indicated that the strains represented two novel genomic species within the genus Arthrobacter and did not belong to A. nicotianae or A. protophormiae (<43 % DNA-DNA relatedness). On the basis of the phylogenetic and phenotypic distinctiveness of the new isolates, these bacteria should be classified as two novel Arthrobacter species, for which the names Arthrobacter bergerei sp. nov. and Arthrobacter arilaitensis sp. nov. are proposed. Type strains have been deposited in culture collections as Arthrobacter bergerei Ca106T (=CIP 108036T=DSM 16367T) and Arthrobacter arilaitensis Re117T (=CIP 108037T=DSM 16368T). PMID- 15653919 TI - Nesterenkonia sandarakina sp. nov. and Nesterenkonia lutea sp. nov., novel actinobacteria, and emended description of the genus Nesterenkonia. AB - Two novel actinobacteria isolates, designated YIM 70009T and YIM 70081T, were characterized in order to determine their taxonomic position. Cells of strains YIM 70009T and YIM 70081T were cocci, although only the latter were motile. The G+C contents of their DNAs were 64.0 and 64.5 mol%, respectively. On the basis of chemotaxonomic characteristics and 16S rRNA gene sequence analysis, the two isolates were classified in the genus Nesterenkonia. DNA-DNA hybridization and comparison of phenotypic characteristics revealed that strains YIM 70009T and YIM 70081T differed from each other and from known species. Therefore, it is proposed that they represent two separate novel species of the genus Nesterenkonia: Nesterenkonia sandarakina sp. nov. (type strain, YIM 70009T=CCTCC AA 203007T=DSM 15664T=KCTC 19011T) and Nesterenkonia lutea sp. nov. (type strain, YIM 70081T=CCTCC AA 203010T=DSM 15666T=KCTC 19013T). PMID- 15653920 TI - Thiobacter subterraneus gen. nov., sp. nov., an obligately chemolithoautotrophic, thermophilic, sulfur-oxidizing bacterium from a subsurface hot aquifer. AB - A novel, thermophilic, obligately chemolithoautotrophic, sulfur/thiosulfate oxidizing bacterium was isolated from subsurface geothermal aquifer water (temperature approximately 70 degrees C) in the Hishikari gold mine, Japan. Cells of the isolate, designated strain C55T, were motile, straight rods with a single polar flagellum. Growth was observed at temperatures between 35 and 62 degrees C (optimum 50-55 degrees C; 60 min doubling time) and pH between 5.2 and 7.7 (optimum pH 6.5-7.0). High growth rate of strain C55T was observed on either thiosulfate or elemental sulfur as a sole energy source, with molecular oxygen as the only electron acceptor. None of the organic compounds tested supported or stimulated growth of strain C55T. The G+C content of the genomic DNA was 66.9 mol%. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain C55T was affiliated to the beta-Proteobacteria, but was distantly related to recognized genera. On the basis of its physiological and molecular properties, strain C55T (=JCM12421T=DSM 16629T=ATCC BAA-941T) is proposed as the type strain of Thiobacter subterraneus gen. nov., sp. nov. PMID- 15653921 TI - Carnobacterium pleistocenium sp. nov., a novel psychrotolerant, facultative anaerobe isolated from permafrost of the Fox Tunnel in Alaska. AB - A novel, psychrotolerant, facultative anaerobe, strain FTR1T, was isolated from Pleistocene ice from the permafrost tunnel in Fox, Alaska. Gram-positive, motile, rod-shaped cells were observed with sizes 0.6-0.7 x 0.9-1.5 microm. Growth occurred within the pH range 6.5-9.5 with optimum growth at pH 7.3-7.5. The temperature range for growth of the novel isolate was 0-28 degrees C and optimum growth occurred at 24 degrees C. The novel isolate does not require NaCl; growth was observed between 0 and 5 % NaCl with optimum growth at 0.5 % (w/v). The novel isolate was a catalase-negative chemoorganoheterotroph that used as substrates sugars and some products of proteolysis. The metabolic end products were acetate, ethanol and CO2. Strain FTR1T was sensitive to ampicillin, tetracycline, chloramphenicol, rifampicin, kanamycin and gentamicin. 16S rRNA gene sequence analysis showed 99.8 % similarity between strain FTR1T and Carnobacterium alterfunditum, but DNA-DNA hybridization between them demonstrated 39+/-1.5 % relatedness. On the basis of genotypic and phenotypic characteristics, it is proposed that strain FTR1T (=ATCC BAA-754T=JCM 12174T=CIP 108033T) be assigned to the novel species Carnobacterium pleistocenium sp. nov. PMID- 15653922 TI - Brevundimonas mediterranea sp. nov., a non-stalked species from the Mediterranean Sea. AB - Six strains of Gram-negative, rod-shaped, non-spore-forming bacteria were isolated from the Mediterranean Sea. 16S rRNA gene sequence analysis indicated that the strains were affiliated within the alphaproteobacterial genus Brevundimonas, with Brevundimonas intermedia (99.4 %) and Brevundimonas vesicularis (99.2 %) as their closest relatives. This affiliation was supported by chemotaxonomic data (major polar lipids: phosphatidyl diacylglycerol, sulfoquinovosyl diacylglycerol and phosphatidyl glucopyranosyl diacylglycerol; major fatty acids: C(18 : 1), C(16 : 0), C(16 : 1), C(15 : 0), C(17 : 1)omega8c, 11-Me-C(18 : 1)omega5t). The results of DNA-DNA hybridization and physiological and biochemical tests allowed genotypic and phenotypic differentiation of the strains from all recognized Brevundimonas species. The strains therefore represent a novel species, for which the name Brevundimonas mediterranea sp. nov. is proposed, with the type strain V4.BO.10T)(=LMG 21911T=CIP 107934T). PMID- 15653925 TI - Strain NBRC (formerly IFO) 3782 is the type strain of Streptomyces rameus Shibata 1959. Opinion 76. AB - The Judicial Commission of the International Committee for Systematics of Prokaryotes decided that strain NBRC (formerly IFO) 3782 (=No. 43797), which was the originally designated type strain, has to replace ATCC 21273 as the type strain of Streptomyces rameus. ATCC 21273 was given as the type strain in the Approved Lists 1980. PMID- 15653924 TI - Sporidiobolus longiusculus sp. nov. and Sporobolomyces patagonicus sp. nov., novel yeasts of the Sporidiobolales isolated from aquatic environments in Patagonia, Argentina. AB - During a survey of carotenogenic yeasts carried out in north-western Patagonia (Argentina), several ballistoconidia-producing strains belonging to the order Sporidiobolales were isolated from aquatic environments. Five strains were found to represent two novel species, for which the names Sporidiobolus longiusculus and Sporobolomyces patagonicus are proposed, with CBS 9654T (=PYCC 5818T=CRUB 1044T) and CBS 9657T (=PYCC 5817T=CRUB 1038T) as the type strains, respectively. The elongated basidia, which are five to six times longer that those of the remaining species of the genus Sporidiobolus, are a particular micromorphological feature of Sporidiobolus longiusculus. On the basis of the sequences of the D1/D2 domains of the 26S rRNA gene, the species most closely related to Sporidiobolus longiusculus is Sporobolomyces bannaensis, whereas Sporobolomyces marcillae is the closest relative of Sporobolomyces patagonicus. Complete internal transcribed spacer sequence analysis confirmed the separate position of Sporidiobolus longiusculus, whereas for Sporobolomyces patagonicus no nucleotide differences were found with respect to Sporidiobolus pararoseus CBS 491T. Negative mating experiments between strains of Sporobolomyces patagonicus and strains of Sporidiobolus pararoseus together with the low DNA-DNA reassociation values for the type strains of the two species validated the proposal of Sporobolomyces patagonicus as a distinct species. Information on additional Patagonian Sporobolomyces isolates is also included in this report. PMID- 15653923 TI - On the monophyly of chromalveolates using a six-protein phylogeny of eukaryotes. AB - A global phylogeny of major eukaryotic lineages is a significant and ongoing challenge to molecular phylogenetics. Currently, there are five hypothesized major lineages or 'supergroups' of eukaryotes. One of these, the chromalveolates, represents a large fraction of protist and algal diversity. The chromalveolate hypothesis was originally based on similarities between the photosynthetic organelles (plastids) found in many of its members and has been supported by analyses of plastid-related genes. However, since plastids can move between eukaryotic lineages, it is important to provide additional support from data generated from the nuclear-cytosolic host lineage. Genes coding for six different cytosolic proteins from a variety of chromalveolates (yielding 68 new gene sequences) have been characterized so that multiple gene analyses, including all six major lineages of chromalveolates, could be compared and concatenated with data representing all five hypothesized supergroups. Overall support for much of the phylogenies is decreased over previous analyses that concatenated fewer genes for fewer taxa. Nevertheless, four of the six chromalveolate lineages (apicomplexans, ciliates, dinoflagellates and heterokonts) consistently form a monophyletic assemblage, whereas the remaining two (cryptomonads and haptophytes) form a weakly supported group. Whereas these results are consistent with the monophyly of chromalveolates inferred from plastid data, testing this hypothesis is going to require a substantial increase in data from a wide variety of organisms. PMID- 15653926 TI - The type species of the genus Paenibacillus Ash et al. 1994 is Paenibacillus polymyxa. Opinion 77. AB - The Judicial Commission of the International Committee for Systematics of Prokaryotes decided that the type species of the genus Paenibacillus is Paenibacillus polymyxa. PMID- 15653927 TI - Rejection of the genus name Pelczaria with the species Pelczaria aurantia Poston 1994. Opinion 78. AB - The Judicial Commission of the International Committee for Systematics of Prokaryotes has decided to place the genus Pelczaria with the species Pelczaria aurantia on the list of nomina rejicienda, due to the lack of an authentic type or neotype strain. PMID- 15653928 TI - The nomenclatural types of the orders Acholeplasmatales, Halanaerobiales, Halobacteriales, Methanobacteriales, Methanococcales, Methanomicrobiales, Planctomycetales, Prochlorales, Sulfolobales, Thermococcales, Thermoproteales and Verrucomicrobiales are the genera Acholeplasma, Halanaerobium, Halobacterium, Methanobacterium, Methanococcus, Methanomicrobium, Planctomyces, Prochloron, Sulfolobus, Thermococcus, Thermoproteus and Verrucomicrobium, respectively. Opinion 79. AB - The Judicial Commission of the International Committee on Systematics of Prokaryotes has corrected the nomenclatural types of 12 orders: Acholeplasmatales, Halanaerobiales, Halobacteriales, Methanobacteriales, Methanococcales, Methanomicrobiales, Planctomycetales, Prochlorales, Sulfolobales, Thermococcales, Thermoproteales and Verrucomicrobiales. PMID- 15653929 TI - The type species of the genus Salmonella Lignieres 1900 is Salmonella enterica (ex Kauffmann and Edwards 1952) Le Minor and Popoff 1987, with the type strain LT2T, and conservation of the epithet enterica in Salmonella enterica over all earlier epithets that may be applied to this species. Opinion 80. AB - The Judicial Commission of the International Committee for Systematics of Prokaryotes has decided that the type species of the genus Salmonella Lignieres 1900 is Salmonella enterica (ex Kauffmann and Edwards 1952) Le Minor and Popoff 1987 and that the type strain of this species is strain LT2T. In addition, the epithet enterica in Salmonella enterica is conserved over all earlier epithets that may be applied to this species. PMID- 15653930 TI - Nomenclature and taxonomy of the genus Salmonella. AB - The nomenclature of the genus Salmonella has reached an unsatisfactory state of affairs, with two systems of nomenclature in circulation. One system, proposed in the 1980s by Le Minor and Popoff, has received wide acceptance, although it does not conform to the rules of the Bacteriological Code. The other system, which conforms to the rules of the Bacteriological Code, is being used by an ever decreasing minority. As a result of a number of recent Requests for an Opinion, the Judicial Commission of the International Committee on the Systematics of Prokaryotes has issued an Opinion (Opinion 80) with the intention that it should solve these discrepancies. However, like all Opinions, it is limited to matters of nomenclature and does not help to interpret the taxonomic consequences. The Judicial Commission has therefore asked experts in the field of nomenclature and taxonomy to write a commentary on the nomenclatural and taxonomic consequences of Opinion 80. The present article explains the nomenclatural consequences of Opinion 80, together with a clear presentation of the taxonomy that results when applying the currently widely accepted interpretation that the genus Salmonella currently includes only two species. PMID- 15653931 TI - Induction of human airway smooth muscle apoptosis by neutrophils and neutrophil elastase. AB - Neutrophils are an important component of airway inflammation and may interact with human airway smooth muscle cells (HASMC). We investigated the effect of neutrophils and of neutrophil-derived proteases on HASMC survival. When co incubated with neutrophils (0.1-1 x 10(6) cells/ml), attachment of human ASMC was reduced to 12.3 +/- 4.3% compared with untreated controls after 72 h. HASMC showed nuclear condensation and fragmentation (41.6 +/- 8.1% compared with baseline of 3.1 +/- 0.4%), and the biochemical markers of apoptosis, annexin V binding (9.7 +/- 0.7%; baseline 1.1 +/- 0.3%) and cleaved caspase-3 expression, were observed. The proteolytic activity released by neutrophils was essential for the proapoptotic effect because inhibition of elastase activity by alpha(1) antitrypsin and MeOSuc-Ala-Ala-Pro-Ala-CMK (MSACK) reduced HASMC apoptosis. Human neutrophil elastase (0.1-3 microg/ml) induced apoptosis of HASMC, as well as other neutrophil serine proteases, cathepsin G, and proteinase 3. Fibronectin degradation products were present in HASMC supernatants exposed to neutrophil conditioned media and to neutrophil elastase. The local release of proteases from neutrophils present in airway smooth muscle cells may lead to HASMC apoptosis as a result of matrix degradation and loss of cell attachment. This may limit pathologic changes such as ASMC hyperplasia and extracellular matrix deposition seen in airway remodeling. PMID- 15653932 TI - Tumor necrosis factor-alpha induces transforming growth factor-beta1 expression in lung fibroblasts through the extracellular signal-regulated kinase pathway. AB - Increased expression of transforming growth factor (TGF)-beta(1) and tumor necrosis factor (TNF)-alpha are thought to play important roles in the development of pulmonary fibrosis. We recently reported that TNF-alpha upregulates TGF-beta(1) expression in primary mouse lung fibroblasts (MLFs), a key cell population in fibrogenesis. In the present study, we have investigated signal transduction pathways involved in TNF-alpha upregulation of TGF-beta(1) in both primary MLFs and the Swiss 3T3 fibroblast cell line. Treatment of fibroblasts with TNF-alpha resulted in a significant increase in TGF-beta(1) protein as measured by ELISA. The increase in protein was preceded by a 200-400% increase in TGF-beta(1) mRNA detected by quantitative, real-time, reverse transcriptase-polymerase chain reaction. Western blot analysis showed that TNF alpha activated the extracellular signal-regulated kinase (ERK), and inhibitors of the ERK-specific mitogen-activated protein kinase pathway (PD98059 or U0126) blocked TNF-alpha induction of TGF-beta(1) mRNA and protein. mRNA stability experiments showed that TNF-alpha increased the half-life of TGF-beta(1) mRNA to more than 24 h compared with approximately 15 h in unstimulated cells. Expression of constitutively active MEK1 that selectively phosphorylates ERK was sufficient for TGF-beta(1) mRNA stabilization in Swiss 3T3 fibroblasts. These results indicate that TNF-alpha activates the ERK-specific mitogen-activated protein kinase pathway leading to increased TGF-beta(1) production in fibroblasts, primarily via a post-transcriptional mechanism that involves stabilization of the TGF-beta(1) transcript. PMID- 15653933 TI - Another milestone: the journal goes monthly. PMID- 15653934 TI - Anxiety and its disorders in late life: moving the field forward. PMID- 15653935 TI - Affective correlates of fear of falling in elderly persons. AB - OBJECTIVE: Fear of falling is common in older people, occurring on average in 50% of those who have fallen in the previous year. Little is known about the psychological correlates of fear of falling. The purpose of this study was to determine whether clinically significant depression and anxiety were independently associated with fear of falling. METHODS: This was a cross sectional study of 105 persons age > or =60 years, admitted to medical or orthopedic wards, who had fallen at least once in the previous 12 months. Fear of falling was assessed using two different constructs: 1) intensity of fear; and 2) self-efficacy. Depressive and anxiety disorders were assessed with the Structured Clinical Interview for DSM-IV. Depression and anxiety severity were assessed with the Hospital Anxiety and Depression Scale. Demographic, physical, functional, and social variables previously found to be associated with fear of falling were also measured. Logistic-regression and multiple-regression analyses were used to examine the independent association of affective variables with fear of falling. RESULTS: Depressive disorders, anxiety disorders, depression severity, and anxiety severity had significant independent associations with both constructs of fear of falling. Of all the variables that were measured, depressive disorders and depression severity had the strongest associations with fear of falling. CONCLUSION: Affective variables had a stronger association with fear of falling than non-affective variables in a hospital-based group of subjects. Further research is needed to determine whether similar findings occur in a community based sample of older people. PMID- 15653936 TI - Correlates of anxiety symptoms in physically disabled older women. AB - OBJECTIVE: The authors describe characteristics that are associated with chronic anxiety symptoms and examine the use of anxiolytic and antidepressant medications in physically disabled women with and without symptoms of anxiety. METHODS: Participants were 791 physically disabled women age 65+ years who participated in the Women's Health and Aging Study for 2-3 years. Anxiety symptoms were measured with four questions from the Hopkins Symptom Checklist, and women were categorized as having no anxiety, intermittent anxiety, and chronic anxiety symptoms. Health-related characteristics, medications, physical functioning, physical activity, and psychosocial variables were also measured. RESULTS: Forty nine percent of women reported no anxiety symptoms; 41% reported intermittent symptoms; and 10% reported chronic symptoms of anxiety. Depressive symptoms and lack of emotional support were significant correlates of intermittent anxiety symptoms, whereas depressive symptoms, negative life events, and lack of emotional support were significant correlates of chronic anxiety symptoms. Over the course of 3 years, 20.3% of women with no anxiety, 33.0% of women with intermittent anxiety, and 48.7% of women with chronic anxiety symptoms took anxiolytic and/or antidepressant medications. CONCLUSION: Anxiety symptoms are common among disabled older women. Psychosocial variables were significantly different in women with intermittent or chronic anxiety symptoms, versus women without anxiety. PMID- 15653937 TI - Age at onset of generalized anxiety disorder in older adults. AB - OBJECTIVE: The authors explored the distribution and correlates of age-at-onset of late-life generalized anxiety disorder (GAD). METHODS: Authors examined the distribution of age at onset in a sample of 67 older adults with GAD recruited for a psychotherapy study. They compared those with an early onset of symptoms (before age 50) to those with a late onset (after 50) on demographic variables and measures of psychopathology and health-related quality of life. RESULTS: There was a bimodal distribution of age at onset, with 57% reporting early onset and 43% reporting a late onset. Patients with an early onset of symptoms had a higher rate of psychiatric comorbidity and psychotropic medication use and more severe worry. Patients with a late onset of symptoms reported more functional limitations due to physical problems. CONCLUSIONS: Although most older GAD patients report an onset in childhood or adolescence, almost half develop the disorder in late life. Older adults with an early onset of GAD appear to have a more severe course, characterized by pathological worry, than those with a later onset. Role disability may be a risk factor for onset of GAD in late life. PMID- 15653938 TI - Depression and generalized anxiety disorder: co-occurrence and longitudinal patterns in elderly patients. AB - OBJECTIVE: The authors sought to establish the natural course and risk-profile of depression, generalized anxiety disorder (GAD), and depression with co-existing GAD in later life. METHODS: A total of 2,173 community-living elderly persons were interviewed at baseline, and at a 3-year follow-up. The course of "pure" depression, "pure" GAD, and depression with coexisting GAD was studied in 258 subjects with baseline psychopathology. Authors assessed bivariate and multivariate relationships between risk factors and course types. The risk profile for onset of pure depression, pure GAD, and the mixed condition at follow up was studied in 1,915 subjects without baseline psychopathology. RESULTS: Remission rate at follow-up was 41% for subjects with depression-only, 48% for pure GAD, and significantly lower (27%) for depression with coexisting GAD. A pattern of temporal sequencing was established, with anxiety often progressing to depression or depression with GAD. Onset of pure depression and depression with co-existing GAD was predicted by loss events, ill health, and functional disability. Onset of pure GAD, and, more strongly, that of depression with coexisting GAD, was associated with longstanding, possibly genetic vulnerability. CONCLUSIONS: In comparison with either depression-only or anxiety-only, the co occurrence of these represents more severe and more chronic psychopathology, associated with longstanding vulnerability. In elderly persons, GAD often progresses to depression or to the mixed condition. These findings mostly favor a dimensional, rather than a categorical, classification of anxiety and depression. PMID- 15653939 TI - Impact of symptoms of generalized anxiety disorder on the course of late-life depression. AB - OBJECTIVE: The authors sought to examine the effect of symptoms of generalized anxiety disorder (GAD) on acute course of depression in a group of older depressed patients. METHODS: Authors assessed GAD symptoms in a sample of 204 patients age 60 and older with unipolar major depression. Patients were treated naturalistically by study geriatric psychiatrists using a treatment guideline for geriatric depression and assessed with the Montgomery-Asberg Depression Rating Scale. They were dichotomized as meeting or not meeting GAD symptom criteria. The groups were analyzed by Cox proportional-hazards models. RESULTS: There were 138 remitters and 66 non-remitters. After analysis controlled for benzodiazepine use, stressful life events, social support, and functional status, having GAD symptoms was associated with longer time-to-remission. CONCLUSIONS: Older depressed patients with GAD have a worse outcome than those without anxiety. Future studies will need to determine the appropriate role of benzodiazepines and other anxiolytics in the treatment of older depressed patients with symptoms of GAD. PMID- 15653940 TI - Impact of comorbid panic and posttraumatic stress disorder on outcomes of collaborative care for late-life depression in primary care. AB - OBJECTIVE: Comorbid anxiety disorders may result in worse depression treatment outcomes. The authors evaluated the effect of comorbid panic disorder and posttraumatic stress disorder (PTSD) on response to a collaborative-care intervention for late-life depression in primary care. METHODS: A total of 1,801 older adults with depression were randomized to a collaborative-care depression treatment model versus usual care and assessed at baseline, 3, 6, and 12 months, comparing differences among participants with comorbid panic disorder (N=262) and PTSD (N=191) and those without such comorbid anxiety disorders. RESULTS: At baseline, patients with comorbid anxiety reported higher levels of psychiatric and medical illness, greater functional impairment, and lower quality of life. Participants without comorbid anxiety who received collaborative care had early and lasting improvements in depression compared with those in usual care. Participants with comorbid panic disorder showed similar outcomes, whereas those with comorbid PTSD showed a more delayed response, requiring 12 months of intervention to show a significant effect. At 12 months, however, outcomes were comparable. Interactions of intervention status by comorbid PTSD or panic disorder were not statistically significant, suggesting that the collaborative care model performed significantly better than usual care in depressed older adults both with and without comorbid anxiety. CONCLUSIONS: Collaborative care is more effective than usual care for depressed older adults with and without comorbid panic disorder and PTSD, although a sustained treatment response was slower to emerge for participants with PTSD. Intensive and prolonged follow-up may be needed for depressed older adults with comorbid PTSD. PMID- 15653941 TI - Randomized, double-blind, placebo-controlled trial of fluoxetine treatment for elderly patients with dysthymic disorder. AB - OBJECTIVE: The authors compared the efficacy and side effects of fluoxetine and placebo in elderly outpatients with dysthymic disorder. METHODS: Patients were randomly assigned to fluoxetine (20 mg-60 mg/day) or placebo for 12 weeks in a double-blind trial. RESULTS: Of 90 randomized patients, 71 completed the trial. In the intent-to-treat sample, random regression analyses of the Hamilton Rating Scale for Depression (Ham-D; 24-item) and Cornell Dysthymia Rating Scale (CDRS) scores at each visit produced significant time x treatment group interactions favoring the fluoxetine group. Analysis of percentage change in Ham-D scores yielded no effect for treatment group, but a similar analysis of percentage change in CDRS scores yielded a main effect for treatment group, favoring fluoxetine over placebo. In the intent-to-treat sample, response rates were 27.3% for fluoxetine and 19.6% for placebo. In the completer sample, response rates were 37.5% for fluoxetine and 23.1% for placebo. CONCLUSION: Fluoxetine had limited efficacy in elderly dysthymic patients. The clinical features of elderly dysthymic patients are typically distinct from those of dysthymic disorder in young adults, and the findings suggest that treatments effective for young adult dysthymic patients may not be as useful in elderly dysthymic patients. Further research is needed to identify efficacious treatments for elderly patients with dysthymic disorder, and investigative tools such as electronic/computerized brain scans and neuropsychological testing may help identify the factors that moderate antidepressant treatment response and resistance. PMID- 15653942 TI - Gambling among older, primary-care patients: an important public health concern. AB - OBJECTIVES: Authors assessed the prevalence rate of gambling in a sample of elderly patients (over age 65) and evaluated the sociodemographic characteristics, health, cognitive status, and psychiatric comorbidities of elderly gamblers. METHODS: This study is a cross-sectional survey of gambling habits. A random sample of older adults with a scheduled primary-care clinic appointment was selected and screened with the General Health Questionnaire and questions about suicidality and alcohol use. Also, participants were asked about their gambling habits. RESULTS: Of 843 screened patients completing the gambling questionnaire, 69.6% reported that they had participated in at least one gambling activity in the last year. At-risk gamblers were defined as those who reported having bet more than $100 on a single bet and/or having bet more than they could afford to lose in the last year. Of those responding, 10.9% were identified as at risk gamblers. The strongest predictors of at-risk gambling behavior were being a binge drinker, presence of current posttraumatic stress disorder symptoms, minority race/ethnicity, and being a VA clinic patient. Subjects with mild-to moderate cognitive impairment were just as likely as those without impairment to gamble and to report at-risk gambling behavior. At-risk gambling behavior was not significantly associated with gender, current or past depressive symptoms, or cigarette smoking. CONCLUSIONS: These data raise a significant public health concern that gambling behaviors are prevalent in older adults. Additional research is needed to further define the determinants of gambling behavior in older adults and evaluate the social, health, and economic costs and benefits of gambling by older adults, especially among those groups determined to be at risk. PMID- 15653943 TI - Generalized anxiety disorder in late life: lifetime course and comorbidity with major depressive disorder. AB - OBJECTIVE: Generalized anxiety disorder (GAD) in elderly persons is highly prevalent, but little is known about its course, age at onset, and relationship to comorbid major depressive disorder (MDD). The authors assessed the course and comorbidity of late-life GAD and MDD. METHODS: Authors assessed elderly subjects in anxiety or depression intervention studies who had a lifetime history of GAD, with current MDD (N=57) or without (N=46). Subjects' lifetime course of illness was charted retrospectively. RESULTS: The 103 subjects had a mean age of 74.1 years, and a mean age at onset of GAD of 48.8 years; 46% were late-onset. GAD episodes were chronic, and 36% were longer than 10 years. Of the comorbid GAD-MDD patients, most had different times of onset and/or offset of the disorders; typically, GAD preceded MDD. CONCLUSIONS: Elderly subjects with GAD tended to have chronic symptoms lasting years-to-decades, without interruption, and many have late onset. Elderly persons with lifetime GAD and MDD tend to have different onset and offset of the two disorders. Findings characterize late-life GAD as a chronic disorder distinct from MDD. PMID- 15653944 TI - Risperidone for resistant anxiety in elderly persons. PMID- 15653946 TI - Incidence of delirium in very old patients after surgery for hip fracture. PMID- 15653949 TI - A dual fate of the hindlimb muscle mass: cloacal/perineal musculature develops from leg muscle cells. AB - The cloaca serves as a common opening to the urinary and digestive systems. In most mammals, the cloaca is present only during embryogenesis, after which it undergoes a series of septation events leading to the formation of the anal canal and parts of the urogenital tract. During embryogenesis it is surrounded by skeletal muscle. The origin and the mechanisms regulating the development of these muscles have never been determined. Here, we show that the cloacal muscles of the chick originate from somites 30-34, which overlap the domain that gives rise to leg muscles (somites 26-33). Using molecular and cell labelling protocols, we have determined the aetiology of cloacal muscles. Surprisingly, we found that chick cloacal myoblasts first migrate into the developing leg bud and then extend out of the ventral muscle mass towards the cloacal tubercle. The development of homologous cloacal/perineal muscles was also examined in the mouse. Concordant with the results in birds, we found that perineal muscles in mammals also develop from the ventral muscle mass of the hindlimb. We provide genetic evidence that the perineal muscles are migratory, like limb muscles, by showing that they are absent in metd/d mutants. Using experimental embryological procedures (in chick) and genetic models (in chick and mouse), we show that the development of the cloacal musculature is dependent on proximal leg field formation. Thus, we have discovered a novel developmental mechanism in vertebrates whereby muscle cells first migrate from axially located somites to the pelvic limb, then extend towards the midline and only then differentiate into the single cloacal/perineal muscles. PMID- 15653950 TI - Obstructive sleep apnea and cardiovascular disease. PMID- 15653951 TI - Minimally invasive is in the eye of the beholder: "big incisions for big operations". PMID- 15653952 TI - A sleeping giant: sleep-disordered breathing in the coronary care unit. PMID- 15653953 TI - Inhaled corticosteroids and fracture risk: having our cake and eating it too. PMID- 15653954 TI - Increased lipase plasma levels in ICU patients: when are they critical? PMID- 15653955 TI - Routine chest radiographs following central line insertion: not always necessary! PMID- 15653956 TI - Glutathione: a radical treatment for cystic fibrosis lung disease? PMID- 15653957 TI - Cardiac rhythm disturbances and ST-segment depression episodes in patients with obstructive sleep apnea-hypopnea syndrome and its mechanisms. AB - STUDY OBJECTIVES: To compare the frequency of daytime and nocturnal cardiac arrhythmias and ST-segment depression episodes among patients with obstructive sleep apnea-hypopnea syndrome (OSAHS), snoring subjects, and healthy subjects, and to analyze the relationship between the cardiac disturbances, sleep characteristics, and sympathetic tone in patients with OSAHS. PATIENTS AND INTERVENTIONS: Twenty-one consecutive patients with OSAHS, 12 snorers without hypersomnolence, and 15 healthy subjects were selected. Polysomnography, 24-h Holter ECG recording, and urinary catecholamine determination were simultaneously performed on all subjects. RESULTS: Patients with OSAHS had more daytime and nocturnal episodes of sinus and supraventricular arrhythmias and couplets than the snoring and control groups. Moreover, nocturnal ST-segment depression episodes were more frequent in the OSAHS group than in control subjects (0.565 +/ 0.826/h vs 0 +/- 0/h [mean +/- SD]). In patients with OSAHS, arousal index and daytime epinephrine levels were related to daytime and nocturnal ST-segment depression episodes, whereas minimum arterial oxygen saturation was related to nocturnal sinus bradycardia and supraventricular tachycardia. Epinephrine and norepinephrine urinary concentrations correlated with sinus and supraventricular arrhythmias. CONCLUSIONS: Patients with OSAHS have a higher frequency of cardiac rhythm disturbances and ST-segment depression episodes than snoring and control subjects. Moreover, ST-segment changes are related to sympathetic tone and sleep fragmentation, whereas most of the rhythm disturbances in patients with OSAHS are associated with sleep fragmentation, nocturnal hypoxemia, and sympathetic tone. PMID- 15653958 TI - Parasympathetic airway response and heart rate variability before and at the end of methacholine challenge. AB - BACKGROUND: The autonomic nervous system plays a primary role in regulating airway caliber, and its dysfunction is likely to contribute to the pathogenesis of airways diseases. Moreover, some findings support the hypothesis that autonomic dysfunction and/or dysregulation contributes to the pathogenesis of airway hyperresponsiveness (AHR). Heart rate variability (HRV) spectral analysis allows identifying noninvasively perturbations of the autonomic system. PURPOSES: We tested the relationship between AHR and cardiac parasympathetic tone assessed by HRV spectral analysis in patients submitted to a diagnostic methacholine bronchial challenge (MBC). METHODS: Fifteen women and 38 men (age range, 18 to 56 years) participated in the study. The principal indications for MBC were suspected asthma, chronic cough, unexplained exercise-induced dyspnea, or cough. The R-R intervals were continuously recorded during the MBC. Autoregressive method was performed on two series of 256 R-R intervals extracted before and after the MBC to obtain low-frequency (LF) and high-frequency (HF) components. RESULTS: The MBC distinguished 29 subjects without airway responsiveness (R-) and 24 responder or hyperresponsive subjects (R+): mean provocative dose of methacholine causing a 20% reduction in mean (+/- SD) FEV1 of 467 +/- 351 microg (range, 70 to 1,426 microg). The HF component expressed in normalized units (n.u.) [the index of parasympathetic modulation] was significantly higher in R+ than in R- at baseline, before MBC (21 +/- 21 n.u. vs 11 +/- 9 n.u., p < 0.05). Interestingly, R+ showed a significant increase of HF component after MBC (243 +/ 30 to 567 +/- 620 ms2 and 21 +/- 21 to 34 +/- 30 n.u., p < 0.01). For all subjects, HF (n.u.) calculated at baseline and after MBC were significantly influenced by the bronchial responsiveness (r2 = -0.28 and -0.51, respectively; p < 0.001). CONCLUSION: In summary, we found that R+ had a significantly higher parasympathetic tone than R- at baseline, and that R+ showed a significant increase in cardiac reactivity after bronchial challenge. These findings demonstrate that the autonomic nervous system, which contributes to the pathogenesis of AHR, is closely linked to cardiac modulation. PMID- 15653959 TI - Nitric oxide is released into circulation with whole-body, periodic acceleration. AB - STUDY OBJECTIVE: To determine if comfortably applied, whole-body, periodic acceleration releases significant amounts of nitric oxide (NO) into the circulation of healthy subjects and patients with inflammatory diseases. MATERIALS: Fourteen healthy adults and 40 adult patients with inflammatory diseases underwent single 45-min trials of whole-body, periodic acceleration with a new "passive exercise" device, while an ECG and a digital pulse wave were obtained with a photoelectric-plethysmograph sensor. METHODS: The position of the dicrotic notch from the pulse waveform was computed from the amplitude of the pulse divided by the height of the dicrotic notch above the end-diastolic level (a/b ratio). Increase of the a/b ratio reflects the vasodilator action of NO that causes downward movement of the dicrotic notch in the diastolic limb of the digital pulse, thereby elevating the a/b ratio. RESULTS: Application of whole body, periodic acceleration was well tolerated in all participants, and all completed the 45-min treatment. The peak value of the a/b ratio markedly rose during periodic acceleration and returned to baseline during a 5-min recovery period in all healthy subjects and patients with inflammatory diseases. CONCLUSIONS: Whole-body, periodic acceleration increased pulsatile shear stress to the endothelium leading to vasodilatation and a fall in the dicrotic notch, consistent with increased NO bioactivity in every healthy adult and adult patient with inflammatory disease so treated. Therefore, passive exercise using whole body, periodic acceleration produces an important benefit that occurs with active exercise. PMID- 15653960 TI - Persistence of lung function abnormalities despite sustained success of percutaneous mitral valvotomy: the need for an early indication. AB - AIMS: We assessed early and long-term pulmonary function changes after percutaneous balloon mitral valvotomy (PBMV). METHODS AND RESULTS: Mitral area, lung function, and exercise capacity were evaluated before, immediately after, and 3 months, 6 months, and 12 months after successful PBMV in 24 patients. PBMV resulted in a significant and sustained increase in mitral area, from 1.0 +/- 0.1 to 1.9 +/- 0.1 cm2 (p = 0.001) [mean +/- SD], with a progressive increase in exercise tolerance at 6-month follow-up (from 22.6 +/- 1.4 to 28.2 +/- 1.2 mL/kg, p = 0.0001). An immediate drop in the diffusing capacity of the lung for carbon monoxide (DLCO) was observed (from 26.7 +/- 1.5 to 22.3 +/- 1.1 mL/min/mm Hg, p = 0.0002) after PBMV, followed by a gradual regression to baseline values at 3 months; at 1 year, the DLCO remained elevated (27.3 +/- 6.3 mL/min/mm Hg). The flow in the small airways was reduced at baseline, and there was no significant change during follow-up. CONCLUSIONS: PBMV produces an initial decrease in DLCO, suggesting a reduction of pulmonary congestion. During follow-up, the regression to the initial lung diffusion values despite a sustained hemodynamic improvement suggests that some irreversible interstitial changes were present. In patients with mitral stenosis, an impairment of lung function parameters suggests that PBMV must be performed early, even if patients have few symptoms. PMID- 15653961 TI - Single-vessel revascularization with minimally invasive direct coronary artery bypass: minithoracotomy or ministernotomy? AB - STUDY OBJECTIVES: To compare the early outcome in patients who underwent off-pump single-vessel revascularization of the left anterior descending coronary artery (LAD) using two different approaches of minimally invasive direct coronary artery bypass grafting (MIDCAB): left anterior small thoracotomy (LAST) and lower ministernotomy. DESIGN: A retrospective analysis of the medical records on length of the skin incision, total operation time, duration of mechanical ventilation, blood transfusion rate, ICU stay, postoperative wound pain, and morbidities. PATIENTS: Thirty-two patients who underwent MIDCAB with the left internal thoracic artery to the LAD for single-vessel disease were studied. LAST was performed in 16 patients, and ministernotomy was performed in 16 patients. For the ministernotomy approach, the lower half of the sternum was split without transverse division, which we called the lower-end sternal splitting (LESS) approach. Postoperative pain was evaluated using a face-rating scale (scale, 1 to 6). RESULTS: There were no significant differences between the two groups in length of the skin incision, duration of mechanical ventilation, and ICU stay. Total operation time was shorter in the LESS group than in the LAST group (p < 0.05). No patients received a blood transfusion in either group. Atrial fibrillation developed in one patient in the LAST group and two patients in the LESS group. Early graft potency was 94% in the LAST group and 100% in the LESS group (p = 0.48). In the LAST group, subcutaneous emphysema developed in three patients and superficial wound dehiscence developed in two patients, but these complications were not observed in the LESS group (p < 0.05). Postoperative pain was significantly higher in the LAST group up to postoperative day 7 (p < 0.05). CONCLUSIONS: Although LAST is the most commonly used approach for MIDCAB, wound complications and postoperative pain with this technique are not insignificant compared with the lower ministernotomy approach. PMID- 15653962 TI - Thromboembolic and bleeding complications following St. Jude Medical valve replacement: results of the German Experience With Low-Intensity Anticoagulation Study. AB - STUDY OBJECTIVES: Due to their inherent thrombogenicity, mechanical cardiac valves necessitate lifelong oral anticoagulation. Less intensive oral anticoagulation than recommended earlier might result in a lower incidence of bleeding complications without increasing the embolic hazard significantly. DESIGN: Comparison of three different intensities of oral anticoagulation in a prospective, randomized multicenter design. Three months after valve replacement, patients were randomly assigned to stratum A, international normalized ratio (INR) 3.0 to 4.5; stratum B, INR 2.5 to 4.0; or stratum C, INR 2.0 to 3.5. PATIENTS: Data from 2,735 patients following aortic valve replacement (AVR; n = 2,024), mitral valve replacement (MVR; n = 553), and combined AVR and MVR (n = 158) with the St. Jude Medical (SJM) valve (St. Jude Medical; St. Paul, MN) between July 1993 and May 1999 were analyzed, covering a total follow-up period of 6,801 patient-years. All complications were registered prospectively. MEASUREMENTS AND RESULTS: Fifty-one thromboembolic events (TEs) were documented, resulting in a linearized incidence of 0.75 TEs per 100 patient-years, 22 of which were minor (0.32% per patient-year), 10 were moderate (0.15% per patient year), and 19 were severe (0.28% per patient-year). Thromboembolism following AVR was significantly lower than after MVR (0.53% per patient-year vs 1.64% per patient-year). Patients reported 1,687 bleeding complications (24.8% per patient year). The vast majority of bleeding complications (n = 1,509; 22.2% per patient year) were classified as minor, 140 were classified as moderate (2.06% per patient-year), and 38 were classified as severe (0.56% per patient-year). The clinically relevant incidences of moderate and severe TEs and bleeding complications were not significantly different between the three prespecified INR strata. CONCLUSIONS: The intention-to-treat analysis of the results of the German Experience With Low Intensity Anticoagulation study leads to the unexpected result that despite a sophisticated reporting system, the incidence of moderate and severe TE and bleeding complications was comparably low in all INR strata and more or less within the so-called background incidence reported for an age related "normal" population. This study supports reexamination of the intensity of anticoagulation in patients with the SJM valve. PMID- 15653963 TI - Use of intraoperative epiaortic ultrasonography to delineate aortic atheroma. AB - OBJECTIVE: A cerebrovascular accident (CVA) is a devastating complication of coronary artery bypass grafting (CABG) and a major cause for morbidity and mortality. Aortic manipulation, cannulation, and clamping during CABG may lead to release of atheromatous material from the ascending aorta, which may cause a CVA. This study assessed the hypothesis that the use of intraoperative epiaortic ultrasonography (EAUS) would supplement imaging information with that derived from manual aortic palpation and influence the surgical decision-making approach accordingly. METHODS: After undergoing a mid-sternotomy for CABG, 105 patients underwent EAUS with an 8-MHz transducer ordinarily used for conventional transthoracic echocardiography. The surgical strategy was decided on at three stages: preoperatively, after manual aortic palpation, and following EAUS. RESULTS: The preoperative strategy had assigned 105 patients to the "touched aorta" group that was planned for either on-pump or off-pump CABG (OPCAB) with proximal anastomosis to the aorta. Pathologic lesions of the atheromatotic ascending aorta were evident in 40 patients (38%), with the lesions detected in 22 patients (21%) by both palpation and EAUS, and in 18 patients (17%) by EAUS alone. The planned surgical strategy was changed in 29 patients (28%): 25 patients (24%) were converted from on-pump CABG to OPCAB, and the EAUS influenced the choice of the aortic cannulation, cross-clamping, and proximal anastomosis site in 4 patients (4%). Among the changes in surgical decision making, changes in 11 patients (10%) were based on lesion detection by both manual palpation and EAUS; in 18 patients (17%), changes resulted from pathologic evidence provided by EAUS alone. CONCLUSIONS: This study showed EAUS to be more sensitive in detecting atherosclerotic lesions than manual intraoperative palpation of the ascending aorta. This investigation contributes new data on the effect of EAUS on intraoperative surgical approach in the era of OPCAB. The use of EAUS has emerged as an important tool in intraoperative decision making, and we recommend its use routinely in CABG procedures. PMID- 15653964 TI - Accuracy of monitoring for sleep-related breathing disorders in the coronary care unit. AB - STUDY OBJECTIVES: To evaluate the frequency of sleep-disordered breathing (SDB) in patients presenting with acute cardiovascular events. DESIGN: Repeat observational study. SETTING: Coronary care unit of a university hospital. PATIENTS: A total of 26 patients presenting with unstable angina, myocardial infarction, or left ventricular failure. MEASUREMENTS: Level 3 portable sleep study performed at the time of acute presentation (study 1; 26 patients) and again > or = 6 weeks later (study 2; 18 patients). RESULTS: SDB (apnea-hypopnea index > or = 15) was identified in 13 of 26 patients (50%) during study 1. One patient had central sleep apnea. Of the 18 who completed the two studies, SDB was confirmed in 10 of 18 patients (56%) during study 1 but in only 5 of 18 patients (28%) during study 2. All five of those patients had obstructive sleep apnea (OSA). Six patients were deemed to have false-positive results for SDB at follow up, and one patient was deemed to have a false-negative result. Detailed analysis suggested that supine posture during study 1 may have contributed to the high false-positive rate, even though only three of six patients fulfilled the criteria for positional OSA. CONCLUSIONS: SDB occurs commonly in patients presenting with an acute cardiovascular event. Consideration of the diagnosis of SDB is an important strategy for secondary prevention. However, our findings indicate that SDB abnormalities may be transient. Sleep studies to investigate SDB as a potential risk factor for cardiovascular morbidity should be carried out when the patient is clinically stable. PMID- 15653965 TI - The French Congenital Central Hypoventilation Syndrome Registry: general data, phenotype, and genotype. AB - OBJECTIVE: To analyze the main clinical features, genetic mutations, and outcomes of patients of the French Congenital Central Hypoventilation Syndrome (CCHS) Registry. DESIGN: A country-wide cohort established throughout a long-term multicenter effort. PATIENTS: Seventy French patients with CCHS (29 male patients and 41 female patients). METHODS: The following items were analyzed: the most important moments of the disease course; the main clinical characteristics; associated pathologic conditions; management; clinical outcome; and genetic mutations. RESULTS: An average of four new cases of CCHS per year was observed in the last 5 years. Thus, the incidence may be estimated to be 1 per 200,000 live births in France. The median age at diagnosis was 3.5 months (range, 0.5 to 15 months) before 1995 and < 2 weeks in the last 5 years (p = 0.01). CCHS occurred in isolation in 58 of 70 patients. In the remainder, it was associated with Hirschsprung disease (HSCR) [nine patients], Hirschsprung and neural crest tumor (two patients), and growth hormone deficiency (one patient). Among the 50 patients who lived beyond 1 year of age, all but one received nighttime ventilation, with 10 of them (20%) receiving it noninvasively. Three patients (6%) required daytime ventilatory support in addition to nighttime ventilation. The overall mortality rate was 38% (95% confidence interval [CI], 27 to 49%). The median age at death was 3 months (range, 0.4 months to 21 years). The 2-year mortality rate was greater in male patients than in female patients (p = 0.02; relative risk [RR], 2.71; 95% CI, 1.14 to 6.47) but was not affected by HSCR (p = 0.93; RR, 0.95; 95% CI, 0.28 to 3.2). The 43 patients who are currently alive (11 men; sex ratio, 0.4) have a mean age of 9 years (range, 2 months to 27 years). Among the 34 patients tested thus far, heterozygous mutations of the paired-like homeobox gene 2B (PHOX2B) gene were found in 31 patients (91%). CONCLUSION: Our four major findings are the extreme rarity of CCHS, the improved recognition over time, the lack of effect of HSCR on the mortality rate, and the high frequency of PHOX2B mutations. PMID- 15653966 TI - Choice of oximeter affects apnea-hypopnea index. AB - STUDY OBJECTIVES: Current Medicare guidelines include an apnea-hypopnea index (AHI) > or = 15 events per hour, in which all hypopneas must be associated with 4% desaturation, to qualify for reimbursement for therapy with continuous positive airway pressure (CPAP). The present data demonstrate the effect of pulse oximeter differences on AHI. DESIGN: Prospective study, blinded analysis. SETTING: Academic sleep disorder center. PATIENTS: One hundred thirteen consecutive patients (84 men and 29 women) undergoing diagnostic sleep studies and being evaluated for CPAP based on the Medicare indications for reimbursement. INTERVENTIONS: Patients had two of four commonly used oximeters with signal averaging times of 4 to 6 s placed on different digits of the same hand during nocturnal polysomnography. MEASUREMENTS AND RESULTS: Apneas and candidate hypopneas (amplitude reduction, > 30%) were scored from the nasal cannula airflow signal without reference to oximetry. Candidate hypopneas then were reclassified as hypopneas by each oximeter if they were associated with a 4% desaturation. Although the use of three oximeters resulted in a similar AHI (bias, < 1 event per hour), the fourth oximeter showed an overall increase in AHI of 3.7 events per hour. This caused 7 of 113 patients to have an AHI of > or = 15 events per hour (meeting the Medicare criteria for treatment) by one oximeter but not when a different oximeter was used. More importantly, when our analysis was limited to those patients whose number of candidate hypopneas made them susceptible to the threshold value of 15 events per hour, 7 of 35 patients who did not meet the Medicare AHI standard for treatment by one oximeter were reclassified when a different oximeter was used. CONCLUSION: In the present study, oximeter choice affected whether the AHI reached the critical cutoff of 15 events per hour, particularly in those with disease severity that was neither very mild nor very severe. As oximetry is not a technique that produces a generic result, there are significant limitations to basing the definition of hypopnea on a fixed percentage of desaturation in determining the eligibility for CPAP therapy. PMID- 15653967 TI - The risk of nonvertebral fracture related to inhaled corticosteroid exposure among adults with chronic respiratory disease. AB - OBJECTIVE: To examine nonvertebral fracture risk in relation to inhaled corticosteroid (ICS) exposure among adults with respiratory disease. DESIGN AND PATIENTS: Nested case-control study within a cohort of 89,877 UnitedHealthcare members aged > or = 40 years with physician insurance claims for COPD or asthma, enrolled for > or = 1 year from January 1, 1997 to June 30, 2001. METHODS: Cases (n = 1,722) represented patients with a first treated nonvertebral fracture (the index date is the first fracture claim). Control subjects (n = 17,220) were randomly selected from the person-time and assigned a random index date. ICS exposure was ascertained 1 month, 3 months, 6 months, and 12 months before the index date, with estimated cumulative dose through 0 to 6 months, 7 to 12 months, and 0 to 12 months. Covariates included demographics, oral corticosteroid and other medication exposure, comorbidities, and indicators of respiratory disease severity. Odds ratios (ORs) adjusted for all covariates were estimated by logistic regression. RESULTS: No increased fracture risk with ICS exposure as a class or with fluticasone propionate alone was detected. ORs for exposure in the preceding 30 days were 1.05 (95% confidence interval [CI], 0.89 to 1.24), 1.13 (95% CI, 0.90 to 1.40), and 0.97 (95% CI, 0.78 to 1.21) for all ICS, fluticasone propionate, and other ICS, respectively. No dose-response effect was present. Among patients with COPD only (n = 6,932), no increased risk was found for recent ICS exposure (OR, 0.86; 95% CI, 0.59 to 1.25). CONCLUSIONS: Concern about nonvertebral fracture risk should not strongly influence the decision to use recommended doses of ICS for adult patients with asthma or COPD in managed-care settings in the United States. This study could not evaluate very-high ICS dose, long-term ICS exposure, or vertebral fracture risk. PMID- 15653968 TI - Transtracheal high-flow insufflation supports spontaneous respiration in chronic respiratory failure. AB - STUDY OBJECTIVES: Transtracheal insufflation of oxygen-enriched air at a high flow rate has been proposed to support ventilation. The purpose of this study was to investigate the physiologic effects of high-flow insufflation unobtrusively with a respiratory inductive plethysmograph in patients with chronic respiratory failure. Using a respiratory inductive plethysmograph also permitted monitoring of end-expiratory lung volume, and respiratory variables could be quantified independently of the tracheal bias flow. DESIGN: Prospective randomized comparison of low-flow vs high-flow transtracheal insufflation. SETTING: Pulmonary division of a tertiary teaching hospital. PATIENTS: Fourteen spontaneously breathing outpatients with chronic hypoxemic respiratory failure carrying a transtracheal catheter for long-term oxygen therapy. INTERVENTIONS AND MEASUREMENTS: Oxygen-enriched air (fraction of inspired oxygen, 0.37) at 15 L/min and oxygen at 1.5 L/min were transtracheally administered for 1 h each. The breathing pattern and the end-expiratory lung volume were monitored by inductive plethysmography along with pulse oximetry and transcutaneous PCO2. Arterial blood gases were also analyzed at the end of the hour of both low-flow and high-flow insufflation. RESULTS: High-flow insufflation decreased the mean (+/- SEM) minute ventilation (Ve) by 20% from 8.37 +/- 0.49 to 6.66 +/- 0.57 L/min, the mean respiratory rate from 19.2 +/- 0.9 to 15.7 +/- 1.0 breaths/min, while mean expiratory time increased from 2.0 +/- 0.1 to 2.8 +/- 0.2 s, and end-expiratory lung volume decreased by 0.55 +/- 0.15 L compared to low-flow oxygen insufflation (p < 0.05 for all comparisons). Mean arterial and transcutaneous PCO2 decreased from 45 +/- 1 to 43 +/- 1 mm Hg and from 54 +/- 2 to 53 +/- 2 mm Hg, respectively (p < 0.05 in both instances), while arterial PaO2 and oxygen saturation did not change. CONCLUSIONS: High-flow transtracheal insufflation of oxygen-enriched air assists ventilation by reducing Ve without compromising gas exchange and by reducing end-expiratory lung volume, possibly through the reversal of dynamic hyperinflation. PMID- 15653970 TI - Exercise outcomes after pulmonary rehabilitation depend on the initial mechanism of exercise limitation among non-oxygen-dependent COPD patients. AB - STUDY OBJECTIVES: Pulmonary rehabilitation (PR) that includes exercise training can improve exercise tolerance and quality of life for patients with COPD. However, the degree of benefit from PR is variable. We hypothesized that the exercise response to PR varies depending on the initial factors that limit exercise. DESIGN, SETTING, PARTICIPANTS, AND MEASUREMENTS: We retrospectively analyzed the change in exercise capacity after PR in 290 nonhypoxemic patients with COPD. We classified patients into the following subgroups based on the primary limitation seen on initial exercise testing: (1) ventilatory-limited (VL); (2) cardiovascular-limited (CVL); (3) mixed ventilatory/cardiovascular limited (VLCVL); and (4) non-cardiopulmonary-limited (NL). We compared outcomes among subgroups. RESULTS: In the entire study population, PR led to increased timed walk distance (30.3%; p < 0.0001) and maximal oxygen consumption (VO2max) [84.8 mL/min; p < 0.0001]. Stepwise multiple regression selected age, ventilatory reserve at peak exercise, and exercise arterial oxygen pressure as individual predictors of improvement in VO2max. VO2max increased in the VL subgroup (30.4 mL/min; p = 0.008), the CVL subgroup (109.0 mL/min; p < 0.0001), the mixed VLCVL subgroup (61.3 mL/min; p < 0.0001), and NL subgroups (110.5 L/min; p < 0.0001). The improvement in VO2max was greater in the CVL subgroup than in the VL subgroup (p < 0.0001). Timed walk distance improved to a similar degree in all subgroups (26 to 36%). CONCLUSIONS: Patients with nonventilatory exercise limitations experience the greatest increase in VO2max after PR. However, even patients with severe ventilatory limitation can improve exercise tolerance with PR. PMID- 15653969 TI - Length and clinical effectiveness of pulmonary rehabilitation in outpatients with chronic airway obstruction. AB - STUDY OBJECTIVE: To assess the clinical effectiveness of pulmonary rehabilitation (PR) after 10 or 20 consecutive sessions in outpatients with chronic airway obstruction (CAO). DESIGN: Observational prospective cohort trial. SETTING: Outpatient clinic of a rehabilitation center. PATIENTS AND INTERVENTIONS: Twenty five outpatients (mean age, 65 +/- 9 years [+/- SD]; FEV1, 64 +/- 12% predicted) admitted to a comprehensive PR program, including exercise training. MEASUREMENTS AND RESULTS: The load reached on a cycloergometer (maximal achieved load [W max]), the maximal and isoload dyspnea and leg fatigue on a Borg scale, 6-min walk distance (6MWD), and the health-related quality of life as assessed using the St. George's Respiratory Questionnaire (SGRQ) [total and components score] have been recorded as outcome measures at baseline, after 10 sessions (T10), and after 20 sessions (T20). The predefined criteria of the clinically significant improvement were as follows: + 15% W-max, + 54 m at 6MWD, - 1 point at isoload dyspnea and leg fatigue, and - 4% at SGRQ scores. There was a mean significant difference between changes at T20 and T10 for 6MWD (- 42.96 m; 95% confidence interval [CI], - 57.79 to - 28.12 m; p = 0.001), total SGRQ (4.80; 95% CI, 2.29 to 7.31; p = 0.001), activity SGRQ (3.60; 95% CI, 0.48 to 6.71; p = 0.025), and symptoms SGRQ (5.96; 95% CI, 2.72 to 9.2; p = 0.001). The percentage of patients who improved was different at T20 as compared with T10 for W-max (68% and 48%, respectively; p = 0.025), 6MWD (76% and 20%, p = 0.001), and total SGRQ (64% and 36%, p = 0.008). CONCLUSIONS: A 10-session course of PR provides only limited clinically significant changes of outcome measures when compared with a 20 session course in outpatients with CAO of mild-to-moderate severity. PMID- 15653971 TI - Step-down compared to fixed-dose treatment with inhaled fluticasone propionate in asthma. AB - BACKGROUND: Inhaled corticosteroids (ICSs) are an effective treatment of asthma even when administered at a low dose. Once asthma is controlled, current guidelines recommend that the dose of ICS be reduced to the lowest possible and effective dose. Although the most appropriate strategy for the stepping down has not yet been defined, quantification of sputum eosinophils and bronchial hyperresponsiveness (BHR) are indeed measures of asthma control. OBJECTIVE: To compare the efficacy of step-down and fixed-dose strategies in the control of BHR to methacholine and eosinophilic inflammation patients with mild-to-moderate asthma. METHODS: We performed a double-blind, randomized study to compare inhaled fluticasone propionate (FP), 1,000 microg/d, then reduced to 200 microg/d (group 1; n = 18) to a fixed dose of FP, 200 microg/d (group 2; n = 17) administered for 6 weeks and then 8 weeks in reducing the provocative dose of methacholine causing a 20% fall in FEV1 (PD20) and sputum eosinophils in 35 patients. The duration of the efficacy was also followed subsequently after 8 weeks of placebo treatment. RESULTS: PD20 remarkably increased with both treatment strategies, but differences between groups were not significant. Sputum eosinophils (median values, percentage) at baseline and after each treatment period were not different (group 1, 16.4 to 1.0 to 2.7%; group 2, 16.7 to 2.8 to 2.8%, respectively). The percentages of patients in whom sputum eosinophilia was normalized (< or = 3%) were as follows: group 1, 69% and 60%; group 2, 50% and 57%. After placebo treatment, sputum eosinophils were still "normalized" in approximately one third of patients. CONCLUSION: Step-down and fixed-dose strategies with FP improved PD20 and sputum eosinophilia to a similar degree. The effect on sputum eosinophils persisted longer than that on methacholine. PMID- 15653972 TI - Phenol-containing saline solution as a diluent for adenosine 5'-monophosphate in bronchial challenge testing. AB - OBJECTIVES: To investigate the effect of dissolving adenosine 5'-monophosphate (AMP) with phenol-containing saline solution on the stability and the bronchoconstrictive properties of this indirect agonist. METHODS: Eleven subjects with asthma well controlled with short-acting inhaled beta2-agonists as required or with inhaled corticosteroids were studied. Bronchial challenge tests with AMP dissolved with either normal saline solution or saline solution containing 0.4% phenol were performed on separate days. Furthermore, to assess the potential influence of the phenol-containing saline solution on the stability of the bronchoconstrictor agent, AMP solutions in concentrations of 40 microg/mL and 400 microg/mL were prepared in saline solution and phenol-containing saline solution and, after 30 min, the AMP levels were determined by high-performance liquid chromatography (HPLC) assay. RESULTS: The geometric mean AMP provocative concentration causing a 20% fall in FEV1 (PC20) was 13.49 mg/mL (95% confidence interval [CI], 6.76 to 26.91) for the saline solution method, and AMP PC20 for the saline solution with phenol method was 8.91 mg/mL (95% CI, 3.39 to 23.44) [p = 0.18]. No significant differences were found between the concentrations of AMP made in saline solution compared to those made in phenol-containing saline solution measured by HPLC. CONCLUSION: These observations indicate that normal saline solution with or without phenol can be used as the diluent for AMP. However, since a potential risk with AMP of industrial sources is the bacterial contamination, adding a preservative such as phenol to a saline solution diluent might be recommended. PMID- 15653973 TI - Effect of spirometric maneuver, nasal clip, and submaximal inspiratory effort on measurement of exhaled nitric oxide levels in asthmatic patients. AB - STUDY OBJECTIVES: The measurement of exhaled nitric oxide (eNO) in asthmatic patients is increasingly being used to aid diagnosis and management. To standardize the measurement techniques, the American Thoracic Society and European Respiratory Society guidelines were published, but these were based mainly on expert opinions without strong clinical evidence on many aspects. We investigated the effect of three different factors on the on-line measurement of eNO. In a clinical setting, we evaluated the effect of prior spirometry, the use of nasal clips, and the influence of submaximal inspiratory effort on on-line eNO readings. Recommended guidelines on these factors have been published but have been supported by scanty research data. METHODS: Three groups of stable asthmatic patients (30 patients in each group) had eNO measurements made on-line (NIOX; Aerocrine AB; Solna, Sweden) before and 5 min after performing spirometric maneuvers, without and with a nasal clip, or with maximal and then submaximal inspiratory efforts. RESULTS: There were no significant differences in mean eNO levels among all three groups, before and after spirometry (68.2 vs 66.0 parts per billion [ppb], respectively; mean difference, 2.2 ppb; 95% confidence interval [CI], -0.4 to 4.9; p = 0.090), without use of a nasal clip compared with its use (46.7 vs 45.6 ppb, respectively; mean difference, 1.1 ppb; 95% CI, -0.7 to 2.8; p = 0.234), and maximal or submaximal inspiratory effort (52.6 vs 51.2 ppb, respectively; mean difference, 1.4 ppb; 95% CI, -0.3 to 3.0; p = 0.096). CONCLUSION: We conclude that on-line eNO measurements in clinical practice are not significantly affected by prior spirometry maneuver, use of a nasal clip, or submaximal inspiratory effort. PMID- 15653974 TI - Deep inspiration avoidance and methacholine response in normal subjects and patients with asthma. AB - BACKGROUND: Deep inspiration (DI) avoidance and time intervals between inhalation and measurement of FEV1 may influence methacholine challenges. OBJECTIVES: (1) To compare the degree of airway response to methacholine when the initial FEV1 measurements are obtained either 30 s or 3 min after inhalation, (2) to evaluate a simplified method to study the influence of DI avoidance before inhalation on the fall in FEV1, and (3) to determine if methacholine has a cumulative effect. PARTICIPANTS/METHODS: Twenty-five patients with asthma and 21 normal subjects without asthma. Four methacholine inhalation tests (MITs) were performed: two standard tidal-breathing MITs, with the first FEV1 measured 30 s (test A) and 3 min (test B) after the end of inhalation; a single-dose MIT, using the last concentration from test B, with no control of DI and the first FEV1 obtained 3 min after inhalation (test C); and an identical single-dose MIT preceded by 20 min of DI avoidance (test D). We compared the provocative concentration of methacholine causing a 20% fall in FEV1 (PC20) from tests A and B (aim 1), the percentage fall in FEV1 from tests C and D (aim 2), and the percentage fall in FEV1 from tests B and C (aim 3). RESULTS: Mean PC20 values from tests A and B were 1.5 mg/mL and 1.0 mg/mL (p = 0.002) in patients with asthma, and 69.8 mg/mL and 29.9 mg/mL (p < 0.0001) in control subjects, respectively. The mean falls in FEV1 for tests C and D were 22.0% and 24.5% (p > 0.05) in patients with asthma, and 22.1% and 38.9% (p = 0.0005) in control subjects, respectively. The mean falls in FEV2 for tests B and C were 30.2% and 22.0% (p = 0.01) in patients with asthma, and 27.5% and 22.1% (p > 0.05) in control subjects, respectively. CONCLUSIONS: In both groups, the longer the time interval between the end of inhalation and the first FEV2 measurement, the greater the fall in FEV2 (lower PC20). DI avoidance before inhalation does not enhance the fall in FEV2 in subjects with asthma, while it does in control subjects. Methacholine has a slight cumulative effect that is significant in patients with asthma (p = 0.007). PMID- 15653975 TI - Cardiac and sternocleidomastoid muscle involvement in Duchenne muscular dystrophy: an MRI study. AB - OBJECTIVE: To examine the extent of cardiac muscle and sternocleidomastoid muscle (SCM) involvement detected by MRI measurement of T2 relaxation time in patients with Duchenne muscular dystrophy (DMD) and no cardiorespiratory symptoms. DESIGN: Prospective controlled study. SETTING: Teaching referral hospital and university hospital. SUBJECTS: Seventeen patients with DMD (age range, 7 to 25 years) and 17 age-matched control subjects. All patients were free of cardiac or respiratory complaints and had normal ECG, echocardiograph, and Holter monitor examination findings. METHODS: We assessed respiratory function by means of standard pulmonary function testing. MRI measurements included the T2 relaxation time of the myocardium and the SCM in patients and control subjects. RESULTS: The FVC and FEV1 values were lower in patients with DMD than in age-matched control subjects, whereas the FEV1/FVC ratio was normal in all subjects. Patients with DMD had lower T2 relaxation time of the heart (37.8 +/- 6.1 ms vs 58.1 +/- 7.1 ms, p < 0.001) and lower T2 relaxation time of the right SCM (24.5 +/- 2.6 ms vs 42.2 +/- 1.3 ms, p < 0.001) and left SCM (23.2 +/- 3.2 ms vs 42.2 +/- 1.6 ms, p < 0.001), compared to control subjects (+/- SD). In children (< 12 years of age), the T2 of the SCM was lower than that of the control subjects, but T2 of the heart did not differ between the two groups. In the patient group, T2 relaxation time of the heart decreased with age (r = - 0.80, p < 0.001). In patients with FVC < 80% of predicted, the T2 values of the heart were lower than the T2 values of patients with FVC > or = 80% of predicted (35.6 +/- 5.8 ms vs 41.8 +/- 4.6 ms, p < 0.05). CONCLUSIONS: MRI measurements of the T2 relaxation time in the myocardium and SCM of patients with DMD and no cardiorespiratory symptoms are abnormal, indicating altered tissue composition. These measurements may prove a clinically useful test for monitoring cardiac and respiratory muscle involvement in these patients. PMID- 15653976 TI - Assessment of airway caliber and bronchodilator responsiveness in subjects with spinal cord injury. AB - STUDY OBJECTIVES: Previous spirometric findings among subjects with chronic tetraplegia that reduction in FEV1 and maximal forced expiratory flow, mid expiratory phase (FEF(25-75%)) correlated with airway hyperresponsiveness to histamine, and that many of these subjects exhibited significant bronchodilator responsiveness, suggested that baseline airway caliber was low in this population. To better evaluate airway dynamics in patients with spinal cord injury, we used body plethysmography to determine specific airway conductance (sGaw), a less effort-dependent and more reflective surrogate marker of airway caliber. DESIGN: Cohort study. SETTING: Veterans Affairs medical center. PARTICIPANTS: Thirty clinically stable subjects with chronic spinal cord injury, including 15 subjects with tetraplegia (injury at C4-C7) and 15 subjects with low paraplegia (injury below T7), participated in the study. Fifteen able-bodied individuals served as a control group. INTERVENTIONS: Subjects underwent baseline assessment of spirometric and body plethysmographic parameters. Repeat measurements were performed among subjects with tetraplegia and paraplegia before and 30 min after receiving aerosolized ipratropium bromide (2.5 mL 0.02% solution; 12 subjects) or normal saline solution (2.5 mL; 6 subjects). MEASUREMENTS AND RESULTS: We found that subjects with tetraplegia had significantly reduced mean values for sGaw (0.16 cm H2O/s), total lung capacity, FVC, FEV1, and FEF(25-75%) compared to subjects in the other two groups. Subjects with tetraplegia who received ipratropium bromide experienced significant increases in sGaw (135%), FEV1 (12%; 260 mL), and FEF(25-75%) (27%). Significant, though far smaller, increases in sGaw (19%) were found among subjects with paraplegia. No discernable change in any pulmonary function parameter was found following the administration of normal saline solution. CONCLUSIONS: Subjects with tetraplegia, as opposed to those with low paraplegia, have reduced baseline airway caliber due to heightened vagomotor airway tone, which we hypothesize is the result of the interruption of sympathetic innervation to the lungs, and/or from low circulating epinephrine levels. PMID- 15653977 TI - Increasing peak expiratory flow time in amyotrophic lateral sclerosis. AB - BACKGROUND: Serial measurements of pulmonary function to indicate decreasing respiratory muscle strength in patients with amyotrophic lateral sclerosis (ALS) is well documented. Quantitative outcome measures include declining FVC, FEV1, maximal inspired pressure, maximal expired pressure, and maximal voluntary ventilation. Increasing peak expiratory flow time (PEFT) may represent a further sensitive measure of declining respiratory muscle strength in ALS. METHODS: Fifty five patients with ALS performed flow-volume loops serially after presentation. The percentage change from baseline values for FVC, peak expiratory flow (PEFR), and PEFT were compared using Spearman correlation coefficients. The prolongation of PEFT with serial tests was analyzed using a Kruskal-Wallis with a Dunn multiple comparison test. Bulbar-onset and limb-onset PEFT was compared using the Mann-Whitney test. RESULTS: PEFT was significantly increased from baseline values at all follow-up tests. However, PEFTs measured at the third, fourth, fifth, and sixth visits, although higher, were not significantly different. Significant negative correlations existed between the increase in PEFT and the decrease in PEFR and FVC. Significant positive correlations existed between the increase in PEFT and days from diagnosis and the decrease in PEFR and decrease in FVC. CONCLUSION: PEFT increases significantly and linearly with time in patients with ALS and may begin to plateau with bulbar symptoms. PEFT increases at a faster rate than the rate of decline in both FVC and PEFR. PEFT is a quantitative measure of decreasing pulmonary function in ALS that is easily measured. PMID- 15653978 TI - Impact of primary graft failure on outcomes following lung transplantation. AB - STUDY OBJECTIVES: Primary graft failure (PGF) is a severe acute lung injury syndrome that occurs following lung transplantation. We compared the clinical outcomes of patients who developed PGF with those who did not. METHODS: We conducted a retrospective cohort study including 255 consecutive lung transplant procedures. PGF was defined as (1) diffuse alveolar opacities developing within 72 h of transplantation, (2) an arterial partial pressure of oxygen/fraction of inspired oxygen (PaO2/FiO2) ratio of < 200 beyond 48 h postoperatively, and (3) no other secondary cause of graft dysfunction. PGF was tested for acceptance with 30-day and all-cause hospital mortality rates, overall survival, hospital length of stay (HLOS), duration of mechanical ventilation, and best 6-min walk test (6MWT) distance achieved within 12 months. SETTING: Academic medical center. RESULTS: The overall incidence of PGF was 11.8% (95% confidence interval [CI], 7.9 to 15.9%). The all-cause mortality rate at 30 days was 63.3% in patients with PGF and 8.8% in patients without PGF (relative risk [RR], 7.15; 95% CI, 4.34 to 11.80%; p < 0.001). A total of 73.3% of patients with PGF died during hospitalization vs 14.2% of patients without PGF (RR, 5.18%; 95% CI, 3.51 to 7.63; p < 0.001). The median HLOS in 30-day survivors was 47 days in patients with PGF vs 15 days in those without PGF (p < 0.001), and the mean duration of mechanical ventilation was 15 days in patients with PGF vs 1 day in those without PGF (p < 0.001). By 12 months, a total of 28.5% of survivors with PGF achieved a normal age-appropriate 6MWT distance vs 71.4% of survivors without PGF at 12 months (p = 0.014). The median best 6MWT distance achieved within the first 12 months was 1,196 feet in patients with PGF vs 1,546 feet in those without PGF (p = 0.009). CONCLUSIONS: PGF has a significant impact on mortality, HLOS, and duration of mechanical ventilation following lung transplantation. Survivors of PGF have a protracted recovery with impaired physical function up to 1 year following transplantation. PMID- 15653979 TI - Psychosocial situation and physical health in 50 patients > 1 year after lung transplantation. AB - BACKGROUND: Lung transplants have been performed worldwide since the early 1980s. While numerous studies have been published on somatic aspects after lung transplantation, there is considerably less information available on psychosocial aspects and on the correlation between the physical and the psychosocial state of health after transplantation. METHODS: Between 1992 and 2002, 125 patients underwent lung transplantation at University Hospital Zurich. To be included into the study, patients had to have received a lung transplant > 12 months previously and to have good knowledge of the German or Italian languages. With the aid of standardized questionnaires, psychosocial variables such as levels of anxiety and depression, self-esteem, and social support were determined. In addition, self assessments of physical and psychological health were obtained. The medical data included information on FEV1, complications such as pulmonary infections, acute or chronic allograft rejection, and assessment of the patient's physical and psychological health by the treating doctors. RESULTS: The overall degree of anxiety and depression of the lung transplant recipients was comparable to standard samples of an average population. However, male lung transplant recipients were significantly more depressed than female recipients. Self-esteem was higher than in clinical comparison samples. Preceding pulmonary complications had long-lasting effects on the level of anxiety, whereas nonpulmonary complications did not have such an effect. CONCLUSIONS: Overall, the psychological well-being of patients after lung transplantation is similar to the normal population. Subgroups of patients with increased psychological distress have been identified. PMID- 15653980 TI - Analyses of efficacy end points in a controlled trial of interferon-gamma1b for idiopathic pulmonary fibrosis. AB - BACKGROUND: Idiopathic pulmonary fibrosis (IPF) is a devastating disease, yet validated, reliable criteria for evaluating patient response to therapies in clinical trials are lacking. METHODS: To optimize selection of end point criteria for the study of interferon (IFN)-gamma1b in patients with IPF, we retrospectively analyzed the components of the primary efficacy end point used in a large, controlled study of 330 patients for reliability, validity, and sensitivity to treatment effect. The primary end point components were death, disease progression defined as a > or = 5 mm Hg increase in resting alveolar arterial oxygen pressure gradient (P[A-a]O2), and disease progression defined as a > or = 10% decrease in percentage of predicted FVC. RESULTS: We found that the P(A-a)O2 criterion was not reliable and was not associated with mortality. In contrast, the FVC criterion was reliable and was associated with a 2.4-fold increase in the risk of death. Of the three measures, only mortality was sensitive to a treatment effect of IFN-gamma1b. Additionally, the tendency for mortality benefit was observed in nearly all patient subgroups defined by baseline physiology. The effect of IFN-gamma1b on mortality was strongest in patients with baseline percentage of predicted FVC > or = 55% (p = 0.004) or percentage of predicted diffusing capacity of the lung for carbon monoxide > or = 30% (p = 0.008). CONCLUSION: We conclude that mortality is the most inclusive end point for future trials of IFN- gamma1b in patients with IPF, and that a > 10% decrement in the percentage of predicted FVC represents a valid measure of disease progression. PMID- 15653981 TI - Desquamative interstitial pneumonia and respiratory bronchiolitis-associated interstitial lung disease. AB - BACKGROUND: Desquamative interstitial pneumonia (DIP) and respiratory bronchiolitis-associated interstitial lung disease (RB-ILD) are uncommon forms of interstitial lung disease and have been incompletely characterized. STUDY OBJECTIVES: To further characterize the clinical features and course of subjects with DIP and RB-ILD. DESIGN: Retrospective study. SETTING: Tertiary care, referral medical center. PATIENTS: Twenty-three subjects with DIP and 12 subjects with RB-ILD seen over a 12-year period between 1990 and 2001. INTERVENTIONS: None. RESULTS: The study population included 19 men (54%) and 16 women (46%). The mean (+/- SD) age at diagnosis was 46 +/- 10 and 43 +/- 7 years, respectively, for patients with DIP and RB-ILD. All subjects were either current or previous smokers except for three subjects with DIP. The diagnosis was confirmed in all cases by surgical lung biopsy. Bronchoscopy with transbronchial lung biopsy had been performed in 12 patients and was nondiagnostic in all. The most common pulmonary function abnormality was a reduced diffusing capacity of the lung for carbon monoxide. A CT scan of the chest revealed ground-glass opacities bilaterally in most patients who had DIP and RB-ILD. No differences were observed between subjects with DIP and RB-ILD with respect to clinical features, radiologic findings, or pulmonary function test results. The clinical course was characterized by relative stability in the majority of patients in both groups and a partial response to corticosteroid therapy. Five deaths were observed, including three resulting from progressive diffuse lung disease, all in subjects with DIP. CONCLUSIONS: We concluded that DIP and RB-ILD are chronic disease processes that in most patients are related to smoking. Persistent abnormalities can be seen on pulmonary function testing and radiologic studies despite smoking cessation and corticosteroid therapy. Corticosteroid therapy appeared to be associated with modest clinical benefit but usually not with resolution of disease. Progressive disease with eventual death can occur in subjects with DIP, especially with continued cigarette smoking. PMID- 15653982 TI - Long-term follow-up CT scan evaluation in patients with pulmonary sarcoidosis. AB - OBJECTIVES: The aim of the present study was to determine how the pattern and extent of sarcoidosis changes over time on serial high-resolution CT (HRCT) scans and to identify CT scan findings that might be helpful in predicting the prognosis of patients with the disease. METHODS: The initial and last HRCT scan findings of 40 patients with pulmonary sarcoidosis during a mean follow-up period of 7.4 years (range, 3 to 18 years) were evaluated retrospectively. HRCT scan findings then were correlated with the pulmonary function tests. RESULTS: Parenchymal abnormalities in most patients with a predominant nodular pattern (18 patients) and multiple large nodular pattern (8 patients) disappeared or decreased in size on long-term follow-up CT scans. A conglomeration pattern (five patients) shrank and evolved into bronchial distortion. The shrinkage of the conglomeration pattern correlated with a decline of FEV1/FVC ratio, despite an improvement in FVC. A ground-glass opacity pattern (five patients) and a consolidation pattern (three patients) evolved into honeycombing. The evolution of the ground-glass opacity and consolidation patterns into honeycombing occurred along with a decline in FVC, although the parenchymal abnormalities became smaller. CONCLUSION: Patients with a predominantly ground-glass opacity pattern and consolidation pattern seen on the initial CT scan had a worse prognosis and were susceptible to developing severe respiratory insufficiency. The predominant patterns seen on the initial HRCT scan may be helpful in predicting the outcomes of patients with sarcoidosis. PMID- 15653983 TI - An assessment of back pain and the prevalence of sacroiliitis in sarcoidosis. AB - OBJECTIVES: Sarcoidosis is a chronic granulomatous multisystem disease in which arthritis is relatively common. Arthritis of the sacroiliac joints (sacroiliitis) has been described in sarcoidosis but is thought to be rare. The objective of this study was to determine the prevalence of sacroiliitis in a secondary-care population of patients with sarcoidosis. METHODS: Patients attending a specialist secondary-care sarcoidosis clinic underwent evaluation of spinal symptoms using a standard back pain questionnaire, examination of spinal mobility, and laboratory measurements of erythrocyte sedimentation rate, C-reactive protein, serum angiotensin-converting enzyme, and neopterin/creatinine ratio. Tissue typing for the presence of the human leukocyte antigen (HLA)-B27 allele was undertaken. Radiographs of the sacroiliac joints were obtained in each patient and reviewed independently by two observers; a further observer reviewed disputed radiographs. RESULTS: Sixty-one patients completed the assessments (80.3% of all patients invited to participate). Forty-nine of 61 patients (80.3%) reported having back pain at some point in their lives. Thirty-one of 61 patients (50.8%) had a score on the back pain questionnaire suggestive of inflammatory spinal disease, but only 3 of these patients had erosive damage of the sacroiliac joints on radiography indicating sacroiliitis. One further patient had erosive damage on radiography, making a total of four individuals with evidence of sacroiliitis, a prevalence of 6.6%. Four patients (one patient with sacroiliitis) were positive for HLA-B27. The back pain questionnaire had a sensitivity of 75% and a specificity of 51% for sacroiliitis in this population. CONCLUSION: The prevalence of spondyloarthropathy in the normal population has been estimated to be 1.9%. In the sarcoid population studied the prevalence was 6.6% suggesting a possible association between these two conditions. The standard back pain questionnaire for the identification of inflammatory spinal disease had a low sensitivity and specificity in this population. PMID- 15653984 TI - Antibody production deficiency with normal IgG levels in bronchiectasis of unknown etiology. AB - BACKGROUND: No defined cause of bronchiectasis is currently found in approximately 50% of cases. Bronchiectasis is a common long-term complication in patients with primary hypogammaglobulinemia. STUDY OBJECTIVES: To ascertain whether antibody production deficiency with normal total serum IgG levels is associated with bronchiectasis. DESIGN: Antibody response to a pneumococcal unconjugate vaccine and an Haemophilus influenzae type b conjugate vaccine was prospectively studied in all consecutive adult patients with bronchiectasis of unknown etiology who were assessed in our chest outpatient clinic from January 1994 to October 2001. Serum-specific antibodies were measured by enzyme-linked immunosorbent assay, and the results were compared with those obtained in a healthy adult control group. Antibody production deficiency was defined as a failure to respond to either vaccine. RESULTS: One hundred seven patients were included in the study (mean age, 46.3 years). Antibody production deficiency was diagnosed in 12 patients (11%). A significantly higher incidence of otitis media, lower serum IgG2 subclass levels, and lower preimmunization antibody levels to Streptococcus pneumoniae and H influenzae type b were observed in patients with antibody production deficiency. The probability of antibody production deficiency in patients with a history of otitis media was 20%, 26% in those with low IgG2 subclass levels, and 58% in those with both a history of otitis media and low IgG2 subclass levels. CONCLUSIONS: Antibody production deficiency with normal IgG levels may be associated with bronchiectasis, making it advisable to evaluate the antibody response to both the H influenzae and pneumococcal vaccines in patients with bronchiectasis of unknown etiology, particularly in those with a history of otitis media, low IgG2 subclass levels, and low levels of baseline specific antibodies. PMID- 15653985 TI - Delays in suspicion and isolation among hospitalized persons with pulmonary tuberculosis at public and private US hospitals during 1996 to 1999. AB - BACKGROUND: While prior studies have shown that public and private hospitals differ in their rates of suspicion and isolation of patients who are at risk for tuberculosis (TB), no study has investigated whether this variation is due to differences in the impact of patient case-mix on hospitals or to variations attributable to specific hospital practice patterns. OBJECTIVE: To investigate patient-level and hospital-level factors associated with delays in TB suspicion and isolation among inpatients with pulmonary TB disease. DESIGN: Retrospective cohort study of patients hospitalized with culture-positive pulmonary TB during 1996 to 1999. SETTING: Patients with culture-proven pulmonary TB treated at three public hospitals (765 patients) and seven not-for-profit private hospitals (172 patients) in Chicago, Los Angeles, and southern Florida that provided care for five or more patients with TB per year during the study period. MEASUREMENTS: Two day rates (within 48 h from admission) of acid-fast bacilli (AFB) smear orders and 1-day rates (within 24 h from admission) of TB isolation. RESULTS: Two-day rates of ordering AFB smears were > 80% at three public and two private hospitals vs 65 to 75% at five private hospitals. One-day rates of TB isolation at the three public hospitals were 64%, 79%, and 86%, respectively, vs 39 to 58% at the seven private hospitals. Delays of > 2 days in ordering AFB smears were associated with patient-level factors: absence of cough (adjusted odds ratio [AOR], 6.02; 95% confidence interval [CI], 3.82 to 9.52), cavitary lung lesion (AOR, 5.17; 95% CI, 1.98 to 13.50), night sweats (AOR, 3.38; 95% CI, 1.90 to 5.99), chills (AOR, 1.70; 95% CI, 1.01 to 2.88), and female gender (AOR, 1.66; 95% CI, 1.06 to 2.60). Delays of > 1 day in ordering pulmonary isolation were associated with patient-level factors: absence of cough (AOR, 3.40; 95% CI, 2.31 to 5.03), cavitary lung lesion (AOR, 2.66; 95% CI, 1.57 to 4.50), night sweats (AOR, 1.98; 95% CI, 1.35 to 2.92), and history of noninjecting drug use (AOR, 1.86; 95% CI, 1.16 to 2.99) and one hospital-level factor: receiving care at a nonpublic hospital. Even after adjustment for patient-level factors, TB patients at private hospitals were half as likely as those at public hospitals to be placed in pulmonary isolation (AOR, 0.47; 95% CI, 0.30 to 0.72), while odds of suspecting TB in these same patients were similar at both hospitals. CONCLUSION: Private hospitals should order TB isolation for all patients for whom AFB smears are ordered, a policy that has been instituted previously at public hospitals in our study. PMID- 15653986 TI - Multicenter study of hospital-acquired pneumonia in non-ICU patients. AB - STUDY OBJECTIVE: To know the incidence, epidemiology, etiology, and outcome of hospital-acquired pneumonia (HAP) in non-ICUs adult patients. SETTING: Twelve Spanish teaching hospitals. INTERVENTIONS: From April 1999 to November 2000, non ICU HAP was prospectively studied by active, bimonthly 1-week surveillance. Epidemiologic data, etiology, and evolution of pneumonia were recorded. Blood and sputum cultures and Legionella pneumophila and Streptococcus pneumoniae urinary antigen tests were performed. RESULTS: We included 186 patients, with complete data available in 165 patients (70.3% male gender; mean age, 63.7 +/- 16.9 years [ +/- SD]) The mean incidence of HAP was 3 +/- 1.4 cases/1,000 hospital admissions. Most patients (64.2%) were in medical wards, had severe underlying diseases (66.6%), and had a hospital stay > 5 days (76.4%). Blood cultures were performed in 139 patients (84.2%), sputum cultures were performed in 89 patients (53.9%), and urinary antigen detection was performed in 123 patients (74.5%). An etiologic diagnosis was obtained in 60 cases (36.4%), and 31 were definitive. The most frequent etiologies were S pneumoniae (16 cases, 14 definitive), L pneumophila (7 cases, 7 definitive), Aspergillus sp (7 cases, 3 definitive), Pseudomonas aeruginosa (7 cases, 2 definitive), and several Enterobacteriaceae (8 cases, 4 definitive). Clinical complications occurred in 52.1% of the cases, and mortality was 26% (13.9% attributed to pneumonia). CONCLUSIONS: Non-ICU HAP is an important cause of hospital morbidity, observed most frequently in medical wards and elderly patients with severe underlying diseases. In this setting, S pneumoniae and Legionella sp should be considered in addition to other nosocomial pathogens; urinary antigen detection is useful in determining the prevalence of these microorganisms. PMID- 15653987 TI - Is chest radiography necessary after uncomplicated insertion of a triple-lumen catheter in the right internal jugular vein, using the anterior approach? AB - STUDY OBJECTIVES: Chest radiographs are required in many institutions by protocol after the insertion of a right internal jugular vein triple-lumen catheter (TLC), even if the anterior approach is used. This study investigates whether correct placement can be predicted during insertion and whether a "routine" postprocedural chest radiograph can be safely omitted. DESIGN: The operators included 18 first-, second-, or third-year medical residents, 3 pulmonary fellows, and a board-certified pulmonary medicine and critical care attending, with at least 1 certified physician present during the procedure. All operators were trained in the "seven number rule." PATIENTS: One hundred consecutive patients who required central venous access. Patients with left internal jugular vein or subclavian catheters were excluded. SETTING: Single institution, medical ICU, step-down unit, and floors. INTERVENTIONS: Right internal jugular vein TLC insertion, anterior approach, with subsequent chest radiograph. MEASUREMENTS AND RESULTS: Eighty-eight patients had uncomplicated insertions, as defined by fewer than four sticks with a 22-gauge pathfinder needle and fewer than four slides with the 18-gauge introducer needle. Ninety-eight catheters were in accurate position, 1 catheter was in the distal superior cava vein, and 1 catheter was in an S-shaped position. CONCLUSIONS: It is safe to omit the routine chest radiograph after uncomplicated insertion of a TLC. i.v. treatment can be initiated early. However, if there is any doubt about the correct position, a chest radiograph should be obtained. PMID- 15653988 TI - Usefulness of ultrasonography in predicting pleural effusions > 500 mL in patients receiving mechanical ventilation. AB - STUDY OBJECTIVE: To assess the accuracy of chest ultrasonography in predicting pleural effusions > 500 mL in patients receiving mechanical ventilation. DESIGN: Prospective study. SETTING: Surgical and medical ICU in a teaching hospital. PATIENTS: Forty-four patients receiving mechanical ventilation with indications of chest drainage of a nonloculated pleural effusion. INTERVENTIONS: Diagnosis of pleural effusion was based on clinical examination and chest radiography. Chest drainage was indicated when considered as potentially useful for the patient (hypoxemia and/or weaning failure). Sonograms were performed before drainage at the bedside, in the supine position, and measurements were performed at the end of expiration. Effusions were classified as > 500 mL or < or = 500 mL according to the drained volume. MEASUREMENTS AND RESULTS: The drained volume ranged from 100 to 1,800 mL (mean, 730 +/- 440 mL [+/- SD]). The distance between the lung and posterior chest wall at the lung base (PLDbase) and the distance between the lung and posterior chest wall at the fifth intercostal space (PLD5) were significantly correlated with the drained volume (PLDbase, r = 0.68, p < 0.001; PLD5, r = 0.56, p < 0.001). A PLDbase > 5 cm predicted a drained volume > 500 mL with a sensitivity of 83%, specificity of 90%, positive predictive value of 91%, and negative predictive value of 82%. Interobserver and intraobserver percentages of error were, respectively, 7 +/- 6% and 9 +/- 6% for PLDbase, and 6 +/- 5% and 8 +/- 5% for PLD5. The PaO2/fraction of inspired oxygen ratio significantly increased after chest drainage in patients with collected volumes > 500 mL (p < 0.01). CONCLUSIONS: Bedside pleural ultrasonography accurately predicted a nonloculated pleural effusion > 500 mL in patients receiving mechanical ventilation using simple and reproducible measurements. PMID- 15653989 TI - Cytomegalovirus infection in critically ill patients: associated factors and consequences. AB - OBJECTIVE: To determine the prevalence, associated findings, and consequences of cytomegalovirus (CMV) antigenemia in critically ill patients. DESIGN: A retrospective, case-control clinical study. SETTING: A 12-bed university hospital medical-surgical ICU. PATIENTS: Two hundred thirty-seven patients with fever for > 72 h, without proven evidence of bacteriologic and/or fungal origin, and whose pp65 antigenemia assays were studied. Patients with HIV infection and transplant recipients were excluded. INTERVENTIONS: None. MEASUREMENTS AND RESULTS: CMV antigenemia was diagnosed within 20 +/- 12 days (mean +/- SD) after ICU admission in 17% patients in whom the pathology was suspected. The 40 patients in the CMV group were matched with 40 other patients in the control group. CMV infection was linked to renal failure (58% vs 33%, respectively; p = 0.02) and steroid use (55% vs 33%, respectively; p = 0.04). Patients with CMV had a significantly longer stay in the ICU (41 +/- 28 days vs 31 +/- 22 days, respectively; p = 0.04), a longer duration of mechanical ventilation (35 +/- 27 days vs 24 +/- 20 days, respectively; p = 0.03), a higher rate of nosocomial infection (75% vs 50%, respectively; p = 0.04), and a higher mortality (50% vs 28%, p = 0.02). CONCLUSIONS: CMV antigenemia is not an uncommon diagnosis in critically ill ICU patients with unexplained prolonged fever after 10 days of hospitalization, regardless of their immune system status. Although associated with a higher morbidity and mortality, the clinical significance of CMV is unknown. Further prospective studies should evaluate the impact on ICU outcome and whether CMV is truly a pathogen or simply another indicator of immunosuppression. PMID- 15653990 TI - Inclusion criteria for clinical trials in sepsis: did the American College of Chest Physicians/Society of Critical Care Medicine consensus conference definitions of sepsis have an impact? AB - BACKGROUND: Over the last 25 years, a growing number of clinical trials have evaluated novel sepsis therapies. To promote uniformity in inclusion criteria for patient enrollment, the American College of Chest Physicians and Society of Critical Care Medicine first published consensus conference definitions for sepsis in 1992. STUDY OBJECTIVES: To characterize (1) the utilization of specific criteria for patient enrollment in sepsis clinical trials and (2) the impact that the consensus conference definitions have had on these criteria. DESIGN: We used MEDLINE to identify clinical trials in sepsis from 1976 to 2001. Clinical trials published after the consensus conference (ACC; from 1993 to 2001) were compared with trials published before the consensus conference (BCC; from 1976 to 1992). RESULTS: We identified 176 clinical trials (ACC, 119 trials; BCC, 57 trials). Clinical trials published ACC were more likely to utilize or reference a previously published standard for inclusion criteria (65% vs 11%, respectively; p < 0.001). The consensus conference definitions were the standards used in 69% of these trials. The utilization of specified values for WBC count, temperature (T), heart rate (HR), and respiratory rate (RR) was significantly increased in the ACC group compared to the BCC group, as follows: WBC count, 62% vs 26%, respectively (p < 0.001); T, 89% vs 56%, respectively (p < 0.001); HR, 77% vs 26%, respectively (p < 0.001); and RR, respectively 76% vs 28% (p < 0.001). ACC, clinical trials were less likely to require blood culture positivity (4 of 119 trials [3%] vs 9 of 57 trials [16%], respectively; p < 0.006) and were more likely to incorporate markers of acute organ dysfunction (81 of 119 trials [68%] vs 28 of 57 trials [49%], respectively; p < 0.03) in the inclusion criteria. CONCLUSIONS: (1) Since 1992 there has been a significant increase in the utilization of predefined sepsis criteria for patient enrollment in clinical trials, and this increase can be attributed to the existence of consensus conference definitions. (2) Compared to inclusion criteria BCC, inclusion criteria ACC were less reliant on blood culture positivity and were more likely to incorporate markers of organ dysfunction. PMID- 15653991 TI - Clinical significance of increased lipase levels on admission to the ICU. AB - STUDY OBJECTIVES: To examine the incidence, risk factors, and sequelae associated with asymptomatic hyperlipasemia in the ICU. SETTING: Medical and surgical ICUs. PATIENTS: Two hundred forty-five adult critically ill patients admitted to an ICU for > 72 h with a diagnosis other than pancreatitis were studied prospectively. MEASUREMENTS: Serum amylase and lipase were measured on ICU admission and every third day until normalized. Clinical parameters including the incidence of ileus, the ability to tolerate enteral feeds, and the results of radiologic studies were also recorded. RESULTS: Hyperlipasemia was present in 40% of patients (peak, 1,183 +/- 175 U/L; range, 209 to 8,620 U/L) [mean +/- SEM]. Increased multiple organ dysfunction scores, hypotension, anemia, mechanical ventilation (MV), bacteremia, elevated liver function test results, and elevated creatinine and triglyceride levels were all associated with increased lipase levels. In multivariate analysis, hypotension, anemia, elevated serum bilirubin, and MV were independently associated with higher lipase levels. Although mortality was not different, ICU length of stay and the duration of MV were significantly greater in patients with increased lipase levels (p < 0.05). Fifty patients underwent imaging studies. Pancreatitis was confirmed in 11 patients. The mean peak lipase value was significantly increased in patients with a positive study finding as compared to those with negative findings: 2,231 +/- 715 U/L and 900 +/- 234 U/L, respectively (p < 0.01). Enteral feedings, when initiated, were tolerated in 94% of patients with increased lipase levels and 97% of patients with normal lipase levels. CONCLUSIONS: Elevated serum lipase levels are frequently encountered in critically ill patients. In the majority of these patients, enteral feedings are well tolerated and there are minimal clinical sequelae. Extremely high lipase levels may be associated with radiologic evidence of pancreatitis. Hypoperfusion and inflammatory processes associated with multiple-organ failure appear to be contribute to these increases. PMID- 15653992 TI - Production of soluble tumor necrosis factor receptors and tumor necrosis factor alpha by alveolar macrophages in sarcoidosis and extrinsic allergic alveolitis. AB - BACKGROUND: Alveolar macrophage (AM)-derived tumor necrosis factor (TNF)-alpha plays a pivotal role in the pathogenesis of sarcoidosis and extrinsic allergic alveolitis (EAA). The effects of TNF-alpha are mediated by membrane TNF receptor (mTNFR)-1 and mTNFR-2, and can be blocked by soluble TNF receptor (sTNFR)-1 and sTNFR-2. METHODS: We measured the production of the two sTNFRs and TNF-alpha in AM culture supernatants from 10 patients with active sarcoidosis, 12 patients with EAA, and 9 control subjects using an enzyme-linked immunosorbent assay method. RESULTS: Compared with control subjects, the spontaneous and lipopolysaccharide (LPS)-stimulated production of sTNFR-1, sTNFR-2, and TNF-alpha was significantly increased in patients with sarcoidosis and EAA. The concentrations of both sTNFRs, but especially of sTNFR-2, were closely related to those of TNF-alpha. The LPS-induced increase was 1.5-fold for sTNFR-1, at least fourfold for sTNFR-2, and at least 25-fold for TNF-alpha in all study populations. CONCLUSION: These results indicate that AMs can release the two sTNFRs in relation to TNF-alpha. sTNFR-2 may be more liable to shedding than sTNFR-1. Both sTNFR-1 and sTNFR-2 may be involved in the pathogenesis of sarcoidosis and EAA, possibly as counterregulators of TNF-alpha. PMID- 15653993 TI - Anti-tissue remodeling effects of corticosteroids: fluticasone propionate inhibits fibronectin expression in fibroblasts. AB - OBJECTIVES: Tissue remodeling often accompanies diseases such as COPD that are caused by or aggravated by tobacco exposure. Inhaled or systemic corticosteroids are frequently used for the treatment of these illnesses, and their beneficial effects are often ascribed to their anti-inflammatory properties. However, their role in tissue remodeling remains unclear. This study was designed to evaluate the role of corticosteroids in matrix expression in vitro. DESIGN: We investigated the effects of the corticosteroid fluticasone propionate (FP) on the production of fibronectin by fibroblasts before and after stimulation by nicotine, a plant alkaloid found in tobacco. Fibronectin is an extracellular matrix glycoprotein found elevated in the alveolar lining fluid and airway walls of subjects with obstructive lung disease, and is considered a marker of tissue remodeling after injury. RESULTS: FP, 1 micromol/L, inhibited the expression of fibronectin messenger RNA and protein in unstimulated NIH-3T3 cells and primary lung fibroblasts, as well as in fibroblasts stimulated with nicotine. The inhibitory effect of FP occurred at the level of gene transcription as demonstrated in lung fibroblasts expressing a construct containing the human fibronectin promoter connected to a luciferase reporter gene, but posttranscriptional effects also appeared involved. Electrophoresis mobility gel shift assays revealed that FP inhibited phosphorylation and DNA binding by the cyclic adenosine monophosphate response element binding protein, a transcription factor required for constitutive and nicotine-induced fibronectin expression. CONCLUSIONS: Together, these data suggest that FP could diminish lung tissue remodeling by inhibiting the production of fibronectin in lung fibroblasts. PMID- 15653994 TI - Expression of apoptotic and antiapoptotic markers in epithelial cells in idiopathic pulmonary fibrosis. AB - STUDY OBJECTIVE: Idiopathic pulmonary fibrosis (IPF) is a chronic, usually fatal lung disease of unknown etiology. A common feature is the presence of microscopic areas of epithelial cell dropout. Increased apoptosis of these cells could elucidate the speculative pathogenesis of the disease. Therefore, the aim of our study was to examine the expression of p53, p21, bcl-2, bax, and caspase-3 in association with DNA strand breaks in bronchial and alveolar epithelial cells in lung specimens from IPF patients and control subjects. PATIENTS AND METHODS: We examined by immunohistochemistry the expression of p53, p21, bax, bcl-2, and caspase-3 in association with DNA strand breaks detected by terminal deoxynucleotide transferase-mediated deoxyuridine triphosphate-biotin nick end labeling (TUNEL) in bronchial and alveolar epithelial cells in lung specimens taken by biopsy in 12 IPF patients and 10 control subjects. An independent tissue evaluation by two pathologists graded semiquantatively the degree of staining present. RESULTS: TUNEL was positive in epithelial cells in all IPF patients and only in one control subject. The expression of p53, p21, bax, and caspase-3 was up-regulated in IPF patients compared to control subjects. Bcl-2 was expressed less in IPF patients than in control subjects. CONCLUSIONS: These results confirm that apoptotic hyperplastic epithelial cells are present in patients with IPF and that the expression of p53, p21, bax, and caspase-3 appears to be up-regulated and that of bcl-2 down-regulated in these cells. The increased expression of proapoptotic molecules in epithelial cells in IPF may be involved in the inadequate and delayed reepithelialization, which in turn contributes to fibroblast proliferation. PMID- 15653995 TI - Idiopathic pulmonary fibrosis: challenges and opportunities for the clinician and investigator. AB - Idiopathic pulmonary fibrosis (IPF) is a relentlessly progressive and typically fatal interstitial lung disease. Besides its grave natural history and prognosis, three aspects of IPF challenge clinicians and investigators: (1) recent changes in the conceptual framework and definition of IPF complicate interpretation of prior clinical investigations; (2) while most patients with suspected IPF do not undergo open-lung biopsy, clinical definitions that do not include biopsy criteria have not been validated prospectively; and (3) available treatments have not been shown to be effective. To optimize clinical care and facilitate clinical investigation, a major goal of IPF research should be to develop validated sets of clinical diagnostic and prognostic criteria. Studies have shown the diagnostic value of high-resolution CT scans and identified important prognostic variables; many of these observations await prospective validation. While previous therapeutic studies have been limited by small sample sizes, lack of a placebo control group, and insufficient attention to patient-centered outcomes, the recent study of interferon gamma-1b demonstrated the feasibility of a large scale, multicenter clinical trial in IPF. In this article, we discuss how overcoming challenges in IPF research will enable future investigators to conduct well-designed observational studies and clinical trials, whose meaningful results will advance our understanding of IPF, its management, and its impact on patients' lives. PMID- 15653996 TI - Health-related quality of life among patients with idiopathic pulmonary fibrosis. AB - The hallmark of idiopathic pulmonary fibrosis (IPF) is relentless and progressive breathlessness culminating in respiratory failure and death. Clinicians and investigators are increasingly aware that many patients with chronic diseases, like IPF, value the quality of their lives as much or more than their length of survival. Despite this growing awareness, little research has focused on quality of life (QOL) in IPF patients. Nevertheless, the few studies that have been performed uniformly show the negative impacts of IPF on QOL, particularly in the areas of physical health, energy, and symptoms. To fill important knowledge gaps, more research is needed. For example, future studies in well-defined IPF patient populations should rigorously assess the psychometric properties of different measures of QOL. Currently, there is no disease-specific instrument for use in patients with IPF. A carefully developed, IPF-specific instrument that includes items most relevant to IPF patients should be more sensitive to change than existing generic or non-IPF respiratory disease-specific instruments. Longitudinal assessments are needed to map the trajectory of QOL in relation to disease progression and to reveal whether different aspects of QOL become impaired over time. Addressing these research opportunities will markedly improve our knowledge of this outcome, which is highly valued by patients. These endeavors will also help clinicians who care for patients with IPF to develop a better understanding of its profound negative impact, and it will help future IPF clinical investigators to select the most valid, reliable, and appropriate QOL instrument to fill the roles their studies require. PMID- 15653997 TI - Anemia, allogenic blood transfusion, and immunomodulation in the critically ill. AB - Anemia and allogenic RBC transfusions are exceedingly common among critically ill patients. Multiple pathologic mechanisms contribute to the genesis of anemia in these patients. Emerging risks associated with allogenic RBC transfusions including the transmission of newer infectious agents and immune modulation predisposing the patient to infections requires reevaluation of current transfusion strategies. Recent data have suggested that a restrictive transfusion practice is associated with reduced morbidity and mortality during critical illness, with the possible exception of acute coronary syndromes. In this article, we review the immune-modulatory role of allogenic RBC transfusions in critically ill patients. PMID- 15653998 TI - A pilot study of the effect of inhaled buffered reduced glutathione on the clinical status of patients with cystic fibrosis. AB - STUDY OBJECTIVES: To assess the impact of inhaled, buffered reduced glutathione (GSH) on clinical indicators of cystic fibrosis (CF) pathophysiology. DESIGN AND PATIENTS: A randomized, double-blind, placebo-controlled pilot study was conducted over an 8-week period. Nineteen subjects, age 6 to 19 years, with CF status documented by positive sweat chloride test results (> 60 mEq/L) were recruited for the trial. After matching on age and sex, 10 patients were randomly assigned to the treatment group and 9 patients to the placebo group. Primary outcomes were FEV1, FVC, forced expiratory flow at 25 to 75% of vital capacity, and peak flow; secondary outcomes were body mass index, 6-min walk distance, and self-reported cough frequency, mucus production/viscosity/color, wellness, improvement, and stamina. INTERVENTIONS AND ANALYSIS: Treatment was buffered GSH, and placebo was sodium chloride with a hint of quinine. The total daily dose of buffered GSH was approximately 66 mg/kg of body weight, and the total daily dose of placebo was approximately 15 mg/kg of body weight (quinine, 25 to 30 microg/kg). Doses were distributed across four inhalation sessions per day and spaced 3- to 4-h apart. General linear mixed models were used to analyze the data. The final sample size was nine subjects in the treatment group and seven subjects in the placebo group. RESULTS: Mean change for peak flow was -6.5 L/min for the placebo group and +33.7 L/min for the GSH group (p = 0.04), and self reported average improvement on a scale from 1 to 5 (1 being much worse and 5 being much better) was 2.8 for placebo and 4.7 for GSH (p = 0.004). Of the 13 primary and secondary outcomes examined, 11 outcomes favored the treatment group over the placebo group (p = 0.002), indicating a general tendency of improvement in the GSH group. No adverse events in the treatment group were noted. CONCLUSION: This pilot study indicates the promise of nebulized buffered GSH to ameliorate CF disease, and longer, larger, and improved studies of inhaled GSH are warranted. PMID- 15653999 TI - Initial and final exercise heart rate transients: influence of gender, aerobic fitness, and clinical status. AB - STUDY OBJECTIVES: To compare the independent and additive data provided by initial and final heart rate (HR) exercise transients, and to analyze both according to gender, aerobic fitness, clinical status, and medication usage. DESIGN: Retrospective study. SETTING: Exercise medicine clinic. PATIENTS: A total of 544 subjects (363 men) with a mean (+/- SD) age of 50 +/- 14 years (age range, 10 to 91 years), including asymptomatic and coronary artery disease patients. MEASUREMENTS AND RESULTS: HR transients were obtained from the following two exercise protocols: 4-s exercise test (4sET) followed by a maximal cardiopulmonary cycling exercise test (CPET). The initial HR transient was represented by the cardiac vagal index (CVI), which was obtained by the 4sET, and the final transient (ie, HR recovery [HRR]) was determined by the following equation: CPET maximal HR - the 1-min postexercise HR. Transients were modestly related (r = 0.22; p < 0.001) when adjusted for age, aerobic fitness, clinical status, and negative chronotropic action drug usage. The transients were unrelated to gender (vs CVI, p = 0.10; vs HRR, p = 0.15). Subjects with a measured maximum oxygen uptake (VO2max) exceeding 100% of the predicted maximal aerobic power showed higher CVIs than those in less aerobically fit subjects (VO2max < 50% subgroup, p = 0.009; VO2max < 75% subgroup, p = 0.034). Both transient results differed for asymptomatic and cardiac subjects (CVI, 1.32 +/- 0.02 vs 1.42 +/- 0.02, respectively [p = 0.001]; HRR, 33 +/- 1 beats/min (bpm) vs 37 +/- 1 bpm, respectively [p = 0.009]). CONCLUSIONS: The initial and final HR transients were modestly related, suggesting a potentially complementary clinical role for both measurements in the assessment of autonomic function in patients with coronary artery disease. Although both HR transients tended to behave similarly under the influence of several variables, the initial HR transient, measured during 4sET, was more likely to discriminate distinct subgroups compared with the final HR transient. PMID- 15654000 TI - Association of depression and life-sustaining treatment preferences in patients with COPD. AB - STUDY OBJECTIVE: Depressive symptoms and reduced health-related quality of life are common in patients with severe COPD. Therefore, understanding the association between preferences for life-sustaining treatment and depression or quality of life is important in providing care. No prior studies have examined the effects of depression and quality of life on treatment preferences in this population. DESIGN AND PATIENTS: Cross-sectional study of 101 patients with oxygen-prescribed COPD. METHODS: Patients completed the St. George's Respiratory Questionnaire, Center for Epidemiologic Studies-Depression survey, and questions regarding their preferences for mechanical ventilation and cardiopulmonary resuscitation if needed to sustain life. RESULTS: Median age was 67.4 years, and median FEV1 was 26.3% predicted. Depression was significantly associated with preferences for resuscitation (50% of depressed patients and 23% of patients without depression refused resuscitation; p = 0.007), but was not associated with preferences for mechanical ventilation. Health-related quality of life was not associated with preferences for either resuscitation or mechanical ventilation. CONCLUSIONS: Clinicians caring for patients with oxygen-prescribed COPD should understand that health-related quality of life does not predict treatment preferences and should not influence clinicians' views of patients' treatment preferences. However, depression does appear to influence patients' treatment decisions for cardiopulmonary resuscitation, and improvement in depressive symptoms should trigger a reassessment of these preferences. PMID- 15654001 TI - Device selection and outcomes of aerosol therapy: Evidence-based guidelines: American College of Chest Physicians/American College of Asthma, Allergy, and Immunology. AB - BACKGROUND: The proliferation of inhaler devices has resulted in a confusing number of choices for clinicians who are selecting a delivery device for aerosol therapy. There are advantages and disadvantages associated with each device category. Evidence-based guidelines for the selection of the appropriate aerosol delivery device in specific clinical settings are needed. AIM: (1) To compare the efficacy and adverse effects of treatment using nebulizers vs pressurized metered dose inhalers (MDIs) with or without a spacer/holding chamber vs dry powder inhalers (DPIs) as delivery systems for beta-agonists, anticholinergic agents, and corticosteroids for several commonly encountered clinical settings and patient populations, and (2) to provide recommendations to clinicians to aid them in selecting a particular aerosol delivery device for their patients. METHODS: A systematic review of pertinent randomized, controlled clinical trials (RCTs) was undertaken using MEDLINE, EmBase, and the Cochrane Library databases. A broad search strategy was chosen, combining terms related to aerosol devices or drugs with the diseases of interest in various patient groups and clinical settings. Only RCTs in which the same drug was administered with different devices were included. RCTs (394 trials) assessing inhaled corticosteroid, beta2-agonist, and anticholinergic agents delivered by an MDI, an MDI with a spacer/holding chamber, a nebulizer, or a DPI were identified for the years 1982 to 2001. A total of 254 outcomes were tabulated. Of the 131 studies that met the eligibility criteria, only 59 (primarily those that tested beta2-agonists) proved to have useable data. RESULTS: None of the pooled metaanalyses showed a significant difference between devices in any efficacy outcome in any patient group for each of the clinical settings that was investigated. The adverse effects that were reported were minimal and were related to the increased drug dose that was delivered. Each of the delivery devices provided similar outcomes in patients using the correct technique for inhalation. CONCLUSIONS: Devices used for the delivery of bronchodilators and steroids can be equally efficacious. When selecting an aerosol delivery device for patients with asthma and COPD, the following should be considered: device/drug availability; clinical setting; patient age and the ability to use the selected device correctly; device use with multiple medications; cost and reimbursement; drug administration time; convenience in both outpatient and inpatient settings; and physician and patient preference. PMID- 15654002 TI - Allergic rhinitis, asthma, and atopy among grape farmers in a rural population in Crete, Greece. AB - STUDY OBJECTIVE: To measure the prevalence of allergic rhinitis, atopy, and asthma among grape farmers, and to compare the respiratory and atopic status in grape farmers with those of nonexposed control subjects. DESIGN: Cross-sectional study. SETTING: Malevisi region in northern Crete, Greece. SUBJECTS AND METHODS: One hundred twenty grape farmers and 100 control subjects living in the Malevisi region were examined. The protocol comprised a questionnaire, skin prick tests for 16 common allergens, measurement of specific IgE antibodies against 8 allergens, and spirometry before and after bronchodilation. RESULTS: Grape farmers were found to have an excess of respiratory symptoms. The comparison with the control group, after adjusting for age, sex, and smoking status, showed that the differences were statistically significant for rhinorrhea (odds ratio [OR], 2.7; 95% confidence interval [CI], 1.5 to 5.1; p < 0.001), sneezing (OR, 2.2; 95% CI, 1.2 to 4.0; p < 0.01), and nasal itching (OR, 1.9; 95% CI, 1.0 to 3.6; p < 0.05), but were nonsignificant for asthma-related symptoms. In the multiple logistic regression model, grape farmers were found to have increased work related symptoms, such as sneezing (OR, 2.9; 95% CI, 1.3 to 6.6; p < 01), rhinorrhea (OR, 2.9; 95% CI, 1.3 to 6.6; p < 0.01), cough (OR, 3.7; 95% CI, 1.2 to 11.4; p < 0.05), and dyspnea (OR, 3.8; 95% CI, 1.1 to 1.3; p < 0.05). The prevalence of allergic rhinitis was 40.8% in grape farmers and 26% in control subjects (OR, 2.0; 95% CI, 1.1 to 3.5; p < 0.02). Increased but statistically nonsignificant values of asthma prevalence were found in grape farmers (6.7%) compared with the control group (2.0%). The prevalence of atopy was 64.2% in grape farmers and 38.0% in the control group (OR, 2.2; 95% CI, 1.2 to 3.5; p < 0.01). Mean FEV1 was significantly lower in grape farmers than in control subjects (p < 0.05), after adjusting for age, sex, and smoking status. Bronchial obstruction was reversible in 23 grape farmers (19.2%) and in 6 control subjects (6%; p < 0.01). CONCLUSIONS: The study mainly demonstrated the high prevalence of allergic rhinitis and work-related respiratory symptoms in grape farmers compared to control subjects. It also suggested that grape farming is possibly associated with increased allergic sensitization to specific pollens, low baseline FEV1, and increased bronchial hyper-responsiveness. Further studies are needed to determine the potential risk factors for these disorders among the farming population. PMID- 15654003 TI - Hemodynamic monitoring utilizing transesophageal echocardiography: the relationships among pressure, flow, and function. PMID- 15654004 TI - A 41-year-old man with altered mental status and acute flaccid paralysis. PMID- 15654005 TI - The air crescent sign: A clue to the etiology of chronic necrotizing pneumonia. PMID- 15654006 TI - Severe refractory hypoxemia following a gunshot injury. AB - We describe the case of a 57-year-old man with severe refractory hypoxemia, despite receiving ventilation therapy with 100% oxygen, following a gunshot wound to his left chest. A limited CT scan of the chest with contrast raised the suspicion of an arteriovenous (AV) fistula. Contrast-enhanced transthoracic echocardiography confirmed the presence of a pulmonary AV fistula. Traumatic pulmonary AV fistula is a rare, but serious and life-threatening condition that should be suspected in patients with thoracic injuries with persistent unexplained hypoxemia. Contrast echocardiography is a relatively simple, inexpensive, and readily available bedside test that can be used to confirm this diagnosis. PMID- 15654007 TI - Identification and treatment of bronchoconstriction induced by a vagus nerve stimulator employed for management of seizure disorder. AB - We evaluated a 63-year-old woman who developed dyspnea with a sensation of chest tightness that was temporally associated with discharges from a vagus nerve stimulator that had been implanted for the control of intractable seizures. Spirometry demonstrated the development of significant airflow obstruction associated with the firing of the stimulator. Adjustment of the stimulator settings resolved the discharge-associated bronchoconstrictive phenomenon. These findings highlight an important association between vagus nerve stimulators and dyspnea that should be considered in the differential diagnosis of patients with these devices who present with dyspnea and/or chest tightness. The relative importance of vagal stimulation to bronchoconstriction is suggested by the findings. PMID- 15654008 TI - Serum KL-6 and surfactant proteins A and D in pediatric interstitial lung disease. AB - OBJECTIVE: To determine if serum KL-6, surfactant protein A (SP-A), and surfactant protein D (SP-D) levels are elevated in pediatric interstitial lung disease (ILD) and associated with pulmonary function and disease severity score. METHODS: Serum KL-6, SP-A, and SP-D levels were measured by enzyme-linked immunosorbent assay in 10 children with ILD and in 10 healthy volunteers. In the ILD group, FEV1 percentage of predicted, FVC percentage of predicted, and ILD disease severity score were measured and correlated with serum KL-6, SP-A, and SP D levels. RESULTS: For the ILD and control groups, respectively, mean serum KL-6 was 4,523 U/mL and 206 U/mL (p = 0.007), mean serum SP-A was 133 ng/mL and 21 ng/mL (p = 0.003), and mean serum SP-D was 304 ng/mL and 75 ng/mL (p = 0.004). There was an inverse relationship between SP-A and FVC (p = 0.05), and between SP D and FEV1 (p = 0.05). There was a direct relationship between SP-D and ILD score (p = 0.05). CONCLUSIONS: Serum KL-6, SP-D and SP-D levels are elevated in children with ILD. SP-A and SP-D levels appear to correlate with some measures of disease severity. PMID- 15654009 TI - Xenobiotic enzymes and genetics of COPD. PMID- 15654010 TI - Inaccurate spirometry results? Let's blame it on the computer! PMID- 15654011 TI - Pitfalls in the interpretation of multivariable models in the critical care literature. PMID- 15654012 TI - The economic impact of late detection of COPD in general practice. PMID- 15654013 TI - Extracorporeal membrane oxygenation and pulmonary disease. PMID- 15654014 TI - Obesity in ICU patients: increase or decrease in mortality? PMID- 15654015 TI - Death receptor signaling. PMID- 15654016 TI - RNA localization mechanisms in oocytes. AB - In many animals, normal development depends on the asymmetric distribution of maternal determinants, including various coding and noncoding RNAs, within the oocyte. The temporal and spatial distribution of localized RNAs is determined by intricate mechanisms that regulate their movement and anchoring. These mechanisms involve cis-acting sequences within the RNA molecules and a multitude of trans acting factors, as well as a polarized cytoskeleton, molecular motors and specific transporting organelles. The latest studies show that the fates of localized RNAs within the oocyte cytoplasm are predetermined in the nucleus and that nuclear proteins, some of them deposited on RNAs during splicing, together with the components of the RNA-silencing pathway, dictate the proper movement, targeting, anchoring and translatability of localized RNAs. PMID- 15654017 TI - The developmental cell biology of Trypanosoma brucei. AB - Trypanosoma brucei provides an excellent system for studies of many aspects of cell biology, including cell structure and morphology, organelle positioning, cell division and protein trafficking. However, the trypanosome has a complex life cycle in which it must adapt either to the mammalian bloodstream or to different compartments within the tsetse fly. These differentiation events require stage-specific changes to basic cell biological processes and reflect responses to environmental stimuli and programmed differentiation events that must occur within a single cell. The organization of cell structure is fundamental to the trypanosome throughout its life cycle. Modulations of the overall cell morphology and positioning of the specialized mitochondrial genome, flagellum and associated basal body provide the classical descriptions of the different life cycle stages of the parasite. The dependency relationships that govern these morphological changes are now beginning to be understood and their molecular basis identified. The overall picture emerging is of a highly organized cell in which the rules established for cell division and morphogenesis in organisms such as yeast and mammalian cells do not necessarily apply. Therefore, understanding the developmental cell biology of the African trypanosome is providing insight into both fundamentally conserved and fundamentally different aspects of the organization of the eukaryotic cell. PMID- 15654018 TI - Remodelling of the nuclear lamina and nucleoskeleton is required for skeletal muscle differentiation in vitro. AB - Changes in the expression and distribution of nuclear lamins were investigated during C2C12 myoblast differentiation. The expression of most lamins was unchanged during myogenesis. By contrast, lamin-B2 expression increased and LAP2alpha expression decreased twofold. These changes were correlated with reduced solubility and redistribution of A-type lamins. When C2C12 myoblasts were transfected with a lamin-A mutant that causes autosomal dominant Emery-Dreifuss muscular dystrophy (AD-EDMD), the mutant protein accumulated in the nucleoplasm and exerted dominant influences over endogenous lamins. Myoblasts transfected with wild-type lamins differentiated, albeit more slowly, whereas myoblasts transfected with mutant lamins failed to differentiate. Myoblast differentiation requires dephosphorylation of the retinoblastoma protein Rb. During myogenesis, Rb was rapidly and progressively dephosphorylated. Underphosphorylated Rb formed complexes with LAP2alpha in proliferating myoblasts and postmitotic myoblasts. In myoblasts transfected with the mutant lamins, this complex was disrupted. These data suggest that remodelling of the nucleoskeleton is necessary for skeletal muscle differentiation and for correct regulation of Rb pathways. PMID- 15654019 TI - Distinct subcellular location of the Ca2+-binding protein S100A1 differentially modulates Ca2+-cycling in ventricular rat cardiomyocytes. AB - Calcium is a key regulator of cardiac function and is modulated through the Ca2+ sensor protein S100A1. S100 proteins are considered to exert both intracellular and extracellular functions on their target cells. Here we report the impact of an increased intracellular S100A1 protein level on Ca2+-homeostasis in neonatal ventricular cardiomyocytes in vitro. Specifically, we compare the effects of exogenously added recombinant S100A1 to those resulting from the overexpression of a transduced S100A1 gene. Extracellularly added S100A1 enhanced the Ca2+ transient amplitude in neonatal ventricular cardiomyocytes (NVCMs) through a marked decrease in intracellular diastolic Ca2+-concentrations ([Ca2+]i). The decrease in [Ca2+]i was independent of sarcoplasmic reticulum Ca2+-ATPase (SERCA2a) activity and was probably the result of an increased sarcolemmal Ca2+ extrusion through the sodium-calcium exchanger (NCX). At the same time the Ca2+ content of the sarcoplasmic reticulum (SR) decreased. These effects were dependent on the uptake of extracellularly added S100A1 protein and its subsequent routing to the endosomal compartment. Phospholipase C and protein kinase C, which are tightly associated with this subcellular compartment, were found to be activated by endocytosed S100A1. By contrast, adenoviral-mediated intracellular S100A1 overexpression enhanced the Ca2+-transient amplitude in NVCMs mainly through an increase in systolic [Ca2+]i. The increased Ca2+-load in the SR was based on an enhanced SERCA2a activity while NCX function was unaltered. Overexpressed S100A1 colocalized with SERCA2a and other Ca2+ regulatory proteins at the SR, whereas recombinant S100A1 protein that had been endocytosed did not colocalize with SR proteins. This study provides the first evidence that intracellular S100A1, depending on its subcellular location, modulates cardiac Ca2+-turnover via different Ca2+-regulatory proteins. PMID- 15654021 TI - Mcp6, a meiosis-specific coiled-coil protein of Schizosaccharomyces pombe, localizes to the spindle pole body and is required for horsetail movement and recombination. AB - We report here that a meiosis-specific gene of Schizosaccharomyces pombe denoted mcp6+ (meiotic coiled-coil protein) encodes a protein that is required for the horsetail movement of chromosomes at meiosis I. The mcp6+ gene is specifically transcribed during the horsetail phase. Green fluorescent protein (GFP)-tagged Mcp6 appears at the start of karyogamy, localizes to the spindle-pole body (SPB) and then disappears before chromosome segregation at meiosis I. In the mcp6Delta strain, the horsetail movement was either hampered (zygotic meiosis) or abolished (azygotic meiosis) and the pairing of homologous chromosomes was impaired. Accordingly, the allelic recombination rates of the mcp6Delta strain were only 10 40% of the wild-type rates. By contrast, the ectopic recombination rate of the mcp6Delta strain was twice the wild-type rate. This is probably caused by abnormal homologous pairing in mcp6Delta cells because of aberrant horsetail movement. Fluorescent microscopy indicates that SPB components such as Sad1, Kms1 and Spo15 localize normally in mcp6Delta cells. Because Taz1 and Swi6 also localized with Sad1 in mcp6Delta cells, Mcp6 is not required for telomere clustering. In a taz1Delta strain, which does not display telomere clustering, and the dhc1-d3 mutant, which lacks horsetail movement, Mcp6 localized with Sad1 normally. However, we observed abnormal astral microtubule organization in mcp6Delta cells. From these results, we conclude that Mcp6 is necessary for neither SPB organization nor telomere clustering, but is required for proper astral microtubule positioning to maintain horsetail movement. PMID- 15654022 TI - Mapping anterograde and retrograde degeneration after stroke. PMID- 15654023 TI - Driving in Parkinson's disease. PMID- 15654020 TI - Neurite extension in central neurons: a novel role for the receptor tyrosine kinases Ror1 and Ror2. AB - Neurite elongation and branching are key cellular events during brain development as they underlie the formation of a properly wired neuronal network. Here we report that the receptor tyrosine kinases Ror1 and Ror2 modulate the growth of neurites as well as their branching pattern in hippocampal neurons. Upon Ror1 or Ror2 suppression using antisense oligonucleotides or RNA interference (RNAi), neurons extended shorter and less branched minor processes when compared to those in control cells. In addition, Ror-depleted cells elongated longer, albeit less branched, axons than seen in control cells. Conversely, Ror overexpression both in non-neuronal cells and in hippocampal neurons resulted in the enhanced extension of short and highly branched processes. These phenotypes were accompanied by changes in the microtubule-associated proteins MAP1B and MAP2. Taken together, these results support a novel role for Ror receptors as modulators of neurite extension in central neurons. PMID- 15654024 TI - Thrombolysis for central retinal artery occlusion. PMID- 15654025 TI - Mapping the onset and progression of atrophy in familial frontotemporal lobar degeneration. AB - BACKGROUND: Frontotemporal lobar degeneration (FTLD) may be inherited as an autosomal dominant disease. Studying patients "at risk" for developing FTLD can provide insights into the earliest onset and evolution of the disease. METHOD: We carried out approximately annual clinical, MRI, and neuropsychological assessments on an asymptomatic 51 year old "at risk" family member from a family with FTLD associated with ubiquitin-positive and tau-negative inclusion bodies. We used non-linear (fluid) registration of serial MRI to determine areas undergoing significant regional atrophy at different stages of the disease. RESULTS: Over the first 26 months of the study, the patient remained asymptomatic, but subsequently developed progressive speech production difficulties, and latterly severe orofacial dyspraxia, dyscalculia, frontal executive impairment, and limb dyspraxia. Regional atrophy was present prior to the onset of symptoms, and was initially centred on the left dorsolateral prefrontal cortex and the left middle frontal gyrus. Latterly, there was increasing asymmetric left frontal and parietal atrophy. Imaging revealed excess and increasing global atrophy throughout the study. Neuropsychological evaluation revealed mild intellectual impairment prior to the onset of these clinical symptoms; frontal executive and left parietal impairment subsequently emerged, culminating in widespread cognitive impairment. Fluid registered MRI allowed the emerging atrophy patterns to be delineated. CONCLUSION: We have demonstrated the onset and progressive pattern of in vivo atrophy in familial FTLD using fluid registered MRI and correlated this with the clinical features. Fluid registered MRI may be a useful technique in assessing patterns of focal atrophy in vivo and demonstrating the progression of degenerative diseases. PMID- 15654027 TI - Quantitative assessment of driving performance in Parkinson's disease. AB - OBJECTIVES: The primary aim of this study was to determine how Parkinson's disease (PD) affects driving performance. It also examined whether changes in driver safety were related to specific clinical disease markers or an individual's self rating of driving ability. METHODS: The driving performance of 25 patients with idiopathic PD and 21 age matched controls was assessed on a standardised open road route by an occupational therapist and driving instructor, to provide overall safety ratings and specific driving error scores. RESULTS: The drivers with PD were rated as significantly less safe (p<0.05) than controls, and more than half of the drivers with PD would not have passed a state based driving test. The driver safety ratings were more strongly related to disease duration (r = -0.60) than to their on time Unified Parkinson's Disease Rating Scale (r = 0.24). Drivers with PD made significantly more errors than the control group during manoeuvres that involved changing lanes and lane keeping, monitoring their blind spot, reversing, car parking, and traffic light controlled intersections. The driving instructor also had to intervene to avoid an incident significantly more often for drivers with PD than for controls. Interestingly, driver safety ratings were unrelated to an individual's rating of their own driving performance, and this was the case for all participants. CONCLUSIONS: As a group, drivers with PD are less safe to drive than age matched controls. Standard clinical markers cannot reliably predict driver safety. Further studies are required to ascertain whether the identified driving difficulties can be ameliorated. PMID- 15654026 TI - The cerebral correlates of different types of perseveration in the Wisconsin Card Sorting Test. AB - OBJECTIVES: To explore the neural substrates corresponding to the perseverative errors in the Wisconsin Card Sorting Test (WCST). METHODS: The study examined the correlations between the WCST performances and the SPECT measurements of regional cerebral blood flow (rCBF) in subjects with neurodegenerative dementia. Negative non-linear correlations between the rCBF and the two different types of the perseverative errors ("stuck-in-set" and "recurrent" perseverative errors) were calculated on a voxel basis and volume-of-interest basis in the mixed groups of 72 elderly and dementia patients. RESULTS: The stuck-in-set perseverative error was associated with the reduced rCBF in the rostrodorsal prefrontal cortex, whereas the recurrent perseverative error was related to the left parietal activity but not to the prefrontal activity. CONCLUSIONS: These findings augment evidence that the rostrodorsal prefrontal cortex crucially mediates attentional set shifting, and suggest that the stuck-in-set perseverative errors would be a true pathognomonic sign of frontal dysfunction. Moreover, this study shows that the recurrent perseverative errors may not be associated closely with the prefrontal function, suggesting that this error and the stuck-in-set error should be differentially estimated in the WCST. PMID- 15654029 TI - Post-micturitional hypotension in patients with multiple system atrophy. AB - BACKGROUND: Patients with multiple system atrophy (MSA) occasionally have episodes of syncope or pre-syncope after micturition. OBJECTIVE: To clarify the mechanism of these episodes by investigating the haemodynamic changes associated with micturition. METHODS: 25 patients with probable MSA and 16 age matched normal controls were studied. Continuous records of blood pressure and heart rate were made during water cystometry, along with the Valsalva manoeuvre, head up tilt testing, measurement of plasma noradrenaline, and calculation of coefficient of variance of RR intervals. RESULTS: Compared with normal controls, MSA patients had a lower baseline blood pressure, smaller blood pressure and heart rate increases during bladder filling, and an abnormal fall in blood pressure for a longer duration after voiding, resulting in significantly lower blood pressure than at baseline (mean systolic blood pressure reduction -15.2 mm Hg), and hypotension compared with control blood pressure (-29.0 mm Hg). The blood pressure fall was greater in patients with micturition syncope/pre-syncope than in those without. It was also greater in patients with abdominal straining resulting from difficulty in voiding. Other cardiovascular indices did not correlate with the fall in blood pressure. CONCLUSIONS: Hypotension after voiding in MSA patients may result from generalised autonomic dysfunction and abnormal abdominal straining, resulting in micturition syncope. PMID- 15654028 TI - Apomorphine in idiopathic restless legs syndrome: an exploratory study. AB - BACKGROUND: Dopaminergic and opioidergic drugs have been found to be effective in patients with restless legs syndrome (RLS). OBJECTIVES: To test the effect of apomorphine--a combined opioidergic and dopaminergic agonist--and subsequent selective antagonism by naloxone and metoclopramide on subjective and objective symptoms in patients with idiopathic RLS. METHODS: Nine patients with RLS were pretreated with oral domperidone for three days. A modified suggested immobilisation test (SIT) was carried out between 8 pm and 1 am under the following conditions of intravenous drug administration: baseline-apomorphine apomorphine plus naloxone-apomorphine plus metoclopramide. Outcome variables were a visual analogue scale (VAS) of subjective RLS symptoms and EMG documented periodic leg movements while awake (PLMW). RESULTS: Compared with baseline, apomorphine resulted in a rapid and significant improvement in subjective RLS symptoms as measured by VAS (54.5% improvement; p = 0.011), and an almost immediate cessation of PLMW, measured by PLMW index (98.0% improvement; p = 0.012). Neither additional naloxone nor metoclopramide blocked this effect significantly. While given apomorphine with metoclopramide, there was a trend to reappearance of PLMW. CONCLUSIONS: Apomorphine may be an effective treatment for idiopathic RLS. Its effectiveness may reflect both to its dopaminergic and its opioidergic activity, and is not diminished significantly by blocking only one of these pathways. The trend to a worsening of the PLMW index with metoclopramide hints at a primarily dopaminergic effect of apomorphine in idiopathic RLS. PMID- 15654031 TI - Comparison of intra-arterial thrombolysis with conventional treatment in patients with acute central retinal artery occlusion. AB - BACKGROUND: Several case series and a recent meta-analysis indicate that intra arterial thrombolysis (IAT) is effective for the treatment of acute central retinal artery occlusion (CRAO). METHODS: A total of 37 patients with acute monocular blindness because of unilateral thromboembolic CRAO were treated with IAT using urokinase within six hours of the onset of symptoms. Visual outcome was compared with a control group of 19 patients, also seen within six hours, who did not undergo thrombolytic treatment. In both groups some patients were treated by paracentesis and/or acetazolamide. Predictors of visual outcome were evaluated. RESULTS: Visual improvement was more likely with IAT (p = 0.01) as were the chances to regain visual acuity of >0.6 significantly better (p = 0.04): 8/37 patients (22%) regained visual acuity of >0.6 in the IAT group and none (0/19) in the control group. Younger patients were more likely to regain some vision with (p = 0.012) or without IAT (p = 0.026). Three patients had minor treatment related cerebral ischaemic events, two had transient ischaemic attacks and one a minor stroke. There were no haemorrhagic complications. CONCLUSIONS: This series of patients with CRAO demonstrated that IAT enhanced the chances of visual improvement compared with conventional treatment only. Furthermore, younger patients have a better chance to achieve some visual recovery. PMID- 15654030 TI - Ischaemic stroke in young adults: predictors of outcome and recurrence. AB - BACKGROUND: There is limited information about predictors of outcome and recurrence of ischaemic stroke affecting young adults. OBJECTIVE: To assess the predictive value of the presenting characteristics for both outcome and recurrence in young stroke victims. METHODS: Clinical and radiological data for 203 patients aged 16 to 45 years were collected prospectively; they comprised 11% of 1809 consecutive patients with ischaemic stroke. The National Institutes of Health stroke scale (NIHSS), the Bamford criteria, and the trial of ORG 10172 in acute stroke treatment (TOAST) classification were used to define stroke severity, subtype, and aetiology. The clinical outcome of 198 patients (98%) was assessed using the modified Rankin scale (mRS) and categorised as favourable (score 0-1) or unfavourable (score 2-6). RESULTS: Stroke was caused by atherosclerotic large artery disease in 4%, cardioembolism in 24%, small vessel disease in 9%, another determined aetiology in 30%, and undetermined aetiology in 33%. Clinical outcome at three months was favourable in 68%, unfavourable in 29%, and lethal in 3%. Thirteen non-fatal stroke, two fatal strokes, and six transient ischaemic attacks (TIA) occurred during a mean (SD) follow up of 26 (17) months. High NIHSS score, total anterior circulation stroke, and diabetes mellitus were independent predictors of unfavourable outcome or death (p<0.0001, p = 0.011, and p = 0.023). History of TIA predicted stroke recurrence (p = 0.02). CONCLUSIONS: Severe neurological deficits at presentation, total anterior circulation stroke, and diabetes mellitus predict unfavourable outcome. Previous TIA are associated with increased risk of recurrence. PMID- 15654033 TI - Kanner's infantile autism and Asperger's syndrome. PMID- 15654032 TI - Longitudinal thalamic diffusion changes after middle cerebral artery infarcts. AB - BACKGROUND: Cerebral infarcts are responsible for functional alterations and microscopic tissue damage at distance from the ischaemic area. Such remote effects have been involved in stroke recovery. Thalamic hypometabolism is related to motor recovery in middle cerebral artery (MCA) infarcts but little is known concerning the tissue changes underlying these metabolic changes. Diffusion tensor imaging (DTI) is highly sensitive to microstructural tissue alterations and can be used to quantify in vivo the longitudinal microscopic tissue changes occurring in the thalamus after MCA infarcts in humans. METHODS: Nine patients underwent DTI after an isolated MCA infarct. Mean diffusivity (MD), fractional anisotropy (FA), and thalamic region volume were measured from the first week to the sixth month after stroke onset in these patients and in 10 age matched controls. RESULTS: MD significantly increased in the ipsilateral thalamus between the first and the sixth month (0.766 x 10(-3) mm(2)/s first month; 0.792 x 10(-3) mm(2)/s third month; 0.806 x 10(-3) mm(2)/s sixth month). No significant modification of FA was detected. In six patients, the ipsilateral/contralateral index of MD was higher than the upper limit of the 95% CI calculated in 10 age matched controls. An early decrease of MD preceded the increase of ipsilateral thalamic diffusion in one patient at the first week and in two other patients at the first month. CONCLUSION: After MCA infarcts, an increase in diffusion is observed with DTI in the ipsilateral thalamus later than 1 month after the stroke onset. This is presumably because of the progressive loss of neurons and/or glial cells. In some patients, this increase is preceded by a transient decrease in diffusion possibly related to an early swelling of these cells or to microglial activation. Further studies in larger series are needed to assess the clinical correlates of these findings. PMID- 15654035 TI - Sensitivity and specificity of the new international diagnostic criteria for migraine with aura. AB - OBJECTIVES: Since 1998, migraine with aura (MA) has been diagnosed according to the operational diagnostic criteria of the International Headache Society (ICHD 1). Here we present the data underlying the new criteria for MA in the ICHD-2 classification. METHODS: Sensitivity of the new criteria was tested in patients with MA and specificity in patients with reversible non-aura visual disturbances. The diagnoses in both groups of patients were made in a validated semistructured physician-conducted interview. We tested five sets of criteria for sensitivity and specificity comparing with the diagnosis according to the ICHD-1 in 200 patients and the selected set of criteria in 274 additional patients. RESULTS: Four sets of criteria had sensitivity/specificity of 46%/100%, 71%/100%, 62%/95%, and 99%/76%. Sensitivity of the selected set of criteria was 84% (95% CI 79% to 90%) and specificity 97% (95% CI 95% to 99%). According to these criteria at least two of the following should be fulfilled: homonymous visual or unilateral sensory symptoms; at least one aura symptom develops gradually over > or =5 minutes and/or different symptoms occur in succession over > or =5 minutes; each symptom lasts > or =5 and < or =60 minutes. In the additional sample sensitivity of the selected criteria was 90% (95% CI 86% to 94%) and specificity 96% (95% CI 91% to 100%). CONCLUSIONS: The diagnostic criteria for MA selected for ICHD-2 had high sensitivity and specificity. The ICHD-2 criteria are more operational and probably delineate a more homogeneous sample of patients than the ICHD-1. The ICHD-2 for MA is intended equally for research and clinical practice and can be used at different levels of specialisation. PMID- 15654034 TI - Axonal damage accumulates in the progressive phase of multiple sclerosis: three year follow up study. AB - BACKGROUND: Neurofilament phosphoforms (Nf) are principal components of the axoskeleton released during axonal injury. Cerebrospinal fluid (CSF) levels of Nf phosphoforms might be useful surrogate markers for disability in multiple sclerosis (MS), aid in distinguishing clinical subtypes, and provide valuable prognostic information. METHOD: Thirty four patients with MS were included in a three year follow up study along with 318 controls with other non-inflammatory neurological diseases. CSF levels of two Nf heavy chain (NfH) phosphoforms (NfH(SMI35), NfH(SMI34)) were quantified at baseline and three year follow up using new ELISA techniques. Levels of NfH phosphoforms, the degree of phosphorylation (NfH(SMI34):NfH(SMI35) ratio), and changes in NfH levels between baseline and follow up (Delta NfH) were related to the clinical phenotype (RR or SP/PP), to three clinical scales (Kurtzke's EDSS, ambulation index (AI), and nine hole peg test (9HPT)), and to progression of disability. RESULTS: A significantly higher proportion (59%) of patients with SP/PPMS experienced an increase in NfH(SMI35) levels between baseline and follow up compared with those with RRMS (14%, p<0.05). CSF NfH(SMI34) levels at baseline were higher in patients with SP/PP (11 pg/ml) compared with RR (7 pg/ml, p<0.05) and NfH(SMI35) levels were higher at follow up in SP/PP (129 pg/ml) compared with levels below assay sensitivity in RR (p<0.05). NfH(SMI35) correlated with the EDSS (r(s) = 0.54, p<0.01), the AI (r(s) = 0.42, p<0.05), and the 9HPT (r(s) = 0.59, p<0.01) at follow up. CONCLUSION: The increase in NfH during the progressive phase of the disease together with the correlation of NfH(SMI35) with all clinical scales at follow up suggests that cumulative axonal loss is responsible for sustained disability and that high NfH(SMI35) levels are a poor prognostic sign. PMID- 15654036 TI - Gamma knife treatment for refractory cluster headache: prospective open trial. AB - BACKGROUND: Since the initial report of Ford et al in 1998 no further study has evaluated radiosurgery of the trigeminal nerve in chronic cluster headache (CCH). METHODS: We carried out a prospective open trial of neurosurgery and enrolled 10 patients (nine men, one woman; mean age 49.8 years, range 32-77) presenting with severe and drug resistant CCH (mean duration 9 years, range 2-33). The cisternal segment of the nerve was targeted with a single 4 mm collimator (80-85 Gy max). RESULTS: The mean follow up was 13.2 months. No improvement was observed in two patients and three patients had no further attacks. Three patients showed dramatic improvement with a few attacks per month or very few attacks over the last six months. Two patients were pain free for only one and two weeks and their headaches recurred with the same severity as before. Three patients developed paraesthesia with no hypoaesthesia, one developed hypoaesthesia, and one developed deafferentation pain. CONCLUSIONS: The rate and severity of trigeminal nerve injury appeared to be significantly higher than in trigeminal neuralgia, and this study does not support the positive results of the study of Ford et al. We consider the morbidity to be significant for the low rate of pain cessation, making this procedure less attractive even for the more severely affected subgroup of patients. PMID- 15654037 TI - Short latency responses in the averaged electro-oculogram elicited by vibrational impulse stimuli applied to the skull: could they reflect vestibulo-ocular reflex function? AB - OBJECTIVES: To investigate whether vibrational impulse stimuli applied to the skull can be used to evoke the vestibulo-ocular reflex (VOR) and detect vestibular lesions. METHODS: Twenty four patients with unilateral vestibular loss (UVD), five with bilateral vestibular loss, two with ocular palsies, and 10 healthy subjects participated. Vibrations of the skull were induced with head taps and with a single period of 160 Hz tone burst on the inion, vertex, and the mastoids while the patients viewed a distant target. Several patients were also examined while viewing a near target, with eccentric gaze and in tilted postures. Responses were recorded by EOG. RESULTS: Responses occurred between 5 ms and 20 ms and seemed to be compensatory to the second phase of the sine wave of vibration impulse and were greatly diminished/absent in patients with bilateral VD and ocular palsies. The patients with UVD had asymmetrical responses in the vertical EOG with stimuli applied on the inion and vertex, with enhancement of the response amplitude on the side of vestibular loss and/or diminution on the healthy side. The asymmetry ratios between the healthy subjects and patients with UVD, and among patients with UVD were statistically significant. Some gaze and positional influences could be demonstrated consistent with otolithic reflexes. CONCLUSION: If the asymmetric responses to skull vibration in UVD result from passive oscillatory movements of the orbital tissues they may reflect the otolith mediated sustained skew torsion. Conversely, if generated by active eye movements, their likely origin is a phasic VOR. PMID- 15654039 TI - Risk-benefit analysis of the treatment of unruptured intracranial aneurysms. AB - OBJECTIVES: To determine under what circumstances repair of unruptured intracranial aneurysms may be beneficial. METHODS: A life expectancy analysis of patients with unruptured aneurysms with and without repair based on prospective data from the International Study of Unruptured Intracranial Aneurysms (ISUIA). RESULTS: Life years are lost at all ages by repairing anterior circulation aneurysms under 7 mm in diameter in patients with no history of a subarachnoid haemorrhage from another aneurysm (incidental). For all other aneurysms the number of life years saved by repair is dependent on the patient's age at the time when repair is undertaken. Between 2 and 40 years are saved by repairing aneurysms in patients aged 20 years. These benefits fall to 0 when remaining life expectancy falls below 15-35 years, corresponding to the age range of 45-70 years. CONCLUSIONS: Repair of unruptured aneurysms benefits patients harbouring them by improving life expectancy except in certain circumstances. The exceptions are patients with remaining life expectancy less than 15-35 years or aged 45-70 (depending on aneurysm size and location) and patients with aneurysms of the anterior circulation under 7 mm in diameter with no history of a previous subarachnoid haemorrhage. These results are based on the findings of the ISUIA and are dependent on their accuracy. PMID- 15654038 TI - Cerebral amyloid angiopathy in traumatic brain injury: association with apolipoprotein E genotype. AB - OBJECTIVE: In view of the association of the apolipoprotein E (APOE) epsilon 4 allele with poor outcome after traumatic brain injury we determined the frequency of cerebral amyloid angiopathy (CAA) and the extent of haemorrhagic pathology in relation to APOE genotype in an autopsy series of 88 head injured cases. METHODS: Tissue sections from the frontal and temporal lobes were immunostained for amyloid-beta peptide (A beta) and stained for Congo red to identify vascular amyloid pathology. A semiquantitative assessment of contusions, the total contusion index, was used to estimate the severity of the haemorrhagic pathology. APOE genotypes were determined by polymerase chain reaction of genomic DNA extracted from paraffin embedded tissue sections. RESULTS: CAA was present in 7/40 (18%) epsilon 4 carriers compared with 1/48 (2%) non-epsilon 4 carriers (p = 0.021, 95% confidence interval (CI) for difference in proportions with CAA 3% to 29%) with 6/40 (4 with CAA) epsilon 4 carriers being homozygotes. Thus the risk of having CAA for epsilon 4 carriers was 8.4 times that for the non-epsilon 4 carriers. However, there was no clear tendency for patients with CAA to have more severe or more numerous contusions (median contusion index 19 (CAA) v 14.5, p = 0.23, 95% CI for difference in medians -5 to 14). CONCLUSIONS: Presence of CAA in head injured cases was significantly associated with possession of an APOE epsilon 4 allele but not with the severity of contusions. PMID- 15654041 TI - Subthalamic nucleus stimulation in tremor dominant parkinsonian patients with previous thalamic surgery. AB - Before the introduction of high frequency stimulation of the subthalamic nucleus (STN), many disabled tremor dominant parkinsonian patients underwent lesioning or chronic electrical stimulation of the thalamus. We studied the effects of STN stimulation in patients with previous ventral intermediate nucleus (VIM) surgery whose motor state worsened. Fifteen parkinsonian patients were included in this study: nine with unilateral and two with bilateral VIM stimulation, three with unilateral thalamotomy, and one with both unilateral thalamotomy and contralateral VIM stimulation. The clinical evaluation consisted of a formal motor assessment using the Unified Parkinson's Disease Rating Scale (UPDRS) and neuropsychological tests encompassing a 50 point frontal scale, the Mattis Dementia Rating Scale, and the Beck Depression Inventory. The first surgical procedure was performed a mean (SD) of 8 (5) years after the onset of disease. STN implantation was carried out 10 (4) years later, and duration of follow up after beginning STN stimulation was 24 (20) months. The UPDRS motor score, tremor score, difficulties in performance of activities of daily living, and levodopa equivalent daily dose significantly decreased after STN stimulation. Neither axial symptoms nor neuropsychological status significantly worsened after the implantation of the STN electrodes. The parkinsonian motor state is greatly improved by bilateral STN stimulation even in patients with previous thalamic surgery, and STN stimulation is more effective than VIM stimulation in tremor dominant parkinsonian patients. PMID- 15654040 TI - Sudden cardiac death in diabetes mellitus: risk factors in the Rochester diabetic neuropathy study. AB - OBJECTIVES: To determine risk factors for sudden cardiac death and the role of diabetic autonomic neuropathy (DAN) in the Rochester diabetic neuropathy study (RDNS). METHODS: Associations between diabetic and cardiovascular complications, including DAN, and the risk of sudden cardiac death were studied among 462 diabetic patients (151 type 1) enrolled in the RDNS. Medical records, death certificates, and necropsy reports were assessed for causes of sudden cardiac death. RESULTS: 21 cases of sudden cardiac death were identified over 15 years of follow up. In bivariate analysis of risk covariates, the following were significant: ECG 1 (evolving and previous myocardial infarctions): hazard ratio (HR) = 4.4 (95% confidence interval (CI), 1.6 to 12.1), p = 0.004; ECG 2 (bundle branch block or pacing): HR = 8.6 (2.9 to 25.4), p<0.001; ECG 1 or ECG 2: HR = 4.2 (1.3 to 13.4), p = 0.014; and nephropathy stage: HR = 2.1 (1.3 to 3.4), p = 0.002. Adjusting for ECG 1 or ECG 2, autonomic scores, QTc interval, high density lipoprotein (HDL) cholesterol, 24 hour microalbuminuria, and 24 hour total proteinuria were significant. However, adjusting for nephropathy, none of the autonomic indices, QTc interval, HDL cholesterol, microalbuminuria, or total proteinuria was significant. At necropsy, all patients with sudden cardiac death had coronary artery or myocardial disease. CONCLUSIONS: Sudden cardiac death was correlated with atherosclerotic heart disease and nephropathy, and to a lesser degree with DAN and HDL cholesterol. Although DAN is associated with sudden cardiac death, it is unlikely to be its primary cause. PMID- 15654042 TI - Reliability of MIBG myocardial scintigraphy in the diagnosis of Parkinson's disease. AB - AIM: To evaluate the reliability of [(123)I]meta-iodobenzylguanidine (MIBG) myocardial scintigraphy for diagnosing Parkinson's disease (PD). PATIENTS/METHODS: A series of 391 outpatients showing one or more parkinsonian like symptoms was longitudinally followed up for accurate clinical diagnosis. MIBG scintigraphy was performed in the patients and 10 normal controls of similar age. The heart to mediastinum uptake ratio was calculated in each person, and the values were considered abnormal if they were greater than two standard deviations below the control mean. RESULTS: MIBG uptake was decreased in most patients with PD (87.7%), and was seen in all advanced cases with Hohen-Yahr stage III or more; the sensitivity and specificity of scintigraphy for detecting PD were 87.7% and 37.4%, respectively. Surprisingly, over half of the patients without PD (66.5%) also exhibited low uptake, resulting in considerable overlap in the ratios between PD and the other disorders. CONCLUSION: MIBG scintigraphy is a sensitive, but not specific, test for PD. Low MIBG uptake does not necessarily indicate PD, but is essential for diagnosing advanced PD. PMID- 15654043 TI - Multiple pathologies in a patient with a progressive neurodegenerative syndrome. AB - A woman presenting with levodopa responsive Parkinsonism developed rapidly progressive bulbar signs, quadriparesis, and upper and lower motor neurone signs. At necropsy, she was found to have three pathological diagnoses: amyotrophic lateral sclerosis, Parkinson's disease, and abundant tau-positive argyrophilic neuritic pathology, known as argyrophilic grain disease. This case raises the possibility that three distinct neuropathological diagnoses share a common aetiology. PMID- 15654045 TI - Brodmann's cortical maps. PMID- 15654044 TI - Postinfectious vasculopathy with evolution to moyamoya syndrome. AB - BACKGROUND: Parainfectious vascular events are a known complication of bacterial meningitis, typically occurring within two weeks of disease onset. Delayed vascular complications are rare. We present a case of progressive vasculopathy following bacterial meningitis. CASE DESCRIPTION: A 20 year old woman developed progressive vasculopathy after successful treatment of pneumococcal meningitis. Within eight months of her infection, angiography revealed the appearance of moyamoya syndrome. Despite aggressive immunomodulation and anticoagulation, she had multiple strokes. Autopsy confirmed severe narrowing of proximal cerebral vasculature with absence of inflammation or atherosclerosis. CONCLUSIONS: The inflammation and subsequent postinfectious autoimmune response associated with meningitis can lead to a progressive vasculopathy and may represent a pathophysiologic mechanism for the arterial occlusions seen in moyamoya syndrome. PMID- 15654046 TI - Satiety dysfunction in Prader-Willi syndrome demonstrated by fMRI. AB - The neurobiology relating to the insatiable appetite observed in Prader-Willi syndrome (PWS) has not been fully characterised. Two functional magnetic resonance imaging (fMRI) scans were performed on each of three adults with PWS. The scans were carried out pre- and post-treatment with the antiepileptic topiramate, which had little effect on body weight and appetite in these subjects. Subjects fasted overnight and drank a 75 g dextrose solution prior to fMRI scans for measurement of brain activation levels during/after glucose ingestion. Following glucose administration, there was a significant delay in activation at the hypothalamus and other brain regions associated with satiety compared with previous data on obese volunteers. These regions include the insula, ventromedial prefrontal cortex, and nucleus accumbens. Individuals with PWS showed a mean latency of 24 min while in a previous study obese volunteers had shown a latency of 15 min and lean volunteers a latency of 10 min in the hypothalamus. Our results provide evidence towards a satiety dysfunction in the central nervous system of PWS patients. PMID- 15654048 TI - Time course of wallerian degeneration after ischaemic stroke revealed by diffusion tensor imaging. AB - Wallerian degeneration (WD) after ischaemic stroke is a well known phenomenon following a stereotypical time course. Whereas conventional magnetic resonance imaging fails to detect signal intensity changes until four weeks after stroke, diffusion tensor imaging (DTI) reveals changes related to WD only after days. DTI was used to monitor the time course of Wallerian degeneration of the pyramidal tract from the early subacute to the late chronic stage of ischaemic stroke in two patients. A progressive decrease of fractional anisotropy was found along the pyramidal tract in the cerebral peduncle below the primary lesion resulting from progressive changes in the principal diffusivities, as well as a slight increase in the orientationally averaged diffusivity in the chronic phase. These signal changes reflect the progressive disintegration of fibre structures resulting from WD. PMID- 15654047 TI - Diagnostic value of the Rotterdam-CAMCOG in post-stroke dementia. AB - BACKGROUND AND OBJECTIVE: Specific screening tests to detect post-stroke dementia are lacking. We recently reported that an adaptation of the Cambridge Cognitive Examination (CAMCOG), the Rotterdam-CAMCOG, had excellent sensitivity and specificity for detecting post-stroke dementia. In this study, we externally validated the diagnostic accuracy of the R-CAMCOG in a new, representative cohort of stroke patients. METHODS: The R-CAMCOG and an extensive neuropsychological examination were administered, independently of each other, in 121 patients aged 55 and over with a stroke in the preceding three to nine months. The gold standard diagnosis of dementia was based on the results of the extensive neuropsychological examination, clinical presentation, and information from a close relative, as well as DSM-IV criteria. RESULTS: Of the 121 patients, 35 had dementia (29%). The diagnostic accuracy at the pre-specified cut-off point of 33/34 was established through receiver operating characteristic (ROC) analyses (sensitivity 66%, specificity 94%). At a cut-off point of 36/37 sensitivity would be 83% and specificity 78%. CONCLUSION: The R-CAMCOG is a useful screening tool for post-stroke dementia in a clinical setting. PMID- 15654049 TI - Checkerboard fields in multiple sclerosis. PMID- 15654050 TI - Insular lesions, ECG abnormalities, and outcome in acute stroke. AB - It has been suggested that lesions in the insula may result in abnormal electrocardiographic (ECG) findings and increase the risk of sudden death. We investigated if computed tomography (CT) detected insular lesions due to acute stroke were related to ECG abnormalities and mortality at three months. Acute insular lesions were diagnosed in 43/179 patients (left insular = 25; right insular = 17; bilateral = 1) with acute stroke (cerebral infarcts = 62 and intracerebral haemorrhage = 17) based on CT scans from 5-8 days after stroke onset; 12 lead ECGs were recorded on admission and ECG telemetry was done in the first 12-24 hours after admission. Information regarding mortality at three months was obtained. Insular lesions were related to sinus tachycardia with heart rate >120 bpm (p = 0.001), ectopic beats >10% (p = 0.032), and ST elevation (p = 0.011). Right insular lesions were related to atrial fibrillation (p = 0.009), atrioventricular block (p = 0.029), ectopic beats >10% (p = 0.016), and inverted T wave (p = 0.040). Right insular lesions, compared with left or no insular lesions, increased the odds of death within three months (OR 6.2, 95% CI 1.5 to 25.2) independent of stroke severity, lesion volume, and age. As the number of patients in the present study is relatively small, our findings need to be confirmed in studies on other populations of stroke patients. PMID- 15654051 TI - Sex hormones modulate brain damage in multiple sclerosis: MRI evidence. AB - BACKGROUND: Sex related differences in the course and severity of multiple sclerosis (MS) could be mediated by the sex hormones. OBJECTIVE: To investigate the relation between serum sex hormone concentrations and characteristics of tissue damage on conventional magnetic resonance imaging (MRI) in men and women suffering from relapsing-remitting MS. RESULTS: Serum testosterone was significantly lower in women with MS than in controls. The lowest levels were found in women with a greater number of gadolinium enhancing lesions. A positive correlation was observed between testosterone concentrations and both tissue damage on MRI and clinical disability. In men, there was a positive correlation between oestradiol concentrations and brain damage. CONCLUSIONS: The hormone related modulation of pathological changes supports the hypothesis that sex hormones play a role in the inflammation, damage, and repair mechanisms typical of MS. PMID- 15654052 TI - Dominance of ipsilateral corticospinal pathway in congenital mirror movements. AB - OBJECTIVE: To clarify the mechanism of congenital mirror movements. DESIGN: The triple stimulation technique (TST) and the silent period were used to investigate a patient with congenital mirror movements. The TST was used to calculate the ratio of ipsilateral to contralateral corticospinal tracts from the two hemispheres to the spinal motor neurones. RESULTS: Transcranial magnetic stimulation over unilateral M1 induced larger ipsilateral than contralateral motor evoked potentials on both sides. Only 9% of spinal motor neurones innervating the abductor digitorum minimi were excited by contralateral primary motor cortex (M1) stimulation, while 94% were excited by the ipsilateral M1 stimulation. The silent period was examined during mirror movements and with voluntary contraction of the right first dorsal interosseus mimicking mirror movements. Left M1 stimulation (through the crossed corticospinal tract) did not show any difference in silent period between the two conditions, while right M1 stimulation (through the uncrossed tract) caused a longer silent period during mirror movements than during voluntary contractions. CONCLUSIONS: The results suggest that mirror movements may be caused by a strong connection between ipsilateral M1 and the mirror movements conveyed through a dominant ipsilateral corticospinal pathway. PMID- 15654053 TI - Persistence of perilymph fistula mechanism in a completely paretic posterior semicircular canal. AB - The three dimensional eye movements (search coil technique) of a patient with a completely paretic left posterior semicircular canal as a result of a perilymph fistula (PLF) were studied. The patient still exhibited pressure induced nystagmus that obeyed Ewald's first law. This finding cannot be explained by otolith stimulation, but might indicate that PLF mechanisms either persist in canal plugging or act on the ampulla by directly deflecting the cupula. PMID- 15654054 TI - Vertical-torsional oscillations and dissociated bilateral horizontal gaze palsy in a patient with a pontine cavernous angioma. AB - We report the case of a 16 year old girl with vertical-torsional oscillations. She had a 4 year history of bilateral horizontal gaze palsy caused by a cavernous angioma in the medial part of the dorsal pons. She presented with vertical oscillopsia that had worsened during the past 3 months. Unilateral three dimensional eye movements and bilateral horizontal eye movements were recorded using a magnetic search coil method and direct current electro-oculography, respectively. She had vertical-torsional oscillations (average frequency: 3.0 Hz) leaving vertical saccades and pursuits intact. The average amplitudes of the vertical and torsional components were 2.0 degrees and 0.6 degrees , respectively. Her horizontal rapid eye movements were severely impaired; however, her horizontal pursuits and slow phases of vestibulo-ocular reflex were only partially impaired (gain<0.3, oculomotor range<+/-9 degrees ). Convergence and divergence were intact. Lesions involving the medial part of the dorsal pons and bilateral paramedian pontine reticular formation can induce vertical and torsional oscillations without disruption of vertical rapid eye movements. PMID- 15654056 TI - "Dropped head syndrome" in syringomyelia: report of two cases. PMID- 15654055 TI - Spanish family with myalgia and cramps syndrome. AB - A Spanish family is reported with dystrophinopathy of myalgia and cramps syndrome type. There were five affected males and three females, and also six asymptomatic carriers. Muscle biopsy showed a dystrophic pattern, but immunohistochemistry carried out with three anti-dystrophin antibodies was normal. Dystrophin analysis by western blot revealed a dystrophin of reduced quantity and molecular weight. DNA analysis showed a deletion of the dystrophin gene involving exons 45-52. The natural history of this disorder and the large intrafamilial clinical variability are discussed. PMID- 15654057 TI - Vitamin B-12, serum folate, and cognitive change between 11 and 79 years. PMID- 15654058 TI - Stent assisted endovascular thrombolysis of internal carotid artery dissection. PMID- 15654059 TI - Bell's palsy: a study of the treatment advice given by Neurologists. PMID- 15654060 TI - Gender influence on the progression of HTLV-I associated myelopathy/tropical spastic paraparesis. PMID- 15654061 TI - Ondine's curse during pregnancy. PMID- 15654062 TI - Acute aspiration pneumonia due to bulbar palsy: an initial manifestation of posterior fossa convexity meningioma. PMID- 15654063 TI - Plasma VEGF as a marker for the diagnosis and treatment of vasculitic neuropathy. PMID- 15654064 TI - Effect of posture on levels of arousal and awareness in vegetative and minimally conscious state patients: a preliminary investigation. PMID- 15654065 TI - Acquired Chiari 1 malformation and syringomyelia following lumboperitoneal shunting for pseudotumour cerebri. PMID- 15654066 TI - Long-term constant light induces constitutive elevated expression of mPER2 protein in the murine SCN: a molecular basis for Aschoff's rule? AB - Circadian rhythms in behavior, metabolism, and physiology are based upon transcriptional/translational feedback loops involving a core set of clock genes that interact to regulate their own expression. In mammals, the SCN is the site of a master biological clock regulating circadian locomotor rhythms. The products of the clock genes mPer1, mPer2, mCry1, and mCry2 form essential components of both negative and positive elements within the SCN oscillator. The primary aims of this study were to examine clock gene abundance under longterm LL in an attempt to provide molecular correlates of the lengthened tau and daily phase delays described by Aschoff's rule. Wheel-running behavior was recorded from mice maintained in either DD or LL for 50 days. The abundance of the clock genes mPer1, mPer2, mCry1, and mCry2 and their protein products was then examined (every approximately 4 h) within the SCN using in situ hybridization and immunocytochemistry. Under LL conditions, mPer1, mPer2, mCry1, and mCry2 messages remained rhythmic, although the waveform of mCry2 was altered compared to DD. In LL, mPER1, mCRY1, and mCRY2 protein levels were also rhythmic and comparable to the patterns observed in DD. However, mPER2 is elevated and constitutively expressed under LL. Thus, rhythmic expression of these clock genes is not dependent on the rhythmic production of mPER2, and the acute up-regulation of mPer1 and mPer2 described for short (nonparametric) light pulses is not sustained under LL conditions. These findings suggest that mPER2 is important for the generation of phase delays in the molecular clockwork, providing a possible molecular explanation for Aschoff's rule: LL lengthens the circadian period by inhibiting the degeneration of mPER2, and constitutively elevated levels of mPER2 enhance the phase-delaying limb of the molecular oscillator. PMID- 15654067 TI - Co-contribution of IP3R and Ca2+ influx pathways to pacemaker Ca2+ activity in stomach ICC. AB - Intracellular Ca2+ oscillations in interstitial cells of Cajal (ICCs) are thought to be the primary pacemaker activity in the gut. In the present study, the authors prepared small tissues of 100-to 300-microm diameter (cell cluster preparation) from the stomach smooth muscle (including the myenteric plexus) of mice by enzymatic and mechanical treatments. After 2 to 4 days of culture, the intracellular Ca2+ concentration ([Ca2+]i) was measured. In the presence of nifedipine, a dihydropyridine Ca2+ channel antagonist, spontaneous [Ca2+]i oscillations were observed within limited regions showing positive c Kitimmunoreactivity, a maker for ICCs. In the majority of cell cluster preparations with multiple regions of [Ca2+]i oscillations, [Ca2+]i oscillated synchronously in the same phase. A small number of cell clusters (8 of 53) showed multiple regions of [Ca2+]i oscillations synchronized but with a considerable phase shift. Neither tetrodotoxin (250 nM) nor atropine (10 microM) significantly affected [Ca2+]i oscillations in the presence of nifedipine. Low concentrations (40 microM) of Ni2+ had little effect on the spontaneous [Ca2+]i oscillation, but SK&F96365 (40 microM) and Cd2+ (120 microM) terminated it. Applications of either 2-aminoethoxydiphenyl borate (10 microM) or xestosponginC(10 microM) completely and rather rapidly (approximately 2 min) abolished the spontaneous [Ca2+]i oscillations. The results suggest that pacemaker [Ca2+]i oscillations in ICCs are produced by close interaction of intracellular Ca2+ release channels, especially inositol 1,4,5-trisphosphate receptor (InsP3R) and Ca2+ influx pathways, presumably corresponding to store-operated type channels. Reverse transcription polymerase chain reaction examinations revealed expression of TRPC2, 4, and 6, as well as InsP3R1 and 2 in ICCs. PMID- 15654068 TI - Ramelteon (TAK-375) accelerates reentrainment of circadian rhythm after a phase advance of the light-dark cycle in rats. AB - In vivo pharmacological effects of ramelteon (TAK-375), a novel, highly MT1/MT2 selective receptor agonist, were studied in rats to determine ramelteon's ability to reentrain the circadian rhythm after an abrupt phase advance. Experiments were also conducted to assess the potential cognitive side effects of ramelteon and its potential to become a drug of abuse. After an abrupt 8-h phase shift, ramelteon (0.1 and 1 mg/kg, p.o.) and melatonin (10 mg/kg, p.o.) accelerated reentrainment of running wheel activity rhythm to the new lightdark cycle. Ramelteon (3-30 mg/kg, p.o.) and melatonin (10-100 mg/kg, p.o.) did not affect learning or memory in rats tested by the water maze task and the delayed match to position task, although diazepam and triazolam impaired both of the tasks. Neither ramelteon (3-30 mg/kg, p.o.) nor melatonin (10-100 mg/kg, p.o.) demonstrated a rewarding property in the conditioned place-preference test, implying that MT1/MT2 receptor agonists have no abuse potential. In contrast, benzodiazepines and morphine showed rewarding properties in this test. The authors' results suggest that ramelteon may be useful for treatment of circadian rhythm sleep disorders without adverse effects typically associated with benzodiazepine use, such as learning and memory impairment, and drug dependence. PMID- 15654069 TI - Scotopic illumination enhances entrainment of circadian rhythms to lengthening light:dark cycles. AB - Endogenously generated circadian rhythms are synchronized with the environment through phase-resetting actions of light. Starlight and dim moonlight are of insufficient intensity to reset the phase of the clock directly, but recent studies have indicated that dim nocturnal illumination may otherwise substantially alter entrainment to bright lighting regimes. In this article, the authors demonstrate that, compared to total darkness, dim illumination at night (< 0.010 lux) alters the entrainment of male Syrian hamsters to bright-light T cycles, gradually increasing in cycle length (T) from 24 h to 30 h. Only 1 of 18 hamsters exposed to complete darkness at night entrained to cycles of T > 26 h. In the presence of dim nocturnal illumination, however, a majority of hamsters entrained to Ts of 28 h or longer. The presence or absence of a running wheel had only minor effects on entrainment to lengthening light cycles. The results further establish the potent effects of scotopic illumination on circadian entrainment and suggest that naturalistic ambient lighting at night may enhance the plasticity of the circadian pacemaker. PMID- 15654070 TI - Binocular interactions in the entrainment and phase shifting of locomotor activity rhythms in Syrian hamsters. AB - To assess binocular interactions and possible ocular dominance in entrainment of circadian rhythms, Syrian hamsters maintained in LL were subjected for several weeks to schedules of eye occlusion with opaque contact lenses. In separate groups, the opaque lens was inserted into the left or right eye for 12 h at the same clock time each day. The left and right eyes of other groups were alternately occluded for 12 h each day, with initial occlusion of either the left or right eye for different groups. A majority of hamsters entrained their locomotor activity rhythm when 1 eye was occluded for 12 h. The modified visual input imposed by covering 1 eye is sufficient to induce entrainment. Locomotor rhythms of most animals in which the 2 eyes were alternately occluded for 12 h each day phasedelayed onset of activity during the 1st few days of the lensing procedure; activity onset then free ran with tau < 24 h for several weeks until entraining with tau of 24 h regardless of whether the left or right eye was initially occluded. Entrainment eventually occurred when activity onset coincided with occlusion of the eye contralateral to the one that was first lensed. Photic and nonphotic explanations for eventual entrainment of locomotor rhythms are discussed, and evidence for asymmetrical photic input from the 2 eyes to the SCN is considered. PMID- 15654071 TI - Nasal versus temporal illumination of the human retina: effects on core body temperature, melatonin, and circadian phase. AB - The mammalian retina contains both visual and circadian photoreceptors. In humans, nocturnal stimulation of the latter receptors leads to melatonin suppression, which might cause reduced nighttime sleepiness. Melatonin suppression is maximal when the nasal part of the retina is illuminated. Whether circadian phase shifting in humans is due to the same photoreceptors is not known. The authors explore whether phase shifts and melatonin suppression depend on the same retinal area. Twelve healthy subjects participated in a within subjects design and received all of 3 light conditions--1) 10 lux of dim light on the whole retina, 2) 100 lux of ocular light on the nasal part of the retina, and 3) 100 lux of ocular light on the temporal part of the retina--on separate nights in random order. In all 3 conditions, pupils were dilated before and during light exposure. The protocol consisted of an adaptation night followed by a 23-h period of sustained wakefulness, during which a 4-h light pulse was presented at a time when maximal phase delays were expected. Nasal illumination resulted in an immediate suppression of melatonin but had no effect on subjective sleepiness or core body temperature (CBT). Nasal illumination delayed the subsequent melatonin rhythm by 78 min, which is significantly (p= 0.016) more than the delay drift in the dim-light condition (38 min), but had no detectable phase-shifting effect on the CBT rhythm. Temporal illumination suppressed melatonin less than the nasal illumination and had no effect on subjective sleepiness and CBT. Temporal illumination delayed neither the melatonin rhythm nor the CBT rhythm. The data show that the suppression of melatonin does not necessarily result in a reduction of subjective sleepiness and an elevation ofCBT. In addition, 100 lux of bright white light is strong enough to affect the photoreceptors responsible for the suppression of melatonin but not strong enough to have a significant effect on sleepiness and CBT. This may be due to the larger variability of the latter variables. PMID- 15654072 TI - The menstrual cycle and susceptibility to virtual simulation sickness. AB - Virtual simulation sickness (VSS) is a form of visually induced motion sickness that can result from immersion in a virtual environment (VE). As in their susceptibility to the sickness induced by real motion, women have been reported to be more susceptible than men to VSS, yet the reason for this difference is not known. The aim of the current study was to investigate the influence of the menstrual cycle on susceptibility to VSS in 16 naturally cycling women and to compare the responses of this group with control groups consisting of 1) 16 premenopausal women taking a combined monophasic oral contraceptive and 2) 16 men. All female participants were immersed in a nauseogenic VE on days 5, 12, 19, and 26 of their menstrual/pill cycle. These days were chosen because they fall in line with peaks and troughs of ovarian hormone levels. Menstrual cycle phase was confirmed by salivary estradiol and progesterone levels. A 4-week "pseudo-cycle" was assigned to the male participants. Hormone analysis revealed that 9 participants in the experimental group had been tested at the desired phases of their cycle. These participants exhibited a significant increase in susceptibility to VSS on day 12 of their cycle. The hormone analysis also showed that the cycles of the 7 remaining members of the experimental group had not precisely followed the expected pattern, and so these people had been tested on days that did not coincide with peaks and troughs of ovarian hormone levels. No consistent variation in susceptibility was observed over the cycle in these volunteers. In addition, no change in susceptibility was observed over the pill cycle of the oral contraceptive group nor over the pseudo-cycle applied to the male control group. The authors conclude that susceptibility to VSS varies over the menstrual cycle as a consequence of hormonal variation. PMID- 15654073 TI - Entrainment in a model of the mammalian circadian oscillator. AB - To adapt the timing of processes regulated by the circadian clock to seasonally varying photoperiods, the phase relation between the circadian clock and dusk or dawn ("phase of entrainment") must be tightly adjusted. The authors use a mathematical model of the molecular mammalian circadian oscillator to investigate the influence of the free-running period (tau) and the shape of the PRC on the phase of entrainment. They find that a phase-dependent sensitivity ("gating") of light-induced period gene transcription enables a constant phase relation to dusk or dawn under different photoperiods. Depending on the freerunning period tau and on the shaping of the PRC by gating, the model circadian oscillator tracks either light onset or light offset under different photoperiods. The study indicates that the phase of entrainment of oscillating cells can be systematically controlled by regulating both gating and the free-running period tau. PMID- 15654074 TI - MEIS C termini harbor transcriptional activation domains that respond to cell signaling. AB - MEIS proteins form heteromeric DNA-binding complexes with PBX monomers and PBX.HOX heterodimers. We have shown previously that transcriptional activation by PBX.HOX is augmented by either protein kinase A (PKA) or the histone deacetylase inhibitor trichostatin A (TSA). To examine the contribution of MEIS proteins to this response, we used the chromatin immunoprecipitation assay to show that MEIS1 in addition to PBX1, HOXA1, and HOXB1 was recruited to a known PBX.HOX target, the Hoxb1 autoregulatory element following Hoxb1 transcriptional activation in P19 cells. Subsequent to TSA treatment, MEIS1 recruitment lagged behind that of HOX and PBX partners. MEIS1A also enhanced the transcriptional activation of a reporter construct bearing the Hoxb1 autoregulatory element after treatment with TSA. The MEIS1 homeodomain and protein-protein interaction with PBX contributed to this activity. We further mapped TSA-responsive and CREB-binding protein dependent PKA-responsive transactivation domains to the MEIS1A and MEIS1B C termini. Fine mutation of the 56-residue MEIS1A C terminus revealed four discrete regions required for transcriptional activation function. All of the mutations impairing the response to TSA likewise reduced activation by PKA, implying a common mechanistic basis. C-terminal deletion of MEIS1 impaired transactivation without disrupting DNA binding or complex formation with HOX and PBX. Despite sequence similarity to MEIS and a shared ability to form heteromeric complexes with PBX and HOX partners, the PREP1 C terminus does not respond to TSA or PKA. Thus, MEIS C termini possess transcriptional regulatory domains that respond to cell signaling and confer functional differences between MEIS and PREP proteins. PMID- 15654075 TI - Interaction of chromatin-associated Plk1 and Mcm7. AB - Plk1 is a multifunctional protein kinase involved in regulation of mitotic entry, chromosome segregation, centrosome maturation, and mitotic exit. Plk1 is a target of DNA damage checkpoints and aids resumption of the cell cycle during recovery from G2 arrest. The polo-box domain (PBD) of Plk1 interacts with phosphoproteins and localizes Plk1 to some mitotic structures. In a search for proteins that interact with the PBD of Plk1, we identified two of the minichromosome maintenance (MCM) proteins, Mcm2 and Mcm7. Co-immunoprecipitation and immunoblot analysis showed an interaction between full-length Plk1 and all other members of the MCM2-7 protein complex. Endogenous Plk1 co-immunoprecipitates with basal forms of Mcm7 as well as with slower migrating forms of Mcm7, induced in response to DNA damage. The strongest interaction between endogenous Plk1 and Mcm7 was detected in a soluble chromatin fraction. These findings suggest a new function for Plk1 in coordination of DNA replication and mitotic events. PMID- 15654076 TI - Silencing near tRNA genes requires nucleolar localization. AB - Transcription by RNA polymerase II is antagonized by the presence of a nearby tRNA gene in Saccharomyces cerevisiae. To test hypotheses concerning the mechanism of this tRNA gene-mediated (tgm) silencing, the effects of specific gene deletions were determined. The results show that the mechanism of silencing near tRNA genes is fundamentally different from other forms of transcriptional silencing in yeast. Rather, tgm silencing is dependent on the ability to cluster the dispersed tRNA genes in or near the nucleolus, constituting a form of three dimensional gene control. PMID- 15654077 TI - Solution structure of enzyme IIA(Chitobiose) from the N,N'-diacetylchitobiose branch of the Escherichia coli phosphotransferase system. AB - The solution structure of trimeric Escherichia coli enzyme IIA(Chb) (34 kDa), a component of the N,N'-diacetylchitobiose/lactose branch of the phosphotransferase signal transduction system, has been determined by NMR spectroscopy. Backbone residual dipolar couplings were used to provide long range orientational restraints, and long range (|i - j| > or = 5 residues) nuclear Overhauser enhancement restraints were derived exclusively from samples in which at least one subunit was 15N/13C/2H/(Val-Leu-Ile)-methyl-protonated. Each subunit consists of a three-helix bundle. Hydrophobic residues lining helix 3 of each subunit are largely responsible for the formation of a parallel coiled-coil trimer. The active site histidines (His-89 from each subunit) are located in three symmetrically placed deep crevices located at the interface of two adjacent subunits (A and C, C and B, and B and A). Partially shielded from bulk solvent, structural modeling suggests that phosphorylated His-89 is stabilized by electrostatic interactions with the side chains of His-93 from the same subunit and Gln-91 from the adjacent subunit. Comparison with the x-ray structure of Lactobacillus lactis IIA(Lac) reveals some substantial structural differences, particularly in regard to helix 3, which exhibits a 40 degrees kink in IIA(Lac) versus a 7 degrees bend in IIA(Chb). This is associated with the presence of an unusually large (230-angstroms3) buried hydrophobic cavity at the trimer interface in IIA(Lac) that is reduced to only 45 angstroms3) in IIA(Chb). PMID- 15654078 TI - Evolution of mitochondrial oxa proteins from bacterial YidC. Inherited and acquired functions of a conserved protein insertion machinery. AB - Members of the Oxa1/YidC family are involved in the biogenesis of membrane proteins. In bacteria, YidC catalyzes the insertion and assembly of proteins of the inner membrane. Mitochondria of animals, fungi, and plants harbor two distant homologues of YidC, Oxa1 and Cox18/Oxa2. Oxa1 plays a pivotal role in the integration of mitochondrial translation products into the inner membrane of mitochondria. It contains a C-terminal ribosome-binding domain that physically interacts with mitochondrial ribosomes to facilitate the co-translational insertion of nascent membrane proteins. The molecular function of Cox18/Oxa2 is not well understood. Employing a functional complementation approach with mitochondria-targeted versions of YidC we show that YidC is able to functionally replace both Oxa1 and Cox18/Oxa2. However, to integrate mitochondrial translation products into the inner membrane of mitochondria, the ribosome-binding domain of Oxa1 has to be appended onto YidC. On the contrary, the fusion of the ribosome binding domain onto YidC prevents its ability to complement COX18 mutants suggesting an indispensable post-translational activity of Cox18/Oxa2. Our observations suggest that during evolution of mitochondria from their bacterial ancestors the two descendents of YidC functionally segregated to perform two distinct activities, one co-translational and one post-translational. PMID- 15654079 TI - Crystal structure of foot-and-mouth disease virus 3C protease. New insights into catalytic mechanism and cleavage specificity. AB - Foot-and-mouth disease virus (FMDV) causes a widespread and economically devastating disease of domestic livestock. Although FMDV vaccines are available, political and technical problems associated with their use are driving a renewed search for alternative methods of disease control. The viral RNA genome is translated as a single polypeptide precursor that must be cleaved into functional proteins by virally encoded proteases. 10 of the 13 cleavages are performed by the highly conserved 3C protease (3C(pro)), making the enzyme an attractive target for antiviral drugs. We have developed a soluble, recombinant form of FMDV 3C(pro), determined the crystal structure to 1.9-angstroms resolution, and analyzed the cleavage specificity of the enzyme. The structure indicates that FMDV 3C(pro) adopts a chymotrypsin-like fold and possesses a Cys-His-Asp catalytic triad in a similar conformation to the Ser-His-Asp triad conserved in almost all serine proteases. This observation suggests that the dyad-based mechanisms proposed for this class of cysteine proteases need to be reassessed. Peptide cleavage assays revealed that the recognition sequence spans at least four residues either side of the scissile bond (P4-P4') and that FMDV 3C(pro) discriminates only weakly in favor of P1-Gln over P1-Glu, in contrast to other 3C(pro) enzymes that strongly favor P1-Gln. The relaxed specificity may be due to the unexpected absence in FMDV 3C(pro) of an extended beta-ribbon that folds over the substrate binding cleft in other picornavirus 3C(pro) structures. Collectively, these results establish a valuable framework for the development of FMDV 3C(pro) inhibitors. PMID- 15654080 TI - Structural basis for metal ion coordination and the catalytic mechanism of sphingomyelinases D. AB - Sphingomyelinases D (SMases D) from Loxosceles spider venom are the principal toxins responsible for the manifestation of dermonecrosis, intravascular hemolysis, and acute renal failure, which can result in death. These enzymes catalyze the hydrolysis of sphingomyelin, resulting in the formation of ceramide 1-phosphate and choline or the hydrolysis of lysophosphatidyl choline, generating the lipid mediator lysophosphatidic acid. This report represents the first crystal structure of a member of the sphingomyelinase D family from Loxosceles laeta (SMase I), which has been determined at 1.75-angstrom resolution using the "quick cryo-soaking" technique and phases obtained from a single iodine derivative and data collected from a conventional rotating anode x-ray source. SMase I folds as an (alpha/beta)8 barrel, the interfacial and catalytic sites encompass hydrophobic loops and a negatively charged surface. Substrate binding and/or the transition state are stabilized by a Mg2+ ion, which is coordinated by Glu32, Asp34, Asp91, and solvent molecules. In the proposed acid base catalytic mechanism, His12 and His47 play key roles and are supported by a network of hydrogen bonds between Asp34, Asp52, Trp230, Asp233, and Asn252. PMID- 15654081 TI - R1, a novel repressor of the human monoamine oxidase A. AB - Monoamine oxidase catalyzes the oxidative deamination of a number of neurotransmitters. A deficiency in monoamine oxidase A results in aggressive behavior in both humans and mice. Studies on the regulation of monoamine oxidase A gene expression have shown that the Sp1 family is important for monoamine oxidase A expression. To search for novel transcription factors, the sequences of three Sp1 sites in the monoamine oxidase A core promoter were used in the yeast one-hybrid system to screen a human cDNA library. A novel repressor, R1 (RAM2), has been cloned. The R1 cDNA encodes a protein with 454 amino acids and an open reading frame at the 5'-end. The transfection of R1 in a human neuroblastoma cell line, SK-N-BE (2)-C, inhibited the monoamine oxidase A promoter and enzymatic activity. The degree of inhibition of monoamine oxidase A by R1 correlated with the level of R1 protein expression. R1 was also found to repress monoamine oxidase A promoter activity within a natural chromatin environment. A gel-shift assay indicated that the endogenous R1 protein in SK-N-BE (2)-C cells interacted with the R1 binding sequence. R1 also bound directly to the natural monoamine oxidase A promoter in vivo as shown by chromatin immunoprecipitation assay. Immunocytochemical analysis showed that R1 was expressed in both cytosol and nucleus, which suggested a role for R1 in transcriptional regulation. Northern blot analysis revealed the presence of endogenous R1 mRNA in human brain and peripheral tissues. Taken together, this study shows that R1 is a novel repressor that inhibits monoamine oxidase A gene expression. PMID- 15654082 TI - Protein phosphatase 2A activity associated with Golgi membranes during the G2/M phase may regulate phosphorylation of ERK2. AB - The extracellular signal-regulated kinase (ERK) 1 and 2 proteins are mitogen activated protein kinase (MAPK) members that regulate cell proliferation and differentiation. ERK proteins are activated exclusively by MAPK kinase 1 and 2 phosphorylation of threonine and tyrosine residues located within the conserved TXY MAPK activation motif. Although dual phosphorylation of Thr and Tyr residues confers full activation of ERK, in vitro studies suggest that a single phosphorylation on either Thr or Tyr may yield partial ERK activity. Previously, we have demonstrated that phosphorylation of the tyrosine residue (Tyr(P) ERK) may be involved in regulating the Golgi complex structure during the G2 and M phases of the cell cycle (Cha, H., and Shapiro, P. (2001) J. Cell Biol. 153, 1355 1368). In the present study, we examined mechanisms for generating Tyr(P) ERK by determining cell cycle-dependent changes in localized phosphatase activity. Using fractionated nuclei-free cell lysates, we find increased serine/threonine phosphatase activity associated with Golgi-enriched membranes in cells synchronized in the late G2/early M phase as compared with G1 phase cells. The addition of phosphatase inhibitors in combination with immunodepletion assays identified this activity to be related to protein phosphatase 2A (PP2A). The increased activity was accounted for by elevated PP2A association with mitotic Golgi membranes as well as increased catalytic activity after normalization of PP2A protein levels in the phosphatase assays. These data indicate that localized changes in PP2A activity may be involved in regulating proteins involved in Golgi disassembly as cells enter mitosis. PMID- 15654083 TI - Identification of dynamic proteome changes upon ligand activation of Trk receptors using two-dimensional fluorescence difference gel electrophoresis and mass spectrometry. AB - The TrkA and TrkB tyrosine kinases are members of the neurotrophin receptor family and mediate survival, differentiation, growth, and apoptosis of neurons in response to stimulation by their ligands, NGF and BDNF, respectively. Expression levels of TrkA/TrkB are important prognostic factors in a variety of embryonal tumors including neuroblastoma, the most common solid tumor of childhood. Because TrkA/TrkB exhibit a high level of sequence similarity and use overlapping pathways for signal transduction, the existence of specific effector molecules crucial for receptor and cell-type-specific response is likely. To identify these effectors by analyzing biological effects of TrkA and TrkB activation in a defined model, we performed a proteome study using the human neuroblastoma SY5Y cell line stably transfected with the TrkA or TrkB cDNA. The use of the recently introduced DIGE (fluorescence two-dimensional difference gel electrophoresis) system (Amersham Biosciences, Piscataway, NJ) allowed us to monitor differences in protein expression between samples in one gel. Proteomic changes were monitored in a time course of 0, 0.5, 1, 6, and 24 h following receptor activation. Using MALDI mass spectrometry, we identified, respectively, 22 and 9 differentially expressed proteins upon the addition of neurotrophin in SY5Y-TrkB and SY5Y-TrkA cells. Functional assignment revealed that the majority of these proteins are involved in organization and maintenance of cellular structures. PMID- 15654084 TI - Report on financing the new model of family medicine. AB - PURPOSE: To foster redesigning the work and workplaces of family physicians, this Future of Family Medicine task force was created to formulate and recommend a financial model that sustains and promotes a thriving New Model of care by focusing on practice reimbursement and health care finances. The goals of the task force were to develop a financial model that assesses the impact of the New Model on practice finances, and to recommend health care financial policies that, if implemented, would be expected to promote the New Model and the primary medical care function in the United States for the next few decades. METHODS: The members of the task force reflected a wide range of professional backgrounds and expertise. The group met in person on 2 occasions and communicated by e-mail and conference calls to achieve consensus. A marketing study was carried out using focus groups to test the concept of the New Model with consumers. External consultants with expertise in health economics, health care finance, health policy, and practice management were engaged to assist the task force with developing the microeconomic (practice level) and macroeconomic (societal level) financial models necessary to achieve its goals. Model assumptions were derived from the published medical literature, existing practice management databases, and discussions with experienced physicians and other content experts. The results of the financial modeling exercise are included in this report. The initial draft report of the findings and recommendations was shared with a reactor panel representing a broad spectrum of constituencies. Feedback from these individuals was reviewed and incorporated, as appropriate, into the final report. RESULTS: The practice-level financial model suggests that full implementation of the New Model of care within the current fee-for-service system of reimbursement would result in a 26% increase in compensation (from 167,457 dollars to 210,288 dollars total annual compensation) for prototypical family physicians who maintain their current number of work hours. Alternatively, physicians could choose to decrease their work hours by 12% and maintain their current compensation. This result is sensitive to physician practice group size. The societal level financial model shows that modifications in the current reimbursement system could lead to further improvements in compensation for family physicians practicing the New Model of care. Reimbursement for e-visits and chronic disease management could further increase total annual compensation to 229,849 dollars for prototypical family physicians maintaining their current number of work hours. The widespread introduction of quality-based physician incentive bonus payments similar to some current programs that have been implemented on a limited basis could further increase total annual compensation up to 254,500 dollars. The adoption of a mixed reimbursement model, which would add an annual per-patient fee, a chronic care bonus, and an overall performance bonus to the current reimbursement system, could increase total annual compensation for the prototypical family physician continuing the current number of hours worked to as much as 277,800 dollars, a 66% increase above current compensation levels. The cost of transition to the New Model is estimated to range from 23,442 dollars to 90,650 dollars per physician, depending on the assumed magnitude of productivity loss associated with implementing an electronic health record. The financial impact of enhanced use of primary care on the costs of health care in the United States was estimated. If every American used a primary care physician as their usual source of care, health care costs would likely decrease by 5.6%, resulting in national savings of 67 billion dollars per year, with an improvement in the quality of the health care provided. CONCLUSIONS: Family physicians could use New Model efficiency to increase compensation or to reduce work time. There are alternative reimbursement methodologies compatible with the New Model that would allow family physicians to share in the health care cost savings achieved as a result of effective and efficient delivery of care. The New Model of care should enhance health care while propelling the US system toward improved performance and results that are satisfying to patients, health care professionals, purchasers, and payers. The New Model needs to be implemented now. Given the recognized need for improvements in the US health care system in the areas of quality, safety, access and costs, there is no reason to delay. PMID- 15654085 TI - The integration of recombination and physical maps in a large-genome monocot using haploid genome analysis in a trihybrid allium population. AB - Integrated mapping in large-genome monocots has been carried out on a limited number of species. Furthermore, integrated maps are difficult to construct for these species due to, among other reasons, the specific plant populations needed. To fill these gaps, Alliums were chosen as target species and a new strategy for constructing suitable populations was developed. This strategy involves the use of trihybrid genotypes in which only one homeolog of a chromosome pair is recombinant due to interspecific recombination. We used genotypes from a trihybrid Allium cepa x (A. roylei x A. fistulosum) population. Recombinant chromosomes 5 and 8 from the interspecific parent were analyzed using genomic in situ hybridization visualization of recombination points and the physical positions of recombination were integrated into AFLP linkage maps of both chromosomes. The integrated maps showed that in Alliums recombination predominantly occurs in the proximal half of chromosome arms and that 57.9% of PstI/MseI markers are located in close proximity to the centromeric region, suggesting the presence of genes in this region. These findings are different from data obtained on cereals, where recombination rate and gene density tends to be higher in distal regions. PMID- 15654086 TI - Genetic control of temperature preference in the nematode Caenorhabditis elegans. AB - Animals modify behavioral outputs in response to environmental changes. C. elegans exhibits thermotaxis, where well-fed animals show attraction to their cultivation temperature on a thermal gradient without food. We show here that feeding-state-dependent modulation of thermotaxis is a powerful behavioral paradigm for elucidating the mechanism underlying neural plasticity, learning, and memory in higher animals. Starved experience alone could induce aversive response to cultivation temperature. Changing both cultivation temperature and feeding state simultaneously evoked transient attraction to or aversion to the previous cultivation temperature: recultivation of starved animals with food immediately induced attraction to the temperature associated with starvation, although the animals eventually exhibited thermotaxis to the new temperature associated with food. These results suggest that the change in feeding state quickly stimulates the switch between attraction and aversion for the temperature in memory and that the acquisition of new temperature memory establishes more slowly. We isolated aho (abnormal hunger orientation) mutants that are defective in starvation-induced cultivation-temperature avoidance. Some aho mutants responded normally to changes in feeding state with respect to locomotory activity, implying that the primary thermosensation followed by temperature memory formation remains normal and the modulatory aspect of thermotaxis is specifically impaired in these mutants. PMID- 15654088 TI - Insertion/deletion and nucleotide polymorphism data reveal constraints in Drosophila melanogaster introns and intergenic regions. AB - Our study of nucleotide sequence and insertion/deletion polymorphism in Drosophila melanogaster noncoding DNA provides evidence for selective pressures in both intergenic regions and introns (of the large size class). Intronic and intergenic sequences show a similar polymorphic deletion bias. Insertions have smaller sizes and higher frequencies than deletions, supporting the hypothesis that insertions are selected to compensate for the loss of DNA caused by deletion bias. Analysis of a simple model of selective constraints suggests that the blocks of functional elements located in intergenic sequences are on average larger than those in introns, while the length distribution of relatively unconstrained sequences interspaced between these blocks is similar in intronic and intergenic regions. PMID- 15654087 TI - The WD40 repeat protein fritz links cytoskeletal planar polarity to frizzled subcellular localization in the Drosophila epidermis. AB - Much of our understanding of the genetic mechanisms that control planar cell polarity (PCP) in epithelia has derived from studies of the formation of polarized cell hairs during Drosophila wing development. The correct localization of an F-actin prehair to the distal vertex of the pupal wing cell has been shown to be dependent upon the polarized subcellular localization of Frizzled and other core PCP proteins. However, the core PCP proteins do not organize actin cytoskeletal polarity directly but require PCP effector proteins such as Fuzzy and Inturned to mediate this process. Here we describe the characterization of a new PCP effector gene, fritz, that encodes a novel but evolutionarily conserved coiled-coil WD40 protein. We show that the fritz gene product functions cell autonomously downstream of the core PCP proteins to regulate both the location and the number of wing cell prehair initiation sites. PMID- 15654089 TI - The heme activator protein Hap1 represses transcription by a heme-independent mechanism in Saccharomyces cerevisiae. AB - The yeast heme activator protein Hap1 binds to DNA and activates transcription of genes encoding functions required for respiration and for controlling oxidative damage, in response to heme. Hap1 contains a DNA-binding domain with a C6 zinc cluster motif, a coiled-coil dimerization element, typical of the members of the yeast Gal4 family, and an acidic activation domain. The regulation of Hap1 transcription-activating activity is controlled by two classes of Hap1 elements, repression modules (RPM1-3) and heme-responsive motifs (HRM1-7). Previous indirect evidence indicates that Hap1 may repress transcription directly. Here we show, by promoter analysis, by chromatin immunoprecipitation, and by electrophoretic mobility shift assay, that Hap1 binds directly to DNA and represses transcription of its own gene by at least 20-fold. We found that Hap1 repression of the HAP1 gene occurs independently of heme concentrations. While DNA binding is required for transcriptional repression by Hap1, deletion of Hap1 activation domain and heme-regulatory elements has varying effects on repression. Further, we found that repression by Hap1 requires the function of Hsp70 (Ssa), but not Hsp90. These results show that Hap1 binds to its own promoter and represses transcription in a heme-independent but Hsp70-dependent manner. PMID- 15654090 TI - Chromosome loss followed by duplication is the major mechanism of spontaneous mating-type locus homozygosis in Candida albicans. AB - Candida albicans, which is diploid, possesses a single mating-type (MTL) locus on chromosome 5, which is normally heterozygous (a/alpha). To mate, C. albicans must undergo MTL homozygosis to a/a or alpha/alpha. Three possible mechanisms may be used in this process, mitotic recombination, gene conversion, or loss of one chromosome 5 homolog, followed by duplication of the retained homolog. To distinguish among these mechanisms, 16 spontaneous a/a and alpha/alpha derivatives were cloned from four natural a/alpha strains, P37037, P37039, P75063, and P34048, grown on nutrient agar. Eighteen polymorphic (heterozygous) markers were identified on chromosome 5, 6 to the left and 12 to the right of the MTL locus. These markers were then analyzed in MTL-homozygous derivatives of the four natural a/alpha strains to distinguish among the three mechanisms of homozygosis. An analysis of polymorphisms on chromosomes 1, 2, and R excluded meiosis as a mechanism of MTL homozygosis. The results demonstrate that while mitotic recombination was the mechanism for homozygosis in one offspring, loss of one chromosome 5 homolog followed by duplication of the retained homolog was the mechanism in the remaining 15 offspring, indicating that the latter mechanism is the most common in the spontaneous generation of MTL homozygotes in natural strains of C. albicans in culture. PMID- 15654091 TI - Detecting amino acid sites under positive selection and purifying selection. AB - An excess of nonsynonymous over synonymous substitution at individual amino acid sites is an important indicator that positive selection has affected the evolution of a protein between the extant sequences under study and their most recent common ancestor. Several methods exist to detect the presence, and sometimes location, of positively selected sites in alignments of protein-coding sequences. This article describes the "sitewise likelihood-ratio" (SLR) method for detecting nonneutral evolution, a statistical test that can identify sites that are unusually conserved as well as those that are unusually variable. We show that the SLR method can be more powerful than currently published methods for detecting the location of positive selection, especially in difficult cases where the strength of selection is low. The increase in power is achieved while relaxing assumptions about how the strength of selection varies over sites and without elevated rates of false-positive results that have been reported with some other methods. We also show that the SLR method performs well even under circumstances where the results from some previous methods can be misleading. PMID- 15654092 TI - Mapping quantitative trait loci in multiple populations of Arabidopsis thaliana identifies natural allelic variation for trichome density. AB - The majority of biological traits are genetically complex. Mapping the quantitative trait loci (QTL) that determine these phenotypes is a powerful means for estimating many parameters of the genetic architecture for a trait and potentially identifying the genes responsible for natural variation. Typically, such experiments are conducted in a single mapping population and, therefore, have only the potential to reveal genomic regions that are polymorphic between the progenitors of the population. What remains unclear is how well the QTL identified in any one mapping experiment characterize the genetics that underlie natural variation in traits. Here we provide QTL mapping data for trichome density from four recombinant inbred mapping populations of Arabidopsis thaliana. By aligning the linkage maps for these four populations onto a common physical map, the results from each experiment were directly compared. Seven of the nine QTL identified are population specific while two were mapped in all four populations. Our results show that many lineage-specific alleles that either increase or decrease trichome density persist in natural populations and that most of this genetic variation is additive. More generally, these findings suggest that the use of multiple populations holds great promise for better understanding the genetic architecture of natural variation. PMID- 15654093 TI - Characterization of Mos1-mediated mutagenesis in Caenorhabditis elegans: a method for the rapid identification of mutated genes. AB - Insertional mutagenesis with a heterologous transposon provides a method to rapidly determine the molecular identity of mutated genes. The Drosophila transposon Mos1 can be mobilized to cause mutations in Caenorhabditis elegans (Bessereau et al. 2001); however, the mutagenic rate was initially too low for use in most forward genetic screens. To increase the effectiveness of Mos1 mediated mutagenesis we examined the conditions influencing Mos1 transposition. First, optimal transposition occurs 24 hr after expression of the transposase and is unlikely to occur in differentiated sperm or oocytes. Second, transposition is limited to germ-cell nuclei that contain donor elements, but the transposase enzyme can diffuse throughout the gonad syncytium. Third, silencing of transposition is caused by changes in the donor array that occur over time. Finally, multiple transposition events occur in individual germ cells. By using screening techniques based on these results, Mos1 mutagenicity was increased to within an order of magnitude of chemical mutagens. PMID- 15654095 TI - Rapid subfunctionalization accompanied by prolonged and substantial neofunctionalization in duplicate gene evolution. AB - Gene duplication is the primary source of new genes. Duplicate genes that are stably preserved in genomes usually have divergent functions. The general rules governing the functional divergence, however, are not well understood and are controversial. The neofunctionalization (NF) hypothesis asserts that after duplication one daughter gene retains the ancestral function while the other acquires new functions. In contrast, the subfunctionalization (SF) hypothesis argues that duplicate genes experience degenerate mutations that reduce their joint levels and patterns of activity to that of the single ancestral gene. We here show that neither NF nor SF alone adequately explains the genome-wide patterns of yeast protein interaction and human gene expression for duplicate genes. Instead, our analysis reveals rapid SF, accompanied by prolonged and substantial NF in a large proportion of duplicate genes, suggesting a new model termed subneofunctionalization (SNF). Our results demonstrate that enormous numbers of new functions have originated via gene duplication. PMID- 15654094 TI - A novel recombination pathway initiated by the Mre11/Rad50/Nbs1 complex eliminates palindromes during meiosis in Schizosaccharomyces pombe. AB - DNA palindromes are rare in humans but are associated with meiosis-specific translocations. The conserved Mre11/Rad50/Nbs1 (MRN) complex is likely directly involved in processing palindromes through the homologous recombination pathway of DNA repair. Using the fission yeast Schizosaccharomyces pombe as a model system, we show that a 160-bp palindrome (M-pal) is a meiotic recombination hotspot and is preferentially eliminated by gene conversion. Importantly, this hotspot depends on the MRN complex for full activity and reveals a new pathway for generating meiotic DNA double-strand breaks (DSBs), separately from the Rec12 (ortholog of Spo11) pathway. We show that MRN-dependent DSBs are formed at or near the M-pal in vivo, and in contrast to the Rec12-dependent breaks, they appear early, during premeiotic replication. Analysis of mrn mutants indicates that the early DSBs are generated by the MRN nuclease activity, demonstrating the previously hypothesized MRN-dependent breakage of hairpins during replication. Our studies provide a genetic and physical basis for frequent translocations between palindromes in human meiosis and identify a conserved meiotic process that constantly selects against palindromes in eukaryotic genomes. PMID- 15654096 TI - A genetic screen for top3 suppressors in Saccharomyces cerevisiae identifies SHU1, SHU2, PSY3 and CSM2: four genes involved in error-free DNA repair. AB - Helicases of the RecQ family and topoisomerase III are evolutionarily conserved proteins important for maintenance of genome stability. In Saccharomyces cerevisiae, loss of the TOP3 gene, encoding topoisomerase III, results in a phenotype of slow growth, DNA damage sensitivity, meiotic defects, and hyperrecombination. The sole RecQ helicase in budding yeast, Sgs1, interacts with Top3 both physically and genetically, and the two proteins are thought to act in concert in vivo. Much recent genetic and biochemical evidence points to the role of RecQ helicases and topoisomerase III in regulating homologous recombination (HR) during DNA replication. Previously, we found that mutations in HR genes partially suppress top3 slow growth. Here, we describe the analysis of four additional mutational suppressors of top3 defects: shu1, shu2, psy3, and csm2. These genes belong to one epistasis group and their protein products interact with each other, strongly suggesting that they function as a complex in vivo. Their mutant phenotype indicates that they are important for error-free repair of spontaneous and induced DNA lesions, protecting the genome from mutation. These mutants exhibit an epistatic relationship with rad52 and show altered dynamics of Rad52-YFP foci, suggesting a role for these proteins in recombinational repair. PMID- 15654097 TI - Seizure suppression by gain-of-function escargot mutations. AB - Suppressor mutations provide potentially powerful tools for examining mechanisms underlying neurological disorders and identifying novel targets for pharmacological intervention. Here we describe mutations that suppress seizures in a Drosophila model of human epilepsy. A screen utilizing the Drosophila easily shocked (eas) "epilepsy" mutant identified dominant suppressors of seizure sensitivity. Among several mutations identified, neuronal escargot (esg) reduced eas seizures almost 90%. The esg gene encodes a member of the snail family of transcription factors. Whereas esg is normally expressed in a limited number of neurons during a defined period of nervous system development, here normal esg was expressed in all neurons and throughout development. This greatly ameliorated both the electrophysiological and the behavioral epilepsy phenotypes of eas. Neuronal esg appears to act as a general seizure suppressor in the Drosophila epilepsy model as it reduces the susceptibility of several seizure-prone mutants. We observed that esg must be ectopically expressed during nervous system development to reduce seizure susceptibility in adults. Furthermore, induction of esg in a small subset of neurons (interneurons) will reduce seizure susceptibility. A combination of microarray and computational analyses revealed 100 genes that represent possible targets of neuronal esg. We anticipate that some of these genes may ultimately serve as targets for novel antiepileptic drugs. PMID- 15654098 TI - Infrequent co-conversion of markers flanking a meiotic recombination initiation site in Saccharomyces cerevisiae. AB - To study the mechanism of meiotic recombination in Saccharomyces cerevisiae, we examined recombination in an interval where the majority of events are initiated at a single hotspot for DNA double-strand breaks (DSBs), with little or no expected contribution by outside initiation events. This interval contained infrequently corrected palindromic markers 300 bp to the left and 600 bp to the right of the DSB hotspot. Conversion of single markers occurred frequently, while conversion of both markers occurred rarely, and many of the tetrads in which both markers converted were the products of multiple events. These data indicate that most meiotic recombination intermediates are asymmetrically positioned around the initiating DSB, with a short (<300 bp) tract of heteroduplex DNA (hDNA) to one side and hDNA on the other side frequently extending 600 bp or more. One consequence of this asymmetry is the preferential concentration of crossovers in the vicinity of the initiating DSB. PMID- 15654099 TI - Bayesian analysis of an admixture model with mutations and arbitrarily linked markers. AB - We introduce here a Bayesian analysis of a classical admixture model in which all parameters are simultaneously estimated. Our approach follows the approximate Bayesian computation (ABC) framework, relying on massive simulations and a rejection-regression algorithm. Although computationally intensive, this approach can easily deal with complex mutation models and partially linked loci, and it can be thoroughly validated without much additional computation cost. Compared to a recent maximum-likelihood (ML) method, the ABC approach leads to similarly accurate estimates of admixture proportions in the case of recent admixture events, but it is found superior when the admixture is more ancient. All other parameters of the admixture model such as the divergence time between parental populations, the admixture time, and the population sizes are also well estimated, unlike the ML method. The use of partially linked markers does not introduce any particular bias in the estimation of admixture, but ML confidence intervals are found too narrow if linkage is not specifically accounted for. The application of our method to an artificially admixed domestic bee population from northwest Italy suggests that the admixture occurred in the last 10-40 generations and that the parental Apis mellifera and A. ligustica populations were completely separated since the last glacial maximum. PMID- 15654100 TI - Systematic, RNA-interference-mediated identification of mus-101 modifier genes in Caenorhabditis elegans. AB - The Mus101 family of chromosomal proteins, identified initially in Drosophila, is widely conserved and has been shown to function in a variety of DNA metabolic processes. Such functions include DNA replication, DNA damage repair, postreplication repair, damage checkpoint activation, chromosome stability, and chromosome condensation. Despite its conservation and widespread involvement in chromosome biogenesis, very little is known about how Mus101 is regulated and what other proteins are required for Mus101 to exert its functions. To learn more about Mus101, we have initiated an analysis of the protein in C. elegans. Here, we show that C. elegans mus-101 is an essential gene, that it is required for DNA replication, and that it also plays an important role in the DNA damage response. Furthermore, we use RNA interference (RNAi)-mediated reverse genetics to screen for genes that modify a mus-101 partial loss-of-function RNAi phenotype. Using a systematic approach toward modifier gene discovery, we have found five chromosome I genes that modify the mus-101 RNAi phenotype, and we go on to show that one of them encodes an E3 SUMO ligase that promotes SUMO modification of MUS-101 in vitro. These results expand our understanding of MUS-101 regulation and show that genetic interactions can be uncovered using screening strategies that rely solely on RNAi. PMID- 15654102 TI - Variogram analysis of the spatial genetic structure of continuous populations using multilocus microsatellite data. AB - A geostatistical perspective on spatial genetic structure may explain methodological issues of quantifying spatial genetic structure and suggest new approaches to addressing them. We use a variogram approach to (i) derive a spatial partitioning of molecular variance, gene diversity, and genotypic diversity for microsatellite data under the infinite allele model (IAM) and the stepwise mutation model (SMM), (ii) develop a weighting of sampling units to reflect ploidy levels or multiple sampling of genets, and (iii) show how variograms summarize the spatial genetic structure within a population under isolation-by-distance. The methods are illustrated with data from a population of the epiphytic lichen Lobaria pulmonaria, using six microsatellite markers. Variogram-based analysis not only avoids bias due to the underestimation of population variance in the presence of spatial autocorrelation, but also provides estimates of population genetic diversity and the degree and extent of spatial genetic structure accounting for autocorrelation. PMID- 15654101 TI - Meiosis-specific regulation of the Saccharomyces cerevisiae S-phase cyclin CLB5 is dependent on MluI cell cycle box (MCB) elements in its promoter but is independent of MCB-binding factor activity. AB - In proliferating S. cerevisiae, genes whose products function in DNA replication are regulated by the MBF transcription factor composed of Mbp1 and Swi6 that binds to consensus MCB sequences in target promoters. We find that during meiotic development a subset of DNA replication genes exemplified by TMP1 and RNR1 are regulated by Mbp1. Deletion of Mbp1 deregulated TMP1 and RNR1 but did not interfere with premeiotic S-phase, meiotic recombination, or spore formation. Surprisingly, deletion of MBP1 had no effect on the expression of CLB5, which is purportedly controlled by MBF. Extensive analysis of the CLB5 promoter revealed that the gene is largely regulated by elements within a 100-bp fragment containing a cluster of MCB sequences. Surprisingly, induction of the CLB5 promoter requires MCB sequences, but not Mbp1, implying that another MCB-binding factor may exist in cells undergoing meiosis. In addition, full activation of CLB5 during meiosis requires Clb5 activity, suggesting that CLB5 may be regulated by a positive feedback mechanism. We further demonstrate that during meiosis MCBs function as effective transcriptional activators independent of MBP1. PMID- 15654103 TI - Statistical tests of the coalescent model based on the haplotype frequency distribution and the number of segregating sites. AB - Several tests of neutral evolution employ the observed number of segregating sites and properties of the haplotype frequency distribution as summary statistics and use simulations to obtain rejection probabilities. Here we develop a "haplotype configuration test" of neutrality (HCT) based on the full haplotype frequency distribution. To enable exact computation of rejection probabilities for small samples, we derive a recursion under the standard coalescent model for the joint distribution of the haplotype frequencies and the number of segregating sites. For larger samples, we consider simulation-based approaches. The utility of the HCT is demonstrated in simulations of alternative models and in application to data from Drosophila melanogaster. PMID- 15654104 TI - Gene clusters for insecticidal loline alkaloids in the grass-endophytic fungus Neotyphodium uncinatum. AB - Loline alkaloids are produced by mutualistic fungi symbiotic with grasses, and they protect the host plants from insects. Here we identify in the fungal symbiont, Neotyphodium uncinatum, two homologous gene clusters (LOL-1 and LOL-2) associated with loline-alkaloid production. Nine genes were identified in a 25-kb region of LOL-1 and designated (in order) lolF-1, lolC-1, lolD-1, lolO-1, lolA-1, lolU-1, lolP-1, lolT-1, and lolE-1. LOL-2 contained the homologs lolC-2 through lolE-2 in the same order and orientation. Also identified was lolF-2, but its possible linkage with either cluster was undetermined. Most lol genes were regulated in N. uncinatum and N. coenophialum, and all were expressed concomitantly with loline-alkaloid biosynthesis. A lolC-2 RNA-interference (RNAi) construct was introduced into N. uncinatum, and in two independent transformants, RNAi significantly decreased lolC expression (P < 0.01) and loline-alkaloid accumulation in culture (P < 0.001) compared to vector-only controls, indicating involvement of lolC in biosynthesis of lolines. The predicted LolU protein has a DNA-binding site signature, and the relationships of other lol-gene products indicate that the pathway has evolved from various different primary and secondary biosynthesis pathways. PMID- 15654105 TI - Modeling quantitative trait Loci and interpretation of models. AB - A quantitative genetic model relates the genotypic value of an individual to the alleles at the loci that contribute to the variation in a population in terms of additive, dominance, and epistatic effects. This partition of genetic effects is related to the partition of genetic variance. A number of models have been proposed to describe this relationship: some are based on the orthogonal partition of genetic variance in an equilibrium population. We compare a few representative models and discuss their utility and potential problems for analyzing quantitative trait loci (QTL) in a segregating population. An orthogonal model implies that estimates of the genetic effects are consistent in a full or reduced model in an equilibrium population and are directly related to the partition of the genetic variance in the population. Linkage disequilibrium does not affect the estimation of genetic effects in a full model, but would in a reduced model. Certainly linkage disequilibrium would complicate the detection of QTL and epistasis. Using different models does not influence the detection of QTL and epistasis. However, it does influence the estimation and interpretation of genetic effects. PMID- 15654106 TI - Genetic structure and diversity in Oryza sativa L. AB - The population structure of domesticated species is influenced by the natural history of the populations of predomesticated ancestors, as well as by the breeding system and complexity of the breeding practices exercised by humans. Within Oryza sativa, there is an ancient and well-established divergence between the two major subspecies, indica and japonica, but finer levels of genetic structure are suggested by the breeding history. In this study, a sample of 234 accessions of rice was genotyped at 169 nuclear SSRs and two chloroplast loci. The data were analyzed to resolve the genetic structure and to interpret the evolutionary relationships between groups. Five distinct groups were detected, corresponding to indica, aus, aromatic, temperate japonica, and tropical japonica rices. Nuclear and chloroplast data support a closer evolutionary relationship between the indica and the aus and among the tropical japonica, temperate japonica, and aromatic groups. Group differences can be explained through contrasting demographic histories. With the availability of rice genome sequence, coupled with a large collection of publicly available genetic resources, it is of interest to develop a population-based framework for the molecular analysis of diversity in O. sativa. PMID- 15654107 TI - The genetic basis for inflorescence variation between foxtail and green millet (poaceae). AB - Grass species differ in many aspects of inflorescence architecture, but in most cases the genetic basis of the morphological difference is unknown. To investigate the genes underlying the morphology in one such instance, we undertook a developmental and QTL analysis of inflorescence differences between the cereal grain foxtail millet and its presumed progenitor green millet. Inflorescence differences between these two species are the result of changes in primary branch number and density, spikelet number, and bristle (sterile branchlet) number; these differences also account for inflorescence variation within the clade of 300+ species that share the presence of bristles in the inflorescence. Fourteen replicated QTL were detected for the four inflorescence traits, and these are suggested to represent genes that control differences between the species. Comparative mapping using common markers from rice and maize allowed a number of candidate genes from maize to be localized to QTL regions in the millet genome. Searches of regions of the sequenced rice genome orthologous to QTL regions on foxtail millet identified a number of transcription factors and hormone pathway genes that may be involved in control of inflorescence branching. PMID- 15654108 TI - Linkage disequilibrium and recent selection at three immunity receptor loci in Drosophila simulans. AB - Immune system genes in a California population sample of Drosophila simulans were shown to bear several hallmarks of the effects of past directional selection. One potential effect of directional selection is an increase in linkage disequilibrium among the polymorphic sites that are linked to the site under selection. In this study, we focus on three D. simulans immunity loci, Hmu, Sr CI/Sr-CIII, and Tehao, for which the polymorphic sites are in nearly perfect linkage disequilibrium, an unusual finding even with respect to other immunity genes sampled from the same lines. The most likely explanation for this finding is that, at each locus, two divergent alleles have been selected to intermediate frequencies in the recent past. The extent to which the linkage disequilibrium extends to the flanks of each of the immunity genes is minimal, suggesting that the favored mutations actually occurred within the immunity genes themselves. Furthermore, the excess linkage disequilibrium found in the California population is not found in an African D. simulans population sample and may be a result of novel pathogen-mediated selection pressures encountered during establishment of non-African populations. PMID- 15654109 TI - Sex-linked differentiation between incipient species of Anopheles gambiae. AB - Emerging species within the primary malaria vector Anopheles gambiae show different ecological preferences and significant prezygotic reproductive isolation. They are defined by fixed sequence differences in X-linked rDNA, but most previous studies have failed to detect large and significant differentiation between these taxa elsewhere in the genome, except at two other loci on the X chromosome near the rDNA locus. Hypothesizing that this pericentromeric region of the X chromosome may be accumulating differences faster than other regions of the genome, we explored the pattern and extent of differentiation between A. gambiae incipient species and a sibling species, A. arabiensis, from Burkina Faso, West Africa, at 17 microsatellite loci spanning the X chromosome. Interspecific differentiation was large and significant across the entire X chromosome. Among A. gambiae incipient species, we found some of the highest levels of differentiation recorded in a large region including eight independent loci near the centromere of the X chromosome. Outside of this region, no significant differentiation was detected. This pattern suggests that selection is playing a role in the emergence of A. gambiae incipient species. This process, associated with efficient exploitation of anthropogenic modifications to the environment, has public health implications as it fosters the spread of malaria transmission both spatially and temporally. PMID- 15654110 TI - Combining data from multiple inbred line crosses improves the power and resolution of quantitative trait loci mapping. AB - Rodent inbred line crosses are widely used to map genetic loci associated with complex traits. This approach has proven to be powerful for detecting quantitative trait loci (QTL); however, the resolution of QTL locations, typically approximately 20 cM, means that hundreds of genes are implicated as potential candidates. We describe analytical methods based on linear models to combine information available in two or more inbred line crosses. Our strategy is motivated by the hypothesis that common inbred strains of the laboratory mouse are derived from a limited ancestral gene pool and thus QTL detected in multiple crosses are likely to represent shared ancestral polymorphisms. We demonstrate that the combined-cross analysis can improve the power to detect weak QTL, can narrow support intervals for QTL regions, and can be used to separate multiple QTL that colocalize by chance. Moreover, combined-cross analysis can establish the allelic states of a QTL among a set of parental lines, thus providing critical information for narrowing QTL regions by haplotype analysis. PMID- 15654111 TI - A multilocus sequence survey in Arabidopsis thaliana reveals a genome-wide departure from a neutral model of DNA sequence polymorphism. AB - The simultaneous analysis of multiple genomic loci is a powerful approach to studying the effects of population history and natural selection on patterns of genetic variation of a species. By surveying nucleotide sequence polymorphism at 334 randomly distributed genomic regions in 12 accessions of Arabidopsis thaliana, we examined whether a standard neutral model of nucleotide sequence polymorphism is consistent with observed data. The average nucleotide diversity was 0.0071 for total sites and 0.0083 for silent sites. Although levels of diversity are variable among loci, no correlation with local recombination rate was observed, but polymorphism levels were correlated for physically linked loci (<250 kb). We found that observed distributions of Tajima's D- and D/D(min)- and of Fu and Li's D-, D*- and F-, F*-statistics differed significantly from the expected distributions under a standard neutral model due to an excess of rare polymorphisms and high variances. Observed and expected distributions of Fay and Wu's H were not different, suggesting that demographic processes and not selection at multiple loci are responsible for the deviation from a neutral model. Maximum-likelihood comparisons of alternative demographic models like logistic population growth, glacial refugia, or past bottlenecks did not produce parameter estimates that were more consistent with observed patterns. However, exclusion of highly polymorphic "outlier loci" resulted in a fit to the logistic growth model. Various tests of neutrality revealed a set of candidate loci that may evolve under selection. PMID- 15654113 TI - Trans events associated with crossovers are revealed in the absence of mismatch repair genes in Saccharomyces cerevisiae. AB - Genetic analysis of recombination in Saccharomyces cerevisiae has revealed products with structures not predicted by the double-strand break repair model of meiotic recombination. A particular type of recombinant containing trans heteroduplex DNA has been observed at two loci. Trans events were originally identified only in tetrads in which the non-Mendelian segregations were not associated with a crossover. Because of this, these events were proposed to have arisen from the unwinding of double Holliday junctions. Previous studies used palindromes, refractory to mismatch repair, as genetic markers whereas we have used a complementary approach of deleting mismatch repair proteins to identify heteroduplex DNA. We found that the markers occurred in trans and were associated with crossovers. In both mlh1Delta and msh2Delta strains, the frequency of trans events associated with a crossover exceeded that predicted from the random association of crossovers with noncrossover trans events. We propose two different models to account for trans events associated with crossovers and discuss the relevance to wild-type DSB repair. PMID- 15654112 TI - Quantitative genetics of vector competence for La Crosse virus and body size in Ochlerotatus hendersoni and Ochlerotatus triseriatus interspecific hybrids. AB - La Crosse virus is a leading cause of pediatric encephalitis in the United States. The mosquito Ochlerotatus triseriatus is an efficient vector for La Crosse virus, whereas the closely related O. hendersoni transmits only at very low rates. Quantitative trait loci (QTL) affecting the ability to orally transmit this virus and adult body size were identified in 164 F(2) female individuals from interspecific crosses of O. hendersoni females and O. triseriatus males using a combination of composite interval mapping (CIM), interval mapping (IM) for binary traits, and single-marker mapping. For oral transmission (OT), no genome locations exceeded the 95% experimentwise threshold for declaring a QTL using IM, but single-marker analysis identified four independent regions significantly associated with OT that we considered as tentative QTL. With two QTL, an increase in OT was associated with alleles from the refractory vector, O. hendersoni, and likely reflect epistatic interactions between genes that were uncovered by our interspecific crosses. For body size, two QTL were identified using CIM and a third tentative QTL was identified using single-marker analysis. The genome regions associated with body size also contain three QTL controlling OT, suggesting that these regions contain either single genes with pleiotropic effects or multiple linked genes independently determining each trait. PMID- 15654114 TI - MLH1 and MSH2 promote the symmetry of double-strand break repair events at the HIS4 hotspot in Saccharomyces cerevisiae. AB - Double-strand breaks (DSBs) initiate meiotic recombination. The DSB repair model predicts that both genetic markers spanning the DSB should be included in heteroduplex DNA and be detectable as non-Mendelian segregations (NMS). In experiments testing this, a significant fraction of events do not conform to this prediction, as only one of the markers displays NMS (one-sided events). Two explanations have been proposed to account for the discrepancies between the predictions and experimental observations. One suggests that two-sided events are the norm but are "hidden" as heteroduplex repair frequently restores the parental configuration of one of the markers. Another explanation posits that one-sided events reflect events in which heteroduplex is formed predominantly on only one side of the DSB. In the absence of heteroduplex repair, the first model predicts that two-sided events would be revealed at the expense of one-sided events, while the second predicts no effect on the distribution of events when heteroduplex repair is lost. We tested these predictions by deleting the DNA mismatch repair genes MSH2 or MLH1 and analyzing the proportion of two-sided events. Unexpectedly, the results do not match the predictions of either model. In both mlh1Delta and msh2Delta, the proportion of two-sided events is significantly decreased relative to wild type. These observations can be explained in one of two ways. Either Msh2p/Mlh1p-independent mispair removal leads to restoration of one of the markers flanking the DSB site or Msh2p/Mlh1p actively promote two sided events. PMID- 15654115 TI - Segregation distortion in hybrids between the Bogota and USA subspecies of Drosophila pseudoobscura. AB - We show that, contrary to claims in the literature, "sterile" males resulting from the cross of the Bogota and USA subspecies of Drosophila pseudoobscura are weakly fertile. Surprisingly, these hybrid males produce almost all daughters when crossed to females of any genotype (pure Bogota, pure USA, hybrid F1). Several lines of evidence suggest that this sex ratio distortion is caused by sex chromosome segregation distortion in hybrid males. We genetically analyze this normally cryptic segregation distortion and show that it involves several regions of the Bogota X chromosome that show strong epistatic interactions with each other. We further show that segregation distortion is normally masked within the Bogota subspecies by autosomal suppressors. Our analysis shows that the genetic basis of hybrid segregation distortion is similar to that of hybrid male sterility between the same subspecies. Indeed the severity of segregation distortion is correlated with the severity of sterility among hybrids. We discuss the possibility that hybrid sterility in this paradigmatic case of incipient speciation is caused by segregation distortion. PMID- 15654116 TI - Rapid concerted evolution of nuclear ribosomal DNA in two Tragopogon allopolyploids of recent and recurrent origin. AB - We investigated concerted evolution of rRNA genes in multiple populations of Tragopogon mirus and T. miscellus, two allotetraploids that formed recurrently within the last 80 years following the introduction of three diploids (T. dubius, T. pratensis, and T. porrifolius) from Europe to North America. Using the earliest herbarium specimens of the allotetraploids (1949 and 1953) to represent the genomic condition near the time of polyploidization, we found that the parental rDNA repeats were inherited in roughly equal numbers. In contrast, in most present-day populations of both tetraploids, the rDNA of T. dubius origin is reduced and may occupy as little as 5% of total rDNA in some individuals. However, in two populations of T. mirus the repeats of T. dubius origin outnumber the repeats of the second diploid parent (T. porrifolius), indicating bidirectional concerted evolution within a single species. In plants of T. miscellus having a low rDNA contribution from T. dubius, the rDNA of T. dubius was nonetheless expressed. We have apparently caught homogenization of rDNA repeats (concerted evolution) in the act, although it has not proceeded to completion in any allopolyploid population yet examined. PMID- 15654117 TI - Serotonin (5HT), fluoxetine, imipramine and dopamine target distinct 5HT receptor signaling to modulate Caenorhabditis elegans egg-laying behavior. AB - Drugs that target the serotonergic system are the most commonly prescribed therapeutic agents and are used for treatment of a wide range of behavioral and neurological disorders. However, the mechanism of the drug action remain a conjecture. Here, we dissect the genetic targets of serotonin (5HT), the selective 5HT reuptake inhibitor (SSRI) fluoxetine (Prozac), the tricyclic antidepressant imipramine, and dopamine. Using the well-established serotonergic response in C. elegans egg-laying behavior as a paradigm, we show that action of fluoxetine and imipramine at the 5HT reuptake transporter (SERT) and at 5HT receptors are separable mechanisms. Even mutants completely lacking 5HT or SERT can partially respond to fluoxetine and imipramine. Furthermore, distinct mechanisms for each drug can be recognized to mediate these responses. Deletion of SER-1, a 5HT1 receptor, abolishes the response to 5HT but has only a minor effect on the response to imipramine and no effect on the response to fluoxetine. In contrast, deletion of SER-4, a 5HT2 receptor, confers significant resistance to imipramine while leaving the responses to 5HT or fluoxetine intact. Further, fluoxetine can stimulate egg laying via the Gq protein EGL-30, independent of SER 1, SER-4, or 5HT. We also show that dopamine antagonizes the 5HT action via the 5HT-gated ion channel MOD-1 signaling, suggesting that this channel activity couples 5HT and dopamine signaling. These results suggest that the actions of these drugs at specific receptor subtypes could determine their therapeutic efficacy. SSRIs and tricyclic antidepressants may regulate 5HT outputs independently of synaptic levels of 5HT. PMID- 15654118 TI - An analysis of genetic diversity across the maize genome using microsatellites. AB - How domestication bottlenecks and artificial selection shaped the amount and distribution of genetic variation in the genomes of modern crops is poorly understood. We analyzed diversity at 462 simple sequence repeats (SSRs) or microsatellites spread throughout the maize genome and compared the diversity observed at these SSRs in maize to that observed in its wild progenitor, teosinte. The results reveal a modest genome-wide deficit of diversity in maize relative to teosinte. The relative deficit of diversity is less for SSRs with dinucleotide repeat motifs than for SSRs with repeat motifs of more than two nucleotides, suggesting that the former with their higher mutation rate have partially recovered from the domestication bottleneck. We analyzed the relationship between SSR diversity and proximity to QTL for domestication traits and observed no relationship between these factors. However, we did observe a weak, although significant, spatial correlation for diversity statistics among SSRs within 2 cM of one another, suggesting that SSR diversity is weakly patterned across the genome. Twenty-four of 462 SSRs (5%) show some evidence of positive selection in maize under multiple tests. Overall, the pattern of genetic diversity at maize SSRs can be explained largely by a bottleneck effect with a smaller effect from selection. PMID- 15654120 TI - Thematic review series: The immune system and atherogenesis. Paying the price for pathogen protection: toll receptors in atherogenesis. AB - Atherosclerosis is a chronic inflammatory response characterized by the accumulation of cells of innate and acquired immune systems within the intima of the arterial wall. Macrophages are the predominant participant in innate immune responses in atherosclerosis. Protein receptors expressed by macrophages and endothelial cells recognize components and products of microorganisms and play a vital role in innate immunity. In particular, the members of the toll-like receptor (TLR) family play a critical role in the inflammatory components of atherosclerosis. Both exogenous ligands involved in microbial recognition as well as endogenous ligands involved in sterile inflammation pathways are implicated in the pathology of atherosclerosis. In this review, we discuss our current understanding of the role of TLRs and their coactivators in atherosclerosis, with particular emphasis on studies in atherosclerosis-prone hypercholesterolemic mice. PMID- 15654119 TI - SepBCTF4 is required for the formation of DNA-damage-induced UvsCRAD51 foci in Aspergillus nidulans. AB - SepB is an essential, conserved protein required for chromosomal DNA metabolism in Aspergillus nidulans. Homologs of SepB include yeast Ctf4p and human hAnd-1. Molecular and bioinformatic characterization of these proteins suggests that they act as molecular scaffolds. Furthermore, recent observations implicate the yeast family members in lagging-strand replication and the establishment of sister chromatid cohesion. Here, we demonstrate that SepB functions in the A. nidulans DNA damage response. In particular, analysis of double mutants reveals that SepB is a member of the UvsC(RAD51) epistasis group. In accord with this prediction, we show that UvsC(RAD51) forms DNA-damage-induced nuclear foci in a manner that requires SepB function. We also provide evidence that implicates SepB in sister chromatid cohesion, thereby suggesting that cohesion may play a role in regulating the localization and/or assembly of UvsC(RAD51) complexes. PMID- 15654121 TI - Structural modification of plasma HDL by phospholipids promotes efficient ABCA1 mediated cholesterol release. AB - It has been suggested that ABCA1 interacts preferentially with lipid-poor apolipoprotein A-I (apoA-I). Here, we show that treatment of plasma with dimyristoyl phosphatidylcholine (DMPC) multilamellar vesicles generates prebeta(1)-apoA-I-containing lipoproteins (LpA-I)-like particles similar to those of native plasma. Isolated prebeta(1)-LpA-I-like particles inhibited the binding of (125)I-apoA-I to ABCA1 more efficiently than HDL(3) (IC(50) = 2.20 +/- 0.35 vs. 37.60 +/- 4.78 microg/ml). We next investigated the ability of DMPC-treated plasma to promote phospholipid and unesterified (free) cholesterol efflux from J774 macrophages stimulated or not with cAMP. At 2 mg DMPC/ml plasma, both phospholipid and free cholesterol efflux were increased ( approximately 50% and 40%, respectively) in cAMP-stimulated cells compared with unstimulated cells. Similarly, both phospholipid and free cholesterol efflux to either isolated native prebeta(1)-LpA-I and prebeta(1)-LpA-I-like particles were increased significantly in stimulated cells. Furthermore, glyburide significantly inhibited phospholipid and free cholesterol efflux to DMPC-treated plasma. Removal of apoA I-containing lipoproteins from normolipidemic plasma drastically reduced free cholesterol efflux mediated by DMPC-treated plasma. Finally, treatment of Tangier disease plasma with DMPC affected the amount of neither prebeta(1)-LpA-I nor free cholesterol efflux. These results indicate that DMPC enrichment of normal plasma resulted in the redistribution of apoA-I from alpha-HDL to prebeta-HDL, allowing for more efficient ABCA1-mediated cellular lipid release. Increasing the plasma prebeta(1)-LpA-I level by either pharmacological agents or direct infusions might prevent foam cell formation and reduce atherosclerotic vascular disease. PMID- 15654122 TI - Delineation of molecular changes in intrahepatic cholesterol metabolism resulting from diminished cholesterol absorption. AB - The absorption of cholesterol by the small intestine is a major route for the net entry of cholesterol into the body and can therefore affect the plasma low density lipoprotein-cholesterol (LDL-C) concentration. These studies used ezetimibe, a potent inhibitor of cholesterol absorption, to delineate the biochemical and molecular changes in intrahepatic metabolism and biliary lipid secretion when there is a major reduction in chylomicron cholesterol delivery to the liver. In female LDL receptor (LDLR)-deficient (LDLR-/-) mice fed a basal diet containing ezetimibe (0-10 mg/day/kg body weight), cholesterol absorption was reduced up to 91%, fecal neutral sterol excretion was increased up to 4.7 fold, and plasma total cholesterol concentrations decreased by up to 18%. Blocking cholesterol absorption prevented the accumulation of very low density lipoproteins and LDL in the circulation of LDLR-/- mice fed a lipid-rich diet. In female LDLR+/+ mice fed the lipid-rich diet with ezetimibe, the relative mRNA level for the LDLR in the liver was 2-fold greater than in matching mice given the lipid-rich diet alone. We conclude that in the mouse the reduction in plasma LDL-C levels induced by blocking cholesterol absorption reflects both a diminished rate of LDL-C production and a modest increase in hepatic LDLR expression. PMID- 15654123 TI - High-level lipoprotein [a] expression in transgenic mice: evidence for oxidized phospholipids in lipoprotein [a] but not in low density lipoproteins. AB - Efforts to elucidate the role of lipoprotein [a] (Lp[a]) in atherogenesis have been hampered by the lack of an animal model with high plasma Lp[a] levels. We produced two lines of transgenic mice expressing apolipoprotein [a] (apo[a]) in the liver and crossed them with mice expressing human apolipoprotein B-100 (apoB 100), generating two lines of Lp[a] mice. One had Lp[a] levels of approximately 700 mg/dl, well above the 30 mg/dl threshold associated with increased risk of atherosclerosis in humans; the other had levels of approximately 35 mg/dl. Most of the LDL in mice with high-level apo[a] expression was covalently bound to apo[a], but most of the LDL in the low-expressing line was free. Using an enzyme linked sandwich assay with monoclonal antibody EO6, we found high levels of oxidized phospholipids in Lp[a] from high-expressing mice but not in LDL from low expressing mice or in LDL from human apoB-100 transgenic mice (P <0.00001), even though all mice had similar plasma levels of human apoB-100. The increase in oxidized lipids specific to Lp[a] in high-level apo[a]-expressing mice suggests a mechanism by which increased circulating levels of Lp[a] could contribute to atherogenesis. PMID- 15654125 TI - Retina expresses microsomal triglyceride transfer protein: implications for age related maculopathy. AB - The principal extracellular lesions of age-related maculopathy (ARM), the leading cause of vision loss in the elderly, involve Bruch's membrane (BrM), a thin vascular intima between the retinal pigment epithelium (RPE) and its blood supply. With age, 80-100 nm solid particles containing esterified cholesterol (EC) accumulate in normal BrM, and apolipoprotein B (apoB) immunoreactivity is detectable in BrM- and ARM-associated lesions. Yet little evidence indicates that increased plasma cholesterol is a risk factor for ARM. To determine if RPE is capable of assembling its own apoB-containing lipoprotein, we examined RPE for the expression of microsomal triglyceride transfer protein (MTP), which is required for this process. Consistent with previous evidence for apoB expression, MTP is expressed in RPE, the ARPE-19 cell line, and, unexpectedly, retinal ganglion cells, which are neurons of the central nervous system. De novo synthesis and secretion of neutral lipid by ARPE-19 was supported by high levels of radiolabeled EC and triglyceride in medium after supplementation with oleate. Lipoprotein assembly and secretion is implicated as a constitutive retinal function and a plausible candidate mechanism involved in forming extracellular cholesterol-containing lesions in ARM. The pigmentary retinopathy and neuropathy of abetalipoproteinemia (Mendelian Inheritance of Man 200100; Bassen-Kornzwieg disease), which is caused by mutations in the MTP gene, may involve loss of function at the retina. PMID- 15654124 TI - Neutral glycolipids of the filamentous fungus Neurospora crassa: altered expression in plant defensin-resistant mutants. AB - To defend themselves against fungal pathogens, plants produce numerous antifungal proteins and peptides, including defensins, some of which have been proposed to interact with fungal cell surface glycosphingolipid components. Although not known as a phytopathogen, the filamentous fungus Neurospora crassa possesses numerous genes similar to those required for plant pathogenesis identified in fungal pathogens (Galagan, J. E., et al. 2003. Nature 422: 859-868), and it has been used as a model for studying plant-phytopathogen interactions targeting fungal membrane components (Thevissen, K., et al. 2003. Peptides. 24: 1705-1712). For this study, neutral glycolipid components were extracted from wild-type and plant defensin-resistant mutant strains of N. crassa. The structures of purified components were elucidated by NMR spectroscopy and mass spectrometry. Neutral glycosphingolipids of both wild-type and mutant strains were characterized as beta-glucopyranosylceramides, but those of the mutants were found with structurally altered ceramides. Although the wild type expressed a preponderance of N-2'-hydroxy-(E)-Delta3-octadecenoate as the fatty-N-acyl component attached to the long-chain base (4E,8E)-9-methyl-4,8-sphingadienine, the mutant ceramides were found with mainly N-2'-hydroxyhexadecanoate instead. In addition, the mutant strains expressed highly increased levels of a sterol glucoside identified as ergosterol-beta-glucoside. The potential implications of these findings with respect to defensin resistance in the N. crassa mutants are discussed. PMID- 15654126 TI - The mitochondrial respiratory complex I is a target for 15-deoxy-delta12,14 prostaglandin J2 action. AB - The prostaglandin J2 derivative 15-deoxy-Delta12,14-prostaglandin J2 (15d-PGJ2) is a very active compound with important effects on inflammation, apoptosis, and cell growth processes. To exert this broad range of effects, 15d-PGJ2 binds and alters the activity of diverse proteins, which consequently are postulated to be mediators of its action. Among them are the transcription factors peroxisome proliferator-activated receptor gamma and nuclear factor kappaB, which are thought to play an essential role in the antitumorigenic and anti-inflammatory actions of 15d-PGJ2. Here, we show that 15d-PGJ2, at micromolar concentrations, efficiently blocks state 3 oxygen consumption in intact nonsynaptic mitochondria isolated from rat cerebral cortex. This effect is attributable to the inhibition by this prostaglandin of the activity of the enzyme NADH-ubiquinone reductase (complex I) of the mitochondrial respiratory chain. In addition to this, 15d-PGJ2 dramatically increases the rate of reactive oxygen species generation by complex I. The inhibition by 15d-PGJ2 of complex I activity was abolished by dithiothreitol, which raises the possibility that adduct formation with a critical component of complex I accounts for the inhibitory effect of this prostaglandin. These results clearly identified mitochondrial complex I as a new target for 15d-PGJ2 actions. PMID- 15654127 TI - Expression, activity, and localization of hormone-sensitive lipase in rat mammary gland during pregnancy and lactation. AB - We examined the presence of hormone-sensitive lipase (HSL) in mammary glands of virgin, pregnant (12, 20, and 21 days), and lactating (1 and 4 days postpartum) rats. Immunohistochemistry with antibody against rat HSL revealed positive HSL in the cytoplasm of both alveolar epithelial cells and adipocytes. In virgin rats, immunoreactive HSL was observed in mammary adipocytes, whereas diffuse staining was found in the epithelial cells. Positive staining for HSL was seen in the two types of cells in pregnant and lactating rats. However, as pregnancy advanced, the staining intensity of immunoreactive HSL increased in the epithelial cells parallel to their proliferation, attaining the maximum during lactation. An immunoreactive protein of 84 kDa and a HSL mRNA of 3.3. kb were found in the rat mammary gland as in white adipose tissue. Both HSL protein and activity were lower in mammary glands from 20 and 21 day pregnant rats than from those of virgin rats, although they returned to virgin values on days 1 and 4 of lactation. Mammary gland HSL activity correlated negatively to plasma insulin levels. Immunoreactive HSL and HSL activity were found in lactating rats' milk. The observed changes indicate an active role of HSL in mammary gland lipid metabolism. PMID- 15654128 TI - Visualization and analysis of apolipoprotein A-I interaction with binary phospholipid bilayers. AB - Apolipoprotein A-I (apoA-I) interaction with specific cell lipid domains was suggested to trigger cholesterol and phospholipid efflux. We analyzed here apoA-I interaction with dimyristoylphosphatidylcholine/distearoylphosphatidylcholine (DMPC/DSPC) bilayers at a temperature showing phase coexistence. Solid and liquid crystalline domains were visualized by two-photon fluorescence microscopy on giant unilamellar vesicles (GUVs) labeled with 6-dodecanoyl-2-dimethyl-amino naphthalene (Laurdan). A decrease of vesicle size was detected as long as they were incubated with lipid-free apoA-I, together with a shape deformation and a relative enrichment in DSPC. Selective lipid removal mediated by apoA-I from different domains was followed in real time by changes in the Laurdan generalized polarization. The data show a selective interaction of apoA-I with liquid crystalline domains, from which it removes lipids, at a molar ratio similar to the domain compositions. Next, apoA-I was incubated with DMPC/DSPC small unilamellar vesicles, and products were isolated and quantified. Protein solubilized both lipids but formed complexes relatively enriched in the liquid component. We also show changes in the GUV morphology when cooling down. Our results suggest that the most efficient reaction between apoA-I and DMPC/DSPC occurs in particular bilayer conditions, probably when small fluid domains are nucleated within a continuous gel phase and interfacial packing defects are maximal. PMID- 15654129 TI - A phytol-enriched diet induces changes in fatty acid metabolism in mice both via PPARalpha-dependent and -independent pathways. AB - Branched-chain fatty acids (such as phytanic and pristanic acid) are ligands for the nuclear hormone receptor peroxisome proliferator-activated receptor alpha (PPARalpha) in vitro. To investigate the effects of these physiological compounds in vivo, wild-type and PPARalpha-deficient (PPARalpha-/-) mice were fed a phytol enriched diet. This resulted in increased plasma and liver levels of the phytol metabolites phytanic and pristanic acid. In wild-type mice, plasma fatty acid levels decreased after phytol feeding, whereas in PPARalpha-/- mice, the already elevated fatty acid levels increased. In addition, PPARalpha-/- mice were found to be carnitine deficient in both plasma and liver. Dietary phytol increased liver free carnitine in wild-type animals but not in PPARalpha-/- mice. Investigation of carnitine biosynthesis revealed that PPARalpha is likely involved in the regulation of carnitine homeostasis. Furthermore, phytol feeding resulted in a PPARalpha-dependent induction of various peroxisomal and mitochondrial beta-oxidation enzymes. In addition, a PPARalpha-independent induction of catalase, phytanoyl-CoA hydroxylase, carnitine octanoyltransferase, peroxisomal 3-ketoacyl-CoA thiolase, and straight-chain acyl-CoA oxidase was observed. In conclusion, branched-chain fatty acids are physiologically relevant ligands of PPARalpha in mice. These findings are especially relevant for disorders in which branched-chain fatty acids accumulate, such as Refsum disease and peroxisome biogenesis disorders. PMID- 15654131 TI - Cholesterol esterase action on human high density lipoproteins and inhibition studies: detection by MALDI-TOF MS. AB - The modification of lipoproteins by lipolytic enzymes such as cholesterol esterase (CEase) is assumed to play an important role in the pathogenesis of atherosclerosis. However, details of the activation and inhibition of CEase are still unknown. In this study, matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDI-TOF MS) was used to investigate the extracts of human lipoproteins after treatment with CEase and to monitor the effects of the inhibitor 2-(diethylamino)-6,7-dihydro-4H,5H-cyclopenta[4,5] thieno[2,3-d][1,3]oxazin-4-one (DOT-3). This approach has the advantage that all lipid classes can be independently detected; therefore, conclusions on the mechanism of the applied enzyme are possible. Besides the expected decrease of cholesteryl esters (CEs) in HDL, a significantly enhanced content of lysophosphatidylcholine (LPC) was also detected, confirming the broad substrate specificity of CEase. It was also demonstrated that DOT-3 significantly inhibited the CEase-catalyzed cleavage of CEs in HDL. Phospholipid (PL) vesicles prepared from phosphatidylcholine (PC) or PC and cholesteryl linoleate were treated with CEase, and the changes in lipid composition were investigated. From the analysis of the generated LPC species in HDL and in the isolated lipid mixtures, it is evident that CEase catalyzes the cleavage of the fatty acid residues in both the sn-1 and sn-2 positions of the PLs. These effects are obvious in the absence as well as in the presence of detergents. PMID- 15654130 TI - Tissue-specific, nutritional, and developmental regulation of rat fatty acid elongases. AB - Of the six fatty acid elongase (Elovl) subtypes expressed in mammals, adult rat liver expresses four subtypes: Elovl-5 > Elovl-1 = Elovl-2 = Elovl-6. Overnight starvation and fish oil-enriched diets repressed hepatic elongase activity in livers of adult male rats. Diet-induced changes in elongase activity correlate with Elovl-5 and Elovl-6 mRNA abundance. Adult rats fed the peroxisome proliferator-activated receptor alpha (PPARalpha) agonist WY14,643 have increased hepatic elongase activity, Elovl-1, Elovl-5, Elovl-6, Delta5, Delta6, and Delta9 desaturase mRNA abundance, and mead acid (20:3,n-9) content. PPARalpha agonists affect both fatty acid elongation and desaturation pathways leading to changes in hepatic lipid composition. Elovl activity is low in fetal liver but increases significantly after birth. Developmental changes in hepatic elongase activity paralleled the postnatal induction of Elovl-5 mRNA and mRNAs encoding the PPARalpha-regulated transcripts, Delta5 and Delta6 desaturase, and cytochrome P450 4A. In contrast, Elovl-6, Delta9 desaturase, and FAS mRNA abundance paralleled changes in hepatic sterol regulatory element binding protein 1c (SREBP 1c) nuclear content. SREBP-1c is present in fetal liver nuclei, absent from nuclei immediately after birth, and reappears in nuclei at weaning, 21 days postpartum. In conclusion, changes in Elovl-5 expression may account for much of the nutritional and developmental control of fatty acid elongation activity in the rat liver. PMID- 15654133 TI - Women living with human immunodeficiency virus or acquired immunodeficiency syndrome: a global epidemic. PMID- 15654132 TI - Physiological importance of SR-BI in the in vivo metabolism of human HDL and LDL in male and female mice. AB - The physiological role of murine scavenger receptor class B type I (SR-BI) was evaluated by in vivo clearances of human HDL3 and LDL in normal and SR-BI knockout (KO) mice. In normal mice, cholesteryl esters (CEs) were removed faster than proteins, indicating a selective uptake process from both HDL3 and LDL. SR BI KO mice showed 80% losses of HDL-CE selective uptake and the complete loss of LDL-CE selective uptake in the first phase of clearance. However, the second phase was characterized by an acceleration of CE disappearance in SR-BI KO mice. Thus, SR-BI is the only murine receptor mediating HDL-CE selective uptake, whereas a SR-BI-independent pathway specific to LDL can rescue SR-BI deficiency. The analysis of LDL recovered 3 h after injection in mice from different genotypes revealed that LDLs are significantly depleted in CE (reduction from 19% to 50% of the CE/protein ratios). A smaller LDL size in comparison with that of noninjected LDL was also detectable but was more evident for LDL recovered from normal mice. All LDL preparations migrate faster than noninjected LDL on agarose barbital gels. Thus, both SR-BI-dependent and -independent pathways lead to substantial changes in LDL. PMID- 15654134 TI - Intrauterine insemination in the management of infertility. PMID- 15654135 TI - Primary prevention: why focus on children & young adolescents? AB - A focus on children and young adolescents in the primary prevention of health risks and disorders such as cancer, hypertension and other cardiovascular diseases, HIV/AIDS, and obesity has been suggested in many reports published throughout the world. Such a focus is important in India as it has a huge adolescents and children population along with the existing economic, social, and health inequalities among the general population. We propose a systematic elucidation of the rationale for such a focus in primary prevention research. We have reviewed studies describing risk factors, the association between risk factors and disease outcomes in affected patients, exposed populations, adolescent samples, as well as reports from studies conducted in India, and the quantitative and qualitative statistical aspects of research. The literature indicates that a lengthy time interval occurs between exposure to high risk factors and the development of disease, and that many such high risk exposures begin in young adolescence. These findings underline the value of targeting children and adolescents for primary prevention efforts in health care and health education for the attainment of overall healthy population in any country including a country like India. PMID- 15654136 TI - A randomised prospective trial of intrauterine insemination versus timed intercourse in superovulated cycles with clomiphene. AB - BACKGROUND & OBJECTIVES: Despite its wide use, the benefits of intrauterine insemination (IUI), its over timed intercourse (TI) in couples with unexplained infertility is a matter of debate. Studies in Indian couples with unexplained infertility showing benefit of IUI over TI are not available. The present study was done with the objective of comparing TI and IUI with husband's sperm in couples with unexplained infertility undergoing superovulation with clomiphene. METHODS: A total of 140 couples with unexplained infertility were subjected to controlled ovarian hyperstimulation (COH) with clomiphene and prospectively randomized to receive either TI (group A) or IUI (group B). Complete follow up was available for 113 couples only. RESULTS: The pregnancy rate and cycle fecundity rate after COH/TI was 41 and 8.8 per cent and after COH/IUI 18 and 3.4 per cent respectively. The difference was statistically not significant. INTERPRETATION & CONCLUSION: The findings of the present study showed that in women with unexplained infertility addition of IUI to ovulation induction does not improve conception rates. COH/TI can help to achieve good results and save the expense and discomfort due to a invasive procedure. PMID- 15654137 TI - Chromobacterium violaceum septicaemia from north India. AB - Though Chromobacterium violaceum is a common inhabitant of soil and water in tropical and sub-tropical regions, human infections are rare but when they do occur result in high mortality. Since the first case from Malaysia in 1927, about 150 cases have been reported in world literature. Till date 6 cases have been reported from southern and eastern parts of India. We report here a case of C. violaceum septicaemia, probably the first case from north India. The patient, a 6 and a half year old boy was admitted with high fever. The patient had anaemia, neutrophilic leucocytosis and bilateral chest infiltrates. Routine and bacteriological investigations were carried out to establish the aetiological diagnosis. C. violaceum was isolated in pure culture from blood and pus. The patient was successfully treated with ciprofloxacin and amikacin. This is probably the first documented case report of C. violaceum infection from north India and the only Indian case with septicaemia which survived. PMID- 15654138 TI - Pre-versus post-formalin effects of intrathecal ketamine on spinal Fos-like immunoreactivity in rats. AB - BACKGROUND & OBJECTIVES: The spinal expression of the c-Fos immediate early gene in response to formalin pain of the hind paw of rat was used as a marker of neuronal activity. Ketamine, a Nmethyl-D-aspartate (NMDA) receptor antagonist produces analgesic action due to the blockade of glutamate action at the NMDA receptor. Earlier study showed that ketamine acts differently depending on its route of administration. We undertook this study to compare a preemptive suppression of noxious stimulation induced spinal Fos-like immunoreactivity (FLI) after receiving intrathecal ketamine before or after formalin pain. METHODS: Male Sprague-Dawley rats received ketamine 1 mg/kg or saline (control group) intrathecally either 5 min before (pre-treatment group) formalin or 5 min after (post-treatment group) formalin (5%, 50 microl) injection. Animals were killed 2 h after the formalin injection, and the lumbar spinal cord was dissected, and processed by immunoperoxidase staining using an antibody against Fos protein. RESULTS: The FLI was significantly reduced in the pre-treatment group, only laminae I-II of the side ipsilateral to the formalin injection (P < 0.05 vs. control). In laminae V-VI, neither of the ketamine treatment groups showed a significant decrease than the control group. INTERPRETATION & CONCLUSION: The results provide evidence that intrathecal ketamine does not have a preemptive blocking effect of FLI expression in whole spinal laminae area. FLI expression of laminae I-II only might not be a good predictor of the ability of agents to produce preemptive effect. The central patterns of activity generated during central sensitization differ regionally in the spinal dorsal horn. PMID- 15654140 TI - Serum lipoprotein (a) levels in liver diseases caused by hepatitis. AB - Lipoprotein (a) [LP(a)] is a genetic variant of low density lipoprotein (LDL) and is mainly synthesized in liver. We conducted a study to evaluate the association of serum [Lp(a)] level with hepatitis viral infections. A total of 130 patients including 50 patients with acute viral hepatitis (AVH), 30 with chronic active hepatitis (CAH), 30 with cirrhosis of liver and 20 patients with fulminant hepatic failure (FHF) were analysed for different hepatitis viral markers and Lp(a) level in their serum samples. For comparison, 50 healthy persons were also tested for Lp(a) level. Serum Lp(a) level in patients in all the disease groups was significantly reduced compared to that observed in controls. Lp(a) level could not be detected in 40 per cent cases with AVH, 46.6 per cent with CAH, 70 per cent with cirrhosis and 80 per cent of FHF patients. On correlating Lp(a) level to viral etiology in these patients, it was found that the extent of diminution in Lp(a) level did not follow any trend with some particular viral infection and was recorded nearly same in all the infections. The findings of this study suggested that serum Lp(a) level was significantly (P<0.001) reduced in patients with liver diseases irrespective of the viral etiology. PMID- 15654139 TI - Effect of sensitization on membrane ion fluxes & intracellular calcium in guineapigs. AB - BACKGROUND & OBJECTIVES: The biochemical mechanisms underlying the development of sensitization-induced airway hyperresponsiveness (AHR) in asthma are poorly defined. Alterations in the regulation of intracellular calcium may play an important role in its pathogenesis. We carried out this study to see the effect of sensitization with ovalbumin on membrane ion fluxes and intracellular calcium in a guinea pig model. METHODS: Airway reactivity to inhaled histamine was measured initially and after sensitization with ovalbumin in 28 guineapigs. Intracellular calcium [Ca(2+)]i was measured in tracheal smooth muscle cells and peripheral leukocytes using fluorescent dye FURA 2AM. Calcium and sodium ion influx across the cell membrane was measured in leukocytes. Ouabain-sensitive Rubidium ((86)Rb) influx was measured in tracheal smooth muscles cells. The activities of Na(+), K(+) ATPase and Ca(2+) ATPase were measured in tracheal smooth muscle cells. Lipid peroxides were measured in plasma. RESULTS: Airway responsiveness was significantly (P<0.001) increased after sensitization along with an increase in [Ca2+]i levels in leukocytes and tracheal smooth muscle cells, higher rates of (45)Ca and (22)Na influx in leukocytes and higher (86)Rb influx rates in tracheal smooth muscle cells, and increased levels of lipid peroxides in plasma. INTERPRETATION & CONCLUSION: In guineapig model of asthma sensitization to allergen increased the membrane permeability to calcium and sodium, and intracellular calcium levels. These alterations may play a role in the pathogenesis of airway hyper-responsiveness following sensitization. PMID- 15654141 TI - In vitro activity of linezolid & quinupristin/dalfopristin against Gram-positive cocci. AB - BACKGROUND & OBJECTIVES: Since the incidence of vancomycin- and methicillin resistant Gram-positive infections continue to increase, novel antimicrobials such as linezolid and streptogramin may provide new options to treat patients. The aim of this study was to investigate in vitro susceptibility of Enterococcus faecium resistant to glycopeptides, coagulase negative staphylococci and S. aureus resistant to methicillin isolated mainly from blood and also rectal swab cultures of patients against quinupristin/dalfopristin and linezolid. METHODS: The in vitro susceptibility to linezolid and quinupristin/dalfopristin for a total of 332 isolates of Gram-positive cocci [127 methicillin-resistant Staphylococcus aureus, 109 methicillin-resistant coagulase negative staphylococci (71 S. epidermidis, 38 S. haemolyticus) and 96 vanA genotype vancomycin-resistant Enterococcus faecium] was investigated by E test. RESULTS: All MRSA and MRCoNS isolates were susceptible to linezolid (MICs < 4.0 mg/l). Ninety per cent of VRE isolates were inhibited by linezolid at concentration of 2.0 mg/l and presented similar activities to quinupristin/dalfopristin. MICs for quinupristin/dalfopristin against staphylococci were also low (MIC(90) = 1.0 mg/l for both MRSA and MRCoNS isolates). INTERPRETATION & CONCLUSION: The results of the present study demonstrated that quinupristin/ dalfopristin and linezolid, have good in vitro activity against MRSA, MRCoNS and vancomycin resistant E. faecium in Turkey. These drugs could be promising therapeutic options in an era of rapidly growing antibiotic resistance in all parts of world. PMID- 15654142 TI - Evaluation of extended spectrum beta lactamase in urinary isolates. AB - BACKGROUND & OBJECTIVES: Urinary tract infection (UTI) remain the common infections diagnosed in outpatients as well as in hospitalized patients. Current knowledge on antimicrobial susceptibility pattern of uropathogens is mandatory for appropriate therapy. Extended spectrum beta lactamases (ESBL) hydrolyse expanded spectrum cephalosporins like ceftazidime, cephotaxime which are used in the treatment of UTI. ESBL producing bacteria may not be detectable by routine disk diffusion susceptibility test, leading to inappropriate use of antibiotics and treatment failure. Not much information on ESBL producing organisms causing UTI is available from India. An effort was therefore made to study the ESBL producing uropathogens and also the susceptibility patterns of ESBL and nonESBL producers. METHODS: Urinary isolates from symptomatic UTI cases attending or admitted to the Indira Gandhi Medical College and Hospital, Nagpur were identified by conventional methods. Antimicrobial susceptibility testing was done by Kirbey Bauer's disc diffusion method. Isolates resistant to cephotaxime were tested for ESBL production by double disc synergy test method. RESULTS: Of the 217 isolates, 87 were cephotaxime resistant Gram-negative bacilli. Of these, 42 (48.3%) were found to be ESBL producers. Escherichia coli, Klebsiella pnuemoniae and Acinetobacter were ESBL producing species. Multidrug resistance was found to be significantly (P<0.05) more in ESBL producing isolates (90.5%) than non ESBL producers (68.9%). INTERPRETATION & CONCLUSION: In the present study a large number of uropathogens were found to be ESBL producers. Most of the ESBL producing isolates were multidrug resistant. Monitoring of ESBL production and antimicrobial susceptibility testing are necessary to avoid treatment failure in patients with UTI. PMID- 15654143 TI - Serodiagnosis of leptospirosis in Delhi using IgM enzyme linked immunosorbent assay. PMID- 15654145 TI - Modern cardiac electrophysiology and the elderly. PMID- 15654147 TI - Antiarrhythmic therapy in elderly persons: a device-based approach. AB - The management of ventricular arrhythmias in elderly persons has undergone a dramatic evolution over the past 10 years. Although life-threatening arrhythmias in elderly persons have been traditionally managed with a variety of pharmacologic agents, this population presents special challenges from pharmacokinetic and pharmacodynamic perspectives. Drug absorption, distribution, metabolism, and efficacy are often altered in elderly patients, resulting in a substantially narrowed therapeutic window. Nonpharmacologic therapy for ventricular arrhythmias has the advantage of not being subject to changes in metabolism, and due to recent technological advances in transvenous lead design as well as improved programming flexibility and reduction in device size, device based therapy for malignant ventricular arrhythmias has become more attractive for use in elderly patients. Several recent studies have suggested that device based therapy provides superior protection from malignant arrhythmias for both primary and secondary prevention indications. The majority of these studies suggest that the benefits of device-based therapy are conferred to patients independent of age. In addition, the complication rate of such therapy appears to be independent of age. PMID- 15654148 TI - Role of electrophysiologic studies, signal-averaged electrocardiography, heart rate variability, T-wave alternans, and loop recorders for risk stratification of ventricular arrhythmias. AB - Ventricular tachycardia and fibrillation are major causes of morbidity and mortality after myocardial infarction. Frequently, sudden cardiac death is the first manifestation of such malignant rhythms. Optimal risk stratification strategies in this population are of utmost importance. In this review the authors discuss the background and clinical use of invasive tests, such as electrophysiologic study and implantable loop recorders, and noninvasive tests, such as signal-averaged electrocardiography, heart rate variability, and T-wave alternans. The utility, indications, and limitations of each test in clinical practice are discussed, especially for the purpose of postmyocardial infarction risk stratification in the elderly population. PMID- 15654150 TI - Rate control vs. pulmonary vein isolation. AB - Atrial fibrillation (AF) is a frequent arrhythmia encountered in clinical practice. It can be asymptomatic in some patients but incapacitating in others. Regardless of symptoms, patients at increased risk of embolism need chronic anticoagulation. In patients with AF and a rapid ventricular response that cannot be controlled with drugs, interventional procedures are required. Radiofrequency catheter ablation of the atrioventricular junction with pacemaker implantation is an effective therapeutic approach that relieves the symptoms associated with a rapid and irregular rhythm. This approach can also improve left ventricular function in patients with tachycardia-induced cardiomyopathy. Due to the irreversible nature of this approach, it is more suitable for older patients and those with advanced left ventricular dysfunction. Many patients with a structurally normal heart have symptomatic paroxysmal AF refractory to antiarrhythmic agents. This form of AF is frequently initiated by premature atrial beats arising from the pulmonary veins. A catheter ablation technique is available to electrically isolate the pulmonary veins. This procedure can eliminate AF in many patients but can result in complications, including embolic events, pulmonary veins stenosis, and cardiac perforation. PMID- 15654151 TI - Cardiac resynchronization therapy: a new therapy for advanced congestive heart failure. AB - Despite medical therapy, many patients with advanced systolic dysfunction remain highly symptomatic. In these patients the presence of a left bundle branch block on electrocardiogram indicates significant dyssynchrony of ventricular contraction. Cardiac resynchronization, by means of biventricular pacing, results in important clinical benefits. Due to the risk for malignant ventricular arrhythmias, this technology is best combined with an implantable cardioverter defibrillator. PMID- 15654152 TI - Choice of pacemakers in patients aged 75 years and older: ventricular pacing mode vs. dual-chamber pacing mode. AB - Half of all pacemakers implanted in the United States are for patients aged 75 years and older. The expectations and needs of an older group are different from patients who are younger, yet it is only recently that different pacing mode benefits for elderly persons have been tested in clinical trials. Some of the results have been surprising and suggest new algorithms for management. Other issues are still on the threshold of investigation. These include pacing for heart failure in elderly patients and pacing combined with cardioverter defibrillator implantation. PMID- 15654153 TI - Ethical issues in the management of geriatric cardiac patients. How do you deal with a difficult patient? PMID- 15654154 TI - Diet, lifestyle, mortality, and memory in the elderly. PMID- 15654149 TI - Ablation therapy of supraventricular tachycardia in elderly persons. AB - Ablation of supraventricular tachycardia in elderly persons presents a challenging problem to electrophysiologists. Friable cardiac structures, prone to catheter perforation, comorbid cardiovascular disease, and the propensity to develop atrial fibrillation and thromboembolic complications, place these patients at high risk. Newer techniques for cardiac mapping and ablation, the establishment of precise ablation lines, and safer approaches to the ablation mechanism (particularly for atrial fibrillation) are presented in this review. In addition, a novel rapid ablation method to permanently cure atrial ventricular nodal re-entry tachycardia is demonstrated. PMID- 15654155 TI - Live three-dimensional transthoracic echocardiographic demonstration of coronary artery to pulmonary artery fistula. PMID- 15654156 TI - Shifting junctional pacemaker with artifactual Q-waves and J(ST) elevations. PMID- 15654157 TI - The infected decubitus ulcer: Superman's kryptonite. PMID- 15654158 TI - The role of tumor necrosis factor-alpha inhibitors in dermatology. PMID- 15654159 TI - Follicular mycosis fungoides: variability of a rare entity. AB - OBJECTIVE: Three patients with follicular mycosis fungoides (FMF), a rare variant of folliculotropic cutaneous T-cell lymphoma, are discussed. Follicular involvement in cutaneous T-cell lymphoma commonly presents clinically with alopecia, follicular cysts, or comedo-like lesions usually associated with follicular mucinosis and strong epidermotropism. In contrast, FMF, in a strict sense, has been defined as a lymphocytic infiltration of hair follicles in the absence of both epidermal invasion and follicular mucin. DISCUSSION: Case 1 shows a clinically stable form of FMF characterized by circumscribed alopecia and its persistent resistance to topical therapeutic approaches. In contrast, Cases 2 and 3 demonstrate that classic mycosis fungoides lesions and FMF can occur simultaneously and that transitions occur from mycosis fungoides to FMF and vice versa. These observations strengthen the concept that FMF and mycosis fungoides represent just two variants of one entity of cutaneous T-cell lymphoma. In Case 3, rapid progression into tumor-stage mycosis fungoides and the follicular infiltration was controlled with bexarotene. CONCLUSIONS: Because it was previously suggested that FMF may have a worse prognosis compared with classic mycosis fungoides, this impressive response to bexarotene is of great clinical interest and may spark new studies employing this novel retinoid for FMF treatment. PMID- 15654160 TI - Hair and nail relationship. AB - Hair and nails are often stated to have much in common in relation to their origin, anatomical structures, and common involvement in many diseases. Hair and nails are predominantly epithelial structures derived from primitive epidermis and made up of keratinous fibrils embedded in a sulfur-rich matrix. It was first noted early in the 20th century that the nail unit was comparable in several respects to a hair follicle sectioned longitudinally and laid on on its side. The epithelial components of hair follicle and nail apparatus are differentiated epidermal structures that may be involved jointly in several ways as congenital and hereditary anomalies and acquired conditions such as alopecia areata, lichen planus, iatrogenic causes, and fungal infection. Multielemental characterization of human hair and nails shows that even after sample washing, many elements are enriched in the surface of the nails. PMID- 15654162 TI - Creating and utilizing a multimedia dermatology medical record for a PocketPC personal digital assistant. AB - Personal digital assistants are lightweight computers that capture and display data via tapping on their screens with a stylus and are easily linked to desktop and network computers. They have been used in medicine for a variety of purposes, and many believe personal digital assistant use can improve the provision of medical care. The author created a multimedia dermatology electronic medical record for a PocketPC (Microsoft Corp., Redmond, WA) personal digital assistant that contains patient images, tables of phototherapy, laboratory and systemic medication data, and typed chart notes. Such a record can be created and utilized but requires more time to assemble than a handwritten note, mostly due to capturing and organizing images. Future challenges involve streamlining record assembly, integrating multimedia records with hospital and office medical records, and assessing how having multimedia data available might affect care. PMID- 15654161 TI - Is there a role for tumor necrosis factor-alpha inhibitors and especially thalidomide in dermatology? AB - Because of the relative shortcomings and their side effects of the available anti inflammatory drugs such as systemic nonsteroids, corticosteroids, and immunosuppressive drugs, and since tumor necrosis factor-a plays a major role in noninfectious inflammatory and autoimmune disorders, tumor necrosis factor-a inhibitory drugs and the available tumor necrosis factor-a inhibitory biologic modifying reagents are described. Among the drugs reviewed are pentoxifylline, thalidomide, etanercept, infliximab, and adalimumab. Their relative effectiveness and side effects are reported and recommendations are made. PMID- 15654163 TI - Light-emitting diode-based therapy. PMID- 15654164 TI - ZODERM (benzoyl peroxide with urea in cream, gel, and cleanser preparations). PMID- 15654165 TI - The influence of proteomics on cosmetic science. PMID- 15654166 TI - Rosacea in a new light. PMID- 15654167 TI - Seborrheic dermatitis. PMID- 15654169 TI - An unusual complication of erythema multiforme and its treatment. PMID- 15654168 TI - The significant threat of patients with cutaneous malignant melanoma to develop ocular melanoma. PMID- 15654170 TI - Erythromelalgia. PMID- 15654171 TI - Twenty-nail dystrophy originating from lichen planus. PMID- 15654172 TI - Treatment of depression and effect of antidepression treatment on nutritional status in chronic hemodialysis patients. AB - BACKGROUND: Depression, which is the most common psychological complication in patients with end-stage renal disease (ESRD), has an impact on the clinical outcome and is associated with malnutrition in chronic hemodialysis patients. This study evaluated the effect of antidepression treatment on nutritional status in depressed chronic hemodialysis patients. METHODS: Sixty-two ESRD patients who underwent dialysis for more than 6 months were interviewed and completed a Beck Depression Inventory assessment. Thirty-four patients who had scores greater than 18 on the Beck Depression Inventory score and met Diagnostic and Statistical Manual of Mental Disorders, 4th Edition criteria for major depressive disorder were selected to receive paroxetine 10 mg/day and psychotherapy for 8 weeks. The remaining 28 patients were assigned to the control group. Change in the severity of depressive symptoms was ascertained by administering the Hamilton Depression Rating Scale. Nutritional status was evaluated by normalized protein catabolic rate, serum albumin and blood urea nitrogen level. RESULTS: All patients successfully completed 8 weeks of antidepression treatment. Antidepression treatment decreased the severity of depressive symptoms (Hamilton Depression Rating Scale score: 16.6 +/- 7.0 versus 15.1 +/- 6.6, P < 0.01) and increased normalized protein catabolic rate (1.04 +/- 0.24 versus 1.17 +/- 0.29 g/kg/day, P < 0.05), serum albumin (37.3 +/- 2.0 versus 38.7 +/- 3.2 g/l, P < 0.005), and prehemodialysis blood urea nitrogen level (24.3 +/- 5.6 versus 30.2 +/- 7.9 mmol/L, P < 0.001). In the control group, no change was noted during the study period. CONCLUSION: This study suggests that antidepressant medication with supportive psychotherapy can successfully treat depression and improve nutritional status in chronic hemodialysis patients with depression. PMID- 15654173 TI - Imported malaria at an inner-city hospital in the United States. AB - BACKGROUND: More than 1000 cases of malaria are reported to the Centers for Disease Control and Prevention each year among travelers or immigrants. METHODS: Retrospective study of patients with malaria seen at Grady Memorial Hospital in Atlanta, Georgia, between October 1988 and September 2000. RESULTS: One hundred twenty-six cases of malaria were diagnosed at Grady Memorial Hospital during the study period. Fourteen patients had seen a physician prior to coming to Grady Memorial Hospital, and in 71% the diagnosis was missed. Half had recently immigrated and half recently traveled to an endemic area, yet only 22% of travelers took prophylaxis. Plasmodium falciparum was the most commonly identified species (52.4%), followed by Plasmodium vivax (23.9%). Seventy-two patients (57.1%) required hospitalization. Presenting symptoms included fever (94%), chills (56%), nausea/vomiting (38%), headache (26%), and abdominal pain (26%). Most patients were diagnosed correctly on the day of admission (79%). Twelve patients (16.7%) had severe malaria, and their complications included severe anemia (9.7%), acute renal failure (4.2%), bleeding and/or disseminated intravascular coagulation (4.2%), shock (2.8%), seizures (2.8%), and hypoglycemia (2.8%). One patient died, and two pregnant women had premature deliveries. Median hospital stay was 3.9 days. One third of the hospitalized patients with P vivax failed to receive primaquine, and in 15% of patients with P falciparum, the treatment was considered to be inappropriate. CONCLUSIONS: Although some patients with malaria have a benign course and a good outcome, many patients require hospitalization, and some have severe complications. Increased efforts are needed to educate travelers about the need for prophylaxis. PMID- 15654174 TI - Simvastatin and L-arginine preserve renal function after ischemia/reperfusion injury. AB - BACKGROUND: HMG-CoA reductase inhibitors have been shown to have beneficial renal hemodynamic effects by increasing renal blood flow, independent of their lipid lowering properties. Currently in organ transplantation, the calcineurin inhibitor cyclosporine A (CyA) is the immunosuppressant of choice. However, its use is limited by its nephrotoxic effects, namely its renal vasoconstrictor properties. The purpose of this study was to determine the effect of an HMG-CoA reductase inhibitor, simvastatin (Zocor), on renal function in rats and on urinary nitrite/nitrate production following ischemia/reperfusion injury (I/R) with concomitant cyclosporine treatment. In addition, L-NAME (N(G)-nitro-L arginine methyl ester) and L-arginine were administered with CyA to the rats to test the hypothesis that simvastatin's beneficial effects were due to nitric oxide. METHODS: Male Wistar rats (250 g) were anesthetized and the supra-aorta clamped for 40 minutes. The right kidney was removed. After recovery, the rats were divided into five groups: 1) controls, no ischemia, no treatment (CTRL, n = 8); 2) ischemia (ISCH) plus cyclosporine A only (CyA, 5 mg/kg/day i.p., n = 8); 3) ischemia plus CyA and simvastatin (SIM, 10 mg/kg/day, gavage, n = 8); 4) ischemia plus simvastatin plus L-NAME plus CyA (10 mg/kg/day, gavage, n = 8), and 5) ischemia plus simvastatin plus L-arginine (2% in drinking water, n = 7) plus CyA. Five to 7 days after I/R injury, the glomerular filtration rate (GFR) was determined using urinary iohexol clearance. Urinary nitrite/nitrate production was determined using nitrate reductase and the Greiss reaction. Data are expressed as mean +/- SEM, and intergroup comparisons were made using one-way analysis of variance. RESULTS: The GFR values (mL/min) for all five groups are as follows: 1) CTRL = 1.25 +/- 0.10; 2) ISCH plus CyA only = 0.45 +/- 0.06 (P < 0.05 versus CTRL, ISCH only and simvastatin and cyclosporine and simvastatin plus L arginine and cyclosporine); 3) CyA and SIM = 0.78 +/- 0.09, CyA and L-NAME = 0.62 +/- 0.12, and CyA and L-arginine and SIM = 1.57 +/- 0.12. Results in the control were significantly different from results in the ischemic only and the L-NAME groups (P < 0.05). The L-arginine plus cyclosporine and simvastatin group was significantly higher than the ischemic only group, ischemic plus simvastatin and cyclosporine and the L-NAME plus cyclosporine group (P < 0.05). No significant differences could be detected in the urinary nitric oxide concentrations. CONCLUSIONS: : After I/R injury and cyclosporine treatment, simvastatin and L arginine preserved renal function, compared with cyclosporine treatment alone, because simvastatin and L-arginine may not have a direct vasoconstrictor effect on the renal microcirculation. They may be suppressing endothelin or increasing other vasodilator mediators such as the vasodilator prostaglandins and/or nitric oxide. PMID- 15654175 TI - Diagnosis of a pheochromocytoma. AB - Although rare, pheochromocytomas are potentially lethal tumors. Thus, it is important that physicians be able to diagnose these tumors. The definitive diagnosis of a pheochromocytoma rests on demonstrating catecholamine overproduction. Once the diagnosis is established, computed tomography scan, magnetic resonance imaging, and metaiodobenzylguanidine studies are utilized for localizing the tumor. This paper reviews the biochemical and radiologic studies useful for evaluating a patient for the possibility of a pheochromocytoma. PMID- 15654176 TI - Improving patient care: learning more from bad outcomes. AB - Many quality improvement efforts look in detail at episodes in which things have gone awry through error, to learn how to prevent repetition of that or related errors, with a particular focus on adverse drug reactions. Looking in a more global fashion at the various scenarios in which matters go amiss, rather than solely at the negative consequences of administering a therapy in error, allows us to derive more useful evidence about best therapeutic practice. An analytic strategy is presented to accomplish this aim, looking at the respective adverse effects associated with giving or with omitting a therapy, in each case both in situations in which the therapy is indicated and in situations in which it is contraindicated. PMID- 15654177 TI - Tachycardia-induced cardiomyopathy: atrial fibrillation and congestive heart failure. AB - Tachycardia-induced cardiomyopathy occurs as a result of prolonged, excessive heart rates. Ventricular function may improve significantly upon control of the heart rate. We present a case of a patient with atrial fibrillation with rapid ventricular response who showed a dramatic improvement in left ventricular function following AV nodal ablation and insertion of a pacemaker. We also review the history and pathophysiology of tachycardia-induced cardiomyopathy. PMID- 15654178 TI - Malignant pleural mesothelioma. AB - Malignant pleural mesothelioma is an uncommon tumor; only about 3000 cases are diagnosed annually in the United States. Cases were described early in the 20th century, but their relationship to asbestos exposure was not documented until 1960. Since then, the incidence has appeared to increase, and numerous epidemiologic studies have confirmed that exposure to asbestos in a variety of settings and occupations is the most significant risk factor for the development of malignant pleural mesothelioma. More recently, the oncogenic virus SV40 has also been implicated as a potential etiologic agent. Surgery, radiotherapy, and chemotherapy have each been used in the treatment of mesothelioma, but generally with little impact on survival. New directions in therapy include aggressive multimodality programs for potentially resectable patients and targeted therapies, including antifolates, antiangiogenesis agents, and drugs directed at epidermal growth factor receptor for the majority of patients presenting with unresectable disease. PMID- 15654179 TI - Pectus excavatum with inspiratory inferior vena cava compression: a new presentation of pulsus paradoxus. AB - A 29-year-old man with pectus excavatum presented with exercise intolerance, pulsus paradoxus, and paradoxically split S2. Chest computed tomography (CT) showed the heart shifted leftward and a pectus severity index of 7.18. Cardiopulmonary exercise study showed reduced VO2max, anaerobic threshold, and oxygen pulse. Echocardiography revealed a decline in mitral and tricuspid valve inflow, and stroke volume during inspiration. Cardiac extrinsic compression and anatomic cardiac abnormalities were not present. Dynamic magnetic resonance imaging (MRI) demonstrated inspiratory inferior vena cava (IVC) compression at the diaphragm. We discuss IVC compression by the diaphragm as a source of patient symptoms and as a mechanism for pulsus paradoxus associated with pectus excavatum. PMID- 15654180 TI - Paraneoplastic neurologic syndrome and autoimmune Addison disease in a patient with thymoma. AB - A 48-year-old man with autoimmune Addison disease developed the following paraneoplastic neurologic syndromes (PNNS): limbic encephalitis, opsoclonus/myoclonus, and sensorimotor and autonomic neuropathies. An anterior mediastinal mass detected on a chest computed tomographic scan was found on resection to be a noninvasive lymphocytic thymoma. The PNNS went into remission 1 year after the thymectomy. This is the first case of thymoma associated with autoimmune Addison disease and PNNS to be described in the literature. PMID- 15654181 TI - Ramipril-induced agranulocytosis confirmed by a lymphocyte cytotoxicity test. AB - A 50-year-old man who had chronic renal failure presented with neutropenic fever four days after ramipril was initiated. Agranulocytosis due to other causes was ruled out after a bone marrow aspiration and biopsy examination were performed. A relationship between the drug and the adverse effect was suggested. It was established by a novel lymphocyte cytotoxicity test. PMID- 15654182 TI - Pancreatic abscess secondary to Alcaligenes faecalis. AB - We report a patient with pancreatic abscesses and necrosis secondary to Alcaligenes faecalis infection. He initially presented with alcohol-induced acute pancreatitis. Twenty days after the initial presentation, he re-presented with increasing pain and was found to have pancreatic necrosis and abscesses. Treatment was initiated with meropenem. Because of persistent fevers, computed tomography-guided drainage was performed. The fluid grew A faecalis resistant to meropenem and the patient continued to be febrile. He recovered only after adequate surgical intervention and appropriate antibiotic coverage. Although this is the first case of A faecalis reported to cause pancreatic abscess, we believe selection of this organism occurred because surgical drainage was delayed while the patient was on the recommended treatment with meropenem. This case emphasizes the need for early surgical drainage of pancreatic abscesses to avoid the selection of such resistant pathogens. PMID- 15654183 TI - The core: where are we and where are we going? PMID- 15654184 TI - Analysis of real-time head accelerations in collegiate football players. AB - OBJECTIVE: To measure and analyze head accelerations during American collegiate football practices and games. METHODS: A newly developed in-helmet 6 accelerometer system that transmits data via radio frequency to a sideline receiver and laptop computer system was implemented. From the data transfer of these accelerometer traces, the sideline staff has real-time data including the head acceleration, the head injury criteria value, the severity index value, and the impact location. Data are presented for instrumented players for the entire 2003 football season, including practices and games. SETTING: American collegiate football. SUBJECTS: Thirty-eight players from Virginia Tech's varsity football team. MAIN OUTCOME MEASUREMENTS: Accelerations and pathomechanics of head impacts. RESULTS: : A total of 3312 impacts were recorded over 35 practices and 10 games for 38 players. The average peak head acceleration, Gadd Severity Index, and Head Injury Criteria were 32 g +/- 25 g, 36 g +/- 91 g, and 26 g +/- 64 g, respectively. One concussive event was observed with a peak acceleration of 81 g, a 267 Gadd Severity Index, and 200 Head Injury Criteria. Because the concussion was not reported until the day after of the event, a retrospective diagnosis based on his history and clinical evaluation suggested a mild concussion. CONCLUSIONS: The primary finding of this study is that the helmet-mounted accelerometer system proved effective at collecting thousands of head impact events and providing contemporaneous head impact parameters that can be integrated with existing clinical evaluation techniques. PMID- 15654185 TI - Height, weight, and BMI predict intra-articular injuries observed during ACL reconstruction: evaluation of 456 cases from a prospective ACL database. AB - OBJECTIVE: To identify demographic and anthropometric risk factors for intra articular (IA) injuries observed during ACL reconstruction. We hypothesize that significant associations exist among height, weight, and body mass index (BMI) with IA injuries when ACL tear occurs. DESIGN: This observational study of a prospective multi-investigator ACL database used logistic and Poisson regression analysis to assess independent predictors of IA injuries. SETTING: Vanderbilt Sports Medicine and affiliated tertiary care center. PATIENTS: Patients undergoing unilateral ACL reconstruction without prior injury to either knee were identified from a detailed prospective ACL reconstruction database. Four hundred fifty-six patients met inclusion/exclusion criteria. INTERVENTIONS: Per inclusion criteria, all patients underwent unilateral ACL reconstruction after assessment of injury profile. MAIN OUTCOME MEASUREMENTS: The ACL database was initiated in 1990 to identify demographic, anthropometric, and mechanistic variables associated with intra-articular injury. RESULTS: Height best predicted tibial and patellar damage. BMI better predicted medial femoral condyle lesions, whereas weight better predicted lateral and patellofemoral injury. BMI and weight equally predicted injury to menisci and medial tibial plateau. Through different outcomes, age (odd ratio [OR], 1.49; 95% CI, 1.02-2.16), height (OR, 2.66; 95% CI, 1.52-4.65), weight (OR, 1.02; 95% CI, 1.01-1.04), and BMI (OR, 1.24; 95% CI, 1.004-1.53) were all significant predictors of intra-articular injury. CONCLUSIONS: This is the first report using multivariable analysis of age, height, weight, and BMI to evaluate associations with IA injuries after ACL rupture observed during ACL reconstruction. We hypothesize that athletes possibly could reduce risk of certain IA pathologies with maintenance of lower body weight and BMI and thus potentially improve long-term functional outcomes after ACL reconstruction. PMID- 15654186 TI - Hip muscle weakness and overuse injuries in recreational runners. AB - OBJECTIVE: To test for differences in strength of 6 muscle groups of the hip on the involved leg in recreational runners with injuries compared with the uninvolved leg and a control group of noninjured runners. DESIGN: Descriptive analysis. SETTING: : Three outpatient physical therapy clinics in the Minneapolis/St. Paul metropolitan area. PARTICIPANTS: Thirty recreational runners (17 female, 13 male) experiencing a single leg overuse injury that presented for treatment between June and September 2002. Thirty noninjured runners (16 female, 14 male) randomly selected from a pool of 46 volunteers from a distance running club served as controls. MAIN OUTCOME MEASURES: Self-report demographic information on running habits, leg dominance demonstrated by preferred kicking leg, and injury information. Muscle strength of the 6 major muscle groups of the hip was recorded using a hand-held dynamometer. The highest value of 2 trials was used, and strength values were normalized to body mass(2/3). RESULTS: Results comparing the injured and noninjured groups showed that leg dominance did not influence the leg of injury (chi(2)(1) = 0.134; P = 0.71). Correlations for internal reliability of muscle measurements between trials 1 and 2 with the hand held dynamometer ranged from 0.80 to 0.90 for the 6 muscle groups measured, and all P values were less than 0.0001. No significant side-to-side differences in hip group muscle strength were found in the noninjured runners (P = 0.62-0.93). Among the injured runners, the injured side hip abductor (P = 0.0003) and flexor muscle groups (P = 0.026) were significantly weaker than the noninjured side. In addition, the injured side hip adductor muscle group was significantly stronger (P = 0.010) than the noninjured side. Duration of symptoms was not a contributing factor to the extent of injury as measured by muscle strength imbalance between injured and uninjured sides. CONCLUSIONS: Although no cause-and-effect relationship has been established, this is the first study to show an association between hip abductor, adductor, and flexor muscle group strength imbalance and lower extremity overuse injuries in runners. Because most running injuries are multifaceted in nature, areas secondary to the site of pain, such as hip muscle groups exhibiting strength imbalances, must also be considered to gain favorable outcomes for injured runners. The addition of strengthening exercises to specifically identified weak hip muscles may offer better treatment results in patients with running injuries. PMID- 15654187 TI - Awareness of genital health in young male athletes. AB - OBJECTIVE: To assess young male athletes' understanding of the need for a genital examination during the sports physical, their knowledge of signs and symptoms of serious testicular pathology, and the type of genital protection they wear for specific sports. Additionally, to identify common sport-specific genital injuries requiring emergency department treatment. DESIGN: Descriptive survey. SETTING: Station-based mass screening preparticipation physical exams performed in northeastern Ohio. PARTICIPANTS: A total of 755 male athletes at the junior high school, high school, and collegiate levels (age 12-25 years). MAIN OUTCOME MEASUREMENTS: Participants' answers to a 5-question survey. Male genital injury occurring in common contact and collision sports identified from the National Electronic Injury Surveillance System. RESULTS: Fifty percent did not understand why the genital examination is done. Forty-seven percent did not wear genital protection during sports. Most were unaware of the risk of testicular cancer and did not appreciate the difference in urgency of seeking medical treatment of painless versus painful testicular swelling. Approximately 20% of emergency department visits for sports-related genital trauma had a risk of permanent injury, and up to 7% resulted in hospitalization or transfer. CONCLUSIONS: Young male athletes are unaware of testicular pathology that affects their age group, and many do not wear genital protection during sports. Better education of young males regarding genital health and protection is necessary. PMID- 15654188 TI - Repeated exposure of tendon to prostaglandin-E2 leads to localized tendon degeneration. AB - OBJECTIVE: To determine whether repeated exposure of rabbit patellar tendon to prostaglandin-E(2) leads to degenerative changes in the tendon. SETTING: Laboratory animal study. MAIN OUTCOME MEASURES: Intratendinous changes including cellularity, matrix organization, collagen fibril packing, and diameter. METHODS: Skeletally mature New Zealand White rabbits (n = 10) were transcutaneously injected in the midsubstance of the patellar tendon with prostaglandin-E(2) (PGE(2); 50 ng or 500 ng). The contralateral tendons were used as 3 different controls (no injection, saline injection, and needlestick only). The injection was repeated once a week for 4 weeks, and the rabbits were killed 1 week after the last injection. The patellar tendons were harvested and examined using hematoxylin and eosin staining and transmission electron microscopy. RESULTS: Compared with the control groups, tendons exposed to PGE(2) by injection showed focal areas of hypercellularity, loss of normal tissue architecture, and focal areas of tendon disorganization and degeneration. Tendons injected with PGE(2) exhibited loosely organized collagen fibrils and had thinner collagen fibril diameter compared with control tendons (P < 0.005). Tendons injected with 500 ng PGE(2) appeared to be more disorganized and degenerated than those injected with 50 ng PGE(2). CONCLUSIONS: Repeated exposure of the tendon to PGE(2) leads to degenerative changes within the tendon. CLINICAL RELEVANCE: It is known that human tendon fibroblasts produce PGE(2) in vitro and in vivo in response to repetitive mechanical loading. This study demonstrates that repetitive exposure of the tendon to PGE(2) can result in degenerative changes within the tendon. Therefore, PGE(2) produced by tendon fibroblasts in response to repetitive mechanical loading in vivo might contribute to the development of exercise induced tendinopathy. PMID- 15654190 TI - Little league elbow. PMID- 15654189 TI - A prospective study of injuries sustained during competitive Muay Thai kickboxing. AB - OBJECTIVE: To investigate prospectively the type and rate of injuries sustained during amateur Muay Thai competition. DESIGN: Prospective cohort study collection of data following clinical examination. SETTING: Amateur Muay Thai competitions in the United Kingdom organized by the International Amateur Muay Thai Federation. PARTICIPANTS: Amateur Muay Thai boxers. Both sexes. Lightweight to super heavyweight. RESULTS: There were 92 participants, 12 females and 82 males. The average age was 17.3 years, and the average previous number of bouts was 3.9. A total of 588.5 minutes of competition time was assessed during a total of 10 events. Injury rates were 1.3 injuries per 100 minutes of competition in the lightweight category, 2.25 per 100 minutes of competition in the middleweight category, 30 per 100 minutes of competition in the heavyweight category, and 2.54 per 100 minutes of competition in the super heavyweight category. CONCLUSIONS: Compared with other reported martial arts, the injury rates are higher in Muay Thai. The head was shown to be the most common site of injury in amateur fighters, but there was an almost complete absence of lower limb injuries, which again is at variance with reported figures for other martial arts. PMID- 15654191 TI - Traumatic pulmonary pseudocyst with hemopneumothorax in a football player. PMID- 15654193 TI - Right ventricular dysfunction in chronic dilated cardiomyopathy and heart failure. AB - In patients with advanced heart failure, the main focus has traditionally been placed on the functional assessment of the left ventricle. Therefore, the current body of literature examining the right ventricle and its influence on the pathophysiological processes in heart failure has been limited. Methods currently employed to assess the size and function of the right ventricle include: high frequency thermodilution, contrast ventriculography, radionuclide ventriculography, echocardiography, and magnetic resonance imaging; however, none of these techniques has proven to be a 'gold standard' for the assessment of right ventricular function. Nevertheless, when these methods are employed, right ventricular dysfunction has been shown to be a powerful predictor of reduced exercise capacity and survival. This relationship holds true for patients with heart failure secondary to either ischemic or non-ischemic dilated cardiomyopathy. PMID- 15654195 TI - The remarkable right ventricle of patients with Eisenmenger syndrome. AB - Patients with Eisenmenger syndrome do much better than patients with all other etiologies of severe pulmonary hypertension. The primary reason, and a major focus of this review, is the right ventricle, a remarkable chamber with preserved systolic function despite a lifetime of systemic level pulmonary hypertension. PMID- 15654196 TI - Right ventricular failure following cardiac surgery. AB - The management of right ventricular failure remains a significant challenge in cardiac surgery. To date, no single clinical study has encompassed all of the principles that can be called upon to manage this problem. This review deals with the physiological management components that may be of value to cardiac medical and surgical clinicians. PMID- 15654194 TI - The right ventricle in pulmonary hypertension. AB - Pulmonary arterial hypertension (PAH) is a term used to classify a variety of conditions that share in common an injury to the pulmonary vasculature that produces elevations in pulmonary arterial pressure. However, it is the integrity of right ventricular function, rather than the degree of vascular injury, that is the major determinant of symptoms and survival in PAH. The article will review the normal structure and function of the right ventricle and summarize the impact of PAH and its treatments on right ventricular function. PMID- 15654197 TI - Increased tissue-type plasminogen activator: a facade in the fibrinolytic system in type 2 diabetes. AB - To determine whether the increased concentration in blood of total tissue-type plasminogen activator (t-PA), accompanying increased plasminogen activator inhibitor type-1 (PAI-1) seen with diabetes reflects increased free t-PA and hence activity or simply increased t-PA complexed with PAI-1, we measured each in 39 people with type 2 diabetes. The increased total t-PA occurred in the absence of an increase in t-PA activity detectable in venous blood samples despite the increase in the concentration of total t-PA protein. BACKGROUND: Type 2 diabetes is known to be associated with increased concentrations in blood of total tissue type plasminogen activator (t-PA) [free t-PA plus t-PA complexed with plasminogen activator inhibitor type-1 (PAI-1)] and PAI-1. This study was performed to determine whether the increased total t-PA is indicative of increased t-PA activity attributable to free t-PA or whether it is simply a manifestation of compensatory processes maintaining t-PA activity detectable in venous blood samples at normal or subnormal levels. METHODS: Total t-PA, free t-PA, and PAI-1 antigen and activity were measured by enzyme-linked immunosorbent assay and immunofunctional chromogenic substrate kinetic assays in peripheral venous blood samples obtained under fasting conditions from 39 people with type 2 diabetes. RESULTS: The results indicate that both PAI-1 and total t-PA antigen concentrations were increased in association with diabetes but the concentration of free t-PA and hence the t-PA activity was not. Thus, the increased total t-PA is a facade potentially masking the impaired fibrinolytic system activity associated with type 2 diabetes. CONCLUSIONS: The results indicate that the increase in the concentration of total t-PA protein in blood of people with diabetes occurs in the absence of an increase in free t-PA. Hence t-PA activity detectable in peripheral venous blood samples does not increase and may even decline modestly. PMID- 15654198 TI - High prevalence of metabolic syndrome among young women with premature coronary artery disease. AB - BACKGROUND: The metabolic syndrome is more prevalent with the use of the recently defined National Cholesterol Education Program Adult Treatment Panel III (NCEP ATP III) criteria and is associated with a greater risk of atherosclerotic cardiovascular disease than any of its individual components. This study evaluated the prevalence of metabolic syndrome in female and male patients with newly diagnosed premature coronary artery disease. METHOD: The study population included 582 consecutive patients (496 men, 86 women) with newly diagnosed premature coronary artery disease (aged < or =45 years). Besides classic major coronary risk factors, all patients were evaluated for the presence of metabolic syndrome based on the NCEP ATP III criteria. RESULTS: The majority of patients were male (85% versus 15%). The overall prevalence of metabolic syndrome was 37%. Women with premature coronary artery disease were found to have a higher prevalence of metabolic syndrome than men (73% versus 31% respectively, p < 0.001). Furthermore, the mean number of components of metabolic syndrome was significantly higher in women compared to men (2.81 +/- 1.09 versus 1.85 +/- 1.08 respectively, p < 0.001). In addition, metabolic syndrome was detected to be the most frequent coronary risk factor in women (73%). Besides, cigarette smoking was found to be significantly higher in males compared to females (70% versus 36% respectively, p < 0.001) and it was the most prevalent coronary risk factor in men with premature coronary artery disease. CONCLUSION: We have shown for the first time a higher prevalence of metabolic syndrome in young females compared with young males with premature coronary artery disease. This data may be useful in directing primary and secondary preventive measures. PMID- 15654199 TI - Distinct mouse coronary anatomy and myocardial infarction consequent to ligation. AB - This study describes the coronary anatomy of C57BL/6 mice and the functional impact of that anatomy on myocardial infarctions induced by ligation of the left coronary artery. In contrast to humans, a distinct septal coronary artery was observed in all mice arising either from a separate ostium from the right sinus of Valsalva or as a branch of the right coronary artery. Ligation of the left coronary artery at the site of its emergence from under the left atrium resulted in reproducible large myocardial infarctions involving the anterolateral, posterior, and apical regions of the heart as evidenced by histology and echocardiography. Interestingly, the septum was spared from infarction as predicted by the presence of the distinct septal branch found in mice. Thus, the distinct mouse coronary artery anatomy results in different regionality of infarction compared with man and large laboratory animals, and this may affect on the associated pathological remodeling of the heart. BACKGROUND: The objective of this study was to describe the coronary anatomy of C57BL/6 mice and determine the functional impact of that anatomy on myocardial infarctions induced by ligation of the left coronary artery. METHODS: C57BL/6 mice were used to visualize mouse coronary anatomy and to generate myocardial infarction. Mouse coronary artery visualization was performed on isolated hearts using injection of Silastic sealant into the aortic arch. After left thoracotomy myocardial infarction was produced by ligating the left coronary artery at the site of the vessels' emergence past the tip of the left atrium. Echocardiography was performed to analyze heart function, and histology was performed to delineate myocardial infarction. RESULTS: The different septal coronary artery was observed in all mice arising either from a separate ostium from the right sinus of Valsalva or as a branch of the right coronary artery. The mouse left coronary artery passed obliquely across the left ventricular free wall similar to the ramus intermedius coronary artery variant in man. Ligation of the left coronary artery as it emerged from under the left atrium resulted in reproducible large infarctions involving the anterolateral, posterior, and apical regions of the heart as demonstrated by histology and echocardiography. Notably, the septum was spared from infarction as predicted by the presence of the distinct septal coronary artery found in mice. CONCLUSIONS: Mouse coronary anatomy is distinct compared to man or large laboratory animals, and myocardial infarctions resulting from ligation of the mouse left coronary artery spare the septum, which may affect the pathological remodeling of the heart. PMID- 15654201 TI - Changes in demographic factors and mortality after out-of-hospital cardiac arrest in Sweden. AB - OBJECTIVE: To describe changes between 1992 and 2003 in age, sex, factors at resuscitation and survival among patients suffering from out-of-hospital cardiac arrest in Sweden. METHODS: This was a prospective observational study including various ambulance organizations in Sweden. Patients suffering from out-of hospital cardiac arrest between 1992 and 2003 included in the Swedish Cardiac Arrest Registry were followed for survival to 1 month. RESULTS: In all 19 791 cases took part in the survey. There was a slight increase in mean age from 68 to 70 years (P = 0.025) and an increase of females from 29 to 32% (P = 0.0001). There was a change in witnessed status (P < 0.0001) with an increase in crew witnessed cases and a decrease in non-witnessed cases. There was a decrease in cases of a cardiac etiology from 75 to 61% (P < 0.0001) and a decrease in the percentage found in ventricular fibrillation from 36 to 25% (P < 0.0001). When crew-witnessed cases were excluded the proportion receiving bystander cardiopulmonary resuscitation (CPR) increased from 30 to 42% (P < 0.0001). There was a slight increase in the overall proportion of patients hospitalized alive from 16 to 20% (P = 0.032). There was no significant change in the overall proportion of survivors at 1 month after cardiac arrest (4.5% in 1992 and 5.0% in 2003). CONCLUSIONS: Among patients suffering from out-of-hospital cardiac arrest in Sweden some changes took place. The most important ones were a decrease in the proportion of patients found in ventricular fibrillation and an increase in the proportion of patients receiving bystander CPR. The proportion of patients admitted alive to hospital increased moderately, whereas the proportion of patients alive after 1 month remained unchanged. PMID- 15654200 TI - Plasma soluble adhesion molecules; intercellular adhesion molecule-1, vascular cell adhesion molecule-1 and E-selectin levels in patients with isolated coronary artery ectasia. AB - Plasma soluble adhesion molecules, intercellular adhesion molecule-1 (ICAM)-1, vascular cell adhesion molecule-1 (VCAM-1) and E-selectin leves of patients with isolated coronary artery ectasia (CAE), patients with obstructive coronary artery disease without CAE and subjects with angiographically normal coronary arteries were evaluated. Patients with isolated CAE were detected to have significantly higher levels of plasma soluble ICAM-1, VCAM-1 and E-selectin in comparison with patients with obstructive coronary artery disease without CAE (ICAM, 673 +/- 153 versus 381 +/- 106, respectively, P < 0.001; VCAM-1, 2366 +/- 925 versus 1136 +/- 208, respectively, P < 0.001; E-selectin, 74 +/- 21 versus 61 +/- 18, respectively, P = 0.01) and subjects with normal coronary arteries (ICAM-1, 673 +/- 153 versus 303 +/- 131, respectively, P < 0.001; VCAM-1, 2366 +/- 925 versus 729 +/- 231, respectively, P < 0.001; E-selectin, 74 +/- 21 versus 49 +/- 9, respectively, P < 0.001), suggesting the presence of a more severe and extensive chronic inflammation in the coronary circulation in patients with isolated CAE. BACKGROUND: The common coexistence of coronary artery ectasia (CAE) with coronary artery disease (CAD) suggests that it may be a variant of CAD. However, it is not clear why some patients with obstructive CAD develop CAE whereas most do not. Inflammation has been reported to be a major contributing factor to both obstructive and aneurysmatic vascular disorders and therefore, in the present study, the plasma soluble adhesion molecules, intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1) and E-selectin levels in isolated CAE were investigated. METHODS: The study population consisted of three groups: the first consisted of 32 patients with isolated CAE without stenotic lesion; the second of 32 patients with obstructive CAD without CAE; and the third group of 30 control subjects with normal coronary arteries. Coronary diameters were measured as the maximum diameter of the ectasic segment by use of a computerized quantitative coronary angiography analysis system. According to the angiographic definition used in the Coronary Artery Surgery Study, a vessel is considered to be ectasic when its diameter is > or =1.5 times that of the adjacent normal segment in segmental ectasia. Plasma soluble ICAM-1, VCAM-1 and E selectin levels were measured in all patients and control subjects using commercially available enzyme-linked immunosorbent assay kits. RESULTS: Patients with isolated CAE were found to have significantly higher levels of plasma soluble ICAM-1, VCAM-1, and E-selectin in comparison with patients with obstructive CAD without CAE (ICAM, 673 +/- 153 versus 381 +/- 106, respectively; P < 0.001; VCAM-1, 2366 +/- 925 versus 1136 +/- 208, respectively; P < 0.001; E selectin, 74 +/- 21 versus 61+/-18, respectively; P = 0.01) and control subjects with normal coronary arteries (ICAM-1, 673 +/- 153 versus 303 +/- 131, respectively;, P < 0.001; VCAM-1, 2366 +/- 925 versus 729 +/- 231, respectively; P < 0.001; E-selectin, 74 +/- 21 versus 49 +/- 9, respectively; P < 0.001). In addition, we detected statistically significant positive correlation between the total length of ectasic segments and the levels of plasma soluble ICAM-1 (r = 0.625; P < 0.001), VCAM-1 (r = 0.548; P = 0.001) and E-selectin (r = 0.390; P = 0.027). Multivariate logistic regression analysis revealed a significant independent relation between isolated CAE and ICAM-1 [odds ratio (OR) = 1.023; 95% confidence interval (CI) = 1.0048-1.0414; P = 0.0129] and VCAM-1 (OR = 1.0057; 95% CI = 1.0007-1.0106; P = 0.0240). CONCLUSIONS: We have shown that patients with isolated CAE have raised levels of plasma soluble ICAM-1, VCAM-1 and E-selectin in comparison with patients with obstructive CAD without CAE and control subjects with normal coronary arteries, suggesting the presence of a more severe and extensive chronic inflammation in the coronary circulation in these patients. PMID- 15654202 TI - Early release of neutrophil markers of activation after direct stenting in patients with unstable angina. AB - OBJECTIVE: To assess polymorphonuclear neutrophils activation after stenting in acute coronary syndromes studied by myeloperoxydase, lactoferrin and elastase release in this clinical setting. METHODS: Myeloperoxydase, lactoferrin, elastase, C-reactive protein and cytokines serum levels were assessed in 20 patients undergoing catheterization for unstable angina. Serial sampling starting before arteriography and continued up to 24 h was carried out in 15 patients undergoing direct stenting (group A) and in five patients assessed by coronary angiography only (group B). RESULTS: Myeloperoxydase, lactoferrin and elastase levels remained unchanged following catheterization, whereas a significant increase in myeloperoxydase (P = 0.0009) and lactoferrin (P = 0.004) was observed after stenting. No change in levels of tumour necrosis factor alpha, interleukin (IL)-8 and IL-11 was found in group B after catheterization at the different sampling times, although IL-8 and IL-11 levels increased transiently following stenting. IL-6 values increased in both groups. Baseline values of C-reactive protein were similar in each group. A progressive increase in C-reactive protein was noted in both groups and appeared to be larger following stenting (group A: P = 0.0002; group B: P = 0.01). CONCLUSIONS: In patients with unstable angina, stenting is associated by immediate neutrophil activation followed by release of inflammatory cytokines (IL-6, IL-8, IL-11) and C-reactive protein elevation. This study points out a potential role of myeloperoxydase as a trigger for inflammatory reaction in patients with unstable coronary syndromes undergoing percutaneous coronary intervention. PMID- 15654204 TI - Bibliography of current world literature. PMID- 15654203 TI - The vascular effects of doxazosin in hypertension complicated by metabolic syndrome. AB - AIMS: To evaluate the vascular effects of doxazosin, an alpha-1 antagonist, in hypertensive patients with metabolic syndrome in whom the drug has previously been shown to exert beneficial metabolic actions on lipids and insulin metabolism. EXPERIMENTAL PROTOCOL: Twelve untreated non-diabetic hypertensive patients with National Cholesterol Education Program (NCEP) ATP-III defined metabolic syndrome were assigned to three-months of treatment with doxazosin (5.5 +/- 1.9 mg/die). Study variables were measured at baseline and after treatment. End-points: forearm blood flow (strain-gauge plethysmography) responses to graded intra-arterial acetylcholine and sodium nitroprusside infusion to test endothelium-dependent and independent vasodilatation respectively. Minimum forearm vascular resistance, the ratio of mean blood pressure and post-ischaemic maximal blood flow, as an index of arteriolar structure; transcapillary albumin escape rate (the 1-h decay rate of I-albumin, 6-8 microC ev) as a measure of systemic capillary permeability. Lipids, fasting and post-glucose insulin were measured at baseline and after treatment. RESULTS: Doxazosin reduced blood pressure, augmented acetylcholine-mediated vasodilatation, decreased minimum resistance and, although not to a statistically significant extent, transvascular albumin leakage increased high-density lipoprotein (HDL) cholesterol while triglycerides and post-stimulative hyperinsulinemia decreased. CONCLUSIONS: Doxazosin improved endothelial-mediated vasomotor function and reversed abnormal arteriolar structure in hypertensive patients with metabolic syndrome while improving lipid profile and blunting post-glucose hyperinsulinemia. PMID- 15654207 TI - New paradigm for the roles of fungi and eosinophils in chronic rhinosinusitis. AB - PURPOSE OF REVIEW: Chronic rhinosinusitis represents a challenge with its poorly understood pathophysiology and limited treatment options. Potential roles of fungi and eosinophils in the etiology and pathophysiology of chronic rhinosinusitis are summarized. RECENT FINDINGS: Previously, the fungal role in chronic rhinosinusitis was limited to the rare subgroup, allergic fungal rhinosinusitis. Critical examination of earlier diagnostic criteria for allergic fungal rhinosinusitis reveals limitations. By using updated diagnostic standards and novel sensitive techniques to detect fungi, a higher number of patients can now be diagnosed with fungal rhinosinusitis. A novel non-IgE-mediated immunologic mechanism in chronic rhinosinusitis patients links the predominant eosinophilic inflammation to certain fungi. Overall, these new findings have implications for surgical and medical approaches, including anti-inflammatory and antifungal medications. SUMMARY: Several classification schemes and diagnostic criteria describe chronic rhinosinusitis and make comparisons difficult. Preselection of patient groups within the chronic rhinosinusitis population and the lack of sensitive diagnostic techniques have prevented a full understanding of the syndrome complex of chronic rhinosinusitis. New results suggest a broader role for fungi in the pathophysiology of chronic rhinosinusitis, linking the eosinophilic inflammation to the presence of certain molds in the nasal and paranasal cavities. Although fungi are commonly found in nearly everyone, only chronic rhinosinusitis patients respond to them with an eosinophilic inflammation. These findings support a shift in the etiologic understanding of chronic rhinosinusitis away from a bacteriologic infectious pathogenesis to a fungal-driven inflammatory pathophysiology. Herein, the authors review earlier studies and describe an updated view on an old paradigm. PMID- 15654208 TI - Role of fungus in eosinophilic sinusitis. AB - PURPOSE OF REVIEW: Chronic rhinosinusitis is characterized by a broad diagnostic and etiologic spectrum. It has been postulated that fungal organisms might represent the immunologic target initiating and maintaining the disease process in patients with chronic rhinosinusitis as a common denominator. This review analyzes the available data to describe the current understanding of the role of fungus in the pathophysiology of chronic rhinosinusitis. RECENT FINDINGS: Recent findings have demonstrated that using highly sensitive methods, the detection rate of fungi in nasal mucin can be increased to approximately 100% in patients and in healthy controls. If this ubiquitous fungal contamination were to be related to a causative mechanism in chronic rhinosinusitis, there would have to be an immunologic sensitization in patients in contrast with healthy individuals, which is yet not fully understood. However, studies were able to demonstrate cytokine production in blood-derived lymphocytes in the presence of fungi in patients, but not in healthy controls. Several questions arise. If this is a relevant finding, in what percentage of chronic rhinosinusitis patients could it be causative? Is chronic rhinosinusitis, then, a systemic disease? How is the extramucosal target in the mucin presented to the immune system? Is intranasal, topical antifungal treatment a successful option? SUMMARY: The role of fungal organisms in the etiology of eosinophilic rhinosinusitis is reviewed with respect to novel findings, and arising questions are discussed in the light of recent investigations. PMID- 15654209 TI - Effect of topical nasal steroid sprays on nasal mucosa and ciliary function. AB - PURPOSE OF REVIEW: This review was conducted to examine current evidence focusing on the effect of nasal steroid sprays on nasal ciliary function. RECENT FINDINGS: Review of current literature suggests that long term effects of topical corticosteroid nasal sprays are safe and fail to cause damage to local nasal structure and function. SUMMARY: The use of corticosteroid nasal sprays for the treatment of allergic rhinitis is widely accepted. Popularity of this class of medications is based on a well-established combination of efficacy, tolerability, and safety. Although current literature suggests that the use of intranasal steroids is indeed safe, increasing indications for prolonged administration continue to fuel debate regarding the long-term effect on local nasal structure and function. The purpose of this article is to review current literature addressing the effects of the components of local intranasal steroid sprays on the structure and function of the nasal mucosa. PMID- 15654210 TI - The low skull base-is it important? AB - PURPOSE OF REVIEW: Little is published regarding variations of skull-base anatomy and their relevance to the performance of endoscopic sinus surgery. Several catastrophic complications have occurred in patients with low-lying skull-base variation. This review's purpose is to make the reader aware of skull-base variation, their recognition, and a surgical plan for sinus surgery in these patients. RECENT FINDINGS: Only a few papers are available for review discussing variations of the low-skull base. Recent findings in reviewing multiple CT scan indicate several variations of the skull. It has also been noted that there are variations of thickening in the skull (ie, lateral ethmoid, thicker bone, more medial, thinner bone). Several case reports have surfaced reflecting how a low lying skull-base can play a role in brain-related complications. These figures are reviewed. SUMMARY: A knowledge of orbital and skull-base variations preoperatively can help plan the surgical procedure and avoid major complication. The preoperative CT scan is the key to preoperative knowledge. The use of special techniques to aid in localization of a low-lying skull-base including image computer guidance is recommended. PMID- 15654211 TI - Powered instrumentation and tissue effects in the nose and paranasal sinuses. AB - PURPOSE OF REVIEW: The technologic cornerstones of endoscopic sinus surgery initially rested on the endoscope and CT scanning. In the mid-1990s, the advent of powered instrumentation, in particular the microdebrider, advanced the surgeon's ability to treat polypoid disease, sinus and nasal disease in a bleeding field, choanal atresia, antral choanal polyps, and inferior turbinate hypertrophy. This review evaluates both positive and negative tissue effects secondary to powered instrumentation. RECENT FINDINGS: Recent reports of deleterious outcomes with powered instrumentation require a reevaluation of the role of powered instrumentation and consideration of factors that lead to injury. The rapidity of injury makes the powered instrument especially dangerous in inexperienced hands. One third of participants in a recent survey were unaware that powered instruments could be involved in orbital injury and half were unaware that cranial injuries could be secondary to powered instrumentation. SUMMARY: Microdebriders will continue to advance the field of endoscopic surgery, providing clearer operative fields and causing less tissue trauma in experienced hands. However, the severity of complications, including the potential for rapidly aspirating orbital and cerebral contents when laminae are violated, must be appreciated and respected. PMID- 15654212 TI - Image-guided surgery: what is the accuracy? AB - PURPOSE OF REVIEW: Use of image-guided surgery (IGS) systems in otolaryngology, particularly rhinology, has grown exponentially in recent years. Central to their use is the understanding of the accuracy of each system. The purpose of this review is to discuss the error inherent in all IGS systems. A standardized technique (currently used in the engineering literature) for understanding and reporting error in IGS systems is reviewed. Using this technique, the error of commercially available IGS systems is reviewed. RECENT FINDINGS: The most commonly used IGS systems depend on the conformation of the skin, as opposed to relying on bone-implanted devices. For these systems, mean accuracies 2 mm or less are routinely reported. This finding is independent of fiducial markers (eg, proprietary headsets, skin-affixed markers, or laser scanning of skin surfaces). Techniques of fiducial localization and registration of CT scans to intraoperative anatomy are proprietary to each company. As such, there is great variability in reporting system specifications-particularly error of IGS systems. This lack of standardization makes comparison of one system to another difficult if not impossible. SUMMARY: Image-guided surgery systems commonly used in rhinology report mean accuracies of 2 mm or less. Surgeons must be aware that this value represents a mean of a distribution of errors. As such, 95% of the time error can be expected to be less than approximately 1.7 times its mean value. However, outliers (errors much larger and much smaller than the mean) may exist for each IGS intervention. As noted, IGS systems function to complement-not replace-knowledge of surgical anatomy. PMID- 15654213 TI - Endoscopic sinus surgery: where we are-and where we're going. AB - PURPOSE OF REVIEW: Improved understanding of sinonasal mucociliary function as well as technological advancements have led to a renaissance in the management of sinonasal disease. The current techniques, and adjunctive intraoperative technologies, have allowed for a more precise and safer surgical dissection, extending this surgical application to include the routine endoscopic management of skull base lesions and sinonasal malignancies. The anatomic boundaries of the nasal endoscope continue to be redefined. This paper will review the recent advances in the field of surgical rhinology as well as discuss directions for the future. RECENT FINDINGS: Advancements in imaging technology including image acquisition, three-dimensional reconstruction, stereotactic navigation, and CT MRI fusion have aided in more thorough preoperative planning, enabling safer and more precise endoscopic surgical dissection. Refinement of endonasal powered instrumentation including microdebriders and high-speed suction irrigation drills have minimized trauma to normal tissues and accelerated postoperative healing. Increased experience with the endoscope has broadened the endoscopic ventral skull base exposure from the odontoid process to foramen ovale to the olfactory bulb. SUMMARY: Endoscopic sinus surgery is no longer exclusively for the management of chronic rhinosinusitis and nasal polyposis. Sinonasal malignancies, as well as anterior skull base lesions have become part of the rhinologist's responsibility. Furthermore, selective lesions managed through traditional craniotomies may now be accessed via trans-sinonasal transcranial endoscopic routes. PMID- 15654214 TI - Superantigen hypothesis for the early development of chronic hyperplastic sinusitis with massive nasal polyposis. AB - PURPOSE OF REVIEW: The pathogenesis, pathophysiology, and immunobiology of chronic hyperplastic sinusitis with massive nasal polyposis are starting to become unraveled. Allergy, viral infection, bacterial infection, fungal infection, and environmental pollution have all been suggested as possible initial triggers that may upregulate inflammation of the lateral wall of the nose to develop nasal polyposis. The purpose of this review is to present data from our laboratory that suggest that one of the possible early events in the development of inflammation of the lateral wall of the nose in chronic hyperplastic sinusitis with massive nasal polyposis is the production of exotoxins from Staphylococcus aureus. The exotoxins may act as superantigens and cause activation and clonal expansion of lymphocytes with specific Vbeta regions, resulting in massive cytokine production. RECENT FINDINGS: Recent published studies suggest that S. aureus is the most common organism isolated from the mucus adjacent to massive nasal polyposis. Staphylococci produce exotoxins. These exotoxins, sometimes known as enterotoxins, include SEA, SEB, and TSST-1. These exotoxins are capable of acting as superantigens and therefore, reacting with T lymphocytes with specific Vbetas in the lateral wall of the nose. Thereafter, it is possible that these lymphocytes are stimulated to produce both TH1 and TH2 cytokines, which have also been demonstrated in the nasal polyp. The consequence of these findings may be the upregulation and increased survival of eosinophils in the nasal polyp. SUMMARY: Staphylococcus aureus is present in the mucin adjacent to nasal polyps in about 60 to 70% of cases of massive nasal polyposis. These organism, as studied up to the present, always produce exotoxins, which may act as superantigens, causing activation and clonal expansion of lymphocytes with specific Vbeta region in the lateral wall of the nose. The present review suggests that activation of these lymphocytes produce both TH1 and TH2 cytokines. The potential damage to the nasal mucosa from eosinophils is briefly discussed. Theoretically, topical antibiotics to suppress the colonization of S. aureus may be a logical approach to downregulate the production of superantigen in the lateral wall of the nose after appropriate endoscopic sinus surgery. PMID- 15654215 TI - Endoscopic sinus surgery rescue of failed osteoplastic flap with fat obliteration. AB - PURPOSE OF REVIEW: The modified endoscopic Lothrop procedure has been used as an alternative to osteoplastic flap with obliteration in the management of chronic frontal sinusitis. More recently it has been used to rescue cases of failed osteoplastic flap. This article reviews the recent literature regarding endoscopic rescue of failed osteoplastic flap with obliteration. RECENT FINDINGS: Endoscopic sinus surgery is indicated in cases of osteoplastic flap failure when there is objective evidence of mucocele formation. All patients require CT and MR imaging preoperatively. Clinical improvement can be achieved in up to 89% of patients in the first 12 to 18 months postoperatively. SUMMARY: The modified endoscopic Lothrop procedure is effective in the management of failed osteoplastic flap with obliteration in the short term. Long-term evaluation of this technique is still required. Endoscopic rescue is technically challenging and requires an experienced surgeon, ideally with access to image guidance equipment. PMID- 15654216 TI - Endoscopic sinus surgery for benign and malignant nasal and sinus neoplasm. AB - PURPOSE OF REVIEW: Endoscopic sinus surgery has become widely accepted as the standard of treatment for chronic inflammatory diseases of the paranasal sinuses unresponsive to medical treatment. With increased skill with endoscopic surgical technique, advanced technologies such as intraoperative imaging systems, and a better understanding of the complex anatomy of the paranasal sinuses and surrounding vital structures, many otolaryngologists have increasingly applied their expertise in endoscopic sinus surgery to the resection of nasal and sinus neoplasms. The following represents a review of the recent literature on the latest trends regarding endoscopic resection of nasal and paranasal sinus neoplasms. RECENT FINDINGS: There has been an increasing popularity in the removal of nasal and paranasal sinus neoplasms through an endoscopic approach. In the recent literature, emphasis has been on the endoscopic surgery of benign tumors, especially inverted papilloma and nasopharyngeal angiofibroma. Other benign neoplasms have also been reported, but only in small case reports. There have been a few recent reports supporting endoscopic removal of malignant neoplasms as well. However, the follow-up is too short and study groups too small to make definitive conclusions. SUMMARY: In the hand of experienced and skilled surgeons, complete endoscopic removal is attainable in most cases. Especially for the more common benign neoplasms, such as inverted papilloma and early stage angiofibroma. En bloc resection is not necessary to achieve oncologic cure. However, several factors have to be considered before selection of this surgical approach. Large tumor size, intracranial or orbital extension, and extensive frontal or infratemporal fossa involvement are relative, but not absolute limitations. PMID- 15654217 TI - Alternative medical treatment strategies for chronic hyperplastic eosinophilic sinusitis. AB - PURPOSE OF REVIEW: Chronic rhinosinusitis (CRS) is one of the most common chronic illnesses in the United States. Although CRS has been viewed traditionally as an infectious disease, treatment focused on antibiotics and surgery has not infrequently provided disappointing results. RECENT FINDINGS: Recently much of CRS has been shown to be an eosinophilic inflammatory disease and new anti inflammatory treatments are being studied. SUMMARY: This review discusses medical management for chronic hyperplastic eosinophilic sinusitis, including antifungal treatment, low-dose macrolide treatment, antilipid mediator therapy, and new immune-modifying treatments. PMID- 15654218 TI - Pediatric sinusitis: when do we operate? AB - PURPOSE OF REVIEW: The role of surgery in the treatment of pediatric sinusitis is still in evolution. This review of recent literature highlights developments in the study of pediatric sinusitis, particularly as it pertains to surgical intervention. RECENT FINDINGS: There is growing support in the literature for adenoidectomy as a first-line surgical intervention for chronic rhinosinusitis in children when maximal medical management fails. Maxillary sinus aspiration or middle meatal culture can be performed at the same sitting to facilitate directed antibiotic therapy. Intravenous antibiotics seem to be a promising alternative to functional endoscopic sinus surgery (FESS), especially in younger children. Current literature continues to support FESS as a safe and effective treatment for pediatric sinus disease. Previous notions that FESS may inhibit midfacial growth have been challenged by several recent studies. There is no clear consensus regarding timing of FESS for chronic rhinosinusitis. However, the current literature seems to support FESS when maximal medical therapy, adenoidectomy, and culture-directed systemic antibiotics have all failed with persistence of sinonasal disease, when anatomic abnormalities predispose to chronic rhinosinusitis by obstructing normal sinonasal drainage pathways, in sinonasal polyposis to facilitate application of topical steroids, as an adjunct to desensitization in aspirin-sensitive patients, when orbital or intracranial complications of sinonasal disease occur, and in selected cystic fibrosis patients to improve quality of life and facilitate application of topical antibiotics with activity against Pseudomonas aeruginosa. SUMMARY: Although the current literature lends some additional clarity to the indications for surgical intervention in pediatric chronic rhinosinusitis, additional research is still needed to elucidate appropriate timing for surgery and more specific indications. PMID- 15654219 TI - Endoscopic Lothrop procedure: when should it be considered? AB - PURPOSE OF REVIEW: The endoscopic Lothrop procedure has proved to be an effective intranasal approach to the treatment of frontal sinus disorders. When successful, the operation re-establishes normal or near-normal frontal sinus function (especially physiologic evacuation of frontal sinus contents). Recently, several authors have proposed the use of this procedure for the treatment of several disease processes in addition to chronic frontal sinusitis. RECENT FINDINGS: Although follow-up has been limited, the endoscopic Lothrop procedure has proved effective in the treatment of patients with the following: (1) failed prior endoscopic surgery, (2) frontal sinus mucocele, (3) frontoethmoid fractures, (4) failed osteoplastic flap frontal sinus obliteration, (5) resection of sinonasal neoplasms, and (6) removal of osteitic foci. SUMMARY: When used by an experienced endoscopic surgeon in selected clinical settings, the endoscopic Lothrop procedure offer acceptable to excellent results with relatively low morbidity and with several advantages over more standard surgical approaches for the treatment of specific disease entities. PMID- 15654220 TI - Bibliography. Current world literature. Nose and paranasal sinuses. PMID- 15654221 TI - Telegenetics: the next phase in the provision of genetics services? PMID- 15654222 TI - Genetic disorders involving molecular-chaperone genes: a perspective. AB - Molecular chaperones are important for maintaining a functional set of proteins in all cellular compartments. Together with protein degradation machineries (e.g., the ubiquitin-proteasome system), chaperones form the core of the cellular protein-quality control mechanism. Chaperones are proteins, and as such, they can be affected by mutations. At least 15 disorders have been identified that are associated with mutations in genes encoding chaperones, or molecules with features suggesting that they function as chaperones. These chaperonopathies and a few other candidates are presented in this article. In most cases, the mechanisms by which the defective genes contribute to the observed phenotypes are still uncharacterized. However, the reported observations definitely point to the possibility that abnormal chaperones participate in pathogenesis. The available data open novel perspectives and should encourage searches for new genetic chaperonopathies, as well as further analyses of the disorders discussed in this article, including detection of new cases. PMID- 15654223 TI - Obstacles and opportunities in meta-analysis of genetic association studies. AB - Genetic association studies have the potential to advance our understanding of genotype-phenotype relationships, especially for common, complex diseases where other approaches, such as linkage, are less powerful. Unfortunately, many reported studies are not replicated or corroborated. This lack of reproducibility has many potential causes, relating to study design, sample size, and power issues, and from sources of true variability among populations. Genetic association studies can be considered as more similar to randomized trials than other types of observational epidemiological studies because of "Mendelian randomization" (Mendel's second law). The rationale and methodology for synthesizing randomized trials is highly relevant to the meta-analysis of genetic association studies. Nevertheless, there are a number of obstacles to overcome when performing such meta-analyses. In this review, the impacts of Type I error, lack of power, and publication and reporting biases are explored, and the role of multiple testing is discussed. A number of special features of association studies are especially pertinent, because they may lead to true variability among study results. These include population dynamics and structure, linkage disequilibrium, conformity to Hardy-Weinberg Equilibrium, bias, population stratification, statistical heterogeneity, epistatic and environmental interactions, and the choice of statistical models used in the analysis. Approaches to dealing with these issues are outlined. The supreme importance of complete and consistent study reporting and of making data readily available is also highlighted as a prerequisite for sound meta-analysis. We believe that systematic review and meta-analysis has an important role to play in understanding genetic association studies and should help us to separate the wheat from the chaff. PMID- 15654224 TI - Telegenetics in Maine: Successful clinical and educational service delivery model developed from a 3-year pilot project. AB - PURPOSE: The goal of this 3-year pilot project was to increase accessibility to genetics educational and clinical services in Maine. METHODS: Southern Maine Genetics Services, Foundation for Blood Research in collaboration with Maine Telemedicine Services established telemedicine capacity to link with rural health care centers located in Northern, Central, and Southern Maine and public health nursing statewide for the provision of genetics clinical and educational services. Core partners included a rural family practice residency program, a rural pediatric practice in northern Maine, and public health nurses statewide. The telegenetics model created was based on development and implementation of a preventive and medical management technology solution, conducting a pilot study to collect data, and approaching insurance companies for reimbursement. Evaluation included surveys on the quality, acceptability, and usefulness of genetics services delivered via telemedicine, telephone interviews, and decision making confidence evaluations. RESULTS: During the project period, 24 rural clinical sites participated. In total, 93 presentations were given, and 125 patients were evaluated. Sixty-four percent of patients evaluated were pediatric. Despite site coordinator efforts to complete satisfaction surveys, the provider and patient response level was low (18% and 25%, respectively). Of those evaluations received, provider and patient response to telegenetics was positive. Decision-making confidence for genetics and neurology consultants was high. Our experience contributes to the development of telegenetics models that can be used in other rural states. PMID- 15654225 TI - Adjusting the estimated proportion of breast cancer cases associated with BRCA1 and BRCA2 mutations: public health implications. AB - PURPOSE: Mutations in BRCA1 or BRCA2 genes increase breast cancer risk. Assuring reliability of information about these mutations is increasingly important to the health care community; mutation testing is becoming more widespread. We describe a methodology for assessing such information. METHODS: Our approach integrates four interdependent epidemiologic parameters: (1) the probability of developing breast cancer, (2) the proportion of breast cancer cases with a BRCA1 or BRCA2 mutation, (3) the proportion of women that carries a mutation, and (4) the proportion of women with a mutation that develops cancer. We assess the plausibility of estimates of these parameters from published reports and commonly accessed information sources. RESULTS: Assuming a fixed probability of developing breast cancer, the following estimates for the other three epidemiologic parameters are derived for women by age 70: 1% to 2% of all breast cancer cases are associated with a BRCA1 or BRCA2 mutation; 1 in 300 to 1 in 465 women carry a mutation; and 35 to 65% of mutation carriers develop breast cancer. Within these ranges, however, only selected combinations are plausible. The proportion of mutation-related breast cancer is lower than listed in some common information sources (1 to 2% vs 6%). Also, penetrance is somewhat lower and the carrier rate somewhat higher. CONCLUSIONS: The four epidemiologic parameters can be integrated to test their plausibility. BRCA1 and BRCA2 mutations are associated with only one-third as many breast cancer cases in the general population as reported by commonly accessed information sources. PMID- 15654226 TI - An evaluation of BRCA1 and BRCA2 founder mutations penetrance estimates for breast cancer among Ashkenazi Jewish women. AB - PURPOSE: Three founder mutations in BRCA1 or BRCA2 genes increase breast cancer risk among Ashkenazi Jewish women. Reported estimates of the magnitude of this risk vary widely. We describe an integrated approach for assessing the plausibility of these estimates. METHODS: Our approach integrates four epidemiologic parameters: (1) the proportion of all breast cancer cases with a founder mutation, (2) the proportion of women that carry one of these mutations, (3) the proportion of women with a mutation that develops cancer, and (4) the number of women who will develop cancer, regardless of mutation status. We then assess the published estimates of the proportion of Ashkenazi Jewish women with a mutation that develops cancer in the context of the other three parameters. RESULTS: Penetrance for the founder mutations by ages 40, 50, and 70 are approximately 7%, 20%, and 40%, respectively. In two of the four published studies that evaluated at least two of the four parameters, penetrance estimates were internally consistent with the other three parameters and were also consistent with our consensus estimate. The third study had incomplete data. In the fourth study, the penetrance estimate was not internally consistent with the other three parameters, nor was it consistent with the consensus estimate. CONCLUSIONS: The four epidemiologic parameters are interdependent and can be used to test the plausibility of any one parameter. Based on the range of breast cancer penetrance estimates for BRCA1 and BRCA2 founder mutations derived by our approach, recently reported penetrance estimates appear to be overestimated. PMID- 15654227 TI - Outcomes from intensive training in genetic cancer risk counseling for clinicians. AB - PURPOSE: Genetic cancer risk assessment is an emerging interdisciplinary practice that requires knowledge of genetics and oncology and specialized patient and family counseling skills. There is a growing need for cancer risk assessment practitioners, but most clinicians have inadequate cross-disciplinary training. An interdisciplinary course was developed to promote practitioner-level competency in cancer risk assessment to community-based clinicians. METHODS Participants were competitively selected from a pool of board-certified/eligible genetic counseling, masters-trained advanced practice nursing and physician applicants. Preference was given to clinicians with strong institutional backing practicing in underserved regions. The Continuing Medical Education/Continuing Education Unit-accredited course included didactic lectures, workshops, counseling practicum, and case conferences. Pre- and postcourse knowledge tests measured cancer genetics knowledge. Six month and one-year postcourse practice outcome surveys measured the impact of the program on professional self-efficacy and continued professional development. RESULTS/CONCLUSIONS: Forty clinicians completed the course (23 genetic counselors, 14 nurses, and three physicians). There was a significant overall increase of 22.6% in postcourse knowledge scores (P < 0.001). Thirty-five (88%) completed prescribed practice development activities. Of 29 respondents to 1-year postcourse survey, 76% reported increased professional self-efficacy; 66% reported increase in number of patients seen, and virtually all indicated interest in additional training. Outcomes demonstrate the value and efficacy of interdisciplinary training in genetic cancer risk assessment targeted to motivated community-based clinicians. Courses such as this can help address the need for competent cancer risk assessment services in communities outside the academic health center. PMID- 15654228 TI - Genetic susceptibility testing versus family history-based risk assessment: Impact on perceived risk of Alzheimer disease. AB - PURPOSE: We examined how an Alzheimer disease (AD) family history assessment as compared to a risk assessment incorporating the absence of a disease-associated susceptibility allele affected risk perception among adult children with a family history of AD. METHODS: The REVEAL study is a clinical trial in which adult children of patients with AD were randomized to receive a risk assessment based upon family history alone or family history plus apolipoprotein E (APOE) disclosure. In this analysis, two subsets of women were identified, each of whom received identical 29% lifetime risk estimates of developing AD. One group received a risk estimate that incorporated APOE epsilon4-negative genetic test results (Genotype Group, n = 30), whereas the other received a risk estimate based on family history and gender (Family History Group, n = 36). Six weeks after risk disclosure, we surveyed participants regarding the impact of the risk assessment on their perceptions of AD risk. RESULTS: 73% of the Genotype Group judged their risk to be lower compared to 25% of the Family History Group (P < 0.0001). 67% of the Genotype Group reported lower anxiety about AD, versus 26% of the Family History Group (P < 0.01). 80% of the Genotype Group indicated that the risk information had a positive impact, versus 36% of the Family History Group (P < 0.001). The Genotype Group was less likely to believe that they would develop AD (13% vs. 36%, P < 0.05) and was more likely to report that the risk assessment removed uncertainty about their chances of developing AD (63% vs. 9%, P < 0.0001). CONCLUSIONS: These data suggest that risk estimates incorporating negative genetic test results affect perceptions of disease susceptibility more strongly than identical estimates based on family history alone. PMID- 15654229 TI - Fluorescence in situ hybridization and chromosome studies after transfusion in newborns: is a waiting period necessary? AB - BACKGROUND: Delaying chromosome studies after transfusion is common practice in many neonatal intensive care units (NICUs). Yet, no evidence exists to support this practice. PURPOSE: To investigate the effects of filtration and irradiation on chromosome detection, and to evaluate donor chromosome interference after transfusion. METHODS: Packed red blood cells (PRBCs) were evaluated by fluorescence in situ hybridization (FISH) and chromosome analyses. To evaluate donor leukocyte survival, blood was collected from female neonates who received male-donated PRBCs. RESULTS: Irradiated, leukodepleted blood had no Y chromosome detection by FISH. Irradiated, microaggregate filtered blood had Y chromosome detection in all samples by FISH but no metaphase growth. No donor chromosomes were detected in neonates after transfusion. CONCLUSIONS: Delaying chromosome or FISH analysis in transfused neonates who have received irradiated blood is unnecessary. PMID- 15654230 TI - PON1 polymorphism, diabetes mellitus, obesity, and risk of myocardial infarction: Modifying effect of diabetes mellitus and obesity on the association between PON1 polymorphism and myocardial infarction. AB - PURPOSE: Previous studies on PON1 gene polymorphism and the risk of coronary atherosclerotic diseases have been inconsistent. This may be in part due to population difference in prevalence of high oxidative stress and its modifying effect on the association. Diabetes and obesity are two major risk factors of myocardial infarction (MI) and associated with high oxidative stress. We investigated the association between PON1 Q192R polymorphism and the risk of myocardial infarction (MI) and in particular, whether the association can be modified by diabetes mellitus (DM) and obesity. METHODS: We conducted a case control study, including 154 MI cases and 154 controls enrolled in Beijing, China. Epidemiological and clinical data and PON1 Q192R genotype were obtained from each subject. RESULTS: Without considering the modifying effect of DM and obesity, PON1 Q192R polymorphism was not associated with MI. When simultaneously examining the joint association of this polymorphism, DM, and obesity with MI, as compared to subjects with QQ genotypes and without DM and obesity, subjects with QR/RR genotypes and with either DM or obesity had significantly higher risk of MI (OR = 3.6, 95% CI: 1.5-8.7). CONCLUSION: Our data suggest that PON1 Q192R polymorphism was not independently associated with MI but further increased the risk of MI among the subjects with DM, obesity, or both, the conditions associated with high oxidative stress. PMID- 15654232 TI - A 6-year survey of HFE gene test for hemochromatosis diagnosis. AB - PURPOSE: A 6-year survey of HFE gene test was conducted to evaluate its helpfulness for hereditary hemochromatosis diagnosis. METHODS: We analyzed C282Y, H63D, and S65C mutations on 3525 individuals. RESULTS: The test produced 89.7% and 30% of positive results for individuals clinically diagnosed hemochromatosis before HFE gene-test availability and those prospectively tested because of elevated serum iron parameter and/or family history, respectively; among them there were 90.4% and 48.7% of C282Y homozygotes. CONCLUSIONS: The HFE gene test confirmed a genetic defect that may lead to iron loading in individuals when iron parameter values, especially for the C282Y/C282Y, were still low as well as for genotypes usually associated with low expressivity and penetrance (C282Y/H63D, H63D/H63D). This gene-test should allow a biochemical follow-up of patients carrying a disease-related genotype. PMID- 15654231 TI - Recurrence risks for neural tube defects in siblings of patients with lipomyelomeningocele. AB - PURPOSE: Neural tube defects (NTDs) are a group of widely varying congenital malformations resulting from incomplete or improper fusion of the neural tube during embryonic development. NTDs are traditionally classified by the presence or absence of a layer of skin covering the spinal defect. Although a genetic component has been well established in the etiology of open NTDs, studies examining the genetics of closed NTDs such as lipomyelomeningocele are rare. We and others have previously observed families in which multiple members were affected with a broad spectrum of NTDs, suggesting the possibility of a common genetic etiology. METHODS: We calculated the sibling recurrence risk in 52 pedigrees in which the proband was diagnosed with lipomyelomeningocele (LMM), defining recurrence broadly to include both closed and open neural tube defects. RESULTS: Although no recurrences of LMM were observed among younger siblings, one younger sibling had myelomeningocele, yielding an estimate of recurrence risk of 0.04 (95% CI 0.01-0.20). When all siblings of the proband were included, two additional affected siblings were identified, one with anencephaly and another with fatty filum, yielding an estimate of recurrence risk of 0.043 (95% CI 0.01 0.12). CONCLUSIONS: Although the sample size is small, these data are not inconsistent with recurrence risks for myelomeningocele, ranging from 2% to 5% in siblings. These data suggest the underlying genetic basis for closed defects may be the same or closely related to that for myelomeningocele in some families, although a larger sample will be necessary before these data are appropriate for use in a clinical setting. Further characterizations, including whether risk for recurrence of NTDs or LMM in families in which the proband is affected with LMM are altered by folate supplementation, may shed light on the underlying genetics. PMID- 15654233 TI - Fryns syndrome: report of eight new cases. PMID- 15654237 TI - Determination of potential uterine (conceptus) doses from axial and helical CT scans. AB - A tissue equivalent Rando Phantom and thermoluminescent dosimeters (TLDs) (LiF) were used to determine potential doses to the uterus at approximately the position of a conceptus from axial and helical CT scans of that region of the body. Doses were measured at the surface of the phantom and at various locations throughout the phantom. The results indicate that measurements taken on the surface of a patient or phantom are generally higher than the level at the location of the uterus within the patient. The results also indicate that the dose to tissue that is 12.7 cm (5 in) out of the direct beam is less than 10% of the dose within the beam. For tissues that are 20 cm (8 in) out of the beam, the dose is approximately 1% of the dose in the beam. PMID- 15654238 TI - A bioassay program for biomedical research programs. AB - An effective radiation safety program for biomedical research minimizes both external and internal exposures. Limited time and resources can make it difficult to find the right balance between the external dosimetry and internal bioassay components: research typically involves chronic use of small activities rather than acute use of large quantities; concern for external exposures, especially extremities, distracts from the potential for intake; regulatory guidance for only tritium and iodine leave the impression there is little need to monitor for other radionuclides; conversely some licensees over rely on frequent bioassays. Too often researchers discount the potential for intake, especially those who repeatedly use small quantities. This program emphasizes minimizing external and internal exposures while providing the radiation protection staff with criteria for determining the need to assay for any radionuclide under a wide variety of working conditions. PMID- 15654239 TI - A study of the personal radiation dose received by nuclear medicine technologists working in a dedicated PET center. AB - The use of dedicated PET scanners is becoming more widespread throughout Australia and the world. PET imaging utilizes short-lived (approximately 108 min), high-energy (511 keV) gamma-ray emitters that could result in a high radiation dose being received by staff. As part of a larger staff and area monitoring project, this paper discusses the personal dose equivalent, H(p)(10), received by PET staff working in a dedicated PET center. The typical H(p)(10) received by staff was approximately 1 microSv per minute of close contact with patients, which resulted in an average daily dose for nuclear medicine technologists of approximately 31 microSv. The average daily administered activity to patients at Austin Health was 1,280 MBq. PMID- 15654240 TI - Lessons learned in dealing with self luminous exit signs. AB - Self-luminous signs are generally licensed consumer products used to identifying exits, handicap accessibility areas, parking lots, no smoking areas, life boat locations, and emergency stair locations. While purchasers of these signs are exempt from the requirement of applying for a specific license, they are not exempt from all other requirements concerning radioactive material. And they might not be exempt from the liability that can result from lost or damaged signs and improper disposal. A review of the potential for contamination from these signs is presented along with suggestions for handling a contaminating event involving a damaged sign. PMID- 15654241 TI - A simple approach to industrial laser safety. AB - Industrial applications of lasers include marking, welding, cutting, and other material processing. Lasers used in these ways have significant power output but are generally designed to limit operator exposure to direct or scattered laser radiation to harmless levels in order to meet the Federal Laser Product Performance Standard (21CFR1040) for Class 1 laser products. Interesting challenges occur when companies integrate high power lasers into manufacturing or process control equipment. A significant part of the integration process is developing engineering and administrative controls to produce an acceptable level of laser safety while balancing production, maintenance, and service requirements. 3M Company uses a large number of high power lasers in numerous manufacturing processes. Whether the laser is purchased as a Class 1 laser product or whether it is purchased as a Class 4 laser and then integrated into a manufacturing application, 3M Company has developed an industrial laser safety program that maintains a high degree of laser safety while facilitating the rapid and economical integration of laser technology into the manufacturing workplace. This laser safety program is based on the requirements and recommendations contained in the American National Standard for Safe Use of Lasers, ANSI Z136.1. The fundamental components of the 3M program include hazard evaluation, engineering, administrative, and procedural controls, protective equipment, signs and labels, training, and re-evaluation upon change. This program is implemented in manufacturing facilities and has resulted in an excellent history of laser safety and an effective and efficient use of laser safety resources. PMID- 15654243 TI - Dose mapping using MCNP5 mesh tallies. AB - Rensselaer Polytechnic Institute has a 69.6 GBq (1.88 Ci) (137)Cs source that is used for research, calibration of various instruments, and teaching. Recently it was calibrated using ion chambers. The source and room were also modeled in Monte Carlo N-Particle transport code (MCNP5) to determine if the use of a new feature called mesh tallies produces a dose map in the entire room that agrees with the measured results. The dose rate in the hallway, while the source is exposed, was also calculated. It was found that the dose rates calculated from the MCNP5 are in reasonable agreement with the measured results and theoretical predictions. It was also confirmed that the dose rates where the user often stays during the measurement are well below the annual limits. This project shows that the MCNP5 mesh tallies are useful tool for dose mapping in many operational radiation protection situations. PMID- 15654244 TI - Evaluation of lead equivalence of patient and hardware materials in medical diagnostic x-ray shielding. AB - In the estimation of additional shielding requirements for primary beam apart from that provided by patient and hardware in the x-ray beam, there is the need to distinguish between attenuation and hardening properties of materials in comparison. In this work, numerical comparison of attenuation and hardening properties of phantom (Lucite, soft tissue, water) and hardware (aluminum and steel) materials with those of lead have been carried out. Results presented show that the shielding affordable by lead attenuation equivalent thicknesses (LAE) and lead hardening equivalent thicknesses (LHE) is not strictly equivalent to that affordable by thicknesses of substitutes (phantom materials, aluminum and steel) when there are differences in attenuation and hardening properties. Even though beams through LAE that are not "exact" have equal exposure values, the half value layers are higher than those through thicknesses of lead substitutes. Example calculations show that the use of lead thickness (LAE) that are not "exact" to account for the shielding afforded by the thickness of the patient (water phantom) produces lesser reduction of the primary radiation level in the area indicated for shielding. The "exact" LAE that will reduce the primary radiation level equally as the patient and radiographic table may be higher by close to 20% or more of that which is not "exact." PMID- 15654245 TI - Comparative study of the influence of anode and filter materials on primary shielding requirements for mammography. AB - Comparison of primary shielding requirements for mammography x-ray beams that are transmitted through different filter materials at equivalent thicknesses has been carried out. In this study, the primary shielding requirement is that which is additional to that provided by the compression paddle, breast tissue, grid and image receptor. Equivalent thickness is defined as the thickness of a filter material (with Z-value between 37 and 55) that will reduce either intensity or total energy or exposure of a given unfiltered incident beam by the same extent or produce equal hardening of the same unfiltered beam as a reference filter (0.03 g cm Mo). The unfiltered incident beams used are those generated by molybdenum (Mo) and tungsten (W) anodes at kVp's of 18, 20, 22,..,42. In most cases (especially for filter with Z-values greater than 42 and values of operating potential greater than 22 kVp), results show that no single thickness was found that could simultaneously match intensity, total energy, exposure, and hardening as the reference filter. There are significant disparities in the values of the additional primary shielding requirements for beams transmitted through the equivalent thicknesses. Structures of the computer software programs (MAFHAMS and MAXRAPS) used to execute the models developed for the computations of the additional primary shielding requirements for the 'equalized' beams are presented. PMID- 15654246 TI - Ensuring the safety of identity. PMID- 15654249 TI - Fast whole-body assessment of metastatic disease using a novel magnetic resonance imaging system: initial experiences. AB - OBJECTIVE: The objective of this study was to investigate the clinical use of a novel whole-body magnetic resonance imaging (MRI) system for comprehensive assessment of tumor spread in clinical routine. MATERIAL AND METHODS: Sixty-five patients with different tumors with known metastatic disease and 6 healthy volunteers were included. High-resolution MRI from head to toe was performed using multiple phased-array surface coil elements, 24 independent receiver channels, and an integrated parallel acquisition technique (iPAT). A total room time of less than 60 minutes was required. Whole-body MRI and conventional spiral computed tomography (CT) were independently evaluated and compared in terms of feasibility, location/number of detected metastases, and therapeutic relevance. RESULTS: Whole-body MRI was successfully performed in 68 of 71 subjects. Compared with CT, more metastases were detected by MRI in 11 of 63 patients (17%), particularly in brain, liver, spleen, lymph nodes, bone marrow, muscle, and subcutaneous fat tissue. According to these findings, therapy had to be modified in 6 of 63 patients (10%). CONCLUSIONS: High-resolution whole-body MRI is feasible in clinical routine within 1 single examination and offers great potential for fast assessment of individual tumor spread and total tumor burden. PMID- 15654250 TI - Effect of inspiratory and expiratory breathhold on pulmonary perfusion: assessment by pulmonary perfusion magnetic resonance imaging. AB - RATIONALE AND OBJECTIVES: The effect of breathholding on pulmonary perfusion remains largely unknown. The aim of this study was to assess the effect of inspiratory and expiratory breathhold on pulmonary perfusion using quantitative pulmonary perfusion magnetic resonance imaging (MRI). METHODS AND RESULTS: Nine healthy volunteers (median age, 28 years; range, 20-45 years) were examined with contrast-enhanced time-resolved 3-dimensional pulmonary perfusion MRI (FLASH 3D, TR/TE: 1.9/0.8 ms; flip angle: 40 degrees; GRAPPA) during end-inspiratory and expiratory breathholds. The perfusion parameters pulmonary blood flow (PBF), pulmonary blood volume (PBV), and mean transit time (MTT) were calculated using the indicator dilution theory. As a reference method, end-inspiratory and expiratory phase-contrast (PC) MRI of the pulmonary arterial blood flow (PABF) was performed. RESULTS: There was a statistically significant increase of the PBF (delta = 182 mL/100 mL/min), PBV (delta = 12 mL/100 mL), and PABF (delta = 0.5 L/min) between inspiratory and expiratory breathhold measurements (P < 0.0001). Also, the MTT was significantly shorter (delta = -0.5 sec) at expiratory breathhold (P = 0.03). Inspiratory PBF and PBV showed a moderate correlation (r = 0.72 and 0.61, P < or = 0.008) with inspiratory PABF. CONCLUSION: Pulmonary perfusion during breathhold depends on the inspiratory level. Higher perfusion is observed at expiratory breathhold. PMID- 15654251 TI - A novel technique to measure splanchnic transit time using microbubble ultrasound. AB - OBJECTIVE: The objective of this study was to measure splanchnic transit time by intravenous injection of a microbubble. MATERIALS AND METHODS: Ten volunteers were examined before and after eating. After Doppler indices of splanchnic circulation were obtained, the superior mesenteric artery (SMA) and vein (SMV) were simultaneously interrogated using power Doppler ultrasound after intravenous injection of a microbubble. Contrast arrival in the SMA and subsequently the SMV was recorded and splanchnic transit time calculated from differences in the time intensity curves. RESULTS: Splanchnic transit time decreased significantly after eating (mean 11 vs. 6.9 seconds; P = 0.007), reflecting splanchnic hemodynamics. Between-subject variability attributable to repeated measurements was least for the SMA resistive index (17%) but 56% for the new index, suggesting poor reproducibility. CONCLUSION: Splanchnic transit time may be measured by microbubble injection but is subject to considerable measurement error. Newer microbubbles and imaging methods may allow more reproducible measurements. PMID- 15654252 TI - Feasibility of pulmonary ventilation visualization with aerosolized magnetic resonance contrast media. AB - OBJECTIVE: The objective of this study was to evaluate a new approach to noninvasive magnetic resonance assessment of human pulmonary ventilation with aerosolized Gd-DTPA. MATERIALS AND METHODS: Fifteen experimental procedures were carried out in 15 healthy volunteers on a 1.5-T imager. For a timespan of 10 minutes, the subjects spontaneously inhaled a commercially available Gd-DTPA magnetic resonance contrast agent in aerosolized form through an inflatable facemask. Gd-DTPA was aerosolized by means of a small-particle generator with integrated heater to increase aerosol production. Respiratory gated dynamic T1 weighted turbo spin echo images were obtained before and after contrast agent aerosol administration. After nebulization, homogeneity of aerosol distribution was graded by 2 experienced readers and pulmonary signal intensity (SI) changes were measured in corresponding regions of both lungs. RESULTS: Pulmonary ventilation visualization, and hence contrast agent delivery, was rated homogeneously distributed by both readers in 14 of 15 cases (93.3%) and slightly inhomogeneous in 1 case (6.7%). Pulmonary SI increased by an average of +37% +/- 8.5 (range, 10-48%). Allergic responses were not noted. CONCLUSIONS: Human ventilation imaging with aerosolized gadolinium-chelates is viable. The presented modality might evolve as an alternative to current nuclear medicine and magnetic resonance image ventilation imaging procedures, avoiding radiation exposure while offering functional ventilation assessment with an acceptable temporal and spatial resolution. Nevertheless, further evaluation is required to define the potential of gadolinium-based ventilation magnetic resonance imaging in illustrating lung disease. PMID- 15654253 TI - Magnetic resonance colonography: comparison of contrast-enhanced three dimensional vibe with two-dimensional FISP sequences: preliminary experience. AB - PURPOSE: The purpose of this study was to compare a dark-lumen magnetic resonance colonography (MRC) approach with a True FISP-based bright-lumen technique concerning presence of artifacts and the detection rate of colorectal pathologies. MATERIALS AND METHODS: Thirty-seven patients with suspected colorectal lesions were included in this trial. The colon was filled with 2500 mL of tap water. Two-dimensional True FISP datasets as well as T1-weighted GRE sequences (pre- and post intravenous contrast) were acquired. The detection rate of colorectal masses and inflammatory lesions was determined for both techniques separately. Besides, image quality was assessed. All patients underwent conventional colonoscopy as the standard of reference. RESULTS: By means of dark lumen MRC datasets, all polyps >5 mm were correctly diagnosed, whereas 4 polyps <5 mm were missed. Sensitivity of dark-lumen MRC amounted to 78.9%. There were no false-positive results: residual stool could correctly be differentiated from colorectal masses. The True FISP-based bright-lumen MRC, however, failed to detect 2 additional polyps resulting in a sensitivity of 68.4%. Furthermore, bright-lumen MRC led to false-positive results in 5 patients. Both techniques visualized inflammatory bowel disease in 5 patients. However, image quality of True FISP was rated superior to that of dark-lumen MRC. CONCLUSION: Dark-lumen MRC proved to be superior over bright-lumen MRC regarding the detection of colorectal masses. However, True FISP imaging can turn out to be helpful as a result of high image quality and motion insensitivity. PMID- 15654254 TI - Pericardial and visceral adipose tissues measured volumetrically with computed tomography are highly associated in type 2 diabetic families. AB - RATIONALE AND OBJECTIVES: Pericardial and visceral adipose tissue volumes can provide new insight into the complex relation between obesity, adult-onset diabetes, and cardiovascular disease. We describe a new method for quantifying pericardial adipose tissue volumes with computed tomography (CT), and present its precision and relation to established measures of adiposity. METHODS: Eighty subjects randomly selected from a family study of sibling pairs concordant for type 2 diabetes and unaffected siblings, 69 with diabetes, had 2 cardiac CT scans with electrocardiographic gating and 1 abdominal scan as part of an examination designed to measure calcified atherosclerotic plaque. Pericardial adipose tissue and visceral adipose tissue were measured using a 3-dimensional analysis technique. Body mass index, waist circumference, waist-to-hip ratio, and percent fat by dual x-ray absorptiometry were measured during the same visit. RESULTS: Pericardial adipose tissue volumes measured independently and in a random order from the 2 sequential cardiac CT scans obtained during the same examination were highly correlated (Spearman R = 0.99; P < 0.0001). The mean +/- standard deviation (median) pericardial adipose tissue volume was 320.5 +/- 147.3 (281.7) mL. Pericardial adipose tissue was highly correlated with total abdominal visceral adipose tissue (R = 0.81; P < 0.0001). CONCLUSION: The significant association between pericardial and visceral adipose tissue volumes in this preliminary study suggests that pericardial, like visceral adipose tissue, may be an important predictor or risk factor for cardiovascular disease and other related illnesses and warrants further evaluation. PMID- 15654255 TI - Functional characterization of prostate cancer by integrated magnetic resonance imaging and oxygenation changes during carbogen breathing. AB - OBJECTIVE: The objective of this study was to evaluate the changes in oxygenation of prostate cancer induced by carbogen breathing using blood oxygen level dependent (BOLD) magnetic resonance image (MRI) with an endorectal coil (eMRI). MATERIALS AND METHODS: In 32 patients with biopsy-proven prostate cancer, endorectal MRI was performed at 1.5 Tesla using the BOLD method. Images were acquired during 4 x 4-minute episodes alternating between room air and carbogen (95% O2/5% CO2) breathing. In each episode, 40 images were acquired (T2*-weighted EPI sequence, 12-14 slices, 3-mm thickness). All patients underwent radical prostatectomy; BOLD-MRI findings were correlated with the histopathologic results. RESULTS: BOLD-MRI could be evaluated in 29 patients, and revealed heterogeneous signal changes of normal prostate and cancer tissue similar to the heterogeneity of prostate tissue in anatomic/pathologic preparation. A significant signal intensity increase (P = 0.004) was found in normal central gland and peripheral zone during carbogen breathing. Signal enhancement in carcinoma was significantly lower (P = 0.004) compared with the contralateral normal side. CONCLUSION: Intrinsic blood-tissue contrast-functional MRI during carbogen breathing may help detect and characterize prostate carcinoma from normal tissue, particularly in small 1-sided carcinomas. This may be useful for identifying candidates for radiotherapy and monitoring noninvasive therapeutic approaches. PMID- 15654256 TI - In vivo visualization of white matter fiber tracts of preterm- and term-infant brains with diffusion tensor magnetic resonance imaging. AB - OBJECTIVE: The goal of this study was to test the feasibility of visualizing a 3 dimensional structure of cerebral white matter fiber tracts in preterm infants, postconceptional age (PCA) 28 weeks to term, by using volumetric diffusion tensor magnetic resonance imaging (DTI) data. MATERIALS AND METHOD: We combined tractography algorithms and visualization methods, currently available for adult DTI data, to trace the pixelated principal direction of a diffusion tensor originating from regions-of-interest with high fractional anisotropy. Consequently, white matter fiber bundles from the genu and the splenium of corpus callosum, the corticospinal tracts, the inferior fronto-occipital fasciculi, and optic radiations were visualized. RESULTS: Our results suggest that major white matter tracts of preterm infant brains, with PCAs ranging from 28 weeks to term (40 weeks old), can be successfully visualized despite the small brain volume and low anisotropy. CONCLUSION: The feasibility of fiber tractography in preterm neonates with DTI may add a new dimension in detection and characterization of white matter injuries of preterm infants. PMID- 15654258 TI - Human alphavbeta3 integrin potency and specificity of TA138 and its DOTA conjugated form (89)Y-TA138. AB - The present study was undertaken to define the alphavbeta3-binding potency and specificity of TA138, a nonpeptide integrin antagonist, and its conjugated form, 89Y-TA138. Various integrin-specific binding and functional assays as well as cell-adhesion assays were used to determine the potency and integrin specificity for TA138 and 89Y-TA138. Both TA138 and 89Y-TA138 inhibited alphavbeta3-mediated [125I]echistatin binding to 293-beta3-transfected cells, with IC50 values of 0.046 and 0.059 microM, respectively, and IC50 values of 0.012 and 0.018 microM, respectively, in inhibiting an alphavbeta3 integrin-mediated 293-beta3 transfected cell adhesion to fibrinogen. TA138 inhibited human umbilical vein endothelial cell adhesion to fibrinogen, with an IC50 value of 0.052 +/- 0.006 microM. Both TA138 and 89Y-TA138 demonstrated a relatively high degree of specificity for human alphavbeta3 integrin as compared with other human integrins, including alphavbeta5, alphaIIbbeta3, and alpha5beta1 (IC50 > 10 microM). Both 89Y-TA138 and TA138 demonstrated comparable alphavbeta3 affinity and specificity as compared with other closely related human integrins such as alphavbeta5, alphaIIbbeta3, or alpha5beta1. PMID- 15654257 TI - Endothelin-1 acts via protein kinase C to block KATP channels in rabbit coronary and pulmonary arterial smooth muscle cells. AB - We investigated the effects of the vasoconstrictor endothelin-1 (ET-1) on the whole-cell ATP-sensitive K+ (KATP) currents of smooth muscle cells that were isolated enzymatically from rabbit coronary artery (CASMCs) and pulmonary artery (PASMCs). The size of the KATP current did not differ significantly between CASMCs and PASMCs. ET-1 reduced the KATP current in a concentration-dependent manner, and this inhibition was greater in PASMCs than in CASMCs (half-inhibition values of 12.20 nM and 1.98 nM in CASMCs and PASMCs, respectively). However, the level of inhibition induced by other vasoconstrictors (angiotensin II, norepinephrine, and serotonin) were not significantly different between CASMCs and PASMCs. Pretreatment with the protein kinase C (PKC) inhibitors staurosporine (100 nM) and GF 109203X (1 microM) prevented ET-1-induced inhibition of the KATP current in both arterial smooth muscle cell preparations. The PKC activators phorbol-12,13-dibutyrate (PDBu) and 1-olelyl-2-acetyl-sn-glycerol (OAG) reduced the KATP current in dose-dependent manner. Although the numbers of ET receptors were not significantly different between the 2 arterial smooth muscle cell preparations, the effects of PDBu and OAG were greater on PASMCs. ET-1-induced inhibition of the KATP current was unaffected by the PKA inhibitor Rp-cAMPs (100 microM) and PKA inhibitory peptide (5 microM). PMID- 15654259 TI - Unfractioned heparin produces vasodilatory action on human internal mammary artery by endothelium-dependent mechanisms. AB - The aim of this study was to investigate whether unfractioned heparin produces a direct vasodilatory effect on the human internal mammary artery (IMA) and the possible underlying mechanisms. Samples of redundant IMA were obtained from 20 patients undergoing coronary artery bypass graft surgery, and concentration response curves to unfractioned heparin were constructed. Unfractioned heparin (0.5-6 U/mL) caused a concentration-dependent relaxation in the endothelium intact human IMA rings precontracted with phenylephrine (10(-6) M). Removal of endothelium significantly inhibited the responses of human IMA to unfractioned heparin (P < 0.05). Nomega-Nitro-L-arginine methyl ester (L-NAME, 10(-4) M), 1H [1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, 10(-5) M) and L-NAME (10(-4) M) plus ODQ (10(-5) M) partially reduced unfractioned heparin-induced vasodilatory response in endothelium-intact rings, whereas indomethacin alone had no effect. The vasodilatory effect of unfractioned heparin was completely inhibited by 40 mM KCl in the presence of L-NAME, ODQ, and indomethacin. These results clearly demonstrated that unfractioned heparin causes a concentration-dependent vasodilatation in human internal mammary artery, and this action seems to be via endothelium-dependent mechanisms, including generation of nitric oxide and endothelium-derived hyperpolarizing factor. PMID- 15654260 TI - Prostacyclin does not inhibit rho-kinase: an implication for the treatment of pulmonary hypertension. AB - Primary pulmonary hypertension continues to be a fatal disease. We have recently demonstrated that long-term inhibition of Rho-kinase, an effector of the small GTPase Rho, is effective for the treatment of pulmonary hypertension (PH) in rats and humans. Prostacyclin has been clinically used for the treatment of PH with moderate success. However, it remains to be examined whether Rho-kinase inhibition is involved in its beneficial effects on PH. In an ELISA assay, neither prostacyclin nor its oral analogue, beraprost sodium, inhibited Rho kinase even at higher concentrations (10(-7) to 10(-5) M, 100 to 10,000 times higher than their clinical concentrations), whereas specific Rho-kinase inhibitors, fasudil and hydroxyfasudil, markedly (approximately 95%) inhibited the Rho-kinase activity at 10(-5) M (near their clinical concentrations). Beraprost sodium did not significantly suppress serotonin-induced vascular smooth muscle cell (VSMC) contractions or Rho-kinase activity of the rat aorta without endothelium, as evaluated by the extent of phosphorylation of the ERM family, a substrate of Rho-kinase, whereas hydroxyfasudil markedly suppressed the VSMC contractions and Rho-kinase activity. These results indicate that prostacyclin lacks direct inhibitory effect on Rho-kinase and suggest that combination therapy with prostacyclin and a Rho-kinase inhibitor could exert further beneficial effects on PH. PMID- 15654261 TI - In vivo and in vitro antiarrhythmic effects of SSR149744C in animal models of atrial fibrillation and ventricular arrhythmias. AB - SSR149744C (2-butyl-3-{4-[3-(dibutylamino)propyl]benzoyl}-1-benzofuran-5 carboxylate isopropyl fumarate) is a new noniodinated benzofuran derivative structurally related to amiodarone and dronedarone that is currently undergoing clinical trials as an antiarrhythmic agent. As SSR149744C exhibits electrophysiological and hemodynamic properties of class I, II, III, and IV antiarrhythmic agents, the aim of this study was to evaluate its acute intravenous (IV) or oral (PO) antiarrhythmic activities in in vitro and in vivo animal models of atrial and ventricular arrhythmias. In vagally induced atrial fibrillation (AF) in anesthetized dogs, SSR149744C (3 and 10 mg/kg IV) terminated AF in all 7 dogs and prevented reinduction in 4 out of 7 dogs; effective refractory periods of right atrium were dose-dependently and frequency independently lengthened. In low-K+ medium-induced AF models, SSR149744C (0.1 to 1 microM) prevented AF in isolated guinea pig hearts in a concentration-dependent manner. At the ventricular level, SSR149744C (0.1 to 10 mg/kg IV and 3 to 90 mg/kg PO) prevented reperfusion-induced arrhythmias in anesthetized rats with a dose-effect relationship, and, at doses of 30 to 90 mg/kg PO, it reduced early (0 24 hours) mortality following permanent left coronary artery ligature in conscious rats. The present results show that SSR149744C is an effective antiarrhythmic agent in atrial fibrillation and in ventricular arrhythmias. Like amiodarone and dronedarone, its efficiency in these animal models of arrhythmias is likely be related to its multifactorial mechanism of action. PMID- 15654262 TI - Mechanisms underlying the endothelium-independent relaxation induced by angiotensin II in rat aorta. AB - It has been suggested that low concentrations of angiotensin II cause vasoconstriction, whereas high concentrations evoke vasodilation. Thus, this work aimed to characterize functionally the mechanisms underlying angiotensin II induced relaxation, at high concentration, in isolated rat aortic rings. Vascular reactivity experiments, using standard muscle bath procedures, showed that angiotensin II (1-30 microM) concentration-dependently induces relaxation of phenylephrine-precontracted rings with intact or denuded endothelium. The relaxation was not altered in the presence of ethylenediamine tetraacetic acid (EDTA), a nonselective inhibitor of metalloprotease. The selective antagonist of AT2 receptors, PD123319, inhibited angiotensin II-induced relaxation. Conversely, losartan or A-779, selective AT1 and Ang1-7 receptor antagonists, respectively, did not alter the relaxation induced by angiotensin II. HOE-140, a selective antagonist of the bradykinin B2 receptor, and amiloride, a Na+/H+ exchanger inhibitor, abolished angiotensin II-induced relaxation. Administration of exogenous bradykinin on precontracted tissues produced concentration-dependent relaxation, which was also inhibited by HOE-140. Preincubation of denuded-rings with NG-nitro-L-arginine methyl ester (L-NAME), 1H-[1,2,4]oxadiazolo[4,3 a]quinoxalin-1-one (ODQ), indomethacin, or tetraethylammonium (TEA) reduced angiotensin II-induced relaxation. The combination of L-NAME, indomethacin, and TEA completely abolished the relaxation induced by angiotensin II. 4 Aminopyridine (4-AP) as well as charybdotoxin reduced angiotensin II-induced relaxation. On the other hand, neither apamin nor glibenclamide altered the relaxation induced by angiotensin II. The major new finding of this work is that it demonstrated functionally the existence of AT2 receptors located on smooth muscle of rat aortic rings that mediated vasorelaxation via stimulation of B2 receptors by bradykinin, which in turns results in the activation of the NO-cGMP pathway, vasodilator cyclooxygenase product(s), and voltage-dependent and Ca+ activated large-conductance K+ channels. PMID- 15654264 TI - Tumor necrosis factor alpha, rapid ventricular tachyarrhythmias, and infarct size in canine models of myocardial infarction. AB - Etanercept (2 mg/kg), a TNFalpha sequestrant, was administered 24 hours and 1 hour before LAD coronary artery ligation to examine the role of TNFalpha on lethal ventricular tachyarrhythmias and myocardial necrosis. Dogs treated with etanercept had decreased very rapid (>360 bpm) ventricular triplets (6 +/- 1/h, n = 8) 2 to 24 hours following coronary artery ligation compared with saline (21 +/ 6/h, n = 10, P < 0.05). This was concordant with 8-fold salvage of beta adrenergic receptor kinase 1 (betaARK) activity compared with control (33.8 +/- 7.2% versus 4.3 +/- 2.2% of unoperated control tissue, P < 0.01, n = 5). Salvage of betaARK occurred without change in the thickness of the epicardial tissue overlying the infarct. In dogs pretreated with etanercept before a 2-hour occlusion/4-hour reperfusion of the LAD coronary artery, infarct mass decreased by 61% (% area at risk) and 55% (% left ventricular mass) in the etanercept group (n = 8) compared with saline (n = 9, P < 0.05). This was concordant with an etanercept-mediated six-fold decrease in leukocyte accumulation within ischemically injured myocardium. TNFalpha antagonism decreases malignant ventricular tachyarrhythmias and may relate to partial protection of normal betaARK-mediated desensitization of beta-adrenergic receptors. TNFalpha sequestration also decreases infarct size in an occlusion/reperfusion model of myocardial ischemia. PMID- 15654263 TI - Effects of dobutamine on left ventriculoarterial coupling and mechanical efficiency in acutely ischemic pigs. AB - This study investigated the effects of dobutamine on left ventriculoarterial (VA) coupling and mechanical efficiency in acutely ischemic pigs. Experiments were performed in 12 pigs in which vascular properties, including peripheral resistance (R2), compliance (C), and arterial elastance (Ea), were estimated with a windkessel model, and left ventricular (LV) function by the slope (Ees) of the end-systolic pressure-volume relationship (ESPVR) and stroke work (SW). VA coupling was defined as Ees/Ea, and mechanical efficiency as SW/pressure-volume area (PVA). In all animals, the left anterior descending coronary artery was ligated after basal measures. The animals were then randomly divided into 2 groups: group CTRL (n = 6) was followed for 180 minutes without other intervention, whereas group DOBU (n = 6) was infused with dobutamine (5 microg x kg(-1) x min(-1)) starting after T60 measures. Coronary occlusion induced a rightward shift of ESPVR and a decrease in Ees from 3.67 +/- 0.33 to 1.92 +/- 0.20 mm Hg x mL(-1), while Ea changed from 3.33 +/- 0.56 to 4.65 +/- 0.29 mm Hg x mL, R2 from 1.72 +/- 0.30 to 2.38 +/- 0.16 mm Hg x s x mL(-1), and C from 0.78 +/ 0.16 to 0.46 +/- 0.08 mL x mm Hg(-1). This altered VA coupling from 1.22 +/- 0.11 to 0.44 +/- 0.07. SW decreased from 4056 +/- 223 to 2372 +/- 122 mm Hg x mL, and PVA and SW/PVA decreased from 5575 +/- 514 to 4830 +/- 317 mm Hg x mL, and from 0.76 +/- 0.04 to 0.49 +/- 0.03, respectively. In group DOBU, dobutamine restored Ees and the position of ESPVR to baseline values, while Ea decreased to 3.39 +/- 0.34 mm Hg x mL(-1) because of an R2 decrease to 1.60 +/- 0.24 mm Hg x s x mL(-1). VA coupling was restored. SW and PVA increased to 3833 +/- 180 mm Hg x mL and to 7498 +/- 442 mm Hg x mL, respectively, while SW/PVA was unchanged. In ischemic pigs, dobutamine restored VA coupling through an increase in LV contractility and decrease in arterial elastance as a result of peripheral vasodilatation. However, myocardial oxygen consumption was increased, and mechanical efficiency impaired. PMID- 15654265 TI - Hypolipidemic drugs affect monocyte IL-1beta gene expression and release in patients with IIa and IIb dyslipidemia. AB - Because atherosclerosis has been proven to be an inflammatory disease, it became obvious that the proper treatment of dyslipidemic patients should not only correct lipid parameters but also inhibit the inflammatory state. One of the crucial proinflammatory and procoagulant cytokines participating in the pathogenesis of atherosclerosis is interleukin-1beta (IL-1beta). Therefore, the aim of the study was to asses the effect of statin and fibrate therapy (for dyslipidemia IIa and IIb, respectively) on IL-1beta gene expression and monocyte release evaluated in each patient. Additionally, the effect of hypolipidemic therapy on fibrinolysis was evaluated. The study was carried out in 37 patients: 12 with biochemically confirmed type IIa dyslipidemia (treated with atorvastatin), 12 with type IIb dyslipidemia (treated with fenofibrate), and 13 age- and sex-matched normolipidemic persons (control). IL-1beta concentrations in cultured monocytes and PAI-1 (Plasminogen Activator Inhibitor) plasma levels were measured using the ELISA method. To evaluate the expression of IL-1beta gene in monocytes, a semiquantitive RT-PCR procedure was performed. The results were normalized with the expression of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) as a housekeeping gene. Although IL-1beta monocyte release was markedly elevated in patients with atherogenic dyslipidemias, IL-1beta gene expression was only slightly and nonsignificantly higher in the studied groups versus control. We have observed significant reduction of IL-1beta mRNA expression after 30-day treatment with the examined drugs (atorvastatin, 2.10 +/- 0.50 versus 1.05 +/- 0.15; P < 0.001, fenofibrate; 2.27 +/- 0.48 versus 1.23 +/- 0.27; P < 0.01). There was no significant difference between statin and fibrate effect on IL-1beta mRNA expression. Similarly, we have noticed significant reduction of IL-1beta release by cultured monocytes after 30-day statin therapy (133.0 +/- 5.7 pg/mL versus 77.0 +/- 3.6 pg/mL; P < 0.01) and fibrate therapy (143.9 +/- 6.5 pg/mL versus 86.2 +/- 5.9 pg/mL; P < 0.01). Besides this antiinflammatory effect, we have observed a 30% reduction of PAI-1 plasma levels in both treated groups. In conclusion, effective 1-month hypolipidemic therapy with atorvastatin or fenofibrate diminished plasma levels of proinflammatory and procoagulatory state markers. PMID- 15654267 TI - Prolongation of QT interval by terbutaline in healthy subjects. AB - In a double-blind, randomized placebo-controlled crossover study, we characterized how terbutaline prolonged cardiac corrected QT interval (QTc). The study was carried out in six young and healthy male subjects in supine position. Escalating terbutaline doses were administered intravenously at infusion rates of 6 mL/h (10 microg terbutaline/min), 12 mL/h (20 microg terbutaline/min), and 18 mL/h (30 microg terbutaline/min). Terbutaline maximally prolonged QTc intervals on average by 60%, from 358 milliseconds (SD 28) to 456 milliseconds (SD 19). The effect was closely associated with a simultaneous decrease in plasma potassium concentration from 4.0 mmol/L (SD 0.1) to 2.5 mmol/L (SD 0.1). The final phase of slow ventricular repolarization, the interval between the apex and the end of T wave, was proven to be highly sensitive to the hypokalemic terbutaline actions, whereas the earlier repolarization phases were not strongly affected by terbutaline. Estimated by using the classic Nernst equation for membrane potentials, terbutaline-induced hypokalemia hyperpolarized ventricular myocardium from the resting level of -90 mV to -110 mV. The prolongation of QTc interval was related to ventricular hyperpolarization with a Pearson correlation coefficient of 0.91. Terbutaline-induced prolongation of QTc interval in healthy volunteers is in conformity with repolarization studies carried out in isolated canine heart ventricular preparations in which the cardiac ventricular cell membrane potential determines the duration of the final phase of slow ventricular repolarization. PMID- 15654268 TI - Endothelin receptor antagonists: clinical realities and future directions. AB - Endothelin (ET) is among the strongest endogenous vasoconstrictors known and a potent mitogen. A rich body of experimental evidence suggests that ET contributes to vascular remodeling and end-organ damage in several cardiovascular conditions. Therefore, blockade of ET receptors has been suggested as an attractive target in a number of acute and chronic cardiovascular indications, including pulmonary arterial hypertension (PAH), systemic hypertension, and heart failure. To date, clinical studies have confirmed expectations in PAH and yielded promising initial results in systemic hypertension, which are currently awaiting confirmation in large-scale trials. In contrast, no added benefit could be demonstrated in large clinical trials on top of current standard treatment in both acute and chronic heart failure. Further clinical development in heart failure has therefore been suspended. Other indications that are currently being studied clinically or would possibly merit clinical trials include acute myocardial ischemia and reperfusion, cerebral vasospasm after intracranial bleeding, glaucoma, acute severe pancreatitis, systemic sclerosis, (diabetic) renal failure, restenosis after angioplasty/stent implantation, and late transplant rejection. This article critically reviews the available clinical data on ET receptor antagonism in cardiovascular indications against the background of the underlying preclinical research. PMID- 15654266 TI - Paradoxical effect of dofetilide on action potential duration and calcium transient amplitude in newborn rabbit ventricular myocytes. AB - The Na+-Ca2+ exchanger (NCX) is up-regulated in the neonatal rabbit heart. Because the duration of membrane depolarization is an important determinant of calcium entry via NCX, pharmacological agents that lengthen the action potential (AP) may significantly increase the amount of activator calcium in newborns. We tested this potentially novel therapeutic strategy by using action potential voltage clamp steps or using dofetilide, a blocker of IKr, to prolong the action potential duration (APD). The effects of changing APD on calcium transients were determined in ventricular myocytes at different developmental stages: newborn (1 4 days), juvenile (9-10 days), and adult ventricular myocytes (35 degrees C; 1 Hz). Calcium transient amplitude in neonatal myocytes increased substantially with clamping with longer APs. In contrast, exposure to dofetilide (0.1, 1, and 10 microM) under current clamp conditions increased APD in a concentration dependent manner but had no significant effect on calcium transient amplitude in either neonates or adults. When the AP was held constant under voltage clamp conditions, dofetilide decreased the calcium transient amplitude in neonates. This effect is likely related to inhibition of sodium-calcium exchanger and L type Ca2+ currents (ICa), as observed in separate experiments. These results suggest that dofetilide has a paradoxical effect on APD and calcium transients in the newborn heart. PMID- 15654269 TI - In the future: when is a rose is a rose is a rose? PMID- 15654270 TI - Conservative therapy in intraocular retinoblastoma: response/recurrence rate. AB - PURPOSE: To evaluate the response/recurrence rate and the outcome in intraocular retinoblastoma treated with chemoreduction and focal therapy, the authors performed a retrospective review of their patients. METHODS: The series included 46 newly diagnosed patients with unilateral or bilateral intraocular retinoblastoma (58 eyes) receiving carboplatin/etoposide chemotherapy associated with focal therapy (laser or cryotherapy). The mean follow-up was 53 months (range 11-125). RESULTS: Fifty-one eyes (88%) presented with complete response after four to eight courses of chemotherapy combined with focal treatment. The response rate was 100% in group 1, 94% in group 2, 100% in group 3, 83% in group 4, and 70% in group 5 (5 vs. 1-4, P < 0.03; 5-4 vs. 1-3, P < 0.025). Twenty-nine eyes (57%) relapsed after a mean of 7 months (range 2-36). The relapse rate was 30% in group 1, 27.% in group 2, 67% in group 3, 80% in group 4, and 100% in group 5 (5 vs. 1-4, P < 0.001; 4-5 vs. 1-3, P < 0.001). Seven of 18 cases achieved a second complete response with further conservative treatment (total courses 8-14). Twenty-nine eyes (50%) were treated without external-beam radiotherapy or enucleation: 90% in group 1, 69% in group 2, 67% in group 3, 33% in group 4, and 6% in group 5 (5 vs. 1-4, P < 0.01; 5-4 vs. 1-3, P < 0.001). Ten eyes (17%) required external-beam radiotherapy and 21 eyes (36%) enucleation. The ocular salvage rate was 67%. CONCLUSIONS: Although all groups of patients with intraocular retinoblastoma responded to carboplatin/etoposide chemotherapy associated with focal therapy, all the cases in group 5 relapsed. This approach is questionable in group 5, in which could be justified to delay aggressive treatment in a very young child. PMID- 15654271 TI - Suppressive effect of intravenous immunoglobulin on peripheral blood neutrophil count in patients with idiopathic thrombocytopenic purpura. AB - PURPOSE: To determine the effect of intravenous immunoglobulin (IVIG) on neutrophil count, the authors studied patients with idiopathic thrombocytopenic purpura (ITP). PATIENTS AND METHODS: Thirty-one of 40 patients with ITP were treated with an IVIG regimen (400 mg/kg/daily for 5 days), while the other 9 were observed without any medications. Peripheral samples were examined during the clinical course. RESULTS: The absolute neutrophil count in 22 patients with no fever as a side effect decreased significantly by day 2 after the start of IVIG, but in most patients it began to increase soon after the last IVIG. There was also a significant difference between the decreased level of the absolute neutrophil count in patients with and without IVIG. However, re-administration of IVIG within 2 weeks did not significantly suppress the neutrophil count. In contrast, the neutrophil count increased in every patient with a fever as a side effect of IVIG. CONCLUSIONS: IVIG induced a decrease in peripheral blood neutrophil count in patients with ITP. PMID- 15654273 TI - Perceptions of distress among adolescents recently diagnosed with cancer. AB - With the goal of studying perceived distress among adolescents recently diagnosed with cancer, 56 adolescents were interviewed by telephone 4 to 8 weeks after diagnosis. The interviews included a structured interview guide, the Hospital Anxiety and Depression Scale, and the subscales Mental Health and Vitality from SF-36. "Losing hair" and "missing leisure activities" were identified as the most prevalent aspects of distress, whereas "missing leisure activities" and "fatigue" were rated with the highest levels of distress. "Worry about not getting well," "mucositis," "nausea," "pain from procedures and treatments," and "worry about missing school" were rated as the overall worst aspects by most adolescents. Twelve percent reached the cutoff score for potential clinical anxiety and 21% for potential clinical depression. Ratings of Mental Health and Vitality were lower than norm values. Prevalence of pain from procedures/treatments was higher among those who scored in the clinical range of depression, and more adolescents who were treated at a local hospital scored in the clinical range of anxiety. The findings show that ratings of prevalence, levels, and aspects perceived as the worst are not necessarily in accordance, that adolescents scoring in the clinical range of psychological distress are in the minority, and that the adolescents experience reduced physical and mental well-being. PMID- 15654272 TI - Prognostic value of quantifying apoptosis factor expression in favorable histology wilms tumors. AB - Important in the homeostasis of normal tissues, apoptosis can be altered to favor cell survival within tumors. High expression of survivin, an inhibitor of apoptosis, and absence of caspase 8, a pro-apoptotic enzyme, independently correlate with poor outcomes in several tumor types. Favorable histology Wilms tumor has a remarkably high cure rate; as a result, the focus of therapy is now aimed at reducing treatment-related morbidity. With the goal of safely reducing therapy in select subgroups of patients, the authors investigated whether the levels of apoptotic factors in tumors could predict the risk for recurrence. Tumor apoptotic factor levels were surveyed in a case-control study from the National Wilms Tumor Study 5 (NWTS-5) and measured via quantitative real-time RT PCR. Survivin and caspase 8 levels were surveyed in 92 primary tumor specimens and SMAC, Bid, and CD95 were surveyed in 24 specimens. All four pro-apoptotic factors studied (caspase 8, SMAC, Bid, and CD95) were analyzed individually and in relation to survivin expression. Although survivin mRNA was present at markedly greater levels than in normal kidney, none of the factors assayed independently or as a ratio was associated with stage of disease or risk for tumor recurrence in this group of tumors. PMID- 15654274 TI - Hypercalcemia and childhood cancer: a 7-year experience. AB - Hypercalcemia is a rare but potentially fatal complication during the management of childhood cancer. The treatment of severe hypercalcemia in children has not been clearly defined. The authors present a retrospective series of 16 children (11 boys and 5 girls) with severe hypercalcemia (>/=2.9 mmol/L) treated between 1997 and 2004 for malignancy. Median serum calcium level was 3.14 mmol/L. Hypercalcemia was present at the initial diagnosis of cancer (eight patients) or occurred during treatment (five patients) or during relapse (three patients). Three children had several episodes of hypercalcemia. All children were treated by hydration for a median of 7 days (range 2-12 days). Eight patients received intravenous pamidronate. The other treatments were adapted to the mechanism of hypercalcemia. Serum calcium levels were lowered to below 3 mmol/L after a median of 2 days and to below 2.7 mmol/L after a median of 4 days after starting treatment. Pamidronate was well tolerated apart from one case of multifactorial renal failure. Intravenous pamidronate is a safe and effective treatment for severe cancer-related hypercalcemia in children. Specific therapy must be initiated as soon as possible. Serum calcium levels must be monitored for a fortnight after administration of pamidronate due to the risk of hypocalcemia. PMID- 15654275 TI - Second malignancy after treatment of pediatric Hodgkin disease. AB - Although treatment of pediatric Hodgkin disease has become highly effective over the past 40 years, a number of patients have developed concerning late effects, such as secondary malignancies. These cancers may occur years to decades after remission and arise in the breast, thyroid, gastrointestinal tract, lung, skin, urogenital tract, and brain. There is also an increased risk of leukemia and non Hodgkin lymphoma. Etiology and risk factors for each cancer type vary but often include certain chemotherapy agents and radiation dosages. Survivorship also varies but is often poor. The authors examined retrospective analyses of these secondary malignancies and present a summary of these findings. The information may allow clinicians to better monitor childhood Hodgkin disease survivors and reduce mortality. PMID- 15654276 TI - Two cases of Ralstonia pickettii bacteremias in a pediatric oncology unit requiring removal of the Port-A-Caths. AB - Ralstonia pickettii is an aerobic, gram-negative bacterium causing bacteremia following the use of contaminated saline vials, respiratory therapy solutions, skin disinfectants, blood culture mediums, and water supplies. It is rarely associated with human infections. The authors report two cases of R. pickettii bacteremia in patients with Port-A-Caths that could be treated only by removal of the ports. PMID- 15654277 TI - Colorectal carcinoma in children, adolescents, and young adults. AB - The authors conducted a retrospective evaluation of patients younger than 30 years with colorectal carcinoma in Argentina. Patients were divided into group 1 (patients treated at pediatric institutions from 0 to 20 years of age, n = 14) and group 2 (patients from 21 to 30, n = 7, treated at adult centers). Group 1 had significantly more advanced disease and a poorer prognosis. Six patients (2/14 in group 1 and 4/7 in group 2) survive disease-free. Thirteen patients died of progressive disease, 1 died of a non-tumor-related cause, and 1 is still on treatment. Patients younger than 20 years have a poorer prognosis, probably because of advanced disease on presentation. PMID- 15654278 TI - Long-term survival in high-grade axial osteosarcoma with bone and lung metastases treated with chemotherapy only. AB - A boy, age 2 years 10 months, with high-grade malignant osteosarcoma of the fifth lumbar vertebra with secondary bilateral pulmonary lesions and bone metastasis at the fifth thoracic vertebra is described. The primary site of disease was inoperable and the patient was treated with chemotherapy only. At present, 83 months from diagnosis and 64 from the end of therapy, he is in very good general condition. Although a surgical approach on the primary and secondary sites is fundamental, this case may be considered an indication of the efficacy of aggressive chemotherapy in treating osteosarcoma. PMID- 15654279 TI - Successful treatment of an unresectable choroid plexus carcinoma in a patient with Li-Fraumeni syndrome. AB - Choroid plexus carcinoma (CPC) is an uncommon central nervous system tumor requiring complete surgical excision for favorable outcome. The authors report the successful treatment of a 2-year-old patient with widely disseminated CPC and Li-Fraumeni syndrome. Following a partial resection of the tumor the patient received chemotherapy consisting of cyclophosphamide, etoposide, and carboplatin. There were no additional surgical procedures and radiation was not administered. Remarkably, the patient remains without evidence of active disease more than 3 years from the completion of therapy. Additional studies are necessary to determine whether this treatment plan can be beneficial to other patients with CPC and whether the patient's p53 mutation had an effect on outcome. PMID- 15654281 TI - Association of chronic symptomatic neutropenia with the triple A syndrome. AB - Chronic neutropenia syndromes include distinct hereditary disorders with varying degrees of neutropenia. Among the more common inherited disorders associated with symptomatic neutropenia are cyclic neutropenia, severe congenital neutropenia (Kostmann disease), and Schwachman-Diamond syndrome. The authors describe a 17 year-old girl with triple A syndrome who developed a progressive decrease in the granulocyte count, finally resulting in long-standing neutropenia. Its probable pathogenesis may be related to dysfunction of ALADIN (the protein known to be mutated in triple A syndrome), resulting in abnormal nucleocytoplasmic transport of essential proteins, in myeloid precursor cells. Chronic neutropenia should therefore be considered among the clinical manifestations of triple A syndrome. PMID- 15654280 TI - Transient neonatal myeloproliferative disorder without Down syndrome and detection of GATA1 mutation. AB - Transient myeloproliferative disorder is a form of self-limited leukemia that occurs almost exclusively in neonates with Down syndrome. The authors report an unusual case of a newborn without constitutional trisomy 21 who developed undifferentiated leukemia and subsequently achieved clinical and molecular remission without chemotherapy. Cytogenetics and molecular analysis have shown trisomy 21 and GATA1 mutation restricted to leukemic cells. G-to-T transversion was detected, which is predicted to result in a premature stop codon (c.119G>T; pGlu67X) in diagnosis samples. These findings emphasize that there must be a powerful interaction between GATA1 and trisomy 21 in leukemogenesis process. PMID- 15654282 TI - High-dose IVIG therapy in a child with idiopathic thrombocytopenic purpura associated with mumps. PMID- 15654283 TI - Neonatal thrombosis associated with maternal antiphospholipid syndrome. PMID- 15654285 TI - A functional polymorphism in the promoter region of the tryptophan hydroxylase gene is associated with alcohol dependence in one aboriginal group in Taiwan. AB - BACKGROUND: Polymorphisms within intron 7 of the tryptophan hydroxylase (TPH1) gene were found to be associated with alcohol dependence in different ethnic groups, including the aboriginal Bunun group in Taiwan. This study aimed to identify genetic variants at the TPH1 locus and to examine their associations with alcoholism. We hypothesized that the polymorphism of TPH1 gene is functional and influences the human circadian rhythm to contribute to the pathophysiology of alcohol dependence. METHODS: DNA from the Taiwanese Han and Bunun was subjected to sequence for screening genetic variation in the coding and promoter regions of the TPH1 locus. Polymorphisms among individuals with alcohol dependence and control subjects in two ethnic groups in Taiwan were investigated. RESULTS: Three variants in the TPH1 promoter region were identified, and the markers are in complete linkage disequilibrium in both populations. Positive associations at both allelic and genotypic levels were obtained between case and control groups in the Bunun. Expression studies demonstrated that the variants indeed affected reporter gene activity in human choriocarcinoma and colon adenocarcinoma cell lines. CONCLUSIONS: Polymorphisms in the promoter region may influence the function of the TPH1 gene and further influence the proclivity of alcohol dependence in one ethnic group in Taiwan. The associations between TPH1 genotypes and alcoholism may deserve further investigation. PMID- 15654286 TI - An expanded evaluation of the relationship of four alleles to the level of response to alcohol and the alcoholism risk. AB - BACKGROUND: Alcoholism is a complex, genetically influenced disorder the cause of which may be better understood through the study of genetically influenced phenotypes that mediate the risk. One such intermediate phenotype is the low level of response (LR) to alcohol. This project used a case-control approach to search for genes that may contribute to LR. METHODS: Data were available from alcohol challenges at approximately age 20 and regarding the development of alcohol use disorders over the subsequent 20 years for 85 men, including 40 reported in a previous genetic analysis. LR was evaluated using oral consumption of 0.75 ml/kg of alcohol, after which changes in subjective feelings of intoxication and body sway were measured. Alcohol abuse and dependence were diagnosed by DSM-III-R criteria through structured interviews administered to both the participant and an informant (usually the spouse) 10, 15, and 20 years after initial testing. Four polymorphisms were evaluated, including the serotonin transporter HTTLPR promoter ins/del, GABAAalpha6 Pro385Ser, NPY Leu7Pro, and catalase 262C>T. Two of these, HTTLPR and GABAAalpha6 Pro385Ser, had been previously associated with LR and alcoholism in a preliminary study. RESULTS: The HTTLPR L allele was significantly related to both the LR and alcoholism in an allele-dosage (stepwise) manner. Furthermore, the association remained when L alleles were subdivided into recently reported functional subtypes: the lowest LR was associated with genotypes correlated with the highest serotonin transporter expression. The GABAAalpha6 Ser385 allele showed a nonsignificant trend for association to a low LR, as had been previously observed, although the Ser385 allele is uncommon, and only 18 heterozygotes were in the current group. However, the six men with both LL and Pro385/Ser385 genotypes had the lowest LR, and each had developed alcoholism during follow-up. Neither NPY nor catalase was associated with either LR or alcoholic outcomes, although the sample did not have sufficient power for definitive conclusions. CONCLUSIONS: This report strengthens the support for a relationship between the HTTLPR L and GABAAalpha6 Ser385 alleles to low alcohol LR and to alcoholism in a prospectively studied cohort evaluated for LR in young adulthood and before the onset of alcohol dependence. PMID- 15654287 TI - Glycine receptors regulate dopamine release in the rat nucleus accumbens. AB - BACKGROUND: The mesolimbic dopamine (DA) system seems to be centrally involved in regulating reward-related behavior and consequently has been implicated in addictive processes, such as alcoholism and drug addiction. This DA system has also been implicated in psychosis and in regulating hedonia/anhedonia, important components of mania and depression. Given the potentially great importance of the mesolimbic DA system for several psychiatric disorders, it is of major interest to delineate the mechanisms and dynamics underlying DA regulation and release. Recently strychnine-sensitive glycine receptors (GlyR) have attracted some interest in this matter. METHODS: Western blot and in vivo microdialysis (couplied to high-pressure liquid chromatography with electrochemical detection), as well as reversed microdialysis, in awake, freely moving, adult male Wistar rats. RESULTS: Here we demonstrate by means of Western blot that alpha GlyR subunit proteins are expressed in the rat nucleus accumbens (nAc), a major target of the mesolimbic DA system. We further show that reversed microdialysis of the competitive GlyR antagonist strychnine into the nAc concentration-dependently (2 200 microM) and in a reversible manner decreases accumbal extracellular DA levels. Conversely, reversed microdialysis of the agonist glycine increases accumbal DA levels in some rats but not others. The strychnine-induced depression of the accumbal DA levels is antagonized by simultaneous local perfusion of glycine. CONCLUSIONS: The present results indicate that GlyRs in the nAc are tonically activated and of importance for regulating extracellular DA levels. The possibility of pharmacologically interfering with GlyRs to combat psychiatric disorders, in which the mesolimbic DA system is implicated, such as alcoholism, drug addiction, and psychosis, should be explored. PMID- 15654288 TI - Accumbal strychnine-sensitive glycine receptors: an access point for ethanol to the brain reward system. AB - BACKGROUND: Ethanol (EtOH), like other drugs of abuse, increases extracellular dopamine (DA) levels in the nucleus accumbens (nAc) of the brain reward system, an effect that may be of importance for alcohol addiction. How this DA increase is produced is not fully understood, although previous studies from the present laboratories indicate that nicotinic acetylcholine receptors in the ventral tegmental area play an important role in mediating this effect. Furthermore, activation of these receptors may be secondary to some priming effect produced by EtOH in the nAc. We recently demonstrated that strychnine-sensitive glycine receptors (GlyRs) are present in the nAc and that they are involved in regulating extracellular DA levels. Here we examine the tentative role of these accumbal GlyRs in the above-mentioned priming mechanism of EtOH. METHOD: In vivo microdialysis (coupled to high pressure liquid chromatography with electrochemical detection) and reversed microdialysis, in awake, freely moving adult male Wistar rats. RESULTS: Local perfusion of strychnine decreased accumbal DA levels per se and completely prevented the increase of accumbal DA levels after both local and systemic EtOH administration. Accumbal perfusion of the GlyR agonist glycine instead increased DA levels in a subpopulation of rats and prevented the EtOH-induced increase after local but not systemic EtOH in all animals. CONCLUSION: The present results suggest that GlyRs in the nAc might constitute targets for EtOH in its mesolimbic DA-activating effect. Gene polymorphism and drug developmental studies that focus on this receptor population and its relation to alcohol dependence are warranted. PMID- 15654289 TI - Involvement of accumbal glycine receptors in the regulation of voluntary ethanol intake in the rat. AB - BACKGROUND: Extracellular dopamine (DA) levels in the nucleus accumbens (nAc) increase after ethanol (EtOH) administration in the rat, a response that may be involved in the positive reinforcing effects of EtOH. The mechanisms underlying this DA activation and how they relate to EtOH reinforcement remain to be elucidated, but recent data indicate that glycine receptors (GlyRs) in the nAc may be involved. Here this hypothesis was further challenged by examining the influence of bilateral accumbal application of glycine (a GlyR agonist), strychnine (a GlyR competitive antagonist), or Ringer on EtOH intake and preference, as well as on the concomitant DA output in the nAc, in EtOH high preferring male Wistar rats. METHODS: EtOH high-preferring male Wistar rats [EtOH preference >60% during continuous access to a bottle of EtOH (6% v/v) and a bottle of water] were limited to drink 1 hr/day (limited access drinking). Thereafter, the animals were equipped bilaterally with microdialysis probes aimed at the mAc, and were subjected to in vivo microdialysis (coupled to high-pressure liquid chromatography with electrochemical detection) and reversed microdialysis (for drug application) during two experimental days (balanced study), during which the animals were allowed a choice between EtOH and water. RESULTS: The EtOH consumption in rats that were perfused with Ringer in the nAc was approximately 0.9 g/kg/hr and associated with a significant increase in extracellular accumbal DA levels. In a subpopulation of rats, bilateral accumbal glycine (100 microM) perfusion produced a significant increase in accumbal DA output and a decrease in EtOH preference and intake. In these glycine responders, the EtOH consumed (approximately 0.7 g/kg/hr) did not produce a further increase of DA levels. In other rats, bilateral glycine perfusion did not change the accumbal DA output, and voluntary EtOH intake was not altered. In these glycine nonresponders, EtOH tended to increase accumbal DA levels. Bilateral accumbal strychnine (20 microM) perfusion significantly decreased DA output in the nAc, and the DA levels remained decreased despite a statistically significant increase of EtOH intake. Finally, the increase in accumbal DA levels observed after EtOH consumption in Ringer-treated rats was significantly larger in glycine responders than in glycine nonresponders. CONCLUSIONS: The present findings suggest that glycine and strychnine alter extracellular DA levels in the nAc, probably via GlyR stimulation and blockade, respectively, and concomitantly glycine and strychnine reciprocally alter also EtOH consumption in EtOH high-preferring male Wistar rats. The possibility of developing selective GlyR agonists and/or antagonists should be explored. Such agents could prove of value in the treatment of alcoholism. PMID- 15654290 TI - Systematic regional study of dopamine, norsalsolinol, and (R/S)-salsolinol levels in human brain areas of alcoholics. AB - BACKGROUND: Dopamine (DA)-derived tetrahydroisoquinolines (TIQs) are discussed as neurochemical factors of addiction processes in alcoholism. In a prospective study, the regional distribution of DA, (R)-salsolinol (SAL), and (S)-SAL, as well as norsalsolinol (NorSAL) was examined systematically in a large collective of human brain samples obtained by autopsy. METHODS: The material comprises 44 brains of alcoholics and 47 controls with 6 standardized specimens in each case. The analytes were determined after solid-phase extraction and enantioselective derivatization using gas chromatography-mass spectrometry. RESULTS: Levels of DA, (R/S)-SAL, and NorSAL in alcoholics did not differ significantly from those of the control group. A relationship between alcohol consumption and SAL formation could not be proved. Topical differences and no ubiquitous occurrence were encountered. Significant amounts of (R)-SAL and (S)-SAL as well as NorSAL only were found in DA-rich areas of the basal ganglia, whereas in other regions of the brain, no TIQs were detected. Especially in the nucleus caudatus, the concentrations of DA, SAL, and NorSAL decreased significantly with rising age. CONCLUSION: These findings do not support the hypothesis that one of the SAL enantiomers or NorSAL is involved in the genesis of alcoholism. However, they suggest that the concentration of the substrate DA may determine the alkaloid level during in vivo formation. The revealed data can serve as reference for other studies in humans concerning the cause of alcoholism or other neurodegenerative diseases with the involvement of TIQs. PMID- 15654291 TI - Alcohol Use Disorders Identification Test (AUDIT): establishment of cut-off scores in a hospitalized Chinese population. AB - BACKGROUND: Alcohol use has become a problem for Taiwanese society. Developing a brief, rapid, and flexible tool to screen an individual's alcohol consumption is important. Many countries use the Alcohol Use Disorders Identification Test (AUDIT) to screen for harmful and dangerous alcohol consumption. The psychometric characteristics of the AUDIT have not been examined in a Chinese population. METHODS: Determination of the cutoff points for the AUDIT Chinese version included three stages: translating the questionnaire, expert review and formal testing on subjects. Participants (N = 112) were recruited from a medical research center of four gastroenterology wards in northern Taiwan. RESULTS: The cutoff point for participants diagnosed as "harmful users" was an AUDIT score of 8. The sensitivity was 0.96, specificity was 0.85, positive predictive value (PPV) was 0.85, negative predictive value (NPV) was 0.96, and area under the receiver-operating characteristic curve (AUROC) was 0.93. The cutoff point for participants diagnosed as "alcohol dependent" was an AUDIT score of 11. The sensitivity was 0.94, specificity was 0.63, PPV was 0.31, NPV was 0.98, and AUROC was 0.84. Furthermore, males had significantly higher AUDIT and AUDIT-C scores. Males were also significantly more likely than females to be diagnosed as harmful users or alcohol dependents. CONCLUSIONS: The Chinese version of the AUDIT gave the same cutoff point for harmful alcohol use by Taiwanese individuals as that set by the World Health Organization. This finding shows that this cutoff point is generally appropriate in screening for problem alcohol consumption. Moreover, the cutoff AUDIT score of 11 for alcohol dependence provides a reference for screening in Taiwanese clinics. PMID- 15654292 TI - Mecamylamine and ethanol preference in healthy volunteers. AB - BACKGROUND: Recent evidence suggests that some of the behavioral effects of alcohol may be mediated through actions on nicotinic acetylcholine receptors. Mecamylamine, a nicotinic acetylcholine receptor antagonist, reduces alcohol preference and consumption in alcohol-preferring rats, and in humans, mecamylamine dampens some of the subjective, or mood-altering, effects of alcohol. This experiment was designed to investigate the effects of mecamylamine on consumption of alcohol in healthy social drinkers. METHODS: Healthy volunteers (12 men, 12 women) participated in a choice procedure in which they chose between an alcoholic beverage and money (low, medium, or high amounts) after pretreatment with mecamylamine (7.5 or 15 mg) or placebo. Outcome measures were the number of alcoholic beverages consumed and the subjective effects of alcohol. RESULTS: Mecamylamine (15 mg) decreased blood alcohol levels (BALs) after a small fixed dose of alcohol (0.2 g/kg). Even when the lower BALs were taken into account, mecamylamine reduced ratings of stimulation after alcohol (Addiction Research Center Inventory A scale). Mecamylamine did not significantly reduce choice for alcohol versus money. However, there was a tendency for the drug to decrease alcohol choice among participants who reported the greatest stimulant-like effects from alcohol. CONCLUSION: These results provide only limited support for the idea that nicotinic acetylcholine receptors are involved in the rewarding effects of alcohol. PMID- 15654293 TI - Electrophysiological effects of allopregnanolone in rats with a history of ethanol exposure. AB - BACKGROUND: Sensitivity to the anticonvulsant effects of allopregnanolone (ALLO) is enhanced during the early phase of ethanol (EtOH) withdrawal. However, it is unclear whether this enhanced sensitivity generalizes to ALLO's neurobehavioral effects during protracted abstinence. The purpose of this study was to examine the neurophysiological effects of ALLO in rats with a history of chronic EtOH exposure after a protracted period of abstinence. METHODS: Male Wistar rats were exposed to EtOH vapor for 14 hr/day for 5 weeks. Blood EtOH levels were maintained between 200 and 250 mg/dl. The effects of ALLO (0.0-10 mg/kg, intraperitoneally) on motor activity, the electroencephalogram (EEG), and auditory event-related potentials then were assessed after 6 to 8 weeks of abstinence from EtOH. RESULTS: ALLO's effects on the EEG were consistent with previous studies and were unaffected by EtOH exposure. ALLO increased high frequency EEG power and shifted peak EEG frequencies in a benzodiazepine- and barbiturate-like manner in both the cortex and the hippocampus. The effects of ALLO on event-related potentials were attenuated in rats with a history of EtOH exposure. Low doses of ALLO (1 and 5 mg/kg) reduced cortical P1 amplitude in response to the standard tone but only in the control group. ALLO also increased N1 amplitude in the hippocampus of the control group while having no significant effect in EtOH-exposed rats. Low doses of ALLO (1 and 5 mg/kg) were found to increase motor activity. CONCLUSIONS: These data indicate that a history of EtOH exposure attenuates some of the neurophysiological effects of ALLO in a manner consistent with cross-tolerance. Taken together, these data suggest that increased sensitivity to ALLO's neurobehavioral effects is limited to the early phases of EtOH withdrawal and may not extend to more protracted periods of abstinence. PMID- 15654294 TI - Effects of topiramate on ethanol and saccharin consumption and preferences in C57BL/6J mice. AB - BACKGROUND: Topiramate has recently been found to be more effective than placebo as an adjunct treatment for alcohol dependence, but it has not yet been investigated in animal models of ethanol consumption. The current experiment examined the effects of topiramate on ethanol drinking in mice using a continuous access, two-bottle choice procedure. METHOD: C57BL/6J male mice were offered a 10% v/v ethanol solution versus tap water over 4 consecutive days per week. Mice were assigned to topiramate (1-50 mg/kg) or saline groups and received injections before the beginning of the dark phase of the light cycle. Topiramate dose increased over 5 successive weeks (1, 5, 10, 25, and 50 mg/kg). Fluid intake was measured 2, 4, and 23 hr after injection. Body weight and food intake were measured at the time of injection. In a second phase, mice were offered saccharin solutions (0.2 and 2.5% w/v) versus tap water after topiramate (50 mg/kg) or saline injections. RESULTS: Results revealed that high topiramate doses (25 and 50 mg/kg) increased water intake and decreased ethanol preference. Compared with saline controls, topiramate produced dose-dependent, bidirectional effects on ethanol dose, with 25 mg/kg of topiramate increasing ethanol dose at 4 and 23 hr after injection but 50 mg/kg topiramate decreasing ethanol dose at 2 hr after injection. During saccharin exposure, topiramate decreased saccharin preference (for 2.5% w/v saccharin solution) and marginally increased water intake but did not directly alter intake of the saccharin solutions. Topiramate had no effects on body weight or daily food intakes. CONCLUSIONS: Topiramate reduced ethanol preference in C57BL/6J mice, but this effect was primarily attributable to elevated water intake. Topiramate also reduced saccharin preference, likely through marginally significant increases in water intake. Increases in water intake and bidirectional effects of topiramate on ethanol dose complicate conclusions with regard to the effects of topiramate on ethanol reward. PMID- 15654295 TI - Performance of asialotransferrin in detecting alcohol abuse. AB - BACKGROUND: The spectrum of alcohol use disorders covers hazardous use, alcohol abuse, and alcohol dependence. The present study evaluated the performance of asialotransferrin, a newly proposed biomarker for alcohol use disorders, in detecting alcohol abuse and alcohol dependence. METHOD: A 4-month trial was conducted in three groups of participants: alcohol abusers and alcohol-dependent patients, as defined in DSM-IV, and a control group. Asialotransferrin was assayed by capillary zone electrophoresis. RESULTS: Asialotransferrin demonstrated a sensitivity of 0.34 and a specificity of 1.00 for alcohol abuse. The sensitivity of asialotransferrin increased to 0.57 in alcohol-dependent patients. CONCLUSION: Despite the high specificity of asialotransferrin in alcohol use disorders, its sensitivity is too low to make it a useful marker of alcohol abuse. PMID- 15654296 TI - Alcohol and atherosclerotic vascular disease risk factors in French men: relationships are linear, J-shaped, and U-shaped. AB - BACKGROUND: Although it is well admitted that alcohol displays a U-shaped relationship with atherosclerotic vascular disease, individual relationships between alcohol and atherosclerosis risk factors may be different and have not been determined precisely for several of them. METHODS: A cross-sectional study within the SU.VI.MAX French cohort study was performed to assess the curve of potential relationships between alcohol and atherosclerosis risk factors in 2126 healthy men. Mean daily alcohol intake was derived from 37 alcoholic beverages in twelve 24-hr dietary recalls. Logistic models were adjusted for age. RESULTS: Apolipoprotein B (ApoB), fasting glucose, body mass index, waist-to-hip ratio, and waist circumference displayed a linear relationship with alcohol. The odds ratios and 95% confidence intervals associated with abnormal values of the markers for the highest quintile of alcohol intake were 1.45 (1.06-1.97) for ApoB, 1.98 (1.40-2.80) for fasting glucose, and 1.74 (1.30-2.34) for body mass index. An inverse J-shaped relationship was assumed for ApoA1 and ApoB/ApoA1 ratio, whereas a U-shaped relationship was observed for serum triglycerides and mixed hyperlipidemia. Only the highest quintile of alcohol was associated with hypertension, although the test for linearity was also significant. No association was observed for Lp(a) or homocysteine. Associations were unmodified by further adjustment for carbohydrates, fiber, lipids, tobacco, or exercise. CONCLUSIONS: The aggregate of the disparate alcohol risk factor relationships suggests probable net benefit at 15 to 25 g of alcohol/day. PMID- 15654297 TI - Trends and determinants of alcohol consumption in Portugal: results from the national health surveys 1995 to 1996 and 1998 to 1999. AB - BACKGROUND: There is very little information on trends and determinants of alcohol consumption in the Portuguese population, which is usually characterized by high wine consumption. METHODS: A cross-sectional studies was conducted in 1995/1996 and 1998/1999 in a representative sample of 0.5% of the mainland Portuguese population (49,768 participants in 1995/1996 and 48,606 in 1998/1999), aged 15 years or more. Alcohol consumption was assessed by asking whether the participants had consumed alcohol in the previous week and how many drinks of wine/beer/whiskey/Port wine they consumed on average during that week. RESULTS: Prevalence of reported alcohol consumption decreased slightly between 1995/1996 and 1998/1999 (men: 65.7 vs. 64.0%, p < 0.001; women: 26.9 vs. 26.0%, p < 0.001). Among drinkers, the most frequently consumed alcoholic beverage was wine, followed by beer, whiskey, and Port wine. The amount of alcohol and wine consumed decreased in both sexes, whereas the amount of beer, whiskey, and Port wine consumed increased in men and the increase in beer consumption was borderline significant in women (p = 0.056). In both sexes, participants <50 years of age tended to consume less wine and more beer, whiskey, and Port wine than their older counterparts. Also, higher education was related to a higher frequency of alcohol consumption, whereas smoking was related to a lower consumption of wine [odds ratio (OR): 0.69 (95% confidence interval [CI]: 0.62-0.77) for men and OR: 0.76 (95% CI: 0.61-0.95) for women] and a higher consumption of beer [OR: 1.43 (95% CI: 1.33-1.54) for men and OR: 2.13 (95% CI: 1.84-2.42) for women and whiskey [OR: 1.28 (95% CI: 1.21-1.35) for men and OR: 2.61 (95% CI: 2.25-3.02) for women]. CONCLUSIONS: The pattern of alcohol consumption is changing in Portugal: the prevalence of drinkers is decreasing, and younger generations are shifting from wine to beer and spirits. Educational level seems to be a powerful mediator in the choice of alcoholic beverage. PMID- 15654298 TI - Drinking, alcohol problems and the five-year recurrence and incidence of male to female and female to male partner violence. AB - BACKGROUND: This study examined the 5-year incidence and recurrence of male to female (MFPV) and female to male partner violence (FMPV) as well as their relationship with drinking and alcohol problems among intact couples in the United States. METHODS: A national sample of couples 18 years of age or older were interviewed in 1995 and again in 2000. RESULTS: Recurrence is slightly higher for FMPV (44%) than MFPV (39%), whereas incidence rates are similar for these two types of violence (MFPV, 5.7%; FMPV, 6%). Cross-tabulations show that a higher frequency of drinking five or more drinks on occasion is positively associated with the overall occurrence of MFPV and with both the recurrence and the overall occurrence of FMPV. Male alcohol problems are associated with a higher recurrence of MFPV and higher overall MFPV. Female alcohol problems are associated with incidence of FMPV. In multivariate analysis, black ethnicity, male unemployment, and severe physical abuse during childhood are associated with recurrence of MFPV. Black ethnicity, male unemployment, male employment status as "retired/other," female age, and couples in which the female drinks more are associated with recurrence of FMPV. Incidence of MFPV is associated with cohabitation, Hispanic ethnicity, and man's observation of violence between parents. Male unemployment, male observation of violence between parents, and man's drinking volume predict incidence of FMPV. CONCLUSIONS: Volume of drinking is the only alcohol indicator associated with intimate partner violence once the effects of other factors are controlled in multivariate analysis. Both MFPV and FMPV are areas of health disparity across whites, blacks, and Hispanics. Factors of risk that predict recurrence and incidence can be identified and used in prevention efforts. PMID- 15654299 TI - Disparities in patterns of alcohol use among reservation-based and geographically dispersed American Indian populations. AB - BACKGROUND: High rates of alcohol use and alcohol-related morbidity and mortality among American Indians (AI) are major public health concerns. The purpose of this paper is to describe patterns of alcohol consumption among three distinct samples of American Indians (AIs) compared to a U.S reference population. METHODS: Data were drawn from two epidemiologic studies: 1) a study of 2,927 AIs living on or near reservations from two culturally distinct tribes in the Southwest (SW-AI) and Northern Plains (NP-AI); and 2) the National Longitudinal Alcohol Epidemiologic Study (NLAES), which included data from a geographically dispersed sample of AIs (n = 780) as well as the US reference population (all-race excluding AIs, n = 30,063). Multivariate analyses were used to assess drinking patterns. RESULTS: After controlling for demographic characteristics, the prevalence of drinking during the past year was similar among males in the NP-AI, NLAES-AI, and the US populations. SW-AI males and females were significantly less likely to drink during the past year (Odds Ratios of 0.74 and 0.41, respectively), while the odds of NP-AI females being current drinkers were twice that of US females. Among those who drank during the past year, the AIs consumed a larger quantity of alcohol per drinking day than the US reference population. However, the reservation-based AIs consumed alcohol less frequently (Odds Ratios between 0.18-0.40, p < 0.01) than the NLAES-AI and US populations. CONCLUSIONS: Patterns of alcohol consumption varied substantially between the two reservation based AI populations, the geographically dispersed sample of AIs, and the US reference population. Careful consideration of these variations may improve the effectiveness of alcohol prevention and treatment programs as they may reflect important underlying differences in the cultures of alcohol consumption across these populations. PMID- 15654300 TI - Vitamin E does not protect against neonatal ethanol-induced cerebellar damage or deficits in eyeblink classical conditioning in rats. AB - BACKGROUND: Rodent studies have shown that heavy binge-like ethanol (EtOH) exposure during the brain growth spurt [postnatal days (PD) 4-9] causes cerebellar neuronal loss and deficits in cerebellar-mediated eyeblink classical conditioning (ECC). Oxidative stress has been implicated in EtOH-mediated brain damage, and studies using vitamin E have reported amelioration of EtOH-induced tissue damage, including protection in rats against EtOH-induced cerebellar Purkinje cell (PC) loss on PD 4 to 5. The purpose of this study was to determine whether dietary supplementation with vitamin E concurrent with binge EtOH exposure on PD 4 to 9 in rats would attenuate the cerebellar cell death and ECC deficits. METHODS: Rat pups were given one of five different neonatal treatments: (1) intubation with EtOH in milk formula (twice daily, total dose 5.25 g/kg/day), (2) intubation with EtOH in milk formula supplemented with vitamin E (12.26 mg/kg/feeding), (3) intubation with milk formula that contained vitamin E only, (4) sham intubations, or (5) normally reared unintubated controls. Between PD 26 and 33, subjects received short-delay ECC for 3 consecutive days. Unbiased stereological cell counts were performed on cerebellar PCs of left cerebellar lobules I to VI and neurons of the interpositus nucleus. In a separate study with PD 4 pups, the effects of vitamin E on EtOH-induced expression of caspase-3 active subunits were assessed using Western blot analysis. RESULTS: EtOH-treated groups showed significant deficits in acquisition of conditioned eyeblink responses and reductions in cerebellar PCs and interpositus nucleus neurons compared with controls. Vitamin E supplementation failed to protect against these deficits. Vitamin E also failed to protect against increases in caspase-3 active subunit expression induced by acute binge EtOH exposure on PD 4. CONCLUSIONS: In contrast to the previously reported neuroprotective potential of antioxidants on EtOH-mediated cerebellar damage, vitamin E supplementation did not diminish EtOH induced structural and functional damage to the cerebellum in this model of binge EtOH exposure during the brain growth spurt in rats. PMID- 15654301 TI - Alcohol consumption in pregnant, black women is associated with decreased plasma and erythrocyte docosahexaenoic acid. AB - BACKGROUND: Inner-city, black women are among those groups that are at higher risk for having infants with fetal alcohol spectrum disorders that can include life-long neurobehavioral and cognitive impairments. Chronic alcohol consumption can decrease amounts of docosahexaenoic acid (DHA), a fatty acid that is essential for optimal infant neural and retinal development in a variety of tissues. METHODS: Black women who presented at an inner-city antenatal clinic for their first prenatal visit were recruited into a longitudinal, observational study. Alcohol intake was determined by a structured interview. Participants provided blood specimens and completed food frequency surveys at 24 weeks of gestation, infant delivery, and 3 months postpartum. Fatty acid composition analyses were completed on 307, 260, and 243 for plasma and 278, 261, and 242 erythrocyte specimens at 24 weeks of gestation, delivery, and 3 months postpartum, respectively. RESULTS: Proportion of drinking days at the first prenatal visit was associated with decreased DHA in plasma and erythrocytes throughout the study. This association was the strongest at 24 weeks of gestation. In addition, an interaction between proportion of drinking days at the time of conception and ounces of absolute alcohol per drinking day at the time of conception was detected and demonstrated that, in daily drinkers, high intakes of alcohol are associated with decreased DHA and arachidonic acid (AA) concentrations in plasma. CONCLUSIONS: Frequent and high intakes of alcohol that have been previously associated with fetal alcohol spectrum disorders are also associated with decreased maternal DHA and AA plasma concentrations. The present findings indicate that maternal DHA deficiency is associated with high-risk drinking and may contribute to the mechanism(s) of alcohol-related neurodevelopmental disorders. PMID- 15654302 TI - Fetal alcohol exposure reduces serotonin innervation and compromises development of the forebrain along the serotonergic pathway. AB - BACKGROUND: We reported previously that a moderate level of fetal alcohol treatment reduces the birth, maturation, and migration of serotonin (5-HT) neurons at embryonic days 11 to 15 (E11-E15). Because 5-HT is known as a differentiation signal for forebrain development, we investigated whether alcohol affects 5-HT innervation to the developing brain and how the target brain areas grow as they receive 5-HT innervation between E15 and E18. METHODS: Pregnant dams were divided into three groups and treated from E7 to E15 or E18 with one of the following conditions: (1) liquid diet that contained 25% ethanol-derived calories (ALC), (2) isocaloric liquid diet pair-fed (PF), or (3) chow fed (Chow). The 5-HT immunostained (5-HT-IM) fibers and size of brain areas were examined as an index of growth along the ascending 5-HT pathway. RESULT: We found that 5-HT-IM fibers innervate the brain regions specifically under active differentiation and that there were three sets of correlated dysmorphology in the ALC group as compared with those of the PF and Chow groups. The three sets are as follows: (1) fewer 5 HT-IM fibers in the medial forebrain bundle and along the projecting pathway through the hypothalamus, septal nucleus, frontal and parietal cortices, and subiculum/hippocampus; (2) underdevelopment of the brain regions along 5-HT fiber projections; and (3) underdevelopment of somatosensory thalamocortical projections, which are known to transiently express 5-HT transporters and to be regulated by 5-HT. No such differences were found between the PF and Chow groups. CONCLUSION: We found that fewer 5-HT fibers grew in the embryos that were exposed to alcohol. As forebrain regions differentiated along the 5-HT projection, we found two reductions: (1) the growth of brain regions along 5-HT projection and (2) the growth of the thalamocortical sensory projection regulated by 5-HT. The reduced 5-HT innervation is in agreement with our previous observation of fewer 5 HT neurons. The subsequent retardation of forebrain growth and sensory thalamocortical fibers along the pathway of reduced 5-HT projection is consistent with the role of 5-HT as a signal for forebrain differentiation. PMID- 15654303 TI - Experimental bypass surgery between the spinal cord and caudal nerve roots for spinal cord injuries. AB - Spinal cord injuries often cause permanent neurological deficits and are still considered as inaccessible to efficient therapy. Injured spinal cord axons are unable to spontaneously regenerate in adult mammalians. Re-establishing functional activity especially in the lower limbs by reinnervating the caudal infra-lesional territories could represent an attractive therapeutic strategy. For several years, we have studied and developed surgical bypasses using peripheral nerve grafts bridging the supra-lesional rostral spinal cord to the caudal infra-lesional lumbar roots. Main objectives were: 1- to overcome the spinal cord lesion and the consecutive glial barrier blocking the axonal regeneration; 2- to find and bring an alternative source of regenerating axons; 3 to guide those axons toward precisely definite targets (for example, lower limb muscles). We report here the results of our experimental research, which led us from animal experimental models (rodents, primates) to the first human experimentation. Limitations of the method (especially technical pitfalls) are numerous. However, we have obtained encouraging results in our attempts to "repair" the motor pathway. Functional recovery with strong evidence of centrifugal axonal regeneration from the spinal cord to the periphery has been observed. Regarding the sensory pathway, we have found evidence of centripetal axonal regeneration from the periphery toward the spinal cord. Further studies are obviously advocated, but our experimental model of spinal cord - nerve roots bypasses may be integrated in future "repair" strategies of both motor and sensory pathways following spinal cord injury. PMID- 15654304 TI - [Pleomorphic xanthoastrocytoma. A review of five observations]. AB - BACKGROUND AND PURPOSE: We present a review of five patients who developed pleomorphic xanthoastrocytomas, presenting the clinical features, pathology findings and therapeutic approach of these rare tumours which almost always occur in children and young adults. METHODS: This paper is a retrospective study of five patients (three males and two females), treated from 1985 to 1996. RESULTS: Seizure was the most frequent symptom. The tumor was located in the temporal or temporo-parietal lobe in two patients, in the parietal lobe in one, the frontal in one and in hypothalamus and third ventricle in the final patient. To our knowledge, no case of pleomorphic xanthoastrocytoma located in the area of hypothalamus/third ventricle has been reported in the literature. All patients underwent surgery which was complete for two patients, subtotal for two others and partial for one. Four were given post-operative cerebral radiation therapy (two recurrences and malignant transformation and two erroneous diagnoses) (mean dose: 44.75 Gy), for two patients surgery was followed by chemotherapy. One patient was lost for follow-up. Two patients presented at least one tumor recurrence. Two died from malignant transformation. Two patients are alive without recurrence at 12 and 16 years from diagnosis with excellent quality-of life (Glasgow Outcome Scale=I). CONCLUSION: Surgery is the gold standard treatment. Prognosis is very good (except for malignant transformation), so that a close long-term clinical and radiological follow-up is mandatory. PMID- 15654306 TI - [Intramedullary tuberculomas. Five cases]. AB - Tuberculosis of the central nervous system is uncommon. The intramedullary localization is exceptional. We report five cases of intramedullary tuberculosis observed in four women and one man with a mean age of 43 years. Two patients had a prior history of tuberculosis. Spinal cord compression was found clinically in all cases. The spinal MRI visualized the tuberculoma in all patients; two had a double tumor. Complete removal of the tuberculoma was possible in only one patient. Outcome was stationary for four patients. PMID- 15654305 TI - [Retro-odontoid synovial cyst with Forestier's disease]. AB - STUDY DESIGN: We report one case of 66-year-old man with Forestier disease and spinal cord compression by retro-odontoid synovial cyst. OBJECTIVES AND METHODS: Although retro-odontoid synovial cysts remain rare, an increasing number have been reported in the literature. Affecting adults in the sixth decade of life, retro-odontoid synovial cysts produce slowly progressive upper spinal cord symptoms. Diagnosis can be achieved by means of magnetic resonance imaging (MRI). Association with Forestier's disease has been reported in only one previous study. Cyst probably develop as a result of enhanced mechanical stress on the only remaining mobile joint. In the literature treatment of retro-odontoid mass associated with Forestier has usually involved occipito-cervical fusion with transoral decompression. RESULTS: In this report we describe a patient treated by the posterolateral route with good short- and mid-term clinical and radiological outcome. CONCLUSIONS: In comparison of transoral route, the advantages of the posterolatral route are shorter hospitalization and lower morbidity especially in elderly patients with operative risk factors. PMID- 15654307 TI - [Perineal pain]. AB - The investigation of patients suffering from perineal pain when sitting led us to perform an anatomical study of the pudendal nerve. We dissected 50 cadavers and found areas of conflict for the nerve fibers. The nerve trunk can become entrapped at the level of the ischiatic spine, in the Alcock's canal and when it crosses the falciform process. Considering the clinical and neurophysiological data, this type of chronic pain may arise from compression of the nerve between the sacro-tuberal and the sacro-spinal ligaments, and/or in the fascia of the internal obturator muscle. Much like treatment of entrapment of the median nerve in the wrist, we decided to treat chronic perineal pain by nerve blocks, and later by surgery. We describe here the clinical symptoms, the neurophysiological data, and the technique of the nerve blocks. For patients with persistent pain, we propose a posterior surgical approach which has provided successful pain relief in two third of patients. PMID- 15654308 TI - [French neurosurgical practice in Neuro-Oncology (national survey--part I)]. AB - OBJECTIVE: The aim of this work is to summarize the elements of the "Cancer Plan" applicable to neurosurgical practice, and to give the results of a national inquiry concerning the daily practice of Neuro-Oncology from the neurosurgical point of view. METHOD: The Neuro-Oncology Group of the French Society of Neurosurgery has submitted a questionnaire to every department of Neurosurgery in France. RESULTS: The response rate of the public centers was 96.5%. Moreover, responses were available from 7 private centers. The results are detailed in the text. CONCLUSION: This national survey highlights the interest and implication of French neurosurgeons in the field of Neuro-Oncology. But also, to be in accordance with the guidelines for good clinical practice, the importance of developing official neuro-oncological networks in order to offer the best access to clinical and fundamental data and hence optimise patient's care. The publication of the "Cancer Plan", the creation of a National Neuro-Oncology Group, and the results of this survey (actual multidisciplinary approach, better information and transparency, individualized care of the patients), are in the line with updating our daily practice, even though discrepancies remain among centers. French neurosurgeons must continue along the same path, but at the same time there is a need for additional help to definitely reach a truly, and homogeneous, optimized care of neuro-oncological patients. PMID- 15654310 TI - [Unemployment and health: a major issue for public statistics]. PMID- 15654311 TI - [Effect of a participation in a community-based nutrition program on children's outcomes]. AB - BACKGROUND: This study aimed to assess the effectiveness of a protein-energetic malnutrition prevention and treatment community program on the nutritional status of 18 to 36-month-old children in the poor communities of the Mono Region in Benin. METHODS: Two cross sectional anthropometric surveys were conducted. The first survey took place in 1998, before the implementation of the program; the second survey was conducted three years later. At each survey, cluster samples proportional to the population size was done; the sizes of the samples were 617 in 1998 and 648 in 2001. The nutritional status of 18 to 36 month-old-children was assessed by z-score of the indexes height-for-age, weight-for-age and weight for-height and the cut-off point of -2 standard deviation was used to identify children who had stunting, wasting or underweight; children with serious stunting, wasting or underweight were identified with the cut-off point of -3 standard deviation. RESULTS: From 1998 to 2001, there was a significant reduction in the prevalence of growth retardation (-13.1%) and underweight (-13.0%). The reductions observed for the three types of protein-energetic malnutrition were more pronounced in the 30 to 36 months age group than in the others age groups. CONCLUSION: Substantial improvement was observed during the three years activities. This improvement cannot be attributed to the program unconditionally, because there was no difference between the nutritional status of participant and non participant children. PMID- 15654312 TI - [Gynecological and breast cancer in the department of Somme]. AB - BACKGROUND: Breast and gynecological tumors are the most common cancers in women. The aim of this study was to show the epidemiologic features of gynecological and breast cancers in the French administrative district of La Somme. METHODS: This study focused on the 1982-1999 period. Incidence, mortality and survival rates were calculated. RESULTS: In 1997-1999, the world standardized breast incidence and mortality rates were 81.6 and 20.2 per 100,000 females per year. Breast and genital tract cancers accounted for 47% of all cancers in women. The incidence and mortality of uterine cervix cancers showed a clear decline over the past 10 years, whereas the trend of breast cancers was dominated by continuing increase. However, mortality was stable for breast cancers. Five year relative survival rates were respectively 80% for breast cancers, and 68%, 76%, 38%, for uterine cervix, uterine body and ovary cancers respectively. Incidence and mortality rates in Somme were in the middle risk range of other cancer French registries. CONCLUSION: The results of this study indicate that genital tract and breast cancers constitute a serious public health problem pointing out the importance of screening activities in the Somme area. PMID- 15654313 TI - [Assessment of the discriminating power of identifiers for record linkage]. AB - BACKGROUND: To reconstruct a patient's medical history, one has often to combine information from different sources, whatever the context of this reconstitution: epidemiological studies or health care. As a linkage using less informative identifiers could lead to linkage errors, it is essential to quantify the information associated to each identifier. METHODS: The aim of this study was to estimate the discriminating power of different identifiers which could be used in a record linkage process based on the means of the likelihood ratio proposed by Jaro as a probabilistic record linkage method. Six identifiers were considered: date of birth, maiden name, usual last name, first and second christian names and the gender. Two types of phonetic treatment were taken into account: the Soundex and a phonetic treatment adapted to the French language. Three situations were considered: 1) and 2) linkage of the data collected during two consecutive years in a university hospital (CHU de Dijon; 100000x100000 records) and a Paris hospital (50000x50000 records), 3) linkage of two files obtained through a healthcare network (Burgundy Perinatal Network; 200x2500 records). RESULTS: Whatever the situation, this work showed the interest of three identifiers when linking data concerning a same patient. The date of birth had the best discriminating power followed by the first and the last names. Including a poorly discriminating identifier like gender did not improve the results. Moreover, adding a second Christian name, often missing, increased linkage errors. On the contrary, it seemed that using a phonetic treatment adapted to the French language could slightly improve the results of linkage in comparison to Soundex. CONCLUSION: Whatever the method used, it seems necessary to improve the quality of identifier collection, in particular of the date of birth and of the first and last names as it could make the linkage of data obtained from different sources easier. Further research is needed to estimate the discriminating power of other identifiers (birth place and parents identifiers). PMID- 15654314 TI - [Linear factor analytic models for reliability analysis of composite variables]. AB - BACKGROUND: Confirmatory factor analysis allows testing whether a composite variable may be considered as a reliable measure of a psychological attribute which is defined within a population. METHODS: Models for parallel, tau equivalent and congeneric measurements are presented along with their reliability coefficients. RESULTS: When the variables are not tau equivalent, the averaged inter-item correlation should be preferred to coefficient alpha, which is not an estimator of the reliability of the corresponding data. As a rule, interpretation of a coefficient as a reliability coefficient requires that the corresponding structural model of the composite be known. Beyond unidimensionality, simultaneous analysis of several congeneric variables through the use of cross sectional or longitudinal hierarchical models entails fragmenting the theoretical variables. CONCLUSION: Interpreting a composite variable whose theoretical structure is corroborated by a hierarchical model may raise some difficulties because of its multidimensionality. Reliability formulae which account for this fragmentation are detailed. PMID- 15654315 TI - [Analysis of repeated binary data: sensitivity to missing data]. AB - BACKGROUND: In longitudinal studies, it is extremely rare that all the planned measurements are actually performed. Missing data are often consecutive to drop outs, but may also be intermittent. In both cases, the analysis of incomplete data necessarily requires assumptions that are generally unverifiable, and the need for sensitivity analyses has been advocated over the past few years. In this article, the attention will be given to longitudinal binary data. METHODS: A method is proposed, which is based on a log-linear model. A sensitivity parameter is introduced that represents the relationship between the response mechanism and the missing data mechanism. It is recommended not to estimate this parameter, but to consider a range of plausible values, and to estimate the parameters of interest conditionally on these plausible values. This allows to assess the sensitivity of the conclusion of a study to various assumptions regarding the missing data mechanism. RESULTS: This method was applied to a randomized clinical trial comparing the efficacy of two treatment regimens in patients with persistent asthma. The sensitivity analysis showed that the conclusion of this study was robust to missing data. PMID- 15654316 TI - [Health of the unemployed in France: literature review]. AB - BACKGROUND: This paper reviews and analyses the main publications concerning health and mortality of the unemployed in France, in order to formulate research orientations for future studies or secondary data analysis. METHODS: The conceptual framework was first set up by assembling the different hypotheses concerning the link between health and unemployment (causality versus selection). A review of the different studies was organized by type: health studies based on data from either cross-sectional health surveys at a national or regional scale, or longitudinal surveys, and mortality studies based on data from follow-up of census samples. RESULTS: Those studies confirm the health disadvantage of the unemployed relative to the employed, in terms of self-perceived health, morbidity, health services utilization and mortality. They provide arguments in favor of both the causality and selection hypotheses. CONCLUSION: The exact nature of the health conditions and causes of death associated with unemployment needs further clarification, as well as the causal pathways. PMID- 15654317 TI - [Drawlines for the surveillance of breastfeeding in France]. PMID- 15654318 TI - Pairing, connecting, exchanging, pausing and pulling chromosomes. PMID- 15654319 TI - Two-fold repeated (betaalpha)4 half-barrels may provide a molecular tool for dual substrate specificity. AB - Some bacterial genomes contain an incomplete set of genes encoding phosphoribosyl isomerases, raising the question of whether there exists broadened substrate specificity for the missing gene products. To investigate the underlying molecular principles of this hypothesis, we have determined the crystal structure of the bifunctional enzyme PriA from Streptomyces coelicolor at 1.8 A resolution. It consists of a (betaalpha)(8)-barrel fold that is assembled by two symmetric (betaalpha)(4) half-barrels. The structure shows how its active site may catalyse the isomerization reactions of two different substrates, and we provide a plausible model of how the smaller of the two substrates could be bound in two different orientations. Our findings expand the half-barrel ancestor concept by demonstrating that symmetry-related half-barrels could provide a smart solution to cope with dual substrate specificity. The data may help to unravel molecular rationales regarding how organisms with miniature genomes can keep central biological pathways functional. PMID- 15654321 TI - Reconstruction of cellular signalling networks and analysis of their properties. AB - The study of cellular signalling over the past 20 years and the advent of high throughput technologies are enabling the reconstruction of large-scale signalling networks. After careful reconstruction of signalling networks, their properties must be described within an integrative framework that accounts for the complexity of the cellular signalling network and that is amenable to quantitative modelling. PMID- 15654320 TI - Crystal structure of Mil (Mth680): internal duplication and similarity between the Imp4/Brix domain and the anticodon-binding domain of class IIa aminoacyl-tRNA synthetases. AB - Proteins of the Imp4/Brix superfamily are involved in ribosomal RNA processing, an essential function in all cells. We report the first structure of an Imp4/Brix superfamily protein, the Mil (for Methanothermobacter thermautotrophicus Imp4 like) protein (gene product Mth680), from the archaeon M. thermautotrophicus. The amino- and carboxy-terminal halves of Mil show significant structural similarity to one another, suggesting an origin by means of an ancestral duplication. Both halves show the same fold as the anticodon-binding domain of class IIa aminoacyl tRNA synthetases, with greater conservation seen in the N-terminal half. This structural similarity, together with the charge distribution in Mil, suggests that Imp4/Brix superfamily proteins could bind single-stranded segments of RNA along a concave surface formed by the N-terminal half of their beta-sheet and a central alpha-helix. The crystal structure of Mil is incompatible with the presence, in the Imp4/Brix domain, of a helix-turn-helix motif that was proposed to comprise the RNA-binding moiety of the Imp4/Brix proteins. PMID- 15654322 TI - Assembly and function of RNA silencing complexes. AB - In the RNA-interference pathway, double-stranded RNA induces sequence-specific mRNA degradation through the action of the RNA-induced silencing complex (RISC). Recent work has provided our first glimpses of the RISC-assembly pathway and uncovered the biochemical roles of critical RISC components. These advances have taken our mechanistic understanding of RNA interference to a new level and promise to improve our ability to exploit this biological process for use in experimental biology and medicine. PMID- 15654323 TI - Epigenetic mechanisms in memory formation. AB - Discoveries concerning the molecular mechanisms of cell differentiation and development have dictated the definition of a new sub-discipline of genetics known as epigenetics. Epigenetics refers to a set of self-perpetuating, post translational modifications of DNA and nuclear proteins that produce lasting alterations in chromatin structure as a direct consequence, and lasting alterations in patterns of gene expression as an indirect consequence. The area of epigenetics is a burgeoning subfield of genetics in which there is considerable enthusiasm driving new discoveries. Neurobiologists have only recently begun to investigate the possible roles of epigenetic mechanisms in behaviour, physiology and neuropathology. Strikingly, the relevant data from the few extant neurobiology-related studies have already indicated a theme - epigenetic mechanisms probably have an important role in synaptic plasticity and memory formation. PMID- 15654324 TI - Working memory in primate sensory systems. AB - Sensory working memory consists of the short-term storage of sensory stimuli to guide behaviour. There is increasing evidence that elemental sensory dimensions - such as object motion in the visual system or the frequency of a sound in the auditory system - are stored by segregated feature-selective systems that include not only the prefrontal and parietal cortex, but also areas of sensory cortex that carry out relatively early stages of processing. These circuits seem to have a dual function: precise sensory encoding and short-term storage of this information. New results provide insights into how activity in these circuits represents the remembered sensory stimuli. PMID- 15654325 TI - Absence of the lipid phosphatase SHIP2 confers resistance to dietary obesity. AB - Genetic ablation of Inppl1, which encodes SHIP2 (SH2-domain containing inositol 5 phosphatase 2), was previously reported to induce severe insulin sensitivity, leading to early postnatal death. In the previous study, the targeting construct left the first eighteen exons encoding Inppl1 intact, generating a Inppl1(EX19-28 /-) mouse, and apparently also deleted a second gene, Phox2a. We report a new SHIP2 knockout (Inppl1(-/-)) targeted to the translation-initiating ATG, which is null for Inppl1 mRNA and protein. Inppl1(-/-) mice are viable, have normal glucose and insulin levels, and normal insulin and glucose tolerances. The Inppl1(-/-) mice are, however, highly resistant to weight gain when placed on a high-fat diet. These results suggest that inhibition of SHIP2 would be useful in the effort to ameliorate diet-induced obesity, but call into question a dominant role of SHIP2 in modulating glucose homeostasis. PMID- 15654326 TI - Toll-like receptor engagement converts T-cell autoreactivity into overt autoimmune disease. AB - Autoimmune diabetes mellitus in humans is characterized by immunological destruction of pancreatic beta islet cells. We investigated the circumstances under which CD8(+) T cells specific for pancreatic beta-islet antigens induce disease in mice expressing lymphocytic choriomeningitis virus (LCMV) glycoprotein (GP) as a transgene under the control of the rat insulin promoter. In contrast to infection with LCMV, immunization with LCMV-GP derived peptide did not induce autoimmune diabetes despite large numbers of autoreactive cytotoxic T cells. Only subsequent treatment with Toll-like receptor ligands elicited overt autoimmune disease. This difference was critically regulated by the peripheral target organ itself, which upregulated class I major histocompatibility complex (MHC) in response to systemic Toll-like receptor-triggered interferon-alpha production. These data identify the 'inflammatory status' of the target organ as a separate and limiting factor determining the development of autoimmune disease. PMID- 15654327 TI - Accelerated re-epithelialization in beta3-integrin-deficient- mice is associated with enhanced TGF-beta1 signaling. AB - The upregulation of TGF-beta1 and integrin expression during wound healing has implicated these molecules in this process, but their precise regulation and roles remain unclear. Here we report that, notably, mice lacking beta(3) integrins show enhanced wound healing with re-epithelialization complete several days earlier than in wild-type mice. We show that this effect is the result of an increase in TGF-beta1 and enhanced dermal fibroblast infiltration into wounds of beta(3)-null mice. Specifically, beta(3)-integrin deficiency is associated with elevated TGF-beta receptor I and receptor II expression, reduced Smad3 levels, sustained Smad2 and Smad4 nuclear localization and enhanced TGF-beta1-mediated dermal fibroblast migration. These data indicate that alpha(v)beta(3)-integrin can suppress TGF-beta1-mediated signaling, thereby controlling the rate of wound healing, and highlight a new mechanism for TGF-beta1 regulation by beta(3) integrins. PMID- 15654328 TI - Structural basis for substrate binding, cleavage and allostery in the tRNA maturase RNase Z. AB - Transfer RNAs (tRNAs) are synthesized as part of longer primary transcripts that require processing of both their 3' and 5' extremities in every living organism known. The 5' side is processed (matured) by the ubiquitously conserved endonucleolytic ribozyme, RNase P, whereas removal of the 3' tails can be either exonucleolytic or endonucleolytic. The endonucleolytic pathway is catalysed by an enzyme known as RNase Z, or 3' tRNase. RNase Z cleaves precursor tRNAs immediately after the discriminator base (the unpaired nucleotide 3' to the last base pair of the acceptor stem, used as an identity determinant by many aminoacyl tRNA synthetases) in most cases, yielding a tRNA primed for addition of the CCA motif by nucleotidyl transferase. Here we report the crystal structure of Bacillus subtilis RNase Z at 2.1 A resolution, and propose a mechanism for tRNA recognition and cleavage. The structure explains the allosteric properties of the enzyme, and also sheds light on the mechanisms of inhibition by the CCA motif and long 5' extensions. Finally, it highlights the extraordinary adaptability of the metallo-hydrolase domain of the beta-lactamase family for the hydrolysis of covalent bonds. PMID- 15654329 TI - Functional annotation and network reconstruction through cross-platform integration of microarray data. AB - The rapid accumulation of microarray data translates into a need for methods to effectively integrate data generated with different platforms. Here we introduce an approach, 2(nd)-order expression analysis, that addresses this challenge by first extracting expression patterns as meta-information from each data set (1(st)-order expression analysis) and then analyzing them across multiple data sets. Using yeast as a model system, we demonstrate two distinct advantages of our approach: we can identify genes of the same function yet without coexpression patterns and we can elucidate the cooperativities between transcription factors for regulatory network reconstruction by overcoming a key obstacle, namely the quantification of activities of transcription factors. Experiments reported in the literature and performed in our lab support a significant number of our predictions. PMID- 15654330 TI - Myosin-XVa is required for tip localization of whirlin and differential elongation of hair-cell stereocilia. AB - Stereocilia are microvilli-derived mechanosensory organelles that are arranged in rows of graded heights on the apical surface of inner-ear hair cells. The 'staircase'-like architecture of stereocilia bundles is necessary to detect sound and head movement, and is achieved through differential elongation of the actin core of each stereocilium to a predetermined length. Abnormally short stereocilia bundles that have a diminished staircase are characteristic of the shaker 2 (Myo15a(sh2)) and whirler (Whrn(wi)) strains of deaf mice. We show that myosin XVa is a motor protein that, in vivo, interacts with the third PDZ domain of whirlin through its carboxy-terminal PDZ-ligand. Myosin-XVa then delivers whirlin to the tips of stereocilia. Moreover, if green fluorescent protein (GFP)-Myo15a is transfected into hair cells of Myo15a(sh2) mice, the wild-type pattern of hair bundles is restored by recruitment of endogenous whirlin to the tips of stereocilia. The interaction of myosin-XVa and whirlin is therefore a key event in hair-bundle morphogenesis. PMID- 15654332 TI - Basic mechanism of three-dimensional collagen fibre transport by fibroblasts. AB - Collagen remodelling by fibroblasts has a crucial role in organizing tissue structures that are essential to motility during wound repair, development and regulation of cell growth. However, the mechanism of collagen fibre movement in three-dimensional (3D) matrices is not understood. Here, we show that fibroblast lamellipodia extend along held collagen fibres, bind, and retract them in a 'hand over-hand' cycle, involving alpha2beta1 integrin. Wild-type fibroblasts move collagen fibres three to four times farther per cycle than fibroblasts lacking myosin II-B (myosin II-B(-/-)). Similarly, myosin II-B(-/-) fibroblasts contract 3D collagen gels threefold less than controls. On two-dimensional (2D) substrates, however, rates of collagen bead and cell movement are not affected by loss of myosin II-B. Green fluorescent protein (GFP)-tagged myosin II-B, but not II-A, restores normal function in knockout cells and localizes to cell processes, whereas myosin II-A is more centrally located. Additionally, GFP-myosin II-B moves out to the periphery and back during hand-over-hand fibre movement, whereas on 2D collagen, myosin II-B is more centrally distributed. Thus, we suggest that cyclic myosin II-B assembly and contraction in lamellipodia power 3D fibre movements. PMID- 15654331 TI - FoxM1 is required for execution of the mitotic programme and chromosome stability. AB - Transcriptional induction of cell-cycle regulatory proteins ensures proper timing of subsequent cell-cycle events. Here we show that the Forkhead transcription factor FoxM1 regulates expression of many G2-specific genes and is essential for chromosome stability. Loss of FoxM1 leads to pleiotropic cell-cycle defects, including a delay in G2, chromosome mis-segregation and frequent failure of cytokinesis. We show that transcriptional activation of cyclin B by FoxM1 is essential for timely mitotic entry, whereas CENP-F, another direct target of FoxM1 identified here, is essential for precise functioning of the mitotic spindle checkpoint. Thus, our data uncover a transcriptional cluster regulated by FoxM1 that is essential for proper mitotic progression. PMID- 15654333 TI - Negative cell-cycle regulators cooperatively control self-renewal and differentiation of haematopoietic stem cells. AB - Haematopoietic stem cells (HSCs) are capable of shifting from a state of relative quiescence under homeostatic conditions to rapid proliferation under conditions of stress. The mechanisms that regulate the relative quiescence of stem cells and its association with self-renewal are unclear, as is the contribution of molecular regulators of the cell cycle to these decisions. Understanding the mechanisms that govern these transitions will provide important insights into cell-cycle regulation of HSCs and possible therapeutic approaches to expand HSCs. We have investigated the role of two negative regulators of the cell cycle, p27(Kip1) and MAD1, in controlling this transition. Here we show that Mad1(-/ )p27(Kip1-/-) bone marrow has a 5.7-fold increase in the frequency of stem cells, and surprisingly, an expanded pool of quiescent HSCs. However, Mad1(-/-)p27(Kip1 /-) stem cells exhibit an enhanced proliferative response under conditions of stress, such as cytokine stimulation in vitro and regeneration of the haematopoietic system after ablation in vivo. Together these data demonstrate that the MYC-antagonist MAD1 and cyclin-dependent kinase inhibitor p27(Kip1) cooperate to regulate the self-renewal and differentiation of HSCs in a context dependent manner. PMID- 15654334 TI - Low LDL cholesterol in individuals of African descent resulting from frequent nonsense mutations in PCSK9. AB - The low-density lipoprotein receptor (LDLR) prevents hypercholesterolemia and atherosclerosis by removing low-density lipoprotein (LDL) from circulation. Mutations in the genes encoding either LDLR or its ligand (APOB) cause severe hypercholesterolemia. Missense mutations in PCSK9, encoding a serine protease in the secretory pathway, also cause hypercholesterolemia. These mutations are probably gain-of-function mutations, as overexpression of PCSK9 in the liver of mice produces hypercholesterolemia by reducing LDLR number. To test whether loss of-function mutations in PCSK9 have the opposite effect, we sequenced the coding region of PCSK9 in 128 subjects (50% African American) with low plasma levels of LDL and found two nonsense mutations (Y142X and C679X). These mutations were common in African Americans (combined frequency, 2%) but rare in European Americans (<0.1%) and were associated with a 40% reduction in plasma levels of LDL cholesterol. These data indicate that common sequence variations have large effects on plasma cholesterol levels in selected populations. PMID- 15654335 TI - A common inversion under selection in Europeans. AB - A refined physical map of chromosome 17q21.31 uncovered a 900-kb inversion polymorphism. Chromosomes with the inverted segment in different orientations represent two distinct lineages, H1 and H2, that have diverged for as much as 3 million years and show no evidence of having recombined. The H2 lineage is rare in Africans, almost absent in East Asians but found at a frequency of 20% in Europeans, in whom the haplotype structure is indicative of a history of positive selection. Here we show that the H2 lineage is undergoing positive selection in the Icelandic population, such that carrier females have more children and have higher recombination rates than noncarriers. PMID- 15654336 TI - Mutations in the gene encoding fibroblast growth factor 10 are associated with aplasia of lacrimal and salivary glands. AB - Autosomal dominant aplasia of lacrimal and salivary glands (ALSG; OMIM 180920 and OMIM 103420) is a rare condition characterized by irritable eyes and dryness of the mouth. We mapped ALSG to 5p13.2-5q13.1, which coincides with the gene fibroblast growth factor 10 (FGF10). In two extended pedigrees, we identified heterozygous mutations in FGF10 in all individuals with ALSG. Fgf10(+/-) mice have a phenotype similar to ALSG, providing a model for this disorder. We suggest that haploinsufficiency for FGF10 during a crucial stage of development results in ALSG. PMID- 15654337 TI - Polyglutamine expansion of huntingtin impairs its nuclear export. AB - Proteins with polyglutamine (polyQ) expansions accumulate in the nucleus and affect gene expression. The mechanism by which mutant huntingtin (htt) accumulates intranuclearly is not known; wild-type htt, a 350-kDa protein of unknown function, is normally found in the cytoplasm. N-terminal fragments of mutant htt, which contain a polyQ expansion (>37 glutamines), have no conserved nuclear localization sequences or nuclear export sequences but can accumulate in the nucleus and cause neurological problems in transgenic mice. Here we report that N-terminal htt shuttles between the cytoplasm and nucleus in a Ran GTPase independent manner. Small N-terminal htt fragments interact with the nuclear pore protein translocated promoter region (Tpr), which is involved in nuclear export. PolyQ expansion and aggregation decrease this interaction and increase the nuclear accumulation of htt. Reducing the expression of Tpr by RNA interference or deletion of ten amino acids of N-terminal htt, which are essential for the interaction of htt with Tpr, increased the nuclear accumulation of htt. These results suggest that Tpr has a role in the nuclear export of N-terminal htt and that polyQ expansion reduces this nuclear export to cause the nuclear accumulation of htt. PMID- 15654338 TI - Mutations in MRAP, encoding a new interacting partner of the ACTH receptor, cause familial glucocorticoid deficiency type 2. AB - Familial glucocorticoid deficiency (FGD), or hereditary unresponsiveness to adrenocorticotropin (ACTH; OMIM 202200), is an autosomal recessive disorder resulting from resistance to the action of ACTH on the adrenal cortex, which stimulates glucocorticoid production. Affected individuals are deficient in cortisol and, if untreated, are likely to succumb to hypoglycemia or overwhelming infection in infancy or childhood. Mutations of the ACTH receptor (melanocortin 2 receptor, MC2R) account for approximately 25% of cases of FGD. FGD without mutations of MC2R is called FGD type 2. Using SNP array genotyping, we mapped a locus involved in FGD type 2 to chromosome 21q22.1. We identified mutations in a gene encoding a 19-kDa single-transmembrane domain protein, now known as melanocortin 2 receptor accessory protein (MRAP). We show that MRAP interacts with MC2R and may have a role in the trafficking of MC2R from the endoplasmic reticulum to the cell surface. PMID- 15654339 TI - The p53QS transactivation-deficient mutant shows stress-specific apoptotic activity and induces embryonic lethality. AB - The role of transcriptional activation in p53 function is highly controversial. To define this role in vivo, we generated a Trp53 knock-in construct encoding a protein carrying mutations of two residues that are crucial for transactivation (L25Q,W26S). Here we show that these mutations have selective effects on the biological functions of p53. Although its ability to activate various p53 target genes is largely compromised, the p53(QS) protein retains the ability to transactivate the gene Bax. The ability of the p53(QS) mutant protein to elicit a DNA damage-induced G1 cell cycle-arrest response is also partially impaired. p53(QS) has selective defects in its ability to induce apoptosis: it is completely unable to activate apoptosis in response to DNA damage, is partially unable to do so when subjected to serum deprivation and retains substantial apoptotic activity upon exposure to hypoxia. These findings suggest that p53 acts through distinct, stimulus-specific pathways to induce apoptosis. The importance of the biological activity of p53(QS) in vivo is underscored by our finding that expression of p53(QS), which cannot bind mdm2, induces embryonic lethality. Taken together, these results suggest that p53 has different mechanisms of action depending on specific contextual cues, which may help to clarify the function of p53 in preventing cancer. PMID- 15654340 TI - A leucine zipper in the N terminus confers membrane association to SLP-65. AB - Membrane recruitment of adaptor proteins is crucial for coupling antigen receptors to downstream signaling events. Despite the essential function of the B cell adaptor SLP-65, the mechanism of its recruitment to the plasma membrane is not yet understood. Here we show that a highly conserved leucine zipper in the SLP-65 N terminus is responsible for membrane association. Alterations in the N terminus abolished SLP-65 membrane localization and activity, both of which were restored by replacement of the N terminus with a myristoylation signal. The N terminus is an autonomous domain that confers specific localization and function when transferred to green fluorescent protein or the adaptor protein SLP-76. Our data elucidate the mechanism of SLP-65 membrane recruitment and suggest that leucine zipper motifs are essential interaction domains of signaling proteins. PMID- 15654341 TI - Inflammatory mediators are insufficient for full dendritic cell activation and promote expansion of CD4+ T cell populations lacking helper function. AB - Dendritic cells (DCs) can be activated directly by triggering of receptors for pathogens or, indirectly, by exposure to inflammatory signals. It remains unclear, however, whether the two pathways result in qualitatively similar DCs or lead to equivalent adaptive immune responses. Here we report that indirect activation by inflammatory mediators generated DCs that supported CD4(+) T cell clonal expansion but failed to direct T helper cell differentiation. In contrast, exposure to pathogen components resulted in fully activated DCs that promoted T helper responses. These results indicate that inflammation cannot substitute for contact with pathogen components in DC activation and suggest that the function of pattern recognition by DCs is to couple the quality of the adaptive immune response to the nature of the pathogen. PMID- 15654342 TI - Calcium oscillations regulate thymocyte motility during positive selection in the three-dimensional thymic environment. AB - The three-dimensional thymic microenvironment and calcium signaling pathways are essential for driving positive selection of developing T cells. However, the nature of calcium signals and the diversity of their effects in the thymus are unknown. We describe here a thymic slice preparation for visualizing thymocyte motility and signaling in real time with two-photon microscopy. Naive thymocytes were highly motile at low intracellular calcium concentrations, but during positive selection cells became immobile and showed sustained calcium concentration oscillations. Increased intracellular calcium was necessary and sufficient to arrest thymocyte motility. The calcium dependence of motility acts to prolong thymocyte interactions with antigen-bearing stromal cells, promoting sustained signaling that may enhance the expression of genes underlying positive selection. PMID- 15654343 TI - Donor impurity band exchange in dilute ferromagnetic oxides. AB - Dilute ferromagnetic oxides having Curie temperatures far in excess of 300 K and exceptionally large ordered moments per transition-metal cation challenge our understanding of magnetism in solids. These materials are high-k dielectrics with degenerate or thermally activated n-type semiconductivity. Conventional super exchange or double-exchange interactions cannot produce long-range magnetic order at concentrations of magnetic cations of a few percent. We propose that ferromagnetic exchange here, and in dilute ferromagnetic nitrides, is mediated by shallow donor electrons that form bound magnetic polarons, which overlap to create a spin-split impurity band. The Curie temperature in the mean-field approximation varies as (xdelta)(1/2) where x and delta are the concentrations of magnetic cations and donors, respectively. High Curie temperatures arise only when empty minority-spin or majority-spin d states lie at the Fermi level in the impurity band. The magnetic phase diagram includes regions of semiconducting and metallic ferromagnetism, cluster paramagnetism, spin glass and canted antiferromagnetism. PMID- 15654344 TI - Molecularly inherent voltage-controlled conductance switching. AB - Molecular electronics has been proposed as a pathway for high-density nanoelectronic devices. This pathway involves the development of a molecular memory device based on reversible switching of a molecule between two conducting states in response to a trigger, such as an applied voltage. Here we demonstrate that voltage-triggered switching is indeed a molecular phenomenon by carrying out studies on the same molecule using three different experimental configurations scanning tunnelling microscopy, crossed-wire junction, and magnetic-bead junction. We also demonstrate that voltage-triggered switching is distinctly different from stochastic switching, essentially a transient (time-dependent) phenomenon that is independent of the applied voltage. PMID- 15654345 TI - Self-assembled microdevices driven by muscle. AB - Current procedures for manual extraction of mature muscle tissue in micromechanical structures are time consuming and can damage the living components. To overcome these limitations, we have devised a new system for assembling muscle-powered microdevices based on judicious manipulations of materials phases and interfaces. In this system, individual cells grow and self assemble into muscle bundles that are integrated with micromechanical structures and can be controllably released to enable free movement. Having realized such an assembly with cardiomyocytes we demonstrate two potential applications: a force transducer able to characterize in situ the mechanical properties of muscle and a self-assembled hybrid (biotic/abiotic) microdevice that moves as a consequence of collective cooperative contraction of muscle bundles. Because the fabrication of silicon microdevices is independent of the subsequent assembly of muscle cells, this system is highly versatile and may lead to the integration of cells and tissues with a variety of other microstructures. PMID- 15654346 TI - Atom inlays performed at room temperature using atomic force microscopy. AB - The ability to manipulate single atoms and molecules laterally for creating artificial structures on surfaces is driving us closer to the ultimate limit of two-dimensional nanoengineering. However, experiments involving this level of manipulation have been performed only at cryogenic temperatures. Scanning tunnelling microscopy has proved, so far, to be a unique tool with all the necessary capabilities for laterally pushing, pulling or sliding single atoms and molecules, and arranging them on a surface at will. Here we demonstrate, for the first time, that it is possible to perform well-controlled lateral manipulations of single atoms using near-contact atomic force microscopy even at room temperature. We report the creation of 'atom inlays', that is, artificial atomic patterns formed from a few embedded atoms in the plane of a surface. At room temperature, such atomic structures remain stable on the surface for relatively long periods of time. PMID- 15654347 TI - Voriconazole therapeutic drug monitoring in allogeneic hematopoietic stem cell transplant recipients. AB - Voriconazole, a new antifungal agent, is increasingly being used after HSCT. The hepatic cytochrome P450 isoenzyme 2C19 plays a significant role in voriconazole metabolism. As CYP2C19 exhibits significant genetic polymorphism, some patients metabolize voriconazole poorly resulting in increased plasma drug levels. The clinical significance of this is unknown, and the utility of monitoring voriconazole levels is unclear. Steady-state trough plasma voriconazole levels were obtained in 25 allogeneic HSCT recipients using an HPLC assay. Patients had drug levels checked once (n=13), twice (n=10), or > or =3 times (n=2) 5-18 days (median 10) after starting voriconazole or dose modification. The 41 voriconazole levels were 0.2-6.8 microg/ml (median 1.6); 6 (15%) were <0.5 (possibly below the in vitro MIC90 for Aspergillus spp.). Voriconazole concentrations correlated with aspartate aminotranferase (AST) (r=0.5; P=0.0009) and alkaline phosphatase (r=0.34; P=0.03), but not with creatinine, bilirubin and alanine aminotransferase (ALT). Since liver dysfunction is common after HSCT, it was not possible to determine if elevated AST and alkaline phosphatase levels were the cause or the consequence of higher voriconazole levels. We conclude that trough voriconazole levels vary considerably between patients, and suggest monitoring levels in patients receiving voriconazole for confirmed fungal infections, and in those with elevated AST or alkaline phosphatase levels. PMID- 15654348 TI - Genotyping of the noncoding control region of BK virus in patients with haemorrhagic cystitis after allogeneic haematopoietic stem cell transplantation. PMID- 15654349 TI - Isolation and transplantation of highly purified autologous peripheral CD34+ progenitor cells: purging efficacy, hematopoietic reconstitution in non-Hodgkin's lymphoma (NHL): results of Japanese phase II study. AB - The purging efficacy of positive selection of autologous CD34+ PBSC with a clinical scale method of magnetic-activated cell sorting system (CliniMACS) was investigated in 48 patients with non-Hodgkin's lymphoma (NHL). The median purity and recovery rate of the CD34+ cells post-selection were 93.3% (range 32.6-99.3) and 72.2% (range 20.5-309.8), respectively. The real-time PCR method to detect the patient-specific monoclonal immunoglobulin heavy chain gene rearrangement (minimal residual tumor; MRT) and CD19 and CD20 positivities were used for the detection of contaminating NHL cells before and after CD34+ selection. After selection, the median (range) depletion rate of MRT was 2.53 (1.52-4.78) log, and that of CD19+ cell and CD20+ cell was 2.46 (0.74-3.64) log and 2.32 (0.40-4.01) log, respectively. In 41 patients, high-dose chemotherapy was performed, followed by the transplantation of the isolated CD34+ cells. Rapid neutrophil recovery as well as platelet recovery was seen with a median time to reach 0.5 x 10(9)/l neutrophils of 10 days (range 8-13) and 20 x 10(9)/l platelets of 14 days (range 10-34), respectively. The present study demonstrated that CliniMACS is a highly effective positive selection method and a high purging efficacy could be obtained without compromising the hematopoietic reconstitution capacity of the graft in NHL patients undergoing high-dose chemotherapy. PMID- 15654351 TI - Use of natural killer cells in hematopoetic stem cell transplantation. AB - Adoptive immunotherapy using natural killer (NK) cells may prove useful, especially in situations where infusion of T cells is impractical such as in recipients of haploidentical stem cell transplantation (HSCT) from haploidentical donors. NK cells may induce potent antileukemic and possibly antirejection activity and may even mitigate graft versus host disease (GvHD). Whether such effects are clinically important and whether they are mediated mainly or exclusively by KIR-HLA class I interactions remains to be determined. Recent advances in graft engineering provide for methods to isolate large numbers of purified NK cells. Several groups have shown that clinical grade NK cells up to a dose of 10(7)/kg may be collected and purified for the purpose of infusion to patients. Early results, in a limited number of patients, show that these cell doses may be administered without adverse events and without inducing GvHD. Whether such infusions will be useful in preventing graft rejection, or exerting graft versus leukemia effects and hastening immune recovery requires further study. PMID- 15654350 TI - Impact of ABO incompatibility on outcome after allogeneic peripheral blood stem cell transplantation. AB - Few studies have addressed the incidence of graft-versus-host disease (GVHD) or survival after ABO-incompatible allogeneic peripheral blood stem cell transplantation (PBSCT). We analyzed the clinical outcome of ABO incompatibility after allogeneic PBSCT. A total of 89 consecutive adult patients with hematological diseases including 49 ABO-identical, 20 major, 15 minor, and five bidirectional ABO-incompatible transplants were enrolled from four medical centers in Korea. No significant difference in engraftment times, graft failure, or transfusion requirements between groups was noted. A clinical diagnosis of severe immune hemolysis or pure red cell aplasia was not made for any patient after transplantation. The incidence of acute or chronic GVHD did not statistically differ between groups. With a median follow-up duration of 13 months (range, 0.5-61 months), the 3-year overall survival estimates for the ABO identical, major/bidirectional, and minor group were 44.6.0+/-9.0, 43.1+/-11.6, and 43.8+/-13.5%, respectively (P=0.8652), while the 3-year disease-free survival estimates were 33.8+/-7.6, 39.9+/-11.4, and 45.7+/-13.1%, respectively (P=0.8546). We observed that time to neutrophil, platelet, and red blood cell engraftment, transfusion requirements, incidence of acute or chronic GVHD, relapse, and survival were not influenced by ABO incompatibility after allogeneic PBSCT from HLA-matched sibling donors. PMID- 15654352 TI - Acute hepatic failure as onset of progressive sclerodermatous chronic graft versus-host disease after donor lymphocyte infusion. PMID- 15654354 TI - Stem cell transplantation for patients with Fanconi anemia with low-dose cyclophosphamide and antithymocyte globulins without the use of radiation therapy. AB - In all, 22 patients with confirmed Fanconi anemia (FA) underwent stem cell transplantation (SCT) from HLA-matched, related donors at KFSHRC. Median age at SCT was 7.6 years (range, 2.5-14.6 years). Conditioning regimen consisted of cyclophosphamide (CY) 15 mg/kg/day intravenously (i.v.) for 4 consecutive days, in addition to equine antithymocyte globulins (ATG) given i.v. at 40 mg/kg/day for four doses pre-SCT. No radiation therapy was given. For graft-versus-host disease prophylaxis, we used cyclosporin at the standard doses; ATG was added at 20 mg/kg/dose i.v. on days 2, 4, 6, 8, 10, and 12 post-SCT (total of six doses). All patients engrafted and are alive and transfusion independent with a median follow-up time of 20.2 months (range, 3.3-59 months). One patient however developed a decrease in her WBC and platelet count. Her work-up revealed slightly hypocellular bone marrow, and a series of chimerism studies over 1 year confirmed that she has stable mixed chimerism; she remains transfusion independent. We conclude that low-dose CY without radiation therapy can be used satisfactorily in the conditioning of patients with FA undergoing related SCT. PMID- 15654353 TI - ESHAP plus G-CSF as an effective peripheral blood progenitor cell mobilization regimen in pretreated non-Hodgkin's lymphoma: comparison with high-dose cyclophosphamide plus G-CSF. AB - The ESHAP (etoposide, methylprednisolone, high-dose cytarabine, and cisplatin) regimen has been shown to be effective as an active salvage therapy for lymphoma. Mobilizing stem cells following ESHAP should decrease time to transplantation by making separate mobilizing chemotherapy (MC) unnecessary, while controlling a patient's lymphoma. We therefore assessed the mobilization potential of ESHAP plus G-CSF in 26 patients (ESHAP group) with non-Hodgkin's lymphoma (NHL) and compared these results with those of 24 patients with NHL who received high-dose (4 g/m2l) cyclophosphamide (HDCY) as MC (HDCY group). The age, sex, and radiotherapy to the axial skeleton were well matched between groups, but the number of patients with poor mobilization predictors was higher in the ESHAP group. Significantly higher numbers of CD34+ cells (x 10(6)/kg) (17.1+/-18.8 vs 5.8+/-5.0, P=0.03) and apheresis day 1 CD34+ cells (x 10(6)/kg) (5.5+/-6.6 vs 1.7+/-2.0, P=0.014) were collected from the ESHAP group than from the HDCY group, and the number of patients who achieved an optimal CD34+ cell target of 5 x 10(6)/kg was higher in the ESHAP group (81 vs 50%, P=0.022). Log-rank test revealed that time to target peripheral blood progenitor cell collection (> or =5 x 10(6)/kg) was shorter in the ESHAP group (P=0.007). These results indicate that ESHAP plus G-CSF is an excellent mobilization regimen in patients with relapsed and poor-risk aggressive NHL. PMID- 15654355 TI - Pulmonary function testing prior to hematopoietic stem cell transplantation. AB - The pretransplant pulmonary function test plays an important role in the management of noninfectious pulmonary complications after hematopoietic stem cell transplantation (HCT). Although these tests are widely used as standard preoperative assessments in the nontransplant population, common conditions associated with the HCT patient requires that particular attention be given to interpretation of pulmonary function testing (PFT) results, such as comparison of serial pulmonary function tests and evaluation of the diffusion capacity. Although their utility in helping to predict the likelihood of developing post transplant pulmonary complications and mortality is not well established, current data indicate that pretransplant PFTs are important as a reference for the interpretation of post transplant PFTs and for identifying patients at high risk for developing pulmonary complications and/or mortality after HCT. Future studies of pretransplant pulmonary function should consider the advances in HCT, so that pretransplant PFTs will become a useful tool in pretransplant risk assessment and help the transplant oncologist to determine the most appropriate conditioning regimen for a patient with compromised lung function. PMID- 15654356 TI - Allogeneic bone marrow transplantation for juvenile myelomonocytic leukemia: a single center experience of 23 patients. AB - Juvenile myelomonocytic leukemia (JMML) is a childhood leukemia for which allogeneic BMT is the only curative therapy. At our pediatric stem cell transplantation unit, we performed 26 BMTs in 23 children (age 0.5-12.7 years). Conditioning was CY/TBI based (1980-1996, n=14) or BU/CY/melphalan based (1996 2001, n=9). Donors were HLA-identical siblings (n=11), unrelated volunteers (n=9) or mismatched family members (n=3). A total of 10 patients survive in CR (median follow-up 6.8 years, range 3.1-22.2 years). Relapse or persistent disease was observed in eight and two patients, respectively. Nine of these patients died, one achieved a second remission following acute nonlymphatic leukemia chemotherapy (duration to date 5.3 years). Transplant-related mortality occurred in four patients. Overall survival at 5 and 10 years was 43.5%. Using T-cell depleted, one-antigen mismatched unrelated donors was the only significant adverse factor associated with relapse in multivariate analysis (P=0.039, hazard ratio 4.9). Together with a trend towards less relapse in patients with graft versus-host-disease and in patients transplanted with matched unrelated donors, this suggests a graft-versus-leukemia effect of allogeneic BMT in JMML. PMID- 15654357 TI - Proteomic characterization of harvested pseudopodia with differential gel electrophoresis and specific antibodies. AB - Malignant gliomas (astrocytomas) are lethal tumors that invade the brain. Invasive cell migration is initiated by extension of pseudopodia into interstitial spaces. In this study, U87 glioma cells formed pseudopodia in vitro as cells pushed through 3 microm pores of polycarbonate membranes. Harvesting pseudopodia in a novel two-step method provided material for proteomic analysis. Differences in the protein profiles of pseudopodia and whole cells were found using differential gel electrophoresis (DIGE) and immunoblotting. Proteins from two-dimensional (2D) gels with M(R)'s of 20-100 kDa and pI's of 3.0-10.0 were identified by peptide mass fingerprinting analysis using mass spectrometry. For DIGE, lysates of pseudopodia and whole cells were each labeled with electrophilic forms of fluorescent dyes, Cy3 or Cy5, and analyzed as mixtures. Analysis was repeated with reciprocal labeling. Differences in protein distributions were detected by manual inspection and computer analysis. Topographical digital maps of the scanned gels were used for algorithmic spot matching, normalization of background, quantifying spot differences, and elimination of artifacts. Pseudopodial proteins in Coomassie-stained 2D gels included isoforms of glycolytic enzymes as the largest group, seven of 24 proteins. Peptide mass fingerprint analysis of DIGE gels demonstrated increased isoforms of annexin (Anx) I, AnxII, enolase, pyruvate kinase, and aldolase, and decreased mitochondrial manganese superoxide dismutase and transketolase in pseudopodia. Specific antibodies showed restricted immunoreactivity of the hepatocyte growth factor (HGF) alpha chain to pseudopodia, indicating localization of its active form. Met (the HGF receptor), actin, and total AnxI were increased in pseudopodial lysates on immunoblots. Increased constituents of the pseudopodial proteome in glioma cells, identified in this study as actin, HGF, Met, and isoforms of AnxI, AnxII, and several glycolytic enzymes, represent therapeutic targets to consider for suppression of tumor invasion. PMID- 15654358 TI - Dendritic cells and macrophages are essential for the retention of lymphocytes in (peri)-insulitis of the nonobese diabetic mouse: a phagocyte depletion study. AB - Dendritic cells (DC) and macrophages (Mphi) are present in high numbers in the pancreas of the non-obese diabetic (NOD) mouse during the diabetogenic process from very early stages onwards. In this study, we used clodronate-loaded liposomes to mediate the temporary systemic depletion of these phagocytic cells and monocytic precursors in order to modulate the pancreatic inflammation. Two intraperitoneal injections given with a 2-day interval to 8-week-old NOD mice depleted monocytes from the circulation and monocytes, DC and Mphi from the spleen within the first days after the injections. Monocytes, DC and Mphi reappeared in the circulation and the spleen within one week and had an unchanged phenotype and antigen presenting function. Interestingly, this treatment caused a delayed disappearance (7-21 days postinjection) of DC and Mphi from the endocrine pancreas at a time when monocytes, DC and Mphi had already repopulated the circulation and the spleen. The depletion of DC and Mphi from the endocrine pancreas was accompanied by a total disappearance of lymphocytes from the pancreas. DC, Mphi and lymphocytes reappeared in the pancreatic inflammatory infiltrates in treated mice from 28 days postdepletion onwards. Importantly, the treatment significantly postponed the onset of diabetes, leading to a strongly decreased incidence by 35 weeks of age. Taken together, our data show an essential role of phagocytic cells, that is, DC and Mphi, in the recruitment of lymphocytes to the pancreatic islets in NOD mice. PMID- 15654360 TI - Distinct role for c-kit receptor tyrosine kinase and SgIGSF adhesion molecule in attachment of mast cells to fibroblasts. AB - Binding of stem cell factor (SCF) to c-kit receptor tyrosine kinase (KIT) transduces signals essential for mast cell development via several pathways including activation of phosphatidylinositol 3-kinase (PI3-K). When cultured mast cells (CMCs) are cocultured with fibroblasts expressing membrane-bound SCF, CMCs with normal KIT adhere to fibroblasts and proliferate, whereas CMCs lacking cell surface expression of KIT do neither. Spermatogenic immunoglobulin superfamily (SgIGSF) was identified as another molecule that participates in mast cell adhesion to fibroblasts. Since the IC-2 mast cell line expressed neither KIT nor SgIGSF, the effect of ectopic expression of KIT or SgIGSF on the adhesion of IC-2 cells was examined. Three forms of KIT with the normal ectodomain were used: wild type (KIT-WT) and two mutant types with a phenylalanine substitution at the tyrosine residue 719 (KIT-Y719F) or 821 (KIT-Y821F). KIT-Y719F does not activate PI3-K, whereas KIT-Y821F does. Firstly, KIT or SgIGSF was expressed singly in IC 2 cells. All three forms of KIT increased the adhesion level of IC-2 cells, whereas SgIGSF did not. Secondly, SgIGSF was coexpressed with one of the three forms of KIT. Coexpression of SgIGSF with KIT-WT or KIT-Y821F increased the adhesion level more markedly than was achieved by KIT-WT or KIT-Y821F alone. The effect was abolished by an antibody that blocks SCF-KIT interaction. In contrast, coexpression of SgIGSF with KIT-Y719F did not increase the adhesion level induced by KIT-Y719F alone. In adhesion of mast cells to fibroblasts, KIT appeared to behave as an adhesion molecule and as an activator of other adhesion molecules through phosphorylating PI3-K. PMID- 15654359 TI - Prolonged beta-catenin stabilization and tcf-dependent transcriptional activation in hyperplastic cutaneous wounds. AB - Mesenchymal cells that accumulate during the proliferative phase of wound healing and that are present in hyperplastic wounds share cytologic similarities with the cells from fibroproliferative lesions in which there is activation of beta catenin-mediated transcription. Re-excision wounds from a previous biopsy and samples from hyperplastic cutaneous wounds were studied along with normal tissues. During normal wound healing, there was an increase in beta-catenin protein level, peaking 4 weeks following the insult and returning towards baseline level by 12 weeks. Hyperplastic wounds exhibited a prolonged duration of elevated beta-catenin, lasting more than 2 years following the initial injury. The level of expression of genes known to be upregulated in the proliferative phase of wound healing (alpha-smooth muscle actin and type three collagen), correlated with beta-catenin protein level. The phosphorylation level of glycogen synthase kinase-3-beta, a kinase important for beta-catenin protein destabilization, correlated with beta-catenin protein level. Beta-catenin was transcriptionally active in these wounds as demonstrated by the expression of the beta-catenin target genes (MMP-7 and FN) and by activation of a tcf-reporter in primary cell cultures. Beta-catenin stabilization increases cell proliferation and motility in fibroblasts in vitro, and likely has a similar function during its transient elevation in the proliferative phase of normal wound healing. In hyperplastic wounds, there is dysregulation of beta-catenin, maintaining the mesenchymal cells in a prolonged proliferative state. As such, beta-catenin likely plays a central role in mesenchymal cells during the healing process, and is an appealing therapeutic target for disorders of wound healing. PMID- 15654361 TI - Stress and weight gain in parents of cancer patients. AB - OBJECTIVE: To investigate the effects of chronic stress on weight changes and related behavioral changes in parents with a child who had just been diagnosed with cancer compared to parents with healthy children. DESIGN: Longitudinal case control study with assessments occurring over a three-month period following the child's diagnosis of cancer. SUBJECTS: In total, 49 parents of healthy children and 49 parents of cancer patients aged 19-58. MEASUREMENTS: Body weight, diet, physical activity, self-reported mood and stress. RESULTS: Parents of cancer patients were more likely to gain weight, and experienced significantly greater weight gain over the 3 months than parents of healthy children. The magnitude of weight gain was related to the degree of psychological distress that the parents experienced. Parents of cancer patients reported lower levels of physical activity and lower caloric intake than parents of healthy children, with the most marked differences between groups occurring in the area of physical activity. CONCLUSION: Findings suggest that a major stressor, such as a child's diagnosis of cancer, is associated with weight gain. Further research is needed to determine how long these weight gains persist and whether other types of stress also produce weight gains. Such studies should focus not only on the effect of stress on eating behavior but also on physical activity. PMID- 15654362 TI - The development of efficient catalysts for palladium-catalyzed coupling reactions of aryl halides. AB - In the mid 1990s, we started a program on the development of more efficient palladium catalysts for the C-C and C-N coupling reactions of aryl halides, especially aryl chlorides. In this contribution, our investigations on new catalyst systems and their application in organic synthesis are summarized. PMID- 15654363 TI - Synthetic self-propelled nanorotors. AB - Self-powered completely synthetic nanorotors have been prepared from barcoded gold-nickel nanorods having the gold end anchored to the surface of a silicon wafer; constant velocity circular movements are observed when hydrogen peroxide fuel is catalytically decomposed to oxygen at the unattached nickel end of the nanorod. PMID- 15654364 TI - Dendritic incorporation of quinacridone: solubility, aggregation, electrochemistry, and solid-state luminescence. AB - The first incorporation of quinacridone, a technologically important organic electroluminescent emitter, into dendrimers increases solubility, decreases aggregation, retards heterogeneous electron transfer, and enhances luminescence in condensed phases (powders and thin films). PMID- 15654365 TI - Selective attachment and release of a chemotherapeutic agent from the interior of a protein cage architecture. AB - The antitumor agent doxorubicin was covalently bound and selectively released in a pH dependent manner from the interior surface of a genetically modified small heat shock protein (Hsp) cage. PMID- 15654366 TI - A cyclometalated palladium-azo complex as a differential chromogenic probe for amino acids in aqueous solution. AB - Solutions of a cyclometalated palladium-azo complex exhibited differential UV-Vis absorption spectra in the presence of alpha-amino acids with different side chain groups. PMID- 15654367 TI - Flip-flop of glycosylphosphatidylinositols (GPI's) across the ER. AB - The transbilayer flip-flop of early intermediates in the glycosylphosphatidylinositol (GPI) biosynthetic pathway has been demonstrated using novel fluorescent GPI probes and a biochemical reconstitution approach. PMID- 15654368 TI - Use of tetrameric cubane aggregates of lithium aryloxides as secondary building units in controlling network assembly. AB - Pre-aggregation of lithium aryloxides into tetrahedral arrangements followed by crystallization with the divergent Lewis base dioxane results in the preparation of three types of coordination polymers: zig-zag chains, (6,3) sheets, and diamondoid lattices. PMID- 15654369 TI - A luminescent linear trinuclear magnesium complex assembled from a phosphorus based tris-hydrazone ligand. AB - A novel linear trinuclear magnesium complex (P(S)[N(CH3)N=CHC6H4-o-O]3)2Mg3 was prepared by the reaction of P(S)[N(CH3))N=CHC6H4-o-OH]3 with MgCl2 x 6H2O in the presence of triethylamine. The trinuclear magnesium complex is fluorescent in solution as well as in the solid-state. PMID- 15654370 TI - Self-assembled helical spherical-nanotubes from an L-glutamic acid based bolaamphiphilic low molecular mass organogelator. AB - An L-glutamic acid based bolaamphiphile was observed to self-assemble into a novel "helical spherical-nanotube" with a molecular monolayered structure within a tubular wall. PMID- 15654371 TI - Helical and polymeric nanostructures assembled from benzene tri- and tetracarboxylic acids associated with terpyridine copper(II) complexes. AB - A ladder-like framework was derived from benzene-1,2,4,5-tetracarboxylic acid with Cu(II) and terpyridine; whereas with benzene-1,3,5-tricarboxylic acid, a helical structure was formed with two CO2- groups bridging opposing (terpyridine)Cu(II) units and the other CO2H connecting the helical cylinders via H-bonding. PMID- 15654372 TI - Spatial organization and patterning of palladium nanoparticles on a self assembled helical ribbon lipid. AB - A cholesterol derivative forms self-assembled helical ribbons in organic solvent, and treatment of this helical ribbon lipid as a template with Pd(Ac)2 provides helically-patterned arrays of palladium nanoparticles followed by reduction. PMID- 15654373 TI - Self-assembled monolayers of optically active Co(III) complexes: a new promoter electrode recognizing the electron transfer site in cytochrome c. AB - A new-class of promoter electrode bearing a molecular recognition ability has been constructed; the chirality and/or orientation of promoter on the Au electrode surface have affected the electron transfer rate of cytochrome c. PMID- 15654374 TI - Efficient relaxivity enhancement in dendritic gadolinium complexes: effective motional coupling in medium molecular weight conjugates. AB - Enhancement of the relaxivities of Gd-based MRI contrast agents is achieved by placing the metal ion at the barycentre of the molecular complex in order to improve motional coupling. PMID- 15654375 TI - A cyclic tetra-nuclear dinitrosyl iron complex [Fe(NO)2(imidazolate)]4: synthesis, structure and stability. AB - A novel cyclic tetra-nuclear dinitrosyl iron complex [Fe(NO)2(Im-H)]4 was isolated and characterized by X-ray crystallography, and in donor solvents this fragments into 17 e- monomeric units that give EPR spectra analogous to the g= 2.03 species seen in mammalian biology. PMID- 15654376 TI - A diiron complex mediates an intramolecular aliphatic hydroxylation by various oxygen donors. AB - In the presence of hydrogen peroxide, m-chloroperbenzoic acid or an iodosyl arene, the tert-butyl group of the ligand H(L-t-Bu) in the complex [Fe2(L-t Bu)(mpdp)]2+ is quantitatively hydroxylated to a butanolate terminally bound to one iron in [Fe2(L-t-Bu - H + O)(mpdp)]2+, and mass spectrometry experiments indicate that the reaction proceeds according to different mechanisms. PMID- 15654377 TI - Reversible 1,4-cycloaddition of singlet oxygen to N-substituted 2-pyridones: 1,4 endoperoxide as a versatile chemical source of singlet oxygen. AB - N-substituted pyridones (1) easily undergo singlet oxygenation to give exclusively the corresponding endoperoxides (2), which decompose to give pyridones again while liberating 1O2 in high yield. PMID- 15654378 TI - Carboxylation of anisole derivatives with CO and O2 catalyzed by Pd(OAc)2 and molybdovanadophosphates. AB - Anisole and its homologues were carboxylated under the influence of CO and O2 catalyzed by Pd(OAc)2 combined with molybdovanadophosphates (HPMoV) under mild conditions to give the corresponding carboxylic acids in fair to good yields; for instance, anisole underwent the carboxylation under a mixed gas of CO (0.5 atm) and O2 (0.5 atm) in the presence of Pd(OAc)2 (5 mol%) and H5PMo10V2O40.nH2O (2 mol%) to form an isomeric mixture of anisic acids in good yield. PMID- 15654380 TI - CD inversion and fluorescence enhancement in organic nanoparticles of (R)-di-2 naphthylprolinol. AB - The positive exciton coupling circular dichroism (ECCD) exhibited by monomer of (R)-di-2-naphthylprolinol in organic solvent is inverted to be negative upon the formation of its nanoparticles. PMID- 15654379 TI - Neutral acetohydroxamic acid coordination to a mononuclear Ni(II) center stabilized by an intramolecular hydrogen-bonding interaction. AB - Treatment of a new chelate ligand having both amide- and phenyl-appended pyridyl moieties with Ni(ClO4).6H2O and acetohydroxamic acid in methanol solution results in the production of a novel pseudo-octahedral Ni(II) complex having a neutral acetohydroxamic acid ligand stabilized by a hydrogen-bonding interaction. PMID- 15654381 TI - Backbone-extended pyrrolidine peptide nucleic acids (bepPNA): design, synthesis and DNA/RNA binding studies. AB - One-carbon extended conformationally constrained pyrrolidine PNA monomer (bepPNA) has been synthesized, incorporated into PNA sequences at predefined positions, and showed selective RNA binding properties. PMID- 15654382 TI - The nature of paramagnetic species in nitrogen doped TiO2 active in visible light photocatalysis. AB - Nitrogen doped TiO2, a novel photocatalyst active in the decomposition of organic pollutants using visible light, contains two different types of paramagnetic centres (neutral NO radicals and NO2(2-) type radical ions respectively) which are likely related to specific properties of the solid. PMID- 15654383 TI - Molybdenum-catalyzed hydrostannations of allenylcarbinols. AB - Allenylcarbinols undergo regioselective hydrostannation in the presence of MoBl3, a catalyst which was developed for the hydrostannation of propargyl alcohols and derivatives; allylstannanes are formed preferentially, which can easily be converted into allyl iodides. PMID- 15654384 TI - Synthesis of anthracene ethers from anthracene methyl ethers via an acid catalyzed exchange reaction. AB - Selective synthesis of anthracene ethers was achieved under mild conditions via an acid-catalyzed ether-ether exchange reaction and its scope and limitations, as well as its potential in synthesizing anthracene-based crown ethers, tested. PMID- 15654385 TI - Solution route to single crystalline SnO platelets with tunable shapes. AB - Square and round single crystalline SnO platelets have been prepared in a solution-based chemical route aided with sonication at room temperature. PMID- 15654386 TI - Coupling of alkynols and a phenyl group to a novel eta5-dihydronaphthalenide ligand on a ruthenium template. AB - We report the highly selective assembly of unprecedented eta5-1-methylene-1,2 dihydronaphthalenide ligands from the stoichiometric coupling of a phenyl group and two equivalents of disubstituted propargylic alcohols; in this reaction, tetraphenylborate acts as a phenylating agent. PMID- 15654387 TI - Fluorescence enhancement of a signaling system in the simultaneous presence of transition and alkali metal ions: a potential AND logic gate. AB - Attachment of a laterally non-symmetric cryptand and a macrocycle at the 9- and 10-positions of anthracene leads to a fluorescent signaling system L1 which gives fluorescence enhancement in the simultaneous presence of alkali and transition metal ions. PMID- 15654388 TI - Identification of hPin1 inhibitors that induce apoptosis in a mammalian Ras transformed cell line. AB - The authors have developed a class of potent inhibitors against the phosphate specific prolyl isomerase hPin1, which induced apoptosis in transformed cell lines. PMID- 15654389 TI - A new approach to construct full-length glycosylphosphatidylinositols of parasitic protozoa and [4-deoxy-Man-III]-GPI analogues. AB - A new [2 + 2 + 2] approach to construct GPI molecules through the efficient synthesis of glucosamine-inositol and tetramannose intermediates led to a total synthesis of a GPI-anchor of Trypanosoma cruzi, and also afforded a key intermediate for the synthesis of valuable [4-deoxy-Man-III]-GPI analogues. PMID- 15654390 TI - The hydrogenation of nitrobenzene to aniline: a new mechanism. AB - The Haber mechanism describing the process of hydrogenating nitrobenzene to aniline is shown to be incorrect and a new mechanism is proposed. PMID- 15654391 TI - PMO[KIT-5]-n: synthesis of highly ordered three-dimensional periodic mesoporous organosilicas with Fm3m symmetry. AB - Divalent surfactant [CH3(CH2)15N(CH3)2(CH2)3N(CH3)3]2+ - 2Br- (C16-3-1) was used as a structure directing agent (SDA) for the synthesis of highly ordered periodic mesoporous organosilicas (PMOs) with cubic Fm3m symmetry from 1,2 bis(triethoxylsilyl)ethane (BTEE) under basic conditions. PMID- 15654393 TI - Protein-assisted synthesis of single-crystal nanowires of bismuth compounds. AB - Lysozyme, a protein, has been found to be a new morphology-directing agent and a simple and mild bio-molecule assisted method has been proposed for the synthesis of single-crystal bismuth sulfide and oxide nanowires. PMID- 15654392 TI - Rapid synthesis of highly luminescent CdTe nanocrystals in the aqueous phase by microwave irradiation with controllable temperature. AB - In this paper, we present a new method for rapid synthesis of high quantum yield CdTe nanocrystals in the aqueous phase by microwave irradiation with controllable temperature. PMID- 15654394 TI - Rational design of azide-bridged bimetallic complexes. Crystal structure and magnetic properties of Fe(III)MFe(III) (M = Ni(II) and Cu(II)) trinuclear species. AB - The first examples of azide-bridged bimetallic trinuclear complexes ([M(cyclam)][FeL(N3)(mu1,5-N3)]2) (H2L = 4,5-dichloro-1,2-bis(pyridine-2 carboxamido) benzene) have been structurally and magnetically characterized. PMID- 15654395 TI - Reactivity of N-alkanoyloxy-2,2,6,6-tetramethylpiperidines (O-acylTEMPOs) towards hydride-transferring or metallic alkylating reagents; unprecedented stability and application to chemoselective transformations. AB - Owing to the unprecedented stability of O-acylTEMPOs towards hydride-transferring and metallic alkylating reagents such as LiAlH4 and RMgX, chemoselective transformation of diacid mixed alkyl/TEMP-1-yl esters, where O-acylTEMPOs remained intact, is achieved with these reagents, giving the corresponding carbinols, respectively. PMID- 15654396 TI - Calix[4]bipyrrole--a big, flexible, yet effective chloride-selective anion receptor. AB - Anion binding studies reveal that, in spite of its big size and flexible structure, calix[4]bipyrrole shows strong anion binding in general and good selectivity towards chloride anion in acetonitrile. PMID- 15654397 TI - Preparation of cyclic alkenylmagnesium reagents via an iodine/magnesium exchange. AB - The reaction of various cyclic alkenyl iodides with i-PrMgCl.LiCl produces the corresponding alkenylmagnesium reagents under mild conditions. After reaction with various electrophiles, like allylic halides, disulfides, aldehydes and acid chlorides, the expected products are obtained in 53-91% yield. Remarkably, the mild conditions of the I/Mg-exchange tolerate the presence of sensitive diene functionalities. PMID- 15654398 TI - Cationic fac-tris(pyrazole) complexes as anion receptors. AB - New receptors fac-[Re(CO)3(pz)3]BAr'4 (pz = 3,5-dimethylpyrazole or 3(5)-tert butylpyrazole, Ar' = 3,5-(CF3)2C6H3), synthesized from [Re(OTf)(CO)5] and the pyrazoles, have been found to show a high affinity for chloride. PMID- 15654399 TI - A dynamic supramolecular system exhibiting substrate selectivity in the catalytic epoxidation of olefins. AB - A dynamic supramolecular system involving hydrogen bonding between a Mn(III) salen catalyst and a Zn(II) porphyrin receptor exhibits selectivity for pyridine appended cis-beta-substituted styrene derivatives over phenyl appended derivatives in a catalytic epoxidation reaction. PMID- 15654400 TI - Patient safety: a global priority. PMID- 15654401 TI - The sector-wide approach: a blessing for public health? PMID- 15654402 TI - The role of religion in tobacco control interventions. PMID- 15654403 TI - Global estimate of the incidence of clinical pneumonia among children under five years of age. AB - OBJECTIVE: Clinical pneumonia (defined as respiratory infections associated with clinical signs of pneumonia, principally pneumonia and bronchiolitis) in children under five years of age is still the leading cause of childhood mortality in the world. In this paper we aim to estimate the worldwide incidence of clinical pneumonia in young children. METHODS: Our estimate for the developing world is based on an analysis of published data on the incidence of clinical pneumonia from community based longitudinal studies. Among more than 2000 studies published since 1961, we identified 46 studies that reported the incidence of clinical pneumonia, and 28 of these met pre-defined quality criteria. FINDINGS: The estimate of the median incidence from those studies was 0.28 episodes per child year (e/cy). The 25-75% interquartile range was 0.21-0.71. We assessed the plausibility of this estimate using estimates of global mortality from acute respiratory infections and reported case fatality rates for all episodes of clinical pneumonia reported in community-based studies or the case-fatality rate reported only for severe cases and estimates of the proportion of severe cases occurring in a defined population or community. CONCLUSION: The overlap between the ranges of the estimates implies that a plausible incidence estimate of clinical pneumonia for developing countries is 0.29 e/cy. This equates to an annual incidence of 150.7 million new cases, 11-20 million (7-13%) of which are severe enough to require hospital admission. In the developed world no comparable data are available. However, large population-based studies report that the incidence of community-acquired pneumonia among children less than five years old is approximately 0.026 e/cy, suggesting that more than 95% of all episodes of clinical pneumonia in young children worldwide occur in developing countries. PMID- 15654404 TI - ICD coding changes and discontinuities in trends in cause-specific mortality in six European countries, 1950-99. AB - OBJECTIVE: To evaluate how often coding changes between and within revisions of the International Classification of Diseases (ICD) complicate the description of long-term trends in cause-specific mortality. METHODS: Data on cause-specific mortality between 1950 and 1999 for men and women aged 60 and older were obtained from Denmark, England and Wales, Finland, the Netherlands, Norway and Sweden. Data were obtained by five-year age groups. We constructed a concordance table using three-digit ICD codes. In addition we evaluated the occurrence of mortality discontinuities by visually inspecting cause-specific trends and country-specific background information. Evaluation was also based on quantification of the discontinuities using a Poisson regression model (including period splines). We compared the observed trends in cause-specific mortality with the trends after adjustment for the discontinuities caused by changes to coding. FINDINGS: In 45 out of 416 (10.8 %) instances of ICD revisions to cause-specific mortality codes, significant discontinuities that were regarded as being due to ICD revisions remained. The revisions from ICD-6 and ICD-7 to ICD-8 and a wide range of causes of death, with the exception of the specific cancers, were especially affected. Incidental changes in coding rules were also important causes of discontinuities in trends in cause-specific mortality, especially in England and Wales, Finland and Sweden. Adjusting for these discontinuities can lead to significant changes in trends, although these primarily affect only limited periods of time. CONCLUSION: Despite using a carefully constructed concordance table based on three-digit ICD codes, mortality discontinuities arising as a result of coding changes (both between and within revisions) can lead to substantial changes in long-term trends in cause-specific mortality. Coding changes should therefore be evaluated by researchers and, where necessary, controlled for. PMID- 15654405 TI - Respiratory syncytial virus infection: denominator-based studies in Indonesia, Mozambique, Nigeria and South Africa. AB - OBJECTIVE: To assess the burden of respiratory syncytial virus (RSV)-associated lower respiratory infections (LRI) in children in four developing countries. METHODS: A WHO protocol for prospective population-based surveillance of acute respiratory infections in children aged less than 5 years was used at sites in Indonesia, Mozambique, Nigeria and South Africa. RSV antigen was identified by enzyme-linked immunosorbent assay performed on nasopharyngeal specimens from children meeting clinical case definitions. FINDINGS: Among children aged < 5 years, the incidence of RSV-associated LRI per 1000 child-years was 34 in Indonesia and 94 in Nigeria. The incidence of RSV-associated severe LRI per 1000 child-years was 5 in Mozambique, 10 in Indonesia, and 9 in South Africa. At all study sites, the majority of RSV cases occurred in infants. CONCLUSION: These studies demonstrate that RSV contributes to a substantial but quite variable burden of LRI in children aged < 5 years in four developing countries. The possible explanations for this variation include social factors, such as family size and patterns of seeking health care; the proportion of children infected by human immunodeficiency syndrome (HIV); and differences in clinical definitions used for obtaining samples. The age distribution of cases indicates the need for an RSV vaccine that can protect children early in life. PMID- 15654406 TI - Religion-based tobacco control interventions: how should WHO proceed? AB - Using religion to improve health is an age-old practice. However, using religion and enlisting religious authorities in public health campaigns, as exemplified by tobacco control interventions and other activities undertaken by WHO's Eastern Mediterranean Regional Office, is a relatively recent phenomenon. Although all possible opportunities within society should be exploited to control tobacco use and promote health, religion-based interventions should not be exempted from the evidence-based scrutiny to which other interventions are subjected before being adopted. In the absence of data and debate on whether this approach works, how it should be applied, and what the potential downsides and alternatives are, international organizations such as WHO should think carefully about using religion-based public health interventions in their regional programmes. PMID- 15654407 TI - The antimicrobial resistance containment and surveillance approach--a public health tool. AB - Antimicrobial drug resistance (AMR) is widely recognized as a global public health threat because it endangers the effectiveness of treatment of infectious diseases. In 2001 WHO issued the Global Strategy for Containment of Antimicrobial Resistance, but it has proved difficult to translate the recommendations of the Global Strategy into effective public health actions. The purpose of the Antimicrobial Resistance Containment and Surveillance (ARCS) approach is to facilitate the formulation of public health programmes and the mobilization of human and financial resources for the containment of AMR. The ARCS approach highlights the fundamental link between rational drug use and containment of AMR. Clinical management of human and animal infections should be improved through better disease control and prevention, high quality diagnostic testing, appropriate treatment regimens and consumer health education. At the same time, systems for supplying antimicrobial drugs should include appropriate regulations, lists of essential drugs, and functional mechanisms for the approval and delivery of drugs. Containment of AMR is defined in the ARCS approach as the continuous application of this package of core interventions. Surveillance of the extent and trends of antimicrobial resistance as well as the supply, selection and use of antimicrobial drugs should be established to monitor the process and outcome of containment of AMR. The ARCS approach is represented in the ARCS diagram (Fig. 2) which provides a simplified, but comprehensive illustration of the complex problem of containment and monitoring of AMR. PMID- 15654408 TI - Do fixed-dose combination pills or unit-of-use packaging improve adherence? A systematic review. AB - Adequate adherence to medication regimens is central to the successful treatment of communicable and noncommunicable disease. Fixed-dose combination pills and unit-of-use packaging are therapy-related interventions that are designed to simplify medication regimens and so potentially improve adherence. We conducted a systematic review of relevant randomized trials in order to quantify the effects of fixed-dose combination pills and unit-of-use packaging, compared with medications as usually presented, in terms of adherence to treatment and improved outcomes. Only 15 trials met the inclusion criteria; fixed-dose combination pills were investigated in three of these, while unit-of-use packaging was studied in 12 trials. The trials involved treatments for communicable diseases (n = 5), blood pressure lowering medications (n = 3), diabetic patients (n = 1), vitamin supplementation (n = 1) and management of multiple medications by the elderly (n = 5). The results of the trials suggested that there were trends towards improved adherence and/or clinical outcomes in all but three of the trials; this reached statistical significance in four out of seven trials reporting a clinically relevant or intermediate end-point, and in seven out of thirteen trials reporting medication adherence. Measures of outcome were, however, heterogeneous, and interpretation was further limited by methodological issues, particularly small sample size, short duration and loss to follow-up. Overall, the evidence suggests that fixed-dose combination pills and unit-of-use packaging are likely to improve adherence in a range of settings, but the limitations of the available evidence means that uncertainty remains about the size of these benefits. PMID- 15654409 TI - Socioeconomic status and obesity in adult populations of developing countries: a review. AB - A landmark review of studies published prior to 1989 on socioeconomic status (SES) and obesity supported the view that obesity in the developing world would be essentially a disease of the socioeconomic elite. The present review, on studies conducted in adult populations from developing countries, published between 1989 and 2003, shows a different scenario for the relationship between SES and obesity. Although more studies are necessary to clarify the exact nature of this relationship, particularly among men, three main conclusions emerge from the studies reviewed: 1. Obesity in the developing world can no longer be considered solely a disease of groups with higher SES. 2. The burden of obesity in each developing country tends to shift towards the groups with lower SES as the country's gross national product (GNP) increases. 3. The shift of obesity towards women with low SES apparently occurs at an earlier stage of economic development than it does for men. The crossover to higher rates of obesity among women of low SES is found at a GNP per capita of about US$ 2500, the mid-point value for lower-middle-income economies. The results of this review reinforce the urgent need to: include obesity prevention as a relevant topic on the public health agenda in developing countries; improve the access of all social classes in these countries to reliable information on the determinants and consequences of obesity; and design and implement consistent public actions on the physical, economic, and sociocultural environment that make healthier choices concerning diet and physical activity feasible for all. A significant step in this direction was taken with the approval of the Global Strategy on Diet, Physical Activity and Health by the World Health Assembly in May 2004. PMID- 15654410 TI - Health in the developing world: achieving the Millennium Development Goals. AB - The Millennium Development Goals depend critically on scaling up public health investments in developing countries. As a matter of urgency, developing-country governments must present detailed investment plans that are sufficiently ambitious to meet the goals, and the plans must be inserted into existing donor processes. Donor countries must keep the promises they have often reiterated of increased assistance, which they can easily afford, to help improve health in the developing countries and ensure stability for the whole world. PMID- 15654412 TI - Arguing the case for strengthening health systems. PMID- 15654413 TI - Macroeconomics and Health Commission findings become reality. PMID- 15654419 TI - Epidemiology of leishmaniasis in Ecuador: current status of knowledge -- a review. AB - Although leishmaniasis is regarded as a significant health problem in Ecuador by the Ministry of Health, and the incidence has increased over the last years, an official map on the geographic distribution of disease and sand fly vectors or a control strategy do not exist yet. This article reviews the current situation based on published information to improve our knowledge and understand the epidemiological situation of leishmaniasis in Ecuador in order to help future research and to develop a national control strategy. The disease is endemic in most provinces throughout Pacific coastal region, Amazonian lowlands, and some inter-Andean valleys with a total 21,805 cases reported during 1990-2003. Whereas cutaneous leishmaniasis (CL) is found throughout Ecuador, mucocutaneous leishmaniasis (MCL) appears to be restricted to the Amazon region; one, parasitologically unconfirmed case of visceral form was reported in 1949. Most human infections are caused by Leishmania (Viannia) spp., which is distributed in the subtropical and tropical lowlands; infections due to L. (Leishmania) spp. are found in the Andean highlands and in the Pacific lowlands as well. The proven vectors are Lutzomyia trapidoi and Lu. ayacuchensis. Canis familiaris, Sciurus vulgaris, Potos flavus, and Tamandua tetradactyla have been found infected with Leishmania spp. It is estimated that around 3000-4500 people may be infected every year, and that 3.1 to 4.5 millions people are estimated to be at risk of contracting leishmaniasis. PMID- 15654420 TI - Distribution and Schistosoma mansoni infection of Biomphalaria glabrata in different habitats in a rural area in the Jequitinhonha Valley, Minas Gerais, Brazil: environmental and epidemiological aspects. AB - This paper examines the distribution and infection of Biomphalaria glabrata with Schistosoma mansoni in all aquatic snail habitats in a rural area in the state of Minas Gerais, Brazil, in relation to physico/biotic and behavioral factors. Snail and environmental surveys were carried out semi-annually between July 2001 and November 2002 at 106 sites. Collected snails were examined in the laboratory for infection. B. glabrata densities were highest in overflow ponds, irrigation ponds, springs, canals and wells, and lowest in fishponds and water tanks. Snail densities were higher during the hot, rainy season except for streams and canals and were statistically associated with the presence of fish, pollution, and vegetation density. Tilapia fish and an unidentified Diptera larva were found to be predators of B. glabrata but ducks were not. Twenty-four of the 25 infected snails were collected in 2001(1.4% infection rate) and only one in 2002, after mass chemotherapy. The occurrence of B. glabrata in all 11 snail habitats both at and away from water contact sites studied indicates widespread risk of human infection in the study area. In spite of the strong association between B. glabrata and tilapia in fishponds we do not recommend its use in schistosomiasis control for ecological reasons and its relative inefficiency in streams and dams. PMID- 15654421 TI - Spatial analysis of the distribution of leprosy in the State of Ceara, Northeast Brazil. AB - The aim of this study was to describe spatial patterns of the distribution of leprosy and to investigate spatial clustering of incidence rates in the state of Ceara, Northeast Brazil. The average incidence rate of leprosy for the period of 1991 to 1999 was calculated for each municipality of Ceara. Maps were used to describe the spatial distribution of the disease, and spatial statistics were applied to explore large- and small-scale variations of incidence rates. Three regions were identified in which the incidence of leprosy was particularly high. A spatial gradient in the incidence rates was identified, with a tendency of high rates to be concentrated on the North-South axis in the middle region of the state. Moran's I statistic indicated that a significant spatial autocorrelation also existed. The spatial distribution of leprosy in Ceara is heterogeneous. The reasons for spatial clustering of disease rates are not known, but might be related to an heterogeneous distribution of other factors such as crowding, social inequality, and environmental characteristics which by themselves determine the transmission of Mycobacterium leprae. PMID- 15654422 TI - Naturally acquired immunity to Haemophilus influenzae type B in healthy Cuban children. AB - We have evaluated the prevalence of antibody to immunogenicity of Haemophilus influenzae type b (Hib) in a group of 4 to 5 years old healthy children, who were too old to be included in the first vaccinated cohort when Hib vaccination begun in Cuba in 1999. Serum capsular polysaccharide specific IgG antibody concentrations were measured in 974 healthy children, between February and May 2002. The prevalence of Hib nasopharyngeal carriage was also estimated. The majority of children (99.7%) had more than 1 microg/ml of antibody. The preliminary report of the nasopharyngeal cultures was positive for H. influenzae in 16 children, but in only one was confirmed as Hib after serotyping (0.1% Hib nasopharyngeal carrier). These results provide evidence that in Cuba the natural active immunity to Hib can be acquired at an early age. PMID- 15654423 TI - Studies of membrane fluidity and heart contractile force in Trypanosoma cruzi infected mice. AB - In Chagas disease serious cardiac dysfunction can appear. We specifically studied the cardiac function by evaluating: ventricle contractile force and norepinephrine response, affinity and density of beta-adrenergic receptors, dynamic properties of myocardial membranes, and electrocardiography. Albino swiss mice (n = 250) were infected with 55 trypomastigotes, Tulahuen strain and studied at 35, 75, and 180 days post-infection, that correspond to the acute, indeterminate, and chronic phase respectively. Cardiac beta-adrenergic receptors' affinity, myocardial contractility, and norepinephrine response progressively decreased from the acute to the chronic phase of the disease (p < 0.01). The density (expressed as fmol/mg.prot) of the receptors was similar to non-infected mice (71.96 +/- 0.36) in both the acute (78.24 +/- 1.67) and indeterminate phases (77.28 +/- 0.91), but lower in the chronic disease (53.32 +/- 0.71). Electrocardiographic abnormalities began in the acute phase and were found in 65% of the infected-mice during the indeterminate and chronic phases. Membrane contents of triglycerides, cholesterol, and anisotropy were similar in all groups. A quadratic correlation between the affinity to beta-adrenergic receptors and cardiac contractile force was obtained. In conclusion the changes in cardiac beta-adrenergic receptors suggests a correlation between the modified beta adrenergic receptors affinity and the cardiac contractile force. PMID- 15654424 TI - In vitro measurement of enzymatic markers as a tool to detect mouse cardiomyocytes injury. AB - Primary cultures of cardiomyocytes represent a useful model for analyzing cardiac cell biology as well as pathogenesis of several cardiovascular disorders. Our aim was to standardize protocols for determining the damage of cardiac cells cultured in vitro by measuring the creatine kinase and its cardiac isotype and lactate dehydrogenase activities in the supernatants of mice cardiomyocytes submitted to different protocols of cell lysis. Our data showed that due to its higher specificity, the cardiac isotype creatine kinase was the most sensitive as compared to the others studied enzymatic markers, and can be used to monitor and evaluate cardiac damage in in vitro assays. PMID- 15654425 TI - Immunological tolerance to pig-serum partially inhibits the formation of septal fibrosis of the liver in Capillaria hepatica-infected rats. AB - Systhematized septal fibrosis of the liver can be induced in rats either by repeated intraperitoneal injections of pig-serum or by Capillaria hepatica infection. The relationship between these two etiological factors, as far as hepatic fibrosis is concerned, is not known, and present investigation attempts to investigate it. C. hepatica-induced septal fibrosis of the liver was considerably inhibited in rats previously rendered tolerant to pig-serum. Pig serum-tolerant rats developed antibodies against pig-serum when infected with C. hepatica, but this did not happen when the infection occurred in normal rats. On the other hand, anti-C. hepatica antibodies failed to recognize any epitope in pig-serum, by Western blot. However, no evidence of an immunological cross reactivity was found, at least at the humoral level. Alternatively, cell-mediated mechanisms may be involved, and further investigations are warranted. PMID- 15654426 TI - Effect of the Aedes fluviatilis saliva on the development of Plasmodium gallinaceum infection in Gallus (gallus) domesticus. AB - Effect of Aedes fluviatilis saliva on the development of Plasmodium gallinaceum experimental infection in Gallus (gallus) domesticus was studied in distinct aspects. Chickens subcutaneously infected with sporozoites in the presence of the mosquito salivary gland homogenates (SGH) showed higher levels of parasitaemia when compared to those ones that received only the sporozoites. However, the parasitaemia levels were lower among chickens previously immunized by SGH or non infected mosquito bites compared to the controls, which did not receive saliva. High levels of anti-saliva antibodies were observed in those immunized chickens. Moreover, 53 and 102 kDa saliva proteins were recognized by sera from immunized chickens. After the sporozoite challenge, the chickens also showed significant levels of anti-sporozoite antibodies. However, the ability to generate anti sporozoites antibodies was not correlated to the saliva immunization. Our results suggest that mosquito saliva components enhance P. gallinaceum parasite development in naive chickens. However, the prior exposure of chickens to salivary components controls the parasitemia levels in infected individuals. PMID- 15654427 TI - Alteration in the endogenous intestinal flora of Swiss Webster mice by experimental Angiostrongylus costaricensis infection. AB - The association between worm infections and bacterial diseases has only recently been emphasized. This study examined the effect of experimental Angiostrongylus costaricensis infection on endogenous intestinal flora of Swiss Webster mice. Eight mice aging six weeks were selected for this experiment. Four were infected with A. costaricensis and the other four were used as controls. Twenty eight days after the worm infection, all mice in both groups were sacrificed and samples of the contents of the ileum and colon were obtained and cultured for aerobic and anaerobic bacteria. In the mice infected with A. costaricensis there was a significant increase in the number of bacteria of the endogenous intestinal flora, accompanied by a decrease in the number of Peptostreptococcus spp. This alteration in the intestinal flora of mice infected by the nematode may help to understand some bacterial infections described in humans. PMID- 15654429 TI - Differences in the stability of the plasmids of Yersinia pestis cultures in vitro: impact on virulence. AB - Plasmid and chromosomal genes encode determinants of virulence for Yersinia pestis, the causative agent of plague. However, in vitro, Y. pestis genome is very plastic and several changes have been described. To evaluate the alterations in the plasmid content of the cultures in vitro and the impact of the alterations to their pathogenicity, three Y. pestis isolates were submitted to serial subculture, analysis of the plasmid content, and testing for the presence of characteristic genes in each plasmid of colonies selected after subculture. Different results were obtained with each strain. The plasmid content of one of them was shown to be stable; no apparent alteration was produced through 32 subcultures. In the other two strains, several alterations were observed. LD50 in mice of the parental strains and the derived cultures with different plasmid content were compared. No changes in the virulence plasmid content could be specifically correlated with changes in the LD50. PMID- 15654428 TI - Trichurid nematodes in ring-necked pheasants from backyard flocks of the State of Rio de Janeiro, Brazil: frequency and pathology. AB - The present investigation is related to the frequency of infection and to the gross and microscopic lesions associated to the presence of trichurid worms in 50 ring-necked pheasants (Phasianus colchicus) from backyard flocks in the state of Rio de Janeiro, Brazil. In the investigated birds, the overall infection rate was of 74%, with the presence of Eucoleus perforans with 72% of prevalence and 21.2 of mean intensity, in the esophageal and crop mucosa and rarely in the junction of the proventriculus and esophagus, E. annulatus with 2% and 3 in the crop mucosa, Capillaria phasianina, with 12% and 4.3 in the cecum and small intestine and Baruscapillaria obsignata, for the first time referred in this host, with 2% and 1 in the small intestine. Clinical signs were absent. The gross lesions observed in the crop and esophagus of 14 (38.9%) pheasants parasitized with E. perforans were thickening, small nodules, congestion, and petechial haemorrhages in the mucosa. These birds presented a mean infection of 37.5 and a range of infection of 10-82. The microscopic lesions revealed chronic esophagitis with diffuse inflammatory process in the lamina propria characterized mostly by a mononuclear cell infiltrate and also with the presence of granulocytes. In the case of the parasitism of pheasants with C. phasianina, the gross lesions were absent; microscopic lesions were characterized by chronic typhlitis with mononuclear infiltrate. Gross and microscopic lesions were absent in the pheasants parasitized with E. annulatus and B. obsignata. PMID- 15654430 TI - Similarity between the association factor of ribosomal subunits and the protein Stm1p from Saccharomyces cerevisiae. AB - A ribosome association factor (AF) was isolated from the yeast Sacchharomyces cerevisiae. Partial amino acid sequence of AF was determined from its fragment of 25 kDa isolated by treating AF with 2-(2-nitrophenylsulfenyl)-3-methyl-3' Bromoindolenine (BNPS-skatole). This sequence has a 86% identity to the product of the single-copy S. cerevisiae STM1 gene that is apparently involved in several events like binding to quadruplex and triplex nucleic acids and participating in apoptosis, stability of telomere structures, cell cycle, and ribosomal function. Here we show that AF and Stm1p share some characteristics: both bind to quadruplex and Pu triplex DNA, associates ribosomal subunits, and are thermostable. These observations suggest that these polypeptides belong to a family of proteins that may have roles in the translation process. PMID- 15654431 TI - Molecular characterization of Mycobacterium kansasii isolates in the State of Sao Paulo between 1995-1998. AB - Mycobacterium kansasii is the most common cause of pulmonary nontuberculous mycobacteria infection and classical identification of this pathogen needs a time consuming phenotypic tests. Polymerase chain reaction-restriction fragment length polymorphism analysis (PRA) of the gene enconding for the 65 kDa heat shock (hsp65) protein offers an easy, rapid, and inexpensive procedure to identify and subtype M. kansasii isolates. In the present study, we performed a retrospective analysis of patients who had mycobacteria identified on the basis of phenotypic tests by means of a review of database at Mycobacteria Laboratory of the Instituto Adolfo Lutz in the period 1995-1998. A total of 9381 clinical isolates were analyzed of which 7777 (82.9%) were identified as M. tuberculosis complex and 1604 (17.1%) as nontuberculous mycobacteria. Of the 296 M. kansasii isolates, 189 (63.8%) isolates obtained from 119 patients were viable and were analyzed by PRA-hsp65. Hundred eight two (98.9%) were classified as M. kansasii type I. Two isolates were classified as type II and III and five isolates were characterized as other Mycobacterium species. Clinical isolates of M. kansasii in the state of Sao Paulo was almost exclusively subtype I regardless of HIV status. PMID- 15654432 TI - Mycobacterium tuberculosis complex differentiation using gyrB-restriction fragment length polymorphism analysis. AB - Mycobacterium tuberculosis complex (MTBC) members are causative agents of human and animal tuberculosis. Differentiation of MTBC members is required for appropriate treatment of individual patients and for epidemiological purposes. Strains from six MTBC species -- M. tuberculosis, M. bovis subsp. bovis, M. bovis BCG, M. africanum, M. pinnipedii, and "M. canetti" -- were studied using gyrB restriction fragment length polymorphism (gyrB-RFLP) analysis. A table was elaborated, based on observed restriction patterns and published gyrB sequences. To evaluate applicability of gyrB-RFLP at Instituto Adolfo Lutz, Sao Paulo, Mycobacterial Reference Laboratory, 311 MTBC clinical isolates, previously identified using traditional methods as M. tuberculosis (306), M. bovis (3), and M. bovis BCG (2), were analyzed by gyrB-RFLP. All isolates were correctly identified by the molecular method, but no distinction between M. bovis and M. bovis BCG was obtained. Differentiation of M. tuberculosis and M. bovis is of utmost importance, because they require different treatment schedules. In conclusion, gyrB-RFLP is accurate and easy-to-perform, with potential to reduce time needed for conventional differentiation methods. However, application for epidemiological studies remains limited, because it cannot differentiate M. tuberculosis from M. africanum subtype II, and "M. canetti", M. africanum subtype I from M. pinnipedii, and. M. bovis from M. bovis BCG. PMID- 15654433 TI - Drug susceptibility of Brazilian strains of Mycobacterium bovis using traditional and molecular techniques. AB - Transmission of Mycobacterium bovis from cattle to humans has been reported and can cause tuberculosis (Tb) and a problem in certain risk populations. Therefore, knowledge of resistance of M. bovis towards antibiotics used for therapy of human Tb could help avoiding cure delay and treatment cost increase when dealing with drug resistant organisms. We therefore evaluated the susceptibility of M. bovis isolates towards streptomycin, isoniazide, rifampicin, ethambutol, and ethionamide, the first line antibiotics for human Tb. Therefore, 185 clinical samples from cattle with clinical signs of tuberculosis were processed and submitted to culturing and bacterial isolates to identification and drug susceptibility testing using the proportion method. Among 89 mycobacterial strains, 65 were identified as M. bovis and none were resistant to any of the antibiotics used. Confirmation of present results by future studies, enrolling a large number of isolates and designed to properly represent Brazilian regions, may favor the idea of using isoniazide preventive therapy as part of a Tb control strategy in special situations. Also, nucleic acids from bacterial isolates were submitted to rifoligotyping, a recently described reverse hybridization assay for detection of mutations causing resistance towards rifampicin. Concordance between the conventional and the molecular test was 100%, demonstrating the use of such methodology for rapid evaluation of drug susceptibility in M. bovis. PMID- 15654434 TI - Antimicrobial activity and chemical investigation of Brazilian Drosera. AB - The antimicrobial activity of three different extracts (hexanic, ethyl acetate, methanol) obtained from Brazilian Drosera species (D. communis, D. montana var. montana, D. brevifolia, D. villosa var. graomogolensis, D. villosa var. villosa, Drosera sp. 1, and Drosera sp. 2 ) were tested against Staphylococcus aureus (ATCC 25923), Enterococcus faecium (ATCC23212), Pseudomonas aeruginosa (ATCC27853), Escherichia coli (ATCC11229), Salmonella choleraesuis (ATCC10708), Klebsiella pneumoniae (ATCC13883), and Candida albicans (a human isolate). Better antimicrobial activity was observed with D. communis and D. montana var. montana ethyl acetate extracts. Phytochemical analyses from D. communis, D. montana var. montana and D. brevifolia yielded 5-hydroxy-2-methyl-1,4-naphthoquinone (plumbagin); long chain aliphatic hydrocarbons were isolated from D. communis and from D. villosa var. villosa, a mixture of long chain aliphatic alcohols and carboxylic acids, was isolated from D. communis and 3b-O-acetylaleuritolic acid from D. villosa var. villosa. PMID- 15654435 TI - Antileishmanial activity of lapachol analogues. AB - The antileishmanial activity of lapachol, isolapachol, and dihydrolapachol, along with soluble derivatives (potassium salt) and acetate was obtained. All the compounds were assayed against metacyclic promastigotes of two different species of Leishmania associated to tegumentar leishmaniasis: L. amazonensis and L. braziliensis. All compounds presented significant activity, being isolapachol acetate the most active against promastigotes, with IC50/24h = 1.6 +/- 0.0 microg/ml and 3.4 +/- 0.5 microg/ml for, respectively, L. amazonensis and L. braziliensis. This compound was also assayed in vivo against L. amazonensis and showed to be active. Its toxicity in vitro was also established, and at concentration similar to the IC50, no toxicity was evidenced. In all experiments, pentamidine isethionate was used as a reference drug. The present results reinforce the potential use of substituted hydroxyquinones and derivatives as promising antileishmanial drugs and suggest a continuing study within this class of compounds. PMID- 15654437 TI - Detection of Entamoeba histolytica/Entamoeba dispar in stool specimens by using enzyme-linked immunosorbent assay. AB - Entamoeba histolytica actually comprises two genetically distinct but morphologically indistinguishable species. E. histolytica can cause invasive intestinal and extra intestinal disease, while E. dispar cannot. Identification and differentiation of E. dispar and E. histolytica in stool sample by microscopy is imprecise. Several weeks of culture and isoenzyme analysis are required to differentiate E. histolytica from E. dispar. In this study, we have used an enzyme-linked immunosorbent assay (ELISA) for detection of E. histolytica/E.dispar and compared it with microscopy. Eighty-eight samples were evaluated, trichrome staining was positive in 20.4% (18) and ELISA was positive in 29.5% (26). Both tests were positive in 14 (15.9%) samples, 4 (4.5%) only with direct microscopy, and 12 (13.6%) only with ELISA. Both tests were negative in 58 (65.9%) samples. Microscopy has low sensitivity and high specificity, low negative predictive value and high positive predictive value in comparison with ELISA. E. histolytica/E. dispar antigen detection ELISA tests are inexpensive compared to the specific tests, yield objective results and do not require experienced microscopists and can therefore be recommended for screening of stools worldwide and the results can be taken for treatment that are fitting with its clinic. PMID- 15654436 TI - Antimicrobial resistance of Enterococcus sp. isolated from the intestinal tract of patients from a university hospital in Brazil. AB - This study reports the results about antimicrobial resistance of Enterococcus spp. isolated from intestinal tract of patients from a university hospital in Brazil. The identification of strains at species level was performed by conventional biochemical tests, API 20 Strep (bioMerieux), and polymerase chain reaction assay. The species distribution was E. faecium (34%), followed by E. faecalis (33%), E. gallinarum (23.7%), E. casseliflavus (5.2%), E. avium (1%), and E. hirae (1%). Intrinsic resistance to vancomycin characterized by presence of vanC genes was found in E. gallinarum and E. casseliflavus. The high prevalence of VanC phenotype enterococci is very important because these species have been reported as causing a wide variety of infections. Vancomycin-resistant E. faecium or E. faecalis were not found and no one isolate of these species was a beta-lactamase producer. Thirteen clinical isolates of enterococci (13.4%) showed multiresistance patterns, which were defined by resistance to three classes of antibiotics plus resistance to at least one aminoglycoside (gentamicin and/or streptomycin). The resistance to several antimicrobials shown by enterococcal strains obtained in this study is of concern because of the decrease in the therapeutic options for treatment of infections caused by enterococci. PMID- 15654438 TI - Intestinal parasitic infections and eosinophilia in an human immunedeficiency virus positive population in Honduras. AB - The occurrence of intestinal parasites, their regional distribution and their relations to eosinophilia were studied in 133 human immunodeficiency virus (HIV) positive individuals from Honduras. After signing an informed consent, participants answered a socio-demographic and risk factor questionnaire, a complete physical examination, medical history, and a series of laboratory tests. All participants were HIV positive but not acquired immunodeficiency syndrome positive. Of them, 67% were co-infected with pathogen and non pathogen parasites. Overall occurrence of nematodes was: 44.3% for Trichuris trichiura, 24% for Ascaris lumbricoides, 12% for Hookworm and 7.5% for Strongyloides stercoralis. No cases of Giardia lamblia, acute amebiasis or cryptosporidiasis were diagnosed. Mean eosinophil percents for participants were consistently and significantly higher in infected than in non infected individuals: 22% for Hookworm vs 7.2% (p < 0.001), 11% for Trichuris compared to 5.2% (p < 0.001), 13.2% compared to 7.5% for S. stercoralis (p < 0.05), and 12% compared to 6% for Ascaris cases (p < 0.05). Helminths and non pathogenic protozoa, as single or mixed infections, occurred among the participants. There was a strong correlation between eosinophilia and helminthiasis infections; however, none was identified between CD4 levels and eosinophilia. Because parasitic infections aggravate malnutrition and promote a disbalanced Th2 response in a potentially immuno-compromised host, their effect on HIV disease progression needs further study, mainly in countries were HIV and parasitic infections are highly prevalent. PMID- 15654440 TI - [Cardiovascular health in the technological era]. PMID- 15654441 TI - [Consolidation of cardiovascular science in Brazil]. PMID- 15654442 TI - [The post graduate program of the school of medicine of UFRGS on cardiology and cardiovascular sciences]. PMID- 15654443 TI - Changes in the parameters of left ventricular diastolic function according to age on tissue Doppler imaging. AB - OBJECTIVE: To determine the correlation between diastolic velocities on tissue Doppler imaging and age in a sample of healthy adults and to correlate age with the velocities of transmitral and pulmonary vein flows. METHODS: Echocardiographic assessment of 51 healthy individuals, whose ages ranged from 21 to 69 years. The diastolic myocardial velocities were recorded on tissue Doppler imaging. The velocities of the transmitral and pulmonary vein flows were also determined. RESULTS: The initial basal septal and basal lateral diastolic myocardial velocities showed an inverse correlation with age [r = -0.40 (P = 0.004), and r = -0.60 (P = 0.0001), respectively]. The atriogenic velocities of tissue Doppler imaging correlated directly with age [r = 0.56 (P = 0.0001) in the basal septal segment, and r = 0.50 (P = 0.0001) in the basal lateral segment]. The velocities of transmitral and pulmonary vein flows also correlated with age. CONCLUSION: Age correlates with the tissue Doppler diastolic myocardial velocities and with the velocities of transmitral and pulmonary vein flows. In healthy individuals, the parameters of left ventricular diastolic function vary with the natural evolution of age. PMID- 15654444 TI - Study of global left atrial shortening in fetuses of diabetic mothers. AB - OBJECTIVE: To test the hypothesis that left atrial shortening fraction is lower in fetuses of diabetic mothers than in fetuses of mothers with no systemic disease. METHODS: Forty-two fetuses of mothers with previous diabetes or gestational diabetes and 39 healthy fetuses of mothers with no systemic disease (controls) underwent echocardiographic examination. Their gestational ages ranged from 25 weeks to term. The left atrial shortening fraction was obtained with the following formula: (left atrial maximum diameter - left atrial minimum diameter)/left atrial maximum diameter. Data were compared using the Student t test, with an alpha level of 0.05. RESULTS: Mean left atrial shortening fractions in fetuses of diabetic mothers and in those in the control group were 0.39 +/- 0.15 and 0.51 +/- 0.11, respectively. This difference was significant with P < 0.001. CONCLUSION: Left atrial dynamics, with a reduction in global left atrial shortening, is increased in fetuses of diabetic mothers. We speculate that this parameter may be useful in assessing fetal left ventricular diastolic function. PMID- 15654445 TI - Comparison of LDL-cholesterol direct measurement with the estimate using the Friedewald formula in a sample of 10,664 patients. AB - OBJECTIVE: To compare direct measurement of LDL-cholesterol (LDL-C) determined by a homogeneous method with LDL-cholesterol estimation determined by the Friedewald formula in a large heterogeneous population. METHODS: The measurements of total cholesterol (TC) and triglycerides (TG) were performed using traditional enzymatic methods. The measurements of HDL-C and LDL-C were performed using direct methods with no precipitation, and the estimation of the LDL-C fraction was calculated using the Friedewald formula. RESULTS: On linear regression analysis, the 2 methods had extremely significant correlation coefficients (P < 0.001). However, the Friedewald formula had a positive bias in regard to the direct method, more pronounced with TC levels > 201 mg/dL. This positive bias also occurred in regard to TG levels < or =150 mg/dL. No bias was observed between the methods for TG levels ranging from 151 to 200 mg/dL and from 201 to 300 mg/dL. On the other hand, for TG levels ranging from 301 to 400 mg/dL, this bias of the Friedewald formula became negative. CONCLUSION: The Friedewald formula did not have a homogeneous performance for estimating LDL-C levels in samples with different TG levels as compared with that of the direct method, what could launch doubts on patients classification on the risk of developing coronary artery disease. PMID- 15654446 TI - Effects of oxidized LDL on in vitro proliferation and spontaneous motility of human coronary artery endothelial cells. AB - OBJECTIVE: To investigate the effects of low concentrations of oxidized LDL (oxLDL) on the proliferation and spontaneous motility of human coronary artery endothelial cells (HCAEC) in culture. METHODS: Cultures of HCAEC were treated with low concentrations of native LDL (nLDL) isolated from human plasma and with LDL minimally oxidized through different chemical methods; the effects were compared. RESULTS: Native LDL had no deleterious effects on in vitro proliferation and motility of HCAEC; however, at its highest concentration and for a longer exposure, nLDL inhibited cell proliferation. The LDL chemically oxidized by spermine nonoate (SNO) and 3-morpholinylsydnonimine (SYN-1) had significant inhibiting effects on in vitro proliferation and motility of HCAEC, which were proportional to the greatest concentrations and degrees of oxidation of LDL. CONCLUSION: OxLDL has a cytotoxic effect, inhibiting the proliferation and spontaneous motility of HCAEC in culture. This effect is proportional to the concentration and degree of oxidation of LDL; native LDL is relatively innocuous. PMID- 15654447 TI - Anomalous origin of a pulmonary artery from the ascending aorta: surgical repair resolving pulmonary arterial hypertension. AB - OBJECTIVE: To emphasize the diagnostic possibility of the anomalous origin of one pulmonary artery from the ascending aorta in infants with clinically refractory heart failure and no intracardiac structural defect. METHODS: Retrospective study of 4 infants with refractory heart failure undergoing 2-dimensional echocardiographic study with subcostal, suprasternal, and parasternal views, and hemodynamic and angiocardiographic study in the anteroposterior projection. RESULTS: Three of the 4 infants had their right pulmonary artery originating from the ascending aorta as their major diagnosis. In the fourth patient, the left pulmonary artery originated from the ascending aorta in association with a large interventricular septal defect. The pressure level in both pulmonary arteries in all infants was that of the systemic level. All patients underwent surgery, which consisted of translocation of the anomalous pulmonary artery from the aorta. Neither immediate nor late cardiac deaths occurred. CONCLUSION: Once the diagnosis of anomalous origin of the pulmonary artery from the ascending aorta in the isolated form is established, the surgical correction should be immediately performed, not only because of the risk of developing pulmonary vascular disease, but also because of the excellent surgical results currently obtained. PMID- 15654448 TI - Lack of effect of captopril on the metabolism of an artificial lipid emulsion similar to chylomicrons in hypertensive hypercholesterolemic patients. AB - OBJECTIVE: To assess the effect of captopril, an angiotensin-converting enzyme inhibitor, on the metabolism of chylomicrons and their remnants and the possible alterations in the concentrations of plasma lipids caused by the drug in hypertensive hypercholesterolemic individuals. METHODS: The metabolism of chylomicrons was tested with the method of artificial lipid emulsion of chylomicrons labeled with 3H-cholesteryl oleate. The emulsion was injected intravenously in 10 patients with mild-moderate arterial hypertension before and 45 days after treatment with captopril (50 mg/day). After injection, blood samples were collected during 60 minutes at pre-established time intervals for determining the decay curve, the fractional catabolic rate (FCR in min-1), and the plasma residence time of the artificial lipid emulsion by analyzing different compartments. The plasma concentrations of the lipids were also assessed before and after treatment. RESULTS: The fractional catabolic rate (min-1) of the lipid emulsion before and after treatment with captopril (0.012 +/- 0.003 and 0.011 +/- 0.003, respectively; p = 0.85, n.s.) and the plasma residence time of the emulsion (83.3 +/- 20.8 and 90.9 +/- 22.5 min, n.s.) did not change, but the total cholesterol and LDL-C levels decreased by 7% and 10%, respectively (p = 0.02). The concentrations of HDL-C, triglycerides, Lp(a), and apolipoproteins AI and B did not change. CONCLUSION: Treatment with captopril, evaluated with the artificial lipid emulsion method, does not cause deleterious changes in the metabolism of chylomicrons and their remnants. PMID- 15654449 TI - Surgical repair of the anomalous origin of the right pulmonary artery from the ascending aorta. AB - The anomalous origin of the right pulmonary artery (AORPA) from the ascending aorta is a rare congenital malformation. We describe an infant who underwent a surgical correction with direct anastomosis between the right pulmonary artery and the pulmonary trunk. Eighteen months later, the patient remains asymptomatic, and no significant residual stenosis was detected on angioresonance. PMID- 15654450 TI - [Endovascular approach of iliac occlusion with acute ischemia of the interior member. A simple and effective procedure in high risk patients]. PMID- 15654452 TI - [Case 10/2004 - Instituto do Coracao do Hospital das Clinicas da FMSUP]. PMID- 15654451 TI - [Case 06/2004 - Sudden death in a 69 year-old woman in the third day after coronary aortic bypass surgery performed in acute phase of myocardial infarction. Instituto do Coracao (InCor) HC-FMUSP]. PMID- 15654453 TI - [Extracranial carotid stenosis]. PMID- 15654454 TI - Factorial validity of the Maslach Burnout Inventory (MBI-HSS) among Spanish professionals. AB - OBJECTIVE: To assess the factorial validity and internal consistency of the Maslach Burnout Inventory (MBI-HSS). METHODS: In a sample consisting of 705 Spanish professionals from diverse occupational sectors (health, education, police and so one), seven plausible factorial models hypothesized were compared using LISREL 8. RESULTS: The four-factor oblique solution and the three-factor oblique solution showed the best and similar fit. Deletion of Item 12 and Item 16, taking into consideration the suggestions in the manual, improved the goodness of fit for both models. The four-factor oblique model suggests that, in addition to Emotional Exhaustion (EE) and Depersonalization (DP), Personal Accomplishment (PA) consists of two components labeled here Self-Competence (Items 4, 7, 17, and 21) and the Existential Component (Items 9, 12, 18, and 19). However, the alpha coefficient was relatively low for the Self-Competence component, suggesting that it is more suitable to estimate the syndrome as a three-dimensional construct. The Cronbach's alpha was satisfactory for PA (alpha =.71) and EE (alpha =.85), and moderate for DP (alpha =.58). CONCLUSIONS: The results show that the MBI-HSS offers factorial validity and its scales present internal consistency to evaluate the quality of working life for Spanish professionals. PMID- 15654455 TI - [Combined exposure to noise and vibration and its effects on workers' hearing]. AB - OBJECTIVE: To assess the exposure of bus drivers to noise and whole-body vibration (WBV) and to examine the possibility of an association between these risk factors for noise-induced hearing loss. METHODS: A cross-sectional study was carried out among 141 bus drivers who underwent an audiometry test. This group was classified and internally stratified in subgroups of "exposed" and "controls" according to cumulative working time as bus drivers. Their exposure to noise and vibration was assessed. The association between noise-induced hearing loss (NIHL) and the set of explanatory variables was analyzed through logistic regression. RESULTS: The average (+/- standard deviation) weekly noise exposure of front engine bus drivers was 83.6 +/- 1.9 dB(A), while rear-engine bus drivers were exposed to 77.0 +/- 1.1 dB(A). The weighted average of vibration acceleration was 0.85/m(2). In the best adjusted model, the multivariable analysis showed that age (>44; OR=2.54; 95% CI=1.15-5.62), diabetes (OR=5.46; 95% CI=0.95-31.4), and the level of noise emission [>86.8 dB(A); OR=2.76; 95% CI=1.24-6.15] were risk factors for NIHL. In another model studied, WBV exposure was significant in determining NIHL. CONCLUSIONS: Bus drivers were exposed to significant WBV levels. The noise exposure was more pronounced in front engine than in rear engine vehicles. No association between WBV exposure and NIHL was observed and no interaction was found between WBV and noise exposure. Further studies are required as other model indicated an association between WBV and NIHL. PMID- 15654456 TI - [Design and validation of a job-exposure matrix to silica]. AB - OBJECTIVE: To develop a population-based matrix of job-exposure to crystalline silica in Brazil and to estimate its validity. METHODS: An epidemiologist and an industrial hygienist developed a matrix of job-exposure in four stages: coding of occupation variable; coding of industry variable; consensual exposure classification between researchers; and estimate of registered workforce in 1995 for each level of exposure. The cross-tabulation of the variables industry (25 columns) and occupation (347 lines) resulted in 8,675 cells, classified according to silica exposure in four levels: non-exposed, possibly exposed, probably exposed, and definitively exposed. For validating the job-exposure matrix, five industries (mining and quarrying, construction, foundry, management of technical personal and textiles), were re-coded according to exposure by external experts. Reliability of the study and external experts was evaluated by agreement measured using kappa analysis. RESULTS: The job-exposure matrix showed high coding agreement, ranging from 64.0% for foundry to 94.0% for mining. Kappa analysis showed good agreement in mining (0.9), and low or average for other sectors (ranging from 0.1 to 0.5). High specificity was found in foundry (86.5%) and mining (100.0%). Construction had 56% specificity. CONCLUSIONS: The study job exposure matrix showed good accuracy and seems to be appropriate for estimating silica exposure among Brazilian workers. PMID- 15654457 TI - [Feeding patterns of Triatoma pseudomaculata in the state of Ceara, Brazil]. AB - OBJECTIVE: T. pseudomaculata, a peridomicilar species, has low rates of T. cruzi infection. The occurrence of this triatomine in the domicile and its feeding patterns was investigated to identify potential T. cruzi reservoirs. METHODS: Nine-hundred and twenty-one specimens of T. pseudomaculata were captured from January 2001 to July 2002 in 13 southern municipalities of the state of Ceara, Brazil. The intestinal contents of the triatomines was removed, spread in a filter paper and tested for the following antisera: bird, cat, cockroach, dog, human, lizard, opossum, ox/goat, pig, and rodent. The presence of T. cruzi was investigated by wet mount microscopic exam and culture (NNN+LIT) of intestinal contents. RESULTS: Of the total studied, 184 (90.6%) were positive for the tested antisera: bird (62.5%)> rodent (33.7%)> dog (20.1%)> opossum (9.8%)> lizard and ox/goat (5%)> cat (2.7%)> pig and cockroach (2.2%)> human (1.6%). Blood meals ranged from none (non-reactive) to four as follows: non-reactive (9.4%), one (57.1%), two (26%), three (7%), and four (0.5%). Only three specimens (1.6%) had T. cruzi infection. CONCLUSIONS: The low incidence of human blood meal shows that T. pseudomaculata is well-adjusted to the peridomicile. However, the epidemiological vigilance in this region is key due to this species' proximity to domiciles. PMID- 15654459 TI - [Incidence of vancomycin-resistant Enterococcus at a university hospital in Brazil]. AB - OBJECTIVE: Vancomycin-resistant Enterococcus (VRE) is today one of the principal microorganisms implicated in nosocomial infections. Thus, a study was carried out with the objective of evaluating its epidemiology at a tertiary-level teaching hospital. METHODS: This was a three-year retrospective epidemiological study conducted from 2000 to 2002. Samples of VRE-positive clinical cultures at a 660 bed university hospital were analyzed. The incidence of VRE and the main anatomical sites and hospital units from which it was isolated were defined. Differences between the variables over the three years of the study were verified, and these were considered significant when p<0.05. RESULTS: There was a progressive increase in the vancomycin resistance in the clinical cultures that were positive for Enterococcus spp., over the three years of the study. In 2000, 9.5% of the samples were vancomycin-resistant, and this increased to 14.7% in 2001 and 15.8% in 2002. The hospital units with the largest numbers of isolates were, respectively, the emergency ward (19.5%) and the general intensive care unit (15%). The anatomical sites with the highest amounts of isolates included: urine (36%) and blood (20%). CONCLUSIONS: With the progressive increase in the incidence of vancomycin resistance and the VRE rate, it is concluded that more effective control measures are needed for deterring the dissemination of VRE. PMID- 15654458 TI - [Aedes aegypti infestation and occurrence of dengue in the city of Belo Horizonte, Brazil]. AB - OBJECTIVE: To assess the association between the proportion of buildings positive for Aedes aegypti larvae measured by means of building infestation rate and dengue incidence rate. METHODS: Autochthonous dengue cases were selected and building infestation rates assessed in the coverage areas of health districts in the city of Belo Horizonte, Brazil, from October 1997 to May 2001. After grouping building infestation rates according to their distribution in quartiles, mean dengue incidence rates (for the month following the building infestation assessment) were compared using ANOVA. RESULTS: A weak though statistically significant correlation was observed between monthly dengue incidence and building infestation rates for the health districts (r=0.21; p=0.02) and coverage areas (r=0.14; p=0.00) in the study period. After grouping building infestation rates in quartiles, coverage areas of building infestation between 0.46% and 1.32% (second quartile) showed a mean monthly dengue incidence rate twice as high as areas of building infestation lower or equal to 0.45% (first quartile). Areas of building infestation between 1.33% and 2.76% (third quartile) and equal to or higher than 2.77% had a mean monthly dengue incidence rate five and seven times, respectively, higher than those areas showing 0.45% or less. CONCLUSIONS: Despite the well-known limitations of building infestation rates for estimating vector infestation and prediction of dengue epidemics, the study results indicate that higher building infestation rates were associated to a higher risk of disease transmission in the health districts and coverage areas of Belo Horizonte. PMID- 15654461 TI - [Trends of homicide death in Brazil in the 90s: the role of firearms]. AB - OBJECTIVE: To describe the growth of homicide death in Brazil and to investigate the contribution of firearms to homicide death growth in the 90s. METHODS: An ecological descriptive time series was carried out for the period 1991 to 2000. External cause mortality data were obtained according to place of residence for the overall population and by gender from the Ministry of Health Mortality Information System. Homicides were classified as firearm, non-firearm and unknown (weapon) deaths. Proportional mortality and mortality rates (by 100,000) were calculated. RESULTS: Homicides accounted for 33% of all external cause deaths in the 90s. Firearms were used in more than 50% of deaths in 1991 and in almost 70% in 2000. This growth was seen for both genders and in all state capitals. Homicide mortality rate grew 27.5% in the whole country, and firearm homicide mortality rate grew 72.5%. During the same period there was a drop in deaths classified as unknown weapon homicide, which could partially explain the growth seen in firearm homicide rates. CONCLUSIONS: Data suggest the magnitude of firearm contribution for the growing tendency of homicide deaths in Brazil in the 90s. Data quality regarding intentionality and type of weapon used has hindered data analysis. PMID- 15654462 TI - [Trends in avoidable causes of infant mortality in Belo Horizonte, Brazil, 1984 to 1998]. AB - OBJECTIVE: To analyze the infant mortality trend in a metropolitan area, from 1984 to 1998. The main focus was on avoidable causes of neonatal and post neonatal mortality. METHODS: Sources of data were the Sistema de Informacoes em Mortalidade do Ministerio da Saude (SIM-MS) [Mortality Information System of the Ministry of Health] and Fundacao Instituto Brasileiro de Geografia e Estatistica (IBGE) [Brazilian Institute of Geography and Statistics Foundation] (official live birth and death records) for the metropolitan region of Belo Horizonte, in the State of Minas Gerais. A simple linear regression model was used to evaluate time-trends of mortality rates. Statistical significance of the inclination of the regression curves was considered for the p<0.05 level. RESULTS: During the 15 year period in question, the infant mortality rate declined from 48.5 to 22.1/1,000 live births. However, the most accentuated decrease was observed during the last four years of the study period. The post-neonatal group was greatly responsible for this decline both in the capital and in the other districts within the metropolitan region of Belo Horizonte. CONCLUSIONS: Although a significant decrease in the infant mortality rate has been observed, particularly in the post-neonatal mortality, it is still larger than the rates found in developed countries. Deaths due to perinatal morbidities as well as the group of causes represented by diarrhea-pneumonia-malnutrition still present an important potential for reduction. The authors discuss the role of the health services in improving the rates of these avoidable causes of infant mortality. PMID- 15654460 TI - [Surveillance of risk factors for chronic diseases through telephone interviews]. AB - OBJECTIVE: To describe methods and initial findings of a surveillance system of risk factors for chronic non-communicable diseases (CNCDs) based on telephone interviews. METHODS: Interviews undertaken in a random sample of the adult population of the Municipality of Sao Paulo living in households with telephone. Sampling was done in two steps and included the random selection of households and the random selection of the household member to be interviewed. The system's questionnaire investigated demographic and socioeconomic characteristics, food consumption and physical activity patterns, smoking, consumption of alcoholic beverages, recalled weight and height and reported medical diagnoses of hypertension and diabetes, among other topics. Prevalence estimates of selected risk factors for CNCDs were calculated for the adult population with telephone and for the city's entire adult population. In this last case, we applied sample weighting factors that took into account demographic and socioeconomic differences between the adult population with telephone and the entire adult population of the municipality. RESULTS: Strong differences between sexes were found for most risk factors: low consumption of fruit and vegetables, high consumption of alcohol and overweight were more frequent among men while sedentary lifestyle and hypertension were more frequent among women. Additional possibilities of stratification of risk factor prevalences allowed by the surveillance system are illustrated using age groups, schooling, and place of residence in the city. CONCLUSIONS: System performance was considered as good and was better than the performance observed in similar systems operating in developed countries when evaluated with basis on the representativeness and reliability of the estimates and on costs. The cost per concluded interview was eight times lower than the cost usually seen in similar systems in developed countries and four to eight times lower than the cost of traditional household surveys undertaken in the city of Sao Paulo. PMID- 15654463 TI - [Impact of influenza vaccination on mortality by respiratory diseases among Brazilian elderly persons]. AB - OBJECTIVE: Respiratory diseases, especially infectious ones, are becoming increasingly representative in the morbidity and mortality patterns of elderly persons. The aim of the present study was to analyze trends in the mortality by respiratory diseases and to observe the impact of influenza vaccination on mortality rates. METHODS: The study was carried out between 1980 and 2000. Subjects were elderly persons living in the State of Sao Paulo, and mortality data were obtained from the Mortality Information System of the Brazilian Ministry of Health. This is an ecological time-series study. We analyzed the time trends of standardized mortality rates by infectious diseases, according to age group (60-64, 65-69, 70-74, 75-79, and 80+ years) and sex, using polynomial regression. We estimated confidence intervals for the mean expected response in the years following the intervention. RESULTS: Rates increased for both sexes among the elderly population. After the intervention, we observed a declining trend with respect to mortality indicators. For older males, the mean rate in the 1980-1998 period was 5.08 deaths per thousand men, with a nonlinear, non-constant increase of 0.13 per year; in 2000, the rate observed was 4.72 deaths per thousand men. The mean annual rate among women over 60 years was 3.18 deaths per thousand women, with a nonlinear, non-constant increase of 0.08 per year; in 2000, the rate observed was 2.99 deaths per thousand women. There was also a significant reduction in mortality rates in all age groups. CONCLUSIONS: Data indicate the importance of respiratory diseases among the elderly population and suggest that specific protection against influenza has a positive effect on the prevention of mortality due to these diseases. PMID- 15654464 TI - [Tuberculosis in Brazil: construction of a territorially based surveillance system]. AB - OBJECTIVE: To analyze the occurrence of tuberculosis and to identify variables that define situations of collective risk that determine the spatial distribution of the disease, as backing for implementing a territory-based surveillance system for tuberculosis control. METHODS: This was an ecological study performed in Olinda, a municipality in the metropolitan region of Recife, State of Pernambuco, between 1996 and 2000. The median number of notified tuberculosis cases in each census tract served as the cutoff point for characterizing areas of high and low transmission. A logistic regression model using this response variable allowed odds ratios for some socioeconomic variables from the 2000 demographic census and other covariates related to the transmission of the disease to be estimated. RESULTS: Tuberculosis in Olinda presented high incidence rates during the study period (average of 111 cases per 100,000 inhabitants). Significant associations with the occurrence of tuberculosis were found for the variables of average number of inhabitants per household (OR=2.2; 95% CI: 1.3; 3.6); existence of families with more than one case during the study period (OR=5.1; 95% CI: 2.3; 11.3); and presence of cases of retreatment (OR=6.8; 95% CI: 2.7; 17.1). The census tract where the latter two events occurred accounted for 45% of the total number of cases during the study period, while representing only 28% of the population of Olinda. CONCLUSIONS: The two explanatory covariates that were strongly associated with higher incidence rates of the disease are events that need to be carefully monitored at a local level by the tuberculosis surveillance system. Simply by mapping out retreatment cases and households with more than one case, attention could be focused on small areas with high priority for intensive intervention, thus facing up to the tuberculosis problem. PMID- 15654465 TI - [Health services utilization in areas covered by the family health program (Qualis) in Sao Paulo City, Brazil]. AB - OBJECTIVE: The Family Health Program (FHP) is a strategy for reorganizing the healthcare attendance system within the Brazilian National Health System. The objective of the study was to assess whether there had been changes in the utilization profile of the healthcare services following implementation of the program, and to identify factors associated with any changes observed. METHODS: Data on service utilization and demand for attendance were analyzed by means of two cluster-based population samples, representing areas covered (n=1865) and not covered (n=2036) by the FHP, in two districts of the municipality of Sao Paulo. The data formed part of a population survey carried out in 2001. Statistical methods for cluster analysis were used. RESULTS: In the area covered by the FHP, no statistically different prevalence ratios (PR) according to schooling and income levels were observed for service utilization. In the area not covered by the FHP, service utilization was positively associated with greater schooling and income. Among individuals with illnesses, the demand for attendance in the area covered by the FHP was higher (higher PR) among those with severe physical limitations. In the area not covered, the PR was higher among those with greater schooling and lower among those who were inactive (unemployed or retired). CONCLUSIONS: In the areas studied, for the population covered by the FHP, income and schooling levels did not constitute factors that significantly differentiated the utilization profile of the healthcare services and the demand for attendance. This indicates that the program may be contributing towards greater equity under these conditions. PMID- 15654467 TI - [Quality of life and depression in women abused by their partners]. AB - OBJECTIVE: To evaluate quality of life and depression among women who suffer domestic violence and to describe the socioeconomic profile of women who were abused by their partners and aspects of these abuses. METHODS: The study sample comprised 100 women who were abused by their partners and filed a complaint at the Police Office for Women of the state of Ceara, Brazil. Three questionnaires were applied: one for collecting demographic and violence data; the second one (GHQ-28) one to evaluate general quality of life; and the third one (Beck) to quantify depression. RESULTS: Abused women are young, married, Catholic, have children, low schooling and low family income. Alcohol use and jealousy were the main reported factors leading to partner's abuse. Of all, 84% of women suffered physical attacks. It was observed that 72% had depression symptoms; 78% had anxiety symptoms and insomnia; 39% had already thought of killing themselves, and 24% started taking anxiety medications after been abused. CONCLUSIONS: Data analysis suggests that domestic violence is associated with a negative perception by women of their mental heath. PMID- 15654466 TI - [Racial, sociodemographic, and prenatal and childbirth care inequalities in Brazil, 1999-2001]. AB - OBJECTIVE: To analyze social inequalities and inequalities in access to and utilization of health care services according to skin color in a representative sample of postpartum women receiving hospital childbirth care. METHODS: A cross sectional study was carried out in a sample of 9,633 postpartum women, of whom 5,002 were white (51.9%), 2,796 mulatto (29.0%), and 1,835 black skin color (19.0%), seen in public maternity hospitals, hospitals contracted out by the Unified National Health System, and private hospitals in the period 1999-2001. Data were collected from medical records and through interviews with the mothers in the immediate postpartum period using standardized questionnaires. Statistical analyses were performed using chi(2) tests to assess homogeneity of proportions and Student's t-test for comparison of measures. The analysis was stratified by maternal schooling. RESULTS: A persistent unfavorable situation was seen for mulatto and black women as compared to white women. Mulatto and black women had the highest rates of adolescent mothers, low schooling, unpaid occupation, and not having a partner. History of physical violence, smoking, attempts to interrupt pregnancy, and visits to several hospitals before being admitted were more frequent among black women, followed by mulatto and then white women of low schooling. High schooling group of women showed better indicators but the same pattern was seen. This variability is also seen in the opposite direction in terms of the level of satisfaction with prenatal and childbirth care. CONCLUSIONS: It was distinguished two forms of discrimination, by educational level and skin color, in care delivered by health services to postpartum women in Rio de Janeiro. PMID- 15654468 TI - [Data and the process of formulating health policies]. AB - OBJECTIVE: To evaluate the use of the available information systems in decision making process involving municipalities' health services, since technical scientific information is becoming an important tool for managers' decision making both in the private and public sectors. METHODS: Four case studies were undertaken in the state of Sao Paulo between 1998 and 2000. The municipalities included in this study varied in size and in terms of the of complexity of their health systems. Research involved the use of both quantitative (survey of epidemic, demographic, economic-financial and social indicators) and qualitative methods (interviews with key actors and focus group). "Triangulation" was adopted in the analysis in order to establish an articulation among the diverse sources of data and methodological procedures utilized. RESULTS: The strategy of implementation of the Unified Health System (SUS) in itself implies in a pattern of consumption of information already available in large data banks within public institutions and local production of information concerning, primarily, the financial dimension of the city or district, whatever its size, the complexity of the local health system and the type of health administration. CONCLUSIONS: The information available on the data banks are, in general, considered outdated with respect to the immediate needs of local health managers. The equipment infrastructure and training of human resources in health data management were considered precarious for use in the decision-making process. PMID- 15654469 TI - [Tuberculosis in Salvador, Brazil: costs to health system and families]. AB - OBJECTIVE: Tuberculosis is one the greatest causes of mortality worldwide, but its economic effects are not well known. This study had the objective of estimating the costs to the public and private healthcare systems and to families of tuberculosis treatment and prevention. METHODS: This study was made in the municipality of Salvador, State of Bahia, Brazil, in 1999. Data for estimating the costs to the healthcare system were collected from the Department of Health, healthcare facilities and a philanthropic institution. The public and private costs were analyzed using cost accounting methodology. Cost data relating to families were collected by means of questionnaires, and included data on transportation, food and other expenses, and also income losses associated with this disease. RESULTS: The average cost of treating one new case of tuberculosis was approximately US$103. The cost of treating one multiresistant patient was 27 higher than this. The cost to the public services consisted of 65% on hospitalization, 32% on treatment, and only 3% on prevention. The families committed around 33% of their income on expenses related to tuberculosis. CONCLUSIONS: Despite the fact that the families did not have to pay for medications and treatment, given that this service is offered by the State, the costs to families related to loss of income due to the disease were very high. The proportion of public service funds utilized for prevention is small. Greater investment in prevention campaigns not only might diminish the numbers of cases but also might lead to earlier diagnosis, thus reducing the costs associated with hospitalization. The lack of an integrated cost accounting system makes it impossible to visualize costs across the various sectors. PMID- 15654470 TI - Influenza vaccination in Brazil: rationale and caveats. AB - Mass vaccination campaigns against influenza in the elderly have been conducted in Brazil since 1999. A search of the literature on influenza in Brazil indicated that data on disease burden are still scarce and inaccurate. Published data seem to indicate that vaccination has produced some impact in the southern and southeastern regions but not in other regions of Brazil. A discussion of the technical and scientific rationale for mass immunization against influenza is presented and it is argued that the current strategy has not taken into account potential differences in disease occurrence in different areas. It is suggested some epidemiological surveillance actions needed to address major concerns regarding mass influenza vaccination and its impact in Brazil. PMID- 15654471 TI - [Infected Lutzomyia edwardsi found in the Greater Sao Paulo area]. PMID- 15654472 TI - [Mass vaccination campaign against serogroup C meningococcal disease, municipality of Itapeva, Sao Paulo]. PMID- 15654473 TI - American trypanosomiasis and electrocardiographic alterations among industrial workers in Sao Paulo, Brazil. AB - In this article, we evaluate electrocardiographic (ECG) alterations among urban workers from a mechanical-metallurgical industry in Sao Paulo, Brazil. In a cross sectional survey carried out in 1980, we found 2.2% positive serological testing for Trypanosoma cruzi infection among 27,081 workers. A comparison between seropositive workers and a random sample of seronegative workers frequency matched by age and occupation revealed that seropositive workers had a much lower educational level, and that a higher proportion of seropositive workers had ECG abnormalities (42.7%) when compared to those negative for T. cruzi infection (19.8%). The high frequency of ECG abnormalities suggests the need to provide medical assistance to these workers, without any kind of discrimination and to bring in a form of management that would decrease this dangerous risk to the workers and people around them. PMID- 15654474 TI - Serological, epidemiological and molecular aspects of hepatitis C virus infection in a population from Londrina, PR, Brazil, 2001-2002. AB - Serological, epidemiological and molecular aspects of hepatitis C virus (HCV) infection were evaluated in 183 subjects from Londrina, Parana, Brazil, and adjacent areas. Serum samples which tested anti-HCV positive by microparticle enzyme immunoassay (MEIA) obtained from eight patients with chronic hepatitis C, 48 blood donors, and 127 patients infected with the human immunodeficiency virus (HIV) were submitted to another enzyme immunoassay (ELISA) and to the polymerase chain reaction (PCR). About 78.7% of samples were also reactive by ELISA, with the greater proportion (70.8%) of discordant results verified among blood donors. A similar finding was observed for HCV-RNA detection by PCR, with 111/165 (67.3%) positive samples, with higher rates among HIV-positive subjects and patients with chronic hepatitis than among blood donors. Sixty-one PCR-positive samples were submitted to HCV genotyping, with 77.1, 21.3 and 1.6% of the samples identified as types 1, 3 and 2, respectively. Finally, analysis of some risk factors associated with HCV infection showed that intravenous drug use was the most common risk factor among HIV/HCV co-infected patients, while blood transfusion was the most important risk factor in the group without HIV infection. The present study contributed to the knowledge regarding risk factors associated with HCV infection and the distribution of HCV genotypes in the population evaluated. PMID- 15654475 TI - Occurrence of Giardia cysts and Cryptosporidium oocysts in activated sludge samples in Campinas, SP, Brazil. AB - Giardia and Cryptosporidium have caused several outbreaks of gastroenteritis in humans associated with drinking water. Contaminated sewage effluents are recognized as a potential source of waterborne protozoa. Due to the lack of studies about the occurrence of these parasites in sewage samples in Brazil, we compared the efficiency of two procedures for concentrating cysts and oocysts in activated sludge samples of one sewage treatment plant. For this, the samples were submitted to i) concentration by the ether clarification procedure (ECP) and to ii) purification by sucrose flotation method (SFM) and aliquots of the pellets were examined by immunofluorescence. Giardia cysts were present in all samples (100.0%; n = 8) when using ECP and kit 1 reagents, while kit 2 resulted in six positive samples (85.7%; n = 7). As for SFM, cysts were detected in 75.0% and 100.0% of these samples (for kit 1 and 2, respectively). Regarding Cryptosporidium, two samples (25.0%; kit 1 and 28.5% for kit 2) were detected positive by using ECP, while for SFM, only one sample (examined by kit 1) was positive (12.5%). The results of the control trial revealed Giardia and Cryptosporidium recovery efficiency rates for ECP of 54.5% and 9.6%, while SFM was 10.5% and 3.2%, respectively. Considering the high concentration detected, a previous evaluation of the activated sludge before its application in agriculture is recommended and with some improvement, ECP would be an appropriate simple technique for protozoa detection in sewage samples. PMID- 15654477 TI - Human vaccinia-like virus outbreaks in Sao Paulo and Goias States, Brazil: virus detection, isolation and identification. AB - Since October 2001, the Adolfo Lutz Institute has been receiving vesicular fluids and scab specimens of patients from Paraiba Valley region in the Sao Paulo and Minas Gerais States and from Sao Patricio Valley, in the Goias State. Epidemiological data suggested that the outbreaks were caused by Cowpox virus or Vaccinia virus. Most of the patients are dairy milkers that had vesiculo-pustular lesions on the hands, arms, forearms, and some of them, on the face. Virus particles with orthopoxvirus morphology were detected by direct electron microscopy (DEM) in samples of 49 (66.21%) patients of a total of 74 analyzed. Viruses were isolated in Vero cell culture and on chorioallantoic membrane (CAM) of embryonated chicken eggs. Among 21 samples submitted to PCR using primers for hemagglutinin (HA) gene, 19 were positive. Restriction digestion with TaqI resulted in four characteristic Vaccinia virus fragments. HA nucleotide sequences showed 99.9% similarity with Cantagalo virus, described as a strain of Vaccinia virus. The only difference observed was the substitution of one nucleotide in the position 616 leading to change in one amino acid of the protein in the position 206. The phylogenetic analysis showed that the isolates clustered together with Cantagalo virus, other Vaccinia strains and Rabbitpox virus. PMID- 15654479 TI - Dapsone hypersensitivity syndrome in an adolescent during treatment during of leprosy. AB - A 12 y old girl was admitted 24 days after start a WHO multidrug therapy scheme for multibacillary leprosy (dapsone, clofazimine and rifampicin) with intense jaundice, generalized lymphadenopathy, hepatoesplenomegaly, oral erosions, conjunctivitis, morbiliform rash and edema of face, ankles and hands. The main laboratory data on admission included: hemoglobin, 8.4 g/dL; WBC, 15,710 cells/mm3; platelet count, 100,000 cells/mm3; INR = 1.49; increased serum levels of aspartate and alanine aminotransferases, gamma-glutamyl transpeptidase, alkaline phosphatase, direct and indirect bilirubin. Following, the clinical conditions had deteriorated, developing exfoliative dermatitis, shock, generalized edema, acute renal and hepatic failure, pancytopenia, intestinal bleeding, pneumonia, urinary tract infection and bacteremia, needing adrenergic drugs, replacement of fluids and blood product components, and antibiotics. Ten days after admission she started to improve, and was discharged to home at day 39th, after start new supervised treatment for leprosy with clofazimine and rifampicin, without adverse effects. This presentation fulfils the criteria for the diagnosis of dapsone hypersensitivity syndrome (fever, generalized lymphadenopathy, exfoliative rash, anemia and liver involvement with mixed hepatocellular and cholestatic features). Physicians, mainly in geographical areas with high prevalence rates of leprosy, should be aware to this severe, and probably not so rare, hypersensitivity reaction to dapsone. PMID- 15654478 TI - Enzootic transmission of Trypanosoma cruzi and T. rangeli in the Federal District of Brazil. AB - The Federal District of Brazil (DF) lies within the Cerrado biome, where open shrubland (savannas) is interspersed with riverside gallery forests and permanent swamps (veredas). Trypanosoma cruzi-infected native triatomines occur in the area, but the enzootic transmission of trypanosomatids remains poorly characterized. A parasitological survey involving sylvatic triatomines (166 Rhodnius neglectus collected from Mauritia flexuosa palms) and small mammals (98 marsupials and 70 rodents, totaling 18 species) was conducted in 18 sites (mainly gallery forests and veredas) of the DF. Parasites were isolated, morphologically identified, and characterized by PCR of nuclear (mini-exon gene) and kinetoplast DNA (kDNA). Six R. neglectus, seven Didelphis albiventris and one Akodon cursor were infected by trypanosomes; wild reservoir infection is documented for the first time in the DF. kDNA PCR detected T. cruzi in five R. neglectus and mini exon gene PCR revealed T. cruzi I in isolates from D. albiventris. Parasites infecting one bug yielded T. rangeli KP1+ kDNA amplicons. In spite of the occurrence of T. cruzi-infected D. albiventris (an important wild and peridomestic reservoir) and R. neglectus (a secondary vector displaying synanthropic behavior), a low-risk of human Chagas disease transmission could be expected in the DF, considering the low prevalence infection recorded in this work. The detection of T. rangeli KP1+ associated with R. neglectus in the DF widens the known range of this parasite in Brazil and reinforces the hypothesis of adaptation of T. rangeli populations (KP1+ and KP1-) to distinct evolutionary Rhodnius lineages. PMID- 15654481 TI - Strain differentiation of Trichophyton rubrum by random amplification of polymorphic DNA (RAPD). AB - Trichophyton rubrum is an important cause of dermatomycoses. Molecular strain typing methods have recently been developed to address questions about epidemiology and source of relapse following treatment. This report describes the application of RAPD for molecular strain differentiation of this fungus utilizing the primers 1- (5'-d[GGTGCGGGAA]-3') and 6- (5'-d[CCCGTCAGCA]-3'). A total of five RAPD patterns were observed among 10 strains of T. rubrum, with each of the primers used. We conclude that RAPD analysis using primers 1 and 6 can be used in epidemiological studies. PMID- 15654483 TI - Fetal programming and future disease. PMID- 15654480 TI - Single step polymerase chain reaction (PCR) for the diagnosis of the Leishmania (Viannia) subgenus. AB - In Brazil, the main etiologic agent of Leishmaniasis that frequently presents with mucosal involvement belongs to the Viannia subgenus. The therapeutic conduct in this disease depends on the parasitological diagnosis, and classical methods are restricted in identifying the agent. In this paper we describe a polymerase chain reaction (PCR), which uses primers designed from mini-exons repetitive sequences. The PCR amplifies a 177bp fragment that can distinguish (Viannia) from (Leishmania) subgenus. This test could be a useful diagnostic tool. PMID- 15654484 TI - Retrospective evaluation of bone pain palliation after samarium-153-EDTMP therapy. AB - PURPOSE: The aim of this study was to evaluate the degree of metastatic bone pain palliation and medullar toxicity associated with samarium-153-EDTMP treatment. METHODS: Seventy-three patients with metastatic bone pain having previously undergone therapy with samarium-153-EDTMP (1 mCi/kg) were retrospectively evaluated. Routine follow-up included pain evaluation and blood counts for 2 months after treatment. Pain was evaluated using a subjective scale (from 0 to 10) before and for 8 weeks after the treatment. Blood counts were obtained before treatment and once a week for 2 months during follow-up. Dosimetry, based upon the urinary excretion of the isotope, was estimated in 41 individuals, and the resulting radiation absorbed doses were correlated with hematological data. RESULTS: Reduction in pain scores of 75% to 100% was obtained in 36 patients (49%), with a decrease of 50% to 75%, 25% to 50%, and 0% to 25% in, respectively, 20 (27%), 10 (14%), and 7 (10%) patients. There was no significant relationship between the pain response and location of the primary tumor (breast or prostate cancer). Mild to moderate myelosuppression was noted in 75.3% of patients, usually with hematological recovery at 8 weeks. The mean bone marrow dose was 347 +/- 65 cGy, and only a weak correlation was found between absorbed dose and myelosuppression (Pearson coefficient = .4). CONCLUSIONS: Samarium-153-EDTMP is a valuable method for metastatic bone pain palliation. A mild to moderate and transitory myelosuppression is the main toxicity observed after samarium therapy, showing a weak correlation with dosimetric measures. PMID- 15654485 TI - Hypertension in employees of a University General Hospital. AB - PURPOSE: To find out the prevalence of hypertension in employees of the Hospital and relate it to social demographic variables. METHODS: Blood pressure measurement was performed with a mercury sphygmomanometer, using an appropriate cuff size for arm circumference, weight, and height in a population sample of 864 individuals out of the 9,905 employees of a University General Hospital stratified by gender, age, and job position. RESULTS: Hypertension prevalence was 26% (62% of these reported being aware of their hypertension and 38% were unaware but had systolic/diastolic blood pressures of >140 and/or >90 mm Hg at the moment of the measurement). Of those who were aware of having hypertension, 51% were found to be hypertensive at the moment of the measurement. The prevalence was found to be 17%, 23%, and 29% (P <.05) in physicians, nursing staff, and "others", respectively. The univariate analysis showed a significant odds ratio for the male gender, age >50 years, work unit being the Institute of Radiology and the Administration Building, educational level 10 years, and body mass index >30 kg/m2. The multivariate logistic regression model revealed a statistically significant association of hypertension with the following variables: gender, age, skin color, family income, and body mass index. CONCLUSIONS: Hypertension prevalence was high, mainly in those who were not physicians or members of the nursing staff. High-risk groups (obese, non white, men, low family income) should be better advised of prevention and early diagnosis of hypertension by means of special programs. PMID- 15654486 TI - Adverse life events and coping skills in panic disorder. AB - Research literature and clinical experience shows that panic patients are often able to identify stressors that preceded the onset of their first attacks. In this study we investigated the relation between life events, coping skills, and panic disorder. METHODS: Forty-tree panic patients were compared with 29 control subjects regarding the occurrence and the impact of stressful life events in a 1 year period preceding the onset of panic attacks using the Social Readjustment Rating Scale and London Life Event and Difficulty Schedule. Coping skills were measured using the Ways of Coping Questionnaire. RESULTS: No differences were observed between panic patients and controls regarding the number of reported stressful life events in the previous year. Panic patients compared to controls reported loss of social support as the most meaningful class of events significantly more often. In response to stressful situations, panic patients more often used coping skills judged as ineffective. CONCLUSIONS: The present study suggests that the type of life event and the coping skills used in response to them, more than the occurrence of stressful events itself, may be associated with the onset of panic disorder. PMID- 15654487 TI - Natural history of stenosis in the iliac arteries in patients with intermittent claudication undergoing clinical treatment. AB - PURPOSE: Inspite of the long experience with the treatment of intermittent claudication, little is known about the natural history of stenotic lesions in the iliac segment. With the advent of endovascular treatment, this knowledge has become important. METHODS: Fifty-two stenosis, diagnosed using arteriography, in 38 claudicant patients were analyzed. After a minimum time interval of 6 months, a magnetic resonance angiography was performed to determine whether there was arterial occlusion. The primary factors that could influence the progression of a stenosis were analyzed, such as risk factors (smoking, hypertension, diabetes, sex, and age), compliance with clinical treatment, initial degree of stenosis, site of the stenosis, and length of follow-up. RESULTS: The average length of follow-up was 39 months. From the 52 lesions analyzed, 13 (25%) evolved to occlusion. When occlusion occurred, there was clinical deterioration in 63.2% of cases. This association was statistically significant (P = .002). There was no statistically significant association of the progression of the lesion with the degree or site of stenosis, compliance with treatment, or length of follow-up. Patients who evolved to occlusion were younger (P = .02). The logistic regression model showed that the determinant factors for clinical deterioration were arterial occlusion and noncompliance with clinical treatment. CONCLUSIONS: The progression of a stenosis to occlusion, which occurred in 25% of the cases, caused clinical deterioration. Clinical treatment was important, but it did not forestall the arterial occlusion. Prevention of occlusion could be achieved by early endovascular intervention or with the development of drugs that might stabilize the atherosclerotic plaque. PMID- 15654488 TI - Weight loss and morphometric study of intestinal mucosa in rats after massive intestinal resection: influence of a glutamine-enriched diet. AB - Short-bowel syndrome is responsible for significant metabolic alterations that compromise nutritional status. Glutamine is considered an essential nutrient for enterocytes, so beneficial effects from supplementation of the diet with glutamine are hypothesized. PURPOSE: In this study, the effect of a diet enriched with glutamine was evaluated in rats undergoing extensive small bowel resection, with analysis of postoperative weight loss and intestinal morphometrics of villi height, crypt depth, and thickness of the duodenal and remnant jejunal mucosa. METHODS: Three groups of male Wistar rats were established receiving the following diets: with glutamine, without glutamine, and the standard diet of laboratory ration. All animals underwent an extensive small bowel resection, including the ileocecal valve, leaving a remnant jejunum of only 25 cm from the pylorus that was anastomosed lateral-laterally to the ascendant colon. The animals were weighed at the beginning and end of the experiment (20th postoperative day). Then they were killed and the remnant intestine was removed. Fragments of duodenal and jejunal mucosa were collected from the remnant intestine and submitted to histopathologic exam. The morphometric study of the intestinal mucosa was accomplished using a digital system (KS 300) connected to an optic microscope. Morphometrics included villi height, crypt depth, and the total thickness of intestinal mucosa. RESULTS: The weight loss comparison among the 3 groups showed no significant loss difference. The morphometric studies showed significantly taller duodenal villi in the glutamine group in comparison to the without glutamine group, but not different from the standard diet group. The measurements obtained comparing the 3 groups for villi height, crypt depth, and thickness of the remnant jejunum mucosa were greater in the glutamine enriched diet group than for the without-glutamine diet group, though not significantly different from with standard-diet group. CONCLUSIONS: In rats with experimentally produced short-bowel syndrome, glutamine-enrichment of an isonitrogenous test diet was associated with an improved adaptation response by the intestinal mucosa but not reduced weight loss. However, the adaptation response in the group receiving the glutamine-enriched diet was not improved over that for the group fed regular chow. PMID- 15654489 TI - Bethanecol chloride for treatment of clomipramine-induced orgasmic dysfunction in males. AB - PURPOSE: To investigate whether bethanecol chloride may be an alternative for the clinical management of clomipramine-induced orgasmic dysfunction, reported to occur in up to 96% of male users. METHODS: In this study, 12 fully remitted panic disorder patients, complaining of severe clomipramine-induced ejaculatory delay, were randomly assigned to either bethanecol chloride tablets (20 mg, as needed) or placebo in a randomized, double-blind, placebo-controlled, two-period crossover study. A visual analog scale was used to assess severity of the orgasmic dysfunction. RESULTS: A clear improvement was observed in the active treatment period. No placebo or carry-over effects were observed. CONCLUSION: These findings suggest that bethanecol chloride given 45 minutes before sexual intercourse may be useful for clomipramine-induced orgasmic dysfunction in males. PMID- 15654490 TI - Prognostic factors in locally advanced colon cancer treated by extended resection. AB - The impact of clinical, pathologic, and surgical variables on the postoperative morbidity, mortality, and survival of patients undergoing extended resections of colon carcinoma were evaluated. METHODS: The medical records of 95 patients who underwent extended resections for colon carcinoma between 1953 and 1996 were reviewed. In all cases, in addition to colectomy, 1 or more organs and/or structures were resected en bloc due to a macroscopically based suspicion of tumor invasion. The clinical, pathologic, and surgical parameters were analyzed. Overall survival rates were analyzed according to the method of Kaplan and Meier. Multivariate analysis was performed using the Cox proportional hazards model. RESULTS: Eighty-six patients were treated by curative surgeries and the remaining by palliative resections. Invasion of the organs and/or adjacent structures and regional lymph nodes was found microscopically in 48 and 31 patients, respectively. The median follow-up without postoperative mortality was 47.7 months. The 5-year overall survival rates was 52.6%. The 5-year overall survival rates for patients undergoing curative and palliative surgeries was 58.3% and 0%, respectively. The mean survival time in the palliative surgery group was 3.1 months. Multivariate analysis showed that Karnofsky performance status was strongly related to the risk of postoperative complications (P = .01), and postoperative deaths were associated with the type of surgery and Karnofsky performance status at the time of admission (P = .001). CONCLUSIONS: Some patients with locally advanced colon adenocarcinomas undergoing extended resections have a 5-year overall survival rates of 58.3%. Patients could benefit from palliative-intent procedures, but these measures should cautiously be indicated and avoided in patients with low Karnofsky performance status due to high rates of postoperative mortality and poor survival. PMID- 15654491 TI - Use of nonradioactive labeling to detect large gene rearrangements in 21 hydroxylase deficiency. AB - PURPOSE: To establish the Southern blotting technique using hybridization with a nonradioactive probe to detect large rearrangements of CYP21A2 in a Brazilian cohort with congenital adrenal hyperplasia due to 21-hydroxylase deficiency (CAH 21OH). METHOD: We studied 42 patients, 2 of them related, comprising 80 non related alleles. DNA samples were obtained from peripheral blood, digested by restriction enzyme Taq I, submitted to Southern blotting and hybridized with biotin-labeled probes. RESULTS: This method was shown to be reliable with results similar to the radioactive-labeling method. We found CYP21A2 deletion (2.5%), large gene conversion (8.8%), CYP21AP deletion (3.8%), and CYP21A1P duplication (6.3%). These frequencies were similar to those found in our previous study in which a large number of cases were studied. Good hybridization patterns were achieved with a smaller amount of DNA (5 mug), and fragment signs were observed after 5 minutes to 1 hour of exposure. CONCLUSIONS: We established a non radioactive (biotin) Southern blot/hybridization methodology for CYP21A2 large rearrangements with good results. Despite being more arduous, this technique is faster, requires a smaller amount of DNA, and most importantly, avoids problems with the use of radioactivity. PMID- 15654492 TI - Effects of medical therapy, alcohol, smoking, and endocrine disruptors on male infertility. AB - Infertility affects up to 15% of the sexually active population, and in 50% of cases, a male factor is involved, either as a primary problem or in combination with a problem in the female partner. Because many commonly encountered drugs and medications can have a detrimental effect on male fertility, the medical evaluation should include a discussion regarding the use of recreational and illicit drugs, medications, and other substances that may impair fertility. With the knowledge of which drugs and medications may be detrimental to fertility, it may be possible to modify medication regimens or convince a patient to modify habits to decrease adverse effects on fertility and improve the chances of achieving a successful pregnancy. Concern is growing that male sexual development and reproduction have changed for the worse over the past 30 to 50 years. Although some reports find no changes, others suggest that sperm counts appear to be decreasing and that the incidence of developmental abnormalities such as hypospadias and cryptorchidism appears to be increasing, as is the incidence of testicular cancer. These concerns center around the possibility that our environment is contaminated with chemicals--both natural and synthetic--that can interact with the endocrine system. PMID- 15654493 TI - Human identification and analysis of DNA in bones. AB - The introduction of molecular biology techniques, especially of DNA analysis, for human identification is a recent advance in legal medicine. Substantial effort has continuously been made in an attempt to identify cadavers and human remains after wars, socio-political problems and mass disasters. In addition, because of the social dynamics of large cities, there are always cases of missing people, as well as unidentified cadavers and human remains that are found. In the last few years, there has also been an increase in requests for exhumation of human remains in order to determine genetic relationships in civil suits and court action. The authors provide an extensive review of the literature regarding the use of this new methodology for human identification of ancient or recent bones. PMID- 15654494 TI - Bone tissue stiffness in the mandibular condyle is dependent on the direction and density of the cancellous structure. AB - Variation in the apparent stiffness of cancellous bone is generally ascribed to variation in cancellous structure and density, while the bone tissue stiffness is assumed to be constant. The purpose of the present study was to examine whether the bone tissue stiffness is dependent on the direction and density of the cancellous structure. Bone tissue stiffness was estimated by combining mechanical testing and micro-finite element (micro-FE) modeling on cylindrical bone specimens obtained from the human mandibular condyle. One set of specimens was tested in the vertical direction of the condyle (n = 39) and another set in the transverse direction (n = 30). The cancellous structure of the specimens was characterized by micro-CT. The apparent bone stiffnesses predicted by the FE model correlated strongly (r2 = 0.91) with the measured apparent bone stiffnesses. Apparent bone stiffness in the transverse direction was considerably smaller than that in the vertical direction. In contrast, the predicted bone tissue stiffness was significantly larger in the transverse direction (E = 13.70 GPa) than in the vertical direction (E = 11.87 GPa). In addition, bone tissue stiffness correlated negatively with the bone volume fraction and directional sensitivity of the bone tissue stiffness increased with a decrease of bone volume fraction. The results suggest that the transversely oriented trabeculae in the mandibular condyle are stiffer and more mineralized than the vertically oriented trabeculae and that bone loss is compensated by an increase in the degree of mineralization. PMID- 15654495 TI - Immunolocalization of heat shock protein 27 in developing jaw bones and tooth germs of human fetuses. AB - 27 kDa Heat shock protein (Hsp27), which is also identified as p29 estrogen receptor associated protein, plays a crucial role in specific growth stages. It also seems to be involved in the balance between differentiation and apoptosis. To determine whether Hsp27 is involved during craniofacial development and odontogenesis, its expression was studied through immunohistochemistry of developing jaw bone as well as the odontogenesis of heads from human fetuses. Formalin-fixed paraffin-embedded specimens of 7 human fetuses (3 female, 4 male), obtained from miscarriages occurring between the 9th and 16th weeks of pregnancy, were examined by using a monoclonal antibody against Hsp27. Staining intensity (weak, +; moderate, ++; strong, +++) was evaluated semiquantitatively. The sample slice was cut through a coronal plane, which included eyes, nasal cavities, tongue, and primitive dental lamina with tooth germs. A transient and spatially restricted expression of Hsp27 in developing human jaw bones and teeth was observed. Osteoblasts around the uncalcified bone matrix showed Hsp27 immunoreaction products (+++), whereas osteocytes were not immunolabeled. In mandibular condyle, immunolabeling was restricted to hypertrophic chondrocytes (++). In developing tooth germs, Hsp27 immunostaining was detected throughout the bud (+++). At the early cap stage, a strong immunolabeling for Hsp27 was seen in the dental lamina (+++), and a moderate staining was seen in the outer dental epithelium (++). At the late cap stage, Hsp27 expression was detected in the outer dental epithelium (++) as well as in the cells of the future stellate reticulum (++). The spatiotemporal-restricted expression of Hsp27 in craniofacial bones during development suggests that this protein could be involved in the balance between differentiation and apoptosis, by modulating the viability of osteoblasts and chondrocytes. The specific regional and temporal expression patterns of Hsp27 during tooth development sustains that this small Hsp might be related to the morphogenesis and cytodifferentiation processes of tooth germs. PMID- 15654496 TI - Kinetic study of early regenerative effects of RGTA11, a heparan sulfate mimetic, in rat craniotomy defects. AB - We previously reported that RGTA, a synthetic heparan sulfate mimetic, induces almost complete closure of craniotomy defects one month after surgery in adult rats. RGTA-treated wounds showed features suggesting unusual cell and matrix interactions reminiscent of developmental events. As healing success or failure is determined shortly after wounding, we examined early events in RGTA-treated wounds. Collagen plasters soaked in a solution of RGTA11 (1.5 Microg per piece) or saline (control) were implanted in rat craniotomy defects. Seven control and seven treated rats were killed daily from days 1 to 7 after surgery. The lesions and adjacent tissues were sampled and processed for morphometry. A layer of type III collagen along the dura mater (DM) thickened up to day 5 in RGTA-treated wounds (p < 0.05 vs day 1), but became thinner in control wounds. Alkaline phosphatase-positive osteoprogenitor cells were detected on day 1 in this layer. Their number increased, and they migrated toward the mid-sagittal sinus and to connective tissue adjacent to the sinus, where they aggregated and differentiated into osteoblasts, forming bone nodules on day 6. These features were not seen in control wounds. Angiogenesis was significantly enhanced in RGTA-treated wounds, especially near the sinus. In vitro, bovine bone endothelial (BBE) cell proliferation was inhibited by RGTA11 in a concentration-dependent manner. In contrast, RGTA11 strongly enhanced the effect of fibroblast growth factor-2 on BBE cell proliferation. These results show that RGTA11, possibly by interacting with heparin-binding growth factors, elicits vascular reactions accompanying the recruitment of a large pool of committed osteoprogenitors from the DM. The DM and the sinus appear to be important centers of organization for craniotomy defect healing. RGTA probably creates an environment that starts a program of directing healing towards bone formation and defect closure. PMID- 15654497 TI - Calcitriol improves streptozotocin-induced diabetes and recovers bone mineral density in diabetic rats. AB - Vitamin D analogs exert a preventative effect on experimental diabetes, but whether or not they are able to halt progress of established diabetes is not yet known. Moreover, it is widely accepted that diabetes may induce osteoporosis, but the efficacy of vitamin D on diabetic osteoporosis is not clear. In order to help clarify these issues, we have tested the efficacy of calcitriol streptozotocin induced diabetes. Streptozotocin (60 mg/Kg body weight) was injected in 3-month old Wistar rats, randomly distributed into two groups: vehicle (olive oil) treated diabetic rats (D) and diabetic rats treated with 1.25-(OH)2D3 250 mg, three times a week (DT). Control animals (C) were treated with vehicle alone. The experiment lasted 8 weeks. The histology of the pancreata was evaluated. Blood glucose and calcium and phosphate in serum and urine were measured. Finally, bone mineral density (BMD) of tibia and lumbar vertebrae were evaluated. After 8 weeks, diabetes persisted in 85% of the diabetic rats (D group), but in only 45% of vitamin D-treated group (DT). At the end of the experiment, DT animals were separated into two groups, those still remaining diabetic (DT-NR) and reversed animals (DT-R). Moreover, bone loss was observed in diabetic animals (D), whereas BMD of DT-R rats showed similar values to those of controls (C). Our results suggest that 1.25(OH)2D3 improves diabetes and, as such, may recover BMD in streptozotocin-induced diabetic rats. PMID- 15654499 TI - Management of polypropylene mesh erosion after intravaginal midurethral sling operation for female stress urinary incontinence. AB - The intravaginal midurethral sling operations have become a trend to treat female stress urinary incontinence. Cases of complications requiring surgical revision are rarely reported. We report on seven patients with complications necessitating surgery. Six patients with vaginal erosion were treated with transvaginal excision of migrated tape and fibrotic tissues around the vaginal wall, and one patient with bladder erosion was treated with suprapubic minimal laparotomy and transvaginal partial excision of the tension-free vaginal tape (TVT). The incidence of polypropylene mesh erosion was 1.1% in this study. All patients recovered well from our surgical intervention, except the bladder erosion patient who needed anticholinergic medication to treat postoperative urgency and urge incontinence. PMID- 15654500 TI - Uterine prolapse in a young nulligravida with scleroderma and premature ovarian failure. AB - We report a case of pelvic organ prolapse quantification (POPQ) stage III uterine prolapse in a 25-year-old nulligravida. Premature ovarian failure was diagnosed after 1 year of amenorrhea. Localized scleroderma was noticed on her thigh and lower back. We discuss the possible role of scleroderma and ovarian failure on the occurrence of uterine prolapse in light of the literature. PMID- 15654501 TI - Evolution of the female pelvis and relationships to pelvic organ prolapse. AB - The female pelvis provides support for the lower limbs as well as for the gastrointestinal tract, the bladder, and the reproductive organs. It must also serve as a passageway for defecation, urination, and, possibly, delivery of an infant. The bones, ligaments, and muscles of the human female pelvis have evolved from our early ancestors. Pelvic organ prolapse may occur because of the limitations involved with adapting the pelvic bones, muscles, and ligaments previously used for other purposes into a supportive role. Here we review these changing roles and functions of nonhuman primate and human female anatomy. PMID- 15654504 TI - Molecular variation of Spiranthes sinensis (Orchidaceae) in Japan, with special reference to systematic treatment of seasonally differentiated groups and a dwarf form, f. gracilis, from Yakushima Island. AB - Molecular variations of Spiranthes sinensis Ames var. australis (R.Br.) H. Hara et Kitam. ex Kitam. in Japan were examined to evaluate the validity of the seasonally differentiated groups and a dwarf form of the species, which is endemic to Yakushima Island, Japan. Sequence differences in the plastid trnL-F locus clearly distinguished Japanese S. sinensis var. australis from S. sinensis var. sinensis collected from Ryukyu. In contrast, the trnL-F sequence of S. sinensis var. australis from Sabah, Malaysia, clearly differed from that of Japanese S. sinensis var. australis, suggesting genetic heterogeneity of Spiranthes sinensis var. australis in Asia. Moreover, a molecular analysis based on the sequences of nuclear ITS1 regions indicated that there are two major groups of S. sinensis var. australis in Japan, with a geographic distribution boundary on Kyushu Island. However, the trnL-F and ITS1 sequences did not support the genetic differentiation of the seasonally differentiated groups or the dwarf form from the other Japanese individuals. Based on these molecular data, the systematic treatment of physiological and morphological variations in the Japanese population of S. sinensis. var. australis is discussed. PMID- 15654505 TI - Association of habitual smoking and drinking with single nucleotide polymorphism (SNP) in 40 candidate genes: data from random population-based Japanese samples. AB - Basic information on the association between lifestyle factors and candidate genes is valuable for genetic-environmental study. We screened the association of habitual smoking or drinking with polymorphism in 40 candidate genes for a total of 153 single nucleotide polymorphisms (SNPs) using a sample of 339 middle-aged, randomly selected Japanese men. Smoking and drinking statuses were elicited during questionnaire-based interviews. Genes were selected based on their possible involvement in genetic-environmental, life-style interactions and constitute the genes expressing xenobiotic metabolism enzymes, DNA repair enzymes, and other stress-related proteins. The P values of odds ratios to habitual smoking for CYP17A1, ESR1, EPHX1, GSTT2, ALDH2, NOS2A, OGG1, and SLC6A4 and those of odds ratios to habitual drinking for CYP1B1, ESR1, HSD17B3, GSTM3, COMT, ADH1C, ALDH2, NOS3, and NUDT1 were under 0.05. These variables were included in a stepwise logistic analysis in order to develop a predictive model for smoking or drinking behavior. In the final model, the only significant variables selected for smoking were OGG1, SLC6A4, EPHX1, ESR1, and CYP17A1, and for drinking, ALDH2 and NUDT1. The findings of the present study suggest that polymorphism in associated candidate genes plays a role in the habitual use of tobacco and alcohol among Japanese men. PMID- 15654502 TI - Symptomatic effect of donepezil, rivastigmine, galantamine and memantine on cognitive deficits in the APP23 model. AB - RATIONALE: APP23 mice are a promising model of Alzheimer's disease, expressing several histopathological, cognitive and behavioural hallmarks of the human condition. A valid animal model should respond to therapeutic interventions in an equivalent manner as human patients. OBJECTIVES: To further validate the APP23 model, we examined whether cognitive deficits could be antagonised by donepezil, rivastigmine, galantamine or memantine, which are approved drugs for symptomatic treatment of dementia. METHODS: Animals were tested at an age at which untreated APP23 mice display severe deficits in visual-spatial learning. Four-month-old APP23 mice and control littermates were administered donepezil (0.3 or 0.6 mg kg( 1)), rivastigmine (0.5 or 1.0 mg kg(-1)), galantamine (1.25 or 2.5 mg kg(-1)), memantine (2 or 10 mg kg(-1)) or saline through daily i.p. injections. After 1 week of treatment, acquisition phase commenced, with daily treatment continuing during cognitive testing. RESULTS: All cholinesterase inhibitors reduced cognitive deficits with the following optimal daily doses: galantamine 1.25 mg kg(-1), rivastigmine 0.5 mg kg(-1) and donepezil 0.3 mg kg(-1). Higher dosages often did not exert beneficial effects in accordance with inverted U-shaped dose response curves described for cholinomimetics. Symptomatic efficacy of memantine on cognition was mild, with significant amelioration manifesting during probe trial. CONCLUSIONS: This is the first study to simultaneously evaluate the efficacy of therapeutically relevant doses of these four compounds in one particular learning and memory paradigm, being the Morris water maze. The fact that symptomatic intervention was able to diminish cognitive impairment, substantially adds to the validity of the APP23 model as a valuable tool to evaluate future therapeutic approaches. PMID- 15654503 TI - Identification of jasmonic acid and its methyl ester as gum-inducing factors in tulips. AB - The purpose of this study was to identify endogenous factors that induce gummosis and to show their role in gummosis in tulip (Tulipa gesneriana L. cv. Apeldoorn) stems. Using procedures to detect endogenous factors that induce gum in the stem of tulips, jasmonic acid (JA) and methyl jasmonate (JA-Me) were successfully identified using gas-liquid chromatography-mass spectrometry. Total amounts of JA and JA-Me designated as jasmonates in tulip stems were also estimated at about 70 80 ng/g fresh weight, using deuterium-labeled jasmonates as internal standards. The application of JA and JA-Me as lanolin pastes substantially induced gums in tulip stems with ethylene production. The application of ethephon, an ethylene generating compound, however, induced no gummosis although it slightly affected jasmonate content in tulip stems. These results strongly suggest that JA and JA Me are endogenous factors that induce gummosis in tulip stems. PMID- 15654506 TI - The enhanced virulence of very virulent infectious bursal disease virus is partly determined by its B-segment. AB - There is a remarkable difference in virulence of infectious bursal disease virus (IBDV) strains ranging from sub-clinical infections for serotype 2 and cell culture adapted serotype 1 strains, to 100% mortality for very virulent serotype 1 strains in young SPF chickens. It is known that cell culture adaptation related attenuation is determined by distinct mutations in the hypervariable region of the VP2 outer capsid protein, encoded on the A-segment. Amino acid mutations in the hypervariable VP2 region however, offer no explanation for the difference in virulence of classical and very virulent serotype 1 strains. Here we show by in vitro and in vivo analysis of rescued segment re-asserted IBDVs that virulence factors are not only located on the A-segment, but on the RNA Dependent RNA Polymerase (VP1) encoding B-segment as well. Insight into the virulence factors of very virulent IBDV will contribute to the improvement of live IBDV vaccines. PMID- 15654507 TI - UV-B irradiation stimulates the promoter activity of the high-risk, cutaneous human papillomavirus 5 and 8 in primary keratinocytes. AB - Human papillomaviruses (HPV) have been implicated in the development of non melanoma skin cancer (NMSC). HPV types 5 and 8 are strongly associated with NMSC in patients with the inherited disease Epidermodysplasia verruciformis (Ev). In these patients tumours arise predominantly on sun-exposed skin and consistently harbour HPV DNAs. To determine whether UV-B irradiation modulates the noncoding region (NCR) promoter activity of the Ev-HPV types 5, 8, 9, 14, 23, 24, and 25 we performed transient transfection assays with NCR luciferase reporter gene constructs in primary human epithelial keratinocytes (PHEKs) and in p53-null RTS3b cells. Each of the HPVs showed different basal NCR activity in both cell types and reacted differently upon UVB treatment and p53 cotransfection in RTS3b cells. The NCR of HPV5 and 8 were the only ones to be activated by UV-B in PHEKs. The stimulation of the NCR activity of the high-risk cutaneous HPV types 5 and 8 by UV-B irradiation may point to a role of this interaction in the development of NMSC. PMID- 15654508 TI - Determination of complete nucleotide sequence of Hibiscus latent Singapore virus: evidence for the presence of an internal poly(A) tract. AB - We have sequenced the complete genome of a hibiscus-infecting tobamovirus, Hibiscus latent Singapore virus (HLSV). The experimental host range of HLSV is similar to that of another distinct species of hibiscus infecting tobamovirus, Hibiscus latent Fort Pierce virus (HLFPV). The genomic structure of HLSV is similar to other tobamoviruses in general. It consists of a 5' untranslated region (UTR), followed by ORFs encoding for a 128 kDa protein and a 186 kDa readthrough protein, a 30 kDa movement protein (MP), 18 kDa coat protein (CP) and a 3' UTR. The unique feature of HLSV is the presence of a poly(A) tract within its 3' UTR. In our previous work, we have reported MP and CP sequences of HLSV and its phylogenetic analysis. Here we report the complete nucleotide sequence of HLSV, phylogenetic analysis of the nucleotide and amino acid sequences of 128/186 kDa ORFs and the presence of a uniquely located poly(A) tract within the 3' UTR. PMID- 15654511 TI - Neutrophil recruitment in mast cell-dependent inflammation: inhibitory mechanisms of glucocorticoids. AB - Mast cells are strategically localized along the microvasculature in tissues in close contact with the external environment, such as the skin, lung and intestines. By releasing a multi-faceted spectrum of proinflammatory mediators, such as cytokines and chemokines, mast cells have the capacity to coordinate trafficking of leukocytes. Mast cells play a pathophysiological role in numerous inflammatory diseases as diverse as hypersensitivity reactions, ischemia/reperfusion injury and rheumatoid arthritis. On the other hand, mast cells act also as tissue sentinels and are critically involved in the host defensive response against microbial infection by stimulating neutrophil recruitment. Glucocorticoids are powerful agents frequently used in mast cell dependent diseases, although the anti-inflammatory mechanisms of these compounds are not completely understood at present. In order to circumvent steroid associated side-effects and develop more specific therapeutics, numerous studies have examined the mechanisms underlying glucocorticoid inhibition of mast cell dependent neutrophil recruitment. Based on recent findings, it may be suggested that glucocorticoids selectively inhibit the expression and function of certain adhesion molecules and chemokines. This review summarizes current insights into the underlying mechanisms of mast cell-regulated tissue accumulation of neutrophils and the inhibitory effects of glucocorticoids. PMID- 15654512 TI - Pathways for the bradykinin B1 receptor-mediated diabetic hyperalgesia in mice. AB - OBJECTIVE: Experimental evidence has shown that the bradykinin B1 receptor (BKB1 R) is involved in the development of hyperalgesia associated with diabetes since specific BKB1-R antagonists significantly inhibited the hyperalgesic activity observed in streptozotocin (STZ)-mice in thermal nociceptive tests. MATERIALS AND METHODS: The involvement of the nitric oxide (NO), the substance P (SP) and the calcitonin gene-related peptide (CGRP) pathways in mediating BKB1-R-induced hyperalgesia was evaluated. Diabetes was induced in male CD-1 mice by injecting STZ (200 mg/kg; i.p.). Nociception was assessed using the hot plate and tail immersion tests, one week following the injection of STZ. RESULTS: The nitric oxide synthase (NOS) inhibitors (L-NNA, 20 mg/kg; L-NMMA, 30 mg/kg and AGUA, 50 mg/kg; i.p.), the SP antagonists (sendide and L-732,138, 100 microg/kg; i.v.) and the CGRP antagonist (hCGRP8-37, 100 microg/kg; i.v.) significantly attenuated the hyperalgesic activity and also reversed the potentiating effect of the BKB1- R agonist, DBK on diabetic hyperalgesia in STZ-mice. CONCLUSIONS: These results support the involvement of BKB1-R in the development of diabetic hyperalgesia in STZ-mice through activation of the NO, SP and CGRP pathways. PMID- 15654513 TI - Evidence that arachidonic acid derived from neutrophils and prostaglandin E2 are associated with the induction of acute lung inflammation by lipopolysaccharide of Escherichia coli. AB - OBJECTIVE: The involvement of arachidonic acid (AA) and PGE2 during the E. coli lipopolysaccharide (LPS)-induced acute lung injury was investigated. MATERIAL: Adult male Wistar rats were used. For in vitro studies, rat neutrophils, bronchoalveolar lavage (BAL) fluid, and lug vascular endothelium were used, as described below. TREATMENT: Rats were given an intratracheal injection of LPS (750 microg). METHODS: Total and differential cell counts in BAL fluid; enzyme linked immunoassay (ELISA) analyses of TNF-alpha, IL-1beta, LTB4 and PGE2 in BAL, and immunohistochemical detection of ICAM-1 on lung vascular endothelium were performed six h after LPS challenge. Fatty acid composition of blood neutrophils and plasma was analyzed by HPLC. RESULTS: Rats instilled with LPS presented a sixty three-fold increase in the number of neutrophils in BAL (from 0.5 x 10(6) to 31.5 x 10(6) cells), accompanied by increased levels of TNF-alpha and IL-1beta (p < 0.001), and a three-fold increase in ICAM-1 expression on vascular endothelium. The content of AA in blood neutrophils was reduced by 50%, whereas the level of PGE2 in BAL was increased by 3.5 fold, without changes in the levels of LTB4. CONCLUSIONS: These findings suggest that AA and PGE2 are associated with LPS challenge. PMID- 15654514 TI - Inhibitory effects of budesonide, desloratadine and dexamethasone on cytokine release from human mast cell line (HMC-1). AB - OBJECTIVE: To determine the inhibitory potency of budesonide on interleukin (IL) 4, 6 and 8, GM-CSF and TNF-alpha release from the human mast cell line (HMC-1) in comparison with the systemic glucocorticosteroid, dexamethasone, and H(1) antagonist, desloratadine. METHODS: HMC-1 was stimulated with 25 ng/ml phorbol 12 myristate 13-acetate (PMA) and 2.5 x 10(-7) M ionomycin (A23187) for 6, 12 and 24 h in both the presence and absence of 10(-6)-10(-10) M concentrations of the test drugs. Culture supernatants were collected and assayed by ELISAs. RESULTS: HMC-1 produced substantial amounts of GM-CSF and IL-8 and smaller amounts of TNF alpha, IL-4 and IL-6 after being stimulated with PMA together with A23187. Budesonide and dexamethasone had potent inhibitory effects and desloratadine had modest inhibitory effects on the release of these cytokines. Budesonide was more potent than dexamethasone at most concentrations and time points. IL-4 was the cytokine which was most susceptible to inhibition by the three tested drugs. The inhibitory effects, in some cases, were time- and concentration-dependent. CONCLUSION: Budesonide had a potent inhibitory effect on cytokine release from HMC-1. Its potency was greater than that of both dexamethasone and desloratadine. PMID- 15654515 TI - Involvement of lymphocytes with a Th1 cytokine profile in bone cell damage associated with MMP-9 production in collagen-induced arthritis. AB - OBJECTIVE: To investigate whether lymphocytes could directly cause the destruction of bone cells in collageninduced arthritis (CIA) mice and the mechanism of this destruction. METHODS: Arthritis was induced with chick type II collagen in Kunming mice. The activities of TNF-alpha and IFN-gamma were measured by biological methods. The mRNA of TNF-alpha, IFN-gamma, T-bet, a Th1 specific transcription factor and Fas was detected by reverse transcript-polymerase chain reaction (RT-PCR). Matrix metalloproteinase (MMP)-9 was measured by gelatin zymography assay. The cytotoxicity of spleen cells to bone cell populations and the proliferation of spleen cells against chick type II collagen (CII) were examined by 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2H-tetrazolium bromide (MTT) method. RESULTS: Arthritis was successfully induced by chick CII in Kunming mice, which had remarkable paw swelling and typical features of arthritis. Compared with naive mice, the arthritic mice showed an increased IFN-gamma and TNF-alpha production in serum, and the spleen cells from the arthritic mice displayed a Th1 cytokine profile and directly damaged bone cells in vitro. Interestingly, naive spleen cells treated by phorbol 12-myristate 13-acetate (PMA) also damaged bone cells as did spleen cells from CIA mice. Cyclosporine A blocked the cytotoxicity of spleen cells from CIA mice to bone cells. Only anti CD4 and complement system together inhibited the effect of spleen cells from CIA mice on bone cells. Only the spleen cells from arthritic mice could proliferate against chick CII. In addition, the culture supernatants from spleen cells of arthritic mice contained a higher activity of MMP-9 and production of NO than those of naive mice. PMA elevated the activity of MMP-9 but not the production of NO in naive spleen cells. Moreover, the activities of TNF-alpha, IFN-gamma and MMP-9 but not the production of NO were higher in joint tissues of arthritic mice than those of naive mice. TNF-alpha and IFN-gamma significantly elevated the activity of MMP-9 in spleen cells from both arthritic and naive mice in vitro. Moreover, only anti-MMP-9 but not anti-MMP-2 antibody inhibited the cytotoxicity of spleen cells from CIA mice to bone cells. CONCLUSIONS: The elevated proinflammatory factors such as TNF-alpha and IFN-gamma may promote the MMP-9 activity in local inflammatory cells that are actually involved in the bone cell damage in CII-induced arthritis. PMID- 15654516 TI - Glucosamine, a naturally occurring amino monosaccharide, suppresses the ADP mediated platelet activation in humans. AB - OBJECTIVE: To evaluate the anti-thrombotic action of glucosamine, a naturally occurring amino monosaccharide, platelets were stimulated with ADP in the presence of glucosamine, and its effects on platelet functions were examined. MATERIALS AND METHODS: Human platelet-rich plasma was stimulated with 2.5 microM ADP in the presence of glucosamine (0.01 approximately 1 mM) or other aminosugars (N-acetyl-glucosamine, galactosamine or N-acetyl-galactosamine, 1 mM), and platelet aggregation was monitored. Furthermore, the effects of glucosamine on the thromboxane A2 production, release of granule contents, intracellular calcium mobilization and phosphorylation of Syk (a 72 kD protein tyrosine kinase) were evaluated following ADP-stimulation. In addition, the binding of [3H] ADP to its receptors was examined. RESULTS: Glucosamine (>0.01 mM) dose-dependently suppressed platelet aggregation in response to ADP (p < 0.05), whereas N-acetyl glucosamine, galactosamine or N-acetyl-galactosamine (1 mM) did not affect the ADP-induced platelet aggregation. Furthermore, glucosamine (>0.1 mM) inhibited the extracellular release of granule contents (ATP and platelet factor 4) and production of thromboxane A2 from ADP-stimulated platelets (p < 0.05). Moreover, glucosamine significantly repressed the intracellular calcium mobilization at >0.1 mM and phosphorylation of Syk at >0.01 mM upon ADP-stimulation (p < 0.05). In addition, glucosamine (>0.1 mM) inhibited the binding of ADP to its receptors (p < 0.05). CONCLUSION: Glucosamine is able to suppress platelet aggregation, release of granule constituents, thromboxane A2 production, calcium mobilization and phosphorylation of Syk possibly via the inhibition of ADP-binding to the receptors. Glucosamine could be expected as a novel anti-platelet agent for thrombotic disorders due to its suppressive actions on platelets. PMID- 15654517 TI - Differential expression of matrix metalloproteinases and cyclooxygenases in synovial cells exposed to Borrelia burgdorferi. AB - OBJECTIVE AND DESIGN: Lyme arthritis is characterized by strong inflammatory reactions probably due to the presence of Borrelia burgdorferi in the joint. It has been suggested that Borrelia adopts different molecular mechanisms that either can amplify the host's inflammatory response or can suppress it. In the present study we analyzed the induction of matrix metalloproteinases (MMPs) and cyclooxygenases (COXs) in human synoviocytes exposed to different B. burgdorferi sensu stricto isolates (Geho and B31). MATERIALS AND METHODS: Synoviocytes were exposed in vitro for 12 h up to 5 days. Semiquantitative reverse transcription polymerase chain reaction was used to assess the mRNA expression of MMP-1 to 13, COX-1 and COX-2. Prostaglandin E2 (PGE2) production was assessed by ELISA. RESULTS: MMP-1 was unchanged in synovial cells exposed to strain Geho, whereas it was downregulated by strain B31. MMP-13 was downregulated by both strains. COX-2 was upregulated by strain B31, which resulted in increased PGE2 concentration in the supernatant. In contrast, COX-1 was slightly upregulated and COX-2 tended to be downregulated by Geho resulting in a decreased PGE2 concentration. CONCLUSIONS: The differential expression of MMPs and COXs suggests that different B. burgdorferi strains influence different molecular mechanisms leading to chronic inflammation. This might be reflected in the clinical variability among Lyme arthritis patients. PMID- 15654518 TI - Sickness behavior of rats with abdominal sepsis can be improved by antibiotic and G-CSF prophylaxis in clinic modeling randomized trials. AB - BACKGROUND AND AIM: In clinical sepsis research nearly all immune-modulators have demonstrated no benefit in regard to the 28-day mortality rate. Other endpoints such as quality of life have become more attractive, but clinically relevant animal models analyzing an equivalent to quality of life by measurement of sickness behavior are extremely rare. The concept of clinic modeling randomized trials was used in an animal trial to model clinical complexity and conditions of a randomized clinical trial. METHODS: 80 adult male Wistar rats were randomly assigned to (1) control: anesthesia and sham operation, (2) sepsis: laparotomy and peritoneal infection with human stool bacteria, (3) sepsis with antibiotic prophylaxis: cefuroxime/metronidazole and (4) sepsis with antibiotic plus a cytokine prophylaxis with granulocyte-colony stimulating factor (GCSF). Endpoints were physiological and behavioral parameters. RESULTS: The combination of antibiotics plus G-CSF was most effective in reducing mortality. All infected animals showed reduced open field activity acutely after infection, and recovery was improved during the 9 day follow-up in rats with prophylactic treatments. In the social interaction test, but not in the elevated plus-maze anxiety test, prophylaxis was also efficient, especially with antibiotics and G-CSF. CONCLUSIONS: The results show that improving sickness behavior in septic rats with G-CSF plus antibiotics may be a promising approach. PMID- 15654519 TI - Screening for colorectal cancer: implications for UK and European initiatives. AB - Colorectal cancer (CRC) is the third most common cancer after lung and prostate cancers, and is the second most common cancer in women in the United Kingdom (UK) after breast cancer. The disease is well suited for prevention with screening programs. CRC is invariably fatal when diagnosed at an advanced stage but curable when diagnosed early, since it passes through a detectable asymptomatic stage. Screening tests with varying degrees of sensitivity and specificity are available. Prior to implementation of a nationwide national programme in the UK, trials are underway to identify the most sensitive and specific screening modality, and to address patient acceptability and resource implications. Screening for CRC using faecal occult blood (FOB) tests may be feasible. There is evidence to show that this modality saves lives at a cost similar to the breast screening programme currently underway in the UK. One-off flexible sigmoidoscopy is an alternative to FOB screening, and pilot data suggest that the technique is logistically feasible in the UK; a multicentre randomised trial is currently underway. Total colonoscopy is the gold standard test, but at present no large randomised controlled trials support this strategy. Colonoscopy is expensive and labour-intensive, and training issues need to be addressed. At the present time, tailored screening (in which intensity is adjusted for individual risk) appears reasonable. FOB and flexible sigmoidoscopy are cost effective but vary in sensitivity and specificity. Public education programmes are essential for any screening strategy to be effective. PMID- 15654520 TI - Colonic motility studies in severe chronic constipation: an organic approach to a functional problem. AB - Chronic constipation is one of the more frequent functional gastrointestinal symptoms. The more severe cases not infrequently undergo a major surgical approach. However, there is still a need for a better diagnostic and therapeutic approach, since the results are still unpredictable, probably due to a poor selection of candidates. This article focuses on the current diagnostic approach to chronic constipation, with an emphasis on the possible role of colonic manometry to help better choose those patients that may benefit from surgery. PMID- 15654521 TI - Different anal pressure profiles in patients with anal fissure. AB - BACKGROUND: A consistent debate exists about the association between anal fissure and hypertonic anal canal. The aim of this study was to determine if the manometric findings in patients with chronic anal fissures varied according to the topography of the fissure. PATIENTS AND METHODS: Seventy-three outpatients (52 men, 71%) with chronic anal fissures and nine healthy volunteers (5 men, 55%) were examined. Patients were classified according to the topography of the anal fissures: posterior midline (group A), anterior midline (group B), and lateral position (group C). We use computerized anorectal manometry to evaluate anal resting pressure, maximal voluntary contraction, recto-anal inhibitory reflex, rectal sensations and rectum compliance. RESULTS: In Group A, the mean pressure was higher than that of controls (p<0.05), and the resting pressure 2 cm from anal verge was higher than that of other groups and controls (p<0.05). Normotonic anal canal was found in 49.1% of patients in group A, in 66% of those in group B and in 57.1% of those in group C. Four elderly patients (7%) of group A had a hypotonic anal canal. No differences were found regarding maximal voluntary contraction, recto-anal inhibitory reflex, rectal sensations and rectum compliance between patients and controls. CONCLUSIONS: Patients with chronic anal fissures may have several anal pressure profiles. The anal canal is often normotonic. Fissures with hypertonic or normo-hypotonic anal canal need different therapies. PMID- 15654522 TI - Elastic one-stage cutting seton for the treatment of high anal fistulas: preliminary results. AB - BACKGROUND: The management of high and complicated anal fistulas remains a therapeutic challenge, and the oldest and theoretically the simplest technique is to use a seton. In this article, we document our recent experience in managing high anal fistulas with a simple modification of the cutting seton. PATIENTS AND METHODS: Surgical outcomes of patients treated by the elastic cutting seton for anal fistulas that involved more than half of the sphincter muscles or for anterior trans-sphincteric fistulas in female patients were analyzed. The elastic seton, which was created by cutting a thin (2-3 mm) circular strip from a surgical glove, including its thicker sleeve, was inserted through the remaining tract in a double-strand fashion, and then tied on the sphincter with two knots in a slightly tight manner. RESULTS: Complete healing was achieved in 9 cases (45%) at 1 month and in all cases (100%) at 3 months postoperatively. Recurrent fistula was noted in a single patient (5%) at 8 months. Worsening of preoperative continence was noted in 4 patients (20%). However, the postoperative incontinence score (0.70+/-1.22) did not differ significantly from the preoperative score (0.41+/-0.41; p=0.059, Wilcoxon's test). CONCLUSIONS: The preliminary results of this series suggest that the elastic cutting seton may be a valid alternative for the treatment of high anal fistulas. The possible positive contribution of the slow and stable cutting of the sphincter on the maintenance of continence is further supported. An important practical disadvantage of the conventional seton treatment, namely the need for postoperative adjustments, is also eliminated. PMID- 15654523 TI - A comparative study between radiofrequency ablation with plication and Milligan Morgan hemorrhoidectomy for grade III hemorrhoids. AB - BACKGROUND: Milligan-Morgan (MM) hemorrhoidectomy is the most favored treatment for prolapsed hemorrhoids. However, it may be associated with severe postoperative pain, long periods of convalescence and other complications. In alternative, I use a procedure of radiofrequency ablation and plication (RAP) of hemorrhoids. The present study compared the two procedures in terms of surgical parameters, postoperative pain and complications. PATIENTS AND METHODS: A total of 60 patients with grade III hemorrhoids were randomized to undergo radiofrequency ablation and plication (31 patients) or MM hemorrhoidectomy (29 patients). The patients were followed up to 2 years. RESULTS: Duration of surgery was significantly longer in the MM group as was postoperative hospitalization (p<0.05). Post-defecation pain and pain at rest were much less in the RAP group (p<0.05). Wound healing period (17 vs. 38 days) and time to return to work (7 vs. 17 days) were the other significant findings favoring RAP procedure. Early complications occurred more frequently in MM group, but late complications like external skin tags (4 vs. 2 patients) were more common in RAP group. One asymptomatic recurrence was noted in RAP group. CONCLUSIONS: Radiofrequency ablation and plication of hemorrhoids is associated with significantly less postoperative pain, shorter hospital stay and earlier return to normal activity. It can be considered as an alternative to the Milligan-Morgan hemorrhoidectomy. PMID- 15654524 TI - Which colorectal cancers are missed by double contrast barium enema? AB - BACKGROUND: The relative merits of either barium enema or colonoscopy for investigating lower gastrointestinal tract symptoms is still unclear. We studied the value of double contrast barium enema (DCBE) as the initial evaluation modality. We reviewed our 10-year experience of double contrast enemas as read by consultant radiologists. The study also aimed to identify which lesions are usually missed. PATIENTS AND METHODS: We reviewed clinical data for all patients who underwent DCBE within the 6 months prior to surgical resection of colorectal cancer between April 1989 and April 1999. Patient demographics and tumour characteristics were analysed for their effects on the likelihood of the lesions being missed at DCBE. RESULTS: There were 706 patients included in the study, 54.2% were male and the mean age was 63.7 years (SEM=0.5 years). The site along the colon and rectum of tumours missed by DCBE corresponded with the frequency of tumour occurrence at each site. The overall rate of missed lesions was 4.1% (29 of 706 patients); these patients were found on subsequent endoscopy to harbour cancer. Tumours less than 3 cm in length and with lesser extent of circumferential involvement were more likely missed at DCBE (p=0.05 and p=0.01, respectively). Age, sex, and tumour grade and stage were not significant predictors of the likelihood of missed lesions. Of the 29 patients with missed lesions, 77.2% had a serum concentration of carcinoembryonic antigen (CEA) above the normal range (3.5 microg/l). The mean follow-up was 65.3 months (SEM=1.8 months). The overall survival for this series was 60.1%. The inaccuracy of the initial DCBE was not found to cause statistically significant differences in the stage of the tumour at diagnosis nor the overall survival of the patients in our series. CONCLUSIONS: Smaller cancers without circumferential involvement may be missed when DCBE is performed to evaluate lower gastrointestinal symptoms. Further evaluation by colonoscopy must be recommended when symptoms persist, especially in the context of a raised CEA level. PMID- 15654525 TI - High-magnification chromoscopic pouchoscopy: a novel in vivo technique for surveillance of the anal transition zone and columnar cuff following ileal pouch anal anastomosis. AB - BACKGROUND: Persistence of underlying disease in the residual rectal mucosa and anal transition zone (ATZ) following ileal pouch-anal anastomosis (IPAA) for ulcerative colitis and familial adenomatous polyposis provides a site for potential malignancy. For this reason endoscopic surveillance is performed, although conventional assessment may be unreliable. We hypothesized that the novel technique of high-magnification chromoscopic pouchoscopy (HMCP) may permit accurate anatomical localization of this high risk zone in vivo and permit improved biopsy accuracy. PATIENTS AND METHODS: We studied 132 patients with IPAA using HMCP. Three distinct zones were defined using magnification endoscopy: ATZ, appearing as a linear cellular marix; columnar cuff, identifiable by a type I crypt pattern; and ileal pouch body, appearing as villous projections. Quadrantic biopsies of these zones were taken in addition to biopsies of any other lesions noted. RESULTS: A total of 1586 biopsies were taken from zones 1-3 (median, 12; range, 5-16 per patient). Overall biopsy-targeting accuracies using magnification guidance as compared with histopathology were 82%, 73% and 91% for the ATZ, cuff and pouch body, respectively. No dysplasia was identified in the quadrantic surveillance biopsies. Histologically confirmed columnar metaplasia was visualized in vivo using magnification chromoscopy. Patients with IPAA >3 years' duration were more likely to have pouch reservoir columnar metaplasia as compared to those <3 years (p<0.01). Pouch reservoir metaplasia was associated with a pre morbid diagnosis of high-grade dysplasia or carcinoma within the premorbid colectomy specimen (p<0.001). CONCLUSIONS: This is the first study to evaluate this novel application of high magnification chromoscopy. Magnification pouchoscopy is a valid predictor of ATZ and cuff anatomy, permitting accurate biopsy targeting. Further randomized studies validating this technique with an emphasis on dysplasia detection in larger cohorts are required. PMID- 15654526 TI - Haemangiopericytoma of the sigmoid mesentery. AB - Haemangiopericytoma is a rare, vascular soft tissue tumour originating from the pericytes surrounding capillaries. We report a case of haemangiopericytoma in the sigmoid mesocolon and are aware of only one previously case. A 61-year-old man was referred with a left iliac fossa mass. At operation, a 10-cm diameter mass was found to be arising from the sigmoid mesentery (Fig. 1). The mass did not involve the bowel wall and there was no clinical evidence of metastatic disease. A sigmoid colectomy with primary anastomosis was performed. The patient made an uneventful recovery. Pathological assessment of the specimen revealed a 95 x 70 x 50 mm(3), purple, lobulated mass within the sigmoid mesocolon adjacent to the bowel. Immunohistological analysis (positive CD34, focal factor VIII) was consistent with a diagnosis of a haemangiopericytoma. Complete excision with adequate margins remains the treatment of choice. We therefore suggest that patients be carefully followed for long periods and advised of the risk of long term relapse. PMID- 15654527 TI - Irreversible lower limb ischaemia following ligation of the inferior mesenteric artery in the surgical treatment of rectal cancer. AB - Rectal cancer and cardiovascular disease are both commoner in the elderly and may coexist. In some severe arteriopaths the blood supply to the lower limbs may be a collateral circulation from the inferior mesenteric artery. Patients with aorto iliac occlusion or severe stenosis may have collaterals from the inferior mesenteric artery to the lower limb blood vessels. Ligation of the inferior mesenteric artery in treating rectal cancer can result in irreversible ischaemia as outlined in this report. Routine palpation of the femoral pulses and awareness of collateral circulation may avoid the disastrous consequences seen in the two cases described. PMID- 15654529 TI - An uncomplicated and safe way to perform on-table colonic lavage. AB - When surgery is performed in patients with acute or sub-acute left-colonic obstruction, the method of antegrade on-table colonic lavage is well established. In this short note, a modification of the original procedure is presented that makes it even more uncomplicated. PMID- 15654528 TI - Pneumomediastinum after dilatation of anal stricture following stapled hemorrhoidopexy. AB - Stapling procedure is a new technique for the surgical management of prolapsing haemorrhoids. Some articles have reported severe adverse effects of this operation. We describe a case of an excessive staple-line stenosis followed stapled haemorrhoidopexy. Proctoscopic dilatation resulted in complications of retropneumoperitoneum, pneumomediastinum, subcutaneous emphysema and perianal abscess. Drainage of the abscess was performed, allowing quick recovery. After discharge from the hospital, the patient continued to perform periodic dilatation. Simple proctoscopic dilatation was conducted in an outpatient setting. PMID- 15654530 TI - Rectovaginal septal repair: case presentations and introduction of a modified reconstruction technique. AB - Rectovaginal fistula are a relatively rare kind of anorectal fistulas. Spontaneous healing is rare and the rectal advancement flap repair is the most popular procedure with success rates ranging between 60% and 80%. We present a new technique for repairing damage in the rectovaginal septum that consists of placing a folded polyglycolic acid mesh (Dexon) between the levator ani muscle closure area and the vaginal wall. This absorbable mesh separates the suture lines on the vaginal and rectal walls, and induces fibrosis and healing. The technique was performed in four women suffering from a rectovaginal fistula due to different causes. It was successful in all cases. PMID- 15654532 TI - Treatment of grade III anal intraepithelial neoplasia with photodynamic therapy: report of a case. Dis Colon Rectum, 2003; 46(11):1555-1559. PMID- 15654533 TI - An injection technique for the treatment of idiopathic pruritus ani. PMID- 15654535 TI - Surgical treatment of anal atresia with vestibular anus in an adult woman. PMID- 15654536 TI - [Sonographic findings in the diagnostic course of vascular access for hemodialysis]. AB - BACKGROUND AND PURPOSE: Hemodialysis and related procedures require a reliable vascular access and the Brescia-Cimino shunt of the nondominant forearm has proven to be its most important form. Today, color Doppler sonography is regarded as the main method in the diagnosis of vascular access dysfunction. The aim of this study was to evaluate the impact of the Resistive Index (RI) of the fistula's feeding artery on the fistula's blood flow. PATIENTS AND METHODS: 47 patients with arteriovenous fistulas were investigated by means of color Doppler sonography. The fistulas' blood flow, morphological details and the RI of the arteries distal as well as proximal to the fistulas were determined. Furthermore, systolic blood pressure of the second and third fingers of both hands was measured. Comparisons were made between the fistula's blood flow and the RI of the feeding artery. RESULTS: On the arm of the fistulas, the proximal and distal RI and the systolic blood pressure were lower than on the contralateral arm. The blood flow of the upper arm fistulas was higher than that of the forearm fistulas due to a higher vascular diameter. No linear correlations existed between the RI and the fistula's blood flow. 50% of the forearm fistulas showed a proximal RI > or = 0.53 and a blood flow < 400 ml/min. CONCLUSION: The RI of the supplying artery of the forearm fistulas is a reliable value to detect insufficient blood flow. In such cases, further diagnostic procedures are necessary. Also, numerous influences on the measurement of the fistula's blood flow exist. Therefore, intraindividual investigations to evaluate the fistulas' further development are recommended to get better options for their control. PMID- 15654537 TI - Influenza vaccination coverage rates in Germany a population-based cross sectional analysis of the seasons 2002/2003 and 2003/2004. AB - BACKGROUND AND PURPOSE: Influenza continues to be a considerable health problem in Europe. Vaccination is the only preventive measure, reducing mortality and morbidity of influenza in all age groups. The objectives of this survey were to assess the level of influenza vaccination coverage in the 2002/2003 season compared with the 2003/2004 season in Germany, to understand the driving forces and barriers to vaccination, and to determine vaccination intentions for the following winter. METHODS: The authors conducted a random-sampling, telephone based household survey among noninstitutionalized individuals representative of the population aged > or = 14. The surveys for 2002/2003 and 2003/2004 used the same questionnaire and were subsequently pooled. Four target groups were determined for analysis: (1) persons aged > or = 60; (2) people working in the medical field; (3) persons suffering from chronic illness; and (4) a group composed of persons aged > or = 60 or working in the medical field or suffering from a chronic illness. RESULTS: The overall sample consisted of 4,011 people. The influenza vaccination coverage rate in Germany increased from 22.3% in 2002/2003 to 25.1% in 2003/2004. This increase is not significant. The most frequent reasons for being vaccinated given by vaccinees were: influenza considered to be a serious illness, which people wanted to avoid (90.1%), having received advice from the family doctor or nurse to be vaccinated (71.3%), and not wanting to infect family and friends (70.4%). Reasons for not being vaccinated mentioned by people who have never been vaccinated were: thinking about it, however, not being vaccinated in the end (47.7%), not expecting to catch influenza (43.6%), and not having received a recommendation from the family doctor to be vaccinated (36.6%). Options encouraging influenza vaccination are: recommendation by the family doctor or nurse (66.6%), more available information on the vaccine regarding efficacy and tolerance (54.2%), and more information available about the disease (52.4%). CONCLUSION: The vaccination coverage in the 2003/2004 season increased in comparison to the 2002/2003 season, although not significantly. The family doctor is the most important source of encouragement for people to be vaccinated against influenza. It seems that the public would be more likely to be vaccinated if they had more information on the efficacy and tolerance of the vaccine, as well as the disease. It is therefore suggested that family doctors be better informed on influenza vaccine and the disease itself, so that they can actively inform their patients on these topics. PMID- 15654538 TI - [Monoclonal antibody therapy for malignant lymphoma]. AB - The treatment options for patients with malignant lymphoma have been substantially enriched by the development of B-cell-specific monoclonal antibodies. One of the reasons for the attractiveness of this approach is the different mode of action of these antibodies compared to chemotherapy: they can exert tumor-suppressive effects by at least three major mechanisms: an intrinsic cytotoxic activity, antibody-dependent cellular cytotoxicity (ADCC), and activation of complement-dependent cytolysis (CDC). These monoclonal antibodies can be applied in an unconjugated form or as a carrier of cytotoxic drugs or radioisotopes. The chimeric anti-CD20 antibody rituximab has a direct anti lymphoma activity, and is highly active in indolent and aggressive lymphoma, in particular in combination with chemotherapy. The anti-CD52 antibody alemtuzumab is effective in the treatment of patients with chronic lymphocytic leukemia (CLL). Another attractive approach is to link anti-CD20 antibodies to radioisotopes, thereby exploiting the radiosensitivity of malignant lymphomas: encouraging results were already presented for the yttrium-90-((90)Y-)labeled anti-CD20 antibody ibritumomab tiuxetan as well as for the iodine-131-((131)I )labeled anti-CD20 antibody tositumomab. PMID- 15654539 TI - [Health care services research]. PMID- 15654540 TI - [DRG Introduction in Germany. Options for action of hospitals]. AB - BACKGROUND AND PURPOSE: The introduction of DRGs in Germany in 2003 enabled the establishment of a compensation system, which combines quality of services and economic viability. This new system leads to an increase in competition among clinics and insurance providers to act as economic as possible. Various options exist for individual hospitals to increase economic viability; these actions, however, need to be identified, categorized and prioritized to enable hospitals to act accordingly. The goal of this analysis is to identify possible actions hospitals can take as a result of the introduction of G-DRGs. METHOD: These actions will be divided into strategic and organizational-operational actions, which have a direct implication on the DRG system or conterminal areas. The strategic actions should provide a first step toward developing a comprehensive system under which individual hospitals can determine the best set of actions to reach their respective goals. The organizational-operative dimension then focuses on the concrete individual actions a hospital can take to reach that goal. RESULTS UND CONCLUSION: As a result, this analysis reveals that hospitals should increasingly attempt to find additional sources of income outside the closed ended sectoral budgets of the public health insurance sector in Germany. Further, hospitals should increasingly focus on optimizing their cost structure according to standard optimization approaches to increase their ability to provide quality services at competitive costs. Finding the right size for each hospital under the new DRG incentivation and compensation structure has become one of the most critical strategic questions each hospital has to answer individually. PMID- 15654541 TI - [Ill tropical traveler returning home]. PMID- 15654542 TI - [S3 guideline by the German Society of Digestive and Metabolic Diseases and the Competence Network of Chronic Inflammatory Bowel diseases on diagnosis and therapy of ulcerative colitis. An update]. PMID- 15654544 TI - [From bench to bedside]. PMID- 15654543 TI - [2005 updates in German coding guidelines and in the total case sum catalogue: cardiology]. PMID- 15654545 TI - [Quantification of minimal residual disease by multiparameter flow cytometry in acute myeloid leukemia. From diagnosis to prognosis]. AB - Today, multiparameter flow cytometry allows the simultaneous assessment of up to five surface and cytoplasmic antigens in different cell populations from peripheral blood and bone marrow samples. Due to the high efficiency of modern flow cytometers a comprehensive characterization is achieved within a short time. In the setting of diagnosing acute myeloid leukemias (AML) this method is used not only to subclassify AML and separate it from acute lymphoblastic and biphenotypic leukemias but also for the identification of leukemia-associated aberrant immunophenotypes (LAIPs). Since these LAIPs are defined individually for each patient, leukemic bone marrow cells can be detected during the course of treatment using the LAIP allowing the quantification of minimal residual disease (MRD) which is not detectable by cytomorphology. Due to its close correlation with the course of the disease and with the risk of relapse the MRD represents an important prognostic parameter which is increasingly used for stratification of therapy in clinical trials. PMID- 15654546 TI - [Magnetic resonance imaging: technical aspects and recent developments]. AB - Magnetic resonance imaging (MR) is based on electromagnetic effects of rotating protons in hydrogen of water and organic materials. With a magnetic field and high frequency electromagnetic pulses, MRI generates data sets to be reconstructed as two-dimensional cross-sectional images or three-dimensional volumes of anatomic structures with excellent soft-tissue contrast. Since modern MR techniques have dramatically reduced acquisition times and motion artifacts, breathhold studies of moving organs such as the stomach and intestinum or heart and lung have become part of the daily routine. MRI does not apply ionizing radiation, therefore dynamic studies and repeated controls may be performed. Original contraindications such as metallic implants or electronic devices (pacemakers) have become relative in many cases. Since MR contrast media are considered not nephrotoxic when applied in regular dose, MRI provides a most valuable alter native for imaging studies and angiography in patients with renal insufficiency. The recent developments in the field of internal medicine include functional imaging of stomach, intestinum, heart and lung. Based on these experiences, protocols for whole-body MRI within reasonable acquisition time and at acceptable costs have been developed. In particular for screening and staging, such MRI proto cols might cover an even broader spectrum of applications in the near future. Based on the knowledge of recent trends and technical backgrounds, many perspectives are opened up for the further development of the method in interdisciplinary cooperation. PMID- 15654547 TI - [Massive ascites of unknown origin. Ovarian tumor]. PMID- 15654548 TI - Deja vu in neurology. AB - The significance of deja vu is widely recognised in the context of temporal lobe epilepsy, and enquiry about deja vu is frequently made in the clinical assessment of patients with possible epilepsy. Deja vu has also been associated with several psychiatric disorders. The historical context of current understanding of deja vu is discussed. The literature reveals deja vu to be a common phenomenon consistent with normality. Several authors have suggested the existence of a "pathological" form of deja vu that differs, qualitatively or quantitatively, from "non pathological" deja vu. The features of deja vu suggesting neurological or psychiatric pathology are discussed. Several neuroanatomical and psychological models of the deja vu experience are highlighted, implicating the perceptual, mnemonic and affective regions of the lateral temporal cortex, hippocampus and amygdala in the genesis of deja vu. A possible genetic basis for a neurochemical model of deja vu is discussed. Clinical approaches to the patient presenting with possible deja vu are proposed. PMID- 15654549 TI - Comparative tolerance of IFN beta-1a regimens in patients with relapsing multiple sclerosis. The EVIDENCE study. AB - The EVIDENCE study was a direct comparative study of two dose regimens of interferon (IFN) beta-1a used in the treatment of relapsing-remitting multiple sclerosis (RRMS): 30 mcg intramuscularly once weekly (qw; n=338) and 44 mcg subcutaneously three times weekly (tiw; n=339). The study continued for an average of 64 weeks. The safety population consisted of all patients receiving at least one dose of study drug. Clinical assessments occurred every 4 weeks for 24 weeks and then every 12 weeks. Blood tests for safety were taken at baseline and at weeks 4 and 12, and every 12 weeks thereafter. Overall adverse events were more common with the 44 mcg tiw regimen (p=0.007), and were due predominantly to differences in injection-site reactions. The majority of adverse events were rated mild by investigators. Hepatic and haematological adverse events and asymptomatic laboratory abnormalities were more common with 44 mcg tiw (p<0.001),with no difference seen for severe events. Flu-like symptoms were more common with 30 mcg qw (p=0.031), were more severe and persisted for longer. Serious adverse events were comparable for both groups, as were drug discontinuations. In conclusion, although adverse events were more common with high-dose, high-frequency IFN therapy, differences were primarily for mild events and did not affect treatment adherence. Based on superior clinical and magnetic resonance imaging outcomes over an average of 64 weeks, coupled with modest safety differences, the risk-benefit ratio for IFN therapy in RRMS favours the 44 mcg tiw regimen over this period of time. PMID- 15654550 TI - Secondary prevention after ischemic stroke. Evolution over time in practice. AB - BACKGROUND AND OBJECTIVE: An optimal management of vascular risk factors, associated with antithrombotic drugs and carotid surgery when appropriate, reduces the risk of a new vascular event after stroke. Although secondary prevention is not optimal in many patients in practice, the question of whether there is an improvement over time remains unanswered. The aim of our study was to test the hypothesis that secondary prevention measures after cerebral ischemia improve over time. METHOD: We included 123 consecutive patients in 1994, and 125 consecutive patients in 2002, who were admitted to a neurological department for any reason and had had an episode of cerebral ischemia less than 6 years earlier. We compared the groups for the management of arterial hypertension, hypercholesterolemia, diabetes mellitus and smoking. We recorded the values of blood pressure, biological parameters, and presence of antithrombotic therapy, lipid-lowering and anti-hypertensive drugs. Whether patients were properly treated or not, was determined by a comparison between their current treatment and guidelines available when recruited. RESULTS: Prevention was not optimal in 96 of 123 (78%) patients in 1994, and in 77 of 125 (62 %) in 2002. Vascular risk factors were better identified and managed in 2002 than in 1994, especially for hypercholesterolemia. Antithrombotic therapies, statins and antihypertensive drugs, except calcium channel blockers, were more often used in 2002. The proportion of patients in whom arterial hypertension and hypercholesterolemia were identified was higher in 2002, but the proportion of patients identified as diabetics remained stable. However, the proportion of patients with blood pressure >140/90 mmHg, glycemia >or = 126 mg/dl, total cholesterol level > or = 240 mg/dl, or being current smokers, were significantly lower in 2002 than in 1994. CONCLUSION: Although most of patients with previous cerebral ischemia did not receive an optimal management of their risk factors in 2002, there was an improvement over an 8-year period. PMID- 15654551 TI - Uncrossed cortico-muscular projections in humans are abundant to facial muscles of the upper and lower face, but may differ between sexes. AB - It is a popular concept in clinical neurology that muscles of the lower face receive predominantly crossed cortico-bulbar motor input, whereas muscles of the upper face receive additional ipsilateral, uncrossed input. To test this notion, we used focal transcranial magnetic brain stimulation to quantify crossed and uncrossed cortico-muscular projections to 6 different facial muscles (right and left Mm. frontalis, nasalis, and orbicularis oris) in 36 healthy right-handed volunteers (15 men, 21 women, mean age 25 years). Uncrossed input was present in 78% to 92% of the 6 examined muscles. The mean uncrossed: crossed response amplitude ratios were 0.74/0.65 in right/left frontalis, 0.73/0.59 in nasalis, and 0.54/0.71 in orbicularis oris; ANOVA p>0.05). Judged by the sizes of motor evoked potentials, the cortical representation of the 3 muscles was similar. The amount of uncrossed projections was different between men and women, since men had stronger left-to-left projections and women stronger right-to-right projections. We conclude that the amount of uncrossed pyramidal projections is not different for muscles of the upper from those of the lower face. The clinical observation that frontal muscles are often spared in central facial palsies must, therefore, be explained differently. Moreover, gender specific lateralization phenomena may not only be present for higher level behavioural functions, but may also affect simple systems on a lower level of motor hierarchy. PMID- 15654553 TI - Memory evaluation with a new cued recall test in patients with mild cognitive impairment and Alzheimer's disease. AB - Free delayed recall is considered the memory measure with the greatest sensitivity for the early diagnosis of dementia. However, its specificity for dementia could be lower, as deficits other than those of pure memory might account for poor performance in this difficult and effortful task. Cued recall is supposed to allow a better distinction between poor memory due to concurrent factors and impairments related to the neurodegenerative process. The available cued recall tests suffer from a ceiling effect. This is a prospective, longitudinal study aiming to assess the utility of a new memory test based on cued recall that avoids the ceiling effect in the early diagnosis of Alzheimer's disease (AD). Twenty-five patients with mild cognitive impairment (MCI), 22 probable AD patients (NINCDS-ADRDA) at a mild stage, 22 elderly patients with subjective memory complaints (SMC) and 38 normal age-matched controls took part in the study. The patients underwent a thorough cognitive evaluation and the recommended screening procedure for the diagnosis of dementia. All patients were re-examined 12-18 months later. A newly devised delayed cued recall test using semantic cues (The RI48 Test) was compared with three established memory tests: the Ten Word-List Recall from CERAD, the "Doors" and the "Shapes" Tests from "The Doors and People Test Battery". Forty-four % of the MCI patients fulfilled criteria for probable AD at follow-up. The RI48 Test classified correctly 88% of the MCI and SMC participants and was the best predictor of the status of MCI and mild AD as well as the outcome of the MCI patients. Poor visual memory was the second best predictor of those MCI patients who evolved to AD. A cued recall test which avoids the ceiling effect is at least as good as the delayed free recall tests in the early detection of AD. PMID- 15654552 TI - Posterior cortical atrophy: variant of Alzheimer's disease? A case series with PET findings. AB - Nine patients with posterior cortical atrophy (PCA), a rare degenerative brain disease of unclear etiology and nosology, were followed over a mean time of 7.4 years. The mean age at onset was low (56.2 years). At onset, eight patients had visuo-spatial and eight had memory impairment. A minority showed early signs of occipital lobe involvement with visual agnosia or hemianopia. Eight patients developed dementia after a mean course of five years. 18F-FDG-PET data of six patients were analysed with statistical parametric mapping. They showed hypometabolism centred on the lateral and medial parietal associative cortex, with variable involvement of the adjacent temporal and occipital associative cortex. A minority showed involvement of the frontal lobes, possibly related to deafferenting of areas related to the control of eye movements. Atrophy and hypometabolism were markedly asymmetric in a subset of cases. Autopsy was performed in one patient. Presenile onset, location, and asymmetry of atrophy suggest that PCA represents a biologically separable variant of Alzheimer's disease. PMID- 15654554 TI - Functional involvement of cerebral cortex in human narcolepsy. AB - The pathophysiology of human narcolepsy is still poorly understood. The hypoactivity of some neurotransmitter systems has been hypothesised on the basis of the canine model. To determine whether narcolepsy is associated with changes in excitability of the cerebral cortex, we assessed the excitability of the motor cortex with transcranial magnetic stimulation (TMS) in 13 patients with narcolepsy and in 12 control subjects. We used several TMS paradigms that can provide information on the excitability of the motor cortex. Resting and active motor thresholds were higher in narcoleptic patients than in controls and intracortical inhibition was more pronounced in narcoleptic patients. No changes in the other evaluated measures were detected. These results are consistent with an impaired balance between excitatory and inhibitory intracortical circuits in narcolepsy that leads to cortical hypoexcitability. We hypothesise that the deficiency of the excitatory hypocretin/orexin-neurotransmitter-system in narcolepsy is reflected in changes of cortical excitability since circuits originating in the lateral hypothalamus and in the basal forebrain project widely to the neocortex, including motor cortex. This abnormal excitability of cortical networks could be the physiological correlate of excessive daytime sleepiness and it could be the substrate for allowing dissociated states of wakefulness and sleep to emerge suddenly while patients are awake, which constitute the symptoms of narcolepsy. PMID- 15654555 TI - LGI1 gene mutation screening in sporadic partial epilepsy with auditory features. AB - Partial epilepsy with auditory features occasionally segregates in families as an autosomal dominant trait. In some families mutations in the leucine-rich glioma inactivated (LGI1) gene have been identified. Sporadic cases might harbour either denovo or low-penetrant LGI1 mutations, which will substantially alter the family risk for epilepsy. We selected sixteen sporadic patients with cryptogenic temporal lobe epilepsy and partial seizures with auditory features. We compared clinical features of these patients with those of published autosomal dominant family cases. We screened these patients for LGI1 mutations. Comparing the sporadic patients with the published familial cases no difference in either the primary auditory features or in the other associated epileptic manifestations was identified. Sequence analysis of the whole LGI1 gene coding regions in sporadic patients did not reveal changes in the LGI1 gene. The genetic analysis demonstrates that LGI1 is not a major gene for sporadic cases of partial epilepsy with auditory features at least in the Italian population. Screening of sporadic patients for LGI1 mutations appears not useful in genetic counselling of these patients. PMID- 15654556 TI - "Anxietas tibiarum". Depression and anxiety disorders in patients with restless legs syndrome. AB - BACKGROUND: Symptoms of anxiety and depression in patients with restless legs syndrome (RLS) have been observed. However, it is unclear whether rates of threshold depression and anxiety disorders according to DSM-IV criteria in such patients are also elevated. METHODS: 238 RLS patients were assessed with a standardized diagnostic interview (Munich-Composite International Diagnostic Interview for DSM-IV) validated for subjects aged 18-65 years. Rates of anxiety and depressive disorders were compared between 130 RLS patients within this age range and 2265 community respondents from a nationally representative sample with somatic morbidity of other types. RESULTS: RLS patients revealed an increased risk of having 12-month anxiety and depressive disorders with particularly strong associations with panic disorder (OR=4.7; 95% CI=2.1-10.1), generalized anxiety disorder (OR=3.5; 95% CI= 1.7-7.1), and major depression (OR=2.6; 95% CI=1.5 4.4). In addition, lifetime rates of panic disorder and most depressive disorders as well as comorbid depression and anxiety disorders were considerably increased among RLS patients compared with controls. CONCLUSIONS: The results suggest that RLS patients are at increased risk of having specific anxiety and depressive disorders. Causal attributions of patients suggest that a considerable proportion of the excess morbidity for depression and panic disorder might be due to RLS symptomatology. PMID- 15654557 TI - Differences in overshoot of blood pressure after head-up tilt in two groups with chronic autonomic failure: pure autonomic failure and multiple system atrophy. AB - On head down tilt to the supine horizontal position (tilt reversal) after head up tilt (HUT), patients with orthostatic hypotension may show an increase in blood pressure (BP) relative to baseline readings. We assessed this BP overshoot in 8 patients with pure autonomic failure (PAF, 64+/-13 years) and 8 patients with multiple system atrophy (MSA, 66+/-10 years). BP was intermittently measured during pre-tilt supine, HUT (60 degrees , 10 min), and post-tilt supine periods. In addition, beat-to-beat BP was measured continuously using the Portapres model 2 device to calculate stroke volume (SV), cardiac output (CO) and total peripheral resistance (TPR). There was systolic BP overshoot of > or = 15 mmHg after tilt reversal in 5 out of 8 PAF, but in only one of 8 MSA. A mean increase of systolic BP in PAF was significantly higher than that in MSA (p<0.01). TPR increased over baseline level after tilt reversal, although there was no significant difference. SV and CO levels during the post-tilt supine period were similar to baseline levels. In conclusion, BP overshoot was prominent in the PAF group but not in the MSA group. The phenomenon of BP overshoot while supine, especially in PAF, may have implications for long term cardiac and vascular damage in such patients. PMID- 15654559 TI - Testing for acanthocytosis A prospective reader-blinded study in movement disorder patients. AB - The presence of acanthocytosis in peripheral blood smears remains the hallmark of the clinical diagnosis of most neuroacanthocytosis syndromes, such as chorea acanthocytosis (ChAc) and McLeod syndrome. Genetic analyses and/or specific laboratory tests are available only for a minority of these disorders. Testing for acanthocytosis is hampered by the lack of data on normal amounts of acanthocytes assessed by a standardized method. We report a prospective reader blinded study designed to establish control values for abnormally shaped erythrocytes in healthy volunteers and patients with movement disorders (MDs) using light microscopic assessment of erythrocyte morphology in standard EDTA and isotonically diluted blood samples. We investigated a total of 100 patients fulfilling clinical criteria of specific MDs, 31 patients with MDs not matching any clinical criteria, and 37 healthy controls. In patients with diagnosed MDs and healthy controls, acanthocytes in dry blood smears were significantly more frequent following isotonic dilution compared with standard EDTA blood. In unfixed wet blood preparations of both EDTA blood and isotonically diluted blood, acanthocyte levels were significantly higher than in standard dry blood smear preparations. There were no statistical differences of acanthocyte levels in all test conditions between diagnosed MDs and healthy volunteers. There was no significant correlation of acanthocyte levels in all blood samples and preparations with age, sex or diagnosis. Thus, normal values were defined as the 99th percentile of combined results of the two groups of volunteers. Diluted blood combined with wet blood preparation showed high specificity (0.98) and the highest sensitivity of all test procedures (all genetically confirmed ChAc patients were detected). The reported method is cheap, readily available, and provides high specificity and sensitivity in respect to clinically relevant acanthocytosis. The use of isotonically diluted blood samples combined with unfixed wet blood preparation with a normal range of <6.3% of total erythrocytes is recommended to search for significant acanthocytosis in movement disorders. PMID- 15654558 TI - Fatigue in patients with lupus is not associated with disturbances in cerebral blood flow as detected by SPECT. AB - OBJECTIVES: Fatigue is a common complaint in patients with systemic lupus erythematosus (SLE). We investigated whether focal or general disturbances of cerebral blood flow (CBF), as assessed by SPECT, were associated with the presence of fatigue in an unselected group of SLE patients. METHODS: Fifty-six patients were included. Mean age was 47.5 years (+/-12.7), mean disease duration 14.7 years (+/-8.9), and disease activity measured by SLE disease activity index (SLEDAI) was 5.7 (+/-5.4). Fatigue was assessed by the Fatigue Severity Scale (FSS) and CBF by Tc-99m-hexamethyl propylamine oxime (HMPAO)-SPECT. The images were read and processed quantitatively by a computer program using the primary visual cortex as reference region and > 15% CBF deviation as definition of abnormality. RESULTS: The mean FSS score was 4.6 (+/-1.8). SPECT revealed focal CBF disturbances in 17 patients (30.4 %). Generalized symmetrical CBF reductions were present in 32 patients (57.1 %). There were no significant associations between CBF disturbances in any region of the brain and the degree of fatigue. CONCLUSIONS: Fatigue in SLE patients is not related to focal or general CBF disturbances. Therefore, factors that do not influence blood flow seem responsible for the fatigue phenomenon. PMID- 15654560 TI - Progression of parkinsonism in multiple system atrophy. AB - BACKGROUND: Progression of parkinsonian motor impairment is usually rapid and relentless in multiple system atrophy (MSA). However, it may also be subject to considerable variation. Prospective natural history studies using validated rating scales are required to accurately determine the progression of parkinsonism in MSA. OBJECTIVE: To assess the progression of parkinsonism in patients with the Parkinson variant of MSA. METHODS: Parkinsonian motor impairment was assessed on regular therapy at two time points (mean follow-up 11.8 months, SD 1.4) using the Hoehn and Yahr scale (H&Y), the Schwab and England ADL scale (SES) and the motor examination section of the UPDRS (UPDRS-III) in 38 patients with clinically probable MSA-P. RESULTS: We examined 38 patients with probable MSA-P (mean age 63.2 years, SD 7.4; mean disease duration 4.1 years, SD 3.0). The mean difference of UPDRS-III between baseline and follow-up was 10.8 (95% CI 8.6-12.9), consistent with an average annual 28.3 % increase of UPDRS-III baseline scores. Several variables were associated with faster progression of parkinsonism including low baseline global motor disability as assessed by H&Y and SES, low baseline UPDRS-III score, and short disease duration. UPDRS-III progression was unrelated to gender, age at symptom onset, and age at baseline visit. CONCLUSION: This is the first observational study on UPDRS rates of decline in MSA. The observed 28.6% annual increase of UPDRS-III scores illustrates the rapid progression of motor impairment in MSA. Furthermore, motor progression appeared to be accelerated during the early disease stages. Our data allow sample size calculations that may be helpful for the planning of future therapeutic trials. PMID- 15654561 TI - Response to "Cervical artery dissection-clinical features, risk factors, therapy and outcome in 126 patients [1]" by Dziewas et al. (2003) in J Neurol 250:1179 1184. PMID- 15654563 TI - Postmortem analysis of pallidotomy in Parkinson's disease. PMID- 15654564 TI - Chorea triggered by hyperglycemia in a maternally inherited diabetes and deafness (MIDD) patient with the A3243G mutation of mitochondrial DNA and basal ganglia calcification. PMID- 15654565 TI - Absence of the Babinski sign in brain death: a prospective study of 144 cases. PMID- 15654566 TI - Resolution of clinical and MR abnormalities in sudden onset massive hypertensive brain stem edema. PMID- 15654567 TI - Extrapyramidal and cerebellar movement disorder in association with heterozygous ceruloplasmin gene mutation. PMID- 15654568 TI - Gheorghe Marinescu (1863-1938). PMID- 15654572 TI - Evaluation of pediatric electrocardiogram diagnosis of ventricular hypertrophy by computer program compared with cardiologists. AB - This study aimed to evaluate the diagnosis of pediatric left ventricular hypertrophy (LVH) and right ventricular hypertrophy (RVH) by the Glasgow electrocardiogram (ECG) interpretation program compared to interpretations provided by two pediatric cardiologists. ECGs had all identifying information removed and were sent to the cardiologists independently with the patient's age and sex and the clinical indication for the ECG, if known. A total of 984 ECGs were included in the study, of which 664 were reported "with clinical indication" and 320 were reported "blind." With respect to an averaged diagnosis of the two cardiologists, the sensitivity of the program for RVH was better when the cardiologists reported blind (73.3%) than with the clinical indication (53.5%), with the same trend for the program compared with individual cardiologists. The specificity of the program was at least 94.4% in all cases. For LVH, the program had high specificity (=95.8%) for "reported blind" and "with clinical indication" cases but low sensitivities throughout (the highest was 44.4% with respect to an averaged diagnosis of the two cardiologists reporting with the clinical indication). Subsequent discussion revealed that if the cardiologists had disagreed with one another initially, their consensus opinion was twice as likely to be in agreement with the program. PMID- 15654573 TI - Hepatofugal portal flow due to acute cellular rejection. AB - We describe a case of living donor liver transplantation with hepatofugal portal flow caused by acute cellular rejection. The reversed portal flow was corrected by splenectomy and ligation of the residual collateral veins. Hepatofugal flow causes ischemic damage to the graft, which does not normalize spontaneously. In this particular case, meticulous Doppler ultrasound examination and prompt response to reversed portal flow salvaged the graft. PMID- 15654574 TI - Cystic ovarian teratoma with intracystic floating globules. AB - Ovarian cystic teratomas are cystic fatty tumors that are often found in patients of reproductive age, and the diagnosis can be easily made radiologically. We present a case of postmenopausal ovarian cystic teratoma with an unusual radiologic appearance of intracystic floating globules. PMID- 15654575 TI - Colon stenosis caused by old portal vein thrombosis. AB - A 58-year-old female with a history of portal vein thrombosis was referred to our hospital with abdominal pain and distention. Colon fiber and enema of the colon showed stenosis at the transverse colon and the ascending colon, with edematous mucosa. Laparotomy revealed no abnormal findings other than chronic ischemia of the colon. To our knowledge, this is the first reported case of colon stenosis caused by portal vein thrombosis. PMID- 15654576 TI - Does iodine concentration affect the diagnostic efficacy of biphasic spiral CT in patients with hepatocellular carcinoma? AB - BACKGROUND: We investigated the effect of iodinated contrast medium concentration on increased neoplastic lesion enhancement and its direct relation to diagnostic efficacy in biphasic spiral computed tomography for detection of hepatocellular carcinoma. METHODS: A pilot, single-center, randomized, double-blind, crossover, comparative study was performed and included 22 participants. Each patient underwent two separate biphasic contrast-enhanced spiral computed tomographic examinations. Scans were performed with iomeprol containing 400 (iomeprol 400) or 300 (iomeprol 300) mg of iodine per milliliter (Iomeron, Bracco Imaging SpA, Milan, Italy) with a 2- to 12-day window scan; patients were given an equal total dose of 45 g of iodine at a fixed injection rate of 4 mL/s. Comparison included assessment of quantitative and qualitative parameters. RESULTS: Lesion density and lesion-to-liver contrast increased more markedly with the higher concentration of contrast medium during the arterial phase (p = 0.0016 and 0.0005, respectively). There was no significant difference in any parameter between the two concentrations during the portal phase. Number of lesions detected during the arterial phase increased from 37 with iomeprol 300 to 42 with iomeprol 400; in the portal phase, the respective numbers were 34 and 36. CONCLUSION: Even though a small number of patients was examined, our study suggests that, in patients with cirrhosis, an increased concentration of iodine improves liver-to-lesion contrast and may improve the detection of hepatocellular carcinoma. PMID- 15654577 TI - Sinistral portal hypertension; imaging findings and endovascular therapy. AB - Sinistral portal hypertension, a rare and localized form of portal hypertension, is the result of splenic vein thrombosis or obstruction and may cause gastrointestinal hemorrhages from the esophageal and gastric varices. This report presents two cases (69- and 10-year-old females) of bleeding gastric varices. The patients were diagnosed as having sinistral portal hypertension. Splenic artery embolization was performed in both patients to overcome intractable bleeding, and the clinical outcome was good. PMID- 15654578 TI - Pancreatic carcinoma versus chronic focal pancreatitis: contrast-enhanced power Doppler ultrasonography findings. AB - BACKGROUND: We assessed the potential usefulness of contrast-enhanced power Doppler ultrasonography (US) for differentiating pancreatic carcinoma from chronic focal pancreatitis. METHODS: Twenty-six consecutive patients with ductal carcinoma (n=16) and chronic focal pancreatitis (n=10) underwent power Doppler US examinations before and after intravenous injection of an air-based contrast agent (Levovist, Schering, Berlin, Germany). Final diagnosis was obtained by surgery in all patients. The following parameters before and after intravenous administration of contrast agent were evaluated for each lesion: number, morphology and course of the vessels within the lesion and time to maximum detectable enhancement. Number of the lesion vessels and time to maximum enhancement for each lesion were compared by with Student t test. RESULTS: After injection of contrast agent, nine of 16 (56.2%) carcinomas showed a larger total number of vessels (p<0.005) and faster enhancement (p<0.0001) compared with chronic focal pancreatitis. Irregular vessels with a serpiginous course that penetrated the tumor was predominant in carcinoma (eight of 16 cases), whereas focal pancreatitis presented mostly as avascular masses (five of 10 cases). CONCLUSION: Contrast-enhanced power Doppler US provides useful findings for differentiating pancreatic carcinoma from chronic focal pancreatitis. PMID- 15654579 TI - Gastritis from NSAIDS to Helicobacter pylori. AB - Gastritis is a histologic diagnosis. To understand gastritis, the radiologist must have some working knowledge of gastric histology and pathology. Therefore, this article first describes normal histologic and radiologic anatomy. The pathology of gastritis is then presented to give the radiologist a basis for understanding the radiologic findings. Finally, gastritis is discussed from a clinical and radiologic perspective. PMID- 15654581 TI - Bone response to treatment with lower doses of conjugated estrogens with and without medroxyprogesterone acetate in early postmenopausal women. AB - Lower doses of conjugated estrogens (CE) alone or combined with lower doses of medroxyprogesterone acetate (MPA) increase mean bone mineral density (BMD) from baseline at the spine and hip in early postmenopausal women. However, not all women on therapy gain BMD. The incidence of continued bone loss (defined as a loss of BMD of > 2% from baseline) among women using lower doses of CE and CE/MPA is unknown. This randomized, double-blind, placebo-controlled, multicenter substudy of the Women's Health, Osteoporosis, Progestin, Estrogen (Women's HOPE) trial investigated the incidence of continued bone loss with lower-dose CE and CE/MPA. Eight hundred twenty-two healthy postmenopausal women with intact uteri received CE 0.625, CE 0.625/MPA 2.5, CE 0.45, CE 0.45/MPA 2.5, CE 0.45/MPA 1.5, CE 0.3, CE 0.3/MPA 1.5 (all doses in mg/day), or placebo for 2 years along with 600 mg/day of calcium. Changes from baseline in spine and total hip BMD were compared among treatment groups in an intent-to-treat analysis. At 12 months, < 10% of women on active treatment lost > 2% of spinal BMD (except CE 0.3/MPA 1.5 [15.6%]), compared with 41.2% of women on placebo. At 24 months, the percentages of women on active treatment who lost > 2% of spine BMD ranged from 4.5% with CE 0.45/MPA 1.5-15.6% with CE 0.3/MPA 1.5, compared with 55.2% of women taking placebo. More than 85% of women on active treatment did not experience continued BMD loss at the hip at 12 months and 24 months, in contrast to 30.6% of women on placebo at 12 months and 36.5% at 24 months. Women receiving active treatment who lost > 2% of spine or hip BMD also had a lesser reduction in biochemical markers of bone turnover. In summary, continued bone loss among early postmenopausal women treated with lower doses of CE or CE/MPA is uncommon. PMID- 15654580 TI - The potential of gene therapy for fracture healing in osteoporosis. AB - Osteoporosis-associated fractures impair a patient's function and quality of life and represent one of the major public health burdens. Demographic changes predict a dramatic increase in osteoporotic fractures. Experimental data have shown that osteoporosis impairs fracture healing. Clinical observations demonstrate high failure rates of implant fixation in osteoporosis. The reduced healing capacity, including impaired bone formation, in osteoporotic humans might be due to defects in mesenchymal stem cells that lead to reduced proliferation and osteoblastic differentiation. Growth factors show remarkable promise as agents that can improve the healing of bone or increase the proliferation and differentiation capacities of mesenchymal stem cells. Their clinical utility is limited by delivery problems. The attraction of gene-transfer approaches is the unique ability to deliver authentically processed gene products to precise anatomical locations at therapeutic levels for sustained periods of time. Unlike the treatment of chronic diseases, it is neither necessary nor desirable for transgene expression to persist beyond the few weeks or months needed to achieve healing. This review presents different approaches of gene therapy to enhance fracture healing and summarizes the promising results of preclinical studies. It focuses on applications of this new technique to fracture healing in osteoporosis. In our opinion, these applications represent some of the few examples in which gene therapy has a good chance of early clinical success. PMID- 15654582 TI - Loss of visual information in neglect: the effect of chromatic- versus luminance contrast stimuli in a "what" task. AB - One to four vertical bars were tachistoscopically presented at various eccentricities along the horizontal meridian to patients with right brain damage and neglect (n=7) and to a control group of right brain damaged patients without neglect (n=4). Luminance contrast stimuli and isoluminant chromatic contrast stimuli were used. The patients' task was to report the number of bars. With luminance stimuli the neglect patients' discrimination in the left visual field was reduced, particularly at the most leftward position tested (ca 20 degrees ) where about 20% of the bars were omitted. The loss of information was limited to luminance contrast stimuli. When chromatic contrast stimuli were used, accuracy was comparable in the two hemifields and between groups of patients. The differential pattern of results for chromatic and luminance contrast stimuli is consistent with the hypothesis of a selective deficit of the magno-cellular pathway in neglect. PMID- 15654583 TI - The influence of perturbation duration and velocity on the long-latency response to stretch in the biceps muscle. AB - Different neural pathways are proposed to mediate the long-latency stretch reflex response (M2) in muscles spanning distal and proximal joints of the upper limb. The M2 at the wrist joint is present only if the duration of the perturbation exceeds a critical time. Lee and Tatton put forward a converging input hypothesis, requiring an interaction of excitatory volleys at the spinal cord, to account for this feature. The goal of the present study was to examine the influence of the duration of perturbation on M2 responses elicited in a muscle spanning the elbow joint. Reflex responses were induced in the biceps brachii muscle by applying ramp-and-hold position displacements to the elbow. It was found that the M2 was strongly dependent on the duration of the perturbation. On average, responses were not elicited following perturbations of less than 35+/-5 ms. Using a novel double-movement paradigm, we were unable to provide support for the converging input hypothesis. The effect of the duration of perturbation on the M2 may account for the conflicting characteristics of the M2 that have been provided by previous studies applying mechanical or electrical perturbations of varying time durations. PMID- 15654585 TI - Observation, imagination and execution of an effortful movement: more evidence for a central explanation of motor imagery. AB - In this study subjects had to imagine, observe and perform a series of 25 squat movements while lifting two dumbbells of 12.5 kg each (one with each hand). This movement is effortful and requires substantial activation of peripheral systems. It was asked whether subjects when they imagined that they were performing the movements or when they observed a model performing the squat movements would show increased activity in EMG, heart rate and respiration compared with a control condition where they sat relaxed in a comfortable chair or a condition where they actually performed the squat movements. Two groups of subjects participated in the experiment: experienced squatters and novices. By employing these two groups we were able to study the differential effect of earlier experience with the target movement on peripheral activation. The results showed that with the exception of respiration no significant peripheral activation could be measured related to motor imagery. Although a clear distinction in experience existed between the experienced squatters versus the novices, no relevant imagery-related differences could be obtained between the two groups. The results are discussed in the light of a central explanation of motor imagery. PMID- 15654586 TI - Modulation of intracortical excitability in human hand motor areas. The effect of cutaneous stimulation and its topographical arrangement. AB - Changes in afferent input can alter the excitability of intracortical inhibitory systems. For example, using paired transcranial magnetic stimulation (TMS), both electrical digital stimulation and muscle vibration have been shown to reduce short-interval intracortical inhibition (SICI). The effects following muscle vibration are confined to the corticospinal projection to the vibrated muscles. The results following digital stimulation are less clear and the relative timing of the cutaneous stimulation and TMS is critical. Here we investigated further whether changes in SICI following digit stimulation exhibit topographic specificity. Eleven normal subjects were investigated (age 28.2+/-7.5 years, mean+/-SD). Electromyographic recordings were made from the right first dorsal interosseous (FDI), abductor digiti minimi (ADM) and abductor pollicis brevis (APB) muscles. SICI was measured, with and without preceding electrical digit II or digit V cutaneous stimulation. The interval between the digital nerve stimulus and test magnetic stimulus was independently set for each subject and established by subtracting the onset latency of the motor evoked potential (MEP) from the latency of the E2 component of the cutaneomuscular reflex. Therefore, measures of intracortical excitability were made at a time at which it is known that cutaneous input is capable of modulating cortical excitability. Single digital nerve stimuli applied to digit II significantly reduced SICI in FDI but not in ADM. Single digital nerve stimuli applied to digit V significantly reduced SICI in ADM but not in FDI or APB. There was a more generalised effect on intracortical facilitation (ICF) with both digit II and digit V stimulation significantly increasing ICF in FDI and ADM. Digital stimulation (either DII or DV) did not significantly affect SICI/ICF in APB. These findings show that appropriately timed cutaneous stimuli are capable of modulating SICI in a topographically specific manner. We suggest that the selective decrease in SICI seen with cutaneous stimulation may be important for focusing of muscle activation during motor tasks. PMID- 15654584 TI - The methionine allele of the COMT polymorphism impairs prefrontal cognition in children and adolescents with ADHD. AB - ADHD is a highly heritable psychiatric disorder of childhood. A functional polymorphism (Val158Met) of the catechol-O-methyltransferase (COMT) gene has attracted interest as a candidate gene for ADHD. The high-activity valine variant of this polymorphism degrades prefrontal dopamine three to four times more quickly than the low-activity methionine variant and could therefore contribute to the proposed hypodopaminergic state in ADHD. Here we tested for association of this polymorphism with ADHD and examined its influence on prefrontal cognition in ADHD. We have previously reported no association of the Val158Met COMT gene polymorphism in 94 Irish ADHD families (Hawi et al. (2000) Am J Med Genet 96:282 284). Here we re-examined this finding with an extended sample of 179 ADHD cases using a family control design. We also examined the performance of children and adolescents with ADHD (n = 61) on a standardised test of sustained attention. Analysis confirmed the absence of an association between the Val158Met COMT gene polymorphism and the clinical phenotype of ADHD. COMT genotype, however, affected prefrontal cognition in ADHD: ADHD children who were homozygous for the valine variant had significantly better sustained attention than those ADHD children possessing at least one copy of the methionine variant. Children possessing the methionine variant performed significantly below age-related norms on tests of sustained attention. Contrary to expectations, the methionine variant of the Val158Met COMT gene polymorphism impaired prefrontally-mediated cognition in ADHD. This effect may be understood by positing a hyper-functioning of prefrontal dopaminergic systems. Against this background, the slower clearance of dopamine associated with the methionine variant of the COMT gene polymorphism may be disadvantageous to cognition in ADHD. PMID- 15654587 TI - The effects of dividing attention on smooth pursuit eye tracking. AB - Attentional processes have traditionally been closely linked to the production of saccadic eye movements, but their role in the control of smooth pursuit eye movements remains unclear. In two experiments we used dual task paradigms to vary the attentional resources available for pursuit eye tracking. In both experiments we found that attentionally demanding secondary tasks impaired smooth pursuit performance, resulting in decreased velocity and increased position error. These findings suggest that attention is important for the maintenance of accurate smooth pursuit, and do not support the hypothesis that pursuit is a relatively automatic function that proceeds optimally in the absence of attentional control. These results add weight to the suggestion that a similar functional architecture underlies both pursuit and saccadic eye movements. PMID- 15654588 TI - Stretch reflex instability compared in three different human muscles. AB - The possibility of causing instability in the stretch reflex has been examined in three different human muscles: biceps, first dorsal interosseous (FDI) of the hand and digastric. Tremor recorded as fluctuation of isometric force was compared with that occurring during contraction against a spring load. The spring compliance was selected to make the natural frequency of the part in each case appropriate for oscillations in the short latency stretch reflex. A computer model of the whole system was used to predict the frequency at which oscillations should be expected and to estimate the reflex gain required in each case to cause sustained oscillations. Estimates were computed of the autospectra of the force records and of the rectified surface EMG signals and of the coherence functions. Normal subjects showed no evidence of a distinct spectral peak during isometric recording from any of the three muscles. However, in anisometric conditions regular oscillations in force occurred in biceps, but not in FDI or digastric. The oscillations in biceps at 8-9 Hz were accompanied by similar oscillations in the EMG which were highly coherent with the force signal. The results are consistent with the presence of a strong segmental stretch reflex effect in biceps and weak or absent reflex in FDI. Digastric is known to contain no muscle spindles and therefore to lack a stretch reflex. In two subjects who volunteered that they had more tremor than normal, but had no known neurological abnormality, there was a distinct peak in the force spectrum at 8-9 Hz in biceps and FDI in isometric conditions with coherent EMG activity. The peak increased in size in anisometric conditions in biceps but not in FDI. This component appears to be of central rather than of reflex origin. No equivalent component was found in digastric records. The results are discussed in relation to the possible role of the short latency stretch reflex in the genesis of physiological tremor in different muscles. PMID- 15654589 TI - The influence of metricality and modality on synchronization with a beat. AB - The great majority of the world's music is metrical, i.e., has periodic structure at multiple time scales. Does the metrical structure of a non-isochronous rhythm improve synchronization with a beat compared to synchronization with an isochronous sequence at the beat period? Beat synchronization is usually associated with auditory stimuli, but are people able to extract a beat from rhythmic visual sequences with metrical structure? We addressed these questions by presenting listeners with rhythmic patterns which were either isochronous or non-isochronous in either the auditory or visual modality, and by asking them to tap to the beat, which was prescribed to occur at 800-ms intervals. For auditory patterns, we found that a strongly metrical structure did not improve overall accuracy of synchronization compared with isochronous patterns of the same beat period, though it did influence the higher-level patterning of taps. Synchronization was impaired in weakly metrical patterns in which some beats were silent. For the visual patterns, we found that participants were generally unable to synchronize to metrical non-isochronous rhythms, or to rapid isochronous rhythms. This suggests that beat perception and synchronization have a special affinity with the auditory system. PMID- 15654590 TI - Manual-aiming bias and the Muller-Lyer illusion: the roles of position and extent information. AB - Several studies have shown that rapid manual aiming movements from one to the other wing of Muller-Lyer illusion figures are biased in a manner consistent with their influence on perceptual judgments. Two experiments examined the role of extent and position information in Muller-Lyer figures in biasing pointing movements. Experiment 1 compared the effect of starting the action either close to the non-target wing or from a position well outside the conventional wings-out (>-<) and wings-in (<->) Muller-Lyer figures. Pointing movements were longer to the target wing of the wings-out than of the wings-in figure but only for actions starting from a position adjacent to the non-target wing. Experiment 2 found no effect of the direction of the target wing on pointing movements between the outer wings of the combined Muller-Lyer illusion figure (<->-<). These findings suggest that perceived extent (of the shaft) and perceived position (of the wings) in Muller-Lyer figures are largely independent of each other, and are consistent with the claim that the illusion is more likely to influence pointing actions if the action is based on extent information than on vertex position information. PMID- 15654591 TI - Effects of stimulus-response uncertainty on saccades to near-threshold targets. AB - Kveraga et al. (2002, Exp Brain Res 146(3):307-14) reported that saccade latencies are immune to the effects of stimulus-response uncertainty and constitute one of the few response systems that violate Hick's law. Similar effects have been reported for keypresses triggered by vibrations of the fingertips, but robust uncertainty effects were subsequently revealed using weak, low-frequency vibrations (Ten Hoopen et al. 1982, Acta Psychol 50:143-157). We wondered whether immunity of saccadic responses would demonstrate a similar intensity-dependency and therefore re-examined the effects of response entropy on saccade latencies using near-threshold visual stimuli. Saccadic latencies remained independent of stimulus-response uncertainty, indicating that saccadic motor programming is unaffected by the duration of the target detection process. PMID- 15654593 TI - Spatially selective reward site responses in tonically active neurons of the nucleus accumbens in behaving rats. AB - To study how hippocampal output signals conveying spatial and other contextual information might be integrated in the nucleus accumbens, tonically active accumbens neurons were recorded in three unrestrained rats as they performed spatial orientation tasks on an elevated round rotatable platform with four identical reward boxes symmetrically placed around the edge. The partially water deprived rats were required to shuttle either between the pair of reward boxes indicated by beacon cues (lights in the boxes) or between the pair of boxes occupying particular locations in relation to environmental landmark cues. In 43/82 neurons, behaviorally correlated phasic modulations in discharge activity occurred, primarily prior to or after water was provided at the reward boxes. Twenty-two had inhibitory modulation, 12 excitatory, and nine were mixed excitatory and inhibitory. Although tonically active neurons (TANs) have rarely been reported in the rodent, the inhibitory and mixed responses correspond to previously reports in the macaque accumbens of tonically active neurons with activity correlated with reward delivery and, following conditioning, to sensory stimuli associated with rewards. Eighteen of the 43 tonically active accumbens neurons showed spatial selectivity, i.e., behaviorally correlated increases or decreases in firing rate were of different magnitudes at the respective reward boxes. This is the first demonstration that the configuration of environmental sensory cues associated with reward sites are also an effective stimulus for these neurons and that different neurons are selective for different places. These results are consistent with a role for the nucleus accumbens in the initiation of goal-directed displacement behaviors. PMID- 15654592 TI - Visual motion due to eye movements helps guide the hand. AB - Movement of the body, head, or eyes with respect to the world creates one of the most common yet complex situations in which the visuomotor system must localize objects. In this situation, vestibular, proprioceptive, and extra-retinal information contribute to accurate visuomotor control. The utility of retinal motion information, on the other hand, is questionable, since a single pattern of retinal motion can be produced by any number of head or eye movements. Here we investigated whether retinal motion during a smooth pursuit eye movement contributes to visuomotor control. When subjects pursued a moving object with their eyes and reached to the remembered location of a separate stationary target, the presence of a moving background significantly altered the endpoints of their reaching movements. A background that moved with the pursuit, creating a retinally stationary image (no retinal slip), caused the endpoints of the reaching movements to deviate in the direction of pursuit, overshooting the target. A physically stationary background pattern, however, producing retinal image motion opposite to the direction of pursuit, caused reaching movements to become more accurate. The results indicate that background retinal motion is used by the visuomotor system in the control of visually guided action. PMID- 15654596 TI - [Commentary on the German adaptation of Consort statements]. PMID- 15654594 TI - Differences in preferred reference frames for postural orientation shown by after effects of stance on an inclined surface. AB - This study reports a postural after-effect of leaning that follows a period of stance on an inclined surface with eyes closed. This leaning after-effect maintained the body-to-surface relationship as if subjects still stood on the incline. We examined the incidence and robustness of the leaning after-effect in 51 healthy subjects. The location of the center of pressure (CoP) under the feet and the alignment of the trunk and legs were measured before, during and after blindfolded subjects stood on a 5 degrees toes-up inclined surface for 2.5 min. When the surface was inclined, all subjects stood with their trunk and legs aligned near to gravity-vertical, similar to the alignment adopted in the pre incline period. When the surface returned to horizontal in the post-incline period, there was a continuum of postural alignment strategies across subjects. At one extreme, subjects leaned forward, with an average trunk lean near 5 degrees . The leaned posture decayed exponentially toward baseline postural alignment across a period of up to 5 min. At the other extreme, subjects did not lean in the post-incline period, but instead, stayed aligned near upright with respect to gravity. Subjects were highly consistent in their post-incline postural behaviors upon repeated testing over days to months and across different directions of surface inclination. Our results suggest that individuals have well established, preferred, sensory strategies for controlling postural orientation when vision is not available. Subjects who leaned in the post-incline period appear to depend more on the geometry of the support surface as a reference frame and to rely more on proprioceptive information to extract kinematic relationships, whereas subjects who did not lean appear to depend more on gravity as a reference frame and to rely more on sensory information related to forces and load. PMID- 15654597 TI - Macaques' (Macaca mulatta) use of numerical cues in maze trials. AB - We tested the ability of number-trained rhesus monkeys to use Arabic numeral cues to discriminate between different series of maze trials and anticipate the final trial in each series. The monkeys' prior experience with numerals also allowed us to investigate spontaneous transfer between series. A total of four monkeys were tested in two experiments. In both experiments, the monkeys were trained on a computerized task consisting of three reinforced maze trials followed by one nonreinforced trial. The goal of the maze was an Arabic numeral 3, which corresponded to the number of reinforced maze trials in the series. In experiment 1 (n=2), the monkeys were given probe trials of the numerals 2 and 4 and in experiment 2 (n=2), they were given probe trials of the numerals 2-8. The monkeys receiving the probe trials 2 and 4 showed some generalization to the new numerals and developed a pattern of performing more slowly on the nonreinforced trial than the reinforced trial before it for most series, indicating the use of the changing numeral cues to anticipate the nonreinforced trial. The monkeys receiving probe trials of the numerals 2-8 did not predict precisely when the nonreinforced trial would occur in each series, but they did incorporate the changing numerals into their strategy for performing the task. This study provides the first evidence that number-trained monkeys can use Arabic numerals to perform a task involving sequential presentations. PMID- 15654595 TI - [Macular hole. Survey and relevant surgical concepts]. AB - The theory of macular hole pathogenesis, which had so far been based on biomicroscopy, has been considerably altered by optical coherence tomography. The precise presentation of vitreofoveal pathology shows that forces acting in different directions are associated with different stages of the disease, making surgical treatment adapted to the different stages possible. Some surgical procedures are still controversial, and there is still no gold standard in macular hole surgery. Especially no agreement exists on the benefit of internal limiting membrane peeling, possibly assisted by staining with indocyanine green. Also details of endotamponade and postoperative positioning are controversial. Therefore, the method of surgical treatment depends a lot on the individual surgeon. This review summarizes the broad spectrum of the literature and the present knowledge in this field. PMID- 15654598 TI - Thaliporphine increases survival rate and attenuates multiple organ injury in LPS induced endotoxaemia. AB - This study addressed the question of whether thaliporphine, a phenolic aporphine alkaloid obtained from Chinese herbs and possessing antioxidant and alpha-1 adrenoceptor antagonistic activity, has protective effects in endotoxaemic rats and we attempted to elucidate the mechanisms contributing to such protective effects. Injection of rats with endotoxin (E. coli lipopolysaccharide, LPS) induced severe hypotension and tachycardia as well as vascular hyporeactivity to noradrenaline. Pretreatment of LPS-treated rats with thaliporphine attenuated the delayed hypotension significantly whilst only a higher dose (1 mg/kg) of thaliporphine decreased LPS-induced tachycardia. LPS significantly increased nitric oxide (NO.) and superoxide anion (O(2).(-)) levels, a response that was reduced by pretreatment with 1 mg/kg thaliporphine. Endotoxaemia for 240 min resulted in a bell-shaped time course for the change of serum tumour necrosis factor-alpha (TNF-alpha) level with a peak at 60 min. Pretreatment of LPS-treated rats with 1 mg/kg thaliporphine significantly reduced the serum TNF-alpha level at 60 min. In addition, LPS caused a biphasic change in blood glucose and thaliporphine attenuated the late-phase decrease in blood glucose. Endotoxaemia induced multiple organ injury in the liver, kidney and heart, as indicated by increases of aspartate aminotransferase (GOT), alanine aminotransferase (GPT), creatinine (CRE), lactate dehydrogenase (LDH) and creatine phosphate kinase muscle-brain (CKMB) levels in serum. These increases of biochemical markers and inflammatory cell infiltration into injured tissues were reduced significantly by treatment with thaliporphine. In addition, thaliporphine increased the survival rate of LPS-treated mice dose-dependently. In conclusion, our results suggest that thaliporphine could be a novel agent for attenuating endotoxin-induced circulatory failure and multiple organ injury and may increase the survival rate. These beneficial effects of thaliporphine may be attributed to the suppression of TNF-alpha, NO. and O(2).(-) production. PMID- 15654599 TI - MHC class I genes in the owl monkey: mosaic organisation, convergence and loci diversity. AB - The MHC class I molecule plays an important role in immune response, pathogen recognition and response against vaccines and self- versus non-self-recognition. Studying MHC class I characteristics thus became a priority when dealing with Aotus to ensure its use as an animal model for biomedical research. Isolation, cloning and sequencing of exons 1-8 from 27 MHC class I alleles obtained from 13 individuals classified as belonging to three owl monkey species (A. nancymaae, A. nigriceps and A. vociferans) were carried out to establish similarities between Aotus MHC class I genes and those expressed by other New and Old World primates. Six Aotus MHC class I sequence groups (Ao-g1, Ao-g2, Ao-g3, Ao-g4, Ao-g5 and Ao g6) weakly related to non-classical Catarrhini MHC were identified. An allelic lineage was also identified in one A. nancymaae and two A. vociferans monkeys, exhibiting a high degree of conservation, negative selection along the molecule and premature termination of the open reading frame at exon 5 (Ao-g5). These sequences' high conservation suggests that they more likely correspond to a soluble form of Aotus MHC class I molecules than to a new group of processed pseudogenes. Another group, named Ao-g6, exhibited a strong relationship with Catarrhini's classical MHC-B-C loci. Sequence evolution and variability analysis indicated that Aotus MHC class I molecules experience inter-locus gene conversion phenomena, contributing towards their high variability. PMID- 15654600 TI - Renal clearance of glycolaldehyde- and methylglyoxal-modified proteins in mice is mediated by mesangial cells through a class A scavenger receptor (SR-A). AB - AIMS/HYPOTHESIS: Glomerular mesangial expansion is a characteristic feature of diabetic nephropathy, and the accumulation of AGE in the mesangial lesion has been implicated as one of its potential causes. However, the route for the AGE accumulation in mesangial lesions in diabetic patients is poorly established. METHODS: Glycolaldehyde-modified BSA (GA-BSA) and methylglyoxal-modified BSA (MG BSA) were prepared as model AGE proteins, and their in vivo plasma clearance was examined in mice, and renal uptake by in vitro studies with isolated renal mesangial cells. RESULTS: Both (111)In-GA-BSA and (111)In-MG-BSA were rapidly cleared from the circulation mainly by both the liver and kidney. Immunohistochemical studies with an anti-GA-BSA antibody demonstrated that intravenously injected GA-BSA accumulated in mesangial cells, suggesting that such cells play an important role in the renal clearance of circulating AGE proteins. Binding experiments at 4 degrees C using mesangial cells isolated from mice showed that (125)I-GA-BSA and (125)I-MG-BSA exhibited specific and saturable binding. Upon incubation at 37 degrees C, (125)I-GA-BSA and (125)I-MG-BSA underwent endocytic degradation by these cells. The binding of the ligands to these cells was inhibited by several ligands for scavenger receptors. The endocytic degradation of GA-BSA by mesangial cells from class A scavenger receptor (SR-A) knock-out mice was reduced by 80% when compared with that of wild type cells. The glomerular accumulation of GA-BSA after its intravenous administration was attenuated in SR-A knock-out mice, as evidenced by immunohistochemical observations. CONCLUSIONS/INTERPRETATION: These results raise the possibility that circulating AGE-modified proteins are subjected to renal clearance by mesangial cells, mainly via SR-A. This pathway may contribute to the pathogenesis of AGE-induced diabetic nephropathy. PMID- 15654601 TI - Inhibition of endogenous SHIP2 ameliorates insulin resistance caused by chronic insulin treatment in 3T3-L1 adipocytes. AB - AIMS/HYPOTHESIS: SHIP2 is a physiologically important negative regulator of insulin signalling hydrolysing the PI3-kinase product, PI(3,4,5)P3, which also has an impact on insulin resistance. In the present study, we examined the effect of inhibiting the endogenous SHIP2 function on the insulin resistance caused by chronic insulin treatment. METHODS: The endogenous function of SHIP2 was inhibited by expressing a catalytically inactive SHIP2 (DeltaIP-SHIP), and compared with the effect of treatments designed to restore the levels of IRS-1 in insulin signalling systems of 3T3-L1 adipocytes. RESULTS: Chronic insulin treatment induced the large (86%) down-regulation of IRS-1 and the modest (36%) up-regulation of SHIP2. Subsequent stimulation by insulin of Akt phosphorylation, PKClambda activity, and 2-deoxyglucose (2-DOG) uptake was markedly decreased by the chronic insulin treatment. Coincubation with the mTOR inhibitor, rapamycin, effectively inhibited the proteosomal degradation of IRS-1 caused by the chronic insulin treatment. Although the coincubation with rapamycin and advanced overexpression of IRS-1 effectively ameliorated subsequent insulin-induced phosphorylation of Akt, insulin stimulation of PKClambda activity and 2-DOG uptake was partly restored by these treatments. Similarly, expression of DeltaIP SHIP2 effectively ameliorated the insulin-induced phosphorylation of Akt without affecting the amount of IRS-1. Furthermore, the decreased insulin-induced PKClambda activity and 2-DOG uptake following chronic insulin treatment were ameliorated by the expression of DeltaIP-SHIP2 more effectively than by the treatment with rapamycin. CONCLUSIONS/INTERPRETATION: Our results indicate that the inhibition of endogenous SHIP2 is effective in improving the state of insulin resistance caused by chronic insulin treatment. PMID- 15654603 TI - High titres of IgA antibodies to enterovirus in fulminant type-1 diabetes. AB - AIMS/HYPOTHESIS: We have recently proposed that fulminant type-1 diabetes is a novel subtype of type-1 diabetes with abrupt onset of insulin-deficient hyperglycaemia without islet-related autoantibodies. The pathogenesis is still unknown, but flu-like symptoms are frequently observed before the onset of disease of this subtype. Enterovirus infection is a candidate environmental factor causing type-1 diabetes. The aim of this study was to determine whether enterovirus infection contributes to the development of fulminant type-1 diabetes. METHODS: We investigated 19 patients with recent-onset fulminant type-1 diabetes, 18 patients with recent-onset typical type-1A diabetes, and 19 healthy controls. IgM, IgG, and IgA subclasses of antibodies to enterovirus were determined by ELISA. RESULTS: IgA antibody titres to enterovirus were significantly higher in fulminant type-1 diabetes than in typical type-1A diabetes (p=0.033) and controls (p=0.0003). IgM antibodies to enterovirus were not detected in any subject. IgG titres were lower in autoimmune diabetes than fulminant type and controls (p=0.014 and 0.019, respectively). CONCLUSIONS/INTERPRETATION: High titres of enterovirus IgA antibodies in serum suggest recurrent enterovirus infection in fulminant type-1 diabetic patients, indicating higher susceptibility to enteroviral infections among them. Such infections might have pathogenetic importance in the triggering of fulminant type 1 diabetes. PMID- 15654604 TI - Percutaneous transluminal angioplasty of an occluded surgical splenorenal shunt in a 4-year-old child after liver transplantation. AB - Percutaneous transluminal angioplasty is increasingly used in children. We present a case of successful balloon angioplasty of an occluded surgical splenorenal shunt in a 4-year-old child who underwent liver transplantation because of biliary atresia. Percutaneous reopening of the shunt is a relatively safe procedure that may spare patients of surgical intervention. PMID- 15654602 TI - Functional and morphological alterations of mitochondria in pancreatic beta cells from type 2 diabetic patients. AB - AIMS/HYPOTHESIS: Little information is available on the insulin release properties of pancreatic islets isolated from type 2 diabetic subjects. Since mitochondria represent the site where important metabolites that regulate insulin secretion are generated, we studied insulin release as well as mitochondrial function and morphology directly in pancreatic islets isolated from type 2 diabetic patients. METHODS: Islets were prepared by collagenase digestion and density gradient purification, and insulin secretion in response to glucose and arginine was assessed by the batch incubation method. Adenine nucleotides, mitochondrial membrane potential, the expression of UCP-2, complex I and complex V of the respiratory chain, and nitrotyrosine levels were evaluated and correlated with insulin secretion. RESULTS: Compared to control islets, diabetic islets showed reduced insulin secretion in response to glucose, and this defect was associated with lower ATP levels, a lower ATP/ADP ratio and impaired hyperpolarization of the mitochondrial membrane. Increased protein expression of UCP-2, complex I and complex V of the respiratory chain, and a higher level of nitrotyrosine were also found in type 2 diabetic islets. Morphology studies showed that control and diabetic beta cells had a similar number of mitochondria; however, mitochondrial density volume was significantly higher in type 2 diabetic beta cells. CONCLUSIONS/INTERPRETATION: In pancreatic beta cells from type 2 diabetic subjects, the impaired secretory response to glucose is associated with a marked alteration of mitochondrial function and morphology. In particular, UCP 2 expression is increased (probably due to a condition of fuel overload), which leads to lower ATP, decreased ATP/ADP ratio, with consequent reduction of insulin release. PMID- 15654605 TI - 18F-FDG-avid sites mimicking active disease in pediatric Hodgkin's. AB - About 1,700 children in the United States are diagnosed yearly with lymphomas; Hodgkin's disease accounts for approximately half of these cases, or 6% of all childhood cancers. Contemporary therapy allows for the achievement of remission in the majority of cases. The fusion of positron emission tomography (PET) with CT provides the most accurate imaging method for disease characterization and treatment response. However, experience with 18F-FDG PET-CT is limited in pediatric Hodgkin's disease. Numerous non-oncologic processes can mimic recurrent or residual tumor. This pictorial addresses mimickers of disease such as uptake in normal structures, infections, transforming germinal canters and effects of therapy on normal tissues. It is essential for radiologists to be familiar with these findings in order to stage disease activity and therapeutic response accurately. PMID- 15654606 TI - Expression levels of hepatic cytochrome P450 enzymes in Aldh2-deficient mice following ethanol exposure: a pilot study. AB - We determined expression levels of hepatic cytochrome P450 (CYP) 2E1, 2B1/2 and 4B1 enzymes in aldehyde dehydrogenase 2 (Aldh2) +/+ and Aldh2 -/- mice by immunoblotting assay following subchronic ethanol exposure for eight days. Using ethanol exposure, the protein expression levels of CYP2E1, 2B1/2 and 4B1 in Aldh2 +/+ mice were increased by factors of 2.61, 1.88 and 2.01 compared with Aldh2 +/+ mice that were not exposed to ethanol, respectively. On the other hand, in the Aldh2 -/- mice, CYP2E1, 2B1/2 and 4B1 protein expression levels after ethanol treatment were shown to be 1.99, 1.05 and 1.33 greater than those of Aldh2 -/- control mice, respectively. We also found an interesting fact in the present study; the Aldh2 -/- mice were shown to have higher CYP2E1 (by a factor of 4.19), 2B1/2 (by a factor of 2.89) and 4B1 (by a factor of 1.53) protein expression levels than Aldh2 +/+ mice despite the lack of ethanol treatment. These results suggest that CYP2E1, 2B1/2 and 4B1 play some role in ethanol metabolism and that Aldh2-deficient individuals may have higher levels of CYP2E1, 2B1/2 and 4B1 enzymes compared to Aldh2 wild-type individuals. PMID- 15654607 TI - Role of diacetyl metabolite in alcohol toxicity and addiction via electron transfer and oxidative stress. AB - There are many gaps in our knowledge of the molecular basis of alcohol toxicity and addiction. Metabolism affords mainly acetic acid via acetaldehyde. A minor metabolite, diacetyl (an alpha-dicarbonyl), arises from the aldehyde. We propose that this C(4) entity and/or its iminium derivatives from condensation with protein amino groups plays important roles in bioresponses. A review of the literature reveals substantial support for this premise. Reduction potentials for diacetyl and its iminium derivatives fall in the range favorable for catalytic electron transfer in vivo, which can generate oxidative stress via reactive oxygen species due to redox cycling. Oxidative stress and reactive oxygen species are linked to toxicity associated with major organs by alcohol. The alpha dicarbonyl moiety in related substances is believed to induce various toxic responses, such as Alzheimer's disease, mutagenesis, and carcinogenesis. In addition to discussion of addiction and computational studies, potential applications for health improvement are suggested. PMID- 15654608 TI - Inappropriate sinus node tachycardia following gastric transposition surgery in children. PMID- 15654611 TI - MIR spectroscopic analysis on sugar metabolic and ethanol productive kinetics of suspension TBY-2 and rice cells pre-cultured in various media. AB - The influence of sugars in pre-cultivation media suspended plant cells on the kinetics of the sugar uptake and the ethanol production was studied by mid infrared spectroscopy using a Fourier transform infrared spectrometer (FT-IR) equipped with an attenuate total reflection accessory (ATR). We performed the plant cell cultivation with Nicotiana tabacum cv. Bright Yellow No.2 (TBY-2) cells and Oryza sativa L., Japonica, cv. Nipponbare (rice) cells, respectively, in pre-culture and culture media, which had various types of glucose, fructose, sucrose or glucose-fructose mixtures. The results confirmed the kinetic differences between the TBY-2 cells and rice cells. These results suggested that the TBY-2 cells consumed sugar before growth and the rice cells consumed sugar after growth, moreover, the ethanol content increased just after cell growth was activated based on the non-dimensional cultivation time for the cell growth behavior. PMID- 15654612 TI - [D-dimer screening in surgical long-term intensive care patients]. AB - BACKGROUND: Clinically unrecognized thrombosis with the danger of a pulmonary embolism represents an underestimated problem in surgical ICU patients. In patients undergoing total hip replacement for instance, over 30% develop a clinically inapparent form of thrombosis, despite initial thrombosis prophylaxis. We tried to recognize clinically inapparent thrombosis in long-term intensive care patients using D-dimer screening and ultrasound imaging. METHODS: All surgical long-term ICU patients received intravenous heparin 5-10 IU/kg body weight and a D-dimer was assay was carried out every 2 days. If the D-dimer level surpassed 2 mg/l, ultrasound imaging of the veins in the legs, pelvis, arms and neck was performed. RESULTS: Included in the study were 50 patients and D-dimer levels above 2 mg/l were detected in 38%. A thrombosis was proven in 63% of the patients with D-dimer values above 2 mg/l and 50% of the thrombosis were detected in the arm and neck veins very often associated with intravenous catheters. CONCLUSION: Routine D-dimer screening and specific use of ultrasound imaging appears to be a valuable method to verify clinically inapparent thromboses in surgical ICU patients. PMID- 15654610 TI - Cajal-like cells in the upper urinary tract: comparative study in various species. AB - The interstitial cells of Cajal (ICC) play an important role in the control of gut motility. The recognition that the ICC cell membrane harbors the c-kit receptor (CD117) sparked rapid advancement in ICC research on the gut and certain pathologies using immunochemical and molecular methods. The question arises whether ICC exist in the upper urinary tract (UUT) and trigger motility. The present study analyzed the distribution of the c-kit receptor in the normal human UUT compared with various species. Immunohistochemistry (alkaline-phosphatase anti-alkaline-phosphatase technique, immunofluorescence) was applied on serial sections using monoclonal and polyclonal antibodies recognizing the c-kit receptor. C-kit staining was compared with standard endothelial, epithelial, neurogenic, histiocytic, mast cell, and smooth muscle markers, as well as a negative control. Normal proximal, middle, and distal ureter segments were analyzed in rodents, carnivores, porcines, cow, and humans. In all species the c kit receptor was detected in either round or spindle-shaped cells. Because of their antigenic profile, the round cells were identified as mast cells occurring in all layers of the ureteral wall except the urothelium and were more frequent in humans. In contrast, the population of spindle-shaped cells was marked only by anti-c-kit receptor antibodies, thus resembling ICC. These ICC-like cells were found among the inner and outer smooth muscle layers and in the lamina propria of all species. In humans, spindle-shaped cells were also found vertically oriented within the urothelium. Our morphological data present for the first time the distribution of ICC in the UUT of various species. The ubiquitous distribution in the entire pyeloureteral complex provides strong evidence that ICC generate electrical pacemaker activity within the UUT as an intrinsic system. Animal studies may help to understand the physiological importance of these ICC-like cells. The significance of these findings needs to be evaluated by functional studies and investigations of certain congenital pathologies with disturbance of the urinary outflow. PMID- 15654613 TI - [Invasive techniques in emergency medicine. IV. Cricothyrotomy in emergency situations]. AB - Cricothyrotomy is a very invasive technique to secure the airway in an emergency but is irreplacable when less invasive techniques fail or cannot be instigated under the prevailing circumstances. Various techniques have been reported which can be subdivided into anatomical-surgical preparation or puncture techniques. The preferred strategy is mostly oriented towards the departmental standard procedure which will be decided by the clinical situation. Training for each procedure can be carried out in intensive care departments, and using autopsy material or a manekin. Various methods of cricothyrotomy will be discussed here, and additionally an anatomical preparation and two puncture techniques will be demonstrated in detail. PMID- 15654615 TI - A novel out-of-frame mutation in the neurofilament light chain gene (NEFL) does not result in Charcot-Marie-Tooth disease type 2E. PMID- 15654614 TI - Investigation of hormone receptor genes in migraine. AB - Migraine is a common neurological condition with a complex mode of inheritance. Steroid hormones have long been implicated in migraine, although their role remains unclear. Our investigation considered that genes involved in hormonal pathways may play a role in migraine susceptibility. We therefore investigated the androgen receptor (AR) CAG repeat, and the progesterone receptor (PR) PROGINS insert by cross-sectional association analysis. The results showed no association with the AR CAG repeat in our study group of 275 migraineurs and 275 unrelated controls. Results of the PR PROGINS analysis showed a significant difference in the same cohort, and in an independent follow-up study population of 300 migraineurs and 300 unrelated controls. Analysis of the genotypic risk groups of both populations together indicated that individuals who carried the PROGINS insert were 1.8 times more likely to suffer migraine. Interaction analysis of the PROGINS variant with our previously reported associated ESR1 594A variant showed that individuals who possessed at least one copy of both risk alleles were 3.2 times more likely to suffer migraine. Hence, variants of these steroid hormone receptor genes appear to act synergistically to increase the risk of migraine by a factor of three. PMID- 15654617 TI - Selective immunoglobulin A deficiency and primary antiphospholipid syndrome. PMID- 15654616 TI - The predictive value of additional late blood pool imaging to the three-phase bone scan in the diagnosis of reflex sympathetic dystrophy in hemiplegic patients. AB - Reflex sympathetic dystrophy (RSD) is a relative common sequel after hemiplegia. The diagnosis of RSD in hemiplegic patients presents difficult clinical problems, as the symptoms and signs of RSD are not specific and RSD may be due to reasons other than hemiplegia. Bone scintigraphy has been routinely used for the diagnosis of RSD; however, the optimal acquisition protocols, diagnostic patterns and the utility of quantitation are controversial. This prospective study was conducted to demonstrate the higher predictive value of an additional late blood pool image to the three-phase bone scan compared to the regular three-phase bone scans in RSD patients associated with hemiplegia. Thirty-four RSD patients were enrolled into the study. Bone scans according to the new protocol were obtained for all patients. Those patients with either negative or positive bone scans with no evidence of RSD were followed for 6 months. The patients had positive bone scan findings and were symptomatic at the time of the study. Of these, seven patients (58.3%) subsequently became symptomatic and five patients (41.7%) remained asymptomatic at 6 months. None of the patients with negative bone scans had symptoms of RSD on presentation except one case. We conclude that the addition of a late blood pool image increases the predictive value and has an impact on initiating early treatment in asymptomatic patients. PMID- 15654619 TI - Reducing by-product formation in L-lactic acid fermentation by Rhizopus oryzae. AB - During L-lactic acid fermentation by Rhizopus oryzae, increasing the phosphate level in the fermentation medium from 0.1 g l(-1) to 0.6 g l(-1) KH2PO4 reduced the maximal concentration of L-lactic acid and fumaric acid from 85 g l(-1) to 71 g l(-1) and from 1.36 g l(-1) to 0.18 g l(-1), respectively; and it decreased the fermentation time from 72 h to 52 h. Phosphate at 0.40 g l(-1) KH2PO4 was suitable for both minimizing fumaric acid accumulation and benefiting L-lactic acid production. PMID- 15654618 TI - Comparison of cardiopulmonary response to endogenous nitric oxide inhibition in pigs inhabited at three levels of altitude. AB - Nitric oxide (NO) plays an important role for the pulmonary circulation in normal and chronic hypoxia. We examined effects of endogenous nitric oxide synthase (NOS) inhibition on pulmonary and systemic vascular resistance in unanesthetized pigs living at three levels of altitude to evaluate the role of NO in adaptation to a hypoxic environment. Unanesthetized male adult pigs in three areas [Matsumoto, Japan (680 m above sea level, n = 5); Xing, China (2,300 m, n = 5); and Maxin, China (3,750 m, n = 5)] were prepared for vascular monitoring. Pulmonary (P(pa)), and systemic artery pressure (P(sa)) were monitored, and pulmonary artery wedge pressure (P(cwp)) and cardiac output (CO) were measured before and after treatment with a non-selective NOS inhibitor, N(w)-nitro-L argine (NLA; 20 mg/kg). Pulmonary vascular resistance (PVR) and systemic vascular resistance (SVR) were (P(pa)-P(cwp))/CO and P(sa)/CO, respectively. Related to altitude baseline P(pa) was elevated. After NLA administration, P(pa) and P(sa) increased and CO decreased in all animals, resulting in increases in PVR and SVR. However, there were no significant differences in the increase in PVR and SVR in the three groups of pigs. Thus, endogenous NO production contributes to regulate the basal pulmonary vascular tone, but the development of hypoxic pulmonary hypertension appears to be independent of the NO pathway in adult pigs. PMID- 15654620 TI - Multiple de novo vascular malformations in relation to diffuse venous occlusive disease: a case report. AB - Brain vascular malformations are dynamic disorders. Although mostly considered to be of congenital origin, the improvement of clinical imaging and vasculogenesis knowledge has shown that they might also result from a biological dysfunction of the remodeling process after birth. Venous occlusive disease and ishemia may represent powerful revealing triggers and support the capillary venous origin of some vascular malformations. We report a unique case of the development of multiple de novo vascular malformations (transverse sinus dural fistula and posterior fossa cavernomas) following acoustic neuroma surgery. PMID- 15654622 TI - Molecular characterization of ONAC300, a novel NAC gene specifically expressed at early stages in various developing tissues of rice. AB - Members of the plant-specific gene family referred to as the NAC family (for NAM ATAF-CUC-related) are involved in various functions including the regulation of plant development. However, no detailed molecular characterization of any member of the NAC family has yet been reported from monocots. Here, we report such a characterization of ONAC300, a novel NAC-family gene identified using a cDNA cloned from microdissected phloem cells of rice. The predicted ONAC300 protein sequence falls into the NAM subgroup, which also contains the proteins CUC1 and CUC2 from Arabidopsis, CUP from snapdragon, CmNACP from pumpkin and NAM from petunia. High levels of ONAC300 mRNA were detected by in situ hybridization in developing shoot apical meristem (SAM) and in the associated young leaves. The use of an ONAC300:: GUS reporter gene revealed that the ONAC300 promoter was expressed predominantly in developing vascular tissues of the leaves and roots. The construct was also expressed in anther filaments, rachis and carpel styles. RT-PCR analysis further revealed that the levels of ONAC300 transcripts were higher in leaves, roots and culms than in panicles. The observed expression pattern of ONAC300 is quite different from those of the dicot NAC genes previously reported. Thus, ONAC300 is a novel member of the NAC family which is expressed at very early developmental stages in the shoot, root and flower, as well as in the mature phloem of vascular tissues in rice. PMID- 15654621 TI - Two cis-acting regulatory elements are involved in the sucrose-inducible expression of the sporamin gene promoter from sweet potato in transgenic tobacco. AB - In this study, we generated transgenic tobacco plants that express the beta glucuronidase (GUS) gene under the control of the 305-bp 5'-upstream region of a gene coding for sporamin A of sweet potato. Expression of GUS in excised tobacco leaves was induced by sucrose, mimicking the sugar-inducible expression of the endogenous sporamin genes in sweet potato. Deletion of the sequences extending from position -305 (relative to the transcription start site) to -283 and from 146 to -87 resulted in an approximately 40-fold enhancement in GUS reporter expression. Furthermore, the sequence from -282 to -165 conferred sucrose inducibility on the -89 core promoter of the Cauliflower Mosaic Virus 35S RNA gene. Analysis of internal deletions, linker scanning and the introduction of base substitutions in the sequence between positions -282 and -165 indicated that sucrose-responsiveness conferred by this region was dependent on the presence of two cis-acting elements, termed CMSREs (carbohydrate metabolite signal responsive elements) 1 and 2, which are separated by a spacer. A sequence similar or identical to the core of CMSRE-1 (TGGACGG) is also present in the promoters of several other sugar-inducible genes, and sequences encopassing the TGGACGG related motifs from two of these could functionally replace the CMSRE-1 in the truncated sporamin A promoter. These results suggest that the TGGACGG element plays an important role in the sucrose-inducible expression of a group of plant genes. PMID- 15654623 TI - Characterisation of Aspergillus niger prolyl aminopeptidase. AB - We have cloned a gene (papA) that encodes a prolyl aminopeptidase from Aspergillus niger. Homologous genes are present in the genomes of the Eurotiales A. nidulans, A. fumigatus and Talaromyces emersonii, but the gene is not present in the genome of the yeast Saccharomyces cerevisiae. Cell extracts of strains overexpressing the gene under the control of its own promoter showed a fourfold to sixfold increase in prolyl aminopeptidase activity, but no change in phenylalanine or leucine aminopeptidase activity. The overexpressed enzyme was subsequently purified and characterised. The enzyme specifically removes N terminal proline and hydroxyproline residues from peptides. It is the first enzyme of its kind from a eukaryotic organism that has been characterised. PMID- 15654624 TI - p53 polymorphisms in Russia and Belarus: correlation of the 2-1-1 haplotype frequency with longitude. AB - Four different polymorphisms in the human p53 gene (a 16-bp duplication in intron 3, and three RFLPs: for Bsh1236I at codon 72, for MspI in intron 6 and for BamHI in the 3' flanking region) and extended haplotypes were studied in nine geographically diverse populations from Russia and Belarus. The Yakuts differed from all other populations, as they had a significantly higher frequency of the BamHI A1 allele. Most populations did not differ significantly from each other in the frequency of the Bsh1236I polymorphism. The 16-bp duplication A1 allele and MspI A2 allele frequencies were significantly higher in the Yakut and Khant populations. Linkage disequilibrium values (D') between BamHI and other polymorphic sites were not significant in many cases; for this reason we have used the 16 bp-Bsh1236I-MspI haplotype frequencies only. Of eight possible haplotypes, five were observed in the populations investigated. Haplotype 1-2-2 was the most frequent in all populations. The next most common haplotype, 1-1-2, was present at very similar frequencies among the Byelorussians and Russians from Smolensk, but was more frequent in other populations. The frequency of haplotype 2-1-1 showed a nearly continuous decrease from West to East (from 17.857% among the Byelorussians to 0.685% in the Yakuts from the Verkhoyansk) and correlated with longitude (Spearman's r = -0.8667, P = 0.0025), which may be due to natural selection and adaptation. The relationships among populations were evaluated by means of Nei's D(A) distances for the 16 bp-Bsh1236I-MspI haplotype frequencies. Based on the multidimensional scaling analysis a correlation between p53 haplotype frequencies and ethnicity is supposed. PMID- 15654625 TI - Allelic expression of IGF2 in live-bearing, matrotrophic fishes. AB - The parental conflict, or kinship, theory of genomic imprinting predicts that parent-specific gene expression may evolve in species in which parental investment in developing offspring is unequal. This theory explains many aspects of parent-of-origin transcriptional silencing of embryonic growth regulatory genes in mammals, but it has not been tested in any other live-bearing, placental animals. A major embryonic growth promoting gene with conserved function in all vertebrates is insulin-like growth factor 2 (IGF2). This gene is imprinted in both eutherians and marsupials, as are several genes that modulate IGF2 activity. We have tested for parent-of-origin influences on developmental expression of IGF2 in two poeciliid fish species, Heterandria formosa and Poeciliopsis prolifica, that have evolved placentation independently. We found IGF2 to be expressed bi-allelically throughout embryonic development in both species. PMID- 15654626 TI - The transmembrane protein, Tincar, is involved in the development of the compound eye in Drosophila melanogaster. AB - We previously cloned and characterized the Drosophila gene, tincar (tinc), which encodes a novel protein with eight putative transmembrane domains. Here, we have studied the expression pattern and functions of tinc during developmental processes. tinc mRNA is expressed in the central and peripheral nervous systems, and midgut during embryogenesis. In the third-instar larval eye disc, tinc mRNA is strongly expressed in all the differentiating ommatidial cells within and in the vicinity of the morphogenetic furrow. Loss-of-function analysis using the RNA interference method revealed severe defects of eye morphogenesis during the late developmental stages. Our results suggested that tinc may have an indispensable role in the normal differentiation of ommatidial cells. PMID- 15654627 TI - Mouse serum factor(s) down-modulate the CD4 and CXCR4 molecules on human T cells conferring resistance to HIV infection in NOG mice. AB - Human cells have developed innate immunity, exploiting several means to block virus infection, and viruses have evolved diverse strategies to resist these. We show here that the human immunodeficiency virus 1 (HIV-1) could neither progressively infect engrafted human leukemic T cells nor repress their growth in NOG mice. However, ED-40515(-) cells infected with HIV-1 before inoculation were found to significantly delay the onset of tumor growth and increased the survival period of NOG mice. ED-40515(-) tumor cells showed resistance to HIV-1 which was apparently correlated with the down-regulation of CD4 and CXCR4 molecules in NOG mice. Serum from three different mouse strains, including NOG, retained a suppressive effect on the CD4 molecule of ED-40515(-) cells in vitro. ED-40515(-) cells obtained from mice re-expressed CD4 and CXCR4 molecules upon in vitro culture and were again successfully infected with HIV-1. These findings indicate that HIV-1 may initially successfully delay or regress tumor growth in NOG mice, but eventually fails to do so because of the evolution of HIV-resistant cells due to a rapid down-modulation of CD4 and CXCR4. Our data also demonstrated that some unknown soluble factor(s) present in mouse serum was responsible for conferring resistance to HIV infection to human T cells. PMID- 15654628 TI - Usefulness of a latex agglutination assay for FDP D-dimer to demonstrate the presence of postmortem blood. AB - D-dimer, a specific fragment resulting from degradation of cross-linked fibrin, is an essential marker for the diagnosis of disseminated intravascular coagulation (DIC). Rapid assay for D-dimer using monoclonal antibody coated-latex particles might be useful for discriminating between postmortem and antemortem blood in bloodstains. We tried to detect D-dimer in nine postmortem blood samples by the rapid latex agglutination assay and to quantify them automatically using the latex photometric immunoassay system. The results showed that all samples were positive and that their amounts of D-dimer were 335-2,800 microg/ml (the normal blood level, <1 microg/ml; the pathogenic blood level with DIC, 1-100 microg/ml). Next, nine stains made of postmortem blood were examined by the rapid latex agglutination assay. The result showed that only one case (D-dimer 335 microg/ml blood) showed weak positive while the others (D-dimer 600-2,800 microg/ml blood) were positive. The present study indicates that the latex agglutination assay for D-dimer can be useful to demonstrate the presence of postmortem blood. PMID- 15654630 TI - Thalamic ependymal cyst presenting with tremor. AB - CASE REPORT: We report the case of a 6-year-old boy presenting with intention tremor. CT and MR scanning showed a large thalamic cyst. TREATMENT: Stereotactic tapping of this cyst was carried out as the first step in surgical management. Following drainage of the cyst the tremor disappeared, but recurred 18 months later. Repeat MRI showed that the thalamic cyst had refilled. As the initial tapping of the cyst had resulted in effective but only transient clinical improvement a cysto-peritoneal shunting procedure was performed. OUTCOME: The clinical symptoms again disappeared and the child has now remained tremor free for 3 years. PMID- 15654629 TI - Frequency-dependent potentiation of voltage-activated responses only in the intact neurohypophysis of the rat. AB - The loose-patch-clamp technique was used with multiple-pulse protocols to study the frequency dependence of currents from the surface of the intact rat neurohypophysis (NH) and hypothalamus. In the NH, but not in the corresponding supraoptic nucleus of the hypothalamus, an initial, single pulse of 3-8 ms duration (long pulse) potentiated a secondary pulse response starting 20-50 ms after the initial pulse. Potentiation was abolished by 4-aminopyridine (4-AP), but not by tetraethylammonium (TEA) chloride or tetrandrine, indicating the participation of A-type potassium currents. Potentiation was also abolished by CdCl2, CoCl2 or 1 microM nicardipine, indicating the participation of calcium currents. The potentiation was reduced significantly in the presence of 4-6 mM extracellular CaCl2, indicating that the potentiation is not due to calcium influx. An initial train with as few as two pulses, each of 0.3-0.7 ms duration (short pulses) at 64-1,100 Hz also potentiated the secondary short pulse response significantly. We conclude that voltage-gated channels underlie this potentiation, which is due to interstitial calcium and potassium homeostasis changes induced by action potential activity and occurs only in the intact NH. A model is proposed for the participation of calcium and potassium channels in the burst patterning that is optimal for secretion from the NH. PMID- 15654631 TI - Total callosotomy for a case of lissencephaly presenting with West syndrome and generalized seizures. AB - CASE REPORT: An 11-month-old girl had an onset of oculogyric crisis at 2 months and she presented with epileptic spasms and generalized tonic seizures with series formation at 3 months. Her seizures were medically intractable and her development had gradually regressed after that. MRI showed severe lissencephaly in bilateral hemispheres. Interictal EEG showed hypsarrhythmia periodically and sporadic spike waves in the right hemisphere. The generalized tonic seizures began with bi-frontal polyspikes on EEG. Our diagnosis was lissencephaly presenting with West syndrome and generalized tonic seizures. A total callosotomy was performed at 11 months. OUTCOME: Postoperative frequency of seizures decreased prominently and developmental progression was resumed. CONCLUSIONS: This case showed that the corpus callosum may play an important role in some types of symptomatic West syndrome. Corpus callosotomy for treating symptomatic West syndrome should be considered as an option after careful selection and consideration of the timing of surgery. PMID- 15654632 TI - Operative indications and neuroendoscopic management of symptomatic cysts of the septum pellucidum. AB - OBJECTIVE: Since there is no current consensus on the diagnostic standards or surgical indications for symptomatic cysts of the septum pellucidum (CSP), we presented our experience treating such cases with neuroendoscopic surgery in order to possibly make a contribution to the attainment of such a consensus. METHODS: In the last 5 years, we effectively treated 5 patients (4 males and 1 female) with CSP by neuroendoscopic surgery. The 3 congenital cases were young: 9, 14, and 29 years old. The 2 secondary cases were adults aged 51 and 65 years old. We were able to fenestrate, and biopsy for pathological analysis, the cyst walls of symptomatic CSP using a minimally invasive neuroendoscopic technique, with postoperative improvement in compression of adjacent structures, as well as clinical signs. CONCLUSIONS: It is necessary to consider the overall picture including clinical signs, progress of the condition, intracranial pressure, and cerebral circulation, in deciding whether surgery is indicated. Neuroendoscopic surgery is an effective, optimal, and convenient therapeutic modality for symptomatic CSP. PMID- 15654634 TI - Leksell gamma knife radiosurgery for cerebral arteriovenous malformations in pediatric patients. PMID- 15654633 TI - Astroblastoma in childhood: pathological and clinical analysis. AB - OBJECT: The object was to describe the clinical, radiologic, and pathologic features of astroblastomas in an unselected group of children who were treated in a single institution during an 11-year period. METHODS: Eight children with astroblastomas of the brain were examined. Diagnosis was based on cell morphology, vascular attachment of the cell main process, lack of an epithelial free surface differentiation, and poor intercellular cohesiveness. In addition to sections, tumor smears and electron microscopy were required for demonstrating or confirming such features. CONCLUSIONS: Clinical findings seem to confirm an apparent predilection of astroblastomas for younger children (median age of onset, 5 years) and the existence of two prognostically different types of tumor well differentiated (low grade) and anaplastic (high grade). Microscopic findings suggest a closer resemblance of tumor cells to astroblasts rather than to "tanycytes" or ependymal cells. It seems, however, that anaplastic astroblasts have a tendency to evolve toward, or be associated with, less differentiated cells, either neuroepithelial or sarcomatous. PMID- 15654635 TI - Leksell gamma knife radiosurgery for cerebral arteriovenous malformations in pediatric patients. AB - OBJECTS: The authors report their experience of gamma knife radiosurgery (GKR) in a large series of pediatric cerebral arteriovenous malformations (cAVMs). The advantages, risks and failures of this approach are presented and discussed. METHODS: Gamma knife radiosurgery was performed on 63 children aged < or =16 years. Haemorrhage was the clinical onset in 50 out of 63 cases. The mean pre-GK cAVM volume was 3.8 cm(3). Fifty-eight out of 63 cAVMs were Spetzler-Martin grades I-III. Most lesions (47 out of 63) were in eloquent or deep-seated brain regions. CONCLUSION: Gamma knife radiosurgery-related complications occurred in 2 out of 47 cases with an available follow-up (1 had transient and 1 permanent morbidity). No bleeding occurred during the latency period. In 39 children with >36-month follow-up, complete cAVM occlusion was angiographically documented in 31, with a 3- and 4-year actuarial obliteration rate of 72 and 77% respectively. High rates of complete obliteration and very low frequency of permanent morbidity with no bleeding during the latency period encourage widespread application of GKR in the treatment of pediatric cAVMs. PMID- 15654636 TI - Cloning and expression of a trehalose synthase from Pseudomonas stutzeri CJ38 in Escherichia coli for the production of trehalose. AB - A novel strain was isolated, Pseudomonas stutzeri CJ38, that enabled direct transformation of maltose to trehalose. In comparison with others reported to date, CJ38 provided a novel trehalose synthase (TSase) without any byproduct, including glucose. Activity analysis, using either maltose or trehalose as a substrate, showed a reversible reaction. There was also no detectable activity of related enzymes with liquid starch and maltooligosaccharides as substrates. Using a malPQ-negative host and MacConkey medium, the TSase gene was cloned in Escherichia coli from CJ38. The resulting sequence contained an open reading frame consisted of 689 amino acids with a calculated molecular mass of 76 kDa. A search for related sequences in various gene and protein data banks revealed a novel family of enzymes that was predicted putatively as a glycosidase or TSase family, with no biochemical evidence. The recombinant enzyme exhibited a high activity toward the substrate maltose, about 50-fold higher than the parent strain and resulted in a high conversion yield (72%) at a relatively high substrate concentration (20%). These results provided the possibility that the strain was effectively used as a potential biocatalyst for the production of trehalose from maltose in a one-step reaction. PMID- 15654637 TI - The two nitrogen mobilisation- and senescence-associated GS1 and GDH genes are controlled by C and N metabolites. AB - In tobacco, the two enzymes of nitrogen metabolism, cytosolic glutamine synthetase (GS1; E.C.6.3.1.2) and glutamate dehydrogenase (GDH; E.C.1.4.1.2), are induced during leaf senescence, whereas the chloroplastic glutamine synthetase (GS2; E.C.6.3.1.2) and nitrate reductase (NR; E.C.1.6.1.1) are repressed in the course of ageing. In this report, we showed in discs of fully expanded Nicotiana tabacum L. cv. Xanthi leaves that sucrose (Suc) and amino acids were involved in the regulation of the expression of GS1 and GDH genes. Suc induced the expression of GS1 and repressed that of GDH. Therefore, we concluded that in response to Suc, GS1 behaved as an "early" Senescence Associated Gene (SAG), whereas GDH behaved as a "late" SAG. Moreover, amino acids induced the expression of both genes. Among the amino acids tested as signal molecules, proline (Pro) and glutamate (Glu) were major inducers of GDH and GS1 expression, respectively. Interestingly, an opposite regulation of GS1 and GS2 by Pro and Glu was shown. The contrary effect of Suc on NIA (NR encoding gene) and GDH mRNA accumulation was also emphasized. PMID- 15654639 TI - Micropropagation of squill (Charybdis numidica) through nodule culture. AB - A micropropagation protocol for squill (Charybdis numidica, Hyacinthaceae) was developed using nodule culture. Nodule formation on leaf sections was induced in liquid Murashige and Skoog (MS) medium supplemented with 20 microM N6 benzylaminopurine (BA) under dark conditions. Nodules were cultured on semi-solid MS medium with factorial combinations of BA (0-40 microM) and alpha naphthaleneacetic acid (NAA) (0-10 microM) under continuous light. Shoot regeneration from nodules occurred at varying degrees on all media. The highest number of shoots was formed on medium containing 2.5 microM NAA and 20 microM BA, while the maximum number of regenerated bulblets per gram nodule was induced on culture medium supplemented with 2.5 microM NAA alone. Regenerated shoots were successfully rooted at approximately 92% on semi-solid MS medium supplemented with 10 microM indole-3-acetic acid (IAA). Plantlets could be hardened and grew well after transfer to the greenhouse. Chemical analyses showed consistent bufadienolide patterns from cloned plantlets and the mother plant. PMID- 15654640 TI - Local-scale density-dependent survival of mobile organisms in continuous habitats: an experimental test using Atlantic salmon. AB - For organisms with restricted mobility, density dependence may occur on spatial scales much smaller than that of the whole population. Averaging densities over whole populations in such organisms gives a more or less inaccurate description of the real variation in competitive intensity over time and space. The potential for local density dependence in more mobile organisms is less well understood, particularly for organisms living in continuous habitats. To test for local density-dependent processes in such an organism, we manipulated egg density (the number of eggs nest(-1)) among ten artificial nests of Atlantic salmon along an 1,848-m long river during two consecutive years. Eggs in different nests were given unique thermal otolith-banding patterns to allow identification of juvenile nest origin. At capture, 1-2 months after emergence, the spatial distribution of juveniles reflected nest locations, with the median absolute dispersal distance being 92 and 41 m in the 2 years. Estimated nest-specific survival rates were strongly negatively related to hatched-egg density in both years (r(2)=0.72 and 0.62), despite dramatic differences in overall mean survival (0.22 and 0.02). Thus, density-dependent survival following emergence in Atlantic salmon juveniles occurs on spatial scales much smaller than that of whole populations. The consistency across years suggests that the phenomenon is likely to occur over most environmental conditions. Our observation of local-scale density dependence is consistent with strong juvenile territoriality, which forces individuals emerging in high-initial density areas to disperse farther, and a high cost (metabolic or predation) of dispersal. We conclude that for mobile organisms with patchy distributions of propagules and constrained juvenile dispersal, increased emphasis on local-scale dynamics should enable a more mechanistic understanding of population regulation even in continuous habitats, and hence increase the predictive power of population models. PMID- 15654638 TI - Identification of pathogen-responsive regions in the promoter of a pepper lipid transfer protein gene (CALTPI) and the enhanced resistance of the CALTPI transgenic Arabidopsis against pathogen and environmental stresses. AB - The 5' flanking region of the CALTPI gene, which encodes a basic lipid transfer protein, was isolated and characterized from the genomic DNA of Capsicum annuum. Four different regions of the promoter sequence of the CALTPI gene were fused to the beta-glucuronidase (GUS) coding region. In an Agrobacterium-mediated transient expression assay, the transcriptional activations of the promoter deletions were examined in tobacco leaves after infection with Pseudomonas syringae pv. tabaci, and treatment with ethylene and salicylic acid. The -808 bp region of the CALTPI gene promoter sequence exhibited full promoter activity. The W-box and ERE-box elements, which are essential for induction by all signals, were localized in the region between -555 bp and -391 bp upstream of the translation initiation site. A CALTPI transgene was then introduced under the control of the 35S promoter into the Arabidopsis ecotype Col-0. Transgenic Arabidopsis lines expressing the CALTPI gene developed rapidly compared to the wild-type plants, indicating that CALTPI may be involved in plant development. Overexpression of the CALTPI gene enhanced the resistance against infection by P. syringae pv. tomato and Botrytis cinerea. The transgenic plants expressing the CALTPI gene also showed high levels of tolerance to NaCl and drought stresses at various vegetative growth stages. No transcription of the PR-1, PR-2, PR-5, thionin, and RD29A genes was observed in untreated leaf tissues of the transgenic plants. The enhanced resistance to pathogen and environmental stresses in transgenic Arabidopsis correlated with the enhanced expression of the CALTPI gene. PMID- 15654641 TI - Correlation between the current TNM staging and long-term survival after curative D1 lymphadenectomy for stomach cancer. AB - BACKGROUND AND AIMS: This study aims to assess the tumour-related factors that influence long-term survival after curative gastrectomy with standard D1 lymphadenectomy for patients with stomach cancer. PATIENTS AND METHODS: Patients who had undergone curative gastrectomy for carcinoma of the stomach at North Shore Hospital between 1990 and 2000 were identified from theatre records and the hospital database. Medical records were reviewed and included tumour location, type of operation, in-hospital mortality, gross morphology of tumour, histological type, and Helicobacter status; pathology slides were reviewed, and tumours were staged according to the new TNM staging. Patients were followed-up for 2-11 years. Length of survival was obtained for each patient from medical records or from family doctors. RESULTS: R0 gastrectomy was performed on 70 patients; median survival was 23 months, and all patients with early gastric cancer are currently still alive. T stage, nodal stage and histological type correlated significantly with survival, but multivariate analysis showed that T stage is the most significant predictor. Five-year survival was 26%. Significant survival difference was seen between T2a and T2b. CONCLUSION: Histological subtype, lymph node metastases and depth of invasion are factors that affect survival of patients with gastric cancer; however, depth of invasion is more important than other variables. Tumour location and type of gastrectomy has no effect on survival. The latest TNM classification (sixth edition) gives a better prognostication than the previous classification. PMID- 15654643 TI - Isolated pelvic perfusion: plasma pharmacokinetics depend primarily on drug dosage and not the type of drug. AB - PURPOSE: Comparison of the pharmacokinetics of four drugs with the isolated pelvic perfusion protocol showed linear relationships between drug dosage and two isolated pelvic plasma parameters, mean AUC (pelvic exposure, microM min) and the mean maximum pelvic drug level (microM). It appears that the pharmacokinetics are sufficiently defined as to predict plasma distribution curves for an additional drug with this protocol. Recent FDA approval of oxaliplatin allowed an evaluation of this premise. METHODS: Linearity of drug dosage with maximum drug levels and exposure (AUC) in the isolated pelvic plasma yields initial estimates of these parameters for additional drugs. Use of an empirical, four-compartment pharmacokinetic model (Wanebo and Belliveau in Cancer Chemother. Pharmacol. 43:427, 1999) allowed the generation of predictive plasma distribution curves. These curves were established by optimizing the initial estimates of maximum drug levels and exposure along with estimates of two additional parameters (half-life of pelvic clearance and pelvic to systemic exposure ratio) from experimental data of the four drugs pharmacokinetically characterized. RESULTS: Calculated plasma distribution curves for oxaliplatin matched the experimental curves from the first three patients receiving oxaliplatin therapy, given the experimental ranges of pharmacokinetic parameters seen with the initial four drugs. CONCLUSION: These results give an overall picture for the plasma pharmacokinetics during the isolation period for the isolated pelvic perfusion protocol. Enough experimental data have been accumulated for five drugs to establish a simple pharmacokinetic model (Wanebo and Belliveau in Cancer Chemother Pharmacol 43:427, 1999) and interdrug relationships (i.e., this report) which can be used to predict reasonable plasma distribution curves for additional drugs with this protocol. PMID- 15654642 TI - Surgical treatment for multinodular goitres in geriatric patients. AB - BACKGROUND: Although age is not a contraindication for thyroid surgery, few elderly patients undergo surgery due to the greater risk of morbidity. The aims of this study are to determine in patients aged >65 years: (1) whether the indications for surgery on multinodular goitre (MG) differ with respect to younger patients; (2) the surgical results; and (3) whether the postsurgery morbidity and mortality rates are higher. PATIENTS AND METHOD: Eighty-one patients aged over 65 years who were receiving surgery for MG were analysed; 40 49%) presented with associated co-morbidities. Sixty percent had thyroid symptoms, either compressive and/or toxic. All underwent programmed surgery following stabilisation and strict control of their co-morbidities. As a control group we used 510 MG patients receiving surgery and aged between 30 and 65 years. RESULTS: Compared with the control group the geriatric patients had a longer time of goitre evolution (P=0.032), greater presence of symptoms (P=0.001) and a higher percentage of intrathoracic component (P=0.001). Compressive symptoms were the major indication for surgery (P=0.001). Postoperative complications occurred in 40% of the patients, a higher rate than in the control group (28%; P=0.011), although a large percentage of those complications were transitory. Definitive complications included two recurrent laryngeal nerve injuries (2.5%). The preoperative symptoms remitted in all the patients, and only three were associated with a thyroid carcinoma, one of which was anaplastic. CONCLUSIONS: MG operated on in elderly patients has a longer evolution and an intrathoracic component, and surgery is indicated restrictively. With close monitoring of the co-morbidities and a programmed operation the results with regard to morbidity and mortality are similar to those obtained at younger ages. PMID- 15654646 TI - Intra-articular cysts and ganglia of the knee: a report of nine patients. AB - Completely intra-articular cysts and ganglia of the knee are rare. They have been found in various locations such as on the anterior or posterior cruciate ligaments, in the infrapatellar fat pad, on the posterior wall of the posteromedial compartment and (very rarely) in connection to the menisci. We analyzed nine patients with intra-articular cysts or ganglia found in a series of 2,400 consecutive arthroscopies. In four patients, the cyst or ganglion was found attached to the anterior part of the ACL, in two patients it was located between the ACL and the PCL, and in the remaining three cases it was found in connection with the meniscus. In three out of the nine patients there was either no or very minor additional pathology found in the knee besides the cyst or the ganglion. We believe that intra-articular cysts and ganglia of the knee can be symptomatic, and excellent or good results after cyst removal can be expected especially when there is little additional pathology. PMID- 15654644 TI - Endothelin receptor antagonists. AB - Hormone refractory prostate cancer remains true to its middle name: it is largely refractory to attempts to delay its progression. New targets and new therapies are demanded. Through a review of the available literature on endothelin and several preclinical observations, the endothelin axis has emerged as one such target. In phase II and III clinical trials of atrasentan, a potent and selective endothelin receptor A subtype (ET(A)) antagonist, disease progression was delayed in some men. This well tolerated, oral agent may help convert advanced prostate cancer to a more chronic disease. This review will discuss the endothelin axis, preclinical rationale and some of the available clinical trial data on this promising new approach. PMID- 15654645 TI - Comparison of the Insall-Salvati ratio of the patella in patients with and without an ACL tear. AB - The object of this prospective study is to compare the Insall-Salvati ratio between the patients who have an anterior cruciate ligament (ACL) tear and receive arthroscopic-assistant ACL reconstruction and the patients who have no ACL tear but do have an internal disorder of the knee and receive arthroscopic surgery. We prospectively and consecutively collected into two groups a total of 217 patients who had sport injuries and received arthroscopic surgery. The study group included 115 patients who had an ACL tear and received arthroscopic assistant ACL reconstruction with middle-third bone-patella tendon-bone graft. The control group included 102 patients with internal disorders of the knee joint, including meniscus tear, plicae, or other chondral lesion, but without an ACL tear. We measured the patellar Insall-Salvati ratio [12] on the pre-operative X-ray films for all patients. The Insall-Salvati ratio in the ACL-tear study group is significantly smaller than the control group of internal disorders of the knee (0.99+/-0.11 vs 1.05+/-0.12, p=0.001). There is no significant difference in age, gender, the side of the involved knee, duration of symptoms, patella length and patella tendon length between the two groups. In conclusion, our study shows that patella infra has an association with ACL tears, and patella infra may be a risk factor for ACL tears. In patients with an ACL tear who had patella baja, the middle-third patellar tendon may not be an ideal graft for reconstruction. PMID- 15654648 TI - [Where are the 1.9 million patients dependent on legal drugs hiding?]. AB - Although the number of patients dependent on legal drugs is estimated to be as high as 1.9 million, very few are in clinical treatment. Research deficits in this area are evident. PMID- 15654647 TI - [Psychological stress in patients with environmentally-induced disorders. A comparison between self concept and expert assessment]. AB - Environmental illnesses raise diagnostic and therapeutic conflicts in scientific discussions and clinical practice. When a patient's health-belief model, based on environmental origins, does not match that of the expert, the therapeutic relationship can be endangered. Our study investigates this discrepancy, which has not been empirically evaluated so far. Patient (n=61) and expert disease concepts were systematically investigated. Our results indicate that in cases in which both concepts are favourable, the patient suffered minor psychiatric disorders with stable psychic structures and the symptoms were associated with medical or environmental causes. If both concepts were unfavourable, a higher proportion of psychiatric disorders with unstable psychic structures were present. In the case of incongruent concepts, the expert evaluations allow a more accurate assessment of the psychiatric diagnoses, psychic states and the psychic attribution of somatic and psychic burden. PMID- 15654651 TI - Subcellular vesicular aggregations of GABAB R1a and R1b receptors increase with age in neurons of the developing mouse brain. AB - GABA(B) receptors play a critical neuromodulatory role in the central nervous system. It has been suggested that both the functional role and the cellular distribution of GABA(B) receptors in the neuronal network change during post natal maturation. In the present study, the cellular and subcellular distribution patterns of the GABA(B) R1a/b receptors have been analysed in different brain regions of the mouse using immunocytochemistry with isoform-specific antisera. GABA(B) R1-immunoreactivity (IR) was present from the first post-natal day (P0) on in most regions of the brain. Neurones exhibited diffuse GABA(B) R1-IR labelling throughout somata and larger proximal dendrites as well as some fine neuronal processes. After P5, distinct punctuated staining was apparent. The number of such GABA(B) IR granules per cell increased with age in a sigmoidal manner from P5 to P60. Electron microscopy revealed GABA(B) IR as clusters of small clear vesicles of 30-50 nm diameter within the cytoplasm and close to the cell membrane at extrasynaptic locations, as well as at pre-synaptic and post synaptic specialisations. The increase in GABA(B) R1-IR punctuate staining during brain maturation points to increasing functional participation and heterogeneity of GABA(B) receptors as the complexity of the central nervous system expands with growth and development. PMID- 15654652 TI - alpha-Gustducin immunoreactivity in the airways. AB - The G-protein subunit alpha-gustducin is a marker of chemoreceptive cells. In the present study, we examined the immunohistochemical localization of alpha gustducin in rat airway epithelium both by light and electron microscopy. alpha Gustducin immunoreactivity was found in solitary cells that presented ultrastructural features of chemoreceptor cells, i.e. flask-shaped or pear shaped, with an apical process with thin microvilli protruding into the lumen. The immunostaining was mainly concentrated in the apical process and along the basolateral cell surface. To investigate whether alpha-gustducin-immunoreactive cells represented a distinct cell subset in rat airways, we performed double label immunocytochemistry with antibodies to protein gene groduct (PGP) 9.5, a marker of neuroendocrine cells, and to phospholipase C beta2 (PLCbeta2), a component of the bitter signalling pathway. alpha-Gustducin-immunoreactive cells were present in a subset of PGP-9.5-immunoreactive elements, although not all alpha-gustducin-positive cells expressed PGP 9.5 labelling. In addition, a subset of alpha-gustducin-expressing cells colocalized PLCbeta2. This work thus demonstrates that solitary alpha-gustducin-immunoreactive cells exist throughout the airways and represent a specialized cell type with morphological and immunohistochemical characteristics of chemoreceptor cells. PMID- 15654653 TI - Estrogenic restoration of functional pancreatic islet cytoarchitecture in diabetes (db/db) mutant C57BL/KsJ mice: relationship to estradiol localization, systemic glycemia, and persistent hyperinsulinemia. AB - The diabetes (db/db) genotype mutation induces a hyperglycemic-hyperinsulinemic endometabolic state in C57BL/KsJ mice, manifesting a type 2 NIDDM diabetes obesity syndrome (DOS) in this hyperphagic, leptin receptor (lf) defective model. The severity of the DOS induced by the single gene, homozygous-recessive mutation may be therapeutically moderated by gonadal steroids and pre-steroidal metabolites. The current studies define the estradiol (E2)-modulated phenotypic, systemic, cytochemical, and cellular metabolic responses to db/db mutation expression as compared to littermate control (+/?) indices. The db/db mutation induced dramatic age- and DOS severity-related increases in body weights, blood glucose, and serum insulin concentrations relative to +/? indices between 4-week old (i.e., initial onset stage of DOS phenotype) and 16-week-old (i.e., chronic stage of DOS) groups. Chronic, low-dose (0.1 microg/3.5 days) E2 treatment (E2 HRx) significantly reduced the obesity mass and blood glucose levels of db/db mutants relative to oil-HRx groups. Similarly, E2-HRx maintained pancreatic glucose utilization rates and pancreatic tissue weights in db/db mutants to near +/? indices. Concurrent amelioration of db/db-enhanced pancreatic lipogenesis and islet hypercytolipidemia occurred following E2-HRx. Pancreatic islet lipo deposition was markedly reduced in db/db mutants following E2-HRx, and the restoration of islet size and cellular insulin concentrations correlated with beta-cell cytoplasmic regranulation of insulin secretory vesicles. In chronic E2 HRx db/db groups, autoradiographic localization of (3)H-E2 was demonstrated in the nuclear compartments of regranulated, nonhypertrophic islet cell populations, including insulin-containing beta-cells. In chronic E2-HRx db/db mutants, beta cell insulin granulation was prominent in mildly hypertrophic pancreatic islets, with cytodistribution patterns and concentrations comparable to normal +/? indices. In contrast, E2-HRx maintained the systemic hyperinsulinemia characteristic of oil-HRx db/db mutants. The results of these studies indicate that the severity of the type 2 NIDDM endometabolic syndrome induced by the db/db genotypic mutation may be influenced by E2-HRx, including reduction of the islet hypercytolipidemia and hypertrophic atrophy which are indicators of impending pancreatic involution in this mutant model. The hypercytolipidemia-induced demise of beta-cell cytoarchitecture was reduced by E2-HRx, including the reestablishment of islet beta-cell cytogranulation. These data suggest that the severity of genomic db/db-mutation expression may be modified by E2-HRx, with the gonadal steroid probably acting as a nuclear-specific stimulatory transcriptional modulator of cellular glucometabolic cascades in the absence of leptin-directed homeostatic influences. PMID- 15654654 TI - Isolation and multilineage differentiation of bovine bone marrow mesenchymal stem cells. AB - The bone marrow harbors a population of mesenchymal stem cells (MSCs) that possess the potential to differentiate into bone, cartilage, and fat, and along other tissue pathways. To date, MSCs from various species have been studied. Despite the bovine experimental model being widely used in experiments in vivo and in vitro, only a limited amount of information regarding bovine MSCs is available. The aim of this study was to isolate and induce the multilineage mesenchymal differentiation of bovine MSCs, thereby initiating further research on these cells. Bovine MSCs were isolated from eight calves, and osteogenic, chondrogenic, and adipogenic differentiation was induced by using a combination of previously reported protocols for other species. The level of differentiation was evaluated by histological examination and by analyzing the expression of tissue-specific genes by a quantitative "real time" reverse transcription/polymerase chain reaction technique. Following osteoinduction, the isolated fibroblast-like cells transformed into cuboidal cells and formed alkaline-phosphatase-positive colonies; during differentiation, these colonies transformed into mineralized nodules. In addition, osteogenesis was followed by osteocalcin and collagen type I mRNA expression. Chondrogenesis was confirmed by the demonstration of collagen type II, aggrecan, and sox9 mRNA expression in the cells stimulated by transforming growth factor beta1 in monolayer culture. After being cultured in an adipogenesis-inducing medium, the MSCs responded by the accumulation of lipid vacuoles and the expression of adipocyte-specific genes. We have therefore demonstrated that cells harvested from bovine bone marrow are capable of in vitro extensive multiplication and multilineage differentiation, making them a relevant and invaluable model in the field of stem cell research. PMID- 15654655 TI - Prostaglandins differently regulate FGF-2 and FGF receptor expression and induce nuclear translocation in osteoblasts via MAPK kinase. AB - We have previously reported that prostaglandin F(2alpha) (PGF(2alpha)) and its selective agonist fluprostenol increase basic fibroblast growth factor (FGF-2) mRNA and protein production in osteoblastic Py1a cells. The present report extends our previous studies by showing that Py1a cells express FGF receptor-2 (FGFR2) and that treatment with PGF(2alpha) or fluprostenol decreases FGFR2 mRNA. We have used confocal and electron microscopy to show that, under PGF(2alpha) stimulation, FGF-2 and FGFR2 proteins accumulate near the nuclear envelope and colocalize in the nucleus of Py1a cells. Pre-treatment with cycloheximide blocks nuclear labelling for FGF-2 in response to PGF(2alpha). Treatment with SU5402 does not block prostaglandin-mediated nuclear internalization of FGF-2 or FGFR2. Various effectors have been used to investigate the signal transduction pathway. In particular, pre-treatment with phorbol 12-myristate 13-acetate (PMA) prevents the nuclear accumulation of FGF-2 and FGFR2 in response to PGF(2alpha). Similar results are obtained by pre-treatment with the protein kinase C (PKC) inhibitor H 7. In addition, cells treated with PGF(2alpha) exhibit increased nuclear labelling for the mitogen-activated protein kinase (MAPK), p44/ERK2. Pre treatment with PMA blocks prostaglandin-induced ERK2 nuclear labelling, as confirmed by Western blot analysis. We conclude that PGF(2alpha) stimulates nuclear translocation of FGF-2 and FGFR2 by a PKC-dependent pathway; we also suggest an involvement of MAPK/ERK2 in this process. PMID- 15654657 TI - Factors predicting long-term graft survival after kidney transplantation: multicenter study in Japan. AB - A multicenter retrospective study was conducted in 936 living donor kidney transplant recipients treated with cyclosporine (CsA) or tacrolimus (FK) from April 1982. The influences of acute rejection, hyperlipidemia, and hypertension were estimated by Kaplan-Meier's analysis and Wilcoxon's analysis. Of 916 recipients, 532 (58.1%) had acute rejections. The 5- and 10-year graft survival rates in the recipients with acute rejection were 75.2% and 55.2%, respectively. The corresponding rates of the recipients without acute rejection were 80.2% and 70.6%, respectively. The graft survival rate was worse in recipients with late phase rejection and multiple rejection episodes (p < 0.00006). Of 451 recipients, 176 (39.0%) had hypercholesterolemia 3 years after kidney transplantation. The 5- and 10-year graft survival rates in the recipients with hypercholesterolemia were 88.7% and 68.7%, respectively. Those of the recipients without hypercholesterolemia were 95.2% and 83.9%, respectively. The graft survival rate in the recipients with hypercholesterolemia was lower than that in the recipients without hypercholesterolemia (p = 0.003). Of 323 recipients, 123 (38.1%) had hypertriglyceridemia 3 years after kidney transplantation. The 5- and 10-year graft survival rates in the recipients with hypertriglyceridemia were 93.7% and 80.5%, respectively. Those in the recipients without hypertriglyceridemia were 95.1% and 86.5%, respectively. The graft survival rate in the recipients with hypertriglyceridemia was lower than that in the recipients without hypertriglyceridemia (p = 0.371). Of 367 recipients, 151 (41.1%) had systolic hypertension 3 years after kidney transplantation. The 5- and 10-year graft survival rates in the recipients with hypertension were 85.6% and 64.7%, respectively. Those of the recipients without hypertension were 95.6% and 83.8%, respectively. The graft survival rate in the recipients with hypertension was lower than that in the recipients without hypertension (p < 0.001). Acute rejection, hyperlipidemia (hypercholesterolemia and hypertriglyceridemia), and hypertension are predictive factors for long-term graft survival. Especially the onset time, number of rejections, and efficacy of treatment for acute rejection would have a significant influence on long-term graft survival. PMID- 15654658 TI - Confidentiality governing surgical research practice. AB - Healthy subjects or patients volunteering to participate in trials expect that their privacy and autonomy will be protected. The aim of this article is to highlight issues related to confidentiality governing surgical research practice. A search of the current relevant literature was undertaken. Consent to the disclosure of any information should be sought wherever practicable, but disclosures should be kept to the minimum necessary. The data should be made anonymous where unidentifiable data serve the purpose. Where the previously described actions are not practicable for various reasons, data may be disclosed for research, provided participants have been given information about access to their records and about their right to object. Personal information may only be disclosed without individual's consent when it is for the protection of the public interest, but this has proved too ambiguous a rubric to be useful without proper clarification. Hampering of noncommercial medical research should also be avoided, as it may cause serious damage to public health. Confidentiality in research is an important issue in the protection of the participants' rights to privacy and autonomy, and it should be considered in the design of each study. Breach of confidentiality is legally justifiable for the sake of the public interest, but proper clarification of the law is required in order to avoid hampering noncommercial medical research that is vital for the public health. PMID- 15654660 TI - Occult intrabiliary rupture of hydatid cysts in the liver. AB - The aim of this study was to investigate the effects of various surgical modalities directed at the cavity of hydatid cysts in patients with occult intrabiliary rupture. In this respect, 324 patients with hydatid cyst of the liver operated on during 1983-2003 were analyzed; among them, 39 patients with occult intrabiliary rupture were included in the study. Clinical symptomatology, physical examination, laboratory findings, results of imaging studies, the localization and size of the cyst, and operative findings were reviewed. Twelve patients had complications, and there was no mortality. The most common complication was bile fistula. The average postoperative hospital stay was 7.0 +/ 3.8 days for patients with omentoplasty and 6.0 +/- 2.5 days for those who underwent cavitary drainage. For patients who do not have bile-stained cystic fluid, the utilization of scolicidal agents is appropriate. Although the opening of the duct is sutured when it is identified, the risk of biliary fistula is not clearly correlated with this approach. In such cases, omentoplasty provides a good alternative to cavitary drainage. PMID- 15654659 TI - A critical evaluation of hepatic resection in cirrhosis: optimizing patient selection and outcomes. AB - Hepatic resection has long been the mainstay of treatment of primary liver cancers, particularly hepatocellular carcinoma (HCC). Because of the high incidence of cirrhosis in patients with HCC, the use of resection was initially limited by the ability of the cirrhotic liver to sustain the surgical insult and the mass reduction. Today, hepatectomy in cirrhosis is undergoing a remarkable evolution. Although surgical and anesthetic improvements have increased the safety of this option, the rapid development of alternative therapies has decreased the need for it. Local excision for small HCC is likely to be replaced by image-guided, percutaneous ablative techniques. Furthermore, total replacement of a cirrhotic liver may be a more effective long-term cure than resection. Unquestionably, resection remains the optimal approach for patients with large tumors and healthy underlying liver function. The role of rapidly evolving new approaches will remain the subject of intensive inquiry in the years to come. In this report, we have attempted to clarify current practice with respect to the evaluation, selection, and technique of resection in cirrhosis, and identify areas of active inquiry. PMID- 15654661 TI - Hemorrhage after duodenopancreatectomy: impact of neoadjuvant radiochemotherapy and experience with sentinel bleeding. AB - Postoperative hemorrhage (PH) after duodenopancreatectomy (DP) is frequently lethal. The aim of this study was to delineate guidelines of management. Between August 1994 and July 2003, 172 patients underwent DP for cancer. Altogether, 26 patients were subjected to an institutional protocol (IP) with standard-dose chemoradiation (CRT) and 4 patients to an extrainstitutional protocol (EIP) with high-dose CRT. Sixteen patients (9.3%) were reoperated for PH. Hemorrhage occurred in 23% of irradiated patients (4 EIP, 3 IP) and in 6% of nonirradiated patients [confidence interval (CI) 1.8-6.5]. Pancreatic leak occurred in nine patients with PH (56%). Sentinel bleeding (SB) was noted in eight patients (50%) with a mean delay of 10 days after DP. Overall mortality after hemorrhage was 56%. Morality rates of patients with EIP or IP were, respectively, 100% and 0%. Mortality rates of patients with or without SB were similar. Mortality rates of axial bleeding (hepatic artery, mesenteric vessels) or lateral bleeding (pancreas remnant, splenic vessels) were, respectively, 88% and 25% (CI 1.6-8.6). Completion of pancreatectomy was achieved in 75% without rebleeding. Preoperative high-dose CRT increased the risk of fatal PH. Because SB occurs before massive hemorrhage, prompt reoperation could reduce mortality. Completion of pancreatectomy was essential during reintervention. Axial bleeding supports high mortality. Moving to the left, the pancreatojejunostomy could avoid contact of pancreatic juice with axial vessels in the case of pancreatic leakage. Ligating the gastroduodenal artery during DP had to leave a stump of around 1 cm to facilitate hemorrhage control without ligating the common hepatic artery. PMID- 15654662 TI - Impact of age on quality of life in patients with rectal cancer. AB - Some studies indicate that age at the time of surgery has a general effect on outcomes. The impact of age on the quality of life (QOL) of patients with rectal cancer, however, has not been investigated. The present study was conducted to address this issue. Over a 5-year period the European Organization for Research and Treatment of Cancer (EORTC)-QLQ-C-30 and a tumor-specific module were prospectively administered to patients before surgery, at discharge, and at 3, 6, 12, and 24 months postoperatively. Comparisons were made between age groups. A total of 519 patients participated in the study. QOL data were available for 253 patients. Significant differences were observed only between patients aged 69 years and younger (< or =69 years) (169/253) and those aged 70 years and older (> or =70 years) (85/253). Physical and role functioning was better for patients < or =69 years; patients > or =70 years suffered from increased pain and fatigue. Younger patients had more difficulty with sexual enjoyment, and over time sexual strain was worse for patients aged > or =70 years during the early postoperative period but improved, whereas patients aged < or =69 years had increasing levels of strain over time. The findings in this study confirmed that QOL is dynamic over time and that age has an impact on QOL and sexuality. Patients aged > or =70 years are affected by impaired physical functioning, global health, and fatigue, whereas increased treatment strain during the early postoperative period improves over time. Patients aged < or =69 years experience increased strain because of impaired sexual function. PMID- 15654663 TI - Addition of rectus sheath relaxation incisions to emergency midline laparotomy for peritonitis to prevent fascial dehiscence. AB - The incidence of fascial dehiscence and incisional hernia after two methods for abdominal wound closure (rectus sheath relaxation incisions and conventional mass closure) was studied in a randomized prospective clinical trial in a consecutive series of 100 patients undergoing midline laparotomy for peritonitis. The two groups were well matched for etiologies of peritonitis, the surgical procedures performed, and the presence of known risk factors for fascial dehiscence. Fifty patients each were randomized either to the conventional continuous mass closure procedure or the rectus sheath relaxation incision technique (designed to increase wound elasticity and decrease tension in the suture line) using identical polypropylene sutures. The incidence of postoperative complications such as duration of ileus, chest infection, and wound infection were not statistically different between the two groups. The intensity of postoperative pain in the rectus sheath relaxation incision group was significantly less. The incidence of wound hematoma was significantly increased in the rectus sheath relaxation incision group. The incidences of fascial dehiscence (16% vs,28%; p < 0.05) and incisional hernia (18% vs, 30%; p < 0.05) were significantly lower after rectus sheath relaxation incisions compared to conventional mass closure. Closure of the midline laparotomy wound in cases of peritonitis using the rectus sheath relaxation technique is safe and less painful, provides increased wound elasticity and decreased tension on the suture line, and significantly decreases the incidence of wound dehiscence. PMID- 15654664 TI - Predictive value of procalcitonin for the diagnosis of bowel strangulation. AB - Intestinal obstruction is responsible for 3% of admissions to hospital emergency surgical departments, but it is difficult to distinguish simple obstruction from strangulation. Simple criteria for this distinction are sought. In this experimental study, procalcitonin, a known marker of bacterial inflammation, was used to detect strangulation. The predictive value of procalcitonin for small bowel strangulation was evaluated. Thirty male New Zealand rabbits (mean weight: 3.0 kg) were divided into three groups. In the first (control) group, only laparatomy was performed. In the second group, simple obstruction was created by ligating a 10-cm distal ileum segment. In the third group, distal strangulation was created by ligating a 10-cm distal ileum segment with the mesentery. Blood (1 cc) was taken from the right auricular vein of each animal for measuring the procalcitonin level. In both the control group and the simple obstruction group the procalcitonin levels were normal. In the strangulation group, elevation of procalcitonin was detected after 30 minutes, and the elevation was statistically significant at 120th minute compared with the control and simple obstruction groups. In patients with small bowel obstruction, measurement of procalcitonin levels is easy to perform and can be used in the follow-up. A more extensive clinical study is needed to evaluate the accuracy of the test as a marker. PMID- 15654665 TI - Gallstone disease after extended (D2) lymph node dissection for gastric cancer. AB - Few studies have reported the incidence and clinical outcomes of gallstone disease after extended (D2) lymph node dissection for gastric cancer. The present study was designed to retrospectively compare limited (D1) and D2 dissections in terms of gallstone formation, presentation of gallstones, and surgery for gallstone disease. A total of 805 Japanese gastric cancer patients (595 male, 210 female) who underwent curative resection with D1 (n = 490) or D2 (n = 315) dissection were retrospectively reviewed. Of those subjects followed for 70.5 +/- 44.3 months (range: 2-196 months), 102 (12.7%) developed gallstones. The incidence of gallstone formation was higher in the D2 group than in the D1 group (17.8% vs. 9.4%, p = 0.001). The interval between gastrectomy and detection of gallstones was shorter in the D2 group than in the Dl group (18.8 +/- 11.4 months vs. 29.4 +/- 18.3 months, p = 0.002). Of those with gallstones followed for 48.0 +/- 28.6 months (range: 1-158 months), 74 (72.5%) remained asymptomatic, and 15 (14.7%) experienced mild biliary pain. Thirteen patients (12.7%) developed recurrent biliary pain (n = 3) or biliary complications (n = 10; 6 acute cholecystitis, 3 obstructive jaundice, and 1 cholangitis), and required surgical treatment. Surgery was more frequently sought in the D2 group than in the D1 group (19.5% vs. 4.3%, p = 0.033). In conclusion, patients with D2 dissection developed gallstones more frequently and earlier than patients with D1 dissection. Of those with gallstones, patients with D2 dissection required surgery more often than patients with D1 dissection. A closer follow-up should be mandatory for gallstone disease after D2 dissection, but further studies are needed before generalizations can be made. PMID- 15654667 TI - [Botulinum toxin type A in the management of spasticity in adults: recommendations of an interdisciplinary group of experts]. PMID- 15654668 TI - [Level of knowledge about epilepsy among Polish patients with epilepsy and their families in the European study SPOKE (Sanofi-Synthelabo Programme for Outcome Knowledge of Epilepsy)]. AB - BACKGROUND AND PURPOSE: The main aim of the European Study SPOKE (Sanofi Synthelabo Programme for Outcome Knowledge of Epilepsy) designed by the Department of Neurosciences, Walton Hospital in Liverpool and the Centre for Health Services Research at the University of Newcastle upon Tyne, England, was to describe levels of knowledge about epilepsy among people with epilepsy and their families. It was equally important to identify gaps in understanding the most important aspects of the illness as well as to identify characteristics of people who score poorly on the Epilepsy Knowledge Questionnaire (EKQ). Another aim was to consider the implications of identified gaps in knowledge for future educational and psychosocial interventions. MATERIAL AND METHODS: The study was conducted in 10 European countries and included 6156 people with epilepsy and 6506 members of their families. Results presented here relate to data for Poland only, where the examined population consisted of 1028 patients with epilepsy. 1033 questionnaires were returned by carers. RESULTS: Over 40% of all respondents had frequent seizures, and one third were seizure free. Scores on the Epilepsy Knowledge Questionnaire were high for the majority of respondents, but generally lower than in most of the countries. With reference to some aspects of illness important gaps in knowledge have been demonstrated. For example, it was shown that a significant number of respondents provided incorrect answers to questions relating to aspects of the etiology of epilepsy and administration of antiepileptic drug medication. There were a number of significant differences between the profiles of high and low scorers on the EKQ; high scorers were likely to have spent longer on education, have lower scores on the impact of epilepsy scale and report better adjustment to their epilepsy. CONCLUSIONS: The results of the study once again underline the importance of continued information among patients with epilepsy and their families in order to achieve. PMID- 15654669 TI - [Analysis of PCNA, Ki67, AgNOR and p53 expression in brain glial tumors]. AB - BACKGROUND AND PURPOSE: The aim of this study was to evaluate clinical usefulness of proliferating cell nuclear antigen (PCNA), Ki67 antigen, p53 protein and silver-binding nucleolar organizer regions (AgNOR) in brain glial tumors. MATERIAL AND METHODS: The investigation of PCNA, Ki67 and p53 was carried out on a group of 120 patients with glial tumors operated on at the Neurosurgical Department of Wroclaw Medical University including 63 patients operated again because of recurrence. AgNOR was evaluated on a group of 64 patients including 38 patients operated again. Classical histological tests, immunohistochemical tests for PCNA, p53 and Ki67 activity with monoclonal antibodies (DACO) and histochemical tests for AgNOR were performed on every specimen of tumor tissue. The level of 40% for PCNA, 2.75 (equal to median) for AgNOR and 5% for Ki67 and p53 was adopted as significant. RESULTS: Mean expression of PCNA of glial tumors grade I and II was 32%, grade III and IV - 44% (p<0.05). Mean expression of AgNOR was 1.88 and 3.16 (p=0.00001), respectively. Average PCNA expression in recurrent tumors to 12 months was 52.7% and for later recurrences - 35.4% (p<0.05). Average expressions of AgNOR were 3.38 and 2.68 (p<0.05), respectively. Differences of Ki67 and p53 expressions were not significant. CONCLUSIONS: PCNA and AgNOR expressions correlate with proliferative activity, growth rate and histological malignancy, reaching high values in highly malignant and early recurrent tumors. Antigens Ki67 and p53 do not seem to be predictive markers of glial tumors. PMID- 15654670 TI - [F-wave amplitude in peripheral nervous system lesions]. AB - BACKGROUND AND PURPOSE: F-wave is a late response recorded from muscle elicited by electric impulse conveyed antidromically to alpha-motor neurons of the spinal cord. F-wave latency and frequency are assessed in routine electroneurography providing information of conduction in the proximal segment of the nerves. F-wave amplitude is rarely analyzed, while it could add valuable information on excitability of motor neurons in different disease states. This study was conducted to determine whether F-wave amplitude is indicative of the level of the peripheral nervous system lesion. MATERIAL AND METHODS: EMG recordings of 204 consecutive patients suspected of peripheral nerves system lesion were retrospectively analyzed. Based on the clinical diagnosis three groups were defined: neuropathy (N=100), myopathy (N=33), ALS (N=18), reference (musculoskeletal pain syndromes: N=53). F-wave amplitude and F/CMAP-ratio (CMAP compound motor action potential) and their relation to parameters of impulse conduction in motor nerve fibers was analyzed. RESULTS: Mean F/CMAP ratio was 11.1% in ALS, 5% in myopathy (p=0.008), 7.1% in neuropathies and 5.6% in the reference group. Giant F-wave (more than 10% of CMAP) was observed in 30% of nerves in ALS, 15% in neuropathy and 10% in myopathy (p=0.036). F-wave amplitude correlated significantly with CMAP amplitude in all groups, while F/CMAP ratio was inversely related to CMAP amplitude in ALS (r=-0.43, p<0.01) and neuropathy (r=-0.37, p<0.0001). F-wave frequency was similar in all groups and correlated with CMAP amplitude. CONCLUSIONS: F-wave amplitude is not indicative of the level of peripheral nervous system lesions. Giant F-wave is observed in neurogenic processes. It reflects an increase of motor unit size in the reinnervation process, but possibly also a change of excitability of motor neuron and its axon. PMID- 15654672 TI - [Results of the operative treatment of intramedullary gliomas]. AB - BACKGROUND AND PURPOSE: To assess the results of the operative treatment of intramedullary gliomas. MATERIAL AND METHODS: In 1995-2002, 30 patients with intramedullary tumors were treated surgically in the Department of Neurosurgery in Sosnowiec. Among them 23 intramedullary gliomas were recognized. There were 8 females and 15 males aged from 11 to 61. The condition of the patients before the operation was assessed according to a modified McCormick's scale. Only two patients were found to be in a good condition -- I degree, 17 patients (74%) -- II degree or III degree and 4 patients were in a serious condition -- IV degree. Among the operated tumors only 9 had a clear border. There were 5 ependymomas (grade I) and 4 anaplastic ependymomas (grade III). The remaining ones were astrocytomas of varying malignancy and infiltration of the white matter of the spinal cord: 8 cases of fibrillary astrocytoma, 3 cases of gemistocytic astrocytoma, but one anaplastic and one case of pilocytic astrocytoma, oligoastrocytoma mixtum and anaplastic oligoastrocytoma. RESULTS: Control MRI revealed a total removal of the lesion in 16 patients (70%), gross total removal in 5 patients (21%), partial removal in two patients (8%). Four patients died among those operated on: two in the perioperative period, the other two 9 months and 2 years after the surgery, respectively. In four patients there was observed regrowth of the lesion and one patient was re-operated on. In the second case fatal recurrence in region C2-Th6 was observed and the patient died, in the last two cases the patients were stable and thus it was decided to delay the re operation. CONCLUSION: Low mortality, morbidity and recurrence rates recommend surgery as an effective treatment for intramedullary gliomas. PMID- 15654666 TI - State of the art in burn treatment. AB - Optimal treatment of burn victims requires deep understanding of the profound pathophysiological changes occurring locally and systemically after injury. Accurate estimation of burn size and depth, as well as early resuscitation, is essential. Good burn care includes also cleansing, debridement, and prevention of sepsis. Wound healing, is of major importance to the survival and clinical outcome of burn patients. An ideal therapy would not only promote rapid healing but would also act as an antiscarring therapy. The present article is a literature review of the most up-to-date modalities applied to burn treatment without overlooking the numerous controversies that still persist. PMID- 15654671 TI - [The effect of rivastigmine on cognitive functions and regional cerebral blood flow in Alzheimer's disease and vascular dementia: follow-up for 2 years]. AB - BACKGROUND AND PURPOSE: The aim of the work was to investigate the effect of treatment with rivastigmine, one of the inhibitors of acetylcholinesterase (AChE I) on the regional cerebral perfusion (rCBF) and the cognitive functions of the brain in patients with Alzheimer's Disease (AD) and Vascular Dementia (VaD). MATERIAL AND METHODS: The investigations of rCBF were carried out using SPECT (Single Photon Emission Computed Tomography). The results given concern investigations of patients carried out at the onset of the investigation, after 12 months, and 24 months of rivastigmine treatment. RESULTS: In patients with AD it was found that treatment with rivastigmine increases rCBF by 5-7% in the temporal areas during the first 12 months. In the frontal areas the increase was by 3-5%. During the next 12 months rCBF with an accuracy of 2% returned to the initial level, with the exception of the motor cortex, where it remained on the level increased by 5-6%. However, the cognitive functions remained constant during the first 12 months of treatment and decreased significantly during the next 12 months. In patients with VaD rCBF increased in all the regions of the brain except for the temporal posterior regions, and remained at an elevated level for the next 12 months. The cognitive functions deteriorated slowly, but to a much lesser degree than in the case of AD. CONCLUSIONS: From the investigations carried out it follows that treatment with rivastigmine during 24 months prevents a decrease of rCBF in patients with AD. However, the cognitive functions deteriorate after 24 months. PMID- 15654673 TI - [Endoscopic treatment of the spontaneous intracerebral hematomas]. AB - BACKGROUND AND PURPOSE: Surgical evacuation of spontaneous intracerebral hematomas (ICH) performed in a traditional way usually increases primary brain tissue damage due to the hemorrhage. On the other hand, symptoms of the intracerebral pressure and secondary brain tissue destruction close to the hematoma are the basis for making a decision about surgical treatment. In order to limit surgical trauma we started research to evaluate the usefulness of endoscopic surgery in the treatment of ICH. MATERIAL AND METHODS: Twenty three cases were included in the study. Patients with consciousness disorders and/or focal neurological deficit and different systemic diseases were qualified for endoscopic evacuation. The diagnosis of hematoma was based on computed tomography (CT). ICHs were lobar and in certain cases they extended to the basal ganglia. All patients were operated on within one week from the onset of symptoms. Neuroendoscope was introduced to the hematoma cavity through the burr hole and the puncture of the cerebral surface over the hematoma. The hematoma was evacuated by fractionated rinsing. Bigger clots were fragmentized but those which were adjacent to the cavity wall were left. Postoperative assessment of the hematoma evacuation was based on CT performed immediately and in the second week after surgery. RESULTS: Total evacuation of the ICH was achieved in 6 patients, and its volume was reduced in 17 cases. Symptoms of brain edema resolved in all patients. A significant trend to reduce focal neurological deficits was observed: 16 patients improved and 3 remained unchanged. Four patients died. CONCLUSIONS: Endoscopic surgery allows a complete hematoma evacuation or reduction of its volume, reduces symptoms of brain edema and accelerates the improvement of focal neurological deficits. PMID- 15654675 TI - [Safety of medicinal products and reporting of adverse drug reactions]. AB - The authors present the role of reports of adverse drug reactions (ADR) in pharmacotherapy. Worldwide monitoring of ADRs together with the European and Polish pharmacovigilance systems are discussed. Some practical information concerning the report form is described i.e. types of reactions, definition of a serious adverse drug reaction, minimum information for the report validation, handling of reports. PMID- 15654674 TI - [Genetic counseling and testing for families with Alzheimer's disease]. AB - With the identification of the genes responsible for autosomal dominant early onset familial Alzheimer's disease (FAD genes), there is a considerable interest in the application of this genetic information in medical practice through genetic testing and counseling. Pathogenic mutations in the PSEN1 and PSEN2 genes encoding presenilin-1 and -2, and the APP gene encoding amyloid b precursor protein, account for 18-50% of familial EOAD cases with autosomal dominant pattern of inheritance. A clinical algorithm of genetic testing and counseling proposed for families with AD has been presented here. A screening for mutations in the APP, PSEN1, and PSEN2 genes is available to individuals with AD symptoms and at-risk children or siblings of patients with early-onset disease determined by a known mutation. In an early-onset family, a known mutation in an affected patient puts the siblings and children at a 50% risk of inheriting the same mutation. The goal of genetic testing is to identify at-risk individuals in order to facilitate early and effective treatments in the symptomatic person based on an individual's genotype and strategies to delay the onset of disease in the presymptomatic mutation carriers. However, there are several arguments against the use of genetic testing both presymptomatically (unpredictable psychological consequences of information about a genetic defect for family members) and as a diagnostic tool for the differential diagnosis of dementia in general practice (a risk of errors in an interpretation of mutation penetrance and its secondary effects on family members, especially for novel mutations; the possibility of coexistence of another form of dementia at the presence of a mutation). Currently, APOE genotyping for presymptomatic individuals with a family history of late-onset disease is not recommended. The APOE4 allele may only confer greater risk for disease, but its presence is not conclusive for the development of AD. PMID- 15654676 TI - [Minimally invasive posterior corpectomy of the lumbar spine with transpedicular fixation]. AB - The authors present their experience in the minimally invasive posterior keyhole lumbar corpectomy with transpedicular stabilization. This technique involves the removal of the posterior part of the affected vertebral body with the pedicle screw fixation through four 2-3 cm long skin incisions on the back. Two cephalad skin incisions provide an approach for corpectomy and instrumentation of the upper pedicles of the construct. Two caudal skin incisions provide an approach for instrumentation of the lower pedicles of the construct. The minimum armamentarium requirement includes classic micro lumbar discectomy retractor set and intraoperative fluoroscopy. According to the authors' best knowledge this is the first minimally invasive posterior keyhole lumbar corpectomy ever reported in the literature (2002). This is also the first minimally invasive transpedicular fixation ever performed in Poland (2002). This technique was presented during EANS Congress (Lisbon, September 2003). Some reports have recently appeared in the literature on percutaneous pedicle screw fixation of the lumbar spine in non traumatic cases. A special instrumentarium system (Sextant by Medtronic) has been developed and used in this type of minimally invasive stabilization. Although this system has not been dedicated for spine fractures it is feasible in trauma cases. We have one case of L2 burst fracture fixed percutaneously with Sextant. PMID- 15654678 TI - [The case of a patient with treatment-refractory obsessive-compulsive disorder operated by stereotactic bilateral cingulotomy]. AB - This case report presents the first patient in Poland with treatment-refractory obsessive-compulsive disorder, operated by stereotactic bilateral cingulotomy. The patient, aged 38 years, with 20-year duration of illness, despite intensive pharmacological and psychotherapeutic treatment was totally unable to function due to obsessive-compulsive symptoms of extreme intensity. The paper presents the neuroimaging documentation of psychosurgery performed on 3rd December 2002. On the basis of nearly two-year follow-up of the patient, the result of the psychosurgery can be assessed as moderately favorable. The patient's subjective mood and social functioning have improved, as well as his cognitive functions measured by means of neuropsychological tests. In patients with severe, chronic and treatment-refractory obsessive-compulsive disorder psychosurgical treatment may be a therapeutic option. PMID- 15654680 TI - [The use of activated recombinant factor VII in the treatment of nasopharyngeal hemorrhage in a patient with extensive fronto-basal injuries. Case report]. AB - The main indication for recombinant activated factor VII (rFVIIa) is the treatment of patients with hemophilia or other hemostatic disturbances. Additionally, rFVIIa has been considered as a universal hemostatic agent prompting its use in the management of severe uncontrolled bleeding in patients without pre-existing coagulopathies. Authors report their own experience in the beneficial effect of rFVIIa (NovoSeven, NovoNordisk, Denmark) administration in a patient with severe bleeding from the nasopharyngeal region after extensive fronto-basal injuries. After one dose of NovoSeven (2.4 mg - 40 microg/kg of body weight) reduction in bleeding, normalization of hemostasis parameters and hemodynamic stabilization of the patient were achieved. PMID- 15654681 TI - [Surgical evacuation of an embolization coil from the middle cerebral artery]. AB - We report the case of a 54-year-old woman with a right intracavernous internal carotid artery aneurysm treated endovascularly with MDS coils. During the procedure, one of the coils migrated to the middle cerebral artery (MCA), resulting in left hemiplegia. The attempt to remove the migrated coil endovascularly failed and the patient had emergency front-temporal craniotomy. After the dissection of the Sylvian fissure, the M1 portion of the MCA was exposed with the coil visible through the distended wall of the artery. Because it was feared that the M1 arteriotomy could tear the MCA, the coil was removed through the anterior temporal artery, a small branch of the M2 segment. Hemiplegia gradually resolved and the patient recovered completely. This case depicts an alternative route via a distal arteriotomy to remove the migrating coil. Additionally, it implies that endovascular procedures in neurosurgical patients should be performed by the interventional neurosurgeon (neurosurgeon trained to perform endovascular procedures) or at least with the support of a neurosurgical department for immediate intervention in the case of life threatening complications. PMID- 15654682 TI - [Alexander Domaszewicz (1887-1948). Professor of neurosurgery at the Warsaw University]. PMID- 15654684 TI - 1J(C,H) couplings to the individual protons in a methyl group: evidence of the methyl protons' engagement in hydrogen bonds. PMID- 15654683 TI - Total synthesis of (+)-leucascandrolide A. PMID- 15654685 TI - Biodegradable, amorphous copolyester-urethane networks having shape-memory properties. PMID- 15654686 TI - Direct access to bicontinuous skeletal inorganic plumber's nightmare networks from block copolymers. PMID- 15654687 TI - Comparison of dexamethasone pharmacokinetics in female rats after intravenous and intramuscular administration. AB - This study seeks a route of drug administration that would produce a pharmacokinetic profile for dexamethasone not significantly different from the intravenous route in female rats and would offer reproducible drug input with minimal stress to the animals. The intramuscular (i.m.) route of drug administration vs intravenous (i.v.) injection were compared in three female Wistar rats administered 1 mg/kg dexamethasone phosphate. Dexamethasone plasma concentrations were measured by a normal phase HPLC assay for 12 h after drug administration. Dexamethasone exhibited monoexponential behavior after intravenous dosing and was absorbed rapidly after intramuscular dosing (absorption half-life of 14 min) with 86% bioavailability. Dexamethasone had a terminal half-life of 2.3 h after drug administration by either route. The volume of distribution of 0.78 l/kg and the clearance of 0.23 l/h/kg are in good agreement with reported pharmacokinetic parameters in male rats. Intravenous dosing can be replaced by intramuscular dosing without causing any marked difference in dexamethasone pharmacokinetics. PMID- 15654689 TI - Guidelines for ethical behavior relating to clinical practice issues in electrodiagnostic medicine. PMID- 15654688 TI - Olfactory neuroblastoma: the 22-year experience at one comprehensive cancer center. AB - BACKGROUND: Olfactory neuroblastoma (ONB) is a rare tumor arising from the olfactory neuroepithelium. There is no universally accepted staging system, and treatment approaches lack uniformity. We present one institution's experience with this tumor and the results of therapy. METHODS: Thirty patients treated for ONB at The University of Texas M. D. Anderson Cancer Center between 1979 and 2002 were retrospectively reviewed. The diagnosis of ONB was histologically confirmed for each patient. RESULTS: The mean follow-up was 7.32 years. In 77% of cases, patients received treatment with surgery followed by postoperative radiation therapy. Sixteen percent received chemotherapy as part of their initial treatment. Overall 5-year and 10-year survival rates were 89% and 81%, respectively. Nine patients whose disease was initially stage C had a recurrence. The mean time for recurrence was 4.67 years. CONCLUSIONS: The M. D. Anderson Cancer Center approach to ONB is complete surgical resection, usually involving a craniofacial approach, with postoperative radiation therapy. This approach seems to be curative in early-stage disease. Late recurrence warrants long-term follow up. PMID- 15654690 TI - Plasticity of dynamic muscle performance with strength training in elderly humans. AB - Data are scarce relating to the plasticity with strength training of dynamic muscle performance in older humans. Hence, we investigated alterations in the torque-velocity relation with strength training in old age, and their origin. Knee extension and leg-press exercises were performed three times per week for 14 weeks. Maximal isokinetic knee extension torque was assessed during concentric and eccentric muscle actions. Agonist-antagonist muscle activation was assessed using electromyography. Vastus lateralis muscle architecture was examined in vivo using ultrasonography. Training increased concentric torque by 22-37% (P < 0.01), but failed to alter eccentric torque (P > 0.05). Increased agonist muscle activation, increased muscle fascicle lengths, and greater elastic energy recovered from tendinous structures may explain the adaptations during concentric actions, whereas the failure of eccentric torque to increase might be explained by the preservation of eccentric force with aging and an underloading of the eccentric movement phase during training. These findings may have important implications for dynamic muscle performance in old age. PMID- 15654691 TI - Meralgia paresthetica after strenuous exercise. AB - We present three patients with signs and symptoms of meralgia paresthetica (MP) after long-distance walking and cycling. No other possible causes of MP, such as trauma or exogenous compression, were present. A neuropathy of the lateral femoral cutaneous nerve was confirmed in all patients with somatosensory evoked potentials. We propose that conduction block due to local ischemia during repetitive muscle stretching was the probable cause for the neuropathy. PMID- 15654693 TI - Central and peripheral motor conduction to cremasteric muscle. AB - The few electrophysiologic studies of the cremasteric muscle (CM) have mainly been restricted to the cremaster reflex with no reference to central and peripheral nerve conduction to the muscle, probably for technical reasons.Twenty six normal adult male volunteers were studied by transcranial magnetic cortical stimulation (TMS) and stimulation of thoracolumbar roots. The genitofemoral nerve (GFN) was stimulated electrically at the anterior superior iliac spine and a needle electrode was inserted into the CM for conduction studies. The motor latency to the CM from the cortical TMS ranged from 20 to 33 ms among the subjects (25.8 +/- 2.9 ms, mean +/- SD). Magnetic stimulation of the lumbar roots produced a motor response of the CM within 9.6 +/- 1.9 ms (range, 6-15). The central motor conduction time to the CM was 16.5 +/- 2.8 ms (range, 10-21). Stimulation of the GFN produced a compound muscle action potential with a mean value of 6.4 +/- 1.8 (range, 4-10) ms in 23 of the 26 cases. Thus, central motor nerve fibers to the CM motor neurons exist, and there may be a representation area for the CM in the cerebral cortex. The GFN motor conduction time to the CM may have clinical utility, such as in the evaluation of the groin pain due to surgical procedures in the lower abdomen. PMID- 15654692 TI - Modulation of biphasic rate of end-plate potential recovery in rat diaphragm. AB - Previous diaphragm studies found that during intermittent stimulation, intratrain end-plate potential (EPP) amplitude rundown is accelerated by increasing stimulation frequency, whereas intertrain EPP rundown is independent of frequency. We hypothesized that increasing stimulation frequency accelerates rundown recovery, and with a biphasic time course. Intracellular recordings were made in vitro from rat phrenic nerve-hemidiaphragm preparations. EPP amplitude recovery after a 100-ms stimulation train and 100 ms of quiescence was significantly greater following stimulation at 200 HZ than at 20-100 HZ, despite larger antecedent EPP decline. EPP amplitudes recovered with a biphasic pattern: an early component with a fast time-constant (0.03-0.06 s) and a late component with a slow time-constant (0.5-5 s). Increased antecedent stimulation frequency accelerated the early component, but stimulation duration or pulse number modulated the late component. When interpreted in the context of vesicle recycling and replenishment models involving multiple pools and pathways, these data suggest that antecedent stimulation frequency regulates predominantly the fast pathways. This may have important implications for the development of respiratory failure in diseases of the neuromuscular junction, such as myasthenia gravis, when the firing duration and frequency are altered in association with changes in breathing pattern. PMID- 15654694 TI - A novel 9 bp deletion in the filamin a gene causes an otopalatodigital-spectrum disorder with a variable, intermediate phenotype. AB - We report a four-generation pedigree with six affected females with cranial hyperostosis and various skeletal abnormalities. The phenotype is similar to frontometaphyseal dysplasia, which is part of the otopalatodigital (OPD) spectrum. We identified a novel in-frame deletion in exon 29 of the Filamin A gene (c.4904_4912del, p.R1635_V1637del) encoding rod domain repeat 14 of the protein. The disorder resulted in early lethality in male children. The phenotype of female individuals in this family is variable and rather mild, and bridges the phenotypes of various OPD-spectrum disorders. PMID- 15654696 TI - Axenfeld-Rieger malformation and distinctive facial features: Clues to a recognizable 6p25 microdeletion syndrome. AB - Deletion of distal 6p is associated with a distinctive clinical phenotype including Axenfeld-Rieger malformation, hearing loss, congenital heart disease, dental anomalies, developmental delay, and a characteristic facial appearance. We report the case of a child where recognition of the specific ocular and facial phenotype, led to identification of a 6p microdeletion arising from a de novo 6:18 translocation. Detailed analysis confirmed deletion of the FOXC1 forkhead gene cluster at 6p25. CNS anomalies included hydrocephalus and hypoplasia of the cerebellum, brainstem, and corpus callosum with mild to moderate developmental delay. Unlike previous reports, hearing was normal. PMID- 15654695 TI - Clinical evidence of decreased olfaction in Bardet-Biedl syndrome caused by a deletion in the BBS4 gene. AB - Recent discoveries have lead to the hypothesis that ciliary dysfunction is a mechanism underlying the pathogenesis of Bardet-Biedl syndrome (BBS). Here, we describe two individuals with decreased olfaction who are members of an extended family affected with BBS caused by a homozygous deletion (c.77-220del) in the BBS4 gene. These findings correlate with the evidence that several BBS proteins, including BBS4, are expressed in the olfactory epithelium (OE). Although the prevalence and the spectrum of impaired olfaction in BBS are not known, the causal relationship of the BBS4 deletion in this family and the decreased olfaction is corroborated by evidence that Bbs2 and Bbs4 knockout mice have severe olfaction deficits and that also patients with BBS caused by mutations in other BBS genes can have impaired olfaction. This finding broadens the spectrum of clinical manifestations associated with BBS, confirms the role of BBS4 in olfaction, and lends support to the hypothesis that ciliary dysfunction is an important aspect of BBS pathogenesis. PMID- 15654697 TI - Antibody response to collagen after functional implantation of different polyester vascular prostheses in pigs. AB - Besides inflammation, specific immune responses are seen also after implantation of biomaterials. The aim was to investigate the humoral response to bovine collagen type I following implantation of various polyester (Dacron) prostheses into pigs. In 24 randomized pigs, the infrarenal aorta was replaced with a segment of collagen-impregnated, woven polyester prosthesis of low, medium, or high porosity. IgG antibodies were detected by immunoassay using native and denatured collagen type I as a target for blood samples taken on day 1 (implantation), 10, 17, 24, 62, and 116. As generally observed, antibodies to native and denatured collagen are of low titer and were significantly correlated with enhanced binding to the denatured form (p < 0.001). The highest overall antibody prevalence to native and denatured collagen was obtained on day 116 with 68% and on day 62 with 59%, respectively. Prostheses with high porosity induced an early immune response on day 10; those with low and medium porosity induced the highest antibody levels later after 2 months. Collagen antibodies neither correlated with serum IgG contents nor with antibodies to the prosthesis polyester matrix. Thus, humoral immune response against implant components may provide a further parameter in describing biocompatibility but also a potential marker that may facilitate monitoring of individual perigraft reaction. PMID- 15654698 TI - Variability in fMRI: a re-examination of inter-session differences. AB - We revisit a previous study on inter-session variability (McGonigle et al. [2000]: Neuroimage 11:708-734), showing that contrary to one popular interpretation of the original article, inter-session variability is not necessarily high. We also highlight how evaluating variability based on thresholded single-session images alone can be misleading. Finally, we show that the use of different first-level preprocessing, time-series statistics, and registration analysis methodologies can give significantly different inter session analysis results. PMID- 15654699 TI - Prevention of postsurgical tissue adhesion by anti-inflammatory drug-loaded pluronic mixtures with sol-gel transition behavior. AB - Sol-gel transition temperature-controllable Pluronic F127/F68 mixtures including mildly crosslinked alginate and nonsteroidal anti-inflammatory drug (ibuprofen) were prepared to evaluate their potential as tissue adhesion barrier gels. The sol-gel transition temperatures of the Pluronic mixtures could be controlled by adjusting F127/F68 ratio and polymer concentration. The mildly crosslinked alginate with still flow property provided the residence stability of Pluronic mixture gels in the body. Ibuprofen was loaded in Pluronic mixtures to reduce inflammatory response in the body and, thus, to prevent tissue adhesion. The gelation temperatures of the Pluronic mixtures were not affected by the alginate but lowered by the addition of ibuprofen. The in vitro drug release behavior and in vivo peritoneal tissue adhesion of the Pluronic mixtures with the sol-gel transition just below body temperatures were investigated. The drug release behavior from the ibuprofen (1 wt%)-loaded Pluronic mixture gels at 37 degrees C was examined using a membrane-less dissolution model. The drug in the mixture gels was released continuously up to about 45-65% of the total loading amount during the first 7 days. For in vivo evaluation of tissue anti-adhesion potential, the Pluronic mixtures with/without drug were coated on the peritoneal wall defects of rats and their tissue adhesion extents and tissue reactions (inflammatory response, granulation tissue formation, and toxicity in organs) were compared. It was observed that ibuprofen has a positive effect for the peritoneal tissue anti-adhesion. The Pluronic F127/F68/alginate/ibuprofen mixture gel (25 wt% of F127/F68 [7/3], 1 wt% ibuprofen) was highly effective for the prevention of peritoneal tissue adhesion and showed a relatively low inflammatory response and non-toxicity, and thus can be a good candidate material as a coatable or injectable tissue adhesion barrier gel. PMID- 15654700 TI - Fabrication and evaluation of nanoporous alumina membranes for osteoblast culture. AB - An understanding of osteoblast response to surface topography is essential for successful bone tissue engineering applications. Alumina has been extensively used as a substrate for bone tissue constructs. However, current techniques do not allow precise surface topography and orientation of the material. In this research, a two-step anodization process was optimized for the fabrication of nanoporous alumina membranes with uniform pore dimension and distribution. The anodization voltage can be varied to create nanoporous alumina membranes with pore sizes ranging from 30 to 80 nm in diameter. The impact of the nanoscale pores on osteoblast response was studied by evaluating cell adhesion, morphology, and matrix production. Scanning electron microscopy and atomic force microscopy were used to characterize the nanoporous alumina membranes. Osteoblast adhesion and morphology were investigated using scanning electron microscopy images and matrix production was characterized using energy dispersive spectroscopy. This research combined the advantages of using alumina, a material with proven biocompatibility and current orthopedic clinical applications, and incorporated porous features on the nanoscale which have been reported to improve osteoblast response. PMID- 15654701 TI - Influence of the power density on the kinetics of photopolymerization and properties of dental composites. AB - Polymer shrinkage during photopolymerization of dimethacrylate monomers, used for many years to produce materials for dental restoration, can induce either the formation of tooth-restoration gaps or the production of residual stress depending on the quality of adhesion between tooth and dental composites. In this work, the effect of the power density, used to photopolymerize three commercial dental composites (Fill Magic, Supra Fill, and Z100), on the kinetics of the reaction was investigated to determine processing conditions in which the generation of residual stress would be reduced by allowing polymer chains and macromers to flow before freezing during gelation of the polymer network. The kinetics of photopolymerization of the dental composites was monitored by real time infrared (FTIR) spectroscopy. Polymer shrinkage and mechanical properties were also investigated by using, respectively, density and microhardness measurements. Results showed that the final conversion (after 200 s), volumetric shrinkage, and microhardness values were not affected by different power densities, mainly because the amount of energy used during photopolymerization was set constant by using different irradiation times. Lower power densities were able to reduce the maximum polymerization rate and delay the formation of a rigid network. Conversion before the formation of the rigid network was also enhanced by using a lower power density. Considering that too premature gelation can lead to residual stress during shrinkage, the results of this work indicated that the use of a lower power density can be effective in terms of delaying the onset of the formation of a rigid network, providing then conditions for macromolecules to flow and relieve stress during shrinkage. PMID- 15654702 TI - A critical comparison of methods for the determination of the aging sensitivity in biomedical grade yttria-stabilized zirconia. AB - Since the recent failure events of two particular series of zirconia femoral heads for total hip replacement prosthesis, a large decrease in the use of zirconia ceramics for orthopaedic implants has been observed. In spite of the biomedical success of this material during the last 10 years, this decrease in use was required for safety reasons, until the cause of the failures is known. It has been shown that these failures were related to the low temperature hydrothermal degradation (also known as aging). Thus, it is crucial to better understand the aging behavior, in order to be able to assess its importance and then control it if required. In this study, various techniques relevant to assess the hydrothermal degradation sensitivity of biomedical grade yttria-stabilized zirconia are discussed and compared. The expected outputs of conventional methods, that is, X-ray diffraction and scanning electron microscopy are examined. More recent methods like optical interferometry and atomic force microscopy are presented, with their respective benefits and drawbacks. An up-to date comparison of these different techniques is provided, and their use for ensuring the long-term reliability of a particular batch of zirconia in terms of aging degradation is demonstrated. PMID- 15654703 TI - Screening for correctable visual acuity deficits in school-age children and adolescents. AB - BACKGROUND: Although the benefits of vision screening seem intuitive, the value of such programmes in junior and senior schools has been questioned. In addition to this there exists a lack of clarity regarding the optimum age, and frequency at which to carry out screening. OBJECTIVES: The objective of this review is to evaluate the effectiveness of vision screening programmes carried out in schools in reducing the prevalence of undetected, correctable visual acuity deficits due to refractive error in school-age children. SEARCH STRATEGY: We searched the Cochrane Central Register of Controlled Trials - CENTRAL (which contains the Cochrane Eyes and Vision Trials Register) on The Cochrane Library (Issue 3 2004), MEDLINE (1966 to August 2004) and EMBASE (1980 to August 2004). No language or date restrictions were placed on these searches. To date it has not been possible to carry out any manual searches but it is hoped to include these in a future update. SELECTION CRITERIA: We planned to include randomised controlled trials including randomised cluster controlled trials. DATA COLLECTION AND ANALYSIS: Two reviewers independently assessed study abstracts identified by the electronic searches. No trials were identified that met the inclusion criteria. MAIN RESULTS: As no trials were identified, no formal analysis was performed. A narrative synthesis of other retrieved studies was undertaken in order to explain current practice. AUTHORS' CONCLUSIONS: At present there are no robust trials available that allow the benefits of school vision screening to be measured. The disadvantage of attending school with a visual acuity deficit also needs to be quantified. The impact of a screening programme will depend on the geographical, and socio-economic setting in which it is conducted. There is therefore clearly a need for well planned randomised controlled trials, in various settings, to be undertaken so that the potential benefits and harms of vision screening can be measured. PMID- 15654704 TI - Oximes for acute organophosphate pesticide poisoning. AB - BACKGROUND: Acute organophosphorus pesticide poisoning causes tens of thousands of deaths each year across the developing world. Standard treatment involves administration of intravenous atropine and oxime to counter acetylcholinesterase inhibition at the synapse. The usefulness of oximes, such as pralidoxime and obidoxime, has been challenged over the past 20 years by physicians in many parts of the world, who have failed to see benefit in their clinical practice. OBJECTIVES: To find the clinical trial evidence for oximes producing clinical benefit in acute organophosphorus pesticide-poisoned patients. SEARCH STRATEGY: We carried out a systematic search to find randomised clinical trials (RCTs) of oximes in acute organophosphorus pesticide poisoning, using MEDLINE, EMBASE and Cochrane databases. All articles with the text words 'organophosphate' or 'oxime' together with 'poisoning' or 'overdose' were examined. (Search last updated November 2003.) SELECTION CRITERIA: Articles that could possibly be randomised clinical trials were retrieved to determine if this was the case. DATA COLLECTION AND ANALYSIS: The published methodology of the possible RCTs located is not clear. One was found in abstract form only and two other published trials also had many gaps in the published methodology. We have attempted to contact the principal authors of all three trials but have been unable to obtain further information. MAIN RESULTS: Two RCTs have been published, involving 182 patients treated with pralidoxime. These trials did not find benefit. However, the studies did not take into account a number of issues important for outcome and the methodology is unclear. Therefore, a generalised statement on effectiveness cannot be supported by the published results. In particular, characteristics at baseline were not evenly balanced, the dose of oxime was much lower than recommended in guidelines, there were substantial delays to treatment, and the type of organophosphate was not taken into account. The abstract of the third trial, a small possible RCT, is uninterpretable without further data. AUTHORS' CONCLUSIONS: Current evidence is insufficient to indicate whether oximes are harmful or beneficial in the management of acute organophosphorus pesticide poisoning. A much larger RCT is required to compare the World Health Organization recommended pralidoxime regimen (>30 mg/kg bolus followed by >8 mg/kg/hr infusion) with placebo. There are many theoretical and practical reasons why oximes may not be useful to patients with overwhelming self-poisoning. Such a study will need to be designed with pre-defined sub-group analysis to allow identification of patient sub-groups that may benefit from oximes. PMID- 15654705 TI - Rofecoxib for osteoarthritis. AB - BACKGROUND: Editor's note: The anti-inflammatory drug rofecoxib (Vioxx) was withdrawn from the market at the end of September 2004 after it was shown that long-term use (greater than 18 months) could increase the risk of heart attack and stroke. Further information is available at www.vioxx.com. Osteoarthritis is a chronic disease of the joints, characterised by joint pain, stiffness and loss of physical function. Its onset is age-related and occurs usually between the ages of 50 and 60. It is the commonest cause of disability in those aged over 65, with OA of the knee and/or hip affecting over 20 per cent of the elderly population. OBJECTIVES: To establish the efficacy and safety of rofecoxib in the management of OA by systematic review of available evidence. SEARCH STRATEGY: We searched the following databases up to August 2004: MEDLINE, EMBASE, Cochrane Database of Systematic Reviews, Cochrane Controlled Trials Register, National Research Register, NHS Economic Evaluation Database, Health Technology Assessment Database. The bibliographies of retrieved papers and content experts were consulted for additional references. SELECTION CRITERIA: All eligible randomised controlled trials (RCTs) were included. No unpublished RCTs were included in this edition of the review. DATA COLLECTION AND ANALYSIS: Data were abstracted independently by two reviewers. A validated checklist was used to score the quality of the RCTs. Comparable trials were pooled using fixed effects model. MAIN RESULTS: Twenty-six RCTs were included. The comparators were placebo, diclofenac, ibuprofen, naproxen, nimesulide, nabumetone, paracetamol, celecoxib and Arthrotec. The evidence reviewed indicated that rofecoxib was more effective than placebo (patient global response RR 1.75 95% CI: 1.35, 2.26) but was associated with more adverse events (RR 1.32 95% CI 1.11, 1.56). There were no consistent differences in efficacy between rofecoxib and any of the active comparators at equivalent doses. Endoscopic studies indicated that compared to ibuprofen 800 mg three times a day, rofecoxib caused fewer erosions and gastric ulcers at doses of 25mg and 50mg; the difference in duodenal ulcers was evident only at a dose of 25mg. Rofecoxib 50mg also caused more endoscopically observed ulcers greater than rofecoxib 25mg (RR 2.48 CI: 1.21, 5.11). Very few of the trials reported overall rates of GI adverse events although rofecoxib was found to cause fewer GI events than naproxen. Only one of the nine trials comparing rofecoxib to celecoxib reported on the overall rates of GI events and this was a comparison of the higher recommended dose of rofecoxib with the lower recommended dose of celecoxib. Similarly, the three trials in older hypertensive patients that examined the cardiovascular safety of rofecoxib and celecoxib used non comparable doses; the results of these studies indicated that rofecoxib caused more patients to have oedema and a clinically significant increase in systolic blood pressure. This difference between rofecoxib and celecoxib was not evident in studies conducted in more general populations. AUTHORS' CONCLUSIONS: Rofecoxib was voluntarily withdrawn from global markets in October 2004 therefore there are no implications for practice concerning its use. There remains a number of questions over both the benefits and risks associated with Cox II selective agents and further work is ongoing. PMID- 15654706 TI - Haloperidol plus promethazine for psychosis induced aggression. AB - BACKGROUND: Health services often manage agitated or violent people and for emergency psychiatric services such behaviour is particularly prevalent (10%). The drugs used in this situation should ensure that the person swiftly and safely becomes calm. OBJECTIVES: To examine whether haloperidol plus promethazine is an effective treatment for psychosis induced agitation/aggression. SEARCH STRATEGY: We searched the Cochrane Schizophrenia Group's Register (July 2004). SELECTION CRITERIA: We included all randomised clinical trials involving aggressive people with psychosis for which haloperidol plus promethazine was being used. DATA COLLECTION AND ANALYSIS: We reliably selected, quality assessed and extracted data from all relevant studies. For binary outcomes we calculated standard estimations of risk ratio (RR) and their 95% confidence intervals (CI). Where possible we estimated weighted number needed to treat or harm (NNT/H). MAIN RESULTS: We identified two relevant high quality studies. One compared the haloperidol plus promethazine mix with midazolam (n=301) and one with lorazepam (n=200). The combined results were largely heterogeneous. In Brazil, haloperidol plus promethazine was an effective means of tranquillisation with over two thirds of people being tranquil or sedated by 30 minutes, but midazolam was more swift (n=301, RR 2.9 CI 1.75 to 4.80, NNH 5 CI 3 to 12). In India, however, 95% of people were tranquil or sedated by 30 minutes if allocated to the combination treatment (vs lorazepam, n=200, RR 0.26 CI 0.10 to 0.68, NNT 8 CI 6 to 17). Over the next few hours of treatment reported differences are negligible. One person given midazolam had respiratory depression (reversed by flumazenil), one given lorazepam had respiratory difficulty. A single person given haloperidol plus promethazine had an epileptic fit. Once the initial tranquillisation was administered, few needed additional medications for continued agitation (n=501, 2 RCTs, RR needing additional tranquillising drugs by four hours 1.67 CI 0.62 to 4.54, 4% vs 2%, I squared 50%) and there were no differences in the low levels of use of restraints. About 28% of people in Brazil in both groups had another episode of aggression in the first day after the initial injection (n=301, RR 0.89 CI 0.62 to 1.29). About half of all people in the Indian study were discharged by four hours (n=200, RR 1.13 CI 0.85 to 1.50) and a similar proportion in Brazil by 15 days (n=301, RR 1.05 CI 0.84 to 1.29). Both studies attained 99% follow up for their primary outcomes. Even by two weeks only 4% of people could not be accounted for (n=501, 2 RCTs, RR 0.91 CI 0.38 to 2.17). AUTHORS' CONCLUSIONS: This review suggests that both benzodiazepines work, but that midazolam has a faster onset and thereby reduces the risk of exposure to violence. Both benzodiazepines have the potential to cause respiratory depression, probably midazolam more so than lorazepam, and we would question the use of this group of drugs outside of those services fully confident of observing for and managing the consequences of respiratory distress. Most evidence, however, exists for the haloperidol plus promethazine mix, with currently more than 400 people randomised to the combination. The onset of action is swift and faster than lorazepam. The combination also seems safe with no clear longer term consequences. We would expect policy makers recommending other drug managements to have equally compelling evidence to support their guidance and hope that this would not be founded in conjecture or consensus, which may be more difficult to defend than evidence from high quality studies. PMID- 15654707 TI - Comparative efficacy of epidural, subarachnoid, and intracerebroventricular opioids in patients with pain due to cancer. AB - BACKGROUND: Since the 1970s, when endogenous opioids and opioid receptors were first isolated in the central nervous system, attempts have been made to optimize opioid therapy by delivering the medication centrally rather than systemically. Although the vast majority of cancer patients obtain satisfactory pain relief from individualized systemic treatment, there remain the few whose pain is refractory to systemic treatments. These patients may obtain relief from neuraxial opioid therapy: intracerebroventricular, epidural or subarachnoid. OBJECTIVES: To compare intracerebroventricular therapy with other neuraxial treatments and to determine whether intracerebroventricular (ICV) has anything to offer over epidural (EPI) and subarachnoid (SA) catheters in terms of efficacy, adverse effects, and complications. SEARCH STRATEGY: A number of electronic databases were searched to retrieve information for inclusion in this review. Non English language reports are awaiting assessment. Unpublished data were not sought. SELECTION CRITERIA: Randomised studies of intracerebroventricular therapy for patients with intractable cancer pain were sought. However, this level of evidence was not available so data from uncontrolled trials, retrospective case series and uncontrolled prospective cohort studies were assessed. DATA COLLECTION AND ANALYSIS: Our search did not retrieve any controlled trials. We therefore used data from uncontrolled studies to compare incidences of analgesic efficacy, adverse effects, and complications. We found 72 uncontrolled trials assessing ICV (13 trials, 337 patients), EPI (31 trials, 1343 patients), and SA (28 trials, 722 patients) in cancer patients. From these we extracted data on analgesic efficacy, common pharmacologic adverse effects, and complications. MAIN RESULTS: Data from uncontrolled studies reported excellent pain relief among 73% of ICV patients compared with 72% EPI and 62% SA. Unsatisfactory pain relief was low in all treatment groups. Persistent nausea, persistent and transient urinary retention, transient pruritus, and constipation occurred more frequently with EPI and SA. Respiratory depression, sedation and confusion were most common with ICV. The incidence of major infection when pumps were used with EPI and SA was zero. There was a lower incidence of other complications with ICV therapy than with EPI or SA. AUTHORS' CONCLUSIONS: Neuraxial opioid therapy is often effective for treating cancer pain that has not been adequately controlled by systemic treatment. However, long-term use of neuraxial therapy can be complicated by problems associated with the catheters. The data from uncontrolled studies suggests that ICV is at least as effective against pain as other neuraxial treatments and may be a successful treatment for patients whose cancer pain is resistant to other treatments. PMID- 15654709 TI - Interventions used to improve control of blood pressure in patients with hypertension. AB - BACKGROUND: It is well recognized that patients with high blood pressure (hypertension) in the community frequently fail to meet treatment goals- a condition labeled as "uncontrolled" hypertension. The optimal way in which to organize and deliver care to patients who have hypertension so that they reach treatment goals has not been clearly identified. OBJECTIVES: To determine the effectiveness of interventions to improve control of blood pressure in patients with elevated blood pressure. To evaluate the ability of reminders to improve the follow-up of patients with elevated blood pressure. SEARCH STRATEGY: All-language search of all articles (any year) in the Cochrane Controlled Trials Register (CCTR), Medline and Embase from June 2000. SELECTION CRITERIA: Randomised controlled trials (RCTs) of patients with hypertension that evaluated the following interventions: (1) self-monitoring (2) educational interventions directed to the patient (3) educational interventions directed to the health professional (4) health professional (nurse or pharmacist) led care (5) organisational interventions that aimed to improve the delivery of care (6) appointment reminder systems. OUTCOMES ASSESSED WERE: (1) mean systolic and diastolic blood pressure (2) control of blood pressure (3) proportion of patients followed up at clinic. DATA COLLECTION AND ANALYSIS: Two authors extracted data independently and in duplicate and assessed each study according to the criteria outlined by the Cochrane Collaboration Handbook. MAIN RESULTS: 59 RCTs met our inclusion criteria. The methodological quality of included studies was variable. An organized system of regular review linked to vigorous antihypertensive drug therapy was shown to reduce blood pressure (weighted mean difference -8.2/-4.2 mmHg, -11.7/-6.5 mmHg, -10.6/-7.6 mmHg for 3 strata of entry blood pressure) and all-cause mortality at five years follow-up (6.38% versus 7.78%, difference 1.4%) in a single large RCT- the Hypertension Detection and Follow-Up study. Other interventions had variable effects. Self-monitoring was associated with moderate net reduction in diastolic blood pressure (weighted mean difference (WMD): -2.03 mmHg, 95%CI: -2.69 to -1.38 mmHg, respectively. Appointment reminders increased the proportion of individuals who attended for follow-up. RCTs of educational interventions directed at patients or health professionals were heterogeneous but appeared unlikely to be associated with large net reductions in blood pressure by themselves. Health professional (nurse or pharmacist) led care may be a promising way of delivering care, with the majority of RCTs being associated with improved blood pressure control, but requires further evaluation. AUTHORS' CONCLUSIONS: Family practices and community-based clinics need to have an organized system of regular follow-up and review of their hypertensive patients. Antihypertensive drug therapy should be implemented by means of a systematic stepped care approach when patients do not reach target blood pressure levels. PMID- 15654708 TI - NSAIDS or paracetamol, alone or combined with opioids, for cancer pain. AB - BACKGROUND: NSAIDs are widely applied to treat cancer pain and are frequently combined with opioids in combination preparations for this purpose. However, it is unclear which agent is most clinically efficacious for relieving cancer related pain, or even what may be the additional benefit of combining an NSAID with an opioid in this setting. OBJECTIVES: To assess the effects of NSAIDs, alone or combined with opioids, for the treatment of cancer pain. SEARCH STRATEGY: We searched the Cochrane Central Register of Controlled Trials (Issue 2, 2002), MEDLINE (January 1966 to March 2003), EMBASE (January 1980 to December 2001), LILACS (January 1984 to December 2001) and reference list of articles. SELECTION CRITERIA: Randomized controlled trials and controlled clinical trials that compared NSAID versus placebo; NSAID versus NSAID; NSAID versus NSAID plus opioid; opioid versus opioid plus NSAID; or NSAID versus opioid. DATA COLLECTION AND ANALYSIS: Two reviewers independently assessed trial quality and extracted data. Study authors were contacted for additional information. Adverse event information was collected from trials. Where there was disagreement between reviewers, the opinion of an additional reviewer was sought to resolve the issue. MAIN RESULTS: Forty-two trials involving 3084 patients were included. Clinical heterogeneity of study methods and outcomes precluded meta-analyses and only supported a qualitative systematic review. Seven of eight papers that compared NSAID with placebo demonstrated superior efficacy of NSAID with no difference in side effects. Thirteen papers compared one NSAID with another; four reported increased efficacy of one NSAID over another. Four different studies found that one NSAID had fewer side effects than one or more others. Twenty-three studies compared NSAIDs and opioids in combination or alone with NSAID/opioid combinations. Thirteen out of 14 studies found no difference, or low clinical difference, when combining an NSAID plus an opioid versus either drug alone. Comparisons between various NSAID/opioid combinations were inconclusive. Nine studies assessed the association between dose and efficacy and safety. Four papers demonstrated increased efficacy with increased dose, but no dose-dependent increase in side effects within the dose ranges studied. Study duration ranged from single dose studies performed over six hours to crossover studies lasting six weeks; however the majority of studies were of less than seven days duration. AUTHORS' CONCLUSIONS: Based upon limited data, NSAIDs appear to be more effective than placebo for cancer pain; clear evidence to support superior safety or efficacy of one NSAID over another is lacking; and trials of combinations of an NSAID with an opioid have disclosed either no difference (4 out of 14 papers), a statistically insignificant trend towards superiority (1 out of 14 papers), or at most a slight but statistically significant advantage (9 out of 14 papers), compared with either single entity. The short duration of studies undermines generalization of their findings on efficacy and safety of NSAIDs for cancer pain. PMID- 15654711 TI - Experimental model for observation of micromotion in cell culture. AB - It is known that the micromotion between implant and bone inhibits direct bone growth either on or into implant surfaces in vivo. Nevertheless, biocompatibility tests in vitro of biomaterials for bone/implant interfaces are mainly performed under static conditions. This work describes a dynamic, in vitro experimental simulation of the effect of mutual, small-scale implant surface-tissue displacement on adhered cells. Disks of simulated tissue (PVP hydrogel) were subjected to cyclic micromotion ranging from 0 at the center to 1000 microm at the periphery at approximately 13 Hz, relative to biomaterial surfaces or tissue culture polystyrene controls populated with human osteoblasts in standard tissue culture plate wells. The effect of the interfacial micromotion on the number of cells remaining attached was quantitated by XTT assay. The activity level of the remaining cells was determined by an alkaline phosphatase assay, and cell stress was evaluated by nitrogen assay. Significantly more cells (ANOVA) became detached from similarly prepared surfaces of titanium, hydroxyapatite, and alumina compared to the polystyrene control, and detachment from alumina was greater than for the other two materials. The activity of the remaining attached cells was lower as compared to the static (no micromotion) control but not significantly different among the biomaterials. All nitrogen assays were negative, suggesting minimal cell stress occurred. The method is proposed as a useful and discriminating in vitro tool for biocompatibility studies focused on cell adhesion to biomaterials under conditions related to those which exist at the implant/bone interface in vivo, and it allows subsequent studies of the still viable cells by other methods. PMID- 15654713 TI - A clinical, histological, and computer-based assessment of the Polaris LV, combination diode, and radiofrequency system, for leg vein treatment. AB - BACKGROUND AND OBJECTIVES: Electro-optical synergy (ELOS) is a novel technology that combines radiofrequency (RF) with optical energy. This study investigated the safety and effectiveness of the Polaris LV system, which is based on combined RF and diode laser (915 nm), for the treatment of leg veins. STUDY DESIGN/MATERIALS AND METHODS: Fifty women (Fitzpatrick II-IV) with red or blue leg veins (1-4 mm in diameter) were treated with the Polaris LV, using a fluence of 60-80 J/cm(2) and conducted RF energy of 100 J/cm(3). Patients received up to three treatment sessions at 2- to 4-week intervals. Both patients and an independent physician graded the level of vessel clearance at 2 months following the last treatment, using pre- and post-treatment photographs. Also, a computer generated assessment of vessel clearance was done in 40 patients. Twenty patients provided biopsy specimens for histologic assessment. RESULTS: Approximately three quarters of patients demonstrated >/= 50% vessel clearance, and about 30% had 75% 100% vessel clearance. Computer-generated scores correlated closely with physician scores. Histologic assessment showed signs of coagulation and prominent endothelial degeneration in all treated vessels, but the epidermis remained normal. There were minimal complications. CONCLUSIONS: The Polaris LV is effective and safe in treating red and blue leg veins up to 4 mm in diameter. PMID- 15654712 TI - Enhanced intrinsic biomechanical properties of osteoblastic mineralized tissue on roughened titanium surface. AB - The biological mechanisms underlying bone-titanium integration and biomechanical properties of the integrated bone are poorly understood. This study assesses intrinsic biomechanical properties of mineralized tissue cultured on titanium having different surface topographies. The osteoblastic phenotypes associated with mineral deposition and collagen synthesis underlying the biomechanical modulation are also reported. Rat bone marrow-derived osteoblastic cells were cultured either on the machined titanium disc or acid-etched titanium disc. Nano indentation study of day 28 culture revealed that the mineralized tissue on the acid-etched surface shows 3-3.5 times greater hardness than that on the machined surface (p < 0.01). Elastic modulus of the mineralized tissue was also 2.5-3 times greater on the acid-etched surface than on the machined surface (p < 0.01). After 28 days of culture, mineralized nodule area was significantly lower on the acid-etched surface than on the machined surface (p = 0.0105), while total calcium deposition did not differ between the two surfaces, indicating denser mineral deposition on the acid-etched surface. Osteopontin and osteocalcin gene expressions assayed by the reverse transcriptase-polymerase chain reaction were upregulated in the acid-etched titanium culture. Collagen synthesis measured by Sirius red stain-based colorimetry was 1.5-10 times higher on the acid-etched surface than on the machined surface in the initial culture period of day 1 to day 14 (p < 0.0001). The amount of collagen synthesis corresponded with the enhanced gene expression of prolyl 4-hydroxylase, a key enzyme for post translational modification of collagen chains. Scanning electron microscopic images revealed that tissue cultured on the acid-etched titanium exhibited plate like, compact surface morphology, while the tissue on the machined titanium appeared porous and was covered by fibrous and punctate structures. We conclude that culturing osteoblasts on rougher titanium surfaces enhances hardness and elastic modulus of the mineralized tissue, associated with condensed mineralization, accelerated collagen synthesis, and upregulated expression of selected bone-related genes. PMID- 15654715 TI - Critique of "Sibpair studies implicate chromosome 18 in essential hypertension" by S. Rutherford, M.P. Johnson, and L.R. Griffiths. 2004. Am J Med Genet 126A:241 247. PMID- 15654714 TI - Evaluation of different temperatures in cold air cooling with pulsed-dye laser treatment of facial telangiectasia. AB - BACKGROUND AND OBJECTIVES: Cold air cooling is widely used in dermatological laser therapy. We investigated the influence of cold air cooling at different skin temperatures on therapeutic outcome and side effects of pulsed dye laser treatment of facial telangiectasia. STUDY DESIGN/MATERIALS AND METHODS: From September 2002 to February 2003, 17 patients with previously untreated facial telangiectasia underwent a single treatment session with flash-lamp pulsed dye laser (3.5 J/cm(2), 585 nm, 0.45 milliseconds pulse length, 10 mm beam diameter, Cynosure V). The treatment area was divided into three sub-areas: no cooling, cold air cooling to 20 degrees C and to 17 degrees C skin temperature. The skin temperature was monitored by a prototype infrared sensor system which controlled the temperature of the cold air stream (Cryo5). In a prospective study, we collected data on purpura, pain, clearance, and patient satisfaction on numerical analog scales (NAS) from 0 (meaning "no") to 3 (meaning "high"). RESULTS: Without cooling, purpura (2.53), pain (2.41), and clearance (2.35) were rated medium to high. Cooling to 20 degrees C reduced purpura (1.12) and pain (1.06), whereas the clearance (2.12) was only slightly affected. Cooling to 17 degrees C reduced purpura (0.88) and pain (0.76) even more, the clearance (2.06) was lowered marginally. Most patients preferred cooling to 20 degrees C skin temperature. CONCLUSION: In dermatological laser therapy of facial telangiectasia, the use of cold air cooling can significantly reduce side effects and increase patient satisfaction while only slightly affecting clearance. Cooling to 20 degrees C skin temperature proved to be a well-balanced middle course. For the practical use of cold air cooling, we thus recommend cooling to a level which the patient can tolerate without problems and to try to increase the energy densities. PMID- 15654716 TI - Clinical trial of a novel non-thermal LED array for reversal of photoaging: clinical, histologic, and surface profilometric results. AB - BACKGROUND AND OBJECTIVES: Photomodulation has been described as a process which modifies cell activity using light sources without thermal effect. The objective of this study was to investigate the use of a non-thermal low dose light emitting diode (LED) array for improving the appearance of photoaged subjects. STUDY DESIGN/MATERIALS AND METHODS: This prospective study investigated a random cohort of patients (N = 90) with a wide range of photoaged skin treated by LED photomodulation using a full panel 590 nm non-thermal full face LED array delivering 0.1 J/cm(2) with a specific sequence of pulsing. Subjects were evaluated at 4, 8, 12, 18 weeks and 6 and 12 months after a series of 8 treatments delivered over 4 weeks. Data collected included stereotactic digital imaging, computerized optical digital profilometry, and peri-ocular biopsy histologic evaluations for standard stains and well as collagen synthetic and degradative pathway immunofluorescent staining. RESULTS: Digital imaging data showed a reduction of signs of photoaging in 90% of subjects with smoother texture, reduction of peri-orbital rhytids, and reduction of erythema and pigmentation. Optical profilometry showed a 10% improvement by surface topographical measurements. Histologic data showed markedly increased collagen in the papillary dermis of 100% of post-treatment specimens (N = 10). Staining with anti-collagen I antibodies demonstrated a 28% (range: 10%-70%) average increase in density while staining with anti-matrixmetalloproteinase (MMP)-1 showed an average reduction of 4% (range: 2%-40%). No side effects or pain were noted. CONCLUSIONS: Photomodulation to reverse photoaging is possible with a specific array of LEDs with a specific fluence using a precise pulsing or "code" sequence. Skin textural improvement by digital imaging and surface profilometry is accompanied by increased collagen I deposition with reduced MMP-1 (collagenase) activity in the papillary dermis. This technique is a safe and effective non painful non-ablative modality for improvement of photoaging. PMID- 15654717 TI - Frequency of potential azole drug-drug interactions and consequences of potential fluconazole drug interactions. AB - PURPOSE: To assess the frequency of potential azole-drug interactions and consequences of interactions between fluconazole and other drugs in routine inpatient care. METHODS: We performed a retrospective cohort study of hospitalized patients treated for systemic fungal infections with an oral or intravenous azole medication between July 1997 and June 2001 in a tertiary care hospital. We recorded the concomitant use of medications known to interact with azole antifungals and measured the frequency of potential azole drug interactions, which we considered to be present when both drugs were given together. We then performed a chart review on a random sample of admissions in which patients were exposed to a potential moderate or major drug interaction with fluconazole. The list of azole-interacting medications and the severity of interaction were derived from the DRUGDEX System and Drug Interaction Facts. RESULTS: Among the 4,185 admissions in which azole agents (fluconazole, itraconazole or ketoconazole) were given, 2,941 (70.3%) admissions experienced potential azole-drug interactions, which included 2,716 (92.3%) admissions experiencing potential fluconazole interactions. The most frequent interactions with potential moderate to major severity were co-administration of fluconazole with prednisone (25.3%), midazolam (17.5%), warfarin (14.7%), methylprednisolone (14.1%), cyclosporine (10.7%) and nifedipine (10.1%). Charts were reviewed for 199 admissions in which patients were exposed to potential fluconazole drug interactions. While four adverse drug events (ADEs) caused by fluconazole were found, none was felt to be caused by a drug-drug interaction (DDI), although in one instance fluconazole may have contributed. CONCLUSIONS: Potential fluconazole drug interactions were very frequent among hospitalized patients on systemic azole antifungal therapy, but they had few apparent clinical consequences. PMID- 15654719 TI - Antidiabetic therapy and the risk of heart failure in type 2 diabetic patients: an independent effect or confounding by indication. AB - PURPOSE: This study assesses the effect of the type of antidiabetic treatment on the risk of developing congestive heart failure (CHF) in type 2 diabetes. METHODS: The study was derived from the U.K.-based General Practice Research Database (GPRD) comprised of 3.5 million subjects followed between 1987 and 2001. A total of 21 888 type 2 diabetic patients were identified. A 6:1 matched nested case-control design was employed. Conditional logistic regression was used to derive adjusted odds ratios (ORs) for the association of drug treatment with CHF controlling for diabetes duration and for diseases known to affect the risk of CHF. Antidiabetic drug exposure was defined as the receipt of at least one prescription for an antidiabetic medication within the 3 months prior to the date of CHF diagnosis. RESULTS: There were 1301 incident cases of CHF in the cohort, matched to 7788 controls. After risk factor adjustment, there was a 1.2-fold increase in the risk of CHF for sulphonylureas (SUs) (OR = 1.17; 95%CI = 1.00 1.37) and metformin monotherapies (OR = 1.22; 95%CI = 0.97-1.52), a 1.6-fold increase with combinations of metformin and SUs (OR = 1.62; 95%CI = 1.30-2.02), a 2.2-fold increase with oral tricombinations (OR = 2.16; 95%CI = 0.96-4.86) and a 1.5-fold increase for insulin compared to no exposure (OR = 1.52; 95%CI = 1.06 2.17). Compared to SUs, bicombinations of metformin and SUs showed a statistically significant 1.4-fold increase in the odds of CHF (OR = 1.38; 95%CI = 1.13-1.69). CONCLUSIONS: All antidiabetic medications were associated with an increased likelihood of CHF compared to no antidiabetic exposure. The risk of CHF increased with the complexity of the antidiabetic regimen suggesting that it is the diabetes severity, which imparts risk and not necessarily the antidiabetic regimen itself. PMID- 15654720 TI - National differences in reporting 'pneumonia' and 'pneumonia interstitial': an analysis of the WHO International Drug Monitoring Database on 15 drugs in nine countries for seven pulmonary conditions. AB - PURPOSE: Increased post-marketing reports of 'interstitial pneumonia' as an adverse drug reaction (ADR) from the use of gefitinib, irinotecan, or leflunomide among patients in Japan have not been noted in other countries. The WHO International Drug Monitoring Database was analyzed to examine Japan's pattern of reporting the term 'pneumonia interstitial' for 15 selected drugs with a mixed history of association with pulmonary ADRs. METHODS: ADR counts from the WHO Database for 1992-2001 were obtained for 15 disparate drugs (three androgen blockers, eight cytotoxics, one proton pump inhibitor, one monoclonal antibody, and two anti-epileptics) from nine countries (Australia, France, Germany, Italy, Japan, Spain, Thailand, U.K., and U.S.A.) for seven pulmonary ADR terms (alveolitis fibrosing, pneumonia, pulmonary fibrosis, lung fibrosis interstitial, pulmonary infiltration, interstitial lung disease, and pneumonia interstitial). Statistical analyses included estimating Poisson-distributed expected rates, observed/expected (O/E) ratios, and 95% confidence intervals (CI). The overlapping nature and changes in definition over time of these terms in medical texts and in the WHO-ART, COSTART, J-ART, and MedDRA coding systems is also noted. RESULTS: Compared to other countries, both Japan and France did not have higher O/E reporting ratios for all seven pulmonary ADRs combined, but did have higher O/E ratios for 'pneumonia interstitial' and lower O/E ratios for 'pneumonia' for the same drugs. CONCLUSIONS: Japan and France were found to preferentially use the term 'pneumonia interstitial' for ADR where other countries used 'pneumonia.' This cultural pattern coincides with the fact that 'pneumonia interstitial' in older versions of COSTART, J-ART, and MedDRA were subsumed under infectious pulmonary diseases. PMID- 15654721 TI - Integrating evidence based medicine into clinical practice. PMID- 15654722 TI - Antivirals and antibiotics for influenza in the United States, 1995--2002. AB - PURPOSE: To measure the rates of antiviral and antibiotic prescribing for patients diagnosed with influenza in the United States. METHODS: We performed a retrospective analysis of visits to ambulatory clinics and emergency departments in the National Ambulatory Medical Care Survey (NAMCS) and the National Hospital Ambulatory Medical Care Survey (NHAMCS) with a diagnosis of influenza that occurred in seven influenza seasons between 1 October 1995 and 31 May 2002 (n=1216). RESULTS: There were an estimated 22 million visits (95%CI, 17--26 million visits) with a diagnosis of influenza to community ambulatory clinics (88% of visits), hospital ambulatory clinics (3%) and emergency departments (9%) in the United States between the 1995--1996 and the 2001--2002 influenza seasons, inclusive. The sample was 63% adults, 44% male and 84% white. Physicians prescribed antivirals in 19% of visits and antibiotics not associated with an antibiotic-appropriate diagnosis in 26% of visits. In multivariable modeling, independent predictors of antiviral prescribing were adult age (OR, 2.1; 95%CI, 1.1--4.0) and Medicare insurance (OR, 0.1 compared to private insurance; 95%CI, 0.0--0.6). Antiviral prescribing was marginally associated with influenza season (OR, 1.2 per influenza season; 95%CI, 1.0--1.4). Independent predictors of antibiotic prescribing were influenza season (OR, 0.8 per influenza season; 95%CI, 0.7--0.9), male sex (OR, 0.6; 95%CI, 0.4--0.9), adult age (OR, 2.3; 95%CI, 1.2--4.2) and emergency department visits (OR, 0.5 compared to community ambulatory visits; 95%CI, 0.3--0.8). CONCLUSIONS: Physicians prescribed antiviral medications to 19% of patients they diagnosed with influenza; the proportion that would have been clinically appropriate is unknown. In contrast, physicians prescribed apparently inappropriate antibiotics to 26% of these same patients, a rate that, encouragingly, decreased over time. PMID- 15654723 TI - Some key points emerging from the COX-2 controversy. PMID- 15654724 TI - Determination of oleanolic acid and ursolic acid in cornel by cyclodextrin modified micellar electrokinetic chromatography. AB - A cyclodextrin-modified micellar electrokinetic chromatography (CD-MEKC) method was established for the determination of oleanolic acid and ursolic acid in cornel. The two components were separated in the running buffer of 40 mmol/L sodium borate containing 5% methanol, 25 mmol/L SDS and 15 mmol/L hydroxypropyl beta-cyclodextrin (HP-beta-CD). The applied voltage was 24 kV. The wavelength of detection was 200 nm. The temperature was kept at 25 C. Cinnamic acid was used as the internal standard. The analytical performance of the method was tested with respect to linearity, precision and recovery. The calibration curves were linear in the range of 10.15-243.6 microg/mL, r=0.9993 (oleanolic acid) and 10.07-241.7 microg/mL, r=0.9994 (ursolic acid); the intra-day precision (RSD) was less than 3.7% (oleanolic acid) and 4.1% (ursolic acid); the inter-day precision (RSD) was less than 4.2% (oleanolic acid) and 4.9% (ursolic acid). The limits of detection were 1.6 microg/mL for both components. The method proved to be sensitive, rapid, accurate and suitable for the determination of oleanolic acid and ursolic acid in cornel. PMID- 15654725 TI - Validation and application of a high-performance liquid chromatography--tandem mass spectrometry assay for mosapride in human plasma. AB - A simple, rapid, sensitive and specific liquid chromatography-tandem mass spectrometry method was developed and validated for quantification of mosapride (I), a novel and potent gastroprokinetic agent that enhances the upper gastrointestinal motility by stimulating 5-HT(4) receptor. The analyte and internal standard, tamsulosin (II), were extracted by liquid-liquid extraction with diethyl ether-dichloromethane (70:30, v/v) using a Glas-Col Multi-Pulse Vortexer. The chromatographic separation was performed on a reversed-phase Waters symmetry C(18) column with a mobile phase of 0.03% formic acid-acetonitrile (10:90, v/v). The protonated analyte was quantitated in positive ionization by multiple reaction monitoring with a mass spectrometer. The mass transitions m/z 422.3 -->198.3 and m/z 409.1 -->228.1 were used to measure I and II, respectively. The assay exhibited a linear dynamic range of 0.5-100.0 ng/mL for mosapride in human plasma. The lower limit of quantitation was 500 pg/mL with a relative standard deviation of less than 15%. Acceptable precision and accuracy were obtained for concentrations over the standard curve ranges. A run time of 2.0 min for each sample made it possible to analyze a throughput of more than 400 human plasma samples per day. The validated method has been successfully used to analyze human plasma samples for application in pharmacokinetic, bioavailability or bioequivalence studies. PMID- 15654726 TI - Preparation and evaluation of uniform-sized molecularly imprinted polymer beads used for the separation of sulfamethazine. AB - Uniform-sized molecularly imprinted polymer (MIP) beads were prepared using a one step swelling and polymerization method. The obtained sulfamethazine (SMZ) imprinted polymer showed high affinity and selectivity toward SMZ and other structurally related sulfonamides in acetonitrile or water-acetonitrile mobile phases, particularly in high aqueous systems. The column performance of the MIPs for SMZ and its analogues could be improved by elevating the column temperature and optimizing the flow rate. The hydrogen-bonding effect plays a significant role in the recognition process of SMZ-imprinted polymer systems in organic media, while the ion-exchange effect, as well as hydrophobic effect, dominates the retention mechanism in aqueous-rich media, in addition to shape recognition. PMID- 15654727 TI - An improved method for proteomics studies in C. elegans by fluorogenic derivatization, HPLC isolation, enzymatic digestion and liquid chromatography- tandem mass spectrometric identification. AB - An improved method for proteomics studies, which includes the fluorogenic derivertization of protein mixtures with 7-chloro-4 (dimethylaminoethylaminosulfonyl)-2,1,3-benzoxadiazole (DAABD-Cl), followed by HPLC isolation, enzymatic digestion and identification of the derivatized proteins by HPLC-electrospray ionization (ESI)-MS/MS with the probability-based protein identification algorithm, identified 103 proteins in the soluble extract (10 microg protein) of Caenorhabditis elegans. PMID- 15654728 TI - Non-extractive procedure followed by LC/APCI MS/MS analysis of trimetazidine in plasma samples for assessing bioequivalence of immediate/modified release formulations. AB - Trimetazidine and internal standard [1-(2,4,5-trimethoxybenzyl)piperazine] were isolated from plasma by protein precipitation with trifluoroacetic acid. The neutralized supernatant was separated on a C(8) column with methanol-aqueous 0.11% triethylamine adjusted to pH 3.3 with formic acid (1:4, v/v) at a flow rate of 0.85 mL/min. The separation was achieved within 8 min and the column ef fluent was transferred into an ion trap analyzer via an atmospheric pressure chemical ionization interface. The mass analyzer was used in the selected reaction monitoring mode, to enhance detection selectivity. The method was fully validated with a quantitation limit for trimetazidine of 1.5 ng/mL. The method was successfully applied to assess bioequivalence of two immediate and two modified commercially available pharmaceutical formulations containing 20 and 35 mg of trimetazidine, respectively. PMID- 15654731 TI - Nanoscale borromean rings. AB - The molecular expression of topologically interesting structures represents a formidable challenge for synthetic chemists. The nontrivial link known as the Borromean rings has long been regarded as one of the most ambitious targets in this field. Of ancient provenance, this symbol comprises three interlocked rings in an inseparable union, but cut any one of the rings and the whole assembly unravels into three separate pieces. This Account delineates different strategies that can be applied to the formation of molecules possessing this distinctive topology, culminating with two successful syntheses of such compounds, thus cutting the Gordian knot of topological chemistry. PMID- 15654732 TI - Confusion approach to porphyrinoid chemistry. AB - N-confused porphyrin (NCP) is a porphyrin isomer that is different largely from the parent porphyrin, particularly in the physical, chemical, structural, and coordination properties. Introduction of the confused pyrrole into the normal and expanded porphyrins leads to generation of the confused porphyrinoids having rich structural diversity. In this Account, we introduce a series of N-confused porphyrinoids recently synthesized and highlight their properties such as fusion, peripheral N coordination, supramolecular assemblies, anion binding, and singlet oxygen sensitization. PMID- 15654733 TI - Investigations on butterfly Fe/S cluster S-centered anions (mu-S-)2Fe2(CO)6, (mu S-)(mu-RS)Fe2(CO)6, and related species. AB - This Account describes the formation and chemical reactivities of the novel butterfly Fe/S cluster anions (mu-S(-))(2)Fe(2)(CO)(6) (I), (mu-S(-))(mu RS)Fe(2)(CO)(6) (II), (mu-S(-))(mu-RS)[Fe(2)(CO)(6)](2)(mu(4)-S) (III), (mu-S( ))(mu-RS)[Fe(2)(CO)(6)](3)(mu(4)-S)(2) (IV), and {(mu-S(-))(mu(4) S)[Fe(2)(CO)(6)](2)}(2)(mu-SZS-mu) (V). It also gives a description of the novel structures, unique properties, and wide applications of the corresponding butterfly Fe/S cluster complexes produced from these anions. Anions I-V along with their products are important in the development of organometallic chemistry, cluster chemistry, catalysis, and material and life sciences. PMID- 15654734 TI - Computational studies on the cyclizations of enediynes, enyne-allenes, and related polyunsaturated systems. AB - Quantum chemical studies of cyclizations of enediynes and enyne-allenes have proven to be computationally tractable thanks to the success of the unrestricted broken spin symmetry (UBS) approach using GGA functionals for the description of open-shell biradicals; the results can further be improved through single-point energy coupled-cluster computations [CCSD(T), BD(T)]. This made comprehensive computational studies on substituent effects and heterosubstituted systems possible. For convenience and predicting new reactions, these transformations can be grouped within larger "families". Alternative cyclization modes are predicted and await experimental realization. PMID- 15654735 TI - Ordering fullerene materials at nanometer dimensions. AB - The combination of the hydrophobic fullerene core with hydrophilic functional groups of both ionic and nonionic nature produces organized structures with sizes that range from nanometer to micrometer length scales. The driving force toward this spontaneous organization is the amphiphilic character of the fullerene derivatives. To control both shape and size of the supramolecular assemblies and ultimately their function, important parameters that must be tuned are (i) the balance between the hydrophobic and the hydrophilic moieties, (ii) the effect of the environment, typically provided by simple solvents, (iii) the interface on which the aggregation occurs or, more precisely, surface templating effects, and (iv) the solvation process. PMID- 15654736 TI - Catalytic transmetalation from group 6 Fischer carbene complexes: an emerging powerful tool in organic synthesis. AB - The chemistry of metal carbene complexes has experienced an enormous development in the past decades. Despite this fact, the use of transition metals as catalysts in reactions involving group 6 Fischer carbene complexes was virtually neglected. Here, we describe how the reactivity of these compounds can be enhanced or modified in the presence of catalytic amounts of a transition metal, leading to new forms of reactivity and others offering clear advantages in terms of efficiency over the uncatalyzed reactions. The key step for these reactions is the transmetalation from the stoichiometric metal carbene reagent to the catalyst. This process generates a new metal-carbene complex that leads to enhanced reactivity of new reaction modes. Two examples of Fischer carbene complexes obtained by transmetalation to Pd and Cu have been isolated during the last 2 years, showing the flexibility if the mechanistic hypothesis for these reactions. The work presented in this Account shows how an area, which was practically unexplored 5 years ago, has emerged as a new and powerful field of research. PMID- 15654737 TI - Bioinspired hydrogen bond motifs in ligand design: the role of noncovalent interactions in metal ion mediated activation of dioxygen. AB - Hydrogen bonds influence secondary coordination spheres around metal ions in many proteins. To duplicate these features of molecular architecture in synthetic systems, urea-based ligands have have been developed that create rigid organic frameworks when bonded to metal ions. These frameworks position hydro-gen bond donors proximal to metal ion(s) to form specific chem-ical microenvironments. Iron(II) and manganese(II) complexes with constrained cavities activate O(2), yielding M(III) (M(III) = Fe and Mn) complexes with terminal oxo ligands. Installation of anionic sites within the cavity assists the formation of complexes with M(II/III)-OH and M(III)-O units derived directly from water. Opening the cavity promotes M(mu-O)(2)M rhombs, as illustrated by isolation of a cobalt(III) analogue, the stability of which is promoted by the hydrogen bonds surrounding the bridging oxo ligands. PMID- 15654738 TI - How to hide zinc in a small protein. AB - Small cysteine-rich proteins (metallothioneins) and related domains of some large proteins (e.g., lysine methyltransferases) bind tri- and tetranuclear zinc clusters with topologies resembling fragments of Zn(II) sulfide minerals. These clusters are ubiquitous in animals, plants, and bacteria. Bacterial metallothioneins can also contain histidines as cluster ligands and embed Zn(II) with a "treble-clef"-like finger fold. This unusual embedded Zn(II) is "hidden" and surprisingly inert toward Zn or Cd exchange. Clearly, proteins can exert fine control over both the thermodynamics and kinetics of zinc binding in thiolate clusters. Genome sequences suggest that related zinc-finger sites are common in a variety of bacteria. PMID- 15654739 TI - Structural properties of the promiscuous VP16 activation domain. AB - Herpes simplex virion protein 16 (VP16) contains two strong activation regions that can independently and cooperatively activate transcription in vivo. We have identified the regions and residues involved in the interaction with the human transcriptional coactivator positive cofactor 4 (PC4) and the general transcription factor TFIIB. NMR and biochemical experiments revealed that both VP16 activation regions are required for the interaction and undergo a conformational transition from random coil to alpha-helix upon binding to its target PC4. The interaction is strongly electrostatically driven and the binding to PC4 is enhanced by the presence of its amino-terminal domain. We propose models for binding of VP16 to the core domains of PC4 and TFIIB that are based on two independent docking approaches using NMR chemical shift changes observed in titration experiments. The models are consistent with results from site-directed mutagenesis and provide an explanation for the contribution of both acidic and hydrophobic residues for transcriptional activation by VP16. Both intrinsically unstructured activation domains are attracted to their interaction partner by electrostatic interactions, and adopt an alpha-helical conformation around the important hydrophobic residues. The models showed multiple distinct binding surfaces upon interaction with various partners, providing an explanation for the promiscuous properties, cooperativity, and the high activity of this activation domain. PMID- 15654740 TI - Calcium and magnesium binding to human centrin 3 and interaction with target peptides. AB - There are four isoforms of centrin in mammals, with variable sequence, tissue expression, and functional properties. We have recently characterized a number of structural, ion, and target binding properties of human centrin isoform HsCen2. This paper reports a similar characterization of HsCen3, overexpressed in Escherichia coli and purified by phase-reversed chromatography. Equilibrium and dynamic binding studies revealed that HsCen3 has one mixed Ca(2+)/Mg(2+) binding site of high affinity (K(d) = 3 and 10 microM for Ca(2+) and Mg(2+), respectively) and two Ca(2+)-specific sites of low affinity (K(d) = 140 microM). The metal-free protein is fragmented by an unidentified protease into a polypeptide segment of 11 kDa, which was purified by HPLC, and identified by mass spectrometry as the segment of residues 21-112. Similarly, controlled trypsinolysis on Ca(2+)-bound HsCen3 yielded a mixture of segments of residues 1 124 and 1-125. The Ca(2+)/Mg(2+) site could be assigned to this segment and thus resides in the N-terminal half of HsCen3. Temperature denaturation experiments, circular dichroism, and utilization of fluorescence hydrophobic probes allowed us to propose that the metal-free protein has molten globule characteristics and that the dication-bound forms are compact with a polar surface for the Mg(2+) form and a hydrophobic exposed surface for the Ca(2+) form. Thus, HsCen3 could be classified as a Ca(2+) sensor protein. In addition, it is able to bind strongly to a model target peptide (melittin), as well as to peptides derived from the protein XPC and Kar1p, with a moderate Ca(2+) dependence. PMID- 15654741 TI - Isolation, solution structure, and insecticidal activity of kalata B2, a circular protein with a twist: do Mobius strips exist in nature? AB - A large number of macrocyclic miniproteins with diverse biological activities have been isolated from the Rubiaceae, Violaceae, and Cucurbitaceae plant families in recent years. Here we report the three-dimensional structure determined using (1)H NMR spectroscopy and demonstrate potent insecticidal activity for one of these peptides, kalata B2. This peptide is one of the major components of an extract from the leaves of the plant Oldenlandia affinis. The structure consists of a distorted triple-stranded beta-sheet and a cystine knot arrangement of the disulfide bonds and is similar to those described for other members of the cyclotide family. The unique cyclic and knotted nature of these molecules makes them a fascinating example of topologically complex proteins. Examination of the sequences reveals that they can be separated into two subfamilies, one of which contains a larger number of positively charged residues and has a bracelet-like circularization of the backbone. The second subfamily contains a backbone twist due to a cis-peptidyl-proline bond and may conceptually be regarded as a molecular Mobius strip. Kalata B2 is the second putative member of the Mobius cyclotide family to be structurally characterized and has a cis peptidyl-proline bond, thus validating the suggested name for this subfamily of cyclotides. The observation that kalata B2 inhibits the growth and development of Helicoverpa armigera larvae suggests a role for the cyclotides in plant defense. A comparison of the sequences and structures of kalata B1 and B2 provides insight into the biological activity of these peptides. PMID- 15654742 TI - Mapping and analysis of the lytic and fusogenic domains of surfactant protein B. AB - Surfactant protein B (SP-B) is a hydrophobic, 79 amino acid peptide that regulates the structure and function of surfactant phospholipid membranes in the airspaces of the lung. Addition of SP-B to liposomes composed of DPPC/PG (7:3) leads to membrane binding, destabilization, and fusion, ultimately resulting in rearrangement of membrane structure. The goal of this study was to map the fusogenic and lytic domains of SP-B and assess the effects of altered fusion and lysis on surface activity. Synthetic peptides were generated to predicted helices and/or interhelical loops of SP-B and tested for fusion, lytic, and surface activities. The N-terminal half of SP-B (residues 1-37), which includes the nonhelical N-terminal amino acids in addition to helices 1 and 2, promoted rapid liposome fusion whereas shorter peptides were significantly less effective. The requirements for optimal surface tension reduction were similar to those for fusion; in contrast, helix 1 (residues 7-22) alone was sufficient for liposome lysis. The C-terminal half of SP-B (residues 43-79), which includes helices 3, 4, and 5, exhibited significantly lower levels of fusogenic, lytic, and surface tension reducing activities compared to the N-terminal region. These results indicate that SP-B fusion, lytic and surface activities map predominantly to the N-terminal half of SP-B. Amino acid substitutions in synthetic peptides corresponding to the N-terminal half of SP-B indicated that, in general, decreased fusion or lytic activities were associated with altered surface tension reducing properties of the peptide. However, the presence of fusion and lytic activities alone could not account for the surface tension reducing property of SP-B. We propose a model in which association of helix 1 with lipids leads to membrane permeabilization but not aggregation; helix 2 mediates membrane cross linking (aggregation), which, in turn, facilitates lipid mixing, membrane fusion, and interfacial adsorption/surface tension reduction. PMID- 15654743 TI - Cholera toxin entry into pig enterocytes occurs via a lipid raft- and clathrin dependent mechanism. AB - The small intestinal brush border is composed of lipid raft microdomains, but little is known about their role in the mechanism whereby cholera toxin gains entry into the enterocyte. The present work characterized the binding of cholera toxin B subunit (CTB) to the brush border and its internalization. CTB binding and endocytosis were performed in organ-cultured pig mucosal explants and studied by fluorescence microscopy, immunogold electron microscopy, and biochemical fractionation. By fluorescence microscopy CTB, bound to the microvillar membrane at 4 degrees C, was rapidly internalized after the temperature was raised to 37 degrees C. By immunogold electron microscopy CTB was seen within 5 min at 37 degrees C to induce the formation of numerous clathrin-coated pits and vesicles between adjacent microvilli and to appear in an endosomal subapical compartment. A marked shortening of the microvilli accompanied the toxin internalization whereas no formation of caveolae was observed. CTB was strongly associated with the buoyant, detergent-insoluble fraction of microvillar membranes. Neither CTB's raft association nor uptake via clathrin-coated pits was affected by methyl-beta cyclodextrin, indicating that membrane cholesterol is not required for toxin binding and entry. The ganglioside GM(1) is known as the receptor for CTB, but surprisingly the toxin also bound to sucrase-isomaltase and coclustered with this glycosidase in apical membrane pits. CTB binds to lipid rafts of the brush border and is internalized by a cholesterol-independent but clathrin-dependent endocytosis. In addition to GM(1), sucrase-isomaltase may act as a receptor for CTB. PMID- 15654744 TI - Allosteric regulation and communication between subunits in uracil phosphoribosyltransferase from Sulfolobus solfataricus. AB - Uracil phosphoribosyltransferase (UPRTase) catalyzes the conversion of 5 phosphate-alpha-1-diphosphate (PRPP) and uracil to uridine 5'-monophosphate (UMP) and diphosphate. The UPRTase from Sulfolobus solfataricus has a unique regulation by nucleoside triphosphates compared to UPRTases from other organisms. To understand the allosteric regulation, crystal structures were determined for S. solfataricus UPRTase in complex with UMP and with UMP and the allosteric inhibitor CTP. Also, a structure with UMP bound in half of the active sites was determined. All three complexes form tetramers but reveal differences in the subunits and their relative arrangement. In the UPRTase-UMP complex, the peptide bond between a conserved arginine residue (Arg80) and the preceding residue (Leu79) adopts a cis conformation in half of the subunits and a trans conformation in the other half and the tetramer comprises two cis-trans dimers. In contrast, four identical subunits compose the UPRTase-UMP-CTP tetramer. CTP binding affects the conformation of Arg80, and the Arg80 conformation in the UPRTase-UMP-CTP complex leaves no room for binding of the substrate PRPP. The different conformations of Arg80 coupled to rearrangements in the quaternary structure imply that this residue plays a major role in regulation of the enzyme and in communication between subunits. The ribose ring of UMP adopts alternative conformations in the cis and trans subunits of the UPRTase-UMP tetramer with associated differences in the interactions of the catalytically important Asp209. The active-site differences have been related to proposed kinetic models and provide an explanation for the regulatory significance of the C-terminal Gly216. PMID- 15654745 TI - On the catalytic role of the conserved active site residue His466 of choline oxidase. AB - The oxidation of alcohols to aldehydes is catalyzed by a number of flavin dependent enzymes, which have been grouped in the glucose-methanol-choline oxidoreductase enzyme superfamily. These enzymes exhibit little sequence similarity in their substrates binding domains, but share a highly conserved catalytic site, suggesting a similar activation mechanism for the oxidation of their substrates. In this study, the fully conserved histidine residue at position 466 of choline oxidase was replaced with an alanine residue by site directed mutagenesis and the biochemical, spectroscopic, and mechanistic properties of the resulting CHO-H466A mutant enzyme were characterized. CHO-H466A showed k(cat) and k(cat)/K(m) values with choline as substrate that were 60- and 1000-fold lower than the values for the wild-type enzyme, while the k(cat)/K(m) value for oxygen was unaffected, suggesting the involvement of His(466) in the oxidation of the alcohol substrate but not in the reduction of oxygen. Replacement of His(466) with alanine significantly affected the microenvironment of the flavin, as indicated by the altered behavior of CHO-H466A with sulfite and dithionite. In agreement with this conclusion, a midpoint reduction potential of +106 mV for the two-electron transfer in the catalytically competent enzyme product complex was determined at pH 7 for CHO-H466A, which was approximately 25 mV more negative than that of the wild-type enzyme. Enzymatic activity in CHO H466A could be partially rescued with exogenous imidazolium, but not imidazole, consistent with the protonated form of histidine exerting a catalytic role. pH profiles for glycine betaine inhibition, the deprotonation of the N(3)-flavin locus, and the k(cat)/K(m) value for choline all showed a significant shift upward in their pK(a) values, consistent with a change in the polarity of the active site. Finally, kinetic isotope effects with isotopically labeled substrate and solvent indicated that the histidine to alanine substitution affected the timing of substrate OH and CH bond cleavages, consistent with removal of the hydroxyl proton being concerted with hydride transfer in the mutant enzyme. All taken together, the results presented in this study suggest that in choline oxidase, His(466) modulates the electrophilicity of the enzyme-bound flavin and the polarity of the active site, and contributes to the stabilization of the transition state for the oxidation of choline to betaine aldehyde. PMID- 15654747 TI - Role of a surface tryptophan in defining the structure, stability, and DNA binding of the hyperthermophile protein Sac7d. AB - Sac7d is a small, chromatin protein from Sulfolobus acidocaldarius which induces a sharp kink in DNA with intercalation of valine and methionine side chains. The crystal structure of the protein-DNA complex indicates that a surface tryptophan (W24) plays a key role in DNA binding by hydrogen bonding to the DNA at the kink site. We show here that substitution of the solvent-exposed tryptophan with alanine (W24A) led to a significant loss in not only DNA binding affinity but also protein stability. The W24A substitution proved to be one of the most destabilizing surface substitutions in Sac7d. A global linkage analysis of the pH and salt dependence of stability indicated that the protein stability surface (DeltaG vs temperature, pH, and salt concentration) was lowered overall by 2 kcal/mol (from 0 to 100 degrees C, pH 0 to 7, and 0 to 0.3 M KCl). The lower free energy of unfolding could not be attributed to significant structural perturbations of surface electrostatic interactions. Residual dipolar coupling of partially aligned protein and the NMR solution structure of W24A confirmed that the surface substitution resulted in no significant change in structure. Stabilization of this hyperthermophile protein and its DNA complex by a surface cluster of hydrophobic residues involving W24 and the two intercalating side chains is discussed. PMID- 15654746 TI - Dynamic motion of helix A in the amino-terminal domain of calmodulin is stabilized upon calcium activation. AB - Calcium-dependent changes in the internal dynamics and average structures of the opposing globular domains of calmodulin (CaM), as well as their relative spatial arrangement, contribute to the productive association between CaM and a range of different target proteins, affecting their functional activation. To identify dynamic structural changes involving individual alpha-helical elements and their modulation by calcium activation, we have used site-directed mutagenesis to engineer a tetracysteine binding motif within helix A near the amino terminus of calmodulin (CaM), permitting the selective and rigid attachment of the fluorescent probe 4',5'-bis(1,3,2-dithioarsolan-2-yl)fluorescein (FlAsH) with full retention of function. The rigid tetracoordinate linkage of FlAsH to CaM, in conjunction with frequency domain fluorescence anisotropy measurements, allows assessment of dynamic changes associated with calcium activation without interference from independent probe motion. Taking advantage of the large fluorescence enhancement associated with binding of FlAsH to CaM, we determined rates of binding of FlAsH to apo-CaM and calcium-activated CaM to be 2800 +/- 80 and 310 +/- 10 M(-)(1) s(-)(1), respectively. There is no difference in the solvent accessibility of the bound FlAsH irrespective of calcium binding to CaM. Thus, given that FlAsH selectively labels disordered structures, the large difference in rates of FlAsH binding indicates that calcium binding stabilizes helix A. Frequency domain anisotropy measurements of bound FlAsH indicate that prior to calcium activation, helix A undergoes large amplitude nanosecond motions. Following calcium activation, helix A becomes immobile, and structurally coupled to the overall rotation of CaM. We discuss these results in the context of a model that suggests stabilization of helix A relative to other domain elements in the CaM structure is critical to defining high-affinity binding clefts, and in promoting specific and ordered binding of the opposing lobes of CaM to target proteins. PMID- 15654748 TI - MAPK modulates the DNA binding of adipocyte enhancer-binding protein 1. AB - Adipocyte enhancer-binding protein 1 (AEBP1) is a down-regulator of adipogenesis through its transcriptional repression activity, as well as through its interaction with mitogen-activated protein kinase (MAPK), which protects MAPK from its specific phosphatases. This study increases our understanding of the mechanisms of DNA binding by AEBP1, the first step in its function as a transcriptional repressor. We show that DNA binding by AEBP1 requires both the N- and C-terminal domains of AEBP1, and MAPK interaction with AEBP1 (through its N terminus) results in enhanced DNA binding. A threonine at position 623 within the C-terminal domain of AEBP1 plays an important role in DNA binding by AEBP1, because the mutation results in decreased DNA binding by AEBP1, which leads to a decrease in the transcriptional repression ability of AEBP1. We also show that in vitro phosphorylation of AEBP1 by MAPK is greatly reduced upon mutation of T623. These results suggest that MAPK regulates the transcriptional activity of AEBP1 by a novel dual mechanism, in which MAPK interaction enhances and subsequent phosphorylation decreases the DNA-binding ability of AEBP1. PMID- 15654749 TI - Integrin-linked kinase complexes with caveolin-1 in human neuroblastoma cells. AB - Integrin-linked kinase (ILK) and caveolin-1 (cav-1) are implicated in the pathogenesis of cancer. Overexpression of ILK leads to altered expression of cell cycle regulators, a decreased level of cell adhesion to the extracellular matrix, a decreased level of apoptosis, in vitro phosphorylation of Akt, and tumor formation in nude mice. Conversely, cav-1 expression is frequently downregulated in many forms of cancer. We examined whether ILK and cav-1 interact in SHEP human neuroblastoma cells because ILK is present in caveolae-enriched membranes and contains a putative cav-binding domain. SHEP cells were stably transfected with vector, wild-type ILK (ILK-wt), kinase-deficient ILK (ILK-kd), or mutant cav binding domain ILK (ILK-mutCavbd). Control SHEP cells and ILK transfectants express high levels of ILK and cav-1. Immunoprecipitation with anti-cav-1 co immunoprecipitates a 59 kDa protein that is immunoreactive with the anti-ILK antibody, and this interaction is partially prevented in cells expressing ILK mutCavbd. Cav-1 and ILK partially colocalize in SHEP cells, also supporting these data. Last, affinity chromatography with a biotinylated cav-scaffolding domain peptide precipitates ILK-wt but not ILK-mutCavbd. These data suggest that the cav binding domain of ILK and the cav-scaffolding domain of cav-1 mediate complex formation in human neuroblastoma cells. PMID- 15654750 TI - Resonance Raman evidence for the presence of two heme pocket conformations with varied activities in CO-bound bovine soluble guanylate cyclase and their conversion. AB - Resonance Raman (RR) spectra of soluble guanylate cyclase (sGC) reported by five independent research groups have been classified as two types: sGC(1) and sGC(2). Here we demonstrate that the RR spectra of sGC isolated from bovine lung contain only sGC(2) while both species are observed in the spectra of the CO-bound form (CO-sGC). The relative populations of the two forms were altered from an initial composition in which the CO-sGC(2) form predominated, with the Fe-CO (nu(Fe)( )(CO)) and C-O stretching modes (nu(CO)) at 472 and 1985 cm(-)(1), respectively, to a composition dominated by the CO-sGC(1) form with nu(Fe)(-)(CO) and nu(CO) at 488 and 1969 cm(-)(1), respectively, following the addition of a xenobiotic, YC 1. Further addition of a substrate, GTP, completed the change. GDP and cGMP had a significantly weaker effect, while a substrate analogue, GTP-gamma-S, had an effect similar to that of GTP. In contrast, ATP had a reverse effect, and suppressed the effects of YC-1 and GTP. In the presence of both YC-1 and GTP, vinyl vibrations of heme were significantly influenced. New CO isotope-sensitive bands were observed at 521, 488, 363, and 227 cm(-)(1). The 521 cm(-)(1) band was assigned to the five-coordinate (5c) species from the model compound studies using ferrous iron protoporphyrin IX in CTAB micelles. Distinct from the 472 cm( )(1) species, both the 488 and 521 cm(-)(1) species were apparently un photodissociable when an ordinary Raman spinning cell was used, indicating rapid recombination of photodissociated CO. On the basis of these findings, binding of YC-1 to the heme pocket is proposed. PMID- 15654751 TI - The aromatic domain of the coronavirus class I viral fusion protein induces membrane permeabilization: putative role during viral entry. AB - Coronavirus (CoV) entry is mediated by the viral spike (S) glycoprotein, a class I viral fusion protein. During viral and target cell membrane fusion, the heptad repeat (HR) regions of the S2 subunit assume a trimer-of-hairpins structure, positioning the fusion peptide in close proximity to the C-terminal region of the ectodomain. The formation of this structure appears to drive apposition and subsequent fusion of viral and target cell membranes; however, the exact mechanism is unclear. Here, we characterize an aromatic amino acid rich region within the ectodomain of the S2 subunit that both partitions into lipid membranes and has the capacity to perturb lipid vesicle integrity. Circular dichroism analysis indicated that peptides analogous to the aromatic domains of the severe acute respiratory syndrome (SARS)-CoV, mouse hepatitis virus (MHV) and the human CoV OC43 S2 subunits, did not have a propensity for a defined secondary structure. These peptides strongly partitioned into lipid membranes and induced lipid vesicle permeabilization at peptide/lipid ratios of 1:100 in two independent leakage assays. Thus, partitioning of the peptides into the lipid interface is sufficient to disorganize membrane integrity. Our study of the S2 aromatic domain of three CoVs provides supportive evidence for a functional role of this region. We propose that, when aligned with the fusion peptide and transmembrane domains during membrane apposition, the aromatic domain of the CoV S protein functions to perturb the target cell membrane and provides a continuous track of hydrophobic surface, resulting in lipid-membrane fusion and subsequent viral nucleocapsid entry. PMID- 15654752 TI - Equilibrium and kinetic analysis of nucleotide binding to the DEAD-box RNA helicase DbpA. AB - The Escherichia coli DEAD-box protein A (DbpA) is an RNA helicase that utilizes the energy from ATP binding and hydrolysis to facilitate structural rearrangements of rRNA. We have used the fluorescent nucleotide analogues, mantADP and mantATP, to measure the equilibrium binding affinity and kinetic mechanism of nucleotide binding to DbpA in the absence of RNA. Binding generates an enhancement in mant-nucleotide fluorescence and a corresponding reduction in intrinsic DbpA fluorescence, consistent with fluorescence resonance energy transfer (FRET) from DbpA tryptophan(s) to bound nucleotides. Fluorescent modification does not significantly interfere with the affinities and kinetics of nucleotide binding. Different energy transfer efficiencies between DbpA-mantATP and DbpA-mantADP complexes suggest that DbpA adopts nucleotide-dependent conformations. ADP binds (K(d) approximately 50 microM at 22 degrees C) 4-7 times more tightly than ATP (K(d) approximately 400 microM at 22 degrees C). Both nucleotides bind with relatively temperature-independent association rate constants (approximately 1-3 microM(-1) s(-1)) that are much lower than predicted for a diffusion-limited reaction. Differences in the binding affinities are dictated primarily by the dissociation rate constants. ADP binding occurs with a positive change in the heat capacity, presumably reflecting a nucleotide-induced conformational rearrangement of DbpA. At low temperatures (<22 degrees C), the binding free energies are dominated by favorable enthalpic and unfavorable entropic contributions. At physiological temperatures (>22 degrees C), ADP binding occurs with positive entropy changes. We favor a mechanism in which ADP binding increases the conformational flexibility and dynamics of DbpA. PMID- 15654754 TI - Heme binding to the histidine-rich protein II from Plasmodium falciparum. AB - The histidine-rich protein II (HRP II) from Plasmodium falciparum has been implicated in the formation of hemozoin, a detoxified, crystalline form of ferric protoporphyrin IX (Fe(3+)-PPIX) produced by the parasite. Fe(3+)-PPIX titrations coupled with quantitative amino acid analysis showed that HRP II binds 15 Fe(3+) PPIX molecules per 30 kDa monomer. Circular dichroism spectroscopy was used to probe the secondary structure of HRP II with and without bound Fe(3+)-PPIX. These studies have revealed large changes in the secondary structure with Fe(3+)-PPIX binding, changing from a random coil in the absence of Fe(3+)-PPIX to a more ordered helical structure in the presence of Fe(3+)-PPIX. The Fe(3+)-PPIX-bound HRP II structure most closely resembles a 3(10)-helix. Coincident with this structural change caused by Fe(3+)-PPIX binding, the formation of an intermolecular disulfide bond occurs between HRP II monomers. In vitro pull-down assays show an interaction between monomers that is dependent on the presence of Fe(3+)-PPIX. One model that best fits with the data reported here requires formation of 15 intermolecular bishistidyl ligated Fe(3+)-PPIX molecules arranged in a head to head fashion, which would then allow for the formation of an intermolecular disulfide bond. The structure best able to accommodate these requirements is a 3(10)-helix. PMID- 15654753 TI - Mass spectrometric identification of lysines involved in the interaction of human replication protein a with single-stranded DNA. AB - Human replication protein A (hRPA), a heterotrimeric single-stranded DNA (ssDNA) binding protein, is required for many cellular pathways including DNA damage repair, recombination, and replication as well as the ATR-mediated DNA damage response. While extensive effort has been devoted to understanding the structural relationships between RPA and ssDNA, information is currently limited to the RPA domains, the trimerization core, and a partial cocrystal structure. In this work, we employed a mass spectrometric protein footprinting method of single amino acid resolution to investigate the interactions of the entire heterotrimeric hRPA with ssDNA. In particular, we monitored surface accessibility of RPA lysines with NHS biotin modification in the contexts of the free protein and the nucleoprotein complex. Our results not only indicated excellent agreement with the available crystal structure data for RPA70 DBD-AB-ssDNA complex but also revealed new protein contacts in the nucleoprotein complex. In addition to two residues, K263 and K343 of p70, previously identified by cocrystallography as direct DNA contacts, we observed protection of five additional lysines (K183, K259, K489, K577, and K588 of p70) upon ssDNA binding to RPA. Three residues, K489, K577, and K588, are located in ssDNA binding domain C and are likely to establish the direct contacts with cognate DNA. In contrast, no ssDNA-contacting lysines were identified in DBD-D. In addition, two lysines, K183 and K259, are positioned outside the putative ssDNA binding cleft. We propose that the protection of these lysines could result from the RPA interdomain structural reorganization induced by ssDNA binding. PMID- 15654755 TI - Amino acid substitution and modification resulting from Escherichia coli expression of recombinant Plasmodium falciparum histidine-rich protein II. AB - The histidine-rich protein II (HRP II) from Plasmodium falciparum is an unusual protein composed of 40% alanine, 36% histidine, and 11% aspartate residues. Expression of HRP II in Escherichia coli results in the isolation of a heterogeneous protein. Mass spectrometry reveals a reduction in mass by multiples of 9 Da from the expected molecular mass that can be attributed to the substitution of glutamine for some histidine residues in the sequence. The extent of the glutamine for histidine substitution can be reduced by slowing the expression rate. Mass spectral analysis of HRP II also revealed alpha-amino methylation of the N-terminal alanine residue of HRP II. PMID- 15654756 TI - Covalent binding and interstrand cross-linking of duplex DNA by dirhodium(II,II) carboxylate compounds. AB - The study of the interactions of double-stranded (ds) DNA with the dirhodium carboxylate compounds Rh(2)(O(2)CCH(3))(4)(H(2)O)(2) (Rh1), [Rh(2)(O(2)CCH(3))(2)(CH(3)CN)(6)](BF(4))(2) (Rh2), and Rh(2)(O(2)CCF(3))(4) (Rh3) supports the presence of covalently linked DNA adducts, including stable DNA interstrand cross-links. The present biochemical study refutes earlier claims that no reaction between dirhodium compounds and dsDNA occurs. The reversal behavior of these interstrand cross-links in 5 M urea at 95 degrees C (for different heating times) implies the presence of various coordination modes involving ax/ax, ax/eq, and eq/eq DNA interactions with the dirhodium core. The reaction rates of the dirhodium compounds with dsDNA were determined spectroscopically and are in the order Rh1 << Rh2 < Rh3. This difference in behavior of the three dirhodium compounds correlates with the lability of the leaving groups and corresponds to the extent of interstrand cross-link formation by these compounds on a 123 bp DNA fragment, as observed by denaturing polyacrylamide gel electrophoresis (dPAGE). Since all three dirhodium compounds form covalent Rh-DNA adducts, including interstrand cross-links, it is important that DNA be considered a potential target for biological activity of these dirhodium carboxylate compounds. PMID- 15654757 TI - Influence of flanking residues on tilt and rotation angles of transmembrane peptides in lipid bilayers. A solid-state 2H NMR study. AB - To gain insight into the parameters that determine the arrangement of proteins in membranes, (2)H NMR experiments were performed to analyze tilt and rotation angles of membrane-spanning alpha-helical model peptides upon incorporation in diacylphosphatidylcholine bilayers with varying thickness. The peptides consisted of the sequence acetyl-GW(2)(LA)(8)LW(2)A-NH(2) (WALP23) and analogues thereof, in which the interfacial Trp residues were replaced by Lys (KALP23) and/or the hydrophobic sequence was replaced by Leu (WLP23 and KLP23). The peptides were synthesized with a single deuterium-labeled alanine at four different positions along the hydrophobic segment. For all peptides a small but systematic increase in tilt angle was observed upon decreasing the bilayer thickness. However, significantly larger tilt angles were obtained for the Lys-flanked KALP23 than for the Trp-flanked WALP23, suggesting that interfacial anchoring interactions of Trp may inhibit tilting. Increasing the hydrophobicity resulted in an increase in tilt angle for the Trp-flanked analogue only. For all peptides the maximum tilt angle obtained was remarkably small (less than 12 degrees ), suggesting that further tilting is inhibited, most likely due to unfavorable packing of lipids around a tilted helix. The results furthermore showed that the direction of tilt is determined almost exclusively by the flanking residues: Trp- and Lys-flanked peptides were found to have very different rotation angles, which were influenced significantly neither by hydrophobicity of the peptides nor by the extent of hydrophobic mismatch. Finally, very small changes in the side chain angles of the deuterated alanine probes were observed in Trp-flanked peptides, suggesting that these peptides may decrease their hydrophobic length to help them to adapt to thin membranes. PMID- 15654758 TI - Apolipoprotein E is the major physiological activator of lecithin-cholesterol acyltransferase (LCAT) on apolipoprotein B lipoproteins. AB - Our previous studies have indicated that lecithin-cholesterol acyltransferase (LCAT) contributes significantly to the apoB lipoprotein cholesteryl ester (CE) pool. Cholesterol esterification rate (CER) in apoA-I(-)(/)(-) apoE(-)(/)(-) mouse plasma was <7% that of C57Bl/6 (B6) mouse plasma, even though apoA-I(-)(/)( ) apoE(-)(/)(-) plasma retained (1)/(3) the amount of B6 LCAT activity. This suggested that lack of LCAT enzyme did not explain the low CER in apoA-I(-)(/)(-) apoE(-)(/)(-) mice and indicated that apoE and apoA-I are the only major activators of LCAT in mouse plasma. Deleting apoE on low-density lipoprotein (LDL) reduced CER (1% free cholesterol (FC) esterified/h) compared to B6 (6% FC esterified/h) and apoA-I(-)(/)(-) (11% FC esterified/h) LDL. Similar sized LDL particles from all four genotypes were isolated by fast protein liquid chromatography (FPLC) after radiolabeling with [(3)H]-free cholesterol (FC). LDLs (1 microg FC) from each genotype were incubated with purified recombinant mouse LCAT; LDL particles from B6 and apoA-I(-)(/)(-) plasma were much better substrates for CE formation (5.7% and 6.3% CE formed/30 min, respectively) than those from apoE(-)(/)(-) and apoE(-)(/)(-) apoA-I(-)(/)(-) plasma (1.2% and 1.1% CE formed/30 min). Western blot analysis showed that the amount of apoA-I on apoE(-)(/)(-) LDLs was higher compared to B6 LDL. Adding apoE to incubations of apoA-I(-)(/)(-) apoE(-)(/)(-) very low density lipoprotein (VLDL) resulted in a 3 fold increase in LCAT CER, whereas addition of apoA-I resulted in a more modest 80% increase. We conclude that apoE is a more significant activator of LCAT than apoA-I on mouse apoB lipoproteins. PMID- 15654759 TI - Residual structure in the repeat domain of tau: echoes of microtubule binding and paired helical filament formation. AB - The microtubule-associated protein tau is found aggregated into paired helical filaments in the intraneuronal neurofibrillary tangle deposits of victims of Alzheimer's disease (AD) and other related dementias. Tau contains a repeat domain consisting of three or four 31-32-residue imperfect repeats that forms the core of tau filaments and is capable of self-assembling into filaments in vitro. We have used high-resolution NMR spectroscopy to characterize the structural properties of the three-repeat domain of tau at the level of individual residues. We find that three distinct regions of the polypeptide corresponding to previously mapped microtubule interaction sites exhibit a preference for helical conformations, suggesting that these sites adopt a helical structure when bound to microtubules. In addition, we directly observe a marked preference for extended or beta-strand-like conformations in a stretch of residues between two of the helical regions, which corresponds closely to a region previously implicated as an early site of beta-strand structure formation and intermolecular interactions leading to paired helical filament (PHF) formation. This observation supports the idea that this region of the protein plays a crucial role in the formation of tau aggregates. We further show that disulfide-bond-mediated dimer formation does not affect and is not responsible for the observed structural preferences of the protein. Our results provide the first high-resolution view of the structural properties of the protein tau, are consistent with an important role for beta structure in PHF formation, and may also help explain recent reports that tau filaments contain helical structure. PMID- 15654760 TI - Kinetic and thermodynamic studies of tet repressor-tetracycline interaction. AB - Stopped-flow measurements have been employed to study the kinetics of the conformational changes in TetR (B) induced by tetracycline binding with and without Mg(2+) ions. Result of stopped-flow fluorometry measurements at pH 8.0 indicate conformational changes in the helix-turn-helix motif in the N-terminal domain and in the C-terminal inducer binding domain. Binding of tetracycline (Tc) to TetR in the absence of Mg(2+) can be described by a simple kinetics process, which is limited to the first step association without any unimolecular conformational change step upon Tc binding. The rate constants for this process are equal to 2.0 x 10(5) M(-)(1) s(-)(1) and 2.1 s(-)(1) for the forward and backward reaction, respectively, and gave the binding constant K(a) = 0.96 x 10(5) M(-)(1). The kinetics of [Tc-Mg](+) binding to TetR can be described by reactions in which the first step describes the association characterized by the rate constants k(a) = 1.4 x 10(5) M(-)(1) s(-)(1) and k(d) = 2.2 x 10(-)(2) s( )(1) and binding constant K(a) = 6.3 x 10(6) M(-)(1). The first step of [Tc Mg](+) association is followed by at least three conformational change steps, which occur in the inducer binding site and then propagate to the surroundings of Trp75 and Trp43 residues. The rate constants for the forward, k(c), and backward, k(-)(c), reaction for each of these conformational steps have been determined. The thermodynamics of the binding of tetracycline with and without Mg(2+) to TetR was investigated by isothermal titration calorimetry (ITC) at pH 8.0 and 25 degrees C. The measurement shows that TetR dimer possesses two equivalent binding sites for tetracycline, characterized by binding constant K(a) = 9.0 x 10(6) M( )(1) and K(a) = 7.0 x 10(4) M(-)(1) for Tc with and without Mg(2+), respectively. The binding of the inducer to TetR, in the presence and absence of Mg(2+) ion, is an enthalpy-driven reaction characterized by DeltaH = -51 kJ mol(-)(1) and DeltaH = -33 kJ mol(-)(1), respectively. The entropy change, DeltaS, for the interaction in the presence of Mg(2+) is equal to -38.9 J K(-)(1) mol(-)(1), and for the tetracycline alone, it was estimated at -17.6 J K(-)(1) mol(-)(1). PMID- 15654761 TI - Interaction of the 20 kDa and 63 kDa fragments of anthrax protective antigen: kinetics and thermodynamics. AB - The action of anthrax toxin begins when the protective antigen (PA(83), 83 kDa) moiety binds to a mammalian cell-surface receptor and is cleaved by a furin family protease into two fragments: PA(20) (20 kDa) and PA(63) (63 kDa). After PA(20) dissociates, receptor-bound PA(63) spontaneously oligomerizes to form a heptameric species, which is able to bind the two enzymatic components of the toxin and transport them to the cytosol. Treatment of PA(83) with trypsin yielded PA(63) and a form of PA(20) lacking unstructured regions at the N- and C-termini. We labeled these fragments with dyes capable of fluorescence resonance energy transfer to quantify their association in solution. We kinetically determined that the equilibrium dissociation constant is 190 nM with a dissociation rate constant, k(off), of 3.3 x 10(-)(2) s(-)(1) (t(1/2) of 21 s). A two-step association process was observed using stopped-flow: a fast bimolecular step (k(on) = 1.4 x 10(5) M(-)(1) s(-)(1)) was followed by a slower unimolecular step (k = 3.5 x 10(-)(3) s(-)(1)) with an equilibrium isomerization constant, K(iso), of 2.1. The two-step mechanism most consistent with the data is one in which the dissociation of the PA(20).PA(63) complex is followed by an isomerization in the PA(63) moiety. Our results indicate that, following the cleavage of PA on the cell surface, PA(20) is largely dissociated within a minute. A slow isomerization step in PA(63) may then potentiate it for oligomerization and subsequent steps in toxin action. PMID- 15654762 TI - Effects of benzo[a]pyrene-deoxyguanosine lesions on DNA methylation catalyzed by EcoRII DNA methyltransferase and on DNA cleavage effected by EcoRII restriction endonuclease. AB - DNA methylation is an important cellular mechanism for controlling gene expression. Whereas the mutagenic properties of many DNA adducts, e.g., those arising from polycyclic aromatic hydrocarbons, have been widely studied, little is known about their influence on DNA methylation. We have constructed site specifically modified 18-mer oligodeoxynucleotide duplexes containing a pair of stereoisomeric adducts derived from a benzo[a]pyrene-derived diol epoxide [(+)- and (-)-r7,t8-dihydroxy-t9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene, or B[a]PDE] bound to the exocyclic amino group of guanine. The adducts, either (+)- or (-)-trans-anti-B[a]P-N(2)-dG (G*), positioned either at the 5'-side or the 3' side deoxyguanosine residue in the recognition sequence of EcoRII restriction modification enzymes (5'-...CCA/TGG...) were incorporated into 18-mer oligodeoxynucleotide duplexes. The effects of these lesions on complex formation and the catalytic activity of the EcoRII DNA methyltransferase (M.EcoRII) and EcoRII restriction endonuclease (R.EcoRII) were investigated. The M.EcoRII catalyzes the transfer of a methyl group to the C5 position of the 3'-side cytosine of each strand of the recognition sequence, whereas R.EcoRII catalyzes cleavage of both strands. The binding of R.EcoRII to the oligodeoxynucleotide duplexes and the catalytic cleavage were completely abolished when G was positioned at the 3'-side dG position (5'-...CCTGG*...). When G* was at the 5' side dG position, binding was moderately diminished, but cleavage was completely blocked. In the case of M.EcoRII, binding is diminished by factors of 5-30 but the catalytic activity was either abolished or reduced 4-80-fold when the adducts were located at either position. Somewhat smaller effects were observed with hemimethylated oligodeoxynucleotide duplexes. These findings suggest that epigenetic effects, in addition to genotoxic effects, need to be considered in chemical carcinogenesis initiated by B[a]PDE, since the inhibition of methylation may allow the expression of genes that promote tumor development. PMID- 15654763 TI - Molecular recognition in a trans excision-splicing ribozyme: non-Watson-Crick base pairs at the 5' splice site and omegaG at the 3' splice site can play a role in determining the binding register of reaction substrates. AB - Trans excision-splicing (TES) ribozymes, derived from a Pneumocystis carinii group I intron, can catalyze the excision of targeted sequences from within RNAs. In this report, the sequence requirements of the splice sites are analyzed. These conserved sequences include a u-G wobble pair at the 5' splice site and a guanosine in the omega position at the 3' splice site (in the substrate). We report that 7 out of 16 base pair combinations at the 5' splice site produce appreciable TES product. This promiscuity is in contrast to results reported for analogous self-splicing reactions using a Tetrahymena ribozyme. At long reaction times TES products dissociate and rebind free ribozyme, at which point product degradation occurs via the 5' cleavage reaction. Unexpectedly, only in cases where Watson-Crick base pairs form at the 5'splice site do we see degradation of TES products at cryptic sites, suggesting that non-Watson-Crick base pairs at the 5' splice site are acting in concert with other factors to precisely determine the binding register of TES reaction substrates within the ribozyme. Moreover, cryptic site degradation does not occur with the corresponding reaction substrates, which additionally contain omegaG, suggesting that omegaG can play a similar role. We report that omegaG cannot be replaced by any other base, so TES substrates require a guanosine as the last (or only) base to be excised. Additionally, we demonstrate that P9.0 and P10 are expendable for TES reactions, suggesting that omegaG is sufficient as a 3' molecular recognition element. PMID- 15654764 TI - Direct evidence that the reaction intermediate of metallo-beta-lactamase L1 is metal bound. AB - In an effort to probe the structure of the reaction intermediate of metallo-beta lactamase L1 when reacted with nitrocefin and other beta-lactams, time-dependent absorption and rapid-freeze-quench (RFQ) EPR spectra were obtained using the Co(II)-substituted form of the enzyme. When using nitrocefin as the substrate, time-dependent absorption spectra demonstrate that Co(II)-substituted L1 utilizes a reaction mechanism, similar to that of the native Zn(II) enzyme, in which a short-lived intermediate forms. RFQ-EPR spectra of this intermediate demonstrate that the binding of substrate results in a change in the electronic properties of one or both of the Co(II)'s in the enzyme that is consistent with a change in the coordination sphere of this metal ion. This observation provides evidence that the reaction intermediate is a metal-bound species. RFQ-EPR studies also demonstrate that other beta-lactams, such as cephalothin, meropenem, and penicillin G, proceed through an electronically similar complex and that the role of metal is similar in all cases. EPR spectroscopy has also identified distinct product-bound species of L1, indicating that reversible product binding must be considered in all future kinetic mechanisms. Consideration of the time-dependent optical and EPR studies in light of available crystallographic information indicates the intimate involvement of the metal ion in the Zn(2)-binding site of L1 in the hydrolytic reaction. PMID- 15654766 TI - Extracellular RNA is a natural cofactor for the (auto-)activation of Factor VII activating protease (FSAP). AB - FSAP (Factor VII-activating protease) is a new plasma-derived serine protease with putative dual functions in haemostasis, including activation of coagulation Factor VII and generation of urinary-type plasminogen activator (urokinase). The (auto-)activation of FSAP is facilitated by polyanionic glycosaminoglycans, such as heparin or dextran sulphate, whereas calcium ions stabilize the active form of FSAP. In the present study, extracellular RNA was identified and characterized as a novel FSAP cofactor. The conditioned medium derived from various cell types such as smooth muscle cells, endothelial cells, osteosarcoma cells or CHO (Chinese-hamster ovary) cells contained an acidic factor that initiated (auto )activation of FSAP. RNase A, but not other hydrolytic enzymes (proteases, glycanases and DNase), abolished the FSAP cofactor activity, which was subsequently isolated by anion-exchange chromatography and unequivocally identified as RNA. In purified systems, as well as in plasma, different forms of natural RNA (rRNA, tRNA, viral RNA and artificial RNA) were able to (auto )activate FSAP into the two-chain enzyme form. The specific binding of FSAP to RNA (but not to DNA) was shown by mobility-shift assays and UV crosslinking, thereby identifying FSAP as a new extracellular RNA-binding protein, the K(D) estimated to be 170-350 nM. Activation of FSAP occurred through an RNA-dependent template mechanism involving a nucleic acid size of at least 100 nt. In a purified system, natural RNA augmented the FSAP-dependent Factor VII activation several-fold (as shown by subsequent Factor Xa generation), as well as the FSAP mediated generation of urokinase. Our results provide evidence for the first time that extracellular RNA, present at sites of cell damage or vascular injury, can serve an important as yet unrecognized cofactor function in haemostasis by inducing (auto-)activation of FSAP through a novel surface-dependent mechanism. PMID- 15654767 TI - Glucosamine induces cell-cycle arrest and hypertrophy of mesangial cells: implication of gangliosides. AB - Alterations in proliferation and hypertrophy of renal mesangial cells are typical features of diabetic nephropathy. The HP (hexosamine pathway) has been proposed as a biochemical hypothesis to explain microvascular alterations due to diabetic nephropathy; however, involvement of HP in the regulation of mesangial cell growth or hypertrophy has been poorly studied. Although gangliosides are known to regulate cell proliferation, their potential role in mesangial cell-growth perturbations has hardly been explored. In the present study, we investigated the effects of the HP activation, mimicked by GlcN (glucosamine) treatment, on mesangial cell growth and hypertrophy and the potential implication of gangliosides in these processes. Our results indicate that GlcN induced hypertrophy of mesangial cells, as measured by an increase in the protein/cell ratio, and it caused cell-cycle arrest by an increase in the expression of cyclin dependent kinase inhibitor p21(Waf1/Cip1). Furthermore, GlcN treatment resulted in a massive increase in the levels of gangliosides G(M2) and G(M1). Treatment of cells with exogenous G(M2) and G(M1) reproduced the effects of 0.5 mM GlcN on p21(Waf1/Cip1) expression, cell-cycle arrest and hypertrophy, suggesting that gangliosides G(M2) and G(M1) are probably involved in mediating GlcN effects. These results document a new role of the HP in the regulation of mesangial cell growth and hypertrophy. They also suggest a potential new mechanism of action of the HP through modulation of ganglioside levels. PMID- 15654769 TI - The trans-Golgi network GRIP-domain proteins form alpha-helical homodimers. AB - A recently described family of TGN (trans-Golgi network) proteins, all of which contain a GRIP domain targeting sequence, has been proposed to play a role in membrane transport. On the basis of the high content of heptad repeats, GRIP domain proteins are predicted to contain extensive coiled-coil regions that have the potential to mediate protein-protein interactions. Four mammalian GRIP domain proteins have been identified which are targeted to the TGN through their GRIP domains, namely p230, golgin-97, GCC88 and GCC185. In the present study, we have investigated the ability of the four mammalian GRIP domain proteins to interact. Using a combination of immunoprecipitation experiments of epitope-tagged GRIP domain proteins, cross-linking experiments and yeast two-hybrid interactions, we have established that the GRIP proteins can self-associate to form homodimers exclusively. Two-hybrid analysis indicated that the N- and C-terminal fragments of GCC88 can interact with themselves but not with each other, suggesting that the GRIP domain proteins form parallel coiled-coil dimers. Analysis of purified recombinant golgin-97 by CD spectroscopy indicated a 67% alpha-helical structure, consistent with a high content of coiled-coil sequences. These results support a model for GRIP domain proteins as extended rod-like homodimeric molecules. The formation of homodimers, but not heterodimers, indicates that each of the four mammalian TGN golgins has the potential to function independently. PMID- 15654768 TI - Pi class glutathione S-transferase genes are regulated by Nrf 2 through an evolutionarily conserved regulatory element in zebrafish. AB - Pi class GSTs (glutathione S-transferases) are a member of the vertebrate GST family of proteins that catalyse the conjugation of GSH to electrophilic compounds. The expression of Pi class GST genes can be induced by exposure to electrophiles. We demonstrated previously that the transcription factor Nrf 2 (NF E2 p45-related factor 2) mediates this induction, not only in mammals, but also in fish. In the present study, we have isolated the genomic region of zebrafish containing the genes gstp1 and gstp2. The regulatory regions of zebrafish gstp1 and gstp2 have been examined by GFP (green fluorescent protein)-reporter gene analyses using microinjection into zebrafish embryos. Deletion and point-mutation analyses of the gstp1 promoter showed that an ARE (antioxidant-responsive element)-like sequence is located 50 bp upstream of the transcription initiation site which is essential for Nrf 2 transactivation. Using EMSA (electrophoretic mobility-shift assay) analysis we showed that zebrafish Nrf 2-MafK heterodimer specifically bound to this sequence. All the vertebrate Pi class GST genes harbour a similar ARE-like sequence in their promoter regions. We propose that this sequence is a conserved target site for Nrf 2 in the Pi class GST genes. PMID- 15654771 TI - Evidence-based medicine in practice: applying intention-to-treat analysis and perprotocol analysis. PMID- 15654770 TI - Arginine methylation regulates IL-2 gene expression: a role for protein arginine methyltransferase 5 (PRMT5). AB - Arginine methylation is a post-translational modification resulting in the generation of aDMAs (asymmetrical omega-NG, NG-dimethylated arginines) and sDMAs (symmetrical omega-NG, N'G-dimethylated arginines). The role of arginine methylation in cell signalling and gene expression in T lymphocytes is not understood. In the present study, we report a role for protein arginine methylation in regulating IL-2 (interleukin 2) gene expression in T lymphocytes. Leukaemic Jurkat T-cells treated with a known methylase inhibitor, 5' methylthioadenosine, had decreased cytokine gene expression, as measured using an NF-AT (nuclear factor of activated T-cells)-responsive promoter linked to the luciferase reporter gene. Since methylase inhibitors block all methylation events, we performed RNA interference with small interfering RNAs against the major PRMT (protein arginine methyltransferases) that generates sDMA (PRMT5). The dose-dependent decrease in PRMT5 expression resulted in the inhibition of both IL 2- and NF-AT-driven promoter activities and IL-2 secretion. By using an sDMA specific antibody, we observed that sDMA-containing proteins are directly associated with the IL-2 promoter after T-cell activation. Since changes in protein arginine methylation were not observed after T-cell activation in Jurkat and human peripheral blood lymphocytes, our results demonstrate that it is the recruitment of methylarginine-specific protein(s) to cytokine promoter regions that regulates their gene expression. PMID- 15654772 TI - Treatment of hepatitis C in injection drug users. PMID- 15654773 TI - Bravo capsule pH monitoring. PMID- 15654774 TI - Assessing symptoms in gastroesophageal reflux disease: how well do clinicians' assessments agree with those of their patients? AB - OBJECTIVES: This study aimed to determine the extent of agreement between clinicians and patients regarding assessments of reflux symptom severity in patients with gastroesophageal reflux disease. METHODS: Data were analyzed from four randomized clinical trials involving 2,674 patients treated with esomeprazole, omeprazole, ranitidine, or placebo. The extent of agreement was determined for symptom severity before and after 4-8 wk of treatment, and for the absence of symptoms after treatment. Agreement was further analyzed by determining weighted kappa values, which were interpreted according to the criteria of Landis and Koch. RESULTS: Before treatment, clinician-patient agreement regarding symptom severity in the four studies was slight to moderate (kappa: 0.17-0.53); 48-52% of assessments agreed for heartburn, 24-35% for epigastric pain, 36-43% for regurgitation, and 63% agreed for dysphagia. Poor agreement reflected clinician underestimation of symptom severity relative to patient ratings in three studies and clinician overestimation in one study. Agreement regarding symptom severity improved following treatment, and was fair to substantial (kappa: 0.31-0.73); 58-78% of assessments agreed for heartburn, 42 60% for epigastric pain, 66-76% for regurgitation, and 86% agreed for dysphagia. After treatment, agreement was greatest for patients reporting absence of symptoms and decreased with increasing severity of symptoms. CONCLUSIONS: The agreement between clinicians and patients in their assessments of the severity of reflux symptoms is poor, particularly before treatment and for more severe symptoms. Improvements in clinician-patient communication may help to bridge this gap, and greater reliance on patient assessments may be appropriate. PMID- 15654775 TI - Measuring gastroesophageal reflux symptoms: musings from Marrakech. AB - The optimum approach for evaluation of reflux symptoms is uncertain. Trials have used a variety of methods, and it is important that a more consistent approach is taken. One piece of the puzzle is whether symptoms should be assessed by a clinician or directly by a patient completed questionnaire. McColl et al. suggest that clinicians tend to underestimate the severity of symptoms and that a more patient-centered approach should be used to elicit symptoms. This has important implications for the development of future questionnaires in gastroesophageal reflux and other gastrointestinal diseases. PMID- 15654776 TI - Crystal violet chromoendoscopy with mucosal pit pattern diagnosis is useful for surveillance of short-segment Barrett's esophagus. AB - BACKGROUND: Because of a rapid increase in the incidence of Barrett's cancer, the appropriate surveillance method for Barrett's esophagus is of interest. Methylene blue chromoendoscopy has been reported to be an effective and inexpensive method to improve biopsy surveillance of Barrett's epithelium. However, the usefulness of this method in short-segment Barrett's esophagus cases is still controversial. AIMS: This study was undertaken to evaluate the abilities of crystal violet and methylene blue chromoendoscopy to detect potentially dysplastic Barrett's epithelium in cases with short-segment columnar-appearing epithelium of the esophago-gastric junction. PATIENTS AND METHODS: Four hundred patients with endoscopically suspected short-segment Barrett's esophagus were enrolled and randomly assigned to receive chromoendoscopy with 0.05% crystal violet, 0.1% crystal violet, 0.5% methylene blue, or 1.0% methylene blue. During crystal violet and methylene blue chromoendoscopy, biopsy specimens were obtained from stained and unstained columnar-appearing epithelium of the esophago-gastric junction, and the detection rates of Barrett's epithelium were evaluated. The value of pit pattern diagnosis was also evaluated as a possible way to detect dysplastic Barrett's epithelium. RESULTS: Chromoendoscopy with 0.05% crystal violet detected histologically confirmed Barrett's epithelium with the highest sensitivity (89.2%) and specificity (85.7%). Crystal violet clearly stained both dysplastic and nondysplastic Barrett's epithelia and made the surface pit pattern easy to observe without using magnifying endoscopy. CONCLUSIONS: The combination of crystal violet chromoendoscopy and pit pattern diagnosis is considered to be useful for the surveillance of short-segment Barrett's esophagus. PMID- 15654777 TI - Feasibility and diagnostic utility of video capsule endoscopy for the detection of small bowel polyps in patients with hereditary polyposis syndromes. AB - OBJECTIVES: At present, surveillance of premalignant small bowel polyps in hereditary polyposis syndromes has a number of limitations. Capsule endoscopy (CE) is a promising new method to endoscopically assess the entire length of the small bowel. METHODS: We prospectively examined 40 patients with hereditary polyposis syndromes (29 familial adenomatous polyposis (FAP), 11 Peutz-Jeghers syndrome (PJS)). Results were compared with push-enteroscopy (PE) results in FAP and with esophagogastroduodenoscopy, PE, (MR)-enteroclysis, and surgical specimen in PJS patients. RESULTS: A total of 76% of the patients with FAP with duodenal adenomas (n = 21) had additional adenomas in the proximal jejunum that could be detected by CE and PE. Moreover, 24% of these FAP patients had further polyps in the distal jejunum or ileum that could only be detected by CE. In contrast, in FAP patients without duodenal polyps (n = 8), jejunal or ileal polyps occurred rarely (12%). CE detected polyps in 10 of 11 patients with PJS, a rate superior to all other reference procedures employed. Importantly, the findings of CE had immediate impact on further clinical management in all PJS patients. CONCLUSIONS: Our results suggest that CE may be of clinical value in selected patients with FAP, whereas in PJS, CE could be used as first line surveillance procedure. PMID- 15654778 TI - Risk factors for pancreatitis in patients with common bile duct stones managed by endoscopic papillary balloon dilation. AB - OBJECTIVES: Endoscopic papillary balloon dilation has been accepted as a possible alternative to endoscopic sphincterotomy, especially in patients with impaired hemostasis. However, pancreatitis associated with endoscopic papillary balloon dilation has remained a controversial, serious issue. The aim of the study was to investigate the risk factors for postendoscopic papillary balloon dilation pancreatitis in a single-center study. METHODS: A total of 304 patients who underwent endoscopic papillary balloon dilation for the management of common bile duct stones were enrolled. The risk of postendoscopic papillary balloon dilation pancreatitis was evaluated and the risk factors were analyzed by univariate and multivariate analysis. Definition and grade of the severity of postendoscopic papillary balloon dilation pancreatitis were based on the 1991 consensus guidelines. RESULTS: Common bile duct was cleared in 292 of 304 patients (96%). Procedure-related pancreatitis occurred in 15 patients (5.0%). The grade was mild in 8 and moderate in 7. Two risk factors, stone diameter and contrast medium injection to the pancreas, were identified by univariate analysis. Finally, only the contrast medium injection to the pancreas was statistically significant by multivariate analysis. Prior history of pancreatitis was identified as a risk factor for postendoscopic papillary balloon dilation pancreatitis in previous studies, but this factor was not identified as a risk factor in our series. CONCLUSIONS: Although the pathogenesis of pancreatitis after endoscopic papillary balloon dilation remains unresolved, the unnecessary injection of contrast medium to the pancreas can certainly be considered to be associated with the increased risk of pancreatitis. PMID- 15654779 TI - Long-term results of partial splenic artery embolization as supplemental treatment for portal-systemic encephalopathy. AB - OBJECTIVES: To present long-term results of angiographic partial splenic artery embolization (PSE) as a supplemental treatment of portal-systemic encephalopathy. METHODS: Twenty-five patients with portal-systemic encephalopathy were divided into two groups: 14 patients underwent transportal obliteration and/or balloon occluded retrograde transvenous obliteration (BRTO) of portal-systemic shunts (PSS), followed by PSE (PSE(+) group), and 11 patients underwent only transportal obliteration and/or BRTO of PSS (PSE(-) group). RESULTS: Portal venous pressures pretreatment was similar to posttreatment in the PSE(+) group, but lower than posttreatment in the PSE(-) group. Serum ammonia levels were higher at pretreatment than at 1 wk posttreatment in both groups, but the levels in the two groups were similar at pretreatment, 1 wk, 3 months, 3 yr, 4 yr, and 5 yr posttreatment. However, serum ammonia levels were lower in the PSE(+) group than in the PSE(-) group 6 months, 9 months, 1 yr, and 2 yr posttreatment. Grades of encephalopathy were higher at pretreatment than at 1 wk posttreatment in both groups, but the levels in the two groups were similar at pretreatment, 1 wk, 2 yr, 3 yr, 4 yr, and 5 yr posttreatment. However, grades of encephalopathy were lower in the PSE(+) group than in the PSE(-) group 3 months, 6 months, 9 months, and 1 yr posttreatment. CONCLUSIONS: Obliteration of PSS followed by PSE benefit patients with portal-systemic encephalopathy. PMID- 15654780 TI - Chronic hepatitis C and type II diabetes mellitus: a prospective cross-sectional study. AB - An epidemiologic link between chronic hepatitis C (HCV) and type II diabetes mellitus (DM) has been established. Our aims were to prospectively determine the prevalence of DM in interferon-naive patients with HCV in comparison with the general population, and to determine the association between DM and impaired fasting glucose (IFG) with histological stage in patients with HCV. A consecutive sample of 179 patients was included in this prospective cross-sectional study. The crude percentage of DM for the cohort was 14.5%, different from the crude rate of 7.8% for the general population (p= 0.0008) and from the rate of 7.3% observed in a matched control group with non-HCV liver disease. The prevalence of DM and IFG (DM/IFG) was higher among HCV-infected patients with advanced versus those with early histological disease (p= 0.0004). Advanced histological disease predicted DM/IFG after controlling for other identified risk factors for DM. Family history was the only other independent predictor of DM/IFG in HCV-infected patients. In conclusion, patients with HCV had a higher prevalence of DM compared to the general population. The presence of advanced histological disease in genetically predisposed HCV-patients is associated with a higher prevalence of DM/IFG. DM and IFG were not associated with anthropomorphic markers of obesity in HCV patients, suggesting a unique multifactorial pathogenesis of DM in HCV. PMID- 15654781 TI - The effect of HIV coinfection on the risk of cirrhosis and hepatocellular carcinoma in U.S. veterans with hepatitis C. AB - OBJECTIVES: This study was conducted to determine whether HIV coinfection increases the risk of cirrhosis in HCV-infected patients in the HAART and pre HAART eras. Further, the risk of hepatocellular carcinoma was also examined. METHODS: This retrospective cohort study was conducted among HCV-infected veterans who were seen at one of the 172 Veterans Health Administration hospitals between October 1, 1991 and September 30, 2000. Patients with prerecorded advanced liver disease were excluded. Incidence rates, cumulative incidence, and Cox proportional hazard ratios were calculated. RESULTS: There were 26,641 patients with HCV-only and 4,761 patients with HCV-HIV coinfection. The unadjusted incidence rate of cirrhosis was lower in patients with coinfection than HCV-only (p < 0.01). After controlling for demographics and confounders (including alcoholism and chronic hepatitis B), coinfection was not significantly associated with cirrhosis. However, there was an increased risk of cirrhosis in patients with coinfection compared to HCV-only during the pre-HAART era (before October 1, 1996) (hazard ratio = 1.48, 1.06-2.07, p= 0.02), but not among patients who entered the cohort during the HAART era. The unadjusted incidence rate of hepatocellular carcinoma in patients with coinfection and HCV-only was 1.3 and 2/1,000 person-years, respectively (p= 0.04). In the multivariate model, coinfection was not associated with hepatocellular carcinoma (hazard ratio = 0.84, p= 0.40). CONCLUSIONS: Coinfection was a significant risk factor for cirrhosis only during the pre-HAART era and was not associated with hepatocellular carcinoma, irrespective of time period. PMID- 15654782 TI - Activation pattern of signal transducers and activators of transcription (STAT) factors in inflammatory bowel diseases. AB - OBJECTIVES: Cytokine signaling pathways involving transcription factors of the signal transducers and activators of transcription (STAT) family play a key role in the pathogenesis of inflammatory bowel diseases (IBD). STAT proteins are latent cytoplasmic transcription factors that induce transcription upon phosphorylation, dimerization, and nuclear translocation. However, their activation pattern in IBD is poorly understood. The aim of our study was to characterize STAT-expression in IBD. METHODS: Mononuclear cells were isolated from 36 colonic specimens of Crohn's disease, ulcerative colitis, or from control patients. Cells were stimulated overnight with antibodies against human CD2 and CD28 and mononuclear cells were analyzed by flow cytometry. Alternatively, CD4(+) T cells were immunomagnetically separated and then assessed by flow cytometry. Intracellular stainings of the following transcription factors were performed: STAT-1, STAT-2, STAT-3, STAT-4, and STAT-6. In addition, immunofluorescence staining on cryosections for phosphorylated STAT-1 and STAT-3 was performed. RESULTS: Average expression of the IFN-gamma inducible transcription factor STAT 1 was increased in Crohn's disease as compared to patients with ulcerative colitis and control patients. However, levels of phospho-STAT-1 were surprisingly not markedly upregulated in IBD as compared to controls. In contrast, STAT-3 and phospho-STAT-3 levels were significantly increased in IBD patients as compared to controls (p < 0.01). No differences could be detected in STAT-6 levels. Finally, average expression of STAT-2, which is involved in type I interferon signalling, was downregulated in IBD as compared to control patients. CONCLUSIONS: The analysis of STAT activation patterns could serve as a helpful tool to characterize intestinal inflammation. Furthermore, the IL-6/STAT-3 rather than the IFN-gamma/STAT-1 signaling pathway emerges as a key target for the development of future therapeutic concepts in IBD. PMID- 15654783 TI - A STAT need for human immunologic studies to understand inflammatory bowel disease. AB - In this issue of The American Journal of Gastroenterology, Mudter et al. present a study elucidating a potential role for STAT proteins in the pathogenesis of the human inflammatory bowel diseases (IBDs). The importance of this study lies in the methodology, as molecular biology was applied to humans to describe expression and activation of STATs in intestinal mucosal cells from IBD patients. The speculative roles for the family members/STAT1 and STAT3 suggest a divergence between human IBD and immunologic hypotheses derived from murine models. This work provides an example of why technology and experiments to understand human immunology are ultimately our best strategy to develop therapies and biomarkers for IBD. PMID- 15654784 TI - Safety and efficacy of adalimumab (D2E7) in Crohn's disease patients with an attenuated response to infliximab. AB - OBJECTIVES: Although infliximab is highly effective in the treatment of Crohn's disease (CD), attenuated response to infliximab may develop over time in a subgroup of patients. The aim of our study was to examine the safety and efficacy of adalimumab (D2E7), a fully humanized anti-TNF-alpha Ab, in CD patients who had experienced an attenuated response to infliximab. METHODS: Fifteen patients with active CD who experienced an attenuated response to infliximab were treated with adalimumab over a 6-month period. Patients, received a loading dose of 80 mg subcutaneously followed by 40 mg every 2 wk. The clinical response to adalimumab was classified as complete response, partial response, or nonresponse. RESULTS: Two patients received the loading dose of adalimumab but did not have adequate follow-up evaluations. Of the remaining 13 patients, 7 (54%) had a complete response, 4 (31%) had a partial response, and 2 (15%) were nonresponders. In six patients, the maintenance dose was increased in order to maintain clinical response. Eight of 11 (73%) patients on concurrent corticosteroids were able to discontinue or significantly decrease the dose of the steroids. Adalimumab was well tolerated without signs or symptoms of allergic reaction except in two patients who developed an injection site reaction. CONCLUSIONS: Our preliminary data suggest that adalimumab may be a safe and effective therapy for patients with CD who have experienced an attenuated response to infliximab. PMID- 15654785 TI - What next after infliximab? AB - The use of infliximab (Remicade) has revolutionized the care of Crohn's disease (CD) patients who have proved refractory to standard treatment. The use of infliximab is very well tolerated in the majority of patients but in a small subset of patients may lead to the production of antibodies (termed "antibodies to infliximab"-ATI). The production of these antibodies has been associated with the development of both acute and delayed infusion reactions, although even in patients who develop ATIs, these reactions are relatively uncommon. Nonetheless, these reactions may occasionally be severe enough to lead to intolerance to infliximab. Another group of patients, after initially having excellent responses to infliximab, experience an attenuated response or loss of response over time. What is the cause of this loss of efficacy? ATIs may play a role in some patients but other potential reasons for this phenomenon have provoked much debate. The importance of other cytokines after TNF-alpha has been neutralized may be relevant as (this has been shown to be the case in rheumatoid arthritis (RA) is the idea of beneficial autoimmunity production to TNF-alpha. (Wildbaum G, Nahir MA, Karin N. Beneficial autoimmunity to proinflammatory mediators restrains the consequences of self-destructive immunity. Immunity 2003;19:679-88.) It has been shown that during the course of an autoimmune condition, the immune system mounts a beneficial autoantibody response to proinflammatory mediators. This response counteracts, to a certain degree, the autoimmune pathology. This natural counteraction has been illustrated in animal models of autoimmunity, and there has been evidence demonstrated that this occurs in human RA. Whether this occurs in Crohn's is unknown; infliximab is a chimeric monoclonal antibody containing an approximately 25% murine region. It had been hoped that the development of humanized or fully human monoclonal antibodies would provide therapeutic antibodies that did not induce an immune response. While this has unfortunately not proven to be the case-these products still have significant immunogenicity these products do present an alternative therapy when infliximab cannot be used. In light of this, adalimumab (Humira) a human monoclonal antibody used for treating rheumatologic conditions has been investigated as an alternate treatment for patients with CD who after initially responding to infliximab experience intolerance or loss of efficacy. Is this a viable alternative? PMID- 15654786 TI - Variants of CARD15 are associated with an aggressive clinical course of Crohn's disease--an IG-IBD study. AB - BACKGROUND: Three major variants of the CARD15 gene confer susceptibility to Crohn's disease (CD). Whether or not these variants correlate with specific clinical features of the disease is under evaluation. AIM: We investigated the possible association of CARD15 variants with specific clinical characteristics, including the occurrence of anti-Saccharomyces cerevisiae antibodies (ASCA) and antineutrophil cytoplasmic antibodies (ANCA), in a large cohort of inflammatory bowel disease (IBD) patients and their unaffected relatives. METHODS: Three hundred and sixteen CD patients (156 with positive family history), 408 ulcerative colitis (UC) patients (206 with positive family history), 588 unaffected relatives, and 205 unrelated healthy controls (HC) were studied. Single nucleotide polymorphisms (SNPs) R702W, G908R, and L1007finsC of the CARD15 gene were investigated and correlated to age at diagnosis, gender, family history, localization, extraintestinal manifestations, previous resective surgery, stenosing/fistulizing pattern, ANCA, and ASCA. RESULTS: Compared to HC, the frequencies of all three variants in CD were significantly increased: 8.7% versus 4.1% for R702W (p < 0.006), 7.3% versus 2.7% for G908R (p < 0.002), 9.3% versus 0.7% for L1007finsC (p < 0.00001). At least one risk allele was found in 38.2% (p < 0.0001, compared to HC), 13.7% (NS), and 15.1% of CD, UC, and HC, respectively. The L1007finsC risk allele was also significantly increased in unaffected relatives of familial (9.5%; p < 0.00001), and sporadic CD (9%; p < 0.00001), compared to HC (0.7%). Sixteen healthy relatives, carriers of two risk alleles, were asymptomatic after 5-8 yr of follow-up. CD carriers of at least one variant were younger (p= 0.03), more likely to have ileal localization (p= 0.0001), stenosing pattern (p= 0.01), previous resective surgery (p= 0.0001), and presence of ASCA (p= 0.0001). No difference in SNPs frequency between familial and sporadic cases of CD was found. CONCLUSION: In our population, both familial and sporadic CD patients carrying at least one major variant of CARD15 had an aggressive clinical course. PMID- 15654787 TI - Comprehensive evaluation of inflammatory and noninflammatory sequelae of ileal pouch-anal anastomoses. AB - BACKGROUND AND AIMS: Ileal pouch-anal anastomosis (IPAA) improves quality of life (QOL) for ulcerative colitis patients who require surgery. Crohn's disease (CD) of the pouch, pouchitis, cuffitis, and irritable pouch syndrome (IPS) have an adverse impact on physical and psychological well-being, which can compromise the gain in QOL after the surgery. Their clinical, endoscopic, and histologic features have not been fully characterized. The aim of this study was to compare demographic, clinical, endoscopic, and histologic features between CD of the pouch, pouchitis, cuffitis, IPS, and normal pouches. METHODS We enrolled 124 patients: normal pouches (N = 26), CD of the pouch (N = 23), pouchitis (N = 22), cuffitis (N = 21), and IPS (N = 32). Symptomatology, endoscopy, histology, and the Cleveland Global QOL and the Irritable Bowel Syndrome-QOL scores were compared among the groups. RESULTS: Univariate analysis of demographic and clinical data showed a possible association between NSAID use and pouchitis, extraintestinal manifestation and cuffitis, and antidepressant use and IPS. There were no differences in the Pouchitis Disease Activity Index symptom scores between the disease groups, with an exception of bleeding, which occurred almost exclusively in cuffitis. Endoscopy was useful in discriminating between CD of the pouch, pouchitis, cuffitis, and normal pouches or IPS. Patients with diseased IPAA had worse QOL scores. CONCLUSIONS: Symptoms largely overlapped among the disease groups of IPAA. Endoscopy is valuable for diagnosis. Inflammatory or noninflammatory sequelae of IPAA adversely affected patients' QOL. PMID- 15654788 TI - The role of breastfeeding in postpartum disease activity in women with inflammatory bowel disease. AB - BACKGROUND: Women with inflammatory bowel disease (IBD) have an increased risk of experiencing a flare in the postpartum period. Work in other autoimmune disorders has found that breastfeeding may be associated with an increased risk for developing postpartum disease relapse. AIM: To assess the association between breastfeeding behavior and postpartum disease activity. METHODS: Women with IBD followed at a tertiary care center with a history of childbirth within the past 5 yr were recruited. Medical records were reviewed for disease type, disease activity during and after pregnancy, medication use, smoking, and breastfeeding behavior. The exposure of interest was breastfeeding prior to the onset of disease activity following a successful asymptomatic pregnancy. RESULTS: One hundred and twenty-two consecutive women who fit eligibility criteria were studied. Overall, only 44% (54/122) of the women had breastfed their infant. Reasons included physician recommendation, fear of medication interactions, and personal choice. Forty-three percent (23/54) of those who breastfed experienced a postpartum flare of their disease. The unadjusted odds ratio for disease activity with a history of breastfeeding was 2.2 (95% CI 1.2-3.9, p= 0.004). When stratified by disease type, the OR for ulcerative colitis was 0.89 (0.29-2.7, p > 0.05) and Crohn's disease 3.8 (1.9-7.4, p < 0.05). When adjusted for medication cessation, the OR became nonsignificant. CONCLUSIONS: A significant number of women with IBD do not breastfeed their children. Any relationship between breastfeeding and disease activity may be more a consequence of discontinuation of IBD therapies. PMID- 15654789 TI - Rectal hyposensitivity: a disorder of the rectal wall or the afferent pathway? An assessment using the barostat. AB - OBJECTIVES: Rectal hyposensitivity (RH) relates to a diminished perception of rectal distension. Diagnosis on the basis of abnormal threshold volumes on balloon distension alone may be inaccurate due to the influence of differing rectal wall properties. The aim of this study was to investigate whether RH was actually due to impaired afferent nerve function or whether it could be secondary to abnormalities of the rectal wall. METHODS: A total of 50 patients were referred consecutively to a tertiary referral unit for physiologic assessment of constipation (Rome II criteria), 25 of whom had associated fecal incontinence. Thirty patients had RH (elevated threshold volumes on latex balloon distension), and 20 patients had normal rectal sensation (NS). Results were compared with those obtained in 20 healthy volunteers (HV). All subjects underwent standard anorectal physiologic investigation, and assessment of rectal compliance, adaptive response to isobaric distension at urge threshold, and postprandial rectal response, using an electromechanical barostat. RESULTS: Mean rectal compliance was significantly elevated in patients with RH compared to NS and HV (p < 0.001). However, 16 patients with RH (53%) had normal compliance. Intensity of the urge to defecate during random phasic isobaric distensions was significantly reduced in patients with RH compared to NS and HV (p < 0.001). The adaptive response at urge threshold was reduced in patients with RH compared to NS and HV (p < 0.001), although spontaneous adaptation at operating pressure was similar in all three groups studied (p= 0.3). Postprandially, responses were similar between groups. CONCLUSIONS: In patients found to have RH on simple balloon distension, impaired perception of rectal distension may be partly explained in one subgroup by abnormal rectal compliance. However, a second subgroup exists with normal rectal wall properties, suggestive of a true impairment of the afferent pathway. The barostat has an important role in the identification of these subgroups of patients. PMID- 15654790 TI - A dose-ranging, phase II study of the efficacy and safety of alosetron in men with diarrhea-predominant IBS. AB - BACKGROUND: A randomized, double blind, placebo-controlled dose-ranging study was conducted to assess the efficacy of alosetron in men with diarrhea-predominant irritable bowel syndrome (IBS). METHODS: Six hundred and sixty-two men were randomized to treatment with alosetron 0.5, 1.0, 2.0, 4.0 mg, or placebo twice daily for 12 wk, followed by a 4-wk posttreatment period. Adequate relief of IBS pain and discomfort during week 5-12 of the treatment phase was the primary endpoint; secondary endpoints included bowel urgency, stool frequency, and consistency, incomplete evacuation, bloating, and abdominal pain or discomfort. RESULTS: Subjects ranked urgency and abdominal pain as their most bothersome IBS symptoms. The average rate of adequate relief during week 5-12 was significantly higher in the alosetron 1.0 mg twice-daily group compared to placebo (53%vs 40%, p= 0.04), and all doses of alosetron significantly reduced stool consistency scores (p < 0.001) indicating firmer stools. No significant effects of alosetron were seen with regard to urgency, number of bowel movements, bloating, and incomplete evacuation. Constipation was the most common adverse event and occurred in a dose-related manner among subjects receiving alosetron, 9% (0.5 mg twice daily), 15% (1.0 mg twice daily), 11% (2.0 mg twice daily), and 21% (4.0 mg twice daily). No serious adverse events of constipation were reported. One subject in the 0.5 mg twice-daily group had an episode of rectal bleeding suggestive of a possible diagnosis of ischemic colitis. CONCLUSIONS: Alosetron 1 mg twice daily provided adequate relief of IBS pain and discomfort, and improved stool consistency in men with diarrhea-predominant IBS. PMID- 15654791 TI - Is constipation associated with decreased physical activity in normally active subjects? AB - BACKGROUND: The effectiveness of physical activity in the management of constipation remains controversial. We examined the associations among physical activity, constipation, and quality of life (QoL) in a population of employed adults to determine whether the risk of constipation is related to physical activity. METHODS: A total of 1,069 employees (age range 24-77) of the Veterans Affairs (VA) Black Hills Health Care System were mailed validated questionnaires (response rate 72%), inquiring about bowel habits, QoL (SF 36), and physical activity (modified Baecke questionnaire). Constipation was defined using the Rome I criteria. RESULTS: One hundred and forty (19.4%, 95% CI 16.2-22.4) employees reported constipation. The average total physical activity and all subscales of physical activity were not significantly different in subjects with and without constipation (all p > or = 0.2). Subjects with constipation had lower QoL scores than subjects without constipation, and physical activity was positively correlated with physical functioning and health perception. CONCLUSION: Physical activity appears to be unrelated to the risk of constipation in employed adults, but higher physical activity was associated with improved QoL. Recommendations to increase physical activity may not alter symptoms of constipation but may improve overall well-being. PMID- 15654792 TI - Characteristics of familial juvenile polyps expressing cyclooxygenase-2. AB - OBJECTIVES: Familial juvenile polyposis (FJP) is a dominant genetic disorder characterized by colorectal, gastric, and small bowel juvenile polyps, and high risk for gastrointestinal cancer. Patients are treated by repeated endoscopic polypectomies and elective surgery. We determined the characteristics of FJP polyps expressing cyclooxygenase-2 (COX-2). METHODS: A total of 115 colorectal and 6 gastric polyps were available from 17 FJP patients. Comparison tissues were 18 sporadic juvenile colorectal polyps, 6 gastric hyperplastic polyps, 9 normal colons, and 3 colorectal cancers (CRCs). Histology sections were classified and stained for COX-2. The polyps' epithelium and stroma and comparison tissues were quantified for COX-2 by: area of staining (0-3) x intensity (0-3). Epithelial and stromal scores (0-9) and total scores (0-18) were evaluated in relationship to patient's age, polyp site, size, dysplasia, and stromal cellularity. RESULTS: Colonic FJP polyps mean total COX-2 score was 10.3 +/- 6.0, and that of sporadic juvenile polyps 3.6 +/- 2.2 (p < 0.01), and in contrast to the latter, FJP COX-2 scores increased significantly (p < 0.01) with polyp size. Linear regression analysis showed significant associations of COX-2 in FJP polyps with dysplasia (p < 0.01), stromal cellularity (p < 0.01), size (> or =1.5 cm) (p= 0.02), and site (right colon) (p= 0.01), and not with age. COX-2 total scores of gastric FJP polyps and hyperplastic polyps were similar. CONCLUSIONS: Expression of COX-2 in FJP polyps and its association with size and dysplasia suggest that, in these patients, chemoprevention with selective COX-2 inhibitors might be a useful adjunct therapy to colonoscopic polypectomy. PMID- 15654793 TI - Prevalence of adenomas among young individuals at average risk for colorectal cancer. AB - OBJECTIVES: We evaluated the prevalence and characteristics of adenomas in a young population not genetically predisposed for the development of colorectal cancer (CRC). METHODS: The databases of the Dutch Hereditary Colorectal Cancer Registry were used. The study population included patients (n = 444) who had regular endoscopy until mutation analysis revealed they did not carry the (Adenomatous Polyposis Coli (APC)/Mismatch Repair) gene defect identified in their family. RESULTS: At first colonoscopy (n = 342; 50% males, mean age 37 yr) a total of 19 adenomas (10 males, mean age 50 yr, range 24-91 yr) and two CRCs (2 males, age 49 and 72 yr) were identified, and at first sigmoidoscopy (n = 102; 53% males, mean age 29 yr) three adenomas (2 males, age 8, 40, and 41 yr) were found. A second colonoscopy was performed in 14 patients with, and in 162 patients without an adenoma. Three of 14 patients (21%) developed a new adenoma (all >50 yr) and 8 of 162 (5%) patients developed their first adenoma during follow-up. In the colonoscopy group, the cumulative proportion of patients free of adenomas at age 50 yr was 86%. Of all adenomas diagnosed during colonoscopy (n = 49), 65% were located distal from the flexura lienalis. Of the adenomas detected during all endoscopies (n = 53), 9.8% were > or =7 mm, 7.5% showed high grade dysplasia, and 7.5% showed tubulovillous features. CONCLUSIONS: On the basis of our findings during colonoscopy we conclude that the risk of developing adenomas/CRC in young individuals without genetic risk factors is low. Adenoma surveillance programs should focus on young individuals with a positive family (or personal) history for adenomas/CRC, or on individuals >50 yr. PMID- 15654794 TI - Usefulness of contrast-enhanced transabdominal ultrasonography in the diagnosis of intraductal papillary mucinous tumors of the pancreas. AB - BACKGROUND: The differentiation of benign from malignant intraductal papillary mucinous tumors (IPMT) is often difficult even by various examination methods. We evaluated the qualitative and quantitative diagnostic ability of contrast enhanced transabdominal ultrasonography (CE-US), mainly in differentiating benign from malignant tumors in patients with IPMT. PATIENTS AND METHODS: There were 21 patients with IPMT who underwent CE-US and endoscopic ultrasonography (EUS). Surgery was performed in all 21 patients. Pathological findings were 4 with carcinoma and 17 with adenoma. CE-US was performed using a contrast agent (Levovist; Tanabe, Osaka, Japan) consisting of galactose microbubbles and a small (0.1%) admixture of palmitic acid, and the following items were evaluated by the following procedure. (1) Two reviewers with experienced sonographic and endosonographic ability evaluated CE-US images before and after contrast enhancement and classified the enhancement effects into three grades. In addition, the presence or absence of enhancement effects by CE-US was compared with that of mural nodules visualized by EUS. (2) In all 21 patients, changes in intensity after contrast enhancement were quantitatively measured using an HDI Lab. HDI Lab was provided by ATL (Philips; Bothell, WA) and these software tools rapidly quantify image characteristics within multiple ROI (regions of interest) and make comparisons between several areas or images. In both the early and late phases, the post-enhancement intensity, difference between pre- and post enhancement intensity, and the percentage change ((post-enhancement value-pre enhancement value)/pre-enhancement value) were compared between malignant and benign lesions, and the ability of CE-US to differentiate between benign and malignant lesions was evaluated in comparison with the ability of EUS to diagnose the degree of malignancy. RESULTS: (1) In both the early and the late phases, both reviewers observed enhancement effects in all 21 patients. And both reviewers observed mural nodules by EUS in all 21 patients. (2) In all 21 patients who underwent resection of IPMT, the intensity increased in both the early and late phases. When the patients with carcinoma were compared with those with adenoma, the post-enhancement intensity was significantly higher, and the difference between pre- and post-enhancement intensity and the percentage change in the early phase and the late phase was significantly more marked in the carcinoma group (p= 0.019, p= 0.002, p= 0.015, p= 0.012, and p= 0.039, respectively). CONCLUSIONS: CE-US was useful for qualitatively diagnosing tumor lesions in patients with IPMT. Moreover, quantitative changes in intensity can be a parameter for the differential diagnosis of benign and malignant tumors. PMID- 15654796 TI - The epidemiology of malignant gastrointestinal stromal tumors: an analysis of 1,458 cases from 1992 to 2000. AB - OBJECTIVES: The epidemiology of gastrointestinal stromal tumor has not been well examined, and prior studies often provide conflicting results. We conducted the first population-based study to evaluate the incidence and survival of malignant gastrointestinal stromal tumor in the United States. METHODS: We utilized the Surveillance, Epidemiology, and End Results registry from the National Cancer Institute to identify all cases of malignant gastrointestinal stromal tumor diagnosed from 1992 to 2000. The age-adjusted incidence rates and the survival rates were calculated. Cox proportional hazards models were used to examine the risk of mortality. RESULTS: Between 1992 and 2000, there were 1,458 cases of diagnosed gastrointestinal stromal tumor. The age-adjusted yearly incidence rate was 0.68/100,000. The mean age at diagnosis was 63 yr. Fifty-four percent were men and 46% were women. The incidence rate was higher among men and among Blacks. Fifty-one percent of cases were in the stomach, 36% small intestine, 7% colon, 5% rectum, and 1% in the esophagus. Fifty-three percent of cases were staged as localized, 19% regional, 23% distant, and 5% unstaged. The 1- and 5-yr relative survival rates were 80% and 45%, respectively. The Cox analysis showed that older age, Black race, advanced stage, and receipt of therapy were independent predictors of mortality. CONCLUSIONS: Malignant gastrointestinal stromal tumors rare, but are more common in the older population, men, and Blacks. Risk factors for mortality include older age, Black race, advanced stage, and no surgical intervention. PMID- 15654795 TI - 99mTc-hexamethylpropylene amineoxime leukocyte scintigraphy in acute pancreatitis: an alternative to contrast-enhanced computed tomography? AB - OBJECTIVE: Contrast-enhanced computed tomography (CECT) is the most efficient imaging technique for the diagnosis and staging of acute pancreatitis (AP); its use, however, may be unfeasible in some patients as a consequence of the drawbacks of intravenous (IV) contrast material. The aim of this study was to test the utility of labeled leukocyte scintigraphy (LLS) as an alternative imaging technique to CECT for the staging of AP. METHODS: Sixty-six patients with AP were prospectively studied. All patients underwent CECT and pancreatic LLS using (99m)Tc-hexamethylpropylene amineoxime as leukocyte label within a time interval of 2 days, in the early phase of AP. In addition, all patients had their serum C-reactive protein (CRP) concentration measured within 48-72 h after admission. CECT images were analyzed for Balthazar's grade of pancreatitis and for the presence or absence of pancreatic necrosis. Scintigraphic activity of 3-4 h planar images was scored on a 0-2 scale in relation to physiological liver uptake. RESULTS: LLS score was significantly related (p < 0.001) to both components of CECT (grade of pancreatitis and pancreatic necrosis). LLS and serum CRP showed similar results for detecting the most severe pancreatic damage as showed by their respective receiver operating characteristic (ROC) curves. Sensitivities and specificities of LLS score of 2 were, respectively, 62% and 96% for the detection of grade D-E pancreatitis and 90% and 89% for the detection of pancreatic necrosis. Scintigraphic score of 2 increased the likelihood of grade D E pancreatitis from 32% (pretest probability) to 87% (posttest probability) (likelihood ratio: 13.9) and that of pancreatic necrosis from 16% to 60% (likelihood ratio: 8.4). CONCLUSIONS: Our results show that leukocytes are related to the severity of local pancreatic damage in AP. Thus, LLS is a potential alternative technique to CECT for staging AP. PMID- 15654797 TI - Severe acute respiratory syndrome associated coronavirus is detected in intestinal tissues of fatal cases. AB - OBJECTIVES: A significant percentage of confirmed severe acute respiratory syndrome (SARS) patients experienced gastrointestinal symptoms, and the viral sequence was detectable in the stool of most patients. At present, the knowledge of the pathology of the digestive system in SARS patients is limited. Because a resurgence of the SARS epidemic is constantly possible, there is an urgent need to understand the involvement of the digestive system in this new disease. METHODS: We performed seven SARS autopsies in which samples of alimentary tract and digestive glands were examined with routine pathology, electron microscopy (EM), in situ hybridization (ISH), immunohistochemistry, and real-time polymerase chain reaction (PCR). RESULTS: The main histopathological finding was atrophy of the mucosal lymphoid tissue. A few mucosal epithelial cells and lymphocytes in the intestine were positively stained for coronavirus with ISH. SARS-coronavirus (CoV)-like particles were found in the mucosal epithelial cells under EM and mild focal inflammation was detected in the alimentary tract. One patient who experienced severe diarrhea had pseudomembranous enteritis of the ileum. Fatty degeneration and central lobular necrosis were observed in the liver. No evidence of direct viral infection was found in the esophagus, the stomach, the salivary gland, the liver, or the pancreas. CONCLUSIONS: In addition to the lungs, the gastrointestinal tract is another target of SARS-CoV infection, as the intestinal epithelial cells and mucosal lymphoid tissue are infected. The findings provide possible explanations for the gastrointestinal symptoms and the presence of virus in the stool of SARS patients. PMID- 15654798 TI - Variability of histologic lesions in relation to biopsy site in gluten-sensitive enteropathy. AB - OBJECTIVES: It is generally believed that in gluten-sensitive enteropathy or celiac disease (CD), mucosal lesions may have a patchy distribution. We wanted to verify this concept and establish whether one or more biopsy samples are needed in order to make a correct diagnosis of CD. METHODS: One hundred and twelve consecutive children with positive antiendomysium (EMA) or antitissue transglutaminase (tTGA) antibodies, referred to us for suspected CD, were enrolled in a prospective fashion. During upper GI endoscopy four to five biopsies were taken from Treitz and/or distal duodenum (D3), intermediate duodenum (D2), proximal duodenum (D1), and duodenal bulb (B). Histologic lesions were classified according to Marsh criteria modified by Oberhuber. RESULTS: A total of 110 patients, all HLA-DQ2 or DQ8 positive, had a final diagnosis of CD (59 classic, 28 atypical, and 23 silent): 102/110 (92.7%) had type 3 lesion-(a) mild, (b) moderate, or (c) severe-in at least one site and 94/110 (85.4%) had villous atrophy (VA) of some degree in all sites. VA of identical degree was present in all biopsy sites in 55/110 (50%) patients. Total VA (type 3c) was present in at least one site in 85/110 (75%), in all sites in 50/110 (45.4%), and significantly increased in aborad direction ((chi(2) > 26.22 with (= 0.01 and d.f. (degrees of freedom) = 12). Eight out of 110 (7.2%) CD patients had exclusively type 1 or 2 lesions, no patient had lesion variability >1 degree and none had normal biopsies. There was no correlation between type or distribution of histologic lesions and clinical presentation of CD. CONCLUSIONS: Mucosal atrophy is present in 85% of patients with CD and total VA is significantly more frequent in distal duodenum or proximal jejunum. Fifty percent of patients have identical VA throughout the duodenum and no duodenal areas are histologically normal. In genetically susceptible children with positive serology, a diagnosis of CD can reliably be made even if biopsies are not taken from the distal duodenum or jejunum. PMID- 15654799 TI - The limitations of gastro-jejunal (G-J) feeding tubes in children: a 9-year pediatric hospital database analysis. AB - BACKGROUND: A gastro-jejunal (G-J) feeding tube is a safe and useful temporizing method of providing enteral access in children. Although G-J tubes are often used to obviate the need for a surgical jejunostomy, their long-term use is often associated with mechanical failure. AIM: To review the clinically effective durability of G-J feeding tubes in providing enteral access in children. METHODS: We performed a retrospective review of 102 patients at the Johns Hopkins Children's Center from 1994-2003 whose underlying diagnosis necessitated the need for postpyloric enteral access. RESULTS: Long-term follow-up was obtained in 85 (48 M; 37 F) patients with a median (range) age of 2.0 (0.1-18.0) yr. The most common indication for G-J tube placement was gastroesophageal reflux with aspiration in 51 patients and feeding intolerance and vomiting in 19 patients. The mean (range) number of tube replacements was 2.2 (1-14) over a median (range) duration of follow-up of 39 (2-474) days. The indication for G-J tube replacement included: tube displacement (58), a clogged tube (41), and a cracked tube or ruptured balloon (35). In 52 cases, the cause for G-J tube replacement was undetermined. CONCLUSIONS: G-J feeding tubes are associated with the frequent need for tube maintenance and replacement and may not be the most feasible clinical option in providing long-term (>1 month) enteral access in children intolerant to gastrostomy tube feeds. Future studies are needed to develop innovative percutaneous jejunostomy tube placement techniques that facilitate long-term enteral access. PMID- 15654800 TI - Updated guidelines for the diagnosis and treatment of gastroesophageal reflux disease. AB - Guidelines for the diagnosis and treatment of gastroesophageal reflux disease (GERD) were published in 1995 and updated in 1999. These and other guidelines undergo periodic review. Advances continue to be made in the area of GERD, leading us to review and revise previous guideline statements. GERD is defined as symptoms or mucosal damage produced by the abnormal reflux of gastric contents into the esophagus. These guidelines were developed under the auspices of the American College of Gastroenterology and its Practice Parameters Committee, and approved by the Board of Trustees. Diagnostic guidelines address empiric therapy and the use of endoscopy, ambulatory reflux monitoring, and esophageal manometry in GERD. Treatment guidelines address the role of lifestyle changes, patient directed (OTC) therapy, acid suppression, promotility therapy, maintenance therapy, antireflux surgery, and endoscopic therapy in GERD. Finally, there is a discussion of the rare patient with refractory GERD and a list of areas in need of additional study. PMID- 15654801 TI - The emerging role of the liver in iron metabolism. AB - Iron is essential in health and well-being and its dysregulation is a common theme in disease. Recent advances in our understanding of the molecular biology underlying hemochromatosis and anemia has provided insight into the complex mechanisms implicated in iron metabolism. The proximal small bowel is the major site of iron absorption and, it is becoming increasingly clear that the regulation of this process involves the liver and, in particular, the hepatic antimicrobial peptide hepcidin. A number of studies have shown hepcidin to have an inhibitory function at the level of small bowel iron absorption, although its exact site of action remains to be elucidated. Clearly, identifying the target of hepcidin is of importance and is likely to lead to the development of therapeutic agents in the treatment of iron disorders. PMID- 15654802 TI - Proton-pump inhibitors and outcome of endoscopic hemostasis in bleeding peptic ulcers: a series of meta-analyses. AB - OBJECTIVE: To perform meta-analyses of studies on outcome of bleeding ulcers of different proton-pump inhibitors (PPIs) regimens, after stratification of patients by endoscopic stigmata, and analysis of studies with and without endotherapy. METHODS: A total of 35 randomized trials comparing PPIs to placebo and/or H2-receptor antagonists (H2RAs) in 4,843 patients with high-risk endoscopic stigmata were retrieved. Outcomes were rebleeding, surgery, and mortality. RESULTS: Monotherapy with oral or bolus PPIs was superior to placebo and H2RAs in reducing rebleeding in both bleeders and nonbleeders at index endoscopy; the need for surgery was reduced only when compared to H2RAs. In nonbleeders, PPI monotherapy was as effective as a combination of endotherapy with H2RAs. A combination of endotherapy with PPIs was superior to monotherapy in reducing bleeding and surgery, and superior to endotherapy alone in minimizing rebleeding, but not surgery; the benefit was lost when confronted to endotherapy plus H2RAs, whether PPIs were given as infusion or bolus. By pooling data from studies comparing high doses of PPIs as continuous infusion versus regular doses as intermittent bolus, rebleeding, surgery, and mortality were not significantly different. CONCLUSIONS: Combination of endotherapy with either PPIs or H2RAs is indicated for nonbleeding ulcers at endoscopy with the intent to reduce rebleeding and surgery. Its value may extend to bleeding lesions, but current data are scanty. The benefit appears to be independent from route and doses of PPIs, as oral, bolus, or infusional methods are all effective. PMID- 15654803 TI - Gastrointestinal histoplasmosis. AB - Gastrointestinal histoplasmosis (GIH) is an uncommon disease with protean manifestations. It may occur as a result of mediastinal histoplasmosis or in the setting of progressive dissemination. GIH may be misdiagnosed as inflammatory bowel disease, malignancy, or other intestinal diseases leading to inappropriate therapies and unnecessary surgical interventions. Patients with bowel obstruction, perforation, or bleeding, and systemic findings suggestive of histoplasmosis should be evaluated for GIH. This is especially true for immunosuppressed patients, especially those with AIDS. Diagnosis first requires consideration of histoplasmosis in the differential in patients with the above types of gastrointestinal abnormalities, and second, familiarity with a battery of mycologic and serologic tests. Progressive disseminated histoplasmosis (PDH) is lethal if left untreated, and treatment is highly effective. This review will focus on the clinical and histopathologic features of GIH, the approach to diagnosis, and recommendations for treatment. PMID- 15654804 TI - Myths and misconceptions about chronic constipation. AB - There are many strongly held beliefs about constipation that are not evidence based. The purpose of this review is to address these beliefs concerning various aspects of constipation. There is no evidence to support the theory that diseases may arise via "autointoxication," whereby poisonous substances from stools within the colon are absorbed. Dolichocolon, defined as an elongated colon, should not be seen as a cause of constipation. The role of sex hormones altering gut function during the menstrual cycle appears to be minimal. During pregnancy they may play a role in slowing gut transit. Hypothyroidism can cause constipation, but among patients presenting with constipation, hypothyroidism is rare. A diet poor in fiber should not be assumed to be the cause of chronic constipation. Some patients may be helped by a fiber-rich diet but many patients with more severe constipation get worse symptoms when increasing dietary fiber intake. There is no evidence that constipation can successfully be treated by increasing fluid intake unless there is evidence of dehydration. In the elderly constipation may correlate with decreased physical activity, but many cofactors are likely to play a role. Intervention programs to increase physical activity as part of a broad rehabilitation program may help. It is unlikely that stimulant laxatives at recommended doses are harmful to the colon. A proportion of patients with chronic constipation is dependent of laxatives to achieve satisfactory bowel function, but this is not the result of prior laxative intake. Tolerance to stimulant laxatives is uncommon. There is no indication for the occurrence of "rebound constipation" after stopping laxative intake. While laxatives may be misused, there is no potential for addiction. PMID- 15654805 TI - Development of macro-aspartate aminotransferase in a patient undergoing specific allergen injection immunotherapy. AB - Macro-aspartate aminotransferase (macro-AST), a complex between normal AST and an immunoglobulin, is recognized as a cause of isolated elevation of AST. Though its pathogenesis is unknown, previous reports have been suggestive of an autoimmune process. We describe a case of macro-AST formation in a patient with previously normal liver enzymes in whom an isolated AST elevation was discovered after initiation of specific allergen injection immunotherapy (SIT) for allergic rhinitis. We propose that SIT in this otherwise healthy patient led to the formation of macro-AST as a consequence of antibody cross-reaction (molecular mimicry). Awareness of this possible mechanism of macroenzyme development may be helpful to physicians evaluating patients with isolated elevations in AST. PMID- 15654806 TI - High-risk behavior and sexual transmission of hepatitis C. PMID- 15654808 TI - Psychiatric disease and functional GI disorders in the community: more evidence for a casual link? PMID- 15654809 TI - Less H. pylori not necessarily more GERD. PMID- 15654810 TI - Treatment for NASH: the value of histology. PMID- 15654811 TI - Re: Surgical volume and long-term survival following surgery for colorectal cancer in the Veterans Affairs Health-Care System. PMID- 15654813 TI - Exploiting T cell receptor genes for cancer immunotherapy. AB - Adoptive antigen-specific immunotherapy is an attractive concept for the treatment of cancer because it does not require immunocompetence of patients, and the specificity of transferred lymphocytes can be targeted against tumour associated antigens that are poorly immunogenic and thus fail to effectively trigger autologous T cell responses. As the isolation and in vitro expansion of antigen-specific lymphocytes is difficult, 'conventional' adoptive T cell therapy can only be carried out in specialized centres in small numbers of patients. However, T cell receptor (TCR) genes isolated from antigen-specific T cells can be exploited as generic therapeutic molecules for 'unconventional' antigen specific immunotherapy. Retroviral TCR gene transfer into patient T cells can readily produce populations of antigen-specific lymphocytes after a single round of polyclonal T cell stimulation. TCR gene modified lymphocytes are functionally competent in vitro, and can have therapeutic efficacy in murine models in vivo. TCR gene expression is stable and modified lymphocytes can develop into memory T cells. Introduction of TCR genes into CD8(+) and CD4(+) lymphocytes provides an opportunity to use the same TCR specificity to produce antigen-specific killer and helper T lymphocytes. Thus, TCR gene therapy provides an attractive strategy to develop antigen-specific immunotherapy with autologous lymphocytes as a generic treatment option. PMID- 15654814 TI - Wiskott-Aldrich syndrome: another piece in the puzzle. PMID- 15654815 TI - A role for cytokine measurement in therapeutic monitoring of immunosuppressive drugs following lung transplantation. PMID- 15654816 TI - A new mouse model of lung allergy induced by the spores of Alternaria alternata and Cladosporium herbarum molds. AB - Asthma is a serious health problem and during the last decade various experimental models of asthma have been developed to study the pathogenesis of this disease. In this study we describe a new mouse model of asthma that uses the spores of Alternaria alternata and Cladosporium herbarum, two allergenic molds recognized as common inducers of rhinitis and asthma in humans. Here we demonstrate that A. alternata and C. herbarum spores are immunogenic when injected into BALB/c mice, and induce the production of specific IgM and IgG1 antibodies and strongly increase IgE serum levels. To induce the allergic response, mice were sensitized by two intraperitoneal (i.p.) injections and then intranasaly (i.n.) challenged with A. alternata and C. herbarum spores. Bronchoalveolar lavages (BALs) from these mice contained numerous macrophages, neutrophils, eosinophils and lymphocytes whereas neutrophils were the predominant BAL inflammatory cells in nonsensitized mice. Histological studies demonstrated an influx of eosinophils in peri-vascular and peri-bronchial areas and the presence of numerous epithelial goblet cells only in sensitized mice. Increased expression of mRNA specific for various chemokines (eotaxin, MIP-1alpha, MIP-2) and chemokine receptors (CCR-1, CCR-2 and CCR-5) was observed in the lungs of nonsensitized mice challenged with the spores. Expression of CCR-3 mRNA in the lungs and Th2 cytokine (IL-4, IL-5 and IL-13) secretion in the BAL was additionally observed in sensitized and challenged mice. Finally we demonstrate through whole-body plethysmography that mold spore sensitization and challenge induce the development of an airway hyperreactivity in response to nebulized methacholine. PMID- 15654817 TI - Correlation of disease evolution with progressive inflammatory cell activation and migration in the IL-4 transgenic mouse model of atopic dermatitis. AB - Atopic dermatitis is a chronic inflammatory skin disease characterized by inflammatory cell infiltration in the skin. In order to assess the roles of inflammatory cells in this disease, we analysed the activation status and surface markers of various leucocytes in the IL-4 transgenic mouse model of atopic dermatitis, by flow cytometry, immuofluorescence microscopy, and T cell proliferation assays. The studies were performed with a nontransgenic mouse control and transgenic mice at three disease stages: before disease onset, early skin disease, and late skin disease, so that we can delineate the immunological sequence of events. As the skin disease evolves, the skin draining lymph node cells from IL-4-Tg mice show a spontaneous proliferation and a progressively enhanced proliferative response to stimulants including anti-CD3, Con A, PHA, and Staphylococcus enterotoxins A and B. As the disease evolves, the percent of lymphoid organ T cells expressing activation molecules (CD44 and CD69) and costimulatory molecules (ICOS and PD-1) are progressively increased; the percent and total number of T cells are reduced in an incremental manner in the secondary lymphoid organs while the number of T cells infiltrating the skin increases in an incremental fashion; the total number of dendritic antigen presenting cells, macrophages, and NK cells gradually increases in the lymphoid organs. Collectively, our results suggest that there is a continued and progressive migration of activated inflammatory cells from the secondary lymphoid organs into the skin where they participate in immune responses resulting in the pathology associated with inflammation. PMID- 15654818 TI - Dietary n-3 polyunsaturated fatty acids suppress splenic CD4(+) T cell function in interleukin (IL)-10(-/-) mice. AB - Our laboratory has demonstrated that down-regulation of proliferation and cytokine synthesis by CD4(+) T cells in mice fed diets rich in n-3 polyunsaturated fatty acids (PUFA) is highly dependent on the involvement of the co-stimulatory molecule, CD28. It has been reported that the inhibitory cytokine interleukin (IL)-10 acts directly on T cells which up-regulate IL-10 receptor (IL 10R) expression following stimulation via CD28 by efficiently blocking proliferation and cytokine production. Thus, it was hypothesized that dietary n-3 PUFA would suppress T cell function through the effects of IL-10. The proliferation of purified splenic CD4(+) T cells activated in vitro with anti-CD3 and anti-CD28 (alphaCD3/CD28) from conventional mice (C57BL/6) fed either a control corn oil (CO)-enriched diet devoid of n-3 PUFA, docosahexaenoic acid (DHA; 22 : 6) or eicosapentaenoic acid (EPA; 20 : 5) for 14 days was suppressed by dietary DHA and EPA. Surprisingly, a similar trend was seen in IL-10 gene knock-out (IL-10(-/-)) mice fed dietary n-3 PUFA. IL-10R cell surface expression was also significantly down-regulated on CD4(+) T cells from both the C57BL/6 and IL-10(-/-) mice fed dietary n-3 PUFA after 72 h of in vitro stimulation with alphaCD3/CD28. Enzyme-linked immunosorbent assay (ELISA) measurements revealed that C57BL/6 mice fed DHA had significantly reduced interferon (IFN)-gamma and IL 10 levels 48 h post-activation. However, CD4(+) T cells from IL-10(-/-) mice fed dietary n-3 PUFA produced significantly greater levels of IFN-gamma than the CO fed group. Our data suggest that in the absence of IL-10, CD4(+) T cells from n-3 PUFA-fed mice may up-regulate IFN-gamma. Suppressed CD4(+) T cells from n-3 PUFA fed C57BL/6 mice may use mechanisms other than IL-10 to down-regulate T cell function. PMID- 15654819 TI - Estren-mediated inhibition of T lymphopoiesis is estrogen receptor-independent whereas its suppression of T cell-mediated inflammation is estrogen receptor dependent. AB - Estrogen has extensive effects on the immune system. The aim of the present experiments was to compare the effects of 17beta-estradiol (E2) and 4-estren 3alpha,17beta-diol (estren) on T lymphopoiesis and T cell-dependent inflammation. In order to investigate the role of estrogen receptors (ER) in the effects of E2 and estren on the immune system, ER knock-out mice lacking both ERalpha and ERbeta (DERKO) were used. T lymphopoiesis and T cell-dependent inflammation were studied by investigating thymus cellularity, the delayed-type hypersensitivity (DTH) reaction, CD4(+) T cells in spleen and serum levels of interleukin (IL)-6. As expected, the presence of ERs was mandatory for all the effects of E2. In contrast, treatment with estren reduced thymus cellularity in ER knock-out mice, indicating an effect through ER-independent pathways. Interestingly, estren suppressed only DTH, the frequency of CD4(+) T cells in spleen and serum levels of IL-6 in wild-type (WT) mice, but not in mice lacking ERs. Thus, our study is the first to show that estren inhibits T lymphopoiesis via ER-independent pathways, whereas its suppressive effects on inflammation are ER-dependent. PMID- 15654820 TI - Interactions between the mannose receptor and thyroid autoantigens. AB - Thyroid autoantigens require internalization and processing by antigen-presenting cells to induce immune responses. Besides pinocytosis, antigen uptake can be receptor-mediated. The mannose receptor (ManR) has a cysteine rich domain (CR) and eight carbohydrate recognition domains (CRD) that bind glycosylated proteins. The TSH receptor (TSHR), thyroid peroxidase (TPO) and thyroglobulin (Tg) are glycoproteins. To investigate a role for the ManR in thyroid autoimmunity, we tested the interaction between these autoantigens and chimeric ManRs. Plasmids encoding the CR-domain linked to IgG-Fc (CR-Fc) and CDR domains 4-7 linked to IgG Fc (CDR4-7-Fc) were expressed and purified with Protein A. Enzyme-linked immunosorbent assay (ELISA) plates were coated with human thyroid autoantigens and CR-Fc or CRD4-7-Fc binding detected with peroxidase-conjugated anti-IgG-Fc. CRD4-7-Fc binding was highest for the TSHR, followed by Tg and was minimal for TPO. CR-Fc bound to Tg but not to TSHR or TPO. The interaction between the TSHR and CRD-Fc was calcium-dependent; it was inhibited by mannose (not galactose), and required a glycosylated TSHR A-subunit. Moreover, precomplexing the TSHR A subunit with CRD-Fc (but not CR-Fc), or adding mannose (but not galactose), decreased in vitro responses of splenocytes from TSHR-immunized mice. Our data indicate that the ManR may participate in autoimmune responses to Tg and the TSHR but not to TPO. Most important, ManR binding of heavily glycosylated TSHR A subunits suggests a mechanism by which the minute amounts of A-subunit protein shed from the thyroid may be captured by antigen-presenting cells located in the gland or in draining lymph nodes. PMID- 15654821 TI - Mycophenolate mofetil and roscovitine decrease cyclin expression and increase p27(kip1) expression in anti Thy1 mesangial proliferative nephritis. AB - The response of mesangial cells to a phlogistic challenge includes cell proliferation and mesangial matrix expansion. Cell proliferation is a highly regulated process which includes enhancing factors such as cyclins, cyclin dependent kinases, and inhibitory proteins, such as p27(kip1). The aim of the study was to evaluate the effects of Mycophenolate mofetil (MMF), and roscovitine (R), on the cell cycle regulatory system when administered in the florid phase of the experimental model of mesangial proliferative nephritis induced by the anti Thy-1 antigen monoclonal antibody. Three days after nephritis induction, different groups were given MMF and R. Rats treated with MMF or R showed a slight decrease in mesangial proliferation and matrix expansion. Samples of cortical tissue were tested by 'real time' RT-PCR in order to study gene expression of cyclins B, D1, D2, D3, E, and the cyclin inhibitor p27(kip1). Localization of mRNA was evaluated by in situ hybridization. Real time RT-PCR analysis showed a significant decrease in cyclins B, D1, D2, and D3 in rats treated with either MMF or R as compared to controls. Both MMF and R treatment induced a significant increase in p27(kip1) mRNA expression. In situ hybridization showed a mesangial endothelial expression pattern in glomeruli. The number of labelled cells per glomerulus, the number of positive glomeruli in each examined slide as well as cyclin D2 and D3 signal intensity was significantly lower in rats treated with MMF or R as compared to controls, whereas MMF or R treatment up-regulated p27(kip1) mRNA expression. Immunohistochemical evaluation of p27(kip1) aimed to examine the influence of MMF or R on protein expression confirmed up-regulation. PMID- 15654822 TI - Alteration of newly induced endochondral bone formation in adult mice without tumour necrosis factor receptor 1. AB - Tumour necrosis factor (TNF)-alpha, a major proinflammatory cytokine, exerts its role on bone cells through two receptors (TNFR1 and TNFR2). TNFR1, but not TNFR2, is expressed by osteoblasts and its function in bone formation in vivo is not fully understood. We compared in vivo new bone formation in TNFR1-deficient (TNFR1(-/-)) mice and wild-type mice, using two models of bone formation: intramembranous ossification following tibial marrow ablation and endochondral ossification induced by bone morphogenetic protein (BMP)-2. Intramembranous osteogenesis in TNFR1(-/-) mice did not differ from the wild-type mice either in histomorphometric parameters or mRNA expression of bone-related markers and inflammatory cytokines. During endochondral osteogenesis, TNFR1(-/-) mice formed more cartilage (at post-implantation day 9), followed by more bone and bone marrow (at day 12). mRNAs for BMP-2, -4 and -7 were increased during the endochondral differentiation sequence in TNFR1(-/-) mice. The expression of receptor activator of NF-kappa B ligand (RANKL) and receptor activator of NF kappa B (RANK), as assessed by quantitative reverse transcription polymerase chain reaction (RT-PCR), was also increased significantly during endochondral ossification in TNFR1(-/-) mice. In conclusion, signalling through the TNFR1 seems to be a negative regulator of new tissue formation during endochondral but not intramembranous osteogenesis in an adult organism. BMPs and RANKL and its receptor RANK may be involved in the change of local environment in the absence of TNFR1 signalling. PMID- 15654823 TI - Production of interleukin (IL)-10 and IL-12 by murine colonic dendritic cells in response to microbial stimuli. AB - Intestinal dendritic cells (DC) are likely to regulate immunity to gut microflora, but little is known about their responses to bacterial antigens. Therefore, DC from normal murine colon were characterized and their cytokine responses to components of Gram-negative and/or Gram-positive bacteria assessed. Cells were obtained by digestion of colonic tissue and contained DC that were identified by flow cytometry as CD11c(+) major histocompatibility complex (MHC) class II(+) cells. Purified DC were obtained by immunomagnetic separation plus cell sorting. DC had the morphology of immature myeloid cells, were endocytically active, expressed low levels of co-stimulatory molecules and stimulated a weak allogeneic mixed leucocyte reaction. Analysis of flow cytometry data by a sensitive subtraction method allowed measurement of production of interleukin (IL)-12 and IL-10 by small numbers of gut DC by intracellular staining. Fewer than 5% of unstimulated DC produced either IL-10 or IL-12. IL-10 production was significantly up-regulated following stimulation with Bifidobacteria longum, but not after exposure to lipopolysaccharide (LPS) or Streptococcus faecium. In contrast, colonic DC produced IL-12 in response to both LPS and B.longum. Thus, colonic DC can produce both IL-12 and IL-10 following bacterial stimulation. Cell wall components from different bacteria stimulate distinct responses and may direct immune responses differentially in the gut. PMID- 15654824 TI - Generation of Th1 T cell responses directed to a HLA Class II restricted epitope from the Aspergillus f16 allergen. AB - The Aspergillus allergen Asp f16 has been shown to confer protective Th1 T cell mediated immunity against infection with Aspergillus conidia in murine models. Here, we use overlapping (11-aa overlap with preceding peptide) pentadecapeptides spanning the entire 427-aa coding region of Asp f16 presented on autologous dendritic cells (DC) to evaluate the ability of this antigen to induce Th1 responses in humans. Proliferative responses were induced in five out of five donors, and one line with a high frequency of interferon (IFN)-gamma-producing CD4(+) T cells in response to the complete peptide pool was characterized. This line was cytotoxic to autologous pool-pulsed and Aspergillus culture extract pulsed targets. Limitation of cytotoxicity to the CD4(+) T cell subset was demonstrated by co-expression of the degranulation marker CD107a in response to peptide pool-pulsed targets. Cytotoxic T lymphocytes (CTL) killed Aspergillus hyphae and CTL culture supernatant killed Aspergillus conidia. By screening 21 smaller pools and individual peptides shared by positive pools we identified a single candidate sequence of TWSIDGAVVRT that elicited responses equal to the complete pool. The defined epitope was presented by human leucocyte antigen (HLA) DRB1-0301. These data identify the first known Aspergillus-specific T cell epitope and support the use of Asp f16 in clinical immunotherapy protocols to prime protective immune responses to prevent or treat Aspergillus infection in immunocompromised patients. PMID- 15654825 TI - Analysis of the susceptibility of CD57 T cells to CD3-mediated apoptosis. AB - After stimulation with anti-CD3 antibody in vitro, CD57(+) T cells showed a greater susceptibility to apoptosis than CD57(-)alphabetaT cell receptor (TCR)(+) T cells (regular alphabeta T cells). The apoptotic fraction of CD57(+) T cells showed an increased production of active caspase-3. An increase in both Fas expression and Fas-ligand (FasL) production was also observed in CD57(+) T cells, whereas the expression of survivin was suppressed in CD57(+) T cells compared to that of regular alphabeta T cells. CD57(+) T cells display a biased expansion of a few Vbeta T cell fractions in individuals, but such Vbeta T cells were not specifically susceptible to CD3-mediated apoptosis. The TCR expression level of CD57(+) T cells was much lower than that of regular T cells and anti-TCR antibody stimulation induced a smaller apoptotic proportion of CD57(+) T cells than did anti-CD3 antibody. Although the CD3epsilon expression levels were similar in both T cell subsets, the CD3zeta level of CD57(+) T cells was significantly higher than that of regular T cells. These results suggest that several apoptotic and anti-apoptotic molecules are involved in the CD3-induced apoptosis of CD57(+) T cells and raise the possibility that the imbalance in expression of the CD3epsilon and CD3zeta chains may also contribute to the susceptibility of CD57(+) T cells to undergo apoptosis. PMID- 15654826 TI - Taurine attenuates CD3/interleukin-2-induced T cell apoptosis in an in vitro model of activation-induced cell death (AICD). AB - Interleukin (IL)-2 immunotherapy is used for the treatment of metastatic melanoma and renal cell carcinoma and mediates its effects through the clonal expansion of lymphocytes. Although IL-2 remains the most effective form of therapy for these cancers, response rates are poor and dose escalation is hampered by side effects, which include vascular leak and lymphopenia. The mechanism underlying T cell loss is currently unidentified but could be the induction of activation-induced cell death (AICD) mediated by FasL. Our previous studies have shown that the amino acid taurine can attenuate apoptosis induced by a number of factors in different cell types. Here, we induced T cell AICD via CD3 and IL-2 stimulation and investigated the effect of taurine on lymphocyte apoptosis. Anti-CD3-activated Jurkat T cells treated with IL-2 significantly increased FasL expression, which was associated with increased apoptosis. Treatment with taurine prior to stimulation down-regulated FasL protein expression and partially inhibited apoptosis. Inhibition of FasL-signalling resulted in an identical reduction in apoptosis. As the kinetics of AICD are completely different in circulating T cells, we repeated these experiments in such cells to confirm our finding. Stimulation of CD4(+) circulating T cells induced apoptosis in sensitized, but not freshly isolated T cells, which was abrogated partially by taurine. In Jurkat cells it was determined that taurine-mediated down-regulation of FasL protein expression was associated with decreased FasL mRNA expression and reduced NFkappaB activation. These results reveal one possible mechanism underlying the lymphopenia observed with IL-2 immunotherapy, involving increased FasL expression leading to apoptosis. Taurine may be of use in reversing the lymphopenia associated with IL-2, thereby augmenting its immunotherapeutic potential. PMID- 15654827 TI - Effects of prolactin and cortisol on natural killer (NK) cell surface expression and function of human natural cytotoxicity receptors (NKp46, NKp44 and NKp30). AB - The surface density of the triggering receptors (e.g. NKp46 and NKp30) responsible for natural killer (NK) cell-mediated cytotoxicity determines the ability of NK cells to kill susceptible target cells. In this study, we show that prolactin up-regulates and cortisol down-regulates the surface expression of NKp46 and NKp30. The prolactin-mediated activation and the cortisol-mediated inhibition of natural cytotoxicity receptor (NCR) surface expression reflects gene regulation at the transcriptional level. NKp46 and NKp30 are the major receptors involved in the NK-mediated killing of K562, a human chronic myelogenous leukaemia cell line. Accordingly, the prolactin dramatically increased the NK-mediated killing of the K562 cell line, whereas cortisol abolished this activity. Our data suggest a mechanism by which prolactin activates the lytic function of NK cells, and cortisol inhibits the NK-mediated attack. PMID- 15654829 TI - Deficiency of the mannan-binding lectin pathway of complement and poor outcome in cystic fibrosis: bacterial colonization may be decisive for a relationship. AB - In cystic fibrosis (CF) prognosis concerning lung damage development is highly variable and difficult to predict. Mannan-binding lectin (MBL) deficiency has been reported to be associated with poor outcome in CF lung disease. MBL is a recognition molecule of the MBL pathway of the complement system and is encoded by a gene characterized by a high degree of polymorphism. Some genotypes result in low serum concentrations of MBL. MBL-associated serine protease 2 (MASP-2) is another protein belonging to the MBL pathway. A mutation resulting in low levels of MASP-2 in serum has been described recently. In the present study, 112 CF patients aged 4-54 years were investigated for MBL and MASP-2 genotypes, serum levels of MBL and MASP-2 and the MBL pathway function in serum. No correlation to reduced lung function or need for lung transplantation was seen, either for MBL deficiency, MASP-2 gene mutation or reduced MBL pathway function. However, in the 27 patients colonized with Staphylococcus aureus, MBL-deficient genotypes were associated with decreased lung function. As expected, MBL pathway function in serum was reduced both in MBL-deficient patients and in patients carrying a mutant MASP-2 allele. An unexpected finding was that CF patients had higher serum levels of MBL than healthy controls when corrected for MBL genotype. In conclusion, MBL pathway function was affected both by MBL and by MASP-2 genotypes. However, MBL or MASP-2 levels in serum did not affect the clinical outcome in the cohort of CF patients studied. PMID- 15654828 TI - Phenotypic perturbation of B cells in the Wiskott-Aldrich syndrome. AB - Wiskott-Aldrich syndrome (WAS) is an X-linked immunodeficiency/platelet disease due to mutations of WASP, a cytoskeletal regulatory protein of blood cells. Patients exhibit a range of immune defects generally attributed to defective T cell function, including poor response to immunization, skewed immunoglobulin isotypes, eczema, recurrent infections, autoimmune disease and increased frequency of malignancies. Here we show a deficit of total B-cells in WAS patients of various ages and identify phenotypic perturbations involving complement receptors and CD27. Whereas B-cells of normal healthy donors are overwhelmingly CD21/CD35-positive, B-cells expressing these receptors are significantly reduced in number in WAS patients, and their paucity may cause suboptimal antigen capture and presentation. The frequencies of IgD(-) and IgG(+) patient B-cells were not different from healthy donors (although absolute numbers were decreased), indicating that isotype switching is occurring. In contrast, the frequency of cells positive for CD27, the marker of post germinal centre B-cells, was significantly decreased even among isotype-switched cells, and B-cells resembling germinal centre progenitors (CD10(+)CD27(-)CD38(bright)) were more frequent in adult patients, suggesting impaired germinal centre maturation/differentiation. The documentation of these phenotypic perturbations and deficit of total cells suggest that defects intrinsic to B-cells contribute to the impaired humoral immunity that characterizes this disease. PMID- 15654831 TI - The chemokine receptor CCR7 is expressed on epithelium of non-inflamed gastric mucosa, Helicobacter pylori gastritis, gastric carcinoma and its precursor lesions and up-regulated by H. pylori. AB - CCR7 chemokine-receptor expression on tumour cells of gastric carcinoma has been associated with lymph-node metastasis and is thought to play an important role in metastasis. However, so far it is unknown whether CCR7 is newly up-regulated on gastric carcinoma or already expressed in non-neoplastic gastric epithelium. Therefore, epithelial CCR7 expression was investigated in the process of gastric carcinogenesis: non-inflamed mucosa --Helicobacter pylori gastritis -- intestinal metaplasia/dysplasia -- gastric carcinoma. CCR7 was expressed by gastric epithelium in non-inflamed gastric mucosa (n = 5), H. pylori gastritis (n = 17), intestinal metaplasia (n = 10), dysplasia (n = 3) and on tumour cells in 20 of 24 patients with gastric carcinoma (13/14 intestinal-type; 7/10 diffuse-type) as tested by immunohistochemistry. As CCR7 expression by gastric epithelium was significantly stronger in H. pylori gastritis than in non-infected mucosa, the influence of H. pylori on CCR7 receptor expression of gastric epithelial cells was investigated by fluorescence activated cell sorter analysis. H. pylori strains up-regulated the CCR7 chemokine-receptor in CCR7-positive cell lines. No difference in CCR7 up-regulation between cag(+) and cag(-)H. pylori strains was found. Epithelial CCR7 up-regulation by H. pylori may alter the metastatic fate of gastric carcinoma. Additionally, CCR7 expression not only on gastric carcinoma, but also on non-neoplastic gastric epithelium, suggests a novel biological function. PMID- 15654830 TI - Impaired interleukin (IL)-4-associated generation of CCR4-expressing T cells in neonates with hereditary allergy risk. AB - Reduced microbial exposure in early life may contribute to the increase of atopic diseases in 'westernized' societies but the underlying mechanisms remain elusive. The objective of this study was to examine how exposure to bacterial lipopolysaccharide (LPS) during early antigen encounter might influence the maturation of neonatal lymphoid cells, and to define possible differences in this respect between neonates with high risk of allergy due to a family history (FH(+)) and controls with no apparent hereditary risk (FH(-)). Cord blood mononuclear cells from the FH(+) or FH(-) group were stimulated with pure LPS or beta-lactoglobulin (beta-LG) in the presence of LPS. T cell expression of chemokine receptors CCR4 and CXCR3 was determined by flow cytometry and reverse transcription-polymerase chain reaction (RT-PCR). Cellular expression of interleukin (IL)-4 was analysed by quantitative RT-PCR, whereas interferon (IFN) gamma was analysed by both quantitative RT-PCR and immunoassay. Stimulation with LPS, or beta-LG together with LPS, induced up-regulation of CCR4 (P < 0.05) and CXCR3 (P < 0.05). For CCR4, such up-regulation was related to the level of IL-4 produced by the same T cells (r(S) = 0.49, P = 0.03), while CXCR3 expression was negatively correlated with the IL-4 levels (r(S) = -0.56, P = 0.02). Compared with the FH(-) group, the FH(+) group showed a significantly lower capacity for generation of CCR4(+) T cells (mean percentage of total T cells: FH(+), 2.42%versus FH(-), 5.74%; P < 0.01), whereas induction of CXCR3 and IFN-gamma did not differ significantly between the two groups. When the immune system in early life encounters antigen together with LPS, the T cell potential for compartmentalized interaction with other immune cells might be increased by elevated CCR4- and CXCR3-expression levels. In neonates at hereditary allergy risk, this putative homeostatic mechanism could theoretically be jeopardized due to decreased up-regulation of CCR4. Conversely, Th1 responses to antigen in the presence of LPS did not appear to be reduced compared with controls. PMID- 15654832 TI - Identification of epithelial auto-antigens associated with periodontal disease. AB - We previously reported evidence that patients with periodontitis have serum antibodies to oral Gram positive bacteria that are cross-reactive with epithelial antigens. In the present report cross-reactive epithelial antigens including CD24, lactate dehydrogenase A [LDM-A], antioxidant protein 2 [AOP 2] and nuclear factor of activated T cells 5 [NFAT 5], were identified by screening a cDNA expression library with pooled patient sera. Titres of antibodies to CD24 peptide correlated negatively with indices of periodontal disease severity. Strong expression of CD24 in the reactive periodontal epithelium and inflamed gingival attachment contrasted with low to undetectable expression in the external gingival epithelium. In periodontitis, a local action of these auto-reactive antibodies could modulate the regulatory potential associated with expression of CD24 in this epithelium. PMID- 15654833 TI - Elevated macrophage migration inhibitory factor (MIF) levels in the urine of patients with focal glomerular sclerosis. AB - The pathogenesis of focal glomerular sclerosis (FGS) is poorly understood. Macrophage migration inhibitory factor (MIF) is a potent pro-inflammatory cytokine released from T cells and macrophages, and is a key molecule in inflammation. To examine further the possible role of MIF in FGS, we measured MIF levels in the urine. The purpose of the present study was to evaluate the involvement of MIF in FGS. Urine samples were obtained from 20 FGS patients. The disease controls included 40 patients with minimal-change nephrotic syndrome (MCNS) and membranous nephropathy (MN). A group of healthy subjects also served as controls. Biopsies were performed in all patients prior to entry to the study. The samples were assayed for MIF protein by a sandwich enzyme-linked immunosorbent assay (ELISA). The levels of MIF in the urine of FGS patients were significantly higher than those of the normal controls and patients with MCNS and MN. In contrast, the levels of urinary MIF (uMIF) in patients with MCNS and MN did not differ significantly from normal values. In the present study, attention also focused on the relationship between uMIF levels and pathological features. Among the patients with FGS, uMIF levels were significantly correlated with the grade of mesangial matrix increase and that of interstitial fibrosis. There was also a significant correlation between uMIF levels and the number of both intraglomerular and interstitial macrophages. Although the underlying mechanisms remain to be determined, our study presents evidence that urinary excretion of MIF is increased in FGS patients with active renal lesions. PMID- 15654834 TI - T cell repertoire in patients with stable scleroderma. AB - At onset of systemic sclerosis (SSc), T cells have been found to oligoclonally expand in the skin, presumably in response to auto-antigens, but the T cell repertoire has not been evaluated at a later stage. To determine whether a perpetuating immune response contributes to the pathogenesis of stable SSc, the T cell repertoire was analysed in patients with diffuse (d) or limited (l) SSc, and compared to patients with primary Raynaud's phenomenon (RP) or healthy volunteers (Ctrl). The T cell repertoire (total, CD4 or CD8 sorted blood T cells) was analysed by qualitative and quantitative immunoscope (14 BV families analysed) in 11 untreated dSSc and 11 untreated lSSc, 10 RP and 11 Ctrl. To better detect in vivo activated cells, repertoire analysis was also performed on sorted CD4 T cells after in vitro culture with IL-2. In parallel, 6 skin biopsies from SSc patients were analysed. After 7-8 years of disease evolution, SSc patients did not show detectable clonal T cell expansions in the skin, even after tentative expansion from the biopsy with IL-2. Total T cell, sorted CD4 and CD8 T cell repertoires from the blood of patients with SSc did not show significant perturbation as compared to patients with RP and Ctrl. After IL-2 culture for 7 days, blood CD4 T cells from the patients did not preferentially expand as compared to RP and Ctrl. These findings suggest that antigen-driven immune responses may play a lesser role in established SSc than at disease onset. PMID- 15654835 TI - Autoantibodies directed against the protease inhibitor calpastatin in psoriasis. AB - Psoriasis is believed to be a T cell-mediated autoimmune disease, but also exhibits autoantibody production. Calpastatin is an endogenous inhibitor of calpain, a ubiquitous protease that regulates inflammatory processes. Anti calpastatin autoantibody was first identified as an autoantibody specific to rheumatoid arthritis, but has been also detected in other autoimmune diseases. In this study, we examined the presence and levels of anti-calpastatin antibody in 77 psoriasis patients by enzyme-linked immunosorbent assay. Compared with normal controls, psoriasis patients exhibited significantly elevated IgG anti calpastatin antibody levels that were similar to those found in rheumatoid arthritis patients. Remarkably, IgG anti-calpastatin autoantibody in sera from psoriasis patients inhibited calpastatin activity. Calpain II expression was up regulated in psoriasis skin lesions compared with normal skin while calpastatin expression was normal. The results of this study reveal the presence of anti calpastatin autoantibody in psoriasis. PMID- 15654836 TI - Excessive expression of Txk, a member of the Tec family of tyrosine kinases, contributes to excessive Th1 cytokine production by T lymphocytes in patients with Behcet's disease. AB - Excessive Th1 cell function is importantly involved in the pathogenesis of Behcet's disease (BD). We previously found that Txk, a member of the Tec family of tyrosine kinases, acts as a Th1 cell specific transcription factor. To investigate immune aberration in the pathogenesis of BD, we studied the expression of Txk and Th1 cytokines in peripheral blood lymphocytes (PBL) and skin lesions in patients with BD. Cytokine production by the lymphocytes was assessed using ELISA. PBL produced excessive Th1 associated cytokines including IFN-gamma and IL-12 spontaneously and in response to exogenous HSP60-derived peptide stimulation, which was shown to induce proliferation of PBL, in patients with BD. Circulating CD4+ T cells expressed excessive Txk protein. A majority of cells infiltrating into skin lesions expressed IFN-gamma in the BD specimens. IL 12 and IL-18 were also expressed in the mononuclear cell aggregates. Lymphocytes accumulating in the skin lesion expressed higher levels of Txk as compared with atopic dermatitis lesions, a typical Th2 disease. IFN-gamma, IL-18 and Il-12 were detected in the BD skin lesions, which may induce preferential development of Th1 cells in patients with BD. The mononuclear cell aggregates contained Txk expressing cells in such skin lesions. Collectively, Txk expressing Th1 cells and the Th1 associated cytokines may play a critical role in the development of skin lesions in BD. PMID- 15654838 TI - Doublecortin expression levels in adult brain reflect neurogenesis. AB - Progress in the field of neurogenesis is currently limited by the lack of tools enabling fast and quantitative analysis of neurogenesis in the adult brain. Doublecortin (DCX) has recently been used as a marker for neurogenesis. However, it was not clear whether DCX could be used to assess modulations occurring in the rate of neurogenesis in the adult mammalian central nervous system following lesioning or stimulatory factors. Using two paradigms increasing neurogenesis levels (physical activity and epileptic seizures), we demonstrate that quantification of DCX-expressing cells allows for an accurate measurement of modulations in the rate of adult neurogenesis. Importantly, we excluded induction of DCX expression during physiological or reactive gliogenesis and excluded also DCX re-expression during regenerative axonal growth. Our data validate DCX as a reliable and specific marker that reflects levels of adult neurogenesis and its modulation. We demonstrate that DCX is a valuable alternative to techniques currently used to measure the levels of neurogenesis. Importantly, in contrast to conventional techniques, analysis of neurogenesis through the detection of DCX does not require in vivo labelling of proliferating cells, thereby opening new avenues for the study of human neurogenesis under normal and pathological conditions. PMID- 15654837 TI - Involvement of Th1 cells and heat shock protein 60 in the pathogenesis of intestinal Behcet's disease. AB - Involvement of excessive Th1 cell functions and heat shock protein expression in the pathogenesis of Behcet's disease (BD) has been reported. In this study we have characterized immune responses in intestinal lesions of BD. Peripheral blood lymphocytes (PBL) of BD and healthy controls (HC) and tissue specimens of intestinal Behcet's disease (intestinal BD), Crohn's disease (CD) and ulcerative colitis (UC) were analysed for mRNA and protein expression by reverse transcriptase-polymerase chain reaction (PCR) and immunohistochemistry, respectively. PBL of BD patients expressed the Th1-related chemokine receptor, CCR5 and CXCR3 preferentially compared with those of healthy controls. Intestinal lesions of BD expressed interferon (IFN)-gamma, tumour necrosis factor (TNF) alpha and interleukin (IL)-12 mRNA, indicating Th1 skewed responses in vivo. mRNA of Txk, a Tec family tyrosine kinase specific to Th1 cells, was expressed in the lesions, suggesting its contribution to the Th1-dominant responses. In the intestinal samples, CCR5 was detected in all the cases with BD, whereas Th2 related CCR3 and CCR4 were detected randomly, mainly in the cases with inactive BD and those receiving large amounts of prednisolone, indicating the Th1-dominant immune responses in the intestinal lesions. As the ligands of CCR5, MIP1alpha and MIP1beta were detected, whereas RANTES was not. Heat shock protein (HSP) 60 was expressed in PBL and intestinal tissues of BD. Th1-dominant immune responses and HSP60 expression may induce the inflammatory responses and thus be associated with the pathogenesis of intestinal BD. PMID- 15654839 TI - The actin-binding protein profilin I is localized at synaptic sites in an activity-regulated manner. AB - Morphological changes at synaptic specializations have been implicated in regulating synaptic strength. Actin turnover at dendritic spines is regulated by neuronal activity and contributes to spine size, shape and motility. The reorganization of actin filaments requires profilins, which stimulate actin polymerization. Neurons express two independent gene products - profilin I and profilin II. A role for profilin II in activity-dependent mechanisms at spine synapses has recently been described. Although profilin I interacts with synaptic proteins, little is known about its cellular and subcellular localization in neurons. Here, we investigated the subcellular distribution of this protein in brain neurons as well as in hippocampal cultures. Our results indicate that the expression of profilin I varies in different brain regions. Thus, in cerebral cortex and hippocampus profilin I immunostaining was associated predominantly with dendrites and was present in a subset of dendritic spines. In contrast, profilin I in cerebellum was associated primarily with presynaptic structures. Profilin I immunoreactivity was partially colocalized with the synaptic molecules synaptophysin, PSD-95 and gephyrin in cultured hippocampal neurons, indicating that profilin I is present in only a subset of synapses. At dendritic spine structures, profilin I was found primarily in protrusions, which were in apposition to presynaptic terminal boutons. Remarkably, depolarization with KCl caused a moderate but significant increase in the number of synapses containing profilin I. These results show that profilin I can be present at both pre- and postsynaptic sites and suggest a role for this actin-binding protein in activity dependent remodelling of synaptic structure. PMID- 15654841 TI - Caspase-independent retinal ganglion cell death after target ablation in the neonatal rat. AB - In neonatal rats, superior colliculus (SC) ablation results in a massive and rapid increase in retinal ganglion cell (RGC) death that peaks about 24 h post lesion (PL). Naturally occurring cell death during normal development, and RGC death after axonal injury in neonatal and adult rats, has primarily been ascribed to apoptosis. Given that normal developmental cell death is reported to involve caspase 3 activation, and blocking caspase activity in adults reduces axotomy induced death, we examined whether blocking caspases in vivo reduces RGC death after neonatal SC lesions. Neither general nor specific caspase inhibitors increased neonatal RGC survival 6 and 24 h PL. These inhibitors were, however, effective in blocking caspases in another well-defined in vitro apoptosis model, the corpus luteum. Caspase 3 protein and mRNA levels in retinas from normal and SC-lesioned neonatal rats were assessed 3, 6 and 24 h after SC removal using immunohistochemistry, western and northern blots and quantitative real-time polymerase chain reaction. Terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL) was used to independently monitor retinal cell death. The polymerase chain reaction data showed a small but insignificant increase in caspase 3 mRNA in retinas 24 h PL. Western blot analysis did not reveal a significant shift to cleaved (activated) caspase 3 protein. There was a small increase in the number of cleaved caspase 3 immunolabelled cells in the ganglion cell layer 24 h PL but this represented only a fraction of the death revealed by TUNEL. Together, these data indicate that, unlike the situation in adults, most lesion-induced RGC death in neonatal rats occurs independently of caspase activation. PMID- 15654840 TI - Dynamic reorganization of the astrocyte actin cytoskeleton elicited by cAMP and PACAP: a role for phosphatidylInositol 3-kinase inhibition. AB - Cyclic AMP (cAMP)-raising agents induce astrocytes grown in vitro to adopt a stellate morphology resembling their in vivo appearance, through the depolymerization of actomyosin stress fibres. The signalling pathways responsible for cAMP-induced astrocyte stellation have thus far remained largely elusive. We showed in this study that the neurotrophic peptide PACAP (pituitary adenylate cyclase-activating polypeptide) mimicked the effect of forskolin, a direct activator of adenylate cyclase, on the actin cytoskeleton of primary rat astrocytes. The depolymerization of stress fibres induced by PACAP or forskolin was prevented by the expression of a constitutively active mutant of RhoA, but not by a protein kinase A (PKA) blocker, indicating that cAMP-raising agents act upstream of RhoA, in a PKA-independent manner. In addition, PACAP and forskolin inhibited basal Akt phosphorylation, and basal and epidermal growth factor (EGF) stimulated phosphatidylinositol 3-kinase (PI 3-K) activities. Incubation with a PI 3-K blocker resulted in the depolymerization of stress fibres. This effect was blocked by the expression of a constitutively active mutant of RhoA, indicating that PI 3-K inhibition acted upstream of RhoA. Together, these data demonstrate for the first time that depolymerization of stress fibres, and the resulting astrocyte stellation, induced by stimulation of cAMP production involves the inhibition of the PI 3-K-RhoA pathway. PMID- 15654842 TI - Expression of the repressor element-1 silencing transcription factor (REST) is influenced by insulin-like growth factor-I in differentiating human neuroblastoma cells. AB - The repressor element-1 (RE-1) silencing transcription factor (REST) interacts with an RE-1 cis element and represses the transcription of neuron-specific genes in neuronal progenitors but is down-regulated in post-mitotic neurons. We report that REST expression is modified, in a time-dependent manner, in SH-SY5Y neuroblastoma cells exposed to insulin-like growth factor I (IGF-I), a polypeptide hormone affecting various aspects of neuronal induction and maturation. REST is increased in cells treated with IGF-I for 2 days and then declines in 5-day-treated cells concomitant with a progressive neurite extension. To investigate any role played by REST in neurodifferentiation by IGF-I, we employed an antisense oligonucleotide (AS-ODN) complementary to REST mRNA. In AS ODN-treated cells, the effects elicited by IGF-I on cell proliferation are not influenced whereas a marked decrease of REST significantly increases neurite elongation without any gross perturbation of neurogenesis. Synapsin I and betaIII tubulin gene promoters contain an RE-1 motif and their transcription is repressed by REST; both of them are increased in cells exposed to IGF-I for 5 days and further elevated by AS-ODN treatment. A parallel increase of growth cone associated protein 43, a protein chosen as a neuronal marker not directly regulated by REST, is also observed. Therefore, REST is elevated during early steps of neural induction by IGF-I and could contribute to down-regulate genes not yet required by the differentiation program while it declines later for the acquisition of neural phenotypes. These results suggest a model in which differentiating neuroblastoma cells determine their extent of neurite outgrowth on the basis of REST disappearance. PMID- 15654843 TI - Photoreceptor morphology is severely affected in the beta,beta-carotene-15,15' oxygenase (bcox) zebrafish morphant. AB - The retinoic acid molecule, a vitamin A derivative, is of key importance for eye and photoreceptor development in vertebrates. Several studies have provided evidence that the ventral part of the retina is particularly susceptible to impairment in retinoid signalling during the period of its development. In zebrafish, targeted gene knockdown of beta,beta-carotene-15,15'-oxygenase (bcox), the key enzyme for vitamin A formation, provokes a loss of retinoid signalling during early eye development that results in microphthalmia at larval stages. Using this model, we analysed the consequences of this for the retinal morphology of the fish larvae in structural details. Our analyses revealed that rods and cones do not express photoreceptor specific proteins (rhodopsin, peanut agglutinin, zpr1) in the peripheral retina. The photoreceptors in the central retina showed shortened outer segments, and electron dense debris in their intermembranal space. The number of phagosomes was increased, and cell death was frequently observed in the outer nuclear layer. Furthermore, the number of Muller cells was significantly reduced in the inner nuclear layer. Thus, we found that the lack of retinoid signalling strongly effects photoreceptor development in the ventral and dorsal retina. In addition, shortened outer segments and cell death of the remaining photoreceptors in the central retina indicate that there is an ongoing need for retinoid signalling for photoreceptor integrity and survival at later developmental stages. PMID- 15654845 TI - Deafferentation and neurotrophin-mediated intraspinal sprouting: a central role for the p75 neurotrophin receptor. AB - Axonal plasticity in the adult spinal cord is governed by intrinsic neuronal growth potential and by extracellular cues. The p75 receptor (p75(NTR)) binds growth-promoting neurotrophins (NTs) as well as the common receptor for growth inhibiting myelin-derived proteins (the Nogo receptor) and so is well situated to gauge the balance of positive and negative influences on axonal plasticity. Using transgenic mice lacking the extracellular NT-binding domain of p75(NTR) (p75-/- mice), we have examined the influence of p75(NTR) on changes in the density of primary afferent (calcitonin gene-related peptide-expressing) and descending monoaminergic (serotonin- and tyrosine hydroxylase-expressing) projections to the dorsal horn after dorsal rhizotomy, with and without concomitant application of exogenous nerve growth factor and NT-3. We found that, in intact p75-/- mice, the axon density of all populations was equal to or less than that in wild-type mice but that rhizotomy-induced intraspinal sprouting was significantly augmented. Monoaminergic axon sprouting was enhanced in both nerve growth factor- and NT-3 treated p75-/- mice compared with similarly treated wild-type mice. Primary afferent sprouting was particularly robust in NT-3-treated p75-/- mice. These in vivo results illustrate the interactions of p75(NTR) with NTs, with their respective tropomyosin-related kinase receptors and with inhibitory myelin derived molecules. Our findings illustrate the pivotal role of p75(NTR) in spinal axonal plasticity and identify it as a potential therapeutic target for spinal cord injury. PMID- 15654844 TI - Sequential activation of p75 and TrkB is involved in dendritic development of subventricular zone-derived neuronal progenitors in vitro. AB - Dendritic arbor development of subventricular zone-derived interneurons is a critical step in their integration into functional circuits of the postnatal olfactory bulb. However, the mechanism and molecular control of this process remain unknown. In this study, we have developed a culture model where dendritic development of purified subventricular zone cells proceeds under serum-free conditions in the absence of added growth factors and non-neural cells. We demonstrate that the large majority of these cells in culture express GABA and elaborate dendritic arbors with spine-like protrusions but they do not possess axons. These neurons expressed receptors for neurotrophins including p75, TrkB and TrkC but not TrkA. Application of exogenous neurotrophins, including brain derived neurotrophic factor (BDNF), neurotrophin-3 (NT3) and nerve growth factor (NGF), to cultures stimulated dendritic growth and led to more complex dendritic arbors during the initial 3 days in culture. Our results suggest that these effects are independent of Trk receptors and mediated by the p75/ceramide signaling pathway. We also show that brain-derived neurotrophic factor is the only neurotrophin that is able to influence late-phase dendritic development via TrkB receptor activation. These results suggest that dendritic arbor development of subventricular zone-derived cells may be regulated by neurotrophins through the activation of p75 and the TrkB receptor signaling pathways in a sequentially defined temporal pattern. PMID- 15654846 TI - Neocortex is the major target of sedative concentrations of volatile anaesthetics: strong depression of firing rates and increase of GABAA receptor mediated inhibition. AB - General anaesthetics cause sedation, amnesia and hypnosis. Although these clinically desired actions are indicative of an impairment of neocortical information processing, it is widely held that they are to a large part mediated by subcortical neural networks. Anaesthetic action on brain stem, basal forebrain and thalamus, all of which are known to modulate cortical excitability, would thus ultimately converge on neocortex, perturbing and reducing action potential activity therein. However, as neocortex harbours molecular targets of anaesthetics in high densities, notably GABA(A) receptors, neocortex itself should be very sensitive to anaesthetics. Here, we performed experiments to reveal the extent to which neocortex proper is a relevant target of the low concentrations of volatile anaesthetics causing sedation and hypnosis. We compared the effects of isoflurane, enflurane and halothane on spontaneous action potential activity of rat neocortical neurons in vivo and in isolated cortical networks in vitro, i.e. in the presence and absence of subcortical arousal systems. We observed that the anaesthetics decreased spontaneous firing of neurons via intracortical mechanisms; concentrations inducing hypnosis in humans reduced discharge rates both in vivo and in vitro to the same extent, approximately 50%. This decrease in neuronal activity was paralleled by a significant enhancement of neocortical GABA(A) receptor-mediated inhibition. These findings challenge the notion of predominantly subcortical effects of volatile anaesthetics and suggest that intracortical targets, among them neocortical GABA(A) receptors, mediate the sedative and hypnotic properties of volatile anaesthetics. PMID- 15654847 TI - Excitotoxic death induced by released glutamate in depolarized primary cultures of mouse cerebellar granule cells is dependent on GABAA receptors and niflumic acid-sensitive chloride channels. AB - Excitotoxic neuronal death has been linked to neurological and neurodegenerative diseases. Several studies have sought to clarify the involvement of Cl(-) channels in neuronal excitotoxicity using either N-methyl-D-aspartic acid (NMDA) or alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate/kainic acid agonists. In this work we induced excitotoxic death in primary cultures of cerebellar granule cells by means of endogenously released glutamate. Excitotoxicity was provoked by exposure to high extracellular K(+) concentrations ([K(+)](o)) for 5 min. Under these conditions, a Ca(2+)-dependent release of glutamate was evoked. When extracellular glutamate concentration rose to between 2 and 4 microM, cell viability was significantly reduced by 30-40%. The NMDA receptor antagonists (MK 801 and D-2-amino-5-phosphonopentanoic acid) prevented cell death. Exposure to high [K(+)](o) produced a (36)Cl(-) influx which was significantly reduced by picrotoxinin. In addition, the GABA(A) receptor antagonists (bicuculline, picrotoxinin and SR 95531) protected cells from high [K(+)](o)-triggered excitotoxicity and reduced extracellular glutamate concentration. The Cl(-) channel blockers niflumic acid and 5-nitro-2-(3-phenylpropylamino)benzoic acid also exerted a neuroprotective effect and reduced extracellular glutamate concentration, even though they did not reduce high [K(+)](o)-induced (36)Cl(-) influx. Primary cultures of cerebellar granule cells also contain a population of GABAergic neurons that released GABA in response to high [K(+)](o). Chronic treatment of primary cultures with kainic acid abolished GABA release and rendered granule cells insensitive to high [K(+)](o) exposure, even though NMDA receptors were functional. Altogether, these results demonstrate that, under conditions of membrane depolarization, low micromolar concentrations of extracellular glutamate might induce an excitotoxic process through both NMDA and GABA(A) receptors and niflumic acid-sensitive Cl(-) channels. PMID- 15654848 TI - GABAergic transmission in the rat paraventricular nucleus of the hypothalamus is suppressed by corticosterone and stress. AB - Parvocellular neurons in the hypothalamic paraventricular nucleus receive hormonal inputs mediated by corticosterone as well as neuronal inputs, prominent among which is a GABAergic inhibitory projection. In the present study we examined the functional properties of this GABAergic innervation when corticosteroid levels fluctuate. Frequency, amplitude and kinetic properties of miniature inhibitory postsynaptic potentials (mIPSCs), mediated by gamma amino butyric acid (GABA) were studied with whole cell recording in parvocellular neurons. Injection of a high dose of corticosterone in vivo suppressed the frequency but did not change the amplitude and kinetic properties of mIPSCs recorded 1-5 h later in vitro. Similar effects were observed after restraint stress. The corticosteroid actions do not require involvement of extrahypothalamic brain regions, because in vitro administration of 100 nM corticosterone (20 min) directly to a hypothalamic slice also suppressed the frequency of mIPSCs recorded several hours later. Corticosterone administration to hypothalamic slices from restraint rats did not result in stronger reduction of mIPSC frequency than either treatment alone, pointing to a common underlying mechanism. Paired pulse response inhibition was reduced by corticosterone, suggesting that the hormone decreases the release probability of GABA-containing vesicles. Unlike neurosteroids, corticosterone induced no rapid effects on mIPSC properties. These results indicate that increases in glucocorticoid level due to stress can slowly but persistently inhibit the GABAergic tone on parvocellular hypothalamic neurons via a hitherto unknown local mechanism independent of limbic projections. PMID- 15654849 TI - Respiratory responses evoked by blockades of ionotropic glutamate receptors within the Botzinger complex and the pre-Botzinger complex of the rabbit. AB - The respiratory role of excitatory amino acid (EAA) receptors within the Botzinger complex (BotC) and the pre-Botzinger complex (pre-BotC) was investigated in alpha-chloralose-urethane anaesthetized, vagotomized, paralysed and artificially ventilated rabbits by using bilateral microinjections (30-50 nL) of EAA receptor antagonists. Blockade of both N-methyl-D-aspartic acid (NMDA) and non-NMDA receptors by 50 mM kynurenic acid (KYN) within the BotC induced a pattern of breathing characterized by low-amplitude, high-frequency irregular oscillations superimposed on tonic phrenic activity and successively the disappearance of respiratory rhythmicity in the presence of intense tonic inspiratory discharges (tonic apnea). KYN microinjections into the pre-BotC caused similar respiratory responses that, however, never led to tonic apnea. Blockade of NMDA receptors by D(-)-2-amino-5-phosphonopentanoic acid (D-AP5; 1, 10 and 20 mM) within the BotC induced increases in respiratory frequency and decreases in peak phrenic amplitude; the highest concentrations caused tonic apnea insensitive to chemical stimuli. Blockade of non-NMDA receptors by 6-cyano 7-nitroquinoxaline-2,3-dione (CNQX; 1, 10 and 20 mM) within the BotC produced only less pronounced increases in respiratory frequency. Responses to D-AP5 in the pre-BotC were similar, although less pronounced than those elicited in the BotC and never characterized by tonic apnea. In the same region, CNQX provoked increases in respiratory frequency similar to those elicited in the BotC, associated with slight reductions in peak phrenic activity. The results show that EAA receptors within the investigated medullary subregions mediate a potent control on both the intensity and frequency of inspiratory activity, with a major role played by NMDA receptors. PMID- 15654850 TI - Inflammation alters somatostatin mRNA expression in sensory neurons in the rat. AB - Proinflammatory neuropeptides, such as substance P and calcitonin gene-related peptide, are up-regulated in primary afferent neurons in acute and chronic inflammation. While these neuropeptides have been intensively studied, potentially anti-inflammatory and/or anti-nociceptive neuropeptides such as somatostatin (SS) have been less widely investigated. Endogenous somatostatin is thought to exert a tonic antinociceptive effect. Exogenous SS is anti inflammatory and antinociceptive and is thought to exert these actions through inhibition of proinflammatory neuropeptide release. In this study we have compared the expression of somatostatin in two inflammatory models: arthritis, a condition associated with increased nociception, and periodontitis, in which there is little evidence of altered nociceptive thresholds. In acute arthritis (< 24 h) SS mRNA was down-regulated in ipsilateral dorsal root ganglia (DRG; 52 +/- 7% of control, P < 0.05), and up-regulated in contralateral DRG (134 +/- 10% of control; P < 0.05). In chronic arthritis (14 days) this pattern of mRNA regulation was reversed, with SS being up-regulated ipsilaterally and down regulated contralaterally. In chronic mandibular periodontitis (7-10 days), SS mRNA was up-regulated in only the mandibular division of the ipsilateral trigeminal ganglion (TG) (day 7, 219 +/- 9% and day 10, 217 +/- 12% of control; P < 0.02) but showed no change in other divisions of the trigeminal ganglion or in the mesencephalic nucleus. These data show that antinociceptive and anti inflammatory neuropeptides are also regulated in inflammation. It is possible that the degree of inflammation and nociception seen may depend on the balance of pro- and anti-inflammatory and nociceptive peptide expression in a particular condition. PMID- 15654851 TI - Depolarization evokes different patterns of calcium signals and exocytosis in bovine and mouse chromaffin cells: the role of mitochondria. AB - This study was planned on the assumptions that different high-voltage activated calcium channels and/or the ability of mitochondria to take up Ca(2+) could be responsible for different cytosolic Ca(2+) concentrations ([Ca(2+)](c)) and catecholamine release responses in adrenal chromaffin cells of bovine and mouse species. Short K(+) pulses (2-5 s, 70 mM K(+)) increased [Ca(2+)](c) to a peak of about 1 microM; however, in bovine cells the decline was slower than in mouse cells. Secretory responses were faster in mouse but were otherwise quantitatively similar. Upon longer K(+) applications (1 min), elevations of [Ca(2+)](c) and secretion were prolonged in bovine cells; in contrast [Ca(2+)](c) in mouse cells declined three-fold faster and failed to sustain a continued secretion. Confocal [Ca(2+)](c) imaging following a 50-ms depolarizing pulse showed a similar Ca(2+) entry, but a rate of [Ca(2+)](c) increase and a maximum peak significantly higher in bovine cells; the rate of dissipation of the Ca(2+) wave was faster in the mouse. The mitochondrial protonophore CCCP (2 microm) halved the K(+)-evoked [Ca(2+)](c) and secretory signals in mouse cells, but had little affect on bovine responses. We conclude that the relative densities of L (15% in bovine and 50% in mouse) and P/Q Ca(2+) channels (50% in bovine and 15% in mouse) do not contribute to the observed differences; rather, the different intracellular distribution of Ca(2+), which is strongly influenced by mitochondria, is responsible for a more sustained secretory response in bovine, and for a faster and more transient secretory response in mouse chromaffin cells. It seems that mitochondria near the plasmalemma sequester Ca(2+) more rapidly and efficiently in the mouse than in the bovine chromaffin cell. PMID- 15654852 TI - Secretion of ATP from Schwann cells in response to uridine triphosphate. AB - The mechanisms by which uridine triphosphate (UTP) stimulates ATP release from Schwann cells cultured from the sciatic nerve were investigated using online bioluminescence techniques. UTP, a P2Y(2) and P2Y(4) receptor agonist, stimulated ATP release from Schwann cells in a dose-dependent manner with an ED(50) of 0.24 microm. UTP-stimulated ATP release occurs through P2Y(2) receptors as it was blocked by suramin which inhibits P2Y(2) but not P2Y(4) receptors. Furthermore, positive immunostaining of P2Y(2) receptors on Schwann cells was revealed and GTP, an equipotent agonist with UTP at rat P2Y(4) receptors, did not significantly stimulate ATP release. UTP-stimulated ATP release involved second messenger pathways as it was attenuated by the phospholipase C inhibitor U73122, the protein kinase C inhibitor chelerytherine chloride, the IP(3) formation inhibitor lithium chloride, the cell membrane-permeable Ca(2+) chelator BAPTA-AM and the endoplasmic reticulum Ca(2+)-dependent ATPase inhibitor thapsigargin. Evidence that ATP may be stored in vesicles that must be transported to the cell membrane for exocytosis was found as release was significantly reduced by the Golgi-complex inhibitor brefeldin A, microtubule disruption with nocodazole, F actin disruption with cytochalasin D and the specific exocytosis inhibitor botulinum toxin A. ATP release from Schwann cells also involves anion transport as it was significantly reduced by cystic fibrosis transmembrane conductance regulator inhibitor glibencamide and anion transporter inhibitor furosemide. We suggest that UTP-stimulated ATP release is mediated by activation of P2Y(2) receptors that initiate an IP(3)-Ca(2+) cascade and protein kinase C which promote exocytosis of ATP from vesicles as well as anion transport of ATP across the cell membrane. PMID- 15654853 TI - Transsynaptic virus tracing from host brain to subretinal transplants. AB - The aim of this study was to establish synapses between a transplant and a degenerated retina. To tackle this difficult task, a little-known but well established CNS method was chosen: trans-synaptic pseudorabies virus (PRV) tracing. Sheets of E19 rat retina with or without retinal pigment epithelium (RPE) were transplanted to the subretinal space in 33 Royal College of Surgeons (RCS) and transgenic s334ter-5 rats with retinal degeneration. Several months later, PRV-BaBlu (expressing E. colibeta-galactosidase) or PRV-Bartha was injected into an area of the exposed superior colliculus (SC), topographically corresponding to the transplant placement in the retina. Twenty normal rats served as controls. After survival times of 1-5 days, retinas were examined for virus by X-gal histochemistry, immunohistochemistry and electron microscopy. In normal controls, virus was first seen in retinal ganglion cells and Muller glia after 1-1.5 days, and had spread to all retinal layers after 2-3 days. Virus labeled cells were found in 16 of 19 transplants where the virus injection had retrogradely labeled the topographically correct transplant area of the host retina. Electron microscopically, enveloped and nonenveloped virus could clearly be detected in infected cells. Enveloped virus was found only in neurons. Infected glial cells contained only nonenveloped virus. Neurons in retinal transplants are labeled after PRV injection into the host brain, indicating synaptic connectivity between transplants and degenerated host retinas. This study provides evidence that PRV spreads in the retina as in other parts of the CNS and is useful to outline transplant-host circuitry. PMID- 15654854 TI - Development of spontaneous mouth/tongue movement and related neural activity, and their repression in fetal mice lacking glutamate decarboxylase 67. AB - Spontaneous body movement starts at early fetal stage, at embryonic day (E) 12-15 in mice. In the present study, the movement of the head region was studied in E13 14 mice by in utero ultrasound imaging, together with the in vitro recording of underlying neural activities in the hypoglossal nerve and the ventral root of the upper cervical cord of an isolated brainstem-spinal cord preparation. The role of gamma-aminobutyric acid (GABA) in the generation of fetal movement was assessed using mice lacking GABA-synthesizing glutamate decarboxylase 67 (GAD67). At E14, mouth opening and tongue withdrawal were observed independently at frequency of 14/h each. This movement was rarely observed in the GAD67-deficient mouse. The intraventricular administration of picrotoxin or 3-mercaptopropionic acid abolished mouth opening in the wild-type mice. In a brainstem-spinal cord preparation, three types of neural discharge were recorded: mouth/tongue-moving burst, respiratory burst and irregular activity on the basis of their waveform, regularity in occurrence and concomitant muscle activity. In the GAD67-deficient mice, the occurrence of mouth/tongue-moving burst and irregular activity was inhibited to about 15 and 40% of those in the wild-type mice, respectively. Respiratory burst was slightly inhibited but the difference was not significant. Picrotoxin greatly reduced the frequency of mouth/tongue-moving burst. These results indicate that GABA is involved in rhythm generation in movement of the head region and support the hypothesis that cleft palate in the GAD67-deficient mouse is due to the impairment of mouth or tongue movement that assists palate formation. PMID- 15654855 TI - Repeated administration of the monoamine reuptake inhibitor BTS 74 398 induces ipsilateral circling in the 6-hydroxydopamine lesioned rat without sensitizing motor behaviours. AB - BTS 74 398 (1-[1-(3,4-dichlorophenyl)cyclobutyl]-2-(3 diaminethylaminopropylthio)ethanone monocitrate) is a monoamine reuptake inhibitor that reverses motor deficits in MPTP-treated (1-methyl-4-phenyl-1,2,3,6 tetrahydropyridine) common marmosets without provoking established dyskinesia. However, it is not known whether BTS 74 398 primes the basal ganglia for dyskinesia induction. In this study, the ability of BTS 74 398 to sensitize 6 hydroxydopamine (6-OHDA)-lesioned rats for the production of abnormal motor behaviours and the induction of striatal DeltaFosB were determined in comparison with l-3,4-dihydroxyphenylalanine methyl ester (L-dopa). Acute administration of BTS 74 398 induced a dose-dependent ipsilateral circling response in unilaterally 6-OHDA-lesioned rats whereas L-dopa produced dose-dependent contraversive rotation. The ipsilateral circling response to BTS 74 398 did not alter during 21 days of administration. In contrast, L-dopa treatment for 21 days caused a marked increase in rotational response. Repeated administration of both L-dopa and BTS 74 398 increased general motor activity and stereotypic behaviour. In L-dopa treated rats, orolingual, locomotive, forelimb and axial abnormal movements developed whereas BTS 74 398 produced only locomotion with a side bias but no other abnormal movements. Sensitization of circling responses and the development of abnormal movements in 6-OHDA-lesioned rats have been associated with the potential of dopaminergic drugs to induce dyskinesia. Furthermore, striatal DeltaFosB immunoreactivity, shown to correlate with dyskinesia induction, was increased by L-dopa but was unaffected by repeated BTS 74 398 administration. The lack of such changes following repeated BTS 74 398 treatment suggests that it may be an effective antiparkinsonian therapy that is unlikely to produce involuntary movements. PMID- 15654856 TI - Matrix metalloproteinase (MMP)-12 expression has a negative impact on sensorimotor function following intracerebral haemorrhage in mice. AB - We investigated the role of matrix metalloproteinases (MMPs) in a mouse model of intracerebral haemorrhage (ICH). Transcripts encoding nine of the 23 known mammalian MMPs were measured. MMP-12 levels were the most elevated. To evaluate the role of MMP-12 in ICH, haemorrhages were induced in wild-type (WT) and MMP-12 null mice. The results show that MMP-12 null mice exhibited significant functional recovery of forelimb reaching and reduced dependence on the ipsilateral forelimb compared to WT mice. There was also a trend for improved sensory function in the tape removal test. With respect to single pellet skilled reaching, MMP-12 null mice recovered to a level that was not significantly different from sham at 14 and 28 days post-ICH. In contrast, WT animals demonstrated a persistent impairment relative to sham controls throughout the survival period (P < 0.05). The cylinder task revealed a lesion-induced reliance on the ipsilateral forelimb that was apparent at day 7 in both MMP-12 null and WT mice (P < 0.05), but only persisted in WT mice at 14 days post-ICH (P < 0.05). Differences in functional outcome could not be explained by tissue sparing. However, Iba1 immunostaining indicated that more cells bearing macrophage morphology were recruited to the lesion area in WT mice. This is the first study to profile the expression patterns of a number of the known MMPs following ICH in mice. The data indicate that MMP-12 expression following haemorrhagic stroke is deleterious and contributes to the development of secondary injury in this disease. PMID- 15654857 TI - Presynaptic NMDA autoreceptors facilitate axon excitability: a new molecular target for the anticonvulsant gabapentin. AB - Gabapentin is a drug with anticonvulsant and analgesic properties causing the reduction of neurotransmitter release. We show that one of the mechanisms implicated in this effect of gabapentin is the reduction of the axon excitability measured as an amplitude change of the presynaptic fibre volley (FV) in the CA1 area of rat hippocampal slices. Interestingly, we found that gabapentin-induced depression of FV is mimicked and occluded by NMDA receptor (NMDA-R) antagonists, indicating that these receptors are located presynaptically and are activated by ambient levels of glutamate. Conversely, NMDA application (20 microM, 10 min) elicits a reversible FV potentiation which is reduced by gabapentin. Both NMDA- and gabapentin-induced FV changes are partially explained by modifications in the firing threshold of individual fibres. Increasing [K(+)](o) does not mimic or occlude (at a concentration of 6.5 mM) the effect of NMDA on FV amplitude, which makes it unlikely that a rise in [K(+)](o) induced by NMDA receptor activation could indirectly participate in the potentiation of the FV. The NMDA-induced FV potentiation is independent of extracellular calcium presence but is completely inhibited in a low-Na(+) solution (50% reduction) or under NMDA channel block (high Mg(2+) or MK 801). These findings suggest that sodium entry through presynaptic NMDA-R channels facilitates axon excitability. The interaction of gabapentin with this newly described mechanism might contribute to its therapeutic benefits. PMID- 15654858 TI - Inhibitory avoidance learning altered ensemble activity of amygdaloid neurons in rats. AB - In this study, we examined single-unit activity in the amygdala before and after a rat had acquired an inhibitory avoidance task. Long-Evans rats with microwires chronically implanted into the central nucleus (CeA) or basolateral complex (BLC) of the amygdala were acclimatized to the apparatus of a step-through inhibitory avoidance task for three sessions. On the fourth session, rats in the experimental group received an inescapable footshock (3 mA, 1 s) as they stepped from the lit side into the dark side of the task apparatus, whereas rats in the control group received the same amount of shock on a different apparatus. All rats were tested for retention in the task apparatus 1 day after shock training. The experimental rats showed better retention than the controls as they stayed longer in the lit side. Ensemble unit activities were recorded in the amygdala nuclei from the indwelling wire bundles during the acclimation and test sessions. The data collected from well-isolated amygdala units showed that neuronal discharge habituated from the first to the third acclimation session. In the test session, the experimental group, but not the control group, showed elevated firing rates in the CeA or BLC neurons located on either side of the brain. These findings provide the first piece of evidence showing that learning of an inhibitory avoidance task leads to an increase in amygdala neuronal discharges during a retention test. PMID- 15654859 TI - Genetic ablation of the mammillary bodies in the Foxb1 mutant mouse leads to selective deficit of spatial working memory. AB - Mammillary bodies and the mammillothalamic tract are parts of a classic neural circuitry that has been implicated in severe memory disturbances accompanying Korsakoff's syndrome. However, the specific role of mammillary bodies in memory functions remains controversial, often being considered as just an extension of the hippocampal memory system. To study this issue we used mutant mice with a targeted mutation in the transcription factor gene Foxb1. These mice suffer perinatal degeneration of the medial and most of the lateral mammillary nuclei, as well as of the mammillothalamic bundle. Foxb1 mutant mice showed no deficits in such hippocampal-dependent tasks as contextual fear conditioning and social transmission of food preference. They were also not impaired in the spatial reference memory test in the radial arm maze. However, Foxb1 mutants showed deficits in the task for spatial navigation within the Barnes maze. Furthermore, they showed impairments in spatial working memory tasks such as the spontaneous alternation and the working memory test in the radial arm maze. Thus, our behavioural analysis of Foxb1 mutants suggests that the medial mammillary nuclei and mammillothalamic tract play a role in a specific subset of spatial tasks, which require combined use of both spatial and working memory functions. Therefore, the mammillary bodies and the mammillothalamic tract may form an important route through which the working memory circuitry receives spatial information from the hippocampus. PMID- 15654860 TI - Hierarchical processing of sound location and motion in the human brainstem and planum temporale. AB - Horizontal sound localization relies on the extraction of binaural acoustic cues by integration of the signals from the two ears at the level of the brainstem. The present experiment was aimed at detecting the sites of binaural integration in the human brainstem using functional magnetic resonance imaging and a binaural difference paradigm, in which the responses to binaural sounds were compared with the sum of the responses to the corresponding monaural sounds. The experiment also included a moving sound condition, which was contrasted against a spectrally and energetically matched stationary sound condition to assess which of the structures that are involved in general binaural processing are specifically specialized in motion processing. The binaural difference contrast revealed a substantial binaural response suppression in the inferior colliculus in the midbrain, the medial geniculate body in the thalamus and the primary auditory cortex. The effect appears to reflect an actual reduction of the underlying activity, probably brought about by binaural inhibition or refractoriness at the level of the superior olivary complex. Whereas all structures up to and including the primary auditory cortex were activated as strongly by the stationary as by the moving sounds, non-primary auditory fields in the planum temporale responded selectively to the moving sounds. These results suggest a hierarchical organization of auditory spatial processing in which the general analysis of binaural information begins as early as the brainstem, while the representation of dynamic binaural cues relies on non-primary auditory fields in the planum temporale. PMID- 15654861 TI - Differential progression of proprioceptive and visual information processing deficits in Parkinson's disease. AB - Indirect evidence suggests that patients with Parkinson's disease (PD) have deficits not only in motor performance, but also in the processing of sensory information. We investigated the role of sensory information processing in PD patients with a broad range of disease severities and in a group of age-matched controls. Subjects were tested in two conditions: pointing to a remembered visual target in complete darkness (DARK) and in the presence of an illuminated frame with a light attached to the index finger (FRAME). Differences in pointing errors in these two conditions reflect the effect of visual feedback on pointing. PD patients showed significantly larger constant and variable errors than controls in the DARK and FRAME condition. The difference of the variable error in the FRAME and DARK condition decreased as a function of the severity of PD. This indicates that any deficits in the processing of proprioceptive information occur already at very mild symptoms of PD, and that deficits in the use of visual feedback develop progressively in later stages of the disease. These results provide a tool for early diagnosis of PD and shed new light on the functional role of the brain structures that are affected in PD. PMID- 15654863 TI - Motor cortex plasticity induced by extensive training revealed by transcranial magnetic stimulation in human. AB - This study examines the effect of high-level skilled behaviour on motor cortex representations of upper extremity muscles of ten sportswomen. We used transcranial magnetic stimulation to map proximal medial deltoid and distal extensor carpi radialis muscle representations on both hemispheres during low level voluntary contraction. We compared cortical representation areas between two groups of subjects and between hemispheres within subjects. The first group comprised five elite volleyball attackers and the second group five runners. Four stimuli were delivered on multiple scalp sites (1.5 cm apart) to induce motor evoked potentials recorded by surface EMG. Maps were described in terms of excitable scalp positions and of motor-evoked potentials. We observed differences in map areas between the two groups. Volleyball players had larger cortical representations of the proximal medial deltoid muscle than runners. Furthermore, the volleyball players had larger map areas for dominant muscles compared with non-dominant muscles. There was no difference, however, in map area for either muscle between the dominant and non-dominant arm in the runner group. Our results show that heavy training in a specific skill induces an expansion of proximal muscle representation in the contralateral primary motor cortex. This enlarged map area for proximal muscle is accompanied by an increase in the overlapping of proximal and distal muscle representations. This could reflect the fact that motor learning of co-ordinated movement involves a common control of both muscles. This reorganization supports the hypothesis of a cortical plasticity driven by activity. PMID- 15654862 TI - The relationship between oscillatory activity and motor reaction time in the parkinsonian subthalamic nucleus. AB - Averaging techniques have demonstrated that movement preparatory cues and movement itself are associated with marked reductions in the oscillatory synchrony of local neuronal populations in the area of the human parkinsonian subthalamic nucleus (STN), as indexed by 8-30 Hz local field potential (LFP) activity. In order to examine the detailed nature and strength of the relationship between reductions in oscillatory activity and movement we examined single-trial LFP activity recorded from the STN area of parkinsonian subjects engaged in a choice reaction task. In this task an initial warning cue was either fully predictive or non-predictive of the hand required to make a later motor response. This motor response was elicited by a second go cue to which data were aligned. We observed a significant linear relationship between the onset time of oscillation reduction after go cues and subsequent motor response time across single trials within subjects. Consistent with this observation we also found a positive correlation of power with response time following go cues. In addition, we observed shorter durations of suppression in fully predictive trials where selection of the response could precede go cue presentation. The results are consistent with the hypothesis that reductions in 8-30 Hz population synchrony in the STN area are related to the processing required for motor preparation, particularly response selection. PMID- 15654864 TI - The astrocytic barrier to axonal regeneration at the dorsal root entry zone is induced by rhizotomy. AB - After dorsal rhizotomy, sensory axons fail to regenerate beyond the astrocytic glia limitans at the dorsal root entry zone (DREZ) but this inhibition can be overcome with the delivery of exogenous neurotrophin-3. We investigated whether axonal inhibition at the DREZ is constitutive or induced after dorsal rhizotomy. Primary afferent neurones from enhanced green fluorescent protein-expressing mice were transplanted into adult rat dorsal root ganglia in the presence or absence of dorsal rhizotomy. In the absence of dorsal rhizotomy mouse axons freely extended into the rat central nervous system. After host dorsal rhizotomy, mouse axons were unable to cross the DREZ. However, in rats that received a dorsal rhizotomy concomitant with intrathecal neurotrophin-3, the mouse axons were able to cross the DREZ. These results indicate that, under normal circumstances, the adult DREZ is permissive to the regeneration of adult sensory axons and that it only becomes inhibitory once dorsal root axons have been injured and astrocytes at the DREZ have become reactive. PMID- 15654865 TI - A rat model of slow Wallerian degeneration (WldS) with improved preservation of neuromuscular synapses. AB - The slow Wallerian degeneration phenotype, Wld(S), which delays Wallerian degeneration and axon pathology for several weeks, has so far been studied only in mice. A rat model would have several advantages. First, rats model some human disorders better than mice. Second, the larger body size of rats facilitates more complex surgical manipulations. Third, rats provide a greater yield of tissue for primary culture and biochemical investigations. We generated transgenic Wld(S) rats expressing the Ube4b/Nmnat1 chimeric gene in the central and peripheral nervous system. As in Wld(S) mice, their axons survive up to 3 weeks after transection and remain functional for at least 1 week. Protection of axotomized nerve terminals is stronger than in mice, particularly in one line, where 95-100% of neuromuscular junctions remained intact and functional after 5 days. Furthermore, the loss of synaptic phenotype with age was much less in rats than in mice. Thus, the slow Wallerian degeneration phenotype can be transferred to another mammalian species and synapses may be more effectively preserved after axotomy in species with longer axons. PMID- 15654866 TI - A unified model of presynaptic release site gating by calcium channel domains. AB - Calcium ions enter through discrete ion channels at presynaptic nerve terminals before binding to and activating transmitter release sites. Opposing models hold that release sites are gated either by calcium domains of single, closely associated channels or by extensive, overlapping domains from many remote channels. At the chick calyx synapse we find a linear relation between transmitter release and the number of open calcium channels, favouring single domain activation. This finding is consistent with results from the squid giant synapse but contrasts with steep power dependences reported in rodent synapses, suggestive of activation by extensive overlapping domains. These different reports were reconciled by plotting 'per cent domain overlap' against the external calcium concentration used for each species. This relationship predicts the involvement of local channels in the activation of release sites in all species. Further, it suggests that each release site is activated by calcium ions from its immediately associated channels and not by ions that enter through channels associated with a neighbouring release site. PMID- 15654867 TI - Activation of extracellular signal-regulated kinase- mitogen-activated protein kinase cascade in the amygdala is required for memory reconsolidation of auditory fear conditioning. AB - Consolidation of new fear memories has been shown to require de novo RNA and protein synthesis in the lateral nucleus of amygdala (LA). Recently we have demonstrated that consolidated fear memories, when reactivated, return to a labile state which is sensitive to disruption by the protein synthesis inhibitor anisomycin. The specific molecular mechanisms that underlie this reconsolidation of fear memories are still largely unknown. The activation of extracellular signal-regulated kinase-mitogen-activated protein kinase (ERK-MAPK) pathway in the LA is required for the consolidation of auditory fear memories. In the present study, we examined the role of ERK-MAPK cascade in the LA during reconsolidation of auditory fear conditioning. We show that intra-LA infusions of the MAPK kinase (MEK) inhibitor U0126, a manipulation which inhibits activation of ERK-MAPK, impairs postreactivation long-term memory (PR-LTM) but leaves the postreactivation short-term memory (PR-STM) intact. The same treatment with U0126, in the absence of memory reactivation, has no effect. Furthermore, we verified that reconsolidation requires translation using a second protein synthesis inhibitor, cycloheximide. Post-reactivation infusions of cycloheximide blocked PR-LTM but not PR-STM and, in the absence of reactivation, had no effect. Our data show that activation of ERK-MAPK signalling pathway and protein synthesis in the LA are required for reconsolidation of auditory fear memories. PMID- 15654868 TI - Dopaminergic control of local interneuron activity in the thalamus. AB - The dorsal lateral geniculate nucleus (dLGN), the major thalamic station of the visual pathway, contains a fairly large number of dopaminergic terminals, and dopamine was found to reduce spontaneous and visually evoked activity in the dLGN in vivo. The cellular basis of this influence remained unknown. Here we have used whole cell patch-clamp techniques to analyse the effects of dopamine (DA) on GABAergic transmission in dLGN slices of juvenile postnatal day (P) 12-P24 Long Evans rats or juvenile (P12-P22) GAD67-GFP (Deltaneo) mice. Spontaneous inhibitory postsynaptic currents (sIPSCs) were increased in frequency by the D(2) like agonist quinpirole (QUIN) in rat (n = 6), as well as in mouse (n = 5) thalamic slices. This effect was blocked in the presence of the D(2)-like antagonist sulpiride (SULP, n = 5) and was absent in the ventrobasal complex (VB) of rat (n = 7) and mouse (n = 4) thalamus, which is devoid of GABAergic interneurons. Direct recordings from labelled GABAergic neurons in the dLGN of GAD67-GFP mice revealed a QUIN-mediated membrane depolarization (n = 12), which was attenuated by SULP (n = 6). These data demonstrate that DA through activation of D(2)-like receptors in GABAergic interneurons induces an increase in inhibitory interactions most likely at F2 dendrodendritic terminals, thereby providing a cellular correlate of the observation made in vivo that DA predominantly acts through inhibition of relay cell activity in the dLGN. PMID- 15654869 TI - Activation of group II mGlu receptors blocks the enhanced drug taking induced by previous exposure to amphetamine. AB - Repeated exposure to amphetamine (AMPH) leads to the development of behavioural sensitization that can be demonstrated in rats as enhanced locomotor responding to and self-administration of the drug. Glutamate systems are known to participate in the induction and expression of sensitization by psychostimulants. Group II metabotropic glutamate receptors (mGluRs), because they negatively regulate both vesicular and nonvesicular glutamate release, are thus well positioned to gate its expression. Here we report that the expression of locomotor sensitization by AMPH is completely prevented by a systemic injection of the selective group II mGluR agonist LY379268 at a dose that produced no effects when administered alone. The activation of group II mGluRs in AMPH sensitized rats also reduced the enhanced overflow of both dopamine and glutamate normally observed in the nucleus accumbens, a brain region critical for the generation of locomotor and drug self-administration behaviours. To directly determine the effect of group II mGluR activation on enhanced drug self administration, AMPH-sensitized rats were allowed to self-administer a mixture of LY379268 and AMPH. These rats continued to self-administer but did not exhibit the enhanced work output and drug intake observed in AMPH-sensitized rats self administering AMPH alone. Thus, activating group II mGluRs prevents the expression of different manifestations of AMPH sensitization including enhanced self-administration of the drug. These receptors may represent a potentially important target for therapeutic intervention directed at drugs of abuse. PMID- 15654870 TI - Data-driven docking for the study of biomolecular complexes. AB - With the amount of genetic information available, a lot of attention has focused on systems biology, in particular biomolecular interactions. Considering the huge number of such interactions, and their often weak and transient nature, conventional experimental methods such as X-ray crystallography and NMR spectroscopy are not sufficient to gain structural insight into these. A wealth of biochemical and/or biophysical data can, however, readily be obtained for biomolecular complexes. Combining these data with docking (the process of modeling the 3D structure of a complex from its known constituents) should provide valuable structural information and complement the classical structural methods. In this review we discuss and illustrate the various sources of data that can be used to map interactions and their combination with docking methods to generate structural models of the complexes. Finally a perspective on the future of this kind of approach is given. PMID- 15654871 TI - Defining the Q-site of Escherichia coli fumarate reductase by site-directed mutagenesis, fluorescence quench titrations and EPR spectroscopy. AB - We have used fluorescence quench titrations, EPR spectroscopy and steady-state kinetics to study the effects of site-directed mutants of FrdB, FrdC and FrdD on the proximal menaquinol (MQH(2)) binding site (Q(P)) of Escherichia coli fumarate reductase (FrdABCD) in cytoplasmic membrane preparations. Fluorescence quench (FQ) titrations with the fluorophore and MQH(2) analog 2-n-heptyl-4 hydroxyquinoline-N-oxide (HOQNO) indicate that the Q(P) site is defined by residues from FrdB, FrdC and FrdD. In FQ titrations, wild-type FrdABCD binds HOQNO with an apparent K(d) of 2.5 nM, and the following mutations significantly increase this value: FrdB-T205H (K(d) = 39 nM); FrdB-V207C (K(d) = 20 nM); FrdC E29L (K(d) = 25 nM); FrdC-W86R (no detectable binding); and FrdD-H80K (K(d) = 20 nM). In all titrations performed, data were fitted to a monophasic binding equation, indicating that no additional high-affinity HOQNO binding sites exist in FrdABCD. In all cases where HOQNO binding is detectable by FQ titration, it can also be observed by EPR spectroscopy. Steady-state kinetic studies of fumarate-dependent quinol oxidation indicate that there is a correlation between effects on HOQNO binding and effects on the observed K(m) and k(cat) values, except in the FrdC-E29L mutant, in which HOQNO binding is observed, but no enzyme turnover is detected. In this case, EPR studies indicate that the lack of activity arises because the enzyme can only remove one electron from reduced MQH(2), resulting in it being trapped in a form with a bound menasemiquinone radical anion. Overall, the data support a model for FrdABCD in which there is a single redox-active and dissociable Q-site. PMID- 15654872 TI - Physicochemical characterization of carboxymethyl lipid A derivatives in relation to biological activity. AB - Lipopolysaccharide (LPS) from the outer membrane of Gram-negative bacteria belongs to the most potent activators of the mammalian immune system. Its lipid moiety, lipid A, the 'endotoxic principle' of LPS, carries two negatively charged phosphate groups and six acyl chain residues in a defined asymmetric distribution (corresponding to synthetic compound 506). Tetraacyl lipid A (precursor IVa or synthetic 406), which lacks the two hydroxylated acyl chains, is agonistically completely inactive, but is a strong antagonist to bioactive LPS when administered to the cells before LPS addition. The two negative charges of lipid A, represented by the two phosphate groups, are essential for agonistic as well as for antagonistic activity and no highly active lipid A are known with negative charges other than phosphate groups. We hypothesized that the phosphate groups could be substituted by other negatively charged groups without changing the endotoxic properties of lipid A. To test this hypothesis, we synthesized carboxymethyl (CM) derivatives of hexaacyl lipid A (CM-506 and Bis-CM-506) and of tetraacyl lipid A (Bis-CM-406) and correlated their physicochemical with their endotoxic properties. We found that, similarly to compounds 506 and 406, also for their carboxymethyl derivatives a particular molecular ('endotoxic') conformation and with that, a particular aggregate structure is a prerequisite for high cytokine-inducing capacity and antagonistic activity, respectively. In other parameters such as acyl chain melting behaviour, antibody binding, activity in the Limulus lysate assay, and partially the binding of 3-deoxy-D-manno-oct-2 ulosonic acid transferase, strong deviations from the properties of the phosphorylated compounds were observed. These data allow a better understanding of endotoxic activity and its structural prerequisites. PMID- 15654874 TI - Role of Tyr84 in controlling the reactivity of Cys34 of human albumin. AB - Cys34 in domain I of the three-domain serum protein albumin is the binding site for a wide variety of biologically and clinically important small molecules, provides antioxidant activity, and constitutes the largest portion of free thiol in blood. Analysis of X-ray structures of albumin reveals that the loop containing Tyr84 occurs in multiple conformations. In structures where the loop is well defined, there appears to be an H-bond between the OH of Tyr84 and the sulfur of Cys34. We show that the reaction of 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) with Tyr84Phe mutant albumin is approximately four times faster than with the wild-type protein between pH 6 and pH 8. In contrast, the His39Leu mutant reacts with DTNB more slowly than the wild-type protein at pH < 8, but at a similar rate at pH 8. Above pH 8 there is a dramatic increase in reactivity for the Tyr84Phe mutant. We also report (1)H NMR studies of disulfide interchange reactions with cysteine. The tethering of the two loops containing Tyr84 and Cys34 not only appears to control the redox potential and accessibility of Cys34, but also triggers the transmission of information about the state of Cys34 throughout domain I, and to the domainI/II interface. PMID- 15654873 TI - Stability of the major allergen Brazil nut 2S albumin (Ber e 1) to physiologically relevant in vitro gastrointestinal digestion. AB - The major 2S albumin allergen from Brazil nuts, Ber e 1, was subjected to gastrointestinal digestion using a physiologically relevant in vitro model system either before or after heating (100 degrees C for 20 min). Whilst the albumin was cleaved into peptides, these were held together in a much larger structure even when digested by using a simulated phase 1 (gastric) followed by a phase 2 (duodenal) digestion system. Neither prior heating of Ber e 1 nor the presence of the physiological surfactant phosphatidylcholine affected the pattern of proteolysis. After 2 h of gastric digestion, approximately 25% of the allergen remained intact, approximately 50% corresponded to a large fragment of M(r) 6400, and the remainder comprised smaller peptides. During duodenal digestion, residual intact 2S albumin disappeared quickly, but a modified form of the 'large fragment' remained, even after 2 h of digestion, with a mass of approximately 5000 Da. The 'large fragment' comprised several smaller peptides that were identified, by using different MS techniques, as deriving from the large subunit. In particular, sequences corresponding to the hypervariable region (Q37-M47) and to another peptide (P42-P69), spanning the main immunoglobulin E epitope region of 2S albumin allergens, were found to be largely intact following phase 1 (gastric) digestion. They also contained previously identified putative T-cell epitopes. These findings indicate that the characteristic conserved skeleton of cysteine residues of 2S albumin family and, particularly, the intrachain disulphide bond pattern of the large subunit, play a critical role in holding the core protein structure together even after extensive proteolysis, and the resulting structures still contain potentially active B- and T-cell epitopes. PMID- 15654875 TI - A single mutation in Escherichia coli ribonuclease II inactivates the enzyme without affecting RNA binding. AB - Exoribonuclease II (RNase II), encoded by the rnb gene, is a ubiquitous enzyme that is responsible for 90% of the hydrolytic activity in Escherichia coli crude extracts. The E. coli strain SK4803, carrying the mutant allele rnb296, has been widely used in the study of the role of RNase II. We determined the DNA sequence of rnb296 and cloned this mutant gene in an expression vector. Only a point mutation in the coding sequence of the gene was detected, which results in the single substitution of aspartate 209 for asparagine. The mutant and the wild-type RNase II enzymes were purified, and their 3' to 5' exoribonucleolytic activity, as well as their RNA binding capability, were characterized. We also studied the metal dependency of the exoribonuclease activity of RNase II. The results obtained demonstrated that aspartate 209 is absolutely essential for RNA hydrolysis, but is not required for substrate binding. This is the first evidence of an acidic residue that is essential for the activity of RNase II-like enzymes. The possible involvement of this residue in metal binding at the active site of the enzyme is discussed. These results are particularly relevant at this time given that no structural or mutational analysis has been performed for any protein of the RNR family of exoribonucleases. PMID- 15654876 TI - Characterization of the secreted chorismate mutase from the pathogen Mycobacterium tuberculosis. AB - The gene encompassing ORF Rv1885c with weak sequence similarity to AroQ chorismate mutases (CMs) was cloned from the genome of Mycobacterium tuberculosis and expressed in Escherichia coli. The gene product (*MtCM) complements a CM deficient E. coli strain, but only if produced without the predicted N-terminal signal sequence typical of M. tuberculosis. The mature *MtCM, which was purified by exploiting its resistance to irreversible thermal denaturation, possesses high CM activity in vitro. The enzyme follows simple Michaelis-Menten kinetics, having a k(cat) of 50 s(-1) and a K(m) of 180 microM (at 30 degrees C and pH 7.5). *MtCM was shown to be a dimer by analytical ultracentrifugation and size-exclusion chromatography. Secondary-structure prediction and CD spectroscopy confirmed that *MtCM is a member of the all-alpha-helical AroQ class of CMs, but it seems to have a topologically rearranged AroQ fold. Because CMs are normally intracellular metabolic enzymes required for the biosynthesis of phenylalanine and tyrosine, the existence of an exported CM in Gram-positive M. tuberculosis is puzzling. The observation that homologs of *MtCM with a predicted export sequence are generally only present in parasitic or pathogenic organisms suggests that secreted CMs may have evolved to participate in some aspect of parasitism or pathogenesis yet to be unraveled. PMID- 15654877 TI - Molecular identification and expression study of differentially regulated genes in the Pacific oyster Crassostrea gigas in response to pesticide exposure. AB - The effects of pesticide contamination on the metabolism of marine molluscs are poorly documented. We investigated the response of a marine bivalve, the Pacific oyster, Crassostrea gigas, using a suppression subtractive hybridization method to identify up- and down-regulated genes after a 30-day exposure period to herbicides (a cocktail of atrazine, diuron and isoproturon, and to the single herbicide glyphosate). A total of 137 unique differentially expressed gene sequences was identified, as well as their associated physiological process. The expression of 18 of these genes was analyzed by RT-PCR under laboratory experimental conditions. The metabolic functions they are associated with include xenobiotic detoxification, energy production, immune system response and transcription. This study provides a preliminary basis for studying the response of marine bivalves to long-term herbicide exposure in terms of regulated gene expression and characterizes new potential genetic markers of herbicide contamination. PMID- 15654878 TI - Probing the access of protons to the K pathway in the Paracoccus denitrificans cytochrome c oxidase. AB - In recent studies on heme-copper oxidases a particular glutamate residue in subunit II has been suggested to constitute the entry point of the so-called K pathway. In contrast, mutations of this residue (E78(II)) in the Paracoccus denitrificans cytochrome c oxidase do not affect its catalytic activity at all (E78(II)Q) or reduce it to about 50% (E78(II)A); in the latter case, the mutation causes no drastic decrease in heme a(3) reduction kinetics under anaerobic conditions, when compared to typical K pathway mutants. Moreover, both mutant enzymes retain full proton-pumping competence. While oxidized-minus-reduced Fourier-transform infrared difference spectroscopy demonstrates that E78(II) is indeed addressed by the redox state of the enzyme, absence of variations in the spectral range characteristic for protonated aspartic and glutamic acids at approximately 1760 to 1710 cm(-1) excludes the protonation of E78(II) in the course of the redox reaction in the studied pH range, although shifts of vibrational modes at 1570 and 1400 cm(-1) reflect the reorganization of its deprotonated side chain at pH values greater than 4.8. We therefore conclude that protons do not enter the K channel via E78(II) in the Paracoccus enzyme. PMID- 15654879 TI - Annexin A2 binds to the localization signal in the 3' untranslated region of c myc mRNA. AB - Messenger RNA trafficking, which provides a mechanism for local protein synthesis, is dependent on cis-acting sequences in the 3' untranslated regions (3'UTRs) of the mRNAs concerned acting together with trans-acting proteins. The C MYC transcription factor is a proto-oncogene product involved in cell proliferation, differentiation and apoptosis. Localization of c-myc mRNA to the perinuclear cytoplasm and its association with the cytoskeleton is determined by a signal in the 3'UTR. Here we show the specific binding of a trans-acting factor to the perinuclear localization element in the 3'UTR of c-myc mRNA and identify this protein as annexin A2. Gel retardation and UV cross-linking experiments showed that proteins in fibroblast extracts formed complexes with the region of c myc 3'UTR implicated in localization; a protein of approximately 36 kDa exhibited specific, Ca(2+)-dependent binding. Binding was reduced by introduction of a mutation that abrogates localization. Using RNA-affinity columns followed by gel electrophoresis and mass spectrometry this protein was identified as annexin A2. The RNA-protein complex formed by cell extracts was further retarded by anti (annexin A2). Purified annexin A2 bound to the same region of the c-myc 3'UTR but binding was reduced by introduction of a mutation, as with cell extracts. It is proposed that binding of annexin A2 to the localization signal in the c-myc mRNA leads to association with the cytoskeleton and perinuclear localization. The data indicate a novel functional role for the RNA-binding properties of annexin A2 in perinuclear localization of mRNA and the association with the cytoskeleton. PMID- 15654880 TI - Effect of deletion of the DNase I hypersensitive sites on the transcription of chicken Ig-beta gene and on the maintenance of active chromatin state in the Ig beta locus. AB - The role of DNase I hypersensitive sites (DHSs) in transcription of the B cell specific Ig-beta gene and in maintenance of active chromatin state in the Ig-beta locus were examined. A total of 10 DHSs were divided into four regions, and each region was deleted separately in chicken B lymphocyte-derived DT40 cells. Deletion of three DHSs located between the Ig-beta promoter and its upstream Na channelgene, resulted in the absence of Ig-beta mRNA. Three regions except the region in the Na channel gene were involved in the transcription of Ig-beta gene. The enhancing activity of DHSs as determined by transient transfection assays did not always correlate with the effect of DHS deletion on the expression level of Ig-beta mRNA. In each deletion, cells contained the same DHSs as observed in the predeletion cells, indicating that deleted DHSs did not participate in the maintenance of DT40-specific DHSs. Enhanced acetylation of H3 and H4 histones at the Ig-beta promoter and at DT40-specific DHSs was observed in cells in which DHSs between the Na channel gene and Ig-beta promoter were deleted; therefore, these DHSs are prerequisite for transcription of the Ig-beta gene but not required for the maintenance of active chromatin state in the Ig-beta locus. Thus, epigenetic factors required for the maintenance of the active chromatin state are suggested to reside in other regions than those deleted in this study. PMID- 15654881 TI - An immunomodulator used to protect young in the pouch of the Tammar wallaby, Macropus eugenii. AB - Eugenin [pGluGlnAspTyr(SO(3))ValPheMetHisProPhe-NH(2)] has been isolated from the pouches of female Tammar wallabies (Macropus eugenii) carrying young in the early lactation period. The sequence of eugenin has been determined using a combination of positive and negative ion electrospray mass spectrometry. This compound bears some structural resemblance to the mammalian neuropeptide cholecystokinin 8 [AspTyr(SO(3))MetGlyTrpMetAspPhe-NH(2)] and to the amphibian caerulein peptides [caerulein: pGluGlnAspTyr(SO(3))ThrGlyTrpMetAspPhe-NH(2)]. Eugenin has been synthesized by a route which causes only minor hydrolysis of the sulfate group when the peptide is removed from the resin support. Biological activity tests with eugenin indicate that it contracts smooth muscle at a concentration of 10( 9) M, and enhances the proliferation of splenocytes at 10(-7) M, probably via activation of CCK(2) receptors. The activity of eugenin on splenocytes suggests that it is an immunomodulator peptide which plays a role in the protection of pouch young. PMID- 15654882 TI - Polarized distribution of inducible nitric oxide synthase regulates activity in intestinal epithelial cells. AB - Inducible nitric oxide synthase (iNOS) functions as a homodimer. In cell extracts, iNOS molecules partition both in cytosolic and particulate fractions, indicating that iNOS exists as soluble and membrane associated forms. In this study, iNOS features were investigated in human intestinal epithelial cells stimulated with cytokines and in duodenum from mice exposed to flagellin. Our experiments indicate that iNOS is mainly associated with the particulate fraction of cell extracts. Confocal microscopy showed a preferential localization of iNOS at the apical pole of intestinal epithelial cells. In particulate fractions, iNOS dimers were more abundant than in the cytosolic fraction. Similar observations were seen in mouse duodenum samples. These results suggest that, in epithelial cells, iNOS activity is regulated by localization-dependent processes. PMID- 15654883 TI - Stimulation of poly(A) synthesis by Escherichia coli poly(A)polymerase I is correlated with Hfq binding to poly(A) tails. AB - The bacterial Lsm protein, host factor I (Hfq), is an RNA chaperone involved in many types of RNA transactions such as replication and stability, control of small RNA activity and polyadenylation. In this latter case, Hfq stimulates poly(A) synthesis and binds poly(A) tails that it protects from exonucleolytic degradation. We show here, that there is a correlation between Hfq binding to the 3' end of an RNA molecule and its ability to stimulate RNA elongation catalyzed by poly(A)polymerase I. In contrast, formation of the Hfq-RNA complex inhibits elongation of the RNA by polynucleotide phosphorylase. We demonstrate also that Hfq binding is not affected by the phosphorylation status of the RNA molecule and occurs equally well at terminal or internal stretches of poly(A). PMID- 15654885 TI - The N-terminus of B96Bom, a Bombyx mori G-protein-coupled receptor, is N myristoylated and translocated across the membrane. AB - In eukaryotic cellular proteins, protein N-myristoylation has been recognized as a protein modification that occurs mainly on cytoplasmic or nucleoplasmic proteins. In this study, to search for a eukaryotic N-myristoylated transmembrane protein, the susceptibility of the N-terminus of several G-protein-coupled receptors (GPCRs) to protein N-myristoylation was evaluated by in vitro and in vivo metabolic labeling. It was found that the N-terminal 10 residues of B96Bom, a Bombyx mori GPCR, efficiently directed the protein N-myristoylation. Analysis of a tumor necrosis factor (TNF) fusion protein with the N-terminal 90 residues of B96Bom at its N-terminus revealed that (a) transmembrane domain 1 of B96Bom functioned as a type I signal anchor sequence, (b) the N-myristoylated N-terminal domain (58 residues) was translocated across the membrane, and (c) two N glycosylation motifs located in this domain were efficiently N-glycosylated. In addition, when Ala4 in the N-myristoylation motif of B96Bom90-TNF, Met-Gly-Gln Ala-Ala-Thr(1-6), was replaced with Asn to generate a new N-glycosylation motif, Asn-Ala-Thr(4-6), efficient N-glycosylation was observed on this newly introduced N-glycosylation site in the expressed protein. These results indicate that the N myristoylated N-terminus of B96Bom is translocated across the membrane and exposed to the extracellular surface. To our knowledge, this is the first report showing that a eukaryotic transmembrane protein can be N-myristoylated and that the N-myristoylated N-terminus of the protein can be translocated across the membrane. PMID- 15654884 TI - Effect of gadolinium on the ryanodine receptor/sarcoplasmic reticulum calcium release channel of skeletal muscle. AB - The effect of gadolinium ions on the sarcoplasmic reticulum (SR) calcium release channel/ryanodine receptor (RyR1) was studied using heavy SR (HSR) vesicles and RyR1 isolated from rabbit fast twitch muscle. In the [(3)H]ryanodine binding assay, 5 microM Gd(3+) increased the K(d) of the [(3)H]ryanodine binding of the vesicles from 33.8 nM to 45.6 nM while B(max), referring to the binding capacity, was not affected significantly. In the presence of 18 nM[(3)H]ryanodine and 100 microM free Ca(2+), Gd(3+) inhibited the binding of the radiolabeled ryanodine with an apparent K(d) value of 14.7 microM and a Hill coefficient of 3.17. In (45)Ca(2+) experiments the time constant of (45)Ca(2+) efflux from HSR vesicles increased from 90.9 (+/- 11.1) ms to 187.7 (+/- 24.9) ms in the presence of 20 microM gadolinium. In single channel experiments gadolinium inhibited the channel activity from both the cytoplasmic (cis) (IC(50) = 5.65 +/- 0.33 microM, n(Hill) = 4.71) and the luminal (trans) side (IC(50) = 5.47 +/- 0.24 microM, n(Hill) = 4.31). The degree of inhibition on the cis side didn't show calcium dependency in the 100 microM to 1 mM Ca(2+) concentration range which indicates no competition with calcium on its regulatory binding sites. When Gd(3+) was applied at the trans side, EGTA was present at the cis side to prevent the binding of Gd(+3) to the cytoplasmic calcium binding regulatory sites of the RyR1 if Gd(3+) accidentally passed through the channel. The inhibition of the channel did not show any voltage dependence, which would be the case if Gd(3+) exerted its effect after getting to the cis side. Our results suggest the presence of inhibitory binding sites for Gd(3+) on both sides of the RyR1 with similar Hill coefficients and IC(50) values. PMID- 15654886 TI - Co-operative effect of the isoforms of type III antifreeze protein expressed in Notched-fin eelpout, Zoarces elongatus Kner. AB - We found that Notched-fin eelpout, which lives off the north east coast of Japan, expresses an antifreeze protein (AFP). The liver of this fish contains DNAs that encode at least 13 type III AFP isoforms (denoted nfeAFPs). The primary sequences of the nfeAFP isoforms were categorized into SP- and QAE-sephadex binding groups, and the latter were further divided into two subgroups, QAE1 and QAE2 groups. Ice crystals observed in HPLC-pure nfeAFP fractions are bipyramidal in shape with different ratios of c and a axes, suggesting that all the isoforms are able to bind ice. We expressed five recombinant isoforms of nfeAFP and analyzed the thermal hysteresis (TH) activity of each as a function of protein concentration. We also examined the change in activity on mixing the isoforms. TH was estimated to be 0.60 degrees C for the QAE1 isoform, 0.11 degrees C for QAE2, and almost zero for the SP isoforms when the concentrations of these isoforms was standardized to 1.0 mm. Significantly, the TH activity of the SP isoforms showed concentration dependence in the presence of 0.2 mm QAE1, indicating that the less active SP isoform becomes 'active' when a small amount of QAE1 is added. In contrast, it does not become active on the addition of another SP isoform. These results suggest that the SP and QAE isoforms of type III AFP have different levels of TH activity, and they accomplish the antifreeze function in a co operative manner. PMID- 15654887 TI - Expression of an alpha-1,3-glucanase during mycoparasitic interaction of Trichoderma asperellum. AB - Trichoderma species have been investigated as biological control agents for over 70 years owing to their ability to antagonize plant pathogenic fungi. Mycoparasitism, one of the main mechanisms involved in the antagonistic activity of Trichoderma strains, depends on the secretion of complex mixtures of hydrolytic enzymes able to degrade the host cell wall. The antifungal activity of an alpha-1,3-glucanase (EC 3.2.1.59, enzymes able to degrade alpha-1,3-glucans and also named mutanases) has been described in T. harzianum and its role in mycoparasitic processes has been suggested. In this study, we report on the purification, characterization and cloning of an exo-alpha-1,3-glucanase, namely AGN13.2, from the antagonistic fungus T. asperellum T32. Expression at the transcription level in confrontation assays against the strawberry pathogen Botrytis cinerea strongly supports the role of AGN13.2 during the antagonistic action of T. asperellum. PMID- 15654888 TI - Molecular characterization, phylogenetic relationships, and developmental expression patterns of prion genes in zebrafish (Danio rerio). AB - Prion diseases are characterized by the accumulation of a pathogenic misfolded form of a prion protein (PrP) encoded by the Prnp gene in humans. In the present study in zebrafish, two transcripts and the corresponding genes encoding prion proteins, PrP1 and PrP2, related to human PrP have been characterized with a relatively divergent deduced amino acid sequence, but a well preserved overall organization of structural prion protein motifs. Whole-mount in situ hybridization analysis performed during embryonic and larval development showed a high level of PrP1 mRNA spatially restricted to the anterior floor-plate of the central nervous system and in ganglia. Transcripts of prp2 were detected in embryonic cells from the mid-blastula transition to the end of the segmentation period. From 24 h postfertilization up to larval stages, prp2 transcripts were localized in distinct anatomical structures, including a major expression in the brain, eye, kidney, lateral line neuromasts, liver, heart, pectoral fins and posterior intestine. The observed differential developmental expression patterns of the two long PrP forms, prp1 and prp2, and the short PrP form prp3, a more divergent prion-related gene previously identified in zebrafish, should contribute to understanding of the phylogenetic and functional relationships of duplicated prion gene forms in the fish genome. Together, the complex history of prion-related genes, reflected in the deduced structural features, conserved amino acid sequence and repeat motifs of the corresponding proteins, and the presence of differential developmental expression patterns suggest possible acquisition or loss of prion protein functions during vertebrate evolution. PMID- 15654889 TI - Comparison of the substrate specificity of two potyvirus proteases. AB - The substrate specificity of the nuclear inclusion protein a (NIa) proteolytic enzymes from two potyviruses, the tobacco etch virus (TEV) and tobacco vein mottling virus (TVMV), was compared using oligopeptide substrates. Mutations were introduced into TEV protease in an effort to identify key determinants of substrate specificity. The specificity of the mutant enzymes was assessed by using peptides with complementary substitutions. The crystal structure of TEV protease and a homology model of TVMV protease were used to interpret the kinetic data. A comparison of the two structures and the experimental data suggested that the differences in the specificity of the two enzymes may be mainly due to the variation in their S4 and S3 binding subsites. Two key residues predicted to be important for these differences were replaced in TEV protease with the corresponding residues of TVMV protease. Kinetic analyses of the mutants confirmed that these residues play a role in the specificity of the two enzymes. Additional residues in the substrate-binding subsites of TEV protease were also mutated in an effort to alter the specificity of the enzyme. PMID- 15654890 TI - Functional expression of olfactory receptors in yeast and development of a bioassay for odorant screening. AB - The functional expression of olfactory receptors (ORs) is a primary requirement to examine the molecular mechanisms of odorant perception and coding. Functional expression of the rat I7 OR and its trafficking to the plasma membrane was achieved under optimized experimental conditions in the budding yeast Saccharomyces cerevisiae. The membrane expression of the receptor was shown by Western blotting and immunolocalization methods. Moreover, we took advantage of the functional similarities between signal transduction cascades of G protein coupled receptor in mammalian cells and the pheromone response pathway in yeast to develop a novel biosensor for odorant screening using luciferase as a functional reporter. Yeasts were engineered to coexpress I7 OR and mammalian G(alpha) subunit, to compensate for the lack of endogenous Gpa1 subunit, so that stimulation of the receptor by its ligands activates a MAP kinase signaling pathway and induces luciferase synthesis. The sensitivity of the bioassay was significantly enhanced using mammalian G(olf) compared to the G(alpha15) subunit, resulting in dose-dependent responses of the system. The biosensor was probed with an array of odorants to demonstrate that the yeast-borne I7 OR retains its specificity and selectivity towards ligands. The results are confirmed by functional expression and bioluminescence response of human OR17-40 to its specific ligand, helional. Based on these findings, the bioassay using the luciferase reporter should be amenable to simple, rapid and inexpensive odorant screening of hundreds of ORs to provide insight into olfactory coding mechanisms. PMID- 15654891 TI - Degradation of chitosans with chitinase B from Serratia marcescens. Production of chito-oligosaccharides and insight into enzyme processivity. AB - Family 18 chitinases such as chitinase B (ChiB) from Serratia marcescens catalyze glycoside hydrolysis via a mechanism involving the N-acetyl group of the sugar bound to the -1 subsite. We have studied the degradation of the soluble heteropolymer chitosan, to obtain further insight into catalysis in ChiB and to experimentally assess the proposed processive action of this enzyme. Degradation of chitosans with varying degrees of acetylation was monitored by following the size-distribution of oligomers, and oligomers were isolated and partly sequenced using (1)H-NMR spectroscopy. Degradation of a chitosan with 65% acetylated units showed that ChiB is an exo-enzyme which degrades the polymer chains from their nonreducing ends. The degradation showed biphasic kinetics: the faster phase is dominated by cleavage on the reducing side of two acetylated units (occupying subsites -2 and -1), while the slower kinetic phase reflects cleavage on the reducing side of a deacetylated and an acetylated unit (bound to subsites -2 and 1, respectively). The enzyme did not show preferences with respect to acetylation of the sugar bound in the +1 subsite. Thus, the preference for an acetylated unit is absolute in the -1 subsite, whereas substrate specificity is less stringent in the -2 and +1 subsites. Consequently, even chitosans with low degrees of acetylation could be degraded by ChiB, permitting the production of mixtures of oligosaccharides with different size distributions and chemical composition. Initially, the degradation of the 65% acetylated chitosan almost exclusively yielded oligomers with even-numbered chain lengths. This provides experimental evidence for a processive mode of action, moving the sugar chain two residues at a time. The results show that nonproductive binding events are not necessarily followed by substrate release but rather by consecutive relocations of the sugar chain. PMID- 15654892 TI - 2-Hydroxyisocaproyl-CoA dehydratase and its activator from Clostridium difficile. AB - The hadBC and hadI genes from Clostridium difficile were functionally expressed in Escherichia coli and shown to encode the novel 2-hydroxyisocaproyl-CoA dehydratase HadBC and its activator HadI. The activated enzyme catalyses the dehydration of (R)-2-hydroxyisocaproyl-CoA to isocaprenoyl-CoA in the pathway of leucine fermentation. The extremely oxygen-sensitive homodimeric activator as well as the heterodimeric dehydratase, contain iron and inorganic sulfur; besides varying amounts of zinc, other metal ions, particularly molybdenum, were not detected in the dehydratase. The reduced activator transfers one electron to the dehydratase concomitant with hydrolysis of ATP, a process similar to that observed with the unrelated nitrogenase. The thus activated dehydratase was separated from the activator and ATP; it catalyzed about 10(4) dehydration turnovers until the enzyme became inactive. Adding activator, ATP, MgCl(2), dithionite and dithioerythritol reactivated the enzyme. This is the first demonstration with a 2-hydroxyacyl-CoA dehydratase that the catalytic electron is recycled after each turnover. In agreement with this observation, only substoichiometric amounts of activator (dehydratase/activator = 10 mol/mol) were required to generate full activity. PMID- 15654893 TI - Detection of native peptides as potent inhibitors of enzymes. Crystal structure of the complex formed between treated bovine alpha-chymotrypsin and an autocatalytically produced fragment, IIe-Val-Asn-Gly-Glu-Glu-Ala-Val-Pro-Gly-Ser Trp-Pro-Trp, at 2.2 angstroms resolution. AB - Chymotrypsin is a prominent member of the family of serine proteases. The present studies demonstrate the presence of a native fragment containing 14 residues from Ile16 to Trp29 in alpha-chymotrypsin that binds to chymotrypsin at the active site with an exceptionally high affinity of 2.7 +/- 0.3 x 10(-11) M and thus works as a highly potent competitive inhibitor. The commercially available alpha chymotrypsin was processed through a three phase partitioning system (TPP). The treated enzyme showed considerably enhanced activity. The 14 residue fragment was produced by autodigestion of a TPP-treated alpha-chymotrypsin during a long crystallization process that lasted more than four months. The treated enzyme was purified and kept for crystallization using vapour the diffusion method at 295 K. Twenty milligrams of lyophilized protein were dissolved in 1 mL of 25 mM sodium acetate buffer, pH 4.8. It was equilibrated against the same buffer containing 1.2 M ammonium sulfate. The rectangular crystals of small dimensions of 0.24 x 0.15 x 0.10 mm(3) were obtained. The X-ray intensity data were collected at 2.2 angstroms resolution and the structure was refined to an R-factor of 0.192. An extra electron density was observed at the binding site of alpha-chymotrypsin, which was readily interpreted as a 14 residue fragment of alpha-chymotrypsin corresponding to Ile-Val-Asn-Gly-Glu-Glu-Ala-Val-Pro-Gly-Ser-Trp-Pro-Trp(16-29). The electron density for the eight residues of the C-terminus, i.e. Ala22-Trp29, which were completely buried in the binding cleft of the enzyme, was of excellent quality and all the side chains of these eight residues were clearly modeled into it. However, the remaining six residues from the N-terminus, Ile16-Glu21 were poorly defined although the backbone density was good. There was a continuous electron density at 3.0 sigma between the active site Ser195 Ogamma and the carbonyl carbon atom of Trp29 of the fragment. The final refined coordinates showed a distance of 1.35 angstroms between Ser195 Ogamma and Trp29 C indicating the presence of a covalent linkage between the enzyme and the native fragment. This meant that the enzyme formed an acyl intermediate with the autodigested fragment Ile16-Trp29. In addition to the O-C covalent bond, there were several hydrogen bonds and hydrophobic interactions between the enzyme and the native fragment. The fragment showed a high complementarity with the binding site of alpha-chymotrypsin and the buried part of the fragment matched excellently with the corresponding buried part of Turkey ovomucoid inhibitor of alpha chymotrypsin. PMID- 15654894 TI - Binding of the volatile general anesthetics halothane and isoflurane to a mammalian beta-barrel protein. AB - A molecular understanding of volatile anesthetic mechanisms of action will require structural descriptions of anesthetic-protein complexes. Porcine odorant binding protein is a 157 residue member of the lipocalin family that features a large beta-barrel internal cavity (515 +/- 30 angstroms(3)) lined predominantly by aromatic and aliphatic residues. Halothane binding to the beta-barrel cavity was determined using fluorescence quenching of Trp16, and a competitive binding assay with 1-aminoanthracene. In addition, the binding of halothane and isoflurane were characterized thermodynamically using isothermal titration calorimetry. Hydrogen exchange was used to evaluate the effects of bound halothane and isoflurane on global protein dynamics. Halothane bound to the cavity in the beta-barrel of porcine odorant binding protein with dissociation constants of 0.46 +/- 0.10 mM and 0.43 +/- 0.12 mM determined using fluorescence quenching and competitive binding with 1-aminoanthracene, respectively. Isothermal titration calorimetry revealed that halothane and isoflurane bound with K(d) values of 80 +/- 10 microM and 100 +/- 10 microM, respectively. Halothane and isoflurane binding resulted in an overall stabilization of the folded conformation of the protein by -0.9 +/- 0.1 kcal.mol(-1). In addition to indicating specific binding to the native protein conformation, such stabilization may represent a fundamental mechanism whereby anesthetics reversibly alter protein function. Because porcine odorant binding protein has been successfully analyzed by X-ray diffraction to 2.25 angstroms resolution [1], this represents an attractive system for atomic-level structural studies in the presence of bound anesthetic. Such studies will provide much needed insight into how volatile anesthetics interact with biological macromolecules. PMID- 15654895 TI - Functional implications of pigments bound to a cyanobacterial cytochrome b6f complex. AB - A highly purified cytochrome b(6)f complex from the cyanobacterium Synechocystis sp. PCC 6803 selectively binds one chlorophyll a and one carotenoid in analogy to the recent published structure from two other b(6)f complexes. The unknown function of these pigments was elucidated by spectroscopy and site-directed mutagenesis. Low-temperature redox difference spectroscopy showed red shifts in the chlorophyll and carotenoid spectra upon reduction of cytochrome b(6), which indicates coupling of these pigments with the heme groups and thereby with the electron transport. This is supported by the correlated kinetics of these redox reactions and also by the distinct orientation of the chlorophyll molecule with respect to the heme cofactors as shown by linear dichroism spectroscopy. The specific role of the carotenoid echinenone for the cytochrome b(6)f complex of Synechocystis 6803 was elucidated by a mutant lacking the last step of echinenone biosynthesis. The isolated mutant complex preferentially contained a carotenoid with 0, 1 or 2 hydroxyl groups (most likely 9-cis isomers of beta-carotene, a monohydroxy carotenoid and zeaxanthin, respectively) instead. This indicates a substantial role of the carotenoid - possibly for strucure and assembly - and a specificity of its binding site which is different from those in most other oxygenic photosynthetic organisms. In summary, both pigments are probably involved in the structure, but may also contribute to the dynamics of the cytochrome b(6)f complex. PMID- 15654896 TI - Probing the mechanism of the bifunctional enzyme ketol-acid reductoisomerase by site-directed mutagenesis of the active site. AB - Ketol-acid reductoisomerase (EC 1.1.1.86) is involved in the biosynthesis of the branched-chain amino acids. It is a bifunctional enzyme that catalyzes two quite different reactions at a common active site; an isomerization consisting of an alkyl migration, followed by an NADPH-dependent reduction of a 2-ketoacid. The 2 ketoacid formed by the alkyl migration is not released. Using the pure recombinant Escherichia coli enzyme, we show that the isomerization reaction has a highly unfavourable equilibrium constant. The reductase activity is shown to be relatively nonspecific and is capable of utilizing a variety of 2-ketoacids. The active site of the enzyme contains eight conserved polar amino acids and we have mutated each of these in order to dissect their contributions to the isomerase and reductase activities. Several mutations result in loss of the isomerase activity with retention of reductase activity. However, none of the 17 mutants examined have the isomerase activity only. We suggest a reason for this, involving direct reduction of a transition state formed during the isomerization, which is necessitated by the unfavourable equilibrium position of the isomerization. Our mechanism explains why the two activities must occur in a single active site without release of a 2-ketoacid and provides a rationale for the requirement for NADPH by the isomerase. PMID- 15654897 TI - Light-induced global structural changes in phytochrome A regulating photomorphogenesis in plants. AB - Phytochromes are photoreceptor proteins that monitor the light environment and regulate a variety of photomorphogenic responses to optimize the growth and development of plants. Phytochromes comprise N-terminal photosensory and C terminal regulatory domains. They are mutually photoconvertible between a red light-absorbing (Pr) and a far-red-light-absorbing (Pfr) form. Their interconversion by light stimuli initiates downstream signaling cascades. Here we report the molecular structures of pea phytochrome A lacking the N-terminal 52 amino-acid residues in the Pr and Pfr forms studied by small-angle X-ray scattering. A new purification protocol yielded monodispersive sample solutions. The molecular mass and the maximum dimension of Pr determined from scattering data indicated its dimeric association. The molecular structure of Pr predicted by applying the ab initio simulation method to the scattering profile was approximated as a stack of two flat bodies, comprising two lobes assignable to the functional regions. Scattering profiles recorded under red-light irradiation showed small but definite changes from those of Pr. The molecular dimensions and predicted molecular structure of Pfr suggest global structural changes such as movement of the C-terminal domains in the Pr-to-Pfr phototransformation. Red light-induced structural changes in Pfr were reversible, mostly due to thermal relaxation processes. PMID- 15654898 TI - Serum ferritin level as a predictor of impaired growth and puberty in thalassemia major patients. AB - OBJECTIVE: Previous studies suggested that in patients with thalassemia major, initiating deferoxamine (DFO) therapy before puberty can prevent iron-induced failure of growth and puberty. However, early initiation of chelation has also been associated with DFO toxicity. The aim of this retrospective study was to determine the prevalence rates of endocrine complications and DFO bone toxicity in our thalassemia major patients and to correlate them with the degree of iron chelation. METHODS: Thirty-nine patients with thalassemia major were followed for a median of 16.3 yr (range 2-28). Individual mean serum ferritin level during the study period was calculated using repeated annual measurements. Bone DFO toxicity was assessed by wrist and spine radiographs; endocrine dysfunction by anthropometric measurements and pubertal stage; and hypogonadotropic hypogonadism by lack of luteinizing hormone response to gonadotropin-releasing hormone. RESULTS: Chelation therapy was initiated at median age 4.9 yr. Mean serum ferritin level during the study period was 2698 +/- 1444 ng/mL. Hypogonadism was noted in 59% of the patients who reached pubertal age, and short stature was found in 36% of patients who reached final height. Mean ferritin level of 2500 ng/mL during puberty was the cut-off for hypogonadism, and ferritin level of 3000 ng/mL during prepuberty was the cut-off for final short stature. None of the patients who attained final height had signs of DFO bone toxicity. CONCLUSIONS: High serum ferritin levels during puberty are a risk factor for hypogonadism, and high serum ferritin levels during the first decade of life predict final short stature. It remains to be determined whether improving chelation by earlier initiation of DFO or by the combined use of DFO and deferiprone will lead to better growth and sexual development without DFO toxicity. PMID- 15654899 TI - Serum IgM, IgG and IgA block by F(ab')-dependent mechanism the binding of natural IgG autoantibodies from therapeutic immunoglobulin preparations to self-antigens. AB - Natural polyreactive IgG autoantibodies are present in the plasma of healthy individuals and as a result in pooled therapeutic intravenous immunoglobulin (i.v.Ig) preparations. The spectrum of self-antigens to which these autoantibodies bind, their fate after intravenous infusion and their biological activity are not well understood. The identity of serum proteins that mask binding of natural autoantibodies to self-proteins is a matter of controversy. The spectrum of native serum proteins bound by i.v.Ig was analyzed by two dimensional electrophoresis. The reactivity of i.v.Ig was directed mainly to circulating immunoglobulins. The binding of the IgG autoantibodies from i.v.Ig to native human liver antigens was blocked not only by a F(ab')2-dependent mechanism by circulating IgM and IgG (as has been previously suggested), but also by serum IgA. This control of anti-self reactivity may be inefficient in some autoimmune diseases. PMID- 15654900 TI - Insulin-like growth factor-1 is a predictor of blood haemoglobin concentration in 70-yr-old subjects. AB - The role of growth hormone (GH) for maintaining normal erythropoiesis among non GH-deficient elderly subjects is not known. OBJECTIVES: To determine relationships between the concentrations of serum insulin-like growth factor-1 (IGF-1) and it's carrier protein insulin-like growth factor binding protein 3 (IGFBP-3), as well as plasma erythropoietin (EPO), and blood haemoglobin (Hb) in elderly subjects. METHODS: Serum IGF-1 and IGFBP-3 and plasma EPO were, in addition to basal haematological tests, measured in a community based representative population sample of 70-yr-olds (n = 619; 317 women and 302 men). Statistical analyses were made before and after exclusion of non-healthy subjects. RESULTS: Mean IGF-1 concentration was higher among men than women (155 vs. 138 mug/L, P = 0.0000), and that of IGFBP3 lower (2.21 vs. 2.65 mg/L, P = 0.0000). Exclusion of subjects with demonstrable disorders did not significantly influence the distributions of serum-IGF-1 and serum-BP-3. Hb concentration was positively correlated to concentrations of IGF-1 (r = 0.15, P < 0.01 for the men; r = 0.34, P < 0.001 for the women), and IGFBP-3 (r = 0.07, n.s. for the men; r = 0.27, P < 0.001 for the women), and negatively correlated to EPO concentration (r = -0.34, P < 0.001 for the men; r = -0.42, P < 0.001 for the women). In multivariate analysis, serum concentrations of IGF-1, iron, and plasma concentrations of EPO were independently correlated to Hb among both men and women. CONCLUSION: GH and/or IGF-1 are, independently from EPO and regardless of health status and sex, a significant determinant of Hb in elderly subjects. PMID- 15654901 TI - Long-term analysis of the palliative benefit of 2-chlorodeoxyadenosine for myelofibrosis with myeloid metaplasia. AB - OBJECTIVE: Therapeutic splenectomy in myelofibrosis with myeloid metaplasia (MMM) may result in extreme thrombocytosis and leukocytosis and accelerated hepatomegaly. We previously described initial palliative benefit from 2 chlorodeoxyadenosine (2-CdA) in such instances. The purpose of this study is to provide long-term follow-up on the durability of response in the initial cohort and in additional subsequent cases. METHODS: We retrospectively identified patients with histologically confirmed MMM who had palliative therapy with 2-CdA. Clinical characteristics and information on subsequent clinical course were abstracted at the time of diagnosis of MMM and at initiation of 2-CdA therapy until death. RESULTS: To date, we have used 2-CdA as palliative therapy in 14 patients with MMM. After a median of four cycles of therapy, responses for hepatomegaly occurred in 56% of patients, thrombocytosis 50%, leukocytosis 55%, and anemia 40%. Cytopenias were frequent but usually transient and without clinical consequence. Responses occurred usually by the second cycle; median duration of response was 6 months (range, 2-19) after completion of 2-CdA therapy. CONCLUSION: This study confirmed relevant and frequently durable palliation of symptoms in about half the patients. 2-CdA is a reasonable palliative option in postsplenectomy patients with MMM who have problematic myeloproliferation. PMID- 15654902 TI - Early and tardive skin adverse events in chronic myeloid leukaemia patients treated with imatinib. AB - Imatinib related non-haematological side-effects are reported in <10% of chronic myeloid leukaemia patients and include oedema, weight gain, nausea, vomiting and muscle cramps. Cutaneous reactions are well-recognized events occurring mostly in patients treated at doses of 600 mg/d and higher, either in stable or progressive disease. We report on our experience relating to dermatological toxicities in imatinib treated CML patients showing a spectrum of skin reactions ranging from rashes to cutaneous carcinoma. PMID- 15654903 TI - Clinical correlates of submicroscopic deletions involving the ABL-BCR translocation region in chronic myeloid leukemia. AB - Recent reports indicate a prognostically detrimental effect of submicroscopic abl bcr deletions associated with the break and fusion points of the derivative chromosome 9 [der(9)] in chronic myeloid leukemia (CML). In a retrospective cohort of 92 patients with CML, the incidence of an atypical D-FISH pattern, that is consistent with a der(9) deletion was 20%. Complete clinical information was available in 82 patients and revealed no significant differences between 18 deleted and 64 non-deleted cases in platelet count, circulating blast percentage, spleen size, or karyotype profile at presentation. However, der(9)-deleted patients presented with significantly lower hemoglobin levels and higher leukocyte counts. At a median follow-up of 31 months, the incidence of disease transformation, drug therapy response, and survival were similar between the two groups. These results are contrary to previous reports that suggested inferior survival as well as poor response to alpha interferon therapy in CML patients carrying der(9) deletions. PMID- 15654904 TI - Preclinical ex vivo expansion of G-CSF-mobilized peripheral blood stem cells: effects of serum-free media, cytokine combinations and chemotherapy. AB - OBJECTIVES: Ex vivo expansion of granulocyte-colony stimulating factor (G-CSF) mobilized peripheral blood stem cells (PBSC) is a promising approach for overcoming the developmental delay of bone marrow (BM) reconstitution after transplantation. This project investigated the effects of culture duration, serum free media, cytokine combinations, and chemotherapy on the outcomes of expansion. METHODS: Enriched CD34+ cells were cultured for 8 or 10 d in serum-free media (QBSF-60 or X-Vivo 10) and four combinations of cytokines consisting of recombinant human pegylated-megakaryocyte growth and development factor, stem cell factor, flt-3 ligand, G-CSF, interleukin (IL)-6, platelet-derived growth factor (PDGF), and IL-1beta. RESULTS: Eight days of culture in QBSF-60 significantly supported efficient expansions of CD34+ cells, CD34+ CD38- cells, colony-forming units (CFU) of myeloid, erythroid, megakaryocytic, and mixed lineages to 3.76-, 14.4-, 28.3-, 24.0-, 38.1-, and 15.7-fold, respectively. Whilst PDGF or IL-6 enhanced the expansion of early, myeloid, and erythroid progenitors, IL-1beta specifically promoted the megakaryocytic lineage. Engraftment of human CD45+ cells were detectable in all non-obese diabetic/severe combined immunodeficient mice transplanted with expanded PBSC from donor samples, being 5.80 +/- 3.34% of mouse BM cells. The expansion and engraftment capacity of CD34+ cells from subjects postchemotherapy were significantly compromised across the panel of progenitor cells. CONCLUSION: Our results provided an optimized protocol for PBSC expansion, applicable to ameliorating neutropenia and thrombocytopenia in post-BM transplant patients by the prompt provision of progenitor cells. For postchemotherapy patients, expansion products might provide committed progenitors for improving short-term engraftment, but not self renewable stem cells. PMID- 15654905 TI - Health related quality of life in patients with multiple myeloma undergoing a double transplantation. AB - OBJECTIVES: To investigate the subjective well-being of patients with newly diagnosed multiple myeloma who were treated in a tandem transplantation programme. METHODS: Fifty-one patients participated in the prospective, longitudinal questionnaire study. The EORTC QLQ-C30 and the EuroQol-5D were administered 2 wk after completion of vincristine, adriamycin and dexamethason/vincristine, adriamycin and methyl prednison (VAD/VAMP) chemotherapy, both at hospital discharge after treatment with high-dose melphalan (HDM) and 1 month after this hospital discharge, at hospital admission, at the day of hospital discharge for peripheral stem cell transplantation (PSCT) and at 6 and 12 months following discharge after PSCT. RESULTS: Overall, patients' functioning improved during treatment and follow-up, with significant decreases shortly following PSCT. Shortly after HDM and PSCT, patients reported a considerable increase in levels of soreness in the mouth (+26/+36 points on a scale ranging form 0 to 100; P < 0.01), change of taste (+23/+21 points; P < 0.05/NS), nausea/vomiting (+26/+27 points; P < 0.01/< 0.05), appetite loss (+40/+43 points; P < 0.001) and diarrhoea (+25/+36 points; P < 0.01). However, none of these symptoms persisted during follow-up. CONCLUSION: The intensive treatment programme was subjectively being well tolerated by the majority of patients. The duration of declined quality of life after administration of HDM seemed to be short. The duration of subjective recovery after PSCT remained uncertain, but in any case was present at the 6 month follow-up. Together with the rather good results in survival, the evaluation of quality of life invites further exploration of double transplantations in multiple myeloma. PMID- 15654906 TI - Conventional versus reduced-intensity conditioning regimen for allogeneic stem cell transplantation in patients with hematological malignancies. AB - BACKGROUND: Allogeneic hematopoietic stem cell transplantation (HSCT) from human leukocyte antigen (HLA)-compatible sibling donors is a potential curative treatment for hematological and non-hematological malignancies. Nevertheless, high mortality rates may be associated with this therapy, especially in older patients, those with other comorbidities or who receive a second HSCT. PATIENTS AND METHODS: We analyzed the factors associated with transplant-related mortality (TRM) and overall survival in 157 consecutive adult patients (104 males and 53 females) who received a HSCT [29 bone marrow (BM) transplantation and 128 peripheral blood (PB) transplantation] from a HLA-identical sibling between January 1995 and March 2002 in our institution. One hundred patients received a standard conditioning prior to HSCT (STAND) and 57 patients received a reduced intensity conditioning (RIC) HSCT. Fifty-eight patients were in an early phase at transplant and 99 in a non-early phase. Median age was 46 yr (16-66), and 90 patients (57%) were >45 yr of age. RESULTS: Patients in the RIC group were older than those in the STAND group, and had a higher proportion of non-early disease phases including a prior autologous HSCT in 39%. Median follow-up for survivors was 28 and 15 months in the STAND and RIC groups (P < 0,001), respectively. Cumulative incidence of TRM at 2 yr was 30% [95% confidence interval (CI) 22-41%] for the STAND group and 22% (95% CI 13-37%) for the RIC group [non-significant (NS)]. Factors associated with a higher TRM in multivariate analysis were: STAND vs. RIC conditioning regimen [relative risk (RR) 5.4; 95% CI 2.3-12.8; P < 0.001]; age > or =45 yr vs. <45 yr (RR 5; 95% CI 2.4-10.8, P < 0.001); second vs. first HSCT (RR 2.8, 95% CI 1.3-6.3, P = 0.01) and non-T-cell-depleted vs. T-cell depleted graft (RR 2.7, 95% CI 1.3-5.8, P = 0.009). Overall survival (OS) at 2 yr was 52.5 +/- 10.4% for STAND group and 59 +/- 16.8% in RIC group. Factors associated with poorer OS in multivariate analysis were: STAND vs. RIC conditioning regimen (RR 3.4, 95% CI 1.7-6.9, P = 0.001); age > or =45 vs <45 yr (RR 2.5, 95% CI 1.4-4.5, P = 0.002) and diagnosis [other than chronic myeloid leukemia (CML) vs. CML] (RR 2.6, 95% CI 1.2-5.7 P = 0.02). CONCLUSIONS: Our results indicate that the introduction of RIC allogeneic HSCT for patients at high risk for TRM (advanced age, prior HSCT and non-T-cell depletion) leads to a reduction in the TRM and improvement in the OS. PMID- 15654908 TI - Prevalence of hepatitis B and C virus infection in haematological malignancies and liver injury following chemotherapy. AB - The aim of this study was to determine the carrier rate of hepatitis virus in patients with haematological malignancies and the incidence of liver injury in these patients following chemotherapy. From January 1996 to September 2002, we studied 601 consecutive, unselected series of patients with haematological malignancies admitted in our hospital unit (Japan). They consisted of 246 cases of acute leukaemia, 218 non-Hodgkin's lymphoma (NHL), 13 adult T-cell leukaemia, and 124 multiple myeloma. Of these 601 patients, 373 were men and 228 were women; their mean age was 61 yr, with a range from 18 to 89 yr. The prevalences of hepatitis B virus (HBV) and hepatitis C virus (HCV) were 7.3% and 10.1%, respectively, in NHL, both higher than those in acute leukaemia (1.7% and 2.9%, P < 0.005) and in general Japanese population (1.2% and 2.6%). The incidence of post-chemotherapy liver injury in 25 HBV carriers (36.0%) was significantly higher than that in 539 non-hepatitis virus carriers (12.6%, P = 0.003) and 37 HCV carriers (10.8%, P = 0.026). Liver injury in HBV carriers was more often present in patients who had been treated with steroids than in those without steroids (72.7% and 0%, P = 0.013). After lamivudine became available in our institution, the incidence of liver injury in HBV carriers was reduced from 53.3% to 10.0% (P = 0.041). The therapeutic strategy for haematological malignancies in hepatitis virus carriers should be further investigated. PMID- 15654907 TI - Relatively favorable outcomes of post-transplant pulmonary function in patients with chronic myeloid leukemia receiving non-myeloablative allogeneic hematopoietic stem cell transplantation. AB - Pulmonary function tests were performed in 20 patients with chronic myeloid leukemia before and after human leukocyte antigen-matched allogeneic sibling hematopoietic stem cell transplantation (HSCT) to identify any conditioning treatment effects on post-transplant function from January 1995 to December 2002. Of 20 patients, eight received non-myeloablative conditioning treatment and 12 received conventional myeloablative conditioning treatment. Pulmonary function tests including forced vital capacity (FVC), forced expiratory volume in the first second (FEV1), and diffusion capacity for carbon monoxide (DLCO) were performed pretransplant, 6 and 12 months post-transplant. Possible pre-HSCT and post-HSCT risk factors were evaluated for association with pulmonary function. The results showed that myeloablative conditioning treatment had greater negative impact on FEV1, FVC, and DLCO than non-myeloablative conditioning therapy. We conclude that non-myeloablative allogeneic HSCT may apply a better transplant choice in patients who need special concern with post-transplant pulmonary function changes. PMID- 15654909 TI - Acute polymyositis after donor lymphocyte infusion. AB - Polymyositis usually occurred along with other manifestations of chronic graft versus-host disease (GVHD) after allogeneic hematopoietic stem cell transplantation (HSCT) had been reported. However, polymyositis with a sole manifestation of acute GVHD following donor lymphocyte infusion (DLI) is rare. We reported a 45-yr-old man of acute lymphoid leukemia post-allogeneic HSCT 6 months developed acute polymyositis after DLI. He did not develop any symptoms, signs of acute or chronic GVHD following allogeneic HSCT, despite withdraw of immuosuppresive agents, cyclosporin A (CsA). As the DNA-STR of bone marrow analysis showed mixed chimerism, he received the DLI for remission on May 16, 2002. Acute polymyositis developed following DLI 22 d later. The clinical presentation of polymyositis is compatible with a manifestation of acute GVHD following DLI. It also responds to the treatment of steroid and CsA. PMID- 15654910 TI - Single-agent thalidomide induces response in T-cell lymphoma. AB - T-cell lymphoma is an aggressive lymphoma that cannot be cured despite aggressive therapy, including autologous stem cell transplantation. Thalidomide is an immunomodulatory drug with numerous properties that has proven effective in relapsed multiple myeloma and, to a lesser extent, in other hematologic diseases. We report three cases of relapsed refractory T-cell lymphoma treated with thalidomide with a good tumor response. PMID- 15654911 TI - A case of disseminated Langerhans' cell histiocytosis treated with thalidomide. AB - Tumor necrosis factor alpha (TNF-alpha) seems to play a key role in the pathogenesis of Langerhans' cell histiocytosis (LCH). Thalidomide is an immunomodulator agent of inflammatory cytokines including TNF-alpha. To our knowledge this is the first case of disseminated LCH successfully treated with thalidomide. PMID- 15654912 TI - Autografting followed by a reduced-intensity conditioning unrelated donor cord blood transplantation for a patient with refractory multiple myeloma: successful engraftment with minimal toxicity. AB - We report on a 59-yr-old man with recurrent multiple myeloma. To reduce treatment related mortality, while retaining the cytoreductive effects of high-dose chemotherapy, as well as graft vs. myeloma effect, we used a reduced-intensity conditioning umbilical cord blood (CB) transplantation following high-dose chemotherapy with autologous stem cell transplantation support. This patient was engrafted rapidly and extramedullary toxicities were acceptable. Although he had local recurrence in the right calf on day +130 after the CB transplantation, the tumor was successfully treated with radiation therapy, and he is alive and well at present (day +480). PMID- 15654913 TI - Epidemiologic study on HFE C282Y mutation in Azerbaijan. PMID- 15654914 TI - Human cyclic thrombocytopenia and Anaplasma spp. infection. PMID- 15654916 TI - Mechanisms regulating phosphoinositide 3-kinase signalling in insulin-sensitive tissues. AB - A great deal of evidence has accumulated indicating that the activity of PI 3 kinase is necessary, and in some cases sufficient, for a wide range of insulin's actions in the cell. Most biochemical, genetic and pharmacological studies have focused on identifying potential roles for the class-Ia PI 3-kinases which are rapidly activated following insulin stimulation. However, recent evidence indicates the alpha isoform of class-II PI 3-kinase (PI3K-C2alpha) may also play a role as insulin causes a very rapid activation of this as well. The basic mechanisms by which insulin activates the various members of the PI 3-kinase family are increasingly well understood and these studies reveal multiple mechanisms for modulating the activity and functionality of PI 3-kinase and for down regulating the signals they generate. These include inhibitory phosphorylation events, lipid phosphatases such as PTEN and SHIP2 and inhibitor proteins of the suppressors of cytokine signalling (SOCS) family. The current review will focus on these mechanisms and how defects in these might contribute to the development of insulin resistance. PMID- 15654917 TI - Regional differences of insulin action in adipose tissue: insights from in vivo and in vitro studies. AB - Adipose tissue is now recognized to have a multitude of functions that are of importance in the regulation of energy balance and substrate metabolism. Different hormones, in particular insulin and catecholamines, govern the storage and utilization of energy in the triglyceride depots. In addition, adipocytes produce several different substances with endocrine or paracrine functions, which regulate the overall energetic homeostasis. With excess energy storage, obesity develops, leading to increased risk for type 2 diabetes and cardiovascular disease. The distribution of body fat appears to be even more important than the total amount of fat. Abdominal and, in particular, visceral adiposity is strongly linked to insulin resistance, type 2 diabetes, hypertension and dyslipidaemia, leading to increased risk of cardiovascular disease. The adverse metabolic impact of visceral fat has been attributed to distinct biological properties of adipocytes in this depot compared with other adipose tissue depots. Indeed, regional variations in the metabolic activity of fat cells have been observed. Furthermore, expression studies aiming at defining the unique biological properties of adipose tissues from distinct anatomical sites have identified depot-related differences in the protein content of fat-produced molecules. In this review we wish to summarize important results from the literature and also some recent data from our own work. The main scope is to describe the biological functions of adipose tissue, and to focus on metabolic, hormonal, and signalling differences between fat depots. PMID- 15654918 TI - Lipid metabolism in human skeletal muscle cells: effects of palmitate and chronic hyperglycaemia. AB - This review focuses on the effect of exogenous factors known to be of importance for the development of insulin resistance in differentiated human myotubes. Recent data from our laboratory on the effects of fatty acid pre-treatment and chronic glucose oversupply on fatty acid and glucose metabolism, without and with acute insulin are presented, and discussed in the context of other recent publications in the field. Pre-treatment of myotubes with palmitate, chronic hyperglycaemia, and acute high concentrations of insulin changed fatty acid metabolism in favour of accumulation of intracellular lipids. Acute insulin exposure increased (14)C-oleate uptake and levels of free fatty acids (FFA) and triacylglycerol (TAG). Palmitate pre-treatment further increased oleate uptake, both under basal conditions and in the presence of insulin, with a marked increase in the phospholipid (PL) fraction, with a concomitant reduction in oleate oxidation. Chronic hyperglycaemia also promoted increased lipogenesis and elevated levels of cellular lipids. Changes in fatty acid metabolism in human muscle, in particular fatty acid oxidation, are probably crucial for the molecular mechanism behind skeletal muscle insulin resistance and impaired glucose metabolism. Differentiated human skeletal muscle cells may be an ideal system to further explore the mechanisms regulating lipid metabolism. PMID- 15654919 TI - Mechanisms regulating GLUT4 glucose transporter expression and glucose transport in skeletal muscle. AB - Skeletal muscle is a major glucose-utilizing tissue in the absorptive state and the major glucose transporter expressed in muscle in adulthood is GLUT4. GLUT4 expression is exquisitely regulated in muscle and this seems important in the regulation of insulin-stimulated glucose uptake by this tissues. Thus, muscle GLUT4 overexpression in transgenic animals ameliorates insulin resistance associated with obesity or diabetes. Recent information indicates that glut4 gene transcription is regulated by a number of factors in skeletal muscle that include MEF2, MyoD myogenic proteins, thyroid hormone receptors, Kruppel-like factor KLF15, NF1, Olf-1/Early B cell factor and GEF/HDBP1. In addition, studies in vivo indicate that under normal conditions the activity of the muscle-specific GLUT4 enhancer is low in adult skeletal muscle compared with the maximal potential activity that it can attain at high levels of the MRF transcription factors, MEF2, and TRalpha1. This finding indicates that glut4 transcription may be greatly up-regulated via activation of this enhancer through an increase in the levels of expression or activity of these transcription factors. Understanding the molecular basis of the expression of glut4 will be useful for the appropriate therapeutic design of treatments for insulin-resistant states. The nature of the intracellular signals that mediate the stimulation of glucose transport in response to insulin or exercise is also reviewed. PMID- 15654920 TI - The brown adipose cell: a model for understanding the molecular mechanisms of insulin resistance. AB - Type 2 diabetes mellitus is a complex metabolic disease that occurs when insulin secretion can no longer compensate insulin resistance in peripheral tissues. At the molecular level, insulin resistance correlates with impaired insulin signalling. This review provides new insights into the molecular mechanisms of insulin action and resistance in brown adipose tissue and pinpoints the role of this tissue in the control of glucose homeostasis. Brown adipocytes are target cells for insulin and IGF-I action, especially during late foetal development when insulin supports survival and promotes both adipogenic and thermogenic differentiation. The main pathway involved in insulin induction of adipogenic differentiation, monitored by fatty acid synthase expression, is the cascade insulin receptor substrate (IRS)-1/phosphatidylinositol 3-kinase (PI3K)/Akt. Glucose transport in these cells is maintained mainly by the activity of GLUT4. Acute insulin treatment stimulates glucose transport largely by mediating translocation of GLUT4 to the plasma membrane, involving the activation of IRS 2/PI3K, and the downstream targets Akt and protein kinase C zeta. Tumour necrosis factor (TNF-alpha) caused insulin resistance on glucose uptake by impairing insulin signalling at the level of IRS-2. Activation of stress kinases and phosphatases by this cytokine contribute to insulin resistance. Furthermore, brown adipocytes are also target cells for rosiglitazone action since they show a high expression of peroxisome proliferator activated receptor gamma, and rosiglitazone increased the expression of the thermogenic uncoupling protein 1. Rosiglitazone ameliorates insulin resistance provoked by TNF-alpha, completely restoring insulin-stimulated glucose uptake in parallel to the insulin signalling cascade. Accordingly, foetal brown adipocytes represent a model for investigating insulin action, as well as for the mechanism by which rosiglitazone increase insulin sensitivity under situations that mimic insulin resistance. PMID- 15654921 TI - Cellular mechanisms of insulin resistance, lipodystrophy and atherosclerosis induced by HIV protease inhibitors. AB - Accumulating clinical evidence now links HIV protease inhibitors (HPIs) to the pathogenesis of insulin resistance, dyslipidaemia, lipodystrophy and atherosclerosis associated with highly active anti-retroviral therapy. Here we briefly describe the evidence for a distinct causative role for HPIs, and explore the cellular mechanisms proposed to underlie these side-effects. Acute inhibition of GLUT4-mediated glucose transport, and defective insulin signalling induced by chronic exposure to nelfinavir, are described as cellular mechanisms of insulin resistance. Interference with adipogenesis and adipocyte apoptosis and nelfinavir induced activation of lipolysis are discussed as potential mechanisms of HPI induced lipodystrophy. HPI-induced free radical production, apoptosis and increased glucose utilization in vascular smooth muscle cells are presented as possible novel mechanisms for atherosclerosis. Common pathways and cause-effect relationships between the various cellular mechanisms presented are then discussed, with emphasis on the role of insulin resistance, free radical production and enhanced lipolysis. Understanding the cellular mechanisms of HPI induced side-effects will enhance the search for improved anti-retroviral therapy, and may also shed light on the pathogenesis of common forms of insulin resistance, dyslipidaemia and atherosclerosis. PMID- 15654922 TI - Cytoplasmic and mitochondrial Ca levels in brown adipocytes. AB - AIM: We elucidated the mitochondrial functions of brown adipocytes in intracellular signalling, paying attention to mitochondrial activity and noradrenaline- and forskolin-induced Ca(2+) mobilizations in cold-acclimated rats. METHODS: A confocal laser-scanning microscope of brown adipocytes from warm or cold-acclimated rats was employed using probes rhodamine 123 which is a mitochondria-specific cationic dye, and the cytoplasmic and mitochondrial Ca(2+) probes fluo-3 and rhod-2. X-ray microanalysis was also studied. RESULTS: The signal of rhodamine 123 in the cells was decreased by antimycin A which effect was less in cold-acclimated cells than warm-acclimated cells. Cytoplasmic and mitochondrial Ca(2+) in cold-acclimated brown adipocytes double-loaded with fluo 3 and rhod-2 were measured. Noradrenaline induced the rise in cytoplasmic Ca(2+) ([Ca(2+)](cyto)) followed by mitochondrial Ca(2+) ([Ca(2+)](mito)), the effect being transformed into an increase in [Ca(2+)](cyto) whereas a decrease in [Ca(2+)](mito) by antimycin A or carbonyl cyanide m-chlorophenylhydrazone (CCCP). Antimycin A induced small Ca(2+) release from mitochondria. CCCP induced Ca(2+) release from mitochondria only after the cells were stimulated with noradrenaline. Further, forskolin also elicited an elevation in [Ca(2+)](cyto) followed by [Ca(2+)](mito) in the cells. The Ca measured by X-ray microanalysis was higher both in the cytoplasm and mitochondria whereas K was higher in the mitochondria of cold-acclimated cells in comparison to warm-acclimated cells. CONCLUSIONS: These results suggest that noradrenaline and forskolin evoked an elevation in [Ca(2+)](cyto) followed by [Ca(2+)](mito), in which H(+) gradient across the inner membrane is responsible for the accumulation of calcium on mitochondria. Moreover, cAMP also plays a role in intracellular and mitochondrial Ca(2+) signalling in cold-acclimated brown adipocytes. PMID- 15654923 TI - Effects of renal nerve stimulation on intrarenal blood flow in rats with intact or inactivated NO synthases. AB - AIM: We studied a possible action of nitric oxide (NO), an intrarenal vasodilator, to buffer a decrease in renal perfusion induced by electrical stimulation of renal nerves (RNS). METHODS: In anaesthetized rats RNS was performed (15 V, 2 ms pulse duration) for 10 s at the frequencies of 2, 3.5, 5 and 7.5 Hz. The total renal blood flow (RBF), an index of cortical perfusion, was measured using a Transonic probe on the renal artery. The outer and inner medullary blood flow (OMBF, IMBF) was measured by laser-Doppler flowmetry. The effect of RNS on RBF, OMBF and IMBF was determined in rats which were either untreated or pre-treated with L-NAME (0.6 or 1.8 mg kg(-1) i.v.), or S-methyl thiocitrulline (SMTC, 20 microg kg(-1) min(-1) i.v.), a selective inhibitor of neuronal NO synthase (nNOS). RESULTS: In untreated rats, RNS decreased IMBF significantly less than RBF and OMBF. High-dose L-NAME treatment significantly enhanced the RNS induced decrease of RBF but not of OMBF or IMBF. SMTC treatment significantly enhanced the decrease of IMBF, without affecting the response of RBF or OMBF. CONCLUSION: At intact NO synthesis the inner medullary circulation is not controlled by renal nerves to the extent observed for the outer medulla or cortex. NO generated by all NOS isoforms present in the kidney buffers partly the intrarenal vasoconstriction triggered by electrical RNS. The NO derived from nNOS seems of particular importance in the control of inner medullary perfusion, interacting with NO generated by endothelial NOS and renal nerves. PMID- 15654924 TI - The acute effects of insulin on the cardiorespiratory responses to hypoxia in streptozotocin-induced diabetic rats. AB - AIMS: We examined whether or not streptozotocin (STZ)-induced diabetic rats, which have a lower heart rate (HR, beats min(-1)) than control rats, could maintain hypoxic ventilatory response. METHODS: Twenty-six Wistar rats, which had been injected with STZ (60 mg kg(-1), EXP) or vehicle (0.1 m citrate buffer, CONT) intraperitoneally at 9 weeks of age, had their cardiorespiratory responses to normoxia and 12%O(2) examined after 5 weeks. RESULTS: Compared with CONT rats, EXP rats had a higher blood glucose [24 +/- 3 vs. 5 +/- 1 (mean +/- SD) mmol L( 1)], a lower body weight (320 +/- 23 vs. 432 +/- 24 g), lower HR (303 +/- 49 vs. 380 +/- 44 in normoxia, and 343 +/- 56 vs. 443 +/- 60 in hypoxia) and a lower mean arterial blood pressure (MAP) (89 +/- 6 vs. 102 +/- 10 mmHg in hypoxia). In contrast, both groups had similar values in ventilation (V(E)), V(E)-metabolic rate (MR) ratio and arterial blood gases (ABGs). In EXP rats, with an acute insulin supplement (i.v., 0.75 U h(-1) for 1.5-2 h), not only blood glucose, but also HR, and MAP were normalized as those obtained in CONT rats, and in hypoxia further increased without affecting V(E)-MR ratio and ABGs. Such acute cardiorespiratory stimulating effects of insulin could not be obtained in non diabetic rats (n = 7, 355 +/- 24 g), in which euglycaemia (mean 6.4 mmol L(-1)) was maintained during the measurements. CONCLUSIONS: Our results suggest that, in STZ-induced diabetic rats: (1) ventilation is hardly suppressed by hyperglycaemia, (2) cardiorespiratory responses can be acutely stimulated by short insulin injection, and (3) the effects, including those through acute blood glucose normalization, are possibly specific for the diabetic impairments. PMID- 15654925 TI - Local cold acclimation of the hand impairs thermal responses of the finger without improving hand neuromuscular function. AB - AIM: To investigate the effects of cold acclimation on the thermal response and neuromuscular function of the hand. METHODS: Ten healthy subjects [three female, seven male, age (mean +/- SD): 27.9 +/- 7.9 years] immersed their right hand in 8 degrees C water for 30 min, 5 days a week for 3 weeks. On the first and the last day, neuromuscular function of the first dorsal interosseus (FDI) muscle was tested. RESULTS: There was no significant change in maximal voluntary contraction strength or evoked contractile characteristics of the FDI after cold acclimation. Minimum finger temperature decreased significantly from 10.6 +/- 1.2 to 9.3 +/- 0.8 degrees C after 3 weeks (P < 0.01), with most of the decrease occurring after a single exposure. Mean finger temperature dropped significantly from 14.2 +/- 1.9 to 11.7 +/- 1.4 degrees C following cold acclimation (P < 0.05), with 90% of this adaptation occurring after 5 days. Onset time of cold-induced vasodilatation increased from 446 +/- 171 to 736 +/- 384 s (P < 0.05) and the amplitude decreased from 5.3 +/- 3.2 to 2.5 +/- 2.1 degrees C (P < 0.05). This was significantly different from the control group, who immersed their right hand on the first and last days only. CONCLUSION: These data suggest that cold acclimation does not enhance hand temperature or function but may put the hands at a greater risk of cold injury when exposed to the cold. PMID- 15654926 TI - Integration of mathematical and experimental approaches to resolve insulin signalling. PMID- 15654927 TI - Bipolar disorder and myo-inositol: a review of the magnetic resonance spectroscopy findings. AB - OBJECTIVES: Myo-inositol is an important component of the phosphatidylinositol second messenger system (PI-cycle). Alterations in PI-cycle activity have been suggested to be involved in the pathophysiology and/or treatment of bipolar disorder. More specifically, lithium has been suggested to act primarily by lowering myo-inositol concentrations, the so-called inositol-depletion hypothesis. myo-Inositol concentrations can be measured in vivo with magnetic resonance spectroscopy (MRS). METHODS: The current review primarily examines animal and human MRS studies that evaluated the role of myo-inositol in bipolar illness and treatment. RESULTS: Studies have been carried out in patients who are manic, depressed, and euthymic, both on and off treatment. However, there are several limitations of these studies. CONCLUSIONS: The preclinical and clinical MRS findings were generally supportive of the involvement of myo-inositol in bipolar disorder and its treatment. Overall, in bipolar patients who are manic or depressed there are abnormalities in brain myo-inositol concentrations, with changes in frontal and temporal lobes, as well as the cingulate gyrus and basal ganglia. These abnormalities are not seen in either euthymic patients or healthy controls, possibly due to a normalizing effect of treatment with either lithium or sodium valproate. There is also increasing evidence that sodium valproate may also act upon the PI-cycle. Nonetheless, it remains uncertain if these changes in myo-inositol concentration are primary or secondary. Findings regarding the specific inositol-depletion hypothesis are also generally supportive in acutely ill patients, although it is not yet possible to definitively confirm or refute this hypothesis based on the current MRS evidence. PMID- 15654928 TI - Bipolar II disorder: a review. AB - OBJECTIVES: To review the current knowledge of bipolar II disorder. METHODS: Literature was reviewed after conducting a Medline search and a hand search of relevant literature. RESULTS: Bipolar II disorder is a common disorder, with a prevalence of approximately 3-5%. Distinct clinical features of bipolar II disorder have been described. The key to diagnosis is the recognition of past hypomania, while depression is the typical presenting feature of the illness. This is responsible for a significant rate of missed diagnosis, and consequent management according to unipolar guidelines. It is unclear if bipolar II disorder is over-represented amongst resistant depression populations and if abrupt offset of antidepressant action is a phenomenon over represented in bipolar II disorder, reflecting induction of predominantly depressive cycling. A few mood-stabilizer studies available provide provisional suggestion of utility. A supportive role for psychosocial therapies is suggested, however, there is a sparsity of published studies specific to bipolar II disorder cohorts. A small number of short-term antidepressant trials have suggested efficacy, however, compelling long-term maintenance data is absent. CONCLUSIONS: An emerging literature on the specific clinical signature and management of the disorder exists, however, this is disproportionately small relative to the epidemiology and clinical significance of the disorder. PMID- 15654929 TI - Psychological therapies for bipolar disorder: the role of model-driven approaches to therapy integration. AB - OBJECTIVES: The psychological and social aspects of bipolar disorder are receiving increasing recognition. Recently, there have been promising developments in psychological interventions, but there is scope for further improvement of therapeutic outcomes. This paper argues for the use of more detailed psychological models of bipolar disorder to inform the further development of therapeutic approaches. METHOD: Evidence for psychological, family and social factors in bipolar disorder is reviewed. The efficacy of current individual and family interventions are discussed. A model, which has potential to synthesize group and individual approaches, is outlined. RESULTS: Psychological, social and family factors have important influences on the onset, course and outcome of bipolar disorder. Interventions based on vulnerability stress models have proved effective. However, to enhance efficacy future developments need to be based on models that integrate current understandings of the multiple levels at which mood fluctuations occur. A particular recent model is discussed which leads to specific proposals for future intervention research. CONCLUSIONS: Psychological and family approaches to BD have much potential. They clearly have a role in conjunction with appropriate pharmacological treatment. If this potential is to be fully realized future developments need to be based on psychological models that can accommodate the complexity of this illness. PMID- 15654930 TI - Comparative analysis of the effects of four mood stabilizers in SH-SY5Y cells and in primary neurons. AB - OBJECTIVES: The mood-stabilizing drug valproic acid (VPA) exerts a neurotrophic effect on the human neuroblastoma cell line, SH-SY5Y. We aimed to establish whether other mood-stabilizing drugs have a similar action and which signalling pathways mediate this process. METHODS: We analysed the effects of the mood stabilizers VPA, lithium, carbamazepine and lamotrigine on proliferation, survival, neurite outgrowth and extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) activation using the SH-SY5Y cell line. We also compared their effects in primary neurons. RESULTS: We found that VPA promotes neurite outgrowth and prevents cell death in SH-SY5Y cells, but has no effect on cell proliferation. This neurotrophic effect does not involve inhibition of glycogen synthase kinase-3, histone deacetylase or prolyl oligopeptidase; the effect also does not seem to involve protein kinase C. In contrast, VPA activates ERK/MAPK and the survival effect of VPA is blocked by inhibition of the ERK/MAPK signalling pathway. Moreover, other activators of ERK/MAPK, such as epidermal growth factor and phorbol 12-myristate 13-acetate, mimic the neurotrophic effects of VPA. Other mood stabilizers do not activate ERK/MAPK and do not promote neurite outgrowth or survival of SH-SY5Y cells. In contrast, both lithium and VPA activate ERK/MAPK in rat primary cortical neurons. CONCLUSIONS: We investigated four mood stabilizers that are effective in the treatment of bipolar disorder. Our results suggest that, while some mood stabilizers may have additional neuroprotective effects, activation of ERK/MAPK does not appear to be a mechanism common to all mood-stabilizing drugs. PMID- 15654931 TI - Interaction of calbindin D28k and inositol monophosphatase in human postmortem cortex: possible implications for bipolar disorder. AB - OBJECTIVES: Therapeutically relevant concentrations of lithium (Li) exert an uncompetitive inhibition on inositol monophosphatase (IMPase). It has recently been shown that calbindin D28k (calbindin) activates IMPase. Purified calbindin attaches to a specific amino acid sequence on purified IMPase enhancing its activity by several hundred fold. We studied whether calbindin activates IMPase in postmortem human brain crude homogenate, whether differences in calbindin levels between lymphocytes and brain may be responsible for our previous finding of reduced IMPase activity in lymphocytes but not brain of bipolar patients, and whether calbindin protein levels are altered in postmortem brain from bipolar patients versus control subjects and schizophrenic and major depressive patients. METHODS: IMPase activity in human postmortem brain specimens with or without 10 microM human recombinant calbindin was quantified spectrophotometrically in an enzyme-linked immunosorbent assay (ELISA) reader. Calbindin protein levels in postmortem brain were determined using Western blot analysis. RESULTS: Supplementation of human recombinant calbindin to postmortem human brain crude homogenate enhanced IMPase activity by 3.5-fold. No difference in postmortem temporal cortex calbindin protein levels was found between bipolar patients versus comparison groups. Two-fold higher calbindin protein levels were found in Li-treated bipolar patients compared with other bipolar patients. Subchronic Li treatment in mice did not affect brain calbindin protein levels significantly. Chronic Li treatment reduced calbindin protein levels in the frontal cortex but not in the hippocampus. CONCLUSIONS: Calbindin is a physiological activator of IMPase in human brain. Protein levels of calbindin are not altered in postmortem temporal cortex of bipolar patients. PMID- 15654932 TI - Content-specificity of dysfunctional cognitions for patients with bipolar mania versus unipolar depression: a preliminary study. AB - OBJECTIVE: Dysfunctional beliefs or cognitions are considered to be fundamental to both the phenomenology and pathogenesis of depression. However, the cognitive aspects of mania have not been as thoroughly investigated. We sought to compare the maladaptive beliefs and cognitions of 23 bipolar manic or hypomanic patients, 28 patients with unipolar major depression, and 24 normal adults. METHOD: The Cognition Checklist for Mania (CCL-M) was used to assess the beliefs. This 61 item self-report instrument is scored for seven subscales measuring (a) self importance, (b) interpersonal grandiosity, (c) inappropriate spending, (d) excitement and risk-taking, (e) interpersonal frustrations, (f) goal-driven activity, and (g) past or future outlooks on life, and also yields a total score. RESULTS: The mean CCL-M total score of the bipolar-manic patients was significantly higher than the mean CCL-M total score of the unipolar-depressed patients, and the patients' mean CCL-M total score was also higher than that of the normal adults. The mean scores of the subscales measuring excitement and past and future memories and expectations were also significantly higher for the bipolar-manic than unipolar-depressed patients. CONCLUSIONS: Bipolar-manic patients endorse with maladaptive beliefs and cognitions that are associated with mania more than do unipolar-depressed patients and normal adults. The implications for the early identification of cognitions associated with prodromal states of mania, and for psychotherapeutic interventions, are discussed. PMID- 15654933 TI - Neuropsychological deficits in adolescents with conduct disorder and comorbid bipolar disorder: a pilot study. AB - OBJECTIVE: We report pilot data on neuropsychological deficits in aggressive juvenile offenders with and without bipolar disorder compared with each other and healthy controls. METHOD: We assessed 52 adolescents and their parent or guardians: 36 incarcerated juvenile offenders and 16 community controls using the Schedule for Affective Disorders and Schizophrenia for School Age Children, Present and Life-Time Version and a neuropsychological testing battery. All incarcerated subjects (n=34) met criteria for Conduct Disorder (CD); 26 are classified as Non-BD-CD, and eight with CD and Bipolar disorder (CD-BD). These subjects were compared to community controls (n=16) matched for age, gender, SES and ethnicity. RESULTS: Relative to controls, the Non-BD-CD subjects' impairments (p<0.05) were in cognitive ability, set shifting/inhibition, planning and verbal memory-language functioning. The CD-BD group displayed impairments (p<0.05) relative to controls in cognitive ability, set shifting, verbal memory-language functioning, and visuospatial tasks. The Non-BD-CD and CD-BD groups however did not display significant differences on most neuropsychological measures compared with each other. When we controlled for Attention Deficit Hyperactivity Disorder, the Non-CD-BP subjects continued to show deficits on Verbal measures where the CD BD subjects maintained deficits in measures of cognitive ability, verbal measures and visual spatial tests. CONCLUSIONS: Juvenile offender with CD displayed a wide range of deficits on neuropsychological testing compared with controls. Although juvenile offenders with and without BD differed on their clinical presentation, differences on neuropsychological measures are not specific and may be related to comorbid diagnoses. PMID- 15654934 TI - Recovery and functional outcomes following olanzapine treatment for bipolar I mania. AB - BACKGROUND: Typical experimental categorizations of treatment responses in bipolar disorder (BPD) patients may have limited relationship to clinical recovery or functional status, and there is inadequate research on such clinically important outcomes. METHODS: We analyzed data from a study of open continuation of olanzapine treatment following a 3-week placebo-controlled trial involving initially hospitalized adult subjects with DSM-IV BP-I mania to estimate rates and times to symptomatic remission (low scores on standardized symptomatic assessments) and clinical recovery (remission sustained>or=8 weeks), associated clinical factors, and functional outcomes. RESULTS: During treatment with olanzapine for 27.9+/-20.1 weeks, symptomatic remission was attained by 70% of subjects, half by 8 weeks (95% CI 6-10) weeks, and later lost by 82% of remitted subjects; remitted (versus non-remitted) subjects had slightly lower baseline clinical global impression scores and greater trial-completion. Sustained clinical recovery was attained by only 40 of 113 (35%) of subjects, half by 36 (95% CI 20-40) weeks, and later lost by 45%. Subjects with above median (>12) initial Hamilton-Depression rating scale depression scores were half as likely to recover (p=0.016) and did so much later (36 versus 12 weeks) than those with lower scores. At final assessment, self-rated well being (SF-36 psychosocial functioning scores) improved substantially more among recovered versus non-recovered subjects (mean changes: 87% versus 23%), and two-thirds of recovered subjects remained unemployed-for-pay while half received disability compensation. CONCLUSIONS: Clinically meaningful symptomatic remission and recovery in relatively severely ill adult bipolar I manic patients were achieved slowly and sustained by only some patients within an average of 7 months of continuous treatment. These clinically relevant outcomes were worse with relatively high initial dysphoria ratings. Well-being was rated higher by recovered subjects, but their ability to work and live independently were markedly impaired. These findings underscore the emerging view that BPD can often be severe, slow to remit, and disabling, particularly in association with prominent depression-dysphoria symptoms. Improved treatments for BPD are needed, guided by longitudinal assessments of clinically meaningful measures of symptomatic recovery and functional outcome. PMID- 15654935 TI - Increased risk for pulmonary embolism in patients with bipolar disorder. AB - OBJECTIVES: The purpose of this study was to investigate the increased risk for pulmonary embolism (PE) in patients with manic-depressive psychosis/bipolar disorder (BD). Affective patients show increased mortality compared with the background population. METHODS: A register study was carried out in which somatic and psychiatric information in 25,834 BD patients and 117,815 controls was extracted from The Danish Psychiatric Central Research Register, The National Register of Patients, The Danish Central Person Register and The Danish Register of Causes of Death, with similar information about patients with schizophrenia and anxiety for comparison. RESULTS: Patients with BD had a significantly increased occurrence of PE [increased incidence rate ratio (IRR)=1.61; 95% confidence interval (CI) (1.38, 1.88)]. An association was also found in schizophrenic patients [IRR=1.78; 95% CI (1.27, 2.51)] and in anxiety patients [IRR=1.49; 95% CI (1.10, 2.02)]. CONCLUSIONS: Increased occurrence of PE in patients with BD is one of the explanations of increased mortality in the affective patient group. A similar finding in females with schizophrenia and females with anxiety suggests 'mental disorder' as the risk factor for PE. The causes for the increased occurrence of PE in BD patients (and other diagnostic groups) need further investigation. PMID- 15654936 TI - African-American participants in a bipolar disorder registry: clinical and treatment characteristics. AB - OBJECTIVES: The goal of this paper was to compare clinical characteristics and treatment history of African-American and Caucasian participants in a bipolar disorder registry. METHODS: The Western Pennsylvania Bipolar Disorder Registry used several recruitment methods to reach individuals self-identified as having bipolar disorder. Individuals who contacted and joined the registry completed an interviewer-administered questionnaire on clinical characteristics and treatment history. A sample of 2,718 registry participants was analyzed in order to compare these characteristics and history by race. RESULTS: African-Americans in the registry reported a greater number of inpatient hospitalizations (9.8 versus 4.4) than Caucasians, as well as a higher suicide attempt rate (64% versus 49%). African-American participants were more likely to report a family member with schizophrenia. With respect to psychotropic medication, African-Americans were less likely to report taking antimanic medication or benzodiazepines, but more likely to report taking antipsychotics than Caucasians. CONCLUSIONS: The present findings reinforce previous reports regarding the chronicity and severity of bipolar disorder among African-Americans. They also support previous studies that found high rates of attempted suicide among African-Americans with bipolar disorder. These findings provide further impetus for specific community and mental health services delivery efforts to reduce barriers to early accurate diagnosis and to appropriate ambulatory treatment for bipolar disorder. PMID- 15654937 TI - Clinical, psychosocial, and treatment differences in minority patients with bipolar disorder. AB - OBJECTIVES: The clinical profile of minorities with bipolar disorder has been largely unexplored. We compared the clinical (e.g. psychiatric and substance use comorbidity), psychosocial, and treatment characteristics between white and minority patients with bipolar disorder (minorities were defined as black or other minority, which included Hispanic, Asian-American, or Native-Americans). METHODS: We collected demographic, diagnosis, and treatment information using the Structured Clinical Interview for DSM-IV (SCID) from 330 inpatients with a current major affective episode across 11 Veterans Affairs (VA) Medical Centers enrolled in the VA Cooperative Study (Reducing the Efficacy-Effectiveness Gap in Bipolar Disorder). RESULTS: Twenty-four percent (n=80) were minority; 9% (n=30) were women, 4% (n=20) were >or=65 years old; and the majority (87%, n=286) had bipolar type I. Minorities compared with whites were no more likely to have a current episode of psychosis (30% versus 37%, respectively; p=0.28). However, minorities were more likely than whites to have a cocaine use disorder (adjusted odd's ratio, OR=2.2; 95% CI: 1.4-3.5; p<0.01) or current alcohol abuse disorder (adjusted OR=1.8; 95% CI: 1.1-3.9;p<0.05). Further breakdown by race/ethnicity revealed that cocaine use disorder was most prevalent among blacks (n=14, 29%), compared with all other minorities (n=2, 6%) or whites (n=10, 4%; p<0.001). Other minorities compared with blacks or whites were more likely involuntarily committed during some part of their index hospitalization (adjusted OR=2.47; 95% CI: 1.1-5.7; p=0.04). CONCLUSIONS: Minorities with bipolar disorder may be a more vulnerable population because of higher rates of substance use disorder and higher rates of involuntary psychiatric commitment. Moreover, the specific profile of vulnerability may differ across minority groups. PMID- 15654938 TI - Dark therapy for mania: a pilot study. AB - BACKGROUND: Recent findings suggest that extended bed rest and darkness could stabilize mood swings in rapid cycling bipolar patients. METHOD: We exposed 16 bipolar inpatients affected by a manic episode to a regimen of 14 h of enforced darkness from 6 p.m. to 8 a.m. each night for three consecutive days [dark therapy (DT)]. Pattern of mood changes were recorded with the Young Mania Rating Scale (YMRS) and compared with a control group of 16 inpatients matched for age, sex, age at onset, number of previous illness episodes and duration of current episode, and were treated with therapy as usual (TAU). RESULTS: Adding DT to TAU resulted in a significantly faster decrease of YMRS scores when patients were treated within 2 weeks from the onset of the current manic episode. When duration of current episode was longer, DT had no effect. Follow-up confirmed that good responders needed a lower dose of antimanic drugs and were discharged earlier from the hospital. CONCLUSIONS: Chronobiological interventions and control of environmental stimuli can be a useful add-on for the treatment of acute mania in a hospital setting. PMID- 15654940 TI - Extracellular 14-3-3sigma protein: a potential mediator of epithelial-mesenchymal interactions. PMID- 15654944 TI - Multiple pathways driving IgE production and chronic dermatitis in mice: a model for atopic dermatitis? PMID- 15654945 TI - Breaking tolerance--another piece added to the vitiligo puzzle. PMID- 15654946 TI - Pathogenesis of photosensitivity in the cutaneous porphyrias. PMID- 15654947 TI - Changing paradigms in cutaneous immunology: adapting with dendritic cells. PMID- 15654948 TI - Hair follicle pigmentation. AB - Hair shaft melanin components (eu- or/and pheomelanin) are a long-lived record of precise interactions in the hair follicle pigmentary unit, e.g., between follicular melanocytes, keratinocytes, and dermal papilla fibroblasts. Follicular melanogenesis (FM) involves sequentially the melanogenic activity of follicular melanocytes, the transfer of melanin granules into cortical and medulla keratinocytes, and the formation of pigmented hair shafts. This activity is in turn regulated by an array of enzymes, structural and regulatory proteins, transporters, and receptors and their ligands, acting on the developmental stages, cellular, and hair follicle levels. FM is stringently coupled to the anagen stage of the hair cycle, being switched-off in catagen to remain absent through telogen. At the organ level FM is precisely coupled to the life cycle of melanocytes with changes in their compartmental distribution and accelerated melanoblast/melanocyte differentiation with enhanced secretory activity. The melanocyte compartments in the upper hair follicle also provides a reservoir for the repigmentation of epidermis and, for the cyclic formation of new anagen hair bulbs. Melanin synthesis and pigment transfer to bulb keratinocytes are dependent on the availability of melanin precursors, and regulation by signal transduction pathways intrinsic to skin and hair follicle, which are both receptor dependent and independent, act through auto-, para- or intracrine mechanisms and can be modified by hormonal signals. The important regulators are MC1 receptor its and adrenocorticotropic hormone, melanocyte stimulating hormone, agouti protein ligands (in rodents), c-Kit, and the endothelin receptors with their ligands. Melanin itself has a wide range of bioactivities that extend far beyond its determination of hair color. PMID- 15654949 TI - Gender differences in mouse skin morphology and specific effects of sex steroids and dehydroepiandrosterone. AB - Sex steroids play an important role in skin morphology and physiology. To evaluate the specific effects of sex steroids, the thickness of each skin layer was measured in intact and gonadectomized (GDX) male and female mice, as well as in GDX animals treated for 3 wk with 17beta-estradiol (E2), dihydrotestosterone (DHT), or their precursor dehydroepiandrosterone (DHEA). Morphological analysis shows that the dorsal skin of intact male is thicker than in the female, whereas the epidermis and hypodermis are thicker in the female. After GDX, epidermal thickness decreases only in the female to become similar to that of the intact male. Epidermal thickness in GDX animals of both sexes increases after E2 treatment to a value similar to that of intact females, whereas an increase is observed only in females after DHEA treatment. Both DHEA and DHT increased dermal thickness whereas E2, DHT, and DHEA markedly reduced hypodermal thickness in GDX animals of both sexes. Under all conditions, the hypodermis remains thicker in females. GDX triggers a rapid hair growth from telogen to anagen with a thicker hair shaft diameter in females. This data shows that DHT, E2, and DHEA exert specific effects on the different skin layers and appendages. PMID- 15654950 TI - Interleukin-6-type cytokines upregulate expression of multidrug resistance associated proteins in NHEK and dermal fibroblasts. AB - Normal human epidermal keratinocytes (NHEK) and dermal fibroblasts express a cell specific pattern of efflux transport proteins. Since regulatory mechanisms for these transporters in cells of the human skin were unknown, we analyzed the influence of inflammatory cytokines on the expression of multidrug resistance associated proteins (MRP1, 3, 4, 5). Using real-time PCR, RT-PCR, cDNA microarray, immunostaining and efflux assays we demonstrated that stimulation of NHEK and primary human dermal fibroblasts with interleukin-6 (IL-6), in combination with its soluble alpha-receptor, or oncostatin M (OSM) for 24-72 h resulted in an upregulation of MRP expression and activity. Both cytokines induced a strong activation of signal transducer and activator of transcription (STAT)1 and STAT3 as well as the mitogen-activated protein kinase (MAPK) Erk1/2. OSM additionally activated proteinkinase B strongly. Using the MAPK/extracellular signal-regulated kinase kinase 1-specific inhibitor U0126 we could exclude a stimulatory effect of MAPK on MRP gene expression. Inhibition of the phosphatidylinositol 3-kinase, however, indicated that this pathway might be involved of OSM-mediated upregulation of MRP4 in dermal fibroblasts. Several inflammatory skin diseases show an enhanced expression of IL-6-type cytokines. Correspondingly, upregulation of MRP expression was found in lesional skin taken from patients with psoriasis and lichen planus. PMID- 15654952 TI - Breaking the connection: caspase 6 disconnects intermediate filament-binding domain of periplakin from its actin-binding N-terminal region. AB - Periplakin is a member of the plakin family of cytolinkers that connect cytoskeletal networks to each other as well as to the cell junctional complexes. Here, we demonstrate a direct molecular interaction between actin and periplakin. Furthermore, the oligomerization state of periplakin was shown to determine specificity of its binding to intermediate filaments (IF) in vitro. Both the filament association and the cell membrane localization of periplakin were confirmed in the cells overexpressing human periplakin. Double labeling of the N- and C-terminally tagged periplakin revealed unexpected lack of co-localization of periplakin ends in a confluent culture, and separation of the periplakin ends was even more pronounced in apoptotic cells. Western analysis revealed that after induction of apoptosis, periplakin becomes cleaved close to its C-terminal tail. Only the distinct cleavage products, but not the full-length periplakin, were present in the cells detached from the solid support during the apoptotic process. We show that caspase 6 cleaves periplakin at an unconventional recognition site, amino acid sequence TVAD. Thus, the separation of periplakin ends disconnects the actin-binding head-rod domain from the IF-binding C-terminal domain. We show that specific cleavage products co-exist with the full-length periplakin in cells, suggesting physiological consequences due to their altered binding specificities. PMID- 15654951 TI - Agonists of proteinase-activated receptor-2 stimulate upregulation of intercellular cell adhesion molecule-1 in primary human keratinocytes via activation of NF-kappa B. AB - Proteinase-activated receptor-2 (PAR2) belongs to a new G protein-coupled receptor subfamily that is activated by various serine proteases. Recent knowledge indicates that PAR2 is involved in cutaneous inflammation and immune response. PAR2 is highly expressed by human keratinocytes (KTC). The underlying mechanisms of PAR2-mediated KTC function and cutaneous immune response are, however, still incomplete. Therefore, we investigated the activation of important signaling cascades in primary human KTC after PAR2-stimulation using specific agonists. Moreover, we compared PAR2-immunoreactivity in the epidermis of inflammatory dermatoses and normal human skin. Electrophoretic mobility shift assays and morphological transduction studies revealed PAR2-induced activation and translocation of nuclear factor kappa B (NF-kappaB) in primary human KTC with a maximum after 1 h. Supershift analysis demonstrated acivation of the p50/p65 heterodimer complex. PAR2 agonists also induced upregulation of intercellular adhesion molecule-1 (ICAM-1) RNA, as shown by RT-PCR. Use of NF-kappaB inhibitors prevented upregulation of the cell adhesion molecule ICAM-1 in KTC indicating a direct role of NF-kappaB in PAR2-mediated upregulation of ICAM-1. Fluorescence activated cell sorter analysis confirmed PAR2-induced and NF-kappaB-mediated upregulation of ICAM-1 protein after 13 h. Moreover, increased expression of PAR2 was detected in KTC of patients with atopic dermatitis suggesting a role of PAR2 in human skin inflammation. In conclusion, PAR2 induces upregulation of cell adhesion molecules such as ICAM-1 in primary human KTC via NF-kappaB activation, and is upregulated in KTC during cutaneous inflammation. Thus, PAR2 may play an important regulatory role of human KTC during inflammation and immune response. PMID- 15654954 TI - Proteome analysis of skin distinguishes acute guttate from chronic plaque psoriasis. AB - Psoriasis is a disease with considerable heterogeneity in clinical presentation. This is the first study using two-dimensional gel electrophoresis to compare global protein expression patterns in lesional and non-lesional skin from subjects with acute guttate psoriasis associated with streptococcal throat infection and chronic plaque psoriasis. Samples from experimentally induced contact eczema and normal skin from healthy controls were also included. Proteins with statistically significant differences in expression were used in hierarchical cluster analyses resulting in separation of the different samples into groups. Chronic plaque and guttate psoriasis samples were distinctly separated, indicating that they represent discrete phenotypes at the protein expression level. Interestingly, there was a trend in which guttate psoriasis lesions clustered closer to eczema than to chronic plaque psoriasis lesions, indicating that the duration of the inflammatory reaction may affect clustering. Several of the differentially expressed proteins were identified by mass spectrometry. PMID- 15654953 TI - The effect of sun exposure in determining nevus density in UK adolescent twins. AB - We report a study of 221 teenage twin pairs to examine the genetic and environmental determinants of nevi representing the most potent phenotypic risk factor for melanoma. Our published heritability analysis estimated that nevi are mainly genetically determined. In this paper we examine the role of sun exposure. We report a correlation between nevus density and sun exposure, particularly that acquired in hotter countries than in the UK (mean nevus density 41 per m2 in those in the highest quartile of exposure vs 24 per m2 in those with no exposure, p<0.0001). We were not able to demonstrate a protective effect for either sun protection cream or shirt wearing. By including phenotypic variables and reported sun exposure into the heritability analysis, we conclude that 66% of the total variance of nevus count is attributable to genetic effects: 7% associated to eye color, 6% to hair color, and 1% to reported skin type, which leaves 52% as to yet unidentified genetic factors. Of the 25% of variation attributable to environmental influences, one-third is estimated to be because of sun exposure on hot holidays. PMID- 15654955 TI - Heat modulation of tropoelastin, fibrillin-1, and matrix metalloproteinase-12 in human skin in vivo. AB - Photoaged skin contains elastotic materials in the upper reticular dermis. This phenomenon is commonly known as solar elastosis. In this study, we investigated the effects of heat on the expression of tropoelastin and fibrillin-1, two main components of elastic fibers, and on matrix metalloproteinase (MMP)-12, the most active MMP against elastin, in human skin in vivo. Heat was found to increase tropoelastin mRNA and protein expression in the epidermis and in the dermis. Fibrillin-1 mRNA and protein expression were increased by heat in the epidermis, but were decreased in the dermis. We found that pre-treatment of skin with N acetyl cysteine or genistein for 24 h prior to heat treatment inhibited the heat induced expression of tropoelastin, but not of fibrillin-1. These data indicate that reactive oxygen species may play a role in tropoelastin expression by heat, but not in fibrillin-1 expression. We also found that heat treatment increases MMP-12 mRNA and protein expression in human skin. Our results suggest that the abnormal production of tropoelastin and fibrillin by heat in human skin and that their degradation by various MMP, such as MMP-12, may contribute to the accumulation of elastotic material in photoaged skin. PMID- 15654957 TI - DNA-based diagnosis of xeroderma pigmentosum group C by Whole-genome scan using single-nucleotide polymorphism microarray. AB - In this study, we have established the molecular basis of xeroderma pigmentosum (XP) in two unrelated Chinese families. In the first patient with consanguineous parents, we mapped the disease-causing locus XPC using single-nucleotide polymorphism microarray. Mutational analysis of the XPC gene showed that the patient is homozygous for a nonsense mutation, E149X. After developing DNA-based diagnosis of XPC, we screened another XP patient for XPC mutations. We found that the second patient is a compound heterozygote of 1209delG and Q554X in this gene. These are the first XPC-causing mutations identified in Chinese patients. PMID- 15654956 TI - Basement membrane reconstruction in human skin equivalents is regulated by fibroblasts and/or exogenously activated keratinocytes. AB - This study was undertaken to examine the role fibroblasts play in the formation of the basement membrane (BM) in human skin equivalents. For this purpose, keratinocytes were seeded on top of fibroblast-free or fibroblast-populated collagen matrix or de-epidermized dermis and cultured in the absence of serum and exogenous growth factors. The expression of various BM components was analyzed on the protein and mRNA level. Irrespective of the presence or absence of fibroblasts, keratin 14, hemidesmosomal proteins plectin, BP230 and BP180, and integrins alpha1beta1, alpha2beta1, alpha3beta1, and alpha6beta4 were expressed but laminin 1 was absent. Only in the presence of fibroblasts or of various growth factors, laminin 5 and laminin 10/11, nidogen, uncein, type IV and type VII collagen were decorating the dermal/epidermal junction. These findings indicate that the attachment of basal keratinocytes to the dermal matrix is most likely mediated by integrins alpha1beta1 and alpha2beta1, and not by laminins that bind to integrin alpha6beta4 and that the epithelial-mesenchymal cross-talk plays an important role in synthesis and deposition of various BM components. PMID- 15654958 TI - The -2518 promotor polymorphism in the MCP-1 gene is associated with systemic sclerosis. AB - Factors influencing the initiation or progression of sclerosis in patients with systemic sclerosis (SSc) are poorly understood. Monocyte chemotactic protein-1 (MCP-1) is a potent chemokine, which is upregulated in fibroblasts during development of sclerosis. In this study, we investigated the frequency of the functional -2518G MCP-1 promoter polymorphism in 18 patients with SSc and 139 healthy controls. In the lesional skin of the same SSc patients, expression of MCP-1 protein was examined by immunohistochemistry. To investigate a genotype/phenotype correlation, basal as well as tumor necrosis factor (TNF) induced MCP-1 expression was analyzed in fibroblasts isolated from the skin of SSc patients with different MCP-1 genotypes by quantitative RT-PCR and ELISA. Genotyping for the -2518 (A/G) MCP-1 promotor polymorphism showed that GG homozygotes were significantly more frequent in patients with SSc than in controls (28%vs 6%). Results of immunohistochemistry revealed that MCP-1 was expressed in keratinocytes, infiltrating inflammatory cells, fibroblasts, and endothelial cells in scleroderma skin, whereas normal control skin showed no MCP 1 expression. MCP-1 expression in fibroblasts from GG-homozygote individuals tended to be stronger as compared to AG or AA genotypes. Furthermore, basal as well as TNF-induced MCP-1 expression of fibroblasts isolated from a GG-homozygote SSc patient was significantly higher than MCP-1 expression of fibroblasts isolated from heterozygote or AA-homozygote donors. The A -2518G polymorphism of the MCP-1 gene appears to affect MCP-1 expression of skin fibroblasts of patients with SSc. In accordance, the G/G genotype may predispose patients to SSc. PMID- 15654959 TI - Haplotype sharing analysis identifies a retroviral dUTPase as candidate susceptibility gene for psoriasis. AB - The psoriasis susceptibility locus 1 (PSORS1) mutation is assumed to reside within a region around human leukocyte antigen-C spanning 250 kb, termed risk haplotype (RH) 1/2. By re-analyzing a published data set with a previously developed method, the haplotype sharing statistic, we confirm localization of PSORS1 to the RH1 region and refine its location to marker M6S168. We replicate this result in an independent patient sample. The target region harbors fragments of a human endogenous retrovirus K (HERV-K) endogenous retrovirus. Two single nucleotide polymorphisms with alleles differing between high- and low-risk haplotypes are located within the HERV-K dUTPase. One of these encodes a predicted non-conserved Glu-Arg exchange. The HERV-K dUTPase is expressed in peripheral blood and in normal as well as lesional psoriatic skin. Our results indicate that an endogenous retroviral dUTPase constitutes a candidate gene for the PSORS1 mutation. PMID- 15654960 TI - The major psoriasis susceptibility locus PSORS1 is not a risk factor for late onset psoriasis. AB - PSORS1 is the major susceptibility locus for psoriasis vulgaris (PV) and lies within an approximately 200 kb segment of the major histocompatibility complex on chromosome 6p21.3. Alleles of candidate genes in this region including human leukocyte antigen (HLA)-C, alpha-helical coiled coil rod (HCR), and corneodesmosin (CDSN) show association with early-onset PV. Late-onset psoriasis (LOP) is defined as a disease with onset after 40 y of age and is typically sporadic. We assessed the role of PSORS1 in genetic susceptibility to LOP. Genotyping for HLA-C alleles and seven single nucleotide polymorphisms (SNP) within the genes HCR and CDSN was performed in LOP (n=145) and normal controls (n=309). Statistical analysis of allelic frequencies included calculation of odds ratio and chi2 comparisons. LOP demonstrated only a weak association to PSORS1 alleles HLA-Cw*6 (p=0.037), CDSN*5 (p=0.041), HCR*WC (p=0.013), and HCR SNP +325 (p=0.038). Patients with age of onset for psoriasis of 50 y or above provided no evidence of association with any of these alleles. These data suggest that the study cohort may include a number of subjects who harbor PSORS1 predisposition to early-onset psoriasis and yet do not present with disease by the age of 40 y. Thus this study demonstrates that PSORS1 is not a major inherited risk factor in the pathogenesis of LOP. These data suggest that the exclusion of LOP subjects from case-control studies will aid further delineation of the PSORS1 locus. Future genome-wide studies will be required to identify loci conferring risk for late-onset disease. PMID- 15654961 TI - Lack of evidence for genetic association to RUNX1 binding site at PSORS2 in different German psoriasis cohorts. AB - A DNA variant, rs734232, altering a RUNX1 binding site was recently reported as susceptibility allele at PSORS2 (17q25) in cohorts of psoriasis patients from the US. A testing of this variant in psoriasis patients from Germany did not confirm this association in 300 trios nor in two case-control studies with 281 patients with psoriasis vulgaris and 375 patients with psoriatic arthritis, respectively. These results fail to support rs734232 as a psoriasis susceptibility factor in German psoriasis patients. PMID- 15654962 TI - Plectin gene mutations can cause epidermolysis bullosa with pyloric atresia. AB - Epidermolysis bullosa with pyloric atresia (EB-PA), manifesting with neonatal blistering and gastric anomalies, is known to be caused by mutations in the hemidesmosomal genes ITGA6 and ITGB4, which encode the alpha6 and beta4 integrin polypeptides, respectively. As part of our molecular diagnostics program, we have now encountered four families with EB-PA in which no mutations could be identified in these two genes. Instead, PCR amplification followed by heteroduplex scanning and/or direct nucleotide sequencing revealed homozygous mutations in the plectin gene (PLEC1), encoding another hemidesmosomal protein previously linked to EB with muscular dystrophy. Our findings provide evidence for additional molecular heterogeneity in EB, and emphasize the importance of screening EB-PA patients not only for alpha6beta4 integrin but also for plectin deficiency. PMID- 15654963 TI - TGFbeta inhibits CD1d expression on dendritic cells. AB - The CD1 family of cell surface glycoprotein has been demonstrated to be a third lineage of antigen-presenting molecules for specific T cell responses. They present lipidic, glycolipidic antigen and hydrophobic peptide to T cells. CD1d restricted T cells play a role in autoimmune disease and in tumor immunity. Transforming growth factor beta (TGFbeta), a member of the family of polypeptide growth factors synthetized by human keratinocytes, has inhibitory effects on proliferation and differentiation of immune cells, especially on CD1d-restricted natural killer T cells. These properties led us to investigate the role of TGFbeta in CD1d expression on dendritic cells (DC), which are known to play a key role in initiation of the immune response. Here, we observed CD1d molecules on DC developed from PBMC with GM-CSF and IL4 but not with GM-CSF, IL4 and TGFbeta for 7 d. RT-PCR and FACS analysis (mAb 42.1) performed at various stages of differentiation on CD34+ HPC show that CD1d mRNA levels and CD1d molecule expression at the cell surface decreased progressively during the differentiation process. Thus, while committing DC-precursors differentiation toward the Langerhans cell (LC) pathway, TGFbeta likely inhibits CD1d transcription. Therefore, LC freshly recovered from epidermal sheet were evaluated by flow cytometry. In accordance with in vitro observation, they did not expressed measurable levels of CD1d molecules at the cell membrane. Thus, TGFbeta produced by keratinocytes contribute to selectively downregulate CD1d expression on intraepidermal-resident LC. PMID- 15654964 TI - Repeated topical challenge with chemical antigen elicits sustained dermatitis in NC/Nga mice in specific-pathogen-free condition. AB - NC/Nga mice are known to develop skin lesions resembling to atopic dermatitis (AD) in conventional but not in specific-pathogen-free (SPF) condition. An epicutaneous application of 2,4-dinitrofluorobenzene (DNFB) increased skin thickness in C3H as well as NC/Nga mice in SPF environment, and the response was enlarged by repeating the challenge at weekly intervals. Although the skin reaction in C3H mice was ameliorated when the challenge was discontinued after the fifth application, the reaction in NC/Nga mice was sustained at least for 3 wk. Analyses of cytokine production by CD4+ cells from the draining lymph node proximal to the lesions revealed that, unlike C3H mice, NC/Nga mice fail to induce T helper 2 (Th2) cytokine interleukin-4 (IL-4), whereas the level of Th1 cytokine interferon-gamma in NC/Nga mice is equivalent to that of C3H mice. In addition, NC/Nga mice highly expressed IL-12, a cytokine-preventing formation of Th2 response, whereas C3H mice did not. Administration of anti-IL-12 antibody reduced duration of dermatitis in DNFB-treated NC/Nga mice. Taken together, our data suggest that IL-12 plays a role in the persistent skin reaction in NC/Nga mice. The action of IL-12 might be mediated by the decrease in IL-4 production. PMID- 15654965 TI - Epitope-specific antibody response to Mel-CAM induced by mimotope immunization. AB - Peptide mimotopes of tumor antigen epitopes have been proposed as components of tumor vaccines. In this study, we determined the immunogenicity of melcam mim1 and melcam mim2, peptide mimics of an epitope of the melanoma cell-adhesion molecule (Mel-CAM). BALB/c mice were vaccinated either with mimotopes or mimotopes coupled to tetanus toxoid (TT). The antibody responses of mice to melcam mim1, melcam mim2, and recombinant Mel-CAM were analyzed by an ELISA and immunoblot analyses. TT-coupled mimotopes led to high titers of IgG mainly of the IgG2a subclass to melcam mim1 and melcam mim2. Immunization with each of the mimotope formulations induced antibodies that cross-reacted with recombinant Mel CAM. Uncoupled mimotopes induced lymphocyte proliferation and cytokine production in spleen cell cultures indicating that both peptide mimotopes also contained T cell epitopes. TT-coupled mimotopes induced T helper (Th)1 (interleukin (IL)-2, interferon-gamma) and Th2 (IL-4, IL-5) cytokines, whereas uncoupled mimotopes induced a Th1-biased T cell response. Our results suggest that mimotopes potentially represent a novel vaccine approach to induce a tumor antigen-specific humoral and cellular response. PMID- 15654967 TI - Mycobacterium vaccae reduces scratching behavior but not the rash in NC mice with eczema: a randomized, blinded, placebo-controlled trial. AB - In a randomized, double-blinded, placebo-controlled trial, we previously showed that intra-dermal administration of a killed Mycobacterium vaccae suspension to school-aged children with atopic dermatitis ameliorates their disease. We wished to test the hypothesis that M. vaccae may also prevent the development of eczema. As it was not possible to do this in children, we studied the NC/Nga eczema mouse model. Thirty NC/Nga mice were randomized into a blinded, placebo-controlled trial where they received either 0.1 or 0.01 mg of M. vaccae (SRP299) or placebo given subcutaneously at 1 and 8 wk of age. Clinical eczema scores, as well as scratching frequency using a digital videotape system were assessed during the 26 wk study. Digital scratch scores correlated with clinical severity (p=0.001). Although there were no significant differences in age of onset or severity of the rash between the three study arms, mice injected with 0.1 mg but not 0.01 mg of SPR299 had significantly lower peak scratch frequencies than controls (Hazard ratio 0.2; 95% confidence interval 0.1-0.7; p=0.01). We conclude that in this NC/Nga mouse model, SRP299 did have a beneficial effect in reducing pruritus, a major clinical symptom of eczema, although it does not prevent the rash from developing. PMID- 15654966 TI - Expression of FcRn, the MHC class I-related receptor for IgG, in human keratinocytes. AB - The neonatal Fc receptor (FcRn) for IgG has been shown to be responsible for IgG transport and to be involved in IgG catabolism. In this study, we show expression of FcRn in normal human epidermal keratinocytes. By RT-PCR, we demonstrate the FcRn alpha-chain mRNA obtained from cultured keratinocytes creating a 457 bp product as confirmed by sequence analysis. Northern blot analysis shows a 1.5 kb transcript. Real-time PCR reveals consistent expression of FcRn alpha-chain mRNA in human keratinocytes from different donors. Anti-FcRn alpha2-extracellular domain and anti-FcRn cytoplasmic tail antibody (Ab) directed against defined antigenic targets were generated and used for immunoblotting and immunoprecipitation revealing protein expression of the 46 kDa FcRn alpha-chain. By immunofluorescence microscopy, we find granular-vesicular staining for FcRn alpha-chain in keratinocytes. Fluorescence-activated cell sorting analysis gives predominantly an intracellular distribution of FcRn in keratinocytes. Biochemically, we demonstrate Fc-dependent binding of human IgG at acidic pH. In normal human epidermis, we find a cytoplasmic vesicular staining of predominantly basal and suprabasal keratinocytes. In summary, we demonstrate expression of a functional FcRn in normal human epidermal keratinocytes. These findings further emphasize the role of keratinocytes as immunomodulating cells in inflammatory and immunologic processes of the skin. PMID- 15654968 TI - Peripheral CD8+ T cell tolerance against melanocytic self-antigens in the skin is regulated in two steps by CD4+ T cells and local inflammation: implications for the pathophysiology of vitiligo. AB - Experimental evidence has suggested a role for CD8+ cytotoxic T lymphocytes (CTL) in the pathophysiology of vitiligo, a pigmentation disorder with focal loss of melanocytes in the skin. The discovery of tyrosinase-related protein 2 (TRP2) as a model melanocytic self-antigen recognized by CD8+ CTL in C57BL/6 mice allowed us to analyze the requirements for CD8+ T cell-mediated autoimmune destruction of melanocytes in an experimental model. Using two different genetic methods for the induction of cellular immunity in vivo, gene gun bombardment of the skin and injection of recombinant adenovirus, we show that peripheral tolerance of CD8+ T cells recognizing a single TRP2-derived H2-Kb-binding peptide is regulated in two steps. In the induction phase, stimulation and expansion of TRP2-specific CD8+ T cells in vivo depend on CD4+ T cell help. In the effector phase, autoimmune destruction of melanocytes in the skin depends on local inflammation. Our results suggest that accidental stimulation of CD8+ CTL recognizing major histocompatibility complex class I-binding peptides derived from melanocytic proteins in the context of an inflammatory skin disease may play an important role in the pathophysiology of vitiligo. PMID- 15654969 TI - A novel T cell receptor transgenic animal model of seborrheic dermatitis-like skin disease. AB - We have characterized a novel animal model of the common inflammatory skin disease seborrheic dermatitis (SD) that involves the expression of the self specific 2C transgenic T cell receptor on the DBA/2 genetic background. Opportunistic fungal pathogens are present in the primary histological lesions and severe disease can be mitigated by the administration of fluconazole, demonstrating a role for infection in disease pathogenesis. Spontaneous disease convalescence occurs at 70-90 d of age and is preceded by an expansion of CD4+ T cells that partially restores the T cell lymphopenia that occurs in these animals. The adoptive transfer of syngeneic CD4+ T cells into pre-diseased DBA/2 2C mice completely abrogates the development of cutaneous disease. The pattern of disease inheritance in DBA/2 backcrosses suggests that one, or a closely linked group of genes, may control disease penetrance. Bone marrow reconstitution experiments demonstrated that the DBA/2 susceptibility factor(s) governing disease penetrance is likely non-hematopoietic since bone marrow from disease resistant 2C mice can adoptively transfer the full disease phenotype to non transgenic DBA/2 animals. This model implicates fungal organisms and CD4+ T cell lymphopenia in the development of a SD-like condition and, as such, may mimic the development of SD in acquired immunodeficiency syndrome. PMID- 15654970 TI - DermaVir: a novel topical vaccine for HIV/AIDS. AB - Human immunodeficiency virus (HIV) vaccines have the potential to improve antiretroviral drug treatment by inducing cytotoxic killing of HIV-infected cells. Prophylactic vaccines utilize new antigens to initiate immunity; however, in HIV-infected individuals the load of viral antigen is not the limiting factor for the restoration of immune responses. Here we describe a novel immunization strategy with DermaVir that improves viral antigen presentation using dendritic cells (DC). DermaVir contains a distinctive plasmid DNA expressing all HIV proteins except integrase to induce immune responses with broad specificity. The DNA is formulated to a mannosilated particle to target antigen-presenting cells and to protect the DNA from intracellular degradation. After topical application, DermaVir-transduced cells migrate from the skin to the draining lymph node and interdigitate as DermaVir-expressing, antigen-presenting DC. We compared the immunogenicity of topical and ex vivo DC-based DermaVir vaccinations in naive rhesus macaques. Both vaccinations induced simian immunodeficiency virus-specific CD4 helper and CD8 memory T cells detected by an in vivo skin test and an in vitro intracellular cytokine-based assay. Topical DermaVir vaccination represents an improvement upon existing ex vivo DC-based immunization technologies and may provide a new therapeutic option for HIV-infected patients. PMID- 15654971 TI - Differentiated keratinocyte-releasable stratifin (14-3-3 sigma) stimulates MMP-1 expression in dermal fibroblasts. AB - Through the use of a keratinocyte/fibroblast co-culture system, we have recently identified a potent keratinocyte-derived anti-fibrogenic factor (KDAF) for dermal fibroblasts. A sequential chromatography of the active fractions of keratinocyte conditioned medium (KCM) and peptide mapping of the candidate proteins identified KDAF as being the keratinocyte-releasable 14-3-3 sigma (14-3-3sigma) protein, which is also known as stratifin. In this study, we hypothesize that differentiated, but not proliferating, keratinocytes are the primary source of releasable 14-3-3sigma in conditioned medium. To address this hypothesis, in a longitudinal study, keratinocyte differentiation was induced by growing these cells in a medium consisting of 50% keratinocyte serum-free medium (KSFM) and 50% Dulbecco's modified eagle's medium without any additives for up to 20 d. When KCM was collected every other day and added to fibroblasts, the level of matrix metalloproteinase (MMP)-1 mRNA expression was markedly increased in fibroblasts receiving KCM and this increase was even greater in cells receiving conditioned media collected at later time points relative to that of controls. The results of a western blot analysis further showed a marked increase in the expression of 14 3-3sigma protein in keratinocytes grown in test medium from day 4 to day 10. This finding was consistent with the levels of 14-3-3sigma mRNA expression in differentiated keratinocytes. In contrast to a very high level of 14-3-3sigma mRNA expression seen in keratinocytes, fibroblasts that are highly responsive to14-3-3sigma were unable to express this factor. Interestingly, the level of 14 3-3sigma mRNA expression was markedly higher in keratinocytes co-cultured with fibroblasts relative to that of mono-cultured keratinocytes. In conclusion, this study provides evidence that keratinocytes express a high level of 14-3-3sigma at the levels of mRNA and protein. But the releasable form of 14-3-3sigma protein was only found in conditioned medium derived from differentiated keratinocytes. Further, our recently purified recombinant 14-3-3sigma protein mimics the collagenase stimulatory effect of KCM in dermal fibroblasts. PMID- 15654973 TI - Phospholipase cgamma1 is required for activation of store-operated channels in human keratinocytes. AB - Store-operated calcium entry depicts the movement of extracellular Ca2+ into cells through plasma membrane Ca2+ channels activated by depletion of intracellular Ca2+ stores. The members of the canonical subfamily of transient receptor potential channels (TRPC) have been implicated as the molecular bases for store-operated channels (SOC). Here we investigate the role of phospholipase C (PLC) in regulation of native SOC and the expression of endogenous TRPC in human epidermal keratinocytes. Calcium entry in response to store depletion with thapsigargin was reversibly blocked by 2-aminoethoxydiphenyl borane, an effective SOC inhibitor, and suppressed by the diacylglycerol analoge, 1-oleoyl-2-acetyl-sn glycerol. Inhibition of PLC with U73122 or transfection of a PLCgamma1 antisense cDNA construct completely blocked SOC activity, indicating a requirement for PLC, especially PLCgamma1, in the activation of SOC. RT-PCR and immunoblotting analyses showed that TRPC1, TRPC3, TRPC4, TRPC5, and TRPC6 are expressed in keratinocytes. Knockdown of the level of endogenous TRPC1 or TRPC4 inhibited store-operated calcium entry, indicating they are part of the native SOC. Co immunoprecipitation studies demonstrated that TRPC1, but not TRPC4, interacts with PLCgamma1 and the inositol 1,4,5-trisphosphate receptor (IP3R). The association of TRPC1 with PLCgamma1 and IP3R decreased in keratinocytes with higher intracellular Ca2+, coinciding with a downregulation in SOC activity. Our results indicate that the activation of SOC in keratinocytes depends, at least partly, on the interaction of TRPC with PLCgamma1 and IP3R. PMID- 15654972 TI - Kinin B2 receptor-coupled signal transduction in human cultured keratinocytes. AB - Kinins are key pro-inflammatory peptides that exhibit mitogenic effects in tissue specific cellular systems. Since the life span of the keratinocyte is regulated by receptors that control proliferation and differentiation, and since both processes are affected during wound healing, we have examined the consequence of kinin B2 receptors (B2R) activation in cultured human keratinocytes. Stimulation of keratinocytes by Lys-bradykinin (LBK) induced a rapid and sustained phosphorylation of 42/44 mitogen-activated protein kinase (MAPK) that translocated to the nucleus, and decreased only after 120 min of stimulation. Kinin B1 and B2 receptor (B1R and B2R) antagonists showed that phosphorylation was mainly because of B2R activation. The GF109203X inhibitor almost completely abolished the effect of LBK, suggesting the involvement of protein kinase C in the signal cascade. MAPK phosphorylation was partially dependent on epidermal growth factor receptor transactivation as assessed by the selective inhibitor, AG1478. LBK stimulation did not result in cell proliferation, but produced a rapid c-Fos expression, nuclear translocation of nuclear factor-kappaB, and a moderated (pro)filaggrin synthesis, indicating that it may modulate cell differentiation. Our results support the view that kinins may affect the life span of human keratinocytes and highlight the importance that kinin peptides may have in the pathogenesis and/or progression of skin diseases. PMID- 15654974 TI - A proteolytic cascade of kallikreins in the stratum corneum. AB - Serine proteases belonging to the kallikrein group may play a central role in desquamation. We have identified human kallikreins 5, 7, and 14 (hK5, hK7, hK14) in catalytically active form in stratum corneum. All three enzymes are produced as inactive precursors. In this work, we prepared recombinant enzymes and enzyme precursors and characterized the catalytic properties of hK5 and hK14. With peptide substrates hK5 and hK14 both showed trypsin-like specificity and alkaline pH-optima. For the substrates tested, hK14 was superior to hK5 as regards maximum catalytic rate as well as catalytic efficiency. hK5, but not hK14, could activate pro-hK7 in a reaction which was optimal at pH 5-7. hK5 could activate its own precursor as well as pro-hK14. This was in contrast to hK14, which could activate pro-hK5 but not its own precursor. The activation of pro-hK5 either by auto activation or by hK14 occurred at maximum rate at neutral or weakly alkaline pH, whereas activation of pro-hK14 by hK5 was optimal at pH 6-7. We conclude that the enzymes studied may be part of a protease cascade in the stratum corneum, and that the observed pH effects may have physiological relevance. PMID- 15654976 TI - Fos and jun proteins are specifically expressed during differentiation of human keratinocytes. AB - Activator protein 1 (AP-1) proteins play key roles in the regulation of cell proliferation and differentiation. In this study we investigated the expression of Fos and Jun proteins in different models of terminal differentiation of human keratinocytes and in skin from psoriasis patients. All Jun and Fos proteins, with the exception of FosB, were efficiently expressed in keratinocytes in monolayer cultures. In contrast, in normal epidermis as well as in organotypic epidermal cultures, the expression pattern of AP-1 proteins was dependent on the differentiation stage. Fos proteins were readily detected in nuclei of keratinocytes of basal and suprabasal layers. JunB and JunD were expressed in all layers of normal epidermis. Interestingly, expression of c-Jun started suprabasally, then disappeared and became detectable again in distinct cells of the outermost granular layer directly at the transition zone to the stratum corneum. In psoriatic epidermis, c-Jun expression was prominent in both hyperproliferating basal and suprabasal keratinocytes, whereas c-Fos expression was unchanged. These data indicate that AP-1 proteins are expressed in a highly specific manner during terminal differentiation of keratinocytes and that the enhanced expression of c-Jun in basal and suprabasal keratinocytes might contribute to the pathogenesis of psoriasis. PMID- 15654975 TI - Characterization and differentiation-dependent regulation of secreted phospholipases A in human keratinocytes and in healthy and psoriatic human skin. AB - Secreted phospholipases A2 (sPLA2) expressed in the skin are thought to be involved in epidermal barrier homeostasis as well as in inflammation. We investigated the expression of the novel sPLA2 subtypes in human skin at mRNA and protein levels in the epidermis and primary keratinocytes from healthy human skin, and in skin sections from patients with psoriasis, where the integrity of the epidermis is drastically affected. Immunofluorescence studies using specific antibodies for the different sPLA2 enzymes show that sPLA2-IB, -IIF, and -X are predominantly expressed in suprabasal layers, whereas sPLA2-V and -IID are detected in the basal and spinous layers. sPLA2-IIA is weakly expressed, and sPLA2-IIE and XIIA are not detectable. Accordingly, in differentiated human primary keratinocyte cultures, the expression of sPLA2-IB, -IIF and -X was increased, whereas that of sPLA2-V and -IID was markedly decreased. In psoriatic skin, sPLA2-X was dramatically downregulated in the epidermis, whereas increased amounts of this enzyme together with sPLA2-IIA, -IID, and -IB appeared in the dermis. An enhanced release of these enzymes with the exception of sPLA2-IID was also observed after treatment of HaCaT keratinocytes with tumor necrosis factor alpha/interferon-gamma. Treatment of HaCaT cells with sPLA2-X and -IB resulted in an increase in prostaglandin E2 formation, suggesting a proinflammatory role of these enzymes during psoriasis. sPLA2-V completely disappeared. The differential locations of the sPLA2 enzymes propose distinct roles of individual enzymes in skin. PMID- 15654977 TI - Selective induction of apoptosis in melanoma cells by tyrosinase promoter controlled CD95 ligand overexpression. AB - Induction of apoptosis has been demonstrated previously by overexpression of CD95 ligand (CD95L) in cultured human melanoma cells. For in vivo approaches based on CD95L, however, targeted expression is a prerequisite and tyrosinase promoters have been considered for selection. Luciferase reporter gene assays performed for a representative panel of melanoma cell lines characterized by strong (SK-Mel 19), moderate (SK-Mel-13, MeWo), weak (A-375), and missing expression (M-5) of endogenous tyrosinase revealed high tyrosinase promoter activities in SK-Mel-19, SK-Mel-13, and MeWo, but only weak activities in A-375 and M-5 as well as in non melanoma cell lines. After transfection of a CMV promoter CD95L expression construct, melanoma cells were found highly sensitive, as compared with non melanoma cells. By applying a tyrosinase promoter CD95L construct, apoptosis was selectively induced in SK-Mel-19, SK-Mel-13, MeWo as well as in A-375, which was characterized by high CD95 surface expression and high sensitivity to agonistic CD95 activation. M5 and non-melanoma cell lines remained uninfluenced. Also, resistance to agonistic CD95 activation seen in MeWo characterized by weak CD95 surface expression was overcome by overexpression of CD95L. Our investigations provide evidence that tyrosinase promoter CD95L constructs may be of value for selective induction of apoptosis in therapeutic strategies for melanoma. PMID- 15654978 TI - Bcl-2 reduced and fas activated by the inhibition of stem cell factor/KIT signaling in murine melanocyte precursors. AB - Stem cell factor (SCF) and its receptor, KIT, are essential to the migration and differentiation of melanocytes during embryogenesis. We previously demonstrated that apoptosis is induced by blocking survival function of the SCF/KIT interaction in a mouse neural crest cell (NCC) primary culture. Using the NCCmelb4 cell line, we investigated the occurrence of apoptosis in the cultured cells when KIT receptors were blocked by the monoclonal anti-KIT antibody (ACK2). Apoptosis following treatment with ACK2 was detected by DNA fragmentation assay, in situ apoptosis detection, and electron microscopy. We noted a decrease in extracellular signal-related kinase (ERK) and ribosomal S6 kinase (RSK) protein expression following ACK2 incubation. Western blot analysis and real-time quantitative RT-PCR revealed an apparent time-dependent reduction in Bcl-2 protein levels with respect to ACK2 within the NCCmelb4 cells. In terms of Bax expression, a difference was not found. Fas and caspase8 proteins increased time dependently in proportion to ACK2 incubation. We noted apoptotic cell death upon addition of ACK2, with evidence of possible involvement of Bcl-2 and Fas in the induction of apoptosis. In contrast, no significant correlation between Fas ligand (Fas-L) expression and ACK2 was found. Fas activation appears to occur independent of Fas-L during ACK2-induced cell death. Therefore, we propose that Fas-L expression in NCCmelb4 cells does not play a major role in facilitating apoptosis. Furthermore, we hypothesize that these molecules combined with SCF/KIT play an important role in regulating the induction of vertebrate NCC apoptosis during embryogenesis. PMID- 15654979 TI - Non-invasive visualization of melanin and melanocytes by reflectance-mode confocal microscopy. AB - In vivo visualization of epidermal melanin was performed by reflectance-mode confocal microscopy (RCM). Firstly, we examined the distribution of epidermal melanin in pigmented animals and compared with that of the human skin. Melanocytes in the skin of pigmented animals were found to accumulate a large amount of melanin that can be easily visualized because of its brightness. Their RCM images correlated well with the Fontana-Masson-stained sections for melanin. In contrast, in the human skin, typical dendritic melanocytes were hardly observed even in pigmented lesions, although supranuclear melanin caps were easily visible. These results suggested that human melanocytes rapidly transfer the produced melanin to keratinocytes and do not accumulate it. Secondly, to elucidate the production of melanin by human melanocytes, we evaluated the changes of melanin after a single ultraviolet (UV) exposure. The melanin accumulating melanocytes were clearly visualized during the skin pigmentation process. The RCM images showed the brightness because of melanin gradually increased from day 4, then dendrite-elongated melanocytes appearing from day 8, and finally melanin caps formed from day 29. In conclusion, RCM successfully evidenced the difference in melanin distribution between the pigmented animals and humans, and the UV-induced pigmentation process in vivo as well. PMID- 15654980 TI - Single UVB overexposure stimulates melanocyte proliferation in murine skin, in contrast to fractionated or UVA-1 exposure. AB - Overexposure to short- and long-wave ultraviolet radiations (UVB, UVA) may contribute to melanoma development through combined genotoxic and mitogenic effects in melanocytes. This study compares the impact of UVA-1 versus UVB, and single versus fractionated exposures on melanocyte proliferation in hairless SKH 2 mice. A single erythemal dose was compared with an equal dose fractionated over 8 d, and dose-dependency was studied. Proliferation (Ki-67 positive-sign) in melanocytes (melanoma antigen recognized by T-cells-1 positive or micropthalmia transcription factor positive) was ascertained in double-labeled skin sections. Single erythemal UVB exposures caused a delayed, dose-dependent increase of melanocyte proliferation. The highest, 17-fold, increase (from 0.05% to 0.8% of melanocytes) occurred 4 d after UVB exposure, without any detectable effect on overall melanocyte numbers. Correspondingly, DNA repair-deficient xeroderma pigmentosum A (Xpa) mice proved exquisitely sensitive to melanocyte proliferation induction by UVB exposure. No discernable effects were measured from fractionated suberythemal UVB exposures, or from any UVA-1 exposure regimen. Hence, melanocyte proliferation appears to be most efficiently induced by a single UVB overexposure. Moreover, the ineffectiveness of UVA-1 radiation and the enhanced sensitivity of Xpa mice point at pyrimidine dimers as causative DNA lesions. Consequently, murine nevi and melanoma are expected to be most effectively induced by intermittent UVB overexposures. PMID- 15654981 TI - Eicosapentaenoic acid and docosahexaenoic acid reduce UVB- and TNF-alpha-induced IL-8 secretion in keratinocytes and UVB-induced IL-8 in fibroblasts. AB - Omega-3 polyunsaturated fatty acids (n-3 PUFA) inhibit ultraviolet B (UVB) induced inflammation and other inflammatory states, in vivo. We examined whether this may be mediated by modulation of interleukin (IL)-8, a chemokine pivotal to skin inflammation induced by UVB, in epidermal and dermal cells. We also explored the ability of n-3 PUFA to protect against tumor necrosis factor (TNF)-alpha induction of IL-8, and assessed relative potencies of the principal dietary n-3 PUFA, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA). Pre supplementation, both HaCaT keratinocyte and CCD922SK fibroblast cell lines showed dose-responses for UVB-induced IL-8 release (p<0.001), assessed 48 h post irradiation. Cells were supplemented with > or =90% purified EPA, DHA, oleic acid (OA) or vehicle control, for 4.5 d. EPA and DHA supplements were bioavailable to keratinocytes and fibroblasts. In keratinocytes, EPA and DHA were shown to reduce basal secretion of IL-8 by 66% and 63%, respectively (p<0.05), and UVB-induced levels by 66% and 65% at 48 h after 100 mJ per cm2, respectively, (p<0.01). A similar pattern occurred in fibroblasts, whereas OA had no influence on IL-8 release in either cell line. In addition, TNF-alpha-induced IL-8 secretion by keratinocytes was reduced by 54% and 42%, respectively, by EPA and DHA (p<0.001). Hence both n-3 PUFA inhibit production of UVB- and TNF-alpha-induced IL-8 in skin cells; this may be important in the photoprotective and other anti-inflammatory effects conferred by these agents. PMID- 15654984 TI - Does recessive EKV exist? PMID- 15654982 TI - Prevention of murine erythropoietic protoporphyria-associated skin photosensitivity and liver disease by dermal and hepatic ferrochelatase. AB - Erythropoietic protoporphyria (EPP) is caused by a defect in ferrochelatase, leading to the accumulation of protoporphyrin predominantly in erythrocytes and hepatocytes, and resulting in skin photosensitivity upon leaching of blood protoporphyrin into the skin. Some patients also develop severe liver damage. Because the respective contributions of hepatic and erythrocytic protoporphyrin to the pathophysiology of EPP remain unclear, we investigated this question using the murine model of EPP. Transplantation of bone marrow from EPP mice to normal recipients resulted in elevated erythrocyte and plasma protoporphyrin levels. However, quantification of serum liver enzymes and bilirubin together with histopathologic examination of liver sections of mice up to 16 months post transplantation showed no evidence of liver damage. Moreover, despite massive elevation of serum protoporphyrin, transplanted mice showed minimal evidence of skin photosensitivity. Photosensitivity could also be prevented locally by implanting skin grafts from normal mice onto the backs of EPP recipients. These data validate the hypothesis that the main source of toxic protoporphyrin originates from the erythrocytes. However, we unexpectedly observed that normal ferrochelatase activity in hepatic and dermal cells of wild-type mice is sufficient to prevent liver disease and significant skin photosensitivity. These findings may provide new strategies for the treatment of EPP. PMID- 15654986 TI - A novel recessive missense mutation in KRT14 reveals striking phenotypic heterogeneity in epidermolysis bullosa simplex. PMID- 15654983 TI - Anti-rejection drug treatment increases basal cell carcinoma burden in Ptch1+/- mice. AB - The development of extensive and severe non-melanoma skin cancer is an extremely common complication of organ transplantation and is assumed to be caused by long term treatment with anti-rejection drugs (ARD). Despite this florid clinical problem, ARD treatments have been reported to affect experimental murine skin carcinogenesis only weakly. We report here that treatment of cesium-137 irradiated Ptch1+/- mice with immunosuppressive doses of cyclosporine A plus prednisolone for 4-1/2 mo increased basal cell carcinoma burden by 2.5-fold. Thus, these mice provide a good model for study of the effects of long-term administration of ARD on at least one type of non-melanoma skin cancer. PMID- 15654987 TI - 2,3,7,8-tetrachlorodibenzo-p-dioxin impairs differentiation of normal human epidermal keratinocytes in a skin equivalent model. PMID- 15654988 TI - Absence of SV40 and other polyomavirus (JCV, BKV) DNA in primary cutaneous B cell lymphomas. PMID- 15654989 TI - Presence of Epstein-Barr virus in Langerhans cells of CTCL lesions. PMID- 15654990 TI - Association of TNF -238 and -308 promoter polymorphisms with psoriasis vulgaris and psoriatic arthritis but not with pustulosis palmoplantaris. PMID- 15654991 TI - Upregulation of CD44 and hyaluronate synthases by topical retinoids in mouse skin. PMID- 15654992 TI - Alopecia areata induced in C3H/HeJ mice by interferon-gamma: evidence for loss of immune privilege. PMID- 15654993 TI - Influence of sperm cell dose and post-insemination backflow on reproductive performance of intrauterine inseminated sows. AB - This study was carried out to evaluate the effects of the sperm cell dose and semen backflow on the pregnancy rate and number of embryos of sows inseminated once at 0-24 h before ovulation, using an intrauterine technique. The results were analysed from a total of 211 sows assigned to three groups inseminated with doses of 0.25 x 10(9) (T1), 0.5 x 10(9) (T2) and 1.0 x 10(9) (T3) spermatozoa. Semen backflow was observed in 95% of the females (143/151) evaluated for this purpose. The percentage of semen backflow is close to two-third of the volume and the percentage of sperm is around 15% of the infused sperm dose. Intrauterine insemination can be successfully performed provided that at least 0.5 billion of sperm cell dose is infused at an interval of 0-24 h before ovulation. PMID- 15654994 TI - Synchronization of ovulation and fixed time intrauterine insemination in ewes. AB - A novel method for oestrus-ovulation synchronization in sheep followed by fixed time insemination is presented herewith. Mature dry ewes (n = 28) of Karagouniko breed being at an unknown stage of the oestrous cycle, were used during the middle of breeding season. The treatment protocol consisted of an initial administration of a GnRH analogue followed 5 days later by a prostaglandin F2alpha injection. Thirty-six hours later a second GnRH injection was administered to synchronize ovulation, and laparoscopic intrauterine insemination was performed 12-14 h later. Three days after insemination, fertile rams were introduced into the flock twice daily and oestrus-mating detection was carried out. For progesterone (P(4)) determination, blood samples were collected on alternate days, starting 2 days before the first GnRH injection and continuing for 17 days after insemination. An additional sample was taken on the day of insemination. Pregnancy diagnosis was carried out by trans-abdominal ultrasonography. Fourteen ewes (50%) conceived at insemination and maintained pregnancy; from the remainder 14 ewes 10 became pregnant at natural service, while four, although they mated at least two to three times, failed to conceive. In response to the first GnRH, P(4) concentration increased at higher levels in ewes that conceived at AI compared with those that failed to conceive (47.54 and 22.44%, respectively; p < 0.05). Significant differences (p < 0.05) in mean P(4) concentration between pregnant and non-pregnant animals were detected 1 day before AI (0.17 +/- 0.06 and 0.26 +/- 0.14 ng/ml, respectively) on the day of AI (0.15 +/- 0.04 and 0.24 +/- 0.08 ng/ml, respectively) as well as 9 and 11 days thereafter (0.48 +/- 0.12 and 0.38 +/- 0.12 ng/ml; 0.68 +/- 0.14 and 0.50 +/- 0.18 ng/ml, respectively). These results indicate that using the proposed protocol, an acceptable conception rate can be achieved which could be further improved by modifying the time intervals between interventions. PMID- 15654996 TI - Addition of recombinant human growth hormone to in vitro maturation medium of bovine oocytes. AB - The aim of this study was to increase the bovine embryonic development rate, adding recombinant human growth hormone (rhGH) to maturation medium of bovine oocytes. Oocytes were matured for 24 h in TCM 199 Earle's salts and five treatments were developed: T1, 0.01 IU/ml of recombinant human follicle stimulating hormone (rhFSH); T2, 0.01 IU/ml of rhFSH + 100 ng/ml of rhGH; T3, 0.01 IU/ml of rhFSH + 1000 ng/ml of rhGH; T4, 100 ng/ml of rhGH; and T5, 1000 ng/ml of rhGH at 39 degrees C and 5% of CO(2) in air and saturated humidity. In vitro fertilization from cumulus-oocyte complexes was conducted in TALP-Fert medium (18-22 h) and spermatozoa were selected by Percoll gradient. Zygotes were incubated in SOFaaci medium in 5% of CO(2) in air, 5% of O(2) at 39 degrees C and saturated humidity for 11 days. There was no statistical difference in cleavage rate and embryo production on day 7 and day 9 among treatments. However, the hatching rate increased significantly in the T4 and T5 treatments (11.0 and 12.8%, respectively), compared with the T1 treatment (4.6%) (p < 0.05). Therefore, the rhGH addition to the oocyte maturation medium showed beneficial effects on the hatching rate of in vitro-produced bovine embryos. PMID- 15654995 TI - Antigens of goat spermatozoa that recognize homologous zona pellucida. AB - Goat sperm surface proteins obtained from purified plasma membrane (PPM) vesicles (purity of membrane checked by marker enzymes and transmission electron microscopy) were size fractionated on an fast protein liquid chromatography (FPLC) gel filtration column. All the seven surface proteins (129, 100, 46, 28, 27, 18 and 10 kDa) obtained were further fractionated and purified on high efficiency gel filtration (GFC-HPLC) as well as ion exchange (DEAE-HPLC) columns. Antibodies were generated against the PPM and the protein fractions. Such resolved and purified surface antigens were tested by Dot Blot Immunoassay and homologous in vitro sperm-zona binding assays. It was revealed that the binding of goat spermatozoa to homologous zona pellucida was inhibited by antisera raised against the five lower molecular weight surface antigens. Further, the components of FPLC-AIII (46 kDa; A represents antigenic protein) and IV (28 kDa) were most promising as the antibodies against these fractions inhibited sperm binding to zona pellucida even at a dilution of 1 : 1000 as tested by the sperm-zona binding assays. PMID- 15654997 TI - Developmental changes of bull (Bos taurus) genitalia as evaluated by caliper and ultrasonography. AB - The objective of the present study is to follow the growth of the testes and the pelvic genitalia in cattle bulls, using ultrasonographic examination and measurements of external genitalia by caliper. Six Friesian bulls were examined periodically from 3 to 24 months of age with 3-month interval. Scrotum circumference (SC) and length were measured by tape. Testicular length, width and depth were measured by caliper. Scrotal contents including testes and epididymis, and the pelvic genitalia including bulbourethral gland (BUG), prostate and seminal glands (SG) were examined using transrectal ultrasonography. Scrotal circumference, length and the three dimensions of the testes showed significant increase with the advancement of age. Significant correlation was present between SC and testes breadth. At the same time, testis breadth as estimated by sonar showed positive correlation with that estimated by caliper. A positive correlation was also found between testicular breadth with each of the BUG and SG. Finally, it was concluded that ultrasonography gave appreciable benefits in studying the developmental changes of the testes and accessory glands of growing bulls. The obtained data could provide a useful tool for predicting bull puberty and fertility. PMID- 15654998 TI - The sow endosalpinx at different stages of the oestrous cycle and at anoestrus: studies on morphological changes and infiltration by cells of the immune system. AB - The aim of this study was to investigate the morphological changes of the sow endosalpinx and the distribution of leukocytes throughout the oestrous cycle and at anoestrus. Nineteen crossbred sows (Swedish Landrace x Swedish Yorkshire) at late dioestrus (three), prooestrus (three), oestrus (three), early dioestrus (three), dioestrus (three) and anoestrus (four) were used. Oviductal samples from three different parts (isthmus, ampulla and infundibulum), taken immediately after slaughter, were fixed, embedded in plastic resin and stained with toluidine blue or stored in a freezer at -70 degrees C until analysed by immunohistochemistry (prooestrus and anoestrus) with an avidin-biotin peroxidase method. Quantitative and qualitative examinations of oviductal epithelium and subepithelial connective tissue were performed by light microscopy. During all stages, a lower degree of morphological changes (pseudostratification, mitosis and secretory granules) was found in the isthmus compared with ampulla and infundibulum. In ampulla and infundibulum, pseudostratification, mitotic activity and secretory granules of the epithelium were high at prooestrus/oestrus. Cytoplasmic protrusions of epithelial cells with some extruded nuclei were prominent in ampulla and infundibulum at all stages except for oestrus and early dioestrus. Lymphocytes as well as CD2- and CD3-positive cells were the predominant immune cells in the epithelial layer. The numbers of lymphocytes and CD3-positive cells did not differ among segments and stages. Numbers of CD2 positive cells did not differ between prooestrus and anoestrus while the numbers were significantly higher in the infundibulum than in ampulla and isthmus. Neutrophils were only occasionally found and mainly in the infundibulum. In the subepithelial connective tissue layer, the two most commonly observed immune cell types were lymphocytes and plasma cells. The numbers of lymphocytes as well as CD2- and CD3-positive cells was lower in isthmus than in the other segments (p < or = 0.001). Higher numbers of plasma cells (p < or = 0.001) were found in infundibulum than in ampulla and isthmus. The numbers of lymphocytes and plasma cells were not significantly different between stages of the oestrous cycle. However, the number of neutrophils differed and were highest at prooestrus in ampulla and infundibulum. The numbers of CD2-, CD3- and CD79-positive cells did not differ between prooestrus and anoestrus whereas for CD14- and SWC3-positive cells, the numbers were higher at prooestrus (p < or = 0.05) than at anoestrus. In the oviduct, the morphology differed in ampulla and infundibulum with oestrous cycle stages, which indicates an effect by ovarian steroid hormones. The immune cell infiltration was less influenced by cyclic changes. However, the immune cell infiltration (in the connective tissue) in the upper part, especially infundibulum, differed significantly from the one in the lower part, isthmus, indicating different immune functions within various parts of the oviduct. PMID- 15654999 TI - Cyclic related and pathological changes in the lectin-binding sites on the swine oviduct. AB - This study was carried out to compare the lectin-binding pattern in the normal and pathological oviduct of sows during the ovarian cycle. Lectin-binding patterns showed differences between segments, phases of ovarian cycle and presence of morphologic lesions. In the infundibulum, it was observed that the cysts, in the follicular phase, reduced Ricinus communis-I (RCA-I) and Ulex europaeus-I (UEA-I) binding. Furthermore, in the pathological oviducts of the luteal-phase group, there is a reduction of Concanavalia ensiformis (Con-A) reactivity in this segment of the tube having wall cysts, adenomyosis and diverticulus. The Arachis hypogaea (PNA) binding in the infundibulum, during the luteal phase, decreased in the tube having adenomyosis. In animals with wall cysts, the Con-A, Triticum vulgaris (WGA) and RCA-I reactivity was minor in the glycocalyx of the isthmus epithelium during follicular phase. Con-A and Dolichos biflorus (DBA) binding pattern was minor in the luteal-phase isthmus of the tube having wall cysts, adenomyosis and diverticulus. In the ampulla, the wall cysts impaired the Con-A reaction only in the basal region of the epithelium, in the follicular phase. Binding with Con-A was decreased in the ampulla of animals in the luteal phase in the tube lesions with cysts and diverticulus. In addition, the diverticulus observed in the ampulla, during the luteal phase, reduced the PNA tubaric binding. The results of this study showed that the morphologic alterations modify the sugar pattern in the oviduct of sows. These modifications in glycoconjugates may be one of the reasons for the failure of fertility in sows. PMID- 15655000 TI - The activity of matrix metalloproteinase-2 and -9 in serum of pregnant and non pregnant bitches. AB - The aim of this study was to investigate, whether the activity of matrix metalloproteinase (MMP)-2 and -9 in the serum of pregnant and non-pregnant bitches differs significantly. For this purpose, 81 blood samples were taken from pregnant bitches at days 5-13, 15-21, 24-31, 34-40 and 41-50 after mating, and 51 samples from non-pregnant animals at corresponding times. Relative enzyme activity, calculated as the percentage of serum enzyme activity on enzyme activity in a control sample, was determined with a commercially available assay after activation of serum MMPs with p-aminophenylmercuric acetate (APMA). In addition, serum oestradiol (E(2)) and progesterone (P(4)) concentrations were measured with an enzymeimmunoassay (EIA). In the pregnant bitches, at days 5-13 and 15-21 after mating, the mean activity of both MMPs was significantly higher than in non-pregnant animals (28.5% vs 24.5% and 27.7% vs 22.6%; p < 0.01). Moreover, in the pregnant bitches, significant correlations were detected between the serum enzyme activity and the serum concentrations of E(2) (-0.900; p < 0.05) and P(4) (+0.667; p < 0.05). PMID- 15655001 TI - Relationship between oxygen consumption and sex of bovine in vitro fertilized embryos. AB - The present study was conducted to examine the relationship between the oxygen consumption rate and sex ratio of bovine in vitro fertilized embryos on each day of blastocyst formation. The quality of blastocysts collected on day 7, 8, and 9 after in vitro fertilization (IVF) were categorized as ranks A and B (excellent and good, respectively) based on microscopic observation of the morphology. The oxygen consumption rate and sex of individual blastocysts were evaluated using two novel techniques: scanning electrochemical microscopy (SECM) and loop mediated isothermal amplification (LAMP), respectively. The oxygen consumption rates of embryos of rank A were significantly higher (p < 0.05) than those of rank B, irrespective of the day of blastocyst appearance after IVF. Neither did the proportion of male embryos of ranks A and B differ significantly from each other at any of the days examined, nor from the average proportion (53%). The oxygen consumption rate of embryos of rank B collected on day 8 was significantly higher (p < 0.05) in female embryos than in male embryos collected on the same day. However, there were no apparent differences of oxygen consumption rates at each day of blastocyst appearance between male and female embryos of rank A. These results indicate that the oxygen consumption rate of individual embryos reflects their quality but does not correlate with the sex ratio of embryos of excellent quality. PMID- 15655002 TI - High porcine parvovirus antibodies in sow herds: prevalence and associated factors. AB - Porcine parvovirus (PPV) is widespread among swine. Our objective was to determine the prevalence of loosely housed sow herds in Finland with at least one animal with high (infection level) PPV antibodies and to gather basic knowledge about vaccination practices. In addition, selected factors associated with high antibody levels found in sows were examined. Altogether, 247 animals were sampled in 21 randomly chosen loosely housed sow herds. Samples were analysed with the haemagglutination inhibition (HI) test. PPV proved to be common; in 17 farms (81%) at least one animal had a high titre (>1 : 512), and 44% of all animals sampled had a high titre. The vaccination programmes had many shortcomings. In the generalised estimation equations (GEE) population-averaged model developed, the factors found to have a significant (p < or = 0.05) effect on HI titres were herd size, parity of two or greater and storage of the vaccine vial after use. Non-returning rate, re-breeding interval and litter size did not differ between herds with no high HI titres (n = 4) and those with at least one high HI titre (n = 17). PMID- 15655003 TI - Timing of ovulation and fertility of heifers after synchronization of oestrus with GnRH and prostaglandin F(2alpha). AB - Synchronization of oestrus and/or ovulation can reduce workload in heifer reproductive management. The objective of this study was to compare two protocols to synchronize oestrus and/or ovulation using GnRH and prostaglandin F(2alpha) (PGF(2alpha)) in dairy heifers concerning their effect on follicular dynamics and reproductive performance. Four trials were carried out. In trial 1, 282 heifers were treated with GnRH and PGF(2alpha) 7 days apart (GP protocol). One group was inseminated on detection of oestrus (IDO 1), and the other group received two timed artificial inseminations (AI) 48 and 72 h after PGF(2alpha) administration (TAI 1). In trial 2, 98 heifers were synchronized with the same GP protocol. Heifers in IDO 2 were treated as in IDO 1, heifers in TAI 2 received two TAI 48 and 78 h after PGF(2alpha) administration. In trial 3, heifers in IDO 3 (n = 71) were again treated as in IDO 1. Heifers in TAI 3 (n = 166) received a second dose of GnRH 48 h after PGF(2alpha) (GPG protocol) and TAI together with this treatment and 24 h later. Trial 4 compared the timing of ovulation after the GP and the GPG protocol, using a subgroup of the heifers from trials 1 to 3. The ovaries of the heifers were scanned via ultrasound at 48, 56, 72, 80, 96 and 104 h after PGF(2alpha) administration. Timing of ovulation and size of the ovulatory follicles were compared between the two groups. In trials 1 to 3, conception rates to first service were between 49 and 66%. They did not differ significantly between IDO and TAI groups within or between trials. Pregnancy rates per synchronization were numerically higher in the TAI groups, but the difference was not significant. Conception rates to breeding on spontaneous oestrus in heifers returning to oestrus were higher than that after synchronized oestrus. In trial 4, more heifers ovulated before the end of the observation period in GPG than in GP (96.5% vs 74.7%; p < 0.001). Overall, ovulatory follicles were smaller in GPG (13.1 +/- 1.9 mm vs 14.3 +/- 1.9 mm; p < 0.001). PMID- 15655004 TI - Associations between energy metabolism, LH pulsatility and first ovulation in early lactating cows. AB - This study was designed to elucidate associations between energy metabolism and LH pulsatility characteristics in early lactation, and days to first ovulation, in order to explain the relationship between energy balance and fertility observed in epidemiological studies. To this end, 10 multiparous HF cows were monitored during lactation, after the application of two different feeding strategies during the dry period. Days to first ovulation was assessed using blood progesterone measurements and LH pulsatility was measured in 8-h windows in the third week postpartum. The association between depth of negative energy balance and days to ovulation was confirmed. However, this study does not support the idea that LH pulsatility characteristics in early lactation are predictive for the interval between parturition and first ovulation. PMID- 15655005 TI - First established pregnancies in Mediterranean Italian buffaloes (Bubalus bubalis) following deposition of sexed spermatozoa near the utero tubal junction. AB - At the time of AI following Ovsynch protocol, a total of 51 buffaloes were randomly divided in a first group (n = 30) subjected to conventional AI into the uterine body with 20 million non-sex sorted frozen-thawed spermatozoa, while a second group (n = 21) was inseminated near the utero-tubal junction (UTJ) ipsilateral to the ovary carrying the preovulatory follicle with 2.5 million live (4 million total) sex-sorted frozen-thawed spermatozoa. The semen used for flowcytometric sorting was collected and processed on a farm in Italy, and then shipped to a laboratory in Germany. Eleven buffaloes were inseminated with X chromosome bearing spermatozoa and 10 with Y-chromosome bearing spermatozoa. Conception rates after conventional and UTJ inseminations were 43.3% (n = 13) and 42.8% (n = 9) respectively (p = 0.97). Eight of the nine foetuses obtained after insemination with sexed spermatozoa corresponded to the sex as predicted by the cell sorting procedure (five male and four female foetuses by ultrasound vs six male and three female foetuses by cell sorting). In conclusion, for the first time buffalo semen has been successfully subjected to procedures for flowcytometric sperm sorting and freezing. Low doses of sexed spermatozoa have been deposited near the UTJ giving conception rates similar to those of conventional AI with full dose. PMID- 15655006 TI - Evaluation of the lipid content in bovine oocytes and embryos with nile red: a practical approach. AB - In this study, the fluorescent lipid dye Nile Red, was used to demonstrate that the lipid content of immature bovine oocytes is correlated with the morphological appearance of the ooplasm. Oocytes with a uniform dark cytoplasm contained significantly more intracellular lipids in lipid droplets compared with oocytes with a granulated or pale cytoplasm (p < 0.05). Furthermore, this lipid-analysing technique was applied for the first time on single bovine in vitro embryos, showing a significant increase of the lipid content in lipid droplets after culture in the presence of serum (p < 0.05). PMID- 15655007 TI - Editorial: Should artemisinin-based combination treatment be used in the home based management of malaria? PMID- 15655008 TI - Developmental impairments following severe falciparum malaria in children. AB - OBJECTIVE: Neurological deficits are reported in children after cerebral malaria (CM) but little is known about the prevalence and characteristics of persisting neurocognitive consequences. The prevalence of developmental impairments following other complications of falciparum malaria, such as multiple, prolonged or focal seizures, is not known. Thus, our objective was to investigate the long term developmental outcome of CM and malaria with complicated seizures (M/S). METHODS: We followed up a cohort of children previously exposed to CM or M/S and children unexposed to either condition. All children between 6 and 9 years of age, exposed to CM, and an equal number of children exposed to M/S were identified from databases of hospital admissions from 1991 to 1998. The unexposed group was randomly selected from a census database. The children's performance was measured using assessments of cognition, motor, speech and language, hearing and vision. A parental questionnaire was used to identify children with epilepsy. RESULTS: CM group scores were significantly lower than unexposed group scores on the assessments of higher level language (adjusted mean difference -1.63, 95% CI: -2.99 to -0.27), vocabulary (-0.02, 95% CI: -0.04 to -0.01), pragmatics (OR 2.81, 95% CI: 1.04-7.6) and non-verbal functioning (-0.33, 95% CI: -0.61 to -0.06). The areas of significantly reduced functioning for the M/S group were concentrated on phonology (OR 2.74, 95% CI: 1.26-5.95), pragmatics (OR 3.23, 95% CI: 1.2-8.71) and behaviour (OR 1.8, 95% CI: 1.0-3.23). The performance of the active epilepsy group was significantly poorer than that of the group without epilepsy on the tests of comprehension, syntax, pragmatics, word finding, memory, attention, behaviour and motor skills. CONCLUSIONS: CM and M/S are associated with developmental impairments. If these impairments persist, this may have implications for least 250,000 children in Sub-Saharan Africa each year. Active epilepsy significantly increases the risk of cognitive and behavioural problems in children with a history of severe malaria. PMID- 15655009 TI - Spatial effects of the social marketing of insecticide-treated nets on malaria morbidity. AB - Randomized controlled trials have shown that insecticide-treated nets (ITNs) have an impact on both malaria morbidity and mortality. Uniformly high coverage of ITNs characterized these trials and this resulted in some protection of nearby non-users of ITNs. We have now assessed the coverage, distribution pattern and resultant spatial effects in one village in Tanzania where ITNs were distributed in a social marketing programme. The prevalence of parasitaemia, mild anaemia (Hb <11 g/dl) and moderate/severe anaemia (Hb <8 g/dl) in children under five was assessed cross-sectionally. Data on ownership of ITNs were collected and inhabitants' houses were mapped. One year after the start of the social marketing programme, 52% of the children were using a net which had been treated at least once. The ITNs were rather homogeneously distributed throughout the village at an average density of about 118 ITNs per thousand population. There was no evidence of a pattern in the distribution of parasitaemia and anaemia cases, but children living in areas of moderately high ITN coverage were about half as likely to have moderate/severe anaemia (OR 0.5, 95% CI: 0.2, 0.9) and had lower prevalence of splenomegaly, irrespective of their net use. No protective effects of coverage were found for prevalence of mild anaemia nor for parasitaemia. The use of untreated nets had neither coverage nor short distance effects. More efforts should be made to ensure high coverage in ITNs programmes to achieve maximum benefit. PMID- 15655010 TI - Experience of targeting subsidies on insecticide-treated nets: what do we know and what are the knowledge gaps? AB - Widespread coverage of vulnerable populations with insecticide-treated nets (ITNs) constitutes an important component of the Roll Back Malaria (RBM) strategy to control malaria. The Abuja Targets call for 60% coverage of children under 5 years of age and pregnant women by 2005; but current coverage in Africa is unacceptably low. The RBM 'Strategic Framework for Coordinated National Action in Scaling-up Insecticide-Treated Netting Programmes in Africa' promotes coordinated national action and advocates sustained public provision of targeted subsidies to maximise public health benefits, alongside support and stimulation of the private sector. Several countries have already planned or initiated targeted subsidy schemes either on a pilot scale or on a national scale, and have valuable experience which can inform future interventions. The WHO RBM 'Workshop on mapping models for delivering ITNs through targeted subsidies' held in Zambia in 2003 provided an opportunity to share and document these country experiences. This paper brings together experiences presented at the workshop with other information on experiences of targeting subsidies on ITNs, net treatment kits and retreatment services (ITN products) in order to describe alternative approaches, highlight their similarities and differences, outline lessons learnt, and identify gaps in knowledge. We find that while there is a growing body of knowledge on different approaches to targeting ITN subsidies, there are significant gaps in knowledge in crucial areas. Key questions regarding how best to target, how much it will cost and what outcomes (levels of coverage) to expect remain unanswered. High quality, well-funded monitoring and evaluation of alternative approaches to targeting ITN subsidies is vital to develop a knowledge base so that countries can design and implement effective strategies to target ITN subsidies. PMID- 15655011 TI - Safety of the combination of chloroquine and methylene blue in healthy adult men with G6PD deficiency from rural Burkina Faso. AB - New drug combinations against falciparum malaria which are both effective and affordable for Sub-Saharan African populations are urgently needed. The combination of the well-known drugs chloroquine (CQ) and methylene blue (MB) is such a promising new regimen. However, there is some concern that MB could cause development of haemolysis in patients with glucose-6-phosphate dehydrogenase (G6PD) deficiency, a condition which is prevalent in malaria-endemic regions. Against this background, 74 G6PD-deficient but otherwise healthy adult men were given a 3-day oral regimen of a total of 1500 mg CQ and 780 mg MB in the District Hospital of Nouna in north-western Burkina Faso. Haemolysis did not occur, haemoglobin levels remained stable or even rose in the study participants, and the drug regimen was well tolerated. Therefore, standard dosages of MB appear to be safe in G6PD-deficient African populations with predominantly class III G6PD deficiency. PMID- 15655012 TI - Comparison of the OptiMAL rapid test and microscopy for detection of malaria in pregnant women in Nigeria. AB - Malaria during pregnancy causes anaemia in pregnant women and low birthweight in infants. The purpose of this study was to evaluate the OptiMAL rapid malaria test in comparison with standard microscopy for malaria diagnosis in 268 pregnant women attending antenatal clinics at the Jos University Teaching Hospital and the Federal Medical Centre-Gombe in Nigeria. Positive results by either method were confirmed using a polymerase chain reaction assay for malaria. Although the OptiMAL assay did not detect malaria in the blood of pregnant women with low levels of parasitaemia, it may be useful for the detection of placental malaria which predisposes to low birthweight infants. PMID- 15655013 TI - Disparities in parasitic infections, perceived ill health and access to health care among poorer and less poor schoolchildren of rural Cote d'Ivoire. AB - Differences in the state of health between rural and urban populations living in Africa have been described, yet only few studies analysed inequities within poor rural communities. We investigated disparities in parasitic infections, perceived ill health and access to formal health services among more than 4000 schoolchildren from 57 primary schools in a rural area of western Cote d'Ivoire, as measured by their socioeconomic status. In a first step, we carried out a cross-sectional parasitological survey. Stool specimens and finger prick blood samples were collected and processed with standardized, quality-controlled methods, for diagnosis of Schistosoma mansoni, soil-transmitted helminths, intestinal protozoa and Plasmodium. Then, a questionnaire survey was carried out for the appraisal of self-reported morbidity indicators, as well as housing characteristics and household assets ownership. Mean travel distance from each village to the nearest health care delivery structure was provided by the regional health authorities. Poorer schoolchildren showed a significantly higher infection prevalence of hookworm than better-off children. However, higher infection prevalences of intestinal protozoa (i.e. Blastocystis hominis, Endolimax nana and Iodamoeba butschlii) were found with increasing socioeconomic status. Significant negative associations were observed between socioeconomic status and light infection intensities with hookworm and S. mansoni, as well as with several self-reported morbidity indicators. The poorest school-attending children lived significantly further away from formal health services than their richer counterparts. Our study provides evidence for inequities among schoolchildren's parasitic infection status, perceived ill health and access to health care in a large rural part of Cote d'Ivoire. These findings call for more equity-balanced parasitic disease control interventions, which in turn might be an important strategy for poverty alleviation. PMID- 15655014 TI - Evaluation of syphilis in patients with HIV infection in Nigeria. AB - OBJECTIVE: To document the manifestations of syphilis among patients with concurrent HIV infection over a 12-month period. METHOD: Descriptive, cross sectional, hospital-based study of all adult patients with syphilis and HIV infection who attended the skin clinic of the University of Nigeria, Teaching Hospital, Enugu, between July 2000 and June 2001. A standardized questionnaire was used to record age, sex, marital status, occupation and risk factor for HIV infection; initial site of onset of rash/ulcers, duration of the illness, any concomitant affection of mucosa, hair and nails as well as treatments received by each patient prior to presentation. Morphological distribution of lesions, mucosal surface (conjuctival, vulval and rectal) examinations and documentation of concomitant disorders with HIV were noted by the examining dermatologist. Lesional biopsy and dark-field microscopy were undertaken to confirm diagnosis where serologic (non-treponemal and treponemal specific) tests for syphilis were inconsistent with clinical suspicion. Each patient had a routine chest x-ray, mantoux and purified protein derivative (PPD) status taken. RESULTS: Thirty-one patients (21 males) with concurrent syphilis and HIV were seen during the study period. Primary syphilis was diagnosed in nine (29%), secondary syphilis in 20 (64.5%) and latent syphilis in two (6.5%). Neurosyphilis was not observed. Prevalence of syphilis for these patients with concurrent HIV was 2.1%. Mean duration of syphilis was 3.9 months +/- 1.4 and lesions of greatest concern occurred mainly on the genitalia. The glans penis was affected in 10 (32.3%) cases, the penile shaft in seven (22.6%), the oral cavity in five (16.1%), the rectum in six (19.4%) and the vulva in three (0.9%) cases. Nine (29.1%) patients had a history of primary syphilitic chancre, 19 (61.3%) had a past history of sexually transmitted disease (STD)--particularly genital ulcers--while three (9.7%) could not recall any past history of STD. Eighteen (59.3%) had a history of unprotected sex, 16 (51.7%) had multiple sexual partners, four (13.3%) had had oral sex, and one anal sex (3.3%); none admitted to being bisexual. Other relevant risk factors for HIV transmission were blood transfusion within 5 years for three (9.7%) and intravenous drug use in two (6.5%). Some patients had more than one condition as a potential source of exposure. Serological tests were weakly reactive in 17 (48.4%), strongly reactive in nine (29%) and non-reactive in five (16.1%) patients. Three patients exhibited prozone phenomenon. Treatment comprised the syndromic approach, which currently is advocated for use in primary healthcare centres without facilities for aetiological diagnosis of sexually transmitted infections. CONCLUSION: Our cases with concurrent syphilis and HIV/AIDS had unusual manifestations, responded to treatment more slowly and died sooner than cases described in Western literature due to generally lower levels of health. PMID- 15655015 TI - Temporal relationship between the occurrence of fresh noma and the timing of linear growth retardation in Nigerian children. AB - BACKGROUND: Fresh noma (cancrum oris) occurs predominantly in children <4 years of age. The key risk factors are poverty, malnutrition and infections. Evolution from an intraoral inflammation to a grotesque oro-facial gangrene is very rapid. OBJECTIVE: We assessed potential relationship between the occurrence of fresh noma and linear growth retardation (LGR) which is prevalent in deprived Third World infants/children between ages 3 and 30 months. LGR is attributed to malnutrition and chronic immunostimulation by environmental antigens. DESIGN: Anthropometric evaluation of children (n = 91) with fresh noma, ages 0-8 years, in relation to US National Center for Health Statistics Reference values was carried out. Age-matched noma-free, poor village children (n = 151) from similar communities as noma cases, and elite urban children (n = 132) served as control groups. Heights and weights were measured and the height for age (HAZ), weight for age (WAZ) and weight for height (WHZ) scores calculated as indices of stunting, underweight and wasting respectively. Serum level of interleukin (IL) 18, a multifunctional cytokine, was also measured. RESULTS: In the age groups 0-4 and 4-8 years, the percentages of noma children <-2.0SD were 91% and 67% respectively. The corresponding values for the village children were 37% and 24% and significantly different (P < 0.001) from the noma group. Only 7% of the elite children aged 4-8 years were stunted. Low body weight and wasting were prominent features of village and noma groups, but more marked in the latter. Associated with noma was a profound increase (P < 0.001) in IL-18 in comparison with urban controls, and a 34% non-statistically significant increase relative to the village control group. Among other functions, IL-18 induces several pro inflammatory cytokines and the matrix metalloproteinases, influences long bone growth, and consequently may be relevant to growth retardation seen in poor village children and noma victims. CONCLUSION: These results suggest that occurrence of fresh noma was probably programmed very early in life by malnutrition and chronic infections resulting from replacement of breast milk with contaminated, inferior substitutes. Although not investigated, we speculate that children with fresh noma might also be victims of intrauterine growth retardation as noma is most prevalent during the infantile phase of child growth which starts at mid-gestation and tails off at 4 years. PMID- 15655016 TI - Pseudomonas aeruginosa isolates recovered from clinical and environmental samples in Buea, Cameroon: current status on biotyping and antibiogram. AB - OBJECTIVE: The study was aimed at determining the prevalent biotypes of Pseudomonas aeruginosa in the environment of Buea and the susceptibility of isolates to antibiotics. METHODS: One hundred and fifty clinical specimens (urine, wound and sputum) collected from patients attending various health institutions in Buea, and 50 environmental swabs from furniture, appliances and surroundings of these institutions were screened for P. aeruginosa using standard microbiological and biochemical methods. Antimicrobial susceptibility of the isolates was determined by the disc diffusion assay. RESULTS: Fifty-one (25.5%) of the 200 specimens were positive for the organism with urine (30%) being the most common source of isolation. Biochemical characterization grouped the isolates into eight biotypes with biotypes B III (25.59%), B II (23.53%) and B I (21.57%) being the most prevalent. Antimicrobial susceptibility results of isolates revealed 13 antibiotype patterns based on resistance to the antimicrobial agents investigated. The resistance pattern, cefotaxime, gentamicin and tetracycline (CTX(R) GEN(R) TET(R)) was the most common (21.6%) amongst the isolates. Over 40% of the isolates exhibited multi-drug resistance to five or more antibiotics, especially environmental isolates. However, there was a significant difference (P < 0.05) in the susceptibility of isolates to ciprofloxacin (98%), amikacin (90.2%) and netilmicin (80.4%) compared with other drugs used in the study. CONCLUSION: A combination of biotyping and antibiogram, which are relatively cheap and routinely available methods in our environment, could be useful for clinical and epidemiological studies particularly in laboratories in the developing world with limited resources. PMID- 15655017 TI - Intersectoral coordination in Aedes aegypti control. A pilot project in Havana City, Cuba. AB - Background The 55th World Health Assembly declared dengue prevention and control a priority and urged Member States to develop sustainable intersectoral strategies to this end. To provide evidence for the reorientation of the dengue prevention policy in Cuba, we launched an intervention study to document the effectiveness of a local-level intersectoral approach. METHODS: We used a quasi experimental design. Social scientists introduced participatory methods to facilitate dialogue in the biweekly meetings of the intersectoral Health Council of the intervention area. This council subsequently developed an intersectoral plan for dengue prevention, of which the core objective was to design and implement activities for communication and social mobilization. In the control area, routine dengue control activities continued without additional input. Knowledge, attitudes and perceptions of dengue, and entomological indices were compared inside and between the areas before and after the 1-year intervention period. RESULTS: In the intervention area the Health Council elaborated an intersectoral plan for dengue prevention focused on source reduction. The Aedes aegypti control methods consisted in eliminating useless containers in the houses and surroundings, covering tanks, and cleaning public and inhabited areas. It was implemented through communication and social mobilization. The Health Council in the control area occasionally discussed dengue issues but did not develop a coordinated action plan. Good knowledge about breeding sites and disease symptoms increased significantly (by 49.7% and 17.1% respectively) in the intervention area as well as the proportion of respondents eliminating containers in and around their houses (by 44%). No changes were observed in the control area. The House Index in the intervention area was 3.72% at baseline and decreased to 0.61% after 1 year. In the control area it remained stable throughout the study period (1.31% and 1.65% respectively). CONCLUSION: The introduction of a participatory approach by social scientists promotes changes in intersectoral management. This facilitates social mobilization which, in its turn, leads to significant changes in knowledge, attitudes and dengue-related practices in the population and eventually to more effective control of Ae. aegypti. PMID- 15655018 TI - Socioeconomic constraints to effective management of Burkitt's lymphoma in south eastern Nigeria. AB - This paper presents health outcomes and associated socioeconomic factors of 41 children admitted to a tertiary care institution in south-east Nigeria with Burkitt's lymphoma (BL) between 1987 and 2004. BL responds well to chemotherapy and does not pose a significant threat to health in industrialized nations. However, in resource-poor settings where it is endemic, socioeconomic factors significantly affect access to care for affected children, making this readily treatable condition a cause of considerable distress and early death in affected children. Half of the children reported in this paper presented with late stage disease. Although laboratory facilities were available, they were not accessible to all the children. Nearly a quarter of parents of these children could not afford the cost of confirmatory tests, and about a fifth (n = 8; 19.5%) of the children received no chemotherapy because of their parents' inability to pay. Only 21 of 41 children (51.2%) remained on treatment long enough (at least 12 weeks) to enable them to be confirmed either as short-term cure (n = 9; 64.3%), or as early relapse (n = 2; 4.9%). Owing to financial constraint, 13 of the parents (31.7%) withdrew their children against medical advice (n = 7; 17.1%) or left the hospital (n = 6; 14.6%). To address the challenge posed by these factors, we call for the establishment of a regional BL programme in Africa to help establish a critical mass of resources (human and material) to facilitate the development of an effective and accessible control programme in the region. PMID- 15655020 TI - Self-reported and observed female genital cutting in rural Tanzania: associated demographic factors, HIV and sexually transmitted infections. AB - OBJECTIVES: To determine (i) the prevalence and type of female genital cutting (FGC) in a rural multi-ethnic village in Tanzania, (ii) its associated demographic factors, (iii) its possible associations with HIV, sexually transmitted infections (STIs) and infertility and (iv) to assess the consistency between self-reported and clinically observed FGC. METHOD: The study was part of a larger community-based, cross-sectional survey with an eligible female population of 1993. All were human immunodeficiency virus (HIV)-tested and asked whether they were circumcised (n = 1678; 84.2%). Participants aged 15-44 years were interviewed (n = 636; 79.7%), and 399 (50.0%) were gynaecologically examined to screen for STIs and determine the FGC status. RESULTS: At a mean age of 9.6 years, 45.2% reported being circumcised. In the age-group 15-44 years, 65.5% reported being cut, while FGC was observed in 72.5% and categorized as clitoridectomy or excision. The strongest predictors of FGC were ethnicity and religion, i.e. being a Protestant or a Muslim. FGC was not associated with HIV infection, other STIs or infertility. A positive, non-significant association between FGC and bacterial vaginosis was found with a crude odds ratio of 4.6. There was a significant decline of FGC over the last generation. An inconsistency between self-reported and clinically determined FGC status was observed in more than one-fifth of the women. CONCLUSION: The data indicate that both women and clinicians might incorrectly report women's circumcision status. This reveals methodological problems in determining women's circumcision status in populations practising the most common type of FGC. The positive association between FGC and bacterial vaginosis warrants further investigation. PMID- 15655019 TI - Field validation of sensitivity and specificity of rapid test for detection of Brugia malayi infection. AB - We conducted a field study of a rapid test (Brugia Rapid) for detection of Brugia malayi infection to validate its sensitivity and specificity under operational conditions. Seven districts in the state of Sarawak, Malaysia, which are endemic for brugian filariasis, were used to determine the test sensitivity. Determination of specificity was performed in another state in Malaysia (Bachok, Kelantan) which is non-endemic for filariasis but endemic for soil-transmitted helminths. In Sarawak both the rapid test and thick blood smear preparation were performed in the field. The rapid test was interpreted on site, whereas blood smears were taken to the district health centres for staining and microscopic examination. Sensitivity of Brugia Rapid dipstick as compared with microscopy of thick blood smears was 87% (20/23; 95% CI: 66.4-97.2) whereas the specificity was 100% (512/512). The lower sensitivity of the test in the field than in laboratory evaluations (> or =95%), was probably due to the small number of microfilaraemic individuals, in addition to difficulties in performing the test in remote villages by field personnel. The overall prevalence of brugian filariasis as determined by the dipstick is 9.4% (95% CI: 8.2-0.5) while that determined by microscopy is 0.90% (95% CI: 0.5-1.3) thus the dipstick detected about 10 times more cases than microscopy. Equal percentages of adults and children were found to be positive by the dipstick whereas microscopy showed that the number of infected children was seven times less than infected adults. The rapid dipstick test was useful as a diagnostic tool for mapping and certification phases of the lymphatic filariasis elimination programme in B. malayi-endemic areas. PMID- 15655021 TI - Evidence-based periodontal disease prevention and treatment: introduction. PMID- 15655022 TI - Evidence-based periodontology, systematic reviews and research quality. PMID- 15655023 TI - Personal oral hygiene and chronic periodontitis: a systematic review. PMID- 15655024 TI - Does oral health promotion improve oral hygiene and gingival health? PMID- 15655025 TI - Effectiveness of mechanical nonsurgical pocket therapy. PMID- 15655026 TI - How effective is surgical therapy compared with nonsurgical debridement? PMID- 15655027 TI - Which reconstructive procedures are effective for treating the periodontal intraosseous defect? PMID- 15655028 TI - Guided tissue regeneration for periodontal intrabony defects--a Cochrane Systematic Review. PMID- 15655029 TI - Systematic review of the effect of smoking on nonsurgical periodontal therapy. PMID- 15655031 TI - Alternative treatment strategies for weight -- the importance of proper evaluation. PMID- 15655032 TI - Prevalence of obesity in the United States. AB - Obesity is a major public health problem in the United States. Data on measured heights and weights indicates that the prevalence of obesity has significantly increased among the US population over the past 30 years. Data collected from 1999 to 2002 estimates that nearly 1/3 of adults are obese (27.6% of men and 33.2% of women) and one in six children and adolescents is overweight. Increased prevalence of excessive weight is noted among all age, gender and racial/ethnic groups; however, disparities exist. There is a need for further research to better understand why these increases have occurred, why the observed disparities exist and how to reverse these trends. PMID- 15655033 TI - Prevalence of obesity in Turkey. AB - Obesity and overweight are increasing in Turkey according to the field surveys that were carried out a decade apart (TEKHARF 1990 and 2000). The overall prevalence of obesity in adults was 18.6% in the year 1990. Ten years later in 2000, the prevalence was 21.9%, which shows a relative increase rate of 17.7%. As it is true for most of the countries, overweight is more common in men and obesity is more prevalent among women in Turkey. PMID- 15655034 TI - Prevalence of obesity in Great Britain. AB - Since 1980 the prevalence of obesity in Great Britain in adults has almost trebled. Latest figures show that 23% of men and 25% of women were obese in 2002. In children, obesity prevalence is lower but the increase in the prevalence of overweight is similar to the rise in obesity in adults. Data from national surveys also show that there are marked differences in the prevalence of obesity that underpin health inequalities. Obesity is higher in low social classes, some ethnic minority groups particularly from South Asia and in Scotland and Wales relative to England. PMID- 15655035 TI - Adiponectin: action, regulation and association to insulin sensitivity. AB - Adiponectin is a novel adipocyte-specific protein, which, it has been suggested, plays a role in the development of insulin resistance and atherosclerosis. Although it circulates in high concentrations, adiponectin levels are lower in obese subjects than in lean subjects. Apart from negative correlations with measures of adiposity, adiponectin levels are also reduced in association with insulin resistance and type 2 diabetes. Visceral adiposity has been shown to be an independent negative predictor of adiponectin. Thus, most features of the metabolic syndrome's negative associations with adiponectin have been shown. Adiponectin levels seem to be reduced prior to the development of type 2 diabetes, and administration of adiponectin has been accompanied by lower plasma glucose levels as well as increased insulin sensitivity. Furthermore, reduced expression of adiponectin has been associated with some degree of insulin resistance in animal studies indicating a role for hypoadiponectinaemia in relation to insulin resistance. The primary mechanisms by which adiponectin enhance insulin sensitivity appears to be through increased fatty acid oxidation and inhibition of hepatic glucose production. Adiponectin levels are increased by thiazoledinedione treatment, and this effect might be important for the enhanced insulin sensitivity induced by thiazolidinediones. In contrast, adiponectin levels are reduced by pro-inflammatory cytokines especially tumour necrosis factor-alpha. In summary, adiponectin in addition to possible anti-inflammatory and anti-atherogenic effects appears to be an insulin enhancer, with potential as a new pharmacologic treatment modality of the metabolic syndrome and type 2 diabetes. PMID- 15655036 TI - Obesity prevention: a proposed framework for translating evidence into action. AB - Obesity as a major public health and economic problem has risen to the top of policy and programme agendas in many countries, with prevention of childhood obesity providing a particularly compelling mandate for action. There is widespread agreement that action is needed urgently, that it should be comprehensive and sustained, and that it should be evidence-based. While policy and programme funding decisions are inevitably subject to a variety of historical, social, and political influences, a framework for defining their evidence base is needed. This paper describes the development of an evidence based, decision-making framework that is particularly relevant to obesity prevention. Building upon existing work within the fields of public health and health promotion, the Prevention Group of the International Obesity Task Force (IOTF) developed a set of key issues and evidence requirements for obesity prevention. These were presented and discussed at an IOTF workshop in April 2004 and were then further developed into a practical framework. The framework is defined by five key policy and programme issues that form the basis of the framework. These are: (i) building a case for action on obesity; (ii) identifying contributing factors and points of intervention; (iii) defining the opportunities for action; (iv)evaluating potential interventions; and (v) selecting a portfolio of specific policies, programmes, and actions. Each issue has a different set of evidence requirements and analytical outputs to support policy and programme decision-making. Issue 4 was identified as currently the most problematic because of the relative lack of efficacy and effectiveness studies. Compared with clinical decision-making where the evidence base is dominated by randomized controlled trials with high internal validity, the evidence base for obesity prevention needs many different types of evidence and often needs the informed opinions of stakeholders to ensure external validity and contextual relevance. PMID- 15655037 TI - Effect of chitosan on weight loss in overweight and obese individuals: a systematic review of randomized controlled trials. AB - This article aims to determine whether chitosan, a popular, over-the-counter, weight loss supplement, is an effective treatment for overweight and obesity. It is designed as a systematic review of randomized controlled trials. The data sources include the electronic databases Medline, EMBASE, Biosis, CINAHL and Cochrane Central Register of Controlled Trials (CCTR); the specialized websites Controlled Trials, International Bibliographic Information on Dietary Supplements (IBIDS), System for Information on Grey Literature in Europe (SIGLE), Reuter's Health Service, Natural Alternatives International and Pharmanutrients; and bibliographies of relevant journal articles. Included were randomized controlled trials of chitosan with a minimum duration of 4 weeks in adults who were overweight or obese and/or had hypercholesterolaemia at baseline. Fourteen trials involving a total of 1071 participants were included in the review. Analyses involving all trials indicated that chitosan preparations result in a small but statistically significant greater reduction in body weight (weighted mean difference -1.7 kg; 95% confidence interval -2.1, -1.3 kg, P < 0.00001) compared with placebo. Analyses restricted to high-quality studies showed that reductions in weight [-0.6 (-1.2, 0.1) kg, P = 0.11] were less than in lower quality studies [-2.3 (-2.7, -1.8) kg, P < 0.00001]. Results obtained from high-quality trials indicate that the effect of chitosan on body weight is minimal and unlikely to be of clinical significance. PMID- 15655038 TI - A review of psychosocial pre-treatment predictors of weight control. AB - Prompted by the large heterogeneity of individual results in obesity treatment, many studies have attempted to predict weight outcomes from information collected from participants before they start the programme. Identifying significant predictors of weight loss outcomes is central to improving treatments for obesity, as it could help professionals focus efforts on those most likely to benefit, suggest supplementary or alternative treatments for those less likely to succeed, and help in matching individuals to different treatments. To date, however, research efforts have resulted in weak predictive models with limited practical usefulness. The two primary goals of this article are to review the best individual-level psychosocial pre-treatment predictors of short- and long term (1 year or more) weight loss and to identify research needs and propose directions for further work in this area. Results from original studies published since 1995 show that few previous weight loss attempts and an autonomous, self motivated cognitive style are the best prospective predictors of successful weight management. In the more obese samples, higher initial body mass index (BMI) may also be correlated with larger absolute weight losses. Several variables, including binge eating, eating disinhibition and restraint, and depression/mood clearly do not predict treatment outcomes, when assessed before treatment. Importantly, for a considerable number of psychosocial constructs (e.g. eating self-efficacy, body image, self-esteem, outcome expectancies, weight specific quality of life and several variables related to exercise), evidence is suggestive but inconsistent or too scant for an informed conclusion to be drawn. Results are discussed in the context of past and present conceptual and methodological limitations, and several future research directions are described. PMID- 15655039 TI - Who succeeds in maintaining weight loss? A conceptual review of factors associated with weight loss maintenance and weight regain. AB - Weight loss is difficult to achieve and maintaining the weight loss is an even greater challenge. The identification of factors associated with weight loss maintenance can enhance our understanding for the behaviours and prerequisites that are crucial in sustaining a lowered body weight. In this paper we have reviewed the literature on factors associated with weight loss maintenance and weight regain. We have used a definition of weight maintenance implying intentional weight loss that has subsequently been maintained for at least 6 months. According to our review, successful weight maintenance is associated with more initial weight loss, reaching a self-determined goal weight, having a physically active lifestyle, a regular meal rhythm including breakfast and healthier eating, control of over-eating and self-monitoring of behaviours. Weight maintenance is further associated with an internal motivation to lose weight, social support, better coping strategies and ability to handle life stress, self-efficacy, autonomy, assuming responsibility in life, and overall more psychological strength and stability. Factors that may pose a risk for weight regain include a history of weight cycling, disinhibited eating, binge eating, more hunger, eating in response to negative emotions and stress, and more passive reactions to problems. PMID- 15655042 TI - Treatment of hepatitis D. AB - Delta virus related chronic hepatitis is difficult to treat. The response to alpha-interferon (IFN), which still represents the only therapy for chronic hepatitis D, varies widely and occurs at different times from the beginning of treatment. The rate of response is proportional to the dose of IFN, with 9 million units (MU) three times a week being more effective than 3 MU thrice weekly. Sustained responses are unusual and are accompanied by the clearance of serum hepatitis B virus surface antigen (HBsAg), seroconversion to anti-HBs and improvement of liver histology. Although disease of a short-standing may respond better to therapy, clear predictors of response are still unidentified. Besides IFN, other therapeutic approaches such as immunosuppressive drugs, acyclovir, ribavirin and thymosin, have been unhelpful. Available evidence does not support the use of deoxynucleotide analogues. Famciclovir has no effect on disease activity and hepatitis D virus (HDV)-RNA levels. Twelve- or 24-month lamivudine treatment does not significantly affect biochemical, virological or histological parameters. Pegylated-IFN could represent a reasonable therapeutic option in the long-term treatment required for chronic hepatitis D. Antisense oligonucleotides and prenylation inhibitors hold promise as therapeutic agents of the future. Liver transplantation provides a valid option for end-stage HDV liver disease; the risk of re-infection is lower for HDV than for HBV under long-term administration of hyperimmune serum against HBsAg. Molecularly tailored drugs capable of interfering with crucial viral replicative processes of HDV appear to be the best prospect in the treatment of hepatitis D. PMID- 15655043 TI - Therapeutic modalities in hepatitis C: challenges and development. AB - Our understanding of the pathogenicity of hepatitis C virus (HCV) is based on patients infected chronically for >20 years. The lack of a suitable animal model, the narrow host range of the virus, and the protracted onset of liver disease induced by HCV have hampered advances in treatment. In spite of these problems, we identified patient and viral characteristics that predict responses to current therapies, including HCV genotype, viral load, body weight, age, liver histology, co-infection with HIV and treatment adherence and tolerance. Interferon (IFN) alpha was the first therapy for chronic HCV infection. The combination of IFN plus ribavirin increases sustained virological response rates compared with IFN alone. Two pegylated IFNs have been developed and are widely approved for the treatment of chronic hepatitis C: peginterferon alpha-2a (40 KD), and pegylated IFN alpha-2b (12 KD). These products have reduced systemic clearance, prolonged half-lives and reduced antigenicity compared with conventional IFN. The reduced clearance results in sustained plasma levels of the drug and allows for once weekly dosing. Pegylated IFN alpha-2b (12 KD) has a small, linear polyethylene glycol (PEG) moiety and has an intermediate duration of activity; peginterferon alpha-2a (40 KD) incorporates a large, branched-chain PEG moiety and has a longer half-life than both conventional IFN alpha and pegylated IFN alpha-2b (12 KD). The combination of a pegylated IFN plus ribavirin significantly increases sustained virological response rates compared with conventional IFN plus ribavirin in patients with chronic hepatitis C and is now recognized as the standard of care for these patients. PMID- 15655044 TI - Morphological alteration and biological properties of hepatocytes not related to tumorigenesis following transfection with HCV core protein. AB - The hepatitis C virus (HCV) core protein is supposed to play a critical role in HCV-mediated human liver disease with its capabilities to regulate the growth rate of hepatocytes and to partially contribute to the pathogenesis of hepatocellular carcinoma in association with cellular oncogenes. In this study, to analyse the possible pathological mechanism of the HCV core protein, human primary embryo hepatocytes transfected with HCV core were monitored by immunofluorescence, reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blot. The morphological changes and biological properties of the transfected hepatocytes were also studied. The results showed that the HCV core gene integrated in the cellular genome and the protein expressed in the transfected hepatocyte, could be detected following serial passage at both the mRNA and protein level. The proliferation assays indicated that hepatocytes transfected with the HCV core gene alone did not exhibit any tumorigenic tendency. Meanwhile, the morphological alterations of these cells demonstrated obvious changes in size, and large vacuolar degeneration. In conclusion, the hepatocytes transfected with the HCV core gene revealed that the core protein expressed induced pathological changes of degeneration, probably related indirectly to tumorigenicity. PMID- 15655045 TI - Gene expression profile of T-cell-specific chemokines in human hepatocyte-derived cells: evidence for a synergistic inducer effect of cytokines and hepatitis C virus proteins. AB - Increased levels of chemokines (CK) in chronic hepatitis C virus (HCV) infection have been found. Given that NS5A and core can function as transcriptional transactivators, we aimed to determine whether these HCV proteins might induce CK gene expression in human hepatocyte-derived cells. We assessed (i) regulated upon activation, normal T cells expressed and activated (RANTES), interferon gamma inducible protein-10 (IP-10), and monokine induced by interferon-gamma (MIG) mRNA levels in NS5A and core stably transfected Chang liver (CHL) cells, stimulated or not with a cytokine mixture (CM), by reverse transcriptase-polymerase chain reaction (RT-PCR) and (ii) quantitative enzyme-linked immunosorbent assay (ELISA) measurements of these CK in the supernatants of CHL cells. Induction of RANTES transcripts in resting HCV-transfected cells was clearly observed, being augmented fourfold in resting NS5A-transfected cells and threefold in resting core-transfected cells over that in resting mock-transfected (control) cells, as well as to a similar extent in CM-stimulated NS5A- and core-transfected cells. Increased RANTES secretion followed the same pattern observed for mRNA expression. Both IP-10 and MIG, such as mRNA and protein levels, were undetectable in resting HCV-transfected and -untransfected cells, whereas IP-10 and MIG mRNA expression was increased by seven- and fivefold in CM-stimulated NS5A-transfected cells and by 10- and 3.5-fold in CM-stimulated core-transfected cells, respectively, above that in CM-stimulated control cells. IP-10 and MIG secretion was enhanced by 2.6- and threefold in CM-stimulated NS5A-transfected cells and by 3.6-fold and 3.7-fold in CM-stimulated core-transfected cells, respectively over that in CM-stimulated control cells. These results demonstrate that NS5A and core proteins, alone or by the synergistic effect of cytokines, are capable of upregulating RANTES, IP-10 and MIG gene expression in cultured human hepatocyte-derived cells. PMID- 15655046 TI - Activation and function of hepatic NK cells in hepatitis B infection: an underinvestigated innate immune response. AB - Natural killer (NK) cells are abundant in the normal liver, accounting for around one-third of intrahepatic lymphocytes and are important in the defence against hepatitis B virus (HBV) infection as innate immune responses. In this review, we discuss the mechanisms of hepatic NK cell activity against HBV. Whether directly activated by HBV infection or indirectly activated by other lymphocytes such as NKT cells or antigen-presenting cells (APCs), hepatic NK cells exert their anti viral functions by natural cytotoxicity and production of high levels of cytokines. However, activated NK cells play an important role in regulating adaptive immune responses by interaction with other lymphocytes such as T, B and APCs. In addition, NK cells may contribute to the lymphocyte-mediated liver injury during HBV infection that was previously considered to be mediated only by CD8+ T cells or/and NKT cells. PMID- 15655047 TI - The minimum number of clones necessary to sequence in order to obtain the maximum information about hepatitis C virus quasispecies: a comparison of subjects with and without liver cancer. AB - Most studies of hepatitis C virus (HCV) quasispecies have reported the results of sequencing only three to five clones per sample. The possibility that sequencing so few clones might not provide a representative picture of the quasispecies present in a sample has never been evaluated. The present study was conducted to evaluate whether sequencing greater numbers of clones results in better information about the HCV quasispecies number and distribution, and to compare the HCV quasispecies in liver cancer cases and controls. RNA was extracted from serial serum samples from six subjects with HCV-associated liver cancer and 11 age- and sex-matched HCV-infected controls without liver cancer. The hypervariable region 1 (HVR1) of the HCV genome was amplified, cloned, and sequenced. For further studies of 12 serum samples from two liver cancer cases and two matched controls, successive groups of 10 additional clones were sequenced up to a total of 50 clones per serum sample. When only 10 clones were sequenced from each specimen, no consistent differences were seen between the number of HCV quasispecies in the six liver cancer cases and the 11 controls. However, sequencing 40 clones from each of 12 samples from two liver cancer cases and two controls revealed a greater number of quasispecies in liver cancer cases than in controls. Testing an additional 10 clones (50 clones per sample) did not significantly increase the number of quasispecies detected. PMID- 15655048 TI - Patient concerns regarding chronic hepatitis C infections. AB - Counselling of patients with chronic hepatitis C infections is often limited to discussions regarding how the virus is transmitted and what can be done to decrease the risk of transmission to others. The purpose of the present study was to document the principal concerns of newly diagnosed and follow-up patients with chronic hepatitis C, and thereby enhance counselling strategies and content. Seventy newly diagnosed and 115 follow-up patients with chronic hepatitis C virus (HCV) infection were initially asked in an open-ended manner (volunteered concerns) and then to prioritize from a prepared list of seven potential concerns (prioritized concerns), to identify those concerns that were of utmost importance to them. The most common volunteered concerns of newly diagnosed patients in decreasing order were: disease progression (27%), premature death (19%), infecting family members (13%), side-effects of treatment (11%) and miscellaneous others. In decreasing order, prioritized concerns included: infecting family members, development of liver cancer, infecting others, development of cirrhosis, social stigma of having liver disease, need for liver transplant and loss of employment. The principal volunteered and prioritized concerns of follow-up patients were similar to those of newly diagnosed patients. Volunteered and prioritized concerns were relatively consistent across the different genders, age groups, ethnic backgrounds, education level, marital status, employment, modes of viral acquisition and in the case of follow-up patients, duration of follow-up. These results indicate that health care providers who focus counselling efforts exclusively on viral transmission are unlikely to address other important concerns of newly diagnosed and follow-up patients with chronic HCV infection. PMID- 15655049 TI - Treatment of histologically mild hepatitis C virus infection with interferon and ribavirin: a multicentre randomized controlled trial. AB - Current guidelines advocate no treatment for patients with histologically mild hepatitis C virus (HCV) infection. This was a UK multicentre randomized controlled trial comparing alpha-interferon (3 MU thrice weekly) + ribavirin (1000-1200 mg/day) for 48 weeks with no treatment in treatment naive, adult patients with histologically mild chronic HCV infection. The aim was to compare benefits, safety and efficacy of combination therapy with alpha-interferon 2b and ribavirin for 48 weeks with no treatment (current standard management) in this patient group. In the treatment group 32 of 98 (33%) patients achieved a sustained virological response (SVR). Patients infected with genotype 1 had a lower SVR than those infected with genotype non-1 (18% vs 49% P = 0.02). No patients who failed to achieve a 2-log drop in viral load at 12 weeks achieved SVR. Improvements in quality of life 24 weeks postcessation of therapy compared with baseline using the SF-36 questionnaire measures were observed in the treated group. For patients with mild HCV infection with viral genotype non-1, the results are sufficiently good to suggest that therapeutic decisions should no longer be biopsy-driven. For patients infected with genotype 1, a liver biopsy is still indicated as the low chance of SVR is outweighed by an unacceptable burden of side-effects. Patients who fail to respond by 12 weeks of therapy should have their treatment curtailed early. PMID- 15655050 TI - Activity of adefovir dipivoxil against all patterns of lamivudine-resistant hepatitis B viruses in patients. AB - One hundred and thirty-one post-liver transplantation patients with chronic hepatitis B and failing lamivudine therapy with detectable serum hepatitis B virus (HBV) deoxyribonucleic acid by hybridization assays or > or =1 x 10(6) copies/mL by polymerase chain reaction, and elevated alanine transaminase levels despite continuous lamivudine, were enrolled in an open-label study of adefovir dipivoxil. The B and C domains of HBV polymerase were sequenced for baseline samples to determine the presence of lamivudine resistance mutations. The results showed that 98% of the samples had tyrosine-methionine-aspartate-aspartate (YMDD) mutations, indicating a strong correlation between the above clinical definition of lamivudine treatment failure and the presence of YMDD mutations. In addition to the rtM204V/I and the rtL180M mutations, the mutation rtV173L was identified in 19% of patients. Four major patterns of lamivudine-resistant HBV were identified: rtL180M + rtM204V (60%), rtV173L + rtL180M + rtM204V (19%), rtM204I (9%) and rtL180M + rtM204I (9%). Treatment with adefovir dipivoxil showed similar antiviral efficacy in patients with lamivudine-resistant virus from all four patterns. PMID- 15655051 TI - Predicting progressive hepatic fibrosis stage on subsequent liver biopsy in chronic hepatitis C virus infection. AB - Retrospective cross-sectional studies indicate that 20% with chronic hepatitis C virus (HCV) infection become cirrhotic within 20 years. Known risk factors for advanced hepatic fibrosis include age at time of infection, male sex, excess alcohol consumption and cytokine polymorphisms. Prospective study to assess and identify factors predictive of change in hepatic fibrosis stage in chronic HCV infection by interval protocol liver biopsy was performed. One hundred and five patients with paired liver biopsy specimens separated by a mean 41 months were recruited from a cohort of 823 HCV carriers. Five per cent developed worsening hepatic fibrosis by more than two stages. In 43% there was no change in fibrosis stage. Excessive alcohol intake currently (P = 0.037) or previously (P = 0.07) predicted progression. In contrast, always having a normal alanine transaminase (P = 0.038) and always being negative in serum for HCV RNA (P =0.067) predicted no progression. Three models were developed to predict outcome. Progressive fibrosis was predicted by baseline fibrosis (P = 0.018), steatosis (P = 0.02) and age (P = 0.017). The rate of progressive fibrosis was predicted by baseline fibrosis (P = 0.0002), steatosis (P =0.039) and lobular inflammation (P = 0.09). Fibrosis stage on the second biopsy was predicted by baseline fibrosis alone (P = 0.01). The rate of progression varies widely. Alcohol misuse is an important co factor. Progressive fibrosis can be predicted at first liver biopsy, where baseline fibrosis is most critical, allowing targeted therapy for those with early disease and a significant risk of progression. PMID- 15655052 TI - Reasons for non-treatment of hepatitis C in veterans in care. AB - We prospectively studied 354 patients with hepatitis C virus (HCV) infection who were referred to a hepatology specialty clinic to find the reasons for non treatment of HCV. The median age was 48 years (range 27-77 years), 98.5% were male and 71% were white. Seventy per cent of the patients were not treated. The most common reasons for non-treatment were non-adherence to follow-up visit (24%), normal liver enzymes (14%), concurrent medical problems (11%), alcohol and drug use (9%), psychiatric problems (7%), advanced liver disease (7%), referral for transplant evaluation (6.4%) and patient refusal, transfer of care to another facility and non-detectable HCV RNA levels (5% each). The reason was not recorded for 5% of the patients and was treatment deferred in 2.4% while waiting for pegylated interferon approval. Non-treatment was more likely in patients with less than 12 years of education and a history of incarceration. Patients who were lost to follow-up and refused treatment were more likely to have current alcohol and drug use and a history of incarceration. PMID- 15655053 TI - The rate of treatment of chronic hepatitis C in patients co-infected with HIV in an urban medical centre. AB - Hepatitis C virus (HCV) and human immunodeficiency virus (HIV) co-infection is common. HIV co-infection results in a higher rate of histologic progression and shorter interval to HCV-related cirrhosis. Successful treatment of HCV with interferon-based therapy reduces the morbidity and mortality of patients. Significant factors may limit the availability of treatment in co-infected patients. The rate of treatment of HCV and limiting factors to treatment in a co infected population in an urban setting were determined. A retrospective review of co-infected patients was conducted at our liver and gastrointestinal (GI) clinics for treatment of HCV from July 2001 to June 2002. Treatment of HCV and reasons for nontreatment were recorded. A total of 104 HCV/HIV co-infected patients were identified. Seventy-two per cent were males. Mean age was 47.2 years (32-72). Seventy-four of the 82 (90%) with identifiable risk factors for HCV infection had a history of intravenous drug use (IVDU). Twenty per cent (21/104) of the total underwent a liver biopsy. Sixty-seven per cent who had a liver biopsy were treated. Overall, sixteen patients were treated. Eighty-eight (85%) patients were not treated for the following reasons: 13 refused treatment, and 75 were ineligible. Of the ineligible patients, 40% were noncompliant with visits, 15% were active substance abusers, 13% had decompensated cirrhosis, 8% had significant active psychiatric conditions and 24% had significant co-morbid disease. A majority of patients co-infected with HCV/HIV had a IVDU history. Most co-infected patients were not eligible for HCV treatment. A majority of noncandidates had potentially modifiable psychosocial factors leading to nontreatment. PMID- 15655054 TI - The effect of adherence to therapy on sustained response in daily or three times a week interferon alpha-2b plus ribavirin treatment of naive and nonresponder chronic hepatitis C patients. AB - The aim was to demonstrate adherence to treatment has been suggested to enhance rates of sustained response in patients with hepatitis C. In this study, we evaluated the effect of drug dosage reduction or the duration of the expected therapy in patients treated with interferon (IFN)-alpha2b plus ribavirin. Virologic response rates were re-analysed according to compliance to therapy in (i) 301 naive and (ii) 142 nonresponders to previous IFN therapy treated with either IFN 5 MU TIW for 8 weeks followed by IFN 3 MU TIW for 40 weeks plus ribavirin or IFN 3 MU QD for 16 weeks followed by IFN 3 MU TIW for 24 weeks plus ribavirin. Patients were separated into those who adhered to > or =80% of their intended treatment schedule (dose of both drugs and duration) and those who did not. Compliance to treatment resulted in significantly higher response rates in both groups of patients: 43.93% compared with 6.90% of noncompliant naive patients and 30.77% compared with 10.53% of nonresponder patients. Compliance to treatment was found to have a similar effect when the results were analysed according to HCV genotype. Our findings suggest that compliance to treatment for > or =80% of the intended treatment schedule results in significantly higher sustained response rates in both naive and nonresponder patients. Consequently, every effort should be made to improve patient adherence to therapy. PMID- 15655055 TI - Prophylactic SSRI during interferon alpha re-therapy in patients with chronic hepatitis C and a history of interferon-induced depression. AB - Only limited data are available on selective serotonin re-uptake inhibitor (SSRI) prophylaxis for antiviral re-treatment in hepatitis C patients with previous interferon-induced major depressive episodes. Therefore, we investigated the efficacy and safety of secondary SSRI prophylaxis in these patients. In a prospective and longitudinal study, repeated psychometric testing (Hospital Anxiety and Depression Scale) was performed before, during, and after antiviral re-treatment. Chronic hepatitis C virus (HCV)-infected patients, who had been psychometrically monitored during an unsuccessful previous antiviral therapy, and had developed major depression were included. Interferon re-therapy with SSRI prophylaxis was started (n = 8). The reference group was comprised of HCV patients without a history of interferon-associated depression and also a group who were previously unsuccessfully treated with interferon and were re-treated without SSRI prophylaxis (n = 9). All patients receiving SSRI prophylaxis were able to complete interferon re-therapy as scheduled. As in the first therapeutic course, depression scores were significantly elevated during re-treatment also (P < 0.001). Depression scores were significantly lower (P =0.036) during interferon re-therapy with SSRI prophylaxis. Reference group subjects showed similar depression scores during first therapy and re-therapy (P > 0.05). In conclusion, hepatitis C patients with a history of interferon-induced major depression can be successfully re-treated with peginterferon/ribavirin and concomitant SSRI prophylaxis. In these patients, SSRI prophylaxis is safe and efficacious and should be considered, if antiviral re-therapy is indicated. PMID- 15655056 TI - Seroepidemiology of hepatitis A in patients with chronic liver disease. AB - Hepatitis A virus (HAV) rarely causes fulminant hepatic failure in the general population. Yet it is a cause of significant morbidity and mortality in patients with chronic liver disease (CLD), in whom routine HAV vaccination is recommended. However, studies of HAV seroprevalence and exposure predictors in populations with CLD are scarce. We have studied a cohort of 473 patients with various causes of CLD between July 2000 and June 2002. Patients were stratified on the basis of age, gender, ethnicity and aetiology of liver disease. The HAV seroprevalence in patients with CLD was compared with that in the general population. We used a logistic regression analysis to identify independent predictors of HAV exposure. Of the 473 patients studied, HAV seroprevalence was available for 454 individuals. HCV, HBV, alcohol, and HCV and alcohol were the causes of CLD in 337, 72, 37 and eight patients, respectively. The overall HAV seroprevalence was 55% in the studied cohort. The age-stratified HAV prevalence for ages 21-30, 31 40, 41-50, 51-60, 61-70 and greater than 70 years was 44, 51, 44, 63, 65 and 64%, respectively. Hispanic ethnicity, Asian ethnicity, alcohol use and ages of 51-70 years were found to be independent predictive variables of prior exposure to HAV. HAV infection in patients with CLD causes considerable morbidity and mortality. We demonstrated that age-stratified seroprevalence of HAV in patients with CLD of various aetiologies is significantly higher than that of the general population, and identified several independent predictors of HAV prior exposure. PMID- 15655057 TI - Usefulness of a new immuno-radiometric assay to detect hepatitis C core antigen in a community-based population. AB - A new immuno-radiometric assay (IRMA) to detect hepatitis C virus (HCV) core antigen (HCVcAg) has been developed. The aim of the present study was to investigate the sensitivity and specificity of this IRMA to measure HCV antigenemia, based on the detection of HCV RNA as the gold standard, and to assess the utility of the IRMA in a community-based population. Anti-HCV positive residents in a hyperendemic area of HCV infection in Japan were studied. Serum levels of HCVcAg were measured using IRMA, and the presence of HCV RNA was determined by a qualitative reverse transcription-polymerase chain reaction (RT PCR) assay. The sensitivity and the specificity of the IRMA were 96.4 and 100%, respectively. The sensitivity of the IRMA was similar between serological HCV group I (HCV genotypes 1a and 1b) (97.6%) and group II (HCV genotypes 2a and 2b) (94.0%). There was a strong correlation between serum HCVcAg level and HCV-RNA measured by a quantitative RT-PCR (r = 0.832, P < 0.0001). There also was a very strong correlation of HCVcAg level between IRMA measurements performed on serum and those performed on plasma (r = 0.984, P < 0.0001). In conclusion, this new IRMA is useful for the detection of HCV core antigen in a community-based population. PMID- 15655058 TI - The discovery of aequorin and green fluorescent protein. PMID- 15655059 TI - Recent developments and new strategies in scanning electron microscopy. AB - In addition to improvements in lateral resolution in scanning electron microscopy, recent developments of interest here concern extension of the incident beam energy, E(0), over two decades, from approximately 20 keV to approximately 0.1-0.5 keV and the possibility of changing the take-off emission, alpha, of detected secondary electrons. These two degrees of freedom for image acquisition permit a series of images of the same field of view of a specimen to be obtained, each image of the series differing from the others in some aspect. The origins of these differences are explored in detail and they are tentatively interpreted in terms of the change in the secondary electron emission yield delta vs. E(0), delta = f(E(0)), and also of the change in delta vs. alpha, partial differentialdelta/ partial differentialalpha. Various origins for the chemical contrast and topographic contrast have been identified. Illustrated by correlating a secondary electron image and a backscattered electron image, use of the scatter diagram technique facilitates image comparison. The difference between the lateral resolution and the size of the minimum detectable detail is outlined to avoid possible errors in nanometrology. Some aspects related to charging are also considered and possible causes of contrast reversal are suggested. Finally, the suggested strategy consists of the acquisition of various images of a given specimen by changing one parameter: primary beam energy and take-off angle for conductive specimens; working distance or beam intensity for high-resolution experiments; scanning frequency for insulating specimens. PMID- 15655060 TI - Imaging fluorescence lifetime heterogeneity applied to GFP-tagged MHC protein at an immunological synapse. AB - Fluorescence imaging of green fluorescent protein (GFP) may be used to locate proteins in live cells and fluorescence lifetime imaging (FLIM) may be employed to probe the local microenvironment of proteins. Here we apply FLIM to GFP-tagged proteins at the cell surface and at an inhibitory natural killer (NK) cell immunological synapse (IS). We present a novel quantitative analysis of fluorescence lifetime images that we believe is useful to determine whether apparent FLIM heterogeneity is statistically significant. We observe that, although the variation of observed fluorescence lifetime of GFP-tagged proteins at the cell surface is close to the expected statistical range, the lifetime of GFP-tagged proteins in cells is shorter than recombinant GFP in solution. Furthermore the lifetime of GFP-tagged major histocompatibility complex class I protein is shortened at the inhibitory NK cell IS compared with the unconjugated membrane. Following our previous work demonstrating the ability of FLIM to report the local refractive index of GFP in solution, we speculate that these lifetime variations may indicate local refractive index changes. This application of our method for detecting small but significant differences in fluorescence lifetimes shows how FLIM could be broadly useful in imaging discrete membrane environments for a given protein. PMID- 15655061 TI - A resin impression SEM technique for examining the glass-ionomer cement chemical fusion zone. AB - Examination of the auto-cure glass-ionomer cement (GIC)-tooth interface using conventional SEM techniques causes severe dehydration and subsequent fracturing of the GIC, hampering accurate assessment of the GIC-tooth interface. A simple, accurate impression technique was developed to examine the GIC chemical fusion zone. Samples of GIC were bonded to prepared cavities and tooth surfaces that had been conditioned with 10% polyacrylic acid for 10 s and allowed to mature for 24 h in neutral buffered saline, sectioned under water and then acid etched with 37% phosphoric acid for 30 s to remove sectioning smear layer and to highlight surface morphology, rinsed and gently dried. Optibond Solo Plus was then applied and photo-polymerized. Increments of posterior hybrid composite were then bonded onto the Optibond and photo-polymerized. The tooth sections with attached GIC were then dissolved off the bonded composite with 18% HCl for 4 days. Subsequent SEM examination of the resin impressions created with this technique provided high-quality, detailed images of the GIC chemical fusion zone, without the development of the fracture artefacts associated with the desiccation of GIC. The resin impression technique described proved to be a simple and successful method for providing accurate SEM images of the GIC-tooth chemical fusion. PMID- 15655062 TI - Shape discrimination by total curvature, with a view to cancer diagnostics. AB - This paper investigates the use of total curvature for shape discrimination of objects via profiles of their planar sections (not assumed to be star shaped). Methods of estimating total curvature from observation of a finite number of points on the boundary of the object are investigated, including a simple discrete approximation method and various interpolation methods. Total curvature is capable of revealing shape differences on a local scale, as demonstrated by the analysis of two data sets of malignant and normal or benign tumour cell nuclear profiles. PMID- 15655063 TI - Statistical analysis of the three-dimensional structure of centromeric heterochromatin in interphase nuclei. AB - Translocation of genes into the pericentromeric heterochromatin occurs during cellular differentiation and leads to a long-term silencing of these genes. Consequently, a structural remodelling of this heterochromatin compartment is observed during differentiation but this remains to be defined from a topological point of view. In a previous study, we analysed the three-dimensional (3D) distribution patterns of centromere clusters (chromocentres) by confocal scanning laser microscopy and found that differentiation of the promyelocytic leukaemia cell line NB4 along the neutrophil lineage is associated with a progressive clustering of centromeres. This clustering was reflected by a decreased number of detectable chromocentres, i.e. groups of centromeres with a distance below the diffraction-limited resolution of optical microscopy. The purpose of this study was to perform a statistical analysis of the 3D distribution of chromocentres in NB4 cells. Several point field characteristics (Ripley's K-function, L-function, pair correlation function, nearest-neighbour distribution function) were investigated to describe the topology of chromocentres during differentiation of NB4 cells. The pair correlation function revealed a higher frequency of chromocentre distances between 350 nm and 800 nm in undifferentiated NB4 cells as compared with differentiated cells. The L-function and the nearest-neighbour distribution function confirmed these results. These data imply the existence of intranuclear heterochromatin zones formed by functionally related centromeric regions. In view of the observed decrease in the number of detectable chromocentres during differentiation, we hypothesize that these zones with a diameter of 350-800 nm in undifferentiated NB4 cells contract into zones with a diameter below 350 nm in differentiated cells. PMID- 15655064 TI - Interactions between saporin, a ribosome-inactivating protein, and DNA: a study by atomic force microscopy. AB - Saporins are enzymes belonging to the PNAG class (polynucleotide: adenosine glycosidase), plant enzymes commonly known as ribosome-inactivating proteins (RIP), as a result of their property of irreversibly damaging eukaryotic ribosomes. Direct imaging with tapping-mode atomic force microscopy (AFM) has been used to study pGEM-4Z plasmid DNA binding to the saporin-SO6 (isoform from Saponaria officinalis seeds). Saporin wrapped the plasmidic DNA, and distribution of the enzyme molecules along the DNA chain was markedly variable; plasmid digested with saporin-SO6 appeared fragmented or topologically modified. The supercoiled DNA strands were cleaved, giving rise to a linearized form and to relaxed forms. Electrophoretic analysis of the effect of standard preparations of saporin-SO6 on pGEM-4S confirmed the presence of DNA strand-cleaving activity. PMID- 15655065 TI - Statistical characterization of TEM images of silica-filled rubber. AB - Transmission electron microscopy is used to study the micro-dispersion of silica fillers within the polymer matrix of rubber. The resulting grey-value images are interpreted as realizations of random fields and are characterized by means of variograms. The so-called Cauchy class is a suitable model for this purpose. Statistical analysis shows that different filler dispersion properties are reflected in different variogram parameters. As a case study, the random field approach is demonstrated for four exemplary rubber compounds. PMID- 15655066 TI - Single-particle visualization of assembly: I. Dimerization in a planar zone. AB - Summary Single-particle fluorescence microscopy of association/dissociation is required for analysis of biological assembly reactions. Toward achieving this goal, Wang et al. (J. Microsc., 2004, 213, 101-109) used molten agarose to concentrate thermally diffusing particles in a thin zone of solution next to the surface of a coverglass (plane of concentration). The present study details the first real-time, single-particle analysis of the association/dissociation of thermally diffusing particles in the plane of concentration. The test particles were procapsids of bacteriophage lambda (radius = 31 nm). Quantification of thermal motion was developed and used to determine whether co-diffusing particles were bound to each other. The data are explained by (1) the presence of a molten agarose-generated barrier that is 93-155 nm from the coverglass surface, and (2) non-random orientation of procapsid dimers in the plane of concentration. PMID- 15655067 TI - Noise suppression of point spread functions and its influence on deconvolution of three-dimensional fluorescence microscopy image sets. AB - The point spread function (PSF) is of central importance in the image restoration of three-dimensional image sets acquired by an epifluorescent microscope. Even though it is well known that an experimental PSF is typically more accurate than a theoretical one, the noise content of the experimental PSF is often an obstacle to its use in deconvolution algorithms. In this paper we apply a recently introduced noise suppression method to achieve an effective noise reduction in experimental PSFs. We show with both simulated and experimental three-dimensional image sets that a PSF that is smoothed with this method leads to a significant improvement in the performance of deconvolution algorithms, such as the regularized least-squares algorithm and the accelerated Richardson-Lucy algorithm. PMID- 15655068 TI - Combined AFM and confocal fluorescence microscope for applications in bio nanotechnology. AB - We present a custom-designed atomic force fluorescence microscope (AFFM), which can perform simultaneous optical and topographic measurements with single molecule sensitivity throughout the whole visible to near-infrared spectral region. Integration of atomic force microscopy (AFM) and confocal fluorescence microscopy combines the high-resolution topographical imaging of AFM with the reliable (bio)-chemical identification capability of optical methods. The AFFM is equipped with a spectrograph enabling combined topographic and fluorescence spectral imaging, which significantly enhances discrimination of spectroscopically distinct objects. The modular design allows easy switching between different modes of operation such as tip-scanning, sample-scanning or mechanical manipulation, all of which are combined with synchronous optical detection. We demonstrate that coupling the AFM with the fluorescence microscope does not compromise its ability to image with a high spatial resolution. Examples of several modes of operation of the AFFM are shown using two-dimensional crystals and membranes containing light-harvesting complexes from the photosynthetic bacterium Rhodobacter sphaeroides. PMID- 15655070 TI - Factors associated with reporting multiple causes of death. AB - BACKGROUND: There is analytical potential for multiple cause of death data collected from death certificates. This study examines relationships of multiple causes of death as a function of factors available on the death certificate (demographics of decedent, place of death, type of certifier, disposal method, whether an autopsy was performed, and year of death). METHODS: Data from 326,332 Minnesota death certificates from 1990-1998 are examined. Underlying and non underlying causes of death are examined (based on record axis codes) as well as demographic and death-related covariates. Associations between covariates and prevalence of multiple causes of death and conditional probability of underlying compared to non-underlying causes of death are examined. The occurrence of ischemic heart disease or diabetes as underlying causes are specifically examined. RESULTS: Both the probability of multiple causes of death and the proportion of underlying cause compared to non-underlying cause of death are associated with demographic characteristics of the deceased and other non-medical conditions related to filing death certificate such as place of death. CONCLUSIONS: Multiple cause of death data provide a potentially useful way of looking for inaccuracies in reporting of causes of death. Differences across demographics in the proportion of time a cause is selected as underlying compared to non-underlying exist and can potentially provide useful information about the overall impact of causes of death in different populations. PMID- 15655071 TI - GATA: a graphic alignment tool for comparative sequence analysis. AB - BACKGROUND: Several problems exist with current methods used to align DNA sequences for comparative sequence analysis. Most dynamic programming algorithms assume that conserved sequence elements are collinear. This assumption appears valid when comparing orthologous protein coding sequences. Functional constraints on proteins provide strong selective pressure against sequence inversions, and minimize sequence duplications and feature shuffling. For non-coding sequences this collinearity assumption is often invalid. For example, enhancers contain clusters of transcription factor binding sites that change in number, orientation, and spacing during evolution yet the enhancer retains its activity. Dot plot analysis is often used to estimate non-coding sequence relatedness. Yet dot plots do not actually align sequences and thus cannot account well for base insertions or deletions. Moreover, they lack an adequate statistical framework for comparing sequence relatedness and are limited to pairwise comparisons. Lastly, dot plots and dynamic programming text outputs fail to provide an intuitive means for visualizing DNA alignments. RESULTS: To address some of these issues, we created a stand alone, platform independent, graphic alignment tool for comparative sequence analysis (GATA http://gata.sourceforge.net/). GATA uses the NCBI-BLASTN program and extensive post-processing to identify all small sub alignments above a low cut-off score. These are graphed as two shaded boxes, one for each sequence, connected by a line using the coordinate system of their parent sequence. Shading and colour are used to indicate score and orientation. A variety of options exist for querying, modifying and retrieving conserved sequence elements. Extensive gene annotation can be added to both sequences using a standardized General Feature Format (GFF) file. CONCLUSIONS: GATA uses the NCBI BLASTN program in conjunction with post-processing to exhaustively align two DNA sequences. It provides researchers with a fine-grained alignment and visualization tool aptly suited for non-coding, 0-200 kb, pairwise, sequence analysis. It functions independent of sequence feature ordering or orientation, and readily visualizes both large and small sequence inversions, duplications, and segment shuffling. Since the alignment is visual and does not contain gaps, gene annotation can be added to both sequences to create a thoroughly descriptive picture of DNA conservation that is well suited for comparative sequence analysis. PMID- 15655069 TI - Mutagenicity testing with transgenic mice. Part I: Comparison with the mouse bone marrow micronucleus test. AB - As part of a larger literature study on transgenic animals in mutagenicity testing, test results from the transgenic mutagenicity assays (lacI model; commercially available as the Big Blue(R) mouse, and the lacZ model; commercially available as the Mutatrade markMouse), were compared with the results on the same substances in the more traditional mouse bone marrow micronucleus test. 39 substances were found which had been tested in the micronucleus assay and in the above transgenic mouse systems. Although, the transgenic animal mutation assay is not directly comparable with the micronucleus test, because different genetic endpoints are examined: chromosome aberration versus gene mutation, the results for the majority of substances were in agreement. Both test systems, the transgenic mouse assay and the mouse bone marrow micronucleus test, have advantages and they complement each other. However, the transgenic animal assay has some distinct advantages over the micronucleus test: it is not restricted to one target organ and detects systemic as well as local mutagenic effects. PMID- 15655072 TI - Prognostic significance of metallothionein expression in renal cell carcinoma. AB - BACKGROUND: Metallothionein (MT) protein expression deficiency has been implicated in carcinogenesis while MT over expression in tumors is indicative of tumor resistance to anti-cancer treatment. The purpose of the study was to examine the expression of MT expression in human renal cell carcinoma (RCC) and to correlate MT positivity, the pattern and extent of MT expression with tumor histologic cell type and nuclear grade, pathologic stage and patients' survival. PATIENTS AND METHODS: The immunohistochemical expression of MT was determined in 43 formalin-fixed and paraffin-embedded RCC specimens, using a mouse monoclonal antibody that reacts with both human MT-I and MT-II. Correlation was sought between immunohistochemical (MT positivity, intensity and extension of staining) and clinico-pathological data (histological cell type, tumor nuclear grade, pathologic stage and patients' survival). RESULTS: Positive MT staining was present in 21 cases (49%), being mild/moderate and intense in 8 and 13 cases, respectively. The pattern was cytoplasmic in 7 cases and was both cytoplasmic and nuclear in 14 cases. MT expression in a percentage of up to 25% of tumor cells (negative MT staining included) was observed in 31 cases, in a percentage 25-50% of tumor cells in 7 cases, and in a percentage of 50-75% of tumor cells in 5 cases. There was no significant correlation of MT intensity of staining to histological type, stage and patients' survival, while it was inversely correlated to higher tumor nuclear grade. MT extent of staining did not correlate with histological type, nuclear grade, and pathologic stage while a statistically significant association was found with patients' survival. CONCLUSIONS: The inverse correlation between MT staining intensity and tumor nuclear grade in RCC suggests a role of MT in tumor differentiation process. Since extent of MT expression is inversely correlated with survival it may be possibly used as a clinical prognostic parameter. PMID- 15655073 TI - Characterization and performance of a toluene-degrading biofilm developed on pumice stones. AB - BACKGROUND: Hydrocarbon-degrading biofilms in the treatment of contaminated groundwaters have received increasing attention due to the role played in the so called "biobarriers". These are bioremediation systems in which a microbial consortium adherent to a solid support is placed across the flow of a contaminated plume, thus promoting biodegradation of the pollutant. RESULTS: A microbial consortium adherent to pumice granules (biofilm) developed from a toluene-enriched microflora in a mini-scale system, following continuous supply of a mineral medium containing toluene, over a 12-month period. Observation by scanning electron microscopy, together with quantification of the biomass attached to pumice, evidenced the presence of abundant exopolymeric material surrounding the cells in the biofilm. Toluene removal monitored during 12-month operation, reached 99%. Identification of the species, based on comparative 16S ribosomal DNA (rDNA) sequence analysis, revealed that Rhodococcus erythropolis and Pseudomonas marginalis were the predominant bacterial species in the microbial consortium. CONCLUSION: A structurally complex toluene-degrading biofilm, mainly formed by Rhodococcus erythropolis and Pseudomonas marginalis, developed on pumice granules, in a mini-scale apparatus continuously fed with toluene. PMID- 15655074 TI - Effect of method of administration on longitudinal assessment of quality of life in gynecologic cancer: an exploratory study. AB - BACKGROUND: Longitudinal assessments of quality of life are needed to measure changes over the course of a disease and treatment. Computer versions of quality of life instruments have increased the feasibility of obtaining longitudinal measurements. However, there remain occasions when patients are not able to complete these questionnaires. This study examined whether changes measured using a computer version of the Functional Assessment of Cancer Therapy - General (FACT G) on two occasions would be obtained if patients completed a paper version on one of the two occasions. METHODS: Gynecologic oncology patients completed a computer version of the FACT-G pre-operatively and at six months. Patients were given the option of using the paper version instead of the computer at either time point. Repeated measures analysis of variance was used. RESULTS: One hundred nineteen patients completed the FACT-G at both time points. Seventy-one (60%) patients used the computer at both visits, 26 (21.8%) used the computer followed by the paper version, 17 (14.3%) used the paper version followed by the computer version, and five patients (4.2%) used the paper version at both visits. Significant effects over time were obtained in the physical, functional, and emotional well-being domains, and in total scores, but there were no effects of method of administration of the questionnaires and no interaction between method of administration and changes over time. CONCLUSIONS: These data indicate that women are responding to the content of the questionnaire and not method of data collection. Although using the same method of administration of instruments over time is desirable, using alternate methods is preferable to forgoing data collection entirely. Large scale studies should be conducted to determine if the multiple methods of data collection that are becoming increasingly available are producing interchangeable information. PMID- 15655075 TI - Clinical and echocardiographic features of aorto-atrial fistulas. AB - Aorto-atrial fistulas (AAF) are rare but important pathophysiologic conditions of the aorta and have varied presentations such as acute pulmonary edema, chronic heart failure and incidental detection of the fistula. A variety of mechanisms such as aortic dissection, endocarditis with pseudoaneurysm formation, post surgical scenarios or trauma may precipitate the fistula formation. With increasing survival of patients, particularly following complex aortic reconstructive surgeries and redo valve surgeries, recognition of this complication, its clinical features and echocardiographic diagnosis is important. Since physical exam in this condition may be misleading, echocardiography serves as the cornerstone for diagnosis. The case below illustrates aorto-left atrial fistula formation following redo aortic valve surgery with slowly progressive symptoms of heart failure. A brief review of the existing literature of this entity is presented including emphasis on echocardiographic diagnosis and treatment. PMID- 15655076 TI - Predominant constitutive CFTR conductance in small airways. AB - BACKGROUND: The pathological hallmarks of chronic obstructive pulmonary disease (COPD) are inflammation of the small airways (bronchiolitis) and destruction of lung parenchyma (emphysema). These forms of disease arise from chronic prolonged infections, which are usually never present in the normal lung. Despite the fact that primary hygiene and defense of the airways presumably requires a well controlled fluid environment on the surface of the bronchiolar airway, very little is known of the fluid and electrolyte transport properties of airways of less than a few mm diameter. METHODS: We introduce a novel approach to examine some of these properties in a preparation of minimally traumatized porcine bronchioles of about 1 mm diameter by microperfusing the intact bronchiole. RESULTS: In bilateral isotonic NaCl Ringer solutions, the spontaneous transepithelial potential (TEP; lumen to bath) of the bronchiole was small (mean +/- sem: -3 +/- 1 mV; n = 25), but when gluconate replaced luminal Cl-, the bionic Cl- diffusion potentials (-58 +/- 3 mV; n = 25) were as large as -90 mV. TEP diffusion potentials from 2:1 NaCl dilution showed that epithelial Cl- permeability was at least 5 times greater than Na+ permeability. The anion selectivity sequence was similar to that of CFTR. The bionic TEP became more electronegative with stimulation by luminal forskolin (5 microM)+IBMX (100 microM), ATP (100 microM), or adenosine (100 microM), but not by ionomycin. The TEP was partially inhibited by NPPB (100 microM), GlyH-101* (5-50 microM), and CFTRInh-172* (5 microM). RT-PCR gave identifying products for CFTR, alpha-, beta , and gamma-ENaC and NKCC1. Antibodies to CFTR localized specifically to the epithelial cells lining the lumen of the small airways. CONCLUSION: These results indicate that the small airway of the pig is characterized by a constitutively active Cl- conductance that is most likely due to CFTR. PMID- 15655078 TI - Web GIS in practice II: interactive SVG maps of diagnoses of sexually transmitted diseases by Primary Care Trust in London, 1997 - 2003. AB - BACKGROUND: The rates of Sexually transmitted diseases (STDs) in England have been rising steadily since the mid 1990s, making them a major public health concern. In 2003, 672,718 people were diagnosed with an STD in England, and around one third of those cases were diagnosed in London. RESULTS: Using GeoReveal v1.1 for Windows, we produced Web-based interactive choropleth maps of diagnoses of STDs by Primary Care Trust (PCT) in London for the years from 1997 to 2003 http://healthcybermap.org/PCT/STDs/. These maps are in Scalable Vector Graphics (SVG) format and require a freely available Adobe SVG browser plug-in to be displayed. They are based on data obtained from the House of Commons Hansard Written Answers for 15 October 2004. They show steadily rising rates of STDs in London over the covered seven-year period. Also, one can clearly see on the maps that PCTs located in central London had the highest numbers of STD diagnoses throughout the mapped seven years. A companion bar chart allows users to instantly compare the STD figure of a given PCT for a given year against the average figure for all 25 mapped PCTs for the same year, and also compare those figures across all seven years. The maps offer users a rich set of useful features and functions, including the ability to change the classification method in use, the number of ranges in the map, and the colour theme, among others. CONCLUSIONS: Wizard-driven tools like GeoReveal have made it very easy to transform complex raw data into valuable decision support information products (interactive Web maps) in very little time and without requiring much expertise. The resultant interactive maps have the potential of further supporting health planners and decision makers in their planning and management tasks by allowing them to graphically interrogate data, instantly spot trends, and make quick and effective visual comparisons of geographically differentiated phenomena between different geographical areas and over time.SVG makes an ideal format for such maps. SVG is a World Wide Web Consortium non-proprietary, XML-based vector graphics format, and is an extremely powerful alternative to Macromedia(R) Flash and bitmap graphics. PMID- 15655077 TI - Cytogenetic abnormalities and fragile-X syndrome in Autism Spectrum Disorder. AB - BACKGROUND: Autism is a behavioral disorder with impaired social interaction, communication, and repetitive and stereotypic behaviors. About 5-10 % of individuals with autism have 'secondary' autism in which an environmental agent, chromosome abnormality, or single gene disorder can be identified. Ninety percent have idiopathic autism and a major gene has not yet been identified. We have assessed the incidence of chromosome abnormalities and Fragile X syndrome in a population of autistic patients referred to our laboratory. METHODS: Data was analyzed from 433 patients with autistic traits tested using chromosome analysis and/or fluorescence in situ hybridization (FISH) and/or molecular testing for fragile X syndrome by Southern and PCR methods. RESULTS: The median age was 4 years. Sex ratio was 4.5 males to 1 female [354:79]. A chromosome (cs) abnormality was found in 14/421 [3.33 %] cases. The aberrations were: 4/14 [28%] supernumerary markers; 4/14 [28%] deletions; 1/14 [7%] duplication; 3/14 [21%] inversions; 2/14 [14%] translocations. FISH was performed on 23 cases for reasons other than to characterize a previously identified cytogenetic abnormality. All 23 cases were negative. Fragile-X testing by Southern blots and PCR analysis found 7/316 [2.2 %] with an abnormal result. The mutations detected were: a full mutation (fM) and abnormal methylation in 3 [43 %], mosaic mutations with partial methylation of variable clinical significance in 3 [43%] and a permutation carrier [14%]. The frequency of chromosome and fragile-X abnormalities appears to be within the range in reported surveys (cs 4.8-1.7%, FRAX 2-4%). Limitations of our retrospective study include paucity of behavioral diagnostic information, and a specific clinical criterion for testing. CONCLUSIONS: Twenty-eight percent of chromosome abnormalities detected in our study were subtle; therefore a high resolution cytogenetic study with a scrutiny of 15q11.2q13, 2q37 and Xp23.3 region should be standard practice when the indication is autism. The higher incidence of mosaic fragile-X mutations with partial methylation compared to FRAXA positive population [50% vs 15-40%] suggests that faint bands and variations in the Southern band pattern may occur in autistic patients. PMID- 15655080 TI - Scientific priorities in North Korea. PMID- 15655079 TI - Expression profile of immune response genes in patients with Severe Acute Respiratory Syndrome. AB - BACKGROUND: Severe acute respiratory syndrome (SARS) emerged in later February 2003, as a new epidemic form of life-threatening infection caused by a novel coronavirus. However, the immune-pathogenesis of SARS is poorly understood. To understand the host response to this pathogen, we investigated the gene expression profiles of peripheral blood mononuclear cells (PBMCs) derived from SARS patients, and compared with healthy controls. RESULTS: The number of differentially expressed genes was found to be 186 under stringent filtering criteria of microarray data analysis. Several genes were highly up-regulated in patients with SARS, such as, the genes coding for Lactoferrin, S100A9 and Lipocalin 2. The real-time PCR method verified the results of the gene array analysis and showed that those genes that were up-regulated as determined by microarray analysis were also found to be comparatively up-regulated by real-time PCR analysis. CONCLUSIONS: This differential gene expression profiling of PBMCs from patients with SARS strongly suggests that the response of SARS affected patients seems to be mainly an innate inflammatory response, rather than a specific immune response against a viral infection, as we observed a complete lack of cytokine genes usually triggered during a viral infection. Our study shows for the first time how the immune system responds to the SARS infection, and opens new possibilities for designing new diagnostics and treatments for this new life-threatening disease. PMID- 15655081 TI - Inflammation and life-span. PMID- 15655082 TI - Why research in family medicine? A superfluous question. AB - The ultimate answer to the question, "Why research in family medicine?" is to provide better care for our patients. Through research we want to improve quality of primary care by improving our understanding and practice of it. This research will inevitably be specific for family medicine as family medicine is a specific discipline. In this article we first explore what makes family medicine a specific discipline. In a second part we present a framework to grasp the various research questions that must be answered to achieve the complex and multifaceted goal of improving quality of care. Family medicine is a specific discipline for 3 reasons: it has a unique epidemiology, the context of care is important, and it has a strong link and responsibility to the community. Quality of care is a complex and multidimensional concept that raises diverse research questions. We propose to map these questions within a framework defined by the 3 dimensions of the Donabedian triangle--structure, process, and outcome-and within each of these dimensions by 5 foci-basic knowledge, diagnostic and therapeutic problem solving, practice implementation, policy context, and education. This framework may help to make the various research questions operational and to point out the gaps in our research. The questions and answers should be relevant to daily practice and comprise all domains of family medicine so that eventually most of our daily actions in practice will be underpinned with medical, contextual, and policy evidence and contribute to the improvement of the quality of care. PMID- 15655084 TI - The research domain of family medicine. AB - This article characterizes the large research domain of family medicine. It is a domain that can be productively explored from different perspectives, including: (1) the ecology of medical care and its focus on the environments of health care and interactions among them; (2) the realm of causation and important opportunities to discover how people lose and regain their health; (3) knowing medicine in different ways, focusing on what things mean in the inner and outer realities of individuals and groups of individuals; (4) the nature of the work of family physicians, such as first-contact care for any type of problem, sticking with patients regardless of their diagnoses, incorporating context into decision making, development of relevant technologies, articulating useful theory, and measuring what happens in family medicine; (5) the standard research categories of basic, clinical, health services, health policy, and educational research; and (6) thinking of family medicine research as both a linear process of translation and a wheel of knowledge with iterative loops of discovery that come from within family medicine. The domain of family medicine research is important and ripe for fuller discovery, and it invites the thinking and imagination of the best investigators. It seems unlikely that medical research can ever be complete without a robust family medicine research enterprise. As the domain of family medicine research is explored, not a few, but billions of people will benefit. PMID- 15655083 TI - Research in family/general practice is essential for improving health globally. PMID- 15655085 TI - Changing research culture. AB - Although there is general agreement that family medicine has a lot to offer to the health care system, the academic dimension is still not widely understood. There are two main reasons why family medicine needs to develop its scientific potential: to address the true nature of the discipline, and to help in its recognition. The academic establishment benefits from academic family medicine by gaining new questions that are necessary to be answered and by gaining new research approaches.Many problems are encountered when introducing family medicine into the academic arena. Two main strategies for developing family medicine research can be identified. The first is to adapt to the existing structure of the academic world by claiming equal rights with the developed disciplines, collaborating with other university departments in their research projects, publishing articles in established journals, and participating in established faculty development programs. The other, more demanding, strategy is to introduce changes to the academic arena by developing specific research questions, by collaborating on research within family medicine, and by developing family medicine's own success criteria for academic excellence. The two approaches are not mutually exclusive. The World Organization of Family Doctors plays an important role in supporting both approaches through its international affiliations and contacts with policy makers. PMID- 15655086 TI - Building family/general practice research capacity. AB - To promote the range of interventions for building family/general practice (family medicine) research capacity, we describe successful international examples. Such examples of interventions that build research capacity focus on diseases and illness research, as well as process research; monitor the output of research in family/general practice (family medicine); increase the number of family medicine research journals; encourage and enable research skills acquisition (including making it part of professional training); strengthen the academic base; and promote research networks and collaborations. The responsibility for these interventions lies with the government, colleges and academies, and universities. There are exciting and varied methods of building research capacity in family medicine. PMID- 15655087 TI - How the philosophies, styles, and methods of family medicine affect the research agenda. AB - Family physicians provide person-centered, continuous, comprehensive care that is accessible and available at the time of need. Although this core philosophy is shared around the world, its translation into actual practice can vary greatly with time and from place to place as family physicians adapt to local constraints and conditions. Factors driving these local variations include entrenched habits and patterns of care, funding systems, patient expectations, public policy, and the availability and quality of other critical health system components. This diversity provides both an opportunity and a challenge for family medicine research. The potential for fruitful comparisons and contrasts arising from natural experiments may require investigators to use multiple research methods capable of evaluating complex interventions and comparisons. Family medicine has the capacity to be an excellent laboratory in which research in representative populations can offer the pragmatic answers needed by practicing physicians. The nature of the research questions and interventions require the involvement of clinicians in the formulation of research questions and evaluation of the applicability of research results. The variations in implementation of the family medicine philosophy can be a potential asset because of the research opportunities they provide. PMID- 15655088 TI - Family medicine research: implications for Wonca. AB - The purpose of the article is to illustrate where in the lifelong curriculum of family medicine ought students, trainees, and family physicians learn about research in family medicine. The role of Wonca to promote this endeavor is discussed. I describe the recruitment and training programs for family physicians and state the implication for Wonca to promote family medicine research in the undergraduate curriculum, during residency, and in the graduate study programs for experienced medical doctors. I describe the possible research fields in family medicine and the special methods that can be used to get reliable and valid answers to relevant questions in family medicine. It is time for Wonca to develop a policy for family medicine research by developing a strategy to promote it. This can be done through a Wonca policy statement about family medicine research in the medical curriculum. Wonca can, in collaboration with national colleges, arrange workshops and master classes in research; create scholarships and fellowships for international family medicine researchers; influence journals and international bodies, such as the World Health Organization, governments, and colleges about the necessity of good family medical research; describe ethical guidelines for international primary care research; and create a number of databases for researchers to use. PMID- 15655090 TI - The 21st century: the age of family medicine research? AB - Family medicine has matured as an academic and scientific discipline with its own core concepts, knowledge, skills, and research domains. It has acquired much expertise in studying common illnesses; the integration of medical, psychological, social, and behavioral sciences; patient-centered care; and health services delivery. Many health care challenges in the 21st century will place a great demand on primary care, which can serve its purpose only if it is of high quality and evidence based. Family medicine research can contribute to many areas of primary care, ranging from the early diagnosis to equitable health care. Stakeholders, such as the World Health Organization, governments, and funding agencies, are becoming more supportive to family medicine research because they recognize its key importance in improving the quality of primary care and bridging the gap between biomedical research and clinical practice. Family medicine can play a leading role in shifting the paradigm of medical research from the laboratory to the person. The 21st century should be a golden age of family medicine research because the time is right for the discipline, the health care environment is most suitable, and stakeholders are supportive. Family medicine must prepare for it by building up its research track record and capacity. PMID- 15655089 TI - Improving health care globally: a critical review of the necessity of family medicine research and recommendations to build research capacity. AB - An invitational conference led by the World Organization of Family Doctors (Wonca) involving selected delegates from 34 countries was held in Kingston, Ontario, Canada, March 8 to 12, 2003. The conference theme was "Improving Health Globally: The Necessity of Family Medicine Research." Guiding conference discussions was the value that to improve health care worldwide, strong, evidence based primary care is indispensable. Eight papers reviewed before the meeting formed the basic material from which the conference developed 9 recommendations. Wonca, as an international body of family medicine, was regarded as particularly suited to pursue these conference recommendations: 1. Research achievements in family medicine should be displayed to policy makers, health (insurance) authorities, and academic leaders in a systematic way. 2. In all countries, sentinel practice systems should be developed to provide surveillance reports on illness and diseases that have the greatest impact on the population's health and wellness in the community. 3. A clearinghouse should be organized to provide a central repository of knowledge about family medicine research expertise, training, and mentoring.4. National research institutes and university departments of family medicine with a research mission should be developed. 5. Practice-based research networks should be developed around the world.6. Family medicine research journals, conferences, and Web sites should be strengthened to disseminate research findings internationally, and their use coordinated. Improved representation of family medicine research journals in databases, such as Index Medicus, should be pursued.7. Funding of international collaborative research in family medicine should be facilitated.8. International ethical guidelines, with an international ethical review process, should be developed in particular for participatory (action) research, where researchers work in partnership with communities. 9. When implementing these recommendations, the specific needs and implications for developing countries should be addressed.The Wonca executive committee has reviewed these recommendations and the supporting rationale for each. They plan to follow the recommendations, but to do so will require the support and cooperation of many individuals, organizations, and national governments around the world. PMID- 15655091 TI - Research in family medicine in developing countries. AB - The output of family medicine research in developing countries varies vastly from country to country and also within countries. Most research originates from academic departments of family medicine or from collaborative initiatives with researchers in developed countries. There is generally a paucity of researchers, resources, and expertise. Many factors affecting the research output in primary care internationally applies in developing countries, but there are also factors that are unique to these countries. Solutions require financial resources and international goodwill. Wonca could play a major role in enhancing research in family medicine in the developing world. PMID- 15655092 TI - Future of research in family medicine: where to from here? AB - Desired research outcomes in family medicine vary according to the developmental stage of the discipline and the context of practice. Several milestones in the evolution of family practice research worldwide have been achieved. Now family medicine researchers face the challenge of discovering how evidence-based primary health care can be delivered in a sustainable way to individuals within communities. To advance family medicine research, we must ensure that trainees have a positive research attitude, develop academic clinician-researchers, lobby for primary care research funding, support practitioners who wish to do research in their own practices, sustain practice-based research networks, and study important questions. PMID- 15655093 TI - Predicting drunk driving: contribution of alcohol use and related problems, traffic behaviour, personality and platelet monoamine oxidase (MAO) activity. AB - AIMS: The aim of the study was to characterize the predictive value of socio economic data, alcohol consumption measures, smoking, platelet monoamine oxidase (MAO) activity, traffic behaviour habits and impulsivity measures for actual drunk driving. METHODS: Data were collected from 203 male drunk driving offenders and 211 control subjects using self-reported questionnaires, and blood samples were obtained from the two groups. RESULTS: We identified the combination of variables, which predicted correctly, approximately 80% of the subjects' belonging to the drunk driving and control groups. Significant independent discriminators in the final model were, among the health-behaviour measures, alcohol-related problems, frequency of using alcohol, the amount of alcohol consumed and smoking. Predictive traffic behaviour measures were seat belt use and paying for parking. Among the impulsivity measures, dysfunctional impulsivity was the best predictor; platelet MAO activity and age also had an independent predictive value. CONCLUSIONS: Our results support the notion that drunk driving is the result of a combination of various behavioural, biological and personality related risk factors. PMID- 15655094 TI - Lower-limb deep-vein thrombosis in a general hospital: risk factors, outcomes and the contribution of intravenous drug use. AB - BACKGROUND: Much of the morbidity associated with deep-vein thrombosis (DVT) is absent from the evidence base used to define best clinical practice. Intravenous (IV) drug use is an increasingly frequent cause of DVT. AIM: To obtain a profile of DVT patients in routine clinical care, and determine the contribution of IV drug use. DESIGN: Retrospective case-note review. METHODS: We reviewed 232 episodes of lower-limb DVT in a large district general hospital during 1996. RESULTS: Patients had mean (range) age 62.8 (21-97) years, with 43.9% aged > 70 years. A large proportion would have been excluded from prospective studies that have contributed to current DVT guidelines. Risk factors included smoking (33.0%), immobility (26.5%), previous DVT (23.6%), surgery in the last 3 months (18.2%), malignancy (16.5%), varicose veins (10.5%) and IV drug use (6.9%). Forty five (19.4%) had multiple risk factors. Postsurgical DVT commonly presented from the community following initial hospital discharge. Intravenous drug use accounted for 48.4% of episodes in patients aged < or =40 years. Thrombosis was right-sided in 68.8% of IV drug users, compared to 38.2% in others (p = 0.034). DISCUSSION: Our patients differed from those in most of the prospective studies used to develop routine clinical care pathways for DVT. Intravenous drug use is an important cause of community-acquired DVT in young adults. PMID- 15655095 TI - Factors influencing medical treatment of heart failure patients in Spanish internal medicine departments: a national survey. AB - BACKGROUND: The medical management of heart failure (HF) in clinical practice varies considerably by country and by medical specialty. AIM: To assess the treatment of HF patients admitted to Internal Medicine departments, and to evaluate out-patient management prior to admission, by specialty. DESIGN: Prospective cross-sectional multi-centre survey. METHODS: Of 55 randomly selected Spanish hospitals, 51 agreed to participate. All patients (n = 2145) consecutively admitted for decompensated HF to the Departments of Internal Medicine of these hospitals, over 5 months, were included. Twenty variables were analysed, including aspects relating to out-patient management prior to admission. RESULTS: Mean +/- SD age was 77.2 +/- 10.5 years, 57.3% were female, 47% had systolic dysfunction. Prescriptions at discharge: loop diuretics 85.6%, spironolactone 29.8%, ACEIs 65.8%, beta-blockers 8.7%, cardiac glycosides 39%. At admission, 86% already had a diagnosis of HF. Of these, 53% (older patients and more women) were being treated on an out-patient basis by primary care physicians. Primary care physicians requested fewer echocardiograms than internists (38% vs. 69%, p<0.001) and prescribed fewer drugs (ACEIs 40% vs. 54%, p<0.001; spironolactone 15% vs. 23%, p<0.05; beta-blockers 6% vs. 13%, p<0.01). The internists treated more incapacitated patients than the cardiologists (p<0.001), prescribed more high-dose ACEIs (20% vs. 13%, p<0.01) and spironolactone (26% vs. 20%, p<0.05), and fewer anticoagulants (32% vs. 39%, p<0.05). DISCUSSION: Patients admitted to medical departments with HF are different to those found in clinical trials. Their management is currently suboptimal. Differences in treatment between internists and cardiologists appear to be accounted for by differences in the patients they treat. PMID- 15655096 TI - Poor prognosis in proteinuric type 2 diabetic patients with retinopathy: insights from the RENAAL study. AB - BACKGROUND: Retinopathy is the clinical hallmark of generalized microangiopathy in diabetes. AIM: To examine the relation of this abnormality to end-stage renal disease (ESRD) and death in type 2 diabetes. DESIGN: Retrospective analysis. METHODS: Of 1513 type 2 diabetic patients with nephropathy participating in the Reduction of Endpoints in NIDDM with the Angiotensin II Antagonist Losartan (RENAAL) study, 1456 (96.5%) were assessed at baseline by ophthalmoscopy or fundus photography. RENAAL was a multinational double masked, randomized, placebo controlled intervention study, whose primary end-point was the composite of a doubling of the baseline serum creatinine concentration, end-stage renal disease (ESRD) or death. RESULTS: Of those assessed at baseline, 65% had diabetic retinopathy. Patients with retinopathy had higher systolic blood pressure, albuminuria and lower glomerular filtration rate (GFR), haemoglobin and serum albumin values than those without. In univariate analyses, the presence of retinopathy was associated with a 44% increase in the primary composite end-point (hazard ratio 1.44, 95%CI 1.22-1.70, p < 0.001). Patients with retinopathy had a 52% increase in doubling of serum creatinine (p < 0.001), a 47% increased risk of ESRD (p = 0.002) and a 33% increase in risk of death (p = 0.026) compared to those without. In multivariate analyses, the presence of retinopathy was associated with a 23% increase (p = 0.015) in the primary composite end-point and a 22% increase in ESRD or death (p = 0.038). DISCUSSION: The presence of diabetic retinopathy at baseline is associated with more proteinuria, lower GFR, and a higher risk for ESRD and death in type 2 diabetic patients. PMID- 15655097 TI - Assessment of a point-of-care test for paracetamol and salicylate in blood. AB - BACKGROUND: Paracetamol and salicylate are commonly taken in acute overdose. Clinicians have a low threshold for excluding the presence of these two drugs, by ordering laboratory tests in any patient suspected of ingesting an overdose or with an altered mental state. AIM: To test the effectiveness of a new point of care test that qualitatively detects paracetamol and salicylate in blood and to examine the potential time saved by its use. DESIGN: Prospective multicentre trial. METHODS: The new test was compared with laboratory analysis in a routine blood sample taken from patients presenting to emergency departments with suspected overdose. RESULTS: The test had sensitivities of 98.5% and 88.5%, and specificities of 74.7% and 92%, for paracetamol and salicylate, respectively, at cut-off levels of 25 mg/l and 100 mg/l, respectively The point of care test results were available 2 h before the laboratory result. DISCUSSION: This point of-care test could be used to rule out an overdose with either of these two drugs, and could thus lead to earlier clinical decisions for suspected overdose patients. Recommendations have been made following this trial that the cut-off value for paracetamol should be reduced from 25 mg/l to 12.5 mg/l in order to increase its usefulness. To prevent the test being misread, we also suggest that each device should be embossed to remind users that the presence of a line indicates there is no drug present. PMID- 15655099 TI - Heparin use in acute ischaemic stroke: does evidence change practice? PMID- 15655098 TI - Primary systemic vasculitis: clinical features and mortality. AB - BACKGROUND: Wegener's granulomatosis (WG), Churg Strauss syndrome (CSS) and microscopic polyangiitis (MPA) are primary systemic vasculitides (PSV), the clinical features of which have been described from tertiary centres. AIM: To provide the first clinical description of MPA from a general hospital and compare clinical features with WG and CSS. DESIGN: Retrospective analysis of patient records. METHODS: Records of 99 PSV patients attending a single hospital, from 1988 to 2000, were reviewed for: clinical features, date/age at diagnosis, sex, duration of illness, anti-neutrophil cytoplasmic antibodies (ANCA), treatment, comorbidity and deaths. Cases were classified using ACR, CHCC and Lanham criteria/definitions. Birmingham vasculitis activity scores (BVAS) and damage index (VDI) were calculated. Survival was assessed using Cox proportional hazards model and standardized mortality ratios (SMRs). RESULTS: Compared to previous reports there was more ENT (29%) and respiratory (29%) but less renal (92%) involvement in MPA, and less ENT involvement in WG (81%). CSS showed high neurological (72%), cardiovascular (28%) and gastrointestinal (17%) involvement and the highest median (range) VDI (p = 0.01 vs. WG; p = 0.001 vs. MPA). BVAS1 was significantly lower in MPA than in WG [median (range) 15 (4-29) vs. 21 (6 39), (p = 0.001)] but not in CSS [20 (7-28), p = 0.08]. SMR (95%CI) for PSV was 4.8 (3.0-6.6); 5-year survival was 45.1% for MPA, 75.9% for WG and 68.1% for CSS. Age was a significant risk, but only to the same extent as in the reference population. When age was adjusted for, no other significant factor was found. DISCUSSION: The clinical characteristics seen here are similar to those in previous series. There are difficulties in using the MPA CHCC definitions in classification. There is a high proportion of neurological involvement in CSS, causing permanent damage. MPA may have a poorer prognosis than WG or CSS. PMID- 15655100 TI - Standards for gene therapy clinical trials based on pro-active risk assessment in a London NHS Teaching Hospital Trust. AB - Conducting gene therapy clinical trials with genetically modified organisms as the vectors presents unique safety and infection control issues. The area is governed by a range of legislation and guidelines, some unique to this field, as well as those pertinent to any area of clinical work. The relevant regulations covering gene therapy using genetically modified vectors are reviewed and illustrated with the approach taken by a large teaching hospital NHS Trust. Key elements were Trust-wide communication and involvement of staff in a pro-active approach to risk management, with specific emphasis on staff training and engagement, waste management, audit and record keeping. This process has led to the development of proposed standards for clinical trials involving genetically modified micro-organisms. PMID- 15655101 TI - Nurse-led adherence support in hypertension: a randomized controlled trial. AB - BACKGROUND: Lack of medication adherence is a common reason for poor control of blood pressure in the community, increasing the risk of heart attacks and strokes. OBJECTIVE: To evaluate the effect of nurse-led adherence support for people with uncontrolled high blood pressure compared with usual care. METHODS: We recruited 245 women and men with uncontrolled hypertension (> or = 150/90 mmHg) from 21 general practices in Bristol, UK. Participants were randomized to receive nurse-led adherence support or usual care alone. Main outcome measures were adherence to medication ('timing compliance') and blood pressure. RESULTS: Mean baseline timing compliance (+/- SD) was high in both the intervention (90.8 +/- 15.6%) and the control group (94.5 +/- 7.6%). There was no evidence of an effect of the intervention on timing compliance at follow-up (adjusted difference in means -1.0%; 95% confidence interval (CI) -5.1 to 3.1). There was also no difference at follow-up between the groups with regard to systolic blood pressure (-2.7 mmHg; 95% CI -7.2 to 1.8) or diastolic blood pressure (0.2 mmHg; 95% CI 1.9 to 2.3). Projected costs for the primary care sector per consultation were 6.60 pound sterling for the intervention compared with 5.08 pound sterling for usual care. CONCLUSION: In this study, adherence to blood pressure medication was much higher than previously reported. There was no evidence of an effect of nurse led adherence support on medication adherence or blood pressure compared with usual care. Nurse-led adherence support was also more expensive from a primary care perspective. PMID- 15655102 TI - Can 'anecdote' ever be research? PMID- 15655103 TI - Copying letters to patients--will it happen? PMID- 15655104 TI - Grain growth and endosperm cell size under high night temperatures in rice (Oryza sativa L.). AB - BACKGROUND AND AIMS: High night temperatures are more harmful to grain weight in rice than high day temperatures. Grain growth rate and growth duration were investigated to determine which was the cause of the decrease in final grain weight under high night temperatures. Endosperm cell number and cell sizes were also examined to determine which might cause the decrease in final grain weight. METHODS: Rice plants were grown outdoors in plastic pots and moved at heading time to three temperature-controlled glasshouses under high night temperature (HNT; 22/34 degrees C), high day temperature (HDT; 34/22 degrees C) and control conditions (CONT; 22/22 degrees C). Grains were sampled periodically, and the time-course of grain growth was divided into rate and duration by logistic regression analysis. Endosperm cell numbers and cell sizes were analysed by digitalized hand-tracing images of endosperm cross-sections. KEY RESULTS: The duration of grain growth was reduced by high temperature both day and night. However, the rate of grain growth was lower in HNT than in HDT. The number of cells in endosperm cross-sections in HNT was similar to that in HDT, and higher than that in CONT. The average cell area was smaller in HNT than in either CONT or HDT. The differences in average cell areas between HNT and HDT were greater at distances 60-80 % from the central point of endosperm towards the endosperm surface. CONCLUSIONS: The results show that HNT compared with HDT reduced the final grain weight by a reduction in grain growth rate in the early or middle stages of grain filling, and also reduced cell size midway between the central point and the surface of endosperm. PMID- 15655105 TI - Influence of the gibberellin-sensitive Rht8 dwarfing gene on leaf epidermal cell dimensions and early vigour in wheat (Triticum aestivum L.). AB - BACKGROUND AND AIMS: The gibberellin-insensitive Rht-B1b and Rht-D1b dwarfing genes are known to reduce the size of cells in culms, leaves and coleoptiles of wheat. Resulting leaf area development of gibberellin-insensitive wheats is poor compared to standard height (Rht-B1a and Rht-D1a) genotypes. Alternative dwarfing genes to Rht-B1b and Rht-D1b are available that reduce plant height, such as the gibberellin-responsive Rht8 gene. This study aims to investigate if Rht8 has a similar dwarfing effect on the size of leaf cells to reduce leaf area. METHODS: The effect of Rht8 on cell size and leaf area was assessed in four types of epidermal cells (interstomatal, long, sister and bulliform) measured on leaf 2 of standard height (rht8) and semi-dwarf (Rht8) doubled-haploid lines (DHLs). The DHLs were derived from a cross between very vigorous, standard height (rht8) ('Vigour18') and less vigorous, semi-dwarf (Rht8) ('Chuan-Mai 18') parents. KEY RESULTS: Large differences were observed in seedling vigour between the parents, where 'Vigour18' had a much greater plant leaf area than 'Chuan-Mai 18'. Accordingly, 'Vigour18' had on average longer, wider and more epidermal cells and cell files than 'Chuan-Mai 18'. Although there was correspondingly large genotypic variation among DHLs for these traits, the contrast between semi-dwarf Rht8 and tall rht8 DHLs revealed no difference in the size of leaf 2 or average cell characteristics. Hence, these traits were independent of plant height and therefore Rht8 in the DHLs. Correlations for leaf and average cell size across DHLs revealed a strong and positive relationship between leaf width and cell files, while the relationships between leaf and cell width, and leaf and cell length were not statistically different. The relative contribution of the four cell types (long, sister, interstomatal and bulliform) to leaf size in the parents, comparative controls and DHLs is discussed. CONCLUSIONS: Despite a large range in early vigour among the DHLs, none of the DHLs attained the leaf area or epidermal cell size and numbers of the vigorous rht8 parent. Nonetheless, the potential exists to increase the early vigour of semi-dwarf wheats by using GA sensitive dwarfing genes such as Rht8. PMID- 15655106 TI - Partitioning of (13)C-photosynthate from spur leaves during fruit growth of three Japanese pear (Pyrus pyrifolia) cultivars differing in maturation date. AB - BACKGROUND AND AIMS: In fruit crops, fruit size at harvest is an important aspect of quality. With Japanese pears (Pyrus pyrifolia), later maturing cultivars usually have larger fruits than earlier maturing cultivars. It is considered that the supply of photosynthate during fruit development is a critical determinant of size. To assess the interaction of assimilate supply and early/late maturity of cultivars and its effect on final fruit size, the pattern of carbon assimilate partitioning from spur leaves (source) to fruit and other organs (sinks) during fruit growth was investigated using three genotypes differing in maturation date. METHODS: Partitioning of photosynthate from spur leaves during fruit growth was investigated by exposure of spurs to (13)CO(2) and measurement of the change in (13)C abundance in dry matter with time. Leaf number and leaf area per spur, fresh fruit weight, cell number and cell size of the mesocarp were measured and used to model the development of the spur leaf and fruit. KEY RESULTS: Compared with the earlier-maturing cultivars 'Shinsui' and 'Kousui', the larger-fruited, later-maturing cultivar 'Shinsetsu' had a greater total leaf area per spur, greater source strength (source weight x source specific activity), with more (13)C assimilated per spur and allocated to fruit, smaller loss of (13)C in respiration and export over the season, and longer duration of cell division and enlargement. Histology shows that cultivar differences in final fruit size were mainly attributable to the number of cells in the mesocarp. CONCLUSIONS: Assimilate availability during the period of cell division was crucial for early fruit growth and closely correlated with final fruit size. Early fruit growth of the earlier-maturing cultivars, but not the later-maturing ones, was severely restrained by assimilate supply rather than by sink limitation. PMID- 15655107 TI - Modelling the effect of fruit growth on surface conductance to water vapour diffusion. AB - BACKGROUND AND AIMS: A model of fruit surface conductance to water vapour diffusion driven by fruit growth is proposed. It computes the total fruit conductance by integrating each of its components: stomata, cuticle and cracks. METHODS: The stomatal conductance is computed from the stomatal density per fruit and the specific stomatal conductance. The cuticular component is equal to the proportion of cuticle per fruit multiplied by its specific conductance. Cracks are assumed to be generated when pulp expansion rate exceeds cuticle expansion rate. A constant percentage of cracks is assumed to heal each day. The proportion of cracks to total fruit surface area multiplied by the specific crack conductance accounts for the crack component. The model was applied to peach fruit (Prunus persica) and its parameters were estimated from field experiments with various crop load and irrigation regimes. KEY RESULTS: The predictions were in good agreement with the experimental measurements and for the different conditions (irrigation and crop load). Total fruit surface conductance decreased during early growth as stomatal density, and hence the contribution of the stomatal conductance, decreased from 80 to 20 % with fruit expansion. Cracks were generated for fruits exhibiting high growth rates during late growth and the crack component could account for up to 60 % of the total conductance during the rapid fruit growth. The cuticular contribution was slightly variable (around 20 %). Sensitivity analysis revealed that simulated conductance was highly affected by stomatal parameters during the early period of growth and by both crack and stomatal parameters during the late period. Large fruit growth rate leads to earlier and greater increase of conductance due to higher crack occurrence. Conversely, low fruit growth rate accounts for a delayed and lower increase of conductance. CONCLUSIONS: By predicting crack occurrence during fruit growth, this model could be helpful in managing cropping practices for integrated plant protection. PMID- 15655108 TI - Not just another hole in the wall: understanding intercellular protein trafficking. AB - Development and differentiation of multicellular organisms requires cell-to-cell communication. In plants direct signaling and exchange of macromolecules between cells is possible through plasmodesmata. Recently direct exchange of membrane bound vesicles and organelles has been demonstrated between animal cells through formation of cytoplasmic bridges (tunneling nanotubes) in vitro. Here we review recent developments in cell-to-cell trafficking of macromolecules in plants and animals. PMID- 15655109 TI - The p400 E1A-associated protein is a novel component of the p53 --> p21 senescence pathway. AB - Adenovirus E1A-associated p400 belongs to the SWI2/SNF2 family of chromatin remodeling proteins. Here, we report that p400 is a component of the p53 p21(WAF1/CIP1/sid1) pathway, regulating the p21 transcription and senescence induction program. Acute depletion of p400 expression by shRNA (short hairpin RNA) synthesis led to premature senescence of untransformed human fibroblasts, whose features include G1 arrest, p21 induction, senescence-associated heterochromatic foci (SAHF), and beta-gal staining. Importantly, p400shRNA induced premature senescence phenotypes were rescued by coexpression of p53-shRNA or p21-shRNA. Furthermore, p400 complex colocalized with p53 on the p21 promoter. These data suggest that the p400 complex inhibits p53 --> p21 transcription and the development of premature senescence. PMID- 15655110 TI - Mad2 prevents aneuploidy and premature proteolysis of cyclin B and securin during meiosis I in mouse oocytes. AB - In mitosis, the spindle checkpoint protein Mad2 averts aneuploidy by delaying anaphase onset until chromosomes align. Here we show that depletion of Mad2 in meiosis I mouse oocytes induced an increased incidence of aneuploidy. Proteolysis of cyclin B and securin commenced earlier in Mad2-depleted oocytes, resulting in a shortened duration of meiosis I. Furthermore, overexpression of Mad2 inhibited homolog disjunction. We conclude that Mad2 delays the onset of cyclin B and securin degradation and averts aneuploidy during meiosis I in mammalian oocytes. The data suggest a link between trisomies such as Down syndrome and defective oocyte spindle checkpoint function. PMID- 15655111 TI - Distinct functions of junD in cardiac hypertrophy and heart failure. AB - Cardiac hypertrophic stimuli induce both adaptive and maladaptive growth response pathways in heart. Here we show that mice lacking junD develop less adaptive hypertrophy in heart after mechanical pressure overload, while cardiomyocyte specific expression of junD in mice results in spontaneous ventricular dilation and decreased contractility. In contrast, fra-1 conditional knock-out mice have a normal hypertrophic response, whereas hearts from fra-1 transgenic mice decompensate prematurely. Moreover, fra-1 transgenic mice simultaneously lacking junD reveal a spontaneous dilated cardiomyopathy associated with increased cardiomyocyte apoptosis and a primary mitochondrial defect. These data suggest that junD promotes both adaptive-protective and maladaptive hypertrophy in heart, depending on its expression levels. PMID- 15655112 TI - High incidence of proviral integrations in the Hoxa locus in a new model of E2a PBX1-induced B-cell leukemia. AB - Relevant mouse models of E2a-PBX1-induced pre-B cell leukemia are still elusive. We now report the generation of a pre-B leukemia model using E2a-PBX1 transgenic mice, which lack mature and precursor T-cells as a result of engineered loss of CD3epsilon expression (CD3epsilon(-/-)). Using insertional mutagenesis and inverse-PCR, we show that B-cell leukemia development in the E2a-PBX1 x CD3epsilon(-/-) compound transgenic animals is significantly accelerated when compared to control littermates, and document several known and novel integrations in these tumors. Of all common integration sites, a small region of 19 kb in the Hoxa gene locus, mostly between Hoxa6 and Hoxa10, represented 18% of all integrations in the E2a-PBX1 B-cell leukemia and was targeted in 86% of these leukemias compared to 17% in control tumors. Q-PCR assessment of expression levels for most Hoxa cluster genes in these tumors revealed an unprecedented impact of the proviral integrations on Hoxa gene expression, with tumors having one to seven different Hoxa genes overexpressed at levels up to 6600-fold above control values. Together our studies set the stage for modeling E2a-PBX1-induced B-cell leukemia and shed new light on the complexity pertaining to Hox gene regulation. In addition, our results show that the Hoxa gene cluster is preferentially targeted in E2a-PBX1-induced tumors, thus suggesting functional collaboration between these oncogenes in pre-B-cell tumors. PMID- 15655113 TI - The control of Spo11's interaction with meiotic recombination hotspots. AB - Programmed double-strand breaks (DSBs), which initiate meiotic recombination, arise through the activity of the evolutionary conserved topoisomerase homolog Spo11. Spo11 is believed to catalyze the DNA cleavage reaction in the initial step of DSB formation, while at least a further 11 factors assist in Saccharomyces cerevisiae. Using chromatin-immunoprecipitation (ChIP), we detected the transient, noncovalent association of Spo11 with meiotic hotspots in wild type cells. The establishment of this association requires Rec102, Rec104, and Rec114, while the timely removal of Spo11 from chromatin depends on several factors, including Mei4 and Ndt80. In addition, at least one further component, namely, Red1, is responsible for locally restricting Spo11's interaction to the core region of the hotspot. In chromosome spreads, we observed meiosis-specific Spo11-Myc foci, independent of DSB formation, from leptotene until pachytene. In both rad50S and com1Delta/sae2Delta mutants, we observed a novel reaction intermediate between Spo11 and hotspots, which leads to the detection of full length hotspot DNA by ChIP in the absence of artificial cross-linking. Although this DNA does not contain a break, its recovery requires Spo11's catalytic residue Y135. We propose that detection of uncross-linked full-length hotspot DNA is only possible during the reversible stage of the Spo11 cleavage reaction, in which rad50S and com1Delta/sae2Delta mutants transiently arrest. PMID- 15655114 TI - Olig2 and Ngn2 function in opposition to modulate gene expression in motor neuron progenitor cells. AB - Spinal motor neurons and oligodendrocytes are generated sequentially from a common pool of progenitors termed pMN cells. Olig2 is a bHLH-class transcription factor in pMN cells, but it has remained unclear how its transcriptional activity is modulated to first produce motor neurons and then oligodendrocytes. Previous studies have shown that Olig2 primes pMN cells to become motor neurons by triggering the expression of Ngn2 and Lhx3. Here we show that Olig2 also antagonizes the premature expression of post-mitotic motor neuron genes in pMN cells. This blockade is counteracted by Ngn2, which accumulates heterogeneously in pMN cells, thereby releasing a subset of the progenitors to differentiate and activate expression of post-mitotic motor neuron genes. The antagonistic relationship between Ngn2 and Olig2 is mediated by protein interactions that squelch activity as well as competition for shared DNA-binding sites. Our data support a model in which the Olig2/Ngn2 ratio in progenitor cells serves as a gate for timing proper gene expression during the development of pMN cells: Olig2(high) maintains the pMN state, thereby holding cells in reserve for oligodendrocyte generation, whereas Ngn2(high) favors the conversion of pMN cells into post-mitotic motor neurons. PMID- 15655115 TI - Asymptomatic left ventricular systolic dysfunction in essential hypertension: prevalence, determinants, and prognostic value. AB - Prevalence, determinants, and prognostic value of asymptomatic left ventricular systolic dysfunction (LVSD) in uncomplicated subjects with essential hypertension are still incompletely known. We studied 2384 initially untreated subjects with hypertension, no previous cardiovascular disease, and no symptoms or physical signs of congestive heart failure (CHF). These subjects were studied at entry and followed for up to 17 years (mean 6.0). Asymptomatic LVSD (ALVSD), defined by an echocardiographic ejection fraction <50%, was found in 3.6% of subjects. Cigarette smoking (P=0.013), increased left ventricular (LV) mass (P=0.001), and higher 24-hour heart rate (P=0.014) were independent correlates of ALVSD. During follow-up, a first cardiovascular event occurred in 227 subjects, and 24 of these events were hospitalizations for symptomatic CHF. Incidence of CHF per 100 persons per year was 0.12 in patients without and 1.48 in patients with ALVSD (log-rank test P=0.0001). In a Cox model, after adjustment for age (P=0.0001), LV mass (P=0.0001), and cigarette smoking (P=0.039), LVSD conferred a markedly increased risk for CHF (odds ratio, 9.99; 95% confidence interval, 3.67 to 27.2). Incidence of coronary (0.84 versus 0.62x100 person years) and cerebrovascular (0.80 versus 0.62x100 person years) events did not differ (all P=NS) between subjects with and without ALVSD. ALVSD is a potent and early marker of evolution toward severe CHF requiring hospitalization in subjects with essential hypertension. PMID- 15655116 TI - Mobilizing endothelial progenitor cells. AB - Mobilization of endogenous endothelial progenitor cells (EPCs) from the bone marrow may be an alternative way to increase neovascularization and may be used as therapeutic option for the treatment of ischemic cardiovascular diseases. In this review, we discuss the EPC mobilizing effects of pro-inflammatory cytokines such as granolocyte monocyte colony-stimulating factor and granulocyte colony stimulating factor, growth factors such as vascular endothelial growth factor, placental growth factor, erythropoietin, and angiopoietin-1, chemokines such as stromal cell-derived factor-1, hormones such as estrogens and lipid-lowering and anti-diabetic drugs, as well as physical activity. PMID- 15655117 TI - Subtle renal injury is likely a common mechanism for salt-sensitive essential hypertension. PMID- 15655118 TI - Different involvement of extracellular matrix components in small and large arteries during chronic NO synthase inhibition. AB - In essential hypertension, conduit arteries present hypertrophic remodeling (increased cross-sectional area), whereas small arteries undergo eutrophic remodeling. The involvement of matrix metalloproteinases (MMPs) and de-adhesion proteins, such as tenascin-C and thrombospondin, has been relatively well characterized in large artery remodeling, but their contribution is not known in small artery remodeling. Rats received N(omega)-nitro-L-arginine methyl ester (L NAME; 50 mg/kg per day) in their drinking water on days 1, 3, 7, 14, and 28. Arterial MMP-2 activity was measured by ELISA, whereas levels of tenascin-C and thrombospondin were assessed by Western blotting. To determine the involvement of MMPs, additional L-NAME rats received the nonselective MMP inhibitor doxycycline (30 mg/kg per day) on days 7, 14, and 28. Already, at day 1, pressure was elevated. Media/lumen ratio of mesenteric arteries and the aorta increased gradually to reach significance at 28 days. However, the cross-sectional area increased only in the aorta, confirming the heterogeneous remodeling process. In small arteries, MMP-2 activity increased after 7 and 14 days of treatment and returned to baseline at 28 days, whereas the elevation was more progressive but sustained in the aorta. The level of thrombospondin paralleled that of MMP-2 in small arteries, whereas tenascin-C levels declined rapidly and stayed below control values. Doxycycline blunted large artery remodeling but had no influence on the development of eutrophic remodeling despite elevation of MMP-2 activity in the process. Thus, in contrast to large artery hypertrophic remodeling, in which the contributions of cellular de-adhesion and matrix breakdown is manifest, the contribution of MMPs in eutrophic remodeling appears less crucial. PMID- 15655119 TI - Dietary linolenic acid is associated with a lower prevalence of hypertension in the NHLBI Family Heart Study. AB - Dietary linolenic acid has been shown to be associated with coronary artery disease. However, limited data are available on its effects on blood pressure. We used data from 4594 white participants (aged 25 to 93 years) in the National Heart, Lung, and Blood Institute (NHLBI) Family Heart Study to evaluate whether dietary linolenic acid was associated with prevalent hypertension and resting blood pressure. We used generalized estimating equations to determine the prevalence odds ratios (ORs) of hypertension and adjusted means of systolic and diastolic blood pressure across quartiles of linolenic acid. Mean dietary linolenic acid intake was 0.81+/-0.35 g per day for men and 0.69+/-0.29 g per day for women. From the lowest to the highest quartile of linolenic acid, multivariable adjusted ORs (95% confidence interval [CI]) for hypertension were 1.0 (reference), 0.73 (0.56 to 0.95), 0.71 (0.53 to 0.95), and 0.67 (0.47 to 0.96), respectively (P for trend 0.04), controlling for age, sex, energy intake, body mass index, risk group, study site, education, smoking, alcohol intake, exercise, and history of coronary artery disease and diabetes mellitus. Dietary linolenic acid was related inversely to resting systolic (P for trend 0.03) but not diastolic blood pressure (P for trend 0.43). Linoleic acid, an omega-6 fatty acid, was not associated with prevalent hypertension or blood pressure. These data suggest that dietary linolenic acid is associated with a lower prevalence of hypertension and lower systolic blood pressure in white subjects. PMID- 15655120 TI - Locating a blood pressure quantitative trait locus within 117 kb on the rat genome: substitution mapping and renal expression analysis. AB - Previously, a blood pressure (BP) quantitative trait locus (QTL) on rat chromosome 9 (RNO9) was localized to a <2.4 cM interval using congenic strains generated by introgressing segments of RNO9 from the Dahl salt-resistant (R) rat into the background of the Dahl salt-sensitive (S) rat. Renal gene expression using Affymetrix gene chips was profiled on S and a congenic strain spanning the 2.4-cM BP QTL interval. This analysis identified 20 differentially expressed genes/expressed sequence tags. Of these, the locus with the greatest differential expression (30- to 35-fold) was regulated endocrine-specific protein 18 (Resp18), which also mapped in the 2.4-cM BP QTL interval. Additional substitution mapping located the QTL to <0.4 cM or approximately 493 kb. This newly defined QTL region still included Resp18. Nucleotide variants were identified between S and R genomic DNA of Resp18 in the coding, 5' regulatory and 3' untranslated regions. The coding sequence variation (T/C) occurs in exon 2 and predicts an amino acid change (Ile/Val) in the protein product. Resp18 was considered a differentially expressed positional candidate for the QTL. To fine-map the BP QTL, we constructed a congenic strain with a smaller introgressed region. Compared with the S rat, this strain (1) had significantly lower BP, (2) did not contain the R form of Resp18, and (3) did not retain the rather spectacular differential expression of Resp18. Together, these results demonstrate that a BP QTL independent of Resp18 exists within the newly defined 117-kb QTL region on RNO9. PMID- 15655121 TI - Beyond blood pressure: endothelial protection against hypercholesterolemia by angiotensin II type-1 receptor blockade. PMID- 15655122 TI - Angiotensin II type-1 receptor antagonism attenuates the inflammatory and thrombogenic responses to hypercholesterolemia in venules. AB - Hypercholesterolemia elicits an inflammatory response in the microvasculature that is accompanied by an increased expression of angiotensin II type-1 receptors (AT1-R) on platelets, leukocytes, and endothelial cells. AT1-R blockade attenuates inflammatory responses to angiotensin II (eg, adhesion molecule expression and reactive oxygen species production). We investigated whether AT1-R antagonism attenuates the platelet and leukocyte recruitment induced by acute hypercholesterolemia in postcapillary venules. Leukocyte and platelet adhesion and oxidative stress were quantified by intravital microscopy in cremaster muscle, and P-selectin and AT1-R expression was determined in mice placed on a normal diet (ND) or high-cholesterol diet (HCD) for 2 weeks. Platelet and leukocyte adhesion was significantly elevated by hypercholesterolemia. In HCD mice receiving losartan (HCD-Los) in drinking water, platelet and leukocyte recruitment was reduced to ND levels. Increased platelet adhesion was observed in HCD mice receiving platelets from HCD-Los mice, consistent with a direct beneficial action of losartan on the vessel wall. Hypercholesterolemia elicited an oxidative stress in venules and an increased expression of P-selectin and AT1 R. The oxidative stress and AT1-R upregulation were reduced by losartan, but the P-selectin response was not. We propose that AT1-R engagement contributes to the prothrombogenic and proinflammatory state induced in venules by hypercholesterolemia. PMID- 15655123 TI - Reduction in albuminuria translates to reduction in cardiovascular events in hypertensive patients: losartan intervention for endpoint reduction in hypertension study. AB - Few data are available to clarify whether changes in albuminuria over time translate to changes in cardiovascular risk. The aim of the present study was to examine whether changes in albuminuria during 4.8 years of antihypertensive treatment were related to changes in risk in 8206 patients with hypertension and left ventricular hypertrophy in the Losartan Intervention For Endpoint reduction in hypertension (LIFE) study. Urinary albumin/creatinine ratio (UACR) was measured at baseline and annually. Time-varying albuminuria was closely related to risk for the primary composite end point (ie, when UACR decreased during treatment, risk was reduced accordingly). When the population was divided according to median baseline value (1.21 mg/mmol) and median year 1 UACR (0.67 mg/mmol), risk increased stepwise and significantly for the primary composite end point from those with low baseline/low year 1 (5.5%), to low baseline/high year 1 (8.6%), to high baseline/low year 1 (9.4%), and to high baseline/high year 1 (13.5%) values. Similar significant, stepwise increases in risk were seen for the components of the primary composite end point (cardiovascular mortality, stroke, and myocardial infarction). The observation that changes in UACR during antihypertensive treatment over time translated to changes in risk for cardiovascular morbidity and mortality was not explained by in-treatment level of blood pressure. We propose that monitoring of albuminuria should be an integrated part of the management of hypertension. If albuminuria is not decreased by the patient's current antihypertensive and other treatment, further intervention directed toward blood pressure control and other modifiable risks should be considered. PMID- 15655124 TI - Antihypertensive therapy increases cerebral blood flow and carotid distensibility in hypertensive elderly subjects. AB - Many physicians are reluctant to lower blood pressure to recommended levels in elderly hypertensive patients because of concern about producing cerebral hypoperfusion. Because hypertension is associated with potentially reversible structural and functional alterations in the cerebral circulation that may improve with treatment, we investigated whether long-term pharmacological reduction of systolic blood pressure will improve, rather than worsen, cerebral blood flow and its regulation. Three groups of elderly subjects 65 years of age or older were studied prospectively: normotensive subjects (N=19), treated hypertensive subjects with systolic pressure <140 mm Hg (N=18), and uncontrolled hypertensive subjects with systolic pressure >160 mm Hg at entry into the study (N=14). We measured beat-to-beat blood flow velocity in the middle cerebral artery (transcranial Doppler ultrasonography), finger arterial pressure (photoplethysmography), and pulsatile distensibility of the carotid artery (duplex Doppler ultrasonography) at baseline and after 6 months of observation or antihypertensive therapy. After baseline hemodynamic measurements, uncontrolled hypertensive subjects underwent aggressive treatment with lisinopril with or without hydroclorothiazide or, if not tolerated, nifedipine or an angiotensin receptor blocker to bring their systolic pressure <140 mm Hg for 6 months. The other 2 groups were observed for 6 months. After 6 months of successful treatment, uncontrolled hypertensive subjects had significant increases in cerebral blood flow velocity and carotid distensibility that was not seen in the other groups. Treatment reduced cerebrovascular resistance and did not impair cerebral autoregulation. Therefore, judicious long-term treatment of systolic hypertension in otherwise healthy elderly subjects does not cause cerebral hypoperfusion. PMID- 15655125 TI - Parecoxib, valdecoxib, and cardiovascular risk. PMID- 15655126 TI - Cyclooxygenases, thromboxane, and atherosclerosis: plaque destabilization by cyclooxygenase-2 inhibition combined with thromboxane receptor antagonism. AB - BACKGROUND: Antagonism or deletion of the receptor (the TP) for the cyclooxygenase (COX) product thromboxane (Tx)A2, retards atherogenesis in apolipoprotein E knockout (ApoE KO) mice. Although inhibition or deletion of COX 1 retards atherogenesis in ApoE and LDL receptor (LDLR) KOs, the role of COX-2 in atherogenesis remains controversial. Other products of COX-2, such as prostaglandin (PG) I2 and PGE2, may both promote inflammation and restrain the effects of TxA2. Thus, combination with a TP antagonist might reveal an antiinflammatory effect of a COX-2 inhibitor in this disease. We addressed this issue and the role of TxA2 in the promotion and regression of diffuse, established atherosclerosis in Apobec-1/LDLR double KOs (DKOs). METHODS AND RESULTS: TP antagonism with S18886, but not combined inhibition of COX-1 and COX 2 with indomethacin or selective inhibition of COX-2 with Merck Frosst (MF) tricyclic, retards significantly atherogenesis in DKOs. Although indomethacin depressed urinary excretion of major metabolites of both TxA2, 2,3-dinor TxB2 (Tx M), and PGI2, 2,3-dinor 6-keto PGF(1alpha) (PGI-M), only PGI-M was depressed by the COX-2 inhibitor. None of the treatments modified significantly the increase in lipid peroxidation during atherogenesis, reflected by urinary 8,12-iso iPF(2alpha)-VI. Combination with the COX-2 inhibitor failed to augment the impact of TP antagonism alone on lesion area. Rather, analysis of plaque morphology reflected changes consistent with destabilization of the lesion coincident with augmented formation of TxA2. Despite a marked effect on disease progression, TP antagonism failed to induce regression of established atherosclerotic disease in this model. CONCLUSIONS: TP antagonism is more effective than combined inhibition of COX-1 and COX-2 in retarding atherogenesis in Apobec-1/LDLR DKO mice, which perhaps reflects activation of the receptor by multiple ligands during disease initiation and early progression. Despite early intervention, selective inhibition of COX-2, alone or in combination with a TP antagonist, failed to modify disease progression but may undermine plaque stability when combined with the antagonist. TP antagonism failed to induce regression of established atherosclerotic disease. TP ligands, including COX-1 (but not COX-2)-derived TxA2, promote initiation and early progression of atherogenesis in Apobec-1/LDLR DKOs but appear unimportant in the maintenance of established disease. PMID- 15655127 TI - Relationship of late loss in lumen diameter to coronary restenosis in sirolimus eluting stents. AB - BACKGROUND: Observed rates of restenosis after drug-eluting stenting are low (<10%). Identification of a reliable and powerful angiographic end point will be useful in future trials. METHODS AND RESULTS: Late loss (postprocedural minimum lumen diameter minus 8-month minimum lumen diameter) was measured in the angiographic cohorts of the SIRIUS (n=703) and E-SIRIUS (n=308) trials. Two techniques, the standard normal approximation and an optimized power transformation, were used to predict binary angiographic restenosis rates and compare them with observed restenosis rates. The mean in-stent late loss observed in the SIRIUS trial was 0.17+/-0.45 mm (sirolimus) versus 1.00+/-0.70 mm (control). If a normal distribution was assumed, late loss accurately estimated in-stent binary angiographic restenosis for the control arm (predicted 35.4% versus observed 35.4%) but underestimated it in the sirolimus arm (predicted 0.6% versus observed 3.2%). Power transformation improved the reliability of the estimate in the sirolimus arm (predicted 3.2% [CI 1.0% to 6.7%]) with similar improvements in the E-SIRIUS trial (predicted 4.0% [CI 1.2% to 7.0%] versus observed 3.9%). In the sirolimus-eluting stent arm, in-stent late loss correlated better with target-lesion revascularization than in-segment late loss (c statistic=0.915 versus 0.665). CONCLUSIONS: Because distributions of late loss with a low mean are right-skewed, the use of a transformation improves the accuracy of predicting low binary restenosis rates. Late loss is monotonically correlated with the probability of restenosis and yields a more efficient estimate of the restenosis process in the era of lower binary restenosis rates. PMID- 15655128 TI - Adrenomedullin enhances angiogenic potency of bone marrow transplantation in a rat model of hindlimb ischemia. AB - BACKGROUND: Previous studies have shown that adrenomedullin (AM) inhibits vascular endothelial cell apoptosis and induces angiogenesis. We investigated whether AM enhances bone marrow cell-induced angiogenesis. METHODS AND RESULTS: Immediately after hindlimb ischemia was created, rats were randomized to receive AM infusion plus bone marrow-derived mononuclear cell (MNC) transplantation (AM+MNC group), AM infusion alone (AM group), MNC transplantation alone (MNC group), or vehicle infusion (control group). The laser Doppler perfusion index was significantly higher in the AM and MNC groups than in the control group (0.74+/-0.11 and 0.69+/-0.07 versus 0.59+/-0.07, respectively, P<0.01), which suggests the angiogenic potency of AM and MNC. Importantly, improvement in blood perfusion was marked in the AM+MNC group (0.84+/-0.08). Capillary density was highest in the AM+MNC group, followed by the AM and MNC groups. In vitro, AM inhibited MNC apoptosis, promoted MNC adhesiveness to a human umbilical vein endothelial cell monolayer, and increased the number of MNC-derived endothelial progenitor cells. In vivo, AM administration not only enhanced the differentiation of MNC into endothelial cells but also produced mature vessels that included smooth muscle cells. CONCLUSIONS: A combination of AM infusion and MNC transplantation caused significantly greater improvement in hindlimb ischemia than MNC transplantation alone. This effect may be mediated in part by the angiogenic potency of AM itself and the beneficial effects of AM on the survival, adhesion, and differentiation of transplanted MNCs. PMID- 15655129 TI - Cholesteryl ester transfer protein TaqIB variant, high-density lipoprotein cholesterol levels, cardiovascular risk, and efficacy of pravastatin treatment: individual patient meta-analysis of 13,677 subjects. AB - BACKGROUND: Several studies have reported that the cholesteryl ester transfer protein (CETP) TaqIB gene polymorphism is associated with HDL cholesterol (HDL-C) levels and the risk of coronary artery disease (CAD), but the results are inconsistent. In addition, an interaction has been implicated between this genetic variant and pravastatin treatment, but this has not been confirmed. METHODS AND RESULTS: A meta-analysis was performed on individual patient data from 7 large, population-based studies (each >500 individuals) and 3 randomized, placebo-controlled, pravastatin trials. Linear and logistic regression models were used to assess the relation between TaqIB genotype and HDL-C levels and CAD risk. After adjustment for study, age, sex, smoking, body mass index (BMI), diabetes, LDL-C, use of alcohol, and prevalence of CAD, TaqIB genotype exhibited a highly significant association with HDL-C levels, such that B2B2 individuals had 0.11 mmol/L (0.10 to 0.12, P<0.0001) higher HDL-C levels than did B1B1 individuals. Second, after adjustment for study, sex, age, smoking, BMI, diabetes, systolic blood pressure, LDL-C, and use of alcohol, TaqIB genotype was significantly associated with the risk of CAD (odds ratio=0.78 [0.66 to 0.93]) in B2B2 individuals compared with B1B1 individuals (P for linearity=0.008). Additional adjustment for HDL-C levels rendered a loss of statistical significance (P=0.4). Last, no pharmacogenetic interaction between TaqIB genotype and pravastatin treatment could be demonstrated. CONCLUSIONS: The CETP TaqIB variant is firmly associated with HDL-C plasma levels and as a result, with the risk of CAD. Importantly, this CETP variant does not influence the response to pravastatin therapy. PMID- 15655130 TI - Irbesartan and lipoic acid improve endothelial function and reduce markers of inflammation in the metabolic syndrome: results of the Irbesartan and Lipoic Acid in Endothelial Dysfunction (ISLAND) study. AB - BACKGROUND: The metabolic syndrome is associated with increased angiotensin II activity, induction of a proinflammatory and oxidative state, and endothelial dysfunction. We evaluated the ability of irbesartan, an angiotensin receptor blocker, and lipoic acid, an antioxidant, to affect endothelial function and inflammation in patients with the metabolic syndrome. METHODS AND RESULTS: We randomized 58 subjects with the metabolic syndrome in a double-blinded manner to irbesartan 150 mg/d (n=14), lipoic acid 300 mg/d (n=15), both irbesartan and lipoic acid (n=15), or matching placebo (n=14) for 4 weeks. Endothelium-dependent and -independent flow-mediated vasodilation was determined under standard conditions. Plasma levels of interleukin-6, plasminogen activator-1, and 8 isoprostane were measured. After 4 weeks of therapy, endothelium-dependent flow mediated vasodilation of the brachial artery was increased by 67%, 44%, and 75% in the irbesartan, lipoic acid, and irbesartan plus lipoic acid groups, respectively, compared with the placebo group. Treatment with irbesartan and/or lipoic acid was associated with statistically significant reductions in plasma levels of interleukin-6 and plasminogen activator-1. In addition, treatment with irbesartan or irbesartan plus lipoic acid decreased 8-isoprostane levels. No significant changes in blood pressure were noted in any of the study groups. CONCLUSIONS: Administration of irbesartan and/or lipoic acid to patients with the metabolic syndrome improves endothelial function and reduces proinflammatory markers, factors that are implicated in the pathogenesis of atherosclerosis. PMID- 15655131 TI - Brugada syndrome: report of the second consensus conference: endorsed by the Heart Rhythm Society and the European Heart Rhythm Association. AB - Since its introduction as a clinical entity in 1992, the Brugada syndrome has progressed from being a rare disease to one that is second only to automobile accidents as a cause of death among young adults in some countries. Electrocardiographically characterized by a distinct ST-segment elevation in the right precordial leads, the syndrome is associated with a high risk for sudden cardiac death in young and otherwise healthy adults, and less frequently in infants and children. Patients with a spontaneously appearing Brugada ECG have a high risk for sudden arrhythmic death secondary to ventricular tachycardia/fibrillation. The ECG manifestations of Brugada syndrome are often dynamic or concealed and may be unmasked or modulated by sodium channel blockers, a febrile state, vagotonic agents, alpha-adrenergic agonists, beta-adrenergic blockers, tricyclic or tetracyclic antidepressants, a combination of glucose and insulin, hypo- and hyperkalemia, hypercalcemia, and alcohol and cocaine toxicity. In recent years, an exponential rise in the number of reported cases and a striking proliferation of articles defining the clinical, genetic, cellular, ionic, and molecular aspects of the disease have occurred. The report of the first consensus conference, published in 2002, focused on diagnostic criteria. The present report, which emanated from the second consensus conference held in September 2003, elaborates further on the diagnostic criteria and examines risk stratification schemes and device and pharmacological approaches to therapy on the basis of the available clinical and basic science data. PMID- 15655132 TI - Patients with depressive symptoms have lower health status benefits after coronary artery bypass surgery. AB - BACKGROUND: Depression is an established independent prognostic factor for mortality, readmission, and cardiac events after CABG surgery. However, limited data exist on whether depression influences functional outcomes after CABG. METHODS AND RESULTS: We followed 963 patients who underwent first CABG between February 1999 and February 2001. At baseline and at 6 months after CABG, we interviewed patients to assess depressive symptoms using the Geriatric Depression Scale (GDS) and physical function using the Short Form-36 Physical Component Scale (PCS). The patient's physical function was considered improved if the PCS score increased > or =5 points at 6 months. Patients with high GDS scores were younger, were more often female, and had worse physical function and higher comorbidity than patients with low GDS scores. Rates of improvement in physical function were 60.1% for a GDS score <5 (below 75th percentile), 49.8% for a GDS score between 5 and 9 (75th to 90th percentile), and 39.7% for a GDS score > or =10 (> or =90th percentile; P=0.002 for the trend). Depressive symptoms remained a significant independent predictor of lack of functional improvement after adjustment for severity of coronary artery disease, angina class, baseline PCS score, and medical history. A GDS score > or =10 was a stronger inverse risk factor for functional improvement after CABG than such traditional measures of disease severity as previous myocardial infarction, heart failure on admission, history of diabetes, and left ventricular ejection fraction. CONCLUSIONS: Higher levels of depressive symptoms at the time of CABG are a strong risk factor for lack of functional benefits 6 months after CABG. PMID- 15655133 TI - Heart failure and death after myocardial infarction in the community: the emerging role of mitral regurgitation. AB - BACKGROUND: In case series, mitral regurgitation (MR) increased the risk of death after myocardial infarction (MI), yet the prevalence of MR, its incremental prognostic value over ejection fraction (EF), and its association with heart failure and death after MI in the community is not known. METHODS AND RESULTS: The prevalence of MR and its association with heart failure and death were examined among 1331 patients within a geographically defined MI incidence cohort between 1988 and 1998. Echocardiography was performed within 30 days after MI in 773 patients (58%), and MR was present in 50% of cases, mild in 38%, and moderate or severe in 12%. Among patients with MR, a murmur was inconsistently detected clinically. After 4.7+/-3.3 years of follow-up, 109 episodes of heart failure and 335 deaths occurred. There was a graded positive association between the presence and severity of MR and heart failure or death. Moderate or severe MR was associated with a large increase in the risk of heart failure (relative risk 3.44, 95% CI 1.74 to 6.82, P<0.001) and death (relative risk 1.55, 95% CI 1.08 to 2.22, P=0.019) among 30-day survivors independent of age, gender, EF, and Killip class. CONCLUSIONS: In the community, MR is frequent and often silent after MI. It carries information to predict heart failure or death among 30-day survivors independently of age, gender, EF, and Killip class. These findings, which are applicable to a large community-based MI cohort, suggest that the assessment of MR should be included in post-MI risk stratification. PMID- 15655134 TI - Vascular endothelial dysfunction and mortality risk in patients with chronic heart failure. AB - BACKGROUND: Endothelial function is known to be impaired in subjects with chronic heart failure (CHF), but the association between endothelial function and subsequent mortality risk in CHF has not been previously reported. METHODS AND RESULTS: Biomarkers of endothelial function in the systemic arterial circulation (flow-mediated dilation [FMD] in the brachial artery) and the pulmonary circulation (exhaled nitric oxide [NO] production during submaximal exercise) were prospectively assessed in 259 subjects with New York Heart Association class II-III CHF. In subjects with FMD measurements (n=149), there were 12 deaths and 5 urgent transplantations over a median follow-up period of 841 days. In subjects with exhaled NO production measurements (n=110), there were 18 deaths and 1 urgent transplantation over a median follow-up period of 396 days. Both decreased FMD and decreased exhaled NO production were associated with increased risk of death or urgent transplantation after adjustment for other known CHF prognostic factors (age, etiology of CHF, functional class, left ventricular ejection fraction) in Cox multivariate proportional-hazards models (adjusted hazard ratio [HR] estimate for a 1% decrease in FMD=1.20; 95% confidence interval [CI], 1.03 to 1.45; P=0.027; adjusted HR estimate for a 1-ppb/min decrease in exhaled NO production=1.31, 95% CI, 1.01 to 1.69, P=0.04). CONCLUSIONS: Endothelial dysfunction in CHF, as assessed by FMD in the brachial artery and exhaled NO production during submaximal exercise, is associated with an increased mortality risk in subjects with both ischemic and nonischemic CHF. PMID- 15655135 TI - Elevated brain natriuretic peptide predicts blood pressure response after stent revascularization in patients with renal artery stenosis. AB - BACKGROUND: A significant number (20% to 40%) of hypertensive patients with renal artery stenosis will not have blood pressure improvement after successful percutaneous revascularization. Identifying a group of patients with refractory hypertension and renal artery stenosis who are likely to respond to renal stent placement would be beneficial. METHODS AND RESULTS: Brain natriuretic peptide (BNP) was measured in 27 patients with refractory hypertension and significant renal artery stenosis before and after successful renal artery stent placement. This neuropeptide was elevated (median, 187 pg/mL; 25th to 75th percentiles, 89 to 306 pg/mL) before stent placement and fell within 24 hours of the successful stent procedure (96 pg/mL; 25th to 75th percentiles, 61 to 182 pg/mL; P=0.002), remaining low (85 pg/mL; 25th to 75th percentiles, 43 to 171 pg/mL) at follow-up. Clinical improvement in hypertension was observed in the patients with a baseline BNP >80 pg/mL (n=22) in 17 patients (77%) compared with 0% of the patients with a baseline BNP < or =80 pg/mL (n=5) (P=0.001). After correction for glomerular filtration rate, BNP was strongly correlated with improvement in hypertension. CONCLUSIONS: BNP is increased in patients with severe renal artery stenosis and decreases after successful stent revascularization. In addition, an elevated baseline BNP level of >80 pg/mL appears to be a good predictor of a blood pressure response after successful stent revascularization. PMID- 15655136 TI - Losartan increases bradykinin levels in hypertensive humans. AB - BACKGROUND: Studies in animals and humans indicate a role for kinins in the actions of angiotensin type 1 (AT1) receptor blockers. However, the effect of these compounds on kinin levels in humans is unknown. METHODS AND RESULTS: We measured angiotensin (Ang), bradykinin (BK), and kallidin peptides in subjects with essential hypertension administered placebo, losartan (50 mg OD), and eprosartan (600 mg OD) in randomized order in a double-blind, 3-period, 3 treatment, crossover trial. Peptides were measured in arterial blood using high performance liquid chromatography-based radioimmunoassays. Losartan increased blood levels of BK-(1-9) and hydroxylated BK-(1-9) by approximately 2-fold and reduced the BK-(1-7)/BK-(1-9) ratio by 55%. There was a trend for eprosartan to produce similar changes in bradykinin levels. There were no changes in blood kallidin levels. Both losartan and eprosartan increased plasma levels of Ang I, Ang II, and Ang-(2-8), and eprosartan increased Ang-(3-8) levels. Ang-(1-7) and Ang-(1-9) levels were unchanged. There was an associated 30% to 35% reduction in Ang II/Ang I ratio and 63% to 69% reduction in Ang-(1-7)/Ang I ratio. Plasma ACE activity was unchanged. CONCLUSIONS: Losartan increases bradykinin levels. The reductions in BK-(1-7)/BK-(1-9), Ang II/Ang I, and Ang-(1-7)/Ang I ratios suggest that the increased bradykinin levels were the result of reduced metabolism by ACE and neutral endopeptidase. Increased bradykinin levels may represent a class effect of AT1 receptor blockers that contributes to their therapeutic actions and may also contribute to the angioedema that may accompany this therapy. PMID- 15655137 TI - Silicone granulomas treated with etanercept. PMID- 15655138 TI - Members of the national psoriasis foundation: more extensive disease and better informed about treatment options. AB - OBJECTIVE: Patient advocacy groups such as the National Psoriasis Foundation (NPF) serve as representatives of those affected by disease and provide information about the condition. Our objective was to assess the extent to which NPF members differ from nonmember patients with psoriasis in their knowledge and use of therapies. PARTICIPANTS: Using random-digit dialing, we identified and interviewed patients with psoriasis in the general US population. Randomly selected NPF members were also interviewed. MAIN OUTCOME MEASURES: Multivariate logistic regression models were used to estimate differences (odds ratios and 95% confidence intervals) in demographic and clinical characteristics and in awareness and use of therapies between members and others diagnosed as having psoriasis. RESULTS: Of 601 individuals with psoriasis identified from the general population survey, 185 provided a second interview and were defined as nonmembers. We interviewed 289 randomly selected members of the NPF. Although members were significantly older and wealthier and had more extensive disease, they reported the disease to be significantly less of a burden and were more satisfied with therapy than others affected. Compared with nonmembers, members were significantly more likely to have heard of and used most of the 10 therapies assessed. However, the proportion of respondents who were aware of a therapy and who also used it did not differ between groups. CONCLUSION: Members of the NPF are better informed and more satisfied with available treatment options than nonmember affected patients. PMID- 15655139 TI - Efficacy and safety observed during 24 weeks of efalizumab therapy in patients with moderate to severe plaque psoriasis. AB - OBJECTIVE: To assess the efficacy and safety of a 24-week course of efalizumab. DESIGN: Phase 3, randomized, double-blind, parallel-group, placebo-controlled 12 week study followed by a 12-week open-label study. SETTING: Outpatient dermatology clinics. Patients A total of 556 patients with moderate to severe chronic plaque psoriasis who were seeing an outpatient dermatologist were included in the study. Intervention For weeks 1 to 12, the 556 patients were randomized to receive 1 mg/kg of efalizumab weekly or placebo subcutaneously. For weeks 13 to 24, 516 of these patients received 1 mg/kg of efalizumab weekly. MAIN OUTCOME MEASURES: Proportion of patients with a 75% or greater improvement in Psoriasis Area and Severity Index (PASI-75), a 50% or greater improvement in PASI (PASI-50), static Physician's Global Assessment (sPGA) rating of minimal or clear, and improvements in Dermatology Life Quality Index (DLQI), itching scale, and Psoriasis Symptom Assessment (PSA) frequency and severity scores at weeks 12 and 24. Safety was evaluated by reviewing adverse events, laboratory parameters, vital signs, and anti-efalizumab antibodies. RESULTS: At week 12, 26.6% of efalizumab-treated patients achieved PASI-75 and 58.5% achieved PASI-50. After 24 weeks of continuous efalizumab therapy, PASI responses increased: 43.8% of patients achieved PASI-75 and 66.6% achieved PASI-50. The percentage of patients who achieved an sPGA rating of minimal or clear increased from 25.7% to 35.9%. The mean percentage of improvement in all patient-reported outcomes (DLQI, itching scale, and PSA frequency and severity scores) at week 12 was maintained at week 24 (DLQI, 49.2%; itching scale, 42.2%; PSA frequency, 47.6%; PSA severity, 47.3%). There was a decline in overall reported adverse events from weeks 1 to 12 (80.4%) to weeks 13 to 24 (63.2%) without evidence of cumulative toxic effects. Conclusion Extending efalizumab treatment from 12 to 24 weeks leads to improved efficacy and maintenance of quality of life with no evidence of cumulative toxic effects noted in patients with moderate to severe chronic plaque psoriasis. PMID- 15655140 TI - Topical tacrolimus ointment combined with 6% salicylic acid gel for plaque psoriasis treatment. AB - BACKGROUND: While oral tacrolimus is effective for the treatment of psoriasis, tacrolimus ointment has shown only spotty efficacy in the treatment of plaque psoriasis. The efficacy of tacrolimus ointment for the treatment of facial and intertriginous psoriasis suggests that if tacrolimus penetration can be increased, the ointment could be used for effective treatment of plaque psoriasis. OBJECTIVE: To assess whether tacrolimus ointment is an effective psoriasis treatment when used in a combination regimen with the penetration enhancer salicylic acid. METHODS: A total of 30 adult subjects with generally symmetrical plaque-type psoriasis were randomized to treatment with 6% salicylic acid gel plus vehicle or 6% salicylic acid gel plus 0.1% tacrolimus ointment in a 12-week left-right comparison study. The primary outcome was the difference between tacrolimus- and vehicle-treated target lesions in the change in the sum of erythema, scale, and thickness scores from baseline to end of treatment. RESULTS: A total of 24 subjects completed the trial. Combination treatment with tacrolimus ointment or vehicle plus salicylic acid gel was well tolerated. There was greater improvement of the sum score in the tacrolimus plus salicylic acid treated target plaques than in the vehicle plus salicylic acid-treated plaques at weeks 1, 2, and 8 (P<.05). The efficacy of this regimen was confirmed by investigator and subject global assessments of plaque severity. CONCLUSIONS: The combination of 0.1% tacrolimus ointment and 6% salicylic acid gel is an effective treatment for psoriasis. Although the results reported herein are from a small exploratory study, the magnitude of the effect was sufficiently large as to be detectable with statistical significance (P<.05). PMID- 15655142 TI - Absence of ribosomal RNA of Mycobacterium tuberculosis complex in sarcoidosis. AB - OBJECTIVE: To determine whether Mycobacterium tuberculosis ribosomal RNA (rRNA) is present in fresh tissue specimens from patients with sarcoidosis. DESIGN: A prospective study. SETTING: A university-based hospital. Patients Thirty-five patients diagnosed as having sarcoidosis at the University Hospital of Bellvitge, Barcelona, Spain, were included in the study. Fresh tissue samples with granulomatous inflammation were prospectively collected between 1997 and 2001 from all patients. For each sample tested, approximately 1 negative control was included. MAIN OUTCOME MEASURES: Mycobacterium tuberculosis rRNA was detected using an isothermal enzymatic amplification system of target rRNA of M tuberculosis complex via DNA intermediates. Smears for acid-fast staining and mycobacteriological cultures were also obtained. RESULTS: A total of 78 biopsy specimens (57 skin, 10 lymph node, 3 lacrimal gland, 2 spleen, 2 lung, 2 muscle, 1 bone, and 1 nerve) collected from 74 patients (35 patients with sarcoidosis and 39 control patients) were included in the study. Stains for acid-fast bacilli and mycobacterial cultures were negative for organisms in all cases. Mycobacterium tuberculosis rRNA was not detected in the specimens from any patients with sarcoidosis or in those from control patients whose cultures were negative for organisms. Ribosomal RNA was detected in 6 tissue specimens from patients with cultures that were positive for M tuberculosis and that were processed in parallel to the samples included in the study. CONCLUSIONS: Although previous studies have reported that mycobacterial antigens may play a role in granuloma formation in some patients with sarcoidosis, our results suggest that M tuberculosis cannot be considered to be the etiologic agent of the disease. PMID- 15655141 TI - Treatment and prophylaxis of seborrheic dermatitis of the scalp with antipityrosporal 1% ciclopirox shampoo. AB - OBJECTIVE: To demonstrate the efficacy, safety, and tolerance of ciclopirox shampoo for treatment and prophylaxis of seborrheic dermatitis of the scalp. DESIGN: Multicenter, randomized, double-blind, vehicle-controlled study. After treatment with ciclopirox shampoo once or twice weekly or vehicle for 4 weeks (study segment A), responders were randomized to a 12-week prophylactic study arm (segment B). SETTING: Forty-five medical centers in Germany (n = 19), France (n = 15), the United Kingdom (n = 8), and Austria (n = 3). Patients A total of 1000 patients with stable or exacerbating seborrheic dermatitis of the scalp. Interventions A total of 949 patients were randomized to receive ciclopirox treatment once or twice weekly or vehicle for 4 weeks. Thereafter, 428 responders received either ciclopirox prophylaxis once weekly or every 2 weeks or vehicle for 3 months. MAIN OUTCOME MEASURES: Primary and secondary: response of "effectively treated" and "cured," with investigators and patients rating acceptability and tolerance. RESULTS: Ciclopirox twice and once weekly produced response rates of 57.9% and 45.4%, respectively, compared with 31.6% for vehicle. Relapses occurred in 14.7% of patients using prophylactic ciclopirox once weekly, 22.1% of those in the prophylactic group shampooing once every 2 weeks, and 35.5% in the vehicle group. The few adverse events were evenly distributed among groups. Local tolerance and cosmetic acceptability were "good" in more than 85% of subjects. CONCLUSIONS: Seborrheic dermatitis of the scalp responds well to 1% ciclopirox shampoo once or twice weekly for 4 weeks. A low relapse rate is maintained by once-weekly shampooing or shampooing once every 2 weeks. These treatments are safe and well-tolerated. PMID- 15655143 TI - Treatment of diffuse basal cell carcinomas and basaloid follicular hamartomas in nevoid basal cell carcinoma syndrome by wide-area 5-aminolevulinic acid photodynamic therapy. AB - OBJECTIVE: To report the use of wide-area 5-aminolevulinic acid photodynamic therapy to treat numerous basal cell carcinomas (BCCs) and basaloid follicular hamartomas (BFHs). DESIGN: Report of cases. SETTING: Roswell Park Cancer Institute. Patients Three children with BCCs and BFHs involving 12% to 25% of their body surface areas. Interventions Twenty percent 5-aminolevulinic acid was applied to up to 22% of the body surface for 24 hours under occlusion. A dye laser and a lamp illuminated fields up to 7 cm and 16 cm in diameter, respectively; up to 36 fields were treated per session. MAIN OUTCOME MEASURES: Morbidity, patient response, and light dose-photodynamic therapy response relationship and durability. RESULTS: Morbidity was minimal, with selective phototoxicity and rapid healing. After 4 to 7 sessions, with individual areas receiving 1 to 3 treatments, the patients had 85% to 98% overall clearance and excellent cosmetic outcomes without scarring. For laser treatments, a sigmoidal light dose-response relationship predicted more than 85% initial response rates for light doses 150 J/cm(2) or more. Responses were durable up to 6 years. Conclusion 5-Aminolevulinic acid photodynamic therapy is safe, well tolerated, and effective for extensive areas of diffuse BCCs and BFHs and appears to be the treatment of choice in children. PMID- 15655144 TI - Prevalence of neurofibromatosis 1 in German children at elementary school enrollment. AB - OBJECTIVE: To determine the prevalence of neurofibromatosis 1 (NF1) among 6-year old children in Germany. SETTING AND PATIENTS: A total of 152819 children aged 6 years in 6 German states were screened for NF1 during routine medical examinations at elementary school enrollment in cooperation with local health departments in 2000 and 2001. MAIN OUTCOME MEASURE: The prevalence of NF1 among 6 year-old German children was estimated to be 1:2996 (95% confidence interval, 1:2260 to 1:3984). RESULTS: Fifty-one NF1 cases were identified and confirmed by evaluation by appropriate medical specialists. Seven other children were found to have multiple cafe au lait spots without other apparent features of NF1. A minimum estimate of the crude prevalence was 3.0 per 10 000 (95% confidence interval, 2.3-4.0 per 10 000 population). The incidence of NF1 in this population was probably in the range of 30 to 38 cases per 100 000 live births. CONCLUSION: Our study obtained a nearly unbiased birth incidence of NF1 of approximately 1 in 2600 to 1 in 3000 and demonstrates that NF1 can be diagnosed by age 6 years in most cases by routine physical examination with special attention to the disease associated skin stigmata. PMID- 15655145 TI - Clinical trial registration: a step forward in providing transparency for the positive and negative results of clinical trials. PMID- 15655146 TI - Clinical trial registration: a statement from the International Committee of Medical Journal Editors. PMID- 15655147 TI - Screening school-aged children for neurofibromatosis 1: dermatology or economics and public policy? PMID- 15655148 TI - Patient education and advocacy groups: a means to better outcomes? PMID- 15655149 TI - Definitions of measures of effect duration for psoriasis treatments. PMID- 15655150 TI - Necrolytic acral erythema: a patient from the United States successfully treated with oral zinc. AB - BACKGROUND: Recently, necrolytic acral erythema (NAE) has been described as a cutaneous marker for hepatitis C virus (HCV) infection. Only 2 cases have been reported in the United States. Successful remission has been induced only with interferon therapy with or without ribavirin. OBSERVATIONS: We describe a 46-year old, HCV-positive African American woman with well-defined, dusky, erythematous plaques on the dorsa of the feet, Achilles tendons, legs, knees, and elbows. Histologic examination revealed confluent upper epidermal necrosis, acanthosis, papillomatosis, and superficial and deep perivascular inflammation. She was diagnosed as having NAE. We induced successful disease remission with oral zinc administration. This is the third NAE case reported in the United States and the first report of disease remission with oral zinc therapy alone. CONCLUSIONS: Since its initial description in Egypt, more cases of NAE are being reported in the United States. Increased awareness of this entity is crucial. Oral zinc might represent a less toxic alternative therapeutic option for patients with NAE. PMID- 15655152 TI - Subcutaneous nodules on the face--quiz case. PMID- 15655151 TI - Clonal T cells in the blood of patients with systemic sclerosis. AB - BACKGROUND: It has been suggested that clonal T cells may play a critical role in the pathogenesis of systemic sclerosis. OBSERVATIONS: A monoclonal population of T cells was found in blood samples from 13 (34%) of 38 consecutive patients with a definite diagnosis of systemic sclerosis who were prospectively examined by T cell receptor gamma gene rearrangement using polymerase chain reaction analysis and denaturating gradient gel electrophoresis. In the healthy control group, the same type of examination revealed a monoclonal population of T cells in the blood samples from only 3 healthy subjects (4%)(odds ratio, 12.28; 95% confidence interval, 2.76-54.64; P = .001). Patients who had a circulating clonal population of T cells were older than those who did not (67 years vs 48 years; P = .04). There was a marked relationship between systemic sclerosis subtypes and the presence of a circulating clonal population of T cells. Twelve (43%) of 28 patients with limited cutaneous sclerosis exhibited a circulating clonal population of T cells, whereas only 1 (10%) of the 10 patients with diffuse cutanous sclerosis had evidence of T-cell clonality (P<.01). CONCLUSIONS: Clonally expanded T cells were more commonly detected in patients with limited cutaneous sclerosis than in those with diffuse cutaneous sclerosis, which is also in accordance with a possible role of clonal T cells in patients with limited cutaneous sclerosis. PMID- 15655153 TI - A slightly erythematous, firm papule on the upper arm--quiz case. PMID- 15655154 TI - Multiple subcutaneous nodules with unknown metastases to the liver--quiz case. PMID- 15655155 TI - Widespread annular eruption in a Black man--quiz case. PMID- 15655156 TI - Evidence of an outbreak of syphilis among men who have sex with men in Rome. PMID- 15655157 TI - Nevus density and atopic eczema. PMID- 15655158 TI - Generalized nummular eczema secondary to peginterferon Alfa-2b and ribavirin combination therapy for hepatitis C infection. PMID- 15655159 TI - Ro/SSA-positive cutaneous lupus erythematosus induced by carbamazepine. PMID- 15655160 TI - Subcutaneous myeloid sarcoma. PMID- 15655161 TI - Telangiectatic reticular erythema unrelated to cardiac devices. PMID- 15655162 TI - Absence of re-epithelialization in a fatal case of toxic epidermal necrolysis: is hyperbilirubinemia a culprit? PMID- 15655163 TI - An unusual presentation of cutaneous leishmaniasis. PMID- 15655164 TI - Sentinel lymph node biopsy in recessive dystrophic epidermolysis bullosa and squamous cell carcinoma. PMID- 15655165 TI - Dermoscopy of dermatofibroma. PMID- 15655167 TI - Quantitative study of tissue-engineered cartilage with human bone marrow mesenchymal stem cells. AB - OBJECTIVES: To assess the possibility of cartilage tissue engineering using human mesenchymal stem cells (hMSCs) and to investigate the quantitative relationship between hMSCs and engineered cartilage. DESIGN: Human mesenchymal stem cells were cultured, cryopreserved, and expanded in vitro. Surface antigens were detected by flow cytometry. In vitro chondrogenesis of hMSCs and cryopreserved hMSCs was performed. The chondrogenesis-induced hMSCs were seeded onto polyglycolic acid scaffolds, cultured in vitro for 3 weeks in chondrogenic medium, and then implanted into nude mice. The implants were harvested after 10 weeks and examined with histologic and immunochemical staining. RESULTS: The construction of cartilages was identified grossly and histologically: 1.9 to 2.5 x 10(7) nucleated cells were obtained from 1 mL of bone marrow, and about 1 to 2 x 10(6) hMSCs were obtained from the primary culture. The number of hMSCs tripled at every passage and reached 1.4 to 2.8 x 10(12) at passage 15. The purity of hMSCs was 95% and 98% at the primary and the fourth passages, respectively. Twenty-one days was the optimal (induction rate, 95%) induction time, with no apparent differences in induction rates among different passages. Based on our findings, hMSCs from 0.07 to 0.14 mL of bone marrow, expanded during 4 passages and induced for 21 days, would be sufficient to engineer 1 cm(2) of cartilage, 3-mm thick. CONCLUSION: Quantitative standards of hMSCs as seed cells for cartilage tissue engineering were established and may have value for later clinical work. PMID- 15655168 TI - Modulation of cutaneous aging with calorie restriction in Fischer 344 rats: a histological study. AB - OBJECTIVE: To examine whether histological changes in skin owing to intrinsic aging in a laboratory rodent model are modulated by caloric restriction (CR). METHODS: The abdominal skin from colony-raised ad libitum-fed Fischer 344 rats and age-matched rats subjected to CR was studied in the light microscope using histological morphometric methods. Animals 4, 12, and 24 months or older were used in this study. We studied the skin to obtain (1) quantitative data on the depth of the epidermis, dermis, and fat layer, the epidermal cellular density, the percentage fraction of dermal collagen, elastic fibers, pilosebaceous units, and capillaries, and the fibroblast density; and (2) qualitative assessment of histological staining for dermal glycosaminoglycans. We analyzed data by means of general linear model 2-way analysis of variance to obtain significance for the effects of age, diet, and age-diet interaction. RESULTS: The ad libitum-fed rats showed age-related increase in the depth of the epidermis, dermis, and fat layer. Calorie restriction prevented these changes, but epidermal nuclear density appeared to be stimulated. A trend toward increased values for collagen and elastic fibers, fibroblasts, and capillaries in skin samples from CR rats was observed. Pilosebaceous units were not modified. Moderately reduced staining for the dermal glycosaminoglycans in the skin of CR rats was noticed. CONCLUSIONS: Histomorphological changes resulting from intrinsic aging affected some of the studied variables in the rat skin, and these changes were delayed or prevented by CR. Some stimulatory effects, such as increased densities of fibroblasts and capillary profiles and higher values of connective tissue fibers resulting from CR, were also observed. Cutaneous morphological changes due to natural aging in this rat model seem to be modified by physiological or metabolic alterations imposed by CR. PMID- 15655170 TI - Viability of crushed and diced cartilage grafts: a study in rabbits. AB - OBJECTIVE: To investigate the effects of dicing and different degrees of crushing on cartilage graft viability and outcome in rhinoplasty. METHODS: Cartilage was harvested from both ears of 29 rabbits. For each animal, 6 cartilage pieces were prepared as follows and inserted into the paraspinal subcutaneous tissue: (1) left intact, (2) diced to approximately 1 x 1-mm pieces and then wrapped in oxidized regenerated cellulose, (3) slightly crushed, (4) moderately crushed, (5) significantly crushed, and (6) severely crushed. Animals were killed at 2, 5, and 10 months, and graft specimens were microscopically examined. RESULTS: As crushing intensity rose, cartilage viability decreased and more cartilage tissue was transformed to connective tissue. The intact and slightly crushed grafts showed significant chondrocyte proliferation. This decreased as crushing intensity increased, and the severely crushed and diced cellulose-wrapped grafts exhibited almost no peripheral chondrocyte proliferation. CONCLUSIONS: Slight crushing of a cartilage graft can produce outstanding graft material that forms softer nasal contours and fills defects well. However, severe crushing of cartilage grafts results in extensive necrosis and eventual reduction in graft volume. The use of oxidized regenerated cellulose to wrap diced cartilage grafts also tends to reduce clinical predictability owing to negative effects on cartilage viability and regeneration. PMID- 15655169 TI - Functional recovery after facial and sciatic nerve crush injury in the rat. AB - OBJECTIVES: To systematically record rat facial nerve recovery following crush injury to the main trunk with respect to ocular and vibrissial function and to compare the rates of facial and sciatic nerve recovery from crush injury in the same animals. This serves as a means of validating the functional parameters of facial nerve recovery against the well-known measure of hind limb function, the Sciatic Function Index. METHODS: The main trunk of the facial nerve and the proximal segment of the sciatic nerve were exposed in all animals. Both nerves were subjected to standardized crush injury and subsequent daily functional testing. After a plateau of functional recovery was achieved, the animals were killed, and the distances between the sites of injury and the end musculature were measured, which allowed determination and comparison of recovery rates in both systems. RESULTS: All crush injuries resulted in loss of electrical conductivity, as proven by intraoperative proximal nerve stimulation. Recovery of ocular and vibrissial motor function occurred starting at postoperative day (POD) 9 and continuing through POD 20. Hind limb function returned later (POD 14-34); however, when corrected for distance, the sciatic recovery rate (2.26 mm/d) appeared to match that of the facial nerve (1.5-2.4 mm/d). CONCLUSIONS: Recovery after facial nerve crush injury follows a predictable time course, and the rate of recovery is consistent with that of sciatic nerve injury. Return of the blink reflex, loss of vibrissial fibrillations, and return of vibrissial sweeping function appear to be internally consistent functional measures of facial recovery. These quantitative measures will be useful for future facial nerve manipulation studies. PMID- 15655171 TI - Effects of copper tripeptide on the growth and expression of growth factors by normal and irradiated fibroblasts. AB - OBJECTIVE: To evaluate the effects of copper tripeptide (GHK-Cu) on the growth and autocrine production of basic fibroblast growth factor, transforming growth factor beta1, and vascular endothelial growth factor by normal and irradiated fibroblasts in a serum-free in vitro environment. METHODS: Primary human dermal fibroblast cell lines were established after explantation from intraoperative specimens obtained from patients who had undergone radiation therapy for head and neck cancer. Normal and irradiated fibroblasts were propagated in serum- and growth factor-free media. Treatment groups were exposed to GHK-Cu (1 x 10(-9) mol/L). We measured cell counts and production of basic fibroblast growth factor, transforming growth factor beta1, and vascular endothelial growth factor. RESULTS: Irradiated fibroblasts survived and replicated in serum-free media. The population-doubling times of normal and irradiated fibroblasts exposed to GHK-Cu were faster than those of nontreated controls. Irradiated fibroblasts treated with GHK-Cu doubled at a rate that approximated that of untreated controls, and produced significantly more basic fibroblast growth factor and vascular endothelial growth factor than untreated controls early after GHK-Cu exposure. CONCLUSIONS: Irradiated fibroblasts survive and replicate in serum-free media, establishing this model as ideal for evaluating growth factor production in vitro. Copper tripeptide accelerates the growth of normal and irradiated fibroblasts to the point where treated irradiated fibroblasts approximate the population-doubling time of normal controls. An early increase in basic fibroblast growth factor and vascular endothelial growth factor production by GHK Cu-treated irradiated fibroblasts may improve wound healing. PMID- 15655172 TI - Rhinoplasty approaches: current state of the art. AB - OBJECTIVE: To survey rhinoplasty surgeons to determine their current approaches and the reason for their use, how they have acquired their knowledge, and trends in the use of open and closed approaches. DESIGN: A rhinoplasty questionnaire on open and closed rhinoplasty approaches was presented to surgeons attending the annual meetings of the American Academy of Facial Plastic and Reconstructive Surgery and the American Academy of Otolaryngology-Head and Neck Surgery. The 178 questionnaires were reviewed for analysis, and the results were tabulated by a statistician. Raw data were analyzed and cross-tabulations of specific subsets were reviewed. Main outcome measures included proportion of surgeons in various demographic groups using each technique, preferred incision, reasons for using each technique, usage over the past 5 years, and expected trend in the next 5 years. RESULTS: Most of the surgeons were in private practice, and most described their practice as otolaryngology (ear, nose, and throat [ENT]), facial plastic surgery (FPS), or ENT/FPS. Of the 178 responding surgeons, 46% had FPS practices and 27% devoted 90% to 100% of their practice to FPS. Most perform 100 or fewer rhinoplasties annually, and 23% perform open rhinoplasty 90% to 100% of the time. Sixty-three percent of FPS surgeons and 55% of ENT/FPS surgeons perform open septorhinoplasty (OSR) more than 50% of the time. Surgeons learn OSR mostly during residency (56%) and in didactic courses (51%) and learn closed rhinoplasty mostly during residency (75%). The most common indications for open rhinoplasty were difficult tip surgery (74%), revision procedures (73%), and grafting procedures (68%). Simple tip (65%) and simple dorsal (73%) procedures were common indications for closed rhinoplasty. The preferred incision for open rhinoplasty was the inverted "V" transcolumellar gull wing (58%); for closed rhinoplasty, cartilage delivery (48%) and intercartilaginous (28%). Most surgeons performed OSR at the same frequency during the past 5 years and expected to use OSR at the same frequency in the next 5 years. CONCLUSIONS: Overall, 53% of respondents used OSR more than 50% of the time. The movement toward open rhinoplasty seems to be plateauing, with a possible slight upward trend in its use. Over the past 5 years, there was still some trend toward the increasing use of the OSR approach, and most surgeons are performing OSR at the same frequency. Those with more than 5 years' experience believe that they are unlikely to change their approach in the next 5 years. Open septorhinoplasty may be indicated for rhinoplasties by a large proportion of surgeons, especially for rhinoplasties that are "difficult" or revisions or those requiring grafting. PMID- 15655174 TI - The effect of rhytidectomy on the nasal valve. AB - OBJECTIVES: To determine the effect of deep-plane rhytidectomy on the cross sectional area of the nasal cavity in the vicinity of the nasal valves and to compare this quantitative measure with patients' subjective assessment of their postoperative nasal airway. DESIGN: An inception cohort of 17 patients undergoing rhytidectomy (either cheek-lift or face-lift) for facial rejuvenation was evaluated with acoustic rhinometry. Initial measurements were taken approximately 1 week prior to surgery, followed by postoperative measurements at 1 week and again at 1 month (a total of 18 measurements per patient). Patients undergoing simultaneous nasal procedures were excluded. Control subjects consisted of patients undergoing facial plastic procedures other than rhytidectomy or septorhinoplasty (n = 3). The main outcome measure was cross-sectional area of both the internal and external valve regions as determined by acoustic rhinometry. The setting was an ambulatory surgery center at a large academic institution. RESULTS: Seventy percent of patients (12 of 17) reported subjective improvement of their nasal airway patency following rhytidectomy, whereas no control subjects (0%) reported any such change. Eighty-eight percent of patients (15 of 17) had a substantial increase in the dimension of their internal nasal valve area as measured with acoustic rhinometry at 1 week, with 70% of patients demonstrating increase at 1 month. Fifty-three percent of patients (9 of 17) demonstrated an increase in their external valve area at 1 week, and 59% had a persistent increase as measured at 1 month. No control subjects demonstrated any significant nasal valve area increases at either time. There was no correlation between age or body mass index and the measurement outcomes among participants. CONCLUSIONS: While there is a statistically significant increase in both the internal and external nasal valve cross-sectional areas at 1 month after rhytidectomy, the permanency of this effect is unknown. In support of these findings, a sizable proportion of patients undergoing rhytidectomy subjectively report an increase in their ability to breathe through their noses, lending credence to a postrhytidectomy melonasal effect. PMID- 15655173 TI - Efficacy of "thick" acellular human dermis (AlloDerm) for lower eyelid reconstruction: comparison with hard palate and thin AlloDerm grafts. AB - OBJECTIVES: To evaluate the efficacy of thick acellular human dermis (thick AlloDerm [LifeCell Corporation, The Woodlands, Tex]) grafts for posterior and middle lamellae reconstruction to correct lower eyelid retraction and to compare the long-term efficacy of thick AlloDerm with thin AlloDerm and hard palate grafts. METHODS: Retrospective analysis of patients undergoing lower eyelid reconstruction, which encompassed subperiosteal midface lifting, middle lamellae scar lysis, and placement of lower eyelid thick AlloDerm graft. Analysis included 21 surgical procedures in 11 patients. All patients had undergone at least 1 previous lower eyelid surgery with resultant lower eyelid retraction and scleral show. Preoperative and postoperative photographs were used for analysis. Measurements of the corneal diameter and distance from pupil center to lower eyelid margin were obtained, standardized, and compared. RESULTS: Of 21 procedures, 16 (8 of 11 patients) demonstrated improvement of lower eyelid position. The mean improvement of the median marginal reflex distance was 1.6 mm (range, 0.4-2.2 mm). The average follow-up after surgery was 215 days (range, 3 12 months). Of 21 procedures (3 patients), 5 failed to demonstrate improvement of lower eyelid position, with the mean final eyelid position lower postoperatively by 0.8 mm (range, 0.4-1.4 mm). CONCLUSIONS: We demonstrated long-lasting improvement of lower eyelid position with placement of thick AlloDerm grafts during lower eyelid reconstruction. The patients in our study had undergone previous lower eyelid blepharoplasty with resultant middle lamellae tethering. Surgical correction included subperiosteal midface-lift and middle lamellae scar lysis, in addition to thick AlloDerm graft placement to the lower eyelid. The results are comparable to hard palate grafts but perhaps superior to thin AlloDerm grafts. PMID- 15655176 TI - Barbed polypropylene sutures for midface elevation: early results. AB - BACKGROUND: In the past decade, the popularity of minimally invasive procedures for midface rejuvenation has greatly increased. We describe a new technique using percutaneous barbed polypropylene sutures to specifically address midface aging. It can be performed in a closed or open approach, depending on the degree of tissue repositioning required. OBJECTIVE: To determine the efficacy of the barbed polypropylene suture in the elevation of the aging midface and the rate of associated early complications. METHODS: Examination of the results obtained in 44 patients in whom we used the barbed polypropylene suture in a private practice plastic surgery facility. Of these patients, 34 underwent an open-approach and 10 a closed-approach procedure. Attention was focused on improvement of the aging midface, including the infraorbital hollow, lower lid position, nasolabial and melolabial creases, and nasojugal groove. Attention was also focused on improvement of the jowls. RESULTS: All patients demonstrated improvement in the midfacial area and jowls, with minimal complications. CONCLUSION: The suture evaluated in this article provides another technique that can be used by facial plastic surgeons in selected patients to address, in particular, ptosis of the midface. PMID- 15655175 TI - Use of the endoscopic forehead-lift to improve brow position in persistent facial paralysis. AB - Traditionally, the asymmetrical brow in facial paralysis has been treated with open procedures. There are few data that support the use of endoscopic procedures to treat patients with facial palsy or paralysis. We sought to evaluate a single surgeon's experience with the use of endoscopic forehead-lifts to treat asymmetrical brow positioning resulting from facial nerve disorders. All cases involving patients who underwent endoscopic brow-lifts by the senior author (Y.D.) from 1997 through 2003 with a minimum follow-up of 12 months were retrospectively reviewed. Demographic data were collected, and patient satisfaction was determined from postoperative interviews conducted at follow-up visits. Standard photographs were used to measure the degree of preoperative and postoperative brow asymmetry. A total of 31 cases were available for review. The average age of our patient population was 47 years (age range, 22-76 years), with a male-female ratio of almost 1.5:1. Twenty-three patients had a complete paralysis, and 8 patients had a palsy. The average preoperative difference in height at the desired apex of brow was 5.9 mm, with a range of 3.0 to 9.0 mm. The average postoperative difference (as measured at 12 months) in brow position was only 1.3 mm, with a range of 0 to 3 mm. Adjunctive periorbital procedures were performed in the majority of patients (90%) at the time of endoscopic brow lifting. All patients felt that their brow position was much improved after surgery. No major complications were encountered. A single patient underwent a secondary open direct browpexy to optimize his result. Endoscopic brow-lifting may be associated with favorable outcomes in the majority of patients with facial nerve palsy or paralysis. Performing concurrent adjunctive periorbital procedures as deemed necessary to optimize lower eyelid position, eyelid closure, and upper eyelid symmetry appears to be safe and reliable. PMID- 15655177 TI - Sir Joshua Reynolds' Lady Caroline Howard. PMID- 15655179 TI - Molecular profiling of tumor progression in head and neck cancer. AB - OBJECTIVE: To assess gene expression changes associated with tumor progression in patients with squamous cell carcinoma of the oral cavity. DESIGN: A microarray containing 17 840 complementary DNA clones was used to measure gene expression changes associated with tumor progression in 9 patients with squamous cell carcinoma of the oral cavity. Samples were taken for analysis from the primary tumor, nodal metastasis, and "normal" mucosa from the patients' oral cavity. SETTING: Tertiary care facility. Patients Nine patients with stage III or stage IV untreated oral cavity squamous cell carcinoma. RESULTS: Our analysis to categorize genes based on their expression patterns has identified 140 genes that consistently increased in expression during progression from normal tissue to invasive tumor and subsequently to metastatic node (in at least 4 of the 9 cases studied). A similar list of 94 genes has been identified that decreased in expression during tumor progression and metastasis. We validated this gene discovery approach by selecting moesin (a member of the ezrin/radixin/moesin [ERM] family of cytoskeletal proteins) and one of the genes that consistently increased in expression during tumor progression for subsequent immunohistochemical analysis using a head and neck squamous cell carcinoma tissue array. CONCLUSION: A distinct pattern of gene expression, with progressive up- or down-regulation of expression, is found during the progression from histologically normal tissue to primary carcinoma and to nodal metastasis. PMID- 15655180 TI - Cost comparison of surgery vs organ preservation for laryngeal cancer. AB - OBJECTIVE: To perform a cost minimization analysis of total laryngectomy with postoperative radiotherapy vs induction chemotherapy with subsequent radiotherapy in patients with advanced (stage III or IV) squamous cell carcinoma of the larynx. DESIGN: Decision-analysis model using data from peer-reviewed trials, case series, meta-analyses, and Medicare diagnosis related group reimbursement rates. SETTING AND PATIENTS: A hypothetical cohort of patients with stage III or IV laryngeal cancer. The perspective is that of a health care payer. INTERVENTIONS: The hypothetical patient cohort could receive (1) surgery (total laryngectomy) with postoperative radiotherapy or (2) induction chemotherapy (fluorouracil and cisplatin) with radiotherapy followed by salvage surgery for patients failing to respond to chemotherapy. MAIN OUTCOME MEASURE: Overall difference in direct medical costs in 2003 US dollars between the 2 treatment arms from initiation to completion of treatment. RESULTS: In the baseline analysis, the direct medical costs for the surgical arm were 30,138 US dollars per patient. For the organ preservation arm, the direct medical costs were 33,052 US dollars per patient. The finding that the surgical arm costs were lower was robust to all sensitivity analyses except for the extreme low estimate for the cost of chemotherapy. CONCLUSIONS: Our results suggest that total laryngectomy with postoperative radiotherapy costs nearly 3000 US dollars less than organ preservation treatment for advanced laryngeal cancer. Given that survival appears equivalent between the 2 modalities, cost consideration and patient preference may be important factors in decision making for the treatment of advanced laryngeal cancer. PMID- 15655181 TI - Comorbidity as a major risk factor for mortality and complications in head and neck surgery. AB - OBJECTIVE: To describe the impact of comorbidity on complications of surgery and mortality in patients with head and neck squamous cell carcinoma (HNSCC). DESIGN: A total of 120 consecutive patients with HNSCC, treated surgically between January 1999 and December 2001, were included. The Adult Comorbidity Evaluation 27 index (ACE-27) and the American Society of Anesthesiologists (ASA) risk classification system were used to describe comorbidity. Major complications were defined and scored by review of the medical records. Univariate and multivariate analyses were performed to determine the impact of 17 clinical variables, including the ACE-27 grade and the ASA class. RESULTS: Twenty-five patients (21.4%) had 1 or more major complications. In the univariate analysis, ACE-27 grade, ASA class, T stage, surgical procedure used for the primary tumor, type of neck dissection, and duration of anesthesia had a significant relation with major complications. In the multivariate analysis, duration of anesthesia and comorbidity reflected by the ACE-27 grade or the ASA class remained significant. The odds ratios (95% confidence intervals) associated with ACE-27 grades of 1 and 2 were 1.9 (0.6-6.8) and 4.6 (1.4-15.2), respectively; with ASA classes 2 and 3, 2.0 (0.5-8.2) and 10.0 (2.2-45.1), respectively. Duration of anesthesia longer than 360 minutes was characterized by an odds ratio of 7.8 (1.8-12.9). CONCLUSIONS: Duration of anesthesia and comorbidity reflected by the ACE-27 grade and the ASA class are important predictors of major complications in head and neck surgery. Optimizing the general condition of patients with HNSCC might reduce morbidity and treatment-related costs. PMID- 15655182 TI - Sentinel lymph node biopsy in N0 squamous cell carcinoma of the oral cavity and oropharynx. AB - OBJECTIVES: To ascertain the feasibility of sentinel lymph node (SLN) localization by preoperative lymphoscintigraphy and intraoperative gamma probe radiolocalization and to determine the predictive value of the SLN for occult metastasis of the neck in N0 squamous cell carcinoma of the oral cavity and oropharynx. DESIGN: A prospective study of 20 consecutive patients with N0 squamous cell carcinoma of the head and neck who underwent lymphoscintigraphy and SLN biopsy. INTERVENTIONS: On the day before surgery, each patient who completed the study underwent a submucosal peritumoral injection of unfiltered technetium 99m sulfur colloid followed by lymphoscintigraphy. Focal areas of radioactivity were marked on the overlying skin. The following day, the patients underwent resection of the primary tumor, elevation of subplatysmal flaps, identification and removal of the SLNs as identified by gamma probe, and complete neck dissections. RESULTS: Lymphoscintigraphy and gamma probe radiolocalization accurately identified 1 or more SLNs in all 20 patients. In 4 (20%) of the 20 patients, the SLN correctly identified metastatic disease. In no instance was the SLN negative when the lymphadenectomy specimen was positive. CONCLUSIONS: In this study, the SLN had a negative predictive value of 100%. Sentinel lymph node biopsy is feasible and appears to accurately predict the presence of occult metastatic disease. Although further study is warranted, SLN biopsy could potentially guide head and neck oncologists to the patient with N0 disease who would benefit most from selective neck dissection and prevent the morbidity of unnecessary neck dissection. PMID- 15655183 TI - Abnormal mesenchymal differentiation in the superior semicircular canal of brn4/pou3f4 knockout mice. AB - OBJECTIVE: To examine the developmental time course of the mutant phenotype and cellular mechanisms that result in malformations of the superior semicircular canal (SSCC) in Brn4 knockout mice. Mutations in the Brn4/Pou3f4 gene result in characteristic inner ear abnormalities in mutant mouse pedigrees, and the findings in these mice are similar to those in human X-linked deafness type III. DESIGN: Mutant and control mice were killed at various neonatal time points to assess the development of the SSCC. Measurements of SSCC diameter were made on paint-perfused specimens at postnatal day (P) 0, P7, P10, and P14. Histologic evaluation of the SSCC was made on hematoxylin-eosin-stained sections at P10. RESULTS: A dysmorphic constriction of the superior arc of the SSCC in Brn4 knockout mice was initially detectable at P14. Interestingly, the mutant SSCC is indistinguishable from control mice at earlier neonatal time points. In mutant neonates, there is persistence of immature woven bone with high cellularity surrounding the perilymphatic space of the SSCC. These findings are not present in control animal specimens, which demonstrate appropriate lamellar bony architecture. CONCLUSIONS: In Brn4 knockout mice, constriction of the SSCC with narrowing of the bony labyrinth develops in the postnatal period at approximately P14. The persistence of immature bone in affected mice indicates that signaling abnormalities disrupt normal mesenchymal differentiation in the SSCC. PMID- 15655184 TI - Vestibular toxic effects induced by once-daily aminoglycoside therapy. PMID- 15655185 TI - A parallel group analysis of tonsillectomy using the harmonic scalpel vs electrocautery. AB - OBJECTIVE: To evaluate efficiency and postoperative morbidity in tonsillectomy using the harmonic scalpel (HS) vs conventional electrocautery (EC). DESIGN: A retrospective medical chart review of all patients who underwent tonsillectomy at a single children's hospital during a 32-month period from January 2001 to August 2003. PATIENTS: The charts of 605 patients who underwent tonsillectomy or adenotonsillectomy were reviewed. The patients were stratified into 2 groups based on the dissecting instrument used (HS vs EC). Each group consisted of 2 subcategories based on age (< or =7 years and >7 years). Data collected included age, sex, operative indication, operative time, postanesthetic recovery room time, pain scale scores, postoperative admissions, and postoperative morbidities. RESULTS: There were significant differences in mean age (P<.001), mean weight (P<.001), and indication for surgery (P<.001) between the 2 methods: HS patients were younger, weighed less, and more often had obstructive symptoms as their primary operative indication compared with the EC group. Average operative times were not significantly different between methods. No differences were noted with respect to recovery time (HS group, P = .10; EC group, P = .30), postoperative oxygen requirements (HS group, P = .07; EC group, P = .09), and postoperative pain scores (HS group, P = .31; EC group, P = .58). There was a significant difference in postoperative admissions between the 2 methods in patients 7 years or younger (P = .005). Finally, we noted a significant difference in postoperative bleeding between the HS (2/292) and EC (13/313) methods when the data were compared per age group (P = .006), but the overall bleeding rate was not statistically significant owing to the small number of total bleeding instances (15/605). CONCLUSIONS: Recent prospective studies indicate that the HS provides advantages over conventional EC with respect to postoperative pain and return to normal activity. This study shows that HS tonsillectomy was as efficient as the conventional EC method. In addition, there was evidence that the rate of postoperative bleeding was significantly reduced by using the HS vs conventional EC. PMID- 15655186 TI - Child behavior and quality of life in pediatric obstructive sleep apnea. AB - OBJECTIVE: To assess behavior and quality of life in children with obstructive sleep apnea (OSA) undergoing tonsillectomy and adenoidectomy compared with control children. DESIGN: Prospective controlled study. SETTING: Hospital-based pediatric otolaryngology practice. PARTICIPANTS: Forty-two children (25 boys and 17 girls; mean [SD] age, 5.8 [2.5] years) with OSA confirmed by positive findings on polysomnography undergoing tonsillectomy and adenoidectomy and 41 control children (29 boys and 12 girls; mean [SD] age, 7.3 [3.8] years) with no history of snoring undergoing unrelated elective surgery. INTERVENTIONS: Parents completed the standardized Child Behavior Checklist and a validated pediatric OSA quality-of-life survey before and 3 months after surgery. MAIN OUTCOME MEASURES: Child Behavior Checklist T scores and score classifications and quality-of-life survey mean scores. RESULTS: Change in mean total problem T score was significantly greater in the OSA group (from 51.6 at baseline to 48.3 at follow up) than in controls (from 45.5 at baseline to 46.7 at follow-up) (P = .03). The improvement in total T score classification (normal vs borderline or abnormal) was significant for children with OSA compared with control children (P = .009). Children with OSA had significant improvements in the quality-of-life survey mean total score and all individual domain scores compared with controls (P<.001). CONCLUSIONS: Behavioral and emotional difficulties are found in children with documented OSA compared with control children, and they improve after treatment. Large improvements in disease-specific quality of life are also found. Scores on a standardized measure of behavior assessment demonstrated significant correlation with scores on a validated quality-of-life instrument. PMID- 15655187 TI - Airway management in conjoined twins: a rare indication for the EXIT procedure. PMID- 15655188 TI - Endovascular treatment of a petrous internal carotid artery aneurysm with hemotympanum and epistaxis using a coronary stent and detachable platinum coils: report of a case. PMID- 15655189 TI - Gefitinib therapy for life-threatening laryngeal papillomatosis. PMID- 15655190 TI - Nonsurgical management of pediatric vocal fold nodules. PMID- 15655193 TI - Radiology quiz case 1. Extracranial internal carotid artery (EICA) pseudoaneurysm. PMID- 15655194 TI - Radiology quiz case 2. Laryngeal plasmacytoma in a patient with known multiple myeloma (MM). PMID- 15655195 TI - Radiology quiz case 3. Retropharyngeal carotid artery. PMID- 15655196 TI - Radiology quiz case 4. A large unicystic ameloblastoma of mandible. PMID- 15655197 TI - Pathology quiz case. Nasal juvenile xanthogranuloma (JXG). PMID- 15655200 TI - Maine Medical Center. PMID- 15655201 TI - Surgery in Oman. AB - During the past 3 decades, Oman (a sultanate), a country with deep-rooted history, culture, and traditions, has undergone a remarkable transformation and modernization in all fields, including education and health care. It has progressively established a nationwide network of modern health services that are accessible even at the village level and have ranked Oman with the developed world. More than 300 surgeons provide a full range of surgical services for a population of 2.5 million. Medical education is firmly established, and accredited surgical residency training has assumed increasing importance during the past decade. Given the sustained growth and dynamic planning, the surgical services will continue to develop, with Omani surgeons playing an increasing role in the future. PMID- 15655202 TI - Initial experience of US Marine Corps forward resuscitative surgical system during Operation Iraqi Freedom. AB - HYPOTHESIS: Modern US Marine Corps (USMC) combat tactics are dynamic and nonlinear. While effective strategically, this can prolong the time it takes to transport the wounded to surgical capability, potentially worsening outcomes. To offset this, the USMC developed the Forward Resuscitative Surgical System (FRSS). By operating in close proximity to active combat units, these small, rapidly mobile trauma surgical teams can decrease the interval between wounding and arrival at surgical intervention with resultant improvement in outcomes. DESIGN: Case series. SETTING: Echelon 2 surgical units during the invasion phase of Operation Iraqi Freedom. PATIENTS: Ninety combat casualties, consisting of 30 USMC and 60 Iraqi patients, were treated in the FRSS between March 21 and April 22, 2003. INTERVENTIONS: Tactical surgical intervention consisting of selectively applied damage control or definitive trauma surgical procedures. MAIN OUTCOME MEASURES: Time to surgical intervention and outcome following treatment in the FRSS. RESULTS: Ninety combat casualties with 170 injuries required 149 procedures by 6 FRSS teams. The USMC patients were received within a median of 1 hour of wounding with the critically injured being received within a median of 30 minutes. Fifty-three USMC personnel were killed in action and 3 died of wounds for a killed in action rate of 13.5% and a died of wounds rate of 0.8% during the invasion phase of Operation Iraqi Freedom. All Marines treated in the FRSS survived. CONCLUSION: The use of the FRSS in close proximity to the point of engagement during the initial, dynamic combat phase of Operation Iraqi Freedom prevented delays in surgical intervention of USMC combat casualties with resultant beneficial effects on patient outcomes. PMID- 15655203 TI - A single-layer, continuous, hand-sewn method for esophageal anastomosis: prospective evaluation in 218 patients. AB - HYPOTHESIS: A 1-layer, continuous technique for esophageal anastomosis after esophagectomy has been in routine use at the University of Hong Kong Medical Centre since 1996. This study aims to document the results of this method and examine factors that may predispose patients to complications associated with esophageal anastomosis. DESIGN: Retrospective study. SETTING: University academic surgical center. PATIENTS AND METHODS: We studied 218 consecutive patients who had an esophageal anastomosis constructed with a 1-layer, continuous technique. Data were prospectively collected. MAIN OUTCOME MEASURES: Morbidity and mortality rates, anastomotic leaks, stricture, and recurrences. RESULTS: Anastomotic leaks affected 7 patients (3.2%), of whom 3 required surgical reexploration and none died. The hospital mortality rate was 0.9% (2 patients), attributed to myocardial infarction and malignancy. Anastomotic strictures developed in 24 patients (11.1%). Multivariate analysis in those with gastric conduits showed that a cervical anastomosis (intrathoracic vs cervical; odds ratio, 0.27; 95% confidence interval, 0.08-0.87; P = .03) and use of the distal stomach (distal stomach vs whole stomach; odds ratio, 5.25; 95% confidence interval, 1.65-16.66; P = .005) were predictive of benign anastomotic stricture formation. Eleven patients (17.5%) who had a cervical anastomosis developed strictures compared with 13 (8.6%) in those who had intrathoracic anastomoses. Strictures developed in 12 patients (7.4%) with a whole stomach conduit and in 9 patients (19.6%) with a distal stomach conduit. Anastomotic recurrence occurred in 8 patients (3.7%); none had a histologically involved resection margin. CONCLUSIONS: The single layer, continuous, hand-sewn technique for esophageal anastomosis is safe and effective. Cervical anastomosis and use of the distal stomach were associated with more benign strictures. PMID- 15655204 TI - Laparoscopic Nissen fundoplication with prosthetic hiatal closure reduces postoperative intrathoracic wrap herniation: preliminary results of a prospective randomized functional and clinical study. AB - BACKGROUND: Postoperative intrathoracic wrap migration is the most frequent morphological complication after laparoscopic antireflux surgery. Previous authors have studied the use of prosthetic materials for hiatal closure to prevent recurrence of hiatal hernia and/or postoperative intrathoracic wrap herniation. HYPOTHESIS: Patients with prosthetic hiatal closure have a higher rate of short-term dysphagia but a significantly lower rate of postoperative intrathoracic wrap herniation at follow-up. DESIGN: Prospective randomized trial. We compared patients who underwent laparoscopic Nissen fundoplication with simple sutured hiatoplasty with those who underwent laparoscopic Nissen fundoplication with prosthetic hiatal closure. SETTING: University-affiliated community hospital. PATIENTS: One hundred consecutive patients undergoing laparoscopic Nissen fundoplication for gastroesophageal reflux disease and hiatal hernia repair. INTERVENTION: Laparoscopic Nissen fundoplication with simple sutured crural closure (n = 50 [group 1]) vs laparoscopic Nissen fundoplication with simple sutured cruroplasty and onlay of a polypropylene mesh (n = 50 [group 2]). MAIN OUTCOME MEASURES: Recurrences; complications; results of esophageal manometry, 24-hour pH monitoring, esophagogastroduodenoscopy, and barium swallow test; and symptomatic outcome. RESULTS: Patients in both groups had similar preoperative values in esophageal manometry, 24-hour pH monitoring, and symptom scoring. At the 3-month and 1-year follow-ups, functional outcome variables (lower esophageal sphincter pressure and DeMeester score) improved significantly compared with the preoperative values. A higher postoperative dysphagia rate could be evaluated in group 2. An intrathoracic wrap migration occurred in 13 patients (26%) in group 1 vs 4 (8%) in group 2 (P<.001). CONCLUSION: Laparoscopic Nissen fundoplication with prosthetic cruroplasty is an effective procedure to reduce the incidence of postoperative hiatal hernia recurrence and intrathoracic wrap herniation. PMID- 15655205 TI - Surgical management of multinodular goiter with compression symptoms. AB - HYPOTHESIS: Multinodular goiter (MG) with compression symptoms has a clinical profile different from that of goiter without these symptoms. The surgical treatment of MG with compression symptoms has a high rate of sternotomy and morbidity. DESIGN: Retrospective study conducted between 1970 and 1999. SETTING: Tertiary referral center. PATIENTS: One hundred fifty-seven patients with MG with compression symptoms were reviewed from 672 patients with MG undergoing surgery in our department. We used 515 patients with MG without compression symptoms as a control group. INTERVENTION: All 157 patients underwent programmed surgery for thyroidectomy. MAIN OUTCOME MEASURES: General patient data, history and symptoms, exploration (both physical and with complementary techniques), data on the surgery and surgeon, and postsurgery morbidity and evolution. The chi2 test, the t test, and a logistic regression test were applied. RESULTS: Multinodular goiter with compression symptoms is characterized by its appearance in persons older than 55 years, a preoperative evolution of more than 10 years, and an intrathoracic component in more than 75% (P<.001). All the patients underwent surgery, with 6 (4%) requiring a sternotomy. Twenty-four percent had complications (n = 37), 3% of which corresponded to 4 cases of permanent recurrent laryngeal nerve injury. Eleven patients (7%) had an associated thyroid carcinoma, 9 of them corresponding to microcarcinomas. However, 5 were multifocal, and there was 1 anaplastic carcinoma, from which the patient died. All the papillary carcinomas are currently asymptomatic. The symptoms were remitted after surgery in all the cases except 1 dysphonia. Of the 32 patients receiving partial surgery, 9 (28%) had recurrence, of whom 6 underwent reoperation to complete the thyroidectomy. CONCLUSIONS: Multinodular goiter with compression symptoms occurs in long-evolving goiters with an intrathoracic component. Surgery is the definitive treatment, as it excludes malignancy, involves low rates of permanent morbidity and mortality, and, if the technique is total thyroidectomy, avoids recurrences. PMID- 15655206 TI - Thoracic outlet decompression for subclavian vein thrombosis: experience in 71 patients. AB - HYPOTHESIS: There is a difference in outcomes when patients have neurogenic thoracic outlet syndrome in addition to subclavian vein thrombosis. METHODS: Analysis of a prospectively developed database, medical record review, and a patient questionnaire were used to summarize clinical experience from December 1990 to December 2001 on the basis of the patient's original evaluation. Patients were stratified on the presence (group 1) or absence (group 2) of additional neurogenic pathologic features. RESULTS: Of 928 patients evaluated for thoracic outlet syndrome, 71 underwent 73 operative procedures for subclavian vein obstruction. Men predominated (55%), and the mean age was 32 years. Group 1 (41%) had more preoperative disability, a higher incidence of persistent pain (24%), and less likelihood of returning to full activity compared with group 2 (67% vs 93%; P = .01). Catheter-directed thrombolysis was used in 65% of veins. Preoperative balloon angioplasty was used selectively (34%), and only 4% required stents. Supraclavicular decompression and venolysis were usually delayed 3 weeks to allow for healing of the venous endothelium. Complications included wound infection (3%) and postoperative hematoma (8%). CONCLUSIONS: Patients with isolated subclavian vein obstruction have a more favorable outcome relative to those with combined neurogenic and venous pathologic features. Decompression following thrombolysis should be delayed to reduce the incidence of postoperative complications. PMID- 15655207 TI - Risk factors for estrogen receptor-positive breast cancer. AB - HYPOTHESIS: Some risk factors associated with breast cancer may be more predictive of estrogen receptor (ER)- positive than ER-negative tumors. DESIGN: Survey of patients enrolled in a study of breast cancer risk factors. SETTING: Community population in a northern California county. PATIENTS: A total of 234 individuals diagnosed as having breast cancer between July 1, 1997, and June 30, 1999, reporting Marin County, California, residence and participating in a questionnaire regarding exposure to breast cancer risk factors. MAIN OUTCOME MEASURE: Diagnosis of ER-positive vs ER-negative breast cancer. RESULTS: Comparison between ER-positive and ER-negative cases showed several factors predictive of ER-positive tumors. In a multivariate model, years of hormone therapy use remained the most significant predictor of ER-positive disease. CONCLUSIONS: Patients diagnosed as having ER-positive breast cancer were more likely to have undergone hormone therapy. The excess of ER-positive breast cancers reported in Marin County could, therefore, in part, be related to hormone therapy. PMID- 15655208 TI - Surgical management of rectal prolapse. AB - BACKGROUND: The problem of complete rectal prolapse is formidable, with no clear predominant treatment of choice. Surgical management is aimed at restoring physiology by correcting the prolapse and improving continence and constipation with acceptable mortality and recurrence rates. Abdominal procedures are ideal for young fit patients, whereas perineal procedures are reserved for older frail patients with significant comorbidity. Laparoscopic procedures with their advantages of early recovery, less pain, and possibly lower morbidity are recently added options. Regardless of the therapy chosen, matching the surgical selection to the patient is essential. OBJECTIVE: To review the present status of the surgical treatment of rectal prolapse. DATA SOURCES: Literature review using MEDLINE. All articles reporting on rectopexy were included. STUDY SELECTION: Articles reporting on prospective and retrospective comparisons were included. Case reports were excluded, as were studies comparing data with historical controls. DATA EXTRACTION: The results were tabulated to show outcomes of different studies and were compared. Studies that did not report some of the outcomes were noted as "not stated." DATA SYNTHESIS: Abdominal operations offer not only lower recurrence but also greater chance for functional improvements. Suture and mesh rectopexy produce equivalent results. However, the polyvinyl alcohol (Ivalon) sponge rectopexy is associated with an increased risk of infectious complications and has largely been abandoned. The advantage of adding a resection to the rectopexy seems to be related to less constipation. Laparoscopic rectopexy has similar results to open rectopexy but has all of the advantages related to laparoscopy. Perineal procedures are better suited to frail elderly patients with extensive comorbidity. CONCLUSIONS: Abdominal procedures are generally better for young fit patients; the results of all abdominal procedures are comparable. Suture and mesh rectopexy are still popular with many surgeons-the choice depends on the surgeon's experience and preference. Similarly, the procedure may be done through a laparoscope or by laparotomy. Perineal procedures are preferable for patients who are not fit for abdominal procedures, such as elderly frail patients with significant comorbidities. The decision between perineal rectosigmoidectomy and Delorme procedures will depend on the surgeon's preference, although the perineal rectosigmoidectomy has better outcomes. PMID- 15655209 TI - Characterizing the general surgery workforce in rural America. AB - BACKGROUND: General surgeons form a crucial component of the medical workforce in rural areas of the United States. Any decline in their numbers could have profound effects on access to adequate health care in such areas. HYPOTHESIS: We hypothesize that the rural areas of the United States are relatively undersupplied with general surgeons. DESIGN AND SETTING: The American Medical Association's Physician Masterfile was used to identify all clinically active general surgeons as well as their locations and characteristics. Their geographic distribution was examined using the ZIP code version of the Rural-Urban Commuting Areas. Surgeons were classified as practicing in urban areas, large rural areas, or small/isolated rural areas. RESULTS: There are currently 17 243 general surgeons practicing in the United States. Nationally, the number of general surgeons per population of 100 000 varies from 6.53 in urban areas to 7.71 in large rural areas and 4.67 in small/isolated rural areas. Only 10.6% of the nation's general surgeons are female. Wide variations in numbers of general surgeons were found between and within individual states. General surgeons in the smallest rural areas are more likely than those in urban areas to be male (92.7% vs 88.3%, P<.001), 50 years of age or older (51.6% vs 42.1%, P<.001), or international medical graduates (25.2% vs 20.1%, P<.001). CONCLUSIONS: The overall size of the rural general surgical workforce has remained static over the last decade, but its demographic characteristics suggest that numbers will decline. Many rural residents have limited access to surgical services. Steps to reverse this trend are needed to preserve the viability of health care in many parts of rural America. PMID- 15655210 TI - Developing and testing competency levels for laparoscopic skills training. AB - HYPOTHESIS: Expert levels can be developed for use as training end points for a basic video-trainer skills curriculum, and the levels developed will be suitable for training. DESIGN: Fifty subjects with minimal prior simulator exposure were enrolled using an institutional review board-approved protocol. As a measure of baseline performance, medical students (n = 11) and surgery residents (n = 39) completed 3 trials on each of 5 validated video-trainer tasks. Four board certified surgeons established as laparoscopic experts (with more than 250 basic and more than 50 advanced cases) performed 11 trials on each of the 5 tasks. The mean score was determined and outliers (>2 SDs) were trimmed; the trimmed mean was used as the competency level. Baseline performance of each subject was compared with the competency level for each task. SETTING: All research was performed in a laparoscopic skills training and simulation laboratory. PARTICIPANTS: Medical students, surgical residents, and board-certified surgeons. MAIN OUTCOME MEASURES: Expert scores based on completion time and the number of subjects achieving these scores at baseline testing. RESULTS: For all tasks combined, the competency level was reached by 6% of subjects by the third trial; 73% of these subjects were chief residents, and none were medical students. CONCLUSIONS: These data suggest that the competency level is suitably challenging for novices but is achievable for subjects with more experience. Implementation of this performance criterion may allow trainees to reliably achieve maximal benefit while minimizing unnecessary training. PMID- 15655211 TI - Identifying risk factors for complications following sentinel lymph node biopsy for melanoma. AB - BACKGROUND: Sentinel lymph node biopsy has become routine in the staging of patients with cutaneous melanoma and is presumed to have fewer complications than elective regional lymph node dissection (RLND). However, little information is available to refute or support this assumption. HYPOTHESIS: Risk factors for complications following sentinel lymph node biopsy (SLNBX) can be identified. DESIGN: Retrospective medical record review. PATIENTS AND METHODS: The medical records of 339 consecutive patients undergoing SLNBX for melanoma between 1996 and 2003 at our institution were reviewed for complications. RESULTS: In our series of 339 patients, 20 complications (5.9%) were observed following SLNBX compared with 15 (19.5%) of 77 patients undergoing RLND during the same period (P<.001). Seroma formation, transient nerve injuries, and minor wound infections were the most frequently observed complications in patients undergoing SLNBX. In contrast, chronic lymphedema and wound infections were the most frequent complications observed in patients undergoing RLND. Patients with comorbid medical conditions had more complications following either SLNBX or RLND than those without. The number of lymph nodes excised and the placement of closed suction drainage were associated with an increased incidence of complications following SLNBX but not RLND. The incidence of annual complications inversely correlated with the cumulative number of SLNBXs performed during this period. CONCLUSIONS: Sentinel lymph node biopsy can be performed with a low incidence of complications. Experience with SLNBX decreases complications. Patients with more than 1 sentinel lymph node excised or a closed-suction drain placed at the time of SLNBX are at an increased risk of complications. PMID- 15655212 TI - The history of anatomy and surgery of the preperitoneal space. AB - Preperitoneal (properitoneal) space is the space between the peritoneum and transversalis fascia. Bogros (1786-1825) described a triangular space in the iliac region between the iliac fascia, transversalis fascia, and parietal peritoneum. In the modern concept, this space lies between the peritoneum and posterior lamina of the transversalis fascia. In 1858, Retzius described the homonymous space, situated anterior and lateral to the urinary bladder (prevesical space). In 1975, Fowler reported that the preperitoneal fascia of the groin is distinct from the transversalis fascia. Preperitoneal herniorrhaphy may be subdivided into 2 approaches: transperitoneal and inguinal. We present herein the evolution of approaches to the preperitoneal space from use of the transperitoneal (or posterior) to use of the anterior preperitoneal and posterior preperitoneal approaches. As anatomic knowledge has increased, the evolution of laparoscopic surgery has paralleled that of open procedures. PMID- 15655213 TI - Image of the month. Stromal cell sarcoma of the breast. PMID- 15655214 TI - Pancreatic head excavation: an update. PMID- 15655215 TI - More on incomplete bibliographies. PMID- 15655216 TI - Video-assisted thoracic surgery sympathectomy for hyperhidrosis. PMID- 15655217 TI - Open access in medical publishing: trends and countertrends. PMID- 15655218 TI - Neural tube defects. PMID- 15655221 TI - Sustainability of family medicine. PMID- 15655222 TI - Neural tube defects. PMID- 15655223 TI - Neural tube defects. PMID- 15655224 TI - Neural tube defects. PMID- 15655225 TI - Neural tube defects. PMID- 15655226 TI - Does the C in CME stand for "Continuing" or "Commercial"? PMID- 15655230 TI - Debate over online recruitment of organ donors. PMID- 15655231 TI - Delisting chiropractic and physiotherapy: false saving? PMID- 15655232 TI - Increasing private delivery of publicly funded services? PMID- 15655233 TI - How do the eyes move together? New understandings help explain eye deviations in patients with stroke. PMID- 15655234 TI - Screening for type 2 diabetes mellitus to prevent vascular complications: updated recommendations from the Canadian Task Force on Preventive Health Care. PMID- 15655235 TI - Is blood transfusion harmful in patients with acute coronary syndromes? PMID- 15655236 TI - Instability of Carbolith (lithium carbonate). PMID- 15655237 TI - A 53-year-old woman with abdominal pain and fullness. PMID- 15655238 TI - Lymphogranuloma venereum in the differential diagnosis of proctitis. PMID- 15655239 TI - Impact of specialist follow-up in outpatients with congestive heart failure. AB - BACKGROUND: There is uncertainty about whether physician specialty influences the outcomes of outpatients with congestive heart failure after adjustment for differences in case mix. Our objective was to determine the impact of physician specialty on outcomes in outpatients with new-onset congestive heart failure. METHODS: The study was a population-based retrospective cohort study involving patients with new-onset congestive heart failure discharged from 128 acute care hospitals in Alberta between Apr. 1, 1998, and July 1, 2000. Outcomes were resource utilization (clinic visits, emergency department visits and hospital admissions) and survival at 30 days and 1 year. RESULTS: A total of 3136 patients were discharged from hospital with a new diagnosis of congestive heart failure (median age 76 years, 50% men). Of these, 1062 (34%) received no follow-up visits for cardiovascular care, 738 (24%) were seen by a family physician (FP) alone, 29 (1%) by a specialist (cardiologist or general internist) alone and 1307 (42%) by both a specialist and an FP. Compared with patients who received no follow-up cardiovascular care, patients who received regular cardiovascular follow-up visits with a physician had fewer visits to the emergency department (38% v. 80%), fewer were admitted to hospital (13% v. 94%), and the adjusted 1-year mortality was lower (22% v. 37%) (all p < 0.001). Compared with patients who received combined specialist and FP care, patients cared for exclusively by FPs had fewer outpatient visits (median 9 v. 17 in the first year), fewer of these patients presented to the emergency department (24% v. 45% in the first year), and fewer were readmitted for cardiovascular care (7% v. 16%) (all p < 0.001). However, the adjusted mortality at 1 year was lower among patients treated with combined care (17% v. 28%, p < 0.001) despite a higher burden of comorbidities. In a multivariate model adjusting for comorbidities (with no cardiovascular follow-up visits as the reference category), the mortality was lower among patients followed on an outpatient basis by an FP alone (odds ratio [OR] 0.66, 95% confidence interval [CI] 0.53-0.82) or by an FP and a specialist (OR 0.34, 95% CI 0.28-0.42). In a proportional hazards model with time-dependent covariates (with adjustment for frequency of follow-up visits), the risk of all-cause mortality was reduced significantly (hazard ratio 0.98, 95% CI 0.97- 0.99) with each specialist visit compared with FP care alone. INTERPRETATION: Patients with congestive heart failure followed by both specialists and FPs had significantly better survival than those followed by FPs alone (or those who received no specific cardiovascular follow-up care). Methods to improve timely and appropriate access to specialists and to improve collaborative care structures are needed. PMID- 15655240 TI - Volume of screening mammography and performance in the Quebec population-based Breast Cancer Screening Program. AB - BACKGROUND: In the Quebec Breast Cancer Screening Program (Programme quebecois de depistage du cancer du sein [PQDCS]), radiologists' and facilities' volumes of screening mammography vary considerably. We examined the relation of screening mammography volume to rates of breast cancer detection and false-positive readings in the PQDCS. METHODS: The study population included 307,314 asymptomatic women aged 50-69 years screened during 1998-2000. Breast cancer detection rates were analyzed by comparing all women with screening-detected breast cancer (n = 1709) and a 10% random sample of those without (n = 30,560). False-positive rates were analyzed by comparing the 3159 women with false positive readings and the 27,401 others in the 10% random sample. Characteristics of participants, radiologists and facilities were obtained from the PQDCS information system. Data were analyzed by means of logistic regression. RESULTS: The rate of breast cancer detection appeared to be unrelated to the radiologist's screening-mammography volume but increased with the facility's screening mammography volume. The breast cancer detection rate ratio for facilities performing 4000 or more screenings per year, compared with those performing fewer than 2000, was 1.28 (95% confidence interval [CI] 1.07-1.52). In contrast, the frequency of false-positive readings was unrelated to the facility's screening volume but was inversely related to the radiologist's screening volume: the rate ratio for readers of 1500 or more screenings per year compared with those reading fewer than 250 was 0.53 (95% CI 0.35-0.79). INTERPRETATION: Radiologists' and facilities' caseloads showed independent and complementary associations with performance of screening mammography in the PQDCS. Radiologists who worked in larger facilities and read more screening mammograms had higher breast cancer detection rates while maintaining lower false-positive rates. PMID- 15655241 TI - How should abridged scientific articles be presented in journals? A survey of readers and authors. AB - SEVERAL SCIENTIFIC AND GENERAL MEDICAL JOURNALS publish full-length articles on their Web sites and abridged versions in their print journals. We surveyed a stratified random sample of BMJ readers and authors to elicit their preferred format for the abridged print version. Each participant received a research paper abridged in 3 different formats: conventional abridged version, journalistic version and enhanced-abstract version. Overall, 45% (95% confidence interval [CI] 42%-48%) of the respondents said they liked the conventional version most, 31% (95% CI 28%-34%) preferred the journalistic version and 25% (95% CI 22%-27%) preferred the enhanced-abstract version. Twenty-eight percent (95% CI 25%-32%) indicated that use of the journalistic format for abridged articles would very likely stop them from submitting papers to BMJ, and 13% (95% CI 11%-16%) said the use of the enhanced-abstract version would stop them from submitting to BMJ. Publishers of general medical journals who publish shortened articles should consider that authors and readers prefer a more conventional style of abridged papers. PMID- 15655242 TI - Patients with treatable malignant diseases--including heart failure--are entitled to specialist care. PMID- 15655243 TI - Breast cancer screening, diagnostic accuracy and health care policies. PMID- 15655244 TI - Oral antihyperglycemic therapy for type 2 diabetes mellitus. AB - Diabetes mellitus is a chronic disease that is growing in prevalence worldwide. Pharmacologic therapy is often necessary to achieve optimal glycemic control in the management of diabetes. Orally administered antihyperglycemic agents (OHAs) can be used either alone or in combination with other OHAs or insulin. The number of available OHAs has increased significantly in the last decade, which translates into more therapeutic options and complex decision-making for physicians. This review article is designed to help with these decisions. We review the mechanism of action, efficacy and side effects of the different classes of OHAs (alpha-glucosidase inhibitors, biguanides, insulin secretagogues, insulin sensitizers and intestinal lipase inhibitor) and discuss the current recommendations for their use. PMID- 15655245 TI - Hepatoma-derived growth factor. Significance of amino acid residues 81-100 in cell surface interaction and proliferative activity. AB - Hepatoma-derived growth factor (HDGF) has proliferative, angiogenic, and neurotrophic activity. It plays a putative role in the development and progression of cancer. When expressed in cells, the mitogenic activity of HDGF depends on its nuclear localization, but it also stimulates proliferation when added to the cell culture medium. A cell surface receptor for HDGF has not been identified so far. We investigated the interaction of various purified recombinant HDGF fusion proteins with the cell surface of NIH 3T3 fibroblasts. We showed that binding of a HDGF-beta-galactosidase fusion protein to the cell surface of NIH 3T3 fibroblasts was saturable, occurred with high affinity (K(D) = 14 nm), and had a proliferative effect. We identified a peptide comprising amino acid residues 81-100 within the amino-terminal part of HDGF that bound to the cell surface of NIH 3T3 cells with saturation and affinity values similar to those of HDGF. When added to primary human fibroblasts, this peptide stimulated proliferation. Substitution of a single amino acid (K96A) within this peptide was sufficient to abolish its binding to the cell surface and its proliferative activity. In contrast, when expressed transiently in NIH 3T3 cells, a HDGF-beta galactosidase fusion protein in which amino acid residues 81-100 were deleted still had proliferative activity, whereas a fusion protein containing only the 81 100 peptide did not. Our results suggest the existence of a plasma membrane located HDGF receptor for which signaling depends on amino acid residues 81-100 of HDGF. This region differs from the one that has been recently identified to be essential for mitogenic activity depending on the nuclear localization of HDGF. Thus, HDGF exerts its proliferative activity via two different pathways. PMID- 15655246 TI - Molecular characterization of N-acylethanolamine-hydrolyzing acid amidase, a novel member of the choloylglycine hydrolase family with structural and functional similarity to acid ceramidase. AB - Bioactive N-acylethanolamines, including anandamide (an endocannabinoid) and N palmitoylethanolamine (an anti-inflammatory and neuroprotective substance), are hydrolyzed to fatty acids and ethanolamine by fatty acid amide hydrolase. Moreover, we found another amidohydrolase catalyzing the same reaction only at acidic pH, and we purified it from rat lung (Ueda, N., Yamanaka, K., and Yamamoto, S. (2001) J. Biol. Chem. 276, 35552-35557). Here we report complementary DNA cloning and functional expression of the enzyme termed "N acylethanolamine-hydrolyzing acid amidase (NAAA)" from human, rat, and mouse. The deduced primary structures revealed that NAAA had no homology to fatty acid amide hydrolase but belonged to the choloylglycine hydrolase family. Human NAAA was essentially identical to a gene product that had been noted to resemble acid ceramidase but lacked ceramide hydrolyzing activity. The recombinant human NAAA overexpressed in HEK293 cells hydrolyzed various N-acylethanolamines with N palmitoylethanolamine as the most reactive substrate. Most interestingly, a very low ceramide hydrolyzing activity was also detected with NAAA, and N lauroylethanolamine hydrolyzing activity was observed with acid ceramidase. By the use of tunicamycin and endoglycosidase, NAAA was found to be a glycoprotein. Furthermore, the enzyme was proteolytically processed to a shorter form at pH 4.5 but not at pH 7.4. Expression analysis of a green fluorescent protein-NAAA fusion protein showed a lysosome-like distribution in HEK293 cells. The organ distribution of the messenger RNA in rats revealed its wide distribution with the highest expression in lung. These results demonstrated that NAAA is a novel N acylethanolamine-hydrolyzing enzyme that shows structural and functional similarity to acid ceramidase. PMID- 15655247 TI - Hyaluronan-CD44 interaction with IQGAP1 promotes Cdc42 and ERK signaling, leading to actin binding, Elk-1/estrogen receptor transcriptional activation, and ovarian cancer progression. AB - In this study, we have examined the interaction of hyaluronan (HA)-CD44 with IQGAP1 (one of the binding partners for the Rho GTPase Cdc42) in SK-OV-3.ipl human ovarian tumor cells. Immunological and biochemical analyses indicated that IQGAP1 (molecular mass of approximately 190 kDa) is expressed in SK-OV-3.ipl cells and that IQGAP1 interacts directly with Cdc42 in a GTP-dependent manner. Both IQGAP1 and Cdc42 were physically linked to CD44 in SK-OV-3.ipl cells following HA stimulation. Furthermore, the HA-CD44-induced Cdc42-IQGAP1 complex regulated cytoskeletal function via a close association with F-actin that led to ovarian tumor cell migration. In addition, the binding of HA to CD44 promoted the association of ERK2 with the IQGAP1 molecule, which stimulated both ERK2 phosphorylation and kinase activity. The activated ERK2 then increased the phosphorylation of both Elk-1 and estrogen receptor-alpha (ER alpha), resulting in Elk-1- and estrogen-responsive element-mediated transcriptional up-regulation. Down-regulation of IQGAP1 (by treating cells with IQGAP1-specific small interfering RNAs) not only blocked IQGAP1 association with CD44, Cdc42, F-actin, and ERK2 but also abrogated HA-CD44-induced cytoskeletal function, ERK2 signaling (e.g. ERK2 phosphorylation/activity, ERK2-mediated Elk-1/ER alpha phosphorylation, and Elk-1/ER alpha-specific transcriptional activation), and tumor cell migration. Taken together, these findings indicate that HA-CD44 interaction with IQGAP1 serves as a signal integrator by modulating Cdc42 cytoskeletal function, mediating Elk-1-specific transcriptional activation, and coordinating "cross-talk" between a membrane receptor (CD44) and a nuclear hormone receptor (ER alpha) signaling pathway during ovarian cancer progression. PMID- 15655248 TI - Long-chain acyl-CoA synthetase 6 preferentially promotes DHA metabolism. AB - Previously we demonstrated that supplementation with the polyunsaturated fatty acids (PUFA) arachidonic acid (AA) or docosahexaenoic acid (DHA) increased neurite outgrowth of PC12 cells during differentiation, and that overexpression of rat acyl-CoA synthetase long-chain family member 6 (Acsl6, formerly ACS2) further increased PUFA-enhanced neurite outgrowth. However, whether Acsl6 overexpression enhanced the amount of PUFA accumulated in the cells or altered the partitioning of any fatty acids into phospholipids (PLs) or triacylglycerides (TAGs) was unknown. Here we show that Acsl6 overexpression specifically promotes DHA internalization, activation to DHA-CoA, and accumulation in differentiating PC12 cells. In contrast, oleic acid (OA) and AA internalization and activation to OA-CoA and AA-CoA were increased only marginally by Acsl6 overexpression. Additionally, the level of total cellular PLs was increased in Acsl6 overexpressing cells when the medium was supplemented with AA and DHA, but not with OA. Acsl6 overexpression increased the incorporation of [(14)C]-labeled OA, AA, or DHA into PLs and TAGs. These results do not support a role for Acsl6 in the specific targeting of fatty acids into PLs or TAGs. Rather, our data support the hypothesis that Acsl6 functions primarily in DHA metabolism, and that its overexpression increases DHA and AA internalization primarily during the first 24 h of neuronal differentiation to stimulate PL synthesis and enhance neurite outgrowth. PMID- 15655249 TI - The E6AP ubiquitin ligase is required for transactivation of the hTERT promoter by the human papillomavirus E6 oncoprotein. AB - Most human cancer cells display increased telomerase activity that appears to be critical for continued cell proliferation and tumor formation. The E6 protein of malignancy-associated human papillomaviruses increases cellular telomerase in primary human keratinocytes at least partly via transcriptional activation of the telomerase catalytic subunit, hTERT. In the present study, we investigated whether E6AP, a ubiquitin ligase well known for binding and mediating some of the activities of the E6 oncoprotein, participated in the transactivation of the hTERT promoter. Our results demonstrate that E6 mutants that fail to bind E6AP are also defective for increasing telomerase activity and transactivating the hTERT promoter. More importantly, E6AP knock-out mouse cells and small interfering RNA techniques demonstrated that E6AP was required for hTERT promoter transactivation in both mouse and human cells. Neither E6 nor E6AP bound to the hTERT promoter or activated the promoter in the absence of the partner protein. With all transactivation-competent E6 proteins, induction of the hTERT promoter was dependent upon E box elements in the core promoter. It appears, therefore, that E6-mediated activation of the hTERT promoter requires a complex of E6-E6AP to engage the hTERT promoter and that activation is dependent upon Myc binding sites in the promoter. The recruitment of a cellular ubiquitin ligase to the hTERT promoter during E6-mediated transcriptional activation suggests a role for the local ubiquitination (and potential degradation) of promoter-associated regulatory proteins, including the Myc protein. PMID- 15655251 TI - MPP+: mechanism for its toxicity in cerebellar granule cells. AB - Cerebellar granule cells constitute the largest homogeneous neuronal population of the mammalian brain. However, they are not often used in studies that involve MPP+-neurotoxicity. Currently, it is known that the toxicity of MPP+ in cerebellar granule cells as well as in other models, including dopaminergic cells, results from activation of the apoptotic machinery after an initial oxidative burst with mitochondrial damage and energetic failure. Therefore, cerebellar granule cells serve as a good model to investigate the MPP+ effects and to study in vitro the molecular mechanism implicated in the genesis of Parkinson's disease. PMID- 15655250 TI - Neuronal migration and the role of reelin during early development of the cerebral cortex. AB - During development, neurons migrate to the cortex radially from periventricular germinative zones as well as tangentially from ganglionic eminences. The vast majority of cortical neurons settle radially in the cortical plate. Neuronal migration requires an exquisite regulation of leading edge extension, nuclear translocation (nucleokinesis), and retraction of trailing processes. During the past few years, several genes and proteins have been identified that are implicated in neuronal migration. Many have been characterized by reference to known mechanisms of neuronal and non-neuronal cell migration in culture; however, probably the most interesting have been identified by gene inactivation or modification in mice and by positional cloning of brain malformation genes in humans and mice. Although it is impossible to provide a fully integrated view, some patterns clearly emerge and are the subject of this article. Specific emphasis is placed on three aspects: first, the role of the actin treadmill, with cyclic formation of filopodial and lamellipodial extensions, in relation to surface events that occur at the leading edge of radially migrating neurons; second, the regulation of microtubule dynamics, which seems to play a key role in nucleokinesis; and third, the mechanisms by which the extracellular protein Reelin regulates neuronal positioning at the end of migration. PMID- 15655252 TI - Molecular mechanism of scorpion neurotoxins acting on sodium channels: insight into their diverse selectivity. AB - Scorpion toxins that affect sodium channel gating traditionally are divided into alpha- and beta-classes. They show vast diversity in their selectivity for phyletic- or isoform-specific sodium channels. This article discusses the molecular mechanism of the selectivity. Moreover, a phylogenetic tree of scorpion toxins has been constructed, which, together with the worldwide distribution of toxins and the zoogeographic dispersion of the studied genera, offers an insight into the evolution of diverse scorpion toxins. PMID- 15655253 TI - Regulation of recombinant and native hyperpolarization-activated cation channels. AB - Ionic currents generated by hyperpolarization-activated cation-nonselective (HCN) channels have been principally known as pacemaker h-currents (Ih), because they allow cardiac and neuronal cells to be rhythmically active over precise intervals of time. Presently, these currents are implicated in numerous additional cellular functions, including neuronal integration, synaptic transmission, and sensory reception. These roles are accomplished by virtue of the regulation of Ih by both voltage and ligands. The article summarizes recent developments on the properties and allosteric interactions of these two regulatory pathways in cloned and native channels. Additionally, it discusses how the expression and properties of native channels may be controlled via regulation of the transcription of the HCN channel gene family and the assembly of channel subunits. Recently, several cardiac and neurological diseases were found to be intimately associated with a dysregulation of HCN gene transcription, suggesting that HCN-mediated currents may be involved in the pathophysiology of excitable systems. As a starting point, we briefly review the general characteristics of Ih and the regulatory mechanisms identified in heterologously expressed HCN channels. PMID- 15655255 TI - Humanin: after the discovery. AB - Humanin (HN) is a novel neuroprotective factor that consists of 24 amino acid residues. HN suppresses neuronal cell death caused by Alzheimer's disease (AD) specific insults, including both amyloid-beta (betaAbeta) peptides and familial AD-causative genes. Cerebrovascular smooth muscle cells are also protected from Abeta toxicity by HN, suggesting that HN affects both neuronal and non-neuronal cells when they are exposed to AD-related cytotoxicity. HN peptide exerts a neuroprotective effect through the cell surface via putative receptor(s). HN activates a cellular signaling cascade that intervenes (at least) in activation of c-Jun N-terminal kinase. The highly selective effect of HN on AD-relevant cell death indicates that HN is promising for AD therapy. Additionally, a recent study showed that intracellularly overexpressed HN suppressed mitochondria-mediated apoptosis by inhibiting Bax activity. PMID- 15655254 TI - Nuclear receptor coregulators are new players in nervous system development and function. AB - Steroid/thyroid hormones and their cognate nuclear receptors (NRs) play important roles in nervous system development and function. The spatial and temporal gene expression that is regulated by NRs in the nervous system requires transcriptional intermediary coregulators, designated as coactivators and corepressors. These coregulators enhance or repress transcriptional activity of NRs and modulate their target gene transcription. Recent progress has largely advanced our understanding of the molecular mechanisms by which NR coregulators function in the nervous system. This article summarizes our current knowledge about the molecular mechanisms, expression patterns, and biological functions of NR coactivators, such as the p160 steroid receptor coactivator family, CBP, p300, BRG1, TRAP220, PGC-1alpha, ERAP140, NIX1, and E6-AP, as well as corepressors such as NCoR and SMRT. Accumulated findings suggest that the functional spectrum of NR coregulators is much broader than was initially speculated, and these coregulators likely contribute to many physiological aspects of nervous system development and function. PMID- 15655256 TI - Gap junctions: their importance for the dynamics of neural circuits. AB - Electrical coupling through gap junctions constitutes a mode of signal transmission between neurons (electrical synaptic transmission). Originally discovered in invertebrates and in lower vertebrates, electrical synapses have recently been reported in immature and adult mammalian nervous systems. This has renewed the interest in understanding the role of electrical synapses in neural circuit function and signal processing. The present review focuses on the role of gap junctions in shaping the dynamics of neural networks by forming electrical synapses between neurons. Electrical synapses have been shown to be important elements in coincidence detection mechanisms and they can produce complex input output functions when arranged in combination with chemical synapses. We postulate that these synapses may also be important in redefining neuronal compartments, associating anatomically distinct cellular structures into functional units. The original view of electrical synapses as static connecting elements in neural circuits has been revised and a considerable amount of evidence suggests that electrical synapses substantially affect the dynamics of neural circuits. PMID- 15655257 TI - Parkinson's and Alzheimer's diseases: protein aggregations and neuroprotection. PMID- 15655258 TI - Multiplication of the alpha-synuclein gene is not a common disease mechanism in Lewy body disease. AB - Lewy body disease (LBD) refers to a heterogeneous group of disorders presenting with parkinsonism and Lewy body (LB) formation. Although the relationship between dementing syndromes with LBs, Parkinson's disease, and Alzheimer's disease is unclear, the former constitute a common form of degenerative dementia and may account for up to 20% of cases in the elderly. We recently demonstrated triplication of the alpha-synuclein gene as the cause of disease in the Spellman Muenter kindred. Neuropathological examination of affected members of the kindred demonstrated extensive LB pathology consistent with diffuse LBD. We examined a large collection of pathologically confirmed LBD cases and found no evidence for multiplication of the alpha-synuclein gene, suggesting that this mechanism is not a common cause of LBD. PMID- 15655259 TI - The Role of alpha-synuclein assembly and metabolism in the pathogenesis of Lewy body disease. AB - Parkinson's disease (PD) and dementia with Lewy bodies (DLB) are members of a family of disorders characterized by the presence of inclusion bodies, or Lewy bodies (LBs), filled with aggregates of alpha-synuclein. These diseases are a leading cause of movement disorders and dementia in the aging population, and it is crucial to understand the factors leading to the accumulation and assembly of these alpha-synuclein aggregates. Previous studies have uncovered much about the factors leading to aggregation and the mechanisms causing neurotoxicity of these inclusion bodies; however, little is known about factors that promote the degradation and prevent the aggregation of alpha-synuclein. The present article provides a review of recent efforts in the investigation of factors involved in alpha-synuclein metabolism and the mechanisms involved in preventing accumulation of alpha-synuclein and degrading this molecule. Understanding these processes might provide targets for the development of novel therapies for disorders such as DLB and PD. PMID- 15655260 TI - Embryonic and adult stem cells as a source for cell therapy in Parkinson's disease. AB - The rationale behind the use of cells as therapeutic modalities for neurodegenerative diseases in general, and in Parkinson's disease (PD) in particular, is that they will improve patient's functioning by replacing the damaged cell population. It is reasoned that these cells will survive, grow neurites, establish functional synapses, integrate best and durably with the host tissue mainly in the striatum, renew the impaired wiring, and lead to meaningful clinical improvement. To increase the generation of dopamine, researchers have already transplanted non-neuronal cells, without any genetic manipulation or after introduction of genes such as tyrosine hydroxylase, in animal models of PD. Because these cells were not of neuronal origin, they developed without control, did not integrate well into the brain parenchyma, and their survival rates were low. Clinical experiments using cell transplantation as a therapy for PD have been conducted since the 1980s. Most of these experiments used fetal dopaminergic cells originating in the ventral mesencephalic tissue obtained from fetuses. Although it was shown that the transplanted cells survived and some patients benefited from this treatment, others suffered from severe dyskinesia, probably caused by the graft's excessive and uncontrolled production and release of dopamine. It is now recognized that cell-replacement strategy will be effective in PD only if the transplanted cells have the same abilities, such as dopamine synthesis and control release, reuptake, and metabolizing dopamine, as the original dopaminergic neurons. Recent studies on embryonic and adult stem cells have demonstrated that cells are able to both self-renew and produce differentiated tissues, including dopaminergic neurons. These new methods offer real hope for tissue replacement in a wide range of diseases, especially PD. In this review we summarize the evidence of dopaminergic neuron generation from embryonic and adult stem cells, and discuss their application for cell therapy in PD. PMID- 15655261 TI - Oxidative stress, induced by 6-hydroxydopamine, reduces proteasome activities in PC12 cells: implications for the pathogenesis of Parkinson's disease. AB - Mutations in familial Parkinson's disease (PD) have been associated with the failure of protein degradation through the ubiquitin-proteasome system (UPS). Impairment of proteasome function has also been suggested to play a role in the pathogenesis of sporadic PD. We examined the proteasome activity in PC12 cells treated with 6-hydroxydopamine (6-OHDA), the dopamine synthetic derivate used in models of PD. We found that 6-OHDA treatment increased protein oxidation, as indicated by carbonyl group accumulation, and increased caspase-3 activity. In addition, there was an increase in trypsin-, chymotrypsin-, and postacidic-like proteasome activities in cells treated with 10-100 microM 6-OHDA, whereas higher doses caused a marked decline. 6-OHDA exposure also increased mRNA expression of the 19S regulatory subunit in a dose-dependent manner, whereas the expression of 20S- and 11S-subunit mRNAs did not change. Administration of the antioxidant N acetylcysteine to 6-OHDA-treated cells prevented the alteration in proteasome functions. Moreover, reduction in cell viability owing to administration of proteasome inhibitor MG132 or lactacystin was partially prevented by the endogenous antioxidant-reduced glutathione. In conclusion, our data indicate that mild oxidative stress elevates proteasome activity in response to increase in protein damage. Severe oxidative insult might cause UPS failure, which leads to protein aggregation and cell death. Moreover, in the case of UPS inhibition or failure, the blockade of physiological reactive oxygen species production during normal aerobic metabolism is enough to ameliorate cell viability. Control of protein clearance by potent, brain-penetrating antioxidants might act to slow down the progression of PD. PMID- 15655262 TI - Iron and alpha-synuclein in the substantia nigra of MPTP-treated mice: effect of neuroprotective drugs R-apomorphine and green tea polyphenol (-)-epigallocatechin 3-gallate. AB - One of the prominent pathological features of Parkinson's disease (PD) is the abnormal accumulation of iron in the substantia nigra pars compacta (SNpc), in the reactive microglia, and in association with neuromelanin, within the melanin containing dopamine (DA) neurons. Lewy body, the morphological hallmark of PD, is composed of lipids, redox-active iron, and aggregated alpha-synuclein, concentrating in its peripheral halo and ubiquitinated, hyperphosphorylated, neurofilament proteins. The capacity of free iron to enhance and promote the generation of toxic reactive oxygen radicals has been discussed numerous times. Recent observations, that iron induces aggregation of inert alpha-synuclein to toxic aggregates, have reinforced the critical role of iron in oxidative stress induced pathogenesis of DA neuron degeneration and protein degradation via ubiquitination. N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)- and 6 hydroxydopamine-induced neurodegeneration in rodents and nonhuman primates is associated with increased presence of iron and alpha-synuclein in the SNpc. The accumulation of iron in MPTP-induced neurodegeneration has been linked to nitric oxide-dependent mechanism, resulting in degradation of prominent iron regulatory proteins by ubiquitination. Radical scavengers such as R-apomorphine and green tea catechin polyphenol (-)-epigallocatechin-3-gallate, as well as the recently developed brain-permeable VK-28 series derivative iron chelators, which are neuroprotective against these neurotoxins in mice and rats, prevent the accumulation of iron and alpha-synuclein in SNpc. This study supports the notion that a combination of iron chelation and antioxidant therapy, as emphasized on several occasions, might be a significant approach to neuroprotection in PD and other neurodegenerative diseases. PMID- 15655263 TI - New aspects of genetic contributions to Parkinson's disease. AB - Over the last few years, several genes for rare, monogenically inherited forms of Parkinson's disease (PD) have been mapped and/or cloned. In dominant families, mutations have been identified in the gene for alpha-synuclein. Aggregation of this protein in intracellular inclusions (Lewy bodies) may be crucial in the molecular pathogenesis of the disease. Three genes have been identified to cause autosomal-recessive early-onset parkinsonism: parkin, DJ1, and PINK1. These genes are thought to be involved in the proteasomal protein degradation pathway, in the cell's response to oxidative stress, and in mitochondrial function, respectively. It is therefore concluded that these cellular mechanisms may play an important role in the degenerative process of PD. There is also accumulating evidence that genetic factors play a role in the common sporadic form of PD, however their precise nature remains unknown. PMID- 15655265 TI - Design and rationale of the Japanese Coronary Artery Disease (JCAD) Study: a large-scale, multicentered prospective cohort study. AB - Since there is in sufficient evidence on patients with coronary artery disease in Japan, the Japanese Coronary Artery Disease (JCAD) Study, in which 217 institutions participate, was designed to collect basic data based on evidence based medicine (EBM). In this study, cardiac catheterization is performed on all cases to select study subjects confirmed as having CAD diagnosed based on the criteria that he or she has stenosis in at least one branch of a coronary artery to the extent of 75% or higher according to the AHA classification. Data including background information, risk factors, clinical management, and medication are to be collected over the web. The follow-up arm of the study consists of following each subject for three years to obtain data on the long term prognosis of patients with CAD while the other arm is for enrolling new subjects every six months who will be followed for six months only for the purpose of determining the latest trend in patients. The two arms of the study have been ongoing since April 2000. As of September 30, 2003, 15,506 subjects have been enrolled in the follow-up arm and the follow-up data have been entered in the database. The authors plan to report data showing any correlation between incidence rate, focusing mainly on cerebrocardiovascular events, and other factors such as the management of risk factors, and type and dosage of medications obtained in the largest cohort ever studied in Japan of patients with a coronary artery lesion confirmed by cardiac catheterization. PMID- 15655264 TI - Pathological proteins in Parkinson's disease: focus on the proteasome. AB - Parkinson's disease (PD) is a multifactorial disease that appears to arise from the effects of both genetic and environmental influences. Pesticides and heavy metals are the principle environmental factors that appear to impact on PD. The known genetic factors include multiple genes that have been identified in related parkinsonian syndromes, as well as alpha-synuclein. Genes associated with either PD or Parkinson-related disorders include parkin, DJ-1, ubiquitin C-terminal hydrolase isozyme L1 (UCH-L1), nuclear receptor-related factor 1, and alpha synuclein. Alpha-synuclein is particularly notable because it aggregates readily and is the main component of Lewy bodies (LBs). Aggregated alpha-synuclein binds the proteasome and potently inhibits proteasomal activity. Because ubiquitin accumulates in LBs, and parkin and UCH-L1 also interact with the ubiquitin proteasomal system, proteasomal dysfunction is thought to contribute to the pathophysiology of PD. Increasing numbers of experiments suggest that neurotoxins might interact with alpha-synuclein or other Parkinson-related proteins to contribute to the pathophysiology of PD. Transgenic animal models overexpressing alpha-synuclein develop age-dependent motor dysfunction and inclusions in the brain stem that contain alpha-synuclein. These models are very helpful in elucidating the pathophysiology of PD but do not completely recapitulate the disease process. The relationship between these transgenic models and PD is a subject of intense investigation. PMID- 15655266 TI - The effect of tirofiban on ST segment resolution in patients with non-ST elevated myocardial infarction. AB - ST segment resolution in ST elevated myocardial infarction has independent predictive value for congestive heart failure and death at 30 days. ST segment depression in unstable angina pectoris (UAP) and non-ST elevated myocardial infarction (NSTEMI) predicts high risk of MI and death and may discriminate patients likely to have greater benefit from aggressive antithrombotic and interventional therapy. This study assessed the effect of tirofiban added to conventional treatment on ST segment resolution in NSTEMI patients. Sixty-four patients were randomized to one of the two groups: 32 patients received conventional treatment while tirofiban was added in the second group of 32 patients. In the first group, 6 patients refused to participate further after giving initial informed consent while 1 patient in the tirofiban group dropped out. We had 26 patients (mean age, 59 years) in the conventional treatment group and 31 patients (mean age, 59 years) received also tirofiban. Tirofiban was administered by intravenous infusion over a 72 hour period. More than 50% regression of depression was considered to be ST segment resolution. The characteristics of the two groups were comparable (Table I). The ST segment resolution evolution did not differ at the 4th and 24th hours between the two groups. Significant differences occurred in the 72nd hour ECG (Table III). ST resolution was present in 67.9% of the tirofiban patients and in 32.1% of the conventional treatment group (P < 0.05). Tirofiban treatment was not associated with an increase in major bleeding even though there was a trend toward an increase in minor bleeding cases and did not influence the occurrence of refractory angina pectoris. PMID- 15655267 TI - Diabetes mellitus has an additional effect on coronary artery disease. AB - We investigated whether plasma levels of adiponectin in patients with both coronary artery disease (CAD) and diabetes mellitus (DM) are lower than in patients with CAD alone. We examined plasma adiponectin levels in 113 patients, 82 with CAD (40 of whom had both CAD and type 2 DM) and 31 normal controls. We found differences in plasma adiponectin levels between CAD patients with and without DM (7.8 +/- 4.75 versus 12.1 +/- 6.87 microg/mL, P = 0.002), between patients with CAD and controls (10.0 +/- 6.27 versus 15.3 +/- 5.38 microg/mL, P < 0.0001), and between men and women (10.2 +/- 6.41 versus 13.1 +/- 6.22 microg/mL, P = 0.017). Plasma adiponectin levels were correlated negatively with body mass index, triglyceride, total cholesterol, hemoglobin A1c, and fibrinogen levels (r = -0.456, P < 0.0001; r = -0.355, P < 0.0001; r = -0.286, P = 0.002; r = -0.299, P < 0.0001; r = -0.400, P < 0.0001, respectively), but were not significantly correlated with high sensitivity C-reactive protein or low density lipoprotein levels (r = -0.088, P = 0.352; r = -0.167, P = 0.077, respectively). Plasma adiponectin levels correlated positively with high density lipoprotein levels (r = 0.410, P < 0.0001). Our study demonstrates that plasma adiponectin levels in patients with both CAD and DM are lower than in patients with CAD alone. We speculate that people who have very low plasma adiponectin levels may be at increased risk of developing both CAD and DM. PMID- 15655268 TI - Simultaneous double external DC shock technique for refractory atrial fibrillation in concomitant heart disease. AB - Atrial fibrillation (AF) has been treated with DC shocks delivered transthoracically, but in 5-30% of patients, the procedures fail to restore sinus rhythm (SR). We hypothesized that applying high energy shock waves to the chest may overcome the inadequate penetration of electrical shock to the atrium. The aim of this study was to evaluate the efficacy of higher energy external DC shock for the treatment of refractory AF coexisting with cardiovascular disease using a synchronized double external defibrillator. Fifteen patients (mean age 65 +/- 8) with refractory AF to standard DC cardioversion (CV) underwent higher energy DC shock using a double external defibrillator. Concomitant heart disease was present in all patients. Warfarin and amiodarone (600 mg/day), were administered for at least three weeks duration before DC CV. Sedation was performed with IV midazolam. Two defibrillator paddles were positioned on the anterior and posterior chest wall in a right lateral decubitus position. Defibrillators were synchronized to the R waves and simultaneously 720 joules of energy was administered to the patients. Amiodarone (200 mg/day) was continuously administered after DC shock to maintain SR. Sinus rhythm was obtained in 13 patients. Sinus rhythm was persistent in 11 patients for six months duration. Creatine kinase MB fractions were normal at 4 (22 +/- 4 IU/L) and 12 hours (18 +/ 4 IU/L). None of the patients developed significant hemodynamic compromise or congestive heart failure, higher AV block, stroke, or transient ischemic cerebral events. The results indicate that higher energy DC shock application using a double external defibrillator is an effective and safe method for the cardioversion of refractory AF. We believe this procedure should be performed before internal atrial cardioversion. PMID- 15655269 TI - Dipyridamole stress echocardiography and ultrasonic myocardial tissue characterization in predicting myocardial ischemia, in comparison with dipyridamole stress Tc-99m MIBI SPECT myocardial imaging. AB - The purpose of this study was to validate whether dipyridamole stress ultrasonic tissue characterization with cyclic variation of integrated backscatter (CVIBS) compared with dipyridamole stress echocardiography and dipyridamole stress Tc99m MIBI SPECT myocardial perfusion scintigraphy could predict myocardial ischemia in patients with chronic coronary artery disease. Twenty patients (16 M, 4 F) who had coronary angiography for stable angina pectoris were included in the study. Mean age was 62 +/- 8 years. The left ventricle was divided into 16 segments. Regional wall motion analysis and CVIBS measurements were obtained from 16 myocardial segments at rest and after dipyridamole (0.84 mg/kg) infusion. After 10 minutes, Tc-99m MIBI (10 mCi) was injected and SPECT myocardial imaging was performed. After 3 hours, 25 mCi Tc-99m MIBI was reinjected and rest images were obtained. A total of 320 ventricular wall segments were evaluated. Two hundred and six ventricular wall segments were supplied by stenotic coronary arteries and 114 segments were supplied by normal coronary arteries. Dipyridamole stress Tc 99m MIBI SPECT studies showed abnormal myocardial perfusion in 176 segments and normal perfusion in 144 segments. Transient regional wall motion abnormality was detected in 116 segments. A significant decrease in CVIBS after dipyridamole stress was detected in 184 segments. The sensitivity and specificity of dipyridamole stress echocardiography, Tc-99m MIBI SPECT, and CVIBS were 56% and 100%, 85% and 92%, and 89% and 100%, respectively, compared with the results from coronary angiography. Dipyridamole stress ultrasonic tissue characterization with CVIBS may provide more sensitive detection of myocardial ischemia than dipyridamole stress echocardiography and may be as valuable as dipyridamole stress myocardial perfusion scintigraphy. PMID- 15655270 TI - Association between angiotensin I-converting enzyme gene insertion/deletion polymorphism and risk of rheumatic heart disease. AB - Scarring and collagen deposition in the valves and destruction of myocytes may result from the combined effects of a smoldering rheumatic process and a constant trauma to the mitral valve or aortic valve by the turbulent flow in rheumatic heart disease (RHD). It has been suggested that angiotensin I-converting enzyme (ACE) may be responsible for the increased valvular fibrosis and calcification in the pathogenesis of RHD. However, the role of ACE genetic variant in RHD has not been studied among the Chinese population in Taiwan. Hence, a case-controlled study was carried out to investigate the possible relationship between the ACE gene insertion/deletion (I/D) and G2350A polymorphisms and RHD. A group of 115 patients with RHD documented by echocardiography and 100 age- and sex-matched normal control subjects were studied. ACE gene I/D and G2350A polymorphisms were identified by polymerase chain reaction-based restriction analysis. There was a significant difference in the distribution of ACE I/D genotypes (P = 0.02) and allelic frequencies (P = 0.04) between RHD cases and normal controls. An odds ratio for the risk of RHD associated with the ACE I/D II genotype was 2.12 (95% CI, 1.21-3.71). An odds ratio for the risk of RHD associated with the ACE I allele was 1.50 (95% CI, 1.02-2.21). The ACE G2350A polymorphism showed no association with RHD (P = 0.90). Further categorization of RHD patients into mitral valve disease and combined valve disease subgroups revealed no statistical difference in these gene polymorphisms when compared between the two subgroups. This study shows that patients with RHD have a higher frequency of ACE II genotype and I allele, which supports a role for ACE I/D gene polymorphisms in determining the risk of RHD in Taiwan Chinese. PMID- 15655271 TI - Comparison of intermittent with continuous simvastatin treatment in hypercholesterolemic patients with end stage renal failure. AB - Coronary artery disease is the most important cause of morbidity and mortality in patients with end-stage renal failure (RF). Hypercholesterolemia is an important risk factor for coronary heart disease. Patients with chronic renal failure (CRF) have difficulties in compliance with their care and treatment. Intermittent simvastatin treatment may help to increase compliance and can be a treatment alternative in patients with CRF at risk of coronary artery disease. We investigated the effects of simvastatin and compared intermittent with continuous simvastatin treatment in hypercholesterolamic patients with CRF. The study group included 40 of 422 CRF patients on dialysis in our clinic. The inclusion criterion was low density lipoprotein cholesterol (LDL-C) of 130 mg/dL or more. Twenty patients received simvastatin 10 mg/day (continuous group) and 20 patients received simvastatin 20 mg three times a week (only dialysis days- intermittent group) for four months. Nineteen patients served as controls and they were given a prescribed diet only. Total cholesterol (TC) and LDL-C decreased markedly in patients receiving intermittent and continuous simvastatin compared to controls. Continuous simvastatin decreased TC by 23% (P < 0.001) and LDL-C by 39% (P < 0.001). Intermittent simvastatin decreased TC by 26% (P < 0.001) and LDL-C by 40% (P < 0.001). The atherogenic index ratios in both the continuous and intermittent groups (TC/High density lipoprotein-cholesterol (HDL-C) and LDL-C/HDL-C) decreased significantly. There was no significant difference in patient compliance between the two groups. Intermittent simvastatin is as effective and reliable as continuous simvastatin treatment and can be an alternative treatment in hypercholesterolemic patients on dialysis. PMID- 15655272 TI - Effects of statins on circulating oxidized low-density lipoprotein in patients with hypercholesterolemia. AB - Recently, it has been reported that circulating oxidized low-density lipoprotein (Ox-LDL) might be a pivotal indicator for coronary artery disease and the severity of acute coronary syndromes. The purpose of this study was to investigate the effects of statins on Ox-LDL in patients with hypercholesterolemia. Sixteen patients with hypercholesterolemia were randomly assigned to 2 groups, one received 10 mg of pravastatin (n = 8) and the other received 20 mg of fluvastatin (n = 8). The plasma level of Ox-LDL was measured using a newly developed sandwich enzyme-linked immunosorbent assay (ELISA) method. There were no differences between the two groups in Ox-LDL, total cholesterol (TC), or LDL cholesterol (LDL-C) at the baseline. The reduction in Ox LDL in the fluvastatin group was significantly higher than that in the pravastatin group (47.5% versus 25.2%, P = 0.033). The reductions in TC and LDL-C did not differ between the two groups. CONCLUSION: The present study has shown for the first time that the level of circulating Ox-LDL was significantly decreased by treatment with statins. In addition, the lowering effect of statins on the circulating Ox-LDL was independent of their lipid-lowering effect. Fluvastatin was more effective than pravastatin with regard to decreasing the circulating Ox-LDL. PMID- 15655273 TI - The effects of atorvastatin treatment on the fibrinolytic system in dyslipidemic patients. AB - Statins have pleiotrophic effects related to the pathogenesis of atherosclerosis and thrombogenicity of the vessel wall beyond lipid lowering. The aim of the present study was to examine the effect of atorvastatin treatment on the fibrinolytic system in patients with dyslipidemia. The investigation was carried out on 41 dyslipidemic patients (21 males and 20 females) with a mean age of 53.8 years (range, 30-76). The patients were divided into subgroups according to their cholesterol and triglyceride levels as hypercholesterolemic (n = 26) and mixed type hyperlipidemic (n = 15) and their risk factors for coronary heart disease including age, sex, hypertension, obesity, smoking, and family history. The patients were started on atorvastatin 10 mg/day, and evaluated within 6-12 weeks to assess the changes in fibrinolytic parameters including global fibrinolytic capacity, plasminogen activator inhibitor type-1 and tissue plasminogen activator, and lipids. After successful lipid-lowering therapy, global fibrinolytic capacity (P = 0.003) and tissue plasminogen activator levels (P = 0.04) were found to be increased and plasminogen activator inhibitor type-1 levels (P = 0.02) decreased in dyslipidemic patients. Global fibrinolytic capacity levels increased (P < 0.001) and plasminogen activator inhibitor type-1 levels decreased (P = 0.01) in patients with hypercholesterolemia (n = 26). However, no significant changes were observed in fibrinolytic parameters in patients with mixed-type hyperlipidemia (n = 15). When the patients were separately evaluated according to risk factors, significant beneficial effects on the fibrinolytic system were observed, especially in patients without obesity and hypertension as well as in older patients and males. These findings suggest that atorvastatin treatment has a beneficial effect on the fibrinolytic system in patients with hypercholesterolemia, but not in patients with mixed-type hyperlipidemia. Further studies are needed to show whether higher doses and longer periods of lipid lowering treatment have beneficial effects in patients with mixed type hyperlipidemia and some risk factors. PMID- 15655274 TI - Concentrations of hepatocyte growth factor, basic fibroblast growth factor, and vascular endothelial growth factor in pericardial fluid and plasma. AB - Some angiogenic factors, including hepatocyte growth factor (HGF), basic fibroblast growth factor (bFGF), and vascular endothelial growth factor (VEGF), have been reported to promote angiogenesis and improve myocardial perfusion in experimental models of ischemic heart disease. These factors are produced in various tissues, including myocardium. We measured the concentrations of HGF, bFGF, and VEGF by enzyme-linked immunosorbent assay in plasma and in pericardial fluid sampled during open heart surgery (12 patients with ischemic heart disease and 17 with nonischemic heart disease). HGF levels were significantly higher in plasma than in pericardial fluid (12.0 +/- 1.8 versus 0.26 +/- 0.04 ng/mL, P < 0.0001). On the other hand, bFGF levels were significantly higher in pericardial fluid than in plasma (243.5 +/- 50.9 versus 49.6 +/- 7.8 pg/mL, P = 0.009). VEGF levels were not significantly different between pericardial fluid and plasma (47.2 +/- 17.6 versus 24.5 +/- 3.6 pg/mL, P = 0.23). Concentrations of angiogenic factors in pericardial fluid and in plasma were not significantly different between patients with ischemic and nonischemic heart disease. These results suggest that the production, secretion, and kinetics of HGF, bFGF, and VEGF are different. These angiogenic factors may have different pathophysiologic roles. PMID- 15655275 TI - Association between mitral annular calcification and stroke. AB - It remains controversial as to whether mitral annular calcification (MAC) is an independent predictor of stroke. The aim of this study was to investigate whether there is an association between the presence of MAC and stroke or whether MAC is one of the predictive factors of carotid atheroma and therefore is a secondary risk for stroke. Fifty-six patients who had MAC demonstrated by echocardiography underwent carotid artery duplex sonography and computed brain tomography with various causes were enrolled in the study. They were compared with 58 control patients without MAC. MAC was defined as a dense, localized, highly reflective area larger than 5 mm at the junction of the atrioventricular groove and posterior mitral valve leaflet. Carotid artery stenosis was defined as lumen diameter narrowing exceeding 60%. Cerebral ischemia was detected by spiral tomography and was classified as infarction and lacunae. A significant association was found between the presence of MAC and carotid atheroma (P = 0.011), MAC and hyperechogen plaque (P = 0.034), and MAC and stenosis (P = 0.008). There was an association between the presence of carotid atheroma and cerebral infarction (P = 0.007). Logistic regression analysis revealed hypertension and diabetes mellitus were independent risk factors (P = 0.030, P = 0.034, respectively) for developing carotid atheroma. MAC was an independent factor for carotid stenosis (P = 0.029). MAC may not be a significant causative factor for stroke, but may be a secondary risk factor. A significant association between the presence of MAC and carotid artery atherosclerotic disease may explain the high prevalence of stroke in patients with MAC. PMID- 15655276 TI - Down-regulation of connexin43 in early myocardial ischemia and protective effect by ischemic preconditioning in rat hearts in vivo. AB - Connexin 43 (Cx43), a primary component of gap junctions, contributes to intercellular electrochemical communication. Cx43 undergoes dephosphorylation in early ischemia. We examined whether Cx43 is degraded in association with dephosphorylation during early myocardial ischemia and whether ischemic preconditioning (IP) affects the degradation after rat coronary artery occlusion. Male Sprague-Dawley rats underwent coronary artery occlusion for 1, 2, or 3 hours, or for 1 hour following treatment either with a calcineurin inhibitor (cyclosporine A), proteasome inhibitor (PSI), or lysosomal inhibitor (E64c), or following IP alone or after protein kinase C (PKC) inhibitor (chelerythrine) pretreatment. The IP was afforded by three cycles of 3 minute ischemia and 5 minute reperfusion. A large portion of the phosphorylated Cx43 (pCx43) in the membrane fraction was dephosphorylated, while a small portion was degraded at 1 hour of ischemia. The effects of the inhibitors were dephosphorylation and degradation by calcineurin and proteasome/lysosome, respectively. IP suppressed the decrease in pCx43 and increase in dCx43, while only the former was inhibited by the PKC inhibitor chelerythrine. The Cx43 mRNA level was reduced at 3 hours, but not at 1 hour of ischemia, irrespective of IP. We believe that Cx43 is dephosphorylated and degraded in early ischemia, whereas Cx43 transcription was suppressed at a later phase of ischemia. PMID- 15655277 TI - Acute myocardial infarction after heroin injections. AB - Information concerning acute myocardial infarctions (AMI) after heroin injection is limited. Only one report has described the association between heroin injections and AMI in a young woman. AMI after heroin injection in a patient with a normal coronary angiogram has not been reported. We report a 38-year-old man who developed AMI after heroin injection. He is probably the first case of AMI with normal coronary artery angiograms associated with heroin abuse. The heroin induced toxic effect and/or coronary spasm are highly suspected to be the causes of the infarction episode. PMID- 15655278 TI - Acute myocardial infarction in a young man complicated with left ventricular thrombi. AB - Premature coronary artery disease is very rare and complication with thrombus formation in the left ventricle is rarer still. A 23-year-old man was admitted to hospital for recent acute myocardial infarction after being struck by a basketball eight days previously. Echocardiography identified two peduncle thrombi at the apex of the left ventricle, which were confirmed with computed tomography. The proximal left anterior descending coronary artery was totally occluded. Following two weeks of treatment with heparin and warfarin, the patient agreed to undergo a coronary artery bypass graft and thrombectomy. The ecchymosed tissue around the coronary artery implied that a trauma injury might have been the cause of the coronary artery disease in this case. This work reviews the pathophysiology and natural history of coronary artery disease in a case of very young myocardial infarction. PMID- 15655279 TI - Reel syndrome and pulsatile liver in a patient with a two-chamber pacemaker. AB - Twiddler's syndrome is characterized by coiling of the pacemaker lead due to the rotation of the pacemaker generator on its long axis. Reel syndrome is another form of Twiddler's syndrome. It occurs due to the rotation of the pacemaker generator on its transverse axis with subsequent coiling of the pacemaker leads around the pulse generator. In this article we describe a patient with a two chamber pacemaker who presented with sudden onset of abdominal pulsation and was subsequently diagnosed as Reel syndrome. To the best of our knowledge, this case is the first case of Reel syndrome that developed in a patient with a two-chamber pacemaker. PMID- 15655280 TI - False-positive exercise test secondary to Wolff-Parkinson-White syndrome in the absence of manifest preexcitation and disappearance of ST depressions after accessory pathway ablation. AB - Wolff-Parkinson-White syndrome with manifest preexcitation is a common cause of false-positive exercise test results. However, false-positive results are extremely rare without manifest preexcitation. We report a case with intermittent Wolff-Parkinson-White syndrome and exercise-induced marked ST depressions in the absence of preexcitation of the QRS complexes. His coronary arteries were normal on angiography and no ST changes were observed in the control exercise test after ablation of the accessory pathway. PMID- 15655281 TI - A case of the toxicity of pilsicainide hydrochloride with comparison of the serial serum pilsicainide levels and electrocardiographic findings. AB - We treated an 88-year-old man with aortic valvular stenosis/insufficiency and paroxysmal atrial fibrillation, who developed ventricular tachycardia due to pilsicainide toxicity. He was treated at the outpatient clinic of his local hospital, and was administered pilsicainide (100 mg/day) for atrial fibrillation. The electrocardiographic findings on admission to our hospital indicated wide QRS with frequent episodes of ventricular tachycardia. We diagnosed him as having pilsicainide toxicity because of a low cardiac output and renal dysfunction. His creatinine level was 2.4 mg/dL and the serum pilsicainide level was 2.42 microg/mL on admission. Fluid infusion and continuous hemodiafiltration were performed to achieve an early reduction in the serum pilsicainide level. His serum pilsicainide concentration was significantly decreased by these treatments, and the prolongation of the QTc and ventricular tachycardia improved in parallel to the decrease in the serum pilsicainide level. The changes in the serum pilsicainide level showed a significant positive correlation with the changes in the electrocardiographic findings (PQ, QRS, ST intervals, and QTc). Pilsicainide should be administered with great care to elderly patients, especially patients with cardiac dysfunction and renal dysfunction. Estimation of the serum level may be possible from the electrocardiographic findings if the pilsicainide toxicity occurs. PMID- 15655282 TI - Pulmonary and aortic valve endocarditis in an adult patient with silent patent ductus arteriosus. AB - Pulmonary and aortic valve endocarditis are uncommon especially in an adult patient with patent ductus arteriosus. A 27-year-old woman diagnosed with pulmonary and aortic valve endocarditis underwent surgical treatment. Here, we report our clinical and surgical experience in treating a case of double valve endocarditis with clinically silent patent ductus arteriosus. PMID- 15655283 TI - A case of deep venous thrombosis with a double inferior vena cava effectively treated by suprarenal filter implantation. AB - Embryogenesis of the inferior vena cava (IVC) is a complex process involving the formation and regression of several anastomoses, thus, various anomalies may occur. We report a case of deep venous thrombosis (DVT) accompanied by a double inferior vena cava (DIVC). A 76-year-old-man was admitted because of right leg edema and pain. Venography revealed two IVC and massive venous thrombus. To avoid massive pulmonary embolism (PE), it was necessary to block both the right and the left IVC. However, the right IVC was too small to implant the filter, so we placed a temporary IVC filter (Antheor filter) in the suprarenal portion of the IVC, after the confluence of the two IVC, and started thrombolytic and anticoagulant therapy. Venography, performed 6 days after filter implantation, showed a considerable amount of remaining thrombus. We replaced the Antheor filter with a Gunther retrievable filter because the former has a catheter and is not suited for long-term use, whereas the latter can be used permanently. Two weeks after filter exchange, thrombus had decreased but remained. We therefore did not remove the Gunther filter. The patient's symptoms gradually improved in response to anticoagulant therapy, and he was discharged with no complications. The present case illustrates the importance of a correct understanding of anatomy and demonstrates the effectiveness of using a suprarenal IVC filter in DVT. PMID- 15655284 TI - Cardiac wall motion abnormalities observed in a patient with transient hyperthyroidism. AB - A 74-year-old woman, with a history of hypertension and hyperlipidemia, was admitted to our hospital. She was found to have a sinus tachycardia with ST segment elevations in leads II, III, (a)V(F), and V(3) through V(6) in electrocardiography, hypokinesis of the left ventricular apex by echocardiography, and normal findings on coronary angiography. Blood analysis revealed an increase in the creatine kinase MB fraction, a significant positive detection in troponin T, and transient elevations in the concentrations of free triiodothyronine, free thyroxine, thyroid globulin antibody, and thyroid peroxidase antibody. Defects in myocardial perfusion and fatty acid metabolism in the apical area were also demonstrated by myocardial scintigraphy. These data suggest that tako-tsubo syndrome or myocardial infarction may be induced in patients with mild and transient hyperthyroidism. PMID- 15655285 TI - Monitoring expression of cytochrome P450 genes during postischemic rat liver reperfusion using DNA microarrays. AB - We investigated the expression of 50 cytochrome P450 (CYP) genes in the postischemic reperfused rat liver using a DNA microarray. Thirteen CYPs showed absent expression in all experiments and 2 CYPs were induced by pentobarbital, which was used as an anesthetic. Eight of the remaining 35 CYPs showed significantly decreased expression following ischemia-reperfusion. Monitoring CYP expression may be a powerful approach for elucidation of pathways regulating drug metabolism that may be involved in postischemic reperfusion liver injury. PMID- 15655286 TI - Induction of apoptosis by Korean medicine Gagam-whanglyun-haedoktang through activation of caspase-3 in human leukemia cell line, HL-60 cells. AB - Gagam-whanglyun-haedoktang (GWH) is a newly designed herbal drug formula based on the traditional oriental pharmacological knowledge for the purpose of treating tumorous diseases. Apoptosis is an evolutionarily conserved suicide program residing in cells. In the present study, apoptosis inducing activities of the decocted water extract of GWH were studied. Results of the 3-(4,5-dimethylthiazol 2-yl)-2,5-diphenyltetrazolium bromide assay showed that GWH had a strong cytotoxic effect on HL-60 cells. The number of live cells was less than 20% after exposure to 1 mg/ml GWH for 48 h. GWH increased cytotoxicity of HL-60 cells in a dose- and time-dependent manner. The percentage of apoptotic cells by flow cytometric analysis of the DNA-stained cells increased to 28%, 31%, and 37% at 24 h and to 37%, 44%, and 81% at 48 h after treatment with 0.01, 0.1, and 1 mg/ml GWH, respectively. DNA fragmentation also occurred in apoptosis and was characterized by a ladder pattern on agarose gel. In addition, GWH increased the secretion of tumor necrosis factor-alpha. GWH-induced apoptosis was accompanied by activation of caspase-3. These results suggest that GWH induces activation of caspase-3 and eventually leads to apoptosis. PMID- 15655287 TI - Increased excretion of urinary 20-HETE in rats with cyclosporine-induced nephrotoxicity. AB - The present study examined the contribution of 20-hydroxy-5,8,11,14 eicosatetraenoic acid (20-HETE) in cyclosporine A (CsA)-induced renal nephrotoxicity. Treatment of rats with CsA (50 mg/kg) for 9 days induced renal damage as indicated by marked increase in urine flow (from 9.0 +/- 0.3 ml/day to 46.6 +/- 7.1 ml/day) and a 3 - 5-fold rise in blood urea nitrogen (BUN) levels. The urinary excretion of 20-HETE increased from 164 +/- 5 ng/day (N = 5) to 2432 +/- 290 ng/day (N = 5, P<0.01) after 9 days of CsA treatment. The increase in the urinary excretion of 20-HETE in the CsA treated rats was highly correlated with the increase in BUN levels (r = 0.819, P<0.001) and urine volume (r = 0.832, P<0.001). Immunohistochemical examination of kidney revealed that expression of cytochrome P450 4A (CYP4A) protein was markedly enhanced in the proximal tubules of CsA-treated rats. These results indicate that CsA-induced nephrotoxicity in rats is associated with a marked elevation in the renal production of 20-HETE and that 20-HETE may contribute to the pathophysiological condition of CsA-induced nephrotoxicity. PMID- 15655288 TI - Effect of crude saponin of Korean red ginseng on high-fat diet-induced obesity in the rat. AB - The anti-obesity effects of crude saponin (CS) of Korean red ginseng (KRG) were investigated in the rat fed a high-fat (HF) diet. Male Sprague-Dawley (SD) rats became obese by feeding the HF diet over 5 weeks, while the control rats were fed a normal diet, and then both groups were treated with CS (200 mg/kg, i.p.) for 3 weeks. The body weight, food consumption, adipose tissues, and expression of appetite peptides such as leptin and neuropeptide Y (NPY) were investigated in rats fed normal and HF diet after treatment of CS. Administration of CS reduced body weight, food intake, and fat content in HF diet rats in a manner similar to those of the normal diet fed rats. The hypothalamic NPY expression and serum leptin level were reduced in HF diet rats after CS treatment. Our results suggest that CS may be useful in the treatment of obesity and related disorders as anti obesity agents. PMID- 15655289 TI - Exogenous histamine stimulates colorectal cancer implant growth via immunosuppression in mice. AB - Results from a limited number of studies suggest a potential role for endogenous histamine in regulating tumor growth in immunocompetent cells. The present study examined the effects of exogenous histamine on colorectal cancer growth and the immune response against tumor tissue in mice. Histamine was administered for 21 days to Colon 38 mouse colon adenocarcinoma-implanted syngeneic mice and tumor volume was measured throughout the experiment. Systemic administration of histamine for 21 days caused a significant increase in tumor implant growth compared with the vehicle. At the end of histamine administration, the interferon (IFN)-gamma / interleukin (IL)-4 ratio in peripheral lymphocytes, as well as histamine and cytokine levels in tumor implants were determined. Histamine levels in tumor implants remained unchanged after exogenous histamine delivery. Mice with tumor implants exhibited significantly elevated IFN-gamma / IL-4 ratios compared with mice lacking tumors. Nonetheless, the increased IFN-gamma / IL-4 ratios were markedly suppressed by histamine administration compared with vehicle. In addition, histamine delivery significantly decreased IFN-gamma and IL 12 mRNA expression, but increased IL-10 mRNA expression in tumor implants. It was concluded that exogenous histamine dysregulates the balance between T-helper 1 (Th1) and T-helper 2 (Th2) cells, attenuating anti-tumor cytokine expression in the tumor microenvironment, thus resulting in stimulated colorectal cancer growth. PMID- 15655290 TI - The antipsychotic and antiemetic drug prochlorperazine delays the ventricular repolarization of the in situ canine heart. AB - Electropharmacological effect of the antipsychotic and antiemetic drug prochlorperazine was assessed using the halothane-anesthetized in vivo canine model (n = 5). Up to 10 times higher than the clinically relevant doses of prochlorperazine (< or = 3 mg/kg, i.v.) did not induce cardiohemodynamic collapse in the model. Meanwhile, clinically relevant to supratherapeutic doses (0.3 - 3 mg/kg, i.v.) prolonged the ventricular repolarization period in a dose-related and reverse-use dependent manner that could become proarrhythmic substrates. Thus, caution has to be paid on the use of prochlorperazine particularly for patients with risks of the elevated plasma drug concentration, compromised cardiac repolarization, and/or frequent ventricular premature beats. PMID- 15655291 TI - Utilization of human liver microsomes to explain individual differences in paclitaxel metabolism by CYP2C8 and CYP3A4. AB - Paclitaxel is widely used for treatment of malignant tumors. Paclitaxel is metabolized by CYP2C8 and CYP3A4, and these enzymes are known to differ between individuals, although the details have not been clarified. Recent progress in pharmacogenetics has shown that genetic polymorphisms of metabolic enzymes are related to these individual differences. We investigated the effect of the polymorphisms on paclitaxel metabolism by analyzing metabolic activities of CYP2C8 and CYP3A4 and expressions of mRNA and protein. Production of 6alpha hydroxypaclitaxel, a metabolite of CYP2C8, was 2.3-fold larger than 3'-p hydroxypaclitaxel, a metabolite of CYP3A4. Significant inter-individual differences between these two enzyme activities were shown. The expressions of mRNA and protein levels correlated well with the enzyme activities, especially with CYP3A4. Although it was previously reported that CYP2C8*3 showed lower activity than the wild type, two subjects that had the CYP2C8*3 allele did not show lower activities in our study. Inter-individual differences in paclitaxel metabolism may be related to CYP2C8 and CYP3A4 mRNA expression. CYP2C8 is the primary metabolic pathway of paclitaxel, but there is a "shifting phenomenon" in the metabolic pathway of paclitaxel in the liver of some human subjects. PMID- 15655292 TI - Lafutidine-induced increase in intracellular ca(2+) concentrations in PC12 and endothelial cells. AB - Lafutidine, a histamine H(2) receptor antagonist, exerts gastroprotective effects in addition to gastric antisecretory activity. The gastrointestinal protective effects of lafutidine are mediated by capsaicin-sensitive neurons, where capsaicin excites neurons by opening a member of the transient receptor potential channel family (TRPV1). Since the effect of lafutidine on the intracellular Ca(2+) concentration ([Ca(2+)](i)) in cells has not been elucidated, we investigated the lafutidine response to [Ca(2+)](i) in rat pheochromocytoma PC12 and human endothelial cells. Lafutidine at pharmacological concentrations greater than 1 mM induced a sustained increase in [Ca(2+)](i) in the presence of extracellular CaCl(2) in PC12 cells, while capsaicin showed dual effects on [Ca(2+)](i) in PC12 cells, where it activated TRPV1 and inhibited store-operated Ca(2+) entry. The thapsigargin (an activator of store-operated Ca(2+) entry) induced increase in [Ca(2+)](i) in PC12 cells was inhibited by capsaicin and SKF96365, an inhibitor of store-operated Ca(2+) entry, and the lafutidine response was inhibited by capsaicin but not by SKF96365. In endothelial cells, lafutidine induced an increase in [Ca(2+)](i) in a SKF96365-insensitive manner. These results suggest that lafutidine stimulates Ca(2+) entry via the capsaicin sensitive pathway but not the SKF96365-sensitive pathway. The possible role of store-operated Ca(2+) entry induced by lafutidine on gastrointestinal function is also discussed. PMID- 15655293 TI - The different roles of 5-HT(2) and 5-HT(3) receptors on antinociceptive effect of paroxetine in chemical stimuli in mice. AB - Serotonin (5-HT) is known to be an important mediator in pain modulation. Some centrally acting agents, like selective serotonin reuptake inhibitors (SSRIs), modulate pain. Activation of the endogenous opioid mechanisms or potentiation of analgesic effect by serotonergic and/or noradrenergic pathways might be involved in antinociception of SSRIs. However, peripheral mechanisms of nociception are not clear. In this study, the antinociceptive effect of paroxetine, its interaction with the opioidergic system and serotonin receptors were tested using the writhing test in mice. Paroxetine (5, 10, 20 mg/kg) induced an antinociceptive effect following i.p. administration in writhing test. For the groups in which the antagonists were tested, the dose of paroxetine that caused a significant and equipotent analgesic effect similar to 0.5 mg/kg morphine was selected. Naloxone significantly antagonized the antinociceptive effects of both paroxetine and morphine in a similar pattern and magnitude. Ketanserin (5-HT(2) receptor antagonist) or ondansetron (5-HT(3)-receptor antagonist) alone did not alter the nociceptive action of acetic acid. While the antinociceptive effect of paroxetine was highly potentiated by ketanserin, ondansetron reduced that antinociception. In conclusion, our results indicate that the antinociceptive effect of paroxetine mainly depends on central opioidergic and serotonergic mechanisms. Peripheral serotonergic mechanisms/receptors may contribute to this antinociceptive effect, especially by 5-HT(3)-receptor subtypes. PMID- 15655294 TI - Transcription factor Nrf2 regulates promoter activity of mouse aldose reductase (AKR1B3) gene. AB - Transcription factor Nrf2 regulates gene expression of drug metabolizing enzymes such as glutathione S-transferase via the antioxidant response element, ARE. Aldose reductase (AR), a member of the aldo-keto reductase (AKR) superfamily, metabolizes various endogenous and exogenous aldehydes. The AR gene 5'-flanking region contains a multiple stress response region (MSRR) composed of two putative AREs (ARE1 and ARE2), an AP1 site, and a tonicity response element (TonE). As this region is highly conserved among species, we examined the involvement of Nrf2 in transcriptional regulation of the AR gene. beta-Naphthoflavone, an Nrf2 activator, elevated the level of AR mRNA in HepG2 cells and increased the promoter activity of the mouse AR (AKR1B3) gene. The promoter activity of the AKR1B3 gene, containing MSRR, was also augmented by overexpression of Nrf2. Deletion and mutation analyses indicated that both ARE1 and the AP1 site were essential for the responsiveness to Nrf2, while ARE2 was nonfunctional. The presence of an ARE1 binding protein complex was revealed by electrophoretic mobility shift assay. These findings indicate that Nrf2 regulates the AKR1B3 promoter activity via ARE1 and the AP1 site. PMID- 15655295 TI - Physiology and pathophysiology of proteinase-activated receptors (PARs): PAR-2 as a potential therapeutic target. AB - PAR-2 is the second member of the family of proteinase-activated receptors activated by trypsin, tryptase, and several other serine proteinases. In order to evaluate the therapeutic potential for PAR-2, we have performed studies on PAR-2 mediated signal transduction and investigated the effects of PAR-2 gene deficiency in disease models. In addition to the G-protein-coupled receptor mediated common signal transduction pathways, inositol 1,4,5-trisphosphate production and mobilization of Ca(2+), PAR-2 can also activate multiple kinase pathways, ERK, p38MAPK, JNK, and IKK, in a cell-type specific manner. The studies using PAR-2-gene-deficient mice highlighted critical roles of PAR-2 in progression of skin and joint inflammation. We also describe the development and evaluation of potent and metabolically stable PAR-2 agonists in multiple assay systems both in vitro and in vivo. The structure-activity relationship analysis indicated the improved potencies of furoylated peptides. Furthermore, the resistance of the furoylated peptide against aminopeptidase contributed to the highly potent and sustained effects of the peptide in vivo. These studies suggest the potential therapeutic importance of PAR-2 in inflammatory diseases. Also, the PAR-2-gene-deficient mice and the potent and metabolically stable agonists are shown to be useful tools for evaluating the potency of PAR-2 as a therapeutic target. PMID- 15655296 TI - Physiology and pathophysiology of proteinase-activated receptors (PARs): regulation of the expression of PARs. AB - The level of receptors expressed on the cell surface determines the cellular responsiveness to agonists. Proteinase-activated receptors (PARs) have been reported to be either up-regulated or down-regulated in response to various types of stimulation and pathological situations. In addition, the transcriptional regulation plays a major role in the alteration of the expression of PARs under pathological conditions, while post-translational mechanisms such as phosphorylation, arresting-binding, internalization, and lysosomal degradation, which desensitize activated PARs and terminate intracellular signaling, also play an important role in regulating the expression of PARs and the cellular responsiveness to the agonists. Elucidating the mechanisms related to the expression of PARs is a critical step to understand the pathophysiology of various diseases and establish new therapeutic strategies. However, the molecular mechanism regulating the expression of PARs still remains to be elucidated. This minireview discusses our current understanding of the mechanisms regulating the expression of PARs. The transcription factors and the regulatory elements in the promoter regions, and the proteins that interact with the receptors and thereby regulate their trafficking and desensitization are the main problems that need to be elucidated. PMID- 15655297 TI - Physiology and pathophysiology of proteinase-activated receptors (PARs): PAR-2 mediated proliferation of colon cancer cell. AB - Proteinase-activated receptor-2 (PAR-2) has been demonstrated to be highly expressed in the gastrointestinal tract. In the present minireview, we summarize the effects of PAR-1 and PAR-2 stimulation using their activating peptides and agonist proteinases on the calcium signaling and the cell proliferation in DLD-1 cell, a human colon cancer cell line. PAR-2 but not PAR-1 stimulation induced the enhancement of cell proliferation, whereas both PAR-1 and PAR-2 stimulation induced the transient increase in [Ca(2+)](i). PAR-2 stimulation induced the phosphorylation of MEK1/2 and ERK1/2, but PAR-1 stimulation did not. The inhibition of MEK1/2 by PD98059 completely abolished the proliferative response to PAR-2 stimulation. Thus, MEK-ERK activation plays major role in the PAR-2 mediated proliferative response. The coupling of PARs to calcium signaling and MEK-ERK activation may be independent, and varied dependent on cell types. PMID- 15655298 TI - Physiology and pathophysiology of proteinase-activated receptors (PARs): PARs in the respiratory system: cellular signaling and physiological/pathological roles. AB - Proteinase-activated receptors (PARs), a family of G protein-coupled receptors, are widely distributed in the mammalian body, playing a variety of physiological/pathophysiological roles. In the respiratory systems, PARs, particularly PAR-2 and PAR-1, are expressed in the epithelial and smooth muscle cells. In addition to the G(q/11)-mediated activation of the phospholipase C beta pathway, epithelial PAR activation causes prompt and/or delayed prostanoid formation, leading to airway smooth muscle relaxation and/or modulation of an inflammatory process. PAR-2 present in the epithelium and smooth muscle is considered primarily pro-inflammatory in the respiratory system, although PAR-2 may also be anti-inflammatory under certain conditions. In the lung epithelial cells, PAR-2 can also be activated by exogenous proteinases including house dust mite allergens, in addition to various possible endogenous agonist proteinases. Clinical evidence also suggests possible involvement of PARs, particularly PAR-2, in respiratory diseases. PARs thus appear to play critical roles in the respiratory systems, and the agonists/antagonists for PARs may serve as the novel therapeutic strategy for treatment of certain respiratory diseases including asthma. PMID- 15655299 TI - Physiology and pathophysiology of proteinase-activated receptors (PARs): role of tryptase/PAR-2 in vascular endothelial barrier function. AB - Proteinase-activated receptor-2 (PAR-2) plays important roles in a variety of pathophysiological functions, including inflammatory responses and nociception. In this minireview, we describe the role of PAR-2 in acute inflammatory responses in lungs associated with iodinated radiographic contrast medium (RCM). Intravenous injection of RCM to rats induces lung injury characterized by vascular hyperpermeability, edema, and respiratory depression. Nafamostat, which is found to be the most potent and specific tryptase inhibitor, prevents RCM induced lung injury. In cultured endothelial cells of human pulmonary artery and bovine aorta, RCM, when applied in combination with mast cells, disrupts barrier function evaluated by the permeability of Evans blue through a monolayer of cultured cells, which is blocked by nafamostat and mimicked by tryptase and PAR-2 activating peptide. The tryptase-induced barrier dysfunction is blocked completely by a phospholipase C inhibitor and partially inhibited by a IP(3) receptor blocker, protein kinase C inhibitor, or Rho kinase inhibitor. Morphological observations reveal the formation of actin stress fibers and disappearance of the intercellular meshwork structure of vascular endothelial cadherin after application of RCM or PAR-2 ligands. Therefore, the release of mast cell tryptase and subsequent activation of endothelial PAR-2 are involved in acute lung injury induced by RCM. PMID- 15655300 TI - Physiology and pathophysiology of proteinase-activated receptors (PARs): Preface. PMID- 15655301 TI - Amyloid pathology and protein kinase C (PKC): possible therapeutics effects of PKC activators. AB - Amyloid beta-protein (Abeta) is one of the most studied peptides in human neurodegenerative disorders. Although much has been learned about the biochemistry of this peptide, fundamental questions such as when and how the Abeta becomes pathologic remain unanswered. In this article we review the recent findings on the biology and pathology of Abeta and the role protein kinase C (PKC) plays in these processes. The potential neuroprotective role of PKC and the possible therapeutic effects of PKC activators in Alzheimer's disease (AD) will be discussed. Briefly, comments will be also addressed on the role of PKC in cell death and neurogenesis in AD. PMID- 15655302 TI - MaxiK channel roles in blood vessel relaxations induced by endothelium-derived relaxing factors and their molecular mechanisms. AB - The endothelium of blood vessels plays a crucial role in the regulation of blood flow by controlling mechanical functions of underlying vascular smooth muscle. The regulation by the endothelium of vascular smooth muscle relaxation and contraction is mainly achieved via the release of vasoactive substances upon stimulation with neurohumoural substances and physical stimuli. Nitric oxide (NO) and prostaglandin I2 (prostacyclin, PGI2) are representative endothelium-derived chemicals that exhibit powerful blood vessel relaxation. NO action involves activation of soluble guanylyl cyclase and PGI2 action is initiated by the stimulation of a cell-surface receptor (IP receptor, IPR) that is coupled with Gs protein-adenylyl cyclase cascade. Many studies on the mechanisms by which NO and PGI2 elicit blood vessel relaxation have highlighted a role of the large conductance, Ca2+-activated K+ (MaxiK, BKCa) channel in smooth muscle as their common downstream effector. Furthermore, their molecular mechanisms have been unravelled to include new routes different from the conventionally approved intracellular pathways. MaxiK channel might also serve as a target for endothelium-derived hyperpolarizing factor (EDHF), the non-NO, non-PGI2 endothelium-derived relaxing factor in some blood vessels. In this brief article, we review how MaxiK channel serves as an endothelium-vascular smooth muscle transducer to communicate the chemical signals generated in the endothelium to control blood vessel mechanical functions and discuss their molecular mechanisms. PMID- 15655303 TI - The role of RhoA-mediated Ca2+ sensitization of bronchial smooth muscle contraction in airway hyperresponsiveness. AB - Smooth muscle contraction is mediated by Ca2+-dependent and Ca2+-independent pathways. The latter Ca2+-independent pathway, termed Ca2+ sensitization, is mainly regulated by a monomeric GTP binding protein RhoA and its downstream target Rho-kinase. Recent studies suggest a possible involvement of augmented RhoA/Rho-kinase signaling in the elevated smooth muscle contraction in several human diseases. An increased bronchial smooth muscle contractility, which might be a major cause of the airway hyperresponsiveness that is a characteristic feature of asthmatics, has also been reported in bronchial asthma. Here, we will discuss the role of RhoA/Rho-kinase-mediated Ca2+ sensitization of bronchial smooth muscle contraction in the pathogenesis of airway hyperresponsiveness. Agonist-induced Ca2+ sensitization is also inherent in bronchial smooth muscle. Since the Ca2+ sensitization is sensitive to a RhoA inactivator, C3 exoenzyme, and a Rho-kinase inhibitor, Y-27632, the RhoA/Rho-kinase pathway is involved in the signaling. It is of interest that the RhoA/Rho-kinase-mediated Ca2+ sensitization of bronchial smooth muscle contraction is markedly augmented in experimental asthma. Moreover, Y-27632 relaxes the bronchospasm induced by contractile agonists and antigens in vivo. Y-27632 also has an ability to inhibit airway hyperresponsiveness induced by antigen challenge. Thus, the RhoA/Rho kinase pathway might be a potential target for the development of new treatments for asthma, especially in airway hyperresponsiveness. PMID- 15655304 TI - Effects of oral glucose intake on gastric myoelectrical activity and gastric emptying. AB - To investigate the effect of oral glucose intake on gastric motility, we measured gastric myoelectrical activity and gastric emptying on two test conditions: 1) glucose intake and 2) water intake in the same 10 healthy male volunteers (20 to 29 years old). Gastric motility was evaluated with cutaneous-recorded electrogastrography (EGG) for 30 min both on fasting and after glucose or water intake, while gastric emptying was measured using acetaminophen-absorption method. There were no significant changes in EGG dominant frequency after water intake, but the frequency increased significantly after glucose intake. A postprandial dip (i.e., a transient decrease in frequency immediately after the food intake) was observed in 3 subjects after water intake and in 8 subjects following glucose intake. The EGG power ratio was significantly larger after glucose than water intake, with delayed gastric emptying in the former case. These results suggest that glucose is one of the components responsible for postprandial gastric motility. PMID- 15655305 TI - Preservative solution for freeze-storage of surgically excised human colon to enable study of smooth muscle function in vitro. AB - We have compared the reactivity to carbachol and high potassium of circular smooth muscle isolated from segments of human colon which was freeze-stored in different preservative solutions for more than one month following surgical resection. Concentration-dependent contractions in response to carbachol were reduced in terms of both their sensitivity (pEC50) and reactivity (Emax), depending on the preservative solutions used. Similar reduction of reactivity to 100 mM KCl was also observed. The best responsiveness was shown when the tissue was freeze-stored in SFM101. It is concluded that the freeze-storage of surgically excised human colon in SFM101 or phosphate buffer solution for more than one month provided the best preservation of smooth muscle function for in vitro pharmacological examination. PMID- 15655306 TI - Effects of endogenous and exogenous nitric oxide on electrical responses of circular smooth muscle isolated from the guinea-pig stomach antrum. AB - The effects of endogenous and exogenous nitric oxide (NO) on electrical activity were investigated in circular smooth muscle preparations isolated from the guinea pig stomach antrum. The actions of endogenous NO were evaluated from the effects of inhibition of NO synthesis by N(omega)-nitro-L-arginine (nitroarginine), while those of exogenous NO were assessed from the effects of SIN-1, an NO donor. Antral circular smooth muscle generated slow potentials periodically at a frequency of about 1 cycle per min (cpm), and unitary potentials were also generated in a random fashion in the interval between slow potentials. Application of nitroarginine (10(-5) M) increased the frequency of slow potentials, with no significant alteration of the resting membrane potential and amplitude of slow potentials. Frequency analysis of unitary potentials revealed that nitroarginine also increased the spectral density at 0.01-1 Hz frequency. The refractory period for the generation of slow potentials evoked by depolarizing pulses was about 10 s, but was decreased to 6 s by nitroarginine. In the presence of nitroarginine, SIN-1 (10(-9)-10(-7) M) reduced the amplitude and frequency of slow potentials: low concentrations (<10(-8) M) reduced only the frequency of slow potentials, while higher concentrations (10(-8)-10(-7) M) reduced both the amplitude and frequency of slow potentials, in a concentration dependent manner, before abolishing the slow potentials. The power spectrum of the unitary potentials indicated that SIN-1 (>10(-8) M) reduced the spectral density at 0.01-1 Hz frequency. The refractory period for the generation of slow potentials was increased again to about 10 s by SIN-1. Thus, the excitatory effects of nitroarginine could be antagonized by SIN-1, suggesting that the inhibitory effects of endogenous NO are comparable to those of exogenous NO produced by SIN-1. The results also suggested that the effects of NO on smooth muscle are insignificant and NO selectively inhibits the activity of intramuscular interstitial cells of Cajal (ICC-IM). PMID- 15655307 TI - Effects of endothelin-1 on the membrane potential and slow waves in circular smooth muscle of rat gastric antrum. AB - Electrophysiological effects of endothelin-1 (ET-1) on circular smooth muscle of rat gastric antrum were investigated by using intracellular membrane potential recording techniques. ET-1 (10 nM) caused an initial hyperpolarization of the membrane which was followed by a sustained depolarization. ET-1 also increased the frequency but not the amplitude of slow waves. In the presence of the endothelin type A (ETA) receptor antagonist, BQ123 (1 microM), ET-1 (10 nM) depolarized the membrane and increased the frequency of slow waves, but without the initial hyperpolarization. The selective endothelin type B (ETB) receptor agonist, sarafotoxin S6c (10 nM), also depolarized the membrane and increased the frequency of slow waves. In the presence of the ETB receptor antagonist, BQ788 (1 microM), ET-1 (10 nM) hyperpolarized the membrane. However, in the presence of BQ788, ET-1 caused neither the depolarization nor the increase in the frequency of the slow waves. The ET-1-induced hyperpolarization was completely abolished by apamin (0.1 microM). In the presence of apamin, ET-1 depolarized the membrane and increased the frequency of slow waves. The ET-1-induced depolarization was significantly attenuated by 4,4'-diisothiocyanatostilbene-2,2'-disulphonic acid (DIDS, 0.3 mM). The increase of the frequency by ET-1 was observed both in the presence and absence of DIDS. These results suggest that, ET-1 hyperpolarizes the membrane by the activation of Ca2+-activated K+ channels via ETA receptors, and depolarizes the membrane by the activation of Ca2+-activated Cl- channels via ETB receptors. ET-1 also appears to increase the frequency of slow waves via ETB receptors, however this mechanism would seem to be independent of membrane depolarization. PMID- 15655308 TI - Oro-pharyngeal chemoreceptor activation induces gastric motor response in healthy volunteer subjects. AB - Previous study has revealed that water-responsive afferent neurons in the superior laryngeal nerve induced inhibition of motility in the proximal and distal stomach using anaesthetized rats. These gastric responses might facilitate the reservoir function of the stomach. To confirm the gastric responses discovered in rats also occur in humans, we evaluated gastric myoelectrical activities in healthy volunteer subjects during fluid intake using electrogastrography. Before human experiments, we recorded the myoelectrical activities in rats to evaluate the response induced by the administration of water into the larynx. A large deflection in the gastric myoelectrical activities was observed just after the administration of water in anesthetized rats. A similar large deflection was also observed just after voluntary swallowing of 20 ml water in humans. The swallowing of saliva did not induce such response. We further observed the gastric response during reflex swallowing elicited by slow infusion of isotonic saline, water or 0.05 M citric acid on to the posterior tongue. Infusion of water and citric acid but not 0.15 M saline induced significant changes in mean relative ratio of the response. These electrogastrographic responses induced by the infusion of liquids strongly suggest that the gastric motor response facilitates reservoir function of the stomach during liquid intake in humans as well as in rats. PMID- 15655309 TI - Studies on light sensitivity in butterflies. PMID- 15655310 TI - Systematic identification of signaling molecules in hydra. PMID- 15655311 TI - MI arrest and apoptosis in starfish oocytes. PMID- 15655312 TI - Studies on biological function and evolutionary origin of symbiotic microorganisms in insects. PMID- 15655313 TI - Different doses of dopamine have heterogeneous effects on cerebral hemodynamics and dopamine receptors in young rabbits as measured with near infrared spectroscopy. AB - BACKGROUND: Fluctuations in cerebral blood volume and cerebral oxygenation may be important in the pathogenesis of intraventricular hemorrhage and hypoxic-ischemic brain injury in the neonate. The cerebral hemodynamic response to dopamine infusion in premature infants is not well established. The newborn rabbit, a rather immature species at birth, is a suitable model for monitoring the physiological changes of the cerebral circulation. METHODS: The effect of dopamine upon cerebral hemodynamics and basal ganglia dopaminergic receptors were studied using four different dopamine doses. RESULTS: No significant changes in near infrared spectroscopy (NIRS) parameters were observed in the animals that received 0.5 (n = 5) and 1 microg/kg/min (n = 4) of dopamine intravenously. In contrast, in those animals that received dopamine at 5 microg/kg/min (n = 7) and 50 microg/kg/min (n = 7), there was a significant decrease in oxygenated hemoglobin. Moreover, this was accompanied by a significant increase in deoxygenated hemoglobin soon after drug infusion. Cerebral blood volume was increased in the group that received 5 microg/kg/min, but significantly decreased in the group that received 50 microg/kg/min. In both groups NIRS parameters returned to baseline values soon after stopping dopamine infusion. CONCLUSION: Despite evidence of a physiological response, we found no difference in the distribution of dopamine receptors between experimental and control animals. We therefore speculate that dopamine has an effect on the cerebrovasculature that could be mediated by factors other than changes in the basal ganglia dopamine receptors. PMID- 15655314 TI - Effect of two amino acid solutions on leucine turnover in preterm infants. AB - OBJECTIVE: To assess the effect of two different parenteral amino acid mixtures, Trophamine and Primene, on leucine turnover in preterm infants. METHOD: Leucine kinetics were measured with [5,5,5 D3]leucine tracer in 15 infants receiving Trophamine (group 'T') (mean birth weight 1,263 g) and 22 who received Primene (group 'P') (mean birth weight 1,336 g) during two study periods, within a few hours after birth but before introduction of parenteral amino acid solution, and again at postnatal day 7. The rate of appearance of leucine was calculated from the enrichment of alpha-ketoisocaproic acid in plasma. RESULTS: There were no significant differences in leucine turnover within a few hours after birth in the two groups. In the infants who received Primene leucine turnover on day 7 was significantly lower than in those who received Trophamine (269 +/- 43 vs. 335 +/- 27, p < 0.05). Despite a higher intake of leucine in the Trophamine group (108 +/ 10 vs. 77 +/- 8 micromol.kg(-1).h(-1)), leucine released from proteins at day 7 was higher in this group compared to Primene (227 +/- 27 vs. 192 +/- 42 micromol.kg(-1).h(-1)). CONCLUSIONS: Primene administration results in lower leucine released from proteins, an estimate of protein breakdown, than Trophamine in preterm infants. Increases in whole body leucine turnover in response to administration of i.v. amino acids is influenced by the composition of the amino acid mixture. The factors responsible for this difference remain to be elucidated. PMID- 15655315 TI - Controversies and issues in hemodiafiltration therapy. AB - Hemodiafiltration appears to be the most effective technique of renal replacement therapy but several drawbacks are not counterbalanced by significant advantages. Although optimal transfer for both small and middle molecules can be achieved, there is no difference in mortality risk between HDF and HD patients. The infusion of a large amount of dialysate containing residual acetate of 2-7 mmol/l could lead to impaired cardiac contractile functions and carbonyl stress whereas loss of amino acids and water-soluble vitamins along with high UF rate could lead to malnutrition. Moreover, as substitution fluid is prepared on-line, contaminated fluid could be inadvertently infused to patients. Stringent maintenance rules are required for the production of sterile and non-pyrogenic dialysis solutions. Finally, daily hemodialysis could be the most promising mode of renal replacement therapy since it leads to a more stable 'milieu interieur' than other techniques whatever the mode of solutes removal when performed three times a week. PMID- 15655316 TI - Clinical results of daily hemofiltration. AB - Various types of daily hemodialysis (HD) have been developed. However, a water treatment system is essential for HD, which markedly prevents daily home HD. Hemofiltration (HF) can be simply performed only with substitution fluid without a water treatment system. This advantage is applied to daily HF. To perform this procedure at home, a simple system is needed. A system in which a dialyzer was combined with a circuit as one cartridge with bag-type substitution fluid was developed by NxStage Inc. This system facilitates not only HF via a supply of 15 l of bag-type substitution fluid, but also automatic washing/priming of the circuit and returning which can be achieved by applying the current CAPD management system. This daily HF may contribute to routine home HD in the future. PMID- 15655317 TI - Kinetics and dose of daily hemofiltration. AB - Daily hemofiltration (D-HF) is a new treatment modality that shows unique solute removal characteristics and possibly provides high quality of life for patients with end-stage renal disease. We evaluated solute removal characteristics of D-HF for 5 patients by kinetic modeling analysis. Five patients treated with normal 4 hour x 3 times/week hemodialysis (HD) were switched to D-HF (2-hour x 6 times/week). Ultrafiltration rates (Q(F)) or small solute clearances ranged from 63 to 106 ml/min. All the pertinent kinetic parameters were determined from patients' physical data and the HD portion of the clinical measurements. The two compartment kinetic model predicted the concentration changes after switching from normal HD to D-HF. In all 5 patients, concentrations of small solutes such as urea-nitrogen (UN) increased whereas that of beta(2)-microglobulin (beta(2) MG) decreased after switching from normal HD to D-HF. Predicted solute concentrations and clinical measurements for UN and beta(2)-MG were in good agreement with average error less than 10%. The model predicted that Q(F) = 155 ml/min may be necessary for the time-averaged concentration (TAC) of UN to be unchanged. The model also predicted that the 7 times/week D-HF should not increase the pretreatment concentration of UN, expecting even much lower beta(2) MG concentration after switching from normal HD to D-HF. D-HF is superior to normal HD for removing larger solutes but may increase the TAC of small solutes. A 7-day treatment (7 times/week) D-HF may improve the solute removal capacity of small solutes. PMID- 15655318 TI - Ultrapure dialysis fluid--direct and indirect benefits in dialysis therapy. AB - The fluid quality description 'ultrapure' means practically free from bacteria and endotoxin. In quantitative terms it is defined as <0.1 CFU/ml and <0.03 EU/ml. The requirements on endotoxin as well as bacteria should be fulfilled, because these two entities are not strictly correlated. Ultrapure dialysis fluid can be prepared from standard quality fluid by a single step of controlled ultrafiltration. Recent clinical studies demonstrate that the use of ultrapure dialysis fluid in hemodialysis is associated with patient benefits indicating a less inflammatory state compared to hemodialysis with standard fluid. By applying one additional step of controlled ultrafiltration, ultrapure dialysis fluid can be further purified to such high microbiological quality that it can be used for infusion. This opens up the possibility for convective therapies, hemodiafiltration and hemofiltration, for which large volumes of sterile infusion solution are needed. With optimal application of these therapies, solute removal is enhanced, qualitatively as well as quantitatively, and fluid management is facilitated through improved hemodynamic stability. PMID- 15655319 TI - Use of ultrapure dialysate in reduction of chronic inflammation during hemodialysis. AB - Chronic inflammation contributes to the pathogenesis of several complications of hemodialysis therapy. It is thought that backfiltration of bacteria-derived contaminations during dialysis may induce a chronic inflammatory state. High sensitivity C-reactive protein (hs-CRP) is one of the tools which can take a hold on such a chronic inflammatory condition. We examined the effect of ultrapure dialysate which contributes to chronic inflammation with hs-CRP and tried to reduce endotoxin (ET) levels at the end of the dialysate from 70 EU/l to <1.0 EU/l (ultrapure dialysate). Other dialysis conditions, except ET level, were fixed. We investigated the hs-CRP of 23 patients receiving regular dialysis before the use of ultrapure dialysate and 1 year after use of it prospectively. The data showed a significant decrease in the median value of the hs-CRP from 0.16 to 0.07 mg/dl (p < 0.05). The value of serum beta(2)-microglobulin decreased from 33.2 to 28.4 mg/dl (p < 0.01) and the hemoglobin level increased from 10.0 to 11.0 g/dl (p < 0.05). These results indicate that even a dialysate containing 70 EU/l of ET level may induce a chronic inflammatory state. hs-CRP is a very useful marker of chronic inflammation and the use of ultrapure dialysate is necessary to improve a chronic inflammatory state. The targeted ET level at the end of the dialysate should be set at < or = 1.0 EU/l. PMID- 15655320 TI - Complementary dialysis for daily dialysis. AB - The greatest benefit of PD ist the continuous maintenance of a stable electrolyte and acid-base balance, as well as a continuous solute, sodium and water removal. Moreover, PD is effective in preserving residual renal function (RRF). However, when RRF declines, complications will occur if not compensated for by an increase in both the dialysis dose and ultrafiltration. Combination therapy PD and HD has been implemented to address the weakness of PD. The weak points of PD are declining UF volume and solute removal over time. The most important weak point of HD is the intermittent nature of the treatment. Based on this, the following concept was developed. At the start of dialysis, PD should be initiated in order to maintain RRF. In this stage, the function of PD is a complement to the RRF. When RRF declines, combination PD and HD should be considered. Subsequently, to achieve sufficient solute removal and ultrafiltration, the number of HD sessions should increase and PD prescription may decrease, resulting in three HD sessions per week and low-volume PD on non-HD days. Thus, PD complements HD in the end. If this integration can become a reality, we can accomplish 'real' daily dialysis. PMID- 15655321 TI - Preservation of renal function: HDF versus PD. The paramount role of peritoneal dialysis. PMID- 15655322 TI - Internal filtration-enhanced hemodialysis is a cost-effective treatment in view of solute removal. AB - By modifying dialyzer module design, internal filtration (IF) within a dialyzer is enhanced to increase convective solute transport. Thus, it can be an alternative to hemodiafiltration with no requirement of substitution fluid or additional complex machines. Cost-effective analysis was conducted in three modes of therapy: high-flux hemodialysis, on-line hemodiafiltration and IF-enhanced hemodialysis. In IF-enhanced hemodialysis, cost-effectiveness of small solute removal is comparable with high-flux hemodialysis and that of beta(2) microglobulin removal is the best. It is concluded that IF-enhanced hemodialysis is the most cost-effective therapy mode in comprehensive overall solute removal. PMID- 15655323 TI - On-line hemodiafiltration as routine treatment of end-stage renal failure: why pre- or mixed dilution mode is necessary in on-line hemodiafiltration today? AB - Hemodiafiltration (HDF) is a well-recognized treatment modality that offers a way of optimizing renal replacement therapy efficacy of end-stage renal disease (ESRD) patients. On-line production of substitution fluid by the 'cold sterilization' process (ultrafiltration) gives access to an unlimited amount of sterile and non-pyrogenic IV grade solution. This advantageous low-cost solution may therefore be employed to develop various forms of high-flux HDF modalities (ol-HDF). High-flux post-dilutional HDF (post-HDF) has mainly been used in clinical practice since it offers the most efficient and best compromise between diffusive and convective clearances. Nowadays, the new targets in anemia correction have created hemorheological conditions that render high filtration rate more difficult to achieve and/or at the expense of higher transmembrane pressure. To overcome this new challenging condition and keeping the same concept, it has been proposed to develop alternative modalities with various sites of fluid substitution (predilution, mixed pre-post with various percentages) in HDF. In this presentation we discuss the benefits of using pre HDF and show how to match performances with post-HDF. Potential advantages of new ol-HDF options (pre-, mixed and mid-dilution) that are advocated have to be demonstrated in clinical trials. On-line HDF is a multipurpose treatment method that is employed to improve care and outcomes of ESRD patients. Due to its versatility, ol-HDF should be considered as a technical platform permitting to personalize and tailor treatment to patients' needs. The mode of substitution (post-, pre-, mixed or mid-dilution) should be established according to hemorheological conditions of the individual patient. PMID- 15655324 TI - Selection of dilutional method for on-line HDF, pre- or post-dilution. AB - Post- and pre-dilution methods equally have advantages and disadvantages, therefore we should choose the more favorable one for clinical use. However, it still remains controversial which technique is better in on-line HDF. In post dilution, the clearances of small molecular uremic toxins increase as well as low molecular proteins, however the risk of albumin leakage caused by high transmembranous pressure (TMP) also increases. We can avoid the risk of albumin leakage because there are several maneuvers which aid in avoiding an increase of TMP, such as glucose infusion and gradual control of Q(f). On the other hand, the pre-dilution method is rather safe in the risk of albumin leakage but has an obvious clearance loss of small molecular substances caused by a decreased dialysate flow rate. The influence of microbial contamination and acetate are rather severe in pre-dilution on-line HDF. These lines of evidence suggest that post-dilution on-line HDF is the best choice for treating chronic HD patients. PMID- 15655325 TI - Preparation of ultrapure dialysate in Japan--clinical usefulness and short-term future. AB - In 1994, the water quality standard of dialysate and substitution fluid for on line HDF was established by the Kyushu Society for HDF. On the other hand, with the widespread use of high-flux membrane, reverse filtration and reverse diffusion became evident, and purification of the dialysate has become essential even for usual hemodialysis. By using ultrapure dialysate, new blood purification methods can be performed, such as internal filtration-enhanced hemodialysis, on line HDF, and on-line HF. As a result, various clinical effects have been reported, such as improvement in anemia and in chronic inflammatory reactions. Suppression of complications involving long-term dialysis is expected, and even the prolongation of life expectancy. By using ultrapure dialysate as substitution fluid for saline solution, a fully automated dialysis machine has been developed. Furthermore, if fully automated consoles can be made smaller in size, they will contribute to the widespread use of home hemodialysis. PMID- 15655326 TI - New trends in HDF therapies: validity of internal filtration-enhanced hemodialysis. AB - Internal filtration-enhanced hemodialysis (IFEHD), defined as HD therapy using a dialyzer designed for enhanced internal filtration, seems to be more convenient in comparison with HDF therapies. In this paper, the validity of IFEHD was discussed during an analytical study and experimental studies. As a result, the maximum internal filtration flow rate (Q(IF)) value increased with a smaller inner diameter (D), a longer effective length (L(eff)), and a larger density ratio (DR) value of the hollow fibers. The validity of IFEHD was clarified by an analytical study and experimental studies. Development of a dialyzer with enhanced internal filtration, however, should take account of the patient's safety, and hemolysis and endotoxin invasion form the dialysate to the patient should be avoided. PMID- 15655327 TI - Clinical benefit of preserving residual renal function in patients after initiation of dialysis. AB - Preserving residual renal function (RRF) after initiation of dialysis therapy is desirable for improving quality of life in ESRD patients. It has been believed that RRF declines more slowly in patients receiving continuous ambulatory peritoneal dialysis (CAPD) than in patients treated with other forms of maintenance dialysis. Episodes of intravascular volume depletion might be expected to cause more rapid loss of RRF, and are more frequent in patients on hemodialysis, which is intermittent therapy. However, recently it was demonstrated that in hemodialysis using high-flux biocompatible membrane and ultrapure water, RRF declines at a rate indistinguishable from that in CAPD. The HDF using ultrapure dialysate and the substitution fluid may show to preserve RRF as well as CAPD patients. In future, it might be a major concern for the assessment of the HDF by a multicenter clinical study for preserving RRF. PMID- 15655328 TI - Effect of hemodiafiltration against radical stress in the course of blood purification. AB - The involvement of radical stress has been suggested as a cause for complications in patients on dialysis, such as arteriosclerosis, dialysis-related amyloidosis, etc. It has been reported that the increase in radical stress is not only seen in renal failure, but that its amplified effect is also seen in the process of blood purification. Our group has reported on the radical stress-reducing effect of HDF. We performed four types of blood purification (HD; on-line HDF; pre, on-line HDF; post, P/P HDF) in patients on maintenance dialysis using the polysulfone (APS) dialyzer. The change in radical related markers such as pentosidine (total, free) and CML (total, free), and the CTL/Cr ratio, and the hydroperoxide radicals were studied. In HDF (post, pre), the amplification rate of hydroperoxide radicals was significantly low, whereas the reduction rate of CTL/Cr ratio as index for hydroxy radicals was significantly higher in on-line HDF than in HD. Both the total CML and T-pentosidine increased in HD but showed a decrease in HDF. As HDF uses large amounts of replacement solution, the following effects can be expected: (a) suppression of the amplification of hydroperoxide radicals and suppression of the amplification of hydroxy radicals, and (b) suppression of fat oxidation by AGEs themselves. These antiradical stress effects are presumed to be exerted by effective removal of radical carrier protein, denatured protein, and complement protein in HDF, by dilution of radicals by massive use of replacement solution, and by the sequential reduction of the excitation and amplification effects. PMID- 15655329 TI - 'Clean dialysate' requires not only lower levels of endotoxin but also sterility of dilution water. AB - Clean dialysate should be used in dialysis with a high-flux dialysis membrane to avoid contamination of endotoxin into blood circuits. For this purpose, we should not only clean up the end stream dialysate by endotoxin-cut filters, but also prevent bacterial growth in dilution water lines. Delivery lines of the prepared dialysate from a central dialysate-supplying machine are sterilized in all dialysis facilities, but those of the dilution water in cases of using personal dialysis machines are not sterilized in most facilities in Japan. In our dialysis department, the endotoxin level in the dilution water lines decreased to <50 EU/l in 3 weeks by sterilization with a low concentration of sodium hypochlorite (30 100 ppm) once a week from the peak level of >1,000 EU/l just after the start of sterilization. PMID- 15655330 TI - Effects of S1P on myoblastic cell contraction: possible involvement of Ca independent mechanisms. AB - Sphingosine-1-phosphate (S1P) is a lipid mediator, which affects many essential processes such as cell proliferation, differentiation and contraction in many cell types. We have previously demonstrated that the lipid mediator elicits Ca(2+) transients in a myoblastic cell line (C2C12) by interacting with its specific receptors (S1PR(s)). In the present study, we wanted to correlate the Ca(2+) response with activation of myoblastic cell contractility. C2C12 cells were first investigated for the expression and cellular organization of cytoskeletal proteins by immunoconfocal microscopy. We found that myoblasts exhibited a quite immature cytoskeleton, with filamentous actin dispersed as a web-like structure within the cytoplasm. To evaluate intracellular Ca(2+) mobilization, the cells were loaded with a fluorescent Ca(2+) indicator (Fluo-3), stimulated with S1P and simultaneously observed with differential interference contrast and fluorescence optics. Exogenous S1P-induced myoblastic cell contraction was temporally unrelated to S1P-induced intracellular Ca(2+) increase; cell contraction occurred within 5-8 s from stimulation, whereas intracellular Ca(2+) increase was evident only after 15-25 s. To support the Ca(2+) independence of myoblastic cell contraction, the cells were pretreated with a Ca(2+) chelator, BAPTA/AM, prior to stimulation with S1P. In these experimental conditions, the myoblasts were still able to contract, whereas the S1P-induced Ca(2+) transients were completely abolished. On the contrary, when C2C12 cells were induced to differentiate into skeletal myotubes, they responded to S1P with a rapid cell contraction concurrent with an increase in the intracellular Ca(2+). These data suggest that Ca(2+)-independent mechanism of cell contraction may be replaced by Ca(2+)-dependent ones during skeletal muscle differentiation. PMID- 15655331 TI - Morphometric quantification of apoptotic stages in cell culture. AB - Apoptosis is an active process of self-destruction, whereby cells undergo physiological cell death. It occurs during development and regulation of tissue homeostasis or as a result of changes in environmental stimuli. Chromatin condensation and nuclear fragmentation, which are typical features of apoptotic nuclei, are usually quantified by fluorescent DNA dyes. The present study reports a reliable method to analyze morphological apoptotic stages in cultured cells, using light microscopy. We used the human neuroblastoma cell line SK-N-BE as a model to study apoptosis induced by inadequate cell-matrix interactions. Apoptosis was detected on cells cultured for different time intervals on polyHEMA, poly-L-lysine or collagen I. Quantitative morphometric and densitometric analysis after hematoxylin nuclear staining and caspase-3 immunocytochemistry, as markers of occurring apoptosis, were performed. Our method identifies different stages of caspase-3 activation and the subsequent DNA fragmentation and condensation. This experimental procedure enables us to detect slight differences in apoptosis progression by morphological analysis. PMID- 15655332 TI - Collagen type I increases bone remodelling around hydroxyapatite implants in the rat tibia. AB - The early interface reaction of cancellous bone to a nanocrystalline hydroxyapatite (HA) cement containing 3 wt% collagen type I (HA/Coll) with a setting under physiological temperature and pH was observed using immunohistochemical techniques. Pure HA served as a control. Cylinders with a diameter of 2 mm were implanted into the proximal tibia of 72 adult Wistar rats. Histological sections of 6 animals were prepared after 1, 2, 4, 6, 14 and 28 days. First, osteoblast-like cells as well as a marked reaction for osteonectin, osteopontin and its ligand CD44 were observed as early as 2 days after implantation at the interface around HA/Coll implants. Further, reactivity for ED1 and cathepsin D, both markers for phagocytotic cells, appeared earlier and stronger around HA/Coll. In cell counts, a significantly higher average number of ED1- and cathepsin D-positive phagocytotic cells was observed around the HA/Coll implants on days 6 (p < 0.01), 14 and 28 (p < 0.05). The number of osteopontin positive cells was significantly higher around HA/Coll implants at days 6 and 14 (p < 0.05). Two weeks after the implantation, first islands of newly formed woven bone were observed around the HA/Coll implant, but not around the control implant. The amount of direct bone contact after 28 days averaged 28% around pure HA and 51% around HA/Coll implants (p < 0.05). While both implants displayed a good osteoconductivity, a higher bone remodelling activity was observed around collagen-containing HA implants compared to pure HA implants. It appears that the addition of collagen to HA implants can enhance both phagocytotic and osteogenic processes. This may result in an earlier acceptance and better osseointegration of the HA/Coll implants into the surrounding tissue. PMID- 15655333 TI - Osteopontin distribution in the canine skeleton during growth and structural maturation. AB - The distribution of osteopontin (OPN) was studied immunohistochemically in cells and extracellular matrix in the humerus, scapula, and lumbar vertebrae of growing (age: 6 weeks, 12 weeks, 4.5 months) and adult dogs. OPN was expressed in hypertrophic chondrocytes of epiphyseal cartilage and in chondrocytes of the deep zone of mature articular cartilage, where extracellular matrix was also stained. OPN expression was strongest in 4.5-month-old puppies in cells of the osteoblastic lineage. It also varied with microlocation and was pronounced in areas prone to resorption due to modelling and remodelling activities. Osteoclasts were always strongly labelled with OPN. OPN deposition in extracellular bone matrix was detected particularly as a delineation of cartilage cores within secondary trabeculae and as a lining of the trabecular surfaces in resorption microlocations. The OPN distribution pattern is discussed here for each cell population with regard to its functional implications. PMID- 15655334 TI - Altered ATP-sensitive P2 receptor subtype expression in the Han:SPRD cy/+ rat, a model of autosomal dominant polycystic kidney disease. AB - The effects of extracellular ATP on fluid secretion and reabsorption by renal epithelial cells, as well as its known effects on cell proliferation and death, are potentially important contributory factors in the development and growth of renal cysts. In this study, we have investigated the protein and mRNA expression of several P2Y receptor subtypes (P2Y(1,2,4,6)), as well as the P2X(5) and P2X(7) receptors, in kidney tissue from the Han:SPRD (cy/+) rat model of polycystic kidney disease. All of the P2Y receptors tested for, and the P2X(5) and P2X(7) subtypes, were located on the cyst-lining cells of Han:SPRD (cy/+) rat polycystic kidneys; most immunostaining was cytosolic and we could not confidently localize it to one or other membrane. However, the staining pattern for P2Y(6) was uniquely granular when compared with the other P2 receptors. P2Y(2) and P2Y(6) receptor mRNA was increased in both homozygote (cy/cy) and heterozygote (cy/+) rat kidneys when compared with unaffected littermates. The protein levels of P2Y(2) and P2Y(6) receptors were also increased, being undetectable or at a low level, respectively, in control tissue. Finally, P2X(7) receptor mRNA was increased in cy/+, but not in cy/cy rat kidneys. Our results show that a number of P2Y receptor subtypes, as well as the P2X(5) and P2X(7) receptors, are clearly expressed in cyst-lining cells in the Han:SPRD (cy/+) rat model of renal cystic disease. Furthermore, P2Y(2) and P2Y(6) receptor mRNA and protein levels are markedly increased in cystic rat kidneys compared with normal rats of the same genetic background. Thus, the most consistent findings were an increase in the expression of P2Y(2), P2Y(6) and P2X(7) receptors in cystic tissue. Given the widely reported effects of stimulating these P2 receptor subtypes in epithelial and other renal cells, they could contribute to the development and growth of renal cysts: extracellular ATP and its products 'trapped' in cyst fluid may activate P2 receptors expressed by cyst-lining cells, causing cyst expansion from increased fluid secretion and/or reduced reabsorption, as well as an increase in cell turnover (re-modeling). PMID- 15655335 TI - Schlemm's canal and the collector channels at different developmental stages in the human eye. AB - We studied Schlemm's canal (SC) and other posttrabecular structures of aqueous humor outflow in eyes from human fetuses (at 24, 26, 33 and 36 weeks of gestation), eyes from normal children (2 months and 8 years old) and normal adult eyes, using mesoscopy/diaphanization and light microscopy. This study points to the presence of a collector canal parallel to the outer wall of SC (the outer collector). At 24 weeks of gestation, the SC was observable throughout the entire circumference (360 degrees) and the anlage of the outer collector was present. At 26 weeks of gestation, the outer collector was still growing. At 33 weeks of gestation, the outer collector was parallel to the SC and had become visible in some sectors. At 36 weeks, the SC and the outer collector were clearly defined and connected by intercanal links. The intrascleral plexus was growing. At 2 months, the deep intrascleral plexus and the episcleral plexus continued to grow, developing by sectors. At 8 years of age, the same structures corresponded to those in adults. The outer collector and the intrascleral plexuses were divided into sectors not greater than 120 degrees. PMID- 15655336 TI - Shortening of the front vibrating part of the vocal folds in phonation. AB - We have tried to make use of the opportunity of repeated observations through the analysis of the recorded movements of the phonating vocal folds by means of the phonating vocal folds through indirect videolaryngostroboscopic technique. The results of our investigation correspond, among others, with Hala and Sovak's diagrammatic description of continual and gradual changes in the vibration of the vocal folds in an uninterrupted sequence of tones of the human voice. In the initial chapter mention has been made of Hala and Honty's works and of Pressmann's work dealing with the described behaviour of the glottis when phonating high-pitched tones. The conclusions analogous to our findings have also been described by means of stereoscopically measuring the length of the phonating vocal folds. In conclusion, by measuring the distances between the anterior commissure and the junction between the membranous part of the vocal fold and vocal process of the arytenoid and the anterior commissure to the posterior nodal point of the vibrating part of the vocal folds we have verified the correctness of the findings published by Czermak, Hala and Honty, Pressmann, and Sovak, who pointed out the phenomenon of the shortening of the front vibrating part of the vocal folds in phonating high-pitched tones of the human voice. PMID- 15655338 TI - Anxiety, depression, and quality of life in mothers of children with cleft lip/palate. AB - Multidisciplinary care for patients with cleft lip and palate (CLP) includes the surgical correction of the facial disfigurement and the rehabilitation of functional deficits to optimize the communicational competence of the affected patient. Although the well-being of the mother of a CLP child is an important protective factor for the child, up to now literature has paid only little attention to this topic. In this study, 50 mothers of CLP children aged 1-10 years were examined using the 36-item Short-Form Health Survey and the Hospital Anxiety and Depression Scale (HADS) to screen for quality of life, anxiety and depression. Surprisingly, the data obtained indicate no group-specific alterations of these items when compared with normal controls. In cases with a deteriorated quality of life, the results of the HADS were pathologic as well. In general, mothers of CLP children aged more than 12 months do not require psychological screening. In individual cases, screening for relevant emotional aspects may be restricted to using the HADS. PMID- 15655337 TI - Features of listeners affecting the perceptions of Mandarin electrolaryngeal speech. AB - The effect of age and gender of listeners on the perceptions of Mandarin electrolaryngeal speech was investigated. Sixty males and 40 females were categorized into five age groups (20-29, 30-39, 40-49, 50-59, and 60-70 years), who were regarded as naive listeners for having no experience with electrolaryngeal speech. They were instructed to score acceptability of a passage and intelligibility of isolated words and embedded words. The results revealed no gender effects but significant age effects on the perceptual evaluation. It was more difficult for the 50-59 and 60-70 groups to understand electrolaryngeal speech. The results were also analyzed for tonal and segmental errors, and errors of tone alone were found to occur more often than segmental errors. In addition, a preliminary study was presented for the perceptions of the four Mandarin tones. Higher percent correct identification was found for the high-level tone compared to the other three tones. PMID- 15655339 TI - Task-based profile of vocal intensity decline in Parkinson's disease. AB - This study examines intensity decay in the phonation of persons with Parkinson's disease (PD). The decline in vocal intensity (determined by linear regression of the intensity envelope) was compared across the following tasks: vowel prolongation, syllable repetition (diadochokinesis, DDK), isolated sentences and conversation. In contrast to previous studies, PD speakers demonstrated no significant differences in intensity decline from healthy speakers in vowel prolongation. The vocal intensity of speakers with PD declined more rapidly than that of controls in DDK tasks. While intensity slopes in conversation were more variable in both groups, some participants with PD exhibited abrupt changes in intensity. Results indicate that the DDK is particularly useful for describing intensity decay associated with PD. However, considering the inconsistent group differences across tasks, and the discrepant findings from previous studies, intensity decay is not a robust symptom of PD. PMID- 15655340 TI - Suitability of acoustic perturbation measures in analysing periodic and nearly periodic voice signals. AB - In recent years, acoustic perturbation measurement has gained clinical and research popularity due to the ease of availability of commercial acoustic analysing software packages in the market. However, because the measurement itself depends critically on the accuracy of frequency tracking from the voice signal, researchers argue that perturbation measures are not suitable for analysing dysphonic voice samples, which are aperiodic in nature. This study compares the fundamental frequency, relative amplitude perturbation, shimmer percent and noise-to-harmonic ratio between a group of dysphonic and non dysphonic subjects. One hundred and twelve dysphonic subjects (93 females and 19 males) and 41 non-dysphonic subjects (35 females and 6 males) participated in the study. All the 153 voice samples were categorized into type I (periodic or nearly periodic), type II (signals with subharmonic frequencies that approach the fundamental frequency) and type III (aperiodic) signals. Only the type I (periodic and nearly periodic) voice signals were acoustically analysed for perturbation measures. Results revealed that the dysphonic female group presented significantly lower fundamental frequency, significantly higher relative amplitude perturbation and shimmer percent values than the non-dysphonic female group. However, none of these three perturbation measures were able to differentiate between male dysphonic and male non-dysphonic subjects. The noise to-harmonic ratio failed to differentiate between the dysphonic and non-dysphonic voices for both gender groups. These results question the sensitivity of acoustic perturbation measures in detecting dysphonia and suggest that contemporary acoustic perturbation measures are not suitable for analysing dysphonic voice signals, which are even nearly periodic. PMID- 15655341 TI - [Development and testing of the Heidelberg preschool screening for auditory perception and speech processing (HVS)]. AB - There is a lack of standardized and quick tests assessing auditory perception and language processing in preschool children in German. Therefore we developed a screening for auditory perception and speech and language processing, analyzed it statistically and evaluated it in 180 children from 15 preschools. The screening comprises 7 tasks: auditory short-term memory, phonological analyses of word initial sounds, syllable segmentation, phonological discrimination,articulatory sequencing, semantic versus phonological classification, and rhyming. Results indicated objectivity and reliability of the test. The comparison of subgroups showed that children having conductive hearing loss, deficits of articulation, deficits of language, and children otherwise considered to be unsuitable for school enrollment all showed significantly poorer achievements in different subtests than children without these problems. PMID- 15655342 TI - Cell cycle-dependent regulation of double-strand break repair: a role for the CDK. AB - DNA Double-Strand Breaks (DSBs) are dangerous lesions that can lead to genomic instability and to cell death. Eukaryotic cells repair DSBs either by nonhomologous end joining (NHEJ) or by homologous recombination (HR). Recent work has allowed to study the ability of yeast cells to repair a single, chromosomal DSB, at different stages of the cell cycle. Yeast cells repair the broken chromosome during the G(1) stage only by NHEJ, whereas HR is the mechanism of choice during the rest of the cell cycle. HR does not require duplicated chromatids or passage through S-phase. Control over the fate of the broken chromosome is exerted by Clb-CDK activity, which is required to carry out the first step of HR, ssDNA resection. Similar results in other organisms suggest that this control is a conserved feature in all eukaryotes. PMID- 15655343 TI - Development and gene expression profiling of a metastatic variant of the human breast cancer MDA-MB-435 cells. AB - We have developed a model system of late stage metastatic progression by isolating a highly malignant variant of human breast cancer cells from the parental MDA-MB-435 cell line. These cells, isolated from early lung metastasis, displayed increased anchorage independent growth in vitro and when transplanted ortho-topically into nude mice showed accelerated tumor growth rate and early lung spontaneous metastasis when compared to its parental counterpart. These cells, designated as MDA-MB-435-F-L, also showed intense wide spread early skeletal metastasis in vertebrae, mandible, femur, tibia and skull as detected by fluorescence imaging in an experimental bone metastasis model. Gene expression profiles from cDNA microarray showed up or downregulation of the expression of several significant genes regulating angiogenesis, apoptosis, ECM remodeling and metastasis in the MDA-MB-435-F-L cells in comparison to the parental cells. Among the up or downregulated genes, some have also been implicated in the survival of breast cancer patients. As such, the candidate genes selected in this breast cancer progression model system may serve as biomarkers of metastatic progression and also as potential tumor targets for breast cancer therapy. PMID- 15655344 TI - IL-6 signaling promotes tumor growth in colorectal cancer. AB - Recent investigations support an important role for TGF-beta in the development of colorectal cancer. However, the molecular consequences of TGF-beta signaling in the colon remains incompletely understood. In a recent study in Immunity, we analyzed the role of TGF-beta in a murine model of colon cancer. Using transgenic mice overexpressing TGF-beta or a dominant negative TGF-beta receptor II under control of the CD2 minigene, we show that TGF-beta signaling in tumor infiltrating T lymphocytes regulates the growth of dysplastic colon epithelial cells, as determined by histology and a novel system for high resolution chromoendoscopy in vivo. At the molecular level, TGF-beta signaling in T cells regulated STAT-3 activation in tumor cells via IL-6. IL-6 signaling required tumor cell derived soluble IL-6R rather than membrane bound IL-6R and suppression of such TGF-beta-dependent IL-6 trans-signaling prevented tumor progression in vivo. Similar to these observations in mice, here we show that human colon cancer tissue expressed only low amounts of membrane bound IL-6R. In contrast, expression and activity of the matrix metalloproteinase TACE were increased. In summary, our data provide novel insights into the role of TGF-beta signaling in colorectal cancer and suggest novel therapeutic approaches for colorectal cancer based on an inhibition of TGF-beta-dependent IL-6 trans-signaling. PMID- 15655345 TI - How cancer could be cured by 2015. AB - As announced by Andrew von Eschenbach, the NCI has set the goal of eliminating suffering and death due to cancer by 2015. Supporting this prediction, I discuss that cancer might be controlled and even cured by combining three potential therapeutic strategies aimed at (i) cancer-specific targets, (ii) universally vital targets with selective protection of normal cells (the selective combinations) and (iii) tissue-specific targets. Although (i) targeting cancer specific pathways (e.g., by imatinib and gefitinib) is probable, it alone will not be sufficient to control cancer. This strategy is limited to oncogene (kinase)-dependent cancers and is further limited by therapy-induced resistance and tumor progression. Thus, targeting cancer-specific pathways needs to be complemented by two divergent therapeutic strategies: (ii) selective combinations and (iii) tissue-selective therapy. With selective protection of normal cells (based on cell cycle and apoptosis manipulation), combinations of selective and chemotherapeutic drugs can be effective in most common cancers. Alternatively, tissue-selective therapy can suppress cancer cells in a tissue-selective manner, sparing other tissues. While alone, each therapeutic strategy may cause drug resistance and even tumor progression; these obstacles can be overcome and even exploited by using all three strategies in sequence. And finally, these strategies will benefit from molecular diagnostics and can be used for chemoprevention. PMID- 15655346 TI - Epigenetic plasticity of hematopoietic cells. AB - In recent years significant evidence was provided for the concept that the developmental potential is engraved in the chromatin of stem cells. This is indicated by low-level expression of lineage specific genes and also by epigenetic alterations that occur prior to gene locus activation. Hence, cell lineage specification involves not only the activation, but also the epigenetic silencing of different genetic programmes. In this article, I summarize recent data from my laboratory that indicate that (i) at the epigenetic level developmental processes occur in a step-wise fashion and (ii) that developmental windows exist, which are associated with a specific chromatin structure, in which such decisions can be reversed. PMID- 15655347 TI - Direct role of PDGF-BB in lymphangiogenesis and lymphatic metastasis. AB - Genetic instability of tumor cells often leads to amplified expression of multiple growth factors that contribute to angiogenesis and tumor growth. Members of the platelet-derived growth factor (PDGF) family are frequently utilized growth factors by many tumors to support their growth. PDGFs have previously been found to induce tumor growth by directly stimulating cell growth of certain types of tumors. We have recently demonstrated that PDGFs are potent angiogenic factors. Particularly, the angiogenic activity of PDGFs can be potentiated in the presence of other angiogenic factors. In addition to stimulation of blood angiogenesis, we have recently found that PDGFs can directly stimulate lymphangiogenesis and lymphatic metastasis. In this review, multiple roles of PDGFs in control of tumor growth and metastasis are discussed. PMID- 15655348 TI - Radiation-stimulated ERK1/2 and JNK1/2 signaling can promote cell cycle progression in human colon cancer cells. AB - The abilities of mutated active K-RAS and H-RAS proteins, in an isogenic human carcinoma cell system, to modulate the activity of signaling pathways and cell cycle progression following exposure to ionizing radiation is largely unknown. Loss of K-RAS D13 expression in parental HCT116 colorectal carcinoma cells blunted basal ERK1/2, AKT and JNK1/2 activity by -70%. P38 activity was not detected. Deletion of the allele to express activated K-RAS nearly abolished radiation-induced activation of all signaling pathways. Expression of H-RAS V12 in HCT116 cells lacking an activated RAS molecule (H-RAS V12 cells) restored basal ERK1/2 and AKT activity to that observed in parental cells, but did not restore or alter basal JNK1/2 and p38 activity. In parental cells radiation (1 Gy) caused stronger ERK1/2 pathway activation compared to that of the PI3K/AKT pathway. In H-RAS V12 cells radiation caused stronger PI3K/AKT pathway activation compared to that of the ERK1/2 pathway. Radiation (1 Gy) promoted S phase entry in parental HCT116 cells within 24h, but not in either HCT116 cells lacking K-RAS D13 expression or in H-RAS V12 cells. In parental cells radiation-stimulated S phase entry correlated with ERK1/2-, JNK1/2- and PI3K-dependent increased expression of cyclin D1 and cyclin A, and to a lesser extent cyclin E, 6-24 h after exposure. Cyclin A and cyclin D1 expression were not increased by radiation in cells lacking K-RAS D13 expression or in H-RAS V12 cells. Radiation (1 Gy) modestly enhanced expression of p53, hMDM2 and p21 in parental cells 2-6 h after exposure, which was abolished in cells lacking K-RAS D13 expression. Introduction of H-RAS V12 into cells lacking mutant active RAS partially restored radiation induced expression of p21 and p53, and enhanced the induction of hMDM2 beyond that observed in parental cells. Collectively, our findings argue that the coordinated activation of multiple signaling pathways, in particular ERK1/2 and JNK1/2, by radiation is required to elevate the expression of G1 and S phase cyclin proteins and to promote S phase entry in human colon carcinoma cells expressing wild type p53. In HCT116 cells H-RAS V12 promotes hMDM2 expression after radiation exposure which correlates with reduced p53 expression and increased cell survival. PMID- 15655349 TI - Selenomethionine regulates cyclooxygenase-2 (COX-2) expression through nuclear factor-kappa B (NF-kappaB) in colon cancer cells. AB - Previously, we showed that selenomethionine (Se-Met) inhibits growth of colon cancer cells via suppressing COX-2 expression at both mRNA and protein level. However, the molecular mechanism by which Se-Met suppresses COX-2 expression remains to be elucidated. To this end, we transiently transfected HCA-7 cells with different COX-2 promoter constructs followed by Se-Met treatment (90 microM) for 12 h. The results suggested the role of nuclear factor-kappa B (NF-kappaB) in transcriptional regulation of COX-2. We also observed complete inhibition of DNA binding activity of NF-kappaB in Se-Met (90 microM) treated HCA-7 cells as shown by electrophoretic mobility shift assay (EMSA). Supershift assays with anti-p65 antibody identified p65 subunit in the protein complex. We further demonstrate dose-dependent inhibition of nuclear translocation of NF-kappaB/p65 in Se-Met treated HCA-7 cells, which could explain the observed reduction in DNA binding of NF-kappaB/p65. These results suggest that Se-Met regulates COX-2 at transcriptional level by modulating the activity of NF-kappaB transcription factor. PMID- 15655350 TI - How the cell deals with DNA nicks. AB - During lagging strand DNA replication, the Okazaki fragment maturation machinery is required to degrade the initiator RNA with high speed and efficiency, and to generate with great accuracy a proper DNA nick for closure by DNA ligase. Several operational parameters are important in generating and maintaining a ligatable nick. These are the strand opening capacity of the lagging strand DNA polymerase delta (Pol delta ), and its ability to limit strand opening to that of a few nucleotides. In the presence of the flap endonuclease FEN1, Pol delta rapidly hands off the strand-opened product for cutting by FEN1, while in its absence, the ability of DNA polymerase delta to switch to its 3'-->5'-exonuclease domain in order to degrade back to the nick position is important in maintaining a ligatable nick. This regulatory system has a built-in redundancy so that dysfunction of one of these activities can be tolerated in the cell. However, further dysfunction leads to uncontrolled strand displacement synthesis with deleterious consequences, as is revealed by genetic studies of exonuclease defective mutants of S. cerevisiae Pol delta. These same parameters are also important for other DNA metabolic processes, such as base excision repair, that depend on Pol delta for synthesis. PMID- 15655351 TI - Ovariectomy aggravates hypersensitivity reactions to paclitaxel in rats. AB - The incidence of hypersensitivity reactions is still a matter of serious concern during chemotherapy with paclitaxel, particularly in patients with ovarian cancer. We recently reported that intravenous injection of paclitaxel causes acute lung injury characterized by vascular hyperpermeability, edema and respiratory dysfunction in rats. In the present study, we investigated the influence of ovariectomy on the paclitaxel-induced acute lung injury in rats. Ovariectomy worsened paclitaxel-induced acute lung injury, which was reversed by 17beta-estradiol. The mRNA expression for endothelial nitric oxide synthase was reduced in lungs of ovariectomized rats. To determine the role for nitric oxide, we examined the effects of several agents that modulate nitric oxide concentration on the pulmonary response to paclitaxel. In ovary-intact rats, a nitric oxide synthase inhibitor NG-nitro-L-arginine methyl ester exaggerated paclitaxel-induced acute lung injury, while nitric oxide donors such as sodium nitroprusside and isosorbide dinitrate attenuated the lung injury. Sodium nitroprusside was also effective in alleviating the paclitaxel-induced acute lung injury in ovariectomized rats. These findings suggest that ovariectomy enhances the susceptibility to paclitaxel hypersensitivity, in which decrease in estrogen and subsequent reduction in nitric oxide synthesis may be involved. PMID- 15655352 TI - The ARF tumor suppressor: keeping Myc on a leash. AB - The ARF tumor suppressor protein acts in a checkpoint that guards against unscheduled cellular proliferation in response to oncogenic signaling. Deregulated expression of c-Myc induces ARF expression and apoptosis through the ARF-Mdm2-p53 axis. Our recent study reveals a new direct role for ARF in controlling c-Myc's oncogenic activity that is independent of p53. ARF binds to and selectively impairs the transactivation ability of c-Myc while leaving its transrepression ability intact. Biologically, ARF prevents hyper-proliferation and transformation caused by c-Myc and enhances c-Myc-induced apoptosis independently of p53. These new findings may be especially relevant for therapeutic strategies targeting c-Myc-induced cancers. PMID- 15655354 TI - Histone H2AX phosphorylation after cell irradiation with UV-B: relationship to cell cycle phase and induction of apoptosis. AB - Damage to DNA that engenders double-strand breaks (DSBs) triggers phosphorylation of histone H2AX on Ser-139. Expression of phosphorylated H2AX (gammaH2AX) can be revealed immunocytochemically; the intensity of gammaH2AX immunofluorescence (IF) measured by cytometry was reported to correlate with the frequency of DSBs induced by X-ray radiation or by DNA damaging antitumor drugs. The aim of the present study was to measure expression of gammaH2AX following exposure of HeLa and HL-60 cells to a wide range of doses of UV-B light (6.1 J/m(2)-3.45 kJ/m(2)) and using multiparameter flow and laser scanning cytometry (LSC) to correlate DNA damage with cell cycle phase and induction of apoptosis. In both cell lines, the highest degree of H2AX phosphorylation induced by UV was seen in S-phase cells, particularly during early portion of S. In cells that did not replicate DNA (G(1), G(2) and M) the degree of H2AX phosphorylation was markedly lower than that in S-phase cells, and was strongly UV dose-dependent. Furthermore, the level of UV-induced gammaH2AX in G(1), G(2) and M was much higher in HeLa- than in HL 60- cells. Apoptotic cells become apparent >2h after exposure to UV and exhibited nearly an order of magnitude higher intensity of gammaH2AX IF than that initially induced by UV; predominantly S-phase cells underwent apoptosis. While the suppression of DNA replication, by aphidicolin prevented the induction of H2AX phosphorylation by UV in most S phase cells, it had no effect on a small cohort of cells that appeared to be entering S-phase, that expressed very high levels of gammaH2AX. Furthermore, aphidicolin itself induced gammaH2AX in early-S phase cells. The induction of gammaH2AX by UV was inhibited, but the incidence of apoptosis increased, by 5 mM caffeine, a known inhibitor of PI-3-related kinases. The data are consistent with the notion that H2AX phosphorylation observed throughout S phase reflects formation of DSBs due to the collision of replication forks with the UV-induced primary DNA lesions. Induction of gammaH2AX in G(1), G(2) and M is likely a response to the primary DSBs generated during UV exposure and/or DNA repair. It is unclear why the latter process was more pronounced in HeLa than in HL-60 cells. PMID- 15655353 TI - The second subunit of the replication factor C complex (RFC40) and the regulatory subunit (RIalpha) of protein kinase A form a protein complex promoting cell survival. AB - Replication Factor C (RFC) is required for the loading of Proliferating Cell Nuclear Antigen (PCNA) onto DNA during DNA replication, repair and recombination. RFC40, the second subunit of the RFC complex, and PCNA have been shown to be overexpressed in gestational trophoblastic diseases. Using RFC40 as the bait in a yeast two-hybrid screening, we have identified a novel interaction between RFC40 and the regulatory subunit (RIalpha) of cAMP-dependent Protein kinase A (PKA). The interaction sites between these two proteins were investigated and mapped to the N-terminus of RIalpha and the C-terminus of RFC40. Moreover, it was demonstrated that the C-subunit of PKA was not associated with the RFC40-RIalpha complex. Furthermore, RFC37, the third subunit of the RFC complex, competes with RIalpha and displaces it from the RFC40-RIalpha complex. Interestingly, downregulation of endogenous RIalpha by 8-chloro cAMP, in MCF7 breast cancer cells led to reduction in the amount of RFC40-RIalpha complex, together with decrease in cell survival. PMID- 15655355 TI - Aneuploidy-cancer predisposition syndromes: a new link between the mitotic spindle checkpoint and cancer. AB - Genetic cancer predisposition syndromes have been crucial to the identification of genes and pathways involved in carcinogenesis. Constitutional gene mutations segregating with distinctive cancer phenotypes provide unequivocal evidence of a gene's causal role in cancer. This type of evidence has been central in proving that oncogenes and tumor suppressor genes can cause human cancers, but has been lacking for genes implicated in generating aneuploidy. However, recently we identified mutations in the mitotic checkpoint gene BUB1B in an autosomal recessive condition characterized by mosaic aneuploidies and childhood cancers. This finding strongly suggests that aneuploidy is causally related to cancer development. PMID- 15655356 TI - A distinct "side population" of cells in human tumor cells: implications for tumor biology and therapy. AB - Stem cells have an extensive capacity to proliferate, differentiate and self renew. In many mammals, including humans, an adult stem cell subpopulation termed the "side population" (SP) has been identified. SP cells can rapidly efflux lipophilic fluorescent dyes to produce a characteristic profile based on fluorescence-activated flow cytometric analysis. Previous studies have demonstrated SP cells in bone marrow obtained from patients with acute myeloid leukemia, suggesting that these cells might be candidate leukemic stem cells, and recent studies have found a SP of tumor progenitor cells in human solid tumors. These new data indicate that the ability of malignant SP cells to expel anticancer drugs may directly improve their survival and sustain their clonogenicity during exposure to cytostatic drugs, allowing disease recurrence when therapy is withdrawn. Identification of a tumor progenitor population with intrinsic mechanisms for cytostatic drug resistance might also provide clues for improved therapeutic intervention. PMID- 15655357 TI - CHK1 affecting cell radiosensitivity is independent of non-homologous end joining. AB - CHK1 is one of the most important checkpoint proteins in mammalian cells for responding to DNA damage. Cells defective in CHK1 are sensitive to ionizing radiation (IR). The mechanism by which CHK1 protects cells from IR-induced killing remains unclear. DNA double strand breaks (DSBs) induced by IR are critical lesions for cell survival. Two major complementary DNA DSBs repair pathways exist in mammalian cells, homologous recombination repair (HRR) and nonhomologous end joining (NHEJ). By using CHK1 kinase dead human cell lines established in our laboratory, we show here that although these human cell lines have different CHK1 activities with different sensitivities to IR-induced killing and G(2) accumulation, all these cell lines show similar inductions and rejoining rates of DNA DSBs. These results indicate that the different radiosensitivities and G(2) checkpoint responses in these cell lines are independent of NHEJ, suggesting that CHK1-regulated checkpoint facilitates HRR and therefore protects cells from IR-induced killing. PMID- 15655358 TI - Glimpsing over the event horizon: evolution of nuclear pores and envelope. AB - The origin of eukaryotes from prokaryotic ancestors is one of the major evolutionary transitions in the history of life. The nucleus, a membrane bound compartment for confining the genome, is a central feature of eukaryotic cells and its origin also has to be a central feature of any workable theory that ventures to explain eukaryotic origins. Recent bioinformatic analyses of components of the nuclear pore complex (NPC), the nuclear envelope (NE), and the nuclear transport systems revealed exciting evolutionary connections (e.g., between NPC and coated vesicles) and provided a useful record of the phyletic distribution and history of NPC and NE components. These analyses allow us to refine theories on the origin and evolution of the nucleus, and consequently, of the eukaryotic cell. PMID- 15655359 TI - Breast cancer cell lines grown in vivo: what goes in isn't always the same as what comes out. AB - Human cell lines constitute powerful model systems for the in vitro and in vivo analysis of cancer. Cancer lines that are invasive in culture often form tumors and metastases in immune deficient mice. It is generally assumed that, in such cases, the principal population of cancer cells in culture corresponds to the tumor-forming cells in vivo. Here, we high-light a recent functional proteomics investigation that suggests the contrary. In this study, cells derived from orthotopic xenograft tumors formed by the invasive breast cancer line MDA-MB-231 were found to exhibit profound differences in their enzyme activity profiles and increased tumor growth rates and metastasis when compared to the parental line. These findings suggest that the in vivo microenvironment of the mouse mammary fat pad cultivates the growth of human breast cancer cells with elevated tumorigenic properties. Characterization of the unique molecular properties of these tumor forming cells may reveal new strategies for the diagnosis and treatment of breast cancer. PMID- 15655360 TI - Inflammatory hypoxia: role of hypoxia-inducible factor. AB - Sites of inflammation are characterized by significant changes in metabolic activity. Shifts in energy supply and demand can result in diminished delivery and/or availability of oxygen, leading to inflammation-associated tissue hypoxia and metabolic acidosis. These shifts in tissue metabolism, as indicated by previous studies, are frequently associated with vasculitis and profound recruitment of inflammatory cell types, particularly myeloid cells such as neutrophils (PMN) and monocytes. Here, we review recent work addressing the influence of hypoxia on development of inflammatory lesions, with particular emphasis on molecular pathways regulated by hypoxia-inducible factor (HIF). PMID- 15655361 TI - Newly identified roles for an old guardian: profound deficiency of the mitotic spindle checkpoint protein BubR1 leads to early aging and infertility. AB - The mitotic spindle checkpoint ensures proper chromosome segregation, and thereby guards against the deleterious effects of aneuploidy. The protein components of the check-point machinery include evolutionarily conserved proteins such as BubR1. While the molecular and cellular biology of this checkpoint is becoming increasingly clarified, the ultimate consequences for overall health of deficiency of specific components such as BubR1 are much less clear--in part due to the embryonic lethality of complete knockouts. Through a clever combination of hypomorphic and knockout alleles, Baker and colleagues were able to engineer mice with graduated levels of BubR1 protein. In doing so, they established the threshold permitting survival to adulthood, but even more intriguingly, they discovered critical roles for BubR1 in preventing early aging and infertility. PMID- 15655363 TI - Poly(ADP-ribosyl)ation and epigenetics. Is CTCF PARt of the plot? AB - Despite the fact that the poly(ADP-ribose) (PAR) modification of proteins has been known for more than four decades, there is no unifying picture of the pathways governed by this process. While the function of poly(ADP-ribosyl)ation (PARlation) has shown to be traditionally associated with DNA repair and genotoxic stress, there is an emerging view that PARlation is also important in epigenetic regulation of chromatin structure and gene expression in the normal context. This view has been exemplified by the recent demonstration that PARlation is essential for the manifestation of the imprinted state of the Igf2 gene. In particular, the PARlation mark was shown to associate preferentially with the maternally inherited H19 ICR allele, this association depended on functional target sites of the chromatin insulator protein CTCF and, importantly, CTCF itself was found to be PARlated. Given that CTCF is currently the only known factor common for all vertebrate chromatin insulators, it is not surprising that the derepression of the maternal Igf2 allele by 3-aminobenzamide (an inhibitor of PAR polymerases) could be linked to a perturbed chromatin insulator function at the H19 ICR. This feature appears to extend to more than 150 chromatin insulators that were isolated due to their in vivo interaction with CTCF. In this review, we discuss in more depth these results and point out that the turnover of the PARlation mark at DNA-bound CTCF is indicative of a novel mode of rheostat control of expression domains possibly by regulating the stability of higher order chromatin conformations. PMID- 15655362 TI - Immunological consequences of macrophage-mediated clearance of apoptotic cells. AB - Apoptosis and the rapid clearance of apoptotic cells by professional or nonprofessional phagocytes are normal and coordinated processes that ensure controlled cell growth with a nonpathological outcome. Defects in clearance of apoptotic cells by macrophages have serious consequences often resulting in autoimmune disorders. Phagocyte-derived immunoregulatory cytokines such as Interleukin-12 and Interleukin-10 play pivotal roles in the etiology and pathology of many autoimmune diseases. Elucidation of the apoptotic cell-mediated signaling mechanisms involved in the control of pro- and anti-inflammatory cytokines during cell turnovers under normal and pathological conditions may help us counter the cytokine dysregulation and control inappropriate host immune reactions in pathological situations such as autoimmunity, infectious diseases, graft-versus-host disease, and cancer. PMID- 15655364 TI - c-Abl in oxidative stress, aging and cancer. AB - c-Abl is activated by oxidative stress but its precise function in cell response to this stress is elusive. Studies of c-Abl(-/-) osteoblasts revealed that c-Abl played a negative role in the induction of peroxiredoxin I (Prx I, Prdx I), an anti-oxidant protein with tumor suppression activity. In contrast, Atm, a signaling molecule that interacts with c-Abl and is required for c-Abl activation, served a totally different function. The significance of these findings is discussed here in the context of aging and tumorigenesis and their links to reactive oxygen species. c-Abl and its derivatives BCR-ABL and v-Abl were discovered more than twenty years ago. BCR-ABL and v-Abl acquire elevated tyrosine kinase activities by fusing to BCR and gag respectively and are capable of transforming myeloid and fibroblast cells. BCR-ABL is also the underlying cause in the development of most cases of chronic myeloid leukemia (CML) in humans. In contrast, c-Abl takes on an auto-inhibiting conformation and its activation requires post-translational modifications such as phosphorylation and myristoylation. The physiological functions of c-Abl remain elusive. PMID- 15655365 TI - Revisiting the "Cdk-centric" view of the mammalian cell cycle. AB - Since the early genetic studies in yeast, regulation of the cell cycle has been associated with the sequential activation of several proline-directed serine threonine protein kinases by cyclins. From yeast to humans, the activity of these cyclin-dependent kinases (Cdks) have been thought to be essential for cell cycle regulation. Recent gene-targeted mouse models for different cyclins and Cdks have shown that members of these families show a certain level of redundancy and that specific complexes are not required for the mitotic cell cycle. However, the complexity of the Cdk-cyclin network and the promiscuity of their members makes it difficult to understand the relative contribution of these proteins to the mammalian cell division cycle. Compensatory roles by non-Cdk activities and Cdk independent functions of cyclins are increasing the complexity of the current simplistic models. We still do not know whether at least one cyclin-dependent kinase activity is required for cell cycle progression in mammalian cells. Indeed, this is a relevant question for cancer therapy. PMID- 15655366 TI - Recruiting substrates to cullin 4-dependent ubiquitin ligases by DDB1. AB - The ubiquitin-proteasome system is the major pathway by which cells target proteins for degradation in a specific manner. The E3 ubiquitin ligase, which brings targeted proteins (substrates) and activated ubiquitin in close proximity, enabling covalent conjugation of ubiquitin to the substrate, is an essential component of this system. Of the E3 ligases, the cullin (CUL) ligases are of high interest because of their capacity to form multiple distinct E3 complexes to ubiquitinate a potentially large number of substrates. Of the six closely related cullins, very little is known about how specific substrates are recruited to CUL4 dependent ligases. A recent paper in Nature Cell Biology may shed some light on this issue as well as on the function of DDB1, a damaged-DNA binding protein that has long been associated with DNA repair. PMID- 15655367 TI - Reprogramming specific gene expression pathways in B-cell lymphomas. AB - The BCL6 transcriptional repressor is normally expressed during the germinal center phase of B-cell differentiation. Germinal center B-cells typically undergo rapid proliferation in spite of accumulating DNA damage caused by class switch recombination and somatic hypermutation. BCL6 is required to license B-cells for the germinal center reaction and its down regulation appears to be important for cells to exit this stage and undergo further differentiation. BCL6 appears to mediate these biological effects by recruiting corepressor complexes to silence critical cell cycle checkpoint and differentiation related genes. Based on our data and recent publications, we propose that these gene pathways are regulated through distinct transcriptional mechanisms, which can be specifically targeted to reprogram B-cells preferentially for either growth suppression and apoptosis, or differentiation. As BCL6 plays a central role in the pathogenesis of diffuse large B-cell lymphoma, we predict that targeting BCL6 transcriptional repression complexes in malignant B-cells may constitute a novel form of transcription therapy for lymphomas and possibly other tumors. PMID- 15655368 TI - Dok1 and SHIP act as negative regulators of v-Abl-induced pre-B cell transformation, proliferation and Ras/Erk activation. AB - The v-Abl tyrosine kinase activates several signaling pathways during transformation of bone marrow cells in mice. Because the SH2-containing inositol 5'-phosphatase (SHIP) and Downstream of tyrosine kinase 1 (Dok1) have been shown to interact with Abl, the effect of SHIP and Dok1 deficiency on v-Abl transformation was investigated. Bone marrow cells from either Dok1- or SHIP deficient mice are more susceptible to transformation by v-Abl. v-Abl-transformed preB cells from these knockout mice show Abl kinase-dependent hyperproliferation and moderate resistance to apoptosis. Elevated activation of Ras, Raf-1, and Erk, but not of Akt, was observed in either SHIP(-/-) or Dok1(-/-) v-Abl-transformed cells. This activation is sensitive to treatment with STI571. Furthermore, treatment of these cells with either a farnesyltransferase inhibitor or a MEK1/2 inhibitor abrogates the increased proliferation of SHIP(-/-) or Dok1(-/-) cells in a dose-dependent manner. Complementation of SHIP(-/-) or Dok1(-/-) cells abrogates their hyperproliferation and intracellular Erk activation. These data indicate that both SHIP and Dok1 functionally regulate the activation of Ras-Erk pathway by v-Abl and affect the mitogenic activity of v-Abl transformed bone marrow cells. PMID- 15655369 TI - Hematopoietic stem cells, leukemic stem cells and chronic myelogenous leukemia. AB - Blood-related cancers, or leukemias, have been shown to arise from a rare subset of cells that escape normal regulation and drive the formation and growth of the tumor. The finding that these so-called cancer stem cells, or leukemic stem cells (LSC), can be purified away from the other cells in the tumor allows their precise analysis to identify candidate molecules and regulatory pathways that play a role in progression, maintenance, and spreading of leukemias. The analyses of the other, numerically dominant, cells in the tumor, while also interesting, do not directly interrogate these key properties of malignancies. Mouse models of human myeloproliferative disorder and acute myelogenous leukemia have highlighted the remarkable conservation of disease mechanisms between both species. They can now be used to identify the LSC for each type of human leukemia and understand how they escape normal regulation and become malignant. Given the clinical importance of LSC identification, the insights gained through these approaches will quickly translate into clinical applications and lead to improved treatments for human leukemias. PMID- 15655370 TI - Proteasome inhibition in the treatment of cancer. AB - The proteasome is the main extralysosomal system involved in intracellular proteolysis. A number of proteasome substrates, including cyclins, IkappaB, and p53, are critical to cell cycle progression and apoptosis. Interruption of the degradation of these substrates through proteasome inhibition is a novel and unique approach to the treatment of malignancies. First-generation proteasome inhibitors lacked usefulness because of broad specificity and irreversible binding to the proteasome. However, the later synthesis of the peptide boronic acid proteasome inhibitor bortezomib allowed for selective, reversible binding. Basic investigations have reported the antitumor activity of bortezomib in a variety of hematologic and solid tumor models and have demonstrated the ability of bortezomib to enhance chemosensitivity and overcome cellular mechanisms of drug resistance attributable, in part, to abrogation of NF-kappaB induction. In patients with relapsed, refractory multiple myeloma who had received a median of six prior regimens, treatment with bortezomib resulted in a 35% response rate (complete plus partial plus minimal response) using criteria of the European Group for Blood and Marrow Transplantation. Encouraging activity has been demonstrated with bortezomib in the first-line treatment of myeloma and in patients with mantle cell lymphoma. Investigations of its utility in the treatment of patients with solid tumors are ongoing. PMID- 15655371 TI - The nucleolus. Playing by different rules? AB - The nucleolus, although an integral part of all eukaryotic nuclei, plays by its own rules in many respects. Cytologically, it is the most prominent subnuclear domain; functionally, it is the site of transcription of 3 of the ribosomal RNAs from the tandemly repeated rDNA, and subsequently of ribosome biogenesis; biochemically, it possesses transcriptional and post-transcriptional machinery not shared with the rest of the nucleus. Making the huge number of ribosomes required by the cell represents an enormous investment of metabolic activity, and so nucleolar function can easily become a de facto limit to cell growth: nucleolar activity must also be actively regulated, but the detailed regulatory networks linking the nucleolus with cellular metabolism are still unclear. Several recent reports have now shown that segregation of the rDNA in yeast, along with telomeric DNA, is also controlled differently from the rest of the genome. This short review describes some of the features of the nucleolus and highlights recent progress in understanding this important, but enigmatic, nuclear structure. PMID- 15655372 TI - ATM and ATR check in on origins: a dynamic model for origin selection and activation. AB - Initiation of DNA replication occurs at origins of replication, traditionally defined by specific sequence elements. Sequence-dependent initiation of replication is the rule in prokaryotes and in the yeast Saccharomyces cereviseae. However, sequence-dependent initiation does not appear to be absolutely required in metazoan eukaryotes. Origin firing is instead likely dependent on stochastic initiation from chromatin-defined loci, despite the demonstration of some specific origins. Based on some recent observations in Xenopus laevis egg extracts and in mammalian cell culture, we propose that timing of origin firing is dependent on feedback from active replicons. This dynamic regulation of replication is mediated by sensing of ongoing replication by the DNA-damage checkpoint kinases ATM and ATR, which in turn downregulate neighboring and distal origins and replicons by inhibition of the S-phase kinases Cdk2 and Cdc7 and by inhibition of the replicative Mcm helicase. Origin selection, activation, and replicon progression are therefore constrained in both space and time via feedback from the cell cycle and ongoing replication. PMID- 15655373 TI - Signalling DNA damage by regulating p53 co-factor activity. AB - In response to DNA damage the related phosphatidylinositol-3-OH-kinase-like kinases ATM and ATR phosphorylate downstream protein targets which facilitate the DNA damage response. A new pathway in which ATM phosphorylates the transcriptional co-factor Strap has been elucidated. Phosphorylation causes the stabilization of nuclear Strap and favours the formation of a stress-responsive co-activator complex. Strap activity enhances p53 acetylation, and augments the response to DNA damage. Most interestingly, in AT cells Strap remains cytoplasmic, and a mutant derivative that cannot be phosphorylated by ATM is similarly localised to the cytoplasm. These results argue that Strap is an important downstream effector in the DNA damage response. PMID- 15655374 TI - A case of selfish nucleolar segregation. AB - Mitotic segregation of nucleolus in fission and budding yeast proceeds without disassembling its complex structure, creating challenging problems for transmission of nucleolus-organizing regions during nuclear division. The SMC complex called condensin, which plays a leading role in organizing mitotic structure of chromosomes in all eukaryotes, is essential for nucleolar segregation in budding yeast, where rDNA chromatin is the main target of mitotic condensin activity. Mitosis-specific condensin targeting to the nucleolus presents an attractive model to study mechanisms controlling condensin binding to specific chromatin domains. Recent reports suggest that the early-anaphase release of Cdc14 from the nucleolus (FEAR pathway) controls the proficiency of nucleolar segregation by promoting the mitotic condensin function in rDNA. This finding uncovers an essential function for the FEAR pathway and postulates the unique nucleolar self-regulatory mechanism, which evolved to recruit two essential enzymatic activities, Cdc14 phosphatase and condensin ATP-dependent supercoiling, for the specific task of segregating nucleoli without their disassembly. PMID- 15655375 TI - Characterization and gene expression profiling of a stable cell line expressing a cell cycle GFP sensor. AB - The use of stable cell lines expressing fusions with green fluorescent protein (GFP) has increased significantly in recent years. In this study we have used a range of complimentary analytical techniques to examine the characteristics of a cell line stably expressing a EGFP cell cycle sensor relative to parental U2OS cells. Analysis of cell cycle duration and cell cycle phase distribution by cell growth assays and flow cytometry revealed that the two cell lines had identical doubling times and cell cycle distributions. Measurement of EGFP fusion protein mRNA by quantitative RT-PCR indicated a EGFP sensor expression level equivalent to endogenous Cyclin B1 (7000 copies/cell in G2). Microarray analysis showed a 0.9% (>2 fold at p<0.001 across 20,000 genes) difference in global gene expression levels between parental and EGFP expressing U2OS cells, with no significant differences in expression of A, B, C, D, E, F, G, H, I, K, L, M or T type Cyclins between the two cell types. These results confirm that engineering a stable cell line for low expression of a EGFP cell cycle sensor is minimally perturbing to the cell cycle and cellular gene expression. PMID- 15655376 TI - Bridging the BMP and Wnt pathways by PI3 kinase/Akt and 14-3-3zeta. AB - BMP, PTEN and Wnt/beta-catenin pathways are the three signaling pathways that control normal development and regeneration of the intestine, and contribute to intestinal polyposis when aberrant inactivation or activation occurs in each of these pathways. Using genetic targeting of BMPR1A in mice, we show that inactivation of BMP signaling results in multiple polyps due to an increased number of crypts and stem cells, accompanied by enhanced Wnt signaling in all proliferating intestine cells. However the increased transcriptional activity of Wnt effecter protein, beta-catenin, is found primarily in intestine stem cells (ISCs). Concurrently, PTEN, an inhibitor of PI3K/Akt pathway, is also primarily inactivated in the ISCs, leading to activation of Akt. Thus, Akt may contribute to activation of beta-catenin in ISCs in coordination with Wnt signaling. By conducting a proteomic analysis of the beta-catenin complex, we show that 14-3 3zeta exists in the beta-catenin complex and facilitates activation of beta catenin by Akt, which, intriguingly, appears to be predominantly in ISCs. Thus, we propose that BMP signaling plays a role in inhibition of ISC self-renewal through suppression of Wnt/beta-catenin signaling in ISC, and this cross-talk is bridged, at least in part, through the PTEN/Akt pathway and further enforced by 14-3-3zeta. PMID- 15655377 TI - Nuclear receptors as negative modulators of STAT3 in multiple myeloma. AB - Multiple myeloma (MM) remains largely incurable despite conventional and high dose therapies. Therefore, novel biologically based treatment approaches are urgently required. Particularly, STAT3 activated by IL-6 has a key role in preventing apoptosis and stimulating growth of multiple myeloma cells. Nuclear receptors, a distinct class of ligand-activated transcriptional factors, can interact and modify the function of transcriptional factors intrinsic to the cytokine signal transduction pathways. We have investigated regulation of two nuclear receptors, peroxisome proliferator-activated receptor gamma (PPARgamma) and estrogen receptor (ER), and their crosstalk with STAT3 in multiple myeloma. These results indicate that ligand-activated nuclear receptors can function as negative modulators of STAT3 through direct mechanisms, or in turn, by facilitating coregulators such as PIAS or SMRT. Therefore, different classes of nuclear receptors affect suppression of STAT3 functions through diverse mechanisms resulting in downregulating IL-6-mediated cell growth and gene expression. Given the importance of IL-6 in multiple myeloma, the estrogen receptor-STAT3 or PPARgamma-STAT3 interaction may have significant therapeutic implications in multiple myeloma. PMID- 15655378 TI - Bub1 multitasking in mitosis. AB - Accurate partition of duplicated genetic material to the daughter cells during mitosis relies on the maintenance of the physical linkage (cohesion) between sister chromatids until their bipolar attachment to the mitotic spindle. In response to a single straying chromatid within a cell, a surveillance mechanism called the spindle checkpoint blocks the ubiquitin ligase activity of the anaphase-promoting complex or cyclosome (APC/C), stabilizes securin (an APC/C substrate and an inhibitor of separase), and delays the activation of separase. This in turn prevents cleavage of cohesin by separase, preserves sister chromatid cohesion, and delays the onset of anaphase. The protein kinase, Bub1, is a key component of the spindle checkpoint. Bub1 has an upstream function in regulating the kinetochore localization of Mad2 and other downstream checkpoint components. In addition, recent biochemical studies have shown that Bub1 directly phosphorylates the APC/C activator, Cdc20, and inhibits APC/C. Finally, Bub1 has a noncheckpoint function at the kinetochores and preserves centromeric cohesion through the MEI-S332/shugoshin family of proteins. Therefore, Bub1 performs multiple tasks in mitosis that ensure the proper inheritance of chromosomes. PMID- 15655379 TI - Protein kinase specificity. A strategic collaboration between kinase peptide specificity and substrate recruitment. AB - Specificity of phosphorylation by protein kinases is essential to the integrity of biological signal transduction. Specificity is determined by two critical elements: (1) peptide specificity of the kinase, i. e., preferential phosphorylation of S/T/Y residues surrounded by particular patterns of amino acids; and (2) recruitment, i. e., increasing the frequency of encounter between kinase and substrate. Historically, the importance of peptide specificity was studied first, but it has been somewhat overshadowed by emerging emphasis on the importance of recruitment. Recent studies confirm and extend understanding of the relative contribution of these two elements. Peptide specificity always constrains the range of sites that can be phosphorylated by a kinase. Only when recruitment is very strong, as in the case with autophosphorylation, can markedly suboptimal substrates be phosphorylated. PMID- 15655380 TI - Early and aggressive treatment of complex hypertension. PMID- 15655381 TI - Angiotensin-converting enzyme i/d polymorphism in patients with malignant hypertension. AB - The angiotensin-converting enzyme (ACE) gene has been implicated in the manifestation of the phenotype of malignant hypertension (MH). In 1990 the ACE gene polymorphism characterized by the insertion or deletion of a 287-base pair fragment in the 17q23 chromosome was identified. The DD genotype is associated with increased tissue and circulating ACE levels and elevated angiotensin II. ACE polymorphism was studied in 48 patients with MH, 25 patients with non-MH, and a control group of 78 normotensive individuals by real-time polymerase chain reaction using the LightCycler system (Roche Diagnostics Corporation, Indianapolis, IN). The DD genotype was found statistically more frequently in MH patients than controls (p=0.028; odds ratio, 2.5; confidence interval, 1.1-5.5). Presence of the DD genotype of the ACE gene is more frequent in MH patients than in controls, indicating that this genotype could be a significant risk factor and a predictor for the development of MH. PMID- 15655383 TI - Roundtable discussion: hypertension in children and adolescents. AB - Following a hypertension symposium held in Philadelphia, PA, on June 8, 2004, a roundtable discussion was held to discuss the treatment of hypertension in adolescents and children. The participants included Thomas D. Giles, MD, of Louisiana State College of Medicine, New Orleans, LA; Bonita Falkner, MD, of the Department of Medicine and Pediatrics, Thomas Jefferson University, Philadelphia, PA; Allan B. Schwartz, MD, of the Division of Nephrology and Hypertension, Drexel University College of Medicine, Philadelphia, PA; Raymond R. Townsend, MD, of the Department of Medicine, University of Pennsylvania School of Medicine, Philadelphia, PA; and Marvin Moser, MD, of the Department of Medicine/Cardiology, Yale University School of Medicine, New Haven, CT. PMID- 15655382 TI - Intraindividual blood pressure responses to angiotensin-converting enzyme inhibition and angiotensin receptor blockade. AB - This study aims to test the hypothesis that in some hypertensive subjects the blood pressure (BP) response to angiotensin-converting enzyme inhibition differs from that to angiotensin receptor blockade (ARB); a responder to angiotensin converting enzyme inhibition may not respond to ARB or the opposite. A randomized, open-label, crossover, comparative trial of lisinopril 20 mg compared with telmisartan 80 mg (5 weeks per treatment period) was conducted in 32 untreated hypertensives using 24-hour ambulatory BP monitoring. Subjects were classified as "responders" and "nonresponders" using an arbitrary threshold of ambulatory BP response (> or =10 mm Hg systolic or > or =5 diastolic) or the median response achieved by each drug. No difference was detected between the drugs in their effect on ambulatory BP (mean difference 1.2+/-7.1/0.7+/-5.1 mm Hg, systolic/diastolic). Significant correlations were found between the antihypertensive responses to the two drugs (r=0.77, p<0.001). Using the arbitrary response criterion, there was a difference between the drugs in the responses in 28%/13% of subjects (9/4 patients) for systolic/diastolic BP (19%/25% using the median response criterion). These data suggest that in some hypertensive patients the BP response to angiotensin-converting enzyme inhibition may fail to predict the response to ARB. It appears that there are differences in the antihypertensive action of angiotensin-converting enzyme inhibitors and ARBs that may be clinically important. PMID- 15655385 TI - Hypertension management: differing points of view. Should we routinely measure renin levels to diagnose and treat patients with hypertension? The argument for. PMID- 15655384 TI - Commentary: The ALLHAT Study: implications for the management of resistant hypertension. PMID- 15655386 TI - Hypertension management: differing points of view. Should we routinely measure renin levels to diagnose and treat patients with hypertension? The argument against. PMID- 15655387 TI - Hypertension icons: Henry R. Black's interview with Marvin Moser, MD. PMID- 15655388 TI - Reflections in hypertension. How should blood pressure be measured during pregnancy? PMID- 15655389 TI - Case studies in hypertension. Dihydropyridine/nondihydropyridine calcium channel blocker combination therapy. PMID- 15655391 TI - Current concepts of pharmacotherapy in hypertension. Carvedilol: new considerations for its use in the diabetic patient with hypertension. PMID- 15655394 TI - Oscillopsia: editorial review. PMID- 15655395 TI - Episodic vertigo. AB - PURPOSE OF REVIEW: This review focuses on three neuro-otological syndromes, which are all marked by rapid scientific progress on the one hand but under-recognition or undertreatment on the other: benign paroxysmal positional vertigo and its variants, superior semicircular canal dehiscence syndrome, and migrainous vertigo. RECENT FINDINGS: The efficacy of Epley's maneuver for treatment of benign paroxysmal positional vertigo has been convincingly demonstrated by a meta analysis of nine randomized controlled trials. Head vibration during Epley's procedure and keeping upright for 48 h after effective treatment do not improve the outcome. Superior canal dehiscence syndrome presents not only with sound and pressure-induced vertigo but also with conductive hearing loss at low frequencies. Migrainous vertigo may present not only with spontaneous attacks but also with positional vertigo or with chronic dizziness and imbalance. Vestibular rehabilitation has been proven to relieve chronic dizziness and visual vertigo. SUMMARY: Recent studies have eliminated several white spots on the neuro otological map. However, many areas are still unexplored, particularly with regard to treatment of specific vestibular syndromes where randomized controlled trials are just at their beginning. PMID- 15655396 TI - Pharmacology of vertigo/nystagmus/oscillopsia. AB - PURPOSE OF REVIEW: To describe recent developments in the pharmacological treatment of vertigo and nystagmus while focusing on vestibular neuritis, Meniere's disease, downbeat nystagmus, periodic alternating nystagmus, acquired pendular nystagmus, and superior oblique myokymia. RECENT FINDINGS: In the last 2 years several studies have been published on possible pharmacological treatment options for nystagmus and oscillopsia. In the treatment of vestibular neuritis two studies showed that cortisone treatment was effective for restoring labyrinthine function. This benefit seems more likely if treatment is started within the first 2 days of onset. For recurrent vertigo attacks due to Meniere's disease, the titration technique with daily or weekly doses of intratympanic gentamicin until onset of vestibular symptoms, change in vertigo or hearing loss rated best for complete vertigo control. A new pharmacological treatment option for downbeat nystagmus is the administration of potassium channel blockers (e.g. 4-aminopyridine). They are thought to reinforce the inhibitory action of cerebellar Purkinje cells. Several case reports have proven the beneficial effect of baclofen on periodic alternating nystagmus, of gabapentin and memantine on acquired pendular nystagmus, and of carbamazepine and gabapentin on superior oblique myokymia. SUMMARY: There have been several new developments in the treatment of nystagmus and vertigo over the last 2 years. These include potassium channel blockers for the treatment of downbeat nystagmus, early cortisone treatment to improve recovery of the labyrinth function in vestibular neuritis, and intratympanic gentamicin treatment for Meniere's disease. Other pharmacological treatment options are baclofen for periodic alternating nystagmus, gabapentin and memantine for acquired pendular nystagmus, and carbamazepine for superior oblique myokymia. PMID- 15655397 TI - A speedy solution for balance and gait analysis: angular velocity measured at the centre of body mass. AB - PURPOSE OF REVIEW: Falls are a serious problem for the elderly and others prone to fall. In particular, those over 65 years of age will suffer at least one fall a year and as a result will need cost-intensive medical treatment. Under these circumstances, an optimal clinical pathway should first identify those with a tendency to fall, second, pinpoint the disease-specific deficits in balance control and walking patterns, and third, offer the possibility through focused biofeedback training to reduce the number of falls suffered. This review describes new methods of assessing balance control in both static and dynamic balance conditions and their applications for balance rehabilitation. RECENT FINDINGS: Technological advances and applied research have shown that body-worn sensors measuring angular velocity (gyroscopes) or the acceleration of the trunk can effectively quantify balance during stance and gait tasks, and can be used to detect potential fallers and discriminate between different balance disorders. Standing on foam support reveals balance deficits regardless of the underlying disease. Disease-specific balance deficits can be characterized by the pattern of trunk movements between various semi-stance and gait tasks. Angular velocity sensors have recently been shown to provide highly relevant information for use as biofeedback, or as an ambulatory device to record balance and gait performance over long periods of time in both clinical and natural living conditions. SUMMARY: Angular velocity sensors can provide balance-relevant information directly to the clinician, and provide an important improvement on the complicated and time/cost-intensive techniques of traditional balance measurement systems. PMID- 15655398 TI - Surgical treatment of Meniere's disease. AB - PURPOSE OF REVIEW: This review comprises new insights from and discusses the impact of recent medical publications on the surgical treatment of Meniere's disease. RECENT FINDINGS: Refining surgical indications through recognition of clinical conditions with similar symptoms and through a more precise estimation of the degree of disability will improve the process of decision making for surgery. Further high-level evidence-based medical data supporting the effectiveness of intratympanic gentamycin has become available. Physiopathological progress, based on animal experiments, towards surgically applied intracochlear drug delivery is addressed. SUMMARY: Studies using level 1 or 2 evidence-based medicine must be conducted to enable better decision making, such as in the application of intratympanic gentamycin or micropressure Meniett therapy at an earlier stage of Meniere's disease. If the results of such studies are conclusive for surgery, this will lead to a shorter duration of discomfort for patients before being offered the possibility of surgery. PMID- 15655399 TI - Motion sickness. AB - PURPOSE OF REVIEW: The public's longstanding resigned tolerance to motion sickness threatens to change, due to the widespread introduction of nauseogenic tilting trains and the increasing use of virtual reality immersion. RECENT FINDINGS: Scientific effort over the last 5 years has focused on precise evaluation of the stimuli that provoke sickness and on the development of behavioural and new pharmacological interventions to suppress sickness. SUMMARY: The precise mechanical ride characteristics of vehicles that provoke sickness have been identified and this will lead to guidelines for future engineering design, especially for suspension systems that compensate for inertial tilt, and recommendations for passengers at risk. The frequency characteristics of motion provoking sickness have been defined with greater precision and identified with shifts in perception of motion versus orientation, and changes in the quality of reflex eye movements. Ability to modify readily the time constant of vestibular 'velocity store' has emerged as a potential candidate marker of successful motion sickness habituation. Behavioural 'autogenic' countermeasures to the development of sickness, such as controlled breathing, which can be implemented readily, are shown to have significant ameliorating effects on nausea and are of value for short term moderate exposures. New classes of pharmacological agents such as N methyl-D-aspartate antagonists and 5HT1a receptor agonists show promise in animals but await trials in humans. PMID- 15655400 TI - Some recent and personal ideas about stroke treatment and prophylaxis. PMID- 15655401 TI - Angioplasty and stenting of intracranial stenosis. AB - PURPOSE OF REVIEW: The review summarizes recent advances in angioplasty and stenting of intracranial atherosclerotic stenosis. RECENT FINDINGS: Several single-center studies show that, with the technological advancements in device technology, intracranial stenting is feasible even in the distal internal carotid artery or middle cerebral artery. However, the clinical outcomes vary between moderate and high risk rates for morbidity and mortality. To date, the only prospective multi-center study that has been conducted is the SSYLVIA (Stenting of Symptomatic Atherosclerotic Lesions in the Vertebral or Intracranial Arteries) study, which demonstrated stroke rates of 6.6% and 13.1% at 30 days and 1 year, respectively. A unique feature of this trial is the assessment of restenosis rates and their potential association with clinical symptoms and risks. A promising new concept for stent-assisted procedures in the intracranial vasculature is the use of self-expanding stents. The high flexibility of these stents makes them suitable for treatment of lesions of the distal internal carotid artery and middle cerebral artery that are either inaccessible or difficult to reach with a balloon-mounted stent. Additionally, these self expanding stents may result in fewer traumas to the parent vessel and in reduced rates of iatrogenic complications and restenosis. SUMMARY: Intracranial angioplasty with or without stenting is a promising treatment option. Patient selection, careful periprocedural medical management, and a highly skilled neuroendovascular surgeon are all required to perform the procedure with an acceptable risk. If stenting is to be shown to be a safe therapeutic alternative, prospective trials comparing stenting with optimal medical treatment need to be performed. PMID- 15655402 TI - Selection of thrombolytic therapy beyond 3 h using magnetic resonance imaging. AB - PURPOSE OF REVIEW: Use of intravenous thrombolytic therapy in ischaemic stroke is restricted to a 3-h time window because of the proof of this time window in pivotal clinical trials. Thrombolysis is aimed at recanalization of occluded arteries and reperfusion of the ischaemic penumbra, a region of critically hypoperfused, functionally impaired, but potentially viable brain. There are a number of current prospective trials that are testing the hypothesis that the presence of the penumbra will predict thrombolytic responders beyond 3 h. RECENT FINDINGS: Using magnetic resonance imaging, a mismatch between a larger perfusion weighted imaging lesion and smaller diffusion-weighted imaging lesion is considered to represent the ischaemic penumbra. Perfusion-weighted imaging provides semiquantitative cerebral blood flow imaging and diffusion-weighted imaging is an index of the largely irreversible ischaemic core. This definition has been modified with the recognition that the perfusion-weighted imaging lesion includes benign oligaemia and that a portion of the diffusion-weighted imaging core is potentially salvageable with rapid reperfusion. Most acute stroke patients have a magnetic resonance imaging-penumbral signature within 6 h of stroke onset. The penumbra is commonly, but not invariably, associated with proximal arterial occlusion and is time-dependent. Preliminary studies have shown benefit from thrombolytic therapy beyond the established 3-h window. SUMMARY: Penumbral imaging using magnetic resonance imaging with perfusion over diffusion weighted imaging mismatch can provide a physiological 'tissue clock' in individual patients. Based on this hypothesis, a number of prospective trials are being performed. These include EPITHET, DEFUSE, DIAS, MR RESCUE and ROSIE. PMID- 15655403 TI - Anticoagulation and platelet antiaggregation therapy in stroke prevention. AB - PURPOSE OF REVIEW: The results of recent large clinical trials have modified treatment plans formerly based on inferred mechanisms of ischemic stroke and hazards of certain forms of therapy. RECENT FINDINGS: Strong data have emerged to support anticoagulation with warfarin for stroke associated with inferred embolism in a setting of atrial fibrillation. No clear advantage for warfarin over aspirin exists for ischemic stroke in a setting of intracranial atheroma, patent cardiac foramen ovale, or elevated levels of antiphospholipid antibody. Among antiplatelet agents, aspirin and clopidogrel have a similar recurrent stroke risk. Combination therapies with aspirin and warfarin show no additional benefits with regard to stroke prevention and carry higher risks of hemorrhage. Treatment with aspirin combined with specially formulated long-acting dipyridamole carries a lower risk of stroke than aspirin alone and does not increase the risk of hemorrhage significantly. The combination of aspirin and clopidogrel does not reduce the risk of stroke over clopidogrel alone and carries a greater risk of bleeding than clopidogrel alone. SUMMARY: Choice of antithrombotic therapy depends on the etiology of the stroke. Oral anticoagulation treatment is the preferred choice for inferred cardioembolism in the setting of atrial fibrillation, while the varying rates of hemorrhage with oral anticoagulants continue to favor antiplatelet therapy in other settings of inferred etiology. Combinations of antithrombotic therapy vary in their lowering of stroke rate, and some raise the risk of hemorrhage. Insufficient data exist to determine whether antithrombotic therapy combined with antihypertensives, statins or other agents will further reduce the risk of stroke in synergistic or supplemental fashion, or give no additional benefit. PMID- 15655406 TI - A preliminary investigation of the effects of a provider network on costs and lost-time in workers' compensation. AB - OBJECTIVE: We sought to compare lost-time days and average and median workers' compensation claims costs between injured workers managed by OMNET Gold (OG) physicians and those managed by physicians not participating in OG. OG is a statewide health care provider network coordinated by occupational medicine physicians and established by the Louisiana Workers' Compensation Corporation (LWCC) to manage the medical care of injured workers. METHODS: We identified and contrasted 158 lost-time claims managed by OG physicians and 1,323 claims managed by physicians not participating in OG during the first year of network operation (August 1, 2003 to July 31, 2004). RESULTS: The average and median costs for a non-OG claim was $12,542 and $5,793, whereas the average and median costs for an OG claim was $6,749 and $3,015. The average and median number of lost-time days for an OG claim was 53.4 and 34.0 and 95.0 and 58.0 for a non-OG claim. The mean differences were statistically significant. CONCLUSIONS: A small network of physicians may have an effect on the duration of lost-time and workers' compensation costs. PMID- 15655404 TI - Adult stem cell therapy in stroke. AB - PURPOSE OF REVIEW: Acute cerebral infarction causes irreversible locally restricted loss of the neuronal circuitry and supporting glial cells with consecutive functional deficits and disabilities. The currently available and effective therapy targets fast vessel recanalization accompanied by symptomatic measures. Research activities focusing on stem cells, which represent a promising source for organotypic cell replacement and functional recovery after stroke, have gained momentum in recent years, making regenerative cell-based therapies a much more feasible realistic approach. This review provides an update about preclinical and clinical cell-based studies in stroke focusing on stem cells derived from the adult central nervous and hematopoetic systems. RECENT FINDINGS: Endogenous neural stem cells, which have been shown to reside throughout life in the central nervous system, have the capacity to replace lost neurons in models for numerous disorders, including cerebral ischemia. Considering adult neural stem cell transplantation as a regenerative strategy after stroke, progress has been made in isolating human adult neural stem cells and demonstrating the feasibility of autologous neural stem cell transplantation. An increasing number of studies provide evidence that hematopoietic stem cells, either after stimulation of endogenous stem cell pools or after exogenous hematopoietic stem cell application (transplantation), improve functional outcome after ischemic brain lesions. Various underlying mechanisms such as transdifferentiation into neural lineages, neuroprotection through trophic support, and cell fusion have been deciphered. SUMMARY: Many preclinical studies employing adult stem cell based strategies hold great promise. For endogenous approaches the correlate of cell replacement underlying functional improvement needs to be demonstrated. Transplantation approaches on the experimental level need further development before clinical application can be considered. PMID- 15655407 TI - Ibandronate in metastatic bone disease: a review of preclinical data. AB - Bisphosphonates are widely used to prevent and treat skeletal complications of metastatic bone disease. There is increasing evidence that, besides inhibiting osteoclast activity and reducing bone resorption, bisphosphonates also have an anti-tumor effect. This paper reviews the preclinical data for ibandronate. Ibandronate increased the proportion of apoptotic tumor cells in vitro and in vivo, possibly following activation of caspase-like proteases. In vitro, ibandronate also prevented adhesion and spreading of tumor cells to bone, and tumor cell invasion. These inhibitory effects were additive when ibandronate was given with paclitaxel or docetaxel. In animal models of tumor-induced osteolysis, ibandronate significantly reduced the development of osteolytic lesions. Efficacy for the prevention and reduction of bone metastases was related to the timing of treatment; ibandronate treatment initiated prior to or shortly after tumor cell inoculation inhibited the growth of bone metastases and preserved skeletal integrity most effectively. As with other bisphosphonates, the influence of ibandronate on soft tissue metastases has been inconsistent. Overall, preclinical evidence supports the rationale for adjuvant treatment with ibandronate for patients at risk of metastatic bone disease. The renal safety profile of ibandronate supports its suitability for long-term adjuvant use, even with intermittent high dosing. Adjuvant clinical trials have been initiated. The ability of bisphosphonates to preserve skeletal integrity is also of benefit in other clinical settings. Recent studies in rat models demonstrate improved osseointegration of joint implants following ibandronate therapy, with potential application in patients with conditions such as degenerative arthritis or osteoporosis. PMID- 15655408 TI - Arsenic in cancer therapy. AB - Arsenic, a natural substance that has been used as a drug for over 2000 years, has been revived because of its remarkable therapeutic efficacy in patients with acute promyelocytic leukemia (APL). Arsenic exerts a dose-dependent dual effect: it causes differentiation at low concentrations and apoptosis at relatively high concentrations. Specific degradation of the leukemogenic PML-RARalpha fusion protein induced by arsenic leads to the differentiation of leukemia cells. The arsenic-induced apoptosis occurs through direct effects on mitochondria, causing the release of apoptotic proteins into the cytosol and the activation of caspases. Preliminary in vitro studies have also extended the potential anti cancer effect of arsenic to non-APL leukemias, lymphoid malignancies and other cancers. In vitro and in vivo studies demonstrate that arsenic exerts a broad spectrum of anti-cancer effects by induction of apoptosis, inhibition of cell proliferation, anti-angiogenesis and possible immunomodulation. Phase I and II clinical trials are underway to evaluate the feasibility, safety and potential effect of arsenic in various cancer types. PMID- 15655409 TI - The Raf kinase inhibitor BAY 43-9006 reduces cellular uptake of platinum compounds and cytotoxicity in human colorectal carcinoma cell lines. AB - Raf kinase plays a central role in oncogenic signaling and acts as a downstream effector of Ras in the extracellular signal-regulated (ERK) kinase pathway. BAY 43-9006 (BAY) is a novel signal transduction inhibitor that prevents tumor cell proliferation and angiogenesis through blockade of the Raf/MEK/ERK pathway at the level of Raf kinase and the receptor tyrosine kinases vascular endothelial growth factor receptor-2 and platelet-derived growth factor receptor-beta. The present study evaluates the effects of combining BAY and platinum derivatives on human colorectal cancer cells using different incubation protocols. Our data show that the combination of oxaliplatin or cisplatin with BAY results in marked antagonism irrespective of the used application schedule. Furthermore, BAY abrogates the cisplatin-induced G2 arrest as well as the G1 arrest induced by oxaliplatin. BAY alone arrests cancer cells in their current cell cycle phase and affects cell cycle regulative genes. Specifically, BAY reduced the protein expression of p21Cip1 as well as cyclin D1, and inhibits the expression of cdc2 (cdk1). Utilizing atom absorption spectrometry, BAY significantly reduced cellular uptake of platinum compounds and thereby the generation of DNA adducts. Taken together, co-incubation with BAY results in reduced cellular uptake of platinum compounds and consecutively reduced generation of DNA adducts, and eventually decreased cellular cytotoxicity in human colorectal cancer cells. Our results indicate that the Raf kinase inhibitor BAY 43-9006 might also directly or indirectly interact with platinum transporter proteins in vitro. PMID- 15655410 TI - The antitumor drug candidate 2-(4-amino-3-methylphenyl)-5-fluorobenzothiazole induces NF-kappaB activity in drug-sensitive MCF-7 cells. AB - 2-(4-Amino-3-methylphenyl)-5-fluoro-benzothiazole (5F 203) potently inhibits MCF 7 breast cancer cell growth in part by activating the aryl hydrocarbon receptor (AhR) signaling pathway. Ligands for the AhR (i.e. dioxin) have also been shown to modulate the NF-kappaB signaling cascade, affecting physiological processes such as cellular immunity, inflammation, proliferation and survival. The objective of this study was to investigate the effect of 5F 203 treatment on the NF-kappaB signaling pathway in breast cancer cells. Exposure of MCF-7 cells to 5F 203 increased protein-DNA complex formation on the NF-kappaB-responsive element as determined by electrophoretic mobility shift assay, but this effect was eliminated in MDA-MB-435 cells, which are resistant to the antiproliferative effects of 5F 203. An increase in NF-kappaB-dependent transcriptional activity was confirmed by a significant increase in NF-kappaB-dependent reporter activity in sensitive MCF-7 cells, which was absent in resistant MDA-MB-435 cells and AhR deficient subclones of MCF-7 cells. Inhibition of NF-kappaB activation enhanced the increase in xenobiotic response element-dependent reporter activity in MCF-7 cells when treated with 5F 203. The drug candidate 5F 203 also induced mRNA levels of IL-6, an NF-kappaB-responsive gene, in MCF-7 cells, but not in MDA-MB 435 cells, as determined by quantitative RT-PCR. These findings suggest that 5F 203 activation of the NF-kappaB signaling cascade may contribute to 5F 203 mediated anticancer activity in human breast cancer MCF-7 cells. PMID- 15655411 TI - Cytotoxicities of three rebeccamycin derivatives in the National Cancer Institute screening of 60 human tumor cell lines. AB - Among the biologically active indolocarbazoles, rebeccamycin, a microbial metabolite produced by Saccharothrix aerocolonigenes, is a well-known topoisomerase (Topo) I poison. In the course of structure-activity relationship studies on rebeccamycin analogs, we have prepared a large number of indolocarbazole derivatives and have shown that, depending on the structural modifications, the cytotoxic effects may be, or not, directly correlated to DNA binding and Topo I inhibition. This suggests that if DNA binding and Topo I play a part in the biological activity of these compounds, other cellular targets might be involved. This paper reports the results of the antiproliferative activities (evaluated in the National Cancer Institute's in vitro panel of 60 tumor cell lines) and the results of a COMPARE analysis run with rebeccamycin derivatives to identify other potential biological targets for these compounds. PMID- 15655412 TI - Effects of retinoic acid steroidal analogs on human leukemic HL60 cell proliferation in vitro and on angiogenesis in vivo. AB - Retinoic acid (RA) can be regarded as a pharmacological agent commonly used for its ability to affect growth and differentiation of a variety of cell types, such as acute promyelocytic leukemic and endothelial cells. In the present work we studied the effect of all-trans-RA (ATRA) and its steroidal analogs EA-4, EA-136 and EA-137 on the growth of human promyelocytic HL-60 cells in vitro. The specific steroidal substrates were chosen in order to further investigate their ability to improve the pharmacological properties of conjugated antileukemic agents. ATRA decreased the number of HL60 cells from the first 24 h after its addition to the cell culture medium. The decrease was significant at concentrations higher than 10(-5) M. All the analogs tested also decreased the number of HL60 cells with an IC50 similar to that of ATRA, except for EA-4 whose IC50 was almost two orders of magnitude lower than that of ATRA, 72 h after its addition to the cell culture medium. Since angiogenesis is important for the growth of hematological malignancies, we furthermore studied the effect of ATRA and its analogs on the formation of new capillaries in the in vivo chicken embryo chorioallantoic membrane (CAM). ATRA, EA-136 and EA-137 induced angiogenesis in the CAM, increased the layer of CAM keratinocytes, and resulted in a significant degree of extravasation. EA-4 had no effect on either angiogenesis or tissue structure in general. It seems that the retinoid EA-4 is a promising agent for the inhibition of human leukemia cell growth. PMID- 15655413 TI - Antagonists of bombesin/gastrin-releasing peptide decrease the expression of angiogenic and anti-apoptotic factors in human glioblastoma. AB - We have investigated the antitumor effects and the mechanism of action of antagonists of bombesin/gastrin-releasing peptide (GRP), RC-3940-II and RC-3940 Et, on the growth of U-118MG human malignant glioma xenografted into nude mice. Tumors volume was measured weekly, and after 6 weeks of treatment with GRP antagonists the tumors were analyzed by Western blot assays for the expression of vascular endothelial growth factor (VEGF), protein kinase C (PKC)-alpha, the anti apoptotic protein Bcl-2 and the pro-apoptotic protein Bax. A radioreceptor assay was used to characterize the receptors for bombesin/GRP. Specific high-affinity receptors for bombesin were found in U-118MG tumors, and their growth was reduced by 52.5% by RC-3940-II and 72.6% by RC-3940-Et (both p<0.01). The tumor doubling time was prolonged by 4.6 and 12 days after treatment with RC-3940-II and RC-3940 Et, respectively, compared to controls (p<0.05). Both antagonists caused a significant (p<0.05) decrease of about 28% in the levels of VEGF protein and a reduction of approximately 35% in the expression of PKCalpha. The relative ratio of Bcl-2:Bax was also diminished by around 70% by both analogs, indicating a net apoptotic gain and the efficacy of treatment. Our results suggest that bombesin/GRP antagonists, RC-3940-II and RC-3940-Et, could be of value for the treatment of human glioblastomas. PMID- 15655414 TI - Enhanced cytotoxicity of bioreductive antitumor agents with dimethyl fumarate in human glioblastoma cells. AB - We compared the cytotoxicity of the bioreductive antitumor agents mitomycin C (MMC) and streptonigrin (SN) with or without the DT-diaphorase (DTD) inducer dimethyl fumarate (DMF) in four human glioblastoma cell lines with the conventional chemotherapeutic agent, 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU). We also examined four other types of cancer cells to compare with glioblastoma cells. Cytotoxicity was measured with the sulforhodamine B (SRB) assay and was represented by 50% inhibition concentration (IC50). Enzymatic activities of DTD, cytochrome b5 reductase and glutathione-S-transferase (GST) in cells were measured spectrophotometrically. IC50 for BCNU was in a range of 28-300 microM in the glioblastoma cell lines. Glioblastoma cells were more sensitive to MMC or SN than to BCNU. Pretreatment with DMF significantly increased cytotoxicity of MMC and SN in glioblastoma cell lines and the NCI-H1299 lung cancer cell line, but had no effect on BCNU cytotoxicity. DMF significantly increased DTD and cytochrome b5 reductase activity, and decreased GST in three of four glioblastoma cell lines. Addition of the DTD inhibitor, dicumarol, significantly inhibited cytotoxicity of MMC and SN, and reversed the increased cytotoxicity seen when DMF was combined with either MMC or SN in all glioblastoma cell lines. Combining inducers of DTD and cytochrome b5 reductase with bioreductive agents may be a potential therapeutic strategy for glioblastoma. PMID- 15655415 TI - Compound 278E, structurally modified from tanshinone, induces monocytic differentiation and regulates proto-oncogene expression in human leukemic HL-60 cells. AB - Tanshinone derivative compounds, isolated from Salvia miltiorrhiza Bunge (Labiatae), have been reported as microtubule inhibitors with antimitotic activity. In this study, we examined the growth-inhibiting and differentiation inducing effect of these compounds on human leukemic HL-60 cells. The expression of protein kinase C (PKC) and proto-oncogenes in 278E-treated cells was also assessed. All tanshinone derivative compounds exhibited growth-inhibitory effects on HL-60 cells, but only 278E induced cell differentiation. Morphological observation of 278E-treated HL-60 cells showed a greater percentage of monocytes and macrophages (Mo/Mphi). Treatment with 5 microg/ml 278E resulted in a marked increase in the percentages of superoxide-producing (up to 95.5+/-1.8%) and non specific esterase-positive cells (up to 80.3+/-9.1%). The differentiated cells also expressed cell surface antigens characteristic of Mo/Mphi, including CD11b, CD14 and CD68. Neither cellular changes in isozymes of PKC nor translocation of these isozymes from cytosol to cell membrane were seen in 278E-treated HL-60 cells. 278E caused a downregulation of c-myc as well as an up-regulation of c fms, c-jun and c-fos. PMID- 15655416 TI - Single-agent trastuzumab versus trastuzumab plus cytotoxic chemotherapy in metastatic breast cancer: a single-institution experience. AB - Trastuzumab has shown significant single-agent activity in patients with Her 2/neu overexpressing metastatic breast cancer, and increased response rates, progression-free and overall survival when added to standard chemotherapy. Despite higher response rates, the combination with chemotherapy has higher toxicity and it remains unknown whether single-agent trastuzumab is equally effective as the combined treatment in terms of progression-free and overall survival. We therefore carried out a retrospective multivariate analysis of 117 patients with Her-2/neu overexpressing metastatic breast cancer who were treated with trastuzumab with or without chemotherapy at a single institution between November 1999 and December 2003. Response rates tended to be higher in patients receiving trastuzumab in combination with chemotherapy (34 versus 8%, p=0.060). However, this did not translate into a benefit in progression-free survival: median (95% confidence interval) progression-free survival was 6.2 (4.45-7.95) months in patients receiving trastuzumab plus chemotherapy versus 4.2 (1.77-6.63) months in those receiving single-agent trastuzumab (p=0.560). Likewise, no significant difference in overall survival was observed: 27.0 (19.9-34.0) versus 23.1 (16.2-30.0) months (p=0.809). We conclude that in the absence of extensive visceral involvement necessitating a higher response rate, single-agent trastuzumab may be a safe and less-toxic alternative to its combined use with other chemotherapy agents. This needs to be confirmed in prospective randomized trials. PMID- 15655417 TI - Treatment of non-small cell lung cancer with gefitinib ('Iressa', ZD1839): the Greek experience with a compassionate-use program. AB - This is a retrospective analysis of 150 patients with advanced non-small cell lung cancer who had failed prior treatment or were unfit for chemotherapy and were treated with oral gefitinib ('Iressa', ZD1839; AstraZeneca) 250 mg/day. Thirty-two patients who received gefitinib for 3 weeks or less were not included in the analysis. For the remaining 118 evaluable patients, the mean age was 63.1 years; most patients had received prior chemotherapy (97.5%), Eastern Cooperative Oncology Group performance status scores 0-2 (97.4%) and stage IV disease (64.4%). The majority were symptomatic (84.6%). Disease control was observed in 30 patients (25.4%), of whom five had a partial response and 25 had stable disease; 18 (15.3%) were not evaluable. Median duration of treatment was 29.9 weeks in responding patients and 11.5 in patients with progressive disease (p<0.0001). Median overall survival was 7.3 months (15.2 months for disease control) and median progression-free survival was 3.2 months. Gefitinib was well tolerated, with grade 3/4 skin rash and diarrhea seen in 2.5 and 4.2% of patients, respectively. Clinical benefit was evaluated using questionnaires before and following treatment with gefitinib. In 82 patients with completed questionnaires, evaluation revealed symptom improvement in 40.1% and improvement in general feeling in 31.4%. Epidermal growth factor receptor (EGFR) analysis found that efficacy did not correlate with tumor EGFR overexpression. Therefore, in this retrospective analysis, gefitinib treatment provided disease control in 25% of patients who derived significant palliative benefit. PMID- 15655418 TI - Long-term remission of excessive liver metastases in a breast cancer patient with chronic alcohol abuse using a monotherapy with trastuzumab. AB - We report on the successful treatment of a 43-year-old breast cancer patient with excessive liver metastases and chronic alcohol abuse. After first occurrence of hepatic metastases, systemic and interventional therapies were performed, and resulted in short-term partial remission. Finally, an excessive progression of the hepatic metastases was diagnosed. A systemic therapy with weekly trastuzumab (Herceptin) infusions was induced and a complete remission was achieved that is ongoing now for over 45 months. PMID- 15655419 TI - Inhibition of breast adenocarcinoma growth by intratumoral injection of lipophilic long-acting lathyrogens. AB - Prevention of the formation of crosslinks and/or disintegration of already formed collagen fibrils in the tumor by known lathyrogens, beta-aminopropionitrile or D penicillamine, may result in the weakening of tumor support, decreasing angiogenesis and promoting tumor regression. This paper reviews our studies with a single intratumoral injection of lipophilic lathyrogens and others, using a systemic administration to investigate the effect of both lathyrogens. Details of our experimental results are also given. PMID- 15655420 TI - New drugs in cancer therapy, National Tumor Institute, Naples, 17-18 June 2004. AB - An international meeting on 'New Drugs in Cancer Therapy' was held at the National Tumor Institute of Naples, on 17-18 June 2004. The first session of the meeting focused on analogs of conventional anti-cancer drugs, such as taxanes, platinum compounds, anthracyclines and topoisomerase I inhibitors. The data of a phase II trial of BMS-247550, an epothilone B analog, in patients with renal cell carcinoma were reported. Data were also presented on BBR-3464, a trinucleate platinum analog which was developed on the grounds of greater potency, a more rapid rate of DNA binding and the ability to induce apoptosis regardless of the p53 status of the cell. Pegylated-coated liposomal formulation doxorubicin (Caelyx) has shown efficacy in metastatic breast cancer and in advanced ovarian cancer; sabarubicin is a third-generation anthracycline with equal or superior potency to doxorubicin or idarubicin in a variety of human tumor cell lines of different histotypes. The main mechanisms of resistance to topoisomerase I inhibitors were discussed; data on diflomotecan were reported, showing a narrow therapeutic index of the drug. The second session of the meeting focused on the ErbB family as a target for anti-cancer therapy. Recent evidence of a correlation between epidermal growth factor receptor (EGFR) mutations at exons 18-21 and clinical response of advanced non-small cell lung cancer to gefitinib therapy was commented on. The issue of the association between ErbB2 expression and gefitinib activity was addressed, while clinical data of a phase II study of gefitinib in advanced breast cancer were presented. Monoclonal antibodies targeting EGFR represent another worthwhile way to interfere with EGFR-driven signal transduction. Cetuximab is reaching market registration in advanced colorectal cancer; in particular, due to the results of the BOND study. The recently presented results of the Bonner study strongly support the activity of this drug in head and neck cancer. A step forward in the research on anti-EGFR monoclonal antibodies may be represented by humanized monoclonal antibodies, such as EMD 72000 and ABX-EGF. Imatinib mesylate is probably the most outstanding example of an effective targeted therapy--its activity in gastrointestinal stromal tumors was so exciting that the drug reached the market without undergoing phase III evaluation. The third session of the meeting was on angiogenesis inhibitors. Drugs may interfere with the angiogenic process via different mechanisms and there is a sound rationale for combining anti-angiogenic agents with chemotherapy or multiple anti-angiogenic strategies. Clinical results obtained with direct anti-angiogenic agents have been negative up to now, but some exciting results have been seen with bevacizumab, a monoclonal antibody targeting vascular endothelial growth factor (VEGF). A few VEGF-tyrosine kinase inhibiting small molecules, such as ZD6474, AZD2171 and PTK/ZK, are undergoing clinical trials. The fourth session of the meeting was on interference with intracellular signal transduction. Farnesyl transferase inhibitors exert their action by interfering with either pro-Ras or RhoB farnesylation. Several clinical studies of different phases with compounds belonging to this class have been carried out, either alone or in combination with chemotherapy; unfortunately, all of them have turned out to be negative. Cell cycle inhibitors, such as CYC-202 and BMS-387032, represent a class of interesting compounds which are in the early phase of development and whose clinical results are eagerly awaited. Another strategy to achieve cell cycle inhibition is to target heat shock protein 90, a molecular chaperone required for protein folding. Clinical data on depsipeptide, a histone deacetylase (HDAC) inhibitor with activity in T cell lymphoma, were presented. Suberoylanilide hydroxamic acid is another small molecular weight inhibitor of HDAC activity. Phase I/II clinical trials have shown low toxicity and evidence of anti-tumor activity; on the other hand, this compound has potential for synergism with radiotherapy, chemotherapy and biologicals. PMID- 15655422 TI - Consumer complaints and nursing home quality. PMID- 15655423 TI - Nursing home consumer complaints and their potential role in assessing quality of care. AB - BACKGROUND: State survey agencies collect and investigate consumer complaints for care in nursing homes and other health care settings. Complaint investigations play a key role in quality assurance, because they can respond to concerns of consumers and families. OBJECTIVE: This study uses 5 years of nursing home complaints data from Massachusetts (1998-2002) to investigate whether complaints might be used to assess nursing home quality of care. RESEARCH DESIGN: The investigator matches facility-level complaints data with On-Line Survey Certification and Reporting (OSCAR) data and Minimum Data Set Quality Indicator (MDS QI) data to evaluate the association between consumer complaints, facility and resident characteristics, and other nursing home quality measures. RESULTS: Consumer complaints varied across facility characteristics in ways consistent with the nursing home quality literature. Complaints were consistently and significantly associated with survey deficiencies, the presence of a serious survey deficiency, and nurse aide staffing. Complaints were not significantly associated with nurse staffing, and associations with 6 MDS QIs were mixed. The number of complaints was significantly predictive of survey deficiencies identified at the subsequent inspection. CONCLUSION: Nursing home consumer complaints provide a supplemental tool with which to differentiate nursing homes on quality. Despite limitations, complaints data have potential strengths when used in combination with other quality measures. The potential of using consumer complaints to assess nursing home quality of care should be evaluated in states beyond Massachusetts. Evaluating consumer complaints also might be a productive area of inquiry for other health care settings such as hospitals and home health agencies. PMID- 15655424 TI - Regional variation and other correlates of Department of Veterans Affairs Disability Awards for patients with posttraumatic stress disorder. AB - BACKGROUND: Minnesota, Minneapolis, Minnesota. Posttraumatic stress disorder (PTSD) is a chronic disabling condition affecting more than 600,000 United States veterans and is the most common psychiatric condition for which veterans seek Veterans Affairs disability benefits. Receipt of such benefits enhances veterans' access to Veteran Affairs health care and reduces their chance of poverty. OBJECTIVES: We sought to determine whether previously identified regional variations in PTSD disability awards are explained by appropriate subject characteristics (eg, differences in PTSD symptomatology or dysfunction) and to estimate the impact of veterans' PTSD symptom severity or level of dysfunction on their odds of obtaining PTSD disability benefits. RESEARCH DESIGN: We used a mailed survey linked to administrative data. SUBJECTS: Subjects included 4918 representative, eligible men and women who filed PTSD disability claims between 1994 and 1998. RESULTS: A total of 3337 veterans returned useable surveys (68%). Before adjustment, PTSD disability claims approval rates ranged from 43% to 75% across regions. After adjustment, rates ranged from 33% to 72% (P <0.0001). Severer PTSD symptoms were associated with greater odds of having PTSD disability benefits (P <0.0001). Unexpectedly, poorer functional status was associated with lower odds of having benefits (P <0.0001). On average, clinical differences between veterans who did and did not have PTSD disability benefits were small but suggested slightly greater dysfunction among those without benefits. CONCLUSIONS: An almost twofold regional difference in claims approval rates was not explained by veterans' PTSD symptom severity, level of dysfunction, or other subject-level characteristics. Veterans who did not obtain PTSD disability benefits were at least as disabled as those who did receive benefits. PMID- 15655425 TI - Changes in prognosis after the first postoperative complication. AB - BACKGROUND: Postoperative complications are common in the Medicare population, yet no study has formally quantified the change in prognosis that occurs after a broad range of first complications. OBJECTIVE: We sought to estimate the relative severity of 24 first postoperative complications. RESEARCH DESIGN: We undertook a multivariate matched, population-based, case-control study of death after surgery in a sample of 1362 Pennsylvania Medicare patients. SUBJECTS: Cases (681 deaths) were selected randomly using claims from 1995-1996. Models were developed to scan all Pennsylvania claims, looking for similar controls that did not die. MEASURES: Charts were abstracted, complications identified, and models were constructed to estimate the odds of dying after any 1 of 24 first postoperative complications. RESULTS: The odds of dying within 60 days increased 3.4-fold (95% confidence interval [CI] 2.5-4.7) in patients with complications as compared with those without complications. A first complication of respiratory compromise was associated with a 7.2-fold increase in the odds of dying (95% CI 4.5-11.6). The first complications of pneumonia or congestive heart failure were associated with, respectively, 5-fold (95% CI 2.1-12.1) and 5.1-fold (2.3-11.1) increases in odds of dying as compared with no complication. CONCLUSIONS: First complications after surgery, even seemingly mild ones, may radically alter the patient's risk of death. First complications often begin the cascade of complications that end in death. Caregivers should consider the first complication as a timely signal of a changed clinical situation demanding a reevaluation of the patient's care. Researchers may use these estimates to determine the relative severity of a broad range of first or early complications. PMID- 15655426 TI - Comorbidity as a predictor of stage of illness for patients with breast cancer. AB - OBJECTIVE: The purpose of this research was to determine whether comorbidity affects the stage at which breast cancer is diagnosed. METHODS: Data from the Surveillance, Epidemiology and End Results (SEER) program of the National Cancer Institute (NCI) was merged with Medicare claims for 17,468 women diagnosed with breast cancer from 1993 to 1995. RESULTS: Women with cardiovascular disease, musculoskeletal disorders, mild-to-moderate gastrointestinal disease, and nonmalignant benign breast disease had a 13%, 7%, 14%, and 24% lower odds, respectively, of being diagnosed with advanced breast cancer. Women with diabetes, other endocrine disorders, psychiatric disorders, or hematologic disorders increased the odds of a late-stage diagnosis by 19%, 11%, 20%, and 19% respectively. Mammography screening and contact with the medical care system decreased the odds of late-stage diagnosis. DISCUSSION: Four hypotheses are suggested to explain this link between comorbid illness and stage at diagnosis: (1) the "surveillance" hypothesis, (2) the "physiological" hypothesis, (3) the "competing demand" hypothesis, and (4) the "death from other causes" hypothesis. CONCLUSIONS: Comorbidity may complicate the diagnostic decision-making process for breast cancer. The results suggest that contact with the medical care system improves the odds of early-stage diagnosis. Thus, barriers to access for people with chronic conditions may exacerbate those chronic conditions and increase the odds of late-stage breast cancer. PMID- 15655427 TI - Racial inequities in the timing of breast cancer detection, diagnosis, and initiation of treatment. AB - BACKGROUND: Recent studies suggest differences in quality and timeliness of care received may be major contributing sources to the racial disparity in breast cancer detection and related outcomes. METHODS: Female patients with breast cancer diagnosed during 1985-1993 (n=400) and followed through June 20, 2001, were included in this retrospective cohort study. Three white patients were selected randomly and matched to each black patient by year of diagnosis. Method and timing of diagnosis and timing of treatment were abstracted from medical records. Initial staging and subsequent breast cancer recurrence and vital status were obtained from the Hospital and Connecticut State Tumor Registry. RESULTS: Black women were more likely than white women to be diagnosed after a patient noted abnormality. Black women were less likely than white women to have completed a diagnostic evaluation within 30 days after a patient-noted abnormality (P <0.01) or after having an abnormality noted on screening mammogram (P=0.0001) and were less likely to have initiated treatment within 30 days of diagnosis (P=0.0001). Women diagnosed after a patient-noted abnormality were more likely to have subsequent breast cancer recurrence and/or death due to breast cancer compared with women diagnosed after a screening mammogram (56% versus 24%, respectively, P <0.05). CONCLUSIONS: Racial differences were identified at each step in the evaluation and treatment clinical pathway, including method of detection, timing from first symptoms of cancer to pathologic diagnosis, and timing from diagnosis to initiation of treatment. The findings highlight the need to provide equal opportunity for timely medical care and treatment. PMID- 15655428 TI - Mortality after cardiac bypass surgery: prediction from administrative versus clinical data. AB - BACKGROUND: Risk-adjusted outcome rates frequently are used to make inferences about hospital quality of care. We calculated risk-adjusted mortality rates in veterans undergoing isolated coronary artery bypass surgery (CABS) from administrative data and from chart-based clinical data and compared the assessment of hospital high and low outlier status for mortality that results from these 2 data sources. STUDY POPULATION: We studied veterans who underwent CABS in 43 VA hospitals between October 1, 1993, and March 30, 1996 (n=15,288). METHODS: To evaluate administrative data, we entered 6 groups of International Classification of Diseases (ICD)-9-CM codes for comorbid diagnoses from the VA Patient Treatment File (PTF) into a logistic regression model predicting postoperative mortality. We also evaluated counts of comorbid ICD-9-CM codes within each group, along with 3 common principal diagnoses, weekend admission or surgery, major procedures associated with CABS, and demographic variables. Data from the VA Continuous Improvement in Cardiac Surgery Program (CICSP) were used to create a separate clinical model predicting postoperative mortality. For each hospital, an observed-to-expected (O/E) ratio of mortality was calculated from (1) the PTF model and (2) the CICSP model. We defined outlier status as an O/E ratio outside of 1.0 (based on the hospital's 90% confidence interval). To improve the statistical and predictive power of the PTF model, selected clinical variables from CICSP were added to it and outlier status reassessed. RESULTS: Significant predictors of postoperative mortality in the PTF model included 1 group of comorbid ICD-9-CM codes, intraortic balloon pump insertion before CABS, angioplasty on the day of or before CABS, weekend surgery, and a principal diagnosis of other forms of ischemic heart disease. The model's c-index was 0.698. As expected, the CICSP model's predictive power was significantly greater than that of the administrative model (c=0.761). The addition of just 2 CICSP variables to the PTF model improved its predictive power (c=0.741). This model identified 5 of 6 high mortality outliers identified by the CICSP model. Additional CICSP variables were statistically significant predictors but did not improve the assessment of high outlier status. CONCLUSIONS: Models using administrative data to predict postoperative mortality can be improved with the addition of a very small number of clinical variables. Limited clinical improvements of administrative data may make it suitable for use in quality improvement efforts. PMID- 15655429 TI - Mortality after noncardiac surgery: prediction from administrative versus clinical data. AB - BACKGROUND: Hospital profiles are increasingly constructed using risk-adjusted clinical data abstracted from patient records. OBJECTIVE: We sought to compare hospital profiles based on risk adjusted death within 30 days of surgery from administrative versus clinical data in a national cohort of surgical patients. DESIGN: This was a cohort study that included 78,546 major noncardiac operations performed between October 1, 1991 and December 31, 1993 in 44 Veterans Affairs hospitals. Administrative data were used to develop and validate multivariable logistic regression models of 30-day postoperative death for all surgery and 4 surgical specialties (general, orthopedic, thoracic, and vascular). Previously developed and validated clinical models were obtained and reproduced for matching operations using data from the VA National Surgical Quality Improvement Program. Observed-to-expected 30-day mortality ratios for administrative and clinical data were calculated and compared for each hospital. RESULTS: In multivariable logistic regression models using administrative data, characteristics such as patient age, race, marital status, admission from a nursing home, interhospital transfer, admission on the weekend, weekend surgery, and risk strata consisting of groups of principal and comorbidity diagnoses were predictive of postoperative mortality (P <0.05). Correlations of the clinical and administrative observed-to expected ratios were 0.75, 0.83, 0.64, 0.78, and 0.86 for all surgery, general, orthopedic, thoracic, and vascular surgery, respectively. When compared with clinical models, administrative models identified outlier hospitals with sensitivity of 73%, specificity of 89%, positive predictive value of 51%, and negative predictive value of 96%. CONCLUSIONS: Our data suggest that risk adjustment of mortality using administrative data may be useful for screening hospitals for potential quality problems. PMID- 15655430 TI - The impact of the SARS outbreak on an urban emergency department in Taiwan. AB - OBJECTIVE: Emergency departments (ED) were on the front lines for possible cases of transmission during the severe acute respiratory syndrome (SARS) epidemic. The purpose of this study was to investigate the impact of the SARS catastrophe on an urban ED. METHODS: The patients' characteristics in an urban ED were collected from March to May 2003 during the SARS outbreak in Taiwan. The crisis period was divided into 2 periods: 30 days before (period 1) and after (period 2) April 21, the date of the first hospital-associated transmission. Problem severity in the ED and stress levels of ED staff during the SARS catastrophe were rated from mild (1 point) to severe (5 points). RESULTS: The number of ED patients declined 33.4% in period 2. There was a 2.1% (95%CI, 0.4-3.8) increase in the percentage of male patients, a 2.5% (95% CI, 1.5-3.7) increase in percentage of fever (>38 degrees C), and a 4.0% (95% CI, 2.6-5.4%) increase in chief complaint of fever in period 2. The number of nontrauma patients younger than 18 years had declined by 44.5% in period 2. The total charge for reimbursement from an insurance institution declined 21.7%. During the SARS outbreak, the most severe stress experienced by either physicians or nurses occurred during emergency resuscitation (median stress rating point, 4; interquartile range, 1). CONCLUSION: The SARS catastrophe affected the ED visit volume, finances, various patient characteristics, and stress levels in the ED physicians and nurses. EDs must be fully prepared to face the challenges of the next outbreak of SARS or other infectious disease. PMID- 15655431 TI - Patient utilities for advanced cancer: effect of current health on values. AB - BACKGROUND: Utilities are critical inputs to economic analyses, yet some things remain unclear about their elicitation and application, particularly the impact of health status on values. Prospect theory predicts that current health affects values, and that losses loom larger than gains. If true, the use of utilities requires that additional complexities be considered. OBJECTIVE: To determine the effect of current health on patient utilities for advanced cancer health states. RESEARCH DESIGN: The research was a cross-sectional survey (n=100 patients) of utilities for 4 hypothetical advanced cancer health states. Chained gamble utilities for gains and losses in health were compared, correlations between current health status and utilities were measured, and patient utilities for experienced versus hypothetical health states were compared. RESULTS: In this sample, gains in health were valued equivalently to losses, health status was not correlated with utility values, and patients' valuation of states equivalent to their current health did not differ from valuations of the same states by patients with better or worse current health. CONCLUSION: This research confirms the uncertainty surrounding the effect of health status on utilities and the question of whose values to use in analyses. The findings suggest that values for health states may be independent of current health status, supporting an objective view of utilities. This research also suggests that patients may be able to provide "experienced utilities" for states other than their own, expanding the population from whom such values can be elicited. These results may dispute prospect theory's predictions regarding health state valuations. PMID- 15655432 TI - Comparison of coding of heart failure and comorbidities in administrative and clinical data for use in outcomes research. AB - BACKGROUND: Despite the potential usefulness of administrative databases for evaluating outcomes, coding of heart failure and associated comorbidities have not been definitively compared with clinical data. OBJECTIVE: To compare the predictive value of heart failure diagnoses and secondary conditions identified in a large administrative database with chart-based records. METHODS: The authors studied 1808 patient records sampled from 14 acute care hospitals and compared clinically recorded data with administrative records from the Canadian Institute for Health Information. The impact of comorbidity coding in the administrative data set according to the Charlson classification was examined in models of 30 day mortality. RESULTS: The positive predictive value (PPV) of a primary diagnosis ICD-9 428 was 94.3% using the Framingham criteria and 88.6% using criteria previously validated with pulmonary capillary wedge pressure. There was reduced prevalence of secondary comorbid conditions in administrative data in comparison with clinical chart data. The specificities and PPV/negative predictive values of administratively identified index comorbidities were high. The sensitivities of index comorbidities were low, but were enhanced by examination of hospitalizations within 1 year prior to the index heart failure admission. Using information from prior hospitalizations modestly enhanced 30-day mortality model performance; however, the odds ratio point estimates of the index and enhanced administrative data sets were consistent with the clinical model. CONCLUSION: The ICD-9 428 primary diagnosis is highly predictive of heart failure using clinical criteria. Examination of hospitalization data up to 1 year prior to the index admission improves comorbidity detection and may provide enhancements to future studies of heart failure mortality. PMID- 15655433 TI - Risk classification of adult primary care patients by self-reported quality of life. AB - BACKGROUND: : Although patient-reported health-related quality of life (HRQOL) is known to predict health services utilization, most risk assessment systems use provider-reported diagnoses as predictors rather than HRQOL. OBJECTIVE: : We sought to classify adult primary care patients prospectively by utilization risk based on age, gender, and HRQOL at a single clinic visit. RESEARCH DESIGN: : Patients completed the Duke Health Profile. Providers completed the Duke Severity of Illness Checklist. Diagnoses were grouped with the Ambulatory Care Groups system. Predictive coefficients for 1-year primary care charges calculated from the age, gender, and HRQOL of 728 reference patients were used to classify 474 test patients into 4 risk classes. Comparisons were made with models that used diagnoses or severity of illness as predictors. RESULTS: : The positive likelihood ratio for predicting highest risk was 2.2 for the HRQOL model, compared with 1.8 for the diagnoses model, 1.6 for the severity model, and 1.5 for age and gender alone. One-year actual primary care visits and charges increased step-wise from lowest to highest risk class. Highest risk patients were older and more likely to be women, black, or Medicaid recipients. Although the highest-risk patients represented only 18.6% of the test group, they accounted for 26.7% of the primary care clinic visits, 31.6% of the clinic charges, 34.6% of the hospital days, 35.1% of hospital charges, and 30.8% of total charges at all healthcare sites. CONCLUSION: : The HRQOL risk classification system can identify primary care patients at risk for high future health services utilization. PMID- 15655434 TI - Vulnerability and the receipt of recommended preventive services: the influence of multiple risk factors. AB - CONTEXT: Previous studies have confirmed the independent associations of race/ethnicity, socioeconomic status, and potential access with the receipt of preventive care. More pragmatic models of vulnerability are needed to examine the concomitant influence of multiple risk factors. OBJECTIVE: To operationalize vulnerability as risk profiles of predisposing (race/ethnicity and education) and enabling (eg, income, health insurance, and having a regular source of care) factors, and their association with the receipt of preventive care. STUDY DESIGN: Cross-sectional data on 14,983 adults from the Household Component of the 1996 Medical Expenditure Panel Survey. MAIN OUTCOME MEASURES: Receipt of recommended preventive care: blood pressure and cholesterol screening, flu shot, Papanicolaou test, mammogram, and dental visit. RESULTS: Controlling for other factors, analyses of risk profiles revealed a clear dose-response relationship with the receipt of preventive care regardless of race/ethnicity. In the total sample, having more risk factors was associated with a lower prevalence of, for example, receiving a cholesterol screening: 1 risk (PR=0.77; CI, 0.71-0.84), 2 risks (PR=0.56; CI, 0.49-0.64), and 3+ risks (PR=0.34; CI, 0.25-0.43). CONCLUSION: Sizeable disparities in the receipt of recommended preventive services were found in relation to increasing vulnerability risk profiles. Without attention to such co-occurring risks, it is unlikely that substantial gains will be made in reducing disparities in the incidence of and mortality from the most common preventable diseases in the United States. PMID- 15655435 TI - Immunosuppression for posterior uveitis. PMID- 15655436 TI - Angiogenesis, choroidal neovascularization, and the coagulation system. PMID- 15655437 TI - Photodynamic therapy for juxtafoveal choroidal neovascularization due to ocular histoplasmosis syndrome. AB - PURPOSE: To report the use of photodynamic therapy with verteporfin in patients with juxtafoveal choroidal neovascularization (CNV) for ocular histoplasmosis syndrome (OHS). METHODS: Retrospective review. Data regarding the following variables were extracted from patient charts: demographic characteristics, previous surgeries, angiographic features, number and time of treatments, follow up time, and change in visual acuity. RESULTS: This study sample consisted of 23 eyes of 23 consecutive patients who were treated with photodynamic therapy for the management of juxtafoveal CNV. When post-treatment visual acuity (mean logMAR acuity=0.321) was compared to baseline acuity (mean logMAR visual acuity=3.89) vision improved by more than three Snellen lines in 30% of eyes, remained the same (+/-2 Snellen lines) in 52% of eyes, and worsened (greater than a two-line loss in visual acuity) in 18% of eyes. Although this series was uncontrolled, the patients had a trend toward a therapeutic benefit when compared to published natural history of similar cases (OR=0.292, P value=0.071 when compared to data from the Macular Photocoagulation Study for treatment of juxtafoveal lesions). CONCLUSION: Photodynamic therapy with verteporfin may be beneficial in patients with juxtafoveal CNV secondary to OHS in terms of both visual stabilization and improvement. PMID- 15655438 TI - Photodynamic therapy with verteporfin for peripapillary choroidal neovascularization. AB - PURPOSE: To evaluate the efficacy of photodynamic therapy with verteporfin in the management of symptomatic extrafoveal peripapillary choroidal neovascularization (CNV). METHODS: Seven eyes of seven patients with symptomatic peripapillary CNV underwent visual acuity testing, ophthalmic examination, color photography, and fluorescein angiography to evaluate the results of photodynamic therapy with verteporfin. Patients were offered treatment following the development of hemorrhage, subretinal fluid, or lipid if it was associated with visual symptoms. A standard nomogram was used to dose Visudyne, application was performed in three separate 30-second zones confluent over the extent of the lesion. The light dose per unit area applied to the retina was approximately 18 J/cm. RESULTS: Five of the seven had CNV limited to the peripapillary area associated with age-related macular degeneration; the remaining two eyes had presumed ocular histoplasmosis syndrome. In five of the seven eyes, two treatments were needed and in two eyes only one treatment was necessary to elicit resolution of active leakage. Retreatment was performed at an average of 76 days after initial treatment. Baseline best-corrected visual acuity (BCVA) ranged from 20/20 to 20/150. Final BCVA ranged from 20/20 to 20/80. In all eyes except for one, which had a pretreatment vision of 20/20, at least two lines of Snellen visual acuity improvement were achieved. Resolution of submacular fluid, hemorrhage, or exudates was noted in six eyes; in the remaining eye there was persistence of subretinal lipid. There were no complications including optic neuropathy in any of the treated eyes. Follow-up ranged from 6 to 13.5 months following last treatment (mean, 10 months). CONCLUSION: Photodynamic therapy with verteporfin for extrafoveal symptomatic peripapillary CNV appears to be effective in improving vision and promoting the resolution of subfoveal exudates, hemorrhage, or fluid. A randomized controlled study with longer follow-up is justified. PMID- 15655439 TI - PVD following plasmin but not hyaluronidase: implications for combination pharmacologic vitreolysis therapy. AB - PURPOSE: To study whether intravitreal injection of plasmin + hyaluronidase safely induces posterior vitreous detachment (PVD). METHODS: Rabbits were randomized into three groups: (A) 20 rabbits, intravitreal injection of plasmin 1 U + hyaluronidase 20 U in balanced salt solution (BSS) 0.1 mL into one eye; (B) 12 rabbits, plasmin alone; (C) 12 rabbits, hyaluronidase alone. The fellow eye of each rabbit was injected BSS 0.1 mL. In Group A, scanning electron microscopy (SEM) was done in four rabbits at 0.5 hour and in four rabbits at 1 hour. After 7 days, all the remaining 36 rabbits received electroretinography, SEM was examined in eight of each group, and immunohistochemistry was done in four of each group. RESULTS: SEM disclosed the eyes of Group A had complete PVD (8/8), Group B partial PVD (7/8), and Group C (8/8) and all the control eyes (24/24) no PVD after 7 days. Partial PVD was found in 4/4 at 0.5 hour and complete PVD was seen in 3/4 at 1 hour in Group A. Immunohistochemistry showed that the amounts of laminin and fibronectin in the vitreoretinal interface were decreased in Group A and B versus the control eyes (P <0.001), but not in Group C versus the control eyes (P >0.05). Electroretinography showed no changes in any group (P >0.05). CONCLUSION: Vitreous injection of plasmin + hyaluronidase induced complete PVD with no obvious toxicity. Plasmin induced partial PVD, but hyaluronidase had no effects. PMID- 15655440 TI - Predictive visual outcome after macula-off retinal detachment surgery using optical coherence tomography. AB - PURPOSE: To determine whether preoperative optical coherence tomography (OCT) is useful in predicting postoperative visual outcome for patients with primary macula-off rhegmatogenous retinal detachment (RD). METHODS: This prospective study included 20 nonconsecutive eyes with macula-off RD and successful reattachment of the retina. Preoperative 5-mm OCT was performed through the center of the fovea. The relationship among preoperative OCT findings, main preoperative clinical variables, and final postoperative visual acuity was statistically analyzed. RESULTS: Final postoperative visual acuity was negatively correlated with three preoperative OCT variables: height of RD at the central fovea (r=0.79; P <0.001), distance from the central fovea to the nearest undetached retina (r=0.75; P <0.00051), and, to a lesser degree, extent of structural changes in the detached retina (r=0.48; P=0.03). Preoperative visual acuity was the only clinical variable statistically correlated with final postoperative visual acuity in this study (r=0.55; P=0.01). Using multivariate logistic regression analyses, the structure of the detached retina combined with the distance from the central fovea to the nearest undetached retina, as determined with OCT, was highly correlated with final postoperative visual acuity (r=0.82; P <0.000051). CONCLUSION: Preoperative macular analysis with OCT may help to predict visual outcome for patients with macula-off RD. PMID- 15655441 TI - Is removal of internal limiting membrane always necessary during stage 3 idiopathic macular hole surgery? AB - PURPOSE: To determine the indications for internal limiting membrane (ILM) removal in stage 3 idiopathic macular holes (MHs). METHODS: Focal posterior vitreous detachments (PVDs) at MH rims were examined preoperatively by optical coherence tomography and binocular slit-lamp fundus examination in 19 patients retrospectively. All eyes underwent pars plana vitrectomy and creation of a PVD, and some eyes underwent a second surgery to remove the ILM. Indications of ILM removal for MH closure were discussed. RESULTS: Preoperatively, 9 eyes did not (non-PVD group) and 10 eyes did (PVD group) have complete focal PVDs. In all nine eyes in the non-PVD group, MHs were closed after the creation of a PVD without ILM peeling (P <0.05, chi test). In the PVD group, 5 eyes (50%) had MHs closed by making PVD complete without ILM removal, and 5 eyes (50%) required ILM removal in a second surgery. In the end, closure of MHs was achieved in all eyes. CONCLUSION: Anatomic closure of stage 3 idiopathic MHs without a PVD at the rim of the hole may be achieved only by creating a PVD without ILM removal. PMID- 15655442 TI - The effects of comfort care on the pain response in preterm infants undergoing screening for retinopathy of prematurity. AB - OBJECTIVE: The aim of the study was to determine if pain and distress during the retinopathy of prematurity (ROP) screening examination could be ameliorated by providing comfort care. STUDY DESIGN: This study was a prospective, randomized, controlled trial of 30 stable preterm infants who underwent initial ROP screening examinations. Fourteen study infants were swaddled, held, and given 24% sucrose solution during the examination. Sixteen controls were examined while lying in their cribs. Vital signs (i.e., pulse rate, respiratory rate, and oxygen saturation), crying time, and time for the vital signs to return to baseline values were recorded at different times during the examination. RESULTS: The vital signs did not vary significantly between the two groups. The participants in the control group had a trend of longer crying time, but this trend did not reach a level of statistical significance. In addition, The time required for the vital signs to return to their baseline values did not vary significantly. CONCLUSION: ROP screening is very distressful for preterm infants. The routine use of comfort care to reduce pain during the examination could not be supported by this study. PMID- 15655443 TI - Midperipheral mottling pigmentation with familial choroidal osteoma. AB - PURPOSE: To describe a rare presentation of familial choroidal osteoma in two siblings. METHODS: The clinical findings in two siblings over 4 years' follow-up. RESULTS: Two brothers (15 and 12 years old) had bilateral choroidal osteomas. Both had bilateral peripapillary yellowish-white lesions and midperipheral mottling pigment appearance, which are not seen in sporadic cases. Extensive midperipheral area with mottling pigment appearance was noted by fluorescein angiography (FA) as scattered multiple hyperfluorescent dots. The yellowish-white lesions showed diffuse hyperfluorescence with FA and hypofluorescence with indocyanine green angiography (ICG). ICG also revealed irregular hyperfluorescent areas within the tumor, indicating abnormal choroidal vessels on the tumor. In the left eye of the younger brother, the subretinal fibrosis due to choroidal neovascularization superior to the macula extended down toward the foveal region over 2 years, resulting in visual deterioration. CONCLUSION: The midperipheral mottling pigment appearance of familial choroidal osteoma cases is unique and different from most sporadic cases, suggesting that familial choroidal osteoma might have separate etiologic or modified factors. PMID- 15655444 TI - X-linked retinoschisis in three females from the same family: a phenotype genotype correlation. AB - PURPOSE: To describe the clinical findings and outcome for three homozygous females affected with X-linked retinoschisis (XLRS) in a large Colombian family with 26 affected males. METHODS: Retrospective review of charts for females from a family with XLRS who underwent complete ophthalmologic examinations, ancillary tests, clinical genetic evaluation, and molecular studies. RESULTS: Three female patients (6 eyes) with clinical findings of XLRS were identified. The patients' ages ranged from 10 to 37 years. Initial visual acuity was equal to or worse than 20/50 in 4 eyes (66%) of 2 patients. Four eyes (66%) were hyperopic. Intraocular pressure was normal in all eyes. Three eyes (50%) had cataracts, and vitreous veils were present in 3 (50%). The optic disk was pale in 6 eyes (100%). Foveal schisis was present in 6 eyes (100%). Peripheral retinal schisis was present in five eyes. A silvery gloss or tapetal-like retinal reflex or sheen was observed in 6 eyes (100%), and dendritic lines were found in 2 (33%). One eye had a retinal break, and one had a retinal detachment. Two eyes underwent cataract extraction; one patient underwent bilateral cryotherapy, one, laser treatment, and one, scleral buckling. Final visual acuity was 20/50 to 20/100 in 2 eyes and 20/200 to 20/400 in 4. Follow-up ranged from 7 to 22 years. Molecular analysis showed that all three female patients were homozygous for the allele 639delG of (on) the XRLS1 gene. CONCLUSIONS: Compared with their affected male relatives, three females from a family with XLRS had similar ocular findings and a more severe course of disease. These findings are explained by the fact that these patients were homozygous for a mutation in the XLRS1 gene. PMID- 15655446 TI - Spontaneous regression of an isolated retinal astrocytic hamartoma. PMID- 15655445 TI - The effect of indocyanine green on cultured retinal glial cells. AB - BACKGROUND: Recently, indocyanine green (ICG) has been utilized to visualize inner limiting membrane in vitreous surgery. However, the safety of ICG injected into the vitreous has not been well established. The possible toxicity of ICG on Muller cells was investigated using cultured rat retinal glial cells (RGCs). METHODS: Rat RGCs were cultured in Dulbecco modified Eagle medium supplemented with 20% fetal calf serum. The cytotoxicity of ICG was assayed with viable cell number and resazurin metabolic assay. The expression of the apoptosis-related gene bcl-2 was examined with real-time polymerase chain reaction analysis. RESULTS: The effects of ICG on the viability of rat RGCs were tested at two different concentrations (0.05% and 0.5%). ICG significantly decreased the viable cell number of RGCs at 0.5%, while there was no significant effect at 0.05%. Similarly, the metabolic activity to resazurin was significantly decreased by exposure to 0.5% ICG. However, ICG showed little effects on resazurin metabolism at 0.05%. The expression levels of bcl-2 mRNA were higher in cells treated with 0.5% ICG than in those treated with 0.05% ICG and untreated control cells. CONCLUSION: The data suggest that ICG initiates the death of RGCs at high concentrations, in part, through apoptosis-related signal pathways. PMID- 15655447 TI - Transient optic disc edema by vitreous traction in a quiescent eye with proliferative diabetic retinopathy mimicking diabetic papillopathy. PMID- 15655448 TI - Purtscher-like retinopathy associated with acute renal allograft rejection. PMID- 15655449 TI - Hypotensive anesthesia in the management of a posterior suprachoroidal foreign body. PMID- 15655450 TI - Unusual complication after temporary use of perfluorohexyloctane (f6h8) in the treatment of complicated retinal detachment. PMID- 15655451 TI - The use of boric acid solution to help in the removal of biodegraded Miragel episcleral buckles. PMID- 15655452 TI - Mucosa-associated lymphoid tissue lymphoma with intraocular involvement. AB - PURPOSE: To report the clinicopathologic features of a patient with mucosa associated lymphoid tissue (MALT) lymphoma of the conjunctiva and associated intraocular involvement. METHODS: This study is a retrospective clinicopathologic correlative case report summarizing the clinical, radiologic, and histopathologic findings of a patient with conjunctival MALT lymphoma and associated intraocular involvement. RESULTS: Ophthalmic examination and fluorescein angiography demonstrated progressive conjunctival infiltration bilaterally, marked uveal effusions in the left eye, and cellular white infiltrates of the choroid in the right eye. MRI of the orbit revealed a diffusely infiltrating intra- and extraocular lesion extending around the globe and optic nerve in the left eye without evidence of intracranial extension. Conjunctival biopsy showed low-grade tumor cells, consistent with the diagnosis of MALT lymphoma. The patient was successfully treated with external beam radiation with marked clinical improvement. CONCLUSION: Conjunctival MALT lymphomas, typically indolent and localized tumors, may extend into the orbit and invade local tissues such as the choroid with devastating consequences. A conjunctival biopsy can provide an easy diagnosis of this treatable entity. PMID- 15655453 TI - Bilateral staphylomas in a patient with Hermansky-Pudlak syndrome. PMID- 15655454 TI - Intravitreal injection of VEGF siRNA. PMID- 15655457 TI - Breast reduction and smoking. AB - To investigate the influence of smoking on the occurrence of complications in patients undergoing breast reduction, a retrospective case note study was performed. Seventy-one patients who had 118 reduction mammoplasties between April 2001 and March 2002 were analyzed; 15.5% (11/71) of all patients suffered a complication. Ten complications were of infective nature. There was no case of partial or total nipple necrosis. Factors like past medical history, breast history, drug history, age, body weight, and resection weight did not influence the development of complications (P > 0.4, respectively). Smoking, however, was found to be a significant risk (P < 0.05). The risk to develop complications after reduction mammoplasty was calculated to be more than 3 times as high when compared with nonsmokers. PMID- 15655455 TI - Photodynamic therapy. PMID- 15655458 TI - Correction of inverted nipple: an alternative method using continuous elastic outside distraction. AB - Inverted nipple, which is defined as a nipple located on a plane lower than the areola, presents both functional and cosmetic problems. It is a source of repeated irritation and inflammation and interferes with nursing. In addition, its abnormal appearance may cause psychologic distress. With consideration of its underlying pathophysiologic components and severity, a number of techniques have been introduced for correction of this anomaly. Most of these techniques involve extensive skin incision around the nipple that may jeopardize the blood and nerve supply to the nipple or create much scar tissue that is esthetically objectionable. For correcting the inverted nipple, the authors introduce an alternative, simple method using continuous elastic outside distraction. Compared with other methods using outside distraction, the authors used an adjustable elastic instrument made of steel wire, spring, and plastic syringe; continuous distraction of the inverted nipples; and sustaining 3 to 6 months. From August 2002 to December 2003, 14 patients (26 nipples) were treated. 12 patients had bilateral inverted nipples. Patient age at operation ranged from 14 to 40 years (mean age, 24 years). All nipples were congenital, and they had no previous operation. Six nipples were grade I, 9 nipples were grade II, and 11 nipples were grade III according to the classification of inverted nipple by Han and Hong. The mean follow-up period was 7.3 months (range, 3-12 months). Follow-up examinations revealed no evidence of recurrence of inversion. There was no complication associated with surgery, such as infection, hematoma, permanent sensory disturbance, or nipple necrosis. All patients were satisfied with their results. The authors conclude that their procedure is reliable, a simple, safe, and effective method for correction inverted nipple. But a long-term follow-up is needed. This technique can be applied to any type of inverted nipple as a primary surgical procedure. PMID- 15655459 TI - Lower abdominal bulge after deep inferior epigastric perforator flap (DIEP) breast reconstruction. AB - The etiology of lower abdominal bulge following breast reconstruction with the DIEP flap is uncertain. Most studies report an incidence that ranges from 0.7% to 5%. The purpose of this study was to review a set of factors that may predispose to a lower abdominal bulge. This was a retrospective review of 123 women who had breast reconstruction with the DIEP flap over a 4-year period. The reconstruction was unilateral in 93 women and bilateral in 30 women, totaling 153 flaps. Etiologic factors that were evaluated included patient age, diabetes mellitus, tobacco use, previous abdominal operations, unilateral or bilateral reconstruction, previous childbirth, aponeurotic plication to improve the natural abdominal contour, and use of Marlex mesh. A lower abdominal bulge occurred in 5 of the 123 women (4%), 2 following 30 bilateral reconstructions (6.6%) and 3 following 93 unilateral reconstructions (3.2%). Analysis of the factors for all women demonstrated diabetes mellitus in 1 (0.8%), tobacco use in 9 (7.3%), a prior abdominal operation in 55 (44.7%), previous childbirth in 95 (77%), aponeurotic plication in 49 (40%), and use of Marlex mesh in 4 (3.3%). Statistical analysis did not show any significant association between the explanatory factors and the occurrence of a lower abdominal bulge, except for a weak trend in women who had not been pregnant (P = 0.08). The results of this study demonstrate that the occurrence of a lower abdominal bulge following the DIEP flap is a random event that can occur in anyone. Pregnancy may confer a preventative effect as the collagen fibers strengthen to overcome the stretching forces. Techniques for prevention and treatment include intraoperative assessment of the anterior rectus sheath, use of an adjuvant material for reinforcement if unstable, and vertical plication for bulge repair. PMID- 15655460 TI - Periareolar-transareolar-perithelial incision for the surgical treatment of gynecomastia. AB - Gynecomastia is an abnormal enlargement of the breast tissue in men. It is the most common disorder of the male breast. Surgical sharp resection of the excess breast tissue is still the mainstay of treatment when medical treatment modalities are proved to be ineffective. The authors believe that areolar incisions give the best results, especially for grades I and IIA gynecomastia. The authors review the ever-increasing areolar incision techniques that have been previously recommended, propose a classification for these techniques, and introduce an alternative technique for areolar resection of the enlarged gland in gynecomastia. An inferior pole, periareolar-transareolar-perithelial (PTP) incision was designed and 15 patients were operated successfully using this technique. Twelve cases were bilateral and 3 were unilateral (27 breasts). A 65 mm access port can be obtained from a 30-mm-diameter areola. No color changes or slough was observed in any of the patients. Areolar access incisions can be classified into 4 main groups: circumareolar, periareolar, transareolar, and circumthelial, and their subgroups. Like every incision proposed, the PTP incision cannot be recommended for every grade of gynecomastia. It is best suited for grades I, IIA, and IIB gynecomastia. Its wide exposure and potential advantage for areolar reduction makes this incision a good alternative to other areolar approaches. PMID- 15655461 TI - Management strategies for peripheral iatrogenic nerve lesions. AB - For diagnostic or therapeutic reasons, various medical procedures may cause iatrogenic injury of peripheral nerves. The authors treated 82 patients for iatrogenic nerve injury between 1990 and 2000. The main reasons for iatrogenic nerve injury were surgical failure, traction or pressure lesions, hematoma, or inadequate positioning of the patient. The authors performed neurolysis in 67 cases, which included reconstruction by nerve graft in 26 patients and direct coaptation of the nerve in 3 patients. Their postoperative results emphasize the importance of early diagnosis and adequate treatment of iatrogenic peripheral nerve damage for optimal functional recovery. The authors highlight the different causes for iatrogenic injury in this study to alert surgeons as well as other medical specialists to simple strategies for avoiding nerve injury. They also indicate the need for physicians to make themselves familiar with early diagnostic steps for detecting iatrogenic injuries, like electroneurographic studies, and to document physical examinations well, to deal with this problem most effectively. PMID- 15655463 TI - Postoperative morphologic analysis of carpal tunnel syndrome using high resolution ultrasonography. AB - The authors evaluated the morphologic changes that follow division of the transverse carpal ligament in patients with carpal tunnel syndrome (CTS) using high-resolution ultrasonography. Ten patients, for a total of 20 hands, underwent high-resolution ultrasonographic studies before the operation and 8 months after the operation. They were all diagnosed with bilateral idiopathic CTS. The authors evaluated the configuration of the median nerve and carpal tunnel at 3 different levels of the wrist: the distal radiocarpal joint level, the pisiform level, and the hook of hamate level. The median nerve gained in thickness to a remarkable extent at 2 distal levels after the operation. The change in morphology of the carpal tunnel at these 2 distal levels was obvious, but the cross-sectional area of the carpal tunnel was increased significantly only at the hook of hamate level. The transverse diameters of the carpal tunnel were not significantly changed. As mentioned, the authors found that the median nerve gained significantly in volume at the distal part of the carpal tunnel postoperatively, and the volumetric increase in the carpal tunnel appears to have resulted from an anterior displacement of newly formed transverse carpal ligament, rather than from a widening of the bony carpal arch. PMID- 15655464 TI - Treatment of idiopathic postmenopausal osteoarthrosis of the trapeziometacarpal joint with the Epping resection arthroplasty technique. AB - Surgical treatment of idiopathic postmenopausal osteoarthrosis of the trapeziometacarpal joint with the Epping resection arthroplasty was performed in 57 cases in 49 women. Data show good pain relief (between 58% and 76%), very good subjective results with 89% patient satisfaction and Disability of Arm, Shoulder, Hand (DASH) scores in the lower third of the scale after 35 months' follow-up. Good functional results with respect to radial abduction (51 degrees) and palmar flexion (45 degrees), as well as improvement in strength measurements, could be achieved. Some patients (13%) reported remaining problems with occasional pain during performance of activities of daily life and work. A significant proximal metacarpal migration (31%) without correlation to objective or subjective outcome was found. The Epping procedure has proven to be a valuable alternative procedure to treat idiopathic postmenopausal trapeziometacarpal arthrosis after a midterm follow-up period. Still, careful patient selection is important and sufficient preoperative information necessary. PMID- 15655466 TI - The middle phalanx in Poland syndrome. AB - The classic deformity of Poland syndrome consists of the combination of unilateral aplasia of the sternocostal head of the pectoralis major muscle and an ipsilateral hypoplastic hand with simple syndactyly and short fingers. The current study specifically investigates the abnormalities of the middle phalanx in a series of 15 patients with Poland syndrome. The degree of hand hypoplasia and finger brachydactyly was classified into 3 grades: mild (n = 2), moderate (n = 10), and severe (n = 3). The middle phalanges of all fingers in all patients showed abnormalities. The degree of deficiency in the middle phalanx (from mild hypoplasia to aplasia) correlated with the degree of brachydactyly, which in turn correlated with the degree of hand hypoplasia. Several other radiologic features of the hypoplastic middle phalanges were documented, such as the central nidus, the rounded or truncated appearance, and the cone-shaped epiphyses. The embryology of limb development was reviewed and it was proposed that abnormalities of the middle phalanx in Poland syndrome occur as a result of a mild ischemic insult during stage 19 of the embryonic life, leading to an arrest or lack of chondrification of the mesenchymal mass of the phalanx. PMID- 15655465 TI - The truly distal lateral arm flap: rationale and risk factors of a microsurgical workhorse in 30 patients. AB - The forearm part of the extended lateral arm flap may be separately raised on the most distal septocutaneous perforator of the posterior collateral radial artery. This truly distal lateral arm flap shares most of the advantages of the radial forearm flap and is associated with less donor site morbidity. From April 2000 to March 2004, we used 30 such flaps as the fasciocutaneous free flap of choice, mostly for reconstructions in the head and neck region. The eventful postoperative course observed in 5 of these flaps motivated us to evaluate the rationale and risk factors of this procedure. We prospectively analyzed the influence on the incidence of partial or complete flap loss of 19 patient-related or procedure-related characteristics that may have acted as risk factors. None were found to be of statistical significance. We found the distal lateral arm flap to have a less robust vascular anatomy than the radial forearm flap, resulting in the need for advanced surgical expertise to raise and handle it. As we recognized the difficulty of this flap to be associated predominantly with this anatomy of its vascular pedicle, we now take a more liberal stand toward the possibility of intraoperative conversion to the use of a radial forearm flap. PMID- 15655467 TI - Transfusion-free pediatric burn surgery: techniques and strategies. AB - More than 1.3 million children sustain burns each year, resulting in 40,000 admissions and more than 3000 pediatric deaths. Pediatric burn surgery has been described as excessively bloody. Strategies to reduce intraoperative blood loss include the use of topical thrombin and epinephrine, extremity tourniquets, acute normovolemic hemodilution, and hypotensive anesthesia. This study reviews the single surgeon pediatric burn experience at a children's hospital and describes a comprehensive blood conservation protocol to achieve transfusion-free pediatric burn surgery. A retrospective chart review of consecutive pediatric burn surgeries from July 2000 to April 2002 was performed. Patient demographics, burn characteristics, treatment, blood loss, laboratory values, transfusion history, and complications were reviewed. Blood loss per percent total body surface area (TBSA) treated as well as percent total blood volume (TBV) loss divided by percent TBSA treated were calculated. A total 31 burn surgeries in 23 patients were reviewed. The average age was 7 years (range, 9 months-17 years). There were 17 extremity, 6 trunk, and 2 head/neck burns. The average TBSA burned was 15% (range, 1-55%). The protocol to reduce intraoperative blood loss consisted of the debridement of full-thickness burns with electrocautery and partial-thickness burns with dermabrasion. All debrided or harvested surgical sites were treated immediately with epinephrine solution-soaked pads. All graft harvest sites were injected with an epinephrine solution before harvesting split-thickness skin grafts. The average TBSA treated per surgery was 7% (range, 1-29%). The average blood loss per percent TBSA treated was 15 mL (range, 0.7-37 mL). The average percent TBV/percent TBSA was 0.76% (range, 0.04-3.6%). All 20 patients underwent surgical debridement, 7 patients were treated with AlloDerm and ultrathin split thickness skin grafts, 2 with full-thickness skin grafts, and 17 with split thickness skin grafts alone. Five children required blood transfusions. These burns averaged 32% TBSA (range, 20-55%). All 5 children receiving transfusions had anemia of thermal injury and demonstrated an average preoperative drop in hematocrit of 12% (range, 10-14%). There was a 29% complication rate, with 7 patients experiencing partial graft loss, and 1 patient who developed a postgraft contracture that required revisional surgery. There was a single mortality secondary to systemic inflammatory response syndrome and acute respiratory distress syndrome. After the proposed pediatric burn treatment protocol, intraoperative blood loss requiring transfusion can be minimized or eliminated. Large TBSA burns must be surveilled for burn wound anemia that may ultimately require blood transfusion. PMID- 15655468 TI - Comparison of preoperative anxiety in reconstructive and cosmetic surgery patients. AB - Surgery is a serious stressor and a cause of anxiety for the patients. Reconstructive surgery patients are mostly operated on because of certain functional impairment or disability; on the contrary, cosmetic surgery patients do not have any physical impairment and they are operated on because of mostly psychologic reasons. The aim of this study was to compare the anxiety levels in the reconstructive surgery patients and cosmetic surgery patients preoperatively. Thirty-two patients in the reconstructive surgery group and 30 patients in the cosmetic surgery group were included in the study. State Trait Anxiety Inventory was used to measure the anxiety levels in these 2 groups preoperatively. The 2 groups were similar in characteristics such as age, gender distribution, number of previous operations, and trait anxiety scores. Mean state anxiety scores obtained for the reconstructive surgery group was 38.0 +/- 8.7, while it was 44.2 +/- 10.79 for the cosmetic surgery group (t test, degrees of freedom = 60, P = 0.015). This study reveals that preoperative anxiety levels in the cosmetic surgery patients are higher than those of the reconstructive surgery patients. Therefore, adequate preoperative preparation for cosmetic surgery should include attempts to cope with anxiety. Anxiolytics may be used more liberally and professional psychologic assistance may be required. PMID- 15655470 TI - Circumferential negative-pressure dressing (VAC) to bolster skin grafts in the reconstruction of the penile shaft and scrotum. AB - This paper presents 4 consecutive cases using negative-pressure dressings (VAC) to bolster skin grafts in male genital reconstruction. In this series reconstruction followed 1 case of tumor ablation and 3 cases of debridement of abscesses or Fornier's gangrene. The VAC was applied circumferentially to the penis to secure skin grafts either directly to the penile shaft or to facilitate skin grafting to the scrotum. Graft areas ranged from 75 to 250 cm. All cases resulted in successful genital wound coverage; minor complications are described. Three practical points are brought forth. First, the VAC facilitates skin grafting to the complex contour of male genitalia. Second, the VAC can be applied circumferentially to the penis without the need for perfusion monitoring or fears of avascular necrosis. Third, with the use of the VAC, bolster use can likely be discontinued as early as 72 hours with good graft adherence and survival. PMID- 15655471 TI - Flap algorithm in vulvar reconstruction after radical, extensive vulvectomy. AB - The objective of this study was to assess the reconstructive options after radical, extensive vulvectomy; relate them to tumor characteristics; and select a choice of flaps able to correct every remaining defect. This study is a retrospective review of a 4-year experience with 31 flaps in 20 consecutive vulvar reconstructions. Three of the 31 flaps presented nonsignificant delayed healing at their tips and 3 other flaps developed a major breakdown related to an infection or an error in flap planning. According to the authors, the size of the defect is the main issue that must be taken into consideration during the establishment of reconstructive needs. Closure of vulvar defects is preferably performed using fasciocutaneous flaps, which are very reliable flaps and can be raised with different techniques to meet different needs. A flap is then chosen with the fewest potential complications. An algorithm has been thus established: Small to medium-size defects are closed with island V-Y flaps, island gluteal fold flaps, or pedicled pudendal thigh flaps. Among them, the island V-Y flap is the workhorse flap for vulvar reconstruction because of its versatility, reliability, and technical simplicity compared with its very low complication rate. If the vulvar defect is large and/or reaches the vulva-crural fold, V-Y flaps are also preferred to close these large and posteriorly extended excisions. If the vulvar defect is very large, extending both anteriorly and posteriorly, the use of a distally based, vertically oriented rectus abdominis muscle flap is recommended. Using this algorithm, immediate vulvar reconstruction with pedicled local or regional flaps can be performed easily and reliably. PMID- 15655472 TI - Histologic characteristics of the human prepuce pertaining to its clinical behavior as a dual graft. AB - The aim of this study was to investigate the unique histologic structure of the normal human prepuce, paying particular attention to the resemblance and dissimilarities between the inner (ie, mucosa) and outer (ie, skin) layers. Histologic sections were stained using hematoxylin-eosin and Van Gieson stains. Transmission electron microscopy was used to evaluate the ultrastructure. Dense capillary networks can be observed in both the upper and lower dermal zones. The dermis lacks a dense collagenous zone. Melanocytes could not be observed in the mucosa. Elastin fibers and bundles were very abundant and dense. Early edema formation can be explained by the loose character of the dermal structure. Better graft "take" in mucosal grafts may be the result of the dense vascular dermal network. Mild hyperpigmentation can be explained by the limited number of melanocytes. However, this can also be observed in mucosal grafts, despite the absence of melanocytes. This may be solely the result of inflammatory hyperpigmentation, which can be seen in skin grafts. The abundance of elastin fibers in the prepuce may be the reason behind the superior wound contraction inhibition. PMID- 15655473 TI - Spontaneous recanalization of the divided lactiferous duct in the rat. AB - The objectives of the current study were to test the hypotheses that ductus lactiferous may regenerate after its transection and to develop a reproducible model for future studies. A total of 168 nipples from 14 Wistar female rats were used in this experimental study. This study was conducted in 2 parts. During part 1, the normal anatomy of the lactiferous duct and its histomorphologic features were studied. This group was also used for histologic and morphometric comparison. During part 2, the lactiferous duct was exposed through a vertical incision and was transected by scissors. A small gap was produced by elastic retraction of the surrounding tissue. The authors assessed patency mainly by histologic evaluation. Galactography was performed to collect additional support for the recanalization process. The patency rate of the lactiferous ducts was 66.6%. Luminal narrowing was observed in all specimens. A significant difference did occur in the mean cross-sectional area (P < 0.01) and the wall thickness (P < 0.05) between 2 groups. A computed stereologic image analyzer was used to estimate the mean surface area of a lactiferous duct and the wall thickness. PMID- 15655474 TI - Dynamic changes appearing in collagen fibers during intrinsic tendon repair. AB - Intrinsic healing of severed tendons shows a delay in a gain in breaking strength and the tendon becomes translucent. The cause of tendon translucence was investigated in suture-repaired rat Achilles tendon. The repair site with adjacent translucent tendon were evaluated histologically on day 10 by immunofluorescence and transmission electron microscopy. The healing tendon translucent region by hematoxylin-eosin staining had few inflammatory cells, polarized light birefringence showed thinner collagen fibers, immunofluorescence showed few myofibroblasts, and transmission electron microscopy revealed frayed, irregular thin collagen fibers. During embryogenesis, tendon fibers grow by the addition of discreet collagen fibril segment structures. The speculation is that collagen fibril segment structures are released from collagen fibers within the translucent tendon region for reuse during the regeneration of tendon collagen fibers during intrinsic tendon repair. Healing tendon translucence is related to a decrease in the diameter of collagen fibers by the release of collagen fibril segments within tendon bundles/fascicles. PMID- 15655475 TI - Combined intraoral and nasal approach to Tessier No:0 cleft with bifid nose. AB - The surgical correction of craniofacial clefts is an extremely difficult field in pediatric plastic surgery. Facial clefts are rare entities that most plastic surgeons will rarely have to manage. Various surgical techniques devised by some authors for facial clefts have not still widely accepted. The bifid nose deformity is generally an indicator of Tessier No:0 clefts, with various degrees of skeletal problems. The nasal roof area is an important key point in determining the appropriate surgical technique for reconstruction. In this report, a case of Tessier No:0 with a moderate nasal bifidity and ours novel surgical repair technique combining oral and nasal incision are presented. PMID- 15655476 TI - Radiation-induced angiosarcoma after mastectomy and TRAM flap breast reconstruction. AB - Radiation-induced angiosarcoma of the breast is being reported with increasing frequency as a result of the increased use of radiation therapy in conjunction with breast conservation surgery. However, this entity has not been well documented in patients undergoing mastectomy. The authors present a case of angiosarcoma occurring in a patient 6 years after undergoing mastectomy for invasive duct carcinoma with immediate transverse rectus abdominis musculocutaneous flap reconstruction followed by postoperative radiation therapy. The diagnosis of angiosarcoma was made by skin biopsy performed by the patient's reconstructive surgeon on routine follow-up examination. This is the first reported case of postradiation angiosarcoma occurring in a postmastectomy breast reconstructed with autogenous tissue and it is unusual in that the cancer invaded the musculocutaneous flap. Diagnosis and management recommendations for radiation induced angiosarcoma are discussed. PMID- 15655477 TI - The Speedo tie-over dressing. AB - A system is presented for the application of tie-over dressings. Following skin grafting, the graft is secured with a tie-over dressing covered with a sheath of 3-way stretch fabric. The fabric is secured with the aid of staples. This system is easy to apply and is useful for securing grafts in otherwise difficult anatomic locations. PMID- 15655479 TI - In vivo visualization of platelet/endothelium cell interaction in muscle flaps. PMID- 15655480 TI - An unexpected complication of steroid use for the treatment of hypertrophic scar. PMID- 15655483 TI - Are nurses expected to be role models too? PMID- 15655482 TI - Alveolar bone regeneration utilizing b-TCP and platelet-rich plasma (PRP) derived from bone marrow aspirate. PMID- 15655484 TI - Taking steps to ensure personal safety. PMID- 15655485 TI - Women's experiences of faecal incontinence: a study. AB - Community nurses are often involved in the assessment of symptoms of incontinence and the provision of continence aids. However, it is less clear to what extent they can and do provide ongoing support and discuss with their patients the everyday experience of living with incontinence. This article focuses on what it means to live with faecal incontinence and, based on the findings from a small study, discusses the range of experience women with this chronic condition have. PMID- 15655486 TI - Effective team management by district nurses. AB - This article considers the key role played by the district nurse in managing the district nursing team in order to provide high quality health care. It considers how the district nurse can use key managerial roles (interpersonal, informational and decision-making) in order to ensure unity within the team. The importance of shared goals and trust to achieve unity is explored and a strategy for managing conflict is discussed. Finally, the article suggests a set of ground rules which could be used to facilitate effective team working. PMID- 15655487 TI - Nurse-led case management for older people with long-term conditions. AB - People in the UK are successfully living longer but demonstrating an increased incidence of long-term conditions. In response to this, there has been a growth of service models that draw on the principles of case management, particularly for those older people with multiple and complex needs. Nurses working in primary care, and district nurses in particular, have always made a significant contribution to the care of older people with long-term, chronic conditions using case management techniques. However, the recent emphasis on case management has created new and expanded roles for nurse in primary care as the providers and coordinators of care. The aim of this article is to provide an overview of service models that use a case management approach and the current evidence for their effectiveness. We will then describe current examples of nurse-led case management in the UK and discuss some key issues for nurses in primary care and their patients. PMID- 15655488 TI - The district nurse radio star who hasn't let fame go to her head. Interview by Tom Pollard. PMID- 15655490 TI - Pressure ulcer risk assessment and prevention. AB - Pressure ulcers were once viewed as an inevitable consequence of being infirm and bed-ridden. As it has been recognized that this is not the case, pressure ulcers have come to be seen much more as an indicator of the quality of care provided, and are consequently high on the political and health agenda. This article provides an overview of the key aspects of pressure ulcer risk assessment and prevention drawn from a variety of national policy documents. PMID- 15655489 TI - An introduction to the law of death and the body as property. PMID- 15655492 TI - A fragile future for primary care nursing? PMID- 15655491 TI - Diabetic retinopathy: protecting the vision of people with diabetes. AB - The aim of this article is to provide an update on the effects, causes and treatment of diabetic retinopathy and to discuss practical ways in which community nurses can help protect people with diabetes from losing their vision and support them if they do. PMID- 15655493 TI - Caring for patients with leg ulcers and an underlying vasculitic condition. AB - It is not uncommon for nurses in the community to encounter patients with leg ulceration combined with rheumatic disease, particularly rheumatoid arthritis (RA). The aetiology of the leg ulcers in these cases is rarely straightforward, and the management of the ulcers is correspondingly complex. Management may be further complicated in the presence of vasculitis, an uncommon disorder in which inflammatory changes cause degradation of blood vessels. Rapid deterioration and pain are the main challenges with these cases. This article discusses the aetiology of vaculitic ulcers, and presents two case studies which were successfully managed using a new hydrogel dressing. PMID- 15655494 TI - Uncommon causes of leg ulceration and lesions not to be missed. AB - While the vast majority of cases of leg ulceration are the result of venous and/or arterial disease, and occur in older people, there are several other, less common conditions that can lead to leg ulceration, often in younger people. This article examines the diagnosis and management of some of these conditions, almost all of which will require referral for specialist assessment and treatment. PMID- 15655495 TI - Pressure area care in primary care: issues in equipment provision. AB - The pressure area care needs of primary care patients are becoming more complex due to changes in health care provision. Enabling patients to remain in their own home and allowing them some independence even though they are at high risk of developing pressure damage, relies on the nurse's knowledge of prevention strategies and equipment provision locally. This article discusses the key issues in relation to the provision of pressure-relieving equipment in a rural care trust in England, and how the trust has overcome the problem of equipment failure, especially out of hours and weekends. PMID- 15655496 TI - Accountability is an issue for all nurses. PMID- 15655497 TI - Fear of the known? District nurses' practice of compression bandaging. AB - District nurses are the acknowledged experts when it comes to the care of leg ulcers in the community setting. An audit in one trust nevertheless highlighted a degree of diagnostic caution among district nurses that led to some patients not receiving the compression therapy that national guidelines recommend. This article reports on a small-scale qualitative study that sought to explore this issue in more depth. The results highlight that there is some conflict between district nurses' practice as expert practitioners and strict adherence to guidelines, which the nurses resolve in various ways. The implications for practice are explored. PMID- 15655498 TI - Nitric oxide relaxes circular smooth muscle of rat distal colon through RhoA/Rho kinase independent Ca2+ desensitisation. AB - 1. The aim of this study in circular smooth muscle of rat distal colon was to determine whether Ca(2+) desensitisation, in addition to mechanisms lowering cytosolic free Ca(2+) concentration ([Ca(2+)](cyt)), was involved in the relaxation elicited by nitric oxide (NO). Changes in isometric tension and [Ca(2+)](cyt) were recorded simultaneously in fura-2-loaded strips. 2. In methacholine (10(-5) M)-precontracted preparations, exogenous NO (10(-4) M), adenosine 5'-triphosphate (ATP; 10(-3) M) and electrical field stimulation (EFS; 1 ms, 40 V, 4 Hz, 1 min) induced a decrease in smooth muscle tension, which was accompanied by a fall in [Ca(2+)](cyt). 3. The sarcoplasmic/endoplasmic reticulum Ca(2+) ATP-ase (SERCA) inhibitor thapsigargin (10(-6) M) did not exert an influence on the decrease in tension produced by exogenous NO, but significantly attenuated the fall in [Ca(2+)](cyt). Both the relaxation and the fall in [Ca(2+)](cyt) to ATP and EFS were unaffected by thapsigargin. 4. Calyculin-A (10( 6) M), a myosin light chain phosphatase (MLCP) inhibitor, significantly reduced the decrease in tension elicited by exogenous NO, but did not alter the fall in [Ca(2+)](cyt) to exogenous NO. Inactivating RhoA by exoenzyme C3 (2 mug ml(-1)) or inhibiting Rho-kinase with (+)-(R)-trans-4-(1-aminoethyl)-N-(4-pyridyl) cyclohexanecarboxamide dihydrochloride monohydrate (Y-27632; 10(-5) M) had no effect on the decrease of both tension and [Ca(2+)](cyt) generated by exogenous NO. 5. This paper demonstrates that a RhoA/Rho-kinase independent Ca(2+) desensitisation pathway contributes to the relaxation by NO in circular smooth muscle strips of rat distal colon. PMID- 15655499 TI - Mediators of coronary reactive hyperaemia in isolated mouse heart. AB - 1. Mechanisms regulating coronary tone under basal conditions and during reactive hyperaemia following transient ischaemia were assessed in isolated mouse hearts. 2. Blockade of NO-synthase (50 muM L-NAME), K(ATP) channels (5 muM glibenclamide), A(2A) adenosine receptors (A(2A)ARs; 100 nM SCH58261), prostanoid synthesis (100 muM indomethacin), and EDHF (100 nM apamin+100 nM charybdotoxin) all reduced basal flow approximately 40%. Effects of L-NAME, glibenclamide, and apamin+charybdotoxin were additive, whereas coadministration of SCH58261 and indomethacin with these inhibitors failed to further limit flow. 3. Substantial hyperaemia was observed after 5-40 s occlusions, with flow increasing to a peak of 48+/-1 ml min(-1) g(-1). Glibenclamide most effectively inhibited peak flows (up to 50%) while L-NAME was ineffective. 4. With longer occlusions (20-40 s), glibenclamide alone was increasingly ineffective, reducing peak flows by approximately 15% after 20 s occlusion, and not altering peak flow after 40 s occlusion. However, cotreatment with L-NAME+glibenclamide inhibited peak hyperaemia by 70 and 25% following 20 and 40 s occlusions, respectively. 5. In contrast to peak flow changes, sustained dilation and flow repayment over 60 s was almost entirely K(ATP) channel and NO dependent (each contributing equally) with all occlusion durations. 6. Antagonism of A(2A)ARs with SCH58261 reduced hyperaemia 20-30% whereas inhibition of prostanoid synthesis was ineffective. Effects of A(2A)AR antagonism were absent in hearts treated with L-NAME and glibenclamide, supporting NO and K(ATP)-channel-dependent effects of A(2A)ARs. 7. EDHF inhibition alone exerted minor effects on hyperaemia and only with longer occlusions. However, residual hyperaemia after 40 s occlusion in hearts treated with L-NAME+glibenclamide+SCH58261+indomethacin was abrogated by cotreatment with apamin+charybdotoxin. 8. Data support a primary role for K(ATP) channels and NO in mediating sustained dilation after coronary occlusion. While K(ATP) channels (and not NO) are also important in mediating initial peak flow adjustments after brief 5-10 s occlusions, their contribution declines with longer 20-40 s occlusions. Intrinsic activation of A(2A)ARs is important in triggering K(ATP) channel/NO-dependent hyperaemia. Synergistic effects of combined inhibitors implicate interplay between mediators, with compensatory changes occurring in K(ATP) channel, NO, and/or EDHF responses when one is individually blocked. PMID- 15655500 TI - Aldosterone does not mediate angiotensin II-induced atherosclerosis and abdominal aortic aneurysms. AB - We have demonstrated previously that infusion of angiotensin II (AngII) into hyperlipidemic mice augments atherosclerosis and results in the formation of abdominal aortic aneurysms (AAA). The purpose of this study was to determine the role of aldosterone in these AngII-induced vascular pathologies. Male apolipoprotein E-/- (apoE) mice were infused with either vehicle or aldosterone (50 or 200 ng kg(-1) min(-1)). Arterial blood pressure was determined throughout the study and serum lipid concentrations and vascular pathology were quantified after 28 days of infusion. Infusion of aldosterone did not influence body weight or serum cholesterol concentrations. Kidney weight was increased dose-dependently by aldosterone infusion. Systolic blood pressure was not significantly altered by aldosterone. Plasma aldosterone concentrations were increased dose-dependently by infusion of aldosterone. However, there was no effect of aldosterone on the extent of atherosclerosis and AAAs were not formed. Implantation of pellets containing spironolactone (16 mg kg(-1) day(-1)) in AngII-infused apoE-/- mice (1000 ng kg(-1) min(-1)) had no effect on AngII-induced elevations in blood pressure. Plasma aldosterone concentration was not influenced by coadministration of spironolactone with AngII. Spironolactone administration did not influence the extent of atherosclerosis. Moreover, spironolactone had no significant effect on AngII-induced AAA (incidence of AAA formation: 80 versus 70% for vehicle versus spironolactone, respectively; not significant). These studies demonstrate that the AngII-induced vascular pathologies of atherosclerosis and AAA formation are not mediated through aldosterone. PMID- 15655501 TI - Differential action of a protein tyrosine kinase inhibitor, genistein, on the positive inotropic effect of endothelin-1 and norepinephrine in canine ventricular myocardium. AB - Experiments were carried out in isolated canine ventricular trabeculae and acetoxymethylester of indo-1-loaded single myocytes to elucidate the role of protein tyrosine kinase (PTK) in the inotropic effect of endothelin-1 (ET-1) induced by crosstalk with norepinephrine (NE). The PTK inhibitor genistein was used as a pharmacological tool. Genistein but not daidzein inhibited the positive inotropic effect and the increase in Ca(2+) transients induced by ET-1 by crosstalk with NE at low concentrations. Genistein and daidzein antagonized the negative inotropic effect and the decrease in Ca(2+) transients induced by ET-1 by crosstalk with NE at high concentrations, but genistein did not affect the antiadrenergic effect of carbachol. Genistein but not daidzein enhanced the positive inotropic effect and the increase in Ca(2+) transients induced by NE via beta-adrenoceptors, while the enhancing effect of genistein was abolished by the protein tyrosine phosphatase inhibitor vanadate. These findings indicate that genistein (1) induces a positive inotropic effect in association with an increase in Ca(2+) transients, (2) inhibits the positive inotropic effect of ET-1 induced by crosstalk with NE, and (3) enhances the positive inotropic effect of NE induced via beta-adrenoceptors by inhibition of PTK. In addition, genistein inhibits the negative inotropic effect of ET-1 induced by crosstalk with NE through a PTK-unrelated mechanism. PTK may play a crucial role in the receptor mediated regulation of cardiac contractile function in canine ventricular myocardium. PMID- 15655502 TI - Positive interaction of the beta2-agonist CHF 4226.01 with budesonide in the control of bronchoconstriction induced by acetaldehyde in the guinea-pigs. AB - Pretreatment of anaesthetized guinea-pigs with either CHF 4226.01 (8-hydroxy-5 [(1R)-1-hydroxy-2-[N-[(1R)-2-(p-methoxyphenyl)-1-methylethyl]amino]ethyl] carbostyril hydrochloride), formoterol or budesonide reduced acetaldehyde (AcCHO) evoked responses in the lungs with a rank order of potency CHF 4226.01 (ED(50) values, from 1.88 to 3.31 pmol) > formoterol (ED(50) values, from 3.03 to 5.51 pmol) >> budesonide (ED(50) values, from 335 to 458 nmol). The duration of action of CHF 4226.01 in antagonizing the airway obstruction elicited by AcCHO was also substantially longer than formoterol (area under the curve) at 10 pmol, 763+/-58 and 480+/-34, respectively; P<0.01). Continuous infusion of a subthreshold dose of AcCHO enhanced the intratracheal pressure (ITP) increases caused by subsequent challenges with substance P (from 9.7+/-0.8 to 27.5+/-1.6 cm H(2)O as a peak, P<0.001). Pretreatment with either CHF 4226.01 or formoterol prevented the sensitizing effect of AcCHO on substance P responses (ED(50) values, 2.85 and 6.11 pmol, respectively; P<0.01). The ED(50) value of budesonide (396 nmol) in preventing AcCHO-evoked ITP increase was reduced when this glucocorticoid was combined with 0.1 pmol CHF 4226.01 (ED(50) 76 nmol; P<0.001). CHF 4226.01/budesonide was two-fold more effective (P<0.01) than the formoterol/budesonide combination. These results suggest that CHF 4226.01/budesonide, by optimizing each other's beneficial potential in the control of pulmonary changes caused by AcCHO in the guinea-pigs, may represent a new fixed combination in asthma. PMID- 15655503 TI - Somatostatin-receptor 2 (sst2)-mediated effects of endogenous somatostatin on exocrine and endocrine secretion of the rat stomach. AB - Somatostatin is a potent inhibitor of gastric acid secretion. Its effects are mediated through five specific receptor subtypes (sst(1-5)), of which sst(2) is dominant on the enterochromaffin-like (ECL) cell and the parietal cell. To study the paracrine mechanisms of somatostatin, the sst(2)-specific antagonist PRL-2903 was used. Effects of PRL-2903 on acid secretion and release of histamine were studied in the totally isolated, vascularly perfused rat stomach. Further, the release of histamine and gastrin after bombesin, alone and in combination with PRL-2903, were studied. Results are presented as mean+/-standard error of the mean (s.e.m.). PRL-2903 concentration-dependently increased the venous histamine concentration from basal 55.6+/-7.5 to 367+/-114 nM at 50 microM PRL-2903. With 10 microM PRL-2903, venous histamine output increased from baseline 6.2+/-0.5 to 20.9+/-4.9 nmol h(-1); P=0.008. The combination of 520 pM gastrin and 10 microM PRL-2903 increased venous histamine output from 41.7+/-7.3 nmol h(-1) with gastrin alone to 95.2+/-9.8 nmol h(-1); P=0.016. Further, 10 microM PRL-2903 increased acid output from baseline 8.5+/-1.8 to 37.4+/-11 micromol h(-1); P=0.017. When combined with 10 microM ranitidine, PRL-2903 did not significantly stimulate acid secretion. Bombesin/PRL-2903 increased venous histamine concentration from 50.4+/-14.8 to 292+/-64.2 nM; P=0.008, and gastrin concentration from 38.6+/-13.1 to 95.8+/-20.3 pM; P=0.037. Endogenous somatostatin exerts a continuous restraint on histamine and gastrin release from the gastric mucosa and significantly reduces baseline acid secretion. PMID- 15655504 TI - The expanding field of cannabimimetic and related lipid mediators. AB - The discovery of the endogenous cannabimimetic lipid mediators, anandamide and 2 arachidonoyl glycerol, opened the door to the discovery of other endogenous lipid mediators similar in structure and function. The majority of these compounds do not bind appreciably to known cannabinoid receptors; yet some of them produce cannabimimetic effects while others exert actions through novel mechanisms that remain to be elucidated. This review explores the growing diversity of recently discovered putative lipid mediators and their relationship to the endogenous cannabinoid system. The possibility that there remain many unidentified signalling lipids coupled with the evidence that many of these yield bioactive metabolites due to actions of known enzymes (e.g. cyclooxygenases, lipoxygenases, cytochrome P450s) suggests the existence of a large and complex family of lipid mediators about which only little is known at this time. The elucidation of the biochemistry and pharmacology of these compounds may provide therapeutic targets for a variety of conditions including sleep dysfunction, eating disorders, cardiovascular disease, as well as inflammation and pain. PMID- 15655505 TI - Vascular effects of ovariectomy and chronic oestrogen treatment in rats: controversy or experimental protocol diversity? AB - Several clinical studies have indicated that oestrogens have protective properties on the cardiovascular system. Although the beneficial effect has been attributable, at least in part, to their ability to stimulate the endothelial formation of nitric oxide (NO) and endothelium-derived hyperpolarizing factor (EDHF), the underlying mechanism still remains unclear. In a study from this issue of British Journal of Pharmacology, Nawate et al. have examined the effects of rat ovariectomy and chronic treatment with 17beta-oestradiol on the endothelial function as assessed ex vivo. The data indicate that acetylcholine induced endothelium-dependent relaxations of the isolated mesenteric artery are affected by neither ovariectomy nor chronic hormonal treatment. Despite the maintained endothelium-dependent relaxation, the contribution of the two major endothelial factors NO and EDHF was changed. Indeed, ovariectomy increased the NO mediated component of the relaxation, most likely as a consequence of the downregulation of the physiological allosteric inhibitor of endothelial NO synthase, caveolin-1. In addition, ovariectomy decreased the EDHF-mediated component of the relaxation and membrane hyperpolarization of the smooth muscle cells, an effect which might be explained by a concomitant decrease of the expression of the gap junction connexin-40 and connexin-43. Furthermore, chronic administration of 17beta-estradiol to ovariectomized rats normalized all these effects. This study provides further experimental evidence indicating that the hormonal status plays a determinant role in the control of the endothelial formation of both NO and EDHF. PMID- 15655507 TI - An investigation of whether agonist-selective receptor conformations occur with respect to M2 and M4 muscarinic acetylcholine receptor signalling via Gi/o and Gs proteins. AB - 1. A range of muscarinic acetylcholine (mACh) receptor agonists (methacholine (MCh), oxotremorine-M (OXO-M), oxotremorine (OXO), arecoline (AREC), bethanechol (BETH), pilocarpine (PILO)) have been investigated with respect to their binding to, and activation of, M(2) and M(4) mACh receptors, recombinantly expressed in Chinese hamster ovary cells, to explore the possibility that these agonists may differentially affect mACh receptor-G(i/o) and -G(s) coupling. 2. M(2)/M(4) mACh receptor coupling to the adenylyl cyclase/cyclic AMP signalling pathway has been explored in intact cells. G(i/o)-mediated negative coupling to adenylyl cyclase was explored functionally by assessing the ability of the mACh receptor agonists to inhibit forskolin-stimulated enzymic activity. Following pertussis toxin treatment (100 ng ml(-1), 18-20 h) to inactivate G(i/o) proteins, each agonist caused a G(s)-mediated enhancement of forskolin-stimulated adenylyl cyclase activity. 3. At both M(2) and M(4) mACh receptors, all agonists tested were more potent in mediating G(i/o)- versus G(s)-coupled responses. This difference (determined as the pIC(50) (G(i/o) coupling) minus pEC(50) (G(s) coupling) value) was greatest for AREC (65-75-fold) and least for BETH and PILO (2 cases) of TdP, (B) drugs causing QT prolongation and/or TdP only, the latter at a very low frequency (< or =2 cases), (C) drugs without reports of TdP or QT prolongation. The use of such an algorithm combined with a database accrued from reference compounds with available clinical data is suggested as a basis for testing new candidate drugs in the early stages of development for proarrhythmic risk prediction. PMID- 15655518 TI - Malonate induces cell death via mitochondrial potential collapse and delayed swelling through an ROS-dependent pathway. AB - 1. Herein we study the effects of the mitochondrial complex II inhibitor malonate on its primary target, the mitochondrion. 2. Malonate induces mitochondrial potential collapse, mitochondrial swelling, cytochrome c (Cyt c) release and depletes glutathione (GSH) and nicotinamide adenine dinucleotide coenzyme (NAD(P)H) stores in brain-isolated mitochondria. 3. Although, mitochondrial potential collapse was almost immediate after malonate addition, mitochondrial swelling was not evident before 15 min of drug presence. This latter effect was blocked by cyclosporin A (CSA), Ruthenium Red (RR), magnesium, catalase, GSH and vitamin E. 4. Malonate added to SH-SY5Y cell cultures produced a marked loss of cell viability together with the release of Cyt c and depletion of GSH and NAD(P)H concentrations. All these effects were not apparent in SH-SY5Y cells overexpressing Bcl-xL. 5. When GSH concentrations were lowered with buthionine sulphoximine, cytoprotection afforded by Bcl-xL overexpression was not evident anymore. 6. Taken together, all these data suggest that malonate causes a rapid mitochondrial potential collapse and reactive oxygen species production that overwhelms mitochondrial antioxidant capacity and leads to mitochondrial swelling. Further permeability transition pore opening and the subsequent release of proapoptotic factors such as Cyt c could therefore be, at least in part, responsible for malonate-induced toxicity. PMID- 15655520 TI - Alpha-2 adrenoceptor subtypes: are more better? AB - The discovery of an additional duplicated alpha-2 adrenoceptor subtype in the zebrafish raises a pesky nomenclature issue, as well as questions about the functions of the alpha-2 adrenoceptors in the zebrafish and how many alpha-2 receptors does an organism really need. PMID- 15655519 TI - The pore-forming subunit of the K(ATP) channel is an important molecular target for LPS-induced vascular hyporeactivity in vitro. AB - ATP-sensitive K(+) (K(ATP)) channel activation is implicated in the vascular hyporeactivity occurring in septic shock. However, channel inhibition with the sulphonylurea receptor (SUR) antagonist, glibenclamide (Glib) fails to reverse lipopolysaccharide (LPS)-induced vascular hyporeactivity in vitro. We investigated whether inhibitors that act by binding to the K(ATP) channel pore could be effective. Ring segments of endothelium-intact rat mesenteric artery were incubated with LPS in culture media for either 6 or 20 h before contractile responses to phenylephrine were assessed in the absence or presence of K(ATP) channel inhibitors. The pore-forming subunit inhibitors barium chloride (BaCl(2); 300 microM) and PNU-37883A (1 microM) significantly reversed hyporeactivity at both time points, although less so at 20 h. In contrast, the SUR inhibitors, Glib (10 microM), tolbutamide (Tolb) (1 mM) and PNU-99963 (1 microM) were ineffective. In LPS-incubated tissues, Glib and Tolb antagonised contractions to the thromboxane A2 mimetic, U46619 (9,11-dideoxy-9alpha, 11alpha-methanoepoxy prostaglandin F(2alpha)) (10(-7) M), whereas the pinacidil-derived inhibitor, PNU 99963, did not. Contractions to 60 mM KCl were unaffected by LPS at 6 h, but were significantly depressed by LPS at 20 h, suggesting that K(+)-channel-independent pathways contribute to hyporeactivity at the later time point. The inducible nitric oxide synthase (iNOS) inhibitor, 1400 W (10 microM) and Tolb inhibited the production of nitrite induced by LPS, whereas BaCl(2) and PNU-37883A had no effect. In conclusion, K(ATP) channels contribute to LPS-induced vascular hyporeactivity via the iNOS pathway in rat mesenteric artery. The effectiveness of pore inhibitors over SUR inhibitors of the K(ATP) channel suggests altered SUR function following LPS administration, which cannot be explained by thromboxane receptor inhibition. PMID- 15655521 TI - 2-Furoyl-LIGRL-NH2, a potent agonist for proteinase-activated receptor-2, as a gastric mucosal cytoprotective agent in mice. AB - 1. Proteinase-activated receptor-2 (PAR(2)), expressed in capsaicin-sensitive sensory neurons, plays a protective role in gastric mucosa. The present study evaluated gastric mucosal cytoprotective effect of 2-furoyl-LIGRL-NH(2), a novel highly potent PAR(2) agonist, in ddY mice and in wild-type and PAR(2)-knockout mice of C57BL/6 background. 2. Gastric mucosal injury was created by oral administration of HCl/ethanol solution in the mice. The native PAR(2)-activating peptide SLIGRL-NH(2), administered intraperitoneally (i.p.) at 0.3-1 micromol kg( 1) in combination with amastatin, an aminopeptidase inhibitor, but not alone, revealed gastric mucosal protection in ddY mice, which was abolished by ablation of capsaicin-sensitive sensory neurons. 3. I.p. administration of 2-furoyl-LIGRL NH(2) at 0.1 micromol kg(-1), without combined treatment with amastatin, exhibited gastric mucosal cytoprotective activity in ddY mice, the potency being much greater than SLIGRL-NH(2) in combination with amastatin. This effect was also inhibited by capsaicin pretreatment. 4. Oral administration of 2-furoyl LIGRL-NH(2) at 0.003-0.03 micromol kg(-1) also protected against gastric mucosal lesion in a capsaicin-reversible manner in ddY mice. 5. I.p. 2-furoyl-LIGRL-NH(2) at 0.1-0.3 micromol kg(-1) caused prompt salivation in anesthetized mice, whereas its oral administration at 0.003-1 micromol kg(-1) was incapable of eliciting salivation. 6. In wild-type, but not PAR(2)-knockout, mice of C57BL/6 background, i.p. administration of 2-furoyl-LIGRL-NH(2) caused gastric mucosal protection. 7. Thus, 2-furoyl-LIGRL-NH(2) is considered a potent and orally available gastric mucosal protective agent. Our data also substantiate a role for PAR(2) in gastric mucosal protection and the selective nature of 2-furoyl-LIGRL-NH(2). PMID- 15655522 TI - Conserved structural, pharmacological and functional properties among the three human and five zebrafish alpha 2-adrenoceptors. AB - 1. Zebrafish has five distinct alpha(2)-adrenoceptors. Two of these, alpha(2Da) and alpha(2Db), represent a duplicated, fourth alpha(2)-adrenoceptor subtype, while the others are orthologue of the human alpha(2A)-, alpha(2B)- and alpha(2C) adrenoceptors. Here, we have compared the pharmacological properties of these receptors to infer structural determinants of ligand interactions. 2. The zebrafish alpha(2)-adrenoceptors were expressed in Chinese hamster ovary cells and tested in competitive ligand binding assays and in a functional assay (agonist-stimulated [(35)S]GTPgammaS binding). The affinity results were used to cluster the receptors and, separately, the ligands using both principal component analysis and binary trees. 3. The overall ligand binding characteristics, the order of potency and efficacy of the tested agonists and the G-protein coupling of the zebrafish and human alpha(2)-adrenoceptors, separated by approximately 350 million years of evolution, were found to be highly conserved. The binding affinities of the 20 tested ligands towards the zebrafish alpha(2)-adrenoceptors are generally comparable to those of their human counterparts, with a few compounds showing up to 40-fold affinity differences. 4. The alpha(2A) orthologues and the zebrafish alpha(2D) duplicates clustered as close pairs, but the relationships between the orthologues of alpha(2B) and alpha(2C) were not clearly defined. Applied to the ligands, our clustering methods segregated the ligands based on their chemical structures and functional properties. As the ligand binding pockets formed by the transmembrane helices show only minor differences among the alpha(2)-adrenoceptors, we suggest that the second extracellular loop--where significant sequence variability is located --might contribute significantly to the observed affinity differences. PMID- 15655523 TI - Anxiogenic properties of an inverse agonist selective for alpha3 subunit containing GABA A receptors. AB - Alpha3IA (6-(4-pyridyl)-5-(4-methoxyphenyl)-3-carbomethoxy-1-methyl-1H-pyridin-2 one) is a pyridone with higher binding and functional affinity and greater inverse agonist efficacy for GABA(A) receptors containing an alpha3 rather than an alpha1, alpha2 or alpha5 subunit. If doses are selected that minimise the occupancy at these latter subtypes, then the in vivo effects of alpha3IA are most probably mediated by the alpha3 subtype. Alpha3IA has good CNS penetration in rats and mice as measured using a [(3)H]Ro 15-1788 in vivo binding assay. At doses in rats that produce relatively low levels of occupancy (12%) in the cerebellum (i.e. alpha1-containing receptors), alpha3IA (30 mg kg(-1) i.p.), like the nonselective partial inverse agonist N-methyl-beta-carboline-3-carboxamide (FG 7142), not only caused behavioural disruption in an operant, chain-pulling assay but was also anxiogenic in the elevated plus maze, an anxiogenic-like effect that could be blocked with the benzodiazepine antagonist Ro 15-1788 (flumazenil). Neurochemically, alpha3IA (30 mg kg(-1) i.p.) as well as FG 7142 (15 mg kg(-1) i.p.) increased the concentration of the dopamine metabolite 3,4 dihydroxyphenylacetic acid in rat medial prefrontal cortex by 74 and 68%, respectively, relative to vehicle-treated animals, a response that mimicked that seen following immobilisation stress. Taken together, these data demonstrate that an inverse agonist selective for GABA(A) receptors containing an alpha3 subunit is anxiogenic, and suggest that since alpha3-containing GABA(A) receptors play a role in anxiety, then agonists selective for this subtype should be anxiolytic. PMID- 15655524 TI - Gastrin promotes human colon cancer cell growth via CCK-2 receptor-mediated cyclooxygenase-2 induction and prostaglandin E2 production. AB - The present study investigates the effects of gastrin-17 on human colon cancer HT 29 cells to examine whether gastrin receptor (CCK-2), cyclooxygenase (COX-1, COX 2) isoforms and prostaglandin receptor pathways interact to control cell growth. Reverse transcription (RT)-polymerase chain reaction (PCR) analysis demonstrated that HT-29 cells are endowed with the naive expression of CCK-2 receptor (short splice variant), COX-1, COX-2 and prostaglandin EP(4) receptor, but not gastrin. Gastrin-17 significantly promoted cell growth and DNA synthesis. Both these stimulating effects were abolished by L-365,260 or GV150013 (CCK-2 receptor antagonists), but were unaffected by SC-560 (COX-1 inhibitor). L-745,337 (COX-2 inhibitor) or AH-23848B (EP(4) receptor antagonist) partly reversed gastrin-17 induced cell growth, while they fully antagonized the enhancing action on DNA synthesis. HT-29 cells responded to gastrin-17 with a significant increase in prostaglandin E(2) release. This enhancing effect was completely counteracted by L-365,260, GV150013 or L-745,337, while it was insensitive to cell incubation with SC-560. Exposure of HT-29 cells to gastrin-17 was followed by an increased phosphorylation of both extracellular regulated kinases (ERK-1/ERK-2) and Akt. Moreover, gastrin-17 enhanced the transcriptional activity of COX-2 gene promoter and stimulated COX-2 expression. These latter effects were antagonized by L 365,260 or GV150013, and could be blocked also by PD98059 (inhibitor of ERK-1/ERK 2 phosphorylation) or wortmannin (inhibitor of phosphatidylinositol 3-kinase). Analogously, gastrin-17-induced prostaglandin E(2) release was prevented by PD98059 or wortmannin. The present results suggest that (a) in human colon cancer cells endowed with CCK-2 receptors, gastrin-17 is able to enhance the transcriptional activity of COX-2 gene through the activation of ERK-1/ERK-2- and phosphatidylinositol 3-kinase/Akt-dependent pathways; (b) these stimulant actions lead to downstream increments of COX-2 expression, followed by prostaglandin E(2) production and EP(4) receptor activation; (c) the recruitment of COX 2/prostaglandin pathways contributes to the growth-promoting actions exerted by gastrin-17. PMID- 15655525 TI - The composition of the GABA receptor at the Caenorhabditis elegans neuromuscular junction. AB - 1. The unc-49 gene of the nematode Caenorhabditis elegans encodes three gamma aminobutyric acid type A (GABA(A)) receptor subunits. Two of these, UNC-49B and UNC-49C, are expressed at high abundance and co-localize at the neuromuscular junction. 2. The UNC-49B subunit is sufficient to form a GABA(A) receptor in vitro and in vivo. Furthermore, all loss-of-function unc-49 alleles lack functional UNC-49B. No mutations specifically inactivate UNC-49C. Thus, UNC-49C appears to be dispensable for receptor function; however, UNC-49C has been conserved among different nematode species, suggesting it plays a necessary role. 3. To ascertain whether UNC-49C is part of the GABA(A) receptor in vivo, we performed patch-clamp electrophysiology on C. elegans muscle cells. Sensitivity to GABA, and to the antagonists picrotoxin and pregnenolone sulfate, matched the UNC-49B/C heteromer rather than the UNC-49B homomer, for both exogenous and synaptically-released GABA. 4. The synaptic localization of UNC-49C requires the presence of UNC-49B, indicative of a physical association between the two subunits in vivo. Thus, the in vivo receptor is an UNC-49B/C heteromer. 5. UNC 49C plays a negative modulatory role. Using the rapid ligand-exchange technique in vitro, we determined that UNC-49C causes accelerated receptor desensitization. Previously, UNC-49C was shown to reduce single-channel conductance in UNC-49B/C heteromers. Thus, the function of UNC-49B is to provide GABA responsiveness and localization to synapses, while the function of UNC-49C is to negatively modulate receptor function and precisely shape inhibitory postsynaptic currents. PMID- 15655526 TI - Endothelin-1 exerts a preconditioning-like cardioprotective effect against ischaemia/reperfusion injury via the ET(A) receptor and the mitochondrial K(ATP) channel in the rat in vivo. AB - In vitro studies have demonstrated that endothelin-1 (ET-1) given before myocardial ischaemia may evoke a preconditioning (PC)-like cardioprotective effect. The first aim of this study was to investigate whether administration of ET-1 before ischaemia exerts cardioprotection against ischaemia/reperfusion injury in vivo and to determine involvement of the ET-1 receptor subtype. The second aim was to examine the role of mitochondrial ATP-sensitive K+ channels (mitoK(ATP)) as a mediator of this cardioprotection. Anaesthetised open-chest Wistar rats were subjected to 30 min of coronary artery occlusion followed by 2 h reperfusion (I/R). In protocol I, the first group was subjected to I/R only (control, n=10). In the second (n=10) group, PC was elicited by three 5 min cycles of coronary artery occlusion, separated by 5 min reperfusion before I/R. The third (n=6) and fourth (n=7) groups were given ET-1 intravenous (i.v.) during three 5 min infusion periods separated by 5 min before I/R. The fourth group was in addition given the ET(A) receptor antagonist LU 135252 5 min before the infusions of ET-1. In protocol II, the first group was I/R control as in protocol I (n=8). The second (n=6), third (n=7) and fourth (n=7) groups were given ET-1 as in protocol I. The third group was in addition given the nonselective K(ATP) channel antagonist glibenclamide (Glib) 30 min before the ET-1 infusions and the fourth group the selective mitoK(ATP) channel antagonist 5-hydroxydecanoic acid (5-HD) 5 min before I/R. There were no significant differences in MAP or heart rate between the groups during I/R. In protocol I, PC reduced IS compared to the control group (10+/-3 vs 35+/-5%, P<0.01). Infusion of ET-1 also reduced IS (to 14+/-3%, P<0.05 vs control). The ET(A) receptor antagonist blocked the reduction in IS induced by ET-1 (IS 47+/-8% after LU+ET-1; P< 0.05 vs ET-1). In protocol II, Glib and 5-HD abolished the cardioprotective effect induced by ET-1 (IS 48+/ 7% after Glib+ET-1 and 42+/-5% after ET-1+5-HD vs 18+/-4% after ET-1 alone; P<0.05). In conclusion, administration of ET-1 before ischaemia resulted in a PC like cardioprotective effect. This effect is mediated via the ET(A) receptor and activation of mitoK(ATP) channels. PMID- 15655528 TI - The selectivity of beta-adrenoceptor antagonists at the human beta1, beta2 and beta3 adrenoceptors. AB - Beta-adrenoceptor antagonists ("beta-blockers") are one of the most widely used classes of drugs in cardiovascular medicine (hypertension, ischaemic heart disease and increasingly in heart failure) as well as in the management of anxiety, migraine and glaucoma. Where known, the mode of action in cardiovascular disease is from antagonism of endogenous catecholamine responses in the heart (mainly at beta1-adrenoceptors), while the worrisome side effects of bronchospasm result from airway beta2-adrenoceptor blockade. The aim of this study was to determine the selectivity of beta-antagonists for the human beta-adrenoceptor subtypes. (3)H-CGP 12177 whole cell-binding studies were undertaken in CHO cell lines stably expressing either the human beta1-, beta2- or the beta3-adrenoceptor in order to determine the affinity of ligands for each receptor subtype in the same cell background. In this study, the selectivity of well-known subtype selective ligands was clearly demonstrated: thus, the selective beta1 antagonist CGP 20712A was 501-fold selective over beta2 and 4169-fold selective over beta3; the beta2-selective antagonist ICI 118551 was 550- and 661-fold selective over beta1 and beta3, respectively, and the selective beta3 compound CL 316243 was 10 fold selective over beta2 and more than 129-fold selective over beta1. Those beta2-adrenoceptor agonists used clinically for the treatment of asthma and COPD were beta2 selective: 29-, 61- and 2818-fold for salbutamol, terbutaline and salmeterol over beta1, respectively. There was little difference in the affinity of these ligands between beta1 and beta3 adrenoceptors. The clinically used beta antagonists studied ranged from bisoprolol (14-fold beta1-selective) to timolol (26-fold beta2-selective). However, the majority showed little selectivity for the beta1- over the beta2-adrenoceptor, with many actually being more beta2 selective. This study shows that the beta1/beta2 selectivity of most clinically used beta-blockers is poor in intact cells, and that some compounds that are traditionally classed as "beta1-selective" actually have higher affinity for the beta2-adrenoceptor. There is therefore considerable potential for developing more selective beta-antagonists for clinical use and thereby reducing the side-effect profile of beta-blockers. PMID- 15655527 TI - Voltage-dependent inhibition of recombinant NMDA receptor-mediated currents by 5 hydroxytryptamine. AB - The effect of 5-HT and related indolealkylamines on heteromeric recombinant NMDA receptors expressed in Xenopus oocytes was investigated using the two-electrode voltage-clamp recording technique. In the absence of external Mg(2+) ions, 5-HT inhibited NMDA receptor-mediated currents in a concentration-dependent manner. The inhibitory effect of 5-HT was independent of the NR1a and NR2 subunit combination. The inhibition of glutamate-evoked currents by 5-HT was use- and voltage-dependent. The voltage sensitivity of inhibition for NR1a+NR2 subunit combinations by 5-HT was similar, exhibiting an e-fold change per approximately 20 mV, indicating that 5-HT binds to a site deep within the membrane electric field. The inhibition of the open NMDA receptor by external Mg(2+) and 5-HT was not additive, suggesting competition between Mg(2+) and 5-HT for a binding site in the NMDA receptor channel. The concentration-dependence curves for 5-HT and 5 methoxytryptamine (5-MeOT) inhibition of NMDA receptor-mediated currents are shifted to the right in the presence of external Mg(2+). The related indolealkylamines inhibited glutamate-evoked currents with the following order of inhibitory potency: 5-MeOT=5-methyltryptamine>tryptamine>7-methyltryptamine>5 HT>>tryptophan=melatonin. Taken together, these data suggest that 5-HT and related compounds can attenuate glutamate-mediated excitatory synaptic responses and may provide a basis for drug treatment of excitoxic neurodegeneration. PMID- 15655529 TI - Purine- and pyrimidine-induced responses and P2Y receptor characterization in the hamster proximal urethra. AB - 1. Purine and pyrimidine compounds were investigated on hamster proximal urethral circular smooth muscle preparations. In situ hybridization studies were carried out to localize P2Y(1), P2Y(2), P2Y(4) and P2Y(6) mRNA. Protein expression was studied using Western blotting analysis with antibodies against P2Y(1) and P2Y(2) receptors. 2. The hamster urethra relaxed with an agonist potency order of: 2 MeSADP>beta,gamma-meATP=ATP=adenosine=ADP>2-MeSATP>alpha,beta meATP>TTP>CTP=UTP>GTP=UDP. The high potency of 2-MeSADP is suggestive of an action via P2Y(1) receptors. Although the order is not characteristic for any known single P2Y receptor subtype, it may represent a combination of P2Y receptor subtypes. 4. The selective P2Y(1) receptor antagonist MRS2179 inhibited ATP-, 2 MeSADP-, 2-MeSATP-, beta,gamma-meATP-, and to a lesser degree alpha,beta-meATP induced responses. 3. Adenosine, but not ATP, was inhibited by the adenosine receptor antagonist 8-phenyltheophylline, indicating that ATP was not acting via adenosine following enzymatic breakdown. 5. Western blotting analysis showed the expression of both P2Y(1) and P2Y(2) receptors, confirming the results obtained with in situ hybridization that showed the expression of both P2Y(1) and P2Y(2), but not P2Y(4) or P2Y(6) mRNA, in smooth muscle layers of the hamster proximal urethra. 6. It is proposed that the relaxant response of the urethra to ATP may be evoked through the activation of the combination of receptors for P2Y(1) and to a lesser extent P2Y(2) receptors, which may mediate a trophic effect in addition. A P2Y subtype responsive to alpha,beta-meATP and P1 receptors may contribute to urethral smooth muscle relaxation. PMID- 15655531 TI - The dual PPARalpha/gamma agonist, ragaglitazar, improves insulin sensitivity and metabolic profile equally with pioglitazone in diabetic and dietary obese ZDF rats. AB - In 6- and 10-week-old obesity-prone (fa/fa) Zucker diabetic fatty (ZDF) rats, effects of prevention and intervention therapies, respectively, were compared between PPARalpha/gamma agonist, ragaglitazar (RAGA) and separate PPARgamma and alpha agonists, pioglitazone (PIO) and bezafibrate (BF). In a separate study, lean (+/+) ZDF rats fed highly palatable chow to induce dietary obesity and insulin resistance were treated similarly. To test insulin-secretory capacity, all animals underwent a hyperglycaemic clamp. Insulin sensitivity was improved equally by RAGA and PIO in fa/fa rats subjected to both prevention and intervention treatments (e.g., prevention HOMA-IR: -71 and -72%, respectively), as was hyperglycaemia (both -68%). BF had no effect on either parameter in any study. Plasma lipids were markedly reduced (by 48-77%) by RAGA in all studies, equivalent to PIO, but to a greater extent than BF. RAGA improved beta-cell function (HOMA-beta) more than three-fold with prevention and intervention therapies, whereas PIO showed improvement only in intervention therapy. Consistent with improved insulin sensitivity, glucose infusion rate during the clamp was 60% higher in RAGA-treated animals subjected to prevention therapy, but there was little additional insulin-secretory response, suggesting that insulin secretion was already maximal.Thus, RAGA and PIO equally improve metabolic profile in ZDF rats, particularly when administered early in the course of diabetes. They also improve beta-cell function, although this is better demonstrated through indices incorporating fasting insulin and glucose concentrations than through the hyperglycaemic clamp technique in this model. PMID- 15655530 TI - Endothelium-dependent contractions in hypertension. AB - 1. Endothelial cells, under given circumstances, can initiate contraction (constriction) of the vascular smooth muscle cells that surround them. Such endothelium-dependent, acute increases in contractile tone can be due to the withdrawal of the production of nitric oxide, to the production of vasoconstrictor peptides (angiotensin II, endothelin-1), to the formation of oxygen-derived free radicals (superoxide anions) and/or the release of vasoconstrictor metabolites of arachidonic acid. The latter have been termed endothelium-derived contracting factor (EDCF) as they can contribute to moment-to moment changes in contractile activity of the underlying vascular smooth muscle cells. 2. To judge from animal experiments, EDCF-mediated responses are exacerbated by aging, spontaneous hypertension and diabetes. 3. To judge from human studies, they contribute to the blunting of endothelium-dependent vasodilatations in aged subjects and essential hypertensive patients. 4. Since EDCF causes vasoconstriction by activation of the TP-receptors on the vascular smooth muscle cells, selective antagonists at these receptors prevent endothelium dependent contractions, and curtail the endothelial dysfunction in hypertension and diabetes. PMID- 15655533 TI - Role of SK(Ca) and IK(Ca) in endothelium-dependent hyperpolarizations of the guinea-pig isolated carotid artery. AB - 1. This study was designed to determine whether the endothelium-dependent hyperpolarizations evoked by acetylcholine in guinea-pig carotid artery involve a cytochrome P450 metabolite and whether they are linked to the activation of two distinct populations of endothelial K(Ca) channels, SK(Ca) and IK(Ca.) 2. The membrane potential was recorded in the vascular smooth muscle cells of the guinea pig isolated carotid artery. All the experiments were performed in the presence of N(omega)-L-nitro arginine (100 microM) and indomethacin (5 microM). 3. Under control conditions (Ca(2+): 2.5 mM), acetylcholine (10 nM to 10 muM) induced a concentration- and endothelium-dependent hyperpolarization of the vascular smooth muscle cells. Two structurally different specific blockers of SK(Ca), apamin (0.5 microM) or UCL 1684 (10 microM), produced a partial but significant inhibition of the hyperpolarization evoked by acetylcholine whereas charybdotoxin (0.1 microM) and TRAM-34 (10 microM), a nonpeptidic and specific blocker of IK(Ca), were ineffective. In contrast, the combinations of apamin plus charybdotoxin, apamin plus TRAM-34 (10 microM) or UCL 1684 (10 microM) plus TRAM-34 (10 microM) virtually abolished the acetylcholine-induced hyperpolarization. 4. In the presence of a combination of apamin and a subeffective dose of TRAM-34 (5 microM), the residual hyperpolarization produced by acetylcholine was not inhibited further by the addition of either an epoxyeicosatrienoic acid antagonist, 14,15-EEZE (10 microM) or the specific blocker of BK(Ca), iberiotoxin (0.1 microM). 5. In presence of 0.5 mM Ca(2+), the hyperpolarization in response to acetylcholine (1 microM) was significantly lower than in 2.5 mM Ca(2+). The EDHF-mediated responses became predominantly sensitive to charybdotoxin or TRAM 34 but resistant to apamin. 6. This investigation shows that the production of a cytochrome P450 metabolite, and the subsequent activation of BK(Ca), is unlikely to contribute to the EDHF-mediated responses in the guinea-pig carotid artery. Furthermore, the EDHF-mediated response involves the activation of both endothelial IK(Ca) and SK(Ca) channels, the activation of either one being able to produce a true hyperpolarization. PMID- 15655532 TI - Protein kinase Cgamma mediates ethanol withdrawal hyper-responsiveness of NMDA receptor currents in spinal cord motor neurons. AB - The present studies were designed to test the hypothesis that neuronal-specific protein kinase Cgamma (PKCgamma) plays a critical role in acute ethanol withdrawal hyper-responsiveness in spinal cord. Patch-clamp studies were carried out in motor neurons in neonatal rat spinal cord slices. Postsynaptic currents were evoked by brief pulses of 2 mM N-methyl-D-aspartic acid (NMDA) in the presence of bicuculline methiodide 10 microM; strychnine 5 microM and tetrodotoxin 0.5 microM. Both ethanol depression and withdrawal hyper responsiveness of NMDA-evoked currents are dependent on increases in intracellular Ca(2+). Blocking intracellular increase in Ca(2+) by 30 mM 1,2 bis(2-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid (BAPTA) not only decreased the ethanol-induced depression of NMDA-evoked currents (33+/-5% in control vs 20+/-3% in BAPTA, P<0.05) but also eliminated acute ethanol withdrawal hyper responsiveness. Immunohistochemistry studies revealed that neonatal spinal cord motor neurons contain an abundance of nuclear PKCgamma. Exposure to ethanol (100 mM) induced PKCgamma translocation from the nucleus to cytoplasm in motor neurons. Pretreatment with the gamma-isozyme-specific peptide PKC inhibitor, gammaV5-3, blocked ethanol-induced translocation and also blocked withdrawal hyper-responsiveness. The results show that PKCgamma mediates ethanol withdrawal hyper-responsiveness in spinal motor neurons; the results may be relevant to some symptoms of ethanol withdrawal in vivo. PMID- 15655534 TI - Captopril improves cerebrovascular structure and function in old hypertensive rats. AB - We examined the effects of an angiotensin-converting enzyme inhibitor (ACEI), captopril, on cerebral arterioles in young and old spontaneously hypertensive rats (SHR). Animals were anesthetized with sodium pentobarbitone (60 mg kg(-1) day(-1)). We measured cerebral blood flow (CBF, arbitrary units) and cerebral arteriolar internal diameter (ID, mum) prior to and during stepwise hypotension (SH) in 6- (WKY-6) and 15-month-old (WKY-15) Wistar Kyoto rats and in age-matched SHR that were untreated (SHR-6 and SHR-15) or treated for 3 months with captopril (SHR-6C, 105+/-2 mg kg(-1) day(-1) and SHR-15C, 94+/-1 mg kg(-1) day(-1)). ID and cross-sectional area of the vessel wall (CSA) were measured in deactivated (EDTA) cerebral arterioles during a second SH. Captopril decreased the lower limit of CBF autoregulation (61+/-6 in SHR-6C and 51+/-2 in SHR-15C vs 52+/-6 in WKY-6 and 62+/-7 in WKY-15 and 83+/-14 mmHg in SHR-6 and 120+/-19 mmHg in SHR-15; P<0.05) and CSA (510+/-21 in SHR-6C and 585+/-25 in SHR-15C vs 529+/-12 in WKY-6 and 549+/-20 in WKY-15 and 644+/-38 mmHg in SHR-6 and 704+/-38 mmHg in SHR-15; P<0.05). Captopril increased cerebral arteriolar external diameter of SHR (105+/ 5 in SHR-6C and 94+/-4 in SHR-15C vs 125+/-8 in WKY-6 and 108+/-3 in WKY-15 and 83+/-2 mmHg in SHR-6 and 80+/-2 mmHg in SHR-15 for a pial arteriolar pressure step of 35-39 mmHg; P<0.05). Captopril attenuated increases in cerebral arteriolar distensibility in young SHR. Thus, ACEIs attenuate eutrophic and hypertrophic inward remodeling of cerebral arterioles in young and old SHR, thus decreasing the lower limit of CBF autoregulation. PMID- 15655535 TI - Transition from placental to air breathing stimulates haem-oxygenase-1 expression without functional consequence for pulmonary vascular adaptation in pigs and mice. AB - 1. In systemic vessels, haem-oxygenase (HO) is induced during oxidative stress and known to modulate vasodilatation and vascular remodelling. At birth, with the transition from placental to air breathing, the pulmonary vessels are exposed to oxidative stress and undergo well-documented remodelling processes. Thus, we investigated the role of HO in the lung during adaptation to extra-uterine life using a pig and mouse model. In addition to the novel data presented with regard to one isoform, HO-1, this study is among the first to describe the pulmonary vascular remodelling in the mouse after birth. 2. We show, for the first time, that another isoform, HO-2, is present constitutively at birth and HO-1 protein is induced in the porcine and murine lung after birth in vascular and airway structures, peaking at 14 days in the pig and at about 4 days in the mouse. Furthermore, we show that HO-1 mRNA declines after birth in the mouse lung. 3. Inhibitors of HO did not modify vasodilator responses in vessels from 14-day-old pigs. 4. Moreover, lungs from HO-1-deficient mice developed normally after birth. 5. HO-1 is induced at birth but plays no role in the development of vasodilator responses or remodelling that occurs at this time. These data suggest that HO-1 expression at birth is a redundant response to oxidative stress in the lungs of healthy mammals. However, it remains possible that this pathway protects if complications occur during or after birth. PMID- 15655537 TI - The prognostic significance of micrometastases in node-negative squamous cell carcinoma of the vulva. AB - Nodal involvement is one of the most significant prognostic factors in squamous cell carcinoma (SCC) of the vulva. We conducted a retrospective analysis of 31 women with histologically node-negative SCC from a population-based cohort of Grampian women. Median follow-up was 42 months after radical vulvectomy with groin node dissection. In total, 13 women (42%) were found to have micrometastases on immunohistochemistry. The risk of recurrence was almost 20 fold higher in those with micrometastases compared to those without (hazard ratio=19.6 (95% CI 2.3-171). PMID- 15655536 TI - Luteinising hormone-releasing hormone analogue reverses the cell adhesion profile of EGFR overexpressing DU-145 human prostate carcinoma subline. AB - Cetrorelix, a luteinising hormone-releasing hormone (LHRH) analogue, has been shown to limit growth of the human androgen-independent prostate cell line DU 145, although other inhibitory actions may also be affected. Both growth and invasion of DU-145 cells are linked to autocrine epidermal growth factor receptor (EGFR) signalling. Invasiveness requires not only cells to migrate to conduits, but also reduced adhesiveness between tumour cells to enable separation from the tumour mass. Thus, we investigated whether Cetrorelix alters the DU-145 cell-cell adhesion and if this occurs via altered EGFR signalling. Pharmacologic levels of Cetrorelix limited the invasiveness of a highly invasive DU-145 subline overexpressing full-length EGFR (DU-145 WT). Extended exposure of the cells to Cetrorelix resulted in increased levels of the cell-cell adhesion complex molecules E-cadherin, alpha- and beta-catenin, and p120. Puromycin blocked the increases in E-cadherin and beta-catenin levels, suggesting that de novo protein synthesis is required. The Cetrorelix effect appears to occur via transmodulation of EGFR by a protein kinase C (PKC)-dependent mechanism, as there were no changes in DU-145 cells expressing EGFR engineered to negate the PKC transattenuation site (DU-145 A654); downregulation of EGFR signalling produced a similar upregulation in adhesion complex proteins, further suggesting a role for autocrine signalling. Cetrorelix increased the cell-cell adhesiveness of DU-145 WT cells to an extent similar to that seen when autocrine EGFR signalling is blocked; as expected, DU-145 A654 cell-cell adhesion also was unaffected by Cetrorelix. The increased adhesiveness is expected as the adhesion complex molecules moved to the cells' periphery. These data offer direct insight into the possible crosstalk pathways between the LHRH and EGFR receptor signalling. The ability of Cetrorelix to downregulate EGFR signalling and subsequently reverse the antiadhesiveness found in metastatic prostate cancer highlights a novel potential target for therapeutic strategies. PMID- 15655538 TI - Evolving treatment strategies for myeloma. AB - In this article, we will discuss how treatment strategies for myeloma have evolved and outline the challenges now faced following the introduction of a number of novel active agents. In particular, we will focus on how achieving a maximum response and maintaining such responses is becoming a key therapeutic strategy and how novel agents can be used to achieve this in the context of current strategies such as autologous transplantation. PMID- 15655540 TI - Phase II study of capecitabine and cisplatin as first-line combination therapy in patients with gastric cancer recurrent after fluoropyrimidine-based adjuvant chemotherapy. AB - To evaluate the efficacy and safety of capecitabine and cisplatin in patients with recurrent gastric cancer after fluoropyrimidine-based adjuvant therapy. Patients with histologically confirmed and measurable advanced gastric cancer that had relapsed after fluoropyrimidine-based adjuvant chemotherapy received oral capecitabine (1250 mg m(-2) twice daily, days 1-14) and intravenous cisplatin (60 mg m(-2) over 1 h, day 1) every 3 weeks. In total, 32 patients were enrolled, of whom 30 were evaluable for efficacy and 32 for safety. A median of 5 cycles (range 1-10) was administered. One patient achieved a complete response and eight had partial responses, giving an overall response rate of 28% (95% CI, 13-44%). The median time to progression and median overall survival were 5.8 months (95% CI, 4.1-7.5 months) and 11.2 months (95% CI, 5.5-16.9 months), respectively. Grade 3 neutropenia and thrombocytopenia were observed in 38 and 6% of patients, respectively. Grade 2/3 nonhaematological toxicities included diarrhoea (19%), stomatitis (19%) and hand-foot syndrome (31%). No grade 4 toxicity, neutropenic fever or treatment-related deaths occurred. Capecitabine in combination with cisplatin was effective and well tolerated as first-line treatment in patients with recurrent gastric cancer after fluoropyrimidine-based adjuvant chemotherapy. PMID- 15655539 TI - Cytotoxic and antiangiogenic activity of AW464 (NSC 706704), a novel thioredoxin inhibitor: an in vitro study. AB - AW464 (NSC 706704) is a novel benzothiazole substituted quinol compound active against colon, renal and certain breast cancer cell lines. NCI COMPARE analysis indicates possible interaction with thioredoxin/thioredoxin reductase, which is upregulated under hypoxia. Through activity on HIF1alpha, VEGF levels are regulated and angiogenesis controlled. A thioredoxin inhibitor could therefore exhibit enhanced hypoxic toxicity and indirect antiangiogenic effects. In vitro experiments were performed on colorectal and breast cancer cell lines under both normoxic and hypoxic conditions and results compared against those obtained with normal cell lines, fibroblasts and keratinocytes. Antiangiogenic effects were studied using both large and microvessel cells. Indirect antiangiogenic effects (production of angiogenic growth factors) were studied via ELISA. We show that AW464 exerts antiproliferative effects on tumour cell lines as well as endothelial cells with an IC(50) of approximately 0.5 microM. Fibroblasts are however resistant. Proliferating, rather than quiescent, endothelial cells are sensitive to the drug indicating potential antiangiogenic rather than antivascular action. Endothelial differentiation is also inhibited in vitro. Hypoxia (1% O(2) for 48 h) sensitises colorectal cells to lower drug concentrations, and in HT29s greater inhibition of VEGF is observed under such conditions. In contrast, bFGF levels are unaffected, suggesting possible involvement of HIF1alpha. Thus, AW464 is a promising chemotherapeutic drug that may have enhanced potency under hypoxic conditions and also additional antiangiogenic activity. PMID- 15655542 TI - Singlet oxygen luminescence as an in vivo photodynamic therapy dose metric: validation in normal mouse skin with topical amino-levulinic acid. AB - Although singlet oxygen ((1)O(2)) has long been proposed as the primary reactive oxygen species in photodynamic therapy (PDT), it has only recently been possible to detect it in biological systems by its luminescence at 1270 nm. Having previously demonstrated this in vitro and in vivo, we showed that cell survival was strongly correlated to the (1)O(2) luminescence in cell suspensions over a wide range of treatment parameters. Here, we extend this to test the hypothesis that the photobiological response in vivo is also correlated with (1)O(2) generation, independent of individual treatment parameters. The normal skin of SKH1-HR hairless mice was sensitised with 20% amino-levulinic acid-induced protoporophyrin IX and exposed to 5, 11, 22 or 50 J cm(-2) of pulsed 523 nm light at 50 mW cm(-2), or to 50 J cm(-2) at 15 or 150 mW cm(-2). (1)O(2) luminescence was measured during treatment and the photodynamic response of the skin was scored daily for 2 weeks after treatment. As observed by other authors, a strong irradiance dependence of the PDT effect was observed. However, in all cases the responses increased with the (1)O(2) luminescence, independent of the irradiance, demonstrating for the first time in vivo an unequivocal mechanistic link between (1)O(2) generation and photobiological response. PMID- 15655541 TI - Survivin expression in ovarian cancer and its correlation with clinico pathological, surgical and apoptosis-related parameters. AB - We investigated the association of survivin expression with prognosis and other apoptosis-related biological factors in 110 primary ovarian cancer patients admitted to the Division of Gynecologic Oncology, Catholic University of Rome. Immunohistochemistry was performed on formalin-fixed, paraffin-embedded sections by using polyclonal antibody ab469 for survivin, and mouse monoclonal antibodies (clone 124 and DO-7), for bcl-2 and p53, respectively. Cytoplasmic survivin immunoreaction was observed in 84.5% cases, while nuclear survivin immunostaining was observed in 29.1% cases. We failed to find any relationship between cytoplasmic survivin positivity rate and any of the parameters examined. Serous tumours showed a lower percentage of nuclear survivin positivity with respect to other histotypes (20.5 vs 48.6%, respectively; P-value=0.004). The percentage of nuclear survivin positivity was higher in cases subjected to primary tumour cytoreduction (43.5%), with respect to patients subjected to exploratory laparotomy (20%) (P=0.024). Bcl-2 and p53 were, respectively, expressed in 27.3 and 60.0% of the cases and their expression was not correlated with survivin status. During the follow-up period, progression and death of disease were observed in 68 (61.8%) and 53 (48.2%) cases, respectively. There was no difference in time to progression and overall survival according to survivin status in ovarian cancer patients. In conclusion, in our experience, the immunohistochemical assessment of survivin status does not seem to be helpful in the prognostic characterisation of ovarian cancer. A more in depth investigation of the complex physiology of divergent survivin variants is needed in order to clarify the biological and the clinical role of differentially located survivin isoforms. PMID- 15655543 TI - Unfavourable expression of pharmacologic markers in mucinous colorectal cancer. AB - Patients with mucinous colorectal cancer generally have worse prognoses than those with the nonmucinous variety. The reason for this disparity is unclear, but may result from a differential response to adjuvant chemotherapy. We examined known molecular markers for response to common chemotherapy in these two histological subtypes. In all, 21 patients with mucinous and 30 with nonmucinous Dukes C colorectal cancer were reviewed for demographic data and outcome. Total RNA from the tumours and adjacent normal mucosa was isolated and reverse transcribed. Quantitative expression levels of drug pathway genes were determined using TaqMan RT-PCR (5-fluorouracil (5-FU): TYMS, DPYD, ECGF1; oxaliplatin: GSTP1 (glutathione S-transferase pi), ERCC1 and 2; irinotecan: ABCB1, ABCG2, CYP3A4, UGT1A1, CES2, TOP1). Mucinous tumours significantly overexpressed both TYMS and GSTP1 relative to nonmucinous tumours and patient-matched normal mucosa. No significant differences in expression of the remaining markers were found. Mean follow-up was 20 months; 17 patients had recurrent disease. Among patients receiving 5-FU, those with mucinous tumours experienced shorter disease-free survival (DFS) than those with nonmucinous tumours (median DFS 13.8 vs 46.5 months, P=0.053). Mucinous colorectal cancer overexpresses markers of resistance to 5-FU and oxaliplatin. Likewise, DFS may be decreased in patients with mucinous tumours who receive 5-FU. The presence of mucin should be carefully evaluated in developmental trials of new agents for treating colorectal cancer. PMID- 15655544 TI - The human papillomavirus E6 and E7 inducible oncogene, hWAPL, exhibits potential as a therapeutic target. AB - Here we show that human papillomavirus (HPV) E6 and E7 oncoproteins induce hWAPL expression. In addition, small interfering RNA (siRNA) of hWAPL suppressed the growth of tumours derived from SiHa cells in nude mice. Thus, hWAPL may be one of the effective targets of uterine cervical cancer therapy. PMID- 15655545 TI - Tyrphostins reduce chemotherapy-induced intestinal injury in mice: assessment by a biochemical assay. AB - Intestinal injury that results from chemotherapy belongs to the major factors of dose-limitation in tumour therapy. The tyrphostins AG1714 and AG1801 reduce cisplatin and 5-FU-induced small intestinal mucosal damage, using a quantitative biochemical assay. The assay is based on the determination of the enzymatic activity of gamma-glutamyl transpeptidase, a marker of the brush border epithelium of the small intestine. PMID- 15655546 TI - Population-based monitoring of cancer patient survival in situations with imperfect completeness of cancer registration. AB - Selective underascertainment of cases may bias estimates of cancer patient survival. We show that the magnitude of potential bias strongly depends on the time periods affected by underascertainment and on the type of survival analysis (cohort analysis vs period analysis). We outline strategies on how to minimise or overcome potential biases. PMID- 15655547 TI - The predictive value of p53, p53R2, and p21 for the effect of chemoradiation therapy on oesophageal squamous cell carcinoma. AB - The p53 family regulates cell-cycle arrest, triggers apoptosis or is involved in repair of DNA damage. In the present study, we analysed the expression of some p53 family proteins and their responses to chemoradiation therapy (CRT) in cases of oesophageal squamous cell carcinoma (ESCC). We immunohistochemically investigated the relationship between p53, p53R2, and p21 expression in biopsy specimens of untreated primary tumours and their clinical and histological responses to CRT in 62 patients with ESCC. Chemoradiation therapy consisted of 5 fluorouracil plus cisplatin and 40 Gy of radiation. The rates of clinical and histological responses (complete or partial) to CRT were 71.0% (clinical) and 52.8% (histological). The rate of positive expression was 43.5% for p53, 37.1% for p53R2, and 54.8% for p21 expression. Statistically significant correlations were found between p53 or p53R2 expression and favourable response to CRT (P=0.0001 or 0.041 clinical, P=0.016 or 0.0018 histological, respectively). Furthermore, in p53-negative tumours, CRT was more effective in tumours with p53R2 negative expression than those with p53R2 positive expression (P=0.0014). We demonstrated that the negative expression of p53 and p53R2 expression was closely related to the effect of CRT and should predict the CRT outcome in patients with ESCC. PMID- 15655548 TI - Years of life lost (YLL) from cancer is an important measure of population burden -and should be considered when allocating research funds. AB - Recently, cancer mortality has been compared to research spending by the National Cancer Research Institute (NCRI), whose research budget is approximately pound sterling 250 million. The analysis shows a mis-match between mortality and research spending. As well as crude mortality rates, other measures of cancer burden should be considered because they contribute additional information. 'Years of life lost' (YLL) summed over each individual dying after a diagnosis of cancer represents a population-based mortality indicator of the impact of that disease on society. Years of life lost divided by the number of deaths for each cancer site produces an additional statistic, the average years of life lost (AYLL), which is a measure of the burden of cancer to the individual patient. For 17 cancer sites where data are available, four tumour sites have a rather large difference in mortality, comparing YLL to crude mortality. Years of life lost shows the population burden from cancers of the ovary, cervix, and CNS to be rather larger than suggested by crude mortality, despite screening programmes for cervix cancer. Using YLL, the underprovision of funding for lung cancer research is similar to that reported using percentage mortality. Breast cancer and leukaemia receive a relatively higher research spend than the population burden of these cancers, and the spending on leukaemia is quite extreme. Prostate cancer has a low per cent YLL but attracts a moderate amount of research spending. The use of AYLL as an indicator of individual cancer burden considerably changes the ranking of the mortality from different tumours. The mean AYLL is 12.5 years. Prostate cancer has the lowest AYLL, only 6.1 years; brain tumour patients have the highest, at just over 20 years. Comparing AYLL to research spending suggests four 'Cinderella' cancer sites with high individual cancer burden but low research spending: CNS tumours, cervix and kidney cancers, and melanoma. Breast cancer and leukaemia have roughly average AYLL but a considerable excess of research spending. YLL emphasises the discrepancy between research spending and mortality, and may be helpful for decisions concerning research support. Average years of life lost measures the burden to individual patients and may be helpful where individuals' needs are relevant, such as palliative care. As well as crude mortality, more subtle and comprehensive calculations of mortality statistics would be useful in debates on research funding and public health issues. PMID- 15655549 TI - Detection of Ki-ras mutations in tissue and plasma samples of patients with pancreatic cancer using PNA-mediated PCR clamping and hybridisation probes. AB - In the present study, we combined the PCR-clamping approach with melting curve analysis using mutant specific hybridisation probes and wild-type specific peptide nucleic acids (PNAs) to determine the genotypes of the most frequent point mutation in codon 12 of the proto-oncogene Ki-ras in tissue and plasma samples of patients with pancreatic cancer. The sensitivity of our assay was 1-5 x 10(-5). The melting curve analysis of tissue samples of four patients revealed two valine mutations, one none-valine mutation and one wild-type sequence. Ki-ras alterations were found in 28% of DNAs (18 out of 64) of nonrelated plasma samples of 10 patients with ductal adenocarcinoma of the pancreas. The valine mutation was the predominantly detected gene alteration (83%). Out of ten patients investigated, four patients (40%) became positive during clinical observation with respect to Ki-ras mutation. All four patients exhibited progressive disease and high levels of tumour marker CA 19-9. In conclusion, the one-step procedure discribed may be a useful clinical tool for analysing Ki-ras point mutations in tissue and plasmas samples. In addition, this method can be adapted for simultanous detection of multiple mutations and quantitation. PMID- 15655550 TI - Survivin, Survivin-2B, and Survivin-deItaEx3 expression in medulloblastoma: biologic markers of tumour morphology and clinical outcome. AB - Survivin is an apoptotic inhibitor that is expressed at high levels in a variety of malignancies. Survivin has four known alternative splice forms (Survivin, Survivin-2B, Survivin-deltaEx3, and Survivin-3B), and the recent literature suggests that these splice variants have unique functions and subcellular localisation patterns. We evaluated 19 fresh-frozen paediatric medulloblastomas for the expression of three Survivin isoforms by quantitative PCR. Survivin was most highly expressed when compared with normal cerebellar tissue. We also investigated Survivin protein expression in 40 paraffin-embedded paediatric medulloblastoma tumours by immunohistochemistry. We found a statistically significant association between the percentage of Survivin-positive cells and histologic subtype, with the large-cell-anaplastic variant expressing Survivin at higher levels than the classic subtype. We also found a statistically significant relationship between the percent of Survivin-positive cells in the tumours and clinical outcome, with higher levels of Survivin correlating with a worse prognosis. In summary, our study demonstrates a role for Survivin as a marker of tumour morphology and clinical outcome in medulloblastoma. Survivin may be a promising future prognostic tool and potential biologic target in this malignancy. PMID- 15655551 TI - Tumour-derived exosomes and their role in cancer-associated T-cell signalling defects. AB - Dendritic and lymphoid 'exosomes' regulate immune activation. Tumours release membranous material mimicking these 'exosomes,' resulting in deletion of reactive lymphocytes. Tumour-derived 'exosomes' have recently been explored as vaccines, without analysis of their immunologic consequences. This investigation examines the composition of tumour-derived 'exosomes' and their effects on T lymphocytes. Membranous materials were isolated from ascites of ovarian cancer patients (n=6) and Western immunoblotting was performed for markers associated with 'exosomes.' Using cultured T cells, 'exosomes' were evaluated for suppression of CD3-zeta and JAK 3 expressions and induction of apoptosis, measured by DNA fragmentation. 'Exosome' components mediating suppression of CD3-zeta were isolated by continuous eluting electrophoresis and examined by Western immunoblotting. 'Exosomes' were shown to be identical with previously characterised shed membrane vesicles by protein staining and TSG101 expression. 'Exosomes' expressed class I MHC, placental alkaline phosphatase, B23/nucleophosmin, and FasL. 'Exosomes' suppressed expression of T-cell activation signalling components, CD3-zeta and JAK 3 and induced apoptosis. CD3-zeta suppression was mediated by two components: 26 and 42 kDa. Only the 42 kDa component reacted with anti-FasL antibody. These results indicate that, while 'exosomes' express tumour antigens, leading to their proposed utility as tumour vaccines, they also can suppress T-cell signalling molecules and induce apoptosis. PMID- 15655552 TI - Lack of EGF receptor contributes to drug sensitivity of human germline cells. AB - Germline mutations have been associated with generation of various types of tumour. In this study, we investigated genetic alteration of germline tumours that affect the drug sensitivity of cells. Although all germline tumour cells we tested were hypersensitive to DNA-damaging drugs, no significant alteration was observed in their DNA repair activity or the expression of DNA repair proteins. In contrast, germline tumours expressed very low level of epidermal growth factor receptor (EGFR) compared to drug-resistant ovarian cancer cells. An immunohistochemical analysis indicated that most of the primary germline tumours we tested expressed very low level of EGFR. In accordance with this, overexpression of EGFR in germline tumour cells showed an increase in drug resistance, suggesting that a lack of EGFR, at least in part, contributes to the drug sensitivity of germline tumours. PMID- 15655553 TI - Making sense of information about HPV in cervical screening: a qualitative study. AB - Introducing human papillomavirus (HPV) testing into cervical cancer screening has the potential to change the way that women understand cervical cancer, the psychological impact of abnormal screening results and the likelihood of future participation in screening. The study used in-depth interviews to examine how women make sense of information about HPV in the context of cervical cancer screening. A total of 74 women were recruited following participation in HPV testing. Women varied widely in their beliefs about the aetiology of cervical cancer and its relationship with sexual activity, as well as in their understanding of the sexually transmitted nature of HPV. While some women who understood that HPV is sexually transmitted were able to integrate this into their existing model of cervical cancer, others were shocked by the link between cervical cancer and sex, of which they had been previously unaware. Women were generally reassured to know that HPV is common, has no symptoms, can lie dormant for many years, can clear up on its own and need not raise concerns about transmission to sexual partners. Women's understanding of HPV varied considerably, even after participation in testing. The way in which information is presented to women will be crucial in minimising the negative psychological impact of testing positive and ensuring that participation in screening remains high. PMID- 15655554 TI - Clinical relevance of genetic instability in prostatic cells obtained by prostatic massage in early prostate cancer. AB - We investigated whether genetic lesions such as loss of heterozygosity (LOH) are detected in prostatic cells obtained by prostatic massage during early diagnosis of prostate cancer (CaP) and discussed their clinical relevance. Blood and first urine voided after prostatic massage were collected in 99 patients with total prostate-specific antigen (PSA) between 4 and 10 ng ml(-1), prior to prostate biopsies. Presence of prostatic cells was confirmed by quantitative RT-PCR analysis of PSA mRNA. Genomic DNA was analysed for LOH on six chromosomal regions. One or more allelic deletions were found in prostatic fluid from 57 patients analysed, of whom 33 (58%) had CaP. Sensitivity and specificity of LOH detection and PSA free to total ratio <15% for positive biopsy were respectively 86.7 and 44% (P=0.002) for LOH, and 55 and 74% (P=0.006) for PSA ratio <15%. Analysis of LOH obtained from prostatic tumours revealed similar patterns compared to prostatic fluid cells in 86% of cases, confirming its accuracy. The presence of LOH of urinary prostatic cells obtained after prostatic massage is significantly associated with CaP on biopsy and may potentially help to identify a set of patients who are candidates for further prostate biopsies. PMID- 15655555 TI - Cellular and complement-dependent cytotoxicity of Ep-CAM-specific monoclonal antibody MT201 against breast cancer cell lines. AB - MT201 is a fully human monoclonal IgG1 antibody with moderate affinity for epithelial cell adhesion molecule (Ep-CAM) being clinically developed for the treatment of carcinomas. Like many other clinically validated IgG1 monoclonal antibodies, MT201 primarily acts by antibody-dependent cellular cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). Here, we analysed ADCC and CDC induced by MT201 and, as reference, trastuzumab against a panel of nine human breast cancer cell lines expressing distinct surface levels of Ep-CAM and human epithelial growth factor receptor type 2 antigen. Maximal cell lysis by ADCC by MT201 and trastuzumab in the presence of peripheral mononuclear cells did not significantly differ when averaged over the nine cell lines, but showed marked differences with respect to individual cell lines. The extent of cell lysis at intermediate surface target density was highly variable, suggesting a dominant influence of other susceptibility factors. Only one breast cancer cell line was eliminated via CDC, but only by MT201. Resistance to CDC appeared to correlate with high expression levels of complement resistance factors. Our present data as well as recent data on the prevalence and prognostic relevance of Ep-CAM expression in metastatic breast cancer suggest that Ep-CAM-specific monoclonal IgG1 antibodies may have a significant therapeutic potential in the treatment of breast cancer. PMID- 15655556 TI - A peptide corresponding to the neuropilin-1-binding site on VEGF(165) induces apoptosis of neuropilin-1-expressing breast tumour cells. AB - There is increasing evidence that vascular endothelial growth factor (VEGF) has autocrine as well as paracrine functions in tumour biology. Vascular endothelial growth factor-mediated cell survival signalling occurs via the classical tyrosine kinase receptors Flt-1, KDR/Flk-1 and the more novel neuropilin (NP) receptors, NP-1 and NP-2. A 24-mer peptide, which binds to neuropilin-1, induced apoptosis of murine and human breast carcinoma cells, whereas a peptide directed against KDR had no effect. Both anti-NP1 and anti-KDR peptides induced endothelial cell apoptosis. Confocal microscopy using 5-(6)-carboxyfluorescein-labelled peptides showed that anti-NP1 bound to both tumour and endothelial cells, whereas anti-KDR bound endothelial cells only. This study demonstrates that NP-1 plays an essential role in autocrine antiapoptotic signalling by VEGF in tumour cells and that NP1-blockade induces tumour cell and endothelial cell apoptosis. Specific peptides can therefore be used to target both autocrine (tumour cells) and paracrine (endothelial cells) signalling by VEGF. PMID- 15655557 TI - Long-term follow-up of breast cancer survivors with post-mastectomy pain syndrome. AB - Post-mastectomy pain syndrome (PMPS) is a recognised complication of breast surgery although little is known about the long-term outcome of this chronic pain condition. In 1996, Smith et al identified a prevalence rate of PMPS of 43% among 408 women in the Grampian Region, Northeast Scotland. The aim of this study was to assess long-term outcome at 7-12 years postoperatively in this cohort of women, to describe the natural history of PMPS and impact of pain upon quality of life. Chronic pain and quality of life were assessed using the McGill Pain Questionnaire (MPQ) and Short Form-36 (SF-36). Of 175 women reporting PMPS in 1996, 138 were eligible for questionnaire follow-up in 2002. Mean time since surgery was 9 years (s.d. 1.8 years). A response rate of 82% (113 out of 138) was achieved; 59 out of 113 (52%) women reported continued PMPS and 54 out of 113 (48%) women reported their PMPS had resolved since the previous survey in 1996. Quality of life scores were significantly lower in women with persistent PMPS compared to those women whose pain had resolved. However, for women with persistent PMPS, SF-36 scores had improved over time. Risk factors for persistent PMPS included younger age and heavier weight. This study found that, of women reporting PMPS in 1996, half of those surveyed in 2002 continued to experience PMPS at a mean of 9 years after surgery. PMID- 15655558 TI - Evaluation of the fibroblast growth factor system as a potential target for therapy in human prostate cancer. AB - Overexpression of fibroblast growth factors (FGFs) has been implicated in prostate carcinogenesis. FGFs function via their high-affinity interactions with receptor tyrosine kinases, FGFR1-4. Expression of FGFR1 and FGFR2 in prostate cancer (CaP) was not found to be associated with clinical parameters. In this report, we further investigated for abnormal FGFR expression in prostate cancer and explore their significance as a potential target for therapy. The expression levels of FGFR3 and FGFR4 in CaP were examined and corroborated to clinical parameters. FGFR3 immunoreactivity in benign prostatic hyperplasia (BPH) and CaP (n=26 and 57, respectively) had similar intensity and pattern. Overall, FGFR4 expression was significantly upregulated in CaP when compared to BPH. A significant positive correlation between FGFR4 expression and Gleason score was noted: Gleason score 7-10 tumours compared to BPH (P<0.0001, Fisher's exact test), Gleason score 4-6 tumours compared to BPH (P<0.0004), and Gleason 7-10 compared to Gleason 4-6 tumours (P<0.005). FGFR4 overexpression was associated with an unfavourable outcome with decreased disease-specific survival (P<0.04, log rank test). FGF-induced signalling is targeted using soluble FGF receptor (sFGFR), potent inhibitor of FGFR function. We have previously shown that sFGFR expression via a replication-deficient adenoviral vector (AdlllcRl) suppresses in vitro FGF-induced signalling and function in human CaP DU145 cells. We tested the significance of inhibiting FGF function along with conventional therapeutic modalities in CaP, and confirmed synergistic effects on in vitro cell growth (proliferation and colony formation) by combining sFGFR expression and treatment with either Paclitaxel (Taxol) or gamma-irradiation. In summary, our data support the model of FGF system as valid target for therapy in CaP. PMID- 15655559 TI - Effect of Helicobacter pylori eradication on ongoing mutation of immunoglobulin genes in gastric MALT lymphoma. AB - Gastric low-grade mucosa-associated lymphoid tissue (low-grade MALT) lymphomas has been associated with Helicobacter pylori (H. pylori) infection. Although infiltrating T cells with specificity for H. pylori are known to stimulate the development of MALT lymphomas, the effect of H. pylori eradication on rearranged immunoglobulin heavy chain (IgH) genes of low-grade gastric MALT lymphomas is unclear. Gastric biopsies from five cases were investigated by cloning and sequence analysis of rearranged IgH genes before and after the treatment for H. pylori. In all cases, IgH genes were mutated from their germline counterpart. The frequency of intraclonal sequence heterogeneity before the eradication of H. pylori varied from 0.25 to 0.49%. Clones obtained from the tumours before the eradication of H. pylori in cases 1 and 2 showed a tendency to display a mutation pattern by positive antigen selection and their monoclonarity disappeared after the eradication. The frequency of intraclonal sequence heterogeneity of the clones obtained from cases 3, 4 and 5 (0.12% in case 3, 0.10% in 4 and 0.18% in 5) after the eradication of H. pylori was lower than that in tumours before the eradication (0.30% in case 3, 0.49% in 4 and not determined in 5). These findings suggest that low-grade gastric MALT lymphomas expand due to the persistent presence of H. pylori in vivo. The characteristic feature of tumour clones obtained from the tumours after the eradication of H. pylori is a very low intraclonal heterogeneity, which may potentially be independent of H. pylori. PMID- 15655560 TI - hMSH2 is the most commonly mutated MMR gene in a cohort of Greek HNPCC patients. AB - Germline mutations in genes encoding proteins involved in DNA mismatch repair are responsible for the autosomal dominantly inherited cancer predisposition syndrome hereditary nonpolyposis colorectal cancer (HNPCC). We describe here analysis of hMLH1 and hMSH2 in nine Greek families referred to our centre for HNPCC. A unique disease-causing mutation has been identified in seven out of nine (78%) families. The types of mutations identified are nonsense (five out of seven) (hMLH1: E557X, R226X; hMSH2: Q158X, R359X and R711X), a 2 bp deletion (hMSH2 1704_1705delAG) and a 2.2 kb Alu-mediated deletion encompassing exon 3 of the hMSH2 gene. The majority of mutations identified in this cohort are found in hMSH2 (77.7%). Furthermore, four of the mutations identified are novel. Finally, a number of novel benign variations were observed in both genes. This is the first report of HNPCC analysis in the Greek population, further underscoring the differences observed in the various geographic populations. PMID- 15655561 TI - Multicolour-banding fluorescence in situ hybridisation (mbanding-FISH) to identify recurrent chromosomal alterations in breast tumour cell lines. AB - Recurrent chromosome breakpoints in tumour cells may point to cancer genes, but not many have been molecularly characterised. We have used multicolour-banding fluorescence in situ hybridisation (mbanding-FISH) on breast tumour cell lines to identify regions of chromosome break created by inversions, duplications, insertions and translocations on chromosomes 1, 5, 8, 12 and 17. We delineate a total of 136 regions of break, some of them occurring with high frequency. We further describe two examples of dual-colour FISH characterisation of breakpoints, which target the 1p36 and 5p11-12 regions. Both breaks involve genes whose function is unknown to date. The mbanding-FISH strategy constitutes an efficient first step in the search for potential cancer genes. PMID- 15655562 TI - BDNF in schizophrenia, depression and corresponding animal models. AB - Understanding the etiology and pathogenesis schizophrenia and depression is a major challenge facing psychiatry. One hypothesis is that these disorders are secondary to a malfunction of neurotrophic factors. Inappropriate neurotrophic support during brain development could lead to structural disorganisation in which neuronal networks are established in a nonoptimal manner. Inadequate neurotrophic support in adult individuals could ultimately be an underlying mechanism leading to decreased capacity of brain to adaptive changes and increased vulnerability to neurotoxic damage. Brain-derived neurotrophic factor (BDNF) is a mediator involved in neuronal survival and plasticity of dopaminergic, cholinergic, and serotonergic neurons in the central nervous system (CNS). In this review, we summarize findings regarding altered BDNF in schizophrenia and depression and animal models, as well as the effects of antipsychotic and antidepressive treatments on the expression of BDNF. PMID- 15655563 TI - Dopaminergic polymorphisms associated with self-report measures of human altruism: a fresh phenotype for the dopamine D4 receptor. PMID- 15655564 TI - Psychopathological symptoms during interferon-alpha and ribavirin treatment: effects on virologic response. PMID- 15655565 TI - Expression of biomarkers modulating prostate cancer progression: implications in the treatment of the disease. AB - OBJECTIVES: To determine whether COX-2, bcl-2 and neoangiogenesis are related to human prostate cancer relapse after definitive surgical treatment and progression toward androgen independence and to evaluate the association between the patterns of these tumoral biomarkers and other standard clinico-pathological parameters (such as Gleason score, PSA, TNM stage). MATERIALS AND METHODS: We retrospectively analyzed the records on 126 prostate cancer samples from patients treated at our University Hospital from 1995 to 2002. The 72 patients with clinically localized disease (group 1) had undergone radical prostatectomy. Another 54 patients (group 2) had metastatic androgen-independent disease. Archived material relating to the subjects was then immunostained for bcl-2, COX 2 and CD-31, using an anti-bcl-2 monoclonal primary antibody, an anti-COX-2 polyclonal rabbit antibody and an anti-CD-31 monoclonal mouse antibody to evaluate neoangiogenesis (MVD, microvessel density). RESULTS: We found that bcl 2, COX-2 and MVD expression increased from group 1 to group 2. The intergroup difference was significant only for high MVD (P < 0.05). On the other hand, high MVD, high bcl-2 and high COX-2 expression was correlated with a higher PSA level (P < 0.01), whereas only a high MVD was also related with Gleason score (P < 0.05). We used univariate analysis to evaluate the prognostic impact of biologic and clinico-pathologic parameters on the disease-free-survival of 72 patients treated by radical prostatectomy. A total of 30 patients (41.6%) experienced biochemical relapse; bcl-2, COX-2 and MVD significantly correlated with disease relapse in these patients. In fact, we observed disease relapse in 24/45 (53%) with high bcl-2 expression, in 15/21 (71%) with a high MVD count and finally, in 30/58 (52%) with high COX-2 expression. Finally, PSA value and Gleason score were the only two biologic markers significantly associated to disease relapse in a multivariate analysis. CONCLUSIONS: Our results strongly support a role for bcl 2, COX-2 and angiogenesis in the development and progression of prostate cancer. Of course, we are aware of the small sample size considered in our study. Further investigations would better clarify the prognostic and therapeutic implications of these findings. PMID- 15655566 TI - Home and office uroflowmetry for evaluation of LUTS from benign prostatic enlargement. AB - A group of 107 patients with lower urinary tract symptoms (LUTS) from benign prostatic enlargement (BPE) participated to the HOUSE Study (Home and Office Uroflowmetry Specific Evaluation). Patients received routine investigation, consisting of medical history taking, physical examination including digital rectal examination, prostate-specific antigen (PSA), assessment of symptoms listed both on the International Prostate Symptom Score and on ICS-male questionnaire. We examined the results of uroflowmetry evaluation in this population; data were analysed to observe if any circadian changes of parameters obtained with home uroflowmetry could be detected. We searched a correlation between Q(max), Q(ave) and ICS-benign prostatic hyperplasia symptom score: a significantly inverse correlation was found only for Q(max), confirming Q(max) as a reliable parameter to quantify subjective symptoms. When examining the multiple flow curves recorded in the same patient with home uroflowmetry, voided volume and flow time had usually higher values during night-time: the existence of circadian changes of uroflowmetry parameters in patients with LUTS from BPE was confirmed, and lower values of average and maximum flow rates during sleep hours were recorded in the same patient. In conclusion, when evaluating the natural history or treatment outcome of individual patients or group of patients in clinical trials for evaluation of BPE and LUTS, an assessment including multiple measurements may be useful and of value. PMID- 15655567 TI - Brachial diplegia as a result of cervical cord injury. AB - OBJECTIVE: To report a case of brachial diplegia with normal lower limb power, after a cervical cord injury. DESIGN: Case report of a 63-year-old man who sustained a neck trauma following a fall. SETTING: Department of Neurology, University Hospitals of Leicester, Royal Infirmary, Leicester, UK. CASE REPORT: The mechanism of the injury was probably one of hyperflexion resulting in central cervical cord contusion. Initial neurological examination revealed brachial diplegia, with posterior column sensory involvement of the lower limbs which had normal power. The patient was also in urinary retention but had no respiratory involvement. Magnetic resonance imaging (MRI) of the spinal cord showed a hyperintense signal in the central mid-cervical cord. CONCLUSION: Pure brachial diplegia represents a rare clinical presentation of cervical cord pathology, including trauma, of which clinicians should be aware. PMID- 15655568 TI - Recurrent vesical calculi, hypercalciuria, and biochemical evidence of increased bone resorption in an adult male with paraplegia due to spinal cord injury: is there a role for intermittent oral disodium etidronate therapy for prevention of calcium phosphate bladder stones? AB - STUDY DESIGN: Clinical case report with comments by colleagues from Sweden, Poland, Spain, Brazil, Japan, Belgium and Switzerland. OBJECTIVES: To discuss the role of disodium etidronate therapy for prevention of calcium phosphate vesical calculi in persons with spinal cord injury, who have hypercalciuria and biochemical evidence of increased bone resorption. SETTING: Regional Spinal Injuries Centre, Southport, UK. METHODS: A 21-year-old male sustained paraplegia (T-10; ASIA scale: A) in a road traffic accident in June 2001. He had an indwelling urethral catheter until the end of August 2001, when he started self catheterisation. He developed bladder stones and electrohydraulic lithotripsy (EHL) was performed in May 2002. All stone fragments were removed. Recurrence of vesical calculi was noted in October 2002. These stones were fragmented by lithoclast lithotripsy in two sessions, in December 2002 and February 2003; all stone fragments were removed at the end of the second session. This patient reverted to indwelling catheter drainage when vesical calculi recurred. In September 2003, X-ray of the abdomen showed recurrence of vesical calculi. By February 2004, the stones had increased in size and number. EHL of vesical calculi was again performed in April 2004. Complete clearance was achieved. RESULTS: A 24-h urinalysis detected hypercalciuria--18.7 mmol/day (reference range: 2.5-7.5). Biochemical analysis of vesical calculus revealed calcium phosphate (85%) and magnesium ammonium phosphate (15%). Plasma C-terminal telopeptide (CTX) was increased - 1.06 ng/ml (reference range: 0.1-0.5 ng/ml). Free deoxypyridinoline/creatinine ratio (fDPD/Cr) in urine was also increased - 20.2 (reference range: 2.3-5.4). In April 2004, this patient was prescribed disodium etidronate 400 mg day. Nearly 3 months after commencing therapy with etidronate, plasma CTX decreased to 0.87 ng/ml. fDPD/Cr in urine also decreased to 12.4. After 4 months of etidronate therapy, 24-h urinary calcium excretion had decreased to 6.1 mmol/day. CONCLUSION: Etidronate (400 mg daily) is a very effective inhibitor of calcium phosphate crystallisation. Etidronate decreased urinary excretion of calcium, an important factor in prevention of calcium phosphate bladder stones. Etidronate therapy is not a substitute for other well established methods for prevention of vesical calculi in spinal cord injury patients, for example, large fluid intake, avoiding long-term catheter drainage. Intermittent therapy with etidronate may be considered in selected patients, in whom hypercalciuria persists after instituting nonpharmacological therapy for an adequate period, for example, early mobilisation, weight-bearing exercises, and functional electrical stimulation. However, possible side effects of etidronate, and the fact that etidronate is not licensed in United Kingdom for prevention of urolithiasis, should be borne in mind. PMID- 15655569 TI - Fifth ventricle: an unusual cystic lesion of the conus medullaris. AB - STUDY DESIGN: Cystic dilatation of the fifth ventricle on its own is an extremely rare pathological event in adults whose pathogenesis is uncertain. The authors describe a personal case of 'fifth ventricle' and review the pertinent literature. OBJECTIVE: To emphasize the importance of including the fifth ventricle in differential diagnosis of lesions of the conus medullaris to ensure proper treatment. SETTING: Italy. METHODS: A 30-year-old woman was referred to us for recurrent low back pain. The patient was assessed by clinical, electrophysiological (motor evoked potential, somatosensorial evoked potential and electromyography of the perineal and lower limb muscles) and urodynamic investigations as well as a magnetic resonance imaging (MRI) of the lumbar-sacral segment with and without gadolinium enhancement, subsequently extended to the entire vertebral column and brain. Follow-up consisted of periodic clinical evaluation and lumbar-sacral MRI after 1 and 2 years. RESULTS: General physical examination, electrophysiological and urodynamic investigations were all negative, confirming the subjective nature of the patient's symptoms. Lumbar sacral MRI demonstrated the presence of a cyst lesion containing cerebrospinal fluid (CSF), which did not enhance after gadolinium, compatible with the diagnosis of the terminal ventricle. By extending the MRI investigation to the entire vertebral column and brain, it was possible to exclude an association with other malformations of the central nervous system. Clinical and radiological follow-up confirmed the nonevolutive nature of the lesion 1 and 2 years later. CONCLUSIONS: The lack of clinical symptoms and the stability of the radiological situation at 1 and 2 years follow-up motivated our choice of conservative treatment. PMID- 15655570 TI - Assessment of health status in children with spina bifida. AB - STUDY DESIGN: Prospective multidimensional study by means of: (1) clinical assessment, (2) parental-administered questionnaire for general health (CHQ PF50), and (3) standardised disability measurements. OBJECTIVES: To assess the health-related quality of life (QoL) and disability in children with spina bifida (SB) and to correlate them with the clinical picture and our previous study on adolescents with SB. SETTING: SB Centre at a University Hospital in Italy. METHODS: A total of 29 consecutive children with SB (mean age 11.4, range 4-14 years)were evaluated through Child Health Questionnaire Parental Form (CHQ-PF50), the FIM instrument, and the Barthel index. RESULTS: Disability was inversely related only (r=0.49; P=0.007) to the physical aspect of the QoL of children. Similarly, the disability was inversely related (r=0.37; P=0.005) to the emotional aspect of QoL of patient's parents. Unexpectedly, for the mental aspects of QoL of patients, major disability was not associated with higher psychological distress and severe role disability due to emotional problems. At clinical examination, findings especially for continence and number of catheterisations were usually related to deterioration of physical aspects of QoL (r=-2.28; P=0.02) in children. CONCLUSION: The multiperspective assessment showed that there is a linear inverse correlation between disability and QoL in children with SB only for physical aspects. Conversely, there is linear inverse correlation between disability and QoL in patient's parents regarding only emotional aspects. Moreover, this study provided useful information for clinical practice underlining that continence problems are those that most affect QoL in children with SB and their parents. PMID- 15655571 TI - [Status report on the chemotherapy of ovarian cancer at special cancer centers in Hungary (2002-2003)]. AB - Data on the first-line treatment of ovarian cancer in special centers of Hungary 2002 and 2003 are presented, involving 283 and 416 patients, respectively. Patients' age, clinical stage and histological type of the tumor were highly similar to literature data, while grades were different. Surgical effectiveness in case of IIIc staged tumors with >1 cm residual mass was 37%. The ratio of interval laparotomy was about 15%. Overall response rates of the first-line treatment of ovarian cancer was 82%, while the rate of complete remissions was 60%. The authors provide detailed analysis of factors that can improve the chemotherapy of ovarian cancer in Hungary. PMID- 15655572 TI - [Recent results of irinotecan therapy in colorectal cancer]. AB - New results presented at ASCO Conference in 2003 added further important data to our knowledge on successful use of irinotecan in colorectal cancer (CRC). Irinotecan - just like oxaliplatin - given as neoadjuvant therapy with 5-FU - folinic acid (FUFA) can render originally unresectable liver or lung metastases of CRC resectable, giving the hope of long-term survival for a proportion of patients. Irinotecan combined with 5-FU is an essential part of the most successful palliative sequential chemotherapy of stage IV CRC. Sequential FOLFIRI before or after FOLFOX combination ensures the longest possible progression-free and overall survival for metastatic CRC patients in the palliative setting. In order to achieve the longest survival time, sequential use of both 5-FU, irinotecan and oxaliplatin is necessary. The French GERCOR Group achieved 26 months median overall survival with the sequential use of continuous infusional FUFA, oxaliplatin and irinotecan combinations in stage IV CRC. The analysis of large phase III trials using 5-FU, irinotecan and oxaliplatin revealed that the higher proportion of patients was treated with all three drugs, the longer overall survival was achieved. If applied with caution, toxicity and efficacy of irinotecan in elderly patients is not significantly different from that seen in younger population. The anti-VEGF bevacizumab increases the efficacy of first line irinotecan therapy, while the addition of cetuximab restores irinotecan sensitivity in second line treatment of stage IV CRC. The combination of irinotecan with oral capecitabine is safe and effective in advanced CRC. PMID- 15655573 TI - [Experiences with Langerhans' cell histiocytosis in children in Hungary]. AB - BACKGROUND: Langerhans cell histiocytosis (LCH) in children is relatively rare, and the long-term analysis of therapy results has not been done yet in Hungary. PURPOSE: In this review we summarise the incidence, clinical features, prognostic risk factors and treatment results of children's LCH in Hungary, using data from the National Childhood Cancer Registry in Hungary in a 20-year period between 1981 and 2000. RESULTS: From January 1981 to December 2000, 111 children under 18 years of age were newly diagnosed with LCH in Hungary. The male-female ratio was 1.36:1, the mean age: 4 years 11 months. The minimal and median follow-up time was 3.48 years and 10.98 years respectively. 38 children had single-system disease, while in 73 cases we found systemic dissemination already at the time of diagnosis. Twenty-two patients were treated only by local surgery, 7 by surgery with local irradiation and 5 children received only local irradiation. In two cases remission was obtained with local steroid administration. 75 patient received chemotherapy. During the twenty years 14 children died, 9 due to the progression of the disease. Sixteen of the 111 patients had relapse with a mean of 2.16+/-1.29 years after the first diagnosis. Three patients with relapse got chemotherapy generally used in lymphoma and remission was achieved. The overall survival of all patients (n=111) was 88.3+/-3.1% at 5 years and 87.3+/-3.2% at 10 and 20 years. CONCLUSION: Childhood LCH is a well treatable disease and the survival rate is high. Even disseminated diseases have a quite good prognosis in childhood. PMID- 15655574 TI - [Teletherapy versus teletherapy and "boost" brachytherapy in the treatment of base of tongue tumors: 5-year results]. AB - AIM: To study the importance of high-dose-rate (HDR) boost brachytherapy (BT) after percutaneous irradiation of base of tongue tumors. METHODS: Between 1992 and 2000 seventy patients with biopsy proven carcinoma of the base of tongue were treated with primary radiation therapy. Fourty patients received a mean dose of 61 Gy (range, 50-72 Gy) external beam irradiation, and afterwards 30 patients were treated with a mean dose of 18 Gy (range, 12-30 Gy) boost HDR BT. Prognostic factors were analyzed in uni- and multivariate model. RESULTS: At a median follow up of 56 (16-108) months, boost BT increased the incidence of local tumor control (LTC) from 38% to 67% (p=0.0145). The 5-year probability of LTC was 60% vs. 36% (p=0.0188), the locoregional tumor control 52% vs. 34% (p=0.0753) and the overall survival (OS) 46% vs. 26% (p=0.0545), respectively, in favor of the boost group. Serious, grade 4 radiation toxicity occurred in 5% (2/40) and 13% (4/30) without or with boost treatment, respectively (p=0.2110). In multivariate analyses for LTC, tumor size (p=0.0042) and boost (p=0.0444), and for OS tumor size (p=0.0047) and nodal status (p=0.0163) had a significant effect. CONCLUSION: Boost BT after teletherapy improves LTC significantly without considerable increase in the risk of side-effects. PMID- 15655575 TI - [Radio-chemotherapy for non-resectable primary esophageal malignant melanoma]. AB - AIM: to demonstrate the simultaneous radio-chemotherapy of primary esophageal malignant melanoma on the basis of one case. PATIENT AND METHODS: 68-years-old male patient with malignant melanoma in middle part of the esophagus. The therapy was started with intraluminal high-dose-rate afterloading brachytherapy for the recanalisation of the esophagus (8 Gy in 0.5 cm deep), followed by percutaneous megavolt therapy two weeks after the last HDR AL session (50 Gy total dose, 5 x 2 Gy/week fractions for 5 week, 3D conformal planning). The chemotherapy was started simultaneously with the percutaneous megavolt irradiation (three courses of Cisplatin-5-Fluorouracil combination, repeated in four-week intervals). After the radio-chemotherapy a supraclavicular metastasis was verified, so the radio chemotherapy was followed with megavolt therapy of the metastasis at 30 Gy dose (5 x 2 Gy/week fractions), and chemotherapy (Cisplatin-Dacarbazine combination in 6-session, four-week intervals) and after them immunotherapy was started. RESULTS: The swallow function has been improved, the supraclavicular metastasis was in partial remission. After the beginning of the radio-chemotherapy the swallow function was good for 16 months, and 18 months after the beginning of radio-chemotherapy the patient died due to pulmonary and hepatic dissemination. CONCLUSION: Radio-chemotherapy of esophageal malignant melanoma has favorable palliative effect with acceptable quality of life. PMID- 15655576 TI - [Prognostic significance of cyclin D1, p27 and p63 expression in oral leukoplakia]. AB - Studies on the expression of genes regulating cell proliferation and apoptosis is of importance in relation to understanding the severity of the process and the possibility of malignant transformation. In the present study immunohistochemical demonstration of cyclin D1, p27 and p63 has been added to our previous investigations on Ki-67, p53 and apoptosis index. Clinical and pathological immunohistochemical studies on oral leukoplakias of 18 patients were performed. Clinically homogenous, non-homogenous or nodular, and erythroleukoplakia were distinguished. Pathologically the grading was made according to the degree of dysplasia. Immunoperoxidase reactions for cyclin D1, p27 and p63 were carried out, and the positivity was expressed in per cent, considering 1000 epithelial cells. Expression of cyclin D1 increased in parallel with the severity of leukoplakia. p27 expression was 14-16% in homogenous and nodular leukoplakias, whereas in erythroleukoplakia it decreased to 1-2%. p63 expression was 10% in average in homogenous, and 5% in nodular leukoplakias. While in erythroleukoplakias it increased to 20 per cent. The characteristic cyclin D1, p27 and p63 phenotype in various forms of leukoplakia may be considered as prognostic factors. PMID- 15655577 TI - [Dual tumours in the GI tract: synchronous and metachronous stromal (GIST) and epithelial/neuroendocrine neoplasms]. AB - Gastrointestinal stromal tumours (GIST) constitute the most frequent group of mesenchymal tumours in the gastrointestinal tract (GI). During the last several decades major advances have been taken in the diagnostics, treatment, and understanding of its pathogenesis. However, much less is known about the either metachronous or synchronous concurrence of GIST and other tumours of different histogenesis. In the present study clinicopathological data of 43 patients with histologically proved gastrointestinal stromal tumour were studied mainly in regard of the occurrence of a secondary neoplasm. Among the 43, 7 cases were found with secondary tumour mainly of epithelial origin. In three cases (cases 3, 5, and 7) GIST concurred with colorectal adenocarcinoma, in one case (case 1) GIST occurred in a patient with a 3-years-history of chronic lymphocytic leukaemia (CLL), in other two (cases 2 and 4) the stromal tumour was combined with in situ adenocarcinoma of the stomach and carcinoid of the pancreas, respectively. In case 6, GIST concurred with a duodenal Brunner gland adenoma. In five cases the stromal tumour and the other neoplasm occurred synchronously, and in four of them, being the stromal tumour clinically silent, GISTs were intraoperative findings. This confirms the importance of surgical intraabdominal control before closure. In one hand the repeated concurrence of GIST and colorectal adenocarcinoma in our series, and on the other hand, that of GIST and adenocarcinoma of the stomach in the literature, may indicate an at least partly common factor, which may be involved in the pathogenesis of these neoplasms. PMID- 15655578 TI - [Radiosensitizing effect of hyperfractionated radiation]. AB - To determine whether hyperfractionated treatment has benefits in the radiation therapy, two melanoma cell lines were irradiated with eight 0.5 Gy fractions as well as one single 4 Gy in vitro. The radiation was performed in air and in hypoxia as well. Cells were also irradiated in the presence of dibromodulcitol, a bifunctional alkylating agent with a weak radiosensitizer effect. The aim of the study was to examine whether hyperfractionation can influence the radiosensitizing effect of the bioreductive agent. Survival of the cells was determined immediately and 24 hours after various treatments by cell counting in hemocytometer and clonogenic assay. The number of the apoptotic cells was determined by the TUNEL assay and was followed up to 72 hours after treatment. Hypoxic cells had higher sensitivity than normoxic cells after 0.5 Gy irradiation. Radiosensitizing enhancement of DBD was higher with fractionated irradiation. The number of the apoptotic cells was significantly higher after hyperfractionated treatments than after single dose treatment combinations. Our results showed the significance of the hyperfractionated irradiation with 0.5 Gy per fraction in vitro. PMID- 15655579 TI - [Mevastatin induced apoptosis in U266 human myeloma cell line]. AB - Statins have been used successfully in the treatment of hypercholesteremia. Moreover, in vitro studies have shown that statins can trigger apoptosis in a variety of tumor cell lines. In the present study we analysed the effect of mevastatin -- a novel inhibitor of HMG-COA reductase, the rate-limiting enzyme of the mevalonate pathway -- on U266 human myeloma cells. Apoptosis induced by mevastatin was associated with increased caspase activity and depolarisation of mitochondrial membrane. Expression of BCL-2 mRNA and protein was down-regulated, with no change in BAX or BCLxL protein production. The mitochondrial program was supported by caspase-8 and cleaved BID activity. None of the antibodies neutralising death-ligand/death-receptor pathway -- TRAIL-R2Fc, anti-TNF-a, anti FASL (NOK-1) -- influenced the mevastatin-induced apoptosis. Mevastatin also stimulated shedding of syndecan-1 from the surface of myeloma cells. PMID- 15655580 TI - Mechano-sensitive orientation of cortical microtubules during gravitropism in azuki bean epicotyls. AB - The orientation of cortical microtubules (cMT) during gravitropism was studied in epidermal cells of azuki epicotyls. The relative proportion of cells with longitudinal cMT increased in the upper epidermis, and those with transverse cMT increased in the lower epidermis. When epicotyls were kept straight during gravistimulation, no change in cMT orientation occurred in either the upper and lower epidermis. When epicotyls were forced to bend downward, cells with transverse cMT increased in the upper epidermis, and those with longitudinal cMT increased in the lower epidermis. When epicotyls were loaded with naphthylphthalamic acid, an inhibitor of auxin transport, both gravitropic bending and change in cMT orientation were inhibited. However, when a change in cMT orientation was induced by forced downward bending, cells with longitudinal cMT increased in the compressed (lower) side and those with transverse cMT increased in the extended (upper) side. It was suggested that cMT orientation was controlled by the bending of the epicotyl and not by a gravity signal per se. Loading with Gd3+, an inhibitor of the stretch-activated channel, did not inhibit gravitropic bending. However, it inhibited cMT reorientation induced by gravitropic bending and by forced bending. Involvement of the stretch-activated channel in mechano-sensitive orientation of cMT was suggested. PMID- 15655581 TI - A novel microsatellite locus isolated from an AFLP fragment in the mangrove species Kandelia obovata (Rhizophoraceae). AB - The study of AFLP analysis in Kandelia obovata, one of the major mangrove species in Japan, revealed the existence of a unique fragment showing stuttered peaks. We cloned this fragment and found a novel microsatellite locus. We report the method used for isolation and the polymorphic nature of this locus among the populations on Iriomote Island. PMID- 15655582 TI - Psoriasis today--a new paradigm. PMID- 15655583 TI - Pathogenesis of psoriasis and current challenges. PMID- 15655584 TI - Incorporating biologics into the treatment of psoriasis. PMID- 15655585 TI - The use of alefacept in the treatment of psoriasis. PMID- 15655586 TI - Progress in the treatment of colorectal cancer. PMID- 15655588 TI - Abdominal wall metastasis following treatment of rectal cancer. AB - We present a 60-year-old man with a metastatic subcutaneous lump in the left lower quadrant of the abdomen, from rectal cancer, which was treated three years earlier with low anterior resection of the rectum. Excision of the abdominal wall metastasis was accomplished with negative histological margins, but six months later a new abdominal wall mass was detected. The patient underwent surgery again, in which the abdominal wall metastasis was resected en bloc with adherent portion of small bowel, along with inguinal lymph node dissection. The patient's condition deteriorated 10 months after the initial diagnosis, presenting again with abdominal wall cancer and dying from disseminated peritoneal disease. PMID- 15655587 TI - Screening for colorectal cancer. AB - A review of the literature regarding the screening strategies for colorectal cancer (CRC), particularly for average risk individuals, is analysed. The advantages and disadvantages or limitations of screening modalities for CRC, such as faecal occult blood testing (FOBT) with guaiac-based tests or the new faecal deoxyribonucleic acid tests, endoscopic screening by flexible sigmoidoscopy, colonoscopy, or CT-colonography and double contrast barium enema examination, are reported. The efficacy and cost of the screening tests are evaluated, and it is found that any of the suggested tests is more cost effective than other medical intervention or treatment as compared with no screening. The reported compliance to any form of screening test was 30-40%, a rate that is low enough. The experience of our surgical department of a screening programme, based on FOBT, on 4189 individuals over 50 years old, and application of colonoscopy in positive subjects, is reported. PMID- 15655589 TI - Effect of ranitidine on healing of normal and transfusion-suppressed experimental anastomoses. AB - BACKGROUND: Histamine has been shown to participate in immune response. Wound healing is a process of immune system. This experimental study was done to find the effect of histamine2 receptor antagonist ranitidine on the healing process of intestinal anastomosis in rats. METHODS: Eighty Wistar rats in four groups of 20 each underwent colon resection and anastomosis. They were given 2 ml saline or blood, twice daily 0.4 ml saline or 0.4 ml saline containing 0.7 mg ranitidine. The animals were killed 3 or 7 days postoperatively and the anastomotic strength assessed by bursting pressure. RESULTS: The ranitidine group developed fewer anastomotic abscesses (p<0.001). Anastomotic strength was significantly reduced either on day 3 or 7 in animals given blood transfusions (p<0.04, p<0.001), whereas in animals given ranitidine this effect was partially reversed. CONCLUSIONS: These data indicate that ranitidine has no influence in anastomotic bursting pressure, but has a lower incidence of septic complications. PMID- 15655590 TI - Large bowel obstruction in women with pelvic malignancies. What is the primary? AB - BACKGROUND: Large bowel obstruction is occasionally the mode of presentation of advanced colorectal or pelvic malignancies, and a prognostic of poor survival. The choices of treatment range from palliative tube decompression to curative or palliative surgery. METHODS: Twelve out of 500 women with various malignancies and symptomatology of large bowel obstruction were studied. All patients required urgent exploratory laparotomies. RESULTS: The primary site was established intra operatively to be the colon (2 cases), stomach (2 cases) and ovaries (2 cases). In the remaining 6 cases there was uncertainty about the origin of primary disease. Routine histology and immuno-histochemistry of the specimens revealed the origin of primary malignancy in all cases. CONCLUSIONS: Meticulous histological examination is essential for the establishment of correct diagnosis and selection of the best available treatment in women with pelvic malignancies, presenting with acute bowel obstruction. PMID- 15655591 TI - Colorectal cancer in patients over 70 years of age: determinants of outcome. AB - BACKGROUND: The aim of this study was to evaluate operative risk factors, the mortality, morbidity and survival in old patients with colorectal cancer. METHODS: From 1160 patients with colorectal cancer, 398 patients aged 70 years or older, from 1970 to 2000, were followed-up. Dukes' classification, differentiation, sex, anatomical site and survival were compared with patients <70 years old. RESULTS: Long-term results have been proved to be similar both in young and old patients. Relative survival rate for patients aged 70-95 (70.5%) were similar to those for patients less than 70 years old (71.6%) and also comparable between male (72.3%) and female (68%) patients. CONCLUSIONS: Elderly patients have a lower capacity to react to postoperative complications, but the relative survival is similar to younger patients. Advanced age alone should not be used as a criterion to deny surgery for colorectal cancer. PMID- 15655592 TI - Perforated diverticulum of the caecum. A difficult preoperative diagnosis. Report of 2 cases and review of the literature. AB - BACKGROUND: Perforation of a solitary caecal diverticulum is a rare cause of acute abdomen and an uncommon differential diagnosis for acute appendicitis. Nine hundred cases have been described since Potiers' first description of perforated caecal diverticulum in 1912. METHODS: We describe 2 cases of perforated diverticulum of the caecum. The first patient was a 50-year-old man diagnosed by subsequent histology, and the second a 77-year-old woman diagnosed intra operatively. Radiography, ultrasound and CT scan of the abdomen pointed at the diagnosis in the second case. A right hemicolectomy was performed. RESULTS: There were no complications, apart from a transient faecal fistula in the first patient managed conservatively. In both patients histology revealed a perforated caecal diverticulum. CONCLUSIONS: The surgeon must be familiar with the diagnosis and management of this rare, inflammatory benign caecal entity. PMID- 15655593 TI - Radio-frequency ablation of hepatic metastases from colorectal cancer. AB - BACKGROUND: The aim of our study is to present the preliminary results of an ongoing radio-frequency (RF) ablation study in patients with hepatic metastases from colorectal cancer. PATIENTS AND METHODS: From November 2003, two patients affected with metachronous liver metastases from colorectal cancer were treated with RF ablation. The mean age of the patients was 66 years (58 and 74 years). Tumours were unifocal right-lobe lesions in one patient and bifocal in the second patient. Under general anaesthesia, a Radionics 200-W RF generator was used to ablate lesions with H2O-cooled electrodes via laparotomy. Patients' follow-up ranged from two to five months including evaluation of salient clinical, radiological and laboratory parameters. RESULTS: The patients experienced moderate-to-severe pain in the right abdomen lasting for 2-3 days and mild fever for 3-6 days after treatment. During the follow-up period no local recurrence was observed. CONCLUSIONS: RF ablation emerges to be a promising method for the treatment of hepatic metastases from colorectal cancer. PMID- 15655594 TI - Prognostic significance of mucinous component in colorectal carcinoma. AB - BACKGROUND: Many factors influence survival in colorectal cancer patients, one of them is the mucinous component of the tumour. Mucinous adenocarcinoma is characterized by the extracellular mucin of more than 50% of the tumour volume. METHODS: From 1970 to 1999, 1160 patients were admitted to our clinic for colorectal cancer. They were divided into four groups according to mucinous character of the tumour, in two time periods of 15 years. RESULTS: There was an increase in the incidence of mucinous tumours from 20.8 to 30.5% in the second period. These tumours were more advanced (Dukes' C) and especially right sided (34.5% vs. 17.9%). Five-year survival was increased during the second period but was of a lesser degree in the mucinous group (51.5% vs. 65.5%). CONCLUSIONS: Colorectal mucinous carcinomas present at a more advanced stage, predominantly in men, with higher right colon location rate, and a worse overall 5-year survival rate than the non-mucinous colorectal cancers. PMID- 15655595 TI - Spontaneous rupture of the rectosigmoid with small bowel evisceration through the anus. AB - Spontaneous rupture of the rectosigmoid with small bowel evisceration through the anus is a rare and acute condition of the gastrointestinal tract. After Brodie's first description, only 52 such cases have been reported in the literature. An 83 year-old woman experienced a sudden, spontaneous rectosigmoid rupture and protrusion of small bowel loops, through the ruptured colon and out of the anus. After an immediate management, an emergency operation was performed, and a small bowel resection and suturing of the ruptured rectosigmoid were carried out for restoration of bowel continuity. On the 41st postoperative day, a massive cerebro vascular accident (CVA) was the cause of the patient's death. Discussion of the aetiology and treatment of this rare condition. PMID- 15655596 TI - Metachronous breast carcinoma (second malignancy), following "cure" from colorectal carcinoma. AB - Metachronous or synchronous breast carcinoma following or co-existing with colorectal carcinoma are well recognised clinicopathological entities, and the risk of developing both possibly underlines the similarities in carcinogenesis pathways for these carcinomas. We present a 60-year-old housewife with a history of a treated primary colon carcinoma (Duke's C) 15 years previously. Six months ago, during the follow-up care a small sub-areolar lesion was determined in a mammogram. A lumpectomy was performed under local anaesthesia, which revealed an infiltrating ductal carcinoma (6 mm in greatest diameter). Immuno-histochemical assays for oestrogen and progesterone receptors and c-erb B2 ongoprotein were performed. Axillary lymphadenectomy showed 1/11 positive node. She received adjuvant radiotherapy and hormone manipulation. To date, seven months later she is disease free. The aim of this report is to emphasise the risk of metachronous second malignancy of breast or colorectal carcinoma following colorectal carcinomas. A second primary colonic malignancy following breast primary carcinoma is more frequent than inverse clinical form. PMID- 15655597 TI - Locally recurrent rectal cancer after curative resection. AB - BACKGROUND: To determine the incidence of local recurrence, after curative resection for rectal cancer, with the application of total mesorectal excision (TME). PATIENTS AND METHODS: During the last ten years, 120 patients underwent curative resection for rectal cancer. As a rule, except for the cases that underwent high anterior resection, TME was applied. In terms of local relapse, routine TME, preoperative radiotherapy, tumour's stage, differentiation grade and number of positive nodes were taken into account. RESULTS: Eight patients (6.7%) presented with local relapse. At 5 years, 91.9% of patients were free of local recurrence and the actuarial disease-free survival was 81%. A significant association between routine TME, tumour's stage, differentiation grade, lymph node invasion and local recurrence was observed. Conversely, preoperative radiotherapy appeared to play no protective role. CONCLUSIONS: The curative resection of rectal cancer, with the application of TME, has led to a very low incidence of local relapse during the last few years. PMID- 15655598 TI - Metastatic breast adenocarcinoma masquerading as colonic primary. Report of two cases. AB - BACKGROUND: Common sites of metastasis for the breast cancer are bones, lungs, the central nervous system and liver. The colon is the rarest site of metastasis for the breast carcinoma. PATIENTS AND RESULTS: We report our recent experience of two female patients, 55 and 57 years old respectively, who presented breast metastasis at the proximal transverse colon. They were operated for breast carcinoma followed by chemotherapy and radiotherapy, four and ten years before the metastasis was diagnosed. The symptoms were anaemia and bowel obstruction. Diagnosis was made by double contrast barium enema and colonoscopy. Postoperatively, both patients received systemic chemotherapy. At the follow-up, two and three years after, there is no evidence of recurrence or metastasis. CONCLUSIONS: Patients with history of breast cancer presenting with anaemia and/or bowel obstruction should be examined for possible metastasis to colon and should be treated surgically followed by systemic chemotherapy. PMID- 15655599 TI - A case of mucinous adenocarcinoma arising in long-standing multiple perianal and presacral fistulas. AB - Chronic anal fistulas are not rare; however, the development of a carcinoma in long-standing, perianal fistulas is rare. We describe a case of an 85-year-old man with multiple, recurring, perianal fistulas, extending to the natal cleft. The patient underwent en bloc resection of the fistulas which were in direct continuity with the middle rectum. Histological examination revealed a mucinous colonic adenocarcinoma. Abdominal CT and colonoscopy revealed an extramural residual rectal mass. The patient refused a radical colorectal operation. Three years later, because of fistula recurrence, he underwent loop sigmoidostomy and radical en bloc excision of the perianal fistula and rectum, with immediate reconstruction by bilateral gluteal flaps. The patient was discharged on the 12th postoperative day, refusing adjuvant radiotherapy. We present this rare malignant entity, successfully treated by staged operations and without any adjuvant therapy. PMID- 15655600 TI - Accuracy of endorectal ultrasonography in the preoperative staging of rectal cancer. AB - BACKGROUND: Preoperative staging of rectal tumours is considered essential to tailoring treatment for individual patients. The aim of the present study was to evaluate the accuracy of 3-D-endorectal ultrasonography for the preoperative staging of rectal cancer. METHODS: Three hundred and fifty-seven patients with rectal adenocarcinomas underwent an endorectal ultrasonography evaluation during a period of eight years. The evaluation was performed by four surgeons. We compared the endorectal ultrasonography staging with the pathology findings. Patients with preoperative chemoradiation were excluded from the study. RESULTS: Overall accuracy in assessing the level of rectal wall invasion was 77.3%, with 9.3% of the tumours overstaged and 8.1% understaged. Accuracy in assessing nodal involvement in 313 patients treated with radical surgery was 74.9%, with 8.9% overstaged and 8.9% understaged. CONCLUSIONS: The accuracy of 3-D-endorectal ultrasonography in assessing the depth of tumour infiltration is good, but it is lower than previously reported. The technique is precise in distinguishing between benign tumours and invasive cancer. The results depend on the experience of the surgeon. PMID- 15655601 TI - Small bowel intussusception due to metastatic malignant melanoma. A case report. AB - Small bowel metastatic deposits attributed to malignant melanoma are found in 2 5% of patients with malignant melanoma of the skin. Ileo-ileo intussusception caused by metastatic melanoma is a very rare condition. The prognosis of metastatic melanoma is poor. We report a case of a cutaneous malignant melanoma which metastasised to the small bowel causing enteroenteric intussusception. This case refers to a 66-year-old male patient who underwent surgery for suspected enteric intussusception. This diagnosis was suggested by computer tomography scan. The patient had had previous surgery for a primary malignant melanoma in the eyelid of the right eye. Segmental intestinal resection with regional lymph node dissection and ileo-ileo anastomosis was performed. Metastatic melanoma in the gastrointestinal tract should be suspected in patients with history of melanoma of the skin and acute gastrointestinal symptoms. Immediate laparotomy and excision of the affected bowel segment is the appropriate treatment. PMID- 15655602 TI - Age-associated prognosis following curative resection for colorectal cancer. AB - BACKGROUND: The aim of this study is to determine the age-associated prognosis in patients who underwent curative resection for colorectal cancer. PATIENTS AND METHODS: Between 1993 and 1999, a total of 136 patients underwent curative surgical procedures for colorectal cancer. The study population was divided into three groups according to the age of the patients. Group A: patients younger than 45 years, group B: patients between 45 and 75 years and group C: patients older than 75 years. Tumour location, Dukes' staging, tumour differentiation and five year survival were evaluated. RESULTS: There was no significant association between age and tumour staging or tumour differentiation (p=0.990, p=0.753 and p=0.308, respectively). The overall survival at 5 years was 85.7% for the young patients, 77.5% for the middle-aged patients and 62.5% for the elderly patients. CONCLUSIONS: This aged-grouped study indicates that prognosis is comparable between younger and middle-aged patients whereas in elderly patients it is worsening but not statistically significantly. PMID- 15655603 TI - Are there any differences in colorectal cancer between young and elderly patients? AB - BACKGROUND: There is a controversy as to whether colorectal cancer (CRC) in the young population is more aggressive than in the elderly population, and hence has a less favourable prognosis. The aim of this study is to review the results of treatment of young and elderly patients diagnosed with CRC during a 10-year period (1994-2003). METHODS: We studied two distinct age groups: young (aged under 40) and elderly (aged over 80), who underwent surgery in the 1st Surgical Department. The young consisted of 11 patients with ages ranging from 37 to 40 (average age 39). The elderly consisted of 45 patients with ages ranging from 81 to 91 (average age 86). We compared clinical signs, pathological grade, Dukes' stage and presence of metastasis, average hospitalisation time, and postoperative mortality. RESULTS: The results of patients admitted to hospital with a diagnosis of acute abdomen were 9% in the young and 7% of the elderly. Those in stage C in Dukes' classification were 54.5% in the young group, and 44.4% in the elderly group. Those with undifferentiated adenocarcinoma were 36.3% in the young, and 8.8% in the elderly. During surgery, distal metastases were found in 18% of the young and 24.4% of the elderly. The average hospitalisation time of young and elderly patients was 13 and 15.2 days, respectively. The postoperative mortality rates of the young and elderly patients were 0% and 4%, respectively. CONCLUSIONS: Except for a higher rate of undifferentiated adenocarcinoma in young patients, there were no particular differences in the compared factors between the two age groups. The mortality rates do not indicate a less favourable prognosis for the young. PMID- 15655604 TI - Adjuvant chemotherapy in colorectal cancer. AB - Our aim was to study the three-fold response to treatment, survival rates and toxicity. MATERIALS AND METHODS: We review 76 patients (median age 66.2 years) affected with the disease. All received adjuvant chemotherapy, while 13 with rectal cancer were subjected to radiation too. Patients were staged after Duke's classification. RESULTS: Recurrence was noted in 15 patients, 3 at stage B and 12 at stage C; median recurrence-free interval was 18.9 months. Patients without relapse had a median disease-free period of 35.9 months. Median overall survival was 35.21 months. The 5-year survival mark have surpassed 14 patients (3 with recurrent disease): 8 at stage B and 6 at stage C. Ten have died; median survival in these patients was 30.5 months. No patients suffered from severe bone marrow toxicity; mucositis and diarrhoea grade III/IV was noted in 10 patients. PMID- 15655605 TI - Five-year survival after curative resection for adenocarcinoma of the colon. AB - BACKGROUND: The purpose of this study was to evaluate the 5-year survival of patients with colon adenocarcinoma that underwent elective or emergency curative surgical treatment. PATIENTS AND METHODS: Between 1993 and 1998, 80 patients underwent a potentially curative colonic resection based on mobilisation along anatomic planes. Among the patients, 26 underwent right colectomy, 3 transverse colectomy, 13 left colectomy and 38 sigmoidectomy. All patients classified as TNM stage III underwent adjuvant chemotherapy. The Kaplan-Meier method was used to analyse survival. RESULTS: Overall 5-year survival was 69.5%. Patient's sex and age, mucinous characteristics of the tumour and tumour location did not significantly affect survival. Patients with higher Duke's classification and TNM stage had significantly worse 5-year survival (p=0.025 and p=0.007, respectively). Although patients with good tumour differentiation had the highest 5-year survival, this difference was not statistically significant (p=0.211). CONCLUSIONS: The treatment of colon adenocarcinoma with curative resection by the end of the 20th century is accompanied with acceptable rates of overall 5-year survival. PMID- 15655606 TI - Colorectal gastrointestinal mesenchymal tumours. Report of a stromal case of the rectum (GIST) and a leiomyosarcoma of the transverse colon. AB - BACKGROUND: Gastrointestinal stromal tumours (GISTs) are the most common mesenchymal tumours of the gastrointestinal tract. They are positive to C-kit (CD 117), more common in the older males, and mostly in the stomach, less in the colon and rectum and oesophagus. Benign tumours are more common than the malignant ones. Classification of GISTs is based on morphology and immunochemistry. METHODS: We report two cases of colorectal gastrointestinal mesenchymal tumours, one on the transverse colon which was found to be immunohistochemically leiomyosarcoma and the other on the rectum which met the GIST criteria. The patients underwent transversectomy and abdominal perineal resection, respectively. CONCLUSIONS: They did not receive Imanitib postoperatively and two years after there is no evidence of recurrence. Surgery is the treatment of choice for resectable GISTs, and other mesenchymal tumours benign or malignant. PMID- 15655607 TI - Repair of parastomal hernia with the use of polypropylene mesh extraperitoneally. AB - Parastomal hernia is the most frequent complication of colostomy. Many surgical techniques have been postulated and prosthetic surgery seems to represents the first-choice treatment. The aim of this study is to report the surgical treatment of 4 patients that developed parastomal hernia, 3-10 months after abdominoperineal excision of the rectum and permanent sigmoidostomy due to carcinoma of the rectum. The repair was made with the use of polypropylene mesh extraperitoneally. One case of limited skin necrosis occurred without any serious consequences. No recurrence has been recorded among the patients, up to this day (follow-up period: 36 months). In conclusion, the suturing of fascial defect and the use of polypropylene mesh extraperitoneally is effective in the treatment of parastomal hernia. PMID- 15655608 TI - Magnetic resonance imaging in rectal cancer: the MERCURY experience. AB - Eleven European clinics, well experienced in a multidisciplinary approach to rectal cancer therapy, were testing magnetic resonance imaging (MRI) as a preoperative tool for staging of unselected rectal cancer patients. Aims and preliminary results of the study are presented. The way of using MRI for patient selection for neoadjuvant therapy is demonstrated from Berlin-Friedrichshain. PMID- 15655609 TI - Obesity and colorectal cancer. AB - BACKGROUND: To correlate obesity and colorectal cancer for Greek living conditions. PATIENTS AND METHODS: We studied 97 patients, who over the last 5 years were diagnosed histopathologically with colorectal cancer. 75.3% of the patients were either overweight or centrally obese; secondly, 21.6% patients had diabetes, percentages higher than those in the population (statistically significant). Hyperinsulinaemia and resistance to insulin have been implicated in colorectal carcinogenesis. CONCLUSIONS: As our sample was small, no statistically significant evidence correlating diet and/or physical activity to colorectal cancer has emerged. PMID- 15655610 TI - Carcinoid: modern aspects on its therapy. AB - In the past two years two patients with carcinoid tumours have been operated in Chios General Hospital. These two cases gave rise to our review of the literature concerning the therapy and follow-up of colorectal carcinoid tumours. We demonstrate the differences among surgeons concerning the acceptable and relevant indications for a hemicolectomy in appendiceal carcinoids and the prevailing aspects in the therapy of carcinoid of the colon and rectum. Finally we present the imaging techniques and the biochemical examinations that are proven of value and therefore should be used in the post-surgery follow-up. PMID- 15655611 TI - Survival after curative resection for rectal cancer by the end of the 20th century. AB - BACKGROUND: The aim of this study was to define the survival rates in patients with rectal carcinoma treated with curative resections. PATIENTS AND METHODS: Between 1993 and 1998, 54 patients with rectal cancer underwent curative resection by conventional technique. Tumour location, TNM staging and tumour differentiation were evaluated. Among the 54 patients, 14 underwent high anterior resection, 28 low anterior resection, 7 abdominoperineal resection and 5 underwent local excision. Survival rates were calculated using the Kaplan-Meier method and long-range analysis. RESULTS: Five-year survival was 70.4%. The survival rate statistically significantly decreased with increasing TNM tumour stage (p=0.009). Patients with poor differentiation of the tumour had the lowest 5-year survival (33%) compared to patients with moderate (72%) and good (78%) tumour differentiation. Sex and age did not affect survival. Location of the tumour in the distal end of the rectum and mucinous characteristics are poor prognostic factors affecting survival. CONCLUSIONS: Curative resection combined with chemoradiotherapy, whenever necessary, is accompanied with acceptable 5-year survival rates. PMID- 15655612 TI - Assessment of local stage in rectal cancer using endorectal ultrasonography (EUS). AB - BACKGROUND: Preoperative staging of rectal cancer is essential for the selection of the optimal treatment. This study aims to evaluate the accuracy of endorectal ultrasonography (EUS) in local staging of rectal cancer. PATIENTS AND METHODS: During a 4-year period, 33 patients with biopsy-proven rectal cancer underwent evaluation of the invasion of the rectal wall, the mesorectal lymph nodes status and the pelvic organs using EUS. We compared the EUS findings (uTN) to the histopathology examination of the resected specimens (pTN) according to TNM classification. RESULTS: Most patients had T3 tumours. Overall accuracy in assessing the depth of rectal wall invasion (T) and the lymph node status was 79% and 59% respectively. Two patients previously treated by preoperative chemoradiation were correctly staged only for N stage. CONCLUSIONS: EUS is a valuable diagnostic tool in local staging of rectal cancer. Progressively increasing experience will overcome the obstacles in accurate interpretation of ultrasound images. PMID- 15655613 TI - Current treatment of colorectal liver metastases. AB - BACKGROUND: The aim of our study is to present our experience in the treatment of liver metastases in patients with colorectal cancer. PATIENTS AND METHODS: Between 1997 and 2003 a total of 12 patients with liver metastases from a primary colorectal cancer were treated in our department. They were 8 males and 4 females with a median age of 64 years (range 56-70 years). RESULTS: Ten patients underwent liver resection. The surgical procedures were 4 major hepatectomies (3 right hepatectomies, 1 left lobectomy) and 9 wedge liver resections. In total, 16 metastatic lesions were resected. Already at the time of the primary tumour, 5 patients presented with a synchronous liver metastasis. In 3 of them, liver metastasis was resected together with the primary tumour, and in the rest, resection was performed 1 month after the initial operation. In 5 patients liver metastases were metachronous and were diagnosed 3-14 months after the initial operation. The median survival of the patients was 39 months. Two patients (one with 2 metastatic lesions) underwent radiofrequency ablation (RFA) of the metachronous metastatic lesions and remain well 3-6 months postoperatively. CONCLUSIONS: Hepatectomy is the treatment of choice for hepatic metastasis of colorectal cancer, whenever feasible. Recent promising treatments such as RFA can further improve the outcome of these patients. PMID- 15655614 TI - Synchronous colorectal cancer. AB - Synchronous carcinomas of the colon and rectum are of considerable clinical significance because of their frequency, the number of extra tumours missed and the difficulty of preoperative diagnosis. A retrospective evaluation of 283 patients with primary colorectal adenocarcinomas was performed. There were 6 patients with 12 synchronous adenocarcinomas (2.12%). Colonoscopy and double contrast barium enema revealed the synchronous cancer in 66.6% of the cases. In two cases the second cancer was found intraoperatively. In one patient an urgent laparotomy was performed because of acute abdomen caused by perforation of the ascending colon. Typical colectomies, depending upon the segment of the location of the lesion, were performed. Second cancers had a significantly more favourable stage than index colorectal adenocarcinomas. The index and the secondary cancers of synchronous colorectal adenocarcinomas showed a better histologic grade (well differentiated type) than the single cancers. Full clinical and radiological investigation is essential, before any operation is undertaken for colorectal cancer. PMID- 15655615 TI - Effects of steroid on the healing of colonic anastomoses in the rat. AB - BACKGROUND: The aim of this experimental study was to evaluate the effects of steroid on colonic anastomosis in a rat model. METHODS: Forty female Wistar rats were randomised into two groups. After resection of a 1-cm segment of the transverse colon, an end-to-end sutured anastomosis was performed. In the rats of the control group 2 ml of solution 0.9% NaCl was injected once daily intraperitoneally perioperatively. Rats of the steroid group received intraperitoneally once daily, and on the same perioperative days, hydrocortisone (5 mg/kg body weight in 2 ml solution NaCl). All the rats were sacrificed on the 8th postoperative day and the anastomoses were examined macroscopically. The bursting pressure measurements were recorded and anastomoses were graded histologically. RESULTS: Dehiscence rate was significantly higher in the steroid group than in the control group (p<0.001). Mean bursting pressure was significantly lower in the steroid group compared to the control group (p<0.001). Colonic healing process assessed as inflammatory cell infiltration and collagen deposition was significantly lower in the steroid group than in the control group (p<0.001, p=0.03 respectively). CONCLUSIONS: Perioperative steroid treatment adversely affects healing of colonic anastomoses in the rat. PMID- 15655616 TI - Application of radionics on primary and metastatic tumours of the liver. AB - The aim of our study is to present our initial experience on the use of radiofrequency ablation (RFA) on hepatic tumours. Four patients with tumours of the liver were submitted to RFA. The first patient underwent an emergency left lobectomy due to rupture of a tumour of the left hepatic lobe. A synchronous tumour was detected on the right lobe. He has been submitted to four sessions of RFA. The second patient was submitted to transverse colon resection (adenocarcinoma) and a year later three metastatic tumours were detected. Two sessions of RFA have been applied in this case. The third patient, with adenocarcinoma of the sigmoid and one metastatic lesion on the right hepatic lobe, and the fourth patient with breast cancer and metastatic lesion of the liver, underwent one session of RFA each. The use of RFA on metastatic or primary tumours of the liver enriches the possibilities of therapeutic treatment. PMID- 15655617 TI - Clinical short-term results of radiofrequency ablation in patients with liver metastases from colorectal cancer. AB - BACKGROUND: The aim of this study was to examine the therapeutic efficacy of radiofrequency tumour ablation in patients with liver metastases from colorectal cancer who are not suitable for hepatic resection. PATIENTS AND METHODS: Between April 2002 and January 2004, 18 patients underwent open radiofrequency ablation (RFA) for colorectal liver metastases. Median lesion size was 5.6 cm (range 3.0 8.0 cm). Patient's follow-up included ultrasound and computed tomography imaging at 3, 6 and 18 months postoperatively. RESULTS: Mean total procedure time was 86 min. The average hospital stay was 5 days. There was no treatment-related mortality. Twelve patients (66.7%) experienced mild right hypochondrium discomfort for 3 days and 6 patients (33.4%) low-grade fever for 4 days. Four patients died within 12 months with extrahepatic disease. In 4 patients lesions increased in size or new lesions developed, 7 patients are alive, symptom-free, with stable disease and 4 patients are free of disease. CONCLUSIONS: RFA is a safe, well tolerated procedure for the treatment of unresectable colorectal liver metastases. PMID- 15655618 TI - Priority of resection in concomitant abdominal aortic aneurysm (AAA) and colorectal cancer (CRC): review of the literature and experience of our clinic. AB - The concomitant occurrence of abdominal aortic aneurysm (AAA) and colorectal cancer (CRC), although rare, always represents a therapeutic dilemma. The incidence of coexistence ranges between 0.49 and 2.1%. Both lesions should be treated to achieve best life expectancy. But the main controversy revolves around whether to treat them simultaneously or as staged procedures. In our institution, we treated seven cases of concomitant AAA and CRC. In five of them, synchronous conventional resection was preferred. In the latest two, which we present, endovascular aortic repair was chosen. No graft infection was documented. PMID- 15655619 TI - Postoperative follow-up of patients with colorectal cancer: a combined evaluation of CT scan, colonoscopy and tumour markers. AB - BACKGROUND: The purpose of this study is to present the experience of our department regarding the importance of the systematic postoperative follow-up of patients with colorectal cancer, early diagnosis and treatment of the recurrence of the disease or a metachronous cancer. METHODS: In a study that took place between October 2001 and February 2004 amongst 67 patients that were operated upon for colorectal cancer, 41 were systematically being followed up through CT scan, colonoscopy and tumour markers. RESULTS: In the 14th and 18th months postoperatively for 2 of the patients the CT scan showed hepatic metastasis, while the colonoscopy was negative. Regarding the tumour markers, one (CEA) was elevated in one patient while three were elevated in the others. For 2 out of the 41 patients the colonoscopy showed recurrence of the disease within one and two years respectively. The CT scan proved to be free of metastasis and the tumour markers were falling within the normal range for one of the patients while for the other CEA was elevated. Both patients underwent additional colectomy. Postoperative increase of the tumour markers was observed in 9 patients. The above patients had normal markers in the immediate postoperative period. For 4 out of the 9 patients recurrence or spread of the disease was observed while the rest of them are still being followed up. CONCLUSIONS: In conclusion, we believe that the systematic postoperative follow-up of the patients with colorectal cancer through CT, colonoscopy and the use of tumour markers contributes decisively to the early diagnosis and treatment of any possible recurrence of the cancer or a metachronous cancer or misdiagnosed concomitant cancer. PMID- 15655620 TI - Mass screening for colorectal cancer: compliance in Almopea Region. AB - Colorectal cancer (CRC) is the second leading cause of cancer-related morbidity and mortality in Europe and the United States. Planning for a CRC screening began in co-operation with local authorities (Pella Prefecture sponsored test kits). Our aims were to develop a screening programme for colorectal cancer using the faecal occult blood test (FOBT) in Almopea province, and to investigate the compliance of local farmers population. Cancer statistics data from Almopea have been analysed and they showed higher colorectal cancer incidence compared to the rest of Greece. We designed a one-time FOBT screening programme on the Surgery Department computer Network, in which we listed 8963 subjects, over 50 years of age. From them, 4189 underwent 3 days FOBT, and the rest were our control group. The method of successive visits to each community by the medical team and educational meeting was chosen. For allocation and gathering of tests, teams of volunteers have been organised. In case of positive FOBT (176 subjects), total colonoscopy was performed. Seventeen (17) polypoids (in 15 patients) and 20 cases of diverticulosis were detected. The compliance of FOBT group was 49% (from 4189). Colonoscopy accepted 89% from 176 patients with positive test. We concluded that our study shows poor compliance of screening population. There is a need for co-operation of medical services, local authorities, media and volunteers support organising. PMID- 15655621 TI - Stage of disease at time of attendance of patients with colorectal cancer at the Department of Surgery of the General Hospital of Edessa. AB - It is acceptable that the stage of disease at time of attendance of patients with colorectal cancer (CRC) is the defining factor for patients' survival rate. From 1986 until 1998, 417 patients were treated in the Department of Surgery of the General Hospital of Edessa with CRC. For diagnosis, established endoscopic procedures were followed. CT, USG, MRI and histology were performed for staging. 85.4% of the patients had stage II, III and IV TNM/UICC tumour at the time of attendance. PMID- 15655622 TI - Treatment of complicated colorectal cancer. Evaluation of the outcome. AB - A comparison of the outcome of 66 patients with complicated colorectal cancer (CRC) who underwent an emergency operation and 217 electively operated patients in an 11-year period was retrospectively determined. The complications presented on admission were obstruction (13.4%), perforation with peritonitis (6.36%), obstruction and perforation (1.4%) and massive bleeding (2.1%). The majority of the patients electively treated were of stages II and III (45.5% and 29%) and those urgently operated on were stages III and IV (67.7% and 14.1%) respectively. Radical resections were performed in 45 patients and palliative in 21. Only ten patients from the urgently treated are still alive and free of the disease. In conclusion, the treatment of complicated CRC carries a considerable incidence of complications, morbidity and mortality, and the overall 5-year survival rate is disappointing compared to electively treated patients. PMID- 15655623 TI - Metachronous colorectal adenocarcinomas. AB - BACKGROUND: The purpose of our study is to emphasise the diagnostic and therapeutic problems of metachronous colorectal cancer. MATERIALS AND METHODS: Between 1990 and February 2004, amongst 185 patients that were treated for colorectal cancer, in four of them a metachronous carcinoma was diagnosed. RESULTS: 1st patient: male 41 years, underwent colectomy of the descending colon for adenocarcinoma. Four years later, a rectosigmoidal cancer was found infiltrating urinary bladder. 2nd patient: male 62 years, underwent right hemicolectomy. Eight years later two synchronous cancers were diagnosed, in the left colic flexure and in the sigmoid colon. 3rd patient: female 73 years, underwent low anterior resection for rectal cancer. Eight years later, caecal and ascending colon cancers were diagnosed with hepatic metastases. 4th patient: female 60 years underwent transversectomy. Six years later caecal cancer was diagnosed with pulmonary metastases. Amongst the four patients, only the fourth had an adequate postoperative follow up. CONCLUSIONS: Extended radical colectomies in young patients and in those where adenomatous polyps coexist will reduce the incidence of metachronous carcinoma. Effective and persistent postoperative surveillance in patients with colorectal cancer will greatly contribute in the detection and treatment of metachronous carcinomas. PMID- 15655624 TI - Manometric and clinical evaluation of patients after low anterior resection for rectal cancer. AB - The aim of this study was to examine the anorectal physiological and clinical changes that occur after low anterior resection for rectal cancer. Since 1998, 30 patients underwent laboratory tests of anorectal function, preoperatively and 1 month and 6 months after low anterior resection. Postoperatively all patients presented with increased bowel frequency, 60% of the patients with mild soiling and 30% with urgency for defecation. Six months after surgery there was a significant improvement of these symptoms. The anal resting pressure was significantly decreased postoperatively, while maximum squeezing pressure remained unchanged. The rectoanal inhibitory reflex was absent in 80% of the patients and at 6 months after surgery it tended to recover. Rectal capacity and compliance were reduced in all patients. In the current study, the majority of patients demonstrated manometric anorectal changes and clinical anorectal function disorders during the first year after surgery. We observed that these disorders correlated with the low level of the anastomosis. PMID- 15655625 TI - Colorectal carcinoma: contribution of growth pattern on prognosis. AB - BACKGROUND: Recent studies have reported poor prognosis in colorectal carcinomas with non-polypoid growth pattern. The aim of the present study is to investigate this issue in the patients referred to "Hatzikostas" General Hospital of Messolonghi, Greece. MATERIALS AND METHODS: A total of 13 patients who were treated for primary colorectal adenocarcinoma in a one-year period were studied. Colorectal carcinomas were classified into two types based on the presence or absence of elevation of tumour as compared with adjacent mucosa: polypoid growth (PG-type) and non-polypoid growth (NPG-type) carcinomas. Clinical and tumour histopathologic parameters were analysed. RESULTS: Unlike PG-type carcinomas, NPG type carcinomas had a high proportion of poorly differentiated adenocarcinoma cells, a high likelihood of being advanced, and a high frequency of lymph node as well as distant metastases at the time of diagnosis. CONCLUSIONS: Colorectal carcinomas with NPG pattern tend to show more malignant characteristics than those with PG pattern, explaining their poorer outcome. PMID- 15655627 TI - Oophorectomy during surgery for colorectal carcinoma. AB - BACKGROUND: The purpose of the study is to identify the effect of synchronous prophylactic oophorectomy in women undergoing colorectal cancer surgery on long term survival, recurrences and sites of failure. PATIENTS AND METHODS: From 1987 to 2003, 124 women, mean age 69+/-10 (35-91) years, with colorectal carcinoma were retrospectively reviewed. In 70 (56.5%) women the ovaries were preserved during surgery and 54 (43.5%) women underwent synchronous prophylactic oophorectomy during primary tumour resection. Univariate and multivariate analysis were used to assess the effect of oophorectomy on long-term survival, recurrences and sites of failure. RESULTS: By univariate analysis it was demonstrated that synchronous oophorectomy had no effect on long-term survival (p=0.7294). By multivariate analysis it was demonstrated that stage was the only factor independently influencing survival (p=0.0061). Twenty-eight patients (23%) developed recurrence and 10 of them developed locoregional recurrence. By univariate analysis it was demonstrated that the number of recurrences was not different between women with or without oophorectomy (p=0.259). Distant and locoregional recurrences were not different between women undergoing resection of primary colorectal carcinoma with or without oophorectomy (p=0.611). CONCLUSIONS: Oophorectomy does not appear to influence long-term survival, the total number of recurrences or the sites of failure. PMID- 15655626 TI - The presence and significance of iron in neoplasia of the colorectum. AB - BACKGROUND: To investigate the presence and distribution of tissue iron in colon adenomas of all grades of dysplasia as well as colorectal carcinomas of all grades of differentiation. MATERIALS AND METHODS: Formalin-fixed, paraffin embedded tissue blocks from 8 biopsies of colon adenomas and 13 surgical specimens of colorectal carcinoma were studied. Consecutive sections were stained using Perls' stain that demonstrates the presence of ferric iron by staining it blue. RESULTS: Iron was absent in all cases of colon adenoma. On the other hand, iron was present in 9 of the 13 cases of colorectal carcinoma (69.23%), regardless of the degree of differentiation, the location and the stage of development. The iron was stored in mesenchymal cells in the stroma and various types of inflammatory cells. CONCLUSIONS: Our results suggest association of iron presence in the environment of malignant cells. Thus, iron may be a carcinogenic factor, possibly facilitating tumour growth and modulating local immune response. PMID- 15655628 TI - Total mesorectal excision with the water-jet-dissection. Technique and results. AB - BACKGROUND: The introduction of total mesorectal excision (TME), in particular, has resulted in a dramatic decrease in postoperative functional disorders. Strict adherence to the dissection planes described above, is the prerequisite for good oncological and functional outcome. To facilitate dissection, we have introduced the high-pressure water jet dissector into our surgical routine for the treatment of rectal cancer. Below, we describe the technique of total mesorectal excision we employ in our department and present our perioperative surgical results. METHODS: We performed total mesorectal excision for rectal cancer on 49 patients, comprising 32 males and 17 females aged between 34 and 88 years. 38 patients underwent low anterior resection of the rectum and 11 patients abdominoperineal resection. RESULTS: The mean operating time was 282 min, mean intraoperative administration of red cell concentrates was 0.55 units. The mean postoperative stay in the ICU was 1.92 days. Anastomotic leakage occurred in 6.1%, wound healing disturbances in 12.2%, urinary tract infection in 12.2% intra-abdominal infection in 4.1% and faecal fistula in 4.1%. Urinary tract infections occurred in 12.2%. Postoperative temporary neurogenic voiding disturbances occurred in 14.3%. Persistent neurogenic bladder dysfunction after three months could be observed in 6.1%. CONCLUSIONS: The perioperative results are comparable to those of centres of excellence in the international literature. The Hydro-Jet dissector significantly facilitated TME. The particular feature of this technical aid is that it permits the rapid early development of the embryological plane between the pelvic nerves and the mesorectal fascia, without doing damage to either of them. This leads to optimal radicality and 1 maximum preservation of the autonomic nerves. Before a definitive pronouncement on voiding disturbances can be made, however, we consider it necessary to carry out a prospective randomised study with preoperative and postoperative urodynamic investigations. PMID- 15655629 TI - Results of laparoscopic treatment of rectal cancer: analysis of 520 patients. AB - BACKGROUND: To evaluate short- and long-term outcomes of laparoscopic resection of rectal carcinoma in curative intent. METHODS: Within a subgroup analysis of a multicentre study, initiated by the "Laparoscopic Colorectal Surgery Study Group (LCSSG)", 520 patients with rectal cancer were included in a prospective, clinical observational study. RESULTS: One hundred and ninety patients underwent an abdominoperineal resection (APR) and 330 patients an anterior resection (AR) of the rectum. The demographic parameters, intraoperative complication rates, morbidity and mortality rates were comparable in both groups. In patients undergoing AR a trend towards limited oncologic radicality, and a significant increase in the anastomotic leak rate related to the distance of the tumour from the anal verge were observed. Calculated survival data revealed stage-related survival rates for UICC stages I, II and III of 82.0%, 68.8% and 63.3%, respectively. CONCLUSIONS: While APR is highly suited to the laparoscopic approach, laparoscopic AR cannot at present be generally recommended. PMID- 15655630 TI - Emergency operation in carcinomas of the left colon: value of Hartmann's procedure. AB - BACKGROUND: Colonic resection according to the procedure by Hartmann is considered a fast and safe surgical intervention, which has been used for years, in particular, in emergency situations. METHODS: Using data of a prospective multicentre study on the operative treatment of colorectal carcinoma over the time period from 1 January 2000 to 31 December 2002, the value of Hartmann's procedure was investigated in carcinoma of the left colon (n=8825) compared with alternative surgical options under emergency circumstances. The significant impact of independent variables on the type of the selected approach was determined by means of logistic regression. RESULTS: While in total 422 primary Hartmann's procedures (4.8%) were executed under curative intention, 213 (50.5%) of those were carried out in emergency situations. Hartmann's procedure was beneficial in cases with tumour-associated obstruction and perforation of the left colon as it resulted in the lowest mortality (7.5%) of the radical operations. Even under palliative intention, Hartmann's procedure was preferred at the left colon but led to a postoperative mortality (32.7%) very similar to that in creation of a colostoma (33.3%) or segmental colonic resection (38.9%). CONCLUSIONS: Hartmann's procedure has been widely accepted as a curative intervention in emergency cases (oncosurgically adequate R0 resection) for the carcinoma of the left colon. Because of the high postoperative morbidity and mortality in emergency situations with only palliative options (R0 resection not possible), alternative endoscopic treatment should be considered more frequently. PMID- 15655631 TI - Perineal wound closure after abdomino-perineal excision of the rectum. AB - Wound infections have a significant impact on the postoperative morbidity after abdomino-perineal rectum resection (APR). However, the technique of perineal wound closure after APR has not been standardised yet. The prospective German multicentre trial "Colorectal Carcinoma (primary tumor)" (study I) enrolled 10 335 patients with rectal cancer over a time period from 1 January 2000 to 31 December 2002. The APR rate was 24.7% (n=2517). Four hundred and forty-five patients (17.6%) developed an infectious complication of the perineal incision. In contrast, the data of the reporting single centre participating in study I were as follows: patients with rectal cancer, n=206; APR, n=37; APR rate, n=18.0% (study II). In our surgical department, a standardised technique of perineal wound closure comprising tight sutures of the tissue in three layers (muscle, ischiorectal and subcutaneous fat), local administration of carriers releasing antibiotics and a redon drainage were used. Using these tools, the rate of septic perineal wound complications was considerably lower with 5.4% vs. 17.6% in the multicentre trial. PMID- 15655632 TI - Emergency operations for carcinoma of the colon. AB - The aim of this retrospective clinical study was to record and analyse the immediate and long term results obtained from emergency operation for carcinoma of the large bowel. In a 12-year period (1991-2003), 154 patients were operated on for carcinoma of the colon. In 29 patients (19%; 11 males, 18 females, m.a. 72.8 years and 67.5 years respectively), the operation was performed urgently. In 21 (72.5%) there was a complete obstruction located in 17 (81%) at the sigmoid, in 2 (9.5%) in right flexure and in 2 (9.5%) in left flexure. Five patients (17%) had developed peritonitis due to perforation proximal to the tumour and three patients (10.5%) had developed necrotic colitis and haemorrhage due to the obstruction. In 23 patients (79.5%) a radical curative operation was feasible [11 (38%) subtotal colectomies with ileo-rectal anastomosis, seven (24%) Hartman's, three (10%) right hemicolectomy and two (7%) on table bowel lavage plus colectomy with ileo-rectal anastomosis] and in six (11%) only palliative operations could be done (transverse loop colostomy). In all cases the postoperative period was uneventful. Three patients with palliative operation were submitted to a radical curative operation 2 months later and the other three patients died 6 months later from causes irrelevant to the primary disease. It is concluded that in this series of patients radical curative operations for carcinoma of the large bowel even under urgent conditions were feasible in the cases without additional complications, eliminating the necessity for reoperations later on. PMID- 15655633 TI - Complications in laparoscopic colorectal surgery: results of a multicentre trial. AB - BACKGROUND: We present a prospective observational multicentre study of 4834 consecutive cases undergoing laparoscopic colorectal procedures. METHODS: The study was initiated on 1 August 1995, with centres from Germany, Austria and Switzerland participating. All patients undergoing laparoscopic colorectal surgery were included, and no selection criteria were applied. RESULTS: Between August 1995 and April 2004, the participating centres treated 4834 patients. Three thousand, one hundred and sixty-six operations were performed for benign indications, and 1668 for cancer, with most operations being done for diverticulitis (2019, 41.8%). The conversion rate was 5.6%, and the overall morbidity was 20.1%. Intra-operative complications occurred in 5.9%, postoperative complications in 14.0%, and the overall anastomotic leak rate was 3.1% (colon 2.6%, rectum 11.9%). The 30-day mortality rate was 1.3%. CONCLUSIONS: It has been our experience that laparoscopic surgery for colorectal disease is safe. Morbidity and mortality are comparable to those of conventional colorectal surgery. PMID- 15655634 TI - Colon cancer: resection standards. AB - The surgeon is one of the most important prognostic factors for patients with colon cancer. Depending on the site of the primary tumour, the potential lymphatic spread is uni-, bi- or tridirectional. Therefore, the extent of surgical resection depends on the site of the tumour and the patient's vascular anatomy. A minimum resection of 10 cm of grossly normal bowel on both sides of the tumour is required to keep the risk of unremoved paracolic lymph node metastasis below 5%. A central ligature of main colic artery (arteries) is strongly recommended, as central lymph nodes are involved in more than 10%. The no-touch isolation technique is easily and quickly performed and has been shown to provide a survival benefit of 6% compared to the conventional technique. Adherent or infiltrated adjacent organs should never be separated from the tumour because the tumour perforation and consecutive tumour cell spillage is afflicted with a reduction in 5-year survival expectancy of up to 40 %. Prophylactic oophorectomy seems to be of no survival benefit, only infiltrated or grossly abnormal ovaries have to be removed. Sentinel lymph node biopsy facilitates an accurate nodal staging and may result in an up-staging in 15-30% and in necessitating adjuvant chemotherapy. Recently published data indicate no oncosurgical disadvantage of laparoscopic colon cancer resection compared to open technique. PMID- 15655635 TI - Ancient Greek medicine as the foundation of contemporary medicine. PMID- 15655636 TI - The surgical approach to locally recurrent rectal cancer. AB - The incidence of rectal cancer recurrence after surgery is 5-45%. Predictive factors relating to local recurrence (LR) are the surgical experience, the location of the tumour, the tumour's pathology, the circumferential margins, the successful total mesorectal excision (TME) and the use of radiochemotherapy. The use of TME and preoperative radiotherapy reduce the risk of LR. Diagnosis is done using imaging techniques. Computed tomography with rectal contrast is the preferred imaging technique. The surgical strategy for LR at and around a previous anastomosis is the resection of the anastomosis with a sphincter-saving procedure. LR isolated to one side after anterior resection is an indication for en bloc resection of the rectum with a portion of the bladder. LR fixed to the sacrum requires a composite abdominosacral resection. Extrapelvic disease is a contraindication to curative resection. The composite sacropelvic resection is a reasonable alternative to palliative radiation with long-term survival of 20-30%. PMID- 15655637 TI - Five fractions of preoperative radiotherapy for selected cases of rectal cancer. AB - In locally advanced colorectal cancer, combined modality treatment is a standard adjuvant therapy in USA and Europe. The basic issue of the timing of radiotherapy and chemotherapy and preoperative radio-chemotherapy versus postoperative radio chemotherapy is being addressed in prospective trials. Preoperative radiotherapy proved that causes more radiosensitivity due to that oxygen tension within the tumour may be higher prior to surgery. Technically, there are two approaches of preoperative radiotherapy. The aim of this article is to report the inclusion criteria, indications, advantages and results of short course preoperative radiotherapy. PMID- 15655638 TI - The management of peritoneal surface malignancy of colorectal cancer origin. AB - Colorectal carcinomas are predominantly spread malignancies. Peritoneal carcinomatosis frequently associates colorectal carcinomas. The tumour grade, the completeness of cytoreduction, the tumour volume, the presence of distant metastases, prior surgery score and the extent of peritoneal implantations are prognostic clinical features of survival. The management of colorectal cancer with peritoneal carcinomatosis is possible by resection of tumour, cytoreduction and intraperitoneal chemotherapy. T3 and T4 colorectal tumours are at risk of developing locoregional recurrence and may be treated by intraperitoneal chemotherapy. Early postoperative intraperitoneal chemotherapy has been used in 40 patients with T3 and T4 tumours, with 15% hospital mortality, and 32.5% morbidity. The overall 3-year survival rate was over 80% and only 15% distant metastases were recorded. PMID- 15655639 TI - Chemotherapy in metastatic colorectal cancer. AB - The treatment of patients with metastatic colorectal cancer (MCRC) has changed dramatically over the years. 5-Fluorouracil (5FU)-based therapies have been routinely included in treatment regimens for colorectal cancer for the past 40 years. A number of options are available to clinicians for the treatment of patients relapsing after surgical excision of their primary tumour, such as 5FU in association with FA, the new drugs such as irinotecan and oxaliplatin and the oral fluoropyrimidines: capecitabine and uracil/tegafur (UFT). It has been shown that combination therapy with 5FU/FA and irinotecan or oxaliplatin is more active than 5FU/FA in the first line of treatment in MCRC. New agents acting on novel targets are under development such as epidermal growth factor inhibitors, vascular endothelial growth factor inhibitors and cyclooxygenase 2 inhibitors. PMID- 15655640 TI - Hepatic resection for metastasis from colorectal cancer. AB - BACKGROUND: Liver metastases from colorectal cancer are treatable and potentially curable when hepatic resection is applied. This paper is to illustrate surgical treatment of metastatic colorectal cancer in China. PATIENTS AND METHODS: Between January 1993 and December 2002, 485 patients with colorectal liver metastases from 6 institutes in China were reviewed. Among them were 340 males and 145 females with average ages of 55.6 years (23-81). Surgical intervention includes primary colorectal cancer resection, hepatic resection, hepatic arterial chemoembolisation and portal vein catheterisation, and systemic chemotherapy. RESULTS: Among 485 cases, data were not complete in 76, and their 3-year survival rate was 35.7%, while from 409 patients who underwent surgical intervention, 11 cases only underwent colorectal cancer resection (group A); 89 with hepatic resection (group B); 204 with hepatic arterial intervention or portal vein catheterisation chemotherapy (group C); 21 with regional ablation by radiofrequency or microwave thermal coagulation (group D); and 84 with systemic chemotherapy (group E). The cumulative 3- and 5-year survival rates were 0% in groups A and E, 43.5% and 32.1% in group B, 27.1% and 0% in group C, and 42.9% and 19.2% in group D. CONCLUSIONS: Surgery can offer long-term survival and resection should be considered when liver metastases can be totally resected with clear margins and when there is no nonresectable extrahepatic disease. The choice between anatomical or wedge resection depends on the number and the location of the metastases. Special multifunctional operative device limit blood loss and increase resectability. PMID- 15655641 TI - Terminology in coloproctology. AB - On the matter of the terminology used for large intestine, its sections and different pathologies, a general observation would be that there is a quantity of different sections, words and interchanges that in the end cause some distress in the medical community. The correct terminology assists in the understanding between doctors, some terms, though are standard and unchangeable. Some of those terms are somewhat false but can always be used in a better way. When the relevant knowledge exists, then cooperation between doctors is easier and could help in avoiding mistakes in the future. PMID- 15655642 TI - Anti-VEGF antibody bevacizumab (Avastin) with 5FU/LV as third line treatment for colorectal cancer. AB - BACKGROUND: To evaluate the activity and safety of bevacizumab when given with standard 5FU/leukovorin (LV) regimens in patients with metastatic colorectal cancer who have failed irinotecan and oxaliplatin-based treatments. METHODS: Bevacizumab was given at 5 mg/kg as an IV infusion every 2 weeks. Patients received 5FU according to Roswell Park or the de Gramont regimen. RESULTS: Nineteen patients enrolled, median age 60, median PS: 1. Most common toxicity attributable to bevacizumab was mild hypertension, epistaxis and mild proteinuria; 1 patient had a CNS haemorrhage. The median number of cycles was 1 (8 weeks). Clinical benefit as disease stabilisation lasting 2-6 months was noted in 9 patients, whereas 10 progressed (median f/u: 5 months). TTP was 16 weeks, and the overall survival has not been reached (24+ weeks). CONCLUSIONS: Bevacizumab may result in growth arrest and clinical benefit in a substantial proportion of patients with colorectal cancer and no alternative treatment. PMID- 15655643 TI - The failed intraperitoneal colon anastomosis after colon resection. AB - BACKGROUND: The purpose of this study is to present the incidence of anastomotic leakage after colon resection and intraperitoneal anastomosis for colorectal cancer. PATIENTS AND METHODS: In the last ten years, 205 patients underwent colonic resection with intraperitoneal anastomosis for colon cancer. The surgical management of colorectal cancer consisted of 66 right hemicolectomies, 3 transverse colectomies, 17 left hemicolectomies, 98 sigmoid colectomies and 21 high anterior resections of the rectum. Diagnosis of leakage was made by clinical features, blood vessel examinations and abdominal CT-scans. RESULTS: Anastomotic leakage occurred in 5 out of 205 patients (2.4%). One of these patients underwent emergency surgery and the other 4 elective surgery; 3 by manual, 2 by mechanical suture. Three patients with anastomotic leakage were reoperated on days 4, 5 and 7, and 2 patients were treated conservatively. Two of the patients (20%) with anastomotic leakage died due to sepsis. CONCLUSIONS: Even though the rate of anastomotic leakage in patients with intraperitoneal anastomosis after colon resection for colorectal cancer is low, it remains a significant complication and a major cause of postoperative morbidity and mortality. PMID- 15655644 TI - Serum levels of bcl-2 in patients with colorectal cancer. AB - BACKGROUND: Many recent studies focus on the immunohistochemical evaluation of Bcl-2 expression, and its prognostic significance in colorectal cancer (CRC). Our aim was to investigate the presence of bcl-2 protein in the serum and to examine the association between its levels, stage and tumour load, in patients with CRC. METHODS: A commercially available ELISA was used for the estimation of bcl-2 levels, in 94 patients with different stages of CRC. Forty-eight healthy blood donors served as controls. Concentrations ranging 2SD above and below the controls median were accepted as "normal". RESULTS: bcl-2 was detected in the serum of patients with CRC. A significantly higher proportion of patients with non-metastatic disease (61%), had high serum bcl-2 values, compared to patients with metastatic disease (28%, p<0.0001). CONCLUSIONS: Serum bcl-2 in patients with CRC may reflect the degree of Bcl-2 expression in cancer tissue. Serum bcl-2 is easily determinable, and could be useful as a prognostic marker in CRC. PMID- 15655645 TI - The effect of aspirin and high fibre diet on colorectal carcinoma: a comparative experimental study. AB - BACKGROUND: The aim of this study is to assess the role of high fibre diet and aspirin on dimethylhydrazine (DMH)-induced colorectal cancer in rats. MATERIALS AND METHODS: Colorectal tumours were induced with DMH. The animals were randomly divided into five groups (15 rats each): I, controls; II, rats receiving only the carcinogen; III, rats receiving the carcinogen and high fibre diet; IVA, rats receiving the carcinogen plus low dose aspirin; IVB, rats receiving the carcinogen plus high dose aspirin. RESULTS: Adenocarcinomas were detected in 100% of the rats in group II, 47% of the rats in group III (chi2, p<0.05), 100% of the rats in group IVA, but the incidence was reduced to 50% in the rats of group IVB (p<0.05). CONCLUSIONS: These data indicate that high fibre diet and aspirin suppress experimental colon carcinogenesis and the protective effect of aspirin is dose related. PMID- 15655646 TI - Anaemia as a symptom of right colon cancer. AB - BACKGROUND: The aim of our study is to determine the proportion of iron deficiency anaemia in patients with right colon cancer at diagnosis and to remind of the need of investigation of the large bowel in patients presenting with anaemia. PATIENTS AND METHODS: From 1988 to 2003, 86 patients with right colon cancer underwent operative management. RESULTS: Seventy-five patients (87.2%) with right colon cancer had anaemia at diagnosis. The mean Ht value was 33.00% (ranging from 16 to 47%). CONCLUSIONS: Iron deficiency anaemia is a common symptom of right colon cancer. During the evaluation of patients with iron deficiency anaemia, examination of the right colon is needed. PMID- 15655647 TI - Serum concentrations of soluble ICAM-1 and VCAM-1 in patients with colorectal cancer. Clinical implications. AB - BACKGROUND: In colorectal cancer (CRC), serum levels of adhesion molecules ICAM-1 and VCAM-1 increase with progressing disease. Our aim was to investigate the effect of treatment in serum ICAM-1 and VCAM-1. METHODS: Serum levels of ICAM-1 and VCAM-1 were determined in 154 patients with CRC. Patients with non-metastatic disease had been treated with "curative" resections, and those with advanced disease were receiving chemotherapy. A group of 48 healthy blood donors served as control. Serum concentrations ranging 2SD above and below the controls median were accepted as "normal". RESULTS: Significantly more patients with advanced disease demonstrated high serum ICAM-1 and VCAM-1 (p<0.001, p=0.043 respectively). However, in some of them serum ICAM-1 and VCAM-1 were below "normal". CONCLUSIONS: Serum ICAM-1 and VCAM-1 may be valuable markers of residual or biologically aggressive disease after curative resections. In advanced disease, they could be useful predictors of response to chemotherapy. PMID- 15655648 TI - Management of colorectal cancer: 20 years' experience. AB - Colorectal cancer is the second most common cause of cancer death in westernised countries. Its treatment is the subject of active, ongoing investigation within all treatment modalities, including surgery, chemotherapy and radiotherapy. We reviewed the charts of all patients treated in this university general surgical department for colorectal carcinoma, between January 1983 and December 2002. A total number of 494 patients with colorectal cancer were operated on, on an elective or emergency basis. Patients' demographics, clinical presentation, location of tumour, type of surgery, operative staging, pathology of tumour, outcome and follow-up were recorded and analysed. PMID- 15655649 TI - Environmental and other risk factors for colorectal carcinogenesis. AB - Diet plays a role in the risk of colon cancer, but exactly how it affects risk is unknown. In western countries diet is low in fibre and high in animal protein, fats and refined carbohydrates such as sugar. Risk seems to be reduced by a diet high in calcium, vitamin D and vegetables such as Brussels sprouts, cabbage and broccoli. Familial polyposis has been studied sufficiently and increases the risk of colon cancer. Moreover, in relatives there is frequently a familial history of an increased incidence of colorectal cancer. There is no conclusive evidence to support any of the hypotheses proposed to explain the role of dietary factors in colorectal carcinogenesis. Prospective trials should generate the information required to develop strategies for diet modification to reduce the incidence of colorectal carcinoma. There is hope that this understanding will lead to a variety of dietary, medical, hormonal and molecular interventions to reduce disease incidence and improve the prognosis of patients with cancers of the colon and rectum. PMID- 15655650 TI - Synchronous polyps in patients with colorectal cancer. AB - BACKGROUND: The aim of this study is to underscore the incidence of synchronous polyps in patients with colorectal cancer and to emphasise the importance of their perioperative detection and management. PATIENTS AND METHODS: Three hundred and seven patients underwent a potentially curative resection for colorectal cancer during the last ten years. A total of 129 synchronous polyps were detected in 72 of the patients (23.5%). Complete preoperative colonoscopy was performed in 62 of the patients. Forty-three polyps (33.4%) in 37 patients were removed preoperatively, while 69 polyps (53.4%) in 25 patients were included in the surgical specimen. In 10 patients the colon was evaluated postoperatively and 17 polyps (13.1%) were removed via endoscopy. RESULTS: A total of 81 polyps were detected in different surgical segments than the index cancer. Furthermore, 15 polyps were detected in the right colon of 55 patients with left colon cancer. CONCLUSIONS: Synchronous polyps in patients with colorectal cancer are a frequent event. Thus, all patients should undergo a perioperative colonoscopy and endoscopic polypectomy, if feasible. The planned surgical procedure may alter as a consequence of the colonoscopic findings in some of the patients. PMID- 15655651 TI - Electrocoagulation: an alternative palliative treatment for rectal cancer. AB - BACKGROUND: We present our experience with palliative transanal electrocoagulation therapy (TEC) for rectal cancer. METHODS: Eight patients with biopsy-proven localised low rectal adenocarcinoma were treated with palliative TEC. Inclusion criteria were: high risk patients with anal adenocarcinoma less than 3 cm in diameter, localised less than 7 cm from the anal verge, limited to the rectal wall. Under local anaesthesia all patients underwent TEC using the traditional cautery. RESULTS: No mortality and morbidity was found. Four patients required a second procedure and one patient had a third session . Two patients died within 2 years from distal metastasis. The remaining six patients are alive and free of local recurrence (follow-up 9 months to 4 years). CONCLUSIONS: In poor surgical candidates, palliative TEC of rectal adenocarcinoma may have a role as an alternative to radical surgical treatment. PMID- 15655652 TI - Anastomotic leakage following anterior resection for rectal cancer. AB - BACKGROUND: The aim of this study is to present the incidence of anastomotic leakage after anterior resection for rectal cancer and to demonstrate the therapeutic approach for the treatment of this complication. PATIENTS AND METHODS: During the last ten years, 93 patients underwent anterior resection of the rectum for rectal cancer. Low anterior resection with total mesorectal excision (TME) was performed in 72, and high anterior resection in 21 patients. The definition of the anastomotic leakage was based on clinical features, peripheral blood investigations and abdominal CT scan. RESULTS: Clinically apparent anastomotic leakage developed in 9 patients (9.7%). Four patients were managed conservatively and five operatively. Postoperative mortality among the patients with anastomotic leakage was not recorded. CONCLUSIONS: The incidence of anastomotic leakage after anterior resection of the rectum for rectal cancer is relatively low. It remains however the most serious complication following rectal resection for cancer. PMID- 15655653 TI - Surgical management of large bowel obstruction due to colonic cancer. AB - BACKGROUND: Bowel obstruction represents a common surgical emergency. The purpose of this study is to highlight our principles while managing cases of large bowel obstruction (LBO) due to colonic carcinoma. METHODS: Twenty-one patients with LBO underwent emergency surgery. Patients with LBO caused by obstructive malignant colonic lesions underwent either with one-stage primary resection and anastomosis (14 patients, 67%) or two-stage operation (7 patients, 33%). RESULTS: There were no operation-related complications. The average length of hospitalisation was 14 days with a range from 10 to 23 days. Postoperative mortality was 14%. CONCLUSIONS: One-stage primary resection and anastomosis of the large bowel, is a feasible option in cases of emergency. LBO caused by colonic carcinoma can be performed with acceptable morbidity and mortality whenever comorbidity of the patient is not a contraindication. PMID- 15655654 TI - Pelvic reconstruction of the retroperitoneum after abdominoperineal resection of the rectum using full-thickness skin grafts. AB - BACKGROUND: To find a safe and effective method of pelvic reconstruction after abdominoperineal resection in order to prevent small intestine from descending into the pelvis. This allows safe delivery of optimal doses of radiation therapy, which exceed radiation tolerance of the small intestine, in advanced stages of colorectal cancer. MATERIALS: Prospective, ongoing study examining patients who underwent abdominoperineal resection and pelvic reconstruction with full thickness skin grafts. METHODS: Nine (9) patients underwent abdominoperineal resection of the rectum and pelvic reconstruction with full-thickness skin grafts for colorectal cancer stage C. Subsequently they were referred for adjuvant radiation therapy and followed up regularly for surgical complications and disease recurrence. RESULTS: All patients successfully completed postoperative radiation therapy and there were no serious surgical complications pertaining to the initial operation and the skin homeotransplantation. None of the patients needed reoperation, so the long-term outcome of the transplantation was not surgically evaluated. CONCLUSIONS: Reconstruction of the peritoneal gap after abdominoperineal resection of the rectum with full-thickness skin graft is a safe and effective method. The small intestine was effectively excluded from the pelvis allowing successful completion of adjuvant radiation therapy with minimal irradiation of the small intestine and effective prevention of radiation enteritis. PMID- 15655655 TI - Is TNM classification related to early postoperative morbidity and mortality after colorectal cancer resections? AB - BACKGROUND: To examine the role of TNM staging system as a predictive factor for postoperative morbidity and mortality, after colorectal cancer resections. PATIENTS AND METHODS: During the last ten years, 368 patients with colorectal cancer were referred to our institution. All patients, who underwent primary treatment elsewhere or defunctioning colostomy only, or who did not undergo surgical therapy were excluded from the analysis. The early postoperative outcomes registration of the remaining 351 patients (197 men, median age 66.2 years) was retrospectively linked to TNM stage. RESULTS: TNM stage had a poor prognostic value for the early postoperative morbidity rate. In addition, according to the statistical analysis, the proportion of early postoperative mortality proved to be higher in patients with TNM stage III or IV colorectal cancer. CONCLUSIONS: TNM classification could be considered as a reliable predictor of early postoperative mortality, but has no role in the prediction of early postoperative morbidity after colorectal resections. PMID- 15655656 TI - Use of intraluminal stapler device for creation of a permanent colostomy. AB - We present our experience with the use of the intraluminal stapler device for the purpose of creating of a dermal permanent colostomy, in cases of acute emergency operations and of regular programmed operations. The advantages of this method for those surgeons who use stapling devices are: controlled safety of the colostomy, shorter operation time and the creation of a stable diameter of the colostomy. All patients of this category as well as those having been operated on for colon cancer have been placed on a protocol follow up for three years and are closely monitored. This has allowed us to closely follow these patients in cases of complications. PMID- 15655657 TI - Synchronous and metachronous colorectal carcinoma. AB - BACKGROUND: Synchronous and metachronous colorectal carcinoma have an incidence of 2-10%. The purpose of the study was to evaluate the clinical characteristics, the accuracy of diagnostic examinations and the survival of these patients. METHODS: From 1970 to 1999, 1160 patients with colorectal cancer were admitted to our Department. During follow-up examination 50 patients (4.3%) were found to present with multiple primary colon cancers. Fifty-two per cent were synchronous and 48% metachronous tumours. RESULTS: The overall 5-year survival of the patients was 45.87%. Mortality was 10% for multiple primaries, while in patients with single cancer was 4.1%. The overall 5-year survival of the patients with multiple primaries tumours was 46.67%. CONCLUSIONS: Patients with colorectal cancer must be fully studied endoscopically. There has been an improvement in survival in recent years due to better surgical techniques, the introduction of more sophisticated examination methods and the meticulous follow-up of patients at risk. PMID- 15655658 TI - [Orthopedic treatment of paraplegics]. PMID- 15655659 TI - Co-existence of Chiari malformation type I and Epstein-Barr virus meningoencephalitis in a 3-year-old child: case report and review of the literature. AB - In the present report we describe an unusual case of a 3-year-old girl who was admitted to our hospital with Epstein-Barr virus meningoencephalitis. Brain magnetic resonance imaging revealed diffuse abnormalities in white matter and Chiari I malformation with cervical and thoracic hydro-syringomyelia. PMID- 15655661 TI - Intracranial hypotension due to leakage of cerebrospinal fluid: could myelography be a therapeutic option? AB - Spontaneous intracranial hypotension (SIH) is an uncommon condition caused by cerebrospinal fluid leakage. We report of a 29-year-old woman with typical symptoms in whom initial cranial MRI and CT were normal . A clinical diagnosis of SIH, but the symptoms did not resolve with conservative therapy or a lumbar epidural blood patch. Repeat MRI revealed (indirect) findings consistent with SIH but failed to directly demonstrate a cerebrospinal fluid fistula. Subsequent CT myelography revealed leakage of contrast medium into the epidural space bilaterally along the nerve roots at C3-7. Following the myelography the patient reported dramatic clinical improvement with complete resolution of symptoms. The adhesive quality of the myelographic contrast medium might have precipitated closure of the meningeal tears. Myelography may be not only of diagnostic value but also therapeutic in SIH. PMID- 15655660 TI - Chemical shift magnetic resonance spectroscopy of cingulate grey matter in patients with minimal hepatic encephalopathy. AB - Minimal hepatic encephalopathy (MHE) is frequently diagnosed in patients with liver cirrhosis who do not show overt clinical cirrhosis-associated neurological deficits. This condition manifests primarily with visuo-motor and attention deficits. We studied the association between visuo-motor deficits and magnetic resonance spectroscopic parameters in cingulate grey matter and white matter of centrum semiovale in patients with liver cirrhosis. The data revealed an increase in the glutamate-glutamine/creatine ratio and a decrease in choline/creatine and inositol/creatine ratios in patients with liver cirrhosis. The analysis of the data showed that cirrhosis-associated deterioration of the visuo-motor function significantly correlates with a decrease in the choline/creatine ratio and an increase in N-acetylaspartate/choline in cingulate grey matter but not in the neighbouring white matter. Furthermore, the increase in the glutamate glutamine/creatine ratio correlated significantly with the increase in the N acetylaspartate/creatine ratio. These data suggest an association between altered choline, glutamate-glutamine and NAA metabolism in cingulate grey matter and symptoms of MHE, and underline the importance of differentiation between grey and white matter in magnetic resonance spectroscopic studies on patients with cirrhosis-associated brain dysfunction. PMID- 15655662 TI - Dynamics of skeleton formation in the Lake Baikal sponge Lubomirskia baicalensis. Part I. Biological and biochemical studies. AB - In ancient Lake Baikal (East Siberia), freshwater sponges have diversified to an extraordinary degree. The skeleton of Lubomirskia baicalensis, which attains a size of up to 1 m, is constructed from spicules, which are cemented into longitudinal bundles. Our X-ray analysis revealed that the architecture of the specimens follows a highly ordered radiate accretive growth pattern. The spicules have a central axial canal with an axial filament inside. This organic filament is composed of silicatein, the major enzyme involved in silica formation of the spicules. We found that the specific activity of silicatein in samples from the non-growing (basal) zone is much lower than in those from the growth zone (tips) and that even the composition of this molecule differs in these regions. The present study shows for the first time that the turnover of silicatein, the major element of the axial canal of sponge spicules, changes within a sponge specimen depending on the region in which it is found. PMID- 15655664 TI - Impact of Cre1, Cre8 and Cre3 genes on cereal cyst nematode resistance in wheat. AB - The cereal cyst nematode (CCN; Heterodera avenae), a root disease of cereal crops, is a major economic constraint in many wheat (Triticum aestivum)-growing areas of the world. The objective of this study was to assess the impact of the Cre1, Cre8 and Cre3 genes on CCN resistance. A population of 92 doubled-haploid (DH) lines derived from a cross between wheat cvs. Frame and Silverstar as well as 1,851 wheat breeding lines were screened for CCN resistance at the Primary Industries Research Victoria (PIRVic). A second population of 9,470 wheat breeding lines was screened at the South Australian Research and Development Institute (SARDI). Cre3 had the largest impact on reducing the number of female cysts, followed by Cre1 and Cre8. There was no significant difference in number of cysts between DH lines with or without the Cre8 marker, suggesting that the marker is not perfectly linked to Cre8. The estimated heritabilities were 0.32 in the DH population, 0.48 in the PIRVic data set and 0.32 in the SARDI data set, which confirm that this is a trait of low heritability. The repeatability of CCN resistance improved with an increase in the number of plants assessed per line-up to ten. However, 85-88% of the improvement was achieved with the assessments of the first five plants. PMID- 15655663 TI - Bowman-Birk inhibitors in Lens: identification and characterization of two paralogous gene classes in cultivated lentil and wild relatives. AB - In order to investigate the genetic structure of lentil Bowman-Birk inhibitors (BBIs), primers were designed on pea BBI sequences. The sequences obtained from lentil DNA, using these primers, indicate that lentil possesses at least two paralogous genes. Protein sequences translated in silico from lentil DNA sequences suggest that the two coded proteins are highly similar to Pisum trypsin inhibitor TI1 and TI6 BBIs, respectively. In fact, both are double-headed inhibitors, one class showing the presence of a trypsin- and a chymotrypsin reactive site, the other showing two trypsin-inhibition sites, similar to pea TI1 and TI6, respectively. The same primers were used to amplify sequences from the DNA of other Lens species. The results strongly support that all species of Lens possess the same classes of BBI coding genes, orthologous to those identified in the cultivated lentil. Lens nigricans showed the most diverging sequences both at the nucleotide and the amino acid level. The similarity of the two gene classes identified in the genus Lens to those of Pisum and the observations that the patterns of expression of the Lens genes are equivalent to those of pea orthologous genes, possibly imply that BBIs in Lens are coded by gene classes with similar genome organization and function to those of pea. Finally, a phyletic analysis, based on the comparison of sequences obtained from other species belonging to the Vicieae tribe of the Fabaceae family, strongly suggests that all Vicieae could have a similar genome organization and function for BBI genes, and that this could be a general rule in all the Fabaceae family. PMID- 15655665 TI - Statistical tests for QTL and QTL-by-environment effects in segregating populations derived from line crosses. AB - Quantitative trait locus (QTL) studies in plants frequently employ phenotypic data on a population of lines (doubled haploid lines, recombinant inbred lines, etc.) tested in multiple environments. An important feature of such data is the genetic correlation among observations on the same genotype in different environments. Detection of QTL-by-environment interaction requires tests which take this correlation into account. In this article, a comparison was made of the properties of several such tests by means of simulation. The results indicate that a split-plot analysis of variance (ANOVA), being an approximate method, tends to be too liberal under departures from the Huynh-Feldt condition. A standard two-way ANOVA, which ignores genetic correlation, yields inappropriate tests and should be avoided. In contrast, mixed model approaches as well as univariate and multivariate repeated-measures ANOVA yield valid results. This supports the use of a flexible mixed model framework in more complex settings, which are difficult to tackle by repeated-measures ANOVA. . PMID- 15655667 TI - Domestication patterns in common bean (Phaseolus vulgaris L.) and the origin of the Mesoamerican and Andean cultivated races. AB - Chloroplast DNA polymorphisms were studied by PCR sequencing and PCR-restriction fragment length polymorphism in 165 accessions of domesticated landraces of common bean from Latin America and the USA, 23 accessions of weedy beans, and 134 accessions of wild beans covering the entire geographic range of wild Phaseolus vulgaris. Fourteen chloroplast haplotypes were identified in wild beans, only five of which occur also in domesticated beans. The chloroplast data agree with those obtained from analyses based on morphology and isozymes and with other DNA polymorphisms in supporting independent domestications of common bean in Mesoamerica and the Andean region and in demonstrating a founder effect associated with domestication in each region. Andean landraces have been classified into three different racial groups, but all share the same chloroplast haplotype. This suggests that common bean was domesticated once only in South America and that the races diverged post-domestication. The haplotype found in Andean domesticated beans is confined to the southern part of the range of wild beans, so Andean beans were probably domesticated somewhere within this area. Mesoamerican landraces have been classified into four racial groups. Our limited samples of Races Jalisco and Guatemala differ from the more widespread and commercially important Races Mesoamerica and Durango in types and/or frequencies of haplotypes. All four Mesoamerican races share their haplotypes with local wild beans in parts of their ranges. Independent domestications of at least some of the races in Mesoamerica and/or conversion of some locally adapted wild beans to cultigens by hybridization with introduced domesticated beans, followed by introgression of the "domestication syndrome" seem the most plausible explanations of the chloroplast and other molecular data. PMID- 15655666 TI - Development and mapping of microsatellite (SSR) markers in wheat. AB - Microsatellite DNA markers are consistently found to be more informative than other classes of markers in hexaploid wheat. The objectives of this research were to develop new primers flanking wheat microsatellites and to position the associated loci on the wheat genome map by genetic linkage mapping in the ITMI W7984 x Opata85 recombinant inbred line (RIL) population and/or by physical mapping with cytogenetic stocks. We observed that the efficiency of marker development could be increased in wheat by creating libraries from sheared rather than enzyme-digested DNA fragments for microsatellite screening, by focusing on microsatellites with the [ATT/TAA]n motif, and by adding an untemplated G-C clamp to the 5'-end of primers. A total of 540 microsatellite-flanking primer pairs were developed, tested, and annotated from random genomic libraries. Primer pairs and associated loci were assigned identifiers prefixed with BARC (the acronym for the USDA-ARS Beltsville Agricultural Research Center) or Xbarc, respectively. A subset of 315 primer sets was used to map 347 loci. One hundred and twenty-five loci were localized by physical mapping alone. Of the 222 loci mapped with the ITMI population, 126 were also physically mapped. Considering all mapped loci, 126, 125, and 96 mapped to the A, B, and D genomes, respectively. Twenty-three of the new loci were positioned in gaps larger than 10 cM in the map based on pre existing markers, and 14 mapped to the ends of chromosomes. The length of the linkage map was extended by 80.7 cM. Map positions were consistent for 111 of the 126 loci positioned by both genetic and physical mapping. The majority of the 15 discrepancies between genetic and physical mapping involved chromosome group 5. PMID- 15655668 TI - Evolutionary conservation and characterization of the bare lymphocyte syndrome transcription factor RFX-B and its paralogue ANKRA2. AB - The extraordinary homology between major histocompatibility complex class II (MHC II) proteins across species from human to bony fish suggests that transcription factors that regulate these proteins might be conserved as well. Deficiencies in four proteins that regulate MHC II genes in humans (RFX-B, RFX5, RFXAP, and CIITA) cause an inherited immunodeficiency disorder known as the bare lymphocyte syndrome (BLS). To understand the structure and mechanism of function of the BLS transcription factors, we analyzed the evolutionary history of RFX-B, the factor deficient in the majority of patients with BLS. Sequence comparison and analysis of the RFX-B proteins showed that RFX-B and a closely related protein, ANKRA2, are present in humans to bony fish and that specific domains are highly conserved. In addition to sequence conservation, functional conservation exists, as mouse and Xenopus RFX-B orthologues, but not the paralogous protein ANKRA2, were able to complement the MHC II deficiency in a BLS-patient-derived cell line deficient in RFX-B. The remarkable conservation of the RFX-B lineage attests to the conservation of the regulation mechanism for this gene system and its importance to precisely regulate MHC class II molecules in both the developing and active immune response. PMID- 15655669 TI - Capillary zone electrophoresis for simultaneous determination of seven nonsteroidal anti-inflammatory drugs in pharmaceuticals. AB - A simple capillary zone electrophoresis (CZE) method has been developed for analyzing seven nonsteroidal anti-inflammatory drugs (NSAIDs)-sulindac (SU), ketoprofen (KE), indomethacin (IN), piroxicam (PI), nimesulide (NI), ibuprofen (IB), and naproxen (NA). The separation was run using borate buffer (60 mmol L( 1), pH 8.5) containing 13% (v/v) methanol at 20 kV, and detected at 200 nm. Several conditions were studied, including concentration and pH of borate buffer, methanol percentage, and separation voltage. In method validation, the calibration plots were linear over the range 40.0-500.0 micromol L(-1). In intra day and inter-day analysis, relative standard deviations (RSD) and relative errors (RE) were all less than 5%. The limits of detection were 10 micromol L(-1) for SU, IN, PI, and 20 micromol L(-1) for KE, NI, IB, NA (S/N = 3, sampling 6 s by pressure). All recoveries were greater than 95%. This method was applied to the quality control of six NSAIDs in pharmaceuticals using NI as internal standard (IS). The assay results were within the labeled amount required by USP 25. PMID- 15655670 TI - Isolation of selenium organic species from antarctic krill after enzymatic hydrolysis. AB - Total selenium content and its distribution in the soluble and insoluble protein bound fractions obtained after aqueous extraction of antarctic krill samples were determined. About 26% of the total selenium (2.4 microg g-1 dry weight) was found in the supernatant; the rest was in the pellet. Isolation of low molecular selenium-containing fractions was also performed by enzymatic digestion of the protein, followed by size-exclusion chromatography in conjunction with atomic absorption spectrometry. From the applied various proteinases (pronase E, subtilisin Carlsberg, trypsin, chymotrypsin, proteinase and proteinase N from Bacillus subtilis and Novo 0.6 MPX enzyme), the treatment with pronase E led to best recovery of selenium. About 96% of the total Se was found in the hydrolysate, mainly in low molecular weight fractions. Eighty percent of the Se species were in fractions with molecular weights in the range of amino acids and short peptides. High-performance liquid chromatography/inductively coupled plasma mass spectrometry (HPLC-ICP-MS) allowed the identification of selenomethionine and the assumption that selenocystine or its derivatives were the main species in these fractions. PMID- 15655671 TI - Pulmonary fat embolism induced by exposure to high ambient temperature in rats with a fatty liver. AB - The aim of this study was to investigate whether a fatty liver contributes to pulmonary embolism under a high ambient temperature. As an experimental model, we exposed fatty liver rats to a high temperature (45 degrees C) and then looked for fat emboli in the alveolar capillaries using the fat-staining method. Fat emboli were detected in the alveolar capillaries of the fatty liver rats, but not in those of the normal liver rats. Moreover, the degree of pulmonary fat embolism tended to become more severe in proportion to the severity of the fatty liver. In addition, fat emboli did not appear at a core body temperature of 40 degrees C, but were detected at a core body temperature of 44 degrees C. From these results, we conclude that a fatty liver may contribute to the formation of pulmonary fat embolism and that high temperatures act as a trigger for the onset of pulmonary embolism. Moreover, it is possible that fatty liver affects the development of heat stroke induced by exposure to a high ambient temperature and that pulmonary fat embolism is a significant finding which helps to enable a diagnosis of heat stroke in autopsy cases. PMID- 15655672 TI - Heat-shock response is associated with decreased production of interleukin-6 in murine aortic vascular smooth muscle cells. AB - Heat shock has been known to change cellular responses to noxious stimuli by inducing heat-shock proteins (Hsps). We hypothesized that a heat-shock response modulates cytokine production in murine aortic vascular smooth muscle cells (VSMCs). VSMCs were exposed to 44 degrees C for 15-60 min, and subjected to interleukin-1beta (IL-1beta) or tumor necrosis factor alpha (TNFalpha), which induced interleukin-6 (IL-6) production. Expression of Hsps was examined with immunoblots, immunocytochemistry, or enzyme-linked immunosorbent assay (ELISA), and that of IL-6 with reverse transcription-polymerase chain reaction (RT-PCR) or ELISA. Heat shock (44 degrees C for 45 min) induced Hsp72 in VSMCs at 4 h and elicited its maximal expression at 8 h after the end of heat shock. Treatment with IL-1beta increased IL-6 transcription in VSMCs up to 24 h in an incubation time-dependent manner. Treatment with IL-1beta or TNFalpha caused a concentration dependent increase in IL-6 production in culture medium, which was attenuated by heat shock. Although treatment with Hsp72 or Hsp60 alone did not significantly affect basal IL-6 release into culture medium statistically, cotreatment with IL 1beta and Hsp72, but not Hsp60 or boiled Hsp72, decreased IL-1beta-induced IL-6 production in culture medium. Introduction of Hsp72, but not Hsp60, into VSMCs decreased IL-1beta-induced IL-6 production in culture medium. These results indicate that the heat-shock response transcriptionally attenuated production of IL-6 in murine aortic VSMCs. PMID- 15655674 TI - MLPA analysis for the detection of deletions, duplications and complex rearrangements in the dystrophin gene: potential and pitfalls. AB - Duchenne muscular dystrophy (DMD) and Becker muscular dystrophy (BMD) are common X-chromosomal recessive disorders caused by mutations in the dystrophin gene. Using the novel multiplex ligation-dependent probe amplification (MLPA) method we performed retrospective and prospective analyses in a total of 193 individuals. Deletions or duplications were identified in 14 out of 90 families previously tested negative by multiplex PCR or FISH analysis. Partially incorrect results were subsequently identified in two families: the loss of exon 38 signal in one case was due to a p.Q1802X nonsense mutation, whilst in another patient an apparent deletion of exon 37 (coinciding with a duplication of exons 46-53) was caused by a p.R1735C polymorphism. In one case we found a complex rearrangement involving a duplication of two regions: dupEX45-48 and dupEX54-55. We conclude that MLPA is a highly sensitive and rapid alternative to multiplex PCR. It can be used on blood samples, chorionic villi and paraffin-embedded tissue. The ease of detection of duplications and the application for female carrier analysis are clearly the main advantages of the method. However, apparent single exon deletions detected by MLPA should be checked by an independent method. Complex rearrangements such as double mutations on the same allele are rare. PMID- 15655675 TI - Biosynthesis of UDP-xylose: characterization of membrane-bound AtUxs2. AB - UDP-xylose (UDP-Xyl) is a sugar donor for the synthesis of glycoproteins, polysaccharides, various metabolites, and oligosaccharides in plants, vertebrates, and fungi. In plants, the biosynthesis of UDP-Xyl from UDP glucuronic acid (UDP-GlcA) appears to be catalyzed by numerous UDP-glucuronic acid decarboxylase (Uxs) isoforms. For example, six Uxs isoforms in Arabidopsis thaliana (L.) and four in rice have been identified. However, the reason/s for the existence of several isoforms that are necessary for the synthesis of UDP-Xyl remains unknown. Here, we describe a Uxs isoform in Arabidopsis, AtUXS2, encoding an integral membrane protein that appears to be localized to the Golgi apparatus. The enzyme is a dimer and has distinct properties. Unlike the UXS3 isoform, which is shown here to be a soluble protein, the UXS2 isoform is membrane bound. The characteristics of the membrane-bound AtUxs2 and cytosolic AtUxs3 support the hypothesis that unique UDP-GlcA-DCs possessing distinct sub-cellular localizations can spatially regulate specific xylosylation events in plant cells. PMID- 15655673 TI - Tandospirone activates neuroendocrine and ERK (MAP kinase) signaling pathways specifically through 5-HT1A receptor mechanisms in vivo. AB - Tandospirone, an azapirone, is a selective serotonin(1A) (5-HT(1A)) receptor agonist. The effects of tandospirone on plasma hormones and on mitogen-activated protein (MAP) kinase activity in the brain of male rats were studied. Tandospirone produced a time- and dose-dependent increase in plasma levels of oxytocin, adrenocorticotropin (ACTH), corticosterone, and prolactin. The minimal dose of tandospirone that led to a significant elevation of plasma oxytocin, ACTH, and prolactin levels was 1.0 mg/kg (s.c.), while the minimal dose for corticosterone release was 3.0 mg/kg (s.c.). The ED(50) of tandospirone was 1.3 mg/kg for oxytocin, 1.2 mg/kg for ACTH, 3.0 mg/kg for corticosterone, and 0.24 mg/kg for prolactin. Pretreatment with the specific 5-HT(1A) receptor antagonist WAY 100,635 (0.3 mg/kg, s.c.) completely blocked the effects of tandospirone on plasma levels of oxytocin, ACTH, and corticosterone but shifted the dose-response curve for prolactin to the right. Tandospirone injection (10 mg/kg, s.c.) stimulated the MAP kinase signaling cascade, specifically the phosphorylation of p42/44 extracellular signal-regulated kinase (ERK). Western blot analysis revealed a significant increase in phosphorylated ERK (p-ERK) levels in the hypothalamic paraventricular nucleus (PVN) as well as the dorsal raphe nucleus 5 min following tandospirone injection. These increases were blocked by pretreatment with WAY 100,635 (0.3 mg/kg). The results are the first evidence that systemic 5-HT(1A) receptor agonist administration produces a rapid increase in p-ERK levels in vivo, providing further insight into the signaling mechanisms of the 5-HT(1A) receptor. PMID- 15655677 TI - 11C-methionine PET versus 99mTc-sestamibi in the pre-operative localisation of hyperfunctional parathyroid tissue. PMID- 15655678 TI - Radiolabelling DOTA-peptides with 68Ga. AB - PURPOSE: A new field of interest is the application of 68Ga-labelled DOTA conjugated peptides for positron emission tomography (PET). The commercially available or house-made generators require time-consuming and tedious handling of the eluate. Radiolabelling at high specific activities without further purification is not possible, while high specific activities are necessary for peptides that potentially display pharmacological side-effects. Here we present the practical aspects and the results of radiolabelling DOTA-peptides with a TiO2 based commercially available 68Ge/68Ga generator. METHODS: Reaction kinetics and parameters influencing the incorporation of the radionuclide at the highest achievable specific activity were investigated. Since high finger doses were anticipated during handling of the high beta-energy emitter 68Ga, finger dosimetric measurements were performed during radiolabelling and in vivo administration. RESULTS: Fractionated elution of the generator revealed that 80% of the radioactivity was recovered in 1 ml. Bi- and trivalent ionic contaminants that compete for the incorporation of the radionuclide were below 50 nM; thus further tedious and time-consuming purification was avoided. Radiolabelling was performed at pH 3.5-4. Plastic shielding (> or =7-mm wall thickness) around the syringe during administration effectively eliminated the positrons. In rats 68GaCl3 had slow clearance from blood, while 68Ga-EDTA was rapidly cleared via the kidneys. Uptake of 68Ga-DOTATOC in somatostatin receptor-positive tissues was high, with no significant difference between 1 and 4 h post injection. CONCLUSION: DOTA-peptides for PET imaging can be labelled with 68Ga up to specific activities of 1 GBq per nmol within 20 min, enabling the clinical application of peptides that display potential pharmacological side-effects. PMID- 15655680 TI - [Vaccinations in adults--who? when? why?]. AB - Despite the success of childhood vaccinations in Europe, many infectious diseases pose a threat to adults, particularly because immunity induced by vaccination is not life long in some cases. This paper presents the rationale for adult boosters for diphtheria, hepatitis B, pertussis, poliomyelitis and tetanus. Moreover indication for adult-vaccinations against measles, mumps, German measles and varicella is discussed, as a significant part of the population in Germany is susceptible or without known immunity/vaccination history. Finally, immunisation of the elderly against infections with influenza-virus and S. pneumoniae will be described. PMID- 15655679 TI - Effect of substrate feed rate on recombinant protein secretion, degradation and inclusion body formation in Escherichia coli. AB - The effect of changes in substrate feed rate during fedbatch cultivation was investigated with respect to soluble protein formation and transport of product to the periplasm in Escherichia coli. Production was transcribed from the P(malK) promoter; and the cytoplasmic part of the production was compared with production from the P(lacUV5) promoter. The fusion protein product, Zb-MalE, was at all times accumulated in the soluble protein fraction except during high-feed-rate production in the cytoplasm. This was due to a substantial degree of proteolysis in all production systems, as shown by the degradation pattern of the product. The product was also further subjected to inclusion body formation. Production in the periplasm resulted in accumulation of the full-length protein; and this production system led to a cellular physiology where the stringent response could be avoided. Furthermore, the secretion could be used to abort the diauxic growth phase resulting from use of the P(malK) promoter. At high feed rate, the accumulation of acetic acid, due to overflow metabolism, could furthermore be completely avoided. PMID- 15655682 TI - [Pathogenesis of the ductal pancreatic adenocarcinoma: implications for future therapies?]. AB - Despite the advances in the knowledge of the molecular pathogenesis, pancreatic cancer remains a devastating disease with an 5-year survival rate below 5%. Current therapeutic approaches are not successful in respect to overall survival. There is a need to transfer the knowledge of basic science to new therapeutic strategies. This review describes the disturbed cell cycle control, anti apoptotic mechanisms, the process of invasion and metastasis and angioneogenesis of pancreatic cancer cells. This knowledge will allow the development of novel therapeutic strategies for the future. PMID- 15655683 TI - [Gastrointestinal oncology]. PMID- 15655681 TI - [Chronic bronchitis, COPD]. AB - Chronic bronchitis is part and precursor of COPD, a complex disease triggered mostly by exposure to cigarette smoke. However COPD develops only in patients with specific susceptibility probably determined by genetic factors or additional risk factors. A specific type of inflammation resides in the bronchial and bronchiolar walls, that infers damage not only to airway structure but also to surrounding alveolar attachments and thus to the lung parenchyma. Chronic bronchitis, fibrosing bronchiolitis and emphysema constitute the three main stems of pathology of the disease but may coexist with varying extent. The clinical picture is therefore quite variable. Treatment exists largely in bronchodilation combining different mechanisms and using long acting drugs usually applied by inhalation. Acute exacerbations promote progression of the disease, which must be counteracted by adaptation and intensification of therapy, in some cases including non invasive or invasive ventilation. Several non-pharmacologic measures such as smoking cessation, rehabilitation, nutritional support, long term oxygen therapy, lung volume reduction and possibly lung transplantation may be available for appropriate patients and have to be considered. PMID- 15655684 TI - [Clinical implications of genetic risk factors of chronic pancreatitis]. AB - The identification of a specific mutation in the human cationic trypsinogen gene in large kindreds with hereditary pancreatitis was the key to understand the genetic background of chronic pancreatitis. Rapidly, other variants within the same gene were identified-even in small families with a minority of patients. Later, mutations of the most important intrapancreatic trypsin inhibitor SPINK1 were found with high prevalence in patients with idiopathic, tropical and alcoholic chronic pancreatitis. We summarize interesting genetic and biochemical findings, point to clinical features and review recommendations for genetic analysis, follow-up and cancer prevention. PMID- 15655685 TI - [Diagnosis and therapy of acute pancreatitis]. AB - In acute pancreatitis the evaluation of severity is as important as the diagnosis. If there is evidence for severe pancreatitis, an immediate intensive care of all organ systems is needed, to avoid complications. Besides clinical signs, serum CRP is the most valuable parameter to define severity. According to present knowledge, a CT-scan is only needed in sepsis or multiorgan failure. Non invasive ventilation should be started early in case of hypoxia. Up to now, no general benefit was detected for antibiotic prophylaxis or enteral nutrition. No consensus exists whether and when endoscopic interventions are superior to surgery in the treatment of infected necrosis. PMID- 15655686 TI - Dynamic patterns of postural sway in ballet dancers and track athletes. AB - We compared the variability and spatiotemporal profile of postural sway of trained ballet dancers to college varsity track athletes under variations in the availability of vision and rigidity of the support surface. We found no differences between the groups according to the variability measures, but variability increased for both groups with eyes closed and on a foam surface. Recurrence quantification analysis revealed that the postural sway of dancers was less regular (lower recurrence), less stable (lower maxline), less complex (lower entropy), and more stationary (lower absolute trend) than that of track athletes. Dancers, possibly as a result of focused balance training, exhibited different dynamic patterns of postural sway. PMID- 15655687 TI - Neck muscle length modulates nociceptive reflex evoked by noxious irritant application to rat neck tissues. AB - The application of mustard oil (MO), a small-fibre excitant and inflammatory irritant, into neck paraspinal muscles of the rat has been shown to produce a significant reflexive increase in electromyographic (EMG) activity in both neck and jaw muscles. It is possible that this nociceptive reflex activity is influenced by muscle length since recent evidence indicates that abnormal neck posture may be associated with cervical musculoskeletal disorders and pain. Therefore, the aim of this study was to test if muscle length modulates this nociceptive reflex response. Three different experimental procedures were employed in rats under halothane anesthesia: (1) MO injected into the left deep neck muscles with the rat placed in a straight body position (Straight group, n = 7); (2) MO injected into lengthened left deep neck muscles with the rat's neck rotated 45 degrees to the right with respect to the head (Stretched group, n = 11); and (3) MO injected into the right deep neck muscles with the rat's neck rotated 45 degrees to the right (Relaxed group, n = 9). The EMG activity of the deep neck, trapezius, and digastric muscles was bilaterally recorded, rectified and integrated into area under the curve (AUC). Control injections of the vehicle, mineral oil, did not evoke any muscle activity but MO evoked EMG activity in the ipsilateral deep neck and trapezius muscles of the Stretched group that was significantly greater than that evoked in the same muscles in the Straight and Relaxed groups. Also, the MO-evoked EMG activity in the contralateral deep neck muscles of the Stretched and Relaxed groups was greater than that of the corresponding muscles in the Straight group. The MO-evoked activity in the digastric, a jaw muscle whose length was not changed, did not show any significant difference between the three groups. These findings indicate that MO application to the rat deep neck muscles results in a larger nociceptive reflex in deep neck and trapezius muscles when they are stretched. This enhanced muscle activity could be associated with changes in the susceptibility of the neck muscles to pain or damage. PMID- 15655688 TI - Do multitrophic interactions override N fertilization effects on Operophtera larvae? AB - We examined how performance of Operophtera brumata (Lepidoptera) larvae was affected by nitrogen (N) fertilization of boreal forest understorey vegetation. We monitored larval densities on Vaccinium myrtillus plants for a period of 7 years in a field experiment. Preliminary results indicated that the N effect on larval densities was weak. To examine if this was due to indirect interactions with a plant pathogen, Valdensia heterodoxa, that share the same host plant, or due to top-down effects of predation, we performed both a laboratory feeding experiment (individual level) and a bird exclusion experiment (population level) in the field. At the individual level, altered food plant quality (changes in plant concentration of carbon, N, phenolics, or condensed tannins) due to repeated infection by the pathogen had no effect on larval performance, but both survival to the adult stage and adult weight were positively affected by N fertilization. Exclusion of insectivorous birds increased the frequency of larval damage on V. myrtillus shoots, indicating higher larval densities. This effect was stronger in fertilized than in unfertilized plots, indicating higher bird predation in fertilized plots. Predation may thus explain the lack of fertilization effect on larval densities in the field experiment. Our results suggest that top-down effects are more important for larval densities than bottom up effects, and that bird predation may play an important role in population regulation of O. brumata in boreal forests. PMID- 15655690 TI - Functional regularity: a neglected aspect of functional diversity. AB - Functional diversity has been identified as a key to understanding ecosystem and community functioning. However, due to the lack of a sound definition its nature and measurement are still poorly understood. In the same way that species diversity can be split into species richness and species evenness, so functional diversity can be split into functional richness (i.e. the amount of functional trait/character/attribute space filled) and functional evenness (i.e. the evenness of abundance distribution in functional trait space). We propose a functional regularity index (FRO) as a measure of functional evenness for situations where species are represented only by a single functional trait value (e.g. mean, median or mode), and species abundances are known. This new index is based on the Bulla O index of species evenness. When dealing with functional types or categorical functional traits, the Bulla O or any other accepted species evenness index may be used directly to measure functional evenness. The advantage of FRO is that it supplies a measure of functional evenness for continuous trait data. The FRO index presented in this paper fulfils all the a priori criteria required. We demonstrate with two example datasets that a range of FRO values may be obtained for both plant and animal communities. Moreover, FRO was strongly related to ecosystem function as seen in photosynthetic biomass in plant communities, and was able to differentiate sampling stations in a lagoon based on the functional traits of fish. Thus, the FRO index is potentially a highly useful tool for measuring functional diversity in a variety of ecological situations. PMID- 15655689 TI - Habitat structural complexity and morphological diversity of fish assemblages in a Neotropical floodplain river. AB - High species richness and evenness in structurally complex habitats has been hypothesized to be associated with niche partitioning. To test this idea, relationships between habitat structural complexity in river littoral-zone habitats and morphological diversity of tropical fishes were examined in the Cinaruco River, Venezuela. Six habitat attributes were quantified in 45 sites spanning a range of structural complexity. Fishes were collected during day and night to estimate species density and relative abundances at each site. Twenty two morphological variables were measured for each species. Principal components analysis (PCA) of physical habitat data yielded two axes that modeled >80% of variation across sites. The first two axes from PCA of fish morphological variables modeled >70% of variation. Species density during both day and night was negatively associated with flow velocity and positively associated with habitat complexity. Similarity of day and night samples from the same site was significantly greater for sites with high habitat complexity and low flow. In general, mean local assemblage morphological PC scores were not significantly associated with habitat PC scores. Average, maximum, and standard deviation of morphological Euclidean distances of local assemblages revealed positive associations with structural complexity and negative associations with flow. These relationships held even when the positive relationship of species density was statistically removed from assemblage morphological patterns. Findings suggest that both species niche compression and assemblage niche space increase when habitat complexity is greater and flow velocity is lower in this tropical lowland river. PMID- 15655691 TI - A genetic analysis of relative growth rate and underlying components in Hordeum spontaneum. AB - Species from productive and unproductive habitats differ inherently in their relative growth rate (RGR) and a wide range of correlated quantitative traits. We investigated the genetic basis of this trait complex, and specifically assessed whether it is under the control of just one or a few genes that can act as 'master switches' by simultaneously affecting a range of traits in the complex. To address this problem, we crossed two Hordeum spontaneum lines originating from two habitats that differ in productivity. The F3 offspring, in which parental alleles are present in different combinations due to recombination and segregation, was analysed for RGR and its underlying components (leaf area ratio, unit leaf rate, photosynthesis, respiration), as well as a number of other physiological and morphological parameters. For this intra-specific comparison, we found a complex of positively and negatively correlated traits, which was quite similar to what is generally observed across species. A quantitative trait loci (QTL) analysis showed three major and one minor QTL for RGR. Most other variables of the growth-trait complex showed fewer QTLs that were typically scattered over various locations on the genome. Thus, at least in H. spontaneum, we found no evidence for regulation of the trait complex by one or two master switches. PMID- 15655692 TI - Assessing determinants of community biomass composition in two-species plant competition studies. AB - A method is proposed for assessing the relative importance of species identity, neighbour species influence and environment as determinants of change in community biomass composition in two-species short-term competition experiments. The method is based on modelling the differences in relative growth rates (RGR) of species (hence called the RGRD method). Using a multiple regression approach it quantifies the effects of initial species' abundance, species identity and environment on RGRD and hence on change in community biomass composition. The RGRD approach is relatively simple to use and deals readily with statistical difficulties associated with correlated responses between species from the same stand. It can be easily adapted to analyse sequential harvest data. An example based on data from two-species mixtures of the annual species Stellaria media and Poa annua is used to illustrate the method. The main determinant of change in community biomass composition was species identity, reflected in the difference in growth rates between the species. Change in community composition was not, in general, significantly affected by the influence of neighbours or fertiliser level. The unimportance of the influence of neighbours in affecting the composition of these communities contrasts with the strong role of intra- and interspecific competition in determining the size of individuals of both species (Connolly et al. in Oecologia 82:513-526, 1990). PMID- 15655693 TI - Evidence for the enemy release hypothesis in Hypericum perforatum. AB - The enemy release hypothesis (ERH), which has been the theoretical basis for classic biological control, predicts that the success of invaders in the introduced range is due to their release from co-evolved natural enemies (i.e. herbivores, pathogens and predators) left behind in the native range. We tested this prediction by comparing herbivore pressure on native European and introduced North American populations of Hypericum perforatum (St John's Wort). We found that introduced populations occur at larger densities, are less damaged by insect herbivory and suffer less mortality than populations in the native range. However, overall population size was not significantly different between ranges. Moreover, on average plants were significantly smaller in the introduced range than in the native range. Our survey supports the contention that plants from the introduced range experience less herbivore damage than plants from the native range. While this may lead to denser populations, it does not result in larger plant size in the introduced versus native range as postulated by the ERH. PMID- 15655694 TI - Inter-specific competitors reduce inter-gender competition in Negev Desert gerbils. AB - We examined gender-dependent competitive interactions between two nocturnal desert gerbil species, Gerbillus andersoni allenbyi and G. pyramidum, by a field manipulation experiment. The study was done in two 1-ha enclosed plots and included allopatric (only G. a. allenbyi) and sympatric (both species together) treatments. Seed trays and thermal imaging cameras were used to observe the gerbils' foraging activities and aggressive interactions. We found that the negative effect of the competitively dominant species, G. pyramidum, on time spent in seed trays, and ability to control these artificial food patches, was stronger on male than on female G. a. allenbyi. Consequently, the aggression of male G. a. allenbyi towards female G. a. allenbyi was markedly reduced, indicating that the dominant species mediated competition between the genders of the subordinate species. Furthermore, this interference-mediated indirect effect was associated with a decrease in the body mass of male G. a. allenbyi and an increase in the survival of female G. a. allenbyi. We suggest that both the reduction in intra-specific aggression and the positive effect on female survival can potentially stabilize competitive interactions and promote coexistence in this small mammal community. PMID- 15655695 TI - Retrovirus mediated gene transduction of human T-cell subsets. AB - PURPOSE: Allogeneic bone marrow transplantation (AlloBMT) can be curative for patients with leukaemia. Graft versus host disease (GVHD) is a potentially life threatening complication of AlloBMT mediated by the T cells contained within the graft. In order to be able to control GVHD, the allogeneic T cells may be transduced with a suicide gene such as herpes simplex virus thymidine kinase (HSV tk). For this strategy to be successful, all subsets of T cells should be transduced to a similar extent. Also, the transduction protocol should not induce expression of unwanted homing receptors, nor should it lead to unwanted skewing of the T-cell receptor repertoire. We have studied the transduction efficiency of naive and memory subsets of CD4+ and CD8+ T cells, and examined the transduced T cell subsets for possible changes in T-cell receptor (TCR) repertoire and homing receptor expression. METHODS: The cells were transduced using a Moloney murine retroviral vector carrying a conjugate of the genes encoding the truncated form of the cell surface marker, low affinity nerve growth factor receptor (DeltaLNGFR) and HSV-tk. Transduction efficiency and homing receptor expression were quantified by flow cytometry. TCR repertoire was determined by spectratyping. RESULTS: We obtained a transduction efficiency of 30-50% of the cells, with no difference between the T-cell subsets. Cell surface receptors responsible for homing to skin, gastrointestinal tract or lymph nodes were practically absent at the end of 2 weeks in culture. The activation procedure seemed to favour the expansion of certain T-cell clones over polyclonal populations. However, there was no difference in the TCR repertoire between transduced and non-transduced cells. CONCLUSION: Changes in the composition of the T-cell subsets at the end of the cell culture were the results of the activation, and not the suicide gene transduction. The transduced T cells did not express unwanted homing receptors. PMID- 15655696 TI - Bone densitometry in pediatric patients treated with pamidronate. AB - BACKGROUND: Increasing numbers of children are being treated with the bisphosphonate pamidronate for low bone mineral density, particularly children with increased risk of fractures caused by bone disorders or low/non-weight bearing. OBJECTIVE: To determine the effect of intravenous pamidronate on the bone mineral density of children with osteogenesis imperfecta and spastic quadriplegic cerebral palsy. MATERIALS AND METHODS: Charts of 38 children with osteogenesis imperfecta (n=20) and spastic quadriplegic cerebral palsy (n=18) treated with pamidronate were retrospectively reviewed. Patients were selected for treatment because of prior fracture and/or abnormally low bone mineral density. All received intravenous pamidronate at two-month to eight-month intervals and were periodically examined using dual energy X-ray absorptiometry. RESULTS: All patients had abnormally low bone mineral density prior to treatment. Lumbar spine bone mineral density and z-scores showed serial improvement in 31 of 32 patients. Spine bone mineral density increased 78+/-38.1% in OI and 47.4+/ 39.0% in children with cerebral palsy. The area of greatest lateral distal femur bone mineral density improvement was in the metaphysis adjacent to the growth plate, with a 96+/-87.8% improvement in the osteogenesis imperfecta group and 65.7+/-55.2% improvement in the cerebral palsy group. Increases in bone mineral density exceeded that expected for age-specific growth. This was demonstrated by improvement in both spine and femur z-scores for both groups. No children with spastic quadriplegic cerebral palsy experienced fractures after the first week of treatment, whereas patients with osteogenesis imperfecta continued to have fractures but at a decreased rate. CONCLUSIONS: Intravenous pamidronate given at 3- to 4-month intervals proved to be effective in increasing bone mineral density in patients with osteogenesis imperfecta and spastic quadriplegic cerebral palsy. The greatest gains in bone mineral density were observed in the children with osteogenesis imperfecta, but they did continue to fracture, albeit at a decreased rate. Children with cerebral palsy gained bone mineral density and did not continue to fracture. PMID- 15655698 TI - Cervical pregnancy treated with transvaginal ultrasound-guided intra-amniotic instillation of methotrexate. AB - AIM: Aim of the study was to investigate the efficacy of single transvaginal ultrasound-guided intraamniotic installation of methotrexate in the management of cervical pregnancy with concurrent review of the literature. MATERIALS AND METHODS: Six patients with cervical pregnancy are included in the study. All patients were treated with single transvaginal ultrasound-guided intraamniotic installation of 70 mg of methotrexate plus folic acid p.o. The main presenting symptoms were mild to moderate vaginal bleeding and lower abdominal cramp-like pain, resembling the clinical presentation of a threatened abortion. The typical ultrasound findings were the absence of intrauterine gestational sac and the detection of a gestational sac within the cervical canal, invading the anterior or the posterior wall of the cervix and normal appearance of the adnexa, bilaterally. The hourglass-shaped cervix was not characteristic at 5 weeks of gestation but it was at 8 weeks of gestation. DISCUSSION: Ultrasound-guided intraamniotic installation of methotrexate in the management of cervical pregnancy appears to be an effective and safe method but the choice of the method should be depended on the gestational age of cervical pregnancy, the presence of active bleeding or not and its severity, the desire for preservation of future fertility, the presence of coexisting valuable intrauterine pregnancy and the experience of the physician in charge. PMID- 15655697 TI - Chorioamnionitis risk and neonatal outcome in preterm premature rupture of membranes. AB - OBJECTIVE: The objective of this study was to compare the neonatal outcome in patients with preterm premature rupture of membranes with and without clinical chorioamnionitis. STUDY DESIGN: This is a retrospective study that included 254 pregnant women with preterm rupture of membranes. The study group was divided according to the presence or absence of clinical chorioamnionitis defined as the presence of two or more of the following criteria: maternal temperature >38 degrees C on two or more occasions > or =1 h apart, maternal tachycardia (> or =120 beats/min), uterine tenderness, foul smelling amniotic fluid, maternal leukocytosis > or =20,000 mm(-3) with bands and positive C reactive protein. Also the study population was divided according to the use of tocolysis. Exclusion criteria included multiple pregnancy, fetal congenital anomalies, diabetes mellitus and severe preeclampsia. Amniotic fluid was collected from the cervix or from the transabdominal amniocentesis. Antibiotics and tocolysis were used according to the hospital protocols. Parametric and nonparametric statistics were used for comparisons. RESULTS: There were no significant differences in birth weight, Apgar scores at 1 and 5 min, rates of respiratory distress syndrome, intraventricular hemorrhage and necrotizing enterocolitis between patients with and without clinical chorioamnionitis or between women who received tocolysis and the ones that did not receive tocolysis. In cases of clinical chorioamnionitis and when tocolysis was used the neonates stayed longer in the neonatal intensive care unit (NICU). CONCLUSION: Patients with preterm premature rupture of membranes and clinical chorioamnionitis have similar neonatal outcomes than the ones without clinical chorioamnionitis. PMID- 15655699 TI - A major site of expression of the ets transcription factor Elf5 is epithelia of exocrine glands. AB - Elf5 belongs to the ets family of transcription factors and was cloned by homology in the DNA binding domain to the related, epithelial-specific ets factor, Elf3. Elf5 mRNA is expressed highly in normal tissue rich in secretory epithelial cells, including mammary gland, lung, kidney, prostate, salivary gland and stomach. The function of Elf5 and the cell types in which it is expressed remain uncharacterised. The presence of Elf5 mRNA in normal tissues, but absence in cancer tissues, may suggest a role for Elf5 in differentiation and development. We have generated a rabbit antiserum directed against a peptide in the Elf5 DNA-binding domain that is conserved between murine and human sequences. The antiserum specifically detects human and murine Elf5 proteins on western blots and shows specific staining on paraffin-embedded sections obtained from tissues including mammary gland, kidney, salivary gland and stomach. Epithelia from the bladder lining, lung and prostate did not stain for the presence of Elf5, though these organs express Elf5 mRNA. We show for the first time that Elf5 is primarily expressed in epithelial cells and is likely to be an epithelial specific protein. The antiserum should prove useful in further analysis of the expression and function of Elf5. PMID- 15655700 TI - Incretin hormones and insulin secretion. PMID- 15655701 TI - A centenary of gastrointestinal endocrinology. AB - Gastrointestinal hormones are peptides released to circulation from endocrine cells as well as neurons in the gastrointestinal tract. More than 30 hormone genes are currently known to be expressed in the stomach and intestines, which makes the gut the largest endocrine organ in the body. Moreover, cell and molecular biology now makes it feasible to conceive gastrointestinal endocrinology under five general headings: 1) The structural homology groups the hormones into eight families, each of which is assumed to originate from a common ancestral gene; 2) the individual hormone gene often have multiple phenotypes due to alternative splicing of the primary transcript, tandem organization of the translational product or differentiated maturation of the prohormone. By a combination of these mechanisms, more than 100 different hormonally active peptides are released from the gastrointestinal tract; 3) in addition, gut hormone genes are also widely expressed outside the gut, some only in neurons and/or in endocrine cells, but others also in other extraintestinal cell-types; 4) the different cell types may express different hormonally active fragments of the same prohormone by variation in the cell-specific posttranslational processing. Finally, 5) endocrine cells, neurons, and spermatozoa display different cell-specific release of gut peptides, so the same peptide may act as a metabolic blood-borne hormone, as a neurotransmitter, as a long-acting growth factor, and as an acute fertility factor. PMID- 15655702 TI - Overview of incretin hormones. AB - Incretins are hormones released by nutrients from the GI tract. They amplify glucose-induced insulin release. By raising circulating incretin levels, oral glucose provokes a higher insulin response than that resulting from intravenous glucose. The two most important incretin hormones are glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1). In patients with type 2 diabetes, the incretin effect is decreased, mainly due to loss of the GIP-regulated second phase of insulin secretion, and because of a decreased secretion of GLP-1. In addition to its insulinotropic effect, GLP-1 inhibits glucagon release, prolongs gastric emptying, and leads to decreases in body weight, all of which explain the marked antidiabetogenic effect of this incretin hormone. PMID- 15655703 TI - On the physiology of GIP and GLP-1. AB - Recent studies have indicated that GIP and GLP-1 are about as important as each other in the incretin effect, being released rapidly after meals and being active already at fasting glucose levels. Although the density of GLP-1 producing cells is higher distally, GlP-1 is normally secreted by jejunal L-cells, explaining the rapid onset of secretion. Moreover, many endocrine cells in the small intestine appear to produce both GIP and GLP-1. Both are metabolized rapidly by the dipeptidyl peptidase IV enzyme, but unlike GIP, about 90 % of GLP-1 is degraded before it reaches the systemic circulation. Apparently, before it is degraded, GLP-1 activates sensory afferents in the gastrointestinal mucosa with cell bodies in the nodose ganglion, signaling onwards to the brain stem and the hypothalamus. A similar mechanism seems to be involved in GLP-1's effect on gastrointestinal motility and secretion, and perhaps its actions on appetite and food intake, all of which may be even more physiologically important than its effects on the beta cells. Cardiovascular and neuroprotective actions of GLP-1 have also recently been reported. Regarding GIP, several lines of evidence suggest that GIP, in addition to its incretin effects, may affect lipid metabolism and promote lipid storage. PMID- 15655704 TI - Nutrient, neural and endocrine control of glucagon-like peptide secretion. AB - The intestinal glucagon-like peptides, GLP-1 and GLP-2, are important regulators of nutrient intake, digestion, absorption and metabolism. Extensive studies have shown that the co-secretion of GLP-1 and GLP-2 is regulated by a variety of dietary nutrient, neural and endocrine inputs. Furthermore, secretion of these peptides is altered in a number of pathological conditions, including type 2 diabetes and obesity. The purpose of this review is to discuss the regulation of GLP-1 and GLP-2 secretion in both health and disease. PMID- 15655706 TI - Physiology of GLP-1--lessons from glucoincretin receptor knockout mice. AB - GLP-1 has both peripheral and central actions, as this hormone is secreted by gut endocrine cells and brainstem neurons projecting into the hypothalamus and other brain regions. GLP-1 has multiple regulatory functions participating in the control of glucose homeostasis, beta-cell proliferation and differentiation, food intake, heart rate and even learning. GLP-1 action depends on binding to a specific G-coupled receptor linked to activation of the adenylyl cyclase pathway. Analysis of mice with inactivation of the GLP-1 receptor gene has provided evidence that absence of GLP-1 action in the mouse, despite this hormone potent physiological effects when administered in vivo, only leads to mild abnormalities in glucose homeostasis without any change in body weight. However, a critical role for this hormone and its receptor was demonstrated in the function of the hepatoportal vein glucose sensor, in contrast to that of the pancreatic beta cells, although absence of both GLP-1 and GIP receptors leads to a more severe phenotype characterized by a beta-cell-autonomous defect in glucose-stimulated insulin secretion. Together, the studies of these glucoincretin receptor knockout mice provide evidence that these hormones are part of complex regulatory systems where multiple redundant signals are involved. PMID- 15655705 TI - Circulation and degradation of GIP and GLP-1. AB - The incretin hormones glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1) are secreted from the intestinal K- and L-cells, respectively, but are immediately subject to rapid degradation. GLP-1 is found in two active forms, amidated GLP-1 (7-36) amide and glycine-extended GLP-1 (7-37), while GIP exists as a single 42 amino acid peptide. The aminopeptidase, dipeptidyl peptidase IV (DPP IV), which is found in the endothelium of the local capillary bed within the intestinal wall, is important for the initial inactivation of both peptides, with GLP-1 being particularly readily degraded. DPP IV cleavage generates N-terminally truncated metabolites (GLP-1 (9-36) amide / (9-37) and GIP (3-42)), which are the major circulating forms. Subsequently, the peptides may be degraded by other enzymes and extracted in an organ-specific manner. However, other endogenous metabolites have not yet been identified, possibly because existing assays are unable either to recognize them or to differentiate them from the primary metabolites. Neutral endopeptidase 24.11 has been demonstrated to be able to degrade GLP-1 in vivo, but its relevance in GIP metabolism has not yet been established. Intact GLP-1 and GIP are inactivated during passage across the hepatic bed by DPP IV associated with the hepatocytes, and further degraded by the peripheral tissues, while the kidney is important for the final elimination of the metabolites. PMID- 15655707 TI - Physiology of GIP--a lesson from GIP receptor knockout mice. AB - A much greater insulin response is observed after oral glucose load than after intravenous injection of glucose. The hormonal factor(s) implicated as transmitters of signals from the gut to pancreatic beta-cells was referred to incretin; gastric inhibitory polypeptide or glucose-dependent insulinotropic polypeptide (GIP) is identified as one of the incretins. GIP exerts its effects by binding to its specific receptor, the GIP receptor, which is expressed in various tissues including pancreatic islets, adipose tissue, and brain. However, the physiological role of GIP has been generally thought to stimulate insulin secretion from pancreatic beta-cells, and the other actions of GIP have received little attention. We have bred and characterized mice with a targeted mutation of the GIP receptor gene. From these studies, we now know that GIP not only mediates early insulin secretion by acting on pancreatic beta-cells, but also links overnutrition to obesity by acting on adipocytes. PMID- 15655708 TI - Beta- and alpha-cell dysfunction in type 2 diabetes. AB - Insulin resistance is a common pathogenetic feature of type 2 diabetes. However, hyperglycemia would not develop if a concomitant defect in insulin secretion were not present. Impaired insulin secretion results from functional and survival defects of the beta-cell. The functional defects can be demonstrated early in the natural history of diabetes and they are hallmarked by abnormal pulsatility of basal insulin secretion and loss of first-phase insulin release in response to a glucose challenge. Moreover, a significant reduction of the beta-cell mass is apparent at the time of the diagnosis of diabetes. The progressive increase in glucose levels, that seems to characterize the natural history of type 2 diabetes, has been claimed to be largely due to progressive reduction of function and mass of beta-cells. Although a genetic predisposition is likely to account for impaired insulin secretion, chronic exposure to hyperglycemia and high circulating FFA is likely to contribute to both functional and survival defects. The disturbance in the endocrine activity of the pancreas is not limited to insulin, since a concomitant increase in fasting plasma glucagon and impaired suppression after the ingestion of an oral glucose load are often observed. This alteration becomes prominent after the ingestion of a mixed meal, when plasma glucagon remains much higher in the diabetic patient as compared to normal individuals. The disproportionate changes in the plasma concentration of the two pancreatic hormones is clearly evident when the insulin:glucagon molar ratio is considered. It is the latter that mainly affects hepatic glucose production. Because of the reduction of the insulin:glucagon molar ratio basal endogenous glucose concentration will be higher causing fasting hyperglycemia, while the hepatic glucose output will not be efficiently suppressed after the ingestion of a meal, contributing to excessive post-prandial glucose rise. Correcting beta- and alpha-cell dysfunction becomes, therefore, an attractive and rational therapeutic approach, particularly in the light of new treatments that may directly act on these pathogenetic mechanisms of type 2 diabetes. PMID- 15655709 TI - Glucose-dependent insulinotropic polypeptide as a regulator of beta cell function and fate. AB - Glucose-dependent insulinotropic polypeptide (GIP) is a gastrointestinal hormone that is secreted in response to food intake and modulates beta cell function. It may also regulate beta cell fate. Released from the nutrient-sensing K-cells of the upper intestine, GIP acts on various tissues, including pancreatic beta cells, via interaction with its G-protein-coupled receptor. Perhaps the most important effect of GIP is its potentiation of insulin secretion. Indeed, pharmacological blockade or genetic knockout of its receptor delays glucose dependent insulin secretion. Exposure to GIP also enhances the beta cell response to future nutrient stimulation and upregulates transcription of key beta cell genes. There is emerging evidence that like the related hormone glucagon-like peptide-1, GIP may function as a beta cell growth factor and anti-apoptotic agent, further supporting a role for this hormone in balancing beta cell function to changing metabolic conditions. Overproduction of GIP in response to increased nutrient loads may, however, contribute to the pathophysiology of obesity. Interestingly, its insulinotropic effect is lost in type 2 diabetes, perhaps because of hyperglycemia-induced receptor desensitization. A better understanding of GIP's effects on the beta cell under normal and pathological conditions may facilitate the design of GIP derivatives for the treatment of metabolic disorders. PMID- 15655712 TI - The role of GLP-1 in the life and death of pancreatic beta cells. AB - Glucagon-like peptide-1 (GLP-1), a peptide hormone produce by intestinal cells, has recently been shown to be capable of modulating islet cell mass. Administration of GLP-1 to rodent models of type 2 diabetes ameliorates insulin secretion, induces the replication of islet cells, and promotes islet-cell neogenesis from pancreatic ductal cells susceptible to transdifferentiate in insulin-producing cells. In addition, an anti-apoptotic effect of GLP-1 has been described in hyperglycemic animal models, using freshly isolated human islets or cultured beta cell lines exposed to various pro-apoptotic stimuli. The aim of this article is to review those reports that have emphasized the role of GLP-1 as a regulator of islet cell mass. PMID- 15655710 TI - New insights concerning the glucose-dependent insulin secretagogue action of glucagon-like peptide-1 in pancreatic beta-cells. AB - The GLP-1 receptor is a Class B heptahelical G-protein-coupled receptor that stimulates cAMP production in pancreatic beta-cells. GLP-1 utilizes this receptor to activate two distinct classes of cAMP-binding proteins: protein kinase A (PKA) and the Epac family of cAMP-regulated guanine nucleotide exchange factors (cAMPGEFs). Actions of GLP-1 mediated by PKA and Epac include the recruitment and priming of secretory granules, thereby increasing the number of granules available for Ca(2+)-dependent exocytosis. Simultaneously, GLP-1 promotes Ca(2+) influx and mobilizes an intracellular source of Ca(2+). GLP-1 sensitizes intracellular Ca(2+) release channels (ryanodine and IP (3) receptors) to stimulatory effects of Ca(2+), thereby promoting Ca(2+)-induced Ca(2+) release (CICR). In the model presented here, CICR activates mitochondrial dehydrogenases, thereby upregulating glucose-dependent production of ATP. The resultant increase in cytosolic [ATP]/[ADP] concentration ratio leads to closure of ATP-sensitive K(+) channels (K-ATP), membrane depolarization, and influx of Ca(2+) through voltage-dependent Ca(2+) channels (VDCCs). Ca(2+) influx stimulates exocytosis of secretory granules by promoting their fusion with the plasma membrane. Under conditions where Ca(2+) release channels are sensitized by GLP-1, Ca(2+) influx also stimulates CICR, generating an additional round of ATP production and K-ATP channel closure. In the absence of glucose, no "fuel" is available to support ATP production, and GLP-1 fails to stimulate insulin secretion. This new "feed forward" hypothesis of beta-cell stimulus-secretion coupling may provide a mechanistic explanation as to how GLP-1 exerts a beneficial blood glucose lowering effect in type 2 diabetic subjects. PMID- 15655711 TI - Glucagon-like peptide-1 and islet lipolysis. AB - A role for glucagon-like peptide 1 (GLP-1) has been suggested in stimulating beta cell lipolysis via elevation of cAMP and activation of protein kinase A, which in turn may activate hormone-sensitive lipase (HSL), thereby contributing to fatty acid generation (FFA) from intracellular triglyceride stores. FFAs may then be metabolized to a lipid signal, which is required for optimal glucose-stimulated insulin secretion. Since HSL is expressed in islet beta-cells, this effect could contribute to the stimulation of insulin secretion by GLP-1, provided that a lipid signal of importance for insulin secretion is generated. To examine this hypothesis, we have studied the acute effect of GLP-1 on isolated mouse islets from normal mice and from mice with high-fat diet induced insulin resistance. We found, however, that although GLP-1 (100 nM) markedly potentiated glucose stimulated insulin secretion from islets of both feeding groups, the peptide was not able to stimulate islet palmitate oxidation or increase lipolysis measured as glycerol release. This indicates that a lipid signal does not contribute to the acute stimulation of insulin secretion by GLP-1. To test whether lipolysis might be involved in the islet effects of long-term GLP-1 action, mice from the two feeding groups were chronically treated with exendin-4, a peptide that lowers blood glucose by interacting with GLP-1 receptors, in order to stimulate insulin secretion, for 16 days before isolation of the islets. The insulinotropic effects of GLP-1 and forskolin were exaggerated in isolated islets from exendin-4 treated mice given a high-fat diet, with a augmented palmitate oxidation as well as islet lipolysis at high glucose levels in these islets. Exendin-4 treatment had less impact on mice fed a normal diet. From these results we conclude that while GLP-1 does not seem to induce beta-cell lipolysis acutely in mouse islets, the peptide affects beta-cell fat metabolism after long-term adaptation to GLP-1 receptor stimulation. PMID- 15655713 TI - The development of beta-cell mass: recent progress and potential role of GLP-1. AB - Over the last decade, remarkable strides in incretin hormone biology have laid the foundation for our more recent appreciation that GLP-1 not only regulates mature beta-cell function but also critically regulates beta-cell differentiation, beta-cell proliferation and beta-cell survival. Dysregulated beta-cell growth and function are central to the pathophysiology of both type 1 and type 2 diabetes. Thus, GLP-1 receptor agonists are being intensively developed for the treatment of human diabetes and are likely to become available to clinical use in the near future. A general overview of beta-cell development will be provided, with particular emphasis on recent contributions to our understanding of pancreas and islet development during the embryonic, fetal and neonatal periods. The transcriptional hierarchy and extracellular signals governing events during these periods will be highlighted. Evidence suggesting a role for endogenous GLP-1 and GLP-1 receptor during beta-cell development will be reviewed. Finally, the therapeutic potential for intervention with GLP1 receptor agonists during the neonatal period will be discussed. PMID- 15655714 TI - New insights into the regulation of glucagon secretion by glucagon-like peptide 1. AB - Glucagon-like peptide-1 (GLP-1) is a potent incretin hormone currently under investigation for use as a novel therapeutic agent in the treatment of type 2 diabetes. One of several therapeutically important biological actions of GLP-1 in type 2 diabetic subjects is ability to induce strong suppression of glucagon secretion. The glucagonostatic action of GLP-1 results from its interaction with a specific G-protein coupled receptor resulting in the activation of adenylate cyclase and an increase in cAMP generation. In the pancreatic alpha-cell, cAMP, via activation of protein kinase A, interacts with a plethora of signal transduction processes including ion-channel activity and exocytosis of the glucagon-containing granules. In this short review, we will focus on recent advances in our understanding on the cellular mechanisms proposed to underlie the glucagonotropic action of GLP-1 and attempt to incorporate this knowledge into a working model for the control of glucagon secretion. Studies on the effects of GLP-1 on glucagon secretion are relevant to the pathogenesis of type 2 diabetes due to the likely contribution of hyperglucagonemia to impaired glucose tolerance in type 2 diabetes. PMID- 15655715 TI - Extrapancreatic effects of GIP and GLP-1. AB - Incretin-based therapy promises to be a useful adjunct in the treatment of diabetes. Glucagon-like peptide-1 (GLP1) and, to a lesser extent, glucose dependent insulinotropic polypeptide (GIP) are potent stimulators of insulin secretion, and consequently have significant effects on the regulation of the glucose metabolism. What has been less clear, however, is whether these hormones exert direct effects on glucose metabolism independent of their effect on pancreatic insulin and glucagon release. Glucose effectiveness and insulin action (the ability of glucose and insulin respectively to stimulate glucose uptake and suppress glucose release) have been reported by some investigators, but not others, to improve during incretin infusion. The purpose of this review is briefly to examine some of the numerous conflicting reports in the literature as to the presence or otherwise of extrapancreatic incretin effects. In addition, we will briefly discuss the gastrointestinal effects of incretins. These effects may be of considerable importance in the treatment of postprandial hyperglycemia although they are not, strictly speaking, the result of a direct incretin effect on glucose metabolism. PMID- 15655716 TI - Effects of glucagon-like peptide 1 on the hepatic glucose metabolism. AB - Glucagon-like peptide 1 has important actions in lowering blood glucose, both through its incretin action and by regulating other systems affecting glucose metabolism. There is good evidence that the coordinate regulation of islet hormones by GLP-1 has significant effects on hepatic glucose metabolism, and this likely contributes to the potent effect of GLP-1 on fasting hyperglycemia in diabetic patients. More controversial are potential effects of GLP-1 on hepatic glucose production or storage independent of insulin and glucagon. There are data from in vitro studies supporting an effect of GLP-1 to promote glycogen synthesis in hepatocytes, and several in vivo studies suggesting that GLP-1 has independent effects on hepatic glucose uptake and/or production. However, these findings must be considered against a backdrop of studies that have not demonstrated islet independent actions of GLP-1. This paper will review the current literature addressing hepatic effects of GLP-1 and identify important gaps in the knowledge base for this topic. PMID- 15655717 TI - GLP-1 and extra-islet effects. AB - The main target of action of glucagon-like peptide-1 (GLP-1) is the islet, where the hormone stimulates insulin secretion, promotes beta cell proliferation and neogenesis, and inhibits glucagon secretion. However, GLP-1 receptors are also expressed outside the islets, increasing the likelihood that GLP-1 also plays a role in other organs. These functions are mainly the inhibition of gastric emptying, gastric acid secretion and exocrine pancreatic secretion, indicating that the hormone acts as an enterogastrone--a hormone released from the distal portion of the small intestine that inhibits proximal gastrointestinal events. Another important action of GLP-1 is to induce satiety. Other effects of the hormone include cardioprotection, neuroprotection, induction of learning and memory, stimulation of afferent, sensory nerves, stimulation of surfactant production in the lung, dilatation of pulmonary vessels, induction of diuresis, and also under some conditions, induction of antidiabetic actions unrelated to islet function. Thus, GLP-1 clearly has several manifestations of activity. The physiological relevance of these actions and their contribution to the overall antidiabetic action of GLP-1 when used in treatment of type 2 diabetes remains to be established. PMID- 15655719 TI - Glucagon-like peptide 1 (GLP-1) in the treatment of diabetes. AB - Glucagon-like peptide 1 (GLP-1) was discovered as an incretin (insulinotropic gut) hormone. Biological actions of GLP-1 in healthy and type 2 diabetic subjects include (a) stimulation of insulin secretion in a glucose-dependent manner, (b) suppression of glucagon, (c) reduction in appetite and food intake, (d) deceleration of gastric emptying. In animal experiments, in addition, (e) stimulation of beta-cell neogenesis, growth and differentiation in animal and tissue culture experiments, and (f) in vitro inhibition of beta-cell apoptosis induced by different agents have been observed. Since the incretin effect--the higher insulin secretory response to oral as compared to intravenous glucose loads - is reduced in patients with Type 2 diabetes, GLP-1 has been used to pharmacologically replace incretin. Intravenous GLP-1 can normalise, and subcutaneous GLP-1 can significantly lower plasma glucose in the majority of patients with Type 2 diabetes. The magnitude of this effect does not greatly depend on patient characteristics such as age, sex, obesity, or baseline insulin and glucagon, with minor influences of previous antidiabetic therapy and actual metabolic control. GLP-1 itself, however, is inactivated rapidly in vivo by the protease DPP IV and can only be used for short-term metabolic control, such as in intensive care units (potentially useful in patients with acute myocardial infarction, coronary surgery, cerebrovascular events, septicaemia, during the perioperative period and while on parenteral nutrition). For more long-term metabolic control, incretin mimetics (agonists at the GLP-1 receptor) with more favourable pharmacokinetic profiles should be used. PMID- 15655718 TI - Gastrointestinal hormones and regulation of food intake. AB - Obesity has been described as the greatest current threat to human health. In order to design drugs to target obesity, it is essential to understand its physiology and pathophysiology. Several peptides synthesised in the gastrointestinal tract which affect food intake have been identified including ghrelin, cholecystokinin (CCK), glucagon-like peptide-1 (7-36) amide (GLP-1), oxyntomodulin, peptide YY (PYY) and pancreatic polypeptide (PP). These peptides represent potential targets for the design of anti-obesity drugs. In this article we review recent advances in our understanding of food intake by these gastrointestinal hormones. PMID- 15655721 TI - GLP-1 receptor agonists and DPP-4 inhibitors in the treatment of type 2 diabetes. AB - Glucagon-like peptide-1 (GLP-1) is an incretin hormone with antidiabetic action through its ability to stimulate insulin secretion, increase beta cell neogenesis, inhibit beta cell apoptosis, inhibit glucagon secretion, delay gastric emptying and induce satiety. It has therefore been explored as a novel treatment of type 2 diabetes. A problem is, however, that GLP-1 is rapidly inactivated by the dipeptidyl peptidase-4 (DPP-4) enzyme, which results in a short circulating half-life of the active form of GLP-1 (< 2 min). Two strategies have been employed to overcome this obstacle as a treatment of diabetes. One is to use GLP-1 receptor agonists that have a prolonged half-life due to reduced degradation by DPP-4. These GLP-1 mimetics include exenatide and liraglutide. Another strategy is to inhibit the enzyme DPP-4, which prolongs the half-life of endogenously released active GLP-1. Both these strategies have been successful in animal studies and in clinical studies of up to one year's treatment. This review will summarize the background and the current (mid 2004) clinical experience with these two strategies. PMID- 15655720 TI - GIP as a potential therapeutic agent? AB - Glucose-dependent insulinotropic polypeptide (GIP) is released from K-cells in the gut after meal ingestion, and acts in concert with glucagon-like peptide 1 (GLP-1) to augment glucose-stimulated insulin secretion. While derivatives of GLP 1 are under active investigation for the treatment of type 2 diabetes, the case is different for GIP. Indeed, the insulinotropic effect of GIP is almost absent in patients with type 2 diabetes. In addition, the unfavourable pharmacokinetic profile of native GIP obviates its clinical application. Different analogues of GIP exhibiting prolonged stability and enhanced biological potency have been generated in order improve the anti-diabetic properties of GIP. However, glucose normalisation, as is typically observed during the intravenous administration of GLP-1 in patients with type 2 diabetes, has not yet been achieved with GIP or its derivatives. Since GIP appears to play a role in lipid physiology and elevated levels of GIP have been associated with obesity, antagonising GIP action has been proposed as a therapeutic strategy for obesity. This concept has recently been reinforced by the observation that GIP receptor knock-out mice are protected from high-fat diet-induced obesity. However, eliminating the effect of endogenous GIP may at the same time impair postprandial insulin secretion, thereby severely disturbing glucose homeostasis. Therefore, therapeutic strategies based on either augmenting or antagonising GIP action are far from being established alternatives for the future therapy of type 2 diabetes or obesity. PMID- 15655722 TI - [Microbiological findings in children with an indwelling central venous catheter]. AB - INTRODUCTION: Besides their obvious advantages for the patient, central venous catheters (CVC) also carry the risk of possible infectious complications. The purpose of our investigation was to carry out a microbiological evaluation of a 5 year set of paediatric patients with indwelling CVCs. PATIENTS AND METHODS: In the group were 218 CVCs inserted to 165 children over a period of 5 years. There were 26 multi-lumen catheters (11.927 %) and 192 single-lumen catheters (88.073 %). The mean indwelling period was 10.1 days per 1 CVC. Blood for microbiology was removed by a physician from the CVC after disinfecting its opening under standard sterile conditions into a commercial sampling vessel HEMOD (Imuna, Sarisske Michalany, Slovak Republic) or into a vessel of an automated haemoculture system BactecPeds PLUS/F (Becton Dickinson and Comp., Spark,MA, USA). When removing the tip of the CVC we disinfected, before removing the CVC, the area around the insertion with isopropyl or ethyl alcohol. We released the fixed CVC and 1 minute after disinfection we pulled out the CVC and cut off the end or rather the tip of the catheter (approx. 1-3 cm of the tip) into a sterile test tube. To establish the diagnosis of infectious complications we used the 1995 Sirges-Serra classification and the CDC criteria. RESULTS: In 5 years (1995 1999) we had 71 infectious complications. There were 31 contaminated catheters, 27 cases of catheter sepsis and 11 cases of catheter bacteraemia. With 147 catheters (67.43%) there were no infectious complications. Dominant microbes were Staphylococcus epidermidis (32 cases - 11 from haemocultures and 21 from CVCs) and Candida spp. (30 cases, 17 from haemocultures and 13 from CVCs). Among the microbiological agents of catheter sepsis predominated Gram-negative bacteria. Out of the whole analysed group 41 children (24.8 %) died. CVC as the cause of death was demonstrated in 6 children (3.636 % of patients with CVC). CONCLUSIONS: Microbiological findings in our group are in line with literary data. To reduce the incidence of infectious complications it is important to limit sampling from CVC to a minimum. Insertion of CVCs under strict sterile conditions and aseptic handling of all entries into the central bloodstream reduces to a minimum the risk of infectious complications. PMID- 15655723 TI - [Comparison of EIA Borrelia recombinant IgM of the 3rd generation with EIA test based on whole cell antigen in the diagnosis of Lyme borreliosis]. AB - OBJECTIVES: Evaluation of a new kit of the 3rd generation, EIA Borrelia recombinant IgM in the serodiagnosis of Lyme borreliosis (LB) and comparison with the EIA test based on the whole cell antigen in the detection of IgM antibodies. MATERIAL AND METHODS: In total 123 children (147 sera) were examined, 71 children (93 sera) of whom showed clinically defined stage of LB. The other 54 sera of 52 children were tested for suspicion for LB which was later excluded, and for cross reacting antibodies. They represented the control group of children with other infectious or autoimmune diseases. The EIA B. recombinant IgM kit (Test-Line, CR) is based on the selected antigen fragments: outer surface protein C (OspC) and internal flagellin (p41i). The fragments were selected according to the most frequent Borrelia spp. in the Czech Republic. RESULTS: Samples compared 101/147 (69 %) with both tests showed correspondent reactions. 46 samples reacted inconsistently, 26 from children with LB and 20 from the control group. The total specificity and sensitivity of the recombinant kit in IgM antibody class was 88,9 % and 26,9 %, respectively. In the case of early disseminated LB infection the sensitivity was 35,6 % and it is comparable with the sensitivity of immunoblot. Statistically, there is no difference in the sensitivity (p = 0,101) and specificity (p = 0,383) of EIA B. recombinant IgM and immunoblot. CONCLUSIONS: The EIA B. recombinant IgM kit shows comparable diagnostic sensitivity in children with acute LB together with significantly higher specificity in children with non-specific response in kits based on the whole cell antigen and at the same time high conformity with results of IgM class immunoblot. This kit of the 3rd generation is reliable for screening of IgM antibodies. PMID- 15655724 TI - [Chemical analysis of bronchoalveolar lavage in diagnostics of pneumonia]. AB - Samples of bronchoalveolar lavages, obtained from 182 patients hospitalised at Vseobecna fakultni nemocnice and Nemocnice Kralovske Vinohrady, Prague, were analysed using solid phase microextraction-gas chromatography. At least 9 %, using less strong criteria up to 20 % of samples, contained higher volatile fatty acids (VFA), indicating the presence of fermenting anaerobic bacteria, found neither by cultivation nor by microscopy. Further compounds appeared in most of chromatographic examinations, tentatively labelled as "alcohol" and "acetoin". The second one occurred almost solely in samples from patients receiving the intensive care including mechanical ventilation. If characterised more precisely, it may serve as a potential marker of patients' status. PMID- 15655725 TI - [Detection of a mutant HSV-1 in a patient presenting efflorescence on an eye lid]. AB - A 9-month-old boy with impetigious blepharitis of the right eye was admitted to the Department of children and adolescents of the Kralovske Vinohrady University Hospital. NA isolated from lesion fluid was screened using real-time PCR based HSV 1/2 detection kit (Roche). Differentiation between HSV types 1 and 2 in this assay is based on variation of double-stranded DNA melting temperature (Tm), being 54 degrees C +/- 2,5 degrees C for HSV-1 and 66,5 degrees C +/- 2,5 degrees C for HSV-2. Since the amplified NA from the sample showed an atypical Tm of 59 degrees C, nested PCR was used for futher analysis: the presence of HSV-1 NA was confirmed by the latter. In spite of the atypical Tm value, the eyelid infection was responsive to acyclovir and resolved after intravenous Herpesin given for 5 days. PMID- 15655733 TI - Is routine testing mandatory or voluntary? PMID- 15655734 TI - Facial lipohypertrophy in HIV-infected subjects who underwent autologous fat tissue transplantation. AB - Of 41 HIV-infected patients with facial lipoatrophy who underwent autologous fat transplantation, disfiguring facial lipohypertrophy at the graft site occurred at the same time as recurrent fat accumulation at the tissue harvest site in 4 patients who had had fat transferred from the dorsocervical fat pad or from subcutaneous abdominal tissue. PMID- 15655735 TI - Seroprevalence of anti-H5 antibody among Thai health care workers after exposure to avian influenza (H5N1) in a tertiary care center. AB - After the initial atypical presentation of a patient with avian influenza (H5N1) infection, paired acute-phase and convalescent-phase serum samples obtained from 25 health care workers (HCWs) who were exposed to the patient were compared with paired serum samples obtained from 24 HCWs who worked at different units in the same hospital and were not exposed to the patient. There was no serologic evidence of anti-H5 antibody reactivity or subclinical infection in either of the groups. PMID- 15655736 TI - Differential interferon- gamma production characterizes the cytokine responses to Leishmania and Mycobacterium leprae antigens in concomitant mucocutaneous leishmaniasis and lepromatous leprosy. AB - BACKGROUND: Tegumentary leishmaniasis and leprosy display similar spectra of disease phenotypes, which are dependent on cell-mediated immunity to specific antigens. Diffuse cutaneous leishmaniasis and lepromatous leprosy represent the anergic end of the spectrum, whereas mucocutaneous leishmaniasis and tuberculoid leprosy are associated with marked antigen-specific cellular immune response. METHODS: We characterized and compared the cell-mediated response to Leishmania and Mycobacterium leprae antigens in a patient with an intriguing association of mucocutaneous leishmaniasis with lepromatous leprosy, which are at opposite ends of the immunopathological spectra of these diseases. This was done by performance of skin tests and by assessment of the cell proliferation and cytokine production of peripheral blood mononuclear cells (PBMCs). RESULTS: Strong skin-test reactions and PBMC proliferation were observed in response to Leishmania antigens but not to M. leprae antigens. The stimulation of PBMCs with Leishmania and M. leprae antigens induced comparable levels of tumor necrosis factor- alpha , interleukin-5, and interleukin-10. However, the interferon- gamma response to Leishmania antigens was remarkably high, and that to M. leprae antigens was almost nil. CONCLUSIONS: We found that concomitant leprosy and tegumentary leishmaniasis can produce opposite polar forms associated, respectively, with absent or exaggerated cell-mediated immune responses to each pathogen. This suggests that independent mechanisms influence the clinical outcome of each infection. Moreover, interferon- gamma appears to play a major role in the clinical expression of these intracellular infections. PMID- 15655737 TI - Influence of bacille Calmette-Guerin vaccination on size of tuberculin skin test reaction: to what size? AB - BACKGROUND: Previous bacillus Calmette-Guerin (BCG) vaccination can confound the results of a tuberculin skin test (TST). We sought to determine a cutoff diameter of TST induration beyond which the influence of BCG vaccination was negligible in evaluating potential Mycobacterium tuberculosis infection in a population of health care workers with a high vaccination rate and low incidence of tuberculosis. METHODS: From 1991 through 1998, all new employees at the University Hospital of Lausanne, Switzerland, underwent a 2-step TST at entry visit. We also gathered information on demographic characteristics, along with factors commonly associated with tuberculin positivity, including previous BCG vaccination, history of latent M. tuberculosis infection, and predictors for M. tuberculosis infection. RESULTS: Among the 5117 investigated subjects, we found that influence of BCG vaccination on TST results varied across categories of age (likelihood ratio test, 0.0001). Prior BCG vaccination had a strong influence on skin test results of or=1 events of LOS (totalling 3462 events). Logistic regression models were used to calculate the crude and adjusted risk for early mortality. RESULTS: Early mortality was associated with 179 LOS events (5.2% of 3,462); the range of pathogens associated with these events included coagulase-negative staphylococci (CoNS), which were the cause of 1.8% of LOS events associated with early mortality, and Pseudomonas species, which were the cause of 22.6% of such events. Early mortality after LOS, adjusted for neonatal risk factors, was significantly associated with sepsis due to certain pathogens: Pseudomonas species (odds ratio [OR], 12.3); Klebsiella species (OR, 6.3); Serratia species (OR, 6.2); Escherichia species (OR, 4.3); Enterobacter species (OR, 4.1); and Candida species (OR, 3.2), compared with sepsis due to CoNS . In addition, lower gestational age, lower chronological age, small size for gestational age, and grade 3-4 intraventricular hemorrhage, each had an independent association with early mortality. CONCLUSIONS: Klebsiella sepsis and Pseudomonas sepsis were associated with a 6.3-fold and 12.3-fold increased risk of early mortality, respectively, and accounted for 41.9% of all early deaths associated with LOS. Considering the aggressive nature of sepsis caused by these pathogens, empiric antibiotic therapy active against these organisms is worth consideration for VLBW infants with presumed LOS. PMID- 15655739 TI - The molecular epidemiology of Streptococcus pneumoniae with quinolone resistance mutations. AB - BACKGROUND: The purpose of this study was to determine the prevalence of fluoroquinolone resistance and quinolone resistance-determining region (QRDR) mutations among Streptococcus pneumoniae isolates in the United States during the period of 2001-2002. A second objective was to examine the genetic relatedness of pneumococcal isolates with parC and/or gyrA mutations during the period of 1994 2002. METHODS: Susceptibility testing was performed for 1902 S. pneumoniae isolates collected in the United States during the period of 2001-2002. On the basis of the minimum inhibitory concentration (MIC) of ciprofloxacin, 146 isolates were selected from the 2001-2002 study for QRDR analysis of parC, parE, gyrA, and gyrB genes. The genetic relatedness of isolates with parC and/or gyrA mutations from 2001-2002 (n=55) and from 3 US surveillance studies conducted during 1994-2000 (n=56) was determined by pulsed-field gel electrophoresis (PFGE). RESULTS: Between 1999-2000 and 2001-2002, there was a 2-fold increase in the rate of ciprofloxacin resistance (MIC, >or=4 micro g/mL), from 1.2% to 2.7%, and in the rate of levofloxacin nonsusceptibility (MIC, >or=4 micro g/mL), from 0.6% to 1.3%. The 111 isolates with parC and/or gyrA mutations were assigned to 48 different PFGE types. Forty-four isolates (40%) belonged to 8 PFGE types that were closely related to widespread clones. Fifteen of the 43 levofloxacin nonsusceptible pneumococci (LNSP) belonged to 4 PFGE types that were closely related to major clones (Spain(23F)-1 [n=6]; Spain(6B)-2 [n=5], Taiwan(19F)-14 [n=2], and Tennessee(23F)-4 [n=2]). CONCLUSION: The population of fluoroquinolone resistant S. pneumoniae in the United States has increased but remains genetically diverse. However, 35% of LNSP were related to widespread pneumococcal clones, increasing the potential for the rapid spread of quinolone resistance in this species. PMID- 15655740 TI - Fluoroquinolone-resistant pneumococci: maybe resistance isn't futile? PMID- 15655741 TI - A European Organization for Research and Treatment of Cancer-International Antimicrobial Therapy Group Study of secondary infections in febrile, neutropenic patients with cancer. AB - BACKGROUND: Neutropenic patients with cancer may develop several episodes of fever and infection during chemotherapy-induced myeloaplasia. METHODS: To identify risk factors for secondary infectious episodes among patients who responded to initial antibiotic therapy, we retrospectively analyzed 2 consecutive, prospective, randomized clinical trials performed by the International Antimicrobial Therapy Group of the European Organization for Research and Treatment of Cancer during 1991-1994. RESULTS: Of 1720 patients with their first episode of febrile neutropenia, 836 responded to the initial antibiotic regimen and were therefore suitable for our analysis. A secondary infection was observed in 129 (15%) of 836 patients that occurred at a median of 10 days (range, 1-28 days) after the onset of the primary febrile episode. Factors at both baseline and day 4 were analyzed. Age of >16 years (odds ratio [OR], 3.46; P<.001), acute leukemia in first induction (OR, 3.17; P<.001), presence of intravenous line (OR, 1.88; P=.04), severe neutropenia (defined as an absolute granulocyte count of <100 cells/mm(3)) on day 4 (OR, 2.72; P<.001), and type of documentation of the primary episode (i.e., microbiologically documented cause or unexplained fever; OR, 2.56; P=.001) were found to be risk factors for secondary infection. The risk of death was higher among patients who developed a secondary infectious episode than among those who did not (5.4% vs. 1.4%; P<.01). CONCLUSIONS: The clinical parameters described above may help to identify neutropenic patients at risk of developing secondary infection. PMID- 15655742 TI - Comparing interferon- gamma release assay with tuberculin skin test readings at 48-72 hours and 144-168 hours with use of 2 commercial reagents. AB - BACKGROUND: Despite widespread use, the tuberculin skin test (TST) has many limitations, including a requirement for a second visit between 48 and 72 hours. The goal of this study was to determine the reliability of a TST reading between 144 and 168 hours. METHODS: Tuberculin antigen was applied into both forearms (Aplisol in one arm and Tubersol in the other, from single lots of each product) by the Mantoux method. Blood samples were obtained for interferon- gamma release assay. Subjects were seen at 48-72 hours for the initial (day 2) TST reading and returned at 144-168 hours for a second (day 7) reading. RESULTS: A total of 116 subjects at increased risk for tuberculosis were studied; 25 (22%) had positive results at day 2 with Tubersol and 27 (23%) had positive results at day 2 with Aplisol. Overall agreement between Tubersol and Aplisol at day 2 was 93% (kappa = 0.80) and at day 7 was 94% (kappa = 0.76). Overall agreement between day 2 and day 7 was 89% for Tubersol and 86% for Aplisol. Discordant results between day 2 and day 7 occurred mostly in persons with a history of bacille Calmette-Guerin vaccination. CONCLUSIONS: Subjects who fail to present at 48-72 hours for TST reading may still have a reliable TST reading at up to 168 hours. Aplisol and Tubersol reagents produce comparable results when compared with the interferon- gamma release assay. PMID- 15655743 TI - Possible animal origin of human-associated, multidrug-resistant, uropathogenic Escherichia coli. AB - BACKGROUND: The multistate occurrence of cases of urinary tract infection (UTI) caused by trimethoprim-sulfamethoxazole (TMP-SMZ)-resistant Escherichia coli strains belonging to a single clonal group (designated as clonal group A [CgA]) in the United States has raised an intriguing hypothesis that these infections may have been spread by contaminated food products. The present study attempted to determine if CgA strains could be traced to food animals. METHODS: A total of 495 animal and environmental E. coli isolates, which belonged to serogroups O11, O17, O73, and O77 and were collected between 1965 and 2002 by the Gastroenteric Disease Center at Pennsylvania State University (University Park, PA), were further subtyped by antimicrobial drug susceptibility, enterobacterial repetitive intergenic consensus (ERIC2) PCR, random amplified polymorphic DNA analysis, pulsed-field gel electrophoresis (PFGE), and virulence profile pattern. RESULTS: Of 495 isolates, 128 (26%) had an ERIC2 PCR electrophoretic pattern indistinguishable from that of the human prototype CgA strain, and 14 CgA isolates were resistant to TMP-SMZ. Cluster analysis of PFGE patterns showed that 1 of these 14 isolates, obtained from a cow in 1988, was 94% similar to a CgA uropathogenic human-associated E. coli strain. The pattern for this isolate was included among a cluster of PFGE patterns for 5 human-associated UTI isolates that were >80% similar to each other. CONCLUSIONS: These observations suggest that drug-resistant, uropathogenic human-associated E. coli strains potentially have an animal origin. The possibility that human drug-resistant UTI could be a foodborne illness has serious public health implications. PMID- 15655744 TI - Is acute uncomplicated urinary tract infection a foodborne illness, and are animals the source? PMID- 15655745 TI - Comparison between LightCycler Real-Time Polymerase Chain Reaction (PCR) assay with serum and PCR-enzyme-linked immunosorbent assay with whole blood samples for the diagnosis of human brucellosis. AB - BACKGROUND: To overcome some of the limitations of conventional microbiological techniques, polymerase chain reaction (PCR)-based assays have been proposed as a useful tool for the diagnosis of human brucellosis. METHODS: A single-blinded comparative study was undertaken that compared 2 different PCR assays: a SYBR Green I LightCycler-based Real-Time PCR assay (LC-PCR; Roche Diagnostic) with serum samples and a PCR-enzyme-linked immunosorbent assay (ELISA) with whole blood samples. Both assays amplify a 223-bp sequence of a gene that codes for the synthesis of an immunogenetic membrane protein specific for Brucella genus (BCSP31). We analyzed the diagnostic yield of these assays with 60 samples obtained from patients with active brucellosis and 37 samples obtained from a control group composed of patients with febrile syndromes of other defined etiologies, asymptomatic subjects with past brucellosis or exposure to Brucella infection who had persistently high titers of anti-Brucella antibodies, and healthy subjects. RESULTS: The sensitivities of LC-PCR with serum samples, PCR ELISA with whole blood samples, and blood cultures were 93.3%, 90%, and 65%, respectively. Three control samples (8.1%) had a positive PCR-ELISA result, and 2 of these samples (5.4%) also had positive LC-PCR results. The specificity and positive likelihood ratios were 94.6% and 17.3, respectively, for LC-PCR and 91.9% and 11.1, respectively, for PCR-ELISA. CONCLUSIONS: The diagnostic yield of LC-PCR with serum samples was higher than that of PCR-ELISA with whole blood samples. The speed and technical simplicity of LC-PCR in serum samples make it a useful alternative to blood cultures for patients with suspected brucellosis and negative or doubtful serological test results. PMID- 15655746 TI - A hospital outbreak of Clostridium difficile disease associated with isolates carrying binary toxin genes. AB - INTRODUCTION: The binary toxin genes cdt and cdtB have been detected in approximately 5% of Clostridium difficile strains. Severe C. difficile disease (CDD) may be associated with strains that carry the binary toxin genes. METHODS: From April 2001 through March 2002, 8 severe and 41 nonsevere cases of nosocomial CDD were studied. Severe cases of CDD were defined by the presence of >or=2 of the following criteria: (1) abdominal pain, (2) a white blood cell count of >20,000 or <1500 cells/mm(3), and (3) ileus or bowel wall thickening with ascites. Underlying disease was assessed by 2 methods: a modified Horn score and the presence of comorbid conditions. The presence of cdtA, cdtB, and the toxin A and toxin B genes was determined, and molecular subtyping was performed. RESULTS: All strains were positive for the toxin A and B genes, and 65.3% of the strains carried the cdtA and cdtB genes. Strains that carried the binary toxin genes accounted for 87.5% of the cases of severe CDD and 61.0% of the nonsevere cases (P=.23). Severity of CDD was not associated with either severe underlying disease or comorbid conditions. The strains that caused severe CDD belonged to 4 protein profile groups and >or=3 restriction endonuclease analysis (REA) groups. All (i.e., 5 of 5) strains in REA group BI, compared with none (i.e., 0 of 7) of the strains in REA group J carried the binary toxin genes (P=.001). Strains that belonged to REA groups BK and BR also carried the binary toxin genes. CONCLUSIONS: The binary toxin genes were present in nearly two-thirds of the C. difficile strains, and they were correlated with the REA group. Severity of CDD was not closely associated with a specific clone or underlying disease, but it may be associated with the presence of the binary toxin genes. Larger studies are needed to discern whether a true association exists and whether the binary toxin alters the pathogenicity of the C. difficile strain. PMID- 15655748 TI - Inducible clindamycin resistance in Staphylococci: should clinicians and microbiologists be concerned? AB - The increasing incidence of a variety of infections due to Staphylococcus aureus- and, especially, the expanding role of community-associated methicillin-resistant S. aureus (MRSA)--has led to emphasis on the need for safe and effective agents to treat both systemic and localized staphylococcal infections. Unlike most previously noted strains of health care-associated MRSA, community-acquired MRSA isolates are often susceptible to several non- beta -lactam drug classes, although they are usually not susceptible to macrolides. Several newer antimicrobial agents and a few older agents are available for treatment of systemic staphylococcal infections, but use may be limited by the relatively high cost of these agents or the need for parenteral administration. Inexpensive oral agents for treatment of localized, community-acquired MRSA infection include clindamycin, trimethoprim-sulfamethoxazole, and newer tetracyclines. Clindamycin has been used successfully to treat pneumonia and soft-tissue and musculoskeletal infections due to MRSA in adults and children. However, concern over the possibility of emergence of clindamycin resistance during therapy has discouraged some clinicians from prescribing that agent. Simple laboratory testing (e.g., the erythromycin-clindamycin "D-zone" test) can separate strains that have the genetic potential (i.e., the presence of erm genes) to become resistant during therapy from strains that are fully susceptible to clindamycin. PMID- 15655747 TI - Quantitative T cell assay reflects infectious load of Mycobacterium tuberculosis in an endemic case contact model. AB - BACKGROUND: Currently, reliable efficacy markers for assessment of new interventions against tuberculosis (TB) are limited to disease and death. More precise measurement of the human immune response to Mycobacterium tuberculosis infection may be important. A qualitative enzyme-linked immunospot assay (ELISPOT) result for early secretory antigenic target 6 (ESAT-6) and culture filtrate protein 10 (CFP-10) offers improved specificity over the purified protein derivative (PPD) skin test reaction in the detection of M. tuberculosis infection. We evaluated the quantitative ELISPOT and PPD skin test responses to recent M. tuberculosis exposure. METHODS: We studied quantitative PPD skin test and PPD ELISPOT results in 1052 healthy household contacts of index patients with cases of sputum smear-positive and culture-positive TB in The Gambia, according to a positive or negative ex vivo interferon gamma ELISPOT response to M. tuberculosis-specific antigens (ESAT-6/CFP-10). We then studied the quantitative PPD skin test and PPD ELISPOT results in patient contacts who had positive ESAT 6/CFP-10 results against a natural exposure gradient according to sleeping proximity to a patient with TB. RESULTS: The number of positive results was significantly greater for both PPD skin test and PPD ELISPOT in ESAT-6/CFP-10 positive subjects, compared with others (P<.0001). However, when quantitative PPD skin test and PPD ELISPOT results were compared in ESAT-6/CFP-10-positive subjects, only the ELISPOT count was sensitive to the exposure gradient, increasing significantly according to exposure (P=.009). CONCLUSIONS: The quantitative ELISPOT response to PPD in specific-antigen-positive contacts of patients with TB reflects the infectious load of M. tuberculosis as a result of recent exposure. This finding offers new possibilities for assessment of the efficacy of new interventions, and adjustment should be made for it when relating the early immune response to progression to disease. PMID- 15655749 TI - International adoption: infectious diseases issues. AB - Nearly 220,000 children have been adopted from other countries by American parents since 1986. Approximately 65,000 children have arrived from China and Russia, mostly in the past 6 years. Most of these children reside in orphanages before adoption, where they may experience malnutrition, environmental deprivation, neglect, and exposure to infectious diseases. After arrival to the United States, international adoptees should undergo specialized screening evaluation for infectious diseases and other conditions. Infectious conditions of special concern include hepatitis B and C, syphilis, human immunodeficiency virus infection, tuberculosis, and presence of intestinal parasites. Before the adoption occurs, the infectious disease consultant may be asked to assist the primary care provider and the adoptive family with advice about travel and review of preadoptive medical records. After the adoption, the infectious diseases consultant may be asked to assess the adequacy of the child's vaccination record from the birth country and to assist in screening, evaluation, and management of infectious diseases. PMID- 15655750 TI - Drug-resistant HIV infection among drug-naive patients in Israel. AB - BACKGROUND: In Israel, <0.06% of the general population is infected with human immunodeficiency virus (HIV), with a much higher prevalence among specific groups. These groups are distinguished demographically by risk behavior category and by virus subtype. We investigated transmission of drug resistance within groups to assess the impact of these factors. METHODS: Plasma samples from >15% of all patients with new diagnoses of HIV infection were randomly collected between June 1999 and June 2003. Sequences from 176 drug-naive patients included 20 of subtype A, 20 of subtype AE, 2 of subtype AC, 29 of subtype B, 100 of subtype C, and 5 of subtype F. RESULTS: Major drug resistance mutations (protease: L90M; reverse transcriptase: M41L, K103N, V106M, M184V, Y181S, G190A, L210W, T215Y/F, and K219R) were detected in 1 subject with A subtype, 3 with subtype B, and 9 with subtype C. In addition, 1 subject with A subtypes, 2 with subtype B, and 10 with subtype C had secondary mutations (protease: M46I; reverse transcriptase: A98G, K101Q, and V108I). Only 1 patient had mutations associated with >1 class of drugs. Among subjects who contracted HIV infection in Israel, 16 of 56 (1 of 7 with subtypes A or AE, 4 of 17 with subtype B, and 11 of 32 with subtype C; P=.7-1.0) carried resistant virus--a significantly higher proportion (P<.001) than in subjects infected in other countries (10 of 120 infected). CONCLUSIONS: Drug-resistant virus was detected in 14.8% of patients with new diagnoses of HIV infection but in 28.6% of patients known to have been infected in Israel. The implications include a need for pretreatment resistance testing and for better programs aimed at prevention of transmission, directed particularly at patients. We did not find significant differences in transmission of resistant virus between those infected with subtypes B and C, despite the different demographic background. A conclusive analysis and interpretation should await a more extensive study. PMID- 15655751 TI - Tolerability of postexposure prophylaxis with the combination of zidovudine lamivudine and lopinavir-ritonavir for HIV infection. AB - Tolerability of the combination of zidovudine-lamivudine and lopinavir-ritonavir as postexposure prophylaxis (PEP) for human immunodeficiency virus infection was prospectively assessed. A total of 121 patients were enrolled in the study; 23 patients discontinued PEP prematurely for reasons other than adverse events. Of the other 98 patients, 58 (59%) experienced adverse effects, which led to premature PEP discontinuation in 20 cases (20%). PMID- 15655752 TI - Prevalence of mixed cryoglobulins in relation to CD4 cell count among patients coinfected with HIV and hepatitis C virus. AB - Cryoglobulinemia was studied in human immunodeficiency virus-positive, hepatitis C virus (HCV)-positive patients in relation to their CD4 cell count. Cryoglobulinemia was found in 18 (31.6%) of 57 patients: 17 (44.7%) of 38 patients with a CD4 cell count of >or=200 cells/ micro L versus 1 (5.3%) of 19 patients with a CD4 cell count of <200 cells/ micro L (P=.0064). Cell-mediated immunity could, therefore, contribute to the production of HCV-associated cryoglobulins. PMID- 15655753 TI - Resistance to dual nucleoside reverse-transcriptase inhibitors in children infected with HIV clade A/E. AB - The prevalence of nucleoside reverse-transcriptase inhibitor (NRTI) mutations was determined among 95 human immunodeficiency virus-infected Thai children who were treated with dual nucleoside reverse-transcriptase inhibitors. Almost all children had resistance to at least 1 NRTI, and approximately half of the children had resistance to multiple NRTIs. Cross-resistance to stavudine and azidothymidine was universal. PMID- 15655754 TI - A new therapeutic challenge for old pathogens: community-acquired invasive infections caused by ceftriaxone- and ciprofloxacin-resistant Salmonella enterica serotype choleraesuis. AB - Recently, antimicrobial resistance among nontyphoid Salmonella serotypes has been increasingly recognized. In southern Taiwan, we encountered 3 cases of invasive infections caused by Salmonella enterica serotype Choleraesuis with resistance to ciprofloxacin and ceftriaxone. Resistance to ciprofloxacin was related to nucleotide mutations in gyrA and parC, and resistance to ceftriaxone was related to the presence of CMY-2 beta -lactamase. PMID- 15655756 TI - Does counseling increase sustained benefit of HAART among prison inmates after release to the community? PMID- 15655758 TI - Sonographic assessment of lipodystrophy in HIV-infected patients: some open questions. PMID- 15655760 TI - Classical diphtheria caused by Corynebacterium ulcerans in Germany: amino acid sequence differences between diphtheria toxins from Corynebacterium diphtheriae and C. ulcerans. PMID- 15655763 TI - Coinfection with HIV and human T lymphotropic virus type 1: what is the real impact on HIV disease? PMID- 15655761 TI - Influence of GB virus type C and HIV coinfection on gamma delta T cells. PMID- 15655765 TI - Fatal Salmonella pulmonary arteritis in a patient with Eisenmenger syndrome. PMID- 15655766 TI - False-positive results of Aspergillus enzyme-linked immunosorbent assays for a patient with gastrointestinal graft-versus-host disease taking a nutrient containing soybean protein. PMID- 15655767 TI - Background for different use of antibiotics in different countries. PMID- 15655768 TI - Treating opportunistic infections among HIV-exposed and infected children: recommendations from CDC, the National Institutes of Health, and the Infectious Diseases Society of America. AB - In 2001, CDC, the National Institutes of Health, and the Infectious Diseases Society of America convened a working group to develop guidelines for therapy of human immunodeficiency virus (HIV)-associated opportunistic infections to serve as a companion to the Guidelines for Prevention of Opportunistic Infections Among HIV-Infected Persons. In recognition of unique considerations related to HIV infection among infants, children, and adolescents, a separate pediatric working group was established. Because HIV-infected women coinfected with opportunistic pathogens might be more likely to transmit these infections to their infants than women without HIV infection, guidelines for treating opportunistic pathogens among children should consider treatment of congenitally acquired infections among both HIV-exposed but uninfected children and those with HIV infection. In addition, the natural history of opportunistic infections among HIV-infected children might differ from that among adults. Compared with opportunistic infections among HIV-infected adults, which are often caused by reactivation of pathogens acquired before HIV infection when host immunity was intact, opportunistic infections among children often reflect primary acquisition of the pathogen and, among children with perinatal HIV infection, infection acquired after HIV infection has been established and begun to compromise an already immature immune system. Laboratory diagnosis of opportunistic infections can be more difficult with children. Finally, treatment recommendations should consider differences between adults and children in terms of drug pharmacokinetics, dosing, formulations, administration, and toxicities. This report focuses on treatment of opportunistic infections that are common in HIV-exposed and infected infants, children, and adolescents in the United States. PMID- 15655769 TI - Coronaviruses in the limelight. PMID- 15655770 TI - Evidence of a novel human coronavirus that is associated with respiratory tract disease in infants and young children. AB - BACKGROUND: The etiological agents responsible for a substantial proportion of respiratory tract diseases have not been identified. We sought to determine whether novel human coronaviruses (HCoVs) are circulating in New Haven, Connecticut, and, if so, whether they are associated with respiratory tract disease in infants and young children. METHODS: We developed a polymerase chain reaction (PCR)-based approach for screening specimens from the respiratory tracts of symptomatic children. PCR probes that target regions of the replicase 1a gene that are conserved among genetically diverse animal CoVs and HCoVs were designed. Using these probes, we identified genomic sequences of a novel HCoV, designated "New Haven coronavirus" (HCoV-NH). Thereafter, we designed specific probes to screen respiratory specimens from children <5 years old for this novel HCoV. Clinical features associated with HCoV-NH infection were identified. RESULTS: Seventy-nine (8.8%) of 895 children tested positive for HCoV-NH. Cough, rhinorrhea, tachypnea, fever, abnormal breath sounds, and hypoxia were the most common findings associated with HCoV-NH infection. Sequence analysis revealed that HCoV-NH is closely related to a novel HCoV recently reported in The Netherlands. CONCLUSIONS: The novel HCoVs identified in New Haven and The Netherlands are similar and likely represent the same species. This newly discovered virus may have worldwide distribution and may account for a significant proportion of respiratory tract disease in infants and young children. PMID- 15655771 TI - Association between a novel human coronavirus and Kawasaki disease. AB - Kawasaki disease is a systemic vasculitis of childhood; its etiology is unknown. We identified evidence of a novel human coronavirus, designated "New Haven coronavirus" (HCoV-NH), in respiratory secretions from a 6-month-old infant with classic Kawasaki disease. To further investigate the possible association between HCoV-NH infection and Kawasaki disease, we conducted a case-control study. Specimens of respiratory secretions from 8 (72.7%) of 11 children with Kawasaki disease and from 1 (4.5%) of 22 control subjects (children without Kawasaki disease matched by age and the time the specimens were obtained) tested positive for HCoV-NH by reverse-transcriptase polymerase chain reaction (Mantel-Haenszel matched odds ratio, 16.0 [95% confidence interval, 3.4-74.4]; P=.0015). These data suggest that HCoV-NH infection is associated with Kawasaki disease. PMID- 15655772 TI - Human coronavirus NL63 infection in Canada. AB - The isolation of human coronavirus NL63 (HCoV-NL63) in The Netherlands raised questions about its contribution to respiratory illness. In this study, a total of 525 respiratory specimens, collected in Canada primarily during the winter months of 2001-2002, were tested for HCoV-NL63; 19 tested positive for HCoV-NL63, demonstrating virus activity during January-March 2002. Patients with HCoV-NL63 were 1 month-100 years old (median age, 37 years). The main clinical presentations were fever (15/19), sore throat (5/19), and cough (9/19), and 4 patients were hospitalized. These results provide evidence for the worldwide distribution of HCoV-NL63. PMID- 15655773 TI - Development and characterization of a severe acute respiratory syndrome associated coronavirus-neutralizing human monoclonal antibody that provides effective immunoprophylaxis in mice. AB - BACKGROUND: Severe acute respiratory syndrome (SARS) remains a significant public health concern after the epidemic in 2003. Human monoclonal antibodies (MAbs) that neutralize SARS-associated coronavirus (SARS-CoV) could provide protection for exposed individuals. METHODS: Transgenic mice with human immunoglobulin genes were immunized with the recombinant major surface (S) glycoprotein ectodomain of SARS-CoV. Epitopes of 2 neutralizing MAbs derived from these mice were mapped and evaluated in a murine model of SARS-CoV infection. RESULTS: Both MAbs bound to S glycoprotein expressed on transfected cells but differed in their ability to block binding of S glycoprotein to Vero E6 cells. Immunoprecipitation analysis revealed 2 antibody-binding epitopes: one MAb (201) bound within the receptor binding domain at aa 490-510, and the other MAb (68) bound externally to the domain at aa 130-150. Mice that received 40 mg/kg of either MAb prior to challenge with SARS-CoV were completely protected from virus replication in the lungs, and doses as low as 1.6 mg/kg offered significant protection. CONCLUSIONS: Two neutralizing epitopes were defined for MAbs to SARS-CoV S glycoprotein. Antibodies to both epitopes protected mice against SARS-CoV challenge. Clinical trials are planned to test MAb 201, a fully human MAb specific for the epitope within the receptor-binding region. PMID- 15655774 TI - Human leukocyte antigen class II alleles and rubella-specific humoral and cell mediated immunity following measles-mumps-rubella-II vaccination. AB - We examined associations between human leukocyte antigen (HLA) class II genes and both rubella-specific immunoglobulin G antibodies and lymphoproliferative responses after measles-mumps-rubella-II (MMR-II) vaccination, in a population based sample of 346 schoolchildren. The DPB1*0301, DPB1*0401, DPB1*1301, and DPB1*1501 alleles were significantly associated with rubella vaccine-induced antibodies. Alleles suggestive of being positively associated with rubella specific lymphoproliferative stimulation indices were DPB1*0301, DQB1*0501, DRB1*0101, and DRB1*1104. Conversely, the DPB1*0401, DPB1*1001, DPB1*1101, DQB1*0202, and DRB1*0701 alleles were negatively associated with rubella-specific lymphoproliferation. This study of HLA class II-restricted humoral and cellular immune responses to rubella provides significant insight into mechanisms of vaccine response and new vaccine development. PMID- 15655775 TI - Immune reconstitution and clearance of human adenovirus viremia in pediatric stem cell recipients. AB - BACKGROUND: Human adenovirus (HAdV) infections are increasingly frequent complications of allogeneic stem-cell transplantation (SCT), especially in children. Only a few data on the correlation between immune recovery and the course of HAdV infection are available, and data on HAdV-specific responses are lacking. METHODS: In a prospective study, we determined the correlation between the HAdV DNA load in plasma and lymphocyte reconstitution in 48 children after allogeneic SCT. Additionally, HAdV-specific humoral and cellular immune responses were investigated. RESULTS: HAdV infection occurred in 21 patients (44%), and, in 6 of these patients, the infection progressed to viremia, as demonstrated by the presence of HAdV DNA in plasma. Low lymphocyte counts at the onset of infection were predictive of HAdV viremia. Survival of patients with HAdV viremia was associated with an increase in lymphocyte counts during the first weeks after infection. In these patients, HAdV-specific CD4+ T cell responses, as well as increases in titers of neutralizing antibody, were detected after clearance of HAdV DNA from plasma. CONCLUSIONS: Lymphocyte reconstitution appears to play a crucial role in clearance of HAdV viremia and survival of the host, warranting further development of therapeutic interventions aimed at improving immune recovery. PMID- 15655776 TI - Differences between T cell-type and natural killer cell-type chronic active Epstein-Barr virus infection. AB - Infections of T cells and natural killer (NK) cells play a central role in the pathogenesis of chronic active Epstein-Barr virus (CAEBV) infection. To characterize the virologic and cytokine profiles of T cell-type and NK cell-type infection, 39 patients with CAEBV infection were analyzed. Patients with T cell type infection had higher titers of immunoglobulin G against early and late EBV antigens, suggesting lytic cycle infection. However, the pattern of EBV gene expression was latency type II; BZLF1, which is a hallmark of lytic cycle infection, could not be detected in any patients, regardless of infection type. Patients with CAEBV infection had high concentrations of proinflammatory, T helper cell type 1, and anti-inflammatory cytokines. The cytokine profile in patients with NK cell-type infection was similar to that in patients with T cell type infection, but the concentration of IL-13 was high in patients with NK cell type infection. These findings should help to clarify the pathogenesis of CAEBV infection and facilitate the development of more-effective treatments. PMID- 15655777 TI - Distribution of varicella-zoster virus DNA and gene products in tissues of a first-trimester varicella-infected fetus. AB - Precise information about varicella-zoster virus (VZV) infection in first trimester fetuses remains sketchy. After varicella infection was diagnosed in a woman, her 12-week-old fetus was aborted and was investigated, by histological examination, virus culturing, polymerase chain reaction, in situ hybridization (ISH), and immunohistochemistry (IHC), for the presence of VZV infection. Only the results of the histological examination suggested the presence of alpha herpesvirus infection, in the gastrointestinal tract and liver; results of ISH were positive for VZV, and results of IHC staining were positive for intermediate early protein 63 (IE63) but negative for glycoprotein E (gE), in the dorsal root ganglia (DRG), meninges, gastrointestinal tract, pancreas, smooth muscle, liver, and placental trophoblast, indicating the presence of a nonproductive, latency like VZV infection. Only the gastrointestinal tract and liver exhibited simultaneous staining for IE63 and gE, a result suggesting that active replication of VZV was present. In conclusion, widespread nonproductive VZV infection in the absence of histological clues is an early event in VZV infection in fetuses. The observed gene-expression pattern in most tissues resembles that of latent VZV infection in DRG. Latency-like infection in nonneural cell types may potentially reactivate, leading to multifocal necrosis, fibrosis, and dystrophic calcifications, as observed in advanced congenital varicella syndrome. PMID- 15655778 TI - Female-to-male infectivity of HIV-1 among circumcised and uncircumcised Kenyan men. AB - BACKGROUND: A lack of male circumcision has been associated with increased risk of human immunodeficiency virus type 1 (HIV-1) acquisition in a number of studies, but questions remain as to whether confounding by behavioral practices explains these results. The objective of the present study was to model per-sex act probabilities of female-to-male HIV-1 transmission (i.e., infectivity) for circumcised and uncircumcised men, by use of detailed accounts of sexual behavior in a population with multiple partnerships. METHODS: Data were collected as part of a prospective cohort study of HIV-1 acquisition among 745 Kenyan truck drivers. Sexual behavior with wives, casual partners, and prostitutes was recorded at quarterly follow-up visits. Published HIV-1 seroprevalence estimates among Kenyan women were used to model HIV-1 per-sex act transmission probabilities. RESULTS: The overall probability of HIV-1 acquisition per sex act was 0.0063 (95% confidence interval, 0.0035-0.0091). Female-to-male infectivity was significantly higher for uncircumcised men than for circumcised men (0.0128 vs. 0.0051; P=.04). The effect of circumcision was robust in subgroup analyses and across a wide range of HIV-1 prevalence estimates for sex partners. CONCLUSIONS: After accounting for sexual behavior, we found that uncircumcised men were at a >2-fold increased risk of acquiring HIV-1 per sex act, compared with circumcised men. Moreover, female-to-male infectivity of HIV-1 in the context of multiple partnerships may be considerably higher than that estimated from studies of HIV-1-serodiscordant couples. These results may explain the rapid spread of the HIV-1 epidemic in settings, found throughout much of Africa, in which multiple partnerships and a lack of male circumcision are common. PMID- 15655779 TI - Antimicrobial-resistant nontyphoidal Salmonella is associated with excess bloodstream infections and hospitalizations. AB - BACKGROUND: Nontyphoidal Salmonella is a leading cause of foodborne illness. Few studies have explored the health consequences of antimicrobial-resistant Salmonella. METHODS: The National Antimicrobial Resistance Monitoring System (NARMS) performs susceptibility testing on nontyphoidal Salmonella isolates. The Foodborne Diseases Active Surveillance Network (FoodNet) ascertains outcomes for patients with culture-confirmed Salmonella infection, in 9 states, each of which participates in NARMS. We analyzed the frequency of bloodstream infection and hospitalization among patients with resistant infections. Isolates defined as resistant to a clinically important agent were resistant to 1 or more of the following agents: ampicillin, ceftriaxone, ciprofloxacin, gentamicin, and/or trimethoprim-sulfamethoxazole. RESULTS: During 1996-2001, NARMS received 7370 serotyped, nontyphoidal Salmonella isolates from blood or stool. Bloodstream infection occurred more frequently among patients infected with an isolate resistant to > or =1 clinically important agent (adjusted odds ratio [OR], 1.6; 95% confidence interval [CI], 1.2-2.1), compared with patients with pansusceptible infection. During 1996-2001, FoodNet staff ascertained outcomes for 1415 patients who had isolates tested in NARMS. Hospitalization with bloodstream infection occurred more frequently among patients infected with an isolate resistant to > or =1 clinically important agent (adjusted OR, 3.1; 95% CI, 1.4-6.6), compared with patients with pansusceptible infection. CONCLUSIONS: Patients with antimicrobial-resistant nontyphoidal Salmonella infection were more likely to have bloodstream infection and to be hospitalized than were patients with pansusceptible infection. Mitigation of antimicrobial resistance in Salmonella will likely benefit human health. PMID- 15655780 TI - Serologic correlates of protection against enterotoxigenic Escherichia coli diarrhea. AB - BACKGROUND: We conducted a nested case-control study in 397 rural Egyptian children <36 months of age to assess the correlation between serum levels of antibodies against toxin and colonization factors (CFs) and the risk of homologous enterotoxigenic Escherichia coli (ETEC) diarrhea. METHODS: Active case detection was performed via semiweekly home visits, and blood was obtained at 3 month intervals. After each serosurvey, case subjects were selected from children experiencing a CF antigen (CFA)/I-, CFA/II-, CFA/IV-, or heat-labile enterotoxin (LT)-ETEC diarrheal episode during the subsequent 3 months. Up to 5 control subjects per case subject were selected from children who did not experience an ETEC diarrheal episode during the corresponding interval. Serum titers of immunoglobulin G antibodies against CFA/I, coli surface antigen (CS) 3, CS6, and LT were measured by enzyme-linked immunosorbant assay. RESULTS: The distribution of serum titers of LT, CS3, and CS6 antibodies did not differ between the case and control subjects. For children <18 months of age, serum titers of CFA/I antibody were inversely related to the risk of CFA/I-ETEC diarrhea; reciprocal serum titers of CFA/I antibody > or =76 were associated with a 77% reduction in the odds of CFA/I-ETEC diarrhea. CONCLUSION: Induction of reciprocal serum titers of antibodies against CFA/I within or above the 76-186 range should be further evaluated as a predictor for assessment of the ability of candidate vaccines to protect against CFA/I-ETEC diarrhea. PMID- 15655782 TI - Host adaptation and immune modulation are mediated by homologous recombination in Helicobacter pylori. AB - Rearrangement of genomic DNA via homologous recombination provides an alternative mechanism of gene regulation that is essential for successful colonization of the gastric mucosa by Helicobacter pylori. Inoculation of outbred mice with the H. pylori SS1 wild-type strain elicited a T helper (Th) 2 response and established a persistent infection. In contrast, inoculation with an isogenic H. pylori strain defective for homologous recombination elicited a Th1-mediated immune response and clearance of infection within 70 days. We, therefore, demonstrate that recombination is critical for mediating persistence of a microbial pathogen through the induction of ineffective immune responses. PMID- 15655781 TI - Expression of tumor necrosis factor- alpha -related apoptosis-inducing ligand and its proapoptotic receptors is down-regulated during gastric infection with virulent cagA+/vacAs1+ Helicobacter pylori strains. AB - BACKGROUND: Infection of the gastric mucosa with Helicobacter pylori leads to increased apoptosis. Cytokines and receptors of the tumor necrosis factor (TNF) family are known to be involved in this process. The role that the death-inducing TNF- alpha -related apoptosis-inducing ligand (TRAIL) and its receptors play, in the context of H. pylori infection, is unknown. METHODS: In 74 H. pylori-infected and 51 H. pylori-uninfected gastric antral biopsy specimens, levels of TRAIL mRNA and TRAIL receptor mRNA were determined quantitatively by TaqMan reverse transcriptase polymerase chain reaction. Recombinant TRAIL-induced apoptosis was measured in human and rat gastric epithelial cells by end-labeling of DNA with fluorescein-dTUP and by fluorescence-activated cell sorter analysis. RESULTS: In patients infected with cagA+/vacAs1+ H. pylori strains, expression of TRAIL and the proapoptotic receptors TRAIL-R1 and -R2 was down-regulated, whereas expression of the antiapoptotic receptors TRAIL-R3 and -R4 was up-regulated. Furthermore, expression of TRAIL and TRAIL-R1 and -R2 correlated inversely with the severity of gastric inflammation. Significant apoptosis of isolated human gastric epithelial cells and highly enriched rat parietal and chief cells was induced by 100 ng/mL TRAIL. CONCLUSIONS: Down-regulation of the TRAIL system, in the context of H. pylori infection, may limit exaggerated apoptosis of gastric epithelial cells and destruction of tissue and, therefore, may enable H. pylori to maintain its niche. PMID- 15655783 TI - Impact of vancomycin resistance on mortality among patients with neutropenia and enterococcal bloodstream infection. AB - We performed a retrospective cohort study to measure the impact of vancomycin resistance on clinical outcome for 83 episodes of enterococcal bloodstream infection (BSI; 22 with vancomycin-resistant enterococci [VRE] and 61 with vancomycin-susceptible enterococci [VSE]) in 77 patients with neutropenia. Cox proportional hazards models showed that vancomycin resistance was an independent predictor of mortality, after controlling for severity of illness, enterococcal species, gram-negative copathogens, sex, race, duration of neutropenia before bacteremia, and early administration of active antibiotics. This effect was evident only > or =10 days after the onset of bacteremia (P=.0263; hazard ratio [HR], 4.969) but not after adjustment for duration of bacteremia. The median duration of bacteremia was 4.5 days for VRE BSI and <1 day for VSE BSI (P=.0001). The only independent predictor of bacteremia duration was vancomycin resistance (P=.0284; HR, 3.863). Vancomycin resistance is associated with increased mortality in patients with neutropenia, possibly because of prolonged duration of bacteremia. PMID- 15655784 TI - Blockade of the Fas/FasL system improves pneumococcal clearance from the lungs without preventing dissemination of bacteria to the spleen. AB - BACKGROUND: The Fas/FasL system is both proapoptotic and proinflammatory. FasL is inhibited by decoy receptor-3 (DcR3), a naturally occurring decoy receptor. We determined the effects of systemic blockade of the Fas/FasL system by a DcR3 analog (DcR3-a) in mice with pneumococcal pneumonia. METHODS: Streptococcus pneumoniae (7.2 x 105 or 1.9 x 107 cfu/mL) was instilled intratracheally into untreated C57Bl/6 mice, C57Bl/6 mice treated with DcR3-a, or Fas-deficient lpr mice, and the mice were studied 48 h later. RESULTS: After instillation of the lower bacterial dose, disruption of the Fas/FasL system by either DcR3-a or the lpr mutation resulted in improved clearance of bacteria in the lungs (mean +/- SE, 4.6+/-2.1 x 10(6) and 3.5 +/- 1.6 x 10(6) cfu/lung, respectively, vs. 21.9+/ 9.3 x 10(6) cfu/lung in untreated C57Bl/6 mice; P<.05) and decreased percentage of polymorphonuclear neutrophils in bronchoalveolar lavage fluid (mean +/- SE, 19.3%+/-9.5% and 20.2%+/-7.8%, respectively, vs. 55.0%+/-12.2% in untreated C57Bl/6 mice; P<.05). These changes were associated with decreased lung concentrations of the proinflammatory cytokines tumor necrosis factor- alpha and macrophage inflammatory protein-2 and with a decrease in apoptotic cells in the alveolar walls. CONCLUSION: Blockade of the Fas/FasL system by DcR3-a in the lungs improves clearance of bacteria in mice with pneumococcal pneumonia. PMID- 15655785 TI - Bartonella quintana in a 4000-year-old human tooth. AB - Bacteria of the genus Bartonella are transmitted by ectoparasites (lice, fleas, ticks) and have mammalian reservoirs in which they cause chronic, asymptomatic bacteremia. Humans are the reservoir of B. quintana, the louse-borne agent of trench fever. We detected DNA of B. quintana in the dental pulp of a person who died 4000 years ago. PMID- 15655786 TI - Helminthic infection down-regulates type 1 immune responses in human T cell lymphotropic virus type 1 (HTLV-1) carriers and is more prevalent in HTLV-1 carriers than in patients with HTLV-1-associated myelopathy/tropical spastic paraparesis. AB - Human T cell lymphotropic virus type 1 (HTLV-1) infection is associated with an exacerbated type 1 immune response and secretion of high levels of proinflammatory cytokines. In contrast, helminthic infection induces a type 2 immune response. In the present study, the cytokine profile in HTLV-1 carriers coinfected with helminths (Strongyloides stercoralis and/or Schistosoma mansoni) was compared with that in HTLV-1 carriers not coinfected with helminths. Levels of interferon (IFN)- gamma were higher in HTLV-1 carriers not coinfected with helminths than in HTLV-1 carriers coinfected with helminths (P<.05). The overall frequency of IFN- gamma -expressing CD8+ and CD4+ cells was decreased in HTLV-1 carriers coinfected with helminths (P<.05). The percentage of interleukin (IL)-5- and IL-10-expressing T cells in HTLV-1 carriers coinfected with helminths was higher than that in HTLV-1 carriers not coinfected with helminths (P<.05). Moreover, we found that the prevalence of helminthic infection was 7-fold higher in HTLV-1 carriers than in patients with HTLV-1-associated myelopathy/tropical spastic paraparesis (P<.05). These data show that helminthic infection decreases activation of type 1 cells, which may influence the clinical outcome of HTLV-1 infection. PMID- 15655787 TI - Calculation of liver-to-blood inocula, parasite growth rates, and preerythrocytic vaccine efficacy, from serial quantitative polymerase chain reaction studies of volunteers challenged with malaria sporozoites. AB - We calculated the number and growth rate of Plasmodium falciparum parasites emerging in recipients of candidate preerythrocytic malaria vaccines and unvaccinated control subjects undergoing mosquito-bite challenge. This was done to measure vaccine efficacy and to distinguish the effects on blood-stage multiplication from those on liver-stage parasites. Real-time polymerase chain reaction measurements of parasite densities were analyzed by nonlinear regression and mixed-effects models. Substantial reductions in numbers of liver parasites resulted from the use of 2 immunization regimens: FP9 boosted by modified virus Ankara (MVA) encoding the malaria epitope-thrombospondin-related adhesion protein insert (92% reduction) and RTS,S/AS02 used in heterologous prime-boost immunization regimens, with MVA encoding the circumsporozoite protein (97% reduction). Forty-eight-hour growth rates in blood from control subjects were not different from those in blood from any vaccination group (mean, 14.4-fold [95% confidence interval, 11-19-fold]). PMID- 15655788 TI - CCR5 plays a critical role in the development of myocarditis and host protection in mice infected with Trypanosoma cruzi. AB - The pathogenesis of myocarditis during Trypanosoma cruzi infection is poorly understood. We investigated the role played by chemokine receptor 5 (CCR5) in the influx of T cells to the cardiac tissue of T. cruzi-infected mice. mRNA and protein for the CCR5 ligands CCL3, CCL4, and CCL5 were detected in the hearts of infected mice in association with CD4+ and CD8+ T cells. There was a high level of CCR5 expression on CD8+ T cells in the hearts of infected mice. Moreover, CCR5 expression on CD8+ T cells was positively modulated by T. cruzi infection. CCR5 deficient mice infected with T. cruzi experienced a dramatically inhibited migration of T cells to the heart and were also more susceptible to infection. These results suggest that CCR5 and its ligands play a central role in the control of T cell influx in T. cruzi-infected mice. Knowledge of the mechanisms that trigger and control the migration of cells to the heart in patients with Chagas disease may help in the design of drugs that prevent myocarditis and protect against the development of severe disease. PMID- 15655790 TI - Are iron-scavenging parasites protective against malaria? PMID- 15655789 TI - A combination of antigenic regions of Toxoplasma gondii microneme proteins induces protective immunity against oral infection with parasite cysts. AB - Infection with Toxoplasma gondii causes morbidity and mortality in congenitally infected and immunocompromised individuals. Both humoral and cell-mediated immunity are involved in host resistance to invasion of the parasite. Among putative vaccine candidates, the T. gondii microneme proteins appear to be promising, because they are responsible for the invasion process. The present work focused on studying the immunogenicity of microneme proteins in infected individuals and in a mouse model of chronic toxoplasmosis. We identified 5 distinct antigenic regions within MIC2, MIC4, MIC2-associated protein, and apical membrane antigen 1 gene products, which were recognized by (1) T cells from both adults with acquired infection and children with congenital infection and (2) antibodies from all patients. Finally, we demonstrated that DNA immunization with microneme fragments elicited effective protection in mice (84% reduction in brain cyst burden), suggesting that a combination of these antigenic regions should be considered in the design of potential vaccines against toxoplasmosis. PMID- 15655791 TI - Tandem mass spectra of transition-metal ion adducts of glycosyl dithioacetals; distinction among stereoisomers. AB - Electrospray ionization mass spectra of some glycosyl dithioacetals recorded in the presence of transition-metal chlorides, XCl2 (where X = Co, Mn and Zn), give abundant adduct ions such as [M+XCl]+ and [2M-H+X]+ and minor ions such as [M H+X]+ and [2M+XCl]+. The tandem mass spectra of these adducts show characteristic elimination of neutral molecules such as H2O, HCl, EtSH, CH2O, C2H4O2/C2H4O. [M+XCl]+ ions fragment readily and the fragmentation appears to be stereochemically controlled as the relative abundances of the fragments are different for three stereoisomers. The added metal is lost as neutral molecules in the form of XCl(OH) and XCl(SEt). This is a predominant pathway in the ZnCl+ adducts. [2M+XCl]+ ions fragment preferentially by elimination of HCl, indicating strong metal interactions in the resulting dimeric [2M-H+X]+ ion. As there are several electron-rich centers in the molecule, the dimeric complex [2M-H+X]+ can have several structures and the observed fragmentations may reflect the sum of those of all these structures. The dimeric complexes fragment by elimination of neutral molecules leaving the dimeric interactions intact. The extent of fragmentation varies for the stereoisomers, leading to stereochemical differentiation. PMID- 15655792 TI - Determination of the ionophoric coccidiostats narasin, monensin, lasalocid and salinomycin in eggs by liquid chromatography/tandem mass spectrometry. AB - A sensitive and selective liquid chromatographic tandem mass spectrometric method (LC/MS/MS) for the simultaneous detection of the ionophoric coccidiostats narasin, monensin, lasalocid and salinomycin in whole eggs has been developed. A very simple sample preparation consisting of an extraction with an organic solvent was carried out. Sample extracts were injected into the LC/MS/MS system on a C18 column and an isocratic elution was performed. Nigericin was used as internal standard. The precursor ions produced by electrospray positive ionisation were selected for collisional dissociation with argon into product ions. Validation of the methods was performed based on Commission Decision 2002/657/EC.1 CC(alpha) was found to be 1 microg/kg for all four compounds. Monitoring of Belgian egg samples in 2004 revealed that residues of salinomycin, lasalocid and monensin could be found. PMID- 15655793 TI - Early prediction of wheat quality: analysis during grain development using mass spectrometry and multivariate data analysis. AB - Matrix-assisted laser desorption/ionisation time-of-flight mass spectrometry and multivariate data analysis have been used for the determination of wheat quality at different stages of grain development. Wheat varieties with one of two different end-use qualities (i.e. suitable or not suitable for bread-making purposes) were investigated. The samples were collected from grains from 15 until 45 days post-anthesis (dpa). Gluten proteins from wheat grains were extracted and subsequently analysed by mass spectrometry. Discrimination partial least-squares regression and soft independent modelling of class analogy were used to determine the quality of new and unknown wheat samples. With these methods, we were able to predict correctly the end-use qualities at every stage investigated. This new fast technique, based on the rapidity of mass spectrometry combined with the objectivity of multivariate data analysis, offers a method that can replace the traditional rather time-consuming ones such as gel electrophoresis. This study focused on the determination of wheat quality at 15 dpa, when the grain is due for harvest 1 month later. PMID- 15655794 TI - Implications of enzymatic, acidic and thermal hydrolysis of DNA on the occurrence of cross-linked melphalan DNA adducts. AB - Calf thymus DNA was treated with melphalan, a nitrogen mustard, and the formation of melphalan cross-linked DNA adducts was investigated. These cross-linked adducts could not be detected either in the enzymatically or in the thermally generated DNA hydrolysates. However, a search for DNA cross-linked adducts in the hydrolysates obtained under acidic conditions revealed the presence of different types of cross-links, mainly containing an adenine moiety. These results are very important because they show that the detection of cross-links is dependent on the hydrolytic procedure used and that these cross-linked adducts are formed under totally different reaction conditions from those in in vivo situations. This can explain the very low abundance or even the absence of cross-linked adducts in nitrogen mustard treated animals. The generally accepted theory that the anti cancer activity of bifunctional mustards such as melphalan is due to cross linking of DNA strands remains therefore from our point of view questionable. PMID- 15655795 TI - Isotope analysis of hydrocarbons: trapping, recovering and archiving hydrocarbons and halocarbons separated from ambient air. AB - It is argued that isotope analysis of atmospheric non-methane hydrocarbons (NMHCs) and, in particular, the analysis of the deuterium/hydrogen (D/H) ratio is valuable because the dominant self-cleansing property of the troposphere is based on the OH radical which removes, e.g., CH4 and other alkanes by H-atom abstraction, which induces large kinetic isotope effects. The major obstacle in applying D/H isotope analysis to atmospheric NMHCs is not only the low abundance of D itself but, in particular, the low concentrations of NMHCs in the parts per trillion range. We show how a selection of NMHCs can be quantitatively separated from 300 L air samples together with CO2 as carrier gas matrix, by using high efficiency cryogenic traps. After diluting the extracted NMHC mixtures with hydrocarbon free air, and determining the mixing ratios, good agreement with original whole air sample analysis exists for alkanes and several halocarbons. For unsaturated hydrocarbons and some other halocarbons the extraction and recovery yield under the given conditions fell considerably, as a function of boiling point. Furthermore, the mixture of NMHCs in the CO2 matrix is proven to remain unchanged over several years when conveniently stored in glass ampoules. The 'extracts' or 'concentrates' of condensables extracted from larger air samples will enable the D/H isotope analysis of ultra trace gases in the atmosphere. PMID- 15655796 TI - Multidimensional statistical analysis applied to electron ionization mass spectra to determine steroid stereochemistry. AB - The differentiation of stereoisomers on the basis of their mass spectra only is usually a difficult challenge even when an informative ionization technique such as electron ionization is used; this is particularly the case for steroids. In this study, multivariate statistical techniques have been applied to the mass spectra of derivatized 5xi-androstane-3xi,17xi-diols (xi = alpha,beta) in order to investigate the possibility of discrimination among the different isomers. After collection of the data from the mass spectra (20 replicates for each of the 8 isomers), each ion was considered as a statistical variable and each mass spectrum as an observation. The more discriminative variables (42 out of the 160 initial ones) were selected using the analysis of variance technique (ANOVA). Thereafter, a linear discriminant analysis (LDA) allowed us to set up a predictive model for stereochemistry determination. The two-dimensional graphical display of the 160 observations on the basis of the canonical variables derived from LDA made it possible to separate the eight isomers. The discrimination of 5alpha and 5beta isomers as well as 3alpha and 3beta was unambiguous, whereas, the discrimination of 17alpha and 17beta epimers was less obvious. The robustness of the model was checked with 40 mass spectra recorded over a 6-month period on different quadrupole mass spectrometers and under different signal acquisition conditions. The percentage of correct assignment of these 'unknown' stereoisomers was 92%; only three 17alpha and 17beta epimers were not correctly plotted in the expected zone. Nevertheless, the performance score was better than those observed with traditional mass spectral libraries. Furthermore, this statistical approach allowed us to identify the main fragment ions involved in the discrimination between isomers: m/z 256 and 421 for isomers 5a-5b; m/z 241 and 331 for isomers 5alpha3alpha-5alpha3beta; m/z 143 and 162 for isomers 5beta3alpha-5beta3beta; and m/z 255 for epimers 17alpha-17beta. PMID- 15655797 TI - Benzene-assisted atmospheric-pressure chemical ionization: a new liquid chromatography/mass spectrometry approach to the analysis of selected hydrophobic compounds. AB - Charge-exchange reactions involving benzene have been successfully exploited to increase the sensitivity of atmospheric-pressure chemical ionization mass spectrometry (APCI-MS) towards hydrophobic compounds of significant environmental relevance which are not detectable with the ordinary APCI techniques. Among them, good sensitivity have been found for (a) highly chlorinated biphenyl derivatives such as dichlorodiphenyltrichloroethane (DDT), dichlorodiphenyldichloroethane (DDD) and dichlorodiphenyldichloroethene (DDE); (b) cyclopentadienes such as Aldrin and its epoxy derivatives Dieldrin and Endrin; and (c) dibenzofurans and dibenzo-para-dioxins such as 2,3,7,8-tetrachlorodibenzofuran (2,3,7,8-TCDF) and 2,3,7,8-tetrachloro-dibenzo-para-dioxin (2,3,7,8-TCDD). The reactant benzene molecules were introduced into the source either through the nebulizer gas or by direct post-column addition of neat liquid, whereas the targeted compounds were analyzed using a high-performance liquid chromatography (HPLC) RP-18 column using methanol/water solutions as mobile phase. By using benzene as post-column reagent, positive ion mode detection was proven to be significantly enhanced as compared with APCI measurements carried out without benzene assistance. PMID- 15655798 TI - Ionization and fragmentation of N-linked glycans as silver adducts by electrospray mass spectrometry. AB - [M+Ag]+ ions were produced by electrospray from neutral high-mannose, hybrid and complex N-linked glycans obtained from bovine ribonuclease, chicken egg glycoproteins, bovine fetuin and porcine thyroglobulin by the addition of silver nitrate to the electrospray solvent. Both singly and doubly charged ions were produced but, as the signals were split between the two silver isotopes, sensitivity was not as high as with the sodium adducts reported earlier. Collision-induced dissociation (CID) spectra were dominated by ions produced by glycosidic cleavages, mainly of the B- and Y-type. Internal cleavage ions involving both B and Y cleavages were very prominent but cross-ring fragments were generally of very low abundance or absent. Silver was very efficient at cleaving the glycosidic bonds, so much so that spectra tended to contain glycosidic ions at most possible combinations of the constituent monosaccharides. PMID- 15655799 TI - Differential adduction of proteins vs. deoxynucleosides by methyl methanesulfonate and 1-methyl-1-nitrosourea in vitro. AB - The reactions of two model mutagenic and carcinogenic alkylating agents, N-methyl N-nitrosourea (MNU) and methyl methanesulfonate (MMS), with proteins and deoxynucleosides in vitro, were investigated. The protein work used an approach involving trypsin digestion and high-performance liquid chromatography/electrospray ionization tandem mass spectrometry (HPLC/ESI-MS/MS). This technique permitted identification of the specific location of protein adduction by both MNU and MMS with commercial apomyoglobin and human hemoglobin, under physiological conditions. MNU treatment resulted in predominantly carbamoylation adducts on the proteins, but in contrast only methylated protein adducts were found following treatment with MMS. Further analyses, using TurboSequest, and the Scoring Algorithm for Spectral Analysis (SALSA), revealed that MNU carbamoylation was specific for modification of either the N-terminal valine or the free amino group in lysine residues of apomyglobin and human hemoglobin. However, MMS methylation modified the N-terminal valine and histidine residues of the proteins. Despite their clear differences in protein modifications, MNU and MMS formed qualitatively the same methylated deoxynucleoside adduct profiles with all four deoxynucleosides in vitro under physiological conditions. In light of their different biological potencies, where MMS is considered a 'super clastogen' while MNU is a 'super mutagen', these differences in reaction products with proteins vs. deoxynucleosides may indicate that these two model alkylating agents work via different mechanisms to produce their mutagenic and carcinogenic effects. PMID- 15655800 TI - Electrospray ionization tandem mass spectrometry of model peptides reveals diagnostic fragment ions for protein ubiquitination. AB - In this work, synthetic peptides were used to determine the fragmentation behavior of ubiquitinated peptides and to find ions diagnostic for peptide ubiquitination. The ubiquitin-calmodulin peptide1 was chosen as the model peptide for naturally occurring ubiquitinated proteins cleaved with endoproteinase gluC. In addition, the fragmentation behavior of model ubiquitinated peptides produced by tryptic digestion was also of great interest since the standard protocols for proteomics-based protein identification use trypsin as the protease. Attachment of ubiquitin to a target protein results in a branched structure, but only ions from the ubiquitin side chain (and the lysine to which it is attached) can be used as diagnostic ions, since fragment ions that contain other amino acids from the parent protein will vary in mass. Characteristic b-type fragment ions from the gluC cleavage of the ubiquitin side chain (designated as b ions) were found which involve only the ubiquitin tail (b2, b3, b4, b5 and b6 ions at m/z 189.06, 302.12, 439.18, 552.30 and 651.30, respectively). Maximum production of these ions occurred at a collision energy of 45 eV in a Q-TOF instrument. Although a non-ubiquitinated peptide may produce isobaric fragment ions, it is unlikely that it can produce these ions in combination. With liquid chromatography/tandem mass spectrometry (LC/MS/MS) experiments, ubiquitinated peptides can readily be determined by surveying the reconstructed or extracted ion chromatograms of the diagnostic fragment ions for common peaks. Characteristic ions resulting from tryptic cleavage of the side chain were found in cleavage products with a missed cleavage, resulting in a LRGG- tag instead of a GG- tag. For the LRGG-tagged peptide, diagnostic MS/MS fragment ions (at m/z 270.17 and 384.21) from the ubiquitin tail (b2 and b4, respectively) were found, along with an internal fragment ion (LRGGK-28) at m/z 484.30. These ions should prove useful in precursor-ion scanning experiments for identifying peptides modified by attachment of ubiquitin, and for locating the site of ubiquitin attachment. PMID- 15655801 TI - Analysis of tetracyclines in honey by high-performance liquid chromatography/tandem mass spectrometry. AB - A confirmatory method coupling liquid chromatography to tandem mass spectrometry (LC/MS/MS) is described for the determination of tetracycline, oxytetracycline, doxycycline and chlortetracycline in honey. Demeclocycline, another tetracycline molecule not reported for its usage in honey, was used as internal standard to quantify the four analytes. The sample preparation entails a clean-up on an Oasis HLB solid-phase extraction cartridge and analyses were realised by LC/MS/MS in selected reaction monitoring mode. The stability of tetracyclines was checked under various storage conditions at -20, +4 and +20 degrees C (both under dark and light exposures). Indeed, tetracyclines are not stable molecules and the epimerisation phenomenon was evaluated in this work. Appropriate correction factors of the MS/MS responses of each epimer were studied for each of the four tetracyclines to accurately quantify them. Moreover, the matrix effects encountered during the LC/MS/MS analyses were also studied in spiked experiments from blank honey samples of various geographical origins and different flower types. PMID- 15655802 TI - Antimycin A-induced killing of HL-60 cells: apoptosis initiated from within mitochondria does not necessarily proceed via caspase 9. AB - BACKGROUND: Antimycin A (AMA) inhibits mitochondrial electron transport, collapses the mitochondrial membrane potential, and causes the production of reactive oxygen species. Previous work by me and my colleagues has demonstrated that AMA causes an array of typical apoptotic phenomena in HL-60 cells. The hypothesis that AMA causes HL-60 apoptosis by the intrinsic apoptotic pathway has now been tested. METHODS: Z-LEHD-FMK and Z-IETD-FMK were used as specific inhibitors of the initiator caspases 9 and 8, respectively. Caspase 3 activation, DNA fragmentation, and cellular disintegration were measured by flow cytometry. Cytochrome c release, chromatin condensation, and nuclear fragmentation were measured by microscopy. RESULTS: AMA caused mitochondrial cytochrome c release and neither Z-LEHD-FMK nor Z-IETD-FMK inhibited that. In the absence of caspase inhibition there was a very close correlation between cytochrome c release and caspase 3 activation. Z-LEHD-FMK blocked caspase 3 activation but enhanced DNA fragmentation and failed to stop nuclear or cellular disintegration. Z-IETD-FMK also blocked caspase 3 activation but, in contrast to Z-LEHD-FMK, delayed DNA fragmentation and disintegration of the nucleus and the cell. CONCLUSIONS: The hypothesis to explain AMA-induced HL-60 apoptosis was clearly inadequate because: (a) caspase 9 inhibition did not prevent DNA fragmentation or cell death, (b) apoptosis proceeded in the absence of caspase-3 activation, (c) the main pathway leading to activation of the executioner caspases was by caspase-8 activation, but caspase 8 inhibition only delayed apoptosis, and (d) activation of caspases 8 and 9 may be necessary for caspase-3 activation. Thus, in this cell model, apoptosis triggered from within the mitochondria does not necessarily proceed by caspase 9, and caspase 3 is not critical to apoptosis. The results provide further evidence that, when parts of the apoptotic network are blocked, a cell is able to complete the program of cell death by alternate pathways. PMID- 15655803 TI - Direct injection of plasmid DNA into the skin induces dermatitis by activation of monocytes through toll-like receptor 9. AB - BACKGROUND: Direct injection of naked DNA into skin can be efficiently used to transfer genes into keratinocytes in vivo. However, bacterial DNA is known to be a potent stimulus for vertebrate immune cells and immune systems. Towards the clinical use of this method, this study examined whether the application of plasmid DNA by direct injection induces any adverse skin effects. METHODS: Several plasmid preparations were prepared and directly injected into rat and human skin. Migration, IL-6 production, and reactive oxygen production assays were performed to determine the type of the primary cells responsible for the reaction. Involvement of toll-like receptor (TLR) 9 was examined by experiments using TLR9-knockout mice. RESULTS: Injection of several plasmid preparations into rat and human skin resulted in an inflammatory reaction at the treated site. Contamination by endotoxin in the plasmid preparation was shown to worsen this skin inflammation reaction. Immunohistochemical analysis showed that the infiltrating cells in the skin lesions were predominantly monocytes and neutrophils. Further experiments examining migration, IL-6 production, and reactive oxygen production indicated that the primary responsible cells were monocytes rather than neutrophils. Since it was recently shown that cytosine guanosine dinucleotide (CpG) motif is critical for immune reaction induction in bacterial DNA and cellular responses were mediated by TLR9, we injected plasmids into the ear skin of TLR9-knockout mice. A decrease in ear swelling was noted in the knockout mice, compared to controls, suggesting that plasmid-DNA-induced dermatitis was mediated mostly by TLR9. CONCLUSIONS: This study demonstrates that injection of plasmid DNA induces skin inflammation initiated by monocyte activation via TRL9. We should therefore attempt to counteract this dermatitis during the clinical use of the naked DNA injection method in skin. PMID- 15655804 TI - Tight control of transgene expression by lentivirus vectors containing second generation tetracycline-responsive promoters. AB - BACKGROUND: The goal of this study was to design improved regulatable lentivirus vector systems. The aim was to design tetracycline (tet)-regulatable lentivirus vectors based on the Tet-on system displaying low background expression in the absence of the doxycycline (DOX) inducer and high transgene expression levels in the presence of DOX. METHODS: We constructed a binary lentivirus vector system that is composed of a self-inactivating (SIN) lentivirus vector bearing inducible first- or second-generation tet-responsive promoter elements (TREs) driving expression of a transgene and a second lentivirus vector encoding a reverse tetracycline-controlled transactivator (rtTA) that activates transgene expression from the TRE in the presence of DOX. RESULTS: We evaluated a number of different rtTAs and found rtTA2S-M2 to induce the highest levels of transgene expression. Regulated transgene expression was stable in human breast carcinoma cells implanted into nude mice for up to 11 weeks. In an attempt to minimize background expression levels, the chicken beta-globin cHS4 insulator element was cloned into the 3' long terminal repeat (LTR) of the transgene transfer vector. The cHS4 insulator element reduced background expression but expression levels following DOX addition were lower than those observed with vectors lacking an insulator sequence. In a second strategy, vectors bearing second-generation TREs harboring repositioned tetracycline operator elements were used. Such vectors displayed greatly reduced leakiness in the absence of DOX and induced transgene expression levels were up to 522-fold above those seen in the absence of DOX. CONCLUSIONS: Inducible lentivirus vectors bearing insulators or second-generation TREs will likely prove useful for applications demanding the lowest levels of background expression. PMID- 15655805 TI - Long-term survival and concomitant gene expression of ribozyme-transduced CD4+ T lymphocytes in HIV-infected patients. AB - BACKGROUND: An anti-HIV-1 tat ribozyme, termed Rz2, has been shown to inhibit HIV 1 infection/replication and to decrease HIV-1-induced pathogenicity in T lymphocyte cell lines and normal peripheral blood T-lymphocytes. We report here the results of a phase I gene transfer clinical trial using Rz2. METHODS: Apheresis was used to obtain a peripheral blood cell population from each of four HIV-negative donors. After enrichment for CD4+ T-lymphocytes, ex vivo expansion and genetic manipulation (approximately equal aliquots of the cells were transduced with the ribozyme-containing (RRz2) and the control (LNL6) retroviral vector), these cells were infused into the corresponding HIV-1-positive twin recipient. Marking was assessed over an initial 24-week period and in total over an approximate 4-year period. RESULTS: The gene transfer procedure was shown to be safe, and technically feasible. Both RRz2- and LNL6-gene-containing peripheral blood mononuclear cells (PBMC) were detected at all time points examined to 4 years. There was concomitant gene construct expression in the absence of the need for ex vivo peripheral blood cell stimulation and there was no evidence of immune elimination of the neoR T-lymphocytes nor of silencing of the Moloney murine leukemia virus long terminal repeat. CONCLUSIONS: The proof of principle results reported here demonstrate safety and feasibility of this type of gene transfer approach. While not specifically tested, T-lymphocytes containing an anti-HIV gene construct may impact on HIV-1 viral load and CD4+ T-lymphocyte count, potentially representing a new therapeutic modality for HIV-1 infection. PMID- 15655806 TI - A risk assessment of atrazine use in California: human health and ecological aspects. AB - A risk assessment of the triazine herbicide atrazine has been conducted by first analyzing the toxicity database and subsequently estimating exposure. Margins of safety (MOS) were then calculated. Toxicity was assessed in animal studies and exposure was estimated from occupational and dietary sources. In acute toxicity studies, atrazine caused developmental toxicity in the rabbit [no observed effect level (NOEL) 5 mg kg(-1) day(-1)] and cardiotoxicity in a dog chronic study (NOEL 0.5 mg kg(-1) day(-1)); cancer (mammary glands) resulted from lifetime exposure. The mammary tumors, which occurred specifically in female Sprague-Dawley rats, were malignant, increased in a dose-dependent manner and were also observed with other, related triazines. Evidence for a genotoxic basis for these tumors was either equivocal or negative. Triazines have been shown to be clastogenic in Chinese hamster ovary cells, in vitro, but without showing a convincing dose/response relationship. Atrazine can be converted into genotoxic N nitrosoatrazine in the environment or the digestive system, suggesting that N nitrosamines derived from triazines could be oncogenic. However, it was concluded that N-nitrosotriazines are unlikely to play a significant role in triazine induced rat mammary gland tumors. An endocrine basis for the mammary tumors, involving premature aging of the female SD rat reproductive system, has been proposed. A suppression of the luteinizing hormone surge during the estrus cycle by atrazine leads to the maintenance of elevated blood levels of 17beta-estradiol (E2) and prolactin. The mechanism for tumor development may include one or more of the following: the induction of aromatase (CYP19) and/or other P450 oxygenases, an antagonist action at the estrogen feedback receptor in the hypothalamus, an agonist action at the mammary gland estrogen receptor or an effect on adrenergic neurons in the hypothalamic-pituitary pathway. None of these has been excluded as a target because there has been a lack of a rigorous attempt to address the mechanism of action for mammary tumors at the molecular level. The potential occupational exposure to atrazine was assessed during mixing, loading and application. Absorbed daily dosage values were 1.8-6.1 microg kg(-1) day(-1). The MOS values (animal NOEL/human exposure) for short-term (acute) exposure were 820-2800. Longer-term occupational exposure and risk were also calculated. Detectable crop residues are generally absent at harvest. Theoretical calculations of acute dietary exposure used tolerance levels, along with secondary residues, and water, for which there is a maximum contamination level; atrazine plus the three main chlorotriazine metabolites were combined. MOS values were above 2000 for all population subgroups. Dietary exposure to atrazine is therefore extremely unlikely to result in human health hazard. Recent publications have reported a possible feminization of frogs, measured in laboratory and field studies. This is assumed to be due to the induction of aromatase, but no measurements of enzyme activity have been reported. In field studies, the water bodies with the greatest numbers of deformed frogs sometimes had the lowest concentrations of atrazine. Other studies have also cast doubt on the feminization theory, except perhaps at very high levels of atrazine. Epidemiology studies have investigated the possibility that atrazine may result in adverse effects in humans. Although some studies have claimed that atrazine exposure results in an elevated risk of prostate cancer, the published literature is inconclusive with respect to cancer incidence. PMID- 15655807 TI - Effects of cyclic AMP-elevating hormones and autacoids on LPS-activated rat peritoneal, bronchoalveolar and hepatic (Kupffer) macrophages. AB - Peritoneal, bronchoalveolar and hepatic (Kupffer) macrophages activated in vitro by endotoxin, exhibit alterations in nitric oxide production when certain hormones or other biologically active agents (autacoids) are present in the culture medium. They also show changes in acid beta-glucuronidase activities and morphological changes concerning cell size and general appearance. Agents known to elevate the intracellular levels of cyclic AMP, e.g. adrenalin, prostaglandin E2 and dopamine, increase the nitric oxide production in all three types of macrophage. The addition of H-89, an inhibitor of protein kinase A, abolishes the increase in nitric oxide production. Adrenalin also increases the extracellular activity of beta-glucuronidase. The results of this work suggest that cyclic AMP elevating hormones and autacoids affect the functions of endotoxin-activated macrophages, such as the production of nitric oxide and the activity of acid beta glucuronidase. PMID- 15655808 TI - Prospective studies on mental status and quality of life in patients with head and neck cancer treated by radiation. AB - Factors affecting the quality of life (QOL) in patients undergoing radiation therapy for head and neck cancer were determined in a prospective study. Full psychiatric interviews and self-report questionnaires concerning patients' QOL, pain, anxiety, mood, coping style and personality were administered to 35 patients with Stage I or II head and neck cancer before radiation therapy (week 0), and approximately one month (week 4) and two months (week 8) after starting of radiation. Anxiety, depression and pain were more severe at week 4 than at baseline. Anxiety was less at week 8, but depressive symptoms remained remarkable. Stepwise regression analyses revealed that changes in depressed mood and pain partially accounted for changes in disease-specific QOL. General health QOL related to a premorbid personality of openness and an emotional aspect of a coping style. Therapeutic interventions including psychiatric management of depression and physical management of pain appear to be crucial for preserving QOL during radiation treatment of patients with head and neck cancer. PMID- 15655809 TI - Current research of hepatic cirrhosis in China. AB - Hepatic cirrhosis is a common disease that poses a serious threat to public health, and is characterized by chronic, progressive and diffuse hepatic lesions preceded by hepatic fibrosis regardless of the exact etiologies. In recent years, considerable achievements have been made in China in research of the etiopathogenesis, diagnosis and especially the treatment of hepatic fibrosis, resulting in much improved prognosis of hepatic fibrosis and cirrhosis. In this paper, the authors review the current status of research in hepatic fibrosis, cirrhosis and their major complications. PMID- 15655810 TI - Increased expression of mitogen-activated protein kinase and its upstream regulating signal in human gastric cancer. AB - AIM: To investigate the expression of mitogen-activated protein kinases (MAPKs) and its upstream protein kinase in human gastric cancer and to evaluate the relationship between protein levels and clinicopathological parameters. METHODS: Western blot was used to measure the expression of extracellular signal-regulated kinase (ERK)-1, ERK-2, ERK-3, p38 and mitogen or ERK activated protein kinaseMEK 1 proteins in surgically resected gastric carcinoma, adjacent normal mucosa and metastatic lymph nodes from 42 patients. Immunohistochemistry was employed for their localization. RESULTS: Compared with normal tissues, the protein levels of ERK-1 (integral optical density value 159 526+/-65 760 vs 122 807+/-65 515, P = 0.001), ERK-2 (168 471+/-95 051 vs 120 469+/-72 874, P<0.001), ERK-3 (118 651+/ 71 513 vs 70 934+/-68 058, P<0.001), P38 (104 776+/-51 650 vs 82 930+/-40 392, P = 0.048) and MEK-1 (116 486+/-45 725 vs 101 434+/-49 387, P = 0.027) were increased in gastric cancer tissues. Overexpression of ERK-3 was correlated to TNM staging (average ratio of integral optic density (IOD)(tumor): IOD(normal) in TNM I, II, III, IV tumors was 1.43+/-0.34, 5.08+/-3.74, 4.99+/-1.08, 1.44+/-1.02, n = 42, P = 0.023) and serosa invasion (4.31+/-4.34 vs 2.00+/-2.03, P = 0.037). In poorly differentiated cancers (n = 33), the protein levels of ERK-1 and ERK-2 in stage III and IV tumors were higher than those in stage I and II tumors (2.64+/-3.01 vs 1.01+/-0.33, P = 0.022; 2.05+/-1.54 vs 1.24+/-0.40, P = 0.030). Gastric cancer tissues with either lymph node involvement (2.49+/-2.91 vs 1.03+/ 0.36, P = 0.023; 1.98+/-1.49 vs 1.24+/-0.44, P = 0.036) or serosa invasion (2.39+/-2.82 vs 1.01+/-0.35, P = 0.022; 1.95+/-1.44 vs 1.14+/-0.36, P = 0.015) expressed higher protein levels of ERK-1 and ERK-2. In Borrmann II tumors, expression of ERK-2 and ERK-3 was increased compared with Borrmann III tumors (2.57+/-1.86 vs 1.23+/-0.60, P = 0.022; 5.50+/-5.05 vs 1.83+/-1.21, P = 0.014). Borrmann IV tumors expressed higher p38 protein levels. No statistically significant difference in expression of MAPKs was found when stratified to tumor size or histological grade (P>0.05). Protein levels of ERK-2, ERK-3 and MEK-1 in metastatic lymph nodes were 2-7 folds higher than those in adjacent normal mucosa. The immunohistochemistry demonstrated that ERK-1, ERK-2, ERK-3, p38 and MEK-1 proteins were mainly localized in cytoplasm. The expression of MEK-1 in gastric cancer cells metastasized to lymph nodes was higher than that of the primary site. CONCLUSION: MAPKs, particularly ERK subclass are overexpressed in the majority of gastric cancers. Overexpression of ERKs is correlated to TNM staging, serosa invasion, and lymph node involvement. The overexpression of p38 most likely plays a prominent role in certain morphological subtypes of gastric cancers. MEK-1 is also overexpressed in gastric cancer, particularly in metastatic lymph nodes. Upregulation of MAPK signal transduction pathways may play an important role in tumorigenesis and metastatic potential of gastric cancer. PMID- 15655811 TI - Expression of Epstein-Barr virus genes in EBV-associated gastric carcinomas. AB - AIM: To understand the expression of latent and lytic genes of Epstein-Barr virus (EBV) in EBV-associated gastric carcinoma (EBVaGC) and to explore the relationship between EBV-encoded genes and development of EBVaGC at molecular level. METHODS: One hundred and seventy-two gastric carcinoma tissues and 172 corresponding para-carcinoma tissues were tested for EBV genome by polymerase chain reaction (PCR)-Southern blotting. EBV-encoded small RNA (EBER) 1 of the PCR positive specimens was detected by in situ hybridization (ISH). Gastric carcinomas with positive EBER1 signals were classified as EBVaGCs. RT-PCR and Southern hybridization were applied to the detection of expression of nuclear antigen (EBNA) promoters (Qp, Wp and Cp), EBNA 1 and EBNA 2, latent membrane proteins (LMP) 1, 2A and 2B and lytic genes (immediate early genes BZLF1 and BRLF1, early genes BARF1 and BHRF1, late genes BcLF1 and BLLF1) in EBVaGCs. RESULTS: Eleven EBV positive samples existed in gastric carcinoma tissues (6.39%). No EBV positive sample was found in corresponding para-carcinoma tissues. The difference between EBV positivity in carcinoma tissues and corresponding para-carcinoma tissues was significant (chi(2) = 9.0909, P = 0.0026). Transcripts of Qp and EBNA1 were detected in all the 11 EBVaGCs, while both Wp and Cp were silent. EBNA2, LMP1 and LMP2B mRNA were absent in all the cases, while LMP2A mRNA was detected in 4 of the 11 cases. Of the 11 EBVaGCs, 7 exhibited BcLF1 transcripts and 2 exhibited BHRF1 transcripts. The transcripts of BZLF1 and BARF1 were detected in 5 cases, respectively. No BLLF1 and BRLF mRNA were detected. CONCLUSION: The latent pattern of EBV in gastric carcinoma corresponds to the latency I/II. Some lytic infection genes are expressed in EBVaGCs tissues. BARF1 and BHRF1 genes may play an important role in tumorigenesis of gastric carcinoma. PMID- 15655812 TI - Selection of optimal antisense accessible sites of survivin and its application in treatment of gastric cancer. AB - AIM: To select the optimal antisense accessible sites of survivin, a highly expressed gene in tumor tissues, in order to explore a novel approach to improve biological therapy of gastric cancer. METHODS: The 20 mer random oligonucleotide library was synthesized, hybridized with in vitro transcribed total survivin cRNA, then digested by RNase H. After primer extension and autoradiography, the antisense accessible sites (AAS) of survivin were selected. Then RNADraw software was used to analyze and choose the AAS with obvious stem-loop structures, according to which the complementary antisense oligonucleotides (AS-ODNs) were synthesized and transferred into survivin highly- expressing gastric cancer cell line MKN-45. Survivin expression was detected by RT-PCR and Western Blotting. Cellular growth activities were assayed by tetrazolium bromide (MTT) colorimetry. Cellular ultrastructure was observed by electronic microscopy, while apoptosis was detected by annexin V-FITC and propidium iodide staining flow cytometry. RESULTS: Thirteen AAS of survivin were selected in vitro. Four AAS with stem-loop structures were chosen, locating at 207-226 bp, 187-206 bp, 126-145 bp and 44-63 bp of survivin cDNA respectively. When compared with non-tranfection controls, their corresponding AS-ODNs (AS-ODN(1), AS-ODN(2), AS-ODN(3) and AS-ODN(4)) could reduce Survivin mRNA levels in MKN-45 cells by 54.3+/-1.1% (t = 6.12, P<0.01), 86.1+/-1.0% (t = 5.27, P<0.01), 32.2+/-1.3% (t = 7.34, P<0.01) and 56.2+/-0.9% (t = 6.45, P<0.01) respectively, while survivin protein levels were decreased by 42.2+/-2.5% (t = 6.26, P<0.01), 75.4+/-3.1% (t = 7.11, P<0.01), 28.3+/-2.0% (t = 6.04, P<0.01) and 45.8+/-1.2% (t = 6.38, P<0.01) respectively. After transfection with 600 nmol/L AS-ODN1-AS-ODN(4) for 24 h, cell growth was inhibited by 28.12+/ 1.54% (t = 7.62, P<0.01), 38.42+/-3.12% (t = 7.75, P<0.01), 21.46+/-2.63% (t = 5.94, P<0.01) and 32.12+/-1.77% (t = 6.17, P<0.01) respectively. Partial cancer cells presented the characteristic morphological changes of apoptosis, with apoptotic rates being 19.31+/-1.16% (t = 7.16, P<0.01), 29.24+/-1.94% (t = 8.15, P<0.01), 11.87+/-0.68% (t = 6.68, P<0.01) and 21.68+/-2.14% (t = 7.53, P<0.01) respectively. CONCLUSION: The AAS of survivin could be effectively selected in vitro by random oligonucleotide library/RNase H cleavage method combined with computer software analysis, this has important reference values for further studying survivin-targeted therapy strategies for gastric cancer. PMID- 15655813 TI - Quantitative assessment model for gastric cancer screening. AB - AIM: To set up a mathematic model for gastric cancer screening and to evaluate its function in mass screening for gastric cancer. METHODS: A case control study was carried on in 66 patients and 198 normal people, then the risk and protective factors of gastric cancer were determined, including heavy manual work, foods such as small yellow-fin tuna, dried small shrimps, squills, crabs, mothers suffering from gastric diseases, spouse alive, use of refrigerators and hot food, etc. According to some principles and methods of probability and fuzzy mathematics, a quantitative assessment model was established as follows: first, we selected some factors significant in statistics, and calculated weight coefficient for each one by two different methods; second, population space was divided into gastric cancer fuzzy subset and non gastric cancer fuzzy subset, then a mathematic model for each subset was established, we got a mathematic expression of attribute degree (AD). RESULTS: Based on the data of 63 patients and 693 normal people, AD of each subject was calculated. Considering the sensitivity and specificity, the thresholds of AD values calculated were configured with 0.20 and 0.17, respectively. According to these thresholds, the sensitivity and specificity of the quantitative model were about 69% and 63%. Moreover, statistical test showed that the identification outcomes of these two different calculation methods were identical (P>0.05). CONCLUSION: The validity of this method is satisfactory. It is convenient, feasible, economic and can be used to determine individual and population risks of gastric cancer. PMID- 15655814 TI - Cytokeratins and carcinoembryonic antigen in diagnosis, staging and prognosis of colorectal adenocarcinoma. AB - AIM: To evaluate the serum levels of cytokeratins and carcinoembryonic antigen (CEA) in diagnosis, staging and prognosis of patients with colorectal adenocarcinoma. METHODS: The sample consisted of 169 patients. One hundred blood donors formed the control group. Radical surgery was performed on 120 patients, with an average follow-up duration of 22.3 mo. Relapses occurred in 23 individuals after an average of 18.09 mo. CEA was assayed via the Delfia method with a limit of 5 ng/mL. Cytokeratins were assayed via the LIA-mat TPA-M Prolifigen method with a limit of 72 U/L. RESULTS: In the diagnosis of patients with colorectal adenocarcinoma, CEA showed a sensitivity of 56%, a specificity of 95%, a positive predictive value of 94%, a negative predictive value of 50% and an accuracy of 76.8%. TPA-M had a sensitivity of 70%, a specificity of 96%, a positive predictive value of 97%, a negative predictive value of 66% and an accuracy of 93.6%. The elevation of one of the markers was shown to have a sensitivity of 76.9%, a specificity of 91%, a positive predictive value of 93.5%, a negative predictive value of 70% and an accuracy of 83.6%. There was no variation in the levels of the markers according to the degree of cell differentiation while there was an elevation in their concentrations in accordance with the increase in neoplastic dissemination. There was a statistically significant difference between the patients with stage IV lesions and those with stages I, II and III tumors. With regard to CEA, the average level was 14.2 ng/mL in patients with stage I lesions, 8.5 ng/mL in patients with stage II lesions, 8.0 ng/mL in patients with stage III lesions and 87.7 ng/mL in patients with stage IV lesions. In relation to TPA-M, the levels were 153.1 U/L in patients with stage I tumors, 106.5 U/L in patients with stage II tumors, 136.3 U/L in patients with stage III tumors and 464.3 U/L in patients with stage IV tumors. There was a statistical difference in patients with a high CEA level in relation to a shorter survival (P<0.05). However, there was no correlation between patients with high TPA-M levels and prognostic indices of patients undergoing radical surgery. CONCLUSION: Cytokeratins demonstrate a greater sensitivity than CEA in the diagnosis of colorectal adenocarcinoma. There is an increase in the sensitivity of the markers with tumor dissemination. Cytokeratins cannot identify the worse prognosis in patients undergoing radical surgery. Cytokeratins constitute an advance in the direction of a perfect tumor marker in the treatment of patients with colorectal cancer. PMID- 15655815 TI - In vitro resistance to interferon of hepatitis B virus with precore mutation. AB - AIM: Chronic hepatitis B virus (HBV) infection is predominantly treated with interferon alpha (IFN-alpha), which results in an efficient reduction of the viral load only in 20-40% of treated patients. Mutations at HBV precore prevail in different clinical status of HBV infection. The roles of precore mutation in the progression of chronic hepatitis and interferon sensitivity are still unknown. The aim of this study was to explore if there was any relationship between HBV precore mutation and sensitivity to interferon in vitro. METHODS: HBV replication-competent recombinant constructs with different patterns of precore mutations were developed. Then the recombinants were transiently transfected into hepatoma cell line (Huh7) by calcium phosphate transfection method. With or without IFN, viral products in culture medium were collected and quantified 3 d after transfection. RESULTS: We obtained 4 recombinant constructs by orientation cloning 1.2-fold-overlength HBV genome into pUC18 vector via the EcoRI and Hind III and PCR mediated site-directed mutagenesis method. All the recombinants contained mutations within precore region. Huh7 cells transfected with recombinants secreted HBsAg and HBV particles into the cell culture medium, indicating that all the recombinants were replication-competent. By comparing the amount of HBV DNA in the medium, we found that HBV DNA in medium reflecting HBV replication efficiency was different in different recombinants. Recombinants containing precore mutation had fewer HBV DNAs in culture medium than wild type. This result showed that recombinants containing precore mutation had lower replication efficiency than wild type. HBV DNA was decreased in pUC18-HBV1.2-WT recombinants after IFN was added while others with precore mutations were not, indicating that HBV harboring precore mutation was less sensitive to IFN in cell culture system. CONCLUSION: These data indicate that HBV harboring precore mutation may be resistant to IFN in vitro. PMID- 15655816 TI - Inhibition of p38 mitogen-activated protein kinase may decrease intestinal epithelial cell apoptosis and improve intestinal epithelial barrier function after ischemia- reperfusion injury. AB - AIM: To investigate the role of p38 mitogen-activated protein kinase in rat small intestine after ischemia-reperfusion (I/R) insult and the relationship between activation of p38 MAPK and apoptotic cell death of intestine. METHODS: Ninety Wistar rats were divided randomly into three groups, namely sham-operated group (C), I/R vehicle group (R) and SB203580 pre-treated group (S). In groups R and S, the superior mesenteric artery (SMA) was separated and occluded for 45 min, then released for reperfusion for 0.25, 0.5, 1, 2, 6, 12 and 24 h. In group C, SMA was separated without occlusion. Plasma D-lactate levels were examined and histological changes were observed under a light microscope. The activity of p38 MAPK was determined by Western immunoblotting and apoptotic cells were detected by the terminal deoxynucleotidyl transferase (TdT)-mediated dUDP-biotin nick end labeling (TUNEL). RESULTS: Intestinal ischemia followed by reperfusion activated p38 MAPK, and the maximal level of activation (7.3-fold vs sham-operated group) was reached 30 min after I/R. Treatment with SB 203580, a p38 MAPK inhibitor, reduced intestinal apoptosis (26.72+/-3.39% vs 62.50+/-3.08% in I/R vehicle, P<0.01) and decreased plasma D-lactate level (0.78+/-0.15 mmol/L in I/R vehicle vs 0.42+/-0.17 mmol/L in SB-treated group) and improved post-ischemic intestinal histological damage. CONCLUSION: p38 MAPK plays a crucial role in the signal transduction pathway mediating post-ischemic intestinal apoptosis, and inhibition of p38 MAPK may attenuate ischemia-reperfusion injury. PMID- 15655818 TI - Toxicity of novel anti-hepatitis drug bicyclol: a preclinical study. AB - AIM: To study the toxicity of bicyclol to animals. METHODS: Acute toxicity test was performed in Kunming strain mice that were orally given bicyclol at the doses of 3 and 5 g/kg body weight, respectively. Wistar rats were orally administered bicyclol at a dose of 5 g/kg body weight. Death and clinical symptoms of animals were recorded within 7 d. Sub-acute toxicity test was carried out in rats that were treated with various doses of bicyclol (150, 300, 600 mg/kg) once daily for 14 d. Animal behaviors, blood biochemical markers, blood and urine pictures were examined. Chronic toxicity test was conducted in 80 Wistar rats of both sexes. The animals were orally administered with various doses of bicyclol (150, 300, 600 mg/kg, 100-400 folds corresponding to the proposed therapeutic dose (1.5 mg/(kg.d)) of bicyclol for patients) once daily for 6 mo except for Sunday. The control group was given the same volume of 0.2% sodium carboxyl methylcellulose (Na-CMC). Twenty-one beagle dogs received bicyclol (25, 75, 225 mg/kg, 16.6, 50, 150 folds corresponding to the proposed therapeutic dose of bicyclol for patients) once a day for 6 mo except for Sunday. The body weight, food intake, urine and feces, blood picture, blood biochemical markers, and pathological examination of main organs were determined. Mutagenicity and teratogenicity were determined. Mutagenicity assay included Ames's test, chromosome aberration test in CHL cells and micronucleus test in mice. For the teratogenicity assay, pregnant Wistar rats weighing 200-250 g were treated with 0.2, 1.0 g/kg bicyclol once daily from the 7th d of gestation for 10 d. RESULTS: The oral LD(50) of bicyclol was over 5 g/kg in mice and rats. No noticeable alterations in subacute and chronic toxicity of rats and dogs were demonstrated. No mutagenicity and teratogenicity of bicyclol were found. CONCLUSION: Bicyclol has no detectable chronic toxicity as well as mutagenicity and teratogenicity in animals. PMID- 15655817 TI - Differences in platelet endothelial cell adhesion molecule-1 expression between peripheral circulation and pancreatic microcirculation in cerulein-induced acute edematous pancreatitis. AB - AIM: To investigate the changes of platelet endothelial cell adhesion molecule-1 (PECAM-1) expression on polymorphonuclear leukocytes (PMNs) in peripheral circulation and pancreatic microcirculation in cerulein-induced acute edematous pancreatitis (AEP). METHODS: Fifty Wistar rats were randomly divided into control group (n = 10) and AEP group (n = 40). A model of AEP was established by subcutaneous injection of cerulein 5.5 and 7.5 mug/kg at 0 and 1 h after the beginning of experiment respectively. PECAM-1 expression on PMNs from splenic vein and inferior vena cava was determined by RT-PCR at mRNA level and determined by flow cytometry at protein level. RESULTS: In experimental rats, an increased PECAM-1 mRNA expression was seen from 4 to 8 h of AEP in peripheral circulation (0.77+/-0.25%, 0.76+/-0.28%, 0.89+/-0.30%, 1.00+/-0.21%), while in pancreatic microcirculation, expression decreased from 2 h and reached the lowest level at 6 h of AEP (0.78+/-0.29%, 0.75+/-0.26%, 0.62+/-0.28%, 0.66+/-0.20%). There were significant differences at 8-h time point of AEP between peripheral circulation and pancreatic microcirculation (1.00+/-0.21% vs 0.66+/-0.20%, P<0.05). Meanwhile, the difference at protein level was also found. CONCLUSION: A reverse expression of PECAM-1 on PMNs was found between peripheral circulation and pancreatic microcirculation, suggesting that inhibition of PECAM-1 expression may improve the pathological change of AEP. PMID- 15655819 TI - Effects of unsaturated fatty acids on calcium-activated potassium current in gastric myocytes of guinea pigs. AB - AIM: To investigate the effects of exogenous unsaturated fatty acids on calcium activated potassium current (I(K(Ca))) in gastric antral circular myocytes of guinea pigs. METHODS: Gastric myocytes were isolated by collagenase from the antral circular layer of guinea pig stomach. The whole-cell patch clamp technique was used to record I(K(Ca)) in the isolated single smooth muscle cells with or without different concentrations of arachidonic acid (AA), linoleic acid (LA), and oleic acid (OA). RESULTS: AA at concentrations of 2,5 and 10 micromol/L markedly increased I(K(ca)) in a dose-dependent manner. LA at concentrations of 5, 10 and 20 micromol/L also enhanced I(K(Ca)) in a dose-dependent manner. The increasing potency of AA, LA, and oleic acid (OA) on I(K(Ca)) at the same concentration (10 micromol/L) was in the order of AA>LA>OA. AA (10 micromol/L) induced increase of I(K(ca)) was not blocked by H-7 (10 micromol/L), an inhibitor of protein kinase C (PKC), or indomethacin (10 micromol/L), an inhibitor of the cyclooxygenase pathway, and 17-octadecynoic acid (10 micromol/L), an inhibitor of the cytochrome P450 pathway, but weakened by nordihydroguaiaretic acid (10 micromol/L), an inhibitor of the lipoxygenase pathway. CONCLUSION: Unsaturated fatty acids markedly increase I(K(ca)), and the enhancing potencies are related to the number of double bonds in the fatty acid chain. The lipoxygenase pathway of unsaturated fatty acid metabolism is involved in the unsaturated fatty acid induced increase of I(K(ca)) in gastric antral circular myocytes of guinea pigs. PMID- 15655820 TI - Hepato-cardiovascular response and its regulation. AB - AIM: To determine the possible existence of a hepato-cardiovascular response and its regulatory mechanism in normal rats. METHODS: Systemic hemodynamic changes following intraportal injection of latex microspheres were studied in two modified rat models of hepatic circulation, in which the extrahepatic splanchnic circulation was excluded by evisceration and the liver was perfused by systemic blood via either the portal vein (model 1) or hepatic artery (model 2) in vivo. RESULTS: In model 1, intraportal injection of two sized microspheres (15-mum and 80-mum) induced a similar decrease in mean arterial pressure, while extrahepatic portal venous occlusion induced an immediate increase in mean arterial pressure. In model 2, microsphere injection again induced a significant reduction in mean arterial pressure, aortic blood flow and aortic resistance. There were no significant differences in these parameters between liver-innervated rats and liver-denervated rats. The degrees of microsphere-induced reduction in mean arterial pressure (-38.1+/-1.9% in liver-innervated rats and -35.4+/-2.1% in liver-denervated rats, respectively) were similar to those obtained by withdrawal of 2.0 mL of blood via the jugular vein (-33.3+/-2.1%) (P>0.05). Injection of 2.0 mL Haemaccel in microsphere-treated rats, to compensate for the reduced effective circulating blood volume, led to a hyperdynamic state which, as compared with basal values and unlike control rats, was characterised by increased aortic blood flow (+21.6+/-3.3%), decreased aortic resistance (-38.1+/-3.5%) and reduced mean arterial pressure (-9.7+/-2.8%). CONCLUSION: A hepato-cardiovascular response exists in normal rats. It acts through a humoral mechanism leading to systemic vasodilatation, and may be involved in the hemodynamic disturbances associated with acute and chronic liver diseases. PMID- 15655822 TI - Ultrastructural changes in non-specific duodenitis. AB - AIM: To investigate the ultrastructural and morphological changes of non-specific duodenitis (NSD) in an attempt to grade them according to the extent of the lesions. METHODS: Biopsies were taken from the mucosa of duodenal bulb of 44 patients selected from the patients undergoing upper gastrointestinal endoscopy for epigastric discomforts. From each patient, two pinch biopsies on the same area were obtained from duodenal bulb. One was for scanning electron microscopy and the other was stained with hematoxylin-eosin, Warthin-Starry silver and both were then examined under light microscope. A total of 12 specimens (three from each degree of the normal and I-III of NSD diagnosed and graded by histology) selected from the 44 patients were dehydrated, critical point dried, coated with gold palladium and examined under a JEOL JSM-30 scanning electron microscope (SEM) at 20 kV. RESULTS: According to the ultrastructural morphologic changes, non-specific duodenitis was divided into normal (as control group), mild, moderate and severe degrees according to results of SEM. The normal villi of duodenal bulb were less than 0.2 mm. There were inflammation cells, occasionally red blood cells and macrophages on the mucosal epithelial surface. Erosion and desquamation of epithelium could be seen. Three cases (25%, 3/12) had gastric metaplasia and Helicobacter pylori (H pylori) infection could be found in 5 cases (41.67%, 5/12) in duodenal bulb mucosa. The most distinctive feature was the ulcer-like defect on the surface of epithelial cells. CONCLUSION: Non-specific duodenitis is a separate entity disease caused by different factors. SEM is of value as an aid in the diagnosis of mucosal diseases of duodenum. PMID- 15655821 TI - Association between polymorphisms in the Toll-like receptor 4, CD14, and CARD15/NOD2 and inflammatory bowel disease in the Greek population. AB - AIM: Crohn's disease (CD) and ulcerative colitis (UC) are multifactorial diseases with a significant genetic background. Apart from CARD15/NOD2 gene, evidence is accumulating that molecules related to the innate immune response such as CD14 or Toll-like receptor 4 (TLR4), are involved in their pathogenesis. In further exploring the genetic background of these diseases, we investigated the variations in the CARD15/NOD2 gene (Arg702Trp, Gly908Arg and Leu1007fsinsC), and polymorphisms in the TLR4 gene (Asp299Gly and Thr399Ile) as well as in the promoter of the CD14 gene (T/C at position -159) in Greek patients with CD and UC. METHODS: DNA was obtained from 120 patients with CD, 85 with UC and 100 healthy individuals. Genotyping was performed by allele specific PCR or by PCR RFLP analysis. RESULTS: The 299Gly allele frequency of the TLR4 gene and the T allele and TT genotype frequencies of the CD14 promoter were significantly higher in CD patients only compared to healthy individuals (P = 0.026<0.05; P = 0.0048<0.01 and P = 0.047<0.05 respectively). Concerning the NOD2/CARD15 mutations the overall presence in CD patients was significantly higher than that in UC patients or in controls. Additionally, 51.67% of the CD patients were carriers of a TLR4 and/or CD14 polymorphic allele and at least one variant of the NOD2/CARD15, compared to 27% of the UC patients. It should be pointed out that both frequencies significantly increased as compared with the 10% frequency of multiple carriers found in healthy controls. A possible interaction of the NOD2/CARD15 with TLR4 and especially CD14, increased the risk of developing inflammatory bowel disease (IBD). CONCLUSION: Our results indicate that co existence of a mutation in either the TLR4 or CD14 gene, and in NOD2/CARD15 is associated with an increased susceptibility to developing CD compared to UC, and to developing either CD or UC compared to healthy individuals. PMID- 15655823 TI - Heme oxygenase-1 alleviates ischemia/reperfusion injury in aged liver. AB - AIM: To investigate if ischemia/reperfusion (I/R) injury in aged liver could be alleviated by heme oxygenase-1 (HO-1). METHODS: Three groups of SD rats (16 mo old) were studied. Group 1: control donors received physiological saline 24 h before their livers were harvested; group 2: donors were pretreated with hemin 24 h before their livers were harvested; and group 3: donors received hemin 24 h before their livers were harvested and zinc protoporphyrin (ZnPP, HO-1 inhibitor) was given to recipients at reperfusion. The harvested livers were stored in University of Wisconsin solution (4 degrees) for 6 h, and then transplanted to syngeneic rats. Serum glutamic oxaloacetic transaminase (SGOT), apoptotic cells, and apoptotic gene were measured 3, 6, 12, 24, 48 h after reperfusion. We measured the apoptotic index by TUNEL, determined the expression of antiapoptotic Bcl-2 and proapoptotic (caspase-3) gene products by Western blot. RESULTS: After 3, 6, 12, 24, and 48 h of reperfusion, the SGOT levels (584.4+/-85.8 u/L, 999.2+/ 125.2 u/L, 423.4+/-161.3 u/L, 257.8+/-95.8 u/L, and 122.4+/-26.4 u/L) in hemin group were significantly (all P<0.05) lower than those in saline group (1082.2+/ 101.2 u/L, 1775.2+/-328.3 u/L, 840.4+/-137.8 u/L, 448.6+/-74.3 u/L, and 306.2+/ 49.3 u/L). Liver HO-1 enzymatic activity correlated with beneficial effects of hemin and deleterious effects of adjunctive ZnPP treatment. Markedly less apoptotic (TUNEL+) liver cells 3, 6, 12, 24, and 48 h after reperfusion (5.16+/ 0.73, 10.2+/-0.67, 9.28+/-0.78, 7.14+/-1.12, and 4.78+/-0.65) (P<0.05) could be detected in hemin liver grafts, as compared to controls (7.82+/-1.05, 15.94+/ 1.82, 11.67+/-1.59, 8.28+/-1.09, and 6.36+/-0.67). We detected the increased levels of Bcl-2 (1.5-fold) expression and compared with saline controls. These differences were most pronounced at 12 h after transplantation. In contrast, an active form of proapoptotic caspase-3 (p20) protein was found to be 2.9-fold lower at 24 h in hemin-pretreated group, as compared to saline liver transplant controls. CONCLUSION: HO-1 overexpression can provide potent protection against cold I/R injury. This effect depends, at least in part, on HO-1-mediated inhibition of antiapoptotic mechanism. PMID- 15655824 TI - Enhanced production of leptin in gastric fundic mucosa with Helicobacter pylori infection. AB - AIM: To determine the concentrations of leptin in plasma and gastric fundic mucosa in humans, with reference to Helicobacter pylori (H pylori) infection, and their association with gastric mucosal levels of interleukin (IL)-1beta, IL-6 and IL-8. METHODS: Plasma leptin concentrations were determined in 135 outpatients with non-ulcer dyspepsia, consisting of 95 H pylori-infected and 40 uninfected subjects, and 13 patients before and after cure of the infection with anti-H pylori regimen. Using biopsy samples that were endoscopically obtained from the middle corpus along the greater curvature, gastric leptin contents were measured by radioimmunoassay and the mucosal concentrations of IL-1beta, IL-6 and IL-8 were measured by enzyme linked immunosorbent assay. We also analysed the expression of leptin in the fundic mucosa by reverse transcriptase-polymerase chain reaction (RT-PCR) and immunohistochemistry. RESULTS: The mucosal levels of leptin in the fundic mucosa of H pylori-infected patients were significantly higher than those of uninfected patients. The amount of gastric leptin correlated positively with the mucosal levels of IL-1beta and IL-6, but not IL-8. Circulating leptin correlated with body mass index, but not with H pylori status, and there was no change in plasma leptin levels following cure of the infection. Leptin immunoreactive cells were noted in the lower half of the fundic glands, and its expression of messenger ribonucleic acid in the oxyntic mucosa was detected by RT-PCR. CONCLUSION: Leptin production is enhanced in H pylori infected gastric mucosa. Gastric leptin may be involved in immune and inflammatory response during H pylori infection, through interaction with proinflammatory cytokines. PMID- 15655825 TI - Tumor vaccine against recurrence of hepatocellular carcinoma. AB - AIM: To investigate the effects of autologous tumor vaccine on recurrence of hepatocellular carcinoma (HCC). METHODS: Sixty patients with HCC who had undergone curative resection, were randomly divided into HCC vaccine group and control group. Three vaccinations at 2-wk intervals were performed after curative hepatic resection. Delayed-type- hypersensitivity (DTH) test was performed before and after vaccination. Primary endpoints were the time of recurrence. RESULTS: Four patients in control group and 6 patients in HCC vaccine group were withdrawn from the study. The vaccine containing human autologous HCC fragments showed no essential adverse effect in a phase II clinical trial and 17 of 24 patients developed a DTH response against the fragments. Three of 17 DTH-positive response patients and 5 of 7 DTH-negative response patients had recurrences after curative resection. After the operation, 1-, 2- and 3-year recurrence rates of HCC vaccine group were 16.7%, 29.2% and 33.3%, respectively. But, 1-, 2- and 3-year recurrence rates of the control group were 30.8%, 53.8% and 61.5%, respectively. The time before the first recurrence in the vaccinated patients was significantly longer than that in the control patients (P < 0.05). CONCLUSION: Autologous tumor vaccine is of promise in decreasing recurrence of human HCC. PMID- 15655826 TI - Tiam1 gene expression and its significance in colorectal carcinoma. AB - AIM: To explore the expression of Tiam1 gene in colorectal carcinoma and its correlation with tumor metastasis. METHODS: Expressions of Tiam1 gene in 8 colorectal carcinoma cell lines were detected by reverse transcriptase-polymerase chain reaction. In vitro invasiveness was determined by means of Matrigel invasion assay. The correlation of Tiam1 expression with the invasive ability was also analyzed. RESULTS: Tiam1 gene was highly expressed in LoVo and SW620, which were established from metastatic colorectal carcinomas in comparison with LS174T, SW480, HCT116, LST, HRT-18 and Hee8693, which were established from primary colorectal carcinomas. In vitro cell invasion demonstrated that LoVo and SW620 had a higher invasive ability than LS174T, SW480, HCT116, LST, HRT-18 and Hee8693. The expression of Tiam1 gene was highly related to the metastatic potential of colorectal carcinoma cells. CONCLUSION: Tiam1 gene may play an important role in invasion and metastasis of colorectal carcinoma and is a metastasis-related gene. PMID- 15655827 TI - Value of portal hemodynamics and hypersplenism in cirrhosis staging. AB - AIM: To determine the correlation between portal hemodynamics and spleen function among different grades of cirrhosis and verify its significance in cirrhosis staging. METHODS: The portal and splenic vein hemodynamics and spleen size were investigated by ultrasonography in consecutive 38 cirrhotic patients with cirrhosis (Child's grades A to C) and 20 normal controls. The differences were compared in portal vein diameter and flow velocity between patients with and without ascites and between patients with mild and severe esophageal varices. The correlation between peripheral blood cell counts and Child's grades was also determined. RESULTS: The portal flow velocity and volume were significantly lower in patients with Child's C (12.25+/-1.67 cm/s vs 788.59+/-234 mm/min, respectively) cirrhosis compared to controls (19.55+/-3.28 cm/s vs 1254.03+/-410 mm/min, respectively) and those with Child's A (18.5+/-3.02 cm/s vs 1358.48+/-384 mm/min, respectively) and Child's B (16.0+/-3.89 cm/s vs 1142.23+/-390 mm/min, respectively) cirrhosis. Patients with ascites had much lower portal flow velocity and volume (13.0+/-1.72 cm/s vs 1078+/-533 mm/min) than those without ascites (18.6+/-2.60 cm/s vs 1394+/-354 mm/min). There was no statistical difference between patients with mild and severe esophageal varices. The portal vein diameter was not significantly different among the above groups. There were significant differences in splenic vein diameter, flow velocity and white blood cell count, but not in spleen size, red blood cell and platelet counts among the various grades of cirrhosis. The spleen size was negatively correlated with red blood cell and platelet counts (r = -0.620 and r = -0.8.34, respectively). CONCLUSION: An optimal system that includes parameters representing the portal hemodynamics and spleen function should be proposed for cirrhosis staging. PMID- 15655828 TI - Existence and significance of hepatitis B virus DNA in kidneys of IgA nephropathy. AB - AIM: To investigate the existence and significance of hepatitis B virus (HBV) DNA in the pathogenesis of IgA nephropathy (IgAN). METHODS: Fifty cases of IgAN with HBV antigenaemia and/or hepatitis B virus antigens (HBAg, or HBsAg, HBcAg) detected by immunohistochemistry in renal tissues were enrolled in our study. The distribution and localization of HBV DNA were observed using in situ hybridization. Southern blot analysis was performed to reveal the state of renal HBV DNA. RESULTS: Among the 50 patients with IgAN, HBs antigenemia was detected in 17 patients (34%). HBAg in renal tissues was detected in 48 patients (96%), the positive rate of HBAg, HBsAg, and HBcAg was 82% (41/50), 58% (29/50), and 42% (21/50) in glomeruli, respectively; and was 94% (47/50), 56% (28/50) and 78% (39/50) in tubular epithelia, respectively. Positive HBV DNA was detected in 72% (36/50) and 82% (41/50) cases in tubular epithelia and glomeruli respectively by in situ hybridization, and the positive signals were localized in the nuclei of tubular epithelial cells and glomerular mesangial cells as well as infiltrated interstitial lymphocytes. Moreover, 68% (34/50) cases were proved to be HBV DNA positive by Southern blot analysis, and all were the integrated form. CONCLUSION: HBV infection might play an important role in occurrence and progress of IgAN. In addition to humoral immune damages mediated by HBAg-HBAb immune complex, renal tissues of some IgAN are directly infected with HBV and express HBAg in situ, and the cellular mechanism mediated by HBV originating from renal cells in situ may also be involved in the pathogenesis of IgAN. PMID- 15655829 TI - Pharmacokinetics of C-1027 in mice as determined by TCA-RA method. AB - AIM: To validate a radioactivity assay, the TCA-RA method, for the measurement of C-1027 in serum and to evaluate its application in determination of pharmacokinetics of C-1027 in mice. METHODS: (125)I-C-1027 was prepared by the Iodogen method and separated by HPLC. The radioactivity assay was established and used to determine (125)I-C-1027 in mice at doses of 10, 50 and 100 microg/kg after precipitation with 20% trichloroacetic acid (TCA-RA method). Several pharmacokinetic parameters were determined after intravenous injection of (125)I C-1027 to mice. RESULTS: After intravenous injection of (125)I-C-1027 to mice, at doses of 10, 50 and 100 microg/kg; the apparent distribution volumes (V(d)) were 0.26, 0.31 and 0.33 L/kg; the biological half-lives (T(1/2)) were 3.10, 3.40 and 3.90 h; the areas under curve (AUC) were 18.41, 103.69 and 202.74 ng/h/mL; the elimination rate constants (K) were 1.04, 1.26 and 0.58/h; and the total body clearance (Cl) were 0.54, 0.48 and 0.49 L/kg/h, respectively. CONCLUSION: TCA-RA is a sensitive, reliable and suitable method for the determination of (125)I-C 1027 in mouse serum. PMID- 15655831 TI - Effect of mutated IkappaBalpha transfection on multidrug resistance in hilar cholangiocarcinoma cell lines. AB - AIM: To explore the expression effect of mutated IkappaBalpha transfection on multidrug resistance gene (MDR-1) in hilar cholangiocarcinoma cells by inhibiting the activity of nuclear transcription factor-kappaB (NF-kappaB). METHODS: We used the mutated IkappaBalpha plasmid to transfect QBC(939)HCVC+ cells and QBC939 cells, and electrophoretic gel mobility shift assay (EMSA) to detect the binding activity of NF-kappaB DNA and the effect of the transfecting mutated IkappaBalpha plasmid on multidrug resistance gene (MDR-1) in hilar cholangiocarcinoma cells and its expression protein (P-GP). RESULTS: Plasmid DNA was digested by restriction enzymes Xbal and Hand III, and its product after electrophoresis showed two bands with a big difference in molecular weight, with a size of 4.9 kb and 1.55 kb respectively, which indicated that the carrier was successfully constructed and digested with enzymes. The radioactivity accumulation of QBC(939)HCVC+ and QBC939 cells transfected with mutated IkappaBalpha plasmid was significantly lower than that of the control group not transfected with mutated IkappaBalpha plasmid. Double densimeter scanning showed that the relative signal density between the tansfection group and non-transfection group was significantly different, which proved that the mutated IkappaBalpha plasmid could inhibit the binding activity of NF-kappaB DNA in hilar cholangiocarcinoma cells. Compared to control group not transfected with m IkappaBalpha plasmid, the expression level of MDR-1mRNA in the QBC939 and QBC939HCVC+ cells transfected with mutated IkappaBalpha plasmid was lower. The expression intensity of P-GP protein in QBC939 and QBC939HCVC+ cells transfected with mutated IkappaBalpha was significantly lower than that of the control group not transfected with mutated IkappaBalpha plasmid. CONCLUSION: The mutated IkappaBalpha plasmid transfection can markedly reverse the multidrug resistance of hilar cholangiocarcinoma cells. Interruption of NF-kappaB activity may become a new target in gene therapy for hilar cholangiocarcinogenesic carcinoma. PMID- 15655830 TI - Correlation between expression of gastrin, somatostatin and cell apoptosis regulation gene bcl-2/bax in large intestine carcinoma. AB - AIM: To explore the correlation between expression of somatostatin (SS), gastrin (GAS) and cell apoptosis regulation gene bcl-2/bax in large intestine carcinoma. METHODS: Sixty-two large intestine cancer tissue samples were randomly and retrospectively selected from patients with large intestine carcinoma. Immunohistochemical staining for bcl-2, bax, GAS, SS was performed according to the standard streptavidin-biotin-peroxidase (S-P) method. According to the semi quantitative integral evaluation, SS and GAS were divided into three groups as follows. Scores 1-3 were defined as the low expression group, 4-8 as the intermediate expression group, 9-16 as the high expression group. Bax and bcl-2 protein expressions in different GAS and SS expression groups of large intestine carcinoma were assessed. RESULTS: The positive expression rate of bax had a prominent difference between SS and GAS high, intermediate and low expression groups (P<0.05, chi(2)(SS) = 9.246; P<0.05, chi(2)(GAS) = 6.981). The positive expression rate of bax in SS high (80.0%, 8/10) and intermediate (76.5%, 13/17) expression groups was higher than that in low expression group (40.0%, 14/35) (P<0.05, chi(2)( high vs low ) = 5.242; P<0.05, chi(2)( middle vs low ) = 6.097). The positive expression rate of bax in GAS high expression group (27.3%, 3/8) was lower than that in low expression group (69.4%, 25/36) (P<0.05, chi(2) = 4.594). However, bax expression in GAS intermediate expression group (46.7%, 7/15) was lower than that in low expression group, but not statistically significant. The positive expression rate of bcl-2 had a prominent difference between SS and GAS high, intermediate and low expression groups (P<0.05, chi(2)(SS) = 7.178; P<0.05, chi(2)( GAS ) = 13.831). The positive expression rate of bcl-2 in GAS high (90.9%, 10/11) and intermediate (86.7%, 13/15) expression groups was higher than that in low expression group (44.4%, 16/36) (P<0.05, chi(2)( high vs low ) = 5.600; P<0.05, chi(2)( middle vs low ) = 7.695). However, the positive expression rate of bcl-2 in SS high (40.0%, 4/10) and intermediate (47.1%, 8/9) expression groups was lower than that in low expression group (77.1%, 27/35) (P<0.05, chi(2)( high vs low ) = 4.710; P<0.05, chi(2)( middle vs low ) = 4.706). There was a significant positive correlation between the integral ratio of GAS to SS and the integral of bcl-2 (P<0.01, r = 0.340). However, there was a negative correlation between the integral ratio of GAS to the SS and bax the integral of (P<0.05, r = -0.299). CONCLUSION: The regulation and control of gastrin, somatostatin in cell apoptosis of large intestine carcinoma may be directly related to the abnormal expression of bcl-2, bax. PMID- 15655832 TI - Treatment of pancreatic pseudocysts in line with D'Egidio's classification. AB - AIM: To explore the implications of underlying diseases in treatment of pancreatic pseudocysts (PPC). METHODS: Clinical data of 73 cases of pancreatic pseudocyst treated in a 12-year period were reviewed comprehensively. Pancreatic pseudocysts were classified according to the etiological criteria proposed by D'Egidio. The correlation between the etiological classification, measure of treatment and clinical outcome of the patients was analyzed. RESULTS: According to the etiological criteria proposed by D'Egidio, 73 patients were divided into three groups. Group I was comprised of 37 patients with type I pseudocyst, percutaneous drainage was successful in the majority (9/11, 82%) while external or internal drainage was not satisfactory with a low success rate (8/16, 50%). Group II was comprised of 24 patients with type II pseudocyst, and internal drainage was curative for most of the cases (11/12, 92%), but the success rate of percutaneous or external drainage was unacceptably low (4/9, 44%). Group III consisted of 12 patients with type III pseudocyst. Internal drainage or pancreatic resection performed in 10 of these patients produced a curative rate of 80% (8/10) with the correction of the ductal pathology as a prerequisite. CONCLUSION: The classification of pancreatic pseudocyst based on its underlying diseases is meaningful for its management. Awareness of the underlying diseases of pancreatic pseudocyst and detection of the ductal pathology in type II and III pancreatic pseudocysts with endoscopic retrograde cholangiopancreatography may help make better decisions of treatment to reduce the rate of complications and recurrence. PMID- 15655833 TI - Effect of glutamate on inflammatory responses of intestine and brain after focal cerebral ischemia. AB - AIM: To study the modulation of glutamate on post-ischemic intestinal and cerebral inflammatory responses in a ischemic and excitotoxic rat model. METHODS: Adult male rats were subjected to bilateral carotid artery occlusion for 15 min and injection of monosodium glutamate intraperitoneally, to decapitate them at selected time points. Tumor necrosis factor alpha (TNF-alpha) level and nuclear factor kappa B (NF-kappaB) activity were determined by enzyme-linked immunosorbant assay (ELISA) and electrophoretic mobility shift assay (EMSA), respectively. Hemodynamic parameters were monitored continuously during the whole process of cerebral ischemia and reperfusion. RESULTS: Monosodium glutamate (MSG) treated rats displayed statistically significant high levels of TNF-alpha in cerebral and intestinal tissues within the first 6 h of ischemia. The rats with cerebral ischemia showed a minor decrease of TNF-alpha production in cerebral and intestinal tissues. The rats with cerebral ischemia and treated with MSG displayed statistically significant low levels of TNF-alpha in cerebral and intestinal tissues. These results correlated significantly with NF-kappaB production calculated at the same intervals. During experiment, the mean blood pressure and heart rates in all groups were stable. CONCLUSION: Glutamate is involved in the mechanism of intestinal and cerebral inflammation responses. The effects of glutamate on cerebral and intestinal inflammatory responses after ischemia are up-regulated at the transcriptional level, through the NF-kappaB signal transduction pathway. PMID- 15655834 TI - Effects of psychological stress on small intestinal motility and expression of cholecystokinin and vasoactive intestinal polypeptide in plasma and small intestine in mice. AB - AIM: To investigate the effects of psychological stress on small intestinal motility and expression of cholecystokinin (CCK) and vasoactive intestinal polypeptide (VIP) in plasma and small intestine, and to explore the relationship between small intestinal motor disorders and gastrointestinal hormones under psychological stress. METHODS: Thirty-six mice were randomly divided into psychological stress group and control group. A mouse model with psychological stress was established by housing the mice with a hungry cat in separate layers of a two-layer cage. A semi-solid colored marker (carbon-ink) was used for monitoring small intestinal transit. CCK and VIP levels in plasma and small intestine in mice were measured by radioimmunoassay (RIA). RESULTS: Small intestinal transit was inhibited (52.18+/-19.15% vs 70.19+/-17.79%, P<0.01) in mice after psychological stress, compared to the controls. Small intestinal CCK levels in psychological stress mice were significantly lower than those in the control group (0.75+/-0.53 microg/g vs 1.98+/-1.17 microg/g, P<0.01), whereas plasma CCK concentrations were not different between the groups. VIP levels in small intestine were significantly higher in psychological stress mice than those in the control group (8.45+/-1.09 microg/g vs 7.03+/-2.36 microg/g, P<0.01), while there was no significant difference in plasma VIP levels between the two groups. CONCLUSION: Psychological stress inhibits the small intestinal transit, probably by down-regulating CCK and up-regulating VIP expression in small intestine. PMID- 15655835 TI - Tumor angiogenesis and its clinical significance in pediatric malignant liver tumor. AB - AIM: To investigate the expression of vascular endothelial growth factor (VEGF) and microvascular density (MVD) count in pediatric malignant liver tumor and their clinical significances. METHODS: Fourteen children with malignant liver tumors including seven hepatocellular carcinomas (HCCs), five hepatoblastomas, one malignant mesenchymoma and one rhabdomyosarcoma were studied. Twelve adult HCC samples served as control group. All samples were examined with streptavidin biotin peroxidase (SP) immunohistochemical staining for VEGF expression and MVD count. RESULTS: VEGF positive expression in all pediatric malignant liver tumors was significantly higher than that in adult HCC (0.4971+/-0.14 vs 0.4027+/-0.03, P<0.05). VEGF expression in pediatric HCC group was also markedly higher than that in adult HCC group (0.5665+/-0.10 vs 0.4027+/-0.03, P<0.01) and pediatric non-HCC group (0.5665+/-0.10 vs 0.4276+/-0.15, P<0.05). The mean value of MVD in pediatric malignant liver tumors was significantly higher than that in adult HCC (33.66+/-12.24 vs 26.52+/-4.38, P<0.05). Furthermore, MVD in pediatric HCC group was significantly higher compared to that in adult HCC group (36.94+/-9.28 vs 26.52+/-4.38, P<0.05), but there was no significant difference compared to the pediatric non-HCC group (36.94+/-9.28 vs 30.37+/-14.61, P>0.05). All 7 children in HCC group died within 2 years, whereas the prognosis in pediatric non-HCC group was better, in which two patients survived more than 5 years. CONCLUSION: Children with malignant liver tumors, especially with HCC, may have extensive angiogenesis that induces a rapid tumor growth and leads to a poor prognosis. PMID- 15655837 TI - Metastasis of primary gallbladder carcinoma in lymph node and liver. AB - AIM: To evaluate the patterns with metastasis of gallbladder carcinoma in lymph nodes and liver. METHODS: A total of 45 patients who had radical surgery were selected. The patterns with metastasis of primary gallbladder carcinoma in lymph nodes and liver were examined histopathologically and classified as TNM staging of the American Joint Committee on Cancer. RESULTS: Of the 45 patients, 29 (64.4%) had a lymph node positive disease and 20 (44.4%) had a direct invasion of the liver. The frequency of involvement of lymph nodes was strongly influenced by the depth of the primary tumor (P = 0.0001). The postoperative survival rate of patients with negative lymph node metastasis was significantly higher than that of patients with positive lymph node metastasis (P = 0.004), but the postoperative survival rate of patients with N1 lymph node metastasis was not significantly different from that of patients with N2 lymph node metastasis (P = 0.3874). The postoperative survival rate of patients without hepatic invasion was significantly better than that of patients with hepatic invasion (P = 0.0177). CONCLUSION: Complete resection of the regional lymph nodes is important in advanced primary gallbladder carcinoma (PGC). The initial sites of liver spread are located mostly in segments IV and V. It is necessary to achieve negative surgical margins 2 cm from the tumor. In patients with hepatic hilum invasion, extended right hepatectomy with or without bile duct resection or portal vein resection is necessary for curative resection. PMID- 15655836 TI - Association of cyclin D1, p16 and retinoblastoma protein expressions with prognosis and metastasis of gallbladder carcinoma. AB - AIM: To investigate the role of cyclin D1, p16 and retinoblastoma in cancerous process of gallbladder carcinomas and to assess the relation between cyclin D1, p16, Rb and the biological characteristics of gallbladder carcinoma. METHODS: Forty-one gallbladder carcinoma, 7 gallbladder adenoma and 14 chronic cholecystitis specimens were immunohistochemically and histopathologically investigated for the relation of cyclin D1, p16 and Rb with Nevin staging and pathologic grading. RESULTS: The expression rates of abnormal cyclin D1 in gallbladder carcinoma (68.3%) and gallbladder adenoma (57.1%) were significantly higher than those in chronic cholecystitis (7.1%) (P<0.05). No significant difference was found both among the pathological grades G(1), G(2) and G(3) and among Nevin stagings S(1)-S(2), S(3) and S(4)-S(5) of gallbladder carcinoma. The positive rates of p16 (48.8%) and Rb (58.5%) in gallbladder carcinoma were significantly lower compared to those in adenoma (100.0%) and cholecystitis (100.0%) (P<0.05). The positive rates of p16 and Rb in Nevin stagings S(1)-S(2) (80.0% and 90.0%) and S(3) (46.2% and 61.5%) gallbladder carcinomas were significantly higher than those in S(4)-S(5) (33.3% and 38.8%) (P<0.05), and those in pathologic grades G(1) (54.5% and 81.8%) and G(2) (50.0% and 62.5%) gallbladder carcinoma were significantly higher than those in G(3) (28.6% and 35.7%) (P<0.05). The protein expression of p16 and Rb had a negative-correlation in gallbladder carcinoma (r = -0.2993, P<0.05), and this negative-correlation was correlated with Nevin staging (P<0.05). Moreover, the protein expression of p16 and cyclin D1 had a negative-correlation in gallbladder carcinoma (r = -0.9417, P<0.05). CONCLUSION: Cyclin D1 may play a role in the early stage of gallbladder carcinoma. Mutation of p16 and Rb genes might be correlated with progression of gallbladder carcinoma. Analysis of p16 and Rb can estimate the prognosis of gallbladder carcinoma. Expression of p16 and Rb may be correlated with Nevin staging and pathologic grading in gallbladder carcinoma. PMID- 15655838 TI - Role of PGI2 in the formation and maintenance of hyperdynamic circulatory state of portal hypertensive rats. AB - AIM: To investigate the role of prostacyclin (PGI(2)) and nitric oxide (NO) in the development and maintenance of hyperdynamic circulatory state of chronic portal hypertensive rats. METHODS: Ninety male Sprague-Dawley rats were divided into three groups: intrahepatic portal hypertension (IHPH) group by injection of CCl(4), prehepatic portal hypertension (PHPH) group by partial stenosis of the portal vein and sham-operation control (SO) group. One week after the models were made, animals in each group were subdivided into 4 groups: saline controlled group (n = 23), Nomega-nitro-L-arginine (L-NNA) group (n = 21) group, indomethacin (INDO) group (n = 22) and high-dose heparin group (n = 24). The rats were administrated 1 mL of saline, L-NNA (3.3 mg/kg.d) and INDO (5 mg/kg.d) respectively through gastric tubes for one week, then heparin (200 IU/Kg/min) was given to rats by intravenous injection for an hour. Splanchnic and systemic hemodynamics were measured using radioactive microsphere techniques. The serum nitrate/nitrite (NO(2)(-)/NO(3)(-)) levels as a marker of production of NO were assessed by a colorimetric method, and concentration of 6-keto-PGF1alpha, a stable hydrolytic product of PGI(2), was determined by radioimmunoassay. RESULTS: The concentrations of plasma 6-keto-PGF1alpha (pg/mL) and serum NO(2)(-)/NO(3)(-) (micromol/L) in IHPH rats (1123.85+/-153.64, 73.34+/-4.31) and PHPH rats (891.88+/-83.11, 75.21+/-6.89) were significantly higher than those in SO rats (725.53+/-105.54, 58.79+/-8.47) (P<0.05). Compared with SO rats, total peripheral vascular resistance (TPR) and spanchnic vascular resistance (SVR) decreased but cardiac index (CI) and portal venous inflow (PVI) increased obviously in IHPH and PHPH rats (P<0.05). L-NNA and indomethacin could decrease the concentrations of plasma 6-keto-PGF1alpha and serum NO(2)(-)/NO(3)(-) in IHPH and PHPH rats (P<0.05). Meanwhile, CI, FPP and PVI lowered but MAP, TPR and SVR increased (P<0.05). After deduction of the action of NO, there was no significant correlation between plasma PGI(2) level and hemodynamic parameters such as CI, TPR, PVI and SVR. However, after deduction of the action of PGI(2), NO still correlated highly with the hemodynamic parameters, indicating that there was a close correlation between NO and the hemodynamic parameters. After administration of high-dose heparin, plasma 6-keto-PGF(1alpha) concentrations in IHPH, PHPH and SO rats were significantly higher than those in rats administrated vehicle (P<0.05). On the contrary, levels of serum NO(2)(-)/NO(3)(-) in IHPH, PHPH and SO rats were significantly lower than those in rats administrated Vehicle (P<0.05). Compared with those rats administrated vehicle, the hemodynamic parameters of portal hypertensive rats, such as CI and PVI, declined significantly after administration of high-dose heparin (P<0.05), while TPR and SVR increased significantly (P<0.05). CONCLUSION: It is NO rather than PGI(2) that is a mediator in the formation and maintenance of hyperdynamic circulatory state of chronic portal hypertensive rats. PMID- 15655839 TI - Knockdown of survivin gene expression by RNAi induces apoptosis in human hepatocellular carcinoma cell line SMMC-7721. AB - AIM: To investigate the survivin gene expression in human hepatocellular carcinoma cell line SMMC-7721 and the effects of survivin gene RNA interference (RNAi) on cell apoptosis and biological behaviors of SMMC-7721 cells. METHODS: Eukaryotic expression vector of survivin gene RNAi and recombinant plasmid pSuppressorNeo-survivin (pSuNeo-SVV), were constructed by ligating into the vector, pSuppressorNeo (pSuNeo) digested with restriction enzymes Xba I and Sal I and the designed double-chain RNAi primers. A cell model of SMMC-7721 after treatment with RNAi was prepared by transfecting SMMC-7721 cells with the lipofectin transfection method. Strept-avidin-biotin-complex (SABC) immunohistochemical staining and RT-PCR were used to detect survivin gene expressions in SMMC-7721 cells. Flow cytometry was used for the cell cycle analysis. Transmission electron microscopy was performed to determine whether RNAi induced cell apoptosis, and the method of measuring the cell growth curve was utilized to study the growth of SMMC-7721 cells before and after treatment with RNAi. RESULTS: The eukaryotic expression vector of survivin gene RNAi and pSuNeo-SVV, were constructed successfully. The expression level of survivin gene in SMMC-7721 cells was observed. After the treatment of RNAi, the expression of survivin gene in SMMC-7721 cells was almost absent, apoptosis index was increased by 15.6%, and the number of cells was decreased in G2/M phase and the cell growth was inhibited. CONCLUSION: RNAi can exert a knockdown of survivin gene expression in SMMC-7721 cells, and induce apoptosis and inhibit the growth of carcinoma cells. PMID- 15655840 TI - Effect of bone marrow-derived monocytes transfected with RNA of mouse colon carcinoma on specific antitumor immunity. AB - AIM: To investigate the effect of bone marrow-derived monocytes transfected with RNA of CT-26 (a cell line of mouse colon carcinoma) on antitumor immunity. METHODS: Mouse bone marrow-derived monocytes were incubated with mouse granulocyte macrophage colony stimulating factor (mGM-CSF) in vitro, and the purity of monocytes was detected by flow cytometry. Total RNA of CT-26 was obtained by TRIzol's process, and monocytes were transfected by TransMessenger in vitro. The activity of cytotoxic T lymphocytes (CTL) in vivo was estimated by the modified lactate dehydrogenase (LDH) release assay. Changes of tumor size in mice and animal's survival time were observed in different groups. RESULTS: Monocytes from mouse bone marrow were successfully incubated, and the positive rate of CD11b was over 95%. Vaccination of the monocytes transfected with total RNA induced a high level of specific CTL activity in vivo, and made mice resistant to the subsequent challenge of parental tumor cells. In vivo effects induced by monocytes transfected with total RNA were stronger than those induced by monocytes pulsed with tumor cell lysates. CONCLUSION: Antigen presenting cells transfected with total RNA of CT-26 can present endogenous? tumor antigens, activate CTL, and effectively induce specific antitumor immunity. PMID- 15655843 TI - The weight of giants. By Caveman. PMID- 15655842 TI - Hematemesis as the initial complication of pancreatic adenocarcinoma directly invading the duodenum: a case report. AB - Pancreatic carcinoma is a debilitating disease and carries a poor prognosis. It is a rare cause of upper gastrointestinal bleeding, even though pancreas, stomach, duodenum and jejunum are adjacent organs. The incidence of pancreatic adenocarcinoma directly invading the gastrointestinal tract leading to gastrointestinal hemorrhage is very low, and most of them present with melena and hematochezia. Here, we describe one unique case manifesting characteristically severe and unremitting hematemesis as an initial presentation of pancreatic adenocarcinoma. This tumor directly invaded the duodenal mucosa as a bleeding protruding tumor mass. Our MEDLINE search has confirmed that this is the first reported case with an initial manifestation of hematemesis from pancreatic adenocarcinoma in Asians. Pancreatic adenocarcinoma directly invading duodenum complicated by hemorrhage can be a rare cause of hematemesis, and clinicians should be reminded of it while they are making differential diagnosis. PMID- 15655841 TI - Acquired hepatocerebral degeneration: a case report. AB - AIM: Acquired hepatocerebral degeneration (AHD) is an exceptional type of hepatic encephalopathies (HE). It is characterized by neuropsychiatric and extrapyramidal symptomatology similar to that seen in hepatolenticular degeneration (Wilson's disease). In this paper, we report a case of AHD with unusual presenting features. METHODS: A 28-year-old man with AHD was described and the literature was reviewed. RESULTS: The man had a history of HBV-related liver cirrhosis. He was admitted to our hospital with apathy, dysarthria, mild consciousness impairment and extrapyramidal symptoms after hematemesis. By review of the literature, cases with AHD often did not present consciousness impairment. So our case was once diagnosed incorrectly as Wilson's disease. CONCLUSION: AHD is a rare syndrome and its variable clinical manifestations make it difficult to be diagnosed. But we believe that extensive examination and thorough understanding of the disease are beneficial to a correct diagnosis. Moreover, biocoene is effective in treating the case. PMID- 15655845 TI - How good is virtual colonoscopy? While effective at finding most lesions, this tool might work best when used with conventional colonoscopy. PMID- 15655846 TI - Nerve stimulation boosts quality of life in people with chronic urinary problems. PMID- 15655847 TI - Humor and the brain. PMID- 15655848 TI - Therapy over the phone beats the blues. PMID- 15655849 TI - Injections offer insight into Alzheimer's disease. PMID- 15655850 TI - Overweight? Here's how to have a successful knee replacement. Develop a healthy diet prior to surgery, so excess weight doesn't ruin your new joint. PMID- 15655852 TI - The mental effects of bad bone breaks. PMID- 15655851 TI - Vitamin C may help prevent arthritis. PMID- 15655853 TI - Wanted: fewer side effects. PMID- 15655854 TI - Radiofrequency procedure blasts kidney tumors. It may help the elderly, but many questions remain. PMID- 15655855 TI - Patients undergoing cancer treatments need monitoring to prevent heart injuries. PMID- 15655856 TI - Cognitive decline in breast cancer patients. PMID- 15655857 TI - Low "good" cholesterol may flag breast cancer risk. PMID- 15655858 TI - Can vitamin therapy prevent restenosis? Conflicting studies indicate the jury is still out on folate therapy after angioplasty. PMID- 15655859 TI - Doubling Zocor dose fails to double its benefit. PMID- 15655860 TI - Coumadin and ginseng: a potentially dangerous combination. PMID- 15655861 TI - Most natural hypertension treatments are not helpful. PMID- 15655862 TI - Reigning in low blood sugar. If you're experiencing hypoglycemia without warning signs, you can train your body to start showing them. PMID- 15655863 TI - Diabetes increases risk for premature death in older adults. But you can take steps to prolong both your lifespan and your quality of life. PMID- 15655864 TI - Metabolic syndrome. Losing weight and exercising can dramatically reduce your risk of dying from this cumulative disease. PMID- 15655865 TI - Fish slows heart disease. PMID- 15655866 TI - Peanuts lower incidence of gall bladder surgery. PMID- 15655867 TI - Which is best: intermittent or continuous exercise? PMID- 15655868 TI - I have coronary artery disease and osteoarthritis. What can you tell me about COX 2 inhibitors and their possible effects on my heart? PMID- 15655869 TI - Are there new ways to test my blood for ovarian cancer? PMID- 15655870 TI - National Institutes of Health unveils open access draft. PMID- 15655871 TI - Researchers identify better hepatitis C treatment for people with HIV. PMID- 15655872 TI - National Institutes of Health's ClinicalTrials.gov Web site wins prestigious award. PMID- 15655873 TI - NIH awards $19 million to University of Alaska. PMID- 15655874 TI - Scientists discover new approaches to manipulating AIDS virus. PMID- 15655875 TI - Hiroshni Nikaido, MD, of University of California, Berkeley wins Bristol-Meyers Squibb Infectious Diseases Research Award. PMID- 15655876 TI - NIAID expands West Nile treatment trials. PMID- 15655877 TI - Oral health and diabetes: what's the link? PMID- 15655878 TI - EFRACom calls for 'significant improvements' to the draft Animal Welfare Bill. PMID- 15655879 TI - BVA to emphasise positive leadership role. PMID- 15655880 TI - [In memory of Dr. Francis Crick--paradigm in theoretical biology]. PMID- 15655881 TI - Health status of the uninsured: opportunities for improvement. PMID- 15655882 TI - Asia, a melting pot for peritoneal dialysis practices: redefining and expanding peritoneal dialysis horizons in Asia. PMID- 15655884 TI - [Benign or malignant jugular vein thrombosis? Differentiation with colour-doppler sonography]. PMID- 15655883 TI - [Endomorphy in childhood and adolescence as predictor of aerobic capacity]. PMID- 15655885 TI - [Guideline for managing suspect and positive cytologic smears of the uterine cervix (revised form, version 2.4)]. PMID- 15655886 TI - Nociception in vertebrates. Proceedings of a symposium at the Society for Experimental Biology. April 1-2, 2004. PMID- 15655887 TI - New perspectives of research on wine and health. Proceedings of an international congress on wine and health. May 8-11, 2003. Marsala, Italy. PMID- 15655888 TI - A way to improve the reproducibility of measurements of myocardial velocity gradients. PMID- 15655889 TI - Long-term survival of patients with single ventricle; which imaging modality? PMID- 15655890 TI - Commentary: a brief history of geriatrics. PMID- 15655891 TI - Commentary: a brief history of geriatrics. PMID- 15655892 TI - Commentary: a brief history of geriatrics. PMID- 15655893 TI - Commentary: a brief history of geriatrics. PMID- 15655894 TI - Commentary: a brief history of geriatrics. PMID- 15655895 TI - Structure function relationship among alpha-crystallin related small heat shock proteins. AB - A sequence alignment is presented which permits the detection of the sequence and structural homology among alpha-crystallin subunits, alphaA and alphaB, and distantly related sHsps, MjHsp16.5 and wheat Hsp16.9. This alignment shows that homology extends beyond the alpha-crystallin domain. Variations in the polydisperse quaternary structure appear, in part, dependent upon the N-terminal 18 and 19 amino acids that are essential for subunit interactions in polydisperse sHsps. The hydrophobic sequence that follows these N-terminal amino acids shares a number of aromatic amino acids and has significant homology with MjHsp16.5. In the second exon of alpha-crystallin, sequence homology is concentrated in a region with chaperone and ANS binding sites. It is clear that the binding site for ANS and its derivative, bis-ANS, requires both positively charged amino acids and hydrophobic interactions. Therefore, its binding is not a true measure of hydrophobic surface exposure. The limited homology and secondary structure in the following C-terminal sequences is related to the pattern of association of other sHsp subunits and/or functional differences. Our study suggests that alphaA has evolved in the lens to chaperone exposed beta-sheet edges of the betagamma crystallins and their proteolytic fragments. Also, both time and a harsh environment such as that in the lens interior, beta-sheet proteins would naturally generate beta-sheet edges. The interaction between such edges results in insoluble, abnormal protein aggregation and in the lens, light scattering elements that cause cataract. PMID- 15655896 TI - Support from hospital to home. PMID- 15655897 TI - [Personality disorders beneath chronic depression--narcissistic depression and schizoid depression]. PMID- 15655898 TI - [Therapeutic strategy for drug resistant depression]. PMID- 15655899 TI - [Therapeutic and pharmacological effects of second-generation antipsychotics on drug-resistant bipolar depression]. PMID- 15655900 TI - [Biological pathophysiology of mood disorders special reference to the neurotransmitter receptors]. PMID- 15655901 TI - [Diagnosis and physiopathology of depression using DNA chip]. PMID- 15655902 TI - [Front line of therapies for depression]. PMID- 15655903 TI - Proceedings of the Third ITI (International Team for Implantology) Consensus Conference. Gstaad, Switzerland, August 2003. PMID- 15655904 TI - [Molecular analytical evaluation of macrolide- and ketolide-resistant Streptococcus pneumoniae--mechanism of action of telithromycin and resistance to it]. AB - PROTEKT (Prospective Resistant Organism Tracking and Epidemiology for the Ketolide Telithromycin) is a worldwide epidemiologic survey for investigating drug susceptibility against major bacterial pathogens in respiratory tract infections, and that is also designed to identify the action mechanism of telithromycin (TEL), a ketolide antibacterial agent, on the resistant Streptococcus pneumoniae and the resistance mechanism for TEL on the TEL resistant S. pneumoniae strain, in addition to determine macrolide/ketolide resistant S. pneumoniae activities of TEL using molecular analysis. TEL exerted the antibacterial action on the macrolide-resistant S. pneumoniae regardless maintaining the macrolide-resistant mechanism and exhibited the potent antibacterial activity against all of ermB gene-positive strains, mefA gene positive strains and ribosome variants. This result was considered to reflect the fact that TEL did not induce resistance to ermB and had extremely low ability to select resistant strain by mutation. These actions of TEL were considered to be derived from its novel chemical structure and might be characteristics of ketolides not possessed by macrolides. In the survey of PROTEKT in 1999 to 2002, among 13,864 strains of S. pneumoniae isolated worldwide, ketolide-resistant strain (TEL MIC > or = 4 microg/ml) was observed in 10 strains (0.07%). MIC of these 10 strains was 4 or 8 microg/mL and all of these strains were ermB-positive strains. Based on this fact, potential involvement of adenine demethylase (ermB gene product) was considered in the background of development of ketolide resistant S. pneumoniae. PMID- 15655905 TI - [Antimicrobial susceptibility of Pseudomonas aeruginosa isolated at Kochi Municipal Central Hospital in the last 3 years]. AB - Antimicrobial susceptibility of Pseudomonas aeruginosa isolated at Kochi Municipal Central Hospital between 2001 and 2003 was assessed according to the NCCLS interpretive criteria. 1. The piperacillin-susceptible rate was 92.9%. 2. Among cephem antibiotics, the ceftazidime-susceptible rate was the highest (96.0%). 3. As for aminoglycosides, susceptibility to tobramycin and amikacin remained with a susceptible rate of 93.2% and 94.8%, respectively. 4. The carbapenem-susceptibility remained high. The susceptible rate for meropenem (94.1%) was higher than that for imipenem (88.3%). 5. Acquisition of resistance was observed in urinary isolates. Four multi-drug resistant P. aeruginosa, which are resistant to all of imipenem, amikacin and ofloxacin were isolated in this study and all were isolated from urine. 6. Of 388 isolates, 34 isolates were resistant to imipenem, but no positive isolate was found in screening of metallo beta-lactamase-producing bacteria. PMID- 15655906 TI - [Bactericial activity of chlorhexidine gluconate against recent clinical isolates of various bacterial species in Japan]. AB - The minimum inhibitory concentration (MIC) and bactericidal activity of chlorhexidine gluconate (CHG) were determined for methicillin-susceptible Staphylococcus aureus (MSSA), methicillin-resistant S. aureus (MRSA), Escherichia coli, Serratia marcescens, Eterobacter cloacae, Pseudomonas aeruginosa and Burkholderia cepacia, isolated from patients in medical institutions all over Japan between 2000 and 2002. The following findings were obtained. 1. The MICs of CHG against MSSA, MRSA, E. coli and B. cepacia were 0.002% or less, and those against S. marcescens, E. cloacae and P. aeruginosa were 0.008% or less. 2. Rapid and strong bactericidal effects of CHG were observed against all clinical isolates of E. coli, E. cloacae and P. aeruginosa tested even at relatively low concentrations (0.02 to 0.05%). 3. Relatively high concentration or prolonged treatment time was required to achieve sufficient bactericidal effect of CHG against some isolates of S. aureus, S. marcescens and B. cepacia. These results suggest that CHG is useful antiseptic agent or disinfectant for recent clinical isolates of various bacterial pathogens. In addition, the selection of treatment concentration and treatment time for each organism and purpose was important to obtain sufficient bactericidal effect. PMID- 15655907 TI - [The electronic personal health record: the spirit of the law of March 4, 2002]. PMID- 15655908 TI - [The fight against smoking. Efficacy of measures against societal factors]. PMID- 15655909 TI - [Smoking cessation--we can be efficient!]. PMID- 15655910 TI - [Smoking behavior in France: recent developments]. AB - The French "cancer fight plan" was launched 2 years ago and has given a new impulse to smoking prevention. Among the different measures proposed, increasing the tobacco price dramatically was the more efficient to reduce the smoking prevalence, and epidemiological effects already exist. In just 2 years, the smoking prevalence has fallen from 35.2 % to 30.4 % in the French adult population and that difference is more importance among youth. The prevalence has dropped from 44.5 % to 36.4 % in this category between 1999 and 2003 after several years of increasing. Health professionals are also concerning by this quick decrease as shown by several recent study. All of these developments are welcome but are very fragile and all of the aspects of prevention are still necessary. PMID- 15655911 TI - [Tobacco-related diseases]. AB - The tobacco smoke intake by the smoker or by the non-smoker is a toxic product and a risk factor for diseases as acne in teenagers, respiratory and non respiratory infection, or histiocytosis X. The four main diseases are: chronic obstructive pulmonary disease, which is more 80% related to tobacco; cancer (lung carcinoma, pharyngeal, laryngeal and oesophageal carcinoma, kidney, bladder carcinoma); cardiovascular diseases (myocardial infarction, cerebral stoke and sudden cardiac death); adverse consequences of smoking related to pregnancy. PMID- 15655913 TI - [Reduction of nicotinic risk--is it possible?]. AB - To decrease the health consequences of tabagism pandemic, quitting is still the only goal. The size of the problem leads to question if harm reduction is possible. Today, this concept, which was successful with illegal drugs, cannot be applied to tobacco because of a lack of proof. There is no safe cigarette and all brands have the same toxicity. Arguments in favor of smokeless tobacco are still insufficient for a public health decision. A voluntary decrease of tobacco consumption does not decrease morbidity and mortality of heavy smokers. A long term use of nicotine replacement therapy is acceptable to reduce the daily smoking under medical supervision in smokers with chronic tobacco related disease who have previously failed in attempts to quit smoking. PMID- 15655912 TI - [Current therapeutic strategies in smoking cessation]. AB - Smoking is a behaviour maintained and enhanced by a dependence mainly induced by nicotine. Despite awareness and knowledge of the associated health risks many smokers find it considerably difficult to quit. The untoward effects of nicotine withdrawal such as apparition of depressive mood, or weight gain, etc. justify the numerous unsuccessful attempts to quit smoking. Treatments with demonstrated efficacy are available and international evidence-based recommendations for cessation interventions have been established. These are: brief advice, assessing the smoking status of each patient and encouraging cessation; nicotine replacement therapies (NRT) [transdermal patch, gum, sublingual tablet or inhalator to be used at sufficiently individualised doses combining, if necessary, two or more NRT products]; bupropion, a more recent treatment: psychotropic drug, a noradrenaline and dopamine re-uptake inhibitor more recently approved for marketing; behavioural and cognitive therapies on their own or combined with pharmacotherapy. Measuring nicotine dependence using the Fagerstrom Test for Nicotine Dependence may help to define the therapeutic strategy. It is obvious that therapies can only work for smokers who are motivated to stop smoking. Before reaching the decision to quit, the smoker goes through a process in the course of which the role of health professionals' advice is paramount. PMID- 15655914 TI - [Smoking in teenagers: a priority issue]. AB - In France, the age that young people light up the first cigarette is particularly early, and thus in terms of public health tobacco use among the youth represents a high stake in the epidemiological projections of the mortality rate of illnesses linked to tobacco for the next twenty years. Measures to "deregulate" tobacco in society must intensify in association with programs that assist the breaking of tobacco dependence among those who have already begun to smoke. In adolescents, the best known treatment is to never start smoking, and to achieve this, what is needed are information campaigns on health education, well-adapted and solidly anchored in the public school curriculum, as well as the accompanying application of existing "deregulation" measures. PMID- 15655915 TI - [The French legislation against smoking]. AB - Smoking is one most common threat of public health around the world. One of the best way to fight it is legislation. The French legislation is probably one of the more sharp and precise for the non-smokers or youths protection. Historically, the law "Veil" voted for in 1976 has defined the French basis of this sort of prevention. The law "Evin" voted for in 1991 has reinforced the non smokers protection by the ban of smoking in public places, has decided the ban of advertising and has allowed very significant tobacco prices increases. Following European directives, new health warning appeared on cigarettes boxes in France in 2003. They are more strict, visible than they were before. It's because the law is implemented and because it's in continuous evolution that the French smoking prevalence will have the right evolution. PMID- 15655916 TI - [Fight against smoking: information and misinformation]. AB - For decades tobacco manufacturers have attempted to trivialise the health risks associated with tobacco, reinforce and enhance its social acceptability using powerful communication and reprehensible strategies. Prevention campaigns have contributed to raise public awareness on the health risks associated with smoking. They have also allowed for an evolution of the social representation of tobacco use. From 2001, to stop tobacco consumption taking off again, media campaigns have become more offensive both in content and form. Today the communication strategy places as priority objectives the reinforcement of the socially unacceptable character of tobacco use (denormalisation) via information on health risks and cessation support for smokers to quit. In 2002, a media campaign (TV, radio, press) denounced the toxicity of tobacco smoke provoking a true remobilisation of public opinion against tobacco. PMID- 15655917 TI - [Patient education: impact of the Internet]. PMID- 15655918 TI - [Neuro-sensory deficit in the elderly. Part II. Eye diseases]. PMID- 15655919 TI - [Vomiting in infants, children and adults, and its treatment. Part II. Vomiting in adults]. PMID- 15655920 TI - [Cancer: epidemiology, carcinogenesis, tumor evolution, classification]. PMID- 15655921 TI - [Medical responsibility: penal, civil, administrative and disciplinary]. PMID- 15655922 TI - [Management of a patient with cancer during all stages of the disease. Symptomatic treatment. Modalities of surveillance. Psychological, ethical and social problems]. PMID- 15655923 TI - [Apparent life threatening events and sudden infant death]. PMID- 15655924 TI - [Organisation and structure of the medical profession in ancient Egypt]. PMID- 15655925 TI - [Comparative surface examinations of morse taper junctions of the MRP-Titan stem shot peened with glass beads]. AB - INTRODUCTION: Shot peening is widely used for surface treatment of hip implants. Shot peening with steel balls followed by a cleaning process with glass beads is used for introduction of negative stress in the production of morse taper junctions of the MRP-Titan stem. An increasing number of publications in maxillofacial surgery and orthopaedic surgery show that there is a significant contamination of Alumina or glass blasted surfaces. Latest research suggested an association between contaminant particles with early loosening of endoprostheses (third body wear). The aim of this study is to evaluate the amount and the effects of surface contamination with glass particles on morse taper junctions of implants and explants of the MRP-Titan stem. MATERIAL AND METHOD: The surface of morse taper junctions of the MRP-Titan stem (5 original-package implants and explants each) are analysed for glass particle contamination. A field emission scanning electron microscopy (LEO 1525) is used for the detection of the glass particles on the implant surface with a backscattered electron detector. The relative surface area covered by particles was calculated by means of an image analyzing software (analySIS, Soft Imaging System GmbH). RESULTS: The surface of the implants showed a considerable contamination with glass particles with a mean of 6.67 +/- 0.82% compared to 2.06 +/- 0.74% on the surface of the explants. The difference was statistically significant (p<0.0001). DISCUSSION: The results of this study show that there is a relative high percentage of contamination with glass particles on shot peened morse taper junctions of the MRP-Titan stem. This contamination is significantly lower on the surface of the explants. With respect to third body wear and osteolysis in total hip arthroplasty further studies are necessary to minimize contamination while maintaining adequate surface quality. PMID- 15655926 TI - [Effects of clinically relevant alumina ceramic, zirconia ceramic and titanium particles of different sizes and concentrations on TNF-alpha release in a human macrophage cell line]. AB - INTRODUCTION: Aseptic loosening is considered to be the main problem of modern endoprothesis. Tumor necrosis factor alpha (TNFalpha) seems to be the initiator protein of particle disease. The aim of our study was to investigate the TNFalpha response of macrophage like cells (MLC) after stimulation with periprosthetic particles, typically found during revision surgery. For this purpose alumina ceramic (Al2O3), zirconia ceramic (ZrO2) and titanium (Ti) particles of different sizes and concentrations were used. Important was to study the effects of different sizes due to TNFalpha secretion and the comparison of the biological effects of alumina ceramic and titanium. METHOD: To obtain an TNFalpha profile we used an established macrophage model (Rader et al.) with THP-1 cells (human monocytic cell line). Therefore 106 MLC were incubated with different particle concentrations and sizes for 6 h. The supernatant was then investigated for TNF using ELISA assay. RESULTS: Ti-particles provoked in both sizes (0.2 microm and 2.5 microm) the greatest TNFalpha response, 8 times and 17 times as high in comparison with control. But substantially more 0.2 microm sized Ti-particles were necessary to get the above mentioned results. Al2O3-particles were not as effective as Ti, but they released fourfold more TNFalpha compared to control. There was no difference in TNFalpha-secretion comparing Al2O3-particles of different sizes (0.6 microm and 2 microm), but a 1000 times greater concentration of the 0.6 microm sized particles were needed. Using Al2O3- and Ti-particles of the same size and concentration, Ti provoked a significant higher TNFalpha response. ZrO2 showed no effects on TNFalpha release. CONCLUSION: Because of our results we recommend ceramic articulating surfaces, which are superior to metal on metal matings ion term of biological reactions. Additionally bigger wear particles should be avoided. Revisionoperation should be done early to avoid huge amount of wear particles and to minimize local osteolysis. PMID- 15655927 TI - [Are angle-fixed implants with elastic properties advantageous for the internal fixation of proximal humerus fractures?]. AB - There is a recent interest for the use of angle-fixed plates in the management of proximal humerus fractures. Rigid implants might be associated with an increased risk of cutting-out. In order to analyse the potential beneficial effects of the implant elasticity on fracture fixation, the biomechanical properties of a rigid and an elastic angle-fixed plating system were assessed in an experimental study. An unstable fracture of the surgical neck was created in 8 pairs of human humeri. Specimens were subjected to axial loading and torque. Stiffness, subsidence and load to failure were assessed. The implant with elastic properties was characterized by a lower torsional stiffness and a higher subsidence during axial loading and torque. This implant failed at lower loads than the rigid implant did. Elastic implant properties of angle-fixed plates have shown not to be advantageous for the management of fractures of the proximal humerus. PMID- 15655928 TI - The diagnostic value of digital subtraction arthrography and radionuclide arthrography in revision total hip arthroplasty. AB - An analysis of plain radiographs, digital subtraction arthrography, and radionuclide arthrography was performed in 25 revision hip arthroplasties to evaluate the efficacy and usefulness of these methods in the diagnosis of loosening. The findings by each method were compared with intraoperative assessment of the status of components and expressed in terms of sensitivity, specificity, and predictive accuracy. Overall accuracy for the acetabular component by plain radiographs was 80%; by digital subtraction arthrography, 88%; by radionuclide arthrography, 68%. Overall accuracy for the femoral component by plain radiographs was 92%; by digital subtraction arthrography, 84%; radionuclide arthrography, 76%. We consider subtraction arthrography and radionuclide arthrography to be adjuvant diagnostic tools which may be indicated in individual cases of suspected implant loosening of total hip arthroplasty. The routine use of these two methods is not warranted when compared to plain radiographs. PMID- 15655929 TI - [Application of a stand-alone interbody fusion cage based on a novel porous TiO2/glass composite. I. Implantation in the sheep cervical spine and radiological evaluation]. AB - Animals are becoming more and more common as in vitro and in vivo models for the human spine. Especially the sheep cervical spine is stated to be of good comparability and usefulness in the evaluation of in vivo radiological, biomechanical and histological behaviour of new bone replacement materials, implants and cages for cervical spine interbody fusion. In preceding biomechanical in vitro examination human cervical spine specimens were tested after fusion with either a cubical stand-alone interbody fusion cage manufactured from a new porous TiO/glass composite (Ecopore) or polymethyl-methacrylate (PMMA) after discectomy. First experience with the use of the new material and its influence on the primary stability after in vitro application were gained. After fusion of 10 sheep cervical spines in the levels C2/3 and C4/5 in each case with PMMA and with an Ecopore-cage, radiologic as well as computertomographic examinations were performed postoperatively and every 4 weeks during the following 2 and 4 months, respectively. Apart from establishing our animal model, we analysed the radiological changes and the degree of bony fusion of the operated segments during the course. In addition we performed measurements of the corresponding disc space heights (DSH) and intervertebral angles (IVA) for comparison among each other, during the course and with the initial values. Immediately after placement of both implants in the disc spaces the mean DSH and IVA increased (34.8% and 53.9%, respectively). During the following months DSH decreased to a greater extent in the Ecopore-segments than in the PMMA-segments, even to a value below the initial value (p>0.05). Similarly, the IVA decreased in both groups in the postoperative time lapse, but more distinct in the Ecopore segments (p<0.05). These changes in terms of a subsidence of the implants, were confirmed morphologically in the radiological examination in the course. The radiologically evaluated fusion, i.e. bony bridging of the operated segments, was more pronounced after implantation of an Ecopore-cage (83%), than after PMMA interposition (50%), but did not gain statistical significance. In this first in vivo examination of our new porous ceramic bone replacement material we showed its application in the spondylodesis model of the sheep cervical spine. Distinct radiological changes regarding evident subsidence and detectable fusion of the segments, operated on with the new biomaterial, were seen. We demonstrated the radiological changes of the fused segments during several months and analysed them morphologically, before the biomechanical evaluation will be presented in a subsequent publication. PMID- 15655930 TI - [In vitro long-term culture of human bone under physiological load conditions]. AB - There is evidence that mechanical loading is an important, if not the most important factor influencing bone mass and architecture. Investigations under in vivo conditions and cell culture methods, performed during the last years, helped to elucidate these mechanisms. However, the mechanisms by which load bearing acts on bone tissue are until now not completely understood. It is well accepted that weight-bearing exercise increases bone mass and on the other hand lower physical activity engenders bone loss. But neither a physiological threshold for bone loss or bone growth nor the character of the mechanical stimulus concerning amount, frequency and duration of the applied load are known. Even more speculative is the idea how this signal is transformed into the biological response of growing bone. Three-dimensional bone-culture-systems with simultaneous applied mechanical load enables to improve the knowledge of regulation of bone metabolism. We show the results of a long-term in vitro experiment with human cancellous bone under physiological load conditions. PMID- 15655931 TI - Biochemical pharmacology of pemetrexed. AB - Pemetrexed (Alimta) is a novel antimetabolite that inhibits the folate-dependent enzymes thymidylate synthase, dihydrofolate reductase, and glycinamide ribonucleotide formyltransferase. Pemetrexed has demonstrated activity in clinical trials in a variety of tumor types, including lung, breast, colon, mesothelioma, pancreatic, gastric, bladder, head and neck, and cervix. Pemetrexed is rapidly metabolized into active polyglutamate forms that are potent inhibitors of several tetrahydrofolate cofactor-requiring enzymes critical to the synthesis of purines and thymidine. Functionally, pemetrexed acts as a prodrug for its polyglutamate forms. Two different transporters are known to take extracellular folates, and some antifolates, into the cell. These are the reduced folate carrier and the folate receptor. One of the many attributes that make pemetrexed unique is that methodology has been developed to eliminate and control the many of its associated clinical toxicities. Multivariate analyses demonstrated that pretreatment total plasma homocysteine levels significantly predicted severe thrombocytopenia and neutropenia, with or without associated grade 3/4 diarrhea, mucositis, or infection. Routine vitamin B12 and folic acid supplementation have resulted in decreased frequency/severity of toxicities associated with pemetrexed without affecting efficacy, making this novel antifolate a safe and efficacious anticancer agent. PMID- 15655932 TI - Overview of phase I/II pemetrexed studies. AB - Pemetrexed (Alimta) is an antifolate that is effective in the inhibition of multiple enzyme targets including thymidylate synthase, dihydrofolate reductase, and glycinamide ribonucleotide formyl transferase. The compound has been evaluated in several phase I trials, both as single agent and in combination with other cytotoxic agents. The initial schedule selected for further investigation in phase II trials was pemetrexed 600 mg/m2 as a 10-minute infusion on day 1 every 21 days. During the subsequent phase II development the dose of pemetrexed was adjusted to 500 mg/m2 due to bone marrow and gastrointestinal toxicities. The adjusted dose of pemetrexed was well tolerated throughout the late-phase drug development program. Preclinical evidence suggests that pemetrexed has additive or synergistic activity when combined with many other clinically important anticancer agents, including gemcitabine (Gemzar), fluorouracil, carboplatin (Paraplatin), oxaliplatin (Eloxatin), paclitaxel, and vinorelbine (Navelbine). Dose-limiting toxicities in these studies were primarily hematologic, and there was no evidence of cumulative hematologic toxicity. During the drug development program it was discovered that supplementation with folic acid and vitamin B12 profoundly increased the tolerability of pemetrexed. The studies discussed in this review demonstrate that pemetrexed is well tolerated as a single agent and will be an important contribution to combination chemotherapy regimens. PMID- 15655934 TI - Pemetrexed in front-line chemotherapy for advanced non-small-cell lung cancer. AB - Pemetrexed (Alimta) possesses broad antitumor activity. It has been evaluated in non-small-cell lung cancer (NSCLC) as front-line chemotherapy in a comprehensive phase II evaluation. While various antifolates have been previously evaluated in clinical trials, drug development was stopped or delayed in light of their lack of efficacy or occurrence of life-threatening toxicities. While similar trends were observed with pemetrexed early in development, investigators instituted folic acid and vitamin B12 supplementation to minimize these toxicities without hampering drug efficacy. This article briefly summarizes the current evidence that supports the role of pemetrexed-based combinations in clinical trials for chemonaive patients with advanced NSCLC. PMID- 15655933 TI - Pemetrexed in malignant pleural mesothelioma. AB - Malignant pleural mesothelioma (MPM) is a disease with a poor prognosis, related in part to the aggressiveness of this disease, and in part due to the lack of drugs that have demonstrated tumor activity. Historically, antifolates such as methotrexate have been the most active drugs in the treatment of mesothelioma. Newer antifolates have recently demonstrated higher efficacy than older regimens in the treatment of this rare disease. One of these agents, pemetrexed (Alimta), has been evaluated both as a single agent and as part of a combination regimen. Pemetrexed has been studied in three trials in patients with MPM, and two phase I trials included patients with MPM. In a phase II trial, pemetrexed was studied as a single agent in patients with MPM. Seven of 64 patients achieved partial responses, with a median overall survival of 10.7 months. A large, randomized, phase III trial was conducted to compare pemetrexed/cisplatin with cisplatin. The response rate was 41.3% compared with 16.7%, median survival was 12.1 months compared with 9.3 months, and 1-year survival was 50.3% vs 38% in the pemetrexed/cisplatin and cisplatin arms, respectively. The combination of pemetrexed/cisplatin also demonstrated superiority in quality of life and pulmonary functioning analysis when compared with cisplatin. PMID- 15655935 TI - Pemetrexed in second-line treatment of non-small-cell lung cancer. AB - According to the updated 2004 guidelines of the American Society of Clinical Oncology (ASCO) on the treatment of advanced non-small-cell lung cancer (NSCLC), docetaxel (Taxotere) can be considered the standard second-line chemotherapy in patients relapsing after frontline therapy. This was based on two phase III trials (TAX 317 and TAX 320) that demonstrated the superiority of docetaxel at 75 mg/m2 in the parameters of survival, quality of life, and disease/symptom control when compared to best supportive care or alternative single-agent chemotherapy. The response rate was approximately 60%, with a median survival time of 7 months and a 1-year survival rate of 30%. Despite the activity demonstrated, this schedule showed an important toxicity profile, with grade 3/4 neutropenia and febrile neutropenia occurring in 70% and 11% of patients, respectively. However, the results obtained by these studies stimulated research interest in new drugs for this disease setting. Pemetrexed (Alimta), a new multitargeted antifolate, has achieved promising results in NSCLC treatment, as a single agent or in combination with other drugs. In the second-line setting, a large phase II study demonstrated good activity of pemetrexed, with an acceptable toxicity profile. This led to a phase III registration trial that compared pemetrexed at 500 mg/m2 to the standard docetaxel dose of 75 mg/m2. While results from this trial demonstrated a similar efficacy of the two regimens in response rate and survival, pemetrexed achieved a better safety profile. These results support the use of pemetrexed as a new option in the second-line treatment of NSCLC. PMID- 15655937 TI - Pemetrexed in transitional cell carcinoma of the urothelium. AB - Currently, the four-drug combination of methotrexate, vinblastine, doxorubicin (Adriamycin), and cisplatin (MVAC) or the two-drug combination of gemcitabine and cisplatin (GC) represents the standard of care for patients with locally advanced and metastatic transitional cell carcinoma of the urothelium. Recently, there has been a plethora of data from other chemotherapeutic regimens. Promising new agents, such as the multitargeted antifolate pemetrexed (Alimta), and new drug combinations have demonstrated increased efficacy and/or decreased toxicity compared with current regimens. Currently, data are available from three phase II studies utilizing pemetrexed or the combination of pemetrexed/gemcitabine (Gemzar) in patients with locally advanced and metastatic transitional cell carcinoma of the urothelium. Further investigation of combinations of pemetrexed and other active drugs in the treatment of patients with locally advanced and metastatic disease is warranted. PMID- 15655936 TI - Pemetrexed in pancreatic cancer. AB - Single-agent gemcitabine (Gemzar) is the standard of chemotherapy for advanced pancreatic cancer, with no phase III trials to date having shown significantly improved survival with gemcitabine-based combinations vs single-agent treatment. The multitargeted antifolate agent pemetrexed (Alimta) shows synergistic effects in vitro in combination with gemcitabine, and activity and good tolerability when used as single-agent treatment in advanced pancreatic cancer. In a phase II trial in patients with advanced pancreatic cancer, the combination of gemcitabine at 1,250 mg/m2 on days 1 and 8 plus pemetrexed at 500 mg/m2 on day 8 after gemcitabine every 21 days resulted in a median survival of 6.5 months and a 1 year survival rate of 29%. Neutropenia was the primary toxicity, with grade 4 toxicity in 51% of patients. The promising results of this trial prompted the initiation of a phase III trial comparing gemcitabine at 1,000 mg/m2 on days 1, 8, and 15 every 28 days vs the 21-day gemcitabine/pemetrexed regimen given with vitamin supplementation in patients with pancreatic cancer. The primary outcome measure was overall survival, with secondary measures including response rate, progression-free survival, and quality of life. While an increase in response and time to progression was reported for the gemcitabine/pemetrexed combination, there were no significant differences in survival between treatment arms. PMID- 15655938 TI - Pemetrexed in gastric cancer. AB - Gastric cancer is a major clinical challenge, with poor overall prognosis and limited life expectancy for patients with advanced disease. Even with surgery and other modalities, palliation is often difficult. Improvement of response rates has evolved with the development of standard regimens and those incorporating newer chemotherapy agents, such as oral fluoropyrimidines, the taxanes, camptothecins, novel platinums (eg, oxaliplatin [Eloxatin]), and antifolates (eg, pemetrexed [Alimta]). Ongoing trials with these regimens aim toward improving survival, as well as improving the safety profile. It is hoped that in conjunction with molecular research in the pathogenesis of gastric cancer and development of targeted therapies in this disease, these trial data might lead to the evolution of treatment strategies that could prove effective. PMID- 15655939 TI - Pemetrexed in advanced colorectal cancer. AB - Pemetrexed (Alimta) shows single-agent activity in advanced colorectal cancer. In two phase II studies in which patients received pemetrexed at 600 mg/m2 or 500 mg/m2 as first-line treatment for metastatic disease, objective response rates were 15.4% and 17.2%. These trials were conducted prior to supplementation with folic acid and vitamin B12, which markedly decreased the frequency of hematologic toxicities of pemetrexed; routine supplementation is now included in all clinical trials of the agent. The marked improvement in toxicity and tolerance with vitamin supplementation suggests the need to reexamine optimal dosing in pemetrexed combination schedules. In a National Surgical Adjuvant Breast and Bowel Project phase II trial in 54 patients with previously untreated advanced colorectal cancer, pemetrexed at 500 mg/m2 plus oxaliplatin (Eloxatin) at 120 mg/m2 every 21 days with folic acid/vitamin B12 supplementation resulted in an objective response of 23%. Three additionalpatients (5.6%) had unconfirmed partial response (partial response at one visit), and 27 patients (50%) had stable disease. Median progression-free survival was 5.3 months and median duration of response was 5.7 months; median overall survival was approximately 11.05 months. Grade 3/4 neutropenia was observed in only 17% of patients and treatment was well tolerated. A phase I/II study is under way to identify and assess the optimal combination of pemetrexed/irinotecan in second-line treatment of advanced colorectal cancer. Planned studies include a phase I study examining the combination of pemetrexed and oxaliplatin given every 2 weeks as first-line treatment, and a phase I/II trial to identify the optimal pemetrexed/ oxaliplatin dose in a 21-day schedule and to compare pemetrexed/ oxaliplatin with FOLFOX4 in first-line treatment of metastatic disease. All of these trials include vitamin supplementation. A phase III trial comparing the every-3-week pemetrexed/oxaliplatin regimen with FOLFOX4 as first-line treatment will be initiated if the outcome of the phase II trial is encouraging. PMID- 15655940 TI - Phase II studies of pemetrexed in metastatic breast and gynecologic cancers. AB - Pemetrexed (Alimta) is active in a variety of solid tumors, including breast and gynecologic cancers. Phase II trials of pemetrexed at a dose of 600 mg/m2 without vitamin B12 and folic acid supplementation in largely pretreated metastatic breast cancer patients demonstrated objective response rates of 21% and 28%, with generally manageable neutropenia constituting the primary toxicity. In phase II trials using 500 mg/m2 with or without vitamin supplementation in anthracycline- and taxane-pretreated patients, response rates were lower (approximately 9%) and treatment was generally well tolerated irrespective of vitamin supplementation status. A phase II trial is currently comparing pemetrexed doses of 600 and 900 mg/m2 with vitamin B12 supplementation in patients with previously untreated advanced breast cancer. In a phase II trial in patients with advanced cervical cancer, pemetrexed at 600 mg/m2 without vitamin supplementation and 500 mg/m2 with supplementation produced similar response rates, with the frequency of neutropenia being somewhat lower among patients receiving the lower dose and vitamin supplementation. Preliminary results in an ongoing phase II trial indicate activity of the regimen of gemcitabine (Gemzar) at 1,000 mg/m2 plus pemetrexed at 500 mg/m2 with vitamin supplementation in patients with ovarian cancer. Ongoing and future studies will establish optimal dosing regimens of pemetrexed and potential benefits of vitamin supplementation in the settings of metastatic breast cancer and gynecologic malignancies. PMID- 15655941 TI - Translational research with pemetrexed in breast cancer. AB - Pemetrexed (Alimta) is a novel folate antimetabolite that primarily inhibits the enzymes thymidylate synthase (TS), dihydrofolate reductase (DHFR), and glycinamide ribonucleotide formyl transferase (GARFT), all of which are involved in pyrimidine and purine synthesis. In a phase II trial of patients with T3/4, N0 2 breast cancer, expression of thymidylate synthase (TS), dihydrofolate reductase (DHFR), glycinamide ribonucleotide formyltransferase (GARFT), p53, and c-erb-B2 (at the mRNA or protein level) was examined in tumor biopsy specimens before and 24 hours after the first dose of pemetrexed and after three cycles of single agent treatment to establish correlations of biomarker levels and changes with clinical outcome and toxicity. Although final data are not available, initial indications are that clinical response may correlate with decreased or low TS expression. The results obtained from clinical data are supported by laboratory results in three cell lines (MDA-231, MCF-7, and ZR-75). These results suggest that in vitro transcript profiling to identify which genes are important predictors of successful cytotoxic chemotherapy, followed by a focused clinical trial to confirm the in vitro results, may be the best approach for translational research. PMID- 15655943 TI - [Biomechanically based planning for joint saving operation of the hip joint]. PMID- 15655942 TI - The promise of pharmacogenomics: gemcitabine and pemetrexed. AB - Although no overall differences in survival have been observed between the many chemotherapy combinations in non-small-cell lung cancer, the clinical application of mRNA expression levels of amplified genes may disclose many genetic influences on cytotoxic drug sensitivity and enable clinicians to tailor chemotherapy according to each individual's gene profile. Specifically, the assessment of ribonucleotide reductase subunit M1 and thymidylate synthase mRNA expression levels might select patients who benefit from gemcitabine (Gemzar) or pemetrexed (Alimta) combinations. Until recently, clinical prognostic factors such as performance status, weight loss, and lactate dehydrogenase were the only parameters used to predict chemotherapy response and survival. However, accumulated data indicate that overexpression of genes involved in cancer glycolysis pathways plays an important role, and might be an independent mechanism of chemoresistance. The dysregulation of glycolytic genes is affected by growth signals involving the PI3K/Akt pathway and downstream genes such as hypoxia-inducible factor-1-alpha. One can thus envision that substantial improvements in therapeutic outcome could benefit from the integration of tailored ribonucleotide reductase-dependent chemotherapy, ribonucleotide reductase antisense therapy, and targeted therapy. PMID- 15655944 TI - [Classification of coxarthrosis as the basis of operative joint salvage therapy. Biomechanics of cranial-lateral hip arthrosis; aim and technique of extension valgisation osteotomy; aim and technique of extension-varus osteotomy, results]. PMID- 15655945 TI - [Long-term results with Chiari pelvic osteotomies in adults]. PMID- 15655946 TI - [Results of the shelf operation after more than 10 years]. PMID- 15655947 TI - [Ball and socket endoprosthesis of the hip joint--resurfacing surgery as a hip joint saving procedure]. PMID- 15655948 TI - [Realignment osteotomies for degenerative arthritis of the knee joint]. PMID- 15655949 TI - [Progression of idiopathic coxa antetorta. a) Spontaneous progression of idiopathic coxa antetorta (controlled clinical study of 148 patients at the end of adolescence)]. PMID- 15655950 TI - [Progression of idiopathic coxa antetorta. B) What happens to the surgically corrected coxa antetorta?]. PMID- 15655951 TI - [Conservative management of congenital hip dislocation. a) 10 years later- congenital hip dislocation]. PMID- 15655952 TI - [Conservative management of congenital hip dislocation. b) Results after 10 or more years of conservative therapy of congenital hip dislocation]. PMID- 15655953 TI - [Conservative management of congenital hip dislocation. c) functioning after early therapy of hip dysplasia and hip dislocation]. PMID- 15655954 TI - [Results in the pelvic region of luxatio coxae congenita. a) 10-year results after Salter's pelvic osteotomy]. PMID- 15655955 TI - [Pelvic surgery in cases of luxatio coxae congenita . b) 10-year results of Salter's pelvic osteotomy partially combined with open reduction and/or femoral osteotomy]. PMID- 15655956 TI - [Pelvic surgery in cases of luxatio coxae congenita. c) Pelvic Chiari's osteotomy with or without bone wedges, 10 year-results]. PMID- 15655957 TI - [Pelvic surgery in luxatio coxae congenita. d) Long-term results of Chiari's pelvic osteotomy performed in children of pre-school age]. PMID- 15655958 TI - [Pelvic surgery in cases of luxation coxae congenita. e) 10-year results of Colonna's hip arthroplasty]. PMID- 15655959 TI - [Legg-Perthes disease. a) Legg-Perthes, introduction to 10-year results]. PMID- 15655960 TI - [Legg-Perthes disease. b) 10-year results of conservative management of Legg Perthes disease]. PMID- 15655961 TI - [5.) 10-year results after epiphyseolysis capitis femoris. a) 10-year results with nails and screws in epiphyseolysis capitis femoris]. PMID- 15655962 TI - [10-year results after epiphyseolysis. b) 10-year results after open reduction and subcapital osteotomy in epiphyseolysis capitis femoris]. PMID- 15655963 TI - [10-year results after epiphyseolysis. c.) 10-year results after epiphyseolysis capitis femoris]. PMID- 15655964 TI - [Coxarthrosis. a) 10-year results with the so-called Setzholz-total prosthesis]. PMID- 15655965 TI - [Coxarthrosis. b) 10-year results with total hip endoprosthesis]. PMID- 15655966 TI - [Coxarthrosis. c) 12-15 year results with intertrochanteric osteotomies in patients with coxarthrosis]. PMID- 15655967 TI - [Coxarthrosis. d) Long-term results with total endoprostheses of the hip]. PMID- 15655968 TI - [Coxarthrosis. e) 10-year results with total hip endoprostheses Balgrist 1962 1968]. PMID- 15655969 TI - [The hip in cerebral athetosis. 10-year results with derotation varisation osteotomies in cerebral palsy]. PMID- 15655970 TI - Destructive periodontal disease in relation to diabetes mellitus, cardiovascular diseases, osteoporosis and respiratory diseases. PMID- 15655971 TI - Periodontal risk management (PRM). PMID- 15655972 TI - Nutritional impact in oral health promotion. PMID- 15655973 TI - Home oral hygiene revisited. Options and evidence. AB - In regard to the limited literature on the subject, and the contradictions observed, we can not conclude that the types of manual brushes produce clinically important effects on the patients' gingival health, or that these effects can be detected consistently. However, the best results have been obtained with new brush designs, and future studies are necessary to clarify the existing contradictions. There is a clear need of long-term studies which comparatively evaluate the ability to reduce gingivitis and plaque with the newly designed brushes. On the other hand, there is evidence that supports the use of powered toothbrushes in the general population, especially those of the oscillating rotating and counter-rotational type, as they have shown their ability to reduce gingival bleeding or inflammation, and dental plaque with greater efficacy than manual brushes. There is a clear need of long-term trials on the efficacy of powered brushes in orthodontic patients. With the existing studies we can conclude that there is limited evidence that orthodontic patients using a powered toothbrush show a slight, but significant, reduction of bleeding, compared with users of manual brushes. No conclusion can be made concerning the type of brush to be used. The techniques of interproximal oral hygiene, fundamentally the use of dental floss and interproximal brushes, appear to add additional benefits, in terms of plaque reduction, when they are associated with conventional manual brushes. Further long-term studies are necessary to confirm their efficacy in the reduction of gingival bleeding or inflammation. The choice of the type of technique must be made in relation to the characteristics of the patient: dental floss could be indicated in individuals with closed interdental spaces, and inter proximal brushes in periodontal patients, or in those with open embrasures. PMID- 15655974 TI - Smoking prevention and cessation. PMID- 15655975 TI - Politically correct CPR. PMID- 15655977 TI - Is splitting up booty turning into a mutiny? PMID- 15655976 TI - Matching glove utility & dexterity. PMID- 15655978 TI - Work station positioning equipment keeps employee safety within reach. Lift, tilt or rotating tables are advantageous from the standpoint of safe ergonomic design. PMID- 15655979 TI - Incentive programs prove rewarding for employees who earn recognition. Injury and workers' compensation cost reductions are possible through cash or gift prizes. PMID- 15655980 TI - Employee health. PMID- 15655981 TI - CPR, first aid training may not meet expectations of federal regulations. PMID- 15655982 TI - EPA focuses on off-site consequences but overlap exists with process safety. PMID- 15655983 TI - State, Fed rules mismatched. PMID- 15655984 TI - Right-to-lifers not terrorists. PMID- 15655986 TI - Not far to fall. PMID- 15655985 TI - Right-to-lifers not terrorists. PMID- 15655987 TI - NIOSH enhances identity with move to Washington. Interview by R. Blake Smith. PMID- 15655988 TI - Corporate reply to traumatic events soothes survivors' troubled minds. PMID- 15655989 TI - Outdated air contaminants system disruptive to industry and labor. PMID- 15655990 TI - Prevention pays costs of cumulative trauma. PMID- 15655991 TI - Contractors' 'band-aid' remedies may defy bloodborne pathogen risks. PMID- 15655992 TI - Temporary worker services face regulatory burden for job safety. PMID- 15655993 TI - Targeting lead in the multimedia environment in the continental United States. AB - The U.S. Environmental Protection Agency's (EPA) lead attainment strategy for air is being expanded to address geographic areas with the potential for multimedia, multipathway exposures to lead. Geographic Information Systems (GIS) technology is used to coordinate information from various databases to identify areas of potential concern. The data retrieval and decision processes used in identifying priority sources from each medium and in evaluating identified areas of concern are described in this paper. Only EPA databases with reliable locational information were used to facilitate accurate mapping and allow correlation with other data sources. The sources of lead loadings to air, water, and soils were mapped using either latitude and longitude or zip code, or county centroids for data lacking longitudinal and latitudinal coordinates (such as the drinking water data). A multimedia cluster of lead sources was identified at the county level, since all the facility data in the five databases could be mapped to this level. An impact factor and weighting system was devised to combine the information on the number of facilities and their relative size in developing a ranking of the multimedia lead clusters of concern in each region. The counties with the highest number of points were considered clusters of highest concern for multimedia lead sources. Two separate lists of the clusters were developed according to a point system. One identified 10 multimedia lead clusters in each of the 10 EPA regions, and the other identified the 100 clusters of highest concern in the country as a whole. The project is designed to be a first step in targeting future efforts to identify potential environmental problems associated with lead. The analyses presented in this paper provide a first look at the areas in the country where there is a potential for multimedia exposure to lead. A more refined analysis at the zip code level was subsequently developed to provide a good understanding of the issues pertaining to potential exposure at the neighborhood level. The results of this analysis will ultimately help the EPA and the states to target implementation and enforcement in areas of high potential lead exposures. PMID- 15655994 TI - Urban emissions measured with aircraft. AB - Detailed knowledge of the quantity and composition of urban emissions is a prerequisite for successful application of atmospheric models to predict transport and distribution of primary and secondary air pollutants in the troposphere. We investigate the prospects and limitations of aircraft measurements in the determination of emission fluxes from urban areas. Our analysis focuses on data collected in September 1994 in and around Athens, Greece. Generally, emission fluxes from cities can be quantified with aircraft and with the minimum acceptable precision (uncertainty better than a factor of 2) only under very favorable meteorological conditions, namely in a homogeneous flow field in a well-mixed boundary layer. Better accuracy can be achieved only through ensemble averaging of repeated measurements. From our measurements in the Athens area, we deduced relative emission ratios of pollutant gases. With the support of ground-based measurements in a street canyon, the emission ratios NOx/CO, SO2/CO, and volatile organic compounds/CO (34 individual VOCs) could be determined with high precision. These results are very useful in analyzing differences between various existing emission inventories. Our data for VOCs reveal that the non-traffic emissions are of the same magnitude as the emissions originating from traffic. PMID- 15655995 TI - SCR performance on a hydrogen reformer furnace. AB - In late 1993, Air Products and Chemicals, Inc. began operating a new steam methane reformer at the Tosco Refining Co.'s Avon refinery in Martinez, CA, to provide hydrogen and steam to the refinery under a long-term supply agreement. The hydrogen plant--owned, operated, and maintained by Air Products--includes a selective catalytic reduction (SCR) unit on the reformer-furnace flue gas for environmental control. SCR is a commercially proven process capable of abating emissions of nitrogen oxides (NOx) to extremely low levels; however, documented experience in a refinery setting has been limited. This paper discusses performance of the SCR, primarily during its first two years of operation; it incorporates theory and prior research findings sufficient to understand the relationship between key system variables and SCR performance. Test results demonstrate that NOx, ammonia (NH3) slip, and carbon monoxide (CO) emissions are in compliance with permit limits. NOx removal efficiency is nearly linear with the inlet NH3:NOx molar ratio up to almost 90% NOx conversion, where ammonia slip begins to rise steeply. The stoichiometric reaction ratio of NH3 to NOx is close to the theoretical 1.0. Catalyst life is estimated at four years, in line with published figures for SCR catalysts in clean-gas service. PMID- 15655996 TI - Fluidized-bed-combustion ash for the solidification and stabilization of a metal hydroxide sludge. AB - Fluidized-bed-combustion (FBC) ash is a by-product from a developing technology for coal-fired power plants that will economically reduce air emissions to meet requirements of the Clean Air Act. FBC ash has physical and chemical properties similar to Portland cement, but only has moderate success as a pozzolan in concrete applications due to low compressive strengths. However, FBC ash has proven effective for use as a binder for the solidification and stabilization (S/S) of metal-bearing sludges. Physical and chemical characterization procedures were used to analyze FBC ash and a metal-bearing sludge obtained from a hazardous waste treatment facility to develop 12 different S/S mix designs. The mix designs consist of four binder designs to evaluate sludge-to-binder ratios of approximately 0, 0.5, and 1. Portland cement is used as a control binder to compare unconfined compressive strengths and Toxicity Characteristic Leaching Procedure (TCLP) analyses from different ratios of the FBC ash streams: fly ash, char, and spent bed material (SBM). Compressive strengths ranging from 84 lbs per square inch (psi) to 298 psi were obtained from various mix designs containing different sludge-to-ash ratios cured for 28 days. All the mix designs passed the TCLP. Recoveries from leaching for each metal were less than 5% for most mix designs. Results of unconfined compressive strengths, TCLP, and percent recovery calculations indicate that the mix design containing approximately a 1:1 ratio of fly ash to char-and-sludge is the best mix design for the S/S of the metal bearing sludge. PMID- 15655997 TI - Nitric oxide formation in an iron oxide pellet rotary kiln furnace. AB - A one-dimensional numerical model was developed to simulate the effects of heat and mass transfer on the formation of oxides of nitrogen (NOx) in a rotary kiln furnace for iron oxide pellet induration. The modeled kiln has a length-to diameter ratio of approximately seven. The principal mechanism of heat transfer is radiation from the flame, which was described by the net radiation method. The well known Zeldovich mechanism was used to predict thermal NOx generation. Temperature fluctuations in the vicinity of the flame were estimated with a clipped Gaussian probability density function. The thermal energy and mass balance model equations were solved numerically. The model is capable of predicting temperature profiles and NOx production rates in agreement with observed plant performance. The model was used to explore the effects of process changes on the total NOx formation in the kiln. It was concluded that the gas temperature as well as the partial pressure of oxygen in the process gases controls the rate of NOx formation. Lowering the temperature of the kiln gases by increasing the secondary air flow rates requires simultaneously decreasing the pellet production rate in order to maintain the pellet temperatures needed for blast furnace conditions. PMID- 15655998 TI - Quantifying NOx for industrial combustion processes. AB - The objectives of this paper are to (1) identify the problems with many of the units that are used to report and regulate NOx, (2) show how to properly correct NOx measurements for oxygen-enhanced combustion, and (3) recommend a preferred type of NOx unit. The current variety of NOx units make comparisons difficult and can cause considerable confusion. NOx may be measured on a wet or dry basis, but it is commonly reported on a dry basis. The reported NOx may differ from the actual measurements, which may be converted to a specific O2 basis level. Nearly all of the measured NOx from industrial combustion systems is in the form of NO, which is converted to NO2 in the atmosphere. However, when given on a mass basis, the measured NO is commonly reported as NO2 for regulatory purposes, but may be reported as NO, NO2, or simply NOx in technical papers. Some existing regulations may penalize combustion technologies with higher efficiencies and lower flue gas volumes, such as oxygen-enhanced combustion. Confusion may occur when applying some of the "conventional" NOx units to oxygen-enhanced processes. A better unit is the mass of NOx generated per unit of production, which also incorporates the overall process efficiency into the emissions. That unit does not penalize more efficient processes that may generate more NOx on a volume basis, but less NOx on a production basis. PMID- 15655999 TI - Preliminary on-road measurement of the effect of oxygenated fuel on CO emissions near Las Vegas, Nevada. AB - A preliminary measurement of an oxygenated fuel effect for in-use vehicles travelling at freeway speed was conducted near Las Vegas, NV, in January 1991 and May 1992. The experimental design was based on two principal factors: (1) the large volume of traffic that visits Las Vegas from Southern California on three day holiday weekends and (2) the fact that, at the time of the study, the Las Vegas area used oxygenated fuels in the winter and Southern California did not. Measurements were conducted at carefully selected sites 20 km southwest of Las Vegas near Sloan, NV, and were accomplished with the Fuel Efficiency Automoblie Test (FEAT) remote sensor developed at the University of Denver. The January 1991 measurements were made during the Las Vegas oxyfuel season, while the May 1992 control measurements were made outside the oxyfuel season. Over 24,500 individual CO concentrations were measured; registration data from over 5,500 of these vehicles were obtained from the license plate numbers. After corrections for differences in velocity and mean age, the Las Vegas outbound (oxyfuel) CO emissions on Monday morning of the January holiday weekend showed a difference of -18% +/- 11% compared to the inbound (non-oxyfuel) CO emissions on Friday evening preceding the holiday weekend. PMID- 15656000 TI - Influence of water content on degradation rates for ethanol in biofiltration. AB - Treatment of ethanol vapor in a peat biofilter with various initial water contents (70%, 59%, 49%, and 35%) was studied. For water contents ranging from 49% to 70%, elimination capacity was about 30 g/m3/h. For a water content of 35%, elimination capacity decreased to 4 g/m3/h. A low mean CO2 yield coefficient (0.35 g CO2 produced per g ethanol consumed) was found for all of the initial water contents. The value was only 20% of the yield coefficient (1.91 g/g) predicted by stoichiometry. When the packing material was dried from 70% to 59% water content during the biofiltration process, elimination capacity dropped from 27 g/m3/h to 4 g/m3/h. After 24 hours of drying, the biofiltration experiment was restarted and run for two more weeks. During this period, the biofilter did not recover. At 59% water content, the rate of water evaporation was estimated at 59.6 g/m3/h. A simplified mass balance permitted calculation of the biological water production rate, approximately 22.1 g/m3/h. PMID- 15656001 TI - An exploratory look at hydrocarbon data from the Photochemical Assessment Monitoring Stations network. AB - This paper describes some characteristics of speciated nonmethane organic compound (NMOC) data collected in 1994 at five Photochemical Assessment Monitoring Stations (PAMS) and archived in the U.S. Environmental Protection Agency's Aerometric Information Retrieval System (AIRS). Topics include data completeness, distribution of individual NMOCs in concentration categories relative to minimum detectable levels, percentage of total NMOC associated with the sum of the 55 PAMS target compounds, and use of scatterplots to diagnose chromatographic misidentification of compounds. This is an early examination of a database that is expanding rapidly, and the insights presented here may be useful to both the producers and future users of the data for establishing consistency and quality control. PMID- 15656002 TI - Real-time, ultrasensitive monitoring of air toxics by laser photoionization time of-flight mass spectrometry. AB - Gas phase concentrations of individual polycyclic aromatic hydrocarbons (PAHs) were measured in real time in combustion products from a co-flow diffusion flame using laser photoionization (LP) time-of-flight mass spectrometry (TOF/MS). In particular, a naphthalene detection sensitivity of 4 parts per billion (ppb) was demonstrated. The use of calibration mixtures with argon indicated the feasibility of naphthalene detection at about 45 parts per trillion (ppt) at a signal-to-noise (S/N) ratio of 20. This suggests the possibility of low-ppt level detection at a S/N of 1. The novelty of the system is the use of a heated sampling probe and a continuously purged, heated-pulse valve that was positioned close to the ionization zone, thereby allowing the generation of photoions in the high-density region of the sample jet, where concentrations of PAH are high. Because the system developed allows for the real time detection of select species, it represents a useful tool in continuous emissions monitoring (CEM) for environmental compliance as well as direct process control. PMID- 15656003 TI - [CME radiology 4]. PMID- 15656004 TI - [Prognostic value of low back pain history with reference to outcome of lumber intervertebral disk displacement operation--a prospective cohort study]. PMID- 15656005 TI - [Barrett's esophagus]. AB - The diagnosis of Barrett's esophagus is rendered based on proof of intestinal metaplasia in the tubular portion of the esophagus. Barrett's develops in a percentage of patients with gastroesophageal reflux disease; risk factors include a long history of the disease, age over 40 years and Caucasian skin. Specifics about a genetic predisposition have not become known to date. Each year, around one out of every 200 patients with Barrett's epithelium develop adenocarcinoma of the esophagus, the incidence of which has risen dramatically over the past two decades. Apart from the early stages, the prognosis for this type of Barrett's carcinoma is extremely unfavorable, even after esophagectomy. It therefore appears sensible to examine patients with a long history of reflux and/or frequently recurrent reflux symptoms and to develop screening strategies for timely detection of persons with Barrett's esophagus along with subsequent monitoring. This would involve regular endoscopic studies accompanied by biopsies aimed at excluding or demonstrating the intraepithelial neoplasms that count as direct precursors to cancer. Treatment of nonneoplastic Barrett's esophagus can be symptomatic. Although theoretically logical, the benefits of normalizing esophageal acid exposure have not been proven. When high-grade intraepithelial neoplasms or mucosal carcinomas have been confirmed, local endoscopical resection and/or ablation appear sufficient, since the risk of lymph node metastasis is extremely low. Previous studies on this subject have been very promising, but should be continued and/or verified. Definitive therapy of more advanced tumor stages is currently given according to multimodal concepts established in an interdisciplinary manner. PMID- 15656006 TI - [Diagnosis of gastro-esophageal reflux disease]. AB - The diagnosis of gastro-esophageal reflux disease (GERD) predominantly relies on the patient's history and endoscopically visible alterations of the esophageal mucosa. Heartburn as the dominating symptom is highly suggestive of GERD though not specific whereas heartburn as one complaint among others is of limited value. Endoscopy enables the definite diagnosis of reflux esophagitis; however, the majority of patients (approximately 60%) does not exhibit erosions (non-erosive form of GERD). In these patients a short course of empiric PPI therapy gives valuable information about the presence or absence of GERD. Only those patients non-responsive to adequate PPI doses should undergo pH-metry. Other diagnostic investigation (e.g. manometry) should be restricted to selected cases. PMID- 15656007 TI - [Reflux disease therapy]. AB - The frequency of gastroesophageal reflux disease is on a constant rise in western countries. Reflux therapy can be subdivided into the treatment of acute erosive reflux esophagitis and the subsequent recurrence prophylaxis, and into the treatment of non-erosive reflux disease as well as the atypical reflux symptoms. Based on conventional large-scale studies, a highly effective therapy is available which offers virtually complete acid blockade using proton pump inhibitors (PPI) of the prazole type. Used for acute treatment, they offer a cure for reflux esophagitis and freedom from symptoms in approx. 90% of the cases, while also showing high efficacy in recurrence prophylaxis. Conversely, the rate of side effects, particularly serious ones, is very low and the long-term safety of this therapy has been good. Since the risk of erosive reflux esophagitis recurring is > 80% without PPI therapy, the majority of the patients will require a long-term, potentially even life-long therapy with PPI. Other therapeutic options are being discussed as alternatives to such long-term prophylaxis. In selected cases, surgical therapy by laparoscopic fundoplication is one option. Alternatively, endoscopical anti-reflux procedures are increasingly coming into use; their merit in long-term prophylaxis however, cannot yet be evaluated. Whenever PPI therapy does not lead to complete freedom from symptoms, an alternative therapeutic approach should be given consideration even in patients with atypical reflux symptoms. The therapy of the Barrett's esophagus, a potentially long-term sequelae of reflux disease, will not be dealt with in this overview, as it is the subject of a separate presentation. PMID- 15656008 TI - [CME ECG 2--solution]. PMID- 15656009 TI - Bulimia symptoms and risk factors in university students. AB - We proposed a model for bulimic symptoms in a sample of 153 female university students attending the Faculty of Psychology, and determined the extent to which such symptoms could be explained by these variables. The following variables were measured during class time: the effects of aesthetic body modelling, the impact of teasing about weight and other aspects of physical appearance, body dissatisfaction, dieting severity, bulimic symptomatology, depression and self esteem. The data underwent Lisrel analysis. The final model was similar to that initially proposed. The most important variables in emergent bulimic symptoms were the perceived pressure of aesthetic body modelling, teasing about weight, body dissatisfaction, self-esteem, depression and dieting. All of these play a role in eating disorder symptomatology through various pathways. PMID- 15656010 TI - Abnormal eating attitudes and sexual abuse experiences in Turkish university women. AB - The aim of this study was to examine the direct and indirect relationship between abnormal eating attitudes and sexual abuse. The subject sample comprised 532 female Turkish undergraduate and nursing students in Istanbul. The Eating Attitudes Test (EAT), Sexual Abuse Questionnaire, the Setting Conditions for Anorexia Nervosa Scale, the Positive and Negative Perfectionism Scale and the Need for Control Scale (NCS) were the instruments of the study. The results showed that sexual abuse has a direct effect on EAT-Bulimia but not on EAT Dieting. It also has some indirect effects on both of these through its effects on family functioning. Perfectionism, on the other hand, was independently linked with both EAT-Dieting and EAT-Bulimia but was not associated with sexual abuse. PMID- 15656012 TI - Body image across three generations of Americans: inter-family correlations, gender differences, and generation differences. AB - Three-hundred-and-eighty-one participants (undergraduates, their parents and grandparents) completed body image (BI; current minus ideal figure ratings) and dieting attitude measures. We found the usual gender gap in BI for the undergraduates (females' BI worse than males), but not for the parents and grandparents. This was due to males' worsening BI with age; females' BI did not differ across generations. The gender gap in dieting attitudes (females more likely to diet) also narrowed with increasing age (again due to males' changing attitudes), but remained significant across generations. In all three generations, females underestimated the size of the figure males found most attractive, whereas males overestimated the analogous figure for females. Finally, we found significant inter-family correlations for BI and dieting for all groups except undergraduate females. We discuss these results within cultural and evolutionary theoretical frameworks. PMID- 15656011 TI - Population at risk for eating disorders in a Spanish region. AB - OBJECTIVE: To estimate the prevalence of adolescent population at risk for eating disorders (EDs), to examine gender differences and their association with non psychotic mental disorders. METHODS: Cross-sectional study using an anonymous, self-reported questionnaire in the classroom and measurement of weight and height in a representative sample of 4334 teenagers of both sexes. A definition of population at risk for EDs has been established, that differentiates those with dieting criteria and those with binge-purge eating behaviour criteria. This definition includes a combination of weight, behaviour, Diagnostic and Statistical Manual of Mental Disorders (DSM-IV) criteria, and Eating Disorder Inventory (EDI) results. The General Health Questionnaire-28 (GHQ-28) was used to assess non-psychotic mental disorders. RESULTS: The prevalence of population at risk for EDs was 2.2% (95% CI: 1.6-2.8) for men and 15.3% (95% CI: 13.8-16.9) for women. The prevalence rate of combining both ED risk and mental disorder was 0.8% (95% CI: 0.4-1.2) for men and 9.9% (95% CI: 8.6-11.2) for women. Non-psychotic mental disorders were more prevalent in the population at risk for EDs than in the rest of the population. CONCLUSIONS: A considerable proportion of adolescent females have eating problems and non-psychotic mental disorders. Their male counterparts also suffer from these disorders, however, a different pattern is followed. PMID- 15656013 TI - Quality of life in patients with binge eating disorder. AB - OBJECTIVE: To examine Health-Related Quality of Life (HRQL) in patients with binge eating disorder (BED). DESIGN: Cross-sectional comparison of HRQL in a BED sample to published normative and clinical samples. Further cross-sectional comparisons within the BED sample comparing obese and non-obese subjects, and depressed and non-depressed subjects. SUBJECTS: Ninety-four patients (73 women and 21 men), who met DSM-IV criteria and attended a medical school based program for BED, participated. Mean body mass index (BMI) was 35.2 (SD = 8.1), and mean age was 44.9 (SD = 8.3); 78.7% of participants were Caucasian. MEASUREMENTS: Subjects completed the Medical Outcomes Study Short Form-36 Health Survey (SF 36), a self-report measure of HRQL. RESULTS: BED patients reported worse functioning in all HRQL domains than US norms, and in some domains of HRQL than obese treatment seekers. Among BED patients, obese BED subjects had significantly worse Physical Component summary scores than non-obese BED subjects, whereas depressed BED subjects had significantly worse Mental Component summary scores than non-depressed BED subjects. CONCLUSION: Obesity status and depression appear to be related to HRQL among BED patients with obesity being related to physical HRQL and depression being related to mental HRQL. PMID- 15656015 TI - The relationship between body image and quality of life in treatment-seeking overweight women. AB - OBJECTIVE: To test the impact of body image on quality of life in overweight and obese people; to evaluate longitudinally the relationship between changes in body image, anthropometrical measurements and quality of life. DESIGN: A cross sectional study and a longitudinal study in a sub-sample after 6 months. SUBJECTS: 308 women in the cross-sectional study and 56 in the longitudinal study, aged 21-65, with body mass index (BMI) > 25 kg/m2 and selected from people who had been sought treatment for overweight in our Institute from 1998 to 2001. MEASUREMENTS: Body weight, body height, waist and hip circumferences. The ORWELL 97 questionnaire to evaluate the obesity related quality of life (ORQL) and the BUT questionnaire to assess the body image (BI) were used. RESULTS: The BUT score was the variable that explained the greatest amount of variance of ORWELL 97 both in cross-sectional and longitudinal studies. Despite no significant differences in athropometrical variables were found between subjects who dropped out and those who did not, psychometrical scores were significantly better in patients still in treatment in follow-up. CONCLUSION: BI is related to subjective ORQL independently of anthropometrical measurements both at baseline and after treatment. Better psychometrical scores in people still in treatment after 6 months suggest that BI could be considered one of the variables which influence compliance to the treatment program, this fact needs more investigations and could be of interest in obesity treatment outcome studies. PMID- 15656014 TI - Patterns of personality disorders in women with chronic eating disorders. AB - The aim of this study was to describe patterns of personality disorders (PDs) in women with chronic eating disorders (EDs). An index group of nineteen women who have had EDs for an average of 8.5 years was compared with a control group of same-aged women from the general population. At the time of the study the index group received treatment at a tertiary treatment center in Stockholm. The PDs were assessed using the DSM-IV part of the DSM-IV and ICD-10 Personality Questionnaire (DIP-Q). In the index group, eighteen of nineteen fulfilled the criteria for one or more PD. The number of PD diagnoses for each women ranged from zero (n = 1) to eight (n = 2) with a median of three. Among the controls, only one woman fulfilled the criteria for one or more PD. The most prevalent disorders in the index group were Borderline, Avoidant, and Obsessive-Compulsive. The index group had significantly higher DIP-Q dimensional scores than the controls in the Paranoid, Schizoid, Schizotypal, Borderline, Histrionic, Avoidant, and Dependent scales. Although the assessment of PD symptoms was limited to self-reports, the high prevalence of PD diagnoses and PD symptoms most probably reflects the severe psychiatric impairments in patients suffering from chronic ED. PMID- 15656016 TI - Assessment of binge eating disorder in morbidly obese patients evaluated for gastric bypass: SCID versus QEWP-R. AB - OBJECTIVE: Binge eating disorder (BED) has been hypothesized to be associated with poor outcome in gastric bypass surgery (GBP). However, past studies have yielded inconsistent results regarding BED and surgical outcome, which may be due to the variety of measures used to assess BED. The present study examines the utility of two commonly used BED diagnostic tools: the Structured Clinical Interview (SCID) and the Questionnaire of Eating and Weight Patterns-Revised (QEWP-R). METHOD: Subjects were 168 adult patients evaluated for GBP. BED was assessed using the SCID and the RESULTS: 27% of the sample received a diagnosis of BED using the QEWP-R, compared with 14% using the SCID. Agreement using Cohen's kappa was 0.37. Compared to sub-non-concordant diagnoses of BED, subjects with concordant diagnoses scored on a measure of self-esteem. DISCUSSION: Although both diagnostic tools use the Diagnostic and Statistical Manual of Mental Disorders-Fourth Edition (DSM-IV) criteria, each yielded different results. Further studies are needed to determine the most accurate method of assessing BED in this population. PMID- 15656017 TI - Self-disclosure in eating disorders. AB - OBJECTIVE: Secrecy and concealment are typical behaviours in individuals with eating problems. This study explored the relationship between eating-related problems and self-disclosure. It examined whether women with greater eating related problems were less willing to disclose. Different types of self disclosure were calculated, considering disclosure related to body appearance and to restrained eating. The role of risk factors which concur to the development and maintenance of eating symptomatology was also explored. METHOD: The Eating Symptoms Inventory was used to investigate the existence of an eventual eating symptomatology, self-disclosure was calculated through the Self-Disclosure Index, while a new scale was validated to assess a self-disclosure related to body image and eating attitudes. Other scales measured the influence of different risk factors, as body dissatisfaction, social pressure to be thin, and restrained eating. RESULTS: A significant inverse relationship was found between general self-disclosure and psychological aspects related to the practice of wrong weight control behaviours and risk factors as dieting, body dissatisfaction, and social pressure to be thin. The significant role of risk factors was confirmed in the development and maintenance of eating disturbances. Interesting results were found using the different self-disclosure indexes as mediators and moderators. Relevant differences were found between Dutch and Italians concerning to their eating attitudes and to the role of different risk factors. CONCLUSION: Some limits are the impossibility to generalize these findings and the use of a non clinical sample. Some new longitudinal studies should be done in this direction to deepen the relationship between self-disclosure and eating disorders. PMID- 15656018 TI - Treatment of adolescent patients with eating disorders: description of a psychodynamic approach in clinical practice. AB - Anorexia nervosa (AN) is now usually considered to be a multifactorial disorder, and there is a consensus among clinicians that its treatment should be aimed at restoring weight, altering anorexic attitudes, treating any medical complications, supporting and treating the family, enhancing autonomy, facilitating identity formation, and increasing self-esteem by means of psychotherapy. The practical aspects of such treatments not only vary from country to country, but sometimes also from one treatment team to another. International meetings dedicate considerable discussion to the subject but, as it seems to be relatively ignored in the published literature, we here describe the main elements of our own method. PMID- 15656019 TI - The Eating Disorder Examination as a retrospective interview. AB - The purpose of this study was to determine the validity of the Eating Disorder Examination 12.0D (EDE) when administered retrospectively. METHODS: Twenty-five female patients suffering from an eating disorder [(10 with anorexia nervosa (AN), 10 with bulimia nervosa (BN), 5 with eating disorders not otherwise specified (EDNOS)] were investigated using the EDE at the time of the first referral to our outpatient ward (T1). Afterwards (mean 1.4 +/- 0.6 years later) each patient was administered again the EDE by the same assessor (T2). At this time the interviewer asked the patients to answer the questions referring to the symptoms and behaviours at the time of the first interview. RESULTS: Test-retest correlation factors were 0.7 or greater for all subscales of the EDE (p < 0.0001) and 0.5 or greater for every single item of the EDE (p < 0.001), except for EDE 1.5 (snack after dinner) and EDE 9A.6 (maximum time free from objective bulimic episodes in the last two months). DISCUSSION: Our results provide evidence that the EDE 12.0D is a reliable interview even when administered retrospectively, suggesting the use of this instrument for the retrospective assessment of eating disorders. PMID- 15656020 TI - The predictive value of body mass index for the weight evolution in anorexia nervosa. AB - BACKGROUND: Low body weight is considered to be an important risk factor for poor outcome in anorexia nervosa (AN). OBJECTIVES: To study the relationship between body mass index (BMI) at the beginning of treatment and at follow-up. A BMI of 13 kg/m2 at admission was hypothesized as cut-off point for better versus worse evolution of weight after treatment. METHOD: The BMI of 232 female AN inpatients was calculated at admission and after 6 months and 1 year. RESULTS: An overall low correlation between BMI at admission and at 6 months (r = 0.27) and 1 year (r = 0.26) follow-up was found. A BMI value of 15 kg/m2 at admission was revealed as the best cut-off point for making predictions. CONCLUSIONS: Patients with an admission BMI below 15 kg/m2 have a significantly greater chance to develop a lower BMI value at follow-up than those with a higher BMI. PMID- 15656021 TI - Hypomagnesemia during refeeding in anorexia nervosa. AB - BACKGROUND: Magnesium deficiency can cause weakness, constipation, seizures and arrhythmias. We frequently observe hypomagnesemia during refeeding in AN. OBJECTIVE: To determine the incidence and time of onset of hypomagnesemia during refeeding in anorexia nervosa (AN). DESIGN: Observational cohort study. SETTING: University teaching hospital in Vancouver, Canada. PATIENTS: Patients with AN (DSM-IV criteria) admitted for refeeding. INTERVENTION: All patients were admitted for supervised refeeding by meal support, in conjunction with our standard medical and psychological treatment. MEASUREMENTS: Serum magnesium was measured daily for 5 days and then 3 times a week. RESULTS: Fifty patients were admitted for an average of 24 days. Sixty percent (30/50) had low serum magnesium during their admission. Hypomagnesemia was present on admission in 16% but as late as the third week of refeeding in others. CONCLUSION: Serum magnesium should be measured on admission and rechecked weekly for the first 3 weeks of refeeding as a minimum. PMID- 15656022 TI - A survey of the use of assessment instruments by eating disorder professionals in clinical practice. AB - Accurate assessment is important in all phases of the treatment process, but there is little information about the assessment procedures utilized by those treating eating disorders in clinical practice. This study surveyed the assessment practices of eating disorder specialists. A brief questionnaire was sent to 480 eating disorder specialists in clinical practice; 95 were returned (19.8% return rate). The questionnaire asked specific questions about what assessment methods and measures they used for initial assessment and diagnosis as well as for treatment outcome. A significant minority of respondents used no validated measures at any stage of the assessment process, and even among those who did use well-validated assessment instruments, only a few instruments were used with any regularity. These results raise some potentially troubling issues concerning the typical assessment practices of clinicians. More collaboration between researchers and clinicians is needed to improve this situation. PMID- 15656023 TI - Cardio-metabolic burden of native asian Indian--India the global capital. PMID- 15656024 TI - Health status of Indian population--current scenario. AB - OBJECTIVE: We aimed at establishing reference intervals for the various biochemical and hematological analytes in healthy population. We also tried to find the percentage of people with coronary artery disease (CAD) and the associated risk factors in 39,940 subjects who had attended the health check up program at our hospital from the years 1996 to 2001. METHODS: The medical record folders of all the subjects were screened manually. Reference values were established using SPSS-8.0 package and the percentiles calculated and with it the corresponding 90% confidence interval (CI). RESULTS: The prevalence of hypertension, diabetes mellitus, and coronary artery disease was found to be 22.5%, 14.2%, and 3.9% respectively. In addition only 41.1% of the population was found to be normolipemic. Most of the analytes showed reference intervals which were in agreement with our reporting values. There was no influence of diet on the reference intervals. Also, there were some analytes like lipids where it was felt that changing the reference values would assign the subjects at greater risk for CAD. CONCLUSION: Implementation of reference intervals in case of lipids poses a dilemma. Lifestyle and diet modifications would have to be implemented to reduce the burden of CAD in this population. PMID- 15656025 TI - Apolipoproteins: correlation with carotid intimamedia thickness and coronary artery disease. AB - BACKGROUND: Lower levels of plasma apolipoprotein AI (Apo A-I) and higher levels of ApoB, and the ratio of ApoB to ApoA-I are considered to be independent risk factors for coronary heart disease, and may assume importance in the definite subset of Indian patients with normal levels of traditional lipid risk factors and an early-onset of coronary artery disease (CAD). Carotid intima-media thickness is considered as a marker of atherosclerosis and in prediction of clinical coronary events and coronary artery disease. With increasing interest in the role of non-traditional lipid risk factors in CAD and few studies reported in Indian subjects, we undertook this study to correlate the apolipoprotein levels with CAD and their impact on arterial thickening utilizing the carotid intima media thickness as a surrogate marker. METHODS AND RESULTS: Traditional lipid profile, apolipoprotein A-I and B and carotid artery Intima-media thickness (IMT) with a B-mode scan were measured in 309 patients recruited for the study (age group 36-64 years), which included 193 males and 116 females. Mean of maximal IMT exceeding 0.8 mm at the far wall of common carotid artery, excluding plaques, was selected as the higher values for comparison. One hundred and twenty two subjects had evidence for CAD as diagnosed by documented hospitalization with myocardial infarction or acute coronary syndrome, coronary angiography when feasible or noninvasive cardiac evaluation. Prevalence of subjects with increased IMT was higher among subjects with an apolipoprotein B: apolipoprotein A-I ratio exceeding one compared to those with a ratio less than one (30.6% vs 16.5%, p = 0.005). Prevalence of CAD was significantly higher among subjects with apolipoprotein B: apolipoprotein A-I ratio exceeding one as compared to those with a ratio less than one (53.7% vs 30.3%, p = 0.0002). Subjects with apolipoprotein B: apolipoprotein A-I ratio exceeding one and carotid IMT more than 0.8 mm had a 2.7-fold prevalence for CAD as against those with a ratio less than one and IMT less than 0.8 mm. On multivariate logistic regression analysis, apolipoprotein B: apolipoprotein A-I ratio exceeding one was significantly associated with increased IMT (odds ratio 2.27) and CAD (odds ratio 2.50) even after inclusion of sex, smoking, body mass index, total cholesterol, LDL cholesterol, HDL-cholesterol, total cholesterol:HDL-cholesterol ratio and serum triglycerides into the model. CONCLUSIONS: We conclude that apolipoprotein B to A I ratio shows a strong association with carotid intimal medial thickening and coronary artery disease in this Western Indian population and may play an important role is assessment of coronary risk in addition to traditional coronary risk factors. PMID- 15656026 TI - Importance of reciprocal leads in acute myocardial infarction. AB - OBJECTIVES: To study the relevance of the ECG changes in the reciprocal leads in patients with acute anterior and inferior wall myocardial infarction, with regard to culprit artery localization and left ventricular (LV) function. METHODS: Three hundred patients of acute myocardial infarction (AMI) (180 anterior, 120 inferior) aged between 30-90 years (mean age - 60 yrs; M:F - 220:80) were studied with regard to the reciprocal lead changes which were correlated with the culprit coronary artery and LV function. 285/300 (95%) patients underwent echocardiography and 62/300 (20.67%) underwent coronary angiography (CAG). RESULTS: In patients with acute anterior wall MI (AWMI), Q wave in inferior leads was found in 38.8% (70/180) patients. Nineteen patients underwent (CAG) and all 70 patients underwent 2D echocardiography. CAG revealed 10/19 and 9/19 patients to have single vessel disease (SVD) and multivessel disease (MVD), respectively. In presence of SVD, 80% (8/10) patients were found to have a distal/mid LAD occlusion. The echocardiogrpahy of these 19 patients showed that 15/19 (78.94%) of these had LV ejection fraction (EF) > 40% as against only 4/19 (21.05%) patients with LVEF < 40%. The echocardiographic study of patients with and without reciprocal ST depression in inferior leads more than or equal to the ST elevation in anterior leads, showed higher incidence of LV dysfunction i.e. LVEF < 40% in patients with reciprocal changes (72.05%) as compared to patients without the reciprocal changes (27.94%). In patients with acute inferior wall MI (IWMI), anterior and lateral ST depression more than or equal to ST elevation in inferior leads was found in 80% (96/120) patients. 27/96 patients underwent coronary angiography and all 96 underwent echocardiography. Patients with ST depression in I, a VL, V4-V6 (apicolateral leads) > or = ST elevation in inferior leads were found to have more occurrence of multivessel disease (21/27) with LVEF < 40% (50/64 i.e. 78.12%, P < 0.001). 12.5% (4/32) patients with reciprocal changes in anterior i.e. VI-V3 leads and 15% (3/20) patients without significant reciprocal changes in anterior and lateral leads had LVEF < 40%. Also, patients with ST depression in I, aVL showed higher incidence of right coronary artery (RCA) lesion (23/27) (P < 0.001). CONCLUSIONS: Patients of acute AWMI with Q waves in inferior leads indicate a smaller infarct with higher incidence of mid/distal LAD occlusion and a relatively preserved LV function. AWMI patients without reciprocal changes in inferior leads have a better LVEF. Patients of acute IWMI with ST depression in apicolateral leads have more occurrence of multivessel disease with significant LV dysfunction. Reciprocal ST depression in I, aVL suggests a possibility of RCA lesion. PMID- 15656027 TI - Chronic hepatitis C in northern India--the pathological and clinical spectrum. AB - BACKGROUND: The aim of the study was to analyse the morphology, clinical presentation and predisposing factors for chronic hepatitis C infection. METHODS: Clinical presentation of 220 patients who presented with hepatitis C virus infection over five years period (January 1996 to December 2000) were recorded. Liver biopsy specimens from 80 adult patients with chronic hepatitis C virus were evaluated using a semiquantitative scoring system. The possible predisposing risk factors for infection in these patients were recorded. RESULTS: Grading of chronic hepatitis C was minimal/mild in 51 (64%) cases. Seven (8.7%) had high grade necroinflammatory activity. Fibrosis was absent in 22(27.5%), mild in 29(36.25%), moderate in 11 (13.75%) and 18(22.5%) had evidence of cirrhosis. No significant correlation was found between the level of transaminases and degree of fibrosis or grade of inflammation. More number of patients with history of alcohol consumption had moderate/severe grade of necroinflammatory activity and cirrhosis as compared to those not taking alcohol at all. Potential predisposing factors were use of unsterile syringes, previous surgery and tattooing in multivariate analysis. CONCLUSION: Nearly two-thirds of patients of chronic hepatitis C where liver biopsy was possible have minimal/mild disease at time of diagnosis; 22% have cirrhosis. Though disease severity can only be assessed by liver histology, liver biopsy may not be possible in a sizeable proportion of patients as they present with advanced liver disease. Most of the patients do not have specific symptoms. Reusable needles/syringes is the commonest potential risk factor. PMID- 15656028 TI - Evaluation of cholinesterase to differentiate pleural exudates and transudates. AB - AIMS AND OBJECTIVES: The present study was undertaken to evaluate the usefulness of pleural fluid cholinesterase (PChE) level in pleural fluid and its ratio to serum cholinesterase (P/SChE) in order to differentiate transudates and exudates and to compare their diagnostic efficacy with the Light's criteria. MATERIAL AND METHODS: A total of 110 patients of pleural effusion of diverse etiology were studied. Eighty patients were of exudative pleural effusion of tubercular, malignant or parapneumonic origin and 30 patients were of transudative effusion. Cholinesterase was estimated in the pleural fluid and serum in all the patients. RESULTS: The mean PChE and P/S ChE were significantly higher in exudates as compared to transudates (p < 0.001). P/S ChE was 0.79 +/- 0.45 and 0.14 +/- 0.11 in exudates and transudates, respectively. When a cut-off value of 469 IU/L for PChE was taken for the diagnosis, it was found that 10% of exudates and 2.5% of transudates were misclassified. However percentage of misclassification decreased to 1.25% in exudates and 3.3% in transudates when the cut-off value of 0.24 for P/S ChE ratio was used. Using Light's criteria, a sensitivity of 91.25% and specificity of 90% with positive predictive value (PPV) of 96.05% and negative predictive value (NPV) of 79.42% was observed. However using P/S ChE, the PPV was 98.75% and NPV was 96.67%. CONCLUSIONS: The estimation of PChE and P/SChE ratio had better discriminatory capacity than Light's criteria. It is cost effective and more specific, therefore its routine estimation is recommended. PMID- 15656029 TI - Pulmonary involvement in chronic arsenic poisoning from drinking contaminated ground-water. AB - OBJECTIVES: Chronic arsenic poisoning, due to ingestion of contaminated ground water, is a major public health problem in West Bengal. It causes multiorgan damage. The present study attempts to objectively investigate the pulmonary involvement by examining the lung function. The nature of lung changes was also evaluated. MATERIAL AND METHODS: One hundred and seven subjects with (cases) and 52 subjects without (controls) chronic arsenic poisoning were examined by spirometry. Forced expiratory volume-I second (FEVI), forced vital capacity (FVC) and peak expiratory flow rate (PEFR) were measured. Bronchoalveolar lavage (BAL) was performed in five cases with and five cases without pulmonary involvement. RESULTS: Thirty three (30.8%) cases and four (7.6%) controls (p<0.01) had respiratory involvement. The pattern of involvement in cases was: obstructive- 20(68.9%) (including three (10%) with bronchiectasis), restrictive- 1(3.5%), mixed- 8(27.6%), malignancy- 4(12.1%) (adenocarcinoma-I, squamous cell- 2, undifferentiated- I). FEVI (69.7+/-25.9 [n=105] vs 83.7+/-15.19 [n=51], p=0.0005), FVC (77.4+/-22.7 [n=105] vs 85.6+/-18.23 [n=51], p=0.025), FEVI/FVC (73.6+/-13.38 [n=105] vs 79.1+/-18.65 [n=52], p=0.007) and PEFR (53.9+/-21.52 [n= 103] vs 67.3+/-18.36 [n=51], p=0.0002) (percent of predicted) were all reduced more in cases compared to controls. Worsening of these parameters correlated with increasing degree of arsenic toxicity. Markers of inflammation (macrophage, lactate dehydrogenase, nitric oxide) were apparently more in the BAL fluid of those with lung involvement than in those without, though the arsenic content did not differ significantly. CONCLUSION: Chronic arsenic poisoning causes pulmonary involvement, predominantly obstructive, the degree of which worsens with increasing degree of arsenic toxicity. Inflammation, rather than direct toxicity, appears to be the underlying mechanism. PMID- 15656030 TI - Acute pulmonary thromboembolism. PMID- 15656031 TI - Venous infarct of jejunum. PMID- 15656032 TI - Imatinib in gastrointestinal stromal tumors. AB - Gastrointestinal stromal tumors (GIST) are the most common mesenchymal neoplasms of gastrointestinal tract. The tumors express the cell surface transmembrane receptor KIT that has a tyrosine kinase activity and is a protein product of KIT protoeoncogene. These tumors occur in the whole of Gastrointestinal tract. Treatment includes surgical resection for localized tumors. For metastatic disease treatment options include systemic chemotherapy, radiation therapy, with a response rate of less than 10%. Presently Imatinib; a tyrosine kinase inhibitor has shown promising result with response rates upto 59-69% in phase II results in metastatic setting; and ongoing phase II & phase III trials in adjuvant setting will help to establish its role as an adjuvant to surgery. We have treated eleven patients of metastatic GIST with Imatinib and we hereby present these cases. PMID- 15656033 TI - Chronic myeloid leukemia. AB - Chronic myeloid leukemia is one of the commonest hematological malignancies seen in clinical practice. It is the result of abnormal and excess cell proliferation due to de-regulated bcr-abl tyrosine kinase activity as a result of Philadelphia chromosome. The present article discusses the various options available to treat the disorder. Allogeneic stem cell transplant remains the gold standard and the only curative option. Hydroxyurea and Busulfan helps in controlling the total leukocyte count but fail to impact on survival. Interferon especially when combined with cytarabine is curative in minority of patients though a substantial number of patients achieve functional cure. Imatinib, a molecular targeted oral therapy, against bcr-abl tyrosine kinase is the latest addition to various treatment options. Early results appear very promising and can be considered as non- transplant standard of care. PMID- 15656034 TI - Melioidosis; the remarkable imitator: recent perspectives. AB - Melioidosis is an important public health problem in some regions of the world. It is endemic in South East Asia. The clinical spectrum of melioidosis is extremely broad, and melioidosis requires awareness on the part of the clinician and the existence of a laboratory capable of isolating and identifying Burkholderia pseudomallei, the etiological agent of melioidosis. Beta-lactams such as ceftazidime is currently the treatment of choice. There is no vaccine licensed for human use. There is an urgent need for rapid diagnostic techniques and effective treatments that are affordable in countries where the disease is endemic. PMID- 15656035 TI - Permanent hypoparathyroidism following radioiodine treatment for hyperthyroidism. AB - Persistent hypoparathyroidism following 131I treatment is a rare but recognized complication. A case is presented, where a relatively small dose of 131I produced persistent hypocalcaemia. PMID- 15656036 TI - Myelodysplasia and acute myeloid leukaemia in a case of rheumatoid arthritis with secondary amyloidosis treated with chlorambucil. AB - Immunosuppressive therapy related secondary haematologic malignancy is well reported. A 52 years lady with established rheumatoid arthritis developed reactive amyloidosis. This was initially treated with colchicine and cyclophosphamide and later with chlorambucil. Ten months after stopping chlorambucil she developed pancytopenia and vitamin B12 deficient megaloblastic anaemia. The pancytopenia was refractory to vitamin B12 supplements and a repeat bone marrow confirmed myelodysplasia (FABI RAEB-T). Within three weeks of this diagnosis she evolved into acute myeloid leukaemia and expired due to refractory thrombocytopenia and uncontrolled bleeding. This case stresses the need for long term follow up of RA patients treated with alkylating agents. PMID- 15656037 TI - Sir David Bruce (1855-1931) postal stamps released to commemorate Anti Brucellosis Congress-Malta 1964. PMID- 15656038 TI - Losartan-ramipril combination treatment. PMID- 15656039 TI - In-vitro activity of linezolid against clinical isolates of Gram-positive cocci in a tertiary care hospital. PMID- 15656040 TI - Risk factors for coronary artery disease in Indians. PMID- 15656041 TI - Lipoprotein(a) as a marker of coronary artery disease and its association with dietary fat. AB - OBJECTIVE: The main objectives of the study were to evaluate the effect of dietary fat on plasma lipoprotein(a) [Lp(a)] levels and to study the potential of Lp(a) as a more reliable marker for CAD compared to other lipids and lipoproteins. METHODS: Twenty CAD patients and 20 healthy controls were recruited for the study. Their fasting plasma Lp(a) levels and complete lipid profile were assayed. The fat intake was calculated using 24 hours dietary recall method. The patients and controls were each divided into two subgroups: Group A consuming dietary fat > 30% and Group B consuming dietary fat < or = 30% of the total kilo calories/day. RESULTS: Results indicated that plasma Lp(a), total serum cholesterol (TC), tryglyceride (TG), low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C) and LDL-C/HDL-C ratio of CAD patients were significantly higher than the controls. High fat intake was found to be associated with higher plasma Lp(a) levels (p<0.05) in patients only. No significant correlation was found between Lp(a) levels and other conventional lipoproteins. CONCLUSION: The lack of correlation between Lp(a) and other lipoproteins indicates its potential as an independent risk factor for CAD. High fat intake led to higher plasma Lp(a) levels in patients; hence it would be worthwhile to evaluate the effect of quality and quantity of fat intake on plasma Lp(a) levels in a larger sample size. PMID- 15656042 TI - Association of major modifiable risk factors among patients with coronary artery disease--a retrospective analysis. AB - BACKGROUND: The relative importance of various risk factors varies in different regions of India. This was a retrospective study of patients with recently diagnosed coronary artery disease to assess four major risk factors: dyslipidemia, hypertension, smoking and diabetes. MATERIAL AND METHODS: A total of 5748 patients (4952 males, 796 females) with recently diagnosed coronary artery disease were analysed from the records of Heart Hospital along with 8103 controls (6092 males and 2011 females). Absolute lipid levels as well as prevalence of dyslipidemia using the ATP III guidelines were assessed. They were classified into two major groups premature CAD (males < 45 years females < 55 years) and CAD at usual age (males > or = 45 years, females > or = 55 years). RESULTS: The most common pattern of CAD was chronic stable angina (n=2773, 48.24%). Mean total cholesterol (TC), LDL cholesterol and TC/HDL ratio were significantly higher in subjects with CAD compared to subjects without CAD controls. The mean HDL cholesterol and triglyceride levels were similar in both groups. Elevated LDL cholesterol, decreased HDL cholesterol, elevated total cholesterol and abnormal TC/HDL ratios were more common in CAD patients as compared to controls (38.8% vs. 33.14%, 29.3% vs 18.2%, 36.9% vs 32.5% and 59.05% vs 44.3% respectively). However lipid abnormalities were not significantly different in females > or = 55 in CAD vs non-CAD group. Smoking was significantly more common in subjects with CAD groups (30.97% vs. 12.72%) as compared to subjects without CAD (P < 0.0001). It was most common in males with premature CAD (44.1 % P < 0.0001). Hypertension was found in 1036 patients (18.02%) and diabetes in 763 (13.28%) as compared to 1126 (13.9%) hypertensives in non-CAD group and 639 diabetics (7.89%) (P = 0.001), both were more common in males > or = 45 and females > or = 55 as compared to those with premature CAD (p < 0.01). CONCLUSION: Among the risk factors assessed, dyslipidemia (particularly abnormal TC/HDL ratio and elevated LDL cholesterol), smoking hypertension and diabetes were associated with coronary artery disease in decreasing order of prevalence. In premature CAD, dyslipidemia and (in males) smoking are of particular importance. PMID- 15656043 TI - Effect of tight glycaemic control on fetal complications in diabetic pregnancies. AB - AIMS: To find the incidence of fetal complications in Indian diabetic mothers with tight glycaemic control (TC), its comparison with other levels of glycaemic control, i.e., acceptable control (AC), uncontrolled (UC), and relevant international data. METHODS: A total of 240 mothers with diabetes mellitus (DM) and pregnancy were risk-matched and selected from the Antenatal Clinic of NRS Medical College, 176 of whom had gestational diabetes mellitus (GDM) and 64 had pregestational diabetes mellitus (PGDM), and were put on exercise, diet and or insulin therapy. Glycaemic parameters monitored include fasting plasma glucose (FPG), 2 hr. postprandial plasma glucose (PPPG) and HbA1C. TC had - FPG < 70 mg/dl, PPPG < 100 mg/dl, HBA1C < 6.5%; AC with FPG 70-95 mg/dl, 2 hr. PPPG 100 120, HBA1C 6.5-7.5% and UC had FPG > 95 mg/ dl, 2 hr. PPPG > 120 mg/dl and HBA1C > 7.5%. Fetal parameters monitored included large-for-date babies (LGA), small for-date babies (SFD), birth asphyxia, perinatal death, neonatal hypoglycemia, neonatal hypocalcaemia and congenital anomalies. RESULTS: (i) LGA-AC had the best results (0% vs. 12.5 and 22.29%); (ii) SFD-TC and AC had worst results (16.7% and 18.18% vs. 0%); (iii) Birth asphyxia-AC fared worse 18.18% vs. 4.16% and 0%; (iv) perinatal death and congenital anomalies showed significant reduction with tight control (4.16% and 0% respectively); (v) Neonatal hypoglycemia is lowered in TC compared with UC while neonatal hypocalcaemia does not show any alteration. For PGDM patients there is little intra-group variability of the parameters. The UC subgroups of GDM fared better than PGDM as far as all complications and congenital anomalies were concerned. Compared with international data, there is a dichotomy of the results of GDM and PGDM. CONCLUSION: For GDM patients all parameters may not be uniformly affected by the same degree of glycaemic control. A tight control may not be theonly factor to decide on the outcomes for PGDM patients. PMID- 15656044 TI - Effect of passive smoking on endothelial function in: healthy adults. AB - BACKGROUND: Active smoking predisposes to atherosclerotic vascular disease but recent evidence that inhalation of environmental tobacco smoke (passive smoking) may also have deleterious cardiovascular effects, has enormous public health implications. Endothelial dysfunction is an important early feature of atherogenic process, which may occur due to passive smoking. OBJECTIVE: To assess the effect of passive smoking on endothelial function (measured by flow-mediated dilatation, a marker of endothelium-dependent arterial dilatation) and compare it with non-smokers. STUDY DESIGN: Case control study. SETTING: Out-Patient Department of Medicine, Government Medical College, Nagpur. PARTICIPANTS: Seventy five young, healthy, male adults between 15-30 years age were studied. There were three groups: (a) Non smokers (n = 25) (b) Passive smokers (n= 25) and (c) Active smokers (n = 25). Subjects with diabetes mellitus, hypertension and ischemic heart disease were excluded. Lipid profile was measured in all. Endothelial function was tested non-invasively by using high frequency linear vascular probe on brachial artery. Resting brachial artery lumen, flow at rest and after hyperemia, flow-mediated dilatation and nitroglycerine-induced dilatation were measured. RESULTS: The mean brachial artery lumen dilatation and flow at rest were similar in all the three groups. Flow-mediated dilatation (FMD%, a marker of endothelium-dependent dilatation and endothelial function) was significantly higher in non-smokers than passive smokers (8.9 +/- 4.8 Vs 5 +/- 2.3, p < 0.01) and also as compared with active smokers (8.9 +/- 4.8 Vs 6.6 +/- 2.2, p < 0.05). Nitroglycerine-induced dilatation, (a marker of endothelium-independent dilatation ) was similar in all the three groups. Serum lipids (mean cholesterol, LDL, and mean LDL/HDL ratio) were statistically significantly higher in passive and active smokers as compared with non-smokers (p < 0.05). CONCLUSION: Like active smoking, passive smoking was also associated with impaired endothelial function, (a key early event in atherogenesis) and altered lipid profile, in healthy young adults. PMID- 15656045 TI - Current status of communicable and non-communicable diseases in India. AB - India is going through a period of transition, both epidemiological and demographic transition. Infectious diseases are still persisting as major health problems in spite of having national programmes for the control of most of these diseases for almost half a century now. This paper focuses on two national programmes: the success story of the National Leprosy Eradication Programme; and the National Anti-Malaria Programme that has failed to achieve its objectives. There are re-emerging infectious diseases which are adding to the burden of diseases. In addition, there is an increasing prevalence of non-communicable diseases as a result of lifestyle changes and urbanization. These are the challenges that are to be tackled in the new millennium. PMID- 15656046 TI - Type 2 diabetes, emphysematous pyelonephritis and emphsematous cholecystitis. PMID- 15656047 TI - Atrial bigeminy presenting as slow regular pulse. PMID- 15656048 TI - Neuromuscular weakness in critically ill. AB - Neuromuscular weakness in critically ill has been increasingly reported in the recent years. Occasionally it may lead to difficulty in weaning the patients from mechanical ventilation, which is difficult to diagnose clinically. Though in well planned studies the incidence has been reported to be high, the diagnosis is often missed due to the presence of various confounding factors in the form of drug effects, underlying disorder and coexisting abnormalities of the central nervous system. A high index of suspicion with detailed neurological and electrophysiological examination is required for an early and accurate diagnosis. A wide spectrum of disorders and drugs can be responsible for the critical illness neuromuscular abnormality. The most frequent and defined disorders include; critical illness polyneuropathy (CIP) which is characterized by a sensorimotor reversible polyneuropathy presenting as distal symmetrical weakness with loss of deep tendon reflexes. Acute myopathy is another important disorder in this group which usually presents with quadriplegia often related to steroid use. Persistent blockade of neuromuscular junction is also defined in critically ill patients. It is, therefore, important to understand these disorders and their implications in the management of these patients. Some of the conditions require prolonged neuro-rehabilitation. The various acquired disorders leading to neuromuscular abnormalities in critical care, and their diagnosis and management are discussed. PMID- 15656049 TI - Dyslipidemia in Asian Indians: determinants and significance. AB - Data suggest that lipid fractions other than total cholesterol, i.e. serum triglycerides (TG) and high-density lipoprotein (HDL) cholesterol are important for the pathogenesis of atherosclerosis. A combination of hypertriglyceridemia, low levels of HDL-cholesterol and high levels of small dense low-density lipoprotein, termed as "atherogenic dyslipidemia', is particularly seen in Asian Indians. Although precise reason for such dyslipidemia is unknown, genetic predisposition and characteristic body composition (excess truncal subcutaneous fat and intraabdominal fat) may be important contributors. A common interface between such body composition and dyslipidemia in Asian Indians is high tendency to develop insulin resistance, more than the other ethnic groups. The general guidelines for the management of dyslipidemia in Asian Indians should be according to National Cholesterol Education Program, Adult Treatment Panel III. However, optimal management requires consideration of ethnic-specific dietary, lifestyle and management factors to formulate individual treatment guidelines. PMID- 15656050 TI - Obstructive sleep apnea: therapies other than CPAP. AB - Nasal application of continuous positive airway pressure (CPAP) is the standard form of therapy for treating obstructive sleep apnea (OSA). Common difficulties associated with CPAP therapy include sense of dryness in the mouth, rhinorrhea, nasal congestion and dryness, mask discomfort, claustrophobia, irritation from device noise, aerophagy, chest discomfort and partner's intolerance. Therefore, many patients are unable to or unwilling to comply with the use of CPAP. This article discusses the various non-CPAP approaches that have been investigated in the management of OSA, which include behavioral therapy (weight loss and positional therapy), pharmacological treatment, mandibular advancement techniques and surgery. However, none of these has been shown to be superior to CPAP. In clinical practice, only selected patients will benefit from therapies other than CPAP. PMID- 15656051 TI - Thrombotic microangiopathies presenting as an obstetric emergency. AB - Thrombotic microangiopathies are disorders that arise due to a diffuse endothelial damage. They predominantly manifest either as thrombotic trombocytopenic purpura (TTP) or hemolytic uremic syndrome (HUS). When they arise as a complication of pregnancy and associated disorders they are associated with a high mortality and morbidity. PMID- 15656052 TI - Acute aortic dissection--De Bakey Type I and Stanford A. AB - We report a rare case of aortic dissection which fits into De Bakey Type I and Stanford A, presented with severe tearing chest pain, paraplegia, stupor, hypotension, and syncope. Echocardiography showed dissection involving aortic root. MRI showed aortic dissection involving ascending, arch and descending aorta. Patient was managed conservatively and he died within 12 hours after the admission. PMID- 15656053 TI - Near-fatal amlodipine poisoning. AB - Amlodipine poisoning is very rare and only few cases have been reported in English literature. We report here a case of severe amlodipine poisoning with non cardiogenic pulmonary edema. PMID- 15656054 TI - Hepatocellular failure in glycogen storage disorder type 3. AB - A case of a 21 years male patient with type 3 glycogen storage disorder diagnosed at necropsy, who died suddenly with hypovolemic shock following a massive upper gastrointestinal bleeding due to hepatocellular failure is reported. Salient features of GSD type 3 are briefly discussed. PMID- 15656055 TI - Solitary plasmacytoma of bone: fourteen years disease-free survival. AB - Solitary plasmacytomas are rare tumors of plasma cell origin categorized as extramedullary plasmacytoma (EP) and solitary plasmacytoma of bone (SPB). We report here a case of SPB who is off treatment for 14 years and is completely disease-free. PMID- 15656056 TI - Medical philately (medical personalities on stamps). Ivan Pavlov (1849-1936). PMID- 15656057 TI - Hand washing in healthcare workers--a challenge that can be met? PMID- 15656058 TI - A potential for interactions between indigenous and prescription drugs. PMID- 15656059 TI - Typhus fever. PMID- 15656060 TI - Docetaxel-induced onycholysis. PMID- 15656062 TI - Self injection of insecticide. PMID- 15656061 TI - Use of sulfonylureas during pregnancy: some incidental observations. PMID- 15656063 TI - Long duration priapism in blast crisis of chronic myeloid leukemia. PMID- 15656064 TI - [Characteristics of suicides in Bogota, 1985-2000]. AB - OBJECTIVE: To establish how suicide's typologies are related with a group of variables described as risk factors and to describe the stability of these typologies during a period of 15 years. METHODS: Observational-analytic study based on autopsy registers of suicides in Bogota in 1985, 1990, 1995 and 2000. In order to determine typologies, the method of multivarioate analysis of multiple correspondences was employed. RESULTS: 171 cases of suicide were detected in 1985, 222 cases in 1990, 273 in 1995 and 329 in 2000. Physical or mental co morbidity, the use of psychoactive substances, access to lethal methods of injury, and the rare use of fire weapons by women, were constant factors during the years studied. At the same time, changes in some typologies were related to the presence of addictions in women and to HIV/AIDS. CONCLUSIONS: Although relatively constant suicide typologies exist during the 15-year period studied, changes in the epidemiological profiles and new life styles generate modifications of these typologies. This suggests that suicide risk factors are a dynamic element that is modified as time passes. PMID- 15656065 TI - [Trends of mortality caused by infectious diseases in Cartagena de Indias, Colombia, 1995-2000: a measure of the epidemiological transition]. AB - OBJECTIVES: Mortality is a definition of, strictly populational nature, different from the concepts of death and decease which reflect the loss of an individ ual biological life. Therefore, mortality expresses the dynamics of deaths along time and space, and only allows for comparisons at this level. Th present study was aimed at describing the tendency of mortality caused b infectious diseases in Cartagena de Indias, Colombia, during the perio 1995-2000, with the purpose of evidencing the epidemiological transition. MATERIALS AND METHODS: The databases of the mortality registers of Cartagena de Indias between 1995 and 2000 were taken as reference. The im pact of mortality was measured by the sum of Years of Potential Life Lost (YPLL) for each group of death causes. To adjust for the temporal preference for the assessment of YPLL a discount rate of 3% was applied in this study. The computerized GESMOR application was used for estimating YPLL, analyzing the mortality reported in the registers using the 9th and 10th versions of the International Classification of Diseases (ICD) to establish the basic causes of death. RESULTS: A mean of 3,506 annual deaths occurred during the period studied, generating on average 60,311 YPLL, which gives a rate of 71.5 YPLL per 1000 inhabitants. This represents a 16% increase which can be explained by an increase in the number of deaths in the lower age groups. For 1995, there was a predominance of group II non-infectious diseases causes (41.9%), followed by group I (30.2%). For the year 2000, the proportional distribution of mortality between the different groups of causes changed, causes in group I constituting the main explanation for causes of YPLL (52.9%). Non-infectious diseases occupied the second place (31.2%). These changes can be explained by the importance of perinatal causes of death, which increased from 14.2% of YPLL in 1995 to 32.7% in 2000. DISCUSSION: It is evident that the greater weight of infectious, maternal, perinatal and nutritional diseases, the slower then the national average decrease in child mortality rates, the greater load of mortality in the male sex, the stability of the group of accidents and injuries at the expense of intentional injuries during the period studied, have caused a stagnation of the epidemiological transition and the instauration of the prolonged polarized model. PMID- 15656066 TI - [Association between the cervico-vaginal inflammatory cytology and the intraepithelial cervical lesion in patients from a sexual and reproductive health clinic in Bogota, Colombia, 1999-2003]. AB - OBJECTIVES: Cervical cancer is the most common malignancy affecting Colombian women. Pap smear screening is used for its early detection. Early stages of cervical cancer can be expressed by the presence of inflammatory atypia in the Pap smear. The main objective of this study is to evaluate the role of inflammatory atypia in Pap smear as a marker of squamous intraepithelial lesions (SIL), given that it is not clear how to deal with the presence of inflammatory cells in Pap smear. MATERIALS AND METHODS: A cross sectional study was carried out to establish the operative characteristics of the cervical Pap smear in the colposcopy clinic of the Clinica Piloto Profamilia, Bogota, between January 1999 and December 2003. We compared patients with biopsies reporting SIL and those with a benign report in the pathologic study. We evaluated sensitivity the and the specificity of the Pap smear in different cutting points compared to the pathology report as gold standard. RESULTS: The prevalence found was 48,7 % for low grade - Squamous Intrepithelial Lesions (SIL) and 9,7 % for high grade - SIL. The best performance for the Pap smear test to detect SIL was when the atypical squamous cells of undetermined significance (ASCUS) were considered as abnormal. Severe inflammatory atypia were associated to SIL in a similar way as to ASCUS. CONCLUSIONS: Sensitivity of the Pap smear as a screening test decreases when excluding Pap smears reporting ASCUS as a cutting point. In our environment, we recommend to report inflammatory atypia on the Pap smear test stratified by severity until the role of severe inflammatory atypia for the diagnosis of SIL is defined. PMID- 15656067 TI - [Effects of child labor upon the health of the child worker]. AB - OBJECTIVES: This article presents a review of the evidence on child labor, in three aspects: 1. The characteristics of children which make them vulnerable to labor; 2. The diseases and accidents related with child labor, and 3. The injuries associated with the childs' occupations, such as agriculture, construction, manufacture, textiles, leather industry, street sales and sales in marketplaces. METHODS: The Medline, EBSCO, PROQUEST and HINARI databases were searched, and material from the International Labor Organization, the American Academy of Pediatrics and the American Association of Psychology was reviewed. RESULTS: Information regarding occupational risk in children is poor as compared with information in adults. Children have characteristics which make them particularly vulnerable to occupational risks and there are no clear standards to protect them. PMID- 15656068 TI - [Incorporation of the Hazard Analysis and Critical Control Point system (HACCP) in food legislation]. AB - The Hazard Analysis and Critical Control Point system (HACCP), recommended by different international organizations as the Codex Alimentarius Commission, the World Trade Organization (WTO), the International Office of Epizootics (OIE) and the International Convention for Vegetables Protection (ICPV) amongst others, contributes to ensuring the innocuity of food along the agro-alimentary chain and requires of Good Manufacturing Practices (GMP) for its implementation, GMP's which are legislated in most countries. Since 1997, Colombia has set rules and legislation for application of HACCP system in agreement with international standards. This paper discusses the potential and difficulties of the legislation enforcement and suggests some policy implications towards food safety. PMID- 15656069 TI - [Validating scales used for measuring factors in medicine]. AB - This article shows the mayor issues related with validation of scales applied in health research and illustrates the steps of this process. Concepts related with the selection of items, translation, validity, reliability and usefulness are discussed in this article. PMID- 15656070 TI - One hundred years of trams. PMID- 15656071 TI - Current concepts in periodontal pathogenesis. AB - Periodontal research over the last 40 years has been remarkably prolific. We now understand that severe periodontitis affects approximately 10-15% of the population (representing a large number of individuals in the UK) and gingivitis and mild periodontitis affect a majority of people. Microbiological research has identified some of the key pathogens that are implicated in periodontal disease. Plaque bacteria exist in biofilms, which have evolved to protect individual organisms within the subgingival bacterial community. For this reason, root surface instrumentation (RSI) remains the cornerstone of periodontal treatment, and is necessary to disrupt the subgingival biofilm mechanically and reduce the bacterial bioburden. Although bacteria are necessary for periodontal disease to occur, a susceptible host is also required. The immune-inflammatory response that develops in the gingival and periodontal tissues in response to the chronic presence of plaque bacteria results in destruction of structural components of the periodontium leading, ultimately, to the clinical signs of periodontitis. The nature of the host response is determined primarily by genetic factors and environmental and acquired factors such as smoking. The host response is essentially protective in nature, but both under-activity (hypo-responsiveness) and over-activity (hyper-responsiveness) of aspects of the host response can result in enhanced tissue destruction. The purpose of this paper is to review current thinking in periodontics with special reference to periodontal epidemiology, microbiology, and pathogenesis. PMID- 15656072 TI - What's new in dentine bonding? Self-etch adhesives. AB - Bonding to dentine is an integral part of contemporary restorative dentistry, but early systems were not user-friendly. The introduction of new systems which have a reduced number of steps--the self-etch adhesives--could therefore be an advantage to clinicians, provided that they are as effective as previous adhesives. These new self-etch materials appear to form hybrid layers as did the previous generation of materials. However, there is a need for further clinical research on these new materials. Advantages of self-etch systems include, no need to etch and rinse, reduced post-operative sensitivity and low technique sensitivity. Disadvantages include, the inhibition of set of self- or dual-cure resin materials and the need to roughen untreated enamel surfaces prior to bonding. PMID- 15656073 TI - A cemento-ossifying fibroma in the maxilla. AB - Cemento-ossifying fibroma is an uncommon, benign lesion, which can lead to swelling and deformity of the face. This report describes an uncommon presentation of this lesion with simple and effective measures to diagnose and treat it. PMID- 15656074 TI - Autotransplantation of an impacted third molar: an orthodontic case report. AB - This case report describes the use of autotransplantation as part of an orthodontic treatment plan where there was early loss of an upper molar tooth secondary to irreversible pulpitis. An impacted lower third molar, which had symptoms of pericoronitis, was transplanted into the upper left second molar socket. Fixed orthodontic treatment, to correct the patient's initial complaint of crowding, was completed in 22 months. The transplanted tooth remained vital and functional throughout. PMID- 15656075 TI - Severe extrusion from an apical abscess: a case report. AB - A 10-year-old boy presented with an acute apical abscess with extensive extrusion of a maxillary central incisor. The tooth, the maxillary right central incisor, had a history of trauma and subsequent apical abscesses. This article describes the presentation, management of the abscess and the subsequent results of treatment. The reasons for the extrusion and success of the treatment are discussed. PMID- 15656076 TI - An overview of bleaching techniques: I. History, chemistry, safety and legal aspects. AB - The use of a variety of bleaching techniques has attracted much interest from the profession, as they are non-invasive and relatively simple to carry out. Coupled with the uncertain legal situation within the European community, and especially within the UK, this series of articles hopes to give a broad overview of bleaching techniques, their efficacy and relative safety, as well as update the current legal situation. This article will give an overview of bleaching: history, chemistry and safety. In addition, it will summarize types of tooth discoloration, along with indications/contra-indications for bleaching. Future articles will address in detail both home and power bleaching techniques, as well as the various ways to bleach non-vital teeth. PMID- 15656077 TI - Management of dental patients on warfarin therapy in a primary care setting. PMID- 15656078 TI - Dental Olympians. PMID- 15656079 TI - Dental Project Peru. PMID- 15656080 TI - [Job stress among Japan Overseas Cooperation volunteers--using the Brief Job Stress Questionnaire]. AB - Mental health problems have recently increased among Japan Overseas Cooperation volunteers since 1965, when the Ministry of Foreign Affairs (Japan International Cooperation Agency) launched this volunteer work project for improving hygiene and socioeconomic conditions in developing countries. There was little research on job stress among them dispatched despite previous surveys indicating job as an important stressor. To investigate stress and job-related stressors among them, we conducted a cross-sectional epidemiological study from October to December in 2003. The subjects were all 1,084 Japan Overseas Cooperation volunteers aged 20 40, who worked in 67 countries worldwide at the time of this study (485 and 599 males and females, 316, 332 and 436 for those staying overseas for 11, 7 and 4 months, respectively). Approximately 80% were involved in their dispatching occupational organizations as professionals in information technology, health & welfare, education, and research. Our main outcome measure used was the Brief Job Stress Questionnaire, which was developed to assess stress and job-related stressors or buffers for Japanese workers. Demographic and personality (Egogram) characteristics as well as other health information were obtained. The response rate was 86.9%. For psychological stress, prevalence was 5.5% (n = 49). Means (+/ SD) were 4.22 (+/- 3.98), and 4.89 (+/- 4.40) for males and females (p < 0.05), and 5.15 (+/- 4.17), 5.05 (+/- 4.45), 3.93 (+/- 4.40) for those staying overseas for 11, 7 and 4 months (p < 0.01), respectively. For physical stress, prevalence was 2.9% (n = 26). Means (+/- SD) were 1.10 (+/- 1.68), and 1.41 (+/- 1.74) for males and females (p < 0.01), and 1.47 (+/- 1.77), 1.35 (+/- 1.89), 1.11 (+/- 1.55) for those staying overseas for 11, 7 and 4 months (p < 0.05), respectively. The factors significantly associated with psychological stress were high job demand, poor human relationships at work, low job suitability, low social support from supervisors and colleagues, and being dissatisfied with their life, according to multiple logistic regression analysis. The present study suggested that psychological stress was more prevalent than physical. It also implied a significant relationship between psychological stress and job-related stressors among the subjects of this study as in employees in Japan. Mental health check ups and counseling in the early stage of psychological stress is important from the viewpoint of prevention of developing stress-related mental health disorders. Education on stress-coping skills should be considered in a training program before they are sent overseas. PMID- 15656081 TI - [Survey on visual and musculoskeletal symptoms in VDT workers]. AB - With the spread of visual display terminals (VDT) in offices, the numbers of workers using VDT and the working hours at such equipment have increased rapidly in recent years. Also, preventive measures for fatigue have been proposed and the office-working environment has been improved. To examine the effects of the rapid changes in working conditions and environment on the health of VDT workers, we conducted a questionnaire survey in 2002. A self-reported questionnaire was distributed to 3,927 office workers; 2,374 (60.5%) responded. Subjects whose questionnaires had missing data were excluded from analysis. As a result, 1,406 (male: 1,069, female: 337) workers aged 20 to 59 were subjected to analysis. By a logistic regression model, we examined the association between VDT use and visual and musculoskeletal symptoms. Prevalence of eye strain and/or pain (72.1%) was the highest, followed by neck stiffness and/or pain (59.3%), low back stiffness and/or pain (30.0%) and hand or arm strain and/or pain (13.9%). Women consistently reported more discomfort than men. As a result of the logistic regression model, eye strain and/or pain was associated with dissatisfaction with airflow, but not with factors affecting visual symptoms as reported in previous studies, for example, reflection of light and blurred characters on the screen. It was thought that airflow appeared as a risk factor because the lighting environment had been improved in offices to prevent reflection of light on the screen. Neck stiffness and/or pain was associated with raising the shoulders during VDT work, the unsuitable shape of the computer mouse for a hand, and the inconvenient arrangement of the mouse in relation to the body. Hand or arm strain and/or pain were associated with the arrangement of the mouse and inappropriate height of the desk. Low back stiffness and/or pain were associated with dissatisfaction with the chair and using the keyboard without a wrist rest. Although measures to prevent fatigue had been implemented for VDT workers, risk factors for musculoskeletal symptoms would be the same as in previous studies. PMID- 15656082 TI - [Measuring workplace climate: reliability and validity of the 12-item Organizational Climate Scale (OCS-12)]. AB - In order to investigate the reliability and validity of the short version of the 30-item Organizational Climate Scale (OCS-30; Toshima and Matsuda, 1992, 1995), a self-administered questionnaire was conducted in a sample of 819 employees of two medium-sized private companies in Japan by using the OCS-30, the Generic Job Stress Questionnaire (GJSQ), and the 12-item General Health Questionnaire (GHQ 12). The OCS has two subscales, i.e., the Tradition Scale (TS) and the Organizational Environment Scale (OES). The organizational climate perceived by each worker can be grouped into four categories based on the subscale scores: low TS and high OES (Active), high TS and high OES (Governed), low TS and low OES (Disorganized), and high TS and low OES (Reluctant). Principal component analysis for the OCS-30 was submitted (varimax rotation, the number of factors = 2), and 6 items for each factor, with factor loadings greater than 0.50, were selected for the short version, which constituted the 12-item Organizational Climate Scale (OCS-12). Cronbach's alpha reliability coefficients of the two subscales of the OCS-12 were acceptable; 0.63 for the TS and 0.71 for the OES. Both two subscales of the OCS-12 were significantly correlated with the GHQ-12 and many subscales of the GJSQ, which indicated the good constructive validity of the OCS-12. Among 4 types of organizational climate categorized by the OCS-12, the "Active" group showed the lowest job stress scores. It is suggested that the OCS-12 could be a reliable and valid instrument for assessing workers' perception of workplace climate. PMID- 15656084 TI - [Methods of examination and diagnosis in ophthalmology]. PMID- 15656083 TI - [Effects of exposure to occupational hand-arm vibration on maintenance of postural balance]. AB - In order to determine the relationship between exposure to hand-arm vibration through the use of vibration tools and dysfunction in the maintenance of postural balance, 106 male forestry workers were examined by stabilometry for deflection in the center of gravity and in the air conduction hearing levels. A questionnaire survey was also conducted among the workers in order to obtain details regarding their age, the types of tools used by them, and the duration for which they had used a chain saw. The vibration acceleration of a chain saw has been limited to a level of 3 G or less since 1976 in accordance with the notification from the Japanese Forestry Agency and the Ministry of Labor. In fact, chain saws with significantly reduced vibration acceleration in comparison with those used before 1976 have been available. Therefore, in 2000, we conducted a test on forestry workers who were divided into two groups-workers who had used a chain saw for 25 years or more (25-yr-or-more group) and workers who had used a chain saw for 24 yr or less (24-yr-or-less group). Compared to the 24-yr-or-less group, the 25-yr-or-more group exhibited significantly higher levels of average deflection in the center of gravity, expressed as the enveloped (aENV) and rectangular (aREC) areas, and in the hearing levels at 500, 1000, 2000, 4000 and 8000 Hz. For the aENV, the correlation coefficients revealed significant relationships between the hearing levels at 4,000 Hz, the duration of use of a chain saw, and age. Since the duration of use of a chain saw exhibited a significant relationship with age, it was necessary to eliminate the effect of age on the aENV. Subsequently, we divided all the workers into age groups spanning ten years each (from 20 to 70 yr) and compared the aENV among the same age groups in both the 25-yr-or-more and the 24-yr-or-less groups. The averages of the aENV for each age group were higher in the 25-yr-or-more group than in the 24-yr-or-less group. In particular, a significant difference was observed in the group of workers in their 40s. Moreover, the average of aENV value was significantly higher in the 25-yr-pr-more group than in the 24-yr-or-less group among the workers between 46 and 68 yr, with their ages being congruent in both groups. While investigating the impact on the workers who had used chain saws, it might be difficult to examine the effect of occupational vibration independent of the noise load. Nevertheless, this study suggests the possibility that a decrease in the maintenance of postural balance has a stronger relationship with exposure to thumping vibration in the past than with the age factor. PMID- 15656085 TI - [Variation of inflammatory reaction of ciliary body--harmony between clinic and basic science]. AB - Histopathological findings of ciliary epithelium and muscle were discussed in the inflammatory conditions that are induced in endophthalmitis, blood-aqueous barrier destruction, cyclocryotherapy and cyclophotocoagulation. The clinical results were also discussed regarding the reduction of aqueous production and uveoscleroplasty which are introduced by the experimental cyclophotocoagulation at the pars plicata/pars plana. Tissue destruction by inflammatory reaction occurred in not only the ciliary body but also in the retina if the endophthalmitis had progressed to the point of severity. The post-inflammatory reactive hypertrophy of nonpigmented and pigmented ciliary epithelium was mild at pars plicata and the anterior portion of the pars plana. On the other hand, it was severe at the posterior portion of the pars plana and the ora serrata, and it was clear that the cyclitic membrane developed into proliferative vitreoretinopathy. Adult pig cultured nonpigmented ciliary epithelium showed proliferated tissue resembling the cyclitic membrane. These results show that nonpigmented ciliary epithelium has a strong proliferative activity as much the same as fibroblasts. The destruction of the blood-aqueous barrier was produced by corneal perforation, paracentesis, prostaglandin E1 (PGE1) subconjunctival injection, and hyperosmotic agent intraophthalmic artery injection. The localization of the tissue destruction was made clear at the beginning portion of the ciliary process and the posterior part of the pars plana. The repetitive instillation of latanoprost and cyclosporin A eye drops showed the same localization of the tissue damage. These results suggest that the beginning portion of the ciliary process is vulnerable to inflammatory agents. Cyclosporin A might produce a toxic reaction on nonpigmented ciliary epithelium, and latanoprost destroys the blood-aqueous barrier. Cyclocryotherapy produces severe necrosis of the pigmented ciliary epithelium and melanocytes, and atrophy of pigmented ciliary epithelium and ciliary muscle, and the proliferation of nonpigmented ciliary epithelium follows. Because cyclocryotherapy is a blind therapy, hypotony and phthisis bulbi will occur if the pars plicata is damaged severely, and the intraocular pressure (IOP) decrease will not be achieved if pars plana is damaged locally. Finally, we demonstrated that it is difficult to predict the therapeutic outcome of cyclocryotherapy. Cyclophotocoagulation of the pars plicata and the pars plana produced severe necrosis of pigmented ciliary epithelium and melanocytes, and atrophy of pigmented ciliary epithelium and ciliary muscle, and the proliferation of nonpigmented ciliary epithelium followed. Cyclitic membrane was developed from the proliferation of nonpigmented ciliary epithelium depending on the severity of photocoagulation. From these experiments the complete destruction of the pars plicata might result in for hypotony and phthisis bulbi. The reduction of aqueous production for angle closure glaucoma was achieved by moderate cyclophotocoagulation of the pars plicata. On the other hand, moderate cyclophotocoagulation of the pars plicata for open-angle glaucoma produced the obstruction of uveoscleral aqueous outflow which compensated for the reduction of aqueous production. From these results we suggested that moderate cyclophotocoagulation of the pars plana for open-angle glaucoma might be necessary to effect the increase of uveoscleral aqueous outflow. Moderate and severe cyclophotocoagulation of the transit area between the pars plicata and the pars plana might bring about reduction of aqueous production and the increase of uveoscleral aqueous outflow. PMID- 15656086 TI - [Progress in pathogenesis and therapeutic research in retinitis pigmentosa and age-related macular degeneration]. AB - Retinitis pigmentosa (RP) and age-related macular degeneration (AMD) are designated special targeted eye diseases by the Welfare and Labor Ministry of Japan. We have been studying the pathogenesis, diagnosis, treatment, and evaluation of these diseases. The development of molecular genetic analyses of RP revealed that the type and frequency of mutations varied with the ethnic population. In our present study, we focused on the genetic analysis and clinical examinations for autosomal dominant retinitis pigmentosa (ADRP). We screened 96 unrelated ADRP families with 9 genes, which included rhodopsin, peripherin/RDS, RP 1, NRL, FSCN 2, PRPF 31, PRPC 8, HPRP 3, IMPDH 1. We also showed the correlations we have found between the phenotype and genotype of hereditary retinal diseases in Japanese patients. Our mutation screenings suggested that Japanese patients with ADRP might have a unique mutation, because the mutation in the FSCN 2 gene has been found only in Japanese patients. On the other hand, the Pro347Leu and Pro23His mutations in the rhodopsin, the Arg677X mutation in the RP 1, and the Asp226Asn mutation in the IMPDH 1 genes are representative mutations for ADRP, and are not found or are very rare in Japanese patients with ADRP. The results of randomized controlled trials of low-dose radiation for wet-type age related macular degeneration located at the fovea centralis indicate the effectiveness of this treatment for maintaining visual acuity and regression of choroidal neovascular membrane (CNV) for at least one-year. Simple surgical removal of CNV or transplantation of autologous cultured iris pigment epithelium (IPE) with vitreous surgery showed some improvement of vision. In either RP or AMD, photoreceptors die, in most cases by apoptosis. Neurotrophic factors (NT) are effective for reducing these processes and preventing photoreceptor cell death in animal models. To apply these methods to humans, the procedures are as follows: 1) obtaining IPE by peripheral iridectomy, 2) culturing it with autologous serum and transfecting the cDNA of NT, and then 3) transplantation of these cells under the retina. We used cDNA of brain-derived neurotrophic factor (BDNF) with adeno-associated virus (AAV) as a vector. These ex vivo procedures were safe and very effective for preventing photoreceptor cell death in animal models, such as RCS rats and light-damaged rats. In the future, these procedures could be applied for RP or AMD and might show some clinical effects for maintaining or improving the vision of patients. PMID- 15656087 TI - [Quantitative assessment of quality of vision]. AB - The importance of quality of vision (QOV) along with quality of life (QOL) in medicine has been recently widely recognized. We have conducted studies to quantitatively analyze factors related to QOV. Irregular astigmatism can be a significant obstacle for achieving satisfactory QOV. Videokeratography data were broken down using Fourier harmonic series analysis into spherical power, regular astigmatism (second harmonic component, n = 2), asymmetry (n = 1), and higher order irregularity (n > or = 3). The irregular astigmatism component calculated by the Fourier analysis significantly correlated with best spectacle-corrected visual acuity. Software was developed to display color-coded maps for the four Fourier indices. The normal range was defined for each Fourier index, and eyes with pathologic and postsurgical conditions were evaluated using the normal range. Progression of keratoconus over time was quantitatively described by Fourier analysis of the videokeratography data. Using the Fourier method, changes in corneal topography following suture removal after penetrating keratoplasty were evaluated. Fourier analysis of videokeratography data significantly facilitated determination of refraction and measurement of best spectacle corrected visual acuity in eyes with corneal irregular astigmatism such as post penetrating keratoplasty eyes. Higher-order wavefront aberrations of the cornea were calculated by expanding videokeratography elevation data into Zernike polynomials, and coma and spherical aberrations were computed. For ocular aberrations, the data obtained with the Hartmann-Shack sensor were decomposed into Zernike polynomials. Coma aberrations of the cornea significantly correlated with age, while corneal spherical aberrations showed no age-related changes. The time-course of changes in corneal higher-order aberrations was reported for photorefractive keratectomy and laser in situ keratomileusis (LASIK). For ocular aberrations, the degree of tilting of the suture-fixed intraocular lens significantly correlated with the amount of coma aberration of the eye. In normal eyes, the ocular coma increased with age mainly because of the increase in the corneal coma, and the ocular spherical aberration increased with age because of the increase in spherical aberration in the internal optics including the crystalline lens. The conventional LASIK significantly increased ocular higher order aberrations, which compromised postoperative contrast sensitivity and low contrast visual acuity. Both corneal and ocular wavefront aberrations were analyzed in cataract, pseudophakic and aphakic eyes, and the equilibrium of spherical aberrations between the cornea and the eye in those conditions was investigated. In pseudophakic eyes, coma aberrations of the cornea, along with the corneal multifocality, significantly contributed to apparent accommodation. Computer simulation indicated that a focus shift of 0.5 diopters deteriorated the retinal image significantly more in eyes without higher-order aberrations than in eyes having a moderate amount of coma aberrations. Clinical results of aspherical intraocular lens were reported for wavefront analysis and contrast sensitivity measurements, and retinal images were analyzed by simulation. For the assessment of vision-related QOL, National Eye Institute Visual Functioning Questionnaire 25 (NEI-VFQ 25) was translated into Japanese. After the validation study, the influence of cataract surgery on QOL was investigated. The QOL score was severely impaired in patients with bilateral cataract, which was significantly and dramatically improved by surgery. Forward scattering, backward scattering, and wavefront aberration induced by the crystalline lens were quantified, and the impact of these factors on visual function was analyzed according to the type of cataract. The degree of QOL improvement by surgery was assessed in relation to the type of cataract and intensity of scattering and aberration. By comprehensively analyzing these factors, quantitative parameters could be developed in the near future to describe progression of cataract and determine the indication for cataract surgery. PMID- 15656088 TI - [Development of a new evaluation system for visual function]. AB - Visual deterioration is caused principally by media opacity, by retinal damage, or by disorders of the higher visual system posterior to the optic nerve. In this article, we focused on media and retinal disorders and clinically evaluated a newly developed system for visual function. The decrease in visual function in cataract is subjectively well evaluated by contrast sensitivity, but is difficult to evaluate objectively. Recently, a wavefront sensor has been developed and ocular higher-order aberration (HOA) can now be measured objectively. We studied the relationship between age and HOA by wavefront sensor and found that HOA, especially spherical aberration in the lens, increased abruptly at the presbyopic age. We also found that the against-the-rule astigmatism in the lens increased at this age. Next, we investigated monocular diplopia which was presumably caused by HOA. Nine eyes with monocular triplopia and with mild nuclear cataract showed significant increase of trefoil aberration and negative spherical aberration (p<0.001). The simulated retinal Landolt's image from these two aberrations showed a triple configuration. Thus we confirmed that triplopia is caused by the combination of trefoil aberration and negative spherical aberration in early nuclear cataract. Next, we investigated whether Area under Log Contrast Sensitivity Function (AULCSF) can be predicted by HOA and light scattering. Backward light scattering (BLS) was evaluated by Schei mpflug image and the forward light scattering (FLS) by a spot size of the Hartmann image compensated for the effect of HOA. HOA was evaluated by root mean square (RMS) value from Hartmann image in a 4 mm pupil. Multiple linear regression revealed that AULCSF could be predicted from BLS, FLS, and HOA, in which each parameter contributed to the prediction significantly (p<0.01). By using this predicted value of AULCSF, the improvement of vision after cataract surgery can be predicted in cataract complicated by retinal disease. It has been reported that by fundus camera equipped with adaptive optics (AO), which is an application of wavefront analysis, photoreceptors could be visualized 2-dimensionally. We developed a compact AO fundus camera and demonstrated that cones were separately analyzed at retinal loci 1 degree temporal to the fovea centralis when the ocular aberration was reduced to less than 0.1 microm RMS in a 6 mm pupil. We are going to use this apparatus for eyes with retinal disease. The functional evaluation of residual retinal ganglion cells (RGCs) in the retina with damage to the photoreceptors is critically important for selecting candidates for artificial retina or regenerative therapy. Transcorneal electrical stimulation (TES) of the retina via contact lens electrodes evokes phosphene and indirect pupillary reflex. The threshold current for evoking phosphene in severely degenerated retinas with visual acuity worse than counting fingers showed a wide distribution, which suggested that TES was useful to evaluate residual RGCs functionally. TES also showed a neuroprotective effect. We suggest that the activation of glial cells by TES up-regulates the production of IGF-1, which eventually protects RGCs. The peripheral retina is important for walking even though the spatial resolution is not high. We developed an apparatus to measure stereopsis in the peripheral retina using a wide screen, and evaluated the peripheral stereopsis of 12 patients after macular translocation surgery and squint surgery. All three patients who showed peripheral stereopsis had only a small amount of squint angle. This method may be useful to evaluate stereopsis in patients with a central scotoma. PMID- 15656089 TI - [A new approach for studying the retinal and choroidal circulation]. AB - One advantage of advanced computer technology is the high throughput with which the retinal and the choroidal circulation can be evaluated from new aspects. To study the choroidal circulation, we first reevaluated indocyanine green video angiography to improve the visualization of indocyanine green (ICG) images, then applied computer technology to analyze images obtained by an ICG video camera system. We also developed a new instrument to measure oxygen saturation levels in the fundus using spectral retinal imaging technology. I. Choroidal circulation. 1. Reevaluation of ICG video camera system: For this purpose, the bio-chemical nature of ICG was studied. 1) Spectral absorption of ICG: The peak absorption of ICG in distilled water was 780 nm as measured with a spectrophotometer. Its maximum absorption shifted from 780 nm to 805 nm after gradually mixing ICG with human serum protein. Conjugation time of ICG as well as fluorescein sodium with human serum protein was then measured by a stopped flowmeter. It was found that fluorescein sodium conjugated with human serum protein within a few milliseconds, while ICG required more than 600 seconds before equilibrium of the binding was reached. From these observations, we developed a new ICG video system with dual light sources; one, a 780 nm diode laser for the early dye filling phase, and the other, a 805 nm diode laser for the later phase of ICG angiography. 2) Binding properties of ICG in human blood: Blood samples were obtained from three healthy volunteers after intravenous administration of ICG. The resulting plasma samples were fractionated by agarose gel immunoelectrophoresis and polyacrylamide gel DISC electrophoresis. The electrophoretic pattern obtained by each method was observed with an ICG fundus video system. We also studied the affinity of ICG for lipids that are common molecular components of lipoproteins such as high-density lipoprotein (HDL) and low-density lipoprotein (LDL). Four kinds of ICG solutions mixed with phospholipid, free cholesterol, esterified cholesterol, and triacrylglycerol were observed with the ICG fundus video system. Both electrophoretic studies showed that ICG bound intensely to HDL and moderately to LDL, and only the solution with phospholipid fluoresced brightly when observed with the ICG fundus video system. 2. Residual fundus ICG fluorescence: Residual fundus fluorescence observed in the late phase of ICG angiography may be delineated differently in normal subjects and in patients with age-related macular degeneration (ARMD). We performed ICG angiography on 8 normal subjects aged below 36 years (8 eyes), 9 normal subjects aged above 62 years (9 eyes), and 21 patients with ARMD aged 50 to 88 years (37 eyes). The intensity and pattern of fluorescence from angiograms obtained in the ultra-late phase, 24 hours after dye injection, was recorded and analyzed. In the ultra-late phase, 95% of ARMD eyes with choroidal neovascularization (CNV) showed geographic hypofluorescent lesions. These hypofluorescent lesions occurred in 73% of ARMD eyes without CN, while age-matched normal subjects had no hypofluorescent lesions. The mean intensity of fluorescence in the normal elder subject group was significantly higher than that seen in the normal younger subject group. These findings may reflect aging change and bio-distribution of lipid on the Bruch-RPE complex. 3. The early dye filling pattern of the choroid: We performed ICG angiography on 10 healthy young volunteers aged 22 to 26 years (23.4+/-1.3; mean+/-standard deviation) using an improved ICG video camera system. ICG (50 mg) dissolved in 2 ml in distilled water was injected through the antecubital vein. Although the choroidal dye filling varied among subjects, it always began in the macular area. In the 10 subjects, initial dye filling had two patterns: reticular (n=8) and flush (n=2). The choroidal circulation filled completely before the retinal circulation did. Bright fluorescence in the macula and fast blood flow may be correlated with ample blood volume and abundant blood vessels in the macular area. 4. The spreading pattern of ICG fluorescence in the choroid: The ICG images obtained for observing the early dye filling pattern of the choroid were further processed with a computer-assisted image analyzer. Subtracted images were made using the early ICG frames with a time interval of 0.12 second. Ninety frames of time-sequential images for 3 seconds starting from the initial dye appearance in the choroid were prepared to construct an animated image. In the initial phase of eye filling, patchy fluorescence appeared in the fovea. The fluorescence then spread centrifugally in all directions in a wave-like pulsatile manner towards the peripheral fundus with increasing brightness. Thus an animated video of subtracted images allowed us to evaluate flow dynamics at the level of the choriocapillaris. Using this new approach, pathogenic involvement of choroidal circulation in varied chorioretinal diseases such as central serous chorioretinopathy can be studied with precision. II. Retinal circulation. We developed a new device to study the oxygen saturation (OS) levels in a wider fundus area. We call this device a spectral retinal image (SRI) system. We are pursuing the possibility of this instrument being in clinics to evaluate chorioretinal diseases. 1. Introduction of the device: The instrument consists of a Sagnac interferometer that has been mounted on top of a fundus camera, and a software module which consists of an acquisition module and an analysis module. The image acquisition process takes 6 seconds during which the fundus is illuminated by the white incandescent light of the fundus camera at the regular power settings. OS values in each pixel of the fundus image with a 35-degree view can then be estimated from the respective spectrum that is obtained by Fourier transforming the interferometer signals. Each OS value is represented by a specific color, and each pixel of the fundus image is painted accordingly. 2. Clinical applications. 1) Control study: OS levels of both retinal arteries and veins within a 1 disc diameter (DD) area around the disc were measured from 20 SRIs obtained from 10 healthy volunteers, and were 96.65+/-3.30% and 56.05+/ 4.69%, respectively. Then 30 healthy volunteers were recruited for further study in which the OS values were calculated in five retinal regions: (1) juxta papillary area within 1.75 DD, (2) fovea within 1.0 DD, (3) papillomacular region within 1.0 DD, (4) superior area of the posterior fundus within 1.0 DD, and (5) inferior posterior area of the posterior fundus within 1.0 DD. The OS level of the juxta-papillary area was the highest, while that of the fovea was the lowest and the other three posterior retinal regions were in the middle. Thus OS levels differed at various areas in the retina with statistical significance. In spite of abundant choroidal circulation in the fovea, the overlying retina may have a relative by low oxygen level. As the retinal pigment epithelium may be efficient enough to block the effect of the choroid optically, our results may indicate that the OS levels represent the OS of the retina. 2) Measurement of OS levels in eyes with retinal circulatory disturbances: Eleven eyes of 10 patients with central retinal vein occlusion (CRVO), which showed various degrees of severity, and 4 fellow unaffected eyes of selected patients were examined by both fluorescein angiography (FAG) and the new SRI system. The fluroescein angiograms were correlated with OS maps that were calculated from the SRI. OS grading demonstrated by OS maps correlated well with severity of CRVO, as estimated by FAG. Thus our SRI system is noninvasive with reproducible results, and may prove to be a useful clinical tool to evaluate the degree of retinal ischemia. 3) Measurement of OS levels in eyes with glaucoma: Forty-seven eyes with open angle glaucoma (OAG) of 47 patients and 21 eyes of 21 age-matched normal subjects were recuited for the study. Twelve eyes with low-tension glaucoma (LTG) were included in the OAG eyes and the rest of the OAG eyes had primary OAG. All patients and normal subjects were examined by SRI. Visual field tests for OAG eyes were done with a Humphrey Field Analyzer using the 30-2 program, Swedish interactive threshold argorithm (SITA). OS levels in the retina at 5 different points: superior, inferior, superio- and inferio-temporal, and nasal region within a juxta-papillary area of 200 microm in diameter were calculated from the SRI. OS levels of retinal arteries were also measured and there were no significant differences between OAG and the control group. OAG eyes showed reduced OS levels in the inferio-temporal retina with statistical significance. This observation was more prominent in LTG eyes. The reduced OS levels observed in OAG eyes correlated well with mean deviation (MD) and the sum of total deviation of the 17 points in corresponding areas in the visual field analysis. PMID- 15656090 TI - [DNA diagnosis in the age of individual made-to-order medications]. AB - Human diseases develop by complex mutual relationships of genetic and environmental factors. In inherited diseases, DNA diagnosis of the disease causing genes provides a confirmation of the disease. On the other hand, DNA diagnosis of the disease-sensitive genes in multifactorial diseases, such as the lifestyle-related diseases (common diseases), provides the risk of developing the disease. Two new technologies are being used for DNA diagnosis in the clinic. The first is called Invader Technology and is a non-PCR method and is useful for detecting well-known genetic variations in large samples efficiently. We have developed a method to quantify the heteroplasmy of mitochondrial DNA mutations by this technique. The second technique, called WAVE, uses denaturing high performance liquid chromatography to screen for mutations in a large number of samples automatically and efficiently. Clinical DNA diagnoses are divided into those for single genetic diseases and those for multifactorial diseases. The purposes for DNA diagnosis in single genetic diseases are: 1) to propose a new clinical classification of the disease, such as TGFB1-related corneal dystrophy or retinitis pigmentosa, based on the genotypes; 2) to confirm a clinical diagnosis, such as Leber's hereditary optic neuropathy (LHON); and 3) to provide an early diagnosis before the development of the disease and thus provide an opportunity to start early treatment. For example, a family history of glaucoma is one of the risk factors for developing glaucoma. The frequency of mutations in the glaucoma genes, myocilin and optineurin, were found to be about 3% and 0.25%, respectively, in Japanese. The significance of DNA diagnosis in multifactorial diseases is that it provides a risk diagnosis for an individual. Single nucleotide polymorphisms (SNPs) of disease-sensitive genes are associated with only a 2- to 3-fold risk of developing the disease. A case-control association study was performed using many SNP markers to identify glaucoma-sensitive genes. A total of 671 Japanese individuals, 201 POAG patients, 234 NTG patients, and 236 normal controls were examined. Fifty-two SNPs in the 38 genes were examined to identify the glaucoma-sensitive genes as candidate genes, and SNPs in AT 1, AT 2, PON 1, GSTT 1, NOS 3, and EDN 1 were associated with glaucoma statistically. Mitochondrial (mt) DNA mutations associated with LHON might be risk factors for open-angle glaucoma, because abnormal optic disc excavations are also found in LHON patients. A total of 651 blood samples were screened for 6 LHON-associated mutations with the Invader assay. Seven patients had one of the five mutations, but none had developed LHON. The 5 mutations were not identified in 236 normal controls. MtDNA mutations may make the optic disc more susceptible to damage in glaucoma patients. The clinical variability in LHON patients suggests that the disease most likely results from multi factorial mechanisms. To determine whether genetic polymorphisms for oxidative stress and apoptosis cause clinical variability in patients with LHON, 12 polymorphisms in 10 genes were analyzed in 87 patients with the 11778 mutation in relation to the age at onset and final visual acuity. LHON patients carrying homozygous His 113 in the EPHX1 gene or homozygous Arg 72 in the TP53 gene developed the disease earlier than those without this genotype. Thus, nuclear genetic polymorphisms related to oxidative stress or apoptosis may modify the age of LHON onset. A clinical trial of 38 healthy volunteers without systemic diseases or eye diseases was performed using an angiotensin II receptor blocker (candesartan cilexetil) as an alternative drug for lowering intraocular pressure (IOP). After a single oral dose of candesartan cilexetil, the IOP fell significantly for 24 hr. There was no association between the effects of oral candesartan cilexetil and the three SNPs in the AT 1 gene. In the 21th century, DNA diagnosis for multifactorial diseases will be required to determine the treatment plan for individuals or to prevent diseases. We have developed a panel of tests by Invader assay for clinical use to detect mutations in the myocilin gene or in LHON. In the future, we will develop a panel to detect SNPs in the glaucoma-sensitive genes to diagnose individuals at risk for developing glaucoma. Such information is expected to help develop new medications. PMID- 15656091 TI - [Structure of parasitic arthropod communities in forest small mammals]. AB - Species composition and structure of ectoparasite arthropod communities were examined all year round six years in the bank vole Clethrionomys glareolus, Ural wood mouse Apodemus uralensis and the common shrew Sorex araneus in forests of the Ilmen'-Volkhov depression. In total, 4500 host samples have been examined and all ectoparasites have been collected. The species composition of ectoparasite community in small mammal species are as follows: the bank vole--29 insect, tick and mite species, the common shrew--23 species, the Ural wood mouse--16 species. In forest biotopes, many temporary ectoparasitic species occur on several host species living in the same habitats under a forest canopy and contacting each other. A parasitic supracommunity in the ecosystem examined has a pool of temporary ectoparasites, which is available for all the community of small mammals. A role of different rodent and shrew species are hosts of insects and ticks changes depending on a density of potential host populations and numerous other environment factors. PMID- 15656092 TI - [Methods for estimating the demographic structure of the taiga tick (Ixodidae) based on results of standard parasitological observations]. AB - A retrospective estimation of the abundance dynamics of the taiga tick Ixodes persulcatus in mixed coniferous and leaf-bearing forests of the Udmurtia Republic in the period 1957-1986 was carried out. A possibility to estimate the absolute number of all stages of I. persulcatus based on relative indices. Females of I. persulcatus lay 20,250 eggs per 1 hectare, and this number of eggs gives birth to 15,000 larvae. From this number, 7870 larvae hibernate and 6550 individuals became fed. The number of nymphs is 5930, among which 5110 individuals live up to spring, and 1390 became fed. The number of adult mites in autumn is 1250; in subsequent spring this number decreases to 780. The mean number of engorged females is 8. The mortality rate of ticks caused by the deficit of hosts increases from preimaginal stages to adults; for larvae, nymphs and imago this index is 16.6, 72.8, and 97.9%, respectively. Quotas of individuals with 3-, 4- and 5-year life cycle among the unfed imago are 70.4, 28.0, and 1.6%, respectively. PMID- 15656093 TI - [Seasonal dynamics of a flea number (Siphonaptera) on the common shrew (Sorex araneus) in the north part of the Novgorod oblast]. AB - In the course of 5-year stationary investigations of the common shrew Sorex araneus in the north part of the Novgorod oblast, 12 flea species have been recorded on this host. Among them, Palaeopsylla soricis and Doratopsilla dasycnema are specific parasites of small insectivores, including the common shrew. Hystrichopsylla talpae is a polyxenous species, parasitizing both rodents and insectivores. Other 9 flea species are not common parasites of S. araneus and apparently have come to this host from other mammal species inhabiting the forest biotopes in the area of investigation. P. soricis and D. dasycnema have similar phenology of parasitism. These fleas appear in spring (April), are present during summer and autumn and disappear in winter. During a year, the abundance of these species shows three pikes, which correspond to three generations: spring (April), summer (June-July) and autumn (September-October) generation. The first species dominates in spring (April-May) and in the autumn-winter period, while the second species predominates in summer. These two species comprise over 90% of total number of fleas collected and determine general dynamics of the flea number on the shrews; during the year the flea number has a sinusoidal increasing from spring to autumn and minimum in winter. PMID- 15656094 TI - [Host-parasite relationships of the genus Hyalomma Koch, 1844 (Acari, Ixodidae) and their connection with microevolutionary process]. AB - Host-parasite relationships of Hyalomma species of the world fauna are analyzed. The majority of species infests predominately various mammals. Birds and reptiles are used as preferred hosts by several Hyalomma species, and only on certain stage: adults of H. aegyptium parasitize tortoises; immature stages of H. marginatum parasitize birds. It is hypothesized that relationships of H. aegyptium adults (subgenus Hyalomma s. str.) with reptiles are secondarily in origin. Immature stages of H. aegyptium retain the primary wide diapason of hosts, which are various small mammals, birds and reptiles. The life cycle of this species is the three-host type that is considered as a primary type in ixodid ticks. A typical scheme of relationships with their hosts in all well examined Hyalommina species has following features: the adult stage parasitize large and medium sized mammals, immature stages parasitize small mammals, three host life cycle. A variety of preferred hosts and types of life cycle is observed in the subgenus Euhyalomma. All species of this subgenus can be arranged into two groups. In the first group, the immature stages infest only small mammals and birds, and the adults parasitize large mammals; this type of host preferences is probably primary host-parasite relationships of Hyalomma. This group includes: H. albiparmatum, H. asiaticum, H. excavatum, H. franchinii, H. impeltatum, H. impressum, H. lusitanicum, H. marginatum, H. nitidum, H. schulzei, and H. truncatum. Hyalomma marginatum and H. schulzei are two-host species; H. excavatum is two- or three-host tick. All the remaining species (except H. albiparmatum, which life cycle is unknown) are three-host ticks. In the second group, the immature stages as well as the adult stage parasitize large mammals. This group includes: H. dromedarii, H. anatolicum, and H. scupense. These species are two- or one-host ticks. PMID- 15656095 TI - [Evolutionary complication of life cycles in Coccidea (Sporozoa: Coccidea)]. AB - Similar strategies to preserve a species were evolved independently in different groups of Coccidea. Polyenergid oocysts and tissue cysts are found in representatives of the orders Protococcidiida and Eimeriida. Hypnozoits are found in Karyolysus lacerate and Plasmodium vivax; transovarial transmission of parasites occurs in life cycles of Coccidea of the genera Karyolysis and Babesia. Formation of heteroxenity in groups of Coccidea apparently was developed by different ways and in different periods. In some groups (Cystoisospora, Toxoplasma, Aggregata, Atoxoplasma, Schelackia, Lankesterella, and Calyptospora), recent definitive hosts were initial hosts; in other groups (Sarcocystis, Karyolysus, Haemogregarina, Hepatozoon, Plasmodium, Haemoproteus, Leucytozoon, Babesiosoma, Theileria, and Babesia), intermediate hosts were initial hosts. PMID- 15656096 TI - [Symmetry of orthonectids and dicyemids]. AB - The analysis of the general structure of orthonectids shows that free-living sexual specimens are bilaterial, while the parasitic plasmodium is anaxonic. All worm-like stages in the life cycle of dicyemids are characterized by monaxonic heteropolar symmetry, while their infusorioform is bilaterial. PMID- 15656097 TI - [Mathevolepis junlanae sp. n. (Cyclophyllidea: Hymenolepididae: Ditestolepidini), a new cestode species from shrews of Far East]. AB - Mathevolepis junlanae sp. n. from shrews (Sorex spp.) of Far East is described given. The new species occupies an intermediate position between the Palaearctic species M. petrotschenkoi Spassky, 1948 (total number of proglottids--4) and M. skrjabini (Sadovskaya, 1965) (10-12 segments in a series) by the number of synchronously developing proglottids (2-4). The new species differs from M. larbi Karpenko, 1982 by the structure and measurements of copulative apparatus in proglottids. In M. larbi, vagina S-shaped, vagina length 0.147-0.154 mm, cirrus length 0.146-0.149 mm, and cirrus bursa reaching aporal excretory vessels. M. junlanae sp. n. has cirrus bursa slightly crossing center line of sexually mature proglottids, cirrus about half as long (0.084-0.092 mm), and smaller sac-shaped vagina (0.075-0.10 mm). The new species differs from the Nearctic representative M. macyi (Locker and Rausch, 1952) by lesser measurements of cirrus bursa (0.12 0.14 mm versus 0.4 mm) and a fewer number of proglottids in series (2-4 versus 5 7). PMID- 15656099 TI - [A first record of microsporidia in the ixodid tick Ixodes ricinus L. (Ixodidae) in the territory of the Commonwealth of Independent States, Republic Moldova]. AB - Spores of microsporidia have been recovered in 5 specimens of 13 ixodid ticks Ixodes ricinus from various populations of the Republic Moldova collected in spring of 2004. Microsporidia were detected by means of fluorescent microscopy. Intensity of infection was 3-6 spores per a micropreparate from one mite. Based on spore size, character of staining and the presence of diplocarion, these spores are referred to the Nosema-like type. Low intensity of infection probably is caused by that fact that ticks were collected in spring period and were unfed. PMID- 15656101 TI - [A redescription of Gyrodactylus atherinae Bychowsky, 1933 based on the collection of B. E. Bychowsky of 1947 from Atherina boyeri pontica in the Black Sea]. AB - A redescription and comparative diagnosis of Gyrodactylus atherinae Bychowsky, 1933 is made based on the material collected of B. E. Bychowsky from Atherina boyeri pontica caught near Karadag (Black Sea) in summer of 1947. PMID- 15656098 TI - [On a species composition of the genus Copiatestes Crowcroft, 1948 (Digenea: Syncoeliidae)]. AB - The genus Corpiatestes Crowcroft, 1948 is revised. It is shown that Corpiatestes filiferus (Leuckart, in Sars, 1885), Syncoelium cypseluri Yamaguti, 1970 and S. regulaci Villarreal and Dailey, 1993 are junior synonyms of the type species C. thyrsitae Crowcroft, 1948. The genus Copiatestes is monotypical. PMID- 15656100 TI - [The influence of the parasite invasion on antioxidant enzyme activity in the liver and muscles of a host, the Black Sea flounder Psetta maxima maeotica]. AB - The influence of the intestine cestode Botriocephalus gregarius on the antioxidant enzyme system in the liver and muscle of its host, the Black Sea flounder Psetta maxima maeotica, was studied. The significant increase of superoxidedismutase activity in examined tissues and changes of catalase and peroxidase activities were detected. High correlation (r > or = 0.6) between enzyme activities and the parasite number was noted. The possibility to use antioxidant enzyme activity as biomarkers of the parasite invasion in fish is discussed. PMID- 15656102 TI - [A new morphological type of nematode larvae of the family Anisakidae (Ascaridida)]. AB - Larvae of anisakid nematodes of a new morphological type from the anadromous mikizha Parasalmo mykiss (Salmonidae) are described. The larvae have 6 sclerotized cogs on the cephalic end of the body. These larvae are referred to the tribe Lappetascarini Rasheed, 1965 as forms with uncertain generic membership. PMID- 15656103 TI - Optimisation in stereotactic radiosurgery of AVMs: I. Mathematical considerations. AB - One forward-planning method and five inverse-planning methods for optimisation of treatment in radiation therapy were compared in the particular case of radiosurgery with micro-multi-leaves collimator (MMLC) and arc therapy. The "manual" method, two matrix methods (singular value decomposition and non negative least square fit), two gradient methods (quasi-Newton and conjugate gradient algorithms) and the "simulated annealing" stochastic method were investigated. The performance of these methods was assessed in terms of the speed of convergence to an optimum, the ability to account for the organs at risk, and probability of targeted success. The study employed an adapted version of the GRATIS treatment planning system. A group of 22 patients previously treated by arc therapy for arteriovenous malformations (AVMs) were studied to evaluate the performance of the various optimisation methods for MMLC and arc therapy. The conjugate gradient method proved to be the most appropriate for most cases. PMID- 15656104 TI - Optimisation in stereotactic radiosurgery of AVMs: II. Comparison of arc and MMLC therapy. AB - Two stereotactic surgery methods, arc and micro-multileave collimator (MMLC) therapy, were compared in the particular case of arteriovenous malformations (AVMs) treatment. Different methods of the treatment optimisation were used. The comparison covered a group of 22 patients suffering from peripheral and central AVMs of different sizes who underwent initially arc therapy. Several parameters were evaluated to compare the two methods: 2D and 3D isodose representations, dose-volume histograms (DVHs) and probability of success. The 3D isodoses were compared for the 22 patients showing a better conformity for the MMLC (three cases are presented). The DVHs of the AVM were also in favour of MMLC. In terms of probability of success, the results showed that are therapy was superior only in the case of small spherical lesions. MMLC therapy proved to be superior to arc therapy in all cases but central spherical small volume AVMs. PMID- 15656105 TI - [Calculation of proton spectra of pure brain tissue types by multiple voxel measurements and image segmentation]. AB - In localized in vivo proton NMR spectroscopy (1H MRS) of the human brain, it often cannot be avoided that the selected volume of interest (voxel) includes both gray matter (GM) and white matter (WM). Since the spectra of GM and WM differ, in general, the acquired spectrum represents a mixed spectrum that depends on the tissue composition inside the voxel. This study describes a method that enables the determination of pure GM spectra and pure WM spectra from mixed spectra. The pure tissue spectra are calculated from measured spectra acquired from several voxels with different mixed tissue compositions. For this purpose, the tissue composition in the voxels must be known. It is determined by segmentation of an additionally acquired 3D image data set with higher spatial resolution. In volunteer examinations, measurements were performed in different regions of the cerebrum, cerebellum, and thalamus. In all examined brain regions, particularly in the cerebellum, clear differences were found between the spectra of WM and GM. The detected differences in the spectra of WM and GM indicate that the tissue composition in the voxel has to be considered in patient studies, in order to distinguish pathological alterations in the spectra from the effects of tissue composition. PMID- 15656106 TI - Pre-treatment verification of intensity modulated photon beams with films and electronic portal imaging--two years of clinical experience. AB - Aim of this report was to summarise clinical experience in the field of pre treatment dosimetric verification of intensity-modulated photon beams (IRMT). From May 2001 to July 2003, 50 patients were irradiated according to IMRT techniques with 6 MV photon beams produced by a Varian Clinac equipped with a 80 leaves multileaf collimator. Dose plans were computed using commercial treatment planning systems, Nucletron Helax-TMS for static cases and Varian Eclipse-Helios for dynamic cases. Pre-treatment dosimetric verification was carried out on a field-per-field basis measuring 2D absolute dose distributions in solid water at 10 cm depth using films or an electronic portal imaging device (EPID). Verification measurements were compared with expected dose maps, and differences were evaluated by means of both a point-to-point analysis and the Gamma Index. Irradiated target volumes (30 head and neck, 8 breast, 12 other patients) ranged from 111 to 2121 cm3 with a mean of 652 +/- 378 cm3. Twenty-nine dose plans were delivered with dynamic technique and 44 with static technique. On average, 5.9 +/ 1.3 fields were applied per plan, with 12.1 +/- 1.6 segments per field in the static mode. Averaging over the whole number of fields we obtained a mean difference (on a pixel-by-pixel basis and per 100 MU delivered) of -0.22 +/- 0.64 cGy between calculation and measurement, with a standard deviation of 1.93 +/- 0.65 cGy. The mean value for the Gamma Index evaluation was 0.47 +/- 0.10, with a mean standard deviation of 0.35 +/- 0.17. The fraction of pixels lying inside the field and showing a gamma index larger than 1 was 5.7% for the triplet Eclipse film-dynamic delivery and 9.9% for the triplet Helax-TMS-EPID-static delivery. The employed IMRT treatments proved that this technique is feasible and dosimetrically accurate. Treatment verification stability and dosimetric analysis of treated plans are highly satisfactory and allow the safe introduction of this modality in the spectrum of techniques offered to a large class of patients. PMID- 15656107 TI - [Measurement of MRI scanner noise]. AB - The present paper describes a simple method for the analysis of MRI scanner noise. Besides the heating of body tissue by strong RF radiation and the formation of circular currents in the body induced bey switching field gradients, a noise level of more than 100 dB(A) during the measurement belongs to the potential risks of MRI. This risk is of particular concern for staff and accompanying persons who remain close to the scanner for different reasons (e. g., monitoring of anesthetized patients, reassuring of children). For this reason, and given the scanty information on noise provided in the manuals of the scanners, it is useful to quantify the noise level more exactly. This applies also to the evaluation of different sound-reducing methods for the patient. This presents the results of noise level measurements in the tomograph and in its surrounding, with and without noise reduction by headphones. PMID- 15656108 TI - Correction of ionic recombination for pulsed radiation according to DIN 6800-2 and TRS-398. AB - The international Agency of Atomic Energy (IAEA) has recently published a new code of practice (TRS-398) for dosimetry of ionizing radiation. In this code, the saturation correction for pulsed radiation is treated differently compared to the German regulation DIN6800-2. The Code provides also an approximation formula that is not identical to that of DIN 6800-2. The present study analyzed the relations between the different formulas and determined the impact on the size of the resulting saturation correction factor. All formulas can be traced back to Boag's theory of ionic recombination, however the formula in DIN 6800-2 uses an additional linear approximation. The approximation in TRS-398, in turn, can be derived from the formula in DIN 6800-2 by applying a further linear approximation. Measurements were performed with four different ionization chambers using photon and electron beams. All formulas yielded the same saturation correction within 0.1%. However, for the medical physicist in a hospital it would be desirable to have the same formula in national regulations and international recommendations, to avoid any uncertainty on which method to apply. PMID- 15656109 TI - Radiation hazard during a manned mission to Mars. AB - The radiation hazard of interplanetary flights is currently one of the major obstacles to manned missions to Mars. Highly energetic, heavy-charged particles from galactic cosmic radiation can not be sufficiently shielded in space vehicles. The long-term radiation effects to humans of these particles are largely unknown. In addition, unpredictable storms of solar particles may expose the crew to doses that lead to acute radiation effects. A manned flight to Mars currently seems to be a high-risk adventure. This article provides an overview on the radiation sources and risks for a crew on a manned flight to Mars, as currently estimated by scientists of the US National Administration for Space and Aeronautics (NASA) and the Space Studies Board (SSB) of the US National Research Council. PMID- 15656110 TI - [Worthiness of Sir Godfrey N. Hounsfield]. PMID- 15656112 TI - [Rabbit.page.nl]. PMID- 15656113 TI - [Corneal diseases in horses (part 2)]. PMID- 15656111 TI - [A horse with a badly-healing wound? A case report]. AB - A 14-year-old Haflinger mare was presented with a wound on the right metatarsus which it had sustained 3 years earlier. The wound had never completely healed but had only recently become a problem. Over a period of several months, the wound became larger, produced a lot of exudate, and the horse became lame on the affected limb. Clinical examination and radiographs failed to reveal the cause of the deterioration. Histological evaluation of tissue removed during debridement of the wound revealed squamous cell carcinoma. Because the tumour had already invaded the bone, the prognosis was unfavourable and the horse was euthanised. Necropsy showed the tumour to have metastasised to the right inguinal area and the adjacent mammary gland. PMID- 15656114 TI - [Diagnostic possibilities for pathological lesions in poultry to trace disease outbreaks of classical fowl pest in the Netherlands in 2003]. PMID- 15656115 TI - [Yearly revaccination of domestic animals]. PMID- 15656116 TI - [Position PAO-D in current market]. PMID- 15656118 TI - The organ trail: matching supply, demand. PMID- 15656117 TI - [A new year--a new future without VKO]. PMID- 15656119 TI - Pfizer touts Medicaid data in arguing that DM works. PMID- 15656120 TI - A better case for quality. Share the savings! PMID- 15656121 TI - When your employees are your focus group. PMID- 15656122 TI - The weight debate, continued. PMID- 15656123 TI - Executive compensation catches up. PMID- 15656124 TI - The role of managed care in patient safety & error reduction. PMID- 15656125 TI - Artificial heart buys precious time. PMID- 15656126 TI - Companies seek guidance as payment options increase. PMID- 15656127 TI - Spitzer just getting started on health care. PMID- 15656128 TI - [Identification of Pterocephalus hookeri]. AB - OBJECTIVE: To study the identification method of Pterocephalus hookeri. METHOD: The microscopical, Physicochemical and TLC methods were used. RESULT AND CONCLUSION: The convenient and effective identification methods for P. hookeri were established, which provide basis for its quality standard and development. PMID- 15656129 TI - [Determination of carvacrol and thymol in Mosla chinensis by HPLC]. AB - OBJECTIVE: To establish a quantitative method of determination of carvacrol and thymol in Mosla chinensis. METHOD: The sample was extracted with 95% ethanol, ODS column was used with methanol-water-acetic acid (60:40:2) as mobile phase. The detection wavelength was set at 274 nm. RESULT: The linearities of carvacrol and thymol were respectively in the range of 0.23-2.15 microg (r = 0.9999) and 0.39 2.36 microg (r = 0.9999); the average recoveries were 99.9% (RSD 1.4%) and 98.6% (RSD 1.3%); the RSD of repeatability were 1.1% and 1.6%. CONCLUSION: The method is reliable, and can be used for quality control of M. chinensis. PMID- 15656130 TI - [Genetic analysis of cultivated ginseng population with the assistance of RAPD technology]. AB - OBJECTIVE: To provide reference for breeding of cultivated ginseng by researching the genetic diversities among different strains. METHOD: To make up the systematic diagram of genetic relationship by calculating the proximity coefficient with SPSS 10.0 software and clustered by between-groups linkage method. RESULT: The selected 18 random primers used in PCR amplification to produce 145 bands in 17 samples, among which 53 bands (36.5%) are polymorphic. The results of cluster analysis show that the genetic relationship between different strains of Da-maya is closer than that of Er-maya, which proves that it is feasible to differentiate the cultivated groups in molecular level on outside character. CONCLUSION: RAPD is a favorable molecular marker to assist ginseng breeding. PMID- 15656131 TI - [Study on quality evaluation methods of pretreatment for polystyrene-type macroporous absorbing resins]. AB - OBJECTIVE: To establish a GC method using wide bore open tubular columns f or controlling pretreatment for polystyrene-type macroporous absorbing resins and its eligible standard. METHOD: A model macroporous absorbing resins made in our lab was eluted by anhydrous alcohol. The residual solvents in elution (xylene, styrene, diethylbenzene, divinybenzene, decane) were assayed by GC. When the residual solvents can not be detected, the pretreatment is eligible. Compare this method with the methods referring in the literatures. RESULT: When using this method to control the pretreatment, the resins need to be elute 11 times with anhydrous alcocol. When using "adding several times water in the alcohol elution will not be turbid" to control the pretreatment, the resins need to be elute 3 times with anhydrous alcocol. When using "the elution have no UV absorption between the wavelength 200-400 nm" to control the pretreatment, the resins need to be elute over 20 times with anhydrous alcocol. CONCLUSION: The method is simple and feasible. PMID- 15656132 TI - [Simultaneous determination of 4 anthraquinones in Kuhuang injection by RP-HPLC]. AB - OBJECTIVE: To develop a new high-performance liquid chromatographic method for simultaneous quantitative determination of 4 anthraquinones in Kuhuang injection. METHOD: The chromatographic separation was performed on a Lichrospher C18 column (4.6 mm x 250 mm, 5 microm,), A linear gradient elution of A (CH3COOH: H2O = 1:100) and B (CH3COOH: H2O: CH3CN = 1:20:80) was used. The flow rate was 0.8 mL x min(-1) and column temperature was set at 35 degrees C. The UV detection wavelength was set at 254 nm. RESULT: The recoveries of rhein, emodin, chrysophanol and physcion were 98.9%, 100.5%, 102.5% and 99.0%, respectively. Their linear ranges were 0.0875-1.75 microg for rhein, 0.0825-1.65 microg for emodin, 0.159-3.17 microg for chrysophanol and 0.0525-1.05 microg for physcion, respectively. CONCLUSION: The method was accurate, repeatable and suitable to determine the contents of 4 anthraquinones in Kuhuang injection. PMID- 15656133 TI - [Study on HPLC fingerprint of Xuelian injection]. AB - OBJECTIVE: To establish the HPLC fingerprint of Xuelian injection. METHOD: HPLC with Diamonsil C18 (4.6 mm x 250 mm, 5 microm) column was used, the methanol-1% acetic acid as a mobile phase and detection wavelength at 270 nm. RESULT: Indicating 11 peaks on HPLC fingerprint. CONCLUSION: This method is simple and acurate with a good reproducibility and can be used as a quality control method for Xuelian injection. PMID- 15656134 TI - [Studies on HPLC fingerprints of tanshinone microemulsion]. AB - OBJECTIVE: To establish the HPLC fingerprints of tanshinone microemulsion. METHOD: HPLC analysis was carried on Hypersil C18(4.6 mm x 250 mm, 5 microm) analytical column; the acetonitrile and 0.5% phosphoric acid solution were used as mobile phases with gradient elution at a flow rate of 1.0 mL x min(-1). The UV detection wavelength was set at 270 nm. RESULT: Six peaks on HPLC fingerprint of Tanshinone Microemulsion are indexed. CONCLUSION: The method developed in the present study is convenient, reliable, and can be used as a quality control method for Tanshinone Microemulsion. PMID- 15656135 TI - [Determination of orientin and vitexin in Trollius chinesis preparation by HPLC]. AB - OBJECTIVE: To establish a HPLC method for determining orientin and vitexin in Trollius chinesis preparation. METHOD: The separation was carried out on Hypersil ODS column with a mixture of acetonitrile-1% acetic acid solution (15:85) as the mobile phase, at a flow rate of 1.0 mL x min(-1). The detection wavelength was set 340 nm. RESULT: Orientin showed a good linear relationship in the range of 8.42-84.2 microg x mL(-1) (r = 0.9999) and the average recovery is 98.3%, RSD is 0.8%. Vitexin showed a good linear relationship in the range of 8.07-80.7 microg x mL(-1) (r = 0.9999) and the average recovery is 102.4%. RSD is 1.7%. CONCLUSION: The method is sensitive, specific, and accurate with a good repeatability and can be used for the quality control of Trollius chinesis preparation. PMID- 15656136 TI - [Studies on the chemical constituents from the aerial parts of Breynia fruticosa]. AB - OBJECTIVE: To investigate the chemical constituents from the aerial parts of Breynia fruticosa. METHOD: Various chromatographic techniques were employed for isolation and purification of the constituents. The structures were elucidated by chemical evidence and spectral methods. RESULT: Seven compounds were obtained and identified by spectroscopic methods and compared with authentic samples as aviculin [(+)-isolariciresinol-9'-rhamno-pyranoside], friedelan-3beta-ol, friedelin, arborinone, isoarborinol, 5-hydroxy-7,8,4'-trimethoxy flavone, 2,4 dihydroxy-6-methoxy-3-methyl-acetophenone. CONCLUSION: All compounds were firstly isolated from B. genus, furthermore, aviculin was isolated from Euphorbiaceae for the first time. PMID- 15656137 TI - [Study on the chemical constituents from the herb of Gentianopsis paludosa]. AB - OBJECTIVE: To study the chemical constituents from the herb of Gentianopsis paludosa. METHOD: Column chromatogrophy and spectral analysis were used to isolate and identify the constituents. RESULT: Six compounds were isolated and identified as 1,7-dihydroxy-3,8-dimethoxyxanthone (I), 1-hydroxy-3, 7, 8 trimethoxyxanthone (II), 1, 8-dihydroxy-3, 7-dimethoxyxanthone (III), 1-hydroxy 3, 7-dimethoxyxanthone (IV), beta-sitosterol (V), daucosterol (VI). CONCLUSION: Compounds III-VI were isolated from the plant for the first time. PMID- 15656138 TI - [Changes in level of organic acids in fructus crataegi after processing]. AB - OBJECTIVE: To compare and analyze the organic acids level in fruit of Crataegus pinnatifida var. major before and after processing. METHOD: The amount of total organic acids and individual citric acid, malate acid, ursolic acid and oleanolic acid in fruit of C. pinnatifida var. major before and after processing were determined by pH titration and HPLC analysis. RESULT: Good linear relationship between peakarea and amount was noted for 0.28-9.02 microg of malate acid, with a correlation coefficient of 0.9998. Regression equation was Y = 2344.7 + 49309.6 X. Good linear relationships between the logarithm of peak area and amount were noted for 0.348-1.74 microg of usolic acid and 0.498-2.48 microg of oleanolic acid, with a correlation coefficient of 0.9998 in each case. Regression equations were Y = 1.3628 X + 5.9508 and Y = 1.4447 X + 5.8236, respectively. CONCLUSION: The lipid-soluble organic acids mostly remain whereas only about 70% of the water soluble acids remain after processing in fruit of C. pinnatifida var. major. PMID- 15656139 TI - [Effects of tianma gouteng fang on transmitter amino acids in the hippocampus extracellular liquids in freely moving rats subjected to brain ischemia]. AB - OBJECTIVE: To observe the effects of Tianma Gouteng Fang (TGF) on the transmitter amino acids in the hippocampus extracellular liquids in freely moving rats subjected to incomplete brain ischemia. METHOD: Hippocampus extracellular liquids was collected continuously by the microdialysis sampling technology in freely moving rats during pre-ischemia, incomplete ischemia and reperfusion periods induced by the occlusion and loose of both common carotid arteries. Each dialysate sample was assayed for GABA, Tau, Glu, Cys and Arg with HPLC electrochemical detector. RESULT: TGF increased the concentrations of GABA and Tau in the extracellular liquids of rat hippocampus. Compared with the model group, the concentration of Glu in the middle and large dosage groups of TGF, during the 120 min of ischemia, reduced by 38.64% and 31.35%, Tau increased by 13.99% and 12.86%, GABA advanced 25.89% and 33.99%, Cys decreased by 40.93% and 42.08%, Arg raised to 116.95% and 108.96%, respectively. After 120 min of reperfusion, the concentration of Glu decreased by 14.55% and 11.48%, Tau increased by 16.13% and 14.03%, GABA increased by 24.41% and 26.22%, respectively. CONCLUSION: TGF can increase the concentration of inhibitory amino acids in hippocampus extracellular liquids of rats and inhibit the excessive release of excitatory amino acids and raise the concentration of the inhibitory amino acids and Arg during the ischemia-reperfusion periods. Therefore, TGF can play the neuroprotective role. PMID- 15656140 TI - [Protective effect of jasminoidin on cascade of damage of cerebral ischemia in rats]. AB - OBJECTIVE: To study the protective effect of jasminoidin on cascade of damage of cerebral ischemia. METHOD: The rats were randomly divided into four groups: sham operated group, ischemic group, the jasminoidin-treatment group and PNS-treatment group. Focal cerebral ischemia was produced by permanent occlusion of left middle cerebral artery (PMCAO) in rats. Radioimmunoassay (RIA) was utilized to identify the content of tumor necrosis factor-alpha (TNF-alpha) and interlukin 1beta (IL 1beta) in brain tissue of rats following ischemia. and that of Neuron-specific enolase (NSE) in rat's serum was observed too. The plasma concentration of vonWillebrand factor (vWF) was identified by enzyme-linked immunoadsordent assay (ELISA). RESULT: After 12 h and 24 h of ischemia, the contents of TNF-alpha and IL-1beta and vWF as well as on NSE showed concomitant increase. Jasminoidin dramatically inhibited the increase of TNF-alpha and IL-1beta after 12 h and 24 h of ischemia, and repressed the increase of vWF after 12 h and 24 h of ischemia too. However, the influence of jasminodin NSE after 12 h and 24 h of ischemia was not significant. CONCLUSION: Jasminoidin had good effect on repressing the expression of TNF-alpha and IL-1beta as well as vWF caused by cerebral ischemia, thus it manifested the effect of relieving the damage to vascular endothelial cell and blocking the progress of cascade damage of cerebral ischemia through inhibiting the process of inflammation. PMID- 15656141 TI - [Protective effect of the extract of Terminalia catappa leaves on acute liver injury induced by D-GalN in mice]. AB - OBJECTIVE: To study the hepatoprotective effect of the extract of Terminala catappa leaves (TCE) and the possible mechanisms underlying its protection on acute liver injury induced by D-Galactosamine (D-GalN). METHOD: In vivo: D-GalN induced liver injury model was used to evaluate the effect of TCE on the activities of serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) in mice. Structure of liver was observed and liver mitochondrial swelling was measured following D-GalN injection without or with TCE. In vitro: D-GalN induced primary cultured hepatocytes injury model was used to value the effect of TCE on cultured hepatocytes. Cell viability was measured by means of MTT assay, and the AST and superoxide dismutase (SOD) activities in supernatant of cultured cells were investigated also. RESULT: In acute hepatic injury test, with oral pretreatment of TCE, remarkable rises in serum AST and ALT activities (2.95 fold and 3.35 fold) induced by D-GalN were obviously reversed and significant morphological changes were remarkably lessened. In addition, the decrease in sensitivity of mitochondrial swelling to the exotic Ca2+ stimulation induced by D GalN was also prevented by TCE. In primary cultured hepatocytes of mice, it was found that incubation with TCE could prevent the decrease in cell viability in a dose-dependent manner. It was also found that both the increase in AST level (1.9 fold) and the decrease in SOD activity (48.0%) in supernatant of primary cultured hepatocytes induced by D-GalN could be inhibited by pretreatment of TCE. CONCLUSION: TCE has hepatoprotective activity and the mechanisms underlying its protective effect may be related to its antioxidant activity and protection on both hepatocytes and liver mitochondria. PMID- 15656143 TI - [The experimental research of Atractylodes macrocephala and Poria cocos on the meridian tropism theory in animals]. AB - OBJECTIVE: The research was designed to study the meridian tropism theory of traditional Chinese drug through experiments in animals. METHOD: We used the mouse model of deficient spleen as the object. After administering Atractylodes macrocephala and Poria cocos respectively, we measured the indexes of MDA, SOD, NO to observe the effects of the drugs on various organs and compared the results with the traditional meridian tropism theory. RESULT: The two drugs had selective effects on the quantity or activity of MDA, SOD, NO in the organs for the normal group and the model group. CONCLUSION: The selective influence of the two drugs has close relativity with the traditional meridian tropism theory. PMID- 15656142 TI - [Effects of shuangshen tongguan (SSTG) on TNF-alpha, ICAM-1 during myocardial ischemia-reperfusion injury]. AB - OBJECTIVE: To observe the effects of SSTG on infarction size and tumor necrosis factor-alpha (TNF-alpha), intercellular adhesion molecular-1 (ICAM-1) levels in serum during reperfusion injury of acute myocardial ischemia. METHOD: Anterior descending branch of coronary artery was ligated and released to create myocardial ischemia-reperfusion injury. The size and weight of infarction area and the contents of TNF-alpha, IGAM-1 in serum were assayed by N-BT staining and ELISA respectively. RESULT: The size and weight of infarct area and the contents of TNF-alpha, ICAM-1 in serum were significantly increased compared with the normal group and were obviously decreased after being treated with SSIG. CONCLUSION: Ischemia-reperfusion stimulated the secretion of TNF-alpha and ICAM 1, which play an important role in ischemia-reperfusion injury. SSTG might protect myocardium from ischemia-reperfusion injury by suppressing over-secretion of TNF-alpha and ICAM-1 and reducing the size and weight of infarction area. PMID- 15656145 TI - [A method for predicting activity of traditional Chinese medicine based on quantitative composition-activity relationship of neural network model]. AB - OBJECTIVE: To study a method for evaluating the quality of traditional Chinese medicine (TCM) according as their activity. METHOD: Combined with partial least squares (PLS), BP and RBF neural networks were selected to establish the model of quantitative composition-activity relationship (QCAR) due to their strong approximation capabilities for nonlinear function respectively. The activity of TCM was predicted with the QCAR model, and the quality of TCM was evaluated according to the predicted activity. RESULT & CONCLUSION: The proposed method was applied to evaluate the quality of Chuanxiong. The results indicated that, in the indexes including training error, prediction error and correlation coefficient, the established model is better than the model established by principal component regression or PIS regression. The new model can accurately represent the complicated nonlinear relationship between the components and the bioactivity of Chuanxiong. Consequently, this method has potential to evaluate the quality of TCM according to bioactivity. PMID- 15656144 TI - [Effect of tanshinone microemulsion on reversing MDR in human tumor cells]. AB - OBJECTIVE: To study the effect of tanshinone microemulsion (Tan-M) on the cytotoxicity to human leukemia-cell-line (K562/ADM) and the reversion of MDR in vitro. METHOD: Microemulsion being supposed as the control group, MT method is adopted to test cytotoxicity and the reverse of MDR. RESULT: Obvious cytotoxicity to K562/ADM was observed for tan-M. Cell non-toxic dosage (growth quotiety > 95%) of Tan-M is 0.2 microg x mL(-1). Low toxic dosage (growth quotiety 85-90%) was 0.7 microg x mL(-1). Cell non-toxic dosage of was 0.7 microg x mL(-1) and low toxic dosage was 1.2 microg x mL(-1). Cell non-toxic dosage of Tan-M (0.2 microg x mL(-1)) significantly lowered the IC50 of K562/ADM by ADM (P < 0.01), and reversed MDR was 3.88 times. Low toxic dosage of Tan-M reversed MDR was 3.97 times. E-M (0.2 microg x mL(-1)) reversed MDR was 2.62 times. CONCLUSION: The result indicates that tanshinone microemulsion possesses cell-toxic effects on human leukemia cell-line and may reverse MDR of tumor cells. PMID- 15656146 TI - [Effects of ginsenoside-Rb on blood lipid metabolism and anti-oxidation in hyperlipidemia rats]. AB - OBJECTIVE: To observe effects of ginsenoside-Rb (G-Rb) on total cholesterol, lipoprotein cholesterol metabolism and anti-oxidation in experimental hyperlipidemia rats. METHOD: Hyperlipidemia rats were respectively given G-Rb 50, 100, 200 mg x kg(-1) x d(-1) ig for twelve days. Total cholesterol, lipoprotein cholesterol and lipid peroxidation (LPO) contents, prostacycline (PGI2), thromboxane (TXA2), superoxide dismutase (SOD) and blood viscosity were measured. Fat accumulation in liver was also observed. RESULT: Triglyceride (TG), total cholesterol (TC), low density lipoprotein cholesterol (LDL-c) in serum, TXA2 in plasma, LPO in serum and liver, and blood viscosity were decreased significantly. High density lipoprotein cholesterol (HDLc) in serum, PGI2 in plasma and SOD in serum and liver were significantly increased by G-Rb (100, 200 mg x kg(-1)) in experimental hyperlipidemia rats. In addition, G-Rb could decrease TC/HDL-c, LDLc/HDL-c ratio, increase PGI2/TXA2 ratio and inhibit fat accumulation in liver. CONCLUSION: G-Rb could have anti-arteriosclerosis effect by improving cholesterol and lipoprotein-cholesterol metabolism, suppressing lipid peroxidation, increasing anti-oxidase activity and PGI2/TXA2 ratio. PMID- 15656147 TI - [Influence of triptolide on neuronal apoptosis in rat with cerebral injury after focal ischemia reperfusion]. AB - OBJECTIVE: To study effect of triptolide (TL) on neuronal apoptosis in cerebral tissue of rat after ischemia-reperfusion. METHOD: Triptolide at dose 0.2 or 0.4 mg x kg(-1) was intraperitoneally injected once a day for 4 d. The focal cerebral ischemia-reperfusion model was established with thread embolism in middle artery before triptolide injection on the fourth day. Neurological deficit score of rats was evaluated; and immunohistochemical techniques were used to count positive cells of express of MPO and TUNEL in cerebraltissue. RESULT: Compared with the control group, the deficit of neural function was significantly improved, and the number of infiltrate of neutrophil and neuronal apoptosis in cerebral tissue was remarkably reduced in two TL-treated groups. CONCLUSION: The results suggested that TL can inhibit infiltration of neutrophil and decrease the degree of neuronal apoptosis in cerebral tissue. PMID- 15656148 TI - [Study on the history of the processing of flos sophorae]. AB - By collecting and analyzing the information of the processing of Flos Sophorae in ancient and recent literatures, we discovered that such methods as steaming, boiling, stir-frying and baking had been used before Qing Dynasty. There were more than 10 kinds of different decoction pieces due to different subsidiary agents and distinction of processing degree. In modern times, besides stir-frying with vinegar used in Jilin, stir-flying with honey used in Henan and Shandong, being crude, yellowing by stir-frying and carbonizing by stir-frying are used in other places. This research has provided useful information for the modern processing study by summarizing the previous experiences earnestly. PMID- 15656149 TI - A time to reflect. PMID- 15656151 TI - Postnatal depression: a review of current literature. AB - The print and other forms of media offer significant amounts of information to women about pregnancy, parenthood and the birth process, but much less information exists about postnatal depression and how to cope with the frequently painful realities involved in childbirth and parenting--especially when debilitated by postnatal depression (PND). Even less information exists about such women's reactions to interventions by health professionals, which is the subject of my Ph.D study in progress. This article reviews current literature about the clinical presentation of postnatal depression, the three major types of mood disorders following childbirth, the risk factors for postnatal depression, detection and treatment of postnatal depression and the need for further research on treatment outcomes for women with PND. PMID- 15656150 TI - Focus on the beat: current fetal monitoring practice in low risk labour. AB - Assessing fetal heart and thus fetal wellbeing during labour and childbirth is an important if contentious, clinical practice. There is debate and concern about optimal methods of fetal monitoring during low risk childbirth. This paper reports on the findings of a qualitative study that investigated current midwives' practices. Data from focus groups conducted with midwives practising in Queensland was subject to a qualitative content analysis. While the ideal method of monitoring the fetal heart in low-risk labour is intermittent auscultation this analysis highlights not only the range of methods but also the rationales, contexts and factors motivating and influencing midwives' practice choices. In the current complex care environment midwives face a range of challenges to practice that facilitate the provision of consistent woman-centered care. PMID- 15656152 TI - Implementation of the SIDS guidelines in midwifery practice. AB - The literature suggests that midwives strongly influence parenting practices immediately after birth and during early postnatal management of the newborn. Midwives must therefore be aware of the current evidence and public health recommendations for reducing the risk of Sudden Infant Death Syndrome (SIDS) and provide consistent information about use of the supine position. Midwives must also include information about environmental factors that are also known to increase the risk of SIDS such as exposure to cigarette smoke, covering the infant's face during sleep and other potential unsafe sleeping practices such as co-sleeping and bed sharing with their infant. The position midwives use to settle infants and place them for sleep is an important example for parents. The position favoured by midwives when placing a newborn to sleep will have a significant impact on parental practice after discharge home. A standardised evidenced based approach to the SIDS Guidelines immediately after birth will facilitate consistency in practice and uniformity in the message parents are given about safe sleeping practices for their newborn infant. PMID- 15656153 TI - Protocol audit of dinoprostone gel induction of labour. AB - This study aimed to evaluate dinoprostone gel induction of labour. A prospective observational study was conducted over six months in 2001. Themes explored include predictive indicators within the Modified Bishop's Score, timing between doses, timing of administration and number of doses administered. Labouring after one dose of dinoprostone gel is significantly related to a Modified Bishop's Score of > 4 (p = 0.005) and position of the cervix (p = 0.031) for all women, and for multiparous women cervical dilatation (p = 0.014). Timing of administration will not affect timing of delivery. There is no benefit in waiting more than six to eight hours between doses. PMID- 15656154 TI - World without end? As America's lifespan continues to grow, so does its reluctance of facing the inevitable. The healthcare industry is caught in the middle. PMID- 15656155 TI - Editorials and opinions "crossing the bar" versus limbo. PMID- 15656156 TI - Using thermal units for estimating critical period of weed competition in off season maize crop. AB - Brazilian off-season maize production is characterized by low yield due to several factors, such as climate variability and inadequate management practices, specifically weed management. Thus, the goal of this study was to determinate the critical period of weed competition in off-season maize (Zea mays L.) crop using thermal units or growing degree days (GDD) approach to characterize crop growth and development. The study was carried out in experimental area of the University of Sao Paulo, Brazil, with weed control (C), as well as seven coexistence periods, 2, 4, 6, 8, and 12 leaves, flowering, and all crop cycle; fourteen treatments were done. Climate data were obtained from a weather station located close to the experimental area. To determine the critical period for weed control (CPWC) logistic models were fitted to yield data obtained in both W and C, as a function of GDD. For an arbitrary maximum yield loss fixed in 2.5%, the CPWC was found between 301 and 484 GDD (7-8 leaves). Also, when the arbitrary loss yield was fixed in 5 and 10%, the period before interference (PBI) was higher than the critical weed-free period (CWFP), suggesting that the weeds control can be done with only one application, between 144 and 410 GDD and 131 and 444 GDD (3-8 leaves), respectively. The GDD approach to characterize crop growth and development was successfully used to determine the critical period of weeds control in maize sown off-season. Further works will be necessary to better characterize the interaction and complexity of maize sown off-season with weeds. However, these results are encouraging because the possibility of the results to be extrapolated and because the potential of the method on providing important results to researchers, specifically crop modelers. PMID- 15656157 TI - Herbicide application timing in preharvest desiccation of soybean cultivars with different growth habits. AB - In order to evaluate the ideal time of herbicide applications to desiccate soybean cultivars with determinate and indeterminate growth habits in preharvest, an experiment was carried out at the University of Ponta Grossa (PR), during 2002 -2003. The experimental design used was a randomized blocks arranged in a factorial scheme 4 x 5 (herbicides and time of application), with three replications, using four soybean cultivars: Spring, V-Max, BR-16, and BRS-133. The herbicides glufosinate, diquat, and paraquat were used at the rate of 400 g ha(-1), when soybean was at stage of R6.0, R6.5, R7.0, R7.5, and R8.0. The assessments included yield, as well as yield components, considering each cultivar separately. The results show that desiccation should be done on cultivars with determinate growth habit at stages between R7.0 and R7.5, independent of herbicide used. For indeterminate growth habit cultivars, desiccation should be done later, because those pods formed after R6.5 stage are important to define yield; although herbicides have different mode of action, yield results were similar. In cultivars, desiccation should start when all pods in the middle of plants become yellow and grains are not held by fibers. PMID- 15656158 TI - Population dynamics of Digitaria spp submitted to selection pressure by herbicides in sugarcane crop. AB - The objective of this research was to study population dynamics of the weed crabgrass, genus Digitaria, submitted to selection pressure by herbicides currently applied in sugarcane crops in Brazil. In the first experiment two crabgrass species (Digitaria nuda and Digitaria ciliaris) and eight herbicide treatments applied in preemergence were used, and control percentage was evaluated at 7, 14, and 21 days after herbicide application (DAA). In the second experiment the level of tolerance through dose-response curve was determined for the species D. nuda and D. ciliaris, to the herbicides imazapyr, tebuthiuron, ametryne, and metribuzin. All the herbicides studied were efficient in controlling D. ciliaris, however, for D. nuda the best results were obtained only with ametryne, metribuzin, and isoxaflutole. The relation (T/S) between the rate required to reduce plant dry biomass (GR50) at 21 DAA of D. nuda and D. ciliaris was 16 for imazapyr and 6.3 for tebuthiuron, showing differential susceptibility of species; however for ametryne the rate T/S of 1.1 showed that D. nuda was not tolerant to this herbicide. For metribuzin, at 1.92 kg a.i. ha(-1), reduction of dry biomass was 80 and 90% to D. nuda and D. ciliaris, respectively. Even being controlled by metribuzin, D. nuda presented a higher level of tolerance to this herbicide, what was confirmed by the relationship T/S 14.4. As general conclusion of the research, it can be stated that the species D. nuda is more tolerant to ALS inhibiting herbicides and substituted urea, when compared with D. ciliaris; probably, D. nuda was selected by repetitive use of these herbicides. PMID- 15656159 TI - Sorption and leaching potential of acidic herbicides in Brazilian soils. AB - Leaching of acidic herbicides (2,4-D, flumetsulam, and sulfentrazone) in soils was estimated by comparing the original and modified AF (Attenuation Factor) models for multi-layered soils (AFi). The original AFi model was modified to include the concept of pH-dependence for Kd (sorption coefficient) based on pesticide dissociation and changes in the accessibility of soil organic functional groups able to interact with the pesticide. The original and modified models, considering soil and herbicide properties, were applied to assess the leaching potential of selected herbicides in three Brazilian soils. The pH dependent Kd values estimated for all three herbicides were observed to be always higher than pH-independent Kd values calculated using average Koc data, and therefore the original AFi model overestimated the overall leaching potential for the soils studied. PMID- 15656160 TI - Sorption, degradation, and leaching of tebuthiuron and diuron in soil columns. AB - A study in small outdoor lysimeters was carried out to determine the leaching of the herbicides tebuthiuron and diuron in different soil types, using undisturbed soil columns. Soil sorption and degradation for both herbicides were also studied in the laboratory. The multi-layered AF (Attenuation Factor) model was evaluated for predicting the herbicides leaching in undisturbed soil columns. Tebuthiuron leached in greater amounts than diuron in both soils. Sorption was well represented by linear and Freundlich equations, however parameters from the linear equations were used in the AF model. In general, both herbicides presented very low sorption, with diuron presenting lower values of sorption coefficient than tebuthiuron in the two soils. Chromatographic data indicated rapid late degradation of diuron and tebuthiuron in both soil types at two different depths. Simple exponential equation was not able to represent degradation, thus a bi exponential equation was used, and some model adjusting was needed. Average measured amounts of each herbicide were compared with amounts predicted by the multi-layered-soil AF model. The AF model was able to predict leaching amounts in the sandy soil, especially for diuron, however it did not perform well in the clayey soil. PMID- 15656161 TI - Effects of herbicide and insecticide interaction on soil entomofauna under maize crop. AB - The aim of this study was to evaluate the impact of the herbicide mixture nicosulfuron + atrazine, with or without the insecticide chlorpyrifos, onto soil entomofauna under maize crop. The treatments, applied 25 days after maize emergence, were represented by a weeded control without insecticide and herbicide, a weeded control with chlorpyrifos, and mixtures of nicosulfuron + atrazine, with or without chlorpyrifos. Arthropods populations, on the soil surface, as well as inside the soil under maize, were principally represented by mites (Arachnida: Acari), decomposer collembolans (Hexapoda:Parainsecta:Collembola) and predator ants (Hymenoptera:Formicidae). The nicosulfuron + atrazine mixture with chlorpyrifos and the isolated chlorpyrifos reduced the population dynamics of all insect groups on the soil surface compared to the weeded control. In the soil, mite and ant populations were reduced after application of the herbicide mixture with chlorpyrifos and of the isolated chlorpyrifos. PMID- 15656162 TI - Influence of soil properties on bioaccumulation of 14C-simazine in earthworms Eisenia foetida. AB - The toxicity of pesticides has been evaluated by several methods including tests with earthworms in both artificial and natural soils treated with the compounds. The ecological niches of earthworms make them good bioindicators of soil contamination. The bioaccumulation of 14C-simazine (6-chloro-N2-N4-diethyl- 1,3,5 triazine-2,4-diamine) was evaluated in earthworms (Eisenia foetida) maintained during three months in two substrates with different physical-chemical characteristics. The substrates were treated with 3.0 mg and 330 kBq of 14C simazine kg(-1) substrate. Results indicated that worms did not influence simazine dissipation in both substrates as indicated by similar recoveries and with no statistical differences with and without earthworms. The radiocarbon recoveries were 86.8 and 95.3%, respectively in the substrates with lower and higher organic matter contents with earthworms, and 91.0 and 107.4% in the same substrates without worms. However, in earthworms the recoveries were statistically higher when they were maintained in the substrate with lower amount of organic matter (0.89%) than from the higher one (0.33%). Consequently, 14C simazine bioconcentration factor (BCF) was also greater in the substrate with lower organic matter (6.89+/-1.55) than in the substrate with higher organic matter content (0.88+/-0.06). The results suggest that the higher soil organic matter content will cause lower probability of contamination of soil organisms with simazine. PMID- 15656163 TI - Alternative herbicides to manage Italian ryegrass (Lolium multiflorum lam) resistant to glyphosate at different phenological stages. AB - During the growing season of 2002--2003, field and greenhouse experiments were conducted with the objective of evaluating the influence of Italian ryegrass phenological stages and management alternatives on the control of resistant biotypes to glyphosate. Three field experiments were conducted in Lagoa Vermelha, RS, Brazil and glyphosate was applied alone and in combinations with alternative herbicides. Two greenhouse experiments were also conducted at the Department of Crop Science, ESALQ/USP, Piracicaba, SP, Brazil. The Italian ryegrass resistant population was collected from Lagoa Vermelha, RS, Brazil. From the results it was possible to conclude that: (i) the more advanced the phenological stage of application, the more difficult the control of resistant Italian ryegrass by glyphosate, mainly by the rate of 960 g a.i. ha(-1); however, this rate applied at earlier phenological stage (five tillers), the control was higher than 90%; (ii) with the increment of glyphosate rate, it significant response was observed on the control at all stages of application; (iii) the mixture of glyphosate + clethodim (1440 + 72 g a.i. ha(-1)), paraquat + diuron (500 + 250 g a.i. ha(-1)), at all stages of application and clethodim (96 g a.i. ha(-1)) and paraquat + diuron (300 + 150 g a.i. ha(-1)) at the initial stages until pre-flowering were excellent alternatives for management of these populations; and (iv) the response of control was much faster for the mixture of glyphosate + clethodim, independently of growth stage. PMID- 15656164 TI - Allelopathic effects of aconitic acid on wild poinsettia (Euphorbia heterophylla) and morningglory (Ipomoea grandifolia). AB - High infestations of alexandergrass (Brachiaria plantaginea) in experimental area have reduced seedbank of some weed species in soil at a greater degree compared to those obtained with the use of an efficient herbicide. Aconitic acid (AA) has been identified in those plants in high concentration. Thus, two experiments were carried out in the laboratory to determine the effects of pure AA on weed seed germination and on endophytic fungi in wild poinsettia (Euphorbia heterophylla) and morningglory (Ipomoea grandifolia) weed species. Doses of AA (0, 0.5, 1.0, and 2.0 mM) were introduced in agar-water 0.5% medium and weed seeds treated with and without external sterilization by sodium hypochloride. Four replications of fifty seeds of each species by treatment were sown in plastic boxes with cover and maintained in germination chamber for 12 days. Reduction in germination rate, root and stem growth were observed in both species, as well as reduction in root number in morningglory were also observed at 2.0 mM concentration of AA. An increase in number of seeds with presence of endophytic fungi was also observed in both species, independently of seed sterilization, mainly of a green type fungi coloration in wild poinsettia. Aconitic acid presents allelopathic activity and can stimulate endophytic fungi species growth in weed seeds. PMID- 15656165 TI - Allelopathic potential and systematic evaluation of organic extracts from Canavalia ensiformis leaves (Jack beans). AB - This article describes the assessment of possible allelopathic potential of organic extracts obtained from leaves of Canavalia ensiformis under laboratory conditions. Furthermore, a systematic evaluation of these extracts was carried out using specific protocols developed in capillary electrophoresis (CE) to determine some groups of secondary metabolites. After the identification and quantification of compounds, the effects of compounds on germination of some common weeds was investigated, which are becoming a real problem in pastures in the state of Para, Brazil. PMID- 15656166 TI - Assessment of aquatic plants in the reservoirs of AES-tiete and development of an integrated control model for the most important species. AB - The general objective of this work was to develop a monitoring and management model for aquatic plants that could be used in reservoir cascades in Brazil, using the reservoirs of AES-Tiete as a study case. The investigations were carried out at the reservoirs of Barra-Bonita, Bariri, Ibitinga, Promissao, and Nova-Avanhandava, located in the Tiete River Basin; Agua Vermelha, located in the Grande River Basin; Caconde, Limoeiro, and Euclides da Cunha, which are part of the Pardo River Basin; and the Mogi-Guacu reservoir, which belongs to the Mogi Guacu River basin. The main products of this work were: development of techniques using satellite-generated images for monitoring and planning aquatic plant control; planning and construction of a boat to move coating plant masses and an airboat equipped with a DGPS navigation and application flow control system. Results allowed to conclude that the occurrence of all types of aquatic plants is directly associated with sedimentation process and, consequently, with nutrient and light availability. Reservoirs placed at the beginning of cascades are more subject to sedimentation and occurrence of marginal, floating and emerged plants, and are the priority when it comes to controlling these plants, since they provide a supply of weeds for the other reservoirs. Reservoirs placed downstream show smaller amounts of water-suspended solids, with greater transmission of light and occurrence of submerged plants. PMID- 15656167 TI - Multiple resistance of acetolactate synthase and protoporphyrinogen oxidase inhibitors in Euphorbia heterophylla biotypes. AB - Resistance to acetolactate synthase (ALS)-inhibiting herbicides in Brazil has been documented for six species. The probability to select biotypes of Euphorbia heterophylla (EPPHL) with multiple resistance increases in the same order of magnitude as the use of other herbicides belonging to only one mechanism of action. The objectives of this work were to evaluate the distribution of resistant populations (R) in the states of the Parana and Santa Catarina; to determine the existence of populations of EPHHL with multiple resistance to ALS and PROTOX inhibitors, and to confirm the occurrence of cross resistance to compounds of these mechanisms of action. Seeds of EPHHL of areas with suspected resistance had been sampled in 97 places during 2003. In the greenhouse experiment samples of each population were sprayed with imazethapyr or fomesafen, at only one rate. To identify the resistant ones they were sprayed with different levels of the herbicides imazethapyr and fomesafen. Later they were sprayed with diverse herbicides of the same mechanisms of action to confirm the multiple/cross resistance. There is widespread distribution in the region of populations with resistance to ALS inhibitors. Some biotypes demonstrated resistance to herbicides from the two mechanisms of action. The resistance factor (FR), or the relation of resistance between R and susceptible biotypes, confirms the existence of two biotypes of EPHHL with cross resistance to several herbicides inhibitors of ALS and PROTOX. PMID- 15656168 TI - Effects of plant densities and management of purple nutsedge on sugarcane yield and effect of growth stages and main way of herbicides contact and absorption on the control of tubers. AB - Field experiments carried out with Cyperus rotundus L. at low (58-246), medium (318-773), and high (675-1198 shoots/m2) densities showed sugarcane yield reductions of 13.5, 29.3, and 45.2%, respectively in relation to the control. In the second field experiment, the integration of a mechanic method with two sequences of plowing and disking operations in the dry season, and complementary applications of trifloxysulfuron-sodium + ametrine and sulfentrazone (rainy season) was studied. Average of the chain connected to original shoot showed 92, 95, and 65% of reduction with trifloxysulfuron-sodium + ametrine and surfactant, at the application stages "early," preflowering, and full flowering, respectively. PMID- 15656169 TI - Population dynamics of weeds in no-tillage and conventional crop systems. AB - Population dynamics of weeds in successive maize and bean crops were evaluated in two soil management systems (conventional and no-tillage), for two maize applications (grain and silage), and in four consecutive growing seasons. Every year, conventional tillage consisted in plowing and harrowing before sowing. In no-tillage, chemical weed desiccation was made with the mixture glyphosate + 2.4 D. To control weeds, the mixture fluazifop-p-butil + fomesafen was applied on the bean crop in all the planting seasons, and the herbicides nicosulfuron + atrazine on maize after crop emergence (1998--1999, 1999--2000, 2001--2002) and atrazine + metolachlor before emergence (2000--2001). Purple nutsedge (Cyperus rotundus) was the most important species under conventional soil tillage; while in no-tillage the dicotyledonous weed species (Amaranthus deflexus, Bidens pilosa, Euphorbia heterophylla, Galinsoga parviflora Ipomoea grandifolia) were the most relevant. Regardless of the maize use, the C. rotundus population and tuber bank, with prevailingly dormant tubers, was considerably reduced in no-tillage compared with the conventional system. PMID- 15656170 TI - Effects of hand weeding strip and nitrogen fertilizer on corn plants. AB - The objective of the present research was to evaluate effects of different strip weed control associated with nitrogen fertilizer on corn applied after planting. The experiment was set and conducted in Botucatu, Sao Paulo State, Brazil, and the hybrid planted was Dekalb 333-B. A completely randomized block design with four replications was used. Experimental plots were disposed as a factorial scheme 2 x 2 x 4, constituted by two types of weeding on row (with or without manual hoeing), two types of weeding on inter-row (with or without manual hoeing), and four nitrogen levels applied after planting (00, 60, 90, and 120 kg ha(-1)). Plots were composed by six rows with 5 m length. Nitrogen fertilizer was applied at 35 days after emergence (d.a.e). For weed community it was evaluated: weed density, dominancy, frequency, and relative importance. The main weed species were: Brachiaria plantiginea, Amaranthus retroflexus, Bidens pilosa, Cyperus rotunds, Brachiaria decumbens, Euphorbia heterofila, Oxalis latifolia, Acanthospermum hispidum, Commelina benghalensis. It was evaluated corn height at 40 and 100 d.a.e., first ear insertion height at 100 d.a.e., and final grain yield at harvesting. Plants and first ear insertion height were affected when nitrogen fertilizer was not applied. Treatments without weed control showed that weed interfered negatively with plants height. There were no correlation between weeds and nitrogen fertilizer for all parameters evaluated. Parcels without weed showed the highest ear weights and final grain production. Treatments that received nitrogen fertilizer, independently of studied arrangement, provided higher yields. PMID- 15656171 TI - Absorption and translocation of 2,4-D in plants of Memora peregrina. AB - The objective of this work was to evaluate absorption and translocation of the herbicide 2,4-D in plants of Memora peregrina. The herbicide 2,4-D was used alone with the formulation DMA 806 BR and associated with the herbicide picloram in the commercial product Padron. Levels of radioactivity on the treated leaves were determined in sample obtained after washing them with methanol and chloroform at different times after the application of the radiolabelled formulation (1, 2, 4, 8, 24, and 48 h). Translocation was evaluated by cutting plants between stem and root. The parts obtained were: root, stem, leaf treated, leaves above the leaf treated, leaves below the leaf treated, and leaf opposite of the leaf treated. These parts were weighted, dried, ground, burnt, and radioactivity in the samples was determined. The results suggest that the translocation of the radioactive herbicide 2,4-D was insignificant in plants of M. peregrina in the two treatments evaluated. Absorption of 14C 2,4-D in the treatment with DMA 806 BR and the mixture of DMA 806 BR plus Padron had the same behavior. These observations explain the inefficient control obtained with this herbicide in plant species under study. PMID- 15656172 TI - Strategies for management of Sicyos polyacanthus Cogn. (Cucurbitaceae) in sugarcane crops of Tucuman, Argentina. AB - Sicyos polyacanthus is one of the most important weed in sugarcane crops of Tucuman (Argentina). The objective of this work was to establish strategies that would decrease the weed incidence in the crop to a minimum level. The study was carried out during 1998--2003 at five localities of sugarcane production of Tucuman (Argentina). The plots were 20 m long (192 m2) with 6 furrows and each plot was replicated five times. Treatments were: (i) Mechanical-chemical cultivation without fire; (ii) Mechanical-chemical culltivation with fire; (iii) Mechanical cultivation with handle pulled and with fire; (iv) Mechanical cultivation with handle pulled without fire; (v) Mechanical and chemical variants with fertilization and without fertilization; (vi) Mechanical and chemical variants with watering and without watering; (vii) Fallow and rotation, at the sugarcane crop renovation; (viii) Mechanical and chemical variants for plant cane and ratoon cane; and (ix) Mulching of harvest rests. The results suggest that besides the use of preemergent herbicides, fire marks the entry point of control, influencing fluxes and seed viability. It appears that fallow, mulching, and rotation of crops is fundamental for eliminating seeds that live short time in the soil and increase the mortality rate of species. PMID- 15656173 TI - Eucalyptus ESTs related to genes for oxidative stress. AB - Oxidative stress generating active oxygen species has been proved to be one of the underlying agents causing tissue injury after the exposure of Eucalyptus (Eucalyptus spp.) plants to a wide variety of stress conditions. The objective of this study was to perform data mining to identify favorable genes and alleles associated with the enzyme systems superoxide dismutase, catalase, peroxidases, and glutathione S-transferase that are related to tolerance for environmental stresses and damage caused by pests, diseases, herbicides, and by weeds themselves. This was undertaken by using the eucalyptus expressed-sequence database (https//forests.esalq.usp.br). The alignment results between amino acid and nucleotide sequences indicated that the studied enzymes were adequately represented in the ESTs database of the FORESTs project. PMID- 15656174 TI - Herbicide leaching on a recharge area of the Guarany aquifer in Brazil. AB - The region of Ribeirao Preto City, located in Southeast of Brazil, Sao Paulo State, is an important sugarcane, soybean, and corn producing area with a high level of pesticides utilization. This region is also an important recharge area for groundwater supply of the Guarany aquifer. Since the past ten years atrazine, simazine, ametryn, tebuthiuron, diuron, 2,4-D, picloram, and hexazinone are the main herbicides used in this area. In order to study a possible leaching of some of these herbicides into the aquifer, surface, and groundwater samples were collected in a watershed during the years of 1996 to 2003, from different locations. To detect and quantify the herbicides a GC-MS (gas chromatograph/mass spectrometry) method was used. The response of the herbicides analyzed was linear over the concentration range of 0.02 to 2.0 microg/L. Analysis of groundwater revealed that the herbicides tebuthiuron, diuron, atrazine, simazine, and ametryn were not present in the samples. In the surface water collected in 1997, ametryn was present in two out of nine locations with concentrations ranging from 0.17 and 0.23 microg/L, which is above the allowable 0.1 microg/L according to the European safety level. The leaching potential of tebuthiuron, diuron, atrazine, simazine, 2,4-D, picloram, and hexazinone has been evaluated using CMLS-94, "Chemical Movement in Layered Soil," as simulation model. No leaching into the depth of the water table at 40 m was found. PMID- 15656175 TI - Detached leaf culture: viability to evaluate 2,4-D toxicity symptoms in cotton apex leaves. AB - To study the viability of detached leaf culture technique, studies were carried out with detached leaves from cotton apex (true trilobed leaves). The prepared leaves were sprayed with 2,4-D amine and ester, at rates of 10, 30, 70, and 100% of the recommended doses. Detached leaves without herbicide spray were used as controls. Simultaneously, a greenhouse experiment was conducted with the same treatments as used for the detached leaves experiment. Toxicity was measured through a 0-to-5 grading according to the percentage of affected leaf area in the detached leaves experiment or examining the affected rate of whole plant as indicated in the greenhouse. Results showed that the ester form of the herbicide induced earlier and more severe toxicity symptoms in detached leaves and greenhouse grown plants. Positive and significant correlations (p < 0.001) were found between toxicity results obtained at 7 and 14 days after application in detached leaves and greenhouse plants (r = 0.97 and 0.92, respectively). Negative, significant correlations (p < 0.005) were found between the toxicity levels found at 7 and 14 days after application in detached leaves and dry matter of cotton plants grown in the greenhouse (r= - 0.92 and -0.92, respectively). PMID- 15656177 TI - Selectivity of the herbicide flazasulfuron applied after postemergence in sugarcane (Saccharum spp. L.) crop. AB - The selectivity of the herbicide flazasulfuron was evaluated when applied at two rates (50 and 100 g/ha) with and without surfactants (Aterbane or Agral at 0.2% v/v). The treatment was applied at early (three leaves) and late (five to six leaves) stages of the postemergence of sugarcane plants (var. RB845257) grown in two soils (sandy and clay) with good moisture status before and after application. Despite the toxicity symptoms, especially at the late stage with the higher rate of application in the sandy soil, the herbicide did not affect the growth nor the stalk yield. The presence of the surfactants had no effect on the toxicity symptoms. PMID- 15656176 TI - Influence of type and amount of straw cover on weed emergence. AB - This research was undertaken during 2003-2004 growing season to evaluate the effects of type [forage sorghum "hybrid Cober Exp" (Sorghum bicolor x Sorghum sudanense), forage millet (Pennisetum americanum "var. BN2"), finger millet (Eleusine coracana), and St Lucia grass (Brachiaria brizantha)] and amount of straw cover (5.5 and 3.0 t ha(-1)) upon the emergence of Bidens pilosa, Chamaesyce spp., Amaranthus spp., and Commelina benghalensis, under field conditions of the Brazilian Cerrado, in the region of Uberlandia--MG. The control consisted additional treatment lacking the straw cover. Emergence of weed depended on the type and amount of straw cover, as well as the weed species. The lowest number of B. pilosa seedlings was found in the presence of forage sorghum straw; Chamaesyce spp. in the lack of straw; Amaranthus spp. in the presence of higher amount of forage sorghum and forage millet, and lower amounts of forage millet and Finger Millet. All the types and amounts of straw reduced the emergency of C. benghalensis, except at the lowest level of St Lucia grass and the lack of straw. PMID- 15656178 TI - Weed interference on maize (Zea mays) under no tillage system. AB - A field trial was carried out in Brazil in March 2002 with the aim to evaluate the effects of different timing and extension of weedy period on maize productivity. The hybrid Pioneer 30K75 was sowed under 7 t ha(-1) mulching promoted by glyphosate spraying. The treatments were divided in two groups: In the first group, weeds were maintained since the maize sowing until different periods in the crop cycle: 0, 14, 28, 42, 56, 70, and 150 days (harvesting time). In the second group, the maize crop was kept weed free for the same periods of the first group. Weed control was done through hand hoeing. A complete randomized blocks experimental design with five replications was used for plots distribution in the field. Nonlinear regression model was used to study the effects of weedy or weedfree periods on maize productivity. Weed community included 13 families and 31 species. Asteraceae, Poaceae, and Euphorbiaceae were the most abundant families. Results showed that under no tillage condition with 7 t ha(-1) mulching at sowing time, the maize crop could cohabit with weed community for 54 days without any yield lost. On the other hand, if the crop was kept weed free for 27 days, the weed interference was not enable to reduce maize production. According to these results one weed control measure between 27 and 54 days after crop emergence could be enough to avoid any reduction in maize productivity. PMID- 15656179 TI - Cover crops and natural vegetation mulch effect achieved by mechanical management with lateral rotary mower in weed population dynamics in citrus. AB - The aim of this study was to obtain information on practical weed management in order to reduce the use of herbicides thereby contributing to the sustainable development of citrus crop. The experiment was carried out under field conditions at the experimental area of the Department of Vegetal Production at the College of Agriculture "Luiz de Queiroz," Piracicaba, SP, Brazil, during the season 2002 2003. Influence of mulches produced by four types of vegetations on the dynamic population of weeds in the line of citrus crop (Citrus sinensis (L.) Osbeck) was evaluated. The experimental design was of factorial randomized blocks (4 x 2), where the treatments were: (i) four types of vegetation: Dolichos lablab L., Cajanus cajan (L.) Millsp, Penisetum glaucum (L.) Leeke, and the natural infestation composed basically by Panicum maximum Jacq.; and (ii) two types of fertilization: directed under canopy and broadcast. Mechanical management of the different vegetations was accomplished using a lateral rotary mower, KAMAQ, I model NINJA MAC 260, projected to release the green cut material under crop canopy, forming a mulch layer. The studied parameters were: (i) counting of weeds per m2 in the crop line after 30, 60, 90, 180, and 210 days following the cutting of existing vegetation; (ii) percentage of covered area by weeds; and (iii) some chemical properties of the soil. It was observed that the natural infestation showed a better weed control when compared with the other treatments, and that the broadcast fertilization, regardless of coverage used, presented a lower number of weeds. PMID- 15656180 TI - Weed seedbank evaluating method to generate spatial distribution maps. AB - The objective of this work was to develop a fast and practical method of weed seedbank evaluation to generate spatially distributed maps for use in site specific weed management. Soil cores were collected at 0.20 m depth, air-dried, and then submitted to seedling growth in greenhouse. The sampling grid of 20 by 20 m was georeferenced by Global Positioning System, obtaining 73 soil cores with three replicates. During the greenhouse trial, there were two peaks of weed seedling growth: one in 119 days after water irrigation and another after KNO3 application. Weeds seedbank maps were obtained at different stages of seedling growth. The Pearson correlation was 0.99 for Brachiaria plantaginea seedbank map, 0.95 for Commelina benghalensis, and 0.85 for Cyperus rotudus generated at 119 days compared with 392 days after seedling growth in the greenhouse. The Brachiaria plantaginea seedbank map evaluated at 35 days presented correlation of 0.97 with 392 days. It was concluded that, for site-specific weed seedbank management, the evaluation of seedling growth in greenhouse until the first emergence peak is enough to generate weed seedbank maps. PMID- 15656181 TI - Periodic inspection on crop sprayers: results according to age of sprayers. AB - The objectives of the IPP Project--Periodic Inspection on Crop Sprayers--are to develop methods for sprayer certification, analyze quality on spray operation, propose an inspection system for crop sprayers in Brazil, improve environmental quality on spray operation, and reduce costs on chemical control for plant protection systems. Periodic inspections on crop sprayers are performed in several countries and are compulsory in most of them, and it is becoming an important tool for improvement and optimization of use of chemicals. The IPP Project in Brazil is funded by FAPESP--Fundacao de Amparo a Pesquisa do Estado de Sao Paulo. The results so far showed that all the sprayers presented failures. However, most of them could be approved with minor services. As an example, 56.6% of the sprayers with more than 2 years of use presented leaks, 47% of them had damaged hoses and 80.5% presented bad tips (nozzles). These results indicate the need for better procedures of use and maintenance of sprayers, justifying the periodic inspection system. PMID- 15656182 TI - Impact of sublethal doses of 2,4-D, simulating drift, on tomato yield. AB - Tomato is considered one of the most sensitive crops regarding 2,4-D drift. In many cases, such susceptibility has led to important restrictions in the use of 2,4-D based products. Field experiments were carried out for two consecutive years in tomato, by applying sublethal doses of 2,4-D (ranging from 0.42 to 13.44 g a.i. ha(-1)) directly to plants, at different stages of growth, as a simulation of eventual drifts to the crop. The range of rates was based on the assumption of a 0.0625-2.0% drift level of a 1 L ha(-1) of the most common formulated herbicides. For this crop, the range of rates between 0.42 and 13.44 g a.i ha(-1) applied at the beginning of flowering caused a linear crop reduction. On the other hand, rates < or = 13.44 g a.i. ha(-1) applied after full development of fourth truss stage or latter had no effect on crop yield or development. For tomato, tolerance to 2,4-D strongly increases with plant age. PMID- 15656183 TI - Deposition and leaching of tebuthiuron on sugar cane straw applied with and without alkyl polyglycoside adjuvant. AB - A laboratory experiment was carried out aiming to study the effects of an alkyl polyglycoside adjuvant (APG) on deposition and leaching of the herbicide tebuthiuron applied on sugar cane straw. Tebuthiuron, at concentration of 1200 mg L(-1), was applied separately and in tank mix with the APG adjuvant, at concentrations of 0.07 and 0.09% (wt v(-1)), using a spraying volume of 204 L ha( 1). A precipitation equivalent to 20 mm of rain was simulated, 24 h after the applications, to evaluate the herbicide leaching. The quantification of tebuthiuron was carried out by the high performance liquid chromatography (HPLC). It was observed that the addition of APG adjuvant at 0.07% (wt v(-1)) provided an increase of 11.5% in the deposition of tebuthiuron on straw, reduction of 50.4% in the drift of the herbicide and it did not affect significantly the leached amount (68.5%), when compared with the treatment where tebuthiuron was applied alone (70.8%). At the concentration of 0.09% (wt v(-1)), the APG adjuvant caused an increase of 22.7% in the deposition; it reduced the drift of the herbicide by 99.9% and reduced the leached amount by 7.6% thereby increasing the retention of the herbicide by straw. PMID- 15656184 TI - Optical properties and potential applications of doped semiconductor nanoparticles. AB - Recent studies on the optical properties, in particular, luminescence, of a variety of doped semiconductor nanoparticles are reviewed. The effects of quantum confinement, temperature and pressure on luminescent properties are discussed. In addition, electroluminescence, cathodoluminescence, magnetoluminescence and related applications involving doped semiconductor nanoparticles are presented. A new phenomenon, upconversion luminescence of doped nanoparticles, is reviewed and its potential applications are discussed. While more research efforts are necessary in order to fully understand the fundamentals and explore the great technological potential behind doped nanoparticles, recent results already show that this is a new and exciting field with applications in many areas. In particular, the emerging field of "spintronics", where spin states are exploited in analogy to conventional electronic states, is discussed and the advantages of using doped semiconductor nanoparticles are elucidated. PMID- 15656185 TI - Nanofabrication with atomic force microscopy. AB - Atomic force microscopy (AFM) was developed in 1986. It is an important and versatile surface technique, and is used in many research fields. In this review, we have summarized the methods and applications of AFM, with emphasis on nanofabrication. AFM is capable of visualizing surface properties at high spatial resolution and determining biomolecular interaction as well as fabricating nanostructures. Recently, AFM-based nanotechnologies such as nanomanipulation, force lithography, nanografting, nanooxidation and dip-pen nanolithography were developed rapidly. AFM tip (typical radius ranged from several nanometers to tens of nanometers) is used to modify the sample surface, either physically or chemically, at nanometer scale. Nanopatterns composed of semiconductors, metal, biomolecules, polymers, etc., were constructed with various AFM-based nanotechnologies, thus making AFM a promising technique for nanofabrication. AFM based nanotechnologies have potential applications in nanoelectronics, bioanalysis, biosensors, actuators and high-density data storage devices. PMID- 15656186 TI - Improvement of thermal contact resistance by carbon nanotubes and nanofibers. AB - Interfacial thermal resistance results of various nanotube and nanofiber coatings, prepared by chemical vapor deposition (CVD) methods, are reported at relatively low clamping pressures. The five types of samples examined include multi-walled and single-walled nanotubes growth by CVD, multi-walled nanotubes grown by plasma enhanced CVD (PECVD) and carbon nanofibers of differing aspect ratio grown by PECVD. Of the samples examined, only high aspect ratio nanofibers and thermally grown multi-walled nanotubes show an improvement in thermal contact resistance. The improvement is approximately a 60% lower thermal resistance than a bare Si-Cu interface and is comparable to that attained by commercially available thermal interface materials. PMID- 15656187 TI - Fabrication of vertical ZnO nanowires on silicon (100) with epitaxial ZnO buffer layer. AB - Vertical ZnO nanowires were successfully grown on epitaxial ZnO (002) buffer layer/Si (100) substrate. The nanowire growth process was controlled by surface morphology and orientation of the epitaxial ZnO buffer layer, which was deposited by radio-frequency (rf) sputtering. The copper catalyzed the vapor-liquid-solid growth of ZnO nanowires with diameter of approximately 30 nm and length of approximately 5.0 microm. The perfect wurtzite epitaxial structure (HCP structure) of the ZnO (0002) nanowires synthesized on ZnO (002) buffer layer/Si (100) substrate results in excellent optical characteristics such as strong UV emission at 380 nm with potential use in nano-optical and nano-electronic devices. PMID- 15656188 TI - Synthesis of DNA-polypyrrole nanocapsule. AB - A template synthesis technique of encapsulating DNA molecules inside polypyrrole (PPy) nanotubes, termed as DNA-PPy nanocapsule, has been described here. The insertion of DNA molecules inside polypyrrole nanotubes is spontaneous in nature. The structural characterizations and possible applications of DNA-PPy nanocapsule in bio-sensing devices have been discussed. PMID- 15656189 TI - Stabilization of single-wall carbon nanotubes in fully sulfonated polyaniline. AB - The interaction of single wall carbon nanotubes (SWNT) with an aqueous solution of the fully sulfonated polyaniline poly(2-methoxyaniline-5-sulfonic acid) (PMAS) and (+)-1-phenylethylamine (PhEA) has been investigated using spectroscopic methods. UV-vis spectral measurements show that the PMAS backbone undergoes conformational changes upon interaction with both SWNT and PhEA. Partial intercalation of PMAS into SWNTbundles was confirmed by Raman spectroscopy and electron microscopy. PMID- 15656190 TI - Ordered nano-lslands on (La,Ba)MnO3 thin film surface by self-organization. AB - Self-organized nanostructures composed of discrete coherent islands were found on La0.67Ba0.33 MnO3 (LBMO) film surfaces. Our results revealed that rippled surface structures on LBMO films appeared during post-annealing at high temperature, and that the step edges of the rippled surfaces offer energetically favorable positions for the nucleation of coherent islands. The rippled surface serves as a template for the arrangement of coherent islands. This provides a new way to prepare ordered surface nanostructures in perovskite oxides without the necessity to employ any preprocessing techniques prior to the growth. PMID- 15656192 TI - Cationic poly(lactide-co-glycolide) nanoparticles as efficient in vivo gene transfection agents. AB - Poly(lactide-co-glycolide) (PLGA), a biocompatible and biodegradable polyester co polymer of PLA and PGA, has been recognized for its ability to deliver genes. However, gene delivery by PLGA nanoparticles is limited by their negative charge and their poor transport through mucosal barriers. In this study, PLGA nanoparticles were surface modified with cationic chitosan in an effort to improve their gene delivery capability. PLGA nanoparticles were synthesized by emulsion-diffusion-evaporation technique using PVA-chitosan (PLGA1) or PVA chitosan-PEG (PLGA2) blend as stabilizers. This method is reproducible and produces nanoparticles with hydrodynamic diameter <200 nm. The nanoparticles were characterized by zetasizer, photon correlation spectroscopy and atomic force microscopy. A549 epithelial cells were transfected in vitro with PLGA particles complexed with a reporter plasmid encoding green fluorescent protein. PLGA particles transferred EGFP gene, but were less efficient than the lipofectamine control. The nanoparticles were also tested for their ability to transport across the nasal mucosa in vivo in mice. The results show that both PLGA1 and PLGA2 facilitate gene delivery and expression in vivo with increased efficiency and without causing inflammation, as measured by IL-6. Together, these results indicate that chitosan-modified PLGA nanoparticles have greater potential as gene carriers. PMID- 15656191 TI - Mesenchymal stem cell adhesion and spreading on nanostructured biomaterials. AB - Bone marrow-derived human mesenchymal stem cells were seeded in serum-free media onto ion beam-deposited nanostructured metalloceramic (Ti-Cr-N) films and plasma nitrided titanium disks, which were left uncoated as well as precoated with fetal bovine serum. Precoating the disks with serum appears to stimulate cell spreading on both the titanium nitride and metalloceramic materials for as little as 1 hour incubation time. The implication is that both of these materials can adsorb serum proteins in amounts sufficient to influence cell adhesion and spreading for potentially improved in vivo response of orthopedic and dental implants. The materials in this study may prove to exhibit enhanced biological and mechanical properties when compared to conventional micron-scale implant materials such as titanium or cobalt-chrome alloys. PMID- 15656193 TI - Extinction coefficients and purity of single-walled carbon nanotubes. AB - Single-walled carbon nanotubes (SWNTs) hold great promise for advanced applications in aerospace, electronics and medicine, yet these industries require materials with rigorous quality control. There are currently no accepted standards for quality assurance or quality control among the commercial suppliers of SWNTs. We briefly discuss the applicability of various techniques to measure SWNT purity and review, in detail, the advantages of near infrared (NIR) spectroscopy for the quantitative assessment of the bulk carbonaceous purity of SWNTs. We review the use of solution phase NIR spectroscopy for the analysis and characterization of a variety of carbon materials, emphasizing SWNTs produced by the electric arc (EA), laser oven (LO) and HiPco (HC) methods. We consider the applicability of Beer's law to carbon materials dispersed in dimethylformamide (DMF) and the effective extinction coefficients that are obtained from such dispersions. Analysis of the areal absorptivities of the second interband transition of semiconducting EA-produced SWNTs for a number of samples of differing purities has lead to an absolute molar extinction coefficient for the carbonaceous impurities in EA-produced SWNT samples. We conclude that NIR spectroscopy is the clear method of choice for the assessment of the bulk carbonaceous purity of EA-produced SWNTs, and we suggest that an absolute determination of the purity of SWNTs is within reach. Continued work in this area is expected to lead to a universal method for the assessment of the absolute bulk purity of SWNTs from all sources--such a development will be of great importance for nanotube science and for future customers for this product. PMID- 15656194 TI - Synthesis of single-walled carbon nanotubes (C-SWNTs) with a plasma torch: a parametric study. AB - In this study, we examine, in detail, the synthesis of single wall carbon nanotubes (C-SWNTs) with a plasma torch, using molecular sources for both carbon and catalyst (typically a carbon-containing gas such as ethylene and ferrocene for the metal catalyst). A comparison of the results obtained by Raman spectroscopy, using two different excitation wavelengths, permitted us to evaluate the importance of certain experimental parameters affecting the quality of the samples; these include the growth temperature of the nanotubes, and the temperature gradient between the flame and the oven. We have found that our method provides results that are qualitatively similar to those obtained using arc and laser techniques: C-SWNTs mixed with catalyst nanoparticles and amorphous carbon. Although the yields of C-SWNTs in the experiments reported here are somewhat lower than other approaches, our method avoids the inconveniences related to the solid phase of the initial material. Observations from scanning electron micrographs (SEM) and transmission electron micrographs (TEM) suggest that the mechanisms involved in the synthesis of the C-SWNTs by our plasma torch are similar to these latter techniques. An industrial-scale process based on a plasma torch could produce large amounts of C-SWNTs with good efficiency, since the present set-up produces continuously 1.5 g/h of deposit while using only 1 kW of power. PMID- 15656195 TI - Highly ordered carbon nanotubes based on porous aluminum oxide. AB - Highly ordered carbon nanotubes (CNTs) are widely pursued due to their unique properties. Anodic aluminum oxide (AAO) exhibits great possibility for this purpose. Here, CNTs based on AAO templates were produced using acetylene or ethylene as the hydrocarbon sources with or without the presence of Co catalysts. CNTs grown on the Co-embedded AAO samples were normally confined within the nanopores of the AAO template. It was found that C2H4 normally requires 100 degrees C higher pyrolysis temperature than C2H2 under otherwise identical conditions. The pyrolysis temperature is greatly reduced with the presence of Co catalysts. CNTs can grow out of the nanopores if Co particles are present at the bottom of the nanopores, and if the nanopores are short in length or large in diameter. The graphitization of AAO-template grown CNTs was studied by Raman spectroscopy. CNTs produced from ethylene are generally better in graphitization than those from acetylene, and CNTs grown with the presence of Co catalysts deposited at the bottom of nanopores are better than those without Co catalysts or with Co catalysts coated on the entire inner wall of nanopores. The growth temperature is found not to play a critical role in graphitization. PMID- 15656196 TI - Biodistribution of carbon single-wall carbon nanotubes in mice. AB - Carbon nanotubes are promising for use in biomedical and pharmaceutical sciences. Therefore, it becomes imperative to know the basic biological properties of carbon nanotubes in vivo. We labeled the water-soluble hydroxylated carbon single wall nanotubes with radioactive 125I atoms, and then the tracer was used to study the distribution of hydroxylated carbon single-wall nanotubes in mice. They moved easily among the compartments and tissues of the body, behaving as small active molecules though their apparent mean molecular weight is tremendously large. This study, for the first time, affords a quantitative analysis of carbon nanotubes accumulated in animal tissues. PMID- 15656197 TI - Visible upconversion emission of Pr3+ doped gadolinium gallium garnet nanocrystals. AB - The luminescence properties of a Pr3+-doped gadolinium gallium garnet (GGG, Gd3Ga5O12) nanocrystalline host were investigated. Dominant blue/green emission was observed emanating from the 3P0 --> 3H4 transition after excitation using a wavelength of 457.9 nm. Continuous wave excitation into the 1D2 level of the Pr3+ ion at 606.9 nm transition produced blue upconversion luminescence spectra, ascribed to emission from the 3P1 --> 3H4 and 3P0 --> 3H4 transitions. The increase in the decay times of the observed transitions following excitation with 606.9 nm is indicative of the dominance of an energy transfer upconversion (ETU) mechanism relative to excited state absorption (ESA). Furthermore, blue, green and red upconversion emission was observed from the 3P0, 3P1 and 1D2 states following excitation into the 1G4 energy level with 980 nm. No change in the decay times of the emitting states was observed following excitation with a wavelength of 980 or 457.9 nm; hence, upconversion was determined to primarily occur through ESA. The luminescence properties of the nanocrystals are compared to a single crystal of GGG:Pr3+ (bulk) with an identical Pr3+ concentration (1%). PMID- 15656198 TI - Cadmium sulfide and cadmium selenide/cadmium sulfide nanoparticles stabilized in water with poly(cysteine acrylamide). AB - Cysteine acrylamide (N-acryloyl L-cysteine) stabilizes CdS nanoparticles as the particles form in aqueous dispersions. Cysteine acrylamide also exchanges for citrate on the surfaces of CdSe and core/shell CdSe/CdS nanoparticles to provide greater stability. Heating of the nanoparticle dispersions polymerizes the cysteine acrylamide on the surface to form a more efficient polydentate stabilizer. The polymer-coated nanoparticle dispersions are colloidally stable even after removal of low molecular weight solutes by dialysis. Emission quantum yields of the polymer-coated CdSe and CdSe/CdS samples were 0.9% and 2.6%, respectively, after aging of the samples in light. CdSe/CdS coated with poly(cysteine acrylamide) is colloidally stable for at least two years in the dark at 5 degrees C. PMID- 15656199 TI - Synthesis of high purity silicon nanoparticles in a low pressure microwave reactor. AB - The formation of pure single crystalline silicon nanoparticles by microwave induced decomposition of silane in a low pressure flow reactor is reported. The morphology and crystal structure of the particles are characterized in situ by particle mass spectrometry (PMS) and ex situ by means of X-ray diffraction, high resolution transmission electron microscopy, electron energy loss spectroscopy, and infrared spectroscopy. The preparation method allows for the adjustment of the mean particle diameter in the range 6 nm < or = dPM < or = 11 nm by controlling the precursor concentration, gas pressure, and microwave power. Spectroscopic investigations reveal that the particles are single crystal silicon. The potential on n- or p-type doping is in progress. PMID- 15656200 TI - Synthesis and characterization of nanostructural hydrogel and template for CdS nanofibers. AB - Hydrogels based on beta-D-glucopyranoside-substituent thiosemicarbazide derivatives were synthesized and used as template to fabricate netlike CdS nanofibers. The self-assembling properties of the hydrogels and their effect on the synthesis of CdS nanostructure were studied. Fourier transform infrared spectra, UV-visible absorption spectra and X-ray diffraction revealed that pi-pi stacking, hydrogen bonds and interdigitated interactions between hydrophobic chains had influence on the formation of self-assembled lamellar hydrogel. Transmission electron microscopic and electron diffraction observation showed the formation of hexagonal stacking nanofiber-based CdS semiconductor materials. The transmission electron microscopic pictures of the CdS nanostructure in difference growth stages revealed the growth mechanism of CdS nanofibers templated by hydrogel. PMID- 15656201 TI - Synthesis and characterization of nanoporous silica using dendrimer molecules. AB - A water soluble porogen, e.g., poly(amido amine) dendrimer, has been used as a structure-directing agent to introduce porosity of nanometer scale in silica based nanocomposite materials. Hydrothermal synthesis was carried out at different elevated temperatures (343-413 K) in a closed teflon-lined stainless steel autoclave under autogenerated pressure. The synthesis time varied from 2 to 6 days, depending upon pH, synthesis temperature, concentration of porogen, etc. X-ray diffraction, transmission and scanning electron microscopic analyses, as well as infrared absorption spectroscopic measurements, were carried out to characterize these materials. Mostly disordered mesostructures were observed. The porous silica particles with sphere-like morphology varied from 30 to 200 nm in size were prepared depending upon the conditions of preparation. The structure and nanoporosity were preserved after the removal of structure-directing porogen through calcination at 823 K. The pore size was in the range of the dendrimer molecule as a template, indicating the structure-directing role of the terminal amino groups in the dendrimer molecule for the synthesis of nanoporous silica, since the nanoporous silica could be prepared by the hydrogen bonding or electrostatic interaction of dendrimer amino groups and tetraethyl orthosilicate. PMID- 15656202 TI - GMR effect in CuCo annealed melt-spun ribbons. AB - A thorough microstructural and magnetic analysis has been performed on as quenched and annealed (475 and 525 degrees C, 1 hour) melt-spun Cu100-xCox (x = 10 and 15) granular alloys, presenting a giant magnetoresistance (GMR) effect. The annealed samples are inhomogeneous with respect to the Co-particle sizes and interparticles distances and, therefore, these particles present superparamagnetic and ferromagnetic behaviours, which determine the GMR response. The samples x = 15, treated at 525 degrees C during 1 hour, presented the best GMR ratio (approximately 5% at room temperature to be the highest value approaching roughly to the saturation under an applied magnetic field of 15 KOe), with the coexistence of Co-particles with both kinds of magnetic behaviour. PMID- 15656203 TI - Synthesis of titanate nanofibers and nanotubes. AB - A simple, mild and low-cost refluxing method was successfully used to synthesize titanate nanofibers and nanotubes. Properties of the as-prepared samples were characterized by means of TEM and XRD techniques. Optimization of the experimental conditions allowed, respectively, the preparation of titanate nanofibers and nanotubes. The photocatalytic activities of the prepared products were also investigated. The results indicated that titanate nanofibers and nanotubes had no photocatalytic activities for the degradation of methyl orange. Calcined at different temperature, the photocatalytic activities of the products increased, accompanying the change of structures. PMID- 15656204 TI - Temperature-dependent phase stability of nanocrystalline SiO2. AB - Nanocrystalline powder of SiO2, synthesized by gas phase condensation, has been characterized by X-ray and electron diffractometric techniques. The composition of the as-received powder is mostly amorphous and a minor amount of alpha crystobalite phase. Annealing at higher temperatures, the amorphous phase gradually transforms into small crystallites of alpha-crystobalite. Subsequently, the tiny crystallites form small domains in the structural form of beta crystobalite. PMID- 15656205 TI - MgO nanowire growth from Mg metal and SiO2. AB - Comparative study on the diameter distribution of MgO nanowires has been carried out. MgO nanowires could be synthesized by the direct reaction between metallic magnesium and silica, and the obtained nanowires have diameters ranging from 50 to 200 nm and lengths of several hundreds nanometers, exhibiting a straight wire. The diameter can be downscaled to smaller than 50 nm, and the nanowire exhibits a curved and twisted one-dimensional structure with lengths up to several micrometers, when a fine support catalyst was used as the reactant. The diameter controlled growth mechanism was also explained in this work. PMID- 15656206 TI - Synthesis and unusual electron paramagnetic resonance spectrum of metastable nanoclusters of ZnO semiconductor crystallites. AB - Metastable nanoclusters of ZnO semiconductor crystallites, 20 to 30 nm diameter, are synthesized by a reconstructive decomposition of a polymer precursor of dispersed Zn2+ cations in poly vinyl alcohol (PVA) polymer molecules. They have EPR (electron paramagnetic resonance) spectrum of distinct excitonic features. Multiple EPR bands appear in prominent intensities in oxygen vacancies VO+ and singly ionized Oi- and Zn(i)+ interstitials. A paramagnetic VO+ vacancy derives from usual diamagnetic O2- vacancy of VO++ (behaves as if doubly charged compared to the lattice) by addition of one electron. The results demonstrate the existence of a surface-interface or surface barrier layer in free-carrier depletion at the crystallite surface in the clusters and its effects on the Oi- and Zn(i)+ ionization states (determine green photoluminescence). Both VO+ and Zn(i)+ are curable by a thermal annealing in O2 gas. A cured sample of equilibrium structure achieved by heating at approximately 550 degrees C has a single EPR in Oi- at g = 1.990. The results are useful in understanding their correlation with EPR and optical properties in ZnO semiconductors and devices. PMID- 15656207 TI - Electronic characteristics of Au-mercaptohexadecanoic acid-Au junction in a capillary. AB - The electrical property of a self-assembled monolayer (SAM) of mercaptohexadecanoic acid (HS-C15H30-COOH) has been investigated with a tunneling junction confined in a capillary. This capillary method can shun the interference of vacuum gap. The contact area can be determined according to the diameter of capillary. The nonlinear current-voltage curve and dl/dV curve are measured in the potential range of +/-0.8 V. The slight asymmetry of I-V curve and dl/dV curve is attributed to the different coupling between the metal and the end groups of molecule. This method will supply a simple way to measure the property of single molecules and monolayers. PMID- 15656208 TI - Influence of magnetic field on electromagnetic instabilities in semiconductor superlattices. AB - Electromagnetic instability criteria in semiconductor superlattices (SL) in the region of a negative differential conductivity (NDC) are studied in the presence of a static transverse field. The regimes with vertical constant electric field in a "short" plate of SL and with vertical current density vector in a "long" plate are considered. The effective differential conductivity tensors for both these regimes are derived and the influence of the magnetic field on the NDC threshold and the growth increment is investigated. The frequencies of the excited waves and the dependence of the growth increment on the direction of propagation for the extraordinary waves are determined. PMID- 15656209 TI - Theoretical studies of amidation reaction at carbon nanotube tips by means of the ONIOM technique: expanding the higher level. AB - In order to study how the choice of model chemistry can affect the results for amidation reaction of carboxylated SWNTs with methylamine, we attempted to expand the higher ONIOM level to six fused aromatic rings at the SWNT edge. The higher theoretical level was described with HF/STO-3G quantum mechanics, and the lower level with UFF universal force field. Introducing additional carboxylic, hydroxyl and quinone groups at the higher ONIOM level gives rise to the formation of new hydrogen bonds. The latter sometimes drastically change the overall geometry of reaction complexes, transition states and products, as well as their energies. In two of the three sets of SWNT models employed in the present study, the energetic preference for amide formation on armchair SWNT tips versus zigzag SWNT tips can be seen, although in all cases, the reaction turns endothermic. For the third set of models, including three hydroxyl and three carboxylic groups at the higher level, the amidation reaction is much more unfavorable for armchair SWNT. PMID- 15656210 TI - Prophylactic antibiotic administration prior to hysterectomy: a quality improvement initiative. AB - OBJECTIVE: To determine whether institution of a preoperative antibiotic policy could increase the use of prophylactic antibiotics prior to hysterectomy. STUDY DESIGN: A retrospective cohort study of 400 women who underwent abdominal, vaginal or laparoscopic hysterectomy for benign indications at Women and Infants Hospital was performed. Rates and timing of prophylactic preoperative antibiotic administration were determined, as were the rates of postoperative febrile morbidity. These data were compared to data collected in a medical record review of 686 hysterectomies performed prior to institution of the antibiotic policy. RESULTS: Prior to the institution of the antibiotic policy, 50% of patients (342/686, 95% CI 46.0, 53.7) received prophylactic preoperative antibiotics. After introduction of the antibiotic policy, 91.2% (95% CI 88.0, 93.8) of patients received prophylactic preoperative antibiotics. Approximately 66% of the antibiotics were administered within the 60 minutes preceding the surgical incision. Postoperative febrile morbidity was noted in 14% of patients prior to the antibiotic policy as compared to 11% of patients after the policy was instituted. Abdominal surgical approach was found to be a clinically and statistically significant risk factor for febrile morbidity (OR = 7.0; 95% CI 2.3, 20.9). CONCLUSION: Rates of prophylactic preoperative antibiotic administration significantly increased after institution of a hospital policy advocating routine antibiotic prophylaxis prior to hysterectomy. Additional steps must be taken to ensure more routine and appropriately timed administration of antibiotics prior to hysterectomy and to continuously monitor the use of prophylactic antibiotics. PMID- 15656211 TI - Short-term postoperative GnRH analogue or danazol treatment after conservative surgery for stage III or IV endometriosis before ovarian stimulation: a prospective, randomized study. AB - OBJECTIVE: To assess the effect of short-term use of a gonadotropin releasing hormone (GnRH) analogue for 3 months before ovarian stimulation in patients with stage III and IV endometriosis after conservative surgery. STUDY DESIGN: Eleven patients were randomly selected to receive intramuscular injections of GnRH analogue, leuprolide acetate (3.75 mg), every 28 days, or 400 mg danazol orally 2 times per day for 3 months before ovarian stimulation after conservative laparoscopic or laparotomy surgeryfor stage III and IV symptomatic endometriosis (group 1), as compared with 30 patients who had received no postoperative treatment with GnRH analogue or danazol but underwent ovarian stimulation immediately after thefirst menses within 3 months postoperatively (group 2). RESULTS: Although the number of oocytes retrieved and number of embryos per cycle were significantly higher in group 1, the pregnancy rate per cycle in group 1 was not significantly different from that in group 2 (18% vs. 20%). The cumulative pregnancy rate at 12 months was 54.5% and 56.7% in group 1 and group 2, respectively. With regard to recurrence of disease after 24 months of follow-up, group 2 had a statistically significantly higher recurrence rate (13.3%) than did group 1 (0%). CONCLUSION: Short-term use of GnRH analogue before ovarian stimulation in women with stage III or IV endometriosis confers no definite benefits on pregnancy rates per cycle when compared with patients who received ovarian stimulation within 3 months after conservative surgery. PMID- 15656212 TI - Early postpartum voiding dysfunction: incidence and correlation with obstetric parameters. AB - OBJECTIVE: To investigate the incidence and obstetric risk factors in early postpartum voiding dysfunction. STUDY DESIGN: Two hundred seventy-seven consecutive women in the early postpartum period were prospectively enrolled. Evaluation included history, physical examination, urinary questionnaire and non invasive uroflowmetry measurements. Patients were interviewed regarding the presence and severity of voiding symptoms and were categorized into symptomatic versus asymptomatic subgroups. Obstetric parameters were compared in symptomatic versus asymptomatic patients as well as in cases of decreased (< 15 mL/sec) versus normal maximum urinary flow rates. RESULTS: One hundred twenty-five women (45% of the study population) hadearly postpartum voiding difficulties. Symptoms were significantly more common after vacuum extraction (38%) than spontaneous vaginal delivery (27%) or cesarean section (15%). The second stage of labor was significantly prolonged in symptomatic versus asymptomatic patients (62+/-52 versus 47+/-48 minutes). Voided volume, maximum and average flow rates and continuous flow pattern were significantly decreased in symptomatic versus asymptomatic patients. Twenty-eight patients (10%) demonstrated decreased (<15 mL/sec) maximum flow rates. The first stage of labor was significantly prolonged in these patients (9.7+/-5.8 versus 6.4+/-3.9 hours). Twenty-five patients (13%) gave birth to a neonate weighing > or =3,800 g. Maximum flow rates among these women were significantly decreased as compared to women with smaller infants (25.6+/-11 versus 32.9+/-19.2 mL/sec, respectively). CONCLUSION: Approximately 50% of patients complained of voiding difficulties in the immediate postpartum period. Main risk factors were prolonged first and second stages of labor, vacuum extraction and birth weight > or = 3,800 g. Long-term follow-up is needed to determine the significance of this clinically common entity. PMID- 15656213 TI - HPV, cervical neoplasia and smoking: knowledge among colposcopy patients. AB - OBJECTIVE: To assess colposcopy patients' knowledge of human papillomavirus (HPV) infection and the role of smoking in cervical neoplasia and, if patients were current smokers, their willingness to quit smoking. STUDY DESIGN: Between January and June 2001, 250 women seen in a colposcopy clinic for the evaluation of an abnormal Pap smear or previously diagnosed cervical neoplasia participated in a survey designed to evaluate knowledge of HPV infection, smoking and neoplasia. Participants who smoked were questioned regarding their smoking behaviors and readiness to quit. In the analysis, the population was stratified by age (< 25 years versus > or = 25 years) to determine differences in knowledge, perception of risk and behaviors. RESULTS: The study population was in general young and racially/ethnically diverse. Fifty-seven percent (143 of 250) thought that HPV was a risk factor in cervical neoplasia, and 58% (146 of 250) associated smoking with neoplasia. Among smokers who discussed risk reduction with their providers, 75% (50 of 66) were advised to stop smoking. Of the 39% (98 of 250) who were current smokers, 63% (62 of 98) agreed that having an abnormal Pap smear would make them consider smoking cessation, and 58% (57 of 98) reported that they would use medical therapy (nicotine replacement or medication) to aid in that process. There were age-related differences in smoking behaviors and attitudes toward cessation (e.g., younger smokers were less likely to desire nicotine replacement). CONCLUSION: Given the high prevalence of smoking and a demonstrated desire by many of the colposcopy patients to stop smoking in the context of a cervical abnormality, further efforts at encouraging cessation are warranted. PMID- 15656214 TI - Obesity and in vitro fertilization: negative influences on outcome. AB - OBJECTIVE: To determine the influence of body mass index (BMI) on in vitro fertilization (IVF) outcome by studying a large cohort of good-prognosis IVF patients. STUDY DESIGN: This was a retrospective study at a university-based IVF center evaluating 920 good-prognosis patients (< 40 years, all with normal ovarian reserve) undergoing luteal leuprolide down-regulation and gonadotropin stimulation for IVF and evaluating IVF outcome, including response to stimulation, cancellation rates and pregnancy outcome based on obesity status. RESULTS: One hundred forty-eight (16.09%) patients were classified as obese (BMI > 27). Obese patients were more likely to undergo cancellation (14.9% vs. 9.1%, p = 0.03) when compared to nonobese patients. There was no difference in the amount of gonadotropins required. Obese patients had fewer oocytes retrieved and lower peak estradiol levels than did nonobese patients. In spite of the lower response, the clinical pregnancy rates per retrieval were no different (56.4% for both groups). CONCLUSION: Obese patients undergoing IVF are more likely to undergo cancellation. If cancellation does not occur, obesity confers a risk of a lower stimulation response. Despite this, the clinical pregnancy rates (per retrieval) were no different in obese patients and nonobese patients. PMID- 15656215 TI - NBME Obstetrics and Gynecology clerkship final examination scores: predictive value of standardized tests and demographic factors. AB - OBJECTIVE: To assess the predictive value of standardized tests and demographic factors on performance on the National Board of Medical Examiners (NBME) Obstetrics and Gynecology (OBGYN) examination. STUDY DESIGN: The study included 171 students who rotated through obstetrics and gynecology from 1992 to 2001. Correlations between NBME OBGYN scores and United States Medical Licensing Examination (USMLE) step 1, MCAT and GPA scores, and temporal and demographic factors were analyzed. RESULTS: The mean Medical College Admission Test (MCAT), NBME and USMLE step 1 scores were 24.03, 67.47 and 194.53, respectively. Significant correlations with NMBE OBGYN were USMLE step 1 scores (r = .517, p < 0.001), MCAT scores (r = .481, p < 0.001), faculty evaluation grade (r=.223, p <0.01), OBGYN residency interest (r= .179, p < 0.05) and female gender (r=.157, p< 0.05) with a significant negative correlation with clerkship duration (r = .208, p < 0. 01). On logistic regression analysis, USMLE, MCAT, academic year and faculty evaluation grade were independently predictive of NBME OBGYN scores. All preclinical standardized tests showed positive correlation with progressive academic years and decreasing student age. and decreasing student age. CONCLUSION: USMLE step 1, MCAT scores and faculty evaluation grades were predictive of NMBE OBGYN scores. They may be helpful in assessing third-year medical students in need of special supervision and assistance. PMID- 15656217 TI - Successful management of cervical pregnancy by selective uterine artery embolization: a case report. AB - BACKGROUND: Cervical pregnancy is potentially associated with life-threatening hemorrhage and often requires hysterectomy to stop the bleeding. Conservative management is becoming more common as treatment of cervical pregnancy. CASE: A case of cervical pregnancy presented with a severe vaginal hemorrhage at 6 weeks' gestation. The patient was conservatively managed with selective bilateral uterine artery embolization (UAE). The vaginal bleeding stopped completely after UAE. The clinical course was uneventful and serum human chorionic gonadotropin decreased immediately. The cervical mass gradually shrank and disappeared on the 31st day after UAE. UAE. CONCLUSION: This was the second case of cervical pregnancy successfully treated with only UAE. PMID- 15656216 TI - Premenarchal recurrent periclitoral abscess: a case report. AB - BACKGROUND: Periclitoral abscesses in premenarchal girls is a serious condition and can have negative implications for future reproductive health. Recurrent periclitoral abscess in premenarchal girls is rare. CASE: An 11-year-old, premenarchal girl presented complaining of dysuria, vulvar swelling, pain and erythema of the clitoral hood and a similar episode 12 months previously. Intravenous antibiotics and local measures resulted in spontaneous drainage of the abscess. On day 3 the area was healing, and the patient was discharged on oral antibiotics and sitz baths. At her 6-month follow-up, she had a normal clitoral hood without tenderness or erythema. CONCLUSION: Broad-spectrum antibiotics and local measures may prevent surgical treatment and potential associated reproductive morbidity. PMID- 15656218 TI - Spontaneous pregnancy in a woman with premature ovarian failure: a case report. AB - BACKGROUND: Women with premature ovarian failure often receive hormonal replacement therapy as part of the management strategy. However, it is very unusual for patients to become pregnant while on hormonal therapy. CASE: A 33 year-old woman with a history of infertility and secondary amenorrhea was diagnosed with premature ovarian failure. Part of her infertility workup included laparoscopy-hysteroscopy, a dye test with methylene blue, and endometrial and bilateral ovarian biopsy. Two months after completion of this procedure, the patient underwent sonography, which showed an intrauterine pregnancy at 14 weeks' gestation. The pregnancy continued to term, and a healthy infant was born. CONCLUSION: In most instances the etiology of premature ovarian failure is unknown. In rare cases this condition undergoes spontaneous, reversible remission. One interesting observation in our patient is that despite the performance of invasive diagnostic tests, there were minimal detrimental effects on the patient's pregnancy and infant. This case illustrates that the remote possibility of a spontaneous pregnancy in women with prema- ture ovarian failure should always be considered, and invasive diagnostic procedures should be performed when an intrauterine pregnancy is ruled out with certainty. PMID- 15656219 TI - Chronic ectopic pregnancy diagnosed incidentally in an infertile woman: a case report. AB - BACKGROUND: Chronic ectopic pregnancy is an enigma. The clinical presentation can be mild, with absent or subtle symptoms. The high incidence of negative pregnancy tests and the poor specificity of sonographic patterns can be misleading, and the correct diagnosis is sometimes established only at surgery or even histopathologically after the operation. We report the first case of a woman who was accidentally diagnosed with chronic ectopic pregnancy during diagnostic laparoscopy performed as part of a routine investigation for primary infertility. CASE: A 28-year-old woman underwent laparoscopyfor infertility. She had a regular menstrual cycle and was asymptomatic. She gave a history of a possible but unconfirmed miscarriage earlier. Her hormone profile was normal apart from a slightly raised prolactin level. An earlier ultrasound showed a polycystic appearance of the ovaries. Laparoscopy was done on the 25th day of the menstrual cycle, and beta-human chorionic gonadotropin was negative. At laparoscopy, a 2-cm mass wasfound in the right fallopian tube. There was no free blood in the pelvis, and no adhesions. Both tubes were patent at hydrotubation. The mass was excised laparoscopically, and histology confirmed a diagnosis of chronic ectopic pregnancy. CONCLUSION: A review of articles on chronic ectopic pregnancy confirmed the difficulty in diagnosing this condition preoperatively. PMID- 15656220 TI - The importance of community leadership. Taking a leadership role in serving the community is the responsibility of all. PMID- 15656221 TI - Enhanced patient understanding: taking steps to improve outcomes. PMID- 15656222 TI - Communication strategies for getting the results you want. PMID- 15656223 TI - Scarred but smarter: lessons learned from Florida's 2004 hurricanes. PMID- 15656224 TI - Responsibility for unsuccessful promotions. Inappropriate promotions result in ethical and personal costs to people and their organizations. PMID- 15656225 TI - Quicker turnaround appeases all customers. Physicians, patients, and management all benefit from a shorter preadmission testing process. PMID- 15656226 TI - Is there a doctor in the house? House call programs address the challenges of chronic care. PMID- 15656227 TI - Implications of the 2004 federal elections. Despite GOP control, significant healthcare policy change may be slow in coming. PMID- 15656228 TI - The importance of interpersonal skills. Emotional intelligence significantly impacts leadership success--and the bottom line. PMID- 15656229 TI - The pros and cons of board compensation. Does the promise justify the means? PMID- 15656230 TI - Leadership education for physicians. Refine your focus and communicate your goals when developing physician leadership programs. PMID- 15656231 TI - Overtime. PMID- 15656233 TI - My "turfdom" for a nurse! PMID- 15656232 TI - Journey to magnet designation: part 2. PMID- 15656234 TI - Back to the basics. PMID- 15656235 TI - [Evidence based surgery of cancer of head of pancreas]. AB - Physicians and surgeons who treat patients with gastrointestinal or hepatic disease must prescribe the most appropriate diagnostic tests, together with an accurate prognosis and effective and safe therapy. This paper examines the best modalities of surgical treatment for cancer of the pancreas, in an evidence-based approach. Evidence was classified as follows: Grade A : evidence from large randomized controlled trials (RCT) or systematic reviews (including meta analyses) of multiple randomized trials which collectively have at least as much data as one single well-defined trial. Grade B: evidence from at least one high quality study of non-randomized cohorts or evidence from at least one high quality case-control study or one high-quality case series. Grade C: opinions from experts without references or access to any of the foregoing The data were obtained from Medline and from controlled randomized trials listed in the Cochrane Library up to the end of 2003. Two series (grade B) showed the superiority of Whipple over total pancreatectomy, with respective median survival times of 12.6 months and 9.6 months. Extensive lymphadenectomy (grade A) in patients with positive lymph nodes gave significantly better survival than standard resection in one trial, but this was not confirmed in the other trial. Results of pylorus-preserving pancreaticoduodenectomy (PPPD) were not different from those of the Whipple procedure on postoperative mortality, morbidity or survival (grade A). Portal vein resection increased the resectability rate. Post operative mortality was not increased: survival was not different in four studies and was shorter in another four studies (grade C). Low-dose postoperative erythromycin accelerates gastric emptying if the right gastric artery is preserved (grade A). One trial suggests that pancreaticogastrostomy reduces the risk of pancreatic fistula. The two other trials are controversial and showed no difference. One prospective non randomized study showed that stenting in pancreaticojejunostomy reduces the risk of pancreatic fistulae and intraabdominal abscess. To prevent this risk of pancreatic fistula, six controlled trials involving patients receiving octreotride were performed Three European trials showed a smaller volume of abdominal drainage fluid and an abnormal amylase concentration; however, two American trials failed to demonstrate a significant difference. Occlusion of the pancreatic duct with fibrin glue did not reduce the risk of pancreatic fistula, but increased the risk of developing diabetes. Intraabdominal drainage after pancreatic resection significantly increased post operative complications (grade A). Surgical resection and reconstruction procedures for pancreatic cancer must be based on evidence-based studies. However, the most important prognostic factor is the surgeon's experience, not only with regard to the post-operative course, but also survival. Specific teaching and training is thus essential. PMID- 15656236 TI - [Orphan lung diseases: from curiosity to concern]. AB - As their name implies, orphan diseases are rare, often poorly studied diseases. Patients with orphan diseases are often deprived of adequate diagnosis and treatment. To promote a comprehensive study of orphan pulmonary diseases, we established a network of pulmonologists, allowing us to collect and study several series of patients. Based on the examples of lymphangioleiomyomatosis and idiopathic eosinophilic pneumonia, we emphasize the value of a global approach to orphan diseases within the context of a medical specialty. PMID- 15656237 TI - [Arterial complications after renal transplantation]. AB - We report our experience of surgical treatment of arterial complications after renal transplantation, in 154 patients operated on between 1970 and 2003. The arterial complication was discovered during a routine post-transplant check-up, or when the patient developed clinical manifestations (arterial hypertension, impaired renal function, arterial symptoms in the lower limbs), or following a septic complication. The following lesions were surgically managed: mycotic aneurysms (N = 5), atherosclerotic aneurysms (N = 13) or obstructions (N = 11) of the aorta and iliac arteries, and stenoses of the artery supplying the grafted kidney (N = 125). The type of arterial repair depended on the complication. There were no postoperative deaths. Three thromboses occurred after reconstruction of a stenotic transplant artery (3/125, 2.4 %), leading to graft loss. During long term follow-up, atherosclerotic lesions of the iliac arteries worsened below an aortic prosthesis in three patients who required repeated surgery. In two other patients, stenosis recurred on a transplant artery and required re-operation. In one patient, an aneurysm developed in a vein graft and was successfully replaced by an arterial autograft. Late deaths among atherosclerotic patients in this setting are usually due to cardiac causes. In all patients, late chronic rejection may lead to kidney loss and hemodialysis. PMID- 15656238 TI - [Dynamic MRI in the preoperative management of genital prolapse. A prospective study]. AB - Forty-three patients with genital prolapse prospectively underwent blinded pre and postoperative MRI, with dynamic assessment of each compartment. MRI was significantly more accurate than physical examination for the diagnosis of posterior defects (rectocele, enterocele) but not for anterior defects or uterine prolapse (hysterocele, cystocele). Postoperative MRI confirmed the surgical outcome and also identified risk factors for recurrence. MRI was particularly reliable for recurrent prolapse and vault prolapse. The authors consider that dynamic MRI can advantageously replace colpocystodefecography, as it is rapid and well tolerated. Technical improvements should help to understand the role of muscular and fascia lesions. PMID- 15656239 TI - [Emergency treatment of hand trauma in France, an important public health challenge]. AB - For decades hand trauma surgery was a small branch of general surgeons' practice. Only after the second world war did some orthopaedic or plastic surgeons with specific training in microsurgery start to sub-specialize in hand surgery. In the past 50 years, National Hand Societies have been created all over the world Hand trauma represents a significant proportion of emergency room attendance, and 20 25 % of traumatic emergencies. Most hand trauma patients are manual workers, who are sometimes injured at work. Hand injuries can result in lengthy periods off work. Incorrect emergency assessment and treatment by a non specialist surgeon can lead to poor outcomes. This can also increase the costs to society, owing to the need for further surgical procedures and physiotherapy. Hand trauma can be prevented by providing a safer environment at home, at work and in the playground Better trauma outcomes can be achieved by early referral to a Hand Center. In France, there are now almost 50 hand trauma centers. These are open 24 hours a day, 365 days a year, with a specialist hand surgeon on call. The latter will have diplomas in microsurgery and in hand and upper limb surgery, plus two years of experience as a fellow in a hand surgery unit. There is now a European Federation of Hand Trauma Centers (FESUM: Federation Europeenne des services d'Urgence-mains) with hand centers in Belgium, Italy, Germany, the UK, Luxembourg, Spain and Switzerland These work under the umbrella of the European Federation of Societies for Surgery of the Hand (FESSH). However, only a small percentage of hand trauma victims are referred to such a centers. Awareness of these centers must be improved among both health care professionals and the public. PMID- 15656240 TI - [The role for new evidence-based methods in patient care]. AB - Are we going to change the way we practice medicine? There are two kinds of medical knowledge: one is impersonal and results from research based on scientific methodology and scientific reasoning; the other is personal and results from learning and apprenticeship with seniors and from individual experience. Evidence-based medicine (EBM) is a new approach to teaching the practice of medicine. Instead of using personal knowledge, intuition, unsystematic clinical experience, and pathophysiological reasoning, it stresses evidence from clinical research. With this new paradigm in mind, clinicians should regularly consult the original literature, critically appraise the methods and the results, and then apply to the patient the optimal knowledge they have at their disposal. This new approach to medicine is gaining worldwide adoption, even if it more or less strictly interpreted. In France, even the translation of the English word "evidence " is controversial. There are two different translations for EBM. The most common is "Medecine fondee sur les preuves", and the other is "Medecine factuelle". The French word "Preuves" is the literal translation of "evidence" Some people argue against this term because its meaning is ambiguous for two reasons: --it does not express the full meaning of the English word "evidence", --there is a fear that the epistemological significance of "preuves" could be misinterpreted, giving birth to legal quibbles. Some people prefer the term "meilleures donnees acquises de la science". Other problems are linguistic and organizational. How can a busy French-speaking practitioner locate, retrieve and critically appraisal original articles, most of which are in English, unless he/she has a library containing the combined international medical literature? French meta-analyses, systematic reviews and books dedicated to evidence-based subjects are alternative options. Even when these practical problems can be solved, the proper place of research-based evidence must be clarified. When evidence is lacking, practitioners have to rely on other forms of medical reasoning for their decisions. Moreover, decision-making is not automatic: in addition to taking scientific knowledge into consideration, each medical decision is the result of a series of judgments based on human and environmental factors. PMID- 15656242 TI - [Advances in comparative pathology: some zoonoses threatening man]. AB - The last major human epidemics of infectious diseases have arisen from animals. Some of them are especially threatening. The authors call attention to the danger of spread of avian influenza, either directly or indirectly through genetic rearrangements. They underline the role of animals in the epidemiology of SARS, West Nile virus, hepatitis E, NIPA and Hendra virus, ehrlichiosis and Lyme disease. The authors recommend health surveillance not only in humans but also in animals; the teaching of zoonoses, and research on animal diseases transmissible to humans. PMID- 15656241 TI - [The role of the council of Europe in the field of health]. AB - To understand the actions of the Council of Europe in the field of human health, it is necessary first to understand the founding ideals, that is to say the creation in 1949 of a European institution capable of anticipating and avoiding differences and conflicts between the member states, whatever their standard of living. For the creation of status objectives, the Council has an appropriate working structure; the Committee of Ministers, the Parliamentary Assembly, non governmental organisations and the European Health Committee, which proposes, develops and oversees the effective application of health measures and social protection. Several hundred recommendations have thus been adopted by the Committee of Ministers. The first steps of this European Health Committee were in the areas of blood transfusion, immunohematology, transplantation and equal access to health services. The European Social Security Code and its Protocol, the Conventions on social and medical assistance, and many international agreements are effective treaties which bind Member States. PMID- 15656243 TI - Continuing the dialogue: a response to Kikuchi's critique of the 2002 CNA Code of Ethics. PMID- 15656244 TI - The Helen files. PMID- 15656245 TI - Challenges to developing and providing nursing leadership. PMID- 15656246 TI - Collaborative nursing education programs: challenges and issues. AB - Collaborative nursing education programs have been offered to facilitate access to baccalaureate-level nursing education. Our Collaborative Nursing Program involved 10 institutional partners and has been one of the largest of such programs. The collaborative approach to nursing education has been identified as an important model; the benefits include optimal use of resources and opportunities to develop and share knowledge across institutions. However, there has been little public discussion of the issues and challenges that emerge, including differing cultures, priorities, vulnerabilities, goals and aspirations between colleges and universities; desire to preserve autonomy and uniqueness; and complexity of approval and accreditation processes. Some of our college partners have chosen to offer an independent applied degree in nursing rather than continuing in a collaborative academic degree program. This paper describes the challenges inherent in maintaining quality of the degrees and strategies to increase the likelihood of continuing collaboration. Clarity and transparency are vital, and supportive programs involving mentorship of educators can foster increasing autonomy of colleges. Collaborative nursing programs pose many challenges, and their future will hinge on understanding mutual goals and expectations. PMID- 15656247 TI - Commentary: The case for national standards. PMID- 15656248 TI - Emotional intelligence: a primal dimension of nursing leadership? AB - Emotional intelligence, a concept well known in the world of business, is quickly gain ing recognition in the healthcare arena. Described as the primal aspect of leadership, emotional intelligence refers to the ability to monitor and discriminate among emotions, and to use the information to guide thought and action. This paper will explore the theoretical and empirical basis of emotional intelligence and its linkages to leadership. Readers will be informed of the origins and theoretical perspectives regarding emotional intelligence, evidence related to the existence of emotional intelligence, benefits and limitations of the concept and implications for nursing leadership. PMID- 15656249 TI - Setting the climate for evidence-based nursing practice: what is the leader's role? AB - Nurses are being challenged today to justify their practice. Many clinical and policy decisions in nursing are based upon isolated, ritualistic and unsystematic forms of clinical practice. The growing movement towards establishing evidence based nursing practice (EBNP) is situated in a systematic appraisal of the best evidence available. Nurse leaders have an obligation to cultivate sound clinical and economic practices leading to quality patient care and positive work life environments for nurses. PMID- 15656250 TI - Investing relational energy: the hallmark of resonant leadership. AB - Recent research has shown that hospital restructuring that included staff layoff has adversely affected the role, health and well-being of nurses who remained employed. Further research found that nurses working in environments that reflected resonant (emotionally intelligent) leadership reported the least negative effects to their healt and well-being following hospital restructuring. What remained unclear was the mechanism by which this mitigation occurred. The purpose of this paper is to explore additional findings from this leadership research and discuss one explanation unique to the academic literature for the mitigation variable--the investment of relational energy by resonant nursing leadership to build relationships with nurses and manage emotion in the workplace. PMID- 15656251 TI - Predictors of nurse managers' health in Canadian restructured healthcare settings. AB - Although nursing leadership roles have been greatly transformed as a result of dramatic changes within healthcare over the past decade, there is little research on the nature of nurse manager work life in current work environments. The purpose of this study was to test a theoretical model derived from Kanter's theory of organizational empowerment: linking nurse managers' perceptions of structural and psychological empowerment to burnout, job satisfaction and physical and mental health. A descriptive, correlational design was used in a sample of 286 first-line (n=202) and middle-level (n=84) hospital-based nurse managers obtained from a provincial registry. Ironically, managers reported high levels of burnout, but good mental and physical health. Middle managers were more empowered and satisfied with their jobs than first-line managers. In both groups, approximately 45% of the variance in job satisfaction and 18-52% of the variance in physical and mental health was explained by empowerment and burnout. Empowered work environments were associated with lower nurse manager burnout and better physical and mental health. The results suggest that creating work environments that provide access to empowerment structures may be a fruitful strategy for creating healthy work environments for nurse managers. PMID- 15656252 TI - Nanotechnology and the challenges it poses for the Federal Government. PMID- 15656253 TI - Application of current research evidence to clinical physiotherapy practice. AB - This paper reviews the nature of physiotherapy intervention studies published in core physiotherapy journals (Australian Journal of Physiotherapy, Physiotherapy Theory and Practice, Physical Therapy, Physiotherapy, and Physiotherapy Canada) between October 2001 and September 2002. The clinical applicability of their evidence was considered in light of the clinical relevance of diagnostic criteria used for subject recruitment, the nature of the interventions tested, and the outcome measures used to determine effectiveness of the intervention. Most studies investigated a "package of care" and used clinician-oriented measures of outcomes to determine the effectiveness of an intervention. This mirrors current clinical practice. However, few studies used tissue-based diagnostic criteria for subject recruitment, tested interventions within an episode-of-care model, or measured outcome from multiple stakeholders' perspectives. These findings highlight potential barriers for clinicians in the uptake and sustained application of research evidence in the clinical setting. Both clinical and research physiotherapists need to be involved in producing generalizable research findings to ensure that evidence-based practice can be widely and readily adopted. PMID- 15656254 TI - Comparing correlates for different types of absence versus lateness behaviors. AB - Using self-report measures of lateness and absence on pilot and main study samples, this study applied Dalton and Mesch's (1991) measurement approach to successfully "dissect" avoidable absence and avoidable lateness behaviors. Avoidable absence behavior was calculated by subtracting unavoidable absence from total absence, and avoidable lateness behavior was calculated by subtracting unavoidable lateness from total lateness. Record-based absence and lateness data, as well as test-retest self-reported absence and lateness data, collected on a subsample of pilot respondents, supported the validity and reliability of this approach. Main sample results included work exhaustion showing stronger positive relationships to absence versus lateness and career commitment exhibiting stronger negative relationships to avoidable lateness and avoidable absence. Two new specific work attitudes, punctuality and sick abuse, were partially successful in explaining lateness and absence behaviors. PMID- 15656255 TI - Allied health deans' and program directors' perspectives of specialized accreditation effectiveness and reform. AB - Criticisms, calls for change, and recommendations for specialized accreditation improvement have been made by individuals or groups external to the daily operations of allied health educational programs, frequently as opinion pieces or articles lacking a research foundation. While there is a great deal of concern related to specialized accreditation, little input has been provided from those within, and integral to, allied health educational programs affected by specialized accreditation standards. The purpose of this study was to explore the perspectives of selected allied health deans and program directors regarding specialized accreditation effectiveness and reform. Survey research was used to study perspectives of allied health deans and program directors located in four year colleges and universities and in academic health centers and medical schools. Surveys were mailed to program directors offering-programs in clinical laboratory sciences and medical technology, nuclear medicine technology, occupational therapy, physical therapy, radiation therapy, and radiography. Simultaneously, allied health deans located within these institutions were surveyed. A total of 773 surveys were mailed and 424 valid responses were received, yielding a response rate of 55%. The results affirmed the role of accreditation as an effective system for assuring quality in higher education. The role of specialized accreditation in improving the quality of allied health programs was clearly articulated by the respondents. Respondents voiced strong opposition to governmental or state-level requirements for accountability and emphasized the vital role of peer evaluators. Significant differences in deans' and program directors' perspectives related to specialized accreditation were evident. Whereas deans and program directors agreed with the purposes of specialized accreditation, they expressed less support for the process and effectiveness, and critique and reform, of specialized accreditation. Within those categories, deans were in greater support of critique and reform efforts related to specialized accreditation, while program directors were more supportive of the purpose, process, and effectiveness of specialized accreditation. PMID- 15656256 TI - Examining the presence of computer-assisted instruction in physical therapy education. AB - The purpose of this study was to compare computer-assisted instruction (CAI) use in physical therapist (PT) and physical therapist assistant (PTA) education programs. Secondary purposes included examining faculty attitudes toward CAI, selection criteria, and evaluation criteria. Questionnaires were mailed to 389 PT and PTA programs. The total return rate was 52% (N = 201). Frequencies, percentages, and modes of responses were calculated for each questionnaire item. Eight percent of PT and 35% of PTA programs indicated that all faculty are using CAI, whereas 3% of PT and 6% of PTA reported that no faculty are using CAI. PT program respondents indicated using nine different types of CAI, whereas PTA programs indicated using eight different types. Fifty-seven percent of PT and 33% of PTA programs provide formal computer instruction. Forty-six percent of PT and 47% of PTA programs have a computer prerequisite. Nine percent of PT and 0% of PTA programs have mandatory computer ownership or lease policies. Positive aspects reported included improved knowledge of technology and independence with information gathering. Negative aspects reported included cost and time. Respondents were unsure whether different students benefit more from CAI (high or low aptitude, learning style) Results indicated that criteria for selecting CAI material are based on cost, ease of use, and compatibility with current systems rather than instructional design, and evaluation procedures included student outcomes and course evaluations. Contextual differences between PT and PTA probably account for differences in types and uses of CAI for PT and PTA programs. Instructor feedback is important when using CAI to teach psychomotor skills. More research is needed to determine if one type of student benefits more than others. Programs should consider the instructional design of CAI materials, in addition to practical aspects such as cost, and ease of use. We need to be aware of the instructional goals, content, and learners when selecting, designing, and using CAI. PMID- 15656257 TI - Training physiotherapy students' abilities in scoring the motor assessment scale for stroke. AB - The purpose of this study was to test the ability of students in scoring an observational tool for measuring physical disability, the Motor Assessment Scale (MAS) for stroke, before and after training in order to establish whether current training and testing procedures for students are adequate. Sixteen students were tested using items 1-8 of the MAS before and after training. The students were required to observe the videotaped performance of four patients with stroke being assessed using the MAS and to score the performance of each patient. After training, the students were retested. The percent exact agreement between students' scores and criterion scores was calculated for each student and for each item. They were then compared before and after training using Student t test. When compared with criterion, the students achieved a mean of 79.6% (standard deviation, 10.8%) exact agreement before training and 83.9% (standard deviation, 6.7%) after training. While this increase in percent exact agreement was not statistically significant (p = 0.10), all students reached at least 75% exact agreement after training. Percent exact agreement on the items increased significantly (p = 0.03) so that seven of the eight items reached at least 75% exact agreement after training. The results of this study suggest that training is important to ensure that all students reach an acceptable standard. PMID- 15656258 TI - Priorities and benchmarks for new faculty in physical therapist education programs: perceptions of program directors. AB - The purpose of this study was to establish development priorities, ideal workload and performance levels, and preferred faculty development activities for new faculty in professional-level physical therapist education programs. A preliminary questionnaire was sent to 183 directors of these programs to identify new faculty and their program directors. Eighty-five new faculty and 79 of their program directors were identified and sent comparable surveys with questions that pertained to the current and ideal teaching, scholarship, and service activities of the new faculty. Results indicated that both new faculty and program directors agreed on ideal workload levels and that workloads for new faculty should shift from teaching to scholarship. New faculty set performance levels that are significantly higher than those identified by their program directors. Development priorities for the new faculty varied from knowledge and skills in instruction as expressed by program directors to scholarship as expressed by new faculty. Individual consultations were the highest preferred faculty development activity reported by both groups. These findings provide direction for the content and delivery method for faculty development activities for new faculty in these and similar education programs. PMID- 15656259 TI - Strategies used to increase lifestyle physical activity in a pedometer-based intervention. AB - The purpose of this study was to determine the self-selected strategies participants used to increase physical activity during an 8-week, pedometer-based lifestyle intervention. Participants were 34 employees of a small northeastern private college, with a mean age of 44.3 (+/- 9.3) years. The intervention consisted of a counseling session (goal setting, strategy selection), daily pedometer usage, and self-monitoring. Measures included height, weight, and pedometer-assessed ambulatory activity. Participants were placed in body mass index (BMI) and weekly physical activity improvement tertiles. BMI categories included normal weight (BMI 18.6-24.9), overweight (BMI 25-29.9), and obese (BMI > or =30). Weekly physical activity improvement categories included small/no improvement (<1%/wk), moderate improvement (1-5%/wk), and large improvement (>5%/wk). Data analysis consisted of descriptive and nonparametric inferential statistics (chi2). Participants in this study used 11 primary strategies on a regular basis to increase daily physical activity. The percentage of participants who used these strategies was as follows: walked to a meeting or work-related errand (64.7%), after work (50.0%), before work (35.3%), at lunch (47.1%), on the weekend (32.4%), while traveling (32.4%), with the dog (32.4%), or to a destination (work/store) (29.4%). Additionally, participants parked farther away (50%), used the stairs rather than an elevator (23.5%), and performed other cardiovascular activity (52.9%). Differences among BMI groups were found for the parking further strategy (p < 0.05), with obese participants using this strategy significantly more than overweight participants. No significant differences were found among weekly physical activity improvement groups. This study indicates that participants in a pedometer-based lifestyle physical activity intervention used a variety of strategies to increase daily physical activity. PMID- 15656260 TI - School-based health centers and academic performance: research, challenges, and recommendations. AB - School-based health centers (SBHCs) provide physical and mental health services on school campuses to improve student health status, and thereby potentially facilitate student academic success. With a growing emphasis on school accountability and the simultaneous dwindling of resources at the federal, state, and local levels, SBHCs face increasing pressures from school administrators and funders to document their impact on student academic achievement. This article reviews the methods, findings, and limitations of studies that have examined the relationship between SBHCs and academic performance. It also describes methodological challenges of conducting and interpreting such research, and discusses factors and intermediate variables that influence student academic performance. Recommendations are offered for SBHC researchers, evaluators, and service providers in response to the pressure they are facing to document the effect of SBHC services on academic outcomes. PMID- 15656261 TI - Implementing evidence-based substance use prevention curricula in North Carolina public school districts. AB - The Safe and Drug-Free Schools and Communities Act (SDFSCA) provides funding for prevention education to nearly every school district in the nation. Recent federal policy requires SDFSCA recipients to implement evidence-based prevention programs. This paper reports the extent to which North Carolina public school districts implement evidence-based substance use prevention curricula. Results showed that while the majority of school districts use evidence-based prevention curricula, they are rarely the most commonly used curricula. Evidence-based curricula are much more likely to be used at the middle school level than at the elementary or high school levels. Urbanicity, coordinator time, and coordinator experience correlated with extensive use of evidence-based curricula in the bivariate analysis, but only time spent on prevention by the Safe and Drug-Free Schools (SDFS) coordinator significantly predicted extensive use in the multivariate analysis. Increasing district SDFSCA coordinator time is a necessary step for diffusing evidence-based curricula. PMID- 15656262 TI - The School Health Portfolio System: a new tool for planning and evaluating coordinated school health programs. AB - The School Health Portfolio System (SHPS), developed originally to evaluate the Florida Coordinated School Health Program Pilot Schools Project, offers a new and innovative system for planning and evaluating a coordinated school health program at the individual school level. The SHPS provides practitioners a detailed but easy-to-use system that enables schools to create new programs or modify existing programs across all eight components of the CSHP model, as well as administrative support critical to sustainability. The System comes packaged as a self contained, notebook-style manual divided into 15 sections. It includes electronic templates of key documents to guide school teams in creating a customized portfolio, and a list of sample goals and artifacts that confirm achievement of a goal related to the school's coordinated school health program. An evaluation rubric provides a structured method to assess a program portfolio's contents, and the extent to which the contents document achievement of program goals. The rubric produces both a qualitative assessment, such as a narrative summary of program strengths and areas for improvement, and a quantitative assessment, such as a numerical score (0-100), letter grade (A-F), or 5-star system (*-*****). The physical structure, function, and scoring of the rubric depend on the method of assessment. The SHPS enables schools to set goals based on individual school needs, and incorporate CSHP goals into school improvement plans--a critical factor in sustainability and accountability. The System also offers teams the option of coordinating their efforts with CDC's School Health Index as a companion assessment measure. This article outlines the process a team would follow in developing a portfolio, and includes a sample assessment for the area of School Health Education. PMID- 15656263 TI - Knowledge and intentions of ninth-grade girls after a breast self-examination program. AB - Breast cancer ranks as the second-leading cause of death for women. Until recently the American Cancer Society advocated teaching breast self-examination (BSE) procedures. The Susan G. Komen Breast Cancer Foundation, active in the fight against breast cancer, continues to support educational efforts to teach breast self-examination skills to adolescent girls. Funded by the Komen Foundation, a BSE education program was developed and delivered by health educators at the Hult Health Education Center. The 50-minute program consisted of a PowerPoint slide presentation of breast cancer and BSE facts, and a video demonstration of proper BSE technique. Using a quasi-experimental design, the program was evaluated for changes in knowledge and intention to perform BSE. Ninth-grade girls at four public high schools (n = 255) in Peoria, Ill., were given a pretest to determine their knowledge about breast cancer and breast self examination. A delayed posttest was administered between five to six weeks later (n = 211). Using the Kruskal-Wallis nonparametric test, girls who participated in the BSE education program (treatment group) recorded a significantly higher overall mean knowledge score than girls who did not participate (comparison group). Those who participated in the program retained that knowledge over a five to six-week period. Analyses of variance revealed that more girls in the treatment group reported performing BSE in the past month as well as a higher intention to perform BSE in the future. PMID- 15656264 TI - Reliability and validity of a brief questionnaire to assess television viewing and computer use by middle school children. AB - Sedentary behaviors, like television viewing, are positively associated with overweight among young people. To monitor national health objectives for sedentary behaviors in young adolescents, this project developed and assessed the reliability and validity of a brief questionnaire to measure weekly television viewing, usual television viewing, and computer use by middle school children. Reliability and validity of the Youth Risk Behavior Survey (YRBS) question on weekday television viewing also were examined. A brief, five-item television and computer use questionnaire was completed twice by 245 middle school children with one week apart. To concurrently assess validity, students also completed television and computer use logs for seven days. Among all students, Spearman correlations for test-retest reliability for television viewing and computer use ranged from 0.55 to 0.68. Spearman correlations between the first questionnaire and the seven-day log produced the following results: YRBS question for weekday television viewing (0.46), weekend television viewing (0.37), average television viewing over the week (0.47), and computer use (0.39). Methods comparison analysis showed a mean difference (hours/week) between answers to questionnaire items and the log of -0.04 (1.70 standard deviation [SD]) hours for weekday television, -0.21 (2.54 SD) for weekend television, -0.09 (1.75 SD) for average television over the week, and 0.68 (1.26 SD) for computer use. The YRBS weekday television viewing question, and the newly developed questions to assess weekend television viewing, average television viewing, and computer use, produced adequate reliability and validity for surveillance of middle school students. PMID- 15656265 TI - School-based approaches to identifying students with asthma. PMID- 15656266 TI - From the search for a molecular code of memory to the role of neurotransmitters: a historical perspective. AB - The history of the neurochemistry of mnesic processes can be divided into two main periods: the first (1946-1978) was inspired by the results of molecular genetics, providing evidence for storage of hereditary information in the DNA of genes. Therefore, the chemical bases for memory were investigated in the macromolecules of the brain. Such attempts were relatively unsuccessful, which led to a second period (starting in 1978) with the research emphasizing, in a less ambitious way, the role of the molecular correlates of mnesic processes, in particular in the main transmitter systems of the brain. PMID- 15656267 TI - Pharmacological findings on the biochemical bases of memory processes: a general view. AB - We have advanced considerably in the past 2 to 3 years in understanding the molecular mechanisms of consolidation, retrieval, and extinction of memories, particularly of fear memory. This advance was mainly due to pharmacological studies in many laboratories using localized brain injections of molecularly specific substances. One area in which significant advances have been made is in understanding that many different brain structures are involved in different memories, and that often several brain regions are involved in processing the same memory. These regions can cooperate or compete with each other, depending on circumstances that are beginning to be identified quite clearly. Another aspect in which major advances were made was retrieval and post-retrieval events, especially extinction, pointing to new therapeutic approaches to fear-motivated mental disorders. PMID- 15656268 TI - Norepinephrine and dopamine as learning signals. AB - The present review focuses on the hypothesis that norepinephrine (NE) and dopamine (DA) act as learning signals. Both NE and DA are broadly distributed in areas concerned with the representation of the world and with the conjunction of sensory inputs and motor outputs. Both are released at times of novelty and uncertainty, providing plausible signal events for updating representations and associations. These catecholamines activate intracellular machinery postulated to serve as a memory-formation cascade. Yet, despite the plausibility of an NE and DA role in vertebrate learning and memory, most evidence that they provide a learning signal is circumstantial. The major weakness of the data available is the lack of a specific description of how the neural circuit modulated by NE or DA participates in the learning being analyzed. Identifying a conditioned stimuli (CS) representation would facilitate the identification of a learning signal role for NE or DA. Describing how the CS representation comes to relate to learned behavior, either through sensory-sensory associations, in which the CS acquires the motivational significance of reward or punishment, thus driving appropriate behavior, or through direct sensory-motor associations is necessary to identify how NE and DA participate in memory creation. As described here, evidence consistent with a direct learning signal role for NE and DA is seen in the changing of sensory circuits in odor preference learning (NE), defensive conditioning (NE), and auditory cortex remodeling in adult rats (DA). Evidence that NE and DA contribute to normal learning through unspecified mechanisms is extensive, but the details of that support role are lacking. PMID- 15656269 TI - Strains of rodents and the pharmacology of learning and memory. AB - Mendelian genetic tools have extensively been used to improve the description of the pharmacological mechanisms involved in learning and memory. The first part of this short review describes experiments involving the bidirectional selection of rats or mice for extreme behavioral characteristics or for sensitivity to pharmacological treatments. The second part focuses specifically on in-breeding. In conclusion, the advantages and the limits of a Mendelian pharmacogenetic approach of learning and memory are discussed. PMID- 15656270 TI - Genetic dissection of learning and memory in mice. AB - In this minireview, we discuss different strategies to dissect genetically the keystones of learning and memory. First, we broadly sketch the neurogenetic analysis of complex traits in mice. We then discuss two general strategies to find genes affecting learning and memory: candidate gene studies and whole genome searches. Next, we briefly review more recently developed techniques, such as microarrays and RNA interference. In addition, we focus on gene-environment interactions and endophenotypes. All sections are illustrated with examples from the learning and memory field, including a table summarizing the latest information about genes that have been shown to have effects on learning and memory. PMID- 15656271 TI - A chronic implant to record electroretinogram, visual evoked potentials and oscillatory potentials in awake, freely moving rats for pharmacological studies. AB - Electroretinogram (ERG), widely used to study the pharmacological effects of drugs in animal models (e.g., diabetic retinopathy), is usually recorded in anesthetized rats. We report here a novel simple method to obtain chronic implantation of electrodes for simultaneous recording at the retinal and cortical levels in freely moving, unanesthetized animals. We recorded cortical (VEPs) and retinal (ERGs) responses evoked by light (flash) stimuli in awake rats and compared the results in the same rats anesthetized with urethane (0.6 mg/kg) before and after the monocular administration of scopolamine methyl bromide (1 per thousand solution). We also compared the retinal responses with those derived from a classic acute corneal electrode. Anesthesia induced consistent changes of several VEP and ERG parameters like an increase of both latency and amplitude. In particular, the analysis of the variation of latency, amplitude, and spectral content of rapid oscillatory potentials could be important for a functional evaluation of the visual system in unanesthetized versus anesthetized animals. PMID- 15656273 TI - The modelling of positron emitter production and PET imaging during carbon ion therapy. AB - At the carbon ion therapy facility of GSI Darmstadt in-beam positron emission tomography (PET) is used for imaging the beta+-activity distributions which are produced via nuclear fragmentation reactions between the carbon ions and the atomic nuclei of the irradiated tissue. On the basis of these PET images the quality of the irradiation, i.e. the position of the field, the particle range in vivo and even local deviations between the planned and the applied dose distribution, can be evaluated. However, for such an evaluation the measured beta+-activity distributions have to be compared with those predicted from the treatment plan. The predictions are calculated as follows: a Monte Carlo event generator produces list mode data files of the same format as the PET scanner in order to be processed like the measured ones for tomographic reconstruction. The event generator models the whole chain from the interaction of the projectiles with the target, i.e. their stopping and nuclear reactions, the production and the decay of positron emitters, the motion of the positrons as well as the propagation and the detection of the annihilation photons. The steps of the modelling, the experimental validation and clinical implementation are presented. PMID- 15656272 TI - All about FAX: a Female Adult voXel phantom for Monte Carlo calculation in radiation protection dosimetry. AB - The International Commission on Radiological Protection (ICRP) has created a task group on dose calculations, which, among other objectives, should replace the currently used mathematical MIRD phantoms by voxel phantoms. Voxel phantoms are based on digital images recorded from scanning of real persons by computed tomography or magnetic resonance imaging (MRI). Compared to the mathematical MIRD phantoms, voxel phantoms are true to the natural representations of a human body. Connected to a radiation transport code, voxel phantoms serve as virtual humans for which equivalent dose to organs and tissues from exposure to ionizing radiation can be calculated. The principal database for the construction of the FAX (Female Adult voXel) phantom consisted of 151 CT images recorded from scanning of trunk and head of a female patient, whose body weight and height were close to the corresponding data recommended by the ICRP in Publication 89. All 22 organs and tissues at risk, except for the red bone marrow and the osteogenic cells on the endosteal surface of bone ('bone surface'), have been segmented manually with a technique recently developed at the Departamento de Energia Nuclear of the UFPE in Recife, Brazil. After segmentation the volumes of the organs and tissues have been adjusted to agree with the organ and tissue masses recommended by ICRP for the Reference Adult Female in Publication 89. Comparisons have been made with the organ and tissue masses of the mathematical EVA phantom, as well as with the corresponding data for other female voxel phantoms. The three dimensional matrix of the segmented images has eventually been connected to the EGS4 Monte Carlo code. Effective dose conversion coefficients have been calculated for exposures to photons, and compared to data determined for the mathematical MIRD-type phantoms, as well as for other voxel phantoms. PMID- 15656274 TI - Measurement of coherent x-ray scatter form factors for amorphous materials using diffractometers. AB - The feasibility of measuring the coherent x-ray scatter form factors of amorphous materials using powder diffractometers has been assessed. A five-step procedure was developed: (i) Low-angle background, consisting of a portion of the incident x-ray beam that passes directly to the detector, is measured using a specially designed replacement for the sample holder which absorbs most of the photons that otherwise would scatter off the sample holder cavity. (ii) Angle-dependent effects including monochromator efficiency and projected beam area are characterized by extracting the incoherent signal from the diffraction pattern of powdered graphite. The incoherent signal divided by the calculated incoherent cross section gives a correction factor as a function of scattering angle theta. (iii) Diffraction patterns are measured for the samples for theta = 2 degrees 150 degrees. (iv) The scattering data are corrected for background and then for angle-dependent effects. (v) The data are normalized to calculated free atom form factors at high theta, and the coherent form factor extracted. The method was implemented on two diffractometers at different energies (Co Kalpha and Cu Kalpha), and the results compared for water and plastics. Over the range 0.117 < x < 5.39 nm(-1), where x = lambda(-1) sin(theta/2), the average form factor ratio for water was 0.93. Systematic errors are difficult to eliminate. While this x ray powder diffractometer technique suffices for a survey measurement of the form factor of a material, its accuracy is probably insufficient for detailed studies. PMID- 15656276 TI - A new test phantom with different breast tissue compositions for image quality assessment in conventional and digital mammography. AB - Our objective is to describe a new test phantom that permits the objective assessment of image quality in conventional and digital mammography for different types of breast tissue. A test phantom, designed to represent a compressed breast, was made from tissue equivalent materials. Three separate regions, with different breast tissue compositions, are used to evaluate low and high contrast resolution, spatial resolution and image noise. The phantom was imaged over a range of kV using a Contour 2000 (Bennett) mammography unit with a Kodak MinR 2190-MinR L screen-film combination and a Senograph 2000D (General Electric) digital mammography unit. Objective image quality assessments for different breast tissue compositions were performed using the phantom for conventional and digital mammography. For a similar mean glandular dose (MGD), the digital system gives a significantly higher contrast-to-noise ratio (CNR) than the screen-film system for 100% glandular tissue. In conclusion, in mammography, a range of exposure conditions is used for imaging because of the different breast tissue compositions encountered clinically. Ideally, the patient dose-image quality relationship should be optimized over the range of exposure conditions. The test phantom presented in this work permits image quality parameters to be evaluated objectively for three different types of breast tissue. Thus, it is a useful tool for optimizing the patient dose-image quality relationship. PMID- 15656275 TI - A comparison of x-ray detectors for mouse CT imaging. AB - There is significant interest in using computed tomography (CT) for in vivo imaging applications in mouse models of disease. Most commercially available mouse x-ray CT scanners utilize a charge-coupled device (CCD) detector coupled via fibre optic taper to a phosphor screen. However, there has been little research to determine if this is the optimum detector for the specific task of in vivo mouse imaging. To investigate this issue, we have evaluated four detectors, including an amorphous selenium (a-Se) detector, an amorphous silicon (a-Si) detector with a gadolinium oxysulphide (GOS) screen, a CCD with a 3:1 fibre taper and a GOS screen, and a CCD with a 2:1 fibre taper and both GOS and thallium doped caesium iodide (CsI:Tl) screens. The detectors were evaluated by measuring the modulation transfer function (MTF), noise power spectrum (NPS), detective quantum efficiency (DQE), stability over multiple exposures, and noise in reconstructed CT images. The a-Se detector had the best MTF and the highest DQE (0.6 at 0 lp mm(-1)) but had the worst stability (45% reduction after 2000 exposure frames). The a-Si detector and the CCD with the 3:1 fibre, both of which used the GOS screen, had very similar performance with a DQE of approximately 0.30 at 0 lp mm(-1). For the CCD with the 2:1 fibre, the CsI:Tl screen resulted in a nearly two-fold improvement in DQE over the GOS screen (0.4 versus 0.24 at 0 lp mm(-1)). The CCDs both had the best stability, with less than a 1% change in pixel values over multiple exposures. The pixel values of the a-Si detector increased 5% over multiple exposures due to the effects of image lag. Despite the higher DQE of the a-Se detector, the reconstructed CT images acquired with the a Si detector had lower noise levels, likely due to the blurring effects from the phosphor screen. PMID- 15656277 TI - Comparing the synergistic effects of oleic acid and dimethyl sulfoxide as vehicles for optical clearing of skin tissue in vitro. AB - Recently, the tissue optical clearing technique has shown great potential in enhancing the capabilities of non-invasive light-based diagnostic and imaging techniques due to increased light penetration into tissue. In order to facilitate the clinical availability of the tissue optical clearing technique by the use of hyperosmotic agents, this study introduces oleic acid, a mono-unsaturated fatty acid which is generally recognised as safe, as an enhancer, and investigates the synergistic effect of oleic acid and propylene glycol on the optical clearing of skin tissue in vitro. Experimental results from near infrared spectroscopy, mass loss measurement and skin permeability assessment show that, when compared with dimethyl sulfoxide (DMSO) as an enhancer, oleic acid has a similar clearing effect. Due to its potential toxicity and the possible side effects, DMSO has been controversial in clinical applications. Therefore, in terms of optical application and clinic safety, the results presented reveal that oleic acid could be an optimal choice as an enhancer for optical clearing of skin tissue. PMID- 15656278 TI - Heat-transfer dynamics during cryogen spray cooling of substrate at different initial temperatures. AB - Cryogen spray cooling (CSC) is used to minimize the risk of epidermal damage during laser dermatologic therapy. However, the dominant mechanisms of heat transfer during the transient cooling process are incompletely understood. The objective of this study is to elucidate the physics of CSC by measuring the effect of initial substrate temperature (T0) on cooling dynamics. Cryogen was delivered by a straight-tube nozzle onto a skin phantom. A fast-response thermocouple was used to record the phantom temperature changes before, during and after the cryogen spray. Surface heat fluxes (q") and heat-transfer coefficients (h) were computed using an inverse heat conduction algorithm. The maximum surface heat flux (q"max) was observed to increase with T0. The surface temperature corresponding to q"max also increased with T0 but the latter has no significant effect on h. It is concluded that heat transfer between the cryogen spray and skin phantom remains in the nucleate boiling region even if T0 is 80 degrees C. PMID- 15656279 TI - Recombination factors for the cylindrical FC65-G ionization chamber in pulsed photon beams and the plane-parallel Roos ionization chamber in pulsed electron beams. AB - The use of ionization chambers in linac radiotherapy dosimetry requires various corrections to the measured charges, one of these being the recombination correction. The recombination correction factor (k(s)) is generally estimated from the two-voltage analysis (TVA) for each beam quality. However, it is possible that the ionization chamber above some threshold polarizing voltage does not follow the accepted Boag theory very well. Secondly the TVA is time-consuming as the chamber needs to stabilize after each polarizing voltage change and since it must be performed for each beam quality. Another approach consists in using the fact that k(s) is predicted to depend linearly on dose per pulse by Boag theory: determining this relationship once and for all using a multi-voltage analysis (MVA), one also checks the range validity of the Boag theory for the chamber considered. This work presents a thorough analysis of k(s) dependence on dose per pulse of FC65-G (cylindrical) and Roos (plane-parallel) ionization chambers in pulsed photon and electron beams, respectively. Within the uncertainties, the recombination factors are found to be independent of beam quality, and no deviation from the Boag theory is observed within the tested range of polarizing voltages. Before adapting the equations given using the MVA other users should check that their ionization chambers show the same dose per pulse dependence using the TVA for a few beam qualities. PMID- 15656280 TI - Simultaneous minimization of leaf travel distance and tongue-and-groove effect for segmental intensity-modulated radiation therapy. AB - This paper introduces a method to simultaneously minimize the leaf travel distance and the tongue-and-groove effect for IMRT leaf sequences to be delivered in segmental mode. The basic idea is to add a large enough number of openings through cutting or splitting existing openings for those leaf pairs with openings fewer than the number of segments so that all leaf pairs have the same number of openings. The cutting positions are optimally determined with a simulated annealing technique called adaptive simulated annealing. The optimization goal is set to minimize the weighted summation of the leaf travel distance and tongue-and groove effect. Its performance was evaluated with 19 beams from three clinical cases; one brain, one head-and-neck and one prostate case. The results show that it can reduce the leaf travel distance and (or) tongue-and-groove effect; the reduction of the leaf travel distance reaches its maximum of about 50% when minimized alone; the reduction of the tongue-and-groove reaches its maximum of about 70% when minimized alone. The maximum reduction in the leaf travel distance translates to a 1 to 2 min reduction in treatment delivery time per fraction, depending on leaf speed. If the method is implemented clinically, it could result in significant savings in treatment delivery time, and also result in significant reduction in the wear-and-tear of MLC mechanics. PMID- 15656281 TI - The impact of inter-fraction dose variations on biological equivalent dose (BED): the concept of equivalent constant dose. AB - Inter-fraction dose fluctuations, which appear as a result of setup errors, organ motion and treatment machine output variations, may influence the radiobiological effect of the treatment even when the total delivered physical dose remains constant. The effect of these inter-fraction dose fluctuations on the biological effective dose (BED) has been investigated. Analytical expressions for the BED accounting for the dose fluctuations have been derived. The concept of biological effective constant dose (BECD) has been introduced. The equivalent constant dose (ECD), representing the constant physical dose that provides the same cell survival fraction as the fluctuating dose, has also been introduced. The dose fluctuations with Gaussian as well as exponential probability density functions were investigated. The values of BECD and ECD calculated analytically were compared with those derived from Monte Carlo modelling. The agreement between Monte Carlo modelled and analytical values was excellent (within 1%) for a range of dose standard deviations (0-100% of the dose) and the number of fractions (2 to 37) used in the comparison. The ECDs have also been calculated for conventional radiotherapy fields. The analytical expression for the BECD shows that BECD increases linearly with the variance of the dose. The effect is relatively small, and in the flat regions of the field it results in less than 1% increase of ECD. In the penumbra region of the 6 MV single radiotherapy beam the ECD exceeded the physical dose by up to 35%, when the standard deviation of combined patient setup/organ motion uncertainty was 5 mm. Equivalently, the ECD field was approximately 2 mm wider than the physical dose field. The difference between ECD and the physical dose is greater for normal tissues than for tumours. PMID- 15656282 TI - Tracking errors in a prototype real-time tumour tracking system. AB - In motion-compensated radiation therapy, radio-opaque markers can be implanted in or near a tumour and tracked in real-time using fluoroscopic imaging. Tracking these implanted markers gives highly accurate position information, except when tracking fails due to poor or ambiguous imaging conditions. This study investigates methods for automatic detection of tracking errors, and assesses the frequency and impact of tracking errors on treatments using the prototype real time tumour tracking system. We investigated four indicators for automatic detection of tracking errors, and found that the distance between corresponding rays was most effective. We also found that tracking errors cause a loss of gating efficiency of between 7.6 and 10.2%. The incidence of treatment beam delivery during tracking errors was estimated at between 0.8% and 1.25%. PMID- 15656283 TI - A finite state model for respiratory motion analysis in image guided radiation therapy. AB - Effective image guided radiation treatment of a moving tumour requires adequate information on respiratory motion characteristics. For margin expansion, beam tracking and respiratory gating, the tumour motion must be quantified for pretreatment planning and monitored on-line. We propose a finite state model for respiratory motion analysis that captures our natural understanding of breathing stages. In this model, a regular breathing cycle is represented by three line segments, exhale, end-of-exhale and inhale, while abnormal breathing is represented by an irregular breathing state. In addition, we describe an on-line implementation of this model in one dimension. We found this model can accurately characterize a wide variety of patient breathing patterns. This model was used to describe the respiratory motion for 23 patients with peak-to-peak motion greater than 7 mm. The average root mean square error over all patients was less than 1 mm and no patient has an error worse than 1.5 mm. Our model provides a convenient tool to quantify respiratory motion characteristics, such as patterns of frequency changes and amplitude changes, and can be applied to internal or external motion, including internal tumour position, abdominal surface, diaphragm, spirometry and other surrogates. PMID- 15656284 TI - A novel four-dimensional image registration method for radionuclide therapy dosimetry. AB - A novel method for registering sequential SPECT scans (4DRRT) is described, whereby all sequential scans acquired in the course of a therapy or a pre-therapy tracer study may be registered in one pass. The method assumes that a monoexponential decay function can be fitted to the series of sequential SPECT scans. Multiple volumes, presenting with different decay rates, are fitted with different mono-exponential functions. The MSSE (mean sum of squared errors in the least-squares fit algorithm), over the volume used for registration, is the cost function minimized at registration. Simulated data were used to assess the effect of thresholding, smoothing, noise and the multi-exponential nature of the four dimensional (4D) SPECT studies on the performance of 4DRRT, resulting in three dimensional (3D) residual registration errors <3.5 mm. The 4DRRT method was then compared to the following 3D registration methods: the correlation coefficient, the sum of absolute differences, the variance of image ratios and the mutual information. The comparisons, using both simulated and clinical data, were based on the standard deviation of the effective decay time distribution, generated from the registered 4D dataset, and showed that image registration using 4DRRT is simpler and more robust compared to the 3D techniques, especially when multiple tumour sites with different decay rates are present. PMID- 15656285 TI - Accuracy of rigid CT-FDG-PET image registration of the liver. AB - Diagnostic and surgical strategies could benefit from accurate localization of liver malignancies via CT-FDG-PET image registration. However, registration uncertainty occurs due to protocol differences in data-acquisition, the limited spatial resolution of positron emission tomography (PET) and the low uptake of 18F-fluorodeoxyglucose (FDG) in normal liver tissue. To assess this uncertainty, methods were presented to estimate registration precision and systematic bias. A semi-automatic, organ-focused method was investigated to minimize the uncertainty well beyond the typical uncertainty of 5-10 mm obtained by commonly available methods. By restricting registration to the liver region and by isolating the liver on computed tomography (CT) from surrounding structures using a thresholding technique, registration was achieved using the mutual information based method as implemented in insight toolkit (ITK). CT and FDG-PET images of 10 patients with liver metastases were registered rigidly a number of times. Results of the organ-focused method were compared to results of three commonly available methods (a manual, a landmark-based and a 'standard' mutual information-based method) where no dedicated image processing was performed. The proposed method outperformed the other methods with a precision (mean+/-s.d.) of 2.5+/-1.3 mm and a bias of 1.9 mm with a 95% CI of [1.0, 2.8] mm. Unlike the commonly available methods, our approach allows for robust CT-FDG-PET registration of the liver, with an accuracy better than the spatial resolution of the PET scanner that was used. PMID- 15656286 TI - Tracing the neuroanatomical profiles of reward pathways with markers of neuronal activation. AB - Functional neuroanatomical tools have played an important role in proposing which structures underlie brain stimulation reward circuitry. This review focuses on studies employing metabolic markers of neuronal and glial activation, including 2 deoxyglucose, cytochrome oxidase, and glycogen phosphorylase, and a marker of cellular activation, the immediate early gene c-fos. The principles underlying each method, their application to the study of brain stimulation reward, and their strengths and limitations are described. The usefulness of this strategy in identifying candidate structures, and the degree of overlap in the patterns of activation arising from different markers is addressed in detail. How these data have contributed to an understanding of the organization of reward circuitry and directed our thinking towards an alternative framework of neuronal arrangement is discussed in the final section. PMID- 15656287 TI - Interference and time: a brief review and an integration. AB - Contemporary theories of associative learning have been preoccupied with the phenomenon of stimulus competition (attenuated responding to a target stimulus outcome association if another stimulus trained together with the target is a better or more reliable predictor of the outcome). In recent years, reports of associative interference between associations trained separately have challenged associative learning theories, which provide no mechanism to account for this type of interference. Moreover, the ever-growing reports of temporal relationships between stimuli being a variable that determines the occurrence of stimulus competition and associative interference call for a reformulation of associative theory that can account for both interference and temporal learning effects. Here, we briefly review some of the central findings in stimulus competition, associative interference, and temporal learning, as well as a recently-proposed integration of these three seemingly disparate families of phenomena. PMID- 15656288 TI - Timed executive functions and white matter in aging with and without cardiovascular risk factors. AB - The present article reviews recent studies of 'healthy aging' with and without cardiovascular risk factors and accompanying white matter changes. Specifically, the relationship between white matter hyperintensities (WMH) and various cognitive functions, in particular executive functions (EF), was subjected to meta-analysis. The results of this analysis indicate that WMH induce a decrease in performance on timed EF tests, while the more basic cognitive abilities (e.g. short-term memory and vocabulary) remained relatively intact. Interestingly, the performance on untimed EF tasks was uniformly unrelated to WMH. This pattern of cognitive dysfunction was observed equally in the 'healthy aged' and the cardiovascular group. These findings suggest that an important distinction exists between untimed and timed EF tests, and that this distinction should be taken into account when interpreting the results of such tests. PMID- 15656289 TI - The SBR and its biofilm application potentials. AB - Twenty plus years of experience, innovation, and research in the field of biological wastewater treatment and biofilm applications lead to the conclusion that biofilms are in many cases more desirable in reactors than suspended activated sludge. Biofilm reactors can provide very long biomass residence times even when the hydraulic influent loading is low. This makes them particularly suitable when treatment requires slow growing organisms with poor biomass yield or when the wastewater concentration is too low to support growth of activated sludge flocs. Regardless of the settling characteristics of biological aggregates or the hydraulic influent loading the metabolic activity in the reactor can be maintained at a high level. This paper reviews the application of biofilms in sequencing batch reactor (SBR) systems to treat non-readily biodegradable substrates, volatile organic waste constituents, complex waste streams requiring co-metabolism, and particulate wastewaters. Recent research using the SBR to form aerobic granular sludge as a special application of biofilms is also discussed. PMID- 15656290 TI - Combining SBR systems for chemical and biological treatment: the destruction of the nerve agent VX. AB - The US Army is pilot testing the neutralization of VX nerve agent stockpiled at Newport, Indiana using caustic hydrolysis in a Sequencing Batch Reactor (SBR). The resulting hydrolysate was tested at the bench-scale for treatment with activated sludge biodegradation in two distinct studies, one in the SBR and another, in the PACT process. The feed to both biological systems was pretreated to enhance the biodegradability of the hydrolysis products. Both biodegradation studies demonstrated that the hydrolysate could easily meet the Chemical Weapons Convention treaty and US environmental regulations following pretreatment. PMID- 15656291 TI - Nitrogen removal from sludge digester liquids by nitrification/denitrification or partial nitritation/anammox: environmental and economical considerations. AB - In wastewater treatment plants with anaerobic sludge digestion, 15-20% of the nitrogen load is recirculated to the main stream with the return liquors from dewatering. Separate treatment of this ammonium-rich digester supernatant significantly reduces the nitrogen load of the activated sludge system. Two biological applications are considered for nitrogen elimination: (i) classical autotrophic nitrification/heterotrophic denitrification and (ii) partial nitritation/autotrophic anaerobic ammonium oxidation (anammox). With both applications 85-90% nitrogen removal can be achieved, but there are considerable differences in terms of sustainability and costs. The final gaseous products for heterotrophic denitrification are generally not measured and are assumed to be nitrogen gas (N2). However, significant nitrous oxide (N2O) production can occur at elevated nitrite concentrations in the reactor. Denitrification via nitrite instead of nitrate has been promoted in recent years in order to reduce the oxygen and the organic carbon requirements. Obviously this "achievement" turns out to be rather disadvantageous from an overall environmental point of view. On the other hand no unfavorable intermediates are emitted during anaerobic ammonium oxidation. A cost estimate for both applications demonstrates that partial nitritation/anammox is also more economical than classical nitrification/denitrification. Therefore autotrophic nitrogen elimination should be used in future to treat ammonium-rich sludge liquors. PMID- 15656292 TI - Nitrogen removal of high strength wastewater via nitritation/denitritation using a sequencing batch reactor. AB - The sequencing batch reactor (SBR) process concept was applied to achieve efficient ammonium removal via nitrite under both laboratory and pilot-scale conditions. Both sets of experimental results show that without pH control or carbon addition the nitritation process consistently converted approximately 50% of the ammonium from biosolids dewatering liquids to nitrite with hydraulic retention times (HRT) as short as 10 h. The results from the pilot-scale study also indicate that the selective oxidation of ammonium to nitrite is a reliable process as the accumulation of nitrate was never an issue during a 330-day trial. The SBR process concept was extended to achieve complete nitrogen removal through nitritation and denitritation in the laboratory scale. The experimental results indicate that a total reduction of 96-98% of the ammonium nitrogen from biosolids dewatering liquids (influent concentration typically 1,200 g m(-3)) was achieved with a short HRT of 1.1 d and a removal rate of 1.05 kgNm(-3)d(-1). This process concept was tested at pilot scale where the nitritation process could be started up without temperature control in a short period of time. Nitrogen removal rates up to 1.2 kgNm(-3)d(-1) at an HRT of 0.88 d have been obtained. COD to nitrogen ratios required in the pilot plant were consistently in the range 1.6-1.9 kgCOD kg(-1)N removed. PMID- 15656293 TI - Nitrite accumulation by aeration controlled in sequencing batch reactors treating domestic wastewater. AB - The feasibility of obtaining and keeping stable nitrite accumulation in Sequencing Batch Reactors (SBRs) treating domestic wastewater is studied. The final product of ammonium oxidation is either reproducible nitrate or nitrite depending on the aeration strategy. With the aerobic-anoxic sequence, two SBRs fed with domestic wastewater are operated in parallel. One SBR (SBR1) is controlled by the aeration control strategy, and the other SBR (SBR2) by alternate aeration control strategy. Based on the on-line indirect measurements of DO and pH, the relationship between pH (or DO) and nitrogen concentration (NH4+-N, NO(3-)-N and NO(2-)-N) is investigated. The result indicates that pH and DO can be used as control parameters for the real-time aeration control strategy to obtain nitritation in SBR treating domestic wastewater. The result of SBR1 indicates that long-term stable nitritation is possible at 32+/-1 degrees C. The result of SBR2 indicates that the aeration control strategy is necessary for nitritation during the acclimation period, because the nitrite accumulation disappears when the aeration is extended. PMID- 15656294 TI - Automatic control strategy for biological nitrogen removal of low C/N wastewater in a sequencing batch reactor. AB - To establish an automatic control system of external carbon addition in biological nitrogen removal, a bench-scale sequencing batch reactor with real time control strategy was designed in this study. An oxidation-reduction potential (ORP) profile was used for automatic control of external carbon addition. The mean removal efficiency of total organic carbon was over 98%. Complete denitrification in an anoxic phase and complete denitrification and nitrification in anoxic and oxic phases were accomplished, respectively, because the oxic and anoxic periods were also appropriately controlled with ORP and pH profiles, respectively. Mean removal efficiency of total nitrogen was over 95%. When concentration of influent wastewater was changed, volume of additional carbon was automatically changed with the influent fluctuation, and flexible hydraulic retention time was achieved in this system. PMID- 15656295 TI - Nitrogen removal from sludge water with SBR process: start-up of a full-scale plant in the municipal wastewater treatment plant at Ingolstadt, Germany. AB - The sludge water obtained from the dewatering processes following anaerobic sludge digestion contains high levels of ammonia. This sludge water is generally returned to the beginning of the waste water treatment plant process, thereby significantly increasing the nitrogen load on the biological process. In this project, the start-up of a full-scale sequencing batch reactor (SBR) process to separately treat the aforementioned sludge water is studied. Two parallel SBRs were operated over 8 hour cycles. The duration of the start-up was approximately 100 days until a hydraulic load of 225 m3/d was reached for each SBR. This paper presents the results of the start-up, highlighting the change in nitrogen concentration with time and the effect of other parameters such as temperature and suspended solids in that period. Following the project period of operation, the ammonium concentration was reduced by more than 95% on average. PMID- 15656296 TI - Performance evaluation of hybrid and conventional sequencing batch reactor and continuous processes. AB - Bench-scale systems, using conventional and compact hybrid activated sludge configurations, were set up to evaluate the systems' nitrification denitrification performance, operating sludge age/MLSS concentration and sludge settleability at a Hong Kong municipal STW. Configurations tested were the continuous clarifier modified Ludzack Ettinger (MLE) and the sequencing batch reactor (SBR) with and without hybrid suspended biofilm carriers. Results demonstrated that the hybrid SBR and MLE systems consistently achieved close to complete nitrification (effluent NH4-N = 2.4 and 6.9 mg/L) and 75% and 67% removal of nitrogen (N) (effluent NO3-N < 10 mg/L) with an overall hydraulic retention time of only 7.5 hours, operating sludge age as short as 5.2 days, and mixed liquor suspended solids concentration of approximately 1,300 mg/L with a sludge volume index of 109 and 229 mL/g, respectively. The most sensitive and slowest growing nitrifiers attached to the hybrid biofilm carriers. This allowed the hybrid processes to be operated at a sludge age shorter than the critical nitrifying sludge age while still retaining near complete nitrification. In contrast, to achieve complete nitrification, the conventional MLE system needed to be operated at 1.5 to 2.5 times the critical sludge age. These results indicate that the hybrid MLE configuration is a suitable process for use in upgrading existing conventional works for N removal and for increasing hydraulic capacity of existing N removal works, without major civil works modifications, in Hong Kong. For new works, consideration might be given to the use of the hybrid SBR, which shows a more stable N removal performance than the MLE process due to its inherent in-basin equalization capacity and better reaction conditions for nitrification in terms of higher initial NH4-N level. It was also observed that the conventional SBR produced better nitrification performance than the hybrid MLE process tested. Design parameters and operating conditions of the hybrid systems should be subjected to further full-scale trial for higher hydraulic capacity and N removal performance. PMID- 15656297 TI - Biological denitrification in a sequencing batch reactor. AB - Biological denitrification was studied in an SBR to investigate the performance. The experimental results showed that granular sludge with high activity and good settling ability can be developed smoothly in thereactor. With the granular sludge, high nitrate loading rate (0.48 kg NO3-N dm(-3) d(-1)) and removal efficiency (more than 99%) can be achieved. Scanning electronic microscope examination showed that only rod-shape bacteria existed on and within the granules, which confirmed indirectly that filamentous bacteria do not utilize nitrate and nitrite as electron acceptors. SBR is an alternative and effective technology for biological denitrification. PMID- 15656298 TI - On-line monitoring for control of a pilot-scale sequencing batch reactor using a submersible UV/VIS spectrometer. AB - A submersible UV/VIS spectrometer was used to monitor a pilot-scale sequencing batch reactor (SBR). The instrument utilises the whole UV/VIS range between 200 and 750 nm. With just one single instrument nitrate, organic matter and suspended solids can be measured simultaneously. The spectrometer is installed directly in the reactor, measures in real-time, and is equipped with an auto-cleaning system using pressured air. The paper shows the calibration results for measurements in the SBR tank, time series for typical SBR cycles, and proposes possible ways for optimisation of the operation by using these measurements. PMID- 15656299 TI - Rate of nitrate production during a two-stage nitrification batch reaction. AB - The two steps of nitrification, namely the oxidation of ammonia to nitrite and nitrite to nitrate, often need to be considered separately in process studies. It has been assumed that these two reactions can be described by single Monod models. In this paper, the suitability of the single Monod model for describing nitrite oxidation to nitrate is discussed. The measured rate of nitrate production during a batch reaction is presented. For the system studied it was found that nitrate production actually increased after the completion of ammonia oxidation. It is suggested that the reason for the increase was a combination of: (i) likely competition for oxygen when both substrates were present, and (ii) a decrease in ammonia inhibition of nitrite oxidisers with the removal of ammonia. The result is that a single Monod expression (based on nitrite as the substrate) could not be used to describe nitrate production. In these types of systems the consequence of oxygen limitation and substrate inhibition should also be considered. PMID- 15656301 TI - Optimal but robust N and P removal in Sbrs: a model-based systematic study of operation scenarios. AB - A systematic approach to determine the optimal operation strategy for nitrogen (N) and phosphorus (P) removal of sequencing batch reactors (SBRs) has been developed and applied successfully to a lab-scale SBR. The methodology developed is based on using a grid of possible scenarios to simulate the effect of the key degrees of freedom in the SBR system. The grid of scenarios is simulated using a calibrated ASM2dN model developed and calibrated in a previous study. Effluent quality in combination with a robustness index for each of the scenarios is used to select the best scenario. With the best scenario, it is possible to improve/increase the current performance of the SBR system by around 54% and 74% for N and P removal respectively. PMID- 15656300 TI - Wastewater nitrogen removal in Sbrs, applying a step-feed strategy: from lab scale to pilot-plant operation. AB - One of the problems of nitrogen removal from wastewater when applying sequencing batch reactor (SBR) technology, is the specific use of organic matter for denitrification purposes. Since easily biodegradable organic matter is rapidly consumed under aerobic or anoxic conditions (i.e. aerobic oxidation or anoxic denitrification, respectively), it is an important factor to consider when scaling up SBRs from the laboratory to real plant operation. In this paper, we present the results obtained in relation to scaling up reactors from lab-scale to pilot-plant scale, treating real wastewater from two different locations: the laboratory and in situ, respectively. In order to make using easily biodegradable organic matter more efficient, the filling phases of SBR cycles were adjusted according to a step-feed strategy composed of 6 anoxic-aerobic events. Feeding only occurred during anoxic phases. The results obtained demonstrated that the methodology may be useful in treating real wastewater with high carbon and nitrogen variations, as it always kept effluent levels lower than the official standards require (effluent total COD lower than 125 mg COD/L and effluent Total Nitrogen lower than 15 mg N/L). PMID- 15656302 TI - Integration of chemical and biological oxidation in a SBBR for tannery wastewater treatment. AB - This paper reports the results of an investigation aimed at evaluating the laboratory-scale performance of an innovative process for treating tannery wastewater. In this process, biological degradation, carried out in a sequencing batch biofilm reactor (SBBR), is combined with chemical oxidation by ozone. Tannery wastewater treatment was carried out, at laboratory scale, on a real primary effluent coming from a centralised plant treating wastewater produced by a large tannery district in Northern Italy. SBBR performance both without and with ozonation, was assessed with very satisfactory results. In particular, in the latter instance the recorded COD, TKN and TSS average removals, (96%), (92%) and (98%) respectively, allowed the maximum allowable concentration values fixed by the Italian regulation in force to be achieved without any additional polishing step. During the investigation biofilm properties (biofilm concentration and biofilm density) and flow dynamics aspects (head loss, shear stress, bed porosity) were also studied. A major feature of the process is that, with or without ozonation, it was characterised by very low specific sludge production (0.05 kgVSS/kgCODremoved) and high biofilm density (i.e. 87-122 gVSS/Lsludge) both contributing to a rather high biofilm concentration (i.e. 31 44 gTSS/Lfilter). PMID- 15656303 TI - Demonstration of enhanced nutrient removal at two full-scale SBR plants. AB - The efficacy of sequencing batch reactors (SBRs) to provide high levels of biological nutrient removal has been extensively demonstrated around the world. Environmental Solutions International (ESI) has now constructed over 20 full scale SBR plants and has confirmed that nutrient removal is enhanced via the process of simultaneous nitrification and denitrification. Over 18 months of operational data from two plants, operating in distinctly different catchments, processing an average of between 2,000 and 2,500 m3/d of wastewater, has clearly shown the efficacy and robustness of the ESI SBR-BNR process. Median effluent total nitrogen and total phosphorus values of 3 mg/L and <0.6 mg/L, respectively, were demonstrated over the 18-month period. This high level of nutrient removal is attributed to the design of the bio-selector which maximises carbon storage for the subsequent denitrification reactions, the effective aeration control which ensures no over-aeration during the air-on cycle as well as the level of operational control provided at these two plants. PMID- 15656304 TI - Enhanced biological phosphorus removal in RBC with SBR modification. AB - The rotating biological contactor (RBC) system was operationally modified with a sequencing batch reactor to achieve biological phosphorus removal from a weak domestic sewage along with nitrogen removal. This study utilized three RBC units, of which two units were the main units to remove phosphorus and NH4N and the third RBC unit was used as the storage of wastewater for its minimal effect to the PAO activities in the anaerobic stage during the operation. It was noticed that the biofilm thickness in RBC must be controlled to be less than 1.8 mm in order to achieve more than 70% of P removal with about 60% of N removal. With a settled sewage representing 200 mg/L of COD and 5 mg/L of P, the predicted P content in biofilm was more than 3% and the effluent P concentration was about 1 mg/L. The %P content in biofilm decreased with an increase of influent COD/TP ratios. The COD requirement for anaerobic P release was similar to reported values for the suspended growth system, however, the overall requirement increased with thicker biofilm. PMID- 15656305 TI - Response of an EBPR population developed in an SBR with propionate to different carbon sources. AB - The effect of different carbon sources (propionate, acetate, butyrate and glucose) on an enhanced biological phosphorus removal biomass developed with propionate as the sole carbon source was studied. Firstly, a group of different cycle studies was carried out using each substrate independently and then, another cycle study was performed with a mixture of substrates. Propionate was found to be the substrate with the highest substrate uptake rate in both sets of experiments. It was also the volatile fatty acid (VFA) which required less reducing power and less P-release to be uptaken. Four different polyhydroxyalkanoate (PHA) monomers produced during the anaerobic phase were detected, and PHB, PHV and PH2MV were quantified. Significant differences in PHA composition were obtained depending on the carbon source. The carbon recovery ratio for the anaerobic phase was also evaluated. The lowest value observed among the different cycle studies was obtained for butyrate, while the highest value was obtained for acetate. PMID- 15656306 TI - Short-term effects of carbon source on the competition of polyphosphate accumulating organisms and glycogen accumulating organisms. AB - The effectiveness of enhanced biological phosphorus removal (EBPR) systems is directly affected by the competition of polyphosphate accumulating organisms (PAOs) and glycogen accumulating organisms (GAOs). This study investigated the short-term effects of carbon source on PAO and GAO performance. The tests were designed to clearly determine the impact of volatile fatty acid (VFA) composition on the performance of two types of biomass, one enriched for PAOs and the other for GAOs. The two populations were enriched in separate reactors using identical operating conditions and very similar influent compositions with acetate as the sole carbon source. The only difference was that a very low level of phosphorus was present in the influent to the GAO reactor. The abundance of PAOs and GAOs was quantified using fluorescence in-situ hybridisation. The results clearly show that there are some very distinctive differences between PAOs and GAOs in their ability to utilise different carbon substrates. While both are able to take up acetate rapidly and completely, the GAOs are far slower at consuming propionate than the PAOs during short-term substrate changes. This provides a potentially highly valuable avenue to influence the competition between PAOs and GAOs. Other VFAs studied seem to be less usable in the short term by both PAOs and GAOs, as indicated by their much lower uptake rates. PMID- 15656307 TI - Monitoring pH and electric conductivity in an EBPR sequencing batch reactor. AB - This paper presents laboratory-scale experimentation carried out to study enhanced biological phosphorus removal. Two anaerobic aerobic (A/O) sequencing batch reactors (SBR) have been operated during more than one year to investigate the information provided by monitoring pH and electric conductivity under stationary and transient conditions. Continuous measurements of these parameters allow detecting the end of anaerobic phosphorus release, of aerobic phosphorus uptake and of initial denitrification, as well as incomplete acetic acid uptake. These results suggest the possibility of using pH and electric conductivity as control parameters to determine the length of both anaerobic and aerobic phases in an A/O SBR. More valuable information provided by monitoring pH and electric conductivity is the relation between the amount of phosphorus released and the conductivity increase observed during the anaerobic stages and which group of bacteria (heterotrophic or polyphosphate accumulating) is carrying out the denitrification process. PMID- 15656309 TI - A novel wastewater treatment process: simultaneous nitrification, denitrification and phosphorus removal. AB - Simultaneous nitrification and denitrification (SND) via the nitrite pathway and anaerobic-anoxic enhanced biological phosphorus removal (EBPR) are two processes that can significantly reduce the COD demand for nitrogen and phosphorus removal. The combination of these two processes has the potential of achieving simultaneous nitrogen and phosphorus removal with a minimal requirement for COD. A lab-scale sequencing batch reactor (SBR) was operated in alternating anaerobic aerobic mode with a low dissolved oxygen concentration (DO, 0.5 mg/L) during the aerobic period, and was demonstrated to accomplish nitrification, denitrification and phosphorus removal. Under anaerobic conditions, COD was taken up and converted to polyhydroxyalkanoates (PHA), accompanied with phosphorus release. In the subsequent aerobic stage, PHA was oxidized and phosphorus was taken up to less than 0.5 mg/L at the end of the cycle. Ammonia was also oxidised during the aerobic period, but without accumulation of nitrite or nitrate in the system, indicating the occurrence of simultaneous nitrification and denitrification. However, off-gas analysis found that the final denitrification product was mainly nitrous oxide (N2O) not N2. Further experimental results demonstrated that nitrogen removal was via nitrite, not nitrate. These experiments also showed that denitrifying glycogen-accumulating organisms rather than denitrifying polyphosphate-accumulating organisms were responsible for the denitrification activity. PMID- 15656308 TI - Influence of ORP variation, carbon source and nitrate concentration on denitrifying phosphorus removal by DPB sludge from dephanox process. AB - The effect of added carbon source and nitrate concentration on the denitrifying phosphorus removal by denitrifying phosphorus removal bacteria sludge was systematically studied using batch experiments, at the same time the variation of ORP was investigated. Results showed that the denitrifying and phosphorus uptake rate in the anoxic phase increased with the high initial anaerobic carbon source addition. However, once the initial COD concentration reached a certain level, which was in excess of the PHB saturation of Poly-p bacteria, residual COD carried over to the anoxic phase inhibited the subsequent denitrifying phosphorus uptake. This was equal to supplementing the external carbon source to the anoxic phase, furthermore the higher the external carbon source concentration the more powerful the inhibition caused. High nitrate concentration in the anoxic phase increased the initial denitrifying phosphorus rate. Oncethe nitrate was exhausted, phosphate uptake changed to phosphate release. Moreover, the time of this turning point occurred later with the higher nitrate addition. On the other hand, through on-line monitoring the variation of the ORP with different initial COD concentration, it was found that ORP could be used as a control parameter for phosphorus release, but it is impossible to utilize ORP for controlling the dinitrification and anoxic phosphorus uptake operations. PMID- 15656310 TI - Optimisation of storage driven denitrification by using on-line specific oxygen uptake rate monitoring during SND in a SBR. AB - This study builds on previous experience of maximising the formation of COD as poly-hydroxybutyrate (PHB) and now describes a feedback technique of preserving the use of PHB for denitrification resulting in enhanced nitrogen removal rather than allowing its wasteful oxidation by oxygen. The feedback technique uses on line SOUR monitoring for detecting the end-point of nitrification and controlling the aerobic phase length accordingly. The laboratory SBR was operated such that all organic substrate (acetate) was rapidly converted to PHB, which then served as the electron donor for nitrogen removal via simultaneous nitrification and denitrification (SND) during the aerobic phase (up to 70% SND). During SBR cycling with a fixed aeration length (240 minutes), PHB was unnecessarily oxidised after ammonium depletion, resulting in little denitrification and poor total nitrogen removal (69%). However, when the aerobic phase length was controlled via the SOUR, up to 1.8 CmM PHB (58 mg L(-1) COD) could be preserved, enabling improved total nitrogen removal (86%). The drop in the SOUR after ammonium depletion was a reproducible event that could be detected even when using raw wastewater and fresh activated sludge. The SOUR-control technique holds promise to build up PHB over a number of SBR cycles. While advanced oxygen control is used for improved N-removal in several existing WWTPs, this study investigates the importance of oxygen control with relevance to PHB driven SND in sequencing batch reactors. PMID- 15656311 TI - The presence of ammonium facilitates nitrite reduction under PHB driven simultaneous nitrification and denitrification. AB - For economic and efficient nitrogen removal from wastewater treatment plants via simultaneous nitrification and denitrification the nitrification process should stop at the level of nitrite such that nitrite rather than nitrate becomes the substrate for denitrification. This study aims to contribute to the understanding of the conditions that are necessary to improve nitrite reduction over nitrite oxidation. Laboratory sequencing batch reactors (SBRs) were operated with synthetic wastewater containing acetate as COD and ammonium as the nitrogen source. Computer controlled operation of the reactors allowed reproducible simultaneous nitrification and denitrification (SND). The oxygen supply was kept precisely at a low level of 0.5 mgL(-1) and bacterial PHB was the only electron donor available for denitrification. During SND little nitrite or nitrate accumulated (< 20% total N), indicating that the reducing processes were almost as fast as the production of nitrite and nitrate from nitrification. Nitrite spiking tests were performed to investigate the fate of nitrite under different oxidation (0.1-1.5 mgL(-1) of dissolved oxygen) and reduction conditions. High levels of reducing power were provided by allowing the cells to build up to 2.5 mM of PHB. Nitrite added was preferentially oxidised to nitrate rather than reduced even when dissolved oxygen was low and reducing power (PHB) was excessively high. However, the presence of ammonium enabled significant reduction of nitrite under low oxygen conditions. This is consistent with previous observations in SBR where aerobic nitrite and nitrate reduction occurred only as long as ammonium was present. As soon as ammonium was depleted, the rate of denitrification decreased significantly. The significance of the observed strongly stimulating effect of ammonium on nitrite reduction under SND conditions is discussed and potential consequences for SBR operation are suggested. PMID- 15656312 TI - Polyhydroxyalkanoates production by activated sludge in a SBR using acetate and propionate as carbon sources. AB - In this work, sludge was submitted to aerobic dynamic substrate feeding. Two sequencing batch reactors were operated, with acetate or propionate as carbon substrates. When acetate was used the system only produced a homopolymer of polyhydroxybutyrate (PHB). In order to maximize the PHB production, tests with different concentrations of acetate and ammonia were preformed. The best results (67.2% of PHB by cell dry weight) were obtained for 0.7 Nmmol/l of ammonia and 180 Cmmol/l of acetate. The PHB cell content was further improved by pulse addition of substrate, three times 60 Cmmol/l of acetate, reaching a value of 78.5%. Propionate can be used as a precursor for hydroxyvalerate. In conjunction with other substrates, it allows for the formation of copolymers, which present better processing properties on commercial applications. Tests with different concentrations of propionate and ammonia were performed. Under the operating conditions used, the maximum PHA accumulated inside cells was 34.8%, with 30 Cmmol/l of propionate and no ammonia. PMID- 15656313 TI - The influence of settling time on the formation of aerobic granules. AB - Aerobic granular sludge, without the addition of carrier material, has only been reported in one suspended growth system, the Sequencing Batch Reactor (SBR) operated with short fill and settling periods. Recent studies have demonstrated that extracellular polysaccharides increased with the formation of aerobic granules, and that the shear force may stimulate production of these polysaccharides. In the study described herein, two SBRs were operated with the same shear force (air flow rate 275 L h(-1)) and two different settling times (2 and 10 min). Only the reactor with 2 min settling formed completely granular sludge, although granules were present in both reactors. Community analysis using 16S rRNA PCR products and DGGE showed that the communities diverged quickly after reactor start-up. For samples taken at steady-state, the granular population was more stable and less diverse than the flocculent reactor. EPS extraction of samples using cation exchange resin yielded similar values for aerobic granular sludge and previously reported anaerobic granules. While differences in the protein and TOC content between the flocculent and granular reactors increased appreciably as the sludge became more granular, the protein to polysaccharide ratio was relatively constant. The experiment confirmed previous theories that short settling times in SBRs select for granular sludge. The settling time results in granular sludge having a higher EPS protein content and a less diverse but more stable population. PMID- 15656314 TI - Mass transfer impacts in flocculent and granular biomass from SBR systems. AB - An experimental study was conducted to describe mass transfer impacts within nitrifying aggregates sourced from sequencing batch reactor (SBR) activated sludge systems. Flocculent and granular sludge with high nitrification activity was obtained in two laboratory SBR systems, supplied with a synthetic, ammonium based feed. The flocculent biomass was fractionated using a sieving procedure, in order to obtain biomass fractions with different particle size distributions. The oxygen uptake rate (OUR) response to changes in dissolved oxygen concentration was measured under highly controlled conditions in a titrimetric and off-gas analysis (TOGA) sensor, and the results used to assess mass transfer effects. As the average particle size of the biomass increased, mass transfer limitations were found to increase significantly. Empirically fitted, apparent K(S,O2) values were demonstrated to be highly dependent on particle size, and reflect the mass transfer limitations occurring in the aggregates within a given system. Such parameters thus have little to do with the actual biokinetic parameter from which they are derived. The results obtained from the TOGA sensor study were consistent with those obtained from a microelectrode study on the same nitrifying granules. Together, these studies add considerable weight to the conclusion that consideration of external and internal mass transfer limitations is vital to the accurate description of activated sludge treatment processes, particularly those with a high oxygen uptake rate. PMID- 15656315 TI - Structure and microbial composition of nitrifying microbial aggregates and their relation to internal mass transfer effects. AB - This paper presents an analysis of the structure and microbial composition of nitrifying aggregates, formed as either flocs or granules, in sequencing batch reactors (SBR) operated with a high ammonium load. The structure and microbial community of the aggregates was determined by fluorescence in situ hybridisation (FISH). The aggregate structure and size was related to mass transfer limitations observed by measurements of OURs measured by either a titrimetric and off-gas analysis sensor (TOGA) or by microsensors. The FISH analysis showed that the spatial arrangement of the microbial consortia correlated well with the oxygen gradients inside the aggregates. In the larger aggregates, the ammonium- and nitrite-oxidising bacteria were mainly concentrated to the outer 100-200 microm, whereas in the floc system, the bacteria were distributed throughout the entire aggregate. This indicates that the internal mass transfer resistance is considerably larger when the aggregate size increases which is directly supported by TOGA measurements. PMID- 15656316 TI - Yannawa wastewater treatment plant (Bangkok, Thailand): design, construction and operation. AB - Yannawa Wastewater Treatment plant (Phase 1) serves a population equivalent of 500,000 and is located on a restricted site within the city of Bangkok, Thailand. Secondary treatment is based on the CASS sequencing batch reactor (SBR) process and the plant is one of the largest multi-storey SBRs in the world. The limitation of available site area, the ground conditions and the characteristics of the wastewater to be treated set a series of challenges for the designers, contractors and commissioning and operational staff. This paper briefly describes the collection system, the process selection and the treatment streams of the wastewater treatment plant. The SBR secondary treatment plant is described in more detail. The problems that arose during commissioning and operation and the solutions made possible by the use of an SBR type of process are discussed. Details of plant performance during performance testing and during the first three years of plant operation are provided. PMID- 15656317 TI - Functional analysis of microbial community in phenol-degrading aerobic granules cultivated in SBR. AB - Phenol-degrading aerobic granules were cultivated in a sequencing batch reactor with an influent phenol concentration of 500 mg l(-1). Eight strains were isolated from aerobic granules to characterize the functional redundancy of the microbial community in the granules. The specific oxygen utilization kinetics show the eight strains possessed different phenol-degrading activities, with half saturation constants (Ks) ranging from 0.4 to 70.5 mg phenol l(-1). Two isolates belonging to dominant populations expressed differing functions. The first strain was linked to the function of phenol degradation as this strain has the highest phenol-degrading ability among all isolates, while the second strain was linked to the maintenance of the granule structure because of its strong self flocculation activity. This study could be used to exploit the granule-based system for treating high-strength wastewaters. PMID- 15656318 TI - Aerobic biodegradation of 3-chlorophenol in a sequencing batch reactor: effect of cometabolism. AB - The aim of the present study was to investigate how phenol modifies, through cometabolism, the biodegrading capability of 3-chlorophenol (3-CP) in a sequencing batch reactor seeded with a mixed culture obtained from a domestic sewage treatment plant. Two laboratory-scale SBRs, one fed 3-CP only and the other fed 3-CP and phenol in the same concentration, were seeded with the partially acclimated biomass. The removal capability in both reactors was measured for progressive increases in the feed organic loading. Cometabolism enhanced biodegradation of 3-CP by reducing both the initial lag period and the time required for the complete removal. 700 mg/L 3-CP was demonstrated to be the highest concentration, which could be completely degraded during the active phase (fill plus react) either in the presence or absence of phenol as the growth substrate even though the lag period was shorter when phenol was present. The operating strategy required modification for the complete removal of 800 mg/L 3 CP. An increase in the phenol to 3-CP ratio did, however, improve 3-CP degradation rate. PMID- 15656319 TI - Two-stage SBR for treatment of oil refinery wastewater. AB - A two-stage sequencing batch reactor (SBR) system was used for treatment of oily wastewater with COD and oil and grease (O&G) concentrations ranging from 1,722 7,826 mg/L and 5,365-13,350 mg/L, respectively. A suitable start-up protocol was developed using gradual increase in oily wastewater composition with methanol as the co-substrate. This strategy enabled a short acclimation period of 12 days for the sludge in the two-stage SBR to adapt to the oily wastewater. After acclimation, the 1st stage and 2nd stage SBRs were able to achieve COD removals of 47.0+/-2.4% and 95.3+/-0.5%, respectively. The 1st stage SBR was able to achieve 99.8+/-0.1% of O&G removal and effluent O&G from the 1st stage SBR was only 6+/-2 mg/L. The 2nd stage SBR was used to further remove COD in the effluent from the 1st stage SBR. The final effluent from the 2nd stage SBR had a COD concentration of 97+/-16 mg/L with no detectable O&G content. Thus, a two-stage SBR system was shown to be feasible for treating high strength oily wastewater to meet the local discharge standards. PMID- 15656320 TI - Variation of the microbial activity during the acclimation phase of a SBR system degrading 4-chlorophenol. AB - The variation of microbial activity during acclimation to 4-chlorophenol (4CP) in an aerobic automated sequencing batch reactor was studied. The results show a reduction in degradation time as the acclimation process occurred. During acclimation for an initial concentration of 50 mg 4CP/L, degradation time was reduced from 40 h to 50 min after 10 cycles. In the case of an initial concentration of 100 mg/L, degradation time was reduced from 52 h to 1.16 h, also after 10 cycles. Doubling the initial concentration of a previously acclimated sludge produces only a slight increase in degradation time. It was found that as acclimation took place, the affinity of the consortia to biodegrade the toxic increased, whereas the ability to biodegrade acetate decreased. The evolution of the substrate uptake rate over time during the acclimation period was highly correlated with an exponential relationship. It was also observed that acclimation decreased both the production of a toxic metabolite and the sludge volumetric index. PMID- 15656321 TI - High COD wastewater treatment in an aerobic SBR: treatment of effluent from a small farm goat's cheese dairy. AB - In France, small goat's cheese dairies using traditional craft methods often have no profitable solution for dealing with the whey byproduct of their cheesemaking activity: it is usually mixed with the cleaning wastewater which, in the absence of other possibilities, is then discharged directly into the environment. The volume of such wastewater is small but it has a high COD of around 12-15 g/L. An aerobic SBR was proposed as a method for treating the mixture of wastewater and whey and the first installation was set up on a farm with 170 goats. Its operations were monitored for 7.5 months, particularly in order to measure any excess volume of sludge and to check that such excess remained within acceptable limits, given the high COD of the effluent requiring treatment. The results obtained show that the treated wastewater was of excellent quality, well within the most rigorous discharge norms. With this type of wastewater, excess sludge was produced in only very low amounts with 0.2 g of SS/g of COD. Moreover, the sludge proved to be quick settling which made it possible to: i) maintain a high level of SS in the reactor (up to 15 g/L); ii) withdraw sludge with concentrations reaching 30 g/L after 2 hours of settling. This resulted in a low volume of excess sludge (less than 5% of treated volume), making such aerobic biological treatment in an SBR competitive when compared to the straightforward spreading of all the wastewater. PMID- 15656322 TI - The performance of a nitrogen-fixing SBR. AB - A laboratory study has successfully demonstrated that a nitrogen deficient thermomechanical pulping wastewater can be effectively treated in a sequencing batch reactor (SBR) operated under conditions of biological nitrogen fixation (the N-ViroTech process). In comparison to continuous stirred tank reactor activated sludge (CSTR-AS) configurations operated under either nitrogen fixing or nitrogen supplemented conditions, slightly lower removals of dissolved organic material were observed in the SBR. However, this was largely offset by significantly better suspended solids removal in the SBR, which contributes to the overall COD discharge. The settleability and dewaterability of sludge produced by the SBR was significantly better than that obtained from the nitrogen fixing CSTR-AS reactors, and comparable to that of a nitrogen supplemented system. Consistently low total and dissolved nitrogen discharges from the N ViroTech systems demonstrated the advantage of this system over ones requiring nitrogen supplementation. The feast-famine regime of an SBR-type configuration has significant potential for the application of this technology in the treatment of nitrogen deficient waste streams, particularly those in which conventional single-stage systems may be susceptible to sludge bulking problems. PMID- 15656323 TI - Sludge settling processes in SBR-related sewage treatment plants according to the Biocos method. AB - This paper describes the investigations in a sedimentation and circulation reactor (SU-reactor) of a three-phase Biocos plant. The aim of these investigations was the determination of the temporal and depth-dependent distribution of suspended solid contents, as well as describing the sludge sedimentation curves. The calculated results reveal peculiarities of the Biocos method with regard to sedimentation processes. In the hydraulically uninterrupted (pre-)settling phase, a sludge level depth was observed, which remained constant over the reactor surface and increased linearly according to the sludge volume. The settling and the thickening processes of this phase corresponded to a large extent to the well-known settling test in a one-litre measuring cylinder. During the discharge phase, the investigated settling rate was overlaid by the surface loading rate and the sludge level changed depending on the difference between those two parameters. The solid distribution of the A-phase indicated a formation of functional zones, which were influenced by the surface loading. The formation was comparable to the formation of layers in secondary settling tanks with vertical flow. The concentration equalisation between the biological reactor and the SU-reactor proved to be problematic during the circulation phase, because a type of internal sludge circulation occurred in the SU-reactor. A permanent sludge recirculation seems to be highly recommendable. PMID- 15656324 TI - Deflocculation effects due to chemical perturbations in sequencing batch reactors. AB - Toxic shock-induced deflocculation was examined for activated sludge exposed to six different classes of industrially relevant chemical toxins: an electrophilic solvent (1-chloro-2,4-dinitrobenzene, CDNB), a heavy metal (cadmium), a hydrophobic chemical (1-octanol), an uncoupling agent (2,4-dinitrophenol, DNP), alkaline pH, and weakly complexed cyanide. The concentrations required to inhibit respiration by 50% were used to shock sequencing batch reactors (SBRs) containing a nitrifying (10-day solids retention time (SRT)) and a non-nitrifying (2-day SRT) biomass. Effluent total suspended solids (TSS) and soluble potassium were monitored to examine deflocculation caused by a bacterial stress response mechanism called glutathione-gated potassium efflux (GGKE). Reactors were monitored for recovery over a period of 3 SRTs or less. At the concentrations tested, CDNB, cadmium and pH 11 were found to cause significant increases in effluent TSS concentrations and showed elevated levels of potassium. In contrast, octanol, DNP and cyanide did not induce severe deflocculation and showed moderate increases in effluent potassium levels. Recovery of effluent TSS and potassium concentrations to control levels generally did not correlate, supporting the hypothesis that reflocculation requires regrowth of biomass. These results suggest that different chemicals induce deflocculation in SBRs, but deflocculation is not necessarily caused by the GGKE mechanism in all cases. PMID- 15656325 TI - Use of an anaerobic sequencing batch reactor for parameter estimation in modelling of anaerobic digestion. AB - The model structure in anaerobic digestion has been clarified following publication of the IWA Anaerobic Digestion Model No. 1 (ADM1). However, parameter values are not well known, and uncertainty and variability in the parameter values given is almost unknown. Additionally, platforms for identification of parameters, namely continuous-flow laboratory digesters, and batch tests suffer from disadvantages such as long run times, and difficulty in defining initial conditions, respectively. Anaerobic sequencing batch reactors (ASBRs) are sequenced into fill-react-settle-decant phases, and offer promising possibilities for estimation of parameters, as they are by nature, dynamic in behaviour, and allow repeatable behaviour to establish initial conditions, and evaluate parameters. In this study, we estimated parameters describing winery wastewater (most COD as ethanol) degradation using data from sequencing operation, and validated these parameters using unsequenced pulses of ethanol and acetate. The model used was the ADM1, with an extension for ethanol degradation. Parameter confidence spaces were found by non-linear, correlated analysis of the two main Monod parameters; maximum uptake rate (k(m)), and half saturation concentration (K(S)). These parameters could be estimated together using only the measured acetate concentration (20 points per cycle). From interpolating the single cycle acetate data to multiple cycles, we estimate that a practical "optimal" identifiability could be achieved after two cycles for the acetate parameters, and three cycles for the ethanol parameters. The parameters found performed well in the short term, and represented the pulses of acetate and ethanol (within 4 days of the winery-fed cycles) very well. The main discrepancy was poor prediction of pH dynamics, which could be due to an unidentified buffer with an overall influence the same as a weak base (possibly CaCO3). Based on this work, ASBR systems are effective for parameter estimation, especially for comparative wastewater characterisation. The main disadvantages are heavy computational requirements for multiple cycles, and difficulty in establishing the correct biomass concentration in the reactor, though the last is also a disadvantage for continuous fixed film reactors, and especially, batch tests. PMID- 15656326 TI - Anaerobic rumen SBR for degradation of cellulosic material. AB - Hydrolysis of organic particulates under anaerobic conditions is generally regarded as the rate limiting step in solid digestion processes. Rumen-based ecosystems appear to achieve very high hydrolysis rates for cellulosic organic material. This study aimed at the development and demonstration of an anaerobic sequencing batch reactor (SBR) process operating with a rumen-based microbial inoculum. Fibrous alpha cellulose was used as sole carbon substrate and the use of an SBR operating cycle allowed the utilisation of a high liquid flow rate (hydraulic retention time of 0.67 d) while maintaining a much longer solids retention time of 7 d. Complete mass balances for carbon and nitrogen, as well as COD balancing allowed the full characterisation of the process stoichiometry and kinetics. Elemental analysis of the biomass revealed a composition of C5H4.8O2.4N0.7, which is quite different from other generic biomass compositions used in the literature. The anaerobic rumen SBR was compared with another rumen based reactor system in the literature which used a continuous filtration process for solid/liquid separation. This comparison showed that the volatile fatty acid production rate from cellulose in the anaerobic SBR was comparable with the performance achieved in the continuous system, although loading, substrate type and media composition were quite different between these two studies. Further evaluation of the anaerobic rumen SBR is required to determine its practical application for other substrates and to demonstrate the scale-up potential of this concept. PMID- 15656328 TI - Gemcitabine and cisplatin following surgical treatment of urinary bladder carcinosarcoma. AB - The clinical case of a 73-year-old man with a history of transitional cell carcinoma of the bladder, an ulcerated mass on the left hemitrigone and left hydronephrosis who underwent radical cystoprostatectomy and urinary diversion followed by cisplatin-gemcitabine chemotherapy is presented. Pathological examination revealed a biphasic mixed tumor characterized by an epithelial and a mesenchymal component. At 24 months of follow-up the patient is alive and free from recurrent disease, with good quality of life and preserved renal function. Carcinosarcoma is highly aggressive and often has a dismal outcome regardless of treatment. Among all the studied prognostic factors, pathological stage is the main predictor of survival. The outcome of our patient suggests that the relatively well tolerated gemcitabine-cisplatin regimen after surgical treatment of invasive carcinosarcoma of the bladder might improve the currently dismal prognosis of selected elderly patients. PMID- 15656327 TI - Efficacy of epoetin alfa in a retrospective non-stratified subgroup analysis of a breast cancer cohort receiving non-platinum chemotherapy. AB - AIMS AND BACKGROUND: More than 60% of patients with metastatic breast cancer receiving non-platinum-based chemotherapy experience anemia, which is associated with fatigue and impaired quality of life. Epoetin alfa treatment in patients with a variety of malignancies has been shown to decrease transfusion requirements and improve hemoglobin levels and quality-of-life efficacy parameters. PATIENTS: Retrospective subgroup analyses were performed in patients with breast cancer who were part of a multinational, randomized (2:1), double blind, placebo-controlled trial of anemic cancer patients (n = 375) undergoing non-platinum-based chemotherapy. RESULTS: In the breast cancer subpopulation (n = 114, 48% with stage IV disease at baseline), the hemoglobin increase was greater for epoetin alfa patients than placebo patients (2.3 versus 0.9 g/dL). Epoetin alfa patients had lower transfusion requirements (28.2% versus 33.3%), improvement or preservation versus deterioration of quality of life, and a higher proportion of responders (patients achieving a > or = 2 g/dL increase in hemoglobin levels unrelated to transfusion) (68.0% versus 22.9% for placebo). The results were similar to those observed in the full study cohort, where statistical analyses showed the differences to be significant (P <0.05 for all). Epoetin alfa treatment was well tolerated. Although the study was not designed or powered for survival as an endpoint, Kaplan-Meier estimates for the full cohort showed a trend in overall survival favoring epoetin alfa treatment (P = 0.13, log rank test); a similar benefit was seen in the breast cancer subpopulation. CONCLUSIONS: In the full study cohort and the breast cancer subpopulation, epoetin alfa effectively treated anemia (increased hemoglobin levels and decreased transfusion requirements) and improved or preserved quality of life. Results concerning potential survival benefits support further study of epoetin alfa in anemic cancer patients. PMID- 15656329 TI - Experimental assessment of chemotherapy-induced early intestinal damage in colon cancer the lactulose-mannitol permeability test. AB - AIMS AND BACKGROUND: Although chemotherapy plays an important role in the management and cure of cancer, it has undesiderable side effects mostly affecting the bone marrow and gastrointestinal tract, which greatly limit patient compliance and treatment efficacy. METHODS: The lactulose-mannitol test was used to assess intestinal mucosa damage 48 hours after the end of the first adjuvant chemotherapy cycle with 5-fluorouracil (5-FU) and levamisole in 12 patients with colon cancer. Fifteen age- and sex-matched subjects were studied as controls. The excreted amount of lactulose and mannitol was expressed as the percentage of the administered doses recovered in the urine as well as their ratio. RESULTS: The percent urinary recovery of lactulose was significantly (P < 0.001) higher in colon cancer patients (1.1 +/- 0.5%) than in the control group (0.3 +/- 0.03%), whereas the mannitol recovery was only slightly reduced in the former. As a result, the lactulose/mannitol excretion ratio was significantly (P < 0.001) higher in colon cancer patients (0.07 +/- 0.03) than in the control group (0.01 +/- 0.01). CONCLUSIONS: As assessed by the lactulose-mannitol test, the combined chemotherapy regimen with 5-FU and levamisole affects mainly the barrier function of the intestinal mucosa rather than its absorption capacity. The toxic effect seems to be attributable to the 5-FU molecule rather than to levamisole. The lactulose-mannitol test is a simple, safe and reliable tool to evaluate chemotherapy-induced early damage to the intestinal epithelium, in particular when new kinds of substances are being administered. Its use in clinical practice seems appropriate to establish the correct timing of drug administration, thereby enhancing treatment efficacy and improving patient compliance. PMID- 15656330 TI - Narrower surgical margins might be sufficient in invasive horizontal growth phase melanoma. AB - BACKGROUND AND AIMS: The delineation of horizontal and vertical growth phases in primary cutaneous melanoma has contributed to our understanding of melanoma progression. Horizontal growth phase invasive melanomas are now believed to metastasize very rarely. Consequently, some of us have started to treat these lesions with very limited surgical margins, assuming that in terms of biological behavior this type of melanoma is more similar to an in situ than an invasive lesion. METHODS: Between January 1997 and December 2001 42 lesions of this type in 41 patients (24 women and 17 men) were treated in the outpatient clinic under local anesthesia. The excision margin was half a centimeter and the subcutaneous fat was cleared in most cases to the deep fascia, which was conserved. Loss was made good by direct tissue closure. All patients had undergone an excisional biopsy before definitive surgery. The size of the lesions ranged from 2 mm to 19 mm in maximum linear extent (median 7 mm). Lesion thickness ranged from 0.11 mm to 0.58 mm (median, 0.27 mm). RESULTS: The median follow-up was 47 months (range, 26-83). During this period none of the patients had locoregional or distant relapses. CONCLUSIONS: This preliminary report seems to corroborate the assumption that horizontal growth phase melanoma is not an aggressive lesion and might therefore be cured by non-aggressive surgery. The proper treatment of such lesions might be a surgical excision at half a centimeter distance from the biopsy scar. This approach may produce very good cosmetic results, while keeping the costs and required resources to a minimum. PMID- 15656331 TI - Gastric cancer: extent of lymph node dissection and requirements for a correct staging. AB - AIMS AND BACKGROUND: Lymphatic spread is an important prognostic factor in gastric cancer. The TNM classification requires at least 15 lymph nodes to stage and identify three prognostic groups according to the number of metastatic lymph nodes: N1 (1-6), N2 (7-15), N3 (> 15). The aim of this study was to investigate which type of lymph node dissection allows an accurate staging. METHODS: From 1996 to 2001, we treated 140 gastric cancer patients, 27 with D1 and 113 with D2 dissection. We evaluated lymph node count, status and ratio between metastatic and total number of excised lymph nodes, keeping 20% as the cutoff value. RESULTS: The mean number of lymph nodes was 18 and 33 respectively for D1 and D2 (P < 0.001), 41% of patients in D1 and 5% in D2 had less than 15 lymph nodes (P < 0.001). 59% in D1 and 73% in D2 (P = 0.145) had lymph node metastases, but this incidence decreased to 36% (P = 0.045) and 16% (P < 0.001) respectively for D1 and D2 when less than 15 lymph nodes were available. Considering the ratio between metastatic and total number of lymph nodes, 45% of D1 versus 3% of D2 (P < 0.001) in the N1 group exceeded 20%. CONCLUSIONS: D2 lymph node dissection is better than D1 in providing at least 15 lymph nodes required for a correct staging. We confirm the risk of a downstage when less than 15 lymph nodes are available. PMID- 15656332 TI - Outcome after conization for cervical intraepithelial neoplasia grade III: relation with surgical margins, extension to the crypts and mitoses. AB - AIMS AND BACKGROUND: Factors linked to residual neoplasia and recurrence following conization of the uterine cervix for treating cervical intraepithelial neoplasia grade III (CIN III), such as the surgical margins, extension of CIN into the crypts and the number of mitoses, have been studied with contradictory results. We evaluated patients submitted to conization for CIN III and analyzed the aforementioned factors, relating them to recurrence and residual neoplasia in hysterectomy specimens. METHODS: The surgical specimen of cold-knife conization for CIN III performed in 63 patients (average age, 37.3 +/- 9.3 years) was fixed in 4% formaldehyde. The ectocervical and endocervical margins were removed and the cone was cut into fragments perpendicular to the surface of the endocervical mucosa (1 mm thick). One histological section (5 microm thick) was cut from each block and stained with hematoxylin-eosin. We studied the total number of fragments from each cone and affected by CIN, endocervical and ectocervical margins, extension to the crypts, number of mitoses and tripolar mitoses in 100 microscope fields using a 100x objective. RESULTS: The endocervical margin was involved in 34.9% vs 9.5% (P = 0.001) of ectocervical margins. Recurrence affected 53.8% of cases presenting involved margins versus 12.9% in the cases without involved margins (P = 0.0078). The average interval to recurrence was 3.2 years. CIN was present in 2.5 to 100% (median, 28%) of the cone fragments (median no., 28; range, 7-95). A median of 44.4% and 25% of cone fragments presented CIN with and without recurrence, respectively. Correlation of the number of mitoses with tripolar mitoses and the percentage of fragments involved by CIN with the number of mitoses and tripolar mitoses showed, respectively, P = 0.02, 0.05 and 0.005. A median of 142 mitosis and 4 tripolar mitosis were observed per case with disease recurrence versus 104 and 3 (P = 0.02, 0.6), respectively, when recurrence did not occur. Of 14 patients who underwent hysterectomy after conization (mean, 3.6 +/- 3 months afterwards) for endocervical or both margins involved by CIN in the cone specimen, 8 (57.1%) presented CIN III and one (7.1%) microinvasive carcinoma. In 96.8% of the conizations, the CIN extended to the crypts. CONCLUSIONS: Involved margins and mitoses are associated with a greater recurrence rate for CIN. Residual neoplasia in the hysterectomy specimen after an involved margin with conization is frequent. PMID- 15656333 TI - Prognostic significance of serum lactate dehydrogenase in osteosarcoma of the extremity: experience at Rizzoli on 1421 patients treated over the last 30 years. AB - AIMS: The study evaluated the correlation between pretreatment serum lactate dehydrogenase (LDH) levels with the stage of disease and its clinical prognostic value. METHODS: Pretreatment serum LDH of 1421 patients with osteosarcoma of the extremity were assessed to investigate whether the enzyme correlates with the stage of the tumor. In 860 assessable patients with localized disease, treated according to 10 different protocols of adjuvant (four) and neoadjuvant chemotherapy (six), we also evaluated the correlation between the serum levels of LDH and outcome. RESULTS: According to the stage of disease, the rate of high serum level of LDH was significantly higher in 199 patients with metastatic disease at presentation than in 1222 patients with localized disease (36.6% vs 18.8%; P < 0.0001). In these patients, the 5-year disease-free survival was 39.5% for patients with high LDH levels and 60% for those with normal values. The 5 year disease-free survival correlated with serum level of LDH at univariate and multivariate analysis, although it lost its significance when histologic response to chemotherapy was also considered in the multivarite analysis. CONCLUSIONS: Serum LDH has a prognostic value and it should be considered in evaluating the results of therapeutic trials of chemotherapy, as well as defining a category of patients at high-risk of relapse to be treated with a more aggressive regimen. PMID- 15656334 TI - Correlation between postoperative infections and long-term survival after colorectal resection for cancer. AB - INTRODUCTION: Predicting long-term survival and cancer recurrence in patients with colorectal cancer is difficult because of the many factors that may affect the prognosis. This study investigated the prognostic significance of postoperative infections for patient outcome. METHODS: From an electronic database we selected 192 patients undergoing elective radical surgery for Dukes' stage B and C colorectal adenocarcinoma. The five-year survival rates were analyzed by the Kaplan-Meier method. Univariate and multivariate analyses were carried out to evaluate the potential prognostic variables using the Cox proportional hazard model. RESULTS: Forty-three patients developed deep incisional or organ/space surgical site infections, while the remaining 149 were complication free. The two groups were comparable for baseline, surgical and histopathological characteristics. At univariate analysis, Dukes' stage and infections were negative prognostic factors, while peritumoral infiltration of lymphocytes and eosinophils and fibrotic tissue appeared as protective variables. However, multivariate analysis showed that only Dukes' stage (P = 0.048) and occurrence of postoperative infectious complications (P = 0.011) were independently associated with outcome. In patients with infectious complications, the survival rate was significantly lower than in patients without infections (log-rank = 0.0004). CONCLUSIONS: The present results suggest the importance of evaluating other variables besides tumor stage in the prediction of long-term outcome. In prognostic studies more attention should be paid to postoperative infections. PMID- 15656335 TI - Differential tumor microenvironment in human ovarian cystic tumors. AB - AIMS AND BACKGROUND: Cells and soluble mediators obtained from tumor effusions are useful in evaluating the tumor microenvironment. Our aim was to examine cytologically and to quantify the leukocyte infiltrate, nitric oxide, cytokines and tumor markers in the intracystic fluid from patients with a cystic adnexal mass, for a possible differentiation between benign and malignant findings. METHODS AND STUDY DESIGN: Sixty-six women who had their cystic fluids collected were prospectively divided into benign tumor (22, 33.3%), malignant tumor (10, 15.2%) or other gynecological alterations (34, 51.5%). Cytology, total and differential leukocyte counts were determined by light microscopy. Tumor markers, cytokines and nitric oxide were assayed in the supernatants using the Immulite system, ELISA and Griess reaction, respectively. RESULTS: The sensitivity and specificity of the cytological analysis was 66.7% and 97.7%, respectively. The levels of CA 19.9, CA 15.3, alpha-fetoprotein, carcinoembryonic antigen, progesterone and beta-HCG were significantly higher in the benign and/or malignant group than in the other gynecological alterations. Also, the local concentrations of CA 15.3 and beta-HCG were significantly higher in malignant than in benign tumors. In malignant tumors, increased leukocyte counts and higher concentrations of IL-6, IL-10 and nitric oxide were detected than in benign tumors or other gynecological alterations. CONCLUSIONS: In malignant tumors, the microenvironment could be differentiated from benign tumors or other gynecological alterations by cystic fluid analysis. PMID- 15656336 TI - Invasive ductal carcinoma of the breast with a large central acellular zone associated with matrix-producing carcinoma. AB - Matrix-producing carcinoma is a variant of metaplastic carcinoma of the breast that consists of overt carcinoma with direct transition to a cartilagenous and osseous stromal matrix. A clinicopathological association between MPC and high grade invasive ductal carcinoma with a large central acellular zone has been recently discussed. However, few reports have ever shown the radiological, pathological and immunohistochemical correlation of this unique tumor. We present a case of 27-year-old woman with matrix-producing carcinoma with a large central acellular zone, which was characterized by a large homogeneous hypodense center with well-marginated, ring-like enhancement on contrast enhanced computerized tomography. PMID- 15656337 TI - Unilateral breast edema in two patients with malignant pleural effusion. AB - Unilateral breast edema usually signifies an underlying pathology of the breast and prompts extensive investigations for the purpose of an early treatment. Although breast edema has been reported with other systemic etiologies, it has not been described in patients with lung cancer. We report two cases of unilateral breast edema occurring in patients with non-small cell lung cancer and ipsilateral pleural effusion. Mammography and ultrasound of the breast both revealed increased interstitial density suggestive of fluid retention without any underlying masses. We performed a therapeutic pleurocentesis on one patient for symptomatic relief, and there was simultaneous improvement of the breast edema. We postulate a possible pathophysiology for the association between breast edema and malignant pleural effusion. The principle of management when one encounters similar cases would be to treat the underlying pleural effusion. PMID- 15656338 TI - A case of bilateral breast cancer and Graves' disease. AB - We report a case of bilateral breast cancer associated with Graves' disease characterized by a large goiter and complicated by a severe ophthalmopathy. The hyperthyroidism was treated initially with methimazole and then with thyroidectomy, the ophthalmopathy with intravenous steroids combined with orbital radiotherapy. The breast tumors underwent surgical resection followed by chemotherapy. We describe this case because of the well-known association between breast cancer and thyroid disease, particularly of the autoimmune type, the causes of this being still unclear. Recent literature on this topic is reviewed, discussing the possible role of the Na(+)-I(-) symporter and anti-TPO antibodies. PMID- 15656339 TI - Unusual breast tumors: primary lymphoma. A case report. AB - An 82-year-old woman presented with a right breast lump with erythematous reaction of the overlying skin and corresponding lymph nodes. Cytological examination of the breast lesion and lymph nodes suggested a lymphoid proliferation. Frozen section revealed carcinoma with lymphoid stroma. Simple mastectomy was performed because of the extent of the lesion. Histological diagnosis was non-Hodgkin type B large-cell lymphoma. Primary breast lymphomas behave similarly to lymphomas of similar histologic types occurring in other sites. The authors illustrate the diagnostic difficulties and the usefulness of complementary techniques in the diagnosis of an unusual breast mass. PMID- 15656340 TI - Nephrotic syndrome and angiotropic lymphoma report of a case. AB - A case of angiotropic lymphoma involving renal glomeruli and interstitial vessels associated with nephrotic syndrome and with minor lesions in the glomerular basal membrane is reported. A 56-year-old woman had fever, weakness and clinical findings of a nephrotic syndrome with normal renal function. Renal biopsy revealed that the glomeruli were infiltrated by neoplastic lymphoid cells positive for CD20 and CD45; the glomerular basement membranes showed a pattern of minimal change disease. This case and our review of the literature suggest that the rare association of intravascular lymphoma and glomerular disease is more than coincidental. PMID- 15656341 TI - Rapid tumor lysis syndrome in a patient with metastatic colon cancer as a complication of treatment with 5-fluorouracil/leucoverin and irinotecan. AB - Tumor lysis syndrome is a potentially fatal complication of anti-cancer therapy that is usually seen in patients with bulky, rapidly proliferating, treatment sensitive tumors such as hematological malignancies, but it rarely occurs in a variety of solid tumors such as colorectal carcinoma. Combination chemotherapy with infusional 5-fluorouracil/leucoverin and irinotecan has been recently accepted as the first treatment option for metastatic colorectal cancer. We present a case of tumor lysis syndrome in a patient with metastatic colon carcinoma that occurred 72 hrs after the initial course of a combination chemotherapy with irinotecan and 5-fluorouracil/leucoverin. Despite the immediate treatment with aggressive hydration by a sodium bicarbonate infusion, followed by forced diuresis and uricolytic therapy, he died of a sudden cardiac arrest complicated by acute renal failure. Our case indicates that administration of 5 fluorouracil/leucoverin and irinotecan for bulky tumors of colorectal origin with a rapid doubling time may induce an acute tumor lysis syndrome, which necessitates frequent laboratory monitoring and a close follow-up of the patient as well as prompt initiation of appropriate therapeutic measures. PMID- 15656342 TI - Cholestatic liver disease with ductopenia (vanishing bile duct syndrome) in Hodgkin's disease: report of a case. AB - Liver involvement is common in advanced stages of Hodgkin's disease. However, only a small percentage of patients with Hodgkin's disease develops jaundice due to several causes. Vanishing bile duct syndrome secondary to Hodgkin's disease is a rare cause of cholestasis in these patients. Only 20 cases, to our knowledge, have been reported so far in adults. We report a case of Hodgkin's disease presenting with obstructive jaundice without detectable liver involvement. Liver biopsies revealed intrahepatic cholestasis and ductopenia. Although the patient was given chemotherapy, he died of sepsis and disseminated intravascular coagulation after 24 weeks of admission to hospital. PMID- 15656344 TI - Cystic liver metastases from lung adenocarcinoma: a case report. AB - Cystic hepatic metastases arising from lung cancer are rare. We herein describe a case of a 71-year-old women admitted to our hospital for abdominal pain 6 months after the resection of a lung adenocarcinoma. Two cystic lesions of the liver were discovered at abdominal ultrasonography and computerized tomography scan. An ERCP excluded a biliary adenoma or adenocarcinoma, and an ultrasound-guided liver biopsy was negative for malignant cells. For persistence of symptoms and lack of a diagnosis, the patient underwent an exploratory laparotomy, a surgical biopsy with a diagnosis of adenocarcinoma, and a consequent right hepatectomy. After 2 years of follow-up, the patient is well and disease free. Although cystic liver metastasis are rare and a differential diagnosis difficult, the malignant nature should always be considered in the differential diagnosis of hepatic cysts to offer the patient the best treatment. PMID- 15656343 TI - An ovarian mucinous adenocarcinoma arising from mature cystic teratoma associated with respiratory type tissue: a case report. AB - Mature cystic teratoma (dermoid cyst) is the most common benign germ cell tumor of the ovary, accounting for approximately 30% of all ovarian tumors. Malignant transformation is rare; the most frequent transformation reported is to squamous cell carcinoma in 80% of cases, whereas transformation to adenocarcinoma is described in about 7% of cases. We report a case of malignant transformation to mucinous adenocarcinoma arising from respiratory-like epithelium in a mature teratoma of the ovary. PMID- 15656345 TI - Lymphangioleiomyomatosis: a case report and review of the literature. AB - Lymphangioleiomyomatosis, a rare disease of unknown etiology that is seen almost exclusively in women of childbearing age, generally presents with features of pulmonary involvement. It may be associated with tuberous sclerosis. Its clinical pulmonary manifestations vary from simple cough to the development of recurrent pneumothorax, hemoptysis, and even complicated pleural effusions. Progressive dyspnea develops as the disease evolves. Most patients eventually require lung transplant. This wide array of symptoms and signs makes the differential diagnosis extensive, and the clinician must be familiar with this disorder to arrive promptly to the correct diagnosis. Here we report a case of a 35-year-old woman with a history of pleuritic effusion with associated dyspnea before being diagnosed with lymphangioleiomyomatosis. A review of the literature pertinent to this case is also provided. PMID- 15656346 TI - [ASCO Annual Meeting 2004, New Orleans. Gemcitabine in metastatic breast carcinoma]. PMID- 15656347 TI - [Some aspects to discuss in the care of the elderly oncologic patient]. PMID- 15656348 TI - [ASCO Annual Meeting 2004, New Orleans. Capecitabine in adjuvant therapy of colorectal carcinoma: first... the evidence]. PMID- 15656349 TI - Thrombin: a paradigm for enzymes allosterically activated by monovalent cations. AB - Enzymes activated by monovalent cations are abundantly represented in plants and in the animal world. The mechanism, of activation involves formation of a ternary intermediate with the enzyme-substrate complex, or binding of the cation to an allosteric site in the protein. Thrombin is a Na+-activated enzyme with procoagulant, anticoagulant and signaling roles. The binding of Na+ influences allosterically thrombin function and offers a paradigm for regulatory control of protease activity and specificity. Here we review the molecular basis of thrombin allostery as recently emerged from mutagenesis and structural studies. The role of Na+ in blood coagulation and the evolution of serine proteases are also discussed. PMID- 15656351 TI - Genetic determinism of parasitic circadian periodicity and subperiodicity in human lymphatic filariasis. AB - The larval parasites of the pantropical lymphatic filariasis exhibit two types of circadian behaviour. Typically, they only appear in the human bloodstream at nighttime, synchronised with their mosquito vectors. In Polynesia and parts of Southeast Asia, free of nocturnal vectors, they are found at all hours, and each population biorhythm differs. Through a geometrical approach, we explain this circadian diversity by a single, dominant mutation: the clocks of individual parasites are set at midnight (ubiquitous) or at 2 p.m. Compared to other circadian genes, this mutation must be very old, as it is shared by four biologically remote genera of parasites. This seniority sheds new light on several theoretical and practical aspects of vector-parasite temporal relations. PMID- 15656350 TI - Fire and death: the pyrin domain joins the death-domain superfamily. AB - Apoptosis and inflammation are important cellular processes that are highly regulated through specific protein-protein interactions (PPI). Proteins involved in these signaling cascades often carry PPI domains that belong to the death domain superfamily. This includes the structurally well-characterized Death Domain (DD), the Death Effector Domain (DED) and the Caspase Recruitment Domain (CARD) subfamilies. Recently, a fourth member of the DD superfamily was identified, the Pyrin Domain (PYD). Based on sequence alignments, homology to other domains occurring in death-signalling pathways, and secondary-structure prediction, the PYD was predicted to have an overall fold similar to other DD superfamily members. Just recently, NMR structures of two PYDs have been determined. The PYD structures not only revealed the DD superfamily fold as previously predicted, but also distinct features that are characteristic exclusively for this subfamily. This review summarizes recent findings and developments regarding structural aspects of the DD superfamily, with a special emphasis on the PPIs of the DD superfamily. PMID- 15656353 TI - Spatial distribution of the copepod Centropages typicus in Ligurian Sea (NW Mediterranean). Role of surface currents estimated by Topex-Poseidon altimetry. AB - A particle-tracking model was used to simulate the dispersion and development of the planktonic copepod Centropages typicus during spring in Ligurian Sea. We show that mesoscale current structure, with a coastal jet and eddies, plays a key role in the transport and dispersion of C. typicus during its life cycle. Although, in the north, offshore Nice, cohorts can be advected southwestward out of Ligurian basin, more to the south others are retained in the central eddy and may give the start to the spring bloom of this species. However, input of individuals from the south through the Corsican Channel and along the west coast of Corsica may also be important in spring. This study shows that the ambit of C. typicus population is larger than the Ligurian Sea. PMID- 15656352 TI - Mitochondrial diseases preferentially involve proteins with prokaryote homologues. AB - The comparison of each of the 393 nuclear-encoded human mitochondrial proteins annotated in the SwissProt databank with 256,953 proteins from 94 prokaryote species showed that two thirds of the mitochondrial proteome were homologous with prokaryotic proteins, whereas one third was not. Prokaryotic mitochondrial proteins differ markedly from eukaryotic proteins, particularly in regard to their size, localization, function, and mitochondrial-targeting N-terminal sequence. Remarkably, the majority of nuclear genes implicated in respiratory chain mitochondrial diseases were found to be of prokaryotic ancestry. Our study indicates that the investigation of the co-evolution of eukaryotic and prokaryotic mitochondrial proteins should lead to a better understanding of mitochondrial diseases. PMID- 15656354 TI - Nuclear-DNA markers confirm the presence of two anchovy species in the Mediterranean. AB - Morphometric, allozymic, and mitochondrial DNA variability previously indicated that the Mediterranean anchovy Engraulis encrasicolus includes two distinct forms, one of the inshore habitat, and the other one of the open-sea habitat. Here, we showed that the two forms significantly differ by several morphological characters. To test the hypothesis, proposed previously, that the two forms are distinct biological species, we used length-polymorphic, exon-primed intronic PCR markers from the creatine-kinase multigenic family and genetically characterized anchovy samples collected in the northwestern Mediterranean and in the eastern Atlantic. Large genetic differences were found between the two forms (Weir and Cockerham's theta = 0.397 to 0.586). In contrast, geographic variation in the open-sea form at the scale of the eastern Atlantic/northwestern Mediterranean was weak (theta = -0.006 to 0.042). This again demonstrated considerable restriction to nuclear gene flow between inshore and open-sea anchovy populations. In addition to previous results from allozymes, this confirmed their status as distinct biological species, namely Engraulis albidus sp. nov. and Engraulis encrasicolus L. PMID- 15656355 TI - [Epidemiology and heat waves: analysis of the 2003 episode in France]. AB - Epidemiology and heat waves: analysis of the 2003 episode in France. The heat wave that struck France in 2003 has been accompanied with an estimated 15,000 excess deaths. This paper stresses the difficulties of the epidemiology of such an event. The relevant clinical and biological information is incomplete or even inaccessible and many of the deaths are due to multiple factors. The data presently available indicate that the deaths occurred in persons already vulnerable, and that the heat wave caused a five- to eight-month loss of lifetime for the affected individuals. There is a noteworthy similarity between the profiles of this exceptional summer mortality surge, and those of many past winters when similar or larger excess mortalities ave occurred without as yet eliciting much public attention. PMID- 15656356 TI - Performance of differential GPS collars in temperate mountain forest. AB - To determine the performance of Global Positioning System (GPS) for habitat studies in free-ranging animals, we tested differential 6- and 8-channel GPS collars under six representative canopies and one open-field reference site in the 'Parc national des Cevennes', southern France. The proportion of successful locations decreased under taller trees and worsened with snow accumulation in mixed coniferous habitats. The mean location success of seven free-ranging red deer fitted with 6-channel GPS collars in the same study area increased with a shorter interval between location attempts and during the leaf-off period. Our data suggested that the differences in location success between leaf-on and leaf off periods might reflect shifts in habitat use rather than a leaf effect under deciduous trees. PMID- 15656357 TI - Troglobitic scorpions: a new genus and species from Brazil. AB - A new genus and species of troglobitic buthid scorpion are described on the basis of a single specimen collected in Brazil. This is the first cavernicolous scorpion ever found in Brazil, and only the second to be found in South America. Some considerations on troglobitic scorpions are proposed. PMID- 15656358 TI - The impacts of logging on the microhabitats used by two species of couas in the western forest of Madagascar. AB - Habitat structure is important to consider in all ecological studies considering the relationships between animals and their environment. Habitat structure can be studied at different scales, from landscape to microhabitat. I studied here the response of two endemic terrestrial birds living in the dry forest of Madagascar. These birds belong to the genus Coua. The study is made at the microhabitat scale in a gallery forest, which has been logged selectively in order to limit the degradation of the forest. Selective logging is promoted to be respectful of the environment by allowing us to exploit the forest without destroying it and the wildlife encountered here. At the microhabitat scale, I underline that selective logging does not affect the Coquerel's coua, which can exploit new microhabitats and increase its density. On the other hand, the giant coua was affected by the restriction of optimal microhabitats for foraging. This species could be adapted to the new habitat by modifying its favourite foraging sites, but by decreasing also the population density. This species was affected by forest degradation, even considered as not destroying. At last, I considered how Coquerel's coua could be used as umbrella species for the endangered mesite. PMID- 15656359 TI - [Evaluations of extreme myopia correction with phakic posterior chamber intraocular lens and laser in situ keratomileusis]. AB - PURPOSE: To compare and evaluate the efficacy, safety and stability of extreme myopia correction with Phakic Posterior Chamber Intraocular Lens (PPCIOL) Keratomileusis (Lasik). and Laser in situ METHODS: This prospective study enrolled 43 cases of extreme myopia. The PPCIOL group was inserted Implantable Contact Lens (ICL) and Lasik group was done excimer laser scanning. The efficacy index and safety index were compared. The rate of regression postoperative was observed and the changes of low contrast vision and Glare vision were statistically studied. RESULT: The mean spherical equivalent refraction of PPCIOL was (-16.77+/-3.37)D(-11.75 approximately -25.75 D)and that of Lasik was (-13.8+/ 2.71) D (-9.37 approximately -23.75 D). There was no statistically difference between efficacy index of two groups postoperatively and a significant higher safety index in PPCIOL(P < 0.001) . The rate of regression was 0 in PPCIOL group and 32.56% in Lasik group (P< 0.001). The increase of low contrast vision and Glare vision from the baseline preoperative was more significant in PPCIOL group than which in Lasik group (P < 0.001). Two eyes of 1 PPCIOL case had anterior capsular cataract in 6 months postoperative. No other complication was found in two groups. CONCLUSION: PPCIOL implantation is an efficient method for extreme myopia correction with more safety and stability, and provides good visual quality. PMID- 15656360 TI - [The application of naphcon eye drops during Lasik surgery]. AB - PURPOSE: To evaluate the effects of Naphcon eye drops for preventing conjunctival bleeding during Lasik surgery. METHODS: One hundred cases (200 eyes) were divided into treating group and control group randomly according to using and not using Naphcon eye drops before Lasik surgery. Treating group patients received three times Naphcon eye drops during 15 minutes before surgery. RESULTS: The incidence of conjunctival bleeding from treating and control group were 8% and 15% respectively. There was significant statistical difference between them. CONCLUSION: Naphcon eye drop is an effective agent to prevent conjunctival bleeding. PMID- 15656361 TI - [Comparison of optic nerve head between normal and glaucoma subjects with Heidelberg Retina Topograph: to introduce a new diagnostic pattern]. AB - PURPOSE: To compare the differences of optic disc topograph between normal and glaucomatous persons, and to establish the score classification pattern for glaucoma using Heidelberg Retina Topograph (HRT). METHODS: One hundred and sixteen eyes from 116 normal controls, 99 eyes from 63 consecutive primary open angle glaucoma(POAG) patients, and 75 eyes from 42 patients with POAG in early stage were included. All subjects underwent HRT exams. The topograph parameters of optic nerve head obtained from HRT were compared between normal and glaucoma, according to the 50%,95%,and 99% distribution of each parameter defining as score -4, -3, -2, -1, 1, 2, 3, 4. The total value of all parameters in one eye were calculated as a diagnostic index. And the cut-off points were determined by specificity of 95% for certain glaucoma and certain normal, the others were classified into suspects. RESULTS: The differences of optic disc topograph between normals and glaucomatous patients were significant. When consecutive glaucoma data were analyzed by the new score classification pattern, miss diagnosis ratio was 6.1%, mis-diagnosis was 5.2%, correctly diagnostic ratio was 57.6%, suspects ratio was 36.4%. When POAG in early stage data were analyzed, the miss-diagnosis ratio and mis-diagnosis ratio were 5.2%, 5.2%, respectively; correctly diagnosis ratio is 36%, suspects ratio is 53.3%. CONCLUSIONS: The differences of optic nerve head topograph between normals and glaucomatous patients are significant. The new diagnostic pattern which will classify subjects into normals, suspects, and glaucoma, may be useful to lower miss-and misdiagnosis rate clinically. PMID- 15656362 TI - [The analysis of various extent visual field defect in chronic glaucoma patients]. AB - PURPOSE: To study the situation for the follow eye visual field defect of patients with severe visual field loss in 1 eye from chronic glaucoma,and to analyze the relative risk factors for visual field defect in such eyes. METHODS: The visual field defect scores of fellow eyes were calculated in 47 cases [primary chronic angle-closed glaucoma (PACG) 23 cases and primary open angle glaucoma (POAG) 24 cases] with severe visual field loss in 1 eye (visual field defect score > or = 12). The relations were analyzed between the visual field defect scores of fellow eyes and the visual acuity,the maximal and the mean intraocular pressure (IOP), ages, refraction, diagnosis, the last periods of glaucoma, gender and the anti-glaucoma surgery history. Spearman correlation analyse was used to estimated the correlation between the visual field defect scores of fellow eyes and the above factors. RESULTS: In the fellow eyes of the 23 cases PACG, 4 cases showed no visual field defect (score 0),5 cases showed the mild visual field defect (score 1-5), 7 cases showed the moderate visual field defect (score 6-11), 7 cases showed the severe or end-stage visual field defect (score 12-20). In the fellow eyes of the 24 cases POAG, 4 cases showed the mild visual field defect (score 1-5), 9 cases showed the moderate visual field defect (score 6-11), 11 cases showed the severe or end-stage visual field defect (score 12-20). The visual field defect score of fellow eyes in the last periods more than or equal to 10 years PACG group was significantly larger than that in the last periods less than 10 years PACG group (Z=2.8380, P=0.0224). The visual field defect score of fellow eyes in the mean IOP more than 18 mmHg PACG group was significantly larger than that in the mean IOP less than or equal to 18 mmHg PACG group (Z=1.9174, P=0.0276). The visual field defect score of fellow eyes in the anti-glaucoma surgery history POAG group was significantly larger than that in the non-anti-glaucoma surgery history POAG group (Z= 2.2377, P=0.0252). The significant positive correlation was found between the visual field defect scores and the mean IOP of the fellow eyes in the PACG group (r=0.4763, P= 0.0216),and the significant negative correlation was found between the visual field defect score and the visual acuity of the fellow eyes in the PACG group (r=-0.4416, P= 0.0349). CONCLUSION: Among the PACG with severe visual field loss in 1 eye, the visual field defect extent in the fellow eyes was related to the mean IOP, the last periods of PACG and the visual acuity of the fellow eyes. The visual field defect score was positively correlated with the mean IOP, and negatively correlated with the visual acuity of the fellow eyes. PMID- 15656363 TI - [The effect of phacoemulsification combined silicone oil remove and intraocular lens implantation on the eyes with silicone oil temponade]. AB - PURPOSE: To study the effect of phacoemulsification combined silicone oil remove and intraocular lens implantation on the eyes with silicone oil temponade. METHODS: Forty cases (41 eyes) performed phacoemulsification combined silicon oil remove and intraocular lens implantation were retrospectively studied and followed back for 2-18 months, analysis the recover of eyesight and complication of postoperation and during the operation. RESULTS: Except three cases had retina detachment after silicone oil remove, all others patients improved the eyesight in some degree. The main complication during the operation was the posterior capsule tear. The complication of postoperation was retinal detachment. CONCLUSION: It is an effective and safe way that the phacoemulsification combined silicone oil remove for eyes with silicone oil temponade. PMID- 15656364 TI - [Acute retinal necrosis: a report of 19 cases]. AB - PURPOSE: To improve the diagnose and treatment capacity on acute retinal necrosis (ARN). METHODS: Retrospectively analyse the manifestation, examination, diagnose, treatment and prognosis of the 19 cases with ARN in our Ophthalmic Center from January, 1996 to November, 2003. RESULTS: Nineteen cases had inflammation in anterior segment, vitritis, necrotizing retinitis and retinal vasculitis, except one patient. After treatment, visual acuity had been improved in 11 eyes (45.8%). Final visual acuity were lower than 0.05 in 11 eyes. Signs were controlled in 21 eyes(87.5%). CONCLUSIONS: The diagnose of ARN should be mainly based on clinical manifestation. Early and sufficient treatment with antiviral agents, corticosteroid and prophylactic laser photocoagulation, sometimes vitrectomy are the key point to control the disease. PMID- 15656365 TI - [The complications and affect factors in the surgery of orbital tumor]. AB - PURPOSE: The surgery complications in 215 orbital tumor cases were analyzed, in order to study the regularity and main affect factors of surgery complications. METHODS: Depending on clinical information, related factors closely with surgery of orbital tumor were analyzed statistically. RESULTS: Fifty-four kinds of various complications occurred within 45 cases (25.12%). Most complications were oculomotor apraxia (31.48%), and then were ptosis (22.22%), retrobulbar hemorrhage (18.51%), visual deterioration (12.96%), mydriasis (12.96%), injury of dura mater encephali (1.85%). CONCLUSIONS: Occurred surgery complications related to location of tumor (P=0.027), the nature of tumor (P=0.045), involved area of tumor and whether tumor with a clear line (P < 0.005). At the same time, it is also related to surgery approaches (P < 0.005). PMID- 15656366 TI - [Normal value of the central corneal thickness measured by non-contact specular microscope]. AB - PURPOSE: To get the normal value of the central corneal thickness (CCT) measured by non-contact specular microscope and to approach relatively influenced factor from different aspects. METHOD: The CCT was determined by the KONAN non-contact specular microscope. 173 normal subjects (322 eyes) were measured to get a normal value of the CCT. In the meantime, the para-central corneal thickness (12 o'clock, 2 o'clock, 10 o'clock, 6 o'clock) were measured and the corneal endothelial cell was analyzed in using the same machine. The statistical analysis was performed in using PC SPSS version 10.0 under windows system. RESULT: Measured by the non-contact specular microscope, the mean value of the CCT in normal eyes is (554.78+/-32.61), microm. There was no significant difference between right eyes and left eyes, and between male and female. There is no linear correlation between age, density and average area of the corneal endothelium cell and the CCT. The peripherial corneal thickness is thicker than the CCT. Among the peripherial corneal thickness, the thickest part is in the position of 12 o'clock,while the thinnest part is in the position of 6 o'clock. CONCLUSION: The CCT can be accurately measured by non-contact specular microscope. The measured value is reliable. PMID- 15656367 TI - [The histopathologic changes of human eyes after laser endoscopic cyclophotocoagulation]. AB - PURPOSE: To observe the changes in ciliary processes in endoscopic cyclophotocoagulation, and propose the principle for choosing energy of laser and access of operating. METHODS: Endoscopic cyclophotocoagulation was performed in the human eyes donated for corneal transplantation through the limber or the pars plana. The coagulated tissue was checked histologically. RESULTS: The ciliary body was viewed clearly with the endoscope (E2). The best choice of energy was shuthe range from 0.4 W to 0.6W, and the most effective reaction was that the processes turned to be pale and shrunk. The main histopathologic changes were that the normal structure of ciliary body was destructed, the pigment epithelial cells and stroma degenerated and the no-pigment epithelial cells kept intact. CONCLUSION: Cyclophotocoagulation could be completed accurately visually under the endoscope. The principle of lowering intraocular pressure was the destruction of ciliary stroma and ciliary pigment epithelial cells. PMID- 15656368 TI - [The adverse reactions and treatment of fundus fluorescein angiography]. AB - PURPOSE: To investigate the safety of fluorescein angiography and to evaluate the examination. METHODS: We retrospectively analyzed 8 344 patients who underwent fluorescein angiography in our hospital in the past four years. The rate of adverse events was calculated and the emergency management for severe cases was discussed. RESULTS: There were 693 cases (8.3%) of adverse events occurring in the patients. Many of them were mild or moderate ones, which were 664 cases in number and were 7.87% in rate. The other 29 cases (0.35%) were severe ones. No special treatment was needed for the mild or moderate adverse events, the symptoms can be relaxed by deep breathing or having a rest for a moment. The symptoms of the severe adverse events can be relieved through anti hypersuscepibility treatments and no sequelae left. No case of death occurred. CONCLUSION: The adverse events could occur in fluorescein angiography examination. It suggests that we should pay attention to it and be aware of the system conditions of the patients. It is also important for us to grasp the indications and contraindications and to prepare for the emergent events. In summary, fluorescein angiography is generally a safe method for the examination of fundus disease. PMID- 15656369 TI - [Efficacy of rigid gas permeable contact lens for the correction of mixed astigmatism]. AB - PURPOSE: To evaluate the efficacy of rigid gas permeable contact lens (RGPCL) for the correction of mixed astigmatism. METHODS: Patients with mixed astigmatism by mydriasis refraction were selected as subjects. Routine eye check was performed to exclude contraindication of contact lens. Investigation included visual acuity, keratometry and corneal topography. Average keratometry value was chose as base curve of the first trial lens. Final base curve, diameter, material and edge of lens were prescribed according to the centralized position, movement and fluorescence pattern of trial lens. After over refraction, final refractive power of lens was prescribed. Corrective visual acuity with spectacle and rigid gas permeable contact lens were compared in 24 patients (31 eyes). RESULTS: Corrective visual acuity of patients with RGPCL was much better than those with spectacle. CONCLUSIONS: RGPCL is safe and effective to correct mixed astigmatism. PMID- 15656370 TI - [Expression of matrix metalloproteinase in human pterygia]. AB - PURPOSE: To determine the cellular distribution and activation of MMP-2 and MMP-9 in human pterygium. METHODS: The expression and activation of MMP-2 and MMP-9 were determined by immunohistochemistry and quantitative gelatinase zymography from 27 pterygia (26 cases, 18 eyes in stationary pterygia and 9 eyes in progressive pterygia) and 6 normal conjunctival specimens. RESULTS: MMP-2 was expressed in all pterygium examined, specifically localized to the epithelium. MMP-9 was the most abundant in pterygium vascular endothelium and inflammatory cell. MMP-2 levels were no difference between the stationary pterygia [(235+/ 18.17) scanning units] and the progressive pterygia [(221+/-22.53) scanning units]. MMP-9 levels in the progressive pterygia [(159+/-19.28) scanning units] were significantly higher than that in the stationary pterygia [(46+/-32.21) scanning units] (P < 0.05). CONCLUSION: This investigation located the expression of MMP-2 and MMP-9 in pterygium of the human eyes. These enzymes may be responsible for the progression of pterygia. PMID- 15656371 TI - [Curcumin inhibits the proliferation of cultured human fetal retinal pigment epithelium cells]. AB - PURPOSE: To investigate the effect of curcumin on the proliferation of human fetal retinal pigment epithelium (hfRPE) cells in vitro. METHODS: Subcultured hfRPE cells were treated with different concentrations of curcumin (2.5, 5, 10, 20 microg/ml) for 24 or 48 hours. MTT assay and flow cytometry were performed to evaluate the effect of curcumin on the proliferation and cell cycle of cultured hfRPE cells. RESULTS: Curcumin was effective in inhibiting the proliferation of subcultured hfRPE, the IC50 value at 24 and 48 hours were found to be 14.27 microg/ml, 12.7 microg/ml, respectively. Cell cycle analysis indicated that CUR blocked cells in G2/M phase. CONCLUSION: Curcumin can inhibit the proliferation of hfRPE cells by cells cycle arrest. PMID- 15656372 TI - [The toxic effect of N-methyl-N- nitrosourea on retina in rats]. AB - PURPOSE: To observe the toxic effect of N-methyl-N-nitrosourea (MNU) on photoreceptor cells within retina of SD rats. METHODS: At 50 days of age, 100 female rats received a single intraperitoneal injection of MNU at different doses of 50 mg/kg, 60 mg/kg, 70 mg/kg and 80 mg/kg body weight, respectively. Each group had 6 rats and 4 untreated rats were used as normal control. At 24, 48 and 72 hours and 7 days after the administration of MNU, the animals were sacrificed and both eyes were enucleated immediately and processed for histological examination. RESULTS: It was found that all doses of MNU could sequentially damage the central and peripheral retina. The first evidence of retinapathy 24 hours after the application of MNU was pyknosis and disruption of photoreceptor cells nuclei and the disorientation of the photoreceptor outer segments; loss of photoreceptor cell deteriorated significantly at 48 hours or 72 hours; the outer nuclear layer and photoreceptor layer were almost completely lost at 7 days. CONCLUSION: The results demonstrated that MNU could selectively damage the photoreceptor cells in the retina of the rats, which was dose-dependent and time dependent. PMID- 15656373 TI - [The culture of retinal muller cells and expression of its glutamate transporter GLAST]. AB - PURPOSE: To culture retinal Muller cells (RMC) of infantile and mice in postnatal day 3 (P3) and study the expression of GLAST. METHODS: RMCs of infantile and mice in P3 were cultured by tissue inoculation and shaken to allow detachment of other retinal cells. Then the morphologic and growth characteristics of the cells was oberseved every day. The second passage cells were identified with GFAP, Vimentin and the expression of GLAST was observed with immunofluorescent staining. RESULT: The cultured RMCs reached confluence in 15-20 days. These cells were positive in GFAP, Vimentin and GLAST antibody staining. CONCLUSIONS: RMCs of infantile and mice in P3 can be successfully cultured by tissue inoculation, GLAST widely expresses on RMCs membrane. PMID- 15656374 TI - [The expression of p53, MDM2 and Ref1 gene in cultured retina neurons of SD rats treated with vitamin B1 and/or elevated pressure]. AB - PURPOSE: To investigate the expression of p53, MDM2 and Ref1 gene in cultured retina neurons of SD rats treated with Vitamin B1 and (or) elevated pressure. METHODS: The retinal neuron of postnatal SD rats were cultured in vivo, the elevated pressure was produced after 7 days, and the total RNA was extracted after another 2 days, expression of p53, MDM2 and Ref1 gene were analyzed with RT PCR. RESULTS: The expression level of p53 and MDM2 gene were increased in elevated pressure group, normal with Ref1 gene expression. But the expression of p53 and MDM2 gene were decreased significantly in elevated pressure group treated with vitamine B1 compare to the elevated group. CONCLUSION: Apoptosis seem to be a mechanism of cell death in retinal neurons of SD rats with elevated pressure.Vitamine B1 have protect effects against elevated pressure. PMID- 15656375 TI - [Decreased chaperone activity of alpha-crystallin by carbamylation in vitro]. AB - PURPOSE: To investigate the effort of carbamylation, an important post translational modification of lens proteins, on chaperone activity of alpha crystallin. METHODS: The alphaL-crystallin and betaL-crystallin were isolated from bovine lens. Carbamylation was performed by different concentrations of potassium [14C] cyanate to alphaL-crystallin at 37 degrees C for 7 days. The binding of cyanate to alphaL-crystallin was determined by measuring the radioactivity incorporated into trichloroacetic acid precipitable protein from potassium [14C] cyanate. In addition, alphaL-crystallin was incubated with unlabelled potassium cyanate (50 and 100 mmol.L(-1)) for 3 and 7 days at 37 degrees C and then its chaperone activity against heat-induced betaL-crystallin aggregation was assayed. Analysis of the carbamylated alphaL-crystallin was further performed by high performance liquid chromatography (HPLC). RESULTS: The rate of binding (cyanate bound /mol of alphaL-crystallin) and the decreased chaperone activity of alphaL-crystallin induced by carbamylation in vitro were dose-dependent and time-dependent. The decreased chaperone activity corresponded with the higher binding of cyanate. After 7 days incubation with 100 mmol.L(-1), almost all chaperone activity was lost compared with a simultaneously incubation control sample. HPLC analysis revealed that aggregation of alphaL-crystallin by potassium cyanate has occurred after 3 days incubation in dose-dependent fashion. CONCLUSION: The alpha-crystallin can be modified in vitro by carbamylation through a high-molecular-weight aggregates formation. The loss of its chaperone activity may result from carbamylation-induced aggregation. PMID- 15656376 TI - Resistance to antibiotics: administrative response to the challenge. AB - The authors discuss antibiotic resistance within a conceptual framework that illustrates the dynamic relationships among antibiotic, patient, and population factors. The complexity of these interactions makes it unlikely that any single intervention or approach will adequately address the problem of increasing rates of antibiotic resistance. A case study focused on Streptococcus pneumoniae in the context of community-acquired pneumonia provides a detailed examination of the manner in which antibiotic use, expenditures, and microbial resistance are affected by an administrative reimbursement restriction implemented by a single government payer. PMID- 15656378 TI - What is next on the political front? PMID- 15656377 TI - Inpatient utilization by dual Medicare-Medicaid eligibles in Medicare risk HMOs and fee for service, California, 1991-1996. AB - Inpatient use among dual Medicare-Medicaid eligible beneficiaries in California Medicare HMOs and fee-for-service plans from 1991 to 1996 was compared, using a unique dataset that links Medicare enrollment data to inpatient discharge data. Dual eligibles in HMOs were found to have lower discharge rates, shorter lengths of stay, and fewer inpatient days than dual eligibles in the traditional fee-for service system. Both, however, had higher discharge rates and inpatient days than non-dual-eligible beneficiaries. The results are consistent with previous findings documenting the high cost of dual eligibles, with the lower use in HMOs likely the result of differences in beneficiary characteristics and delivery of care between systems. PMID- 15656379 TI - Does disease management reduce health care costs? PMID- 15656381 TI - Health divides. PMID- 15656380 TI - The economic value of lamotrigine as a mood stabilizer: a U.S. managed care perspective. AB - This study attempts to provide real-world evidence of the economic value of lamotrigine as a mood stabilizer for patients with bipolar disorder. Researchers examined longitudinal administrative claims data obtained for patients with a diagnosis of affective disorder between 1997 and 2001. The study compared outcomes for the 12 months preceding initiation of lamotrigine with the 12 months following initiation of lamotrigine in patients previously treated with lithium, carbamazepine or valproic acid, other anticonvulsants, selective serotonin reuptake inhibitors, or other antidepressants. Outcomes were analyzed using logistic and Tobit regression models. The data suggest that initiation of lamotrigine is associated with reductions in hospital days, a reduction in hospital days of greater than 17 days for some patients, and net cost savings of more than $400 per patient per year. PMID- 15656382 TI - Peas of mind. PMID- 15656384 TI - Professionals' addiction to medical labels is changing our children's experience of childhood. PMID- 15656383 TI - Small talk. PMID- 15656385 TI - A home of their own. PMID- 15656386 TI - Tackling them young. PMID- 15656387 TI - Same story--different tale. PMID- 15656388 TI - Make dreams come true. PMID- 15656389 TI - Diatribe. PMID- 15656390 TI - [The principles of action and use of phlebotropic agents]. PMID- 15656391 TI - [Electrophysiological atrial and sinus nodal remodelling phenomenon: mechanisms of development and pathogenesis]. PMID- 15656393 TI - [Coronary stenting in patients with chronic forms of coronary heart disease]. AB - The paper presents the results of treatment in patients with chronic heart disease by coronary stenting. Endovascular interventions have been found to be highly effective in attenuating the clinical manifestations, in improving the results of exercise tests, and in reducing needs for medical treatment. The late postoperative factors influencing the higher incidence of stent stenosis have been studied. PMID- 15656392 TI - [Potentialities of cytoprotection in the treatment of chronic heart failure in patients with coronary heart disease]. AB - The study was undertaken to evaluate the effects of long-term (6-month) therapy with the selective beta-blocker bisoprolol, the cytoprotector trimetazidine and their combination on the clinical course of disease, the morphofunctional parameters of the left ventricle (LV) and life quality in 71 patients with coronary heart disease (CHD) concurrent with functional classes (FC) III-IV chronic heart failure (CHF) (ejection fraction (EF) < 35%, as evidenced by echoCG). In Groups 1, 2, and 3, basic therapy was supplemented by bisoprolol, trimetazidine, and bisoprolol plus trimetazidine, respectively. The initial dose of bisoprolol was 1.25 mg with its subsequent titration to an individually tolerable. Trimetazidine was given in a dose of 20 mg thrice daily. The patients' clinical status and echoCG were monthly assessed. There was a decrease in FC of CHD in all the groups, a reduction in LV end diastolic and systolic volumes, and an increase in shortening faction, LV EF, and an improvement of LV diastolic function. There was evidence that it should be expedient and safe to use the cytoprotector trimetazidine in the treatment of CHF in patients with CHD. In patients with CHD concurrent with CHF, therapy using a combination of bisoprolol and trimerazidine was found to have the most pronounced impact, which yielded the maximum clinical and hemodynamic effect (LV EF increased by 42.6% of the baseline values, the rate of early and late ventricular diastolic filling decreased by 59% of the baseline values). The revealed regularities of the positive effect on the clinical and hemodynamic parameters of long-use of bisoprolol, trimetazidine, and their combination in patients with CHD concurrent with FC III-IV CHF show it expedient to include these drugs and their combination into therapy for CHF that develops in the presence of CHD. PMID- 15656394 TI - [Myocardial ischemia and paroxysmal cardiac arrhythmia in different severity of bronchial asthma]. AB - The study was undertaken to define the prevalence and pattern of myocardial ischemia and cardiac arrhythmia in patients with persistent bronchial asthma (BA) in relation to its severity. Eighty-one patients with persistent BA, including 21 patients with mild BA, 39 with moderate BA, and 21 with severe BA, were examined. ECG and Holter 24-hour monitoring were performed. Myocardial ischemic episodes were detected in 33% of the patients with moderate BA and in 57% of the patients with severe BA. Silent and painful myocardial ischemia occurred at the equal rate in moderate BA and painful episodes predominated in severe BA. The incidence and degree of supraventricular and ventricular extrasystoles increased with the severity of BA. The paroxysms of ventricular tachycardia, atrial fibrillation and flutter appeared in the presence of BA concurrent with myocardial ischemia. PMID- 15656395 TI - [The duodenal epithelium in duodenal diseases]. AB - The purpose of the study was to define duodenal epithelial function in duodenal diseases and to assess the potentialities of correction of detected dysfunctions. The electrical properties (EPs) of the duodenal epithelium were explored in patients with primary chronic duodenitis, duodenal peptic ulcer (DPU) on an exacerbation and in the late postoperative period after selective proximal vagotomy. Mucosal biopsy specimens were obtained at fibrogastroduodenoscopy with biopsy forceps and placed in a sign-alternating field with the parameters, adequate bioelectric epitheliocytic characteristics: 28 V at a voltage sign change rate of 1 Hz, and a current strength of 200 microA. Bioelectrical cell reactions (nuclear fluctuation amplitude) were recorded by light microscopy using a MOB-1A ocular micrometer at minutes 10, 30, and 60 of the onset of electrical field action. A significant reduction was found in the amplitude of duodenal epithelial nuclear fluctuations in the examinees, which was most pronounced in DPU on an exacerbation. Duodenal epithelial EPs were shown to undergo regular changes during therapy and may serve as an additional test in evaluating the efficiency of therapy. PMID- 15656397 TI - [Mycotic infection in gastric peptic ulcer]. AB - To assess the risk factors of mycotic infection on the gastric mucosa (GM) in patients with gastric peptic ulcer, 34 patients who had much mycelium on the bottom of the defect and 122 patients without fungal infection were examined in detail. The occurrence of mycotic infection in patients with ulcerative defects was associated with a variety of both local (the depth, number, large sizes of defects, diminished gastric acid-producing ability) and total (age, trophological insufficiency, iron-deficiency anemia, oral mycotic disease, autonomic dysfunction) factors. Mycotic involvement of ulcerative defects is secondary and regarded as a focal manifestation of mycotic infection. The likely impact of mycotic infection on the quality of ulcerative defect scarring makes us consider whether antimycotic agents should be added to antiulcerative therapy. PMID- 15656396 TI - [Efficacy of derivatives of 3-hydroxypyridine and succinic acid in diabetic patients with the diabetic foot]. AB - A prospective placebo-controlled study was performed to examine the effects of emoxipine, reamberine, and mexidole on the severity of neuropathic and anxiety depressive symptom clusters, by simultaneously evaluating the parameters of the lipid peroxidation-antioxidative defense protection (LPO-AOD) system in diabetic patients with the diabetic foot. The 14-day use of all the test drugs was found to cause a significant attenuation of diabetic neuropathy and to abolish concomitant anxiety and depression. The positive clinical effect of emoxipine and reamberine was shown to be associated with the stimulation of AOD and with the limitation of LPO. Mexidole is not inferior to emoxipine and reamberine in its clinical efficacy, but it did not change the parameters of the LPO-AOD system as compared with placebo. PMID- 15656399 TI - [Dipyridamole prevention of outbreaks of respiratory infections in the homogeneous population]. AB - The purpose of the study was to evaluate the effectiveness and prevention of outbreaks of community-acquired pneumonia (CAP) and acute respiratory diseases (ARD). The study included conscripts arrived at a training center. An 8-week course of prevention (curantyl, 100 mg a week) was assessed. The end point of the study was a new case of CAP and ARD. The study covered 2333 persons, of them 2226 completed the study. In the control group (n = 1034), CAP developed in 11.9% of the examinees (95% confidence interval (CI), 10-14%), clinical cases of ARD were observed in 18.7% (95% CI, 16-21%. The same parameters in the prophylaxis group (n = 1192) were 9.6% (95% CI, 8-11%) and 11,7% (95 CI, 10-14%). The relative risk of the diseases for a prevention group was 0.8 (95% CI, 0.7-1.1) (p = 0.34) and 0.6% (95% CI, 0.5-0.8) (p < 0.0001), respectively. The study has demonstrated that dipyridamole is effective and safe as a drug for preventing ARD. The findings were slightly more scarce than those obtained by other authors, which may explain the performance of the study in summer when the role of respiratory viruses and related disease is not so significant. PMID- 15656398 TI - [Pharmacological capacities of the prevention of colorectal cancer]. AB - The paper discusses the results of researches and clinical cases of the use of some drugs and diet in the prevention of colorectal cancer (CRC). Particular emphasis is laid on the efficacy and mechanism of antitumor action of aspirin and nonsteroidal antiinflammatory drugs, folic acid, calcium, ursodeoxycholic acid, epidermal growth factor receptor inhibitors, alpha-difluoromethylornitine, selenium. The paper underlines the importance of dietary addition of products containing the high levels of dietary fibers (available in vegetables, fruits, wheat bran, porridges, etc.) and nonhydrogenated unsaturated fats and omega3 fatty acids. To keep to the general routine involving high physical activity, obesity prevention is of great value in the prophylaxis of CRC. PMID- 15656400 TI - [Systemic vasculitis as an interdisciplinary problem]. AB - Systemic vasculitis (SV) is characterized by generalized vascular bed lesion involving vessels of different sizes into a pathological process. The paper presents the results of a follow-up of 500 patients with different forms of SV, by making studies of immunity and the hemostatic system, angioscanning, Doppler ultrasound study of vessels, electrophysiological studies (rheoencephalography, encephalography), computed and magnetic resonance imaging of the brain, and visceral ultrasonography. A variety of clinical symptoms and involvement of different organs determine the interest of physicians of different specialties in the diagnosis and treatment of SV. The involvement of the nervous system in the process occurs in all forms of vasculitis, by afflicting the central, peripheral, and autonomic nervous systems with the development of regulatory and functional disorders. Lesions of the visual organ are typical of nonspecific aortoarteritis (Takayasu's disease), Wegener's granulomatosis, giant-cell arteritis. Recurrent uveitis is characterized in Behcet's syndrome. Cutaneous manifestations are included into the classification criteria of nodal polyartheritis, hemorrhagic vasculitis, and Kawasaki's disease. ENT and oral involvement are observed in Wegener's granulomatosis. PMID- 15656401 TI - [Migrating subcutaneous myasis with systemic manifestations caused by gadfly larvae]. AB - The paper describes a case of cutaneous myiasis caused by the larvae of Hypoderma bovis. Its prolonged recurrent course over 4 months was observed. It appeared as migrating mass lesions, toxicoallergic reactions, and syncope. The paper is the first to show it possible to perform chemotherapy of myiasis with the antibiotic ivermectin in combination with aversectin ointment. PMID- 15656402 TI - [Fibrosing alveolitis as a complication due to D-penicillamine therapy for Wilson Konovalov disease]. PMID- 15656403 TI - [Bilateral spontaneous pneumothorax in an adolescent suffering from histiocytosis X]. PMID- 15656405 TI - [The 11th Russian National Congress on Man and Drug (Moscow, April 19-23, 2004)]. PMID- 15656404 TI - [Clinical and economical studies: real or fictitious benefit?]. PMID- 15656406 TI - [Computer monitoring- based algorithm for the discrimination between pathological and compensatory changes in acute disorders of blood circulation]. AB - A differential diagnosis algorithm for pathological and compensatory changes was implemented on the basis of a general-access "Microsoft Access 2002" data bank (DB). The DB algorithm makes the soft more flexible; the interpretation of findings can be changed unassistedly in the real time mode with respect to specific needs encountered in the treatment of cardiologic patients. With DB a high-skilled practitioner can swiftly alter the algorithm while processing a complex gauging; it is also possible to make corrections in the reference data, to predetermine the stages and objects of analysis and to use applications in dealing with strategic tasks. Information is supplied from BD in the form of reports generated by Microsoft Access, it is integrated with other MS Office tools, which makes it accessible to layman users. PMID- 15656407 TI - [Correlation between the precision of analogue-to-digital converter and discretion frequency at the P wave base of the electrocardiogram signal]. AB - Described is the interplay between the quantifications of the ECG signal by time, on the one hand, and by amplitude, on the other hand, at the P wave base. Ratios are suggested that can be used to choose a precision of the analogue-to-digital converter for preset values of P-wave parameters ensuring the detection of the P wave with an error not exceeding one time-quantification pitch. PMID- 15656408 TI - [The results of the monitoring of characteristics of digital receivers for X-ray images by using enterprise (VNIIMT) reference standard 01-22-04]. AB - Described are the results of testing of methods of the new 01-22-04 standard designed for monitoring the image-quality parameters of digital X-ray converter receivers (made alongside with tests of digital X-ray diagnostic units ProScan 7000 and ProScan-2000). PMID- 15656409 TI - [The study of the possible use of image reconstruction algorithms based on integral radon changes in small-angle X-ray computer tomography]. AB - Described are the results of a study of how a quantity of aspect angles affects the computer algorithm (deprojection with convolution filtration _ DCF) as well as of how symmetry and asymmetry in the aspect-angle position affect the space- and density resolutions of tomographic images. Examinations were made by using the reconstruction algorithm model (DCF) with fan X-ray geometry. PMID- 15656411 TI - [Portable device for ribs traction]. AB - Described in the paper is a portable device designed by the author for the traction of ribs in the complex treatment of multiple rib fractures. It is autonomous and easy in use. The devise does not damage the skin and tissues. PMID- 15656410 TI - [Modern evaluation of the accuracy of determination of the acid-forming function of the stomach by the method of intragastric pH-metry]. AB - Described are the results of research of how the standard location (antral region stomach body-cardium) of the comparison electrode in the 3-electrode probe affects the clinical evaluation of the acid-forming function of the stomach (AFC) in patients with gastric and duodenal ulcer. It was found that a sufficient accuracy (+/-0.2) in measurements of basal pH of the stomach body and, consequently, in clinical evaluation of AFC for the main standard locations is ensured by pH-probes with the chlorosilver skin comparison electrode. PMID- 15656412 TI - [Interactive input and correction of data about the structure of irradiated organism in the planning of radiotherapy for malignant tumor]. AB - A method is suggested for the interactive input, conversion and presentation of data about the structure of irradiated organism preset as Data Reference Planes (DRP). The possibility to build a library of DRP and the simplicity of their adjustment to each disease case witness to that the method of input, storage, conversion and presentation of data about the irradiated organism is promising in tumor radiotherapy. PMID- 15656413 TI - [Automated management of physical load in health prophylaxis]. AB - Described in the paper is a research case of the core of an automated support system for the decision-making in assigning a physical-load schedule that would take account of each specific patient and provide the practitioner with the related comprehensive data in diagnosis and therapy. PMID- 15656414 TI - [About optimized designs and circuits of autonomous electric stimulators for the gastrointestinal tract]. AB - Described in the paper are the key principles of designing of autonomous electrodes for the gastrointestinal tract (AE GT) as well as circuits of stimulating-pulse generators. A shape for the electric-stimulator frame, its geometric dimensions and choice of a material for electrodes are substantiated. The electric- and trauma-safety of AE GT is discussed. The main stimulating current parameters, as well as the flowchart and design of the electric stimulator are presented. PMID- 15656415 TI - [Checking of the automated collimation system of the X-ray beams in diagnostic equipment]. PMID- 15656416 TI - [Use of blue and green systems of image visualization in roentgenology]. AB - The main peculiarities of two image visualization systems related with the specificity of intensifying screens and of radiographic films in each of them are discussed. Specific features of kinetic development of modern orthochromatic general-purpose radiographic films were studied versus those of the traditional films; differences related with radiation hardness of some of the intensifying screen manufactured in Russia were investigated. Some practical advice was suggested on the basis of a conducted analysis of the "green" system specificity; such advice provides for reorienting the X-ray examination room, in Russia, for gadolinium screens and modern radiography films. PMID- 15656417 TI - [The experience in using of the induced otoacoustic emission in the practice of audiologist]. AB - Described in the paper is the possibility of using the induced otoacoustic emission (registered by different devices) in the practice of audiologist. The devices used for the purpose confirm the method as promising. PMID- 15656418 TI - [The world prepares for influenza epidemics]. PMID- 15656419 TI - [Research of psychotherapy is clinical research]. PMID- 15656420 TI - [Therapy of mind is a form of art, can one make science out of art?]. PMID- 15656421 TI - [Research of effectiveness is a challenge in psychotherapy]. PMID- 15656422 TI - [Processing of visual data in the cerebral cortex]. PMID- 15656423 TI - [Circadian clocks]. PMID- 15656424 TI - [Attention deficit-hyperactivity disorder among children in a neuropsychiatric problem]. PMID- 15656425 TI - [Primary bone lymphomas. Experiences in Kuopio University Central Hospital during 1990-2003]. PMID- 15656426 TI - [Non-islet cell tumor hypoglycemia]. PMID- 15656427 TI - [Aroma therapy and massage for relieving symptoms of cancer patients]. PMID- 15656428 TI - [Repeated periodic fever of an aluminium factory worker]. PMID- 15656429 TI - [The theory behind medicine is leaking and draft comes through the windows]. PMID- 15656430 TI - The cost of good health. PMID- 15656431 TI - Presidential address. "Trilogy of Learning" - the 22nd Biennial Conference of the New Zealand Dental Association. PMID- 15656432 TI - Amelogenesis imperfecta--multidisciplinary management from eruption to adulthood. Review and case report. AB - Amelogenesis imperfecta (AI) is a group of hereditary conditions that affect enamel formation. It is associated with a high morbidity for the patients and may present major restorative and sometimes orthodontic challenges for the dental team. Early recognition followed by appropriate preventive and restorative care is essential in the successful management of AI. A multidisciplinary approach with careful planning from early childhood will maximise the treatment options available for the permanent dentition and optimise the final outcome. In this case, a team consisting of two paediatric dentists, an orthodontist, a restorative dentist, and an oral and maxillofacial surgeon were involved in the management of the patient over a 12-year period. Treatment included preventive advice, interim composite restorations, two phases of orthodontic treatment, orthognathic surgery and placement of cast crowns. The patient is extremely happy with the result. PMID- 15656433 TI - Dentinal crack incidence following ultrasonic vibration to intra-radicular posts. AB - OBJECTIVE: To determine the incidence of dentinal cracks on the root-face of extracted teeth containing intra-radicular posts subjected to prolonged ultrasonic vibration. Design & Setting This in-vitro experiment, conducted at the University of Manchester, UK, used post type and ultrasonic vibration as dependent variables. MATERIALS AND METHODS: Forty canine teeth were decoronated and root-filled with gutta-percha and sealer. A parallel shaped post-space of 1.4 mm diameter and 10 mm depth was prepared in each root. The teeth were grouped into morphologically similar pairs, and for each pair one tooth was randomly allocated to receive a zirconium ceramic post, the other a stainless steel post. The pairs of teeth were randomly assigned to a control or to a test group. The root-face was examined for dentine cracks or fractures with direct vision and at 20x magnification with transillumination. Samples were then stored in sterile water for one week. Ultrasonic vibration was applied continuously for 30 minutes to the top of the posts in the test groups. All samples were then re-examined for dentine cracks or fractures. RESULTS: Following storage, no new cracks were evident for the control groups. Seven new cracks occurred in the test groups (following application of ultrasonic vibration), six of which were on the root face of samples with ceramic posts. The difference in crack distribution between the two groups was significant (Fisher's Exact Test, p=0.029). CONCLUSION: Prolonged application of ultrasonic vibration to ceramic posts increased the incidence of root-face cracks. PMID- 15656434 TI - Reference manual 2004-2005. PMID- 15656435 TI - Oral health policies. PMID- 15656436 TI - Clinical guideline on oral and dental aspects of child abuse and neglect. AB - In all states, physicians and dentists recognize their responsibility to report suspected cases of abuse and neglect. The purpose of this statement is to review the oral and dental aspects of physical and sexual abuse and dental neglect and the role of physicians and dentists in evaluating such conditions. The statement also addresses the oral manifestations of sexually transmitted diseases and bite marks, including the collection of evidence and laboratory documentation of these injuries. In all 50 states, physicians and dentists are required to report suspected cases of child abuse and neglect to social service or law enforcement agencies. Physicians receive minimal training in oral health and dental injury and disease, and thus may not detect dental aspects of abuse or neglect as readily as they do child abuse and neglect involving other areas of the body. Therefore, physicians and dentists should collaborate to increase the prevention, detection, and treatment of these conditions. PMID- 15656437 TI - Clinical guideline on infant oral health care. PMID- 15656438 TI - Clinical guideline on adolescent oral health care. PMID- 15656439 TI - Clinical guideline on management of persons with special health care needs. PMID- 15656440 TI - Clinical guideline on periodicity of examination, preventive dental services, anticipatory guidance, and oral treatment for children. PMID- 15656441 TI - Clinical guideline on the role of dental prophylaxis in pediatric dentistry. PMID- 15656442 TI - Clinical guideline on fluoride therapy. PMID- 15656443 TI - Clinical guideline on behavior management. PMID- 15656444 TI - Clinical guideline on the elective use of minimal, moderate, and deep sedation and general anesthesia for pediatric dental patients. PMID- 15656445 TI - Clinical guideline on use of anesthesia-trained personnel in the provision of general anesthesia/ deep sedation to the pediatric dental patient. PMID- 15656446 TI - Clinical guideline on pediatric restorative dentistry. PMID- 15656447 TI - Guideline on pulp therapy for primary and young permanent teeth. PMID- 15656448 TI - Clinical guideline on management of acute dental trauma. PMID- 15656449 TI - Clinical guideline on management of the developing dentition in pediatric dentistry. PMID- 15656450 TI - Clinical guideline on acquired temporomandibular disorders in infants, children, and adolescents. PMID- 15656451 TI - Clinical guideline on record-keeping. PMID- 15656452 TI - Clinical guideline on appropriate use of antibiotic therapy. PMID- 15656453 TI - Clinical guideline on antibiotic prophylaxis for patients at risk. PMID- 15656454 TI - Clinical guideline on dental management of pediatric patients receiving chemotherapy, hematopoietic cell transplantation, and/or radiation. PMID- 15656455 TI - What difference can I make? Our Scholarship winners speak out. PMID- 15656456 TI - The open abdomen: aetiology, classification and current management strategies. AB - A new classification system has been devised for open abdominal wounds. This will help identify an appropriate management strategy, and indicate the associated morbidity and outcome. In all cases, early intervention is vital. PMID- 15656457 TI - Spotting the signs: a case of late diagnosed painful calciphylaxis. AB - Although the number of cases of calciphylaxis is increasing, it is often not diagnosed until a late stage, increasing the risk of mortality. A characteristic is purple, mottled and painful lesions, which have a tendency to become necrotic. PMID- 15656458 TI - Linguistic validation of the Cardiff Wound Impact Schedule into French, German and US English. AB - OBJECTIVE: Chronic leg wounds can represent a major health problem and have a negative emotional impact on patients' lives. The Cardiff Wound Impact Schedule (CWIS) was developed by the Wound Healing Research Unit at University of Wales College of Medicine to assess the quality of life in patients with chronic wounds. METHOD: To perform the linguistic validation (translation) of the CWIS into German, French and US English as a first step in making the questionnaire available globally. The standard linguistic validation process employed by the Mapi Research Institute was used to translate the CWIS into French and German. An adjusted process was used for the US English translation. RESULTS: The linguistic validation of the US English version was straightforward as it shares the same root language with the original UK version. The problems encountered in the French and German translations were mostly related to semantics and syntax. CONCLUSION: The CWIS is now available for further validation testing by the international community (the psychometric evaluation of each translated questionnaire). These translations will be useful to health-care providers undertaking formal quality-of-life assessments of patients with chronic leg wounds. PMID- 15656459 TI - A short report on the prevention of slippage in donor site dressings. AB - Gauze is often a component of donor site dressings, and its removal can cause patients severe pain. This study set out to examine the extent of the problem and to investigate whether the method of anchoring the dressing influences slippage. PMID- 15656460 TI - Matrix metalloproteinases: their function in tissue repair. AB - Matrix metalloproteinases are involved in all stages of cutaneous wound repair and play a key role in regulating the balance between tissue synthesis and tissue destruction. Greater understanding of this may result in better treatment options. PMID- 15656461 TI - An investigation of the long-term bioactivity of endogenous growth factor in OASIS Wound Matrix. AB - OBJECTIVE: To examine the retention and bioactivity of endogenous FGF-2 after prolonged storage within a complex matrix. FGF-2 is a growth factor found in OASIS Wound Matrix, a purified collagen wound-care product containing other endogenous components of the extracellular matrix. METHOD: FGF-2 content was measured by ELISA. FGF-2 activity was measured using an in vitro bioassay in rat pheochromocytoma (PC12) cells. RESULTS: FGF-2 content ranged from 15.3 ng/g to 84.3 ng/g. The bioassay showed that the FGF-2 retained in the matrix was present in a bioactive form able to cause differentiation of cells in culture. CONCLUSION: The results demonstrate that wound-care products can be developed to retain their bioactivity over time and that inherently unstable purified growth factors are preserved if stored as bound factors within their natural extracellular matrix. The results also suggest that use of acellular matrices containing active growth factors would have advantages in terms of simplicity and cost over purified recombinant growth factor therapies. PMID- 15656462 TI - Choice of dressing has a major impact on blistering and healing outcomes in orthopaedic patients. AB - OBJECTIVE: To investigate the effect of three postoperative dressings on orthopaedic wound healing. METHOD: Three hundred orthopaedic patients were divided into three treatment groups and allocated to management with one of three dressings: Primapore, Tegaderm with pad, and OpSite Post-Op. Staff completed a questionnaire to evaluate the wound progression. Outcome measures were the presence of infection, blistering and the number of dressing changes required. RESULTS: There was a significantly lower incidence of blistering with OpSite Post Op (6%) than Tegaderm with pad (16%) and Primapore (24%) (p<0.001). Patients in the OpSite Post-Op group had the lowest exudate levels. CONCLUSION: Dressings that employ a clear film and have a high moisture vapour transmission rate have been shown to reduce both the rate of blistering and wound discharge. The additional expense inherent in using such dressings may, in reality, prove cost effective because of the reduced need for dressings changes and the subsequent earlier discharge of these patients from hospital with an uncomplicated wound. PMID- 15656463 TI - Wound-contact layers: a re-evaluation. PMID- 15656464 TI - Silver dressings: the need for clarity. PMID- 15656465 TI - Prognostic factors for venous ulcer healing in a non-selected population of ambulatory patients. AB - OBJECTIVE: To identify wound prognostic indicators in a non-selected patient population with leg ulcers. METHOD: This was a prospective observational survey involving 151 physicians. Ambulatory patients with venous leg ulcers were treated with a non-adherent foam dressing and usual leg ulcer management. At follow-up after three to six weeks, ulcer healing parameters and local treatment tolerability were recorded and concordance with compression therapy was validated. RESULTS: A > or = 40% reduction in the largest wound length was selected as an indicator of a favourable healing outcome. A total of 330 patients were included and seen at follow-up. The mean largest wound length was 4.9 +/- 3.6 cm. A > or = 40% decrease in this parameter was noted in 178 patients. Dressing acceptability and tolerability were excellent. Compression therapy, while heterogeneous among physicians, was used regularly and applied correctly (by patients and practitioners) in 81.2% of patients. Predictors for not reaching the main outcome were the presence of an arterial lower limb disease, an ulcer duration of more than three months and an initial ulcer length of 10 cm or more. Linear regression showed that old age and a high body mass index were independent predictors of a poor outcome. CONCLUSION: Clinical indicators such as simple ulcer length measurement and ulcer duration may help physicians to detect patients with a prognosis of poor healing. PMID- 15656467 TI - Prophylactic antibiotic use in clean neurosurgery: of potential benefit or harm to the patient? AB - The increasing occurrence of antibiotic resistance is causing alarm, and efforts are being made to prevent the inappropriate use of antibiotics. But discussion on whether or not prophylaxis is needed in clean neurosurgery continues. PMID- 15656466 TI - The prevalence of pressure ulcers in hospitalised patients in a university hospital in India. AB - OBJECTIVE: To estimate the prevalence of pressure ulcers in hospitalised patients and any underlying or predisposing factors to ulceration. METHOD: This cross sectional study took place in a university hospital in Varanasi, India. A total of 445 patients hospitalised in medical and surgical wards were examined in a single day for the number, site and grade of pressure ulcers. Haemoglobin, serum albumin and blood sugar levels of patients with pressure ulcers were recorded. RESULTS: The prevalence of pressure ulcers was high (4.94%). Anaemia, malnutrition and diabetes were important risk factors, while morbidity due to pressure ulcers in long-stay wards, such as neurology, was exceptionally high (40.9%). CONCLUSION: Pressure ulcers remain one of the most neglected aspects of health-care provision in India and identifying their associated risk factors at an early stage may go a long way in preventing their occurrence. PMID- 15656469 TI - The doctor's dilemma. PMID- 15656468 TI - The effectiveness of a hydrogel dressing compared with standard management of pressure ulcers. AB - OBJECTIVE: To compare a hydrogel dressing with a conservative topical wound-care regimen (gauze with povidone-iodine) for the treatment of pressure ulcers. METHOD: A prospective study comprising 27 (24 male and three female) spinal-cord injury patients with a total of 49 pressure ulcers. Wounds were randomised into a treatment group (n=25) or control group (n=24). The primary outcome measure was the rate of wound healing (cm2/days). RESULTS: Healing rates in the treatment group were higher than in the control group, although this was not statistically significant. Some 84% of the wounds in the treatment group and 54% in the control group epithelialised, which was statistically significant (p=0.04). The mean healing time in the treatment group was 48 days compared with 45 days for the controls. CONCLUSION: Our results suggest that hydrogel occlusive wound dressings facilitate healing by promoting more rapid epithelialisation of pressure ulcers, when compared with conservative wound care. PMID- 15656470 TI - Clinical academic medicine: the way forward. PMID- 15656471 TI - Infective endocarditis--new guidance recommends a more aggressive approach. PMID- 15656472 TI - NICE guidance on type 1 diabetes in adults. PMID- 15656473 TI - Recently introduced antibiotics: a guide for the general physician. PMID- 15656474 TI - Management of meningitis. PMID- 15656475 TI - The returned traveller. PMID- 15656476 TI - Bone and joint infection. PMID- 15656477 TI - Intra-abdominal infections. PMID- 15656478 TI - The competent doctor: a paper for discussion. AB - The clinical competence of doctors needs to be defined and assessed in relation to the circumstances of individual practice, in terms of knowledge and skills, both clinical and communicative, as well as attitudes and behaviour. The need to identify the role of the medical practitioner in the context of team working is important and this position requires the skills of diagnosis, synthesis of information and prioritisation of investigation and treatment. Completion of training and acquisition of the Certificate of Completion of Specialist Training (CCST) lead to inclusion in the Specialist Register thus defining the specialist; elevation of a specialist to consultant status in the NHS requires the potential to acquire with maturity, both of person and experience, the confidence to take responsibilities for handling difficult situations, to manage uncertainties in clinical encounters and to guide younger doctors in their careers. PMID- 15656479 TI - The Assisted Dying for the Terminally Ill Bill, 2004. PMID- 15656480 TI - Symptoms and the perception of disease. AB - Any symptom represents a perception of an abnormal internal body state. The threshold for perceiving the internal body state as abnormal varies and depends particularly on psychological influences. As a result, a symptom can either reflect pathology, whether serious or not, or be generated wholly psychologically. Intuition allowing discrimination between these possibilities is central to the physician's art. Particular difficulty arises in differentiating between those psychologically generated symptoms which are produced unconsciously, often as a result of anxiety or depression, and those that constitute deliberate deception. Such malingering has the unstated intent of accessing a secondary gain, such as welfare benefit. The art of diagnosis includes estimation of whether symptoms resonate with known pathophysiological processes, using history-taking as a story which unfolds logically towards a diagnosis, assessing how a patient reacts to their symptoms compared to neutral matters, detecting exaggeration or falsification, and documenting evidence of psychologically generated abnormalities during examination. Scientific ability is only one of the attributes of a good diagnostician; equally important are abilities to notice things, to weigh up human nature and to recognise dilemmas. Our procedures for selecting medical students and physicians need to assess these skills as well as scientific qualifications. PMID- 15656481 TI - Prophylaxis and treatment of infective endocarditis in adults: a concise guide. AB - Infective endocarditis (IE) is a life-threatening disease with substantial morbidity and mortality which affects individuals with underlying structural cardiac defects who develop bacteraemia, often as a result of dental, gastrointestinal, genitourinary, respiratory or cardiac invasive/surgical procedures. Prompt recognition of the clinical diagnosis by a wide variety of medical personnel, early involvement of specialist cardiologists, cardiac surgeon and a microbiologist, and prompt treatment with the most appropriate antimicrobial agents offer the greatest chance of improving the outcome for these patients. The guidance given here to clinicians involved in the management of patients with IE briefly covers diagnosis, antibiotic prophylaxis, medical treatment and the indications for surgery. PMID- 15656482 TI - Chronic obstructive pulmonary disease, neutrophils and bacteria: from science to integrated care pathways. AB - The incidence of chronic obstructive pulmonary disease (COPD) and its effect on morbidity and mortality is rising. Extensive research implicates the role of inflammation and particularly the effect of neutrophil proteinases in generating many of the pathological features seen as part of the COPD syndrome. Exacerbations of COPD are an important cause of morbidity, mortality and progressive deterioration in lung function. These episodes are poorly defined but bacteria in the airways drives neutrophilic inflammation and proteinase release and hence may play a key role. The differentiation between benign colonisation and heightened neutrophilic inflammation can be determined by observation of sputum purulence. This reflects the likelihood of bacteria being present and particularly the number of bacteria. This simple clinical observation enables decision making about the need for antibiotic therapy to be as much a part of patient self-management as physician directed management in the integrated care pathway for COPD. PMID- 15656483 TI - Fluid, electrolytes and nutrition. AB - Nutrition and fluid and electrolyte balance are inextricably linked through ingestion, digestion, absorption and intermediary metabolism. Studies are described showing that man's physiological capacity to excrete excess salt and water is limited under normal conditions; it is further reduced by starvation and injury, so that patients are easily overloaded, resulting in increased complications of illness and surgery. Return of gastrointestinal function postoperatively is delayed by moderate saline overload. Illness not only influences external fluid and electrolyte balance but also internal balance between the extracellular and intracellular spaces and between the intravascular and extravascular components of the extracellular space. The mechanisms and management of these changes are discussed. The importance of fluid and electrolyte balance in nutritional support is emphasised--indeed, the benefits of nutrition may be negated by electrolyte imbalance. The relationships between serum albumin concentration, illness, nutrition and fluid balance are discussed and the limited role of albumin infusions described. Surveys among doctors have revealed a poor knowledge of fluid and electrolyte balance. Measures are needed to improve training. PMID- 15656484 TI - Treatment in art. PMID- 15656485 TI - A lateral gaze at pain: the science of attention. PMID- 15656486 TI - Mentoring. PMID- 15656487 TI - The 2001 guidelines on cardiopulmonary resuscitation. PMID- 15656488 TI - Excessive daytime sleepiness and driving: regulations for road safety. PMID- 15656489 TI - Excessive daytime sleepiness and driving: regulations for road safety. PMID- 15656490 TI - Effects of 6 degree elevation of the heels on 3D kinematics of the distal portion of the forelimb in the walking horse. AB - REASONS FOR PERFORMING STUDY: Understanding of the biomechanical effects of heel elevation remains incomplete because in vivo studies performed with skin markers do not measure the actual movements of the 3 digital joints. OBJECTIVE: To quantify the effects of 6 degree heel wedge on the 3-dimensional movements of the 4 distal segments of the forelimb in the walking horse. METHODS: Four healthy horses were used. Kinematics of the distal segments was measured invasively with a system based on ultrasonic triangulation. Three-dimensional rotations of the digital joints were calculated by use of a 'joint coordinate system' (JCS). Data obtained with heel wedges were compared to those obtained with standard shoes during the stance phase of the stride. RESULTS: Heel wedges significantly increased maximal flexion of the proximal (PIPJ) and distal (DIPJ) interphalangeal joints and maximal extension (mean +/- s.d. +0.8 +/- 0.3 degrees) of the metacarpophalangeal joint (MPJ). Extension of the PIPJ and DIPJ was decreased at heel-off. Few effects were observed in extrasagittal planes of movement. CONCLUSIONS: Heel wedges affect the sagittal plane kinematics of the 3 digital joints. POTENTIAL RELEVANCE: Controversial effects previously observed on the MPJ may be explained by the substantial involvement of the PIPJ, which was wrongly neglected in previous studies performed on the moving horse. PMID- 15656491 TI - Verification of skin-based markers for 3-dimensional kinematic analysis of the equine tarsal joint. AB - REASONS FOR PERFORMING STUDY: Kinematic studies are usually based on tracking markers attached to the skin. However, complex joints, such as the tarsal joint, function in 3-dimensions (3D), and have therefore necessitated application of the invasive bone pin technique, limiting kinematic studies to the research laboratory. This study investigates the feasibility of using skin-based markers for 3D analysis of tarsal joint motion. HYPOTHESIS: Three-dimensional motions of the tarsal joint can be measured with an acceptable degree of accuracy using skin markers. METHODS: Retroreflective markers were attached over the tibial and metatarsal segments. Markers were tracked automatically at trot. Three dimensional skin correction algorithms were used for correction of skin displacement, and 3D motions derived from the corrected (CSD) and uncorrected (USD) skin displacement were compared with data from a previous study in which those motions were described using bone-fixed markers (BFM) by correlation, root mean square errors (RMS) and shape agreement (SA) of the curves. RESULTS: The RMS of BFM and CSD were smaller than those of BFM and USD for all motions. The correlation coefficients of BFM and CSD were higher than those of BFM and USD. SA was good or fair for all motions except internal/external rotation and medial/lateral translation. CONCLUSIONS AND POTENTIAL RELEVANCE: With appropriate correction for skin movement relative to skeletal landmarks, skin markers can identify tarsal 3D motions for flexion/extension, abduction/adduction, cranial/caudal translation, and proximal/distal translation, allowing analysis and comparison of information between horses during swing and stance phases. PMID- 15656492 TI - Three-dimensional kinematic analysis of horses with induced tarsal synovitis. AB - REASONS FOR PERFORMING STUDY: Techniques for 3D analysis have recently been developed. This study applied 3D kinematic analysis to describe the effects of lameness in the distal intertarsal (DIT) and tarsometatarsal (TMT) joints; complex joints such as this may show measurable amounts of movement outside the sagittal plane that may change in response to pathological conditions. HYPOTHESIS: Three-dimensional motions of the tarsal joint change after the induction of synovitis at the DIT and TMT joints. METHODS: Twelve retroreflective markers, attached on the skin over the right limb, were used to develop 3D coordinate systems for tibial and metatarsal segments. Data were collected at trot from 4 horses on 2 occasions, before (sound condition) and after (lame condition) induction of synovitis of the DIT and TMT joints. Ranges of 3D motions of the metatarsal segment relative to the tibial segment were compared between sound and lame conditions using t tests. RESULTS: There were significant decreases (P<0.05) in tarsal joint flexion and cranial translation of the metatarsus relative to the tibia during stance, and in proximal translation of the metatarsus during swing. CONCLUSIONS AND POTENTIAL RELEVANCE: In the lame condition, reduced cranial translation at the DIT and TMT joints may lead to a reduction in the motion of the articular cartilage surfaces relative to each other, which may have adverse effects on cartilage nutrition and function. Further studies of tarsal 3D kinematics in horses with bone spavin could allow quantification of the biomechanical effects of the special shoes used for conservative treatment. PMID- 15656493 TI - Three-dimensional analysis of patterns of skin displacement over the equine radius. AB - REASONS FOR PERFORMING STUDY: Surface markers are usually used to track bone movement. However, skin movement related to the bone has a large effect on the analysis of skeletal kinematics. A 2-dimensional (2D) skin displacement correction model has been successfully developed, but no 3D skin displacement model exists. OBJECTIVES: To develop a 3-dimensional (3D) skin displacement model for the equine radial segment during trot. METHODS: The 3D trajectories of 6 skin based markers and a bone-fixed triad were captured at trot in 4 horses. Skin displacements in the bone-based coordinate system were calculated using a singular-value decomposition method. The truncated Fourier series models were developed for the skin displacements using a generalised cross-validatory spline. RESULTS: Mean + /- s.d. of peak skin displacement of the 3 markers on the proximal radius as percentage of radial length was 10.7 +/- 0.5, 4.6 +/- 1.5 and 14.5 +/- 2.9% in x, y and z direction, respectively. For the 3 markers on the distal radius, the equivalent displacements were 4.7 +/- 0.6, 1.7 +/- 0.8 and 7.3 +/- 18% in x, y and z direction, respectively. CONCLUSIONS: The 3D skin displacement model for correction of skin marker motion over the equine radius relative to the bone can be established using a truncated Fourier series, which has previously been used successfully to develop 2D models. POTENTIAL RELEVANCE: This method of determining 3D skin displacement correction needs to be extended to the entire fore- and hindlimbs to provide a more sensitive measure of kinematic analysis. Accurate descriptions of the 3D motions of the limb segments and interactions between adjacent segments at the joints are necessary for understanding of the mechanics of different gaits and the gait aberrations that manifest as lameness. PMID- 15656494 TI - Three-dimensional carpal kinematics of trotting horses. AB - REASONS FOR PERFORMING STUDY: Descriptions of 3D kinematics assist in understanding joint function and dysfunction, and are an essential step toward 3D inverse dynamic analysis. OBJECTIVES: To measure 3D carpal joint motion during trotting. METHODS: Three-dimensional trajectories of bone-fixed markers on the radius and third metacarpus of the right forelimb of 3 healthy horses were recorded at 120 Hz using a 6-camera analysis system. Joint kinematics were calculated in terms of helical angles between the 2 segments using a spatial attitude method. RESULTS: All horses showed carpal extension and internal rotation of the metacarpus relative to the radius as the carpus assumed the close packed position. In late stance, the carpus began a cycle of flexion that continued through midswing, accompanied by a small cycle of internal rotation. The direction of abduction/adduction varied between horses. The predominant rotational movement was flexion/extension, which showed a range of motion of 15 +/- 6 degrees in stance and 76 +/- 13 degrees in swing. CONCLUSIONS: Carpal motions were generally similar between horses with the exception of abduction/adduction. POTENTIAL RELEVANCE: Knowledge of carpal joint motion should assist in understanding the pathogenesis of carpal injuries. However, it seems probable that real differences exist between individuals; therefore, further investigations of the effect of conformation on carpal motion should be performed in a much larger population of horses. PMID- 15656495 TI - Development of a 3D model of the equine distal forelimb and of a GRF shoe for noninvasive determination of in vivo tendon and ligament loads and strains. AB - REASONS FOR PERFORMING STUDY: As critical locomotion events (e.g. high-speed and impacts during racing, jump landing) may contribute to tendinopathies, in vivo recording of gaits kinematic and dynamic parameters is essential for 3D reconstruction and analysis. OBJECTIVE: To propose a 3D model of the forelimb and a ground reaction force recording shoe (GRF-S) for noninvasively quantifying tendon and ligament loads and strains. METHODS: Bony segments trajectories of forelimbs placed under a power press were recorded using triads of ultrasonic kinematic markers linked to the bones. Compression cycles (from 500-6000 N) were applied for different hoof orientations. Locations of tendon and ligament insertions were recorded with regard to the triads. The GRF-S recorded GRF over the hoof wall and used four 3-axis force sensors sandwiched between a support shoe and the shoe to be tested. RESULTS: Validation of the model by comparing calculated and measured superficial digital flexor tendon strains, and evaluation of the role of proximal interphalangeal joint in straight sesamoidean ligament and oblique sesamoidean ligament strains, were successfully achieved. Objective comparisons of the 3 components of GRF over the hoof for soft and hard grounds could be recorded, where the s.d. of GRF norm was more important on hard ground at walk and trot. CONCLUSIONS: Soft grounds (sand and rubber) dissipate energy by lowering GRF amplitude and diminish bounces and vibrations at impact. At comparable speed, stance phase was longer on soft sand ground. POTENTIAL RELEVANCE: The conjugate use of the GRF-S and the numerical model would help to quantify and analyse ground/shoe combination on comfort, propulsion efficiency or lameness recovery. PMID- 15656496 TI - In vitro measurement of internal hoof strain. AB - REASONS FOR PERFORMING STUDY: Strains during stance on the hoof wall surface have been measured by a number of authors in vitro and in vivo. Histological structure and mechanical properties vary through the wall thickness (radially); radial strain measurements may therefore aid the understanding of mechanical function of the capsule and adjacent tissues. OBJECTIVES: To develop instrumentation capable of measuring internal hoof strain, and to carry out a preliminary comparison of normal and laminitic hooves. METHODS: Six forelimbs from 4 horses, including 2 with laminitis from the same horse, were tested using an Instron test rig designed to simulate the walk at impact, midstance and breakover. Internal strains were measured at a dorsal site using strain gauges moulded into a plug made of 007 fast-set structural adhesive. In addition, kinetic and kinematic data were collected from each specimen. RESULTS: When simulating the walk, a significant (P<0.0001) increase in gradient of radial tensile strain was found in a normal hoof wall, from 5.6 +/- 73.9 microepsilon at the outer gauge to 418.5 +/ 170.6 microepsilon at the inner gauge. However, radial strains measured at the inner gauge site in limbs with laminitis were found to be significantly (P<0.0001) compressive, with values of -406.7 +/- 156.3 and -109.9 +/- 72.4 microepsilon for Specimens 1 and 2, respectively. CONCLUSIONS AND POTENTIAL RELEVANCE: These preliminary data indicate that a marked redistribution may well occur in the wall of laminitic hooves. With a larger sample size, the results should have relevance to the treatment and management of laminitis. PMID- 15656497 TI - Surface strains around the midshaft of the third metacarpal bone during turning. AB - REASONS FOR PERFORMING STUDY: Bone strains quantify skeletal effects of specific exercise and hence assist in designing training programmes to avoid bone injury. OBJECTIVE: To test whether compressive strains increase on the lateral surface of the inside third metacarpal bone (McIII) and the medial surface of the outside McIII in a turn. METHODS: Rosette strain gauges on dorsal, medial and lateral surfaces of the midshaft of the left McIII in 2 Thoroughbred geldings were recorded simultaneously during turning at the walk on a bitumen surface. RESULTS: Medial surface: Compression peaks were larger in the outside limb. Tension peaks were larger in the inside limb and in a tighter turn. On the lateral surface compression and tension peaks were larger on the inside limb, which showed the largest recorded strains (compression of -1400 microstrains). Dorsal compression strains were larger on the outside limb and on a larger circle. Tensile strains were similar in both directions and larger on a larger circle. CONCLUSIONS: Compressive strains increased on the lateral surface of the inside McIII and medial surface of the outside McIII in a turn. POTENTIAL RELEVANCE: Slow-speed turning exercise may be sufficient to maintain bone mechanical characteristics in the inside limb lateral McIII cortex. Further work is needed to confirm these findings and to determine whether faster gaits and/or tighter turns are sufficient to cause bone modelling levels of strain in the medial and lateral McIII cortex. PMID- 15656498 TI - In vitro assessment of movements of the sacroiliac joint in the horse. AB - REASONS FOR PERFORMING STUDY: Sacroiliac joint (SIJ) disease is associated with poor hindlimb action, lameness and poor performance in horses. However, little is known about the biomechanics of this low-motion joint. OBJECTIVES: To determine in vitro the capacities of movement of the SIJ in the sagittal plane, and to test the effect of a sacrosciatic and sacrotuberal desmotomy on its stabilisation. METHODS: Six anatomical specimens underwent cycles of flexion-extension of the lumbosacral joint (LSJ) before and after desmotomy. Kinematic triads were linked rigidly to the sacrum, spinous process of vertebra L5 and iliac wing. Angles were measured using a joint coordinate system based on anatomical frames. RESULTS: The LSJ underwent regular movements of flexion and extension (overall mean +/- s.d. range 23.4 +/- 1.6 degrees). The only recorded movement of the SIJ was a nutation during LSJ flexion (overall mean +/- s.d. 0.8 +/- 0.5 degrees). Desmotomy induced an increase of that nutation (overall mean +/- s.d. 1.7 +/- 0.2 degrees). CONCLUSIONS AND POTENTIAL RELEVANCE: Movements of the SIJ were small and coupled only with the flexion of the LSJ. The ligaments surrounding the SIJ have a strong effect on the stabilisation of this joint. Due to the limited amount of movement, its biomechanical study in vivo seems to be difficult. Further in vitro studies would be useful to determine the role of each ligament, to better understand the clinical consequences of the tears frequently observed during necropsy. PMID- 15656499 TI - Determination of the stiffness of the equine spine. AB - REASONS FOR PERFORMING STUDY: Mechanical properties of the equine back are the bases for realistic modelling of the back, which is recognised as an important step towards improved understanding of the pathogenesis of equine back conditions. HYPOTHESIS: The stiffness of the equine back depends on the direction of the applied force and on the position of the spine. METHODS: Fourteen dissected spines were tested in a tensile testing machine. In 3 different positions, simulating dorsoventral, laterolateral and dorsoventral-30 degree rotated movement, force was applied on the dorsal spinous process of T12 to reach an excursion of 4 cm in each direction. The normal distribution of the stiffness coefficients was tested with a Kolmogorov-Smirnov test and the stiffness coefficients were compared with paired t tests. RESULTS: Mean +/- s.d. dorsoventral stiffness was 2093 +/- 611 N/m for the nonrotated spine and 2182 +/- 459 N/m for the 30 degree rotated spine. Mean laterolateral stiffness was significantly lower than dorsoventral stiffness at 1454 +/- 156 N/m. CONCLUSIONS: The stiffness of the spine depends on the direction of loading. POTENTIAL RELEVANCE: The stiffness of the spine under loading may be a relevant factor in the development of back disorders. PMID- 15656500 TI - Influence of the load of a rider or of a region with increased stiffness on the equine back: a modelling study. AB - REASONS FOR PERFORMING STUDY: Knowledge of load effects is crucial for the understanding of the aetiology and pathogenesis of equine back problems. OBJECTIVE: To investigate different load scenarios of the equine back, such as being ridden or increased muscle tone, using biomechanical simulations. METHODS: Kinetic and kinematic data of 15 sound horses and the electromyelograph of their long back muscles were recorded. A biomechanical simulation model was used for simulations under different biomechanical scenarios (ridden/unridden, localised increased stiffness) using ADAMS. RESULTS: The vertical forces acting through a rider were: walk 3.83 N/kg, trot 5.18 N/kg and gallop 5.60 N/kg. No significant changes in transversal forces were found between ridden and unridden horses. Profound changes were seen in the torques at the segment following a region of increased stiffness: in walk, lateral peak torques increased from 342 to 1723 Nm, and in trot from 393 to 1004 Nm, and dorsoventral from 386 to 3705 Nm (walk) and 458 to 4340 Nm (trot). CONCLUSIONS AND POTENTIAL RELEVANCE: The simulation shows that the stress of a rider is lower than that of pathological processes such as partial increased stiffness of the back. Study of revised models with improved anatomical realism might help to raise the plausibility of model results. PMID- 15656501 TI - Kinematic evaluation of the back in the sport horse with back pain. AB - REASONS FOR PERFORMING STUDY: Earlier studies have developed a clinical tool to evaluate objectively the function of the equine back. The ability to differentiate horses with back pain from asymptomatic, fully functioning horses using kinematic measures from this tool has not been evaluated. OBJECTIVES: To compare the kinematics of the back at walk and trot in riding horses with back dysfunction to the same parameters in asymptomatic sport horses. METHODS: The kinematics of the back in 12 horses with impaired performance and back pain were studied at walk and trot on a treadmill. Data were captured for 10 sees at 240 Hz. Range of movement (ROM) and intravertebral pattern symmetry of movement for flexion and extension (FE), lateral bending (LB) and axial rotation (AR) were derived from angular motion pattern data and the results compared to an earlier established database on asymptomatic riding horses. RESULTS: At walk, horses with back dysfunction had a ROM smaller for dorsoventral FE in the caudal thoracic region (T13 = 7.50 degrees, T17 = 7.71 degrees; P<0.05), greater for LB at T13 (8.13 degrees; P<0.001) and smaller for AR of the pelvis (10.97 degrees; P<0.05) compared to asymptomatic horses (FE-T13 = 8.28 degrees, FE-T17 = 8.49 degrees, LB T13 = 6.34 degrees, AR-pelvis = 12.77 degrees). At trot, dysfunctional horses had a smaller (P<0.05) ROM for FE at the thoracic lumbar junction (T17 = 2.46 degrees, L1 = 2.60 degrees) compared to asymptomatic horses (FE-T17 = 3.07 degrees, FE-L1 = 3.12 degrees). CONCLUSIONS: The objective measurement technique can detect differences between back kinematics in riding horses with signs of back dysfunction and asymptomatic horses. The clinical manifestation of back pain results in diminished flexion/extension movement at or near the thoracic lumbar junction. However, before applying the method more extensively in practice it is necessary to evaluate it further, including measurements of patients whose diagnoses can be confirmed and long-term follow-ups of back patients after treatment. POTENTIAL RELEVANCE: Since the objective measurement technique can detect small movement differences in back kinematics, it should help to clinically describe and, importantly, objectively detect horses with back pain and dysfunction. PMID- 15656502 TI - Detection of spinal ataxia in horses using fuzzy clustering of body position uncertainty. AB - REASONS FOR PERFORMING STUDY: Subjective neurological evaluation in horses is prone to bias. An objective method of spinal ataxia detection is not subject to these limitations and could be of use in equine practice and research. HYPOTHESIS: Kinematic data in the walking horse can differentiate normal and spinal ataxic horses. METHODS: Twelve normal and 12 spinal ataxic horses were evaluated by kinematic analysis walking on a treadmill. Each body position signal was reduced to a scalar measure of uncertainty then fuzzy clustered into normal or ataxic groups. Correct classification percentage (CCP) was then calculated using membership values of each horse in the 2 groups. Subsequently, a guided search for measure combinations with high CCP was performed. RESULTS: Eight measures of body position resulted in CCP > or = 70%. Several combinations of 4-5 measures resulted in 100% CCP. All combinations with 100% CCP could be obtained with one body marker on the back measuring vertical and horizontal movement and one body marker each on the right fore- and hindlimb measuring vertical movement. CONCLUSIONS AND POTENTIAL RELEVANCE: Kinematic gait analysis using simple body marker combinations can be used objectively to detect spinal ataxia in horses. PMID- 15656503 TI - The role of electromyography in clinical diagnosis of neuromuscular locomotor problems in the horse. AB - REASONS FOR PERFORMING STUDY: Systematically performed EMG needle examination of muscles provides essential information about the functional aspects of the motor unit. However, clinical studies in which information is given on the diagnostic and discriminative values of electromyography (EMG) in the horse are scarce. OBJECTIVES: To determine to what extent inclusion of EMG analysis in clinical examination contributes to determination of type and localisation of abnormality. METHODS: EMG analysis, complete clinical examination and diagnosis of 108 horses (mean +/- s.d. age 75 +/- 3.8 years; bodyweight 548 +/- 86 kg; height 1.67 +/- 0.07 m) were performed, and results without and with EMG analysis compared. RESULTS: Without EMG, myopathy and neuropathy were diagnosed in 20 and 58 horses, respectively, and with EMG in 17 and 82 horses. EMG changed localisation in myopathy and neuropathy in 12 and 37% of cases, respectively. Lesions in the C1 T2, T2-L3 and L3-S3 segments were, respectively, diagnosed without EMG in 7, 11 and 30%, and with EMG in 27, 7 and 17% of cases. Where no clinical diagnosis could be made prior to EMG, many patients appeared to be suffering from localised cervical lesions (29%) or generalised neuropathy (54%). CONCLUSIONS AND POTENTIAL RELEVANCE: The assistance of EMG in discriminating between normal, neuropathy and myopathy, and in locating pathology, contributes to diagnosis of neuromuscular problems. PMID- 15656505 TI - Compensatory load redistribution of horses with induced weightbearing hindlimb lameness trotting on a treadmill. AB - REASONS FOR PERFORMING STUDY: The compensatory mechanisms of horses with weightbearing hindlimb lameness are still not fully understood. HYPOTHESIS: That weightbearing, unilateral hindlimb lameness would not only alter stride characteristics to diminish structural stress in the affected limb but also induce compensatory load adjustments in the other supporting limbs. OBJECTIVE: To document the load and time shifting mechanisms of horses with unilateral weightbearing hindlimb lameness. METHODS: Reversible lameness was induced in 8 clinically sound horses by applying a solar pressure model. Three degrees of lameness (subtle, mild and moderate) were induced and compared with the nonlame (sound) control measurement. Vertical ground reaction forces were recorded for all 4 limbs simultaneously on an instrumented treadmill. RESULTS: Compared to the sound situation, moderate hindlimb hoof lameness induced a decrease in stride duration (-3.3%) and stride impulse (-3.1%). Diagonal impulse decreased selectively in the lame diagonal stance (-7.7%). Within the diagonal limb pair, vertical impulse was shifted to the forelimb during the lame diagonal stance (+6.5%) and to the hindlimb during the sound diagonal stance (+3.2%). Peak vertical force and vertical impulse decreased in the lame limb (-15%), but only vertical impulse increased in the contralateral hindlimb (+5.7%). Stance duration was prolonged in both hindlimbs (+2.5%). Suspension duration was reduced to a greater extent after push-off of the lame diagonal limb pair (-21%) than after the sound diagonal limb pair (-9.2%). CONCLUSIONS: Four compensatory mechanisms could be identified that served to reduce structural stress, i.e. peak vertical force on the affected limb: 1) reduction of the total vertical impulse per stride; 2) diagonal impulse decreased selectively in the lame diagonal; 3) impulse was shifted within the lame diagonal to the forelimb and in the sound diagonal to the hindlimb; and 4) the rate of loading and peak forces were reduced by prolonging the stance duration. POTENTIAL RELEVANCE: Load shifting mechanisms are not only effective in diminishing peak forces in the affected limb, but also suppress compensatory overload in other limbs. Selected force and time parameters allow the unequivocal identification of the lame limb. Future studies have to examine how far these compensatory mechanisms may be generalised for other defined orthopaedic problems in the hindlimb. PMID- 15656504 TI - Mepivacaine local anaesthetic duration in equine palmar digital nerve blocks. AB - REASONS FOR PERFORMING STUDY: Perineural analgesics are used for lameness diagnosis but the duration of effect, knowledge of which would provide valuable information when performing subsequent blocks, is unknown. OBJECTIVE: To evaluate the duration of a palmar digital nerve block using force plate measurements. METHODS: Ten horses diagnosed with unilateral navicular syndrome were trotted at range of 3 +/- 0.15 m/sec over a force plate to record ground reaction forces for 5 trials of each forelimb. Data were recorded before nerve block, and then at 15 mins, 1, 2 and 24 h post nerve block. RESULTS: Before nerve block, peak vertical force (mean +/- s.e.) was significantly higher in the contralateral forelimb (CL = 5345 +/- 188 N) than in the lame forelimb (L = 4256 +/- 204 N; P<0.05). At 15 mins post nerve block there was no significant difference between the 2 forelimbs (CL = 5140 +/- 184 N; L = 5126 +/- 129 N), and this remained the case for 1 h. By 2 h, the mean score for the lame leg had decreased (L = 4642 +/- 182 N) but was still greater than preblock. By 24 h, vertical forces had returned to preblock values. CONCLUSIONS: The palmar digital nerve block was fully effective between 15 mins and 1 h. The analgesic effect began to subside between 1 and 2 h but sufficient analgesia persisted to affect gait characteristics beyond 2 h. POTENTIAL RELEVANCE: When using a palmar digital nerve block, it is important to perform lameness evaluations between 15 mins and 1 h to be sure of effective nerve blockade. PMID- 15656506 TI - Influence of rider on lameness in trotting horses. AB - REASONS FOR PERFORMING STUDY: Equine lameness is commonly evaluated when the horse is being ridden, but the influence of the rider on the lameness has not been documented. OBJECTIVE: To document the effect of 2 riders of different training levels on the vertical movement of the head and croup. METHODS: Twenty mature horses were ridden at trot by an experienced dressage rider and a novice rider, as well as trotted in hand. Kinematic measurements of markers placed on the horse's head and sacral bone were carried out. The asymmetries of the vertical head and sacral bone motion were calculated as lameness parameters and compared with paired t tests. RESULTS: Trotting in hand, 17 horses showed forelimb lameness (1-4/10) and 13 hindlimb lameness (1-2/10). Intra-individually, 11 horses showed significant differences in forelimb lameness and 4 horses showed significant differences in hindlimb lameness when ridden. Over all horses, hindlimb lameness increased significantly under the dressage rider compared to unridden horses. CONCLUSIONS: The presence of a rider can alter the degree of lameness; however, its influence cannot be predicted for an individual horse. POTENTIAL RELEVANCE: In order to evaluate mild lameness, horses should be evaluated at trot both under saddle and in hand. If lameness is exacerbated, a second rider may be helpful; the level of training of the rider should be taken into consideration. PMID- 15656507 TI - Relationships between fore- and hindlimb ground reaction force and hoof deceleration patterns in trotting horses. AB - REASONS FOR PERFORMING STUDY: The transmission of shockwaves following hoof impact is proposed to be one major source of stress to the limb. In the forelimb, there are indications that the period of horizontal deceleration of the hoof is related to the attenuation of shockwaves. In the hindlimb, information about the hoof deceleration has been lacking. OBJECTIVE: To compare hoof deceleration patterns between the fore- and hindlimbs. METHODS: Seven Standardbreds were trotted by hand over a force plate covered with sand, with triaxial accelerometers mounted on the fore and hind hooves. Variables representative of decelerations (first 2 main vertical deceleration peaks; characteristic minimum and maximum values in the craniocaudal deceleration; hoof braking time) and ground reaction forces (vertical loading rates; maximum and the following local minimum of the craniocaudal force) of the initial part of the stance phase, and the differences between individual fore- and hindlimb time and amplitude variables were used for statistical analyses. RESULTS: Force plate data showed significantly greater vertical loading rate (mean +/- s.d. 6.5 +/- 5.9 N/sec) and horizontal loads (190.4 +/- 110.2 N) in the forelimb than the hindlimb, but the parameters from accelerometer data showed no significant differences. CONCLUSIONS: No significant difference was found in the hoof deceleration, but the deceleration curves displayed a common pattern that described in detail the kinematics of the fore and hind hooves during the initial period of hoof braking. POTENTIAL RELEVANCE: These results contribute to further knowledge about the characteristics of these potential risk factors in the development of subchondral bone damage in the horse. Further studies are required on the influence of hoof braking pattern at higher speed, different shoeing and ground surfaces with different properties. PMID- 15656508 TI - Ground reaction forces and limb function in tolting Icelandic horses. AB - REASONS FOR PERFORMING STUDY: Gaited horses employ 4-beat stepping (singlefoot) gaits that extend into speeds typical of trots. Ground reaction force (GRF) patterns of these specialised gaits have not been reported; therefore, appraisal of these gaits using nongaited horse kinetics may lead to clinical misjudgements. HYPOTHESIS: GRFs of tolting Icelandic horses will be comparable in profile and magnitude with those of trotting horses. METHODS: Forelimb and hindlimb GRFs were obtained for 10 Icelandic horses ridden at a tolt. These data were evaluated across 3 speed ranges: <2, 2.5-5 and >5 m/sec. RESULTS: Virtually all vertical force tracings were single-peaked. Forelimbs typically had greater peak vertical forces and impulses compared with hindlimbs. Support duration and forelimb vertical impulse were correlated negatively with speed, whereas peak vertical, braking and propulsive forces and hindlimb braking and propulsive impulses were correlated positively with speed. CONCLUSIONS: GRF profiles of tolting Icelandic horses are more similar to profiles of trots than walks, suggesting that tolts follow bouncing mechanics. POTENTIAL RELEVANCE: Greater overlap of limb support in 4-beat gaits (even at high speeds) is associated with lower peak vertical force magnitudes of tolts compared with those reported for trots at comparable speeds, which may help limit the occurrence of overloading injuries in Icelandic horses. PMID- 15656509 TI - Influence of individual competition level on back kinematics of horses jumping a vertical fence. AB - REASONS FOR PERFORMING STUDY: The costs and investments required for the purchase and training of showjumpers justify the need to find selection means for jumping horses. Use of objective kinematic criteria correlated to jumping ability could be helpful for this assessment. OBJECTIVES: To compare back kinematics between 2 groups of horses of different competition levels (Group 1, competing at high level; Group 2 competing at low level) while free jumping over a 1 m vertical fence. METHODS: Three-dimensional recordings were performed using 2 panning cameras. Kinematic parameters of the withers and tuber sacrale (vertical displacement, vertical and horizontal velocities), backline inclination and flexion-extension motion of the 3 main dorsal segments (thoracic, thoracolumbar and lumbosacral) were analysed. RESULTS: Group 2 horses had a lower displacement of their withers and tuber sacrale from the end of the last approach stride until the first departure stride (P<0.05). As a result, they increased the flexion of their thoracolumbar and lumbosacral junctions during the hindlimb swing phase before take-off (P<0.05). However, withers and tuber sacrale velocities were slightly modified. Group 1 horses pitched their backline less forward during the forelimb stance phase before take-off and straightened it more after landing (P<0.05), probably indicating a more efficient strutting action of their forelimbs. CONCLUSIONS AND POTENTIAL RELEVANCE: Because significant differences in back motion were found between good and poor jumpers when jumping a 1 m high fence, criteria based on certain back kinematics can be developed that may help in the selection of talented showjumpers. PMID- 15656510 TI - Evaluation of pressure distribution under an English saddle at walk, trot and canter. AB - REASONS FOR PERFORMING STUDY: Basic information about the influence of a rider on the equine back is currently lacking. HYPOTHESIS: That pressure distribution under a saddle is different between the walk, trot and canter. METHODS: Twelve horses without clinical signs of back pain were ridden. At least 6 motion cycles at walk, trot and canter were measured kinematically. Using a saddle pad, the pressure distribution was recorded. The maximum overall force (MOF) and centre of pressure (COP) were calculated. The range of back movement was determined from a marker placed on the withers. RESULTS: MOF and COP showed a consistent time pattern in each gait. MOF was 12.1 +/- 1.2 and 243 +/- 4.6 N/kg at walk and trot, respectively, in the ridden horse. In the unridden horse MOF was 172.7 +/- 11.8 N (walk) and 302.4 +/- 33.9 N (trot). At ridden canter, MOF was 27.2 +/- 4.4 N/kg. The range of motion of the back of the ridden horse was significantly lower compared to the unridden, saddled horse. CONCLUSIONS AND POTENTIAL RELEVANCE: Analyses may help quantitative and objective evaluation of the interaction between rider and horse as mediated through the saddle. The information presented is therefore of importance to riders, saddlers and equine clinicians. With the technique used in this study, style, skill and training level of different riders can be quantified, which would give the opportunity to detect potentially harmful influences and create opportunities for improvement. PMID- 15656511 TI - Effects of girth, saddle and weight on movements of the horse. AB - REASONS FOR PERFORMING STUDY: Although the saddle is seen as one of the biggest causes of back pain, and weightbearing is seen as an important aetiological factor in 'kissing spine' syndrome (KSS), the effects of a saddle and weight on the back movements of the horse have never been studied. OBJECTIVE: To determine the effects of pressure on the back, exerted by tack and weight, on movements of the horse. HYPOTHESIS: Weight has an extending effect on the horse's back and, as a compensatory mechanism to this extension, an alteration in pro- and retraction angles was expected. A similar but smaller effect was expected from a saddle only and a lungeing girth. METHODS: Data were captured during treadmill locomotion at walk, trot and canter under 4 conditions: unloaded; with lungeing girth; saddle only; and saddle with 75 kg of weight. Data were expressed as maximal extension, maximal flexion angles, range of motion of L3 and L5 and maximal pro- and retraction angles of the limbs. RESULTS: At walk and trot, there was a significant influence on back kinematics in the 'saddle with weight' situation, but not in the other conditions. Overall extension of the back increased, but the range of movement remained the same. Limb kinematics changed in the sense that forelimb retraction increased. At canter, both the 'saddle with weight' and 'saddle only' conditions had a significant extending effect on the back, but there was no effect on limb kinematics. CONCLUSIONS AND POTENTIAL RELEVANCE: Weight and a saddle induce an overall extension of the back. This may contribute to soft tissue injuries and the KSS. The data from this study may help in understanding the reaction of the equine back to the challenges imposed by man when using the animal for riding. PMID- 15656513 TI - Influence of support boots on fetlock joint angle of the forelimb of the horse at walk and trot. AB - REASONS FOR PERFORMING STUDY: Support boots are thought to reduce tension on the superficial digital flexor tendon (SDTF) of the horse and are frequently recommended for horses convalescing after tendonitis, but evidence of their effectiveness is conflicting. OBJECTIVE: To document the effects of 4 different types of support boots on fetlock joint angle in comparison to the unprotected fetlock. METHODS: In 26 horses, the kinematics of the forelimb fetlock joint angle was measured at walk and trot on a treadmill when wearing 3 different types of support boots and 1 protective boot, as well as without boots. As outcome parameters, maximum extension of the fetlock joint and the moment at which maximum extension occurred in the stride cycle were determined. RESULTS: At walk, 2 of the support boots reduced the maximum extension significantly by 0.8 and 0.9 degrees, respectively (P<0.05). Additionally, one type of boots also delayed the occurrence of maximal extension within the stride cycle. At trot, all support boots reduced maximum extension significantly by 0.56-1.44 degrees (P<0.01), and the protective boot reduced maximum extension by 0.56 degrees (P<0.05). CONCLUSIONS AND POTENTIAL RELEVANCE: The results demonstrate the effectiveness of support boots in reducing maximum extension of the fetlock, which can be assumed to reduce tension in the suspensory apparatus and SDFT. The delay of the moment of maximal extension may be relevant in reducing dynamic forces. However, it should be noted that the long-term consequences of reduction of maximum fetlock extension are still uncertain. Such a reduction over a prolonged period might negatively affect fibre alignment in the healing tendon. PMID- 15656512 TI - Effects of athletic taping of the fetlock on distal limb mechanics. AB - REASONS FOR PERFORMING STUDY: Athletic taping is used frequently by human athletes to stabilise, maintain or strengthen soft tissue structures, but empirical evidence supporting any changes in equine kinematics is lacking. OBJECTIVES: To assess the effects of athletic taping of the fetlock applied by an experienced athletic trainer on forelimb mechanics in healthy horses. HYPOTHESES: That athletic taping of the distal forelimb reduces 1) hyperextension of the fetlock joint during stance, 2) flexion of the fetlock joint during swing and 3) ground reaction forces during stance. METHODS: Ground reaction force and kinematic data were obtained for 6 healthy horses trotting at 3 m/sec for 4 sequential conditions (baseline, untaped; pre-exercise, taped; post exercise, taped post 30 mins trotting exercise; transfer, 4 h after tape removal). Data were analysed using 2-way mixed ANOVAs (condition; joint). RESULTS: A statistically significant interaction was identified for the fetlock during the swing phase (mean +/- s.d. peak flexion at baseline 157 +/- 4 degrees, reduced with taping to 172 +/- 4 degrees; P<0.05) compared with no differences across conditions for the other joints. Peak vertical force reduced significantly (P<0.05) with taping. CONCLUSIONS: Athletic taping of the fetlock does not alter the kinematics of the forelimb during stance, but does limit flexion of the fetlock during the swing phase. The decreased peak vertical force may be due to an increased proprioceptive effect. POTENTIAL RELEVANCE: Reduced peak vertical forces may be of benefit in preventing or reducing injury. Further investigation remains necessary before it can be concluded that taping should be applied for tendinous or ligamentous rehabilitation in equine patients. PMID- 15656514 TI - Energetic and kinematic consequences of weighting the distal limb. AB - REASON FOR PERFORMING STUDY: It is well known that adding a load to a horse's back increases its energetic costs of locomotion, but the magnitude of increase obtained by loading the most distal portion of limb has not been measured. OBJECTIVES: To measure oxygen consumption in horses with mass added to the back and hooves. Because such mass distribution alters inertial parameters of the limbs, kinematic measurements were made to quantify the magnitude of change in limb movement. METHODS: Steady-state oxygen consumption was measured in 6 horses with a load of 2.4 kg. The load was either carried on the back or distributed equally between the 4 limbs. Modified bell boots kept the mass at the level of P3. Horses trotted on a treadmill at speeds ranging from 2 to 5 m/sec (in 0.5 m/sec increments). High-speed (250 Hz) digital images were recorded in a sagittal plane and the positions of retroreflective markers located on standard positions on the limbs were digitised for kinematic analysis. RESULTS: Loading of the distal limbs produced a 6.7% increase in metabolic rate, an order of magnitude higher than when the mass was added over the back. Although the stride period was 2% longer in horses with loads on the distal limbs, time of contact and duty factor were not different. Distal limb loading increased the range of motion in hind- but not forelimbs. CONCLUSIONS: The costs of swinging the limbs in the horse are considerable and the addition of weights to the distal limb can have a profound effect on not only the energetics of locomotion but also the kinematics, at least in the hindlimb. POTENTIAL RELEVANCE: The use of weighted shoes, intended to increase animation of the gait, increases the metabolic effort of performance horses a disproportionate amount. The additional mass also increases the joint range of motion and, potentially, the likelihood of injury. PMID- 15656515 TI - Dynamic pressure measurements for the detailed study of hoof balance: the effect of trimming. AB - REASONS FOR PERFORMING STUDY: Studies on hoof balance have, so far, only assessed the effects of strong and relatively unrealistic interventions due to technical limitations of measuring equipment. For the assessment of more subtle interventions, other techniques are necessary. OBJECTIVES: To test a sensitive pressure measurement system during locomotion and to set a standard for further studies by using the system to evaluate the effects of trimming. METHODS: Eighteen horses were measured before and after trimming with an interval of 4 weeks. Trimming was standardised to a straight hoof-pastern axis. The horses trotted over a pressure/force measuring system, with a temporal resolution of 240 Hz and a spatial resolution of 0.39 cm2. RESULTS: The preferred way of landing was lateral, asymmetrical in both front and hind feet. The duration of landing was shorter in forelimbs than in hindlimbs with an equal decreasing trimming effect. Horses had a fixed hoof-unrollment pattern; the centre of pressure (CoP) travelled towards a maximum lateral deviation and returned towards the dorsopalmar/plantar axis of the hoof. Trimming decreased the intra-individual left/right difference in maximum lateral displacement. CONCLUSIONS: The technique used provided easily accurate data to quantify hoof balance characteristics and to measure short-term trimming effects. POTENTIAL RELEVANCE: Determination of CoP patterns has added value in gait analysis and can improve our understanding of the effects of different interventions on hoof balance. PMID- 15656517 TI - [In vitro effect of pulse Nd: YAG laser plus NaF on ultrastructure and fluoride distribution of enamel]. AB - OBJECTIVE: To investigate the effect of pulse Nd: YAG laser combined with NaF on caries resistance of human enamel in vitro. METHODS: 24 caries-free, constant premolares were cut into enamel mass. Surface ultrastructure and fluoride distribution of enamel in laser-NaF treated group, laser treated group, NaF treated group and control group were observed by scanning electron microscope (SEM) and electron probe microanalyzer. RESULTS: (1) SEM observation: In laser NaF treated group, the smear layer was almost removed, the enamel surface showed partially melted-like and bubble-like appearance, diffuse white-small particles distributed on enamel surface, and melted position, microcrack and microhole appeared whitely blocking-reflection. (2) Electron probe microanalyzer analysis: In enamel samples, the highest average Ci of F- and the hightest depth which F- entered in enamel were recorded in the laser-NaF treated group. The average Ci of F- in laser-NaF treated group was significantly increased compared with that in NaF-treated group or laser-treated group or control group (P < 0.001). CONCLUSION: The pulsed Nd:YAG laser combined with sodium fluoride could make the enamel surface partially melt, change the structure of the crystal, enhance the absorption of fluoride, and clear smear layer. This technology may resist the caries. PMID- 15656516 TI - [Expression related to vascular endothelial growth factor C and induced nitride oxide synthesizase in lymph node micrometastasis of oral squamous cell carcinoma]. AB - OBJECTIVE: To investigate the relationship of vascular endothelial growth factor C (VEGF-C) and induced nitride oxide synthesizase (iNOS) expression in lymph node micrometastasis of oral squamous cell carcinoma. METHODS: Samples were obtained from 47 cases of oral squamous cell carcinoma and 15 cases with normal oral mucosa, VEGF-C and iNOS mRNA expression were detected by RT-PCR method. Lymph node micrometastasis of 10 normal lymph nodes and 355 lymph nodes from 47 cases of oral squamous cell carcinoma was detected with immunohistochemical reaction in cytokeratin antibody. RESULTS: The percentages in tumors with higher expression were 57.4% for VEGF-C, 68.1% for iNOS (P < 0.05). They were significantly higher than that of normal groups. Significant positive relationship was found between VEGF-C and iNOS (P < 0.01). The positive rate of cytokeratin (CK) was 48.9%. Significant positive relationship was found between VEGF-C and CK, iNOS and CK (P < 0.01). The expression rates of CK in positive group of VEGF-C and iNOS were 63.0%, 65.6% respectively, and were significant higher than negative groups. CONCLUSION: Expression of VEGF-C and iNOS in lymph node micrometastasis of oral squamous cell carcinoma is significant related. PMID- 15656518 TI - [Inhibition effect of glycyrrhizin on transplanted mandibular gland fibrosarcoma of mice]. AB - OBJECTIVE: To study glycyrrhizin's anticancer effect and its mechanism. METHODS: 3-methylcholanthrene were injected into mice's submandibular glands to induce tumor, then transplanted the tumor pieces (1 mm3) to mice. The transplanted tumors were measured, and flow cytometry analysis and cytomorphology observation were conducted. RESULTS: Glycyrrhizin (GL) inhibited the transplanted mandibular gland fibro-sarcoma of mice and the suitable GL dose for inhibiting fibrosarcoma of mice was 1.61 mg per 20 g weight. The GL dose below 3.22 mg per 20 g weight didn't produce remarkable toxicity and side effects. GL induced cytomorphological changes of tumor cells and enhanced immunosuppression of macrophage on fibrosarcoma. The result of flow cytometry showed that tumor cell counts of GL1 and GL2 groups increased remarkably in DNA synthetic prophase, and decreased in DNA synthetic phase. CONCLUSION: GL can inhibit transplanted mandibular gland fibro-sarcoma of mice. The anticancer mechanism of GL may be acting on related enzymes with phagocytosis. The result of flow cytometry showed that the shift of fibrosarcoma cells from G1 phase to S phase was blocked. This suggests that the anticancer action of GL is related to its inhibition of ribonucleotide reductase, a rate-limiting enzyme in DNA synthesis. PMID- 15656519 TI - [Cytocompatibility of nanophase hydroxyapatite ceramics]. AB - OBJECTIVE: To evaluate the cytocompatibility of nanophase hydroxyapatite ceramics in vitro. METHODS: Hydroxyapatite (HA) was prepared via wet method. The grain size of the hydroxyapatite in the study was determined by scanning electron microscope and atomic force microscope with image analysis software. Primary osteoblast culture was established from rat calvaria. Cell adherence and proliferation on nanophase hydroxyapatite ceramics and conventional hydroxyapatite ceramics were examined at 1, 3, 5, 7 days. Morphology of the cells was observed by microscope. RESULTS: The average grain size of the nanophase and conventional HA was 55 nm and 780 nm, respectively. Throughout 7 days period, osteoblast proliferation on the HA was similar to that on tissue culture borosilicate glass controls, osteoblasts could attach, spread and proliferate on HA. However, compared to conventional ceramics, osteoblast proliferation on nanophase HA was significantly better after 1, 3, 5 and 7 days. CONCLUSION: Cytocompatibility of nanophase HA was significantly better than conventional ceramics. PMID- 15656520 TI - [The positive effect of transforming growth factor beta on ectomesenchymal stem cells of embryonic facial processes differentiating to smooth muscle cells]. AB - OBJECTIVE: To investigate the effect of transforming growth factor beta (TGF beta) on ectomesenchymal stem cells differentiating to smooth muscle cells. METHODS: 60 pmol/L TGF-beta was added to the ectomesenchymal stem cells of embryonic facial processes. Immunohistochemistry assay and image analysis were used to value the expression extent of a smooth muscle actin (alpha-SMA) and quantitative RT-PCR was used to value the quantity of alpha-SMA. RESULTS: 2 days later, about 95% cells in TGF-beta group and 65% cells in control group without differentiation inhibitor expressed alpha-SMA. Expression of alpha-SMA in TGF beta group was stronger than that of control group after one and two days. Quantitative RT-PCR showed the quantity of alpha-SMA mRNA in treated group cells was more than that of in control group. CONCLUSION: Quantity of alpha-SMA in TGF beta group is more than that of spontaneous differentiation group. TGF-beta has positive effect on ectomesenchymal stem cells differentiating to smooth muscle cells. PMID- 15656521 TI - [The study of viscoelastic residual stresses of ceramic-metal bond]. AB - OBJECTIVE: To study the residual stresses of ceramic-metal bond at viscoelastic and elastic phases during cooling of porcelain-fused-to-metal in order to precisely calculate the ceramic-metal bond strength and improve the restorations. METHODS: The finite element model was set up according the crack initiation test (three-point flexure bond test) based on ISO Standard, elements of viscoelastic and themal-displacement were used to part the model. The result at viscoelastic phases was used as initiation condition of elastic phases to add up. RESULTS: The compressive stress was caused by metal during cooling occurred in the ceramic. The shear stress induced by loading was offset by thermal shear stress. Load tensile stress and the thermal compressive stress vertical of interface concentrated at the end of the bond interface, but the tensile was greatly higher. CONCLUSION: The residual stress is very important to metal-ceramic restorations, and the viscoelastic behavior of porcelain greatly influences it. If the metal and ceramic are compatible,the components stresses of the residual stresses may benefit to ceramic-metal bond, and can be taken as a part of bond stresses. PMID- 15656522 TI - [The study of the relationship of malodor and microbial composition of interdental and subgingival plaques in periodontitis patients]. AB - OBJECTIVE: The study is to investigate the microbial composition of interdental and subgingival plaques of periodontitis patients with or without malodor, to explore the relationships between periodontitis and oral malodor. METHODS: 20 patients of periodontitis with malodor were chosen from 210 patients of periodontitis, and the clinical parameter of plaque index (PLI), gingival bleeding index (GBI) and probing depth (PD) were measured and compared with the control group which had periodontal disease without malodor. During the experiment, the interdental and subgingival microbial samples in both groups were collected and sent to anaerobic culture for 48 hrs, then the total CFU/ml of each sample were counted, and each type of bacteria was separated and identified. All of the data were analyzed by using the statistical software SPSS 10.0. RESULTS: (1) There were no satistical differences on PLI, GBI, PD between experimental group and control group. (2) The percents of leptospira in both interdental and subgingival plaques of test group were significantly higher than that of the control group (P < 0.01). (3) Either the interdental or in subgingival plaques, the count results of CFU/ml were similar in both groups (P > 0.05). (4) The proportions of malodor producing anaerobic bacteria in interdental gingival plaque, such as P. gingivalis and Veillonelia, were singnificantly different between test group and control group. CONCLUSION: The proportions of VSC's producing anaerobic bacteria in interdental gingival plaque may be play the significant roles in oral malodor. Further studies should be taken to elucidate the relationship between malodor and periodontitis. PMID- 15656523 TI - [A study of the related factors and prophylactic measures of endodontic interappointment emergencies]. AB - OBJECTIVE: To estimate the related factors and prophylactic measures of endodontic interappointment emergencies (EIE). METHODS: The clinical cases were divided into three groups. Cases in the routine group were treated according to the routine procedures of root canal therapy. Cases in the medicament group took dexamethasone tablets after the instrumentation. Root canals in the drainage group were prepared at the first visit and had been left open for two or three days. The related factors were recorded. The incidences and flare-up index (FUI) value of EIE were statistically analyzed. RESULTS: The EIE incidence of the routine group was 19.1%. The EIE incidence and FUI value of the overinstrumented teeth were significantly higher than those of other teeth (P < 0.05). EIE were not significantly related to the age, gender, tooth location and pathosis of the pulpal and periapical tissue (P > 0.05). The FUI value of the medicament group was significantly lower than that of the routine group (P < 0.05). Less severe EIE occurred in the cases of the drainage group than those of the control group (P > 0.05). CONCLUSION: The occurrence of EIE was significantly related to overinstrumentation. Taking dexamethasone may relieve EIE. PMID- 15656524 TI - [Comparative study of resosolv and chloroform in retreatment of canal obturated with resinifying agent]. AB - OBJECTIVE: To evaluate the possibility for removal of resinifying agent, time required for removal and the working length loss by Resosolv or Chloroform. METHODS: 40 human teeth (80 root canals) obturated with FR phenolaldehyde agent were divided into four groups, 20 root canals per group. Group A: Resosolv with K file; group B: chloroform with K file; group C: Resosolv with Ultrasonic K file; group D: Chloroform with ultrasonic K file. Calculating the pereentage for removal of resinifying agent, time required for removal and the working length loss. RESULTS: The effectiveness of Resosolv for removing resinifying agent was better than chloroform. 87.5% of canals could be negotiated by Resosolv; 45% of canals be negotiated by chloroform. CONCLUSION: Resosolv is an effective solvent for canals obturated with resinifying agent. PMID- 15656525 TI - [Analysis of 30 consecutive free flap transplantation for head and neck reconstruction in paediatric patients]. AB - OBJECTIVE: To analyze the reliability and feasibility of free flap transfer for head and neck reconstruction in the paediatric patients. METHODS: Thirty consecutive free flap transfer performed in 28 patients younger than 15 years-old were reviewed. Data concerning the operation included date of surgery, defect description and site, stage and histology of tumor, flap type, recipient vessel and complications. RESULTS: The average age of patients was 12.3 years-old, with age ranged from 5 to 15. There were 19 males and 9 females. Four kinds of free flap were used. The free fibula flap was most commonly used, followed by free radial forearm flap, rectus abdominis myocutaneous flap, and free scapular flap. The overall success rate of the flaps was 93.3%. The overall complication rate was 36.7%. The vessel thrombosis rate was 6.7%. CONCLUSION: Free flap transfer in the paediatric patients for head and neck reconstruction is safe and reliable. There is no difference between the normal patients and paediatric patients in the success rate and complication rate. PMID- 15656526 TI - [Observation and analysis on coronal MR images of cadaver TMJ]. AB - OBJECTIVE: To investigate the MR image features of TMJ coronal structures. METHODS: 28 TMJs from 15 cadavers dipped in formalin over 1 year were included. Their mandibles were fixed at intercuspal position when TMJs oblique coronal MR scan was taken. Baseline was draw on the oblique coronal imaging along the ramus of mandible through its center. The corresponding dimensions and areas were measured with electronic ruler. RESULTS: The dimensions were in accordance from large to small as: lateral-medial dimension of fossa, lateral-medial dimension of condyle, medial dimension of fossa, medial dimension of condyle, lateral dimension of condyle and lateral dimension of fossa. Three significant co relative relationship were found: the lateral dimension of condyle with lateral dimension of fossa, the medial dimension of condyle with medial dimension of fossa, and the lateral-medial dimension of condyle with lateral-medial dimension of fossa. CV of lateral dimension of fossa was higher than that of lateral and medial dimension of condyle, and then all the three were higher than other dimensions. The size of the area of joint space shown on oblique coronal imagings was positively related to that of the area of coronal disc imagings. CONCLUSION: Significant co-ordinate relationship of condyle and fossa of TMJ coronally was existed, and the condyle position to fossa lateral-medially varied in great. PMID- 15656527 TI - [The orthognathic treatment of mandibular asymmetry]. AB - OBJECTIVE: To evaluate the methods of orthognathic treatment for mandibular asymmetry. METHODS: 110 cases of mandibular asymmetry had been treated in past thirteen years. The type of deformity, the management of the cases, and the effects of orthognathic treatment were summarized in this paper. Orthodontic therapy was taken before and after operation to keep the treatment effect. RESULTS: According to the type of the asymmetry deformity, different methods were used. After treatment, all the patients obtained satisfactory facial appearances and oral function. CONCLUSION: It should be diagnosed before surgery whether the development of deformity is steady or not. If it is steady, the effects of orthognathic and orthodontic therapy will be satisfactory; if not, we should remove the affected condyles and simulataneously ameliorate facial appearance. PMID- 15656528 TI - [Clinical analysis of badly necrotic pyogenic infection in cervical part with pneumatosis of vomica]. AB - OBJECTIVE: To analyze the clinical character, diagnosis and treatment of badly necrotic pyogenic infection in cervical part with pneumatosis of vomica. METHODS: The clinical character, diagnosis and treatment of badly necrotic pyogenic infection in cervical part with pneumatosis of vomica of 46 cases were investigated retrospectively. RESULTS: Lateral and bilateral neck infection cases were 38 and 8 respectively. 30 cases formed primary pyogenic infection in cervical part with pneumatosis of vomica, and 16 did from adjacent sites. Besides the characters of the acute infection, gas storage in deep cervical part abscess was notable. CT and B Ultrasonic examination provided useful informations such as sizes, shapes, capacity, extents of abscess and the relationship between the abscess and vessel or vital organ. Diagnosis puncture and germiculture were performed before and after operation. The results showed that 25 of 46 cases were infected by staphylococcus or streptococcus, and 21 cases did by other bacterium. Exploration and drainage treatments were performed. All cases were cured except 2 died. CONCLUSION: Diagnostic puncture, CT and/or B Ultrasonic examination are essential for diagnosis and presurgical planning. Germiculture provides reliable evidence for finding pathogeny and therapy. The most possibilities of pyogenic infection with pneumatosis of vomica in cervical part are the action of aerogenic bacterium, infection both in cervical part and chest or swallowing movement of pharynx. PMID- 15656529 TI - [The study on color space of the VINTAGE & UNIBOND standard color]. AB - OBJECTIVE: To research the color space of the standard color board, guide the clinic work of color match, and establish an foundation for deeper researching. METHODS: Through taking analysis of reflecting spectrum and spectral tristimulus, each parameter of the color on standard board was found out. The color difference between border upon colors on board was worked out through matching the color parameters. The number of color space between each border upon color on standard color board, divided by deltaE = 1.5, was found. RESULTS: The number of color space between each border upon color was 3-6. The color space was bigger in group B, but smaller in group D. The color space was bigger in high lightness area, but smaller in low lightness area. CONCLUSION: Some color space has been found between standard color borders of VINTAGE and UNIBOND. Color of natural teeth is unable to be completely included into standard color border, especially in clinical care. PMID- 15656530 TI - [A statistic analysis on absence of third molar germs in orthodontic patients]. AB - OBJECTIVE: To examine the absence of the third molar germs in orthodontic patients and to evaluate the relationship between third molar germs and malocclusion. METHODS: The subjects comprised 234 patients (male 92, female 142) from the orthodontic clinic whose ages were 14-18. The assessments of the third molar germs were made from panoramic radiographs, and the assessments of ANB angle were made from lateral cephalograms. All the data were analyzed by statistic chi2 test. RESULTS: The percentage of male who missed one or more third molar gems (37.0%) was higher than that of female (24.6%). There was no significant difference between the absent frequencies of third molar germs on left and right sides in either maxilla or mandible. The absent percentage of third molar germs in skeletal III subjects was higher than those in both skeletal class I and II subjects. The absent difference of third molar germs was in upper arches (P < 0.05), but not in lower arches (P > 0.05). There was no significant difference in absent percentage of third molar germs between skeletal class I and II subjects. CONCLUSION: Male patients have higher absent frequencies of third molar germs than female ones. Skeletal class III patients have higher absence of third molar germs in upper jaws than skeletal class I and II patients. PMID- 15656531 TI - [The study on characteristics of soft tissue profile for different malocclusion]. AB - OBJECTIVE: To study the coordination of facial soft tissue in Angle's Class I, II1, III malocclusion, providing reference for the clinical practice. METHODS: 60 lateral cephalometric radiographs of three classes of Angle's malocclusion were included. 13 measurements were analyzed by SPSS 10.0. RESULTS: Protrusion of the upper and lower lips increased in Angle's II1 comparing with Angle's I and protrusion of the upper lip was larger than the lower lip, no difference was found in the facial convexity. Increase of upper lip inclination and underdevelopment of maxillary in Angle's III comparing with Angle's I. No significant different was found in the form of lower lip, soft tissue facial angle and Z angle. Comparing Angle's II1 with Angle's III, every measurements were significantly different except upper lip inclination, nasolabial angle, representing the formation mechanism of malocclusion. CONCLUSION: Some differences were found in form and position of lip in three classes of Angle's malocclusion. However, the deformity was not obvious in Angle's II1 and Angle's III because of compensation mechanism. Nasolabial angle only reflected change of upper lip, but could not reflect characters of facial profile. PMID- 15656532 TI - [Expression of telomerase activity and c-myc and stimulatory protein 1 in human ameloblastoma]. AB - OBJECTIVE: To study the oncogene transcriptor c-myc, stimulatory protein 1 (SP1) expression in ameloblastoma (AB) and their relation with telomerase reverse transcripase (hTERT), and to investigate the clinical biological characteristics of AB. METHODS: The expression was observed in AB by in situ hybridization and SP method. RESULTS: The positive rates of c-myc mRNA, hTERT mRNA and SP1 protein were 81.5% (44/54), 94.4% (51/54) and 83.3% (45/54), respectively. Their positive rates increased as AB recurred and transformed malignantly. A strong correlation was found between hTERT and c-myc, hTERT and SP1 (rs = 0.853, P < 0.001; rs = 0.900, P < 0.001). CONCLUSION: Activity of telomerase plays an important role in the tumorigenesis development of AB. Increasing of hTERT expression may be related to c-myc and SP1. The expression of these three parameters has a significant correlation with the clinical biological characteristics of AB. PMID- 15656533 TI - [Construction of eukaryotic expression vector for KGPcd gene from Porphyromonas gingivalis and expression in mammalian cells]. AB - OBJECTIVE: This study aimed at constructing secretory eukaryotic expression vector of KGPcd gene encoding whole amino acid residues of mature KGPcd from Porphyromonas gingivalis and investigating the transcription and expression of recombined plasmid VR1020/KGPcd in mammalian cells. METHODS: Eukaryotic expression plasmid VR1020/KCPcd was constructed by using molecular cloning methods. Then, the VR1020/KGPcd was transfected into mammalian cell COS7 with Lipofectamine 2000 according to the manufacturer's instruction. The transcription of VR1020/KGPcd was assayed by reverse transcription polymerase chain reaction (RT-PCR). The expression product of VR1020/KGPcd was analyzed by using indirect immunofluorescence. The protein secretion in cultural medium was detected by ELISA method. RESULTS: It proved that the VR1020/KGPcd could be transcribed and translated into transfected COS7 cells. The expressed targeted protein could be secreted into cultural supernatant and could be detected by ELISA. CONCLUSION: The eukaryotic expression plasmid of VR1020/KGPcd was constructed successfully and its product can be expressed in mammalian cells. The results indicated that the recombinant plasmid has antigenicity and may be acted as candidate gene vaccine. This laid a basis for its use as gene vaccine candidates in the development of anti-periodontitis and paved the way for further study. PMID- 15656534 TI - [A study on myogenic differentiation of human adipose tissue-derived stromal cells]. AB - OBJECTIVE: To isolate human adipose tissue-derived stromal cells and study the potential of osteogenic differentiation after inductive culture. METHODS: Liposuction human adipose tissues were minced and digested with collagenase type I. The obtained stromal cells were plated in BGJb medium as primary culture for ten days. The second passage cells were harvested and plated in DMEM/F12 medium supplemented with 10% FBS, 5% horse serum and 50 micromol/L hydrocortisone for myogenic induction culture. The cell-anchored slips were removed and fixed in 4% formaldehydam polymerisatum. Toluidine blue, Mallory's phosphotungstic hematoxylin staining and monoclonal antibody to human skeletal muscle myosin heavy chain immunocytochemical methods were used to assay the differentiation of cells. RESULTS: It was observed that the size and shape of induced cells were much different from those of non-induced cells. Toluidine blue, Mallory's phosphotungstic hematoxylin staining demonstrated there were many basophilic striations within cytoplasm and multinucleated myotubes were formed. Immunocytochemical stain indicated that characterastic skeletal myosin heavy chain was positive in myogenic induced cells. CONCLUSION: It seems that human adipose tissue represents an abundant reservoir of adult stem cells that have multi-germline potential to differentiate into myoblasts. Adipose tissue derived stromal cells will be another alternative source for cell-based tissue engineering in skeletal muscle reconstruction. PMID- 15656535 TI - [Changes of taste bud and fungiform papillae after 60Co radiation in rat]. AB - OBJECTIVE: To observe the morphological changes and the regenerating ability of the fungiform papillae and taste buds after 60Co radiation with clinical doses in rats. METHODS: The heads, faces and necks of 30 SD rats were radiated with a large dose and one time of 60Co in the clinical radiation. The general living condition and the number and shape of the fungiform papillae and taste buds of the tongues were observed after the radiation in rats. RESULTS: In the group of 60Co radiation, the animals had wilting, decreasing appetite, losing weight. The heads, faces and necks of animals appeared redness, peeling of hair, increasing of secretions in 5 days after the 60Co radiation. The changes reached the summit in 10 days and the general living condition of the animals recovered in 60 days. The fungiform papillae and taste buds of the animals appeared degeneration, atrophy and collapsing in 5 days after the 60Co radiation. The injuries reached the summit in 10-20 days and the fungiform papillae and taste buds regenerated partially, and the some atrophied fungiform papillae and taste buds were not regenerated in 60 days. CONCLUSION: The damage to fungiform papillae and taste buds of tongue following the 60Co radiation with the clinical doses was very serious. The damaged fungiform papillae and taste buds can regenerate partially, but not completely. PMID- 15656536 TI - [Cloning and sequencing of the upstream of mouse dentin sialophosphoprotein promoter]. AB - OBJECTIVE: To clone and sequence the upstream of mouse dentin sialophosphoprotein promoter. METHODS: Genomic DNA was obtained from Balb/c mouse blood. The upstream of mouse dentin sialophosphoprotein promoter segments was obtained by PCR. Then the segments were inserted into T-vector. The plasmids were identified by digestion with the restriction enzyme analysis. The positive clone was sequenced and compared with Genebank. RESULTS: The upstream of mouse dentin sialophosphoprotein promoter was divided into three sequences and three different target segments with 997 bp, 1004 bp and 674 bp in length were obtained. After identified, sequenced and compared with Genebank, the sequences of the segments were consistent with those displayed on Genebank by 99%. CONCLUSION: The clone of the upstream of mouse dentin sialophosphoprotein promoter was successful. This work will help to study the regulation of DSPP expression. PMID- 15656537 TI - [Effect of mold temperature on mechanical properties of Ti-Zr alloy castings]. AB - OBJECTIVE: To investigate the influence of mold temperature on the mechanical properties of Ti-Zr alloy castings. METHODS: Ti-Zr alloy was cast into a mold which was made of a zircon (ZrO2.SiO2) investment with a casting machine (China) designed as vacuum, pressure and centrifuge. At three mold temperatures (room temperature, 300 degrees C, 600 degrees C) the Ti-Zr alloy was cast separately. The ultimate tensile strength (sigmab), 0.2% offset yield point strength (sigma0.2), and elongation to fracture (delta) were determined by a universal testing machine with 15 mm pre-marked gauge interval. The surface micro structures of Ti-Zr alloy castings at three mold temperatures were observed by optical microscope. RESULTS: At a higher mold temperature during casting, the tensile strength and yield strength of Ti-Zr alloy were significantly increased, while the elongations were significantly decreased. The micro-structure of the surface was different from the inner part. There was a reacted layer in the outermost surface. The thickness of reacted layers at the three mold temperatures was about 35.5 man (room temperature), 55.0 microm (300 degrees C) and 65.5 microm (600 degrees C), respectively. The inner part of microstructure composed of main a phases within the prior alpha grain boundaries. CONCLUSION: The mold temperature is one of the major factors influencing Ti-Zr alloy mechanical properties. Comparing with Ti6Al4V alloy and Co-Cr alloy castings, Ti-Zr alloy is more suitable for denture frame. PMID- 15656539 TI - Finding the gold in MEDLINE: clinical queries. PMID- 15656538 TI - [A survey on knowledge and habits of oral health in freshmen of Sichuan University]. AB - OBJECTIVE: To investigate the oral health knowledge and habits of freshmen of Sichuan University and promote the prophylactic therapy of dental diseases in the university. METHODS: The table and the standard on oral health survey authorized by WHO were employed in this study. RESULTS: Only 13.1% of all the investigated freshmen had periodic oral health examination. 74.4% brush teeth twice per day, only 7.7% brush teeth over 3 minutes, 16.6% insist in fluoridated toothpaste, and only 6% had applied dental floss. 51.9% do not often take snack food and 94.7% are non-smokers; (4) Tests of prophylactic therapy of dental caries and periodontal diseases and oral health knowledge have poor results. CONCLUSION: The freshmen have limited knowledge of oral health and lack serious attitude toward oral cleaning behaviors. PMID- 15656540 TI - Peginterferon alpha-2a improved the hepatitis C virologic response in concurrent HIV and chronic hepatitis C virus infections. PMID- 15656541 TI - Rapid measurement of B-type natriuretic peptides reduced time to discharge and treatment costs in patients with acute dyspnea. PMID- 15656542 TI - Review: hormone replacement therapy may reduce the risk for death in younger but not older postmenopausal women. PMID- 15656543 TI - Volunteers trained in CPR and the use of automated external defibrillators increased survival after out-of-hospital cardiac arrest. PMID- 15656544 TI - An early invasive strategy reduced death or MI better than a conservative strategy in unstable angina and non-STEMI at advanced age. PMID- 15656545 TI - Review: sedative-hypnotic agents reduce mortality and duration of delirium in the alcohol withdrawal syndrome. PMID- 15656546 TI - Methylprednisolone caused an increase in death after head injury. PMID- 15656547 TI - Esomeprazole was not better than omeprazole for resolving heartburn in endoscopy negative reflux disease. PMID- 15656548 TI - Below-knee elastic compression stockings reduced development of the postthrombotic syndrome in proximal deep venous thrombosis. PMID- 15656550 TI - Long-term use of combination DMARDs did not sustain disease remissions, but delayed joint damage in early rheumatoid arthritis. PMID- 15656549 TI - Review: Foam-based, constant low-pressure mattresses are better than standard hospital mattresses for reducing pressure ulcers. PMID- 15656551 TI - Peginterferon alpha-2a improved the hepatitis C virologic response in concurrent HIV and chronic hepatitis C virus infections. PMID- 15656553 TI - Review: clonidine is more effective than placebo for long-term smoking cessation, but has side effects. PMID- 15656554 TI - Review: replacement therapy is effective in both men and women. PMID- 15656555 TI - Review: pharmacotherapy improves cognitive symptoms in dementia. PMID- 15656556 TI - Review: limited evidence supports the use of atypical antipsychotic drugs in behavioral and psychological symptoms of dementia. PMID- 15656557 TI - Ezetimibe plus simvastatin lowered lipid levels more than simvastatin monotherapy in primary hypercholesterolemia. PMID- 15656558 TI - A dietitian-led intervention reduced weight and waist circumference in obese patients with type 2 diabetes. PMID- 15656559 TI - Review: fluoxetine, orlistat, and sibutramine modestly reduce weight in type 2 diabetes. PMID- 15656560 TI - Review: orlistat and sibutramine are modestly effective for weight loss at 1 year. PMID- 15656561 TI - Review: antioxidant supplementation does not reduce gastrointestinal cancer. PMID- 15656562 TI - Review: anticonvulsants are better than placebo for reducing the frequency of migraine attacks. PMID- 15656563 TI - Nebulized lidocaine before nasogastric tube insertion reduced patient discomfort but increased risk for nasal bleeding. PMID- 15656564 TI - Magnetic resonance imaging was more sensitive than mammography for detecting breast cancer in high-risk women. PMID- 15656566 TI - Loop diuretics and angiotensin-converting enzyme inhibitors increased risk for hospitalization for lithium toxicity. PMID- 15656565 TI - Mortality rate from early prostate cancer increased 3 fold after 15 years following the diagnosis. PMID- 15656568 TI - Next generation, high relaxivity gadolinium MRI agents. PMID- 15656569 TI - Ligand-carrying gas-filled microbubbles: ultrasound contrast agents for targeted molecular imaging. PMID- 15656570 TI - Improving the integrity of three-dimensional vascular patterns by poly(ethylene glycol) conjugation. AB - Development of functional tissue-engineering constructs may require that multiple cell types be organized in controlled three-dimensional (3-D) microarchitectures with proper nutrient diffusion and vascularization. In the past few years, a variety of microscale techniques have demonstrated the ability to control protein and cell attachment in defined patterns. Nevertheless, maintenance of these patterns over time has been a significant challenge due to nonspecific protein adsorption and cell migration. To this end, we have investigated the effectiveness of poly(ethylene glycol) (PEG) thin films in maintaining the integrity of 3-D cellular patterns, using human umbilical vein endothelial cells (HUVEC) as a model system. These HUVEC constructs were created using extracellular matrix (ECM)-based microfluidic patterning. Our results indicated that PEG-conjugated substrates improve cell pattern integrity as compared to control silicon. The compliance multifactor (a measure of pattern integrity; higher value means lower pattern integrity) was about 3.66 +/- 0.29 on day 5 for PEG-conjugated surfaces, compared with 8.23 +/- 0.42 for control surfaces ECM based microfluidic patterning coupled with stable PEG-conjugated surfaces may serve as a vital tool for vascularized tissue engineering. PMID- 15656572 TI - Positron-labeled dopamine agonists for probing the high affinity states of dopamine subtype 2 receptors. AB - It is well documented that guanidine nucleotide-coupled dopamine subtype 2 receptors (D2) are configured in high and low affinity states for the dopamine agonist in vitro. However, it is still unclear whether these functional states exist in vivo. We hypothesized that positron-labeled D2 agonist and Positron Emission Tomography can be used to probe these functional states noninvasively. Recently, we demonstrated in nonhuman primates that N-[11C]propyl-norapomorphine (NPA), a full D2 agonist, is a suitable tracer for imaging the high affinity states of D2 receptors in vivo. We also developed kinetic modeling method to derive receptor parameters, such as binding potential (BP) and specific uptake ratios (V3''). When coupled with a dopamine releasing drug, amphetamine, NPA was found to be more sensitive than antagonist tracers, such as [11C]raclopride (RAC), to endogenous dopamine concentration changes (by about 42%). This finding suggests that NPA is a superior tracer for reporting endogenous DA concentration. In addition, the difference of the BP or V3'' of NPA and RAC under control and amphetamine challenge conditions could be used to estimate the functional states of D2 receptors in vivo. On the basis of our findings and the assumptions that NPA binds only to the high affinity states and RAC binds equally to both affinity states, we proposed that about 70% of the D2 receptors are configured in the high affinity states in vivo. PMID- 15656571 TI - Human serum albumin bearing covalently attached iron(II) porphyrins as O2 coordination sites. AB - Tetrakis{(alpha,alpha,alpha,alpha-o-pivalamido)phenyl}porphinatoiron(II) with a bifunctional tail possessing an axially coordinated imidazolyl group and a protein attachable succinimidyl(glutamyl) group (FeP-GluSu) has been synthesized. It can efficiently react with the lysine residues of recombinant human serum albumin (rHSA), giving a new albumin-heme conjugate [rHSA(FeP-Glu)]. MALDI-TOFMS showed a distinct molecular ion peak at m/z 70 643, which indicates that three FeP-Glu molecules were covalently linked to the rHSA scaffold. The binding number of FeP-Glu is approximately three (mol/mol) and independent of the mixing ratio. The CD spectrum and Native PAGE revealed that the albumin structure remained unaltered after the covalent bonding of the hemes. This rHSA(FeP-Glu) conjugate can bind and release O2 reversibly under physiological conditions (pH 7.3, 37 degrees C) in the same manner as hemoglobin and myoglobin. The O2-adduct complex had a remarkably long lifetime (tau(1/2): 5 h). The O2-binding affinity [P(1/2)O2: 27 Torr] was identical to that of human red cells. Laser flash photolysis experiments gave the O2- and CO-association rate constants and suggested that there are two different geometries of the imidazole binding to the central ion. PMID- 15656573 TI - Chlorophyll-a analogues conjugated with aminobenzyl-DTPA as potential bifunctional agents for magnetic resonance imaging and photodynamic therapy. AB - A clinically relevant photosensitizer, 3-devinyl-3-(1 hexyloxyethyl)pyropheophorbide-a (HPPH, a chlorophyll-a derivative), was conjugated with Gd(III)-aminobenzyl-diethylenetriaminepentaacetic acid (DTPA), an experimental magnetic resonance (MR) imaging agent. In vivo reflectance spectroscopy confirmed tumor uptake of HPPH-aminobenzyl-Gd(III)-DTPA conjugate was higher than free HPPH administered intraveneously (iv) to C3H mice with subcutaneously (sc) implanted radiation-induced fibrosarcoma (RIF) tumor cells. In other experiments, Sprague-Dawley (SD) rats with sc implanted Ward Colon Carcinoma cells yielded markedly increased MR signal intensities from tumor regions-of-interest (ROIs) 24 h post-iv injection of HPPH-aminobenzyl-Gd(III) DTPA conjugate as compared to unconjugated HPPH. In both in vitro (RIF tumor cells) and in vivo (mice bearing RIF tumors and rats bearing Ward Colon tumors) the conjugate produced significant increases in tumor conspicuity at 1.5 T and retained therapeutic efficacy following PDT. Also synthesized were a series of novel bifunctional agents containing two Gd(III) atoms per HPPH molecule that remained tumor-avid and PDT-active and yielded improved MR tumor conspicuity compared to their corresponding mono-Gd(III) analogues. Administered iv at a MR imaging dose of 10 micromol/kg, these conjugates produced severe skin phototoxicity. However, by replacing the hexyl group of the pyropheophorbide-a with a tri(ethylene glycol) monomethyl ether (PEG-methyl ether), these conjugates produced remarkable MR tumor enhancement at 8 h post-iv injection, significant tumoricidal activity (80% of mice were tumor-free on day 90), and reduced skin phototoxicity compared to their corresponding hexyl ether analogues. The poor water-solubility characteristic of these conjugates was resolved by incorporation into a liposomal formulation. This paper presents the synthesis of tumor-avid contrast enhancing agents for MR imaging and thus represents an important milestone toward improving cancer diagnosis and tumor characterization. More importantly, this paper describes a new family of bifunctional agents that combine two modalities into a single cost-effective "see and treat" approach, namely, a single agent that can be used for contrast agent-enhanced MR imaging followed by targeted photodynamic therapy. PMID- 15656575 TI - Multivalent carbocyanine molecular probes: synthesis and applications. AB - Synergistic multivalent interactions can amplify desired chemical or biological molecular recognitions. We report a new class of multicarboxylate-containing carbocyanine dye constructs for use as optical scaffolds that not only serve as fluorescent antennas but also participate in structural assembly of the multivalent molecular construct. Three generations of carboxylate-terminating multivalent near-infrared carbocyanine probes from a dicarboxylic acid precursor dye (cypate) were prepared via its imino diacetic acid derivatives. Conjugation of the probes with D-(+)-glucosamine afforded dendritic arrays of the carbohydrates on an inner NIR chromophore core. All the multicarboxylate probes and their glucosamine conjugates have similar NIR spectral properties because conjugation occurred at distal positions to the inner chromophore core, thereby providing consistent and predictable spectral properties for their biological applications. Although light-induced photodamage equally affected the precursor dye, multicarboxylate probes, and their glucosamine derivatives, we observed that octacarboxylcypate (multivalent probe) was remarkably stable in different mediums at physiologically relevant temperatures relative to cypate, especially in basic mediums. Biodistribution studies in tumor-bearing nude mice show that all the glucosamine conjugates localized in the tumor but cypate was almost exclusively retained in the liver at 24 h postinjection. The tumor uptake does not correlate with the number of glucosamine tether on the multicarboxylate probe. Overall, the triglucosamine derivative appears to offer the best balance between high tumor uptake and low retention in nontarget tissues. These results suggest that multivalent molecular beacons are useful for assessing the beneficial effects of multivalency and for optimizing the biological and chemical properties of tissue specific molecular probes. PMID- 15656574 TI - A new high affinity technetium-99m-bombesin analogue with low abdominal accumulation. AB - 99mTc-labeled bombesin analogues have shown promise for noninvasive detection of many tumors that express bombesin (BN)/gastrin-releasing peptide (GRP) receptors. 99mTc-labeled peptides, however, have a tendency to accumulate in the liver and intestines due to hepatobiliary clearance as a result of the lipophilicity of the 99mTc chelates. This makes the imaging of lesions in the abdominal area difficult. In this study, we have synthesized a new high affinity 99mTc-labeled BN analogue, [DTPA1, Lys3(99mTc-Pm-DADT), Tyr4]BN, having a built-in pharmacokinetic modifier, DTPA, and labeled with 99mTc using a hydrophilic diaminedithiol chelator (Pm-DADT) to effect low hepatobiliary clearance. In vitro binding studies using human prostate cancer PC-3 cell membranes showed that the inhibition constant (Ki) for [DTPA1, Lys3(99Tc-Pm-DADT), Tyr4]BN was 4.1 +/- 1.4 nM. Biodistribution studies of [DTPA1, Lys3(99mTc-Pm-DADT), Tyr4]BN in normal mice showed very low accumulation of radioactivity in the liver and intestines (1.32 +/- 0.13 and 4.58 +/- 0.50% ID, 4 h postinjection, respectively). There was significant uptake (7.71 +/- 1.37% ID/g, 1 h postinjection) in the pancreas which expresses BN/GRP receptors. The uptake in the pancreas could be blocked by BN, partially blocked by neuromedin B, but not affected by somatostatin, indicating that the in vivo binding was BN/GRP receptor specific. Scintigraphic images showed specific, high contrast delineation of prostate cancer PC-3 xenografts in SCID mice. Thus, the new peptide has a great potential for imaging BN/GRP receptor-positive cancers located even in the abdomen. PMID- 15656576 TI - Rapid binding of concanavalin A and maltose-polyrotaxane conjugates due to mobile motion of alpha-cyclodextrins threaded onto a poly(ethylene glycol). AB - Maltose-polyrotaxane conjugates (Mal-PRXs), in which maltose-conjugated alpha cyclodextrins (alpha-CDs) are threaded onto a poly(ethylene glycol) (PEG) chain capped with benzyloxycarbonyl-L-tyrosine, were characterized in terms of their molecular motion and the relation to multivalent interactions between the maltose moiety and concanavalin A from Canavalia ensiformis (Con A). Spin-lattice relaxation time (T1) and spin-spin relaxation time (T2) of alpha-CD C(1), maltosyl C(1), and PEG methylene protons in the Mal-PRXs revealed that the mobile motion of alpha-CDs in the polyrotaxane governed the molecular motion of maltosyl groups in alpha-CDs and threading PEG chain. The association constant (Ka) of the Mal-PRXs with 22, 38 and 53% of alpha-CD threading was 5.7 x 10(4), 1.1 x 10(6), and 5.3 x 10(5) (M(-1)-maltose), respectively. The largest Ka value of the Mal PRX with 38% of alpha-CD threading was well correlated with the T1 and T2 values of maltosyl groups and alpha-CD, suggesting that the mobile motion of maltose conjugated alpha-CDs in the Mal-PRX contributes to the highest affinity with Con A. Initial rate of binding with Con A was also governed by the mobile motion of maltose-conjugated alpha-CDs. Therefore, we concluded that both highly molecular motion due to the mobile motion of maltose-conjugated alpha-CDs and multivalency of the Mal-PRXs contributes to inducing rapid Con A binding. PMID- 15656577 TI - Noninvasive molecular imaging of MYC mRNA expression in human breast cancer xenografts with a [99mTc]peptide-peptide nucleic acid-peptide chimera. AB - Human estrogen receptor-positive breast cancer cells typically display elevated levels of Myc protein due to overexpression of MYC mRNA, and elevated insulin like growth factor 1 receptor (IGF1R) due to overexpression of IGF1R mRNA. We hypothesized that scintigraphic detection of MYC peptide nucleic acid (PNA) probes with an IGF1 peptide loop on the C-terminus, and a [99mTc]chelator peptide on the N-terminus, could measure levels of MYC mRNA noninvasively in human IGF1R overexpressing MCF7 breast cancer xenografts in nude mice. We prepared the chelator-MYC PNA-IGF1 peptide, as well as a 4-nt mismatch PNA control, by solid phase synthesis. We imaged MCF7 xenografts scintigraphically and measured the distribution of [99mTc]probes by scintillation counting of dissected tissues. MCF7 xenografts in nude mice were visualized at 4 and 24 h after tail vein administration of the [99mTc]PNA probe specific for MYC mRNA, but not with the mismatch control. The [99mTc]probes distributed normally to the kidneys, livers, tumors, and other tissues. Molecular imaging of oncogene mRNAs in solid tumors with radiolabel-PNA-peptide chimeras might provide additional genetic characterization of preinvasive and invasive breast cancers. PMID- 15656579 TI - Synthesis of a lipid conjugate of SO3Le(a) and its enhancement on liposomal binding to activated platelets. AB - 3'-O-Sulfated Le(a) (SO3Le(a)) is one of the most potent natural oligosaccharide ligands of selectins. The specific interactions between SO3Le(a) and E-/P selectins are critical in the inflammation process. This paper described an efficient synthesis of a lipid conjugate of SO3Le(a) and its combination with phospholipid and cholesterol to form SO3Le(a)-coated liposomes by the freeze-thaw and extrusion method. The size (D = 78 nm) and stability of the resultant glycoliposomes were comparable to that of liposomes without the glycoconjugate. It was further observed that the incorporation of SO3Le(a) into liposomes could significantly enhance their adhesion to activated platelets as a result of the specific binding between SO3Le(a) on the glycoliposome and the P-selectin on activated platelets. The glycoliposome constructs may be useful for antiinflammation or for targeted delivery of drugs to endothelial cells that express E- and P-selectins. PMID- 15656578 TI - Structure-photodynamic activity relationships of substituted zinc trisulfophthalocyanines. AB - To identify optimal features of metalated sulfophthalocyanine dyes for their use as photosensitizers in the photodynamic therapy of cancer, we synthesized a series of alkynyl-substituted trisulfonated phthalocyanines and compared their amphiphilic properties to a number of parameters related to their photodynamic potency. Varying the length of the substituted alkynyl side-chain modulates the hydrophobic/hydrophilic properties of the dyes providing a linear relationship between their n-octanol/water partition coefficients and retention times on reversed-phase HPLC. Aggregate formation of the dyes in aqueous solution increased with increasing hydrophobicity while monomer formation was favored by the addition of serum proteins or organic solvent. Trisulfonated zinc phthalocyanines bearing hexynyl and nonynyl substituents exhibited high cellular uptake with strong localization at the mitochondrial membranes, which coincided with effective photocytotoxicity toward EMT-6 murine mammary tumor cells. Further increase in the length of the alkynyl chains (dodecynyl, hexadecynyl) did not improve their phototoxicity, likely resulting from extensive aggregation of the dyes in aqueous medium and reduced cell uptake. Aggregation was evident from shifts in the electronic spectra and reduced capacity to generate singlet oxygen. When monomerized through the addition of Cremophor EL all sulfonated zinc phthalocyanines gave similar singlet oxygen yields. Accordingly, differences in the tendency of the dyes to aggregate do not appear to be a determining factor in their photodynamic potency. Our results confirm that the latter in particular relates to their amphiphilic properties, which facilitate cell uptake and intracellular localization at photosensitive sites such as the mitochondria. Combined, these factors play a significant role in the overall photodynamic potency of the dyes. PMID- 15656580 TI - Affinity capture of a mammalian DNA polymerase beta by inhibitors immobilized to resins used in solid-phase organic synthesis. AB - The application of resins normally used in solid-phase organic synthesis to the affinity capture of a mammalian DNA polymerase beta (pol beta) is reported. Lithocholic acid (LCA), an inhibitor of pol beta, was immobilized on various solid supports, and the batch affinity purification of pol beta from a mixture of proteins using these LCA-immobilized resins was examined. Of the resins tested, TentaGel was the most effective at purifying pol beta and at resisting nonspecific absorption of proteins. The immobilized LCA recognized pol beta specifically, which resulted in pol beta binding to the resin. Using the LCA immobilized resin, it was possible to purify pol beta from a mixture of proteins. Furthermore, it was possible to concentrate pol beta from a crude nuclear extract of human T lymphoma Molt4 cells. To facilitate the immobilization of compounds on TentaGel resins, we also designed and prepared photoaffinity beads containing a photoreactive group at the free termini of the TentaGel resin. The pol beta inhibitors LCA, C18-beta-SQDG, and epolactaene were immobilized on the photoaffinity beads by photoreaction. The batch affinity purification of pol beta from a protein mixture could be also achieved with these beads. PMID- 15656581 TI - Synthesis and in vitro characterization of organometallic rhenium and technetium glucose complexes against Glut 1 and hexokinase. AB - A series of nine organometallic technetium-99m and rhenium complexes of glucose are presented and characterized in vitro regarding their potential as surrogates of [18F]-2-fluoro-desoxy glucose ([18F]-FDG). The glucose derivatives are functionalized at positions C-1, C-2, C-3, and C-6. Different spacer lengths and chelating systems have been introduced at these sites. For the (radio)labeling, the organometallic precursors [99mTc(H2O)3(CO)3]+ and [ReBr3(CO)3](2-) respectively have been used. The resulting complexes have been characterized chemically and radiochemically. The formation of uniform products has been observed on the macroscopic (Re) and no-carrier-added level (99mTc). The Tc-99m complexes revealed good inertness against ligand exchange (Cys and His) and excellent stability in physiological buffered saline as well as in human plasma over a period of 24 h at 37 degrees C. The rhenium complexes have been tested for competitive inhibition of the (yeast) hexokinase. Only for C-2 derivatized glucose complexes with extended spacer functionalities Ki values in the millimolar and sub-millimolar range have been observed. In silico molecular docking experiments supported these experimental findings. However, the competitive inhibitors are not recognized as a pseudosubstrate of hexokinase. The cellular uptake of all 99mTc-complexes into HT-29 colon carcinoma cells via Glut1 was generally low and unspecific independent of the position at the hexose ring, the chelating systems, or the overall charge of the corresponding metal complexes. The current results seem to preclude the use of these compounds as [18F]-FDG surrogates primarily due to the low cellular uptake via Glut1. PMID- 15656582 TI - Characterization of site-specific ScFv PEGylation for tumor-targeting pharmaceuticals. AB - New radiopharmaceuticals are possible using site-specific conjugation of small tumor binding proteins and poly(ethylene glycol) (PEG) scaffolds to provide modular multivalent, homo- or heterofunctional cancer-targeting molecules having preferred molecular size, valence, and functionality. Residence time in plasma can be optimized by modification of the size, number, and charge of the protein units. However, random PEG conjugation (PEGylation) of these small molecules via amine groups has led to variations of structural conformation and binding affinity. To optimize PEGylation, scFvs have been recombinantly produced in a vector that adds an unpaired cysteine (c) near the scFv carboxy terminus (scFv c), thus providing a specific site for thiol conjugation. To evaluate the general applicability of this unpaired cysteine for PEGylation of scFv-c, conjugation efficiency was determined for four different scFvs and several PEG molecules having thiol reactive groups. The effect of the PEG molecular format on scFv-c PEG malignant cell binding was also addressed. ScFvs produced as scFv-c and purified by anti E-TAG affinity chromatography were conjugated using PEG molecules with maleimide (Mal) or o-pyridyl disulfide (OPSS). Conjugations were performed at pH 7.0, with 2 molar excess TCEP/scFv and PEG-(Mal) or PEG-OPSS, using 5:1 (PEG/scFv). PEG-Mal conjugation efficiency was also evaluated with 1:5 (PEG/scFv). PEGylation efficiency was determined for each reaction by quantitation of the products on SDS-PAGE. ScFv-c conjugation with unifunctional maleimide PEGs resulted in PEG conjugates incorporating 30-80% of the scFv-c, but usually above 50%. Efficiency of scFv-c conjugation to both functional groups of the bifunctional PEG-(Mal)2 varied between the PEG and scFv-c molecules studied. A maximum of 45% of scFv-c protein was conjugated as PEG- (scFv-c)2 using the smallest PEG-(Mal)2 (2 kDa). No significant increase in scFv-c conjugation was observed by the use of greater than a 5 molar excess of PEG/scFv-c. Under the same conjugation conditions, PEG as OPSS yielded less than 10% PEG-scFv-c. PEG (scFv)2 conjugates had increased binding in ELISA using malignant cell membranes, when compared with unmodified scFv-c. PEGylated-scFv binding was comparable with unmodified scFv-c. In summary, scFv-c can be PEGylated in a site-specific manner using uni- or bivalent PEG-Mal, either linear or branched. ScFv-c was most efficiently conjugated to smaller PEG-Mal molecules, with the smallest, 2 kDa PEG Mal, usually PEGylating 60-90% of the scFv-c. ScFv-c conjugation to form PEG (scFv-c)2 reached greatest efficiency at 45%, and its purified form demonstrated greater binding than the corresponding scFv-c. PMID- 15656583 TI - Preparation and biological characterization of polymeric micelle drug carriers with intracellular pH-triggered drug release property: tumor permeability, controlled subcellular drug distribution, and enhanced in vivo antitumor efficacy. AB - A novel intracellular pH-sensitive polymeric micelle drug carrier that controls the systemic, local, and subcellular distributions of pharmacologically active drugs has been developed in this study. The micelles were prepared from self assembling amphiphilic block copolymers, poly(ethylene glycol)-poly(aspartate hydrazone adriamycin), in which the anticancer drug, adriamycin, was conjugated to the hydrophobic segments through acid-sensitive hydrazone linkers. By this polymer design, the micelles can stably preserve drugs under physiological conditions (pH 7.4) and selectively release them by sensing the intracellular pH decrease in endosomes and lysosomes (pH 5-6). In vitro and in vivo studies show that the micelles have the characteristic properties, such as an intracellular pH triggered drug release capability, tumor-infiltrating permeability, and effective antitumor activity with extremely low toxicity. The acquired experimental data clearly elucidate that the optimization of both the functional and structural features of polymeric micelles provides a promising formulation not only for the development of intracellular environment-sensitive supramolecular devices for cancer therapeutic applications but also for the future treatment of intractable cancers with limited vasculature. PMID- 15656584 TI - Role of biotin-binding affinity in streptavidin-based pretargeted radioimmunotherapy of lymphoma. AB - One pretargeting approach to cancer radioimmunotherapy utilizes an antibody streptavidin conjugate that is first localized to the tumor. A "clearing agent" is then administered to remove the excess bioconjugate from blood, followed by injection of the radiolabeled biotin therapeutic. In this study, the role of streptavidin-biotin affinity in this pretargeting system was investigated for the first time in vivo, with a reduced affinity, site-directed streptavidin mutant and with radiolabeled bis-biotin reagents. The S45A streptavidin mutant (SA S45A), which displays a faster off-rate for biotin, was utilized with a bivalent biotin carrier that retains high avidity for the streptavidin mutant. Mice were fed either a normal or biotin-deficient diet, yielding serum endogenous biotin concentrations of 31 nM and 5 nM, respectively. Lymphoma-bearing nude mice pretargeted with 1F5 Antibody-SA-Wild Type (WT) bioconjugates produced (125)I-bis biotin tumor concentrations of 2.2%ID/g and 7.0%ID/g in mice fed normal diets vs biotin-deficient diets. (125)I-bis-biotin tumor concentrations of mice pretargeted with 1F5-SA-S45A were 12%ID/g and 10%ID/g for mice fed normal and biotin-deficient diets, respectively. However, poor clearance of the 1F5-SA-S45A with the biotinylated clearing agent led to high normal organ concentrations of (125)I-bis-biotin. A galactosylated human serum albumin (HSA) modified with bis biotin was then tested, and normal organ (125)I-bis-biotin concentrations were significantly reduced. Tumor-to-organ ratios achieved for 1F5-SA-S45A with the HSA-bis-biotin clearing agent in mice with high serum biotin were similar to those achieved with 1F5-SA-WT in mice with low serum biotin. These results demonstrate that exchange of bound endogenous biotin with lower affinity streptavidin mutants is possible, and that corresponding use of bis-biotin carriers can nearly eliminate the differences in therapeutic radioactivity at the tumor site in animals on normal vs biotin-deficient diets. The results also interestingly demonstrate, however, that improved clearance agents capable of removing the lower affinity streptavidin-antibody conjugate are needed to achieve comparable specificity in tumor to blood or normal organ ratios. PMID- 15656586 TI - PEG-modified protamine with improved pharmacological/pharmaceutical properties as a potential protamine substitute: synthesis and in vitro evaluation. AB - Cardiopulmonary bypass (CPB) procedures are frequently associated with massive inflammatory responses, resulting in a high rate of morbidity and mortality in routine cardiac operations. One recognized attribute of these deleterious responses is the synergic effect of heparin and protamine, which elicit the activation of the complement system in vivo. To circumvent such toxic effects following protamine reversal of heparin anticoagulation in the CPB procedures, we proposed that poly(ethylene glycol) (PEG)-modified protamine could retain the heparin-neutralization ability and yet diminish the induced complement activation by the formed heparin-protamine complexes (HPC), thereby providing highly improved pharmacological properties. PEGylation of protamine was carried out by utilizing N-hydroxysuccinimidyl (NHS) conjugation chemistry. Size exclusion chromatography (SEC), reverse-phase high performance liquid chromatography (RP HPLC), and matrix-assisted laser desorption mass spectrometry (MALDI-MS) were used to assess the conjugation stiochiometry, the purity of the conjugates, and the site of PEG modification, respectively. The heparin-neutralizing activity was determined by using heparin affinity chromatography and various biological assays including the plasma-activated partial thromboplastin time (aPTT), anti-Xa, and anti-IIa methods. The potency in inducing complement activation was examined in vitro using the CH50 hemolytic assay. The PEG-modified protamine was successfully synthesized with a PEG/protamine stiochiometry of 1:1. Only one conjugation site for PEG that was located at the N-terminal end of protamine was obtained. In the biological evaluations, the PEG-modified protamine displayed a full retention of the heparin-neutralizing ability of protamine and a significantly reduced activity in complement activation following its complexation with heparin. Results from studies of the particle size and zeta potential indicated that the PEG-modified protamine formed substantially smaller aggregates with heparin, rendering them less effective in triggering the size-dependent complement responses. As with protamine, PEG-modified protamine exhibited an enhanced aqueous solubility, therefore attaining significantly improved pharmaceutical properties. These preliminary results suggested that the PEG-modified protamine conjugate might serve as a potential protamine substitute with improved therapeutic and pharmaceutical properties in heparin reversal. PMID- 15656585 TI - Amphiphilic core-shell nanoparticles with poly(ethylenimine) shells as potential gene delivery carriers. AB - Spherical, well-defined core-shell nanoparticles that consist of poly(methyl methacrylate) (PMMA) cores and branched poly(ethylenimine) shells (PEI) were synthesized via a graft copolymerization of methyl methacrylate from branched PEI induced by a small amount of tert-butyl hydroperoxide. The PMMA-PEI core-shell nanoparticles were between 130 to170 nm in diameter and displayed zeta-potentials near +40 mV at pH 7 in 1 mM aqueous NaCl. Plasmid DNA (pDNA) was mixed with nanoparticles and formed complexes of approximately 120 nm in diameter and was highly monodispersed. The complexes were characterized with respect to their particle size, zeta-potential, surface morphology, and DNA integrity. The complexing ability of the nanoparticles was strongly dependent on the molecular weight of the PEI and the thickness of the PEI shells. The stability of the complexes was influenced by the loading ratio of the pDNA and the nanoparticles. The condensed pDNA in the complexes was significantly protected from enzymatic degradation by DNase I. Cytotoxity studies using MTT colorimetric assays suggested that the PMMA-PEI (25 kDa) core-shell nanoparticles were three times less toxic than the branched PEI (25 kDa). Their transfection efficiencies were also significantly higher. Thus, the PEI-based core-shell nanoparticles show considerable potential as carriers for gene delivery. PMID- 15656587 TI - Cationic lipids with increased DNA binding affinity for nonviral gene transfer in dividing and nondividing cells. AB - Effect of headgroup structure on catonic lipid-mediated transfection was investigated with either a (i) tertiary amine, (ii) quaternary amine with a hydroxyl, or (iii) quaternary amine with mesylate as headgroups. Liposomes were formulated using cholesterol or dioleoyl phosphatidyl ethanolamine (DOPE) as colipids, and transfection efficiencies were determined in rapidly dividing colon carcinoma (CT 26) and rat aortic smooth muscle (RASM) cells as well as in nondividing human pancreatic islets using luciferase and green fluorescent protein expression plasmids, pcDNA3-Luc and pCMS-EGFP, respectively. Liposome/pDNA complexes were evaluated for DNA conformational state by circular dichroism (CD), DNA condensation by electrophoretic mobility shift assay (EMSA), particle size and zeta potential by laser diffraction technique, and surface morphology by transmission electron microscopy (TEM). Encouraging transfection results were obtained with the mesylate headgroup based lipid in liposome formulations with DOPE as a colipid, which were higher than the commercially available Lipofectamine formulation. We hypothesize that the additional hydrogen bonding or covalent interactions of the headgroup with the plasmid DNA, leading to higher binding affinity of the cationic lipids to pDNA, results in higher transfection. This hypothesis is supported by TEM observations where elongated complexes were observed and more lipid was seen associated with the DNA. PMID- 15656588 TI - Template-directed assembly and characterization of metallosalen-DNA conjugates. AB - Nucleic acid template-directed synthesis represents a powerful method for the encoded synthesis of new bioconjugates. Our laboratory previously reported a strategy for the synthesis of a new metal-DNA hybrid, metallosalen-DNA, by the DNA or RNA template-directed cross-linking of two salicylaldehyde-modified DNA oligonucleotides. The current manuscript describes the optimal assembly requirements and biophysical characterization of metallosalen-DNA conjugates containing nickel and manganese ions. Competitive assembly reactions demonstrated the template-directed nature of metallosalen-DNA formation. A single metallosalen DNA conjugate was assembled selectively in the presence of two pairs of salicylaldehyde precursor strands and a single DNA template. Assembly reactions were sensitive to base pair mismatches in the pairing arms. Single base mismatches resulted in a loss of metallosalen-DNA conjugate yield. Metallosalen DNA assembly yields depended on the identity and length of the template spacer, the reaction pH, and the type of metal and diamine utilized in the assembly reaction. Metallosalen-DNA conjugates were stable to a variety of conditions, including extended incubation at 50 degrees C. Nickel metallosalen-DNA remained unchanged after incubation at 80 degrees C for 24 h, while decomposition of manganese metallosalen-DNA was observed under the same conditions. Circular dichroism (CD) spectroscopy indicated that DNA duplexes containing internal metallosalen moieties adopted B-form double helices. UV thermal denaturation analysis demonstrated that 32-nucleotide duplexes containing internal metallosalen modifications displayed melting temperatures approximately 5 degrees C less than unmodified DNA duplexes. PMID- 15656589 TI - Synthesis of a metallopeptide-PNA conjugate and its oxidative cross-linking to a DNA target. AB - A nickel(II)-PNA bioconjugate was prepared by formation of a salicylaldimine complex with the amino terminus of a peptide-PNA hybrid with the sequence Arg-His Gly-[TACCTAGCAT]PNA-Arg-CONH2. Hybridization to complementary oligodeoxynucleotides was demonstrated, and covalent adduct formation was observed upon addition of KHSO5 as oxidant. In the absence of PNA, the reactivity of the phenolic radical generated as an intermediate was found to be G >> T >> C, A; by inclusion of the PNA delivery agent, cross-links between the two oligomers could be observed with T and C bases in the vicinity of the nickel complex, although G was still the most reactive site. The metal complex could be removed by treatment with EDTA following which the Schiff base linkage was readily hydrolyzed. The final result in this case is a salicylaldehyde moiety appended at the target site in DNA. PMID- 15656590 TI - Synthesis of new bivalent peptides for applications in the Affinity Enhancement System. AB - The feasibility of two-step radioimmunotherapy (RIT) of cancer by the Affinity Enhancement System (AES) has been demonstrated in experimental and clinical studies. This technique, associating a bispecific antibody and a bivalent peptide radiolabeled with iodine-131, has been developed to reduce toxicity and to improve therapeutic efficacy compared to one-step targeting methods. The use of AES with different beta-emitters such as rhenium-188, samarium-153, or lutetium 177 or alpha-emitters such as actinium-225 or bismuth-213 is now considered. Thus three new peptides, designed to allow for the coupling of a variety of bifunctional chelating agents BCA, were synthesized by associating two glycyl succinyl-histamine (GSH) arms, which are recognized by the 679 monoclonal antibody (mAb-679), with different binding agents, such as p-nitrophenylalanine or N,N-bis(carboxymethyl)-4-N'-(9-fluorenylmethyloxycarbonyl)aminobenzylamine. Immunoreactivity and serum stability evaluation were performed for each synthesized peptide. One of the three peptides (LM218) proved to be more stable than the others, and three different BCAs were coupled to LM218 (CITC-DTPA, CITC TTHA, and CITC-CHXA''DTPA). One of these products, LM218-BzTTHA was radiolabeled with indium-111 without loss of immunoreactivity toward the mAb-679. These new peptides will allow pretargeted RIT with a large variety of radionuclides, to adapt the choice of the radionuclide (LET, half-life, penetrating emission) to the nature and size of targeted tumors. PMID- 15656591 TI - Fluorescent nanometer microspheres as a reporter for sensitive detection of simulants of biological threats using multiplexed suspension arrays. AB - We succeeded in using 40 nm FRET (fluorescence resonance energy transfer) microspheres conjugated to antibodies as the fluorescent reporters to perform the multiplexing suspension array measurements on two simulants of biological threats, ricin (A chain) and a crude spore preparation of Bacillus globigii (Bg). The microspheres were impregnated with two types of fluorophores in equal number (approximately 140 fluorophores in total per microsphere) and displayed bright PE like fluorescence via a fluorescence resonance energy transfer mechanism. Activated microspheres (aldehyde groups) were directly coupled to antibodies and used to form sandwich-type immunoassays in a suspension array. For the crude preparations of Bg, the assay sensitivity using antibody-conjugated microspheres is an order of magnitude higher than that using the conventional fluorescent reporter, R-phycoerythrin (PE). Using the microspheres, Bg at the concentration of 5 ng/mL can be easily detected. For ricin, the assay sensitivity was similar to that obtained using PE as the reporter, but washing the reaction mixtures resulted in the fluorescence signals that were 2-3 times higher compared to those using PE. Ricin at a concentration of 1 ng/mL can be readily identified. Importantly, the two simulants do not interfere with each other in the multiplexing experiments. The 40 nm FRET microspheres are a new sensitive alternative as fluorescent reporters for detection in suspension arrays. PMID- 15656592 TI - A long-acting, highly potent interferon alpha-2 conjugate created using site specific PEGylation. AB - Recombinant interferon alpha-2 (IFN-alpha2) is used clinically to treat a variety of viral diseases and cancers. IFN-alpha2 has a short circulating half-life, which necessitates frequent administration to patients. Previous studies showed that it is possible to extend the circulating half-life of IFN-alpha2 by modifying lysine residues of the protein with amine-reactive poly(ethylene glycol) (PEG) reagents. However, amine-PEGylated IFN-alpha2 comprises a heterogeneous product mixture with low specific activity due to the large number and critical locations of lysine residues in IFN-alpha2. In an effort to overcome these problems we determined the feasibility of creating site-specific, mono PEGylated IFN-alpha2 analogues by introducing a free (unpaired) cysteine residue into the protein, followed by modification of the added cysteine residue with a maleimide-PEG reagent. IFN-alpha2 cysteine analogues were expressed in Escherichia coli and purified, and their in vitro bioactivities were measured in the human Daudi cell line growth inhibition assay. Several cysteine analogues were identified that do not significantly affect in vitro biological activity of IFN-alpha2. Certain of the cysteine analogues, but not wild-type IFN-alpha2, reacted with maleimide-PEG to produce mono-PEGylated proteins. The PEG-Q5C analogue retained high in vitro bioactivity (within 3- to 4-fold of wild-type IFN alpha2) even when modified with 20- and 40-kDa PEGs. Pharmacokinetic experiments indicated that the 20-kDa PEG-Q5C and 40-kDa PEG-Q5C proteins have 20-fold and 40 fold longer half-lives, respectively, than IFN-alpha2 following subcutaneous administration to rats. These studies demonstrate the feasibility of using site specific PEGylation technology to create a long-acting, mono-PEGylated IFN-alpha2 protein with high specific activity. PMID- 15656594 TI - A new highly efficient photoreactive analogue of dCTP. Synthesis, characterization, and application in photoaffinity modification of DNA binding proteins. AB - A new base-substituted analogue of dCTP, exo-N-{2-[N-(4-azido-2,5-difluoro-3 chloropyridine-6-yl)-3-aminopropionyl]aminoethyl}-2'-deoxycytidine-5' triphosphate (FAP-dCTP) has been synthesized and characterized. FAP-dCTP is an efficient substrate of mammalian DNA polymerase beta in the reaction of primer elongation displaying substrate properties as an analogue of dCTP and dTTP. FAP dCTP was used for the photoaffinity modification of mammalian DNA polymerase beta. Two approaches to photoaffinity labeling were utilized. In one approach, photoreactive FAP-dCTP was first incorporated into radiolabeled primer-template, and photoreactive DNA was UV-irradiated in the presence of DNA polymerase beta, which resulted in the polymerase labeling by photoreactive primer. In an alternate approach, FAP-dCTP was first UV-cross-linked to the enzyme; subsequently, radiolabeled primer-template was added, and the enzyme-linked FAP dCTP was incorporated into the 3'-end of radioactive primer. This "catalytic" modification pathway was shown to be less specific in recognition of FAP-dCTP as an analogue of dCTP than dTTP. FAP-dCTP was used as substrate of endogenous DNA polymerases of HeLa cell extract to synthesize photoreactive DNAs for photoaffinity modification of cell proteins. UV irradiation results in modification of DNA binding proteins of cell extract. The level of photoaffinity labeling of protein targets in the cell extract was strongly dependent on the efficiency of synthesis of photoreactive DNA. PMID- 15656593 TI - Transfection activity of polyamidoamine dendrimers having hydrophobic amino acid residues in the periphery. AB - We designed poly(amidoamine) dendrimers with phenylalanine or leucine residues at their chain ends. Thereby, we achieved efficient gene transfection of cells through synergy of the proton sponge effect, which is induced by the internal tertiary amines of the dendrimer, and hydrophobic interaction by the hydrophobic amino acid residues in the dendrimer periphery. Dendrimers having 16, 29, 46, and 64 terminal phenylalanine residues were prepared by the reaction of the amine terminated poly(amidoamine) G4 dendrimer and L-phenylalanine using condensing reagent 1,3-dicyclohexylcarbodiimide. Transfection activity of these phenylalanine-modified dendrimers for CV1 cells, an African green monkey kidney cell line, increased concomitant with the increasing number of the terminal phenylalanine residues, except for the dendrimer with 64 phenylalanine residues, which showed poor water solubility and hardly formed a complex with DNA at neutral pH. However, under weakly acidic conditions, the dendrimer with 64 phenylalanine residues formed a complex with DNA, thereby achieving highly efficient transfection. In contrast, the attachment of L-leucine residues was unable to improve the transfection activity of the parent dendrimer, probably because of the relatively lower hydrophobicity of this amino acid. The phenylalanine-modified dendrimer exhibited a higher transfection activity and a lower cytotoxicity than some widely used transfection reagents. For that reason, the phenylalanine-modified dendrimers are considered to be promising gene carriers. PMID- 15656595 TI - Synthesis and characterization of poly(ethylene oxide)-block-poly(methylidene malonate 2.1.2) block copolymers bearing a mannose group at the PEO chain end. AB - A poly(ethylene oxide)-block-poly(methylidene malonate 2.1.2) block copolymer (PEO-b-PMM 2.1.2) bearing a mannose moiety at the poly(ethylene oxide) chain end was synthesized by sequential anionic polymerization of ethylene oxide (EO) and methylidene malonate 2.1.2 (MM 2.1.2), followed by a coupling reaction between its poly(ethylene oxide) amino- or aldehyde-end group and a sugar derivative. Different coupling procedures, either in organic media or in aqueous micellar solutions, were examined in order to optimize the poly(ethylene oxide) end glycosylation yield. The micellar size of the functionalized block copolymers was determined by dynamic light scattering. PMID- 15656596 TI - Copoly(styrene-maleic acid)-pirarubicin micelles: high tumor-targeting efficiency with little toxicity. AB - The copolymer of styrene-maleic acid (SMA) was used to construct micelles containing pirarubicin (4'-O-tetrahydropyranyladriamycin, or THP) as a new anticancer drug formulation. The procedure for the preparation of the micelles was simple, the component consisting of only SMA and pirarubicin in a noncovalent association, possibly by hydrophobic interaction between the styrene portion of SMA and pirarubicin chromophore. This method ensures more than 80% recovery of pirarubicin by weight, and 60% of drug loading (by weight) was achieved. The micelles obtained (SMA-THP) showed high solubility in water and a constant pirarubicin release rate of about 3-4%/day in vitro. SMA-THP micelles had an average molecular size of about 34 kDa according to gel chromatography; this size is a marked increase from the 627.6 Da of free THP, which suggests the formation of a micellar structure. When albumin was added, the molecular size of the micelles increased to about 94 kDa, which indicates binding to albumin, a unique characteristic of SMA. SMA-THP micelle preparation had a cytotoxic effect (93 101%) on MCF-7 breast cancer cells and SW480 human colon cancer cells in vitro that was comparable to that of free THP. An in vivo assay of SMA-THP at doses of 20 mg/kg in ddY mice bearing S-180 tumor revealed complete tumor eradication in 100% of tested animals. Mice survived for more than 1 year after treatment with micellar drug doses as high as 100 mg/kg pirarubicin equivalent. This marked antitumor activity can be attributed to the enhanced permeability and retention (EPR) effect of macromolecular drugs seen in solid tumors, which enables selective delivery of drugs to tumor and thus much fewer side effects. Complete blood counts, liver function test, and cardiac histology showed no sign of adverse effects for intravenous doses of the micellar preparation. These data thus suggest that intravenous administration of the SMA-THP micellar formulation can enhance the therapeutic effect of pirarubicin more than 50-fold. PMID- 15656597 TI - Conjugation of DOTA using isolated phenolic active esters: the labeling and biodistribution of albumin as blood pool marker. AB - A convenient method for the functionalization of proteins with DOTA (1,4,7,10 tetraazacyclododecane-1,4,7,10-tetraacetic acid) has been developed. For this purpose DOTA was converted into a series of different monoreactive activated phenolic esters. The esters were prepared in a single step from commercially available DOTA, using 1-ethyl-3-[3-(dimethylamino)propyl]carbodiimide or 1,3 dicyclohexylcarbodiimide as coupling agent. The resulting activated esters were isolated by HPLC, lyophilized, and stored for future applications. In solid form the compounds exhibit high hydrolytic stability. The reactions with proteins proceeded in good yields. The conjugation and subsequent radiolabeling of the 4 nitrophenol ester of DOTA with 67Ga was investigated with rat serum albumin. A time-dependent biodistribution study in tumor bearing rats was conducted to demonstrate the integrity of the albumin conjugate. These results suggest that phenolic esters of DOTA represent versatile reagents to conjugate DOTA with proteins and other biomolecules in high yields. PMID- 15656598 TI - A high molar extinction coefficient sensitizer for stable dye-sensitized solar cells. AB - An amphiphilic heteroleptic polypyridyl ruthenium complex with a high molar extinction coefficient was synthesized and demonstrated as an efficient, thermostable sensitizer in nanocrystalline dye-sensitized solar cells. PMID- 15656599 TI - Complementary quadruple hydrogen bonding in supramolecular copolymers. AB - This paper describes a supramolecular copolymer based on quadruple hydrogen bonding between self-complementary ureido-pyrimidinones and complementary 2,7 diamido-1,8-naphthyridines. In contrast to previously reported polymer systems solely based on complementary hydrogen-bonding units, these polymers retain a high DP over a broad composition range. PMID- 15656600 TI - Near-IR luminescence of monolayer-protected metal clusters. AB - Visible-near-IR luminescence spectra of gold MPCs that are similar, irrespective of the number of core atoms (all <2 nm diameter) and different monolayers, are reported. The luminescence can be quantitatively invoked by introducing polar ligands into nonpolar MPC monolayers and by galvanic exchange of metal atoms on the MPC core surface with different metals. The observed emissions are believed to result from surface-localized states that depend on both the core metal of the nanoparticle and the ligands attached to the metal surface. PMID- 15656601 TI - Bonding in HNO-myoglobin as characterized by X-ray absorption and resonance raman spectroscopies. AB - The EXAFS and resonance Raman spectra on the HNO-myoglobin adduct, 1, are consistent with the presence of HNO bound to a heme center. The three-dimensional structure about the heme center of 1 obtained from multiple-scattering (MS) analysis of the EXAFS of the heme protein yielded an Fe-N-O bond angle of 131 degrees and an Fe-N bond length of 1.82 A, which compare well with published values for model complexes containing RNO ligands. Resonance Raman spectra identified the nu(N=O) stretch at 1385 cm-1 (confirmed by 15N labeling), which corresponds well with those reported for small molecule HNO complexes. The wavelength of the nu(Fe-N) at 636 cm-1 of 1 is significantly higher than those of MbIINO and MbIIINO (554 and 595 cm-1, respectively). The XAFS, XANES, and resonance Raman data are all consistent with the structure deduced from the NMR experiments, providing more detail on the bonding between HNO and the metal center. PMID- 15656602 TI - Guanidinium ion as a symmetrical template in the formation of cubic hydrogen bonded borate networks with the boracite topology. AB - Reaction between boric acid and guanidinium salts in methanolic solution at room temperature in the presence of various bases and anions gives well-formed solvated crystals of composition {[B(OCH3)4]3[C(NH2)3]4}+X-. The products in which X- = Cl- and PF6- are characterized by single-crystal X-ray diffraction. The tetramethoxyborate and guanidinium components form a highly symmetrical (cubic) hydrogen-bonded 3D network having the "boracite topology" [i.e., the (63)(6284) topology]. Highly disordered solvent molecules and anions occupy methyl-surfaced cavities and channels that represent more than 30% of the space. PMID- 15656603 TI - Design and synthesis of zinc-selective chelators for extracellular applications. AB - Zinc (Zn2+) is found in every cell in human bodies. A few millimolar of free Zn2+ exists in the vesicles of presynaptic neurons in the mammalian brain and is released by synaptic activity or depolarization, modulating the function of certain ion channels and receptors. Although various chemical tools for measuring Zn2+ in biological samples, such as fluorescent probes for Zn2+, have been developed, Zn2+-selective chelators have room to be improved. Research on Zn2+ signals in the brain has traditionally employed several chelators, which have several shortcomings for biological applications. Here we report the design, synthesis, and properties of new membrane-impermeable chelators selective for Zn2+ and describe biological applications in hippocampal slices. As a result, our newly designed chelator revealed the first biological implication that presynaptic Zn2+ can be released in the CA1 region. This confirms the utility of these new chelatotrs as extracellular Zn2+ chelators for biological applications. PMID- 15656604 TI - Water-soluble [2.2]paracyclophane chromophores with large two-photon action cross sections. AB - A series of alpha,omega-donor-substituted distyrylbenzene dimers held together by the [2.2]paracyclophane core were designed, synthesized, and characterized. Different substituents were chosen to modulate the strength of the donor nitrogen groups and to allow the molecules to be either neutral and soluble in nonpolar organic solvents or charged and water-soluble. The specific neutral structures are (in order of decreasing donor strength) 4,7,12,15-tetra[N,N-bis(6' ' chlorohexyl)-4'-aminostyryl]-[2.2]paracyclophane (1N), 4,7,12,15-tetra[(N-(6' ' chlorohexyl)carbazol-3'-yl)vinyl]-[2.2]paracyclophane (2N), and 4,7,12,15 tetra[N,N-bis(4' '-(6' ''-chlorohexyl)phenyl)-4'-aminostyryl]-[2.2]paracyclophane (3N). The charged species are 4,7,12,15-tetra[N,N-bis(6' '-(N,N,N trimethylammonium)hexyl)-4'-aminostyryl]-[2.2]paracyclophane octaiodide (1C), 4,7,12,15-tetra[(N-(6' '-(N,N,N-trimethylammonium)hexyl)carbazol-3'-yl)vinyl] [2.2]paracyclophane octaiodide (2C), and 4,7,12,15-tetra[N,N-bis(4' '-(6' '' (N,N,N-trimethylammonium)hexyl)phenyl)-4'-aminostyryl]-[2.2]paracyclophane octaiodide (3C). Two-photon excitation spectra, measured using the two-photon induced fluorescence technique, show in toluene the following trend for the two photon cross sections (delta): 3N > 2N > 1N. In water the delta values follow the same order, 3C approximately 2C > 1C, but are smaller (approximately one-third). Significantly, the fluorescence quantum yield (eta) in water decreases much more for 1, relative to 2 and 3. The two-photon action cross sections (deltaeta) of 2C and 3C are 294 GM and 359 GM, respectively. These values are among the highest reported thus far. These results show that, to maximize the deltaeta in this class of chromophores, one needs to fine-tune the magnitude of the charge transfer character of the excited state, to minimize fluorescence quenching in polar media. PMID- 15656606 TI - A nuclear isotope effect for interfacial electron transfer: excited-state electron injection from Ru ammine compounds to nanocrystalline TiO2. AB - The coordination compounds Ru(deeb)(NH3)4(PF6)2 and Ru(deeb)(NH2(CH2)2NH2)4(PF6)2, where deeb is 4,4'-(CO2CH2CH3)2-2,2'-bipyridine, were synthesized and attached to optically transparent nanocrystalline (anatase) TiO2 films. The compounds were found to be nonemissive in fluid acetonitrile and when attached to TiO2 with excited-state lifetimes <10 ns. Infrared measurements showed the expected isotopic substitution of the deuterated compounds on TiO2 thin films. A small 10-15 mV shift in the RuIII/II reduction potentials was measured upon deuteration. Metal-to-ligand charge-transfer (MLCT) excitation resulted in interfacial electron transfer into the TiO2 semiconductor with quantum yields that were dependent on the excitation wavelength and deuteration of the ammine ligands. The quantum yields were optimized with blue light excitation (417 nm) and deuterium substitution. In contrast, the kinetic rate constants for charge recombination were insensitive to deuteration and the excitation wavelength. Control experiments with Ru(deeb)(bpy)2(PF6)2 indicated that deuteration of the TiO2 surface alone does not affect the injection or recombination processes. A model is proposed wherein electron injection occurs in competition with vibrational relaxation and/or intersystem crossing of the excited states. Exchange of hydrogen by deuterium slows vibrational relaxation and/or intersystem crossing, resulting in higher injection yields. PMID- 15656605 TI - Light regulation of aptamer activity: an anti-thrombin aptamer with caged thymidine nucleobases. AB - "Caged" derivatives of a 15 nucleotide ssDNA anti-thrombin aptamer have been synthesized in which thymidine nucleotides are modified with photolabile protecting groups. One caged thymidine in a key location is enough to completely mask the aptamer's function in respect to their affinity for thrombin and their inhibition of the blood clotting cascade. With light (366 nm) the caging group can be removed, yielding the unmodified active aptamer. PMID- 15656608 TI - Changes in calmodulin main-chain dynamics upon ligand binding revealed by cross correlated NMR relaxation measurements. AB - The fast dynamics of protein backbones are often investigated by nuclear magnetic relaxation experiments that report on the degree of spatial restriction of the amide bond vector. By comparing calmodulin in the peptide-bound and peptide-free states with these classical methods, we observe little difference in the dynamics of the polypeptide main chain (average order parameter decrease of 0.01 unit upon binding). However, when using NMR methods that monitor the mobility of the CO Calpha bond vector, we reveal a significant reduction of dynamics of the protein main chain (average order parameter decrease of 0.048 units). Previous investigations have suggested that the side-chain dynamics is reduced by an average of 0.07 order parameter units upon ligand binding (Lee, A. L.; Kinnear, S. A.; Wand, A. J. Nat. Struct. Biol. 2000, 7, 72-77). The current findings suggest that the change of the CO-Calpha bond vector dynamics is intermediate between the changes in NH and side-chain dynamics and report a previously undetected loss of main-chain entropy. Weak site-to-site correlations between the different motional indicators are also observed. PMID- 15656607 TI - Structural and spectroscopic characterization of (mu-hydroxo or mu-oxo)(mu peroxo)diiron(III) complexes: models for peroxo intermediates of non-heme diiron proteins. AB - (mu-Hydroxo or oxo)(mu-1,2-peroxo)diiron(III) complexes having a tetradentate tripodal ligand (L) containing a carboxylate sidearm [Fe2(mu-OH or mu-O)(mu O2)(L)2]n+ were synthesized as models for peroxo-intermediates of non-heme diiron proteins and characterized by various physicochemical measurements including X ray analysis, which provide fundamental structural and spectroscopic insights into the peroxodiiron(III) complexes. PMID- 15656609 TI - Preparation of Superconducting Na(x)(H2O)yCoO2 using NaMnO4 as the deintercalation and oxidation agent. AB - We have used aqueous NaMnO4 solution as the deintercalation and oxidation agent to treat gamma-Na0.7CoO2 powders and to successfully obtain superconducting sodium cobalt oxyhydrates, Nax(H2O)yCoO2, with onset Tc approximately 4.6 K without using highly toxic Br2/CH3CN solution. Chemical analyses indicate that the sodium content x decreases with increasing concentration of NaMnO4 solution and depends slightly on the immersion time. Unlike using a high concentration of aqueous KMnO4 as the deintercalation and oxidation agent, all the hydrated products are the c approximately 19.6 A phase with bilayers of water molecules intercalated between the CoO2 layers and sodium layers because of the absence of K+ in the Na+ layers. PMID- 15656610 TI - Direct observation of the participation of flavin in product formation by thyX encoded thymidylate synthase. AB - The synthesis of thymine for DNA is catalyzed by the enzyme thymidylate synthase (TS). A family of flavin-dependent TSs encoded by the thyX gene has been discovered recently. These newly discovered TSs require a reducing substrate in addition to 2'-deoxyuridine monophosphate (dUMP) and 5,10 methylenetetrahydrofolate (CH2THF), suggesting that the enzyme-bound flavin is a redox intermediary in catalysis. The oxidation of the reduced flavin of the TS from Campylobacter jejuni has been observed directly upon mixing with dUMP and CH2THF under anaerobic conditions, thus providing the first direct demonstration of its redox role in catalysis. Product analysis showed that the one mole of 2' deoxythymidine monophosphate is formed along with one mole of tetrahydrofolate for each mole of reduced enzyme-bound flavin. The classic TS inactivator 5-fluoro 2'-deoxyuridine monophosphate (FdUMP) was able to bind to the reduced enzyme but was unable to oxidize the flavin, even in the presence of CH2THF. Furthermore, the nucleotide binding site of the enzyme treated with FdUMP and CH2THF was irreversibly blocked, suggesting the formation of a stable substrate adduct analogous to that formed by the well-studied thyA-encoded TS. The formation of inactivated enzyme without flavin oxidation indicates that methylene transfer from the folate to the nucleotide occurs prior to flavin redox chemistry. PMID- 15656611 TI - Unique oxidation reaction of amides with pyridine-N-oxide catalyzed by ruthenium porphyrin: direct oxidative conversion of N-acyl-L-proline to N-acyl-L-glutamate. AB - Oxidations of alkanes, alkenes, and aromatic rings with pyridine N-oxides are efficiently catalyzed by ruthenium porphyrins under mild conditions. We show here that the oxidation of N-acyl cyclic amines with RuIVtetraarylporphyrin dichloride 2,6-substituted pyridine N-oxides directly gives N-acyl amino acids in modest to good yield via oxidative C-N bond cleavage. N-Acylpyrrolidines and N acylpiperidines were converted to N-acyl-gamma-aminobutyric acids and N-acyl delta-aminovaleric acids, respectively. This type of reaction is a novel one in which the C-N bond is cleaved selectively at the less substituted carbon. Notably, the proline residue in proline-containing peptides was selectively converted to glutamate. A large intramolecular kinetic isotope effect (kH/kD = 9.8) was observed in the oxidation of N-benzoyl[2,2,-d2]pyrrolidine, indicating that the reaction should involve an alpha-hydrogen atom abstraction process as the rate-determining step. N-Acylcarbaldehyde, the putative intermediate ring opened form of alpha-hydroxylated N-acyl cyclic amine, was readily oxidized with the oxidizing system to afford the corresponding N-acylamino acid in good yield. Further, lactams (1-methyl-2-pyrrolidone and 1-methyl- 2-piperidone) were also oxidized to give the corresponding imides (1-methylsuccinimide and 1 methylpiperidine-2,6-dione). PMID- 15656612 TI - One-step synthesis of labeled sugar nucleotides for protein O-GlcNAc modification studies by chemical function analysis of an archaeal protein. AB - Herein we present the chemical function analysis of a recombinant sugar nucleotidyltransferase from the hyperthermophile Pyrococcus furiosus and its use in the one-pot synthesis of chloroacetyl- and alkyne-tagged analogues of uridinediphospho-N-acetylglucosamine (UDP-GlcNAc). The gene was originally annotated as a glucose-1-phosphate deoxythymidylyltransferase; however, kinetic analysis of a panel of sugar-1-phosphates with the protein shows that it is better described as a bifunctional protein that synthesizes UDP-GlcNAc from glucosamine-1-phosphate and acetyl coenzyme A (CoA). A new mass-spectrometry based assay for the rapid analysis of the acyltransferase activity demonstrates that the enzyme can also accept cheaper truncated N-acetylcysteamine thioester substrates in place of the natural acetyl CoA. The enzyme can tolerate alkyne or chloride substitutions in the acyl moiety, thereby allowing the facile synthesis of tagged sugar nucleotides for future use in protein O-GlcNAc modification studies. PMID- 15656613 TI - 14-azacamptothecin: a potent water-soluble topoisomerase I poison. AB - On the basis of an analysis of luotonin A and its D-ring deaza analogue as topoisomerase I poisons and topoisomerase I-dependent cytotoxic agents, a novel analogue of the structurally related antitumor antibiotic camptothecin (CPT) was prepared. 14-Azacamptothecin was found to have much greater aqueous solubility than CPT, to inhibit topoisomerase I-mediated DNA relaxation more efficiently than CPT, and to stabilize the covalent binary complex to almost the same extent. 14-Aza CPT was found to be slightly less active than CPT in mediating cytotoxicity toward yeast expressing human topoisomerase I, possibly as a consequence of its greater off-rate from the CPT-topoisomerase I-DNA ternary complex. PMID- 15656614 TI - The pyridine ring of NAD is formed by a nonenzymatic pericyclic reaction. AB - The biosynthesis of quinolinate 3, the precursor to the pyridine ring of NAD, is still poorly understood. Two pathways have been identified, one involving the direct formation of quinolinic acid from aspartate and dihydroxyacetone phosphate, the other requiring a five-step degradation of tryptophan. The final step in this degradation is catalyzed by the non-heme Fe(II)-dependent enzyme 3 hydroxyanthranilate-3,4-dioxygenase (HAD). This enzyme catalyzes the oxidative ring opening of 3-hydroxyanthranilate (1) to 2-amino-3-carboxymuconic semialdehyde (ACMS, 2) which then cyclizes to quinolinate (3). In this communication, we demonstrate the following: (1) cyclization of ACMS to 3 is not HAD catalyzed, (2) the most stable form of ACMS in solution is an all trans isomer which undergoes facile cis to trans isomerization about the C2-C3 and C4 C5 double bonds via transient formation of its enol tautomer (6), (3) a model study on the ring opening of N,N-dimethylcarbamoylpyridinium with hydroxide and methoxide suggests that the cyclization of ACMS occurs by an electrocyclization reaction of its enol tautomer 6. Thus, the biosynthesis of quinolinic acid, by the tryptophan pathway, is likely to be a member of a growing family of natural products whose biosynthesis involves a pericyclic reaction. PMID- 15656615 TI - Resist-free patterning of surface architectures in polymer-based microanalytical devices. AB - The ability to form patterns of chemically reactive surface functionalities in microanalytical devices using a simple photopatterning approach without the need for photoresist-based methods is described. Direct UV exposure of the surfaces of poly(methyl methacrylate), PMMA, and poly(carbonate), PC, microfluidic devices through optical masks leads to the production of patterns of near monolayer quantities of surface carboxylic acid groups as determined by surface coverage, X ray photoelectron spectroscopy, and fluorescence microscopy experiments. Formation of the reactive carboxylic acid groups without significant physical (topographical) damage to the polymer device substrates is achieved by use of low UV fluence and exposure times. Modification of the patterned, surface carboxylic acid groups with metals, thermally responsive polymers, and antibodies results in microfluidic devices possessing metallic interconnects and detection electrodes and the ability to capture intact biological cells and proteins from solution. PMID- 15656616 TI - Reduction of trichloroethylene by outer-sphere electron-transfer agents. AB - A series of well-characterized outer-sphere electron-transfer agents were used to study the reduction of trichloroethylene (TCE). Through comparison of the final product ratios of cis-dichloroethylene (cDCE) to trans-dichloroethylene (tDCE) of the outer-sphere reductants to that observed for vitamin B12, it was found that an outer-sphere electron-transfer mechanism is not the likely major pathway in the reduction of TCE by B12. The results provide kinetic support for a two-step reduction mechanism by outer-sphere reductants that involves both vinyl radicals and vinyl anions. The results suggest that the [cDCE]:[tDCE] ratio can be used as a diagnostic for the mechanism of TCE reduction. PMID- 15656617 TI - Caged phosphoproteins. AB - We present the chemical and biological synthesis of caged phosphoproteins using the in vitro nonsense codon suppression methodology. Specifically, phosphoamino acid analogues of serine, threonine, and tyrosine with a single photocleavable o nitrophenylethyl caging group were synthesized as the amino acyl tRNA adducts for insertion into full-length proteins. For this purpose, a novel phosphitylating agent was developed. The successful incorporation of these bulky and charged amino acids into the alpha-subunit of the nicotinic acetyl choline receptor (nAChR) and the vasodilator-stimulated phosphoprotein (VASP) using an in vitro translation system is reported. PMID- 15656618 TI - The total synthesis of gambierol. AB - This communication describes the total synthesis of the marine polyether toxin, gambierol. This work couples our iterative C-glycoside/enol ether-olefin metathesis strategy to the subunits with a unique olefin metathesis/carbonyl olefination reaction to bring the subunits together. PMID- 15656619 TI - Dihydrogen complexes of electrophilic metal centers: observation of Cr(CO)5(H2), W(CO)5(H2) and [Re(CO)5(H2)]+. AB - Photolysis of dichloromethane solutions of M(CO)6 (M = Cr, W) at low temperature in the presence of hydrogen gas affords W(CO)5(H2) (1) and Cr(CO)5(H2) (2). Complexes 1 and 2 are characterized as dihydrogen complexes based on short T1 values for the hydride resonances and a large HD coupling of 35.3 Hz (W) and 35.8 Hz (Cr) in the HD derivatives. A cationic analogue, [Re(CO)5(H2)]+ (3), was prepared by reaction of Re(CO)5Cl with [Et3Si][B(C6F5)4] in fluorobenzene under hydrogen. Complex 3-d1 exhibits JHD = 33.9 Hz. Complex 3 is strongly acidic, with complete deprotonation by diethyl ether; complexes 1 and 2 are moderately acidic. Deprotonation of 1 is complete in the presence of one equivalent of triethylamine. PMID- 15656620 TI - A family of alkynylgold(III) complexes [AuI(mu-{CH2}2PPh2)2Au(III)(C triple bond CR)2] (R = Ph, tBu, Me3Si): facile and reversible comproportionation of gold(I)/gold(III) to digold(II). AB - The symmetric digold(II)dichloride bis(ylide) complex [Au2Cl2(mu-{CH2}2PPh2)2] reacts with acetylides to form the asymmetric heterovalent gold(I)/gold(III) complexes [AuI(mu-{CH2}2PPh2)2AuIII(CCR)2] [R = Ph, tBu, and SiMe3], the phenyl analogue of which was characterized by X-ray crystallography. These compounds represent the first examples of gold(III) complexes containing two acetylide ligands. [AuI(mu-{CH2}2PPh2)2AuIII(CCPh)2] undergoes a reversible comproportionation reaction upon treatment with [Ag(ClO4)tht] to give the symmetric digold(II) cationic complex [Au2(tht)2(mu-{CH2}2PPh2)2](ClO4)2. If this complex is treated with phenylacetylene in the presence of base, the heterovalent gold(I)/gold(III) complex is re-formed. This reversible interconversion between binuclear gold(I)/gold(III) and digold(II) bis(ylide) complexes is unprecedented. PMID- 15656621 TI - Approaching zero: using fractured crystals in metrology for replica molding. AB - This report presents a simple and convenient method to generate nanoscale fractures (cracks) in smooth, single-crystalline Si substrates. The cracks propagated as approximately straight lines along the {100} crystal planes with controllable length defined by a stabilizing backlayer. Close to its tip, the crack presented a vertical offset of the two planes as step of smoothly decreasing height, ranging from the microscale to the atomic scale. The edges of a crack were in close contact at the tip of the crack but were separated at the edge where the crack was initiated. These steps served as ideal test features for probing the limits of the replication of soft lithography. Analysis of topography of original and replicated features (in "hard" poly(dimethylsiloxane and polyurethane) by atomic force microscopy demonstrated that steps down to 0.4 nm could be reproduced; these features approach the dimensions of atoms. PMID- 15656622 TI - Heterogeneous binding of lipoteichoic acid to the surface of titanium dioxide as determined with 31P solid-state NMR spectroscopy. AB - The adsorption of lipoteichoic acid onto the surface of titanium dioxide is shown to be heterogeneous. 31P CPMAS solid-state NMR reveals two distinct phosphate species. The chemical shift anisotropy, asymmetry parameter, and rotating-frame spin-lattice relaxation suggest that 50% of the phosphates are bound to the surface. The remaining phosphates also exhibit restricted molecular motion, but do not have a direct surface bond. PMID- 15656624 TI - Highly efficient, nonpeptidic oligoguanidinium vectors that selectively internalize into mitochondria. AB - Oligoguanidinium-based cell delivery systems have gained broad interest in the drug delivery field since one decade ago. Thus, arginine-containing peptides as Tat or Antp, oligoarginine peptides, and derived peptoids have been described as shuttles for delivering nonpermeant drugs inside cancer cells. Herein we report a new family of tetraguanidinium cell penetrating vectors efficiently internalized in human tumor cells. Their high internalization, studied by confocal microscopy and flow cytometry, as well as their specific accumulation in mitochondria makes these new vectors likely vehicles for the targeted delivery of anticancer drugs to mitochondria. PMID- 15656623 TI - Mechanism of inactivation of inducible nitric oxide synthase by amidines. Irreversible enzyme inactivation without inactivator modification. AB - Nitric oxide synthases (NOS) are hemoproteins that catalyze the reaction of L arginine to L-citrulline and nitric oxide. N-(3-(Aminomethyl)benzyl)acetamidine (1400W) was reported to be a slow, tight-binding, and highly selective inhibitor of iNOS in vitro and in vivo. Previous mechanistic studies reported that 1400W was recovered quantitatively after iNOS fully lost its activity and modification to iNOS was not detected. Here, it is shown that 1400W is a time-, concentration , and NADPH-dependent irreversible inactivator of iNOS. HPLC-electrospray mass spectrometric analysis of the incubation mixture of iNOS with 1400W shows both loss of heme cofactor and formation of biliverdin, as was previously observed for iNOS inactivation by another amidine-containing compound, N5-(1-iminoethyl)-L ornithine (L-NIO). The amount of biliverdin produced corresponds to the amount of heme lost by 1400W inactivation of iNOS. A convenient MS/MS-HPLC methodology was developed to identify the trace amount of biliverdin produced by inactivation of iNOS with either 1400W or L-NIO to be biliverdin IXalpha out of the four possible regioisomers. Two mechanisms were previously proposed for iNOS inactivation by L NIO: (1) uncoupling of the heme peroxide intermediate, leading to destruction of the heme to biliverdin; (2) abstraction of a hydrogen atom from the amidine methyl group followed by attachment to the heme cofactor, which causes the enzyme to catalyze the heme oxygenase reaction. The second mechanistic proposal was ruled out by inactivation of iNOS with d3-1400W, which produced no d2-1400W. Detection of carbon monoxide as one of the heme-degradation products further excludes the covalent heme adduct mechanism. On the basis of these results, a third mechanism is proposed in which the amidine inactivators of iNOS bind as does substrate L-arginine, but because of the amidine methyl group, the heme peroxy intermediate cannot be protonated, thereby preventing its conversion to the heme oxo intermediate. This leads to a change in the enzyme mechanism to one that resembles that of heme oxygenase, an enzyme known to convert heme to biliverdin IXalpha. This appears to be the first example of a compound that causes irreversible inactivation of an enzyme without itself becoming modified in any way. PMID- 15656625 TI - Relation between pore sizes of protein crystals and anisotropic solute diffusivities. AB - The diffusion of a solute, fluorescein, into lysozyme protein crystals with different pore structures was investigated. To determine the diffusion coefficients, three-dimensional solute concentration fields acquired by confocal laser scanning microscopy (CLSM) during diffusion into the crystals were compared with the output of a time-dependent 3-D diffusion model. The diffusion process was found to be anisotropic, and the degree of anisotropy increased in the order: triclinic, tetragonal and orthorhombic crystal morphology. A linear correlation between the pore diffusion coefficients and the pore sizes was established. The maximum size of the solute, deduced from the established correlation of diffusion coefficients and pore size, was 0.73 +/- 0.06 nm, which was in the range of the average diameter of fluorescein (0.69 +/- 0.02 nm). This proves that size exclusion is the key mechanism for solute diffusion in protein crystals. Hence, the origin of solute diffusion anisotropy can be found in the packing of the protein molecules in the crystals, which determines the crystal pore organization. PMID- 15656626 TI - Demonstration of an alternative mechanism for G-to-G cross-link formation. AB - The cross-link dG-to-dG is an important product of DNA nitrosation. Its formation has commonly been attributed to nucleophilic substitution of N2 in a guaninediazonium ion by guanine, while recent studies suggest guanine addition to a cyanoamine derivative formed after dediazoniation, deprotonation, and pyrimidine ring-opening. The chemical viability of the latter mechanism is supported here by the experimental demonstration of rG-to-aG formation via rG addition to a synthetic cyanoamine derivative. Thus, all known products of nitrosative guanine deamination are consistent with the postulate of pyrimidine ring-opening. This postulated mechanism not only explains what is already known but also suggests that other products and other cross-links also might be formed in DNA deamination. The study suggests one possible new product: the structure isomer aG(N1)-to-rG(C2) of the classical G(N2)-to-G(C2) cross-link. While the formation of aG(N2)-to-rG(C2) has been established by chemical synthesis, the structure isomer aG(N1)-to-rG(C2) has been assigned tentatively based on its MS/MS spectrum and because this assignment is reasonable from a mechanistic perspective. Density functional calculations show preferences for the amide iminol tautomer of the classical cross-link G(N2)-to-G(C2) and the amide-amide tautomer of G(N1)-to-G(C2). Moreover, the results suggest that both cross-links are of comparable thermodynamic stability, and that there are no a priori energetic or structural reasons that would prevent the formation of the structure isomer in the model reaction or in DNA. PMID- 15656627 TI - Molecular control of macroscopic cubic, columnar, and lamellar organizations in luminescent lanthanide-containing thermotropic liquid crystals. AB - The connection of lipophilic gallic acid derivatives at the 5,5'- or 6,6' positions of the rigid 2,6-bis(1-ethyl-benzimidazol-2-yl)pyridine core provides two pro-mesogenic tridentate ligands L10 and L12, whose molecular shapes, anisometries, and directional intermolecular pi-stacking can be tuned. X-ray diffraction data in the crystalline state, combined with solution 1H NMR measurements, show that complexation with trivalent lanthanides, Ln(III), produces the neutral hemi-disklike complexes [Ln(Li)(NO3)3] (i = 10, 12), which dimerize to give the rodlike bimetallic complexes [Ln2(Li)2(NO3)6] at lower temperature. The relevant thermodynamic parameters for the latter process depend on the nature of the ligand, the size of the metal ion, and the strength of the intermolecular interactions involved in the condensed phase. These three dimensional models obtained for the complexes in the crystals and in solution are eventually confronted with small-angle XRD profiles recorded in the intermediate thermotropic liquid crystalline phase, in which the rigidity of the packed polyaromatic cores is maintained, while the alkyl chains are molten. According to the specific geometries and nuclearities of the molecular complexes, three types of mesophases (lamellar, columnar, and cubic) can be induced, which provides a direct correlation between the microscopic arrangements and the macroscopic ordering in lanthanide-containing metallomesogens. PMID- 15656628 TI - Nanomolar protein sensing with embedded receptor molecules. AB - A new concept of protein sensing at the air-water interface is introduced, based on amphiphilic receptor molecules embedded in a lipid monolayer. The process begins with incorporation of a small amount (0.13 equiv) of one or two different calix[4]arenes, adorned with charged functional groups at their upper rims, into a stearic acid monolayer. These doped monolayers are subsequently shown to attract peptides and proteins from the aqueous subphase. Depending on the host structure, the monolayers can be made selective for basic or acidic proteins. A working model is proposed, which explains the large observed p/A shifts with reincorporation of excess receptor molecules into the lipid monolayer after complex formation with the oppositely charged protein. This requires a self assembly of multiple calixarene units over the protein surface, which bind the protein in a cooperative fashion. Oppositely charged calixarene derivatives do not form molecular capsules inside the monolayer, but rather remain separate inside the lipid layer, adopting a perpendicular orientation. They combine their hydrogen bond donor and acceptor capacities, and thus markedly enhance the sensitivity of the sensor system toward proteins, pushing the detection limits to 10 pM concentrations. The response pattern obtained from various receptor units inside the monolayer toward the same protein creates a fingerprint for this protein, which can hence be selectively detected at nanomolar concentrations (pattern recognition). PMID- 15656629 TI - Direct observation of ligand binding to membrane proteins in living cells by a saturation transfer double difference (STDD) NMR spectroscopy method shows a significantly higher affinity of integrin alpha(IIb)beta3 in native platelets than in liposomes. AB - About 30% of the proteins in mammalian systems are membrane bound or integrated (e.g., GPCRs). It is inherently difficult to investigate receptor-ligand interactions on a molecular level in their natural membrane environment. Here, we present a new method based on saturation transfer difference (STD) NMR to characterize at an atomic level binding interactions of cell surface proteins in living cells. Implemented as a double difference technique, STD NMR allows the direct observation of binding events and the definition of the binding epitopes of ligands. The binding of the pentapeptide cyclo(RGDfV) to the surface glycoprotein integrin alpha(IIb)beta3 of intact human blood platelets can be detected by saturation transfer double difference (STDD) NMR in less than an hour. A 5-fold higher STD response reflects a significantly higher affinity of integrin alpha(IIb)beta3 in native platelets than in liposomes, which demonstrates the importance of studying membrane proteins in their natural environment. Also, the binding mode of cyclo(RGDfV) in the arginine glycine region is slightly different when interacting with native integrin in platelets compared to integrin reintegrated into liposomes. PMID- 15656630 TI - Pseudo-para-dinitro[2.2]paracyclophane radical anion, a mixed-valence system poised on the Class II/Class III borderline. AB - Analysis of optical spectra of a pseudo-para-dinitro[2.2]paracyclophane radical anion using Marcus-Hush theory reveals that its off diagonal coupling element, H(ab), is large enough, relative to its reorganization energy, to change it from a localized class II compound to a delocalized class III compound by changing solvents. The optical spectra, along with frontier orbital analysis, show that the assumption that E(op) = 2H(ab) for class III compounds is not true in this case. PMID- 15656631 TI - Phase stabilization through electronic tuning: electron-poorer alkali-metal indium compounds with unprecedented In/Li clusters. AB - Three alkali-metal-indium compounds K34In(92.30)Li(12.70) (I), K14Na20In(91.82)Li(13.18) (II), and K14Na20In(96.30) (III) (all Rm) have been synthesized and characterized by structural and physical property measurements and electronic structure calculations. Novel mixed In/Li anionic icosahedra and fused icosahedra form in I and II. All three contain In28 as the first triply fused In icosahedra, which are further linked into (In28)In(In28) sandwich adducts in compounds I and II and (In28)In2(In28) in III. Stabilization of these electron-poorer phases through electronic tuning occurs via two different structural (redox) perturbations, either by substitution of certain indium atoms in the clusters by electron-poorer lithium atoms or by the introduction of defects and disorder in the fused cluster (III). The preferential occurrence of either substitutions or defect formation in the clusters is consistent with extended Huckel band calculation results for both the ideal pure indium phase and the Li-substituted equivalent. Model (ideal) and experimental EF values (based on stoichiometries) fall around a pseudogap in DOS. All three compounds are metallic according to both EHTB band calculations and measured resistivities. The cations (A = K, Na) in all the three structures generate A136 clathrate-IotaIotatype networks with remarkably specific and transferable cation dispositions around the two types of anionic cluster units. PMID- 15656632 TI - Synthesis of dicyclopenta[a,e]pentalenes via a molybdenum carbonyl mediated tandem allenic Pauson-Khand reaction and the X-ray crystal structure of a planar dicyclopenta[a,e]pentalene. AB - Two 14 pi cross-linked annulenes which belong to the family of dicyclopenta[a,e]pentalenes have been synthesized, 14 pi bis enol triflate ester 27 and the 3,7-diisopropylsilyl substituted 14 pi dicyclopenta[a,e]pentalene 30. The new allenic tandem Pauson-Khand reaction mediated by Mo(CO)(6) was employed as the key process to construct the core of the tetracycles. The two linear dicyclopenta[a,e]pentalenes 27 and 30 underwent significant electronic delocalization, perhaps even aromaticity, as revealed by the X-ray structure of 27. The tetracyclic rings in 27 assumed a flat geometry (Figure 4); the bond lengths of the tetracycle in 27 also fit well into the criteria for aromatic compounds. A comparison of the NMR and UV spectra of both 27 and 30 demonstrated that they both exhibited similar electronic properties, therefore, the purple colored 14 pi cross linked annulene 30 is planar as well as delocalized. PMID- 15656633 TI - Solvent-assisted new reaction pathways for the (THF)W(CO)5-promoted endo- and exo cycloisomerization of 4-pentyn-1-ol: a theoretical investigation. AB - New solvent-assisted mechanistic routes were located for the (THF)W(CO)5-promoted endo- and exo-cycloisomerization of 4-pentyn-1-ol using the B3LYP/6-31G (with the LANL2DZ relativistic pseudopotential for W) theory level. A mixed model was used by explicitly including a THF molecule as a component of the reactive system and taking into account the effect of bulk solvent by means of the PCM-UAHF method. This THF molecule plays a crucial role by making the endo route leading to the cyclic carbene complex the most favorable one because of an important catalytic effect. These new mechanisms allow a satisfactory rationalization of experimental facts. PMID- 15656634 TI - Reactivity of peroxo forms of the vanadium haloperoxidase cofactor. A DFT investigation. AB - Density functional theory has been used to investigate structural, electronic and reactivity properties of complexes related to the peroxo forms of vanadium haloperoxidases (VHPO). In particular, the reactivity of the cofactor as a function of protonation state and environment, which are two factors thought to be crucial in modulating the activity of the enzyme, has been examined. In full agreement with experimental data, results highlight the role of protonation in the activation of the peroxo-vanadium complexes and show that the oxo-transfer step involves the unprotonated axial peroxo oxygen atom, which is easily accessible to substrates in the peroxo form of the enzyme. The role of Lys353, which in the X-ray structure of the peroxide-bound form of vanadium chloroperoxidase is hydrogen bonded to the equatorial oxygen atom of the peroxo group, has been also explored. It is concluded that Lys353 can play a role similar to a H+ in the activation of the peroxo form of the cofactor. PMID- 15656635 TI - Single-site anionic polymerization. Monomeric ester enolaluminate propagator synthesis, molecular structure, and polymerization mechanism. AB - The synthesis and molecular structure of the first examples of monomeric lithium ester enolaluminates that serve as structural models for single-site anionic propagating centers, as well as the mechanism of their polymerization of methacrylates catalyzed by conjugate organoaluminum Lewis acids, are reported. Reactions of isopropyl alpha-lithioisobutyrate (2) with suitable deaggregating and stabilizing organoaluminum compounds such as MeAl(BHT)2 (BHT = 2,6-di-tert butyl-4-methylphenolate) in hydrocarbons cleanly generate lithium ester enolaluminate complexes such as Li+[Me2C=C(OiPr)OAlMe(BHT)2]- (3). Remarkably, complex 3 is isolable and exists as a monomer in both solid and solution states. Unlike the uncontrolled polymerization of methacrylates by the aggregating enolate 2, the methacrylate polymerization by the monomeric 3 is controlled but exhibits low activity. However, the well controlled and highly active polymerization can be achieved by using the 3/MeAl(BHT)2 propagator/catalyst pair, which is conveniently generated by in situ mixing of 2 with 2 equiv of MeAl(BHT)2. The structure of the added organoaluminum compounds has marked effects on the degree of monomer activation, enolaluminate formation and reactivity, and polymerization control. Kinetics of the polymerization by the 3/MeAl(BHT)2 pair suggest a bimolecular, activated-monomer anionic polymerization mechanism via single-site ester enolaluminate propagating centers. The molecular structures of activated monomer 1, aggregated initiator 2, and monomeric propagator 3 have been determined by X-ray diffraction studies. PMID- 15656636 TI - On the origin of optical activity in tris-diamine complexes of Co(III) and Rh(III): a simple model based on time-dependent density function theory. AB - Time-dependent density functional theory (TD-DFT) is applied to the CD spectra of Lambda(deltadeltadelta)-(+)-[Co(S-pn)n(en)(3-n)]3+ (n = 1, 2, 3) and Lambda(deltadeltadelta)-(+)-[Co(en)3]3+ as well as the stereoisomers Delta ((delta)n(lambda)(3-n))-(-)-[Co(S-pn)n(en)(3-n)]3+ (n = 1, 2, 3) and Delta(deltadeltadelta)-(-)-[Co(en)3]3+. Theory is able to reproduce the major differences in the CD spectra of the species with a Lambda-configuration and their isomers with a Delta-configuration in both the d-d and ligand-to-metal CT region. It is further possible to rationalize the trend in terms of a larger azimuthal distortion away from the octahedral geometry in the Lambda-conformation compared to the Delta-configuration. Considerations were also given to the CD spectra of the lel3-isomer, Delta(lambdalambdalambda)-(-)-[Rh(R-pn)3]3+ and the ob-isomer, Lambda(lambdalambdalambda)-(+)-[Rh(S-pn)3]3+. PMID- 15656637 TI - Silicon-nitrogen bonding in silatranes: assignment of photoelectron spectra. AB - Silicon-nitrogen bonding and the photoelectron spectra of hydro-silatrane and methyl-silatrane, XSi[OCH2CH2]3N (X = H and Me), were studied with ab initio electron propagator theory, many-body methods, and density functional models. A linear vibronic coupling (LVC) model was employed to estimate vibrational widths of the ionization bands and to study the dependence of the ionization energies on the molecular geometry. Particular attention was given to coordinates that change the Si-N distance and the strength of the donor-acceptor interaction between these two atoms. The ionization energy of the highest occupied molecular orbital has a very strong geometrical dependence which leads to an unusually large vibrational width in the corresponding photoelectron band. The assignment of this band in methyl-silatrane, which was controversial for a long time, is resolved by the present study. The calculated photoelectron spectra allow for clear assignment of at least three more bands in the observed spectra. The present results demonstrate the important role of electrostatic interactions in Si <-- N bonding and in the outer-valence ionization energies of the silatranes. PMID- 15656639 TI - High nuclearity iridium-platinum clusters: synthesis, structures, bonding, and reactivity. AB - The reaction of Ir4(CO)12 with an excess of Pt(PBu(t)3)2 at room temperature yielded the bis-Pt(PBu(t)3) adduct Ir4(CO)12[Pt(PBu(t)3)]2 (9), which contains two Pt(PBu(t)3) groups bridging opposite edges of a central Ir4 pseudotetrahedron. The same reaction at 110 degrees C yielded two new higher nuclearity complexes, Ir8(CO)12[Pt(PBu(t)3)]4 (10) and Ir6(CO)10[Pt(PBu(t)3)]4 (11). Compound 10 consists of a central Ir4(CO)4 tetrahedron with four edge bridging Ir(CO)2 groups and four Pt(PBu(t)3) groups that are each bonded to Ir3 triangles of the Ir4 tetrahedron and two of the Ir(CO)2 groups. Compound 11 consists of a central Ir4(CO)4 pseudotetrahedron with two edge-bridging Ir(CO)2 groups and four Pt(PBu(t)3) groups; one Pt(PBu(t)3) group is bonded to five iridium atoms as found in 10; two are bonded to four iridium atoms, and one is bonded to one of the outer Ir2Pt triangles. Compound 11 reacted with hydrogen at 97 degrees C to give the new tetrahydrido complex Ir6(CO)8[Pt(PBu(t)3)]4(mu-H)4 (12). Compound 12 is formed by the loss of the two bridging carbonyl ligands from 11 and the addition of four hydrido ligands. All four new compounds were characterized by both 1H and 31P NMR and by single-crystal X-ray diffraction analyses. The bonding in 9 was studied by Fenske-Hall molecular orbital calculations, which in this case provides a delocalized bonding description for the Ir-Ir and Ir-Pt bonding, where the attachment of the 0 e- fragments of Pt(PR3) use Ir-Ir bonding orbitals of the Ir4(CO)12 cluster to form multicenter Pt-Ir bonds. PMID- 15656638 TI - Interaction of the human prion PrP(106-126) sequence with copper(II), manganese(II), and zinc(II): NMR and EPR studies. AB - The synthetic peptide encompassing residues 106-126 (PrP106-126, KTNMKHMAGAAAAGAVVGGLG) of the human prion protein was considered for its binding properties toward copper(II), manganese(II) and zinc(II) at pH 5.7. 1H and 13C 1D spectra, 1H spin-lattice relaxation rates, and 1H-15N and 1H-13C HSQC 2D experiments were obtained in the absence and in the presence of metal ions. While Zn(II) was found to yield negligible effects upon any NMR parameter, metal peptide association was demonstrated by the paramagnetic effects of Cu(II) and Mn(II) upon 1D and 2D spectra. Delineation of structures of metal complexes was sought by interpreting the paramagnetic effect on 1H spin-lattice relaxation rates. Exchange of peptide molecules from the metal coordination sphere was shown to provide sizable contribution to the observed relaxation rates. Such contribution was calculated in the case of Cu(II); whereas the faster paramagnetic rates of peptide molecules bound to Mn(II) were determining spin lattice relaxation rates almost exclusively dominated by exchange. Proton-metal distances were therefore evaluated in the case of the Cu(II) complex only and used as restraints in molecular dynamics calculations where from the structure of the complex was obtained. The peptide was shown to bind copper through the imidazole nitrogen and the ionized amide nitrogen of His-111 and the amino terminal group with the terminal carboxyl stabilizing the coordination sphere through ionic interactions. The data were interpreted as to demonstrate that the hydrophobic C-terminal region was not affecting the copper-binding properties of the peptide and that this hydrophobic tail is left free to interact with other target molecules. As for the complex with Mn(II), qualitative information was obtained on carbonyl oxygens of Gly-124 and Leu-125, beyond the terminal Gly-126 carboxyl, being at close distance from the metal ion, that also interacts, most likely, through a hydrogen bond of metal-bound water, with the imidazole ring of His-111. PMID- 15656640 TI - Synthesis, characterization, and structure-selective extraction of 1-3-nm diameter AuAg dendrimer-encapsulated bimetallic nanoparticles. AB - The synthesis and characterization of 1-3-nm diameter, structurally well-defined, bimetallic AuAg dendrimer-encapsulated nanoparticles (DENs) are reported. Three different bimetallic structures were examined: AuAg alloys synthesized by cocomplexation and subsequent reduction of dendrimer-encapsulated Au3+ and Ag+ and core/shell [Au](Ag) and [AuAg alloy](Ag) structures (for structured materials, brackets indicate the core metal and parentheses indicate the shell metal) synthesized by a sequential loading method. Depending on the shell metal and its oxidation state, the AuAg nanoparticles can be extracted from the dendrimer into an organic phase using different surfactants. This provides a means for analyzing the composition of the shell. UV-vis, TEM, and single particle X-ray energy dispersive spectroscopy (EDS) were used to characterize the bimetallic DENs before and after extraction and show that the extraction step does not alter the size or composition of the bimetallic nanoparticles. PMID- 15656641 TI - Substrate hydroxylation in methane monooxygenase: quantitative modeling via mixed quantum mechanics/molecular mechanics techniques. AB - Using broken-symmetry unrestricted density functional theory quantum mechanical (QM) methods in concert with mixed quantum mechanics/molecular mechanics (QM/MM) methods, the hydroxylation of methane and substituted methanes by intermediate Q in methane monooxygenase hydroxylase (MMOH) has been quantitatively modeled. This protocol allows the protein environment to be included throughout the calculations and its effects (electrostatic, van der Waals, strain) upon the reaction to be accurately evaluated. With the current results, recent kinetic data for CH3X (X = H, CH3, OH, CN, NO2) substrate hydroxylation in MMOH (Ambundo, E. A.; Friesner, R. A.; Lippard, S. J. J. Am. Chem. Soc. 2002, 124, 8770-8771) can be rationalized. Results for methane, which provide a quantitative test of the protocol, including a substantial kinetic isotope effect (KIE), are in reasonable agreement with experiment. Specific features of the interaction of each of the substrates with MMO are illuminated by the QM/MM modeling, and the resulting effects upon substrate binding are quantitatively incorporated into the calculations. The results as a whole point to the success of the QM/MM methodology and enhance our understanding of MMOH catalytic chemistry. We also identify systematic errors in the evaluation of the free energy of binding of the Michaelis complexes of the substrates, which most likely arise from inadequate sampling and/or the use of harmonic approximations to evaluate the entropy of the complex. More sophisticated sampling methods will be required to achieve greater accuracy in this aspect of the calculation. PMID- 15656643 TI - Where does the planar-to-nonplanar turnover occur in small gold clusters? AB - Several levels of theory, including both Gaussian-based and plane wave density functional theory (DFT), second-order perturbation theory (MP2), and coupled cluster methods (CCSD(T)), are employed to study Au6 and Au8 clusters. All methods predict that the lowest energy isomer of Au6 is planar. For Au8, both DFT methods predict that the two lowest isomers are planar. In contrast, both MP2 and CCSD(T) predict the lowest Au8 isomers to be nonplanar. PMID- 15656642 TI - Application of molecular absorption properties of horseradish peroxidase for self indicating enzymatic interactions and analytical methods. AB - In this paper an in depth study is presented of the use of the horseradish peroxidase (HRP) enzyme as a self-indicating biorecognition reagent in UV-vis molecular absorption spectrometry. The HRP/H2O2 reaction mechanism in the absence of an external substrate has been clarified, and the interaction between HRP and glucose oxidase (GOx) has been studied. It has been demonstrated that GOx can act as a substrate of HRP; in both cases the kinetic constants have been obtained and mathematical models have been developed. Second, the HRP/H2O2 reaction is used to follow a H2O2-producing enzymatic reaction, the glucose reaction with GOx being used as a model. As an application of this, two methodologies have been proposed for glucose determination: with or without previous incubation of glucose with GOx. In both cases mathematical models relating HRP absorbance changes to glucose concentration have been developed and tested; both methods have been optimized, analytically characterized, and tested for glucose determination in samples. The methodology described could be applied to other heme-proteins and to other H2O2 producing enzymatic reactions. The models permit the reaction constants to be calculated. From the analytical chemistry point of view the models allow the prediction of the method sensitivity for other analytes involved in this type of reaction if the kinetic constants are known and can be used in the design of optical sensors. PMID- 15656644 TI - Glycosidic bond cleavage of pyrimidine nucleosides by low-energy electrons: a theoretical rationale. AB - DNA damage by attachment of low-energy secondary electrons is a very interesting and important mechanism. Electron capture and subsequent base release are thought to be the elementary steps of this mechanism. The process of the N1-glycosidic bond breaking of anion radicals of pyrimidine nucleosides, specifically the 2' deoxyribothymidine (dT) and 2'-deoxyribocytidine (dC) anions, has been investigated theoretically at the B3LYP/DZP++ level of theory. The release of nucleobases by the attachment of low-energy electrons depends on the formation of a stable anion radical of the nucleoside. The lower bond-breaking activation energy and the higher vertical electron detachment energy for dT enables the heterolytic cleavage of the N1-glycosidic bond. However, with the higher bond breaking activation energy and the lower vertical electron detachment energy for dC, the release of cytosine might be impractical when the incident electrons have high kinetic energy. Furthermore, the release of cytosine would have a quantum yield much lower than that of dT when the incident electrons have lower kinetic energy. This study also demonstrates the importance of the proton at O5' of 2' deoxyribose in the base release process. Extending this investigation from dT to dC advances the insight into the mechanism of the N1-glycosidic bond-breaking process. The information from this extensive investigation should be valuable for further experimental studies of cytosine release in irradiated DNA. PMID- 15656646 TI - Validation of a tomato-specific gene, LAT52, used as an endogenous reference gene in qualitative and real-time quantitative PCR detection of transgenic tomatoes. AB - Toward the development of reliable qualitative and quantitative Polymerase Chain Reaction (PCR) detection methods of transgenic tomatoes, one tomato (Lycopersicon esculentum) species specific gene, LAT52, was selected and validated as suitable for using as an endogenous reference gene in transgenic tomato PCR detection. Both qualitative and quantitative PCR methods were assayed with 16 different tomato varieties, and identical amplified products or fluorescent signals were obtained with all of them. No amplified products and fluorescent signals were observed when DNA samples from 20 different plants such as soybean, maize, rapeseed, rice, and Arabidopsis thaliana were used as templates. These results demonstrated that the amplified LAT52 DNA sequence was specific for tomato. Furthermore, results of Southern blot showed that the LAT52 gene was a single copy gene in the different tested tomato cultivars. In qualitative and quantitative PCR analysis, the detection sensitivities were 0.05 and 0.005 ng of tomato genomic DNA, respectively. In addition, two real-time assays employing this gene as an endogenous reference gene were established, one for the quantification of processed food samples derived from nontransgenic tomatoes that contained degraded target DNA and the other for the quantification of the junction region of CaMV35s promoter and the anti-sense ethylene-forming enzyme (EFE) gene in transgenic tomato Huafan No. 1 samples. All of these results indicated that the LAT52 gene could be successfully used as a tomato endogenous reference gene in practical qualitative and quantitative detection of transgenic tomatoes, even for some processed foods derived from transgenic and nontransgenic tomatoes. PMID- 15656647 TI - A metabonomic strategy for the detection of the metabolic effects of chamomile (Matricaria recutita L.) ingestion. AB - A metabonomic strategy, utilizing high-resolution 1H NMR spectroscopy in conjunction with chemometric methods (discriminant analysis with orthogonal signal correction), has been applied to the study of human biological responses to chamomile tea ingestion. Daily urine samples were collected from volunteers during a 6-week period incorporating a 2-week baseline period, 2 weeks of daily chamomile tea ingestion, and a 2-week post-treatment phase. Although strong intersubject variation in metabolite profiles was observed, clear differentiation between the samples obtained before and after chamomile ingestion was achieved on the basis of increased urinary excretion of hippurate and glycine with depleted creatinine concentration. Samples obtained up to 2 weeks after daily chamomile intake formed an isolated cluster in the discriminant analysis map, from which it was inferred that the metabolic effects of chamomile ingestion were prolonged during the 2-week postdosing period. This study highlights the potential for metabonomic technology in the assessment of nutritional interventions, despite the high degree of variation from genetic and environmental sources. PMID- 15656648 TI - Intramolecular carbon isotopic composition of monosodium glutamate: biochemical pathways and product source identification. AB - Monosodium glutamate (MSG) obtained as trade samples from several manufacturers was studied to determine the range of its intramolecular 13C/12C composition. Although the carbon isotopic composition of the total MSG molecule did not differ among manufacturers in most instances, significant differences were observed in the isotopic composition of the alpha-carboxyl carbon, suggesting that different proprietary strains of industrial microorganisms or MSG purification methods may impart unique isotopic fingerprints upon their products. The 13C depletion of the alpha-carboxyl carbon relative to the rest of the molecule helps constrain the identity of the potential anapleurotic carboxylating enzymes responsible for its fixation. PMID- 15656649 TI - Rapid method for simultaneous quantitative determination of four major essential oil components from oregano (Oreganumsp.) and thyme (Thymus sp.) using FT-Raman spectroscopy. AB - A simple and rapid method for the quantitative determination of four major components found in oregano and thyme essential oils is presented. The method correlates the Raman peak intensity in the spectral region from 1800 to 600 cm( 1) and the concentration percentage of each particular constituent in the sample. To achieve accurate quantification results and avoid the risk of overlapping peaks of unknown Raman-active substances in natural essential oils, the peaks must be analyzed. For this purpose, PEAKSOLVE software (Ver. 1.0.5) was used. Unknown samples were measured with the FT-Raman method, and the results were compared to those of the gas chromatographic (GC) analysis. The comparison was made at a confidence level of 99%, and the two methods scored equally in terms of repeatability and accuracy even at the edge of the method specifications. The new method can provide accurate results in very short times once the setup is complete and could be utilized in areas where vast amounts of samples must be analyzed. PMID- 15656650 TI - FT-Raman spectroscopic simultaneous determination of fructose and glucose in honey. AB - A new method for mass percentage determination of fructose and glucose based on FT-Raman spectroscopy is evaluated with a standard HPLC-based method. FT-Raman spectra manipulation is done via the spectrometer software, and a PLS (partial least squares) method is developed with the TQ Analyst software (Ver 1. 1a). The simultaneous quantitative determination uses an input range from 1700 to 700 cm( 1) without correction or baseline factors. The standards used in the PLS method are honey samples previously analyzed by HPLC to obtain their mass percentage concentrations in fructose and glucose. The returned results are statistically tested with those of the HPLC method. Both methods appear to score equally in terms of reproducibility. The honey content of the two sugars in total was found up to 40-74%. The honey samples content in fructose and glucose was determined by HPLC (24.1-42.9% and 16.2-33.1%, respectively) and FT-Raman (24.0-40.8% and 21.1 32.2%, respectively). PMID- 15656652 TI - Ultrasensitive quantitative determination of paraquat: application to river, ground, and drinking water analysis in an agricultural area. AB - The water specimens were collected from wells and irrigation ditches in the agricultural area to the south of Milan and from Olona River and Mantua Lake and analyzed for paraquat detection. The assay was performed using a specific polyclonal antibody raised in sheep and rabbit anti-sheep IgG conjugated with a chelating molecule 4,7-bis(chlorosulfophenyl)-1,10-phenanthroline-2,9 dicarboxylic acid complexed with Eu3+ as a fluorescent marker. Bovine serum albumin conjugated with 5-(1'-methyl[4,4']bipyridinyl-1-yl)pentanoic acid was used in solid phase preparation. The sensitivity achieved was 20 ng L(-1). The recovery in samples spiked with three different PQ concentrations was between 88 and 108%. PMID- 15656651 TI - New PVC nitrate-selective electrode: application to vegetables and mineral waters. AB - The paper refers the analytical potentialities of the oxa-azamacrocycles as potentiometric ionophores for the construction of electrodes selective to nitrate. Afterward, the membrane selective to nitrate is designed and optimized using a [3.3.3.3]oxazane as an ionophore. The membrane was prepared using dibutylphthalate as a solvent mediator, tetraoctylammonium as a lipophilic membrane additive, and poly(vinyl chloride (PVC) as a polymeric matrix, applied directly onto a conductive graphite/epoxy resin support. The electrodes presented a slope of 60 +/- 0.1 mV log(-1), a low limit of linear response of 4.2 x 10(-6) mol L(-1), a useful lifetime of 1 year, and improved selectivity characteristics when compared with other nitrate electrodes. The good working characteristics of this electrode, constructed without inner-reference solution, made possible its application to the determination of nitrate in different types of vegetables and bottled mineral waters without the use of a conditioning solution. The application of a significant F test proved that the results obtained were similar to those attained by application of the brucine spectrophotometric method adopted as a reference technique. Linear regression analysis also showed good agreement between the results obtained by the proposed method and the reference one. PMID- 15656653 TI - Simple and inexpensive method for the reliable determination of additions of soybean proteins in heat-processed meat products: an alternative to the AOAC official method. AB - Despite the existence of an AOAC official method based on an enzyme-linked immunosorbent assay (ELISA) for the determination of additions of soybean proteins in meat products, its use for quantitative assessment is limited. Accordingly, a simple and inexpensive method has been developed and validated in this work. The method involves defatting the meat samples with acetone, solubilization of soybean proteins in a 30 mM Tris-HCl buffer (pH 8) containing 0.5% (v/v) 2-mercaptoethanol, and the identification of two peaks from soybean proteins in the chromatogram obtained by perfusion reversed-phase chromatography and UV detection. Determination of soybean proteins by the proposed method did not suffer from matrix interferences, with a good linear correlation up to a concentration of 12.50 mg/mL soybean proteins being observed. The proposed method was proven to be specific, precise, accurate, robust, and sensitive, making possible the detection and the quantitation of additions of 0.07% (w/w) and 0.25% (w/w), respectively, of soybean proteins in meat products (related to 1 g of initial product). The method has been applied to the determination of the soybean protein content in commercial heat-processed meat products, obtaining results that were statistically similar to those obtained by the official ELISA method but with a higher reliability and simplicity and a lower cost and analysis time. PMID- 15656654 TI - Metabolism of antioxidant and chemopreventive ellagitannins from strawberries, raspberries, walnuts, and oak-aged wine in humans: identification of biomarkers and individual variability. AB - Ellagitannins (ETs) are dietary polyphenols, containing ellagic acid (EA) subunits, with antioxidant and cancer chemopreventive activities that might contribute to health benefits in humans. However, little is known about their metabolic fate. We investigate here the metabolism of different dietary ETs and EA derivatives in humans. Forty healthy volunteers were distributed in four groups. Each group consumed, in a single dose, a different ET-containing foodstuff, i.e., strawberries (250 g), red raspberries (225 g), walnuts (35 g), and oak-aged red wine (300 mL). After the intake, five urine fractions (F) were collected at 8 (F1), 16 (F2), 32 (F3), 40 (F4), and 56 (F5) h. Neither ETs nor EA were detected in urine after LC-MS/MS analysis. However, the microbial metabolite 3,8-dihydroxy-6H-dibenzo[b,d]pyran-6-one (urolithin B) conjugated with glucuronic acid was detected along the fractions F3-F5 in all of the subjects, independently of the consumed foodstuff. The mean percentage of metabolite excretion ranged from 2.8 (strawberries) to 16.6% (walnuts) regarding the ingested ETs. Considerable interindividual differences were noted, identifying "high and low metabolite excreters" in each group, which supported the involvement of the colonic microflora in ET metabolism. These results indicate that urolithin B (a previously described antiangiogenic and hyaluronidase inhibitor compound) is a biomarker of human exposure to dietary ETs and may be useful in intervention studies with ET-containing products. The antioxidant and anticarcinogenic effects of dietary ETs and EA should be considered in the gastrointestinal tract whereas the study of potential systemic activities should be focused on the bioavailable urolithin B derivatives. PMID- 15656655 TI - Identification and antioxidant potential of flavonoids and low molecular weight phenols in olive cultivar chemlali growing in Tunisia. AB - Increasing interest in phenolic compounds in olives is due to their antioxidant and health-enhancing properties. In this study the phenolics in fruits of the Tunisian olive cultivar Chemlali were extracted by methanol-water and fractionated using Sephadex LH-20 column chromatography. The identification of phenolic monomers and flavonoids was based on separation by high-performance liquid chromatography equipped with a diode array detector followed by liquid chromatography-mass spectrometry and gas chromatography-mass spectrometry analysis. Oleuropein, a secoiridoid glycoside esterified with a phenolic acid, was the major compound. Eight phenolic monomers and 12 flavonoids were also identified in Chemlali olives. Five flavonoids were isolated and purified using Sephadex LH-20 column chromatography and preparative paper chromatography. The antioxidant activity of the extract and the purified compounds was evaluated by measuring the radical scavenging effect on 1,1-diphenyl-2-picrylhydrazyl and by using the beta-carotene-linoleate model assay. Acid hydrolysis of the extract enhanced its antioxidant activity. Hydroxytyrosol and quercetin showed antioxidant activities similar to that of 2,6-di-tert-butyl-4-methylphenol. A hydroxyl group at the ortho position at 3' on the B ring of the flavonoid nucleus could contribute to the antioxidant activity of the flavonoids. PMID- 15656656 TI - Germination of peanut kernels to enhance resveratrol biosynthesis and prepare sprouts as a functional vegetable. AB - Bioactive benefits of resveratrol in the diets have attracted extensive interests of the public. Peanut is one of the potent natural sources of resveratrol. In this study, germination of peanut kernels to enhance resveratrol biosynthesis and preparation of sprouts as a functional vegetable was conducted. When the rehydrated kernels of three peanut cultivars were germinated at 25 degrees C and relative humidity 95% in dark for 9 days, resveratrol contents increased significantly from the range of 2.3 to 4.5 microg/g up to the range of 11.7 to 25.7 mug/g depending upon peanut cultivar. In comparison with the sprout components, resveratrol contents were highest in the cotyledons, slightly lower in the roots, and not detected in the stems. When the sprouts were heated in boiling water for 2 min, resveratrol contents varied in a limited range. Methanol extracts of the freeze-dried sprouts exhibited potent 1,1-diphenyl-2-picryl hydrazyl scavenging activity and antioxidative potency against linoleic acid oxidation. These activities increased with an increase of germination time. After 9 days of germination, total free amino acid, sucrose, and glucose contents increased significantly while crude protein contents decreased and the large sodium dodecyl sulfate polyacrylamide gel electrophoresis protein molecules of the kernels were extensively degraded. From a practical viewpoint, it is of potency to prepare peanut sprouts as a functional vegetable. PMID- 15656657 TI - Antifungal and insect antifeedant 2-phenylethanol esters from the liverwort Balantiopsis cancellata from Chile. AB - A chemical study of a dichloromethane extract of Balantiopsis cancellata led to the isolation of four known 2-phenylethanol esters (1-4) and a phenylethanediol benzoate (5). Antifeedant activity toward Spodoptera littoralis (disk-choice bioassay) and growth inhibition of the phytopathogen Cladosporium herbarum in TLC bioautography assays were determined. The results show that the antifeedant and antifungal activity of the extract is attributable mainly to the trans-beta methylthioacrylate 4. PMID- 15656658 TI - Differential exudation of two benzoxazinoids--one of the determining factors for seedling allelopathy of Triticeae species. AB - Benzoxazinoids (Bx) are natural phytotoxins that function as chemical defense compounds in several species. The release of Bx by intact plant roots associated these compounds with root allelopathy in Triticeae species; however, the significance of exudate concentrations of Bx for plant-plant interactions is still a controversial question. A biological screening of 146 cultivars of four Triticeae species (Triticum aestivum L., Triticum durum Desf., Triticum spelta L., and Secale cereale L.) demonstrated a high cultivar dependence to suppress the root growth of Sinapis alba L. by root allelopathy in a dose-response bioassay. Only a few cultivars possessed a marked high or low allelopathic activity, whereby high-performance liquid chromatography-diode array detection analysis of root exudates revealed that these cultivars differed considerably in their ability to exude the two Bx aglucones, DIBOA [2,4-dihydroxy-2H-1,4 benzoxazin-3(4H)-one] and DIMBOA [2,4-dihydroxy-7-methoxy-2H-1,4-benzoxazin-3(4H) one]. The total amount of DIBOA and DIMBOA exuded showed a significant correlation to the growth inhibition in bioassay with a statistically estimated contribution to the overall allelopathic effect of 48-72%. In a bioassay with pure phytotoxins, Bx concentrations consistent with the amounts quantified in the screening bioassay caused detrimental effects on S. alba and almost reproduced the statistically estimated contribution. The observed causal association between the allelopathic activity under laboratory conditions and the exudate concentrations of Bx suggests that this association might have implications for the interference of Triticeae species in natural plant communities. PMID- 15656659 TI - Intracellular mediators of procyanidin-induced lipolysis in 3T3-L1 adipocytes. AB - We have previously reported that grape seed procyanidins stimulate long-term lipolysis on 3T3-L1 fully differentiated adipocytes. To unravel the molecular mechanism by which procyanidins exert this effect, we checked the involvement of two main cellular targets in adipose cells: protein kinase A (PKA) and peroxisome proliferator-activated receptor-gamma (PPAR-gamma). Procyanidin treatment increased intracellular cAMP levels in 3T3-L1 adipocytes, and their lipolytic effect was inhibited by simultaneous treatment with H89, a PKA specific inhibitor. BRL49653, a very highly specific ligand of PPAR-gamma, totally abolished the lipolytic effect of procyanidins. Simultaneous to this long-term lipolytic effect, the mRNA levels of some differentiation adipocyte markers decreased, although there were no changes in the triglyceride content of the cells. BRL49653 did not antagonize the decrements of differentiation markers. These results support a mediation of PPAR-gamma and PKA on the lipolytic effects of procyanidins on 3T3-L1 adipocytes. PMID- 15656660 TI - Four new prenylated isoflavonoids in Tadehagi triquetrum. AB - Investigation on the anthelminthic bioactive compounds of the ethanol extract of Tadehagi triquetrum resulted in the isolation of three new prenylated isoflavones, triquetrumones A (1), B (2), and C (3), and one new prenylated biisoflavanone, (R)-triquetrumone D (4), along with 16 known compounds, cyclokievitone (5), yukovanol (6), aromadendrin (7), kaempferol (8), astragalin (9), 2-O-methyl-l-chiro-inositol (10), galactitol (11), p-hydroxycinnamic acid (12), ursolic acid (13), betulinic acid (14), beta-sitosterol (15), daucosterol (16), stigmasterol (17), stigmasta-5,22-dien-3-O-beta-d-glucopyranoside (18), saccharose (19), and docosanoic acid (20). The structures of 1-4 were elucidated on the basis of spectroscopic and spectrometric methods. Compounds 1-3 displayed mild anthelminthic bioactivity, and compound 3 showed a significant binding ability to the estrogen receptor. PMID- 15656662 TI - Castanea sativa Mill. leaves as new sources of natural antioxidant: an electronic spin resonance study. AB - The antioxidant potential of Castanea sativa Mill. leaf (sweet chestnut) was explored as a new source of active extracts. The capacity of the different fractions issued from aqueous, methanol, and ethyl acetate extracts to inhibit the stable free radical 2,2-diphenyl-1-pycryl-hydrazyl, superoxide anion, and hydroxyl radical was measured by electronic spin resonance. Their scavenging potential was analyzed versus their amount of phenolic compounds. Among the active fractions, the most effective one was A6, an ethyl acetate fraction, which contained a high level of total phenolic compounds (29.1 g/100 g). Thus, a different extraction procedure was performed to concentrate the active compounds of A6 in the new C. sativa leaf extract (CSLE). Compared to reference antioxidants (quercetin and vitamin E) and standard extracts (Pycnogenol, from French Pinus maritima bark, and grape marc extract), it was observed that A6 and CSLE have high antioxidant potentials, equivalent to at least those of reference compounds. PMID- 15656661 TI - Ectopic expression of anthocyanin 5-o-glucosyltransferase in potato tuber causes increased resistance to bacteria. AB - The principal goal of this paper was to investigate the significance of anthocyanin 5-O-glucosyltransferase (5-UGT) for potato tuber metabolism. The ectopic expression of a 5-UGT cDNA in the tuber improved the plant's defense against pathogen infection. The resistance of transgenic lines against Erwinia carotovora subsp. carotovora was about 2-fold higher than for nontransformed plants. In most cases the pathogen resistance was accompanied by a significant increase in tuber yield. To investigate the molecular basis of transgenic potato resistance, metabolic profiling of the plant was performed. In tuber extracts, the anthocyanin 3,5-O-substituted level was significantly increased when compared to that of the control plant. Of six anthocyanin compounds identified, the highest quantity for pelargonidin 3-rutinoside-5-glucoside acylated with p coumaric acid and peonidin 3-rutinoside-5-glucoside acylated with p-coumaric acid was detected. A significant increase in starch and a decrease in sucrose level in transgenic tubers have been detected. The level of all other metabolites (amino acids, organic acids, polyamines, and fatty acids) was quite the same as in nontransformants. The plant resistance to bacterial infection correlates with anthocyanin content and sucrose level. The properties of recombinant glucosyltransferase were analyzed in in vitro experiments. The enzyme kinetics and its biochemical properties were similar to those from other sources. PMID- 15656663 TI - Three new furostanol saponins from the leaves of Lycianthes synanthera ("Chomte"), an edible Mesoamerican plant. AB - Three new furostanol oligoglycosides, 3-O-{alpha-L-rhamnopyranosyl-(1-->2)-[alpha L-rhamnopyranosyl-(1-->4)]-beta-D-glucopyranosyl}-26-O-beta-D-glucopyranosyl 22alpha-methoxy-25R-furost-5-ene-3beta,17alpha,26-triol (1), 3-O-{alpha-L rhamnopyranosyl-(1-->2)-[alpha-L-rhamnopyranosyl-(1-->4)]-beta-D-glucopyranosyl} 26-O-beta-D-glucopyranosylfurost-5-ene-3beta,17alpha,22alpha,25,26-pentol (2), and 3-O-{alpha-L-rhamnopyranosyl-(1-->2)-[alpha-L-rhamnopyranosyl-(1-->4)]-beta-D glucopyranosyl}-26-O-beta-D-glucopyranosylfurost-5-ene-3beta,22alpha,25,26-tetrol (3), named lycianthosides A-C, together with known flavone glycosides were isolated from Lycianthes synanthera leaves, an edible plant of the Solanaceae family that grows naturally in Guatemala. The nutrient composition of the raw leaves was also evaluated. PMID- 15656664 TI - Radical scavenging potential of phenolic compounds encountered in O. europaea products as indicated by calculation of bond dissociation enthalpy and ionization potential values. AB - The radical scavenging potential of phenolic compounds occurring in Olea europaea and of recently identified hydroxytyrosol metabolites was evaluated by means of quantum chemical calculations. The bond dissociation enthalpy (BDE) of phenolic hydroxyl groups and the ionization potential (IP) were calculated as descriptors to predict the H-atom-donating and electron-donating abilities of antioxidants, respectively. Catechol derivatives had the lowest BDE values (77.7-80.1 kcal/mol) whereas the lignans, pinoresinol and 1-acetoxypinoresinol, and other monophenols had much higher BDE values (85.1-88.0 kcal/mol), which suggested a lower potential for radical scavenging. Side chain characteristics were not found to affect the size of BDE values although differences in lipophilicity (on the basis of calculated Log P values) indicate variability in the activity in real systems. Conclusions for the antioxidant potential could not be drawn based on the IP values. Lack of experimental data for most of the studied compounds due to oxidative instability and difficulties in synthesis or isolation supports the usefulness of a computational approach for those interested in the antioxidant potential of phenolics encountered in O. europaea products. PMID- 15656665 TI - The antiproliferative and differentiating effects of human leukemic U937 cells are mediated by cytokines from activated mononuclear cells by dietary mushrooms. AB - Proliferation of human leukemic U937 cells was remarkably inhibited by conditioned medium (CM) of human peripheral blood mononuclear cells (MNC-CM) stimulated with cold-water extracts (CWE) (10-800 microg/mL of medium) of dietary mushrooms, Hypsizigus mamoreus (HM), Agrocybe aegerita (AA), Flammulina velutipes (FV), whereas insignificant results were observed when cells were cultured in the presence of CWE at the corresponding level. Water extracts from mushrooms were fractionated by Sephadex G-50 chromatography, and the pooled high molecular weight fraction (F1) (200 microg/mL) of HM (HM1) and AA (AA1) exhibited growth inhibitions >80% on U937 cells. Interestingly, the thus-cultured U937 cells showed high nitroblue tetrazolium (NBT) positive (>68%) and nonspecific esterase (NSE) positive (>47%) percentages, revealing the remarkable differentiation into monocytes/macrophages upon incubation with HM1- and AA1-stimulated MNC-CM. In addition, assays for the expressions of monocyte-associated antigens, CD11b, CD14, and CD68, also evidenced the remarkable differentiation of U937 cells into monocytes/macrophages by presenting high CD maker positive percentages (>50%). Tumor necrosis factor (TNF)-alpha and interleukin (IL)-1beta in CM of HM1 stimulated MNC for 1 day (MNC-CM-1) were 1350 and 1374 pg/mL, respectively, revealing the potent antitumor and differentiation-inducing activities of HM. Of note, MNC-CM-1 appeared to be more effective than day 5 MNC-CM (MNC-CM-5) in both antitumor and differentiation-inducing activities. PMID- 15656666 TI - Effect of preparation conditions on protein secondary structure and biofilm formation of kafirin. AB - Various extraction and drying conditions for the isolation of kafirin from dry milled, whole grain sorghum have been investigated, with a view to optimizing extraction of the protein for commercial food coatings and packaging films. The addition of sodium hydroxide to an aqueous ethanol extractant increased the yield and solubility of kafirin. Subsequent heat drying at 40 degrees C was shown to cause the kafirin to aggregate as indicated by an increase in intermolecular beta sheets. Extraction of the flour using ethanol (70%, w/w) with 0.5% (w/w) sodium metabisulfite and 0.35% (w/w) sodium hydroxide at 70 degrees C followed by freeze drying of the protein was found to produce a yield of 54% kafirin with good film forming properties. The kafirin films were assessed for their sensory properties, tensile strength, strain, and water vapor permeability. Fourier transform infrared spectroscopy was used to study the secondary structure of the extracted kafirins. The best films were made with kafirin containing a large proportion of nativelike alpha-helical structures with little intermolecular beta-sheet content as indicated by the Fourier transform infrared reflectance peak intensity ratios associated with these secondary structures. The principal factor affecting the secondary structure of the protein appeared to be the temperature at which the protein was dried. Heat drying resulted in a greater proportion of intermolecular beta-sheets. Any industrial-scale extraction must therefore minimize protein aggregation and maximize native alpha-helical structures to achieve optimal film quality. PMID- 15656667 TI - Proteome-level investigation of Brassica carinata-derived resistance to Leptosphaeria maculans. AB - Plants resistant to the fungal pathogen Leptosphaeria maculans were generated by an interspecific cross between the highly susceptible Brassica napus (canola) and the highly resistant Brassica carinata. Changes in the leaf protein profiles of these lines were investigated in order to understand the biochemical basis for the observed resistance. Two-dimensional electrophoresis followed by tandem mass spectrometry led to the identification of proteins unique to the susceptible (5 proteins) and resistant genotypes (7 proteins) as well those that were differentially expressed in the resistant genotype 48 h after challenge with the pathogen (28 proteins). Proteins identified as being unique in the resistant plant material included superoxide dismutase, nitrate reductase, and carbonic anhydrase. Photosynthetic enzymes (fructose bisphosphate aldolase, triose phosphate isomerase, sedoheptulose bisphosphatase), dehydroascorbate reductase, peroxiredoxin, malate dehydrogenase, glutamine synthetase, N-glyceraldehyde-2 phosphotransferase, and peptidyl-prolyl cis-trans isomerase were observed to be elevated in the resistant genotype upon pathogen challenge. Increased levels of the antioxidant enzyme superoxide dismutase were further validated and supported by spectrophotometric and in-gel activity assays. Other proteins identified in this study such as nitrate reductase and peptidylprolyl isomerase have not been previously described in this plant-pathogen system, and their potential involvement in an incompatible interaction is discussed. PMID- 15656668 TI - Relationship of carotenoids and tocopherols in a sample of carrot root-color accessions and carrot germplasm carrying Rp and rp alleles. AB - Carotenoids and tocopherols are powerful antioxidants synthesized in plants from a common precursor. They may offer significant health benefits to humans. Seed oils have been shown to possess high levels of tocopherols, but little is known about their levels in the edible portions of most vegetable crops. A two-year field experiment was conducted at two locations to assess levels of major carotenoids and tocopherols in carrot (Daucus carota) root and leaf tissue. Levels of compounds in root tissue reported on a dry weight basis were as follows: alpha-tocopherol, 0.04-0.18 ppm; lycopene, 0.00-52.94 ppm; alpha carotene, 10.63-1504.76 ppm; and beta-carotene, 26.69-1673.76 ppm. Higher levels of all carotenoids were measured in phloem tissue than in xylem. Leaf tissue levels of tocopherols measured on a dry weight basis ranged from 0.02 to 0.85 ppm, whereas levels of carotenoids ranged from 12.81 to 411.66 ppm. In xylem tissue, alpha-tocopherol was significantly (P < or =0.001) positively correlated with alpha-carotene (r = 0.65) and with beta-carotene (r = 0.52). This positive correlation indicates it may be possible to select for both increased alpha tocopherol and carotenoids in carrot. The reduced pigment (rp) mutation of carrot exhibited a 96% reduction in levels of alpha- and beta-carotene and a 25-43% reduction in alpha-tocopherol when compared to a near-isogenic line. In plants homozygous for rp, a substantial increase was observed in phytoene, a precursor to carotenoids, suggesting the location of the rp lesion in the carotenoid synthesis pathway. PMID- 15656669 TI - Enrichment of yeast thioredoxin by green tea extract through activation of Yap1 transcription factor in Saccharomyces cerevisiae. AB - Thioredoxin (TRX) is an important antioxidant present in all types of organisms. Besides its role as an antioxidant, TRX protects the gastric mucosa due to its antiinflammatory effect. In addition, TRX decreases allergenicity; therefore, the oral administration of TRX is of considerable interest with respect to its clinical use as well as the development of functional foods containing TRX. We have attempted to enrich the cellular TRX content in Saccharomyces cerevisiae, and found that green tea extract (Sunphenon), which is rich in catechins (polyphenols), activates the Yap1 transcription factor, leading to the induction of TRX2, a target of Yap1. Production of yeast TRX was monitored by both a TRX2 lacZ reporter expression assay and Western blotting using an anti-yeast TRX antibody. Maximal production of TRX was achieved in a medium containing 0.1% green tea extract at pH 7.6. We discuss the underlying mechanism by which green tea extract activates Yap1. PMID- 15656670 TI - Analysis of the early stages of lipid oxidation in freeze-stored pork back fat and mechanically recovered poultry meat. AB - An analytical method that can detect low levels of oxidation in food earlier than a sensory panel would be a valuable tool for food manufacturers as well as research institutes. Two model matrixes, pork back fat and mechanically recovered poultry meat (MRPM), were freeze-stored in air at -20 degrees C for 26 weeks. Peroxide value, thiobarbituric acid reactive substances, volatiles analyzed with dynamic headspace gas chromatography-mass spectrometry (GC-MS) and a gas-sensor array technique (electronic nose), chemiluminescence, and front-face fluorescence were evaluated against sensory analysis with regard to detection of early oxidation and correlation with sensory data. Fluorescence and GC-MS could detect oxidative changes in pork back fat earlier than the sensory panel and the electronic nose at the same time. The three methods were highly correlated with sensory attributes (r = 0.8-0.9). GC-MS gave the best results with regard to detection of small oxidative changes in MRPM. PMID- 15656671 TI - Uracil as an index of lactic acid bacteria contamination of tomato products. AB - The aim of this research was to evaluate the suitability of uracil as an hygienic quality index of tomato products. Whereas uridine was naturally present throughout tomato fruits' ripening, uracil appeared only after microbial contamination. In tomato pulp inoculated with nine different microbial strains, all five lactic acid bacteria (LAB) studied released relevant quantities of uracil (150-1040 mg/kg of dm), with a correlated partial or total decrease of uridine. Uracil production by yeasts and molds was very low or nonexistent; the starting uridine concentration (approximately 960 mg/kg of dm) remained constant or increased. Uracil thermostability was also verified. Twenty-six samples of tomato paste (30 degrees Brix) were collected from bag-in-drums produced in an industrial processing plant, some with evident swelling symptoms. All of the samples with high microbial count presented uracil. Uracil was also present in samples with microbial contamination under the detection limit and Howard mold count below legislation limits, implying the reprocessing, at least partial, of altered tomato product. The results indicate that uracil presence in tomato products is an index of LAB contamination that has occurred before heat treatment. PMID- 15656672 TI - Inhibiting effects of resveratrol and its glucoside piceid against Venturia inaequalis, the causal agent of apple scab. AB - Resveratrol is a phytoalexin produced in different unrelated plant species such as grapevine and peanut. The effect of resveratrol and its glucoside trans resveratrol-3-O-beta-D-glycopyranosid (piceid) against Venturia inaequalis, the causal agent of apple scab, was evaluated using a newly established test based on enzymatically isolated cuticular membranes (CMs) from apple (Malus domestica Borkh.) leaves. The test substances resveratrol and piceid were either sorbed to CMs before inoculation with spores or were applied simultaneously with the Venturia inaequalis spores to the CMs, and their effect on germination, appressoria formation, and penetration was examined. Resveratrol had no influence on spore germination but a significant inhibiting effect on penetration when applied simultaneously as well as before. A percentage inhibition of 89.7 +/- 11.5 and 61.8 +/- 35.1 was observed for simultaneous and preapplication, respectively. The resveratrol glucoside piceid had a significant inhibitory effect on germination and completely inhibited penetration of the fungus at concentrations between 200 and 400 microg mL(-1) when applied simultaneously with the spores to the CMs. On piceid-enriched CM (preapplication), spores germinated but penetration was inhibited nearly completely (96.1 +/- 5.1%). Thus, in vitro experiments showed that resveratrol and its glucoside in fact could contribute to improving the pathogen resistance of apple leaves. PMID- 15656673 TI - Available nitrogen and arginine deaminase activity in groundnut (Arachis hypogaea L.) fields after imidacloprid, diazinon, and lindane treatments. AB - Plant available nitrogen and arginine deaminase activities were determined in insecticide-treated groundnut (Arachis hypogaea L.) fields between July and November for three consecutive years (1997-1999). Diazinon was applied for both seed and soil treatments. However, imidacloprid and lindane were used only for the seed treatments. An average half-life (t(1/2)) of diazinon in seed- and soil treated fields was 29.32 and 34.87 days, respectively. Its residues were detected till 60 days in both seed- and soil-treated fields. Diazinon treatments had shown stimulatory effects on available nitrogen in both types of treatments. However, the increase in arginine deaminase activity was only observed in diazinon soil treated field. Residues of imidacloprid and lindane were detected in seed-treated fields till 90 and 120 days with an average half-life (t(1/2)) of 40.96 and 53.39 days, respectively. Imidacloprid had stimulatory effects, and lindane had adverse effects on both available nitrogen and arginine deaminase activities. PMID- 15656674 TI - Perchlorate accumulation in forage and edible vegetation. AB - The accumulation of perchlorate in vegetation is becoming a concern, with increasing numbers of sites reporting the presence of perchlorate in groundwater and surface water. This study investigated potential perchlorate uptake and distribution by a variety of forage and edible crops in both the laboratory and the field. Perchlorate concentrations in soybean leaves grown in the greenhouse were significantly higher than perchlorate concentrations in soybean seeds and pods. Perchlorate concentrations in alfalfa grown in sand were significantly lower than those in alfalfa grown in soil. The concentration of perchlorate in tomato was lower in the fruit than the leaves. Commercially grown wheat and alfalfa samples all contained perchlorate, 0.72-8.6 mg/kg of fresh weight (FW) in the wheat stems, 0.71-4.4 mg/kg of FW in the wheat heads, and 2.9 mg/kg of FW in alfalfa. All field garden samples tested (including cucumber, cantaloupe, and tomato) that were irrigated with perchlorate-tainted water contained perchlorate at various concentrations ranging from 0.040 to 1.65 mg/kg of FW. Bioconcentration factors (BCF), ratios of plant fresh weight concentrations to estimated or measured groundwater concentrations [(microg/kg of FW)/microg/L], were all in the same order of magnitude ranging from 215 +/- 126 for wheat stems to 233 +/- 264 for wheat heads and to 380 +/- 89 for alfalfa. BCF for garden fruit samples were much lower (0.5-20). Results from this study highlight the potential for perchlorate exposure by routes other than drinking water. PMID- 15656675 TI - Spectroscopic and thermal investigations on compost improved by iron salt addition. AB - A quality compost obtained from sewage sludge (one part in weight) and yard trimmings and sawdust (two parts) has been investigated as a potential carrier of iron to plants. At the end of the thermophilic phase, the composting materials were added with crystalline FeSO4.7H2O (97%). Chemical properties, respiratory indices, and seed germination tests proved the compost to be suitable as an iron carrier in agriculture. Fourier transform infrared spectroscopy (FT-IR) and fluorescence spectroscopy provided evidence of the effective linking of the iron ion to the organic molecule functional groups, thus preventing the loss of iron ion by leaching and precipitation phenomena and allowing the metal ion to be available to plants as both mineral and organic species. The thermogram obtained on compost without iron was similar to that previously obtained for composted materials of different origins, whereas samples with added iron ion exhibited in addition an exotherm in the medium-temperature region. FT-IR spectra carried out on samples heated at different temperatures indicated a loss of iron ion linked to carboxyl groups. PMID- 15656676 TI - Determination of the odor threshold concentrations of chlorobrominated anisoles in water. AB - Trihalophenols, which are drinking water disinfection byproducts (DBPs) formed by chlorination or chloramination practices, can be biomethylated into trihalogenated anisoles. These latter compounds have traditionally been suspected of causing odor episodes in drinking water around the world. The odor threshold concentration (OTC) of mixed chlorobrominated anisoles, which were previously synthesized, was determined by flavor profile analysis (FPA) performed by an experienced panel trained to identify odors and tastes in water. The odor threshold amount (OTA) was evaluated by using a gas chromatograph equipped with olfactometry (GC-O) and electron capture detectors (ECD). FPA results for mixed chlorobromoanisoles gave a theoretical OTCs range from 2 to 30 ng/L, the 2,6-diBr 3Cl-anisole being the most odorous compound. Rubber is the general descriptor described by panelists for these compounds, although earthy and musty are the following most cited descriptors. PMID- 15656677 TI - The effect of solvent interactions on alpha-, beta-, and gamma cyclodextrin/flavor molecular inclusion complexes. AB - Three commonly used flavor industry solvents (propylene glycol, triacetin, and triethyl citrate) were tested for their capacity to interfere with the ability of alpha-, beta-, and gamma-cyclodextrin to form molecular inclusion complexes with flavors. Six flavor compounds (ethyl butyrate, ethyl heptanoate, l-menthol, methyl anthranilate, neral, and geranial) were measured by headspace gas chromatography above 2:1 water/ethanol containing appropriate additions of cyclodextrin and flavor solvent. The smallest and most polar solvent molecule represented by propylene glycol had the least effect on cyclodextrin/flavorant complex formation. In contrast, triacetin, intermediate in size among the three flavor diluents studied, had the greatest effect, even though, based on at least some computed molecular parameters, it appears to be more polar than triethyl citrate. The explanation for this apparent anomaly may lie in differences in the extent to which triacetin and triethyl citrate are able to interact with cyclodextrins by means of partial interaction with the hydrophobic cavities of the latter. PMID- 15656678 TI - Identification and aroma impact of norisoprenoids in orange juice. AB - Four norisoprenoids, alpha-ionone, beta-ionone, beta-cyclocitral, and beta damascenone, along with their putative carotenoid precursors, were identified in Valencia orange juice using time-intensity GC-O, GC-MS, and photodiode array HPLC. alpha-Ionone and beta-cyclocitral are reported in orange juice for the first time. GC-O aroma peaks were categorized into seven groups with similar sensory qualities: citrus/minty, metallic/mushroom/geranium, roasted/cooked/meaty/spice, fatty/soapy/green, sulfury/solventy/medicine, floral, and sweet fruity. The four norisoprenoids contributed approximately 8% of the total aroma intensity and 78% of the total floral aroma category. The putative carotenoid norisoprenoid precursors, alpha- and beta-carotene, alpha- and beta cryptoxanthin, and neoxanthin, were identified in the same orange juice using photodiode array HPLC retention times and spectral characteristics. PMID- 15656679 TI - Inhibition of key aroma compound generated during ultrahigh-temperature processing of bovine milk via epicatechin addition. AB - The ability of epicatechin (EC) to inhibit the thermal development of aroma compounds (i.e., Maillard reaction products) formed during ultrahigh-temperature (UHT) processing of bovine milk was evaluated. Volatile extracts were prepared for two UHT-processed milk samples made from (1) raw milk and (2) raw milk containing 0.1% EC by solvent-assisted flavor evaporation (SAFE) and subsequently analyzed by aroma extract dilution analysis (AEDA). Sensory evaluation was also conducted by a trained panel on the intensity of cooked flavor and bitterness in four UHT-processed milk samples (0.00, 0.01, 0.10, and 0.20% EC added prior to processing), as well as a commercial pasteurized milk sample for comparison. AEDA indicated that addition of EC to raw fluid milk prior to UHT processing reduced the overall thermal formation of key aroma-active compounds in comparison to the traditional UHT milk sample. The largest changes in FD values were reported for methional, furfural, 2-isopropyl-3-methoxypyrazine, 2-acetyl-1-pyrroline, and 2 acetyl-2-thiazoline (Maillard-type aroma compounds) with 32-, 8-, 8-, 4-, and 4 fold reductions in formation, respectively. Sensory evaluation also revealed that all EC-containing UHT milk samples had statistically (P < 0.05) lower cooked flavor intensity in comparison to the control, whereas the 0.2% EC sample was statistically similar to a pasteurized milk sample. Furthermore, addition of EC at or below 0.1% in UHT fluid milk did not significantly increase the bitterness intensity. PMID- 15656680 TI - Aroma release and retronasal perception during and after consumption of flavored whey protein gels with different textures. 1. in vivo release analysis. AB - The influence of gel texture on retronasal aroma release during mastication was followed by means of real-time proton-transfer reaction mass spectrometry and compared to sensory perception of overall aroma intensity. A clear correlation was found between individual-specific consumption patterns and the respective physicochemical release patterns in vivo. A modified data analysis approach was used to monitor the aroma changes during the mastication process. It was found that the temporal resolution of the release profile played an important role in adequate description of the release processes. On the basis of this observation, a hypothesis is presented for the observed differences in intensity rating. PMID- 15656681 TI - Use of chemical profiling to differentiate geographic growing origin of raw pistachios. AB - The objective of this study was to demonstrate the feasibility of chemical profiling methods combined with multivariate methods to differentiate the geographical growing regions of pistachios (Pistachia vera). Elemental analysis (Ba, Be, Ca, Cu, Cr, K, Mg, Mn, Na, V, Fe, Co, Ni, Cu, Zn, Sr, Ti, Cd, and P) of pistachios samples was performed using inductively coupled plasma atomic emission spectrometry. Analysis of inorganic anions and organic acids (selenite, bromate, fumarate, malate, selenate, pyruvate, acetate, phosphate, and ascorbate) of pistachio samples was performed using capillary electrophoresis. Bulk carbon and nitrogen isotope ratios were performed using stable isotope MS. There were nearly 400 pistachio samples analyzed from the three major pistachio growing regions: Turkey, Iran, and California (United States). A computational evaluation of the trace element data sets was carried out using statistical pattern recognition methods including principal component analysis, canonical discriminant analysis, discriminant analysis, and neural network modeling. Several linear discriminant function models classified the data sets with 95% or higher accuracy. We report the development of a method combining elemental analysis and classification techniques that may be widely applied to the determination of the geographical origin of foods. PMID- 15656682 TI - Beta-lactoglobulin-dextran conjugates: effect of polysaccharide size on emulsion stability. AB - A series of dextrans and beta-lactoglobulin were covalently conjugated and screened for their ability to stabilize oil-in-water emulsions. Dextrans with the molecular mass of 19.6 kDa, 87 kDa, 150 kDa, 500 kDa, and 2000 kDa were attached to beta-lactoglobulin via the Maillard reaction. The conjugates were then purified and evaluated as emulsifiers under neutral conditions. The ability to stabilize emulsions was determined by monitoring oil droplet size over time. Adsorption of the conjugates to the droplet surface was characterized by determining the protein surface load. The results show that increasing polysaccharide size increases emulsion stability up to 150 kDa before leveling off. Conversely, surface protein density remains constant until 150 kDa before decreasing with polysaccharide size. A model is presented to interpret the results. PMID- 15656683 TI - Hop as an interesting source of resveratrol for brewers: optimization of the extraction and quantitative study by liquid chromatography/atmospheric pressure chemical ionization tandem mass spectrometry. AB - Nowadays, hop is used almost exclusively by brewers for bitterness and flavor. Although hop polyphenols have been widely studied in the past decade for their antioxidant activity in the boiling kettle, very little is known about their real impact on health. The discovery of resveratrol in hop pellets highlights the potential health-promoting effect of moderate beer consumption. Here, we have optimized a quantitative extraction procedure for resveratrol in hop pellets. Preliminary removal of hydrophobic bitter compounds with toluene and cyclohexane at room temperature allows 99% trans-resveratrol recovery by ethanol:water (75:25, v/v) solid/liquid extraction at 60 degrees C. Reverse phase liquid chromatography proves an excellent means of separating isomers. In addition, we have compared two mass spectrometry ionization methods-atmospheric pressure chemical ionization (APCI) and electrospray ionization (ESI)-in both the positive and the negative modes. On the basis of standard additions applied with the optimized extraction procedure and reverse phase high-performance liquid chromatography-APCI(+)-tandem mass spectrometry, it appears that Tomahawk hop pellets (T90, harvest 2002) contain 0.5 ppm trans-resveratrol, 2 ppm trans piceid, no cis-resveratrol, and 0.9 ppm cis-piceid. PMID- 15656684 TI - Anti-inflammatory potential of flavonoid contents from dried fruit of Crataegus pinnatifida in vitro and in vivo. AB - The dried fruits of Crataegus pinnatifida, a local soft drink material and medical herb, demonstrated antioxidant effect in a previous study. The present study investigates the anti-inflammatory potential of flavonoid contents from dried fruit of C. pinnatifida (CF-Fs). The preliminary investigation showed that CF-Fs (0.25-0.75 mg/mL) decreased the release of PGE2 and nitric oxide as induced by lipopolysaccharide (LPS, an endotoxin) in macrophage RAW 264.7 cells. The in vivo assay showed that pretreatment of rats with CF-Fs (50-200 mg/kg dosed by gavage) for 5 days significantly decreased the serum levels of the hepatic enzyme markers alanine aminotransferase and aspartate aminotransferase induced by the 6 h treatment with LPS (i.p.; 5 mg/kg). Histopathological evaluation of the rat livers revealed that CF-Fs reduced the incidence of liver lesions such as neutrophil infiltration and necrosis induced by LPS. Furthermore, it was found that pretreatment with CF-Fs decreased the hepatic expression of iNOS and COX-2 induced by LPS in rats. These results demonstrate that CF-Fs present anti inflammatory potential in vitro and in vivo and that they may play a role in hepatoprotection. PMID- 15656685 TI - Linolenic acid-modified chitosan for formation of self-assembled nanoparticles. AB - Chitosan was modified by coupling with linolenic acid through the 1-ethyl-3-(3 dimethylaminopropyyl)carbodiimide-mediated reaction. The degree of substitution was measured by 1H NMR, and it was 1.8%, i.e., 1.8 linolenic acids group per 100 anhydroglucose units. The critical aggregation concentration (CAC) of the self aggregate of hydrophobically modified chitosan was determined by measuring the fluorescence intensity of the pyrene as a fluorescent probe. The CAC value in phosphate-buffered saline (PBS) solution (pH 7.4) was 5 x 10(-2) mg/mL. The average particle size of self-aggregates of hydrophobically modified chitosan in PBS solution (pH 7.4) was 210.8 nm with a unimodal size distribution ranging from 100 to 500 nm. A transmission electron microscopy study showed that the formation of near spherical shape nanoparticles had enough structural integrity. The loading ability of hydrophobically modified chitosan (LA-chitosan) was investigated by using bovine serum albumin (BSA) as a model protein. Self aggregated nanoparticles exhibited an increased loading capacity (19.85 +/- 0.04 to 37.57 +/- 0.25%) with an increasing concentration of BSA (0.1-0.5 mg/mL). PMID- 15656686 TI - Color, betalain pattern, and antioxidant properties of cactus pear (Opuntia spp.) clones. AB - Total phenolics, ascorbic acid, and betalain contents of differently colored cactus pear clones (nine Opuntia ficus-indica [L.] Mill. clones and one O. robusta Wendl. clone) were investigated and related to their respective antioxidant potential assessed by Trolox-equivalent antioxidant capacity (TEAC) and oxygen radical absorbance capacity (ORAC) assays. TEAC and ORAC values were very highly correlated with each other and also with values for total phenolics, betalain contents, and ascorbic acid concentrations. Total phenolics had the greatest contribution to ORAC and TEAC values. High-performance liquid chromatography (HPLC)-diode array detector (DAD)-tandem mass spectrometry (MS/MS) measurements of cactus pear juices permitted the differentiation of the clones based on variations in pigment patterns and betalain concentrations. The red and yellow betalains were absent in lime green colored cactus fruits. The ratio and concentration of these pigments were responsible for the yellow, orange, red, and purple colors in the other clones. Progeny of purple and lime green colored parents were characterized by 12% and 88% of plants bearing lime green and purple fruit, respectively. This implies that the genes for betalain production were lacking in the lime green fruits but could be provided by a parent with a complete set of genes, that is, purple fruits. Besides known pigments typical of Cactaceae, two unexpected betalains were identified. Whereas gomphrenin I was found for the first time in tissues of cactus plants, methionine-betaxanthin has never been described before as a genuine betalain. In addition to their alleged health-promoting properties, various combinations of yellow betaxanthins and red purple betacyanins may allow the development of new food products without using artificial colorants. PMID- 15656687 TI - Proteolytic activity in byproducts from cod species caught at three different fishing grounds. AB - Proteolytic activity in byproducts from cod species caught at three different fishing grounds has been characterized and compared. The overall highest activity in the byproduct fractions was found in viscera at pH 3 (35 degrees C). Cut off and liver fractions also show maximum activity at pH 3, 35 and 50 degrees C, respectively. Proteolytic activity in viscera and cut off fractions are more affected by fishing ground than by species. Proteolytic activity at pH 3 in viscera is higher in samples from the south coast of Ireland, while viscera samples from the Icelandic Sea have the highest activity at pH 7. For cut off, the activity is higher in samples from the south coast of Ireland than the other two fishing grounds. PMID- 15656688 TI - Aggregation of casein micelles by interactions with chitosans: a study by Monte Carlo simulations. AB - Recently, it was found that the addition of chitosan, a cationic polymer, to whole or skim milk produces the destabilization and coagulation of casein micelles which takes place without modifications in the milk pH or in the stability of most of the whey proteins. In the present work, Monte Carlo simulations are employed to show that the phase separation of casein micelles induced by chitosan can be explained by a depletion mechanism, where an effective attraction between the casein micelles is induced by the presence of chitosan molecules. This interaction is described on the basis of Vrij's model, where the depletion of polymer from the gap between neighboring casein micelles originates an effective attractive interaction that leads to a phase transition. This model, that considers volume restriction effects, accounts for several qualitative and even quantitative aspects of the experimental data for the coagulation of casein through chitosan addition. PMID- 15656689 TI - Antioxidant properties of casein calcium peptides and their effects on lipid oxidation in beef homogenates. AB - The antioxidant activity of casein calcium peptides in several in vitro assay systems was investigated. Casein calcium peptides were prepared by the microbial enzymic hydrolysis of casein calcium. The main peak of the molecular mass distribution of the peptides was about 3 kDa. Casein calcium peptides showed strong antioxidant activity with the beta-carotene bleaching method, and they also showed scavenging activity against radicals such as superoxide radicals, 1,1 diphenyl-2-picrylhydrazyl (DPPH) radicals, and hydroxyl radicals. Antioxidant activity was increased with an increasing peptide concentration. Casein calcium peptides also showed strong antioxidant activity against lipid oxidation in ground beef homogenates. These results suggest that casein calcium peptides are a suitable natural antioxidant that prevents the lipid oxidation of meat and related food ingredients. PMID- 15656690 TI - Characterization and functional study of Antrodia camphorata lipopolysaccharide. AB - Lipopolysaccharide (LPS) is a highly proinflammatory molecule isolated from bacteria. This study demonstrated the existence of LPS in a medicinal fungus, Antrodia camphorata. Because no LPS had been identified in any fungus organism, the purification of LPS from A. camphorata was attempted. LPSs from six strains of A. camphorata (35396, 35398, 35716, B71, B85, and B86) were isolated. Chemical and functional properties were investigated on the fungus LPS. Compositional analysis revealed that sorbitol, fucose, galactose, and glucose were the neutral sugars in LPS of A. camphorata. Galactosamine, glucosamine, galactose, and glucose were the predominant monosaccharide species in E. coli O129 LPS molecules, whereas galactosamine and glucosamine were absent in A. camphorata LPS. Because these properties are different from those of bacterial LPS, the functions between fungus and bacterial LPS are also discussed. The vascular endothelial lining of blood vessels, which controls leucocyte traffic and activation, may be one of the primary targets of LPS action during sepsis. Assays for biological activity were performed on endothelial cells with anti inflammatory effects associated with sepsis. A. camphorata LPS apparently showed a lesser extent of cytotoxicity than bacterial LPS. In contrary to the proinflammatory property of bacterial LPS, LPS from A. camphorata differentially reversed bacterial LPS-induced intercellular adhersion molecule-1 and monocyte adhesion; both were indicators during inflammatory process. In conclusion, basic chemical properties categorized A. camphorata extracts into lipopolysaccharide. However, the detailed functional structures and bioactivities of A. camphorata LPS were totally different from those of bacterial LPS. The investigation of the existence and anti-inflammatory effect of fungus LPS is at present a truly novel and important finding. These results show that LPS isolated from A. camphorata offers a novel therapeutic target for anti-inflammation against E. coli infection. PMID- 15656691 TI - Determination of anthocyanins in Ruscus aculeatus L. berries. AB - Red berries of Ruscus aculeatus L., a wild shrub typical of Mediterranean Europe and Africa, were investigated for the first time in order to determine the profile of anthocyanins. The pigments were extracted from the skins of the berries with 0.1% HCl in methanol, purified using a C-18 solid phase cartridge, and identified by means of high-performance liquid chromatography (HPLC)-diode array detection-mass spectrometry analysis. Information from HPLC profiles, saponification, and acid hydrolysis of the anthocyanins showed that the major anthocyanins were pelargonidin 3-O-rutinoside (64%), pelargonidin 3-O-glucoside (16%), and pelargonidin 3-O-trans-p-coumarylglucoside (13%). The attractive color of R. aculeatus berries and the great abundance of the plant in the south of Italy make these berries a new and promising source of natural colorants. PMID- 15656692 TI - Comparative studies on the hypolipidemic and growth suppressive effects of oolong, black, pu-erh, and green tea leaves in rats. AB - The four major commercial teas, oolong, black, pu-erh, and green teas, have been manufactured in southeast Asia. In this study, we evaluated the growth suppressive and hypolipidemic effect of these four different tea leaves by oral feeding to male Sprague-Dawley rats for 30 weeks. The results showed that the suppression of body weights of tea leaves-fed groups were in the order: oolong tea > pu-erh tea > black tea > green tea. Pu-erh tea and oolong tea could lower the levels of triglyceride more significantly than that of green tea and black tea, but pu-erh tea and green tea were more efficient than oolong tea and black tea in lowering the level of total cholesterol. In lipoprotein, 4% pu-erh tea could increase the level of HDL-C and decrease the level of LDL-C, but other teas simply decrease the levels of both. The activity of antioxidant enzyme SOD is increased in all tea-fed groups as compared to the basal diet-fed group. Finally, relative weight ratios of liver to epididylmal adipose tissue were lower in feeding oolong tea and pu-erh tea groups. On the basis of these findings, it seemed that the fully fermented pu-erh and black tea leaves and partially fermented oolong tea leaves were more effective on their growth suppressive and hypolipidemic effects as compared to the nonfermented green tea leaves. PMID- 15656693 TI - Susceptibility to hypoxia/reoxygenation of aged rat cardiomyocytes and its modulation by selenium supplementation. AB - Since in the aged heart an increased basal production of reactive oxygen species (ROS) has been demonstrated, and the resistance to ROS attack could be ameliorated by antioxidant supplementation, we verified the protective effect of selenium, as sodium selenite (SS) or seleno methionine (SM), in cultured rat cardiomyocytes aged in vitro. In normoxia, glutathione peroxidase (GPx) activity and total antioxidant activity were higher in old than in young cardiomyocytes, suggesting the existence of a compensatory increase of antioxidant defenses. When aged cells were submitted to hypoxia/reoxygenation, GPx activity was not modified; while total antioxidant activity decreased, conjugated diene level increased. Selenium supplementation, particularly as SM, was able to increase GPx, and consequently total antioxidant activity, and to decrease conjugated diene production. The observed ability of selenium supplementation to protect aged cardiomyocytes from hypoxia/reoxygenation damage underlines the importance of an optimal selenium dietary intake, particularly in the elderly. PMID- 15656696 TI - Pharmacokinetics, pharmacodynamics and drug interaction potential of enfuvirtide. AB - Enfuvirtide, the first fusion inhibitor approved for the treatment of HIV-1 infection, is a synthetic peptide that binds to HIV-1 glycoprotein 41, blocking the fusion of viral and cellular membranes. When administered subcutaneously at the recommended dose of 90 mg twice daily with optimised background antiretroviral therapy, enfuvirtide significantly reduces plasma HIV-1 RNA levels up to 48 weeks compared with optimised background therapy alone. Enfuvirtide exhibits a small volume of distribution (5.48 L), low systemic clearance (1.4 L/h) and high plasma protein binding (92%). Less than 17% of enfuvirtide is converted to a minimally active deaminated form of the parent drug. Both enfuvirtide and its metabolite are primarily eliminated via catabolism to amino acid residues. Following subcutaneous administration, enfuvirtide is almost completely absorbed, and exposure increases almost linearly with dose over the range 45-180 mg. When administered at the recommended dose in adults, subcutaneous absorption is slow and protracted, resulting in relatively flat steady-state plasma concentration-time profiles. Bioavailability is high (84.3%) and the elimination half-life (3.8 hours) supports twice-daily administration. Comparable absorption was observed from three different anatomical injection sites. The pharmacokinetic-pharmacodynamic relationship indicates that the recommended dose, in combination with other active antiretrovirals, is optimal. Enfuvirtide clearance is influenced to a small extent by sex and bodyweight but this does not necessitate dosage adjustment. In vitro and in vivo studies indicate that enfuvirtide has a low potential to interact with concomitantly administered drugs. Enfuvirtide did not influence concentrations of drugs metabolised by cytochrome P450 (CYP) 3A4, CYP2D6 or N-acetyltransferase, and had only minimal effects on those metabolised by CYP1A2, CYP2E1 or CYP2C19. Coadministration of ritonavir, ritonavir-boosted saquinavir or rifampicin (rifampin) did not result in clinically significant changes in enfuvirtide pharmacokinetics. In HIV-1-infected paediatric patients, subcutaneous dosages based on bodyweight (2 mg/kg twice daily) produce pharmacokinetics broadly similar to those observed in adults administered 90 mg twice daily. PMID- 15656695 TI - Individualised cancer chemotherapy: strategies and performance of prospective studies on therapeutic drug monitoring with dose adaptation: a review. AB - Therapeutic drug monitoring (TDM) is increasingly used in clinical practice for the optimisation of drug treatment. Although pharmacokinetic variability is an established factor involved in the variation of therapeutic outcome of many chemotherapeutic agents, the use of TDM in the field of oncology has been limited thus far. An important reason for this is that a therapeutic index for most anticancer agents has not been established; however, in the last 20 years, relationships between plasma drug concentrations and clinical outcome have been defined for various chemotherapeutic agents. Several attempts have been made to use these relationships for optimising the administration of chemotherapeutics by applying pharmacokinetically guided dosage. These prospective studies, individualising chemotherapy dose during therapy based on measured drug concentrations, are discussed in this review. We focus on the way a target value is defined, the methodologies used for dose adaptation and the way the performance of the dose-adaptation approach is evaluated. Furthermore, attention is paid to the results of the studies and the applicability of the strategies in clinical practice. It can be concluded that TDM may contribute to improving cancer chemotherapy in terms of patient outcome and survival and should therefore be further investigated. PMID- 15656697 TI - The hepatic sinusoid in aging and cirrhosis: effects on hepatic substrate disposition and drug clearance. AB - The fenestrated sinusoidal endothelium ('liver sieve') and space of Disse in the healthy liver do not impede the transfer of most substrates, including drugs and oxygen, from the sinusoidal lumen to the hepatocyte. Plasma components transfer freely in both directions through the endothelial fenestrations and into the space of Disse. The endothelium is attenuated, there is no basement membrane and there is minimum collagen in the space of Disse, thus minimising any barriers to substrate diffusion. Both cirrhosis and aging are associated with marked structural changes in the sinusoidal endothelium and space of Disse that are likely to influence bulk plasma transfer into the space of Disse, and diffusion through the endothelium and space of Disse. These changes, termed capillarisation and pseudocapillarisation in cirrhosis and aging, respectively, impede the transfer of various substrates. Capillarisation is associated with exclusion of albumin, protein-bound drugs and macromolecules from the space of Disse, and the progressive transformation of flow-limited to barrier-limited distribution of some substrates. There is evidence that the sinusoidal changes in cirrhosis and aging contribute to hepatocyte hypoxia, thus providing a mechanism for the apparent differential reduction of oxygen-dependent phase I metabolic pathways in these conditions. Structural change and subsequent dysfunction of the liver sieve warrant consideration as a significant factor in the impairment of overall substrate handling and hepatic drug metabolism in cirrhosis and aging. PMID- 15656694 TI - Interactions between antiretrovirals and antineoplastic drug therapy. AB - Despite the established impact of highly active antiretroviral therapy (HAART) in reducing HIV-related morbidity and mortality, malignancy remains an important cause of death. Patients who receive the combination of cancer chemotherapy and HAART may achieve better response rates and higher rates of survival than patients who receive antineoplastic therapy alone. However, the likelihood of drug interactions with combined therapy is high, since protease inhibitors (PIs) and non-nucleoside reverse transcriptase inhibitors (NNRTIs) are substrates and potent inhibitors or inducers of the cytochrome P450 (CYP) system. Since many antineoplastic drugs are also metabolised by the CYP system, coadministration with HAART could result in either drug accumulation and possible toxicity, or decreased efficacy of one or both classes of drugs. Although formal, prospective pharmacokinetic interaction studies are not available in most instances, it is possible to infer the nature of drug interactions based on the metabolic fates of these agents. Paclitaxel and docetaxel are both metabolised by the CYP system, although differences exist in the nature of the isoenzymes involved. Case reports describing adverse consequences of concomitant taxane-antiretroviral therapy exist. Although other confounding factors may have been present, these cases serve as reminders of the vigilant monitoring necessary when taxanes and HAART are coadministered. Similarly, vinca alkaloids are substrates of CYP3A4 and are, thus, vulnerable to PI- or NNRTI-mediated changes in their pharmacokinetics. Interactions with the alkylating agents cyclophosphamide and ifosfamide are complicated as a result of the involvement of the CYP3A4 and CYP2B6 isoenzymes in both the metabolic activation of these drugs and the generation of potentially neurotoxic metabolites. Existing data regarding the metabolic fate of the anthracyclines doxorubicin and daunorubicin suggest that clinically detrimental interactions would not be expected with coadministered HAART. Commonly used endocrine therapies are largely substrates of the CYP system and may, therefore, be amenable to modulation by concomitant HAART. In addition, tamoxifen itself has been associated with reduced concentrations of both anastrozole and letrozole, raising the concern that similar inducing properties may adversely affect the outcome of PI- or NNRTI-based therapy. Similarly, dexamethasone is both a substrate and concentration-dependent inducer of CYP3A4; enhanced corticosteroid pharmacodynamics may result with CYP3A4 inhibitors, while the efficacy of concomitant HAART may be compromised with prolonged dexamethasone coadministration. Since PIs and NNRTIs may also induce or inhibit the expression of P-glycoprotein, the potential for additional interactions to arise via modulation of this transporter also exists. Further research delineating the combined safety and pharmacokinetics of antiretrovirals and antineoplastic therapy is necessary. PMID- 15656698 TI - Pharmacokinetic/pharmacodynamic modelling of antibacterials in vitro and in vivo using bacterial growth and kill kinetics: the minimum inhibitory concentration versus stationary concentration. AB - BACKGROUND: The minimum inhibitory concentration (MIC) is the in vitro reference value to describe the activity of an antibacterial against micro-organisms. It does not represent the dynamic effect of the antimicrobial at any point in time, but rather the total antimicrobial effect over the incubation period at a fixed concentration. OBJECTIVE: To explore the concentration-effect relationship of antimicrobial concentrations against micro-organisms in relation to the MIC. METHODS: Time-kill curves were generated for ceftazidime, meropenem and tobramycin against Pseudomonas aeruginosa. The Hill equation with variable slope was fit to the time-kill data, and mathematical models of growth and kill were explored with reference to the MIC. RESULTS: With declining concentrations, bacterial killing will decrease until a specific threshold concentration is reached. This concentration, at which bacteria are neither killed nor able to grow, is named the stationary concentration (SC) and is not equal to the MIC. Pharmacokinetic/pharmacodynamic simulations over a range of kill rates, growth rates and slope factors showed that for beta-lactam antibacterials, the SC is close to the MIC value, which may explain why concentrations in vivo need to be above the MIC, while regrowth of bacteria occurs when concentrations decline below the MIC. For concentration-dependent antibacterials, such as aminoglycosides and quinolones, the SC is shown to be markedly different from the MIC and, in general, is much lower. CONCLUSION: The MIC is not a good pharmacodynamic parameter to characterise the concentration effect relationship of a given antimicrobial. For 'concentration independent' antimicrobials the SC is likely to be close to the MIC, but may be much lower for 'concentration dependent' antimicrobials, and may explain sub-MIC effects. PMID- 15656699 TI - Oral bioavailability of posaconazole in fasted healthy subjects: comparison between three regimens and basis for clinical dosage recommendations. AB - BACKGROUND AND OBJECTIVE: Posaconazole is a potent, extended-spectrum triazole antifungal agent currently in clinical development for the treatment of invasive fungal infections. This study was conducted to compare the bioavailability and resulting serum concentrations of posaconazole 800 mg following administration of three different dose regimens to fasting adults. STUDY DESIGN: This was a randomised, open-label, three-way crossover study. METHODS: Subjects fasted 12 hours before and 48 hours after the administration of posaconazole oral suspension (800 mg) given as a single dose (regimen A), 400 mg every 12 hours (regimen B) or 200 mg every 6 hours (regimen C). Plasma posaconazole concentrations were determined for 48 hours after the initial dose and subjects completed a 1-week washout period between treatment regimens. A one-compartment oral model with first-order rate of absorption and first-order rate of elimination was fitted to the plasma concentration-time data. Differences in exposure were investigated by allowing the bioavailability fraction to vary among regimens. STUDY PARTICIPANTS: A total of 18 healthy men were enrolled in and completed the study. MAIN OUTCOME MEASURES AND RESULTS: Posaconazole relative bioavailability was estimated to be significantly different among regimens (p < 0.0001) and increased with the number of doses, such that regimen B/regimen A = 1.98 +/- 0.35, representing a 98% increase, and regimen C/regimen A = 3.20 +/- 0.69, or a 220% increase. With use of the one-compartment model, the population steady-state values for area under the concentration-time curve over 24 hours were predicted to be 3900, 7700 and 12 400 microg.h/L, with average plasma concentrations of 162, 320 and 517 microg/L for regimens A, B and C, respectively. CONCLUSION: These data suggest that divided daily dose administration (every 12 or 6 hours) significantly increases posaconazole exposure under fasted conditions. PMID- 15656700 TI - Reactions of excited states of phenoxazin-3-one dyes with amino acids. AB - The interaction with amino acids of the excited states of the N-oxide resazurin and its deoxygenation product resorufin, has been studied in aqueous solution at pH 7.5. Steady-state and time-resolved studies show that the fluorescence is quenched by amino acids. Complexation of the dyes in the ground state with aromatic amino acids was also observed. The singlet quenching is attributed to electron transfer from the amino acids to the excited dye based on the dependence of the bimolecular rate constants with the ionization potential of quenchers. Flash photolysis experiments allowed determination of the quenching rate constants for the triplet deactivation of dyes by several amino acids, as well as the characterization of the transients formed in the process. These data show that the triplet is also deactivated by an electron transfer process. However, the deactivation of the N-oxide dye by tryptophan can be described by a hydrogen atom transfer. The protolytic dissociation constants of the dye radical ions are reported. The irradiation of rezasurin in the presence of amino acids leads to deoxygenation of the dye to give resorufin. This process involves the triplet excited state of resazurin and is efficient only in the presence of amino acids containing the -SH group. PMID- 15656701 TI - Update on the positive effects of light in humans. AB - The adverse effects of sunlight, from melanoma to cataracts, are well known and frequently reported (1). However, because humans evolved under sunlight, it is not surprising that there are many positive effects of light on human health. Light that reaches the human eye has two fundamental biological functions: regulation of the visual cycle and of circadian rhythm. We report here the most recent developments in both of these areas. PMID- 15656703 TI - Ambulatory Pediatric Association policy statement: Ensuring integrity for research with children. PMID- 15656704 TI - Creating an analytic voice in the policy storm. PMID- 15656705 TI - Responsible research involving children. PMID- 15656706 TI - Health literacy among Spanish-speaking Latino parents with limited English proficiency. AB - BACKGROUND: Health literacy measures the degree to which individuals understand health information. It has not been studied among parents with limited English proficiency (LEP). OBJECTIVE: We aimed to determine how well Spanish-speaking Latino parents with LEP understood the written instructions accompanying a routinely prescribed medication. DESIGN AND METHODS: We conducted a cross sectional survey of parents of young children. We showed subjects a medicine bottle with an English prescription label and a Spanish drug information sheet (DIS). Subjects demonstrated how much medicine they would give and stated how often they would give it (Medication Dosing). Then they answered 5 questions regarding information from the DIS (DIS comprehension). We coded responses dichotomously as correct or incorrect. We compared Medication Dosing and DIS comprehension by age, comfort with speaking English, birthplace, number of years in the United States, and education. Regression analyses were performed to adjust for these potential confounders. RESULTS: Of 100 participants, 22% correctly dosed the medication; 29% correctly answered all questions regarding the Spanish DIS. Of subjects comfortable speaking English, 50% correctly demonstrated the amount of medicine to give. Overall, higher education and comfort speaking English were associated with better Medication Dosing. Higher education and birth in South America were associated with better DIS comprehension. CONCLUSIONS: Few parents with LEP were able to understand routinely dispensed written medication instructions. Pediatricians should not assume that Spanish-speaking Latino parents who are comfortable speaking English will understand a prescription label written in English, or that Latino parents who speak Spanish will understand drug information written in Spanish. PMID- 15656708 TI - Delayed or forgone care among children with special health care needs: an analysis of the 2001 National Survey of Children with Special Health Care Needs. AB - OBJECTIVE: To examine the associations of sociodemographic characteristics with both the prevalence and the causes of delayed or forgone care in a nationally representative sample of children with special health care needs. METHODS: Data were abstracted from the 2001 National Survey of Children with Special Health Care Needs. The families of children with special health care needs (CSHCN) who reported delayed or forgone care were asked about the reasons. The 12 reasons in the questionnaire were grouped into 5 categories. Bivariate and multivariate logistic regression analyses were conducted in SUDAAN to examine the relationship between sociodemographic characteristics of CSHCN and the incidence of delayed or forgone care by its reasons. RESULTS: Nearly 10% of CSHCN had experienced delayed or forgone health care in the past 12 months in 2001. Logistic regression showed that delayed or forgone care was more likely to be reported by the families of CSHCN who were adolescents, who had more severe limitations, lived in the South or West, lacked medical insurance, and who lived in families under or near the federal poverty line. Hispanics were more likely to report "lack of medical specialty" and "had language, communication, or cultural problems with provider." Both Hispanics and non-Hispanic others were twice as likely to report "provider not accessible" as reasons for the delayed or forgone care compared with non Hispanic whites or blacks. conclusion: CSHCN with certain socioeconomic status and sociodemographic characteristics, as well as those with severe limitations in activity, were more likely to be affected by circumstances that result in delayed or forgone care. PMID- 15656707 TI - Health care for children and youth in the United States: annual report on patterns of coverage, utilization, quality, and expenditures by income. AB - OBJECTIVES: To examine differences by income in insurance coverage, health care utilization, expenditures, and quality of care for children in the United States. METHODS: Two national health care databases serve as the sources of data for this report: the 2000-2002 Medical Expenditure Panel Survey (MEPS) and the 2001 Nationwide Inpatient Sample (NIS) from the Healthcare Cost and Utilization Project (HCUP). In the MEPS analyses, low income is defined as less than 200% of the federal poverty level and higher income is defined as 200% of the federal poverty level or more. For the HCUP analyses, median household income for the patient's zip code of residence is used to assign community-level income to individual hospitalizations. RESULTS: Coverage. Children from low-income families were more likely than children from middle-high-income families to be uninsured (13.0% vs 5.8%) or covered by public insurance (50.8% vs 7.3%), and less likely to be privately insured (36.2% vs 87.0%). Utilization. Children from low-income families were less likely to have had a medical office visit or a dental visit than children from middle-high-income families (63.7% vs 76.5% for office-based visits and 28.8% vs 51.4% for dental visits) and less likely to have medicines prescribed (45.1% vs 56.4%) or have utilized hospital outpatient services (5.2% vs 7.0%), but more likely to have made trips to the emergency department (14.6% vs 11.4%). Although low-income children comprise almost 40% of the child population, one quarter of total medical expenditures were for these children. Hospital Discharges. Significant differences by community-level income occurred in specific characteristics of hospitalizations, including admissions through the emergency department, expected payer, mean total charges per day, and reasons for hospital admission. Leading reasons for admission varied by income within and across age groups. Quality. Low-income children were more likely than middle-high income children to have their parents report a big problem getting necessary care (2.4% vs 1.0%) and getting a referral to a specialist (11.5% vs 5.3%). Low-income children were at least twice as likely as middle-high-income children to have their parents report that health providers never/sometimes listened carefully to them (10.0% vs 5.1%), explained things clearly to the parents (9.6% vs 3.4%), and showed respect for what the parents had to say (9.2% vs 4.2%). Children from families with lower community-level incomes were more likely to experience ambulatory-sensitive hospitalizations. Racial/Ethnic Differences Between Income Groups. Use and expenditure patterns for most services were not significantly different between low- and middle-high-income black children and were lower than those for white children. CONCLUSIONS: While health insurance coverage is still an important factor in obtaining health care, the data suggest that efforts beyond coverage may be needed to improve access and quality for low-income children overall and for children who are racial and ethnic minorities, regardless of income. PMID- 15656709 TI - Two-photon excitation induced fluorescence of a trifluorophore-labeled DNA. AB - Two-photon excitation of a trifluorophore (6-carboxyfluorescein, N,N,N',N' tetramethyl-6-carboxyrhodamine and cyanine-5 monofunctional dye) labeled DNA, which has a scaffold of 26 nucleotides, was achieved using focused laser light of a Q-switched Nd-YAG laser (1064 nm). The observed fluorescence signature (emission ratio from the three fluorophores) of the labeled DNA after two-photon excitation is very different from the fluorescence signatures produced by one photon excitation at different wavelength. The additional fluorescence signatures produced by two-photon excitation of the fluorescent oligonucleotides will facilitate their use as combinatorial fluorescence energy transfer tags for multiplex genetic analysis. PMID- 15656711 TI - Inflammatory bowel disease. Part II: Crohn's disease--pathophysiology and conventional and alternative treatment options. AB - Crohn's disease, a subcategory of inflammatory bowel disease, contributes to significant morbidity, particularly in industrialized nations. It can affect people of any age, but is more commonly diagnosed in adolescence and young adulthood. Inflammation and ulceration occurs primarily in the terminal ileum and colon, although any portion of the intestinal tract can be affected. No etiology has been identified for Crohn's disease, although a number of factors contribute to its etiopathogenesis, including genetic, microbial, inflammatory, immune, and permeability abnormalities. Conventional medications are not curative but can contribute to resolution of acute flare-ups and help maintain remission. Because significant side effects are associated with many these medications, more natural interventions to help maintain remission should be considered. Associated nutrient deficiencies, dietary interventions, and nutrient and botanical supplementation are discussed. PMID- 15656712 TI - Pharmacologically active natural compounds for lung cancer. AB - This article consists of an analysis of the available scientific research on botanically derived compounds that have potential efficacy in the treatment of lung cancer. The mechanisms of activity reviewed include alkylating agents, topoisomerase poisons, DNA synthesis inhibitors, protein synthesis inhibitors, immunoceuticals, and lipoxygenase inhibitors. Selection criteria include: (1) products whose activity have at least minimal scientific confirmation - preclinical (in vitro, in vivo) or clinical; (2) products with a well-defined chemical composition; or (3) products with a well-known or scientifically plausible mechanism of activity. PMID- 15656713 TI - Evaluation of the effects of Neptune Krill Oil on the clinical course of hyperlipidemia. AB - OBJECTIVE: To assess the effects of krill oil on blood lipids, specifically total cholesterol, triglycerides, low-density lipoprotein (LDL), and high-density lipoprotein (HDL). METHODS: A multi-center, three-month, prospective, randomized study followed by a three-month, controlled follow-up of patients treated with 1 g and 1.5 g krill oil daily. Patients with hyperlipidemia able to maintain a healthy diet and with blood cholesterol levels between 194 and 348 mg per dL were eligible for enrollment in the trial. A sample size of 120 patients (30 patients per group) was randomly assigned to one of four groups. Group A received krill oil at a body mass index (BMI)-dependent daily dosage of 2-3 g daily. Patients in Group B were given 1-1.5 g krill oil daily, and Group C was given fish oil containing 180 mg eicosapentaenoic acid (EPA) and 120 mg docosahexaenoic acid (DHA) per gram of oil at a dose of 3 g daily. Group D was given a placebo containing microcrystalline cellulose. The krill oil used in this study was Neptune Krill Oil, provided by Neptune Technologies and Bioresources, Laval, Quebec, Canada. OUTCOME MEASURES: Primary parameters tested (baseline and 90-day visit) were total blood cholesterol, triglycerides, LDL, HDL, and glucose. RESULTS: Krill oil 1-3 g per day (BMI-dependent) was found to be effective for the reduction of glucose, total cholesterol, triglycerides, LDL, and HDL, compared to both fish oil and placebo. CONCLUSIONS: The results of the present study demonstrate within high levels of confidence that krill oil is effective for the management of hyperlipidemia by significantly reducing total cholesterol, LDL, and triglycerides, and increasing HDL levels. At lower and equal doses, krill oil was significantly more effective than fish oil for the reduction of glucose, triglycerides, and LDL levels. PMID- 15656714 TI - Monograph. Angelica sinensis. PMID- 15656715 TI - Monograph. Boron. PMID- 15656716 TI - Monograph. Valeriana officinalis. PMID- 15656717 TI - Domestic violence and employment: a qualitative study. AB - This exploratory study sought to gather detailed information about how domestic violence affects women's employment, specifically to identify the types of job interference tactics used by abusers and their consequences on women's job performance; identify and understand the context associated with disclosure about victimization to employers and coworkers; and identify the supports offered to employees after disclosure. Qualitative analyses, guided by grounded theory, revealed that perpetrators exhibited job interference behaviors before, during, and after work. Abuser tactics reduced women's job performance as measured by absenteeism, tardiness, job leavings, and terminations. Among women who disclosed victimization to employers, informal and formal job supports were offered. Workplace supports led to short-term job retention, but fear and safety issues mitigated employers' attempts to retain workers. PMID- 15656720 TI - Predicting occupational coping responses: the interactive effect of gender and work stressor context. AB - This study examined whether gender and work stressor context interacted to affect reports of occupational coping strategies. Participants (N=113) were asked to indicate how they would cope with both self-focused (i.e., solitary) and interpersonal (i.e., group) work overload contexts by providing responses to the Cybernetic Coping Scale (J. Edwards & A. J. Baglioni, 1993). Consistent with the hypothesized interaction, male and female participants evidenced similar coping strategies in the self-focused work overload context, whereas female participants showed significantly higher use of coping (T(2)=.08, p<.05) and, in particular, greater emotional-expressive coping (T(2)=.07, p<.01) in the interpersonal work overload context as compared with male participants. Theoretical and practical implications were discussed. PMID- 15656719 TI - Working conditions and three types of well-being: a longitudinal study with self report and rating data. AB - The mean of self-report and observer ratings of working conditions was used to predict 3 types of well-being in 52 young workers: general well-being, job related well-being, and spillover from work to nonwork domains. Longitudinally, job control predicted spillover. There was no strong evidence for reverse causation. Synchronously, Time 2 job stressors predicted all types of well-being, and job control predicted general well-being. Because dependent variables at Time 1 are controlled for, this indicates short-term effects. Results for stressors are in line with a stress reaction model, indicating a rather quick symptom development but reversibility. The effect of control on spillover, however, suggests a sleeper effect model, with symptoms appearing with delay. PMID- 15656718 TI - Do the effects of job stressors on health persist over time? A longitudinal study with observational stressor measures. AB - Observational work analyses were conducted among 222 office workers (131 women, 91 men) from 12 German companies to measure stressors independently of worker appraisal. Stressors were determined twice (1990, 1991), along with 7 health indicators separately assessed by questionnaire. Health indicators were assessed again in 1992 and once more in 1998. This 2-wave, 2-variable design with 2 follow up measurements of health indicators determines (a) the causal directions using cross-lagged partial correlations (stressors seem to have an impact on health); (b) the strength of the impact (cross-lagged correlations for Waves 1 to 4 range from .18Ser mutation in peripherin/rds causes a loss-of-function phenotype in transgenic mice. AB - P/rds (peripherin/retinal degeneration slow) is a photoreceptor-specific membrane glycoprotein necessary for outer segment disc morphogenesis. Mutations in P/rds are associated with different blinding diseases. A C214S (Cys214-->Ser) missense mutation has been shown to be the cause for a late-onset form of ADRP (autosomal dominant retinitis pigmentosa) in humans. In the present study, we generated transgenic mice expressing P/rds with the C214S mutation and crossed them into rds mutant mice to elucidate the mechanism underlying the pathology of ADRP. Although an ample amount of transgene message was formed in C214S retinas from all transgenic lines, only a trace amount of the mutant protein was detected by Western blotting and immunoprecipitation. C214S mice on the wild-type or rds+/- backgrounds exhibited no signs of negative effects of the mutation on retinal structure or function, suggesting a loss-of-function phenotype. This phenotype is further supported by the absence of outer segment formation in the C214S mice on the rds-/- background. In contrast, expression of C214S protein in the inner retinal cells of transgenic mice or in COS cells resulted in the formation of a substantial amount of mutant protein, signifying a possible photoreceptor specific regulation of P/rds. These results provide evidence that the loss-of function phenotype seen in C214S transgenic mice shows a disease progression that correlates with ADRP patients carrying the same mutation, indicating that the C214S mutation on one allele of P/rds results in haploinsufficiency. PMID- 15656786 TI - The CYP2B2 phenobarbital response unit contains binding sites for hepatocyte nuclear factor 4, PBX-PREP1, the thyroid hormone receptor beta and the liver X receptor. AB - A 163 bp enhancer in the CYP2B2 5' flank confers PB (phenobarbital) inducibility and constitutes a PBRU (PB response unit). The PBRU contains several transcription factor binding sites, including NR1, NR2 and NR3, which are direct repeats separated by 4 bp of the nuclear receptor consensus half-site AGGTCA, as well as an ER (everted repeat) separated by 7 bp (ER-7). Constitutive androstane receptor (CAR)-RXR (retinoic X receptor) heterodimers are known to bind to NR1, NR2 and NR3. Electrophoretic mobility-shift analysis using nuclear extracts from livers of untreated or PB-treated rats revealed binding of several other proteins to different PBRU elements. Using supershift analysis and in vitro coupled transcription and translation, the proteins present in four retarded complexes were identified as TRbeta (thyroid hormone receptor beta), LXR (liver X receptor), HNF-4 (hepatocyte nuclear factor 4) and heterodimers of PBX-PREP1 (pre B cell homoeobox-Pbx regulatory protein 1). LXR-RXR heterodimers bound to NR3 and TRbeta bound to NR3, NR1 and ER-7, whereas the PBX-PREP1 site is contained within NR2. The HNF-4 site overlaps with NR1. A mutation described previously, GRE1m1, which decreases PB responsiveness, increased the affinity of this site for HNF-4. The PBRU also contains a site for nuclear factor 1. The PBRU thus contains a plethora of transcription factor binding sites. The profiles of transcription factor binding to NR1 and NR3 were quite similar, although strikingly different from, and more complex than, that of NR2. This parallels the functional differences in conferring PB responsiveness between NR1 and NR3 on the one hand, and NR2 on the other. PMID- 15656788 TI - Role of cis-acting elements in the control of SERCA2b Ca2+ pump mRNA decay by nuclear proteins. AB - Alternative splicing at position 3495 b yields SERCA2 (sarco/endoplasmic reticulum Ca2+ pump 2) RNA species, namely SERCA2a and SERCA2b which differ in 3' end regions. This results in SERCA2b RNA being less stable. In vitro decay experiments show that, in the presence of protein extracts from nuclei of LVMs (left ventricular myocytes), the rate of decay of both SERCA2b RNA and synthetic RNA from its 3'-region is greater than that of the corresponding SERCA2a RNA. To search for cis-acting instability elements in the 3'-region of SERCA2b, we examined the effects of LVM nuclear protein extracts on the in vitro decay of six short overlapping capped [m7G(5')ppp(5')Gm] and polyadenylated (A40) RNA fragments from the 3'-end region (3444-4472) of SERCA2b. The proximal fragment 2B1 (3444-3753) was the most unstable. 2B1 RNA without a cap or a polyadenylated tail was analysed further in electrophoretic mobility-shift assays, and was observed to bind to protein(s) in the nuclear extracts. Based on competition for binding to nuclear proteins between radiolabelled 2B1 RNA and short unlabelled RNA fragments, the cis-acting element involved in this binding was the sequence 2B1-4. 2B1-4 is a 35-base (3521-3555, CCAGUCCUGCUCGUUGUGGGCGUGCACCGAGGGGG) GC rich region just past the splice site (3495). Nuclear extracts decreased the electrophoretic mobility of the radiolabelled 2B1-4 RNA which bound to two proteins (19 and 21 kDa) in cross-linking experiments. Excess 2B1-4 RNA decreased the decay of the 2B1 RNA by the nuclear protein extract. 2B1-del 4 RNA (2B1 with the 2B1-4 domain deleted) also decayed more slowly than the control 2B1 RNA. Thus SERCA2b contains a novel GC-rich cis-acting element involved in its decay by nuclear proteins. PMID- 15656789 TI - Biosynthesis of agmatine in isolated mitochondria and perfused rat liver: studies with 15N-labelled arginine. AB - An important but unresolved question is whether mammalian mitochondria metabolize arginine to agmatine by the ADC (arginine decarboxylase) reaction. 15N-labelled arginine was used as a precursor to address this question and to determine the flux through the ADC reaction in isolated mitochondria obtained from rat liver. In addition, liver perfusion system was used to examine a possible action of insulin, glucagon or cAMP on a flux through the ADC reaction. In mitochondria and liver perfusion, 15N-labelled agmatine was generated from external 15N-labelled arginine. The production of 15N-labelled agmatine was time- and dose-dependent. The time-course of [U-15N4]agmatine formation from 2 mM [U-15N4]arginine was best fitted to a one-phase exponential curve with a production rate of approx. 29 pmol x min(-1) x (mg of protein)(-1). Experiments with an increasing concentration (0- 40 mM) of [guanidino-15N2]arginine showed a Michaelis constant Km for arginine of 46 mM and a Vmax of 3.7 nmol x min(-1) x (mg of protein)(-1) for flux through the ADC reaction. Experiments with broken mitochondria showed little changes in Vmax or Km values, suggesting that mitochondrial arginine uptake had little effect on the observed Vmax or Km values. Experiments with liver perfusion demonstrated that over 95% of the effluent agmatine was derived from perfusate [guanidino 15N2]arginine regardless of the experimental condition. However, the output of 15N-labelled agmatine (nmol x min(-1) x g(-1)) increased by approx. 2-fold (P<0.05) in perfusions with cAMP. The findings of the present study provide compelling evidence that mitochondrial ADC is present in the rat liver, and suggest that cAMP may stimulate flux through this pathway. PMID- 15656790 TI - AlphaII-spectrin interacts with Tes and EVL, two actin-binding proteins located at cell contacts. AB - The spectrin-based membrane skeleton, a multi-protein scaffold attached to diverse cellular membranes, is presumed to be involved in the stabilization of membranes, the establishment of membrane domains as well as in vesicle trafficking and nuclear functions. Spectrin tetramers made of alpha- and beta subunits are linked to actin microfilaments, forming a network that binds a multitude of proteins. The most prevalent alpha-spectrin subunit in non-erythroid cells, alphaII-spectrin, contains two particular spectrin repeats in its central region, alpha9 and alpha10, which host an Src homology 3 domain, a tissue specific spliced sequence of 20 residues, a calmodulin-binding site and major cleavage sites for caspases and calpains. Using yeast two-hybrid screening of kidney libraries, we identified two partners of the alpha9-alpha10 repeats: the potential tumour suppressor Tes, an actin-binding protein mainly located at focal adhesions; and EVL (Ena/vasodilator-stimulated phosphoprotein-like protein), another actin-binding protein, equally recruited at focal adhesions. Interactions between spectrin and overexpressed Tes and EVL were confirmed by co immunoprecipitation. In vitro studies showed that the interaction between Tes and spectrin is mediated by a LIM (Lin-11, Isl-1 and Mec3) domain of Tes and by the alpha10 repeat of alphaII-spectrin whereas EVL interacts with the Src homology 3 domain located within the alpha9 repeat. Moreover, we describe an in vitro interaction between Tes and EVL, and a co-localization of these two proteins at focal adhesions. These interactions between alphaII-spectrin, Tes and EVL indicate new functions for spectrin in actin dynamics and focal adhesions. PMID- 15656791 TI - Probing a putative dantrolene-binding site on the cardiac ryanodine receptor. AB - Dantrolene is an inhibitor of intracellular Ca2+ release from skeletal muscle SR (sarcoplasmic reticulum). Direct photoaffinity labelling experiments using [3H]azidodantrolene and synthetic domain peptides have demonstrated that this drug targets amino acids 590-609 [termed DP1 (domain peptide 1)] of RyR1 (ryanodine receptor 1), the skeletal muscle RyR isoform. Although the identical sequence exists in the cardiac isoform, RyR2 (residues 601-620), specific labelling of RyR2 by dantrolene has not been demonstrated, even though some functional studies show protective effects of dantrolene on heart function. Here we test whether dantrolene-active domains exist within RyR2 and if so, whether this domain can be modulated. We show that elongated DP1 sequences from RyR1 (DP1 2s; residues 590-628) and RyR2 (DP1-2c; residues 601-639) can be specifically photolabelled by [3H]azidodantrolene. Monoclonal anti-RyR1 antibody, whose epitope is the DP1 region, can recognize RyR1 but not RyR2 in Western blot and immunoprecipitation assays, yet it recognizes both DP1-2c and DP1-2s. This suggests that although the RyR2 sequence has an intrinsic capacity to bind dantrolene in vitro, this site may be poorly accessible in the native channel protein. To examine whether it is possible to modulate this site, we measured binding of [3H]dantrolene to cardiac SR as a function of free Ca2+. We found that > or =10 mM EGTA increased [3H]dantrolene binding to RyR2 by approximately 2 fold. The data suggest that the dantrolene-binding site on RyR2 is conformationally sensitive. This site may be a potential therapeutic target in cardiovascular diseases sensitive to dysfunctional intracellular Ca2+ release. PMID- 15656792 TI - Direct comparison of the specificity of gene silencing using antisense oligonucleotides and RNAi. AB - RNAi (RNA interference) and ASO (antisense oligonucleotide) technologies are the most commonly used approaches for silencing gene expression. However, the specificity of such powerful tools is an important factor to correctly interpret the biological consequences of gene silencing. In the present study, we examined the effects of acute loss of Ser/Thr kinase PDK1 (3-phosphoinositide-dependent kinase 1) expression using ASO and RNAi, and compared, for the first time, these two techniques using Affymetrix microarrays. We show that both ASO- and siRNA (small interfering RNA)-mediated knock-down of PDK1 expression strongly inhibited cell proliferation, although by different mechanisms, thereby questioning the specificity of these reagents. Using microarray analysis, we characterized the specificity of the ASO- and siRNA-mediated gene silencing of PDK1 by examining expression profiles 48 and 72 h following oligonucleotide transfection. At 48 h, a PDK1-dependent pattern of gene alterations was detectable, despite a large number of non-specific changes due to transfection of control nucleic acids. These non-specific alterations became more apparent at the 72 h time point, and obscured any PDK1-specific pattern. This study underscores the importance of defining appropriate control ASOs and siRNAs, using multiple oligonucleotides for each target and preferably short time points following transfection to avoid misinterpretation of the phenotype observed. PMID- 15656796 TI - Updates from the British Association of Dermatologists 84th annual meeting, 6-9 July 2004, Belfast, U.K. AB - Herein is a synopsis of the main research and clinical findings presented at the British Association of Dermatologists meeting held during 6-9 July 2004, in Belfast, U.K. The conference highlighted the progress that has been made in understanding the increasing biological, epidemiological and therapeutic advances that have been made recently in the field of dermatology. The authors highlight the more important advances or summaries, but this is not meant as a substitute for reading the conference proceedings and related references quoted in this article. PMID- 15656793 TI - H11 has dose-dependent and dual hypertrophic and proapoptotic functions in cardiac myocytes. AB - We have shown previously that H11, a serine/threonine kinase, is up-regulated in a heart subjected to ischaemia/reperfusion. In the present study, we have characterized the cellular function of H11, using neonatal rat cardiac myocytes. Although transduction of adenovirus harbouring H11 at low doses increased the cell size, at higher doses it induced apoptosis in cardiac myocytes. Apoptosis was not observed when adenovirus harbouring H11-KI (kinase-inactive mutant of H11) was used, suggesting that the proapoptotic effect of H11 is kinase dependent. The hypertrophic effect of H11 at high doses was unmasked when apoptosis was inhibited by the caspase inhibitor DEVD-CHO, suggesting that H11 stimulates both hypertrophy and apoptosis in parallel. H11-KI induced hypertrophy even at high doses, indicating that H11 stimulates hypertrophy through kinase independent mechanisms. H11-KI activated Akt, and cardiac hypertrophy induced by H11-KI was blocked by LY294002, an inhibitor of phosphoinositide 3-kinase. Co immunoprecipitation analyses indicated that H11 interacts with the alpha subunit of CK2 (casein kinase 2). Overexpression of H11 decreased the kinase activity of CK2. DRB (5,6-dichloro-1-beta-D-ribofuranosyl-benzimidazole), an inhibitor of CK2, mimicked the effect of H11, whereas DRB and H11 failed to exhibit additive effects on apoptosis, suggesting that H11 and DRB utilize a common mechanism to induce apoptosis, namely inhibition of CK2. In summary, H11 is a dual-function kinase in cardiac cells: it induces hypertrophy at low doses through kinase independent activation of Akt, whereas it causes apoptosis at high doses through protein kinase-dependent mechanisms, in particular by physical interaction with and subsequent inhibition of CK2. PMID- 15656795 TI - Pruritus precipitated by hydroxyethyl starch: a review. AB - Hydroxyethyl starch (HES) is widely used for fluid management in broad populations of patients, particularly in the surgery and intensive care settings. Pruritus, often manifested as pruritic crises, is increasingly being recognized as a common major adverse effect of HES administration. This iatrogenic form of pruritus is frequently severe and protracted with a serious negative impact on patient quality of life, including sleep disturbance, disruption of daily routine and mental distress. Such pruritus is generally refractory to available therapies and can persist for up to 12-24 months. All currently clinically available HES solutions entail the risk of pruritus, including those of diverse molecular weights and substitutions. Although dose dependent, HES-induced pruritus nevertheless can often be provoked by relatively low routine doses. The pathophysiological basis for pruritus is the widespread tissue deposition of HES, prominently in macrophages. HES tissue deposits are long lasting and sometimes massive. Usually several weeks elapse between HES exposure and the onset of pruritus. Consequently, it is important to inquire about prior HES exposure in the diagnostic evaluation of pruritus sine materia. Awareness about the scope of the pruritus problem needs to increase among physicians administering HES. Well designed clinical outcome studies are needed to assess more fully the incidence, dose dependency and mechanisms of pruritus with particular HES solutions. PMID- 15656797 TI - Suggestive linkage of familial primary cutaneous amyloidosis to a locus on chromosome 1q23. AB - BACKGROUND: There is a high incidence of primary cutaneous amyloidosis (PCA) in South America, South-east Asia and Taiwan. To date, the aetiology of PCA remains unknown, but it is believed to be multifactorial. Although most cases are sporadic, some patients have a family history. Familial aggregation and different susceptibility to PCA among ethnic groups suggest that genetic factors may play an important role in its pathogenesis. However, no genetic loci for familial PCA (FPCA) have been identified so far. OBJECTIVES: In order to identify the susceptibility gene of FPCA, we took a candidate gene approach and performed linkage analysis on chromosome 1q21.3-24.2, including the 1q23.2 region where the gene encoding serum amyloid P component (APCS) is located. PATIENTS AND METHODS: Nine FPCA families including 29 individuals affected with PCA were recruited for this linkage study. Initially, 11 highly polymorphic microsatellite markers spanning the region from 1q21.3 to 1q24.2 were genotyped and revealed a suggestive linkage region. This region was further fine-mapped with seven additional markers. We also re-sequenced the 2.5-kb genomic region of the APCS gene in 29 affected and 42 control individuals. Two-point and multipoint linkage analyses were performed using the LINKAGE program. Nonparametric linkage (NPL) analysis and reconstruction of haplotypes were performed with the GENEHUNTER program. RESULTS: Both two-point and multipoint linkage analysis for all 11 markers generated negative or small positive total lod scores for all nine families. However, when we considered only three families, a maximum two-point total lod score of 2.09 was obtained for the marker D1S2844 at theta = 0.01. A plateau of multipoint total lod score between D1S2768 and D1S2878 with a maximum of 2.48 at the marker D1S2844 was observed. A maximum NPL score of 3.11 (P = 0.008) was also obtained for the marker D1S2878. However, re-sequencing of the APCS gene identified no functional mutation. CONCLUSIONS: Both parametric and nonparametric linkage evidence suggested that a possible susceptibility locus for a subset of FPCA might exist on chromosome 1q23. This is the first report demonstrating suggestive evidence of linkage of FPCA to a locus in this candidate region. No functional sequence variations of the APCS gene were found to be associated with this disease among the study families. Our data imply the existence of at least one additional locus responsible for FPCA in these families, confirming genetic heterogeneity of this skin disorder. PMID- 15656798 TI - The mitogen-activated protein kinases p38 and ERK1/2 are increased in lesional psoriatic skin. AB - BACKGROUND: Alterations in specific signal transduction pathways may explain the hyperproliferation and abnormal differentiation of the keratinocytes as well as the increased expression of inflammatory cytokines seen in psoriasis. Major signalling pathways used by eukaryotic cells to transduce extracellular signals into cellular responses impinge on the mitogen-activated protein kinases (MAPKs). OBJECTIVES: To investigate the expression of the MAPK p38, extracellular signal regulated kinase (ERK) and c-Jun NH2-terminal kinase (JNK) in psoriatic skin. METHODS: Keratome biopsies were taken from patients with plaque-type psoriasis. Western blot analysis was used to determine p38, ERK and JNK activity and protein levels, whereas kinase assays were used to examine the kinase activity of p38. RESULTS: We demonstrated increased levels of the phosphorylated forms of p38 and ERK1/2 in lesional psoriatic skin compared with nonlesional psoriatic skin. No abnormality was found in the activation and expression of JNK1/2. Ex vivo kinase assays confirmed the increased activation of p38, and furthermore demonstrated increased kinase activity of the p38 isoforms p38alpha, p38beta and p38delta in lesional compared with nonlesional psoriatic skin. p38gamma was not detected in the psoriatic skin. Clearance of the psoriatic lesions, induced by climatotherapy at the Dead Sea for 4 weeks, led to a normalization in the activity of both p38 and ERK1/2. CONCLUSIONS: Taken together, our results demonstrate that the activity of the MAPKs p38alpha, p38beta and p38delta and ERK1/2 are increased in lesional psoriatic skin compared with nonlesional psoriatic skin, and that clearance of psoriasis normalizes the p38 and ERK1/2 activity. Thus, p38 and ERK1/2 might be potential targets in the treatment of psoriasis. PMID- 15656800 TI - Analysis of genes responding to ultraviolet B irradiation of HaCaT keratinocytes using a cDNA microarray. AB - BACKGROUND: Ultraviolet (UV) B irradiation causes many important biological changes in skin, which lead to pathophysiological alterations of the homeostatic environment. OBJECTIVES: To gain more insight into the molecular events provoked by UVB irradiation, we performed cDNA microarray analysis. METHODS: Immortalized HaCaT keratinocytes were irradiated with a high cytotoxic dose of UVB (50 mJ cm( 2)), and total RNA was isolated. Fluorescently labelled probes were prepared by reverse transcription and were hybridized with cDNA microarray slides made using 840 cDNA clones. RESULTS: Time-course cDNA microarray analysis revealed the global gene expression profile after UVB exposure. Of 840 genes tested, 192 genes showed changes in their expression levels at one or more of four time points. The genes were clustered into four groups according to their expression patterns in a self-organizing maps analysis. Classification of these genes into nine functional categories revealed that UVB irradiation affected several biological processes. The genes that were first upregulated and then returned to normal levels included several genes related to the inhibition of cell growth and the proteasome pathway. Conversely, the expressions of many genes involved in the cytoskeleton, signal transduction, metabolism and transcription were first downregulated or unchanged and then upregulated later, reflecting the recovery of UVB-damaged cellular activities. CONCLUSIONS: These results demonstrate the complexity of the transcriptional profile of the UVB response, and provide a basis for the global characterization of UV-regulated gene expression. PMID- 15656799 TI - Somatic mutations in the PTCH, SMOH, SUFUH and TP53 genes in sporadic basal cell carcinomas. AB - BACKGROUND: Basal cell carcinoma (BCC) of the skin is the most common human cancer. The genetic alterations underlying BCC development are only partly understood. OBJECTIVES: To investigate further the molecular genetics of sporadic BCCs, we performed mutation analyses of 10 skin cancer-associated genes in 42 tumours. METHODS: Single-strand conformational polymorphism analysis followed by DNA sequencing was used to screen for mutations in the sonic hedgehog pathway genes PTCH, SMOH, SUFUH and GLI1, in the TP53 tumour suppressor gene, and in the proto-oncogenes NRAS, KRAS, HRAS, BRAF and CTNNB1. Microsatellite markers flanking the PTCH, SUFUH and TP53 loci at 9q22, 10q24 and 17p13, respectively, were studied for loss of heterozygosity (LOH). RESULTS: PTCH mutations were found in 28 of 42 tumours (67%). Microsatellite analysis revealed LOH on 9q22 in 20 of 38 tumours investigated (53%), including 14 tumours with and six tumours without PTCH mutations. SMOH mutations were identified in four of the 42 BCCs (10%) while two tumours demonstrated mutations in SUFUH, including one missense mutation and one silent mutation. None of the BCCs showed LOH at markers flanking the SUFUH locus. Seventeen BCCs (40%) carried TP53 mutations, with only three tumours showing evidence of biallelic TP53 inactivation. TP53 mutations were present in BCCs with and without mutations in PTCH, SMOH or SUFUH. Interestingly, 72% of the TP53 alterations were presumably ultraviolet (UV)-induced transition mutations. In contrast, only 40% of the PTCH and SMOH alterations corresponded to UV signature mutations. No mutations were identified in GLI1, NRAS, KRAS, HRAS, BRAF or CTNNB1. CONCLUSIONS: Our data confirm the importance of PTCH, SMOH and TP53 mutations in the pathogenesis of sporadic BCCs. SUFUH alterations are restricted to individual cases while the other investigated genes do not appear to be important targets for mutations in BCCs. PMID- 15656801 TI - Blood flow in psoriatic plaques before and after selective treatment of the superficial capillaries. AB - BACKGROUND: Blood flow is substantially raised in psoriatic plaques. In addition, mechanisms of vasoconstriction and vasodilatation (locally and neurally mediated), although intact, are altered in magnitude. The elevated blood flow is considered to be a result of abnormalities (increase in vessel number, width and length) in the superficial capillary loops rather than changes in the deeper feeding vessels (arterioles). OBJECTIVES: To determine if selective thermolysis of psoriatic capillaries with a pulsed dye laser (PDL) leads to normalization of blood flow and also if the vasoconstrictor and vasodilator responses are returned to normal magnitude. METHODS: Laser Doppler red cell flux was recorded from plaques on the forearm or elbow (untreated plaque site) and from clinically uninvolved skin at an equivalent site on the opposite limb. Plaques were treated on three occasions, at 2-weekly intervals, with a PDL. Laser Doppler red cell flux measurements were then repeated, 2 weeks after the final laser treatment was performed (treated plaque site). RESULTS: There was significant clinical improvement in plaques after treatment (P = 0.02), but complete clearance of lesions did not occur. Blood flow in plaques under basal conditions remained significantly elevated above blood flow in clinically uninvolved skin, despite laser treatment (P < 0.001). The physiological tests of resistance vessel function showed that the laser did not impair the ability of psoriatic resistance vessels to constrict and dilate. However, there was only partial resolution of the percentage responses to the provocation tests towards the values recorded in clinically uninvolved skin. CONCLUSIONS: The results indicate that it is unlikely that the reduced resistance of the expanded superficial capillary bed is solely responsible for the massively elevated blood flow in plaque skin. It is more likely that the vascular abnormalities in psoriasis also extend to involve the deeper, larger resistance vessels (arterioles). PMID- 15656802 TI - Is ultrasound lymph node examination superior to clinical examination in melanoma follow-up? A monocentre cohort study of 373 patients. AB - BACKGROUND: There is still lack of consensus regarding the most effective follow up for stage I and II melanoma patients although some consensus conferences have provided guidelines stating that clinical examination should be the standard. OBJECTIVES: Our aim was to study the value of adding ultrasound lymph node examination (7.5 MHz) to the routine clinical examination recommended by French guidelines in melanoma follow-up. METHODS: A cohort of melanoma patients was enrolled between 1 July 1995 and 1 July 2000 in a follow-up protocol including clinical examination performed four times a year for thick melanomas (Breslow index > or = 1.5 mm) and twice a year for thin melanomas (Breslow index < 1.5 mm) according to French guidelines, and ultrasound lymph node examination performed every 6 months for thick melanomas and every year for thin melanomas. Follow-up was continued up to 1 July 2003. When clinical or ultrasound examination indicated signs of node recurrence, surgical biopsy of the involved node was performed. When ultrasound examination was only suspicious, another ultrasound examination was performed within the following 3 months. The results of both clinical and ultrasound examinations were compared with histopathology examination when node biopsy was performed. RESULTS: Ultrasound follow-up was performed for 373 patients (213 females and 160 males). Mean age at diagnosis of melanoma was 59 years (range 14-90, SD 15). In total, 1909 ultrasound examinations combined with clinical examination were analysed. Node biopsy was performed in 65 patients and demonstrated melanoma metastases in 54. Sensitivity of clinical examination and ultrasound examination was 71.4%[95% confidence interval (CI) 55.4-84.3] and 92.9 (95% CI 80.5-98.5), respectively, P = 0.02. Specificity of clinical examination and ultrasound examination was 99.6% (95% CI 99.2-99.8) and 97.8% (95% CI 97.0-98.4), respectively. Despite this apparent superiority of ultrasound examination over palpation, only 7.2% of the patients really benefited from ultrasound examination (earlier lymph node metastasis detection or avoidance of unnecessary surgery), while 5.9% had some deleterious effect from ultrasound examination (unnecessary stress caused by repetition of ultrasound examination for benign lymph nodes, useless removal of benign lymph node). CONCLUSIONS: This study confirms the greater sensitivity of ultrasound examination to clinical examination in the diagnosis of node metastases from cutaneous melanoma. However, the place of ultrasound in routine follow-up is at least questionable as only a very small proportion of patients (1.3%) really benefited from adding ultrasound examination to clinical examination. PMID- 15656803 TI - Cutaneous manifestations of the malignant carcinoid syndrome. AB - BACKGROUND: The incidence of carcinoid tumours is approximately 1.5 per 100 000 of the population. The malignant carcinoid syndrome, which is caused by circulating neuroendocrine mediators produced by the tumour, occurs in less than 10% of patients. Cutaneous involvement, although recognized in this rare syndrome, has rarely been reported. OBJECTIVES: To examine a series of patients with the malignant carcinoid syndrome, to establish the prevalence and types of cutaneous involvement and to assess whether these could be used as indicators of disease activity, progression and prognosis. METHODS: Patients with the malignant carcinoid syndrome who attended a multidisciplinary clinic for neuroendocrine tumours over a 23-month period between February 2001 and December 2002 were invited to participate in the study. This involved completion of a standard history proforma and a detailed cutaneous examination with findings recorded by clinical photography. RESULTS: Twenty-five patients were enrolled. All but one had experienced flushing, three had rosacea, five had clinical features of pellagra and two had scleroderma. Flushing was generally an early manifestation of the syndrome, whereas both the pellagra and scleroderma tended to occur in more advanced disease. CONCLUSIONS: This descriptive case series indicates that cutaneous features are not uncommon in the syndrome. They are readily diagnosed on clinical examination, and may be useful indicators of disease activity and prognosis. PMID- 15656804 TI - Clinicopathological features of cutaneous lesions of adult T-cell leukaemia/ lymphoma. AB - BACKGROUND: Adult T-cell leukaemia/lymphoma (ATLL) is a human malignancy associated with human T-cell leukaemia virus type I (HTLV-I). ATLL frequently involves the skin. OBJECTIVES: To correlate the clinicopathological features and prognosis in patients with ATLL and cutaneous lesions. METHODS: We examined the HTLV-I proviral state and the clinicopathological features of the cutaneous lesions in 80 patients with serum anti-ATL antibody, to clarify the correlation between macroscopic/histopathological findings and prognosis. Southern blot analysis was performed in all cases to detect monoclonal HTLV-I proviral DNA integration. RESULTS: The cutaneous lesions of 46 patients were positive for proviral DNA integration. The median survival time of patients with monoclonal proviral DNA integration in cutaneous lesions was 14 months, which was markedly shorter than that of patients negative for proviral DNA integration (72 months). Of the 46 patients with proviral DNA, 21 had solitary or multiple red nodules (including three with subcutaneous induration), eight had multiple red papules and 17 had erythema. Patients with papules and nodules had poorer prognosis than those with erythema. Histopathologically, the prognosis was poorer in patients with nodular or diffuse infiltration of medium-sized to large lymphoma cells, compared with those with perivascular infiltration of small to medium-sized lymphoma cells. CONCLUSIONS: Our results show a close correlation between clinicopathological features of HTLV-I-associated cutaneous lesions and prognosis. PMID- 15656805 TI - Lymphoma in patients with dermatitis herpetiformis and their first-degree relatives. AB - BACKGROUND: The risk for lymphoma is increased in both dermatitis herpetiformis (DH) and in coeliac disease. The lymphoma most associated with coeliac disease is enteropathy-associated T-cell lymphoma. OBJECTIVES: To study the occurrence and type of lymphoma in a large series of patients with DH and their first-degree relatives. METHODS: The occurrence of lymphoma was studied in 1104 patients consecutively diagnosed with DH in two university hospitals during 1969-2001. A questionnaire was sent to 341 patients to examine the occurrence of lymphoma in their 1825 first-degree relatives. To analyse whether the DH patients with lymphoma had adhered to a gluten-free diet similarly to the patients without lymphoma, two age- and sex-matched patients with DH served as controls for each index case. Data on the gluten-free diet were collected from prospectively completed dietary forms and also from medical records. RESULTS: Eleven (1%) patients contracted lymphoma 2-31 years after the diagnosis of DH. Eight had B cell-type lymphoma, two enteropathy-associated T-cell lymphoma and one remained unclassified due to missing material. Three (0.2%) of the first-degree relatives contracted lymphoma, all B-cell type. The 11 DH patients with lymphoma had adhered to a gluten-free diet significantly less strictly than the DH controls without lymphoma. CONCLUSIONS: The present study documents that patients with DH can have both B- and T-cell lymphoma. The DH patients with lymphoma had not adhered as strictly to the gluten-free diet as the control patients without lymphoma. The occurrence of lymphoma in the first-degree relatives was lower than in the patients with DH. PMID- 15656806 TI - Surveillance of patients at high risk for cutaneous malignant melanoma using digital dermoscopy. AB - BACKGROUND: Dermoscopy has improved the sensitivity and specificity of clinical diagnosis of melanoma from 60% to over 90%. However, in order not to miss melanoma a certain percentage of suspicious but benign lesions has to be excised. OBJECTIVES: To evaluate the dermoscopic changes and the rates of excision in benign melanocytic naevi and cutaneous malignant melanoma in long-term follow-up of high-risk patients using digital dermoscopy. METHODS: Digital dermoscopic images of 2015 atypical melanocytic naevi in 196 high-risk patients were analysed retrospectively. Among others, the following data were collected for each naevus: changes in surface area, overall architecture, dermoscopic patterns and distribution of pigmentation. All tumours suspicious for melanoma or showing asymmetrical changes were excised. RESULTS: During a median follow-up time of 25 months 128 (6.4%) of all naevi showed changes in size or architecture. Eighty-six per cent of all changes in patients who attended more than one visit were observed at the first follow-up visit. Thirty-three lesions showing changes were excised and two melanomas in situ and 31 melanocytic naevi were diagnosed. CONCLUSIONS: Follow-up examinations using digital dermoscopy revealed unchanged morphology in the large majority of melanocytic naevi. Excisions were only performed in cases of asymmetrical growth, asymmetrical changes of pigmentation, or development of dermoscopic features indicative of melanoma. The ratio of 33 lesions excised in order to identify two melanomas in situ seems reasonable and may be further reduced in future. PMID- 15656807 TI - Relation between diagnoses on severity, sick leave and loss of job among patients with occupational hand eczema. AB - BACKGROUND: Epidemiological studies indicate that occupational hand eczema (OHE) often is associated with persistent dermatitis and prolonged sick leave, which may lead to unemployment. Previous studies suggest that OHE caused by allergic contact dermatitis and atopic dermatitis (AD) carries the worst prognosis. OBJECTIVES: To evaluate and compare the severity and consequences of recognized OHE in different diagnostic and subdiagnostic groups. METHODS: Between October 2001 and November 2002, all new cases of recognized OHE were identified from the Danish National Board of Industrial Injuries Registry (758 cases). Severity was graded from 0 to 2 depending on the intensity of skin response and the frequency of relapse. To supplement the information from the Registry, we surveyed the study population using a postal questionnaire which included questions about disease duration, sick leave, current occupation and loss of job. RESULTS: The overall response rate to the questionnaire was 82%. We observed substantially greater severity among those with occupational irritant contact dermatitis (ICD) and AD than for any other diagnoses. Age above 50 years was also associated with increased severity of OHE. Prolonged sick leave due to OHE was reported by 19.9% and was associated with AD and severe OHE. We found a higher proportion of prolonged sick leave among those in food-related occupations (27.2%) compared with those in wet occupations (20.1%) and other occupations (16.5%). Twenty-three per cent reported that they had lost their job at least once during the past 12 months due to OHE. The only strong association with loss of job was food-related occupations. CONCLUSIONS: Occupational ICD and AD appear to be strongly associated with severity of OHE. AD and severity of OHE were independently associated with prolonged sick leave. Having a food-related occupation appears to be associated with increased risk of loss of job. PMID- 15656808 TI - Evaluation of a novel 308-nm monochromatic excimer light delivery system in dermatology: a pilot study in different chronic localized dermatoses. AB - BACKGROUND: Recently, units have been developed that are capable of delivering large fluences of narrowband ultraviolet (UV) B selectively to cutaneous lesions within a reasonable time. OBJECTIVES: To analyse the efficacy of a novel nonlaser 308-nm monochromatic excimer light (MEL) delivery system in various dermatoses usually treated by narrowband UVB phototherapy. METHODS: Fifty-four patients with chronic and resistant localized dermatoses were enrolled in a prospective study: 17 with palmoplantar pustular psoriasis, seven with plaque-type psoriasis, four with nail psoriasis, eight with chronic atopic dermatitis of the hands, 10 with chronic nonatopic dermatitis of the hands and eight with alopecia areata. The 308 nm xenon chloride MEL delivery system (Excilite; DEKA, Florence, Italy) was used to produce an average incident dose rate of 50 mW cm(-2) at a tube-to-skin distance of 15 cm and with a maximum irradiating area of 512 cm2. The initial dose was based on multiples of a predetermined minimal erythema dose (MED), and subsequent doses were based on the response to treatment. Treatments were scheduled weekly for a maximum of 10 weeks. Clinical responses were evaluated using photographic documentation and (except for alopecia areata) clinical score. RESULTS: The MED ranged from 250 to 350 mJ cm(-2) (mean +/- SD 318.2 +/- 28.4). MEL at 308 nm was the most effective for palmoplantar pustular psoriasis with a mean improvement of 79% after a mean of 5.3 treatments and a mean dose of 11.8 MED per treatment. Plaque-type psoriasis was significantly less sensitive to treatment and nail psoriasis demonstrated no benefit from treatment. Chronic palmar atopic dermatitis was cleared in two patients and the mean improvement was 54% as compared with 46% in patients with chronic nonatopic dermatitis of the hands. Four complete regrowths among the eight patients with alopecia were observed after a mean of 5.1 treatments. The percentages of improvement had significantly decreased at the 6-month visit, and only four patients (24%) with palmoplantar pustular psoriasis still demonstrated a significant improvement. Common side-effects included intense erythema and, more rarely, blisters, but these were well tolerated. CONCLUSIONS: Our preliminary results confirm the efficacy of this novel 308-nm MEL delivery system, which appears to be effective and safe for palmoplantar pustular psoriasis. To a lesser extent, plaque-type psoriasis, chronic atopic and nonatopic dermatitis of the hands and alopecia may also benefit from this treatment. PMID- 15656809 TI - Thick melanoma: the challenge persists. AB - BACKGROUND: Considerable resources have been channelled into primary and secondary prevention of cutaneous melanoma over the past 20 years. These efforts have been associated with a significant increase in the proportion of thin, good prognosis lesions and this is felt to be the principal reason for the current overall improvement in melanoma survival. OBJECTIVES: Analysis of Scottish Melanoma Group (SMG) data was carried out to identify the proportion of thick melanomas presenting over time. SMG data were used to characterize the patients presenting with thick melanoma. METHODS: Using data from the SMG database 915 patients (392 male and 523 female) first diagnosed with invasive melanoma > or = 3.5 mm thick in the two decades between 1979 and 1998, inclusive, were identified. The patients were from regions designated South-east Scotland, Tayside, Grampian and Highland, which together form half of all Scottish cases. RESULTS: The analysis shows that, although the proportion of thick, poor prognosis melanomas has decreased over time, the number presenting per year has not significantly altered. In the first decade, 50.5% of registrations were thick lesions and these fell to 31.0% in the second decade. In the first decade there were 419 cases (173 male), median age 66 years (range 5-99). Fifty-five patients were under the age of 40 years. Two hundred and twelve melanomas were nodular, 116 superficial spreading (SSM), 34 acral and 26 lentigo maligna melanoma. Sixty nine patients had either lymph node involvement or distant spread at presentation. Despite a 93.3% increase in the total number of melanoma registrations by the end of the second decade, there was relatively little change in the absolute numbers of thick lesions. The total number of thick lesions was 496 (220 male), an increase of 18.4%. Median age was greater, at 70 years (range 1-98), and 31 patients were under the age of 40 years. Nodular was still the commonest type but its proportion had dropped significantly compared with the first decade, with a corresponding increase in SSM and acral types. CONCLUSIONS: Over a 20-year period there was little change in the absolute number of patients presenting with thick melanoma each year, though these form a diminishing proportion of the rising number of total melanomas. This thick melanoma group is characterized by an increasingly older age group and a changing type profile, nodular and SSM being the most common types. This work suggests that the resources currently directed at public and professional education on melanoma are having no effect on this group of patients and that alternative strategies may need to be considered. PMID- 15656810 TI - Prevalence of atopic dermatitis in Japanese elementary schoolchildren. AB - BACKGROUND: Although there have been several reports on the prevalence of atopic dermatitis (AD) in Japanese schoolchildren based on questionnaires, there has been no nation-wide study of the frequency of this condition diagnosed by dermatologists in regular health check-ups of schoolchildren. OBJECTIVES: The objective of this work was to evaluate precisely the prevalence of AD in elementary schoolchildren in Japan based on regular health check-ups by dermatologists. METHODS: In 2001/2, elementary schoolchildren: first graders (age 6-7 years) and sixth graders (age 11-12 years) were examined by dermatologists in eight prefectures of Japan (Hokkaido, Iwate, Tokyo, Gifu, Osaka, Hiroshima, Kochi and Fukuoka). In each prefecture, public elementary schools were randomly selected from urban and rural districts. We planned to examine about 700 schoolchildren in each of urban first, urban sixth, rural first and rural sixth grades from the eight areas, a total of 22 400 children (700 x 4 x 8). AD was diagnosed by the dermatologists based on the Japanese Dermatological Association criteria for the disease. RESULTS: The point prevalence of AD was 11.2% overall (2664 of 23 719) ranging from 7.4% (Iwate) to 15.0% (Fukuoka) in the eight areas. Seventy-four per cent, 24%, 1.6% and 0.3% of those afflicted were in the mild, moderate, severe and very severe groups, respectively. Overall, the prevalence of first graders was slightly higher than that of sixth graders (11.8% vs. 10.5%, P < 0.01). There was no apparent difference in prevalence between urban and rural districts, or between boys and girls. CONCLUSIONS: The prevalence of AD in Japanese elementary schoolchildren was about 10%, three-quarters of those being mildly affected. This is the first nation-wide study made of Japanese elementary schoolchildren examined by dermatologists to evaluate the frequency of AD. PMID- 15656811 TI - Ultraviolet B irradiation of human skin induces an angiogenic switch that is mediated by upregulation of vascular endothelial growth factor and by downregulation of thrombospondin-1. AB - BACKGROUND: We have previously demonstrated that skin-specific overexpression of the endogenous angiogenesis inhibitor thrombospondin (TSP)-1 prevented chronic ultraviolet (UV) B-induced angiogenesis, inflammatory cell infiltration and cutaneous photodamage in mice. OBJECTIVES: To elucidate the mechanisms by which acute UVB-induced angiogenesis induces dermal damage, and to study the molecular regulation of acute UVB-induced angiogenesis in human skin. METHODS: We subjected five healthy volunteers to acute UVB irradiation (2 minimal erythema doses) and performed histological analysis at 48 h after UVB irradiation. RESULTS: Histology revealed epidermal hyperplasia, infiltration of elastase-producing neutrophils and elastin fibre damage. Immunohistochemistry for CD31 demonstrated pronounced angiogenesis with a significant increase in both vascular density and vessel size, associated with increased endothelial cell proliferation. Whereas constitutive expression of TSP-1 but only weak expression of vascular endothelial growth factor (VEGF) were detected in normal human epidermis, pronounced downregulation of TSP-1 and upregulation of VEGF were observed in epidermal keratinocytes after acute UVB irradiation. These findings were confirmed by quantitative reverse transcription-polymerase chain reaction analysis after UVB irradiation of cultured HaCaT keratinocytes in vitro. CONCLUSIONS: Together, these data indicate that a disruption of the balance between VEGF and TSP-1 expression leads to a UVB-induced angiogenic switch, facilitating the infiltration of elastase-producing leucocytes and cutaneous photodamage. PMID- 15656812 TI - Imiquimod cream 5% for recalcitrant cutaneous warts in immunosuppressed individuals. AB - BACKGROUND: Viral warts may cause significant morbidity in individuals unable to mount an adequate T-helper 1 cell-mediated immune response to human papillomavirus. Imiquimod is a potent inducer of antiviral cytokine activity which has shown significant efficacy in the treatment of genital warts. Similar efficacy in cutaneous warts is not yet established. OBJECTIVES: To assess the response of persistent cutaneous warts to 5% imiquimod cream in immunosuppressed individuals. METHODS: Fifteen immunosuppressed patients with warts on the hands and/or feet present for more than 18 months, which had failed to respond to a minimum of 12 weeks of topical salicylic acid and four cycles of cryotherapy, were recruited. Imiquimod 5% cream was applied in an open label, right vs. left comparison study for 24 weeks (three times weekly for 8 weeks, daily for 8 weeks, then daily with occlusion for 8 weeks). RESULTS: Twelve (80%) patients completed the study protocol. Benefit was seen in five patients [36% in the intent-to-treat analysis (14 patients)], including more than 30% clearance of warts in three patients and reduction in overall size of warts in two further cases. Local skin reactions occurred in four (29%) patients and were usually mild. A transient rise in creatinine (11-29% above baseline) was measured in three renal transplant recipients, but we did not consider that this was related to imiquimod exposure. CONCLUSIONS: This is the first controlled study to assess therapeutic efficacy of topical 5% imiquimod cream in persistent warts associated with immunosuppression. It provides preliminary evidence that topical imiquimod may benefit a subgroup of immunosuppressed patients with recalcitrant cutaneous warts. PMID- 15656813 TI - Topical corticosteroids for atopic eczema: clinical and cost effectiveness of once-daily vs. more frequent use. AB - BACKGROUND: Topical corticosteroids remain the mainstay of treatment for atopic eczema, yet there is uncertainty over the frequency of their use in terms of clinical and cost effectiveness. OBJECTIVES: To assess the clinical and cost effectiveness of once-daily vs. more frequent use of same-potency topical corticosteroids in atopic eczema. METHODS: A systematic review of the clinical and cost-effectiveness literature was undertaken, together with a cost minimization analysis. RESULTS: The review identified a sparse literature, comprising one previous systematic review and 10 randomized controlled trials (RCTs). No published cost-effectiveness studies were identified. RCTs were focused on potent topical corticosteroids (eight RCTs), with no trials (RCTs/controlled clinical trials) identified on mild potency products. There was broad heterogeneity in trial methods, and therefore we considered outcomes according to: (i) at least a good response or 50% improvement, and (ii) eczema rated as cleared or controlled. Studies found little difference between once daily and more frequent use of topical corticosteroids. The literature on moderately potent and potent corticosteroids offered no basis for favouring once daily or more frequent use, although some significant differences favouring twice daily treatment were identified. One RCT on very potent products favoured three times daily use on the basis of clinical response, but reported no difference in the numbers with at least a good response. Given the similar outcomes seen in clinical effectiveness a cost-minimization approach was adopted to consider cost effectiveness, in order to identify the least-cost option. However, cost minimization analysis proved complex due to wide variations in product price, with the relative cost of product comparisons by frequency proving the most important factor in determining the least-cost alternative. CONCLUSIONS: This review has not identified any clear differences in outcomes between once-daily and more frequent application of topical corticosteroids. We would encourage prescribing clinicians to consider the once-daily use of topical corticosteroids when making treatment decisions for patients with atopic eczema. However, we find that the literature on clinical effectiveness is limited and a broader understanding of compliance and phobia associated with topical steroids is needed to inform on this issue. PMID- 15656815 TI - Large-scale DNA microarray analysis of atopic skin lesions shows overexpression of an epidermal differentiation gene cluster in the alternative pathway and lack of protective gene expression in the cornified envelope. AB - BACKGROUND: Atopic dermatitis (AD)-specific genes have not yet been clarified. Objectives To identify gene expression specific to active atopic skin lesions. METHODS: We analysed 23,000 genes in skin biopsy samples from 17 patients with AD and four normal controls using Affymetrix oligonucleotide arrays. RESULTS: Four of the 10 genes with the greatest differences in expression between patients and controls, S100A8 and S100A7 (upregulated), and loricrin and filaggrin (downregulated), were epidermal differentiation genes located on 1q21, a locus previously reported to have a genetic linkage with AD. CONCLUSIONS: Our results, showing downregulation of the cornified envelope genes and upregulation of the alternative keratinization pathway, are the first to suggest abnormal epidermal differentiation and defective defences as key abnormalities in AD. PMID- 15656814 TI - Detection of 1733insC mutations in an Asian family with Birt-Hogg-Dube syndrome. AB - BACKGROUND: Birt-Hogg-Dube syndrome (BHD) is a rare autosomal dominant genodermatosis characterized by skin tumours, including multiple fibrofolliculomas, trichodiscomas and acrochordons. BHD patients also may suffer from associated renal and colonic carcinomas. The defective gene in BHD has been recently identified and is suspected of being a tumour suppressor gene. Several mutations of the BHD gene have been reported only in Caucasian patients. OBJECTIVES: This study reports the first Asian family that has been demonstrated to carry a BHD mutation. PATIENTS/METHODS: The proband was a 26-year-old Japanese man with multiple asymptomatic, soft skin-coloured papules on his face, neck and trunk, which were clinically thought to be acrochordon. His father was also affected. Histopathologically, the papules revealed a fibrofolliculoma that had a circumscribed proliferation of fibroblasts and collagen fibres surrounding an abnormal hair follicle. RESULTS: Mutational analysis of the BHD gene of the proband and the father detected 1733insC, a cytosine insertion mutation in an eight-cytosine tract (nucleotides 1733-1740) in exon 11. Analysis of fibrofolliculoma in the proband showed heterozygous 1733insC mutation, suggesting the absence of loss of heterozygosity. Interestingly, previous mutational analysis in Caucasian patients revealed that both1733insC and 1733delC mutations were hot spots. CONCLUSIONS: This study is the first to find the same hot-spot 1733insC mutation in Asian kindred. The mutations in this polycytosine tract may have a wide, global distribution despite their arising from a different ethnic background. PMID- 15656816 TI - Toxic epidermal necrolysis in a premature infant of 27 weeks' gestational age. AB - Toxic epidermal necrolysis (TEN) is very rare in the newborn period. So far, three cases of TEN in newborns have been reported worldwide. We report a premature infant of 27 weeks' gestational age with TEN at 4 weeks of age. Sepsis treated by an antibiotic combination regimen preceding the TEN was a common feature of all four cases. In our patient, coagulase-negative staphylococci could be identified by blood culture, whereas the previously reported patients suffered from Klebsiella pneumoniae sepsis or Escherichia coli sepsis. Possibly, the uniform association with septic infection in the cases of TEN in the neonatal period might hint at a causal association, thus differentiating it from TEN in older children or adults. PMID- 15656817 TI - Does topical tacrolimus induce lentigines in children with atopic dermatitis? A report of three cases. AB - Three children with severe atopic dermatitis were noted at routine follow-up to have developed multiple small pigmented macules during long-term therapy with topical tacrolimus 0.1% (Protopic, Fujisawa). Representative lesions in two of the three cases were confirmed histologically as simple lentigines. The focal distribution of lentigines to sites of tacrolimus use, and the temporal association between use of tacrolimus and development of lesions, suggest that topical tacrolimus is of direct aetiological relevance to their development. Careful long-term follow-up will be required to assess the clinical implications of these findings and whether they represent an increase in risk for melanocytic neoplasia. PMID- 15656818 TI - A new type of erythrokeratoderma. AB - We describe a Dutch man suffering from a previously undescribed erythrokeratoderma associated with palmoplantar keratoderma and circular constrictions of the fingers. No mutations were identified in the genes encoding loricrin, connexin 26, 30, 30.3, 31 and 31.1, and ARS/complex B. There are some similarities between the disorder described here and other palmoplantar keratodermas and erythrokeratodermas, but assignment to a particular disease category is not possible. Hence we propose that we have delineated a novel type of keratoderma. PMID- 15656819 TI - Netherton syndrome: report of two Taiwanese siblings with staphylococcal scalded skin syndrome and mutation of SPINK5. AB - Netherton syndrome (NS) is a severe autosomal recessive ichthyosis. It is characterized by congenital ichthyosiform erythroderma, trichorrhexis invaginata, ichthyosis linearis circumflexa, atopic diathesis and frequent bacterial infections. Pathogenic mutations in SPINK5 have recently been identified in NS. SPINK5 encodes lymphoepithelial Kazal-type-related inhibitor (LEKTI), a new type of serine protease inhibitor involved in the regulation of skin barrier formation and immunity. We report two Taiwanese brothers with NS. The patients had typical manifestations of NS with an atopic diathesis and recurrent staphylococcal infections, including staphylococcal scalded skin syndrome (SSSS) since birth. Horny layers were obtained by skin surface biopsy for electron microscopy from lesional skin of both patients and from normal controls. All 33 exons and flanking intron boundaries of SPINK5 were amplified for direct sequencing. The ultrastructure of the stratum corneum (SC) was characterized by premature degradation of corneodesmosomes (CDs) with separation of corneocytes. A homozygous 2260A --> T (K754X) mutation of SPINK5 was found in both patients. Staphylococcal exfoliative toxin A (ETA) is a serine protease capable of cleaving desmoglein 1, an important adhesive molecule of CDs, and can cause separation of the SC, resulting in SSSS. The premature degradation of CDs found in our patients may be attributable to insufficient LEKTI, and possibly also to colonization/infection of ETA-producing Staphylococcus aureus. Mechanisms involved in the pathogenesis of the skin barrier defect in NS are proposed. Further study is needed to prove this hypothesis. PMID- 15656820 TI - Cannabis arteritis. AB - The main causes of arteriopathy in young patients include drugs, metabolic diseases, pseudoxanthoma elasticum and Buerger's disease. Arteritis due to Cannabis indica was first reported in 1960, and the role of this drug as a risk factor for arteritis was confirmed in several subsequent publications. A 38-year old smoker with no previous contributory medical history except for long-standing cannabis abuse developed a dry necrotic lesion of the left big toe. Imaging examinations revealed proximal arteriopathy of the lower limbs that predominated on the left side. He had no atherogenic or thrombogenic risk factors, and no signs of pseudoxanthoma elasticum were found. Remarkably, the development of arteritis paralleled cannabis abuse. The course was slowly favourable after weaning from the drug, vasodilator treatment and hyperbaric oxygen therapy. Despite some subtle clinical differences (more proximal than distal involvement), cannabis arteritis may be considered as a particular form of Buerger's disease, where cannabis, along with tobacco, seems to cause arterial lesions. Along with the noxious effects of cannabis on vessels, a role for contaminating arsenic is also possible. Cannabis arteritis is not widely known, but may prove not to be so rare if one considers consumption of cannabis besides that of tobacco. PMID- 15656821 TI - Bowen's disease on the sole: p16INK4a overexpression associated with human papillomavirus type 16. AB - We report a case of Bowen's disease on the sole presenting clinically as an exophytic, blackish-grey, verrucous tumour, and showing human papillomavirus (HPV) type 16 on analysis with polymerase chain reaction. Positive stains for HPV particles by immunohistochemical analysis were limited to several cell nuclei at the upper stratum Malpighii. However, all the tumour cells in the epidermis exhibited strong and diffuse nuclear and cytoplasmic stains for the tumour suppressor protein p16INK4a. We speculate that dysregulation of the retinoblastoma/p16INK4a pathway may be involved in the pathogenesis of the lesion, and p16INK4a overexpression might serve as a useful surrogate marker for identifying Bowen's disease harbouring high-risk types of HPV infection. PMID- 15656822 TI - A Korean case of oculocutaneous albinism type IV caused by a D157N mutation in the MATP gene. PMID- 15656823 TI - Sentinel node biopsy in the management of primary cutaneous melanoma. PMID- 15656824 TI - Dermatologists should exert a greater influence over sentinel lymph node biopsy for melanoma in the U.K. PMID- 15656825 TI - Angiokeratomas, Fabry disease and enzyme replacement therapy: still a challenge. PMID- 15656827 TI - Diffuse cutaneous mastocytosis treated with psoralen photochemotherapy: case report and review of the literature. PMID- 15656828 TI - Localized scleroderma: response to occlusive treatment with tacrolimus ointment. PMID- 15656830 TI - Topical use of tacrolimus and squamous cell carcinoma on the penis. PMID- 15656829 TI - Meyerson's phenomenon induced by interferon-alfa plus ribavirin in hepatitis C infection. PMID- 15656831 TI - Unilateral remission of psoriasis following traumatic nerve palsy. PMID- 15656832 TI - Pimecrolimus 1% cream for perianal atopic dermatitis. PMID- 15656833 TI - Safety and efficacy study on etanercept in patients with plaque psoriasis. PMID- 15656834 TI - Pioglitazone: a promising therapy for psoriasis. PMID- 15656835 TI - X-linked recessive ichthyosis in a girl: strategy for identifying the causal mechanism. PMID- 15656836 TI - Follicular cutaneous T-cell lymphoma: beneficial effect of isotretinoin for persisting cysts and comedones. PMID- 15656837 TI - Chemical exposures other than arsenic are probably not important risk factors for squamous cell carcinoma, basal cell carcinoma and malignant melanoma of the skin. PMID- 15656842 TI - Hospitalized elders and family caregivers: a typology of family worry. AB - OBJECTIVE: This qualitative study explored the kinds of worry that family caregivers experience when their older relatives are hospitalized. BACKGROUND: Little is known about what kinds of worries family caregivers may have in association with the hospitalizations of older relatives. An understanding of the different patterns of family worry may help health care teams intervene more effectively to meet family caregiver's needs by reducing their anxiety. DESIGN: A qualitative descriptive design with Loftland and Loftland (1984) approach for the study of a phenomenon occurring in a social setting was used. METHOD: A purposeful sample of 10 participants was obtained that included six family caregivers and four nurses. Participants were recruited from two hospitals in the northwest US. Intensive interviews and participant observations were used for data collection, and Loftland and Loftland's (1984) qualitative approach was used for data analysis. RESULTS: Family worry was defined as family caregivers' felt difficulty in fulfilling their roles because of worry. Four categories of family worry were identified as a result of this study: (i) worry about the patient's condition; (ii) worry about the patient's care received from the health care team; (iii) worry about future care for the patient provided by the family caregiver; and (iv) worry about finances. CONCLUSIONS: The findings of this pilot study provide nurses with the initial knowledge of the typology of family worry associated with elderly relatives' hospitalizations. RELEVANCE TO CLINICAL PRACTICE: The findings of this study may sensitize the nurses to more precisely evaluate family caregivers' worry about their hospitalized elders and provide more effective nursing interventions to improve outcomes of both patients and their family caregivers. PMID- 15656843 TI - Work satisfaction and dissatisfaction--caregivers' experiences after a two-year intervention in a newly opened nursing home. AB - AIMS AND OBJECTIVES: The aim of the study was to investigate, from the narratives of nine enrolled nurses and one nurses' aide directly involved in patient care, the deeper meaning of work satisfaction and dissatisfaction when working with the older people. BACKGROUND: Both nationally and internationally, there is little research documented regarding the working situation of the enrolled nurses and nurses' aides who make up the majority of care for older people today. With this in mind, it is important to focus on how these occupational groups experience their work with the older residents in municipal care, following a two-year intervention. DESIGN: The study is part of a larger longitudinal study, with a quasi-experimental design within the municipal system of care for older people in Sweden. The investigation was carried out following a two-year intervention, which included: education, support and clinical supervision. METHOD: The interviews were performed 12 and 24 months after start of the intervention and were analysed with a phenomenological-hermeneutic method inspired by Ricoeur's philosophy. RESULTS: The findings from these narratives illustrated a change compared with the findings from the first interviews, when the nursing home had just opened. There was a shift from a dominance of dissatisfaction with work, to a dominance of work satisfaction and this was expressed in the following themes: experience of a changed perspective, experience of open doors, and experience of closed doors. Each theme emerged from several different subthemes and each subtheme that had been expressed in the caregivers' narratives was interpreted. CONCLUSIONS: The study shows that the caregivers' experience of work satisfaction in the workplace exceeded their experience of dissatisfaction and that the intervention, consisting of: education, support, and supervision might have facilitated this positive development where the older residents were prioritized. It also shows that communication and understanding between management and staff had increased as the nursing home had opened. RELEVANCE TO CLINICAL PRACTICE: The findings can be used to help to prevent work dissatisfaction, and thereby increase work satisfaction for caregivers working in nursing homes. PMID- 15656844 TI - Health promoting hospitals: the role and function of nursing. AB - BACKGROUND: In 1986, the World Health Organisation's (WHO) Ottawa Charter for Health Promotion sought to create a framework that conveyed the notion of capacity building, related to specific settings, and a structured process for health promotion action. It provided the platform from which the health promoting hospital movement was later launched. Nearly two decades on, the health promoting hospital (HPH) movement has grown considerably and continues to expand, against the backdrop of having to adapt to the changing needs and demands of clients and the evolving social context of their communities. Many nurses, it is argued here, are often unaware of health promoting hospitals concepts or, when they are, do little to contribute to them. METHOD: A critical review of the available health promoting hospital and related literature has been conducted for discussion. FINDINGS: The literature revealed that hospitals are being urged to reject practices based purely on health-limiting and limited biomedical frameworks, in favour of moving towards models based on health promoting hospitals and public health-orientated hospitals. This requires radical reform that focuses on the social and health policy context of organizations, the socio-political empowerment of its employers and clients, and the personal/collective positive health and welfare of its employers and clients. Many health service agencies are beginning to emphasize population health within communities as part of a concerted move away from an original primary focus on acute inpatient hospital based service provision. CONCLUSION: Hospitals need to adapt and expand their efforts to focus on health promotion activities, in collaboration with the ever widening community networks of health and social agencies. This requires the commitment of all health care professional groups. Nurses who practice in the hospital setting should be aiming to initiate and promote radical health promotion reform as set out in the health promoting hospital movement. This paper argues that nursing per se could be making much larger inroads and efforts to affect and implement wide-ranging health promotion activities in hospital organizations. Nurses should view the HPH movement as another opportunity truly to embrace evolving broad-based health promotion concepts, as a means to forge and own their own health agenda, and also as a means to move beyond the traditional reliance of a limited health education role. RELEVANCE TO CLINICAL PRACTICE: Hospitals and their employees must be seen to advocate, support and implement wide-reaching social and community-based reform, as part of a necessary commitment to 'seamless' health care provision. The health promoting hospital movement represents a collective vehicle for enabling such activities. If nurses wish to be at the forefront of current health service strategies they must be seen to embrace the radical health promotion reforms that are emerging from the current literature and put forward in this article. PMID- 15656845 TI - SARS (severe acute respiratory syndrome): reflective practice of a nurse manager. AB - AIMS AND OBJECTIVES: This paper describes the reflective practice of a nurse manager in Hong Kong in supporting frontline nurses to overcome the crisis of SARS. BACKGROUND: SARS infection was a crisis for everyone in endemic areas because of its threat to physical and emotional health. Hong Kong was the second leading endemic area in the world. Inadequate supplies of protective devices and the death of a nurse infected with SARS triggered nurses' negative emotions. METHODS: A model of structured reflection was adopted to examine one's practice. A problem-solving model for crisis intervention was integrated into the reflective stage of structured reflection. RESULTS: Promotion of nurses' safety and emotional stability were the major goals in handling the crisis. Strategies were employed including self-awareness, empowerment and team building, information sharing, provision of personal protective equipment and emotional support for frontline nurses. CONCLUSIONS: SARS infection threatens the physical and emotional health of nurses. From a positive perspective, such a crisis created an opportunity to learn and grow in terms of ethical, personal and aesthetic arenas. RELEVANCE TO CLINICAL PRACTICE: SARS epidemic raised worldwide attention and challenged the Hong Kong's health care system. Reflective practice is useful to guide and examine nurses' professional action during the crisis, and to put the experience into a learning perspective. PMID- 15656846 TI - Patient participation in nursing care: an interpretation by Swedish registered nurses. AB - AIMS AND OBJECTIVES: The aim of this study was to clarify Registered Nurses' understanding of patient participation in nursing care. Objectives were to investigate ward nurses' interpretation of the elements of patient participation and its implementation. BACKGROUND: One goal of health care is patient participation. Registered Nurses must encourage the participation of their patients in nursing care situations. A right for patients to make their own informed choices is laid down in Swedish laws and national and local directives. The concept of patient participation can be interpreted differently and is thus difficult to implement and evaluate. Current nursing literature provides little clarity of the elements and processes of patient participation and conceptual definitions differ. DESIGN AND METHODS: A qualitative approach was applied and the Grounded Theory method used. Thirty-one Registered Swedish Nurses described the meaning of patient participation and its implementation in nursing care. Seven focus group interviews with nurses providing inpatient somatic care were carried out in five hospitals in west Sweden. RESULTS: Four properties, describing nurses' approaches and procedures, constitute patient participation. The properties are: interpersonal procedure, therapeutic approach, focus on resources, opportunities for influence. These properties were considered crucial for patient participation in a nursing care context. CONCLUSIONS AND RELEVANCE TO CLINICAL PRACTICE: The results clarify the concept of patient participation from a nursing perspective. A comprehensive description of important factors for patient participation could be made on the basis of nursing experience. This comprehensive description can be used in nursing care practice and its evaluation. The developed categories can be used in nursing care documentation of how patient participation is realized. PMID- 15656847 TI - Telling tales: a qualitative exploration of how children's nurses interpret work with unaccompanied hospitalized children. AB - AIMS AND OBJECTIVES: To explore how qualified children's nurses define, interpret and give meaning to their work with unaccompanied children in hospital. BACKGROUND: Unaccompanied children are amongst the most vulnerable of the hospital population yet there is little insight into the strategies nurses use to work with them. DESIGN: A qualitative study. METHODS: Tape recorded unstructured interviews with four qualified children's nurses. The interviews focused on stories from the participants' experiences of working with unaccompanied hospitalized children. Transcribed interviews were analyzed for narrative structure to expose how the participants interpreted and gave meaning to their experiences. FINDINGS: The participants' interpretations of the child 'alone' advance previous understandings. The stories unfold into interpreted meanings that shed light on the participant's experiences of sadness and distress. RELEVANCE TO CLINICAL PRACTICE: The strategy of differentiating between being a nurse and being a parent emerges as an important strategy in interpreting professional boundaries. The conventional meaning of 'emotional distancing' is further advanced by the participants' resiliency at remaining emotionally available to the children and parents in their stories. Further research into the emotional aspects of working with children in hospital is warranted. PMID- 15656848 TI - Effects of a relaxation breathing exercise on fatigue in haemopoietic stem cell transplantation patients. AB - AIM: Our purpose was to investigate the effect of a relaxation breathing exercise on fatigue in allogenic haemopoietic stem cell transplantation patients. BACKGROUND: Exercise is a promising approach in ameliorating the fatigue associated with cancer and its treatment. DESIGN: A randomized design with control, experimental groups being assessed pre- and postintervention. METHODS: Thirty-five patients were randomly selected, with 18 assigned to an exercise group and 17 to a control group. The exercise intervention lasted for 30 minutes every day for six weeks. It consisted of physical exercise combined with relaxation breathing exercise. Fatigue was measured by the revised version of the Piper Fatigue Scale. RESULTS: The exercise group had a greater decrease in fatigue than the control group. CONCLUSION: These findings indicate that a relaxation breathing exercise would improve fatigue in allogenic haemopoietic stem cell transplantation patients. RELEVANCE TO CLINICAL PRACTICE: A relaxation breathing exercise by a nurse can improve fatigue in allogenic haemopoietic stem cell transplantation patients. PMID- 15656849 TI - Malignant wounds in women with breast cancer: feminine and sexual perspectives. AB - AIMS AND OBJECTIVES: . The aim of this article is to investigate the way malignant fungating wounds affect femininity, sexuality and daily life in women with progressive breast cancer. Malignant wounds occur when the underlying localized tumour filtrates into the skin, blood capillaries and/or lymph vessels. DESIGN: The study was a prospective and exploratory intervention study. The intervention was tested on 12 consecutively selected women with progressive breast cancer and malignant fungating wounds. METHODS: The 12 women participated in a 4-week wound care intervention programme that comprised evidence-based wound care principles and psychosocial support. Data were generated by means of interviewing prior to and following the intervention period. RESULTS: The women described how malodorous and oozing wounds trigger anxiety about seepage, prevent them from wearing feminine attire and cause them to suppress the need for physical closeness and sexual activity. The results showed that by using modern wound care products, the patients could be secured against seepage and odour. The women experienced a sense of comfort, were able to dress again as they wished, no longer felt caged in and isolated and were given a sense of freedom which they had not felt for a long time. CONCLUSIONS: The intervention succeeded in increasing breast cancer patients' psychosocial well-being and reducing social isolation. RELEVANCE TO CLINICAL PRACTICE: There is a lack of standard methods of practice for care of malignant wounds in women with breast cancer. This article describes an intervention for wound care which could improve daily live in women with progressive breast cancer. PMID- 15656850 TI - Ulcerative colitis: factors affecting difficulties of life and psychological well being of patients in remission. AB - BACKGROUND: Ulcerative colitis is a chronic disease of unknown aetiology characterized by alternating periods of remission and relapses. The difficulties in the patients with ulcerative colitis daily life mostly arise from symptoms associated with bowel inflammation but there are many patients who have difficulties despite being in the remission phase. This study was conducted to elucidate factors that influence perception of difficulties of life and psychological well being of patients with ulcerative colitis in remission. METHODS: A questionnaire survey was carried out in 72 outpatients with ulcerative colitis in remission. The perception of difficulties of life was assessed using the scale developed by Tanaka et al. Psychological well being was assessed using the Japanese version of the 'profile of mood states'. Physical condition, demographic attributes and psychosociological states were also investigated as related factors. RESULTS: There was no outstanding aspect of psychological well being, but a relatively large number of patients perceived a 'decline of vitality or vigour' despite being in the remission phase. In the presence of irritable bowel syndrome-like symptoms, the scores for 'difficulties of life in society' or 'difficulties concerned with bowel movements' were high. Scores for 'decline of vitality or vigour' were high when the emotive coping scores were high, social support was lower, disease durations were short and relapses occurred more than once. When the emotive coping scores were high, psychological well being was not fine. RELEVANCE TO CLINICAL PRACTICE: Strategies need to be developed to allow patients to recover and maintain their strength in the remission phase. Further, the strategies should take the above factors into consideration. PMID- 15656851 TI - Living with a stoma: a review of the literature. AB - AIMS AND OBJECTIVES: This systematic review looks at the psychological and social impact of stoma surgery on peoples' lives. BACKGROUND: The formation of a stoma can have a negative effect on a person's quality of life and affect lifestyle in a number of ways. METHODS: The review examines nursing literature from 1990 to date and focuses on how stoma patients' lives are affected by the presence of a stoma. It informs nursing practice so nurses are better able to help individuals improve, maintain or recover their health following ostomy surgery. RESULTS: The findings of the review indicate that stoma surgery can impact on individuals' lives in many different ways. RELEVANCE TO CLINICAL PRACTICE: Knowledge of the problems stoma patients can experience can help nurses plan care in an individualized way. Recommendations for nursing practice and future research are made. PMID- 15656852 TI - Assessing competency in nursing: a comparison of nurses prepared through degree and diploma programmes. AB - AIMS AND OBJECTIVES: The present study aimed to investigate the competencies of qualifiers from three-year degree and three-year diploma courses in England at one, two and three years after qualification. BACKGROUND: The provision of three year preregistration nursing degrees in the UK has increased in recent years and in many colleges degrees are offered alongside the existing three-year diploma courses. Yet little is known about the relationship between these different education programmes and the competence of qualifiers. METHODS: A cross-sectional survey design was employed to make comparisons of both self-reported and line manager-rated competencies of graduate and diplomate nurses who had qualified up to three years previously. INSTRUMENTS: A revised version of the Nursing Competencies Questionnaire was used to measure both overall competence and also eight specific nursing competencies. A shortened version of this scale was also used to assess internal consistency across measures. Two additional competencies, research awareness and policy awareness, were also measured. RESULTS: Structural equation modelling found very little difference in the overall competence and specific competencies of graduates and diplomates. Where differences were found in the self-report data, diplomates scored more highly than graduates in the constructs of planning and social participation; however, these differences became non-significant when background variables were controlled for. LIMITATIONS: The findings are interpreted with caution due to the size of differences, the size of some of the samples of respondents and the developmental stage of the instrument used. CONCLUSIONS: It does not appear that graduates and diplomates in England differ in their level of competence to any great extent as measured by the Nursing Competencies Questionnaire. Areas of further work are discussed in the light of the findings. RELEVANCE TO CLINICAL PRACTICE: While this may alleviate concerns about clinical disparities between the two groups, it raises questions about the proposed benefits to nursing of three-year preregistration degrees in terms of quality of care during the first three years of qualification. PMID- 15656853 TI - Perinatal nursing education for single-room maternity care: an evaluation of a competency-based model. AB - AIMS AND OBJECTIVES: To evaluate the success of a competency-based nursing orientation programme for a single-room maternity care unit by measuring improvement in self-reported competency after six months. BACKGROUND: Single-room maternity care has challenged obstetrical nurses to provide comprehensive nursing care during all phases of the in-hospital birth experience. In this model, nurses provide intrapartum, postpartum and newborn care in one room. To date, an evaluation of nursing education for single-room maternity care has not been published. DESIGN: A prospective cohort design comparing self-reported competencies prior to starting work in the single-room maternity care and six months after. METHODS: Nurses completed a competency-based education programme in which they could select from a menu of learning methods and content areas according to their individual needs. Learning methods included classroom lectures, self-paced learning packages, and preceptorships in the clinical area. Competencies were measured by a standardized perinatal self-efficacy tool and a tool developed by the authors for this study, the Single-Room Maternity Care Competency Tool. A paired analysis was undertaken to take into account the paired (before and after) nature of the design. RESULTS: Scores on the perinatal self efficacy scale and the single-room maternity care competency tool were improved. These differences were statistically significant. CONCLUSIONS: Improvements in perinatal and single-room maternity care-specific competencies suggest that our education programme was successful in preparing nurses for their new role in the single-room maternity care setting. This conclusion is supported by reported increases in nursing and patient satisfaction in the single-room maternity care compared with the traditional labour/delivery and postpartum settings. RELEVANCE TO CLINICAL PRACTICE: An education programme tailored to the learning needs of experienced clinical nurses contributes to improvements in nursing competencies and patient care. PMID- 15656854 TI - Sleep disturbance: the patient care activities applied at the night shift in the intensive care unit. AB - AIMS AND OBJECTIVES: The aim of the study was to determine the frequency and types of nursing activities on sedated and non-sedated patients in the intensive care unit at night shift. BACKGROUND: The complex intensive care unit and nursing activities has been described as an environment in which patients enter in a physiological crisis. This is even more so when the patient is unable to communicate or heavily sedated or unable to move. Nursing activities, fear and apprehension cause low morale and sleep disturbance. METHOD: The study was carried out on a descriptive basis in the Surgical Intensive Care Unit. Thirty sedated and 30 non-sedated patients to whom the nursing care was applied during 19.00-07.00 hours for three consecutive nights were chosen with simple random sampling. Data were gathered by means of retrospective examination of the standard nursing chart notes about the 60 patients for three consecutive nights. Data were analysed using percentage calculations and chi-square test. RESULTS: The results demonstrate that patient records indicated a mean of 51 interactions per patient per night. Nursing activities were more frequent between 02.00 and 05.00 hours. It was detected that nursing activities such as mouth and eye care, decubitus ulcer care, change of dressing, bed bath, catheter change for three night shifts were more frequently between 24.00 and 05.00 hours. Mechanical ventilation control and endotracheal suctioning were more frequently performed on sedated patients than on non-sedated patients (P < 0.05). CONCLUSION: These findings of this study indicate that the nursing activities in the intensive care unit were focused on the activities, which will maximize what is seen to be physiological stability. RELEVANCE TO CLINICAL PRACTICE: This study has provided further data that identify the activities of nurses working in the intensive care unit at night shift. We hope to contribute to the development of the sleep protocol in surgical intensive care units. PMID- 15656855 TI - Sleep quality in hospitalized patients. AB - AIMS AND OBJECTIVES: The objective of this study was to evaluate and compare sleep quality of the hospitalized patients and matched healthy controls. BACKGROUND: Although the functions of sleep are not clearly understood, it is generally accepted that it is necessary for the maintenance of good health. Hospitalized patients' sleep may not be refreshing or restorative. The reasons for this can be categorized into three groups: environmental, physiological and psychological. DESIGN AND METHODS: This research was conducted at the Cumhuriyet University Hospital in Turkey. One hundred and fifty hospitalized patients (psychiatry = 50; orthopaedic + general surgery + cardiovascular surgery + urology = 50; internal medicine + chest diseases + infectious diseases + physical therapy and rehabilitation = 50) and 50 healthy controls constituted the sample. The researchers administered to the patient and control groups Sociodemographic Information Form and the Pittsburgh Sleep Quality Index. We compared sociodemographic and illness variables with sleep characteristics. The following statistical analyses were used in order to evaluate the data: variance analysis, Tukey HSD test, Student's t-test, Kruskall-Wallis test. RESULTS: We found that patients in psychiatric ward experienced worse sleep quality than the other patients, worse in female patients than male patients, and worse sleep characteristics in patients than controls. CONCLUSIONS: Health professionals must be educated about sleep and must provide intervention when needed. Relevance to clinical practice. The enhancing of sleep quality accelerates to the recovery from illness. PMID- 15656856 TI - Patient teaching prior to coronary angiography in Hong Kong: a pilot study. PMID- 15656857 TI - Experiences expressed by artificially ventilated patients. PMID- 15656858 TI - Lower limb ulcerations in older people in Hong Kong. PMID- 15656859 TI - Young medical patients' experience of physical restraint: an exploratory study. PMID- 15656860 TI - Commentary on Wiman E & Wikblad K (2004) Caring and uncaring encounters in nursing in an emergency department. Journal of Clinical Nursing 13, 422-429. PMID- 15656862 TI - Commentary on Dempsey J (2004) Falls prevention revisited: a call for a new approach. Journal of Clinical Nursing 13, 479-485. PMID- 15656866 TI - Commentary on Chan S & Levy V (2004) Postnatal depression: a qualitative study of the experiences of a group of Hong Kong Chinese women. Journal of Clinical Nursing 13, 120-123. PMID- 15656864 TI - Hallstrom I & Elander G (2004) Decision-making during hospitalization: parents' and children's involvement. Journal of Clinical Nursing 13, 367-375. PMID- 15656868 TI - Commentary on Skovdahl K et al. (2004) Dementia and aggressiveness: stimulated recall interviews with caregivers after video recorded interactions. Journal of Clinical Nursing13, 515-525. PMID- 15656871 TI - The inflammatory reflex--introduction. AB - Sepsis is the third leading cause of death in the developed world. Despite recent advances in intensive care treatment and the discovery of antibiotics, sepsis remains associated with a high mortality rate. The pathogenesis of sepsis is characterized by an overwhelming systemic inflammatory response that is central to the development of lethal multiple organ failure. This volume of the Journal of Internal Medicine contains three reviews addressing novel aspects of a system we are only beginning to understand - the interactions between the immune and the nervous systems, the 'neuro-immune axis'. Tracey (Nature 2002; 420: 853) recently discovered that the nervous system, through the vagus nerve, can modulate circulating TNF-alpha levels induced by microbial invasion or tissue injury. This cholinergic anti-inflammatory pathway is mediated primarily by nicotinic acetylcholine receptors on tissue macrophages - the pathway leads to decreased production of proinflammatory cytokines. The author reports that treatment with the acetylcholine receptor agonist, nicotine, modulates this system and reduces mortality in 'established' sepsis. Watkins and Maier (J Intern Med 2005; 257: 139) illustrate that pathological pain (induced by inflammation) is not simply a strict neuronal phenomenon, but is a component of the immune response, and is modulated by peripheral immune cells and spinal cord glia cells. This may be of importance for future development of novel drugs for neuropathic pain as well as our understanding of increased risks for infections in anaesthetic skin areas. Blalock (J Immunol 1984; 132: 1067) elucidates the possibility that the immune system actually functions as the sixth sense, sensing microbes and microbial toxins that we cannot see, hear, taste, touch or smell. Activation of the sympathetic nervous system also has predominantly anti-inflammatory effects that are mediated through direct nerve to immune cell interaction or through the adrenal neuro-endocrine axis. PMID- 15656872 TI - The immune system as the sixth sense. AB - One of the truly remarkable discoveries in modern biology is the finding that the nervous system and immune system use a common chemical language for intra- and inter-system communication. This review will discuss some of the pivotal results that deciphered this chemical language. Specifically the nervous and immune systems produce a common set of peptide and nonpeptide neurotransmitters and cytokines that act on a common repertoire of receptors in the two systems. The paper will also review more recent studies that have delineated hardwired and humoral pathways for such bidirectional communication. This is discussed in the context of the idea that the sharing of ligands and receptors allows the immune system to serve as the sixth sense that notifies the nervous system of the presence of entities, such as viruses and bacteria, that are imperceptible to the classic senses. Lastly, this review will suggest ways to apply the newfound knowledge of the sixth sense to understand a placebo effect and to treat human disease. PMID- 15656873 TI - Immune regulation of central nervous system functions: from sickness responses to pathological pain. AB - Classically, the central nervous system (CNS) and the immune system are thought to operate independently of each other. This simplistic view has been corrected in recent years, first with the recognition that the brain dynamically modulates the immune system, and later with the reverse; that is, that the immune system modulates the CNS as well. The evidence that the immune system regulates CNS functions is first reviewed. This immune-to-brain communication pathway triggers the production of a constellation of CNS-mediated phenomena, collectively referred to as 'sickness responses'. These sickness responses are created by immune-to-brain signals activating CNS glia to release glial proinflammatory cytokines. The most recently recognized member of this constellation of changes is enhanced pain responsivity. The hypothesis is then developed that pathological, chronic pain may result from 'tapping into' this ancient survival oriented circuitry, including the activation of immune and glial cells and the release of immune/glial proinflammatory cytokines. This can occur at the level of peripheral nerves, dorsal root ganglia, spinal cord, and likely at higher brain areas. The implications of this model for human chronic pain syndromes and clinical resolution of these chronic pain states are then discussed. PMID- 15656874 TI - Autonomic neural regulation of immunity. AB - The 'cytokine theory of disease' states that an overproduction of cytokines can cause the clinical manifestations of disease. Much effort has been expended to determine how cytokines are regulated in normal health. Transcriptional, translational and other molecular control mechanisms protect the host from excessive cytokine production. A recent discovery revealed an unexpected pathway that inhibits macrophage cytokine production. The inflammatory reflex is a physiological pathway in which the autonomic nervous system detects the presence of inflammatory stimuli and modulates cytokine production. Afferent signals to the brain are transmitted via the vagus nerve, which activates a reflex response that culminates in efferent vagus nerve signalling. Termed the 'cholinergic anti inflammatory pathway', efferent activity in the vagus nerve releases acetylcholine (ACh) in the vicinity of macrophages within the reticuloendothelial system. ACh can interact specifically with macrophage alpha7 subunits of nicotinic ACh receptors, leading to cellular deactivation and inhibition of cytokine release. This 'hard-wired' connection between the nervous and immune systems can be harnessed therapeutically in animal models of inflammatory disease, via direct electrical stimulation of the vagus nerve, or through the use of cholinergic agonists that specifically activate the macrophage alpha7 subunit of the ACh receptor. Autonomic dysfunction has been associated with human inflammatory diseases including rheumatoid arthritis, diabetes and sepsis; whether this dysfunction results from the inflammatory component of these diseases, or is actually an underlying cause, is now less clear. The description of the cholinergic anti-inflammatory now brings to the fore several new therapeutic strategies for inflammatory disease, and suggests that many of these diseases may actually be diseases of autonomic dysfunction. PMID- 15656875 TI - Adiponectin--journey from an adipocyte secretory protein to biomarker of the metabolic syndrome. AB - Adiponectin is an adipocyte-derived hormone that was discovered in 1995. Unlike leptin, which was identified around the same time, the clinical relevance of adiponectin remained obscure for a number of years. However, starting in 2001, several studies were published from different laboratories that highlighted the potential antidiabetic, antiatherosclerotic and anti-inflammatory properties of this protein complex. Methods to measure the protein with high throughput assays in clinical samples were developed shortly thereafter, and as a result hundreds of clinical studies have been published over the past 3 years describing the role of adiponectin in endocrine and metabolic dysfunction. Furthermore, adiponectin research has expanded to include a role for adiponectin in cancer and other disease areas. Although it is an impossible task to summarize the findings from all these studies in a single review, we aim to demonstrate the utility of circulating adiponectin as a biomarker of the metabolic syndrome. Evidence for this relationship will include how decreased levels of plasma adiponectin ('hypoadiponectinaemia') are associated with increased body mass index (BMI), decreased insulin sensitivity, less favourable plasma lipid profiles, increased levels of inflammatory markers and increased risk for the development of cardiovascular disease. Therefore, adiponectin levels hold great promise for use in clinical application serving as a potent indicator of underlying metabolic complications. PMID- 15656876 TI - Effects of long-term treatment with loop diuretics on bone mineral density, calcitropic hormones and bone turnover. AB - BACKGROUND: Loop diuretics (LD) are widely used in the treatment of cardiovascular diseases and disorders with fluid accumulation. LD are known to increase renal calcium losses and may thereby affect calcium homeostasis and bone metabolism. OBJECTIVE: We studied to what extent long-term treatment with LD affects calcium homeostasis and bone metabolism. DESIGN AND SUBJECTS: In a cross sectional design we compared 140 postmenopausal women treated with a LD for more than 2 years with 140 age-matched women not in diuretic therapy. RESULTS: Treatment with LD was associated with significantly increased urinary calcium, plasma parathyroid hormone (PTH) and 1,25-dihydroxyvitamin D levels. Per 40 mg day(-1) of furosemide, urinary calcium was increased by 17% (P < 0.05) and plasma PTH levels were increased by 28% (P = 0.04). Users of LD had a 17% higher body weight (P < 0.001) compared with nonusers. This was due to a 32% higher fat mass (P < 0.001) and a 6% higher lean tissue mass (P < 0.001). Moreover, users of LD had a higher bone mineral density (BMD) at the spine (+7.5%, P < 0.001), hip (+4.8%, P = 0.004), forearm (+3.7%, P = 0.01) and whole body (+2.5%, P = 0.06). However, after adjustment for body weight differences, BMD did not differ between groups. Nevertheless, duration of LD treatment was positively associated with BMD at the spine (P = 0.03) and whole body (P < 0.05). BMD at the spine increases by 0.3% per 1 year of treatment. CONCLUSIONS: The increased renal calcium losses in users of LD are compensated for by a PTH-dependent increase in 1,25(OH)(2)D levels. Thereby calcium balance remains neutral without major effects on bone metabolism. PMID- 15656877 TI - The G-250A substitution in the promoter region of the hepatic lipase gene is associated with the conversion from impaired glucose tolerance to type 2 diabetes: the STOP-NIDDM trial. AB - OBJECTIVES: Dyslipidaemia that includes high levels of triglycerides and low high density lipoprotein cholesterol is a risk factor for type 2 diabetes. Hepatic lipase gene encoding a lipolytic enzyme participating in remodelling of plasma lipoproteins and formation of serum lipid profile is a promising candidate gene for type 2 diabetes. The purpose of the study was to investigate whether the G 250A promoter polymorphism of the LIPC gene predicts the conversion from impaired glucose tolerance (IGT) to type 2 diabetes. SUBJECTS AND DESIGN: Study population comprised of subjects who participated in the STOP-NIDDM trial aiming to investigate the effect of acarbose compared with placebo on the prevention of type 2 diabetes in subjects with IGT. RESULTS: Compared with subjects carrying the G-250G genotype, subjects with the A-250A genotype of the LIPC gene had a 2.35-fold [95% confidence interval (CI) 1.27-4.33, P = 0.006] higher risk of developing type 2 diabetes. Subjects in the placebo group and all women carrying the A-250A genotype had an especially high risk for the conversion to type 2 diabetes [odds ratio (OR) 2.74, 95% CI 1.14-6.61, P = 0.024 and OR 3.70, 95% CI 1.35-10.1, P = 0.011 respectively]. CONCLUSION: The G-250A promoter polymorphism of the LIPC gene is associated with an increased risk of development of type 2 diabetes in high-risk subjects with IGT. Therefore, genes regulating atherogenic dyslipidaemia are promising candidate genes for type 2 diabetes. PMID- 15656878 TI - Favourable association of leg fat with cardiovascular risk factors. AB - OBJECTIVES: To assess the association of trunk and leg fat mass (FM) and leg lean tissue mass (LTM) with conventional cardiovascular risk factors. DESIGN: Cross sectional study. SETTING AND SUBJECTS: We studied 1249 men and 3007 women (age 20 79 years) who attended a research institute for a health checkup between October 1995 and February 2004. MAIN OUTCOME MEASURES: Body mass index, waist circumference (WC), hip circumference (HC), systolic and diastolic blood pressure, total cholesterol, HDL cholesterol, triglycerides, glucose and haemoglobin A(1C) were measured. Trunk FM, leg FM and leg LTM were obtained by dual-energy X-ray absorptiometry. We evaluated the associations between the indices for regional body composition and cardiovascular risk factors, which included hypertension, hypercholesterolaemia, hypo-HDL cholesterolaemia, hypertriglyceridaemia, dyslipidaemia and diabetes mellitus. RESULTS: Increase in WC and HC respectively showed increase and decrease in odds ratios of cardiovascular risk factors. Increase in trunk FM by 1 kg significantly increased the risk of the cardiovascular risk factors with the odds ratios ranging between 1.11 and 1.45. Increase in leg FM by 1 kg significantly decreased the risk with the odds ratios ranging between 0.52 and 0.90, except for the nonsignificant results for hypercholesterolaemia and hypo-HDL cholesterolaemia in men. Odds ratios of 1 kg increase in leg LTM were only significant for dyslipidaemia in men and hypercholesterolaemia in women (both 0.93). CONCLUSIONS: WC and HC showed opposite and independent associations with cardiovascular risk factors. The favourable association of HC was mainly attributable to that of leg FM. PMID- 15656879 TI - Factors associated with development of stroke long-term after myocardial infarction: experiences from the LoWASA trial. AB - OBJECTIVE: To describe factors associated with the development of stroke during long-term follow-up after acute myocardial infarction (AMI) in the LoWASA trial. PATIENTS: Patients who had been hospitalized for AMI were randomized within 42 days to receive either warfarin 1.25 mg plus aspirin 75 mg daily or aspirin 75 mg alone. DESIGN: The study was performed according to the probe design, that is open treatment and blinded end-point evaluation. SETTING: The study was performed in 31 hospitals in Sweden. The mean follow-up time was 5.0 years with a range of 1.7-6.7 years. RESULTS: In all, 3300 patients were randomized in the trial, of which 194 (5.9%) developed stroke (4.2% nonhaemorrhagic, 0.5% haemorrhagic and 1.3% uncertain. The following factors appeared as independent predictors for an increased risk of stroke: age, hazard ratio and 95% confidence interval (1.07; 1.05-1.08), a history of diabetes mellitus (2.4; 1.8-3.4), a history of stroke (2.3; 1.5-3.5), a history of hypertension (2.0; 1.5-2.7) and a history of smoking (1.5;1.1-2.0). Most of these factors were also predictors of a nonhaemorrhagic stroke whereas no predictor of haemorrhagic stroke was found. CONCLUSION: Risk indicators for stroke long-term after AMI were increasing age, a history of either diabetes mellitus, stroke, hypertension or smoking. PMID- 15656881 TI - Diagnostic immunohistochemistry in neuromuscular disorders. AB - Most neuromuscular disorders display only non-specific myopathological features in routine histological preparations. However, a number of proteins, including sarcolemmal, sarcomeric, and nuclear proteins as well as enzymes with defects responsible for neuromuscular disorders, have been identified during the past two decades, allowing a more specific and firm diagnosis of muscle diseases. Identification of protein defects relies predominantly on immunohistochemical preparations and on Western blot analysis. While immunohistochemistry is very useful in identifying abnormal expression of primary protein abnormalities in recessive conditions, it is less helpful in detecting primary defects in dominantly inherited disorders. Abnormal immunohistochemical expression patterns can be confirmed by Western blot analysis which may also be informative in dominant disorders, although its role has yet to be established. Besides identification of specific protein defects, immunohistochemistry is also helpful in the differentiation of inflammatory myopathies by subtyping cellular infiltrates and demonstrating up-regulation of subtle immunological parameters such as cell adhesion molecules. The role of immunohistochemistry in denervating disorders, however, remains controversial in the absence of a reliable marker of muscle fibre denervation. Nevertheless, as well as the diagnostic value of immunocytochemical analysis it may also widen understanding of muscle fibre pathology as well as help in the development of therapeutic strategies. PMID- 15656882 TI - Expression of cell adhesion molecules, CD44s and E-cadherin, and microvessel density in invasive micropapillary carcinoma of the breast. AB - AIMS: Invasive micropapillary carcinoma (IMPC) is a rare variant of ductal carcinoma of the breast and is characterized by high metastatic potential and an aggressive clinical course. This tumour is hence ideal for studying the mechanism underlying tumour biological behaviour, especially metastasis. Cell adhesion molecules, such as CD44 and E-cadherin (Ecad), and angiogenesis are considered important in the invasion and metastasis of tumours. METHODS AND RESULTS: We immunohistochemically analysed 23 IMPCs for expression of a standard form of CD44 (CD44s), Ecad, and CD34 to measure microvessel density (MVD). Results are compared with the changes observed in 23 tubular carcinomas (TCs), another variant of ductal carcinoma that rarely metastasizes. Evaluation of haematoxylin and eosin (H&E) sections showed a higher prevalence of lymph-vascular invasion (19/23, 83%) and regional lymph node involvement (12/15, 80%) in IMPCs; whereas no lymph-vascular invasion or lymph node metastasis was identified in TCs. Loss or reduction of CD44s immunoreactivity was significantly frequent in IMPC (39%) compared with TC (4%) (P = 0.0098), and was associated with positive axillary lymph nodes and lymph-vascular invasion. All cases of IMPC and TC strongly expressed Ecad. MVD (in five 200x fields) was significantly higher in IMPC (88 +/ 37) than in TC (57 +/- 16) (P = 0.001). In the IMPC group, MDV was higher in cases with positive lymph node(s) (P = 0.048), and cases with loss or reduction of CD44s expression (P = 0.011). The same trend was also demonstrated in cases with lymph-vascular invasion (P = 0.077). Moreover, the vessels in IMPC had much smaller calibres with thinner walls than those in TC. CONCLUSIONS: Loss of the CD44 adhesion molecule and high MVD may play a significant role in the high incidence of lymph-vascular permeation and metastasis in IMPC. PMID- 15656883 TI - Hypoxia-inducible factor-1alpha is associated with angiogenesis, and expression of bFGF, PDGF-BB, and EGFR in invasive breast cancer. AB - AIMS: Hypoxia-inducible factor-1 (HIF-1) is the key transcription factor regulating the cellular response to hypoxia, including angiogenesis. Growth factors play an important role in tumour growth and angiogenesis and some have been shown to be induced by HIF-1 in vitro. This study investigated if angiogenesis or growth factors or their receptors are associated with HIF-1alpha in invasive breast cancer. METHODS AND RESULTS: High levels of HIF-1alpha, detected by immunohistochemistry in 45 breast cancers, were positively associated with increased microvessel density (as a measure of angiogenesis) (P = 0.023). Furthermore, high levels of HIF-1alpha were associated with epithelial expression (> or = 10%) of epidermal growth factor receptor (EGFR) (P = 0.011), platelet derived growth factor (PDGF)-BB (P < 0.001), and basic fibroblast growth factor (bFGF) (P = 0.045). A positive, yet insignificant, trend for HIF-1alpha to be associated with epithelial expression of transforming growth factor (TGF)-alpha (P = 0.081) and vascular endothelial growth factor (VEGF) (P = 0.109) was noticed as well as an inverse association with stromal expression of TGF-beta-R1 (P = 0.070). CONCLUSIONS: In invasive breast cancer, HIF-1alpha is associated with angiogenesis, and expression of growth factors bFGF and PDGF-BB, and the receptor EGFR. Thus, agents targeting HIF-1 may combine different pathways of inhibiting breast cancer growth, including angiogenesis and growth factors. PMID- 15656884 TI - PPARgamma expression in breast cancer: clinical value and correlation with ERbeta. AB - AIMS: To examine the expression of peroxisome proliferator-activated receptor gamma (PPARgamma) in invasive breast carcinoma in relation to known clinicopathological features, ERbeta, and relapse-free and overall patient survival. PPARgamma is a ligand-activated transcriptional factor that regulates the transcription of various target genes and has been implicated in human breast cancer. METHODS AND RESULTS: We performed immunohistochemistry to detect PPARgamma, ERalpha, PR and ERbeta in 170 infiltrative breast carcinomas. The results were subjected to statistical analysis. PPARgamma was detected in the cytoplasm of 58% of breast carcinoma samples. PPARgamma did not differ with regard to any of the clinicopathological parameters except for histological grade, to which it was found to be inversely correlated (P = 0.019), and ERbeta, to which it was positively related (P = 0.016). As regards relapse-free survival, in univariate statistical analysis PPARgamma was found to exert a marginally favourable impact on all the patients (P = 0.076), but a strong one on patients with ductal carcinoma (P = 0.027), whereas Cox's regression analysis depicted PPARgamma to be an independent prognosticator for patients with ductal carcinoma (P = 0.039). No association was found between PPARgamma expression and overall survival. CONCLUSION: These results indicate the favourable impact of PPARgamma expression on disease-free survival of patients with ductal breast carcinoma and its possible cooperation with ERbeta in exerting that favourable effect. PMID- 15656885 TI - Cystic hypersecretory carcinoma: rare and poorly recognized variant of intraductal carcinoma of the breast. Report of five cases. AB - AIMS: To report five cases of a rare variant of intraductal carcinoma of the breast, so-called cystic hypersecretory carcinoma. The clinical and pathological characteristics of the lesion are described, along with a review of the literature. METHODS AND RESULTS: The patients were females aged between 53 and 78 years (average 66.8 years). The size of the lesions ranged between 70 and 80 mm in largest dimension. In two cases, the development of high-grade invasive ductal carcinoma was observed; in one additional case there was recurrence of high-grade in-situ carcinoma after 3 years. This emphasizes the importance of correct diagnosis of this potentially aggressive lesion. Strong over-expression of HER 2/neu protein was observed in three cases, including the two with an invasive component. Protein p53 was variably positive in all cases. Steroid receptor immunohistochemistry yielded variable results with only one case being positive for both oestrogen and progesterone receptors. Interestingly, in most cases (4/5) staining for androgen receptors was observed. CONCLUSIONS: Cystic hypersecretory ductal carcinoma of the breast is a rare distinctive variant of ductal carcinoma in situ. It has the potential for invasive growth and the development of metastases. PMID- 15656886 TI - Transitional cell carcinoma of the bladder mimicking lobular carcinoma of the breast: a discohesive variant of urothelial carcinoma. AB - AIMS: To describe a series of 10 cases of transitional cell carcinoma which show morphological features which mimic lobular carcinoma of the breast and diffuse carcinoma of the stomach. METHODS AND RESULTS: Ten cases were identified from the files at Southampton University Hospitals NHS Trust and from the authors' consultation files. Immunostains were performed and clinical information was obtained. Eight of the patients were male and two female. Ages ranged from 52 to 77 years at presentation. All of the tumours showed areas where the tumour was composed of uniform cells with a discohesive single-cell, diffusely invasive growth pattern. In areas the tumour cells were arranged in linear single-cell files and in separate areas solid sheets of discohesive cells. In all of the cases some tumour cells showed prominent intracytoplasmic vacuoles. In addition to this pattern, four cases showed typical transitional cell carcinoma or carcinoma in situ. The majority of the tumours expressed cytokeratin 20 but not oestrogen receptors. CONCLUSION: This study highlights a pattern of diffusely invasive transitional cell carcinoma not previously described and one which is important to recognize in order to avoid misdiagnosis of metastatic lobular carcinoma of the breast, especially in small biopsies. PMID- 15656887 TI - Small cell carcinoma of the bladder: a contemporary clinicopathological study of 51 cases. AB - AIMS: We present 51 cases of primary small cell carcinoma of the bladder in a clinicopathological study with emphasis on features that aid in the initial recognition and diagnosis of small cell carcinoma of the bladder. METHODS AND RESULTS: The patients were 40 men and 11 women between the ages of 39 and 87 years (mean age 67 years). Clinical data were available in 41 cases. The most common symptomatology was haematuria in 63% of the patients while dysuria was present in 12%. Thirty-eight patients were caucasians; seven patients were Hispanics; two patients were Asian; one patient was African-American; in the three additional patients no racial information was obtained. Biopsy material was obtained in all of the patients. Cystectomy was performed in 20 patients. At diagnosis, clinical stage was as follows: stage I in two (5%), stage II in 18 (44%), stage III in 10 (24%), and stage IV in 11 (27%). Histologically, urothelial carcinoma was present in 70% of the cases, adenocarcinoma in 8%, and squamous cell carcinoma in 10% of the cases. Small cell carcinoma was the only histology present in only 12% of the cases studied. Immunohistochemical studies using chromogranin, synaptophysin and chromogranin were positive in 30-70% of the cases. CONCLUSIONS: The present study highlights the unusual phenomenon of pure small cell carcinoma of the bladder and its association with other non-small cell carcinomas in that anatomical location. In addition, the study highlights the different modalities employed to treat patients in whom there is a component of small cell carcinoma of the bladder. PMID- 15656888 TI - Elevated expression of hyaluronan and its CD44 receptor in the duodenal mucosa of coeliac patients. AB - AIMS: Since hyaluronan (HA) metabolism is disturbed in some malignant tumours and in inflammatory diseases, we analysed HA and its receptor CD44 as well as the expression of the Ki67 nuclear protein, a marker of cell proliferation, in histological sections of duodenal biopsies of coeliac disease patients and controls. METHODS AND RESULTS: The study group consisted of 52 patients with coeliac disease in remission, 40 patients with newly diagnosed disease and 10 healthy control subjects. HA was detected with a specific biotinylated probe prepared from cartilage aggrecan and link protein, and CD44 with an antibody recognizing all forms of CD44 and another specific for its v6 variant. For the expression of the nuclear protein, monoclonal antibody MIB-1 was used. The percentage of HA-positive cells in surface epithelium was higher in newly diagnosed patients (13%) compared with patients in remission (11%) and controls (2%). In addition, HA intensity in the lamina propria was decreased in the newly diagnosed patients. In patients with active disease, 22-26% of the surface epithelium was CD44+, whereas the corresponding figure in patients in remission was 5%, and that of controls 1%. The more intensive MIB-1 labelling in the duodenal epithelium of coeliac patients without treatment was normalized after gluten-free diet. CONCLUSIONS: The HA-positive coat on surface epithelium seen even in patients in remission suggests persistent or even permanent changes in the epithelial permeability barrier in coeliac disease. PMID- 15656889 TI - Hypertrophic eosinophilic gastroenteropathy is associated with reduced enterocyte apoptosis. AB - AIMS: To investigate the cause of grossly elongated villi in four children presenting with obstruction due to a novel form of eosinophilic gastroenteropathy in which there was profound hyperplasia of the intestinal villi with grossly increased villous/crypt ratio and prominent mucosal eosinophilia. Increased eosinophils were also present in the muscularis propria and submucosa. All had intermittent diarrhoea and signs of a protein-losing enteropathy. METHODS AND RESULTS: The cause of the grossly elongated villi was investigated by studying enterocyte proliferation (Ki67), survival factors (bcl-2) and apoptosis (TUNEL) in these patients (n = 4) and normal (jejunum n = 6, ileum n = 6) and disease (n = 6) controls. The most remarkable finding was that apoptotic enterocytes were undetectable in the elongated villi. CONCLUSIONS: It seems likely that a defect in the regulation of apoptosis of the epithelium occurs which could explain the remarkable hyperplasia of the villi seen. PMID- 15656890 TI - Colonic angiodysplasia and true diverticula: is there an association? AB - AIMS: To investigate the pathology of colectomy specimens, from patients presenting with lower gastrointestinal haemorrhage, who had undergone preoperative mesenteric angiography. The angiography diagnoses ranged from active bleeding of unknown aetiology to angiodysplasia. METHODS AND RESULTS: The macroscopic and microscopic pathology of 11 colectomy specimens was studied. All the specimens contained blood on receipt with no obvious macroscopic source of haemorrhage identified. In all cases the large bowel demonstrated diverticula with an unusual topography. These were wide-mouthed, up to 20 mm across and exhibited a partly scattered distribution across the bowel; in two cases the diverticula contained blood clot. Histology demonstrated true diverticula invested by all muscle layers, exhibiting thick-walled vessels, within the submucosa and within the muscularis propria fibres, resembling the vascular elements identified in congenital vascular malformations. The two cases that contained blood clot revealed the thick-walled vessels to have ruptured into the diverticular lumen. Histology from the background colon between the diverticula demonstrated vascular lesions fulfilling the histological criteria for angiodysplasia; these features were also represented in the true-type diverticula. Most cases showed right-sided angiodysplasia and right-sided true diverticula. However, left-sided angiodysplastic lesions were also seen in association with left-sided true-type diverticula. CONCLUSIONS: We have identified the dual pathologies of angiodysplasia and multiple true-type diverticula, and suggest a topographical relationship between these two lesions. Furthermore, we propose that the thick-walled vessels within the true diverticula are congenitally abnormal, and raise the possibility that these vessels exert local haemodynamic effects, that effectively predispose the colon to acquired angiodysplastic phenomena. PMID- 15656891 TI - Immunohistochemical differentiation of eosinophilic heart diseases using antibodies against eosinophil activation markers. AB - AIMS: Eosinophilic heart syndromes are rare in Western countries and include endocarditis parietalis fibroplastica (EPF) and hypersensitivity myocarditis (HM). There are striking differences in natural history and morphological findings. Since diagnosis can be difficult when analysing small myocardial biopsies lacking the characteristic histological features, we studied a set of immunohistochemical markers in order to characterize the activation status of the infiltrating eosinophils to distinguish between these two entities. METHODS AND RESULTS: This study is based on the investigation of seven explanted hearts and one left ventricular specimen collected during implantation of a left ventricular assist device from a total of seven patients with HM. Also investigated were three right and three left ventricular specimens from five patients with EPF. We used antibodies (Ab) against EG1, and EG2, CD44, and CD69 which have been described as markers to distinguish between resting and activated eosinophils. The EG1 to EG2 ratio of eosinophils and the immunoreactivity against CD44 showed no differences between the two entities. However, eosinophils in the EPF were completely negative for CD69, whereas eosinophils reacted positively within the HM group. CONCLUSION: The immunohistochemical investigation of eosinophilic heart diseases using antibodies against CD69 can be a useful tool to distinguish between hypersensitivity myocarditis and endocarditis parietalis fibroplastica. PMID- 15656892 TI - Fat in the right ventricle of the normal heart. AB - AIMS: Fibrofatty replacement of the right ventricle wall, often with associated inflammation, is the hallmark of arrhythmogenic right ventricular cardiomyopathy (ARVC), a rare but established cause of sudden cardiac death in young adults. Fatty infiltration of the right ventricle alone without fibrosis may also occur but its relation to sudden death is not well established. In this study we assessed the amount of epicardial and intramyocardial fat in the right ventricle of 'normal' hearts from subjects who had died of non-cardiac causes. METHODS AND RESULTS: Hearts (n = 148) were examined from 81 males and 67 females, with an age range of 6 months to 68 years, who had died of non-cardiac causes. The extent and distribution of right ventricular epicardial and intramyocardial fat was assessed macro- and microscopically, respectively. The majority of hearts (85%) contained at least some intramyocardial fat with significantly more fat replacement noted in the right ventricles of older subjects and in females than in males. There was no significant fibrosis or inflammation in any of the 148 cases. CONCLUSION: Variable amounts of intramyocardial fat may be seen in the right ventricle of subjects dying of non-cardiac related causes. Care should be taken not to confuse this relatively common simple fatty infiltration with ARVC. PMID- 15656893 TI - Radiation-induced atypical squamous metaplasia of the urinary bladder mucosa with involvement of subepithelial tissue mimicking metastatic cervical squamous carcinoma. PMID- 15656895 TI - Trichogerminoma: further evidence to support a specific follicular neoplasm. PMID- 15656897 TI - Osteosarcoma arising in a metastatic leiomyosarcoma. PMID- 15656896 TI - The prognostic relevance of estimates of proliferative activity in early breast cancer. PMID- 15656898 TI - Plasmablastic lymphoma arising at a cutaneous site. PMID- 15656899 TI - Abdominopelvic perivascular epithelioid cell sarcoma (malignant PEComa) mimicking gastrointestinal stromal tumour of the rectum. PMID- 15656900 TI - The stability of life satisfaction in a 15-year follow-up of adult Finns healthy at baseline. AB - BACKGROUND: While physical health has improved considerably over recent decades in Finland, the disease burden of mental health, especially that of depression, has become increasingly demanding. However, we lack long-term data on the natural course of subjective well-being in the general population. The aim of this study was to investigate the long-term course of self-reported life satisfaction. METHODS: This was a 15-year prospective cohort study on a nationwide sample of adult Finnish twins (N = 9679), aged 18-45 and healthy at baseline, who responded to postal questionnaires in 1975, 1981 and 1990 including a 4-item life satisfaction scale (happiness/easiness/interest in life and feelings of loneliness). Life satisfaction score (range: 4-20) was classified into three categories: satisfied (4-6), intermediate (7-11) and dissatisfied group (12-20). The associations between life satisfaction scores during the follow-up were studied with linear/logistic regression. RESULTS: Moderate stability and only a slight effect of age or birth-cohort on mean life satisfaction score (LS) were detected. In 1990, 56% of all and 31% of the dissatisfied remained in the same LS category as at baseline. Only 5.9% of the study subjects changed from being satisfied to dissatisfied or vice versa. Correlations between continuous scores (1975, 1981 and 1990) were 0.3-0.4. Baseline dissatisfaction (compared to satisfaction) predicted dissatisfaction in 1981 (OR = 10.4; 95%CI 8.3-13.1) and 1990 (5.6; 4.6-6.8). Multiple adjustments decreased the risk only slightly. CONCLUSIONS: Life satisfaction in adult Finns was moderately stable during 15 years. Among an identifiable group (i.e. the dissatisfied) life dissatisfaction may become persistent, which places them at a greater risk of adverse health outcomes. PMID- 15656901 TI - Risk adjustment methods for Home Care Quality Indicators (HCQIs) based on the minimum data set for home care. AB - BACKGROUND: There has been increasing interest in enhancing accountability in health care. As such, several methods have been developed to compare the quality of home care services. These comparisons can be problematic if client populations vary across providers and no adjustment is made to account for these differences. The current paper explores the effects of risk adjustment for a set of home care quality indicators (HCQIs) based on the Minimum Data Set for Home Care (MDS-HC). METHODS: A total of 22 home care providers in Ontario and the Winnipeg Regional Health Authority (WRHA) in Manitoba, Canada, gathered data on their clients using the MDS-HC. These assessment data were used to generate HCQIs for each agency and for the two regions. Three types of risk adjustment methods were contrasted: a) client covariates only; b) client covariates plus an "Agency Intake Profile" (AIP) to adjust for ascertainment and selection bias by the agency; and c) client covariates plus the intake Case Mix Index (CMI). RESULTS: The mean age and gender distribution in the two populations was very similar. Across the 19 risk-adjusted HCQIs, Ontario CCACs had a significantly higher AIP adjustment value for eight HCQIs, indicating a greater propensity to trigger on these quality issues on admission. On average, Ontario had unadjusted rates that were 0.3% higher than the WRHA. Following risk adjustment with the AIP covariate, Ontario rates were, on average, 1.5% lower than the WRHA. In the WRHA, individual agencies were likely to experience a decline in their standing, whereby they were more likely to be ranked among the worst performers following risk adjustment. The opposite was true for sites in Ontario. CONCLUSIONS: Risk adjustment is essential when comparing quality of care across providers when home care agencies provide services to populations with different characteristics. While such adjustment had a relatively small effect for the two regions, it did substantially affect the ranking of many individual home care providers. PMID- 15656903 TI - Identification of amplified and highly expressed genes in amplicons of the T-cell line huT78 detected by cDNA microarray CGH. AB - BACKGROUND: Conventional Comparative Genomic Hybridization (CGH) has been widely used for detecting copy number alterations in cancer and for identifying regions containing candidate tumor responsible genes. Recently, several studies have shown the utility of cDNA microarray CGH for studing gene copy changes in various types of tumors. However, no such studies on T-cell lymphomas have been performed. To date T-cell lymphomas analyzed by the use of chromosome CGH have revealed only slight copy number alterations and not gene amplifications. RESULTS: In the present study, we describe the characterization of three amplicons of the T-cell line huT78 located at 2q34-q37, 8q23-q24 and 20p, where new amplified and overexpressed genes are found. The use of a cDNA microarray containing 7.657 transcripts allowed the identification of certain genes, such as BCLX, PCNA, FKBP1A, IGFBP2 and cMYC, that are amplified, highly expressed, and also contained in the amplicons on 20p and 2q. The expresion of these genes was analyzed in 39 T-cell lymphomas and 3 other T-cell lines. CONCLUSION: By the use of conventional CGH and CGH and expression cDNA microarrays we defined three amplicons in the T-cell line huT78 and identified several novel gene amplifications (BCLX, PCNA, FKBP1A, IGFBP2 and cMYC). We showed that overexpression of the amplified genes could be attributable to gene dosage. We speculate that deregulation of those genes could be important in the development of T-cell lymphomas and/or in the maintenance of T-cell lines. PMID- 15656902 TI - Microarray analysis of gene expression profiles of cardiac myocytes and fibroblasts after mechanical stress, ionising or ultraviolet radiation. AB - BACKGROUND: During excessive pressure or volume overload, cardiac cells are subjected to increased mechanical stress (MS). We set out to investigate how the stress response of cardiac cells to MS can be compared to genotoxic stresses induced by DNA damaging agents. We chose for this purpose to use ionising radiation (IR), which during mediastinal radiotherapy can result in cardiac tissue remodelling and diminished heart function, and ultraviolet radiation (UV) that in contrast to IR induces high concentrations of DNA replication- and transcription-blocking lesions. RESULTS: Cultures enriched for neonatal rat cardiac myocytes (CM) or fibroblasts were subjected to any one of the three stressors. Affymetrix microarrays, analysed with Linear Modelling on Probe Level, were used to determine gene expression patterns at 24 hours after (the start of) treatment. The numbers of differentially expressed genes after UV were considerably higher than after IR or MS. Remarkably, after all three stressors the predominant gene expression response in CM-enriched fractions was up regulation, while in fibroblasts genes were more frequently down-regulated. To investigate the activation or repression of specific cellular pathways, genes present on the array were assigned to 25 groups, based on their biological function. As an example, in the group of cholesterol biosynthesis a significant proportion of genes was up-regulated in CM-enriched fractions after MS, but down regulated after IR or UV. CONCLUSION: Gene expression responses after the types of cellular stress investigated (MS, IR or UV) have a high stressor and cell type specificity. PMID- 15656904 TI - A methodology for distinguishing divergent cell fates within a common progenitor population: adenoma- and neuroendocrine-like cells are confounders of rat ileal epithelial cell (IEC-18) culture. AB - BACKGROUND: IEC-18 cells are a non-transformed, immortal cell line derived from juvenile rat ileal crypt cells. They may have experimental advantages over tumor derived gastrointestinal lineages, including preservation of phenotype, normal endocrine responses and retention of differentiation potential. However, their proclivity for spontaneous differentiation/transformation may be stereotypical and could represent a more profound experimental confounder than previously realized. We hypothesized that IEC-18 cells spontaneously diverge towards a uniform mixture of epigenetic fates, with corresponding phenotypes, rather than persist as a single progenitor lineage. RESULTS: IEC-18 cells were cultured for 72 hours in serum free media (SFM), with and without various insulin-like growth factor agonists to differentially boost the basal rate of proliferation. A strategy was employed to identify constitutive genes as markers of divergent fates through gene array analysis by cross-referencing fold-change trends for individual genes against crypt cell abundance in each treatment. We then confirmed the cell-specific phenotype by immunolocalization of proteins corresponding to those genes. The majority of IEC-18 cells in SFM alone had a loss in expression of the adenomatous polyposis coli (APC) gene at the mRNA and protein levels, consistent with adenoma-like transformation. In addition, a small subset of cells expressed the serotonin receptor 2A gene and had neuroendocrine like morphology. CONCLUSIONS: IEC-18 cells commonly undergo a change in cell fate prior to reaching confluence. The most common fate switch that we were able to detect correlates with a down regulation of the APC gene and transformation into an adenoma-like phenotype. PMID- 15656905 TI - Lymphocyte subsets in hemophilic patients with hepatitis C virus infection with or without human immunodeficiency virus co-infection: a nested cross-sectional study. AB - BACKGROUND: With chronic infection, hepatitis C virus (HCV) RNA can be detected in B cells and associated with B-cell disorders, but these are not well defined. METHODS: The relationship between HCV infection and lymphocyte subpopulations was evaluated rigorously in 120 asymptomatic hemophilic patients, randomly selected from a prospective cohort study. CD4+ T cells, CD8+ T cells, CD19+ B cells, and CD56+ NK cells were quantified by flow cytometry using cryopreserved peripheral blood mononuclear cells of 24 hemophilic patients in each of five age-matched groups [uninfected; chronic HCV with or without human immunodeficiency virus (HIV); and cleared HCV with or without HIV]. RESULTS: As expected, patients with HIV had significantly reduced CD4+ and increased CD8+ T cells. Irrespective of HIV, patients with chronic HCV infection had approximately 25% fewer CD19+ B cells than those without chronic HCV infection. CONCLUSIONS: These data support the hypothesis that asymptomatic patients with chronic HCV infection have an altered B-lymphocyte population. PMID- 15656906 TI - MDM2 negatively regulates the human telomerase RNA gene promoter. AB - BACKGROUND: We have previously demonstrated that NF-Y and Sp1 interact with the human telomerase RNA (hTR) promoter and play a central role in its regulation. We have also shown that pRB activates the hTR promoter, but the mechanism of pRb directed activation is unknown. It has recently been reported that pRB induces Sp1 activity by relieving inhibition mediated by mdm2. The aim was to investigate possible roles for mdm2 in hTR promoter regulation. METHODS: Chromatin immunoprecipitation was used to determine binding of mdm2 to the hTR promoter. Transfection and luciferase assays were used to investigate mdm2 repression of the promoter activity and interaction with known transcriptional modulators. RESULTS: Here we show using chromatin immunoprecipitation that mdm2 specifically binds the hTR promoter in vivo. Transient co-transfection experiments using an hTR promoter luciferase reporter construct show that hTR promoter activity is inhibited by over-expression of mdm2 in 5637 bladder carcinoma cells (p53 and pRB negative, low mdm2). Titration of mdm2 was able to antagonise activation of hTR promoter activity mediated by pRB or Sp1 over-expression, although in the presence of pRB, mdm2 could not repress promoter activity below basal levels. Using an Sp1 binding site mutation construct we showed that mdm2 repression did not absolutely require Sp1 binding sites in the hTR promoter, suggesting the possibility of pRB/Sp1 independent mechanisms of repression. Finally, we show that NF-Y mediated transactivation of the hTR promoter was also suppressed by mdm2 in a dose-dependent manner. CONCLUSIONS: These studies suggest that mdm2 may inhibit the hTR promoter by multiple mechanisms. Mdm2 may directly repress activation by both pRB and Sp1, or activation by NF-Y. Furthermore, the ability of mdm2 to interact and interfere with components of the general transcription machinery might partly explain the general repressive effect seen here. Elucidation of new regulators affecting hTR basal promoter activity in cancer cells provides a basis for future studies aimed at improving our understanding of the differential hTR expression between normal and cancer cells. PMID- 15656907 TI - Randomized clinical trial of surgery versus conservative therapy for carpal tunnel syndrome [ISRCTN84286481]. AB - BACKGROUND: Conservative treatment remains the standard of care for treating mild to moderate carpal tunnel syndrome despite a small number of well-controlled studies and limited objective evidence to support current treatment options. There is an increasing interest in the usefulness of wrist magnetic resonance imaging could play in predicting who will benefit for various treatments. METHOD AND DESIGN: Two hundred patients with mild to moderate symptoms will be recruited over 3 1/2 years from neurological surgery, primary care, electrodiagnostic clinics. We will exclude patients with clinical or electrodiagnostic evidence of denervation or thenar muscle atrophy. We will randomly assign patients to either a well-defined conservative care protocol or surgery. The conservative care treatment will include visits with a hand therapist, exercises, a self-care booklet, work modification/ activity restriction, B6 therapy, ultrasound and possible steroid injections. The surgical care would be left up to the surgeon (endoscopic vs. open) with usual and customary follow-up. All patients will receive a wrist MRI at baseline. Patients will be contacted at 3, 6, 9 and 12 months after randomization to complete the Carpal Tunnel Syndrome Assessment Questionnaire (CTSAQ). In addition, we will compare disability (activity and work days lost) and general well being as measured by the SF-36 version II. We will control for demographics and use psychological measures (SCL-90 somatization and depression scales) as well as EDS and MRI predictors of outcomes. DISCUSSION: We have designed a randomized controlled trial which will assess the effectiveness of surgery for patients with mild to moderate carpal tunnel syndrome. An important secondary goal is to study the ability of MRI to predict patient outcomes. PMID- 15656909 TI - Effects of the cyclooxygenase-2 inhibitor nimesulide on cerebral infarction and neurological deficits induced by permanent middle cerebral artery occlusion in the rat. AB - BACKGROUND: Previous studies suggest that the cyclooxygenase-2 (COX-2) inhibitor nimesulide has a remarkable protective effect against different types of brain injury including ischemia. Since there are no reports on the effects of nimesulide on permanent ischemic stroke and because most cases of human stroke are caused by permanent occlusion of cerebral arteries, the present study was conducted to assess the neuroprotective efficacy of nimesulide on the cerebral infarction and neurological deficits induced by permanent middle cerebral artery occlusion (pMCAO) in the rat. METHODS: Ischemia was induced by permanent occlusion of the middle cerebral artery in rats, via surgical insertion of a nylon filament into the internal carotid artery. Infarct volumes (cortical, subcortical and total) and functional recovery, assessed by neurological score evaluation and rotarod performance test, were performed 24 h after pMCAO. In initial experiments, different doses of nimesulide (3, 6 and 12 mg/kg; i.p) or vehicle were administered 30 min before pMCAO and again at 6, 12 and 18 h after stroke. In later experiments we investigated the therapeutic time window of protection of nimesulide by delaying its first administration 0.5-4 h after the ischemic insult. RESULTS: Repeated treatments with nimesulide dose-dependently reduced cortical, subcortical and total infarct volumes as well as the neurological deficits and motor impairment resulting from permanent ischemic stroke, but only the administration of the highest dose (12 mg/kg) was able to significantly (P < 0.01) diminish infarct volume. The lower doses failed to significantly reduce infarction but showed a beneficial effect on neurological function. Nimesulide (12 mg/kg) not only reduced infarct volume but also enhanced functional recovery when the first treatment was given up to 2 h after stroke. CONCLUSIONS: These data show that nimesulide protects against permanent focal cerebral ischemia, even with a 2 h post-treatment delay. These findings have important implications for the therapeutic potential of using COX-2 inhibitors in the treatment of stroke. PMID- 15656908 TI - Inhibition of early steps in the lentiviral replication cycle by cathelicidin host defense peptides. AB - BACKGROUND: The antibacterial activity of host defense peptides (HDP) is largely mediated by permeabilization of bacterial membranes. The lipid membrane of enveloped viruses might also be a target of antimicrobial peptides. Therefore, we screened a panel of naturally occurring HDPs representing different classes for inhibition of early, Env-independent steps in the HIV replication cycle. A lentiviral vector-based screening assay was used to determine the inhibitory effect of HDPs on early steps in the replication cycle and on cell metabolism. RESULTS: Human LL37 and porcine Protegrin-1 specifically reduced lentiviral vector infectivity, whereas the reduction of luciferase activities observed at high concentrations of the other HDPs is primarily due to modulation of cellular activity and/ or cytotoxicity rather than antiviral activity. A retroviral vector was inhibited by LL37 and Protegrin-1 to similar extent, while no specific inhibition of adenoviral vector mediated gene transfer was observed. Specific inhibitory effects of Protegrin-1 were confirmed for wild type HIV-1. CONCLUSION: Although Protegrin-1 apparently inhibits an early step in the HIV-replication cycle, cytotoxic effects might limit its use as an antiviral agent unless the specificity for the virus can be improved. PMID- 15656910 TI - Evidence for preferential copackaging of Moloney murine leukemia virus genomic RNAs transcribed in the same chromosomal site. AB - BACKGROUND: Retroviruses have a diploid genome and recombine at high frequency. Recombinant proviruses can be generated when two genetically different RNA genomes are packaged into the same retroviral particle. It was shown in several studies that recombinant proviruses could be generated in each round of HIV-1 replication, whereas the recombination rates of SNV and Mo-MuLV are 5 to 10-fold lower. The reason for these differences is not clear. One possibility is that these retroviruses may differ in their ability to copackage genomic RNAs produced at different chromosomal loci. RESULTS: To investigate whether there is a difference in the efficiency of heterodimer formation when two proviruses have the same or different chromosomal locations, we introduced two different Mo-MuLV based retroviral vectors into the packaging cell line using either the cotransfection or sequential transfection procedure. The comparative study has shown that the frequency of recombination increased about four-fold when the cotransfection procedure was used. This difference was not associated with possible recombination of retroviral vectors during or after cotransfection and the ratios of retroviral virion RNAs were the same for two variants of transfection. CONCLUSIONS: The results of this study indicate that a mechanism exists to enable the preferential copackaging of Mo-MuLV genomic RNA molecules that are transcribed on the same DNA template. The properties of Mo-MuLV genomic RNAs transport, processing or dimerization might be responsible for this preference. The data presented in this report can be useful when designing methods to study different aspects of replication and recombination of a diploid retroviral genome. PMID- 15656911 TI - Gene expression profiling revealed novel mechanism of action of Taxotere and Furtulon in prostate cancer cells. AB - BACKGROUND: Both Taxotere and Capecitabine have shown anti-cancer activity against various cancers including prostate cancer. In combination, Taxotere plus Capecitabine has demonstrated higher anti-cancer activity in advanced breast cancers. However, the molecular mechanisms of action of Taxotere and Capecitabine have not been fully elucidated in prostate cancer. METHODS: The total RNA from PC3 and LNCaP prostate cells untreated and treated with 2 nM Taxotere, 110 microM Furtulon (active metabolite of Capecitabine), or 1 nM Taxotere plus 50 microM Furtulon for 6, 36, and 72 hours, was subjected to Affymetrix Human Genome U133A Array analysis. Real-time PCR and Western Blot analysis were conducted to confirm microarray data. RESULTS: Taxotere and Furtulon down-regulated some genes critical for cell proliferation, cell cycle progression, transcription factor, cell signaling, and oncogenesis, and up-regulated some genes related to the induction of apoptosis, cell cycle arrest, and differentiation in both cell lines. Taxotere and Furtulon also up-regulated some genes responsible for chemotherapeutic resistance, suggesting the induction of cancer cell resistance to these agents. CONCLUSIONS: Taxotere and Furtulon caused the alternation of a large number of genes, many of which may contribute to the molecular mechanisms by which Taxotere and Furtulon inhibit the growth of prostate cancer cells. This information could be utilized for further mechanistic research and for devising optimized therapeutic strategies against prostate cancer. PMID- 15656912 TI - An evaluation of the metabolic syndrome in the HyperGEN study. AB - BACKGROUND: In 2001 the National Cholesterol Education Program (NCEP) provided a categorical definition for metabolic syndrome (c-MetS). We studied the extent to which two ethnic groups, Blacks and Whites were affected by c-MetS. The groups were members of the Hypertension Genetic Epidemiology Network (HyperGEN), a part of the Family Blood Pressure Program, supported by the NHLBI. Although the c-MetS definition is of special interest in particular to the clinicians, the quantitative latent traits of the metabolic syndrome (MetS) are also important in order to gain further understanding of its etiology. In this study, quantitative evaluation of the MetS latent traits (q-MetS) was based on the statistical multivariate method factor analysis (FA). RESULTS: The prevalence of the c-MetS was 34% in Blacks and 39% in Whites. c-MetS showed predominance of obesity, hypertension, and dyslipidemia. Three and four factor domains were identified through FA, classified as "Obesity," "Blood pressure," "Lipids," and "Central obesity." They explained approximately 60% of the variance in the 11 original variables. Two factors classified as "Obesity" and "Central Obesity" overlapped when FA was performed without rotation. All four factors in FA with Varimax rotation were consistent between Blacks and Whites, between genders and also after excluding type 2 diabetes (T2D) participants. Fasting insulin (INS) associated mainly with obesity and lipids factors. CONCLUSIONS: MetS in the HyperGEN study has a compound phenotype with separate domains for obesity, blood pressure, and lipids. Obesity and its relationship to lipids and insulin is clearly the dominant factor in MetS. Linkage analysis on factor scores for components of MetS, in familial studies such as HyperGEN, can assist in understanding the genetic pathways for MetS and their interactions with the environment, as a first step in identifying the underlying pathophysiological causes of this syndrome. PMID- 15656913 TI - Hinderin, a five-domains protein including coiled-coil motifs that binds to SMC3. AB - BACKGROUND: The structural maintenance of chromosome proteins SMC1 and SMC3 play an important role in the maintenance of chromosomal integrity by preventing the premature separation of the sister chromatids at the onset of anaphase. The two proteins are constitutive components of the multimeric complex cohesin and form dimers by interacting at their central globular regions. RESULTS: In order to identify proteins that by binding to SMC3 may interfere with the protein dimerization process, a human cDNA library was screened by the yeast two-hybrid system by using the hinge region of SMC3 as bait. This has lead to the identification of Hinderin, a novel five domains protein including two coiled coil motifs and sharing a strikingly structural similarity to the SMC family of proteins. Hinderin is ubiquitously expressed in human tissues. Orthologue forms of the protein are present in other vertebrates but not in lower organisms. A mapping of the interaction sites revealed that the N- and C-terminal globular domains mediate the binding of Hinderin to SMC3. Hinderin/SMC3 complexes could be recovered by immunoprecipitation from cell lysates using an anti-SMC3 antibody, thus demonstrating that the two proteins interact in vivo. On the contrary, Hinderin did not interact with SMC1. In vivo the rate of SMC1/SMC3 interaction was decreased by the ectopic expression of Hinderin. CONCLUSIONS: Hinderin is a novel binding partner of SMC3. Based on its ability to modulate SMC1/SMC3 interaction we postulate that Hinderin affects the availability of SMC3 to engage in the formation of multimeric protein complexes. PMID- 15656914 TI - Genome-scale analysis of positional clustering of mouse testis-specific genes. AB - BACKGROUND: Genes are not randomly distributed on a chromosome as they were thought even after removal of tandem repeats. The positional clustering of co expressed genes is known in prokaryotes and recently reported in several eukaryotic organisms such as Caenorhabditis elegans, Drosophila melanogaster, and Homo sapiens. In order to further investigate the mode of tissue-specific gene clustering in higher eukaryotes, we have performed a genome-scale analysis of positional clustering of the mouse testis-specific genes. RESULTS: Our computational analysis shows that a large proportion of testis-specific genes are clustered in groups of 2 to 5 genes in the mouse genome. The number of clusters is much higher than expected by chance even after removal of tandem repeats. CONCLUSION: Our result suggests that testis-specific genes tend to cluster on the mouse chromosomes. This provides another piece of evidence for the hypothesis that clusters of tissue-specific genes do exist. PMID- 15656917 TI - The third time's a charm! PMID- 15656916 TI - Adaptogenic and nootropic activities of aqueous extract of Vitis vinifera (grape seed): an experimental study in rat model. AB - BACKGROUND: The aerial parts of Vitis vinifera (common grape or European grape) have been widely used in Ayurveda to treat a variety of common and stress related disorders. In the present investigation, the seed extract of V. vinifera was evaluated for antistress activity in normal and stress induced rats. Furthermore, the extract was studied for nootropic activity in rats and in-vitro antioxidant potential to correlate its antistress activity. METHODS: For the evaluation of antistress activity, groups of rats (n = 6) were subjected to forced swim stress one hour after daily treatment of V. vinifera extract. Urinary vanillylmandelic acid (VMA) and ascorbic acid were selected as non-invasive biomarkers to assess the antistress activity. The 24 h urinary excretion of vanillylmandelic acid (VMA) and ascorbic acid were determined by spectrophotometric methods in all groups under normal and stressed conditions. The nootropic activity of the extract as determined from acquisition, retention and retrieval in rats was studied by conditioned avoidance response using Cook's pole climbing apparatus. The in vitro antioxidant activity was determined based on the ability of V. vinifera to scavenge hydroxyl radicals. RESULTS: Daily administration of V. vinifera at doses of 100, 200 and 300 mg/kg body weight one hour prior to induction of stress inhibited the stress induced urinary biochemical changes in a dose dependent manner. However, no change in the urinary excretion of VMA and ascorbic acid was observed in normal animals at all the doses studied. The cognition, as determined by the acquisition, retention and recovery in rats was observed to be dose dependent. The extract also produced significant inhibition of hydroxyl radicals in comparison to ascorbic acid in a dose dependent manner. CONCLUSION: The present study provides scientific support for the antistress (adaptogenic), antioxidant and nootropic activities of V. vinifera seed extract and substantiate the traditional claims for the usage of grape fruits and seeds in stress induced disorders. PMID- 15656915 TI - Upregulated expression of human neutrophil peptides 1, 2 and 3 (HNP 1-3) in colon cancer serum and tumours: a biomarker study. AB - BACKGROUND: Molecular markers for localized colon tumours and for prognosis following therapy are needed. Proteomics research is currently producing numerous biomarker studies with clinical potential. We investigate the protein composition of plasma and of tumour extracts with the aim of identifying biomarkers for colon cancer. METHODS: By Surface Enhanced Laser Desorption/Ionisation--Time Of Flight/Mass spectrometry (SELDI-TOF/MS) we compare the protein profiles of colon cancer serum with serum from healthy individuals and the protein profiles of colon tumours with normal colon tissue. By size exclusion chromatography, we investigate the binding of HNP 1-3 to high mass plasma proteins. By microflow we investigate the effect of HNP 1-3 on mammalian cells. RESULTS: Human Neutrophil Peptides -1, -2 and -3 (HNP 1-3), also known as alfa-defensin-1, -2 and -3, are present in elevated concentrations in serum from colon cancer patients and in protein extracts from colon tumours. A fraction of HNP 1-3 in serum is bound to unidentified high mass plasma proteins. HNP 1-3 purified from colon tumours are lethal to mammalian cells. CONCLUSIONS: HNP 1-3 may serve as blood markers for colon cancer in combination with other diagnostic tools. We propose that HNP 1-3 are carried into the bloodstream by attaching to high mass plasma proteins in the tumour microenvironment. We discuss the effect of HNP 1-3 on tumour progression. PMID- 15656919 TI - Obstetric maternal outcomes at Bella Coola General Hospital: 1940 to 2001. AB - OBJECTIVE: To describe obstetric procedures (episiotomy, forceps, vacuum extraction, caesarean section) and maternal outcomes for patients who gave birth in an isolated, rural hospital. DESIGN: A retrospective cohort study. STUDY POPULATION: Women beyond 20 weeks' gestation who gave birth between Mar. 7, 1940, and June 9, 2001, inclusive, at the Bella Coola General Hospital (BCGH). MAIN OUTCOME MEASURES: Data collected included maternal age, date of delivery, mode of delivery (vaginal delivery v. cesarean section), whether an episiotomy was performed or not, if forceps or vacuum extraction were used, whether analgesia, sedation or anesthesia was used, and maternal mortality. RESULTS: There were 2373 deliveries, including 12 sets of twins. There were no maternal mortalities. Cesarean sections were not routinely performed until the 1970s. Since then, there has been an increase in cesarean section rates to 11% of all deliveries in the 1990s. In the 1940s 28% of deliveries involved an episiotomy. This increased to 47% in the 1970s and was followed by a sharp decline to 4% in the 1990s. There was an increase, followed by a more gradual decrease in the use of forceps, and there was a recent increase in the use of vacuum extraction. The changes in procedure rates appear to reflect best practice guidelines of the times. In the case of episiotomies, the data suggest rural physicians are capable of rapid incorporation of recent recommendations. Rates for all procedures tended to be lower than those reported elsewhere in Canada and the United States. Narcotics, sedatives, inhalation agents and regional anesthetics were used to relieve the pain of labour and delivery throughout the study period. CONCLUSIONS: Women giving birth in the low technology environment of the BCGH experienced relatively low obstetric procedural rates with excellent maternal outcomes. PMID- 15656920 TI - Perinatal outcomes at Bella Coola General Hospital: 1940 to 2001. AB - OBJECTIVE: To describe perinatal outcomes (mortality, weight, condition at birth) at an isolated, rural hospital. DESIGN: A retrospective cohort study. STUDY POPULATION: Neonates born to women beyond 20 weeks' gestation who delivered in the Bella Coola General Hospital (BCGH) between Mar. 7, 1940, and June 9, 2001, inclusive. MAIN OUTCOME MEASURES: Information collected from the labour and delivery case room record book includes Aboriginal status, date of delivery, birth weight, newborn mortality, and newborn condition at birth. RESULTS: There were 2373 deliveries, including 12 sets of twins. Total newborn mortality rates declined from approximately 4.7% in the 1940-1954 time period to 0.7% in the 1970 1984 time period and have remained near that level ever since. From 1940-1960 BCGH's perinatal mortality rate was higher than Canada's; it was lower than Canada's in the 1970s, higher in the 1980s and about the same for the 1990s. The condition of the vast majority (approximately 90%) of newborns was described as being "good" at birth. Approximately 5% of newborns had birth weights < 2500 g, and this has not changed much over the years. In the 1951-1962 time period Aboriginal women had a higher percentage (8%) of infants with birth weight < 2500 g compared with non-Aboriginal women (5%), but this percentage has declined over time to the point where the rate for both groups is now around 5%. CONCLUSIONS: Women giving birth in the low technology environment of the BCGH experienced acceptable neonatal outcomes. Trends in perinatal mortality, morbidity and low birth-weight rates mirror those recorded for Canada. PMID- 15656921 TI - Sustaining rural maternity care--don't forget the RNs. AB - INTRODUCTION: Registered nurses provide intrapartum care to women who choose to have their babies in hospital. Considering the current national shortage of nurses, the ability of registered nurses to continue to care for women, especially in small rural hospitals, is a critical concern. PURPOSES: The purposes of the study were 1) to conduct a systematic review of the maternal child-nursing literature in rural locations; and 2) to identify one rural Ontario hospital where nurses and physicians deliver care to women with low-risk pregnancies, and then conduct an institutional ethnography to understand the enablers and barriers to low-risk rural maternity care. METHODS: A literature search was conducted to determine the state of rural registered nurses; and a telephone survey of 25 rural Ontario hospitals was undertaken to locate a hospital in which an institutional ethnography study could be conducted. RESULTS: Registered nurses in rural areas are more likely to be multi-specialists than generalists because of the need to adapt to emergencies across the life continuum. To care for pregnant women and their families, registered nurses require many of the same considerations that physicians have outlined: access to continuing education, appropriate call-back schedules, support from other health care professionals and administrators, and a value system that respects their expertise. Results from the ethnography of one Ontario health care institution revealed that when these aforementioned considerations are addressed, registered nurses are able to provide safe, comprehensive low-risk care in a rural maternity programme. CONCLUSIONS: Registered nurses play an important collaborative role in maternity care. We need Canadian data on registered nurses so that we can educate, recruit and retain them to care for women with low-risk pregnancies in rural and remote ares of Canada. Nursing services should be reviewed. Collaborative care models integrating newer professionals such as midwives, as well as understanding the role of doulas, may help in developing sustainable care to rural women. PMID- 15656924 TI - The occasional breech. PMID- 15656922 TI - A comparison of Canadian medical students from rural and non-rural backgrounds. AB - CONTEXT: Very little is known about medical students from rural areas currently enrolled in Canadian medical schools. PURPOSE: We aimed to compare rural and non rural students in terms of demographics, socioeconomic status, financial status and career choices. METHODS: As part of a larger Internet survey of all students at Canadian medical schools outside Quebec, conducted in January and February 2001, we conducted post-hoc analyses to compare students from rural and non-rural areas. Canada Post's classification system was used to determine rural status. To compare differences between rural and non-rural students, we used logistical regression models for categorical variables and factorial analysis of variance for continuous variables. RESULTS: We received responses from 2994 (68.5%) of 4368 medical students. Eleven percent of Canadian medical students come from rural backgrounds. Rural students tend to be older and originate from families of lower socioeconomic status. Students from rural areas report higher levels of debt, increased rates of paid part-time and summer employment, and greater stress from their finances. Nevertheless, rural students are not more likely to state that financial considerations will affect their choice of specialty or practice location. CONCLUSIONS: Canadian medical students who come from rural backgrounds are different from their non-rural counterparts. Students from rural areas face numerous financial barriers in obtaining a medical education and report greater levels of financial stress. Medical schools should examine and address barriers to admission of rural students and they should consider directing more financial resources toward this financially vulnerable group. PMID- 15656927 TI - Clinical presentation, diagnosis, and management of gastroesophageal reflux disease. AB - GERD is ubiquitous throughout the adult population in the United States. It commonly adversely affects quality of life and occasionally causes life threatening complications. The new and emerging medical and endoscopic therapies for GERD and the new management strategies for BE should dramatically reduce the clinical toll of this disease on society. PMID- 15656926 TI - Pathogenesis of reflux esophagitis and Barrett's esophagus. AB - An understanding of the pathogenesis of reflux esophagitis and Barrett's esophagus requires knowledge of the noxious elements in gastric juice and the three major esophageal defenses designed to protect against them. When the esophageal epithelium cannot prevent gastric acid from acidifying the intercellular spaces, the foundation is set for the development of the major symptoms, signs, and complications of reflux esophagitis. Inadequate defense by the epithelium can occur by exposure to the acidic refluxate for a prolonged period of time, because of defects in the antireflux or luminal clearance mechanisms, or by exposure to ingested products that directly impair the epithelium's intrinsic defenses, rendering it vulnerable to injury from even physiologic levels of acid reflux. PMID- 15656928 TI - Epidemiology, pathophysiology, and treatment of Barrett's esophagus: reducing mortality from esophageal adenocarcinoma. AB - The definition of BE has evolved over time. BE is the key premalignant lesion for developing EAC. The epidemiology and pathophysiology of BE is outlined, and risk factors for BE and EAC are reviewed. GERD plays a crucial role in the pathophysiology and the clinical identification of BE. Endoscopy with biopsy is the best tool for diagnosing and surveying patients with BE. Detection of early neoplasia is the present approach to reduce EAC mortality. Novel technology should assist in the early detection of dysplasia to enable targeted therapy. Effective chemopreventive strategies may reduce the risk of progression to EAC. PMID- 15656929 TI - The relation of Helicobacter pylori to gastric adenocarcinoma and lymphoma: pathophysiology, epidemiology, screening, clinical presentation, treatment, and prevention. AB - Helicobacter pylori infection may be the most common chronic bacterial infection worldwide; however, the prevalence varies between countries and is usually linked to socioeconomic conditions. Gastric cancer is one of the most frequent cancers in developing countries and usually about the seventh most common in developed countries. This article explores the relation of H. pylori to gastric adenocarcinoma and lymphoma. The pathophysiology, epidemiology, screening, clinical presentation, treatment, and prevention are discussed. PMID- 15656931 TI - The progression of hepatitis B- and C-infections to chronic liver disease and hepatocellular carcinoma: epidemiology and pathogenesis. AB - HCC is a major problem in certain regions of the world, and an increasing problem in the United States. Although the precise mechanisms are not delineated, chronic viral hepatitis B and C clearly cause HCC. Epidemiologic evidence provides strong associations with both viral infections. The predominant pathway to HCC is through cirrhosis, but direct viral carcinogenic effects may also contribute, particularly for hepatitis B. Understanding of this epidemiology and pathophysiology provides the basis for the clinical evaluation, diagnosis, screening, and treatment of HCC. PMID- 15656930 TI - The progression of hepatitis B- and C-infections to chronic liver disease and hepatocellular carcinoma: presentation, diagnosis, screening, prevention, and treatment of hepatocellular carcinoma. AB - Much information has been gained in the diagnosis and treatment of HCC during the last 15 years. Ever improving imaging technology has made nonhistologic diagnostic criteria possible, albeit controversial. Liver transplantation, resection, and RFA are considered curative options. Yet, HCC incidence is steadily rising because of limited progress on disease prevention. Accurate and cost-effective screening is necessary. Presently, only 10% to 15% of HCC patients present with a curative stage of disease. Because the field of HCC is rapidly changing, patients with HCC should be referred to liver centers with a full array of services, from surgical to oncologic. The prognosis for HCC patients will surely improve with a multi-disciplinary approach to care and further clinical research. Better screening and prevention of recurrence should eventually improve survival. It is hoped that antiviral treatment studies will lower the risk of HCC, and that these changes will occur soon enough to help the many patients at risk for or diagnosed with HCC over the next several years. PMID- 15656932 TI - Relation of hemochromatosis with hepatocellular carcinoma: epidemiology, natural history, pathophysiology, screening, treatment, and prevention. AB - HH is a common inherited disorder of iron metabolism affecting about 1 out of 250 individuals of Northern European decent. Many of these patients do not have evident phenotypic expression and do not develop significant iron loading. Some patients, however, develop progressive iron overload and cirrhosis. These individuals are at risk of developing HCC. Cirrhotics with hemochromatosis should undergo regular screening for HCC. If HCC is identified early, treatment with either resection or liver transplantation is optimal. Palliative measures, including ablative therapy and chemoembolization, can be used. With increasing clinical recognition,hemochromatosis should be diagnosed earlier and progression to cirrhosis and HCC should be minimized. PMID- 15656934 TI - Nassarius kraussianus shell beads from Blombos Cave: evidence for symbolic behaviour in the Middle Stone Age. AB - Since 1991, excavations at Blombos Cave have yielded a well-preserved sample of faunal and cultural material in Middle Stone Age (MSA) levels. The uppermost MSA phase, M1, is dated to c. 75 ka by optically stimulated luminescence (OSL) and thermoluminescence, and the middle M2 phase to a provisional c. 78 ka. Artefacts unusual in a MSA context from these phases include bifacial points, bone tools, engraved ochre and engraved bone. In this paper, we describe forty-one marine tick shell beads recovered from these MSA phases and tick shell beads from Later Stone Age (LSA) levels at Blombos Cave and the Die Kelders site. Thirty-nine shell beads come from the upper M1 phase and two from M2. Morphometric, taphonomic and microscopic analysis of modern assemblages of living and dead tick shell demonstrate that the presence of perforated Nassarius kraussianus shells in the Blombos MSA levels cannot be due to natural processes or accidental transport by humans. The types of perforation seen on the MSA shells are absent on modern accumulations of dead shells and not attributable to post-depositional damage. Their location, size, and microscopic features are similar to those obtained experimentally by piercing the shell wall, through the aperture, with a sharp bone point. Use-wear, recorded on the perforation edge, the outer lip, and the parietal wall of the aperture indicates the shells having being strung and worn. MSA shell beads differ significantly in size, perforation type, wear pattern and shade compared to LSA beads and this eliminates the possibility of mixing across respective levels. Thirty-one beads were found in four groups of five to twelve beads, each group being recovered in a single square or in two adjacent sub squares during a single excavation day. Within a group, shells display a similar shade, use-wear pattern and perforation size suggesting their provenance from the same beadwork item, lost or disposed during a single event. The likely symbolic significance of these finds suggests levels of cognitively modern behaviour not previously associated with MSA people. PMID- 15656935 TI - The metabolic costs of 'bent-hip, bent-knee' walking in humans. AB - The costs of different modes of bipedalism are a key issue in reconstructing the likely gait of early human ancestors such as Australopithecus afarensis. Some workers, on the basis of morphological differences between the locomotor skeleton of A. afarensis and modern humans, have proposed that this hominid would have walked in a 'bent-hip, bent-knee' (BHBK) posture like that seen in the voluntary bipedalism of untrained chimpanzees. Computer modelling studies using inverse dynamics indicate that on the basis of segment proportions AL-288-1 should have been capable of mechanically effective upright walking, but in contrast predicted that BHBK walking would have been highly ineffective. The measure most pertinent to natural selection, however, is more likely to be the complete, physiological, or metabolic energy cost. We cannot measure this parameter in a fossil. This paper presents the most complete investigation yet of the metabolic and thermoregulatory costs of BHBK walking in humans. Data show that metabolic costs including the basal metabolic rate (BMR) increase by around 50% while the energy costs of locomotion and blood lactate production nearly double, heat load is increased, and core temperature does not return to normal within 20 minutes rest. Net effects imply that a resting period of 150% activity time would be necessary to prevent physiologically intolerable heat load. Preliminary data for children suggest that scaling effects would not significantly reduce relative costs for hominids of AL-288-1's size. Data from recent studies using forwards dynamic modelling confirm that similar total (including BMR) and locomotor metabolic costs would have applied to BHBK walking by AL-288-1. We explore some of the ecological consequences of our findings. PMID- 15656936 TI - Evolution of the brainstem orofacial motor system in primates: a comparative study of trigeminal, facial, and hypoglossal nuclei. AB - The trigeminal motor (Vmo), facial (VII), and hypoglossal (XII) nuclei of the brainstem comprise the final common output for neural control of most orofacial muscles. Hence, these cranial motor nuclei are involved in the production of adaptive behaviors such as feeding, facial expression, and vocalization. We measured the volume and Grey Level Index (GLI) of Vmo, VII, and XII in 47 species of primates and examined these nuclei for scaling patterns and phylogenetic specializations. Allometric regression, using medulla volume as an independent variable, did not reveal a significant difference between strepsirrhines and haplorhines in the scaling of Vmo volume. In addition, correlation analysis using independent contrasts did not find a relationship between Vmo size or GLI and the percent of leaves in the diet. The scaling trajectory of VII volume, in contrast, differed significantly between suborders. Great ape and human VII volumes, furthermore, were significantly larger than predicted by the haplorhine regression. Enlargement of VII in these taxa may reflect increased differentiation of the facial muscles of expression and greater utilization of the visual channel in social communication. The independent contrasts of VII volume and GLI, however, were not correlated with social group size. To examine whether the human hypoglossal motor system is specialized to control the tongue for speech, we tested human XII volume and GLI for departures from nonhuman haplorhine prediction lines. Although human XII volumes were observed above the regression line, they did not exceed prediction intervals. Of note, orang-utan XII volumes had greater residuals than humans. Human XII GLI values also did not differ from allometric prediction. In sum, these findings indicate that the cranial orofacial motor nuclei evince a mosaic of phylogenetic specializations for innervation of the facial muscles of expression in the context of a generally conservative scaling relationship with respect to medulla size. PMID- 15656938 TI - Developmental processes and canine dimorphism in primate evolution. AB - Understanding the evolutionary history of canine sexual dimorphism is important for interpreting the developmental biology, socioecology and phylogenetic position of primates. All current evidence for extant primates indicates that canine dimorphism is achieved through bimaturism rather than via differences in rates of crown formation time. Using incremental growth lines, we charted the ontogeny of canine formation within species of Eocene Cantius, the earliest known canine-dimorphic primate, to test whether canine dimorphism via bimaturism was developmentally canalized early in primate evolution. Our results show that canine dimorphism in Cantius is achieved primarily through different rates of crown formation in males and females, not bimaturism. This is the first demonstration of rate differences resulting in canine dimorphism in any primate and therefore suggests that canine dimorphism is not developmentally homologous across Primates. The most likely interpretation is that canine dimorphism has been selected for at least twice during the course of primate evolution. The power of this approach is its ability to identify underlying developmental processes behind patterns of morphological similarity, even in long-extinct primate species. PMID- 15656937 TI - Vocal production mechanisms in a non-human primate: morphological data and a model. AB - Human beings are thought to be unique amongst the primates in their capacity to produce rapid changes in the shape of their vocal tracts during speech production. Acoustically, vocal tracts act as resonance chambers, whose geometry determines the position and bandwidth of the formants. Formants provide the acoustic basis for vowels, which enable speakers to refer to external events and to produce other kinds of meaningful communication. Formant-based referential communication is also present in non-human primates, most prominently in Diana monkey alarm calls. Previous work has suggested that the acoustic structure of these calls is the product of a non-uniform vocal tract capable of some degree of articulation. In this study we test this hypothesis by providing morphological measurements of the vocal tract of three adult Diana monkeys, using both radiography and dissection. We use these data to generate a vocal tract computational model capable of simulating the formant structures produced by wild individuals. The model performed best when it combined a non-uniform vocal tract consisting of three different tubes with a number of articulatory manoeuvres. We discuss the implications of these findings for evolutionary theories of human and non-human vocal production. PMID- 15656939 TI - Reconstitution of glycopeptide export in mixed detergent-solubilised and resealed microsomes depleted of lumenal components. AB - Export of macromolecules from the endoplasmic reticulum (ER) lumen into the cytosol is a major aspect of the quality control systems operating within the early secretory system. Glycopeptides are exported from the ER by an ATP- and GTP dependent pathway, which shares many similarities to the protein export system. Significantly, for glycopeptides, there is no requirement for cytosolic factors, biochemically distinguishing the glycopeptide and protein paths and probably reflecting the lower conformational complexity of the former substrate. Genetic studies in yeast, and biochemical data from higher eukaryotes, indicate that glycopeptides utilise the Sec61 translocon. Here, we report a new system allowing access to lumenal ER components, facilitating assessment of their importance in glycopeptide retrotranslocation and potentially other processes. Saponin, in combination with CHAPS, but not saponin alone, facilitated removal of >95% of lumenal protein disulphide isomerase (PDI) and BiP. Upon resealing, these microsomes retained glycopeptide export competence. These data suggest that the majority of lumenal components of the ER are most likely nonessential for glycopeptide export. In addition, export competence was highly sensitive to the addition of external protease, indicating a role for protein factors with cytoplasmically exposed determinants. PMID- 15656940 TI - The comparison of the estimation of enzyme kinetic parameters by fitting reaction curve to the integrated Michaelis-Menten rate equations of different predictor variables. AB - The estimation of enzyme kinetic parameters by nonlinear fitting reaction curve to the integrated Michaelis-Menten rate equation ln(S(0)/S)+(S(0) S)/K(m)=(V(m)/K(m))xt was investigated and compared to that by fitting to (S(0) S)/t=V(m)-K(m)x[ln(S(0)/S)/t] (Atkins GL, Nimmo IA. The reliability of Michaelis Menten constants and maximum velocities estimated by using the integrated Michaelis-Menten equation. Biochem J 1973;135:779-84) with uricase as the model. Uricase reaction curve was simulated with random absorbance error of 0.001 at 0.075 mmol/l uric acid. Experimental reaction curve was monitored by absorbance at 293 nm. For both CV and deviation <20% by simulation, K(m) from 5 to 100 micromol/l was estimated with Eq. (1) while K(m) from 5 to 50 micromol/l was estimated with Eq. (2). The background absorbance and the error in the lag time of steady-state reaction resulted in negative K(m) with Eq. (2), but did not affect K(m) estimated with Eq. (1). Both equations gave better estimation of V(m). The computation time and the goodness of fit with Eq. (1) were 40-fold greater than those with Eq. (2). By experimentation, Eq. (1) yielded K(m) consistent with the Lineweaver-Burk plot analysis, but Eq. (2) gave many negative parameters. Apparent K(m) by Eq. (1) linearly increased, while V(m) were constant, vs. xanthine concentrations, and the inhibition constant was consistent with the Lineweaver-Burk plot analysis. These results suggested that the integrated rate equation that uses the predictor variable of reaction time was reliable for the estimation of enzyme kinetic parameters and applicable for the characterization of enzyme inhibitors. PMID- 15656941 TI - Measurement of mitochondrial respiration in permeabilized murine neuroblastoma (N 2alpha) cells, a simple and rapid in situ assay to investigate mitochondrial toxins. AB - Most mitochondria-based methods used to investigate toxins require the use of relatively large amounts of material and hence compromised sensitivity in assay. We adopted procedures from methods initially developed to diagnose mitochondrial encephalomyopathies and unified these into a single assay. Eukaryotic cell membranes are selectively permeabilized with digitonin to render a system in which mitochondrial respiration can be measured rapidly and with considerable sensitivity. Mitochondria remain intact, uninjured, and in their natural environment where mitochondrial respiration can be measured in situ under physiologically relevant conditions. This approach furthermore allows measurement of toxin effects on individual mitochondrial complexes. Numerous compounds at varying concentrations can be screened for mitochondrial toxicity, while the site of mitochondrial inhibition can be determined simultaneously. We used this assay to investigate, in murine neuroblastoma (N-2alpha) cells, the mitochondrial inhibitory properties of the parkinsonian-inducing proneurotoxin, 1-methyl-4 phenyl-1,2,3,6-tetrahydropyridine (MPTP), and its neurotoxic monoamine oxidase-B (MAO-B)-generated metabolite, the 1-methyl-4-phenylpyridinium species (MPP(+)). Within the time frame of each measurement (15 min), MPTP (< or = 1 mM) did not interfere with in situ mitochondrial respiration. As expected, MPP(+) was found to be a potent Complex I inhibitor but surprisingly also found to inhibit Complex IV. Optimized conditions for performing this assay are provided. PMID- 15656942 TI - Circular dichroism analysis of destruxins from Metarhizium anisopliae. AB - Destruxins are secondary metabolites secreted by Metarhizium anisopliae [Y. Kodaira, Toxic substances to insects, produced by Aspergillus ochraceus and Oopsra destructor, Agric. Biol. Chem., 25 (1961) 261-262. D.W. Roberts, Toxins from the entomogenous fungus Metarhizium anisoplaie: Isolation from submerged cultures, J. Invertebr. Pathol., 14 (1969) 82-88. D.W. Roberts, Toxins from the entomogenic fungi in microbial control of pest and plant disease, Academic press, New York, 1981, pp441-464.]. In recent research, other than being used as insecticides, destruxins exhibited great potential in therapeutical applications such as antitumor, antivirus, and animal cell immunization effectiveness, etc. In this study, the conformations purified destruxins were determined by circular dichroism (CD). The results indicated that these cyclic peptides have the type I beta-turn conformation. In addition, different types of destruxins exhibited different CD spectra in acetonitrile. Therefore, these characters can be used as fingerprints to identify each type of destruxin. To further investigate the interactions among destruxins, various combinations of destruxins in 10 mM phosphate-buffered saline (PBS) were also studied by CD. The results strongly suggested that destruxins might work independently in vivo. To our knowledge, this is the first report presenting the CD analysis of purified destruxins. PMID- 15656943 TI - Transcription of T7 DNA immobilised on latex beads and Langmuir-Blodgett film. AB - The recognition of DNA is the first and most important condition for biological applications, including transcription and translation regulators and DNA sensors. For this purpose, we have developed few systems where we were able to immobilize long double-stranded DNA (dsDNA) successfully to the surfaces of different solid substrates. To achieve this, we have chosen polystyrene beads and standard Langmuir-Blodgett monolayer of Zn-arachidate. In the first attempt, variant of T7 DNA containing one strong promoter A1 for Escherichia coli RNA polymerase was immobilised on uniform polystyrene microspheres (0.31 microm diameter) by covalent grafting. In the latter case, Zn(II) is bound to arachidic acid through charge neutralization. Since tetrahedral Zn(II) participates in DNA recognition through coordination, we have been able to layer DNA over the Zn-arachidate monolayer. The successful immobilization of DNAs on these different substrates was visualized under fluorescence microscope. These immobilized DNAs were used as a template to study in vitro transcription reaction and thus we introduce a new strategy for the study of transcription in heterogeneous phase. PMID- 15656944 TI - The inhibitory effect of pentobarbitone on reverse transcription-PCR. AB - Pentobarbitone sodium (Sodium 5-ethyl-5[1-methylbutyl]-pentobarbitone) is a short acting barbiturate that is commonly used to euthanase animals. As part of our studies into the molecular genetics of copper toxicosis in Bedlington terrier dogs, reverse-transcription (RT)-PCR was noted to always fail on RNA samples collected from livers of dogs sacrificed by pentobarbitone injection. When samples were collected without pentobarbitone, however, RT-PCR was always successful. We suspected the possible inhibition by pentobarbitone sodium of either reverse transcriptase or Taq polymerase. In vitro studies showed that pentobarbitone interference of PCR occurred at >4 microg/microl. To identify if pentobarbitone produced competitive inhibition, each components (Taq polymerase, MgCl(2), dNTP, etc.) of the PCR was individually altered. However, inhibition still persisted, suggesting that multiple PCR components may be affected. Also it was shown that pentobarbitone interference was not dependent on the PCR product size. Simple dilution of pentobarbitone contaminated DNA solutions, and the addition of bovine serum albumin (BSA) to the PCR mix overcame pentobarbitone interference. In vivo, PCR by pentobarbitone was found to be compounded by high DNA concentration and pentobarbitone contamination. In addition, both high DNA concentration and pentobarbitone contamination could be overcome through dilution and the addition of BSA. Further work is required to quantify pentobarbitone concentration in the liver-extracted DNA and RNA samples before this inhibition effect on PCR can be fully elucidated. PMID- 15656945 TI - Analytical and semipreparative resolution of enatiomers of albendazole sulfoxide by HPLC on amylose tris (3,5-dimethylphenylcarbamate) chiral stationary phases. AB - Broad spectrum anthelmintic agent-albendazole sulfoxide (ABZSO) have been separated and semiprepared on amylose tris (3,5-dimethylphenylcarbamate) chiral stationary phases by HPLC using mobile phases contained with n-hexane and different alcohols. For analytical separation the influence of the nature and content of alcoholic modifiers on separation were systemically studied. Then, the analytical methods were scaled up to semipreparative loading to obtain small quantities (about 1 g) of both ABZSO enantiomers. Especially, different loading amounts were investigated for their effect on various parameters of semipreparative HPLC. In addition, optical rotation and circular dichroism (CD) of both ABZSO enantiomers collected were determined and single enantiomers were found stable in configuration for 1 year. PMID- 15656946 TI - Polyvinylferrocenium modified Pt electrode for anaerobic glucose monitoring. AB - An amperometric enzyme electrode for the determination of glucose under anaerobic solution conditions was developed by immobilizing glucose oxidase and then by adsorbing ferrocene in polyvinylferrocenium matrix coated on a Pt electrode surface. The amperometric response due to the electrooxidation of ferrocene that the reduced flavin adenine dinucleotide centers of glucose oxidase was measured at a constant potential. The response characteristics of the enzyme electrode were investigated. The effects of the thickness of the polymeric film, the amount of the enzyme immobilized, the amount of the mediator, the glucose concentration, the applied potential, operating pH and temperature on the response of the enzyme electrode were studied. The response time and the optimum pH were found to be 30 40 s and pH 7.4 at 25 degrees C, respectively. The linear response was observed up to 5.0 mM glucose concentration that the produced detectable current was 0.0075 mM glucose concentration. The activation energy (E(a)) of immobilized enzyme reaction was calculated to be 41.3 kJ mol(-1) from the Arrhenius plot. The apparent Michaelis-Menten constant (K(Mapp)) was found to be 6.05 mM glucose according to the Lineweaver-Burk graph of the Michaelis-Menten equation under the optimum conditions. The interference signal due to the most common electrochemical interfering species was also evaluated. PMID- 15656947 TI - Congenital salivary gland anlage tumor: a case series and review of the literature. AB - OBJECTIVE: To understand the clinical presentation and management of salivary gland anlage tumor (SGAT). DESIGN AND METHODS: This case series includes a report of a newborn male who presented with acute airway obstruction secondary to a nasopharyngeal mass which was discovered in the course of the clinical evaluation. Six additional cases of SGAT from the pathology consultation files of one of the authors (LPD) presenting in similar fashion are also included. The relevant literature from 1966 to the present has been reviewed through a Medline keyword search utilizing terms "salivary gland anlage tumor", "neonatal", and "nasopharynx." RESULTS: Endoscopic evaluation identified a nasopharyngeal mass tethered to the posterior septum. Although CT and MRI were helpful in identifying the mass and excluding involvement of the surrounding structures, the imaging characteristics of the mass itself were nonspecific. The patient was taken to the operating room and the polypoid mass was removed transorally after lysis of its septal attachment. Pathologic examination revealed a SGAT, a recently described entity in neonates and young infants, who present with early onset respiratory distress. Since the initial report of nine cases by one of the co-authors (LPD), seven additional cases including the present one have been seen in consultation. CONCLUSIONS: Salivary gland anlage tumor of the nasopharynx is a rare cause of neonatal airway obstruction. Endoscopic evaluation and imaging studies are helpful in the exclusion of other etiologies, some of which may have intracranial extension. Simple excision has been curative to date. There have been no reported recurrences in any of the previously studied cases with clinical follow-up dating more than 5 years. PMID- 15656948 TI - Long-term data on children implanted with a short electrode array. AB - Ossification and cochlear malformations are no longer seen as a contra-indication to cochlear implantation. The MED-EL COMBI 40+ short electrode was designed specifically for cases where major ossification has occurred, or where full insertion of an electrode may not be possible due to abnormal structure of the cochlea. This study investigates outcomes of 18 children implanted with the short electrode array. These children were assessed using the EARS test battery pre operatively and at a number of intervals thereafter. Results show a consistent improvement in time on most tests; these results appear to be independent of aetiology. Data from these children were compared to 18 matched pairs implanted with the standard COMBI 40+ electrode array. The short electrode children do not perform as well as the standard children initially, but do tend to catch-up at later test intervals. Results indicate the benefit of using a short electrode when complete insertion of the standard electrode is not viable. PMID- 15656950 TI - Young deaf children with hearing aids or cochlear implants: early assessment package for monitoring progress. AB - BACKGROUND: Very few assessment measures exist for evaluating progress in young deaf children with hearing aids and cochlear implants. OBJECTIVE: To introduce and describe an early assessment package that covers auditory perception, communication/language development, and speech production in very young deaf children. MAIN OUTCOME MEASURES: Seven of the assessment measures (Listening Progress Profile, Categories of Auditory Performance, Tait Video Analysis, Stories-Narratives Assessment Procedure, Profile of Actual Linguistic Skills, Speech Intelligibility Rating, and the Profile of Actual Speech Skills) have been specifically developed at the Nottingham Cochlear Implant Programme, and a further one (Meaningful Auditory Integration Scale) was modified for use within the package. Moreover, two commercially available tests (Pragmatics Profile of Everyday Communication Skills and Preschool Language Scale) are included to complete the package. METHODS: The present paper describes each measure, how to use it, and its time frame. In addition, two case studies demonstrate the usefulness of the package as a whole. RESULTS AND CONCLUSIONS: The Nottingham Early Assessment Package (NEAP) offers a framework with which to assess in young deaf children the use of audition and language and communication in real-life situations. Being simple, reliable, and time effective can be used in everyday clinical practice. NEAP is innovative in design and offers a structured approach to monitor very young deaf children, both in short and long term. In addition, it allows the identification of additional problems and areas of difficulty as well as specific abilities and skills. This enables the clinician to determine appropriate intervention strategies. PMID- 15656949 TI - Hearing impairment in Dutch patients with connexin 26 (GJB2) and connexin 30 (GJB6) mutations. AB - OBJECTIVE: Despite the identification of mutations in the connexin 26 (GJB2) gene as the most common cause of recessive nonsyndromic hearing loss, the pattern of hearing impairment with these mutations remains inconsistent. Recently a deletion encompassing the GJB6 gene was identified and hypothesized to also contribute to hearing loss. We hereby describe the hearing impairment in Dutch patients with biallelic connexin 26 (GJB2) and GJB2+connexin 30 (GJB6) mutations. METHODS: The audiograms of patients who were screened for GJB2 and GJB6 mutations were analysed retrospectively. Standard statistical testing was done for symmetry and shape, while repeated measurement analysis was used to assess the relation between mutation and severity. Progression was also studied via linear regression analysis. RESULTS: Of 222 hearing-impaired individuals, 35 exhibited sequence variations; of these 19 had audiograms for study. Hearing loss in patients with biallelic "radical" (i.e. deletions, nonsense and splice site) mutations was significantly worse than in the wild type and heterozygotes (SAS proc GENMOD, p=0.013). The presence of at least one missense mutation in compound heterozygotes tends to lead to better hearing thresholds compared to biallelic radical mutations (p=0.08). One patient with the [35delG]+[del(GJB6-D13S1830)] genotype was severely impaired. Non-progressive hearing impairment was demonstrated in five 35delG homozygotes in individual longitudinal analyses. However a patient with the [299A>C]+[416G>A] genotype showed significant threshold progression in the lower frequencies. Findings on asymmetry and shape were inconclusive. CONCLUSIONS: Our data support the hypothesis that severity is a function of genotype and its effect on the amino acid sequence. A bigger cohort is required to establish non-progressivity more definitively. PMID- 15656951 TI - Effects of inhalant anesthesia: tympanometry validation (R98/217). AB - OBJECTIVES: The purpose of this study was to determine the best time to assess middle ear status with tympanometry for tympanometric instrument validation. The research question addressed three logical times for tympanometric reading prior to myringotomy. METHODS: Fifty-one children, ranging in age from nine months to 10 years, were recruited for this investigation. Participants underwent inhalant anesthesia for myringotomy and grommet placement with or without adenoidectomy and/or tonsillectomy. Tympanometry data was analyzed across three time periods. RESULTS: Findings indicated that tympanometric readings prior to anesthesia produced the most accurate results. CONCLUSIONS: Implicated in findings, in order to ensure that tympanometric instruments are reliable, typanometric measurements should be taken prior to the administration of any anesthesia. PMID- 15656952 TI - Prevention of gentamicin induced ototoxicity by trimetazidine in animal model. AB - OBJECTIVE: To show the efficacy of intra-peritoneally administered trimetazidine to prevent gentamicin ototoxicity, which is still an important cause of profound deafness among children in different parts of the world. METHODS: Two groups of Swiss albino mice received daily intra-muscular injections of gentamicin for 30 days. One of the groups received trimetazidine intra peritoneally in addition to the gentamicin. Auditory thresholds of the animals were measured by evoked brain stem response at the beginning and the end of the study. Results were compared to the results of the control group, which received intra peritoneal saline injections. RESULTS: Both groups receiving gentamicin injections had significant auditory threshold shifts, but in the group receiving additional trimetazidine, the threshold shift was not statistically significant when compared to control group. Threshold shift in gentamicin group significantly differed from that of the control group (p=0.0001) and gentamicin+trimetazidine group (p=0.0001), on the other hand there was no statistically significant difference between control group and trimetazidine+gentamicin group (p=0.102). CONCLUSION: Gentamicin ototoxicity can be prevented by intra peritoneal trimetazidine injections in animal model. This treatment modality may be a mode of protection from gentamicin ototoxicity in children. PMID- 15656953 TI - Treatment results of 59 young patients with nasopharyngeal carcinoma. AB - PURPOSE: To evaluate the outcome of patients 17 (90.5% versus 64.7%; p=0.02). Patients aged <16 years had longer 5 year DMFS (90.5% versus 67.7%; p=0.05) with a borderline significance. DMFS is significantly shorter in patients initially presented with N3 disease compared to N0-2 disease (58.8% versus 84.4%; p=0.04). CONCLUSIONS: We suggest that RT combined with multiagent chemotherapy is effective in achieving satisfactory DFS and comparable OS in young patients with NPC. Results of both previous series and ours are evoking the necessity of early administration of efficient chemotherapeutic agents in addition to radiotherapy, but indistinct is the precise regimen and the intensity along the radiotherapy. All efforts towards decreasing late side effects of treatment should be encouraged in this long life expected group. PMID- 15656954 TI - Pure tone audiometry in children. AB - OBJECTIVE: Our purpose was to evaluate performance of the children on pure tone audiometry (PTA), and role of age, concentration level (CL) and otitis media with effusion (OME) in testing process. METHODS: This study was prospectively performed on 100 consecutive children referred for PTA between 24 and 120 months of age. After the audiologists' decision on test technique, the followings were noted: CL (by using a visual analog scale), test duration, test convenience (TC) and the problems (interruption, delay, crying) during the test. By using pure tone averages (PTAv), mean hearing level (MHL) and mean gap (MG) were calculated. RESULTS: PTA was achieved in 90% of the children and PTAv of bone and air conduction in the both ears were obtained in 86% (play audiometry with earphones (PAE): 73%, conventional audiometry (CA): 22%). These ratios were not different between the children with and without OME. About 55% of the younger children than 36 months were failed in PTA, while this ratio was 4.5% in the children between 36 and 120 months. Only difference between PAE and CA tests was in MHL. In OME group, only TC was found to be lower. Correlation analysis showed that CL was correlated with age. Test duration was correlated with MG and MHL, but found to be dependent on only MG. Further, it was found that test choice of the audiologist was correlated with age and CL, but dependent on only CL. CONCLUSIONS: PTA is available in children depending on their age and particularly CL; hence, performance in PTA is directly related with audiologists' assessment the children and cooperation before starting the test. Meanwhile, test duration appears to be directly associated with the work on detection of the gap between air and bone conduction thresholds. PMID- 15656955 TI - Pediatric video laryngo-stroboscopy. AB - OBJECTIVE: Laryngo-stroboscopy remains as the clinical gold standard for assessing properties of the glottal phonatory and valvular function. This includes deficits of closure as well as mucosal wave irregularities secondary to abnormal zones of pliability and symmetry. Per-oral stroboscopy has technical limitations in children due to the size of the telescope and issues of patient compliance. However, flexible laryngoscopy is readily performed in newborns and young children. This paper describes the use of a new trans-nasal, digital flexible laryngoscope, which allows for laryngo-stroboscopy in children. METHODS AND RESULTS: A prospective longitudinal series was done on 25 children ages 19 months-13 years (mean age, 7.0 years) with this new technology. All 25 were successfully examined. CONCLUSIONS: New technological advancements in the design of digital flexible endoscopes has allowed for laryngo-stroboscopy, and therefore, provides the potential for expanding the population of children with vocal disorders in whom stroboscopic imaging is possible. Larger studies will be necessary to determine its limitations related to age, development, and disease. As the study of pediatric voice disorders continues to evolve, accurate diagnosis is essential to apply state of the art non-operative and phonosurgical interventions. Further longitudinal studies are currently underway to continue to refine techniques of pediatric voice assessment and to define limitations of this new technology. PMID- 15656956 TI - Assessment of information processing in children with functional articulation disorders. AB - OBJECTIVE: The aim of this study was to investigate the information processing skills in children who have functional articulation disorder by using a cognitive linguistic test battery. MATERIAL-METHODS: Thirty-three children with functional articulation disorder were enrolled in this study. They ranged in age from 6 to 10 years. A control group without articulation disorders was composed of 160 children with similar properties. In the first stage of this study, both groups were administered the Ross Information Processing Assessment Test-Primary. In this test, the scores obtained from eight subtests; namely immediate memory, recent memory, recall of general information, spatial orientation, temporal orientation, organization, problem solving and abstract reasoning, was calculated for both groups. The scores of the eight subtests are combined to form the four composites. These composites are memory quotient, orientation quotient, thinking and reasoning quotient and information processing quotient. Information processing quotient is the best and the most comprehensive estimate of a child's overall information processing ability. In the second stage of the study the articulation-disordered group was divided into two subgroups according the number of the mistakes, which was done at the level of the phoneme. The scores obtained from both subgroups were compared with the scores of the control group. RESULTS: The overall statistical analysis of the scores revealed that; the study group had significantly lower scores than the control group from memory quotient, thinking and reasoning quotient and information processing quotient. When compared to control group, the subgroup who cannot pronounce one phoneme had similar scores from all four composites. Moreover, the children who cannot pronounce multiple phonemes had significantly lower scores from memory quotient, thinking and reasoning quotient and information processing quotient. The results obtained from this study seem to be suggesting that information processing skills of children with functional articulation disorder are significantly low as compared to normal children. CONCLUSIONS: These results are revealing that the information processing skills of children with functional articulation disorder should be investigated in a detailed manner. According to the results obtained from this investigation these children should be put on deficit oriented education programs in addition to articulation therapy. PMID- 15656958 TI - The clinical significance of adenoid-choanae area ratio in children with adenoid hypertrophy. AB - OBJECTIVE: We designed a method to compare the adenoid-choanae area ratio (A/C ratio) measured in children with symptoms of adenoid hypertrophy with those from children without any symptoms related to adenoid hypertrophy. METHODS: A prospective analysis of A/C ratio of 42 patients undergoing adenoidectomy and tonsillectomy from January 2000 to September 2003 in a medical center was compared with 28 patients receiving treatment other than the above-mentioned procedures in the same period. We used 0 degrees telescope to take pictures of the nasopharynx under general anesthesia preoperatively. The pictures were transformed digitally. The A/C ratio was calculated by the program we developed after these pictures were processed by the Photoshop 6.0 (Adobe, USA) software. RESULTS: The A/C ratio in adenoid hypertrophy group (0.863+/-0.0983) is significantly higher (p<0.001) than that of control group (0.445+/-0.1431). Mouth breathing, nasal obstruction and snoring were found to be more severe in children with high A/C ratio (p<0.001). CONCLUSIONS: The A/C ratio provided the two dimensional information of the nasopharyngeal airway. The A/C area ratio is significantly higher in the group of symptomatic adenoid hypertrophy when compared with symptomatic-free group. For practical purposes, the adenoid with an A/C ratio more than 2/3 is considered a pathological enlargement. PMID- 15656957 TI - Measuring the correlation between adenoidal-nasopharyngeal ratio (AN ratio) and tympanogram in children. AB - OBJECTIVE: To evaluate the correlation between adenoidal-nasopharyngeal ratio (AN ratio) and tympanogram in children. STUDY DESIGN: A prospective clinical study from June 2002 to May 2003. METHOD: A total of 64 children, aged 6-9 years who presented with nasal obstruction, snoring, mouth breathing, and hyponasal speech were examined and AN ratio was calculated by using the lateral neck radiograms and compared with the tympanometric values. The relationship between AN ratio and middle-ear pressure was evaluated, regarding the AN ratio of 0.71. The chi-square test was used to analyze the correlation between AN ratio and middle ear pressures and Wilcoxon test was used to compare the changes between the mean AN ratio values, and mean middle ear pressures (including A and C type tympanograms) before and after medical therapy. RESULTS: Middle-ear effusions and C type tympanograms in impedance audiometry were both related to eustachian tube dysfunction resulting from enlargement of the adenoids with AN ratios higher than 0.71. Middle ear pressures were found lower in children with AN ratio greater than 0.71 than in children AN ratio less than 0.71 and the difference was highly significant (p<0.001). Although medical treatment of large adenoids was rather effective to shrink the adenoid tissue (p<0.001), it did not cause a statistically significant change in tympanometric values (p>0.05). CONCLUSIONS: Antibiotherapy is effective in reducing adenoid size without signs and symptoms of infection. The reduction of the adenoids in size after 3 weeks of antibiotherapy has an positive effect on recovery of eustachian tube function but is not sufficient in patients with middle ear effusion. Early ventilation tube insertion may be an alternative therapy for the middle ear effusions not improving by 3 weeks medical therapy. PMID- 15656959 TI - The validity of the OSA-18 among three groups of pediatric patients. AB - OBJECTIVE: To compare the signs and symptoms of obstructive sleep apnea syndrome in three groups of pediatric patients; solid organ transplant recipients, healthy children, and children with leukemia; in order to examine the effects of chronic illness on the obstructive sleep apnea-18-item questionnaire and to investigate its validity as a screening tool for obstructive sleep apnea in the pediatric solid organ transplant population. METHODS: In this cross-sectional study, there were two hundred and six subjects; 46 kidney transplant recipients, 59 liver transplant recipients, 34 patients with leukemia, and 67 healthy children. Adenotonsillar enlargement was assessed by using the obstructive sleep apnea-18 item questionnaire and by performing a focused physical examination of the oral and nasal cavity at the time of the child's routine visit in either the transplant clinic, outpatient oncology center, or general pediatric clinic. RESULTS: Comparison of questionnaire scores amongst the three groups showed significant differences between the healthy children and liver transplant recipients as well as those with leukemia. There was a significant difference in the physical examination scores of the children with leukemia as compared to the other groups. CONCLUSIONS: Adenotonsillar enlargement in pediatric transplant recipients can be an early indication of post-transplantation lymphoproliferative disorder. However, the prevalence of adenotonsillar enlargement in the transplant population does not appear to differ from that of the healthy population. Additionally, scores on the OSA-18 in the transplant population were confounded by chronic illness. Further prospective studies need to be performed to develop a screening tool to identify transplant recipients at risk for post-transplantation adenotonsillar lymphoma. PMID- 15656961 TI - Malformations of the first branchial cleft: duplication of the external auditory canal. AB - OBJECTIVE: Malformations of the first branchial cleft are uncommon and only sporadic reported in the literature. The may be present as a swelling or inflammatory opening on the neck, blind cysts or fistula associated with the external auditory canal. In this retrospective study, clinical features, diagnostic and therapeutic pitfalls are described in nine pediatric cases. PATIENTS AND RESULTS: Between 1998 and 2002, duplication of the external auditory canal were diagnosed in nine patients aged from 7 months to 14 years. Three infants had cysts in the external ear canal; one had a sinus on the anteroinferior canal wall. One girl had two canals divided by skin and both were blind ended. Three children had clinically swelling or abscess formation with persistent drainage below the earlobe or on the neck and one girl had swelling behind the auricle. All patients were treated surgically. CONCLUSION: The first branchial cleft malformation may be unrecognized or may be mistaken for tumors or other inflammatory lesions in the periauricular region. Surgical treatment might then be inadequate leading to recurrence or secondary infection. The distinct clinical features, which can be derived from embryologic development, usually lead to the correct diagnosis and favorable surgical treatment. PMID- 15656960 TI - An unusual case of rhabdomyosarcoma presenting as orbital apex syndrome. AB - PRECIS: A 12-year-old female presented with symptoms and signs of orbital apex syndrome (OAS), secondary to stage IV alveolar rhabdomyosarcoma (RMS) originating in the sphenoid and ethmoid sinuses. OBJECTIVE: To present a case of alveolar rhabdomyosarcoma, unusual in its presentation as orbital apex syndrome and also its origin from the sphenoid and ethmoid sinuses. DESIGN: : Observational case report. METHODS: Ophthalmologic findings, neuroimaging, medical and surgical intervention, histopathologic analysis, and clinical course are described. RESULTS: A 12-year-old female presented with progressive visual loss in her left eye, difficulty with eye movements, and mild headache. Her examination was consistent with orbital apex syndrome. Imaging with contrast revealed a mass originating in the left sphenoid and ethmoid sinuses invading the left optic canal. Emergent biopsy was interpreted as alveolar rhabdomyosarcoma; subsequent metastatic work-up revealed bone marrow metastases. The patient was diagnosed with stage IV alveolar rhabdomyosarcoma and immediately started on combination orbital radiation therapy (RT) and systemic chemotherapy. She experienced gradual improvement of ocular motility, though her optic neuropathy persisted. CONCLUSION: Alveolar rhabdomyosarcoma of paranasal origin, specifically from the sphenoid and ethmoid sinuses, should be included in the differential diagnosis for orbital apex syndrome in children. PMID- 15656962 TI - Tracheostomy in children with nemaline core myopathy. AB - Two children with nemaline core myopathy (NM) who required tracheostomy are reported. One had a severe neonatal form requiring tracheostomy in the neonatal period for continuous mechanical ventilation. The other had a milder form with obstructive sleep apnoea, who underwent tracheostomy at age 22 months after adenoidectomy and a trial of nocturnal nasal continuous positive airway pressure ventilation. These cases highlight the clinical heterogeneity of the condition and the important role of the otolaryngologist in its management. PMID- 15656963 TI - Rhabdomyosarcoma presenting as an anterior neck mass and possible thyroid malignancy in a seven-month-old. AB - A seven-month-old male presented with a one month history of an enlarging left neck mass and worsening inspiratory stridor. Upon excision of the mass, pathologic examination was consistent with embryonal rhabdomyosarcoma (RMS). Preoperative imaging and intraoperative exploration were consistent with tumor replacing the left lobe of the thyroid. No cases of either anterior neck rhabdomyosarcoma or thyroid rhabdomyosarcoma have been explicitly described in the literature. The distinction between the two malignancies becomes important when considering prognosis and treatment protocols. PMID- 15656964 TI - Left ventricular hypertrophy with outflow tract obstruction-a complication of dexamethasone treatment for subglottic stenosis. AB - To our knowledge, this is the 1st reported case of steroid-induced obstructive cardiomyopathy in a child being treated for subglottic stenosis. As well, although born at 27 weeks, our patient was over 4 kg and 4 months of age (44 weeks corrected gestational age). This appears to be a phenomenon chiefly involving premature babies in the neonatal period with no reports of this occurring in patients greater than 40 weeks corrected gestational age. Signs of cardiac hypertrophy include tachycardia, new cardiac murmur, increased oxygen requirements, decreased urine output, and decreased peripheral perfusion. Diagnosis and eventual recovery was confirmed with serial echocardiograms. Knowledge of this serious side effect of dexamethasone will allow otolaryngologists to intervene early and prevent a potentially deadly complication. PMID- 15656965 TI - Sphenoid sinusitis disguised by precocious puberty. AB - Isolated sphenoid sinusitis in childhood is a rare clinical entity which is often very difficult to diagnose, hence, it can easily be misdiagnosed. The rarity of this entity is related to the absence at the sinus in children and the lack of development in patients with sinopathology. The first presentation of this disorder might be disguised by another disease. We present a case sphenoid sinusitis in ten years old girl where precocious puberty and slight headache were the main symptoms. Surgical drainage and antimicrobial treatment were administered. Clinical features, diagnostic tools and treatment options for this entity are discussed. PMID- 15656966 TI - Unusual association of congenital middle ear cholesteatoma and first branchial cleft anomaly: management and embryological concepts. AB - OBJECTIVE: To report two cases of an undescribed association of first branchial cleft fistula and middle ear congenital cholesteatoma and to discuss management and embryological hypothesis. DESIGN: Retrospective study and review of the literature RESULTS: Both patients were young girls free of past medical or surgical history. Surgical removal of the first cleft anomaly found in the two cases a fistula routing underneath the facial nerve. Both cholesteatomas were located in the hypotympanum, mesotympanum. In one case, an anatomical link between the two malformations was clearly identified with CT scan. DISCUSSION: The main embryological theories and classification are reviewed. A connection between Aimi's and Michaels' theories (congenital cholesteatoma) and Work classification might explain the reported clinical association. PMID- 15656968 TI - Function of alternative splicing. AB - Alternative splicing is one of the most important mechanisms to generate a large number of mRNA and protein isoforms from the surprisingly low number of human genes. Unlike promoter activity, which primarily regulates the amount of transcripts, alternative splicing changes the structure of transcripts and their encoded proteins. Together with nonsense-mediated decay (NMD), at least 25% of all alternative exons are predicted to regulate transcript abundance. Molecular analyses during the last decade demonstrate that alternative splicing determines the binding properties, intracellular localization, enzymatic activity, protein stability and posttranslational modifications of a large number of proteins. The magnitude of the effects range from a complete loss of function or acquisition of a new function to very subtle modulations, which are observed in the majority of cases reported. Alternative splicing factors regulate multiple pre-mRNAs and recent identification of physiological targets shows that a specific splicing factor regulates pre-mRNAs with coherent biological functions. Therefore, evidence is now accumulating that alternative splicing coordinates physiologically meaningful changes in protein isoform expression and is a key mechanism to generate the complex proteome of multicellular organisms. PMID- 15656969 TI - Of Fox and Frogs: Fox (fork head/winged helix) transcription factors in Xenopus development. AB - Transcription factors of the Fox (fork head box) family have been found in all metazoan organisms. They are characterised by an evolutionary conserved DNA binding domain of winged helix structure. In the South African clawed frog, Xenopus laevis, more than 30 Fox genes have been found so far. This review summarises our present knowledge regarding the general structure and common features of the fork head box and will then characterise Fox genes that have been described in Xenopus. Special attention was paid to the temporal and spatial expression patterns during early embryonic development. For some of these genes, the molecular mechanisms leading to their regulation after the onset of zygotic transcription are known. We also report on functional aspects including target gene regulation, cell or tissue specification and interference with the cell cycle. Finally, Fox proteins serve as mediators of signalling pathways and they might function as checkpoint molecules for the cross-regulatory interactions of different intracellular signal transduction chains. PMID- 15656970 TI - Analysis of 101 nuclear transcriptomes reveals 23 distinct regulons and their relationship to metabolism, chromosomal gene distribution and co-ordination of nuclear and plastid gene expression. AB - Post-endosymbiotic evolution of the proto-chloroplast was characterized by gene transfer to the nucleus. Hence, most chloroplast proteins are nuclear-encoded and the regulation of chloroplast functions includes nuclear transcriptional control. The expression profiles of 3292 nuclear Arabidopsis genes, most of them encoding chloroplast proteins, were determined from 101 different conditions and have been deposited at the GEO database (http://www.ncbi.nih.gov/geo/) under . The 1590 most-regulated genes fell into 23 distinct groups of co-regulated genes (regulons). Genes of some regulons are not evenly distributed among the five Arabidopsis chromosomes and pairs of adjacent, co-expressed genes exist. Except regulons 1 and 2, regulons are heterogeneous and consist of genes coding for proteins with different subcellular locations or contributing to several biochemical functions. This implies that different organelles and/or metabolic pathways are co-ordinated at the nuclear transcriptional level, and a prototype for this is regulon 12 which contains genes with functions in amino acid and carbohydrate metabolism, as well as genes associated with transport or transcription. The co-expression of nuclear genes coding for subunits of the photosystems or encoding proteins involved in the transcription/translation of plastome genes (particularly ribosome polypeptides) (regulons 1 and 2, respectively) implies the existence of a novel mechanism that co-ordinates plastid and nuclear gene expression and involves nuclear control of plastid ribosome abundance. The co-regulation of genes for photosystem and plastid ribosome proteins escapes a previously described general control of nuclear chloroplast proteins imposed by a transcriptional master switch, highlighting a mode of transcriptional regulation of photosynthesis which is different compared to other chloroplast functions. From the evolutionary standpoint, the results provided indicate that functional integration of the proto-chloroplast into the eukaryotic cell was associated with the establishment of different layers of nuclear transcriptional control. PMID- 15656971 TI - Drag&Drop cloning in yeast. AB - We have developed a set of vectors that have enhanced capabilities for efficiently constructing and expressing differentially tagged fusion proteins using Drag&Drop cloning in the yeast Saccharomyces cerevisiae. The pGREG vectors are based on the pRS series with an additional general kanR selection marker. In vivo homologous recombination is used to introduce genes of interest into galactose-inducible expression vectors (pGREGs), permitting the formation of amino-terminal fusions. The vectors all contain common regions for recombination that flank the stuffer fragment. Introduction of common recombination sequences at the end of PCR fragments will permit the cloning of genes without the need for specific restriction sites. In this process, the selectable stuffer HIS3 gene is replaced by successful gene integration, and a screen for loss of the selection marker identifies potential recombinants. Due to the modular structure of the vectors, genes introduced into one vector can be readily transferred by in vivo recombination to all other members of the vector system, thus permitting rapid and easy Drag&Drop construction of a series of tagged proteins. The pGREG series combines features for expression, tagging, integration, localization and library construction with the advantage of obtaining immediate results from sub-sequent experiments. This Drag&Drop system also allows efficient cloning and expression of heterologous genes in large-scale experiments. PMID- 15656972 TI - Isolation and characterization of the human Cdc2L1 gene promoter. AB - CDK11 (cyclin-dependent kinase 11, formerly known as PITSLRE) is a member of the p34cdc2-related kinases. It has been previously shown to be involved in a variety of different cellular processes including RNA processing, apoptosis, and cell cycle progression. It is encoded by two different but highly similar genes, Cdc2L1 (cell division control 2 like 1) and Cdc2L2 (cell division control 2 like 2). Previous studies from our group identified and characterized the transcriptional regulation of the human Cdc2L2 gene promoter. The current studies identify and characterize the Cdc2L1 gene promoter. We cloned the promoter and elucidated the different transcriptional regulatory elements that reside within the 5' region of the gene. Deletion analysis of the promoter showed a region of nucleotides -152 to +11 to be necessary for basal transcription of the Cdc2L1 gene. Sequencing analysis found this region of the promoter to be highly GC-rich but is lacking both TATA and CAAT boxes. There are several different transcription factor binding sites that are consensus or near consensus found within this region. The potential binding sites include two Ets-1 sites, one Skn 1 site, and one E2F-1 site. Transfection studies of various site-directed mutagenesis clones for these different sites revealed that both Ets-1 sites play critical roles in sustained transcriptional activity as well as Skn-1. Chromatin immunoprecipitation of the endogenous promoter with Ets-1 and Skn-1 verified an in vivo association of Ets-1 and Skn-1 transcription factors with the endogenous promoter. These results, in addition to our Cdc2L2 results, lead to the further comprehension of the fundamental mechanisms dictating CDK11 gene expression through the Cdc2L1 gene promoter. PMID- 15656973 TI - A new additional reporter enzyme, dinoflagellate luciferase, for monitoring of gene expression in mammalian cells. AB - Dinoflagellate luciferase (DL) catalyses the oxidation of dinoflagellate luciferin by molecular oxygen, resulting in an electronically excited species that emits blue light (lambda(max)=474 nm). Luciferase has three catalytic domains in its single polypeptide chain (M(r)=ca. 140 kDa), and each domain (about 40 kDa) is enzymatically active when expressed individually in recombinant fusion proteins in E. coli. Thus, DL should be useful as a reporter enzyme in studies of gene expression in mammalian cells. Expression plasmids consisting of one domain of luciferase (dDL) cDNA linked to different several promoters were introduced into a series of mammalian cell lines. Following transfection, dDL activities in cell extracts were determined by a rapid light emission assay of luciferase activity. For dual and multiple reporter assays, it is possible to exchange dDL for the firefly or renilla luciferases, and use the new luciferase for control or target reporter genes. Thus, the triple-reporter assay can identify three transcriptional activities of different genes at the same time. This work establishes the DL gene as a new efficient marker of gene expression in mammalian cells. PMID- 15656974 TI - Gene expression profiling of E2F-1-induced apoptosis. AB - It has been shown that adenovirus-mediated overexpression of E2F-1 can efficiently induce apoptosis in cancer cells with little effect on normal cells. However, the mechanisms by which E2F-1 induces apoptosis remains poorly understood. The goal of this study was to evaluate changes in gene expression in response to E2F-1 in order to help elucidate the mechanisms by which E2F-1 causes apoptosis. Therefore, we used a quantitative microarray assay to identify the genes regulated by E2F-1 in melanoma cells. By gene expression profiling, we first screened a proprietary list of about 12,000 genes. Overexpression of E2F-1 in melanoma cells resulted in two-fold or greater alteration in the level of expression of 452 genes compared to vehicle-treated control cells. Most of the affected genes were not known to be responsive to E2F-1 prior to this study. E2F 1 adenoviral infection of these cells was found to affect the expression of a diverse range of genes, including oncogenes, transcription factors and genes involved in signal transduction, cell cycle regulation, cell proliferation and apoptosis, as well as other genes with unknown function. Changes in expression of 17 of these genes were confirmed by quantitative real-time polymerase chain reaction (PCR). This is first application of the microarray technique in the study of the global profile of genes regulated by E2F-1 in melanoma cells. This study leads to an increased understanding of the biochemical pathways involved in E2F-1-induced apoptosis and possibly to the identification of new therapeutic targets. PMID- 15656976 TI - Functional XPB/RAD25 redundancy in Arabidopsis genome: characterization of AtXPB2 and expression analysis. AB - The xeroderma pigmentosum complementation group B (XPB) protein is involved in both DNA repair and transcription in human cells. It is a component of the transcription factor IIH (TFIIH) and is responsible for DNA helicase activity during nucleotide (nt) excision repair (NER). Its high evolutionary conservation has allowed identification of homologous proteins in different organisms, including plants. In contrast to other organisms, Arabidopsis thaliana harbors a duplication of the XPB orthologue (AtXPB1 and AtXPB2), and the proteins encoded by the duplicated genes are very similar (95% amino acid identity). Complementation assays in yeast rad25 mutant strains suggest the involvement of AtXPB2 in DNA repair, as already shown for AtXPB1, indicating that these proteins may be functionally redundant in the removal of DNA lesions in A. thaliana. Although both genes are expressed in a constitutive manner during the plant life cycle, Northern blot analyses suggest that light modulates the expression level of both XPB copies, and transcript levels increase during early stages of development. Considering the high similarity between AtXPB1 and AtXPB2 and that both of predicted proteins may act in DNA repair, it is possible that this duplication may confer more flexibility and resistance to DNA damaging agents in thale cress. PMID- 15656975 TI - Identification and expression analysis of alternative transcripts of the mouse GA binding protein (Gabp) subunits alpha and beta1. AB - The erythroblast transformation specific (ETS) transcription factor GA-binding protein (Gabp) is widely expressed and acts on a diverse range of target genes, including nuclear-encoded mitochondrial proteins and neuromuscular-specific genes. The GABPalpha subunit contains an ETS DNA binding domain and the beta subunit contains a nuclear localization signal (NLS) and transactivation domain. Here, we show coincident expression of Gabpalpha and beta1 throughout mouse embryogenesis, consistent with the gene products functioning in a complex. We have also identified 2 alternatively spliced, tissue-specific exons 1 (5' untranslated regions) of mouse Gabpalpha and 4 alternative 3' polyadenylation signals that, in combination, result in 12 transcripts for Gabpalpha. These alternative transcripts are suggested to have altered stability, subcellular localization and/or translation efficiency. Further, we identified nine differentially expressed splice variants of mouse Gabpbeta1 that encode beta protein forms lacking functional domains, suggesting a dominant negative function. Together, alternative transcripts of Gabpalpha and beta1 provide a mechanism for tissue-specific regulation of Gabp activity. PMID- 15656977 TI - Unusual structure of ribosomal DNA in the copepod Tigriopus californicus: intergenic spacer sequences lack internal subrepeats. AB - Eukaryotic nuclear ribosomal DNA (rDNA) is typically arranged as a series of tandem repeats coding for 18S, 5.8S, and 28S ribosomal RNAs. Transcription of rDNA repeats is initiated in the intergenic spacer (IGS) region upstream of the 18S gene. The IGS region itself typically consists of a set of subrepeats that function as transcriptional enhancers. Two important evolutionary forces have been proposed to act on the IGS region: first, selection may favor changes in the number of subrepeats that adaptively adjust rates of rDNA transcription, and second, coevolution of IGS sequence with RNA polymerase I transcription factors may lead to species specificity of the rDNA transcription machinery. To investigate the potential role of these forces on population differentiation and hybrid breakdown in the intertidal copepod Tigriopus californicus, we have characterized the rDNA of five T. californicus populations from the Pacific Coast of North America and one sample of T. brevicornicus from Scotland. Major findings are as follows: (1) the structural genes for 18S and 28S are highly conserved across T. californicus populations, in contrast to other nuclear and mitochondrial DNA (mtDNA) genes previously studied in these populations. (2) There is extensive differentiation among populations in the IGS region; in the extreme, no homology is observed across the IGS sequences (>2 kb) from the two Tigriopus species. (3) None of the Tigriopus IGS sequences have the subrepeat structure common to other eukaryotic IGS regions. (4) Segregation of rDNA in laboratory crosses indicates that rDNA is located on at least two separate chromosomes in T. californicus. These data suggest that although IGS length polymorphism does not appear to play the adaptive role hypothesized in some other eukaryotic systems, sequence divergence in the rDNA promoter region within the IGS could lead to population specificity of transcription in hybrids. PMID- 15656978 TI - Molecular and functional characterization of CaLEA6, the gene for a hydrophobic LEA protein from Capsicum annuum. AB - We used differential screening to isolate a full-length dehydration-responsive cDNA clone encoding a hydrophobic late embryogenesis abundant (LEA)-like protein from PEG-treated hot pepper leaves. Named CaLEA6 (for Capsicum annuum LEA), this gene belongs to the atypical hydrophobic LEA Group 6. The full-length CaLEA6 is 709 bp long with an open reading frame encoding 164 amino acids. It is predicted to produce a highly hydrophobic, but cytoplasmic, protein. The putative M(r) of CaLEA6 protein is 18 kDa, with a theoretical pI of 4.63. Based on our Southern blot analysis, CaLEA6 appears to exist as a small gene family. CaLEA6 was not expressed prior to any treatment, but its transcript was rapidly and greatly increased following trials with PEG, ABA, and NaCl. Chilling also induced its rapid induction, but to a much lesser extent. Accumulation of CaLEA6 protein occurred soon after NaCl applications, but considerably delayed after treatment with PEG. Tobacco plants that overexpressed CaLEA6 showed enhanced tolerance to dehydration and NaCl but not to chilling, as defined by their leaf fresh weights, Chl contents, and the general health status of the leaves. Therefore, we suggest that CaLEA6 protein plays a potentially protective role when water deficit is induced by dehydration and high salinity, but not low temperature. PMID- 15656979 TI - Apisalpha2, Apisalpha7-1 and Apisalpha7-2: three new neuronal nicotinic acetylcholine receptor alpha-subunits in the honeybee brain. AB - Acetylcholine is the principal excitatory neurotransmitter in the central nervous system of insects. Nicotinic acetylcholine receptors, which belong to the ligand gated ion channel family, constitute important targets for insecticides. In the honeybee Apis mellifera, pharmacological evidence supports the existence of several nicotinic acetylcholine receptors. In this paper, we report the identification of three new genes that encode nicotinic acetylcholine receptor alpha-subunits in the honeybee. Phylogenetic comparisons with other ligand-gated ion channel subunit sequences support their classification as Apisalpha2, Apisalpha7-1 and Apisalpha7-2 subunits. Based on in situ hybridization experiments, we determined their expression patterns in the different brain regions of pupae and adult honeybees. Our results show that these nicotinic acetylcholine receptor subunits are differently expressed among the brain regions and that they appear at different stages of honeybee development. PMID- 15656980 TI - The top 10 most abundant transcripts are sufficient to characterize the organs functional specificity: evidences from the cortex, hypothalamus and pituitary gland. AB - Using serial analysis of gene expression, we have identified the most abundant mRNA transcripts in parietal cortex, hypothalamus and pituitary gland in adult male mice. High mRNA abundance of neurogranin (cell signalling and communication) was characteristic of the cortex. The common molecular features of cortex and hypothalamus were high abundance of mRNA encoding mitochondrial enzymes such as reduced form of nicotinamide adenine dehydrogenase (NADH) 4 and cytochrome c oxidase 2 (energy metabolism), brain creatine kinase (energy metabolism) and myelin basic protein (cell structure). In the hypothalamus, mRNA levels of apolipoprotein E (lipid metabolism), prostaglandin D2 (cell signalling and communication) and secreted acidic cysteine-rich glycoprotein (extracellular matrix) were especially high. A common molecular feature of the hypothalamus and pituitary was high mRNA abundance of guanine nucleotide binding protein alpha stimulating complex locus (cell signalling and cell communication). The pituitary gland was characterized by high expression of genes encoding hormones such as growth hormone, pro-opiomelanocortin and prolactin, as well as neuronatin (cell differentiation) and four potential novel transcripts. Thus, these results show that the cortex, hypothalamus and pituitary gland can be specifically characterized according to their 10 most abundant transcripts. In addition, the current study serves as a basis for future studies on the potential novel transcripts and the transcripts with unclear functions despite their extremely high abundance, as well as studies on physiology and pathology of the two brain regions and pituitary gland. PMID- 15656982 TI - Characterization of the promoter of 1A6/DRIM, a novel cancer-related gene and identification of its transcriptional activator. AB - 1A6/DRIM (Down-regulated in Metastasis) has been reported to express at a high level in the gastric cancer tissues and the premalignant lesions implicating the involvement of 1A6/DRIM in cell transformation. Although the information regarding the putative functions and distribution of the 1A6/DRIM in different tissues and cell lines has been increasing recently, its promoter and promoter regulating factors remain unknown. In this study, the transcription initiation site of 1A6/DRIM was confirmed to be located at 147 bp upstream of the ATG codon using the primer extension analysis. The minimal promoter region of the 1A6/DRIM is located between -47 and +42 of the transcription initiation site measured by luciferase reporter assays using a set of deletion constructs. In addition, an E box is shown to be an essential element for transcriptional regulation of 1A6/DRIM demonstrated by luciferase assay with different deletion and mutation constructs. Finally, a transcription factor, upstream stimulatory factor 2 (USF2) was found to be an activator of the 1A6/DRIM through binding to the E-box demonstrated by luciferase reporter assay, electrophoretic mobility shift assay, and chromatin immunoprecipitation (ChIP) assay. The structural analysis of the 1A6/DRIM promoter and the identification of its potential regulatory effecter may help us to understand its biological functions in regulating cancer development. PMID- 15656981 TI - Dual promoter structure of ZFP106: regulation by myogenin and nuclear respiratory factor-1. AB - The WD40 repeats containing zinc finger protein 106 (ZFP106) is a conserved mammalian protein of unknown function. However, its cDNA shares an extended region of identity with the scr homology domain 3 binding protein 3 (Sh3bp3) cDNA encoding a protein implicated in the insulin signaling pathway. Asking, whether Zfp106 and Sh3bp3 are products of the same gene, we characterized the structures and transcriptional regulation of Zfp106 and its human homologue, ZFP106. A TATA less, CpG island associated promoter (P1), was mapped by 5'-RACE to a region 19 kb upstream of the ZFP106 translation start site. P1 is active throughout development and at low levels in all adult tissues examined. A conserved cis element in the proximal P1 region showed specific binding to nuclear respiratory factor-1 (NRF-1). Mutagenesis of this site and transfection of a dominant negative NRF-1 both revealed the crucial role of NRF-1 in activation of P1. The broad tissue expression of P1 was in contrast to the high level of ZFP106 mRNA observed in striated muscle. This prompted additional 5'-RACE experiments that established a second, TATA box-containing promoter (P2) upstream of the third coding exon. P1 and P2 transcripts encode proteins with distinct N-terminal sequences, with Sh3bp3 corresponding to a rare, alternatively spliced P2 transcript. P2 initiated transcripts are specifically expressed in striated muscle and their level is strongly upregulated during myogenic, but not adipogenic differentiation. By deletion analysis, the region between nucleotides 296 to +96 was sufficient for robust P2 responsiveness to myogenic differentiation. This response is mediated by the additive effect of binding of myogenin to three critical E boxes within this region. In addition, transcriptional enhancer factor-1 family factors contribute to both basal and myogenesis induced P2 activity. In situ hybridization of mouse embryos confirmed predominant expression of Zfp106 in tissues with high developmental expression of either NRF-1 (brown fat and developing brain) or myogenin (striated muscle). Our results suggest distinct roles of tissue-specific ZFP106 isoforms in growth related metabolism and provide the foundation for further studies into the regulation and function of ZFP106. PMID- 15656983 TI - The OsLti6 genes encoding low-molecular-weight membrane proteins are differentially expressed in rice cultivars with contrasting sensitivity to low temperature. AB - Rice (Oryza sativa L.) is sensitive to chilling particularly at early stages of seedling establishment. Two closely related genes (OsLti6a, OsLti6b), which are induced by low temperature during seedling emergence were isolated from a cold tolerant temperate japonica rice cultivar. These genes are closely related to the Arabidopsis rare cold-inducible (RCI2) and barley low-temperature-inducible (BLT101) genes. Based on direct biochemical and indirect physiological evidence and similarity with a conserved protein domain in the Cluster of Orthologous Groups (COG) database (e.g., yeast PMP3), the rice genes belong to a class of low molecular-weight hydrophobic proteins involved in maintaining the integrity of the plasma membrane during cold, dehydration and salt stress conditions. Both genes exhibit a genotype-specific expression signature characterized by early and late stress-inducible expression in tolerant and intolerant genotypes, respectively. The differences in temporal expression profiles are consistent with cultivar differences in cold-induced membrane leakiness and seedling vigor. The presence of CRT/DRE promoter cis-elements is consistent with the synchronized expression of OsLti6 genes with the C-repeat binding factor/drought responsive element-binding protein (CBF/DREB) transcriptional activator. The present results indicate that the Oslti6 genes are part of a battery of cold stress defense related genes regulated by a common switch. PMID- 15656984 TI - A W-box is required for full expression of the SA-responsive gene SFR2. AB - Transcripts of SFR2, a member of the S family of receptor kinase genes, accumulate rapidly in Brassica oleracea leaves in response to wounding, bacterial infection and following treatment with salicylic acid (SA). Expression of a chimeric gene consisting of the SFR2 5' flanking sequence fused to the gusA reporter gene is also induced in wounded and SA-treated Arabidopsis plants indicating that the observed response is conferred by the SFR2 promoter. We show here that, in Arabidopsis plants carrying the salicylate hydroxylase (NahG) transgene, wound induction of the SFR2 promoter-gusA reporter fusion was abolished, indicating that, as has previously been shown for the response to bacterial infection, SA is required for the response to wounding. Deletion analysis of the SFR2 promoter identified a region necessary for full expression following SA treatment. This region, which includes two putative W-boxes, is conserved in the promoter of the Arabidopsis SFR2 homologue, ARK3. Deletion of a 12 bp region containing the two W-box motifs reduced the response to SA treatment. Tandem repeats of the W-box-containing element fused upstream of a CaMV 35S minimal promoter enhanced reporter gene expression in transgenic Arabidopsis both in the absence and presence of SA. Gel-mobility shift assays showed that Arabidopsis leaf extracts contained factors that bound to a fragment of the promoter spanning the putative W-boxes and that a fragment in which these motifs were mutated was unable to compete for binding. In summary, induction of the SFR2 promoter in response to bacterial infection and wounding requires SA, and full expression of the induced gene requires the presence of a functional element containing W-box motifs in the SFR2 promoter. The involvement of two W boxes indicates that transcription factors of the WRKY family may play a key role in mediating these responses. PMID- 15656985 TI - Gene expression of Toll-like receptor-2, Toll-like receptor-4, and MD2 is differentially regulated in rabbits with Escherichia coli pneumonia. AB - Sepsis, a common sequela to Gram-negative pneumonia, results in considerable morbidity and mortality in hospitalized patients. The goal of this study was to determine whether Gram-negative pneumonia alters the expression TLR2, TLR4, and MD2 in lungs or in organs distant to the site of the primary infection. The cDNA sequence coding open reading frames for rabbit TLR2, TLR4, and MD2 were cloned and expressed in Escherichia coli, and specific polyclonal antibodies and polymerase chain reaction (PCR) probes were produced to identify changes in these receptors in rabbits with Gram-negative pneumonia. Using tissues from lungs and distant organs, we show that TLR2, TLR4, and MD2 gene expression is differentially regulated in rabbits with E. coli pneumonia. The increased expression of TLR2 and TLR4 could play an important role in the innate immune response to bacterial infection in the lungs, and improve pathogen recognition and bacterial clearance. In contrast, the increased gene expression of TLR2, TLR4, and MD2 in organs distant to the primary site of infection may contribute to the deleterious systemic inflammatory response observed in patients with sepsis. PMID- 15656986 TI - Short tandem repeat (STR) replacements in UTRs and introns suggest an important role for certain STRs in gene expression and disease. AB - Some untranslated sequence (UTR)-localized, short tandem repeats (STRs) exhibit evidence of selection pressure, including STR-coupling preferences, STR conservation, interspecies STR-STR replacements, and STR variants implicated in certain diseases. We wished to determine if STR replacements occurred near disease-related genes, including previously unstudied STRs as well as some STRs already implicated in disease. Among nine strong-candidate prostate cancer (CaP) predisposing genes, three [steroid 5-alpha-reductase 2 (Srd5A-2), macrophage scavenger receptor-1 (MSR-1), and tumor necrosis factor receptor-21 (Tnfr-21)] exhibited striking STR replacements (P<0.001). The glomerular disease-related gene, CD2AP, exhibited an STR replacement flanked by well-conserved sequences, suggesting an STR-focused process. Another glomerular disease-related gene, rabphilin 3A, exhibited at least two STR replacements at the same UTR position comparing Drosophila melanogaster, Mus musculus, and Homo sapiens. Two genes implicated in blood-clotting disorders, von Willebrand factor (vWA) and fibrinogen alpha (FGA), exhibited multiple-intron STR replacements among mammals, extending STR replacement phenomena to introns. Among primates, a tyrosine hydroxylase (THO1) intron STR, previously implicated in both schizophrenia and drug withdrawal delirium, exhibited frequent replacements. Some STR replacements were early events in gene divergence. When STR sequences of closely related species were available, STR replacement was observed to be nearly as rapid as speciation. STR replacements expand the list of STR sequences that may contribute to genetic activity and to disease processes. PMID- 15656987 TI - Effect of 3'UTR length on the translational regulation of 5'-terminal oligopyrimidine mRNAs. AB - In Vertebrates, all genes coding for ribosomal proteins, as well as those for other proteins implicated in the production and function of translation machinery, are regulated by mitogenic and nutritional stimuli, at the translational level. A cis-regulatory element necessary for this regulation is the typical 5'UTR, common to all ribosomal protein mRNAs, which always starts at the 5' end with several pyrimidines. Having noticed that the 3'UTR of all ribosomal protein mRNAs is much shorter than most cellular mRNAs, we have now studied the possible implication of this 3'UTR feature in the translational regulation. For this purpose, we constructed a number of chimeric genes whose transcribed mRNAs contain: (1) the 5'UTR of ribosomal protein S6 mRNA or, as a control, of beta-actin mRNA; (2) the EGFP reporter coding sequence from the starting AUG to the stop codon; (3) different 3'UTRs of various lengths. These constructs have been stably transfected in human HEK293 cells, and the translation regulation of the expressed chimeric mRNAs has been analyzed for translation efficiency, in growing and in serum starved cells, by the polysome association assay. The results obtained indicate that, while the typical growth associated translational regulation is bestowed on an mRNA by the pyrimidine sequence containing 5'UTR, the stringency of regulation depends on the short size of the 3'UTR. PMID- 15656988 TI - Dimerization of bacteriophage P2 integrase is not required for binding to its DNA target but for its biological activity. AB - Coliphage P2 integrates into the host chromosome upon lysogenization via site specific recombination mediated by the phage integrase (Int). P2 integrase belongs to the tyrosine family of recombinases. In this work, it is shown that P2 integrase forms dimers but not oligomers in the absence of its DNA target. Furthermore, the C-terminal end of the protein and amino acid (aa) E197 have been found to be involved in dimerization. Amino acid E197 is located in a conserved region of the tyrosine recombinases that has not previously been implicated in dimerization. The dimerization deficient mutants were unaffected in binding to its phage attachment site (attP) substrate, but had a reduced ability to complement an int-defective prophage. PMID- 15656989 TI - Expression of the mouse WNK1 gene in correlation with ganglioside GD3 and functional analysis of the mouse WNK1 promoter. AB - WNK1 is one of WNK (With No K=Lysine) protein kinases which comprise a newly described subfamily. Our studies showed that expression of the mouse WNK1 gene was dramatically suppressed in a tumor cell line when its phenotype was altered by suppression of the GD3-synthase gene expression. The mouse WNK1 gene was expressed at a high level at early stage of embryonic brain and its expression decreased as brain developed, similar to the expression pattern of the GD3 synthase gene. To study transcriptional regulation, we cloned a 5'-flanking 1239 bp fragment of the mouse WNK1 gene. This fragment contains a number of potential consensus binding sites for transcription factors, including Sp1, AP2, CCAAT, Est 1, Oct-1, CNBP, and NFkB, but lacks a TATA box. Primer extension identified multiple putative transcriptional initiation sites, including several sites downstream of the ATG codon. Activities of the promoter fragments were assessed in mouse breast Sa/R-MT cells by transient transfection and the results showed that the promoter elements between -700 and -977 is required for maintaining a high level of promoter activity of the TATA-less mouse WNK1 gene. PMID- 15656990 TI - Identification of two novel human genes, DIPLA1 and DIPAS, expressed in placenta tissue. AB - Here we report the identification and expression analysis of two novel human genes--DIPLA1 (Differentially expressed in placenta 1) and DIPAS (DIPLA1 Antisense). These genes are located at chromosomal region 9q33.1, in opposite orientations, and are flanked by the pregnancy-associated plasma protein-A (PAPP A) and astrotactin 2 (ASTN2) genes. The mRNA sequences of both genes contain several upstream AUGs (uAUG) and various potential open reading frames (ORFs). DIPLA1 mRNA is 1.8 kb long and contains a 285 nt ORF coding for a polypeptide designated as replicative senescence up-regulated (RSU) protein. Antisense DIPAS mRNA is 2.7 kb long and contains a 309 nt ORF coding for a protein with partial similitude to the gamma isoform variant of the human Ca(2+)/calmodulin (CaM) dependent protein kinase II. Both genes are conserved in placental-species and are presumably transcribed from initiator (Inr) promoter elements located at opposite strands. In 20 human normal tissues tested, DIPLA1 mRNA expression was placenta-specific, whereas DIPAS mRNA expression was higher in placenta, brain, kidney and testis. In addition, DIPAS mRNA hybridizes with the 3'UTR region from PAPP-A mRNA, which spans over 4 kb more than previously reported, forming a potential sense-antisense double stranded RNA (dsRNA) duplex. Our results are of interest for placenta gene expression regulation and for the identification of novel genes in the human genome. PMID- 15656991 TI - Identification of the gene for the monomeric alkaline phosphatase of Vibrio cholerae serogroup O1 strain. AB - Alkaline phosphatase (APase) of Vibrio cholerae is the first monomeric alkaline phosphatase reported [Roy, N.K., Ghosh, R.K., Das, J., 1982a. Monomeric alkaline phosphatase of V. cholerae. J. Bacteriol. 150, 1033-1039.]. The gene (phoA(VC)) encoding this enzyme is not identified in the published genome sequence of the V. cholerae serogroup O1 El Tor strain N16961 [Heidelberg et al., 2000, DNA sequence of both the chromosome of cholera pathogen V. cholerae. Nature 406, 477-484.]. However two genes (phoB(VC) and phoR(VC)) regulating the synthesis of alkaline phosphatase in this organism, equivalent to phoB and phoR of Escherichia coli, are located in tandem on chromosome I of V. cholerae. We have identified the phoA(VC) gene on the N16961 genome sequence by amino acid sequence analysis of the purified alkaline phosphatase of V. cholerae classical strain 569B followed by BLAST search. The gene was found to be located on the hypothetical protein locus VCA0033 of chromosome II. The identity of the gene was confirmed by expressing the cloned VCA0033 locus in phoA mutant E. coli E15 and JC9223 cells and isolating V. cholerae monomeric alkaline phosphatase. Insertional inactivation of the gene also resulted in complete loss of the phenotype. Unlike in E. coli where phoB, phoR and phoA are closely linked, phoA(VC) is not linked to phoB(VC) and phoR(VC). PMID- 15656993 TI - Biochemical characterization of the mammalian Cux2 protein. AB - The CCAAT displacement protein (CDP) and cux (Cut homeobox) genes were originally identified as the human and mouse orthologs of Drosophila melanogaster cut. More recently, vertebrates were found to possess a second cut orthologs that was generated by gene duplication: Cux2. We report the initial biochemical characterization of the Cux2 protein in tissue culture and in vitro. We generated four polyclonal antibodies that were able to recognize the human and mouse Cux2 protein but displayed little or no cross-reactivity towards CDP1 and Cux1. The expression of the Cux2 protein was convincingly detected in only one among 19 neuronal cell lines: the SH-SY5Y human neuroblastoma cell line. CDP/Cux proteins contain four DNA binding domains, three Cut repeat (CR1, CR2 and CR3) and one Cut homeodomain (HD). Purified fusion proteins containing either CR1CR2, CR2CR3HD or CR3HD exhibited similar DNA binding specificities as the corresponding domains of Cux1, but their DNA binding kinetics were much more rapid. Similarly, the full length Cux2 protein made rapid but transient interactions with DNA. We did not observe an N-terminally processed Cux2 isoform equivalent to the Cux1 p110 isoform. Whereas Cux1 can function as a repressor or activator in a promoter specific manner, Cux2 functioned exclusively as a transcriptional repressor in NIH3T3 cells. Overall, our results suggest that the Cux1 and Cux2 proteins carry distinct biochemical functions. Cux2 is able, like Cux1, to perform the CCAAT displacement activity. However, Cux2 is unlikely to execute transcriptional regulatory functions that require stable interaction with DNA. PMID- 15656992 TI - Molecular cloning and characterization of gene for Golgi-localized syntaphilin related protein on human chromosome 8q23. AB - Loci for several human genetic diseases including glaucoma have been mapped to q23 region on chromosome 8. We carried out homology search analysis of the genomic sequence of a bacterial artificial chromosome (BAC) clone, KB1590E11, on 8q23 region, and mapped a previously described cDNA, KIAA1472, to this BAC clone. In this study, we determined the complete genomic structure of the KIAA1472 gene and its expression in various tissues and cell lines. Four mRNA species (types 1a, 1b, 1c, and 2) were produced from this gene by use of alternative transcription start sites and alternative-splicing events. These mRNAs were expressed in various tissues, except for type 1a, which was found only in the brain. Further, type 1 mRNA could be translated into two protein isoforms with different N-terminal sequences; and type 2 mRNA, into another type of isoform. All three of these KIAA1472 gene products were localized in Golgi apparatus and contained a C-terminal hydrophobic segment characteristic of a transmembrane domain, thus indicating them to be Golgi membrane-bound proteins. Furthermore, these proteins were homologous to syntaphilin, a molecule involved in guiding vesicular transport. These results indicate that KIAA1472 gene products may play an important role in vesicular traffic in various tissues including the brain. PMID- 15656996 TI - Symptoms of post-traumatic stress disorder among emergency medicine residents. AB - There have been anecdotal reports of post-traumatic stress disorder (PTSD) in physicians responding to mass casualty events. No formal, prospective study has addressed the presence of PTSD symptoms as a result of the work of Emergency Medicine residents in non-mass casualty settings. The purpose of this study is to evaluate the presence of symptoms of PTSD among Emergency Medicine residents (EMR). The study was a survey of EMR, administered in an anonymous, voluntary format in late June 2001. The survey was conducted at an Emergency Medicine residency program that serves a large, urban, county hospital. Four groups, incoming interns and three EM resident classes were surveyed. PTSD symptoms were divided into three categories according to the DSM IV. The Jonckheere-Terpstra test for trends was applied to each of the three categories of symptoms. Sixty three surveys were administered, with a 93.6% response rate. All respondents reported experience with patient death or dying. Seven residents reported sufficient symptoms to meet the DSM IV criteria for PTSD. Each of the three symptom categories showed a statistically significant increase in the proportion of positive responses as the resident time in training increased (p < 0.01). In conclusion, many EM residents reported symptoms of PTSD. Symptoms of PTSD significantly increased as resident level of training increased. PMID- 15656994 TI - Functional characterization of the human SOX3 promoter: identification of transcription factors implicated in basal promoter activity. AB - SRY-related HMG-box genes (Sox genes) constitute a large family of developmentally regulated genes involved in the decision of cell fates during development and implicated in the control of diverse developmental processes. Sox3, an X-linked member of the family, is expressed in the central nervous system (CNS) from the earliest stages of development. It is considered to be one of the earliest neural markers in vertebrates playing the role in specifying neuronal fate. The aim of this study has been to determine and characterize the promoter of the human SOX3 gene and to elucidate molecular mechanisms underlying the regulation of its expression. In this study, we have isolated and performed the first characterization of the human SOX3 promoter. We have identified the transcription start point (tsp) and carried out the structural and functional analysis of the regulatory region responsible for SOX3 expression in NT2/D1 cell line. Using promoter-reporter constructs, we have determined the minimal SOX3 promoter region that confers the basal promoter activity, as well as two regulatory elements which have positive effects on the promoter activity. We have investigated in detail the functional properties of three conserved motifs within the core promoter sequence that bind transcription factors specificity protein 1 (Sp1), upstream stimulatory factor (USF) and nuclear factor Y (NF-Y). By mutational analysis, we have shown that all three sites are of functional relevance for constitutive SOX3 expression in NT2/D1 cells. We have also shown that, besides the TATA motif, at least one other essential regulatory element is required for the basal transcription of the human SOX3. Taken together, data presented in this paper suggest that transcription factors such as Sp1, USF and NF-Y could function as key regulators for the basal activation of the human SOX3 gene. PMID- 15656995 TI - Molecular markers for animal biotechnology: sea bass (Dicentrarchus labrax, L.) HMG-CoA reductase mRNA. AB - Modern technologies may improve fish production and quality and, at the same time, reduce environmental impact with benefits on the public perception of the industry. To be economically profitable, these modern technologies request an increase of rearing density that, however, could affect fish welfare. With the aim to search for molecular biomarkers to describe fish welfare, we have recently compared gene expression of sea bass farmed at different population densities by differential display obtaining six bands differentially expressed. In this paper, we have cloned the mRNA corresponding to one of those differentially expressed bands obtaining a 3860-bp sequence with an ORF of 2664 bp. Its virtual translation originated a 887-aa polypeptide that, by comparison with the other sequences available in the public data bases, resulted to be the 3-hydroxil-3 methyl-glutaryl coenzyme A reductase (HMGCR). In sea bass, as for the other species, the N- and C-terminus portions are the most conserved and are linked by an hydrophilic region that appears to be quite variable. Due to its role in the synthesis of cholesterol, HMGCR mRNA could be a good biomarker for detecting fish welfare. For this reason, we also followed, by real-time PCR, its expression after crowding stress comparing it with mRNA levels of HSP 70 and 90: HMGCR mRNA resulted highly expressed in the fishes farmed at 100 kg/m(3). PMID- 15656997 TI - The effect of vinegar on colorimetric end-tidal carbon dioxide determination after esophageal intubation. AB - This study examines the effect of vinegar placement in the stomach on colorimetric end-tidal carbon dioxide (ETCO(2)) determinations after esophageal intubation. Using a blinded, prospective, before and after post-mortem swine model, colorimetric ETCO(2) was determined after aspiration of the stomach contents and after placement of aliquots of saline and vinegar. Data were compiled from 12 swine within 120 min post-mortem. In 12 of 12 trials, the ETCO(2) detector turned yellow, indicating "positive" determination of CO(2), but did not return to "purple" with multiple insufflations with 100% O(2). We conclude that esophageal intubation with a small amount of vinegar in the stomach can cause an irreversible color change of the detector to yellow. Color change indicating the presence of carbon dioxide without subsequent color change back to purple with insufflation with 100% oxygen should arouse suspicion of improper placement of the endotracheal tube. PMID- 15656998 TI - On-scene physician assessment of thromboembolic etiology in out-of-hospital cardiac arrest. AB - Out-of-hospital cardiac arrest (OOHCA) treatment produces dismal recovery rates. Newer, directed therapies such as thrombolysis may be best considered if possible etiologies can be assessed immediately. We conducted a prospective, pilot, feasibility study of on-scene physician assessments in non-traumatic OOHCA. Physicians responded to the scene and reported likelihood of thromboembolic etiology based on detailed history and physical assessments. Included were 136/148 OOHCAs during the 6-month study period; median age was 69.5 years and 72% were men. Physicians judged 103/136 (76%) of arrests to be of thromboembolic etiology and would have recommended thrombolytic bolus in 83/136 (61%). Among 19 instances of physician-reported contraindications, 17 (90%) were not true contraindications. Median age was lower in the group recommended for thrombolysis. Thromboembolic etiology as judged by on-scene physicians was common and physicians recommended thrombolytic bolus commonly. Contraindications were highly overestimated. These data may be useful in the consideration of innovative, directed therapies such as thrombolysis in attempts to improve outcomes from OOHCA. PMID- 15656999 TI - Myocardial infarction and cardiac complications in emergency department patients admitted to the intensive care unit with gastrointestinal hemorrhage. AB - Patients admitted with significant gastrointestinal hemorrhage (GIH) often experience in-hospital cardiac complications. This retrospective study examined 68 patients admitted from the Emergency Department to the Intensive Care Unit (ICU) over a 1-year period. The patients were 75% Caucasian, 60% male, with a mean age of 57 +/- 19 years. Medical co-morbidity was noted in 70%, and 54% of patients had a history of significant alcohol use. A systolic blood pressure < 100 mm Hg was present in 26%, hemoglobin < 7 mg/dL in 32%, and three patients (4%) expired. Death, acute myocardial infarction or other cardiac complications were noted in 32% of patients. Patients older than 60 years were three times more likely to have a complicated course than were younger patients, and those with a co-morbidity were 14.8 times more likely. Patients with a history of significant alcohol use were 31% less likely to have an inpatient complication than those without such a history. Regression analysis supported the protective effect of a history of significant alcohol use and also demonstrated that a history of peptic ulcer disease was predictive of inpatient complications. Older GIH patients and those with co-morbidities may benefit from ICU disposition given their greater risk. Younger patients presenting with hematemesis and a history of significant alcohol use tended to have fewer complications such that it may be possible to manage these patients outside of the ICU if hemodynamically stable. PMID- 15657000 TI - Synchronized emergency department cardioversion of atrial dysrhythmias saves time, money and resources. AB - The strategy of elective synchronized cardioversion (EDCV) of new onset atrial fibrillation/flutter (AF/flutter) compares favorably to that of Emergency Department (ED) rate control and inpatient admission. This 1-year study comprised consecutive ED synchronized cardioversions performed on patients with new onset (< 48 h) AF/flutter; all were hemodynamically stable. A control group was obtained by chart review of all patients meeting the inclusion criteria admitted in the same year who were managed with rate control in the ED and inpatient admission. Thirty ED cardioversions were performed on 24 patients. Twenty-nine of 30 (97%) of ED cardioversions were successful. The mean hospital length of stay (LOS) for the EDCV group, including those admitted, was 22.8 h (95% CI: 1.7-44.0) compared to the control group: 55.6 h (all admitted) (95% CI: 41.6-69.6). Median LOS for the entire EDCV group was 4 h, compared with 39.3 h for the controls (p < 0.001). There was also a significant difference in median hospital charge, including ED care: EDCV group: $1598 vs. controls $4271 (p < 0.001). All of the study patients were contacted by telephone a minimum of 4 weeks after cardioversion to assess for complications, recidivism, and satisfaction. There were no complications in the EDCV group, and all expressed satisfaction with the procedure. Elective synchronized cardioversion in the ED is an effective strategy for management of new-onset AF/flutter and is associated with significant decreases in charges and length of stay as well as a high degree of patient satisfaction. PMID- 15657001 TI - Vocal cord dysfunction: an uncommon cause of stridor. AB - We present a case of vocal cord dysfunction syndrome (VCDS) presenting as acute angioedema of the upper airway. The presentation of this syndrome and its differentiation from other upper airway conditions that require far different and more urgent treatment is discussed. PMID- 15657002 TI - Evidence-based evaluation of psychiatric patients. AB - There has been much discussion in the literature concerning the medical evaluation of the psychiatric patient. The medical evaluation of the psychiatric patient involves obtaining a history, performing an appropriate physical and mental status examination, and testing, when indicated. This review applies the rigors of evidence-based evaluation of the medical literature that is currently available to determine when it is safe to place a patient with behavioral complaints under psychiatric care. PMID- 15657003 TI - Acute appendicitis of the appendiceal stump. AB - We report a case of a 27-year-old man, status post open appendectomy as an infant, in whom the diagnosis of acute appendicitis of the appendiceal stump was made by computed tomography (CT). A coronal reformatted CT image demonstrated both the inflamed appendix and a normal terminal ileum. Although rare, stump appendicitis may present with signs and symptoms typical of acute appendicitis in patients status post appendectomy and should be considered in the differential diagnosis. PMID- 15657005 TI - Altered mental status and ataxia secondary to acute Kava ingestion. AB - Kava has traditionally been used for a variety of purposes in the tropical islands of Polynesia but is becoming more frequently available in the United States health supplement market due to its calming effects in patients with anxiety. The side effect profile is poorly known but has recently gained the attention of the Food and Drug Administration (FDA). Although hepatitis and liver failure have been described with chronic ingestion, the effects of acute overdose are poorly described. We present a case of acute Kava overdose resulting in altered mental status and ataxia similar to that seen with ethanol intoxication. PMID- 15657004 TI - Pneumoperitoneum after percutaneous endoscopic gastrostomy: a case report and review. AB - A complication of percutaneous endoscopic gastrostomy (PEG) is perforation of a hollow viscus. This is typically detected by finding of pneumoperitoneum (PP) on radiographs. However, PP can occasionally be a benign finding. A review of the literature shows many causes for a benign PP, and it has been noted to occur frequently after PEG placement. In the absence of signs or symptoms of peritoneal inflammation, PP usually requires no further investigation or treatment. PMID- 15657007 TI - The epidemiology of skydiving injuries: World freefall convention, 2000-2001. AB - The purpose of this study is to describe the prevalence and types of injuries incurred by civilian skydivers using contemporary equipment under conventional conditions. Injury data were collected at the World Freefall skydiving convention (WFFC), during two consecutive periods of operation, August 4-13, 2000 and August 3-12, 2001. During the study periods, 8976 skydivers made 117,000 skydives. The First Aid Station at the WFFC treated 204 patients for injuries related to skydiving, at a rate of 17.4/10,000 (injuries/skydives). Most injuries were minor (66%) and required only simple first aid. Significant injuries, defined as those requiring treatment in the emergency department, occurred at a rate of 6.0/10,000 (injuries/skydives). The rate of hospitalization was 1.8/10,000 skydives. There was one fatality during this study. We believe these results provide a current update regarding the risk and types of injury related to recreational skydiving. PMID- 15657006 TI - Mycotoxins revisited: Part I. AB - Mushrooms are ubiquitous in nature. They are an important source of nutrition; however, certain varieties contain chemicals that can be highly toxic to humans. Industrially cultivated mushrooms are historically very safe, but foraging for mushrooms or accidental ingestion of mushrooms in the environment can result in serious illness and death. The emergency department is the most common site of presentation for patients suffering from acute mushroom poisoning. Although recognition can be facilitated by identification of a characteristic toxidrome, the presenting manifestations can be variable and have considerable overlap with more common and generally benign clinical syndromes. The goal of this two-part article is to review the knowledge base on this subject and provide information that will assist the clinician in the early consideration, diagnosis and treatment of mushroom poisoning. Part I, presented in this issue of the Journal, reviews the epidemiology and demographics of mushroom poisoning, the physical characteristics of the most toxic varieties, the classification of the toxic species, and an overview of the cyclopeptide-containing mushroom class. Part II, to be published in the next issue of the Journal, will be focused on the presentation of the other classes of toxic mushrooms along with an up-to-date review of the most recently identified poisonous varieties. PMID- 15657008 TI - Exertional chest pain. PMID- 15657010 TI - Ocular foreign body. PMID- 15657009 TI - Escalator injuries. PMID- 15657011 TI - Shortness of breath in an infant with history of acute lymphoblastic leukemia. PMID- 15657012 TI - Puzzling groin pain in a 15-year-old boy. PMID- 15657013 TI - A rare manifestation of ascariasis: encephalopathy. PMID- 15657014 TI - A child's hand caught in a bathtub plughole-anesthetic implications of an unusual clinical scenario. PMID- 15657015 TI - The UCSD Research Associate Program: a recipe for successfully integrating undergraduates with emergency medicine research. AB - Previous reports have documented the successful integration of undergraduates into the Emergency Department (ED) to assist in data collection for various research projects, with resultant improvements in departmental academic productivity. These reports have not detailed the exact procedures required to institute such a program. Over 100 undergraduates from three institutions currently participate in the UCSD Research Associate Program. Here we document our experience with the inception of such a program and define the various components required to successfully conduct an ED undergraduate research associate program, defining the roles of key personnel, detailing the administrative component, discussing the selection of appropriate research projects and the design of data and instruction sheets, addressing issues related to training and didactics, and suggesting strategies for recruitment and upkeep of interest. PMID- 15657016 TI - Trends in substance abuse by teenagers in the Czech Republic. AB - Drug Epidemiology Headquarters of the Hygienic Station of the Capital Prague has created a complex drug information system in the Czech Republic. This department collects and analyzes data concerning problem drug users, non-fatal emergencies, and occurence of infectious diseases, and all these surveys are provided on a routine basis. Data are gathered from the national network of treatment and contact centers for drug users, through repeated representative epidemiological surveys, obligatory reports on infectious diseases (hepatitis A, B and C, HIV/AIDS), and a sentinel system of data collection is used for non-fatal emergencies. In 1994, 1997, and 2000, a repeated study concerning trends of substance abuse by teenagers (aged 14 to 19 years of age) from all types of schools and from all regions of the Czech Republic was carried out by this department in order to study changes in substance abuse and teenagers' opinions related to drugs. In 1994 there were 8334 respondents in the study, in 1997 there were 14,282 respondents, and in 2000 there were 13,018 respondents, which is about 1.5% of the population group of teenagers from 14-19 years old in each year of the study. Recommendations of EMCDDA (European Monitoring Center for Drugs and Drug Addiction in Lisbon, Portugal), a body of European Council, are followed by Drug Epidemiology Headquarters. PMID- 15657017 TI - A medical student's perspective: selecting the best emergency medicine residency program. PMID- 15657018 TI - Functional limitations and survival following stroke: psychological and clinical predictors of 3-year outcome. AB - Previous studies predicting functional outcomes of stroke have mainly used clinical and demographic measures. This study examines the additional predictive value of psychological predictors. A cohort of patients were assessed on 6 occasions between admission to hospital and 3 years post-stroke, with 40 of the original sample of 101 being available at the final assessment. Demographic variables, clinical indexes, and psychological measures were used to predict survival to 3 years and 3-year functional recovery was assessed using measures derived from the Barthel Index which took account of initial levels of performance. For functional recovery, multiple regression analyses showed that fixed clinical and demographic indexes explained 16% to 40% of the variance in recovery. Perceptions of control 6 months after discharge added significantly to the predictive equations. Because these cognitions are modifiable, they offer opportunities for intervention to improve patient outcomes. Multiple regression showed that no psychological variables predicted survival. PMID- 15657019 TI - The role of purpose in life in recovery from knee surgery. AB - This study examined the role of a sense of purpose in life (PIL) in recovery from knee replacement surgery in 64 surgery patients. Each of the surgery patients had been diagnosed with severe osteoarthritis of the knee. Regression analyses were conducted predicting changes in health 6 months after surgery. When considered alone, PIL was related to less anxiety, depression, negative affect, functional disability, stiffness, and more positive affect. When optimism, pessimism, and emotionality were controlled, PIL was still related to less negative affect, depression, and anxiety, and more positive affect. The results suggest that PIL may be an important positive personal characteristic and target for interventions. PMID- 15657020 TI - Heartbeat sensitivity in adults with congenital heart disease. AB - This study tested the hypothesis that patients with a congenital heart disease are sensitive regarding heartbeat perception, reflected in enhanced attention for heartbeat, estimation of own heart rate, and a vulnerability to become anxious by listening to heartbeat sounds. Twenty adults with a congenital heart disease, and 20 healthy controls conducted 3 experimental tasks: a concentration task during distraction by heartbeat sounds, own heart rate estimation, and exposure to different patterns of heartbeat sounds. The results showed that patients were more distracted by heartbeat, and were also worse at estimating heart rate than controls. However, heartbeat sounds did not evoke anxiety. In conclusion, patients with a congenital heart disease may differ from controls in heartbeat perception, but there was no support for obsessive monitoring for heartbeat or excessive reactions to heartbeat sounds. PMID- 15657022 TI - Steps and sitting in a working population. AB - This study aimed to assess sitting time and number of steps taken each day, and the relationships between these variables, in a sample of working Australian adults. Workers (N = 185) wore a pedometer for 7 days and recorded the number of steps taken and time spent sitting each day. Average time spent sitting on weekdays was 9.4 (SD = 2.40) hr, with about half spent sitting at work. Despite this, the average steps taken each day (M = 8,873, SD = 2,757) was higher on weekdays than on weekend days. There was a clear inverse relationship between sitting time at work and number of steps taken on weekdays, r = -.34, p < .001); those in the highest tertile for sitting time reported about 3,000 fewer daily steps. Workers in managerial and professional occupations reported more time sitting at work (M = 6.2 hr per day) and lower weekday step counts (M = 7,883, N = 43) than technical (M = 3.3 hr sitting at work and 10,731 weekday steps, N = 33) and blue collar workers (M = 1.6 hours sitting and 11,784 steps, N = 11). The findings suggest those whose daily work involves long hours of sitting should be the focus of efforts to promote physical activity both within and outside the workplace. PMID- 15657021 TI - Hypoglycemia anticipation, awareness and treatment training (HAATT) reduces occurrence of severe hypoglycemia among adults with type 1 diabetes mellitus. AB - Severe hypoglycemia (SH) can be a significant problem for patients around the world with Type 1 Diabetes Mellitus (T1DM). To avoid SH, patients need to better manage, and reduce the occurrence of, preceding mild hypoglycemia. Hypoglycemia Anticipation, Awareness and Treatment Training (HAATT), developed in the United States specifically to address such issues, was evaluated at short- and long-term follow-up in a medically, economically and culturally different setting; Bulgaria. Sixty adults with T1DM and a history of recurrent SH (20 each from Sofia, Russe, and Varna, Bulgaria) were randomized to Self-Monitoring of Blood Glucose (SMBG) or SMBG+ HAATT. For 6 months before and 1 to 6 and 13 to 18 months after intervention, participants recorded occurrence of moderate, severe, and nocturnal hypoglycemia. For 1-month pre- and post-intervention, participants completed daily diaries concerning their diabetes management. Relative to SMBG, HAATT produced significant improvement in occurrence of low BG, moderate, severe, and nocturnal hypoglycemia, and detection and treatment of low BG (p values < .05 to < .001), with no compromise in metabolic control. At long-term follow-up, HAATT participants continued to have significantly fewer episodes of moderate and severe hypoglycemia. These findings suggest that a structured, specialized psycho educational treatment program (HAATT) can be highly effective in managing hypoglycemia. PMID- 15657024 TI - The value of sedentary alternatives influences child physical activity choice. AB - The choice to be physically active or sedentary depends in part on the value of the alternatives that are available. The shift from sedentary to active alternatives may be a function of the value of the sedentary alternatives. To evaluate the influence of the value of the sedentary alternatives on the choice to be physically active or sedentary, 30 nonobese 8-12-year-old youth were randomized in groups that provided a choice between 4 active alternatives or 4 sedentary alternatives (Group 1), their least valued sedentary activity (Group 2), or their most valued sedentary activity (Group 3), on computerized and questionnaire versions of a behavioral choice task. The computer task required participants to work for access to the alternatives. The work required to obtain access to the active alternatives remained constant for all choices, while the work required to gain access to the sedentary alternatives progressively increased. Compared to Groups 1 and 3, participants in Group 2 chose to be sedentary less often as they had to make a choice between being physically active or sedentary on the computerized (p < .005) and the questionnaire version (p < .05) of the behavioral choice task, which correlated r = .54, p < .01. These results suggest interventions designed to increase physical activity by reducing access to sedentary behaviors may need to consider the value of the targeted sedentary behavior and the extent to which the sedentary behaviors compete with physical activity. PMID- 15657023 TI - Are stress related factors associated with alcohol intake? AB - Moderate alcohol consumption is related to reduced risks of coronary heart disease and all-cause mortality. Our goal is to advance our understanding of the associations between stress-related factors and alcohol consumption, using cutoff points for alcohol intake that reflect health benefits rather than health risks. Cross-sectional data were used from 4,131 respondents (age 20-65 years) participating in a cohort study in the Netherlands on psychosocial factors and cancer risk. Analyses were performed among drinkers only, for men and women separately. Heavy alcohol intake (>/= 3 glasses per day for men, >/= 2 glasses per day for women) was associated with only a few stress-related factors in multivariate analyses. No significant associations between the total amount of stressors and alcohol intake were found. We conclude that stress-related factors are only marginally associated with a heavy alcohol intake compared with fair drinking, using the safe limits of drinking as cutoff point. PMID- 15657025 TI - Depression and its 6-month course in untreated hemodialysis patients: a preliminary prospective follow-up study in Turkey. AB - The major goal of this study is to assess the frequency of psychiatric disorders in end-stage renal disease (ESRD) patients, who were on hemodialysis (HD) treatment in Turkey. Additionally, it aims to determine whether depression, anxiety, and functional and occupational levels of patients who did not receive any psychiatric treatments change at the end of 6 months. We conducted Structured Clinical Interview for the DSM-IV, Clinical Version on 50 HD patients, and 16 (32%) fulfilled the criteria for a psychiatric disorder. Depressive disorder, not otherwise specified, was observed in 12 (24%) patients. Adjustment disorder with depressed mood and dysthymic disorder were observed in 8% of our population. Fourteen patients were followed up for 6 months. All patients were assessed with Hamilton Depression (HDRS), Anxiety Rating Scale (HARS), and Global Assessment of Functioning (GAF) at baseline and at 6 months. The comparisons of baseline and 6 months HDRS, HARS, and GAF scale scores did not reveal any statistically significant differences in 14 depressed patients. The level of depression, anxiety, and functional and occupational impairment did not remit spontaneously in our untreated depressed HD patients. We believe that further studies regarding categorization, prognosis, and treatment of these patients are needed to better understand what to do when we encounter depressive ESRD patients. PMID- 15657028 TI - Geotrichum silvicola sp. nov., a novel asexual arthroconidial yeast species related to the genus Galactomyces. AB - Four strains of an asexual arthroconidial yeast species were isolated from Drosophila flies in two Atlantic rain forest sites in Brazil and two strains from oak tasar silkworm larvae (Antheraea proylei) in India. Analysis of the sequences of the D1/D2 large subunit rRNA gene showed that this yeast represented a novel species of the genus Geotrichum, described as Geotrichum silvicola sp. nov. The novel species was related to the ascogenous genus Galactomyces. The closest relatives of Geotrichum silvicola were Galactomyces sp. strain NRRL Y-6418 and Galactomyces geotrichum. The type culture of Geotrichum silvicola is UFMG-354-2T (=CBS 9194T=NRRL Y-27641T). PMID- 15657030 TI - Heterodimerization of mu- and delta-opioid receptors occurs at the cell surface only and requires receptor-G protein interactions. AB - Homo- and heterodimerization of the opioid receptors with functional consequences were reported previously. However, the exact nature of these putative dimers has not been identified. In current studies, the nature of the heterodimers was investigated by producing the phenotypes of the 1:1 heterodimers formed between the constitutively expressed mu-opioid receptor (MOR) and the ponasterone A induced expression of delta-opioid receptor (DOR) in EcR293 cells. By examining the trafficking of the cell surface-located MOR and DOR, we determined that these two receptors endocytosed independently. Using cell surface expression-deficient mutants of MOR and DOR, we observed that the corresponding wild types of these receptors could not rescue the cell surface expression of the mutants, whereas the antagonist naloxone could. Furthermore, studies with constitutive or agonist induced receptor internalization also indicated that MOR and DOR endocytosed independently and could not "drag in" the corresponding wild types or endocytosis deficient mutants. Additionally, the heterodimer phenotypes could be eliminated by the pretreatment of the EcR293 cells with pertussis toxin and could be modulated by the deletion of the RRITR sequence in the third intracellular loop that is involved in the receptor-G protein interaction and activation. These data suggest that MOR and DOR heterodimerize only at the cell surface and that the oligomers of opioid receptors and heterotrimeric G protein are the bases for the observed MOR-DOR heterodimer phenotypes. PMID- 15657031 TI - Stabilization of the unliganded glucocorticoid receptor by TSG101. AB - The glucocorticoid receptor (GR) has been shown to undergo hormone-dependent down regulation via transcriptional, post-transcriptional, and posttranslational mechanisms. However, the mechanisms involved in modulating GR levels in the absence of hormone remain enigmatic. Here we demonstrate that TSG101, a previously identified GR-interacting protein, stabilizes the hypophosphorylated form of GR in the absence of ligand. We found that a non-phosphorylated version of GR (S203A/S211A) showed enhanced interaction with TSG101 as compared with the wild type GR, suggesting that TSG101 interacts more favorably with GR when it is not phosphorylated. A significant accumulation of GR S203A/S211A protein is detected in the absence of ligand when TSG101 is overexpressed, whereas no increase in the wild type phosphorylated GR or phosphomimetic GR S203E/S211E was observed in mammalian cells. In contrast, down-regulation of TSG101 expression by siRNA renders the hypophosphorylated form of GR unstable. We further show that TSG101 stabilizes GR by impeding its degradation by the proteasome and extending receptor half-life. Thus, in absence of a ligand, TSG101 binds GR and protects the non-phosphorylated receptor from degradation. PMID- 15657029 TI - A random peptide library fused to CCR5 for selection of mimetopes expressed on the mammalian cell surface via retroviral vectors. AB - A random peptide library was expressed on the surface of a mammalian cell by applying retroviral vectors. The random sequence was fused to the CCR5 chemokine receptor, which served as a scaffold to present the library at the cell surface. We used this library to isolate an epitope mimetope in a proof of principle system. This approach can become a tool for rapid creation of peptidic expression domains in a eukaryotic environment. Applications include the creation of decoys for receptors in cell-cell interactions, screening for molecules that drive ligand expression on target cells in two-cell interaction screens, among other utilities. PMID- 15657033 TI - A complex mechanism determines polarity of DNA replication fork arrest by the replication terminator complex of Bacillus subtilis. AB - Two dimers of the replication terminator protein (RTP) of Bacillus subtilis bind to a chromosomal DNA terminator site to effect polar replication fork arrest. Cooperative binding of the dimers to overlapping half-sites within the terminator is essential for arrest. It was suggested previously that polarity of fork arrest is the result of the RTP dimer at the blocking (proximal) side within the complex binding very tightly and the permissive-side RTP dimer binding relatively weakly. In order to investigate this "differential binding affinity" model, we have constructed a series of mutant terminators that contain half-sites of widely different RTP binding affinities in various combinations. Although there appeared to be a correlation between binding affinity at the proximal half-site and fork arrest efficiency in vivo for some terminators, several deviated significantly from this correlation. Some terminators exhibited greatly reduced binding cooperativity (and therefore have reduced affinity at each half-site) but were highly efficient in fork arrest, whereas one terminator had normal affinity over the proximal half-site, yet had low fork arrest efficiency. The results show clearly that there is no direct correlation between the RTP binding affinity (either within the full complex or at the proximal half-site within the full complex) and the efficiency of replication fork arrest in vivo. Thus, the differential binding affinity over the proximal and distal half-sites cannot be solely responsible for functional polarity of fork arrest. Furthermore, efficient fork arrest relies on features in addition to the tight binding of RTP to terminator DNA. PMID- 15657032 TI - Modulation of endosomal cholesteryl ester metabolism by membrane cholesterol. AB - Cells acquire cholesterol in part by endocytosis of cholesteryl ester containing lipoproteins. In endosomes and lysosomes cholesteryl ester is hydrolyzed by acidic cholesteryl ester hydrolase producing cholesterol and fatty acids. Under certain pathological conditions, however, such as in atherosclerosis, excessive levels of cholesteryl ester accumulate in lysosomes for reasons that are poorly understood. Here, we have studied endosomal and lysosomal cholesteryl ester metabolism in cultured mouse macrophages and with cell-free extracts. We show that net hydrolysis of cholesteryl ester is coupled to the transfer of cholesterol to membranes. When membrane cholesterol levels are low, absorption of cholesterol effectively drives cholesteryl ester hydrolysis. When cholesterol levels in acceptor membranes approach saturation or when cholesterol export is blocked, cholesterol is re-esterified in endosomes. These results reveal a new facet of cellular cholesterol homeostasis and provide a potential explanation for cholesteryl ester accumulation in lysosomes of atherosclerotic cells. PMID- 15657034 TI - Intracellular positioning of isoforms explains an unusually large adenylate kinase gene family in the parasite Trypanosoma brucei. AB - Adenylate kinases occur classically as cytoplasmic and mitochondrial enzymes, but the expression of seven adenylate kinases in the flagellated protozoan parasite Trypanosoma brucei (order, Kinetoplastida; family, Trypanosomatidae) easily exceeds the number of isoforms previously observed within a single cell and raises questions as to their location and function. We show that a requirement to target adenylate kinase into glycosomes, which are unique kinetoplastid-specific microbodies of the peroxisome class in which many reactions of carbohydrate metabolism are compartmentalized, and two different flagellar structures as well as cytoplasm and mitochondrion explains the expansion of this gene family in trypanosomes. The three isoforms that are selectively built into either the flagellar axoneme or the extra-axonemal paraflagellar rod, which is essential for motility, all contain long N-terminal extensions. Biochemical analysis of the only short form trypanosome adenylate kinase revealed that this enzyme catalyzes phosphotransfer of gamma-phosphate from ATP to AMP, CMP, and UMP acceptors; its high activity and specificity toward CMP is likely to reflect an adaptation to very low intracellular cytidine nucleotide pools. Analysis of some of the phosphotransfer network using RNA interference suggests considerable complexity within the homeostasis of cellular energetics. The anchoring of specific adenylate kinases within two distinct flagellar structures provides a paradigm for metabolic organization and efficiency in other flagellates. PMID- 15657035 TI - Search for apoptotic nucleases in yeast: role of Tat-D nuclease in apoptotic DNA degradation. AB - DNA fragmentation/degradation is an important step for apoptosis. However, in unicellular organisms such as yeast, this process has rarely been investigated. In the current study, we revealed eight apoptotic nuclease candidates in Saccharyomyces cerevisiae, analogous to the Caenorhabditis elegans apoptotic nucleases. One of them is Tat-D. Sequence comparison indicates that Tat-D is conserved across kingdoms, implicating that it is evolutionarily and functionally indispensable. In order to better understand the biochemical and biological functions of Tat-D, we have overexpressed, purified, and characterized the S. cerevisiae Tat-D (scTat-D). Our biochemical assays revealed that scTat-D is an endo-/exonuclease. It incises the double-stranded DNA without obvious specificity via its endonuclease activity and excises the DNA from the 3'- to 5'-end by its exonuclease activity. The enzyme activities are metal-dependent with Mg(2+) as an optimal metal ion and an optimal pH around 5. We have also identified three amino acid residues, His(185), Asp(325), and Glu(327), important for its catalysis. In addition, our study demonstrated that knock-out of TAT-D in S. cerevisiae increases the TUNEL-positive cells and cell survival in response to hydrogen hyperoxide treatment, whereas overexpression of Tat-D facilitates cell death. These results suggest a role of Tat-D in yeast apoptosis. PMID- 15657036 TI - Molecular characterization of the human Calpha-formylglycine-generating enzyme. AB - Calpha-formylglycine (FGly) is the catalytic residue in the active site of sulfatases. In eukaryotes, it is generated in the endoplasmic reticulum by post translational modification of a conserved cysteine residue. The FGly-generating enzyme (FGE), performing this modification, is an endoplasmic reticulum-resident enzyme that upon overexpression is secreted. Recombinant FGE was purified from cells and secretions to homogeneity. Intracellular FGE contains a high mannose type N-glycan, which is processed to the complex type in secreted FGE. Secreted FGE shows partial N-terminal trimming up to residue 73 without loosing catalytic activity. FGE is a calcium-binding protein containing an N-terminal (residues 86 168) and a C-terminal (residues 178-374) protease-resistant domain. The latter is stabilized by three disulfide bridges arranged in a clamp-like manner, which links the third to the eighth, the fourth to the seventh, and the fifth to the sixth cysteine residue. The innermost cysteine pair is partially reduced. The first two cysteine residues are located in the sequence preceding the N-terminal protease-resistant domain. They can form intramolecular or intermolecular disulfide bonds, the latter stabilizing homodimers. The C-terminal domain comprises the substrate binding site, as evidenced by yeast two-hybrid interaction assays and photocross-linking of a substrate peptide to proline 182. Peptides derived from all known human sulfatases served as substrates for purified FGE indicating that FGE is sufficient to modify all sulfatases of the same species. PMID- 15657037 TI - Type I transforming growth factor beta receptor binds to and activates phosphatidylinositol 3-kinase. AB - We have examined the interaction of transforming growth factor (TGF)beta receptors with phosphatidylinositol 3-(PI3) kinase in epithelial cells. In COS7 cells, treatment with TGFbeta increased PI3 kinase activity as measured by the ability of p85-associated immune complexes to phosphorylate inositides in vitro. Both type I and type II TGFbeta receptors (TbetaR) associated with p85, but the association of TbetaRII appeared to be constitutive. The interaction of TbetaRI with p85 was induced by treatment with TGFbeta. The receptor association with PI3 kinase was not direct as (35)S-labeled rabbit reticulocyte p85 did not couple with fusion proteins containing type I and type II receptors. A kinase-dead, dominant-negative mutant of TbetaRII blocked ligand-induced p85-TbetaRI association and PI3 kinase activity. In TbetaRI-null R1B cells, TGFbeta did not stimulate PI3 kinase activity. This stimulation was restored upon reconstitution of TbetaRI by transfection. In R1B and NMuMG epithelial cells, overexpression of a dominant active mutant form of TbetaRI markedly enhanced ligand-independent PI3 kinase activity, which was blocked by the addition of the TbetaRI kinase inhibitor LY580276, suggesting a causal link between TbetaRI function and PI3 kinase. Overexpressed Smad7 also prevented ligand-induced PI3 kinase activity. Taken together, these data suggest that 1) TGFbeta receptors can indirectly associate with p85, 2) both receptors are required for ligand-induced PI3 kinase activation, and 3) the activated TbetaRI serine-threonine kinase can potently induce PI3 kinase activity. PMID- 15657038 TI - Folding competence of N-terminally truncated forms of human procathepsin B. AB - Besides acting as an inhibitor, the propeptide of human cathepsin B exerts an important auxiliary function as a chaperone in promoting correct protein folding. To explore the ability of N-terminally truncated forms of procathepsin B to fold into enzymatically active proteins, we produced procathepsin B variants progressively lacking N-terminal structural elements in baculovirus-infected insect cells. N-terminal truncation of the propeptide by up to 22 amino acids did not impair the production of activable procathepsin B. Secreted forms lacking the first 20, 21, or 22 amino acids spontaneously generated mature cathepsin B through autocatalytic processing, demonstrating that the first alpha-helix (Asp11 Arg20) is necessary for efficient inhibition of the enzyme by its propeptide. In contrast, proenzymes lacking the N-terminal part including the first beta-sheet (Trp24-Ala26) of the propeptide or containing an amino acid mutation directly preceding this beta-sheet were no longer properly folded. This shows that interactions between Trp24 of the propeptide and Tyr183, Tyr188, and Phe180 of the mature enzyme are important for stabilization and essential for procathepsin B folding. Thus, proenzyme forms missing more than the N-terminal 22 amino acids of the propeptide (notably truncated cathepsin B produced by the mRNA splice variant lacking exons 2 and 3, resulting in a propeptide shortened by 34 amino acids) are devoid of proteolytic activity because they cannot fold correctly. Thus, any pathophysiological involvement of truncated cathepsin B must be ascribed to properties other than proteolysis. PMID- 15657039 TI - Homeobox-clock protein interaction in zebrafish. A shared mechanism for pineal specific and circadian gene expression. AB - In non-mammalian vertebrates, the pineal gland is photoreceptive and contains an intrinsic circadian oscillator that drives rhythmic production and secretion of melatonin. These features require an accurate spatiotemporal expression of an array of specific genes in the pineal gland. Among these is the arylalkylamine N acetyltransferase, a key enzyme in the melatonin production pathway. In zebrafish, pineal specificity of zfaanat2 is determined by a region designated the pineal-restrictive downstream module (PRDM), which contains three photoreceptor conserved elements (PCEs) and an E-box, elements that are generally associated with photoreceptor-specific and rhythmic expression, respectively. Here, by using in vivo and in vitro approaches, it was found that the PCEs and E box of the PRDM mediate a synergistic effect of the photoreceptor-specific homeobox OTX5 and rhythmically expressed clock protein heterodimer, BMAL/CLOCK, on zfaanat2 expression. Furthermore, the distance between the PCEs and the E-box was found to be critical for PRDM function, suggesting a possible physical feature of this synergistic interaction. OTX5-BMAL/CLOCK may act through this mechanism to simultaneously control pineal-specific and rhythmic expression of zfaanat2 and possibly also other pineal and retinal genes. PMID- 15657040 TI - Effects of p47phox C terminus phosphorylations on binding interactions with p40phox and p67phox. Structural and functional comparison of p40phox and p67phox SH3 domains. AB - The neutrophil NADPH oxidase produces superoxide anions in response to infection. This reaction is activated by association of cytosolic factors, p47phox and p67phox, and a small G protein Rac with the membranous flavocytochrome b558. Another cytosolic factor, p40phox, is associated to the complex and is reported to play regulatory roles. Initiation of the NADPH oxidase activation cascade has been reported as consecutive to phosphorylation on serines 359/370 and 379 of the p47phox C terminus. These serines surround a polyproline motif that can interact with the Src homology 3 (SH3) module of p40phox (SH3p40) or the C-terminal SH3 of p67phox (C-SH3p67). The latter one presents a higher affinity in the resting state for p47phox. A change in SH3 binding preference following phosphorylation has been postulated earlier. Here we report the crystal structures of SH3p40 alone or in complex with a 12-residue proline-rich region of p47phox at 1.46 angstrom resolution. Using intrinsic tryptophan fluorescence measurements, we compared the affinity of the strict polyproline motif and the whole C terminus peptide with both SH3p40 and C-SH3p67. These data reveal that SH3p40 can interact with a consensus polyproline motif but also with a noncanonical motif of the p47phox C terminus. The electrostatic surfaces of both SH3 are very different, and therefore the binding preference for C-SH3p67 can be attributed to the polyproline motif recognition and particularly to the Arg-368p47 binding mode. The noncanonical motif contributes equally to interaction with both SH3. The influence of serine phosphorylation on residues 359/370 and 379 on the affinity for both SH3 domains has been checked. We conclude that contrarily to previous suggestions, phosphorylation of Ser-359/370 does not modify the SH3 binding affinity for both SH3, whereas phosphorylation of Ser-379 has a destabilizing effect on both interactions. Other mechanisms than a phosphorylation induced switch between the two SH3 must therefore take place for NADPH oxidase activation cascade to start. PMID- 15657042 TI - Atomic force microscopy imaging demonstrates that P2X2 receptors are trimers but that P2X6 receptor subunits do not oligomerize. AB - P2X receptors are cation-selective channels activated by extracellular ATP. The architecture of these receptors is still not completely clear. Here we have addressed this issue by both chemical cross-linking and direct imaging of individual receptors by atomic force microscopy (AFM). Cross-linking of the P2X(2) receptor produced higher order adducts, consistent with the presence of trimers. The mean molecular volume of the receptor determined by AFM (409 nm(3)) also points to a trimeric structure. P2X(2) receptors bearing His(6) epitope tags were incubated with anti-His(6) antibodies, and the resultant complexes were imaged by AFM. For receptors with two bound antibodies, the mean angle between the antibodies was 123 degrees , again indicating that the receptor is a trimer. In contrast, cross-linking of the P2X(6) receptor did not produce higher order adducts, and the mean molecular volume of the receptor was 145 nm(3). We conclude that P2X(2) receptors are trimers, whereas the P2X(6) receptor subunits do not form stable oligomers. PMID- 15657041 TI - Steric accessibility of the HIV-1 gp41 N-trimer region. AB - During human immunodeficiency virus entry, gp41 undergoes a series of conformational changes that induce membrane fusion. Immediately prior to fusion, gp41 exists in a prehairpin intermediate in which the N- and C-peptide regions of gp41 are exposed. Rearrangement of this intermediate into a six-helix bundle composed of a trimeric coiled coil from the N-peptide region (N-trimer) surrounded by three peptides from the C-peptide region provides the driving force for membrane fusion, whereas prevention of six-helix bundle formation inhibits viral entry. Because of its central role in mediating viral entry, the N-trimer region of gp41 is a key vaccine target. Extensive efforts to discover potent and broadly neutralizing antibodies (Abs) against the N-trimer region have, thus far, been unsuccessful. In this study, we attached a potent C-peptide inhibitor that binds to the N-trimer region to cargo proteins of various sizes to examine the steric accessibility of the N-trimer during fusion. These inhibitors show a progressive loss of potency with increasing cargo size. Extension of the cargo/C peptide linker partially restores inhibitory potency. These results demonstrate that the human immunodeficiency virus defends its critical hairpin-forming machinery by steric exclusion of large proteins and may explain the current dearth of neutralizing Abs against the N-trimer. In contrast, previous results suggest the C-peptide region is freely accessible during fusion, demonstrating that the N- and C-peptide regions are in structurally distinct environments. Based on these results, we also propose new strategies for the generation of neutralizing Abs that overcome this steric block. PMID- 15657043 TI - Mutational analysis of target enzyme recognition of the beta-trefoil fold barley alpha-amylase/subtilisin inhibitor. AB - The barley alpha-amylase/subtilisin inhibitor (BASI) inhibits alpha-amylase 2 (AMY2) with subnanomolar affinity. The contribution of selected side chains of BASI to this high affinity is discerned in this study, and binding to other targets is investigated. Seven BASI residues along the AMY2-BASI interface and four residues in the putative protease-binding loop on the opposite side of the inhibitor were mutated. A total of 15 variants were compared with the wild type by monitoring the alpha-amylase and protease inhibitory activities using Blue Starch and azoalbumin, respectively, and the kinetics of binding to target enzymes by surface plasmon resonance. Generally, the mutations had little effect on k(on), whereas the k(off) values were increased up to 67-fold. The effects on the inhibitory activity, however, were far more pronounced, and the K(i) values of some mutants on the AMY2-binding side increased 2-3 orders of magnitude, whereas mutations on the other side of the inhibitor had virtually no effect. The mutants K140L, D150N, and E168T lost inhibitory activity, revealing the pivotal role of charge interactions for BASI activity on AMY2. A fully hydrated Ca(2+) at the AMY2-BASI interface mediates contacts to the catalytic residues of AMY2. Mutations involving residues contacting the solvent ligands of this Ca(2+) had weaker affinity for AMY2 and reduced sensitivity to the Ca(2+) modulation of the affinity. These results suggest that the Ca(2+) and its solvation sphere are integral components of the AMY2-BASI complex, thus illuminating a novel mode of inhibition and a novel role for calcium in relation to glycoside hydrolases. PMID- 15657044 TI - Acceptor RNA cleavage profile supports an invasion mechanism for HIV-1 minus strand transfer. AB - We previously proposed that HIV-1 minus strand transfer occurs by an acceptor invasion-initiated multi-step mechanism. During synthesis of minus strong stop DNA, reverse transcriptase (RT) transiently pauses at the base of TAR before continuing synthesis. Pausing promotes RT-RNase H cleavage of the donor RNA, exposing regions of the cDNA. The acceptor RNA then invades at these locations to interact with the minus strong stop DNA. Whereas primer extension continues on the donor RNA, the cDNA-acceptor hybrid expands by branch migration until transfer of the primer terminus is completed. We present results here showing that the interaction of the acceptor RNA and the cDNA can be determined by examining the time-dependent cleavage of the acceptor RNA by RNase H. Our approach utilizes a combination of RT-RNase H and Escherichia coli RNase H to allow assessment of acceptor-cDNA interactions at high sensitivity. Results show an initial interaction of the acceptor RNA with cDNA at the base of TAR. We observe a time-dependent shift in RNase H susceptibility along the length of the acceptor toward the 5' end, suggesting hybrid propagation from the initial invasion point. Control experiments validate that the RNase H cleavage profile represents the formation and expansion of the acceptor-DNA interaction and that the process is promoted by the nucleocapsid. Observations with this new approach lend additional support to the proposed multistep transfer mechanism. PMID- 15657045 TI - Lipid rafts control signaling of type-1 cannabinoid receptors in neuronal cells. Implications for anandamide-induced apoptosis. AB - Several G protein-coupled receptors function within lipid rafts plasma membrane microdomains, which may be important in limiting signal transduction. Here we show that treatment of rat C6 glioma cells with the raft disruptor methyl-beta cyclodextrin (MCD) doubles the binding efficiency (i.e. the ratio between maximum binding and dissociation constant) of type-1 cannabinoid receptors (CB1R), which belong to the rhodopsin family of G protein-coupled receptors. In parallel, activation of CB1R by the endogenous agonist anandamide (AEA) leads to approximately 3-fold higher [35S]GTPgammaS binding in MCD-treated cells than in controls, and CB1R-dependent signaling via adenylate cyclase, and p42/p44 MAPK is almost doubled by MCD. Unlike CB1R, the other AEA-binding receptor TRPV1, the AEA synthetase NAPE-PLD, and the AEA hydrolase FAAH are not modulated by MCD, whereas the activity of the AEA membrane transporter (AMT) is reduced to approximately 50% of the controls. We also show that MCD reduces dose-dependently AEA-induced apoptosis in C6 cells but not in human CHP100 neuroblastoma cells, which mirror the endocannabinoid system of C6 cells but are devoid of CB1R. MCD reduces also cytochrome c release from mitochondria of C6 cells, and this effect is CB1R dependent and partly mediated by activation of p42/p44 MAPK. Altogether, the present data suggest that lipid rafts control CB1R binding and signaling, and that CB1R activation underlies the protective effect of MCD against apoptosis. PMID- 15657046 TI - Cannabinoid receptor-induced neurite outgrowth is mediated by Rap1 activation through G(alpha)o/i-triggered proteasomal degradation of Rap1GAPII. AB - The G(alpha)o/i-coupled CB1 cannabionoid receptor induces neurite outgrowth in Neuro-2A cells. The mechanisms of signaling through G(alpha)o/i to induce neurite outgrowth were studied. The expression of G(alpha)o/i reduces the stability of its direct interactor protein, Rap1GAPII, by targeting it for ubiquitination and proteasomal degradation. This results in the activation of Rap1. G(alpha)o/i induced activation of endogenous Rap1 in Neuro-2A cells is blocked by the proteasomal inhibitor lactacystin. G(alpha)o/i stimulates neurite outgrowth that is blocked by the expression of dominant negative Rap1. Expression of Rap1GAPII also blocks the G(alpha)o/i-induced neurite outgrowth and treatment with proteasomal inhibitors potentiates this inhibition. The endogenous G(alpha)o/i coupled cannabinoid (CB1) receptor in Neuro-2A cells stimulates the degradation of Rap1GAPII; activation of Rap1 and treatment with pertussis toxin or lactacystin blocks these effects. The CB1 receptor-stimulated neurite outgrowth is blocked by treatment with pertussis toxin, small interfering RNA for Rap, lactacystin, and expression of Rap1GAPII. Thus, the G(alpha)o/i-coupled cannabinoid receptor, by regulating the proteasomal degradation of Rap1GAPII, activates Rap1 to induce neurite outgrowth. PMID- 15657047 TI - A mathematical model for the branched chain amino acid biosynthetic pathways of Escherichia coli K12. AB - As a first step toward the elucidation of the systems biology of the model organism Escherichia coli, it was our goal to mathematically model a metabolic system of intermediate complexity, namely the well studied end product-regulated pathways for the biosynthesis of the branched chain amino acids L-isoleucine, L valine, and L-leucine. This has been accomplished with the use of kMech (Yang, C. R., Shapiro, B. E., Mjolsness, E. D., and Hatfield, G. W. (2005) Bioinformatics 21, in press), a Cellerator (Shapiro, B. E., Levchenko, A., Meyerowitz, E. M., Wold, B. J., and Mjolsness, E. D. (2003) Bioinformatics 19, 677-678) language extension that describes a suite of enzyme reaction mechanisms. Each enzyme mechanism is parsed by kMech into a set of fundamental association-dissociation reactions that are translated by Cellerator into ordinary differential equations. These ordinary differential equations are numerically solved by Mathematica. Any metabolic pathway can be simulated by stringing together appropriate kMech models and providing the physical and kinetic parameters for each enzyme in the pathway. Writing differential equations is not required. The mathematical model of branched chain amino acid biosynthesis in E. coli K12 presented here incorporates all of the forward and reverse enzyme reactions and regulatory circuits of the branched chain amino acid biosynthetic pathways, including single and multiple substrate (Ping Pong and Bi Bi) enzyme kinetic reactions, feedback inhibition (allosteric, competitive, and non-competitive) mechanisms, the channeling of metabolic flow through isozymes, the channeling of metabolic flow via transamination reactions, and active transport mechanisms. This model simulates the results of experimental measurements. PMID- 15657048 TI - Structural requirements of the extracellular to transmembrane domain junction for erythropoietin receptor function. AB - The erythropoietin receptor (EpoR) is crucial for erythrocyte formation. The x ray crystal structures of the EpoR extracellular domain lack the juxtamembrane (JM) region and the junction to the transmembrane (TM) domain. Yet the JM-TM regions are important for transmitting the conformational change imposed on the receptor dimer by Epo binding. Cysteine-scanning mutagenesis of the JM-TM regions identified three novel constitutively active mutants, demonstrating close disulfide-bonded juxtapositioning of these residues in the JM (L223C) and N terminal TM domain (L226C, I227C). Chemical cross-linking defined the interface of the active helical TM dimer and revealed that the JM-TM segment encompassing Leu(226)-Leu(230) is non-helical. Molecular dynamics and NMR studies indicated that the TM-JM junction forms an N-terminal helix cap. This structure is important for EpoR function because replacement of this motif by consecutive leucines rendered the receptor constitutively active. PMID- 15657049 TI - Cdc24 regulates nuclear shuttling and recruitment of the Ste5 scaffold to a heterotrimeric G protein in Saccharomyces cerevisiae. AB - The Saccharomyces cerevisiae guanine nucleotide exchange factor Cdc24 regulates polarized growth by binding to Cdc42, a Rho-type GTPase that has many effectors, including Ste20 kinase, which activates multiple MAPK cascades. Here, we show that Cdc24 promotes MAPK signaling during mating through interactions with Ste5, a scaffold that must shuttle through the nucleus and bind to the beta subunit (Ste4) of a G protein for Ste20 to activate the tethered MAPK cascade. Ste5 was basally recruited to growth sites of G1 phase cells independently of Ste4. Loss of Cdc24 inhibited nuclear import and blocked basal and pheromone-induced recruitment of Ste5. Ste5 was not basally recruited and the MAPK Fus3 was not basally activated in the presence of a Cdc24 mutant (G168D) that still activates Cdc42, suggesting that Cdc24 regulates Ste5 and the associated MAPK cascade through a function that is not dependent on its guanine nucleotide exchange factor activity. Consistent with this, Cdc24 bound Ste5 and coprecipitated with Ste4 independently of Far1 and Ste5. Loss of Cdc24 decreased Ste5-Ste4 complex formation, and loss of Ste4 stimulated Cdc24-Ste5 complex formation. Collectively, these findings suggest that Cdc24 mediates site-specific localization of Ste5 to a heterotrimeric G protein and may therefore ensure localized activation of the associated MAPK cascade. PMID- 15657050 TI - The pro-inflammatory mediator leukotriene D4 induces phosphatidylinositol 3 kinase and Rac-dependent migration of intestinal epithelial cells. AB - Inflammatory bowel diseases are associated with increased risk of developing colon cancer. A possible role of the pro-inflammatory leukotriene D4 (LTD4) in this process has been implicated by the findings that LTD4 can signal increased proliferation and survival, both hallmarks of a cancer cell, in non-transformed intestinal epithelial cells. Here we make the novel finding that LTD4 can also signal increased motility in these cells. In parallel, we found that LTD4 induced a simultaneous transient 10-fold increase in Rac but not Cdc42 activity. These data were also supported by the ability of LTD4 to activate the Rac GDP/GTP exchange factor Vav2. Further, LTD4 triggered a 3-fold transient increase in phosphatidylinositol 3-kinase (PI3K) phosphorylation, a possible upstream activator of the Vav2/Rac signaling pathway. The activation of Rac was blocked by the PI3K inhibitors LY294002 and wortmannin and by transfection of a kinase negative mutant of PI3K or a dominant-negative form of Vav2. Furthermore, Rac was found to co-localize with actin in LTD4-generated membrane ruffles that were formed by a PI3K-dependent mechanism. In accordance, the inhibition of the PI3K and Rac signaling pathway also blocked the LTD4-induced migration of the intestinal cells. The present data reveal that an inflammatory mediator such as LTD4 cannot only increase proliferation and survival of non-transformed intestinal epithelial cells but also, via a PI3K/Rac signaling pathway, trigger a motile response in such cells. These data demonstrate the capacity of inflammatory mediators to participate in the process by which inflammatory bowel conditions increase the risk for colon cancer development. PMID- 15657051 TI - Amplification of telomeric arrays via rolling-circle mechanism. AB - Alternative (telomerase-independent) lengthening of telomeres mediated through homologous recombination is often accompanied by a generation of extrachromosomal telomeric circles (t-circles), whose role in direct promotion of recombinational telomere elongation has been recently demonstrated. Here we present evidence that t-circles in a natural telomerase-deficient system of mitochondria of the yeast Candida parapsilosis replicate independently of the linear chromosome via a rolling-circle mechanism. This is supported by an observation of (i) single stranded DNA consisting of concatameric arrays of telomeric sequence, (ii) lasso shaped molecules representing rolling-circle intermediates, and (iii) preferential incorporation of deoxyribonucleotides into telomeric fragments and t circles. Analysis of naturally occurring variant t-circles revealed conserved motifs with potential function in driving the rolling-circle replication. These data indicate that extrachromosomal t-circles observed in a wide variety of organisms, including yeasts, plants, Xenopus laevis, and certain human cell lines, may represent independent replicons generating telomeric sequences and, thus, actively participating in telomere dynamics. Moreover, because of the promiscuous occurrence of t-circles across phyla, the results from yeast mitochondria have implications related to the primordial system of telomere maintenance, providing a paradigm for evolution of telomeres in nuclei of early eukaryotes. PMID- 15657052 TI - Mouse testican-2. Expression, glycosylation, and effects on neurite outgrowth. AB - Mouse testican-2 was cloned, sequenced, and shown to be a proteoglycan with a multidomain structure closely similar to that of the human ortholog, previously described as a calcium binding extracellular matrix molecule of the BM 40/SPARC/osteonectin family (Vannahme, C., Schubel, S., Herud, M., Gosling, S., Hulsmann, H., Paulsson, M., Hartmann, U., and Maurer, P. (1999). J. Neurochem. 73, 12-20). Recombinant mouse testican-2 was used to prepare specific antibodies that allowed the detection of testican-2 in various brain structures but also in lung, testis, and in several endocrine glands. Although the testican-2 expressed in EBNA-293 cells carried both heparan sulfate and chondroitin/dermatan sulfate glycosaminoglycan chains, the tissue form always contained only heparan sulfate. Both tissue-derived and recombinant testican-2 carried N-linked glycans. Tissue derived forms of testican-2 were detected as proteoglycans of varying size, whereas a portion of the molecules produced by EBNA-293 cells were core proteins, lacking glycosaminoglycans. Both the proteoglycan and core protein forms of testican-2 inhibited neurite extension from cultured primary cerebellar neurons and may play regulatory roles in the development of the central nervous system. PMID- 15657053 TI - Physicochemical characterization of casein phosphopeptide-amorphous calcium phosphate nanocomplexes. AB - Milk caseins stabilize calcium and phosphate ions and make them available to the neonate. Tryptic digestion of the caseins yields phosphopeptides from their polar N-terminal regions that contain clusters of phosphorylated seryl residues. These phosphoseryl clusters have been hypothesized to be responsible for the interaction between the caseins and calcium phosphate that lead to the formation of casein micelles. The casein phosphopeptides stabilize calcium and phosphate ions through the formation of complexes. The calcium phosphate in these complexes is biologically available for intestinal absorption and remineralization of subsurface lesions in tooth enamel. We have studied the structure of the complexes formed by the casein phosphopeptides with calcium phosphate using a range of physicochemical techniques including x-ray powder diffraction, scanning electron microscopy, transmission electron microscopy, and equilibrium binding analyses. The amorphous nature of the calcium phosphate phase was confirmed by two independent methods: x-ray powder diffraction and selected area diffraction. In solution, the ion activity product of a basic amorphous calcium phosphate phase was the only ion product that was a function of bound phosphate independent of pH, consistent with basic amorphous calcium phosphate being the phase stabilized by the casein phosphopeptides. Detailed investigations of calcium and calcium phosphate binding using a library of synthetic homologues and analogues of the casein phosphopeptides have revealed that although the fully phosphorylated seryl-cluster motif is pivotal for the interaction with calcium and phosphate, other factors are also important. In particular, calcium binding and calcium phosphate stabilization by the peptides was influenced by peptide net charge, length, and sequence. PMID- 15657055 TI - A five-coordinate metal center in Co(II)-substituted VanX. AB - In an effort to structurally probe the metal binding site in VanX, electronic absorption, EPR, and extended x-ray absorption fine structure (EXAFS) spectroscopic studies were conducted on Co(II)-substituted VanX. Electronic spectroscopy revealed the presence of Co(II) ligand field transitions that had molar absorptivities of approximately 100 m(-1) cm(-1), which suggests that Co(II) is five-coordinate in Co(II)-substituted VanX. Low temperature EPR spectra of Co(II)-substituted VanX were simulated using spin Hamiltonian parameters of M(S) = |+/-1/2), E/D = 0.14, g(real(x,y)) = 2.37, and g(real(z)) = 2.03. These parameters lead to the prediction that Co(II) in the enzyme is five-coordinate and that there may be at least one solvent-derived ligand. Single scattering fits of EXAFS data indicate that the metal ions in both native Zn(II)-containing and Co(II)-substituted VanX have the same coordination number and that the metal ions are coordinated by 5 nitrogen/oxygen ligands at approximately 2.0 angstroms. These data demonstrate that Co(II) (and Zn(II) from EXAFS studies) is five coordinate in VanX in contrast to previous crystallographic studies (Bussiere, D. E., Pratt, S. D., Katz, L., Severin, J. M., Holzman, T., and Park, C. H. (1998) Mol. Cell 2, 75-84). These spectroscopic studies also demonstrate that the metal ion in Co(II)-substituted VanX when complexed with a phosphinate analog of substrate D-Ala-D-Ala is also five-coordinate. PMID- 15657054 TI - Pim-1 ligand-bound structures reveal the mechanism of serine/threonine kinase inhibition by LY294002. AB - Pim-1 is an oncogene-encoded serine/threonine kinase primarily expressed in hematopoietic and germ cell lines. Pim-1 kinase was originally identified in Maloney murine leukemia virus-induced T-cell lymphomas and is associated with multiple cellular functions such as proliferation, survival, differentiation, apoptosis, and tumorigenesis (Wang, Z., Bhattacharya, N., Weaver, M., Petersen, K., Meyer, M., Gapter, L., and Magnuson, N. S. (2001) J. Vet. Sci. 2, 167-179). The crystal structures of Pim-1 complexed with staurosporine and adenosine were determined. Although a typical two-domain serine/threonine protein kinase fold is observed, the inter-domain hinge region is unusual in both sequence and conformation; a two-residue insertion causes the hinge to bulge away from the ATP binding pocket, and a proline residue in the hinge removes a conserved main chain hydrogen bond donor. Without this hydrogen bond, van der Waals interactions with the hinge serve to position the ligand. The hinge region of Pim-1 resembles that of phosphatidylinositol 3-kinase more closely than it does other protein kinases. Although the phosphatidylinositol 3-kinase inhibitor LY294002 also inhibits Pim 1, the structure of the LY294002.Pim-1 complex reveals a new binding mode that may be general for Ser/Thr kinases. PMID- 15657057 TI - Microfibrils at basement membrane zones interact with perlecan via fibrillin-1. AB - Mutational defects in fibrillin-rich microfibrils give rise to a number of heritable connective tissue disorders, generally termed microfibrillopathies. To understand the pathogenesis of these microfibrillopathies, it is important to elucidate the supramolecular composition of microfibrils and their interaction properties with extracellular matrix components. Here we demonstrate that the proteoglycan perlecan is an associated component of microfibrils typically close to basement membrane zones. Double immunofluorescence studies demonstrate colocalization of fibrillin-1, the major backbone component of microfibrils, with perlecan in fibroblast cultures as well as in dermal and ocular tissues. Double immunogold labeling further confirms colocalization of perlecan to microfibrils in various tissues at the ultrastructural level. Extraction studies revealed that perlecan is not covalently associated with microfibrils. High affinity interactions between fibrillin-1 and perlecan were found by kinetic binding studies with dissociation constants in the low nanomolar range. A detailed mapping study of the interaction epitopes by solid phase binding assays primarily revealed interactions of perlecan domains I and II with a central region of fibrillin-1. Analysis of perlecan null embryos showed less microfibrils at the dermal-epidermal junction as compared with wild-type littermates. The data presented indicate a functional significance for perlecan in anchoring microfibrils to basement membranes and in the biogenesis of microfibrils. PMID- 15657056 TI - Mechanisms responsible for the promoter-specific effects of myocardin. AB - Understanding the mechanism of smooth muscle cell (SMC) differentiation will provide the foundation for elucidating SMC-related diseases such as atherosclerosis, restenosis, and asthma. Recent studies have demonstrated that the interaction of SRF with the co-activator myocardin is a critical determinant of smooth muscle development. It has been proposed that the specific transcriptional activation of smooth muscle-restricted genes (as opposed to other SRF-dependent genes) by myocardin results from the presence of multiple CArG boxes in smooth muscle genes that facilitate myocardin homodimer formation. This proposal was further tested in the current study. Our results show that the SMC specific telokin promoter, which contains only a single CArG box, is strongly activated by myocardin. Furthermore, myocardin and a dimerization defective mutant myocardin induce expression of endogenous telokin but not c-fos in 10T1/2 fibroblast cells. Knocking down myocardin by small interfering RNA decreased telokin promoter activity and expression in A10 SMCs. A series of telokin and c fos promoter chimeric and mutant reporter genes was generated to determine the mechanisms responsible for the promoter-specific effects of myocardin. Data from these experiments demonstrated that the ets binding site in the c-fos promoter partially blocks the activation of this promoter by myocardin. However, the binding of ets factors alone was not sufficient to explain the promoter-specific effects of myocardin. Elements 3' of the CArG box in the c-fos promoter act in concert with the ets binding site to block the ability of myocardin to activate the promoter. Conversely, elements 5' and 3' of the CArG box in the telokin promoter act in concert with the CArG box to facilitate myocardin stimulation of the promoter. Together these data suggest that the promoter specificity of myocardin is dependent on complex combinatorial interactions of multiple cis elements and their trans binding factors. PMID- 15657058 TI - The role of the regulatory subunit of fission yeast calcineurin for in vivo activity and its relevance to FK506 sensitivity. AB - Calcineurin, a protein phosphatase required for Ca2+ signaling in many cell types, is a heterodimer composed of catalytic and regulatory subunits. The fission yeast genome encodes a single set of catalytic (Ppb1) and regulatory (Cnb1) subunits, providing an ideal model system to study the functions of these subunits in vivo. Here, we cloned the cnb1+ gene and showed that the cnb1 knock out (Deltacnb1) exhibits identical phenotypes with Deltappb1 and that overexpression of Ppb1 failed to suppress the phenotypes of Deltacnb1. Interestingly, overexpression of the C-terminal-deleted Ppb1 (Ppb1DeltaC), the constitutively active form of Ppb1, also failed to suppress the phenotypes of Deltacnb1. FK506 caused MgCl2 sensitivity to the wild-type cells in an FKBP12 dependent manner. Co-overexpression of Ppb1 and Cnb1 suppressed the FK506-induced MgCl2 sensitivity, but the suppression was only partial, suggesting that an excess amount of the Ppb1-Cnb1 complex cannot compete out the FKBP12-FK506 complex. Although overexpression of Ppb1DeltaC alone had little effect on cell growth, co-overexpression of Ppb1DeltaC and Cnb1 caused a distinct growth defect. FK506 suppressed the growth defect when Cnb1 was co-expressed using the attenuated nmt1 promoter, but it failed to suppress the defect when Cnb1 was co expressed using the wild-type nmt1 promoter. Knock-out of the prz1+ gene, encoding a downstream target transcription factor of calcineurin, suppressed the growth defect irrespective of the promoter potency. These results suggest that Cnb1 is essential for the activation of calcineurin and that the activated calcineurin is the pharmacological target of the FKBP12-FK506 complex in vivo. PMID- 15657059 TI - Ecdysone-mediated up-regulation of the effector caspase DRICE is required for hormone-dependent apoptosis in Drosophila cells. AB - The Drosophila steroid hormone ecdysone mediates cell death during metamorphosis by regulating the transcription of a number of cell death genes. The apical caspase DRONC is known to be transcriptionally regulated by ecdysone during development. Here we demonstrate that ecdysone also regulates the transcription of DRICE, a major effector caspase and a downstream target for DRONC in the fly. Using RNA interference in an ecdysone-responsive Drosophila cell line, we show that drice up-regulation is essential for apoptosis induced by ecdysone. We also show that drice expression is specifically controlled by the ecdysone-regulated transcription factor BR-C. Combined with previous observations, our results indicate that transcriptional regulation of the components of the core apoptotic machinery plays a key role in hormone-regulated programmed cell death during Drosophila development. PMID- 15657060 TI - c-Abl tyrosine kinase regulates caspase-9 autocleavage in the apoptotic response to DNA damage. AB - Activation of the initiator caspase-9 is essential for induction of apoptosis by developmental signals, oncogenic transformation, and genotoxic stress. The c-Abl tyrosine kinase is also involved in the apoptotic response to DNA damage. The present results demonstrate that c-Abl binds directly to caspase-9. We show that c-Abl phosphorylates caspase-9 on Tyr-153 in vitro and in cells treated with DNA damaging agents. Moreover, inhibition of c-Abl with STI571 blocked DNA damage induced autoprocessing of caspase-9 to the p35 subunit and activation of caspase 3. Caspase-9(Y153F) also attenuated DNA damage-induced processing of caspase-9 to p35, activation of caspase-3, and apoptosis. These findings indicate that caspase 9 autoprocessing is regulated by c-Abl in the apoptotic response to genotoxic stress. PMID- 15657061 TI - High affinity interaction with filamin A protects against calcium-sensing receptor degradation. AB - Calcium-sensing receptors (CaR) regulate cell proliferation, differentiation, and apoptosis through the MAPK pathway. MAPK pathway activation requires the cytoskeletal scaffold protein filamin A. Here we examine the interactions of CaR with filamin A in HEK-293 and M2 or A7 melanoma cells to determine how interactions with filamin A facilitate signaling. Filamin A interacts with CaR through two predicted beta-strands from residues 962 to 981; interactions between filamin A and CaR are greatly enhanced by exposure to 5 mM Ca2+. Truncations or deletions (from 972 to 997 or 962 to 981) of the CaR carboxyl terminus eliminate high affinity interactions with filamin A, but CaR-mediated MAPK pathway activation still occurs. CaR-mediated ERK phosphorylation can be localized to a predicted alpha-helix proximal to the membrane, which has been shown to be important for G protein-mediated signaling (residues 868-879). In M2 cells ( filamin A), CaR expression levels are very low; cotransfection of CaR with filamin A increases total cellular CaR and increases plasma membrane localization of CaR, facilitating CaR signaling to the MAPK pathway; similar results were obtained in HEK-293 cells. Interaction with filamin A increases cellular CaR by preventing CaR degradation, thereby facilitating CaR signaling. In addition, filamin A facilitates signaling to the MAPK pathway even by CaR truncations or deletion mutants that cannot engage in high affinity interactions with filamin A, suggesting the targeting of critical signaling proteins to CaR. PMID- 15657062 TI - The molecular chaperone, ClpA, has a single high affinity peptide binding site per hexamer. AB - Substrate recognition by Clp chaperones is dependent on interactions with motifs composed of specific peptide sequences. We studied the binding of short motif bearing peptides to ClpA, the chaperone component of the ATP-dependent ClpAP protease of Escherichia coli in the presence of ATPgammaS and Mg2+ at pH 7.5. Binding was measured by isothermal titration calorimetry (ITC) using the peptide, AANDENYALAA, which corresponds to the SsrA degradation motif found at the C terminus of abnormal nascent polypeptides in vivo. One SsrA peptide was bound per hexamer of ClpA with an association constant (K(A)) of 5 x 10(6) m(-1). Binding was also assayed by changes in fluorescence of an N-terminal dansylated SsrA peptide, which bound with the same stoichiometry of one per ClpA hexamer (K(A) approximately 1 x 10(7) m(-1)). Similar results were obtained when ATP was substituted for ATPgammaS at 6 degrees C. Two additional peptides, derived from the phage P1 RepA protein and the E. coli HemA protein, which bear different substrate motifs, were competitive inhibitors of SsrA binding and bound to ClpA hexamers with K(A)' > 3 x 10(7) m(-1). DNS-SsrA bound with only slightly reduced affinity to deletion mutants of ClpA missing either the N-terminal domain or the C-terminal nucleotide-binding domain, indicating that the binding site for SsrA lies within the N-terminal nucleotide-binding domain. Because only one protein at a time can be unfolded and translocated by ClpA hexamers, restricting the number of peptides initially bound should avoid nonproductive binding of substrates and aggregation of partially processed proteins. PMID- 15657064 TI - The transcriptional activity of estrogen receptor-alpha is dependent on Ca2+/calmodulin. AB - Estrogen binds to estrogen receptors in cells, thereby activating the receptors and eliciting biological effects. One of the best characterized effects of estrogen receptor-alpha (ERalpha) is transcriptional activation that regulates selected target genes in the nucleus. Work from several laboratories has documented a Ca2+-dependent interaction between ERalpha and calmodulin. In addition, we previously showed that antagonism of calmodulin function in cells prevented estradiol from inducing ERalpha transcriptional activity, suggesting that association of ERalpha with calmodulin participates in ERalpha function. In this study we adopted a multifaceted approach to directly address this hypothesis. The calmodulin binding domain on ERalpha was identified and several mutant ERalpha constructs unable to bind calmodulin were generated. Elimination of calmodulin binding prevented estradiol from stimulating ERalpha transcriptional activation. Essentially identical results were obtained when intracellular Ca2+ was chelated with the cell-permeable chelator 1,2-bis(o aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetra(acetoxymethyl)ester (BAPTA AM). Moreover, CaM1234, a calmodulin mutant that is unable to bind Ca2+, functioned as a dominant negative construct. Transfection of cells with CaM1234 reduced estradiol-stimulated ERalpha transcriptional activity. These data indicate that binding to calmodulin is required for normal transcriptional function of ERalpha. PMID- 15657063 TI - Acidic extracellular pH induces matrix metalloproteinase-9 expression in mouse metastatic melanoma cells through the phospholipase D-mitogen-activated protein kinase signaling. AB - The extracellular pH (pHe) of tumor tissues is often acidic, which can induce the expression of several proteins. We previously showed that production of matrix metalloproteinase-9 (MMP-9) was induced by culturing cells at acidic pHe (5.4 6.5). Here we have investigated the signal transduction pathway by which acidic pHe induces MMP-9 expression. We found that acidic pHe (5.9) activated phospholipase D (PLD), and inhibition of PLD activity by 1-butanol and Myr-ARF6 suppressed the acidic pHe-induced MMP-9 expression. Exogenous PLD, but not phosphatidylinositol-specific PLC or PLA2, mimicked MMP-9 induction by acidic pHe. Western blot analysis revealed that acidic pHe increased the steady-state levels of phosphorylated extracellular signal-regulated kinases 1/2 and p38 and that the PLD inhibitors suppressed these increases. Using 5'-deletion mutant constructs of the MMP-9 promoter, we found that the acidic pHe-responsive region was located at nucleotide -670 to -531, a region containing the NF kappa B binding site. A mutation into the NF kappa B binding site reduced, but not completely, the acidic pHe-induced MMP-9 promoter activity, and NF kappa B activity was induced by acidic pHe. Pharmacological inhibitors specific for mitogen-activated protein kinase kinase 1/2 (PD098059) and p38 (SB203580) attenuated the acidic pHe-induced NF kappa B activity and MMP-9 expression. These data suggest that PLD, mitogen-activated protein kinases (extracellular signal regulated kinases 1/2 and p38), and NF kappa B mediate the acidic pHe signaling to induce MMP-9 expression. A transcription factor(s) other than NF kappa B may also be involved in the MMP-9 expression. PMID- 15657065 TI - Tyrosine nitration on p65: a novel mechanism to rapidly inactivate nuclear factor kappaB. AB - NO is an important factor that induces post-translational modifications of proteins by cellular reduction and oxidation mechanism: cysteinyl-nitrosylation or Tyr nitration. Nuclear factor (NF)-kappaB activity can be rapidly suppressed by sodium nitroprusside, a NO donor. This effect was effectively reversed by peroxynitrite scavenger deferoxamine, suggesting a Tyr nitration-mediated mechanism. Western blot with nitrotyrosine-specific antibody demonstrated that the p65 subunit of NF-kappaB was predominantly nitrated on Tyr residues. Tyr nitration of p65 induced its dissociation from p50, its association with IkappaBalpha, and subsequent sequestration of p65 in the cytoplasm by IkappaBalpha-mediated export. Liquid chromatography-coupled nanoelectrospray mass spectrometry revealed specific nitration on Tyr-66 and Tyr-152 residues of p65. Mutation studies confirmed that both Tyr-66 and Tyr-152 residues were important for the direct effects of NO on p65, which resulted in more p65 export and inactivation of NF-kappaB activity. This study identified a novel and efficient pathway where NO rapidly inactivated NF-kappaB activity by inducing Tyr nitration on p65. PMID- 15657066 TI - Coregulator interactions with the thyroid hormone receptor. AB - The thyroid hormone receptor (TR) directly regulates the transcription of thyroid hormone-responsive genes in response to changing levels of thyroid hormone. Mechanistically TR utilizes a complex set of binding interactions, with hormone, response elements, and coregulatory proteins, to provide specific local control of patterns of transcriptional response that are partially responsible for inducing the tissue-selective responses to the circulating hormone. One of the apparently dominant phenomena in the regulation of thyroid hormone responses is the protein interactions between TR and its coregulators. This review summarizes the current state of knowledge with respect to the identity of these coregulators, their interaction with TR, and the consequences of those interactions. PMID- 15657067 TI - Phosphotyrosine signaling networks in epidermal growth factor receptor overexpressing squamous carcinoma cells. AB - Overexpression and enhanced activation of the epidermal growth factor (EGF) receptor are frequent events in human cancers that correlate with poor prognosis. Anti-phosphotyrosine and anti-EGFr affinity chromatography, isotope-coded muLC MS/MS, and immunoblot methods were combined to describe and measure signaling networks associated with EGF receptor activation and pharmacological inhibition. The squamous carcinoma cell line HN5, which overexpresses EGF receptor and displays sustained receptor kinase activation, was used as a model system, where pharmacological inhibition of EGF receptor kinase by erlotinib markedly reduced auto and substrate phosphorylation, Src family phosphorylation at EGFR Y845, while increasing total EGF receptor protein. Diverse sets of known and poorly described functional protein classes were unequivocally identified by affinity selection, comprising either proteins tyrosine phosphorylated or complexed therewith, predominantly through EGF receptor and Src family kinases, principally 1) immediate EGF receptor signaling complexes (18%); 2) complexes involved in adhesion and cell-cell contacts (34%); and 3) receptor internalization and degradation signals. Novel and known phosphorylation sites could be located despite the complexity of the peptide mixtures. In addition to interactions with multiple signaling adaptors Grb2, SHC, SCK, and NSP2, EGF receptors in HN5 cells were shown to form direct or indirect physical interactions with additional kinases including ACK1, focal adhesion kinase (FAK), Pyk2, Yes, EphA2, and EphB4. Pharmacological inhibition of EGF receptor kinase activity by erlotinib resulted in reduced phosphorylation of downstream signaling, for example through Cbl/Cbl B, phospholipase Cgamma (PLCgamma), Erk1/2, PI-3 kinase, and STAT3/5. Focal adhesion proteins, FAK, Pyk2, paxillin, ARF/GIT1, and plakophillin were down regulated by transient EGF stimulation suggesting a complex balance between growth factor induced kinase and phosphatase activities in the control of cell adhesion complexes. The functional interactions between IGF-1 receptor, lysophosphatidic acid (LPA) signaling, and EGF receptor were observed, both direct and/or indirectly on phospho-Akt, phospho-Erk1/2, and phospho-ribosomal S6. PMID- 15657068 TI - Can cardiac pacemakers and magnetic resonance imaging systems co-exist? PMID- 15657069 TI - Folic acid supplementation during methotrexate treatment: nonsense? PMID- 15657070 TI - A randomized controlled trial of intra-articular triamcinolone and/or physiotherapy in shoulder capsulitis. AB - OBJECTIVE: To assess the effectiveness of intra-articular triamcinolone injection and physiotherapy singly or combined in the treatment of adhesive capsulitis of the shoulder. METHODS: Eighty patients with adhesive capsulitis of less than 6 months duration were randomized to one of four groups: Group A, injection of triamcinolone 20 mg and eight sessions of standardized physiotherapy; Group B, injection of triamcinolone 20 mg alone; Group C, placebo injection and eight sessions of standardized physiotherapy; or Group D, placebo injection alone. All subjects were given an identical home exercise programme. Outcome measures were assessed at 6 weeks and 16 weeks. The primary outcome measure was Shoulder Disability Questionnaire (SDQ) score. Secondary outcomes were measurement of pain using a visual analogue scale (VAS), global disability using VAS and range of passive external rotation. A two-way analysis of variance was used to explore the effects of corticosteroid injection and physiotherapy. RESULTS: At 6 weeks, the SDQ had improved significantly more in the groups receiving corticosteroid injection (P = 0.004). Physiotherapy improved passive external rotation at 6 weeks (P = 0.02) and corticosteroid injection improved self-assessment of global disability at 6 weeks (P = 0.04). There was no interaction effect between injection and physiotherapy. At 16 weeks, all groups had improved to a similar degree with respect to all outcome measures. CONCLUSION: Corticosteroid injection is effective in improving shoulder-related disability, and physiotherapy is effective in improving the range of movement in external rotation 6 weeks after treatment. PMID- 15657071 TI - Rituximab inefficiency during type I cryoglobulinaemia. PMID- 15657072 TI - Leflunomide in rheumatoid arthritis: recommendations through a process of consensus. AB - OBJECTIVES: To determine, by consensus, the optimal use of leflunomide in rheumatoid arthritis (RA), using a multidisciplinary panel of experts and performing meta-analyses of available data. METHODS: A multidisciplinary panel of experts in RA was convened. Important questions, pertinent to the use of leflunomide in the treatment of RA, were defined by consensus at an initial meeting. Each question was allocated to subgroups of two or three members, who worked separately to prepare a balanced opinion, based on published literature, data from individual patients taking part in phase II and phase III clinical trials provided by Aventis, and data from a USA-based medical claims database (AETNA). The full group then reconvened to agree on an overall consensus statement. Recommendations concerning efficacy and tolerability versus comparator drugs and placebo were derived from two new meta-analyses. RESULTS: Leflunomide was at least as effective as sulphasalazine and methotrexate, and equally well tolerated on meta-analysis of trial data. Overall withdrawal rates for all adverse events were similar for all three drugs. Avoidance of the loading dose reduces 'nuisance' side-effects (e.g. nausea), but probably delays the onset of action. Adverse events could usually be managed by dose reduction and/or symptomatic therapy. CONCLUSIONS: On the basis of efficacy, safety and cost, leflunomide should be considered in patients with RA who have failed first-line DMARD drug therapy. In refractory cases, leflunomide may be used in combination with, for example, methotrexate before biological agents. Therapy should be initiated by a specialist, but repeat prescribing in general practice on a shared care basis is acceptable using agreed protocols. Clear mechanisms are required to monitor toxicity, with good communication between the patient and rheumatologist to manage nuisance side-effects and avoid unnecessary discontinuation of leflunomide. PMID- 15657073 TI - Downregulation of Par3 and aPKC function directs cells towards the ICM in the preimplantation mouse embryo. AB - Generation of inside cells that develop into inner cell mass (ICM) and outside cells that develop into trophectoderm is central to the development of the early mouse embryo. Critical to this decision is the development of cell polarity and the associated asymmetric (differentiative) divisions of the 8-cell-stage blastomeres. The underlying molecular mechanisms for these events are not understood. As the Par3/aPKC complex has a role in establishing cellular polarity and division orientation in other systems, we explored its potential function in the developing mouse embryo. We show that both Par3 and aPKC adopt a polarized localization from the 8-cell stage onwards and that manipulating their function re-directs cell positioning and consequently influences cell fate. Injection of dsRNA against Par3 or mRNA for a dominant negative form of aPKC into a random blastomere at the 4-cell stage directs progeny of the injected cell into the inside part of the embryo. This appears to result from both an increased frequency by which such cells undertake differentiative divisions and their decreased probability of retaining outside positions. Thus, the natural spatial allocation of blastomere progeny can be over-ridden by downregulation of Par3 or aPKC, leading to a deceased tendency for them to remain outside and so develop into trophectoderm. In addition, this experimental approach illustrates a powerful means of manipulating gene expression in a specific clonal population of cells in the preimplantation embryo. PMID- 15657074 TI - Expression of junctional adhesion molecule-A prevents spontaneous and random motility. AB - Junctional adhesion molecule-A (JAM-A) is a cell-surface glycoprotein that localizes to intercellular junctions and associates with intracellular proteins via PSD95-Dlg-ZO1-binding residues. To define the functional consequences of JAM A expression, we have produced endothelial cells from JAM-A-deficient mice. We report here that the absence of JAM-A enhanced spontaneous and random motility. In turn, the enhanced motility of JAM-A-negative cells was abrogated either on transfection of exogenous JAM-A or on treatment with inhibitors of glycogen synthase kinase-3beta (GSK-3beta). In addition, in JAM-A-positive cells, motility was enhanced on inactivation of protein kinase Czeta (PKCzeta), which is an inhibitor of GSK-3beta. Although these findings suggested that JAM-A might inhibit GSK-3beta, we found that expression per se of JAM-A did not change the levels of inactive GSK-3beta. Thus, JAM-A expression may regulate effectors of motility that are also downstream of the PKCzeta/GSK-3beta axis. In support of this view, we found that JAM-A absence increased the number of actin-containing protrusions, reduced the stability of microtubules and impaired the formation of focal adhesions. Notably, all the functional consequences of JAM-A absence were reversed either on treatment with GSK-3beta inhibitors or on transfection of full length JAM-A, but not on transfection of a JAM-A deletion mutant devoid of the PSD95-Dlg-ZO1-binding residues. Thus, by regulating cytoskeletal and adhesive structures, JAM-A expression prevents cell motility, probably in a PSD95-Dlg-ZO1 dependent manner. PMID- 15657075 TI - Direct transport across the plasma membrane of mammalian cells of Leishmania HASPB as revealed by a CHO export mutant. AB - Leishmania HASPB is a lipoprotein that is exported to the extracellular space from both Leishmania parasites and mammalian cells via an unconventional secretory pathway. Exported HASPB remains anchored in the outer leaflet of the plasma membrane mediated by myristate and palmitate residues covalently attached to the N-terminal SH4 domain of HASPB. HASPB targeting to the plasma membrane depends on SH4 acylation that occurs at intracellular membranes. How acylated HASPB is targeted to the plasma membrane and, in particular, the subcellular site of HASPB membrane translocation is unknown. In order to address this issue, we screened for clonal CHO mutants that are incapable of exporting HASPB. A detailed characterization of such a CHO mutant cell line revealed that the expression level of the HASPB reporter molecule is unchanged compared to CHO wild-type cells; that it is both myristoylated and palmitoylated; and that it is mainly localized to the plasma membrane as judged by confocal microscopy and subcellular fractionation. However, based on a quantitative flow cytometry assay and a biochemical biotinylation assay of surface proteins, HASPB transport to the outer leaflet of the plasma membrane is largely reduced in this mutant. From these data, we conclude that the subcellular site of HASPB membrane translocation is the plasma membrane as the reporter molecule accumulates in this location when export is blocked. Thus, these results allow us to define a two-step process of HASPB cell surface biogenesis in which SH4 acylation of HASPB firstly mediates intracellular targeting to the plasma membrane. In a second step, the plasma membrane-resident machinery, which is apparently disrupted in the CHO mutant cell line, mediates membrane translocation of HASPB. Intriguingly, the angiogenic growth factor FGF-2, another protein secreted by unconventional means, is shown to be secreted normally from the HASPB export mutant cell line. These observations demonstrate that the export machinery component defective in the export mutant cell line functions specifically in the HASPB export pathway. PMID- 15657076 TI - Integrin-dependent PLC-gamma1 phosphorylation mediates fibronectin-dependent adhesion. AB - Although integrin engagement initiates signaling events such as focal-adhesion kinase (FAK) and Src kinase activation, the role of phosphoinositide turnover in cell adhesion is less clear. To assess PLC-gamma1 function in this process, Plcg1(-/-) fibroblasts (Null) were compared with the same fibroblasts in which PLC-gamma1 was re-expressed (Null+). Following plating on fibronectin, Null cells displayed a significantly impaired rate of adhesion compared with Null+ cells. This defect was detected at low concentrations of fibronectin; at high fibronectin concentrations, the Null and Null+ cells displayed equivalent adhesion characteristics. The differences were not due to PLC-gamma1-dependent changes in integrin subunit expression, nor was integrin receptor clustering impaired with the absence of PLC-gamma1. Experiments with site-specific antibodies and PLC-gamma1 mutants showed that fibronectin selectively increased phosphorylation of Tyr783 and that mutagenesis of this residue, but not Tyr771 or Tyr1253, abrogated fibronectin-dependent adhesion. The SH2 domains of PLC-gamma1 were also required for maximal adhesion on fibronectin. Adhesion to fibronectin induced PLC-gamma1 tyrosine phosphorylation that was inhibited by a Src-kinase inhibitor, but not an epidermal-growth-factor-receptor kinase inhibitor. Moreover, in cells null for Src family members, but not in cells null for FAK family members, integrin-dependent PLC-gamma1 tyrosine phosphorylation was greatly reduced. Finally, the data demonstrated that PLC-gamma1 co immunoprecipitated with Src following fibronectin-induced integrin activation, and this association did not depend on FAK expression. PMID- 15657078 TI - Induction of apoptosis by TNF receptor 2 in a T-cell hybridoma is FADD dependent and blocked by caspase-8 inhibitors. AB - Previously we reported that both human TNFR1 and TNFR2 mediate TNF-induced apoptosis in the transfected rat/mouse T cell hybridoma PC60. We show here that TNFR2-mediated apoptosis in PVC60 cells can be blocked by the broad-spectrum caspase inhibitor zVAD-fmk, the caspase-8 inhibitor zIETD-fmk and by CrmA, a viral inhibitor of caspase-1 and caspase-8. This suggests an involvement of caspase-8 in TNFR2-mediated apoptosis. The upstream adaptor of caspase-8, FADD, is also involved in TNFR2-induced cell death, since transient overexpression of a dominant negative deletion mutant of FADD inhibited apoptosis induced by this receptor. TNFR2-induced apoptosis is independent of endogenous TNF or other death inducing ligand production and subsequent activation of TNFR1 or other death receptors. Furthermore, TNFR2 stimulation does not enhance sensitivity for a subsequent TNFR1-induced apoptotic signal, as has been reported for Jurkat cells. TRAF2 downregulation, which has been proposed as the mechanism by which TNFR2 enhances TNFR1 signaling, was observed in PC60 cells, but the TNRF1 signal was not modulated. These data confirm the capacity of TNFR2 to generate an apoptotic cell death signal independent of TNFR1. PMID- 15657077 TI - TRIP6 is a RIP2-associated common signaling component of multiple NF-kappaB activation pathways. AB - Receptor-interacting protein 2 (RIP2) is a member of the RIP kinase family that has been shown to be crucially involved in inflammation, innate and adaptive immune responses. The physiological and pathological roles of RIP2 are mediated through its involvement in multiple NF-kappaB activation pathways, including those triggered by tumor necrosis factor (TNF), interleukin 1 (IL-1), Toll-like receptor 2 (TLR2), TLR3, TLR4 and Nod1. In this report, we identified the LIM domain-containing protein TRIP6 as a RIP2-interacting protein in yeast two-hybrid screens. In mammalian cells, TRIP6 interacts with RIP2 in a TNF- or IL-1 dependent manner. Overexpression of TRIP6 potentiates RIP2-mediated NF-kappaB activation in a dose-dependent manner. The LIM domains of TRIP6 are responsible for its interaction with RIP2. TRIP6 also interacts with TRAF2, a protein that is crucially involved in TNF signaling, as well as the IL-1 receptor, TLR2 and Nod1. Overexpression of TRIP6 potentiates NF-kappaB activation by TNF, IL-1, TLR2 or Nod1, whereas a dominant negative mutant or RNA-interference construct of TRIP6 inhibits NF-kappaB activation by TNF, IL-1, TLR2 or Nod1. Moreover, TRIP6 also potentiates RIP2- and Nod1-mediated ERK activation. These data have established a physical and functional association between TRIP6 and RIP2, and suggest that RIP2's involvement in multiple NF-kappaB and ERK activation pathways is mediated through TRIP6. PMID- 15657079 TI - Mechanism of early biphasic activation of poly(ADP-ribose) polymerase-1 in response to ultraviolet B radiation. AB - The damage to DNA caused by ultraviolet B radiation (280-320 nm) contributes significantly to development of sunlight-induced skin cancers. The susceptibility of mice to ultraviolet B-induced skin carcinogenesis is increased by an inhibitor of the DNA damage-activated nuclear enzyme poly(ADP-ribose) polymerase-1 (PARP), hence PARP activation is likely to be associated with cellular responses that suppress carcinogenesis. To understand the role of activated PARP in these cellular functions, we need to first clearly identify the cause of PARP activation in ultraviolet B-irradiated cells. Ultraviolet B, like ultraviolet C, causes direct DNA damage of cyclobutane pyrimidine dimer and 6, 4-photoproduct types, which are subjected to the nucleotide excision repair. Moreover, ultraviolet B also causes oxidative DNA damage, which is subjected to base excision repair. To identify which of these two types of DNA damage activates PARP, we examined mechanism of early PARP activation in mouse fibroblasts exposed to ultraviolet B and C radiations. The ultraviolet B-irradiated cells rapidly activated PARP in two distinct phases, initially within the first 5 minutes and later between 60-120 minutes, whereas ultraviolet C-irradiated cells showed only the immediate PARP activation. Using antioxidants, local irradiation, chromatin immunoprecipitation and in vitro PARP assays, we identified that ultraviolet radiation-induced direct DNA damage, such as thymine dimers, cause the initial PARP activation, whereas ultraviolet B-induced oxidative damage cause the second PARP activation. Our results suggest that cells can selectively activate PARP for participation in different cellular responses associated with different DNA lesions. PMID- 15657080 TI - Stromal-epithelial interactions in aging and cancer: senescent fibroblasts alter epithelial cell differentiation. AB - Cellular senescence suppresses cancer by arresting cells at risk of malignant tumorigenesis. However, senescent cells also secrete molecules that can stimulate premalignant cells to proliferate and form tumors, suggesting the senescence response is antagonistically pleiotropic. We show that premalignant mammary epithelial cells exposed to senescent human fibroblasts in mice irreversibly lose differentiated properties, become invasive and undergo full malignant transformation. Moreover, using cultured mouse or human fibroblasts and non malignant breast epithelial cells, we show that senescent fibroblasts disrupt epithelial alveolar morphogenesis, functional differentiation and branching morphogenesis. Furthermore, we identify MMP-3 as the major factor responsible for the effects of senescent fibroblasts on branching morphogenesis. Our findings support the idea that senescent cells contribute to age-related pathology, including cancer, and describe a new property of senescent fibroblasts - the ability to alter epithelial differentiation - that might also explain the loss of tissue function and organization that is a hallmark of aging. PMID- 15657081 TI - Phototactic activity in Chlamydomonas 'non-phototactic' mutants deficient in Ca2+ dependent control of flagellar dominance or in inner-arm dynein. AB - In the mechanism underlying the phototactic behavior of Chlamydomonas, Ca(2+) has been thought to control the dominance between the two flagella so as to steer the cell to correct directions. A newly isolated mutant, lsp1, that displays weak phototaxis was found to be defective in this Ca(2+)-dependent shift in flagellar dominance; in demembranated and reactivated cell models, the trans flagellum (the flagellum farthest from the eyespot) beat more strongly than the other (the cis flagellum) in about half of the cells regardless of the Ca(2+) concentration between <10(-9) M and 10(-6) M, a range over which wild-type cell models display switching of flagellar dominance. This is unexpected because ptx1, another mutant that is also deficient in flagellar dominance control, has been reported to lack phototactic ability. We therefore re-examined ptx1 and another reportedly non phototactic mutant, ida1, which lacks inner arm dynein subspecies f (also called I1). Both were found to retain reduced phototactic abilities. These results indicate that both Ca(2+)-dependent flagellar dominance control and inner-arm dynein subspecies f are important for phototaxis, but are not absolutely necessary. Analysis of the flagellar beat frequency in lsp1 cell models showed that both of the flagella beat at the frequency of the cis flagellum in wild type. In addition, lsp1 and ptx1 were found to be deficient in determining the sign of phototactic migration. Hence, the Ca(2+)-dependent flagellar dominance control detected in demembranated cells might be involved in the determination of the sign of phototaxis. The gene responsible for the lsp1 mutation was identified by phenotype rescue experiments and found to have sequences for phosphorylation. PMID- 15657082 TI - Endofin recruits clathrin to early endosomes via TOM1. AB - TOM1 and its related proteins, TOM1-like1 (TOM1-L1) and TOM1-like2 (TOM1-L2), constitute a subfamily of the VHS domain protein family. We have recently shown that endofin, a FYVE domain protein associated with the early endosome, is able to recruit cytosolic TOM1 onto endosomal membranes. To reveal the biological consequence of endofin-mediated endosomal recruitment of TOM1, we have identified the clathrin heavy chain as a major interacting protein for TOM1. Optimal clathrin binding by TOM1 involves three sites: residues 300-321, 321-326 and a putative clathrin-binding box at residues 362-366 ((362)LEDEF(366)). Although residues 321-326 could function independently as a weak clathrin-binding motif, deletion of amino acids 300-321 or mutation of (362)Leu and (364)Asp to Ala residues reduced the binding of clathrin to TOM1. A fragment lacking amino acids 300-322 and containing (362)Leu and (364)Asp to Ala mutations lost the ability to interact with clathrin. Remarkably, overexpression of endofin led to a massive and specific recruitment of clathrin [but not dynamin, or the adaptor protein (AP) complexes, AP1, AP2 or AP3] onto endofin-positive endosomes. Although SARA is homologous to endofin, it did not interact with the C-terminal region of TOM1. Examination of chimeric proteins of endofin and SARA suggests that the C-terminal half of endofin is responsible for interaction with the C-terminal region of TOM1 and for recruitment of TOM1 and clathrin to endosomes. The correlation between the ability of endofin to interact with the C-terminal domain of TOM1 and clathrin recruitment suggests that endofin may recruit clathrin via TOM1. Indeed, a chimeric protein consisting of TOM1 fused to two FYVE domains derived from endofin has the ability to recruit clathrin onto endosomal structures. Moreover, we show that affinity-purified TOM1 antibody can abolish binding of clathrin to the C-terminal region of TOM1. Upon microinjection into cells, this antibody reduced the membrane association of clathrin. These results, taken together, suggest that TOM1 is an important molecule for membrane recruitment of clathrin, and that endofin is able to exploit this recruitment at the endosome. PMID- 15657083 TI - Dissection of brefeldin A-sensitive and -insensitive steps in apicoplast protein targeting. AB - The apicoplast is a relict plastid found in many apicomplexans, including the pathogens Toxoplasma gondii and Plasmodium falciparum. Nucleus-encoded apicoplast proteins enter the ER, and after cleavage of the signal sequence, are routed to the apicoplast by virtue of a transit peptide, which is subsequently removed. To assess the mechanisms of localization we examined stable transfectants of T. gondii for the localization and processing of various GFP fusion proteins. GFP fusions bearing apicoplast targeting sequences targeted efficiently to the plastid, with no retention in the ER, even when an ER retention/retrieval sequence was added. Incubation with brefeldin A, which blocks ER-to-Golgi trafficking by inhibiting a GTP exchange factor required for retrograde trafficking, blocked the processing of the protein. Surprisingly, it did not affect the immunofluorescence pattern. To avoid the potentially misleading presence of pre-existing GFP fusion protein in the apicoplast, we used a ligand regulated aggregation system to arrest the GFP fusion protein in the ER prior to trafficking. Upon addition of ligand to promote disaggregation, the fusion protein targeted to the plastid, even in the presence of brefeldin A. Ligand release at 15 degrees C, which blocks trafficking of Golgi-routed proteins, also allowed significant localization to the plastid. Our data indicate that apicoplast proteins can localize to the region of the plastid when Golgi trafficking is inhibited, but suggest that some steps in import or maturation of the proteins may require a brefeldin A-sensitive GTP exchange factor. PMID- 15657084 TI - Papillote and Piopio: Drosophila ZP-domain proteins required for cell adhesion to the apical extracellular matrix and microtubule organization. AB - Adhesion between epithelial cells and extracellular substrates is normally mediated through basal adhesion complexes. However, some cells also possess comparable junctions on their apical surface. Here, we describe two new Drosophila proteins, Piopio and Papillote, that are required for the link between the apical epithelial surface and the overlying apical extracellular matrix (aECM). The two proteins share a zona pellucida (ZP) domain with mammalian aECM components, including the tectorins found in the vertebrate inner ear. Tagged versions of both proteins localized to the apical epithelial surface. Mutations in piopio, papillote and dumpy (another gene encoding a ZP-domain protein) cause defects in the innermost layer of the aECM and its detachment from the epidermis. Loss of Piopio, but not Papillote or Dumpy, causes the absence of specialized microtubule bundles from pupal wings, suggesting that Piopio plays a role in microtubule organization. Thus, ZP domain-containing proteins may have shared functions within the aECM, while also exhibiting specific interactions with the cytoskeleton. PMID- 15657085 TI - Satellite DNA binding and cellular localisation of RNA helicase P68. AB - We purified a 68-kDa protein from the mouse nuclear matrix using ion exchange and affinity chromatography. Column fractions were tested for specific binding to mouse minor satellite DNA using a gel mobility shift assay. The protein was identified by mass spectrometry as RNA helicase P68. In fixed cells, P68 was found to shuttle in and out of SC35 domains, forming fibres and granules in a cell-cycle dependent manner. Analysis of the P68 sequence revealed a short potential coiled-coil domain that might be involved in the formation of P68 fibres. Contacts between centromeres and P68 granules were observed during all phases of the cycle but they were most prominent in mitosis. At this stage, P68 was found in both the centromeric regions and the connections between chromosomes. Direct interaction of P68/DEAD box RNA helicase with satellite DNAs in vitro has not been demonstrated for any other members of the RNA helicase family. PMID- 15657086 TI - Dynamics and interaction of caveolin-1 isoforms with BMP-receptors. AB - Caveolae are small invaginations of the cell membrane that are thought to play a role in important physiological functions such as cell surface signaling, endocytosis and intracellular cholesterol transport. Caveolin-1 is a key protein in these domains and contributes to the organization of cholesterol and saturated lipids within these vesicular invaginations of the plasma membrane. Caveolae are thought to be involved in the signaling of tyrosine kinase receptors and serine threonine receptors. In this article we focus on the involvement of caveolae in the signal transduction of bone morphogenetic proteins (BMPs). BMPs play important roles during embryonic development and especially in chondrogenesis, osteogenesis, neurogenesis and hematopoiesis. The initiation of the signal tranduction starts by the binding of a BMP to a corresponding set of BMP receptors. Using image cross-correlation spectroscopy, we show that the BMP receptors BRIa and BRII colocalize with caveolin-1 isoforms alpha and beta on the cell surface. BRIa colocalizes predominantly with the caveolin-1 alpha isoform. Coexpression of BRII leads to a redistribution of BRIa into domains enriched in caveolin-1 beta. After stimulation with BMP-2, BRIa moves back into the region with caveolin-1 alpha. BRII is expressed in regions enriched in caveolin-1 alpha and beta. Stimulation of cells with BMP-2 leads to a redistribution of BRII into domains enriched in caveolin-1 alpha. Immunoprecipitation studies using transfected COS-7 cells indicate that BRII binds to caveolin-1 alpha and beta. The binding of BRII to caveolin-1 was verified using A431 cells. Stimulation of starved A431 cells with BMP-2 lead to a release of caveolin-1 from the BMP receptors. We show further that the caveolin-1 beta isoform inhibits BMP signaling whereas the alpha isoform does not. PMID- 15657087 TI - Differential effects of IL-10 on prostaglandin H synthase-2 expression and prostaglandin E2 biosynthesis between spleen and bone marrow macrophages. AB - Different populations of mononuclear phagocytes (MO) show considerable diversity of cellular function including prostaglandin E2 (PGE2) biosynthesis. Certain bacterial components enhance PGE2 biosynthesis differentially in selected populations of MO. Interleukin (IL)-10 is proposed to inhibit modulation of PGE2 biosynthesis by down-regulating prostaglandin G/H synthase-2 (PGHS-2) expression. To assess whether IL-10 regulates PGE2 biosynthesis and PGHS-2 expression, splenic and bone marrow MO were isolated from IL-10-deficient (IL-10(-/-)), C57Bl/6 [wild-type (WT) control], and Balb/c (comparison control) mice and were treated with lipopolysaccharide (LPS) and/or interferon-gamma (IFN-gamma) as a model of bacterial inflammation. LPS-induced PGHS-2 expression was similar for splenic MO isolated from the three strains of mice. However, PGE2 released by LPS treated splenic MO was significantly higher in IL-10(-/-) and Balb/c than in WT cells. In the presence of LPS and IFN-gamma, PGHS-2 expression and PGE2 release by IL-10(-/-) and Balb/c splenic MO were enhanced compared with stimulation with LPS alone or IFN-gamma alone. However, there was no significant increase in PGE2 release from WT splenic MO treated with LPS plus IFN-gamma despite increased PGHS 2 expression. In sharp contrast, PGHS-2 expression and PGE2 release by bone marrow MO were greatly enhanced in IL-10(-/-) cells compared with control cells. Our results indicate that IL-10 regulation of MO PGE2 biosynthesis and PGHS-2 expression is compartment-dependent and that PGE2 production is not linked directly to PGHS-2 levels. Furthermore, our findings emphasize strain-specific differences between C57Bl/6 and Balb/c mice, and Balb/c appears more similar to the IL-10(-/-) than to the C57Bl/6 with respect to prostanoid production. PMID- 15657088 TI - Contraction- and hypoxia-stimulated glucose transport is mediated by a Ca2+ dependent mechanism in slow-twitch rat soleus muscle. AB - Increases in contraction-stimulated glucose transport in fast-twitch rat epitrochlearis muscle are mediated by AMPK- and Ca2+/calmodulin-dependent protein kinase (CAMK)-dependent signaling pathways. However, recent studies provide evidence suggesting that contraction-stimulated glucose transport in slow-twitch skeletal muscle is mediated through an AMPK-independent pathway. The purpose of the present study was to test the hypothesis that contraction-stimulated glucose transport in rat slow-twitch soleus muscle is mediated by an AMPK independent/Ca2+-dependent pathway. Caffeine, a sarcoplasmic reticulum (SR) Ca2+ releasing agent, at a concentration that does not cause muscle contractions or decreases in high-energy phosphates, led to an approximately 2-fold increase in 2 deoxyglucose (2-DG) uptake in isolated split soleus muscles. This increase in glucose transport was prevented by the SR calcium channel blocker dantrolene and the CAMK inhibitor KN93. Conversely, 5-aminoimidazole-4-carboxamide-1-beta-D ribofuranoside (AICAR), an AMPK activator, had no effect on 2-DG uptake in isolated split soleus muscles yet resulted in an approximately 2-fold increase in the phosphorylation of AMPK and its downstream substrate acetyl-CoA carboxylase. The hypoxia-induced increase in 2-DG uptake was prevented by dantrolene and KN93, whereas hypoxia-stimulated phosphorylation of AMPK was unaltered by these agents. Tetanic muscle contractions resulted in an approximately 3.5-fold increase in 2 DG uptake that was prevented by KN93, which did not prevent AMPK phosphorylation. Taken in concert, our results provide evidence that hypoxia- and contraction stimulated glucose transport is mediated entirely through a Ca2+-dependent mechanism in rat slow-twitch muscle. PMID- 15657089 TI - Arginine synthesis does not occur during first-pass hepatic metabolism in the neonatal piglet. AB - We have shown that first-pass intestinal metabolism is necessary for approximately 50% of whole body arginine synthesis from its major precursor proline in neonatal piglets. Furthermore, the intestine is not the site of increased arginine synthesis observed during dietary arginine deficiency. Primed constant intravenous (iv) and intraportal (ip) infusions of L-[U-14C]proline, and iv infusion of either L-[guanido-14C]arginine or L-[4,5-3H]arginine were used to measure first-pass hepatic arginine synthesis in piglets enterally fed either deficient (0.20 g.kg(-1).day(-1)) or generous (1.80 g.kg(-1).day(-1)) quantities of arginine for 5 days. Conversion of arginine to other urea cycle intermediates and arginine recycling were also calculated for both dietary treatments. Arginine synthesis (g.kg(-1).day(-1)) from proline was greater in piglets (P < 0.05) fed the deficient arginine diet in both the presence (generous: 0.07; deficient: 0.17; pooled SE = 0.01) and absence (generous: 0.06; deficient: 0.20; pooled SE = 0.01) of first-pass hepatic metabolism. There was no net arginine synthesis from proline during first-pass hepatic metabolism regardless of arginine intake. Arginine conversion to urea, citrulline, and ornithine was significantly greater (P < 0.05) in piglets fed the generous arginine diet. Calculated arginine fluxes were significantly lower (P = 0.01) for [4,5-3H]arginine than for [guanido 14C]arginine, and the discrepancy between the values was greater in piglets fed the deficient arginine diet (35% vs. 20%). Collectively, these findings show that first-pass hepatic metabolism is not a site of net arginine synthesis and that piglets conserve dietary arginine in times of deficiency by decreasing hydrolysis and increasing recycling. PMID- 15657090 TI - Calcium-sensing receptor activation induces nitric oxide production in H-500 Leydig cancer cells. AB - Nitric oxide (NO) is a versatile second messenger. NO is produced by Leydig cells, where NO is a negative regulator of steroidogenesis. In cancer cells, NO is thought to have mutagenic and proliferative effects. We have previously shown that the calcium-sensing receptor (CaR) has promalignant effects in rat H-500 Leydig cancer cells, a model for humoral hypercalcemia of malignancy. Calcium, the major physiological ligand of the CaR, is a recognized intracellular cofactor in the process of NO production by virtue of its positive modulation of neuronal and endothelial nitric oxide synthase (NOS), but importantly, not of inducible (i) NOS activity. iNOS activity is regulated by changes in its expression level. Therefore, we investigated whether CaR activation changes iNOS expression. We found that high extracellular calcium (Cao2+) upregulates the level of mRNA for iNOS, whereas no change was seen in neuronal or endothelial NOS, as assessed by microarray and real-time PCR, respectively. The high Cao2+-induced iNOS upregulation was also detected by Northern and Western blotting. By quantitative real-time PCR, we showed that calcium maximally upregulates iNOS at 18 h. The effect of calcium was abolished by overexpression of a dominant-negative CaR (R185Q), confirming that the effect of Cao2+ was mediated by the CaR. Cells treated with high calcium had higher NO production than those treated with low calcium, as detected with the NO-specific DAF2-AM dye. This was confirmed in single-cell fluorescence determinations using confocal microscopy. In conclusion, high calcium upregulates the levels of iNOS mRNA and protein as well as NO production in H-500 cells, and the effect of Cao2+ on iNOS expression is mediated by the CaR. PMID- 15657091 TI - Defective insulin signaling in skeletal muscle of the hypertensive TG(mREN2)27 rat. AB - Essential hypertension is frequently associated with insulin resistance of skeletal muscle glucose transport, with a potential role of angiotensin II in the pathogenesis of both conditions. The male heterozygous TG(mREN2)27 rat harbors the mouse transgene for renin, exhibits local elevations in angiotensin II, and is an excellent model of both hypertension and insulin resistance. The present study was designed to investigate the potential cellular mechanisms for insulin resistance in this hypertensive animal model, including an assessment of elements of the insulin-signaling pathway. Compared with nontransgenic, normotensive Sprague-Dawley control rats, male heterozygous TG(mREN2)27 rats displayed elevated (P < 0.05) fasting plasma insulin (74%), an exaggerated insulin response (108%) during an oral glucose tolerance test, and reduced whole body insulin sensitivity. TG(mREN2)27 rats also exhibited decreased insulin-mediated glucose transport and glycogen synthase activation in both the type IIb epitrochlearis (30 and 46%) and type I soleus (22 and 64%) muscles. Importantly, there were significant reductions (approximately 30-50%) in insulin stimulation of tyrosine phosphorylation of the insulin receptor beta-subunit and insulin receptor substrate-1 (IRS-1), IRS-1 associated with the p85 subunit of phosphatidylinositol 3-kinase, Akt Ser473 phosphorylation, and Ser9 phosphorylation of glycogen synthase kinase-3beta in epitrochlearis and soleus muscles of TG(mREN2)27 rats. Soleus muscle triglyceride concentration was 25% greater in the transgenic group compared with nontransgenic animals. Collectively, these data provide the first evidence that the insulin resistance of the hypertensive male heterozygous TG(mREN2)27 rat can be attributed to specific defects in the insulin-signaling pathway in skeletal muscle. PMID- 15657092 TI - Marked attenuation of production of collagen type I from cardiac fibroblasts by dehydroepiandrosterone. AB - Dehydroepiandrosterone (DHEA) is a type of adrenal steroid. The concentrations of DHEA and its sulfate (DHEA-S) in serum reach a peak between the ages of 25 and 30 yr and thereafter decline steadily. It was reported that DHEA-S concentration in humans is inversely related to death from cardiovascular diseases. In this study, we examined the effects of DHEA on regulation of collagen mRNA and collagen synthesis in cultured cardiac fibroblasts. Treatment with DHEA (10(-6) M) resulted in a significant decrease in procollagen type I mRNA expression compared with controls. This was accompanied by a significant decrease in procollagen type I protein accumulation in the medium and also a significant decrease in procollagen type I protein synthesis in the cellular matrix. Furthermore, to confirm in vitro results, we administered DHEA to Sprague-Dawley rats, which were treated with angiotensin II for 8 wk to induce cardiac damage. Procollagen type I mRNA expression was significantly decreased and cardiac fibrosis significantly inhibited in DHEA-treated rat hearts without lowering the systolic blood pressure. These results strongly indicate that DHEA can directly attenuate collagen type I synthesis at the transcriptional level in vivo and in vitro in cardiac fibroblasts. PMID- 15657093 TI - Whole body and forearm substrate metabolism in hyperthyroidism: evidence of increased basal muscle protein breakdown. AB - Thyroid hormones have significant metabolic effects, and muscle wasting and weakness are prominent clinical features of chronic hyperthyroidism. To assess the underlying mechanisms, we examined seven hyperthyroid women with Graves' disease before (Ht) and after (Eut) medical treatment and seven control subjects (Ctr). All subjects underwent a 3-h study in the postabsorptive state. After regional catheterization, protein dynamics of the whole body and of the forearm muscles were measured by amino acid tracer dilution technique using [15N]phenylalanine and [2H4]tyrosine. Before treatment, triiodothyronine was elevated (6.6 nmol/l) and whole body protein breakdown was increased 40%. The net forearm release of phenylalanine was increased in hyperthyroidism (microg.100 ml( 1).min(-1)): -7.0 +/- 1.2 Ht vs. -3.8 +/- 0.8 Eut (P = 0.04), -4.2 +/- 0.3 Ctr (P = 0.048). Muscle protein breakdown, assessed by phenylalanine rate of appearance, was increased (microg.100 ml(-1).min(-1)): 15.5 +/- 2.0 Ht vs. 9.6 +/- 1.4 Eut (P = 0.03), 9.9 +/- 0.6 Ctr (P = 0.02). Muscle protein synthesis rate did not differ significantly. Muscle mass and muscle function were decreased 10-20% before treatment. All abnormalities were normalized after therapy. In conclusion, our results show that hyperthyroidism is associated with increased muscle amino acid release resulting from increased muscle protein breakdown. These abnormalities can explain the clinical manifestations of sarcopenia and myopathy. PMID- 15657094 TI - Pairwise local structural alignment of RNA sequences with sequence similarity less than 40%. AB - MOTIVATION: Searching for non-coding RNA (ncRNA) genes and structural RNA elements (eleRNA) are major challenges in gene finding today as these often are conserved in structure rather than in sequence. Even though the number of available methods is growing, it is still of interest to pairwise detect two genes with low sequence similarity, where the genes are part of a larger genomic region. RESULTS: Here we present such an approach for pairwise local alignment which is based on foldalign and the Sankoff algorithm for simultaneous structural alignment of multiple sequences. We include the ability to conduct mutual scans of two sequences of arbitrary length while searching for common local structural motifs of some maximum length. This drastically reduces the complexity of the algorithm. The scoring scheme includes structural parameters corresponding to those available for free energy as well as for substitution matrices similar to RIBOSUM. The new foldalign implementation is tested on a dataset where the ncRNAs and eleRNAs have sequence similarity <40% and where the ncRNAs and eleRNAs are energetically indistinguishable from the surrounding genomic sequence context. The method is tested in two ways: (1) its ability to find the common structure between the genes only and (2) its ability to locate ncRNAs and eleRNAs in a genomic context. In case (1), it makes sense to compare with methods like Dynalign, and the performances are very similar, but foldalign is substantially faster. The structure prediction performance for a family is typically around 0.7 using Matthews correlation coefficient. In case (2), the algorithm is successful at locating RNA families with an average sensitivity of 0.8 and a positive predictive value of 0.9 using a BLAST-like hit selection scheme. AVAILABILITY: The program is available online at http://foldalign.kvl.dk/ PMID- 15657095 TI - Modeling of signal-response cascades using decision tree analysis. AB - MOTIVATION: Signal transduction cascades governing cell functional responses to stimulatory cues play crucial roles in cell regulatory systems and represent promising therapeutic targets for complex human diseases. however, mathematical analysis of how cell responses are governed by signaling activities is challenging due to their multivariate and non-linear nature. diverse computational methods are potentially available, but most are ineffective for protein-level data that is limited in extent and replication. RESULTS: We apply a decision tree approach to analyze the relationship of cell functional response to signaling activity across a spectrum of stimulatory cues. as a specific example, we studied five intracellular signals influencing fibroblast migration under eight conditions: four substratum fibronectin levels and presence versus absence of epidermal growth factor. we propose techniques for preprocessing and extending the experimental measurement set via interpolative modeling in order to gain statistical reliability. for this specific case study, our approach has 70% overall classification accuracy and the decision tree model reveals insights concerning the combined roles of the various signaling activities in governing cell migration speed. we conclude that decision tree methodology may facilitate elucidation of signal-response cascade relationships and generate experimentally testable predictions, which can be used as directions for future experiments. PMID- 15657096 TI - PIBASE: a comprehensive database of structurally defined protein interfaces. AB - MOTIVATION: In recent years, the Protein Data Bank (PDB) has experienced rapid growth. To maximize the utility of the high resolution protein-protein interaction data stored in the PDB, we have developed PIBASE, a comprehensive relational database of structurally defined interfaces between pairs of protein domains. It is composed of binary interfaces extracted from structures in the PDB and the Probable Quaternary Structure server using domain assignments from the Structural Classification of Proteins and CATH fold classification systems. RESULTS: PIBASE currently contains 158,915 interacting domain pairs between 105,061 domains from 2125 SCOP families. A diverse set of geometric, physiochemical and topologic properties are calculated for each complex, its domains, interfaces and binding sites. A subset of the interface properties are used to remove interface redundancy within PDB entries, resulting in 20,912 distinct domain-domain interfaces. The complexes are grouped into 989 topological classes based on their patterns of domain-domain contacts. The binary interfaces and their corresponding binding sites are categorized into 18,755 and 30,975 topological classes, respectively, based on the topology of secondary structure elements. The utility of the database is illustrated by outlining several current applications. AVAILABILITY: The database is accessible via the world wide web at http://salilab.org/pibase SUPPLEMENTARY INFORMATION: http://salilab.org/pibase/suppinfo.html. PMID- 15657097 TI - Dynamic model based algorithms for screening and genotyping over 100 K SNPs on oligonucleotide microarrays. AB - MOTIVATION: A high density of single nucleotide polymorphism (SNP) coverage on the genome is desirable and often an essential requirement for population genetics studies. Region-specific or chromosome-specific linkage studies also benefit from the availability of as many high quality SNPs as possible. The availability of millions of SNPs from both Perlegen and the public domain and the development of an efficient microarray-based assay for genotyping SNPs has brought up some interesting analytical challenges. Effective methods for the selection of optimal subsets of SNPs spanning the genome and methods for accurately calling genotypes from probe hybridization patterns have enabled the development of a new microarray-based system for robustly genotyping over 100,000 SNPs per sample. RESULTS: We introduce a new dynamic model-based algorithm (DM) for screening over 3 million SNPs and genotyping over 100,000 SNPs. The model is based on four possible underlying states: Null, A, AB and B for each probe quartet. We calculate a probe-level log likelihood for each model and then select between the four competing models with an SNP-level statistical aggregation across multiple probe quartets to provide a high-quality genotype call along with a quality measure of the call. We assess performance with HapMap reference genotypes, informative Mendelian inheritance relationship in families, and consistency between DM and another genotype classification method. At a call rate of 95.91% the concordance with reference genotypes from the HapMap Project is 99.81% based on over 1.5 million genotypes, the Mendelian error rate is 0.018% based on 10 trios, and the consistency between DM and MPAM is 99.90% at a comparable rate of 97.18%. We also develop methods for SNP selection and optimal probe selection. AVAILABILITY: The DM algorithm is available in Affymetrix's Genotyping Tools software package and in Affymetrix's GDAS software package. See http://www.affymetrix.com for further information. 10 K and 100 K mapping array data are available on the Affymetrix website. PMID- 15657098 TI - Mtreemix: a software package for learning and using mixture models of mutagenetic trees. AB - SUMMARY: Mixture models of mutagenetic trees constitute a class of probabilistic models for describing evolutionary processes that are characterized by the accumulation of permanent genetic changes. They have been applied to model the accumulation of chromosomal gains and losses in tumor development and the development of drug resistance-associated mutations in the HIV genome.Mtreemix is a software package for estimating mutagenetic trees mixture models from observed cross-sectional data and for using these models for predictions. We provide programs for model fitting, model selection, simulation, likelihood computation and waiting time estimation. AVAILABILITY: Mtreemix, including source code, documentation, sample data files and precompiled Solaris and Linux binaries, is freely available for non-commercial users at http://mtreemix.bioinf.mpi sb.mpg.de/ PMID- 15657099 TI - Online predicted human interaction database. AB - MOTIVATION: High-throughput experiments are being performed at an ever-increasing rate to systematically elucidate protein-protein interaction (PPI) networks for model organisms, while the complexities of higher eukaryotes have prevented these experiments for humans. RESULTS: The Online Predicted Human Interaction Database (OPHID) is a web-based database of predicted interactions between human proteins. It combines the literature-derived human PPI from BIND, HPRD and MINT, with predictions made from Saccharomyces cerevisiae, Caenorhabditis elegans, Drosophila melanogaster and Mus musculus. The 23,889 predicted interactions currently listed in OPHID are evaluated using protein domains, gene co-expression and Gene Ontology terms. OPHID can be queried using single or multiple IDs and results can be visualized using our custom graph visualization program. AVAILABILITY: Freely available to academic users at http://ophid.utoronto.ca, both in tab-delimited and PSI-MI formats. Commercial users, please contact I.J. CONTACT: juris@ai.utoronto.ca SUPPLEMENTARY INFORMATION: http://ophid.utoronto.ca/supplInfo.pdf. PMID- 15657100 TI - Integrative data analysis for functional prediction: a multi-objective optimization approach. AB - An integrative classification system for functional genomics is introduced. A comparison with a previous study of the yeast mitochondrial proteome is presented. AVAILABILITY: A demonstration prototype, interSearch, is available on request. SUPPLEMENTARY INFORMATION: http://ijsr32.infj.ulster.ac.uk/~e10110731/interSearch. PMID- 15657101 TI - IntegratedMap: a Web interface for integrating genetic map data. AB - SUMMARY: IntegratedMap is a Web application and database schema for storing and interactively displaying genetic map data. Its Web interface includes a menu for direct chromosome/linkage group selection, a search form for selection based on mapped object location and linkage group displays. An overview display provides convenient access to the full range of mapped and anchored object types with genetic locus details, such as numbers, types and names of mapped/anchored objects displayed in a compact scrollable list box that automatically updates based on selected map location and object type. Also, multilinkage group and localized map views are available along with links that can be configured for integration with other Web resources. AVAILABILITY: IntegratedMap is implemented in C#/ASP.NET and the package, including a MySQL schema creation script, is available from http://cggc.agtec.uga.edu/Data/download.asp PMID- 15657102 TI - Use of within-array replicate spots for assessing differential expression in microarray experiments. AB - MOTIVATION: Spotted arrays are often printed with probes in duplicate or triplicate, but current methods for assessing differential expression are not able to make full use of the resulting information. The usual practice is to average the duplicate or triplicate results for each probe before assessing differential expression. This results in the loss of valuable information about genewise variability. RESULTS: A method is proposed for extracting more information from within-array replicate spots in microarray experiments by estimating the strength of the correlation between them. The method involves fitting separate linear models to the expression data for each gene but with a common value for the between-replicate correlation. The method greatly improves the precision with which the genewise variances are estimated and thereby improves inference methods designed to identify differentially expressed genes. The method may be combined with empirical Bayes methods for moderating the genewise variances between genes. The method is validated using data from a microarray experiment involving calibration and ratio control spots in conjunction with spiked-in RNA. Comparing results for calibration and ratio control spots shows that the common correlation method results in substantially better discrimination of differentially expressed genes from those which are not. The spike-in experiment also confirms that the results may be further improved by empirical Bayes smoothing of the variances when the sample size is small. AVAILABILITY: The methodology is implemented in the limma software package for R, available from the CRAN repository http://www.r-project.org PMID- 15657103 TI - EPIMHC: a curated database of MHC-binding peptides for customized computational vaccinology. AB - SUMMARY: EPIMHC is a relational database of MHC-binding peptides and T cell epitopes that are observed in real proteins. Currently, the database contains 4867 distinct peptide sequences from various sources, including 84 tumor associated antigens. The EPIMHC database is accessible through a web server that has been designed to facilitate research in computational vaccinology. Importantly, peptides resulting from a query can be selected to derive specific motif-matrices. Subsequently, these motif-matrices can be used in combination with a dynamic algorithm for predicting MHC-binding peptides from user-provided protein queries. AVAILABILITY: The EPIMHC database server is hosted by the Dana Farber Cancer Institute at the site http://immunax.dfci.harvard.edu/bioinformatics/epimhc/ PMID- 15657104 TI - Co-occurrence based meta-analysis of scientific texts: retrieving biological relationships between genes. AB - MOTIVATION: The advent of high-throughput experiments in molecular biology creates a need for methods to efficiently extract and use information for large numbers of genes. Recently, the associative concept space (ACS) has been developed for the representation of information extracted from biomedical literature. The ACS is a Euclidean space in which thesaurus concepts are positioned and the distances between concepts indicates their relatedness. The ACS uses co-occurrence of concepts as a source of information. In this paper we evaluate how well the system can retrieve functionally related genes and we compare its performance with a simple gene co-occurrence method. RESULTS: To assess the performance of the ACS we composed a test set of five groups of functionally related genes. With the ACS good scores were obtained for four of the five groups. When compared to the gene co-occurrence method, the ACS is capable of revealing more functional biological relations and can achieve results with less literature available per gene. Hierarchical clustering was performed on the ACS output, as a potential aid to users, and was found to provide useful clusters. Our results suggest that the algorithm can be of value for researchers studying large numbers of genes. AVAILABILITY: The ACS program is available upon request from the authors. PMID- 15657105 TI - Testing association of a pathway with survival using gene expression data. AB - MOTIVATION: A recent surge of interest in survival as the primary clinical endpoint of microarray studies has called for an extension of the Global Test methodology to survival. RESULTS: We present a score test for association of the expression profile of one or more groups of genes with a (possibly censored) survival time. Groups of genes may be pathways, areas of the genome, clusters from a cluster analysis or all genes on a chip. The test allows one to test hypotheses about the influence of these groups of genes on survival directly, without the intermediary of single gene testing. The test is based on the Cox proportional hazards model and is calculated using martingale residuals. It is possible to adjust the test for the presence of covariates. We also present a diagnostic graph to assist in the interpretation of the test result, visualizing the influence of genes. The test is applied to a tumor dataset, revealing pathways from the gene ontology database that are associated with survival of patients. AVAILABILITY: The Global Test for survival has been incorporated into the R-package globaltest (version 3.0), available at http://www.bioconductor.org PMID- 15657106 TI - Prediction of unfolded segments in a protein sequence based on amino acid composition. AB - MOTIVATION: Partially and wholly unstructured proteins have now been identified in all kingdoms of life--more commonly in eukaryotic organisms. This intrinsic disorder is related to certain critical functions. Apart from their fundamental interest, unstructured regions in proteins may prevent crystallization. Therefore, the prediction of disordered regions is an important aspect for the understanding of protein function, but may also help to devise genetic constructs. RESULTS: In this paper we present a computational tool for the detection of unstructured regions in proteins based on two properties of unfolded fragments: (1) disordered regions have a biased composition and (2) they usually contain either small or no hydrophobic clusters. In order to quantify these two facts we first calculate the amino acid distributions in structured and unstructured regions. Using this distribution, we calculate for a given sequence fragment the probability to be part of either a structured or an unstructured region. For each amino acid, the distance to the nearest hydrophobic cluster is also computed. Using these three values along a protein sequence allows us to predict unstructured regions, with very simple rules. This method requires only the primary sequence, and no multiple alignment, which makes it an adequate method for orphan proteins. AVAILABILITY: http://genomics.eu.org/ PMID- 15657107 TI - Interacting genetic loci cause airway hyperresponsiveness. AB - Airway hyperresponsiveness (AHR) is a key physiological component of asthma, and the genetic basis of this complex trait has remained elusive. We created recombinant congenic mice with increased naive AHR by serially backcrossing A/J mice (which have elevated naive AHR) with C57BL/6J mice and selecting for mice with an elevated naive AHR phenotype. The seventh backcross-generation hyperresponsive mice retained A/J loci in three regions. Quantitative trait linkage (QTL) analysis of 123 unselected N8 progeny demonstrated that the AHR phenotype was not associated with any single locus but was significantly associated with an interaction of loci on chromosomes 2 and 6. These findings were confirmed in an independent analysis of chromosome substitution strain mice. The identification of genomic regions containing loci causally associated with AHR and the demonstration that this trait requires their interaction have important implications for the dissection of the genetic etiology of asthma in humans. PMID- 15657108 TI - Are radon gas measurements adequate for epidemiological studies and case control studies of radon-induced lung cancer? AB - The lung dose derived from radon is not attributed to the radon gas itself, but instead to its short-lived progeny. However, in many epidemiological studies as well as in case control studies of the radon risk, the excess number of cancers are related to the radon gas exposure, and not to the radon progeny exposure. A justification for such an approach has resorted to the assumption that there is self-compensation between the radiation doses from the unattached and attached fractions. In the present study, we used the Jacobi model to calculate the radon progeny concentrations in a room by varying the attachment rate and then calculated the resulting lung dose. It was found that self-compensation was not fully realised, and the effective dose can vary by a factor up to approximately 2 for the same radon gas concentration. In conclusion, the radon gas concentration alone does not provide adequate information on the effective dose. PMID- 15657109 TI - Radiation doses to patients from enteroclysis. AB - The purpose of this study was to determine the patient doses during enteroclysis and compare them with the available bibliographical data. For 14 enteroclysis examinations, the dose-area product (DAP) meter readings, fluoroscopy time, number of radiographs and exposure data were recorded. From these data, the fluoroscopy and radiography contributions to DAP, the entrance surface dose (ESD) and the effective dose (E) for each examination were estimated. The mean DAP was 81 Gy cm(2) and the mean fluoroscopy time was 19.5 min. The fluoroscopy contribution to DAP was 77% and 8.7 films were acquired in each examination on average. The mean ESD and E were estimated to be 428 mGy and 21 mSv, respectively. The mean DAP and fluoroscopy time calculated in this study are quite high when compared with those reported in the literature, suggesting that the examination technique should be reviewed and the ways to reduce patient exposure without compromising the diagnostic quality should be acquired. PMID- 15657110 TI - Radon concentration measurements in bituminous coal mines. AB - Radon measurements were carried out in Kozlu, Karadon and Uzulmez underground coal mines of Zonguldak bituminous coal basin in Turkey. Passive-time integrating method, which is the most widely used technique for the measurement of radon concentration in air, was applied by using nuclear etched track detectors (CR-39) in the study area. The radon concentration measurements were performed on a total of 42 points in those three mines. The annual exposure, the annual effective dose and lifetime fatality risk, which are the important parameters for the health of workers, were estimated based on chronic occupational exposure to the radon gas, which is calculated using UNCEAR-2000 and ICRP-65 models. The radon concentrations at several coal production faces are higher than the action level of 1000 Bq m(-3). It is suggested that the ventilation rates should be rearranged to reduce the radon concentration. PMID- 15657111 TI - Natural radioactivity in drinking water in private wells in Finland. AB - Natural radioactivity in drinking water was determined in population-based random study of 472 private wells. The mean concentrations of (222)Rn, (226)Ra, (234)U, (238)U, (210)Pb and (210)Po in drilled wells were 460, 0.05, 0.35, 0.26, 0.04 and 0.05 Bq l(-1), and in wells dug in the soil were 50, 0.016, 0.02, 0.015, 0.013 and 0.007 Bq l(-1), respectively. Approximately 10% of the drilled wells exceeded a radon concentration of 1000 Bq l(-1) and 18% a uranium concentration of 15 microg l(-1). The mean annual effective dose from natural radionuclides for a drilled well user was 0.4 mSv and 0.05 mSv for a user of a well dug in the soil. The effective dose arising from (222)Rn was 75% of the total of all natural radionuclides for drilled well users. As regards long-lived radionuclides, (210)Po and (210)Pb caused the largest portion of the effective dose. The dose arising from (238)U, (234)U and (226)Ra was only 8% of the total of all natural radionuclides. PMID- 15657112 TI - Three Mile Island epidemiologic radiation dose assessment revisited: 25 years after the accident. AB - Over the past 25 years, public health concerns following the Three Mile Island (TMI) accident prompted several epidemiologic investigations in the vicinity of TMI. One of these studies is ongoing. This commentary suggests that the major source of radiation exposure to the population has been ignored as a potential confounding factor or effect modifying factor in previous and ongoing TMI epidemiologic studies that explore whether or not TMI accidental plant radiation releases caused an increase in lung cancer in the community around TMI. The commentary also documents the observation that the counties around TMI have the highest regional radon potential in the United States and concludes that radon progeny exposure should be included as part of the overall radiation dose assessment in future studies of radiation-induced lung cancer resulting from the TMI accident. PMID- 15657113 TI - PTTL method applied to UV radiation detection during refractive surgery using excimer laser. AB - The method of phototransferred thermoluminescence (PTTL), using CaSO(4):Dy pellets produced at IPEN as sensitive material, was used to detect the spread laser radiation inside the surgery room during refractive surgical procedures using ArF excimer lasers. The purpose of this work was to study the viability of performing the ultraviolet radiation (UVR) exposure detection of patients and the hospital's surgical staff during a refractive surgery. The CaSO(4):Dy pellets were positioned at different distances from the laser source inside the surgery room: patient's ( congruent with 0.15 m), surgeon's ( congruent with 0.5 m) and nurse's ( congruent with 1.0 m) foreheads, lateral ( congruent with 1.5 m) and back ( congruent with 4.0 m) walls. The measurements of PTTL were carried out at two different conditions: five surgeries, each one taking approximately 10 min, and during a period of 4 h (cumulative), when several operations were performed. The detectors positioned as far as 4.0 m from the UV laser source were sensitised, making the UVR detection feasible at large source-detector distances. The absorbed energy was detected in the range from 40 microJ to 30 mJ during a surgery. This result indicates that the method studied can be used to detect the spread UVR. PMID- 15657114 TI - Experimental microdosimetric distributions of protons, from 19 to 65 MeV, using an A150-walled Low Pressure Proportional Counter. AB - Microdosimetric distributions of protons, from 19 to 65 MeV, were measured using an A150-walled Low Pressure Proportional Counter at the cyclotron facility in the National Institute of Radiological Sciences. The measured distributions were found to be very different from the spectra assumed by the Continuous Slowing Down Approximation. The measurements consisted of determining the energy deposition by direct incident protons, by secondary electrons produced in the wall of the detector and by scattered protons at the wall. The secondary electron and the scattered proton distributions were studied using analytical functions and the MCNPX Monte Carlo code, respectively. PMID- 15657115 TI - Cost-effectiveness of adherence-enhancing interventions: a quality assessment of the evidence. AB - OBJECTIVE: To determine whether the current cost-effectiveness evidence on adherence-enhancing interventions (AEIs) was of sufficient quality to aid in decision-making regarding medication adherence policies. DATA SOURCES: A computerized search of Embase, MEDLINE, Cinahl, Econlit, NHSEED, Psychlit, EPIC, and Cochrane databases (1980-April 2004) was performed. English-language human subject articles were identified using the key words compliance, adherence, concordance, patient assistance, therapeutic alliance, costs, economics, efficiency, resource use/utilization, cost-of-illness, cost-effectiveness, cost minimization, cost-utility, and cost-benefit. STUDY SELECTION AND DATA EXTRACTION: Studies that appeared to assess the cost-effectiveness of medication AEIs were included. Methodologic rigor was assessed using 15 minimum quality criteria. DATA SYNTHESIS: We found 45 comparative studies in 43 publications. Asthma (14 studies) and psychiatric illness (12 studies) were most commonly investigated. In 33 studies, interventions were educational, 18 had multiple components, and 23 did not appear to be linked to proven reasons for nonadherence. Reporting of adherence and outcome results was often unclear. Cost data were poorer quality than outcome data, using average or estimated costs and omitting some cost elements. Nine studies carried out incremental economic analysis. No study met all quality criteria. CONCLUSIONS: We were not able to make definitive conclusions about the cost-effectiveness of AEIs due to the heterogeneity of the studies found and incomplete reporting of results. Important policy decisions need to be made about nonadherence; however, they are currently being made in a vacuum of adequate information. AEIs must be based on reasons for nonadherence and be evaluated using accepted clinical and economic quality criteria. PMID- 15657116 TI - Potential for toxicity with use of bitter orange extract and guarana for weight loss. PMID- 15657117 TI - Insulin detemir--a new basal insulin analog. AB - OBJECTIVE: To review the pharmacology, pharmacokinetics, clinical trial data, adverse effects, and role in therapy of insulin detemir. DATA SOURCES: Articles and meeting abstracts were identified through searches of MEDLINE (1996-June 2004), EMBASE (1980-June 2004), and International Pharmaceutical Abstracts (1970 June 2004) databases, and unpublished information was provided by the manufacturer. STUDY SELECTION AND DATA EXTRACTION: All available studies relating to insulin detemir's pharmacology were selected. Only human studies were used for pharmacokinetic, drug interaction, efficacy, and safety data. DATA SYNTHESIS: Insulin detemir is a basal insulin analog that has been shown to improve glycemic control in patients with type 1 and type 2 diabetes. CONCLUSIONS: Insulin detemir offers some benefits over NPH for use as basal insulin in patients with type 1 and type 2 diabetes. PMID- 15657118 TI - The N-terminal to C-terminal motif in protein folding and function. AB - Essentially all proteins known to fold kinetically in a two-state manner have their N- and C-terminal secondary structural elements in contact, and the terminal elements often dock as part of the experimentally measurable initial folding step. Conversely, all N-C no-contact proteins studied so far fold by non two-state kinetics. By comparison, about half of the single domain proteins in the Protein Data Bank have their N- and C-terminal elements in contact, more than expected on a random probability basis but not nearly enough to account for the bias in protein folding. Possible reasons for this bias relate to the mechanisms for initial protein folding, native state stability, and final turnover. PMID- 15657120 TI - Pathogenic hantaviruses bind plexin-semaphorin-integrin domains present at the apex of inactive, bent alphavbeta3 integrin conformers. AB - The alphavbeta3 integrins are linked to human bleeding disorders, and pathogenic hantaviruses regulate the function of alphavbeta3 integrins and cause acute vascular diseases. alphavbeta3 integrins are present in either extended (active) or dramatically bent (inactive) structures, and interconversion of alphavbeta3 conformers dynamically regulates integrin functions. Here, we show that hantaviruses bind human alphavbeta3 integrins and that binding maps to the plexin semaphorin-integrin (PSI) domain present at the apex of inactive, bent, alphavbeta3-integrin structures. Pathogenic hantaviruses [New York-1 virus (NY 1V) and Hantaan virus (HTNV)] bind immobilized beta3 polypeptides containing the PSI domain, and human (but not murine) beta3 polypeptides inhibit hantavirus infectivity. Substitution of human beta3 residues 1-39 for murine beta3 residues directed pathogenic hantavirus infection of nonpermissive CHO cells expressing chimeric alphavbeta3 receptors. Mutation of murine beta3 Asn-39 to Asp-39 present in human beta3 homologues (N39D) permitted hantavirus infection of cells and specified PSI domain residue interactions with pathogenic hantaviruses. In addition, cell-surface expression of alphavbeta3 locked in an inactive bent conformation conferred hantavirus infectivity of CHO cells. Our findings indicate that hantaviruses bind to a unique domain exposed on inactive integrins and, together with prior findings, suggest that this interaction restricts alphavbeta3 functions that regulate vascular permeability. Our findings suggest mechanisms for viruses to direct hemorrhagic or vascular diseases and provide a distinct target for modulating alphavbeta3-integrin functions. PMID- 15657119 TI - Metabolizing enzyme toxicology assay chip (MetaChip) for high-throughput microscale toxicity analyses. AB - The clinical progression of new chemical entities to pharmaceuticals remains hindered by the relatively slow pace of technology development in toxicology and clinical safety evaluation, particularly in vitro approaches, that can be used in the preclinical and early clinical phases of drug development. To alleviate this bottle-neck, we have developed a metabolizing enzyme toxicology assay chip (MetaChip) that combines high-throughput P450 catalysis with cell-based screening on a microscale platform. The MetaChip concept is demonstrated by using sol-gel encapsulated P450s to activate the prodrug cyclophosphamide, which is the major constituent of the anticancer drug Cytoxan, as well as other compounds that are activated by P450 metabolism. The MetaChip provides a high-throughput microscale alternative to currently used in vitro methods for human metabolism and toxicology screening based on liver slices, cultured human hepatocytes, purified microsomal preparations, or isolated and purified P450s. This technology creates opportunities for rapid and inexpensive assessment of ADME/Tox (absorption, distribution, metabolism, excretion/toxicology) at very early phases of drug development, thereby enabling unsuitable candidates to be eliminated from consideration much earlier in the drug discovery process. PMID- 15657121 TI - Recent biological invasion may hasten invasional meltdown by accelerating historical introductions. AB - Biological invasions are rapidly producing planet-wide changes in biodiversity and ecosystem function. In coastal waters of the U.S., >500 invaders have become established, and new introductions continue at an increasing rate. Although most species have little impact on native communities, some initially benign introductions may occasionally turn into damaging invasions, although such introductions are rarely documented. Here, I demonstrate that a recently introduced crab has resulted in the rapid spread and increase of an introduced bivalve that had been rare in the system for nearly 50 yr. This increase has occurred through the positive indirect effects of predation by the introduced crab on native bivalves. I used field and laboratory experiments to show that the mechanism is size-specific predation interacting with the different reproductive life histories of the native (protandrous hermaphrodite) and the introduced (dioecious) bivalves. These results suggest that positive interactions among the hundreds of introduced species that are accumulating in coastal systems could result in the rapid transformation of previously benign introductions into aggressively expanding invasions. Even if future management efforts reduce the number of new introductions, given the large number of species already present, there is a high potential for positive interactions to produce many future management problems. Given that invasional meltdown is now being documented in natural systems, I suggest that coastal systems may be closer to this threshold than currently believed. PMID- 15657122 TI - Pseudomonas syringae manipulates systemic plant defenses against pathogens and herbivores. AB - Many pathogens are virulent because they specifically interfere with host defense responses and therefore can proliferate. Here, we report that virulent strains of the bacterial phytopathogen Pseudomonas syringae induce systemic susceptibility to secondary P. syringae infection in the host plant Arabidopsis thaliana. This systemic induced susceptibility (SIS) is in direct contrast to the well studied avirulence/R gene-dependent resistance response known as the hypersensitive response that elicits systemic acquired resistance. We show that P. syringae elicited SIS is caused by the production of coronatine (COR), a pathogen-derived functional and structural mimic of the phytohormone jasmonic acid (JA). These data suggest that SIS may be a consequence of the previously described mutually antagonistic interaction between the salicylic acid and JA signaling pathways. Virulent P. syringae also has the potential to induce net systemic susceptibility to herbivory by an insect (Trichoplusia ni, cabbage looper), but this susceptibility is not caused by COR. Rather, consistent with its role as a JA mimic, COR induces systemic resistance to T. ni. These data highlight the complexity of defense signaling interactions among plants, pathogens, and herbivores. PMID- 15657123 TI - Expression of a viral polymerase-bound host factor turns human cell lines permissive to a plant- and insect-infecting virus. AB - Tospoviruses are the only plant-infecting members of the Bunyaviridae family of ambisense ssRNA viruses. Tomato spotted wilt tospovirus (TSWV), the type-member, also causes mild infection on its main insect vector, Frankliniella occidentalis. Herein, we identified an F. occidentalis putative transcription factor (FoTF) that binds to the TSWV RNA-dependent RNA polymerase and to viral RNA. Using in vitro RNA synthesis assays, we show that addition of purified FoTF improves viral replication, but not transcription. Expression of FoTF deletion mutants, unable to bind the RNA-dependent RNA polymerase or viral RNA, blocks TSWV replication in F. occidentalis cells. Finally, expression of FoTF wild-type turns human cell lines permissive to TSWV replication. These data indicate that FoTF is a host factor required for TSWV replication in vitro and in vivo, provide an experimental system that could be used to compare molecular defense mechanisms in plant, insect, and human cells against the same pathogen (TSWV), and could lead to a better understanding of evolutionary processes of ambisense RNA viruses. PMID- 15657124 TI - A form of DISC1 enriched in nucleus: altered subcellular distribution in orbitofrontal cortex in psychosis and substance/alcohol abuse. AB - Disrupted-In-Schizophrenia 1 (DISC1) was identified as the sole gene whose ORF is truncated and cosegregates with major mental illnesses in a Scottish family. DISC1 has also been suggested, by association and linkage studies, to be a susceptibility gene for schizophrenia (SZ) in independent populations. However, no analysis of DISC1 protein in human brains, especially those of patients with SZ, has yet been conducted. Here we performed a biochemical analysis of DISC1 protein in a well characterized set of autopsied brains, including brains of patients with SZ, bipolar disorder, and major depression (MD), as well as normal control brains. We identified an isoform of DISC1 by using MS and demonstrated that it is enriched in the nucleus of HeLa cells. In the orbitofrontal cortex, the subcellular distribution of this DISC1 isoform, assessed by the nuclear to cytoplasmic ratio in the immunoreactivity of the isoform, is significantly changed in brains from patients with SZ and MD. This altered distribution is also observed in those subjects with substance and alcohol abuse. The changes in MD brains are significantly influenced by substance/alcohol abuse as well as postmortem interval; however, the alteration in SZ brains is free from brain associated confounding factors, although an interaction with substance/alcohol abuse cannot be completely ruled out. These results suggest that DISC1 may be implicated in psychiatric conditions in other populations than the unique Scottish family. PMID- 15657125 TI - Anticipating the three-dimensional consequences of eye movements. AB - Rapid eye movements called saccades give rise to sudden, enormous changes in optic information arriving at the eye; how the world nonetheless appears stable is known as the problem of spatial constancy. One consequence of saccades is that the directions of all visible points shift uniformly; directional or 2D constancy, the fact that we do not perceive this change, has received extensive study for over a century. The problems raised by 3D consequences of saccades, on the other hand, have been neglected. When the eye rotates in space, the 3D orientation of all stationary surfaces undergoes an equal-and-opposite rotation with respect to the eye. When presented with a an optic simulation of a saccade but with the eyes still, observers readily perceive this depth rotation of surfaces; when simultaneously performing the corresponding saccade, the 3D orientations of surfaces are perceived as stable, a phenomenon I propose calling 3D spatial constancy. In experiments presented here, observers viewed ambiguous 3D rotations immediately before, during, or after a saccade. The results show that before the eyes begin to move the brain anticipates the 3D consequences of saccades, preferring to perceive the rotation opposite to the impending eye movement. Further, the anticipation is absent when observers fixate while experiencing optically simulated saccades, and therefore must be evoked by extraretinal signals. Such anticipation could provide a mechanism for 3D spatial constancy and transsaccadic integration of depth information. PMID- 15657127 TI - Three-pronged genomic analysis reveals yeast cell-type regulation circuitry. PMID- 15657126 TI - Redefining implicit and explicit memory: the functional neuroanatomy of priming, remembering, and control of retrieval. AB - We used event-related functional MRI to study awareness of prior episodes during memory retrieval and its relationship to the intention to retrieve memories. Participants completed cues with words from a prior list (intentional test) or with the first words that came to mind (incidental test). During both tests, explicit memory was separated from priming in the absence of explicit memory. Priming was associated with hemodynamic decreases in left fusiform gyrus and bilateral frontal and occipital brain regions; explicit memory was associated with bilateral parietal and temporal and left frontal increases. Retrieval intention did not change these patterns but was associated with activity in right prefrontal cortex. Our results provide firm evidence that implicit and explicit memory have distinct functional neuroanatomies, and that strategic control of retrieval engages brain structures distinct from those involved in both implicit and explicit memory. They have critical implications for theories of memory and consciousness, which often equate consciousness with control. PMID- 15657128 TI - Mutation of the androgen receptor causes oncogenic transformation of the prostate. AB - Recent evidence demonstrates that the androgen receptor (AR) continues to influence prostate cancer growth despite medical therapies that reduce circulating androgen ligands to castrate levels and/or block ligand binding. Whereas the mutation, amplification, overexpression of AR, or cross-talk between AR and other growth factor pathways may explain the failure of androgen ablation therapies in some cases, there is little evidence supporting a causal role between AR and prostate cancer. In this study, we functionally and directly address the role whereby AR contributes to spontaneous cancer progression by generating transgenic mice expressing (i) AR-WT to recapitulate increased AR levels and ligand sensitivity, (ii) AR-T857A to represent a promiscuous AR ligand response, and (iii) AR-E231G to model altered AR function. Whereas transgenes encoding either AR-WT or AR-T857A did not cause prostate cancer when expressed at equivalent levels, expression of AR-E231G, which carries a mutation in the most highly conserved signature motif of the NH2-terminal domain that also influences interactions with cellular coregulators, caused rapid development of prostatic intraepithelial neoplasia that progressed to invasive and metastatic disease in 100% of mice examined. Taken together, our data now demonstrate the oncogenic potential of steroid receptors and implicate altered AR function and receptor coregulator interaction as critical determinants of prostate cancer initiation, invasion, and metastasis. PMID- 15657129 TI - Age-specific incidence of inherited versus sporadic cancers: a test of the multistage theory of carcinogenesis. AB - Knudson [Knudson, A. G. (1971) Proc. Natl. Acad. Sci. USA 68, 820-823] suggested that progression of retinoblastoma follows from two mutational events. Individuals who inherit one mutated gene copy should follow an age-onset pattern set by only a single rate-limiting step for transformation, whereas normal individuals should follow an age-onset pattern set by two rate-limiting events. Knudson's analysis of inherited and sporadic cases of retinoblastoma supported this prediction. However, retinoblastoma has a peculiar age-onset pattern concentrated in early life, because the retinal tissue completes most of its cell division by 5 years of age. Here, I compare age-specific incidences of inherited and sporadic forms of colon cancer, a much more typical form of human cancer. My simple mathematical analysis based on multistage theory explains the observed differences in age-onset patterns between inherited and sporadic cases. I also analyze recent retinoblastoma data and provide a mathematical analysis and interpretation. My analysis supports Knudson's two-hit theory but is much simpler and easier to understand than the original mathematical theory, which was based on a complicated model of cell division in the retina. My simpler theory for retinoblastoma makes clear the common basis for understanding multistage progression in tissues as different as the retina and colon. PMID- 15657130 TI - Development and evaluation of a microdevice for amino acid biomarker detection and analysis on Mars. AB - The Mars Organic Analyzer (MOA), a microfabricated capillary electrophoresis (CE) instrument for sensitive amino acid biomarker analysis, has been developed and evaluated. The microdevice consists of a four-wafer sandwich combining glass CE separation channels, microfabricated pneumatic membrane valves and pumps, and a nanoliter fluidic network. The portable MOA instrument integrates high voltage CE power supplies, pneumatic controls, and fluorescence detection optics necessary for field operation. The amino acid concentration sensitivities range from micromolar to 0.1 nM, corresponding to part-per-trillion sensitivity. The MOA was first used in the lab to analyze soil extracts from the Atacama Desert, Chile, detecting amino acids ranging from 10-600 parts per billion. Field tests of the MOA in the Panoche Valley, CA, successfully detected amino acids at 70 parts per trillion to 100 parts per billion in jarosite, a sulfate-rich mineral associated with liquid water that was recently detected on Mars. These results demonstrate the feasibility of using the MOA to perform sensitive in situ amino acid biomarker analysis on soil samples representative of a Mars-like environment. PMID- 15657131 TI - A fluorescent probe designed for studying protein conformational change. AB - The usefulness of fluorescence in studying protein motions derives from its sensitivity, kinetic resolution, and compatibility with both live cells and physiological assays. Recent advances in microscopy and membrane protein purification have permitted the observation of fluorescence changes that accompany the functional transitions of complex eukaryotic membrane proteins. These techniques rely on probes that can clearly report the environmental changes of specific residues, but most commonly available side-chain-reactive probes are not well suited for this purpose. Here, we introduce a red Cys-reactive probe, aminophenoxazone maleimide (APM), designed with improved chemical and spectral properties for reporting protein conformational change. APM is compact, uncharged, and has a short linker between probe and protein, all of which ensure that it can closely track the motions of the side chain to which it is attached. It undergoes large polarity-dependent changes in Stokes shift, as well as large bathochromic shifts in both excitation maximum (from 521 nm in toluene to 598 nm in water) and emission maximum (580 nm to 633 nm). These polarity-dependent spectral changes offer a potentially simple means of relating fluorescence to local structure and motion, although they are partially offset by some complicating factors in APM fluorescence. We find that, like a rhodamine maleimide, APM senses the conformational changes underlying voltage sensing in the Shaker potassium channel, and it is superior at a site that shows limited reactivity to the rhodamine. The spectral characteristics of APM can also report subtle differences between aqueous positions in purified preparations of the beta2 adrenergic receptor. PMID- 15657132 TI - Clonal expansion of hepatocytes during chronic woodchuck hepatitis virus infection. AB - Chronic hepadnavirus infections cause liver damage with ongoing death and regeneration of hepatocytes. In the present study we set out to quantify the extent of liver turnover by measuring the clonal proliferation of hepatocytes by using integrated viral DNA as a genetic marker for individual hepatocyte lineages. Liver tissue from woodchucks with chronic woodchuck hepatitis virus (WHV) infection was assayed for randomly integrated viral DNA by using inverse PCR. Serial endpoint dilution of viral-cell junction fragments into 96-well plates, followed by nested PCR and DNA sequencing, was used to determine the copy number of specific viral cell junctions as a measure of the clonal distribution of infected cell subpopulations. The results indicated that the livers contained a minimum of 100,000 clones of >1,000 cells containing integrated DNA, representing at least 0.2% of the hepatocyte population of the liver. Because cells with integrated WHV DNA comprised only 1-2% of total liver cells, it is likely that the total number of clones far exceeds this estimate, with as much as one-half of the liver derived from high copy clones of >1,000 cells. It may be inferred that these clones have a strong selective growth or survival advantage. The results provide evidence for a large amount of hepatocyte proliferation and selection having occurred during the period of chronic WHV infection ( approximately 1.5 years) in these animals. PMID- 15657133 TI - Loss of perivascular aquaporin 4 may underlie deficient water and K+ homeostasis in the human epileptogenic hippocampus. AB - An abnormal accumulation of extracellular K+ in the brain has been implicated in the generation of seizures in patients with mesial temporal lobe epilepsy (MTLE) and hippocampal sclerosis. Experimental studies have shown that clearance of extracellular K+ is compromised by removal of the perivascular pool of the water channel aquaporin 4 (AQP4), suggesting that an efficient clearance of K+ depends on a concomitant water flux through astrocyte membranes. Therefore, we hypothesized that loss of perivascular AQP4 might be involved in the pathogenesis of MTLE. Whereas Western blot analysis showed an overall increase in AQP4 levels in MTLE compared with non-MTLE hippocampi, quantitative ImmunoGold electron microscopy revealed that the density of AQP4 along the perivascular membrane domain of astrocytes was reduced by 44% in area CA1 of MTLE vs. non-MTLE hippocampi. There was no difference in the density of AQP4 on the astrocyte membrane facing the neuropil. Because anchoring of AQP4 to the perivascular astrocyte endfoot membrane depends on the dystrophin complex, the localization of the 71-kDa brain-specific isoform of dystrophin was assessed by immunohistochemistry. In non-MTLE hippocampus, dystrophin was preferentially localized near blood vessels. However, in the MTLE hippocampus, the perivascular dystrophin was absent in sclerotic areas, suggesting that the loss of perivascular AQP4 is secondary to a disruption of the dystrophin complex. We postulate that the loss of perivascular AQP4 in MTLE is likely to result in a perturbed flux of water through astrocytes leading to an impaired buffering of extracellular K+ and an increased propensity for seizures. PMID- 15657134 TI - alpha-lytic protease can exist in two separately stable conformations with different His57 mobilities and catalytic activities. AB - alpha-Lytic protease is a bacterial serine protease widely studied as a model system of enzyme catalysis. Here we report that lyophilization induces a structural change in the enzyme that is not reversed by redissolution in water. The structural change reduces the mobility of the active-site histidine residue and the catalytic activity of the enzyme. The application of mild pressure to solutions of the altered enzyme reverses the lyophilization-induced structural change and restores the mobility of the histidine residue and the enzyme's catalytic activity. This effect of lyophilization permits a unique opportunity for investigating the relationship between histidine ring dynamics and catalytic activity. The results demonstrate that His57 in resting enzymes is more mobile than previously thought, especially when protonated. The histidine motion and its correlation to enzyme activity lend support to the reaction-driven ring flip hypothesis. PMID- 15657135 TI - Statistical coevolution analysis and molecular dynamics: identification of amino acid pairs essential for catalysis. AB - Molecular dynamics (MD) simulations of HhaI DNA methyltransferase and statistical coupling analysis (SCA) data on the DNA cytosine methyltransferase family were combined to identify residues that are coupled by coevolution and motion. The highest ranking correlated pairs from the data matrix product (SCA.MD) are colocalized and form stabilizing interactions; the anticorrelated pairs are separated on average by 30 A and form a clear focal point centered near the active site. We suggest that these distal anti-correlated pairs are involved in mediating active-site compressions that may be important for catalysis. Mutants that disrupt the implicated interactions support the validity of our combined SCA.MD approach. PMID- 15657136 TI - Abelson-interactor-1 promotes WAVE2 membrane translocation and Abelson-mediated tyrosine phosphorylation required for WAVE2 activation. AB - WAVE2 is a member of the Wiskott-Aldrich syndrome protein family of cytoskeletal regulatory proteins shown to link Rac activation to actin remodeling via induction of Arp 2/3 activity. WAVE2 is thought to be regulated by its positioning in a macromolecular complex also containing the Abelson-(Abl) interactor-1 (Abi-1) adaptor, but the molecular basis and biologic relevance of WAVE2 inclusion in this complex are ill defined. Here we show that Abi-1 binding to WAVE2 is mediated by discrete motifs in the Abi-1 coiled-coil and WAVE2 WAVE homology domains and increases markedly in conjunction with Abi-1-WAVE2 translocation and colocalization at the leading edge in B16F1 cells after fibronectin stimulation. Abi-1 also couples WAVE2 to Abl after cell stimulation, an interaction that triggers Abl membrane translocation with WAVE2, Abi-1, and activated Rac, as well as Abl-mediated tyrosine phosphorylation and WAVE2 activation. By contrast, mutation of tyrosine residue Y150, identified here as the major site of Abl-mediated WAVE2 tyrosine phosphorylation, as well as disruption of WAVE2-Abi-1 binding, impairs induction of WAVE2-driven actin polymerization and its membrane translocation in association with activated Rac. Similarly, WAVE2 tyrosine phosphorylation and induction of membrane actin rearrangement are abrogated in fibroblasts lacking the Abl family kinase. Together, these data reveal that Abi-1-mediated coupling of Abl to WAVE2 promotes Abl-evoked WAVE2 tyrosine phosphorylation required to link WAVE2 with activated Rac and with actin polymerization and remodeling at the cell periphery. PMID- 15657138 TI - The economics of ideas and intellectual property. AB - Innovation and the adoption of new ideas is fundamental to economic progress. Here we examine the underlying economics of the market for ideas. From a positive perspective, we examine how such markets function with and without government intervention. From a normative perspective, we examine the pitfalls of existing institutions, and how they might be improved. We highlight recent research by us and others challenging the notion that government awards of monopoly through patents and copyright are "the way" to provide appropriate incentives for innovation. PMID- 15657137 TI - Gene delivery of human apolipoprotein E alters brain Abeta burden in a mouse model of Alzheimer's disease. AB - Apolipoprotein E (apoE) alleles are important genetic risk factors for Alzheimer's disease (AD), with the epsilon4 allele increasing and the epsilon2 allele decreasing risk for developing AD. ApoE has been shown to influence brain amyloid-beta peptide (Abeta) and amyloid burden, both in humans and in transgenic mice. Here we show that direct intracerebral administration of lentiviral vectors expressing the three common human apoE isoforms differentially alters hippocampal Abeta and amyloid burden in the PDAPP mouse model of AD. Expression of apoE4 in the absence of mouse apoE increases hippocampal Abeta(1-42) levels and amyloid burden. By contrast, expression of apoE2, even in the presence of mouse apoE, markedly reduces hippocampal Abeta burden. Our data demonstrate rapid apoE isoform-dependent effects on brain Abeta burden in a mouse model of AD. Gene delivery of apoE2 may prevent or reduce brain Abeta burden and the subsequent development of neuritic plaques. PMID- 15657139 TI - EsxA and EsxB are secreted by an ESAT-6-like system that is required for the pathogenesis of Staphylococcus aureus infections. AB - Mycobacterium tuberculosis secretes ESAT-6, a virulence factor that triggers cell mediated immune responses and IFN-gamma production during tuberculosis. ESAT-6 is transported across the bacterial envelope by a specialized secretion system with a FSD (FtsK-SpoIIIE domain) membrane protein. Although the presence of ESAT-6 like genes has been identified in the genomes of other microbes, the possibility that they may encode general virulence functions has hitherto not been addressed. Herein we show that the human pathogen Staphylococcus aureus secretes EsxA and EsxB, ESAT-6-like proteins, across the bacterial envelope. Staphylococcal esxA and esxB are clustered with six other genes and some of these are required for synthesis or secretion of EsxA and EsxB. Mutants that failed to secrete EsxA and EsxB displayed defects in the pathogenesis of S. aureus murine abscesses, suggesting that this specialized secretion system may be a general strategy of human bacterial pathogenesis. PMID- 15657140 TI - In vivo modulation of morphogenetic movements in Drosophila embryos with femtosecond laser pulses. AB - The complex biomechanical events associated with embryo development are investigated in vivo, by using femtosecond laser pulse-induced ablation combined with multimodal nonlinear microscopy. We demonstrate controlled intravital ablations preserving local cytoskeleton dynamics and resulting in the modulation of specific morphogenetic movements in nonmutant Drosophila embryos. A quantitative description of complex movements is obtained both in GFP-expressing systems by using whole-embryo two-photon microscopy and in unlabeled nontransgenic embryos by using third harmonic generation microscopy. This methodology provides insight into the issue of mechano-sensitive gene expression by revealing the correlation of in vivo tissue deformation patterns with Twist protein expression in stomodeal cells at gastrulation. PMID- 15657141 TI - Drosophila peptidoglycan recognition protein LC (PGRP-LC) acts as a signal transducing innate immune receptor. AB - Drosophila peptidoglycan recognition protein LC (PGRP-LC), a transmembrane protein required for the response to bacterial infection, acts at the top of a cytoplasmic signaling cascade that requires the death-domain protein Imd and an IkappaB kinase to activate Relish, an NF-kappaB family member. It is not clear how binding of peptidoglycan to the extracellular domain of PGRP-LC activates intracellular signaling because its cytoplasmic domain has no homology to characterized proteins. Here, we demonstrate that PGRP-LC binds Imd and that its cytoplasmic domain is critical for its activity, suggesting that PGRP-LC acts as a signal-transducing receptor. The PGRP-LC cytoplasmic domain is also essential for the formation of dimers, and results suggest that dimerization may be required for receptor activation. The PGRP-LC cytoplasmic domain can mediate formation of heterodimers between different PGRP-LC isoforms, thereby potentially expanding the diversity of ligands that can be recognized by the receptor. PMID- 15657142 TI - The Muller-Lyer illusion explained by the statistics of image-source relationships. AB - The Muller-Lyer effect, the apparent difference in the length of a line as the result of its adornment with arrowheads or arrow tails, is the best known and most controversial of the classical geometrical illusions. By sampling a range image database of natural scenes, we show that the perceptual effects elicited by the Muller-Lyer stimulus and its major variants are correctly predicted by the probability distributions of the possible physical sources underlying the relevant retinal images. These results support the conclusion that the Muller Lyer illusion is a manifestation of the probabilistic strategy of visual processing that has evolved to contend with the uncertain provenance of retinal stimuli. PMID- 15657143 TI - Natural-scene geometry predicts the perception of angles and line orientation. AB - Visual stimuli that entail the intersection of two or more straight lines elicit a variety of well known perceptual anomalies. Preeminent among these anomalies are the systematic overestimation of acute angles, the underestimation of obtuse angles, and the misperceptions of line orientation exemplified in the classical tilt, Zollner, and Hering illusions. Here we show that the probability distributions of the possible real-world sources of projected lines and angles derived from a range-image database of natural scenes accurately predict each of these perceptual peculiarities. These findings imply that the perception of angles and oriented lines is determined by the statistical relationship between geometrical stimuli and their physical sources in typical visual environments. PMID- 15657144 TI - Reconstitution of uridine-deletion precleaved RNA editing with two recombinant enzymes. AB - Uridine insertion/deletion RNA editing in trypanosomatid mitochondria is a posttranscriptional RNA modification phenomenon required for translation of mitochondrial mRNAs. This process involves guide RNA-mediated cleavage at a specific site, insertion or deletion of Us from the 3' end of the 5' mRNA fragment, and ligation of the two mRNA fragments. The Leishmania major RNA ligase containing complex protein 2 expressed in insect cells has a 3'-5' exoribonuclease activity and was therefore renamed RNA editing exonuclease 1 (REX1). Recombinant REX1 specifically trims 3' overhanging Us and stops at a duplex region. Evidence is presented that REX1 is responsible for deletion of the 3' overhanging Us from the bridged mRNA 5' cleavage fragment and that RNA editing ligase 1 is responsible for the ligation of the two mRNA cleavage fragments in U deletion editing. The evidence involves both in vivo down-regulation of REX1 expression in Trypanosoma brucei by RNA interference and the reconstitution of precleaved U-deletion in vitro editing with only two recombinant enzymes: recombinant REX1 and recombinant RNA editing ligase 1. PMID- 15657145 TI - Breaking sieve for steric exclusion of a noncognate amino acid from active site of a tRNA synthetase. AB - The genetic code is fixed in aminoacylation reactions catalyzed by aminoacyl-tRNA synthetases. Amino acid discrimination occurs at two sites: one for amino acid activation and aminoacylation and one for editing misactivated amino acids. Although the active site sieves out bulkier amino acids, misactivation occurs with substrates whose side chains are smaller than the cognate one. Paradoxically, although alanyl-tRNA synthetase activates glycine as well as alanine, the sterically larger (than alanine) serine is also misactivated. Here, we report crystal structures of an active fragment of Aquifex aeolicus alanyl tRNA synthetase complexed, separately, with Mg2+-ATP, alanine, glycine, and serine. Ala and Gly are bound in similar orientations in a side-chain accommodating pocket, where alpha-amino and carboxyl groups are stabilized by salt bridges, and the carboxyl by an H-bond from the side chain NH2 of Asn-194. In contrast, whereas the same two salt bridges stabilize bound Ser, H-bonding of the highly conserved (among class II tRNA synthetases) Asn-194 side chain NH2 to the Ser OH, instead of to the carboxyl, forces pocket expansion. Significantly, in the Mg2+-ATP complex, Asn-194 coordinates a Mg2+-alpha-phosphate bridge. Thus, the sieve for Ser exclusion is broken because of selective pressure to retain Asn 194 for Mg2+-ATP and Ala binding. PMID- 15657146 TI - A partial atomic structure for the flagellar hook of Salmonella typhimurium. AB - The axial proteins of the bacterial flagellum function as a drive shaft, universal joint, and propeller driven by the flagellar rotary motor; they also form the putative protein export channel. The N- and C-terminal sequences of the eight axial proteins were predicted to form interlocking alpha-domains generating an axial tube. We report on an approximately 1-nm resolution map of the hook from Salmonella typhimurium, which reveals such a tube made from interdigitated, 1-nm rod-like densities similar to those seen in maps of the filament. Atomic models for the two outer domains of the hook subunit were docked into the corresponding outermost features of the map. The N and C termini of the hook subunit fragment are positioned next to each other and face toward the axis of the hook. The placement of these termini would permit the residues missing in the fragment to form the rod-like features that form the core domain of the hook. We also fit the hook atomic model to an approximately 2-nm resolution map of the hook from Caulobacter crescentus. The hook protein sequence from C. crescentus is largely homologous to that of S. typhimurium except for a large insertion (20 kDa). According to difference maps and our fitting, this insertion is found on the outer surface of the hook, consistent with our modeling of the hook. PMID- 15657147 TI - Human maintenance DNA (cytosine-5)-methyltransferase and p53 modulate expression of p53-repressed promoters. AB - DNA (cytosine-5)-methyltransferase (DNMT) 1 participates in transcriptional repression of genes by methylation-dependent and -independent mechanisms. Here, DNMT1 is shown to bind p53 and colocalize in the nucleus. DNMT1-mediated methylation is stimulated by p53 in vitro. Upon p53 induction, a reporter construct containing the antiapoptotic gene survivin promoter, which contains a natural p53 binding site, was methylated in WT HCT116 cells but not in DNMT1 null or p53 null cells. Endogenous survivin gene repression involves cooperation between DNMT1 and p53 and is relieved by introduction of DNMT1- or p53-specific small inhibitory RNA. DNMT1 null cells did not exhibit a significant repressive effect for p53 responsive survivin and cdc25C gene expression compared with the parental cells. Normal human fibroblasts also exhibited similar DNMT1- and p53 mediated methylation of the survivin promoter, suggesting cooperation between p53 and DNMT1 in gene silencing. PMID- 15657148 TI - Mesoscopic self-organization of a self-assembled supramolecular rectangle on highly oriented pyrolytic graphite and Au(111) surfaces. AB - A self-assembled supramolecular metallacyclic rectangle was investigated with scanning tunneling microscopy on highly oriented pyrolytic graphite and Au(111) surfaces. The rectangles spontaneously adsorb on both surfaces and self-organize into well ordered adlayers. On highly oriented pyrolytic graphite, the long edge of the rectangle stands on the surface, forming a 2D molecular network. In contrast, the face of the rectangle lays flat on the Au(111) surface, forming linear chains. The structures and intramolecular features obtained through high resolution scanning tunneling microscopy imaging are discussed. PMID- 15657150 TI - Summaries for patients. What is the best way to test for colorectal cancer? PMID- 15657149 TI - Glutamate regulation of DARPP-32 phosphorylation in neostriatal neurons involves activation of multiple signaling cascades. AB - Dopamine- and cAMP-regulated phosphoprotein of 32 kDa (DARPP-32) plays a central role in medium spiny neurons in the neostriatum in the integration of various neurotransmitter signaling pathways. In its Thr-34-phosphorylated form, it acts as a potent protein phosphatase-1 inhibitor, and, in its Thr-75-phosphorylated form, it acts as a cAMP-dependent kinase inhibitor. Here, we investigated glutamate-dependent signaling cascades in mouse neostriatal slices by analyzing the phosphorylation of DARPP-32 at Thr-34 and Thr-75. Treatment with glutamate (5 mM) caused a complex change in DARPP-32 Thr-34 phosphorylation. An initial rapid increase in Thr-34 phosphorylation was NMDA/alpha-amino-3-hydroxy-5-methyl-4 isoxazolepropionic acid (AMPA)/metabotropic glutamate-5 receptor-dependent and was mediated through activation of a neuronal nitric oxide synthase/nitric oxide/cGMP/cGMP-dependent kinase signaling cascade. A subsequent decrease in phosphorylation was attributable to activation of an NMDA/AMPA receptor/Ca2+/protein phosphatase-2B signaling cascade. This decrease was followed by rephosphorylation via a pathway involving metabotropic glutamate-5 receptor/phospholipase C and extracellular receptor kinase signaling cascade. Treatment with glutamate initially decreased Thr-75 phosphorylation through activation of NMDA/AMPA receptor/Ca2+/protein phosphatase-2A signaling. Thereafter, glutamate slowly increased Thr-75 phosphorylation through activation of metabotropic glutamate-1 receptor/phospholipase C signaling. Our analysis of DARPP-32 phosphorylation in the neostriatum revealed that glutamate activates at least five different signaling cascades with different time dependencies, resulting in complex regulation of protein kinase and protein phosphatase activities. PMID- 15657151 TI - Summaries for patients. Use of colorectal cancer screening by doctors in practice. PMID- 15657152 TI - Summaries for patients. Targeting cholesterol treatment toward increasing high density lipoprotein levels. PMID- 15657153 TI - Summaries for patients. Long-term effects of antiviral treatment for hepatitis C. PMID- 15657154 TI - Summaries for patients. Wide range of drugs now available to effectively treat chronic heart failure. PMID- 15657155 TI - Accuracy of screening for fecal occult blood on a single stool sample obtained by digital rectal examination: a comparison with recommended sampling practice. AB - BACKGROUND: Many expert panels recommend colorectal cancer screening for average risk asymptomatic individuals older than 50 years of age. Recent studies have found that 24% to 64% of primary care providers use only the digital fecal occult blood test (FOBT) as their primary screening test. The effectiveness of a single digital FOBT is unknown. OBJECTIVE: To compare the sensitivity and specificity of digital FOBT and the recommended 6-sample at-home FOBT for advanced neoplasia in asymptomatic persons. DESIGN: Prospective cohort study. SETTING: 13 Veterans Affairs medical centers. PATIENTS: 3121 asymptomatic patients 50 to 75 years of age. INTERVENTION: 2665 patients had 6-sample at-home FOBT and digital FOBT, followed by complete colonoscopy. MEASUREMENTS: We measured the sensitivity of digital and 6-sample FOBT for advanced neoplasia and the specificity for no neoplasia. We calculated predictive values and likelihood ratios for advanced neoplasia, defined as tubular adenomas 10 mm or greater, adenomas with villous histology or high-grade dysplasia, or invasive cancer. RESULTS: Of all participants, 96.8% were men; their average age was 63.1 years. The 6-sample FOBT and the single digital FOBT had specificities of 93.9% and 97.5%, respectively, as defined by studying 1656 patients with no neoplasia. Sensitivities for detection of advanced neoplasia in 284 patients were 23.9% for the 6-sample FOBT and 4.9% for the digital FOBT. The likelihood ratio for advanced neoplasia was 1.68 (95% CI, 0.96 to 2.94) for positive results on digital FOBT and 0.98 (CI, 0.95 to 1.01) for negative results. LIMITATIONS: Most patients were men. CONCLUSIONS: Single digital FOBT is a poor screening method for colorectal neoplasia and cannot be recommended as the only test. When digital FOBT is performed as part of a primary care physical examination, negative results do not decrease the odds of advanced neoplasia. Persons with these results should be offered at-home 6-sample FOBT or another type of screening test. PMID- 15657156 TI - A national survey of primary care physicians' methods for screening for fecal occult blood. AB - BACKGROUND: Screening with the fecal occult blood test (FOBT) has been shown to reduce colorectal cancer incidence and mortality in randomized, controlled trials. Although the test is simple, implementation requires adherence to specific techniques of testing and follow-up of abnormal results. OBJECTIVE: To examine how FOBT and follow-up are conducted in community practice across the United States. DESIGN: Cross-sectional national surveys of primary care physicians and the public. SETTING: The Survey of Colorectal Cancer Screening Practices in Health Care Organizations and the 2000 National Health Interview Survey. PARTICIPANTS: 1147 primary care physicians who ordered or performed FOBT and 11 365 adults 50 years of age or older who responded to questions about FOBT use. MEASUREMENTS: Self-reported data on details of FOBT implementation and follow-up of positive results. RESULTS: Although screening guidelines recommend home tests, 32.5% (95% CI, 29.8% to 35.3%) of physicians used only the less accurate method of single-sample in-office testing; another 41.2% (CI, 38.3% to 44.0%) used both types of test. Follow-up of positive test results showed considerable nonadherence to guidelines, with 29.7% (CI, 27.1% to 32.4%) of physicians recommending repeating FOBT. Furthermore, sigmoidoscopy, rather than total colon examination, was commonly recommended to work up abnormal findings. Nearly one third of adults who reported having FOBT said they had only an in office test, and nearly one third of those who reported abnormal FOBT results reported no follow-up diagnostic procedures. LIMITATIONS: The study was based on self-reports. Data from the National Health Interview Survey may underestimate the prevalence of in-office testing and inadequate follow-up. CONCLUSIONS: Mortality reductions demonstrated with FOBT in clinical trials may not be realized in community practice because of the common use of in-office tests and inappropriate follow-up of positive results. Education of providers and system level interventions are needed to improve the quality of screening implementation. PMID- 15657157 TI - A randomized trial of a strategy for increasing high-density lipoprotein cholesterol levels: effects on progression of coronary heart disease and clinical events. AB - BACKGROUND: The high-density lipoprotein (HDL) cholesterol level is a strong predictor of cardiovascular events in epidemiologic studies. Until recently, it has been less extensively studied as a therapeutic target. OBJECTIVE: To assess the angiographic and clinical effects of a pharmacologic strategy to increase HDL cholesterol levels. DESIGN: Randomized, double-blind, placebo-controlled trial conducted from 1993 to 1996. SETTING: Outpatient specialty clinic of a large U.S. military medical center. PARTICIPANTS: 143 military retirees younger than 76 years of age with low HDL cholesterol levels and angiographically evident coronary disease. INTERVENTION: Gemfibrozil, niacin, and cholestyramine or corresponding placebos, with aggressive dietary and lifestyle intervention at baseline. MEASUREMENTS: Change from baseline to 30 months and a composite measure of clinical events that included hospitalization for angina, myocardial infarction, transient ischemic attack and stroke, death, and cardiovascular procedures. RESULTS: At baseline, mean (+/-SD) lipid values were as follows: total cholesterol, 5.1 +/- 0.8 mmol/L (196 +/- 31 mg/dL); low-density lipoprotein (LDL) cholesterol, 3.3 +/- 0.7 mmol/L (128 +/- 27 mg/dL); and HDL cholesterol, 0.9 +/- 0.2 mmol/L (34 +/- 6 mg/dL). Compared with placebo, the pharmacologically treated group experienced a 20% (95% CI, 14.8% to 24.3%) decrease in total cholesterol level, a 36% (CI, 28.4% to 43.5%) increase in HDL cholesterol level, a 26% (CI, 19.1% to 33.7%) decrease in LDL cholesterol level, and a 50% (CI, 40.5% to 59.2%) reduction in triglyceride levels. Focal coronary stenosis increased by 1.4% in the placebo group but decreased by 0.8% in the drug group (difference, -2.2 percentage points [CI, -4.2 to -0.1 percentage points]). A composite cardiovascular event end point was reached in 26% of patients in the placebo group and 13% of those in the drug group (difference, 13.7 percentage points [CI, 0.9 to 26.5 percentage points]). Side effects, particularly flushing and gastrointestinal intolerance, were more common in the drug group but rarely led to withdrawal from the study. LIMITATIONS: The study was small and used a composite clinical outcome. Whether improvements in angiographic findings were due to reductions in LDL cholesterol or increases in HDL cholesterol was not established. Flushing may have led to inadvertent unblinding in patients who were randomly assigned to active study drugs. CONCLUSIONS: A combination regimen aimed at increasing HDL cholesterol levels improves cholesterol profiles, helps prevent angiographic progression of coronary stenosis, and may prevent cardiovascular events in some people who exercise regularly and eat low-fat diets. PMID- 15657158 TI - Antiviral therapy for cirrhotic hepatitis C: association with reduced hepatocellular carcinoma development and improved survival. AB - BACKGROUND: Although cirrhosis is a major risk factor for development of hepatocellular carcinoma, no definitive prospective analyses have assessed the long-term efficacy of antiviral therapy in cirrhotic patients. OBJECTIVE: To elucidate the role of antiviral therapy in the suppression of liver tumors and survival over a long-term follow-up period. DESIGN: Prospective cohort study. SETTING: 25 clinical centers. PATIENTS: 345 patients with chronic hepatitis C and cirrhosis enrolled in previous trials. INTERVENTION: 271 patients received 6 to 9 million U of interferon 3 times weekly for 26 to 88 weeks; 74 received no treatment. MEASUREMENTS: Blood tests and abdominal ultrasonography were done regularly to detect hepatocellular carcinoma. RESULTS: Hepatocellular carcinoma was detected in 119 patients during a 6.8-year follow-up: 84 (31%) in the interferon-treated group and 35 (47%) in the untreated group. Cumulative incidence of hepatocellular carcinoma among interferon-treated patients was significantly lower than in untreated patients (Cox model: age-adjusted hazard ratio, 0.65 [95% CI, 0.43 to 0.97]; P = 0.03), especially sustained virologic responders. A total of 69 patients died during follow-up: 45 (17%) in the treated group and 24 (32%) in the untreated group. Interferon-treated patients had a better chance of survival than the untreated group (Cox model: age-adjusted hazard ratio, 0.54 [CI, 0.33 to 0.89]; P = 0.02). This was especially evident in sustained virologic responders. LIMITATION: This was not a randomized, controlled study. Patients enrolled in the control group had declined to receive interferon treatment even though they were eligible for treatment. CONCLUSION: Interferon therapy for cirrhotic patients with chronic hepatitis C, especially those in whom the infection had been cured, inhibited the development of hepatocellular carcinoma and improved survival. PMID- 15657159 TI - Improving patient care. The cognitive psychology of missed diagnoses. AB - Cognitive psychology is the science that examines how people reason, formulate judgments, and make decisions. This case involves a patient given a diagnosis of pharyngitis, whose ultimate diagnosis of osteomyelitis was missed through a series of cognitive shortcuts. These errors include the availability heuristic (in which people judge likelihood by how easily examples spring to mind), the anchoring heuristic (in which people stick with initial impressions), framing effects (in which people make different decisions depending on how information is presented), blind obedience (in which people stop thinking when confronted with authority), and premature closure (in which several alternatives are not pursued). Rather than trying to completely eliminate cognitive shortcuts (which often serve clinicians well), becoming aware of common errors might lead to sustained improvement in patient care. PMID- 15657160 TI - Improving patient care. My right knee. AB - Despite some impressive recent gains, improving the glaring deficiencies in health care quality is proving to be very hard. Improvement is local, rather than system-wide, and is sustained with difficulty, rather than becoming an intrinsic feature of care. My right knee will probably need to be replaced soon. This has given me the opportunity to define, in very personal terms, 5 specific dimensions of "total quality" that I will require from the medical institution that does my surgery and that every patient has the right to require of their encounters with the health care system. Don't kill me (no needless deaths). Do help me, and don't hurt me (no needless pain). Don't make me feel helpless. Don't keep me waiting. And don't waste resources, mine or anyone else's. Given my requirements, it is not clear that any health care institution in the United States will want to take me on as a patient. Although at this point individual institutions can meet some of these requirements, no single institution can deliver on all of them. Generating the energy, insight, and courage we need to get to "total quality" may require those of us who work in health care to get much better at seeing images of ourselves in the people we help. As Gandhi said, "You must be the change you wish to see in the world." PMID- 15657161 TI - Update in endocrinology. PMID- 15657162 TI - Narrative review: pharmacotherapy for chronic heart failure: evidence from recent clinical trials. AB - Heart failure is an important cause of morbidity and mortality. Clinical trials over the past 2 decades have revolutionized the care of patients with systolic heart failure, and substantial data support the use of angiotensin-converting enzyme inhibitors, beta-blockers, angiotensin-receptor blockers, and aldosterone blockers in the management of this serious condition. This article reviews the evidence on the pharmacologic treatment of heart failure, with a focus on recent clinical trials. PMID- 15657163 TI - Office-based testing for fecal occult blood: do only in case of emergency. PMID- 15657164 TI - Mainstream and alternative medicine: converging paths require common standards. PMID- 15657165 TI - A home visit. PMID- 15657166 TI - Racial and ethnic disparities in health care. PMID- 15657167 TI - Racial and ethnic disparities in health care. PMID- 15657168 TI - Limitations of virtual colonoscopy. PMID- 15657169 TI - Diabetes care in the Veterans Affairs system and in managed care. PMID- 15657170 TI - Prolonged coagulopathy related to superwarfarin overdose. PMID- 15657173 TI - Bioengineered lung epithelium: implications for basic and applied studies in lung tissue regeneration. PMID- 15657174 TI - Effects of cigarette smoke on pulmonary homeostasis. PMID- 15657175 TI - A common Fanconi anemia mutation in black populations of sub-Saharan Africa. AB - Fanconi anemia (FA) is a genetically heterogeneous chromosomal instability syndrome associated with multiple congenital abnormalities, aplastic anemia, and cancer. We report that a deletion mutation in the FANCG gene (c.637_643delTACCGCC) was present in 82% of FA patients in the black populations of Southern Africa. These patients originated from South Africa, Swaziland, Mozambique, and Malawi. The mutation was found on the same haplotype and was present in 1% of controls from the black South African population. These data indicate that the birth incidence of FA in this population is higher than 1 in 40 000, which is much higher than previously supposed, and suggest that the FANCG deletion is an ancient founder mutation in Bantu-speaking populations of sub Saharan Africa. Diagnostic screening is now possible by means of a simple DNA test. PMID- 15657176 TI - Cyclosporin A and tacrolimus, but not rapamycin, inhibit MHC-restricted antigen presentation pathways in dendritic cells. AB - The main targets for the immunosuppressive calcineurin inhibitors, cyclosporin A (CsA) and tacrolimus, have been considered to be activated T cells, but not antigen-presenting cells. Here we demonstrate that CsA and tacrolimus, but not rapamycin, inhibit major histocompatibility complex (MHC)-restricted antigen presentation in dendritic cells (DCs). Microencapsulated ovalbumin (OVA) was efficiently captured, processed, and presented on both class I MHC molecules (cross-presentation) as well as on class II MHC molecules. Addition of CsA and tacrolimus, but not rapamycin, to cultures of DCs inhibited both the class I processing pathway and the class II processing pathway of exogenous OVA. In addition, CsA and tacrolimus, but not rapamycin, also inhibited the classic class I processing pathway of endogenous OVA. CsA and tacrolimus did not inhibit presentation of exogenously added OVA peptide, SIINFEKL, phagocytic activity of DCs, or the total level of expression of class I MHC (H-2Kb) molecules. CsA and tacrolimus, however, inhibited profoundly the expression of SIINFEKL-H-2Kb complexes in OVA-phagocytized DCs. These results demonstrate clearly that CsA and tacrolimus inhibit intracellular processing events of antigens, and further suggest that the immunosuppressive activity of CsA and tacrolimus is at least in part due to inhibition of antigen processing pathways. PMID- 15657177 TI - Phosphorylation of RasGRP3 on threonine 133 provides a mechanistic link between PKC and Ras signaling systems in B cells. AB - B-cell receptor (BCR) signaling activates a number of intracellular signaling molecules including phospholipase C-gamma2 (PLC-gamma2), which generates membrane diacylglycerol (DAG). DAG recruits both protein kinase C (PKC) and RasGRP family members to the membrane and contributes to their activation. We have hypothesized that membrane colocalization facilitates activation of RasGRP3 by PKC. Here we demonstrate that PKC phosphorylates RasGRP3 on Thr133 in vitro, as determined by mass spectrometry. RasGRP3 with a Thr133Ala substitution is a poor PKC substrate in vitro and a poor Ras activator in vivo. Antiphosphopeptide antibodies recognize Thr133-phosphorylated RasGRP3 in B cells after BCR stimulation or DAG analog treatment, but much less so in resting cells. PKC inhibitors block RasGRP3 Thr133 phosphorylation and Ras-extracellular signal-related kinase (Erk) signaling with a similar pattern. After stimulation of T-cell receptor (TCR) or DAG analog treatment of T cells, PKC-catalyzed phosphorylation of RasGRP1 occurs on the homologous residue, Thr184. These studies shed light on the proposed "PKC Ras pathway" and support the hypothesis that RasGRP phosphorylation by PKC is a mechanism that integrates DAG signaling systems in T and B cells. PKC-mediated regulation of RasGRPs in lymphocytes may generate cooperative signaling in response to increases in DAG. The mast- and myeloid-selective family member RasGRP4 is regulated by different means. PMID- 15657178 TI - The effect of first-line imatinib interim therapy on the outcome of allogeneic stem cell transplantation in adults with newly diagnosed Philadelphia chromosome positive acute lymphoblastic leukemia. AB - Previously, we suggested that imatinib incorporation into conventional chemotherapy as an alternative (imatinib interim therapy) might be a useful strategy for bridging the time to allogeneic stem cell transplantation (SCT) for newly diagnosed Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph(+) ALL). Here, we provide an updated report on this strategy in 29 patients. At the time of enrollment, 23 patients (79.3%) achieved complete remission (CR). After the first imatinib cycle, the median breakpoint cluster region-Abelson oncogene locus (BCR-ABL)/ABL ratios decreased by 0.77 log in 25 (86.2%) responders, and their BCR-ABL/ABL ratios decreased further by 0.34 log after the second imatinib cycle, which included 7 molecular CR. One patient (4.3%) relapsed during the imatinib therapy. The remaining 3 patients were primarily refractory to both imatinib and chemotherapy. Twenty-five (86.2%) of the 29 patients received transplants in first CR. With a median follow-up duration of 25 months after SCT, the 3-year estimated probabilities of relapse, nonrelapse mortality, disease-free survival, and overall survival were 3.8%, 18.7%, 78.1%, and 78.1%, respectively. In comparison to our historical control data, first-line imatinib interim therapy appears to provide a good quality of CR and a survival advantage for patients with Ph(+) ALL. Further long-term follow-up is needed to validate the results of this study. PMID- 15657180 TI - Activity and specificity of toxin-related mouse T cell ecto-ADP ribosyltransferase ART2.2 depends on its association with lipid rafts. AB - Adenosine diphosphate (ADP)-ribosyl-transferases (ARTs) transfer ADP-ribose from nicotinamide adenine dinucleotide (NAD) onto target proteins. T cells express ART2.2, a toxin-related, glycosylphosphatidylinositol (GPI)-anchored ecto-enzyme. After the release of NAD from cells, ART2.2 ADP-ribosylates the P2X7 purinoceptor, lymphocyte function-associated antigen (LFA-1), and other membrane. Using lymphoma transfectants expressing either ART2.2 with its native GPI anchor (ART2.2-GPI) or ART2.2 with a grafted transmembrane anchor (ART2.2-Tm), we demonstrated that ART2.2-GPI but not ART2.2-Tm associated with glycosphingolipid enriched microdomains (lipid rafts). At limiting substrate concentrations, ART2.2 GPI exhibited more than 10-fold higher activity than ART2.2-Tm. On intact cells, ART2.2-GPI ADP-ribosylated a small number of distinct target proteins. Strikingly, the disruption of lipid rafts by cyclodextrin or membrane solubilization by Triton X-100 increased the spectrum of modified target proteins. However, ART2.2 itself was a prominent target for ADP-ribosylation only when GPI anchored. Furthermore, cholesterol depletion or detergent solubilization abolished the auto-ADP-ribosylation of ART2.2. These findings imply that ART2.2 GPI, but not ART2.2-Tm, molecules are closely associated on the plasma membrane and lend support to the hypothesis that lipid rafts exist on living cells as platforms to which certain proteins are admitted and others are excluded. Our results further suggest that raft association focuses ART2.2 on specific targets that constitutively or inducibly associate with lipid rafts. PMID- 15657179 TI - PD166326, a novel tyrosine kinase inhibitor, has greater antileukemic activity than imatinib mesylate in a murine model of chronic myeloid leukemia. AB - Imatinib mesylate is highly effective in newly diagnosed chronic myeloid leukemia (CML), but BCR/ABL (breakpoint cluster region/abelson murine leukemia)-positive progenitors persist in most patients with CML treated with imatinib mesylate, indicating the need for novel therapeutic approaches. In this study, we have used the murine CML-like myeloproliferative disorder as a platform to characterize the pharmacokinetic, signal transduction, and antileukemic properties of PD166326, one of the most potent members of the pyridopyrimidine class of protein tyrosine kinase inhibitors. In mice with the CML-like disease, PD166326 rapidly inhibited Bcr/Abl kinase activity after a single oral dose and demonstrated marked antileukemic activity in vivo. Seventy percent of PD166326-treated mice achieved a white blood cell (WBC) count less than 20.0 x 10(9)/L (20,000/microL) at necropsy, compared with only 8% of imatinib mesylate-treated animals. Further, two thirds of PD166326-treated animals had complete resolution of splenomegaly, compared with none of the imatinib mesylate-treated animals. Consistent with its more potent antileukemic effect in vivo, PD166326 was also superior to imatinib mesylate in inhibiting the constitutive tyrosine phosphorylation of numerous leukemia-cell proteins, including the src family member Lyn. PD166326 also prolonged the survival of mice with imatinib mesylate-resistant CML induced by the Bcr/Abl mutants P210/H396P and P210/M351T. Altogether, these findings demonstrate the potential of more potent Bcr/Abl inhibitors to provide more effective antileukemic activity. Clinical development of PD166326 or a related analog may lead to more effective drugs for the treatment of de novo and imatinib mesylate-resistant CML. PMID- 15657181 TI - A neurotrophin axis in myeloma: TrkB and BDNF promote tumor-cell survival. AB - Multiple myeloma (MM) is a B-cell neoplasm that is characterized by the clonal expansion of malignant plasma cells and is frequently associated with chromosomal translocations placing an oncogene under the control of the immunoglobulin heavy chain enhancer. Despite these pathogenic translocations, MM cells remain dependent on external cues for survival. We present evidence that brain-derived neurotrophic factor (BDNF), a member of the neurotrophin family of growth factors, and its high-affinity receptor, tropomyosin receptor kinase B (TrkB), contribute to these survival cues. MM cells express TrkB, and respond to BDNF by activating mitogen-activated protein kinase (MAPK) and phosphatidylinositol-3 kinase-a PI3K target (PI3K/Akt) signaling cascades. Addition of BDNF protects human MM cell lines (HMCLs) from apoptosis induced by dexamethasone or bortezomib and prolongs the survival of primary MM cells cultured alone or with human bone marrow (BM) stroma. As BDNF and TrkB are expressed by osteoblasts, stromal cells, and endothelial cells within the BM microenvironment, a BDNF-TrkB axis may be critical to the interactions of MM with bone and stroma that contribute to MM tumor progression. Finally, BDNF is expressed by malignant plasma cells isolated from a subset of patients with MM, as well as by most HMCLs, suggesting a potential role for this neurotrophin axis in autocrine as well as paracrine support of MM. PMID- 15657182 TI - Aberrant subcellular targeting of the G185R neutrophil elastase mutant associated with severe congenital neutropenia induces premature apoptosis of differentiating promyelocytes. AB - Mutations in the ELA2 gene encoding neutrophil elastase (NE) are present in most patients with severe congenital neutropenia (SCN). However, the mechanisms by which these mutations cause neutropenia remain unknown. To investigate the effects of mutant NE expression on granulopoiesis, we used the HL-60 promyelocytic cell line retrovirally transduced with the G185R NE mutant that is associated with a severe SCN phenotype. We show that the mutant enzyme accelerates apoptosis of differentiating but not of proliferating cells. Using metabolic labeling, confocal immunofluorescence microscopy, and immunoblot analysis of subcellular fractions, we also demonstrate that the G185R mutant is abnormally processed and localizes predominantly to the nuclear and plasma membranes rather than to the cytoplasmic compartment observed with the wild-type (WT) enzyme. Expression of the G185R mutant appeared to alter the subcellular distribution and expression of adaptor protein 3 (AP3), which traffics proteins from the trans-Golgi apparatus to the endosome. These observations provide further insight into potential mechanisms by which NE mutations cause neutropenia and suggest that abnormal protein trafficking and accelerated apoptosis of differentiating myeloid cells contribute to the severe SCN phenotype resulting from the G185R mutation. PMID- 15657183 TI - Ligands for natural killer cell-activating receptors are expressed upon the maturation of normal myelomonocytic cells but at low levels in acute myeloid leukemias. AB - Natural killer (NK) cell-mediated cytolytic activity against tumors requires the engagement of activating NK receptors by the tumor-associated ligands. Here, we have studied the role of NKG2D and natural cytotoxicity receptors (NCRs) in the recognition of human leukemia. To detect as-yet-unknown cell-surface molecules recognized by NCRs, we developed soluble forms of NKp30, NKp44, and NKp46 as staining reagents binding the putative cognate ligands. Analysis of UL16-binding protein-1 (ULBP1), ULBP2, and ULBP3 ligands for NKG2D and of potential ligands for NKp30, NKp44, and NKp46 in healthy hematopoietic cells demonstrated the ligand-negative phenotype of bone marrow-derived CD34(+) progenitor cells and the acquisition of cell-surface ligands during the course of myeloid differentiation. In acute myeloid leukemia (AML), leukemic blasts from approximately 80% of patients expressed very low levels of ULBPs and NCR-specific ligands. Treatment with differentiation-promoting myeloid growth factors, together with interferon gamma, upregulated cell-surface levels of ULBP1 and putative NCR ligands on AML blasts, conferring an increased sensitivity to NK cell-mediated lysis. We conclude that the ligand-negative/low phenotype in AML is a consequence of cell maturation arrest on malignant transformation and that defective expression of ligands for the activating NKG2D and NCR receptors may compromise leukemia recognition by NK cells. PMID- 15657185 TI - Identification of research priorities in occupational health. PMID- 15657184 TI - Uptake and presentation of hepatitis C virus-like particles by human dendritic cells. AB - Hepatitis C virus (HCV) is a major cause of chronic hepatitis worldwide. Interaction of dendritic cells (DCs) with viral particles may play an important role in the immunopathogenesis of HCV infection. Since the synthesis or purification of infectious virions is limited, we used HCV-like particles (HCV LPs) to study the interaction of HCV with human DCs. Immature DCs exhibited an envelope-specific and saturable binding of HCV-LPs, indicating receptor-mediated DC-HCV-LP interaction. Confocal microscopy revealed that HCV-LPs were rapidly taken up by DCs in a temperature-dependent manner. Competition experiments demonstrated that C-type lectins such as mannose receptor or DC-SIGN (DC-specific intercellular adhesion molecule 3-grabbing nonintegrin) were not sufficient for mediating HCV-LP binding. HCV-LP uptake was followed by DC activation. DCs pulsed with HCV-LPs stimulated HCV core-specific CD4(+) T cells, indicating that uptake of HCV-LPs by DCs leads to antigen processing and presentation on major histocompatibility complex (MHC) class II molecules. Finally, HCV-LP-derived antigens were efficiently cross-presented to HCV core-specific CD8(+) T cells. These findings demonstrate that HCV-LPs represent a novel model system to study HCV-DC interaction allowing definition of the molecular mechanisms of HCV uptake, DC activation, and antigen presentation to T cells. Furthermore, HCV-LP-mediated DC activation and efficient antigen presentation may explain the marked immunogenicity of HCV-LPs in vivo. PMID- 15657186 TI - Mental challenge in the workplace and risk of dementia in old age: is there a connection? PMID- 15657188 TI - Mortality of workers at a nickel carbonyl refinery, 1958-2000. AB - BACKGROUND: Excess risks of respiratory cancer have been shown in some groups of nickel exposed workers. It is clear, however, that not all forms of nickel exposure are implicated in these excess risks. AIM: To determine whether occupational exposures received in a modern nickel carbonyl refinery lead to increased risks of cancer, in particular nasal cancer and lung cancer. METHODS: The mortality experienced by a cohort of 812 workers employed at a nickel refinery was investigated. Study subjects were all male workforce employees first employed in the period 1953-92 who had at least five years' employment with the company. Observed numbers of cause specific deaths were compared with expectations based on national mortality rates; SMRs were also calculated by period from commencing employment, year of commencing employment, and type of work. RESULTS: Overall, standardised mortality ratios (SMRs) were close to 100 for all causes (Obs 191, SMR 96, 95% CI 83 to 111), all neoplasms (Obs 63, SMR 104, 95% CI 80 to 133), non-malignant diseases of the respiratory system (Obs 18, SMR 97, 95% CI 57 to 153), and diseases of the circulatory system (Obs 85, SMR 94, 95% CI 75 to 116). There were no significantly increased SMRs for any site of cancer. There was a non-significant excess for lung cancer (Obs 28, Exp 20.17, SMR 139, 95% CI 92 to 201), and in subgroup analyses a significantly increased SMR of 231 (Obs 9) was found for those 142 workers with at least five years' employment in the feed handling and nickel extraction departments. In the total cohort there was a single death from nasal cancer (Exp 0.10). CONCLUSIONS: The non-significant excess of lung cancer deaths may well be a chance finding, but in light of previous studies some role for nickel exposures cannot be excluded. PMID- 15657187 TI - A comparison of self-reported sickness absence with absences recorded in employers' registers: evidence from the Whitehall II study. AB - AIM: To compare self-reported sickness absence days in the last 12 months with recorded absences from the employers' registers for the same period. METHODS: Self-reported sickness absence data over the 12 months preceding baseline (1985 88) were compared with absence records from the employers' registers over the same period for 2406 women and 5589 men, participants in the Whitehall II study of British civil servants. Associations with self-rated health, longstanding illness, minor psychiatric disorder, physical illness, and prevalent coronary heart disease at baseline were determined. RESULTS: In general, women reported less sickness absence over the last year than was recorded in the employers' registers, while men, with the exception of those in the lower employment grades, reported more. Agreement between self-reported and recorded absence days decreased as the total number of days increased. After adjustment for employment grade and the average number of recorded and self-reported absence days, the total number of self-reported absence days was within two days of the recorded number of days for 63% of women and 67% of men. Associations between annual self reported sickness absence days and self-rated health, longstanding illness, minor psychiatric disorder, physical illness, and prevalent coronary heart disease were as strong as those for recorded absence days. CONCLUSION: These findings suggest that agreement between the annual number of self-reported and the annual number of recorded sickness absence days is relatively good in both sexes and that associations with health are equivalent for both measures. PMID- 15657189 TI - Health of children working in small urban industrial shops. AB - AIMS: To explore associations between work status and multidimensional health indices in a sample of urban Lebanese children. METHODS: A cross-sectional survey was used to compare 78 male children (aged 10-17 years) working full time in small industrial shops, and a comparison group of 60 non-working male schoolchildren. All children lived and worked or studied in the poor neighbourhoods of three main Lebanese cities. RESULTS: Working children reported frequent abuses. They smoked and dated more than the comparison group. They also reported a higher number of injuries (last 12 months) and recent skin, eye, and ear complaints (last two weeks). Physical examination revealed more changes in their skin and nails, but no differences in height or weight compared to non working group. A higher blood lead concentration was detected among working children, but no differences in haemoglobin and ferritin. No differences were noted between the two groups of children regarding anxiety, hopelessness, and self-esteem. The drawings of the working children, however, revealed a higher tendency to place themselves outside home and a wider deficit in developmental age when compared to non-working children. CONCLUSION: Significant differences were found between working and non-working children with respect to physical and social health parameters, but differences were less with regard to mental health. Future research should focus on (1) more sensitive and early predictors of health effects, and (2) long term health effects. The generality of findings to other work settings in the developing world should also be tested. PMID- 15657190 TI - Gender differences in the association of age with physical workload and functioning. AB - AIMS: To test whether (1) physically demanding work is less frequent for older than younger employees, and whether (2) the association of physically demanding work with decline of physical functioning is stronger for older employees than their younger counterparts. The gender differences in these associations were examined. METHODS: Subjects of the study were 40-60 year old employees of the City of Helsinki. Data (n = 5802) were collected with mail questionnaires in 2000 and 2001. Functioning was measured with the Role Limitations due to Physical Health Problems scale of the SF36 health questionnaire. Logistic regression models were used to analyse the data. RESULTS: There was a linear trend of less physically demanding work in older than in younger age groups. This trend was more marked for men than women. Age and physically demanding work were associated with poor functioning. In women the association of physically demanding work with poor functioning tended to be stronger for older than for younger age groups, while the opposite was observed in men. CONCLUSIONS: Results suggest that physically demanding work causes more ailments in women of high age than men. It is possible that less men than women are still employed in physically demanding occupations at high age, even though direct evidence of exit from physically demanding work cannot be obtained from cross-sectional data. In these data the physically demanding occupations for men and women were largely different. High physical workload among women working in social and health care is likely to contribute to the gender differences. PMID- 15657192 TI - The relationship between job satisfaction and health: a meta-analysis. AB - BACKGROUND: A vast number of published studies have suggested a link between job satisfaction levels and health. The sizes of the relationships reported vary widely. Narrative overviews of this relationship have been published, but no systematic meta-analysis review has been conducted. METHODS: A systematic review and meta-analysis of 485 studies with a combined sample size of 267 995 individuals was conducted, evaluating the research evidence linking self-report measures of job satisfaction to measures of physical and mental wellbeing. RESULTS: The overall correlation combined across all health measures was r = 0.312 (0.370 after Schmidt-Hunter adjustment). Job satisfaction was most strongly associated with mental/psychological problems; strongest relationships were found for burnout (corrected r = 0.478), self-esteem(r = 0.429), depression (r = 0.428), and anxiety(r = 0.420). The correlation with subjective physical illness was more modest (r = 0.287). CONCLUSIONS: Correlations in excess of 0.3 are rare in this context. The relationships found suggest that job satisfaction level is an important factor influencing the health of workers. Organisations should include the development of stress management policies to identify and eradicate work practices that cause most job dissatisfaction as part of any exercise aimed at improving employee health. Occupational health clinicians should consider counselling employees diagnosed as having psychological problems to critically evaluate their work-and help them to explore ways of gaining greater satisfaction from this important aspect of their life. PMID- 15657191 TI - Effect of different approaches to treatment of smoking as a potential confounder in a case-control study on occupational exposures. AB - AIM: To evaluate the effect of different approaches to treatment of smoking as a potential confounder in an occupational study of lung cancer. METHODS: Data were used from a case-control study on 956 men with lung cancer and 1253 population controls recruited in two northern Italian areas during 1990-1992. The risk of lung cancer associated with 11 selected job titles and eight selected industrial activities was estimated using seven different methods to treat smoking history. To evaluate the confounding effect of smoking, odds ratios obtained using the first six models were compared with estimates from the seventh and most complex model, in which cumulative tobacco consumption and time since cessation were considered. RESULTS: Although crude odds ratios for some of the occupational categories were biased by up to 25%, such bias decreased to less than 10% when a simple model including smoking status (never, ex-, current) was used. CONCLUSIONS: In occupational studies on lung cancer risk, information on smoking status may allow satisfactory control of the potential confounding effect of the habit. PMID- 15657193 TI - Insomnia is more common among subjects living in damp buildings. AB - BACKGROUND: Insomnia is a condition with a high prevalence and a great impact on quality of life. Little is known about the relation between and sleep disturbances and the home environment. AIM: To analyse the association between insomnia and building dampness. METHODS: In a cross-sectional, multicentre, population study, 16 190 subjects (mean age 40 years, 53% women) were studied from Reykjavik in Iceland, Bergen in Norway, Umea, Uppsala, and Goteborg in Sweden, Aarhus in Denmark, and Tartu in Estonia. Symptoms related to insomnia were assessed by questionnaire. RESULTS: Subjects living in houses with reported signs of building dampness (n = 2873) had a higher prevalence of insomnia (29.4 v 23.6%; crude odds ratio 1.35, 95% CI 1.23 to 1.48). The association between insomnia and different indicators of building dampness was strongest for floor dampness: "bubbles or discoloration on plastic floor covering or discoloration of parquet floor" (crude odds ratio 1.96, 95% CI 1.66 to 2.32). The associations remained significant after adjusting for possible confounders such as sex, age, smoking history, housing, body mass index, and respiratory diseases. There was no significant difference between the centres in the association between insomnia and building dampness. CONCLUSION: Insomnia is more common in subjects living in damp buildings. This indicates that avoiding dampness in building constructions and improving ventilation in homes may possibly have a positive effect on the quality of sleep. PMID- 15657194 TI - Patient satisfaction with occupational health physicians, development of a questionnaire. AB - AIMS: To develop a questionnaire that measures specific aspects of patient satisfaction with occupational health physicians. METHODS: General patient satisfaction questionnaires, a literature survey, and interviews with patients were used. An initial questionnaire was distributed among sick listed patients (n = 432) of occupational physicians (n = 90) from different occupational health services. To reduce items and to develop scales exploratory factor analysis and reliability analysis was used. A linear regression model was used to predict satisfaction ratings from the scales of the questionnaire. RESULTS: Questions about independence of the occupational physician were difficult to ask unambiguously. The factor analysis revealed five relevant factors which were named "being taken seriously as a patient", "attitude towards occupational health services", "trust and confidentiality", "expectations", and "comfort and access". All scales could be reduced to a maximum of five items without reducing the scale reliability too much. In the regression analysis, 71% of the variance of satisfaction ratings was explained by the first four scales and most by the first scale. "Comfort and access" did not contribute significantly to the model. CONCLUSIONS: A short questionnaire was developed to measure different aspects of patient satisfaction specific for occupational health. Whether the questionnaire can effectively lead to quality improvement in occupational health services should be investigated. PMID- 15657195 TI - Haloacetic acids in drinking water and risk for stillbirth. AB - BACKGROUND: Trihalomethanes (THMs) occurring in public drinking water sources have been investigated in several epidemiological studies of fetal death and results support a modest association. Other classes of disinfection by-products found in drinking water have not been investigated. AIMS: To investigate the effects of haloacetic acid (HAA) compounds in drinking water on stillbirth risk. METHODS: A population based case-control study was conducted in Nova Scotia and Eastern Ontario, Canada. Estimates of daily exposure to total and specific HAAs were based on household water samples and questionnaire information on water consumption at home and work. RESULTS: The analysis included 112 stillbirth cases and 398 live birth controls. In analysis without adjustment for total THM exposure, a relative risk greater than 2 was observed for an intermediate exposure category for total HAA and dichloroacetic acid measures. After adjustment for total THM exposure, the risk estimates for intermediate exposure categories were diminished, the relative risk associated with the highest category was in the direction of a protective effect, and all confidence intervals included the null value. CONCLUSIONS: No association was observed between HAA exposures and stillbirth risk after controlling for THM exposures. PMID- 15657196 TI - World at work: charcoal producing industries in northeastern Brazil. PMID- 15657197 TI - Are sickness absence frequencies in the study of EU countries underestimates? PMID- 15657199 TI - Retirement on grounds of ill health. PMID- 15657198 TI - Changing trends in US mesothelioma incidence. PMID- 15657202 TI - Hypoplastic coronary artery disease: report of one case. AB - Hypoplastic coronary artery disease (HCAD) is a rare abnormality with a high rate of sudden death and poor outcome. HCAD was revealed by myocardial infarction in a teenager with objective evidence of silent ischaemia on myocardial scintigraphy. After four years of follow up, he suddenly collapsed during exercise and subsequently died. Although HCAD is very uncommon, it should be actively excluded in children and young adults who experience sudden cardiac death. Aggressive treatment such as implantable cardioverter-defibrillator or heart transplantation may be indicated for this rare coronary abnormality. PMID- 15657201 TI - Treatment of unprotected left main coronary artery stenosis with a drug eluting stent in a heart transplant patient with allograft vasculopathy. AB - High risk angioplasty with drug eluting stent placement into an unprotected left main coronary artery in a heart transplant recipient with allograft vasculopathy is reported. Ten month angiographic follow up is reported. The literature is reviewed and current methods of revascularisation are described in detail. This is the first report of drug eluting stent use in this clinical situation. PMID- 15657203 TI - Contained myocardial rupture: a variant linking complete and incomplete rupture. AB - Myocardial rupture is an uncommon complication of myocardial infarction, often with devastating haemodynamic consequences. Although rupture is usually fatal, when patients do survive, the majority present with a pseudoaneurysm in which the rupture is sealed by a haematoma on the epicardial surface of the heart. Cases in which all myocardial layers are dissected except the epicardium or visceral pericardium have been included under this subheading. The authors describe such a case and suggest the pathological description of a "contained myocardial rupture". This link between complete and incomplete myocardial rupture may allow a more conservative management approach to be pursued. PMID- 15657200 TI - Staphylococcus lugdunensis infective endocarditis: description of 10 cases and analysis of native valve, prosthetic valve, and pacemaker lead endocarditis clinical profiles. AB - OBJECTIVE: To evaluate the incidence and the clinical and echocardiographic features of infective endocarditis (IE) caused by Staphylococcus lugdunensis and to identify the prognostic factors of surgery and mortality in this disease. DESIGN: Prospective cohort study. SETTING: Study at two centres (a tertiary care centre and a community hospital). PATIENTS: 10 patients with IE caused by S lugdunensis in 912 consecutive patients with IE between 1990 and 2003. METHODS: Prospective study of consecutive patients carried out by the multidisciplinary team for diagnosis and treatment of IE from the study institutions. English, French, and Spanish literature was searched by computer under the terms "endocarditis" and "Staphylococcus lugdunensis" published between 1989 and December 2003. MAIN OUTCOME MEASURES: Patient characteristics, echocardiographic findings, required surgery, and prognostic factors of mortality in left sided cases of IE. RESULTS: 10 cases of IE caused by S lugdunensis were identified at our institutions, representing 0.8% (four of 467), 1.5% (two of 135), and 7.8% (four of 51) of cases of native valve, prosthetic valve, and pacemaker lead endocarditis in the non-drug misusers. Native valve IE was present in four patients (two aortic, one mitral, and one pulmonary), prosthetic valve aortic IE in two patients, and pacemaker lead IE in the other four patients. All patients with left sided IE had serious complications (heart failure, periannular abscess formation, or shock) requiring surgery in 60% (three of five patients) of cases with an overall mortality rate of 80% (four of five patients). All patients with pacemaker IE underwent combined medical treatment and surgery, and mortality was 25% (one patient). In total 59 cases of IE caused by S lugdunensis were identified in a review of the literature. The combined analysis of these 69 cases showed that native valve IE (53 patients, 77%) is characterised by mitral valve involvement and frequent complications such as heart failure, abscess formation, and embolism. Surgery was needed in 51% of cases and mortality was 42%. Prosthetic valve endocarditis (nine of 60, 13%) predominated in the aortic position and was associated with abscess formation, required surgery, and high mortality (78%). Pacemaker lead IE (seven of 69, 10%) is associated with a better prognosis when antibiotic treatment is combined with surgery. CONCLUSIONS: S lugdunensis IE is an uncommon cause of IE, involving mainly native left sided valves, and it is characterised by an aggressive clinical course. Mortality in left sided native valve IE is high but the prognosis has improved in recent years. Surgery has improved survival in left sided IE and, therefore, early surgery should always be considered. Prosthetic valve S lugdunensis IE carries an ominous prognosis. PMID- 15657204 TI - Subacute coronary stent thrombosis in a patient developing clopidogrel associated thrombotic thrombocytopenic purpura. AB - Clopidogrel, in combination with aspirin, is commonly used for the prevention of thrombosis in patients who have received coronary artery stents. As a rare but critical complication, clopidogrel associated thrombotic thrombocytopenic purpura (TTP) has previously been described. A 78 year old man presented with unstable angina and filiform subtotal stenosis of the left anterior descending artery. He was treated with balloon angioplasty and stent implantation. After four days the patient again had angina caused by stent thrombosis, which was treated with balloon angioplasty. During hospital stay the typical course of clopidogrel associated TTP was observed with thrombocytopenia and petechial purpura occurring 14 days after drug initiation and prompt response to therapeutic plasma exchanges. These findings strongly suggest that clopidogrel may have increased platelet activation and aggregation in this immunologically susceptible patient, ultimately leading to a stent thrombosis. PMID- 15657205 TI - Delayed severe multivessel spasm and aborted sudden death after Taxus stent implantation. AB - Sudden cardiac arrest associated with major spasm of three coronary arteries was observed about 10 hours after Taxus stent insertion in a three vessel lesion and was successfully treated by intracoronary glyceryl trinitrate infusion. This case illustrates a potential risk associated with drug eluting stent and alerts clinicians to the life threatening risk of spasm when stenting multiple vessels with drug eluting stent (especially the Taxus stent). PMID- 15657206 TI - Oblique vein of Marshall. PMID- 15657207 TI - Large Eustachian valve and kyphoscoliosis. PMID- 15657208 TI - Contrast echocardiography during alcohol septal ablation: friend or foe? PMID- 15657210 TI - Posterior mitral valve leaflet prolapse diagnosed with multislice spiral computed tomography. PMID- 15657211 TI - Myocardial metabolic manipulation: a new therapeutic approach in heart failure? AB - Myocardial metabolic manipulation using drugs such as trimetazidine may offer a new therapeutic approach to the treatment of heart failure. PMID- 15657212 TI - Quantifying myocardial perfusion using contrast echocardiography. AB - There is a complex relation between what can be seen using perfusion imaging techniques, and what can be measured. PMID- 15657213 TI - Loffler's endomyocarditis. PMID- 15657215 TI - Ring in the heart. PMID- 15657214 TI - Idiopathic ventricular outflow tract tachycardia. AB - Although the pathogenesis of ventricular outflow tract tachycardia has not been fully elucidated, recent findings suggest that defects in cAMP signalling may be involved. PMID- 15657216 TI - The changing state of surgery for adult congenital heart disease. AB - The number of patients with adult congenital heart disease is expected to increase by 25% during this decade, so that by 2010 there will be more patients undergoing surgery for congenital heart disease over 16 years of age than under. PMID- 15657218 TI - Pulmonary artery dissection: an emerging cardiovascular complication in surviving patients with chronic pulmonary hypertension. AB - Pulmonary arterial dissection is an extremely rare and usually lethal complication of chronic pulmonary hypertension. The condition usually manifests as cardiogenic shock or sudden death and is therefore typically diagnosed at postmortem examination rather than during life. However, recent isolated reports have described pulmonary artery dissection in surviving patients. The first case of pulmonary artery dissection in a surviving patient with cor pulmonale caused by chronic obstructive pulmonary disease is presented. The aetiology, pathophysiology, and clinical presentation of pulmonary artery dissection are reviewed and factors that may aid diagnosis during life are discussed. PMID- 15657219 TI - An unusual case of ventricular tachycardia. PMID- 15657220 TI - Pre-discharge stress echocardiography and exercise ECG for risk stratification after uncomplicated acute myocardial infarction: results of the COSTAMI-II (cost of strategies after myocardial infarction) trial. AB - OBJECTIVE: To compare in a prospective, randomised, multicentre trial the relative merits of pre-discharge exercise ECG and early pharmacological stress echocardiography concerning risk stratification and costs of treating patients with uncomplicated acute myocardial infarction. DESIGN: 262 patients from six participating centres with a recent uncomplicated myocardial infarction were randomly assigned to early (day 3-5) pharmacological stress echocardiography (n = 132) or conventional pre-discharge (day 7-9) maximum symptom limited exercise ECG (n = 130). RESULTS: No complication occurred during either stress echocardiography or exercise ECG. At one year follow up there were 26 events (1 death, 5 non-fatal reinfarctions, 20 patients with unstable angina requiring hospitalisation) in patients randomly assigned to early stress echocardiography and 18 events (2 reinfarctions, 16 unstable angina requiring hospitalisation) in the group randomly assigned to exercise ECG (not significant). The negative predictive value was 92% for stress echocardiography and 88% for exercise ECG (not significant). Total costs of the two strategies were similar (not significant). CONCLUSION: Early pharmacological stress echocardiography and conventional pre-discharge symptom limited exercise ECG have similar clinical outcome and costs after uncomplicated infarction. Early pharmacological stress echocardiography should be considered a valid alternative even for patients with interpretable baseline ECG who can exercise. PMID- 15657221 TI - Strain rate imaging after dynamic stress provides objective evidence of persistent regional myocardial dysfunction in ischaemic myocardium: regional stunning identified? AB - OBJECTIVE: To investigate whether persistent ischaemic dysfunction of the myocardium after dynamic stress can be diagnosed from changes in ultrasonic strain rate and strain. DESIGN: Prospective observational study, with age matched controls. SETTING: University hospital. PATIENTS AND METHODS: 26 patients (23 men, mean (SD) age 58.9 (8.1) years) with coronary artery disease but no infarction and 12 controls (9 men, aged 56.1 (8.8) years) with normal coronary arteriography and negative exercise test underwent treadmill exercise (Bruce protocol). Tissue Doppler echocardiography was performed at baseline, at peak exercise, and at intervals up to one hour. Systolic and diastolic velocity, strain, and strain rate were recorded in the basal anterior segment of 16 patients with proximal left anterior descending coronary artery disease. RESULTS: Patients developed ischaemia, since they experienced angina, exercised for less time, and reached a lower workload than the control group, and had ST segment depression (-2.4 mm). Myocardial systolic velocity immediately after exercise increased by 31% and strain rate fell by 25% compared with increases of 92% and 62%, respectively, in the control group (p < 0.05). During recovery, myocardial systolic velocity and strain rate normalised quickly, whereas systolic strain remained depressed at 30 and 60 minutes after exercise, by 21% and 23%, respectively, compared with baseline (p < 0.05 versus controls). Myocardial diastolic velocities and strain rate normalised but early diastolic strain remained depressed by 32% compared with controls for 60 minutes (p < 0.05). Strain during atrial contraction was abnormal for 30 minutes. CONCLUSIONS: Myocardial strain shows regional post-ischaemic dysfunction in systole and diastole and may become a useful diagnostic tool in patients presenting with chest pain with a normal ECG. PMID- 15657224 TI - Traumatic pericardial effusion caused by a safety pin. PMID- 15657223 TI - Long term cardioprotective action of trimetazidine and potential effect on the inflammatory process in patients with ischaemic dilated cardiomyopathy. AB - OBJECTIVE: To investigate the long term effects of trimetazidine in patients with dilated ischaemic cardiomyopathy. The effects of trimetazidine on left ventricular function as well as its tolerability profile and potential anti inflammatory effects were studied. DESIGN: 61 patients were randomly assigned either to receive trimetazidine (20 mg thrice daily) in addition to their conventional treatment or to continue their usual drug treatment for 18 months. Patients were evaluated at baseline and at 6, 12, and 18 months with a clinical examination, echocardiography, and biochemical analysis (C reactive protein). RESULTS: Trimetazidine added to the usual treatment significantly improved the patients' functional status (assessed by New York Heart Association functional class). The functional improvement of trimetazidine treated patients was associated with a significant increase in left ventricular ejection fraction (30 (6)%, 32 (8)%, 38 (7)%, and 37 (6)% v 31 (8)%, 30 (7)%, 28 (6)%, and 26 (9)% in control patients at baseline and at 6, 12, and 18 months, respectively) and with a significant effect on ventricular remodelling. C reactive protein plasma concentrations remained stable throughout the study in patients receiving trimetazidine (2.5 (1.0), 2.7 (2.0), 2.7 (3.0), and 3.0 (2.0) mg/l at baseline and at 6, 12, and 18 months, respectively) but increased significantly in the control group (2.4 (1.0), 3.4 (1.2), 6.0 (4.0), and 7.0 (5.0) mg/l, respectively). No significant adverse event or changes in clinical or biochemical parameters were detected. CONCLUSION: Treatment with trimetazidine added to the usual treatment for up to 18 months was well tolerated and induced a functional improvement in patients with dilated cardiomyopathy. Trimetazidine treatment was associated with a significant improvement of left ventricular function and the remodelling process. Results also suggest that the inflammatory response was limited in patients treated with trimetazidine. PMID- 15657226 TI - Evaluation of a hand carried cardiac ultrasound device in an outpatient cardiology clinic. AB - OBJECTIVE: To determine the diagnostic potential of a hand carried cardiac ultrasound (HCU) device (OptiGo, Philips Medical Systems) in a cardiology outpatient clinic and to compare the HCU diagnosis with the clinical diagnosis and diagnosis with a full featured standard echocardiography (SE) system. METHODS: 300 consecutive patients took part in the study. The HCU examination was performed by an experienced echocardiographer before patients visited the cardiologist. The echocardiographer noted whether the HCU device was able to confirm or reject the referral diagnosis, which abnormality was detected, and whether SE investigation was necessary. Physical examination by a cardiologist followed and thereafter, whenever required, a complete study with an SE was carried out. The HCU data were compared with the clinical diagnosis of the cardiologist and the SE diagnosis in a blinded manner. RESULTS: The cardiologist referred 203 of 300 patients for an SE study and 13 patients for transoesophageal echocardiography. In 84 patients no further examination was considered necessary. HCU echocardiography was able to confirm or reject the suspected clinical diagnosis in 159 of 203 (78%) patients. In 44 of 203 (22%) patients SE Doppler was needed. Agreement between the HCU device and the SE system for the detection of major abnormalities was excellent (98%). The HCU device missed 4% of the major findings. Among the 84 patients not referred for an SE, the HCU device detected unsuspected major abnormalities missed with the physical examination in 14 (17%). CONCLUSION: Integration of an HCU device with the physical examination augments the yield of information. PMID- 15657225 TI - Expression of connexins 40 and 43 in human left atrium in atrial fibrillation of different aetiologies. AB - OBJECTIVE: To test the hypothesis that atrial fibrillation (AF) is associated with changes in the expression of connexins 40 and 43 in the left atrium with more pronounced changes in mitral valve disease than in lone AF. METHODS: Protein concentrations of connexin 40 and connexin 43 were analysed in left atrial tissue of patients undergoing cardiac surgery. One group of patients had lone AF (n = 41), one group had AF and mitral valve repair (n = 36), and one group in sinus rhythm served as controls (n = 15). RESULTS: Western blot analysis of connexin 40 and connexin 43 expression showed an increase of both gap junctional proteins (connexin 43 > connexin 40) in patients with AF of all forms compared with patients in sinus rhythm (p = 0.01 and p = 0.011, respectively). Subgroup analysis showed increased concentrations of connexin 40 in lone AF and AF with mitral valve disease compared with sinus rhythm (p = 0.06 and p = 0.029, respectively), whereas the same analysis for connexin 43 reached significance only in the mitral valve disease group (p = 0.031). No differences in connexin 40 and connexin 43 expression were detectable between lone AF and AF with mitral valve disease. Within the groups connexin 40 and connexin 43 expression did not differ between patients with paroxysmal AF and patients with chronic AF. CONCLUSION: The present study shows for the first time that AF can induce changes in the left atrium with increased connexin expression. Furthermore, no systematic differences between patients with paroxysmal and chronic AF were detected. PMID- 15657227 TI - Dilated coronary arterial lesions in the late period after Kawasaki disease. AB - OBJECTIVES: There are two types of late coronary dilated lesions after Kawasaki disease: new aneurysms and expanding aneurysms. The development of coronary dilated lesions late after Kawasaki disease was investigated. METHODS: Between 1978 and 2003, 562 patients with coronary arterial lesions underwent selective coronary angiography on at least two occasions. RESULTS: Of the 562 patients studied, 17 new dilated or expanding lesions were found in 15 patients (3%, 11 boys, four girls). The time of detection of new aneurysms after Kawasaki disease ranged from 1.9-19.2 years (median 11.4 years) and their diameters ranged from 2.0-6.5 mm (median 4.4 mm). Thirteen new aneurysms occurred in vessels in which previous aneurysms had regressed and all new aneurysms were associated with localised stenosis. A new aneurysm at the bifurcation or in the branches was seen in 14 (93%) and 13 were eccentric (87%). Of two expanding aneurysms, one involved the right coronary artery in one patient and the other the left anterior descending coronary artery. One expanding aneurysm increased from 4.4 mm to 19.5 mm over 17 years, and the other expanding aneurysm increased from 10 mm to 15 mm in one year. CONCLUSIONS: Neither new nor expanding aneurysms have caused cardiac events. New aneurysms often develop as a pre-stenotic or post-stenotic dilatation secondary to localised stenosis. New and expanding aneurysms may be caused by haemodynamic factors in addition to the abnormality of the coronary arterial wall after severe acute vasculitis. Coronary arterial wall abnormalities were stenosis as well as, rarely, dilatation of the vessels in the late period. It is important to recognise that the changes of the coronary arterial wall persist late after regression of a large aneurysm. PMID- 15657229 TI - A rare cause of ST segment elevation. PMID- 15657228 TI - Novel quantitative assessment of myocardial perfusion by harmonic power Doppler imaging during myocardial contrast echocardiography. AB - OBJECTIVE: To test the hypothesis that the power of the received signal of harmonic power Doppler imaging (HPDI) is proportional to the bubble concentration under conditions of constant applied acoustic pressure, and to determine whether a new quantitative method can overcome the acoustic field inhomogeneity during myocardial contrast echocardiography (MCE) and identify perfusion abnormalities caused by myocardial infarction. METHODS: The relation between Levovist concentration and contrast signal intensity (CI) of HPDI was investigated in vitro under conditions of constant acoustic pressure. MCE was performed during continuous infusion of Levovist with intermittent HPDI every sixth cardiac cycle in 11 healthy subjects and 25 patients with previous myocardial infarction. In the apical views myocardial CI (CI(myo)) was quantified in five myocardial segments. The CI from the left ventricular blood pool adjacent to the segment was also measured in dB and subtracted from the CI(myo) (relative CI (RelCI)). RESULTS: CI had a logarithmic correlation and the calculated signal power a strong linear correlation with Levovist concentration in vitro. Thus, a difference in CI of X dB indicates a microbubble concentration ratio of 10(X/10). In normal control subjects, CI(myo) differed between the five segments (p < 0.0001), with a lower CI(myo) in deeper segments. However, RelCI did not differ significantly between segments (p = 0.083). RelCI was lower (p < 0.0001) in the 39 infarct segments (mean (SD) -18.6 (2.8) dB) than in the 55 normal segments (mean (SD) -15.1 (1.6) dB). RelCI differed more than CI(myo) between groups. CONCLUSIONS: The new quantitative method described can overcome the acoustic field inhomogeneity in evaluation of myocardial perfusion during MCE. RelCI represents the ratio of myocardium to blood microbubble concentrations and may correctly reflect myocardial blood volume fraction. PMID- 15657230 TI - Sinus node revisited in the era of electroanatomical mapping and catheter ablation. AB - OBJECTIVE: To study the architecture of the human sinus node to facilitate understanding of mapping and ablative procedures in its vicinity. METHODS: The sinoatrial region was examined in 47 randomly selected adult human hearts by histological analysis and scanning electron microscopy. RESULTS: The sinus node, crescent-like in shape, and 13.5 (2.5) mm long, was not insulated by a sheath of fibrous tissue. Its margins were irregular, with multiple radiations interdigitating with ordinary atrial myocardium. The distances from the node to endocardium and epicardium were variable. In 72% of the hearts, the whole nodal body was subepicardial and in 13 specimens (28%) the inner aspect of the nodal body was subendocardial. The nodal body cranial to the sinus nodal artery was more subendocardial than the remaining nodal portion, which was separated from the endocardium by the terminal crest. In 50% of hearts, the most caudal boundaries of the body of the node were at least 3.5 mm from the endocardium. When the terminal crest was > 7 mm thick (13 hearts, 28%), the tail was subepicardial or intramyocardial and at least 3 mm from the endocardium. CONCLUSIONS: The length of the node, the absence of an insulating sheath, the presence of nodal radiations, and caudal fragments offer a potential for multiple breakthroughs of the nodal wavefront. The very extensive location of the nodal tissue, the cooling effect of the nodal artery, and the interposing thick terminal crest caudal to this artery have implications for nodal ablation or modification with endocardial catheter techniques. PMID- 15657231 TI - Early and late outcome of skeletonised bilateral internal mammary arteries anastomosed to the left coronary system. AB - OBJECTIVES: To investigate in a retrospective study the technical aspects of using the in situ bilateral internal mammary arteries (IMAs), with the right IMA (RIMA) used for revascularisation of the circumflex system, and to evaluate early and late outcome. MATERIALS AND METHODS: Between January 1997 and July 2003, 552 consecutive patients underwent grafting of the circumflex artery system with an in situ skeletonised RIMA routed through the transverse sinus (eventually retrocaval). Mean (SD) age was 63.8 (11) years. 331 (60%) patients underwent total arterial myocardial revascularisation. Mean follow up was 26 (9) months. RESULTS: The success rate of skeletonised RIMA grafting to the circumflex branch was 100%. There were 19 (3.4%) in-hospital deaths. Perioperative myocardial infarction occurred in 12 (2.2%) patients. In 155 patients undergoing postoperative angiography, two had an occluded RIMA and a string-like phenomenon was seen in three RIMA and one left IMA (LIMA). Three RIMA and three LIMA had stenotic lesions. The patency rates of RIMA and LIMA were 94% and 97.4%, respectively. Strong predictors of non-functional IMA grafts were a recipient coronary artery diameter of < 1.5 mm (p = 0.022), < 60% stenosis of the recipient coronary artery (p = 0.015), diffuse stenotic lesions of the recipient coronary artery (p = 0.018), and a small IMA calibre (p = 0.0001). Cumulative actuarial survival at three years was 96.4% and event-free cumulative survival was 93.8%. CONCLUSIONS: Use of the bilateral IMAs offers the possibility of constructing various configurations, making total arterial myocardial revascularisation possible with a minimum number of arterial conduits. Use of the skeletonised RIMA through the transverse sinus and eventually retrocavally can reach most branches of the circumflex system and is associated with an excellent patency rate. Patients who received bilateral IMA grafts for left coronary system revascularisation had improved early and late outcomes and decreased risk of death, reoperation, and angioplasty. PMID- 15657232 TI - Coronary artery spasm in lymphocytic myocarditis: a rare cause of acute myocardial infarction. PMID- 15657233 TI - Fractional flow reserve for the prediction of cardiac events after coronary stent implantation: results of a multivariate analysis. AB - OBJECTIVE: To determine the prognostic value of fractional flow reserve (FFR) measurements after coronary stent implantation including multiple clinical and angiographic parameters collected in one centre. METHODS: 119 consecutive patients were enrolled who had a stent implanted with the use of a pressure wire as a guidewire. Patients were followed up for at least six months. Any death, myocardial infarction, and target vessel revascularisation were considered major adverse cardiac events (MACE). Multivariate logistic regression was used to determine adjusted odds ratios (OR) and 95% confidence intervals (CI) for FFR and covariates. RESULTS: Complete follow up data were available for all 119 patients. Pre-interventional FFR increased from 0.65 (0.15) to 0.94 (0.06) (p < 0.0001) after stent implantation. Eighteen MACE (15%) occurred during follow up including 15 (12.6%) target vessel revascularisations. Final FFR was significantly higher in patients without than in patients with an event (0.95 (0.05) v 0.88 (0.08), p = 0.001). In the multivariate logistic regression analysis, only final FFR < 0.95 (OR 6.22, 95% CI 1.79 to 21.62, p = 0.004) and reduced left ventricular function (OR 0.95, 95% CI 092 to 0.99, p = 0.021) remained as significant independent predictors for MACE. CONCLUSION: These results including multiple parameters underline that FFR after coronary stenting is a strong and independent predictor for subsequent cardiac events after six months' follow up. PMID- 15657235 TI - Pulmonary artery obstruction mimicking the clinical features of acute pulmonary embolism. PMID- 15657234 TI - Changing practice of cardiac surgery in adult patients with congenital heart disease. AB - OBJECTIVES: To review 13 years' data from a unit for grown ups with congenital heart disease (GUCH) to understand the change in surgical practice. METHODS: Records were reviewed of patients over 16 years of age undergoing surgery between 1 January 1990 and 31 December 2002 in a dedicated GUCH unit. Patients with atrial septal defects were included but not those with Marfan's syndrome or undergoing a first procedure for bicuspid aortic valves. Three equal time periods of 52 months were analysed. RESULTS: Of 474 operations performed, 162 (34.2%) were repeat operations. The percentage of repeat operations increased from 24.8% (41 of 165) in January 1990-April 1994 to 49.7% (74 of 149) in September 1998 December 2002. Mortality was 6.3% (n = 30). The median age decreased from 25.4 years (interquartile range 18.7) in January 1990-April 1994 to 23.9 (interquartile range 17.3) in September 1998-December 2002 (p = 0.04). The proportion of patients with a "simple" diagnosis decreased from 45.4% (74 or 165) in January 1990-April 1994 to 27.5% (41 of 149) in September 1998-December 2002 (p = 0.013). Pulmonary valve replacements in operated tetralogy of Fallot increased from one case in January 1990-April 1994 to 23 cases in September 1998 December 2002 and conduit replacement increased from five cases to 17. However, secundum atrial septal defect closures decreased from 35 cases to 14 (p < 0.0001). The estimated cost (not including salaries and prosthetics) incurred by an adult patient with congenital heart disease was pound2290 compared with pound2641 for a patient undergoing coronary artery bypass grafting. CONCLUSION: Despite the impact of interventional cardiology, the total number of surgical procedures remained unchanged. The complexity of the cases increased particularly with repeat surgery. Nevertheless, the patients do well with low mortality and the inpatient costs remain comparable with costs of surgery for acquired disease. PMID- 15657236 TI - Health related quality of life in adults with repaired tetralogy of Fallot: psychosocial and cognitive outcomes. AB - OBJECTIVE: To assess the health related quality of life of patients who were operated on during childhood for total correction of tetralogy of Fallot, focusing on the psychosocial and cognitive outcomes. PATIENTS: 54 patients (24 men and 30 women, mean (SD) age 32 (4) years), operated on for total correction of tetralogy of Fallot at mean age of 8.2 years, underwent a cardiological examination, psychological assessment (semistructured interview, Minnesota multiphasic personality inventory), evaluation of quality of life (36 item short form health survey), and neuropsychological assessment with an extensive neuropsychological battery of tests. RESULTS: Psychological characteristics-(1) a lower than normal academic level, (2) a job inadequate for educational level, (3) a preference for an overprotective familiar setting, and (4) a difficulty communicating own corporal image. Denial of the cardiopathy was found to be a common behaviour to normalise functioning. Very few patients had a deficit in memory, learning, or attention functions; rather, patients had a deficit in the executive functions, problem solving, and planning strategies. CONCLUSIONS: Despite a satisfactory health related quality of life, there are residual psychological and social problems in addition to impaired cognitive outcomes in the presence of a normal intelligence quotient. PMID- 15657237 TI - Cardiac troponin I release after transcatheter atrial septal defect closure depends on occluder size but not on patient's age. AB - OBJECTIVE: To examine whether transcatheter closure of secundum atrial septal defect (ASD) with the Amplatzer septal occluder leads to more myocardial injury in children than in adults. DESIGN: In a prospective study with children and adults cardiac troponin I (cTnI) serum concentrations were determined by immunoassay (AxSYM, Abbott Laboratories) before, during, and up to 20 months after surgical or transcatheter ASD closure. PATIENTS: Four groups of patients were studied: transcatheter ASD closure (group 1: 22 children, age range 3.26 14.7 years; group 2: 22 adults, 18.0-67.3 years), surgical ASD closure (group 3: 18 children, 3.12-13.5 years), and diagnostic catheterisation (group 4: 12 children, 2.68-15.0 years). RESULTS: cTnI concentrations were significantly increased after occluder implantation with higher serum concentrations in children than in adults (immediately after implantation: group 1, 3.2 (4.4) microg/l; group 2, 1.1 (4.2) microg/l; four hours after implantation: group 1, 4.8 (5.0) microg/l; group 2, 1.7 (2.3) microg/l; both p < 0.01, group 1 v group 2; one day after implantation: group 1, 3.0 (5.7) microg/l; group 2, 2.2 (5.2) microg/l) but were less than 20% of those after surgical ASD closure (group 3; p < 0.001) where the highest cTnI concentration was found (37.1 (26.3) microg/l). Diagnostic catheterisation (group 4) was not associated with detectable cTnI increase. From the cTnI concentrations the total amount of cTnI released after ASD closure was estimated for each patient. This was dependent on the size of the occluder (p < 0.05) but not on the patient's age or procedural duration. CONCLUSION: In regard to interventional ASD closure our data do not provide evidence that the child's myocardium is more vulnerable. Transcatheter ASD closure induces minor myocardial lesion, the extent of which depends on the size of the Amplatzer septal occluder but is irrespective of the patient's age. PMID- 15657238 TI - Coronary ulcer. PMID- 15657239 TI - Spontaneous dissection of native coronary arteries. PMID- 15657240 TI - Pseudoaneurysm compressing the ascending aorta on a recurrent aortic type A dissection. PMID- 15657241 TI - Cycle exercise causes a lower ventilatory response to exercise in chronic heart failure. PMID- 15657243 TI - Value of the unipolar electrogram in the diagnosis of right ventricular perforation following pacemaker implantation. PMID- 15657242 TI - Exercise modulates myocardial protein kinase B/Akt in Zucker obese rats. PMID- 15657244 TI - The health care burden of acute chest pain. PMID- 15657245 TI - Intracoronary brachytherapy for the treatment of complex in-stent restenosis. PMID- 15657246 TI - Hypoplastic aortic arch in newborns rapidly adapts to post-coarctectomy circulatory conditions. PMID- 15657247 TI - Large coronary artery aneurysms following sirolimus eluting stent implantation. PMID- 15657248 TI - No association between anti-Borrelia immunoglobulin G and cardiac disorders: results from a population based sample. PMID- 15657249 TI - Evidence of gender bias when applying the new diagnostic criteria for myocardial infarction. PMID- 15657250 TI - Evolution of aortitis after cardiac catheterisation. PMID- 15657251 TI - A promoter polymorphism of the insulin-like growth factor-I gene is associated with left ventricular hypertrophy. PMID- 15657252 TI - The putative satiety hormone PYY is raised in cardiac cachexia associated with primary pulmonary hypertension. PMID- 15657253 TI - Severe tricuspid regurgitation following a stab to the heart. PMID- 15657254 TI - Impact of stroke volume on mitral annular velocities derived from tissue Doppler imaging. PMID- 15657255 TI - Percutaneous closure of ruptured sinus of Valsalva aneurysm using an Amplatzer occluder device. PMID- 15657256 TI - Transoesophageal ultrasonography: a new approach to imaging the thoracic aorta. PMID- 15657257 TI - Heterogeneous geographic distribution of patients with aortic valve stenosis: arguments for new aetiological hypothesis. PMID- 15657258 TI - Right to left shunt 20 years after sinus venous atrial septal defect closure. PMID- 15657259 TI - Left ventricular hypertrophy in hypertension: its arrhythmogenic potential. PMID- 15657260 TI - New insights into the pathology of inherited cardiomyopathy. PMID- 15657261 TI - Non-cardiac vascular disease. PMID- 15657262 TI - TRKing signals through the Golgi. AB - The subcellular localization of transmembrane receptors and other signaling proteins has emerged as a key component in the regulation of the intensity and specificity of their activity. Recent research indicates that immature TrkA neurotrophin receptors are transactivated in the Golgi apparatus after stimulation of neuropeptide pituitary adenylate cyclase-activating polypeptide PAC1 receptors or adenosine A(2A) receptors. Transactivation occurs independently of the TrkA extracellular ligand, nerve growth factor (NGF), through a signaling pathway that is distinct from that used in the transactivation of other receptor tyrosine kinases and has consequences distinct from those elicited by NGF at the plasma membrane. PMID- 15657263 TI - Stable isotope labeling with amino acids in cell culture (SILAC) for studying dynamics of protein abundance and posttranslational modifications. AB - Stable isotope labeling with amino acids in cell culture (SILAC) is a simple and straightforward approach for in vivo incorporation of a tag into proteins for relative quantitation by mass spectrometry. SILAC is a simple, yet powerful, method for investigating the dynamics of protein abundance and posttranslational modifications. Here, we provide detailed instructions for using this method to study protein complexes, protein-protein interactions, and the dynamics of protein abundance and posttranslational modifications. We expect that SILAC will become a routine technique because of its applicability to most areas of cell biology. We have also developed a Web site (http://www.silac.org) to provide researchers with updated information about this method and related resources. PMID- 15657265 TI - Teaching resources. A model for fast-track exocytosis of synaptic vesicles. AB - The animation illustrates one model for "kiss-and-run" or fast-track release of neurotransmitter through a calcium-triggered transient pore formed between the synaptic vesicle and plasma membrane. The vesicles remain attached to the plasma membrane throughout the cycle. The animation illustrates a role for SNARE proteins and dynamin in the formation and closure of the pore. The animation would be useful in both neurobiology courses, but also any cell biology course that explores calcium-regulated secretion events. PMID- 15657264 TI - Teaching resources. Calcium-triggered exocytosis and clathrin-mediated endocytosis of synaptic vesicles. AB - The animation illustrates the recruitment, calcium-triggered exocytosis, and clathrin-mediated endocytosis of synaptic vesicles. The animation shows not only the proteins involved in synaptic vesicle fusion and recycling, but also changes in the lipid composition that are important for targeting of proteins to sites of membrane internalization. The animation would be useful in both neurobiology courses, but also any cell biology course that explores clathrin-mediated endocytic and calcium-regulated secretion events. PMID- 15657266 TI - Histologic characterization of canine dilated cardiomyopathy. AB - Dilated cardiomyopathy (DCM), characterized by chamber dilatation and myocardial systolic and diastolic dysfunction, is one of the most common heart diseases in dogs. The clinical diagnosis is based on findings on echocardiographic and Doppler examinations, with the active exclusion of other acquired or congenital heart diseases. However, the echocardiographic criteria for the diagnosis of DCM are not wholly specific for the disease, and histologic examination may be necessary for final diagnosis. Review of reports on histologic findings in dogs with clinically diagnosed DCM reveals two histologically distinct forms of DCM: 1) cardiomyopathy of Boxers and Doberman Pinschers, corresponding to the "fatty infiltration-degenerative" type and 2) the form seen in many giant, large-, and medium-sized breeds, including some Boxers and Doberman Pinschers, classified as the "attenuated wavy fiber" type of DCM. The histologic changes of the attenuated wavy fiber type of DCM may precede clinical and echocardiographic signs of heart disease, thus indicating an early stage of DCM. PMID- 15657267 TI - Sequential pathology after experimental infection with marine viral hemorrhagic septicemia virus isolates of low and high virulence in turbot (Scophthalmus maximus L). AB - Three marine viral hemorrhagic septicemia virus isolates were used to bath challenge turbot with the purpose of studying mortality and the pathology and antigen distribution over time. Two high-virulence isolates, 860/94, 4p168 and a low-virulence isolate 1p3 from a Baltic Sea herring were used. Organ samples were collected sequentially at 2, 4, 7, 10, 15, 20, 25, and 45 days postinfection. Specimens were processed for virology, histopathology, and immunohistochemistry. Organs during the early stages of infection (from 2 to 7 days) had virus isolation from all three groups only on day 7. Virus titer in kidney and heart sampled at day 25 was higher for the two virulent isolates compared with the low virulence isolate. The viral distribution in situ of the two more virulent isolates from turbot (860/94) and herring (4p168) resembled viral hemorrhagic septicemia in rainbow trout with regard to the target organs. Early infection of endothelial cells in both kidney and heart was observed. Accumulated mean mortality was 41.5% for the turbot isolate 860/94, 48% for the herring isolate 4p168, and 3.5% for the herring isolate 1p3. This study revealed that the isolates from turbot (860/94) and herring (4p168) induced significantly higher mortality compared with the virus-free control and the herring isolate (1p3). The onset of mortality is markedly later in turbot compared with what is seen in rainbow trout. PMID- 15657268 TI - Trichomonad gastritis in rhesus macaques (Macaca mulatta) infected with simian immunodeficiency virus. AB - In a retrospective study, 51 cases of gastritis (14%) were identified from among 341 necropsies performed on simian immunodeficiency virus (SIV)-infected rhesus macaques (Macaca mulatta) at the New England Primate Research Center from 1993 to 2001. Protozoa were seen in the stomach of 13 monkeys (25%) with gastritis. Two histopathologic manifestations of gastritis were observed: seven cases of lymphoplasmacytic gastritis with trichomonad trophozoites within lumens of gastric glands and four cases of necrosuppurative gastritis containing intralesional periodic acid-Schiff-positive protozoa; two cases of gastritis had morphologic features of both types of gastritis. In instances of necrosuppurative and combined lymphoplasmacytic and necrosuppurative gastritis, protozoa were 4-35 microm in diameter and round to tear-shaped. Because of the unusual morphology of the protozoa in these latter cases, transmission electron microscopy and polymerase chain reaction (PCR) were used to further identify these organisms. The protozoa were definitively identified as Tritrichomonas in all cases on the basis of ultrastructural characteristics (flagella and undulating membranes) and amplification of a 347-bp product of the 5.8S ribosomal RNA gene of Tritrichomonas foetus, Tritrichomonas suis and Tritrichomonas mobilensis by PCR using DNA extracted from stomach tissue. On the basis of these observations, we conclude that Tritrichomonas can be a significant cofactor in the development of necrosuppurative gastritis in SIV-infected rhesus macaques. PMID- 15657269 TI - Overexpression of HER-2 in feline invasive mammary carcinomas: an immunohistochemical survey and evaluation of its prognostic potential. AB - The role of c-erbB-2 protooncogene status in feline invasive mammary carcinomas (FMCs) was assessed through the HER-2 receptor immunohistochemical expression. The HER-2 overexpression was then correlated with some relevant histologic parameters and with the clinical course of the disease during a 2-year follow-up. Forty-seven FMCs from surgically treated queens were considered. Tumors were classified according to the WHO criteria and stromal or lymphatic invasion (or both) and histologic grading were recorded. The immunohistochemical staining was performed on paraffin sections and a well-defined scoring system based upon numbers of HER-2 receptors expressed on the cell surface was applied according to standard guidelines. Overall survival (OS) distributions were generated with the Kaplan-Meier method. HER-2 overexpression was detected in 28 of the 47 carcinomas (59.6%). This parameter was demonstrated to be significantly correlated with the shorter OS (P = 0.02). However, the HER-2 overexpression did not show significant correlation with histologic type, tumor grading, or presence of lymphatic invasion. Furthermore, the HER-2 overexpression appeared with a higher percentage in FMCs than what is reported in canine or human mammary carcinomas. The significant correlation with a shorter OS suggests a possible role of HER-2 as an additional marker of malignancy in FMCs and as a reliable prognostic indicator. As in the human oncology practice, the identification of the FMCs that overexpress HER-2 may also promote new therapeutic strategies. PMID- 15657270 TI - Interstitial lung disease in West Highland White Terriers. AB - Progressive respiratory failure and pulmonary fibrosis in West Highland White Terriers (WHWT) is an apparently genetic disorder of unknown pathogenesis. This study characterizes the light microscopic, ultrastructural, and immunohistochemical features of affected WHWT in comparison with lesions in usual interstitial pneumonia (UIP) of humans. Lesions in WHWT were confined to the expansion of the interstitial space of alveolar septa by extracellular matrix (ECM) determined to be mixtures of type-I and -III collagens. Features of UIP such as intra-alveolar fibroblastic foci, subpleural distribution, and honeycombing were not observed in six WHWT. Comparison with normal dogs showed no apparent increase in septal myofibroblasts. Ultrastructually, the ECM in alveolar septa consisted of large aggregates of periodic collagen filaments underlying alveolar capillaries that were surrounded by thick bands of amorphous to fine fibrillar matrix. This study suggests that chronic pulmonary disease of WHWT is a result of aberrant collagen regulation. PMID- 15657271 TI - Comparison of diagnostic detection methods for Mycobacterium avium subsp. paratuberculosis in North American bison. AB - Tissues and fecal material were collected from 14 North American bison (Bison bison) that were suspected of having Johne's disease and analyzed for the presence of Mycobacterium avium subsp. paratuberculosis (M. paratuberculosis). Sections of ileum, ileal-cecal lymph node, and three sequential sections of jejunum with their associated mesenteric lymph nodes were taken from each animal. Fecal culture indicated that 5 of 14 (35.7%) animals were infected, whereas cultures from tissues detected 12 of 14 (85.7%) animals as infected and 59 of 111 (53.2%) of the tissues as positive for M. paratuberculosis. Polymerase chain reaction analysis identified infection in 14 of 14 (100%) animals and in 91 of 112 (81.2%) tissues. In addition, tissues were processed for Ziehl-Neelsen acid fast staining, auramine O/acridine orange fluorescent staining, and immunohistochemical staining. Ziehl-Neelsen and auramine O staining identified 7 of 14 (50%) and 5 of 14 (35.7%) animals as infected and 24 of 112 (21.4%) and 28 of 112 (25%) tissues as positive, respectively. Immunohistochemical analyses of bison tissues, using antisera collected from rabbits immunized with four different preparations of M. paratuberculosis, identified a greater percentage of infected animals (ranging from 57 to 93%) and positive tissues (ranging from 28 to 46%). Collectively, these data indicate that DNA-based detection of M. paratuberculosis was more sensitive than bacterial culture or staining, identified infection in all the bison, and detected the greatest number of positive tissues within each animal. PMID- 15657272 TI - Metastatic tumors to the adrenal glands in domestic animals. AB - Although metastases to the adrenals are common in humans, they have not been thoroughly studied in animals. The purpose of this retrospective study was to document the types of malignant tumors that metastasize to canine, feline, equine, and bovine adrenals, and the rate at which they do so. The average rate of adrenal involvement in metastatic cancer was 112/534 (21.0%) in dogs, 12/81 (14.8%) in cats, 18/67 (26.9%) in horses, and 5/16 (31.3%) in cattle. In dogs, 26 different tumor types metastasized to the adrenals. Pulmonary, mammary, prostatic, gastric, and pancreatic carcinomas, and melanoma had the highest rates of metastasis to the adrenal glands in dogs. Hemangiosarcoma and melanoma had high rates of adrenal involvement in horses. In cats and cattle, relevant data were only available for lymphoma. Adrenal metastases usually occurred in the late stages of the disease. One dog had developed Addison's disease (hypoadrenocorticism) secondary to lymphoma. Metastatic lesions represented 126/472 (26.7%) of canine, 12/20 (60.0%) of feline, 21/80 (26.3%) of equine, and 5/9 (55.5%) of bovine adrenal neoplasms. This study shows that adrenal glands should be thoroughly examined during both clinical work-up and postmortems when disseminated neoplasia is suspected. PMID- 15657273 TI - An epizootic of avian pox in endemic short-toed larks (Calandrella rufescens) and Berthelot's pipits (Anthus berthelotti) in the Canary Islands, Spain. AB - Between January 2002 and November 2003, 50% (n = 395) of short-toed larks (Calandrella rufescens) and 28% (n = 139) of Berthelot's pipits (Anthus berthelotti) examined on the islands of Fuerteventura and Lanzarote, Canary Islands, had gross lesions compatible with avian pox. However, Spanish sparrows (Passer hispaniolensis, n = 128) and trumpeter finches (Bucanetes githagineus, n = 228), which inhabit the same steppe habitats associated with goat husbandry, did not have poxlike lesions. Histopathology and electron microscopy confirmed poxvirus in the lesions, whereas serology using standard, fowl poxvirus-and pigeon poxvirus-based diagnostic agar gel immunodiffusion techniques was negative, likely because of the limited (74.6% pipit; 74.9% lark) similarity between the viruses in our species and fowlpox virus on which the serologic tests rely. On the basis of polymerase chain reaction analyses, the virus isolated from dried lesions of C. rufescens has 80.5% similarity with the virus isolated from A. berthelotti and 91.3% similarity with canarypox, whereas A. berthelotti poxvirus has only 80% similarity with canarypox. We have two distinct and possibly new avian poxviruses. Both poultry and the wild birds on the farms were heavily infested by fleas, which may have acted as vectors in transmission of poxvirus. Disease prevalence in these Canary Island passerines is higher than that described in song birds in Hawaii that are now threatened, endangered, or extinct. Environmental and biological factors contributing to increased disease susceptibility of these isolated populations must be investigated. PMID- 15657274 TI - Immunohistochemical studies of epithelial cell proliferation and p53 mutation in bovine ocular squamous cell carcinoma. AB - Bovine ocular squamous cell carcinoma (OSCC) is the second most common cause of rejection due to neoplasia in slaughterhouses on Sao Miguel Island, Azores, and accounts for significant economic losses. To obtain a better insight into the genesis and neoplastic transformation process of bovine OSCC, abnormal protein expression and proliferation index were assessed by the immunohistochemical evaluation of p53 and Ki67, respectively. OSCC samples were collected from 15 bovines and were classified histologically according to the degree of differentiation into three categories: poorly, moderately, and well differentiated. Immunohistochemistry using polyclonal anti-human p53 antibody and polyclonal anti-human Ki67 antibody was performed. Ten of 15 tumors tested were immunoreactive for p53. Twelve tumors demonstrated Ki67 expression. As in human squamous cell carcinoma, p53 overexpression is frequent in bovine OSCC, providing support for a possible role of the protein in the pathogenesis of this neoplasia. No correlation between the percentage of p53 stained nuclei and the degree of differentiation was observed, although different patterns of staining were seen according to the degree of keratinization of the tumor cells. With the exception of the moderately differentiated OSCC group, Ki67 index showed significant correlation with the histologic pattern, increased proliferation being found in poorly differentiated OSCC (P = 0.013). PMID- 15657275 TI - Papillary adenocarcinoma of the extrahepatic bile duct in a Holstein cow. AB - A 15-year-old female Holstein cow was presented with a history of anorexia and weight loss. Abnormal physical examination findings included poor body condition, dehydration, icterus, and photodermatitis on nonpigmented areas of the skin of the dorsum. At necropsy, the common bile duct was greatly enlarged, with thickened walls, and tightly adhered to the hepatic capsular surface and serosa of adjacent loops of the small intestine. Two sessile, yellowish, friable, well circumscribed, cauliflower-like, bulging masses (14 and 8 cm in diameter) were attached to the inner surface of the common bile duct blocking its lumen. Microscopically, the tumor was composed of well-differentiated, columnar epithelial cells with oval or round, vesicular, basal nuclei arranged in papillary projections. Periodic acid-Schiff-positive material accumulated in the apical cytoplasm of the neoplastic cells. Immunostaining of the tumor cells was positive for cytokeratin (CK) of wide-spectrum screening and for CK 7. The diagnosis of papillary adenocarcinoma of the extrahepatic bile duct was based on the morphologic features of the neoplasm and evidence of local invasion. To the best of our knowledge, extrahepatic bile duct carcinomas have not been described in cattle. PMID- 15657276 TI - Invasive epithelial mesothelioma in a dog. AB - This report describes the gross, microscopic, and immunohistochemical features of an invasive epithelial mesothelioma in an 11-year-old neutered male Golden Retriever. The tumor involved the pericardium, pleura, mediastinum, and peritoneum and invaded into submesothelial tissues. Neoplastic cells in the thoracic fluid showed prominent features of malignancy in a background of mixed inflammatory cells and scattered erythrocytes. Histologically, the tumor consisted of nests of epithelioid cells with frequent mitotic figures and multinucleation that infiltrated submesothelial tissues. Neoplastic cells strongly coexpressed vimentin and cytokeratin intermediate filaments, which assisted in the differentiation from other epithelial tumors of nonmesothelial origin. PMID- 15657277 TI - Systemic AL amyloidosis associated with multiple myeloma in a horse. AB - AL amyloidosis is the most common type of systemic amyloidosis in humans, and it is frequently associated with multiple myeloma. But, AL amyloidosis is very rare in domestic animals. A 16-year-old Quarter horse gelding was diagnosed with systemic AL amyloidosis associated with multiple myeloma. Clinical problems were rapid weight loss, muscle atrophy, soft unformed stool, and ventral edema. Grossly, diffuse gastrointestinal hemorrhage, markedly thickened jejunal mucosa, and splenomegaly were present. Microscopically, diffuse severe amyloid deposits were present in the lamina propria of glandular stomach, duodenum, and jejunum. Much of the spleen and sternal bone marrow was replaced by neoplastic round cells, and multiple foci of amyloid were also present in the spleen and bone marrow. Electron microscopy revealed the neoplastic round cells to be of plasma cell origin, and the amyloid showed a strongly positive immunoreactivity with polyclonal anti-human immunoglobin lambda light-chain antisera. To our knowledge, this is the second report describing systemic AL amyloidosis in domestic animals associated plasma cell neoplasia and the first associated with multiple myeloma, as is common in humans. PMID- 15657278 TI - Thyroid carcinosarcoma in a dog. AB - An adult male mixed breed dog developed pain, on swallowing, that lasted for 1 week. Physical exam and radiographs revealed a subcutaneous mass in the cervical area and three metastatic nodules in the lung. The cervical mass was surgically removed and fixed in 10% buffered formalin. Histopathologic diagnosis was carcinosarcoma supported by positive immunohistochemistry (IHC) results for cytokeratin and vimentin. IHC for thyroglobulin proved that the tissue of origin was the thyroid gland. This is the fifth canine case of thyroid carcinosarcoma to be documented, but the first one to be confirmed by specific cell markers. It is an extremely rare neoplasm that also occurs in people. PMID- 15657279 TI - Paraneoplastic pemphigus in a dog with splenic sarcoma. AB - Paraneoplastic pemphigus (PNP) is an autoimmune blistering skin disease of humans that consists of characteristic skin lesions associated with concurrent neoplasia. In this study we provide histologic and serologic evidence to support a diagnosis of PNP in a dog with splenic sarcoma. Skin lesions consisted of widespread erosions involving haired skin, mucocutaneous junctions, and oral mucosa. Microscopic examination of skin and mucosae revealed lesions consistent with both pemphigus vulgaris and erythema multiforme. Immunoprecipitation confirmed that circulating IgG autoantibodies from this patient recognized five distinct antigens, presumed to represent epidermal plakins. Clinical, histopathologic, and immunologic findings in this patient were similar to those observed in human patients with PNP. The splenic neoplasia in this dog was diagnosed as a phenotypically variable spindle cell sarcoma. To date, only one other dog has been reported with PNP. This is the second reported case of canine PNP and the first patient in whom skin lesions were identified in association with splenic neoplasia. PMID- 15657280 TI - Melanotroph pituitary adenoma in a cat with diabetes mellitus. AB - A 13-year-old male, castrated, crossbred cat was referred for insulin-resistant diabetes mellitus. The cat had a ravenous appetite and a dull coat. Basal urinary corticoid/creatinine ratios were normal. In the low-dose dexamethasone suppression test there was no suppression of the (nonelevated) plasma cortisol concentration, whereas the (nonelevated) plasma adrenocorticotropic hormone (ACTH) concentration declined to low values. Basal plasma alpha-melanocyte stimulating hormone (alpha-MSH) concentrations were highly elevated (> 1,500 ng/liter). Computed tomography revealed a pituitary tumor originating from the pars intermedia (PI). After microsurgical transsphenoidal hypophysectomy, the clinical signs resolved and the cat no longer required insulin administration. Microscopic examination of the surgical specimen revealed a pituitary adenoma originating from the PI with infiltration into the neural lobe. The adenoma immunostained intensely positive for alpha-MSH and only weakly for ACTH. It is concluded that the ACTH-independent cortisol production was probably due to the (weak) glucocorticorticotropic effects of the extremely high plasma concentration of alpha-MSH and related peptides. PMID- 15657281 TI - Copper-associated chronic hepatitis and cirrhosis in a European Shorthair cat. AB - A 2-year-old, castrated, male European Shorthair cat was presented with inappetence, vomiting, and fever. Ascites and an irregularly nodular liver were observed on laparotomy; in view of the poor prognosis, the animal was euthanatized. Histopathologic examination of the liver showed a chronic hepatitis with cirrhosis associated with massive accumulation of copper in hepatocytes and macrophages, particularly in the fibrotic areas between the regenerative nodules. Electron microscopy and energy-dispersive X-ray analysis revealed lysosomal accumulation of copper in the hepatocytes. Analytical determination showed very high copper concentrations in the liver. No evidence for increased dietary or environmental copper could be found. This and the pattern of hepatic copper storage, the associated inflammation and fibrosis, strongly suggest a primary copper storage disorder. PMID- 15657282 TI - Estrogen-dependent induction of cyclooxygenase-2 in the canine prostate in vivo. AB - Cyclooxygenase (COX)-2 is involved in several physiologic and pathologic processes. COX-2 is overexpressed in human and canine prostate cancer, but little is known about COX-2 inducers in the prostate. Our objective was to investigate the effect of sex steroid hormones on COX-2 expression in the canine prostate in vivo. COX-2 expression was evaluated by immunohistochemistry in intact and castrated dogs treated with exogenous androgen or estrogen. Results showed that no COX-2 staining was observed in prostates of untreated or androgen-treated castrated or intact dogs. However, treatment of intact and castrated dogs with estrogen resulted in squamous metaplasia with intense COX-2 expression observed in both squamous epithelial cells and in cells of acini without metaplasia. This is the first report to demonstrate the induction of COX-2 by estrogen in the prostate in vivo. PMID- 15657283 TI - Comment on "Follicular stem cell carcinoma: histologic, immunohistochemical, ultrastructural, and clinical characterization in 30 dogs". PMID- 15657284 TI - Comment on "Failure to detect prion protein (PrPres) by immunohistochemistry in striated muscle tissues of animals experimentally inoculated with agents of transmissible spongiform encephalopathy". PMID- 15657285 TI - Broadening the horizons for yellow fever: new uses for an old vaccine. AB - The vaccine against yellow fever is one of the safest and most effective ever developed. With an outstanding record in humans, has this live attenuated vaccine been overlooked as a promising vector for the development of vaccines against pathogens outside its own genus? Recent studies, including a report by Tao et al. on page 201 of this issue, have sparked renewed interest. PMID- 15657286 TI - IL-12- and IL-23-induced T helper cell subsets: birds of the same feather flock together. AB - Traditionally, CD4(+) T cells have been separated into two different subsets named T helper (Th)1 and Th2. A new IL-23-driven subset of Th cells called Th(IL 17) has now been described. The data suggest that IL-23 plays an important role in the differentiation of autoreactive pathogenic T cells. Whether these IL-23 induced Th(IL-17) cells are a unique subset or are related to other Th subsets is discussed. PMID- 15657287 TI - Prevention of UV radiation-induced immunosuppression by IL-12 is dependent on DNA repair. AB - The immunostimulatory cytokine IL-12 is able to antagonize immunosuppression induced by solar/ultraviolet (UV) radiation via yet unknown mechanisms. IL-12 was recently found to induce deoxyribonucleic acid (DNA) repair. UV-induced DNA damage is an important molecular trigger for UV-mediated immunosuppression. Thus, we initiated studies into immune restoration by IL-12 to discern whether its effects are linked to DNA repair. IL-12 prevented both UV-induced suppression of the induction of contact hypersensitivity and the depletion of Langerhans cells, the primary APC of the skin, in wild-type but not in DNA repair-deficient mice. IL-12 did not prevent the development of UV-induced regulatory T cells in DNA repair-deficient mice. In contrast, IL-12 was able to break established UV induced tolerance and inhibited the activity of regulatory T cells independent of DNA repair. These data identify a new mechanism by which IL-12 can restore immune responses and also demonstrate a link between DNA repair and the prevention of UV induced immunosuppression by IL-12. PMID- 15657288 TI - NFATc2 and NFATc3 transcription factors play a crucial role in suppression of CD4+ T lymphocytes by CD4+ CD25+ regulatory T cells. AB - The phenotype of NFATc2(-/-) c3(-/-) (double knockout [DKO]) mice implies a disturbed regulation of T cell responses, evidenced by massive lymphadenopathy, splenomegaly, and autoaggressive phenomena. The population of CD4(+) CD25(+) T cells from DKO mice lacks regulatory capacity, except a small subpopulation that highly expresses glucocorticoid-induced tumor necrosis factor receptor family related gene (GITR) and CD25. However, neither wild-type nor DKO CD4(+) CD25(+) regulatory T cells (T reg cells) are able to suppress proliferation of DKO CD4(+) CD25(-) T helper cells. Therefore, combined NFATc2/c3 deficiency is compatible with the development of CD4(+) CD25(+) T reg cells but renders conventional CD4(+) T cells unresponsive to suppression, underlining the importance of NFAT proteins for sustaining T cell homeostasis. PMID- 15657289 TI - Impact of DNA ligase IV on nonhomologous end joining pathways during class switch recombination in human cells. AB - Class switch recombination (CSR) is a region-specific, transcriptionally regulated, nonhomologous recombinational process that is initiated by activation induced cytidine deaminase (AID). The initial lesions in the switch (S) regions are subsequently processed and resolved, leading to recombination of the two targeted S regions. The mechanisms by which repair and ligation of the broken DNA ends occurs is still elusive. Recently, a small number of patients lacking DNA ligase IV, a critical component of the nonhomologous end joining (NHEJ) machinery, have been identified. We show that these patients display a considerably increased donor/acceptor homology at Smu-Salpha junctions compared with healthy controls. In contrast, Smu-Sgamma junctions show an increased frequency of insertions but no increase in junctional homology. These altered patterns of junctional resolution may be related to differences in the homology between the Smu and the downstream isotype S regions, and could reflect different modes of switch junction resolution when NHEJ is impaired. These findings link DNA ligase IV, and thus NHEJ, to CSR. PMID- 15657290 TI - Yellow fever 17D as a vaccine vector for microbial CTL epitopes: protection in a rodent malaria model. AB - The yellow fever vaccine 17D (17D) is safe, and after a single immunizing dose, elicits long-lasting, perhaps lifelong protective immunity. One of the major challenges facing delivery of human vaccines in underdeveloped countries is the need for multiple injections to achieve full efficacy. To examine 17D as a vector for microbial T cell epitopes, we inserted the H-2K(d)-restricted CTL epitope of the circumsporozoite protein (CS) of Plasmodium yoelii between 17D nonstructural proteins NS2B and NS3. The recombinant virus, 17D-Py, was replication competent and stable in vitro and in vivo. A single subcutaneous injection of 10(5) PFU diminished the parasite burden in the liver by approximately 70%. The high level of protection lasted between 4 and 8 wk after immunization, but a significant effect was documented even 24 wk afterwards. Thus, the immunogenicity of a foreign T cell epitope inserted into 17D mimics some of the remarkable properties of the human vaccine. Priming with 17D-Py followed by boosting with irradiated sporozoites conferred sterile immunity to 90% of the mice. This finding indicates that the immune response of vaccine-primed individuals living in endemic areas could be sustained and magnified by the bite of infected mosquitoes. PMID- 15657291 TI - Dynamic regulation of PU.1 expression in multipotent hematopoietic progenitors. AB - PU.1 is an Ets family transcription factor that is essential for fetal liver hematopoiesis. We have generated a PU.1(gfp) reporter strain that allowed us to examine the expression of PU.1 in all hematopoietic cell lineages and their early progenitors. Within the bone marrow progenitor compartment, PU.1 is highly expressed in the hematopoietic stem cell, the common lymphoid progenitor, and a proportion of common myeloid progenitors (CMPs). Based on Flt3 and PU.1 expression, the CMP could be divided into three subpopulations, Flt3(+) PU.1(hi), Flt3(-) PU.1(hi), and Flt3(-) PU.1(lo) CMPs. Colony-forming assays and in vivo lineage reconstitution demonstrated that the Flt3(+) PU.1(hi) and Flt3(-) PU.1(hi) CMPs were efficient precursors for granulocyte/macrophage progenitors (GMPs), whereas the Flt3(-) PU.1(lo) CMPs were highly enriched for committed megakaryocyte/erythrocyte progenitors (MEPs). CMPs have been shown to rapidly differentiate into GMPs and MEPs in vitro. Interestingly, short-term culture revealed that the Flt3(+) PU.1(hi) and Flt3(-) PU.1(hi) CMPs rapidly became CD16/32(high) (reminiscent of GMPs) in culture, whereas the Flt3(-) PU.1(lo) CMPs were the immediate precursors of the MEP. Thus, down-regulation of PU.1 expression in the CMP is the first molecularly identified event associated with the restriction of differentiation to erythroid and megakaryocyte lineages. PMID- 15657292 TI - IL-23 drives a pathogenic T cell population that induces autoimmune inflammation. AB - Interleukin (IL)-23 is a heterodimeric cytokine composed of a unique p19 subunit, and a common p40 subunit shared with IL-12. IL-12 is important for the development of T helper (Th)1 cells that are essential for host defense and tumor suppression. In contrast, IL-23 does not promote the development of interferon gamma-producing Th1 cells, but is one of the essential factors required for the expansion of a pathogenic CD4(+) T cell population, which is characterized by the production of IL-17, IL-17F, IL-6, and tumor necrosis factor. Gene expression analysis of IL-23-driven autoreactive T cells identified a unique expression pattern of proinflammatory cytokines and other novel factors, distinguishing them from IL-12-driven T cells. Using passive transfer studies, we confirm that these IL-23-dependent CD4(+) T cells are highly pathogenic and essential for the establishment of organ-specific inflammation associated with central nervous system autoimmunity. PMID- 15657293 TI - High frequency of antitumor T cells in the blood of melanoma patients before and after vaccination with tumor antigens. AB - After vaccination of melanoma patients with MAGE antigens, we observed that even in the few patients showing tumor regression, the frequency of anti-vaccine T cells in the blood was often either undetectable or <10(-5) of CD8 T cells. This frequency being arguably too low for these cells to be sole effectors of rejection, we reexamined the contribution of T cells recognizing other tumor antigens. The presence of such antitumor T cells in melanoma patients has been widely reported. To begin assessing their contribution to vaccine-induced rejection, we evaluated their blood frequency in five vaccinated patients. The antitumor cytotoxic T lymphocyte (CTL) precursors ranged from 10(-4) to 3 x 10( 3), which is 10-10,000 times higher than the anti-vaccine CTL in the same patient. High frequencies were also observed before vaccination. In a patient showing nearly complete regression after vaccination with a MAGE-3 antigen, we observed a remarkably focused antitumoral response. A majority of CTL precursors (CTLp's) recognized antigens encoded by MAGE-C2, another cancer-germline gene. Others recognized gp100 antigens. CTLp's recognizing MAGE-C2 and gp100 antigens were already present before vaccination, but new clonotypes appeared afterwards. These results suggest that a spontaneous antitumor T cell response, which has become ineffective, can be reawakened by vaccination and contribute to tumor rejection. This notion is reinforced by the frequencies of anti-vaccine and antitumor CTLs observed inside metastases, as presented by Lurquin et al. (Lurquin, C., B. Lethe, V. Corbiere, I. Theate, N. van Baren, P.G. Coulie, and T. Boon. 2004. J. Exp. Med. 201:249-257). PMID- 15657294 TI - Contrasting frequencies of antitumor and anti-vaccine T cells in metastases of a melanoma patient vaccinated with a MAGE tumor antigen. AB - Melanoma patients have high frequencies of T cells directed against antigens of their tumor. The frequency of these antitumor T cells in the blood is usually well above that of the anti-vaccine T cells observed after vaccination with tumor antigens. In a patient vaccinated with a MAGE-3 antigen presented by HLA-A1, we measured the frequencies of anti-vaccine and antitumor T cells in several metastases to evaluate their respective potential contribution to tumor rejection. The frequency of anti-MAGE-3.A1 T cells was 1.5 x 10(-5) of CD8 T cells in an invaded lymph node, sixfold higher than in the blood. An antitumor cytotoxic T lymphocyte (CTL) recognizing a MAGE-C2 antigen showed a much higher enrichment with a frequency of approximately 10%, 1,000 times higher than its blood frequency. Several other antitumor T clonotypes had frequencies >1%. Similar findings were made on a regressing cutaneous metastasis. Thus, antitumor T cells were approximately 10,000 times more frequent than anti-vaccine T cells inside metastases, representing the majority of T cells present there. This suggests that the anti-vaccine CTLs are not the effectors that kill the bulk of the tumor cells, but that their interaction with the tumor generates conditions enabling the stimulation of large numbers of antitumor CTLs that proceed to destroy the tumor cells. Naive T cells appear to be stimulated in the course of this process as new antitumor clonotypes arise after vaccination. PMID- 15657295 TI - Cyclical modulation of sphingosine-1-phosphate receptor 1 surface expression during lymphocyte recirculation and relationship to lymphoid organ transit. AB - Sphingosine-1-phosphate receptor 1 (S1P(1)) was recently shown to be required for lymphocyte egress from lymphoid organs. Here we have examined the relationship between S1P(1) abundance on the cell and egress efficiency. Using an integrin neutralization approach to separate the processes of entry and exit, we show that pertussis toxin treatment reduces lymphocyte egress from lymph nodes. Retrovirally mediated S1P(1) overexpression is sufficient to reduce B cell accumulation in the splenic white pulp and to promote egress of activated T cells from lymph nodes, whereas S1P(1)(+/-) cells have reduced lymph node exit efficiency. Furthermore, lymphocyte S1P(1) is down-regulated in the blood, up regulated in lymphoid organs, and down-regulated again in the lymph. We propose that cyclical ligand-induced modulation of S1P(1) on circulating lymphocytes contributes to establishing their lymphoid organ transit time. PMID- 15657296 TI - Preferential phosphorylation of R-domain Serine 768 dampens activation of CFTR channels by PKA. AB - CFTR (cystic fibrosis transmembrane conductance regulator), the protein whose dysfunction causes cystic fibrosis, is a chloride ion channel whose gating is controlled by interactions of MgATP with CFTR's two cytoplasmic nucleotide binding domains, but only after several serines in CFTR's regulatory (R) domain have been phosphorylated by cAMP-dependent protein kinase (PKA). Whereas eight R domain serines have previously been shown to be phosphorylated in purified CFTR, it is not known how individual phosphoserines regulate channel gating, although two of them, at positions 737 and 768, have been suggested to be inhibitory. Here we show, using mass spectrometric analysis, that Ser 768 is the first site phosphorylated in purified R-domain protein, and that it and five other R-domain sites are already phosphorylated in resting Xenopus oocytes expressing wild-type (WT) human epithelial CFTR. The WT channels have lower activity than S768A channels (with Ser 768 mutated to Ala) in resting oocytes, confirming the inhibitory influence of phosphoserine 768. In excised patches exposed to a range of PKA concentrations, the open probability (P(o)) of mutant S768A channels exceeded that of WT CFTR channels at all [PKA], and the half-maximally activating [PKA] for WT channels was twice that for S768A channels. As the open burst duration of S768A CFTR channels was almost double that of WT channels, at both low (55 nM) and high (550 nM) [PKA], we conclude that the principal mechanism by which phosphoserine 768 inhibits WT CFTR is by hastening the termination of open channel bursts. The right-shifted P(o)-[PKA] curve of WT channels might explain their slower activation, compared with S768A channels, at low [PKA]. The finding that phosphorylation kinetics of WT or S768A R-domain peptides were similar provides no support for an alternative explanation, that early phosphorylation of Ser 768 in WT CFTR might also impair subsequent phosphorylation of stimulatory R domain serines. The observed reduced sensitivity to activation by [PKA] imparted by Ser 768 might serve to ensure activation of WT CFTR by strong stimuli while dampening responses to weak signals. PMID- 15657297 TI - Reversible silencing of CFTR chloride channels by glutathionylation. AB - The cystic fibrosis transmembrane conductance regulator (CFTR) is a phosphorylation- and ATP-dependent chloride channel that modulates salt and water transport across lung and gut epithelia. The relationship between CFTR and oxidized forms of glutathione is of potential interest because reactive glutathione species are produced in inflamed epithelia where they may be modulators or substrates of CFTR. Here we show that CFTR channel activity in excised membrane patches is markedly inhibited by several oxidized forms of glutathione (i.e., GSSG, GSNO, and glutathione treated with diamide, a strong thiol oxidizer). Three lines of evidence indicate that the likely mechanism for this inhibitory effect is glutathionylation of a CFTR cysteine (i.e., formation of a mixed disulfide with glutathione): (a) channels could be protected from inhibition by pretreating the patch with NEM (a thiol alkylating agent) or by lowering the bath pH; (b) inhibited channels could be rescued by reducing agents (e.g., DTT) or by purified glutaredoxins (Grxs; thiol disulfide oxidoreductases) including a mutant Grx that specifically reduces mixed disulfides between glutathione and cysteines within proteins; and (c) reversible glutathionylation of CFTR polypeptides in microsomes could be detected biochemically under the same conditions. At the single channel level, the primary effect of reactive glutathione species was to markedly inhibit the opening rates of individual CFTR channels. CFTR channel inhibition was not obviously dependent on phosphorylation state but was markedly slowed when channels were first "locked open" by a poorly hydrolyzable ATP analogue (AMP-PNP). Consistent with the latter finding, we show that the major site of inhibition is cys-1344, a poorly conserved cysteine that lies proximal to the signature sequence in the second nucleotide binding domain (NBD2) of human CFTR. This region is predicted to participate in ATP-dependent channel opening and to be occluded in the nucleotide-bound state of the channel based on structural comparisons to related ATP binding cassette transporters. Our results demonstrate that human CFTR channels are reversibly inhibited by reactive glutathione species, and support an important role of the region proximal to the NBD2 signature sequence in ATP-dependent channel opening. PMID- 15657299 TI - The "dashpot" mechanism of stretch-dependent gating in MscS. AB - The crystal structure of the small conductance mechanosensitive channel (MscS) has been an invaluable tool in the search for the gating mechanism, however many functional aspects of the channel remain unsettled. Here we characterized the gating of MscS in Escherichia coli spheroplasts in a triple mutant (mscL-, mscS-, mscK-) background. We used a pressure clamp apparatus along with software developed in-lab to generate dose-response curves directly from two-channel recordings of current and pressure. In contrast to previous publications, we found that MscS exhibits essentially voltage-independent activation by tension, but at the same time strong voltage-dependent inactivation under depolarizing conditions. The MscS activation curves obtained under saturating ramps of pressure, at different voltages, gave estimates for the energy, area, and gating charge for the closed-to-open transition as 24 kT, 18 nm2, and +0.8, respectively. The character of activation and inactivation was similar in both K+ and Na+ buffers. Perhaps the most salient and intriguing property of MscS gating was a strong dependence on the rate of pressure application. Patches subjected to various pressure ramps from 2.7 to 240 mmHg/s revealed a midpoint of activation almost independent of rate. However, the resultant channel activity was dramatically lower when pressure was applied slowly, especially at depolarizing pipette voltages. It appears that MscS prefers to respond in full to abrupt stimuli but manages to ignore those applied slowly, as if the gate were connected to the tension-transmitting element via a velocity-sensitive "dashpot." With slower ramps, channels inactivate during the passage through a narrow region of pressures below the activation midpoint. This property of "dumping" a slowly applied force may be important in environmental situations where rehydration of cells occurs gradually and release of osmolytes is not desirable. MscS often enters the inactivated state through subconducting states favored by depolarizing voltage. The inactivation rate increases exponentially with depolarization. Based on these results we propose a kinetic scheme and gating mechanism to account for the observed phenomenology in the framework of available structural information. PMID- 15657298 TI - Ca(2+)- and volume-sensitive chloride currents are differentially regulated by agonists and store-operated Ca2+ entry. AB - Using patch-clamp and calcium imaging techniques, we characterized the effects of ATP and histamine on human keratinocytes. In the HaCaT cell line, both receptor agonists induced a transient elevation of [Ca2+]i in a Ca(2+)-free medium followed by a secondary [Ca2+]i rise upon Ca2+ readmission due to store-operated calcium entry (SOCE). In voltage-clamped cells, agonists activated two kinetically distinct currents, which showed differing voltage dependences and were identified as Ca(2+)-activated (I(Cl(Ca))) and volume-regulated (I(Cl, swell)) chloride currents. NPPB and DIDS more efficiently inhibited I(Cl(Ca)) and I(Cl, swell), respectively. Cell swelling caused by hypotonic solution invariably activated I(Cl, swell) while regulatory volume decrease occurred in intact cells, as was found in flow cytometry experiments. The PLC inhibitor U-73122 blocked both agonist- and cell swelling-induced I(Cl, swell), while its inactive analogue U-73343 had no effect. I(Cl(Ca)) could be activated by cytoplasmic calcium increase due to thapsigargin (TG)-induced SOCE as well as by buffering [Ca2+]i in the pipette solution at 500 nM. In contrast, I(Cl, swell) could be directly activated by 1-oleoyl-2-acetyl-sn-glycerol (OAG), a cell-permeable DAG analogue, but neither by InsP3 infusion nor by the cytoplasmic calcium increase. PKC also had no role in its regulation. Agonists, OAG, and cell swelling induced I(Cl, swell) in a nonadditive manner, suggesting their convergence on a common pathway. I(Cl, swell) and I(Cl(Ca)) showed only a limited overlap (i.e., simultaneous activation), although various maneuvers were able to induce these currents sequentially in the same cell. TG-induced SOCE strongly potentiated I(Cl(Ca)), but abolished I(Cl, swell), thereby providing a clue for this paradox. Thus, we have established for the first time using a keratinocyte model that I(Cl, swell) can be physiologically activated under isotonic conditions by receptors coupled to the phosphoinositide pathway. These results also suggest a novel function for SOCE, which can operate as a "selection" switch between closely localized channels. PMID- 15657300 TI - PKC-alpha-mediated remodeling of the actin cytoskeleton is involved in constitutive albumin uptake by proximal tubule cells. AB - One key role of the renal proximal tubule is the reabsorption of proteins from the glomerular filtrate by constitutive receptor-mediated endocytosis. In the opossum kidney (OK) renal proximal tubule cell line, inhibition of protein kinase C (PKC) reduces albumin uptake, although the isoforms involved and mechanisms by which this occurs have not been identified. We used pharmacological and molecular approaches to investigate the role of PKC-alpha in albumin endocytosis. We found that albumin uptake in OK cells was inhibited by the pan-PKC blocker bisindolylmaleimide-1 and the isoform-specific PKC blockers Go-6976 and 2',3,3',4,4'-hexahydroxy-1,1'-biphenyl-6,6'-dimethanol dimethyl ether, indicating a role for PKC-alpha. Overexpression of a kinase deficient PKC-alpha(K368R) but not wild-type PKC-alpha significantly reduced albumin endocytosis. Western blot analysis of fractionated cells showed an increased association of PKC-alpha-green fluorescent protein with the membrane fraction within 10-20 min of exposure to albumin. We used phalloidin to demonstrate that albumin induces the formation of clusters of actin at the apical surface of OK cells and that these clusters correspond to the location of albumin uptake. These clusters were not present in cells grown in the absence of albumin. In cells treated either with PKC inhibitors or overexpressing kinase-deficient PKC-alpha(K368R) this actin cluster formation was significantly reduced. This study identifies a role for PKC-alpha in constitutive albumin uptake in OK cells by mediating assembly of actin microfilaments at the apical membrane. PMID- 15657301 TI - Enhanced Ca2+ response to AVP in preglomerular vessels from rats with genetic hypertension during different hydration states. AB - Exaggerated arginine vasopressin (AVP)-induced calcium signaling and renal vasoconstriction, characteristic in young spontaneously hypertensive rats (SHR) during euvolemia, are related to greater amounts of V1a receptor mRNA and V1a protein in preglomerular resistance arterioles. The present study determined whether V1a receptor density and calcium signal transduction in the renal vasculature of young SHR is regulated appropriately during physiological changes in hydration state. [3H]AVP ligand binding documented two- to threefold greater density of V1a receptors in euvolemic SHR vs. Wistar-Kyoto (WKY) rats. Parallel changes in V1a receptor density were observed in both strains during chronic water loading (plus approximately 50 fmol/mg) and during dehydration (minus approximately 50 fmol/mg). Affinity was unchanged. Real-time RT-PCR demonstrated that V1a mRNA in preglomerular arterioles was three times greater in euvolemic SHR. Dehydration decreased expression approximately 50% in renal vessels independent of rat strain; water loading increased V1a mRNA. Thus V1a receptor regulation correlated with changes in mRNA in a normal manner in response to chronic changes in AVP concentration, albeit set at a higher level in SHR. In dehydrated animals, AVP increased the cytosolic Ca2+ concentration ([Ca2+]i) by 60 +/- 5 and 112 +/- 13 nM cytosolic Ca2+ in WKY and SHR, respectively (P < 0.01), whereas in hydrated animals the [Ca2+]i increase was 168 +/- 10 and 220 +/ 18 nM, respectively (P < 0.05). In all hydration states, calcium signaling was greater in SHR compared with WKY (P < 0.05). Calcium signaling paralleled changes in the receptor density and mRNA. Mechanisms other than hydration state per se are likely to be responsible for the two- to threefold difference in the V1a receptor density between WKY and SHR in the renal vasculature at the critical age of 6 wk. PMID- 15657302 TI - A SAGE-based comparison between glomerular and aortic endothelial cells. AB - Endothelial cells have many characteristics in common, but significant morphological and functional differences exist between endothelial cells from different anatomic sites. The specific glomerular endothelial (GEn) cell transcript repertoire is unknown. We sought to determine whether endothelial cells derived from bovine glomeruli display a distinct transcriptional profile compared with bovine aortic endothelium (BAE) under identical conditions. Serial analysis of gene expression (SAGE), which includes known and unknown transcripts, was used to make the comparison. The GEn and BAE SAGE libraries contain 36,844 and 26,452 total tag sequences, respectively. Among 6,524 unique tag sequences represented at least 2 times in the 2 libraries, 2,094 (32%) were matched to well characterized bovine cDNA sequences (358 tags) or expressed sequence tags (EST). Identification of the human homolog was achieved for 1,035 of these tags. Forty two tags were differentially expressed in GEn. For 25 of these, the bovine cDNA or EST, and for 17 the human homolog was identified. Among all transcripts with a known bovine and human tag, seven were expressed at levels more than 10-fold higher in cultured GEn cells compared with all other SAGE libraries. The transcript "DKFZp564B076" was localized by in situ hybridization to glomerular endothelium in vivo and was shown by real-time RT-PCR to be highly abundant in glomeruli compared with aortic intima. This work supports the concept that differences in the transcriptional profile of endothelial cells from distinct origins are observed under otherwise equivalent conditions. Furthermore, we have identified the first known transcript predominant in glomerular endothelium in vivo. PMID- 15657303 TI - The cover. Home Sweet Home. PMID- 15657304 TI - Women shoulder growing HIV/AIDS burden. PMID- 15657305 TI - Depression-heart disease link probed. PMID- 15657306 TI - Down syndrome protein deters cancer: scientists reveal molecular mechanism. PMID- 15657307 TI - Infectious prions hitch ride on ferritin. PMID- 15657315 TI - Correction: actual causes of death in the United States, 2000. PMID- 15657316 TI - Length of stay, functional outcome, and mortality following medical rehabilitation. PMID- 15657317 TI - Physicians with opioid dependence. PMID- 15657318 TI - Physicians with opioid dependence. PMID- 15657319 TI - Regionalization of coronary angioplasty and travel distance. PMID- 15657320 TI - Premutation alleles and fragile X-associated tremor/ataxia syndrome. PMID- 15657321 TI - Diversion and abuse of methadone prescribed for pain management. PMID- 15657322 TI - Quality of cardiopulmonary resuscitation during out-of-hospital cardiac arrest. AB - CONTEXT: Cardiopulmonary resuscitation (CPR) guidelines recommend target values for compressions, ventilations, and CPR-free intervals allowed for rhythm analysis and defibrillation. There is little information on adherence to these guidelines during advanced cardiac life support in the field. OBJECTIVE: To measure the quality of out-of-hospital CPR performed by ambulance personnel, as measured by adherence to CPR guidelines. DESIGN AND SETTING: Case series of 176 adult patients with out-of-hospital cardiac arrest treated by paramedics and nurse anesthetists in Stockholm, Sweden, London, England, and Akershus, Norway, between March 2002 and October 2003. The defibrillators recorded chest compressions via a sternal pad fitted with an accelerometer and ventilations by changes in thoracic impedance between the defibrillator pads, in addition to standard event and electrocardiographic recordings. MAIN OUTCOME MEASURE: Adherence to international guidelines for CPR. RESULTS: Chest compressions were not given 48% (95% CI, 45%-51%) of the time without spontaneous circulation; this percentage was 38% (95% CI, 36%-41%) when subtracting the time necessary for electrocardiographic analysis and defibrillation. Combining these data with a mean compression rate of 121/min (95% CI, 118-124/min) when compressions were given resulted in a mean compression rate of 64/min (95% CI, 61-67/min). Mean compression depth was 34 mm (95% CI, 33-35 mm), 28% (95% CI, 24%-32%) of the compressions had a depth of 38 mm to 51 mm (guidelines recommendation), and the compression part of the duty cycle was 42% (95% CI, 41%-42%). A mean of 11 (95% CI, 11-12) ventilations were given per minute. Sixty-one patients (35%) had return of spontaneous circulation, and 5 of 6 patients discharged alive from the hospital had normal neurological outcomes. CONCLUSIONS: In this study of CPR during out-of-hospital cardiac arrest, chest compressions were not delivered half of the time, and most compressions were too shallow. Electrocardiographic analysis and defibrillation accounted for only small parts of intervals without chest compressions. PMID- 15657323 TI - Quality of cardiopulmonary resuscitation during in-hospital cardiac arrest. AB - CONTEXT: The survival benefit of well-performed cardiopulmonary resuscitation (CPR) is well-documented, but little objective data exist regarding actual CPR quality during cardiac arrest. Recent studies have challenged the notion that CPR is uniformly performed according to established international guidelines. OBJECTIVES: To measure multiple parameters of in-hospital CPR quality and to determine compliance with published American Heart Association and international guidelines. DESIGN AND SETTING: A prospective observational study of 67 patients who experienced in-hospital cardiac arrest at the University of Chicago Hospitals, Chicago, Ill, between December 11, 2002, and April 5, 2004. Using a monitor/defibrillator with novel additional sensing capabilities, the parameters of CPR quality including chest compression rate, compression depth, ventilation rate, and the fraction of arrest time without chest compressions (no-flow fraction) were recorded. MAIN OUTCOME MEASURE: Adherence to American Heart Association and international CPR guidelines. RESULTS: Analysis of the first 5 minutes of each resuscitation by 30-second segments revealed that chest compression rates were less than 90/min in 28.1% of segments. Compression depth was too shallow (defined as <38 mm) for 37.4% of compressions. Ventilation rates were high, with 60.9% of segments containing a rate of more than 20/min. Additionally, the mean (SD) no-flow fraction was 0.24 (0.18). A 10-second pause each minute of arrest would yield a no-flow fraction of 0.17. A total of 27 patients (40.3%) achieved return of spontaneous circulation and 7 (10.4%) were discharged from the hospital. CONCLUSIONS: In this study of in-hospital cardiac arrest, the quality of multiple parameters of CPR was inconsistent and often did not meet published guideline recommendations, even when performed by well-trained hospital staff. The importance of high-quality CPR suggests the need for rescuer feedback and monitoring of CPR quality during resuscitation efforts. PMID- 15657325 TI - Folate intake and the risk of incident hypertension among US women. AB - CONTEXT: Folate has important beneficial effects on endothelial function, but there is limited information about folate intake and risk of incident hypertension. OBJECTIVE: To determine whether higher folate intake is associated with a lower risk of incident hypertension. DESIGN, SETTING, AND PARTICIPANTS: Two prospective cohort studies of 93,803 younger women aged 27 to 44 years in the Nurses' Health Study II (1991-1999) and 62,260 older women aged 43 to 70 years in the Nurses' Health Study I (1990-1998), who did not have a history of hypertension. Baseline information on dietary folate and supplemental folic acid intake was derived from semiquantitative food frequency questionnaires and was updated every 4 years. MAIN OUTCOME MEASURE: Relative risk of incident self reported hypertension during 8 years of follow-up. RESULTS: We identified 7373 incident cases of hypertension in younger women and 12,347 cases in older women. After adjusting for multiple potential confounders, younger women who consumed at least 1000 microg/d of total folate (dietary plus supplemental) had a decreased risk of hypertension (relative risk [RR], 0.54; 95% confidence interval [CI], 0.45-0.66; P for trend <.001) compared with those who consumed less than 200 microg/d. Younger women's absolute risk reduction (ARR) was approximately 8 cases per 1000 person-years (6.7 vs 14.8 cases). The multivariable RR for the same comparison in older women was 0.82 (95% CI, 0.69-0.97; P for trend = .05). Older women's ARR was approximately 6 cases per 1000 person-years (34.7 vs 40.4 cases). When the analysis was restricted to women with low dietary folate intake (<200 microg/d), the multivariable RR for younger women with total folate intake at least 800 microg/d compared with less than 200 microg/d was 0.55 (95% CI, 0.32 0.94; P for trend = .03), and 0.61 (95% CI, 0.34-1.11; P for trend = .05) in the older cohort. Among women who did not take folic acid-containing supplements, dietary folate intake of 400 microg/d or more was not significantly associated with risk of hypertension. CONCLUSION: Higher total folate intake was associated with a decreased risk of incident hypertension, particularly in younger women. PMID- 15657324 TI - Effectiveness of a quality improvement intervention for adolescent depression in primary care clinics: a randomized controlled trial. AB - CONTEXT: Depression is a common condition associated with significant morbidity in adolescents. Few depressed adolescents receive effective treatment for depression in primary care settings. OBJECTIVE: To evaluate the effectiveness of a quality improvement intervention aimed at increasing access to evidence-based treatments for depression (particularly cognitive-behavior therapy and antidepressant medication), relative to usual care, among adolescents in primary care practices. DESIGN, SETTING, AND PARTICIPANTS: Randomized controlled trial conducted between 1999 and 2003 enrolling 418 primary care patients with current depressive symptoms, aged 13 through 21 years, from 5 health care organizations purposively selected to include managed care, public sector, and academic medical center clinics in the United States. INTERVENTION: Usual care (n = 207) or 6 month quality improvement intervention (n = 211) including expert leader teams at each site, care managers who supported primary care clinicians in evaluating and managing patients' depression, training for care managers in manualized cognitive behavior therapy for depression, and patient and clinician choice regarding treatment modality. Participating clinicians also received education regarding depression evaluation, management, and pharmacological and psychosocial treatment. MAIN OUTCOME MEASURES: Depressive symptoms assessed by Center for Epidemiological Studies-Depression Scale (CES-D) score. Secondary outcomes were mental health-related quality of life assessed by Mental Health Summary Score (MCS-12) and satisfaction with mental health care assessed using a 5-point scale. RESULTS: Six months after baseline assessments, intervention patients, compared with usual care patients, reported significantly fewer depressive symptoms (mean [SD] CES-D scores, 19.0 [11.9] vs 21.4 [13.1]; P = .02), higher mental health related quality of life (mean [SD] MCS-12 scores, 44.6 [11.3] vs 42.8 [12.9]; P = .03), and greater satisfaction with mental health care (mean [SD] scores, 3.8 [0.9] vs 3.5 [1.0]; P = .004). Intervention patients also reported significantly higher rates of mental health care (32.1% vs 17.2%, P<.001) and psychotherapy or counseling (32.0% vs 21.2%, P = .007). CONCLUSIONS: A 6-month quality improvement intervention aimed at improving access to evidence-based depression treatments through primary care was significantly more effective than usual care for depressed adolescents from diverse primary care practices. The greater uptake of counseling vs medication under the intervention reinforces the importance of practice interventions that include resources to enable evidence-based psychotherapy for depressed adolescents. PMID- 15657326 TI - Effect of estrogen therapy on gallbladder disease. AB - CONTEXT: Estrogen therapy is thought to promote gallstone formation and cholecystitis but most data derive from observational studies rather than randomized trials. OBJECTIVE: To determine the effect of estrogen therapy in healthy postmenopausal women on gallbladder disease outcomes. DESIGN, SETTING, AND PARTICIPANTS: Two randomized, double-blind, placebo-controlled trials conducted at 40 US clinical centers. The volunteer sample was 22,579 community dwelling women aged 50 to 79 years without prior cholecystectomy. INTERVENTION: Women with hysterectomy were randomized to 0.625 mg/d of conjugated equine estrogens (CEE) or placebo (n = 8376). Women without hysterectomy were randomized to estrogen plus progestin (E + P), given as CEE plus 2.5 mg/d of medroxyprogesterone acetate (n = 14,203). MAIN OUTCOME MEASURES: Participants reported hospitalizations for gallbladder diseases and gallbladder-related procedures, with events ascertained through medical record review. Cox proportional hazards regression was used to assess hazard ratios (HRs) and 95% confidence intervals (CIs) using intention-to-treat and time-to-event methods. RESULTS: The CEE and the E + P groups were similar to their respective placebo groups at baseline. The mean follow-up times were 7.1 years and 5.6 years for the CEE and the E + P trials, respectively. The annual incidence rate for any gallbladder event was 78 events per 10,000 person-years for the CEE group (vs 47/10,000 person-years for placebo) and 55 per 10,000 person-years for E + P (vs 35/10,000 person-years for placebo). Both trials showed greater risk of any gallbladder disease or surgery with estrogen (CEE: HR, 1.67; 95% CI, 1.35-2.06; E + P: HR, 1.59; 95% CI, 1.28-1.97). Both trials indicated a higher risk for cholecystitis (CEE: HR, 1.80; 95% CI, 1.42-2.28; E + P: HR, 1.54; 95% CI 1.22 1.94); and for cholelithiasis (CEE: HR, 1.86; 95% CI, 1.48-2.35; E + P: HR, 1.68; 95% CI, 1.34-2.11) for estrogen users. Also, women undergoing estrogen therapy were more likely to receive cholecystectomy (CEE: HR, 1.93; 95% CI, 1.52-2.44; E + P: HR, 1.67; 95% CI, 1.32-2.11), but not other biliary tract surgery (CEE: HR, 1.18; 95% CI, 0.68-2.04; E + P: HR, 1.49; 95% CI, 0.78-2.84). CONCLUSIONS: These data suggest an increase in risk of biliary tract disease among postmenopausal women using estrogen therapy. The morbidity and cost associated with these outcomes may need to be considered in decisions regarding the use of estrogen therapy. PMID- 15657327 TI - Adolescents' reports of parental knowledge of adolescents' use of sexual health services and their reactions to mandated parental notification for prescription contraception. AB - CONTEXT: Legislation has been proposed that would mandate parental notification for adolescents younger than 18 years (minors) obtaining prescription contraception from federally funded family planning clinics. OBJECTIVE: To determine the extent to which parents are currently aware that their teenage daughters are accessing reproductive health services and how minors would react in the face of mandated parental involvement laws for prescription birth control. DESIGN, SETTING, AND PARTICIPANTS: A total of 1526 female adolescents younger than 18 years seeking reproductive health services at a national sample of 79 family planning clinics were surveyed between May 2003 and February 2004. MAIN OUTCOME MEASURES: Proportions of minor females who reported that a parent or guardian was aware that they were at the family planning clinic and, under conditions of mandated parental involvement, proportions of minors who would access prescription contraceptives at family planning clinics or engage in unsafe sex. RESULTS: Sixty percent of minors reported that a parent or guardian knew they were accessing sexual health services at the clinic. Fifty-nine percent of all adolescents would use the clinic for prescription contraception even if parental notification were mandated. This response was less common (29.5%) among adolescents whose parents were unaware of their clinic visits and more common (79%) among those whose parents were aware. Many adolescents gave more than 1 response to mandated parental involvement. Forty-six percent would use an over the-counter method, and 18% would go to a private physician. Seven percent said that they would stop having sex as one response, but only 1% indicated this would be their only reaction. One in 5 adolescents would use no contraception or rely on withdrawal as one response to mandated notification. CONCLUSIONS: Most minor adolescent females seeking family planning services report that their parents are aware of their use of services. Most would continue to use clinic services if parental notification were mandated. However, mandated parental notification laws would likely increase risky or unsafe sexual behavior and, in turn, the incidence of adolescent pregnancy and sexually transmitted diseases. PMID- 15657328 TI - A simplified approach to the management of non-ST-segment elevation acute coronary syndromes. AB - CONTEXT: While current practice guidelines provide an evidence-based approach to management of acute coronary syndromes (ACS), application of the evidence by individual physicians has been suboptimal. OBJECTIVE: To assess and synthesize the evidence regarding optimal management of non-ST-segment elevation ACS (NSTE ACS). DATA SOURCES: Systematic searches of peer-reviewed publications were performed in MEDLINE and the Cochrane Database from January 1990 through November 2004, with consultation by content experts. Search terms included antiplatelet therapy, antithrombotic therapy, angiotensin-converting enzyme inhibition, angiotensin receptor blockade, beta-blockade, hypertension, hyperlipidemia, cigarette smoking, diet, diabetes mellitus, exercise, myocardial ischemia, and coronary artery disease. STUDY SELECTION AND DATA EXTRACTION: Criteria for selection of studies included controlled study design, English language, and clinical pertinence. Data quality was based on the publishing journal and relevance to clinical management of NSTE-ACS. DATA SYNTHESIS: While outcomes of controlled studies support a comprehensive approach in the management of patients with NSTE-ACS, many physicians perceive existing guidelines as lengthy and complex. After risk stratification to identify those patients most likely to benefit from an early invasive vs early conservative strategy, a comprehensive management plan can be assembled through an "ABCDE" approach. The elements of this include "A" for antiplatelet therapy, anticoagulation, angiotensin converting enzyme inhibition, and angiotensin receptor blockade; "B" for beta blockade and blood pressure control; "C" for cholesterol treatment and cigarette smoking cessation; "D" for diabetes management and diet; and "E" for exercise. CONCLUSION: An "ABCDE" approach for the management of NSTE-ACS provides a practical and systematic means to implement evidence-based medicine into clinical practice. PMID- 15657330 TI - Cardiopulmonary resuscitation in the real world: when will the guidelines get the message? PMID- 15657329 TI - A perspective on US drug reimportation. PMID- 15657331 TI - JAMA patient page. Cardiopulmonary resuscitation (CPR). PMID- 15657332 TI - More evidence that locoregional radiation therapy improves survival: what should we do? PMID- 15657333 TI - Measuring DNA repair capacity: small steps. PMID- 15657334 TI - Anastrozole data show continued delay in relapse, but no clear survival advantage. PMID- 15657335 TI - Which came first? Studies clarify role of aneuploidy in cancer. PMID- 15657336 TI - Cancer's big sleep: senescence may be potential target for cancer therapies. PMID- 15657337 TI - Stat bite: Stomach cancer in males by world region, 2002. PMID- 15657338 TI - Redesigning NCI's clinical trials system: integration is the goal. PMID- 15657339 TI - Designing the Selenium and Vitamin E Cancer Prevention Trial (SELECT). AB - Prostate cancer continues to be a major health threat, especially among African American men. The Selenium and Vitamin E Cancer Prevention Trial (SELECT), which opened on July 25, 2001, was planned to study possible agents for the prevention of prostate cancer in a population of 32,400 men in the United States, including Puerto Rico, and Canada. SELECT is a phase III randomized, placebo-controlled trial of selenium (200 microg/day from L-selenomethionine) and/or vitamin E (400 IU/day of all rac alpha-tocopheryl acetate) supplementation for a minimum of 7 years (maximum of 12 years) in non-African American men at least 55 years of age and African American men at least 50 years of age. SELECT is a large, simple trial that conforms as closely as possible with community standards of care. This commentary discusses the design problems the SELECT investigators had to resolve in developing the trial, including the role of prostate cancer screening, the best forms and doses of the study agents, and estimation of the event (prostate cancer) rate of men on the placebo arm. PMID- 15657340 TI - Epigenetic changes in prostate cancer: implication for diagnosis and treatment. AB - Prostate cancer is the most common noncutaneous malignancy and the second leading cause of cancer death among men in the United States. DNA methylation and histone modifications are important epigenetic mechanisms of gene regulation and play essential roles both independently and cooperatively in tumor initiation and progression. Aberrant epigenetic events such as DNA hypo- and hypermethylation and altered histone acetylation have both been observed in prostate cancer, in which they affect a large number of genes. Although the list of aberrantly epigenetically regulated genes continues to grow, only a few genes have, so far, given promising results as potential tumor biomarkers for early diagnosis and risk assessment of prostate cancer. Thus, large-scale screening of aberrant epigenetic events such as DNA hypermethylation is needed to identify prostate cancer-specific epigenetic fingerprints. The reversibility of epigenetic aberrations has made them attractive targets for cancer treatment with modulators that demethylate DNA and inhibit histone deacetylases, leading to reactivation of silenced genes. More studies into the mechanism and consequence of demethylation are required before the cancer epigenome can be safely manipulated with therapeutics as a treatment modality. In this review, we examine the current literature on epigenetic changes in prostate cancer and discuss the clinical potential of cancer epigenetics for the diagnosis and treatment of this disease. PMID- 15657342 TI - DNA repair capacity of lymphoblastoid cell lines from sisters discordant for breast cancer. AB - BACKGROUND: Interindividual differences in DNA repair capacity may influence cancer risk. We tested whether the nucleotide excision repair pathway was deficient in breast cancer case patients by analyzing sister pairs. METHODS: Cell lines derived from sisters discordant for breast cancer (137 families containing 158 case patients and 154 control sisters) were obtained from the Metropolitan New York Registry of Breast Cancer Families. Lymphoblastoid cells were treated with benzo[a]pyrene diolepoxide (BPDE) for 30 minutes and were either harvested immediately or were washed and cultured in complete medium for 4 hours to allow DNA repair. Immunofluorescence using a polyclonal anti-BPDE-DNA primary antibody was used to quantify BPDE-DNA adducts. Percent DNA repair capacity was calculated from the difference between staining immediately after treatment minus that after 4 hours of repair, divided by the initial damage and was categorized into quartiles based on control values. Odds ratios and 95% confidence intervals (CIs) were calculated using conditional logistic regression models adjusted for age at blood donation, body mass index, and smoking. Statistical tests were two-sided. RESULTS: Mean percent DNA repair capacity was lower in breast cancer case patients than in control subjects (difference = 8.6, 95% CI = 4.3 to 13.8, P = .001). Using the quartile with the highest percent DNA repair capacity as the referent group, adjusted odds ratios of breast cancer increased from 1.23 (95% CI = 0.57 to 2.65) to 2.38 (95% CI = 1.17 to 4.86) to 2.99 (95% CI = 1.45 to 6.17) (P(trend) = .002) as DNA repair capacity decreased. CONCLUSIONS: Deficient DNA repair capacity is associated with increased breast cancer risk. PMID- 15657341 TI - Locoregional radiation therapy in patients with high-risk breast cancer receiving adjuvant chemotherapy: 20-year results of the British Columbia randomized trial. AB - BACKGROUND: The British Columbia randomized radiation trial was designed to determine the survival impact of locoregional radiation therapy in premenopausal patients with lymph node-positive breast cancer treated by modified radical mastectomy and adjuvant chemotherapy. Three hundred eighteen patients were assigned to receive no further therapy or radiation therapy (37.5 Gy in 16 fractions). Previous analysis at the 15-year follow-up showed that radiation therapy was associated with a statistically significant improvement in breast cancer survival but that improvement in overall survival was of only borderline statistical significance. We report the analysis of data from the 20-year follow up. METHODS: Survival was analyzed by the Kaplan-Meier method. Relative risk estimates were calculated by the Wald test from the proportional hazards regression model. All statistical tests were two-sided. RESULTS: At the 20 year follow up (median follow up for live patients: 249 months) chemotherapy and radiation therapy, compared with chemotherapy alone, were associated with a statistically significant improvement in all end points analyzed, including survival free of isolated locoregional recurrences (74% versus 90%, respectively; relative risk [RR] = 0.36, 95% confidence interval [CI] = 0.18 to 0.71; P = .002), systemic relapse-free survival (31% versus 48%; RR = 0.66, 95% CI = 0.49 to 0.88; P = .004), breast cancer-free survival (48% versus 30%; RR = 0.63, 95% CI = 0.47 to 0.83; P = .001), event-free survival (35% versus 25%; RR = 0.70, 95% CI = 0.54 to 0.92; P = .009), breast cancer-specific survival (53% versus 38%; RR = 0.67, 95% CI = 0.49 to 0.90; P = .008), and, in contrast to the 15-year follow up results, overall survival (47% versus 37%; RR = 0.73, 95% CI = 0.55 to 0.98; P = .03). Long-term toxicities, including cardiac deaths (1.8% versus 0.6%), were minimal for both arms. CONCLUSION: For patients with high-risk breast cancer treated with modified radical mastectomy, treatment with radiation therapy (schedule of 16 fractions) and adjuvant chemotherapy leads to better survival outcomes than chemotherapy alone, and it is well tolerated, with acceptable long term toxicity. PMID- 15657344 TI - The role of overdiagnosis and reclassification in the marked increase of esophageal adenocarcinoma incidence. AB - BACKGROUND: The incidence of esophageal adenocarcinoma is rising dramatically. This increase may reflect increased disease burden, reclassification of related cancers, or overdiagnosis resulting from increased diagnostic intensity, particularly upper endoscopy for patients with gastroesophageal reflux disease or Barrett esophagus. METHODS: We used the National Cancer Institute's Surveillance, Epidemiology, and End Results database to extract information on incidence, stage distribution, and disease-specific mortality for esophageal adenocarcinoma as well as information on related cancers. RESULTS: From 1975 to 2001, the incidence of esophageal adenocarcinoma rose approximately sixfold in the United States (from 4 to 23 cases per million), a relative increase greater than that for melanoma, breast, or prostate cancer. Reclassification of squamous cell carcinoma is an unlikely explanation for the rise in incidence, because the anatomic distribution of esophageal cancer in general has changed. The only location with increased incidence is the lower third of the esophagus-the site where adenocarcinoma typically arises. Reclassification of adjacent gastric cancer is also unlikely because its incidence has also increased. Because there has been little change in the proportion of patients found with in situ or localized disease at diagnosis since 1975 (from 25% to 31%) and because esophageal adenocarcinoma mortality has increased more than sevenfold (from 2 to 15 deaths per million), overdiagnosis can be excluded as an explanation for the rise in incidence. CONCLUSION: The rising incidence of esophageal adenocarcinoma represents a real increase in disease burden. PMID- 15657343 TI - Role of human cripto-1 in tumor angiogenesis. AB - BACKGROUND: Human cripto-1 (CR-1) promotes cell transformation and increases migration and invasion of various mouse and human epithelial cell lines. We investigated whether CR-1 also stimulates angiogenesis. METHODS: We used human umbilical vein endothelial cells (HUVECs) to measure in vitro migration with fibronectin-coated Boyden chambers, invasion with Matrigel-coated Boyden chambers, proliferation with a tetrazolium salt, and differentiation with an in vitro Matrigel assay. We investigated new blood vessel formation in vivo by use of Matrigel-filled silicone cylinders implanted under the skin of nude mice and by use of a breast cancer xenograft model with CR-1-transfected or control Neo transfected MCF-7 human breast cancer cells. We also used a blocking anti-CR-1 monoclonal antibody to investigate the role of CR-1 in angiogenesis in vivo and in vitro. All statistical tests were two-sided. RESULTS: CR-1 stimulated HUVEC proliferation, migration, and invasion and induced HUVEC differentiation into vascular-like structures on Matrigel. In vivo, recombinant CR-1 protein induced microvessel formation in Matrigel-filled silicone cylinders, and microvessel formation was statistically significantly inhibited with a blocking anti-CR-1 monoclonal antibody (CR-1 and antibody = 127% of microvessel formation compared with that in untreated control cylinders and CR-1 alone = 259%; difference = 132%, 95% confidence interval [CI] = 123% to 140%; P<.001). Tumors formed by CR-1 transfected MCF-7 cells in the cleared mammary fat pad of nude mice had higher microvessel density than tumors formed by control Neo-transfected MCF-7 cells (CR 1-transfected cells = 4.66 vessels per field and Neo-transfected cells = 2.33 vessels per field; difference = 2.33 vessels per field, 95% CI = 1.2 to 2.8; P = .004). CONCLUSION: CR-1 appears to have an important role in the multistep process of angiogenesis. PMID- 15657346 TI - Re: New guidelines to evaluate the response to treatment in solid tumors (ovarian cancer). PMID- 15657345 TI - A study of the impact of adding HPV types to cervical cancer screening and triage tests. AB - Use of human papillomavirus (HPV) testing in cervical cancer prevention is increasing rapidly. A DNA test for 13 HPV types that can cause cervical cancer is approved in the United States for co-screening with cytology of women >or=30 years old and for triage of women of all ages with equivocal cytology. However, most infections with HPV are benign. We evaluated trade-offs between specificity and sensitivity for approximately 40 HPV types in predicting cervical intraepithelial neoplasia 3 and cancer in two prospective studies: a population based screening study that followed 6196 women aged 30-94 years from Costa Rica for 7 years and a triage study that followed 3363 women aged 18-90 years with equivocal cytology in four U.S. centers for 2 years. For both screening and triage, testing for more than about 10 HPV types decreased specificity more than it increased sensitivity. The minimal increases in sensitivity and in negative predictive value achieved by adding HPV types to DNA tests must be weighed against the projected burden to thousands of women falsely labeled as being at high risk of cervical cancer. PMID- 15657347 TI - Re: Polymorphisms associated with circulating sex hormone levels in postmenopausal women. PMID- 15657348 TI - Induction of caspase-dependent, p53-mediated apoptosis by apigenin in human neuroblastoma. AB - Neuroblastoma is a pediatric tumor accounting for 15% of childhood cancer deaths and has a poor prognosis in children >1 year of age. We investigated the ability of apigenin, a nonmutagenic dietary flavonoid that has been shown to have antitumor effects in various tumor cell lines, to inhibit growth and induce apoptosis of the human neuroblastoma cell lines NUB-7, LAN-5, and SK-N-BE(2). Apigenin inhibited colony-forming ability and survival, and induced apoptosis of NUB-7 and LAN-5 cells. The presence of the C2-C3 double bond and the 4'-OH group on the flavonoid structure correlated with the growth-inhibitory potential of apigenin. Furthermore, apigenin inhibited NUB-7 xenograft tumor growth in anonobese diabetic/severe combined immunodeficiency mouse model, likely by inducing apoptosis. Apigenin did not inhibit survival of primary sympathetic neurons, suggesting that it is not toxic to nontransformed cells. The mechanism of action of apigenin seems to involve p53, as it increased the levels of p53 and the p53-induced gene products p21WAF1/CIP1 and Bax. Furthermore, apigenin (15-60 micromol/L) induced cell death and apoptosis of neuroblastoma cells expressing wild-type but not mutant p53. Apigenin increased caspase-3 activity and PARP cleavage, and Z-VAD-FMK, a broad-spectrum caspase-3 inhibitor, rescued NUB-7 cells from apigenin-mediated apoptosis indicating that apigenin induced apoptosis in acaspase-dependent manner. Overexpression of Bcl-X(L) rescued NUB-7 from apigenin-induced cell death, suggesting that Bax activity is important for the action of apigenin. Apigenin is thus a candidate therapeutic for neuroblastoma that likely acts by regulating a p53-Bax-caspase-3 apoptotic pathway. PMID- 15657350 TI - (-)-Gossypol acts directly on the mitochondria to overcome Bcl-2- and Bcl-X(L) mediated apoptosis resistance. AB - Aberrant overexpression of antiapoptotic members of the Bcl-2 protein family, including Bcl-2 and Bcl-X(L), contributes to malignant transformation and subsequent resistance to traditional chemotherapeutics. Thus, these proteins represent attractive targets for novel anticancer agents. The small molecule, gossypol, was initially investigated as a contraceptive agent, but subsequently has been shown to possess anticancer properties in vitro and in vivo. Recently gossypol has been found to bind to Bcl-X(L) and, with less affinity, to Bcl-2. Here we investigate the ability of the (-) enantiomer of gossypol, (-)-gossypol, to overcome the apoptosis resistance conferred by Bcl-2 or Bcl-X(L) overexpression in Jurkat T leukemia cells. (-)-Gossypol potently induced cell death in Jurkat cells overexpressing Bcl-2 (IC50, 18.1+/-2.6 micromol/L) or Bcl X(L) (IC50, 22.9+/-3.7 micromol/L). Vector-transfected control cells were also potently killed by (-)-gossypol (IC50, 7.0+/-2.7 micromol/L). By contrast, the chemotherapy drug etoposide only induced efficient killing of vector-transfected cells (IC50, 9.6+/-2.3 micromol/L). Additionally, (-)-gossypol was more efficient than etoposide at inducing caspase-3 activation and phosphatidylserine externalization in the setting of Bcl-2 or Bcl-X(L) overexpression. (-)-Gossypol induced apoptosis was associated with Bak activation and release of cytochrome c from mitochondria, suggesting a mitochondrial-mediated apoptotic mechanism. Moreover, (-)-gossypol treatment of isolated mitochondria purified from Bcl-2 overexpressing cells also resulted in cytochrome c release, indicating a possible direct action on Bcl-2 present in the mitochondrial outer membrane. Taken together, these results suggest that (-)-gossypol is a potent and novel therapeutic able to overcome apoptosis resistance by specifically targeting the activity of antiapoptotic Bcl-2 family members. (-)-Gossypol may be a promising new agent to treat malignancies that are resistant to conventional therapies. PMID- 15657349 TI - Preclinical studies of a nonpeptidic small-molecule inhibitor of Bcl-2 and Bcl X(L) [(-)-gossypol] against diffuse large cell lymphoma. AB - Overexpression of Bcl-2/Bcl-X(L) protein has been observed in more than 80% of B cell lymphomas. Diffuse large cell lymphoma (DLCL) is the most common subtype of non-Hodgkin's lymphoma. (-)-Gossypol, a natural product isolated from cottonseeds, was discovered as a potent small-molecule inhibitor of Bcl-2 and Bcl X(L) proteins, with a Ki value in the nanomole per liter range for both. In vitro, (-)-gossypol showed significant growth inhibition effect against WSU-DLCL2 lymphoma cell line and fresh cells obtained from a lymphoma patient with no effect on normal peripheral blood lymphocytes. As expected (-)-gossypol induced complete cytochrome c release from mitochondria, increased caspases-3 and -9 activity, and caused apoptotic death without affecting protein levels of Bcl-2, Bcl-X(L), Bax, and Bak. The addition of cyclophosphamide-Adriamycin-vincristine prednisolone (CHOP) regimen to lymphoma cells preexposed to (-)-gossypol enhanced killing significantly. The maximum tolerated dose of (-)-gossypol in severe combined immunodeficient (SCID) mice was 40 mg/kg for three i.v. injections when given alone and 20 mg/kg x 3 when given in combination with CHOP. Using WSU-DLCL2 SCID mouse xenograft model, the tumor growth inhibition, the tumor growth delay, and the log10 kill of mice treated with (-)-gossypol + CHOP were better than CHOP or (-)-gossypol alone. We conclude that adding Bcl-2/Bcl-X(L) small-molecule inhibitor to standard chemotherapy may prove an effective strategy in lymphoma therapy. PMID- 15657351 TI - The nuclear factor kappa B inhibitor parthenolide restores ICI 182,780 (Faslodex; fulvestrant)-induced apoptosis in antiestrogen-resistant breast cancer cells. AB - The molecular mechanisms underlying the acquisition of resistance to the antiestrogen Faslodex are poorly understood, although enhanced expression and activity of nuclear factor kappaB (NFkappaB) have been implicated as a critical element of this phenotype. The purpose of this study was to elucidate the mechanism by which NFkappaB up-regulation contributes to Faslodex resistance and to determine whether pharmacologic inhibition of NFkappaB by the small molecule parthenolide could restore Faslodex-mediated suppression of cell growth. Basal expression of multiple NFkappaB-related molecules in MCF7-derived LCC1 (antiestrogen-sensitive) and LCC9 (antiestrogen-resistant) breast cancer cells was determined, and cells were treated with Faslodex or parthenolide. The effect of these drugs either singly or in combination was assessed by cell proliferation, estrogen receptor (ER)-dependent transcriptional activation, cell cycle analysis, and apoptosis assays. Expression of the p65 NFkappaB subunit and the upstream NFkappaB regulator IkappaB kinase gamma/NFkappaB essential modulator were increased in the resistant MCF7/LCC9 cells (P=0.001 and 0.04, respectively). Whereas MCF7/LCC9 cells were unresponsive to Faslodex alone, parthenolide effectively inhibited MCF7/LCC9 cell proliferation and the combination of Faslodex and parthenolide resulted in a 4-fold synergistic reduction in cell growth (P=0.03). This corresponded to a restoration of Faslodex-induced apoptosis (P=0.001), with no observable changes in ER-dependent transcription or cell cycle phase distribution. Because parthenolide has shown safety in Phase I clinical trials, these findings have direct clinical relevance and provide support for the design of clinical studies combining antiestrogens and parthenolide in ER positive breast cancer. PMID- 15657352 TI - Up-regulation of c-Jun-NH2-kinase pathway contributes to the induction of mitochondria-mediated apoptosis by alpha-tocopheryl succinate in human prostate cancer cells. AB - Previously, alpha-tocopheryl succinate (alpha-TOS) has been reported to induce caspase-mediated apoptosis in PC-3 human prostate cancer cells. Caspase-9 was among several initiator caspases activated by alpha-TOS, suggesting a potential contribution of the intrinsic apoptotic pathway in mediating the response to alpha-TOS. Gene expression microarray was carried out as a screen to identify novel signaling molecules modulated by alpha-TOS, with a special focus on those known to play a role in mitochondria-mediated apoptosis. We discovered that Ask1, GADD45beta, and Sek1, three key components of the stress-activated mitogen activated protein kinase pathway, are novel targets of alpha-TOS. Western blot analysis showed increased levels of phospho-Sek1 and phospho-c-Jun-NH2-kinase (JNK) in addition to total Ask1, GADD45beta, and Sek1. alpha-TOS also altered JNK specific phosphorylation of Bcl-2 and Bim in a manner consistent with enhanced mitochondrial translocation of Bax and Bim. Because the expression level of most Bcl-2 family members remained unchanged, the posttranslational modification of Bcl-2 and Bim by JNK is likely to be a driving force in alpha-TOS activation of the intrinsic apoptotic pathway. Based on our findings, we propose a working model to capture the salient features of the apoptotic signaling circuitry of alpha-TOS. PMID- 15657353 TI - Selective inhibitors of MEK1/ERK44/42 and p38 mitogen-activated protein kinases potentiate apoptosis induction by sulindac sulfide in human colon carcinoma cells. AB - The nonsteroidal anti-inflammatory drug (NSAID) sulindac prevents experimental colon cancer and can regress precancerous polyps in humans. Sulindac sulfide inhibits cyclooxygenase (COX)-mediated prostaglandin synthesis and retards the growth of cultured colon cell lines primarily by inducing apoptosis. Given the known role of mitogen-activated protein kinase (MAPK) in signal transduction and the regulation of cell survival and death, we determined the effect of sulindac sulfide on MAPK activation, COX-2 expression, and apoptosis induction in HCA-7 human colon cancer cells. Sulindac sulfide treatment was associated with activation of ERKp44/42 and p38 MAPK in a dosage- and time-dependent manner, and also activated upstream MEK. Similar results were seen in HCT-15 cells and also with the selective COX-2 inhibitor NS398. ERKp44/42 and p38 activation were accompanied by an induction of COX-2 protein expression. Selective inhibitors of sulindac sulfide-induced ERKp44/42 (PD98059) and p38 MAPK (SB203580) activation also suppressed the induction of COX-2 by this NSAID. Furthermore, both MAPK inhibitors significantly augmented sulindac sulfide-induced apoptosis, as did suppression of constitutive COX-2 using antisense oligonucleotides. In conclusion, MEK/ERK and p38 MAPK activation mediate COX-2 induction by sulindac sulfide. Selective inhibitors of these MAPKs potentiate apoptosis induction by this NSAID, suggesting a novel strategy for the prevention or treatment of colorectal cancer. PMID- 15657354 TI - Epigenetic regulation of O6-methylguanine-DNA methyltransferase gene expression by histone acetylation and methyl-CpG binding proteins. AB - Transcriptional silencing of the DNA repair gene, O6-methylguanine-DNA methyltransferase (MGMT) in a proportion of transformed cell lines is associated with methylated CpG hotspots in the MGMT 5' flank. The goal of the study was to evaluate the mechanism by which CpG methylation of theMGMT promoter region influenced silencing of the gene. Analysis of histone acetylation status in two regions of the promoter using chromatin immunoprecipitation assay showed that a higher level of histone acetylation was associated with expression in three MGMT expressing cell lines (HeLa CCL2, HT29, and Raji) compared with three MGMT silenced cell lines (HeLa S3, BE, and TK6). To determine how the modulation of CpG methylation and histone acetylation influenced MGMT expression, we exposed the cells to 5-aza-2'deoxycytidine (5-Aza-dC), inhibitor of DNA methylation, which strongly up-regulated MGMT expression in three MGMT-silenced cell lines whereas trichostatin A, inhibitor of histone deacetylase, weakly induced MGMT. However, combined treatment with 5-Aza-dC and trichostatin A significantly up regulated MGMT RNA expression to a greater extent than in cells treated with either agent alone suggesting that histone deacetylation plays a role in MGMT silencing but that CpG methylation has a dominant effect. Consistent with enhanced MGMT expression, 5-Aza-dC increased the association of acetylated histone H3 and H4 bound to the MGMT promoter. Chromatin immunoprecipitation analysis of methyl-CpG binding domain containing proteins detected a greater amount of MeCP2, MBD1, and CAF-1 bound to the MGMT promoter in MGMT-silenced cells. Our findings implicate specific MBD proteins in methylation-mediated transcriptional silencing of MGMT. PMID- 15657355 TI - Covalent binding of antitumor benzoacronycines to double-stranded DNA induces helix opening and the formation of single-stranded DNA: unique consequences of a novel DNA-bonding mechanism. AB - The majority of DNA-binding small molecules known thus far stabilize duplex DNA against heat denaturation. A high, drug-induced increase in the melting temperature (Tm) of DNA is generally viewed as a good criterion to select DNA ligands and is a common feature of several anticancer drugs such as intercalators (e.g., anthracyclines) and alkylators (e.g., ecteinascidin 743). The reverse situation (destabilization of DNA to facilitate its denaturation) may be an attractive option for the identification of therapeutic agents acting on the DNA structure. We have identified the tumor-active benzoacronycine derivative S23906 1 [(+/-)-cis-1,2-diacetoxy-6-methoxy-3,3,14-trimethyl-1,2,3,14-tetrahydro-7H benzo[b]pyrano[3,2]acridin-7-one] as a potent DNA alkylating agent endowed with a helicase-like activity. Using complementary molecular approaches, we show that covalent binding to DNA of the diacetate compound S23906-1 and its monoacetate analogue S28687-1 induces a marked destabilization of the double helix with the formation of alkylated ssDNA. The DNA-bonding properties and effects on DNA structure of a series of benzoacronycine derivatives, including the dicarbamate analogue S29385-1, were studied using complementary biochemical (electromobility shift assay, nuclease S1 mapping) and spectroscopic (fluorescence and Tm measurements) approaches. Alkylation of guanines in DNA by S28687-1 leads to a local denaturation of DNA, which becomes susceptible to cleavage by nuclease S1 and significantly decreases the Tm of DNA. The drug also directly alkylates single-strand DNA, but mass spectrometry experiments indicate that guanines in duplexes are largely preferred over single-stranded structures. This molecular study expands the repertoire of DNA-binding mechanisms and provides a new dimension for DNA recognition by small molecules. PMID- 15657356 TI - Epigallocatechin-3-gallate induces apoptosis in estrogen receptor-negative human breast carcinoma cells via modulation in protein expression of p53 and Bax and caspase-3 activation. AB - Epigallocatechin-3-gallate (EGCG) has been shown to have anticarcinogenic effects in in vitro and in vivo models, and this effect is mediated at least in part by its ability to induce apoptosis in cancer cells without affecting normal cells. It has been recognized that estrogen receptor (ER)-dependent breast cancers generally have a better prognosis and are often responsive to antiestrogen therapy; however, ER-independent breast cancers are more aggressive and unresponsive to antiestrogens. Using the MDA-MB-468 human breast cancer cell line as an in vitro model of ER-negative breast cancers, we found that treatment of EGCG resulted in dose-dependent (5-80 microg/mL) and time-dependent (24-72 hours) inhibition of cellular proliferation (15-100%) and cell viability (3-78%) in MDA MB-468 cells. Decrease in cell viability was associated with the induction of apoptosis (18-66%) which was analyzed by DNA ladder assay, fluorescence staining, and flow cytometry. Induction of apoptosis by EGCG could be corroborated to the increased expression of tumor suppressor protein p53 and its phosphorylation at Ser 15 residue. EGCG decreased the expression of antiapoptotic protein Bcl-2 but increased proapoptotic protein Bax in these cells. The increased ratio of Bax/Bcl 2 proteins after EGCG treatment may have resulted in increased release of cytochrome c from mitochondria into cytosols, increased expression of Apaf-1, and activation of caspase-3 and poly(ADP-ribose) polymerase, which may lead to apoptosis in MDA-MB-468 cells. Together, the results of this study provide evidence that EGCG possesses anticarcinogenic effect against ER-negative breast cancer cells and thus provide the molecular basis for the future development of EGCG as a novel and pharmacologically safe chemopreventive agent for breast cancer prevention. PMID- 15657357 TI - Anti-alpha4 integrin monoclonal antibody inhibits multiple myeloma growth in a murine model. AB - In a syngeneic murine model of multiple myeloma with many of the characteristics of the human disease, a monoclonal antibody (mAb) to the integrin very late antigen-4 (VLA-4), given after the myeloma has already homed to and begun to establish itself within the bone marrow compartment, produces statistically significant effects on multiple disease variables. These include reductions in circulating levels of IgG2b; percentage of IgG2b-positive myeloma cells circulating in blood; spleen weight; and myeloma cell burden in spleen, bone marrow, and liver. mAb therapy had no effect on nonmalignant hematopoietic cells. An acute 6-day regimen of mAb treatment, initiated very late in disease to avoid mAb elimination in the immunocompetent animals, still significantly reduced spleen and blood myeloma cell burden. The ability of the (VLA-4) mAb to affect multiple variables in this model, even as monotherapy, suggests this pathway plays a central role in disease progression. PMID- 15657358 TI - Combination therapy of inhibitors of epidermal growth factor receptor/vascular endothelial growth factor receptor 2 (AEE788) and the mammalian target of rapamycin (RAD001) offers improved glioblastoma tumor growth inhibition. AB - Malignant gliomas are highly lethal tumors that display striking genetic heterogeneity. Novel therapies that inhibit a single molecular target may slow tumor progression, but tumors are likely not dependent on a signal transduction pathway. Rather, malignant gliomas exhibit sustained mitogenesis and cell growth mediated in part through the effects of receptor tyrosine kinases and the mammalian target of rapamycin (mTOR). AEE788 is a novel orally active tyrosine kinase inhibitor that decreases the kinase activity associated with the epidermal growth factor receptor and, at higher concentrations, the vascular endothelial growth factor receptor 2 (kinase domain region). RAD001 (everolimus) is an orally available mTOR inhibitor structurally related to rapamycin. We hypothesized that combined inhibition of upstream epidermal growth factor receptor and kinase domain region receptors with AEE788 and inhibition of the downstream mTOR pathway with RAD001 would result in increased efficacy against gliomas compared with single-agent therapy. In vitro experiments showed that the combination of AEE788 and RAD001 resulted in increased rates of cell cycle arrest and apoptosis and reduced proliferation more than either agent alone. Combined AEE788 and RAD001 given orally to athymic mice bearing established human malignant glioma tumor xenografts resulted in greater tumor growth inhibition and greater increases in median survival than monotherapy. These studies suggest that simultaneous inhibition of growth factor receptor and mTOR pathways offer increased benefit in glioma therapy. PMID- 15657359 TI - Roscovitine-induced up-regulation of p53AIP1 protein precedes the onset of apoptosis in human MCF-7 breast cancer cells. AB - We reported recently that roscovitine arrested human MCF-7 cancer cells at G2-M phase of the cell cycle and concomitantly induced apoptosis. After roscovitine treatment, the level of wild-type p53 protein strongly increased and p53 was accumulated in the nucleus. Here, we raised the question of which pathway would be involved in roscovitine-induced apoptosis in MCF-7 cells, which are known to be caspase-3-deficient, and whether roscovitine-mediated activation of p53 protein might positively affect the execution of cell death. Roscovitine induced a depolarization of mitochondrial potential beginning at 6 hours posttreatment as evidenced by changes in J-aggregate formation and release of the mitochondrial proteins cytochrome c and apoptosis-inducing factor. Interestingly, roscovitine stimulated a site-specific phosphorylation of wild-type p53 protein in a time dependent manner. p53 protein was specifically phosphorylated at Ser46. P-Ser46 activated wild-type p53 tumor suppressor up-regulated p53AIP1 protein, its downstream target known to mediate the depolarization of mitochondria. The onset of phosphorylation of p53 at Ser46 preceded the up-regulation of p53AIP1 protein and the depolarization of mitochondrial potential. We compared the kinetics of roscovitine-mediated p53 activation between caspase-3-deficient parental MCF-7 cells and cells reconstituted with caspase-3. The kinetics and the extent of p53 protein activation in caspase-3-proficient cells differed from those observed in caspase-3-deficient parental cells. Remarkably, roscovitine failed to induce phosphorylation at Ser46 in caspase-3-reconstituted MCF-7 cells. Our results indicate that, depending on the status of caspase-3 in MCF-7 cells, different apoptotic pathways were initialized. PMID- 15657360 TI - Metabolism and pharmacokinetics of the cyclin-dependent kinase inhibitor R roscovitine in the mouse. AB - R-roscovitine (seliciclib, CYC202) is a cyclin-dependent kinase inhibitor currently in phase II clinical trials in patients with cancer. Here, we describe its mouse metabolism and pharmacokinetics as well as the identification of the principal metabolites in hepatic microsomes, plasma, and urine. Following microsomal incubation of R-roscovitine at 10 microg/mL (28 micromol/L) for 60 minutes, 86.7% of the parent drug was metabolized and 60% of this loss was due to formation of one particular metabolite. This was identified as the carboxylic acid resulting from oxidation of the hydroxymethyl group of the amino alcohol substituent at C2 of the purine core present in R-roscovitine. Identification was confirmed by chemical synthesis and comparison of an authentic sample of the R roscovitine-derived carboxylate metabolite (COOH-R-roscovitine). Other minor metabolites were identified as C8-oxo-R-roscovitine and N9-desisopropyl-R roscovitine; these accounted for 4.9% and 2.6% of the parent, respectively. The same metabolic pattern was observed in vivo, with a 4.5-fold lower AUC(infinity) for R-roscovitine (38 micromol/L/h) than for COOH-R-roscovitine (174 micromol/L/h). Excretion of R-roscovitine in the urine up to 24 hours post-dosing accounted for an average of only 0.02% of the administered dose of 50 mg/kg, whereas COOH-R-roscovitine represented 65% to 68% of the dose irrespective of the route of administration (i.v., i.p., or p.o.). A partially deuterated derivative (R-roscovitine-d9) was synthesized to investigate if formation of COOH-R roscovitine could be inhibited by replacement of metabolically labile protons with deuterium. After 60 minutes of incubation of R-roscovitine-d9 or R roscovitine with mouse liver microsomes, formation of COOH-R-roscovitine-d9 was decreased by approximately 24% compared with the production of COOH-R roscovitine. In addition, the levels of R-roscovitine-d9 remaining were 33% higher than those of R-roscovitine. However, formation of several minor R roscovitine metabolites was enhanced with R-roscovitine-d9, suggesting that metabolic switching from the major carbinol oxidation pathway had occurred. Synthetic COOH-R-roscovitine and C8-oxo-R-roscovitine were tested in functional cyclin-dependent kinase assays and shown to be less active than R-roscovitine, confirming that these metabolic reactions are deactivation pathways. PMID- 15657361 TI - Role of phorbol ester localization in determining protein kinase C or RasGRP3 translocation: real-time analysis using fluorescent ligands and proteins. AB - The diacylglycerol signaling pathway, involving protein kinase C (PKC) and RasGRP, is a promising therapeutic target for both cancer and other indications. The phorbol esters, ultrapotent diacylglycerol analogues, bind to and activate PKC and RasGRP. Here, using fluorescent phorbol esters and complementary fluorescent PKC and RasGRP constructs, we determined the localization of the phorbol ester as a function of time after addition and how the resultant PKC or RasGRP3 translocation related to ligand localization. For these studies, we prepared fluorescently labeled phorbol esters of varying lipophilicities based on the BODIPY FL (green) or BODIPY 581/591 (red) fluorophores, and by using fusion constructs of green fluorescent protein or DsRed with PKC isoforms or RasGRP3 expressed in Chinese hamster ovary cells, we simultaneously compared the kinetics and pattern of localization of PKC or RasGRP3 with that of the fluorescent red or green phorbol esters. Binding assays showed that the fluorescent derivatives were potent ligands. Uptake followed a one-compartment pharmacokinetic model with a half-time of minutes to hours, depending on the ligand, and all of the fluorescent phorbol esters localized primarily to intracellular membranes, with little plasma membrane localization. The fluorescent phorbol esters induced translocation of and generally colocalized with PKCdelta or RasGRP3. However, PKCalpha and, initially, PKCdelta, translocated to the plasma membrane, in which little phorbol ester accumulated. The findings argue that the rate of uptake of phorbol esters influences the subsequent pattern of PKCdelta translocation, and that the specificity for PKCalpha translocation is dominated by factors other than the localization of the ligand. PMID- 15657362 TI - Distinct gene expression patterns in a tamoxifen-sensitive human mammary carcinoma xenograft and its tamoxifen-resistant subline MaCa 3366/TAM. AB - The reasons why human mammary tumors become resistant to tamoxifen therapy are mainly unknown. Changes in gene expression may occur as cells acquire resistance to antiestrogens. We therefore undertook a comparative gene expression analysis of tamoxifen-sensitive and tamoxifen-resistant human breast cancer in vivo models using Affymetrix oligonucleotide arrays to analyze differential gene expression. Total RNAs from the tamoxifen-sensitive patient-derived mammary carcinoma xenograft MaCa 3366 and the tamoxifen-resistant model MaCa 3366/TAM were hybridized to Affymetrix HuGeneFL and to Hu95Av2 arrays. Pairwise comparisons and clustering algorithms were applied to identify differentially expressed genes and patterns of gene expression. As revealed by cluster analysis, the tamoxifen sensitive and the tamoxifen-resistant breast carcinomas differed regarding their gene expression pattern. More than 100 transcripts are changed in abundance in MaCa 3366/TAM as compared with MaCa 3366. Among the genes that are differentially expressed in the tamoxifen-resistant tumors, there are several IFN-inducible and estrogen-responsive genes, and genes known to be involved in breast carcinogenesis. The genes neuronatin (NNAT) and bone marrow stem cell antigen 2 (BST2) were sharply up-regulated in MaCa 3366/TAM. The differential expression of four genes (NNAT, BST2, IGFBP5, and BCAS1) was confirmed by Taqman PCR. Our results provide the starting point for deriving markers for tamoxifen resistance by differential gene expression profiling in a human breast cancer model of acquired tamoxifen resistance. Finally, genes whose expression profiles are distinctly changed between the two xenograft lines will be further evaluated as potential targets for diagnostic or therapeutic approaches of tamoxifen-resistant breast cancer. PMID- 15657363 TI - A therapeutic model for advanced endometrial cancer: systemic progestin in combination with local adenoviral-mediated progesterone receptor expression. AB - Cancer of the uterine endometrium is a frequent gynecologic malignant disease for which few therapeutic options are available for advanced disease. Progesterone is the normal female hormone that limits growth and proliferation of endometrial cancers; however, progesterone receptors are frequently down-regulated, leading to treatment failures. The current studies explored the effectiveness of adenoviral-mediated progesterone receptor gene transduction in combination with progestin therapy in mouse xenograft models. Pretreatment of cells with progesterone receptor-encoding adenovirus and progestin inhibited the development of s.c. tumors in athymic mice. In the i.p. xenograft model, replacement of both isoforms of progesterone receptor, PRA and PRB, in combination with progestin treatment resulted in a significant 2.6-fold increase in overall survival time compared with control animals. These studies indicate that when progesterone receptor levels are maintained, progestin therapy is effective in limiting tumor growth. Future therapeutic regimens targeted at enhancing progesterone receptor expression have the potential to improve outcomes in women with endometrial cancer. PMID- 15657364 TI - Carotenoids activate the antioxidant response element transcription system. AB - Epidemiologic studies have found an inverse association between consumption of tomato products and the risk of certain types of cancers. However, the mechanisms underlying this relationship are not completely understood. One mechanism that has been suggested is induction of phase II detoxification enzymes. Expression of phase II enzymes is regulated by the antioxidant response element (ARE) and the transcription factor Nrf2 (nuclear factor E2-related factor 2). In this study, we determined the role of this transcription system in the induction of phase II enzymes by carotenoids. We found that in transiently transfected cancer cells, lycopene transactivated the expression of reporter genes fused with ARE sequences. Other carotenoids such as phytoene, phytofluene, beta-carotene, and astaxanthin had a much smaller effect. An increase in protein as well as mRNA levels of the phase II enzymes NAD(P)H:quinone oxidoreductase and gamma glutamylcysteine synthetase was observed in nontransfected cells after carotenoid treatment. Ethanolic extract of lycopene containing unidentified hydrophilic derivatives of the carotenoid activated ARE with similar potency to lycopene. The potency of the carotenoids in ARE activation did not correlate with their effect on intracellular reactive oxygen species and reduced glutathione level, which may indicate that ARE activation is not solely related to their antioxidant activity. Nrf2, which is found predominantly in the cytoplasm of control cells, translocated to the nucleus after carotenoid treatment. Interestingly, part of the translocated Nrf2 colocalized with the promyelocytic leukemia protein in the promyelocytic leukemia nuclear bodies. The increase in phase II enzymes was abolished by a dominant-negative Nrf2, suggesting that carotenoid induction of these proteins depends on a functional Nrf2 and the ARE transcription system. PMID- 15657366 TI - Benzodiazepines affect channel opening of GABA A receptors induced by either agonist binding site. AB - Benzodiazepines are widely used as anxiolytics, sedatives, muscle relaxants, and anticonvulsants. They allosterically modulate GABA type A (GABA(A)) receptors by increasing the apparent affinity of the agonist GABA to elicit chloride currents. Such an increase in apparent affinity of channel gating could either be caused by an increase in affinity for GABA or by a facilitation of channel opening. In the first case, conformation of the affected sites would have to be altered. In the second case, the affected sites are not necessarily altered, because diazepam could facilitate conformational changes leading to the open channel. It is controversial as to whether benzodiazepines affect only channel opening induced by the occupation of one of the two agonist binding sites or by both. We used receptors formed by concatenated subunits to selectively destroy one of the two agonist sites by point mutation. Both of the receptors harboring only one active agonist site could be stimulated by diazepam. We therefore present evidence that binding of diazepam can affect channel opening induced by either agonist binding site. PMID- 15657365 TI - Cytoplasmic confinement of breast cancer resistance protein (BCRP/ABCG2) as a novel mechanism of adaptation to short-term folate deprivation. AB - The unique capability of breast cancer resistance protein (BCRP/ABCG2) to export mono-, di-, and triglutamates of folates should limit cellular proliferation under conditions of folate deprivation, particularly upon BCRP overexpression. Here, we explored the mode of adaptation of BCRP-overexpressing cells to short term folate deprivation. MCF-7/MR cells grown in high folate medium (2.3 muM folic acid) containing mitoxantrone had 62% of their overexpressed BCRP in the plasma membrane and only 38% in the cytoplasm. In contrast, cells grown for 2 weeks in folic acid-free medium followed by an adaptation week in low folate medium (1 nM folic acid) had 86% of BCRP in the cytoplasm and only 14% in the plasma membrane. Unlike BCRP, various transmembrane proteins retained their normal plasma membrane localization in folate-deprived cells. Folate deprivation was also associated with a 3-fold decrease in BCRP and multidrug resistance protein 1 (MRP1/ABCC1) levels. Confocal microscopy with folate-deprived cells revealed that cytoplasmic BCRP colocalized with calnexin, an established endoplasmic reticulum resident. The loss of BCRP from the plasma membrane in folate-deprived cells consistently resulted in a 4.5-fold increase in [(3)H]folic acid accumulation relative to MCF-7/MR cells. Hence, cellular adaptation to shortterm folate deprivation results in a selective confinement of BCRP to the cytoplasm along with a moderate decrease in BCRP and MRP1 levels aimed at preserving the poor intracellular folate pools. These results constitute a novel mechanism of cellular adaptation to short-term folate deprivation and provide further support to the possible role of BCRP in the maintenance of cellular folate homeostasis. PMID- 15657368 TI - Hyperghrelinemia does not accelerate gastric emptying in Prader-Willi syndrome patients. AB - Prader-Willi syndrome (PWS) is the most common form of syndromic obesity associated with hyperphagia. Because ghrelin stimulates gastric motility in rodents, and PWS patients have 3- to 4-fold higher fasting plasma ghrelin concentrations than normal subjects, we hypothesized that hyperphagia associated with PWS may be partly explained by rapid gastric emptying due to the increased gastric motility caused by ghrelin. We determined gastric emptying times (GETs) and measured ghrelin levels in 11 PWS children and 11 age-, sex-, and body mass index-matched controls using a standard meal containing [(99m)Tc]diaminetriaminepentacetate. Median plasma ghrelin levels before (precibum) and after the GET study were higher in PWS patients than in controls (P = 0.004 and P = 0.001, respectively). Median percent gastric retentions at 90 min after the standard meal were 57.1% (range, 34.0-83.2%) in PWS patients and 40.2% (range, 27.2-60.2%) in controls (P = 0.03). In particular, precibum ghrelin concentrations were not significantly correlated with the rate of gastric emptying in PWS patients (P = 0.153; r = 0.461) or controls (P = 0.911; r = 0.048). Our results show that gastric emptying in PWS is reduced despite higher ghrelin levels, and that the voracious appetite associated with PWS is related to another mechanism. PMID- 15657367 TI - Regulation of the Cyp2a5 gene involves an aryl hydrocarbon receptor-dependent pathway. AB - We have investigated the role of the aryl hydrocarbon receptor (AHR) in the regulation of the Cyp2a5 gene. The C57BL/6 and DBA/2 mouse strains with a genetically determined difference in AHR function were used to study the CYP2A5 induction by typical AHR ligands, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) and 3-methylcholanthrene. The CYP2A5 mRNA up-regulation in these mouse strains showed a difference in response, typical for AHR-regulated genes, both by TCDD in cultured primary hepatocytes and by 3-methylcholanthrene in vivo. In primary hepatocytes, TCDD caused a 3-fold elevation of the CYP2A5 protein level and a similar induction of the CYP2A5-catalyzed coumarin 7-hydroxylation activity. In reporter gene assays, the Cyp2a5 promoter region -3033 to +10 mediated a 2- to 5 fold induction of luciferase activity by TCDD treatment in primary hepatocytes and in Hepa-1 hepatoma cells with an intact AHR/AHR nuclear translocator (ARNT) complex. In Hepa-1 variant cell lines with deficiencies in the AHR/ARNT complex, the absence of ARNT abolished the induction. A putative AHR response element (XRE) was identified in the Cyp2a5 promoter at the position -2514 to -2492 and found to interact with the AHR/ARNT heterodimer. Transfection experiments combined with mutation of the XRE site indicated that the site partly mediates the TCDD induction of Cyp2a5. An additional AHR-dependent mechanism also regulates the proximal promoter of the Cyp2a5 gene. In conclusion, our studies showed that AHR ligands up-regulate Cyp2a5 transcriptionally by an AHR/ARNT dependent mechanism and established Cyp2a5 as a novel AHR-regulated gene. PMID- 15657369 TI - Characterization of ghrelin-like immunoreactivity in human plasma. AB - Ghrelin is a gastric peptide hormone that has an important role in appetite control and GH release. Circulating ghrelin levels are inversely correlated with body mass index and postprandially suppressed. However, the molecular form of circulating ghrelin has not been fully characterized. We studied circulating ghrelin-like immunoreactivity (Ghr-LI) using three RIAs: one specific for only the active, acylated ghrelin (antibody G0-1) and the other two detecting both active and inactive, des-acylated ghrelin (antibody SC and the commercially available Phoenix Pharmaceuticals assay). Plasma ghrelin levels were measured in healthy subjects after a test breakfast (n = 8). Ghr-LI detected by SC and the commercial assay fell significantly at 90 and 120 min post meal (P < 0.01). G0-1 Ghr-LI decreased significantly at 30 min (P < 0.05) post meal and had returned to basal levels at 90 min. Gel permeation chromatography identified three Ghr-LI peaks in plasma. Two G0-1 Ghr-LI peaks with a molecular weight much larger than ghrelin peptide were detected. Only one Ghr-LI peak was detected by the SC and commercial RIA, at the same elution position as synthetic des-acylated ghrelin. These results suggest that the majority of circulating acylated ghrelin is bound to larger molecules, whereas des-acylated ghrelin circulates as free peptide. Assays measuring specific forms of ghrelin may be more useful in determining its physiological role than those that detect both acylated and des-acylated forms. PMID- 15657370 TI - Efficacy and safety of pravastatin in children and adolescents with heterozygous familial hypercholesterolemia: a prospective clinical follow-up study. AB - Heterozygous familial hypercholesterolemia (HeFH) is associated with elevated cholesterol levels and early-onset atherosclerosis. We assessed the efficacy and safety for up to 2 yr of pravastatin treatment in 19 girls and 11 boys (age range, 4.1-18.5 yr) with HeFH. Pravastatin was started at 10 mg/d, with a forced titration by 10 mg at 2, 4, 6, and 12 months until the target cholesterol level [< or =194 mg/dl (< or =5 mmol/liter)] was reached. By 2, 4, 6, 12, and 24 months of treatment, the total cholesterol levels had, respectively, decreased by 19, 20, 23, 27, and 26%, and the low-density lipoprotein cholesterol levels had decreased by 25, 27, 29, 33, and 32% compared with the dietary baseline values. Seventeen percent of patients had lipid deposits (carotid plaque, xanthomas, or corneal arcus) at baseline, and 27% had deposits at 1 yr. The side effects were mild, and no clinically significant elevations in alanine aminotransferase, creatine kinase, or creatinine were seen. Growth and pubertal maturation remained normal in all subjects. In conclusion, pravastatin treatment was safe and well tolerated. The efficacy in children with slight or moderate hypercholesterolemia was satisfactory, but in children with severe hypercholesterolemia, it was insufficient. PMID- 15657371 TI - Expression of human prostaglandin transporter in the human endometrium across the menstrual cycle. AB - Prostaglandins (PGs) are important regulators of reproductive function. The mechanism by which PGs are transported across the biological membrane is a new emerging field of investigation. Prostaglandin transporter (PGT) has been identified as a functional PG carrier. The aim of our study was to outline the expression of PGT in the human endometrium across the menstrual cycle. Quantitative RT-PCR showed human PGT (hPGT) expression to be strong in the proliferative and early secretory phases and low in the middle to late secretory phase. Northern blot analysis revealed hPGT mRNA transcript of 4 kb in the human endometrium. A peptide-directed polyclonal antibody was generated in rabbits against the 22 amino acids forming the C terminus of hPGT. Antibody specificity was demonstrated by Western blot. Immunoblots of endogenous hPGT in the human endometrium revealed a 70-kDa protein in endometrial cells. Endometrial biopsies collected across the menstrual cycle were used to assess hPGT protein expression by immunohistochemistry. hPGT was immunolocalized to luminal, glandular epithelial, and stromal cells. Because it was observed at the mRNA level, semiquantitative analysis showed a higher protein expression in proliferative and early secretory phases than in the mid-late secretory phase. In conclusion, our study revealed that hPGT expression is modulated in epithelial and stromal cells of the human endometrium at both mRNA and protein levels during the menstrual cycle. These findings support a role for hPGT as an important new player in the regulation of PG action in the human endometrium. PMID- 15657372 TI - Dietary soy containing phytoestrogens does not activate the hemostatic system in postmenopausal women. AB - The soybean is rich in isoflavone phytoestrogens, which are ligands for estrogen receptors, but it is unknown whether soy/phytoestrogens have similar procoagulant effects to estrogen. In this randomized double-blind trial, 40 healthy postmenopausal women of age 50-75 yr received soy protein isolate (40 g soy protein, 118 mg isoflavones) (n = 19) or casein placebo (n = 21). Plasma markers of coagulation, fibrinolysis, and endothelial dysfunction were measured at baseline and 3 months. The baseline characteristics of the two groups were similar. Compared with casein placebo, soy decreased triglycerides (P < 0.005) and low-density lipoprotein/high-density lipoprotein ratio (P < 0.001) and increased lipoprotein (a) (P < 0.05). Activity of coagulation factor VII (VIIc) decreased similarly in both groups (P < 0.005). Prothrombin fragments 1 + 2 (a marker of thrombin generation) decreased in the soy group (P < 0.005), but the change was not different from the casein group. There was no effect of soy on soluble fibrin (a marker of fibrin production), plasminogen activator inhibitor-1 (a marker of fibrinolytic inhibitory potential), D-dimer (a marker of fibrin turnover), or von Willebrand factor (a marker of endothelial damage). In conclusion, the results of the current study do not support biologically significant estrogenic effects of soy/phytoestrogens on coagulation, fibrinolysis, or endothelial function. PMID- 15657373 TI - Ghrelin, peptide YY, glucose-dependent insulinotropic polypeptide, and hunger responses to a mixed meal in anorexic, obese, and control female adolescents. AB - To determine whether peptide YY (PYY), ghrelin, glucose-dependent insulinotropic polypeptide (GIP), and satiety responses to food intake are impaired in anorexia or obesity, we studied 30 female adolescents with anorexia nervosa [body mass index (BMI) 16.3 kg/m2], obesity (BMI 34.3 kg/m2), or normal weight (BMI 20.2 kg/m2). PYY, ghrelin, GIP, insulin, and glucose concentrations and four markers of satiety were measured for 240 min after a mixed meal. The area under the curve for glucose was similar in obese (OB) and normal-weight control (C) subjects but was 15% lower in anorexic (AN) subjects. The area under the curve for insulin was 47% lower in AN and 87% higher in OB subjects, compared with C subjects. After the meal, PYY increased significantly in C (+41%, P < 0.05) but not in AN or OB adolescents. Ghrelin concentrations were highest in AN subjects and lowest in the OB group, compared with C subjects and fell significantly by 25% in all three groups. GIP concentrations were lower in AN subjects throughout the test and increased in all three groups after the mixed meal. AN adolescents reported being less hungry than OB and C adolescents. There was a negative correlation between fasting ghrelin (but not PYY or GIP) and BMI and insulin (r2= 0.33) and a positive correlation between the decrease in hunger 15 min after the meal and PYY concentrations at 15 min (r2= 0.20). In conclusion, the blunted PYY response to a meal in OB adolescents suggests that PYY plays a role in the pathophysiology of obesity. Ghrelin is unlikely to play a causal role in anorexia nervosa or obesity. The lower GIP observed in AN subjects despite a similar caloric intake may appropriately prevent an excessive insulin response in these patients. PMID- 15657374 TI - Expression and localization of adipophilin and perilipin in human fetal membranes: association with lipid bodies and enzymes involved in prostaglandin synthesis. AB - Lipid storage droplets (LSDs) are subcellular storage depots for triglycerides (TGs) and cholesterol esters surrounded by specific populations of proteins that are necessary for their formation. We have previously described the appearance of LSDs in human fetal membranes with advancing gestation and labor. Perilipin and adipophilin are functional/structural proteins located on the surfaces of intracellular LSDs. Adipophilin and perilipin were both immunolocalized to the amnion epithelium and amnion fibroblasts in human fetal membranes. Adipophilin was also localized to the choriodecidual layer, whereas perilipin was localized to the chorion trophoblasts. Although immunohistochemical data show an apparent increase in adipophilin, but not perilipin, expression in fetal membranes with advancing gestation and labor, Western analysis of tissue homogenate supernatant revealed no significant changes in adipophilin and perilipin expression. However, Western analysis of the floating lipid-rich layer from the tissue homogenate revealed an abundance of adipophilin and perilipin as well as other enzymes (cytosolic phospholipase A2, prostaglandin endoperoxide, and microsomal associated prostaglandin E synthase-1) involved in prostaglandin synthesis. The association of these enzymatically active proteins with LSDs suggests that LSDs may be foci for signaling via the arachidonic acid cascade in fetal membranes. The structural and functional roles of adipophilin and perilipin in gestation and labor remain to be determined. PMID- 15657375 TI - Elevated serum interferon-gamma-inducible chemokine-10/CXC chemokine ligand-10 in autoimmune primary adrenal insufficiency and in vitro expression in human adrenal cells primary cultures after stimulation with proinflammatory cytokines. AB - Chemokines are a large family of cytokines involved in the pathogenesis of inflammatory and autoimmune diseases. Among CXC chemokines, CXC chemokine ligand 10 (CXCL10) has been identified to play an important role in several endocrinological autoimmune diseases, such as Hashimoto's thyroiditis, Graves' disease, and type 1 diabetes mellitus. Although the mechanisms leading to glandular autoimmune process may be at least in part shared by different endocrine organs, the role of CXCL10 in autoimmune adrenal insufficiency is unknown. The aim of this study was to evaluate the role of CXCL10 in Addison's disease (AD). Serum CXCL10 levels were assayed in 64 patients with clinically evident autoimmune AD, 20 patients with autoimmune subclinical AD, nine patients with nonautoimmune AD, and 48 healthy volunteers. Clinically evident and subclinical AD, but not nonautoimmune AD patients, showed a significant increase in serum CXCL10 levels compared with healthy subjects: 119.9 pg/ml (range, 39.8 427.6) and 124.0 pg/ml (range, 37.0-384.7) vs. 75.6 pg/ml (range, 22.4-164.0; P < 0.001 for both groups). Comparable serum CXCL10 levels were found between patients with an isolated form of AD and patients with other autoimmune conditions associated with AD, suggesting a specific influence of the adrenal autoimmune process in determining elevated CXCL10 concentrations in such patients. No relationship was found between serum CXCL10 levels and anti-21 hydroxylase or adrenal cortex autoantibody titers or between CXCL10 levels and duration of disease. The role of CXCL10 in the adrenal gland was also evaluated in vitro in human zona fasciculata cells (hZFC). CXCL10, although not basally detected in cultured hZFC, was strongly induced by interferon-gamma and synergistically increased by TNF-alpha addition. Hydrocortisone or ACTH alone had no effect on CXCL10 secretion in hZFC, but they both significantly inhibited cytokine-induced CXCL10 secretion. Taken together, these data suggest a potential role of hZFC, through the production of CXCL10, in regulating the recruitment of specific subsets of activated lymphocytes in autoimmune AD. PMID- 15657376 TI - Hypogonadism in hereditary hemochromatosis. AB - Hypogonadism, usually hypogonadotropic in origin, is the most common nondiabetic endocrinopathy in hereditary hemochromatosis (HH). Early studies, usually evaluating small numbers of patients with advanced HH, report prevalence rates of 10-100%. The clinical presentation of HH has changed in recent years as a result of increased awareness and screening. We assessed the prevalence of hypogonadism in a large group of patients with HH diagnosed in a single center over the past 20 yr, the period of follow-up spanning the time before and after widespread screening was introduced and the HFE gene was recognized. Abnormally low plasma testosterone levels, with low LH and FSH levels, were found in nine of 141 (6.4%) male patients tested. Eight of nine (89%) had associated hepatic cirrhosis; three of nine (33%) had diabetes. Inappropriately low LH and FSH levels were found in two of 38 females (5.2%) in whom the pituitary-gonadal axis could be assessed. This is the largest detailed study of hypogonadism reported in HH. The lower prevalence of hypogonadism compared with other reported series reflects the earlier diagnosis of HH in an unselected group of patients attending a single center. Patients with lesser degrees of hepatic siderosis at diagnosis are unlikely to develop hypogonadism. PMID- 15657377 TI - Cardiology patient pages. Red wine and your heart. PMID- 15657378 TI - Images in cardiovascular medicine. Singular coronary artery aneurysm: imaging with coronary angiography versus 16-slice computed tomography, transesophageal echocardiography, and magnetic resonance tomography. PMID- 15657379 TI - Images in cardiovascular medicine. False aneurysm of the ascending aorta caused by a sternal wire. PMID- 15657380 TI - Letters regarding article by Bibbins-Domingo et al, "B-type natriuretic peptide and ischemia in patients with stable coronary disease: data from the Heart and Soul study". PMID- 15657382 TI - Letter regarding article by Abbate et al, "Widespread myocardial inflammation and infarct-related artery patency". PMID- 15657383 TI - Letter regarding article by Gazzaruso et al, "Relationship between erectile dysfunction and silent myocardial ischemia in apparently uncomplicated, type 2 diabetic patients". PMID- 15657384 TI - We think we see a pattern emerging here. PMID- 15657385 TI - Cardiovascular magnetic resonance in cardiac amyloidosis. PMID- 15657386 TI - Optimizing percutaneous coronary intervention outcomes: the next steps. PMID- 15657387 TI - Drug therapy in the heart transplant recipient: part IV: drug-drug interactions. PMID- 15657388 TI - Role of permanent pacing to prevent atrial fibrillation: science advisory from the American Heart Association Council on Clinical Cardiology (Subcommittee on Electrocardiography and Arrhythmias) and the Quality of Care and Outcomes Research Interdisciplinary Working Group, in collaboration with the Heart Rhythm Society. AB - This advisory summarizes the current database on pacing modalities and algorithms used to prevent and terminate atrial fibrillation (AF). On the basis of the evidence indicating that ventricular pacing is associated with a higher incidence of AF in patients with sinus node dysfunction, a patient who has a history of AF and needs a pacemaker for bradycardia should receive a physiological pacemaker (dual chamber or atrial) rather than a single-chamber ventricular pacemaker. For patients who need a dual-chamber pacemaker, efforts should be made to program the device to minimize the amount of ventricular pacing when atrioventricular conduction is intact. Many pacemakers and implantable defibrillators have features designed to prevent AF and to terminate AF with rapid atrial pacing. The evidence to support their use is limited, although these algorithms appear to be safe and usually add little additional cost. For patients who have a bradycardia indication for pacing and also have AF, no consistent data from large randomized trials support the use of alternative single-site atrial pacing, multisite right atrial pacing, biatrial pacing, overdrive pacing, or antitachycardia atrial pacing. Even fewer data support the use of atrial pacing in the management of AF in patients without symptomatic bradycardia. At present, permanent pacing to prevent AF is not indicated; however, additional studies are ongoing, which will help to clarify the role of permanent pacing for AF. PMID- 15657389 TI - Going in GTP cycles in the nucleolus. AB - Proteins are directed to cellular compartments by specific localization signals. A GTP-driven cycle has now been identified as a mechanism for protein targeting to the nucleolus. The involvement of a GTP switch suggests that nucleolar localization can be regulated and may be responsive to extracellular stimuli via signaling pathways. The uncovered mechanism also implies that localization is determined by increased retention rather than directed targeting. PMID- 15657390 TI - A multistep, GTP-driven mechanism controlling the dynamic cycling of nucleostemin. AB - Nucleostemin (NS) was identified as a stem cell- and cancer cell-enriched nucleolar protein that controls the proliferation of these cells. Here, we report the mechanism that regulates its dynamic shuttling between the nucleolus and nucleoplasm. The nucleolar residence of nucleostemin involves a transient and a long-term binding by the basic and GTP-binding domains, and a dissociation mechanism mediated by the COOH-terminal region. This cycle is propelled by the GTP binding state of nucleostemin. We propose that a rapid nucleostemin cycle is designed to translate extra- and intra-cellular signals into the amount of nucleostemin in the nucleolus in a bidirectional and fast manner. PMID- 15657391 TI - Cell growth-dependent coordination of lipid signaling and glycosylation is mediated by interactions between Sac1p and Dpm1p. AB - The integral membrane lipid phosphatase Sac1p regulates local pools of phosphatidylinositol-4-phosphate (PtdIns(4)P) at endoplasmic reticulum (ER) and Golgi membranes. PtdIns(4)P is important for Golgi trafficking, yet the significance of PtdIns(4)P for ER function is unknown. It also remains unknown how localization of Sac1p to distinct organellar membranes is mediated. Here, we show that a COOH-terminal region in yeast Sac1p is crucial for ER targeting by directly interacting with dolicholphosphate mannose synthase Dpm1p. The interaction with Dpm1p persists during exponential cell division but is rapidly abolished when cell growth slows because of nutrient limitation, causing translocation of Sac1p to Golgi membranes. Cell growth-dependent shuttling of Sac1p between the ER and the Golgi is important for reciprocal control of PtdIns(4)P levels at these organelles. The fraction of Sac1p resident at the ER is also required for efficient dolichol oligosaccharide biosynthesis. Thus, the lipid phosphatase Sac1p may be a key regulator, coordinating the secretory capacity of ER and Golgi membranes in response to growth conditions. PMID- 15657392 TI - Muscle expression of a local Igf-1 isoform protects motor neurons in an ALS mouse model. AB - Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease characterized by a selective degeneration of motor neurons, atrophy, and paralysis of skeletal muscle. Although a significant proportion of familial ALS results from a toxic gain of function associated with dominant SOD1 mutations, the etiology of the disease and its specific cellular origins have remained difficult to define. Here, we show that muscle-restricted expression of a localized insulin-like growth factor (Igf) -1 isoform maintained muscle integrity and enhanced satellite cell activity in SOD1(G93A) transgenic mice, inducing calcineurin-mediated regenerative pathways. Muscle-specific expression of local Igf-1 (mIgf-1) isoform also stabilized neuromuscular junctions, reduced inflammation in the spinal cord, and enhanced motor neuronal survival in SOD1(G93A) mice, delaying the onset and progression of the disease. These studies establish skeletal muscle as a primary target for the dominant action of inherited SOD1 mutation and suggest that muscle fibers provide appropriate factors, such as mIgf-1, for neuron survival. PMID- 15657393 TI - Spindle-independent condensation-mediated segregation of yeast ribosomal DNA in late anaphase. AB - Mitotic cell division involves the equal segregation of all chromosomes during anaphase. The presence of ribosomal DNA (rDNA) repeats on the right arm of chromosome XII makes it the longest in the budding yeast genome. Previously, we identified a stage during yeast anaphase when rDNA is stretched across the mother and daughter cells. Here, we show that resolution of sister rDNAs is achieved by unzipping of the locus from its centromere-proximal to centromere-distal regions. We then demonstrate that during this stretched stage sister rDNA arrays are neither compacted nor segregated despite being largely resolved from each other. Surprisingly, we find that rDNA segregation after this period no longer requires spindles but instead involves Cdc14-dependent rDNA axial compaction. These results demonstrate that chromosome resolution is not simply a consequence of compacting chromosome arms and that overall rDNA compaction is necessary to mediate the segregation of the long arm of chromosome XII. PMID- 15657394 TI - Nuclear transport of single molecules: dwell times at the nuclear pore complex. AB - The mechanism by which macromolecules are selectively translocated through the nuclear pore complex (NPC) is still essentially unresolved. Single molecule methods can provide unique information on topographic properties and kinetic processes of asynchronous supramolecular assemblies with excellent spatial and time resolution. Here, single-molecule far-field fluorescence microscopy was applied to the NPC of permeabilized cells. The nucleoporin Nup358 could be localized at a distance of 70 nm from POM121-GFP along the NPC axis. Binding sites of NTF2, the transport receptor of RanGDP, were observed in cytoplasmic filaments and central framework, but not nucleoplasmic filaments of the NPC. The dwell times of NTF2 and transportin 1 at their NPC binding sites were 5.8 +/- 0.2 and 7.1 +/- 0.2 ms, respectively. Notably, the dwell times of these receptors were reduced upon binding to a specific transport substrate, suggesting that translocation is accelerated for loaded receptor molecules. Together with the known transport rates, our data suggest that nucleocytoplasmic transport occurs via multiple parallel pathways within single NPCs. PMID- 15657397 TI - Generation of nonidentical compartments in vesicular transport systems. AB - How can organelles communicate by bidirectional vesicle transport and yet maintain different protein compositions? We show by mathematical modeling that a minimal system, in which the basic variables are cytosolic coats for vesicle budding and membrane-bound soluble N-ethyl-maleimide-sensitive factor attachment protein receptors (SNAREs) for vesicle fusion, is sufficient to generate stable, nonidentical compartments. A requirement for establishing and maintaining distinct compartments is that each coat preferentially packages certain SNAREs during vesicle budding. Vesicles fuse preferentially with the compartment that contains the highest concentration of cognate SNAREs, thus further increasing these SNAREs. The stable steady state is the result of a balance between this autocatalytic SNARE accumulation in a compartment and the distribution of SNAREs between compartments by vesicle budding. The resulting nonhomogeneous SNARE distribution generates coat-specific vesicle fluxes that determine the size of compartments. With nonidentical compartments established in this way, the localization and cellular transport of cargo proteins can be explained simply by their affinity for coats. PMID- 15657396 TI - Role of mitochondria in the pheromone- and amiodarone-induced programmed death of yeast. AB - Although programmed cell death (PCD) is extensively studied in multicellular organisms, in recent years it has been shown that a unicellular organism, yeast Saccharomyces cerevisiae, also possesses death program(s). In particular, we have found that a high doses of yeast pheromone is a natural stimulus inducing PCD. Here, we show that the death cascades triggered by pheromone and by a drug amiodarone are very similar. We focused on the role of mitochondria during the pheromone/amiodarone-induced PCD. For the first time, a functional chain of the mitochondria-related events required for a particular case of yeast PCD has been revealed: an enhancement of mitochondrial respiration and of its energy coupling, a strong increase of mitochondrial membrane potential, both events triggered by the rise of cytoplasmic [Ca2+], a burst in generation of reactive oxygen species in center o of the respiratory chain complex III, mitochondrial thread-grain transition, and cytochrome c release from mitochondria. A novel mitochondrial protein required for thread-grain transition is identified. PMID- 15657395 TI - Actin-myosin-based contraction is responsible for apoptotic nuclear disintegration. AB - Membrane blebbing during the apoptotic execution phase results from caspase mediated cleavage and activation of ROCK I. Here, we show that ROCK activity, myosin light chain (MLC) phosphorylation, MLC ATPase activity, and an intact actin cytoskeleton, but not microtubular cytoskeleton, are required for disruption of nuclear integrity during apoptosis. Inhibition of ROCK or MLC ATPase activity, which protect apoptotic nuclear integrity, does not affect caspase-mediated degradation of nuclear proteins such as lamins A, B1, or C. The conditional activation of ROCK I was sufficient to tear apart nuclei in lamin A/C null fibroblasts, but not in wild-type fibroblasts. Thus, apoptotic nuclear disintegration requires actin-myosin contractile force and lamin proteolysis, making apoptosis analogous to, but distinct from, mitosis where nuclear disintegration results from microtubule-based forces and from lamin phosphorylation and depolymerization. PMID- 15657398 TI - ARFGAP1 plays a central role in coupling COPI cargo sorting with vesicle formation. AB - Examining how key components of coat protein I (COPI) transport participate in cargo sorting, we find that, instead of ADP ribosylation factor 1 (ARF1), its GTPase-activating protein (GAP) plays a direct role in promoting the binding of cargo proteins by coatomer (the core COPI complex). Activated ARF1 binds selectively to SNARE cargo proteins, with this binding likely to represent at least a mechanism by which activated ARF1 is stabilized on Golgi membrane to propagate its effector functions. We also find that the GAP catalytic activity plays a critical role in the formation of COPI vesicles from Golgi membrane, in contrast to the prevailing view that this activity antagonizes vesicle formation. Together, these findings indicate that GAP plays a central role in coupling cargo sorting and vesicle formation, with implications for simplifying models to describe how these two processes are coupled during COPI transport. PMID- 15657401 TI - Maximizing therapeutic benefit of rituximab: maintenance therapy versus re treatment at progression in patients with indolent non-Hodgkin's lymphoma--a randomized phase II trial of the Minnie Pearl Cancer Research Network. AB - PURPOSE: To compare the benefit of maintenance rituximab therapy versus rituximab re-treatment at progression in patients with previously treated indolent non Hodgkin's lymphoma. PATIENTS AND METHODS: Between June 1998 and August 2002, 114 patients who had received previous chemotherapy for indolent non-Hodgkin's lymphoma were treated with a standard 4-week course of rituximab. Patients with objective response or stable disease were randomly assigned to receive either maintenance rituximab therapy (standard 4-week courses administered at 6-month intervals) or rituximab re-treatment at the time of lymphoma progression. The duration of rituximab benefit was measured from the date of first rituximab treatment until the date other treatment was required. RESULTS: Ninety (79%) of 114 patients had objective response or stable disease after initial rituximab treatment, and were randomly assigned to treatment. Progression-free survival was prolonged in the maintenance group (31.3 v 7.4 months; P = .007). Final overall and complete response rates were higher in the maintenance group. Duration of rituximab benefit was similar in the maintenance and re-treatment groups (31.3 v 27.4 months, respectively). More maintenance patients remain in continuous remission, and more are currently in complete remission. Both treatment approaches were well tolerated. CONCLUSION: In patients who have objective response or stable disease with single-agent rituximab therapy, duration of rituximab benefit is substantially prolonged with either scheduled maintenance treatment or rituximab re-treatment at the time of progression. At present, the magnitude of benefit with either approach appears similar. However, additional follow-up of this trial is required, and completion of phase III randomized trials is necessary to definitively answer this question. PMID- 15657399 TI - Effects of Arp2 and Arp3 nucleotide-binding pocket mutations on Arp2/3 complex function. AB - Contributions of actin-related proteins (Arp) 2 and 3 nucleotide state to Arp2/3 complex function were tested using nucleotide-binding pocket (NBP) mutants in Saccharomyces cerevisiae. ATP binding by Arp2 and Arp3 was required for full Arp2/3 complex nucleation activity in vitro. Analysis of actin dynamics and endocytosis in mutants demonstrated that nucleotide-bound Arp3 is particularly important for Arp2/3 complex function in vivo. Severity of endocytic defects did not correlate with effects on in vitro nucleation activity, suggesting that a critical Arp2/3 complex function during endocytosis may be structural rather than catalytic. A separate class of Arp2 and Arp3 NBP mutants suppressed phenotypes of mutants defective for actin nucleation. An Arp2 suppressor mutant increased Arp2/3 nucleation activity. Electron microscopy of Arp2/3 complex containing this Arp2 suppressor identified a structural change that also occurs upon Arp2/3 activation by nucleation promoting factors. These data demonstrate the importance of Arp2 and Arp3 nucleotide binding for nucleating activity, and Arp3 nucleotide binding for maintenance of cortical actin cytoskeleton cytoarchitecture. PMID- 15657400 TI - A role for myosin VI in postsynaptic structure and glutamate receptor endocytosis. AB - Myosin VI (Myo6) is an actin-based motor protein implicated in clathrin-mediated endocytosis in nonneuronal cells, though little is known about its function in the nervous system. Here, we find that Myo6 is highly expressed throughout the brain, localized to synapses, and enriched at the postsynaptic density. Myo6 deficient (Snell's waltzer; sv/sv) hippocampus exhibits a decrease in synapse number, abnormally short dendritic spines, and profound astrogliosis. Similarly, cultured sv/sv hippocampal neurons display decreased numbers of synapses and dendritic spines, and dominant-negative disruption of Myo6 in wild-type hippocampal neurons induces synapse loss. Importantly, we find that sv/sv hippocampal neurons display a significant deficit in the stimulation-induced internalization of alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid-type glutamate receptors (AMPARs), and that Myo6 exists in a complex with the AMPAR, AP-2, and SAP97 in brain. These results suggest that Myo6 plays a role in the clathrin-mediated endocytosis of AMPARs, and that its loss leads to alterations in synaptic structure and astrogliosis. PMID- 15657402 TI - Phase II trial of individualized rituximab dosing for patients with CD20-positive lymphoproliferative disorders. AB - PURPOSE: To determine the feasibility and efficacy of pharmacokinetic (PK) -based maintenance dosing of rituximab and possibly design a more rational maintenance schedule. PATIENTS AND METHODS: Patients with CD20-positive lymphoproliferative disorders were treated with four weekly infusions of rituximab 375 mg/m(2). All patients without progressive disease were then monitored for 1 year and received a single infusion of 375 mg/m(2) when the level decreased below 25 microg/mL. RESULTS: Twenty-nine of 31 patients were assessable with a variety of histologic subtypes. The overall response rate (ORR) for the entire group was 59% with 27% complete responses (CRs) and 32% partial responses. The median PFS for all patients was 19 months, with a median follow-up of 25 months. In 22 patients with low-grade non-Hodgkin's lymphoma (LGNHL), the ORR was 63% with 36% CR and median progression-free survival (PFS) has not been reached. Of 29 assessable patients, 22 were available for PK-based maintenance. The median time to repeat bolus was 5 months (range, 1 to 9 months) for the first, 3.5 months (range, 2 to 5 months) for the second, and 3 months (range, 2 to 4 months) for the third infusion. Ninety-five percent of patients required three or fewer infusions to be maintained in the therapeutic range. CONCLUSION: Individualized PK dosing for rituximab produced efficacy comparable to other published maintenance strategies. PK data from this trial suggest that a rational maintenance strategy in patients with LGNHL would be a single dose of 375 mg/m(2) of rituximab every 3 to 4 months. PMID- 15657403 TI - Long-term survival after cisplatin-based induction chemotherapy and radiotherapy for nasopharyngeal carcinoma: a pooled data analysis of two phase III trials. AB - PURPOSE: To evaluate the long-term outcome in patients with nasopharyngeal carcinoma (NPC) treated with induction chemotherapy and radiotherapy (CRT) versus radiotherapy alone (RT). PATIENTS AND METHODS: The data from two phase III studies comparing CRT with RT in NPC were updated and pooled together for analysis. A total of 784 patients were included for analysis, with an equal number of patients in both arms. Induction chemotherapy consisted of two to three cycles of cisplatin, bleomycin, and fluorouracil, or cisplatin and epirubicin. RT was given to the nasopharynx and neck using megavoltage radiation (median dose, 70 Gy). The median follow-up time for surviving patients was 67 months. Analysis was based on intention to treat. RESULTS: The addition of induction chemotherapy to RT was associated with a decrease in relapse by 14.3% and cancer-related deaths by 12.9% at 5 years. The 5-year relapse-free survival rate was 50.9% and 42.7% in the CRT and RT arm, respectively (P = .014), and the 5-year disease specific survival rate was 63.5% and 58.1% in the CRT and RT arm, respectively (P = .029). The 5-year overall survival rate was 61.9% and 58.1% in CRT and RT arm, respectively (P = .092). The incidence of locoregional failure and distant metastases was reduced by 18.3% and 13.3% at 5 years, respectively, with induction chemotherapy. There was no significant difference in the treatment failure patterns between the two arms. CONCLUSION: The addition of cisplatin based induction chemotherapy to RT was associated with a modest but significant decrease in relapse and improvement in disease-specific survival in advanced stage NPC. However, there was no improvement in overall survival. PMID- 15657404 TI - Rituximab therapy for patients with newly diagnosed, advanced-stage, follicular grade I non-Hodgkin's lymphoma: a phase II trial in the North Central Cancer Treatment Group. AB - PURPOSE: Patients with newly diagnosed, advanced-stage, follicular grade 1 non Hodgkin's lymphoma (NHL) are often asymptomatic and can be observed without immediate chemotherapy. The goals of this study were to assess the overall response rate (ORR) to rituximab in this patient population and to determine the time-to-progression (TTP) and time-to-subsequent-chemotherapy (TTSC). PATIENTS AND METHODS: Eligible patients had untreated follicular grade 1 NHL, and measurable stage III/IV disease. Patients received rituximab 375 mg/m(2) intravenous weekly x 4 doses and were then followed for response and progression; no maintenance therapy was provided. RESULTS: Thirty-seven patients were accrued; one patient was ineligible. The median age was 59 years (range, 29 to 83 years). Six patients (18%) had elevated lactate dehydrogenase levels. The ORR was 72%, with 36% complete remissions. Fourteen (39%) of 36 patients remain in unmaintained remission, two died without disease progression, and three died with disease progression. Twenty (56%) of 36 patients have disease progression. The median TTP was 2.2 years (95% CI, 1.3 to not yet reached). Eighteen patients have subsequently been treated with chemotherapy, with a median TTSC of 2.3 years (95% CI, 1.6 to not yet reached). Patients with a high lactate dehydrogenase level had a lower ORR of 33% and a short TTP of only 6 months. CONCLUSION: Rituximab can be safely administered to patients with advanced-stage follicular grade 1 NHL with efficacy and minimal toxicity. This therapy is highly active and offers an acceptable alternative to observation in this patient population. Patients with high LDH should not be considered for rituximab monotherapy. PMID- 15657406 TI - Role of cytochrome P450 phenotyping in cancer treatment. PMID- 15657405 TI - Randomized pharmacokinetic and pharmacodynamic study of docetaxel: dosing based on body-surface area compared with individualized dosing based on cytochrome P450 activity estimated using a urinary metabolite of exogenous cortisol. AB - PURPOSE: Docetaxel is metabolized by cytochrome P450 (CYP3A4) enzyme, and the area under the concentration-time curve (AUC) is correlated with neutropenia. We developed a novel method for estimating the interpatient variability of CYP3A4 activity by the urinary metabolite of exogenous cortisol (6-beta-hydroxycortisol [6-beta-OHF]). This study was designed to assess whether the application of our method to individualized dosing could decrease pharmacokinetic (PK) and pharmacodynamic (PD) variability compared with body-surface area (BSA) -based dosing. PATIENTS AND METHODS: Fifty-nine patients with advanced non-small-cell lung cancer were randomly assigned to either the BSA-based arm or individualized arm. In the BSA-based arm, 60 mg/m(2) of docetaxel was administered. In the individualized arm, individualized doses of docetaxel were calculated from the estimated clearance (estimated clearance = 31.177 + [7.655 x 10(-4) x total 6 beta-OHF] - [4.02 x alpha-1 acid glycoprotein] - [0.172 x AST] - [0.125 x age]) and the target AUC of 2.66 mg/L . h. RESULTS: In the individualized arm, individualized doses of docetaxel ranged from 37.4 to 76.4 mg/m(2) (mean, 58.1 mg/m(2)). The mean AUC and standard deviation (SD) were 2.71 (range, 2.02 to 3.40 mg/L . h) and 0.40 mg/L . h in the BSA-based arm, and 2.64 (range, 2.15 to 3.07 mg/L . h) and 0.22 mg/L . h in the individualized arm, respectively. The SD of the AUC was significantly smaller in the individualized arm than in the BSA-based arm (P < .01). The percentage decrease in absolute neutrophil count (ANC) averaged 87.1% (range, 59.0 to 97.7%; SD, 8.7) in the BSA-based arm, and 87.4% (range, 78.0 to 97.2%; SD, 6.1) in the individualized arm, suggesting that the interpatient variability in percent decrease in ANC was slightly smaller in the individualized arm. CONCLUSION: The individualized dosing method based on the total amount of urinary 6-beta-OHF after cortisol administration can decrease PK variability of docetaxel. PMID- 15657407 TI - Is there still a role for induction chemotherapy for head and neck cancer? PMID- 15657408 TI - Optimizing rituximab in B-cell lymphoma. PMID- 15657409 TI - Changing HIV and AIDS-related behavior: promising approaches at the individual, group, and community levels. PMID- 15657411 TI - Group intervention to reduce HIV transmission risk behavior among persons living with HIV/AIDS. AB - Results of a randomized controlled trial show that a behavioral intervention grounded in social cognitive theory reduces unprotected sexual behaviors among men and women living with HIV infection, with the greatest reductions in HIV transmission risk behaviors occurring with non-HIV-positive sex partners. In this article, the authors describe the intervention development and intervention content of the social cognitive risk reduction intervention for HIV-positive persons. The effective five group session intervention focused on enhancing motivation through self-reflection and developing coping efficacy skills for HIV disclosure decision making, active listening, assertiveness, and problem solving for disclosure and transmission risk reduction behaviors. Intervention components were tailored for gender and sexual orientation and integrated skills practice sessions used role-plays couched within scenes from popular films. This intervention was demonstrated to be effective in a community-service delivery setting and can be adapted for implementation in HIV-related services delivered within support groups. PMID- 15657410 TI - Prevention for HIV-seropositive persons: successive approximation toward a new identity. AB - This article presents a three-module intervention based on social action theory that focuses on health promotion and social identity formation for seropositive youth. The modules are designed to reduce transmission of HIV by reducing sexual and substance abuse acts, increasing healthy acts and adherence to care, and maintaining positive behavioral routines. Components of the modules are described, including examples of how these components are implemented in the actual intervention sessions. The importance of using successive approximation to consolidate changes in behavior by defining social roles and personal identities that are consistent with positive behavioral routines is demonstrated. Outcomes of the intervention are presented as well as issues of cost-effectiveness, feasibility, and alternative implementation strategies. PMID- 15657412 TI - Development and implementation of a medication adherence training instrument for persons living with HIV: the MATI. AB - Recent advances in the medical management of HIV offer the potential for increased longevity, improved quality of life, and the treatment of HIV as a chronic, rather than terminal, illness. The advent of highly active antiretroviral therapy (HAART) has required the necessity of strict adherence to complex medication regimens. As a vital factor in the successful outcome of HAART therapy, adherence-focused treatment represents a teaching opportunity for practitioners involved in the care and management of persons who are HIV positive. Scores of articles have been published regarding nonadherence, and numerous strategies have been employed to encourage adherence. Despite these efforts, medication adherence continues to present problems for patients and health care providers. This article discusses prior and existing adherence interventions for people living with HIV and outlines the rationale related to the development of a structured protocol, the Medication Adherence Training Instrument (MATI), for the evaluation and enhancement of HIV medication adherence by health care providers. The key components to the MATI throughout all sessions are to provide information, enhance motivation, and assess current levels of adherence and HIV-related knowledge. PMID- 15657413 TI - An intervention to promote the female condom to sexually transmitted disease clinic patients. AB - This article describes a 1-hour behavioral intervention designed to promote female condoms and safer sex to women at a high risk for sexually transmitted diseases (STDs). The intervention includes a promotional videotape; a skills oriented counseling session with a nurse clinician; assorted take-home items, including a videotape for men; and free supplies of female and male condoms. Designed for women ages 18 to 34 attending public STD clinics, the intervention is developed using a systematic process of formative evaluation influenced by principles of social marketing and drawing on the social cognitive theory. The effect of the intervention on female and male condom use is evaluated using a pretest-posttest design with 1,159 women. Most elements of the intervention could be replicated in settings other than STD clinics and delivered by persons other than nurse clinicians. PMID- 15657414 TI - Collaborating with an urban community to develop an HIV and AIDS prevention program for black youth and families. AB - This article describes a collaboration between academic researchers and residents of a low-income, inner-city community to develop and deliver an HIV and AIDS prevention program for Black youth. The Chicago HIV Prevention and Adolescent Mental Health Project (CHAMP) Program was developed and implemented to decrease HIV and AIDS risk exposure among youth living in a community that has been dramatically affected by HIV and AIDS. The article outlines (a) phases in the collaborative process to develop the program; (b) strategies used to embed contextually relevant themes and activities that address individual and systemic factors influencing HIV and AIDS risk; (c) a process model, based on the CHAMP experience, that can be replicated to develop programs for other youth problems; (d) descriptions of the CHAMP preadolescent and early adolescent curricula; (e) and how university- and community-based facilitators were trained to collaborate as a team to implement the CHAMP Program. Information is also provided about delivering the program in a distressed urban setting. PMID- 15657415 TI - A web-based therapeutic workplace for the treatment of drug addiction and chronic unemployment. AB - This article describes a Web-based therapeutic workplace intervention designed to promote heroin and cocaine abstinence and train and employ participants as data entry operators. Patients are paid to participate in training and then to perform data entry jobs in a therapeutic workplace business. Salary is linked to abstinence by requiring patients to provide drug-free urine samples to gain access to the workplace. Prior data show that a prototype of the intervention could promote drug abstinence. Preliminary data on the Web-based intervention suggest that it should be able to teach adults with histories of chronic unemployment and drug addiction to become skilled data entry operators in about 3 to 6 months. Early experience in the business provides preliminary evidence that it might become financially successful. The therapeutic workplace intervention may serve as an effective and practical long-term treatment for chronic unemployment and heroin and cocaine addiction. PMID- 15657416 TI - Direct and indirect interactions between calcineurin-NFAT and MEK1-extracellular signal-regulated kinase 1/2 signaling pathways regulate cardiac gene expression and cellular growth. AB - MEK1, a member of the mitogen-activated protein kinase (MAPK) cascade that directly activates extracellular signal-regulated kinase (ERK), induces cardiac hypertrophy in transgenic mice. Calcineurin is a calcium-regulated protein phosphatase that also functions as a positive regulator of cardiac hypertrophic growth through a direct mechanism involving activation of nuclear factor of activated T-cell (NFAT) transcription factors. Here we determined that calcineurin-NFAT and MEK1-ERK1/2 signaling pathways are interdependent in cardiomyocytes, where they directly coregulate the hypertrophic growth response. For example, genetic deletion of the calcineurin Abeta gene reduced the hypertrophic response elicited by an activated MEK1 transgene in the heart, while inhibition of calcineurin or NFAT in cultured neonatal cardiomyocytes also blunted the hypertrophic response driven by activated MEK1. Conversely, targeted inhibition of MEK1-ERK1/2 signaling in cultured cardiomyocytes attenuated the hypertrophic growth response directed by activated calcineurin. However, targeted inhibition of MEK1-ERK1/2 signaling did not directly affect calcineurin-NFAT activation, nor was MEK1-ERK1/2 activation altered by targeted inhibition of calcineurin-NFAT. Mechanistically, we show that MEK1-ERK1/2 signaling augments NFAT transcriptional activity independent of calcineurin, independent of changes in NFAT nuclear localization, and independent of alterations in NFAT transactivation potential. In contrast, MEK1-ERK1/2 signaling enhances NFAT dependent gene expression through an indirect mechanism involving induction of cardiac AP-1 activity, which functions as a necessary NFAT-interacting partner. As a second mechanism, MEK1-ERK1/2 and calcineurin-NFAT proteins form a complex in cardiac myocytes, resulting in direct phosphorylation of NFATc3 within its C terminus. MEK1-ERK1/2-mediated phosphorylation of NFATc3 directly augmented its DNA binding activity, while inhibition of MEK1-ERK1/2 signaling reduced NFATc3 DNA binding activity. Collectively, these results indicate that calcineurin-NFAT and MEK1-ERK1/2 pathways constitute a codependent signaling module in cardiomyocytes that coordinately regulates the growth response through two distinct mechanisms. PMID- 15657417 TI - Identification of putative new splicing targets for ETR-3 using sequences identified by systematic evolution of ligands by exponential enrichment. AB - ETR-3 (also know as BRUNOL3, NAPOR, and CUGBP2) is one of six members of the CELF (CUG-BP1- and ETR-3-like factor) family of splicing regulators. ETR-3 regulates splicing by direct binding to the pre-mRNA. We performed systematic evolution of ligands by exponential enrichment (SELEX) to identify the preferred binding sequence of ETR-3. After five rounds of SELEX, ETR-3 selected UG-rich sequences, in particular UG repeats and UGUU motifs. Either of these selected motifs was able to restore ETR-3 binding and responsiveness to a nonresponsive splicing reporter in vivo. Moreover, this effect was not specific to ETR-3 since minigenes containing either of the two motifs were responsive to two other CELF proteins (CUG-BP1 and CELF4), indicating that different members of the CELF family can mediate their effects via a common binding site. Using the SELEX-identified motifs to search the human genome, we identified several possible new ETR-3 targets. We created minigenes for two of these genes, the CFTR and MTMR1 genes, and confirmed that ETR-3 regulates their splicing patterns. For the CFTR minigene this regulation was demonstrated to be dependent on the presence of the putative binding site identified in our screen. These results validate this approach to search for new targets for RNA processing proteins. PMID- 15657418 TI - Mechanisms of transcription-replication collisions in bacteria. AB - While collisions between replication and transcription in bacteria are deemed inevitable, the fine details of the interplay between the two machineries are poorly understood. In this study, we evaluate the effects of transcription on the replication fork progression in vivo, by using electrophoresis analysis of replication intermediates. Studying Escherichia coli plasmids, which carry constitutive or inducible promoters in different orientations relative to the replication origin, we show that the mutual orientation of the two processes determines their mode of interaction. Replication elongation appears not to be affected by transcription proceeding in the codirectional orientation. Head-on transcription, by contrast, leads to severe inhibition of the replication fork progression. Furthermore, we evaluate the mechanism of this inhibition by limiting the area of direct contact between the two machineries. We observe that replication pausing zones coincide exactly with transcribed DNA segments. We conclude, therefore, that the replication fork is most likely attenuated upon direct physical interaction with the head-on transcription machinery. PMID- 15657419 TI - Rejoining of DNA double-strand breaks as a function of overhang length. AB - The ends of spontaneously occurring double-strand breaks (DSBs) may contain various lengths of single-stranded DNA, blocking lesions, and gaps and flaps generated by end annealing. To investigate the processing of such structures, we developed an assay in which annealed oligonucleotides are ligated onto the ends of a linearized plasmid which is then transformed into Saccharomyces cerevisiae. Reconstitution of a marker occurs only when the oligonucleotides are incorporated and repair is in frame, permitting rapid analysis of complex DSB ends. Here, we created DSBs with compatible overhangs of various lengths and asked which pathways are required for their precise repair. Three mechanisms of rejoining were observed, regardless of overhang polarity: nonhomologous end joining (NHEJ), a Rad52-dependent single-strand annealing-like pathway, and a third mechanism independent of the first two mechanisms. DSBs with overhangs of less than 4 bases were mainly repaired by NHEJ. Repair became less dependent on NHEJ when the overhangs were longer or had a higher GC content. Repair of overhangs greater than 8 nucleotides was as much as 150-fold more efficient, impaired 10-fold by rad52 mutation, and highly accurate. Reducing the microhomology extent between long overhangs reduced their repair dramatically, to less than NHEJ of comparable short overhangs. These data support a model in which annealing energy is a primary determinant of the rejoining efficiency and mechanism. PMID- 15657420 TI - Recruitment of the extracellular signal-regulated kinase/ribosomal S6 kinase signaling pathway to the NFATc4 transcription activation complex. AB - Integration of protein kinases into transcription activation complexes influences the magnitude of gene expression. The nuclear factor of activated T cells (NFAT) group of proteins are critical transcription factors that direct gene expression in immune and nonimmune cells. A balance of phosphotransferase activity is necessary for optimal NFAT activation. Activation of NFAT requires dephosphorylation by the calcium-mediated calcineurin phosphatase to promote NFAT nuclear accumulation, and the Ras-activated extracellular signal-regulated kinase (ERK) mitogen-activated protein (MAP) kinase, which targets NFAT partners, to potentiate transcription. Whether protein kinases operate on NFAT and contribute positively to transcription activation is not clear. Here, we coupled DNA affinity isolation with in-gel kinase assays to avidly pull down the activated NFAT and identify its associated protein kinases. We demonstrate that p90 ribosomal S6 kinase (RSK) is recruited to the NFAT-DNA transcription complex upon activation. The formation of RSK-NFATc4-DNA transcription complex is also apparent upon adipogenesis. Bound RSK phosphorylates Ser(676) and potentiates NFATc4 DNA binding by escalating NFAT-DNA association. Ser(676) is also targeted by the ERK MAP kinase, which interacts with NFAT at a distinct region than RSK. Thus, integration of the ERK/RSK signaling pathway provides a mechanism to modulate NFATc4 transcription activity. PMID- 15657421 TI - Stable binding of ATF6 to BiP in the endoplasmic reticulum stress response. AB - Endoplasmic reticulum (ER) stress-induced activation of ATF6, an ER membrane bound transcription factor, requires a dissociation step from its inhibitory regulator, BiP. It has been generally postulated that dissociation of the BiP ATF6 complex is a result of the competitive binding of misfolded proteins generated during ER stress. Here we present evidence against this model and for an active regulatory mechanism for dissociation of the complex. Contradictory to the competition model that is based on dynamic binding of BiP to ATF6, our data reveal relatively stable binding. First, the complex was easily isolated, in contrast to many chaperone complexes that require chemical cross-linking. Second, ATF6 bound at similar levels to wild-type BiP and a BiP mutant form that binds substrates stably because of a defect in its ATPase activity. Third, ER stress specifically induced the dissociation of BiP from ER stress transducers while the competition model would predict dissociation from any specific substrate. Fourth, the ATF6-BiP complex was resistant to ATP-induced dissociation in vitro when isolated without detergents, suggesting that cofactors stabilize the complex. In favor of an active dissociation model, one specific region within the ATF6 lumenal domain was identified as a specific ER stress-responsive sequence required for ER stress-triggered BiP release. Together, our results do not support a model in which competitive binding of misfolded proteins causes dissociation of the BiP-ATF6 complex in stressed cells. We propose that stable BiP binding is essential for ATF6 regulation and that ER stress dissociates BiP from ATF6 by actively restarting the BiP ATPase cycle. PMID- 15657422 TI - RAD51-dependent break-induced replication differs in kinetics and checkpoint responses from RAD51-mediated gene conversion. AB - Diploid Saccharomyces cells experiencing a double-strand break (DSB) on one homologous chromosome repair the break by RAD51-mediated gene conversion >98% of the time. However, when extensive homologous sequences are restricted to one side of the DSB, repair can occur by both RAD51-dependent and RAD51-independent break induced replication (BIR) mechanisms. Here we characterize the kinetics and checkpoint dependence of RAD51-dependent BIR when the DSB is created within a chromosome. Gene conversion products appear within 2 h, and there is little, if any, induction of the DNA damage checkpoint; however, RAD51-dependent BIR occurs with a further delay of 2 to 4 h and cells arrest in response to the G(2)/M DNA damage checkpoint. RAD51-dependent BIR does not require special facilitating sequences that are required for a less efficient RAD51-independent process. RAD51 dependent BIR occurs efficiently in G(2)-arrested cells. Once repair is initiated, the rate of repair replication during BIR is comparable to that of normal DNA replication, as copying of >100 kb is completed less than 30 min after repair DNA synthesis is detected close to the DSB. PMID- 15657423 TI - Mapping and functional characterization of the TAF11 interaction with TFIIA. AB - TFIIA interacts with TFIID via association with TATA binding protein (TBP) and TBP-associated factor 11 (TAF11). We previously identified a mutation in the small subunit of TFIIA (toa2-I27K) that is defective for interaction with TAF11. To further explore the functional link between TFIIA and TAF11, the toa2-I27K allele was utilized in a genetic screen to isolate compensatory mutants in TAF11. Analysis of these compensatory mutants revealed that the interaction between TAF11 and TFIIA involves two distinct regions of TAF11: the highly conserved histone fold domain and the N-terminal region. Cells expressing a TAF11 allele defective for interaction with TFIIA exhibit conditional growth phenotypes and defects in transcription. Moreover, TAF11 imparts changes to both TFIIA-DNA and TBP-DNA contacts in the context of promoter DNA. These alterations appear to enhance the formation and stabilization of the TFIIA-TBP-DNA complex. Taken together, these studies provide essential information regarding the molecular organization of the TAF11-TFIIA interaction and define a mechanistic role for this association in the regulation of gene expression in vivo. PMID- 15657424 TI - Common effects of acidic activators on large-scale chromatin structure and transcription. AB - Large-scale chromatin decondensation has been observed after the targeting of certain acidic activators to heterochromatic chromatin domains. Acidic activators are often modular, with two or more separable transcriptional activation domains. Whether these smaller regions are sufficient for all functions of the activators has not been demonstrated. We adapted an inducible heterodimerization system to allow systematic dissection of the function of acidic activators, individual subdomains within these activators, and short acidic-hydrophobic peptide motifs within these subdomains. Here, we demonstrate that large-scale chromatin decondensation activity is a general property of acidic activators. Moreover, this activity maps to the same acidic activator subdomains and acidic-hydrophobic peptide motifs that are responsible for transcriptional activation. Two copies of a mutant peptide motif of VP16 (viral protein 16) possess large-scale chromatin decondensation activity but minimal transcriptional activity, and a synthetic acidic-hydrophobic peptide motif had large-scale chromatin decondensation activity comparable to the strongest full-length acidic activator but no transcriptional activity. Therefore, the general property of large-scale chromatin decondensation shared by most acidic activators is not simply a direct result of transcription per se but is most likely the result of the concerted action of coactivator proteins recruited by the activators' short acidic hydrophobic peptide motifs. PMID- 15657425 TI - Desmosomal cadherin misexpression alters beta-catenin stability and epidermal differentiation. AB - Desmosomal adhesion is important for the integrity and protective barrier function of the epidermis and is disregulated during carcinogenesis. Strong adhesion between keratinocytes is conferred by the desmosomal cadherins, desmocollin (Dsc) and desmoglein. These constitute two gene families, members of which are differentially expressed in epidermal strata. It has been suggested that this stratum-specific expression regulates keratinocyte differentiation. We tested this hypothesis by misdirecting the expression of the basally abundant desmosomal cadherins Dsc3a and Dsc3b to suprabasal differentiating keratinocytes in transgenic mice. No phenotype was apparent until adulthood, when mice developed variable ventral alopecia and had altered keratinocyte differentiation within affected areas. The follicular changes were reminiscent of changes in transgenic mice with an altered beta-catenin stability. Stabilized beta-catenin and increased beta-catenin transcriptional activity were demonstrated in transgenic mice prior to the phenotypic change and in transgenic keratinocytes as a consequence of transgene expression. Hence, a link between desmosomal cadherins and beta-catenin stability and signaling was demonstrated, and it was shown that desmocollin cadherin expression can affect keratinocyte differentiation. Furthermore, the first function for a "b-type" desmocollin cadherin was demonstrated. PMID- 15657426 TI - Regulation of Rho and Rac signaling to the actin cytoskeleton by paxillin during Drosophila development. AB - Paxillin is a prominent focal adhesion docking protein that regulates cell adhesion and migration. Although numerous paxillin-binding proteins have been identified and paxillin is required for normal embryogenesis, the precise mechanism by which paxillin functions in vivo has not yet been determined. We identified an ortholog of mammalian paxillin in Drosophila (Dpax) and have undertaken a genetic analysis of paxillin function during development. Overexpression of Dpax disrupted leg and wing development, suggesting a role for paxillin in imaginal disc morphogenesis. These defects may reflect a function for paxillin in regulation of Rho family GTPase signaling as paxillin interacts genetically with Rac and Rho in the developing eye. Moreover, a gain-of-function suppressor screen identified a genetic interaction between Dpax and cdi in wing development. cdi belongs to the cofilin kinase family, which includes the downstream Rho target, LIM kinase (LIMK). Significantly, strong genetic interactions were detected between Dpax and Dlimk, as well as downstream effectors of Dlimk. Supporting these genetic data, biochemical studies indicate that paxillin regulates Rac and Rho activity, positively regulating Rac and negatively regulating Rho. Taken together, these data indicate the importance of paxillin modulation of Rho family GTPases during development and identify the LIMK pathway as a critical target of paxillin-mediated Rho regulation. PMID- 15657427 TI - MOZ-TIF2 inhibits transcription by nuclear receptors and p53 by impairment of CBP function. AB - Chromosomal rearrangements associated with acute myeloid leukemia (AML) include fusions of the genes encoding the acetyltransferase MOZ or MORF with genes encoding the nuclear receptor coactivator TIF2, p300, or CBP. Here we show that MOZ-TIF2 acts as a dominant inhibitor of the transcriptional activities of CBP dependent activators such as nuclear receptors and p53. The dominant negative property of MOZ-TIF2 requires the CBP-binding domain (activation domain 1 [AD1]), and coimmunoprecipitation and fluorescent resonance energy transfer experiments show that MOZ-TIF2 interacts with CBP directly in vivo. The CBP-binding domain is also required for the ability of MOZ-TIF2 to extend the proliferative potential of murine bone marrow lineage-negative cells in vitro. We show that MOZ-TIF2 displays an aberrant nuclear distribution and that cells expressing this protein have reduced levels of cellular CBP, leading to depletion of CBP from PML bodies. In summary, our results indicate that disruption of the normal function of CBP and CBP-dependent activators is an important feature of MOZ-TIF2 action in AML. PMID- 15657428 TI - Filamin A-bound PEBP2beta/CBFbeta is retained in the cytoplasm and prevented from functioning as a partner of the Runx1 transcription factor. AB - The heterodimeric transcription factor PEBP2/CBF is composed of a DNA-binding subunit, called Runx1, and a non-DNA-binding subunit, called PEBP2beta/CBFbeta. The Runx1 protein is detected exclusively in the nuclei of most cells and tissues, whereas PEBP2beta is located in the cytoplasm. We addressed the mechanism by which PEBP2beta localizes to the cytoplasm and found that it is associated with filamin A, an actin-binding protein. Filamin A retains PEBP2beta in the cytoplasm, thereby hindering its engagement as a Runx1 partner. The interaction with filamin A is mediated by a region within PEBP2beta that includes amino acid residues 68 to 93. The deletion of this region or the repression of filamin A enables PEBP2beta to translocate to the nucleus. Based on these observations, we propose that PEBP2beta has two distinct domains, a newly defined regulatory domain that interacts with filamin A and the previously identified Runx1-binding domain. PMID- 15657429 TI - Human papillomavirus oncoprotein E7 targets the promyelocytic leukemia protein and circumvents cellular senescence via the Rb and p53 tumor suppressor pathways. AB - Cellular senescence can be triggered by a variety of signals, including loss of telomeric integrity or intense oncogenic signaling, and is considered a potent, natural tumor suppressor mechanism. Previously, it was shown that the promyelocytic leukemia protein (PML) induces cellular senescence when overexpressed in primary human fibroblasts. The mechanism by which the PML IV isoform elicits this irreversible growth arrest is believed to involve activation of the tumor suppressor pathways p21/p53 and p16/Rb; however, a requirement for either pathway has not been demonstrated unequivocally. To investigate the individual contributions of p53 and Rb to PML-induced senescence, we used oncoproteins E6 and E7 from human papillomaviruses (HPVs), which predominantly target p53 and Rb. We show that E7, but not E6, circumvents PML-induced senescence. Using different E7 mutant proteins, dominant negative cyclin dependent kinase 4, and p16 RNA interference, we demonstrate that Rb-related and Rb-independent mechanisms of E7 are necessary for subversion of PML-induced senescence and we identify PML as a novel target for E7. Interaction between E7 and a functional prosenescence complex composed of PML, p53, and CBP perturbs transcriptional activation of p53, thus highlighting a significant effect also on the p53 tumor suppressor pathway. Given the importance of HPV in the pathogenesis of cervical cancer, our results warrant a more detailed analyses of PML in HPV infections. PMID- 15657430 TI - Inhibition of macroautophagy triggers apoptosis. AB - Mammalian cells were observed to die under conditions in which nutrients were depleted and, simultaneously, macroautophagy was inhibited either genetically (by a small interfering RNA targeting Atg5, Atg6/Beclin 1-1, Atg10, or Atg12) or pharmacologically (by 3-methyladenine, hydroxychloroquine, bafilomycin A1, or monensin). Cell death occurred through apoptosis (type 1 cell death), since it was reduced by stabilization of mitochondrial membranes (with Bcl-2 or vMIA, a cytomegalovirus-derived gene) or by caspase inhibition. Under conditions in which the fusion between lysosomes and autophagosomes was inhibited, the formation of autophagic vacuoles was enhanced at a preapoptotic stage, as indicated by accumulation of LC3-II protein, ultrastructural studies, and an increase in the acidic vacuolar compartment. Cells exhibiting a morphology reminiscent of (autophagic) type 2 cell death, however, recovered, and only cells with a disrupted mitochondrial transmembrane potential were beyond the point of no return and inexorably died even under optimal culture conditions. All together, these data indicate that autophagy may be cytoprotective, at least under conditions of nutrient depletion, and point to an important cross talk between type 1 and type 2 cell death pathways. PMID- 15657431 TI - Silencing of unpaired chromatin and histone H2A ubiquitination in mammalian meiosis. AB - During meiotic prophase in male mammals, the X and Y chromosomes are incorporated in the XY body. This heterochromatic body is transcriptionally silenced and marked by increased ubiquitination of histone H2A. This led us to investigate the relationship between histone H2A ubiquitination and chromatin silencing in more detail. First, we found that ubiquitinated H2A also marks the silenced X chromosome of the Barr body in female somatic cells. Next, we studied a possible relationship between H2A ubiquitination, chromatin silencing, and unpaired chromatin in meiotic prophase. The mouse models used carry an unpaired autosomal region in male meiosis or unpaired X and Y chromosomes in female meiosis. We show that ubiquitinated histone H2A is associated with transcriptional silencing of large chromatin regions. This silencing in mammalian meiotic prophase cells concerns unpaired chromatin regions and resembles a phenomenon described for the fungus Neurospora crassa and named meiotic silencing by unpaired DNA. PMID- 15657432 TI - c-Jun N-terminal kinase contributes to aberrant retinoid signaling in lung cancer cells by phosphorylating and inducing proteasomal degradation of retinoic acid receptor alpha. AB - Retinoic acid (RA) is the ligand for nuclear RA receptors (RARs and RXRs) and is crucial for normal epithelial cell growth and differentiation. During malignant transformation, human bronchial epithelial cells acquire a block in retinoid signaling caused in part by a transcriptional defect in RARs. Here, we show that activation of c-Jun N-terminal kinase (JNK) contributes to RAR dysfunction by phosphorylating RARalpha and inducing degradation through the ubiquitin proteasomal pathway. Analysis of RARalpha mutants and phosphopeptide mapping revealed that RARalpha residues Thr181, Ser445, and Ser461 are phosphorylated by JNK. Mutation of these residues to alanines prevented efficient ubiquitination of RARalpha and increased the stability of the protein. We investigated the importance of RARalpha phosphorylation by JNK as a mediator of retinoid resistance in lung cancer. Mice that develop lung cancer from activation of a latent K-ras oncogene had high intratumoral JNK activity and low RARalpha levels and were resistant to treatment with an RAR ligand. JNK inhibition in a human lung cancer cell line enhanced RARalpha levels, ligand-induced activity of RXR RAR dimers, and growth inhibition by RA. These findings point to JNK as a key mediator of aberrant retinoid signaling in lung cancer cells. PMID- 15657433 TI - POT1 and TRF2 cooperate to maintain telomeric integrity. AB - Mammalian telomeric DNA contains duplex TTAGGG repeats and single-stranded overhangs. POT1 (protection of telomeres 1) is a telomere-specific single stranded DNA-binding protein, highly conserved in eukaryotes. The biological function of human POT1 is not well understood. In the present study, we demonstrate that POT1 plays a key role in telomeric end protection. The reduction of POT1 by RNA interference led to the loss of telomeric single-stranded overhangs and induced apoptosis, chromosomal instability, and senescence in cells. POT1 and TRF2 interacted with each other to form a complex with telomeric DNA. A dominant negative TRF2, TRF2(DeltaBDeltaM), bound to POT1 and prevented it from binding to telomeres. POT1 overexpression protected against TRF2(DeltaBDeltaM)-induced loss of telomeric single-stranded overhangs, chromosomal instability, and senescence. These results demonstrate that POT1 and TRF2 share in part in the same pathway for telomere capping and suggest that POT1 binds to the telomeric single-stranded DNA in the D-loop and cooperates with TRF2 in t-loop maintenance. PMID- 15657434 TI - Genetic replacement of cyclin D1 function in mouse development by cyclin D2. AB - D cyclins (D1, D2, and D3) are components of the core cell cycle machinery in mammalian cells. It is unclear whether each of the D cyclins performs unique, tissue-specific functions or the three proteins have virtually identical functions and differ mainly in their pattern of expression. We previously generated mice lacking cyclin D1, and we observed that these animals displayed hypoplastic retinas and underdeveloped mammary glands and a presented developmental neurological abnormality. We now asked whether the specific requirement for cyclin D1 in these tissues reflected a unique pattern of D cyclin expression or the presence of specialized functions for cyclin D1 in cyclin D1 dependent compartments. We generated a knock-in strain of mice expressing cyclin D2 in place of D1. Cyclin D2 was able to drive nearly normal development of retinas and mammary glands, and it partially replaced cyclin D1's function in neurological development. We conclude that the differences between these two D cyclins lie mostly in the tissue-specific pattern of their expression. However, we propose that subtle differences between the two D cyclins do exist and they may allow D cyclins to function in a highly optimized fashion. We reason that the acquisition of multiple D cyclins may allow mammalian cells to drive optimal proliferation of a diverse array of cell types. PMID- 15657435 TI - Nuclear oncoprotein prothymosin alpha is a partner of Keap1: implications for expression of oxidative stress-protecting genes. AB - Animal cells counteract oxidative stress and electrophilic attack through coordinated expression of a set of detoxifying and antioxidant enzyme genes mediated by transcription factor Nrf2. In unstressed cells, Nrf2 appears to be sequestered in the cytoplasm via association with an inhibitor protein, Keap1. Here, by using the yeast two-hybrid screen, human Keap1 has been identified as a partner of the nuclear protein prothymosin alpha. The in vivo and in vitro data indicated that the prothymosin alpha-Keap1 interaction is direct, highly specific, and functionally relevant. Furthermore, we showed that Keap1 is a nuclear-cytoplasmic shuttling protein equipped with a nuclear export signal that is important for its inhibitory action. Prothymosin alpha was able to liberate Nrf2 from the Nrf2-Keap1 inhibitory complex in vitro through competition with Nrf2 for binding to the same domain of Keap1. In vivo, the level of Nrf2 dependent transcription was correlated with the intracellular level of prothymosin alpha by using prothymosin alpha overproduction and mRNA interference approaches. Our data attribute to prothymosin alpha the role of intranuclear dissociator of the Nrf2-Keap1 complex, thus revealing a novel function for prothymosin alpha and adding a new dimension to the molecular mechanisms underlying expression of oxidative stress-protecting genes. PMID- 15657436 TI - Eukaryotic translation initiation factor 4E activity is modulated by HOXA9 at multiple levels. AB - The eukaryotic translation initiation factor 4E (eIF4E) alters gene expression on multiple levels. In the cytoplasm, eIF4E acts in the rate-limiting step of translation initiation. In the nucleus, eIF4E facilitates nuclear export of a subset of mRNAs. Both of these functions contribute to eIF4E's ability to oncogenically transform cells. We report here that the homeodomain protein, HOXA9, is a positive regulator of eIF4E. HOXA9 stimulates eIF4E-dependent export of cyclin D1 and ornithine decarboxylase (ODC) mRNAs in the nucleus, as well as increases the translation efficiency of ODC mRNA in the cytoplasm. These activities depend on direct interactions of HOXA9 with eIF4E and are independent of the role of HOXA9 in transcription. At the biochemical level, HOXA9 mediates these effects by competing with factors that repress eIF4E function, in particular the proline-rich homeodomain PRH/Hex. This competitive mechanism of eIF4E regulation is disrupted in a subset of leukemias, where HOXA9 displaces PRH from eIF4E, thereby contributing to eIF4E's dysregulation. In regard to these results and our previous finding that approximately 200 homeodomain proteins contain eIF4E binding sites, we propose that homeodomain modulation of eIF4E activity is a novel means through which this family of proteins implements their effects on growth and development. PMID- 15657437 TI - Negative regulation of NF-kappaB signaling by PIAS1. AB - The NF-kappaB family of transcription factors is activated by a wide variety of signals to regulate a spectrum of cellular processes. The proper regulation of NF kappaB activity is critical, since abnormal NF-kappaB signaling is associated with a number of human illnesses, such as chronic inflammatory diseases and cancer. We report here that PIAS1 (protein inhibitor of activated STAT1) is an important negative regulator of NF-kappaB. Upon cytokine stimulation, the p65 subunit of NF-kappaB translocates into the nucleus, where it interacts with PIAS1. The binding of PIAS1 to p65 inhibits cytokine-induced NF-kappaB-dependent gene activation. PIAS1 blocks the DNA binding activity of p65 both in vitro and in vivo. Consistently, chromatin immunoprecipitation assays indicate that the binding of p65 to the promoters of NF-kappaB-regulated genes is significantly enhanced in Pias1-/- cells. Microarray analysis indicates that the removal of PIAS1 results in an increased expression of a subset of NF-kappaB-mediated genes in response to tumor necrosis factor alpha and lipopolysaccharide. Consistently, Pias1 null mice showed elevated proinflammatory cytokines. Our results identify PIAS1 as a novel negative regulator of NF-kappaB. PMID- 15657438 TI - Similar effects of Brca2 truncation and Rad51 paralog deficiency on immunoglobulin V gene diversification in DT40 cells support an early role for Rad51 paralogs in homologous recombination. AB - BRCA2 is a tumor suppressor gene that is linked to hereditary breast and ovarian cancer. Although the Brca2 protein participates in homologous DNA recombination (HR), its precise role remains unclear. From chicken DT40 cells, we generated BRCA2 gene-deficient cells which harbor a truncation at the 3' end of the BRC3 repeat (brca2tr). Comparison of the characteristics of brca2tr cells with those of other HR-deficient DT40 clones revealed marked similarities with rad51 paralog mutants (rad51b, rad51c, rad51d, xrcc2, or xrcc3 cells). The phenotypic similarities include a shift from HR-mediated diversification to single nucleotide substitutions in the immunoglobulin variable gene segment and the partial reversion of this shift by overexpression of Rad51. Although recent evidence supports at least Xrcc3 and Rad51C playing a role late in HR, our data suggest that Brca2 and the Rad51 paralogs may also contribute to HR at the same early step, with their loss resulting in the stimulation of an alternative, error prone repair pathway. PMID- 15657439 TI - Muscle-specific Pten deletion protects against insulin resistance and diabetes. AB - Pten (phosphatase with tensin homology), a dual-specificity phosphatase, is a negative regulator of the phosphoinositide 3-kinase (PI3K)/Akt signaling pathway. Pten regulates a vast array of biological functions including growth, metabolism, and longevity. Although the PI3K/Akt pathway is a key determinant of the insulin dependent increase in glucose uptake into muscle and adipose cells, the contribution of this pathway in muscle to whole-body glucose homeostasis is unclear. Here we show that muscle-specific deletion of Pten protected mice from insulin resistance and diabetes caused by high-fat feeding. Deletion of muscle Pten resulted in enhanced insulin-stimulated 2-deoxyglucose uptake and Akt phosphorylation in soleus but, surprisingly, not in extensor digitorum longus muscle compared to littermate controls upon high-fat feeding, and these mice were spared from developing hyperinsulinemia and islet hyperplasia. Muscle Pten may be a potential target for treatment or prevention of insulin resistance and diabetes. PMID- 15657440 TI - Phosphorylation of Par-4 by protein kinase A is critical for apoptosis. AB - Despite distinct dissimilarities, diverse cancers express several common protumorigenic traits. We present here evidence that the proapoptotic protein Par 4 utilizes one such common tumorigenic trait to become selectively activated and induce apoptosis in cancer cells. Elevated protein kinase A (PKA) activity noted in cancer cells activated the apoptotic function of ectopic Par-4 or its SAC (selective for apoptosis induction in cancer cells) domain, which induces apoptosis selectively in cancer cells and not in normal or immortalized cells. PKA preferentially phosphorylated Par-4 at the T155 residue within the SAC domain in cancer cells. Moreover, pharmacological-, peptide-, or small interfering RNA mediated inhibition of PKA activity in cancer cells resulted in abrogation of both T155 phosphorylation and apoptosis by Par-4. The mechanism of activation of endogenous Par-4 was similar to that of ectopic Par-4, and in response to exogenous stimuli, endogenous Par-4 induced apoptosis by a PKA- and phosphorylated T155-dependent mechanism. Enforced elevation of PKA activity in normal cells resulted in apoptosis by the SAC domain of Par-4 in a T155-dependent manner. Together, these observations suggest that selective apoptosis of cancer cells by the SAC domain of Par-4 involves phosphorylation of T155 by PKA. These findings uncover a novel mechanism engaging PKA, a procancerous activity commonly elevated in most tumor cells, to activate the cancer selective apoptotic action of Par-4. PMID- 15657442 TI - The deubiquitylation activity of Ubp8 is dependent upon Sgf11 and its association with the SAGA complex. AB - Covalent modifications of the histone tails and the cross talk between these modifications are hallmark features of gene regulation. The SAGA histone acetyltransferase complex is one of the most well-characterized complexes involved in these covalent modifications. The recent finding that the removal of the ubiquitin group from H2B is performed by a component of SAGA, Ubp8, is intriguing as it assigns two posttranslation modification processes to one complex. In this work, we characterize the association of Ubp8 with SAGA and the effect that acetylation and deubiquitylation have on one another in vitro and in vivo. We found not only that Ubp8 is a part of the SAGA complex, but also that its deubiquitylation activity requires Ubp8's association with SAGA. Furthermore, we found that the Ubp8 association with SAGA requires Sgf11 and that this requirement is reciprocal. We also found that the acetylation and deubiquitylation activities of SAGA are independent of one another. However, we found that preacetylating histone H2B inhibited subsequent deubiquitylation. Additionally, we found that increasing the ubiquitylation state of H2B inhibited the expression of the ARG1 gene, whose repression was previously shown to require the RAD6 ubiquitin ligase. Taken together, these data indicate that the expression of some genes, including ARG1, is regulated by a balance of histone H2B ubiquitylation in the cell. PMID- 15657441 TI - H2B ubiquitin protease Ubp8 and Sgf11 constitute a discrete functional module within the Saccharomyces cerevisiae SAGA complex. AB - The SAGA complex is a multisubunit protein complex involved in transcriptional regulation in Saccharomyces cerevisiae. SAGA combines proteins involved in interactions with DNA-bound activators and TATA-binding protein (TBP), as well as enzymes for histone acetylation (Gcn5) and histone deubiquitylation (Ubp8). We recently showed that H2B ubiquitylation and Ubp8-mediated deubiquitylation are both required for transcriptional activation. For this study, we investigated the interaction of Ubp8 with SAGA. Using mutagenesis, we identified a putative zinc (Zn) binding domain within Ubp8 as being critical for the association with SAGA. The Zn binding domain is required for H2B deubiquitylation and for growth on media requiring Ubp8's function in gene activation. Furthermore, we identified an 11-kDa subunit of SAGA, Sgf11, and showed that it is required for the Ubp8 association with SAGA and for H2B deubiquitylation. Different approaches indicated that the functions of Ubp8 and Sgf11 are related and separable from those of other components of SAGA. In particular, the profiles of Ubp8 and Sgf11 deletions were remarkably similar in microarray analyses and synthetic genetic interactions and were distinct from those of the Spt3 and Spt8 subunits of SAGA, which are involved in TBP regulation. These data indicate that Ubp8 and Sgf11 likely represent a new functional module within SAGA that is involved in gene regulation through H2B deubiquitylation. PMID- 15657443 TI - A single domain in human DNA polymerase iota mediates interaction with PCNA: implications for translesion DNA synthesis. AB - DNA polymerases (Pols) of the Y family rescue stalled replication forks by promoting replication through DNA lesions. Humans have four Y family Pols, eta, iota, kappa, and Rev1, of which Pols eta, iota, and kappa have been shown to physically interact with proliferating cell nuclear antigen (PCNA) and be functionally stimulated by it. However, in sharp contrast to the large increase in processivity that PCNA binding imparts to the replicative Pol, Poldelta, the processivity of Y family Pols is not enhanced upon PCNA binding. Instead, PCNA binding improves the efficiency of nucleotide incorporation via a reduction in the apparent K(m) for the nucleotide. Here we show that Poliota interacts with PCNA via only one of its conserved PCNA binding motifs, regardless of whether PCNA is bound to DNA or not. The mode of PCNA binding by Poliota is quite unlike that in Poldelta, where multisite interactions with PCNA provide for a very tight binding of the replicating Pol with PCNA. We discuss the implications of these observations for the accuracy of DNA synthesis during translesion synthesis and for the process of Pol exchange at the lesion site. PMID- 15657444 TI - TIEG1 null mouse-derived osteoblasts are defective in mineralization and in support of osteoclast differentiation in vitro. AB - Transforming growth factor beta-inducible early gene 1 (TIEG1) is a member of the Kruppel-like transcription factor family. To understand the physiological role of TIEG1, we generated TIEG(-/-) (null) mice and found that the TIEG(-/-) mice had increased osteoblast numbers with no increased bone formation parameters. However, when calvarial osteoblasts (OBs) were isolated from neonatal TIEG(-/-) and TIEG(+/+) mice and cultured in vitro, the TIEG(-/-) cells displayed reduced expression of important OB differentiation markers. When the OBs were differentiated in vitro by treatment with bone morphogenic protein 2, the OBs from TIEG(+/+) calvaria displayed several mineralized nodules in culture, whereas those from TIEG(-/-) mice showed no nodules. To characterize the OBs' ability to support osteoclast differentiation, the OBs from TIEG(+/+) and TIEG(-/-) mice were cultured with marrow and spleen cells from TIEG(+/+) mice. Significantly fewer osteoclasts developed when TIEG(-/-) OBs were used to support osteoclast differentiation than when TIEG(+/+) OBs were used. Examination of gene expression in the TIEG(-/-) OBs revealed decreased RANKL and increased OPG expression compared to TIEG(+/+) OBs. The addition of RANKL to these cocultures only partially restored the ability of TIEG(-/-) OBs to support osteoclast differentiation, whereas M-CSF alone or combined with RANKL had no additional effect on osteoclast differentiation. We conclude from these data that TIEG1 expression in OBs is critical for both osteoblast-mediated mineralization and osteoblast support of osteoclast differentiation. PMID- 15657447 TI - Mother Teresa versus Ebenezer Scrooge: mortality salience leads proselfs to endorse self-transcendent values (unless proselfs are reassured). AB - Recently, Jonas, Schimel, Greenberg, and Pyszczynski demonstrated that mortality salience (MS) increased contributions to (an ingroup) charity (i.e., the Scrooge effect). The authors examined whether individual differences in social value orientations would moderate the Scrooge effect. In line with an Ebenezer shift hypothesis, proselfs were less likely than prosocials to endorse self transcendent values in a dental pain control condition but were indistinguishable from prosocials in an MS condition as proselfs increased endorsement of self transcendent values under MS (Experiments 1 and 2a). However, when participants gave their impressions of an unfavorable prosocial or a favorable proself prior to the MS manipulation, proselfs were again less likely than prosocials to endorse self-transcendent values (Experiments 2a and 2b), suggesting that proselfs are unlikely to transform into prosocials under conditions of MS when given reasons to disidentify with prosocial values or identify with proself values. PMID- 15657446 TI - Mediation and moderation of the relationship between intrinsic religiousness and mental health. AB - The present study examined specific aspects of individuals' personal strivings as mediators, and religious tradition as a moderator, of the relationship between intrinsic religiousness and mental health. In a sample of 268 university students, the negative relationship between intrinsic religiousness and hostility was mediated by the degree of sanctification within individuals' strivings. The relationships between intrinsic religiousness and both anxiety and depression were moderated by religious tradition, with Catholics' intrinsic religiousness significantly associated with greater anxiety and depression but Protestants' intrinsic religiousness not significantly associated with either of these mental health variables. Implications of these results for future research on religiousness are discussed. PMID- 15657448 TI - Victim and perpetrator accounts of interpersonal transgressions: self-serving or relationship-serving biases? AB - Two studies investigated perpetrator and victim biases in reported transgressions. Study 1 showed that in nonromantic relationships, perpetrators were more likely than victims to emphasize details that minimized their transgressions, whereas victims were more likely to exaggerate the severity of the event. Study 2 examined these perspective-related differences in romantic relationships and their relationship to forgiveness. Although victims were less likely than perpetrators to include features that minimized the event, they were no more likely to include features that magnified the event; individuals in highly satisfying relationships were less likely to exhibit self-serving biases than were individuals in less satisfying relationships. The data also were consistent with a causal sequence in which positive relationship quality led to more benign interpretations of a transgression, which in turn, promoted forgiveness. PMID- 15657445 TI - A direct intersection between p53 and transforming growth factor beta pathways targets chromatin modification and transcription repression of the alpha fetoprotein gene. AB - We purified the oncoprotein SnoN and found that it functions as a corepressor of the tumor suppressor p53 in the regulation of the hepatic alpha-fetoprotein (AFP) tumor marker gene. p53 promotes SnoN and histone deacetylase interaction at an overlapping Smad binding, p53 regulatory element (SBE/p53RE) in AFP. Comparison of wild-type and p53-null mouse liver tissue by using chromatin immunoprecipitation (ChIP) reveals that the absence of p53 protein correlates with the disappearance of SnoN at the SBE/p53RE and loss of AFP developmental repression. Treatment of AFP-expressing hepatoma cells with transforming growth factor-beta1 (TGF-beta1) induced SnoN transcription and Smad2 activation, concomitant with AFP repression. ChIP assays show that TGF-beta1 stimulates p53, Smad4, P-Smad2 binding, and histone H3K9 deacetylation and methylation, at the SBE/p53RE. Depletion, by small interfering RNA, of SnoN and/or p53 in hepatoma cells disrupted repression of AFP transcription. These findings support a model of cooperativity between p53 and TGF-beta effectors in chromatin modification and transcription repression of an oncodevelopmental tumor marker gene. PMID- 15657449 TI - Attitudinal ambivalence, rumination, and forgiveness of partner transgressions in marriage. AB - Although positive and negative attitudes toward a transgressor are related to increased and decreased forgiveness, respectively, prior research has failed to investigate forgiveness among those who feel both positively and negatively toward a transgressor. Therefore, the authors examined such ambivalence and its relationship to forgiveness. It was hypothesized that spouses with ambivalent attitudes toward their partner will be less forgiving of a partner transgression because such an event is likely to prime the negative component of their ambivalence. Because ruminating about a transgression also has the potential to prime the negative component of one's ambivalence, an interaction between rumination and ambivalence was predicted. Data from 87 married couples showed that greater attitudinal ambivalence toward the partner was associated with decreased forgiveness only when husbands and wives thought about the transgression frequently; ambivalence was not related to forgiveness in the absence of rumination. The implications of these findings for understanding forgiveness in marriage and for increasing forgiveness among married couples are discussed. PMID- 15657450 TI - Attachment style, excessive reassurance seeking, relationship processes, and depression. AB - The authors examined the association between excessive reassurance seeking (ERS) in couple relationships and depression, hypothesizing that this association can be explained by attachment anxiety. In Study 1, 72 couples completed questionnaires about ERS, depression, attachment style, and relationship quality. In Study 2, 61 couples completed the same measures in addition to completing daily diary assessments for 14 days. In both studies, ERS was related to depression, but only because of its association with attachment anxiety. The association between attachment anxiety and depression was not mediated by either partner's relationship quality, even though partners noticed each other's ERS. Instead, relationship quality was related primarily to avoidant attachment. These findings were replicated at a daily level of analysis, where interesting details of the underlying processes were revealed. At least in young unmarried couples, depression is associated with attachment anxiety--and one of its facets, ERS- mostly for intrapsychic reasons. PMID- 15657452 TI - Marital quality, forgiveness, empathy, and rumination: a longitudinal analysis. AB - McCullough, Rachal, et al.'s (1998) social-psychological framework of forgiveness informed a longitudinal study that examined the extent to which marital forgiveness is determined by social-cognitive (the offended spouse's rumination and emotional empathy) and relationship variables (the quality of the relationship in which the offense took place). In the study, 119 husbands and 124 wives from long- and medium-term marriages in north Italy provided data at two time points separated by a 6-month interval. Structural equation models showed that rumination and empathy independently predicted concurrent marital forgiveness. Forgiveness in turn predicted concurrent marital quality. Finally, reciprocal directions of effect emerged between forgiveness and marital quality over time. These results are discussed in terms of their implications for promoting forgiveness, and future research directions are outlined. PMID- 15657451 TI - On the eve of war: authoritarianism, social dominance, and American students' attitudes toward attacking Iraq. AB - In the week before the 2003 American attack on Iraq, the effects of authoritarianism and the social dominance orientation on support for the attack were examined. Based on prior research on the nature of these constructs, a structural model was developed and tested. As predicted, authoritarianism strengthened support for the attack by intensifying the perception that Iraq threatened America. Social dominance increased support by reducing concern for the likely human costs of the war. Both also increased blind patriotism, which in turn reduced concern for the war's human costs and was reciprocally related to the belief that Iraq threatened America. PMID- 15657453 TI - Promoting success or preventing failure: cultural differences in motivation by positive and negative role models. AB - In two studies, cross-cultural differences in reactions to positive and negative role models were examined. The authors predicted that individuals from collectivistic cultures, who have a stronger prevention orientation, would be most motivated by negative role models, who highlight a strategy of avoiding failure; individuals from individualistic cultures, who have a stronger promotion focus, would be most motivated by positive role models, who highlight a strategy of pursuing success. In Study 1, the authors examined participants' reported preferences for positive and negative role models. Asian Canadian participants reported finding negative models more motivating than did European Canadians; self-construals and regulatory focus mediated cultural differences in reactions to role models. In Study 2, the authors examined the impact of role models on the academic motivation of Asian Canadian and European Canadian participants. Asian Canadians were motivated only by a negative model, and European Canadians were motivated only by a positive model. PMID- 15657454 TI - Grandparent-grandchild contact and attitudes toward older adults: moderator and mediator effects. AB - Two studies tested the intergroup contact hypothesis in the context of the grandparent-grandchild relationship. The hypothesis suggests that contact with an out-group member has more influence on attitudes toward the out-group when group memberships are salient. In Study 1, the predicted link was found but only for grandparents with whom the grandchild had more frequent contact. The second study examined only the most frequent grandparent relationship and replicated the effect. This study also investigated the role of various mediators of the link between quality of contact and attitudes, as well as quality of contact and perceived out-group variability. Perspective taking, anxiety, and accommodation mediated the effects of contact on attitudes, whereas individuation and self disclosure mediated the effects of contact on perceived out-group variability. Moderated mediational analysis indicated that the moderating effect of group salience occurs between quality of contact and the mediator, not between the mediator and attitudes. PMID- 15657455 TI - The gaze of the optimist. AB - Two studies used eye tracking to investigate the attentional preferences of optimists and pessimists to negative emotional stimuli. In both studies, optimistic and pessimistic college students viewed three types of visual stimuli while having their eye movements tracked: skin cancer (melanoma) images, matched schematic line drawings, and neutral faces. In the first study, participants were asked to view the images naturally, whereas in the second study, some participants received a relevance manipulation. Percentage of fixation time to the different images was measured. Optimists showed selective inattention to the skin cancer images, even after controlling for attention to matched schematic line drawings. This relationship remained significant in both studies after controlling for the effects of neuroticism, affect, anxiety, relevance, and perceptual variables. These data suggest that optimists may indeed wear "rose colored glasses" in their processing of information from the world. PMID- 15657456 TI - Antecedents and reactions to health-related social control. AB - A model of the antecedents and reactions to health-related social control is proposed. This model suggests health behavior characteristics, including type, frequency, and severity of consequences, are social control antecedents. Social control is then thought to elicit better health behavior and emotions. Attributions to explain social control are proposed to effect emotional reactions and behavior. Undergraduates read hypothetical scenarios to test the proposed model. Study 1 found that health-compromising behaviors and behaviors with more severe consequences elicited more social control. Study 2 found that, compared to negative tactics, positive social control tactics elicited more behavior change, and compared to social/appearance concerns, attributions to health elicited positive emotions. Attributions did not moderate the impact of social control on emotions or behavior. PMID- 15657457 TI - Social identity complexity and outgroup tolerance. AB - Social identity complexity refers to the way in which individuals subjectively represent the relationships among their multiple ingroup memberships. More specifically, individuals with low social identity complexity see their ingroups as highly overlapping and convergent, whereas those with high complexity see their different ingroups as distinct and cross-cutting membership groups. The present study tested the hypothesis that perceived overlap among ingroup memberships would be negatively related to ingroup inclusiveness and tolerance for outgroups, such that individuals with high overlap (low complexity) would be less tolerant and accepting of outgroups in general than those with low overlap (high complexity). Results from a telephone interview survey of adult residents of the state of Ohio supported this hypothesis. Individual differences in complexity of perception of their national, religious, occupational, political, and recreational social identities were systematically related to their attitudes toward ethnic outgroups and diversity. PMID- 15657459 TI - Airway remodeling and inflammation in symptomatic infants with reversible airflow obstruction. AB - RATIONALE: We hypothesized that the epithelial reticular basement membrane (RBM) thickening and eosinophilic inflammation characteristic of asthma would be present in symptomatic infants with reversible airflow obstruction. METHODS: RBM thickness and numbers of inflammatory cells were determined in ultrathin sections of endobronchial biopsies obtained from 53 infants during clinical bronchoscopy for severe wheeze and/or cough. Group A: 16 infants with a median age of 12 months (range 3.4-26 months), with decreased specific airway conductance (sGaw) and bronchodilator reversibility; Group B: 22 infants with a median age of 12.4 months (5.1-25.9 months), with decreased sGaw but without bronchodilator reversibility; and Group C: 15 infants with a median age of 11.5 months (3.4-24.3 months) with normal sGaw. Additional comparisons were made with the following groups. Group D: 17 children, median age 10.3 years (6-16 years), with difficult asthma; Group E: 10 pediatric control subjects without asthma, median age 10 years (6-16 years); and Group F: nine adult normal, healthy control subjects, median age 27 years (21-42 years). MAIN RESULTS: There were no significant differences in RBM thickness or inflammatory cell number between the infant groups. RBM thickness was similar in the infants and Groups E and F. However, the RBM in all infant groups (Group A: median 4.3 microm [range 2.8-9.2 microm]; Group B: median 4.15 microm [range 2.7-5.8 microm]; Group C: median 3.8 microm [range 2.7-5.5 microm]) was significantly less thick than that in the older children with asthma (Group D: median 8.3 microm [range 5.3-12.7 microm]; p < 0.001). CONCLUSION: RBM thickening and the eosinophilic inflammation characteristic of asthma in older children and adults are not present in symptomatic infants with reversible airflow obstruction, even in the presence of atopy. PMID- 15657460 TI - Needs and opportunities for research in hypersensitivity pneumonitis. AB - Hypersensitivity pneumonitis (HP) develops after inhalation of many different environmental antigens, causing variable clinical symptoms that often make diagnosis uncertain. The prevalence of HP is higher than recognized, especially its chronic form. Mechanisms of disease are still incompletely known. Strategies to improve detection and diagnosis are needed, and treatment options, principally avoidance, are limited. A workshop recommended: a population-based study to more accurately document the incidence and prevalence of HP; better classification of disease stages, including natural history; evaluation of diagnostic tests and biomarkers used to detect disease; better correlation of computerized tomography lung imaging and pathologic changes; more study of inflammatory and immune mechanisms; and improvement of animal models that are more relevant for human disease. PMID- 15657461 TI - Silica particles enhance peripheral thrombosis: key role of lung macrophage neutrophil cross-talk. AB - RATIONALE: Inflammation and thrombosis are related via interactions between leukocytes, platelets, the vasculature, and the coagulation system. However, the mechanisms behind these interactions remain poorly understood. OBJECTIVES: We have investigated the effects of the well known pulmonary inflammation induced by silica for the development of peripheral thrombogenicity in a hamster model of thrombosis. In addition, the consequences of pulmonary macrophage and circulating monocyte and neutrophil depletion on the thrombogenicity were investigated. METHODS: Silica particles (2-200 mug/hamster) were intratracheally instilled, and experimental thrombosis in photochemically induced femoral vein lesions was assessed 24 hours later, in association with cellular infiltration in the lung. MEASUREMENTS AND MAIN RESULTS: Intratracheally instilled silica particles (20 and 200 mug/hamster) triggered pulmonary inflammation, together with stimulation of peripheral platelet-rich thrombus formation. Both the selective depletion of lung macrophages by intratracheal administration of clodronate liposomes and the combined depletion of circulating monocytes and neutrophils by intraperitoneal injection of cyclophosphamide significantly reduced silica-induced influx of macrophages and neutrophils in bronchoalveolar lavage, and reduced peripheral thrombogenicity. Silica-induced lung inflammation was accompanied by increased neutrophil elastase levels in bronchoalveolar lavage and in plasma. Specific neutrophil elastase inhibition in the lung did not affect lung inflammation but reduced peripheral thrombogenicity. CONCLUSION: These findings uncover pulmonary macrophage-neutrophil cross-talk releasing neutrophil elastase into the blood circulation. Elastase, triggering activation of circulating platelets, may then predispose platelets to initiate thrombotic events on mildly damaged vasculature. PMID- 15657462 TI - Effect of macrolides on in vivo ion transport across cystic fibrosis nasal epithelium. AB - Fourteen- and 15-member macrolide antibiotics are under investigation as potential therapeutic agents for cystic fibrosis (CF). The nonantibiotic mechanisms of action of these compounds in CF are not understood. We used nasal potential difference (NPD) measurements to test the effect of macrolides on airway epithelial ion (chloride, sodium) transport of CF mice and humans. We tested clarithromycin and azithromycin in mice, and clarithromycin in patients with CF. Baseline and post-treatment NPD was measured in two strains (C57Bl6 and BalbC) of CF transmembrane regulator "knockout" and littermate control mice, and in DeltaF508/DeltaF508 mice. In addition, NPD was measured in 18 human subjects with CF (17 DeltaF-508/DeltaF-508 and 1 DeltaF-508/other) who were undergoing a 12-month, randomized, double-blind crossover study of the effects of clarithromycin on pulmonary outcome in CF. Neither clarithromycin nor azithromycin affected ion transport characteristics of normal or CF nasal epithelium in either mouse or humans. We conclude that the apparent beneficial effects of macrolides on pulmonary outcome in CF are not mediated by their modulation of ion transport. PMID- 15657463 TI - Hypogonadism in men with chronic obstructive pulmonary disease: prevalence and quality of life. AB - We recently reported that hypogonadism does not affect respiratory muscle performance and exercise capacity in men with chronic obstructive pulmonary disease (COPD). In COPD, however, the relationship between exercise capacity and quality of life is controversial, making it unreliable to extrapolate about quality of life from exercise data. Accordingly, we determined prevalence and impact of hypogonadism on health-related quality of life in men with COPD. We enrolled 101 stable outpatient men (FEV1 1.34 +/- 0.04 L) older than 54 years; 38 patients were hypogonadal-a prevalence similar to that reported in the general population. The degree of airflow limitation did not predict levels of free testosterone. Quality of life, as quantified by a disease-specific instrument (St. George's Respiratory Questionnaire) and a general-health instrument (Veterans Short Form-36) were equivalent in the hypogonadal and eugonadal groups. Both groups demonstrated large decrements in perceived physical health and smaller decrements in perceived emotional and mental health. No relationship was found between free testosterone level and physical activity, respiratory symptoms, or quality of life. In conclusion, hypogonadism, although common among men older than 54 years with COPD, does not worsen the severity of respiratory symptoms or quality of life. PMID- 15657464 TI - Lung parenchyma remodeling in a murine model of chronic allergic inflammation. AB - This study tested the hypotheses that chronic allergic inflammation induces not only bronchial but also lung parenchyma remodeling, and that these histologic changes are associated with concurrent changes in respiratory mechanics. For this purpose, airway and lung parenchyma remodeling were evaluated by quantitative analysis of collagen and elastin, immunohistochemistry (smooth-muscle actin expression, eosinophil, and dendritic cell densities), and electron microscopy. In vivo (airway resistance, viscoelastic pressure, and static elastance) and in vitro (tissue elastance, resistance, and hysteresivity) respiratory mechanics were also analyzed. BALB/c mice were sensitized with ovalbumin and exposed to repeated ovalbumin challenges. A marked eosinophilic infiltration was seen in lung parenchyma and in large and distal airways. Neutrophils, lymphocytes, and dendritic cells also infiltrated the lungs. There was subepithelial fibrosis, myocyte hypertrophy and hyperplasia, elastic fiber fragmentation, and increased numbers of myofibroblasts in airways and lung parenchyma. Collagen fiber content was increased in the alveolar walls. The volume proportion of smooth muscle specific actin was augmented in distal airways and alveolar duct walls. Airway resistance, viscoelastic pressure, static elastance, and tissue elastance and resistance were significantly increased. In conclusion, prolonged allergen exposure induced remodeling not only of the airway wall but also of the lung parenchyma, leading to in vivo and in vitro mechanical changes. PMID- 15657465 TI - Hypocapnia is not a predictor of central sleep apnea in patients with cirrhosis. AB - During sleep, maintenance of rhythmic breathing is critically dependent on the level of PCO(2), such that if the prevailing spontaneous PCO(2) decreases below the apneic threshold, central sleep apnea (CSA) occurs. Several studies have shown that in patients with systolic heart failure (SHF), presence of a low, awake arterial PCO(2) (Pa(CO(2))) increases the likelihood of developing CSA during sleep. We therefore sought to determine if a low Pa(CO(2)) is a predictor of CSA in patients with cirrhosis of the liver and with normal left ventricular systolic function. In 13 hypocapnic (Pa(CO(2)) < 36 mm Hg, mean = 33 mm Hg) patients with SHF and a mean left ventricular ejection fraction of 23%, the mean apnea-hypopnea index, was 28/hour. CSA accounted for most of the breathing disorders. In 10 hypocapnic (Pa(CO(2)) < 36 mm Hg, mean = 32 mm Hg) patients with cirrhosis and a normal left ventricular ejection fraction (60%), the mean apnea hypopnea index was 2/hour. The maximum central apnea index was 0.2/hour. There were no significant differences in age, demographics, pulmonary function tests, Pa(O(2)), Pa(CO(2)), minute and alveolar ventilation, and ventilatory responses to CO(2) between the two groups. We conclude that, in contrast to SHF, presence of hypocapnia does not predict CSA in cirrhosis. PMID- 15657466 TI - Characterization of cytokine/chemokine profiles of severe acute respiratory syndrome. AB - RATIONALE: There is currently no optimal treatment or effective drug for severe acute respiratory syndrome (SARS), because the immunopathologic mechanism is poorly understood. OBJECTIVES: To explore the immune mechanism underlying the pathogenesis of SARS, we studied the expression profile of cytokines/chemokines in the blood and the immunopathology of the lung and lymphoid tissues. METHODS: Fourteen cytokines/chemokines in the blood of 23 patients with SARS were dynamically screened, using a bead-based multiassay system. Reverse transcription polymerase chain reaction was performed to amplify mRNA. Histopathology of the lung and lymphoid tissues at autopsy was examined, using methods of immunohistochemistry and double immunofluorescence staining. MAIN RESULTS: Interferon-inducible protein-10 (IP-10) was markedly elevated in the blood during the early stage of SARS, and remained at a high level until convalescence. Moreover, IP-10 was highly expressed in both lung and lymphoid tissues, where monocyte-macrophage infiltration and depletion of lymphocytes were observed. The levels of interleukin-6, interleukin-8, and monocyte chemoattractant protein-1 were concomitantly increased in the blood of the patients with superinfection, and the mRNAs for these cytokines were also increased in lung tissues. CONCLUSIONS: Induction of IP-10 is a critical event in the initiation of immune mediated acute lung injury and lymphocyte apoptosis during the development of SARS. Superinfection after the immune injury is the main cause of death. The prompt elevation of interleukin-6, interleukin-8, and monocyte chemoattractant protein-1 is a sign of superinfection, indicating a high risk of death. PMID- 15657467 TI - Agonists of toll-like receptors 2 and 4 activate airway smooth muscle via mononuclear leukocytes. AB - RATIONALE: Toll-like receptors 2 and 4 (TLR2, TLR4) enable cellular responses to bacterial lipoproteins, LPS, and endogenous mediators of cell damage. They have an established role in the activation of leukocytes, endothelial cells, and some smooth muscle cell types, but their roles in airway smooth muscle are uncertain. OBJECTIVES: To determine the roles of TLRs in activation of airway smooth muscle. METHODS: Airway smooth muscle cells were cultured with TLR agonists, in the presence or absence of mononuclear leukocytes. MEASUREMENTS AND MAIN RESULTS: We observed expression of TLR2 and TLR4 mRNAs, which could be upregulated by treatment with proinflammatory cytokines in primary human airway smooth muscle, but no important functional responses to agonists of these TLRs were seen. Coincubation of airway smooth muscle with peripheral blood mononuclear cells, at concentrations as low as 250 mononuclear cells/ml, resulted in a marked cooperative response to TLR stimuli, and synergistic production of cytokines, including chemokines (interleukin [IL-]-8) and IL-6. This cooperative response was greater when monocytes were enriched and was transferable using supernatants from LPS-stimulated peripheral blood mononuclear cells. Activation of cocultures required IL-1 generation from mononuclear cells, and was blocked by IL-1 receptor antagonist, though IL-1 generation alone was not sufficient to account for the magnitude of mononuclear cell-dependent coculture activation. CONCLUSIONS: These data indicate that potent amplification of inflammation induced by TLR agonists, such as LPS, may be achieved by cooperativity between airway smooth muscle and leukocytes involved in immune surveillance or inflammation. PMID- 15657468 TI - Effect of maltose-containing sports drinks on exercise performance. AB - This study examined the effect of maltose-containing sports drinks on exercise performance. Ten subjects completed 4 trials. Each trial consisted of a glycogen depletion protocol, followed by a 15-min refueling, after which subjects performed an 1-h performance test while consuming one of the experimental drinks (HGlu, glucose; HMal, maltose; MalMix, sucrose, maltose, and maltodextrin; Plac, placebo). Drinks provided 0.65 g/kg body weight carbohydrates during refueling and 0.2 g/kg every 15 min during the performance test. Although no significant differences were found in performance (HGlu: 67.2 +/- 2.0; HMal: 68.6 +/- 1.7; MalMix: 66.7 +/- 2.0; Plac: 69.4 +/- 3.0 min, P> 0.05), subjects completed the MalMix trial 3.9%; faster than the Plac. Carbohydrate drinks caused comparable plasma glucose values that were significantly higher during refueling and at the end of exercise, compared to Plac. The data suggest that although carbohydrate drinks help to maintain plasma glucose at a higher level, no differences in performance could be detected after glycogen-depleting exercise. PMID- 15657469 TI - Effects of caffeine ingestion on exercise testing: a meta-analysis. AB - This study used the meta-analytic approach to examine the effects of caffeine ingestion on exercise testing. Forty double-blind studies with 76 effect sizes (ES) met the inclusion criteria. The type of exercise test was classified as endurance, graded, or short-term. In comparison with placebo, caffeine improved test outcome by 12.3 % (95 % CI, 9.1 to 15.4), which was equivalent to an overall ES of 0.41 (95 % CI, 0.31 to 0.51). Endurance exercise significantly improved test outcome (P < 0.05) more than either graded or short-term exercise. When exercise protocol was examined, time-to-exhaustion (Tlim) protocols had a significantly greater (P < 0.05) ES than either the graded or the non-Tlim protocol(s). The results from this meta-analysis confirm the ergogenic effects of caffeine, particularly for endurance testing that use Tlim protocols. PMID- 15657470 TI - The effects of pre-exercise high carbohydrate meals with different glycemic indices on blood leukocyte redistribution, IL-6, and hormonal responses during a subsequent prolonged exercise. AB - The purpose of this study was to determine the effect of pre-exercise high carbohydrate meals with high glycemic index (HGI) or low glycemic index (LGI) on blood leukocyte redistribution during subsequent endurance exercise. Eight male subjects performed a 90-min run on a treadmill at 70 % VO2max 3 h after ingesting an isocaloric HGI or LGI meal with GI values of 77 and 37, respectively. Blood counts of leukocytes, and neutrophils and the neutrophil/lymphocyte ratio were significantly lower in LGI than HGI at 90 min of exercise (P < 0.05). The plasma glucose concentrations were significantly higher in LGI than HGI between 15 min and 45 min of exercise. There were, however, no differences in plasma cortisol, growth hormone, and interleukin-6 concentrations between trials. Thus, the GI of a pre-exercise meal influences leukocyte trafficking and plasma glucose but has limited effects on circulating stress hormone and cytokine responses to exercise. PMID- 15657471 TI - Comparison of DXA, Siri's 2C, and Lohman' s Db-mineral models for estimating the body fat of physically active adults. AB - The authors used 3-component (3C) Db-mineral-model (Lohman, 1986) reference measures to cross-validate Siri's (1961) 2-component (2C) conversion formula and dual-energy X-ray absorptiometry (DXA) estimates of relative body fat (%BF) for physically active adults. Participants varied in age (18 to 59 y), body fatness, ethnicity (black, Hispanic, white), and physical activity level. The 3C Db mineral model was used to obtain reference measures of % BF (%BF3C) for comparison with body-composition measures from DXA and hydrodensitometry. For men (n = 110) and women (n= 110), %BF3C (14.0 % BF and 24.4 % BF, respectively) was more accurately estimated by Siri's 2C formula (% FSiri; men, r = 0.97, SEE = 1.77 % BF; women, r = 0.98, SEE = 1.56 % BF) than by DXA (% FDXA; men, r = 0.86, SEE = 3.54 % BF; women, r =0.88, SEE = 3.73 % BF). The average %BFSiri (men, 15.8 % BF; women, 24.7 % BF) and % FDXA (men, 16.2 % BF; women, 26.0 %BF) differed significantly (P < 0.001) from %BF3C. Siri's 2C model estimated the average % BF3C in this sample more accurately than DXA did. PMID- 15657472 TI - Gastrointestinal discomfort during intermittent high-intensity exercise: effect of carbohydrate-electrolyte beverage. AB - This study investigated whether different beverage carbohydrate concentration and osmolality would provoke gastrointestinal (GI) discomfort during intermittent, high-intensity exercise. Thirty-six adult and adolescent athletes were tested on separate days in a double-blind, randomized trial of 6 % and 8 % carbohydrate electrolytes (CHO-E) beverages during four 12-min quarters (Q) of circuit training that included intermittent sprints, lateral hops, shuttle runs, and vertical jumps. GI discomfort and fatigue surveys were completed before the first Q and immediately after each Q. All ratings of GI discomfort were modest throughout the study. The cumulative index for GI discomfort, however, was greater for the 8 % CHO-E beverage than for the 6 % CHO-E beverage at Q3 and Q4 (P < 0.05). Averaging across all 4 quarters, the 8 % CHO-E treatment produced significantly higher mean ratings of stomach upset and side ache. In conclusion, higher CHO concentration and osmolality in an ingested beverage provokes stomach upset and side ache. PMID- 15657474 TI - The impact of prior coffee consumption on the subsequent ergogenic effect of anhydrous caffeine. AB - This study examined whether the prior consumption of coffee (COF) decreased the ergogenic effect of the subsequent ingestion of anhydrous caffeine (CAF). Thirteen subjects performed 6 rides to exhaustion at 80 % VO2max 1.5 h after ingesting combinations of COF, decaffeinated coffee (DECOF), CAF, or placebo. The conditions were DECOF + placebo (A), DECOF + CAF (5 mg/kg) (B), COF (1.1 mg/kg caffeine) + CAF (5 mg/kg) (C), COF + CAF (3 mg/kg) (D), COF + CAF (7 mg/kg) (E), and colored water + CAF (5 mg/kg) (F). Times to exhaustion were significantly greater for all trials with CAF versus placebo (trial A). Exercise times (in minutes) were: 21.7 +/- 8.1, 29.0 +/- 7.4, 27.8 +/- 10.8, 25.1 +/-7.9, 26.4 +/- 8.0 and 26.8 +/- 8.1 for trials A through F, respectively. In conclusion, the prior consumption of COF did not decrease the ergogenic effect of the subsequent ingestion of anhydrous CAF. PMID- 15657473 TI - Nutrient intake and eating habits of triathletes on a Brazilian diet. AB - The objectives of this study were to determine the dietary habits, energy balance, and macro- and micronutrient dietary adequacy of triathletes, and to provide an athletes' food pyramid. Thirty-eight Brazilian triathletes, had anthropometric measurements taken. Mean (+/- standard deviation) body weight, height, and percentage body fat were 71.2 kg (+/- 9.4), 176.7 cm (+/- 5.5), and 12.3 % +/- ( 3.6) for men and 55.8 kg (+/- 5.2), 161.6 cm (+/- 4.5), and 24.3 % (+/- 4.2) for women, respectively. A 24-h recall and a food-frequency questionnaire showed that mean total intakes of energy (MJ), carbohydrate, protein, and fat (g x kg(-1) x d(- 1)) were, respectively, 15.4, 7.3, 2.0, and 1.6 for men and 9.9, 5.9, 1.6, and 1.3 for women. The number of meals and intake of some food groups were insufficient, resulting in inadequate intake of carbohydrate and some micronutrients. Based on this study, athletes need help to achieve their sports-related nutrition goals, especially during intense training. PMID- 15657475 TI - Food and macronutrient intake of elite kenyan distance runners. AB - The food and macronutrient intake of elite Kenyan runners was compared to recommendations for endurance athletes. Estimated energy intake (EI: 2987 +/- 293 kcal; mean +/- standard deviation) was lower than energy expenditure (EE: 3605 +/ 119 kcal; P < 0.001) and body mass (BM: 58.9 +/- 2.7 kg vs. 58.3 +/- 2.6 kg; P < 0.001) was reduced over the 7-d intense training period. Diet was high in carbohydrate (76.5%, 0.4 g/kg BM per day) and low in fat (13.4 %). Protein intake (10.1 %; 1.3 g/kg BM per day) matched recommendations for protein intake. Fluid intake was modest and mainly in the form of water (1113 +/- 269 mL; 0.34 +/- 0.16 mL/kcal) and tea (1243 +/- 348 mL). Although the diet met most recommendations for endurance athletes for macronutrient intake, it remains to be determined if modifying energy balance and fluid intake will enhance the performance of elite Kenyan runners. PMID- 15657477 TI - Protein nanotechnology: the new frontier in biosciences. AB - The combination of nanotechnology and molecular biology has led to a new generation of nanoscale-based devices and methods for probing the cell machinery and elucidating intimate life processes occurring at the molecular level that were heretofore invisible to human inquiry. This chapter provides a brief overview of the field of nanotechnology and its applications to the study, design, and use of protein systems in biology and medicine. PMID- 15657476 TI - Tricarboxylic-acid-cycle intermediates and cycle endurance capacity. AB - The purpose of this study was to determine whether ingestion of a multinutrient supplement containing 3 tricarboxylic-acid-cycle intermediates (TCAIs; pyridoxine alpha-ketoglutarate, malate, and succinate) and other substances potentially supporting the TCA cycle (such as aspartate and glutamate) would improve cyclists' time to exhaustion during a submaximal endurance-exercise test (approximately 70 % to 75 % VO2peak) and rate of recovery. Seven well-trained male cyclists (VO2max 67.4 2.1 mL x kg(-1) x in(-1), 28.6 +/- 2.4 y) participated in a randomized, double-blind crossover study for 7 wk. Each took either the treatment or a placebo 30 min before and after their normal training sessions for 3 wk and before submaximal exercise tests. There were no significant differences between the TCAI group (KI) and placebo group (P) in time to exhaustion during cycling (KI = 105 +/- 18, P = 113 +/- 11 min); respiratory-exchange ratio at 20 min intervals; blood lactate and plasma glucose before, after, and at 30-min intervals during exercise; perceived exertion at 20-min intervals during exercise; or time to fatigue after the 30-min recovery (KI = 16.1 +/- 3.2, P = 15 +/- 2 min). Taking a dietary sport supplement containing several TCAIs and supporting substances for 3 wk does not improve cycling performance at 75 % VO2peak or speed recovery from previously fatiguing exercise. PMID- 15657478 TI - Kinetics and mechanisms of protein crystallization at the molecular level. AB - This chapter focuses on the processes by which a protein molecule in a supersaturated solution joins a protein crystal. The pair of proteins ferritin/apoferritin is used as an example. The most commonly used technique in such investigations has been atomic force microscopy. I discuss the modifications and tests of the atomic force microscope necessary to obtain real-time, in situ molecular resolution imaging. Then, I review tests that establish a quantitative correspondence between the continuous models of crystal growth and the discrete nature of the processes at the molecular level. I address the issue of whether the incorporation of a molecule from a solution into a growth site on the crystal surface is limited by the slow rate of decay of an Eyring-type transition state. The conclusion is that, on the contrary, a scenario, envisioned by Smoluchowski and Debye, is followed in which the kinetics is only limited by the rate diffusion over the free-energy barrier of interaction between two molecules, or an incoming molecule and a surface. Review of the data for many other protein and nonprotein systems suggests that this conclusion is valid not only for the crystallization of ferritin/apoferritin, but also for many other protein and small-molecule crystallization systems. Finally, I review results establishing that the pathway that a molecule from a solution takes on its way to an incorporation site on the crystal surface is indirect: it includes adsorption on the surface and two-dimensional diffusion toward the incorporation site. These results will likely contribute to the understanding at the molecular level not only of the processes of crystallization of proteins and small molecules, but also of the deposition of layers of proteins and other soft materials on substrates, and of other processes of self-assembly in solution. PMID- 15657479 TI - Nanostructured systems for biological materials. AB - The sol-gel process is a chemical technique for immobilizing biomolecules in an inorganic, transparent matrix. The dopant biomolecules reside in an interconnected mesoporous network and become part of the nanostructured architecture of the entire material. In this chapter, we review the sol-gel immobilization approach and discuss how it leads to the stabilization of a number of proteins against aggressive chemical and thermal environments. We also review the sensor applications of this material that result from having analyte molecules diffuse through the matrix and reach the immobilized biomolecule. PMID- 15657480 TI - Nanomaterials of drug delivery systems for tissue regeneration. AB - A new therapeutic trial aimed at assisting tissue regeneration at a body defect in size too large for self-repair has recently begun. The objective is to substitute the biological functions of damaged and injured organs by taking advantage of cells. For successful tissue regeneration, it is absolutely indispensable not only to have cells of high proliferation and differentiation potential, but also to create an environment suitable for inducing regeneration. Such creation can be artificially achieved only by providing various biomaterials to promote cell proliferation and differentiation, such as cell scaffold and growth factors. Growth factors are often required to promote tissue regeneration because they can induce angiogenesis, which promotes a sufficient supply of oxygen and nutrients to effectively maintain the biological functions of cells transplanted for organ substitution. However, because of their poor in vivo stability, the biological effects of growth factors cannot always be expected unless these drug delivery systems (DDSs) are contrived. In this chapter, several research approaches to tissue regeneration are reviewed to emphasize the significance of biomaterials and DDS technologies in regenerative medicine. PMID- 15657481 TI - Nanotechnology with S-layer proteins. AB - The cross-fertilization of biology, chemistry, material sciences, and solid-state physics is opening up a great variety of new opportunities for innovation in nanosciences. One of the key challenges is the technological utilization of self assembly systems wherein molecules spontaneously associate under equilibrium conditions into reproducible supramolecular aggregates. The attractiveness of such processes lies in their capability to build uniform, ultrasmall functional units and the possibility of exploiting such structures at meso- and macroscopic scale for life and nonlife science applications. The use of crystalline bacterial cell-surface proteins (S-layer proteins) provided innovative approaches for the assembly of supramolecular structures and devices with dimensions of a few to tens of nanometers. S-layers have proven to be particularly suited as building blocks in a molecular construction kit involving all major classes of biological molecules. The immobilization of biomolecules in an ordered fashion on solid substrates and their controlled confinement in definite areas of nanometer dimensions are key requirements for many applications including the development of bioanalytical sensors, biochips, molecular electronics, biocompatible surfaces, and signal processing among functional membranes, cells, and integrated circuits. PMID- 15657482 TI - Folding of beta-structured fibrous proteins and self-assembling peptides. AB - Natural fibrous proteins include families found in natural materials such as wool and silk; in tissue components such as collagen and elastin; or in virus and bacteriophage adhesins. They have long fascinated scientists and engineers because of their mechanical and elastic properties, and considerable efforts have been made in order to produce artificial materials inspired from these natural proteins. The understanding of their structure, folding, and assembly properties is necessary in order to achieve these objectives. However, because of their complexity, structural information is quite often extremely difficult to obtain for these proteins. In this chapter, we focus on a particular family of fibrous proteins: trimeric, beta-stranded viral adhesins. We describe folding strategies that led to the identification of stable domains that could be crystallized, and the novel structural motifs that are emerging. We also discuss self-assembling peptides derived from these motifs. Finally, we review the possibilities of designing novel macroscopic materials as well as nanoscale fibrous objects with controlled dimensions and properties based on these novel structural motifs. PMID- 15657483 TI - Application of NMR methods to identify detection reagents for use in development of robust nanosensors. AB - Nuclear magnetic resonance (NMR) spectroscopy is a powerful technique for studying bimolecular interactions at the atomic scale. Our NMR laboratory is involved in the identification of small molecules, or ligands, that bind to target protein receptors such as tetanus neurotoxin (TeNT) and botulinum neurotoxin, anthrax proteins, and HLA-DR10 receptors on non-Hodgkin lymphoma cancer cells. Once low-affinity binders are identified, they can be linked together to produce multidentate synthetic high-affinity ligands (SHALs) that have very high specificity for their target protein receptors. An important nanotechnology application for SHALs is their use in the development of robust chemical sensors or biochips for the detection of pathogen proteins in environmental samples or body fluids. Here we describe a recently developed NMR competition assay based on transferred nuclear Overhauser effect spectroscopy that enables the identification of sets of ligands that bind to the same site, or a different site, on the surface of TeNT fragment C (TetC) than a known "marker" ligand, doxorubicin. Using this assay, one can identify the optimal pairs of ligands to be linked together for creating detection reagents, as well as estimate the relative binding constants for ligands competing for the same site. PMID- 15657484 TI - Studying 3D subdomains of proteins at the nanometer scale using fluorescence spectroscopy. AB - Databases devoted to the crystal structure of proteins have dramatically increased in size during the last two decades. Moreover, X-ray and NMR technology studies have shown that proteins belonging to the same family generally share the same global 3D architecture. These results suggest that the need for experimental determination of protein structure will be reduced to those that are suspected to have sufficiently novel structures. Furthermore, NMR and other techniques have demonstrated that a protein in solution experiences constant random thermal motions that occur over large time scales, ranging from picoseconds to seconds and perhaps hours. Such changes may have important functional consequences, but identifying which changes are functionally relevant remains a difficult task even if this problem has been addressed both with experimental and computational methods. For that specific purpose, there is a need for methods allowing a fast and accurate monitoring of conformation changes (that occur at specific sub domains of proteins. Fluorescence resonance energy transfer (FRET) is a suitable tool for monitoring conformational changes at the nanoscale level. This chapter describes the various FRET methods that are used for monitoring the 3D sub-domain conformation of proteins in solution, in single living cells and at the single molecular level. PMID- 15657485 TI - Carbon nanotubes and nanowires for biological sensing. AB - This chapter reviews the recent development in biological sensing using nanotechnologies based on carbon nanotubes and various nanowires. These 1D materials have shown unique properties that are efficient in interacting with biomolecules of similar dimensions, i.e., on a nanometer scale. Various aspects including synthesis, materials properties, device fabrication, biofunctionalization, and biological sensing applications of such materials are reviewed. The potential of such integrated nanobiosensors in providing ultrahigh sensitivity, fast response, and high-degree multiplex detection, yet with minimum sample requirements is demonstrated. This chapter is intended to provide comprehensive updated information for people from a variety of backgrounds but with common interests in the fast-moving interdisciplinary field of nanobiotechnology. PMID- 15657486 TI - Carbon nanotube systems to communicate with enzymes. AB - The efficient transfer of electrons between enzymes and electrodes is important for understanding the intrinsic redox properties as well as for developing protein-based biosensors and bioelectronic devices. One strategy to achieve efficient electron transfer to proteins is to build up the electrode inside the protein so that it is close to the redox-active center of the protein. To achieve this requires exceedingly small electrodes. Carbon nanotubes, which are as small as 1 nm in diameter, have the potential to be such electrodes. This chapter outlines recent research toward this goal via the self-assembly of vertically aligned single-walled carbon nanotubes on electrode surfaces followed by the subsequent attachment of proteins to the free ends of the tubes. PMID- 15657487 TI - Molecularly imprinted polymers for biomolecular recognition. AB - Molecular imprinting of polymers is a concept for the synthetic formation of structurally organized materials providing binding sites with molecular selectivity. Compared to biological receptors, these polymeric recognition systems have the advantage of superior chemical and mechanical stability with potential applications in areas such as biomimetic catalysis and engineering, biomedical analysis, sensor technology, or the food industry. In particular, molecularly imprinted polymers (MIPs) providing selectivity for biorelated molecules are gaining substantial importance. In this context, a self-assembly approach for the synthesis of imprinted polymers against the flavonol quercetin is presented, which is exemplary for the biologically relevant group of flavonoid compounds. The creation of synthetic selective recognition sites for this biomolecule is demonstrated by comparing the separation capabilities of imprinted and nonimprinted polymer particles for several structurally related molecules via high-performance liquid chromatography experiments. The developed quercetin-MIP enables selective extraction of quercetin even from complex mixtures, demonstrating the potential for designing biomimetic recognition materials with improved selectivity for biomolecules with tunable functionality at a nanoscale. PMID- 15657488 TI - Plasmonics-based nanostructures for surface-enhanced Raman scattering bioanalysis. AB - Surface-enhanced Raman scattering (SERS) spectroscopy is a plasmonics-based spectroscopic technique that combines modern laser spectroscopy with unique optical properties of metallic nanostructures, resulting in strongly increased Raman signals when molecules are adsorbed on or near nanometer-size structures of special metals such as gold, silver, and transition metals. This chapter provides a synopsis of the development and application of SERS-active metallic nanostructures, especially for the analysis of biologically relevant compounds. Some highlights of this chapter include reports of SERS as an immunoassay readout method, SERS gene nanoprobes, near-field scanning optical microscopy SERS probes, SERS as a tool for single-molecule detection, and SERS nanoprobes for cellular studies. PMID- 15657489 TI - Bacterial virus phi29 DNA-packaging motor and its potential applications in gene therapy and nanotechnology. AB - A controllable, 30-nm imitating DNA-packaging motor was constructed. The motor is driven by six synthetic adenosine triphosphate (ATP)-binding RNA (packaging RNA [pRNA]) monomers, similar to the driving of a bolt with a hex nut. Conformational change and sequential action of the RNA with fivefold (viral capsid)/sixfold (pRNA hexamer) mismatch could ensure continuous rotation of the motor with ATP as energy. In the presence of ATP and magnesium, a 5-microm synthetic DNA was packaged using this motor. On average, one ATP was used to translocate two bases of DNA. The DNA-filled capsids were subsequently converted into up to 109 PFU/mL of infectious virus. The three-dimensional structures of pRNA monomer, dimer, and hexamer have been probed by photoaffinity crosslinking, chemical modification interference, cryo-atomic force microscopy, and computer modeling. The pRNA's size and shape can be controlled and manipulated at will to form stable dimers and trimers. Cryo-atomic force microscopy revealed that monomers, dimers, and trimers displayed a checkmark outline, elongated shape, and triangular structure, respectively. The motor can be turned off by gamma-S-ATP or EDTA and turned on again with the addition of ATP or magnesium, respectively. The formation of ordered structural arrays of the motor complex and its components, the retention of motor function after the 3'-end extension of the pRNA, and the ease of RNA dimer, trimer, and hexamer manipulation with desired shape and size make this RNA containing motor a promising tool for drug and gene delivery and for use in nanodevices. PMID- 15657490 TI - Construction of ordered protein arrays. AB - Artificially ordered protein arrays provide a facile approach to a variety of problems in biology and nanoscience. Current demonstration systems use either nucleic acid tethers or methyltransferase fusions in order to target proteins or peptides of interest to nucleic acid scaffolds. These demonstrations point to the large number of useful devices and assemblies that can be envisioned using this approach, including smart biological probes and drug delivery systems. In principle, these systems are now capable of imitating the earliest forms of prebiotic organisms and can be expected to reach the complexity of a small virus in the near future. Third-generation methyltransferase inhibitors provide an example of a smart chemotherapeutics that can be constructed with this approach. We describe the use of mechanistic enzymology, computer-aided design, and microfluidic chip-based capillary electrophoresis in assessing the final assembly and testing of designs of this type. PMID- 15657491 TI - Bioengineering and characterization of DNA-protein assemblies floating on supported membranes. AB - This chapter describes the design, practical construction, and characterization of P-DNA and their applications in building a new generation of DNA chips. P-DNAs are artificial covalent assemblies involving a histidine tag head able to bind to modified phospholipids, a core protein domain derived from cytochrome b5 by genetic engineering that features specific spectroscopic and electrochemical properties useful for detection, a synthetic linker acting as a spacer, and an oligonucleotide acting as a probe. P-DNA has the property of being able to efficiently self-associate to a supported bilayer including nickel-iminodiacetate modified phospholipids. The construction of P-DNA and its interaction with a complementary oligonucleotide sequence can be monitored in real time by surface plasmon resonance using a Biacore system or equivalent. P-DNA chips feature unique properties including tunable surface density of probes; very low nonspecific interaction with external DNA; lateral mobility, minimizing-steric interaction; optimization of hybridization efficiency; and, potentially, recognition by multiple probes of a single target and perfectly defined and homogeneous structure, permitting high density up to a compact monolayer. Potential applications of this new device are multiple, including high sensitivity and high-selectivity chips for DNA-DNA, DNA-RNA, or DNA-protein interactions. PMID- 15657492 TI - Nanosystems for biosensing: multianalyte immunoassay on a protein chip. AB - This chapter describes the construction of addressable two-dimensional (2D) microarrays via the random fluidic self-assembly of metallic particles and the use of these arrays as platforms for constructing protein chips for bioassays. These arrays will be useful as platforms for constructing protein chips for bioassays in a broad range of applications. The basic units in the assembly are microfabricated particles, which carry a straightforward visible code, and the corresponding array template patterned on a glass substrate. On one face, the particles consist of a hydrophobic and magnetic Ni-polytetrafluoroethylene (Ni PTFE) composite layer; the other face has a gold layer that was modified for biomolecular attachment. We use photoresist patterning to create an array template with spatially discrete microwells into which an Ni-PTFE hydrophobic composite layer and a hydrophobic photoadhesive coating are electrodeposited. After biomaterial attachment and binding processes in bulk, the particles are randomly self-assembled onto the lubricated bonding sites on the chip substrate. This self-assembly process is driven by a combination of magnetic, hydrophobic, and capillary interactions. The encoding symbol carried by each particle is used to identify the target attached to the particle surface. This model system demonstrates the utility of the protein chip array for conducting simultaneous multianalyte immunoassays of human immunoglobulins (IgA, IgG, and IgM). PMID- 15657493 TI - Optical nanosensors for detecting proteins and biomarkers in individual living cells. AB - Recently, nanotechnology has been revolutionizing important areas in molecular biology and medicine, especially diagnostics and therapy at the molecular and cellular levels. The combination of nanotechnology, biology, advanced materials, and photonics opens up the possibility of detecting and manipulating atoms and molecules using nanodevices. This capability has the potential for a wide variety of medical uses at the cellular level. One of the most recent technological advances has been in the area of nanosensors. This chapter describes the principle of optical nanosensors, their development, and their applications for in vivo analysis of proteins and biomarkers in individual living cells. Nanosensors were fabricated with optical fibers pulled down to tips with distal ends in nanoscale dimensions. Nanosensors with immobilized bioreceptor probes (e.g., antibodies, enzyme substrate) that are selective to target analyte molecules are also referred to as nanobiosensors. Laser light is launched into the fiber, and the resulting evanescent field at the tip of the fiber is used to excite target molecules bound to the antibody molecules. A photometric detection system is used to detect the optical signal (e.g., fluorescence) originating from the analyte molecules or from the analyte-bioreceptor reaction. PMID- 15657494 TI - Nanoelectrodes integrated in atomic force microscopy cantilevers for imaging of in situ enzyme activity. AB - For investigation of laterally resolved information on biological activity, techniques for simultaneously obtaining complementary information correlated in time and space are required. In this context, recent developments in scanning probe microscopy are aimed at information on the sample topography and simultaneously on the physical and chemical properties at the nanometer scale. With the integration of submicro- and nanoelectrodes into atomic force microscopy (AFM) probes using microfabrication techniques, an elegant approach combining scanning electrochemical microscopy with AFM is demonstrated. This instrumentation enables simultaneous imaging of topography and obtainment of laterally resolved electrochemical information in AFM tapping mode. Hence, topographical and electrochemical information on soft surfaces (e.g., biological species) and polymers can be obtained. The functionality of tip-integrated electrodes is demonstrated by simultaneous electrochemical and topographical studies of an enzyme-modified micropattern. PMID- 15657496 TI - Near-field scanning optical microscopy for bioanalysis at nanometer resolution. AB - The nondestructive imaging of biomolecules in nanometer domains in their original location and position as adsorbed or deposited on a surface is of garners considerable experimental interest. Near-field scanning optical microscopy (NSOM) is an emerging technique with its astonishing resolving power of <100-nm domains, and nondestructive nature compared with other scanning probe microscopic techniques is an emerging technique to achieve this goal. At the single-molecule level of resolution, it is possible to use the NSOM as a critical tool for visualization of proteins on surfaces to obtain more fundamental information about their orientation and locality without disturbing their original orientation and position, and level of interaction with the surface. Several areas of science and medicine can benefit from this type of study especially for biomedical and biochip applications. To illustrate possible applications, imaging of green fluorescent proteins and biomolecules associated with multidrug resistance proteins in tumor cells will be demonstrated using NSOM. PMID- 15657495 TI - Protein amyloidose misfolding: mechanisms, detection, and pathological implications. AB - A variety of diseases result because of misfolded protein that deposits in extracellular space in the body. These deposits can be amorphous (disordered) or fibrillar (ordered). Inclusion bodies are an example of amorphous aggregates, and amyloid fibril is an example of fibrillar or ordered aggregates. In this chapter, we discuss a class of diseases caused by fibrillar aggregate deposits or amyloid fibrils called amyloidosis. We also review mechanisms by which different proteins misfold to form amyloid fibrils. Each amyloid fibril formed from a different protein causes a different disease by affecting a different organ in the body. However, the characteristics of different amyloid fibrils, namely structure and morphology, observed by electron microscopy and X-ray fiber diffraction appear to be quite similar in nature. We present therapeutic strategies developed to eliminate amyloid fibril formation. These strategies could possibly avert a whole class of fatal diseases caused by amyloid fibril deposition owing to similar characteristics of the amyloid fibrils. PMID- 15657498 TI - A new glossary for nuclear medicine. PMID- 15657499 TI - Imaging the dopaminergic system in suspected parkinsonism, drug induced movement disorders, and Lewy body dementia. AB - This review discusses the role of pre- and post-synaptic dopaminergic imaging in suspected Parkinson's disease, vascular parkinsonism, the parkinsonian syndromes, drug induced movement disorder, and Lewy body dementia. It is envisaged that this information may be useful to neurologists, psychiatrists, physicians/geriatricians, radiologists and nuclear medicine physicians. PMID- 15657500 TI - Usefulness of meta-[123I]iodobenzylguanidine myocardial scintigraphy for predicting cardiac events in patients with dilated cardiomyopathy who receive long-term beta blocker treatment. AB - OBJECTIVE: Large-scale clinical trials have demonstrated that beta blocker therapy improves the prognosis of dilated cardiomyopathy (DCM), but cardiac events sometimes occur even in patients showing favourable response to the therapy. In this study, the usefulness of meta-iodobenzyguanidine (MIBG) in predicting cardiac events was investigated in DCM patients successfully receiving long-term treatment with beta blockers. METHODS: The subjects were 53 patients with DCM (including 10 women; mean age, 56.5+/-10.9 years) who could continue beta blocker therapy for more than 6 months. MIBG was performed within 1 year of commencing the therapy. The extent score, severity score and washout rate were obtained from single photon emission computed tomography images. At the same time, left ventricular ejection fraction (LVEF) and left ventricular end diastolic diameters were measured by echocardiography. The endpoints were cardiac events (cardiac death and admission to hospital due to heart failure or arrhythmia), and patients were observed for an average of 1314+/-986 days (150 4100 days). RESULTS: Cardiac events occurred in nine patients during the observation period. The multivariate statistical analysis demonstrated that the delayed extent score was the strongest significant predictive factor, (hazard ratio 1.036, P<0.01). while LVEF was not a predictive factor. Both the improvement of LVEF and MIBG were significant predictive factors. The improvement of washout rate was the strongest. CONCLUSION: Parameters of MIBG but not of LVEF were useful in predicting cardiac events in DCM patients whose condition had been successfully stabilized by the introduction of beta blockers. PMID- 15657501 TI - Left ventricular function parameters obtained from gated myocardial perfusion SPECT imaging: a comparison of two data processing systems. AB - BACKGROUND AND AIM: The Cedars-Sinai Quantitative Gated Single Photon Emission Computed Tomography (SPECT) (QGS) program, used to quantify left ventricular function parameters from gated myocardial perfusion scintigraphy (MPS), has been extensively validated and compared with other methods of quantification. However, little is known about the reproducibility of QGS on different processing systems. This study compared the findings of QGS running on workstations provided by two different manufacturers. METHODS: Gated rest MPS studies of 50 patients were analysed retrospectively. Filtered back-projection (FBP) was performed using identical parameters on Philips Pegasys and Nuclear Diagnostics Hermes workstations to produce gated short-axis (SA) slices. In addition, the gated SA slices reconstructed on the Pegasys were transferred to the Hermes. QGS was used to calculate the end-diastolic volume (EDV), end-systolic volume (ESV) and left ventricular ejection fraction (LVEF) in each case. RESULTS: The mean+/-standard deviation differences between the Pegasys and Hermes function parameters were 7.06+/-3.91 ml (EDV), -5.54+/-3.21 ml (ESV) and +1.14%+/-1.43% (LVEF) when data were reconstructed on different systems, and -0.16+/-1.58 ml (EDV), -0.10+/-1.02 ml (ESV) and +0.14%+/-0.73% (LVEF) when data were reconstructed on the same system. Bland-Altman plots showed definite trends for EDV and ESV for data reconstructed on different systems, but no trends were seen for data reconstructed on the same system. CONCLUSIONS: When data were reconstructed on two separate systems, the difference between the function parameters obtained from Pegasys and Hermes could be ascribed to differences in the reconstruction process on each system despite the use of identical parameters (filters, etc). However, when the same reconstructed data were analysed on both systems, no significant difference in left ventricular function parameters was observed. PMID- 15657503 TI - Prone SPECT myocardial perfusion imaging is associated with less cardiac drift during the acquisition duration than imaging in the supine position. AB - BACKGROUND AND AIM: Cardiac cranial drift, a slow vertical upward displacement seen during the acquisition of myocardial single photon emission tomography (SPECT), is a source of image artefacts that may lead to erroneous interpretation. Changes in breathing pattern and depth throughout image acquisition are believed to cause cardiac cranial drift. As the physiology of respiration probably differs with postural changes, we hypothesized that cardiac drift may be different for supine vs. prone acquisitions. Our aim was to assess the magnitude of cardiac displacement for prone and supine SPECT acquisitions in patients undergoing stress myocardial perfusion imaging. METHODS: We enrolled prospectively 15 subjects undergoing exercise myocardial perfusion imaging. Subjects had post-stress images acquired in both the prone and supine positions. Motion was assessed in the horizontal (x) and vertical (y) axes for both camera heads at all 64 projections at which images were obtained. Pixel displacement (number of pixels from the baseline of zero) in either the cranial/caudal or left/right direction was quantified using the automated camera motion correction algorithm. RESULTS: Supine imaging was associated with more cranial drift than prone imaging (1.20+/-0.40 pixels vs. 0.92+/-0.24 pixels, P<0.05). There was no significant difference in cardiac displacement in the horizontal axis (1.03+/-0.5 pixels vs. 1.12+/-0.22 pixels, P=NS). CONCLUSIONS: Prone imaging is associated with less cardiac cranial drift than imaging in the supine position, suggesting that the former is associated with a more constant and reliable diaphragmatic breathing pattern. Acquisitions in the prone position may thus be associated with fewer motion artefacts than supine acquisitions for cardiac SPECT imaging. PMID- 15657502 TI - The influence of post-exercise cardiac changes on thallium-gated myocardial perfusion scintigraphy findings in normal subjects. AB - BACKGROUND AND AIM: During recovery after exercise, the heart rate and blood pressure return to a resting state more rapidly than the end-systolic left ventricular dimensions and fractional shortening. The aim of this study was to assess how exercise-related cardiac changes affect the interpretation of myocardial perfusion images in normal subjects. Systolic cardiac parameters on gated stress and rest images were evaluated in healthy young and elderly subjects. METHODS: Twenty-six healthy young and 20 healthy elderly subjects participated in the study. An injection of 111-130 MBq of thallium-201 (201Tl) was given at peak exercise. Rest images were acquired 2.5 h after stress acquisition, 15 min after a second injection of 18.5-37 MBq of 201Tl. Data were analysed using automatic-processing software for quantitative gated single photon emission computed tomography (SPECT) (QGS). The parameters derived from QGS were the end-systolic volume (ESV), end-diastolic volume (EDV), left ventricular ejection fraction (LVEF), end-systolic surface area (ESSA) and end-diastolic surface area (EDSA). The difference between wall thickening in the basal and apical segments (Delta WT) was also calculated. Perfusion images were visually assessed for differences in cardiac size, evidence of reversible hypoperfusion and hot spots. RESULTS: In the young group, LVEF was approximately 6% higher at stress than at rest. EDV, ESV, ESSA and EDSA were all significantly lower, and Delta WT was significantly higher, at stress than at rest. In the elderly group, the mean LVEF at stress was slightly higher than the finding at rest (P<0.05). Visual evaluation of perfusion images revealed mild reversible stress hypoperfusion in the inferoseptal region in eight young male subjects. CONCLUSIONS: In healthy young subjects, post-exercise cardiac changes affect systolic functions detected on gated thallium myocardial perfusion scintigraphy, resulting in a smaller heart size during stress. This finding, accompanied by a significant difference in apex to base counts during stress, may cause basal portions of the heart to appear ischaemic. The absence of these findings in the elderly suggests a decrease in contractility with age. PMID- 15657504 TI - Illustrations of abdominal abnormalities on 99mTc tetrofosmin gated cardiac SPECT. AB - BACKGROUND: 99mTc tetrofosmin is predominantly cleared by the hepatobiliary system and its scan is analogous to a cholescintigraphy. The radiopharmaceutical may be localized in the bone marrow. During cardiac acquisition, when the area being viewed includes the lower thorax and abdomen, abnormalities of the liver, gallbladder and kidneys, and the radioactivity in the oesophagus, stomach and bowel, as well as bone marrow uptake, can be depicted by routine raw data cine images. Excluding the intrathoracic abnormalities, these abdominal abnormalities are illustrated in this paper. METHODS AND MATERIALS: Retrospectively, we reviewed the raw data cine images from 566 patients. These images were 180 degrees or 360 degrees rotation and showed the abdominal organs and thoracic cage, including the ribs, sternum and spine. We found 234 abnormalities, which were correlated with radiography, laboratory and clinical findings. RESULTS: Intra-abdominal abnormalities included bone marrow visualization, 93 occurrences (39.7%); duodeno-gastric (DG) and entero-gastric (EG) reflux, 47 (20.1%); non visualization of the gallbladder, including possible causes of cholelithiasis, acute cholecystitis, or cholecystectomy, 31 (13.2%); elevation of the right diaphragm and depression of diaphragm, 30 (12.8%); renal abnormalities, including absent kidney, renal atrophy and cyst, 12 (5.1%); splenomegaly, 10 (4.3%); liver, including hepatomegaly and cirrhosis of the liver, eight (3.4%); and breast attenuation resulting in photopenia in the liver, three (1.2%). CONCLUSIONS: (1) The bone marrow uptake of the vertebrae accounts for almost 40% of the abdominal abnormalities, and is usually concordant with anaemia; (2) DG and EG refluxes represent 20% of abdominal abnormalities; they may have a clinical impact and may or may not interfere with SPECT processes and result in artifacts of the inferior wall of the left ventricle; (3) non-visualization of the gallbladder should prompt a search for cholelithiasis; and (4) benign renal cyst or cirrhosis of the liver could also be demonstrated. These coincidental findings may be included in routine cardiac SPECT reports that may have clinical impact because they give the referring physician the chance to pursue further clinical investigation. PMID- 15657505 TI - Ablative treatment of thyroid cancer with high doses of 131I without pre-therapy scanning. AB - OBJECTIVE: To determine the success of ablative treatment using fixed high doses of 131I in patients with thyroid cancer. METHODS: A retrospective study was conducted on 274 patients who received ablative treatment without previous scanning, with the dose being based on surgical staging: stage I patients (tumour restricted to the thyroid) received 3.7 GBq, and stage II (lymph node metastases) and stage III patients (extra-thyroid invasion) were treated with 5.5 GBq. Successful treatment was defined as a negative control scan. RESULTS: One hundred and sixty patients were classified as stage I and 114 as stages II or III. Forty six patients presented ectopic uptake on post-therapy scans (10% in stage I and 26% in stage II or III). Among stage I patients, the efficacy of treatment was 78.7%. A 47% failure rate was associated with metastases, and among patients without metastases who did not respond to treatment, thyroid bed uptake >5% on post-therapy scans was observed in 61%. Patients with stage II or III showed 62.2% efficacy. Twenty-three of the 43 (53%) unsuccessfully treated patients in this group had metastases and of the 20 patients without metastases, 14 (70%) presented thyroid bed uptake >5%. There were no differences between the responsive and non-responsive groups in terms of age, gender, histological type or size of the primary tumour. CONCLUSION: Empirical treatment presented 72% efficacy (higher in stage I) and failure was associated with the presence of metastases and large thyroid remnants; factors that can be evaluated by pre therapy whole-body scanning. PMID- 15657506 TI - Localization of parathyroid adenomas using 11C-methionine positron emission tomography. AB - BACKGROUND AND AIM: In symptomatic hyperparathyroidism, pre-surgical localization of the suspected site of adenoma is desirable. All widely available techniques may have difficulty in localizing the site. The aim of this study was to determine whether 11C-methionine positron emission tomography (PET) could accurately localize parathyroid adenomas in patients in whom conventional imaging had failed. PATIENTS AND METHODS: Fifty-one patients presenting with hyperparathyroidism, and in whom other imaging techniques had failed to definitely identify the site of adenoma, were reviewed retrospectively after 11C methionine PET scanning. Patients were followed up by surgical histology, or clinically if surgery was not performed. RESULTS: 11C-Methionine PET scanning was found to have a sensitivity of 83%, a specificity of 100% and an accuracy of 88% in successfully locating parathyroid adenomas. Most false negatives were due to adenomas in the lower mediastinum that was outside the area of scanning. CONCLUSIONS: 11C-Methionine PET is a reliable and highly accurate technique for localizing parathyroid adenomas in patients in whom conventional imaging techniques have failed. It is necessary to image to the level of the lower mediastinum. PMID- 15657507 TI - How often do patients undergo repeat PET or PET/CT examinations? Experience from a UK institution. AB - BACKGROUND AND AIM: According to the report of the Intercollegiate Standing Committee on Nuclear Medicine, the UK requires 40-60 positron emission tomography (PET) machines in the next decade (Intercollegiate Standing Committee on Nuclear Medicine). Positron Emission Tomography: a Strategy for Provision in the UK. London: Royal College of Physicians of London; 2003, pp. 1-9). This figure is based mainly on patients receiving only one examination and restricting the clinical indication to three primary diagnoses. The aim of this study was to assess the appropriateness of this figure and the assumptions made in the Intercollegiate report on UK PET provision. METHODS: We examined retrospectively our institution's entire PET and PET/computed tomography (CT) database, which spans 4 years and 9 months. We recorded the number of patients who received repeat examinations. RESULTS: Reports were available for 3354 PET/CT or PET-only studies; 418 of 2268 patients (18.4%) received at least one repeat PET/CT examination. The three main indications for PET examination in the Intercollegiate report only accounted for approximately 60% of the examinations undertaken. CONCLUSION: Our records suggest that basing the UK's future PET provision on a single examination and on three clinical indications only is no longer realistic. PMID- 15657508 TI - The conditions for which the geometric mean method revealed a more accurate calculation of relative renal function in 99mTc-DMSA scintigraphy. AB - AIMS: (1) To compare the results of calculating relative renal function (RRF) by using only posterior images (POST) with the geometric mean (GM) through both anterior and posterior imaging on dimercaptosuccinic acid (DMSA) scintigraphy. (2) To determine whether there was an age-related difference between them and whether some renal pathologies or asymmetrical renal function cause an error in the RRF calculation by using posterior images only. METHODS: Eight hundred and ninety-one DMSA scans were studied retrospectively. The patients were divided into five age groups: group I, < or =2 years; group II, >2 to < or =5 years; group III, >5 to < or =10 years; group IV, >10 to < or =18 years; and group V, >18 years. The RRF of the right kidney (RKF) was calculated from the POST and GM counts. The differences between RKFGM and RKFPOST were calculated in all the patients. RESULTS: Among the 891 patients, nine had malrotated or malpositioned kidneys, 373 had renal pathologies of pyelonephritis, hydronephrosis, cortical scarring and atrophy, 247 had asymmetrically functioning kidneys and 509 had normal kidneys. When the patients were analysed according to different age groups, significant differences were found between all groups (P<0.05) except groups I-II and IV-V (P>0.05) with the F-test. The clinically meaningful RRF variance (> or =5% difference between two methods) rate differed significantly between groups I, II and III, and groups IV and V (chi-squared test, P<0.05). In patients aged < or =10 years, a clinically meaningful RRF variance (> or =5%) rate was significantly higher in the groups with pathological or asymmetrically low (< or =40% RRF) functioning kidneys than in the groups without pathological or asymmetrically low functioning kidneys, respectively (P<0.05). CONCLUSIONS: According to our findings, the calculation of RRF using the GM method differs significantly from that using posterior images. Calculation of the GM can effectively correct the RRF measurement not in only adults, but also in the patient population musical #10 years of age. In addition, a significant correction can be made in patients aged musical #10 years who have a renal pathology or an asymmetrically low functioning kidney. PMID- 15657509 TI - Synthesis, biodistribution and effects of farnesyltransferase inhibitor therapy on tumour uptake in mice of 99mTc labelled epidermal growth factor. AB - OBJECTIVE: The goal of this study was to develop a 99mTc labelled human epidermal growth factor (hEGF) for the in-vivo prediction of cancer cell response to farnesyltransferase inhibitor (FTI) therapy. This is based on the observation that internalization of EGF receptors is inhibited by FTIs. METHODS: We describe the radiolabelling of 99mTc-hEGF using the hydrazinonicotinamide (HYNIC) linker. Binding characteristics of 99mTc-HYNIC-hEGF to the EGF receptor are explored using an in-vitro binding assay. Biodistribution data of the compound in mice and tumour uptake in LoVo tumour bearing athymic mice before and after farnesyltransferase inhibitor therapy are presented. RESULTS: No colloid formation was observed. Binding parameters and LoVo tumour uptake of 99mTc-HYNIC hEGF did not differ significantly from directly labelled 123I-hEGF values. However, the biodistribution data of the 99mTc-HYNIC-hEGF showed higher uptake in liver and intestines and decreased stomach uptake compared to its 123I analogue. Eight hours after farnesyltransferase inhibitor therapy with R115777, LoVo tumour uptake of 99mTc-HYNIC-hEGF decreased significantly, as shown using planar gamma scintigraphy (the ratio tumour vs. thigh dropped from 2.54+/-0.83 to 0.99+/ 0.18). These data confirm the results obtained using 123I-hEGF. CONCLUSION: These data suggest that 99mTc-HYNIC-hEGF is a promising and selective new radiotracer for in-vivo monitoring of the EGF receptor with SPECT. Moreover, 99mTc-HYNIC-hEGF is a possible tool for early therapy response prediction of farnesyltransferase inhibitors. PMID- 15657510 TI - 99mTc carbonyl t-butyl isonitrile: a potential new agent for myocardial perfusion imaging. AB - OBJECTIVE: The recent development of mixed ligand complexes using a 99mTc tricarbonyl synthon has prompted us to revisit the first generation product, 99mTc-t-butyl isonitrile (TBI), for possible myocardial imaging after modification of the 99mTc core to a mixed ligand core of carbonyl and t-butyl isonitrile. The easy availability of TBI from commercial sources and the recent promising development of a 'kit' procedure to prepare the 99mTc tricarbonyl aqua synthon/precursor [99mTc(H2O)3(CO)3]+ were other factors that triggered this work. METHODS: The carbonyl precursor (37-370 MBq/0.5 ml) was synthesized and reacted with TBI (3 mg.ml-1) at room temperature and at pH 8 for 1 h. [99mTc(CO)3(TBI)3]+ was characterized by C18 reverse phase HPLC in gradient mode with water and acetonitrile as solvent. Biodistribution studies were carried out in normal mice and planar images were acquired in rabbit at 5 min, 30 min, 1 h, 2 h and 4 h post-injection to assess heart uptake and soft tissue retention. [99mTc(CO)3(TBI)3]+was formed as a single species in >95% yield and was found to be stable. Biodistribution studies in mice revealed 2.3 (+/-0.2)% uptake in heart at 5 min p.i. with heart/liver, heart/lung and heart/blood ratios of 1.5, 2.0 and 30, respectively. Imaging studies in rabbits showed high uptake in myocardium, with negligible activity in blood and lungs, at 5 min p.i. that washed out of the heart after 4 h. CONCLUSION: [99mTc(CO)3(TBI)3]+ could be prepared in >95% yields. The complex showed high myocardial uptake with desirable rate of washout from heart in rabbits. [99mTc(CO)3(TBI)3]+ has potential to extend to larger animal studies and later for clinical evaluation as a myocardial imaging agent. PMID- 15657511 TI - A comparison of techniques for analysing 90Y-Zevalin thin-layer chromatography plates. AB - OBJECTIVES: To simulate 90Y-Zevalin thin-layer chromatograms representing a range of radiochemical purities, to compare the radiochemical purities obtained with five techniques used to quantify 90Y on the plates and to measure the reproducibility of the five techniques at the minimum acceptable radiochemical purity of 95%. METHODS: Yttrium-90 solutions were pipetted onto the origin and solvent front lines of thin-layer chromatography (TLC) plates to simulate radiochemical purities of 90%, 92%, 94%, 95%, 96%, 98% and 100%. Each plate was analysed using three TLC scanners (Bioscan AR2000, Bioscan Mini-scan and an instrument constructed in-house) and two cut-and-count techniques: one using a sodium iodide well detector and the other a liquid scintillation counter. The reproducibility of each technique was measured by analysing the 95% plate 10 times. RESULTS: The radiochemical purities measured by the five techniques agreed well. The means of the seven results obtained with each agreed within 0.7%. The reproducibility of each technique was excellent. The coefficient of variation for 10 measurements was < or =0.3%. The signal to background ratios were satisfactory, ranging from 24 to 2.1 x 10(5). CONCLUSION: Each technique is suitable for analysing 90Y-Zevalin TLC plates. PMID- 15657512 TI - Implications of the UK NHS consent policy for nuclear medicine practice. AB - To comply with government policy on consent, the Sheffield Teaching Hospitals (STH) National Health Service (NHS) Trust introduced a new consent policy in February 2002. Verbal or written consent (depending on the level of risk) must be obtained prior to each study. The patient must be fully informed and given time to reach a decision. Consideration needs to be given to the following: to whom, when and how to provide such information and obtain consent. Each study type and patient circumstance needs to be classified according to risk. Consideration of the risks resulted in a local policy in which written consent is required for the following: therapeutic procedures, studies on pregnant women, studies in which pregnancy needs to be avoided, research procedures, cardiac stress for myocardial perfusion scintigraphy and intrathecal administration. Patient information leaflets have been updated with new information about the study and any risks. Information is now available for both patients and hospital staff. Compliance with the consent policy in a service department provides logistic challenges, but it is possible to fully inform patients in advance about their treatment, allowing them to give informed consent. PMID- 15657513 TI - Guidelines for the measurement of glomerular filtration rate using plasma sampling. PMID- 15657516 TI - History of clinical trials for breast cancer in Kyushu and Okinawa. PMID- 15657517 TI - Prediction of response to docetaxel by immunohistochemical analysis of CYP3A4 expression in human breast cancers. AB - BACKGROUND: Docetaxel (DOC) is inactivated by CYP3A4, high expression of which in tumor tissue might serve as a resistance mechanism. In the present study, the CYP3A4 protein level in breast cancers was determined by immunohistochemistry, and its relationship with the response to DOC treatment was studied. MATERIALS AND METHODS: Thirty-one patients with locally advanced (n = 21) or recurrent (n = 10) breast cancers underwent tumor biopsy, followed by DOC treatment (60 mg/m2 q3w). Expression of CYP3A4 was studied by immunohistochemistry. RESULTS: Patients with CYP3A4 negative tumors (n = 15) by immunohistochemistry showed a significantly (P < 0.01) higher response rate (67%) to DOC treatment than those with CYP3A4 positive tumors (n = 16, 19%). The positive predictive value, negative predictive value, and diagnostic accuracy of CYP3A4 expression by immunohistochemistry in the prediction of response to DOC were 67%, 81%, and 74%, respectively. CONCLUSION: Immunohistochemical analysis of CYP3A4 expression in tumor cells might be clinically useful in the prediction of tumor response to DOC. PMID- 15657518 TI - Breast conserving surgery with primary volume replacement using a lateral tissue flap. AB - Breast conserving surgery (BCS) is now a standard surgical treatment for early breast cancer. The number of patients with tumors under 3 cm who underwent breast conserving surgery overtook the number of patients who underwent total mastectomy for the first time in Japan in 2003. We have been employing breast conserving surgery with primary reconstruction using a lateral tissue flap (LTF), and have performed breast conserving surgery for 266 patients from 1990 to 2002. The incidence of local relapse was 5.6%. Although we did not irradiate a low risk group of 101 patients, our method is not inferior to other reports in which all cases underwent irradiation. Primary reconstruction with LTF has three advantages. The first is that we can avoid poly-surgery for breast reconstruction. The second is that the volume of the graft is maintained longer than reconstruction with a musculo-cutaneous flap. The third is that patients can avoid allergic reactions or granulomas as seen with artificial prosthesies. In conclusion, breast conserving surgery with immediate volume replacement with a LTF is a reasonable surgical procedure and has the advantage of avoiding unnecessary surgical procedures for reconstruction and surgical invasion without delaying the diagnosis of local relapse. Moreover, an adequate assessment of risk can spare low risk groups irradiation. PMID- 15657519 TI - Team approach to providing the multidisciplinary medical treatment derived by the patients and their family. AB - To date, the biological approach to breast cancer, such as pathologic subtype genetic analysis has been well investigated, and is considered to be the most important approach to under breast cancer treatment. Recently, the importance of a team approach to multidisciplinary medical treatment and holistic medical treatment has been recognized. The five following points are important: 1) recognition of patients' needs, 2) clarifying responsibility, 3) respect for each other, 4) maintaining good communication, 5) updating the system. Our original 'team approach path' is useful as a communication tool between a patient and the staff. Patient satisfaction is the purpose and a team approach is one method of carrying out medical treatment led by a patient as well as the medical treatment and informed decision based on narrative. PMID- 15657520 TI - Open-configuration MR-guided microwave thermocoagulation therapy for metastatic liver tumors from breast cancer. AB - BACKGROUND: Liver metastases from breast cancer are associated with a poor prognosis, however, local control with microwave thermocoagulation therapy has been used in certain subgroups of these patients in the past decade. In this study, open-configuration magnetic resonance (MR) -guided microwave thermocoagulation therapy was used for metastatic liver tumors from breast cancer, and the efficacy of this treatment was assessed. METHODS: Between June 2000 and April 2004, we used MR-guided microwave thermocoagulation therapy on 11 nodules in 8 patients with metastatic liver tumors from breast cancer. The procedure was carried out under general anesthesia. A 0.5 T open-configuration MR system and a microwave coagulator were used. Near-real-time MR images and real time temperature images were collected and displayed on the monitor. The MR compatible thoracoscope was used and combined with MR imaging guidance. Navigation software, a 3D Slicer, was installed and customized. RESULTS: The customized navigation software displayed near-real-time MR images. The percutaneous puncture into the tumors was successful in all cases. No mortality or major complications occurred as a result of the procedures. Five of the 8 patients are alive with new metastatic foci with a mean observation period of 25.9 months. CONCLUSIONS: We developed several devices to allow safe, easy, and accurate MR-guided microwave thermocoagulation therapy of liver tumors. Open configuration MR-guided microwave thermocoagulation therapy appears to be a feasible method for tumor ablation of metastatic liver tumors from breast cancer. PMID- 15657521 TI - The use of MR imaging guided focused ultrasound in breast cancer patients; a preliminary phase one study and review. AB - The use of high energy focused ultrasound (FUS) waves to destroy tumor tissue in breast cancer is explored. High energy FUS destroys cells by raising the temperature of the treated volume high enough to denature cell proteins and bring about cellular death. The absorbed energy results in extremely high tissue gradients between target cells and surrounding tissue, so the effect of the focused energy is concentrated only at the target, leaving the healthy tissue unscathed. This phase one trial has examined the possibility of ablating breast carcinoma using MRI Guided FUS (MRIgFUS) in place of lumpectomy. Ten female patients underwent the procedure at the Chaim Sheba Medical Center between September 2002 and August 2004, using the ExAblate 2000 (InSightec, Haifa Israel Ltd.). Seven to 10 days after the procedure, all patients underwent standard lumpectomy and axillary sampling to complete standard treatment and to allow pathological evaluation of the procedure. Two patients had a complete pathological response. The remaining 8 patients had varying amounts of residual tumor; 2 had microscopic foci of residual carcinoma, 3 had 10% residual tumor, and 3 had 10-30% of residual tumor. Although still in its infancy, the future role of this type of ablation in breast cancer and other tumors is discussed. PMID- 15657522 TI - General aspects and specific issues of informed consent on breast cancer treatments. AB - Informed consent (IC) is the process by which a patient can make choices about his/her health care; therefore it is considered to be a voluntary authorization given by the patient to the physician. To ensure the patient's right to self determination, what can the physicians do? When treating breast cancer, there are several specific issues that must be clarified by the IC. We have selected and evaluated the basic elements of IC and mentioned the basic concepts of IC in details. First of all, complete information must be disclosed to the patient (physician's responsibility for medical accountability). The information to be disclosed is summarized in the following three elements: 1) The nature of the treatment/procedure, 2) The relevant risks/benefits, and 3) Reasonable alternatives to the proposed intervention (alternative treatments/procedures). However, the physician is not obliged to persuade the patient to accept the proposed intervention. IC information should be documented in detail on the patient's chart without delay. These issues include IC regarding surgical procedures (mastectomy or breast conservation treatment), IC regarding clinical studies (description of randomized controlled trials), IC regarding genetic diagnosis (ethical issues), and the like. IC means informed decision-making, close relationships between physicians and patients are needed. PMID- 15657523 TI - Color Doppler ultrasound as an objective assessment tool for chemotherapeutic response in advanced breast cancer. AB - BACKGROUND: In our part of the world, the majority of the patients with breast cancer present with locally advanced disease and require neo-adjuvant chemotherapy as the primary treatment modality. It is essential to monitor the response to chemotherapy in these patients. Clinical examination as the sole criterion of response assessment is entirely subjective and fallacious. Magnetic Resonance Imaging (MRI) and Computed Tomography (CT) are expensive. The role of Doppler ultrasonography as an imaging modality for this purpose is therefore being evaluated. METHODS: A prospective study was undertaken of 25 cases of locally advanced breast carcinoma (LABC) and Color Doppler sonography was used for the sequential assessment of chemotherapeutic response. The response assessed on the basis of clinical examination and Color Doppler was compared with the histological response. The parameters assessed on color Doppler were a change in the number of flow signals, maximum flow velocity (Vmax), pulsatility index (PI) and resistivity index (RI). Responses were analysed statistically using the Pearson correlation coefficient and Kappa statistics (kappa). The sensitivity, specificity, positive predictive & negative predictive values for predicting complete histological response were calculated. RESULTS: Color Doppler showed a sensitivity of 88.88 % for predicting complete histological response. The negative predictive value of color Doppler was 92.3 %. A significant correlation was obtained between color Doppler and histopathological response. CONCLUSIONS: Color Doppler was found to be an objective and effective tool or modality compared with clinical evaluation in sequential response assessment, especially for predicting complete histological response. PMID- 15657525 TI - A case of amyotrophic lateral sclerosis and breast cancer. AB - We report a case of breast cancer occurring in a patient with amyotrophic lateral sclerosis (ALS). A 58-year-old Japanese woman diagnosed with ALS 6 years previously noticed a mass in the left breast. We performed a modified radical mastectomy for the mass lesion. Invasive ductal carcinoma without lymph node metastasis was diagnosed. During the operation, she had no worsening of her neurological symptoms. The patient has been cancer-free for 11 months since her operation and no improvement has been seen in her neurological condition. To the best of our knowledge, this is the first case of breast cancer occurring in a patient with ALS in Japan. PMID- 15657524 TI - Two cases of breast cancer with cartilaginous and osseous metaplasia. AB - Invasive breast cancer (IBC) with cartilaginous or osseous metaplasia is rare. Here we report two cases of this unusual variation. Case 1: The patient was a 33 year-old woman with a right breast tumor, 2.2 cm in size. Mammograms (MMG) presented no specific findings, but ultrasound (US) showed a cystic-like lesion. Excisional biopsy confirmed IBC with cartilaginous and osseous metaplasia. Biopsy was followed with a modified radical mastectomy. One lymph node was positive, and both estrogen receptor (ER) and progesterone receptor (PgR) were negative. Case 2: The patient was a 43-year-old woman with a left breast tumor, 4.2 cm in size. MMGs presented no findings but US showed an irregular shaped, low-echoic area, suggesting malignancy. Core needle biopsy confirmed IBC with cartilaginous metaplasia. A total adenectomy and lymph node dissection with breast reconstruction using a lattisimus dorsi muscle flap were performed. Two of 18 lymph nodes were positive for metastasis and both ER and PgR were negative. IBC with cartilaginous or osseus metaplasia seem to be divided into two types pathologically, with or without intervening spindle cells, which is related to the prognosis. Matrix producing carcinoma (MPC) has no intervening spindle cells and a better prognosis than other types, however, MPC has been reported to have the same prognosis as ordinary breast cancer after for adjusting its stage. Our two cases were MPC's and no recurrence has been detected 5 and 3 years from the initial therapy, respectively. PMID- 15657526 TI - [Shedding some light on the management of patients with unexplained gastrointestinal bleeding]. PMID- 15657527 TI - Push enteroscopy for gastrointestinal bleeding: diagnostic yield and long-term follow-up. AB - AIM: Little is known about the long-term course of patients explored by push enteroscopy for gastrointestinal bleeding of obscure origin. This study aimed to determine the diagnostic yield and the therapeutic impact of enteroscopy, the rate of rebleeding and predictive factors of rebleeding in these patients. PATIENTS AND METHODS: One hundred nineteen patients underwent push enteroscopy for overt bleeding (N=66) or anemia (N=53). RESULTS: Enteroscopy was positive in 42% of patients (colon 17%, stomach 13%, small bowel 12%) and diagnosed arteriovenous malformations in two-thirds of patients. Twenty-five additional diagnoses were established during the 2-month follow-up. Treatment was definitive in 13% of patients, without recurrent bleeding. Rebleeding occurred in 45% of patients, and was more frequent when a lesion was visualized (73% vs 28% after 5 years, P=0.02). In multivariate analysis, a lesion visualized by enteroscopy was the only independent predictive factor. CONCLUSION: Enteroscopy is not a high performance diagnostic tool for obscure gastrointestinal bleeding and enables definitive treatment in less than 15% of patients. PMID- 15657528 TI - Diagnostic yield and therapeutic implications of capsule endoscopy in obscure gastrointestinal bleeding. AB - AIM: The main aim of this study was to evaluate efficacy and therapeutic impact of capsule endoscopy (CE) in obscure gastrointestinal bleeding (OGIB). In addition, we evaluated the software of automatic detection of red zones (SBI, Given Imaging). PATIENTS AND METHODS: From June 2002 to June 2003, thirty-five patients with OGIB underwent capsule endoscopy after negative upper and lower digestive endoscopy. Capsule endoscopy was performed following a 12-hour fasting period and some received 2 L of PEG the night before for bowel preparation. RESULTS: CE was performed for occult (N=18) or overt (N=17) OGIB. Potentially bleeding lesions were found in 16/35 patients (45.7%). Lesions were angiodysplasias (N=8), ulcerations (N=4), tumors (N=2) and active bleeding without visible lesion (N=2). Lesions were located in gastric antrum (N=1), duodenum (N=2) and jejuno-ileum (N=13). Endoscopic (N=10), surgical (N=2) or medical (N=1) treatments were performed in 13/35 (37%). SBI was retrospectively evaluated in 24 patients with sensitivity, specificity, positive and negative predictive value of respectively 45%, 73%, 50% and 69%. CE retention during 10 days occurred in a patient with a small bowel NSAID-induced stricture. CONCLUSION: CE is a safe and effective procedure in the management of OGIB and had a therapeutic impact in more than one third of patients. PMID- 15657529 TI - Prevalence, characteristics and prognosis of MEN 1-associated glucagonomas, VIPomas, and somatostatinomas: study from the GTE (Groupe des Tumeurs Endocrines) registry. AB - Few studies have concerned the rare functioning endocrine pancreatic tumors associated with multiple endocrine neoplasia type 1 (MEN 1). When sporadic, these tumors have a poor prognosis. AIM: To analyze the frequency, characteristics and prognosis of MEN 1-associated glucagonomas, VIPomas and somatostatinomas recorded in the GTE (Groupe des Tumeurs Endocrines) registry. METHODS: Records of the patients whose GTE registry codes included glucagonoma, VIPoma or somatostatinoma were reviewed. The diagnosis was confirmed when there were clinical signs of a functioning tumor and/or when blood levels of the peptide were higher than twice the upper limit of normal. RESULTS: Among 580 patients with MEN 1, duodeno pancreatic involvement was present in 307 (52.9%). Five (1.6%) had a glucagonoma, 3 (0.98%) a VIPoma and 2 (0.65%) a somatostatinoma. A clinical syndrome was present in 1 patient with glucagonoma, in the 3 with VIPomas and in 1 with somatostatinoma. Tumor size was greater than 3 cm more often for these rare tumours (67%) than in patients with other type of duodeno-pancreatic involvement (28%) (P=0.02) and visceral metastases were more frequent (40% vs 15%; P=0.056). Ten-year survival of patients with glucagonomas, VIPomas or somatostatinomas (53.8%; CI95%: 15.5-92.1) was poorer than that of patients with insulinomas (91.4%; CI95%: 83.399.5; P=0.01) or gastrinomas (81.7%; CI95%: 74.9-88.5; P=0.20) and close to that of patients with non-functioning tumors (62.2%, CI95%: 41.0 83.9; NS). CONCLUSION: Glucagonomas, VIPomas and somatostatinomas, especially the functioning type, are very rare in patients with MEN 1. Prognosis is poor, probably because of large tumor size and high rate of metastasis. Survival is similar to that in patients with non-functioning tumors. PMID- 15657530 TI - [How to improve treatment of resectable pancreatic adenocarcinomas? Surgical resection, histopathological examination, adjuvant therapies]. PMID- 15657531 TI - [Therapeutic alternatives after failure of primary treatment in digestive parasitic diseases in immune competent adult]. PMID- 15657532 TI - [The return of the "great imitator": is there cause for alarm?]. PMID- 15657533 TI - Management of hepatitis C patients: a French population-based study. AB - AIMS: Our aim was to assess the proportion of patients in a well-defined population reaching specialized medical care after hepatitis C diagnosis. METHODS: Hepatitis C-positive patients recorded in the population-based registry of Cote-d'Or, an administrative district in France, constituted the study population. RESULTS: Between 1994 and 1999, new hepatitis C-positive serology was diagnosed in 847 patients, of whom 690 were eligible for this study. A total of 135 patients had not been given specialized medical care after diagnosis; among them, 50.4% had a normal serum alanine transferase level at diagnosis, 62.2% had risk factors related to lifestyle (drug addiction, sexual risk...), and 26.7% were current alcoholics. The 555 other patients were involved in specialized medical care after diagnosis: 42.7% had a liver biopsy and 27.0% were treated. Treatment was carried out more often in males than in females (OR: 1.67; P<0.005), and in patients less than 65 years old (OR: 2.94; P<0.0002). Nearly 30.5% of patients with a Metavir score greater than A1F1 did not undergo treatment. CONCLUSION: This study shows that in a general population at least one patient out of five with hepatitis C infection remains outside the health care system. It also reveals that management practices vary with gender. Further surveys are needed to better understand this phenomenon. PMID- 15657534 TI - [Overlap syndrome between primary biliary cirrhosis and autoimmune hepatitis]. PMID- 15657536 TI - [Liver and parasitic diseases]. PMID- 15657537 TI - [Altered general condition, hypereosinophilia and highly specific liver nodules]. PMID- 15657538 TI - [Amoebic liver abcess]. PMID- 15657539 TI - [Alveolar echinococcosis in the Paris area]. PMID- 15657542 TI - [Endoscopic drainage of cystic dystrophy of the duodenal wall: report of three cases]. AB - Optimal management of cystic dystrophy of a heterotopic duodenal pancreas or cystic dystrophy of the duodenal wall has not yet been established. Surgical treatment by pancreaticoduodenectomy or by-pass procedure is indicated in patients with the most serious symptoms. Endoscopic cystic drainage is an alternative to surgery. We report three cases of cystic dystrophy of the duodenal wall successfully treated by endoscopic drainage. Symptoms disappeared immediately in all cases. No complications were observed. In one case, a second drainage was necessary 15 months after the first one. The 3 patients were free of symptoms after 6, 36, and 44 months of follow-up, respectively. No recurrence was found with CT-scan. The long-term efficiency of the endoscopic procedure must to be evaluated. PMID- 15657543 TI - [Ovarian adenocarcinoma, primary or metastasic tumour of an adenocarcinoma of the colon: the role of molecular biology]. AB - After the primary diagnosis of an adenocarcinoma, diagnosis of the origin of a second adenocarcinoma is a problem (primitive or metastatic). Although the clinical evolution or location sometimes makes it possible to determine the origin of the new lesion, in some cases the diagnosis cannot be confirmed. Although a pathological examination is essential for diagnosis, it may still be inconclusive, making management difficult. The use of molecular biology can help solve this problem. We report the case of a patient who presented with an ovarian adenocarcinoma of an undetermined origin one year after an adenocarcinoma of the colon. This clinical case illustrates the contribution of molecular biology in the diagnosis of the origin of an ovarian adenocarcinoma by characterizing allelic losses in 5 chromosome segments using microsatellite markers genotyping in the two lesions. The comparative analysis suggested the primitive origin of the ovarian lesion. PMID- 15657544 TI - [Primary leiomyoma of the liver: a rare benign tumor]. AB - Primary hepatic leiomyoma is a very rare tumor secondary to benign smooth muscle proliferation. The primary location in the liver is usually found in adult women. A 36-year-old woman with right upper quadrant abdominal pain had primary hepatic leiomyoma. The presenting features of primary leiomyoma and the diagnostic approach for these lesions are discussed, in particular the role of immunohistochemistry. PMID- 15657545 TI - [Autoimmunization induced by interferon alpha therapy in chronic hepatitis C]. AB - We report the case of a 56 year-old woman with post-transfusion chronic hepatitis C who presented with a severe ALT flare up associated with a rapid progression of liver fibrosis during interferon alpha 2b therapy. Several hypotheses were considered to explain the etiology of this ALT flare: there was no viral super infection by other hepatotropic viruses, no toxic hepatitis, no metabolic disease, and no other specific liver diseases could be identified. HLA typing showed a specific profile A1 B8 DR3 (risk factor of auto-immunization during interferon alpha therapy) with antinuclear antibodies and anti smooth muscle antibodies. This case suggests that auto-immunization induced by interferon alpha should be investigated in case of ALT flare that is not followed by an HCV breakthrough. PMID- 15657546 TI - [Acute incarcerated rectal prolapse following bowel preparation with polyethylene glycol]. PMID- 15657547 TI - [Gastric high-grade B-cell lymphoma associated with primary biliary cirrhosis regression after eradication of Helicobacter Pylori]. PMID- 15657548 TI - [Hemorrhagic cystic lymphangioma of the duodenum treated successfully by argon plasma]. PMID- 15657549 TI - [Idiopathic ileocolonic varices: a rare cause of lower gastrointestinal bleeding]. PMID- 15657550 TI - [Gallstone ileus disclosed by a stone in the rectum]. PMID- 15657552 TI - [Anicteric febrile cholestasis in a homosexual patient: beware of a breakout of syphilis]. PMID- 15657551 TI - [Autoimmune hepatitis associated with multiple sclerosis]. PMID- 15657554 TI - Liver aquaporins: significance in canalicular and ductal bile formation. AB - Abstract Bile is primarily secreted in hepatocytes (i.e. the canalicular bile) and subsequently delivered to the intrahepatic bile ducts, where is modified by cholangiocytes (i.e. the ductal bile). Bile formation is the result of the coordinated interactions of membrane-transport systems that generate the vectorial movement of solutes and osmotically driven water molecules. Hepatocytes and cholangiocytes express aquaporins, specialized membrane channel proteins that facilitate the osmotic transport of water. In this review, we provide a summary of what is known on liver AQPs and their significance in canalicular and ductal bile formation under normal and pathological conditions. PMID- 15657556 TI - Clinical and epidemiological features of 147 Chilean patients with chronic hepatitis C. AB - Prevalence, modes of transmission, clinical characteristics and outcomes of hepatitis C (HCV) infection vary in different geographical areas. We aim to describe clinical and epidemiological features of Chilean patients infected with hepatitis C virus. An analysis of demographic, epidemiological, clinical and laboratory data of patients referred to a liver clinic and blood donors with chronic hepatitis C was carried out. 147 patients were evaluated, 68 (46%) were male. Median age was 56 years, median infection age was 27 years and median duration of infection was 27 years. 52.5% of the patients were cirrhotic, and estimated risk of progression to cirrhosis was 16% at 20 years from infection. Risk factors for acquisition of the disease among patients were: Blood transfusion 54%, injection drug use 5%, and risky sexual behavior 2%. No factor was identified in 43% of the patients. Twelve of 64 (18.8%) family members tested positive for HCV antibodies. Genotype 1b was predominant (82%), and 52% of patients had high viral load (>850.000 IU/mL). Liver biopsy was available in 50 patients, showing advanced fibrosis in 54%. These patients were in average 10 years older and tended to have longer duration of infection. Hepatocellular carcinoma was present at the moment of enrollment in 7 patients and developed in 4 more patients during follow up (2.4 years). In conclusion, the natural history and clinical characteristics of HCV infection in Chilean patients is similar to that described elsewhere. The main risk factor was blood transfusion. A significant proportion of patients had advanced liver disease or hepatocellular carcinoma at time of diagnosis. PMID- 15657557 TI - In whom, how and how often is surveillance for hepatocellular carcinoma cost effective? AB - BACKGROUND: Despite well known worldwide differences in hepatocellular carcinoma incidence, which reflect different risk profiles, current recommendation of surveillance with ultrasound and alpha-fetoprotein twice-a-year has been restricted to cirrhotic patients. To evaluate the generalizability of this recommendation, we reviewed the clinical charts of hepatocellular carcinoma cases in a Mexican scenario. To evaluate efficiency, we performed a literature based cost-effectiveness analysis. METHODS: Charts pertaining to 174 consecutive patients with histologically proven hepatocellular carcinoma, seen at a tertiary health care centre were analysed. A decision tree, based on the surveillance and recall algorithm of the European Association for the Study of the Liver was constructed. Ultrasound and/or alpha-fetoprotein, performed every six or twelve months were the diagnostic alternatives, and accurate diagnoses, direct medical costs and cost-effectiveness ratios were the outcomes of interest. RESULTS: Male:female ratio was 1.2:1, underlying liver disease was secondary to alcohol in 44% and to hepatitis C virus in 26%, documented cirrhosis was present in 42%. Cost-effectiveness ratios for twice-a-year ultrasound and alpha-fetoprotein ranged from $303.09 to $346.22 U.S. dollars per accurate diagnosis, and for annual ultrasound from $115.86 to $116.42 U.S. dollars. CONCLUSIONS: Male gender, hepatitis C and cirrhosis were not predominant characteristics in our series. If a hepatocellular carcinoma surveillance program were to be instituted in our setting, or where patient characteristics are similar to ours, it probably should not be restricted to cirrhotic patients. Recommended performance of ultrasound and alpha-fetoprotein every six months is the least cost-effective surveillance strategy. Instead, annual ultrasound optimises diagnoses and costs. PMID- 15657555 TI - Microarrays, antiobesity and the liver. AB - In this review, the microarray technology and especially oligonucleotide arrays are exemplified with a practical example taken from the perilipin-/- mice and using the dChip software, available for non-lucrative purposes. It was found that the liver of perilipin-/- mice was healthy and normal, even under high-fat diet when compared with the results published for the scd1-/- mice, which under high fat diets had a darker liver, suggestive of hepatic steatosis. Scd1 is required for the biosynthesis of monounsaturated fatty acids and plays a key role in the hepatic synthesis of triglycerides and of very-low-density lipoproteins. Both models of obesity resistance share many similar phenotypic antiobesity features, however, the perilipin-/- mice had a significant downregulation of stearoyl CoA desaturases scd1 and scd2 in its white adipose tissue, but a normal level of both genes inside the liver, even under high-fat diet. Here, different microarray methodologies are discussed, and also some of the most recent discoveries and perspectives regarding the use of microarrays, with an emphasis on obesity gene expression, and a personal remark on my findings of increased expression for hemoglobin transcripts and other hemo related genes (hemo-like), and for leukocyte like (leuko-like) genes inside the white adipose tissue of the perilipin-/- mice. In conclusion, microarrays have much to offer in comparative studies such as those in antiobesity, and also they are methodologies adequate for new astounding molecular discoveries [free full text of this article Online]. PMID- 15657559 TI - Images in hepatology. Telangiectatic adenoma: a variant of hepatocellular adenoma. PMID- 15657558 TI - Bile duct reconstruction after iatrogenic injury in the elderly. AB - Bile duct injuries occur with a frequency of 1 to 5 per 1000 cases as a result of an increase in the number of procedures performed. Elderly patients have more severe lithiasis- related diseases than the younger population. This fact increases the likelihood of conversion from laparoscopic to open surgery, and the probability of injury. We report the results of bile duct reconstruction after injury in these patients. SETTING: A tertiary care academic university hospital. METHODS: The files of patients over 65 years of age who had biliary tract reconstruction after iatrogenic injury were retrospectively reviewed. Post operative results, quality of life and failures from the repair were analyzed. RESULTS: 20 patients over the age of 65 were referred for biliary tract reconstructive surgery over a ten year period. Mean age was 71 (65 - 83). Nineteen cases were referred from other hospitals. Fourteen cases (60%) had comorbidities (type 2 diabetes mellitus, systemic hypertension). All patients were treated by means of Roux en Y hepatojejunostomy. No operative morbility was recorded and only one major complication (abdominal collection) was found. Two long term mortalities unrelated to surgery were found. Treatment success (as defined by the Johns Hopkins criteria) was obtained in 17 cases (85%) and only one patient (5%) required reoperation two years after the initial attempt. CONCLUSIONS: Elderly patients that survive biliary injury and are reconstructed have long-term results comparable to the younger population; Roux en Y hepatojejunostomy offers the best surgical choice. PMID- 15657560 TI - Acute liver failure and the Molecular Adsorbents Recirculating System: early experience in a tertiary care hospital in Mexico City. AB - Acute liver failure is a clinical condition associated with high mortality despite recent technological advances. Supportive devices such as the Molecular Adsorbents Recirculating System (MARS) provide therapeutic strategies to add time to find an organ for orthotopic liver transplantation or to allow the native liver to recover sufficiently to make transplantation unnecessary. In this series of cases, we discuss our initial experiences with three patients with acute liver failure. One patient had high bilirubin levels caused by Epstein-Barr virus infection and responded well after three MARS sessions. In a second patient, MARS therapy was used to treat acute-on-chronic liver failure caused by chronic hepatitis B virus infection that had not been treated previously; because of severe hemodynamic compromise, only one MARS session was performed. The third patient had an initial diagnosis of acute liver failure and cryptogenic hepatitis, and was treated with five MARS sessions as a supportive measure until the definitive diagnosis (metastatic disease) was performed. In all patients, MARS therapy was well tolerated and induced only mild hypokalemia. In conclusion, although MARS therapy was an effective strategy for these cases of liver failure and greatly improved the biochemical variables, its impact on the mortality rate has not yet been determined. PMID- 15657561 TI - Pericarditis and possible antiphospholipid syndrome on primary biliary cirrhosis. PMID- 15657562 TI - Tribute to Paul LoGerfo, MD. PMID- 15657563 TI - Intraoperative electromyographic monitoring of the recurrent laryngeal nerve in reoperative thyroid and parathyroid surgery. AB - BACKGROUND: Injury to the recurrent laryngeal nerve (RLN) is a rare complication of initial thyroid and parathyroid surgery, but the prevalence is much higher in the reoperative setting. The use of continuous, intraoperative electromyographic monitoring of the RLN has been suggested to improve the safety of cervical explorations. METHODS: Outcomes of a group of reoperative thyroid and parathyroid cases that used EMG monitoring with endoscopically applied hook-wire electrodes were compared with a group of cervical reoperations without monitoring. Office laryngoscopy (indirect or fiberoptic) was used to evaluate and follow suspected RLN complications. RESULTS: Electromyography was used in 52 cervical reexploration procedures. Patients averaged 1.8 previous explorations (range, 1-7 explorations) and underwent procedures for parathyroid (31%) and/or thyroid (77%) disease (overall, 72% malignant). The non-monitored group had 59 patients with similar characteristics. Only 1 permanent nerve complication in each group was unintended (electromyography, 1.9%; non-electromyography, 1.7%). Seven false negative and 2 false-positive electromyographic findings occurred. No complications resulted from placement of the electromyography electrodes. CONCLUSIONS: Intraoperative electromyographic monitoring of the RLN in reoperative neck surgery can be performed safely but did not decrease RLN complications in this study. Experience and routine nerve exposure remain crucial to the minimization of RLN complications. PMID- 15657564 TI - Hereditary medullary thyroid carcinoma in patients greater than 50 years old. AB - BACKGROUND: Despite near complete penetrance and frequent early evaluation of medullary thyroid carcinoma (MTC) in multiple endocrine neoplasia 2 (MEN 2) variants, a significant minority of patients are evaluated later in life. METHODS: With the aim of characterizing the expression of hereditary MTC in an older cohort, MEN 2 patients from our institutional database who were evaluated after age 50 years were identified, and clinical data were reviewed. RESULTS: Thirty-nine patients (36 MEN 2A, 3 FMTC, and no MEN 2B) who were evaluated after age 50 years were identified; they represented 9% of all MEN 2 patients who were enrolled in our program. Most of the patients (79%) had abnormal screening examinations, and the AJCC staging was significantly higher in this cohort compared with younger patients. Overall, 43% of the patients had normal calcitonin levels after operation. There were 3 observed MTC-related deaths, all from distant metastases; the overall survival rate was 86% at 5 years and 74% at 10 years. The distribution of RET mutations in this cohort was similar to younger patients. CONCLUSIONS: These results suggest the presence of modifiers of MTC expression. Despite the tendency of older patients with hereditary MTC to have advanced stage disease at evaluation, they have high rates of biochemical cure, and the overall survival is excellent. PMID- 15657565 TI - Squamous cell carcinoma-related oncogene is highly expressed in developing, normal, and adenomatous adrenal tissue but not in aggressive adrenocortical carcinomas. AB - BACKGROUND: Our previous work has demonstrated squamous cell carcinoma-related oncogene (SCCRO) expression in adult murine adrenocortical tissue. The aim of this study was to assess patterns of SCCRO expression in the embryonic murine adrenal gland, and in normal and neoplastic human adrenocortical tissues in order to determine its role as a marker of differentiation in adrenocortical development and neoplastic progression. METHODS: Murine embryos were procured at developmental stages E8 to E18. A tissue microarray was constructed containing 38 normal, 39 adenomatous, and 87 carcinomatous human adrenocortical specimens. Immunohistochemistry for SCCRO was performed and its expression was graded in suitable tissues. RESULTS: SCCRO expression was detected in the murine adrenal cortex as early as E15 and persisted into the postnatal period. High-level SCCRO expression was identified in 94% of normal (32/34) and adenomatous (29/31) adrenocortical specimens but in only 63% (45/72) of adrenocortical carcinoma (ACC) specimens ( P = .001). Loss of SCCRO expression in primary ACC (13/34 (34%)) correlated with advanced stage ( P = .06), presence of M1 disease at presentation ( P = .03), and worse overall survival ( P = .006). CONCLUSIONS: SCCRO appears to be a marker of adrenocortical differentiation in both murine and human systems. SCCRO expression may be useful in distinguishing adrenocortical adenomas from ACC. Moreover, loss of SCCRO expression in primary ACC is associated with worse outcome and may be a marker of progressive dedifferentiation in these tumors. PMID- 15657566 TI - Should pheochromocytoma size influence surgical approach? A comparison of 90 malignant and 60 benign pheochromocytomas. AB - BACKGROUND: Diagnostic tests cannot reliably distinguish malignant from benign pheochromocytomas. Laparoscopic adrenalectomy for pheochromocytomas >6 cm is controversial because of a perceived increased risk of malignancy that is based on anecdotal reports and small series. The aim of this study was to determine if pheochromocytoma size should affect the choice of surgical approach. METHODS: Malignant pheochromocytomas in the Surveillance Epidemiology and End Results (SEER) database (1988-2000) were compared to benign pheochromocytomas in our institutional database (1993-2003). The sensitivity, specificity and likelihood ratio for tumor size to predict malignancy were calculated for both groups. RESULTS: Ninety malignant and 60 benign pheochromocytomas were compared. Overall, malignant pheochromocytomas were larger than benign pheochromocytomas (7.6 +/- 4.2 cm vs 5.3 +/- 2.3 cm). However, tumor size was not significantly different between malignant pheochromocytomas without local invasion or metastases and benign pheochromocytomas (6.1 +/- 3.1 cm vs 5.3 +/- 2.3 cm). In pheochromocytomas with local disease only, maximum likelihood ratio to predict malignancy was at a tumor size of greater than 8 cm (2.84). CONCLUSIONS: Although risk of malignancy increases with size for all pheochromocytomas, size does not reliably predict malignancy in pheochromocytomas with local disease only. Regardless of tumor size, laparoscopic adrenalectomy for pheochromocytoma should be converted to open adrenalectomy for difficult dissection, invasion, adhesions, or surgeon inexperience. PMID- 15657568 TI - Does propofol anesthesia affect intraoperative parathyroid hormone levels? A randomized, prospective trial. AB - BACKGROUND: Intravenous propofol (2,6-diisopropylphenol) infusion is used commonly for sedation/anesthesia during operations. Several authors have reported that propofol can interfere with intact parathyroid hormone (PTH) testing in vitro. Therefore, many surgeons avoid propofol during parathyroidectomy. METHODS: To determine whether propofol affects intraoperative PTH levels in vivo, we randomly assigned 34 patients (80% power; alpha < .05) with secondary hyperparathyroidism to undergo surgery for dialysis access. Patients were assigned randomly to local anesthesia with either propofol (n = 17 patients) or midazolam (n = 17 patients) sedation. PTH values were obtained before the procedure and at 10 minutes and 30 minutes after the start of the propofol or midazolam. RESULTS: Median preoperative serum PTH and calcium levels were 175 pg/mL (range, 27-2646 pg/mL) and 9.2 mg/dL (range, 8.1-10.8 mg/dL), respectively. There was no statistically significant difference between the PTH levels in the 2 groups at each of our time points. There was also no difference in the percentage of change from baseline in the PTH values between our 2 groups. No patient in either group had a sustained drop in their PTH level of greater than 50%. CONCLUSIONS: Intravenous propofol infusion does not alter PTH levels significantly during the operation. Therefore, we believe the intraoperative PTH assay can be used safely during propofol sedation when parathyroid surgical procedures are being performed. PMID- 15657569 TI - The coming of age of radiation-induced hyperparathyroidism: evolving patterns of thyroid and parathyroid disease after head and neck irradiation. AB - BACKGROUND: This study investigates the association of thyroid and parathyroid disease in radiation-exposed patients and tests the hypothesis that hyperparathyroidism (HPT) occurs after a longer latency period. METHODS: Routine questioning identified 40 patients in the endocrine surgery clinic with a history of radiation exposure. Patients with thyroid disease were screened for HPT and vice versa. RESULTS: The known diagnosis at initial referral was thyroid disease alone in 26 of 40 patients, HPT alone in 9 of 40 patients, and both in 5 of 40 patients. After screening was completed, a total of 18 of 40 patients were found to have both thyroid disease and HPT. HPT manifested an average of 17 years later than thyroid disease (52.6 +/- 10.0 years vs 35.5 +/- 13.8 years after radiation therapy [ P < .05]). One third of the patients with HPT had normal serum calcium levels despite elevated parathyroid hormone levels and abnormal parathyroid glands. CONCLUSION: As the widespread use of x-ray therapy for benign childhood conditions becomes more remote, practitioners should expect a decreasing number of patients with radiation-associated thyroid disease, with an increase in HPT. Patients with radiation exposure and thyroid disease should be evaluated carefully and followed for HPT. Those patients who undergo surgical procedures should have close inspection of the parathyroid glands, with biopsy and excision when appropriate. PMID- 15657570 TI - Intraoperative PTH monitoring in parathyroid hyperplasia requires stricter criteria for success. AB - BACKGROUND: Intraoperative parathyroid hormone (IOPTH) monitoring in parathyroidectomy for multigland disease is less clear than for single-gland disease. This study assesses the role of IOPTH for hyperplasia. METHODS: A prospective database revealed 45 patients with hyperplasia undergoing parathyroidectomy utilizing IOPTH from February 1999 to August 2003. RESULTS: Twenty-six females and 19 males had a mean age of 55 years. Twenty-two patients underwent total parathyroidectomy. Twenty-three patients underwent subtotal parathyroidectomy. Twenty-seven patients (60%) had a drop of IOPTH greater than 50% at 10 minutes after removal of all presumably abnormal parathyroid tissue. Nine additional patients (20%) had an IOPTH drop greater than 50%, but continued exploration revealed more abnormal tissue. Nine patients failed to decrease greater than 50%, and exploration was continued. A final IOPTH less than 35 pg/mL or a greater than 90% decrease from baseline was predictive of a successful operation in 40 patients. The 5 patients who did not meet this criteria remained hyperparathyroid. CONCLUSIONS: IOPTH identifies sporadic hyperplasia and guides completeness of resection for patients with known hyperplasia. However, more rigid criteria are required than for adenomas. Failure to achieve appropriate decreases in IOPTH should prompt further neck exploration or a search for a mediastinal gland. PMID- 15657571 TI - Molecular analysis of Hurthle cell neoplasms by gene profiling. AB - BACKGROUND: Though Hurthle cell tumors are considered a variant of follicular lesions, recent data have suggested that Hurthle cell carcinomas may be more closely related to papillary thyroid carcinomas (PTCs). These studies were conducted to determine if molecular profiling can enhance our understanding of Hurthle cell neoplasms. METHODS: Thirteen Hurthle cell tumors (9 adenomas, 4 carcinomas) were analyzed with the Affymetrix HU-95 gene chips. Molecular profiles obtained were compared to 14 follicular adenomas (FAs), 7 follicular carcinomas (FCs), 10 PTCs, 11 follicular variant PTCs, and 9 hyperplastic nodules. Hierarchical cluster analysis defined potential groupings and differences among samples. RESULTS: Hurthle cell carcinomas grouped with FCs 100% of the time. Surprisingly, Hurthle cell adenomas clustered with FCs when compared to FAs and FCs in 8/9 (88%) cases. All 13 Hurthle cell lesions migrated as a distinct group separate from PTCs and FVPTCs. Finally, all Hurthle cell lesions clustered with FCs, rather than PTCs, when compared to both groups. CONCLUSIONS: Molecular profiles of Hurthle cell adenomas and carcinomas are more similar to FCs than benign lesions or PTCs. Although Hurthle cell adenomas generally behave in a benign fashion, the molecular signature of these lesions suggests a more malignant phenotype. PMID- 15657572 TI - The influence of sample site on intraoperative PTH monitoring during parathyroidectomy. AB - BACKGROUND: Intraoperative parathyroid hormone (IOPTH) monitoring is common during parathyroidectomy. We hypothesized that sample site (peripheral vs central vein) may impact IOPTH interpretation. METHODS: Two hundred and one patients underwent curative parathyroidectomy for single-gland disease. IOPTH was drawn peripherally (PV) in 114 patients and centrally (CV, jugular vein) in 87 patients. Decrease from baseline IOPTH and the presence of a normal value at 10 and 15 minutes were determined. The slope of IOPTH decay was calculated. These data were compared between sample sites. RESULTS: Median baseline IOPTH was 268 pg/mL (CV) and 191 pg/mL (PV, P = .003). The mean IOPTH decay slopes were -0.75 (PV) and -0.76 (CV, P = NS), and the mean percent IOPTH decrease at 10 minutes was 79% PV and 80% CV (P = NS). At 10 minutes, a > or =50% drop from baseline was seen in 94% (CV) versus 97% (PV) of patients ( P = NS), resulting in a median IOPTH of 40 pg/mL (CV) versus 34 pg/mL (PV, P = .09). By 15 minutes, the central IOPTH had decreased by > or =50% of baseline in 98% of patients ( P = NS when compared to the 10-minute PV site). CONCLUSIONS: IOPTH kinetics are largely the same for PV and CV sample sites, but baseline values are higher with central sampling. Consequently, CV IOPTH levels are generally higher at 10 minutes, but this discrepancy resolves by 15 minutes. The surgeon utilizing CV samples may need to extend the sampling period. PMID- 15657573 TI - Isolated hepatic perfusion for the treatment of patients with advanced liver metastases from pancreatic and gastrointestinal neuroendocrine neoplasms. AB - BACKGROUND: We report results of using isolated hepatic perfusion (IHP) in patients with advanced progressive liver metastases (LM) from pancreatic and gastrointestinal neuroendocrine neoplasms (NENs). METHODS: Thirteen patients with LM from NENs (mean percent hepatic replacement, 30; range, 10-60) were treated with a 1-hour hyperthermic IHP via a laparotomy with the use of 1.5 or 2.0 mg/kg melphalan and/or 1 mg tumor necrosis factor. An oxygenated extracorporeal circuit with inflow through the gastroduodenal artery and common hepatic artery, and outflow to a segment of the inferior vena cava was used. Portal flow and inferior vena cava flow were shunted to the axillary vein. Radiographic response, recurrence pattern, and survival were assessed. RESULTS: Mean operative time was 9 hours (8-11 hours), and a median hospital stay was 10 days (6-64 days). Fifty percent of evaluable patients had a radiographic partial response in the liver (mean duration, 15 months; range, 6-26 months; 2 ongoing). Four had a marginal response (25%-49% reduction in the neoplasm). The median, hepatic, progression free survival was 7 months (range, 3-27 months). The median actuarial survival was 48 months including 1 treatment mortality (median follow-up, 23 months). CONCLUSIONS: For patients with advanced LM from NENs, IHP provides a reasonable response rate and duration with acceptable morbidity; continued clinical evaluation is important. PMID- 15657574 TI - Preventable reoperations for persistent and recurrent papillary thyroid carcinoma. AB - BACKGROUND: Cervical recurrence occurs in up to 25% of patients with papillary thyroid carcinoma (PTC) due either to the aggressive biology of PTC or to inadequate treatment. This retrospective study was designed to determine the frequency of inadequate surgical or medical therapy as a cause of persistent or recurrent PTC. METHODS: We identified all patients who underwent reoperation for persistent (within 6 months of initial or pre-referral operation) or recurrent (greater than 6 months after initial or pre-referral operation) PTC from 1992 to 2003. Medical records including initial preoperative imaging, operative, and histopathology reports were reviewed. The initial surgical procedure was considered incomplete if all gross neoplasm was not removed or if "node plucking" was performed, and a subsequent recurrence occurred in the same cervical compartment. RESULTS: Seventy-two consecutive patients underwent reoperation for persistent (17) or recurrent (55) PTC. Of the 17 patients with persistent PTC, reoperation was judged to have been possibly preventable in 14 (82%) due to inadequate preoperative imaging or incomplete initial surgery. Of the 55 patients with recurrent PTC, reoperation was judged to have been possibly preventable due to incomplete initial surgery in 14 (25%). Based on the National Comprehensive Cancer Network guidelines, 33 (46%) of 72 patients with persistent or recurrent PTC received inadequate initial local treatment. CONCLUSIONS: Reoperation in 28 (39%) of 72 patients with persistent or recurrent PTC was potentially preventable. Accurate preoperative staging and adherence to the suggested National Comprehensive Cancer Network treatment guidelines may minimize the need for cervical reoperation. PMID- 15657575 TI - Profile and outcome of pure androgen-secreting adrenal tumors in women: experience of 21 cases. AB - BACKGROUND: The aim of the study was to determine the outcome and possible prognostic factors of pure androgen-secreting adrenal tumors (PASATs). METHODS: In a review of 801 adrenal operations from 1970 through 2003, 21 women with PASATs were divided into 2 groups, benign and undetermined tumors (Weiss score < or = 3) (BT = 11) and malignant tumors (Weiss > or =4 or nonresectable) (MT = 10). RESULTS: In both groups, age at presentation was similar. There were no differences concerning type of secretion, but increase in testosterone level was 2.6-fold greater in malignant tumors (MT) than benign tumors (BT). Imaging studies allowed diagnosis of malignancy in 4 of 10 MT. Size and weight were greater for MT than for BT (average, 13.7 vs 9.2 cm and 1462 vs 206 g). At follow up (median, 17 y; range, 1 to 33 y) 2 of 11 patients with BT died of unrelated causes, and 9 of 11 are alive without recurrence; 5 of 10 patients with MT died of disease, and 3 are alive with disease; 7 of those 8 patients had stage III or IV disease and/or had a Weiss score of 6 or greater. Two patients with MT are alive without disease; both were Weiss 7, stage II and received mitotane postoperatively. CONCLUSIONS: PASATs of undetermined benign condition share the good prognosis of BT. Weiss score is diagnostic of malignancy. MacFarlane classification determines the prognosis, and long-term disease-free survivors at stages III/IV are never observed after operation only. Postoperative mitotane therapy might be beneficial at stage II. PMID- 15657576 TI - Effect of calcium channel blockers on the sensitivity of preoperative 99mTc-MIBI SPECT for hyperparathyroidism. AB - BACKGROUND: Technetium 99m ( 99m Tc)-methoxyisobutylisonitrile (MIBI) single photon emission computed tomography (SPECT) is frequently used in the evaluation of patients with hyperparathyroidism. Calcium channel blockers (CACBs) may affect 99m Tc-MIBI uptake by parathyroid cells. This study examines the effect of CACB therapy on the sensitivity of 99m Tc-MIBI SPECT localization for hyperparathyroidism. METHODS: Two hundred fifty-three operated patients with hyperparathyroidism were retrospectively reviewed. The potential effect of CACB therapy on 99m Tc-MIBI scan sensitivity was examined by using logistic regression analysis. Possible confounding factors were considered. RESULTS: Among 235 patients, those with multiple endocrine neoplasia, type I (MEN-I), MEN-IIA, 4 gland hyperplasia, secondary hyperparathyroidism, and tertiary hyperparathyroidism exhibited no difference associated with CACB use. Of the remaining 198 patients with primary hyperparathyroidism, 7/30 (23%) with negative 99m Tc-MIBI SPECT scans compared to 24/168 (14%) with positive scans used CACBs. After correcting for age, gender and gland weight, the odds ratio (OR) for a negative study in patients taking CACBs was 2.88 (95% CI, 1.03-8.10; P = .045). Atherosclerosis, hypertension, diabetes mellitus, preoperative calcium and parathyroid hormone levels, and thyroid hormone use were not confounding factors. CONCLUSIONS: CACB therapy reduces the sensitivity of 99m Tc-MIBI parathyroid SPECT in patients with primary hyperparathyroidism. Further studies are required to determine the potential reversibility of this effect with termination of CACB therapy. PMID- 15657577 TI - The surgical management of MEN-1 pancreatoduodenal neuroendocrine disease. AB - BACKGROUND: The management of multiple endocrine neoplasia, type 1 (MEN-1) pancreatoduodenal neuroendocrine neoplasms (NENs) is controversial. An aggressive surgical approach is intended to control the functional syndromes and malignant potential for nodal or distant metastasis. METHODS: The results of treating 39 patients with MEN-1 pancreatoduodenal NENs over a 35-year period are available from chart reviews and patient interviews. This study focuses on pattern of disease, disease recurrence, and long-term functional outcomes. RESULTS: Between 1967 and 2003, 39 patients ages 19 to 58 years (mean age, 37) had abdominal operations for their pancreatoduodenal NENs: 26 with Zollinger-Ellison syndrome, 4 with hypoglycemia, 3 with both Zollinger-Ellison syndrome and hypoglycemia, and 6 with nonfunctional neoplasms. Fifteen of these 39 patients had malignant disease on initial abdominal operation; 24 of 39 patients have not required abdominal reoperation, 17 of whom have available follow-up data. Of these 17 patients, 11 have biochemical evidence of disease recurrence (increased serum concentrations of gastrin, insulin, or pancreatic polypeptide), while 6 have no biochemical evidence of recurrence. A total of 30 abdominal reoperations were performed in 15 patients; 14 of 15 patients undergoing 1 or more reoperations developed evident malignant disease by their most recent operation. Nine of 13 reoperative patients with follow-up data have evidence of disease recurrence. Functional outcomes available in 20 patients showed that 10 patients require insulin and that 6 require oral hypoglycemic medications. Ninety percent have no abdominal pain or nausea/vomiting, while 4 are unable to return to work secondary to this disease. CONCLUSIONS: Treatment of MEN-1 pancreatoduodenal NENs is met with frequent recurrence and some treatment-related morbidity and mortality. Most patients (22 of 39) eventually demonstrated malignant growth, but, with this strategy, few died of this disease. PMID- 15657578 TI - RET signals through focal adhesion kinase in medullary thyroid cancer cells. AB - BACKGROUND: The RET proto-oncogene is implicated in medullary thyroid cancer (MTC) and has been shown to signal indirectly to focal adhesion kinase (FAK) in cell types other than MTC. We have previously shown that FAK is phosphorylated in MTC cells. We hypothesized that inhibition of RET with pharmacologic inhibitors or by depletion with siRNA would decrease FAK phosphorylation in MTC cells, thereby implicating a RET-FAK signaling pathway. METHODS: Human MTC cells (TT cells) were treated with pharmacologic inhibitors or transfected with RET siRNA. Total protein was detected by immunoblotting. Phosphorylated FAK was detected by immunoprecipitating total FAK and immunoblotting with antiphosphotyrosine. RESULTS: Treatment of MTC cells with the inhibitor PP2 significantly inhibited RET phosphorylation and, to a lesser extent, FAK phosphorylation. Imatinib mesylate inhibited FAK phosphorylation only at high doses. RET siRNA significantly decreased RET expression and FAK phosphorylation. CONCLUSIONS: RET signals through FAK in MTC cells. Whether this is due to a direct or indirect interaction is not yet clear. PP2 or a similar inhibitor might be a useful treatment for MTC. PMID- 15657579 TI - The palliative role of 131I-MIBG and 111In-octreotide therapy in patients with metastatic progressive neuroendocrine neoplasms. AB - BACKGROUND: Radiolabeled octreotide and metaiodobenzylguanidine (MIBG) have demonstrated limited antitumor effect on neuroendocrine neoplasms (NENs). The purpose of this study was to assess the palliative benefit of radionuclide therapy (RNT) in NENs. METHODS: Since April 2001, RNT for progressive, nonsurgically resectable NENs was utilized. NENs that were MIBG positive received 131 I-MIBG therapy, whereas octreotide-only-positive neoplasms received 111 In octreotide therapy. Symptomatic, quality of life (QoL), biochemical, and radiographic responses to RNT were evaluated. RESULTS: Twenty-four patients had either MIBG or octreotide avid NENs. A mean (range) of 4 (1-7) 131 I-MIBG treatments were given to 13 patients over a duration of 18 months (6-42). The group consisted of 2 medullary thyroid cancer (MTC), and 2 foregut and 9 small bowel carcinoids; 12 patients (92%) had symptomatic improvement. Stability of tumor size was confirmed in 6, regression in 2, and progression in 5. Biochemical responses were demonstrated in 2. Five (2-16) treatments of 111 In-octreotide was given to 11 patients over 17 months (6-40). There were 1 MTC, 1 insulinoma, 2 gastrinomas, and 3 small-bowel and 4 foregut carcinoids. Symptomatic benefit was seen in 6 (55%), biochemical response in 3. Tumor regression was seen in 1, stability in 5, and progression in 5. CONCLUSIONS: Radionuclide therapy appears to offer good palliation to patients with progressive NENs. PMID- 15657580 TI - Role for adrenal venous sampling in primary aldosteronism. AB - BACKGROUND: The aim of this study was to determine the effect of adrenal venous sampling (AVS) on the management of patients with primary aldosteronism. METHODS: From September 1990 through October 2003, 203 patients with primary aldosteronism (mean age, 53 years; range, 17-80; 163 men) were selected prospectively for AVS on the basis of degree of aldosterone excess, age, desire for surgical treatment, and computed tomographic (CT) findings. RESULTS: Both adrenal veins were catheterized in 194 patients (95.6%). Notable among the 110 patients (56.7%) with unilateral aldosterone hypersecretion were 24 (41.4%) of 58 patients with normal adrenal CT findings, 24 (51.1%) of 47 with unilateral micronodule (< or =10 mm) apparent on CT (7 had unilateral aldosterone hypersecretion from the contralateral adrenal), 21 (65.6%) of 32 with unilateral macronodule (>10 mm) apparent on CT (1 had unilateral aldosterone hypersecretion from the contralateral adrenal), 16 (48.5%) of 33 with bilateral micronodules, and 2 (33%) of 6 with bilateral macronodules. CONCLUSIONS: On the basis of CT findings alone, 42 patients (21.7%) would have been incorrectly excluded as candidates for adrenalectomy, and 48 (24.7%) might have had unnecessary or inappropriate adrenalectomy. AVS is an essential diagnostic step in most patients to distinguish between unilateral and bilateral adrenal aldosterone hypersecretion. PMID- 15657581 TI - Early prediction of postthyroidectomy hypocalcemia by one single iPTH measurement. AB - BACKGROUND: We prospectively evaluated the possibility to make an early prediction of postthyroidectomy hypocalcemia by postoperative intact parathyroid hormone (iPTH) measurements. METHODS: Fifty-three consecutive patients who underwent bilateral thyroid resection were included; iPTH was measured preoperatively, at the end of the surgical procedure, and at 2, 4, 6, 24, and 48 hours after the operation. Patients who had hypocalcemia (serum total calcium, <8.0 mg/dL) were compared with normocalcemic patients. RESULTS: Sixteen patients experienced hypocalcemia. Six patients experienced symptoms. No significant difference was found between hypocalcemic and normocalcemic patients concerning demographic, pathologic, and preoperative laboratory data, surgical procedure, and intraoperative findings. Postoperative iPTH levels were reduced in hypocalcemic patients at the end of the procedure and at 2, 4, 6, 24, and 48 hours after the operation ( P < .001). IPTH levels below the normal range (<10 pg/mL) at 4 and 6 hours after the operation correctly predicted postoperative hypocalcemia and symptoms in all but 1 patient with a self-limiting, asymptomatic hypocalcemia (serum calcium concentration, 7.8 mg/dL) (specificity, 100%; sensitivity, 94%; overall accuracy, 98%). CONCLUSIONS: One single iPTH measurement reliably can predict, early after thyroidectomy, which patients are prone to clinically relevant postoperative hypocalcemia and necessitate supplementation treatment and which patients are eligible for a safe early discharge. PMID- 15657582 TI - Normocalcemic hyperparathyroidism in patients with osteoporosis. AB - BACKGROUND: Diagnosing primary hyperparathyroidism (PHP) in patients with osteoporosis is important because of the benefits of surgery. Screening patients with osteoporosis for PHP with only total serum calcium level will fail to diagnose PHP in patients with intermittent or no elevation of the total calcium level. METHODS: This is a retrospective study of 140 patients who had a preoperative bone density study of the 223 patients who had surgery for PHP from January 1995 to June 1999. Normocalcemic hyperparathyroidism was defined as having all normal total calcium values or only intermittent elevation defined as at least 40% of the total calcium values in the normal range. RESULTS: Osteoporosis was identified in 64 of these 140 patients (46%). Fifteen patients with osteoporosis and PHP had normocalcemic hyperparathyroidism. Six of these patients had all preoperative total calcium values in the normal range, and the remaining 9 patients had intermittent elevation of the total calcium. This group consisted of 12 women and 3 men aged 55 to 79 years. Forty-four concomitant ionized and total calcium values were available in the patients with all normal preoperative total serum calcium values. Forty-two serum ionized calcium values (95%) were elevated. Fifty-one concomitant values were available in the patients with intermittent elevation of the total calcium, and only 20 total calcium values (39%) were elevated and 47 of ionized values (92%) were elevated ( P < .01). Intact parathyroid hormone was also significantly better than total calcium in identifying PHP. Twenty of 23 intact serum parathyroid hormone values (87%) were elevated ( P < .05). CONCLUSIONS: Screening patients with osteoporosis for PHP with only total calcium levels will fail to identify patients with no elevation of total calcium level and many patients with only intermittent elevation of the total calcium level. Ionized calcium and intact parathyroid hormone were comparable and significantly better than total calcium level in the detection of PHP in patients with osteoporosis. Ionized calcium and intact parathyroid hormone should be used to diagnose hyperparathyroidism in patients with osteoporosis and normal serum total calcium levels. PMID- 15657583 TI - Predictive factors for recurrence after thyroid lobectomy for unilateral non toxic goiter in an endemic area: results of a multivariate analysis. AB - BACKGROUND: The aim of the study was to identify the factors that are predictive of recurrence after thyroid lobectomy for unilateral non-toxic thyroid goiter in an endemic region through a multivariate analysis. METHODS: Two hundred sixty eight consecutive patients who underwent thyroid lobectomy and who were evaluated by the same endocrinologist were included. Univariate and multivariate analysis analyzed the relationship between sex, age, preoperative thyroid-stimulating hormone, duration of disease, duration of levothyroxine (LT4) preoperative therapy, cytologic results, histologic results, resected thyroid weight, numbers and diameters of thyroid nodules, morphologic alterations of the remnant lobe, follow-up length, postoperative LT4 therapy, ultrasonographic evidence of recurrence, and reoperation. RESULTS: The incidence of recurrence was 33.9% (91/268 patients) after a mean follow-up time of 79.9 months (range, 12-251 months), female sex ( P = .016), multiple nodules ( P = .017), and lack of postoperative LT4 therapy ( P = .0009) were predictive factors of recurrence. Reoperation was performed in 20 patients (7.4%); factors that were predictive of reoperation were the presence of multiple nodules ( P = .008), resected thyroid weight ( P = .00006), and lack of postoperative hormonal therapy ( P = .0005). CONCLUSIONS: Thyroid lobectomy for unilateral non-toxic goiter, when combined with suppressive or substitutive thyroxin therapy, resulted in a low rate of recurrence and reoperation in an endemic area. PMID- 15657584 TI - Near-total parathyroidectomy is beneficial for patients with secondary and tertiary hyperparathyroidism. AB - BACKGROUND: Despite several options for the extent of parathyroidectomy in secondary and tertiary hyperparathyroidism, <5% of patients are referred for surgical procedures. The purpose of this study was to identify beneficial outcomes of near-total parathyroidectomy (NTPTX) in this population. METHODS: NTPTX (a vascularized parathyroid remnant is left in situ, and cryopreservation is performed) was completed in 142 patients with secondary (n = 93 patients) and tertiary (n = 49 patients) hyperparathyroidism, and was guided by intraoperative intact PTH (IOPTH). RESULTS: One hundred thirty-six of 142 patients (96%) remain cured at a mean follow-up time of 23 +/- 26 months. Six patients had missed mediastinal (n = 4 patients) or cervical (n = 2 patients) parathyroid glands. No patient required re-resection of a cervical parathyroid remnant. One patient had hypoparathyroidism and underwent autografting with cryopreserved tissue. NTPTX reduced parathyroid mass (4526 +/- 4515 mg to 77 +/- 56 mg) and IOPTH (1171 +/- 1252 pg/mL to 101 +/- 127 pg/mL). Fatigue and musculoskeletal pain resolved promptly after operation. Patients with tertiary hyperparathyroidism experienced an improvement in bone density of 3.6% per year. NTPTX achieved full healing of extremity ulcers without amputations or perioperative death in 22 patients with calciphylaxis. CONCLUSIONS: NTPTX effectively treats secondary and tertiary hyperparathyroidism in >95% patients. IOPTH monitoring and cryopreservation are valuable procedural adjuncts. Limb salvage in calciphylaxis, symptomatic relief, and improved bone density are substantial benefits to recommend this procedure to patients with renal hyperparathyroidism. PMID- 15657585 TI - GCMB gene, a master regulator of parathyroid gland development, expression, and regulation in hyperparathyroidism. AB - BACKGROUND: The glial cell missing gene, GCMB , encodes a transcription factor, which is a master regulator of parathyroid development. We postulated that the GCMB gene might play a role in parathyroid tumorigenesis in hyperparathyroidism. METHODS: We used real-time quantitative reverse transcriptase polymerase chain reaction to study GCMB mRNA expression in parathyroid tissue: normal (n = 3), hyperplastic (n = 16), adenomas (n = 19), and cancers (n = 8). In primary parathyroid culture, the effect of CaCl 2 on parathyroid hormone secretion and GCMB mRNA expression was studied by using enzyme-linked immunosorbent assay and reverse transcriptase polymerase chain reaction, respectively. RESULTS: GCMB mRNA expression was lower in normal (0.4 +/- 0.1, mean +/- standard error of mean) parathyroid glands as compared to adenoma (3.5 +/- 1.7), hyperplasia (3.2 +/- 1.3 primary hyperparathyroidism [n = 11] and 7.6 +/- 4.8 secondary hyperparathyroidism [n = 5]), and cancer (3.6 +/- 1.3) ( P = .001). There was no difference in the level of GCMB mRNA expression between parathyroid adenoma, hyperplasia, and cancer. In primary culture of parathyroid adenoma (n = 9) and hyperplasia (n = 2), parathyroid hormone secretion was increased 2- to 15-fold with low calcium concentration (0.5 to 4.0 mmol/L CaCl 2 from 2 to 6 hours, P < .005). The level of GCMB mRNA expression was down-regulated with lower extracellular CaCl 2 concentration ( P < .005). CONCLUSIONS: GCMB expression is upregulated in abnormal parathyroid glands of hyperparathyroidism and decreases in response to hypocalcemia. The GCMB transcription factor might mediate the effect of calcium on parathyroid cell parathyroid hormone expression/secretion. PMID- 15657586 TI - Gastric carcinoid tumors in multiple endocrine neoplasia-1 patients with Zollinger-Ellison syndrome can be symptomatic, demonstrate aggressive growth, and require surgical treatment. AB - BACKGROUND: Gastric carcinoid tumors occur in 15% to 50% of patients with multiple endocrine neoplasia-1/Zollinger-Ellison syndrome (MEN-1/ZES) but are thought to be benign. We report 5 patients with MEN-1/ZES with symptomatic, aggressive gastric carcinoid tumors that required surgical procedures. METHODS: This was a retrospective chart review. RESULTS: Each patient had MEN-1/ZES. Each patient had innumerable gastric carcinoid tumors with symptoms. The fasting gastrin level was 47,000 pg/mL (normal, <200 pg/mL); the basal acid output was 79 mEq/hr (n = 3), and the age at surgical exploration was 47 +/- 6 years, with a duration of MEN-1 of 21 +/- 3 years and of ZES of 15 +/- 2 years. All patients had elevated 5-HIAA or serotonin levels. Somatostatin receptor scintigraphy showed increased stomach uptake in 4 patients (80%). Four patients had a total gastrectomy; 4 patients had lymph node metastases removed, and 3 patients had liver metastases resected. One patient who did not have a total gastrectomy had liver carcinoid metastases. CONCLUSIONS: These results demonstrate that gastric carcinoid tumors in patients with longstanding MEN-1/ZES may be symptomatic, aggressive, and metastasize to the liver. With increased long-term medical treatment and life expectancy, these tumors will become an important determinant of survival. PMID- 15657587 TI - Large remnant 131I ablation as an alternative to completion/total thyroidectomy in the treatment of well-differentiated thyroid cancer. AB - BACKGROUND: An alternative to completion thyroidectomy for well-differentiated thyroid carcinoma is to ablate the remnant lobe with 131 I. The purpose of this study is to review our own experience with large remnant ablation. METHODS: A retrospective review of 169 patients with well-differentiated thyroid cancer treated at one institution over a 14-year period was undertaken. Seventy-one patients who underwent partial thyroidectomy (PT) followed by 131 I ablation were identified. This group was compared to 98 patients treated with total thyroidectomy (TT). RESULTS: Mean follow-up was 6.2 years for the 71 PT + 131 I versus 4.7 years for the 98 TT patients (P = .184). Recurrence occurred in 4 of 71 PT + I 131 patients (5.6%) versus 9 of 98 TT patients (9.2%) (P = .393). Other than a tendency for the size of the primary to be slightly larger and for the histology to be follicular carcinoma in the PT + 131 I patients, the 2 groups were nearly identical in age, gender, and other prognostic factors such as capsular invasion and metastases. CONCLUSIONS: Large-dose ablation with 131 I is a viable alternative to completion thyroidectomy. Recurrence rates over an average 6-year period are similar to TT. Long-term monitoring of these cohorts is required. PMID- 15657588 TI - Early parathyroidectomy increases bone mineral density in patients with mild primary hyperparathyroidism: a prospective and randomized study. AB - BACKGROUND: There is an ongoing controversy regarding how to take care of patients with mild primary hyperparathyroidism (PHPT) and how to grade their disease activity in terms of bone parameters. This prospective and randomized study was undertaken to evaluate skeletal effects of delayed surgical treatment in such patients. METHODS: Fifty patients with mild PHPT (serum calcium, 2.55 to 2.95 mmol/L; 10.2 to 11.8 mg/dL) were randomized to parathyroidectomy either at diagnosis or 1 year later. Hip and spine bone mineral density (BMD, determined by dual energy x-ray absorptiometry), bone alkaline phosphatase in serum, osteocalcin and beta-CrossLaps in plasma, and calcium in urine were measured in all patients at inclusion in the study and 1 and 2 years later. RESULTS: The skeletal effects of mild PHPT varied with anatomy and time of exposure. Parathyroidectomy decreased all biochemical bone markers ( P < .0001) and increased lumbar spine BMD ( P < .05) equally in both groups, even in patients without overt osteoporosis, whereas hip BMD was increased ( P < .05) in the early intervention group only. CONCLUSION: Prolonged exposure to mild and seemingly stable PHPT is a risk factor for hip fractures, which adds to other reasons for surgical treatment of this condition without delay regardless of serum calcium levels. PMID- 15657589 TI - Importance of adrenal incidentaloma in patients with a history of malignancy. AB - BACKGROUND: Adrenal incidentaloma presents a frequent finding in patients with a history of malignancy. This study was carried out to determine whether imaging techniques can discriminate between a malignant and a benign adrenal tumor and subsequently select candidates for adrenal surgery. METHODS: Beginning in July 1995, oncologic patients with adrenal incidentaloma underwent abdominal ultrasonography, computed tomography (CT), magnetic resonance imaging (MRI), and [ 18 ]fluoro-2-deoxy-D-glucose positron emission tomography (18-FDG-PET). Consecutively in all patients adrenalectomy was performed. Histologic findings were compared with the imaging results. RESULTS: In 42 patients, 33 to 79 years old (mean age, 58 years), 44 adrenal lesions were detectable. Two patients presented with bilateral adrenal masses. At operation, 43 adrenal resections and 3 biopsies were performed. Histologic examination revealed 31 metastases (71%) and 13 adrenal adenomas (29%). In metastases the sensitivity of ultrasonography, CT, MRI, and 18-FDG-PET was 66%, 81%, 100%, and 100%, respectively. For adrenal adenoma, the sensitivity of ultrasonography, CT, MRI, and 18-FDG-PET was 46%, 39%, 100%, and 54%, respectively. CONCLUSIONS: In oncologic patients with indeterminate adrenal tumors MRI and 18-FDG-PET provide accurate differentiation between metastases and benign adenomas. Positive results of these 2 imaging modalities are highly suggestive for metastatic disease. In cases of negative MRI and 18-FDG-PET results the adrenal lesion corresponds to a benign adenoma that needs no surgical intervention. PMID- 15657590 TI - Insulin-like growth factor 1 signaling in human gastrointestinal carcinoid tumor cells. AB - Background Insulin-like growth factor 1 (IGF-1) is an autocrine regulator of carcinoid tumors. Blockade of IGF-1 signaling has been proposed as a therapeutic target in the treatment of patients with carcinoid syndrome. We hypothesized that the induction of parallel raf-1/MEK1 pathways will block IGF-1-mediated chromogranin A (CgA) maintenance. Methods Human gastrointestinal carcinoid tumor cells (BON) were treated with IGF-1 (0-500 ng/mL). Raf-1/MEK1 activation was achieved with an estrogen-inducible raf-1 vector that was transduced into BON cells. Activation of IGF-1/raf-1 pathways was determined by phosphorylation of downstream targets p70s6 and ERK1/2. The secreted and intercellular levels of CgA were measured in conditioned media and whole cell extracts by Western and enzyme linked immunosorbent assay analysis. Results IGF-1 and raf-1 pathways were activated successfully in BON cells, as shown by high levels of phosphorylated p70s6 and phosphorylated ERK1/2, respectively. Treatment of BON cells with IGF-1 stimulated the release of CgA, while high intracellular CgA levels were maintained. The activation of raf-1/MEK1 reversed the effect of IGF-1 treatment by the depletion of intracellular CgA. Conclusions The induction of the raf 1/MEK1 pathway blocks IGF-1-mediated intracellular neuroendocrine hormone regulation. Therefore, raf-1/MEK1 activation may be a viable method to block IGF 1-mediated cellular effects and serve as a therapeutic target in gastrointestinal carcinoid tumors. PMID- 15657591 TI - Detection of multiple gland primary hyperparathyroidism in the era of minimally invasive parathyroidectomy. AB - BACKGROUND: A focused surgical approach for primary hyperparathyroidism relies on the ability of preoperative imaging and intraoperative parathyroid hormone monitoring (IOPTH) to detect multiple gland disease (MGD). The study objective was to determine the best predictor for MGD. METHODS: First time parathyroidectomy was performed on 233 patients with primary hyperparathyroidism who underwent preoperative sestamibi imaging, ultrasound, and IOPTH between December 1999 and January 2004. RESULTS: Single gland disease (SGD) was found in 204 (88%) and MGD in 23 (10%) patients. Hyperparathyroidism persisted in 6 of 233 patients (2.6%). For patients with MGD, sestamibi imaging correctly predicted MGD in 2 of 23 (9%) patients, incorrectly showed SGD in 9 of 23 (39%), and was negative in 12 of 23 (52%). Ultrasound correctly predicted MGD in 6 of 23 (26%) patients, incorrectly predicted SGD in 6 of 23 (39%), and was negative in 8 of 23 (35%). Together sestamibi imaging and ultrasound predicted MGD in 7 of 23 (30%) patients, incorrectly predicted SGD in 7 of 23 (30%), was negative in 7 of 23 (30%), and was discordant in 10 of 23 (5%). IOPTH indicated MGD in 15 of 18 (83%) patients but falsely predicted cure after single gland excision in 3 of 18 (17%). The combination of sestamibi imaging, ultrasound, and IOPTH detected MGD in 16 of 18 (89%) patients. CONCLUSION: Ultrasound was more sensitive for detecting MGD than sestamibi imaging. Ultrasound and sestamibi imaging together provided information warranting a bilateral approach in 70% of patients with MGD. IOPTH was the most sensitive for MGD, but combining all 3 tests was the best predictor, identifying the majority of patients with MGD. PMID- 15657592 TI - Risk factors of paralysis and functional outcome after recurrent laryngeal nerve monitoring in thyroid surgery. AB - BACKGROUND: Recurrent laryngeal nerve monitoring (RLNM) has been suspected to reduce postoperative RLN paralysis (RLNP). However, functional outcome of RLNM in comparison with no nerve identification and visual nerve identification only has not been analyzed. METHODS: Analysis of 16,448 consecutive multi-institutional operations resulted in 29,998 nerves at risk. Three groups of different RLN treatment were compared: group 1, no RLN identification; group 2, visual RLN identification; and group 3, visual RLN identification and electromyographic monitoring. RLNM was performed with a bipolar needle electrode that was placed through the cricothyroid ligament into the vocal muscle. RESULTS: Risk factors for permanent RLNP were recurrent benign and malignant goiter (odds ratios, [ORs]), 4.7, and 6.7, respectively), primary surgery in thyroid malignancy (OR, 2.0), lobectomy (OR, 1.8), no nerve identification (OR, 1.4), low or medium volume hospital (OR, 1.3), and low volume surgeons (OR, 1.2). CONCLUSIONS: Based on these data, visual nerve identification was identified to be the gold standard of RLN treatment in thyroid surgery. RLNM is a promising tool for nerve identification and protection in extended thyroid resection procedures. However, because of the overall low frequency of RLNP, no statistical difference compared with visual nerve identification only was reached in the setting of this study. PMID- 15657595 TI - Comm function in commissural axon guidance: cell-autonomous sorting of Robo in vivo. AB - Commissureless (Comm) controls axon guidance across the Drosophila melanogaster midline by regulating surface levels of Robo, the receptor for the midline repellent Slit. Two different models have been proposed for how Comm regulates Robo: a 'sorting' model and a 'clearance' model, both based on studies using heterologous cells in vitro. Here, we test these two models in vivo. We establish a genetic rescue assay for Comm, and use this assay to show that midline crossing does not require the presence of Comm in midline cells, as proposed by the clearance model. Moreover, by monitoring the trafficking of a Robo-green fluorescent protein (GFP) fusion in living embryos, we demonstrate that Comm prevents the delivery of Robo-GFP to the growth cone, as predicted by the sorting model. It has also been suggested that Comm must be ubiquitinated by the Nedd4 ubiquitin ligase. We show here, however, that ubiquitination of Comm is not required for its function in vitro or in vivo, and that Nedd4 is unlikely to function in axon guidance at the midline. PMID- 15657596 TI - Coactivation and timing-dependent integration of synaptic potentiation and depression. AB - Neuronal synaptic connections can be potentiated or depressed by paired pre- and postsynaptic spikes, depending on the spike timing. We show that in cultured rat hippocampal neurons a calcium/calmodulin-dependent protein kinase II (CaMKII) mediated potentiation process and a calcineurin-mediated depression process can be activated concomitantly by spike triplets or quadruplets. The integration of the two processes critically depends on their activation timing. Depression can cancel previously activated potentiation, whereas potentiation tends to override previously activated depression. The time window for potentiation to dominate is about 70 ms, beyond which the two processes cancel. These results indicate that the signaling machinery underlying spike timing-dependent plasticity (STDP) may be separated into functional modules that are sensitive to the spatiotemporal dynamics (rather than the amount) of calcium influx. The timing dependence of modular interaction provides a quantitative framework for understanding the temporal integration of STDP. PMID- 15657597 TI - A representation of the hazard rate of elapsed time in macaque area LIP. AB - The capacity to anticipate the timing of environmental cues allows us to allocate sensory resources at the right time and prepare actions. Such anticipation requires knowledge of elapsed time and of the probability that an event will occur. Here we show that neurons in the parietal cortex represent the probability, as a function of time, that a salient event is likely to occur. Rhesus monkeys were trained to make eye movements to peripheral targets after a light dimmed. Within a block of trials, the 'go' times were drawn from either a bimodal or unimodal distribution of random numbers. Neurons in the lateral intraparietal area showed anticipatory activity that revealed an internal representation of both elapsed time and the probability that the 'go' signal was about to occur (termed the hazard rate). The results indicate that the parietal cortex contains circuitry for representing the time structure of environmental cues over a range of seconds. PMID- 15657598 TI - Large-scale changes in dendritic structure during reorganization of adult somatosensory cortex. AB - In adult rat somatosensory cortex (S1), neurons are biased and have less dendritic arbor close to the border between the forepaw and lower jaw representations. Changes in sensory experience cause changes in the functional organization of the neocortex. Therefore, we examined the morphology of neurons in the reorganized region of S1 after forepaw denervation. We found that during reorganization dendritic arbors changed to reflect the new location of the border. PMID- 15657599 TI - Central amygdala ERK signaling pathway is critical to incubation of cocaine craving. AB - Using a rat model of craving and relapse, we have previously found time-dependent increases in cue-induced cocaine seeking over the first months of withdrawal from cocaine, suggesting that drug craving incubates over time. Here, we explored the role of the amygdala extracellular signal-regulated kinase (ERK) signaling pathway in this incubation. Cocaine seeking induced by exposure to cocaine cues was substantially higher after 30 withdrawal days than after 1 withdrawal day. Exposure to these cues increased ERK phosphorylation in the central, but not the basolateral, amygdala after 30 d, but not 1 d, of withdrawal. After 30 d of withdrawal from cocaine, inhibition of central, but not basolateral, amygdala ERK phosphorylation decreased cocaine seeking. After 1 d of withdrawal, stimulation of central amygdala ERK phosphorylation increased cocaine seeking. Results suggest that the incubation of cocaine craving is mediated by time-dependent increases in the responsiveness of the central amygdala ERK pathway to cocaine cues. PMID- 15657601 TI - Research network: EUROGLYCANET: a European network focused on congenital disorders of glycosylation. PMID- 15657600 TI - Dynamics of motion signaling by neurons in macaque area MT. AB - Most neurons in macaque area MT are selective for the direction of stimulus motion. By comparing direction selectivity for gratings and plaids, we classified MT neurons as pattern direction selective (PDS) or component direction selective (CDS). We compared the time course of responses in CDS and PDS neurons in opiate anesthetized macaques, using a rapid pseudorandom sequence of gratings and plaids that moved in different directions. On average, responses began 6 ms earlier in CDS neurons than in PDS neurons. More importantly, the pattern-selective responses of PDS neurons did not reach their fully selective state until 50-75 ms after the responses of CDS neurons had stabilized. The population motion response of MT is therefore initially dominated by component motion signals, and does not completely represent pattern motion until substantially later. The circuits that compute pattern motion take more time to finish their work than those signaling component motion. PMID- 15657602 TI - Statistical genetics: usual suspects in complex disease. PMID- 15657603 TI - Identification of a novel mevalonate kinase gene mutation in combination with the common MVK V377I substitution and the low-penetrance TNFRSF1A R92Q mutation. AB - The hyperimmunoglobulinemia D and periodic fever syndrome (HIDS) is an autosomal recessively inherited autoinflammatory disease caused by mutations in the mevalonate kinase (MVK) gene on chromosome 12q24, which lead to a depressed enzymatic activity of mevalonate kinase (MK). TNF-receptor associated periodic syndrome (TRAPS), on the other hand, is the most frequent autosomal dominantly inherited periodic fever syndrome due to mutations in exons 2-4 and 6 of the TNFRSF1A gene on chromosome 12p13.2. We describe a girl with heterozygosity for the common MVK V377I mutation and for a novel T(1132) --> C transition, leading to the exchange of serine (TCC) by proline (CCC) at amino-acid position 378. Interestingly, our patient presented only with mild clinical features typical of HIDS and slightly increased immunoglobulin D levels, but a distinctly diminished MK activity. The girl was also heterozygous for the TNFRSF1A R92Q low-penetrance mutation, which may have significant proinflammatory effects. However, at the time of presentation, the patient had no TRAPS-associated symptoms. PMID- 15657604 TI - Association of BDNF with restricting anorexia nervosa and minimum body mass index: a family-based association study of eight European populations. AB - Eating disorders (ED), such as anorexia nervosa (AN) and bulimia nervosa (BN), are complex psychiatric disorders where different genetic and environmental factors are involved. Several lines of evidence support that brain-derived neurotrophic factor (BDNF) plays an essential role in eating behaviour and that alterations on this neurotrophic system participates in the susceptibility to both AN and BN. Accordingly, intraventricular administration of BDNF in rats determines food starvation and body weight loss, while BDNF or its specific receptor NTRK2 knockout mice develop obesity and hyperphagia. Case-control studies also suggest a BDNF contribution in the aetiology of ED: we have previously reported a strong association between the Met66 variant within the BDNF gene, restricting AN (ANR) and minimum body mass index (minBMI) in a Spanish sample, and a positive association between the Val66Met and -270C/T BDNF SNPs and ED in six different European populations. To replicate these results, avoiding population stratification effects, we recruited 453 ED trios from eight European centres and performed a family-based association study. Both haplotype relative risk (HRR) and haplotype-based haplotype relative risk (HHRR) methods showed a positive association between the Met66 allele and ANR. Consistently, we also observed an effect of the Met66 variant on low minBMI and a preferential transmission of the -270C/Met66 haplotype to the affected ANR offspring. These results support the involvement of BDNF in eating behaviour and further suggest its participation in the genetic susceptibility to ED, mainly ANR and low minBMI. PMID- 15657605 TI - Genetic and clinical characteristics of maturity-onset diabetes of the young in Chinese patients. AB - In Caucasians, maturity-onset diabetes of the young (MODY) is mostly caused by mutations in the hepatocyte nuclear factor (HNF)-1alpha (MODY3) and glucokinase (MODY2) genes. Most Japanese MODY patients, however, are not linked to known MODY genes. In this study, we examined the genetic and clinical characteristics of Chinese subjects with MODY. The study included 146 unrelated families fulfilling the minimum criteria for MODY: two consecutive generations of type II diabetes with at least one member diagnosed under the age of 25. We screened for mutations in the HNF-4alpha (MODY1), MODY2 and MODY3 genes by direct sequencing. Antibody to glutamic acid decarboxylase (GAD-Ab) was measured in subjects with MODY of unknown cause (MODYX). Insulin resistance index and other clinical data were compared in sex-, age- and duration-matched MODY3 and MODYX patients. In all, 13 families had MODY3 mutations and two had MODY2 mutations. No MODY1 mutation was found. Four of the 12 different MODY3 mutations were newly identified novel mutations (Q243E, A311D, P379R and P488fsdelC). In subjects with MODYX, 3% were GAD-Ab positive and 60% were overweight. Compared to MODY3 patients, MODYX patients had higher body mass index (P<0.02), higher insulin resistance index (P=0.001) and triglyceride level (P<0.02), lower HDL level (P=0.001) and more hypertension (P<0.05), but no significant difference in the prevalence of diabetic complications. In conclusion, MODY3 and MODY2 account for only 9 and 1%, respectively, of Chinese MODY. A majority of Chinese MODY patients are due to defects in unknown genes and appear to be characterized by insulin resistance. PMID- 15657607 TI - Information and informed consent in a longitudinal screening involving children: a questionnaire survey. AB - This empirical study explores participants' perceptions of information and understanding of their children's and their own involvement in a longitudinal screening, the ABIS Study. ABIS (All Babies In Southeast Sweden) is a multicentre, longitudinal research screening for Type 1 diabetes and multifactorial diseases involving 17 005 children and their families. For this study, a random selection of mothers was made, using perinatal questionnaire serial numbers from the ABIS study. In total, 293 of these mothers completed an anonymous questionnaire (response rate 73.3%). Our findings from the questionnaire indicate a marked difference between the reported satisfaction with and understanding of the information provided on the one hand and the significant lack of knowledge of some of the aims and methods of the ABIS screening on the other, namely concerning high-risk identification of involved children, potential prevention and future questionnaires. Two questions evoked by our results are: (1) what information is required for participants in longitudinal studies involving children? and (2) how do we ensure and sustain understanding, and thus in a prolonging, informed consent in these studies? This study underlines the importance of an increased understanding of the ethical issues that longitudinal research on children raise and the need to discuss how information and informed consent strategies should be analysed and designed in longitudinal studies. PMID- 15657606 TI - X-chromosome as a marker for population history: linkage disequilibrium and haplotype study in Eurasian populations. AB - Linkage disequilibrium (LD) structure is still unpredictable because the interplay of regional recombination rate and demographic history is poorly understood. We have compared the distribution of LD across two genomic regions differing in crossing-over activity -- Xq13 (0.166 cM/Mb) and Xp22 (1.3 cM/Mb) -- in 15 Eurasian populations. Demographic events predicted to increase the LD level -- genetic drift, bottleneck and admixture - had a very strong impact on extent and patterns of regional LD across Xq13 compared to Xp22. The haplotype distribution of the DXS1225-DXS8082 microsatellites from Xq13 exhibiting strong association in all populations was remarkably influenced by population history. European populations shared one common haplotype with a frequency of 25-40%. The Volga-Ural populations studied, living at the geographic borderline of Europe, showed elevated LD as well as harboring a significant fraction of haplotypes originating from East Asia, thus reflecting their past migrations and admixture. In the young Kuusamo isolate from Finland, a bottleneck has led to allelic associations between loci and shifted the haplotype distribution, but has much less affected single microsatellite allele frequencies compared to the main Finnish population. The data show that the footprint of a demographic event is longer preserved in haplotype distribution within a region of low crossing-over rate, than in the information content of a single marker, or between actively recombining markers. As the knowledge of LD patterns is often chosen to assist association mapping of common disease, our conclusions emphasize the importance of understanding the history, structure and variation of a study population. PMID- 15657608 TI - Evolutionary genetics: genetics of lactase persistence--fresh lessons in the history of milk drinking. PMID- 15657609 TI - CNGB3 mutations account for 50% of all cases with autosomal recessive achromatopsia. AB - Achromatopsia is a congenital, autosomal recessively inherited disorder characterized by a lack of color discrimination, low visual acuity (<0.2), photophobia, and nystagmus. Mutations in the genes for CNGA3, CNGB3, and GNAT2 have been associated with this disorder. Here, we analyzed the spectrum and prevalence of CNGB3 gene mutations in a cohort of 341 independent patients with achromatopsia. In 163 patients, CNGB3 mutations could be identified. A total of 105 achromats carried apparent homozygous mutations, 44 were compound (double) heterozygotes, and 14 patients had only a single mutant allele. The derived CNGB3 mutation spectrum comprises 28 different mutations including 12 nonsense mutations, eight insertions and/or deletions, five putative splice site mutations, and three missense mutations. Thus, the majority of mutations in the CNGB3 gene result in significantly altered and/or truncated polypeptides. Several mutations were found recurrently, in particular a 1 bp deletion, c.1148delC, which accounts for over 70% of all CNGB3 mutant alleles. In conclusion, mutations in the CNGB3 gene are responsible for approximately 50% of all patients with achromatopsia. This indicates that the CNGB3/ACHM3 locus on chromosome 8q21 is the major locus for achromatopsia in patients of European origin or descent. PMID- 15657610 TI - Segmental haplosufficiency: transmitted deletions of 2p12 include a pancreatic regeneration gene cluster and have no apparent phenotypic consequences. AB - Segmental aneuploidy usually has phenotypic consequences but unbalanced rearrangements without phenotypic consequences have also been reported. In particular, harmless deletions of G-dark bands 5p14 and 16q21 have each been found in more than one independent family. Here, we report two families that were ascertained at prenatal diagnosis and had similar overlapping deletions that removed most of the gene poor G-dark band 2p12. PCR mapping showed that the deletions had a minimum size of 6.1 and 6.9 Mb with at least 13 hemizygous loci including a cluster of six pancreatic islet-regenerating genes. These deletions had no apparent phenotypic consequences in eight family members. In contrast, a third family was ascertained through a child with Wilm's tumour; both the child and his mother had more proximal deletions, developmental delay and some dysmorphic features. The deletion had a minimum size of 5.7 Mb and extended into the gene-rich area of 2p11.2. These results are consistent with the idea that there may be segments of the genome that are consistently haplosufficient. The introduction of higher resolution methods of dosage analysis into diagnostic laboratories is already revealing more transmitted abnormalities of uncertain significance. As a result, published cases of transmitted imbalances have been collected as a guide to the possible significance of such findings in the future (see the 'Chromosome Anomaly Collection' at www.som.soton.ac.uk/research/geneticsdiv). PMID- 15657611 TI - A 2.3 Mb duplication of chromosome 8q24.3 associated with severe mental retardation and epilepsy detected by standard karyotype. AB - Chromosome duplications are found in about 2% of subjects with a typical chromosomal phenotype but their frequency is likely to be higher, as suggested by the first array-CGH data. According to the orientation of the duplicated segment, duplications may be in tandem or inverted. The latter are usually associated with a distal deletion. We studied a de novo 2.3 Mb inverted duplication of 8q24.3 without apparently associated deletion in a subject with profound psychomotor retardation, idiopathic epilepsy and growth delay. In spite of its small size, the presence of the rearrangement was suspected on standard karyotypes (approximately 400 bands) and later confirmed by Fluorescent in situ hybridization (FISH) analysis. We hypothesize that the GRINA gene, a glutamate binding subunit of NMDA receptor ion channel lying within the duplicated segment, may be responsible for the epilepsy. This paper confirms that small subtelomeric de novo duplications may be responsible for mental retardation, facial dysmorphisms and/or congenital malformations, although their presence may be overlooked by FISH analysis. PMID- 15657612 TI - Linkage disequilibrium patterns vary substantially among populations. AB - A major initiative to create a global human haplotype map has recently been launched as a tool to improve the efficiency of disease gene mapping. The 'HapMap' project will study common variants in depth in four (and to a lesser degree in up to 12) populations to catalogue haplotypes that are expected to be common to all populations. A hope of the 'HapMap' project is that much of the genome occurs in regions of limited diversity such that only a few of the SNPs in each region will capture the diversity and be relevant around the world. In order to explore the implications of studying only a limited number of populations, we have analyzed linkage disequilibrium (LD) patterns of three 175-320 kb genomic regions in 16 diverse populations with an emphasis on African and European populations. Analyses of these three genomic regions provide empiric demonstration of marked differences in frequencies of the same few haplotypes, resulting in differences in the amount of LD and very different sets of haplotype frequencies. These results highlight the distinction between the statistical concept of LD and the biological reality of haplotypes and their frequencies. The significant quantitative and qualitative variation in LD among populations, even for populations within a geographic region, emphasizes the importance of studying diverse populations in the HapMap project to assure broad applicability of the results. PMID- 15657613 TI - Haplotype structure of TNFRSF5-TNFSF5 (CD40-CD40L) and association analysis in systemic lupus erythematosus. AB - Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease that is caused by genetic and environmental factors. The tumour necrosis factor (TNF) superfamily of genes play a central role in immune regulation and have been proposed to be involved in the development of SLE. TNFRSF5 (CD40) falls on 20q11 13, a region linked with SLE in three independent genome-wide studies. TNFSF5 (CD40L) falls on Xq26 and is the ligand for TNFRSF5. Seven single-nucleotide polymorphisms (SNPs) in CD40 and eight SNPs in CD40L were looked at for linkage disequilibrium (LD) and haplotype analysis in European-Caucasians. Limited haplotype diversity was observed across CD40 and CD40L, and >97% of the diversity was captured. We also examined the association of SNPs and haplotypes in CD40 and CD40L with SLE in European-Caucasians. There was no evidence of association for CD40 or CD40L in 408 European-Caucasian families with SLE from UK. Haplotype tagging SNPs (htSNPs) are made known, which will facilitate analysis for susceptibility in other autoimmune diseases and risk for infectious disease. PMID- 15657614 TI - LHON/MELAS overlap syndrome associated with a mitochondrial MTND1 gene mutation. AB - Pathogenic point mutations in the mitochondrial MTND1 gene have previously been described in association with two distinct clinical phenotypes -- Leber hereditary optic neuropathy (LHON) and mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes (MELAS). Here we report the first heteroplasmic mitochondrial DNA (mtDNA) point mutation (3376G>A) in the MTND1 gene associated with an overlap syndrome comprising the clinical features of both LHON and MELAS. Muscle histochemistry revealed subtle mitochondrial abnormalities, while biochemical analysis showed an isolated complex I deficiency. Our findings serve to highlight the growing importance of mutations in mitochondrial complex I structural genes in MELAS and its associated overlap syndromes. PMID- 15657615 TI - Polymorphisms in APOA1 and LPL genes are statistically independently associated with fasting TG in men with CAD. AB - The objective of this paper was to identify the single nucleotide polymorphisms (SNPs) that show unshared effects on plasma triglyceride (TG) levels and to investigate whether these SNPs show statistically independent effects on plasma TG levels. In total, 59 polymorphisms in 20 genes involved in lipid metabolism were investigated. Polymorphisms were selected for a multivariate ANOVA model if they showed an univariate association with TG (after adjustment for HDL-C and LDL C) in more than 50% of bootstrap samples that were made from the original data. The multivariate model included 512 men with coronary artery disease from the REGRESS study who were completely genotyped for eight polymorphisms selected in the univariate procedure (ie, APOA1 G(-75)A, ABCA1 C(-477)T, ABCA1 G1051A, APOC3 T3206G, APOE Arg158Cys, LIPC C(-514)T, LPL Asn291Ser and LPL Ser447Stop). The gene variants APOA1 G(-75)A (P=0.04) and LPL Asn291Ser (P=0.03) were significantly associated with plasma TG levels in this multivariate analysis. The eight polymorphisms explained 8.9% of the variation in plasma TG levels. In conclusion, this study showed statistically independent effects of gene variants in the APOA1 and LPL genes on fasting plasma levels of TG. Nevertheless, only a small part of variation in TG levels could be explained by the polymorphisms. PMID- 15657616 TI - Defective protein glycosylation in patients with cutis laxa syndrome. AB - Congenital cutis laxa is a genetically heterogeneous condition presenting with loose and redundant skin folds, decreased elasticity of the skin, connective tissue involvement and a highly variable spectrum of associated features. The most common forms are inherited in an autosomal recessive or dominant fashion. Fibulin 5 and elastin mutations were detected in a limited number of patients, but in most cases the etiology is not known. Based on a previous observation of an abnormal transferrin isoelectric focusing pattern in a patient with cutis laxa indicating an N-glycosylation defect, we performed a screening for disorders of protein glycosylation in unrelated children with cutis laxa syndrome, including a recently developed test for defective O-glycosylation. Here, we describe five patients from consanguineous marriages with a cutis laxa syndrome with skeletal and joint involvement, developmental delay and neurological findings. Three of these five children have an inborn error of glycan biosynthesis affecting the synthesis of both N- and O-linked glycans. Two patients had normal glycosylation patterns. All known causes of secondary glycosylation disorders were excluded in the children. No mutations were found in the FBLN5 gene. In conclusion, we have identified a new combined glycosylation defect with a distinct clinical phenotype. Our results suggest that a combined defect of glycosylation might be a causative factor in congenital cutis laxa. This is the first report where abnormal N- and O-linked glycosylation is implicated in the etiology of cutis laxa syndrome. PMID- 15657617 TI - Cascade testing in familial hypercholesterolaemia: how should family members be contacted? AB - Cascade testing or screening provides an important mechanism for identifying people at risk of a genetic condition. For some autosomal dominant conditions, such as familial hpercholesterolaemia (FH), identifying relatives allows for significant health-affecting interventions to be administered, which can extend a person's life expectancy significantly. However, cascade screening is not without ethical implications. In this paper, we examine one ethically contentious aspect of cascade screening programmes, namely the alternative methods by which relatives of a proband can be contacted. Should the proband be invited to contact his or her family members, or should the screening programme contact family members directly? We argue that direct contact is an ethically justifiable method of contact tracing in cascade screening for FH. Not only has this method already been utilised without adverse effects, an examination of the ethical arguments against it shows these are unsubstantiated. We describe several criteria that, if met, will allow an appropriate balance to be struck between maximising the efficiency of family tracing and respecting the interests of probands and their relatives. PMID- 15657619 TI - Sequence variation in the ATP8B1 gene and intrahepatic cholestasis of pregnancy. AB - Intrahepatic cholestasis of pregnancy (ICP) is a cholestatic condition that may affect women during the third trimester of pregnancy. Symptoms experienced by these women generally resolve spontaneously following delivery, but prior to delivery the fetus is at increased risk of intrauterine distress and sudden intrauterine death. The genetic etiology of most cases of ICP is unknown, although heterozygous carriers of mutations causing progressive familial intrahepatic cholestasis (PFIC) diseases may experience ICP. When examining linkage to known cholestasis genes, affected members of four Finnish ICP families shared haplotypes around ATP8B1, the gene responsible for PFIC1. This gene was subsequently screened in 176 familial and sporadic ICP patients. A total of 17 sequence changes were detected, five exonic and 12 intronic. No intronic change was associated with ICP in sporadic cases. Four intronic changes segregated with ICP in three families, a different change in each of two families and three changes in another family, although the significance of this is currently unknown. Three exonic changes were nonsynonymous, one (in exon 23) is probably a polymorphism while two predict novel amino-acid replacements (N45T and K203R). These changes, in exons 2 and 7, were detected in one individual each, and may have predisposed these individuals to ICP. In conclusion, although the exon 2 and 7 changes may have functioned as risk alleles, ATP8B1 is probably not a major gene contributing to the occurrence of ICP. PMID- 15657618 TI - A common CTLA4 haplotype associated with coeliac disease. AB - Coeliac disease is a common enteropathy with a strong inherited risk characterised by dietary wheat, rye and barley induced T-cell activation. Although there is replicated linkage to 2q33, results are inconsistent from association studies of the most promising candidate genes: the CD28/CTLA4/ICOS cluster. CTLA4 plays a key role in regulating T lymphocyte mediated inflammatory responses, and variants in the 3' region influence development of diabetes and thyroid disease. We genotyped CTLA4 variants (-1722 C/T, -658 T/C, -318 C/T, +49 A/G, +1822 C/T, CT60 A/G) to tag all common haplotypes (>5% frequency) and an ICOS variant (IVS+173 C/T) in 340 white UK Caucasian coeliac disease cases. Strict ascertainment criteria for coeliac cases required both villous atrophy at diagnosis and positive serology. In total, 973 healthy controls were available for SNP, and 705 for CTLA4 haplotype, based association analyses. Coeliac disease showed weak association with the CTLA4 +1822T (P=0.019) and CT60 G (P=0.047) alleles. Strong association was seen with a common CTLA4 haplotype (P=0.00067, odds ratio 1.41) of frequency 32.7% in coeliac disease and 25.5% in healthy controls. A common CTLA4 haplotype shows strong association with coeliac disease, and contains multiple alleles reported to affect immunological function. Loss of tolerance to dietary antigens in coeliac disease may be mediated in part by heritable variants in co-signalling genes regulating T-cell responses. PMID- 15657620 TI - Patients with familial biparental hydatidiform moles have normal methylation at imprinted genes. AB - In molar tissues from patients with recurrent biparental hydatidiform moles, we could previously demonstrate that differentially methylated regions (DMRs) of four imprinted genes are abnormally methylated on the maternal alleles. It remained unclear if this abnormal methylation originated de novo in the molar tissues or if it is even recognizable in the patient somatic tissues. To address this question, we investigated the DNA methylation of four imprinted genes in total blood from the two sister-patients. Here, we show that both patients retain normal methylation levels at the DMRs of the four genes in blood tissues. For two maternally expressed genes, we could use informative SNPs to follow the inheritance of the abnormally methylated maternal alleles in the molar tissues. We find that the transmitted abnormally methylated maternal alleles to the moles originated from the maternal grandmother and that the same alleles are not methylated in the patients. Our data suggest that the abnormal methylation in familial biparental molar tissues was acquired de novo in the patients'germline as a result of a false reprogramming or during the postzygotic development of the conceptuses that led to moles. PMID- 15657622 TI - Confidentiality and serious harm in genetics. PMID- 15657623 TI - Determination of the 'critical region' for cat-like cry of Cri-du-chat syndrome and analysis of candidate genes by quantitative PCR. AB - Cri-du-chat (CDC, OMIM 123450) is a chromosomal syndrome that results from partial deletions on the short arm of chromosome 5. The clinical features of CDC normally include high-pitched cat-like cry, mental retardation, microcephaly, hypertelorism and epicanthic folds. The cat-like cry is the most prominent clinical characteristic in newborn children and is usually considered as diagnostic for the CDC syndrome. Using a strategy of 'phenotype dissection', the critical region for cat-like cry was mapped to the chromosomal segment 5p15.3 5p15.2 in previous reports. In this study, the distal breakpoints of two interstitial deletions in two clinical distinctive CDC patients are analysed, one with and one without the cat-like cry. Using PCR, the critical region for the cat like cry is mapped to a short 640 kbp region on chromosome 5p. Genome analysis of this critical region reveals a gene-rich sequence containing five known genes, five putative genes and three spliced EST sequences, altogether 71 predicted exons. Three genes, FLJ25076, a homolog to a ubiquitin-conjugating enzyme UBC-E2, FLJ20303, a nucleolar protein NOP2, which may play a role in the regulation of the cell cycle and MGC5309, a protein with similarity to Nut2, a Drosophila transcriptional coactivator, have been characterized and expression profiles determined by quantitative PCR. These results suggest that one candidate gene, FLJ25076, encodes a ubiquitin-conjugated enzyme E2 type, which is locally expressed in thoracic and scalp tissues. The other two genes are expressed uniformly in all tissues tested, which suggest that they are housekeeping genes. PMID- 15657624 TI - Year in review: communicable disease surveillance, 2003. PMID- 15657625 TI - A large outbreak of norovirus gastroenteritis linked to a catering company, New South Wales, October 2003. PMID- 15657627 TI - A review of Salmonella surveillance in New South Wales, 1998-2000. PMID- 15657626 TI - Risk factors for sporadic Salmonella Birkenhead infection in Queensland and northern New South Wales: a case control study. PMID- 15657628 TI - Tobacco and health fact sheet: Car and home smoke-free zone. PMID- 15657629 TI - Communicable diseases report, New South Wales, for July and August 2004. PMID- 15657632 TI - Etanercept (Enbrel) -- an update. AB - Etanercept is a tumor necrosis factor antagonist with anti-inflammatory effects. It is currently approved in the US for psoriasis, psoriatic arthritis, ankylosing spondylitis, rheumatoid arthritis and juvenile rheumatoid arthritis. Clinical trials have shown this agent to have an excellent safety profile and to be well tolerated by both adult and pediatric patients. PMID- 15657633 TI - A review of prednicarbate (Dermatop). AB - Prednicarbate is a nonhalogenated corticosteroid that is used in the treatment of inflammatory skin diseases, for example atopic dermatitis. It has a favorable benefit-risk ratio, with an inflammatory action similar to that of a medium potency corticosteroid, but with a low potential to cause skin atrophy. Thus, prednicarbate may be a consideration in the treatment of atopic dermatitis in children, and other inflammatory disorders in children and adults. PMID- 15657634 TI - Treatment of photoaging with topical aminolevulinic acid and light. AB - Photodynamic therapy (PDT) has been used for several years for the treatment of actinic keratoses and prevention of invasive nonmelanoma cancers. More recently, increasing physician expertise with the topical sensitizers and light sources employed in PDT has led to expanded applications, including its use for improvement of the visible signs of photoaging. Aesthetic treatment of photoaged skin with brief application of topical 5-aminolevulinic acid followed by well tolerated light sources, such as intense pulsed light or pulsed-dye laser, can enhance the effectiveness of nonablative laser treatment without increasing adverse effects or downtime. PMID- 15657635 TI - Anatomical bases of the bypass-flap: study of the thoracodorsal axis. AB - Cutaneous tissue loss in patients with lesions on the arterial axes remains difficult to treat. Currently, combined surgery associating distal bypass and free flap seems to be the technique that yields the best results. The hemodynamic advantages of this technique, recently demonstrated, are the distal resistance and the increase in bypass flow. Nevertheless, it is complex and its indications limited. Two major drawbacks can be noted: The increasing risk of thrombosis due to the multiplication of anastomoses on the same arterial axis and the deterioration in venous autograft. To overcome these inconveniences we propose a new technique that we call bypass-flap (BF): the graft of an anatomical entity comprised of one artery and one flap. This graft secures the cover of tissue loss and the revascularization of the limb. Apart from its combined nature this technique presents three major advantages. The arterial autograft is superior to the venous graft, the gradually decreasing diameter of the artery secures the congruence of the anastaomoses, and the arterial flow of the graft is higher than a simple bypass due to the joint vascularization of the flap. The arterial graft includes the subscapular and the thoracodorsal arteries. The free flap is composed of serratus anterior muscle supplied by branches of the graft. This investigation studied the feasibility of the bypass flap and determined the length and diameter of the arterial graft and its muscular branch. Forty anatomical preparations were performed on 20 cadavers. The dissections were performed after injection of Rhodorsil. The anatomical feasibility of the bypass flap was confirmed in 37 cases. The total length of the arterial graft that preserved an external diameter above 2 mm was measured at 13 cm (8.5-15.5). This includes the subscapular artery and the thoracodorsal artery with its intramuscular part (if external diameter of that part always above 2 mm). The length of the pedicle of the serratus anterior flap was measured at 7.5 cm (3.0 12.5 cm). PMID- 15657637 TI - Femtosecond laser for glaucoma treatment: a study on ablation energy in pig iris. AB - Safe and effective laser ophthalmic surgery requires a fine balance between the efficiency of laser delivered and the degree of collateral side damage. The laser ocular tissue interaction process is reliant on three main variables, namely, wavelength, pulse duration, and deposited energy. A certain amount of energy is needed to achieve ablation, while too much energy can result in unwanted collateral thermal damage. In our work the relationship between energy deposition and ablation effect is studied by an in-vitro experiment using an 800 nm wavelength 150 fs-pulse-duration laser system. This experiment aims to validate the probability of decreasing the supplied energy during glaucoma surgery by femtosecond laser. Our results show that less energy is needed using femtosecond laser than that using a longer pulse laser. PMID- 15657636 TI - Risk factors for mesh erosion after transvaginal surgery using polypropylene (Atrium) or composite polypropylene/polyglactin 910 (Vypro II) mesh. AB - The objective of this study was to identify the risk factors associated with the occurrence of mesh erosion (ME) during the first 6 post-operative months in patients having undergone transvaginal repair of pelvic organ prolapse (POP) with Atrium or Vypro II mesh. We retrospectively reviewed the records of 198 consecutive patients who underwent vaginal reconstructive surgery reinforced either by Atrium or Vypro II mesh between February 1999 and July 2003. Univariable and multivariable logistic regression was performed to assess associations between measured covariates and ME. Fourteen patients [7.1%, 95% confidence interval (CI): 3.9-11.6] developed vaginal ME. We found that, in the Vypro II mesh patients, the surgeon experienced in the technique had less erosions than less experienced surgeons (2.9 vs 15.6%, p=0.02). There was no statistically significant difference between the monofilament polypropylene (Atrium) and the composite polypropylene/polyglactin 910 (Vypro II) mesh (7.2 vs 6.9%, p=0.41) when adjusted for surgeon experience and patient age. There was no association between mesh type (Atrium or Vypro II) and vaginal ME following transvaginal repair of POP. Surgeon experience and patient age were associated with ME. PMID- 15657639 TI - Insulin inhibits AMPA-induced neuronal damage via stimulation of protein kinase B (Akt). AB - We designed a series of experiments to explore the neuroprotective effects of insulin. Insulin significantly inhibited the alpha-amino-3-hydroxy-5 methylisoxazole-4-propionic acid (AMPA)-induced neuronal cell damage as evidenced by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium-bromide (MTT) assay. However, insulin had little affect on the AMPA-induced glial cell damage. To determine whether insulin inhibits AMPA-induced excitotoxicity, we performed grease-gap recording assays using rat brain slices. In these experiments, insulin also significantly inhibited AMPA-induced depolarization. Flow cytometry and DNA fragmentation assays showed that insulin inhibits AMPA-induced apoptosis and DNA fragmentation, respectively. Insulin stimulated protein kinase B (Akt) activity, whereas AMPA pretreatment did not alter the insulin-stimulated Akt activity. On the contrary, insulin blocked induction of SAPK/JNK, which AMPA stimulated. Taken together, these results suggest that insulin exerts neuroprotective effects by inhibiting AMPA-induced excitotoxicity and apoptosis, possibly by activating Akt and blocking SAPK/JNK. PMID- 15657638 TI - Phylogenetic relationships among some Ateles species: the use of chromosomic and molecular characters. AB - As with most platyrrhines, the systematics of Ateles is under discussion. In order to help clarify its systematic, we employed chromosomic and molecular characters to analyze the phylogenetic relationship among some species of the genus Ateles. Chromosomic studies were conducted on 14 atelid specimens: eight Ateles from A. paniscus, A. chamek, A. belzebuth and A. geoffroyi, and six Alouatta caraya. Ateles paniscus showed 2N=32, whereas A. chamek, A. belzebuth and A. geoffroyi presented 2N=34, XX/XY (with a submetacentric X and a variable Y) corroborated by male meiosis. Nucleotide sequence variation at the mitochondrial cytochrome c oxidase subunit II gene (COII) was analyzed in ten New World monkey specimens. Parsimony trees showed consistent phylogenetic relationships using both chromosomic forms and mitochondrial COII gene sequences as characters. Particularly, chromosomic phylogenies showed A. hybridus as a divergent taxon from the remaining group, whereas A. chamek, A. belzebuth and A. marginatus form an unresolved clade with A. geoffroyi as sister group. PMID- 15657640 TI - Botulinum toxin type A in experimental neuropathic pain. AB - A peripheral application of botulinum toxin type A (7 U/kg) has significantly reduced thermal and mechanical hypersensitivity in rats with the partial sciatic nerve transection as a classical model of surgical neuropathy. PMID- 15657641 TI - Amyloid precursor protein (APP) and its derivatives change after cellular energy depletion. An in vitro-study. AB - To study the relationship between the metabolism of amyloid precursor protein (APP) and cellular energy failure, HEK 293 cells stably transfected with betaAPP 695 underwent graded energy failure induced either by i) hypoxia (pO(2) 25 mm Hg), ii) inhibition of the respiratory chain by sodium azide (NaN(3)), or iii) by combined glucose deprivation/hypoxia of different duration and severity. Secreted APP (APPs) and the derivative betaA4 were quantified autoradiographically by immunoprecipitation, and [(35)S] methionine labeling. APP holoprotein (APPh) was determined by Western blot analysis. The concentrations of the energy-rich metabolites ATP, ADP, creatine phosphate (CrP), and adenosine were measured by high performance liquid chromatography. Mild to moderate energy failure after NaN(3) treatment (2h, 4h) and hypoxia (2h, 8h) was characterized by normal ATP concentration but also by a high reduction in CrP. A stress condition indicated by an increased ATP turnover and adenosine increase was obtained. Intracellular APPh increased but its metabolites APPs and betaA4 as measured in the extracellular compartment decreased. These changes may point to a compensatory response of APP but also to a initial disturbance in intracellular APP metabolism. Severe abnormalities in both energy formation and utilization after 8h NaN(3) and hypoxia glucose deprivation were found to be accompanied by a drastic fall in intracellular APPh concentration by at least 50%, paralleled by an accelerating reduction in the extracellular concentrations of both APPs and betaA4.A significant linear correlation between APPh and ATP and between CrP and betaA4 became obvious. The data of the present study indicate that abnormalities in APP metabolism were generated in an energy-dependent manner. The obvious similarities to sporadic Alzheimer s disease are discussed. PMID- 15657643 TI - Event-related potential correlates selectively reflect cognitive dysfunction in schizophrenics. AB - Schizophrenics show event-related potential (ERP) and particularly P3 abnormalities. To study the more detailed relationships between these ERP alterations and cognitive dysfunction we recorded and analyzed ERPs using a particular experimental approach. In 34 schizophrenics and 25 controls ERPs were obtained by a visual Go/Nogo task requiring response inhibition and were decomposed into temporally independent topographical components using Independent Component Analysis (ICA). ICA disentangled different subcomponents of P3. Subcomponent P3b with a parietal maximum amplitude was significantly reduced in the schizophrenics, probably reflecting their attentional deficits. Subcomponent P3ng with a frontal maximum amplitude and enhanced during Nogo condition appeared as an electrophysiological index of response inhibition. A significantly reduced P3ng enhancement, found in schizophrenics, probably reflects their impaired response control. CONCLUSIONS: ICA can successfully identify ERP subcomponents with distinct scalp topographies representing significant differential indices of normal and abnormal cognitive processing. Involvement of frontal brain areas in disturbed executive control in schizophrenics is supported by our ICA findings. PMID- 15657642 TI - Amelioration of the cerebrovascular amyloidosis in a transgenic model of Alzheimer's disease with the neurotrophic compound cerebrolysin. AB - Increased production and reduced clearance of amyloid beta (Abeta) plays a central role in the pathogenesis of Alzheimer's disease (AD). We have recently shown that the neurotrophic peptide mixture Cerebrolysin (Cbl) has the ability of improving synaptic functioning and reducing amyloid deposition in a transgenic (tg) animal model of Alzheimer's disease (AD). Since in AD, potentially toxic Abeta aggregates accumulate not only around neurons but also in the blood vessels, then it is important to investigate whether bioactive compounds such as Cbl might have the capacity to ameliorate the age-related cerebral amyloid angiopathy (CAA) in tg models. To this end, tg mice expressing mutant human amyloid precursor protein (APP) under the Thy1 promoter were treated with Cbl or saline alone starting at 7 or 12 months of age for a total of three months. Neuropathological analysis with an antibody against Abeta showed that Cbl decreased amyloid deposition around the blood vessels in a time dependent manner. These effects were accompanied by a reduction in perivascular microgliosis and astrogliosis and increased expression of markers of vascular fitness such as CD31 and ZO-1. No lymphocytic infiltration was observed associated with Abeta in the vessels. Consistent with these findings, ultrastructural analysis showed that while in tg mice treated with saline alone there was an abundant accumulation of amyloid fibers in the vascular wall accompanied by thickening of the basal membrane and endothelial cell damage, in Cbl-treated mice there was considerable reduction in the subcellular alterations of endothelial and smooth muscle cells with preservation of basal membranes and intercellular junctions. Taken together, these results suggest that Cbl treatment might have beneficial effects in patients with cognitive impairment due to cerebrovascular amyloidosis by reducing Abeta accumulation and promoting the preservation of the cerebrovasculature. PMID- 15657644 TI - CSF homocysteine is not elevated in schizophrenia. AB - Homocysteine is a neurotoxic amino acid originally found to be an independent risk factor for cardiovascular and cerebral vascular disease and more recently suggested to be a risk factor for Alzheimer's disease. Several authors have observed high plasma homocysteine levels among schizophrenia patients. We reported that such high levels characterize young male schizophrenia patients. We now studied two groups of schizophrenia patients (N=41) and controls (N=29) for CSF homocysteine levels. No difference was found for CSF homocysteine levels between schizophrenia patients and controls (p=.041 for Study A and p=.52 for Study B). PMID- 15657645 TI - Association study of the frizzled-3 (FZD3) gene with schizophrenia and mood disorders. AB - Two research groups have recently reported a significant association between schizophrenia and genetic variants of Frizzled-3 (FZD3) gene. We examined a possible association in a Japanese sample of schizophrenia, bipolar disorder, unipolar depression and controls with four single nucleotide polymorphisms (SNPs), tested in previous reports. We failed to find significant association in the four SNPs or haplotype analysis. The FZD3 gene might not play a role in conferring susceptibility to major psychosis in our sample. PMID- 15657646 TI - Relationships between serotonin transporter promoter polymorphism, platelet serotonin transporter binding and clinical phenotype in suicidal and non-suicidal adolescent inpatients. AB - Relationships between the serotonin transporter promoter polymorphism (5-HTTLPR), platelet serotonin transporter (SERT) binding and clinical phenotype were examined in 32 suicidal and 28 non-suicidal Ashkenazi Israeli adolescent psychiatric inpatients. The 5-HTTLPR polymorphism was not associated with transporter binding or with suicidality or other clinical phenotypes. However, in the suicidal group, a significant positive correlation between platelet SERT density and anger scores (n=32, r=.40; p=.027) and a negative correlation between platelet count and trait anxiety (n=32, r=-.42; p=.034) were observed. PMID- 15657647 TI - Practice parameters for the management of anal fissures (revised). PMID- 15657648 TI - Who, why, what, and how? PMID- 15657649 TI - Assessment of operative risk in colorectal cancer surgery: the Cleveland Clinic Foundation colorectal cancer model. AB - INTRODUCTION: Predictive models play a pivotal role in the provision of risk adjusted, operative mortality rates. The purpose of the study was to describe the development of a dedicated prognostic index for quantifying operative risk in colorectal cancer surgery. METHODS: Data were collected from 5,034 consecutive patients undergoing major surgery in a single center from October 1976 to July 2002. Primary end point was 30-day operative mortality. A multilevel Bayesian logistic regression model was developed to adjust for case-mix and accommodate the variability of outcomes between surgeons. The model was internally validated (split-sample) and tested using measures of discrimination, calibration, and subgroup analysis. RESULTS: The patients' median age was 66 (range, 18-98) years. Operative mortality was 2.3 percent with no significant variability between surgeons or through time. Multivariate analysis identified the following independent risk factors: age (odds ratio = 1.5 per 10-year increase), American Society of Anesthesiologists grade (odds ratio for ASA II, III, IV-V vs. I = 2.6, 4.3, 6.8), TNM staging (odds ratio for Stage IV vs. I-III = 2.6), mode of surgery (odds ratio for urgent vs. nonurgent = 2.1) no-cancer resection vs. cancer resection (odds ratio = 4.5), and hematocrit level. The model offered adequate discrimination (area under receiver operator characteristic curve = 0.801) and excellent agreement between observed and model-predicted outcomes over ten major colorectal procedures (P = 0.191). CONCLUSIONS: The colorectal cancer model provided an accurate means of estimating risk for individual patients in the preoperative setting. It has important implications in everyday practice, because it may be used as an adjunct in the process of informed consent and for monitoring surgical performance through time. PMID- 15657650 TI - Preoperative chemoradiotherapy and total mesorectal excision surgery for locally advanced rectal cancer: correlation with rectal cancer regression grade. AB - PURPOSE: Preoperative long-course chemoradiotherapy is recommended for rectal carcinoma when there is concern that surgery alone may not be curative. Downstaging of the tumor can be measured as rectal cancer regression grade (1-3) and may be of importance when estimating the prognosis. The aim of this study was to look at the long-term results of tumor regression in patients receiving long course chemotherapy before surgical resection of rectal cancer. METHODS: We reviewed those patients who received preoperative chemoradiotherapy followed by surgical resection for carcinoma of the mid rectum or distal rectum found to be stage T3/4 between January 1995 and November 1999. Patients received 45 to 50 Gy irradiation in 2-Gy fractions and an infusion of 5-fluorouracil. Surgical specimens were assessed for rectal cancer regression grade. Patients were followed up routinely with clinical examination, computed tomography, and colonoscopy. RESULTS: Sixty-five patients with a mean age 65 (range, 32-83) years underwent chemoradiotherapy before surgical resection. Thirty patients (46 percent) were classified as rectal cancer regression Grade 1, with 9 patients (14 percent) having complete sterilization of the tumor. Fifty-three patients (82 percent) underwent a curative resection. Overall survival, with a median follow up of 39 (range, 24-83) months, was 67 percent and was associated with tumor downstaging. The local recurrence rate was 5.8 percent in those patients who underwent a curative resection and was significantly lower with rectal cancer regression Grade 1 tumors (P = 0.03). Eight of nine patients (89 percent) whose tumor had been sterilized were alive and well with no recurrence of tumor at a median follow-up of 41 (range, 24-70) months. CONCLUSIONS: Preoperative chemoradiotherapy resulted in significant regression of tumor. Overall survival was high and was associated with downstaging of tumor. The local recurrence rate was significantly lower with rectal cancer regression Grade 1 tumors and was not seen in patients with sterilized tumors. PMID- 15657652 TI - Outcome of salvage abdominoperineal resection after failed endocavitary radiation in patients with rectal cancer. AB - PURPOSE: Endocavitary radiation is a treatment option for selected patients with rectal cancer, but concern exists for the effectiveness of salvage abdominoperineal resection. This study was designed to examine outcomes after salvage abdominoperineal resection for recurrence after endocavitary radiation. METHODS: A prospective database was used to identify patients undergoing abdominoperineal resection after endocavitary radiation from 1985 to 2001. Office records and a tumor registry were used for disease status and survival data. Survival was calculated using the Kaplan-Meier method and groups compared using the Mantel-Haenszel test. RESULTS: Thirty-eight patients underwent salvage abdominoperineal resection. The mean time to recurrence after completion of endocavitary radiation was 21 +/- 27 months, with 29 percent having persistent disease, 63 percent recurrent disease, and 8 percent a second primary. At abdominoperineal resection, 47 percent had tumor transection, specimen perforation, or injury to the genitourinary or gynecologic tract. Nine patients (24 percent) had positive radial margins. The mean time to perineal wound healing was 56 +/- 74.1 days postoperatively, with 36.8 percent taking more than 60 days. Seventeen patients (45 percent) re-recurred at a mean of 21 +/- 25 months after salvage, with a local control rate of 26 percent at 45 +/- 37 months of follow up. Median disease-specific survival from completion of endocavitary radiation was 115.5 months, with a five-year, disease-specific survival rate of 66 percent. Patients with recurrent disease after endocavitary radiation had significantly (P = 0.025) better disease-specific survival than those with persistent disease (median survival 115 vs. 25 months). CONCLUSIONS: Although technically difficult and associated with a high morbidity, abdominoperineal resection can salvage a significant fraction (55 percent) of patients failing endocavitary radiation. A high index of suspicion for recurrence and a tenacious approach to its diagnosis are essential for optimal outcomes. PMID- 15657651 TI - The efficacy of a nerve stimulator (CaverMap) to enhance autonomic nerve identification and confirm nerve preservation during total mesorectal excision. AB - PURPOSE: Sexual dysfunction after total mesorectal excision may be caused by injury to the autonomic nerves. During surgery, nerve identification is not always achieved, and, to date, there has been no method to objectively confirm nerve preservation. The aim of this study was to assess the efficacy of a nerve stimulating device (CaverMap) to assist in the intraoperative identification of the autonomic nerves during total mesorectal excision, and objectively confirm nerve preservation after proctectomy is completed. PATIENTS AND METHODS: Sexually active consecutive male patients undergoing total mesorectal excision were prospectively enrolled in this study. During pelvic dissection, the surgeon attempted to localize the hypogastric and cavernous nerves. CaverMap was used to confirm these findings and to facilitate the identification in cases of uncertainty. At the completion of proctectomy, the nerves were restimulated to ensure preservation. Factors that could affect the surgeon's ability to localize the nerves and CaverMap to confirm this were evaluated. RESULTS: Twenty-nine male patients with a median age of 58 years were enrolled in this study. An attempt to visualize the hypogastric nerves during dissection was made in 26 patients; the surgeon was able to identify the nerves in 19 (73 percent) patients. CaverMap successfully identified the nerves in six of the seven remaining patients, and failed to identify them in only one case. An attempt to localize the cavernous nerves during dissection was made in 13 patients, of which localization was successful in 8 (61.5 percent) patients. CaverMap improved the identification rate in four of the remaining five patients. After proctectomy, CaverMap successfully confirmed the preservation of both hypogastric and cavernous nerves in 27 of 29 (93 percent) patients. A history of previous surgery statistically correlated with failure to identify the hypogastric nerves by the surgeon (P = 0.005). There were no adverse events related to use of the device. CONCLUSION: CaverMap may be a useful tool to facilitate identification of the pelvic autonomic nerves during total mesorectal excision and to objectively confirm nerve preservation. PMID- 15657654 TI - Comparison of multivisceral resection and standard operation for locally advanced colorectal cancer: analysis of prognostic factors for short-term and long-term outcome. AB - PURPOSE: The aim of the present study is to clarify the characteristics of multivisceral resection and to discuss strategies for improving the overall outcome of multivisceral resection for locally advanced colorectal cancer. METHODS: The study included 323 patients who electively underwent curative surgery for pT3-pT4 colorectal carcinoma without distant metastasis. We evaluated the short-term and long-term outcome of multivisceral resection relative to that of the standard operation by means of multivariate analysis of the prognostic factors. RESULTS: Of 323 patients, 53 (16.4 percent) received multivisceral resection because of adhesion to other organs. Multivisceral resection was significantly associated with tumor size, depth of invasion, operative blood loss, operation time, and blood transfusion (all: P < 0.0001). Overall morbidity rates were 49.1 percent after multivisceral resection vs. 17.8 percent after the standard operation (P < 0.0001), and postoperative mortality rate was 0 percent in both groups (not significant). Only multivisceral resection (odds ratio, 2.725; 95 percent confidence interval, 1.125-6.623; P = 0.0264) was an independent factor for overall postoperative complications. The survival rate of patients after multivisceral resection was similar to that after the standard operation (5-year rate, 76.6 percent vs. 79.5 percent, P = 0.9347). Lymph node metastasis (hazard ratio, 2.510; 95 percent confidence interval, 1.460-4.315; P = 0.0009) and blood transfusion (hazard ratio, 2.353; 95 percent confidence interval, 1.185-4.651; P = 0.0145) were independently associated with patient survival. CONCLUSIONS: For locally advanced colorectal cancer, the long-term outcome after multivisceral resection is comparable to that after the standard operation. However, it should be recognized that multivisceral resection is associated with higher postoperative morbidity. In addition, a reduction in the incidence of blood transfusion may contribute to improving patient survival. PMID- 15657653 TI - Total pelvic exenteration with distal sacrectomy for fixed recurrent rectal cancer in the pelvis. AB - PURPOSE: This study evaluates the effectiveness of total pelvic exenteration with distal sacrectomy for fixed recurrent tumor that developed from primary rectal cancer. METHODS: We investigated surgical indications, techniques to minimize blood loss and reduce complications, and oncological outcomes in 57 patients who underwent total pelvic exenteration with distal sacrectomy between 1983 and 2001. RESULTS: Forty-eight patients (84 percent) had negative margins. A comparison between two periods (1983-1992 and 1993-2001) showed that mean blood loss decreased from 4,229 to 2,500 ml (P = 0.002), indicating a favorable learning curve in minimizing blood loss. Two hospital deaths were observed in the earlier period and none in the later period. The most common sacral amputation level was the S3 superior margin, followed by the S4 inferior margin and the S2 inferior margin. The most frequent complication was sacral wound dehiscence in 51 percent, followed by pelvic sepsis in 39 percent. The incidence of pelvic sepsis in the later period was significantly decreased to 23 percent, compared with 72 percent in the earlier period (P = 0.046). Multivariate analysis showed that negative margins and negative carcinoembryonic antigen predicted improved survival. In 48 patients with negative margins, three-year and five-year disease-specific survival rates were 62 percent and 42 percent, respectively. CONCLUSION: Strict patient selection makes total pelvic exenteration with distal sacrectomy a feasible radical approach for fixed recurrent tumor. Careful performance of this surgical procedure along with the proper steps to decrease blood loss should achieve a favorable learning curve and low rate of surgical complications. PMID- 15657655 TI - Are survival rates different for young and older patients with rectal cancer? AB - PURPOSE: Although it is generally believed that young patients with rectal cancer have worse survival rates, no comprehensive analysis has been reported. This study uses a national-level, population-based cancer registry to compare rectal cancer outcomes between young vs. older populations. METHODS: All patients with rectal carcinoma in the Surveillance, Epidemiology, and End Results cancer database from 1991 to 1999 were evaluated. Young (range, 20-40 years; n = 466) and older groups (range, 60-80 years; n = 11,312) were compared for patient and tumor characteristics, treatment patterns, and five-year overall and stage specific survival. Cox multivariate regression analysis was performed to identify predictors of survival. RESULTS: Mean ages for the groups were 34.1 and 70 years. The young group was comprised of more black and Hispanic patients compared with the older group (P < 0.001). Young patients were more likely to present with late stage disease (young vs. older: Stage III, 27 vs. 20 percent respectively, P < 0.001; Stage IV, 17.4 vs. 13.6 percent respectively, P < 0.02). The younger group also had worse grade tumors (poorly differentiated 24.3 vs. 14 percent respectively, P < 0.001). Although the majority of both groups received surgery (85 percent for each), significantly more young patients received radiation (P < 0.001). Importantly, overall and stage-specific, five-year survival rates were similar for both groups (P = not significant). CONCLUSIONS: Although previous studies have found young rectal cancer patients to have poorer survival compared with older patients, this population-based study shows that young rectal cancer patients seem to have equivalent overall and stage-specific survival. PMID- 15657656 TI - Review of survival curves for colorectal cancer. AB - PURPOSE: Actuarial and Product Limited (i.e., Kaplan-Meier) estimates of survival are commonly used in the literature to describe outcomes in patients treated for cancer. Terms such as cancer-specific and cancer-free survival are frequently quoted, although often without clear definitions. This study was designed to compare survival estimates using the Kaplan-Meier method on the same population of patients but using different definitions of what constitutes an event. This was to highlight some of the variation that can occur when different techniques are used to perform these calculations. METHODS: Data were obtained from a prospective database that had recorded all patients presenting with colorectal cancer from 1996 to 2002. Using this information, we calculated the 1) overall (all-cause mortality), 2) cancer-specific, 3) cancer-free, 4) recurrence-free, and 5) relative survival (and 95 percent confidence intervals) at five years postpresentation. RESULTS: The study included 497 patients with a mean age of 68 years, and a male-to-female ratio of 1.3:1. They were followed for a mean of 2.2 years (standard deviation, +/-1.1), with 50 patients (10.1 percent) followed for more than five years. The various survivals at five years were: 1) overall survival, 55.6 percent (95 percent confidence interval, 49.1-62.1 percent), 2) cancer-specific survival, 67 percent (95 percent confidence interval, 60.9-73.1 percent), 3) cancer-free survival, 49.9 percent (95 percent confidence interval, 43.6-56.2 percent), 4) recurrence-free survival, 43.5 percent (95 percent confidence interval, 37.2-49.8 percent), and 5) relative survival, 73.4 percent (95 percent confidence interval, 65.4-81.4 percent). CONCLUSIONS: The five-year survival calculations for this group of patients with colorectal cancer varied by as much as 30 percent depending on how the data was censored. This highlights that there needs to be a clear and accountable definition on how survival curves are calculated and presented in the literature to allow for meaningful interpretation and comparisons. PMID- 15657657 TI - Free colorectal cancer cells on the peritoneal surface: correlation with pathologic variables and survival. AB - BACKGROUND: Clinicopathologic staging of colorectal cancer remains the best predictor of survival. Prognostication for an individual with colorectal cancer remains elusive. This study was designed to investigate the incidence of free surface colorectal cancer cells detected by cytology during elective open curative resection, to correlate their presence with particular clinicopathologic variables and determine whether their presence was predictive of cancer-specific survival. METHODS: Over a six-year period in one institution, all elective colon and intraperitoneal rectal cancer specimens were assessed during primary resection for the presence of free colorectal cancer cells by means of a simple and tested specimen imprint cytology methodology. Clinicopathologic variables were assessed prospectively and blinded to cytology results. All patients were followed up routinely until death and if the patient was not seen within the last six months, information was obtained from the New South Wales Registry of Births, Deaths and Marriages in Australia. RESULTS: Overall, 26 of 281 (9.25 percent) colorectal cancers had positive cytology for cancer cells on the peritoneal surface of the bowel. Poorly differentiated tumors were significantly associated with positive cytology. Tumor penetration, presence of vascular or neural invasion, mucinous characteristics, lymph node status, and operative procedure performed were not statistically significant predictors of positive cytology. Overall, 43 of the 281 patients (15.3 percent) died during the mean follow-up period of 49.2 months from cancer-related deaths. Of these patients, 8 had positive cytology and 35 had negative cytology results. Cancer-specific survival assessed with the log-rank test was significantly associated with positive cytology in univariate (P = 0.008) and multivariate analysis (P < 0.001). CONCLUSION: In this study, the presence of free surface colorectal cancer cells has been shown to be predictive of survival and is independent of direct peritoneal invasion and lymph node status. Thus, further assessment of this simple prognostic variable is warranted and selection of patients with positive cytology for possible adjuvant therapies may be beneficial. PMID- 15657658 TI - Determination of the peritoneal reflection using intraoperative proctoscopy. AB - PURPOSE: Rectal carcinomas are amenable to transanal excision in 3 to 5 percent of cases. Location below the peritoneal reflection is one requirement for transanal excision and transanal endoscopic microsurgery. The location of the peritoneal reflection has not been extensively studied in living patients. METHODS: This study investigated the location of the peritoneal reflection in 50 patients undergoing laparotomy. The distance from the anal verge to the peritoneal reflection was measured in each patient via simultaneous intraoperative proctoscopy and intra-abdominal visualization of the peritoneal reflection. The mean distance to the peritoneal reflection, range of measurements, and complications of proctoscopy were recorded. RESULTS: Intraoperative proctoscopy was performed on 50 patients after informed consent. The mean lengths of the peritoneal reflection were 9 cm anteriorly, 12.2 cm laterally, and 14.8 cm posteriorly for females, and 9.7 cm anteriorly, 12.8 cm laterally, and 15.5 cm posteriorly for males. The lengths of the anterior, lateral, and posterior peritoneal measurements were statistically different from one another, regardless of gender (P < 0.01). There were no complications of proctoscopy. CONCLUSIONS: Our data indicated that the peritoneal reflection is located higher on the rectum than reported in autopsy studies, and that there is no difference between males and females. Knowledge of the location and position of a rectal carcinoma in relationship to the peritoneal reflection will help the surgeon optimize the use of transanal techniques of resection. PMID- 15657659 TI - Prognostic implications of hMLH1 and p53 immunohistochemical status in right sided colon cancer. AB - PURPOSE: Extensive research into the molecular biology of colorectal cancer has identified a plethora of molecular markers reputed to provide independent prognostic information. p53 mutational status has been associated with both improved and reduced survival; however, tumors expressing a particular phenotype associated with defective mismatch repair consistently do better. This study was designed to examine site-specific survival implications of p53 and mismatch repair status. METHODS: Mismatch repair (hMLH1 and hMSH2) and p53 status was investigated immunohistochemically in 111 proximal colon cancers along with tumor TNM stage, grade, and extramural vascular invasion. Fisher's exact test was used to assess categoric data; univariate and multivariate models compared survival between the respective tumor phenotypes. RESULTS: Thirty-two percent of tumors showed loss of expression of hMLH1 and in a multivariate analysis were associated with a significant survival advantage after adjustment for tumor stage, p53 status, and extramural vascular invasion (hazard ratio, 0.29; 95 percent confidence interval, 0.1-0.87; P = 0.027). Only two tumors showed loss of expression of hMSH2, which was not related further to survival. Aberrant p53 expression was detected in 39 percent of tumors. Such expression was found to be associated with a significantly reduced survival in univariate analysis (P = 0.037, log-rank test) but not in a multivariate model. Subgroup analysis showed no association between survival and p53 expression in mismatch repair proficient tumors. CONCLUSIONS: Loss of hMLH1 expression is an independent predicator of improved survival in this series and perhaps the underlying cause of the observed survival difference associated with p53 expression. PMID- 15657660 TI - Specific enhanced expression of platelet-derived endothelial cell growth factor in submucosa of human colorectal cancer. AB - PURPOSE: Platelet-derived endothelial cell growth factor, identified to be an angiogenic factor, has been implicated in metastases of colorectal cancer. This study aimed to clarify the role and localization of platelet-derived endothelial cell growth factor associated with human colorectal cancer invasion. METHODS: Thirty-two patients with colorectal cancer who had undergone surgery were analyzed. Platelet-derived endothelial cell growth factor enzyme activities in the colorectal cancer specimens were measured. Cells that expressed platelet derived endothelial cell growth factor were identified and localized by immunohistochemical analysis with anti-human platelet-derived endothelial cell growth factor antibody and by in situ hybridization with specific RNA probe. RESULTS: Platelet-derived endothelial cell growth factor enzyme activity increased significantly in cancer tissues compared with normal colonic mucosa at various distances from the cancer. Immunohistochemical analysis and in situ hybridization demonstrated platelet-derived endothelial cell growth factor expression in stromal macrophages and fibroblasts located in cancer tissues and surrounding noncancerous tissues, although the tumor cells and normal colonic mucosa were negative. The value of platelet-derived endothelial cell growth factor expression was highest at the border of the colorectal cancer (35.3 +/- 8.9 percent), followed by the cancer nest (15.2 +/- 9.2 percent) and normal mucosa (7.7 +/- 3.4 percent). In the border area, the highest value of platelet derived endothelial cell growth factor expression was observed in the submucosa (35.3 +/- 8.9 percent), followed by the muscular propria (21.9 +/- 7.7 percent) and the subserosa (14.9 +/- 5.5 percent). CONCLUSIONS: Stromal macrophages and fibroblasts are responsible for elevated platelet-derived endothelial cell growth factor activity in colorectal cancer. The significance of enhanced expression of platelet-derived endothelial cell growth factor in the submucosa at the cancer border remains unclear. Cancer stroma may be an important factor for cancer angiogenesis and may serve as a treatment target through specific modulation of angiogenic factors. PMID- 15657662 TI - Liposome-mediated adenomatous polyposis coli gene therapy: a novel anti-adenoma strategy in multiple intestinal neoplasia mouse model. AB - PURPOSE: Familial adenomatous polyposis is a highly penetrant, autosomal dominant disease resulting from a germline mutation of the adenomatous polyposis coli gene. Besides colorectal polyps and cancer, more than 90 percent of familial adenomatous polyposis patients also develop duodenal polyposis with an approximately 5 percent lifetime risk of malignant transformation. Because adenomatous polyposis coli protein has a "gatekeeper role" in the adenoma carcinoma sequence, replacing its function may reduce polyp formation. We studied the functional outcome of per-oral, liposome-mediated adenomatous polyposis coli gene replacement therapy in a multiple intestinal neoplasia mouse model. METHODS: Twenty multiple intestinal neoplasia mice, heterozygous for the human homologue adenomatous polyposis coli gene, were randomly assigned to three groups: no treatment (n = 8); control plasmid containing green fluorescence protein reporter gene (n = 6); and plasmid containing the full-length adenomatous polyposis coli gene (n = 6). For the adenomatous polyposis coli-treated and green fluorescence protein reporter gene-treated groups, each mouse received the appropriate plasmid complexed with liposome, administered twice per week by oral gavage regime. Treatment lasted four weeks and all animals were killed at the end of treatment period with harvesting of intestinal tissue for polyp number estimation. RESULTS: There was a statistically significant 25 percent reduction in the total number of polyps in the adenomatous polyposis coli-treated (73.1 +/- 1.4) group compared with untreated control (97.8 +/- 5.3, P < 0.01, Tukey test) and multiple intestinal neoplasia mice treated with control green fluorescence protein gene (103.3 +/- 1.7, P < 0.01, Tukey test). CONCLUSION: Adenomatous polyposis coli gene dysfunction underlies tumorigenesis in familial adenomatous polyposis patients and multiple intestinal neoplasia mice. This in vivo study provides evidence to support a novel anti-adenoma strategy using enteral adenomatous polyposis coli gene replacement therapy. PMID- 15657661 TI - Hyperplastic polyposis and the risk of colorectal cancer. AB - PURPOSE: Hyperplastic polyps are usually considered to be an innocent finding with little or no potential to progress to colorectal cancer. However, recent literature suggests that some of these polyps may be morphologically and genetically distinct and lead to microsatellite unstable colorectal cancers. The purpose of this study was to define the cancer risk associated with hyperplastic polyposis. METHODS: All patients with hyperplastic polyposis diagnosed by a single colorectal surgeon at a university hospital were followed prospectively. The diagnosis of hyperplastic polyposis was made by the presence of more than 20 hyperplastic polyps distributed throughout the colon and/or a hyperplastic polyp at least 1 cm in size in diameter in the right colon. Patient demographics, family history, size, location, and distribution of polyps and the development of colorectal cancer were noted. RESULTS: Thirteen patients who met the criteria for hyperplastic polyposis were identified and followed prospectively. All of these patients had at least 30 polyps distributed throughout the colon, often > 100. Nine of 13 also had a hyperplastic polyp at least 1 cm in size, usually in the right colon. Of particular note, 7 of 13 patients (54 percent) were diagnosed with colorectal cancer during the study period. Four had cancer on initial diagnosis and three patients developed cancer despite frequent colonoscopic surveillance. Five of seven colorectal cancers were located in the right colon. CONCLUSIONS: Patients with hyperplastic polyposis are at high risk for colorectal cancer. Failure to identify this subset of patients could have dire consequences. PMID- 15657663 TI - Distilled water peritoneal lavage after colorectal cancer surgery. AB - PURPOSE: Tumor cells exfoliated into the peritoneal cavity during colorectal cancer surgery are viable and tumorigenic and may contribute to peritoneal recurrence. Although commonly used, the tumoricidal effectiveness of antiseptics in peritoneal lavage is doubted because of their chemical alteration by peritoneal secretions. In contrast, osmotic lysis by incubation in distilled water may offer an effective tumoricidal activity. Data defining the susceptibility of colorectal carcinoma cells to osmotic lysis are lacking and hence there is no consensus on optimal lavage methodology. METHODS: We examined the cytocidal activity of water on colorectal cancer cell lines in culture and determined the effect of peritoneal secretions in vivo on the tumoricidal effectiveness of water. RESULTS: Incubation of cells in distilled water resulted in cell lysis, with 100 percent lysis achieved after 14 minutes of incubation. In vivo, contamination of lavage water by peritoneal secretions produced a resultant solution with an osmolality of 50 mM. Sequential lavages reduced this contamination, enabling a final resultant solution with an osmolality of 10 mM, which produced 100 percent cell lysis after 32 minutes of incubation. CONCLUSIONS: Current peritoneal lavage methodology is inadequate because complete cell lysis requires water incubation for longer time periods than is currently practiced. Solutions to this problem are discussed. PMID- 15657664 TI - Azathioprine or ileocolic resection for steroid-dependent terminal ileal Crohn's disease? A Markov analysis. AB - INTRODUCTION: The objective of this study was to determine whether initial azathioprine therapy, followed by ileocolic resection if azathioprine fails, or initial ileocolic resection without a trial of azathioprine is the preferred treatment strategy in steroid-dependent, terminal ileal Crohn's disease. METHODS: A Markov, decision analytic model was developed to simulate a 36-month course for a patient with steroid-dependent, terminal ileal Crohn's disease who would initially take azathioprine or have ileocolic resection. Clinically important outcomes in the model included side effects and effectiveness of azathioprine and postoperative complications, mortality, and recurrence following ileocolic resection. The probabilities and utilities for these variables were derived from previously published studies. RESULTS: Initial azathioprine therapy offered a relatively small benefit of 0.45 quality-adjusted life-months over initial ileocolic resection. The model was sensitive to utility for being symptom-free on azathioprine and utility for being symptom-free postoperatively. CONCLUSIONS: Initial azathioprine therapy and initial ileocolic resection are both reasonable treatment strategies in this setting. The preferred treatment strategy is highly dependent on the quality of life that can be achieved with each treatment option. Therefore, individual response and symptom control with each treatment must be strongly considered in this treatment decision. PMID- 15657665 TI - Health-related quality of life and pouch function in continent ileostomy patients: a 30-year perspective. AB - PURPOSE: The principal aim of this study was to assess long-term pouch durability and health-related quality of life in an original series of patients operated on with a continent ileostomy. PATIENTS: Data from 68 of 88 patients who had a continent ileostomy performed at Sahlgrenska University Hospital between 1967 and 1974 were analyzed. Median age at follow-up was 60 (range, 40-89) years and median follow-up was 31 (range, 29-36) years. METHODS: Patients were sent a questionnaire on pouch function along with the Short Form-36 Health Survey 2.0. A random age-matched and gender-matched sample was drawn from the Swedish national Short Form-36 Health Survey norm database to compare with the patient group. RESULTS: The majority of the patients reported good physical condition and satisfactory pouch function. Patients evacuated the pouch a median of four times every 24 hours. Twelve patients (18 percent) had leakages. Forty-four patients (65 percent) had had at least one postoperative revision to restore continence. Generally minor peristomal skin irritation occurred in seven patients (10 percent). Patients with concurrent complaints (mostly age related) reported poorer health-related quality of life. Nevertheless, 78 percent of the patients rated their overall health as good, very good, or excellent. The patients' Short Form-36 Health Survey scores were comparable to reference values. CONCLUSIONS: Although revisional operations may be needed to restore continence, continent ileostomy has a good durability. Pouch function was satisfactory and patients' satisfaction was high. Health-related quality of life levels were similar to those of the general population. PMID- 15657666 TI - Injectable silicone biomaterial for fecal incontinence caused by internal anal sphincter dysfunction is effective. AB - PURPOSE: Fecal incontinence caused by a weak or disrupted internal anal sphincter is common but there has been no effective treatment. This prospective study evaluates the medium-term clinical effects of an injectable silicone biomaterial, PTP (Bioplastique), used to augment the internal anal sphincter. METHOD: Eighty two patients (64 females; median age, 66 years) with severe fecal incontinence and a low anal resting pressure caused by internal anal sphincter dysfunction (defect, n = 11; intact, n = 71) were randomized to PTP injection into intersphincteric space and internal anal sphincter with (Group A, n = 42) or without (Group B, n = 40) guidance by endoanal ultrasound. Both groups were similar in terms of age, gender, past anorectal surgery, duration of follow-up (median, 6 months; range, 1-12 months), and baseline continence score. Sixty-two percent of Group A and 55 percent of Group B had prolonged pudendal nerve terminal motor latency. RESULTS: There was no significant complication. Two patients in Group A and four patients in Group B noted minor discomfort at injection sites. At one month postprocedure, endoanal ultrasound confirmed retention of silicone biomaterial without migration. In both groups, fecal continence was significantly improved by PTP implants 1 month after injection, but continued to improve significantly for up to 12 months in Group A and 6 months in Group B (P < 0.001). Improvement in fecal continence and maximum anal resting pressure was significantly greater in Group A, in whom injection was guided by endoanal ultrasound, than in Group B. At three months after injection, significantly more Group A patients than Group B patients achieved >50 percent improvement in Wexner's continence score (69 percent vs. 40 percent; P = 0.014). Ninety-three percent of Group A and 92 percent of Group B had >50 percent improvement in global quality of life scores (visual analog scale). At a median follow-up of 6 months, all domains of the fecal incontinence quality of life scale improved significantly in both groups; however, the physical function and mental health scores of Short Form-12 only improved in Group A. A prolonged pudendal nerve terminal motor latency had no effect on functional outcome in either group. CONCLUSION: Injection of silicone biomaterial provided a marked improvement in fecal continence and quality of life in patients with internal sphincter dysfunction, despite the presence of pudendal neuropathy. PMID- 15657667 TI - The physical properties of rectal contents have effects on anorectal continence: insights from a study into the cause of fecal spotting on orlistat. AB - PURPOSE: The intermittent loss of oil or stool ("spotting") is an adverse effect that occurs in patients taking orlistat; the pathophysiology is unknown. This study was designed to investigate the local effects of orlistat, free fatty acids, and the effects of the physical properties of rectal contents on anorectal function and continence. METHODS: Anorectal physiology and continence function were assessed in ten healthy patients after the application of four test enemas: 1) high-viscosity stool substitute, 2) stool substitute with free fatty acid, 3) low-viscosity oil with placebo, 4) oil with orlistat. Rectal function and capacity were assessed by barostat techniques. Anal resting pressure, squeeze pressure, and squeeze duration were assessed by manometry. A retention test was performed using the same enemas as a quantitative assessment of continence. RESULTS: Orlistat and free fatty acid had no adverse effects on anorectal function or continence. For each enema, the maximum volume retained correlated with rectal capacity (r = 0.85; P < 0.01). Continence during rectal filling was better maintained for high-viscosity stool substitute than low-viscosity oil enemas (P < 0.03). Patients able to maintain effective squeeze pressure retained more of the low-viscosity enemas than those with short squeeze duration (P < 0.01); in contrast, the volume retained of high-viscosity enemas was unaffected by anal sphincter function. CONCLUSIONS: The physical properties of rectal contents, rectal capacity, and voluntary anal sphincter function have effects on continence function in healthy patients. The occurrence of spotting may depend on both intrinsic anorectal function and the effects of orlistat on the volume and physical properties of stool. PMID- 15657668 TI - Prevention of peritoneal adhesions by intraperitoneal administration of vitamin E: an experimental study in rats. AB - PURPOSE: Previous studies have shown dietary supplements of vitamin E to reduce the incidence of postoperative peritoneal adhesions. The objective of this study was to show the effect of intramuscular or intraperitoneal administration of vitamin E on peritoneal adhesions. METHODS: Eighty rats were divided into four groups: Group A (control), Group B (intramuscular vitamin E), Group C (intraperitoneal olive oil, the vehicle/diluent of vitamin E), and Group D (intraperitoneal vitamin E diluted in olive oil). The same experimental method was used in all rats to produce adhesions, consisting of cecal abrasion and ligature of the adjacent parietal peritoneum. The rats were killed at 14 days to assess the adhesions occurring. The results were analyzed using a chi-squared test. RESULTS: All animals in Groups A, B, and C had substantial adhesions. In Group D, 11 rats had insubstantial adhesions and only 4 had substantial adhesions. There were no significant differences between Groups A, B, and C in terms of percent formation of adhesions. A significant difference was found between Group D (vitamin E plus olive oil by the intraperitoneal route) and each of the experimental groups, A, B, and C (P < 0.0005). CONCLUSIONS: Our results show that intraperitoneal administration of vitamin E just before closing the laparotomy was effective for reducing adhesion formation. By contrast, the same effect was not achieved after intramuscular administration. PMID- 15657669 TI - Robotic and laparoscopic surgery for treatment of colorectal diseases. AB - PURPOSE: In the last ten years, several robotic systems have been developed to overcome the loss of the three-dimensional view and dexterity characteristic of laparoscopic surgery. The aim of this study was to compare the traditional laparoscopic approach and robotic techniques in the treatment of colorectal diseases. METHODS: The study compares a consecutive series of patients treated surgically for colorectal disease from June 2001 to May 2003 with the da Vinci robotic system (Intuitive Surgical) and a matched number of patients who underwent conventional laparoscopy during the same time interval. The factors analyzed were the time required to prepare the patient and the room, total time of surgery, length of specimens, number of lymph nodes retrieved, blood loss, complications, and postoperative results. RESULTS: The study included 106 patients (53 in each group). No differences were observed in total time of surgery (laparoscopic group, 222 +/- 77 minutes vs. robotic group, 240 +/- 61 minutes), specimen length (laparoscopic group, 29 +/- 11 cm vs. robotic group, 27 +/- 13 cm), or number of lymph nodes retrieved (laparoscopic group, 16 +/- 9 vs. robotic group, 17 +/- 10). It took significantly longer to prepare the operating room and patient in the robotic group (24 +/- 12 minutes) than in the laparoscopic group (18 +/- 7 minutes). There were three conversions to laparotomy in the laparoscopic group; in the robotic group, two cases were converted to laparoscopy and three to hand-assisted laparoscopy. No significant differences were observed between the two groups in terms of recovery of bowel function and postoperative hospital stay. CONCLUSIONS: Robot-assisted surgery proved to be as safe and effective as laparoscopic techniques in the treatment of colorectal diseases. Because of its dexterity and three-dimensional view, the da Vinci system was particularly useful in specific stages of the procedure, e.g., takedown of the splenic flexure, dissection of a narrow pelvis, identification of nervous plexus, and handsewn anastomosis. The cost-effectiveness of the procedure still needs to be evaluated. PMID- 15657670 TI - A ten-year study of penetrating injuries of the colon. AB - BACKGROUND: Colon injury has been associated with a high risk of septic complications and mortality. We prospectively studied the pattern, management, outcome, and prognostic factors in patients who sustained penetrating colon injuries. PATIENTS AND METHODS: Sixty patients who presented to our hospital with penetrating colon injuries over a ten-year period (1992 to 2001) were studied. RESULTS: Colon wounds were caused by gunshots in 55 (91.7 percent) patients and knife stabs in 5 (8.3 percent). There was a delay of more than 12 hours before laparotomy in 30 (50 percent) patients. Moderate or major fecal contamination of the peritoneal cavity occurred in 58 (96.7 percent) patients. The average penetrating abdominal trauma index score was 25.9 and 20 (33.3 percent) patients sustained Flint Grade 3 colon injury. Associated intra-abdominal injuries occurred in the small bowel (73.3 percent), liver (25 percent), stomach (23.3 percent), and mesentery (16.7 percent). Right colon wounds (35) were managed by primary repair in 24 (68.6 percent) patients and proximal diverting colostomy in 11 (31.4 percent), whereas left colon wounds (25) were managed by diverting colostomy in 22 (88.0 percent) patients and primary repair in 3 (12.0 percent) patients. Common complications included wound infection (56.7 percent), septicemia (31.7 percent), and enterocutaneous fistula (16.7 percent). The overall mortality rate was 33.3 percent and colon injury-related mortality was 21.7 percent. Presence of destructive colon injury was associated with a greater than fourfold increased incidence of death. Other significant risk factors included shock on admission, major fecal contamination, duration of operation more than four hours, penetrating abdominal trauma index score >25, and more than two postoperative complications. There was no difference in outcome between patients who had primary repair and those undergoing diverting colostomy. Colostomy closure-related morbidity was 21 percent and mortality was 5.3 percent. CONCLUSION: A more liberal use of primary repair is required in our patients with penetrating injuries of the colon. PMID- 15657671 TI - Anocutaneous V-Y advancement flap for the treatment of complex perianal fistula. AB - PURPOSE: The treatment of intersphincteric and low transsphincteric fistula is well defined, but controversy remains around the management of complex perianal fistula. This study was designed to assess the utility of anocutaneous flap repair in complex types of perianal fistula. METHODS: Sixty-five perianal fistula in 65 patients treated with anocutaneous advancement flap for the complex fistula, between April 1998 and December 2002, are included this prospective study. Mean age was 34 +/- 2.1 (range, 24-53) years. Magnetic resonance imaging was used for the diagnosis of fistula. Excision of the internal opening and the overlying anoderm, curettage of the fistula tract, closure of internal opening with absorbable polyglactin 3/0 suture, and drainage of the external opening(s) by insertion of penrose drain were common operational steps. Outcome was evaluated in terms of healing and incontinence. RESULTS: Successful healing of 59 of 65 complex fistulas was achieved using this technique with no disturbance of continence and minimal complications. Mean follow-up and complete healing time were 32 +/- 0.6 (range, 12-52) months and 5.4 +/- 0.8 (range, 3-7) weeks respectively. CONCLUSIONS: Although the study cases were relatively small in number, this report showed that clinical results of anocutaneous advancement flap are acceptable. However, large studies are needed to reach an ultimate conclusion for assessing the place of anocutaneous flap advancement in complex fistula. PMID- 15657672 TI - Improvement of colonic healing by preoperative rectal irrigation with short-chain fatty acids in rats given radiotherapy. AB - BACKGROUND: We investigated the effect of preoperative rectal irrigation with short-chain fatty acids on irradiated colonic anastomosis in rats. METHODS: Sixty male Wistar rats were divided into four groups. Group I (control group, n = 15) underwent left colon resection and primary anastomosis. Group II (Short-chain fatty acids pretreatment group, n = 15) had short-chain fatty acids rectal irrigation for five days preoperatively. Group III (preoperative radiotherapy group, n = 15) underwent irradiation to the whole pelvis eight and four days before the operation, for a total dose of 20 Gy. Group IV (preoperative radiotherapy group + short-chain fatty acids pretreatment group, n = 15) had rectal irrigation with short-chain fatty acids for five days after the second irradiation. Within each group, animals were anesthetized to assess the clinical, mechanical, histologic, and biochemical parameters of anastomotic healing on either the third or seventh postoperative days. RESULTS: The mean bursting pressure was significantly low in Group III on Day 3 and was significantly high in Group IV on Day 7 (P = 0.001, P = 0.021). The burst occurred at the anastomoses in all animals tested on the third postoperative day, and outside of the anastomoses in all animals tested on the seventh postoperative day. The histologic parameters of anastomotic healing, such as epithelial regeneration and formation of granulation tissue, were significantly improved by use of preoperative rectal irrigation with short-chain fatty acids on Day 7. The amount of total and salt-soluble collagen concentrations significantly increased in Group IV compared with the control group on Day 3 (P = 0.008, P = 0.004). CONCLUSION: Some mechanical and histologic aspects of colonic anastomotic healing can be adversely affected by preoperative radiotherapy, but rectal irrigation with short-chain fatty acids may improve anastomotic healing. PMID- 15657673 TI - Quality assessment and improvement in colon and rectal surgery. PMID- 15657674 TI - Angiosarcoma of the colon and rectum: report of a case and review of the literature. AB - Angiosarcoma is a malignancy that occurs rarely in the gastrointestinal tract. We present a case of a 77-year-old male who had rectal bleeding and obstructive bowel symptoms. A large near-obstructing mass was seen endoscopically, but biopsies were inconclusive. A CT scan showed a large sigmoid lesion, and the patient had surgical resection. A large hemorrhagic-appearing tumor was found at operation. Angiosarcoma of the sigmoid colon was diagnosed on histologic examination. After colonic resection, the patient rapidly developed numerous liver metastases and died six months later. A review of the literature reveals only 12 other reported cases of colorectal angiosarcoma; 62 percent of these patients died within one year of surgical resection. The role of adjuvant radiation and/or chemotherapy is unclear. In conclusion, colorectal angiosarcomas are rare tumors that behave very aggressively, and the outcome is generally poor. PMID- 15657675 TI - Rectal duplication cyst presenting as perianal sepsis: report of two cases and review of the literature. AB - INTRODUCTION: Recurrent perianal sepsis is a difficult problem to manage in colorectal surgical practice. One cause is rectal duplication cyst, a rare congenital lesion that is easily overlooked. Many cases have associated congenital defects, especially musculoskeletal anomalies, and may provide a clue to the underlying condition. Early diagnosis is important because these cysts do not resolve spontaneously and may undergo malignant change. METHODS: We present two cases of middle-aged females who presented with perianal sepsis secondary to rectal duplication cyst. The first case had numerous surgical procedures for a perianal fistula during a ten-year period. She had associated sacral anomalies consistent with Currarino syndrome. The second case presented with a perineal mass after a bout of perianal inflammation. Both cases had the entire cyst surgically excised. RESULTS: There were no complications postoperatively and no recurrence at follow-up. Histopathology revealed no malignancy in the cyst. CONCLUSIONS: Rectal duplication cyst is a rare cause of recurrent perianal sepsis that should be considered in difficult cases, especially in those with associated musculoskeletal anomalies. Complete surgical excision is the preferred treatment to prevent recurrence and the risk of malignant degeneration. PMID- 15657676 TI - Retroperitoneal perforation of the colon caused by colonic tuberculosis: report of a case. AB - We present a 25-year-old, HIV-negative patient from Kosovo, with no significant past medical history, who was admitted to a local hospital for nonspecific upper abdominal discomfort. He was transferred to us after a retroperitoneal mass with contact to the right colonic flexure had been found during workup. Colonoscopy demonstrated an edemateous area with a central fistula in the right flexure, and histology showed caseous necrosis. Although neither bacteriology nor histology could detect any germs, gastrointestinal tuberculosis seemed to be very probable. Laparotomy with a segmental resection of the colon was performed to remove the fistula-bearing segment, and histologic examination of the resected specimen confirmed the intraoperative suspect of a retroperitoneal colonic perforation. Again, all cultures from the specimen were negative for tuberculosis, but polymerase chain reaction of a regional lymph node revealed acid-fast bacilli of the Mycobacterium tuberculosis/bovis species. Although the patient had no other sites of tuberculosis infection like pulmonary or urinary, he received adjuvant standard tuberculosis treatment for six months. At control examination one year after the operation, the patient was free of recurrence and in very good general condition. We report this extremely rare presentation of gastrointestinal tuberculosis to sensitize physicians to tuberculosis again, because incidence rates are increasing and this disease will certainly play a more important role in the future. PMID- 15657677 TI - German artificial sphincter system: first report of a novel and highly integrated sphincter prosthesis for therapy of major fecal incontinence. PMID- 15657678 TI - Pharmacologic treatments maintain an important role in the treatment of anal fissure. PMID- 15657682 TI - [Dermal chondroid melanoma]. AB - This case describes the history of disease of a 53-year old man, who has been treated for a putative recurrent nailbed granuloma of the right big toe since 1996. In 2000 an enlarged inguinal lymph node was excised. The light microscopic examination showed a metastasis of a malignant melanoma. In 2003 we received a tumor of the right big toe for histopathological examination. The histological and immunohistochemical results proved a dermal chondroid melanoma. This extremely rare variant of malignant melanoma occurs particularly in subungual location and is possibly related to a previous trauma. We discuss the spectrum of differential diagnoses and the importance of immunohistochemistry. PMID- 15657681 TI - [Leiomyosarcoma of the inferior vena cava. Review of a rare disease]. AB - Primary sarcomas of the great vessels are rare. Their most common site is the inferior vena cava. We report a primary leiomyosarcoma of a 41 year old female patient localised just below the right renal vein. The resected tumour showed the histological signs of a well differentiated, progesterone positive leiomyosarcoma. The case is discussed on the data of the reviewed literature. PMID- 15657683 TI - [Perivascular neoplasms of skin and soft tissues. A review]. AB - Perivascular neoplasms of skin and soft tissues comprise a group of benign and malignant mesenchymal neoplasms showing a perivascular myogenic differentiation. Whereas glomus tumour including its variants represents a clearly defined clinicopathological entity, the whole concept of haemangiopericytoma has been questioned in the last years, and no clear diagnostic criteria are established at the moment. Myopericytomas and myofibromas represent a morphological spectrum of perivascular spindle-shaped lesions. Infantile myofibroma/infantile myofibromatosis and solitary myofibroma of adults are characterized by a biphasic growth of mature, spindled, myofibroblastic cells and smaller, immature, mesenchymal cells associated with numerous haemangiopericytoma-like vessels. Benign and rare malignant myopericytomas of skin and soft tissues are characterized by a concentric, perivascular growth of plump spindled and round tumour cells staining positively for muscle actin, alpha-smooth muscle actin and more rarely for desmin. PEComas represent an enigmatic family of neoplasms composed of perivascular epithelioid, clear, and spindled tumour cells characterized by a coexpression of melanocytic and myogenic markers. PMID- 15657684 TI - [Desmoid-type fibromatosis (aggressive fibromatosis)]. AB - Desmoid-type fibromatoses (aggressive fibromatoses) represent infiltrative, locally destructively growing soft tissue tumors with a high potential for recurrence. Desmoid tumors of 33 adult patients were analysed regarding clinical and morphological aspects (sex, age distribution, site, size, mitotic rate, tumor microvessel density, surgical margins, additional radiotherapy). Possible statistical correlations were examined using log-rank-tests. No prognostic significance of tumor microvessel density was evident. A correlation between mitotic index (1 or more mitoses per 50 high power fields) and local relapse rate was notably striking, but not statistically significant (log-rank: 0.17). Additional postoperatively applied radiotherapy proved to be statistically significant to avoid local recurrences (log-rank: 0.01). The presented results may indicate an increased risk for local relapse in those desmoid-type fibromatoses which are mitotically active. Postoperative radiotherapy seems to be effective in the treatment of aggressive fibromatosis to avoid tumor recurrence. Differential diagnosis of desmoid-type fibromatosis/aggressive fibromatosis in adulthood include various fibroblastic/myofibroblastic soft tissue tumors such as nodular fasciitis, fibrosarcoma, low-grade fibromyxoid sarcoma, myofibroblastic sarcoma as well as leiomyosarcoma and soft tissue leiomyoma. PMID- 15657685 TI - [Tenosynovial giant cell tumor]. AB - Morphological, ultrastructural, and immunohistochemical findings of 12 diffuse type-tenosynovial giant cell tumors/pigmented villonodular synovitis are presented compared to 30 localized tenosynovial giant cell tumors (giant cell tumor of tendon sheath). Diffuse-type-tenosynovial giant cell tumor is characterized by a striking vascularisation pattern composed of densely arranged thin-walled, partly slit-like and partly hyalinized small blood vessels within the papillary synovial fronds. These vessels may show abnormal structures with incompletely arranged endothelial cells/pericytes. The fibrohistiocytic tumor cells probably cause considerable compression/distortion or destruction of the small vessels which might be responsible for an increased blood deposition and massive hemosiderosis. Accompanying multinucleated osteoclast-like giant cells seemingly are recruited from circulating blood monocytes. Microhemorrhagic foci with multinucleated giant cells could be detected in 83% of diffuse-type and 67% of localized-type tumors. Apart from the described vessels, typical morphological findings in diffuse-type tenosynovial giant cell tumors included "giant" hemosiderotic granules, (at least 2-3 times the diameter of an erythrocyte) "giant" siderophages, pseudoalveolar clefts and irregularly anastomosing synovial fronds. Neither mitotic rate nor the amount of giant cells/amount of nuclei of giant cells revealed statistically significant differences between localized-type and diffuse-type of tenosynovial giant cell tumor. Immunohistochemically, the diffuse-type exhibited focal expression of CD31 (in 75% of tumors) and calretinin (in 63%) besides CD68-staining. PMID- 15657686 TI - [Low-malignant peripheral nerve sheath tumors of nasal and sinonasal mucous membranes]. AB - Sinonasal malignant peripheral nerve sheath tumors (MPNST) are infrequent neoplasms. 16 cases of low-malignant MPNST in this localization were retrieved from the files of soft tissue tumors established in Jena. They were examined with regard to their morphology and immunohistochemistry. The importance of an only partial immunostaining by S100 protein antibodies for diagnosis and differential diagnostic discrimination to benign peripheral nerve sheath tumors (schwannomas and neurofibromas) is explained. Finally, the differential diagnosis of spindle cell tumors in the sinonasal region in general is briefly discussed. PMID- 15657687 TI - Radiotherapy, morphine, and bone pain: a difficult relationship? PMID- 15657689 TI - What are cancer patients' preferences about treatment at the end of life, and who should start talking about it? A comparison with healthy people and medical staff. AB - GOALS OF THE WORK: In order to strengthen cancer patients' autonomy and to improve quality of palliative care, it is necessary to know what are the patients' preferences for treatment at the end of life, whether they accept the idea of advance directives, and who should initiate the process of fulfilling such a document. PATIENTS AND METHODS: We compared cancer patients' preferences with respect to particular treatment options at the end of life, acceptance of the idea of advance directives, and preferences for whom should initiate writing such a document with that of healthy controls, nursing staff, and physicians (n=100 each group) using a structured questionnaire. RESULTS: Cancer patients wanted treatment with antibiotics and infringing treatments such as chemotherapy and dialysis significantly more often than healthy controls, nursing staff, and physicians (p<0.01 and p<0.001, respectively). Determinants associated with the wish to opt for these treatments were reduced health condition and older age. The groups did not differ with respect to their acceptance of advance directives; 58 75% of all those surveyed wanted their physicians to initiate a discussion about writing such a document if they thought it appropriate. CONCLUSIONS: Cancer patients' preferences for treatment at the end of life significantly differ compared to other groups. Oncologists should initiate a discussion about an advance directive when/if the course of the illness seems to make this appropriate, which corresponds to the wish of the majority of cancer patients, healthy controls, and medical staff. PMID- 15657688 TI - Problem-solving and distress in prostate cancer patients and their spousal caregivers. AB - GOALS OF WORK: Prostate cancer, the most common life-threatening cancer among American men, increases risk of psychosocial distress and negatively impacts quality of life for both patients and their spouses. To date, most studies have examined the relationship between patient coping and distress; however, it is also likely that what the spouse does to cope, and ultimately how the spouse adjusts, will affect the patient's adjustment and quality of life. The present study examined the relationships of spouse problem-solving coping, distress levels and patient distress in the context of prostate cancer. The following mediational model was tested: Spouses' problem-solving coping will be significantly inversely related to patients' levels of distress, but this relationship will be mediated by spouses' distress levels. PATIENTS AND METHODS: One hundred seventy-one patients with prostate cancer and their spousal caregivers were assessed for mood; spouses were assessed for problem-solving coping skills. Structural equation modeling was used to test model fit. MAIN RESULTS: The model tested was a good fit to the data. Dysfunctional spousal problem-solving was a significant predictor of spouse distress level but constructive problem-solving was not. Spouse distress was significantly related to patient distress. Spouse dysfunctional problem-solving predicted patient distress, but this relationship was mediated by spouse distress. The same mediational relationship did not hold true for constructive problem-solving. CONCLUSIONS: Spouse distress mediates the relationship between spouse dysfunctional coping and patient distress. Problem-solving interventions and supportive care for spouses of men with prostate cancer may impact not only spouses but the patients as well. PMID- 15657690 TI - Procalcitonin--a marker of invasive fungal infection? AB - Procalcitonin (PCT) has been described as a marker of bacterial sepsis. However, little is known of its diagnostic value in fungal infections. We calculated the sensitivity of PCT for detection of invasive fungal infections (IFI) by analyzing 55 episodes of proven or probable IFI (three in our series, 52 reported in the recent literature). In the early phase of IFI, PCT was elevated in fewer than half of invasive candidiasis episodes and in only one patient (5.3%) with invasive aspergillosis. Due to low sensitivity and specificity, PCT adds little to the diagnosis of IFI. PMID- 15657692 TI - [Exceptional conjunctival tumor in a young allergic woman]. PMID- 15657691 TI - [Best's disease. Overview of pathology and its causes]. AB - Best's disease is a hereditary affection with reduced penetrance and juvenile onset. The fundus may be unremarkable or present various stages up to scarring of the macula. Histopathology of advanced stages discloses deposits of lipofuscin on Bruch's membrane, the innermost layer of which is the basal membrane of the retinal pigment epithelium (RPE). The deposits correlate with lipofuscin and melanofuscin granulae in the RPE. Photoreceptors correlating with the lesions have lost their outer segments and the RPE as well as the photoreceptors appear edematous. Loss of photoreceptor function does not necessarily follow progression of fundus appearance. Loss of function usually correlates with a reduced Arden ratio of the electro-oculogram even in the absence of funduscopic changes.Best's disease is caused by mutations in VMD2 (hBEST1). Bestrophin, the gene product of hBEST1, is a regulatory part of a Ca(2+) channel or a Ca(2+)-dependent Cl( )channel. In this paper the relevant data on clinical and genetic pathology are summarized and evaluated. PMID- 15657693 TI - [Ranking of systemic steroids after normal-risk keratoplasty. Results of a randomized prospective study]. AB - BACKGROUND: The aim of this study was to evaluate the efficacy of sole application of topical steroids after normal-risk keratoplasty. PATIENTS AND METHODS: This randomized prospective clinical study assessed 40 patients who had undergone penetrating normal-risk keratoplasty. Twenty patients were treated exclusively with prednisolone acetate 1% eye drops 5x/day for 6 months postoperatively. Another 20 patients additionally received systemic fluocortolone 1 mg/kg body weight per day tapered within 3 weeks postoperatively. The main outcome measures included clear graft survival, ratio of graft rejection, and side effects. RESULTS: The mean postoperative follow-up was 18+/-9 months. Three graft rejections were observed in the group receiving only topical steroids. Two graft rejections were observed in the group administered combined systemic and topical steroid therapy. None of the patients has developed irreversible graft failure so far. CONCLUSION: Sole topical steroid application seems to be an effective immune prophylaxis in patients undergoing penetrating normal-risk keratoplasty. PMID- 15657695 TI - Transabdominal repair of prolapsed pelvic ileal J-pouch after restorative proctocolectomy. PMID- 15657694 TI - [Best's disease with normal EOG. Case report of familial macular dystrophy]. AB - Best's disease is an autosomal dominant disorder with incomplete penetrance and variable expression. A typical characteristic of Best's disease is a pathological EOG. We describe four members of a family with bilateral, subfoveal vitelliform lesions. The EOG was normal in all cases. Genetic analysis of the oldest son indicated a heterozygotic mutation Ala234Val in the VMD2 gene, so-called bestrophin gene, which is associated with Best's disease. Molecular genetic analysis also found Best's disease with a normal EOG. A normal EOG cannot exclude Best's disease. The family members should receive genetic consultation and if wished analysis of the VMD2 gene. PMID- 15657696 TI - Interactions between interlimb and intralimb coordination during the performance of bimanual multijoint movements. AB - The simultaneous performance of movements involving different effectors gives rise to neural and biomechanical interactions between and within limbs. The present study addressed the role of interlimb and intralimb constraints during the control of bimanual multijoint movements. Thirteen participants performed eight tasks involving the bilateral elbows and wrists under different coordination conditions. With respect to interlimb coordination, coordination patterns referred to the in-phase and anti-phase coordination modes, involving the simultaneous timing of homologous versus non-homologous muscles, respectively. With respect to inter-segmental (intralimb) coordination, the isodirectional mode referred to simultaneous flexions and extensions in the ipsilateral wrist and elbow joints, whereas the non-isodirectional mode involved simultaneous flexion in one joint together with extension in the other joint, or vice versa. The analysis of the data focused upon measures of relative phasing between proximal and distal joints within a limb as well as between the homologous joints of both limbs. With respect to interlimb coordination, findings revealed that adoption of the in-phase mode resulted in a higher quality of interlimb coordination than the anti-phase mode. However, the mode adopted in the distal joints had a larger impact on the quality of interlimb coordination than the mode adopted in the proximal joints. More specifically, in-phase coordination of the distal joints had a positive, and anti-phase coordination a negative, influence on the global coordinative behavior of the system. Minor effects of intralimb coordination modes on interlimb coordination were observed. With respect to intralimb coordination between the ipsilateral elbow and wrist, the isodirectional mode was performed with higher stability than the non isodirectional mode. The mode of interlimb coordination also affected the quality of intralimb coordination, such that generating anti-phase coordination patterns in the distal joints had a negative influence on the accuracy and stability of intralimb coordination. Taken together, the present findings suggest a hierarchical structure whereby interlimb coordination constraints have a stronger impact on the global coordinative behavior of the system than intralimb coordination constraints. Moreover, the global coordinative state of the system is more affected by the coordination between the distal than between the proximal joints. Overall, the findings suggest that the mirror-image symmetry constraint has a powerful influence on bimanual multijoint coordination. PMID- 15657697 TI - Localization of the plane of regard in space. AB - When we fixate an object in space, the rotation centers of the eyes, together with the object, define a plane of regard. People perceive the elevation of objects relative to this plane accurately, irrespective of eye or head orientation (Poljac et al. (2004) Vision Res, in press). Yet, to create a correct representation of objects in space, the orientation of the plane of regard in space is required. Subjects pointed along an eccentric vertical line on a touch screen to the location where their plane of regard intersected the touch screen positioned on their right. The distance of the vertical line to the subject's eyes varied from 10 to 40 cm. Subjects were sitting upright and fixating one of the nine randomly presented directions ranging from 20 degrees left and down to 20 degrees right and up relative to their straight ahead. The eccentricity of fixations relative to the pointing location varied by up to 40 degrees . Subjects underestimated the elevation of their plane of regard (on average by 3.69 cm, SD=1.44 cm), regardless of the fixation direction or pointing distance. However, when the targets were shown on a display mounted in a table, to provide support of the subject's hand throughout the trial, subjects pointed accurately (average error 0.3 cm, SD=0.8 cm). In addition, head tilt 20 degrees to the left or right did not cause any change in accuracy. The bias observed in the first task could be caused by maintained tonus in arm muscles when the arm is raised, that might interfere with the transformation from visual to motor signals needed to perform the pointing movement. We conclude that the plane of regard is correctly localized in space. This may be a good starting point for representing objects in head-centric coordinates. PMID- 15657698 TI - Learning a throwing task is associated with differential changes in the use of motor abundance. AB - This study sought to characterize changes in the synergy of joint motions related to learning a Frisbee throwing task and, in particular, how the use of abundant solutions to joint coordination changed during the course of learning for successful performance. The latter information was helpful in determining the relative importance of different performance-related variables (PVs) to performance improvement. Following a pre-test, the main experiment consisted of six subjects practicing a Frisbee throw to a laterally-placed target for five days, 150 throws per day, followed by a post-test. A subgroup of three subjects continued to practice for an extended period of extensive practice amounting to 1800-2700 additional throws each, followed by a second post-test. Motor abundance was addressed through the uncontrolled manifold approach (UCM), which was used to partition the variance of joint configurations into two components with respect to relevant PVs, one component leading to a consistent value of the PV across repetitions, and a reflection of motor abundance, and a second component resulting in unstable values of the relevant PV. The method was used to test hypotheses about the relative importance of controlling the PVs that have an impact on successful task performance: movement extent, movement direction, hand path velocity, and the hand's orientation to the target. In addition, the amount of self-motion, or apparently extraneous joint motion having no effect on the hand's motion, compared to joint motion that does affect the hand's motion, was determined. After a week of practice, all subjects showed improvement in terms of targeting accuracy. Hand movement variability also decreased with practice and this was associated with a decrease in overall joint configuration variance. This trend continued to a greater extent in the three subjects who participated in extended practice. Although the component of joint configuration variance that was consistent with a stable value of all PVs was typically substantially higher than variance leading to unstable values of those PVs, both components decreased with practice. However, the decrease in joint configuration variance reflecting motor abundance was less than the other variance component only in relation to control of movement direction and the hand's orientation to the target. These results indicate that improvement of throwing performance in this experiment was more related to improved stabilization of movement direction and to the hand's orientation to the target than to movement extent and hand velocity. Nonetheless, the relative values of the two joint variance components were such that the instantaneous value of both hand path velocity and movement extent were stabilized throughout the experiment and showed a consistent compensatory relationship at the time of Frisbee release, despite not changing with practice. Finally, the amount of self-motion increased significantly with practice, possibly reflecting better compensation for perturbations due to the limb's dynamics. The results are consistent with other studies, suggesting the need to reevaluate Bernstein's hypothesis of freeing and freezing DOFs with learning. PMID- 15657699 TI - [Diabetic muscle infarction-an orthopedic disease pattern?]. AB - Diabetic muscle infarction (DMI) is a largely unfamiliar disease. It affects mainly patients around 40 years of age with long-standing diabetes and concomitant end-organ complications. The symptoms represent a classic pattern of a musculoskeletal disease with muscle pain without trauma, swelling, and functional impairment. Although its short-term prognosis is good, with improvement of the symptoms over weeks or months under analgesia and rest, a high recurrence rate of up to 60% can be observed. Additionaly, the long-term survival of patients after DMI is reduced mostly due to major vascular complications. Since many diabetic patients are in orthopedic care for musculoskeletal disorders, the orthopedic surgeon should be aware of this disease to avoid unnecessary invasive diagnostic procedures and initiate suitable therapy. Furthermore, a better knowledge of the disease could lead to definite conclusions regarding its real incidence and aid in establishing new therapeutic measures for prophylaxis and better long-term survival. PMID- 15657700 TI - [Neurological and functional recovery from spinal cord injury. Progress and evaluation standards in paraplegic medicine]. AB - Today, there is accumulating evidence from animal experiments that axonal regeneration and an enhanced level of functional repair can be induced after a spinal cord injury (SCI). Consequently, in the near future, new therapeutic approaches will be developed for the treatment of patients with SCI. The aim of the project presented here is to provide the required clinical basis for the implementation of novel interventional therapies. Refined and combined clinical and neurophysiological measures are needed for a precise qualitative and quantitative assessment of spinal cord function in patients with SCI at an early stage. This represents a basic requirement for recognizing any improvement in the recovery of function and to monitor any significant effect of a new treatment. The paper presents objective and refined tools as a basis for monitoring the effects of new treatment strategies. PMID- 15657701 TI - Polypyrrole modified stainless steel frits for on-line micro solid phase extraction of ochratoxin A. AB - Polypyrrole (PPy) was electrochemically synthesized on stainless steel frits as a sorbent for the micro solid phase extraction (muSPE) of ochratoxin A (OTA). Using 20 microl of standard solution under a fast flow rate of 0.5 ml/min, 80% recovery of OTA was achieved in the concentration range from 0.1-10 pg/mul. This good recovery was achieved within a short residence time of 1.2 s. A binding capacity of 1 ng OTA was estimated for each PPy-modified frit, or 2 ng OTA for two frits in series. The bound OTA could be pulsed eluted (PE) with 20 microl of 1% triethylamine in acetonitrile. On-line coupling of this PPy-on-a-frit and PE technique to high performance liquid chromatography (HPLC) was straightforward. On-line muSPE-PE-HPLC results clearly demonstrated the capability of PPy-on-a frit to bind OTA in the presence of red wine, beer, and orange juice components. PMID- 15657703 TI - Sample preparation for speciation. PMID- 15657702 TI - DNA separation with low-viscosity sieving matrix on microfabricated polycarbonate microfluidic chips. AB - Microfluidic devices have been fabricated on polycarbonate (PC) substrates by use of a hot embossing method using a silicon master template. By adding auxiliary lines around the functional channel on the silicon master, its lifetime was significantly prolonged and the bonding strength of the PC cover plate to the microfluidic chip was greatly improved. More than 300 polycarbonate microfluidic chips have been replicated with the same silicon mold. CE separation of Phi X 174/HaeIII DNA restriction fragments, with high resolution efficiency and good reproducibility, was achieved on these devices using the low-viscosity sieving matrix HPMC-50. Temperature was found to have a significant effect on separation efficiency. PMID- 15657704 TI - A dynamic dilution system-based evaluation of the procedure adopted for determining ozone precursor volatile compounds. AB - A dynamic dilution system was created to evaluate the performance and the reliability of ozone precursor volatile organic compound (VOC) sampling ("TO-Can" canisters) and analysis (thermal desorption/gas chromatography/flame ionisation detection) techniques used by the "Laboratoire Interregional de Chimie du Grand Est (LIC)". Different atmospheres of VOCs were generated at concentrations between 0.8 and 25 ppb, with temperatures of 0, 10, 20 and 30 degrees C, and with relative humidities of 0, 30, 50, 70 and 90%. These conditions are generally representative of those commonly observed in ambient air in the eastern France. This dynamic dilution allows the simulation of a wide range of scenarios (concentrations, temperatures and relative humidities). After assessing the capacity and performance of the system, it was applied in order to evaluate the recoveries and stabilities of VOCs from canisters used for the collection and analysis of two mixtures of VOCs. The first mixture contained six alkanes (ethane, propane, butane, pentane, hexane and heptane), and the second contained five alkenes (ethene, propene, butene, 1-pentene and 1-hexene), five aromatics (benzene, toluene, ethylbenzene, m-xylene and o-xylene), acetylene, and 1,3 butadiene. No significant losses of alkanes from the canisters were observed after 21 days of storage, and good recoveries of alkanes from the canisters (>80%) were obtained regardless of the concentration, the temperature and the relative humidity. However, losses of certain aromatics were noted at low relative humidity. PMID- 15657705 TI - A new determining method of copper(II) ions at ng ml(-1) levels based on quenching of the water-soluble nanocrystals fluorescence. AB - CdTe nanocrystals (NCs) were prepared and modified in aqueous solution by using 3 mercaptopropionic acid (MPA). Compared with previous fluorescent sensors based on organic fluorophores, these NCs have the following merits. First, they are sensitive and the synthetic procedure is simple; second, they have narrow, tunable, symmetric emission spectra and are photochemically stable. The applicability of these NCs to the determination of Cu(II) ions was studied successfully. Maximum fluorescence intensity was produced at pH 7.8, with excitation and emission wavelengths at 365 and 532 nm, respectively. Under optimal conditions, linear relationships were found between the relative fluorescence intensity and the concentration of Cu(II) ions in the range 0-256 ng ml(-1) and the limit of detection was 0.19 ng ml(-1). This method was applied to determine real samples successfully. In addition, the quenching mechanism is also described. PMID- 15657707 TI - Spectrofluorimetric determination of phenyl-beta-naphthylamine used as rubber antioxidant. AB - Phenyl-beta-naphthylamine (PBN) used as rubber antioxidant was found to have native fluorescence. A spectrofluorimetric method for determination of PBN in multicomponent mixtures of polymer additives is described. The apparent excitation and fluorescence wavelengths used are 348 and 413.5 nm, respectively. Maximum fluorescence intensity is obtained by irradiating PBN dissolved in ethanol, at room temperature. The fluorescence varies linearly with the concentration of PBN in the range of 0.04-4 microg mL(-1). The accuracy and precision of the method are reported. PMID- 15657706 TI - Glucose biosensor based on glucose oxidase immobilized in sol-gel chitosan/silica hybrid composite film on Prussian blue modified glass carbon electrode. AB - An improved amperometric glucose biosensor based on glucose oxidase immobilized in sol-gel chitosan/silica hybrid composite film, which was prepared from chitosan (CS) and methyltrimethoxysilane (MTOS), on the surface of Prussian blue (PB)-modified glass carbon electrode was developed. The film was characterized by FT-IR. Effects of some experimental variables such as ratio of CS to silica, buffer pH, temperature, and applied potential on the current response of the biosensor were investigated. The biosensor fabricated under optimal conditions had a linear response to glucose over the range 5.0 x 10(-5) to 2.6 x 10(-2) M with a correlation coefficient of 0.9948 and a detection limit of 8.0 x 10(-6) M based on S/N = 3. The biosensor had a fast response time of less than 10 s, a high sensitivity of 420 nA mM(-1), a long-term stability of over 60 days, and a good selectivity. The apparent Michaelis-Menten constant K(m) was found to be 3.2 x 10(-3) M. The activation energy for enzymatic reaction was calculated to be 21.9 kJ mol(-1). This method has been used to determine the glucose concentration in real human blood samples. PMID- 15657708 TI - Determination of pesticides and PCBs in honey by solid-phase extraction cleanup followed by gas chromatography with electron-capture and nitrogen-phosphorus detection. AB - A multiresidue method for determination of 15 organochlorine pesticides (OCPs), six polychlorinated biphenyls (PCBs), and seven organophosphorus pesticides (OPPs) is implemented for routine determinations of residues in honey. The method involves solid-phase extraction cleanup and determination by GC-ECD/NPD. Quantitation limits ranged from 0.1 to 0.6 microg kg-1 honey for OCPs and PCBs, and from 5.0 to 25.0 microg kg-1 honey for OPPs. Recoveries of OCPs ranged between 77.4 and 94.0%; for PCBs they were from 63.8 to 73.5%. Recovery assays for OPPs varied from 66.7 to 98.1%. The method was applied to the analysis of 111 honey samples from Aragon, Spain. The results obtained indicated a low level of contamination by pesticide residues and PCBs, which can contribute to ensuring the consumer has a safe wholesome supply of honey. PMID- 15657709 TI - Novel potentiometric immunosensor for determination of diphtheria antigen based on compound nanoparticles and bilayer two-dimensional sol-gel as matrices. AB - A novel method for fabrication of a diphtheria potentiometric immunosensor has been developed by means of self-assembling compound nanoparticles to a thiol containing sol-gel network. A cleaned gold electrode was first immersed in a 3 mercaptopropyltrimethoxysilane (MPS) sol-gel solution to assemble a silica sol gel monolayer. The silane entities were then polymerized into a two-dimensional sol-gel network (2D network) by dipping into aqueous NaOH. The second silane layer was formed by re-immersion in the MPS sol-gel solution overnight. The compound nanoparticles (nanocompounds) containing gold nanoparticles and silver nanoparticles were then chemisorbed on to the thiol groups of the second silane layer. Finally, diphtheria antibody (anti-Diph) was adsorbed on to the surface of the compound nanoparticles. The modified process was characterized by cyclic voltammetry (CV). Detection is based on the change in the potentiometric response before and after the antigen-antibody reaction. A direct potentiometric response to diphtheria antigen (Diph) was obtained from the immobilized diphtheria antibody. The potentiometric response of the resulting immunosensor was rapid and the linear range was from 22 to 800 ng mL-1 with the linear regression equation DeltaE=-79.5+69.4 log [Diph] and a detection limit of 3.7 ng mL-1 (at 3delta). Up to 19 successive assay cycles with retention of sensitivity were achieved for probes regenerated with 0.2 mol L-1 glycine-hydrochloric acid (Gly-HCl) buffer solution. Moreover, analytical results from several serum samples obtained using the developed technique were in satisfactory agreement with those given by the ELISA method, implying a promising alternative approach for detecting diphtheria antigen in clinical diagnosis. PMID- 15657710 TI - Determination of selected polycyclic aromatic hydrocarbons and oxygenated polycyclic aromatic hydrocarbons in aerosol samples by high-performance liquid chromatography and liquid chromatography-tandem mass spectrometry. AB - Fine and ultrafine particles are probably responsible for numerous health effects, but it is still unclear whether and to what extent the particle itself or organic compounds adsorbed or condensed on the particle are responsible for the effects observed. One important class of particle-bound substances are the polycyclic aromatic hydrocarbons (PAH) and their oxygenated derivatives. To improve the tools used for chemical characterization of particulate matter analytical methods for the determination of PAH and oxygenated PAH in aerosol samples of different origin have been developed and optimized. PAH on high-volume filters and on soot aerosols were analyzed by using accelerated solvent extraction for extraction and high-performance liquid chromatography with fluorescence detection for separation and quantification. Total PAH concentrations were in the range 0.3-9.3 ng m(-3). For analysis of selected oxygenated PAH on high-volume filters a liquid chromatography-tandem mass spectrometric method was developed and optimized. Preliminary investigations showed that oxygenated PAH at pg m(-3) concentrations can be determined. PMID- 15657712 TI - Electrosynthesis and analytical characterization of PMMA coatings on titanium substrates as barriers against ion release. AB - The performance of polyacrylic coatings as barrier films against corrosion of titanium-based orthopaedic implants was investigated. In particular, poly(methyl methacrylate) (PMMA) was electrosynthesized on titanium substrates by electro reductive processes from aqueous monomer solutions. The obtained PMMA coatings were characterized by Fourier transform infrared spectroscopy (FTIR) and X-ray photoelectron spectroscopy (XPS). The effect of an annealing treatment on the morphology of coatings with respect to uniformity and porosity of films was assessed by scanning electron microscopy (SEM). An inductively coupled plasma mass spectrometry (ICP-MS) technique was used for ion concentration measurements in ion release tests performed on TiAlV sheets modified with PMMA coatings (annealed and unannealed). Results indicated that the annealing process produces coatings with considerable anticorrosion performances. PMID- 15657713 TI - [Introduction to the topic: mild hypothermia. Are we too hopeful?]. PMID- 15657711 TI - Spectroscopic and voltammetric studies of pefloxacin bound to calf thymus double stranded DNA. AB - Spectral and electrochemical studies have been carried out on the interaction of pefloxacin with calf thymus double-stranded dsDNA. The voltammetric behavior of pefloxacin was investigated at glassy carbon, carbon paste and dsDNA-modified carbon paste electrodes using cyclic voltammetry. Pefloxacin was oxidized, yielding one irreversible oxidation peak. The modification of the carbon paste surface with dsDNA allowed an accumulation process to take place for pefloxacin such that higher sensitivity was achieved compared with the bare surface. The response was characterized with respect to ionic strength, accumulation time, pefloxacin concentration, and other variables. The stripping differential pulse voltammetric response showed a linear calibration curve in the range 1.0 x 10(-7) 1.0 x 10(-5) mol l(-1) with a detection limit of 5.0 x 10(-8) mol l(-1) at the dsDNA modified electrode. The method was applied to the direct determination of pefloxacin in diluted urine samples. PMID- 15657714 TI - [Decreased inspiratory time during ventilation of an unprotected airway. Effect on stomach inflation and lung ventilation in a bench model]. AB - BACKGROUND: In an unprotected airway during cardiopulmonary resuscitation, two ventilations with an inspiratory time of 2 s after 15 chest compressions are recommended. Therefore, approximately 30% of the resuscitation attempt is spent on ventilation. Since survival rates did not decrease sharply when minute ventilation levels were relatively low, and uninterrupted chest compressions with a constant rate of approximately 100/min have been shown to be lifesaving, it may be beneficial to decrease the time spent on ventilation and instead, increase the time for chest compressions. METHODS: In an established bench model of a simulated, unprotected airway with increased airway resistance, we evaluated if inspiratory time can be decreased from 2 to 1 s at different lower oesophageal sphincter pressure (LOSP) levels during ventilation with a bag-valve-mask device. RESULTS: An inspiratory time of 2 vs. 1 s resulted in significantly lower peak airway pressure, while lung tidal volume was significantly higher at an inspiratory time of 2 s and a LOSP of 5 cm H(2)O (480+/-20 vs. 380+/-30 ml) and 10 cm H(2)O (630+/-50 vs. 440+/-20 ml) and significantly lower at a LOSP of 15 cm H(2)O (470+/-70 vs. 540+/-20 ml). While neither ventilation strategy produced stomach inflation at 20 cm H(2)O LOSP, 1 vs. 2 s inspiratory time produced significantly higher stomach inflation at 15 cm H(2)O LOSP (8+/-11 vs. 0 ml) and significantly lower stomach inflation at a LOSP of 5 cm H(2)O (359+/-31 vs. 375+/ 29 ml) and 10 cm H(2)O (28+/-13 vs. 36+/-12 ml) per breath. CONCLUSION: In this model of a simulated, unprotected airway, a reduction of inspiratory time from 2 to 1 s resulted in a significant increase of peak airway pressure, while lung tidal volumes and stomach inflation volumes were statistically different but clinically comparable. PMID- 15657715 TI - Transgenic tomato plants expressing an Arabidopsis thionin (Thi2.1) driven by fruit-inactive promoter battle against phytopathogenic attack. AB - Tomato is one of the most important crop plants; however, attacks by pathogens can cause serious losses in production. In this report, we explore the potential of using the Arabidopsis thionin (Thi2.1) gene to genetically engineer enhanced resistance to multiple diseases in tomato. Potential thionin toxicity in fruits was negated by the use of a fruit-inactive promoter to drive the Thi2.1 gene. In transgenic lines containing RB7/Thi2.1, constitutive Thi2.1 expression was detected in roots and incidentally in leaves, but not in fruits. Disease assays revealed that the transgenic lines that were tested conferred significant levels of enhanced resistance to bacterial wilt (BW) and Fusarium wilt (FW). Further studies indicated that BW disease progression in transgenic lines was delayed by a systemic suppression of bacterial multiplication. By adopting a safe genetic engineering strategy, the present investigation is another step forward demonstrating thionin practicality in crop protection. PMID- 15657716 TI - Decreased sucrose-6-phosphate phosphatase level in transgenic tobacco inhibits photosynthesis, alters carbohydrate partitioning, and reduces growth. AB - The aim of this work was to examine the role of sucrose-6-phosphate phosphatase (SPP; EC 3.1.3.24) in photosynthetic carbon partitioning. SPP catalyzes the final step in the pathway of sucrose synthesis; however, until now the importance of this enzyme in plants has not been studied by reversed-genetics approaches. With the intention of conducting such a study, transgenic tobacco plants with reduced SPP levels were produced using an RNA interference (RNAi) strategy. Transformants with less than 10% of wild-type SPP activity displayed a range of phenotypes, including those that showed inhibition of photosynthesis, chlorosis, and reduced growth rates. These plants had strongly reduced levels of sucrose and hexoses but contained 3-5 times more starch than the control specimens. The leaves were unable to export transient starch during extended periods of darkness and as consequence showed a starch- and maltose-excess phenotype. This indicates that no alternative mechanism for carbon export was activated. Inhibition of SPP resulted in an approximately 1,000-fold higher accumulation of sucrose-6-phosphate (Suc6P) compared to wild-type leaves, whereas the content of hexose-phosphates was reduced. Although the massive accumulation of Suc6P in the cytosol of transgenic leaves was assumed to impair phosphate-recycling into the chloroplast, no obvious signs of phosphate-limitation of photosynthesis became apparent. 3 Phosphoglycerate (3-PGA) levels dropped slightly and the ATP/ADP ratio was not reduced in the transgenic lines under investigation. It is proposed that in SPP deficient plants, long-term compensatory responses give rise to the observed acceleration of starch synthesis, increase in total cellular Pi content, decrease in protein content, and related reduction in photosynthetic activity. PMID- 15657717 TI - Relationship between expression of the PM H+-ATPase, growth and ion partitioning in the leaves of salt-treated Medicago species. AB - The role of the plasma membrane (PM) H(+)-ATPase (E.C. 3.6.1.3) in the plant's response to salt stress was studied in the perennial leguminosae forage Medicago arborea L. and its close relative Medicago citrina (Font-Quer) Greuter, a species exposed to saline conditions in its original habitat. Plants were solution cultured for 8 days in 1 or 100 mM NaCl. Leaf growth and CO(2) assimilation were more inhibited by salt in M. arborea than in M. citrina. Both species were able to osmoregulate, and salt-treated plants maintained turgor potentials, with no differences between species. Contrasting ion distribution patterns showed that M. citrina was able to exclude Na(+) from the leaves more selectively, while M. arborea had a greater buildup of leaf blade Na(+). Isolation of purified PM and quantification of H(+)-ATPase protein by Western blot analysis against the 46E5B11D5 or AHA3 antibodies showed an increase in response to salt stress in the expanding (92%) and expanded leaves (87%) of M. citrina, while no differences were found in the corresponding leaves of M. arborea. The assay of H(+)-ATPase specific activity of the two leaf types in salinized M. citrina confirmed this increase, as activities increased with 55% and 104% for the expanded and expanding leaves, respectively, while no significant differences were found for either leaf type of salinized M. arborea. A possible role of the increased expression of the PM H(+)-ATPase for leaf expansion and ion exclusion in salt stressed plants is discussed. PMID- 15657718 TI - [Endoscopic therapy of acute and chronic pancreatitis]. AB - Endoscopic therapy is valuable for both acute and chronic pancreatitis. Early endoscopic papillotomy appears, in the case of a severe course of acute biliary pancreatitis, to be advantageous. Endoscopic drainage can be considered in cases of acute fluid retention and necrosis as well as subacute, non-healing pancreatitis or cyst development. By acute chronic pancreatitis with strictures or bile duct stones, papillotomy, dilation and stent insertion can lead to an improvement in pain symptoms. An improvement in endo- or exocrine function, however, is not expected. Studies on the endoscopic therapy of pancreatitis are still very limited, and recommendations can usually only be made based on retrospective case series. PMID- 15657719 TI - [Upper gastrointestinal bleeding after long term, high dose NSAID medication: a wolf in sheep's clothing?]. AB - A 71-year-old patient with melena, abdominal and lower back pain was admitted to hospital under suspicion of upper gastrointestinal bleeding. He had mild anemia and an elevated C-reactive protein. Endoscopy and ultrasound failed to localize the source of bleeding. The patient died 2 1/2 days after admission on an exsanguinating hemorrhage. On pathologic examination a fistula between the small aortic aneurysm and duodenum and periaortic inflammation in contact with a spondylitis were found. When clinical presentation is subtle, with a herald bleeding followed by a period of grace, the diagnosis of aortoenteric fistula may be extremely difficult. PMID- 15657722 TI - Characterization and potential application of purified aldehyde oxidase from Pseudomonas stutzeri IFO12695. AB - The molecular weight of purified aldehyde oxidase from Pseudomonas stutzeri IFO12695 was estimated to be 160 kDa by a gel filtration method. SDS-PAGE showed that the enzyme consisted of three non-identical subunits with molecular weights of 18, 38, and 83 kDa. The enzyme exhibited an absorption spectrum with maxima at 277, 325, 365, 415, 450, 480, and 550 nm and possessed molybdenum, CMP, iron, sulfur, and FAD as its cofactors, indicating that it belonged to the xanthine oxidase family. A variety of aliphatic and aromatic aldehydes were oxidized; and among them n-hexylaldehyde gave the most rapidly action. When 10 mM formaldehyde was treated with the aldehyde oxidase in the presence of catalase for 240 min, the formaldehyde concentration was reduced to 0.8 mM, suggesting this enzyme might be effective for the removal of formaldehyde contained in wastewater. PMID- 15657721 TI - [Status febrilis and unconsciousness]. AB - Cerebral malaria with Plasmodium vivax is uncommon. Normally Plasmodium falciparum is the cause of cerebral malaria. We report about a 18 year old patient from Pakistan with a history of intermittent fever for several months. The patient recovered within a few days; however prognosis can be severe when cerebral malaria is complicating the course of Plasmodium vivax infection. PMID- 15657723 TI - Haematococcus pluvialis cultivation in split-cylinder internal-loop airlift photobioreactor under aeration conditions avoiding cell damage. AB - The effects of superficial gas velocity in the riser (U(Gr)) and gas entrance velocity (v) on the growth of Haematococcus pluvialis cultivated in a split cylinder internal-loop airlift photobioreactor were investigated. Cell growth decreased when U(Gr) and v were increased above 12 mm s(-1) and 22.8 m s(-1), respectively. The maximum cell density of H. pluvialis was 110 x 10(4) vegetative cells ml(-1) and the chlorophyll-a titer was 7 mg l(-1). The cell damage in the photobioreactor was greater when v was increased by an increase in U(Gr) rather than by a decrease in sparger internal diameter. The overall volumetric mass transfer coefficient (k(L)a) of the photobioreactor was measured at the same U(Gr) (6-24 mm s(-1)) and v (12-80 m s(-1)). The k(L)a values reached in the airlift photobioreactor were between 10 h(-1) and 32 h(-1). PMID- 15657724 TI - Applications of zeolite inorganic composites in biotechnology: current state and perspectives. AB - The purpose of this short review is to introduce applications of inorganic composites, zeolites, in biotechnology. Although inorganic chemistry is generally considered distant from biotechnology, the two could be harmoniously integrated for biopolymer chromatography. New chromatographic carriers have been developed based on principles differing from those underlying conventional chromatography. Some can be used for the purification of proteins according to novel physicochemical principles, according to their isoelectric point (pI), molecular weight and shape. The amount of protein adsorbed is related to the pore size of the composites, which can recognize biomolecules with reference to these three parameters. Proteins adsorbed at their pI have been found to be desorbed at the pI by polyethylene glycol, but not by high ionic medium (NaCl), SDS, non-ionic detergents, ATP or urea. Therefore, inorganic composites synthesized in consideration of pore size and three-dimensional structure are suitable as new chromatographic carriers. Selective fractionation of biomaterials including proteins and nucleic acids should provide useful information regarding whether conjugated proteins in a precipitated state can be separated on net charge and whether cells can be directly fractionated in future. PMID- 15657725 TI - [OP-hospitation program of VOD and DDG]. PMID- 15657727 TI - [The 100 years since discovery of Spirochaeta pallida]. AB - On 3 March 1905, Fritz Schaudinn, Erich Hoffmann and Fred Neufeld, working in the women's ward of the Department of Dermatology at the Charite Hospital in Berlin, became the first people in the world to observe the causative agent of syphilis, Treponema pallidum. The pathogen's etiological significance was subsequently demonstrated by Schaudinn and Hoffmann and other scientists. The detection of Treponema pallidum was the first decisive step towards the development of diagnostic and therapeutic procedures in subsequent years. PMID- 15657726 TI - [Serum markers for melanoma]. AB - Tumor markers in the serum of cancer patients have an important role in clinical diagnosis and in prognosis, and also in the monitoring of the patients' disease and response to therapy over time. The serum markers currently available for melanoma have only limited clinical use. Those most widely used in clinical applications are S100-beta, melanoma inhibitory activity, and lactate dehydrogenase; there are close correlations between the serum concentrations of these and tumor load. Regular determination of S100-beta and MIA levels during follow-up can therefore be used for early detection of a tumor relapse in melanoma patients, increased serum concentrations of these marker proteins being indicative of tumor growth. Patients with distant metastases from melanoma who present with elevated serum levels of S100-beta, MIA, or LDH have poorer overall survival than do patients whose serum concentrations are within normal ranges. These three markers can also be used to monitor the course of disease and therapy outcome in patients with distant metastases. Since there are no marker proteins for melanoma that are not dependent on tumor load, it is not currently possible to forecast the survival of patients who are tumor free after surgery. Serum markers are also not suitable for screening or for the diagnosis of primary melanomas. PMID- 15657728 TI - [Inflammatory lesion with a black scab on the face of a 16-year old girl]. PMID- 15657729 TI - [Rare types of vasculitis as markers of plasmocytoma]. AB - We report on two female patients who presented with painful recurrent palpable purpura, ulcers and necroses on the extremities. The results of all examinations and laboratory tests considered together suggested a diagnosis of necrotizing leukocytoclastic vasculitis. Leukocytoclastic vasculitis is an inflammatory necrotizing condition of the superficial dermal vessels, presenting with variable clinical symptoms. In most cases it becomes manifest as palpable purpura, but hemorrhagic-necrotizing, bullous, nodular and urticarial presentations also occur. Common etiological factors include bacterial, viral or drug antigens, chronic infections (hepatitis B and C), non-Hodgkin lymphomas (monoclonal gammopathy, multiple myeloma), leukemia (hairy cell leukemia), and tumors (bronchial, breast, and gastric cancer) and also connective tissue disorders. In the course of the work-up, a plasmocytoma was discovered as the cause of the leukocytoclastic vasculitis, presenting in a similar way to livedo reticularis in one case and to pyoderma gangraenosum in the other. PMID- 15657730 TI - [Facial pigmentation following therapy with terbinafine]. AB - A 65-year-old patient presented with grey-brownish maculae localized on the face. He had been treated with oral terbinafine due to onychomycosis, and the first spots manifested after 4 weeks of therapy. Other drugs were not taken by the patient, who was otherwise in a healthy condition. Histology showed melanin localized within macrophages in the upper and lower dermis. Cutaneous side effects are well described in patients treated with terbinafine and usually present as urticaria or eczema. Severe reactions may occur in rare cases. Hyperpigmentation has not yet been described as a consequence of oral terbinafine. Grey hyperpigmentation as it occurred in our patient has been described as a side effect of therapy with minocycline, amiodarone, tricyclic antidepressants, or heavy metals. This case report shows that drug-induced hyperpigmentation should also be considered if the patient takes drugs not known for this kind of side effect. PMID- 15657731 TI - [Superficial X-ray therapy of lip cancer after chronic excessive sun exposure of a 47-year-old autistic painter]. AB - A 47-year-old autistic painter developed lip cancer after chronic excessive sun exposure. As she feared sun withdrawal during inpatient surgical excision, she refused this first-line treatment and therefore received outpatient X-ray therapy. The reason why she developed premature lip cancer and refused to stay in a hospital and the way she illustrated her "sun addiction" in her pictures is presented. Two years later she was free of recurrence. She has remained a sun worshipper until now, but after X-ray therapy she used daily sunscreens with very high sun protection factors, which attenuated the actinic damage. Considering the specific personality of the autistic painter, outpatient radiation therapy was a good alternative to inpatient surgery. PMID- 15657732 TI - [Allergic contact urticaria caused by a chameleon. Expression of sensitization to Ficus benjamina]. AB - A 31-year-old man presented with a long history of rhinoconjunctivitis and sneezing that lasted from March to May. The man kept a chameleon (Chamaeleo calyptratus) as a pet and reported about recurrent urticarial lesions, erythema, and itch after skin contact with the animal. The reactions started within a few minutes and were restricted specifically to the contact sites with the animal's claws. No comparable reactions occurred in other subjects. Allergy testing confirmed allergy to hazel, alder, birch, and ash pollen, and additionally revealed sensitization to house dust mite, cat, and Ficus benjamina. Apparently the contact dermatitis originated from passive transfer of Ficus benjamina allergens to the patient's skin by the reptile which habitually climbed on a big Ficus benjamina tree in the patient's home, thereby contaminating its claws with the plant's allergenic milky sap. Careful examination revealed strong perforation of many leaves by the pointed claws. The patient denied respiratory symptoms from Ficus benjamina and intolerance of Ficus-associated fruits. PMID- 15657733 TI - [Multiple subcutaneous painful hyperpigmented nodules on both lower limbs]. PMID- 15657734 TI - [Prevention of atopic eczema. Evidence based guidelines]. AB - With an estimated prevalence of 12% for preschool children and 3% for adults, atopic eczema is a serious public health problem. This disease severely jeopardizes quality of life and is associated with considerable costs. Since there is still no causal therapy, primary and secondary prevention are especially important. Here the evidence basis for recommendations on prevention of atopic eczema is discussed on the basis of the first evidence-based consensus guideline (S3) on allergy prevention. This recommends that babies should be breastfed exclusively for at least 4 months and exposure to passive smoking be avoided even during pregnancy; restriction of the maternal diet during pregnancy has no influence, though during breastfeeding it can lower the incidence of eczema among babies at risk. Thereby this measure should be balanced with potential consequences of malnutrition. There seems to be a positive correlation between keeping small rodents (rabbits, guinea pigs), and possibly cats, and the occurrence of atopic eczema, while keeping dogs has no effect or even a protective effect. Avoidance of an unfavorable indoor climate is probably also helpful in preventing eczema. There is no evidence to support deviating from the current recommendations of the standing committee for vaccination. PMID- 15657735 TI - [Socio-legal evaluation of UV-induced skin tumors]. AB - The present work deals with insurance and legal issues on the prevention of UV induced skin tumors. We are convinced that squamous cell carcinoma of the skin fulfils the socio-legally required conditions according to paragraph 9 Abs. 2 SGB VII for approval as an occupational disease. In malignant melanoma evidence also exists for its induction through UV exposure and increased risk for occupational UV exposure, thus, making approval as an occupational disease possible in individual cases. According to the currently available medical knowledge on basal cell carcinoma and malignant melanoma, there is no sufficient basis for the approval of these as occupational disorders. Therefore, significant actions should not only be taken in the context of primary disease prevention, but also within the framework of secondary and tertiary disease prevention in occupational UV exposure. PMID- 15657736 TI - [Albert Neisser's expeditions to Java in 1905 and 1907. Syphilis research and travel experiences]. AB - Albert Neisser, the noted dermatologist from Breslau, went on study tours to Java in 1905 and 1907 in order to conduct experiments on monkeys to investigate a number of open questions concerning etiology, course and therapy of syphilis. These large-scale research efforts brought many results, which were somewhat overshadowed by more up-to-date investigations of other groups. Neisser considered his main achievement to be new insights into the immunity and therapy of syphilis. PMID- 15657737 TI - Distribution of the stem cells (neoblasts) in the planarian Dugesia japonica. AB - It has been postulated that the high regeneration ability of planarians is supported by totipotent stem cells, called neoblasts. There have been a few reports showing the distribution of neoblasts in planarians. However, the findings were not completely consistent. To determine the distribution of neoblasts, we focused on proliferating cell nuclear antigen (PCNA), which is present in proliferative cells. We cloned and sequenced the cDNA of PCNA from the planarian Dugesia japonica and produced an antiserum recognizing the gene product. X-ray irradiation caused rapid loss of all PCNA-positive cells and loss of the neoblasts (which were morphologically defined by the presence of the chromatoid body), strongly suggesting that all PCNA-positive cells were true neoblasts. Using the antiserum, we were successful in identifying the neoblasts more clearly than any previous work. In addition to their dispersed distribution in the dorsal and ventral mesenchyme, the neoblasts were distributed as clusters along the midline and bilateral lines in the dorsal mesenchyme. We also examined the behavior of the neoblasts after decapitation. Decapitation did not seem to affect the migration of neoblasts far from the wound. We demonstrated here that DjPCNA is a powerful tool for identifying planarian neoblasts. PMID- 15657739 TI - Two classes of 5S rDNA unit arrays of the silver fir, Abies alba Mill.: structure, localization and evolution. AB - The structure and organization of the 5S ribosomal DNA units of the silver fir, Abies alba Mill., as well as their position in the chromosome complement were investigated. PCR amplification of the gene and nontranscribed spacer region, sequence analysis and Southern hybridization, using a homologous probe, detected DNA sequences of approximately 550 bp and 700 bp. Sequence analysis of the spacers revealed that the difference in length between the sequences occurred in the middle spacer region as a result of the amplification of a 75-bp sequence of the short unit class, which is organized in four 54- to 68-bp tandem repeats in the long spacer unit. The 5S rDNA transcribed region is 120 bp long and shows high sequence similarity with other gymnosperm species. The comparative analysis of 5' and 3' flanking sequences of 5S rRNA genes of silver fir and other gymnosperms indicates that A. alba spacer units have the same rate of evolution and are more closely related to Larix and Pseudotsuga than to Pinus and Picea. Southern hybridization and fluorescence in situ hybridization of metaphase chromosomes of A. alba suggest that the short and long spacer units are organized as separate tandem arrays at two chromosomal loci on chromosomes V and XI. PMID- 15657740 TI - Fine mapping of the FT1 locus for soybean flowering time using a residual heterozygous line derived from a recombinant inbred line. AB - Fine-mapping of loci related to complex quantitative traits is essential for map based cloning. A residual heterozygous line (RHL) of soybean (Glycine max) derived from a recombinant inbred line (RIL) was used for fine-mapping FT1, which is a major quantitative trait locus (QTL) responsible for soybean flowering time. The residual heterozygous line RHL1-156 was selected from the RILs that were derived from two distantly related varieties, Misuzudaizu and Moshidou Gong 503. The genome of RHL1-156 contains a heterozygous segment (approximately 17 cM) surrounding the FT1 locus but is homozygous in other regions, including three other loci affecting flowering time. A large segregating population of 1,006 individuals derived by selfing of RHL1-156 included two homozygous genotypes for the nearest marker of FT1 whose flowering time differed by 26 days. No continuous range of phenotypes was observed, in contrast to the F2 population, suggesting that a single FT1 locus affected the flowering time in the RHL1-156 line. Linkage analysis revealed that the FT1 locus mapped as a single Mendelian factor between two tightly linked DNA markers, Satt365 and GM169, at distances of approximately 0.1 cM and 0.4 cM, respectively. Our results show that a RHL derived from RILs can be used to fine-map a QTL and that RHLs can be an efficient tool for a systematic fine-mapping of QTLs. PMID- 15657738 TI - Dynamics of blood chylomicron fatty acids in a marine carnivore: implications for lipid metabolism and quantitative estimation of predator diets. AB - Blubber fatty acid(s) (FA) signatures can provide accurate estimates of predator diets using quantitative FA signature analysis, provided that aspects of predator FA metabolism are taken into account. Because the intestinal absorption of dietary FA and their incorporation into chylomicrons (the primary transport lipoproteins for dietary FA in the blood) may influence the relationship between FA composition in the diet and adipose tissue, we investigated the metabolism of individual FA at these early stages of assimilation. We also investigated the capacity of chylomicron signatures to provide quantitative estimates of prey composition of an experimental meal. Six captive juvenile grey seals (Halichoerus grypus) were fed either 2.3 kg (n = 3) or 4.6 kg (n = 3) of Atlantic herring (Clupea harengus). Although chylomicron FA signatures resembled diet signatures at all samplings, absolute differences were smallest at 3-h post-feeding, when chylomicrons were likely largest and had the greatest ratio of triacylglycerol to phospholipid FA. Specific FA that differed significantly between diet and chylomicron signatures reflected either input from endogenous sources or loss through peroxisomal beta-oxidation. When these aspects of metabolism were accounted for, the quantitative predictions of diet composition generated using chylomicron signatures were extremely accurate, even when tested against 28 other prey items. PMID- 15657741 TI - Development of sequence characterized amplified region (SCAR) primers for the detection of Phyto.5.2, a major QTL for resistance to Phytophthora capsici Leon. in pepper. AB - Phytophthora capsici causes devastating disease on many crop species, including Capsicum. Resistance in Capsicum annuum is genetically and physiologically complex. A panel of Capsicum germplasm that included genotypes from both C. annuum and C. chinense showing highly resistant, highly susceptible and intermediate or tolerant responses to the pathogen, respectively, was screened with a series of randomly amplified polymorphic sequence primers to determine which genomic regions contribute to the highest level of resistance. One primer, OpD04, amplified a single band only in those C. annuum and C. chinense genotypes showing the highest level of resistance. The amplified product was cloned, sequenced and used to design longer primers in order to generate a sequence characterized amplified region marker which was then mapped in a reference mapping population and a screened population segregating for resistance to P. capsici. These primers were observed to define a locus on pepper chromosome 5 tightly linked to Phyto.5.2, one of six quantitative trait loci (QTL) previously reported to contribute to P. capsici resistance. These results indicate that the Phyto.5.2 QTL may be widely distributed in highly resistant germplasm and provide improved resolution for this QTL. This work also defines the first breeding tools for this system, allowing for the rapid selection of genotypes likely to be highly resistant to P. capsici. PMID- 15657742 TI - Genetic and biochemical analysis of common wheat cultivars lacking puroindoline a. AB - Puroindoline a (Pin-a) and puroindoline b (Pin-b), two basic isoforms encoded by the Pina-D1 and Pinb-D1 loci respectively, involved in controlling grain texture in wheat, were isolated from starch granules of soft wheat cultivars using three different extraction procedures, and fractionated by acidic polyacrylamide gel electrophoresis (A-PAGE). Tris buffer containing 1% Triton X-114 extracted Pin-a and small amounts of Pin-b, whereas 1% SDS preferably extracted Pin-b. Large amounts of both puroindolines were isolated by a solution containing 50% propan-2 ol and 50 mM NaCl. This solution extracted reduced amounts of Pin-b and no traces of Pin-a from starch granules of 20 hard common wheats containing the null allele Pina-D1b. The absence of Pin-a was confirmed by immunostaining with an anti-Pin-a antiserum. With the exception of two cultivars, null Pin-a cultivars gave no PCR fragment with three primer pairs specific to either the coding region or the promoter region of Pina-D1a, suggesting that major changes had occurred at the Pina-D1 locus in these genotypes. Cultivars Fortuna and Glenman were unique in giving size-specific PCR fragments with all primer pairs for the allele Pina-D1a and showed a cytosine deletion at position 267 in the coding region of the Pin-a gene, which resulted in a TGA stop codon at position 361. However, there was no evidence of a mutated protein in the A-PAGE or SDS-PAGE patterns of Fortuna and Glenman. The novel gene, provisionally named Pina-D1c, is the first null allele due to a point mutation that has been identified at the Pina-D1 locus. PMID- 15657743 TI - The carrot, not the stick: appetitive rather than aversive gustatory stimuli support associative olfactory learning in individually assayed Drosophila larvae. AB - The ability to learn is universal among animals; we investigate associative learning between odors and "tastants" in larval Drosophila melanogaster. As biologically important gustatory stimuli, like sugars, salts, or bitter substances have many behavioral functions, we investigate not only their reinforcing function, but also their response-modulating and response-releasing function. Concerning the response-releasing function, larvae are attracted by fructose and repelled by sodium chloride and quinine; also, fructose increases, but salt and quinine suppress feeding. However, none of these stimuli has a nonassociative, modulatory effect on olfactory choice behavior. Finally, only fructose but neither salt nor quinine has a reinforcing effect in associative olfactory learning. This implies that the response-releasing, response-modulating and reinforcing functions of these tastants are dissociated on the behavioral level. These results open the door to analyze how this dissociation is brought about on the cellular and molecular level; this should be facilitated by the cellular simplicity and genetic accessibility of the Drosophila larva. PMID- 15657745 TI - HPLC with laser-induced native fluorescence detection for morphine and morphine glucuronides from blood after immunoaffinity extraction. AB - A new immunoaffinity solid phase extraction of morphine and its phase II metabolites, morphine-3-beta-D-glucuronide and morphine-6-beta-D-glucuronide is described. An immunoadsorber was applied which was created for the first time by the immobilisation of specific antibodies (polyclonal, host: rabbit) by the sol gel method. The extraction method in combination with high performance liquid chromatography-fluorescence determination has been validated and shown to be applicable to blood samples of heroin victims in a low concentration range. Blood extracts were essentially free of interfering matrix components when compared to C8-extracts. Additionally, a novel, sensitive and selective detection system for wavelength-resolved analysis of laser-induced fluorescence coupled to HPLC was developed. The analytes were excited with a frequency tripled Ti:Sa laser (lambda=244 nm quasi cw). The total emission spectrum was recorded with a detection system consisting of an imaging spectrograph and a back-illuminated CCD camera. This technique of detection, combined with an extended optical path (at least 6 mm could be illuminated by the laser), resulted in an optimal fluorescence intensity of the analytes. The method permitted the analysis of morphine, morphine-3-beta-D-glucuronide and morphine-6-beta-D-glucuronide in a low concentration range and could be applied to a complex matrix such as postmortem blood samples because analyte peaks could be discriminated from matrix peaks by their characteristic emission spectra. PMID- 15657744 TI - Differential gene expression in yellow-necked mice Apodemus flavicollis (Rodentia, Mammalia) with and without B chromosomes. AB - Most B chromosomes are heavily heterochromatic, promoting the general idea that they are genetically inert. The B chromosomes of Apodemus flavicollis are euchromatic and show a high degree of homology with the A chromosomes. The euchromatic nature of B chromosomes in A. flavicollis suggests that they may carry active genes and have transcriptional activity. We applied the differential display reverse transcription-polymerase chain reaction (DD RT-PCR) in order to analyze and compare gene expression in animals possessing B chromosomes and animals without B chromosomes. After a second and third round of amplification, three cDNA fragments were differentially expressed in +B mice compared with 0B animals. These cDNAs were Chaperonin containing TCP-1, subunit 6b (zeta) (CCT6B), Fragile histidine triad gene (FHIT) and hypothetical gene XP transcript. The differential expression pattern was confirmed by Real Time RT-PCR. We suggest that altered expression of these important genes is due to the presence of B chromosomes. In elevating the expression of these genes, B chromosomes of A. flavicollis affect some of the crucial processes in the cell. The significance of these effects and the nature of B chromosomes of A. flavicollis are discussed in the context of the data presented. PMID- 15657746 TI - Internal carotid artery injury during functional endoscopic sinus surgery and its management. AB - Rupture of the internal carotid artery (ICA) during functional endoscopic sinus surgery (FESS) is a rare complication, which can potentially result in death. Only a few cases have been reported in the literature thus far. We present four cases with an ICA bleeding during sphenoidotomy. The treatment is discussed and an emergency plan to manage the heavy arterial bleeding is presented. PMID- 15657747 TI - [Infectious mononucleosis]. AB - The primary infection with Epstein-Barr virus in an immunocompetent individual leads to infectious mononucleosis with symptoms of diphtheroid angina, lymph node swelling in the neck and hepatosplenomegaly. The most common age of infection lies between 15 and 25 years. The illness can affect a number of organs simultaneously and thus requires interdisciplinary diagnostics. For differential diagnosis, a differential blood analysis and a EBV quick test are required. The presence of IgM antibodies demonstrates the presence of the infection. Ultrasound of the abdomen can be made to determine the involvement of additional organs. In most cases, recovery occurs without complications. Acute cases can usually be handled successfully with medication. If symptomatic treatment fails, pharyngeal airway obstruction is possible and a tonsillectomy may be necessary. Otherwise, surgical treatment is obsolete. Generally, the prognosis is good. Severe courses and complications are rare. PMID- 15657748 TI - [Tinnitus from the point of view of an ENT practitioner]. AB - Tinnitus lasting for more than 3 months is considered chronic. It is not a self contained illness as such, but a symptom of a central information processing malfunction with individually distinguishing emotional reactions. However, it can, as chronically decompensated tinnitus, manifest itself as an ailment which is more or less pronounced in the person afflicted, and it can also, as a subjective sensation, be the symptom of an objective illness which must be excluded by differential diagnosis. In the majority of cases of chronic tinnitus, psychological components play a substantial role and must be taken into account in diagnosis. As a rule, no therapeutic impact can be made on chronic tinnitus. The doctor's target is therefore to achieve and stabilise habituation in the person afflicted so that, although the tinnitus will continue to exist, it will no longer be perceived as burdensome. Amongst the causes or contributory factors of tinnitus are dysfunctions of the upper cervical vertebral column and/or of the mandibular joints. In cases of tinnitus, these should always be diagnostically clarified and, wherever possible, eliminated to make habituation easier and to diminish the likelihood of recidivism. PMID- 15657749 TI - [Quality control of audiologic results in a university phoniatric/pediatric institution. An example of expert assessment of occupational noise induced hearing loss]. AB - AIM: An investigation was made to reveal whether suspicion of occupational hearing loss can be satisfactorily determined by an otolaryngologist or workplace audiological measurement. These were compared with a formal audiometrical assessment at a university clinic. METHODS AND RESULTS: A retrospective study was made of 95 cases of noise induced hearing loss. A total of 78 individuals were investigated by an otolaryngologist and 70 by workplace audiometry. Using workplace audiometry, 27% of the tests showed a reduction in working capacity of at least 20%. In only five of these was a specialist opinion sought within a year. In 50%, this took longer than 5 years. A comparison of audiometric data from expert opinion revealed that there was conformity in only 47% with workplace audiometry and 48% with otolaryngologist testing. In some cases (27% workplace and 33% ENT practice), the measured hearing loss and calculation of disability exceeded that determined by the experts. CONCLUSIONS: The results of workplace audiometry demonstrated that hearing loss was frequently reported only after the workers had received a disability of at least 20%. Possible reasons for discrepancies in audiological testing might be the exaggeration of hearing loss by the worker, insufficient recovery time after noise exposure, or inexperienced audiologists. Our data show that audiometric testing in workplace audiometry, as well as in ENT practice, often reveal a higher disability rating compared to formal audiological (university) assessment, even if these discrepancies do not reach statistical significance. PMID- 15657750 TI - [Hearing impairment in children and adolescents with Down's syndrome]. AB - BACKGROUND: There are controversial opinions about the incidence of hearing impairment of children and adolescents with Down's syndrome as an additional cause for obstruction in social and communicative live. Next to congenital diseases of the inner ear, functional limitations can often lead to conductive hearing disorders in patients with Down's syndrome. PATIENTS AND METHODS: The hearing ability, otoscopic and audiometric characteristics of 115 patients aged 3.2+/-2.9 years were analysed retrospectively over 42 months. RESULTS: A total of 86 children showed morphologic otoscopic peculiarities: transitory evoked otoacoustic emissions (TEOAE) were often non-reproducible, even in patients with normal hearing abilities(14.1% of cases), and were not evaluable in 32.8% of cases. Half of the patients showed hearing loss, 82% of whom suffered from a conductive hearing loss, 6.9% from a combined, and 8.6% from isolated sensorineural hearing loss. Hearing loss was first diagnosed when the children were 4.6+/-3.4 years old. CONCLUSION: The frequent occurrence of hearing loss justifies and requires an early and continuous pedaudiological examination. As a common screening examination, TEOAE is limited due to its cost, however, it fulfils its purpose as a means of distinguishing healthy children from those with possible hearing disorders. In most cases, objective and subjective audiometric methods must be combined in order to precisely define the hearing ability. PMID- 15657751 TI - [Effects of magnetic resonance tomography on upper eyelid implants]. AB - BACKGROUND: For the therapy of lagophthalmos in facial palsy, gold (99.99%), platinum (99.95%) and platinum/iridium (97%/3%) alloy weights are implanted into the upper eyelid. Investigations on the MRI compatibility of the different eyelid implants with 1.5 and 3.0 Tesla-MRT-systems, which are often used for the diagnosis of head and neck tumours, are still missing. METHODS: Three different eyelid implants made of pure gold, pure platinum and a platinum/iridium alloy were tested in vitro. As parameters, temperature changes, movements of the implants and picture artefacts of the different implants were determined using a 1.5 and 3.0 Tesla MR system. The deflection forces of the implants were evaluated by thread test, lying in a Petri dish and floating in a water bath. RESULTS: No rise in temperature was recorded in any implant. On the water surface, all three implants changed their positions and floated towards the magnetic field. The velocity of the implants increased in the 3.0 Tesla system. Increasing the frictional resistance by lying the implants in a Petri dish, there was no movement in the 1.5 or 3.0 Tesla systems. In addition, no movement of the implants could be observed in the thread test. All implants made few artefacts, with gold showing the least artefacts. CONCLUSION: MR imaging at 1.5 or 3.0 Tesla may be performed safely for warming up or dislocation of the implant in patients with eyelid implants made of pure gold, pure platinum or a platinum/iridium alloy. PMID- 15657752 TI - [Spinal manipulative therapy and cervical artery dissections]. AB - Severe complications after cervical spine manipulation are rare. As experts for medical treatment errors, we received between July 2002 and February 2004 cases with serious complications in the central nervous system after manipulation. 5 vertebral artery dissections with subsequent brain infarction were registered. In all cases, the patients showed complete persisting remission of symptoms. In addition, a kinematic estimation model was developed to study the possible causes of vertebral artery damage. We were able to demonstrate that material extension is dependent on cervical rotation and the "free length" of the vertebral artery in the upper cervical spine. PMID- 15657753 TI - [Significance of dendritic cells for the immunotherapy of tumors]. PMID- 15657754 TI - [Double vision and impediments to nasal breathing]. PMID- 15657755 TI - [Cochlear implantation with preservation of residual deep frequency hearing]. AB - BACKGROUND: The aim of the present paper is to evaluate the clinical parameters in patients implanted for combined, ipsilateral electric-acoustic stimulation of the auditory system. METHODS: A total of 18 patients with residual deep frequency hearing were implanted with a Combi 40+cochlear implant (MED-EL, Austria). Insertion depths ranged from 18 to 22 mm (360 degrees ). A modified surgical technique should contribute to hearing preservation in low frequency regions of the cochlea. Pure-tone audiometric thresholds were measured pre- and postoperatively. A speech audiometric evaluation was performed on two subjects. RESULTS: Utilizing adapted surgical procedures, the preservation of low frequency hearing was accomplished in 16 of 18 subjects (88.9%). Seven (38.9%) patients had complete and nine (50.0%) partial preservation of residual hearing. The speech discrimination scores of two patients documented an increase in sentence intelligibility when compared with only the cochlear implant. CONCLUSIONS: Hearing preservation in cochlear implant surgery is possible. Insertions of 360 degrees provide a full functioning cochlear implant to stimulate sufficient neural structures for above average discrimination scores with the implant alone. A synergistic effect of the electric and the acoustic stimulation modes leads to high discrimination scores in background noise. PMID- 15657756 TI - [Sudden hearing loss as the leading symptom of an infarction of the left anterior inferior cerebellar artery]. AB - The sudden onset of hearing impairment or hearing loss can be a characteristic sign of a vertebrobasilar circulatory disturbance. We report on a 65 year old male patient with an acute left-sided tinnitus followed by hearing loss as an initial symptom of an infarction of the left anterior inferior cerebellar artery (AICA). Successively, additional symptoms with vertigo, nausea, vomiting and a transient dysarthria and ataxia of the left upper extremity occurred. In the course of the illness, the hearing loss, ataxia and dysarthria completely recovered. MRI of the brain showed an infarction in the area of the anterior inferior cerebellar artery; neurosonographic examination of the basilar and vertebral arteries was normal. Therefore, in patients with acute hearing impairment or hearing loss, an AICA-ischemia should be considered and the patient carefully examined for additional brainstem symptoms, since this can be the first sign of an life-threatening basilar artery thrombosis. PMID- 15657758 TI - Sequential predator effects across three life stages of the African tree frog, Hyperolius spinigularis. AB - While theoretical studies of the timing of key switch points in complex life cycles such as hatching and metamorphosis have stressed the importance of considering multiple stages, most empirical work has focused on a single life stage. However, the relationship between the fitness components of different life stages may be complex. Ontogenetic switch points such as hatching and metamorphosis do not represent new beginnings--carryover effects across stages can arise when environmental effects on the density and/or traits of early ontogenetic stages subsequently alter mortality or growth in later stages. In this study, I examine the effects of egg- and larval-stage predators on larval performance, size at metamorphosis, and post-metamorphic predation in the African tree frog Hyperolius spinigularis. I monitored the density and survival of arboreal H. spinigularis clutches in the field to estimate how much egg-stage predation reduced the input of tadpoles into the pond. I then conducted experiments to determine: (1) how reductions in initial larval density due to egg predators affect larval survival and mass and age at metamorphosis in the presence and absence of aquatic larval predators, dragonfly larvae, and (2) how differences in mass or age at metamorphosis arising from predation in the embryonic and larval environments affect encounters with post-metamorphic predators, fishing spiders. Reduction in larval densities due to egg predation tended to increase per capita larval survival, decrease larval duration and increase mass at metamorphosis. Larval predators decreased larval survival and had density-dependent effects on larval duration and mass at metamorphosis. The combined effects of embryonic and larval-stage predators increased mass at metamorphosis of survivors by 91%. Larger mass at metamorphosis may have immediate fitness benefits, as larger metamorphs had higher survival in encounters with fishing spiders. Thus, the effects of predators early in ontogeny can alter predation risk even two life stages later. PMID- 15657759 TI - Using animal movement paths to measure response to spatial scale. AB - Animals live in an environment that is patchy and hierarchical. I present a method of detecting the scales at which animals perceive their world. The hierarchical nature of habitat causes movement path structure to vary with spatial scale, and the patchy nature of habitat causes movement path structure to vary throughout space. These responses can be measured by a combination of path tortuousity (measured with fractal dimension) versus spatial scale, the variation in tortuousity of small path segments along the movement path, and the correlation between tortuousities of adjacent path segments. These statistics were tested using simulated animal movements. When movement paths contained no spatial heterogeneity, then fractal D and variance continuously increased with scale, and correlation was zero at all scales. When movement paths contained spatial heterogeneity, then fractal D sometimes showed a discontinuity at transitions between domains of scale, variation showed peaks at transitions, and correlations showed a statistically significant positive value at scales smaller than patch size, decreasing to below zero at scales greater than patch size. I illustrated these techniques with movement paths from deer mice and red-backed voles. These new analyses should help understand how animals perceive and react to their landscape structure at various spatial scales, and to answer questions about how habitat structure affects animal movement patterns. PMID- 15657760 TI - Respiration characteristics in temperate rainforest tree species differ along a long-term soil-development chronosequence. AB - We measured the response of dark respiration (R(d)) to temperature and foliage characteristics in the upper canopies of tree species in temperate rainforest communities in New Zealand along a soil chronosequence (six sites from 6 years to 120,000 years). The chronosequence provided a vegetation gradient characterised by significant changes in soil nutrition. This enabled us to examine the extent to which changes in dark respiration can be applied across forest biomes and the utility of scaling rules in whole-canopy carbon modelling. The response of respiration to temperature in the dominant tree species differed significantly between sites along the sequence. This involved changes in both R(d) at a reference temperature (R(10)) and the extent to which R(d) increased with temperature (described by E(o), a parameter related to the energy of activation, or the change in R(d) over a 10 degrees C range, Q(10)). Site averaged E(o) ranged from 44.4 kJ mol(-1) K(-1) at the 60-year-old site to 26.0 kJ mol(-1) K( 1) at the oldest, most nutrient poor, site. Relationships between respiratory and foliage characteristics indicated that both the temperature response of respiration (E(o) or Q(10)) and the instantaneous rate of respiration increased with both foliar nitrogen and phosphorus content. The ratio of photosynthetic capacity (Whitehead et al. in Oecologia 2005) to respiration (A(max)/R(d)) attained values in excess of 15 for species in the 6- to 120-year-old sites, but thereafter decreased significantly to around five at the 120,000-year-old site. This indicates that shoot carbon acquisition is regulated by nutrient limitations in the retrogressing ecosystems on the oldest sites. Our findings indicate that respiration and its temperature response will vary according to soil age and, therefore, to soil nutrient availability and the stage of forest development. Thus, variability in respiratory characteristics for canopies should be considered when using models to integrate respiration at large spatial scales. PMID- 15657757 TI - Genetic and epigenetic features in radiation sensitivity Part I: cell signalling in radiation response. AB - Recent progress especially in the field of gene identification and expression has attracted greater attention to genetic and epigenetic susceptibility to cancer, possibly enhanced by ionising radiation. It has been proposed that the occurrence and severity of the adverse reactions to radiation therapy are also influenced by such genetic susceptibility. This issue is especially important for radiation therapists since hypersensitive patients may suffer from adverse effects in normal tissues following standard radiation therapy, while normally sensitive patients could receive higher doses of radiation offering a better likelihood of cure for malignant tumours. This paper, the first of two parts, reviews the main mechanisms involved in cell response to ionising radiation. DNA repair machinery and cell signalling pathways are considered and their role in radiosensitivity is analysed. The implication of non-targeted and delayed effects in radiosensitivity is also discussed. PMID- 15657761 TI - Spermatogeny as a life-history index in parasitoid wasps. AB - Reproduction is a major life-history trait but it has been studied mostly in relation to female reproductive effort. Recently, an ovigeny index using the proportion of oocytes ready to be oviposited at eclosion has been proposed for female insect parasitoids. Here, we propose a spermatogeny index for male parasitoid wasps. Prospermatogenic species have an index of 1, have all their spermatozoids mature at emergence and do not produce more later in life. At the other end of the spectrum, synspermatogenic species have no spermatozoids at emergence and produce them later in life. The level of spermatogeny should be linked to several other life-history parameters such as longevity, size, nutrition, distribution of mating opportunities and dispersion before and after mating. Data presented for some parasitoid species support the presence of variability in this male life-history parameter. PMID- 15657762 TI - Individual quality, survival variation and patterns of phenotypic selection on body condition and timing of nesting in birds. AB - Questions about individual variation in "quality" and fitness are of great interest to evolutionary and population ecologists. Such variation can be investigated using either a random effects approach or an approach that relies on identifying observable traits that are themselves correlated with fitness components. We used the latter approach with data from 1,925 individual females of three species of ducks (tufted duck, Aythya fuligula; common pochard, Aythya ferina; northern shoveler, Anas clypeata) sampled on their breeding grounds at Engure Marsh, Latvia, for over 15 years. Based on associations with reproductive output, we selected two traits, one morphological (relative body condition) and one behavioral (relative time of nesting), that can be used to characterize individual females over their lifetimes. We then asked whether these traits were related to annual survival probabilities of nesting females. We hypothesized quadratic, rather than monotonic, relationships based loosely on ideas about the likely action of stabilizing selection on these two traits. Parameters of these relationships were estimated directly using ultrastructural models embedded within capture-recapture-band-recovery models. Results provided evidence that both traits were related to survival in the hypothesized manner. For all three species, females that tended to nest earlier than the norm exhibited the highest survival rates, but very early nesters experienced reduced survival and late nesters showed even lower survival. For shovelers, females in average body condition showed the highest survival, with lower survival rates exhibited by both heavy and light birds. For common pochard and tufted duck, the highest survival rates were associated with birds of slightly above-average condition, with somewhat lower survival for very heavy birds and much lower survival for birds in relatively poor condition. Based on results from this study and previous work on reproduction, we conclude that nest initiation date and body condition covary with both reproductive and survival components of fitness. These associations lead to a positive covariance of these two fitness components within individuals and to the conclusion that these two traits are indeed correlates of individual quality. PMID- 15657763 TI - Genetic transformation of Leymus chinensis with the PAT gene through microprojectile bombardment to improve resistance to the herbicide Basta. AB - Chinese leymus [Leymus chinensis (Trin.) Tzvel.] is a perennial grass (tribe Gramineae) that is widely distributed throughout northern China and Mongolia where it is produced as a forage product. Severe production losses due to weed growth have serious economic consequences, and as non-selective herbicides not only kill the weeds but are also harmful to this forage grass, the introduction of a foreign gene for resistance to the herbicide Basta is necessary since this species lacks herbicide resistance. We have investigated the transformation of a gene for phosphinothricin acetyltransferase (PAT) through microprojectile bombardment in Chinese leymus. Calli from immature inflorescences cultured on N6 medium supplemented with 2.0 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D) and 5.0 mg/l of glutamine were bombarded. The bombarded calli survived on selection medium with 1.0 mg/l of phosphinothricin (PPT). Twenty-three plantlets regenerated from resistant calli on differentiation medium supplemented with 1.0 mg/l 6-benzylaminopurine, 1.0 mg/l kinetin, and 1.0 mg/l PPT, and five of these regenerated plantlets survived on rooting medium with 1.0 mg/l of PPT. PCR and Southern blotting analyses indicated that the PAT gene had been integrated into the genomes of two Chinese leymus plantlets and that the gene was stably transferred to its clonal offsprings. There were no other phenotypic effects associated with transgene expression during vegetative growth except tolerance to the herbicide Basta. PMID- 15657764 TI - Protection of the bone-tendon-bone graft during anterior cruciate ligament reconstruction. AB - Damage of graft fibers is a complication that can happen during arthroscopic reconstruction of the anterior cruciate ligament. It usually occurs while pulling the neoligament through the tibial tunnel, inserting the femoral screw, or doing additional notchplasty. Protection of the graft with a polyethylene tube can reduce the risk of graft laceration. PMID- 15657765 TI - Use of vascularized pedicle iliac bone graft combined with transtrochanteric rotational osteotomy in the treatment of avascular necrosis of the femoral head. AB - INTRODUCTION: A vascularized pedicle iliac bone graft combined with transtrochanteric anterior rotational osteotomy was performed in patients with extensive necrosis in whom the necrotic area occupied more than two-thirds of the weight-bearing zone of the femoral head. The purpose of this procedure is to supply vascularity and mechanical strength to the avascular femoral head. MATERIALS AND METHODS: Seventeen hips in 14 patients (8 male, 6 female) whose average age at surgery was 37.9 (range 21-51) years underwent this procedure between June 1992 and December 2002. These hips (stage 2 in 3 hips, stage 3A in 13 hips, and stage 3B in 1 hip) were examined for changes according to the Japanese Orthopedic Association (JOA ) score and the presence or absence of progression of the clinical stage at least 1 year after surgery. RESULTS: The mean JOA score improved from 67.8 points preoperatively to 78.1 points by 18-133 (mean 50.7) months postoperatively. There was no disease progression to a more advanced stage in 12 of 17 hips (71%) postoperatively. CONCLUSION: A vascularized pedicle iliac bone graft combined with transtrochanteric anterior rotational osteotomy to treat avascular necrosis of the femoral head is considered promising for joint preservation. PMID- 15657766 TI - Aerobic exercise performance correlates with post-ischemic flow-mediated dilation of the brachial artery in young healthy men. AB - In older healthy men, aerobic exercise capacity is related to postischemic flow mediated dilation of the brachial artery (FMD), but corresponding data in a younger population is not available. In addition, whether submaximal aerobic exercise performance also correlates with this kind of vasomotor reactivity is not known. Therefore, in 15 nonsmoking young healthy men [age 27 (5) years; body mass index: 24 (2) kg/m(2); mean (SD)] with different levels of ordinary physical activity, but not performing upper-extremity training, we measured FMD at 1 min after reactive hyperemia, and pulmonary oxygen uptake (VO(2)) at ventilatory anaerobic threshold (VO(2)AT) and at peak effort (peak VO(2)) during an incremental exercise on a treadmill. In our participants, FMD was 9.1 (3.4)%, VO(2)AT was 40.72 (5.92) ml/kg per min, and peak VO(2) was 52.95 (8.13) ml/kg per min. Using bivariate Pearson's correlation, and in separate multivariate regression analyses, VO(2)AT and peak VO(2) showed a significant and reasonably good correlation with FMD (r = 0.84, P < 0.001 and r = 0.77, P = 0.001, respectively), independent of age, body mass index and serum total cholesterol (beta = 0.77, P < 0.001, R(2) of the overall model = 0.79 and beta = 0.70, P < 0.005, R(2) of the overall model = 0.69, respectively). Our data provide evidence suggesting that in young healthy men a higher submaximal and maximal aerobic exercise performance is associated with a greater FMD of peripheral conduit arteries. PMID- 15657767 TI - The generalized force-velocity relationship explains why the preferred pedaling rate of cyclists exceeds the most efficient one. AB - The most efficient pedaling rate (lowest oxygen consumption) at a workload of 50 300 W has been reported to be in the range of 42-60 rpm. By contrast, most competitive cyclists prefer a pedaling rate of more than 90 rpm. The reason for this difference is still unknown. We assume that the high pedaling rate preferred by cyclists can be explained by the inherent properties of muscle fibers. To obtain statements which do not depend on muscle's cross-section and length, we generalized Hill's characteristic equations where muscle force and heat liberation are related to shortening velocity. A pedaling rate of f (etamax) yields to maximal efficiency, whereas the higher pedaling rate f (Pmax) leads to maximal power. The ratio f (Pmax)/f (etamax) between these two pedaling rates ranges from 1.7 to 2.4, and it depends on the muscle's fiber-type composition. In sprints and competitions of very short duration, f (Pmax) is more advantageous because energy supply is not the predominant limiting factor. The price to be paid for the most powerful pedaling rate is lower efficiency and higher energy cost. In longer exercises, economy is more important and the optimal pedaling rate shifts toward f (etamax). We conclude that the optimal pedaling rate, representing the fastest race performance, is not fixed but depends on race duration; it ranges between f (etamax) and f (Pmax). Our results are not only of interest for competitive cyclists but also for investigations using cycle ergometers: maximum power might not be reached by using a pedaling rate near the most efficient one. PMID- 15657768 TI - Combination treatment of CC531-lac-Z rat liver metastases by chemoembolization with pemetrexed disodium and gemcitabine. AB - PURPOSE: The aim of this study was to evaluate the combination effect of pemetrexed disodium (MTA; Alimta; LY 231514) and gemcitabine (GEM) administered by hepatic artery and portal vein chemoembolization (HACE and PVCE) in a colorectal cancer rat liver metastasis model. MATERIALS AND METHODS: Proliferation studies on CC531-lac-Z rat colon cancer cells were performed using the MTT assay to obtain the optimal combination schedule of the two antineoplastic agents. To generate diffuse liver metastasis, 4 x 10(6) tumor cells were implanted into the portal vein of male WAG/Rij rats. MTA (30 mg/kg, 60 mg/kg, and 90 mg/kg) was administered locoregionally by portal vein chemoembolization (PVCE) and compared with repeated systemic intravenous injection. GEM (50 mg/kg) was also given locoregionally by hepatic artery chemoembolization (HACE) as well as systemically. All routes of administration were examined alone as well as in combination. Efficacy of treatment in terms of liver metastases burden was determined at the end of the experiment by measuring the beta-galactosidase activity of CC531-lac-Z cells with a chemoluminescence assay. RESULTS: Combination experiments in vitro showed a more than additive tumor cell reduction after sequential exposure to MTA preceding GEM (observed/expected ratio [O/E] = 0.73). Experiments with the reverse sequence (GEM-->MTA) resulted only in additive combination effects (O/E ratio = 1.08). Simultaneous drug exposure showed less than additive combination effects (O/E ratios > or = 1.25). In vivo, locoregional administration by HACE with GEM was significantly more effective than systemic intravenous bolus treatment (P = 0.03). Portal vein chemoembolization with MTA performed immediately after tumor cell inoculation was ineffective. Repeated systemic treatment with MTA yielded a slight reduction in tumor cell load that was significant versus control at the medium and high doses (60 mg/kg, P = 0.009; 90 mg/kg, P = 0.046) but not versus intraportal chemoembolization. The combination treatment of systemic (60 and 90 mg/kg) or locoregional (60 mg/kg) MTA with HACE using GEM (50 mg/kg) resulted in more than 80% tumor growth inhibition; this antineoplastic combination effect was maximally additive. CONCLUSION: A regimen-dependent synergistic combination effect of both drugs was found in vitro. In animals, hepatic artery chemoembolization with GEM was superior to systemic intravenous bolus treatment. Portal vein chemoembolization with MTA was ineffective. The optimal in vitro regimen of MTA (intravenous or PVCE) preceding GEM (HACE) resulted in a maximally additive tumor growth inhibition. The results indicate that MTA and GEM can successfully be combined and favor further evaluation in patients. PMID- 15657769 TI - [Intraspinal cystic space occupying lesion after cranial subarachnoid hemorrhage]. PMID- 15657770 TI - [Functional imaging of neurocognitive dysfunction in attention deficit hyperactivity disorder]. AB - Attention Deficit Hyperactivity Disorder (ADHD) is a neurobiological disorder of early childhood onset. Defining symptoms are chronic impairments of attention, impulse control and motor hyperactivity that frequently persist until adulthood. Miscellaneous causes of the disorder have been discussed. Accumulating evidence from imaging- and molecular genetic studies strengthened the theory of ADHS being a predominantly inherited disorder of neurobiological origin. In the last 15 years, non-invasive brain imaging methods were successfully implemented in pediatric research. Functional magnetic resonance imaging studies gave major insight into the neurobiological correlates of executive malfunction, inhibitory deficits and psychomotoric soft signs. These findings are in good accordance with brain morphometric data indicating a significant volumetric decrease of major components of striato-thalamo-cortical feedback loops, primarily influencing prefrontal executive functioning (e.g. basal ganglia). Empirical evidence points to a broad array of associated behavioral disturbances like deficient visuomotor abilities and oculomotor dysfunctions. This paper reviews the current empirical evidence derived from prior imaging studies. Special emphasis is given to the relevance of oculomotor dysfunctions in clinical and research settings, as well as their assessment in the MR environment. PMID- 15657771 TI - Differentiation of the follicle-associated epithelium in ileal Peyer's patch and production of 50-nm particles are maintained in B-cell-depleted fetal sheep. AB - To evaluate the dependence of the differentiation of the follicle-associated epithelium (FAE) on the presence of follicular B-cells, the FAE of ileal Peyer's patch follicles was examined in B-cell-depleted fetal lambs. The FAE of these rudimentary follicles, which are devoid of lymphocytes, showed normal differentiation, including carbonic anhydrase reactivity and ultrastructural characteristics of transcytosis, extensive interdigitation of the lateral plasma membrane and the shedding of membrane-bounded particles, approximately 50 nm in size, resembling exosomes. These 50-nm membrane-bounded particles were abundant in the extracellular space of the epithelium and the dome but no particles were found in the rudimentary follicles. This study confirms that the rudimentary follicles consist of clusters of follicular dendritic cells. Our findings suggest that the differentiation of FAE of ileal Peyer's patch and the production of the 50-nm particles constitute features that appear to be independent of B-cells. PMID- 15657772 TI - Progression of varicella-zoster virus necrotizing retinopathy in an HIV-negative patient with transient immune deviation. AB - PURPOSE: To report a case of unilateral varicella-zoster virus (VZV) necrotizing retinopathy that progressed from outer retinitis with features of progressive outer retinal necrosis (PORN) to typical acute retinal necrosis (ARN) in an HIV negative patient with a transient decrease in CD4 lymphocyte counts and CD4/CD8 ratio. METHOD: Case report. RESULTS: A 41-year-old Chinese man presenting with blurred vision in the right eye was diagnosed with herpetic necrotizing retinitis without vitritis. Fundus examination revealed retinal arteritis and extensive deep whitish retinal lesions in the mid-periphery with minimal vitritis. Aqueous humor and vitreous PCR were positive for VZV. His CD4 count on presentation was depressed (239 cells/ul) and the CD4/CD8 ratio was low (0.8). The referring ophthalmologist had treated him with prednisolone 60 mg/day. At our institution, when intravenous acyclovir was started and the steroid therapy discontinued, he developed severe vitritis and the deep retinal lesions progressed to full thickness retinitis typical of ARN. Repeat CD4 count was 512 cells/ul at day 14. In total, he was treated with 14 days of i.v. acyclovir (12 mg/kg 8-hourly) followed by oral valaciclovir 500 mg three times a day for 3 months. Prednisolone 30 mg once daily was restarted and tapered over 3 months. Despite prophylactic argon retinal photocoagulation to the edge of the retinitis, the patient developed a total retinal detachment at 3 months. CONCLUSIONS: VZV retinal infection in an HIV-negative patient with transient immune deviation can manifest initially as outer retinitis with features similar to PORN and progress to typical ARN when CD4 counts return to normal. PMID- 15657774 TI - Pars plana vitrectomy with internal limiting membrane removal for macular hole associated with proliferative diabetic retinopathy. AB - BACKGROUND: We describe the outcome of vitreous surgery in three eyes of three patients with macular hole associated with proliferative diabetic retinopathy (PDR) in the absence of fibrovascular proliferation, a combination of conditions where efficacy is incompletely known. METHODS: The patients, all male were 62, 65, and 66 years old. Panretinal photocoagulation had been performed preoperatively in all, and one eye had undergone vitreous surgery. No fibrovascular tissue causing macular traction was observed in any case. Fluorescein angiography and optical coherence tomography (OCT) demonstrated persistent diabetic macular edema surrounding the hole. Affected eyes were treated with vitrectomy including internal limiting membrane (ILM) peeling; 20% sulfur hexafluoride gas (SF(6)) was introduced for tamponade. RESULTS: Anatomical closure of the macular hole as well as resolution of macular edema was achieved in all cases, and vision improved considerably by more than two Snellen lines. CONCLUSIONS: Vitreous surgery with ILM peeling was effective for macular hole associated with PDR, attaining not only macular hole closure but also resolution of persistent diabetic macular edema. PMID- 15657773 TI - Response of retinal blood flow to systemic hyperoxia in smokers and nonsmokers. AB - BACKGROUND: We investigated the influence of chronic smoking on ocular vascular reactivity during breathing of 100% oxygen. METHODS: Retinal vascular reactivity was tested during inhalation of 100% oxygen over 10 min. The observer-masked two cohort study was performed in 24 healthy male volunteers (12 smokers and 12 nonsmokers) using the Zeiss Retinal Vessel Analyzer and laser Doppler velocimetry. From these parameters retinal blood flow was calculated. RESULTS: Hyperoxia significantly decreased arterial (smokers: p<0.001 vs baseline; nonsmokers: p=0.003 vs baseline) and venous (smokers: p<0.001 vs baseline; nonsmokers: p<0.001 vs baseline) diameters. This decrease was significantly more pronounced in smokers (arterial diameter: p<0.001, venous diameter: p=0.003). Hyperoxia decreased venous blood flow velocity (smokers: p=0.02 vs baseline; nonsmokers: p<0.001 vs baseline) to a comparable degree (p=0.51). The two groups showed a comparable decrease in retinal blood flow during hyperoxia (smokers: p<0.001 vs baseline; nonsmokers: p<0.001 vs baseline; p=0.76 between groups). The decrease of PCO(2) during inhalation of 100% oxygen was significantly more pronounced in smokers than in nonsmokers (p=0.038). CONCLUSION: The present study indicates an abnormal retinal vascular response to hyperoxia in smokers. Further studies are needed to identify possible neural or humoral factors involved in this shifted vasoconstrictory status in smokers. PMID- 15657775 TI - Correlation between capillary blood flow of retina estimated by SLDF and circulatory parameters of retrobulbar blood vessels estimated by CDI in diabetic patients. AB - BACKGROUND: The aim of this study was to investigate the correlation between the recordings of scanning laser Doppler flowmetry (SLDF) of the retina and the recordings of color Doppler imaging (CDI) of the retrobulbar circulatory parameters in diabetic patients without diabetic retinopathy. METHODS: Twenty three diabetic patients without diabetic retinopathy were evaluated using SLDF for the apparent retinal circulation and using CDI for the apparent retrobulbar circulation in the central retinal artery, the central retinal vein and the short posterior ciliary artery. The circulatory parameters estimated in the retinal tissue using SLDF were velocity, volume and flow. The Circulatory parameters that were recorded using the CDI method were peak systolic velocity (PSV), end diastolic velocity (EDV), mean velocity (MV), pulsatility (PI) and resistivity index (RI). We obtained the correlation coefficients between parameters of SLDF and CDI. Multiple regression analysis was performed with "flow" parameter of SLDF recordings as a dependent variable and all estimated CDI parameters as independent variables. Multiple linear regression was also performed, including the "flow" parameter of SLDF recordings as a dependent variable and PI of the CDI parameters of all the measured blood vessels as independent variables. RESULTS: The "velocity" parameter of SLDF was significantly correlated with the PI in the central retinal artery (P=0.02), PI and RI in the central retinal vein (P=0.01; P=0.01) and the PSV, MV, PI and RI in the short posterior ciliary artery, as recorded by CDI (P=0.003; P=0.02; P=0.002; P=0.01). The "volume" parameter of SLDF was significantly correlated with the PI and RI in the central retinal vein (P=0.03; P=0.03) and the PSV in the short posterior ciliary artery (P=0.03), as recorded by CDI. The "flow" parameter of SLDF was significantly correlated with the PI and RI in the central retinal vein (P=0.01; P=0.01) and the PSV, MV, PI and RI in the short posterior ciliary artery (P=0.003; P=0.03; P=0.002; P=0.01) as measured by CDI. The multiple regression analysis was statistically non significant (P=0.86). The multiple linear regression analysis indicated that from among the PI of the evaluated blood vessels, the PI of the short posterior ciliary artery was the most significant predictor of the "flow" parameter of SLDF (P=0.01). CONCLUSION: This study suggests a positive correlation between the recordings of SLDF of the retinal tissue and the retrobulbar circulatory parameters of the CDI in diabetic patients without diabetic retinopathy. PMID- 15657777 TI - Toxic clozapine serum levels during acute urinary tract infection: a case report. PMID- 15657776 TI - Angioedema induced by tramadol--a potentially life-threatening condition. PMID- 15657778 TI - Adverse reactions related to drugs for headache treatment: clinical impact. AB - OBJECTIVE: To assess the clinical impact of adverse reactions related to drugs for primary headache treatment. METHODS: We examined the adverse reactions to 360 medications prescribed by the specialists of the Headache Centre of the University of Modena and Reggio Emilia to 256 consecutive outpatients (214 female, 42 male; mean age: 38.88 +/- 14.06 years; range 10-72 years). Adverse reactions were reported by patients during scheduled follow-up visits, classified by specialists and reassessed by a clinical pharmacologist. RESULTS: Adverse reactions with a causal relationship classified as definite/probable/possible were 202 (56%): 62% (80/129) were due to acute treatments and 53% (122/231) to prophylactic treatments (chi2 test, P = 0.115 ns). More than 90% of the adverse reactions were of limited intensity [mild (58%) or moderate (36%)]. Only 5% were severe, and two reactions (1%) were serious. The most affected apparatus was the nervous system (41%). Of these adverse reactions, 43% caused the discontinuance of the treatment, especially of prophylaxis (54%). Patients evaluated 70% of the medications as effective, but, at the same time, they considered most of the adverse reactions (69%) unacceptable. CONCLUSION: Adverse reactions related to headache medications have a strong impact on patients' management, even if their real intensity and severity are usually very limited. Drugs for headache treatment are still far from being ideal drugs. To prevent the discontinuance of effective medications, the physician, prior to prescribing, should assess, together with the patient, the acceptability of the more common adverse drug reactions. PMID- 15657780 TI - Dextromethorphan metabolic phenotyping in an Iranian population. AB - OBJECTIVE: CYP2D6 polymorphism of drug metabolism represents an important source of interindividual and interethnic variation in drug response. Since this polymorphism has not been studied in an Iranian population, the present study was undertaken. METHODS: Two hundred healthy unrelated Iranian subjects participated in this study. Phenotyping was based on high-performance liquid chromatography determination of the dextromethorphan/total dextrorphan molar ratios as metabolic ratios (MRs) in plasma samples collected at 3 h after oral administration of 30 mg dextromethorphan hydrobromide. Since the dextromethorphan detection limit of 5 ng/ml achieved in the simultaneous assay for dextromethorphan and its metabolites was not adequate to identify intermediate metabolizers (IMs), 80 of 200 samples selected randomly were also assayed using a modified, more sensitive procedure with a dextromethorphan detection limit of 1 ng/ml. RESULTS: Poor and extensive metabolizers (EMs) could be identified distinctly. A 520-fold interindividual variation in dextromethorphan MRs was observed in this study. In contrast to undetectable dextrorphan and hydroxymorphinan concentrations, clearly determinable dextromethorphan concentrations higher than 10 ng/ml were observed in plasma samples of poor metabolizers (PMs). Considering the antimode of 0.3, five (2.5%, 95% confidence interval of 0.34-4.66) volunteers were identified as PMs. Using the more sensitive method, dextromethorphan was quantified in 4 (one PM) of 80 samples. Excluding the PM, a Shapiro-Wilk test indicated a non-normal distribution of MRs (P < 0.01) in the latter population. CONCLUSIONS: From this study it can be concluded that the frequency of PMs in an Iranian population is 2.5% (95% confidence interval of 0.34-4.66). IMs could be identified using dextromethorphan plasma assays with detection limits of at least 1 ng/ml. However, the phenotype-genotype relationships in this respect remain to be established. PMID- 15657781 TI - Effect of benzodiazepines on the metabolism of buprenorphine in human liver microsomes. AB - OBJECTIVE: To determine whether enzyme inhibition explains the clinical adverse interaction of benzodiazepines and buprenorphine. METHODS: Buprenorphine was incubated in the presence of benzodiazepines (or metabolites) with human liver microsomes (HLMs). A number of benzodiazepines were screened at therapeutic concentrations after 0-min and 15-min preincubation times. For tentative metabolically activated inhibitors, the kinetics of inhibition was studied in a secondary incubation system. Buprenorphine and norbuprenorphine were quantified by means of liquid chromatography-mass spectrometry. RESULTS: Buprenorphine elimination and norbuprenorphine formation were at most reduced by 26% (i.e., weak or negligible inhibition). Evidence of metabolically activated inhibition suggested the need for further studies on the inhibitory kinetics. Midazolam caused time- and concentration-dependent inhibition of norbuprenorphine formation with pseudo-first-order kinetics, and K(I) and k(inact) values of 10.5 microM and 0.045 min(-1), respectively. Mixed-type inhibition of buprenorphine elimination (K(i) = 30-35 microM) and a noncompetitive inhibition of norbuprenorphine formation were also observed. For clonazepam (up to 10 microM), 3-hydroxy-7 acetamidoclonazepam (up to 10 microM), and alpha-hydroxy-triazolam (up to 1.0 microM), no time- or concentration-dependent inhibition of buprenorphine metabolism was found. CONCLUSION: A single benzodiazepine, midazolam, is a moderate mechanism-based inactivator of buprenorphine N-dealkylation. It is anticipated that repeated exposures to midazolam might alter the in vivo metabolism of buprenorphine. PMID- 15657782 TI - Variations of CYP3A activity induced by antiretroviral treatment in HIV-1 infected patients. AB - OBJECTIVE: To measure the in vivo variations of CYP3A activity induced by anti HIV drugs in human immunodeficiency virus (HIV)1-positive patients. METHODS: A low oral dose of midazolam (MID) (0.075 mg) was given to the patients and the 30 min total 1-OH midazolam (1-OHMID)/MID ratio was determined. Patients were phenotyped either before the introduction of antiretroviral treatments (control group, 90 patients) or after a variable period of antiretroviral treatment (56 patients). Twenty-one subjects underwent multiple phenotyping tests (before and during the course of the treatment). RESULTS: The median MID ratio was 3.51 in the control group (range 0.20-14.6). It was 5-fold higher in the group with efavirenz (28 patients; median, range: 16.0, 3.81-367; P < 0.0001), 13-fold lower with nelfinavir (18 patients; 0.27, 0.06-36.3; P < 0.0001), 17-fold lower with efavirenz + ritonavir (three patients; 0.21, 0.05-0.47; P = 0.006), 50-fold lower with ritonavir (four patients; 0.07, 0.06-0.17; P = 0.0007), and 7-fold lower with nevirapine + (ritonavir or nelfinavir or grapefruit juice) (three patients; 0.48, 0.03-1.83; P = 0.03). CYP3A activity was lower in the efavirenz + ritonavir group (P = 0.01) and in the ritonavir group (P = 0.04) than in the nelfinavir group, although already strongly inhibited in the latter. CONCLUSION: The low dose MID phenotyping test was successfully used to measure the in vivo variations of CYP3A activity induced by antiretroviral drugs. Efavirenz strongly induces CYP3A activity, while ritonavir almost completely inhibits it. Nelfinavir strongly decreases CYP3A activity, but to a lesser extent than ritonavir. The inhibition of CYP3A by ritonavir or nelfinavir offsets the inductive effects of efavirenz or nevirapine administered concomitantly. Finally, no induction of CYP3A activity was noticeable after long-term administration of ritonavir at low dosages (200 mg/day b.i.d.) or of nelfinavir at standard dosages (2,500 mg/day b.i.d.). PMID- 15657783 TI - Pharmacokinetics of N-acetylcysteine following repeated intravenous infusion in haemodialysed patients. AB - OBJECTIVE: N-acetylcysteine (NAC) is a mucolytic agent with anti-oxidant properties. It might have potential positive effects in renal patients and, therefore, its pharmacokinetics and safety in haemodialysis was investigated. METHODS: Twelve dialysis patients received 2 g NAC (10 ml NAC 20% solution i.v.) mixed with 500 ml saline during the first 3 h of the session for six dialysis sessions. A bolus of heparin was injected intravenously as LWH-heparin. In six patients, one session was repeated with NAC mixed with heparin and infused through the heparin pump. RESULTS: Baseline NAC was on average 454 ng ml(-1); its concentration increased to 9,253 ng ml(-1) at the second infusion and attained a steady state between 14,000 ng ml(-1) and 17,000 ng ml(-1) at the fourth dose. We observed a C (max) of 53,458 ng ml(-1) with a t (max) of 3.0 h. Plasma clearance was 1.25 l h(-1) and dialytic clearance 5.52 l h(-1). No side effects were observed. CONCLUSION: In the case of repeated doses, the NAC pre-dose concentration after repeated infusion of 2 g of the drug during the first 3 h of a dialysis session reached the steady state at the fourth infusion, without further accumulation. The dialytic clearance is effective, the total body clearance being reduced to 1.25 l h(-1). In dialysis patients, 2 g NAC given intravenously over 3 h is a safe dosage, with no short-term side effects. PMID- 15657784 TI - The psychopharmacology of herbal extracts: issues and challenges. PMID- 15657785 TI - Occult spinal dysraphism and Pacinian hamartomas. AB - CASES: The authors report two patients with occult spinal dysraphism who, following histological analysis, were found to harbor associated Pacinian hamartomas. One patient's hamartoma was found in conjunction with their lipomyelomeningocele and the other was combined with a presacral myxopapillary ependymoma associated with a fatty filum terminale. DISCUSSION AND CONCLUSION: We review the literature and believe this to be only the third report of Pacinian hamartomas in association with occult spinal dysraphism. Although Pacinian hamartomas are seemingly rare in conjunction with occult spinal dysraphism, mesodermal elements are often found in this disease entity. PMID- 15657786 TI - Reliability of a new adjustable shunt device without the need for readjustment following 3-Tesla MRI. AB - INTRODUCTION: 3-Tesla (T) magnetic resonance imaging (MRI) offers high resolution imaging with improved signal quality. It also allows for improved functional investigations, most prominently with respect to fiber tracts and their relation to pathological lesions. Up to now, patients with adjustable shunt systems were not eligible for high field power magnetic resonance imaging. We have evaluated the effects of this technique upon a newly developed adjustable shunt valve. METHOD: Ten adjustable shunt devices were examined during routine and functional 3-T high field MRI examinations. Pressure settings were checked after 17 examinations each for all valves. There were no changes in pressure setting at all possible levels in any of the tested devices. No problems with the adjusting mechanism were observed during 340 examinations. CONCLUSION: This new shunt device offers the diagnostic benefit of high field magnetic resonance imaging in shunt-dependent patients who need an adjustable valve. The valve is not affected by higher magnetic field power and does not require readjustment of the pressure settings after MRI examination. PMID- 15657787 TI - Intracranial peripheral primitive neuroectodermal tumor manifesting as abducens nerve palsy. AB - INTRODUCTION: Peripheral primitive neuroectodermal tumor (PNET) occurring as an extra-axial lesion within the intracranial space and extending to the subarachnoid space is extremely rare. CASE REPORT: An 18-month-old girl presented with an intracranial peripheral PNET manifesting as abducens nerve palsy. Magnetic resonance imaging on admission revealed a lesion affecting the trigeminal and abducens nerves. The tumor was partially removed via the subtemporal approach. Histological examination showed a high-grade, undifferentiated neoplasm of small cell type with positive immunostaining for MIC2. The histological diagnosis was peripheral PNET. OUTCOME: Craniospinal radiotherapy reduced the tumor size, but adjuvant chemotherapy designed for Ewing's sarcomas and PNETs was not effective. She died 1 month after the last chemotherapy, despite whole craniospinal irradiation (total dose 53.2 Gy) and chemotherapy. PMID- 15657788 TI - Distribution territories and causative mechanisms of ischemic stroke. AB - Ischemic stroke prognosis, risk of recurrence, clinical assessment, and treatment decisions are influenced by stroke subtype (anatomic distribution and causative mechanism of infarction). Stroke subtype diagnosis is better achieved in the early phase of acute ischemia with the use of multimodal MR imaging. The pattern of brain lesions as shown by brain MR imaging can be classified according to a modified Oxfordshire method, based on the anatomic distribution of the infarcts into six groups: (1) total anterior circulation infarcts, (2) partial anterior circulation infarcts, (3) posterior circulation infarcts, (4) watershed infarcts, (5) centrum ovale infarcts, and (6) lacunar infarcts. The subtype of stroke according to its causative mechanism is based on the TOAST method, which classifies stroke into five major etiologic groups: (1) large-vessel atherosclerotic disease, (2) small-vessel atherosclerotic disease, (3) cardioembolic source, (4) other determined etiologies, and (5) undetermined or multiple possible etiologies. The different MR imaging patterns of acute ischemic brain lesions visualized using diffusion-weighted imaging and the pattern of vessel involvement demonstrated with MR angiography are essential factors that can suggest the most likely causative mechanism of infarction. This information may have an impact on decisions regarding therapy and the performance of additional diagnostic tests. PMID- 15657790 TI - Diagnostic value of pelvic radiography in the initial trauma series in blunt trauma. AB - The purpose of the study was to evaluate the diagnostic value of pelvic radiography in the initial trauma series when compared to multidetector CT (MDCT) findings in serious blunt trauma. Inclusion criteria were blunt trauma and pelvic radiography in the initial trauma series, followed by a whole-body MDCT. A total of 1386 patients (874 male, 512 female, age 16-91 years, mean 41 years) met the inclusion criteria. Imaging studies were evaluated retrospectively by anatomical region and classified, when possible, using the Tile classification. Based on MDCT, a total of 629 injuries occurred in 226 (16%) of these 1386 patients. Radiography depicted 405 fractures in these 226 patients, giving an overall sensitivity of 55%. In 24 patients (11%) radiography was false-negatively normal. The sensitivity of radiography was mainly good in the anteroinferior parts of the pelvis, fair in the acetabulum and ileum, and poor in the posterior ring. By MDCT 141 (62%) patients were classified using the Tile classification and by radiography 133 patients (59%) were classified. MDCT and radiography showed the same type of pelvic injury in 72 patients (59%) and the subtype in 17 patients (14%). In 48 patients (40%) the pelvis was shown to be stable by radiography but unstable by MDCT. In conclusion, the sensitivity of pelvic radiography is low, and it is not reliable for determining if the pelvic injury is stable or not. PMID- 15657789 TI - Ischemic stroke: carotid and vertebral artery disease. AB - Ischemic strokes may have distinct aetiologies, including several different intrinsic arterial pathological disorders. The diagnosis and understanding of these arterial diseases is critical for the correct management of stroke as different treatment approaches are undertaken according to the aetiology. Atherosclerosis is by far the most common arterial disease among adults, and other pathological processes include arterial dissection, small vessel disease, inflammatory and non-inflammatory vasculopathy and vasomotor disorders. In children, there are several vasculopathies responsible for vaso-occlusive disease such as sickle-cell anemia, acute regressive angiopathy and Moya-Moya disease, neurofibromatosis, dissections, vasculitis associated with intracranial and systemic infections. An overview of the major carotid and vertebral pathological diseases responsible for ischemic stroke in adults and children, highlighting the accuracy of the different imaging modalities for its diagnosis and the imaging appearance of these diseases, is given. PMID- 15657791 TI - How is pulmonary arterial blood flow affected by pulmonary venous obstruction in children? A phase-contrast magnetic resonance study. AB - BACKGROUND: Hemodynamic changes within a stenosed pulmonary vein might not reflect the severity of the obstruction if redistribution of pulmonary artery flow occurs. OBJECTIVE: The purpose was to investigate flow changes in branch pulmonary arteries in the presence of pulmonary vein stenosis. MATERIALS AND METHODS: Twelve children (age range 3-180 months) who had undergone MRI for pulmonary vein stenosis were identified. The severity of vein stenosis was assessed from percentage diameter reduction. Pulmonary artery flow distribution was correlated with the severity of pulmonary vein stenosis. Nine of the patients had unilateral stenosis; three had bilateral involvement. RESULTS: Unilateral vein stenosis was associated with diastolic reversal in the ipsilateral branch pulmonary artery (mean flow reversal 12.3%, range 2.4-42%) and continuous diastolic forward flow in the contralateral pulmonary artery in seven of nine patients. Severe stenosis was associated with decreased systolic flow to the ipsilateral lung. The ratio of net forward flow through pulmonary arteries correlated well with the cross-sectional area ratio of pulmonary arteries (r=0.75, P=0.006). CONCLUSIONS: Severe pulmonary vein obstruction results in redistribution of pulmonary arterial flow. When investigating pulmonary vein stenosis by MR, an evaluation of the pulmonary arterial system should be included to assess the functional importance of an obstruction. PMID- 15657793 TI - Imaging findings in pleuropulmonary blastoma. AB - BACKGROUND: Pleuropulmonary blastoma (PPB) is a rare tumor of the chest seen in young children and recently recognized as distinct from the pulmonary blastoma typically encountered in adults. OBJECTIVE: The purpose of this study is to review and describe the findings of PPB on radiography and CT in four patients. METHODS: Radiographs and CT findings were reviewed in four patients with pathologically proven PPB. RESULTS: All four cases demonstrated large masses in the right hemithorax with heterogeneous low attenuation, pleural effusion, contralateral mediastinal shift, and lack of chest wall invasion. CONCLUSION: When a large pleural-based mass is identified in a young child, PPB should be considered. Suggestive findings include absence of chest wall invasion, presence of pleural fluid, right-sided location, and heterogeneous low attenuation. PMID- 15657792 TI - Atelectasis of the left lung induced by subcarinal pulmonary artery sling. AB - Pulmonary artery sling is an unusual cause of respiratory distress in the neonatal period. Despite the use of conventional diagnostic tools such as ECHO or angiography, delineation of the course of the pulmonary artery and its relationship with the neighbouring bronchus may be difficult. We describe an infant with complete collapse of the left lung due to an aberrant left pulmonary artery in whom the combination of axial CT and three-dimensional reconstruction was of great value in facilitating the diagnosis and surgical planning. PMID- 15657794 TI - Modelling cell death in human tumour cell lines exposed to the anticancer drug paclitaxel. AB - Most anti-cancer drugs in use today exert their effects by inducing a programmed cell death mechanism. This process, termed apoptosis, is accompanied by degradation of the DNA and produces cells with a range of DNA contents. We have previously developed a phase transition mathematical model to describe the mammalian cell division cycle in terms of cell cycle phases and the transition rates between these phases. We now extend this model here to incorporate a transition to a programmed cell death phase whereby cellular DNA is progressively degraded with time. We have utilised the technique of flow cytometry to analyse the behaviour of a melanoma cell line (NZM13) that was exposed to paclitaxel, a drug used frequently in the treatment of cancer. The flow cytometry profiles included a complex mixture of living cells whose DNA content was increasing with time and dying cells whose DNA content was decreasing with time. Application of the mathematical model enabled estimation of the rate constant for entry of mitotic cells into apoptosis (0.035 per hour) and the duration of the period of DNA degradation (51 hours). These results provide a dynamic model of the action of an anticancer drug that can be extended to improve the clinical outcome in individual cancer patients. PMID- 15657796 TI - The effect of a non-uniform turning kernel on ant trail morphology. AB - An ordinary differential equation model is constructed for the formation of pheromone trails by ants on a pre-determined network. At each junction of the trails the probability that an ant will turn through any particular angle is given by a turning kernel. We prove analytically using analogies with thermodynamics that turning behaviour determines trail morphology when the turning kernel is steep. We conjecture that this is also true in general for non uniform turning kernels and present numerical simulations as evidence. Using this conjecture we show the existence of three types of collective foraging: individuals exploring without the use of a trail network, and two distinct types of trail networks; one that consists of low pheromone concentration trails that bend, branch and dissipate and one that consists of high pheromone concentration, straight, unbranched trails. We show that the form of the pheromone response function is crucial in determining the existence and stability of the steady states corresponding to these three foraging strategies, and examine the bifurcations between different trail morphologies as a function of turning kernel steepness for a particular response function. PMID- 15657795 TI - Stability analysis of Turing patterns generated by the Schnakenberg model. AB - We consider the following Schnakenberg model on the interval (-1,1): [formula see text] where D1 > 0, D2 > 0, B > 0. We rigorously show that the stability of symmetric N-peaked steady-states can be reduced to computing two matrices in terms of the diffusion coefficients D1, D2 and the number N of peaks. These matrices and their spectra are calculated explicitly and sharp conditions for linear stability are derived. The results are verified by some numerical simulations. PMID- 15657797 TI - [Allocating and scheduling operating room time]. AB - During the past decade many scientific advances have been made concerning the development of methodologies to maximize efficiency of surgical facilities through allocating and scheduling of operating rooms. In this article such a methodology is described. Using the analysis of historical data of surgical activity in a facility, future demand is predicted and planned. Part of the methodology includes principles and rules needed for the daily organization and operative management of surgical facilities. They are also derived from the same science and therefore the basis for rational and structured decision making. Medical aspects such as patient safety and free choice of day for surgery have higher priority than the economic goal of maximizing operating room efficiency. PMID- 15657799 TI - Biological activity and physicochemical parameters of marine halogenated natural products 2,3,3',4,4',5,5'-heptachloro-1'-methyl-1,2'-bipyrrole and 2,4,6 tribromoanisole. AB - Physicochemical parameters (vapor pressure, water solubility, Henry's law constant) and biological activities of two halogenated natural products frequently detected in marine samples and food were determined. Synthetic 2,3,3',4,4',5,5'-heptachloro-1'-methyl-1,2'-bipyrrole (Q1) and 2,4,6 tribromoanisole (TBA) were available in pure form. The physicochemical parameters were in the range of anthropogenic chlorinated compounds of concern. The aqueous solubilities at 25 degrees C (S(w,25)) of Q1 and TBA were 4.6 microg/L and 12,200 microg/L, respectively, whereas subcooled liquid vapor pressures were 0.00168 Pa (Q1) and 0.06562 Pa (TBA) as measured by the gas chromatographic-retention time technique. Q1 was negative by established test systems for the determination of ethoxyresorufin-O-deethylase (EROD) induction and by sulforhodamine B assay. EROD induction potency was at least 10(-7) times lower than that of 2,3,7,8 tetrachlorodibenzo-p-dioxin (TCDD). At a relatively high concentration (20 microM), Q1 inhibited specific binding of 2 nM [(3)H]TCDD to the in vitro expressed human aryl hydrocarbon receptor (AHR) by 18%; lower concentrations showed no effect. Molecular modeling showed that Q1 is nonplanar, consistent with its relatively modest affinity as an AHR ligand. When tested for cell-growth inhibitory/cytocidal activity in human tumor cells, Q1 was only marginally, if at all, active with an IC(50) value >50 microM compared with five to ten times lower IC(50) values for potent cytotoxins tested in the test system used. Furthermore, standard pesticide tests on insecticidal, herbicidal, and fungicidal activity did not provide any significant activity at highest concentrations. For TBA, the results in all tests were comparable with Q1. The SRB assay was also applied to the halogenated natural product 4,6-dibromo-2-(2',4'-dibromo)phenoxyanisole, but no toxic response was found. Although it was apparent that Q1 and TBA had been proven to have relatively low biological activity in all tests performed, further research is necessary to clarify whether metabolites of the compounds eventually may possess a risk to humans or other living organisms. Nevertheless, the role of Q1 in nature remains uncertain. PMID- 15657798 TI - Apolipoprotein E is associated with age at onset of amyotrophic lateral sclerosis. AB - Apolipoprotein E (APOE) is a confirmed risk factor for Alzheimer disease. APOE is also involved in several other neurodegenerative disorders, including Parkinson disease and multiple sclerosis. Previous studies of amyotrophic lateral sclerosis (Lou Gehrig disease, ALS) have investigated the effect of APOE on the risk of developing ALS, age at onset, site of onset, and duration of the disease. The results have been inconsistent, possibly due to small sample sizes and complete reliance on case-control data. No family-based association studies were performed. To address these limitations, we investigated the relationship between APOE functional polymorphisms and age at onset of ALS in a large set of 508 families. We treated age at onset as a quantitative trait and performed family based association analysis using the TDTQ5 method. APOE-2 is protective against earlier onset (P =0.001) with an average age at onset of APOE-2 carriers approximately 3 years later than that of non-APOE-2 carriers. Similar to our previous report, we did not find APOE associated with ALS risk. Our findings suggest that APOE may express its strongest effect through age at onset rather than on risk. PMID- 15657800 TI - Hexachlorocyclohexane use in the former Soviet Union. AB - Because of the large size of the former Soviet Union (FSU) and the heavy use of organochlorine pesticides (OCPs) in the FSU, usage information regarding OCPs in the FSU is important in compiling global emission inventories and thus in studying the transport of OCPs among different environmental compartments worldwide. The availability of such information is limited. By analyzing the available 1,2,3,4,5,6-hexachlorocyclohexane (HCH) data in the FSU, this article presents estimates of HCH usage in this region from 1950 to 1990, when HCH was officially banned for agricultural use by the FSU government. The creation of HCH usage inventories for the FSU has paved the way to produce HCH emission inventories for this region. Total HCH usages for agricultural purposes in the FSU from 1950 to 1990 were estimated to be 1,960 kt for technical HCH and 40 kt for lindane. The total usage for the isomers was 270 kt for gamma-HCH, 1,270 kt for alpha-HCH, and 170 kt for beta-HCH. Use of HCH reached a peak in 1965: 130 kt for technical HCH, 2.7 kt for lindane, 18 kt for gamma-HCH, 86 kt for alpha-HCH, and 11 kt for beta-HCH. Gridded usage data sets in the FSU of technical HCH and lindane-and the alpha-HCH, beta-HCH and gamma-HCH isomers-on a 1degree x 1degree longitude and latitude grid system for 1980 are freely available to all users at http://www.msc.ec.gc.ca/data/gloperd/. PMID- 15657801 TI - A pilot study of the measurement and control of deep dust, surface dust, and lead in 10 old carpets using the 3-spot test while vacuuming. AB - This pilot study measured and examined the relationship between surface dust, deep dust, lead (Pb), and the 3-spot test during vacuuming of carpets. The 3-spot test measures the total time in seconds for the indicator light on a Hoover vacuum with dirt detector (HVDD) to turn from red to green on three spots 3 feet apart at least 4 feet from an entrance door. Ten older carpets were sampled in the Seattle area by using the following: (1) a 3-spot test to estimate the deep dust; (2) measuring the surface dust in g/m(2) with the High-Volume Small Simplified Surface Sampler; (3) vacuuming with an HVDD to extract a portion of the deep dust in g/m(2); and (4) repeating this cycle of a 3-spot test, surface dust sample, and deep dust sample until the clean-carpet criteria was reached. Dust particles <150 mum were analyzed for Pb. The surface dust, deep dust, and dust collection rate (g/min) decreased rapidly at first and then much more slowly as vacuuming continued. The initial 3-spot test ranged from 12 to 110 seconds (median 40). The starting surface dust loading was 0.7 to 21.1 g/m(2) (median 2.9 g/m(2)), and it decreased by 84% to 99% when the deep dust was removed. Total dust-the sum of the surface dust and deep dust-ranged from 8.3 to 465 g/m(2) (median 63.2 g/m(2)). It took from 2.3 to 92 min/m(2) (median of 7.5 min/m(2)) to remove the total dust. The starting dust collection rate of 3.8 to 37 g/min decreased to final rates of 0.5 to 4.3 g/min. The 3-spot test (seconds) correlated with total dust (g/m(2)) (r = 0.63, p = 0.037) and cleaning time (min/m(2)) (r = 0.50, p = 0.12) when the data were log transformed. This study supports the utility of the 3-spot test. It tends to provide families and professional carpet cleaners with a quick and low-cost estimate of the deep dust present and the time required to clean carpets as well as indicating when the carpet is clean. Deep dust tends to accumulate in older carpets and becomes surface and airborne dust after activity on a carpet. Monitoring and removing the deep dust in old carpets may decrease the exposure of infants and sensitive adults to Pb, allergen, and other pollutants in house dust. PMID- 15657803 TI - Sorption of chlorophenolates in soils and aquifer and marine sediments. AB - This article describes the sorption behavior of 3 hydrophobic ionizable chlorophenols-2,4,6-trichlorophenol, 2,3,4,6-tetrachlorophenol, and pentachlorophenol-in different types of natural sorbents. A series of experiments was carried out with 11 topsoil samples, 9 aquifer sediments, and 12 marine sediments differing in pH, organic-matter content, and mineral composition and presumably also in type of organic matter due to their differing origins. Ionized forms of chlorophenols dominated in almost all sorption experiments. Freundlich isotherm coefficients K(f) and 1/n, as well as organic-matter sorption coefficient (log K(om)) and free-energy change (DeltaG(o)), were calculated for all 3 compounds in all sorbents. The sorption intensity of predominantly ionized chlorophenols increased linearly with the increase of sorbent organic-matter content and decreased with the increasing sorbent pH. Different sorption behavior of all 3 compounds in marine sediments with respect to topsoils and aquifer sediments was indicated by significant differences in K(f) and 1/n coefficients as well as in log K(om) and DeltaG(o) values. The highest K(f) and log K(om) values were obtained for sorption of chlorophenolic compounds in topsoils and the lowest in marine sediments, although both groups of sorbents had similar organic matter content. The 1/n coefficient, reflecting the isotherm nonlinearity, was considerably lower than unity for all compounds in almost all sorbents. The most significant deviation of sorption isotherms from linearity was observed in marine sediments. Only marine sediments showed a linear increase in sorption intensity of all 3 compounds with the increase in sorbent-specific surface area. These results pointed to a different mechanism of sorption in marine and terrestrial sorbents and confirmed that the capacity of sorption was related to amount as well as type and origin of organic matter. PMID- 15657802 TI - Pulsed losses and degradation of aldicarb in a South Florida agricultural watershed. AB - The objectives of these studies were to characterize patterns of movement of aldicarb, aldicarb sulfoxide, and aldicarb sulfone from a typical canalized South Florida watershed and to evaluate aldicarb dissipation in surface water in situ within a citrus grove. Surface water samples were collected daily or every other day from the discharge point for the watershed beginning May 15, 2001, through August 15, 2002. Of 457 samples collected, aldicarb, aldicarb sulfoxide, and aldicarb sulfone were detected in 6, 1, and 13, respectively. Aldicarb was detected from February through May 2002, corresponding to the legal application season of January 1 through April 30 in Florida. Aldicarb concentrations ranged from <0.16 to 4.97 ng ml(-1). A single detection (0.99 ng ml(-1)) of aldicarb sulfoxide occurred in March 2001. The majority of aldicarb sulfone detections occurred during June and July, 2001, after the application season, and ranged from <0.22 to 0.89 ng ml(-1). The half-life for aldicarb in fortified, native surface water ranged from 1.86 to 3.64 days depending on the source of water and the presence of sediments. These results demonstrated the utility of sampling on a frequent basis (compared with monthly or quarterly) for better characterizing pesticide discharges, especially in flashy systems such as canal-drained watersheds within South Florida. PMID- 15657804 TI - Effects of heavy-metal stress on cyanobacterium Anabaena flos-aquae. AB - The influence of two metals, copper and cadmium, was studied on the growth and ultrastructures of cyanobacterium Anabaena flos-aquae grown at three different temperatures: 10 degrees C, 20 degrees C, and 30 degrees C. The highest concentration of chlorophyll a was observed at 20 degrees C and the lowest at 10 degrees C. Both toxic metal ions, Cu(2+) and Cd(2+), inhibited growth of the tested cyanobacterium. Chlorophyll a concentration decreased with the increase of metal concentration. A 50% decrease in the growth of A. flos-aquae population, compared with the control, was reached at 0.61 mg l(-1) cadmium and at 0.35 mg l( 1) copper (at 20 degrees C). Copper at all temperatures tested was proven to be more toxic than cadmium. At 3 mg l(-1), the lysis and distortion of cells was observed; however, after incubation at 9 mg l(-1) cadmium, most of the cells were still intact, and only intrathylakoidal spaces started to appear. Copper caused considerably greater changes in the protein system of A. flos-aquae than did cadmium; in this case, not only phycobilins but also total proteins were destructed. The aim of this study was also to identify the place of metal accumulation and sorption in the tested cyanobacterium. Analysis of the energy dispersion spectra of the characteristic x-ray radiation of trichomes and their sheaths showed that cadmium was completely accumulated in cells but was not found in the sheath. Spectrum of the isolated sheath after treatment with copper exhibited only traces of the metal, but isolated cells without a sheath showed a high peak of copper. PMID- 15657805 TI - Toxicity assessment of pesticide mixtures typical of the Sacramento-San Joaquin Delta using Chironomus tentans. AB - This study examined the effects of nine commonly detected pesticides in the Sacramento-San Joaquin Delta on the aquatic midge Chironomus tentans. Pesticides were chosen from a variety of chemical classes including organophosphate (OP) insecticides as well as triazine, triazinone, and substituted urea herbicides. Both single toxicant and binary mixture bioassays were performed. In addition, midges were pre-exposed to DDE at environmentally relevant concentrations and then challenged by exposing them to chlorpyrifos or diazinon in single-toxicant acute bioassays. Results indicate that most of the binary mixtures elicited additive responses in C. tentans, whereas OP insecticides in combination with various herbicides caused greater-than-additive responses. Pre-exposures with DDE did not have a significant impact on subsequent OP challenges at DDE concentrations of 0.2, 2, and 20 microg/kg. This study represents an important first step in understanding the interactions among various pesticides commonly detected throughout the Delta. PMID- 15657807 TI - Influence of ploidy and metal-metal interactions on the accumulation of Ag, Cd, and Cu in oysters Crassostrea gigas thunberg. AB - The present study was designed to compare the response to contaminants in diploid with triploid specimens of the oyster Crassostrea gigas. The reproduction investment in bivalve molluscs has priority on somatic growth. Thus, genetic sterilization by triploidy induction enables the energy flux to be directed toward somatic growth and glucide storage. Bioaccumulation was examined for Ag (10 microg/L), Cd (10 microg/L), and Cu (30 microg/L) to determine if the response to metals follows similar patterns in diploid (2n) and triploid (3n) groups. The effect of ploidy was also evaluated as a function of dry weight of soft tissue and condition index. Moreover, the reciprocal influence of these metals on their incorporation was studied. The results showed that the major factor governing the degree of metal bioaccumulation by oysters is the nature of the metal introduced in the experimental medium. Thus, the uptake of Cd is proportionally more important than in the case of Ag and even more in Cu. For Cu treated samples, the influence of ploidy on weight and metal body burden (and Cu concentration) was not significant, whereas for Ag and Cd, significant differences according to genetic type were evidenced by higher tissue weight and lower concentrations in triploid than diploid specimens. Metal-metal interactions study especially showed a reciprocal antagonism between Ag and Cu. PMID- 15657806 TI - Assessment of a 2,4,6-trinitrotoluene-contaminated site using Aporrectodea rosea and Eisenia andrei in mesocosms. AB - Polynitro-organic compounds such as 2,4,6-trinitrotoluene (TNT) can be released into the environment from production and processing facilities and military firing ranges as well as through field use and disposal practices. Based on laboratory toxicity data, TNT has lethal (at >/=260 mg TNT/kg dry soil) and sublethal effects (at >/=59 mg TNT/kg dry soil) to the earthworm. However, field studies are needed to relate exposure of organisms to explosives in mixed contaminated soil under field conditions and to define effects-based ecotoxicologic benchmarks for TNT-contaminated soil. In the present study, the lethal and sublethal effects of a 10-day in situ exposure at a TNT-contaminated field site using mesh-bag mesocosms were assessed. In addition to the survival end point, the biomarkers of earthworm exposure and effect-including tissue residues, lysosomal neutral red retention time (NRRT), and total immune activity (TIA)-were measured. Concentrations of TNT in soil mesocosms ranged from 25 to 17,063 mg/kg. Experiments indicated a trend toward decreasing survival of caged Aporrectodea rosea and Eisenia andrei as the concentration of TNT and total nitroaromatic compounds increased. E. andrei tolerated higher concentrations of TNT (up to 4050 mg/kg dry soil) in mesocosms than did indigenous earthworms, who survived only at /=6.3 microg cypermethrin/L), spinal curvatures, and delayed or absence of swim bladder inflation (>/=12.5 microg cypermethrin/L). The exposure of the gametes (stage 0) or animals during earlier embryonic development (stages 9 or 29) was not a critical window for cypermethrin exposure. Although the incidence of edemas in embryos occurred mainly during exposure of these early developmental stages, embryo and larva lethality and the incidence of transient sublethal effects in hatchlings showed that the later exposure window (stage 34) was the most sensitive. The stage 34 group involved advanced organogenetic stages in which the chorion partially degraded before hatching. Our studies reinforced the idea that a combination of morphologic and functional impairment evaluation is a more sensitive response to developmental toxicants than morphologic defects alone. PMID- 15657811 TI - A biopsy procedure for determining filet and predicting whole-fish mercury concentration. AB - Although mercury contamination of fish is a widespread phenomenon, its regional evaluation is hindered by the reluctance of permitting agencies to grant collection permits, problems in securing adequate freezer space, and time to process whole, large fish or filets. We evaluated mercury concentrations in 210 filet biopsies from 65 sites in 12 western states relative to whole-body mercury concentration in the same fish. We found a highly significant relationship (r(2) = 0.96) between biopsy and whole-fish mercury concentrations for 13 piscivorous and nonpiscivorous fish species. We concluded that relative to conventional fish tissue sampling and analysis procedures for whole fish or filets, the biopsy procedure for mercury in fish tissue is nonlethal, less cumbersome, more likely to be permitted by fisheries agencies, and a precise and accurate means for determining both filet and whole-fish mercury concentrations. PMID- 15657812 TI - Zinc and lead poisoning in wild birds in the tri-state mining district (Oklahoma, Kansas, and Missouri). AB - The Tri-State Mining District (Oklahoma, Kansas, and Missouri) is contaminated with Pb, Cd, and Zn from mining, milling and smelting. Metals have been dispersed heterogeneously throughout the District in the form of milled mine waste ("chat"), as flotation tailings and from smelters as aerial deposition or slag. This study was conducted to determine if the habitat has been contaminated to the extent that the assessment populations of wild birds are exposed to toxic concentrations of metals. American robins (Turdus migratorius), northern cardinals (Cardinalis cardinalis), and waterfowl had increased Pb tissue concentrations (p < 0.05) compared with Pb tissue concentrations from reference birds, and the exposure of songbirds to Pb was comparable with that of birds observed at other sites severely contaminated with Pb. Mean activities of the Pb sensitive enzyme delta-aminolevulinic acid dehydratase (ALAD) were decreased by >50% in red blood cells in these birds (p < 0.05). Several birds had tissue concentrations of Pb that have been associated with impaired biological functions and external signs of poisoning. Cadmium was increased in kidneys of songbirds (p < 0.05), but no proximal tubule cell necrosis associated with Cd poisoning was observed. Zinc concentrations in liver and kidney of waterfowl were significantly higher (p < 0.05) than reference values. The increased environmental concentrations of Zn associated with mining in the District accounted for the pancreatitis previously observed in five waterfowl from the District. The District is the first site at which free-flying wild birds have been found to be suffering severe effects of Zn poisoning. PMID- 15657813 TI - Effects of azinphos-methyl on cholinergic responses and general health in zebra finches (Taeniopygia guttata) after previous treatment with p,p'-DDE. AB - Although organochlorine (OC) pesticides were replaced with organophosphates (OPs) in the early 1970s, they continue to persist in orchard environments today. Extensive research has been conducted to determine the effects of currently used OPs on cholinesterase (ChE) activity; however, although OCs continue to be prevalent in areas of previous use, few studies have looked at the toxicity of a combination of residual OC compounds with currently used OP pesticides. The focus of our study was to determine the effects of azinphos-methyl (a common OP used in apple orchards today) on ChE activity and general health in zebra finches (Taeniopygia guttata) previously exposed to p,p'-DDE (a commonly detected metabolite of DDT). The main results of our study were as follows: (1) azinphos methyl alone caused a dose-dependent inhibition of plasma and brain ChE activity; (2) p,p'-DDE in combination with azinphos-methyl did not change azinphos-methyl inhibition of ChE activity; and (3) there were suggestions of immunostimulation in birds dosed 1 year previously to p,p'-DDE and of anemia when p,p'-DDE was combined with azinphos-methyl; however, there was no dose-response for these parameters in birds subsequently dosed with p,p'-DDE. PMID- 15657815 TI - [Has the development of PET come to an end with PET-CT?]. PMID- 15657816 TI - Multi-slice CT for visualization of acute pulmonary embolism: single breath-hold subtraction technique. AB - PURPOSE: The purpose of our preliminary animal study was to evaluate the feasibility of a new subtraction technique for visualization of perfusion defects within the lung parenchyma in segmental and subsegmental pulmonary embolism (PE). MATERIALS AND METHODS: In three healthy pigs, PE were artificially induced by fresh human clot material. Within a single breath-hold, CT angiography (CTA) was performed on a 16-slice multi-slice CT scanner (SOMATOM Sensation 16; Siemens, Forchheim, Germany) before and after intravenous application of 80 mL of contrast medium, followed by a saline chaser. Scan parameters were 120 kV and 100 mAs (eff.), using a collimation of 16 x 1.5 mm and a table speed/rot. of 36 mm (pitch: 1.5; rotation time: 0.5 s). A new 3D subtraction technique was developed, which is based on automated segmentation, non-linear spatial filtering and non rigid registration. Data were analysed using a color-encoded "compound view" of parenchymal enhancement and CTA information displayed in axial, coronal and sagittal orientation. RESULTS: Subtraction was technically feasible in all three data sets. The mean scan time for each series was 4.7 s, interscan delay was 14.7 s, respectively. Therefore, an average breath-hold of approximately 24 s was required for the overall scanning procedure. Downstream of occluded segmental and subsegmental arteries, perfusion defects were clearly assessable, showing lower or missing enhancement compared to normally perfused lung parenchyma. In all pigs, additional peripheral areas with triangular shaped perfusion defects were delineated, considered typical for PE. CONCLUSIONS: Our initial results from the animal model studied show that perfusion imaging of PE is feasible within a single breath-hold. It allows a comprehensive assessment of perfusion deficits as the direct proof of a pulmonary embolus, can be combined with an indirect visual quantification of the density changes in the adjacent lung tissue. PMID- 15657814 TI - Evaluation of skin and respiratory doses and urinary excretion of alkylphosphates in workers exposed to dimethoate during treatment of olive trees. AB - This article describes a study of exposure to dimethoate during spraying of olive trees in Viterbo province in central Italy. Airborne concentrations of dimethoate were in the range 1.5 to 56.7 nmol/m(3). Total skin contamination was in the range 228.4 to 3200.7 nmol/d and averaged 96.0% +/- 3.6% of the total potential dose. Cotton garments afforded less skin protection than waterproof ones, which were in turn associated with higher skin contamination than disposable Tyvek overalls. Total potential doses and estimated absorbed doses, including their maxima, were below the acceptable daily intake of dimethoate, which is 43.6 nmol/kg body weight (b.w.). Urinary excretion of alkylphosphates was significantly higher than in the general population, increasing with exposure and usually showing a peak in the urine sample collected after treatment. Metabolite concentrations were influenced by the type of individual protection used: minimum levels were associated with the closed cabin and maximum levels with absence of any respiratory or hand protection. Urinary alkylphosphates showed a good correlation with estimated absorbed doses and are confirmed as sensitive biologic indicators of exposure to phosphoric esters. PMID- 15657817 TI - [Classification and treatment of endoleaks after endovascular treatment of abdominal aortic aneurysms]. AB - This article describes the classification of endoleaks after endovascular treatment of abdominal aortic aneurysms, thereby summarizing the most important problems of this endovascular technique. The correct classification of endoleaks is a prerequisite for interdisciplinary discussion. It is indispensable for professional reporting of the pathological findings and for the decision making as to the adequate treatment of endoleaks. Irrespective of the types of stent graft and property of the material, five endoleak types are defined in the literature: leakage at the anchor sites (type I); leakage due to collateral arteries (type II); defective stent grafts (type III); leakage due to porosity of the graft material (type IV); and endotension (type V). The causes of endoleaks are discussed and treatment options are reviewed for the diverse pathologic findings. PMID- 15657818 TI - [Visualization and quantification of large bowel motility with functional cine MRI]. AB - PURPOSE: To develop and evaluate a method to visualize and quantify large bowel motility using functional cine MRI. METHODS: Fifteen healthy individuals (8 males, 7 females, 20 to 45 years old) with no history or present symptoms of bowel disorders were enrolled in a functional cine MRI examination at 6 a. m. after a starving phase for at least eight hours before and after oral administration of Senna tea (mild stimulating purgative). Two consecutive sets of repeated measurements of the entire abdomen were performed using a 1.5 T MRI system with coronal T2-weighted HASTE sequences anatomically adjusted to the course of the large bowel. A navigator technique was used for respiratory gating at the level of the right dorsal diaphragm. The changes in diameter (given in cm) were measured at 5 different locations of the ascending (AC), transverse (TC) and descending colon (DC), and assessed as parameters for the bowel motility. RESULTS: The mean values as a statistical measure for large bowel relaxation were determined. Before ingestion of Senna tea, the mean diameter measured 3.41 cm (ascending colon), 3 cm (transverse colon) and 2.67 cm (descending colon). After the ingestion of Senna tea, the mean diameter increased to 3.69 cm (ascending colon) to 3.4 cm (transverse colon) and to 2.9 cm (descending colon). A statistically significant difference was demonstrated with the Wilcoxon test (level of confidence 0.05). For the determination of dynamic increase, the changes of the statistical scatter amplitude to the mean value were expressed as percentage before and after the ingestion of Senna tea. Thereby, an increase in variation and dynamic range was detected for the AC (112.9 %) and DC (100 %), but a decrease in the dynamics for the TC (69 %). CONCLUSION: A non-invasive method for the assessment of bowel motility was developed for the first time. The use of functional cine MRI utilizing a prokinetic stimulus allowed visualisation and quantification of large bowel motility. Further studies have to determine whether this technique is clinically relevant. PMID- 15657819 TI - [Detection of pulmonary nodules with breath-hold magnetic resonance imaging in comparison with computed tomography]. AB - PURPOSE: Evaluation of sensitivity and false positive findings of two fast MRI sequences for the detection of pulmonary nodules in comparison with spiral CT by two independent observers. MATERIALS AND METHODS: All 30 enrolled patients had a spiral CT or MSCT as base line study. MRI was performed with a 1.5 T MR scanner (Sonata, Siemens) using a transverse 3D gradient echo sequence (3D-GRE: TR/TE/flip = 2.9 ms/1.1 ms/5 degrees ) and a half-Fourier single-shot fast spin echo sequence (HASTE: TR/TE/flip = 800/25/150 degrees ) acquired in three planes. A separate analysis for both sequences was carried out prospectively by two independent readers (A and B) with different experience regarding pulmonary MRI. Additionally, a retrospective reading with knowledge of the CT scans was done. Results were calculated for all lesions and for lesions larger than 4 mm. RESULTS: The sensitivities were 73 %, 70 % and 84 % for the 3D-GRE sequence (reader A, reader B, retrospective reading) and 65 %, 68 % and 81 % for the HASTE sequence. For lesions larger than 4 mm, the sensitivities were 93 %, 89 %, 96 % for the 3D-GRE sequence and 85 %, 85 %, 96 % for the HASTE sequence. The rate of false positive findings depended on the reader's experience, but was generally lower for the 3D-GRE sequence with 2 and 16 (reader A and B) false positive nodules compared to 4 and 40 false positive findings for the HASTE sequence. The 3D-GRE sequence was more accurate for both readers (reader A: p = 0.08, reader B: p = 0.00003). CONCLUSION: The sensitivity of MRI for the detection of lung nodules was only acceptable for lesions larger than 4 mm. The 3D-GRE sequence is superior to the HASTE sequence due to the reduced amount of false positive findings with comparable sensitivity. PMID- 15657821 TI - [Left ventricular functional parameters: comparison of 16-slice spiral CT with MRI]. AB - PURPOSE: The quantitative measurement of left ventricular functional parameters using multislice computed tomography (MSCT) with retrospective ECG-gating and comparison of the results with magnetic resonance imaging (MRI). MATERIALS AND METHODS: Thirty-one patients with suspected or known coronary artery disease underwent MSCT angiography with retrospective ECG-gating (Sensation 16, Siemens). Based on the CT data set, short axis reformations of the left ventricle were performed for functional analysis. On a commercially available workstation, end diastolic- (EDV), end-systolic- (ESV), stroke volume (SV), ejection fraction (EF) and myocardial mass (MM) were calculated from MSCT (temporal resolution 105 - 210 ms) data according to the modified Simpson's rule and compared to MRI (1.5 T scanner, temporal resolution 48 ms) using a 2D TrueFISP cine sequence with respiration hold. RESULTS: In all cases, the quality was adequate for both MSCT and MRI. MSCT and MRI had an excellent correlation for EDV (r = 0.86), ESV (r = 0.91), EF (r = 0.87) and MM (r = 0.88), and a good correlation for SV (r = 0.70). The mean difference was 13.2 +/- 21.9 ml for EDV, 8.7 +/- 15.9 ml for ESV, 4.6 +/ 12.3 ml for SV, 1.4 +/- 5.2 % for EF, and 11.9 +/- 13.8 g for MM. However, EDV (p = 0.002), ESV (p = 0.005), SV (p = 0.048), and MM (p < 0.0001) were significantly overestimated with MSCT compared to MRI. For EF, no significant difference between MSCT and MRI was found (p = 0.15). CONCLUSION: For left ventricular functional parameters, MSCT of the heart with retrospective ECG gating showed a high correlation with MRI, which has an important implication when using MSCT for non-invasive cardiac imaging. Despite the high correlation, overestimation of EDV, EVS, SV, and MM with MSCT has to be taken into account when applying this technology in clinical practice. EF was not significantly different between both modalities. PMID- 15657820 TI - [Calcium scoring and coronary angiography performed with multislice spiral CT -- clinical experience]. AB - PURPOSE: To report our clinical experience in calcium scoring and coronary angiography with multislice computer tomography (MSCT). MATERIAL AND METHODS: Cardiac four-row MSCT (Siemens, Volume Zoom, Erlangen, Germany) was performed in 60 patients, comprising 45 patients without known coronary vessel disease (CVD) and falling in an intermediate risk (group I) by calcium scoring, and 15 patients with multivessel disease by the combination of calcium scoring and MSCT coronary angiography (group II). Group I underwent analysis of risk factors as well as patient management in the form of risk profile modulation, indication for invasive selective coronary angiography (SCA) and supplemental myocardial diagnostic evaluation (e. g., SPECT). Agatston and volume scores (Virtuoso, Siemens, Erlangen, Germany) were calculated for calcium scoring. Group II had the calcium scoring and diagnostic accuracy of MSCT in detecting coronary artery stenosis evaluated according to established American Heart Association (AHA) criteria. SCA was used as gold standard. RESULTS: In group I, calcium scoring was positive in 27/45 (60 %) patients, with 14/27 (52 %) consecutively referred to SCA and 2/27 (7 %) to SPECT. SCA revealed significant coronary stenosis (> 60 %) in 5/27 (19 %) patients and SPECT detected myocardial ischemia in 0/2 patients. Patients with obstructive coronary vessel disease showed a significantly increased calcium score of > 300 together with a high risk profile. Patients with negative calcium score showed significantly less risk factors (p < 0.05). The use of calcium scoring in therapeutic procedures in the form of increased modulation of risk factors was documented in 21/45 (47 %) patients. In group II, sensitivity, specificity and diagnostic accuracy for the detection of high grade coronary artery stenosis were 64 %, 99 % and 96 %, and the Agatston and Volume scores were 333 +/- 123 and 334 +/- 136, respectively. CONCLUSION: MSCT can be applied as risk profile module for coronary screening of patients with intermediate risk. As non-invasive alternative for the evaluation of coronary vessel disease, it can be useful in some cases by providing additional information. PMID- 15657822 TI - [Digital mammography with high-resolution storage plates (CR) versus full-field digital mammography (CCD) (DR) for microcalcifications and focal lesions -- a retrospective clinical histologic analysis (n = 102)]. AB - PURPOSE: To determine the diagnostic accuracy of microcalcifications and focal lesions in a retrospective clinical-histological study using high-resolution digital phosphor storage plates (hard copy) and full-field digital mammography (hard copy). MATERIALS AND METHODS: From May 2003 to September 2003, 102 patients underwent digital storage plate mammography (CR), using a mammography unit (Mammomat 3000 N, Siemens) in combination with a high resolution (9 lp/mm) digital storage phosphor plate system (pixel size 50 microm) (Fuji/Siemens). After diagnosis and preoperative wire localization, full-field digital mammography (CCD) (DR) was performed with the same exposure parameters. The full field digital mammography used a CCD-detector (SenoScan) (Fisher Imaging) with a resolution of 10 Ip/mm and a pixel size of 50 microm. Five investigators determined the diagnosis (BI-RADS I - V) retrospectively after the operation from randomly distributed mediolateral views (hard copy reading). These results were correlated with the final histology. RESULTS: The diagnostic accuracy of digital storage plate mammography (CR) and full-field digital mammography (CCD) (DR) was 73 % and 71 % for all findings (n = 102), 73 % and 71 % for microcalcifications (n = 51), and 72 % and 70 % for focal lesions (n = 51). The overall results showed no difference. CONCLUSION: Our findings indicate the equivalence of high resolution digital phosphor storage plate mammography (CR) and full-field digital mammography (CCD) (DR). PMID- 15657823 TI - [CT-guided steroid injection into disc herniation: a causative therapy for lower back pain due to disc herniation]. AB - PURPOSE: To evaluate the therapeutic benefit of CT-guided steroid injection into disc herniations. MATERIALS AND METHODS: In 64 patients, 2.5 mg Dexamethasone was injected into a symptomatic disc herniation under CT-guidance. Conservative treatment 3 to 12 months before injection was unsuccessful. Classified as "complete relief", "strong relief", "mild relief" and "no relief" of pain, the change of discomfort and pain was registered at 14 days, 3 months and 6 months after injection while the conservative regimen continued. RESULTS: At 14 days after injection, 36 patients (56 %) showed complete relief, 5 strong relief (8 %), 12 mild relief (19 %) and 11 no relief (17 %). At 6 months after injection, 25 patients showed complete relief (39 %), 16 strong relief (25 %), and 23 no relief (36 %). One complicating spondylodiscitis was observed. CONCLUSION: CT guided steroid injection into symptomatic disc herniation represents a therapeutic alternative with good therapeutic results. It can be recommended as an alterative to other minimal invasive therapeutic regimens. The disc material acts as reservoir for the drug with good long term depot effect. PMID- 15657824 TI - [Potential advantages of the MRI for the radiofrequency ablation of liver tumors]. AB - PURPOSE: To present first results of radiofrequency ablation of liver tumors using a new MR compatible applicator. MATERIALS AND METHODS: We performed 37 interventions in 20 patients (mean age 58.6 years) with primary intrahepatic malignancies or metastases: colorectal carcinoma n = 6, hepatocellular carcinoma n = 3, pancreatic carcinoma n = 4, sarcoma n = 2, cholangiocellular carcinoma n = 1, carcinoma of the tonsil n = 1, breast carcinoma n = 1, gastric carcinoma n = 1, and gastrointestinal stroma tumor n = 1. Interventions were performed under CT guidance with CT fluoroscopy (n = 32) and under MR-guidance (n = 5) using fast T1 weighted sequences in breath-hold technique. RFA was performed with the RF generator (150 W) under local anesthesia and sedation using MR compatible applicators (Starburst XL, Rita Medical Systems, USA) together with the appropriate Soft Tissue Introducer System. Intra-interventional control was performed with intrahepatically or intralesionally placed introducer system or applicator. MRI was performed with plain breath-triggered T2-weighted turbo spin echo sequences (TSE T2) with fat saturation. RESULTS: All interventions were performed without major events. The mean diameter of induced coagulation was 4.0 (+/- 0.7) cm. Repositioning was necessary in 8 interventions (21 %) after detection of residual tumor on an intra-interventional MRI. After a mean follow up of 6.5 (+/- 1.2) months, the local tumor control rate was 92 %. CONCLUSION: MR compatible RF applicators offer the opportunity for intra-interventional detection of residual tumor during RF ablations by use of sensitive MRI sequences. These procedures may lead to a higher confidence in tumour ablation and may reduce the number of re-interventions and local recurrences of intrahepatic tumors. PMID- 15657825 TI - [Technical feasibility of the implantation of a monorail stent system into the renal arteries without pre-dilatation]. AB - PURPOSE: To evaluate the technical feasibility of the implantation of the monorail RX Herculink system into the renal arteries without pre-dilatation. MATERIALS AND METHODS: Forty-two patients (mean age 71 years) from four centers with a total of 44 renal artery stenoses underwent implantation of the RX Herculink stent. The mean grade of the stenosis was 83.8 %, the mean length 7.5 mm. The stenoses were ostial in 38 cases and in immediate proximity to the ostium in 6 cases. The mean follow-up-period was 57 weeks (24 - 176 weeks). RESULTS: In 42 cases, the implantation was successful without pre-dilatation. In 2 cases, pre dilatation was carried out. In none of the cases, detachment of the stent from the balloon was observed. In one stenosis with a length of 17 mm, implantation of two stents was performed. In 9 cases, post-dilatation with a larger balloon or higher balloon pressure was necessary. Residual stenoses exceeding 30 % were not observed. Two patients developed local bleeding at the puncture site. During the follow-up, restenoses were observed in 5 stents after 26 to 126 weeks, which necessitated a second intervention in 3 cases (PTA in 2 cases, re-stenting in 1 case). The primary patency rate after 6 and 12 months was 0.92 +/- 0.056 according to Kaplan-Meier, the secondary patency rate after 6 and 12 months was 1.0 +/- 0.0. CONCLUSION: Implantation of the RX Herculink stent system into the renal arteries without pre-dilatation is technically feasible and safe. Even without pre-dilatation, the stent-system can be advanced through the stenosis without detachment. The complication rate is low. Our clinical results are comparable to previous studies. PMID- 15657826 TI - [Transarterial embolization for uterine fibroids: clinical success rate and results of magnetic resonance imaging]. AB - PURPOSE: To analyze the clinical success rate and the findings of magnetic resonance imaging (MRI) after uterine artery embolization of symptomatic leiomyomas (fibroids) of the uterus. MATERIALS AND METHODS: This is a prospective single-center case study of 80 consecutively treated patients, followed for 3 - 6 months (group I), 7 - 12 months, (group II), and 13 - 25 months (group III). MRI was used to determine the uterine volume and size of the dominant leiomyoma. Symptoms and causes requiring repeat interventions were analyzed. RESULTS: Significant (p < 0.01) volume reduction of the uterus (median: 34.95 % confidence interval [CI]: 30.41 - 41.76 %) and dominant leiomyoma (median: 52.07 %, CI: 47.71 - 61.57 %) was found. The decrease in uterine volume (I-III: 22.68 %, 33.56 %, 47.93 %) and dominant leiomyoma volume (I-III: 41.86 %, 62.16 %, 73.96 %) progressed with the follow-up time. Bleeding resolved significantly (p < 0.0001) in all three follow-up groups (groups I-III: 92.86 %, 95.23 %, 96.67 %). Furthermore, urinary frequency (groups I-III: 70 %, 75 %, 82.35 %) and sensation of pelvic pressure (groups I-III: 42.86 %, 60 %, 93.75 %) improved, which was statistically significant in group III (p < 0.01). The number of leiomyomas correlated (p < 0.05) with improvement of the bleeding and the pelvic pressure. Repeat therapy was necessary for complications in four patients (5 %) and for therapeutic failure in three patients (3.8 %). Permanent amenorrhea was observed in four patients (5 %) of age 45 years or older. CONCLUSION: Uterine artery embolization of uterine leiomyomas has a high clinical success rate with an acceptable incidence of complications and repeat interventions. PMID- 15657827 TI - [Diffusion-weighted imaging in the diagnostic evaluation of the hydrocephalus in patients with acute or chronic increase in cerebral pressure]. AB - PURPOSE: To investigate whether diffusion-weighted imaging (DWI) in magnetic resonance imaging (MRI) provides additional information about the periventricular white matter for the assessment of hydrocephalus. MATERIALS AND METHODS: Sixteen MRI examinations (11 with acutely increased cerebral pressure, 5 without symptoms) on 15 patients with hydrocephalus (4 patients with communicating hydrocephalus and 11 patients with obstructive hydrocephalus) were analyzed. One symptomatic patient subsequently became asymptomatic. We investigated the "apparent diffusion coefficient" (ADC) in the subcortical and periventricular white matter. The ADCs of the study patients were compared with those of a healthy control group. RESULTS: Symptomatic patients with hydrocephalus, 6/11 showed periventricular edema and a significantly higher ADC values in the periventricular region than in the subcortical white matter. 5/11 symptomatic patients showed significantly higher ADC values even in the absence of periventricular interstitial edema (both groups contained patients with communicating and obstructive hydrocephalus). All 5 asymptomatic patients with hydrocephalus did not have a significantly higher ADC values in the periventricular region. CONCLUSION: In patients with hydrocephalus and acutely increased cerebral pressure, DWI showed a significantly higher ADC values in the periventricular region even without visible interstitial edema on conventional MRI sequences. PMID- 15657828 TI - [Reliability of quantifying vascular white matter brain lesions -- a contribution to reproducible quantitative diagnosis]. AB - PURPOSE: Microangiopathic lesions of the brain tissue correlate with the clinical diagnosis of vascular subcortical dementia. The "experience-based" evaluation is insufficient. Rating scales may contribute to reproducible quantification. MATERIALS AND METHODS: In MRI studies of 10 patients, 9 neuroradiologists quantified vascular white matter lesions (WMLs) at two different points in time for 12 anatomically defined regions with respect to number, size and localization (score). For 9 observers and 10 studies, 90 intra-observer differences were obtained for each of the 12 WML scores. To calculate the inter-observer reliability, rating pairs were formed. Furthermore, 360 differences were computed for each score and rating for 12 anatomically defined WML scores, and the intraclass correlation (ICC) was calculated as a measure of agreement (reliability). RESULTS: As to the intra-observer reliability, the median of the differences was 1.5 for the entire brain as opposed to 0 for defined brain regions. The corresponding values for the inter-observer reliability were 3 and 1, respectively. The mean intra-class correlation coefficient for the 10 studies was 0.88, whereas the mean interclass correlation concerning the inter-observer reliability was 0.70, with the first and second rating being averaged. The rating of each study took about 6 minutes. CONCLUSION: The rating scale with high intra- and inter-observer reliability can dependably quantify WMLs and correlates with the clinical diagnosis of vascular dementia. Using a reliable rating scale, the diagnostic distinction of age-associated physiological vs. pathological size of the WML can make a contribution to the reproducible quantifiable diagnostic evaluation of vascular brain tissue lesions within the framework of dementia diagnostics. PMID- 15657830 TI - [Referrer satisfaction as a quality criterion: developing an questionnaire for measuring the quality of services provided by a radiology department]. AB - PURPOSE: To develop a questionnaire for measuring referring physician satisfaction and to conduct a pilot study in which this questionnaire is given to all physicians referring patients to the authors' radiology department. MATERIALS AND METHODS: After qualitative pre-testing and adjustment of the prototype questionnaire, data were collected using the finalized standardized questionnaire comprising 29 indicators rated on a 4-point ordinal scale mailed with a personalized cover letter to the total referring physician population of a radiology department (n = 727). The replies, rated 1 - 4, were entered into a data entry mask for statistical analysis. RESULTS: The response rate was 33.8 %. The indicators with the highest satisfaction rating were the range of examinations offered ("very satisfied": 79.3 % mean 3.79), the quality of the technical equipment used for MRI and CT (79.3 %, 3.79) and mammography (82.5 %, 3.82), and the quality of the images yielded by these procedures (74.5 %, 3.73 and 82.2 %, 3.83). Dissatisfaction was relatively high with the indicators "time to receipt of the written report" (28.3 % "not very satisfied" or "not at all satisfied", mean 2.97), "time to receipt of the X-ray images" (18.2 %, 3.07) and "availability of previous findings" (20.9 %, 3.05); satisfaction was higher among external referring physicians (p < 0.05). Physicians rated the importance of these three indicators as relatively high ("very important": 62.4 %, 54.3 % and 49.6 %). Other indicators showing a similar level of dissatisfaction were "car parking availability" (24.1 %, 3.01), "patient waiting time" (27.4 %, 2.87) and "patient environment" (21.2 %, 2.99), although these factors were rated as less important ("very important": 33.0 %, 33.7 % and 40.4 %). CONCLUSION: This questionnaire constitutes a standardized validated instrument for assessing referring physician satisfaction with a radiology department. The data from this pilot study highlight areas for potential improvement. Deployment of such a questionnaire in different radiology departments could serve to establish best practice benchmarks. PMID- 15657829 TI - [Pseudonormalization of diffusion weighted images: magnetic resonance imaging in an animal model (C6-glioma)]. AB - PURPOSE: Previous studies on intracranial tumors indicate that a high apparent diffusion coefficient (ADC) is due to low cellularity and that lower values indicate a dense, highly cellular tumor. Diffusion is affected by three major factors: cell density, existence and distribution of vasogenic edema, and hypoxic tissue. Therefore, we studied the characteristics of diffusion-weighted magnetic resonance imaging (DWI) in a rat brain C6 glioma during tumor progression. MATERIALS AND METHODS: In male Wistar rats, C6 gliomas were implanted in the caudoputamen. At day 9, 11, 13 and 15 after tumor inoculation, conventional DWI was performed on a 2.35 Tesla small bore MRI unit (Biospec 24/40, BRUKER Medizintechnik, Ettlingen, Germany). RESULTS: On conventional T2-weighted and contrast-enhanced T1-weighted images, all tumors could well be delineated from the surrounding brain tissue and showed significant progression. On DWI, the tumors were isointense or slightly hypointense compared to the surrounding brain. On the ADC maps, the tumors could be well visualized due to increasing ADC values from day 9 to 15. The mean ADC of brain tumor tissue was 0.76 +/- 0.4 x 10 ( - 3) mm (2)/s at day 9 and 0.91 +/- 0.03 x 10 ( - 3) mm (2)/s at day 15. The mean ADC of the normal contralateral caudoputamen was 0.59 +/- 0.007 x 10 ( - 3)mm (2)/s. CONCLUSION: T2 prolongation and increased water diffusion can be balanced on DWI in C6 gliomas, resulting in isointensity on DWI (T2 shine-through washout phenomenon). ADC maps are indispensable for the correct interpretation of tumor tissue diffusion behavior. PMID- 15657831 TI - [A solution to add digital signatures to medical images according to the DICOM standard: embedded systems]. AB - Radiology departments often underestimate the importance of protecting medical data during transmission, including the precautions taken to ensure data protection. In teleradiology, transmitted as well as stored patient data have to be signed digitally according to the currently valid regulation (Rontgenverordnung, RoV). The DICOM standard facilitates a digital signature. So far, medical image manufacturers only announced to support this security feature. We introduce a solution that extends the feature of digital signing to older modalities. PMID- 15657832 TI - [Value of postmortem computed tomography in comparison to autopsy]. AB - PURPOSE: To assess the diagnostic value of postmortem computed tomography (CT) in comparison to autopsy. MATERIALS AND METHODS: Twenty-seven cadavers were examined by sequential cranial CT and helical CT through the neck, thorax and abdomen and subsequently underwent an autopsy with histomorphologic examination of the pathologic specimens. The findings of CT, autopsy and histology were registered and compared by three radiologists and one specialist for forensic medicine, using a data entry form. RESULTS: In 19 of 27 cases, the findings explaining the cause of death were concordant for CT and autopsy. Intracranial, intraspinal and intracardiac gas accumulations (n = 12) were registered by CT alone. The detection of skull fractures was equal for both methods (n = 3). CT showed diagnostic problems in the assessment of pneumonic infiltrations (n = 16) and pulmonary edema (n = 21). CONCLUSION: CT is a useful and complementary method to autopsy. PMID- 15657833 TI - [Aneurysm of the petrous part of the A. carotis interna with recurrent life threatening epistaxis]. PMID- 15657834 TI - [Cystic impressive space-occupying lesion dorsal to the pancreas head]. PMID- 15657835 TI - [Interventional recanalization of renal artery stenosis as a complication of endovascular therapy of aortic aneurysm]. PMID- 15657836 TI - [Chronic Marchiafava-Bignami syndrome in alcoholism of many years]. PMID- 15657837 TI - [Lemierre syndrome]. PMID- 15657838 TI - [Head and neck tumors -- PET-CT is superior to PET alone in the diagnosis and therapy planning]. PMID- 15657839 TI - [Rectal carcinoma -- water in the rectum makes ultrasonography easier]. PMID- 15657841 TI - [Portal hypertension -- portosystemic shunts prolong life]. PMID- 15657840 TI - [Bronchial carcinoma: how are false positive results to be avoided]. PMID- 15657843 TI - [Forecasting value -- spiral CT and endosonography supplement each other in the staging of esophageal carcinoma]. PMID- 15657842 TI - [Computed tomography -- Xenon CT in the diagnosis of liver tumors]. PMID- 15657844 TI - [Periaortic hematoma -- blunt thoracic trauma in CT]. PMID- 15657845 TI - [Intestinal cancer screening -- omissions in virtual colonoscopy]. PMID- 15657846 TI - [Digital radiographic image presentation -- automatic or semiautomatic processing?]. PMID- 15657851 TI - Endoscopic ultrasonography. AB - This paper covers the literature published between June 2003 and August 2004 on endoscopic ultrasonography (EUS) and EUS-guided fine-needle aspiration (FNA). During this period, more than 120 original studies were published on the topic, not to mention editorials and case reports--indirect evidence of the ever increasing importance of EUS in everyday clinical practice. However, due to limitations of space, this review will focus only on a few articles selected for their importance and originality. The studies presented deal either with the outcome of EUS--i. e., its real importance in affecting the choices physicians are faced with in their clinical work--or definitively clarify certain technical issues that have often been a matter of debate at EUS meetings. Finally, a few papers describing new potential diagnostic or interventional applications of EUS are presented. Many other relevant papers in addition to those discussed in detail are cited in the references. PMID- 15657852 TI - Gastroesophageal reflux disease and Barrett's esophagus. AB - Several articles have been published during the last year that may affect the management of patients with gastroesophageal reflux disease (GERD) and/or Barrett's esophagus in the near future. A new method of measuring esophageal pH has been introduced that does not require an indwelling transnasal catheter and may allow a more physiological assessment of esophageal acid exposure. Several articles discussed the use of endoscopic antireflux procedures, and a sham controlled randomized study was published concerning the Stretta procedure. A long-term follow-up study and a decision analysis study have again fueled the debate concerning the relevance of surveillance of Barrett's patients, whereas other studies focused on techniques that may improve the detection of specialized intestinal metaplasia and dysplasia within the Barrett's segment. Finally, several studies have reported promising results with the endoscopic treatment of Barrett's metaplasia and early neoplasia using ablation techniques or endoscopic resection modalities. This review summarizes the most important articles in the field of GERD and Barrett's esophagus that have been published in peer-reviewed journals during the last year that are relevant to the practicing endoscopist. PMID- 15657853 TI - Treatment of oesophagogastric tumours. PMID- 15657854 TI - Diagnosis of esophagogastric tumors. AB - With regard to esophageal tumors, important reports on several topics have been published recently. 1) The place of endoscopic ultrasonography (EUS) as the best locoregional staging technique for cancer of the esophagus has been further consolidated. The addition of fine-needle aspiration makes EUS more sensitive than computed tomography (CT) and more accurate than CT or EUS alone for nodal staging. 2) High-resolution endoscopy with chromoendoscopy has been found to be very effective for mucosal lesions, but not for submucosal lesions. In combination with EUS, the sensitivity for submucosal tumors increases up to 60 %. 3) Autofluorescence-guided biopsy has been reported to be a good tool for detecting high-grade dysplasia. A narrow-band imaging system improved the overall accuracy for depth of invasion. 4) The incidence of hypopharyngeal cancer increases after resection for esophageal carcinoma. Patients with a scattered staining pattern after application of Lugol's solution are more prone to develop upper lesions. 5) Fluorescence imaging makes it possible to detect low-grade intraepithelial neoplasia in Barrett's mucosa, with fewer biopsies. 6) Patients with Barrett's esophagus with a length of over 3 cm had a significantly greater prevalence of dysplasia in comparison with those in the whom the Barrett's segment was shorter than 3 cm (23 % vs. 9 %, P = 0.0001). With regard to gastric tumors, 1) Helicobacter pylori eradication can significantly reduce the development of gastric cancer, but only in patients without precancerous lesions. 2) Intestinal metaplasia types II and III have been shown to have a higher rate of progression to low-grade dysplasia than type I. 3) With regard to screening in asymptomatic individuals, serum pepsinogen may represent an alternative to conventional fluoroscopy methods. 4) In patients who have undergone esophagectomy for esophageal cancer, annual follow-up endoscopies are vital for detecting early secondary gastric cancer and ulcerations in which curative treatment is possible. 5) High-resolution endoscopy allows more precise diagnosis of early gastric cancer. The presence of irregular minute vessels and variations in vessel caliber were found to be specific of early gastric cancer. The small regular pattern of sulci and ridges was observed significantly more frequently in differentiated carcinoma than in undifferentiated carcinoma. 6) Infrared-ray electronic endoscopy combined with indocyanine green injection appears to be effective in detecting sentinel nodes that contain metastases in patients with gastric cancer. 7) Gastric adenocarcinoma was found to show specific changes in the fluorescence spectra emitted, in comparison with normal gastric mucosa. However, there was wide variation in the emitted autofluorescence spectra in gastric cancer with signet-ring cells in comparison with normal mucosa. PMID- 15657855 TI - Diagnostic laparoscopy. AB - Each diagnostic procedure needs to be viewed in the context of all the other available diagnostic tools, and therefore has to be reevaluated periodically. This is also true of diagnostic laparoscopy, whether performed by gastroenterologists in patients under sedoanalgesia or by surgeons in patients under general anesthesia. Publications during the previous year have shed light on many important issues. Despite its greater expense, fluorescence laparoscopy may have advantages over white-light laparoscopy in the diagnosis of small tumor seeding. The unchanged superiority of laparoscopy over other imaging modalities for detecting small superficial liver metastases and peritoneal seeding has been clearly demonstrated. Whether cytological investigation of peritoneal washing can add relevant information to laparoscopy remains a matter of debate. Diagnostic laparoscopy before surgery has proved to be effective in many fields, including traumatology. The use of diagnostic laparoscopy in certain clinical circumstances, such as the early postoperative period after major cardiac surgery using extracorporeal circulation, appears promising. A transgastric approach to the peritoneal cavity for diagnostic and therapeutic purposes, the feasibility of which has been demonstrated in a porcine model, is an exciting innovation with as yet unforeseeable implications. PMID- 15657856 TI - The future of surgical endoscopy. PMID- 15657857 TI - Inflammatory bowel disease. AB - This review summarizes important publications that have appeared during the last year dealing with imaging techniques and endoscopy, as well as the management of low-grade dysplasia and stenosis in inflammatory bowel disease. Magnetic resonance enteroclysis and capsule endoscopy are currently emerging as new imaging techniques for the small bowel in Crohn's disease. While magnetic resonance enteroclysis is, at least in Europe, increasingly being used as a reference method, the value of capsule endoscopy for the management of inflammatory bowel disease is still being evaluated. Chromoendoscopy is being studied in patients with long-lasting ulcerative colitis and may be a promising and sensitive technique for the diagnosis of dysplasia. However, there are conflicting data regarding the appropriate management when low-grade dysplasia is diagnosed in patients with ulcerative colitis. Endoscopic dilation can often be successfully carried out in cases of intestinal stenosis. The results of long term follow-up studies indicate that several dilation procedures are often necessary and that a relatively high percentage of patients still have to undergo surgery. PMID- 15657858 TI - Unsedated transnasal percutaneous endoscopic gastrostomy placement in selected patients. AB - BACKGROUND AND STUDY AIMS: The placement of a percutaneous endoscopic gastrostomy (PEG) is the procedure of choice for the long-term management of dysphagic patients with neurological disease or with trauma or tumors of the head and neck. It is not always possible to perform conventional upper gastrointestinal endoscopy in such patients due to stenosis and/or occlusion of the mouth or pharynx and/or partial or complete trismus. The aim of this study was to show whether transnasal esophagogastroduodenoscopy (EGD) offers a feasible and effective alternative method for PEG placement in these selected patients. PATIENTS AND METHODS: PEG placement was required for 155 patients at our institution during a 27-month period. In 12 patients oral access of an endoscope into the esophagus was not possible. Unsedated transnasal EGD (T-EGD) was then performed using an ultrathin video gastroscope, which had a distal-end diameter of 5.9 mm. A 16-Fr polyurethane PEG tube with a conical, flexible, soft distal end and a collapsible bumper was used in all cases. The Gauderer-Ponsky pull technique was used for PEG placement. RESULTS: T-EGD and perendoscopic transnasal placement of a PEG tube was successfully performed in all 12 patients. No patient required sedation during the procedure. No immediate or late-onset procedure related complications occurred in any of the 12 patients. CONCLUSIONS: In some dysphagic patients in whom the oral route is not accessible with a standard endoscope, a transnasal endoscopic approach allows the placement of a PEG tube. In these selected patients this technique has been shown to be safe and effective and does not require the use of sedation. PMID- 15657859 TI - A novel diagnostic method for evaluation of vascular lesions in the digestive tract using infrared fluorescence endoscopy. AB - BACKGROUND AND STUDY AIMS: We have developed an infrared fluorescence endoscope to evaluate gastrointestinal vascular lesions. Infrared endoscopy (IRE) after intravenous administration of indocyanine green (ICG) is used at present to examine vascular lesions such as esophageal varices. However, no previous study has compared the sensitivity of infrared fluorescence endoscopy (IRFE) with that of IRE. In this study, we compared the usefulness of IRFE and IRE. PATIENTS AND METHODS: For IRFE we used an infrared endoscope equipped with excitation and barrier filters and an intensified charge-coupled device camera. In preliminary experiments, the observable tissue depth was assessed by wrapping increasing numbers of layers of commercially available pork around a syringe containing a uniform concentration of ICG or by changing the concentration of ICG in a syringe covered by a piece of pork of uniform thickness. In the clinical part of the study, ICG was administered intravenously at different concentrations to patients with esophageal varices and the resulting infrared fluorescent images were evaluated. RESULTS: The preliminary experiments revealed that the depth of tissue that could be visualized was significantly greater in IRFE than it was in IRE (11.2 mm in IRFE vs. approximately 3.2 mm in IRE). Clear infrared fluorescence was obtained by IRFE at lower concentrations of ICG than the concentrations required to obtain clear images using IRE. In the clinical part of the study, clear infrared fluorescence was observed in a region where esophageal varices had been detected by conventional endoscopy when ICG was administered in doses of 0.005 mg/kg to 0.01 mg/kg, which was lower than the doses used in IRE. CONCLUSIONS: Compared with conventional IRE, IRFE facilitated the observation of deeper layers, and esophageal varices were observed by IRFE following the intravenous administration of a markedly reduced dose of ICG. IRFE, in combining the characteristics of reflected infrared light and fluorescence, may be a useful novel procedure in the diagnosis of vascular lesions in the gastrointestinal tract. PMID- 15657860 TI - Precut papillotomy versus persistence in difficult biliary cannulation: a prospective randomized trial. AB - BACKGROUND AND STUDY AIMS: Failed biliary cannulation occurs in up to 10% of patients undergoing ERCP. There is some controversy as to the safety and efficacy of using precut techniques to achieve biliary cannulation in difficult cases. To date, no randomized trial has compared the success and complication rates of precut with the rates for persistence when biliary cannulation is difficult. The aim of this study was to compare the success rates and complication rates of precut with the success rates and complication rates of persistence in cases of difficult biliary cannulation. PATIENTS AND METHODS: Patients without prior sphincterotomy who required biliary cannulation were screened. A "difficult biliary cannulation" was arbitrarily defined as failed cannulation after 12 minutes. These patients were then randomized to continue treatment by needle knife cut over the roof of the papilla or by persistence with a non-wire-guided, single-lumen papillotome. "Primary" success was defined as deep cannulation within 15 minutes of randomization. Primary and final success rates and complication rates within 30 days after ERCP were compared. RESULTS: Over a 38 month period a total of 642 patients were screened. Patients in whom biliary cannulation was successful within a time period of 12 minutes or less formed the reference group (n = 580). The remainder of the patients were randomly assigned to the "precut" arm (n = 32) or to the "persistence" arm (n = 30). Primary success rates and complication rates were similar in the precut and persistence arms (75% and 4% respectively for the precut arm vs. 73% and 9% for the persistence arm). The final successful cannulation rate in the entire group of 642 patients was 99.5%. CONCLUSIONS: In experienced hands, precut papillotomy and persistence in cannulation are equally effective in cases of difficult cannulation, with a similar complication rate. PMID- 15657861 TI - Push-and-pull enteroscopy using the double-balloon technique: method of assessing depth of insertion and training of the enteroscopy technique using the Erlangen Endo-Trainer. AB - BACKGROUND AND STUDY AIMS: This study was conducted to test a method of measuring the depth of insertion into the small bowel during push-and-pull enteroscopy using the Erlangen Endo-Trainer. Furthermore, the Erlangen Endo-Trainer model for training in the new method of push-and-pull enteroscopy using the double-balloon technique was also evaluated. MATERIALS AND METHODS: Specially prepared packages of porcine upper visceral organs were used, including the esophagus, stomach, duodenum, and small bowel, implanted into the Erlangen Endo-Trainer. In the first step of this study, all of the modifications needed to obtain a model useful for training in the new enteroscopy technique were tested, including different lengths of small bowel. In the next step, the Erlangen Endo-Trainer was used to evaluate a special method of measuring the depth of insertion during push-and pull enteroscopy by comparing estimated insertion depths of 100 cm and 200 cm, marked on the porcine small bowel, with endoscopic insertion depths determined afterwards by measuring the length of the small bowel from the pylorus to the mark on the porcine small bowel. Additionally, the Endotrainer was used to demonstrate the principle of double-balloon enteroscopy and to provide training in this new enteroscopy technique with a lifelike simulation. RESULTS: The modified Erlangen Endo-Trainer proved its value for demonstrating the principle of push-and-pull enteroscopy and for providing training in this enteroscopic technique as well as for testing the measurement method. The evaluation of the measurement method showed that the estimation of the insertion depths was accurate, with a mean deviation of less than 10 %. A total of 13 workshops (seven national and six international), including a total of 97 participants, were carried out between January and August 2004. Under the supervision of one of the authors, pairs of trainees were able to reach the ileal valve or the end of the small bowel from the oral route. CONCLUSIONS: The modified Erlangen Endo-Trainer is useful for training in the push-and-pull enteroscopy technique. The new specially developed method of measuring the depth of insertion during push-and pull enteroscopy seems to be valid. PMID- 15657862 TI - On writing (6): Writing a book. PMID- 15657863 TI - Enteral stents in the gastric outlet and duodenum. PMID- 15657864 TI - Duodenal obstruction by gallstones (Bouveret's syndrome): a review of the literature. AB - Bouveret's syndrome, first described in 1896, is gastric obstruction by a gallstone following a cholecystoduodenal fistula. Endoscopy is the mainstay of diagnosis, but radiographic examination such as upper gastrointestinal contrast series and abdominal radiography can also contribute to the diagnosis. Diagnosis by computed tomography and ultrasonography has also been described. The syndrome can be diagnosed and treated endoscopically, with stone extraction or mechanical lithotripsy. Extracorporeal shockwave lithotripsy has also been used successfully. Surgery is required in over 90% of cases, with mortality rates ranging from 19% to 24%. One-stage and two-stage procedures have been described, including enterolithotomy, cholecystectomy, and fistula repair, no convincing data are available to show which of these two approaches provides a better outcome. Although the condition is rare, Bouveret's syndrome should be considered in elderly patients with a history of chronic cholecystitis who present with pain, vomiting or haematemesis. PMID- 15657865 TI - Successful endoscopic management of iatrogenic mediastinal infection and subsequent esophagomediastinal fistula, following endosonographically guided fine needle aspiration biopsy. AB - Complications following endoscopic ultrasound-guided fine-needle aspiration (EUS FNA) biopsy are rare. A 75-year-old man underwent EUS-FNA biopsy of an enlarged mediastinal lymph, which histologic investigation revealed to be a metastasis of a hepatocellular carcinoma. The patient developed the postinterventional complication of suppurative infection within the mediastinum. Under EUS guidance, a pigtail catheter and a soft tube were inserted to respectively drain and rinse the mediastinal lesion for 8 days. The remaining esophagomediastinal fistula was closed by gathering the fistula margins, using band ligations and an Endoloop. The fistula healed with no further complaints or dysphagia. Infection is a possible complication of endoluminal FNA biopsy. An endoscopically guided therapeutic approach can be favored as the initial treatment of choice and as a reasonable alternative that avoids surgical intervention. PMID- 15657866 TI - Ischemic duodenitis. PMID- 15657867 TI - Ingestion of an unusual foreign body. PMID- 15657868 TI - Arterioportal fistula complicating endoscopic retrograde cholangiography. PMID- 15657869 TI - Intestinal bleeding associated with antiphospholipid antibody syndrome. PMID- 15657870 TI - A janus-headed polyp: adenoma and carcinoma with a single stalk. PMID- 15657871 TI - Guidelines of the French Society for Digestive Endoscopy (SFED). Role of endoscopy in microscopic colitis. PMID- 15657872 TI - Association testing in a linked region using large pedigrees. AB - This report describes computer implementation of a scheme for joint linkage and association analysis. The model implemented in the computer package Mendel estimates both recombination and linkage-disequilibrium parameters and conducts likelihood-ratio tests for (1) linkage alone, (2) linkage and association simultaneously, and (3) association in the presence of linkage. Application of the method to data from Finnish pedigrees with familial combined hyperlipidemia illustrates its potential for identification of associated SNP haplotypes in the presence of linkage. For the test results to be valid, good estimates of haplotype frequencies must be used in the analysis. PMID- 15657873 TI - PHOX2B genotype allows for prediction of tumor risk in congenital central hypoventilation syndrome. AB - The Phox2b gene is necessary for autonomic nervous-system development. Phox2b-/- mice die in utero with absent autonomic nervous system circuits, since autonomic nervous system neurons either fail to form or degenerate. We first identified the Phox2b human ortholog, PHOX2B, as the gene underlying congenital central hypoventilation syndrome (CCHS, or Ondine curse), with an autosomal dominant mode of inheritance and de novo mutation at the first generation. We have subsequently shown that heterozygous mutations of PHOX2B may account for several combined or isolated disorders of autonomic nervous-system development--namely, tumors of the sympathetic nervous system (TSNS), such as neuroblastoma and late-onset central hypoventilation syndrome. Here, we report the clinical and molecular assessments of a cohort of 188 probands with CCHS, either isolated or associated with Hirschsprung disease and/or TSNS. The mutation-detection rate was 92.6% (174/188) in our series, and the most prevalent mutation was an in-frame duplication leading to an expansion of +5 to +13 alanines in the 20-alanine stretch at the carboxy terminal of the protein. Such findings suggest PHOX2B mutation screening as a simple and reliable tool for the diagnosis of CCHS, independent of the clinically variable phenotype. In addition, somatic mosaicism was detected in 4.5% of parents. Most interestingly, analysis of genotype-phenotype interactions strongly supports the contention that patients with CCHS who develop malignant TSNS will harbor either a missense or a frameshift heterozygous mutation of the PHOX2B gene. These data further highlight the link between congenital malformations and tumor predisposition when a master gene in development is mutated. PMID- 15657874 TI - A 4-bp deletion in the Birt-Hogg-Dube gene (FLCN) causes dominantly inherited spontaneous pneumothorax. AB - Primary spontaneous pneumothorax (PSP), a condition in which air enters the pleural space and causes secondary lung collapse, is mostly sporadic but also occurs in families. The precise etiology of PSP remains unknown, although it is associated with emphysemalike changes (bullae) in the lungs of almost all patients. We describe the results of a genetic study of a large Finnish family with a dominantly inherited tendency to PSP. A genomewide scan suggested linkage to chromosome 17p11. Screening of the best candidate gene, FLCN, revealed a 4-bp deletion in the first coding exon, which causes a frameshift that predicts a protein truncation 50 missense amino acids downstream. All carriers of the deletion had bullous lung lesions. Mutations in FLCN are also responsible for Birt-Hogg-Dube (BHD) syndrome (a dominantly inherited disease characterized by benign skin tumors, PSP, and diverse types of renal cancer) and, rarely, are detected in sporadic renal and colorectal tumors. Unlike other FLCN mutations, the exon 4 deletion seems to be associated with bullous lung changes only with 100% penetrance. These results suggest that changes in FLCN may have an important role in the development of PSP and, more importantly, of emphysema, a chronic pulmonary disease that often leads to formation of bullous lesions and lowered pulmonary function. Additionally, given the strong association of PSP and BHD, the connection between these conditions needs to be investigated further, particularly in patients with familial PSP, who may be at a greater risk of developing renal cancer. PMID- 15657875 TI - Polymorphisms in the tyrosine kinase 2 and interferon regulatory factor 5 genes are associated with systemic lupus erythematosus. AB - Systemic lupus erythematosus (SLE) is a complex systemic autoimmune disease caused by both genetic and environmental factors. Genome scans in families with SLE point to multiple potential chromosomal regions that harbor SLE susceptibility genes, and association studies in different populations have suggested several susceptibility alleles for SLE. Increased production of type I interferon (IFN) and expression of IFN-inducible genes is commonly observed in SLE and may be pivotal in the molecular pathogenesis of the disease. We analyzed 44 single-nucleotide polymorphisms (SNPs) in 13 genes from the type I IFN pathway in 679 Swedish, Finnish, and Icelandic patients with SLE, in 798 unaffected family members, and in 438 unrelated control individuals for joint linkage and association with SLE. In two of the genes--the tyrosine kinase 2 (TYK2) and IFN regulatory factor 5 (IRF5) genes--we identified SNPs that displayed strong signals in joint analysis of linkage and association (unadjusted P<10(-7)) with SLE. TYK2 binds to the type I IFN receptor complex and IRF5 is a regulator of type I IFN gene expression. Thus, our results support a disease mechanism in SLE that involves key components of the type I IFN system. PMID- 15657877 TI - Screw-sense inversion characteristic of alpha-helical poly(beta-p-chlorobenzyl L aspartate) and comparison with other related polyaspartates. AB - This is one of a series of studies on the reversal of the helix sense of polyaspartates originated from the pioneering work of Goodman and his associates in 1960s. Poly(beta-p-chlorobenzyl L-aspartate) (PClBLA) is one of the well studied polyaspartate derivatives in both solution and the solid state. The chemical structure of PClBLA differs from those of poly(beta-benzyl L-aspartate) (PBLA) and poly(beta-phenethyl L-aspartate) (PPLA) only at the terminal of the relatively long side chain. PBLA takes a left-handed form (L) in conventional helicoidal solvents and does not exhibit any screw-sense inversion. In contrast to PBLA, both PClBLA and PPLA form a right-handed helix (R) in chlorinated alkane solvents and exhibits a reversal of alpha-helix sense at higher temperatures. Yet the transition behaviors in the presence of denaturant acid are quite different between these two polymers. While PPLA exhibits transitions such as R --> L --> coil by lowering temperature, PClBLA directly goes into the coil state without showing the reentrant L form. The cause of these phenomenological differences among these polymers has been investigated by constructing the phase diagram. PMID- 15657876 TI - Assessing adenoviral hammerhead ribozyme and small hairpin RNA cassettes in neurons: inhibition of endogenous caspase-3 activity and protection from apoptotic cell death. AB - Antisense technology, including ribozyme and small interfering RNA, is being developed to mediate the down-regulation of specific intracellular genes. It was observed in this study that both antiluciferase ribozymes and short hairpin RNAs (shRNAs) could significantly reduce the activity of exogenously expressed luciferase in primary hippocampal neurons in a viral titer-dependent manner. shRNAs were more effective gene-silencing agents than ribozymes, although they exhibited some nonspecific gene-silencing effects at high viral titers. We also attempted to increase ribozyme efficacy by using a woodchuck hepatitis posttranscriptional regulatory element (WPRE) in the ribozyme expression cassette. The results showed that adenoviral vectors encoding specific ribozymes could silence the cellular expression of luciferase and endogenous procaspase-3 significantly. Furthermore, the antiprocaspase-3 ribozyme was shown to inhibit staurosporine-mediated cell death. The addition of a WPRE did not, however, increase or decrease ribozyme activity. As far as we are aware, this is the first example of adenovirally mediated delivery of hammerhead ribozymes being used to manipulate gene expression in primary neurons. The results therefore suggest that hammerhead ribozymes may be useful tools for studying neuronal gene function and have potential as therapeutic agents to treat CNS diseases. PMID- 15657878 TI - Nanomaterials and chip-based nanostructures for capillary electrophoretic separations of DNA. AB - Capillary electrophoresis (CE) and microchip capillary electrophoresis (MCE) using polymer solutions are two of the most powerful techniques for the analysis of DNA. Problems, such as the difficulty of filling polymer solution to small separation channels, recovering DNA, and narrow separation size ranges, have put a pressure on developing new techniques for DNA analysis. In this review, we deal with DNA separation using chip-based nanostructures and nanomaterials in CE and MCE. On the basis of the dependence of the mobility of DNA molecules on the size and shape of nanostructures, several unique chip-based devices have been developed for the separation of DNA, particularly for long DNA molecules. Unlike conventional CE and MCE methods, sieving matrices are not required when using nanostructures. Filling extremely low-viscosity nanomaterials in the presence and absence of polymer solutions to small separation channels is an alternative for the separations of DNA from several base pairs (bp) to tens kbp. The advantages and shortages of the use of nanostructured devices and nanomaterials for DNA separation are carefully addressed with respect to speed, resolution, reproducibility, costs, and operation. PMID- 15657879 TI - The structure of antimicrobial pexiganan peptide in solution probed by Fourier transform infrared absorption, vibrational circular dichroism, and electronic circular dichroism spectroscopy. AB - Pexiganan (Gly-Ile-Gly-Lys-Phe-Leu-Lys-Lys-Ala-Lys-Lys-Phe-Gly-Lys-Ala-Phe-Val Lys-Ile-Leu-Lys-Lys), a 22 amino acid peptide, is an analogue of the magainin family of antimicrobial peptides present in the skin of the African clawed frog. Conformational analysis of pexiganan was carried out in different solvent environments for the first time. Organic solvents, trifluoroethanol (TFE) and methanol, were used to study the secondary structural preferences of this peptide in the membrane-mimicking environments. In addition, aqueous (D2O) and dimethyl sulfoxide (DMSO) solutions were also investigated to study the role of hydrogen bonding involved in the secondary structure formation. Fourier transform infrared absorption, vibrational circular dichroism (VCD), and electronic circular dichroism (ECD) measurements were carried out under the same conditions to ascertain the conformational assignments in different solvents. All these spectroscopic measurements suggest that the pexiganan peptide has the tendency to adopt different structures in different environments. Pexiganan appears to adopt an alpha-helical conformation in TFE, a sheet-stabilized beta-turn structure in methanol, a random coil with beta-turn structure in D2O, and a solvated beta-turn structure in DMSO. PMID- 15657880 TI - CD spectral study of Dnp derivatives of amino acids and peptides for their configurational and conformational analysis. AB - Rules relating the stereochemistry of N-Dnp (Dnp: 2,4-dinitrophenyl) derivatives of alpha-amino acids and peptides and the sign of the Cotton effects at the longest wavelength band (ca. 400 nm) are surveyed. Some new data and insights concerning the CD spectra of Dnp-alpha-amino acids are included: i.e., the spectra of Dnp derivatives as the composite of the corresponding o-nitrophenyl and p-nitrophenyl derivatives; the crystal structure of Dnp-I-phenylalanine and its solid-state CD spectra; the CD spectra of Dnp-alpha-amino acids containing sulfur atom on their side chains; and the theoretical approach to the CD spectra using molecular orbital method-based calculation. Conformational analyses of cyclic and linear peptides by the CD spectra of their Dnp derivatives are also discussed. PMID- 15657881 TI - End-labeled free-solution electrophoresis of DNA. AB - DNA is a free-draining polymer. This subtle but "unfortunate" property of highly charged polyelectrolytes makes it impossible to separate nucleic acids by free flow electrophoresis. This is why one must typically use a sieving matrix, such as a gel or an entangled polymer solution, in order to obtain some electrophoretic size separation. An alternative approach consists of breaking the charge to friction balance of free-draining DNA molecules. This can be achieved by labeling the DNA with a large, uncharged molecule (essentially a hydrodynamic parachute, which we also call a drag-tag) prior to electrophoresis; the resulting methodology is called end-labeled free-solution electrophoresis (ELFSE). In this article, we review the development of ELFSE over the last decade. In particular, we examine the theoretical concepts used to predict the ultimate performance of ELFSE for single-stranded (ssDNA) sequencing, the experimental results showing that ELFSE can indeed overcome the free-draining issue raised above, and the technological advances that are needed to speed the development of competitive ELFSE-based sequencing and separation technologies. Finally, we also review the reverse process, called free-solution conjugate electrophoresis (FSCE), wherein uncharged polymers of different sizes can be analyzed using a short DNA molecule as an electrophoretic engine. PMID- 15657882 TI - Acid-base titration of melanocortin peptides: evidence of Trp rotational conformers interconversion. AB - Tryptophantime-resolved fluorescence was used to monitor acid-base titration properties of alpha-melanocyte stimulating hormone (alpha-MSH) and the biologically more potent analog [Nle4, D-Phe7]alpha -MSH (NDP-MSH), labeled or not with the paramagnetic amino acid probe 2,2,6,6-tetramthylpiperidine-N-oxyl-4 amino-4-carboxylic acid (Toac). Global analysis of fluorescence decay profiles measured in the pH range between 2.0 and 11.0 showed that, for each peptide, the data could be well fitted to three lifetimes whose values remained constant. The less populated short lifetime component changed little with pH and was ascribed to Trp g+ chi1 rotamer, in which electron transfer deactivation predominates over fluorescence. The long and intermediate lifetime preexponential factors interconverted along that pH interval and the result was interpreted as due to interconversion between Trp g- and trans chi1 rotamers, driven by conformational changes promoted by modifications in the ionization state of side-chain residues. The differences in the extent of interconversion in alpha-MSH and NDP-MSH are indicative of structural differences between the peptides, while titration curves suggest structural similarities between each peptide and its Toac-labeled species, in aqueous solution. Though less sensitive than fluorescence, the Toac electron spin resonance (ESR) isotropic hyperfine splitting parameter can also monitor the titration of side-chain residues located relatively far from the probe. PMID- 15657883 TI - The efficiency of electrokinetic pumping at a condition of maximum work. AB - Numerical methods are employed to examine the work, electric power input, and efficiency of electrokinetic pumps at a condition corresponding to maximum pump work. These analyses employ the full Poisson-Boltzmann equations and account for both convective and conductive electric currents, including surface conductance. We find that efficiencies at this condition of maximum work depend on three dimensionless parameters, the normalized zeta potential, normalized Debye layer thickness, and a fluid property termed the Levine number indicating the nominal ratio of convective to conductive electric currents. Efficiencies at maximum work exhibit a maximum for an optimum Debye layer thickness when the zeta potential and Levine number are fixed. This maximum efficiency increases with the square of the zeta potential when the zeta potential is small, but reaches a plateau as the zeta potential becomes large. The maximum efficiency in this latter regime is thus independent of the zeta potential and depends only on the Levine number. Simple analytical expressions describing this maximum efficiency in terms of the Levine number are provided. Geometries of a circular tube and planar channel are examined. PMID- 15657884 TI - Motion of single long DNA molecules through arrays of magnetic columns. AB - We present a videomicroscopy study of T4 DNA (169 kbp) in microfluidic arrays of posts formed by the self-assembly of magnetic beads. We observe DNA moving through an area of 10 000 microm(2), typically containing 100-600 posts. We determine the distribution of the contact times with the posts and the distribution of passage times across the field of view for hundreds of DNA per experiment. The contact time is well approximated by a Poisson process, scaling like the inverse of the field strength, independent of the density of the array. The distribution of passage times allows us to estimate the mean velocity and dispersivity of the DNA during its motion over distances long compared to our field of view. We compare these values with those computed from a lattice Monte Carlo model and geometration theory. We find reasonable quantitative agreement between the lattice Monte Carlo model and experiment, with the error increasing with increasing post density. The deviation between theory and experiment is attributed to the high mobility of DNA after disengaging from the posts, which leads to a difference between the contact time and the total time lost by colliding. Classical geometration theory furnishes surprisingly good agreement for the dispersivity, while geometration theory with a mean free path significantly overestimates the dispersivity. PMID- 15657885 TI - A triple-injection method for microchip electrophoresis. AB - We report here a novel triple injection method for microchip electrophoresis (micro-CE) that results in a higher intensity of DNA peaks. This new method includes a triple-repeated process of a combination of a sample loading voltage and a separation voltage in each interval, namely (loading time) + (separation time) + (loading time) + (separation time) + (loading time), prior to electrophoretic separation. All these injections were electrokinetically controlled by a software. Although the usual sample injection, which included the process of one 60 s electrokinetically application, was limited by the amount of sample, peaks of 40% higher intensity were obtained using the new method within half of the conventional injection time compared to the conventional method. Maximum peak intensity was successfully achieved with integration of the intensities of the triple-repeated peaks by adjusting the application period of the separation voltage. Repetition of the sample loading voltage for an adjusted period with a further adjusted period of separation voltage in each interval may be an effective method for injection of samples that results in peaks with higher intensity. PMID- 15657886 TI - Impact of reservoir potentials on the analyte behavior in microchip electrophoresis: computer simulation and experimental validation for DNA fragments. AB - Fundamental understanding of the impact of reservoir potentials on the analyte behavior on the microfluidic chips is an important issue in microchip electrophoresis (MCE) for suitable injection and separation of analytes, since the applied potentials may significantly affect the shape of sample plug, sample leakage from the injection channel to the separation channel, injected sample amount, and separation efficiency. This study addressed this issue for the case of a conventional cross-geometry microchip with four reservoirs using computer simulations, the results of which were verified by the analysis of DNA fragments. For the microchip with a definite structure and migration distance, the injected sample amount was shown to be the vital parameter for improving the limit of detection and resolution. During injection, the shape of the sample plug could be adjusted by varying the reservoir potentials. It was demonstrated that a "magnified injection" (applying high voltage on the three reservoirs to the sample reservoir) is useful to enhance the detection sensitivity depending on the analyte composition, although such injection was previously avoided because of introducing too large amounts of the analyte in comparison with two established modes, floating and pinched injection. Optimal magnified injection was proved to improve the sensitivity for about 4 times over that of pinched injection for the analysis of DNA step ladders using microchip gel electrophoresis (MCGE). Sample leakage of DNA fragments could be suppressed by applying a high positive voltage on injection channel during separation, but the voltage degraded the injected amount and resolution. PMID- 15657887 TI - Perspective on concentration polarization effects in electrochromatographic separations. AB - This work illustrates the appearance and electrohydrodynamic consequences of concentration polarization in the particulate and monolithic fixed beds used in capillary electrochromatography and related electrical-field assisted processes. Key property of most porous materials is the co-existence of bulk, quasi electroneutral macroporous regions and mesoporous compartments which are ion permselective (due to electrical double-layer overlap) causing different transport numbers for co-ionic and counterionic species, e.g., background electrolyte components, or the analytes. For a cathodic electroosmotic flow the (cation) permselectivity, together with diffusive and electrokinetic transport induces depleted and enriched concentration polarization zones at the anodic and cathodic interfaces, respectively, in dependence of the mobile phase ionic strength and applied electrical fields. At high field strength a secondary, nonequilibrium electrical double layer may be created in the depleted concentration polarization zones of a material stimulating electroosmosis of the second kind. The potential of this induced-charge electroosmosis with respect to nonlinear flow velocities and electrokinetic instability mixing (basically destroying the concentration polarization zones) is analyzed in view of the pore space morphology in random-close packings of spherical-shaped, porous particles and hierarchically structured monoliths. Possible applications based on a fine tuning of the illustrated effects emerge for microfluidic pumping and mixing, or the intensification of sample recovery in adsorption processes. With this perspective we want to focus the attention on concentration polarization in electrochromatographic systems by presenting and discussing original data acquired on relevant microscopic as well as macroscopic scales, and point towards the importance of related effects in colloid and membrane science.* PMID- 15657889 TI - Monomeric and polymeric anionic gemini surfactants and mixed surfactant systems in micellar electrokinetic chromatography. Part I: characterization and application as novel pseudostationary phases. AB - Sodium di(undecenyl) tartarate monomer (SDUT), a vesicle-forming amphiphilic compound possessing two hydrophilic carboxylate head groups and two hydrophobic undecenyl chains gemini surfactant, was prepared and polymerized to form a polymeric gemini surfactant (i.e., poly-SDUT). These anionic surfactant systems with carboxylate (SDUT and poly-SDUT) and sulfate (sodium dodecyl sulfate, SDS) head groups as well as mixed surfactant systems (SDS/SDUT, SDS/poly-SDUT, and SDUT/poly-SDUT) were then applied as novel pseudostationary phases in micellar electrokinetic chromatography (MEKC). The SDUT and poly-SDUT were characterized using various analytical techniques. Retention factors of 36 benzene derivatives were calculated in 20 mM phosphate buffer of each surfactant system. The retention factor values of the test solutes show that there are distinctive selectivity differences between the surfactant systems. Solute-pseudostationary phase interactions in MEKC were also examined by determining the free energy of transfer of the substituted functional groups from the aqueous buffer phase into the pseudostationary phase. PMID- 15657888 TI - Field gradient electrophoresis. AB - The class of equilibrium gradient methods utilizes the opposition of two forces, at least one of which changes in magnitude with position, to separate and concentrate analytes. The drawback of many methods of this type is that the production of two opposing forces requires in comparison to standard methods, such as capillary electrophoresis, a relatively complex apparatus. In addition, for techniques such as electric field gradient focusing, hydrodynamic flow leads to Taylor dispersion, which limits the attainable concentration factor. We propose a new method, gradient field electrophoresis, which achieves analyte separation and focusing with only one spatially varying force, an electric field gradient. A model for the method is developed and used to analyze peak capacity. Experimental results for a protein (R-phycoerythrin) are given and compared to the model. PMID- 15657890 TI - Monomeric and polymeric anionic gemini surfactants and mixed surfactant systems in micellar electrokinetic chromatography. Part II: characterization of chemical selectivity using two linear solvation energy relationship models. AB - Sodium di(undecenyl) tartarate monomer (SDUT), a vesicle-forming amphiphilic compound possessing two hydrophilic carboxylate headgroups and two hydrophobic undecenyl chains, was prepared and polymerized to form a polymeric vesicle (i.e., poly-SDUT). The anionic surfactants of SDUT and poly-SDUT (carboxylate head group) and sodium dodecyl sulfate, SDS (sulfate head groups) as well as mixed surfactant systems (SDS/SDUT, SDS/poly-SDUT, and SDUT/poly-SDUT) were applied as pseudostationary phases in micellar electrokinetic chromatography (MEKC). Two linear solvation energy relationship (LSER) models, i.e., solvatochromic and solvation parameter models, were successfully applied to investigate the effect of the type and composition of pseudostationary phases on the retention mechanism and selectivity in MEKC. The solvatochromic and solvation parameter models were used to help understand the fundamental nature of the solute-pseudostationary phase interactions and to characterize the properties of the pseudostationary phases (e.g., solute size and hydrogen bond-accepting ability for all pseudostationary phases). The solute types were found to have a significant effect on the LSER system coefficients and on the predicted retention factors. Although both LSER models provide the same information, the solvation parameter model is found to provide much better results both statistically and chemically than the solvatochromic model. PMID- 15657891 TI - System zones in capillary zone electrophoresis: moving boundaries caused by freely migrating hydrogen ions. AB - We demonstrate that system zones (SZs) can be expected in background electrolytes (BGEs) with a low buffer capacity. The mobilities of this type of SZ (m(SZ)) could be determined by calculations both based on a mathematical model and by a simulation program. The values of m(SZ) are increasing for decreasing buffer capacities and lower concentrations of the BGEs. For completely unbuffered BGEs with a pH below 7, the m(SZ) reached values up to 350x10(-9) m(2)V(-1)s(-1). This value indicates that the existence of this type of SZ originates from migrating hydrogen ions. Although both the mathematical model and simulation program do not consider the influence of the pH of the sample solution, experiments have shown that the pH of the sample solution is also very important. The lower the pH of the sample solution, the larger the m(SZ) in a specific BGE for cationic SZs. Using completely unbuffered BGEs the hydrogen ions present in the original sample start to migrate freely through the capillary tube with the mobility of a single hydrogen ion and cause stepwise disturbances in the base line of the detector trace. It is remarkable that this type of SZ appears not to affect the electromigration dispersion (EMD) in a strong way and so far only dips could be obtained experimentally. PMID- 15657892 TI - Identification of rat Gas7 isoforms differentially expressed in brain and regulated following kainate-induced neuronal injury. AB - The growth arrest-specific gene 7 (Gas7) is expressed primarily in the brain and is necessary for the formation of neurite in cultured cerebellar preneurons. The endogenous rat Gas7 (rGas7) is transiently elevated before nerve growth factor promoted neurite outgrowths emerge in cultured PC12 cells. We report three Gas7 isoforms (a, b, and c) in rat tissues. Peptide microsequencing identified two Gas7 forms, rGas7-a (38 kDa) and rGas7-b (47 kDa). rGas7-c can be predicted from a transcription variant by alternative splicing. Although two open reading frames were predicted, a cloned rGas7 cDNA encoded mostly rGas7-a in mammalian cells. The overexpression of the rGas7 cDNA in PC12 cells sufficed to promote small lamellipodia- and filopodia-like cell processes that resemble the initial stages of neurite formation. Three rGas7 isoforms were differentially expressed in all of the brain subregions. Only rGas7-a was detected in rat cerebellum, as in mouse cerebellum. Kainate injury did not affect the level of rGas7-b, but the level of isoform c was substantially suppressed in the hippocampus. Immunohistochemistry reveals that Gas7 was expressed primarily in the pyramidal neurons of the hippocampus and was quickly attenuated before recovery in the CA3 area after kainate was administered. These results suggest that differential expression and unique regulation of Gas7 isoforms in brain subregions may be important in specialized brain functions. Conservation of Gas7 isoforms by alternative splicing in mammals is also considered. PMID- 15657893 TI - Helix-helix interconversion rates of short 13C-labeled helical peptides as measured by dynamic NMR spectroscopy. AB - The rates at which a peptide hexamer and a peptide octamer interconvert between left- and right-handed helical forms in CD2Cl2 solution have been characterized by 13C dynamic NMR (DNMR) spectroscopy. The peptide esters studied are Fmoc (Aib)n-OtBu (n = 6 and 8), where Fmoc is 9-fluorenylmethyoxycarbonyl and Aib is the strongly helix-forming residue alpha-aminoisobutyric acid. Because the Aib residue is itself achiral, homooligomers of this residue form a 50/50 mixture of enantiomeric 3(10)-helices in solution. It has been demonstrated (R.-P. Hummel, C. Toniolo, and G. Jung, Angewandte Chemie International Edition, 1987, Vol. 26, pp. 1150-1152) that oligomers of Aib interconvert on the millisecond timescale. We have performed lineshape analysis of 13C-NMR spectra collected for our peptides enriched with 13C at a single residue. Rate constants for the octamer range from 6 s(-1) at 196 K to about 56,500 s(-1) at 320 K. At all temperatures, the hexamer interconverts about three times faster than the octamer. Eyring plots of the data reveal experimentally indistinguishable DeltaH++ values for the hexamer and octamer of 37.8 +/- 0.6 and 37.6 +/- 0.4 kJ mol(-1) respectively. The difference in the rates of interconversion is dictated by entropic factors. The hexamer and octamer exhibit negative DeltaS++ values of -29.0(-1) +/- 2.5 and 37.3 +/- 1.7 J K(-1) mol(-1), respectively. A mechanism for the helix-helix interconversion is proposed. and calculated DeltaG++ values are compared to the estimate for a decamer undergoing a helix-helix interconversion. PMID- 15657894 TI - Direct observation of spectral differences between normal and basal cell carcinoma (BCC) tissues using confocal Raman microscopy. AB - Raman spectroscopy has strong potential for providing noninvasive dermatological diagnosis of skin cancer. In this study, confocal Raman microscopy was applied to the dermatological diagnosis for one of the most common skin cancers, basal cell carcinoma (BCC). BCC tissues were obtained from 10 BCC patients using a routine biopsy and used for confocal Raman measurements. Autofluorescence signals from tissues, which interfere with the Raman signals, were greatly reduced using a confocal slit adjustment. Distinct Raman band differences between normal and BCC tissues for the amide I mode and the PO2- symmetric stretching mode showed that this technique has strong potential for use as a dermatological diagnostic tool without the need for statistical treatment of spectral data. It was also possible to precisely differentiate BCC tissue from surrounding noncancerous tissue using the confocal Raman depth profiling technique. We propose that confocal Raman microscopy provides a novel method for dermatological diagnosis since direct observations of spectral differences between normal and BCC tissues are possible. PMID- 15657895 TI - A new type of migrating zone boundary in electrophoresis: 1. General description of boundary behavior based on electromigration dispersion velocity profiles. AB - Till now two principal types of electrophoretic boundaries have been recognized: a steady-state one showing self-sharpening properties and an unsteady-state one showing electromigration dispersion (EMD). In this contribution, the existence of a third (hybrid) type of electrophoretic boundary is revealed, that shows simultaneously: (i) steady-state character with self-sharpening properties in a certain part of the boundary, demarcated by a certain range of its composition; (ii) unsteady-state character with EMD in the resting part. A new theoretical approach to the study of the structure and properties of electrophoretic boundaries is presented, based on EMD velocity profiles representing the dependence of the EMD velocity on the composition in any point of the transition region across the boundary. According to this approach, the linearity or nonlinearity of the EMD velocity profile is recommended as the criterion for distinguishing the actual type of the boundary in question. It is shown that the new type of electrophoretic boundary is not an exotic case but may be observed even in simple systems as, e.g., formed by adjacent zones containing mixtures of current species such as picrate and acetate. Computer simulations are presented which confirm the theoretical conclusions and illustrate the three types of electrophoretic boundaries. PMID- 15657896 TI - Implications of galactocerebrosidase and galactosylcerebroside metabolism in cancer cells. AB - Galactosylcerebroside is known to be overexpressed upon the cellular surface of a variety of cancers. In squamous cell carcinomas of the head and neck, one explanation for galactosylcerebroside accumulation has been identified as a transcriptional repression of the galactocerebrosidase gene. Galactocerebrosidase is the enzyme responsible for degrading galactosylcerebroside to ceramide. Ceramide is an important apoptosis activator, whereas galactosylcerebroside functions as an inhibitor. A shift of the ceramide metabolism balance in favor of glycosylated forms has been identified as a mechanism of drug resistance for several antineoplastic agents. Our review elaborates on possible explanations for galactocerebrosidase suppression and on other explanations for increased glycosphingolipid concentration within cancer cell membranes. Furthermore, conjecturable influences of a repressed galactocerebrosidase expression on tumor biology are to be explained. The inhibiting transcription factors YY1 and AP2 have been identified as potential galactocerebrosidase gene suppressors. The resulting accumulation of galactosylcerebroside promotes a reduction of cellular adhesion and inhibits apoptosis, leading to increased cellular growth, migration and prolonged cell survival contributing to carcinogenesis. PMID- 15657897 TI - Effects on proliferation and melanogenesis by inhibition of mutant BRAF and expression of wild-type INK4A in melanoma cells. AB - Activating BRAF mutations and loss of wild-type INK4A expression both occur at high frequencies in melanomas. Here, we present evidence that BRAF and INK4A have different effects on melanogenesis, a marker of melanocytic differentiation. Human melanoma cell line 624Mel harbors mutations in both BRAF and INK4A. The in vitro and in vivo growth of these cells was inhibited by either reduced expression of mutant BRAF using stable retroviral RNA interference (RNAi) or retrovirus-mediated stable expression of wild-type INK4A cDNA. Consistent with the observed growth inhibition, phosphorylation of S780 and S795 in pRB, both CDK4/6 targets, was suppressed in cells expressing either mutant BRAF RNAi or wild-type INK4A. Interestingly, melanoma cells expressing mutant BRAF RNAi had increased pigmentation, produced more mature melanosomes and melanin and expressed higher levels of tyrosinase and tyrosinase-related protein-1, whereas melanogenesis was not induced by wild-type INK4A. We found that the melanocyte lineage-specific master control protein microphthalmia-associated transcription factor was upregulated by inhibition of mutant BRAF, which may be the cause for the melanogenic effect of BRAF RNAi. The results suggest that, although both BRAF and INK4A lesions promote cell growth and tumor formation, mutant BRAF may also induce dedifferentiation in melanoma cells. PMID- 15657898 TI - Analyte and system eigenpeaks in nonaqueous capillary zone electrophoresis: theoretical description and experimental confirmation with methanol as solvent. AB - A mathematical model developed for aqueous solutions and adapted to methanol as solvent was applied to predict the electromigration characteristics of analytes and background electrolytes in capillary zone electrophoresis. These characteristics are the effective mobility, and the tendency of the analyte to undergo peak-broadening due to electromigration dispersion. The input parameters for calculation like limiting mobilities and dissociation constants were experimentally determined or taken from the literature. By the aid of the model, the molar response for conductivity detection was calculated as well as the transfer ratio when indirect UV detection was used. They allow depicting the electropherogram by computer simulation. An additional important program output is the prediction of the occurrence of system- or eigenpeaks that mimic peaks of analytes or electroosmotic flow markers. The measured electropherograms were in agreement with those theoretically predicted. Deviations were attributed to ion pairing in methanolic solutions, which was not implemented in the model. PMID- 15657899 TI - Presence of neuroglobin in cultured astrocytes. AB - Neuroglobin (Ngb), a recently discovered intracellular respiratory globin in neurons, may play a crucial role in oxygen homeostasis in the brain. We report preliminary findings indicating the presence of functional neuroglobin in primary cultures of cerebral cortical astrocytes. Reverse transcription real-time polymerase chain reaction (RRT-PCR) and immunostaining confirmed such presence in cultured astrocytes isolated from newborn mouse brain. Ngb antisense treatment increased apoptosis in ischemic astrocytes. The discovery of Ngb in astrocytes may provide some insight into how oxygen homeostasis is regulated in the brain. PMID- 15657900 TI - Pharmacological and small interference RNA-mediated inhibition of breast cancer associated fatty acid synthase (oncogenic antigen-519) synergistically enhances Taxol (paclitaxel)-induced cytotoxicity. AB - The relationship between breast cancer-associated fatty acid synthase (FAS; oncogenic antigen-519) and chemotherapy-induced cell damage has not been studied. We examined the ability of C75, a synthetic slow-binding inhibitor of FAS activity, to modulate the cytotoxic activity of the microtubule-interfering agent Taxol (paclitaxel) in SK-Br3, MDA-MB-231, MCF-7 and multidrug-resistant MDR-1 (P Glycoprotein)-overexpressing MCF-7/AdrR breast cancer cells. When the combination of C75 with Taxol in either concurrent (C75 + Taxol 24 hr) or sequential (C75 24 hr --> Taxol 24 hr) schedules were tested for synergism, addition or antagonism using the isobologram and the median-effect plot analyses, co-exposure of C75 and Taxol mostly demonstrated synergistic effects, whereas sequential exposure to C75 followed by Taxol mainly showed additive or antagonistic interactions. Because the nature of the cytotoxic interactions was definitely schedule-dependent in MCF 7 cells, we next evaluated the effects of C75 on Taxol-induced apoptosis as well as Taxol-activated cell death and cell survival-signaling pathways in this breast cancer cell model. An ELISA for histone-associated DNA fragments demonstrated that C75 and Taxol co-exposure caused a synergistic enhancement of apoptotic cell death, whereas C75 pre-treatment did not enhance the apoptosis-inducing activity of Taxol. Co-exposure to C75 and Taxol induced a remarkable nuclear accumulation of activated p38 mitogen-activated protein kinase (p38 MAPK), which was accompanied by a synergistic nuclear accumulation of the p53 tumor-suppressor protein that was phosphorylated at Ser46, a p38 MAPK-regulated pro-apoptotic modification of p53. As single agents, FAS blocker C75 and Taxol induced a significant stimulation of the proliferation and cell survival mitogen-activated protein kinase extracellular signal-regulated kinase (ERK1/ERK2 MAPK) activity, whereas, in combination, they interfered with ERK1/ERK2 activation. Moreover, the combined treatment of C75 and Taxol inactivated the anti-apoptotic AKT (protein kinase B) kinase more than either agent alone, as evidenced by a synergistic down regulation of AKT phosphorylation at its activating site Ser(473) without affecting AKT protein levels. To rule out a role for non-FAS C75-mediated effects, we finally used the potent and highly sequence-specific mechanism of RNA interference (RNAi) to block FAS-dependent signaling. Importantly, SK-Br3 and multi-drug resistant MCF-7/AdrR cells transiently transfected with sequence specific double-stranded RNA oligonucleotides targeting FAS gene demonstrated hypersensitivity to Taxol-induced apoptotic cell death. Our findings establish for the first time that FAS blockade augments the cytotoxicity of anti-mitotic drug Taxol against breast cancer cells and that this chemosensitizing effect is schedule-dependent. We suggest that the alternate activation of both the pro apoptotic p38 MAPK-p53 signaling and the cytoprotective MEK1/2 --> ERK1/2 cascade, as well as the inactivation of the anti-apoptotic AKT activity may explain, at least in part, the sequence-dependent enhancement of Taxol-induced cytotoxicity and apoptosis that follows inhibition of FAS activity in breast cancer cells. If chemically stable FAS inhibitors demonstrate systemic anticancer effects of FAS inhibition in vivo, these findings may render FAS as a valuable molecular target to enhance the efficacy of taxanes-based chemotherapy in human breast cancer. PMID- 15657902 TI - CLN3, the protein associated with batten disease: structure, function and localization. AB - Batten disease, an inherited neurodegenerative storage disease affecting children, results from the autosomal recessive inheritance of mutations in Cln3. The function of the CLN3 protein remains unknown. A key to understanding the pathology of this devastating disease will be to elucidate the function of CLN3 at the cellular level. CLN3 has proven difficult to study as it is predicted to be a membrane protein expressed at relatively low levels. This article is a critical review of various approaches used in examining the structure, trafficking, and localization of CLN3. We conclude that CLN3 is likely resident in the lysosomal/endosomal membrane. Different groups have postulated conflicting orientations for CLN3 within this membrane. In addition, CLN3 undergoes several posttranslational modifications and is trafficked through the endoplasmic reticulum and Golgi. Recent evidence also suggests that CLN3 traffics via the plasma membrane. Although the function of this protein remains elusive, it is apparent that genetic alterations in Cln3 may have a direct affect on lysosomal function. PMID- 15657904 TI - Large-volume stacking in capillary electrophoresis using pH hysteresis of the electroosmotic flow in a bare fused-silica capillary. AB - For large-volume stacking with the electroosmotic flow pump (LVSEP) in capillary electrophoresis of anionic analytes it is required that the electroosmotic mobility (EOM) should be smaller than the magnitudes of the effective mobilities of the analytes. When a fused-silica capillary is treated with an acidic solution, the silanoate group on the silica surface is neutralized to silanol and the EOM is suppressed. Due to the slow deprotonation equilibrium of the silanol group at an intermediate pH, this reduced EOM can be retained during a number of electrophoresis runs. Using a bare fused-silica capillary preconditioned with 0.01 M HCl, successful LVSEP at pH 6.0 was achieved for weakly acidic compounds with two orders of magnitude enhancements in the concentration sensitivity. The repeatability in migration times of ten analytes stacked by LVSEP in a single day was excellent with the relative standard deviation (RSD) less than 1% (n = 6). The day-to-day repeatability was also excellent with RSD less than 3% (n = 3 x 6) when the capillary was preconditioned each day. PMID- 15657903 TI - Conductivity properties of carrier ampholyte pH gradients in isoelectric focusing. AB - The conductivity properties of natural pH gradient created by carrier ampholytes were studied during the process of isoelectric focusing (IEF). IEF was performed in capillaries (10-30 mm long) or in microchips with the same channel length. A 10-30x reduction of the conductivity of the separation medium was observed during the establishment of pH gradient. Results obtained using different IEF voltages indicate that there is a nonlinear relationship between the conductivity of an established pH gradient and the applied electric field. Our theoretical analysis using a simplified model generated values that reasonably agree with the experimental data. In addition, we found that above a certain electric field ( approximately 300 V/cm), resolution does not increase with the applied voltage as predicated; we observed band-broadening and gel breakdown. The approach presented in this work can be used for optimization of the IEF separation and judicious selection of IEF conditions. PMID- 15657905 TI - Direct calculation of interconversion barriers in dynamic chromatography and electrophoresis: Isomerization of captopril. AB - Dynamic capillary electrophoresis (DCE) and direct calculation of the rate constants of isomerization has been applied to determine the cis-trans isomerization barriers of the angiotensin-converting enzyme inhibitor captopril. The separation of the rotational cis-trans isomeric drug has been performed in an aqueous 50 mM borate buffer at pH 9.3. Interconversion profiles featuring plateau formation, peak-broadening, and peak coalescence were observed. To determine the rate constants of the forward and backward reaction (k(cis-->trans) and k(trans- >cis)) of the isomerization process in dynamic capillary electrophoresis, a novel straightforward calculation method using the experimental parameters plateau height, h(plateau), peak width at half height w(h), the total migration times of the cis-trans isomers t(R) and the electroosmotic break-through time t(0) as well as the peak ratio of the cis-trans isomers is presented for the first time. From temperature dependent measurements the rate constants k(cis-->trans) and k(trans- >cis) and the kinetic activation parameters DeltaG( not equal), DeltaH( not equal), and DeltaS( not equal) of the cis-trans isomerization of captopril were obtained. From the activation parameters the isomerization barriers of captopril at 37 degrees C under basic conditions were calculated to be DeltaG( not equal) (cis-->trans) = 90.3 kJ.mol(-1)and DeltaG( not equal) (trans-->cis) = 90.0 kJ.mol(-1*). PMID- 15657906 TI - Analysis of biological motors via multidimensional fractionation: a strategy. AB - Past strategies for the analysis of ATP-fueled motors include single-motor analysis. Single-motor analysis bypasses limitations caused by motor asynchrony during the traditional ensemble averaging analysis. The present communication describes revised ensemble averaging analysis that also can bypass asynchrony derived limitations. This revised analysis makes measurements of one motor variable dependent on the others. One example is nondenaturing gel electrophoresis with more than one dimension. Each dimension measures one of the motor variables. This multidimensional procedure is used to obtain the values of "conformational" motor variables as a function of a "clock" motor variable. In theory, the cycle of the motor can be analyzed from a single multidimensional analysis of a collection of asynchronous motors sampled at only one time. That is to say, motor asynchrony becomes an asset, rather than a liability. PMID- 15657908 TI - Combination of feature-reduced MR spectroscopic and MR imaging data for improved brain tumor classification. AB - The purpose of this paper is to evaluate the effect of the combination of magnetic resonance spectroscopic imaging (MRSI) data and magnetic resonance imaging (MRI) data on the classification result of four brain tumor classes. Suppressed and unsuppressed short echo time MRSI and MRI were performed on 24 patients with a brain tumor and four volunteers. Four different feature reduction procedures were applied to the MRSI data: simple quantitation, principal component analysis, independent component analysis and LCModel. Water intensities were calculated from the unsuppressed MRSI data. Features were extracted from the MR images which were acquired with four different contrasts to comply with the spatial resolution of the MRSI. Evaluation was performed by investigating different combinations of the MRSI features, the MRI features and the water intensities. For each data set, the isolation in feature space of the tumor classes, healthy brain tissue and cerebrospinal fluid was calculated and visualized. A test set was used to calculate classification results for each data set. Finally, the effect of the selected feature reduction procedures on the MRSI data was investigated to ascertain whether it was more important than the addition of MRI information. Conclusions are that the combination of features from MRSI data and MRI data improves the classification result considerably when compared with features obtained from MRSI data alone. This effect is larger than the effect of specific feature reduction procedures on the MRSI data. The addition of water intensities to the data set also increases the classification result, although not significantly. We show that the combination of data from different MR investigations can be very important for brain tumor classification, particularly if a large number of tumors are to be classified simultaneously. PMID- 15657909 TI - Dissipation of acetochlor and its distribution in surface and sub-surface soil fractions during laboratory incubations. AB - Pesticides in soil are subject to a number of processes that result in transformation and biodegradation, sorption to and desorption from soil components, and diffusion and leaching. Pesticides leaching through a soil profile will be exposed to changing environmental conditions as different horizons with distinct physical, chemical and biological properties are encountered. The many ways in which soil properties influence pesticide retention and degradation need to be addressed to allow accurate predictions of environmental fate and the potential for groundwater pollution. Degradation and sorption processes were investigated in a long-term (100 days) study of the chloroacetanilide herbicide, acetochlor. Soil cores were collected from a clay soil profile and samples taken from 0-30 cm (surface), 1.0-1.3 m (mid) and 2.7 3.0 m (deep) and treated with acetochlor (2.5, 1.25, 0.67 microg acetochlor g(-1) dry wt soil, respectively). In sterile and non-sterile conditions, acetochlor concentration in the aqueous phase declined rapidly from the surface and subsoil layers, predominantly through nonextractable residue (NER) formation on soil surfaces, but also through biodegradation and biotic transformation. Abiotic transformation was also evident in the sterile soils. Several metabolites were produced, including acetochlor-ethane sulphonic acid and acetochlor-oxanilic acid. Transformation was principally microbial in origin, as shown by the differences between non-sterile and sterile soils. NER formation increased rapidly over the first 21 days in all soils and was mainly associated with the macroaggregate (>2000 microm diameter) size fractions. It is likely that acetochlor is incorporated into the macroaggregates through oxidative coupling, as humification of particulate organic matter progresses. The dissipation (ie total loss of acetochlor) half-life values were 9.3 (surface), 12.3 (mid) and 12.6 days (deep) in the non-sterile soils, compared with 20.9 [surface], 23.5 [mid], and 24 days [deep] in the sterile soils, demonstrating the importance of microbially driven processes in the rapid dissipation of acetochlor in soil. PMID- 15657912 TI - Alterations in gene expression profiles and the DNA-damage response in ionizing radiation-exposed TK6 cells. AB - Identifying genes that are differentially expressed in response to DNA damage may help elucidate markers for genetic damage and provide insight into the cellular responses to specific genotoxic agents. We utilized cDNA microarrays to develop gene expression profiles for ionizing radiation-exposed human lymphoblastoid TK6 cells. In order to relate changes in the expression profiles to biological responses, the effects of ionizing radiation on cell viability, cloning efficiency, and micronucleus formation were measured. TK6 cells were exposed to 0.5, 1, 5, 10, and 20 Gy ionizing radiation and cultured for 4 or 24 hr. A significant (P < 0.0001) decrease in cloning efficiency was observed at all doses at 4 and 24 hr after exposure. Flow cytometry revealed significant decreases in cell viability at 24 hr in cells exposed to 5 (P < 0.001), 10 (P < 0.0001), and 20 Gy (P < 0.0001). An increase in micronucleus frequency occurred at both 4 and 24 hr at 0.5 and 1 Gy; however, insufficient binucleated cells were present for analysis at the higher doses. Gene expression profiles were developed from mRNA isolated from cells exposed to 5, 10, and 20 Gy using a 350 gene human cDNA array platform. Overall, more genes were differentially expressed at 24-hr than at the 4-hr time point. The genes upregulated (> 1.5-fold) or downregulated (< 0.67 fold) at 4 hr were those primarily involved in the cessation of the cell cycle, cellular detoxification pathways, DNA repair, and apoptosis. At 24 hr, glutathione-associated genes were induced in addition to genes involved in apoptosis. Genes involved in cell cycle progression and mitosis were downregulated at 24 hr. Real-time quantitative PCR was used to confirm the microarray results and to evaluate expression levels of selected genes at the low doses (0.5 and 1.0 Gy). The expression profiles reflect the cellular and molecular responses to ionizing radiation related to the recognition of DNA damage, a halt in progression through the cell cycle, activation of DNA-repair pathways, and the promotion of apoptosis. PMID- 15657913 TI - Persistence of chromosome aberrations following acute radiation: II, does it matter how translocations are scored? AB - Chromosome breaks and rearrangements resulting from ionizing radiation can be much more complicated than many investigators thought possible some years ago. The realization that not all translocations are reciprocal, that multiway exchanges occur, and that some double-strand breaks are not repaired prior to mitosis have all contributed to the difficulty of knowing how best to identify, record, evaluate, and report chromosome translocations. Here we describe the results of a series of experiments in which blood from two normal healthy subjects was obtained, irradiated with 137Cs gamma-rays in vitro at doses ranging from 0 (controls) to 4 Gy, and cultured. Cells from each dose group and donor were harvested at days 2, 2.5, 3, 4, 5, and 7 and evaluated for chromosome damage by simultaneously painting chromosomes 1, 2, and 4 in red and 3, 5, and 6 in green. The persistence of dicentrics, fragments, rings, insertions, and PAINT translocations are reported separately by us in this issue. In this article, we focus on translocations, characterizing the various types in detail and comparing and contrasting their persistence across all dose groups for both donors. The results indicate that the persistence of all translocation types was sufficient to be used for retrospective dosimetry, although nonreciprocal translocations exhibited diminished persistence compared to the other types. We also characterize the kinetics of the radiation dose responses of the two donors who exhibited significant differences in the induction as well as the persistence of translocations. Based on the evidence presented here, we hypothesize that these individuals differ in the recognition and repair of radiation-induced damage as well as in cell cycle checkpoint control. Despite these differences, the temporal frequency of translocation losses at both the high and low doses was similar for both subjects. PMID- 15657914 TI - Troglitazone and liver injury: in search of answers. PMID- 15657915 TI - Persistence of chromosome aberrations following acute radiation: I, PAINT translocations, dicentrics, rings, fragments, and insertions. AB - Chromosome translocations are used to estimate the doses of radiation received following occupational or accidental exposure. Biodosimetry relies on the assumption that translocations are not cell-lethal and persist with little or no loss over time. While translocations do exhibit substantially greater persistence than other aberration types (e.g., dicentrics), there is evidence that translocation frequencies also decline over time, at least following acute doses above 1 Gy. To the extent that translocation frequencies decline, the predicted absorbed doses will be underestimated. Yet unknown is whether translocations induced by ionizing radiation at doses below 1 Gy also show significant declines. Here we report on the persistence of translocations induced by 137Cs gamma-rays at acute doses ranging from 0.2 to 4 Gy using peripheral blood lymphocytes from two unrelated healthy male donors. Chromosome aberrations were evaluated by simultaneously painting chromosomes 1, 2, and 4 in red and 3, 5, and 6 in green in cells harvested 2-7 days following exposure and were scored using the PAINT system. Translocations were also enumerated using several other methods and these results are reported separately by us in this issue. For comparison, the persistence of dicentrics, rings, acentric fragments, and color junctions was also evaluated and showed rapid losses with time. The results from both donors provide evidence that translocation frequencies decline with time in a statistically significant manner at doses as low as 0.2-0.3 Gy. The frequency of translocations for all dose groups declined from day 2 to 7 by averages of 39% and 26% for donors 1 and 2, respectively. These data emphasize the importance of considering translocation loss in biological dosimetry long times after exposure. PMID- 15657916 TI - In vivo mutagenesis induced by benzo[a]pyrene instilled into the lung of gpt delta transgenic mice. AB - Benzo[a]pyrene (B[a]P) is a ubiquitous airborne pollutant whose mutagenicity has been evaluated previously by oral and intraperitoneal administration to experimental animals. In this study, mutagenesis in the lungs, the target organ of air pollutants, was examined after a single intratracheal instillation of 0-2 mg B[a]P into gpt delta transgenic mice. Intratracheal injection of B[a]P resulted in a statistically significant and dose-dependent increase in gpt mutant frequency as measured by 6-thioguanine selection. The mutant frequencies at B[a]P doses of 0.5, 1, and 2 mg were 2.8, 4.2, and 6.8 times higher than the frequency seen in nontreated mice (0.60 +/- 0.13 x 10(-5)). The most frequent mutations induced by B[a]P treatment were G:C-->T:A transversions, which are characteristic of B[a]P mutagenesis in other models, and single-base deletions of G:C base pairs. To characterize the hotspots of B[a]P-induced mutations in the gpt gene, we analyzed sequences adjacent to the mutated G:C base pairs. Guanine bases centered in the nucleotide sequences CGT, CGA, and CGG were the most frequent targets of B[a]P. Our results indicate that intratracheal instillation of B[a]P into gpt delta mice causes a dose-dependent increase in gpt mutant frequency in the lung, and that the predominant mutation induced is G:C-->T:A transversion. PMID- 15657917 TI - Optimizing outcomes and bridging biliary atresia into adulthood. PMID- 15657918 TI - Associations between XPC expression, genotype, and the risk of head and neck cancer. AB - Squamous cell carcinoma of the head and neck (SCCHN) is a relatively rare cancer with a poor prognosis. In the present study, we investigated susceptibility biomarkers for SCCHN in 155 Koreans (73 SCCHN cases and 82 controls). Expression of the xeroderma pigmentosum group C (XPC) DNA-repair gene was measured by TaqMan fluorogenic real-time RT-PCR using RNA isolated from peripheral blood samples. In addition, the XPC-PAT genotype [an intronic and biallelic poly(AT) insertion/deletion polymorphism] was determined by PCR of peripheral blood DNA. Among known environmental and physical risk factors, age, gender, tobacco smoking, and alcohol consumption were associated with the SCCHN risk (P < 0.01). Lower XPC expression levels were found for the SCCHN cases, particularly in the larynx, than for the controls (P < 0.05); the geometric means (standard deviations) of XPC expression normalized by 18S rRNA in the cases and controls were 5.89 (8.13) and 15.14 (9.77), respectively. However, the distributions of the XPC-PAT genotypes were similar in the cases and controls. In addition, there were no associations between the expression of XPC and the PAT genetic polymorphism. XPC expression was not affected by age, gender, tobacco smoking, or alcohol consumption. After adjusting for SCCHN-associated lifestyle factors, the effective strength of XPC expression on SCCHN risk was weaker (0.05 < P < 0.1) than those of tobacco smoking and age (P < 0.05). There was a positive association between XPC mRNA expression in blood and SCCHN tissues (n = 9; P < 0.01); therefore, peripheral blood appears to be a reasonable surrogate tissue for XPC expression in tumor tissue. In conclusion, the XPC-PAT polymorphism had no effect on XPC expression or SCCHN risk. However, XPC expression may influence SCCHN risk. PMID- 15657919 TI - Somatic mutant frequency at the HPRT locus in children associated with a pediatric cancer cluster linked to exposure to two superfund sites. AB - The somatic mutant frequency (Mf) of the hypoxanthine phosphoribosyl transferase (HPRT) gene has been widely used as a biomarker for the genotoxic effects of exposure but few studies have found an association with environmental exposures. We measured background Mfs in 49 current and former residents of Dover Township, New Jersey, who were exposed during childhood to industrially contaminated drinking water. The exposed subjects were the siblings of children who developed cancer after residing in Dover Township, where the incidence of childhood cancer has been elevated since 1979. Mfs from this exposed group were compared to Mfs in 43 age-matched, presumably unexposed residents of neighboring communities with no known water contamination and no increased cancer incidence. Statistical comparisons were based on the natural logarithm of Mf (lnMF). The mean Mf for the exposed group did not differ significantly from the unexposed group (3.90 x 10( 6) vs. 5.06 x 10(-6); P = 0.135), but unselected cloning efficiencies were higher in the exposed group (0.55 vs. 0.45; P = 0.005). After adjustment for cloning efficiency, lnMf values were very similar in both groups and age-related increases were comparable to those previously observed in healthy children. The results suggest that HPRT Mf may not be a sensitive biomarker for the genotoxic effects of environmental exposures in children, particularly when substantial time has elapsed since exposure. PMID- 15657920 TI - Insight into hepatotoxicity: The troglitazone experience. PMID- 15657921 TI - Genotoxicity of inorganic lead salts and disturbance of microtubule function. AB - Lead compounds are known genotoxicants, principally affecting the integrity of chromosomes. Lead chloride and lead acetate induced concentration-dependent increases in micronucleus frequency in V79 cells, starting at 1.1 microM lead chloride and 0.05 microM lead acetate. The difference between the lead salts, which was expected based on their relative abilities to form complex acetato cations, was confirmed in an independent experiment. CREST analyses of the micronuclei verified that lead chloride and acetate were predominantly aneugenic (CREST-positive response), which was consistent with the morphology of the micronuclei (larger micronuclei, compared with micronuclei induced by a clastogenic mechanism). The effects of high concentrations of lead salts on the microtubule network of V79 cells were also examined using immunofluorescence staining. The dose effects of these responses were consistent with the cytotoxicity of lead(II), as visualized in the neutral-red uptake assay. In a cell-free system, 20-60 microM lead salts inhibited tubulin assembly dose dependently. The no-observed-effect concentration of lead(II) in this assay was 10 microM. This inhibitory effect was interpreted as a shift of the assembly/disassembly steady-state toward disassembly, e.g., by reducing the concentration of assembly-competent tubulin dimers. The effects of lead salts on microtubule-associated motor-protein functions were studied using a kinesin gliding assay that mimics intracellular transport processes in vitro by quantifying the movement of paclitaxel-stabilized microtubules across a kinesin coated glass surface. There was a dose-dependent effect of lead nitrate on microtubule motility. Lead nitrate affected the gliding velocities of microtubules starting at concentrations above 10 microM and reached half-maximal inhibition of motility at about 50 microM. The processes reported here point to relevant interactions of lead with tubulin and kinesin at low dose levels. PMID- 15657925 TI - Cytomics: an entry to biomedical cell systems biology. PMID- 15657926 TI - Chromatin supraorganization and extensibility in mouse hepatocytes following starvation and refeeding. AB - BACKGROUND: The effect of 48 h of starvation and of 48 h of refeeding subsequent to starvation on chromatin supraorganization and extensibility was studied in hepatocytes from adult mice. METHODS: Methods used involved topochemical assays, image analysis, gravity action, and polarization microscopy. RESULTS: Starvation increased the chromatin packing states, especially in areas of noncondensed chromatin, and induced drastic decreases in concanavalin A reactivity due to nuclear matrix glycoproteins and the frequency of nuclei with chromatin extensibility under gravity. Changes in chromatin packing state were accompanied by shifts of nuclear areas of part of the nuclear population to smaller values but did not affect the respective Feulgen-DNA amounts except for a few nuclei. The extent of chromatin unpackaging, but not of frequency of nuclei with formation of extended chromatin fibers, in starved mice that were refed was greater than in well-fed controls. Refeeding induced increase in Feulgen-DNA amounts and regain and redistribution of concanavalin A-reactive nuclear glycoproteins. However, the duration of refeeding used was probably insufficient to reestablish the stereo arrangement of the chromatin-nuclear matrix and to restore chromatin fluidity to the level seen in well-fed mice. CONCLUSIONS: The changes in the liver cell nuclei associated with starvation and refeeding of adult mice involved chromatin supraorganization, hepatocyte proliferation (refeeding), and the loss, regain, and redistribution of nuclear proteins, especially nuclear matrix components, related to chromatin organization and extensibility. These changes are suggested as favoring the silencing and reactivation of transcriptional activities, depending on the organism's nutritional state. PMID- 15657927 TI - Selective targeting of indel-inferred differences in spatial structures of highly homologous proteins. AB - Recent findings have shown that the protein elongation factor-1alpha (EF-1alpha) from the eukaryotic pathogen Leishmania donovani possesses virulence properties. This was unexpected, since it has greater than 80% sequence identity with its human homologue. Given that EF-1alpha is essential for cell survival, in principle, it can be considered an attractive drug target. However, the challenge is to be able to selectively target the protein so as not to affect function of the human homologue. While a limited number of discrete differences were scattered throughout the sequence, most of the difference between these 2 homologues could be attributed to a 12-amino acid insert present in human EF 1alpha and absent from the leishmania sequence. In the present study, we modeled the spatial differences in structures of human and L. donovani EF-1alpha's inferred by this insertion-deletion (or "indel"). The protein models were used to develop antibodies directed specifically toward the deletion region of the pathogen protein. The strategy described allowed successful selective targeting of this putative leishmania virulence factor while avoiding recognition of the highly similar human EF-1alpha homologue. These findings may establish a new strategy for the development of antagonists directed against certain pathogenic targets having close human homologues. PMID- 15657928 TI - YbiV from Escherichia coli K12 is a HAD phosphatase. AB - The protein YbiV from Escherichia coli K12 MG1655 is a hypothetical protein with sequence homology to the haloacid dehalogenase (HAD) superfamily of proteins. Although numerous members of this family have been identified, the functions of few are known. Using the crystal structure, sequence analysis, and biochemical assays, we have characterized YbiV as a HAD phosphatase. The crystal structure of YbiV reveals a two-domain protein, one with the characteristic HAD hydrolase fold, the other an inserted alpha/beta fold. In an effort to understand the mechanism, we also solved and report the structures of YbiV in complex with beryllofluoride (BeF3-) and aluminum trifluoride (AlF3), which have been shown to mimic the phosphorylated intermediate and transition state for hydrolysis, respectively, in analogy to other HAD phosphatases. Analysis of the structures reveals the substrate-binding cavity, which is hydrophilic in nature. Both structure and sequence homology indicate YbiV may be a sugar phosphatase, which is supported by biochemical assays that measured the release of free phosphate on a number of sugar-like substrates. We also investigated available genomic and functional data in an effort to determine the physiological substrate. PMID- 15657929 TI - David Blow (1931-2004). PMID- 15657930 TI - Crystal structure of an alanine-glyoxylate aminotransferase from Anabaena sp. at 1.70 A resolution reveals a noncovalently linked PLP cofactor. PMID- 15657931 TI - Properties of polyproline II, a secondary structure element implicated in protein protein interactions. AB - The polyproline II (PPII) conformation of protein backbone is an important secondary structure type. It is unusual in that, due to steric constraints, its main-chain hydrogen-bond donors and acceptors cannot easily be satisfied. It is unable to make local hydrogen bonds, in a manner similar to that of alpha helices, and it cannot easily satisfy the hydrogen-bonding potential of neighboring residues in polyproline conformation in a manner analogous to beta strands. Here we describe an analysis of polyproline conformations using the HOMSTRAD database of structurally aligned proteins. This allows us not only to determine amino acid propensities from a much larger database than previously but also to investigate conservation of amino acids in polyproline conformations, and the conservation of the conformation itself. Although proline is common in polyproline helices, helices without proline represent 46% of the total. No other amino acid appears to be greatly preferred; glycine and aromatic amino acids have low propensities for PPII. Accordingly, the hydrogen-bonding potential of PPII main-chain is mainly satisfied by water molecules and by other parts of the main chain. Side-chain to main-chain interactions are mostly nonlocal. Interestingly, the increased number of nonsatisfied H-bond donors and acceptors (as compared with alpha-helices and beta-strands) makes PPII conformers well suited to take part in protein-protein interactions. PMID- 15657932 TI - Understanding the energetics of helical peptide orientation in membranes. AB - Understanding the energetic factors determining the positioning and orientation of single-helical peptides in membranes is of fundamental interest in structural biology. Here, a simple 5-slab continuum dielectric model for the membrane is examined that distinguishes between the solvent, headgroup, and core regions. An analytical solution for the electrostatic solvation of a single dipole and an all atom model of N-methylacetamide are used to demonstrate the effect of the dielectric boundaries in the system on peptide dipole orientation. The dipole orientation energy is shown to dominate the electrostatic solvation energy of a polyalanine helix in the membrane. With an additional surface-area-dependent term to account for the cavity formation in the aqueous region, the continuum electrostatics description is used to examine several helical peptides, the atoms of which are explicitly represented with a molecular mechanics force field. The experimentally determined tilt angles of a number of peptides of alternating alanine and leucine residues, and of glycophorin and melittin, are accurately reproduced by the model. The factors determining the tilt angles and their fluctuations are analyzed. The tilt angles of the simpler peptides are found to increase approximately linearly with peptide length; this effect is also rationalized. The analysis and model presented here provide a step toward the prediction of helical membrane protein structure. PMID- 15657933 TI - The Protein Coil Library: a structural database of nonhelix, nonstrand fragments derived from the PDB. AB - Approximately half the structure of folded proteins is either alpha-helix or beta strand. We have developed a convenient repository of all remaining structure after these two regular secondary structure elements are removed. The Protein Coil Library (http://roselab.jhu.edu/coil/) allows rapid and comprehensive access to non-alpha-helix and non-beta-strand fragments contained in the Protein Data Bank (PDB). The library contains both sequence and structure information together with calculated torsion angles for both the backbone and side chains. Several search options are implemented, including a query function that uses output from popular PDB-culling servers directly. Additionally, several popular searches are stored and updated for immediate access. The library is a useful tool for exploring conformational propensities, turn motifs, and a recent model of the unfolded state. PMID- 15657934 TI - 14q32.3 deletion syndrome with autism. PMID- 15657935 TI - Small fiber diameter fibro-porous meshes: tissue response sensitivity to fiber spacing. AB - The purpose of this research was to determine if fiber spacing for small fiber diameter fibro-porous meshes affected tissue response in vivo. Disk-shaped polyurethane meshes, with mean fiber diameters of 7.6 microm and fiber spacing between 6 and 68 microm, were implanted in rat subcutaneous dorsum for 5-week intervals and then prepared for light microscopy and morphological analysis. Results showed that implants with 12- to 68-microm spacing had no histologically apparent fibrous capsule around the perimeter, a result different from that for 6 microm spacing samples that had a capsule around a mean of 34.2% of the perimeter. For the 12- to 68-microm spacing range, a mean of 21.0% of individual fibers within the meshes were encapsulated. Qualitatively, it appeared that larger fibers were encapsulated more frequently than smaller ones. When nodeless or baggy meshes were implanted, cells tended to cluster three or more fibers into groups and then encapsulate each group. Over the 6- to 68-microm spacing range, cell nuclei volume fraction within the meshes increased from the 6- to the 29 microm spacing (p = 0.000) and then decreased from the 29- to the 68-microm spacing (p = 0.015). There was a trend of an increase in local vessel volume fraction with spacing over the 6- to 68-microm range, though the relationship was weak. The results indicate that the reason for the lack of encapsulation of small fiber fibro-porous meshes is not exclusively a pore boundary explanation, as is proposed for small-pore porous meshes. PMID- 15657936 TI - Flow perfusion culture induces the osteoblastic differentiation of marrow stroma cell-scaffold constructs in the absence of dexamethasone. AB - Flow perfusion culture of scaffold/cell constructs has been shown to enhance the osteoblastic differentiation of rat bone marrow stroma cells (MSCs) over static culture in the presence of osteogenic supplements including dexamethasone. Although dexamethasone is known to be a powerful induction agent of osteoblast differentiation in MSC, we hypothesied that the mechanical shear force caused by fluid flow in a flow perfusion bioreactor would be sufficient to induce osteoblast differentiation in the absence of dexamethasone. In this study, we examined the ability of MSCs seeded on titanium fiber mesh scaffolds to differentiate into osteoblasts in a flow perfusion bioreactor in both the presence and absence of dexamethasone. Scaffold/cell constructs were cultured for 8 or 16 days and osteoblastic differentiation was determined by analyzing the constructs for cellularity, alkaline phosphatase activity, and calcium content as well as media samples for osteopontin. For scaffold/cell constructs cultured under flow perfusion, there was greater scaffold cellularity, alkaline phosphatase activity, osteopontin secretion, and calcium deposition compared with static controls, even in the absence of dexamethasone. When dexamethasone was present in the cell culture medium under flow perfusion conditions, there was further enhancement of osteogenic differentiation as evidenced by lower scaffold cellularity, greater osteopontin secretion, and greater calcium deposition. These results suggest that flow perfusion culture alone induces osteogenic differentiation of rat MSCs and that there is a synergistic effect of enhanced osteogenic differentiation when both dexamethasone and flow perfusion culture are used. PMID- 15657937 TI - CNP is required for maintenance of axon-glia interactions at nodes of Ranvier in the CNS. AB - Axoglial interactions underlie the clustering of ion channels and of cell adhesion molecules, regulate gene expression, and control cell survival. We report that Cnp1-null mice, lacking expression of the myelin protein cyclic nucleotide phosphodiesterase (CNP), have disrupted axoglial interactions in the central nervous system (CNS). Nodal sodium channels (Nav) and paranodal adhesion proteins (Caspr) are initially clustered normally, but become progressively disorganized with age. These changes are characterized by mislocalized Caspr immunostaining, combined with a decrease of clustered Na+ channels, and occur before axonal degeneration and microglial invasion, both prominent in older Cnp1 null mice. We suggest that CNP is a glial protein required for maintaining the integrity of paranodes and that disrupted axoglial signaling at this site underlies progressive axonal degeneration, observed later in the CNS of Cnp1-null mice. PMID- 15657938 TI - Metabotropic P2 receptor activation regulates oligodendrocyte progenitor migration and development. AB - To gain insights into the role of purinergic receptors in oligodendrocyte development, we characterized the expression and functional activity of P2 receptors in cultured rat oligodendrocyte progenitors and investigated the effects of ATP and its breakdown products on the migration and proliferation of this immature glial cell population. Using Western blot analysis, we show that oligodendrocyte progenitors express several P2X (P2X(1,2,3,4,7)) and P2Y (P2Y(1,2,4)) receptors. Intracellular Ca(2+) recording by Fura-2 video imaging allowed to determine the rank potency order of the P2 agonists tested: ADPbetaS = ADP = Benzoyl ATP > ATP > ATPgammaS > UTP, alpha,beta-meATP ineffective. Based on the above findings, on pharmacological inhibition by the antagonists oxATP and MRS2179, and on the absence of alpha,betameATP-induced inward current in whole cell recording, P2X(7) and P2Y(1) were identified as the main ionotropic and metabotropic P2 receptors active in OPs. As a functional correlate of these findings, we show that ATP and, among metabotropic agonists, ADP and the P2Y(1) specific agonist ADPbetaS, but not UTP, induce oligodendrocyte progenitor migration. Moreover, ATP and ADP inhibited the proliferation of oligodendrocyte progenitors induced by platelet-derived growth factor, both in purified cultures and in cerebellar tissue slices. The effects of ATP and ADP on cell migration and proliferation were prevented by the P2Y(1) antagonist MRS2179. By confocal laser scanning microscopy, P2Y(1) receptors were localized in NG2-labeled oligodendrocyte progenitors in the developing rat brain. These data indicate that ATP and ADP may regulate oligodendrocyte progenitor functions by a mechanism that involves mainly activation of P2Y(1) receptors. PMID- 15657939 TI - Glial restricted precursors protect against chronic glutamate neurotoxicity of motor neurons in vitro. AB - We have examined the expression of glutamate transporters in primary and immortalized glial precursors (GRIPs). We subsequently transduced these cells with the GLT1 glutamate transporter and examined the ability of these cells to protect motor neurons in an organotypic spinal cord culture. We show that glial restricted precursors and GRIP-derived astrocytes predominantly express glutamate transporters GLAST and GLT1. Oligodendrocyte differentiation of GRIPs results in downregulation of all glutamate transporter subtypes. Having identified these precursor cells as potential vectors for delivering glutamate transporters to regions of interest, we engineered a line of GRIPS that overexpress the glutamate transporter GLT1. These cells (G3 cells) have a nearly fourfold increase in glutamate transporter expression and at least a twofold increase in the V(max) for glutamate transport. To assess whether G3 seeding can protect motor neurons from chronic glutamate neurotoxicity, G3s were seeded onto rat organotypic spinal cord cultures. These cultures have previously been used extensively to understand pathways involved in chronic glutamate neurotoxicity of motor neurons. After G3 seeding, cells integrated into the culture slice and resulted in levels of glutamate transport sufficient to enhance total glutamate uptake. To test whether neuroprotection was related to glutamate transporter overexpression, we isolated GRIPS from the GLT1 null mouse to serve as controls. The seeding of G3s resulted in a reduction of motor neuron cell death. Hence, we believe that these cells may potentially play a role in cell-based neuroprotection from glutamate excitotoxicity. PMID- 15657941 TI - Proliferating resident microglia express the stem cell antigen CD34 in response to acute neural injury. AB - Reactive microgliosis is a highly characteristic response to neural injury and disease, which may influence neurodegenerative processes and neural plasticity. We have investigated the origin and characteristics of reactive microglia in the acute phase of their activation in the dentate gyrus following transection of the entorhino-dentate perforant path projection. To investigate the possible link between microglia and hematopoietic precursors, we analyzed the expression of the stem cell marker CD34 by lesion-reactive microglia in conjunction with the proliferation marker bromodeoxyuridine (BrdU) and the use of radiation bone marrow (BM) chimeric mice. We found that CD34 is upregulated on early-activated resident microglia, rather than by infiltrating bone marrow-derived cells. The number of CD34(+) microglia peaked at day 3 when 67% of the resident CD11b/Mac 1(+) microglia co-expressed CD34, and all CD34(+) cells co-expressed Mac-1, and decreased sharply toward day 5, unlike Mac-1, which was maximally expressed at day 5. Approximately 80% of the CD34(+) cells in the denervated dentate gyrus had incorporated BrdU into their nuclei at day 3. We also showed that CD34 is upregulated on early-activated microglia in the facial motor nucleus following peripheral axotomy. The results suggest lesion-reactive microglia to consist of functionally distinct subpopulations of cells; a major population of activated resident CD34(+)Mac-1(+) microglia with a high capacity for self-renewal, and a subpopulation of CD34(-)Mac-1(+) microglia which has a mixed extrinsic and intrinsic origin and whose proliferative capacity is unknown. PMID- 15657940 TI - Intermediate filament protein synemin is present in human reactive and malignant astrocytes and associates with ruffled membranes in astrocytoma cells. AB - Synemin, a very unique type VI intermediate filament (IF) protein, exhibits alternative splice variants termed alpha and beta. Unlike other IF proteins, synemin binds to actin-associated proteins, including alpha-actinin, vinculin, and alpha-dystrobrevin. Our previous work has demonstrated the presence of synemin in differentiating astrocytes. In this study, we have examined the presence of synemin in human astrocytes under pathological conditions, using rabbit antibodies raised against the C-terminal domain of human synemin produced in bacteria. Western blotting shows that astrocytic tumors contain greater amounts of alpha-synemin than do normal brain tissues. These tumors also contain beta-synemin, which is not detectable in normal brain. Immunohistochemistry demonstrates that, while synemin is present in normal adult brain only in vascular smooth muscle cells, it is newly synthesized by reactive and neoplastic astrocytes. Alpha- and beta-Synemins have also been detected by Western blotting and polymerase chain reaction in several human glioblastoma cell lines. In these cell lines, surprisingly, synemin is associated with ruffled membranes in addition to being distributed along the IF network. In ruffled membranes, synemin was found to co-localize with alpha-actinin. This unusual cellular localization for an IF protein is maintained after nocodazole-induced perinuclear coiling of the vimentin IF network. In addition, immunoprecipitation experiments demonstrate that synemin forms a complex with alpha-actinin in glioblastoma cells. Taken together with synemin localization within ruffled membranes, this finding suggests that synemin plays a role in motility of glioblastoma cells. PMID- 15657942 TI - Differential susceptibility of C2C12 myoblasts and myotubes to group II phospholipase A2 myotoxins from crotalid snake venoms. AB - Group II phospholipase A(2) (PLA(2)) myotoxins isolated from Viperidae/Crotalidae snake venoms induce a rapid cytolytic effect upon diverse cell types in vitro. Previous studies suggested that this effect could be more pronounced on skeletal muscle myotubes than on other cell types, including undifferentiated myoblasts. This study utilized the murine skeletal muscle C2C12 cell line to investigate whether differentiated myotubes are more susceptible than myoblasts, and if this characteristic is specific for the group II myotoxic PLA(2)s. The release of lactic dehydrogenase was quantified as a measure of cytolysis, 3 h after cell exposure to different group II PLA(2)s purified from Bothrops asper, Atropoides nummifer, Cerrophidion godmani, and Bothriechis schlegelii venoms. In addition, susceptibility to lysis induced by synthetic melittin and group III PLA(2) from bee (Apis mellifera) venom, as well as by anionic, cationic, and neutral detergents, was comparatively evaluated on the two cultures. Myotubes were significantly more susceptible to group II PLA(2) myotoxins, but not to the other agents tested, under the same conditions. Moreover, the increased susceptibility of myotubes over myoblasts was also demonstrated with two cytolytic synthetic peptides, derived from the C-terminal region of Lys49 PLA(2) myotoxins, that reproduce the action of their parent proteins. These results indicate that fusion and differentiation of myoblasts into myotubes induce changes that render these cells more susceptible to the toxic mechanism of group II PLA(2) myotoxins, but not to general perturbations of membrane homeostasis. Such changes are likely to involve myotoxin acceptor site(s), which remain(s) to be identified. PMID- 15657943 TI - Effect of D-penicillamine on rat lung elastin cross-linking during the perinatal period. AB - This study was designed to clarify the effects of D-penicillamine (DPA), a drug used for treatment of various pathological events, on lung elastin formation and maturation of the newborn in the perinatal period. The investigation was conducted on 20 newborn rats bred from 40 female and six male rats. DPA doses 400 mg kg(-1) day(-1) and physiological saline were given intraperitoneally (i.p) to experimental and control groups. To assess newborn maturation, their body and lung weights were determined. Serum Cu levels were measured by atomic absorption spectroscopy and ceruloplasmin (Cp) activities were measured spectrophotometrically. Newborn lung tissue elastin, desmosine (DES) and isodesmosine (IDES) levels were measured by HPLC. The results showed that DPA treatment caused loss of skin elasticity and reduction in body and lung weight in newborns of the experimental group. The serum Cu levels and Cp activity were found to be significantly lower in both maternal and newborn of the experimental groups compared with the control group. The lung DES, IDES and elastin values of newborns in the experimental group were decreased compared with the control group. In conclusion, our results indicate that 400 mg kg(-1) day(-1) DPA, a dose that is used in the treatment of Wilson's disease, rheumatoid arthritis and cystinuria, caused the retardation of newborn maturation, a decrease in DES-IDES cross-links and levels of lung elastin of offspring in the perinatal period. Another conclusion to be drawn from this study is that even low levels of Cu depletion due to DPA administration induces a change in cross-linking in lung elastin during the perinatal period. PMID- 15657944 TI - Prefractionation techniques in proteome analysis: the mining tools of the third millennium. AB - The present review deals with prefractionation protocols used in proteomic investigation in preparation for mass spectrometry (MS) or two-dimensional electrophoresis (2-DE) map analysis. Briefly, reported methods focus on cell organelle differential centrifugation and on chromatographic approaches, to continue in extenso with a panoply of electrophoretic methods. In the case of chromatography, procedures useful as a prefractionation step, including affinity, ion-exchange, and reversed-phase resins, revealed several hundreds of new species, previously undetected in unfractionated samples. Novel chromatographic prefractionation methods are also discussed such as a multistaged fractionation column, consisting in a set of immobilized chemistries, serially connected in a stack format (an assembly of seven blocks), each capable of harvesting a given protein population. Such a method significantly simplifies the complexity of treated samples while concentrating species, all resulting in a larger number of visible proteins by MS or 2-DE. Electrophoretic prefractionation protocols include all those electrokinetic methodologies which are performed in free solution, essentially all relying on isoelectric focusing steps (although some approaches based on gels and granulated media are also discussed). Devices associated with electrophoretic separation are multichamber apparatus, such as the multicompartment electrolyzers equipped with either isoelectric membranes or with isoelectric beads. Multicup device electrophoresis and several others, exploiting the conventional technique of carrier ampholyte focusing, are reviewed. This review also reports approaches for sample treatments in order to detect low-abundance species. Among others, a special emphasis is made on the reduction of concentration difference between proteins constituting a sample. This latter consists in a library of combinatorial ligands coupled to small beads. Such a library comprises hexameric ligands composed of 20 amino acids, resulting in millions of different structures. When these beads are impregnated with complex proteomes (e.g., human sera) of widely differing protein compositions, they are able to significantly reduce the concentration differences, thus greatly enhancing the possibility to evidence low-abundance species. It is felt that this panoply of methods could offer a strong step forward in "mining below the tip of the iceberg" for detecting the "unseen proteome". PMID- 15657945 TI - HLA-B27: portraying immunodominant viral epitopes. AB - Although the crystal structure of HLA-B27 has been known for a long time, only recently have X-ray diffraction studies of this molecule in complex with individual peptides become available. The report of three such structures involving viral epitopes that are immunodominant in HLA-B27-restricted T cell responses against influenza, Epstein-Barr and HIV viruses significantly improves our perception of critical aspects of the immunological and pathogenetic roles of HLA-B27, including (1) the molecular basis of its peptide-binding specificity and how this is modulated by subtype polymorphism, (2) the relationship between the structural and the antigenic features of immunodominant viral epitopes, (3) the basis for long term non-progression to AIDS of HIV-infected HLA-B27+ individuals, and (4) the structural features of microbial peptides influencing NK receptor engagement. Here, I discuss the implications of this and related studies for the relevance of HLA-B27 in host defense and as a pathogenetic molecule in spondyloarthritis. PMID- 15657946 TI - Procoagulant phenotype of endothelial cells after coculture with biomaterial treated blood cells. AB - Understanding endothelial cell (EC)/blood/biomaterial interactions is crucial for the advancement of cardiovascular devices that often fail because of the lack of nonthrombogenic biomaterials. To begin to assess these interactions, a static EC/blood cell/biomaterial model was used. Isolated blood cells were pretreated with model biomaterial beads with different surface chemistries: polystyrene (PS), and PS beads grafted with 3-kDa polyethylene glycol (PEG) with either a hydroxyl (PS-PEG-OH) or amine (PS-PEG-NH(2)) terminal group at 5.4 or 54 x 10(4) beads/mL. Biomaterial-treated monocytes, neutrophils, or platelets were applied to human umbilical vein ECs (HUVECs) for 5 or 24 h of static coculture, and the resultant procoagulant HUVEC phenotype was characterized using several methods. Flow cytometry was used to assess surface expression of tissue factor (TF), adenosine triphosphate diphosphohydrolase, phosphatidylserine, and thrombomodulin, a functional TF assay was used to assess TF activity, and a plasma recalcification assay examined clotting times on HUVECs. Static coculture of HUVEC with biomaterial-treated neutrophils induced a procoagulant phenotype as exemplified by upregulation of TF expression and total functional activity, and downregulation of adenosine triphosphate diphosphohydrolase and thrombomodulin expression. The plasma recalcification assay demonstrated that HUVECs cocultured with biomaterial-treated monocytes significantly shortened clotting times, with some effect of similarly treated neutrophils. PMID- 15657947 TI - A critical role for lipophosphoglycan in proinflammatory responses of dendritic cells to Leishmania mexicana. AB - Recognition of pathogen-associated molecular patterns (PAMP) influences the response of dendritic cells (DC) and therefore development of innate and adaptive immunity. Different forms of Leishmania mexicana have distinct effects on DC, with promastigotes and amastigotes being activating and apparently neutral, respectively. We investigated whether stage-specific differences in surface composition might account for these distinct effects. Amastigotes and promastigotes lacking the lpg1 gene needed for lipophosphoglycan (LPG) biosynthesis could not activate DC in vitro. Genome-wide transcriptional profiling of DC infected with wild-type or mutant promastigotes or wild-type amastigotes revealed that wild-type promastigotes induce an inflammatory signature that is lacking in DC exposed to the other parasite forms. The proinflammatory response pattern was partly recovered by reconstitution of lpg1 expression in lpg1-/- parasites, and exposure to purified LPG increased the expression of MHC class II and CD86 on DC. Infection with wild-type but not lpg1 /- promastigotes increased the number of activated DC in draining lymph nodes, and this was correlated with lower early parasite burdens in wild-type-infected animals. These in vivo and in vitro results suggest an LPG-dependent activation of DC that contributes to host defense and agree with the notion that the parasites evolved under immune pressure to down-regulate PAMP expression in mammalian hosts. PMID- 15657948 TI - Crystal structures and KIR3DL1 recognition of three immunodominant viral peptides complexed to HLA-B*2705. AB - We have solved the crystal structures of three HLA-B*2705-peptide complexes with the immunodominant viral peptides: EBV EBNA3C 258-266 (RRIYDLIEL), influenza (flu) nucleoprotein NP383-391 (SRYWAIRTR), and HIV gag 264-273 (KRWIILGLNK). Long term non-progression during HIV infection has been associated with presentation by HLA-B*2705, and T cell recognition, of the highly immunodominant KRWIILGLNK peptide. The tight hydrogen-bonding network observed between the HLA-B*2705 B pocket and the peptide P2 arginine guanadinium anchor explains why mutation of this residue during HIV infection results in loss of peptide binding, immune escape and progression to AIDS. Prominent, solvent-exposed structures within these peptides may participate in generating T cell responses to these immunodominant epitopes. In the HLA-B*2705 complex with flu NP383-391, the amino acid side chains of residues 4, 7 and 8 are solvent-exposed whilst in the HIV decamer, the main-chain bulges into the solvent around P7. Thus, HLA-B*2705 presents viral peptides in a range of conformations. Tetrameric complexes of HLA B*2705 with the HIV and flu but not EBV peptides bound strongly to the killer-Ig like receptor (KIR)3DL1. Substitution of EBV P8 glutamate to threonine allowed recognition by KIR3DL1. In the HLA-B*2705-EBV structure the P8 glutamate side chain is solvent-exposed and may inhibit KIR3DL1 binding through electrostatic forces. PMID- 15657949 TI - Virus infection-associated bone marrow B cell depletion and impairment of humoral immunity to heterologous infection mediated by TNF-alpha/LTalpha. AB - We previously showed that influenza virus infection of mice induces a depletion of bone marrow B lineage cells due to apoptosis of early B cells mediated by a mechanism involving TNF-alpha/LTalpha. Here we demonstrate that this effect is also observed with acute lymphocytic choriomeningitis virus (LCMV) infection and resulted in a deficiency of both splenic transitional B cells and mature follicular B cells. To determine whether there was an associated impairment of humoral immunity, we infected mice with LCMV and 10 days later at the peak of the B cell depletion, inoculated them with influenza virus. We found that influenza virus-specific antibody titers were dramatically reduced in mice recovering from LCMV infection compared to those in mice infected with influenza virus alone. Further, we showed that there was no reduction of the influenza virus-specific antibody response in LCMV-infected TNF-alpha/LTalpha-deficient mice, suggesting that TNF-alpha/LTalpha-mediated effects on bone marrow and/or peripheral lymphocytes were responsible for the observed impairment in humoral immunity. These results show that the TNF-alpha/LTalpha production induced following infection with diverse viruses has detrimental effects on early B cells in the bone marrow, and may be among the factors that lead to the severely compromised humoral immunity observed to subsequent heterologous infections. PMID- 15657950 TI - Poxvirus semaphorin A39R inhibits phagocytosis by dendritic cells and neutrophils. AB - The poxvirus A39R protein is a member of the semaphorin family that binds to Plexin C1, a molecule expressed on neutrophils and dendritic cells (DC). We previously showed that binding of A39R to Plexin C1 induces local rearrangement of the actin cytoskeleton and inhibits integrin-mediated adhesion, leading to cell retraction. As phagocytosis is dependent on both cytoskeleton integrity and integrin function, we tested the effect of A39R on DC and neutrophil phagocytosis. We found that A39R treatment strongly inhibits phagocytosis by DC and neutrophils in vitro in a Plexin C1-dependent fashion. Moreover, A39R treatment inhibited the capacity of CD8alpha+ DC to take up apoptotic bodies in vivo. As a consequence, A39R impaired the ability of CD8alpha+ DC to cross-prime CD8+ T cells ex vivo. In contrast, A39R had no effect on direct priming of CD8+ T cells by peptide-pulsed CD8alpha+ DC in vitro. These results suggest that poxviruses may use semaphorin homologs as a means to evade the immune system. PMID- 15657951 TI - Characterization of the proximal enhancer element and transcriptional regulatory factors for murine recombination activating gene-2. AB - Recombination-activating gene (RAG)-1 and RAG-2 are essential for V(D)J recombination and are expressed specifically in lymphoid cells. We previously identified two putative enhancer elements, the proximal and distal enhancers, located at -2.6 and -8 kb, respectively, 5' upstream of mouse RAG-2, and characterized the distal enhancer element in detail. In this study, to characterize the proximal enhancer in vitro as well as in vivo, we first defined a 170-bp core enhancer element within the proximal enhancer (Ep) and determined its activity in various cells. Ep conferred enhancer activity only in B-lymphoid cell lines, but not in T- or non-lymphoid cell lines. Analysis of the transgenic mice carrying an EGFP reporter gene linked with Ep revealed that Ep activated the transcription of the reporter gene in bone marrow and spleen, but not in thymus or non-lymphoid tissues. Ep was active in both B220+IgM- and B220+IgM+ subpopulations in the bone marrow and in the B220+ subpopulation in the spleen. Using electrophoretic mobility shift assays and mutational assays, we found that Ikaros and CCAAT/enhancer binding protein cooperatively bind Ep and function as the transcription factors responsible for B cell-specific enhancer activity. These results demonstrate the role of Ep as a cis-regulatory enhancer element for RAG-2-specific expression in B-lymphoid lineages. PMID- 15657952 TI - The immunomodulator FTY720 interferes with effector functions of human monocyte derived dendritic cells. AB - The potent immunomodulator FTY720 elicits immunosuppression via acting on sphingosine 1-phosphate receptors (S1PR), thereby leading to an entrapment of lymphocytes in the secondary lymphoid tissue. To elucidate the potential in vitro effects of this drug on human monocyte-derived DC, we used low nanomolar therapeutic concentrations of FTY720 and phosphorylated FTY720 (FTY720-P) and investigated their influence on DC surface marker expression, protein levels of S1PR and DC effector functions: antigen uptake, chemotaxis, cytokine production, allostimulatory and Th-priming capacity. We report that both FTY720 and FTY720-P reduce chemotaxis of immature and mature DC. Mature DC generated in the presence of FTY720 or FTY720-P showed an impaired immunostimmulatory capacity and reduced IL-12 but increased IL-10 production. T cells cultured in the presence of FTY720- or FTY720-P-treated DC showed an altered cytokine production profile indicating a shift from Th1 toward Th2 differentiation. In treated immature and mature DC, expression levels for two S1PR proteins, S1P1 and S1P4, were reduced. We conclude that in vitro treatment with FTY720 affects DC features that are essential for serving their role as antigen-presenting cells. This might represent a new aspect of the overall immunosuppressive action of FTY720 and makes DC potential targets of further sphingolipid-derived drugs. PMID- 15657953 TI - Effect of hexaflumuron on mortality of the Western subterranean termite (Isoptera: Rhinotermitidae) during and following exposure and movement of hexaflumuron in termite groups. AB - The effects of hexaflumuron consumption on the mortality of workers of the Western subterranean termite, Reticulitermes hesperus Banks, were observed following various exposures to a substrate treated with 14C-labeled hexaflumuron at 5 g kg(-1). Uptake of hexaflumuron by workers was rapid, peaking at approximately 280 ng hexaflumuron per termite 12 days after exposure. The onset of mortality began at day 8, with cumulative mortality reaching approximately 96% by day 45. Excretion of hexaflumuron from worker termites was rapid following various exposure periods, half-lives ranging from 2.1 to 4.7 days. Trophallaxis among worker termites was efficient, hexaflumuron levels in recipient termites approaching two-thirds of levels present in donor termites feeding continuously on hexaflumuron-treated filter paper. The effect of donor-to-recipient group ratios was negligible on the amounts of hexaflumuron transferred from donors to recipients. In laboratory tests, hexaflumuron diffused from a feeding source within 7 days. The presence of additional food sources appeared to inhibit movement of hexaflumuron. Movement of hexaflumuron by cannibalism and coprophagy occurred, but was significant only when termites were starved. Hexaflumuron also appeared to adversely affect egg development by preventing hatch. Hexaflumuron was readily transferred through termite groups, effectively suppressing laboratory populations of R hesperus. Variable efficacy in field situations employing baiting with hexaflumuron in southern California may be a consequence of sporadic feeding at bait stations, the rapid clearance of hexaflumuron from individual termites, and the difficulties in bait presentation (low foraging activity, poor bait station foraging fidelity) inherent in the foraging characteristics of the Western subterranean termite. PMID- 15657954 TI - A novel role for CD28 in thymic selection: elimination of CD28/B7 interactions increases positive selection. AB - While the importance of the CD28/B7 costimulation pathway is well established for mature T cells, the role of CD28 in thymocyte selection is less well defined. The role of CD28 in both negative and positive selection was assessed using H-Y specific TCR-transgenic (Tg) RAG-2-deficient (H-Yrag) mice. Negative selection in male H-Yrag mice was not affected by deficiency in CD28 or B7. Surprisingly, absence of CD28 or B7 in H-Yrag females resulted in increased numbers of CD8 single-positive (SP) thymocytes. The CD8 SP thymocytes found in these females were mature and functionally competent. Furthermore, double-positive (DP) thymocytes from CD28-knockout (CD28KO) or B7.1/B7.2 double-KO (B7DKO) females had higher levels of both CD5 and TCR than those from WT females, consistent with a stronger selecting signal. CD28KO H-Yrag fetal thymic organ cultures also had elevated numbers of thymic CD8 SP cells, reflecting increased thymic differentiation and not recirculation of peripheral T cells. Finally, increased selection of mature CD4 and CD8 SP T cells was observed in non-TCR-Tg CD28KO and B7DKO mice, indicating that this function of CD28-B7 interaction is not unique to a TCR-Tg model. Together these findings demonstrate a novel negative regulatory role for CD28 in inhibiting differentiation of SP thymocytes, probably through inhibition of thymic selection. PMID- 15657956 TI - Comparative acaricide susceptibility and detoxifying enzyme activities in field collected resistant and susceptible strains of Tetranychus urticae. AB - A field-collected strain (MR-VL) of the two-spotted spider mite, Tetranychus urticae Koch, exhibited strong resistance to bifenthrin, dicofol and fenbutatin oxide in comparison with a susceptible laboratory strain (LS-VL). The MR-VL strain was screened for cross-resistance with several currently used acaricides. Cross-resistance was detected with clofentezine (RR = 2631), dimethoate (RR = 250), chlorfenapyr (RR = 154), bromopropylate (RR = 25), amitraz (RR = 17), flucycloxuron (RR = 15) and azocyclotin (RR = 7). Abamectin, acequinocyl, bifenazate, tebufenpyrad and spirodiclofen did not show any signs of cross resistance. Enhanced detoxification by increased activity of mono-oxygenases (MO) and esterases is at least partially responsible for the observed resistance and cross-resistance. MO assays with 7-ethoxycoumarin (7-EC) were optimised and 7 ethoxy-4-trifluoromethylcoumarin (7-EFC), a new MO-substrate, was evaluated for the first time in T urticae and proved to be a good alternative to 7-EC. Approximately 3- and 4-fold higher MO activity was detected with 7-EFC and 7-EC respectively in the MR-VL strain. Kinetic parameters of general esterase assays with 4-nitrophenyl acetate and 1-naphthyl acetate as substrate indicated that more esterases were present in the MR-VL strain. A first attempt was made to classify the esterases present in T urticae. Acetyl-, aryl- and carboxyl esterases were detected with the use of inhibitors after separation by native PAGE. Glutathione-S-transferases did not seem to play any role in the observed resistance and no differences were detected when the general oxidative capacities of the two strains were compared. PMID- 15657958 TI - Pushing the treatment envelope for chronic hepatitis C--is more necessarily better? PMID- 15657957 TI - Involvement of a sodium channel mutation in pyrethroid resistance in Cydia pomonella L, and development of a diagnostic test. AB - Populations of the codling moth, Cydia pomonella L (Lepidoptera, Tortricidae) have developed resistance to several classes of insecticide such as benzoylureas, juvenile hormone analogues, ecdysone agonists and pyrethroids, but the corresponding resistance mechanisms have not been extensively studied. Knockdown resistance (kdr) to pyrethroid insecticides has been associated with point mutations in the para sodium channel gene in a great variety of insect pest species. We have studied two susceptible strains (S and Sv) and two resistant strains (Rt and Rv) of C pomonella that exhibited 4- and 80-fold resistance ratios to deltamethrin, respectively. The region of the voltage-dependent sodium channel gene which includes the position where kdr and super-kdr mutations have been found in Musca domestica L was amplified. The kdr mutation, a leucine-to phenylalanine replacement at position 1014, was found only in the Rv strain. In contrast, the super-kdr mutation, a methionine-to-threonine replacement at position 918, was not detected in any C pomonella strain. These data allowed us to develop a PCR-based diagnostic test (PASA) to monitor the frequency of the kdr mutation in natural populations of C pomonella in order to define appropriate insecticide treatments in orchards. PMID- 15657962 TI - 7Li, 31P, and 1H pulsed gradient spin-echo (PGSE) diffusion NMR spectroscopy and ion pairing: on the temperature dependence of the ion pairing in Li(CPh3), fluorenyllithium, and Li[N(SiMe3)2] amongst other salts. AB - 7Li, 31P, and 1H variable-temperature pulsed gradient spin-echo (PGSE) diffusion methods have been used to study ion pairing and aggregation states for a range of lithium salts such as lithium halides, lithium carbanions, and a lithium amide in THF solutions. For trityllithium (2) and fluorenyllithium (9), it is shown that ion pairing is favored at 299 K but the ions are well separated at 155 K. For 2 lithio-1,3-dithiane (13) and lithium hexamethyldisilazane (LiHMDS 16), low temperature data show that the ions remain together. For the dithio anion 13, a mononuclear species has been established, whereas for the lithium amide 16, the PGSE results allow two different aggregation states to be readily recognized. For the lithium halides LiX (X = Br, Cl, I) in THF, the 7Li PGSE data show that all three salts can be described as well-separated ions at ambient temperature. The solid state structure of trityllithium (2) is described and reveals a solvent separated ion pair formed by a [Li(thf)4]+ ion and a bare triphenylmethide anion. PMID- 15657963 TI - The rich stereochemistry of eight-vertex polyhedra: a continuous shape measures study. AB - A stereochemical study of polyhedral eight-vertex structures is presented, based on continuous shape measures (CShM). Reference polyhedra, shape maps, and minimal distortion interconversion paths are presented for eight-vertex polyhedral and polygonal structures within the CShM framework. The application of these stereochemical tools is analyzed for several families of experimental structures: 1) coordination polyhedra of molecular transition-metal coordination compounds, classified by electron configuration and ligands; 2) edge-bonded polyhedra, including cubane structures, realgar, and metal clusters; 3) octanuclear transition-metal supramolecular architectures; and 4) coordination polyhedra in extended structures in inorganic solids. Structural classification is shown to be greatly facilitated by these tools, and the detection of less common structures, such as the gyrobifastigium, is straightforward. PMID- 15657964 TI - From discrete particles to spherical aggregates: a simple approach to the self assembly of Au colloids. AB - Here we demonstrate a simple, template-free approach to the formation of spherical gold aggregates through the reduction of HAuCl4 by NaBH4, in the presence of cysteine (Cys) as a capping agent. The resulting aggregates are quite stable in solution. The pH of the solution and the molar ratio of Au:Cys are two key empirical factors in the formation of such highly ordered aggregates. At slightly alkaline pH (7-10) and with Au:Cys ratios ranging from 1:0.5 to 1:2, spherical Au aggregates of 30-80 nm are formed. At lower Cys ratios (Au:Cys> or =1:0.5) very loosely linked aggregates are formed; however, at very high Cys ratios (Au:Cys< or =1:3), highly dispersed Au particles of 2-4 nm are obtained, which are virtually indistinguishable from the original colloidal form. Aggregate size is influenced markedly by component concentration; a 3-fold increase in standard levels resulted in Au spherical aggregates of a larger size, 200-500 nm. In addition, we used a combination of Cys and lysine (Lys) as a capping agent/cross-linker and found that the morphology of the Au colloid aggregates can be easily manipulated from a linear to a spherical form by adjusting the proportions of Cys and Lys in the capping agent/cross-linker mixture. The introduction of mercapto (SH)-containing organic acids reduced the cross-linking ability of Cys, especially in the case of long-chain acids. Complete disruption of the spherical aggregates highlights the importance of Cys per se. An explanation of this ordered self-assembly process is proposed, in the context of the known surface chemistry of Au colloids. PMID- 15657966 TI - Catalysts for cross-coupling reactions with non-activated alkyl halides. AB - Despite the problems inherent to metal-catalyzed cross-coupling reactions with alkyl halides, these reactions have become increasingly important during the last few years. Detailed mechanistic investigations have led to a variety of novel procedures for the selective cross-coupling of non-activated alkyl halides bearing beta hydrogen atoms with a variety of organometallic nucleophiles under mild reaction conditions. This Minireview highlights selected examples of metal catalyzed coupling methods and is intended to encourage chemists to exploit the potential of these approaches in organic synthesis. PMID- 15657967 TI - Dithiol proteins as guardians of the intracellular redox milieu in parasites: old and new drug targets in trypanosomes and malaria-causing plasmodia. AB - Parasitic diseases such as sleeping sickness, Chagas' heart disease, and malaria are major health problems in poverty-stricken areas. Antiparasitic drugs that are not only active but also affordable and readily available are urgently required. One approach to finding new drugs and rediscovering old ones is based on enzyme inhibitors that paralyze antioxidant systems in the pathogens. These antioxidant ensembles are essential to the parasites as they are attacked in the human host by strong oxidants such as peroxynitrite, hypochlorite, and H2O2. The pathogen protecting system consists of some 20 thiol and dithiol proteins, which buffer the intraparasitic redox milieu at a potential of -250 mV. In trypanosomes and leishmania the network is centered around the unique dithiol trypanothione (N1,N8 bis(glutathionyl)spermidine). In contrast, malaria parasites have a more conservative dual antioxidative system based on glutathione and thioredoxin. Inhibitors of antioxidant enzymes such as trypanothione reductase are, indeed, parasiticidal but they can also delay or prevent resistance against a number of other antiparasitic drugs. PMID- 15657968 TI - Single-triggered trimeric prodrugs. PMID- 15657969 TI - Biomimetic total synthesis of gambogin and rate acceleration of pericyclic reactions in aqueous media. PMID- 15657970 TI - Enantioselective organocatalyzed alpha sulfenylation of aldehydes. PMID- 15657971 TI - Highly efficient dynamic kinetic resolution of azlactones by urea-based bifunctional organocatalysts. PMID- 15657974 TI - A relational database application in support of integrated neuroscience research. AB - The development of relational databases has significantly improved the performance of storage, search, and retrieval functions and has made it possible for applications that perform real-time data acquisition and analysis to interact with these types of databases. The purpose of this research was to develop a user interface for interaction between a data acquisition and analysis application and a relational database using the Oracle9i system. The overall system was designed to have an indexing capability that threads into the data acquisition and analysis programs. Tables were designed and relations within the database for indexing the files and information contained within the files were established. The system provides retrieval capabilities over a broad range of media, including analog, event, and video data types. The system's ability to interact with a data capturing program at the time of the experiment to create both multimedia files as well as the meta-data entries in the relational database avoids manual entries in the database and ensures data integrity and completeness for further interaction with the data by analysis applications. PMID- 15657975 TI - Graph theoretical characterization and tracking of the effective neural connectivity during episodes of mesial temporal epileptic seizure. AB - Via a detailed case study of mesial temporal lobe epilepsy, we show that a method of determining the direction of information flow among signals is able to provide focal localization via the simultaneous analysis of multiple EEG channels. This determination is accomplished by representing information flow direction via directed graphs, where focal electrodes are associated with high observed rates of pertinence to strongly connected subgraphs. Further clinical support to this finding is provided by results for an additional 9 cases of focal epilepsy cases. The graph theoretical approach is a tool for describing and analyzing the effective connectivity dynamics behind epileptic seizures and may provide a common language for studying other complex dynamic relationships between neural structures. PMID- 15657976 TI - Dynamics of everyday life: rigorous modular modeling in neurobiology based on Bloch's dynamical theorem. AB - Natural, everyday sensorimotor behaviors, such as rising from sitting, typically have an intrinsic organization of several levels of analysis. Taking this intrinsic organization as key to understanding neural dynamics is neither a top down nor a bottom-up approach, but rather a meshing of multiple centers and levels of analysis. Motor control requires body dynamics that are consistent with physical dynamics, besides the more microscopic levels of neural dynamics. The dynamics of separate movements have been investigated as if the ends can be capped off, separated from the rest of the individual's life. Is this dynamically correct? Even chaotic behavior is deterministic. However, the mathematics of nonlinear oscillations is not all of dynamics. This paper relates Bloch's dynamical theorem to the modular, conditional approach to sensorimotor and other neural functioning. Bloch's dynamical theorem lays a foundation for the piecewise study of structurally accurate dynamics in theoretical neurobiology. Piecewise studies can be used as a modeling option complementary to the methods of nonlinear oscillator dynamics. By applying Bloch's theorem, dynamics of movements analyzed piecewise can be extended into a smooth flow on any manifold, either as a whole or conditionally. Conditional dynamics makes dynamical modeling options explicit, often depending on what variables the organism can control, and allows one to take different modeling options at different junctures in analyzing the same phenomenon. For example, this approach allows the study of complex motor control problems to be reduced to modular constructions using singularities and flow lines. Dynamical contingencies are expressed using the mathematics of ordered structures. This paper presents Bloch's dynamical theorem and its relevance to model construction in theoretical neurobiology. Specific examples, integrated into physiological and behavioral context, are cited from the literature. PMID- 15657977 TI - Automated morphometric analysis of the cat retinal alpha/Y, beta/X and delta ganglion cells using wavelet statistical moment and clustering algorithms. AB - Computational morphological analysis comprises the development of measures (indicators) that describe different form attributes of a neuron and provides additional parameters for classification algorithms. Our work addressed the problem of small group sizes often encountered in neuromorphological and neurophysiological research, automated classification tasks (unsupervised learning) and introduced a new morphological measure: the wavelet statistical moment. We analysed cat alpha/Y, beta/X and delta Golgi-stained retinal ganglion cells using six different shape features (circularity, 2(nd) statistical moment and entropy of Gaussian blurred images, wavelet statistical moment, number of terminations and the fractal dimension). This allowed us to compare the sensitivity of the methods in uniquely describing morphological attributes of these cells. PMID- 15657978 TI - Recalibration of somesthetic plantar information in the control of undisturbed upright stance maintenance. AB - To assess the effects of changes in somesthetic plantar information on upright quiet stance, a rotary plantar massage was applied under the feet of healthy subjects for ten minutes. The controlling variable, the centre of pressure (CP) displacements, were recorded, before and after massage, through a force platform and decomposed into two elementary motions: the vertical projection of the centre of gravity (CG(v)) and the difference between the latter and the CP (CP-CG(v)) along medio-lateral ML and antero-posterior AP directions. These motions were processed through frequency analysis and modelled as fractional Brownian motion. For CP-CG(v) motions, the frequency analysis shows that massage under the plantar soles induces a decrease of the amplitudes along the ML direction suggesting reduced overall muscular activity (abductor-adductor muscles of the hip according to Winter et al.). A general trend is that the CG(v) amplitudes are also diminished after massage especially in the ML direction, indicating a better distribution of the body weight on the two supports. On the other hand, the effects tend to vanish after about 8 minutes. Conversely, when the massage was given under the toes, no particular effect on any elementary motion was observed, suggesting that the plantar mechanoreceptors under the toes necessitate stronger stimulation to respond significantly and/or that the greater sensitivity obtained was not used by the CNS. Overall, this data emphasises the fact that a recalibration of somesthetic cues may occur when enhanced afferent information is fed to the postural system. PMID- 15657979 TI - Neurophysiologically-based mean-field modelling of tonic cortical activity in post-traumatic stress disorder (PTSD), schizophrenia, first episode schizophrenia and attention deficit hyperactivity disorder (ADHD). AB - A recently developed quantitative model of cortical activity is used that permits data comparison with experiment using a quantitative and standardized means. The model incorporates properties of neurophysiology including axonal transmission delays, synaptodendritic rates, range-dependent connectivities, excitatory and inhibitory neural populations, and intrathalamic, intracortical, corticocortical and corticothalamic pathways. This study tests the ability of the model to determine unique physiological properties in a number of different data sets varying in mean age and pathology. The model is used to fit individual electroencephalographic (EEG) spectra from post-traumatic stress disorder (PTSD), schizophrenia, first episode schizophrenia (FESz), attention deficit hyperactivity disorder (ADHD), and their age/sex matched controls. The results demonstrate that the model is able to distinguish each group in terms of a unique cluster of abnormal parameter deviations. The abnormal physiology inferred from these parameters is also consistent with known theoretical and experimental findings from each disorder. The model is also found to be sensitive to the effects of medication in the schizophrenia and FESz group, further supporting the validity of the model. PMID- 15657980 TI - Sacrococcygeal teratoma. PMID- 15657981 TI - Treating the injured and burying the dead. PMID- 15657982 TI - Financing the relief effort. PMID- 15657983 TI - Lauriston Taylor. PMID- 15657984 TI - A liver transplant recipient with an infected explanted liver. PMID- 15657985 TI - After the tsunamis. PMID- 15657987 TI - Sepsis induces the transcription of the glucocorticoid receptor in skeletal muscle cells. PMID- 15657986 TI - New insights into DNA triplexes: residual twist and radial difference as measures of base triplet non-isomorphism and their implication to sequence-dependent non uniform DNA triplex. AB - DNA triplexes are formed by both isomorphic (structurally alike) and non isomorphic (structurally dissimilar) base triplets. It is espoused here that (i) the base triplet non-isomorphism may be articulated in structural terms by a residual twist (Delta(t) degrees), the angle formed by line joining the C1'...C1' atoms of the adjacent Hoogsteen or reverse Hoogsteen (RH) base pairs and the difference in base triplet radius (Delta(r) A), and (ii) their influence on DNA triplex is largely mechanistic, leading to the prediction of a high (t + Delta(t))degrees and low (t - Deltat)degrees twist at the successive steps of Hoogsteen or RH duplex of a parallel or antiparallel triplex. Efficacy of this concept is corroborated by molecular dynamics (MD) simulation of an antiparallel DNA triplex comprising alternating non-isomorphic G*GC and T*AT triplets. Conformational changes necessitated by base triplet non-isomorphism are found to induce an alternating (i) high anti and anti glycosyl and (ii) BII and an unusual BIII conformation resulting in a zigzag backbone for the RH strand. Thus, base triplet non-isomorphism causes DNA triplexes into exhibiting sequence-dependent non-uniform conformation. Such structural variations may be relevant in deciphering the specificity of interaction with DNA triplex binding proteins. Seemingly then, residual twist (Delta(t) degrees) and radial difference (Deltar A) suffice as indices to define and monitor the effect of base triplet non isomorphism in nucleic acid triplexes. PMID- 15657988 TI - In memoriam Edward Byrd Truitt, Jr., PhD 1922-2004. PMID- 15657989 TI - [Prevention of thromboembolism in surgery]. AB - Prophylactic treatment against deep vein thrombosis has become a routine part of surgical treatment. The indications and the form of prophylaxis selected depend on the patient's individual risk profile, which is determined in turn by a combination of exposing and predisposing risk factors. The exposing risk factors depend on the type of surgery and trauma the patient is exposed to, while the predisposing risks are determined by factors peculiar to the patient. This review deals with the modalities of prophylaxis current-ly available, pharmacological details relating to these, and their clinical significance. In addition, evidence based data, recommendations for the duration of prophylaxis derived from official guidelines, and medicolegal aspects are discussed. The development of new anticoagulants is expanding the range of prophylactic methods, which means further in-formation is needed. PMID- 15657990 TI - [Stop smoking]. PMID- 15657991 TI - [Instruments, books and other objects memorable to (almost) forgotten opinions, therapies buildings etc: insemination apparatus]. PMID- 15657992 TI - Three steps to effective volunteer management. Simply asking people to help may not be enough. PMID- 15657994 TI - They answered the call: Nebraska nurses join the ranks in World War II. AB - On December 7, 1941, there were fewer than 1,000 nurses in the Army Nurse Corps. That infamous day, 82 of those brave nurses were stationed in Hawaii. Their bravery, leadership and calmness under extreme duress foreshadowed the amazing role nurses would play in World War II. In the months and years that followed Japan's bombing of Pearl Harbor, over 59,000 American nurses would answer the call and join the Army Nurse Corps. Courageous Nebraskan women were among them. PMID- 15657997 TI - Multi-family group therapy may hold the key to long-term recovery from anorexia nervosa. All in the family. PMID- 15657998 TI - Homerton Hospital is home to a unique collection of outsider art. Tate east. PMID- 15658000 TI - Disability or symptoms, which comes first? PMID- 15657999 TI - This is slow advocacy--not like the advocacy where our partners are able to tell us what they want. PMID- 15658001 TI - Enhanced induction of the immunoproteasome by interferon gamma in neurons expressing mutant Huntingtin. AB - Huntington disease (HD) inclusions are stained with anti-ubiquitin and anti proteasome antibodies. This, together with proteasome activity studies on transfected cell, suggested that alterations in the ubiquitin-proteasome system (UPS) might contribute to HD pathogenesis. In previous work we reported that in a conditional mouse model of Huntington's disease (HD94 mice), the chymiotrypsin- and trypsin-"like" activities of the proteasome are increased selectivity in the affected and aggregate-containing brain regions: striatum, and cortex. Moreover, in these areas a neuronal increase in the interferon-inducible subunits of the immunoproteasome LMP2 and LPM7 was observed. In order to test if the expression of N-terminal mutant huntingtin (htt) by itself is sufficient to induce the change in proteasome catalytic activities as well as in LMP2 subunit expression, we performed activities of the proteasome and western blot experiments in striatal cultured neurons from HD94 mice free of glial contamination. We found no changes in any of the activities in these cells. Furthermore, western blot analysis performed with specific antibody against LMP2 subunits, revealed no difference in levels of this subunit in striatal neurons from HD94 compared to control cultures were treated with interferon-gamma (IFN-gamma) during 72 hours, a clear increase in LMP2 levels was observed in control neuronal cultures. Interestingly, this increase was much more pronounced (95% higher) in HD94 striatal cultures. These results indicate that although expression of mutant htt is not sufficient to induce the changes in proteasome catalytic core observed in HD, it synergizes the changes induced by IFN-gamma. Furthermore, immunocytochemical studies revealed that HD94 striatal neuron expressing high levels of LMP2 subunit showed a pre-apoptotic appearance. These results suggest that the correlation between neuronal induction of the immunoproteasome and neurodegeneration found in HD brains is secondary to inflammatory processes. PMID- 15658003 TI - The tenth anniversary of the death of Severo Ochoa. PMID- 15658004 TI - Sex, drugs and cardiac care. Interview by David Crouch. PMID- 15658002 TI - Stress kinases involved in tau phosphorylation in Alzheimer's disease, tauopathies and APP transgenic mice. AB - Hyperphosphorylation and accumulation of tau in neurons (and glial cells) is one of the main pathologic hallmarks in Alzheimer's disease (AD) and other tauopathies, including Pick's disease (PiD), progressive supranuclear palsy, corticobasal degeneration, argyrophilic grain disease and familial frontotemporal dementia and parkinsonism linked to chromosome 17 due to mutations in the tau gene (FTDP-17-tau). Recent studies have shown increased expression of select active kinases, including stress-activated kinase, c-Jun N-terminal kinase (SAPK/JNK) and kinase p38 in brain homogenates in all the tauopathies. Strong active SAPK/JNK and p38 immunoreactivity has been observed restricted to neurons and glial cells containing hyperphosphorylated tau, as well as in dystrophic neurites of senile plaques in AD. Moreover, SAPK/JNK- and p38-immunoprecipitated sub-cellular fractions enriched in abnormal hyperphosphorylated tau have the capacity to phosphorylate recombinat tau and c-Jun and ATF-2 which are specific substrates of SAPK/JNK and p38 in AD and PiD. Interestingly, increased expression of phosphorylated SAPK/JNK and p38 in association with hyperphosphorylated tau containing neurites have been observed around betaA4 amyloid deposits in the brain of transgenic mice (Tg2576)carrying the double APP Swedish mutation. These findings suggest that betaA4 amyloid has the capacity to trigger the activation of stress kinases which, in turn, phosphorylate tau in neurites surrounding amyloid deposits. Reduction in the amyloid burden and decreased numbers of amyloid plaques but not of neurofibrillary degeneration has been observed in the brain of two AD patients who participated in an amyloid-beta immunization trial. Activation of stress kinases SAPK/JNK and p38 were reduced together with decreased tau hyperphosphorylation of aberrant neurites in association with decreased amyloid plaques. These findings support the amyloid cascade hypothesis of tau phosphorylation mediated by stress kinases in dystrophic neurites of senile plaques but not that of neurofibrillary tangles and neuropil threads in AD. PMID- 15658005 TI - Bilateral volume reduction of the insular cortex in patients with schizophrenia: a volumetric MRI Study. AB - The morphologic changes of the insular cortex have been described in schizophrenia, but with inconsistencies between reports. We investigated the insular cortex volume by magnetic resonance imaging in 59 schizophrenia patients (31 males, 28 females) and 62 age- and gender-matched healthy controls (31 males, 31 females). The insular cortex volume was measured on consecutive coronal 1-mm slices. Volumes of the left and right insular cortex were significantly reduced in schizophrenia patients compared with control subjects. There were no effects of gender on the insular cortex volume in the patient group or control subjects. Bilateral insular cortex volumes were correlated negatively with illness duration in the patient group. The findings of this study suggest that there is a possible progressive loss of the gray matter volume of the bilateral insular cortices subsequent to the onset of schizophrenia. PMID- 15658007 TI - Diabetic macular edema: an OCT-based classification. AB - PURPOSE: More than ten years after ETDRS, Optical Coherence Tomography (OCT) greatly enhanced our ability to detect macular thickening and has brought new insights on the morphology of edema and on the presence of vitreal traction. In this study we propose a new classification of macular edema based on OCT findings to better catalogue and follow this complex clinical entity. METHODS: Since January 2000 we analysed with OCT 2 (Zeiss Inc.) more than one thousand and two hundred eyes with DME. RESULTS: The classification takes into account five parameters: retinal thickness, diffusion, volume, morphology and presence of vitreous traction. Standard figures and numerical values for every parameter are given. CONCLUSION: Although ETDRS guidelines for laser treatment of DME still remain the only proven therapy for this condition, many other strategies are now on trial, and the vast majority of authors use OCT as the best indicator of therapeutic benefit. The amount of information given by OCT demonstrates that macular edema is a complex clinical entity with various morphology and gravity, and disclaimed the limitations of a simple "clinical" definition. As in many other examples such as macular holes and choroidal neovascularization, a uniform and precise definition of macular edema would increase the possibility to compare and judge the result of different therapeutic strategies. Aim of this classification is to implement the ETDRS clinical definition of DME with the precise and useful data given by OCT to better diagnose, catalogue and follow macular edema. PMID- 15658008 TI - [Ultrasonography in the urinary tract: understand the need well]. PMID- 15658009 TI - [Recent studies on atypical psychoses]. PMID- 15658006 TI - Diode laser, vitrectomy and intravitreal triamcinolone. A comparative study for the treatment of diffuse non tractional diabetic macular edema. AB - PURPOSE: To determine the effectiveness and safety of diode laser, pars plana vitrectomy (PPV) with gas tamponade, and intravitreal triamcinolone (IVT) as possible treatments for diffuse diabetic macular edema (dDME). To determine whether the new macular edema Classification previously proposed by some of the authors may help as a guide in the choice of treatment. METHODS: A retrospective, comparative study of 169 eyes with dDME that underwent treatment. The eyes divided into 3 groups: in the PPV Group, 59 eyes received PPV with gas tamponade; in the DIODE Group, 53 eyes received a laser grid; in the IVT Group, 57 eyes received an intravitreal injection of 4 mg of triamcinolone acetonide. The follow up ranged from 6 to 24 months. The eyes were classified according to the new DME Classification based on OCT. MAIN OUTCOMES: Change in foveal thickness as determined by Optical Coherence Tomography (OCT); change in visual acuity; intra and postoperative complications. RESULTS: Mean visual acuity (VA) improved at 3 months with every treatment. At one year only eyes which underwent PPV still had better VA than the pre-op value, while eyes which underwent IVT and laser treatment showed regression. PPV show the greater percentage of eyes which gain 3 or more lines of VA at one year (17%). Mean foveal thickness improved at 3 months with every treatment. Only the eyes into the PPV Group showed relatively low foveal thickness at one year. Better final VA and foveal thickness were obtained if preop VA is > or = 0.3 and if earlier stages of DME were treated according to the new DME Classification. PPV was the treatment which offered the most stable results with at one year or longer. Similar results were observed into the PPV Groups as a whole and into a subgroup of eyes with preoperative PVD. No complications were encountered with laser diode treatment. Long term complications into the IVT Group were elevated IOP (8%), retinal detachment (3.5%) and posterior cataract (15%). Long term complications into the PPV Group were retinal detachment (3.4%) and cataract (90%). CONCLUSIONS: Diode laser, PPV with gas tamponade and IVT are effective alternative treatments to decrease foveal thickness and improve visual acuity in eyes with DME. However while the results of PPV are stable in the long term follow-up, diode laser and IVT do not offer stable results. Complications may be severe with PPV and IVT. It is necessary to carefully select cases which would benefit from these types of treatments. The authors think that the OCT Classification may serve as a guide for the choice of treatment. PMID- 15658010 TI - [Prognosis in olfaction disorders]. PMID- 15658011 TI - Designing babies: human research issues. PMID- 15658012 TI - Commercial tissue repositories: HIPAA raises sponsors' fears. PMID- 15658014 TI - Correction and clarification. PMID- 15658013 TI - Quebec physicians' knowledge and opinions regarding substitute consent for decisionally incapacitated older adults. PMID- 15658015 TI - Some precision would be helpful. PMID- 15658016 TI - By any other name: the many iterations of "patient advocate" in clinical research. PMID- 15658017 TI - Costs to subjects for research participation and the informed consent process: regulatory and ethical considerations. PMID- 15658018 TI - So what are we going to do about research using clinical information and samples? PMID- 15658019 TI - Global health and the scientific research agenda. PMID- 15658020 TI - Queries on nutrition and hydration. PMID- 15658022 TI - No single integrating organ? PMID- 15658021 TI - A future ethical issue. PMID- 15658023 TI - Pro-life nurses and abortion. PMID- 15658024 TI - Washington insider. PMID- 15658025 TI - The role of the priest in bioethical decision making. PMID- 15658026 TI - Managed care and justice: compatible or adversarial in achieving the ends of medicine? PMID- 15658027 TI - The moral flaw in the "pro-choice" position. PMID- 15658028 TI - Immediate hominization from the systems perspective. PMID- 15658029 TI - Oregon's oxymoron: the Death with Dignity Act. PMID- 15658030 TI - On the status of parthenotes: defining the developmental potentiality of a human embryo. PMID- 15658031 TI - Reflections on artificial nutrition and hydration: colloquium of the Canadian Catholic Bioethics Institute (CCBI). PMID- 15658032 TI - Protecting patients from unethical, incompetent and criminal acts by health care practitioners. PMID- 15658033 TI - [Myocardial scintigraphy for the assessment of perfusion in diabetic patients]. PMID- 15658034 TI - Direct-to-consumer genetic tests, government oversight, and the First Amendment: what the government can (and can't) do to protect the public's health. PMID- 15658035 TI - [Newly emerging infections: Tetanus]. PMID- 15658037 TI - [General cognitive ability. Current state and prospects for further investigations]. AB - The paper reviews general cognitive ability (g), the ways of its measurement, and genetic and behavioural aspects of its investigations. The criteria for assessment of cognitive abilities in animals are also suggested and described. Based on these criteria the new behavioural methods and procedures are developed for investigation of general cognitive ability. PMID- 15658036 TI - Voting by residents of nursing homes and assisted living facilities: State law accommodations. PMID- 15658038 TI - [A possible mechanism of participation of dopaminergic cells and striatal cholinergic interneurons in the conditioned selection of motor activity]. AB - A possible mechanism of participation of cholinergic striatal interneurons and dopaminergic cells in conditioned selection of a certain types of motor activity is proposed. This selection is triggered by simultaneous increase in the activity of dopaminergic cells and a pause in the activity of cholinergic interneurons in response to a conditioned stimulus. This pause is promoted by activation of striatal inhibitory interneurons and action of dopamine at D2 receptors on cholinergic cells. Opposite changes in dopamine and acetylcholine concentration synergistically modulate the efficacy of corticostriatal inputs, modulation rules for the "strong" and "weak" corticostriatal inputs are opposite. Subsequent reorganization of neuronal firing in the loop cortex--basal ganglia--thalamus- cortex results in amplification of activity of the group of cortical neurons that strongly activate striatal cells, and simultaneous suppression of activity of another group of cortical neurons that weakly activate striatal cells. These changes can underlie a conditioned selection of motor activity performed with involvement of the motor cortex. As follows from the proposed model, if the time delay between conditioned and unconditioned stimuli does not exceed the latency of responses of dopaminergic and cholinergic cells (about 100 ms), conditioned selection of motor activity and learning is problematic. PMID- 15658039 TI - [Recognition of emotions based on facial expressions in subjects with different personality traits]. AB - A reaction time and accuracy of visual recognition of emotions of joy, anger and fear in their relation to personality traits was studied in 68 healthy subjects. According to scores of Kettell Questionnaire all the participants were divided into two groups: emotionally unstable and emotionally stable, which differed in their emotional and communication traits. It was shown that in the stable group recognition of fear was significantly worse and more slowly than in the unstable group. Besides, the emotionally stable subjects recognized the frightened facial expression less accurate and slowly than they did the joyous and threatening ones. The reaction time and recognition level was found to be closely correlated with some personality traits. These traits were different in two groups and differed from data in the control session of gender recognition. The conjunction between recognition of fearful facial expression and the personality traites and its adaptive significance were discussed. The data seems to be essential for understanding of individual strategy of communication. PMID- 15658040 TI - [Participation of the motor cortex in the bimanual unloading task: a study by transcranial magnetic stimulation]. AB - Motor potentials of m. biceps brachii evoked by transcranial magnetic stimulation of the contralateral motor cortex have been recorded in postural adjustment during arm unloading in humans. During active unloading, the amplitude of the motor evoked potential decreases simultaneously with the decreasing of the muscle activity. During load keeping, the muscle response changes simultaneously with the load changes. When the other arm has lifted the other load during load keeping, the amplitude of the motor evoked potential decreases in the m. biceps of the keeping arm without muscle activity changes. Passive unloading results in the same changes of the motor evoked potential as active unloading. A possible role of the direct corticospinal volley and the motor command mediated by some subcortical structures in the decrease of the muscle activity preceding active unloading (postural adjustment) is discussed. PMID- 15658042 TI - [Impulsation of hypothalamic and amygdalar neurons in bilateral derivations in the state of food motivation]. AB - The character of impulsation of single neurons in the lateral hypothalamic and amygdalar central nucleus of rabbits recorded in bilateral derivations during quiet wakefulness, after 24-h food deprivation, and after satiation was studied by plotting autocorrelation histograms and by counting the mean frequency of discharges. During the transition from hunger to satiation, the character of impulsation of neurons in hypothalamus and amygdala changed in different ways: (1) a greater number of hypothalamic than amygdalar neurons changed their mean discharge frequency (85 versus 56%, respectively); (2) in hunger, the number of hypothalamic neurons with delta-frequency oscillations decreased as compared to quiet wakefulness and satiation, and in the amyglala the number of neurons with beta2-frequency oscillations increased; (3) in hunger, the number of hypothalamic neurons with bursting and periodic discharges decreased and the number of amygdalar neurons with equiprobabilistic discharges increased. During state alternation (according to the autocorrelation histograms) the strongest changes in the character of neuronal discharges took place in the left hypothalamus and left amygdala. The maximum differences in neuronal impulsation between the left and right hypothalamus were observed in the state of hunger and between the left and right amygdala, after satiation. PMID- 15658041 TI - [Simulation of the role of intracortical inhibition as a source of sensitivity to cross-like figures]. AB - Simulation of receptive fields of striate neurons sensitive to cross-like figures under the blockade of intracortical inhibition was performed. It was shown that without the inhibition, a neuron with convergence of signals from two orientation detectors widens its tuning to a cross in such a way that the tuning becomes invariant to the shape and orientation of the cross. Detector of a cross whose orientation is based on the disinhibition mechanisms becomes a bar orientation detector under conditions of inhibition blockade. Another scheme of receptive field is advanced, in which the inhibitory zones mask tuning to cross-like figures, but the blockade of inhibition unveils such sensitivity. We specified the features of the receptive fields (shape, localization, and weight of their zones), which, being applied in simulation, imitate properties of the real striate neurons sensitive to shape and orientation of a cross-like figures. PMID- 15658043 TI - [Neonatal Semax and saline injections induce open-field behavior changes in mice of different genotypes]. AB - DBA/2, CBA mice, and their F1 hybrids (first series) and 101/HY and C3H mice (second series) were injected as neonates (2-7 days of life) with Semax (sc., 7 microg per animal). Semax is a peptide analogue of ACHT4-10 fragment which is resistant to degradation. The common feature of remote effects of both Semax and saline injections was the set of changes in the open-field behavior in adult (2.5 to 3-month-old) animals as compared to intact mice. Unexpectedly, the neonatal saline injections induced many changes in adult behavior, part of these effects being genotype-dependent. The most conspicuous shifts (genotype-dependent increase or decline) in freezing, grooming and rearing scores were displayed by DBA/2 and C3H mice, whereas the hole-poke frequencies were significantly changed in CBA and C3H mice. Squares crossed in the center of arena and rearing number were significantly increased in saline group of DBA/2 mice, whereas in Semax injected DBA/2 group they were approximately equal to the level of intact mice. This means that the remote effects of noxious stimulation (injections of saline) were in some ways "compensated" as the result of concomitant peptide effect. At the same time, the numbers of freezing and grooming episodes were also increased in these groups. Because exploratory behavior and manifestations of anxiety increased or decreased simultaneously, it proves to be difficult to ascribe these changes to behavioral modulation along the "novelty seeking--anxiety" axis. In mice of other genotypes, changes in the same indices of the open-field behavior were revealed, but these changes were different in their direction. It was suggested that the complex patterns of postnatal behavior was the result of neonatal injections modulating subsequent brain development. PMID- 15658044 TI - [Protein synthesis-dependent reactivation of environmental conditioned reflex in terrestrial snails]. AB - We investigated influence of anisomycine injection on reconsolidation of contextual memory after development of environmental conditioned reflex in terrestrial snail Helix. Testing the amplitude of behavioral reactions (tentacle withdrawal) in response to standard tactile stimulation of the skin in two contexts: a) when the snail was fixed by the shell and was moving on the surface of the ball floating in water, or b) was moving on the flat surface of glass terrarium, has shown no difference in response amplitudes. After a session of electric shocks (5 days) in one context only (ball) the associative learning was clearly observed as the significant difference of response amplitudes in two contexts. On the other day following testing was performed a session of "reminding", immediately after which the snails were injected by anisomycine (control snails were injected by saline solution). Testing has shown that injection of anisomycine led to impairment of the context conditioning. Results suggest that the mechanisms of consolidation of new memory and memory reconsolidation after retrieval are not identical. PMID- 15658045 TI - [Effect of changes in extracellular calcium concentration on electrical characteristics of command neurons after defensive reflex conditioning in snail]. AB - Studies of electrical characteristics of command neurons of the defensive reflex have shown that membrane potential does not reliable differ for both naive and learned snails with changing of the calcium concentrations in external solution. With increasing of the concentrations of calcium ions it is observed the increases of threshold potential from 14 + 0.7 mV in 2.5 mM Ca2+ to 21.8 + 0.9 mV in 20 mM Ca2+ for naive snails. Threshold potential decreases from 16.8 + 0.6 mV (saline solution for snail--10 mM Ca2+) to 13.3 + 0.6 mV in 20 mM Ca2+ with increasing of calcium concentrations for learned snails, and under its reduction it is also decreased, but less remarkably, than for naive, to 11.8 + 0.8 mV in 2.5 mM Ca2+. On the same scheme it is changed a critical level of depolarization: for naive snails it decreases with increasing of the calcium concentrations, and for learned snails it increases either with increasing of the calcium concentrations, and so with its reduction. PMID- 15658047 TI - [Natural head-forelimb coordination in dogs after vestibular de-afferentation]. AB - We studied the influence of the vestibular lesion on the natural head--forelimb coordination. This coordination exists in intact dogs at the early stage of acquisition of the instrumental feeding reaction of tonic forelimb flexion in order to hold a cup with meat during eating when the head is bent down to foodwell. In untrained dogs, the forelimb flexion is preceded by lifting the head bent down to the food; the following lowering of the head leads to extension of the flexed forelimb. For performing the instrumental reaction, the innate coordination has to be rearranged into the opposite one. It is achieved only by learning. After the lesion of the primary motor cortex contralateral to the "working" forelimb in trained dogs, the innate coordination reappears, whereas the learned coordination breaks down steadily. It was shown that bilateral vestibular lesion do not disturb the innate coordination in intact dogs at the early stage of learning and in trained dogs after the motor cortex lesion. It was concluded that the studied natural head--forelimb coordination is not connected with the vestibular reflex. PMID- 15658046 TI - [Neuronal activity in a honey bee (Apis mellifera L.) with kynurenine deficiency]. AB - Neuronal activity of the antennal lobes, mushroom bodies, and cervical connective in wild-type honey bees and snowlaranija mutants was recorded at different stages of the ontogeny (on the 1st, 3rd, 7th, and 25th days). The mutation snowlaranija affects the structural gene of tryptophane oxygenase, the first key exzyme in the kynurenine pathway of tryptophane metabolism, and leads to a deficit of kynurenines. Changes in neuronal activity in nutant bees were most pronounced in the cervical connective. A significant decrease in the pulse rate was revealed only in homozygous but not in heterozygous individuals. This finding is in accordance with previously reported inhibitory effect of the mutation at the behavioral level. Less pronounced effects were obtained when the neuronal activity was recorded in the antennal lobes or mushroom bodies. This may be related to a complex character of biochemical changes in different parts of mutants brain. PMID- 15658048 TI - [Comparative analysis of EEG spatial organization in the rats with different level of genetically determinated anxiety under ethanol]. AB - The pecularities of EEG changes in the rats with geneticaly different level of anxiety was studed after ethanol injection. The biopotentials spatial synchronization and entropy in the "high anxious" rats (strain MR) did not increase under low dose; on the contrary the above changes were characteristic for the rats without anxiety signs (strain MNRA). After injection of middle and high ethanol doses the "high anxious" rats as compared with "low anxious" ones had the "simplification" of reaction type--the increase of narrow frequency coherence (i.e. linear biopotentials relationship) and spectral power of theta activity (5.75-7.25 Hz), which was combined with the decrease of nonlinear biopotential characteristics. This fact is interpreted as the predominance of subcortical mechanisms of the brain functional state regulation over cortical ones. The significant decrease of high frequency EEG components in "high anxious" rats takes place under more high ethanol doses, presumably reflecting the increase of the anxiolytic level of ethanol effect. PMID- 15658049 TI - [Effect of acute melipramine administration on motor activity and defensive conditioned reflexes of passive and active avoidance in rats]. AB - The administration of threecyclic antidepressant melipramine to Wistar rats (15 mg/kg, intraperitonaly, 2 h before of experiments) increases time of an "open field" centre leaving. Thus melipramine does not influence horizontal and vertical activity, and also the number of bolus. At the development of a passive avoidance conditioned reflex melipramine significantly slows down realization of a unconditional mink reflex by untrained rats, increasing the latency of call in a dark compartment of the chamber. After training significant deterioration of a reflex reproduction is observed. At research of a defensive conditioned reflex of active avoidance melipramine worsens both development and reproduction of a reflex. The comparative analysis of the literary data of imipramine action on uptake of serotonine and noradrenaline and the analysis of the literary data on a role of these systems in the processes of learning and memory allows to suggest, that the effect of melipramine is connected mainly to amplification activity of serotoninergic system of a brain. It is supposed, that acute administration of melipramine creates emotionally negative state, worsens processes of learning and memory, strengthening mainly activity of a brain serotoninergic system. It specifies that serotoninergic system of a brain is system of punishment. Its activation interferes with formation and consolidation of connections between conditional and unconditional irritation. PMID- 15658050 TI - [The effect of preceding mild hypoxia on the alteration of acquisition and retention of the conditioned passive avoidance behavior caused by severe hypobaric hypoxia in rats]. AB - The purpose of this study was to determine whether mild hypobaric hypoxic preconditioning provides protection against learning deficit caused by subsequent more severe hypoxia insult. Learning was examined using a passive avoidance task. Three groups of Wistar male rats: the intact and exposed to either severe hypoxia (160 Torr, exposition 3 h) or mild hypobaric hypoxic preconditioning (360 Torr, exposition 2 h, repeated three or six times daily) followed by severe hypoxia, were included in this study. In experiment 1 a passive avoidance response was acquired in 15 min immediately after hypoxia. In experiment 2 rats were exposed to hypoxia in 60 min after the acquisition of passive avoidance response. The mild hypobaric hypoxic preconditioning significantly attenuated the hypoxia induced learning deficit in rats in Experiments 1 and 2. In experiment 1 the mild hypobaric hypoxic preconditioning repeated six times was more effective in protection against learning deficit in hypoxia exposed rats than in the case of triple mild hypobaric hypoxic preconditioning. The amount of rats suffered irreversible respiratory arrest was also assessed in this study. It was found that 50% of rats exposed to severe hypoxia died in consequence of this pathology, whereas in rats preconditioned before the severe hypoxia only 15% died for this reason. The overall results indicate that the mild hypobaric hypoxic preconditioning significantly increases CNS resistance to severe hypoxia in rats. PMID- 15658051 TI - [General cognitive ability. Some new approaches in mice]. AB - In this paper some new methods for studying of general cognitive ability, g, and their experimental testing on mice are proposed. Instead of techniques based on learning processes with application of many trials and gradual elaboration of a habit, the proposed methods allow to assess behaviour by using only one or a few trials and to compare such behaviour with conceptual activity. PMID- 15658052 TI - [Psychology of learning, cognitive neurology and ecology of the education medium]. PMID- 15658053 TI - [Image of the month. Pancreatic cancer and ivory vertebra]. PMID- 15658054 TI - [Image of the month. Perineal ulcer in a spinal bifida patient]. PMID- 15658055 TI - [Clinical case of the month. MacLeod syndrome]. AB - MacLeod syndrome is a rare cause of localized hypertransradiancy of the lung. This syndrome is defined by radiological features: localized hypertransradiancy due to oligemia and presence of air-trapping on expiratory chest radiography. Involvement of one entire lung is called "unilateral hyperlucent lung". Whereas the etiology is different, the physiopathology is probably identical to that of the panacinar emphysema of chronic obstructive pulmonary disease. The syndrome is believed to be related to acute bronchiolitis during infancy. Clinical manifestations and prognosis depend mainly on the presence of other lesions due to the same infectious agent like bronchiectasis. Pulmonary function tests, chest CT-scan, ventilation and perfusion scintigraphy and, if necessary, bronchoscopy help the differential diagnosis and detect associated bronchiectasis. PMID- 15658057 TI - [Prostate carcinoma cell lines and apoptosis: a review]. AB - Prostate cancer is a major pathology in industrialized countries. Tumor growth usually results from increased cell proliferation, conjugated with an inhibition of programmed cell death (apoptosis). In this paper, after a short description of the apoptotic mechanisms and their methods of investigation, we review the present knowledge of the implication of different molecular actors in the regulation of apoptosis in prostate cancer cells. This review notably summarizes the present knowledge of the (de)regulation of the effects of androgens, p53, Bcl 2, Bcl-xL, Bax, Akt, PTEN, Par-4, clusterine, caspases and NF-kappaB in prostate adenocarcinoma cell lines and provides an appraisal of their therapeutic potential. A better knowledge of the apoptotic pathways in these cells could indeed allow the development of new selective and effective anti-cancer strategies. PMID- 15658056 TI - [Aspirin: recent advances in cardiovascular prevention]. AB - More than a century after being launched onto the market, aspirin still remains a fascinating drug, both for its demonstrated antalgic, antipyretic, antiinflammatory and antithrombotic properties and, also, for newer, yet conjectural, applications mentioned in recent publications. The role of aspirin, as an irreversible COX-1 inhibitor and antiplatelet agent, is well elucidated and established. Our purpose is to review the value of aspirin for primary and secondary prevention of ischemic cardiovascular events. The clinician constantly has to manage a trade off between the protective effects of aspirin and its possible hemorrhagic, notably gastrointestinal, side-effects. The Task Force of the ESC recommends the use of doses no higher than 75-100 mg/d. New antiplatelet agents (thienopyridin derivatives), which have a totally different mode of action, have been introduced and were compared with aspirin. Although clopidogrel may be slightly superior to the latter, according to the European experts: "the size of any additional benefit is statistically uncertain and the drug has not been granted a claim, of superiority". Economical considerations reinforce this view. Clopidogrel is undoubtedly a good alternative when aspirin is contra indicated, poorly tolerated, or not efficacious. Resistance to aspirin and resistance to clopidogrel have been described. In some high-risk patients, the combined use of aspirin and clopidogrel is deemed justified. PMID- 15658058 TI - [Percutaneous coronary angioplasty in diabetic patients: new prospects with drug eluting stents]. AB - Coronary revascularization procedures are associated with less favourable outcomes in diabetic patients as compared to non-diabetic individuals. Especially, percutaneous coronary angioplasty (PTCA) is associated with a high level of restenosis and recurrent cardiac morbidity and mortality. In diabetic patients, PTCA should ideally be combined with stents. Bare-metal stents reduce by almost half the risk of restenosis, but this favourable effect decreases with the vessel calibre, a common finding in diabetic patients. Drug-eluting stents containing pharmacological agents that can reduce the risk of restenosis (sirolimus, paclitaxel) provide better angiographic results, including in small coronary arteries, and this effect has been shown to be accompanied by significant reduction of both morbidity and mortality. Such preliminary results obtained in the general population (including around 20% of diabetic subjects) deserve further confirmation in a large clinical trial specifically devoted to diabetic patients. Drug-eluting stents may represent a major advance in the management of diabetic patients with coronary heart disease in the near future. PMID- 15658059 TI - [Medical studies at the University of Liege: renewing the art of teaching]. AB - The Faculty of Medicine of the University of Liege has undergone since 2000, a striking change in the medical school, aiming at a better correlation between medical education and community needs and a shift from teaching to learning approach for education. Consequently, teaching matters related to the community medical practice interface have been reinforced. Furthermore, problem based learning has been implemented all along the cursus. PMID- 15658060 TI - [Anatomo-clinical confrontation. Vulvar melanosis]. AB - Vulvar melanosis is a benign disorder that may suggest a malignant melanoma. We report the case of a woman in whom partial vulvectomy was performed to eradicate the melanotic macule. The lesion corresponds to increased accumulation of melanin inside keratinocytes in the absence of any melanocytic neoplasm. PMID- 15658061 TI - [How I explore...significance of ophthalmoscopy in patients with arterial hypertension]. AB - The ocular fundus is the only examination which allows a simple and non invasive visualisation of the terminal vascular system. Feature analysis of fundus examination supposes a similar vascular state in the other organs and permits direct diagnostic, prognostic and therapeutic choices. This article will emphasize the value of an ophthalmoscopy examination in patients with arterial hypertension. PMID- 15658062 TI - Evaluation of schistosomicidal activity of myrrh extract: parasitological and histological study. AB - Ninety mice were divided into three main groups: G. I (non-infected control), G. II (infected non treated control) and G. III (infected treated), which was further subdivided into 4 subgroups: IIIA, IIIB, IIIC & IIID, where the drug was administered in a dose of 500 mg/kg body weight for five days before infection for subgroup IIIA, on the 1st day post infection (PI) for subgroup IIIB, 21 days PI for subgroup IIIC and 45 days post infection for subgroup IIID. All animals were sacrificed 80 days after the start of the experiment. Anti-schistosomal activity was assessed parasitologically by estimating the percentage reducetion of worm burden, egg count/gm tissues (liver & intestine), and the oogram pattern; histologically & histopathologically by examination of jejunum using different stains. The percentage reduction of worm burden was 30.35%, 64.54%, 76.92% and 98.46% respectively when compared to the infected non-treated control G. The maximum effect of the drug was observed in subgroups IIIC & IIID. Also, there was marked reduction in the egg count in tissues (liver & intestine). In addition the oogram pattern showed that myrrh had high antischistosomal activity. The histologically and histopathologically infected untreated (GII), when compared to non-infected non-treated control G. showed apparent shortening and flattening of the jejunal villi with focal loss of the epithelial covering. Loss of PAS positive brush border of many enterocytes with goblet cells hyperplasia was seen. Bilharzial granulomas were frequently encountered in the submucosa and the musculosa with numerous eosinophils content. In subgroups IIIA & IIIB, there was mild amelioration of the mucosal structural abnormalities. The granulomas were less frequently seen with decrease of their eosinophils. In subgroups IIIC & IIID there was restoration of the jejunal mucosal continuity, marked decrease in the granulomas and paucity of eosinophils. The present data proved that myrrh has a valuable schistosomicidal effect against different maturation stages of S. mansoni. The chemotherapeutic effect was more evident when the drug was given to the infected mice on the 21st as well as on the 45th day PI. The drug proved a promising chemoprophylactic agent when used five days before exposure to infection. PMID- 15658064 TI - Detection of IgA anti-Entamoeba histolytica in the patients' saliva. AB - E. histolytica is a protozoan parasite with worldwide distribution. The diagnosis of this parasite still depends on the microscopic examination. Determination of the sIgA in the different body fluid proves to be of diagnostic value in many pathogens. To find out the potentials of salivary sIgA in diagnosis of intestinal amoebiasis, 30 patients with parasitologic proved E. histolytica and 30 cross matched negative controls were selected for levels of salivary sIgA using radial immunodiffuion technique. The result showed a high significant increase in the mean concentration of salivary sIgA in patients rather controls. However, six negative controls gave positive sIgA. So, salivary IgA concentration has a predictive diagnostic values of intestinal amoebiasis, when the stool examination is practically impossible as well as in tissue amoebiasis. PMID- 15658063 TI - Effect of Ferula assafoetida on experimental murine Schistosoma mansoni infection. AB - Ferula assafoetida is a hard, resinous, oily herbaceous gum belonging to plant family Umbelliferae. It is used as a traditional medicine in many parts of the world and its wide use in medicine was listed by many authors. In the current study, the effect of F. assafoetida on Schistosoma mansoni in experimentally infected mice is investigated. F. assafoetida was given orally via intragastric tube in an oil-form and a powder-form in different concentrations. Four test groups of 30 mice each were studied. Gs I & II mice were given F. assafoetida in an oil-form in different concentrations at 4 and 6 weeks post infection (PI) respectively. Mice in Gs III & IV were given re-constituted F. assafoetida powder in different concentrations (conc.) at 4 & 6 weeks PI. respectively like the previous groups. Oil-form F. assafoetida was given at conc. of 50, 25 & 15 mg/ml. Powder-form was given at conc. of 32, 16 & 8 mg/ml. A highly significant statistical difference between the test Gs (I, II, III & IV) was recorded in comparison to the infection control G with p value < 0.0001 and also between powder and oil forms of F. assafoetida (p < 0.0001) as regards the mean worm burden and tissue egg count. The highest reduction in worm burden and egg counts was found with powder form of F. assafoetida (Gs III & IV) when compared to oil form (Gs I & II), as confirmed histopathologically and by ultrastructural profile alteration. PMID- 15658065 TI - Induction of protective antibody response in rabbits against fascioliasis with Toxocara/Fasciola cross- reactive defined antigen. AB - A method of affinity chromatography purification of Toxocara vitulorum antigen cross- reacts with Fasciola gigantica antiserum is described. Characterization of the isolated cross- reactive fraction by SDS-polyacrylamide gel electrophoresis, isoelectric focusing and amino acid analysis resulted in a fraction consists of five polypeptides of 137.7KDa, 81KDa, 75KDa, 48KDa and 21.6KDa with isoelectric points of 8, 7.5, 7.2, 6.7 and 6.6. Seventeen amino acids were identified in the fraction with high proportions of only two of them (tyrosine and glutamic). Rabbits immunization with this identified T. vitulorum cross- reactive antigen in Freund's adjuvant followed by challenge with F. gigantica metacercariae resulted in 60% reduction in worm burden over control infected rabbits. Higher IgG level was detected in vaccinated rabbits four weeks post first immunization than control infected ones and remained high up to the end of the trial. PMID- 15658066 TI - Five years after IOM report, experts gauge progress of patient safety. PMID- 15658067 TI - Billing codes for pharmacy clinical services advance. PMID- 15658068 TI - Interventions to improve compliance with guidelines on surgical prophylaxis. PMID- 15658069 TI - Look-alike, sound-alike drug errors with Reminyl and Amaryl. PMID- 15658070 TI - Medications during the perioperative period. PMID- 15658071 TI - Use of colistin in the treatment of multiple-drug-resistant gram-negative infections. AB - PURPOSE: The use of colistin for the treatment of infections caused by multiple drug-resistant (MDR) gram-negative microorganisms was studied. METHODS: The efficacy of colistin for treating infections caused by MDR gram-negative microorganisms and the development of renal toxicity were studied in hospitalized adult patients in Spain. Patients treated between January 2001 and October 2001 were included. RESULTS: Over the study period, 71 courses of inhaled colistin, 12 courses of i.v. or intramuscular (i.m.) colistin, and 2 courses of intrathecal colistin were administered to 80 patients. All were infected by MDR organisms: 69 (86%) by Acinetobacter baumannii and 11 (14%) by Pseudomonas aeruginosa. In 41 patients (51%), the episodes were caused by A. baumannii strains susceptible exclusively to colistin. The causative organisms were cleared in 92% of the patients from whom posttreatment repeat specimens were obtained. The in-hospital mortality rate was 18% (14 patients). There were no significant changes in mean serum urea or creatinine concentrations in patients receiving i.v. or i.m. therapy. CONCLUSION: Colistin was used in 80 patients infected with A. baumannii or P. aeruginosa and appeared to be efficacious and safe. PMID- 15658072 TI - Bioavailability of intranasal butorphanol administered from a single-dose sprayer. AB - PURPOSE: The bioavailability and tolerability of single doses of intranasal butorphanol tartrate using a single-dose, metered sprayer were studied. METHODS: In this open-label, randomized, three-way crossover study, 24 healthy volunteers received three treatments: (1) 2 mg of i.v. butorphanol (treatment A), (2) 2 mg of intranasal butorphanol (treatment B), and (3) 1 mg of intranasal butorphanol (treatment C). The three treatments received by each subject were separated by six-day washout periods. Venous blood samples (10 mL each) were obtained from an indwelling catheter at 0 (predose), 5, 10, 15,20,30, and 45 minutes and 1,2,3,4,6,8, 12, and 16 hours after butorphanol administration. Pharmacokinetic parameters were determined using standard noncompartmental methods with log linear least-squares regression analysis to determine the elimination-rate constants. RESULTS: Intranasal butorphanol 1 and 2 mg administered using unit dose sprayers had a mean bioavailability of approximately 80%, which is higher than the percentage reported with the commercially available multidose product (61-69%). The absorption of intranasal butorphanol was rapid, with a median time to reach maximum concentration of 20 minutes (range, 10-60 minutes). Elimination profiles were comparable among all treatments. There were no clinically significant changes in the results of physical examinations, nasal evaluations, or laboratory tests related to butorphanol treatment. Most adverse effects reported were mild to moderate and as expected for this drug. CONCLUSION: Single dose intranasal butorphanol was rapidly absorbed and had high absolute bioavailability in healthy volunteers. PMID- 15658073 TI - Utilization of darbepoetin alfa and epoetin alfa for chemotherapy-induced anemia. AB - PURPOSE: The patterns of use and effectiveness of therapy with darbepoetin alfa and epoetin alfa for chemotherapy-induced anemia (CIA) in hospital outpatient and community settings were evaluated. METHODS: Data were collected from medical charts at 65 oncology clinics in hospital outpatient and community settings for consecutive patients who received the first dose of either darbepoetin alfa or epoetin alfa between August 1, 2002, and February 15, 2003, and were to have had 12 weeks of follow-up data. RESULTS: Data from the charts of 3123 patients were abstracted. Of these patients, 2785 were treated with only one erythropoietic agent (1444 with darbepoetin alfa and 1341 with epoetin alfa) and were included in the analysis. The most common initial dosage of darbepoetin alfa was 200 microg every two weeks (61% of darbepoetin alfa recipients), and the most common initial dosage of epoetin alfa was 40,000 units weekly (72%). With these regimens, the dosage was escalated for 22% of darbepoetin alfa recipients and 23% of epoetin alfa recipients at a median of six weeks after the initial dose. The mean change from baseline in hemoglobin concentration after 12 weeks of therapy was similar for both groups, as was the percent of patients with red-blood-cell transfusions during treatment. CONCLUSION: The most common initial dosage of darbepoetin alfa for CIA was 200 microg every two weeks, and the most common initial dosage of epoetin alfa was 40,000 units weekly. At these dosages, the two agents appear to have similar clinical effectiveness. PMID- 15658075 TI - Effects of media coverage of Women's Health Initiative study on attitudes and behavior of women receiving hormone replacement therapy. AB - PURPOSE: The effects of media coverage of the Women's Health Initiative (WHI) study on the attitudes and behavior of women regarding hormone replacement therapy (HRT) were studied. METHODS: This cross-sectional observational study took place in 12 counties in western North Carolina from 1997 to 1999. Participants were recruited from among women who participated in a previous study, which examined the effects of osteoporosis education on adherence to prevention guidelines. Women in the current study were limited to those previously reporting postmenopausal status and use of HRT. Participants were interviewed by telephone with a 41-item scripted survey. Main outcomes included primary sources of information regarding HRT, perception of the accuracy of information from various sources, and changes in HRT use. RESULTS: Every woman in the study (n = 97) reported having heard about the WHI study, and 52% reported that it had affected their use of HRT. Women using estrogen alone were as likely as women using an estrogen-plus-progestin combination to have had their use affected (48% versus 59%, respectively). Women making a change after hearing about the WHI study results were significantly less likely to trust information from their physicians regarding HRT. CONCLUSION: A survey of women who had received HRT found that media reports on the WHI study had a significant influence on their use of HRT. PMID- 15658077 TI - Perceived accuracy of drug orders transmitted orally by telephone. PMID- 15658076 TI - Controlling antimicrobial use and decreasing microbiological laboratory tests for urinary tract infections in spinal-cord-injury patients with chronic indwelling catheters. AB - PURPOSE: The effect of replacing the indwelling catheter of patients suspected of having a urinary tract infection (UTI) before collecting a urine sample on the number of organisms isolated in cultures and on drug and microbiology laboratory costs was studied. METHODS: Data were collected for all patients hospitalized in two spinal cord injury (SCI) units between October 2001 and March 2002 who had an indwelling catheter or suprapubic catheter and were suspected of having a UTI. Urine samples were obtained through a port of the indwelling catheter in one SCI unit, while the indwelling catheter was replaced immediately before each urine sample was obtained in the second SCI unit. Patient demographics, history of antimicrobial use, bacterial isolate sensitivity data, and current antimicrobial treatment were recorded. RESULTS: A total of 85 patients, 41 in the control group and 44 in the intervention group, were enrolled during the six-month study period. In the control and intervention groups, 93 and 79 organisms were isolated, respectively, with an average of 2 isolates per patient in the control group and 1 per patient in the intervention group. Patients in the control group had significantly more multidrug-resistant organisms in their urine, with 34 isolated from 26 patients (63%) (p < 0.001). Changing the indwelling catheter decreased antimicrobial and microbiology laboratory costs, resulting in a cost saving of $15.64 per patient. CONCLUSION: Replacement of the indwelling catheter before collecting a urine sample for culture and conducting susceptibility testing reduced the pathogens identified, the number of toxic antimicrobials prescribed to treat the infection, and the costs of antimicrobials and microbiology laboratory technician time. PMID- 15658074 TI - Meta-analysis of trials comparing postoperative recovery after anesthesia with sevoflurane or desflurane. AB - PURPOSE: Results of published, randomized controlled trials comparing sevoflurane and desflurane were pooled to measure differences in times until patients obeyed commands, were extubated, were oriented, were discharged from the postanesthesia care unit (PACU), and were ready to be discharged to home, as well as the occurrence of postoperative nausea and vomiting (PONV). METHODS: We reviewed all randomized clinical trials in MEDLINE through December 18, 2003, with a title or abstract containing the words sevoflurane and desflurane. Two reviewers independently extracted study data from papers that met inclusion criteria. Endpoints were pooled using random-effects meta-analysis. RESULTS: Twenty-two reports of 25 studies (3 reports each described 2 studies) met our inclusion criteria. A total of 746 patients received sevoflurane, and 752 received desflurane. Patients receiving desflurane recovered 1-2 minutes quicker in the operating room than patients receiving sevoflurane. They obeyed commands 1.7 minutes sooner (p < 0.001; 95% confidence interval [CI], 0.7-2.7 minutes), were extubated 1.3 minutes sooner (p = 0.003; 95% CI, 0.4-2.2 minutes), and were oriented 1.8 minutes sooner (p < 0.001; 95% CI, 0.7-2.9 minutes). No significant differences were detected in the phase I or II PACU recovery times or in the rate of PONV. CONCLUSION: Meta-analysis of studies in which the duration of anesthesia was up to 3.1 hours indicated that patients receiving either desflurane or sevoflurane did not have significant differences in PACU time or PONV frequency. Patients receiving desflurane followed commands, were extubated, and were oriented 1.0-1.2 minutes earlier than patients receiving sevoflurane. PMID- 15658078 TI - Comparison of sargramostim and filgrastim in the treatment of chemotherapy induced neutropenia. PMID- 15658080 TI - Dog days. PMID- 15658079 TI - Stability and compatibility of levofloxacin and metronidazole during simulated and actual Y-site administration. PMID- 15658081 TI - Mental capacity: in search of alternative perspectives. PMID- 15658083 TI - Capacity and competence in child and adolescent psychiatry. AB - Capacity and competence in the field of child and adolescent psychiatry are complex issues, because of the many different influences that are involved in how children and adolescents make treatment decisions within the setting of mental health. This article will examine some of the influences which must be considered, namely: developmental aspects, the paradoxical relationship between the need for autonomy and participation and the capacity of children, family psychiatry, and the duty of care towards children and adolescents. The legal frameworks relevant to consideration of consent and competence will be briefly considered, as well as some studies of children's consent, participation and competence. A case vignette will be used as a focus to consider the complexity of the issue of competence in child and adolescent psychiatry, in the particular mental disorder of anorexia nervosa. PMID- 15658082 TI - The cognitive based approach of capacity assessment in psychiatry: a philosophical critique of the MacCAT-T. AB - This article gives a brief introduction to the MacArthur Competence Assessment Tool-Treatment (MacCAT-T) and critically examines its theoretical presuppositions. On the basis of empirical, methodological and ethical critique it is emphasised that the cognitive bias that underlies the MacCAT-T assessment needs to be modified. On the one hand it has to be admitted that the operationalisation of competence in terms of value-free categories, e.g. rational decision abilities, guarantees objectivity to a great extent; but on the other hand it bears severe problems. Firstly, the cognitive focus is in itself a normative convention in the process of anthropological value-attribution. Secondly, it misses the complexity of the decision process in real life. It is therefore suggested that values, emotions and other biographic and context specific aspects should be considered when interpreting the cognitive standards according to the MacArthur model. To fill the gap between cognitive and non cognitive approaches the phenomenological theory of personal constructs is briefly introduced. In conclusion some main demands for further research to develop a multi-step model of competence assessment are outlined. PMID- 15658085 TI - The contribution of narrative ethics to issues of capacity in psychiatry. AB - Cognitive and rational assessments of competence do not fully capture the way in which individuals normally make decisions. Human beings have always used stories to explain their experiences and values. Narrative ethics should be used to understand the perspective in context of a patient whose competence is in question, and so avoid a destructive clash. Psychiatry and professionals within it also have a narrative that may join with that of science, but there is no special privilege for these narratives unless survival is at stake. The narrative approach should be used to try to make different stories compatible. This article examines the background to this approach, and indicates some ways in which it could be used in the specific cases addressed in the series. PMID- 15658084 TI - Competence in mental health care: a hermeneutic perspective. AB - In this paper we develop a hermeneutic approach to the concept of competence. Patient competence, according to a hermeneutic approach, is not primarily a matter of being able to reason, but of being able to interpret the world and respond to it. Capacity should then not be seen as theoretical, but as practical. From the perspective of practical rationality, competence and capacity are two sides of the same coin. If a person has the capacity to understand the world and give meaning to the situation, he or she is able to make decisions, and is thus competent. People can fail in the area of practical rationality. They can feel ill at ease, uncomfortable or not at home in the situation. Under such conditions, they appear as incompetent, and urge caregivers to respond in such a way that their competence can be raised. The issue is not how to measure their incompetence, but how to help them to become more competent, that is to get a practical grip on their situation and to be able to live out their lives in such a way that they develop their identity in relations with others. From a hermeneutic point of view, assessing a patient's capacity implies focusing on the patient's way of meaning making and regarding her behavior from the perspective of practical rationality. The focus should not be on the assessment as a matter of fact, but on improving capacity. This requires allowing the patient to experience the world meaningfully and affording her, in the context of a supportive and trustful institutional environment, the possibility of developing a personal narrative where her choices are expressed verbally or non-verbally. PMID- 15658086 TI - Feminist ethic of care: a third alternative approach. AB - A man with Alzheimer's who wanders around, a caregiver who disconnects the alarm, a daughter acting on het own, and a doctor who is not consulted set the stage for a feminist reflection on capacity/competence assessment. Feminist theory attempts to account for gender inequality in the political and in the epistemological realm. One of its tasks is to unravel the settings in which actual practices, i.c. capacity/competence assessment take place and offer an alternative. In this article the focus will be on a feminist ethics of care in which relationality, care, vulnerability, and responsibility are privileged concepts and attitudes. The emphasis on these notions leads to a specific view of autonomy that has consequences for both carereceivers (patients, clients) and caregivers (professional and not professional). These concepts constitute a default setting that shapes the context for capacity/competence assessment. Whereas this notion is meant to distinguish between those who need to be taken care of and those who do not, reflection on what it means to say 'those who need to be taken care of is also required. The feminist analysis presented here emphasizes the necessity of the contextualization of assessment of competence. It sketches the multifold and complex grid that comprehends capacity assessment. PMID- 15658087 TI - Patient capacity in mental health care: legal overview. AB - The discriminatory effects of categorizing psychiatric patients into competent and incompetent, have urged lawyers, philosophers and health care professionals to seek a functional approach to capacity assessment. Dutch and English law have produced some guidelines concerning this issue. So far, most legal systems under investigation have concentrated on alternatives for informed consent by the patient in case of mental incapacity, notably substitute decision-making, intervention of a judge and advance directives. It is hard to judge the way in which the law may further adapt to a more functional assessment of capacity, because the nature of law shows that legal reforms usually take place only when new methods have been accepted by the field. This is not yet the case today. PMID- 15658088 TI - Preservation of hearing in patients undergoing microsurgery for vestibular schwannoma: degree of meatal filling. AB - OBJECT: Preservation of hearing has become a standard goal in selected patients undergoing surgery for a vestibular schwannoma (VS). This study was aimed at analyzing the role played by filling of the internal auditory canal (IAC) as well as those played by preoperative hearing quality, and tumor size in the postoperative preservation of serviceable hearing (SH). METHODS: Three hundred eighty-six patients with VS were treated. Hearing preservation was attempted in 128 cases (33.2%) by using intraoperative monitoring and following a retrosigmoid approach. The maximal extrameatal size of the tumor, its extension within the IAC, and pre- and postoperative hearing quality, according to the Gardner Robertson classification, were evaluated. Preservation of SH was achieved in 24.2% of the 128 patients. With respect to tumor size, SH was preserved in 39% of 77 patients harboring a tumor 15 mm wide or smaller and in 2% of 51 patients with lesions 16 mm wide or larger (p < 0.001). With regard to filling of the IAC, among 63 patients harboring a tumor 15 mm or smaller, in whom magnetic resonance images were available, SH was preserved in 52.8% of 36 patients with partial filling and in 25.9% of 27 patients with complete filling (p = 0.032). Concerning preoperative hearing quality, in the patients with tumors 15 mm or smaller, SH was preserved in 46.5% of 43 patients with Gardner-Robertson Class I hearing and 29.4% of 34 patients with Class II hearing (p = 0.126). Both tumor size and the extent of IAC filling proved statistically significant in a multivariable analysis (p < 0.001 and p = 0.026, respectively). CONCLUSIONS: Incomplete filling of the IAC and a tumor size of 15 mm or smaller are independent favorable factors in SH preservation. Excellent preoperative hearing appears to have a positive impact but does not have statistical significance. Intraoperative monitoring is useful in guiding the dissection; however, the surgeon's knowledge of topographical landmarks and meticulous surgical technique remain the essential factors of success. PMID- 15658089 TI - Long-term hearing preservation after surgery for vestibular schwannoma. AB - OBJECT: Vestibular schwannomas (VSs) are now amenable to resection with excellent hearing preservation rates. It remains unclear whether immediately postoperative hearing is a durable result and will not diminish over time. The aim of this study was to determine the rate of long-term preservation of functional hearing following surgery for a VS and to examine factors influencing hearing preservation. METHODS: All patients eligible for hearing preservation (Gardner Robertson Class I or II) who had undergone resection of a VS by a single surgeon were reviewed retrospectively. Follow-up audiograms and magnetic resonance images were obtained. Of 142 patients deemed eligible for hearing preservation surgery, 38 had immediate postoperative hearing confirmed by an audiogram. In these patients with preserved hearing, the audiographic results demonstrated functional hearing in 30 (85.7%) of 35 patients who underwent repeated testing at a mean follow-up time of 7 years. Delayed hearing loss occurred in five (14.3%) of the 35 patients and did not correlate significantly with the size of the tumor. Hearing improved one Gardner-Robertson class postoperatively in three (7.9%) of the 38 patients. CONCLUSIONS: Long-term functional hearing was maintained in 85.7% of patients when it was preserved immediately postoperatively and the result was independent of tumor size. The results of this study emphasize that long-term preservation of functional hearing is a realistic goal following VS surgery and should be attempted in all patients in whom preoperative hearing is determined to be Gardner-Robertson Class I or II. PMID- 15658090 TI - Long-term outcomes in patients with vestibular schwannomas treated using gamma knife surgery: 10-year follow up. AB - OBJECT: Gamma knife surgery (GKS) has been a safe and effective treatment for vestibular schwannomas in both the short and long term, although less is known about long-term outcomes in the past 10 years. The aim of this study was to clarify long-term outcomes in patients with vestibular schwannomas treated using GKS based on techniques in place in the early 1990s. METHODS: Eighty patients harboring a vestibular schwannoma (excluding neurofibromatosis Type 2) were treated using GKS between May 1991 and December 1993. Among these, 73 patients were assessed; seven were lost to follow up. The median duration of follow up was 135 months. The mean patient age at the time of GKS was 56 years old. The mean tumor volume was 6.3 cm3, and the mean maximal and marginal radiation doses applied to the tumor were 28.4 and 14.6 Gy, respectively. Follow-up magnetic resonance images were obtained in 71 patients. Forty-eight patients demonstrated partial tumor remission, 14 had tumors that remained stable, and nine demonstrated tumor enlargement or radiation-induced edema requiring resection. Patients with larger tumors did not fare as well as those with smaller lesions. The actuarial 10-year progression-free survival rate was 87% overall, and 93% in patients with tumor volumes less than 10 cm3. No patient experienced malignant transformation. CONCLUSIONS: Gamma knife surgery remained an effective treatment for vestibular schwannomas for longer than 10 years. Although treatment failures usually occurred within 3 years after GKS, it is necessary to continue follow up in patients to reveal delayed tumor recurrence. PMID- 15658092 TI - Clinical implications of associated venous drainage in patients with cavernous malformation. AB - OBJECT: The authors reviewed angiograms obtained in patients with cavernous malformations to identify and characterize coexisting venous drainage. METHODS: Fifty-seven patients with cavernous malformations treated at the authors' institutions between 1994 and 2002 were classified into three groups according to the venous system adjacent to the malformation on angiography studies. In Group A patients (23 patients) the malformations had no venous drainage; in Group B patients (14 patients) the lesions were associated with typical venous malformations; and in Group C patients (20 patients) the lesions had atypical venous drainage (AVD). The risk of hemorrhage based on the type of associated venous drainage was analyzed, and the usefulness of magnetic resonance (MR) imaging compared with digital subtraction (DS) angiography in demonstrating associated AVD was determined. Fifty-seven patients harbored 67 cavernous malformations: Group A patients had 29 cavernous malformations with no associated venous drainage; Group B patients had 17 lesions associated with venous malformations; and Group C patients harbored 21 lesions, 20 of which manifested AVD. Symptomatic hemorrhage was present in 10 (43.5%) of 23 Group A patients and in 28 (82.4%) of 34 Groups B and C patients. Although high-resolution MR imaging revealed the presence of associated venous malformations in 11 (78.6%) of 14 Group B patients, such studies demonstrated AVD in only two (10%) of 20 Group C patients. CONCLUSIONS: Patients harboring cavernous malformations plus venous malformations or AVD are more likely to present with symptomatic hemorrhage than are patients with cavernous malformation alone. The actual incidence of associated venous drainage may be underestimated when MR imaging alone is used rather than combined with DS angiography. PMID- 15658091 TI - Meningiomas of the cerebellopontine angle with extension into the internal auditory canal. AB - OBJECT: Only some meningiomas of the cerebellopontine angle (CPA) extend into the internal auditory canal (IAC) or arise from its dural lining. The authors investigated cases of CPA tumors in which the meningioma was inserted in the dura mater in or at the ICA or infiltrated a cranial nerve. METHODS: The authors reviewed patient charts including surgical and clinical records, intraoperative recordings of auditory evoked potentials, records of postoperative auditory examinations, and imaging studies. In a series of 421 patients harboring CPA meningiomas, 72 patients in whom there was dural involvement of the IAC were investigated. Total tumor resection was achieved in 86.1%. In 34 patients, opening of the IAC was required for total tumor removal; this procedure did not influence the patient functional outcome. Among patients with secondary involvement of the IAC, anatomical preservation of the facial and cochlear nerves was obtained in 94%, whereas among patients in whom the lesion arose from the dura in or at the IAC these values were 80 and 75%, respectively. Functional preservation of the seventh and eighth cranial nerves in cases of tumor extension within the IAC was 86 and 77%, respectively, whereas in cases in which the IAC was involved it was only 60%. In four of five patients in whom the tumor had its origin in the dura mater within the IAC, the seventh or eighth cranial nerve had to be sacrificed to achieve tumor removal because of the lesion's infiltrative behavior. Facial nerve reconstruction by sural grafting was performed in the same operative procedure. CONCLUSIONS: Meningiomas of the CPA involving the IAC require special surgical management. Dural involvement of the IAC requires opening by using a diamond drill, a procedure that does not influence cranial nerve outcome. The increased rate of cranial nerve morbidity is attributed to the infiltrative behavior of these meningiomas. If affected nerve segments have to be sacrificed, immediate reconstruction enables satisfactory long-term results. PMID- 15658093 TI - Frequency and management of recurrent stenosis after carotid artery stent implantation. AB - OBJECT: To determine the rate of hemodynamically significant recurrent carotid artery (CA) stenosis after stent-assisted angioplasty for CA occlusive disease, the authors analyzed Doppler ultrasonography data that had been prospectively collected between October 1998 and September 2002 for CA stent trials. METHODS: Patients included in the study participated in at least 6 months of follow-up review with serial Doppler studies or were found to have elevated in-stent velocities (> 300 cm/second) on postprocedure Doppler ultrasonograms. Hemodynamically significant (> or = 80%) recurrent stenosis was identified using the following Doppler criteria: peak in-stent systolic velocity at least 330 cm/second, peak in-stent diastolic velocity at least 130 cm/second, and peak internal carotid artery/common carotid artery velocity ratio at least 3.8. Follow up studies were obtained at approximate fixed intervals of 1 day, 1 month, 6 months, and yearly. Angiography was performed in the event of recurrent symptoms, evidence of hemodynamically significant stenosis on Doppler ultrasonography, or both. Treatment was repeated because of symptoms, angiographic evidence of severe (> or = 80%) recurrent stenosis, or both of these. Stents were implanted in 142 vessels in 138 patients (all but five patients were considered high-risk surgical candidates and 25 patients were lost to follow-up review). For the remaining 112 patients (117 vessels), the mean duration of Doppler ultrasonography follow up was 16.42+/-10.58 months (range 4-54 months). Using one or more Doppler criteria, severe (> or = 80%) in-stent stenosis was detected in six patients (5%). Eight patients underwent repeated angiography. Six patients (three with symptoms) required repeated intervention (in four patients angioplasty alone; in one patient conventional angioplasty plus Cutting Balloon angioplasty; and in one patient stent-assisted angioplasty). CONCLUSIONS: In a subset of primarily high risk surgical candidates treated with stent-assisted angioplasty, the rates of hemodynamically significant restenosis were comparable to surgical restenosis rates cited in previously published works. Treatment for recurrent stenosis incurred no instance of periprocedure neurological morbidity. PMID- 15658094 TI - Postoperative cerebral hyperperfusion associated with impaired cognitive function in patients undergoing carotid endarterectomy. AB - OBJECT: Cognitive impairment occurs in 20 to 30% of patients following carotid endarterectomy (CEA). The purpose of the present study was to determine whether postoperative cerebral hyperperfusion is associated with impairment of cognitive function in patients undergoing that procedure. METHODS: Cerebral blood flow (CBF) was measured using single-photon emission computerized tomography scanning before and immediately after CEA and on the 3rd postoperative day in 92 patients with ipsilateral internal carotid artery stenosis of 70% or greater. Hyperperfusion post-CEA was defined as a 100% increase or greater in CBF compared with preoperative values. Neuropsychological testing was also performed preoperatively and at the 1-, 3-, and 6-month follow-up examinations. At the 1 month postoperative neuropsychological assessment, 11 patients (12%) displayed evidence of cognitive impairment. In addition, the incidence of postoperative cognitive impairment in patients with post-CEA hype perfusion (seven [58%] of 12 patients) was significantly higher than that in patients without post-CEA hyperperfusion (four [5%] of 80 patients; p < 0.0001). A logistic regression analysis demonstrated that post-CEA hyperperfusion was the only significant independent predictor of postoperative cognitive impairment. Of the seven patients in whom post-CEA hyperperfusion and cognitive impairment were identified 1 month postoperatively, four (including three patients with hyperperfusion syndrome) remained cognitively impaired at the 3- and 6-month follow-up examinations. CONCLUSIONS: Postoperative cerebral hyperperfusion is associated with impairment of cognitive function in patients undergoing CEA. Furthermore, the development of hyperperfusion syndrome is associated with the persistence of postoperative cognitive impairment. PMID- 15658095 TI - Proximal intracranial internal carotid artery branches: prevalence and importance for balloon occlusion test. AB - OBJECT: The aim of this study was to determine the prevalence of angiographically identifiable skull base arterial branches that potentially serve as collateral conduits during a balloon occlusion test (BOT) of the internal carotid artery (ICA). The authors posited that neurological deficits in patients who had previously tolerated the occlusion test may be attributable to an unrecognized collateral support through these channels (operant during proximal ICA BOT) when permanent ICA occlusion was performed more distally. METHODS: In 481 cases (962 ICAs), cerebral angiograms obtained during routine Wada testing were retrospectively reviewed. Two hundred sixty-one patients had at least one angiographically identifiable ICA branch; 109 patients had two or more branches. A meningohypophyseal branch of the cavernous ICA was identified on the right side in 108 patients and on the left in 122. A vidian artery originated from the petrous portion of the ICA on the right side in 58 patients and on the left in 85. The inferolateral trunk revealed itself as a branch of the cavernous ICA on the right side in 17 patients and on the left in 33. A caroticotympanic artery arose from a left cavernous ICA. A persistent trigeminal artery was situated on the right side in two patients and on the left in three. More than half of the patients had angiographically identifiable and perhaps hemodynamically significant skull base branches of the ICA, and approximately one quarter had more than one identifiable branch. CONCLUSIONS: The authors recommend that patients be screened during angiography studies performed prior to BOT in branches of the proximal intracranial ICA and that the site of BOT be moved distally if such branches are identified. PMID- 15658096 TI - Long-term deep brain stimulation in elderly patients with cardiac pacemakers. AB - OBJECT: Deep brain stimulation (DBS) has become an accepted therapy for movement disorders such as Parkinson disease (PD) and essential tremor (ET), when these conditions are refractory to medical treatment. The presence of a cardiac pacemaker is still considered a contraindication for DBS in functional neurosurgery. The goal of this study was to evaluate the technical and clinical management of DBS for the treatment of movement disorders in elderly patients with cardiac pacemakers. METHODS: Six patients with cardiac pacemakers underwent clinical and cardiac examinations to analyze the safety of DBS in the treatment of movement disorders. Four patients suffered from advanced PD and two patients had ET. The mean age of these patients at surgery was 69.5 years (range 63-79 years). The settings of the pacemakers were programmed in a manner considered to minimize the chance of interference between the two systems. There were no adverse events during surgery. Four patients underwent stimulation of the thalamic ventralis intermedius nucleus (VIM), and two patients stimulation of the subthalamic nucleus. In general, bipolar sensing was chosen for the cardiac pacemakers. In all but one patient the quadripolar DBS electrodes were programmed for bipolar stimulation. Several control electrocardiography studies, including 24-hour monitoring, did not show any interference between the two systems. At the time this paper was written the patients had been followed up for a mean of 25.3 months (range 4-48 months). CONCLUSIONS: In certain conditions it is safe for patients with cardiac pacemakers to receive DBS for treatment of concomitant movement disorders. Cardiac pacemakers should not be viewed as a general contraindication for DBS in patients with movement disorders. PMID- 15658097 TI - Risk factors for aneurysmal subarachnoid hemorrhage in patients in Izumo City, Japan. AB - OBJECT: The annual incidence of aneurysmal subarachnoid hemorrhage (SAH) in Izumo City, Japan, appears to be the highest rate among those reported; therefore the author investigated the risk factors for SAH in patients in this city. METHODS: A case-control study of 247 patients (108 men and 139 women with ages ranging from 28-96 years) with aneurysmal SAH was conducted in Izumo between 1980 and 1998. Hypertension, diabetes mellitus, heart disease, liver disease, cigarette smoking, alcohol consumption, and serum levels of total cholesterol, aspartate aminotransferase, alanine aminotransferase, and urea nitrogen were assessed as possible risk factors for SAH by using conditional logistic regression. After adjustment for other risk factors, results of multivariate analysis showed that hypertension was the most powerful risk factor, regardless of age and sex. The odds ratio for hypertension was higher in women than in men. The second greatest risk factors were cigarette smoking in those 59 years of age or younger and in men and hypercholesterolemia in those 60 years of age or older and in women. Among individuals 60 years of age or older and among women, diabetes mellitus and heart disease were inversely associated with the risk of SAH. When analyses were performed in 219 cases of confirmed ruptured cerebral aneurysm, very similar results were obtained. CONCLUSIONS: Among patients in Izumo, hypertension was the most notable risk factor for aneurysmal SAH, regardlessof age and sex, followed by cigarette smoking in younger men and hypercholesterolemia in older women. In older women, diabetes mellitus and heart disease decreased the risk of SAH. PMID- 15658098 TI - Influence of endothelial nitric oxide synthase T-786C single nucleotide polymorphism on aneurysm size. AB - OBJECT: Among patients with aneurysms, those with heterozygous (T/C) endothelial nitric oxide synthase (eNOS) T-786C single nucleotide polymorphism (SNP), a mutation reducing endothelial nitric oxide synthesis, are reported to have larger ruptured intracranial aneurysms (IAs) than those with homozygous (C/C or T/T) genotype. The authors tested patients harboring aneurysms for eNOS T-786C SNP in two populations--Japanese and Korean. METHODS: The eNOS T-786C SNP was genotyped through direct sequencing in genomic DNA obtained from 336 Japanese and 191 Korean patients with lAs and 214 Japanese and 191 Korean control volunteers. Differences in genotype frequencies among the various aneurysm sizes were evaluated using the Fisher exact test. There was no significant difference in heterozygous (T/C) eNOS T-786C SNP between aneurysms 5 mm or smaller and those from 6 to 9 mm, and between lesions 5 mm or smaller and those 10 mm or larger in 336 Japanese patients harboring aneurysms--220 with ruptured and 116 with unruptured lesions--and in 191 Korean patients with ruptured aneurysms. CONCLUSION: The eNOS T-786C SNP genotype does not influence the size of aneurysms. PMID- 15658099 TI - Enlarging vertebrobasilar nonsaccular intracranial aneurysms: frequency, predictors, and clinical outcome of growth. AB - OBJECT: Vertebrobasilar nonsaccular intracranial aneurysms (NIAs) are characterized by elongation, dilation, and tortuosity of the vertebrobasilar arteries. The goal of this study was to define the frequency, predictors, and clinical outcome of the enlargement of vertebrobasilar NIAs. METHODS: Patients with vertebrobasilar fusiform or dolichoectatic aneurysms demonstrated on imaging studies between 1989 and 2001 were identified. In particular, patients who had undergone serial imaging were included in this study and their medical records were retrospectively reviewed. Prospective information was collected from medical records or death certificates when available. Both initial and serial imaging studies were reviewed. The authors defined NIA enlargement as a change in lesion diameter greater than 2 mm or noted on the neuroradiologist's report. A Cox proportional hazards regression was used to model time from diagnosis of the vertebrobasilar NIA to the first documented enlargement as a function of various predictors. The Kaplan-Meier method was used to study patient death as a function of aneurysm growth. Of the 159 patients with a diagnosis of vertebrobasilar NIA, 52 had undergone serial imaging studies including 25 patients with aneurysm enlargement. Lesion growth significantly correlated with symptomatic compression at the initial diagnosis (p = 0.0028), lesion type (p < 0.001), and the initial maximal lesion diameter (median 15 mm in patients whose aneurysm enlarged compared with median 8 mm in patients whose aneurysm did not enlarge; p < 0.001). The mortality rate was 5.7 times higher in patients with aneurysm growth than in those with no enlargement after adjustment for patient age (p = 0.002). CONCLUSIONS: Forty-eight percent of vertebrobasilar NIAs demonstrated on serial imaging enlarged, and this growth was associated with significant morbidity and death. Significant risk factors for aneurysm enlargement included symptomatic compression at the initial diagnosis, transitional or fusiform vertebrobasilar NIAs, and initial lesion diameter. Further studies are necessary to determine appropriate treatments of this disease entity once enlargement has been predicted or occurs. PMID- 15658100 TI - Early transplantation of an encapsulated glial cell line-derived neurotrophic factor-producing cell demonstrating strong neuroprotective effects in a rat model of Parkinson disease. AB - OBJECT: Glial cell line-derived neurotrophic factor (GDNF) has been shown to confer neuroprotective effects on dopaminergic neurons. The authors investigated the effects of GDNF on 6-hydroxydopamine (6-OHDA)-treated dopaminergic neurons in vitro and in vivo. METHODS: First, the authors examined how 1, 10, or 100 ng/ml of GDNF, administered to cells 24 hours before, simultaneously with, or 2 or 4 hours after 6-OHDA was added, affected dopaminergic neurons. In a primary culture of E14 murine ventral mesencephalic neurons, earlier treatment with the higher dosage of GDNF suppressed 6-OHDA-induced loss of dopaminergic neurons better than later treatment. Next, the authors examined whether continuous infusion of GDNF at earlier time points would demonstrate a greater neuroprotective effect in a rat model of Parkinson disease (PD). They established a human GDNF-secreting cell line, called BHK-GDNF, and encapsulated the cells into hollow fibers. The encapsulated cells were unilaterally implanted into the striatum of adult rats 1 week before; simultaneously with; or 1, 2, or 4 weeks after 6-OHDA was given to induce lesions of the same striatum. With the earlier transplantation of a BHK GDNF capsule, there was a significant reduction in the number of amphetamine induced rotations displayed by the animals. Rats that had received earlier implantation of BHK-GDNF capsules displayed more tyrosine hydroxylase-positive neurons in the substantia nigra pars compacta and a tendency for glial proliferation in the striatum. CONCLUSIONS: These neuroprotective effects may be related to glial proliferation and signaling via the GDNF receptor alpha1. The results of this study support a role for this grafting technique in the treatment of PD. PMID- 15658101 TI - Real-time in vivo imaging of the convective distribution of a low-molecular weight tracer. AB - OBJECT: Convection-enhanced delivery (CED) is increasingly used to distribute therapeutic agents to locations in the central nervous system. The optimal application of convective distribution of various agents requires the development of imaging tracers to monitor CED in vivo in real time. The authors examined the safety and utility of an iodine-based low-molecular-weight surrogate tracer for computerized tomography (CT) scanning during CED. METHODS: Various volumes (total volume range 90-150 microl) of iopamidol (MW 777 D) were delivered to the cerebral white matter of four primates (Macaca mulatta) by using CED. The distribution of this imaging tracer was determined by in vivo real-time and postinfusion CT scanning (< or = 5 days after infusion [one animal]) as well as by quantitative autoradiography (14C-sucrose [all animals] and 14C-dextran [one animal]), and compared with a mathematical model. Clinical observation (- 5 months) and histopathological analyses were used to evaluate the safety and toxicity of the tracer delivery. Real-time CT scanning of the tracer during infusion revealed a clearly definable region of perfusion. The volume of distribution (Vd) increased linearly (r2 = 0.97) with an increasing volume of infusion (V.). The overall Vd/Vi ratio was 4.1+/-0.7 (mean+/-standard deviation) and the distribution of infusate was homogeneous. Quantitative autoradiography confirmed the accuracy of the imaged distribution for a small (sucrose, MW 359 D) and a large (dextran, MW 70 kD) molecule. The distribution of the infusate was identifiable up to 72 hours after infusion. There was no clinical or histopathological evidence of toxicity in any animal. CONCLUSIONS: Real-time in vivo CT scanning of CED of iopamidol appears to be safe, feasible, and suitable for monitoring convective delivery of drugs with certain features and low infusion volumes. PMID- 15658103 TI - Immobilization of basic fibroblast growth factor on a platinum microcoil to enhance tissue organization in intracranial aneurysms. AB - OBJECT: To enhance tissue organization in an aneurysm lumen, the authors prepared a platinum microcoil carrying basic fibroblast growth factor (bFGF) and analyzed its effectiveness in the treatment of aneurysms. METHODS: Ultrathin multiorganic layers were assembled on a platinum coil through successive deposition of cationic polyethylenimine and anionic heparin, and then bFGF was immobilized through an affinity interaction with heparin. The bFGF was effectively immobilized on the surface of the platinum coil without deterioration of the coil's mechanical properties. Coil embolization of aneurysms constructed using a canine common carotid artery was performed via the endovascular approach. The aneurysms together with parent arteries were harvested 2 weeks after coil embolization. Platinum coils unmodified, coated with heparin, or immobilized with heparin and bFGF were examined. The percentage of occlusion at the aneurysm orifice in animals treated with bFGF-immobilized coils (92.99+/-7.94%) was significantly greater than that in animals treated with heparin-coated coils (57.26+/-10.76%) or unmodified coils (52.86+/-8.54%). The histological score of the aneurysms treated with bFGF-immobilized coils was also significantly greater than the scores in the control group. CONCLUSIONS: These results indicated that bFGF-immobilized microcoils may be beneficial in the obliteration of aneurysms. PMID- 15658104 TI - Microsurgical anatomy of cerebral revascularization. Part I: anterior circulation. AB - OBJECT: Revascularization is an important component of treatment for complex aneurysms that require parent vessel occlusion, skull base tumors that involve major vessels, and certain ischemic diseases. In this study, the authors examined the microsurgical anatomy of cerebral revascularization in the anterior circulation by demonstrating various procedures for bypass surgery. METHODS: Twenty-five adult cadaveric specimens were studied, using 3 to 40 magnification, after the arteries and veins had been perfused with colored silicone. The microsurgical anatomy of cerebral revascularization in the anterior circulation was examined with the focus on the donor, recipient, and graft vessels. The techniques discussed in this paper include the superficial temporal artery (STA) middle cerebral artery (MCA), middle meningeal artery (MMA)-MCA, and side-to-side anastomoses; short arterial and venous interposition grafting; and external carotid artery/internal carotid artery (ICA)-M2 and ICA-ICA bypasses. Bypass procedures for cerebral revascularization are divided into two categories depending on their flow volume: low-flow and high-flow bypasses. A low-flow bypass, such as the STA-MCA anastomosis, is used to cover a relatively small area, whereas a high-flow bypass, such as the ICA-ICA anastomosis, is used for larger areas. Cerebral revascularization techniques are also divided into two types depending on the graft materials: pedicled arterial grafts, such as STA and occipital artery grafts, and free venous or arterial grafts, which are usually saphenous vein and radial artery grafts. Pedicled arterial grafts are mainly used for low-flow bypasses, whereas venous or arterial grafts are used for high-flow bypasses. CONCLUSIONS: It is important to understand the methods of bypass procedures and to consider indications in which cerebral revascularization is needed. PMID- 15658102 TI - Isolation of immortalized, INK4a/ARF-deficient cells from the subventricular zone after in utero N-ethyl-N-nitrosourea exposure. AB - OBJECT: Brain tumors, including gliomas, develop several months after rats are exposed in utero to N-ethyl-N-nitroso-urea (ENU). Although pathological changes cannot be detected until these animals are several weeks old, the process that eventually leads to glioma formation must begin soon after exposure given the rapid clearance of the carcinogen and the observation that transformation of brain cells isolated soon after exposure occasionally occurs. This model can therefore potentially provide useful insights about the early events that precede overt glioma formation. The authors hypothesized that future glioma cells arise from stem/progenitor cells residing in or near the subventricular zone (SVZ) of the brain. METHODS: Cells obtained from the SVZ or corpus striatum in ENU-exposed and control rats were cultured in an epidermal growth factor (EGF)-containing, chemically defined medium. Usually, rat SVZ cells cultured in this manner (neurospheres) are nestin-positive, undifferentiated, and EGF-dependent and undergo cell senescence. Consistent with these prior observations, control SVZ cells undergo senescence by the 12th to 15th doubling (20 of 20 cultures). In contrast, three of 15 cultures of cells derived from the SVZs of individual ENU treated rats continue to proliferate for more than 60 cell passages. Each of these nestin-expressing immortalized cell lines harbored a common homozygous deletion spanning the INK4a/ARF locus and was unable to differentiate into neural lineages after exposure to specific in vitro stimuli. Nevertheless, unlike the rat C6 glioma cell line, these immortalized cell lines demonstrate EGF dependence and low clonogenicity in soft agar and did not form tumors after intracranial transplantation. CONCLUSIONS: Data in this study indicated that immortalized cells may represent glioma precursors that reside in the area of the SVZ after ENU exposure that may serve as a reservoir for further genetic and epigenetic hits that could eventually result in a full glioma phenotype. PMID- 15658105 TI - Microsurgical anatomy of cerebral revascularization. Part II: posterior circulation. AB - OBJECT: Revascularization is an important component of treatment for complex aneurysms, skull base tumors, and vertebrobasilar ischemia in the posterior circulation. In this study, the authors examined the microsurgical anatomy related to cerebral revascularization in the posterior circulation and demonstrate various procedures for bypass surgery. METHODS: The microsurgical anatomy of cerebral and cerebellar vessels as they relate to revascularization procedure and techniques, including extracranial-to-intracranial bypass grafting, arterial interposition grafting, and side-to-side anastomosis, were examined by performing stepwise dissections in 22 adult cadaveric specimens. The arteries and veins in the specimens were perfused with colored silicone. Dominant cerebral and cerebellar revascularization procedures in the posterior circulations include superficial temporal artery (STA)-posterior cerebral artery (PCA), STA-superior cerebellar artery (SCA), occipital artery (OA)-anterior inferior cerebellar artery, OA-posterior inferior cerebellar artery (PICA), and PICA-PICA anastomoses. These procedures are effective in relatively small but critical areas including the brainstem and cerebellum. For revascularization of larger areas a saphenous vein graft is used to create a bypass between the PCA and the external carotid artery. Surgical procedures are generally difficult to perform in deep and narrow operative spaces near critical vital structures. CONCLUSIONS: Although a clear guideline for cerebral revascularization procedures has not yet been established, it is important to understand various microsurgical techniques and their related anatomical structures. This will help surgeons consider surgical indications for treatment of patients with vertebrobasilar ischemia caused by aneurysms, tumors, or atherosclerotic diseases in the posterior circulation. PMID- 15658106 TI - Incontinence after brain glioma surgery: new insights into the cortical control of micturition and continence. Case report. AB - Cortical control of micturition and continence remains poorly understood. The authors report two cases of patients who presented with prolonged urinary disturbances after resection of a brain glioma. Accurate anatomofunctional correlations determined using postoperative magnetic resonance imaging support the following: 1) the implication of the posterior portion of the right anterior cingulate gyms in the perception of bladder sensation and maintenance of continence; 2) the involvement of the right anterior insula in bladder relaxation; and 3) the role of the right inferior frontal cortex in the decision concerning whether to initiate a micturition. On the basis of these results, a preliminary model of a cortical network associated with micturition and continence is proposed. PMID- 15658107 TI - Cerebral vasospasm following intracranial hypotension caused by cerebrospinal fluid leak from an incidental lumbar durotomy. Case report. AB - The authors report on the unusual case of a patient with intracranial hypotension following an incidental durotomy complicated by an extensive but reversible cerebral vasospasm. Despite the dural tear repair and correction of the intracranial hypotension, the vasospasm ran its course. The precise mechanism of the cerebral vasospasm in this patient is unclear. PMID- 15658108 TI - Cerebellopontomandibular vascular malformation: a rare type of cerebrofacial arteriovenous metameric syndrome. Case report. AB - The concept of a cerebrofacial arteriovenous metameric syndrome (CAMS) recently has been posited. A 22-year-old woman presented with gait ataxia. She had unilateral angiomatous lesions involving the lower face, the auricle, and the retromastoid area, as well as the tongue on the right side. Angiography revealed the presence of extensive arteriovenous malformations (AVMs) in the ipsilateral cerebellum and pons, in addition to those located in the facial region. The authors report a case of multiple AVMs of the face, cerebellum, and pons, corresponding to CAMS Type 3, and include a review of the literature. PMID- 15658109 TI - Antegrade recanalization of a completely embolized vertebral artery after endovascular treatment of a ruptured intracranial dissecting aneurysm. Report of two cases. AB - Occlusion of the parent artery is a traditional method of treatment of unclippable cerebral aneurysms. Surgical or endovascular occlusion of the parent artery proximal to the aneurysm has been recommended for the treatment of dissecting aneurysms located in the vertebrobasilar circulation. Nevertheless, occlusion of the parent artery may not result in permanent exclusion of the aneurysm from the systemic circulation because, occasionally, postoperative rebleeding occurs after proximal occlusion. Alternatively, endovascular occlusion of the affected site, including the aneurysmal dilation, and parent artery, is a safe and reliable treatment for dissecting aneurysms. The authors present two rare cases of ruptured vertebral artery (VA) dissecting aneurysms that were treated by endovascular occlusion of the affected site including the aneurysm and parent artery by using Guglielmi detachable coils. In both cases the VA recanalized in an antegrade fashion during the follow-up period. Based on these unique cases, the authors suggest that a careful angiographic follow up of dissecting aneurysms is required, even in patients successfully treated with endovascular occlusion of the affected artery and aneurysm. PMID- 15658110 TI - Multiple meningiomas in a patient with Rubinstein-Taybi syndrome. Case report. AB - The authors report a case of multiple meningiomas in a 37-year-old woman with Rubinstein-Taybi syndrome. The patient harbored a bifrontal ossifying meningioma and multiple intracranial meningiomas. She underwent surgery for the frontal ossifying meningioma and a right frontoparietal meningioma. PMID- 15658111 TI - Discrepant findings for Wada test and functional magnetic resonance imaging with regard to language function: use of electrocortical stimulation mapping to confirm results. Case report. AB - The Wada test is still considered the gold standard for determining the language dominant hemisphere prior to brain surgery. The authors report on a 34-year-old right-handed woman whose Wada test results indicated that the right hemisphere was dominant for language. In contrast, functional magnetic resonance (fMR) imaging was indicative of bilaterally represented language functions. Activation in the left hemisphere demonstrated on fMR imaging was most pronounced in the Broca area. Importantly, fMR imaging results in this area were confirmed on electrocortical stimulation mapping. These contradictory findings indicated that a right hemispherre dominance for language according to the Wada test should be questioned and verified using electrocortical stimulation. Nonetheless, the question remains whether involvement of these areas in the left frontal hemisphere is critical for language, as these were spared during surgery. PMID- 15658112 TI - Cortical osteotomy technique for mobilizing the temporal muscle in pterional craniotomies. Technical note. AB - Although many procedures have already been performed to mobilize and reconstruct the temporal muscle for pterional craniotomies, the authors described a novel cortical osteotomy technique for creating and mobilizing a cortical bone slat along the superior temporal line with the temporal muscle attached to it. Screw fixation of the cortical bone slat then provides secured temporal muscle reconstruction. As such, this new technique minimized damage to the temporal muscle and prevented the formation of an anterior temporal hollow. In addition, key hole and parietal burr hole defects were covered by the cortical bone slat. PMID- 15658113 TI - Papaverine and vasospasm. PMID- 15658114 TI - Dissecting basilar aneurysms. PMID- 15658115 TI - Mild head injury. PMID- 15658116 TI - Mild head injury. PMID- 15658117 TI - Mild head injury. PMID- 15658118 TI - Phase 1 trial of oscillating field stimulation for complete spinal cord injury in humans. PMID- 15658119 TI - Oscillating field stimulation for complete spinal cord injury in humans: a phase 1 trial. AB - OBJECT: An electrical field cathode (negative pole) has trophic and tropic effects on injured spinal cord axons in in vitro and in vivo models of sea lamprey, rodent, and canine spinal cord injury (SCI) and it improves functional outcome. A human oscillating field stimulator (OFS) was built, a Food and Drug Administration (FDA) exemption number was obtained, and institutional review board approval was given for a Phase 1 trial to study 10 humans with complete motor and sensory SCI. METHODS: Entry criteria were complete SCI between C-5 and T-10 in patients 18 to 65 years of age and no transection demonstrated on magnetic resonance imaging. All participants received the National Acute Spinal Cord Injury Study (NASCIS) III methylprednisilone protocol. Cord compression and/or vertebral instability was treated before study entry. After treatment complete SCI (according to the American Spinal Injury Association [ASIA] score) remained in all patients with no somatosensory evoked potentials (SSEPs) below the injury level after surgery or for 48 hours. All patients underwent implantation of the OFS within 18 days. Patients underwent evaluation every 2 weeks postimplantation; the OFS was explanted at 15 weeks. Independent neurological status was assessed based on the ASIA score, visual analog scale (VAS) pain score, and SSEPs at 6 weeks, 6 months, and 1 year. Statistical analyses were performed using the two-tailed Wilcoxon test and analysis of variance (ANOVA). There were no complications at insertion of the OFS; there was one case of wound infection after explantation (5% infection rate). One patient was lost to follow up after 6 months. In all 10 patients the mean VAS pain score was 8 at implantation, 2 at 6 months, and in the nine attending follow up for 1 year it remained 2. At 1 year, the mean improvement in light touch was 25.5 points (ANOVA p < 0.001, Wilcoxon test p = 0.02), the mean improvement in pinprick sensation was 20.4 points (ANOVA p < 0.001, Wilcoxon test p = 0.02), and the mean improvement in motor status was 6.3 points (ANOVA p < 0.01, Wilcoxon test p = 0.02). Of five cases involving cervical cord injuries, bilateral upper extremity SSEPs were normal in one, unilateral upper-extremity SSEPs were recovered in four, bilateral upper-extremity SSEPs were recovered in one, and abnormal lower-extremity SSEPs resolved in one case. In one of the five cases involving thoracic injuries an abnormal lower-extremity SSEP resolved. CONCLUSIONS: The use of OFS treatment in patients with SCI is safe, reliable, and easy. Compared with the outcomes obtained in compliant NASCIS III plegic patients, the results of the present study indicate efficacy, and the FDA has given permission for enrollment of 10 additional patients. PMID- 15658120 TI - Experience with cervical stenosis and temporary paralysis in athletes. AB - OBJECT: Transient spinal cord injury (TSCI) in athletes presents one of the most challenging clinical scenarios. Management difficulties in and subsequent return to-play decisions are especially important in those with cervical canal stenosis. METHODS: Ten athletes (nine male and one female patients) were evaluated for TSCI. The diagnostic survey included physical and neurological examinations, plain radiographs with flexion-extension dynamic studies, computerized tomography, and magnetic resonance (MR) imaging. Clinical courses were followed and, in those who returned to contact sports activities, subsequent experience was noted. Symptoms consisted of paralysis, weakness, or numbness in all four extremities, their duration ranging from 15 minutes to 48 hours. Radiography revealed no evidence of fracture/dislocation or ligamentous instability. Spinal stenosis of 8 to 13 mm in length at three or more levels was evident in all cases. Four patients returned to competition without recurrent TSCI (mean follow up duration 40 months); six individuals retired. The occurrence of TSCI is not uncommon in athletes involved in contact sports. The diagnostic workup focuses on excluding fracture/dislocation, cord contusion, ligamentous infolding or instability, herniated nucleus pulposus, syrinx, or other surgically correctable lesions. There appear to be two groups of athletes who sustain TSCI: those who experience TSCI yet in whom radiographic studies are normal, and those with cervical stenosis, the most difficult management group. CONCLUSIONS: It does not appear that a single episode of TSCI in an athlete with spinal stenosis will substantially increase the risk of subsequent catastrophic spinal cord injury in those in whom MR imaging demonstrates preservation of cerebrospinal fluid signal. PMID- 15658122 TI - Impact of interlaminar graft materials on the fusion status in atlantoaxial transarticular screw fixation. AB - OBJECT: Structural interlaminar graft materials were used for atlantoaxial transarticular screw fixation (TSF), and its impact on the fusion status was investigated. METHODS: Forty-two patients (10 men, 32 women, mean age 51 years, mean follow-up period 45 months; 30 with rheumatoid arthritis, and 12 with os odontoideum) underwent TSF and modified Brooks posterior wiring involving titanium cables. As interlaminar graft materials, autologous bone from posterior iliac crest alone was used in 20 patients (Group A), and a structural spacer (13 ceramic spacers, nine titanium mesh cages) in 22 (Group B). Lateral radiographs were evaluated to determine bone fusion, alignment of the cervical spine, and wire loosening. Solid osseous fusion was obtained in 95% of Group A and 96% of Group B patients. The mean atlantoaxial angle was 19.1+/-9.7 degrees and 16.7+/ 10.4 degrees before surgery (p = 0.45), and 27.4+/-7.8 degrees and 22.1+/-5.5 degrees after surgery (p = 0.02) in Groups A and B, respectively. Atlantoaxial hyperlordosis (atlantoaxial angle > or = 30 degrees) was observed in 32% of Group A and 18% of Group B patients (p = 0.26). Postoperative kyphosis occurred in 40% of Group A and 23% of Group B patients (p = 0.28). Loosening of the cable was demonstrated in 50% of Group A and 36% of Group B patients(p = 0.37). In Group B patients maintenance of cervical lordosis was more likely than in those in Group A, although the differences did not reach statistical significance. CONCLUSIONS: These results indicate that structural interlaminar spacers can maintain proper cervical alignment without a decease in the fusion rate; the authors recommend their use in conjunction with TSF. PMID- 15658121 TI - Comparison of spinal and general anesthesia in lumbar laminectomy surgery: a case controlled analysis of 400 patients. AB - OBJECT: Despite a history of safety and efficacy, spinal anesthesia is rarely used in lumbar surgery. Application of regional anesthetics is widely preferred for lower-extremity surgery, but general anesthesia is used almost exclusively in spine surgery, despite evidence that spinal anesthesia is as safe and may offer some advantages. METHODS: In this case-controlled study the authors analyzed outcomes obtained in 400 patients in whom either spinal anesthesia or general anesthesia was induced to perform a lumbar decompression. Patients were matched for anesthesia-related class, preoperative diagnosis, surgical procedure, and perioperative protocols. All aspects of surgery, recovery, postanesthesia care, and pain management were uniform irrespective of the anesthetic type. Case complexity was equivalent. An independent observer performed analysis of the data. Data from the intraoperative period through hospital discharge were collected and compared. Two hundred consecutive patients meeting inclusion criteria were included in each group. Patients were treated for either lumbar stenosis or herniated nucleus pulposus. Demographically, both groups were well matched. Anesthetic and operative times were longer for patients receiving a general anesthetic (p < 0.05), in whom more nausea and greater requirements for antiemetics and pain medication were also present during recovery (p < 0.05). Overall complication rates and, specifically, the incidences of urinary retention were significantly lower in spinal anesthesia--induced patients (p < 0.05). There were no neural injuries in either group, and the incidence of spinal headache was lower in patients receiving a spinal anesthetic (1.5% compared with 3%). CONCLUSIONS: Spinal anesthesia was as safe and effective as general anethesia for patients undergoing lumbar laminectomy. Potential advantages of spinal anesthsia include a shorter anesthesia duration, decreased nausea, antiemetic and analgesic requirements, and fewer complications. Successful surgery can be performed using either anesthesia type. PMID- 15658123 TI - Calcium phosphate cement leakage after percutaneous vertebroplasty for osteoporotic vertebral fractures: risk factor analysis for cement leakage. AB - OBJECT: The purpose of this study was to analyze the risk factors for leakage of calcium phosphate cement (CPC) after vertebroplasty for osteoporotic vertebral fractures and to determine whether the vertebral body (VB) leakage caused any changes in the therapeutic benefits. METHODS: Between August 2000 and April 2002, the authors performed 65 CPC-assisted vertebroplasty procedures in 55 patients with thoracic or lumbar osteoporotic vertebral fractures. Back and low-back pain were evaluated using the visual analog scale and the duration of analgesic medication requirement. Factors related to CPC leakage and the postoperative outcome were analyzed. There was a small amount of VB CPC leakage in 23 cases. In 10 of 23 cases, leakage into the epidural space was found. Although VB CPC leakage was independently associated with high initial age, female sex, high bone mineral density (BMD), short injury-surgery interval, and injection via the unipedicular route in the logistic regression analysis, there was no factor associated with CPC leakage into the epidural space. Cement leakage into the epidural space reduced the immediate therapeutic effects on fracture-related pain (p = 0.0128). All patients in whom cement leaked into the epidural space had improved by the 2-week follow-up examination. CONCLUSIONS: Advanced initial age, female sex, high BMD, a short interval from injury to surgery, and injection via the unipedicular route may increase the incidence of CPC leakage. Cement leakage into the epidural space attenuated only the immediate therapeutic effects of CPC assisted vertebroplasty. PMID- 15658124 TI - Surgical experience with symptomatic thoracic ossification of the ligamentum flavum. AB - OBJECT: Symptomatic thoracic ossification of the ligamentum flavum (OLF) is rare, and its prognostic factors remain unclear. The authors retrospectively studied 24 patients with surgically treated thoracic OLF to delineate its prognostic factor. METHODS: The clinical manifestations, radiological studies, surgical records, and pathological findings were reviewed. Preoperative and postoperative neurological data were reappraised using the American Spinal Injury Association and modified Japanese Orthopaedic Association (JOA) scoring systems. Spearman rank-correlation coefficients and nonparametric tests were used to analyze the correlations between the variables of patient characteristics, preoperative duration of symptoms, preoperative neurological status, associated spinal disorder(s) other than thoracic OLF, and the final functional outcome. CONCLUSIONS: Decompressive surgery is indicated in patients in whom symptomatic thoracic spinal cord compression is caused by intruding OLF. Magnetic resonance imaging can provide sufficient clues for the diagnosis of thoracic OLF. Higher preoperative modified JOA scores of 3 and 4 are positively correlated with better postoperatiVe functional recovery than lower scores. Surgery should be performed as soon as possible before independent ambulatory function is impaired. PMID- 15658125 TI - Use of "MAPs" for determining the optimal surgical approach to metastatic disease of the thoracolumbar spine: anterior, posterior, or combined. Invited submission from the Joint Section Meeting on Disorders of the Spine and Peripheral Nerves, March 2004. AB - The surgical treatment of thoracolumbar metastases is controversial, and various approaches have been described. No single approach, however, is always applicable, and the optimal surgical strategy for any individual is determined by several interrelated factors. The authors have grouped these factors into four preoperative planning considerations that form the mnemonic "MAPS": 1) method of resection; 2) anatomy of spinal disease; 3) patient's level of fitness; and 4) stabilization. The choice of approach is also considered in light of the goals of surgery, including the relief of pain, neurological palliation, spinal stabilization, and oncological control. PMID- 15658126 TI - Postoperative fibrosis after surgical treatment of the porcine spinal cord: a comparison of dural substitutes. Invited submission from the Joint Section Meeting on Disorders of the Spine and Peripheral Nerves, March 2004. AB - OBJECT: Postoperative adhesion- and fibrosis-induced spinal cord tethering is not uncommon and may be associated with delayed clinical sequelae. Multiple dural substitutes have been used in surgery without a full appreciation of the grafts' adverse effects. The authors conducted a comparative animal experimental study to evaluate the degree of chronic inflammatory reactions, adhesions, and fibrosis caused by the use of four dural substitutes--Surgicel, Durasis, DuraGen, and Preclude. METHODS: Twenty-six pigs weighing 30 to 40 kg underwent a two-level lumbar laminectomy (a midline durotomy, implantation of a 2-cm dural substitute in the subarachnoid space, and watertight dural closure). After 8 weeks the animals were killed, and two independent neuropathologists blinded to the dural substitute group evaluated several sites along the implants, providing descriptions and quantitative scoring of fibrosis, chronic inflammatory reactions, foreign-body reactions, and spinal cord changes. Kruskal-Wallis one way analysis of variance for ranks corrected for multiple comparisons was used to examine differences among the materials. CONCLUSIONS: The DuraGen dural substitute produced the least amount of inflammation in the subarachnoid space and Preclude generated the most (p < 0.001). Surgicel and DuraGen were completely resorbed on histological sections, but both produced some inflammation, which diminished gradually from the dural implant center. Histological evaluation of the nonresorbed grafts demonstrated that Durasis caused the least degree of inflammatory cell infiltration (p < 0.001). The Preclude dural substitute consistently demonstrated encapsulation and arachnoidal reaction. There was no evidence of implant-related adverse effects on the underlying pia mater and white matter regardless of the substitute type. PMID- 15658128 TI - Biomechanical comparison: stability of lateral-approach anterior lumbar interbody fusion and lateral fixation compared with anterior-approach anterior lumbar interbody fusion and posterior fixation in the lower lumbar spine. AB - OBJECT: The stability of lateral lumbar interbody graft-augmented fusion and supplementary lateral plate fixation in human cadavers has not been determined. The purpose of this study was to investigate the immediate biomechanical stabilities of the following: 1) femoral ring allograft (FRA)-augmented anterior lumbar interbody fusion (ALIF) after left lateral discectomy combined with additional lateral MACS HMA plate and screw fixation; and 2) ALIF combined with posterior transpedicular fixation after anterior discectomy. METHODS: Sixteen human lumbosacral spines were loaded with six modes of motion. The intervertebral motion was measured using a video-based motion-capturing system. The range of motion (ROM) and the neutral zone (NZ) in each loading mode were compared with a maximum of 7.5 Nm. The ROM values for both stand-alone ALIF approaches were similar to those of the intact spine, whereas NZ measurements were higher in most loading modes. No significant intergroup differences were found. The ROM and NZ values for lateral fixation in all modes were significantly lower than those of intact spine, except when NZ was measured in lateral bending. All ROM and NZ values for transpedicular fixation were significantly lower than those for stand alone anterior ALIF. Transpedicular fixation conferred better stabilization than lateral fixation in flexion, extension, and lateral bending modes. CONCLUSIONS: Neither approach to stand-alone FRA-augmented ALIF provided sufficient stabilization, but supplementary instrumentation conferred significant stabilization. The MACS HMA plate and screw fixation system, although inferior to posterior transpedicular fixation, provided adequate stability compared with the intact spine and can serve as a sound alternative to supplementary spinal stabilization. PMID- 15658127 TI - Role of granulocyte-macrophage colony-stimulating factor in preventing apoptosis and improving functional outcome in experimental spinal cord contusion injury. AB - OBJECT: Granulocyte-macrophage colony-stimulating factor (GM-CSF) is a potent hemopoietic cytokine that stimulates stem cell proliferation in the bone marrow and inhibits apoptotic cell death in leukocytes. Its effects in the central nervous system, however, are still unclear. The present study was undertaken to determine if GM-CSF can rescue neuronal cells from apoptosis and improve neurological function in a spinal cord injury (SCI) model. METHODS: To study the effect of GM-CSF on apoptotic neuronal death, the authors used a staurosporine induced neuronal death model in an N2A cell line (in vitro) and in a rat SCI model (in vivo). The N2A cells were preincubated with GM-CSF for 60 minutes before being exposed to staurosporine for 24 hours. To inhibit GM-CSF, N2A cells were pretreated with antibodies against the GM-CSF receptor for 60 minutes. Clip compression was,used to induce SCI. Animals were treated with daily doses of GM CSF (20 microg/day) for 5 days. The number of apoptotic cells in the spinal cord and neurological improvements were assessed. Pretreatment with GM-CSF was found to protect N2A cells significantly from apoptosis, and neutralizing antibodies for the GM-CSF receptors inhibited the rescuing effect of GM-CSF on apoptosis. In the rat SCI model, neurological function improved significantly in the GM-CSF treated group compared with controls treated with phosphate-buffered saline. Terminal deoxynucleotidyl transferase-mediated deoxyuridine triphosphate nick-end labeling staining showed that GM-CSF administration reduced apoptosis in the injured spinal cord. CONCLUSIONS: Treatment of SCI with GM-CSF showed beneficial effects. Neuronal protection against apoptosis is viewed as a likely mechanism underlying the therapeutic effect of GM-CSF in SCI. PMID- 15658129 TI - Lipoma with dumb-bell extradural extension through the intervertebral foramen into the spinal canal. Case report. AB - The authors present the case of a 60-year-old woman with a neck lipoma that developed dumb-bell extradural extension, causing radiculopathy. To the best of the authors' knowledge, this is the first report of a lipoma originating in the neck with dumb-bell extradural extension through the intervertebral foramen and into the spinal canal. The lipoma was first excised from the foramen via a posterior approach to allow decompression of the nerve roots. The remaining lipomatous tissue was then resected via an anterior approach to avoid the region around the vertebral artery. PMID- 15658130 TI - Diagnosis-related pitfall of a lateral sacral cyst. Case report. AB - Because physical examination typically demonstrates normal findings in cases of low-back pain, diaglosis of the cause can be challenging. Frequent magnetic resonance imaging studies of the lumbosacral spine can typically lead to discovery of benign diseases and thus misinterpretation of these images. The authors report an unusual case in which a functional ovarian cyst was incidentally associated with a perineural cyst and mimicked a lateral sacral meningocele. In light of this, the authors recommend repeated examinations to avoid mistakes. PMID- 15658131 TI - Foraminal deposition of calcium pyrophosphate dihydrate crystals in the thoracic spine: possible relationship with disc herniation and implications for surgical planning. Report of two cases. AB - The authors report two cases of nodular calcium pyrophosphate dihydrate (CPPD) crystal deposition close to the thoracic neural foramen, which caused chronic radiculopathy. Preoperatively, the lesions were interpreted as calcified disc herniations. Both patients underwent surgery in which an extended transfacet pedicle-sparing approach was used. Incision of the posterior longitudinal ligament released soft degenerated material. In both cases, histological examination showed abundant degenerative debris along with CPPD crystals. Spinal CPPD deposition is a comparatively rare disease that almost invariably involves the posterior aspect of the spinal canal, typically the ligamentum flavum. The exceptional foraminal location of the lesions reported here, combined with the surgical findings, indicated that the CPPD crystals were deposited on a laterally herniated disc fragment. A distinctive feature in both cases was the soft consistency of the resected tissue. The consistency of the disc material and the location of the lesion in the axial plane (that is, median compared with lateral) are key factors in determining the optimal surgical approach to thoracic disc herniations. In describing consistency, terms such as "calcified" and "hard" have been used interchangeably in the literature. In the cases reported here, what appeared on computerized tomography and magnetic resonance imaging studies to be densely calcified lesions were shown intraoperatively to be soft herniations. The authors' experience underscores that not all densely calcified herniated discs are hard. Although detection of this discrepancy would have left surgical planning for the lateral disc herniations unchanged, it could have altered planning for centrally or centrolaterally located disc herniations. PMID- 15658132 TI - Rare cause of subarachnoid hemorrhage: spinal meningeal carcinomatosis. Case report. AB - Subarachnoid hemorrhage (SAH) of spinal origin is a rare entity accounting for approximately 1% of all cases of SAH. Its most frequent causes are trauma and vascular malformations. Although primary spinal tumors, especially ependymomas, are also relatively common causes, SAH secondary to a metastatic spinal tumor arising from outside the central nervous system is an extremely rare condition; only one case has been reported in the literature. The authors present a case of spinal meningeal carcinomatosis secondary to cutaneous malignant melanoma in which the patient presented with only symptoms of SAH. Although very rare, this case underscores several factors. 1) Spinal SAH due to spinal metastases should be considered in the differential diagnosis of patients with previously known malignancy. 2) Spinal SAH may manifest without paraparesis or sensory deficit. 3) Magnetic resonance imaging of the spinal cord may be important to determine the source of SAH in patients in whom four-vessel cerebral angiography demonstrates no abnormal findings. PMID- 15658133 TI - Novel treatment of basilar invagination resulting from an untreated C-1 fracture associated with transverse ligament avulsion. Case report and description of surgical technique. AB - The authors describe the case of a traumatic C-1 ring fracture and transverse ligament injury in an otherwise healthy adult woman; the lesion was essentially untreated for 3 months and resulted in basilar invagination. On presentation 3 months postinjury, the patient complained of severe increasing suboccipital pain and a grinding sensation in her upper neck. Axial computerized tomography (CT) scans revealed a C-1 ring fracture, basilar invagination with the dens abutting the clivus, and significant lateral splaying of the C-1 lateral masses. Flexion extension radiography demonstrated abnormal motion at the atlantoaxial junction. A unique surgical technique was used to address simultaneously the C1-2 instability, the displaced C-1 fracture, and basilar invagination without having to perform occipital fixation. The authors believe that an understanding of the mechanism of the cranial settling in this case (further splaying of the C-1 lateral masses and downward migration of the occipital condyles) permitted full reduction of the deformity; this was accomplished by performing a horizontal reduction of the C-1 lateral masses, using direct C-1 lateral mass screws, a rod compressor, and a cross-link. Postoperative CT scanning confirmed the success of reduction. The results in this report highlight a rare but important complication of untreated C-1 fracture and ligament disruption, and the authors describe a novel treatment technique with which to restore vertical alignment and preserve occipital C-1 motion. A variation of this technique may also be used to treat Type II transverse ligament injuries associated with C-1 ring fractures as an alternative to halo immobilization. PMID- 15658134 TI - Microsurgical infrapedicular paramedian approach for retrovertebral lumbar disc herniations. Technical note. AB - The author performed a microsurgical infrapedicular paramedian approach in 35 patients (23 men and 12 women) to remove herniated lumbar retrovertebral discs that did not have an apparent origin at either the superior or inferior disc level. The goal of this surgery was to minimize the bone resection, preserve the facet joint, and avoid the risk of secondary vertebral instability. The Macnab outcome classification was used to assess all patients who attended follow-up examination for at least 15 months. The clinical results were excellent or good in 34 (97%) of 35 cases. This minimally invasive lumbar spine technique resulted in minimal morbidity, excellent clinical benefits, and a long-term outcome without evidence of secondary segmental instability. PMID- 15658135 TI - The development of techniques for resection of spinal cord tumors by Harvey W. Cushing. AB - Harvey Cushing's refinement of Halsted's meticulous surgical techniques facilitated safe resection of intradural spinal tumors. Although Cushing focused his attention on brain tumors at the Peter Bent Brigham Hospital, his numerous contributions to the treatment of intradural spinal tumors include the description of these tumors' natural histories and their histological classifications. The application of his experienced intracranial techniques to the resection of spinal tumors improved outcomes. The authors review selected operative notes and sketches to demonstrate his technique in the excision of the spinal cord tumors. PMID- 15658136 TI - The physician and the dying patient: a question of control? PMID- 15658137 TI - Planning care for non-oncologic terminal illness in advanced age. AB - Due to the increase in longevity today, advanced illness in the elderly exists together with severe disability and often dementia that generally become less responsive to known treatment. This leads to repeated admissions to an internal ward in a general hospital, which results not only in a lack of treatment continuity but also in inappropriate management resulting in over- or under treatment. Towards the end of their lives, the treatment problems of non oncologic elderly patients with advanced diseases stem from a number of factors: multiple pathology, difficulty in predicting irreversibility, staff reluctance to discontinue active specific treatment and resort to palliative care only, and the lack of a framework to ensure continuity of treatment in the community or hospital. These advanced systemic illnesses are characterized by fluctuating exacerbations and remissions, making it very difficult to assess irreversibility. This article proposes the establishment of advanced centralized care planning, based on community care, the geriatric hospital and, in particular, a geriatric support unit within the skilled nursing department, catering holistically for the ongoing needs of the patient and his/her family and supplying a backup to the community care. PMID- 15658138 TI - Characteristics of road traffic accidents treated in an urban trauma center. AB - BACKGROUND: Road traffic accidents are the leading cause of accidental injury and death for persons under the age of 35. The medical literature presents surprisingly little information on the general characteristics of such accidents in the urban setting. OBJECTIVES: To characterize RTA patients arriving at an urban trauma center. METHODS: We prospectively examined the charts of all patients admitted to the Tel Aviv Sourasky Medical Center due to RTA injuries during two periods in 1995. RESULTS: Of the 1,560 patients examined, the male:female ratio was 1:1 and median age was 27 years (47% aged 20-30 years); 51% of the accidents took place between 8 a.m. and 4 p.m. and on working week days; automobiles comprised 47.1% of the vehicles involved, motorized two-wheel vehicles 37.1%, bicycles 3.8%, and pedestrians 12%. The Glasgow Coma Scale was 15 on arrival in 98.7% of the patients. The trunk was the most commonly injured body part (84.7%); whiplash injury to the neck was diagnosed in 343 patients (22%), and brain concussion in 183 (11.7%). Computed tomography studies were performed in 34 patients (2.2%). The vast majority of patients (1,438, 92.2%) was discharged home; 14 (0.9%) were admitted to the intensive care unit, and 2 (0.13%) died during hospitalization. The average time spent in the emergency department in the morning shift was 2.1 hours. CONCLUSIONS: We could identify distinguishing factors of this population: equal gender distribution, peak RTA incidence in the young adult working population during working hours, automobile drivers being the most injured subgroup, a disproportionate number of motorcycle and motor scooter involvement, and a relatively extensive amount of time and resources spent treating these injuries despite their generally minor nature. PMID- 15658139 TI - Determinants affecting initiation of continuous positive airway pressure treatment. AB - BACKGROUND: Continuous positive airway pressure is the treatment of choice for patients with obstructive sleep apnea syndrome. OBJECTIVE: To determine the factors influencing treatment initiation with a CPAP device in a healthcare system in which co-payment is required. METHODS: A total of 400 adult patients with OSAS who required CPAP therapy completed questionnaires at three different stages of the diagnostic and therapeutic process: CPAP titration study (stage 1), patient adaptation trial (stage 2), and purchase of a CPAP device (stage 3). Logistic regression was used to analyze the variables influencing CPAP use at the different stages of the diagnostic and therapeutic processes. RESULTS: Only 32% of the patients who underwent CPAP titration study purchased a CPAP device. The number of subjects who purchased a CPAP device increased gradually as monthly income increased, 28% vs 62% in the "very low" and "very high" income levels respectively. Reporting for the titration increased in patients with excessive daytime sleepiness and an Epworth Sleepiness Scale score above 9 (odds ratio = 1.9, P = 0.015). Higher socioeconomic status increased reporting to stage 2 (OR = 1.23, P = 0.03) and CPAP purchase (stage 3, OR = 1.35, P = 0.002). Excessive daytime sleepiness increased reporting to stage 2 (OR = 2.28, P = 0.006). Respiratory disturbance index above 35 increased CPAP purchasing (OR = 2.01, P = 0.022). Support from the bed partner, referring physician and sleep laboratory team increased CPAP purchasing. CONCLUSIONS: A supportive environment for a patient with OSAS requiring CPAP is crucial to increase initiation of CPAP treatment. Minimizing cost sharing for the CPAP device will reduce inequality and may increase CPAP treatment initiation. PMID- 15658140 TI - Hepatic lymphoma: an imaging approach with emphasis on image-guided needle biopsy. AB - BACKGROUND: Involvement of the liver by lymphoma is almost always secondary. A definite diagnosis can be made only after histologic examination. Recently there has been a trend to replace surgical biopsies with percutaneous core-needle biopsies for the diagnosis of lymphoproliferative disorders. OBJECTIVES: To describe our experience with percutaneous image-guided needle biopsies of the liver in 15 cases of primary and secondary hepatic lymphoma. METHODS: Between 1997 and 2002, 15 of all the percutaneous computerized tomography-guided core needle liver biopsies performed at our institution yielded the diagnosis of lymphoma. We retrospectively reviewed the medical records of these patients. RESULTS: Seven patients had primary hepatic lymphoma (all non-Hodgkin's lymphoma) and eight had secondary (three Hodgkin's disease and five non-Hodgkin's lymphoma). No major complications were caused by the percutaneous biopsies, and all biopsies were diagnostic. The imaging findings were non-specific but were characteristic and similar to previously described series. Imaging demonstrated hypodense lesions by CT, or hypoechoic or anechoic lesions by ultrasound in all but two cases in which hilar lesions resulted in biliary dilatation, both demonstrated by ultrasound. CONCLUSIONS: Review of our primary cases indicated no association with cirrhosis or AIDS in contradistinction to the worldwide experience. There were no significant complications in the 15 patients in the study, and a definite diagnosis of lymphoma was made in all the cases with no need to proceed to surgical biopsy. We highly recommend image-guided core-needle biopsy of the liver as a reliable and useful tool for the diagnosis of hepatic lymphoma. PMID- 15658141 TI - Restorative proctocolectomy with ileal pouch-anal anastomosis for ulcerative colitis and familial adenomatous polyposis: twenty years follow-up in 174 patients. AB - BACKGROUND: Restorative proctocolectomy with ileal pouch-anal anastomosis has become the surgical procedure of choice for patients with ulcerative colitis and familial adenomatous polyposis. OBJECTIVES: To evaluate the long-term functional outcome of patients who underwent this surgical procedure. METHODS: We performed this observational study in 174 consecutive patients: 146 with UC and 28 with FAP. The patients, 91 males and 83 females with a mean age of 34.1+/-10.6 years (range 6-67 years), underwent the procedure between January 1984 and January 2004 (mean follow-up 64.8 months, range 1-240 months). The indications for surgery were intractable disease in 124 patients (71%), dysplasia in 36 (21%), severe bleeding in 8 (5%), and perforation in 6 (3%). RESULTS: A protective ileostomy was performed in 140 patients (96%) with UC and 12 (43%) with FAP. An urgent three-stage procedure was necessary in 14 patients (8.4%). A mucosal proctectomy was performed in 94 (54%), and a double stapling technique in 80 (46%). Mean length of hospital stay was 9.4+/-6.6 days (range 5-34 days, median 8). Complications included pelvic sepsis in 7 patients (4.2%), anastomotic leakage in 8 (4.8%), bowel obstruction in 22 (13.2%), incisional hernia in 12 (7.2%), anastomotic stenosis that usually responded to manual dilatation in 46 (27.6%), pouchitis in 106 (61%), recto-vaginal fistula in 3 (1.8%), retrograde ejaculation in 3 (1.8%), and impotence in 2 (1.2%). There was no mortality in this group of patients. The median number of bowel movements per 24 hours was six in UC patients and five in FAP patients, with at least one bowel movement during the night. Complete daytime and night-time continence was documented in 124 patients (71%). Overall satisfaction was 95%. CONCLUSIONS: Restorative proctocolectomy with ileal pouch-anal anastomosis confers a long-term good quality of life to both UC and FAP patients, and the majority of patients are fully continent with five to six bowel movements per day. PMID- 15658142 TI - Ultrasonography as a diagnostic modality in acromioclavicular joint pathologies. AB - BACKGROUND: Ultrasound is useful in detecting acromioclavicular pathologies in cases of trauma, inflammations and degenerative changes. OBJECTIVES: To describe the sonographic findings of acromioclavicular joint pathology in patients with anterior shoulder pain. METHODS: Sonographic examination of the ACJ was used to examine 30 adults with anterior shoulder pain. As a control group we studied 30 asymptomatic patients and compared the findings to plain radiographs of the symptomatic group. RESULTS: The pathologic findings consisted of swelling of the joints, bone irregularities, widening and/or narrowing of the ACJ, soft tissue cyst formation, excessive fluid collection, and calcification. All these signs represent degenerative changes compatible with early osteoarthritis. We encountered one case of septic arthritis that required joint aspiration and antibiotic treatment. CONCLUSIONS: It is our belief that ultrasonography should be used routinely in cases of anterior shoulder pain since it demonstrates various pathologies undetected by plain radiographs. PMID- 15658143 TI - Colchicine inhibits heterotopic ossification: experimental study in rabbits. AB - BACKGROUND: Heterotopic ossification is a common complication of hip surgery and musculoskeletal or brain trauma. OBJECTIVES: To confirm by in vivo study that colchicine inhibits osteoblast cell proliferation with marked decrease in tissue mineralization. METHODS: Heterotopic ossification was induced in three groups of New Zealand white rabbits (females, 6 months old, weight 3-3.5 kg) by injecting 2 ml bone marrow drawn from the iliac crest into their right thigh muscle. To prevent heterotopic ossification, colchicine (0.25 mg/ day) was administered orally for 4 weeks to two groups of adult rabbits: group A (preload group)--1 week preceding bone marrow injection; group B--on day of injection; and group C- control group. RESULTS: After 4 weeks the rabbits were evaluated by radiographs and ultrasound for evidence of heterotopic ossification. At the end of the study histologic samples were taken from all the thighs. Imaging and histologic studies showed, with statistical significance, almost complete prevention of heterotopic ossification formation in group A (preload) and a marked decrease in group B, when compared with the controls where large new bone had formed at the injection site. These results indicated the inhibitory effects of colchicine on a bone forming process in soft tissue such as heterotopic ossification. CONCLUSIONS: The role of colchicine in preventing heterotopic ossification in other bone-forming conditions, such as hip arthroplasty or pelvic trauma, and after brain trauma, remains to be evaluated in a clinical setting. PMID- 15658144 TI - Prenatal exposure to influenza and the risk of subsequent development of schizophrenia. AB - Various events occurring during pregnancy might influence the normal neurogenesis of fetus brain, including exposure to the influenza virus. Several studies have attempted to find a relationship between exposure to influenza virus and the onset of schizophrenic behavior in childhood or adulthood, however results remain contradictory. In this review we describe several animal and human studies that show or do not show a relationship between exposure to the influenza virus during pregnancy and the subsequent development of schizophrenia. PMID- 15658145 TI - The who and where of care: non-oncologic terminal illness in the elderly. PMID- 15658146 TI - Ending in a crash. PMID- 15658147 TI - Revisiting old ghosts: prenatal viral exposure and schizophrenia. PMID- 15658148 TI - Delayed recurrence of group D salmonella mycotic aneurysm of the abdominal aorta presenting as an aorto-duodenal fistula. PMID- 15658150 TI - Recurrent hemarthrosis of the knee mimicking pigmented villonodular synovitis. PMID- 15658149 TI - Osteoporosis and amenorrhea in a young patient with von Hippel-Lindau disease. PMID- 15658151 TI - Fracture and migration of a sternal metal suture: a possible cause of post coronary bypass intermittent chest pain. PMID- 15658152 TI - An observant Jewish physician working in a secular ethical society: ethical dilemmas. PMID- 15658153 TI - Medical students' ethical, legal and cross-cultural experiences during their clinical studies. AB - BACKGROUND: Many medical school curricula include training for ethical considerations, legal comprehension, implementation of patients' rights, awareness of cultural differences, and communication skills (ELCE). OBJECTIVES: To explore medical students' perceptions of their ELCE training during the clinical phase as well as the relationship between humanistic practice skills' experiences and the quality of clinical training. METHODS: A cross-sectional survey was carried out in two cohorts during their clinical year period at Tel Aviv University's Sackler Faculty of Medicine at the end of their Internal Medicine and Surgery clerkships in the 2002 academic year. The research tool was an 18 item Likert-type questionnaire (ELCEQ), based on the literature of biomedical ethics, legal aspects and behavior of practice skills. The content validation of the questionnaire was established by consulting experts among the school's faculty. It was circulated among the students by representatives of the Unit of Medical Education. RESULTS: The response rate was 88%. Students reported only a few opportunities for gaining experience in humanistic practice skills. A weak correlation was found between students' assessment of the quality of clinical training and their experiences in humanistic practice skills. CONCLUSIONS: A wider and more relevant range of active experiences in humanistic practice skills should be available to students during the clerkships. Correspondingly, there is a need for the clinical faculty to find innovative ways to internalize their task as role models and ensure that students acquire and are able to practice those skills. PMID- 15658154 TI - Lead removal from foundry waste by solvent extraction. AB - Solvent extraction is used to reduce lead concentrations from millpond wastewater solids, a type of foundry process waste. Toluene and toluene mixed with di-(2 ethyl-hexyl) phosphoric acid (HDEHP) have been tried as leaching solvents. Toluene is ineffective as a solvent in extracting lead, but the toluene-HDEHP mixture effectively removes lead from solid foundry waste. The effects of the HDEHP concentration, the contact time, and the amount of solvent used on lead extraction have been investigated. The mass transfer process is rapid: contact time of 1/2 hour has been found to be sufficient to accomplish the leaching process. The concentration of HDEHP significantly impacts lead removal. The optimum concentration of HDEHP is determined to range from 0.05 to 0.1 mol/l. The Toxicity Characteristic Leaching Procedure (TCLP) test of the treated samples gives leachable lead in much lower quantities than those found in the untreated samples. Thus the solvent extraction process appears to be an effective method to significantly reduce the lead content of millpond wastewater solids. PMID- 15658155 TI - Wastes of multilayer containers as substrate in composting processes. AB - An evaluation of the influence and degradation of ground multilayer containers, such as Tetra Brik Aseptic (TBA), in the composting process of municipal solid wastes (MSW) is presented. Two composting piles were formed by mixing, in different proportions, an organic fraction of MSW and TBA material ground as flakes or stripes. Piles were periodically aerated by mechanical mixing. Evolution of the composting was followed by frequent measurements of suitable parameters such as temperature, moisture, ashes, total organic carbon (TOC), nitrogen, C/N ratio, pH, conductivity, nutrients, and heavy metals. As expected, aluminum and polyethylene films from TBA were not degraded during the processes, but the cardboard fraction of TBA showed a partial decomposition that was more intense in the material ground as flakes. After two months of curing, the composts were refined and analyzed to determine their nutrient and heavy-metals contents. Despite their poor aesthetic properties, mainly resulting from the presence of little pieces of plastic and aluminum films, the composts contained the required amounts of nutrients; furthermore, their heavy-metals content was below the limits proposed by some official European organizations. PMID- 15658156 TI - An observation-based model for analyzing ozone precursor relationships in the urban atmosphere. AB - An Observation-Based Model (OBM) is described, which uses in-situ atmospheric observations to determine the sensitivity of ozone concentrations in an urban atmosphere to changes in the emissions of ozone precursors (i.e., volatile organic compounds and nitrogen oxides). The model is formulated following the concept of Relative Incremental Reactivity (RIR) developed by Carter and Atkinson. In the OBM, however, observed concentrations rather than emission inventories are used to drive the photochemical simulations and thereby ensure that the calculations are carried out for the proper mix of nitrogen oxides and volatile organic compounds. From these calculations, a series of sensitivity factors, or RIRs, are inferred that can be used to (1) determine whether reducing emissions of nitrogen oxide or emissions of hydrocarbons would be most effective in abating ozone in a given urban area, and (2) identify the most critical subset of hydrocarbons present in an urban atmosphere causing ozone exceedances. Because the OBM is relatively easy and inexpensive to operate and makes use of data that are increasingly available, it can be used to analyze a wide array of ozone episodes and, thus, could prove to be a relatively cost-effective tool for the analysis of ozone precursor relationships in an urban atmosphere. On the other hand, because the OBM is diagnostic rather than prognostic, it cannot be used in a predictive mode to estimate exactly how much emission reduction is needed to reduce ozone concentrations. For this reason, the OBM should be viewed as a complement to, rather than a substitute for, more sophisticated gridded, emission based models. To illustrate the characteristics of the OBM and to demonstrate its applicability, we first compare the results of the OBM to those obtained from a series of simulations of the Atlanta metropolitan area using the Urban Airshed Model (UAM), a three-dimensional Eulerian grid model. The OBM is then used to analyze a dataset obtained from the 1990 Atlanta Ozone Study, an EPA field sampling program conducted during the summer of 1990. Because of limitations and potential flaws in the 1990 Atlanta dataset, the results of this OBM analysis are largely illustrative rather than definitive. Nevertheless, a few important issues are elucidated by the analysis. These include (1) the importance of accounting for biogenic hydrocarbons produced from urban vegetation; (2) the potential flaw in using early-morning VOC-to-NOx ratios to infer whether ozone production is limited by VOC or NOx; (3) the critical need for high-sensitivity nitrogen oxide measurements to quantify the sub-ppbv concentrations of NO during the afternoon hours; and (4) the need to consider a number of individual ozone episodes in studying an urban atmosphere because of the possibility that the degree of VOC- and NOx-limitation may vary from one episode to another. PMID- 15658157 TI - Theoretical study of liquid droplet dispersion in a venturi scrubber. AB - The droplet concentration distribution in an atomizing scrubber was calculated based on droplet eddy diffusion by a three-dimensional dispersion model. This model is also capable of predicting the liquid flowing on the wall. The theoretical distribution of droplet concentration agrees well with experimental data given by Viswanathan et al. for droplet concentration distribution in a venturi-type scrubber. The results obtained by the model show a non-uniform distribution of drops over the cross section of the scrubber, as noted by the experimental data. While the maximum of droplet concentration distribution may depend on many operating parameters of the scrubber, the results of this study show that the highest uniformity of drop distribution will be reached when penetration length is approximately equal to one-fourth of the depth of the scrubber. The results of this study can be applied to evaluate the removal efficiency of a venturi scrubber. PMID- 15658158 TI - Estimations of real-world N2O emissions from road vehicles by means of measurements in a traffic tunnel. AB - Ambient air measurements of N2O, NOx, CO, and HC based on grab sampling were conducted in a major traffic tunnel in Sweden, that carries up to 4,000 vehicles per hour, in order to estimate real-world emissions of N2O for road traffic. Two different methods--relative and mass balance-were used to calculate a N2O emission factor for the mixed vehicle fleet, which gave an average emission factor, at average speeds of 30-70 km/h, of approximately 25 mg N2O/ km, with a range of 7-56 mg/km. PMID- 15658159 TI - Application of dispersion modeling to indoor gas release scenarios. AB - Many complex models are available to study the dispersion of contaminants or ventilation effectiveness in indoor spaces. Because of the computationally complex numerical schemes employed, most of these models require mainframe computers or workstations. However, simple design tools or guidelines are needed, in addition to complicated models. A dispersion model based on the basic governing equations was developed and uses an analytical solution. Because the concentration is expressed by an analytical solution, the grid size and time steps are user definable. A computer program was used to obtain numerical results and to obtain release history from a thermodynamic source model. The model can be used to estimate three-dimensional spatial and temporal variations in concentrations resulting from transient gas releases in an enclosure. The model was used to study a gas release scenario from a pressurized cylinder into a large ventilated building, in this case, a transit parking and fueling facility. PMID- 15658160 TI - Evaluation of VOC emission measurement methods for paint spray booths. AB - Interest in regulations to control solvent emissions from automotive painting systems is increasing, especially in ozone nonattainment areas. Therefore, an accurate measurement method for VOC emissions from paint spray booths used in the automotive industry is needed to ascertain the efficiency of the spray booth capture and the total emissions. This paper presents the results of a laboratory study evaluating potential VOC sampling and analytical methods used in estimating paint spray booth emissions, and discusses these results relative to other published data. Eight test methods were selected for evaluation. The accuracy of each sampling and analytical method was determined using test atmospheres of known concentration and composition that closely matched the actual exhaust air from paint spray booths. The solvent mixture to generate the test atmospheres contained a large proportion of polar, oxygenated hydrocarbons such as ketones and alcohols. A series of identical tests was performed for each sampling/analytical method with each test atmosphere to assess the precision of the methods. The study identified significant differences among the test methods in terms of accuracy, precision, cost, and complexity. PMID- 15658161 TI - Validation of a test method for the measurement of methanol emissions from stationary sources. AB - Title III of the 1990 Clean Air Act Amendments designated methanol as a pollutant to be regulated. The U.S. Environmental Protection Agency (EPA), through a contract with Research Triangle Institute, has developed a method for measuring methanol emissions from stationary sources. The methanol sampling train (MST) consists of a glass-lined heated probe, two condensate knockout traps, and three sorbent cartridges packed with Anasorb 747. Samples are desorbed with a 1:1 mixture of carbon disulfide (CS2) and N,N-dimethylformamide (DMF). Condensate water and CS2/ DMF samples are analyzed by gas chromatography with flame ionization detection. The MST has a practical quantitation limit of approximately 3 ppm for a 20-L sample. Samples were shown to be stable for at least two weeks after collection. Field tests of the MST and the National Council of the Paper Industry for Air and Stream Improvement (NCASI) methanol sampling method were conducted at two pulp and paper mills. Sampling and analysis procedures followed EPA Method 301 requirements. The sampling location for the first field test was the inlet vent to a softwood bleach plant scrubber, where the methanol concentration was approximately 30 ppm. The mean recovery of spike was 108.3% for the MST method and 81.6% for the NCASI method. Although neither method showed significant bias at the 95% confidence level, the between-methods bias was significantly different. A second field test was conducted at a vent from a black liquor oxidation tank where the methanol concentration was approximately 350 ppm. Mean spike recoveries were 96.6 and 94.2% for the MST and NCASI methods, respectively. The biases of the two methods and the between-methods bias were not significantly different for the second field test. PMID- 15658162 TI - Pilot-scale demonstration of a two-stage methanotrophic bioreactor for biodegradation of trichloroethylene in groundwater. AB - A two-stage methanotrophic bioreactor system was developed for remediation of water contaminated with TCE and other chlorinated, volatile, aliphatic hydrocarbons. The first stage of the reactor was a suspended-growth culture vessel using a bubbleless methane-transfer device. The second stage was a plug flow bioreactor supplied with contaminated groundwater and cell suspension from the culture vessel. The test objectives were to determine the applicability of microbial culture conditions reported in the literature for continuous, pilot scale TCE treatment; the technical feasibility of plug-flow bioreactor design for treatment of TCE; and the projected economic competitiveness of the technology considering the cost of methane for growth of methanotrophs. The methanotrophic organism used in the study was Methylosinus trichosporium OB3b. Information on system operation was obtained in bench tests prior to conducting the pilot tests. In bench- and pilot-scale tests, variability in the degree of TCE degradation and difficulty in maintaining the microbial culture activity led to short periods of satisfactory biotreatment. Further development of the microbial culture system will be required for long-term operation. During transient periods of high TCE degradation activity, the bioreactor concept proved feasible by exhibiting both a high degree of TCE biodegradation (typically about 90% at influent TCE concentrations of 0.5-4 ppm) and a close approximation to first-order reactor kinetics throughout the length of the reactor. Actual methane usage in the pilot scale reactor resulted in projected methane costs of $0.33 per 1000 gallons of water treated. This cost theoretically would be reduced by system modifications. The theoretical minimum methane cost was approximately $0.05 per 1000 gallons. PMID- 15658163 TI - Assessing ozone-precursor relationships based on a smog production model and ambient data. AB - A semi-empirical model, Johnson's smog production model (SPM), which relates precursor emissions to ozone levels and estimates the relative effectiveness of volatile organic compounds (VOC) and NOx emission controls, has been evaluated and a modified version of SPM has been introduced. Both versions have been applied to routine data from 1989-1991 in five areas in the United States. In particular, extent parameters, which reveal the relative merit of VOC and NOx controls in reducing high ozone levels, have been calculated. Preliminary applications of SPM reveal interesting features with respect to VOC vs. NOx controls in reducing high ozone levels. For hourly data with ozone > or =0.08 ppm, distributions of extent parameters resulting from the modified SPM show the effectiveness of VOC controls at more monitoring sites than those from Johnson's SPM; however, relative features between the two versions are similar. On the other hand, for hourly data with ozone > or =0.12 ppm, the two SPM versions show very similar relative effectiveness of VOC and NOx controls with chosen values of model parameters. To improve the credibility of SPM, the range of validity of relationships between maximum smog produced or maximum ozone and NOx concentrations must be determined, and the parameters in these relationships must be better determined for typical VOC mixtures. Another essential parameter, which determines the fractional loss of NOy (NO and its oxidation products) from the gas phase must be better determined. PMID- 15658164 TI - Evaluation of an intensive sampling and analysis method for carbon monoxide. AB - This paper describes a portable, low-cost whole air sampler capable of collecting 12 1-L samples in tedlar bags and using a single "D" cell for power. The results are presented from two tests of an intensive sampling and analysis method for measuring ambient carbon monoxide (CO) concentrations in urban areas using this sampler. The method is evaluated in comparison with an approved continuous CO analyzer, in intercomparisons of the results of re-analysis of the samples, and by examination of the results from co-located samplers. The precision of the analytical method was found to be +/-0.30 ppmv. The precision of the sampling method was found to be +/-0.73 ppmv and therefore is the limiting factor in the method's overall precision. These values are sufficient to verify attainment of National Ambient Air Quality Standards (NAAQS) levels in urban areas. Improvements in the sampler and analytical procedure are discussed. PMID- 15658165 TI - Growth response of bigcone Douglas fir (Pseudotsuga macrocarpa) to long-term ozone exposure in southern California. AB - Long-term radial growth of bigcone Douglas fir (Pseudotsuga macrocarpa) was studied throughout its range in the San Bernardino Mountains of southern California, where ambient ozone has been high for approximately the past 40 years. A gradient of both ozone concentration and precipitation exists from west (high) to east (low). Growth rates of bigcone Douglas fir are considerably lower since 1950 throughout the San Bernardino Mountains, with the largest growth reductions in the western part of the range where ozone exposure is highest. Needle retention is also somewhat lower at high ozone sites. Lower annual precipitation since 1950 may have some impact on long-term growth reductions, and short-term growth reductions induced by drought are an important component of long-term growth reductions at sites with high ozone exposure. An ozone-climate stress complex may be responsible for recent reductions in the growth of bigcone Douglas fir. PMID- 15658166 TI - Vapor pressure response to denaturant and water in E10 blends. AB - On December 16, 1993, the U.S. Environmental Protection Agency (EPA) released the final rule on reformulated gasoline (RFG). This rule will affect the composition of as much as 45% of the gasoline used in the United States by the summer of 1995. The acceptance of any gasoline component lies in its ability to contribute to the RFG program's environmental goals. This study was conducted to determine the effect of water and ethanol denaturant on gasoline Reid vapor pressure (RVP) for which little quantitative data are available. This paper addresses two new areas where environmental goals may be achieved while maintaining the use of ethanol-blended gasolines within ozone nonattainment areas. PMID- 15658167 TI - Bench-scale testing of selected remediation alternatives for contaminated sediments. AB - The Assessment and Remediation of Contaminated Sediments (ARCS) Program within the U.S. Environmental Protection Agency's Great Lakes National Program Office (GLNPO) contained a component for demonstrating and evaluating sediment remediation technologies. Toward this end, bench-scale tests of solvent extraction, thermal desorption, and wet air oxidation technologies were conducted. Contaminated sediments were tested from the Grand Calumet River, Indiana; Buffalo River, New York; Saginaw River, Michigan; and Ashtabula River, Ohio. The primary contaminants of concern in these sediments were polychlorinated biphenyls (PCBs) and polynuclear aromatic hydrocarbons (PAHs). The solvent extraction tests were conducted with sediments from the Grand Calumet, Buffalo, and Saginaw rivers. The thermal desorption studies were conducted with sediments from the Grand Calumet, Buffalo, and Ashtabula rivers. The wet air oxidation testing was performed with the Grand Calumet River sediment. Raw sediment contaminant concentrations ranged from 0.32-21.9 mg/kg dry mass for PCBs and 2.70 266 mg/kg dry mass for PAHs. PCB removal or destruction efficiencies ranged from approximately 6-99%. PAH removal or destruction efficiencies ranged from 65-99%. Mass balance closures ranged from 40-99% for solids; 59-139% for water; 29-3500% for oil; 16-129% for PCBs; and 69-3170% for PAHs. PMID- 15658168 TI - Determining temporal and spatial variations in ozone air quality. AB - Unless the change in emissions is substantial, the resulting improvement in ozone air quality can be easily masked by the meteorological variability. Therefore, the meteorological and chemical signals must be separated in examining ozone trends. In this paper, we discuss the use of the Kolmogorov-Zurbenko filter in evaluating the temporal and spatial variations in ozone air quality utilizing ozone concentration data from several monitoring locations in the northeastern United States. The results indicate a downward trend in the ozone concentrations during the period 1983-1992 at most locations in the northeastern United States. The results also reveal that ozone is a regional-scale problem in the Northeast. PMID- 15658169 TI - Linear least squares method for time series analysis with an application to a methane time series. AB - A simple method of time series analysis, based upon linear least squares curve fitting, is developed. The method's advantages and disadvantages are discussed, and an example is presented using the Vostok Core methane record. PMID- 15658170 TI - Look before you leap. AB - Learning to critically evaluate therapeutic claims is vital to the success of the practicing veterinarian and the veterinary profession as a whole. Until such time as good scientific data in support of therapies are obtained, veterinarians should be careful in their advocacy and employment of new and unproved practices. They should also be aware of the many reasons why therapeutic "success" may be the result of a variety of factors unrelated to the treatment modality itself. Only by relying on rigorous standards of evidence can equine veterinarians prevent a return to the sincere but misguided ways of yesteryear. PMID- 15658172 TI - Practice management software for the equine practitioner. AB - Practice management software benefits the equine practitioner in many ways. Ready access to patient records, rapid entry and lookup of clinical findings, easy invoicing and monitoring of revenues, inventory management, rapid search and sorting of data, reminder and recall maintenance, and integrated word processing are just some of the software functions that assist the successful practitioner. Computer use in veterinary practice may have seemed to many to be a gimmick at one time. Now that computers have become a part of our everyday life, it is clear that they can be an essential component of the modem, successful, and profitable equine practice. PMID- 15658171 TI - Advances in equine practice management. AB - As equine practices move forward in the 21st century, we feel increasing pressure to manage our businesses like business people. This requires outside expertise, in-house business-trained managers, and continuing education on various aspects of practice life. Aside from the AAEP, several other organizations are providing continuing education in management, including the Association of Veterinary Practice Managers, the Veterinary Management Institute offered at Purdue University Krannert Graduate School of Management, the American Animal Hospital Association through (The Virginia-Maryland Regional College of Veterinary Medicine's) on-line practice management course, and the University of Illinois Equine Practice Management modules. PMID- 15658173 TI - Gene therapy: future therapies in osteoarthritis. AB - The field of equine veterinary practice is in an ever-evolving state, requiring current technologies to be constantly evaluated for new applications. The specific use of gene therapy in the horse is a novel application. The authors want to help familiarize the equine practitioner with the concept of gene therapy, and introduce its use and potential future benefits for the equine industry in the treatment of osteoarthritis. PMID- 15658174 TI - The future of antiinflammatory therapy. AB - The cells and mediators that make up the inflammatory response have the potential to injure tissues and contribute to the pathophysiology of many inflammatory diseases. Strategies to reduce neutrophil migration into sites of inflammation and subsequent activation by inhibiting integrin-mediated adhesion hold promise for successful treatment of a variety of inflammatory diseases. New pharmacologic agents that specifically target prostanoid mediators of inflammation by specifically inhibiting the activity of cyclooxygenase 2 are potent antiinflammatory agents with fewer gastrointestinal side effects than nonspecific cyclooxygenase inhibitors. These areas of antiinflammatory research are rapidly yielding drugs with diverse future applications in equine medicine. PMID- 15658175 TI - Advances in information technology. AB - Advances in digital technology have vastly increased the speed and content of information transfer. Equine practitioners of the future should be in contact with their office and other information resources at all times through an array of digital networks. Patients and practitioners alike should benefit from better information access. PMID- 15658177 TI - The internet and the future of equine practice. AB - The Internet is a fast-growing technology that allows information (text as well as images) to be shared across the world in an instant. As computer systems, software, and Internet services become less expensive options, the world becomes more experienced and comfortable with these technologies. In the foreseeable future, equine veterinarians are likely to be better educated about diseases and treatments, better connected with specialists for consultations on difficult cases, able to respond to client communications more efficiently, able to accurately educate clients, and able to communicate with colleagues from across the country and the world. Additionally, there is likely to be a time when using the Internet and computers is profitable in all these ways as well as economically. Improving doctor-client relationships and patient care are goals for all veterinarians, and the Internet holds the technology to render these now and in the future. PMID- 15658176 TI - Digital imaging advances and the future. AB - Digitization of clinical data is becoming increasingly important in veterinary medicine. Current veterinary applications range from digital cameras to computed radiography. The opportunity for increased productivity and diagnostic capabilities makes the computerization of veterinary medicine especially exciting. Future trends seem to indicate that more veterinarians should have better technology available in their practices and more veterinarians should be connected to experts, clients, and each other. PMID- 15658178 TI - Advances in equine ultrasonography. AB - The digital storage and transfer of images coupled with the development of the Internet have revolutionized diagnostic imaging. With the press of a button, practitioners can exploit the diagnostic skills of specialists anywhere in the world and enlist their help with challenging cases. This easy transfer of images and digital data should stimulate a greater number of multicenter studies so that the sensitivity and specificity of ultrasonographic techniques can be established in larger and more meaningful cohorts of patients. There is now no doubt that we can identify a plethora of conditions of the horse with ultrasonography; however, there is a continuing need to look critically at how we use such information to improve patient management and outcome. PMID- 15658179 TI - Equine dental advances. AB - The reintroduction and development of safe motorized instruments, the increased availability of continuing education, and the understanding and implementation of appropriate procedures allow practitioners to provide better dental care. Veterinarians realize that sedation, analgesia, a full-mouth speculum, and proper instrumentation are necessary to provide these services. Continued instrument design, future research, and new treatment and prophylactic protocols should have a positive impact on the future of equine dental health. New and rediscovered procedures for equilibrating equine occlusion are allowing horses to masticate more efficiently, carry a bit more comfortably, and experience improved performance. The horse, the horse owner, and the veterinary profession all benefit from providing complete equine dental care. PMID- 15658180 TI - Respiratory applications for the future: one perspective. AB - Respiratory diagnostic instruments previously available only to referral centers should be converted into miniature, portable, wireless medical devices for use in practice. Internet and wireless connections of these instruments should expedite the process of seeking second opinions and in general information sharing. We are moving into a world of greater objectivity that should free us from speculation in some areas, yet open new avenues for research and innovation in others. PMID- 15658182 TI - Behavior and misbehavior of the horse. AB - For decades after the discipline of psychiatry had been established as an accepted specialty, many medical schools continued to fail to train their students in the fundamentals of this discipline. Medical students all have at least cursory exposure to psychiatric principles and basic psychology. Unfortunately, the veterinary profession has lagged behind human medicine in this regard. Until recently, veterinary students received no training in animal behavior, and there were no available residencies within our schools for developing board-certified behavioral specialists. PMID- 15658181 TI - Frontier medicine: the future and integrative medicine. AB - Vigorous and prolonged effort is required to gain true mastery of the healing arts. Conventional and complementary medicine have complementary strengths and weaknesses. Like the yin and yang of traditional Chinese medicine, they naturally flow into one another by a process of induction, creating balance. Integrative medicine is the frontier; it is the future. If we are to progress beyond our current understanding and ability to heal, we must work with theoretic models that allow us and our perception to operate "outside the box." For some, this understanding is intuitive. It is through cooperative and collaborative efforts of intuitively adept and technologically adept minds that we can integrate and advance our understanding; increase our ability to predict, prevent, and diagnose disease; and expand our therapeutic options. PMID- 15658183 TI - Limitations to veterinary applications of new technologies in treatment and diagnostics. AB - Technology promises to improve the lifestyle and life quality of humankind. As a rule, wherever human medicine goes, veterinary medicine is sure to follow. Nevertheless, the promise of technologic advances does not shine as bright for veterinarians as for human physicians. This trend is echoed in the business of animal health as pharmaceutic company after pharmaceutic company spins off or otherwise eliminates their animal health division. Instead, a small group of strictly animal health-oriented companies compete for the animal health dollar, promising that fewer and fewer expensive technologies are likely to be incorporated into the standard of veterinary practice. All is not lost, however, because as progress is made in the field of human biotechnology, the cost of the technology should eventually come down, permitting at least some of the advances in human medicine to become available to the veterinarian. PMID- 15658185 TI - A confidential inquiry estimating the number of patients affected with sickle cell disease and thalassemia major confirms the need for a prevention strategy in the Netherlands. AB - We have conducted a broad confidential inquiry among 401 hospital departments trying to estimate the number of patients affected with severe forms of hemoglobinopathies living in The Netherlands. With less than 30% response we have registered 559 patients in all age categories of whom 77.0% are affected with sickle cell disease and 17.5% with beta-thalassemia (thal) major. We estimate that the real figure could be around 800 patients, a figure more than six times higher than the number published in 1995 on which the reluctance to offer prevention was based. The actual figures and the incidence estimation of approximately 60 patients a year underline the urgent need for the official implementation of a prevention strategy in The Netherlands. During the last 5 years we have been working towards the implementation of a multi-intervention strategy for primary prevention using the existing structures of public health. The obstacles we have encountered to endorse such a strategy are discussed as a possible guide for other immigration countries. PMID- 15658184 TI - Hb Hope [beta136(H14)Gly-->Asp (GGT-->GAT)]: interactions with Hb S [beta6(A3)Glu ->Val (GAG-->GTG)], other variant hemoglobins and thalassemia. AB - Hb Hope [beta136(H14)Gly-->Asp (GGT-->GAT)] was first described in an African American family in 1965. Since then, it has been found in combination with several different globin gene mutations in many other families of divergent ethnic backgrounds. The basis for its relatively frequent occurrences remains unexplained. This variant hemoglobin (Hb) is mildly unstable and has reduced oxygen affinity, but is generally innocuous clinically. This variant Hb can present as a confounding factor in arriving at a correct diagnosis by either electrophoresis or high performance liquid chromatography (HPLC), particularly during the neonatal period. DNA-based diagnostics can help solve this potential problem. PMID- 15658186 TI - Prevalence of the sickle cell gene in Yemen: a pilot study. AB - To determine the prevalence of the sickle cell gene (betaS) in Yemen and amongst people from different regions of the country living in the capital, Sana'a City, cord blood samples from 1,500 consenting mothers were collected from hospitals in Sana'a City between July and December 2001. The names and original homes of the parents were recorded. Cation exchange high performance liquid chromatography (HPLC) analysis was used for screening, while isoelectric focusing (IEF) and DNA polymerase chain reaction (PCR) were used to confirm Hb S [beta6(A3)Glu-->Val]. Thirty-three samples were found to show Hb FAS, giving an overall likely betaS gene frequency of 0.011. The betaS gene frequency varied with the part of the country from which the parents came. Amongst people from Taiz and Haja in the west, the gene frequency was more than 0.04, but less than 0.004 amongst people from Ibb, adjacent to the governorate (administrative division) of Taiz. Of 66 chromosomes from babies carrying the betaS gene, only 1.5% also carried the -158 (C-->T) Ggamma-globin gene XmnI site compared with 16.1% of 168 chromosomes from babies without the betaS gene from the same regions. The results of this study show a higher betaS gene frequency in the western coastal part of Yemen than in the central mountainous and eastern desert areas. The incidence of affected homozygous births may therefore reach 20/10,000 in certain areas, although it is much lower than this overall. Limited health resources can best be invested in developing a program of education, screening and health care, initially prioritizing those communities residing in the western areas of Yemen with the highest betaS gene frequency. PMID- 15658187 TI - A novel approach to rapid determination of betaS-globin haplotypes: sequencing of the Agamma-IVS-II region. AB - beta-Globin gene cluster haplotypes were originally determined by restriction endonuclease mapping with Southern blots of polymorphic sites around the gene cluster. Over the years, haplotyping has been found to be useful, not only in population genetics but also in predicting the severity of hemoglobinopathies such as sickle cell disease. The sickle mutation occurs on five distinct haplotypes. The hitherto used methods are cumbersome and time-consuming, making haplotype determination a tedious procedure. We report our experience with a novel, rapid approach to haplotyping based on sequence polymorphisms in the Agamma-IVS-II region. We provide an algorithm that allows rapid assignment of the four African haplotypes carrying the sickle mutation. PMID- 15658188 TI - Study on molecular epidemiology of the alpha-thalassemias in Liuzhou City, Guangxi Autonomous Region, China. AB - Guangxi is one of the provinces of Southern China with the highest incidence of alpha-thalassemia (thal). Liuzhou is the second biggest city in Guangxi. To find out the incidence of the various alpha-thal genotypes, and their distribution in the Liuzhou area, an F820 Blood Cell Analysis System was used to measure the parameters of red blood cells. A SPIFE Rapid Auto-Electrophoresis System was used to analyze the normal and abnormal hemoglobins (Hbs). Multiplex polymerase chain reaction (mPCR) was used to detect the alpha-globin genotypes. Thirty-two (2.05%) out of 7805 young couples undergoing pre-marriage counseling, were diagnosed as having an Hb H (beta4) disease. The study of 1228 cord blood samples revealed 138 newborn children carrying an alpha-thal determinant with nine different genotypes, thus making the total incidence of alpha-thal 11.24%. Among 185 cases of Hb H, 119 (64.1%) were confirmed as being deletional, and 66 cases (35.7%) nondeletional types. The severity of the Hb H diseases could be classified in the following order: alphaCSalpha/--SEA (alphaConstant (Spring)alpha/--Southeast Asia); alpha(-4.2)/--SEA; alpha(-3.7)/--SEA. Ten cases of alpha-thal determinants were found in combination with beta-thal. The mPCR technique can detect all kinds of combinations of the three common large deletions (--SEA, alpha(-4.2) and alpha(-3.7)) accurately and conveniently. PMID- 15658189 TI - Hb Johnstown [beta109(G11)Val-->Leu]: A high oxygen affinity variant associated with beta0-thalassemia. AB - Hb Johnstown [beta109(G11)Val-->Leu], a high oxygen affinity hemoglobin (Hb) variant associated with beta0-thalassemia (thal) [IVS-I-1 (G-->A)], was identified in an 8-year-old girl referred to our laboratory because of erythrocytosis and a left-shifted oxygen dissociation curve (ODC). The phylogenetic tree showed that the mother was heterozygous for the Hb variant and the father was a beta0-thal carrier. This Hb variant, with normal electrophoresis, was characterized at the DNA level by beta gene sequencing. The amino acid substitution potentially disrupts alpha1beta1 contacts i n the deoxyHb conformation, thus shifting the equilibrium towards the high affinity oxyHb conformation. The erythrocytosis and low values for actual P50 due to Hb Johnstown were more marked due to the co-inheritance of the beta0-thal. PMID- 15658190 TI - Molecular characterization and diagnosis of Hb Crete [beta129(H7)Ala-->Pro]. AB - We report the molecular characterization of Hb Crete [beta129(H7)Ala-->Pro] in a female subject from the Greek island of Crete. DNA sequence analysis revealed a 1368 GCC-->CCC base substitution in exon 3 of the beta-globin gene, leading to the Ala-->Pro amino acid change at codon 129. Both the proband and her mother, who were found to be heterozygotes for Hb Crete, presented with mild microcytic anemia and normal Hb A2 levels and iron metabolism indices. This is the first description of an heterozygous Hb Crete case, and also the first report on the molecular basis of Hb Crete. Moreover, the proposed NlaVI restriction enzyme based detection of Hb Crete at the DNA level is a fast and accurate approach, useful for molecular diagnostics. PMID- 15658191 TI - Prenatal diagnosis in a family at risk for beta-thalassemia and hemophilia A: an uncommon association. AB - beta-Thalassemia (thal) is an autosomal recessive disorder with a prevalence of 2 3% in Indians, while hemophilia A is X-linked with a prevalence of 1 in 5,000 10,000 male births. The chances of both these disorders being present together is extremely rare (1 in 250,000). We report an interesting consanguineous family from Western India with a combination of these two disorders, which was referred to us for prenatal diagnosis. PMID- 15658192 TI - A new highly unstable alpha chain variant causing alpha(+)-thalassemia: Hb Zurich Albisrieden [alpha59(E8)Gly-->Arg (alpha2)]. AB - A new alpha-globin mutation causing persistent mild hypochromic microcytosis and erythrocytosis is described. Hb Zurich Albisrieden [alpha59(E8)Gly-->Arg (alpha2)] is not detected at the protein level and leads to alpha(+)-thalassemia (thal). PMID- 15658193 TI - Hb Dhonburi (Neapolis) [beta126(H4)Val-->Gly] identified in a family from northern Iran. AB - Thalassemias are the most common hereditary diseases in Iran, resulting from synthesis defects in one or more hemoglobin (Hb) subunits. The majority of patients suffer from beta-thalassemia (thal), but cases with microcytic hypochromic anemia and normal electrophoretic patterns are suspected to have alpha- or silent beta-thal. A family from the northern part of Iran, an area highly prevalent for thalassemias, was referred to us for prenatal diagnosis. The hematological data of the father indicated a pattern of beta-thal minor. Reverse hybridization analysis for the most common beta-globin mutations identified IVS II-1 (G-->A) in the heterozygous state. The maternal laboratory data indicated a case more compatible with alpha-thal. Iron deficient anemia was ruled out, and common alpha-thal point mutations and deletions were investigated. As no mutation was detected, chain synthesis was performed and showed an alpha/beta chain ratio of 2.1, that was in the range of beta-thal minor. DNA sequencing of the entire beta-globin gene identified a heterozygous GTG-->GGG (Val-->Gly) mutation at codon 126, also known as Hb Dhonburi (Neapolis). Prenatal diagnosis of the fetal DNA showed the absence of the IVS-II-1 and codon 126 anomalies. This result demonstrates the importance of screening of individuals with mild microcytic hypochromic anemia for both alpha- and silent beta-thal mutations. PMID- 15658194 TI - Hemoglobinopathies in the Christmas Island population. AB - Christmas Island is a remote Australian territory 2,400 km north of Perth. Health care is administered from Perth. The population is predominantly Chinese, with some Malay, Indian and European. As hemoglobinopathies are known to be common amongst these ethnic groups, a study was performed to determine their prevalence and significance in the Christmas Island population. Three-hundred and sixty-four individuals (adults and children) were tested. All subjects were assessed by full blood count, alpha-globin multiplex polymerase chain reaction (PCR) and PCR testing for Hb Constant Spring [alpha142, Term-->Gln, TAA-->CAA (alpha2)]. Microcytic patients (MCV <80 fL) were further investigated by high performance liquid chromatography (HPLC) and serum ferritin was determined. Where present, beta-thalassemia (thal) mutations were characterised by PCR. Thirty-four subjects (9.3%) were microcytic and of these five were iron deficient. The remainder were heterozygous for a hemoglobinopathy, giving a 9.1% incidence of hemoglobinopathies in Christmas Islanders. alpha-Thalassemia was identified in 23 subjects, seven of whom were heterozygous for alpha(-3.7); the remaining 16 were heterozygous for the - -SEA deletion. One case of heterozygous deltabeta-thal and one case of heterozygous Hb E [beta26(B8)Glu-->Lys] was detected. Of the eight subjects heterozygous for beta-thal, at least five mutations are represented, indicating a diverse and heterogeneous origin for this population. PMID- 15658195 TI - What it takes to be a hero. PMID- 15658197 TI - The ABCs of buying or selling a dental practice. PMID- 15658196 TI - A supernumerary tooth fused to the distal surface of a mandibular third molar. AB - Fusion is a tooth anomaly considered to be the result of the union between two adjacent tooth buds. The incidence of fusion is less than 1 percent of the population. The following is a case report of a unique form of a supernumerary tooth fused to the distal surface of a mandibular third molar. Radiographic evaluation demonstrates the communication of the pupal tissues. A differential diagnosis of developmental anomalies is discussed. PMID- 15658198 TI - Preparedness: a healthcare provider's role.... PMID- 15658199 TI - Don Quixote, DMD. PMID- 15658200 TI - Oral pathology quiz #35. Case number 1. Mucoepidermoid carcinoma. PMID- 15658201 TI - Victory for the vulnerable. PMID- 15658202 TI - NP care vs. physician care: an analysis of recent outcomes research. PMID- 15658204 TI - Transitioning from the military. PMID- 15658203 TI - Giving up the ghost. PMID- 15658205 TI - Congress ponders health issues. PMID- 15658206 TI - Primary and secondary prevention of coronary heart disease. Evidence offers multiple strategies. PMID- 15658207 TI - Butts out. Incorporating smoking cessation into your practice. PMID- 15658208 TI - United we stand. Using grassroots efforts to achieve legislative change. PMID- 15658209 TI - Addressing the health care needs of children in foster care. An innovative approach. PMID- 15658210 TI - Urinary control devices for stress incontinence in women. New options are on the horizon. PMID- 15658211 TI - Constipation in the elderly. Nonpharmacologic approaches are safe and effective. PMID- 15658212 TI - Rock bottom. Recognizing alcoholism in your patients. PMID- 15658213 TI - Treating the recovering addict. PMID- 15658214 TI - A portable fiber-optic chemical device for the quantitative determination of carbon monoxide from automobile exhaust emissions. AB - A colorimetric method for the quantitative determination of CO by diffuse reflectance is described. This method is based on the reduction by CO of Mo (VI) from the indicator reagent molybdosilicic acid (H8Si[Mo2O7]6). The reduction yielded a change of color from clear yellow to dark green on white disk filter chart paper wetted with reagent indicator solution. The gaseous mixture containing CO was forced to pass through this chart paper, initiating the reaction. The intensity of the color produced, measured by diffuse reflectance, was proportional to the CO concentration present in exhaust gases in the range from 0.02 to 12% volume/volume (v/v). A 650-nm light-emitting diode was used as a light source. A two-fiber-optic system carried the light from the source to the detection system, which was composed of a photodiode, an amplification circuit, and a digital display. The method was applied with success in field measurements for automobiles in the Otto cycle. In a previous paper, this method was used for the quantitative determination of exhaust emissions from diesel-fueled vehicles. PMID- 15658215 TI - Monitoring odorous sulfur emissions using self-organizing maps for handling ion mobility spectrometry data. AB - Possibilities for monitoring emissions of reduced sulfur compounds in pulp and paper mills were investigated using ion mobility spectrometry (IMS) and a self organizing map (SOM) algorithm. The reduced sulfur compounds measured were hydrogen sulfide (H2S), dimethyl sulfide (DMS), and methyl mercaptan (MM). Attention was paid to momentary concentrations because there is no monitoring device able to measure peak concentrations of reduced sulfur compounds under field conditions. These methods were evaluated by measuring the reduced sulfur compounds first in the laboratory and then at a process monitoring site at a pulp factory. The aim was to find out whether it would be possible to use the laboratory measurements to recognize the same reduced sulfur compounds at the monitoring site. Data collection was followed by analysis using the SOM algorithm and Sammon's mapping. The results showed that the IMS spectra of reduced sulfur compounds and their mixtures can be distinguished from each other by computationally intelligent methods and that the spectra from the process monitoring site corresponded with the laboratory measurements to a certain extent. PMID- 15658216 TI - Analysis of atmospheric conditions during air pollution episodes in Ankara, Turkey. AB - This paper presents a detailed analysis of the meteorological conditions that are associated with strong air pollution episodes in Ankara, Turkey. Based on climatological and air quality data [SO2 and TSP (total suspended particulates)] obtained for the winter months during 1989-1994, the analysis showed that the presence of weak atmospheric pressure gradients and warm air advection were the most important factors leading to high SO2 and TSP concentrations. In addition, the onset of the high air pollution episodes was generally associated with a trend toward negative vorticity at the 850-hPa level. PMID- 15658217 TI - The capture of heavy metals from incineration using a spray dryer integrated with a fabric filter using various additives. AB - This study investigated the effects of feedstock additives [polyvinyl chloride (PVC) and NaCl] and spray dryer additives (SiO2, CaCl2, NaHCO3) on heavy metal and fly ash removal efficiencies, and on particle size distribution of heavy metals. A spray dryer with an integrated fabric filter was used as an air pollution control device (APCD). Removal efficiencies for fly ash and heavy metals were greater than 95 and 90%, respectively. When additives of PVC or NaCl were used, the concentration of heavy metals distributed in fly ash apparently varied when the particle diameter was <1 microm. Although the effects of the additives SiO2, CaCl2, and NaHCO3 on the elemental size distribution of Cr were insignificant, these additives did slightly increase concentrations of Cd, Zn, and Pb partitioning in coarser particles (>1 microm). PMID- 15658218 TI - Speciated hydrocarbon and carbon monoxide emissions from an internal combustion engine operating on methyl tertiary butyl ether-containing fuels. AB - In the present work, engine and tailpipe (after a three-way catalytic converter) emissions from an internal combustion engine operating on two oxygenated blend fuels [containing 2 and 11% weight/weight (w/w) methyl tertiary butyl ether (MTBE)] and on a nonoxygenated base fuel were characterized. The engine (OPEL 1.6 L) was operated under various conditions, in the range of 0-20 HP. Total unburned hydrocarbons, carbon monoxide, methane, hexane, ethylene, acetaldehyde, acetone, 2-propanol, benzene, toluene, 1,3-butadiene, acetic acid, and MTBE were measured at each engine operating condition. As concerns the total HC emissions, the use of MTBE was beneficial from 1.90 to 3.81 HP, which were by far the most polluting conditions. Moreover, CO emissions in tailpipe exhaust were decreased in the whole operation range with increasing MTBE in the fuel. The greatest advantage of MTBE addition to gasoline was the decrease in ethylene, acetaldehyde, benzene, toluene, and acetic acid emissions in engine exhaust, especially when MTBE content in the fuel was increased to 11% w/w. In tailpipe exhaust, the catalyst operation diminished the observed differences. Ethylene, methane, and acetaldehyde were the main compounds present in exhaust gases. Ethylene was easily oxidized over the catalyst, while acetaldehyde and methane were quite resistant to oxidation. PMID- 15658219 TI - Development of emission factors for polyamide processing. AB - Emission factors for selected volatile organic compounds (VOCs) and particulate material were developed during processing of commercial grades of polyamide 6, polyamide 66, and polyamide 66/6 resins. A small commercial-type extruder was used, and melt temperatures ranged from 475 to 550 degrees F. An emission factor was calculated for each substance measured and is reported as pounds released to the atmosphere per million pounds of polymer resin processed. Scaled to production volumes, these emission factors can be used by processors to estimate emission quantities from similar polyamide extrusion operations. PMID- 15658220 TI - Economic comparison of emission control systems for glass manufacturing furnaces with heat recovery. AB - Glass manufacturing, like other process industries, is faced with air pollution compliance problems due to ever stricter emission limits. Several waste gas cleaning equipment options are available for air pollution control (APC) in glass plants, the most common arrangements being based on electrostatic precipitator (ESP) or fabric filter (FF) dust collectors and semi-wet or dry processes for acid gas removal. However, several counteracting aspects affect the choice of gas cleaning technologies, which are confirmed by the discrepancies encountered in actual suppliers' bids. In this paper, the main pollution control options are analyzed by carrying out a critical comparison under the cost-effectiveness point of view to select the lowest cost arrangement considering capital investment, operating expenses, and energy-saving revenues from heat recovery processes. The analysis is carried out with reference to a case study involving actual float glass production lines at Pilkington plants in Italy. PMID- 15658221 TI - Modeling the effects of VOC/NOx emissions on ozone synthesis in the cascadia airshed of the Pacific Northwest. AB - A modeling system consisting of MM5, Calmet, and Calgrid was used to investigate the sensitivity of anthropogenic volatile organic compound (VOC) and oxides of nitrogen (NOx) reductions on ozone formation within the Cascadia airshed of the Pacific Northwest. An ozone episode that occurred on July 11-14, 1996, was evaluated. During this event, high ozone levels were recorded at monitors downwind of Seattle, WA, and Portland, OR, with one monitor exceeding the 1 hr/120 ppb National Ambient Air Quality Standard (at 148 ppb), and six monitors above the proposed 8 hr/80 ppb standard (at 82-130 ppb). For this particular case, significant emissions reductions, between 25 and 75%, would be required to decrease peak ozone concentrations to desired levels. Reductions in VOC emissions alone, or a combination of reduced VOC and NOx emissions, were generally found to be most effective; reducing NOx emissions alone resulted in increased ozone in the Seattle area. When only VOC emissions were curtailed, ozone reductions occurred in the immediate vicinity of densely populated areas, while NOx reductions resulted in more widespread ozone reductions. PMID- 15658222 TI - An extended self-organizing map network for modeling and control of pulse jet fabric filters. AB - Pulse jet fabric filters (PJFFs) have become an attractive option of particulate collection utilities, because they can meet stringent particulate emission limits regardless of variation in operating conditions. Despite their wide applications, the present control algorithm for PJFFs can best be described as rudimentary. In this paper, a modeling and control strategy based on the local model network (LMN) is proposed. An extended self-organizing map (ESOM) network is developed to construct the LMN model of the filtration process using the filter's input-output data. Subsequently, these ESOM local models are incorporated into the design of local generalized predictive controllers (GPC), and the proposed controller design is obtained as the weighted sum of these local controllers. Simulation results show that the proposed controller design yields a better performance than both conventional GPC and proportional plus integral (PI) controllers yield. PMID- 15658223 TI - Washing of marine coastal sand in a batch reactor: sorption and desorption of BTEX. AB - This study addresses the issues related to decontamination of marine beach sand accidentally contaminated by petroleum products. Sorption and desorption of BTEX (i.e., benzene, toluene, ethylbenzene, and xylene) onto the sand from Uran Beach, located near the city of Mumbai, India, were studied, and isotherms were determined using the bottle point method to estimate sorption coefficients. Alternatively, QSARs (i.e., quantitative structure activity relationships) were developed and used to estimate the sorption coefficients. Experiments for kinetics of volatilization as well as for kinetics of sorption and desorption in the presence of volatilization were conducted in a fabricated laboratory batch reactor. A mathematical model describing the fate of volatile hydrophobic organic pollutants like BTEX (via sorption and desorption in presence of volatilization) in a batch sediment-washing reactor was proposed. The experimental kinetic data were compared with the values predicted using the proposed models for sorption and desorption, and the optimum values of overall mass transfer coefficients for sorption (K(s)a(s)) and desorption (K(d)a(d)) were estimated. This was achieved by minimization of errors while using the sorption coefficients (Kp) obtained from either laboratory isotherm studies or the QSARs developed in the present study. Independent experimental data were also collected and used for calibration of the model for volatilization, and the values of the overall mass transfer coefficient for volatilization (K(g)a(g)) were estimated for BTEX. In these exercises of minimization of errors, comparable cumulative errors were obtained from the use of Kp values derived from experimental isotherms and QSARs. PMID- 15658225 TI - Toxic emissions from mobile sources: a total fuel-cycle analysis for conventional and alternative fuel vehicles. AB - Mobile sources are among the largest contributors of four hazardous air pollutants--benzene, 1,3-butadiene, acetaldehyde, and formaldehyde--in urban areas. At the same time, federal and state governments are promoting the use of alternative fuel vehicles as a means to curb local air pollution. As yet, the impact of this movement toward alternative fuels with respect to toxic emissions has not been well studied. The purpose of this paper is to compare toxic emissions from vehicles operating on a variety of fuels, including reformulated gasoline (RFG), natural gas, ethanol, methanol, liquid petroleum gas (LPG), and electricity. This study uses a version of Argonne National Laboratory's Greenhouse Gas, Regulated Emissions, and Energy Use in Transportation (GREET) model, appropriately modified to estimate toxic emissions. The GREET model conducts a total fuel-cycle analysis that calculates emissions from both downstream (e.g., operation of the vehicle) and upstream (e.g., fuel production and distribution) stages of the fuel cycle. We find that almost all of the fuels studied reduce 1,3-butadiene emissions compared with conventional gasoline (CG). However, the use of ethanol in E85 (fuel made with 85% ethanol) or RFG leads to increased acetaldehyde emissions, and the use of methanol, ethanol, and compressed natural gas (CNG) may result in increased formaldehyde emissions. When the modeling results for the four air toxics are considered together with their cancer risk factors, all the fuels and vehicle technologies show air toxic emission reduction benefits. PMID- 15658224 TI - Demonstrating attainment of the air quality standards: integration of observations and model predictions into the probabilistic framework. AB - This paper introduces an integrated observational-modeling approach to transform the deterministic nature of attainment demonstrations of the National Ambient Air Quality Standard (NAAQS) into the probabilistic framework. While the methods presented here can be used to address any air quality standard that is based on extreme values, this paper focuses on the application to the 1-hr and 8-hr NAAQS for ozone. Extreme value statistics and resampling techniques are applied to estimate the probability of exceeding the NAAQS for both 1-hr and 8-hr ozone concentrations. Within the integrated observation-modeling analysis approach, we show that the model-to-model differences in the predicted responses to emission reductions are smaller than the model-to-model differences in predicted absolute ozone concentrations. We illustrate that the emission reductions stemming from a real-world emission control strategy would substantially reduce the probability of exceeding the NAAQS over a large portion of the eastern United States, especially for the 8-hr average ozone concentrations. PMID- 15658226 TI - Sensitivity analysis and evaluation of microFacCO: a microscale motor vehicle emission factor model for CO emissions. AB - This paper presents a sensitivity analysis of a microscale emission factor model (MicroFacCO) for predicting real-time site-specific motor vehicle CO emissions to input variables, as well as a limited field study evaluation of the model. The sensitivity analysis has shown that MicroFacCO emission estimates are very sensitive to vehicle fleet composition, speed, and ambient temperature. For the present U.S. traffic fleet, the CO emission rate (g/mi) is increased by more than 500% at 5 mph in comparison with a speed greater than 40 mph and by approximately 67% at ambient temperatures of 45 degrees F and > or = 95 degrees F in comparison with an ambient temperature of 75 degrees F. The input variable "emission failure standard rate" is more sensitive to estimating emission rates in the 1990s than in the 2000s. The estimation of emission rates is not very sensitive to relative humidity. MicroFacCO can also be applied to examine the contribution of emission rates per vehicle class and model year. The model evaluation is presented for tunnel studies at five locations. In general, this evaluation study found good agreement between the measured and the modeled emissions. These analyses and evaluations have identified the need for additional studies to update the high speed (>35 mph) air conditioning (A/C) correction factor and to add effects due to road grades. MicroFacCO emission estimates are very sensitive to the emission standard failure rate. Therefore, the model performance can be greatly improved by using a local emission standard failure rate. PMID- 15658227 TI - Health effects of waste incineration: a review of epidemiologic studies. AB - There is an increasing trend toward using incineration to solve the problem of waste management; thus, there are concerns about the potential health impact of waste incineration. A critical review of epidemiologic studies will enhance understanding of the potential health effects of waste incineration and will provide important information regarding what needs to be investigated further. This study reviews the epidemiologic research on the potential health impact of waste incineration. Previous studies are discussed and presented according to their study population, incinerator workers or community residents, and health end points. Several studies showed significant associations between waste incineration and lower male-to-female ratio, twinning, lung cancer, laryngeal cancer, ischemic heart disease, urinary mutagens and promutagens, or blood levels of certain organic compounds and heavy metals. Other studies found no significant effects on respiratory symptoms, pulmonary function, twinning, cleft lip and palate, lung cancer, laryngeal cancer, or esophageal cancer. In conclusion, these epidemiologic studies consistently observed higher body levels of some organic chemicals and heavy metals, and no effects on respiratory symptoms or pulmonary function. The findings for cancer and reproductive outcomes were inconsistent. More hypothesis-testing epidemiologic studies are needed to investigate the potential health effects of waste incineration on incinerator workers and community residents. PMID- 15658228 TI - Do trust inspections truly reflect hospital hygiene? PMID- 15658229 TI - Brian Belle-Fortune a voice from acute nursing. PMID- 15658230 TI - Has the public health white paper gone too far? PMID- 15658231 TI - Has the public health white paper gone too far? PMID- 15658232 TI - Consent--obtaining permission to care. PMID- 15658233 TI - Breathing new life into COPD care. PMID- 15658234 TI - The comfort zone. PMID- 15658235 TI - Understanding the importance to health of a balanced diet. AB - Obesity is one of the fastest-developing public health problems. The general population needs to have a balanced, healthy diet as it provides the energy and nutrients required to survive and stay healthy. Combining a healthy diet with an active lifestyle has huge health benefits and helps reduce the risk of major health problems such as heart disease, cancer and obesity. PMID- 15658236 TI - Links between drug and alcohol misuse and psychiatric disorders. AB - Substance use is common in a wide range of psychiatric disorders. Psychiatric disorder can increase the risks of such use developing into a substance use disorder. Conversely, substance use disorders are frequently accompanied by psychiatric symptoms. In some people such symptoms indicate formal psychiatric disorder. Whatever the causes, comorbidity may complicate treatment and resolution of both disorders, often presenting as a vicious circle that is difficult to break. PMID- 15658237 TI - Improving primary care services for people with learning disability. AB - The health of people with a learning disability is significantly poorer than that of the general population. In this project, two senior nurses developed a programme of inclusive health care aimed at enabling people who have learning disabilities to lead healthier lives. The project demonstrated that a joined-up approach is required across agencies to tackle the exclusion of people with a learning disability from mainstream health care services. PMID- 15658239 TI - Improving care of patients with inflammatory bowel disease. PMID- 15658240 TI - Time to grow. PMID- 15658238 TI - Four-step method of interpreting arterial blood gas analysis. AB - Arterial blood gas analysis can be complex. However, in many clinical areas the nurse is one of the first to see the results so they need to know whether immediate action is required. This article identifies a simple four-step approach that enables ABG interpretation even when all the complexities are not fully understood. PMID- 15658241 TI - A culture of respect. PMID- 15658242 TI - Health is the key. PMID- 15658243 TI - Relieving the pressure of childcare. PMID- 15658244 TI - Reaping the benefits of teamwork. PMID- 15658245 TI - Exploring new ways to learn. PMID- 15658246 TI - Healthier working lives across the UK. PMID- 15658247 TI - How will IWL affect the private sector? PMID- 15658248 TI - Mark Radcliffe welcomes the new year with a smile. PMID- 15658249 TI - [Overview: Atherogenic lipoproteins and their clinical significance]. PMID- 15658250 TI - [Clinical significance of plasma total cholesterol, free cholesterol and esterified cholesterol levels]. PMID- 15658251 TI - [HDL cholesterol]. PMID- 15658252 TI - [LDL cholesterol]. PMID- 15658253 TI - [Clinical significance of remnant-lipoprotein]. PMID- 15658254 TI - [Remnant-like particle-cholesterol (RLP-C)]. PMID- 15658255 TI - [Triglycerides (monoacylglycerol, diacylglycerol, triacylglycerol)]. PMID- 15658256 TI - [Chylomicron]. PMID- 15658258 TI - [Lipoprotein X (LP-X)]. PMID- 15658257 TI - [Lipoprotein and lipoprotein subfraction]. PMID- 15658259 TI - [Lipoprotein (a) and its phenotype]. PMID- 15658260 TI - [Beta-lipoproteins, sialic acid beta-lipoproteins]. PMID- 15658261 TI - [Phospholipid and phospholipid fraction]. PMID- 15658262 TI - [Serum total free fatty acid and its fatty acid species]. PMID- 15658263 TI - [Long-chain polyunsaturated fatty acid]. PMID- 15658264 TI - [Bile acid, bile acid profile, and various conjugated fractions]. PMID- 15658265 TI - [Small-dense low-density lipoproteins]. PMID- 15658266 TI - [Analysis of lipoprotein lipase activity, immunoreactive mass and gene]. PMID- 15658267 TI - [Hepatic triglyceride lipase (HTGL)]. PMID- 15658268 TI - [Lecithin-cholesterol acyltransferase (LCAT)]. PMID- 15658269 TI - [Phospholipase A]. PMID- 15658270 TI - [Acyl-coenzyme A: cholesterol acyltransferase (ACAT)]. PMID- 15658271 TI - [Cholesteryl ester transfer protein (CETP)]. PMID- 15658272 TI - [Fatty acid binding proteins]. PMID- 15658273 TI - [Apolipoprotein A-I, A-II, A-IV]. PMID- 15658274 TI - [Lipid-poor apolipoprotein A-I, glycated apolipoprotein A-I]. PMID- 15658275 TI - [Apolipoprotein B]. PMID- 15658276 TI - [Apolipoprotein C-I, C-II, C-III]. PMID- 15658278 TI - [Apolipoprotein J (apo J)]. PMID- 15658277 TI - [Apolipoprotein E and apolipoprotein E phenotype]. PMID- 15658279 TI - [Other apolipoproteins (apolipoprotein F, G, H, L, M and so on)]. PMID- 15658280 TI - [Icosapentanoic acid (EPA), docosahexanoic acid (DHA)]. PMID- 15658281 TI - [LDL receptor activity]. PMID- 15658282 TI - [Vitamin A (retinoids)]. PMID- 15658283 TI - [Retinol-binding protein (RBP)]. PMID- 15658285 TI - [Vitamin B1 (thiamin)]. PMID- 15658284 TI - [Beta-carotene]. PMID- 15658286 TI - [Vitamin B2 (riboflavin)]. PMID- 15658287 TI - [Vitamin B6 (pyridoxin)]. PMID- 15658288 TI - [Vitamin B12]. PMID- 15658289 TI - [Vitamin B12 absorption test]. PMID- 15658290 TI - [Vitamin C (ascorbic acid)]. PMID- 15658291 TI - [Vitamin D metabolites]. PMID- 15658292 TI - [Vitamin D-binding protein]. PMID- 15658293 TI - [Vitamin E and its subfraction]. PMID- 15658294 TI - [Vitamin K compounds]. PMID- 15658296 TI - [Folic acid]. PMID- 15658295 TI - [Vitamin P (flavonoid)]. PMID- 15658297 TI - [Nicotinic acid (niacin)]. PMID- 15658298 TI - [Biotin (vitamin H)]. PMID- 15658299 TI - [Pantothenic acid]. PMID- 15658300 TI - [Inositol]. PMID- 15658302 TI - [Bone alkaline phosphatase (BAP)]. PMID- 15658301 TI - [Ubiquinone (UQ), coenzyme Q (CoQ)]. PMID- 15658303 TI - [Tartrate resistant acid phosphatase]. PMID- 15658304 TI - [Hydroxyproline]. PMID- 15658305 TI - [Pyridinoline, deoxypyridinoline]. PMID- 15658306 TI - [Osteoclast-activating factors (OAF)]. PMID- 15658307 TI - [N-terminal propeptide of type I procollagen (PINP)]. PMID- 15658308 TI - [N-terminal crosslinking telopeptide of type I collagen]. PMID- 15658309 TI - [Pyridinoline cross-linked carboxy-terminal telopeptide of type I collagen (ICTP)]. PMID- 15658310 TI - [Carboxy-terminal propeptide of type I procollagen (PICP)]. PMID- 15658311 TI - [Beta-crosslaps]. PMID- 15658312 TI - [Sodium, potassium]. PMID- 15658313 TI - [Chloride]. PMID- 15658314 TI - [Magnesium (Mg)]. PMID- 15658315 TI - [Calcium, inorganic phosphate]. PMID- 15658316 TI - [Osmolarity]. PMID- 15658317 TI - [PCO2, PO2, SO2 in arterial blood]. PMID- 15658318 TI - [pH, plasma HCO3- in arterial blood]. PMID- 15658319 TI - [Exhaled nitric oxide (NO), carbon monoxide (CO)]. PMID- 15658320 TI - [Base excess and buffer base]. PMID- 15658321 TI - [Iron, total iron binding capacity (TIBC), unsaturated iron binding capacity (UIBC)]. PMID- 15658322 TI - [Copper]. PMID- 15658323 TI - [Zinc (Zn)]. PMID- 15658324 TI - [Chromium]. PMID- 15658325 TI - [Manganese]. PMID- 15658326 TI - [Cobalt]. PMID- 15658327 TI - [Selenium]. PMID- 15658329 TI - [Vanadium]. PMID- 15658328 TI - [Molybdenum]. PMID- 15658331 TI - [Lead]. PMID- 15658330 TI - [Aluminium]. PMID- 15658332 TI - [Cadmium]. PMID- 15658333 TI - [Mercury]. PMID- 15658334 TI - [Bismuth]. PMID- 15658336 TI - [Clinical evaluation of therapeutic drug monitoring]. PMID- 15658335 TI - [Nickel (Ni)]. PMID- 15658337 TI - [Doping control in sports]. PMID- 15658338 TI - [Benzodiazepine (clonazepam, nitrazepam, diazepam)]. PMID- 15658339 TI - [Hypnotic (barbiturate, non-barbiturate)]. PMID- 15658340 TI - [Antiepileptic drugs]. PMID- 15658341 TI - [Antidepressants]. PMID- 15658342 TI - [Butyrophenone derivatives]. PMID- 15658343 TI - [Benzamide-type anti-psychotic drugs]. PMID- 15658344 TI - [Lithium carbonate]. PMID- 15658345 TI - [Acetaminophen]. PMID- 15658346 TI - [Salicylic acid, aspirin]. PMID- 15658347 TI - [Steroidal anti-inflammatory drugs]. PMID- 15658348 TI - [Non steroidal anti-inflammatory drugs]. PMID- 15658349 TI - [Digoxin]. PMID- 15658350 TI - [Antiarrhythmic agents]. PMID- 15658351 TI - [Vasodilator: nitrate and nitrite]. PMID- 15658352 TI - [Bronchodilator: theophylline]. PMID- 15658353 TI - [Anti-allergy drugs]. PMID- 15658354 TI - [Glycopeptides (vancomycin, teicoplanin)]. PMID- 15658355 TI - [Aminoglycoside antibiotics]. PMID- 15658356 TI - [Beta-lactam antibiotics]. PMID- 15658357 TI - [Fluoroquinolones]. PMID- 15658358 TI - [Anti-HIV drugs]. PMID- 15658359 TI - [Zanamivir, oseltamivir]. PMID- 15658360 TI - [Alkylating agents, antimetabolites, alkaloids]. PMID- 15658361 TI - [Platinum derivatives]. PMID- 15658362 TI - [Topoisomerase I inhibitor (irinotecan hydrochloride)]. PMID- 15658363 TI - [Cyclosporine]. PMID- 15658364 TI - [Tacrolimus hydrate (tacrolimus)]. PMID- 15658365 TI - [Opiates]. PMID- 15658366 TI - [Amphetamines]. PMID- 15658367 TI - [Blood ethanol concentration]. PMID- 15658368 TI - [Trichloroacetic acid and trichloroethanol in urine]. PMID- 15658369 TI - [Mandelic acid in urine]. PMID- 15658370 TI - [Hippuric acid and methylhippuric acid in urine]. PMID- 15658371 TI - [N-methylformamide in urine]. PMID- 15658372 TI - [Endocrine disrupting chemicals--major EDCs and their analytical methods]. PMID- 15658373 TI - [p-Nitrophenol]. PMID- 15658374 TI - [Dioxin]. PMID- 15658375 TI - [Cotinine]. PMID- 15658376 TI - [Toluene]. PMID- 15658377 TI - [Dimethylamine]. PMID- 15658378 TI - [Phenol]. PMID- 15658379 TI - [General remarks: Qualitative and semi-quantitative analyses of medicinal and poisonous substances and their clinical significance]. PMID- 15658380 TI - [Arsenic]. PMID- 15658381 TI - [Cyanide]. PMID- 15658382 TI - [Methylmercury]. PMID- 15658383 TI - [Detection of paraquat dichloride and diquat dibromide]. PMID- 15658384 TI - [Sodium azide]. PMID- 15658385 TI - [Carbamates]. PMID- 15658386 TI - [Methanol]. PMID- 15658387 TI - [Peripheral blood analysis]. PMID- 15658388 TI - [Reticulocyte]. PMID- 15658389 TI - [Blood specific gravity]. PMID- 15658390 TI - [Erythrocyte sedimentation rate]. PMID- 15658391 TI - [Viscosity of blood and plasma]. PMID- 15658392 TI - [Neutrophil functions]. PMID- 15658393 TI - [Bleeding time, capillary resistance test]. PMID- 15658394 TI - [Clotting time and kaolin clotting time]. PMID- 15658395 TI - [Euglobulin lysis time (ELT)]. PMID- 15658396 TI - [Prothrombin time]. PMID- 15658397 TI - [Activated partial thromboplastin time (APTT), partial thromboplastin time (PTT)]. PMID- 15658398 TI - [Thrombo test (TBT), hepaplastin test (HPT)]. PMID- 15658399 TI - [Fibrinogen (factor I) and dys (functional) -fibrinogen]. PMID- 15658400 TI - [Soluble fibrin monomer complex (SFMC), soluble fibrin monomer (SFM)]. PMID- 15658401 TI - [Fibrin x fibrinogen degradation products (FDP)]. PMID- 15658402 TI - [FDP-D dimer and FDP-E fraction (FDP-E)]. PMID- 15658403 TI - [Fibrinogen and fibrin degradation products by leukocyte elastase]. PMID- 15658404 TI - [Fibrinopeptide A (FPA), fibrinopeptide B (FPB) and fibrinopeptide Bbeta(15-42) (FPBbeta15-42)]. PMID- 15658405 TI - [Cross-linked fibrin polymer and degradation products by plasmin (XDP)]. PMID- 15658406 TI - [Prothrombin (factor II) and prothrombin fragment 1+2 (F1+2)]. PMID- 15658407 TI - [Thrombin, antithrombin, thrombin-antithrombin complex (TAT)]. PMID- 15658408 TI - [TF (tissue factor), TFPI (tissue factor pathway inhibitor)]. PMID- 15658409 TI - [Factor V]. PMID- 15658410 TI - [Factor VII]. PMID- 15658411 TI - [Factor VIII and factor VIII inhibitor]. PMID- 15658412 TI - [von Willebrand factor (antigen and activity), multimer structure, and ristocetin cofactor activity]. PMID- 15658413 TI - [Factor IX and factor IX inhibitor]. PMID- 15658414 TI - [Factor X]. PMID- 15658415 TI - [Factor XI]. PMID- 15658416 TI - [Factor XII (Hageman factor)]. PMID- 15658417 TI - [Factor XIII]. PMID- 15658418 TI - [Fletcher factor, Fitzgerald factor]. PMID- 15658419 TI - [Heparin cofactor II (HCII)]. PMID- 15658420 TI - [Protein C inhibitor (PCI)]. PMID- 15658421 TI - [Molecular marker for anticoagulant protein C pathway (APC-PCI complex)]. PMID- 15658422 TI - [Activated protein C resistance (APC resistance)]. PMID- 15658423 TI - [Protein C, protein S, protein Z]. PMID- 15658424 TI - [Anti-phospholipid antibody (aPL)]. PMID- 15658425 TI - [Anticardiolipin antibody (aCL)]. PMID- 15658426 TI - [Lupus anticoagulant (LA)]. PMID- 15658427 TI - [Plasminogen (PG) and plasmin (PM)]. PMID- 15658428 TI - [Tissue plasminogen activator (t-PA) and urokinase plasminogen activator (u-PA)]. PMID- 15658430 TI - [Alpha2-plasmin inhibitor (alpha2PI)]. PMID- 15658429 TI - [Plasminogen activator inhibitor-1 (PAI-1)]. PMID- 15658431 TI - [Plasmin-alpha2 plasmin inhibitor complex (PIC)]. PMID- 15658432 TI - [t-PA x PAI-1 complex]. PMID- 15658434 TI - [Platelet function tests (aggregation, release reaction, adhesion)]. PMID- 15658433 TI - [TAFI]. PMID- 15658435 TI - [Analysis of platelet membrane glycoprotein]. PMID- 15658436 TI - [Platelet factor 4 (PF4)]. PMID- 15658438 TI - [Platelet-associated IgG]. PMID- 15658437 TI - [Beta-thromboglobulin (beta-TG)]. PMID- 15658439 TI - [Thrombomodulin]. PMID- 15658440 TI - [Hemoglobin (Hb) and its components]. PMID- 15658441 TI - [Abnormal Hb (hemoglobin)]. PMID- 15658442 TI - [Sulfhemoglobin]. PMID- 15658443 TI - [Methemoglobin]. PMID- 15658444 TI - [Measurement of glutathione]. PMID- 15658445 TI - [Genetics and biochemistry of NADH-cytochrome b5 reductase]. PMID- 15658446 TI - [Erythrocyte SOD]. PMID- 15658448 TI - [Leukocyte acid phosphatase]. PMID- 15658447 TI - [Na,K-ATPase in human red blood cell membranes]. PMID- 15658449 TI - [Myeloperoxidase]. PMID- 15658450 TI - [Neutrophil alkaline phosphatase score]. PMID- 15658451 TI - [Hexosaminidase isoenzyme pattern in leukemia]. PMID- 15658452 TI - [Eosinophil granule protein]. PMID- 15658453 TI - [Asialo GM1]. PMID- 15658454 TI - [Glucose-6-phosphate dehydrogenase (G6PD)]. PMID- 15658455 TI - [Phosphoglucomutase (PGM)]. PMID- 15658456 TI - [Erythrocyte glutathione reductase (EGR)]. PMID- 15658457 TI - [2' 5'-oligoadenylate synthetase (2-5AS)]. PMID- 15658458 TI - [Terminal deoxynucleotidyl transferase (TdT)]. PMID- 15658460 TI - [Erythrocyte transketolase]. PMID- 15658459 TI - [2,3-Diphosphoglycerate (2,3-DPG)]. PMID- 15658461 TI - [Sorbitol dehydrogenase (SDH)]. PMID- 15658462 TI - [Biochemistry and enzymatic diagnosis of red blood cell pyruvate kinase deficiency]. PMID- 15658463 TI - [Phosphofructokinase (PFK)]. PMID- 15658465 TI - Reinvestigation of clinical value of Rivalta reaction of puncture fluid. AB - Rivalta reaction is still used as a puncture fluid test for differentiation of exudate and transudate. However, the test method of Rivalta reaction has not been standardized, or positive precipitates for the reaction have not been investigated. Thus, we clarified the measurement method, and investigated Rivalta reaction-positive proteins. Rivalta reaction-positive punctuates converted to negative when pH increased to 4.6 or higher, showing the necessity of pH adjustment of acetic acid solution to 3.6-4.2 in Rivalta reaction. Using pH 4.0 acetic acid solution, 8 types of proteins were identified in Rivalta reaction positive turbid precipitates: C-reactive protein (CRP), alpha1-antitrypsin (alpha1-AT), alpha1-acid glycoprotein (alpha1-AG), haptoglobin (Hp), transferrin (Tf), ceruloplasmin (Cp), fibrinogen (Fg), and hemopexin (Hpx). Since these are acute reactive proteins, or proteins increased in malignant tumors and infections, positivity for Rivalta reaction at the specified pH may suggest pathological inflammation. PMID- 15658464 TI - [Other major red cell enzyme activities]. PMID- 15658466 TI - [Evaluation of SMBG values using various instruments and clinical significance of forearm SMBG measurement]. AB - Whole blood or plasma glucose values were compared between those measured using various instruments for SMBG and automated analyses in samples (antecubital vein) from 20 approximately 70 years old obtained at cookie tests. A good correlation between values in whole blood SMBG and plasma automated analyses values greater than r = 0.94 was observed for instruments A, C, F, and H, and the per cent difference from the automated values was less than 5% for A, B, E and H. Per cent difference of SMBG values by I was -16% in whole blood and -9% in plasma, suggesting the possibility of measuring real values in whole blood. With plasma, a good correlation greater than r = 0.95 was noted for A, C, F and K, and the per cent difference less than 10% was noted for C, E, F and I. A relatively good correlation (r: 0.63 approximately 0.90) between forearm SMBG value and plasma automated values was noted with the % difference of the mean less than 11%. During cookie test, there is no significant difference between forearm SMBG values and antecubital plasma automated values. Values at finger tips are significantly greater by 5 approximately 20% over automated plasma values. In conclusion, whole blood values by most of the SMBG instruments are well correlated with plasma automated values, although some disagreement was noted. Values at forearm are not different from plasma val ues of antecubital vein, while at finger tip SMBG values showed higher levels. The measurement at forearm is therefore recommended, and in addition the pain is less. PMID- 15658467 TI - Development of an enzyme-linked immunosorbent assay for fecal bile acid. AB - It has been suggested that the bile acids in the feces act as a promoter of colon cancer. Among the bile acids, deoxycholic acid (DCA), which is one kind of the secondary bile acid, is said to have strong influence. DCA/cholic acid (CA) ratio in feces is also said to have a diagnostic significance in colon cancer. With this in mind, we created a CA and DCA's monoclonal antibody (MoAb) to measure them through the enzyme linked immunosorbent assay (ELISA) method. Using these MoAb, we were able to measure CA and DCA concentrations with low cross-reaction to other bile acids compared with the method with polyclonal antibody (PoAb). We measured CA and DCA concentrations and calculated the DCA/CA ratios in healthy subjects and patients with colon cancer. All subjects had been screened for colon cancer. We then compared the healthy subjects, the cancer patients before surgery and the same cancer patients after surgery. Cancer patients after surgery had significantly low DCA/CA ratios compared to before surgery, whereas there was no significant difference between healthy subjects and the pre-operative colon cancer patients. PMID- 15658468 TI - [Relationship between aneuploidy of chromosome 17 and amplification of HER-2/neu gene in the different histological type of breast cancer]. AB - We examined the relationship between the aneuploidy of chromosome 17 and the amplification of HER-2/neu gene by fluorescence in situ hybridization (FISH) in the different histological type of breast cancer (n = 55). 51 patients recognized the aneuploidy that consisted of 37 patients of monosomy and 14 patients of polysomy. In the scirrhous carcinoma, HER-2/neu gene was amplified in 15% of monosomy cases, but it was not amplified in all polysomy cases. In the papillotubular carcinoma, HER-2/neu gene was contrastively amplified in all polysomy cases, but it was not amplified in all monosomy cases. No case of solid tubular carcinoma recognized the amplification. The relation was different according to the histological type of breast cancer. PMID- 15658469 TI - [Accreditation of clinical laboratories based on ISO standards]. AB - International Organization for Standardization (ISO) have published two international standards (IS) to be used for accreditation of clinical laboratories; ISO/IEC 17025:1999 and ISO 15189:2003. Any laboratory accreditation body must satisfy the requirements stated in ISO/IEC Guide 58. In order to maintain the quality of the laboratory accreditation bodies worldwide, the International Laboratory Accreditation Cooperation (ILAC) has established the mutual recognition arrangement (MRA). In Japan, the International Accreditation Japan (IAJapan) and the Japan Accreditation Board for Conformity Assessment (JAB) are the members of the ILAC/MRA group. In 2003, the Japanese Committee for Clinical Laboratory Standards (JCCLS) and the JAB have established the Development Committee of Clinical Laboratory Accreditation Program (CLAP), in order to establish the CLAP, probably starting in 2005. PMID- 15658470 TI - [Development of the reimbursement system based on DPC]. AB - In the health insurance system of Japan, a fee-for-service system has been applied to individual treatment services, but this fee system involves a structural problem of causing increases in examinations and drug administration. Various attempts have been made to solve this structural problem in several treatment fields, in which a flat payment system can be introduced with the fee for-service system as the basis. The diagnosis procedure combination (DPC) system introduced in April 2003, which is a flat payment system using a diagnosis classification, is the first large scale revision of the medical fee system of Japan. This diagnosis classification is considered to be effective for simplifying the medical fee system, within the framework of EBM, and for providing patients with information. However, since there are also structural problems in the flat payment system, such as examination and treatment of low quality, selection of patients, and upcoding, its introduction should be performed with sufficient caution. We will make more efforts to establish a better medical fee system by evaluating these problems. PMID- 15658471 TI - [Monoclonal antibody therapy and laboratory medicine]. AB - Monoclonal antibodies (mAbs) therapies have been introduced to the many kinds of malignancies. Rituximab (Rituxan), a chimeric anti-CD20 monoclonal antibody, has improved clinical outcome in the patients with B cell non-Hodgkin's lymphoma (NHL) as a single agent as well as combination with chemotherapies including CHOP and stem cell transplantation. The efficacy has been reported in the patients with follicular lymphoma as well as aggressive NHL. The expression of CD20 should be confirmed by immunopathological staining or flow cytometry before the treatment. Gemtuzumab ozogamicin (Mylotarg), calicheamicin conjugates of anti CD33 mAb, has been introduced in the treatment of AML. CD33 is not expressed by immature pluripotent stem cells even though expressed on myeloid progenitor cells. Mylotarg is internalized via CD33, however, detouched calicheamicin might be pumped out by multi-drug resistant related p glycoprotains (p-gp). The expression of CD33 and p-gp should be analyzed by flow cytometry before the treatment. We should give another caution to tumor-lysis syndrome and veno occlusive disease. PMID- 15658472 TI - [Report of cytological diagnosis: new reporting form for breast cytology]. AB - New reporting form for breast cytology was reported. The reporting form was consisted of diagnostic category and cytological findings. Diagnostic category was classified roughly into 2 of inadequate and adequate. Adequate was divided into four items of "normal or benign", "indeterminate", "suspicious of malignancy" and "malignant". Inadeguate was less than 10% of all cytological cases. "Indeterminate" cases established less than 10% of the adequate cases. The cases diagnosed to be "suspicious of malignancy" provided that it was desirable that more than 90% were malignant by the later histological diagnosis. In the cytological findings, expected histological type of the breast should be included. PMID- 15658473 TI - [Reporting system in surgical pathology]. AB - The questionnaire survey on quality assurance in pathology departments and certified hospitals was performed by Japanese Society of Pathology in 2001 and it showed that most institutions do not have guidelines for the reporting system. In this paper, how to write histopathology reports is described. The reports are composed of 1) clinical data, 2) pathology data, 3) macroscopic findings, 4) microscopic findings, 5) diagnoses, and 6) comments. Neither of the components should be made light of. We also describe the results of questionnaire survey on histopathology reports we performed. PMID- 15658474 TI - [Present status of and future aspects on geriatric care]. PMID- 15658475 TI - [Necessity and role of geriatric health services]. PMID- 15658476 TI - [Comprehensive geriatric care--concept and assessment of QOL]. PMID- 15658477 TI - [Comprehensive geriatrics--concept of terminal care]. PMID- 15658478 TI - [Comprehensive geriatrics--primary prevention]. PMID- 15658479 TI - [Primary geriatric care]. PMID- 15658480 TI - [Hypertension in elderly patients]. PMID- 15658481 TI - [Ischemic heart disease in elderly patients]. PMID- 15658482 TI - [Cerebrovascular disorders in elderly patients]. PMID- 15658483 TI - [Sleep apnea syndrome in elderly patients]. PMID- 15658484 TI - [Pneumonia in elderly patients]. PMID- 15658485 TI - [Diabetes in elderly patients]. PMID- 15658486 TI - [Urogenital disorders in elderly patients]. PMID- 15658487 TI - [Osteoporosis and other bone diseases in elderly patients]. PMID- 15658488 TI - [Outline of long-term care insurance system in Japan]. PMID- 15658490 TI - [Home care services covered by long-term care insurance system in Japan]. PMID- 15658489 TI - [Role of geriatric physicians in long-term care insurance system in Japan]. PMID- 15658491 TI - [Present status of geriatric care in the world]. PMID- 15658492 TI - [Genes in the aging process]. PMID- 15658493 TI - [Present status of and future aspects on clinical geriatrics in Japan (discussion)]. PMID- 15658494 TI - [Hepatic portal venous gas--a report of two cases successfully treated with conservative management]. PMID- 15658495 TI - [Early diagnosis of Legionella pneumonia with mental disturbance by urinary antigen]. PMID- 15658496 TI - [Masked diabetes insipidus manifested by administration of adrenocortical steroid hormone for erythroderma]. PMID- 15658497 TI - [Clinical safety management in internal medicine]. PMID- 15658498 TI - [Molecular mechanism in regulation of IGE production]. PMID- 15658500 TI - [Revised classification of lupus nephritis]. PMID- 15658499 TI - [Clinical study on bronchial asthma and leukotriene antagonists]. PMID- 15658501 TI - [Pharmacology of second-generation antipsychotics: a validity of the serotonin dopamine hypothesis]. AB - New atypical antipsychotic drugs such as risperidone, olanzapine and quetiapine, that have been modeled on the prototype agent clozapine and developed since the 1990's, are now referred to as second-generation antipsychotics (SGA). It has been proposed by Meltzer (1989) that the interaction between serotonin (5-HT) and dopamine (DA) systems may play a critical role in the mechanism of action for atypical antipsychotics because potent 5-HT2A receptor antagonism together with relatively weak D2 receptor antagonism could differentiate most atypical antipsychotics from typical antipsychotics. This serotonin-dopamine hypothesis has become a useful model for studying and developing new drugs to achieve a significant antipsychotic effect with lower incidence of extrapyramidal side effects compared to first-generation antipsychotics. In contrast, Kapur and Seeman (2001) argued the alternative "fast-off" theory that clozapine occupies D2 receptors to a similar extent as typical antipsychotics do and then rapidly dissociates from D2 receptors. This paper reviews the current issues on the serotonin-dopamine hypothesis and recent research on the role of 5-HT receptor subtypes in the mechanism of action for SGA. In particular, SGA-induced DA release in the prefrontal cortex, possibly through the functional activation of 5 HT1A receptors by 5-HT2A and D2 receptor-mediated interaction, may be the basis for the cognitive effects of SGA. PMID- 15658502 TI - [Neuropsychiatric disorders and GABA]. AB - In the mammalian central nervous system (CNS), the inhibitory GABAergic system is composed of different signaling molecules such as glutamate decaroxylase, vesicular GABA transporters, GABA receptors, GABA transporters and GABA transaminase. A prevailing view is that the balance between excitatory signaling mediated by glutamate and inhibitory signaling mediated by GABA plays a pivotal role in mechanisms underlying the modulation and maintenance of a variety of neural functions. Therefore, abnormalities in a GABAergic signaling molecule would lead to a crisis of severe symptoms relevant to a number of neuropsychiatric disorders. These include epilepsy, depression, schizophrenia, stiff-person syndrome, drug addiction and so on. In this review article, we will summarize recent studies on the relationship between the malfunction of GABAergic signaling molecules and the etiology of these neuropsychiatric disorders. We will also refer to novel strategies on GABAergic signaling molecules other than GABA receptors for therapeutic usefulness in the future. PMID- 15658503 TI - [Neuropsychological and neuropathological interactions in affective and cognitive geriatric syndromes]. AB - This paper reviews recent empirical findings related to depressive moods and cognitive functions in elderly persons. Vascular depression and major depression in Alzheimer disease (AD) are particularly common among elderly persons and, therefore, are discussed in light of the interactions among the cognitive deficits, the depressed moods and the neuropathologies. Functional imaging studies with depressed patients in several therapeutic or cognitive challenges have revealed that depression may involve impaired reciprocal interactions among the several important cognitive brain areas such as the hippocampus, anterior cingulate cortex and dorsolateral prefrontal cortex. Such impairment may be produced by the small ischemic changes in the subcortical areas of the frontal cortex in vascular depression and by the particular neurodegenerative processes in major depression in AD. PMID- 15658504 TI - The effectiveness of lithium augmentation of milnacipran: preliminary data using the modified Japanese Psychopharmacology Algorithm. AB - Medication algorithms based on the best evidence available together with expert consensus are considered to promote logical consistent clinical decision making in the choice of antidepressant drugs. We report our preliminary results using the modified algorithm established by the Japanese Psychopharmacology Algorithm Project (JPAP). Subjects were 24 patients with major depressive disorders who presented to the outpatient clinic of the Department of Psychiatry at the National Defense Medical College prior to any treatment for the current episode. Ultimately, 15 patients recovered with treatment according to our protocol, including 7 who recovered upon treatment with first-line drugs; the most effective of these was paroxetine, followed by fluvoxamine and then milnacipran. Six patients recovered with second-line treatments. Among these, a combination of milnacipran and lithium was most effective, with recovery of 4 of 4 patients. We have formed a strong impression that augmentation therapy, especially with milnacipran plus lithium, is likely to be effective if the first-line antidepressant is ineffective. Investigation of more cases will be needed to confirm or refine details of the algorithm and, more generally, to determine the best approach to antidepressant medication. PMID- 15658505 TI - [An animal model of posttraumatic stress disorder and its validity: effect of paroxetine on a PTSD model in rats]. AB - To better understand neuroscientific aspects of posttraumatic stress disorder (PTSD), it is necessary to establish an animal model of PTSD in which behavioral changes persist after initial traumatization. We administered inescapable footshock (IS) to male Wistar rats in a shuttle-box (inescapable stress session), and after 2 weeks we performed an avoidance/escape task session in the shuttle box using signal lights as anxiogenic external stimuli. Rats exposed to IS beforehand exhibited PTSD-like bi-directional behavioral changes, that is, "avoidance/numbing" (e.g. decreased activity, reactivity, and interest in surroundings) and "hyperarousal" (e.g. irritability and exaggerated responsiveness to external stimuli). Concretely speaking, in a relatively calm situation, spontaneous locomotion decreased during a 5-min adaptation period just before the avoidance/escape task session. On the other hand, in a stressful situation after starting the task session, not only avoidance responses to external stimuli (signal lights) but also gate-crossings during inter-trial interval increased. Accordingly, the paradigm used here could serve as a useful model of PTSD. We administered paroxetine (PRX) to rats just after IS for 2 weeks to examine its chronic effect on our animal model. Two weeks of PRX treatment significantly reduced hyperarousal-like behaviors, but no effect on avoidance/numbing-like ones was manifested. PMID- 15658506 TI - [Nicotine actions on behavior of man]. PMID- 15658507 TI - Nociceptin/orphanin FQ, hedonic state and the response to abused drugs. AB - The mesolimbic dopamine and endogenous opioid systems are foremost in our understanding of the neurochemical systems implicated in the control of motivation and reward. Several decades of study indicate that these systems interact to balance hedonic state and drive seeking for both natural and artificial and rewarding stimuli, such as food and abused drugs. Several classes of endogenous opioids have been identified, of which nociceptin (known also as orphanin FQ) is the most recently discovered. The relative contribution that each class of endogenous opioid peptides makes in determining basal and drug stimulated changes in hedonic state is still unclear. mu and delta opioid receptor agonists induce increases in hedonic state and are therefore rewarding. Kappa opioid receptor agonists, on the other hand, typically decrease hedonic state and are aversive. Although nociceptin is mostly devoid of hedonic properties when administered centrally, it effectively blocks the acquisition of conditioned place preferences to various rewarding drugs, suggesting that nociceptin reverses stimulated increases in hedonic state. Nociceptin also blocks the seeking of some rewarding stimuli in drug self-administration paradigms. Neurobiological explanations for these behavioral effects are incomplete; though a key finding has been that nociceptin suppresses basal and drug-stimulated activity in the mesolimbic dopamine system. Thus far, little is known about the role of naturally occurring endogenous nociceptin in the control of reward and motivated behaviors. Gaining an understanding in this area could contribute significantly to our understanding of how the brain regulates hedonic balance and motivation in both normal and perturbed states. have formed a strong impression that augmentation therapy, especially with milnacipran plus lithium, is likely to be effective if the first-line antidepressant is ineffective. Investigation of more cases will be needed to confirm or refine details of the algorithm and, more generally, to determine the best approach to antidepressant medication. PMID- 15658508 TI - [Japanese Genetics Initiative for Drug Abuse (JGIDA)]. PMID- 15658509 TI - [Current trend in physical dependence on benzodiazepine drugs by small animals]. PMID- 15658510 TI - [Antidepressants and substance dependence? A preclinical aspect (action mechanism, dependence characteristics, adverse effects and interactions of antidepressants)]. PMID- 15658511 TI - [Survey on the abuse of uncontrolled drugs by using the Internet]. PMID- 15658512 TI - [Spontaneous recrudescence of psychotic diseases caused by methamphetamine (a stimulant): the stress type and the role of noradrenaline and dopamine systems]. PMID- 15658513 TI - [Kidney protection--pain control--infection prevention. So that quality of life and cancer are not a contradiction]. PMID- 15658514 TI - [During immunosuppression threaten infections. Calculated antibiotic therapy in neutropenic patients]. PMID- 15658515 TI - [Coronary disease patient with infarction symptoms and ST-elevations in ECG. Should you have used thrombolysis, too?]. PMID- 15658516 TI - [Do the colleagues grasp laser too fast? "Age spot" abolished by laser -- melanoma grows further]. PMID- 15658517 TI - [The pain becomes continuously worse. Soft groin or pubic bone inflammation?]. PMID- 15658518 TI - [Childhood allergies are increasing enormously. Is it genetics or the environment?]. PMID- 15658519 TI - [How can we recognize allergies in childhood?]. AB - There is an increase in the prevalence of atopic disorders such as atopic dermatitis, allergic rhinitis/rhinoconjunctivitis. Apart from a positive family history, environmental and genetic factors play a major role in their development. The measures of choice for the diagnosis of allergic conditions is the history (personal or as reported by family members, for example). For in vivo testing in children, prick tests are employed. For in vitro diagnosis, determination of total IgE and specific IgE is used. If bronchial asthma is suspected, children older than 5 years, may be submitted to a diagnostic lung function test. Provocative testing is reserved for specific problems. PMID- 15658520 TI - [Strategies for the prevention and treatment of allergies in children and adolescents]. AB - Over and beyond the avoidance of allergies, prophylactic measures can also prevent the development of lower airway disease from upper airway disease, for example, bronchial asthma from allergic rhinitis. A further protective effect can be achieved by reducing both prenatal and postnatal allergens. A major concept of prevention--not merely of pulmonary function changes and the development of allergies--is the avoidance of exposure to passive smoking. With regard to pharmacotherapeutic agents, both official approval for their use by the respective age group, and the dearth of study-based data need to be taken into account. To ensure the successful application of inhalative agents, the use of appropriate inhalation aids as also ensuring the correct inhalation technique are urgently recommended, particularly in the case of children and adolescents. PMID- 15658521 TI - [Genetics and allergies: consequences for the practitioner?]. AB - Atopic disorders (asthma, hay fever, atopic dermatitis) may develop on the basis of a genetic predisposition towards environmental factors. These complex genetic conditions do not follow Mendelian inheritance patterns. Rather, the genetic predisposition for atopic diseases is based on a combination of changes affecting several genes which are involved in a variety of pathophysiological mechanisms. A number of candidate genes for asthma and allergies have already been identified. Today, it seems possible to identify genetic risk profiles related to specific environmental conditions which should lead to an improvement in diagnostic, preventive and therapeutic measures. However, before genetic diagnostic can be implemented in clinical practice, further studies are needed. PMID- 15658522 TI - [Evidence-based pharmacotherapy of neuropathic pain syndromes]. AB - Neuropathic pain can severely reduce quality of life. Double blind, placebo controlled studies confirm the efficacy of the treatment of painful neuropathy, postherpetic neuralgia and trigeminal neuralgia with tricyclic antidepressants, ion channel blockers, opioids and lipoic acid. The numbers-needed-to-treat (NNT) with monotherapy to achieve pain reduction of at least 50% are in the range of 2 to 4. Recent studies indicate that patients can benefit from combinations of opioids and tricyclic antidepressants or opioids and gabapentin. PMID- 15658523 TI - [Improvement in quality of life in the elderly. Results of a placebo-controlled study on the efficacy and tolerability of lecithin fluid in patients with impaired cognitive functions]. PMID- 15658524 TI - [New Chairman of the German Hypertension League: new goals, old problems, full engagement]. PMID- 15658525 TI - [Quality management brought family physician Rohde-Kampmann more free time and more money. How fulfilling obligations improved his practice]. PMID- 15658526 TI - [The 10-minute consultation. Incidental finding -- increased liver values]. PMID- 15658528 TI - [Bene diagnoscitur, dene curatur]. PMID- 15658529 TI - [Potentialities of non-invasive diagnosis of coronary artery anomalies by electron-beam tomography and multislice computed tomography]. AB - AIM: To assess diagnostic value of electron-beam computed tomography and multislice computed tomography (EBT and MSCT) for diagnosis of congenital anomalies of coronary arteries (CA). MATERIAL AND METHODS: Non-invasive coronary angiography with EBT and MSCT has been performed in 1162 patients. RESULTS: Different anomalies of coronary arteries have been found in 10 (0.9%) patients. Results of EBT and MSCT agreed with those of coronary angiography performed in 8 cases with CA anomalies. Though 720 of 1162 patients had coronary stenoses, no significant atherosclerotic lesions were found in anomalous CA. CONCLUSION: EBT and MSCT enable non-invasive screening and diagnosis of coronary arteries anomalies. PMID- 15658527 TI - [Look diagnosis. Pantyhose killer]. PMID- 15658530 TI - [Diagnosis of coronary atherosclerosis using echocardiography with bicycle ergometry exercise]. AB - AIM: To determine sensitivity and specificity of stress echocardiography (SEC) in diagnosis of stenosing atherosclerosis of coronary arteries (CA), especially in patients recovered from Q-myocardial infarction. MATERIAL AND METHODS: The above sensitivity and specificity were studied in 75 patients (70 males and 5 females; mean age 53.7 +/- 7 years) with coronary atherosclerosis (CS). RESULTS: The sensitivity of impaired local contractility index (ILCI) in univessel lesion was 77.8%, in multivessel lesion--90.4%, specificity--85.7%. Sensitivity of other parameters studied was for development of an anginal episode 65.4%, increment of ejection fraction under 5%--63.9%, increased end systolic volume--61.1%, depression of ST segment--48.1%, increased end diastolic volume 38.9%. A total of 3 parameters had 100% sensitivity: impossible a > 5% rise of EF in response to exercise, an increase of end systolic and diastolic volumes, anginal attack (85.7%), ST depression (78.6%). CONCLUSION: Hemodynamic parameters must be assessed in the course of SECG for objectivization of the result and improvement of its specificity. PMID- 15658531 TI - [Diagnosis of myocardial infarction complicated by complete block of the left branch of His bundle as shown by computed electrocardiotopography]. AB - AIM: To assess efficacy of computed electrocardiotopography (CECT) in diagnosis of myocardial infarction (MI) complicated by a complete block of His' bundle left branch (HBLB). MATERIAL AND METHODS: Twenty two patients with a complete block of HBLB have undergone CECT (hardware Rhythm-M, software "Rhythm" and "Heart"). RESULTS: Five patients had no scars. The rest 17 patients had MI. The diagnosis of MI was definite by ECG in 6 (35%) patients. In 11 (65%) patients MI diagnosis by ECG was doubtful. Only CECT detected typical for MI changes in the QRS changes. CONCLUSION: CECT can significantly detect scars in the myocardium in a complete block of HBLB and improves diagnostic potential of ECG. PMID- 15658532 TI - [Long-term orthostatic and bicycle ergometry exercise tests in differential diagnosis of syncopal conditions of unclear origin]. AB - AIM: To study efficiency of bicycle exercise test and long-term orthostatic test in differential diagnosis of syncopes. MATERIAL AND METHODS: The study included 113 patients suffering from recurrent syncopes of unclear origin. Twelve healthy volunteers were selected for assessment of the tests specificity. All the patients and controls were examined by means of long-term orthostatic test (LTOT) and bicycle exercise test (BET). RESULTS: Specificity of both tests was 100%. In LTOT syncopes were observed in 52 (46%) cases, including vasovagal syncopes (51 patients), epileptic seizure (1 patient). BET induced vasovagal syncopes in 54 (48%) patients. The results of both tests by inducing syncopes coincided in 30 (26%) cases. In 24 (21%) patients syncopes occurred only in BET while LTOT appeared uninformative. In 21 (18.6%) cases syncopes occurred only in LTOT while BET produced a negative result. 38 (34%) patients had syncopes neither in LTOT nor BET. Both tests induced syncopes in 75 (66%) patients. No complications were observed. CONCLUSION: LTOT and BET are safe and complementary methods in differential diagnosis of syncopes. PMID- 15658533 TI - [Clinical features and diagnosis of congenital stenosis of the pulmonary artery and the right ventricular outflow tract]. AB - AIM: To analyse clinical manifestations of congenital pulmonary artery stenosis (CPAS) and right ventricular outlet stenosis (RVOS), development of its diagnosis depending on anatomic variants of heart disease. MATERIAL AND METHODS: The data on 539 patients examined with conventional clinical and highly informative novel techniques were studied. RESULTS: The analysis of the CPAS and RVOS clinical picture depending on the anatomic variant of the heart disease has shown that clinical symptomatology was of the same type. The data of the tests have some specific features allowing identification of some forms of the heart disease. CONCLUSION: Diagnostic and prognostic significance of the findings was determined which gave rise to an optimal examination scheme with focus on wider use of highly informative techniques. PMID- 15658534 TI - [Leukocytosis as an indicator of risk of ischemic heart disease and its exacerbations (review)]. PMID- 15658535 TI - [Arterial hypertension in adolescents with different physical constitution including those with signs of metabolic syndrome]. AB - AIM: Detection of clinico-paraclinical features of arterial hypertension (AH) in adolescents with different physical constitution including signs of metabolic syndrome (MS). MATERIAL AND METHODS: Blood levels of uric acid, cholesterol, triglycerides (TG), high and low density lipoproteins were examined and glucose tolerance test was made in 108 adolescents with arterial hypertension (AH) and different physical constitution (normal, with overweight and obese). Waist and hip distribution of fat was assessed. The control group consisted of 108 healthy adolescents. RESULTS: Most of the hypertensive children had hyperuricemia (HU), some of the patients combined HU with changes in glycemic profile of blood. The above abnormalities were especially evident in patients with MS who were characterized by abdominal obesity, higher blood pressure, subnormal high density lipoproteins, higher triglycerides and total cholesterol in blood, more severe HU and defects in glycemic profile of blood. CONCLUSION: MS in adolescents may be of genetic origin as well as constitutional HU and insulin resistance may be mutually dependent signs combination of which with other factors presents the whole spectrum of clinical and paraclinical manifestations of MS. Detection of HU and insulin resistance with their correction in a preclinical disease stage (in children and adolescents) may represent a new line in prevention of AH and MS in adults. PMID- 15658537 TI - [Exacerbated chronic obstructive pulmonary disease: current trends in diagnosis and treatment (review)]. PMID- 15658536 TI - [Lisinopril effects on cerebral blood flow and blood rheology in hypertensive patients]. AB - AIM: To access the effect of lisinopril (diroton) on cerebral circulation and blood rheology in patients with arterial hypertension stage II. MATERIAL AND METHODS: The trial included 37 patients (16 males, 21 females) with a mean arterial hypertension (AH) history 15.9 +/- 5.6 years. Diroton was given in a dose 10-40 mg/day for 6 months. Cerebral circulation (total cerebral circulation and venous outflow--TCC and VOF) was accessed by means of doppler ultrasonography. Blood and plasm rheology was determined using a rotational viscozymeter ACP-2. Instrumental tests were performed at baseline and at the end of the study. RESULTS: Rheology tests showed that diroton-treated patients achieved a significant decrease in blood viscosity in high, moderate and low shear stress and plasma viscosity, a decrease in platelet aggregation index and an increase in the index of erythrocytic deformability. All these changes were accompanied with a significant fall in fibrinogen and hematocrit. Doppler ultrasound revealed an insignificant increase in TCC and VOF. CONCLUSION: Diroton significantly improved impaired blood rheology and viscosity in AH patients as well as cerebral hemodynamics in patients with subnormal cerebral circulation and venous outflow at baseline. PMID- 15658538 TI - [Levofloxacin: role and place in the treatment of infections of the lower respiratory tracts]. PMID- 15658540 TI - [Humoral and cellular adaptation mechanisms in development of intoxication syndrome in patients with acute viral hepatitis]. AB - AIM: To study endotoxin-binding function of blood in patients with acute viral hepatitis (AVH) in the course of the disease. MATERIAL AND METHODS: A total of 335 patients with AVH were examined. The titers of IgG antiendotoxin antibodies to glycolipid of Re-chemotype were estimated in enzyme immunoassay (Diaendotox-2 kit), endotoxin-binding activity (EBA) of high density lipoproteins (HDLP) in EIA with the kit Liprobind (Microecos). The number of polymorphonuclear leukocytes (PL) binding lipopolysaccharides were counted by M. Yu. Yakovlev et al. method. For detection of PL binding endotoxin of gram-negative bacteria, kits Endim (Microecos) were used, beta2-microglobulin was estimated by a radioimmunoassay using kits made in Belarus Institute of Bioorganic Chemistry. Registration of chemoluminescence was conducted on chemiluminometer LBK 1251 (Finland) by M. Wilson (1985) method. RESULTS: In moderate AVH at the height of intoxication a low Re-antibodies level was recorded irrespective of the disease etiology. To convalescence this level went up. PL reserves in AVH patients were low except patients with AVH C who had unaffected neutrophil reverves in relation to LPS binding. At the height of AVH, HDLP EBA was very high in all the groups with normalization to convalescence. In VHC high HDLP EBA persisted as long as the disease course. A significant rise of beta2-microglobulin was seen in all the groups of AVH patients, its severity correlating with the degree of the disease severity. CONCLUSION: One of the leading factors in development of indoxication in AVH patients is a lipopolysaccharide complex of gram-negative intestinal bacteria actively coming into blood flow due to hepatocyte cytolysis. In response to LPS action there appeared a stereotypic adaptation complex of reactions in the form of activation of cellular and humoral immune response. PMID- 15658539 TI - [How to diagnose functional (non-ulcerative) dyspepsia (review)]. PMID- 15658541 TI - [Invasive streptococcal (group A) infection: an outlook (review)]. PMID- 15658542 TI - [Combined antiretroviral therapy with nevirapine of HIV-infected adults]. PMID- 15658543 TI - [Antiphospholipid syndrome and exogenic risk factors in thromboses]. AB - AIM: To examine relationships between incidence rate of thromboses in antiphospholipid syndrome and exogenic risk factors (RF) of thrombosis. MATERIAL AND METHODS: The trial enrolled 131 patients (105 females and 26 males). They were divided into three groups: 23 patients with systemic lupus erythematosus (SLE, group 1), 63 patients with SLE and antiphospholipid syndrome (APS, group 2), 45 patients with primary APS (PAPS, group 3). Thrombosis RF questionnaire survey was made. Effects of corticosteroids and cyclophosphamide on occurrence of thrombosis were assessed. RESULTS: Such exogenic RF as intake of coffee, fat food, alcohol were not related to thromboses. Hypodynamia was more typical for APS patients (21.3%) than for SLE patients free of APS (8.7%). Overweight for was characteristic for APS patients (49 and 34.7%, respectively). The proportion of smokers was higher in APS patients, though smoking did not provoke thrombotic complications. A direct correlation was found between occlusion and corticosteroids administration, while occlusion and cyclophosphamide treatment correlated inversely. CONCLUSION: Venous thrombosis RF in patients with APS and PAPS are obesity and treatment with glucocorticosteroids. PMID- 15658544 TI - [Immunological features of rheumatoid arthritis in patients infected with viruses of hepatitis B, C and in patients with cryoglobulinemia]. AB - AIM: To specify immunological features of rheumatoid arthritis (RA) associated with hepatitis B, C viruses (HBV, HCV) and cryoglobulinemia (CGE). MATERIAL AND METHODS: Four groups of patients with verified RA were examined immunologically: infected with HCV, infected with HBV, with CGE diagnosed by the capillary method, free of HBV, HCV, CGE. CGE was estimated by the spectrophotometric method, functional activity of the complement components--by hemolytic micromethod. Polymerase chain reaction, identification of serological markers of viral hepatitides B and C were conducted with application of commercial kits produced in Russia. RESULTS: In RA patients infected with HBV and HCV, functional activity of the complement was significantly reduced. RA patients with hepatitis C and CGE showed hypoactivity of all the complement components. A classic course of RA was associated with elevated levels of C1 and C3 components in normal values of C1q, C2, C4 and C5 components. CGE was highest in RA+HCV and RA+CGE groups. CONCLUSION: The study of immunological features of RA associated with HCV, HBV and CGE is important not only in scientific but also in practical aspect as it indicates the necessity of treating the above conditions with drugs which do not suppress complement system significantly. PMID- 15658545 TI - [Polymorphism of clinical symptoms of connective tissue dysplasia syndrome]. PMID- 15658546 TI - [Connective tissue dysplasia and pregnancy (review)]. PMID- 15658547 TI - [Genital prolapse in women and articular hypermobility syndrome in connective tissue dysplasia]. AB - AIM: To examine relationships between genital prolapse and joint hypermobility (JHM), between GP severity and degree of JHM, to reveal causes of GP in JHM patients. MATERIAL AND METHODS: A total of 208 females with GP (mean age 38.9 years) entered the trial. They were divided into three groups by severity of connective tissue dysplasia (CTD): with mild CTD--16.3%, moderate CTD--35.6% and severe CTD--48.1%. The following methods of investigation were used: podometric Freedland's technique, ultrasonic investigation of the gall bladder, kidneys, echocardiography, morphological study of platelets, morphological and immunohistochemical study of the ligaments. RESULTS: In GP patients JHM occurred in 41.8% patients. Hypermobility of large joints reaches 38.9%, large--20.7%. Such associated manifestations of CTD as flatfoot, JHM, deformation of the spine, varicosity, predisposition to vegetovascular dysfunctions, mitral prolapse, arrhythmia, impaired conduction, refraction, gastroptosis, nephroptosis, hernias were observed in 69.2, 46.1, 38, 53.8, 49, 43.3, 38, 19.2, 12, 6.7, 14.4%, respectively. The morphological changes are explained by 40 +/- 15% type 1-3 collagen loss in the interstitial substance. Severe forms of GP in patients with JHM were observed in 84%. 52.4% females with JHM developed severe GP within 3 years after delivery of a child. Most of the examinees carried undifferentiated congenital dysplasias. CONCLUSION: JHM is a criterion of CTD diagnosis. PMID- 15658551 TI - [Hypomagnesemia in patients with chronic alcoholism in the course of alcohol withdrawal syndrome]. AB - AIM: To ascertain hypomagnesemia (HM) rate in patients with chronic alcoholism (CA) with alcohol withdrawal syndrome (AWS); correlation between AWS severity and HM rate. MATERIAL AND METHODS: Mg in plasm was measured at photometry in 129 CA patients treated in Kaunas Mental Hospital. RESULTS: Plasma Mg in CA patients with AWS was reduced (< 0.749 mmol/l), normal (0.750-1.250 mmol/l) or high (> 1.251 mmol/l) in 28.7, 65.1 and 6.2% patients, respectively. HM was diagnosed in 42.3% with a severe, 19.1% with a moderate and 20.0% with weak AWS. CONCLUSION: HM in AWS was registered in 28.7% patients. It occurred significantly more frequently (p < 0.05) in patients with a severe AWS. Pathogenetic mechanisms of HM in AWS are described. PMID- 15658548 TI - [Vitrum osteomag in prevention of osteoporosis in postmenopausal women: results of the comparative open multicenter trial]. AB - AIM: To investigate efficacy, tolerance and safety of the drug vitrum osteomag one tablet of which contains 600 mg calcium (1500 mg calcium carbonate), 200 IU of cholecalcepherol, 40 mg of magnesium, zinc (7.5 mg), copper (1 mg), manganese (1.8 mg) and boron (250 mcg) in women with osteopenia for prevention of osteoporosis. MATERIAL AND METHODS: A multicenter comparative open trial of vitrum osteomag influence on mineral bone density (MBD), change of pain syndrome in bones, index of calcium-phosphorous metabolism covered 334 postmenopausal women with osteopenia. MBD was measured in low-back spine and proximal part of the hip with DEXA method. All the patients were divided into 3 groups: 125 women taking 2 tablets of vitrum osteomag daily for 12 months (group 1); 111 women taking 1500 mg calcium carbonate (group 2); 96 women--control group (only observation). RESULTS: Vitrum osteomag relieved pain in the back and joints, had a positive effect on bone density (+1.5%) and proximal parts of the hip (0.6 0.93%) exceeding the effect of calcium carbonate only which preserves the initial MBD in low back spine but does not prevent bone loss in the hip. MBD dynamics in patients given vitrum osteomag differs essentially from one in the control group (from -1.9 to -2.91%) which demonstrates a reliable preventive anti-osteoporotic effect of this medication. The drug increases the level of general and ionized calcium in blood but does not cause hypercalcemia lowering the level of parathormone in blood. The rate of side effects in group 1 was 14.4% and did not differ much from that in group 2 (16.2%). CONCLUSION: The results of the study allow to recommend vitrum osteomag for prophylaxis of a rapid loss of bone tissue mineral density. PMID- 15658549 TI - [Structum therapy of patients with rheumatoid arthritis]. AB - AIM: To study efficacy of the chondroprotector chondroitin sulphate (structum, Pier Fabr Medicament Production, France) in patients with rheumatoid arthritis (RA) and secondary osteoarthrosis of the knee joints. MATERIAL AND METHODS: 15 women with a long history of RA (mean duration 11.9 years) entered an open non randomized trial of structum. The patients had a severe progressive highly active RA with a definite x-ray stage of the disease. 13 patients had a positive rheumatoid factor (1:80 to 1:1280) and involved knee joints which had been affected for 1 to 10 years (mean 5.3 years). The second x-ray stage was in 8 patients, the third stage of knee joints arthrosis was in 7 ones. A marked pain syndrome in the knee joints upon movement (mean 64.7 mm by VAS) was observed in all the examinees and at rest (mean 28 mm by VAS) in 13 of 15 patients. Structum was given according to a standard scheme: 500 mg 3 times a day for 3 weeks than 500 mg 2 times a day for up to 6 months. Basic drugs for RA were the same for all the observation period. RESULTS: Structum noticeably improved knee joint function (mean Leken's index 12.8, 11.3 and 9.4 scores before the treatment, on treatment month 3 and 6. Movement pain syndrome VAS reduced from 64.7 mm at the start to 51 mm 3 months and 37.5 mm 6 months later, rest VAS--from 19 to 10.3 and 6.4 mm, respectively. The demand in intraarticular glucocorticoids went down from 52 injections at the start of therapy to 6 after 6 months. Side effects for 6 months were absent. Overall efficacy was good (73.3% and 80%) as judged by the doctors and patients, respectively. After 6 months of therapy control x-rays found no progression of destructive changes in the knee joints (by MRI--in 4 patients). CONCLUSION: Structum has a marked positive therapeutic effect in patients with severe and long-term course of RA with associated pronounced secondary joint arthrosis. PMID- 15658552 TI - [Iatrogeny: role of E. M. Tareev and his school in the modern understanding of the problem]. PMID- 15658550 TI - [Characteristics of erythropoiesis regulation in population living at high altitude]. AB - AIM: To study characteristics of iron deficiency anemia (IDA) in native population of high altitude territories. MATERIAL AND METHODS: 1300 women living in Tien-Shan areas were examined for serum and erythrocytic ferritins (SF and EF), general iron-binding capacity of the serum, transferrin saturation with iron, transferrin, soluble transferrin receptor (TR), erythropoietin (EP), Hb, erythrocytes, erythrocytic indices. RESULTS: Regulation of erythropoiesis in women living at high altitude is specific, i.e. disagreement between complete depletion of iron (by SF and EF) and normal level of Hb, erythrocytes, EP and TR. CONCLUSION: It is suggested that in population living at high altitudes (3000 m above the sea level and higher) long-term adaptation to hypoxia gave rise to an original mechanism of erythropoiesis regulation when all coming iron participates in hemoglobin synthesis without iron deposition. Under hypoxic hypoxia regulation of erythropoiesis is directed to prevention of tissue hypoxia. PMID- 15658553 TI - [Olfaction and its receptors. History of a Nobel prize]. PMID- 15658554 TI - [Cochlear implant in postlingual adults with progressive hearing loss]. AB - OBJECTIVES: To analize the results of patients with cohlear implants in postlingual adults with progressive hearing loss and to analize various predictive factors. MATERIAL AND METHODS: Prospective study of 42 patients with profound and progressive hearing loss. We analized the outcomes in the first two years of follow-up. We examined the results in regards to the of age at time of implantation, other handicaps, cause of hearing loss, age of onset, cochlear ossification and actives electrodes. We use the logo-auditory protocol developed at the University Spanish of navarra. RESULTS: All tests presented a significative improvement in the first six months (p<0.01 Student t or Wilcoxon test). After six months the results were not statistically significant. The more important predictive factors were percentage of life with hearing loss, percentage of life with profound hearing loss, cochlear ossification and actives electrodes thus the significancy was more relevant at a follow-up period of 3 months (p<0.05 Student t or Mann-Withney test). CONCLUSIONS: These patients showed clear benefits in a short period of time and the patients with unfavourable circumstances had a more slow progression but a similar final outcome. PMID- 15658555 TI - [Anticochlear antibodies titles measured by Western-blot and hearing recovery level after corticoid therapy in patients with sudden deafness]. AB - OBJECTIVES: Detection of antichoclear antibodies in patients with sudden deafness (SD) ranges from five to forty one percent depending on authors. We tried to correlate the level of antibodies measured by Western-blot (WB) and hearing loss. PATIENTS AND METHODS: Forty nine subjects bearing of SD and a positive WB for bovine cochlear antigen. Hearing loss average was measured at the onset and after treatment with deflazacort 1 mg/kg per day by means of audiometry, and antibody titles were quantified in the diluted sera. RESULTS: Initial loss was 49.4+/ 20.6%, and after corticotherapy it was 15.9+/-18.0%. There was no statistically significative correlation titles-hearing loss (y=0.1122x+30.613; R2=0.7552) nor titles-recovery (y=-0.0818x+8.,65; R2=0.3229). CONCLUSIONS: Although predictive capability for treatment response is accepted for WB, quantifying of the antibody titles in patients with SD does not allow to make a prognosis about the average hearing recovery. PMID- 15658556 TI - [Malignant tumours of the external auditory canal and of the middle ear]. AB - OBJECTIVES: To review our experience and results in the treatment of a low incidence pathology such as the malignant ear tumors. METHODS: The study reviewed 36 patients with malignant tumors of the EAC and middle ear treated between 1977 and 2000 in our hospital, excluding cancer of the pinna, metastatic tumours and sarcomas. It was used the staging system proposed by the M.D. Anderson. RESULTS: The most common histological type in our series is the squamous cell carcinoma, and the otorrhea and pain are the primary symptoms in 100% of patients. Surgery combined with radiotherapy obtained a 41% 5-year survival rate. CONCLUSIONS: The staging system is an important prognostic factor and it is important an early diagnosis to achieve a better therapeutical result. PMID- 15658557 TI - [Combined nasal approach: maxilla-premaxilla approach and external approach]. AB - INTRODUCTION: A major aim of septo-rhinoplasty is the treatment of the internal and external nasal defects overall. We describe our technique with a combined external approach and maxilla-premaxilla approach in one surgical process. We present our functional and aesthetic results. METHODS: We analyse 16 patients that underwent septo-rhinoplasty using a combined approach; external approach and maxilla-premaxilla approach between 1999 and 2003. Anterior active rhinomanometry (RAA) and Anderson's scale to grade nose defects were performed before the operation and once again a year later. Furthermore, patients answered a visual analogical scale in order to know if they had noticed an improvement in nasal a year after the operation. RESULTS: RAA showed significant improvement in nose flow by 100 Pa pressure with a total flow mean of 730 cm3/s after surgery. After the operation, the majority of patients felt a definitive improvement in their nasal respiration. Scores on Anderson's scale of nose defects showed a significant improvement in nasal defects after surgery p<0.001. We achieved also a significant improvement for each pyramidal subunit. There were no complications in the healing of incisions neither patients complained of them. CONCLUSION: Nasal combined approach; external approach and maxilla-premaxilla approach in septo-rhinoplasty is a safe technique and allows functional and aesthetic improvement in nose defects, mainly in nasal base defects combined with septal defects. PMID- 15658558 TI - [Epidermal growth factor receptor gene amplification and E-cadherin expression in head and neck carcinomas]. AB - INTRODUCTION: The E-cadherin adhesion molecule is fundamentally involved in the maintenance of normal epithelial morphology and differentiation. The scattering and invasion of cancer cells induced by epidermal growth factor receptor (EGFR) activation has been suggested to be probably by affecting E-cadherin function. The aim of this study is to confirm whether EGFR amplification is related to E cadherin expression in head and neck squamous cell carcinomas. MATERIAL AND METHODS: Fifty patients with head and neck squamous cell carcinoma were studied. EGFR amplification was analyzed by semiquantitative PCR. E-cadherin expression was determined by immunohistochemistry. RESULTS: EGFR amplification was found in 9 cases (18%). E-cadherin expression was generally weaker in tumors than in adjacent normal epithelium. No relationship was found between EGFR amplification and E-cadherin expression. CONCLUSION: EGFR amplification did not affect the level of E-cadherin expression, suggesting a complementary role of both of these molecules in reduction of cellular adhesion in head and neck squamous cell carcinomas. PMID- 15658559 TI - [Angiomyoma of the retropharyngeal space]. AB - We report the case of a 65-year-old male patient with an angiomyoma of the retropharyngeal space. Angiomyoma is a tumour that arises from the smooth muscle in the wall of blood vessels. It is a relatively rare tumour of the head and neck, mostly in deep locations. This one is the second reported angiomyoma of the retropharyngeal space. Although image examinations and fine needle aspiration (FNA) did point to a vascular tumour, the diagnosis of this lesion is made by on excisional biopsy and histological staining with vimentin, desmin, actin or myosin. The treatment is excision of the tumour and recurrence is exceptional. PMID- 15658560 TI - [Patient with rhinorrea, destruction of nasal cartilage and hypochronic lesions of the skin]. AB - The leprosy is a chronic illness caused by M. leprae, which affects mucosa mainly at the level of the nasal region. The important destruction of the bone produces the facies leonina. We present the case of a patient with nasal leprosy, describing the clinical characteristics and the diagnostic and therapeutic methods that were carried out on her. PMID- 15658561 TI - Serosurvey of selected zoonotic agents in polar bears (Ursus maritimus). AB - Between 1982 and 1999 blood samples were collected from 500 polar bears (Ursus maritimus) captured in the Beaufort and Chukchi seas, to determine the seroprevalence of Brucella species, Toxoplasma gondii, and Trichinella species infections. The bears were classified into four age groups, cubs, yearlings, subadults and adults. Brucella and Toxoplasma antibodies were detected by agglutination (a buffered acidified card antigen and rapid automated presumptive test for brucellosis and a commercial latex agglutination test for toxoplasmosis); an ELISA was used to detect Trichinella antibodies. The overall seroprevalence of Brucella species was 5 per cent, and subadults and yearlings were 2-62 times (95 per cent confidence interval 1.02 to 6.82) more likely to be seropositive for Brucella species than adults and their cubs. The antibody prevalence for Toxoplasma gondii was 6 per cent, and for Trichinella species 55.6 per cent. The prevalence of antibodies to Trichinella species increased with age (P<0.001). PMID- 15658562 TI - Keratomycosis in six horses in the United Kingdom. AB - Six horses with keratomycosis were examined and three different clinical expressions of the disease were recognised. The diagnostic work-up and response to treatment is described. PMID- 15658563 TI - Effects of breed and feed supplementation on the fertility of cows developed for milk production in Zimbabwe. AB - Forty-six indigenous Sanga-type (Nkone and Tuli breeds) cows and 46 crossbred (Nkone x Jersey and Tuli x Jersey) cows were randomly allocated to four treatment combinations in a 2 x 2 factorial arrangement with two breeds and two dietary levels, a control diet and a diet supplemented with dairy meal containing 14 per cent crude protein at the rate of 2 kg per cow per day. The progesterone concentration was measured in milk samples taken three times a week from 10 days postpartum for up to 200 days, and the cows' bodyweights and body condition scores were recorded fortnightly. The pregnancy rate in the crossbred cows was significantly higher (P<0.05) than in the indigenous cows, and the assumed pregnancy loss rate 30 days after conception was significantly higher (P<0.05) in the indigenous cows than in the crossbred cows. The supplemented crossbred cows had a lower pregnancy loss rate than the supplemented indigenous cows (P<0.05). All the supplemented indigenous cows that lost pregnancies were in their first parity, whereas all the crossbred cows that lost pregnancies were multiparous and were not supplemented. The indigenous cows weighed significantly more (P<0.05) than the crossbred cows irrespective of diet, and the supplemented cows of both breeds weighed more (P<0.05) than the control cows. The supplemented indigenous cows had significantly higher (P<0.05) body condition scores than the control cows. The mean dairy milk yield of all the breeds was generally low but significantly higher (P<0.05) in the crossbred than in the indigenous cows. PMID- 15658564 TI - Evaluation of on-farm faecal worm egg counting in sheep. PMID- 15658565 TI - Subcutaneous granuloma associated with Macrophomina species infection in a cat. PMID- 15658567 TI - Withdrawal period for Ovitelmin S&C. PMID- 15658566 TI - Cystolithiasis secondary to intravesical foreign body in a horse. PMID- 15658568 TI - Equine viral arteritis in a stallion. PMID- 15658569 TI - Practice standards. PMID- 15658570 TI - Greyhound welfare. PMID- 15658571 TI - Sheep and goats. PMID- 15658572 TI - Renal rescue therapy in early stage of severe sepsis: a case study approach. AB - OBJECTIVE: The aim of this study was to evaluate the efficacy of noradrenaline and furosemide in combination for the treatment of impending acute renal failure in early stage of severe sepsis. DESIGN: observational case study. SETTING: Nine bed general ICU in university-affiliated cancer institute. PATIENTS: Severe septic cancer patients admitted to the ICU. MEASURMENTS AND RESULTS: 17 severe septic patients with multiorgan dysfunction syndrome (admission SOFA score, mean 9.1+/-3.0 p, and APACHE II score, mean 20.4+/-5.1 p.) received full intensive treatment including volume expansion, hemodynamic support with noradrenaline infusion and low-dose hydrocortisone. Severe sepsis was documented by proven infection, site of infection and high levels of procalcitonin (mean value 69.8 ng/ml, 7.1-588 ng/ml), C-reactive protein (mean 210 mg/l, range 49-370 mg/l) and low total cholesterol levels (mean 2.36 mmol/l, range 1.3-3.9 mmol/l). Acute renal injury and acute renal failure syndrome were detected in 14 patients (82 %) out of 17. The combination of noradrenaline continuous infusion (0.06-0.12 microg/kg/min) and furosemide infusion (10-30 mg/hr) was used for hemodynamic and renal support. We induced polyuria and reverse acute tubular necrosis to nonoliguric acute renal failure in 11 patients (78.5 %) from 14 septic cancer patients with acute renal injury/failure syndrome. We recorded 35.2 % hospital mortality due to the severe sepsis and septic shock. We used no renal replacement therapy. CONCLUSION: We consider renal rescue protocol as an effective method in the treatment for acute renal injury/failure syndrome in early phase of severe sepsis, when it is instituted very early with low/moderate dosage of noradrenaline and furosemide. (Tab. 6, Ref. 29.) PMID- 15658573 TI - Effects of terguride treatment on glucose abnormalities induced by ischemic brain damage in SHR/N-cp lean Koletsky strain and in rats of Wistar strain. AB - BACKGROUND: Severe head injury is associated with a stress response that includes hyperglycemia, which has been shown in both experimental and clinical studies to exacerbate the severity of brain injury during ischemic conditions. OBJECTIVES: This study evaluated the possible protective effect of Terguride (trans dihydrolisuride) on glucose metabolism against hyperglycemia. METHODS: The experiments were performed in male and female rats of Wistar and Koletsky strain. Glucose intolerance was induced in both strains by 4-hour-occlusion of both common carotid arteries followed by 44-hour reperfusion. RESULTS: Brain ischemia induced glucose intolerance in both rat strains. Basal glycemia was significantly increased by the brain ischemia in male and female Wistar rats, but not in Koletsky rats. The analysis of the effect of Terguride treatment of glucose abnormalities on the "area under the glucose tolerance curve" (AUC) has shown significant decrease of AUC in both sexes of Wistar strain and in females of Koletsky strain. Basal glycemia was significantly decreased only in males of Wistar strain. CONCLUSION: Terguride (trans-dihydrolisuride) decreases hyperglycemia in rats with ischemic brain damage. (Fig. 4, Ref. 29.) PMID- 15658574 TI - Human eosinophils as potent inflammatory cells and their apoptosis. AB - A significant association was established between the eosinophil and a number of disease conditions, including helminthiasis, allergy, asthma, drug hypersensitivity, certain neoplasias, and graft rejection. Activation of eosinophils and release of proinflammatory lipid mediators, cytokines, free oxygen radicals, highly-charged cationic proteins contribute to the onset and maintenance of tissue inflammation. Eosinophil accumulation in blood and tissues has been related to a defect in their apoptotic death. Decisive events during the apoptotic process involve mitochondrial permeabilization and caspase activation. Clearance of apoptotic cells depends on the ability of phagocytes to recognize their cell targets and, subsequently, to engulf them. (Tab. 2, Ref. 32.) PMID- 15658575 TI - Immunohistochemical detection of MRP1 protein in normal and hyperplastic human thyroid gland. AB - In this study we describe the localization of MRP1 in normal and hyperplastic thyroid tissues. To demostrate this protein we have used the enzyme imunohistochemical method with monoclonal antibody MRPm6. We have found MRP1 to be present in follicular epithelial cells of normal thyroid tissue and higher expression in the same cells of hyperplastic thyroid tissue. The brown colour of the diaminobenzidine reaction product allowed us to define the distribution of stain clearly. (Fig. 4, Ref. 16.) PMID- 15658576 TI - Operative treatment of cervical premalignant lesions and the presence of high risk human papilloma virus as etiologic agent. AB - The conization as an excisional method is used as an operative treatment of high risk premalignant cervical lesions. Early detection and removal may stop the developing process that leads to invasive carcinoma. The study presents a group of 100 women, operated with "cold knife" conization during the year 2002 at the 1st Operative Oncologic Department of the Gynecology and Obstetrician Clinic - Medical Faculty, Skopje. The operative material showed CIN 2 in 2 and CIN 3 in 31 women. In 7 of them, CIN 3 lesions were removed by punch biopsy or ECC, thus the cone showed only CIN 2. In two women out of the total of 67 with the preoperative diagnosis of CIS, CIN 3 lesions were postoperatively diagnosed. This is the result of the adequate punch biopsy during the diagnostic procedure. Three cones showed Ca microinvasivum and other 3 displayed Ca invasivum which then needed other surgical treatment. In 93 % of the women the conization had a therapeutic effect and in 7 % it was only of diagnostic value. HPV testing was made in 55 out of the operatively treated women. The most common type HPV 16 was identified in 27 % of the women. The second was HPV 31 in 25.5 %, then HPV 66 in 14.5 %, HPV 18 in 11%, HPV CP 8304 and HPV 38 in 5.5 %, HPV 58 as well as MM9 in 2 (3.6 %). The dichotomic presence of HPV 31+18 and HPV 6+16 was identified in two women. (Tab. 3, Ref: 10.) PMID- 15658577 TI - Close association between parathyroid hormone and left ventricular function and structure in end-stage renal failure patients under maintenance hemodialysis. AB - OBJECTIVES: Cardiovascular risk factors are a significant burden in end-stage renal disease patients under hemodialysis and cardiovascular-related diseases are the leading cause of death among these patients and are responsible for almost half of all deaths in dialysis patients. In this study we aimed to consider the role of excess PTH in the development of left ventricilar hypertrophy (LVH) and LV ejection fraction in patients with end-stage renal disease under regular hemodialysis. PATIENTS AND METHODS: This study is cross-sectional, and was done in patients with end-stage renal disease (ESRD) undergoing maintenance hemodialysis treatment. Calcium, Phosphorus, Alkalin phosphatase and Intact PTH (iPTH) were measured. Hypertensive patients were stratified into three stages. The total of 73 patients (F=28, M=45) consisted of 58 non-diabetic hemodialysis patients (F=22, M=36) and 15 diabetic hemodialysis patients (F=6, M=9). RESULTS: Significant inverse correlation of serum ALP with percent age of LV ejection fraction, marginal correlation of serum ALP with LVH and marginal correlation of serum iPTH with LVH were observed. Also significant inverse correlation of serum iPTH with percent age of LV ejection fraction in non diabetic HD patients was observed. CONCLUSIONS: Adverse effects of secondary hyperparathyroidism on LV function and structure in this study show the role of excess PTH in the development of left ventricilar (LV) hypertrophy as well as low LV ejection fraction in patients with end-stage renal disease under hemodialysis which needs more attention to control of secondary hyperparathyroidism to reduce the risk of cardiovascular morbidity and mortality in dialysis patients. (Tab. 5, Fig. 3, Ref: 29.) PMID- 15658579 TI - Velopharyngeal insufficiency and its associated pathologies in cleft patients. AB - OBJECTIVE: A daily contact with cleft patients affected by velopharyngeal insufficiency (VPI) of various types and origins has stimulated us to analyze the associated disorders participating in the achieved functional results. In our experience, an early palate closure (in the first year of life) reduces the incidence of VPI and therefore the need of pharyngoplasty. (Tab. 6, Fig. 1, Ref. 12.) PMID- 15658578 TI - Cost-effectiveness analysis of switching from intravenous to oral administration of antibiotics in elderly patients. AB - OBJECTIVE: The aim of the introduced work was to evaluate pharmacoeconomic advantages of timely switching from intravenous to oral administration of antibiotics (ATB). METHODS: The evaluated group was selected out of 2870 patients, who were hospitalised at the Clinic of Geriatric Medicine of the Faculty of Medicine of Comenius University in Bratislava from January 1st 1999 to December 31st 2001. In our retrospective study we analysed 96 patients with community-acquired pneumonia successfully treated by ATB. In 43 of them ATB were given intravenously and in 53 the therapy was switched, i.e. the intravenous administration was used at the beginning and oral administration when the condition improved. We applied a cost-effectiveness analysis to the pharmacoeconomic evaluation. The cost-effectiveness coefficient was calculated as the ratio of ATB price (Slovak Crowns) to the effectiveness criterion (number of asymptomatic days in month). RESULTS: According to the cost-effectiveness coefficient, the switch therapy was significantly less expensive in all evaluated ATB (except for pefloxacin) in comparison with intravenous administration: ampicillin-sulbactam 93.9 vs 168.1; cefuroxime 90.0 vs 123.3; amoxicillin clavulanate 74.0 vs 116.3; ciprofloxacin 31.7 vs 54.1. CONCLUSIONS: A timely switching from intravenous to oral administration of ATB in a suitable patient is an effective way to save financial resources. (Tab. 5, Ref: 21.) PMID- 15658580 TI - Intraabdominal pressure after rectal resections. AB - The mean correlation curves of the intraabdominal pressure over time were evaluated by regression analysis (value 0.05) in both groups. The curves had a rising character in both groups, but in the control group the curve rose more steeply and reached a higher level than the curve of studied group. The onset of peristalsis, first flatus and stool took a shorter time in the studied group. (Tab. 1, Fig. 5, Ref. 5.) PMID- 15658581 TI - 20-year experience in operations for subclavian steal syndrome. AB - The aim of this article is to summarize our experience in operations for subclavian steal syndrome. We recommend anaesthesia with preservation of patient's consciousness and mobility, and we prefer transposition of arteries to prosthesis implantation. The main characteristic of this approaches is an attempt to increase safety of operation. A feasible procedure is suggested also in the case of reconstruction occlusion: the axillo-axillary bypass. (Fig. 6, Ref. 3.) PMID- 15658582 TI - Iatrogenic perforation during an endoscopic examination of the gastrointestinal tract. AB - OBJECTIVE: The study was aimed to highlighting the situations leading to increased risk of iatrogenic perforation during an endoscopic examination of the GIT. The optimization of surgical care procedure following intestinal perforation was suggested as well. METHODS: We analysed 3897 colonoscopic examinations performed during the past 5 years. We have found 6 cases (0.15 %) of iatrogenic colon perforation. RESULTS: All six cases of iatrogenic GIT perforations were followed by surgical revision. A suture was done twice, stomy three times, resection once, and restomisation also once. CONCLUSIONS: Polypectomy of thick polyps with a wider base and more rigid consistency is dangerous. The longer is the time and stronger the coagulation, the higher is the chance of iatrogenic perforation. Therefore we recommend laparoscopically assisted procedure. (Ref. 10.) PMID- 15658584 TI - Medical information portal. AB - The classic function of the library was to build and maintain a knowledge base and to provide an access to that collective memory for the purposes of learning, teaching, caring for patients, research work or managing an organization. The formats and presentation of that knowledge base are changing rapidly, as the methods and techniques for accessing information. (Fig. 1, Ref. 1.) PMID- 15658583 TI - Oncologic markers in patients with colorectal cancer after a complex therapy. AB - The authors present a prospective study on a value of oncologic markers CEA and CA 19-9 in patients after curative therapy for colorectal adenocarcinoma. During a five-year follow-up in 320 patients, a significant elevation of CEA or CA 19-9 was documented in 71 patients (22.8 %), and resulted in tumour detection in 39/71 patients (55 %). Although the levels were defined as false positive in 32 patients (45 %), the importance of CEA and CA 19-9 monitoring is documented by elevated levels of oncomakers in 39/55 patients (71%) with metastases or local regional recurrence of colorectal adenocarcinoma. (Tab. 3, Ref: 21.) PMID- 15658585 TI - [Happy end...and a Happy New Year. The centenarian, Lakartidningen, is doing well -in spite of tough times]. PMID- 15658586 TI - [Drug-eluting stents in coronary arteries. The risk of restenosis is drastically reduced]. PMID- 15658588 TI - [Episodes of severe hypoglycemia--increasing problem in type 1 diabetes]. AB - A cross-sectional survey of severe hypoglycaemia was performed in type 1 diabetes mellitus patients in 1984 and repeated in 1998 at the diabetes out-patient clinic of a Swedish university hospital. The study revealed that the prevalence of severe hypoglycaemia had increased by more than 50 per cent over 14 years, in spite of more frequent use of multiple insulin injection therapy and daily self monitoring of blood glucose. A multiple logistic regression analysis of risk factors for severe hypoglycaemia explained less than 10% of the variance, implicating only unawareness of hypoglycaemia and HbA1c. It is concluded that long duration type 1 diabetes mellitus patients are increasingly vulnerable with respect to severe hypoglycaemia and that this should be taken into account when individual treatment goals are being proposed to patients. Novel short-acting and long-acting analogs of insulin as well as insulin pumps may prove useful to minimize the risk of severe hypoglycaemic episodes. It is argued that the ability of currently marketed glucose monitoring systems to sensitively and specifically detect hypoglycaemia is limited. PMID- 15658589 TI - [A patient from Turkana]. PMID- 15658587 TI - [Influenza vaccination for all older than 65 years--utopia or possibility]. PMID- 15658590 TI - [2004--a jubilee year for organ transplantation. Fifty years of development from a questioned activity to an established care routine]. PMID- 15658591 TI - [...among evasive syndromes and conditions. A crowd turbulent with pain, fatigue, anxiety, headache, vertigo, stomach, pain...]. PMID- 15658592 TI - [From starvation cures to pancreas transplantations. Treatment of diabetes seen in a perspective of 100 years]. PMID- 15658593 TI - [The still frustrating mysterious gallstone. A century of gallstone disease in Lakartidningen]. PMID- 15658594 TI - [Prevention goes astray. Stop subsidizing of drugs for primary prevention when other alternatives are available]. PMID- 15658595 TI - [Is reduce mental capacity cheating?]. PMID- 15658596 TI - [How strict should editors be? Nothing is done for the sake of errors]. PMID- 15658597 TI - [SBU's conclusions concerning treatment of depression are not well-founded]. PMID- 15658598 TI - [The National Social Insurance Board should change sickness certificates instructions]. PMID- 15658599 TI - [Reply from the SBU about salt and high blood pressure: More controlled long-term trials are required]. PMID- 15658600 TI - Effects of palladacycle complex on hematopoietic progenitor cells proliferation in vivo and in vitro and its relation with the inhibitory properties of this compound on the angiotensin-I converting enzyme activity. AB - In the present study, we introduce a new class of organometallic compound, the Biphosphinic Palladacycle Complex [Pd (C2, N-S(-)(dmpa)(dppf)] Cl (BPC), as an angiotensin-I converting-enzyme inhibitor (ACEI) with hematological regulation properties. When BPC was assayed as a competitive inhibitor over the hydrolysis of Abz-YRK (Dnp)-P-OH (Km = 7.0 microM), it showed a Kiapp = 0.2259 ng and a Ki value of 94.12 pg. Using murine long-term bone marrow cultures (LTBMCs) and clonal culture techniques, we also evaluated the capacity of this drug (1.18 microM) to module haematopoietic progenitor cells proliferation in vitro and in vivo. Our results demonstrated that BPC produces no toxicity to bone marrow cells, as determined by the unchanged cell number in the non-adherent layer at weeks 1, 2, and 8 and the increased number of adherent cells present in the BPC treated LTBMCs. However, the proportion of CFU-Cs in the non-adherent cell layer was reduced at weeks 5, 6, 8, and 9. In vivo studies using the dose of 1 mg/kg of BPC, administered by subcutaneous route, presented similar result as those found in vitro, in the number of CFU-Cs. This latter finding may be explained by the inhibitory effects of this drug on the ACE activity, which probably result in increased levels of its substrate AcSDKP, a negative regulator of hematopoiesis. PMID- 15658601 TI - Stabilization of red blood cell membranes by thalidomide in vitro. AB - The anti-inflammatory effect of thalidomide has been well established. The mechanism of this anti-inflammatory action is still not completely understood. Certain drugs exert their anti-inflammatory action by stabilizing the membranes of polymorphonuclear neutrophils (PMN) thereby reducing the production of reactive oxygen intermediates. We evaluated the effect of thalidomide on cell membranes by using red blood cells (RBC), PMN and the monocyte-like cell line THP 1. Osmotic fragility of RBC showed that in vitro, thalidomide stabilized the membrane of RBC from plasma free blood; whereas, it did not affect RBCs from whole blood. Red blood cells taken from subjects before and after ingestion of thalidomide were not affected after exposure to different concentrations of hypotonic NaCl solution. Thalidomide did not affect the membrane stability of PMNs as well as THP-1 in a significant manner. These data suggest that the anti inflammatory mechanism of thalidomide is not related to events associated with the oxidative burst of PMNs or monocytes. PMID- 15658602 TI - Ehrlich ascites tumor as a tool in the development of compounds with immunomodulatory properties. AB - In previous works, we have demonstrated that the myeloprotective properties of several natural and synthetic compounds are partly responsible for their antitumor activity in the Ehrlich ascites tumor (EAT) model. In this work, we present information that may be useful to the study of pharmacological and toxicological properties of compounds that affect the hematological compartment. Clonogenic studies in EAT-inoculated mice demonstrated a rapid decrease in bone marrow CFU-GM, whereas a progressive increase in splenic CFU-GM and cellularity was observed, followed by splenomegaly. Bone marrow cellularity declined on the third day after tumor challenge, returning to normal values thereafter. Serum from EAT-bearing mice produced detectable colony-stimulating activity in vitro. Similar results were observed with the conditioned medium from Ehrlich tumor cell cultures, but not with the cell-free Ehrlich tumor ascitic fluid. Tumor inoculation also resulted in a more striking depletion in the number of non adherent cells in long-term bone marrow cell cultures (LTBMCs) with no bone marrow stroma formation. We speculate that the physiological alterations induced by the EAT growth can be used to assess the ability of compounds to modulate the hematopoietic response. PMID- 15658603 TI - Induction of coronary arteritis with administration of CAWS (Candida albicans water-soluble fraction) depending on mouse strains. AB - The intraperitoneal administration of CAWS (water-soluble extracellular polysaccharide fraction obtained from the culture supernatant of Candida albicans) to mice induces coronaritis similar to Kawasaki disease. We analyzed differences in the production of cytokines involved in the occurrence of coronary arteritis among mouse strains, C3H/HeN, C57BL/6, DBA/2 and CBA/J that were injected with CAWS at 4 mg/mouse for 5 consecutive days in the first week and the fifth week of administration. The incidence of arteritis was 100% in C57BL/6, C3H/HeN and DBA/2 mice, but only 10% in CBA/J mice. The coronary arteritis observed in DBA/2 mice was the most serious, with several mice expiring during the observation period. The CAWS-sensitive strains revealed increased levels of IL-6 and IFN-gamma during the course of a specific response to CAWS by spleen cells. In contrast, IL-10 levels were observed to increase markedly in CAWS resistant CBA/J mice, but not the CAWS-sensitive strains. However, TNF-alpha levels were more elevated only in DBA/2 mice. The difference in disease development and cytokine production strongly suggests that the genetic background of the immune response to CAWS contributes to the occurrence of coronary arteritis. PMID- 15658605 TI - Human hemoglobin shares bioactivities ascribed to human tumor necrosis factor alpha. AB - Hemoglobin mediated cytotoxicity and apoptosis has been evaluated in Tumor necrosis factor-alpha (TNF-alpha) sensitive cell line, U937 and compared with TNF alpha. Both species of hemoglobin, Hemoglobin A2 and Hemoglobin A0 induced apoptosis and cytotoxicity in U937 cell as measured by flow cytometry and 3-(4,5 dihydro-6-(4-(3,4-dimethoxybenzooyl)-1-piperazinyl)-2(1H)-quinoline (MTT) assay respectively. Different concentration of Hemoglobin A0 (4 ng/mL to 4000 ng/mL) induced apoptosis ranging from 9% to 16% in U937 cells. 4000 ng/mL hemoglobin A0 showed maximal apoptotic cells. TNF-alpha showed 87% apoptotic U937 cells at concentration of 1 pg/mL. HbA0 displayed cytotoxicity in U937 cell line at higher concentration in comparison to TNF-alpha. 4000 ng/mL of hemoglobin A0 showed optimal cytotoxic response in U937 cells. A dose response curve was also observed with varying doses of hemoglobin A0. U937 cells pretreated with serum activated LPS for 1 hr and incubated with different concentration of hemoglobin or human TNF-alpha for 24 h reduced the cytotoxic effect on U937. Dexamethasone treatment of U937 cells helped in protecting the HbA0 and HbA2 mediated cytotoxicity and anti-TNF-alpha antibody neutralized the hemoglobin mediated apoptosis and cytotoxicity. It is therefore apparent that human hemoglobin shares some of the bioactivities previously ascribed to TNF-alpha. Sharing of bioactivities of TNF alpha by hemoglobin is interesting and suggests that cell free hemoglobin can mimic TNF-alpha functionally. PMID- 15658606 TI - Maturation of fish erythrocytes coincides with changes in their morphology, enhanced ability to interact with Candida albicans and release of cytokine-like factors active upon autologous macrophages. AB - Erythrocytes from the rainbow trout Salmo gairdneri Richardson (Salmo g.R.) were classified into immature and mature populations, respectively, by measuring longitudinal diameters. More elongated fish erythrocytes (FE), classified as mature cells, were those interacting with Candida albicans (CA) in a higher frequency in terms of either binding to the fungus or its intracellular engulfment. At the same time, in the rosetting phenomenon more elongated mature FE surrounded macrophages (Mo) phagocytosing CA. Finally, FE activated by CA released in the supernatants cytokine-like factors able to modulate Mo functions. In particular, these active supernatants were analyzed for their capacity to inhibit Mo migration Macrophage Inhibition Factor (MIF) activity and enhance Mo phagocytosis. Both activities were detected in supernatants from CA stimulated FE but not in control supernatants. MIF activity could play a role in the accumulation of Mo in the context of functional rosettes, while the factor enhancing Mo phagocytosis could promote clearance of CA in a more efficacious way. PMID- 15658604 TI - Suppression of nuclear translocation of nuclear factor-kappaB and nuclear factor of activated T cells by Younggaechulgam-tang. AB - Younggaechulgam-tang (YGCGT) is known to suppress inflammatory and autoimmune responses, and it has clinically been used among Oriental medical doctors in South Korea. We investigated YGCGT-mediated changes in downstream T cell signal transduction. The expression levels of nuclear factor-kappaB (NFkappaB) subunit RelA and nuclear factor of activated T cells (NFATc1) in cytoplasm and nucleus were examined by western blot analysis. Interlukin-2 (IL-2) expression in MOLT-4 cells activated by phytohemagglutinin (PHA) was determined by reverse transcription-polymerase chain reaction (RT-PCR) and Western blot analysis. IL-2 secretion was measured by an enzyme-linked immunosorbent assay (ELISA). PHA induced translocation of Rel A and NFATcl to the nucleus were markedly reduced by YGCGT treatment. Furthermore, IL-2 mRNA and protein levels and IL-2 secretion were significantly diminished by YGCGT treatment. In conclusion, YGCGT treatment of T cells inhibits selectively nuclear translocation of RelA and NFATc1, resulting in diminished production of IL-2. These results suggest that YGCGT may have potential as immunosuppressive drugs with improved efficacy and reduced side effects. PMID- 15658607 TI - Inhibitory effect of inflammatory cytokines production from activated mast cells by Gamisopoonghwanghyul-tang. AB - Rheumatoid arthritis (RA) is a chronic inflammatory disease and its exact cause and pathophysiological process remain unclear. Because the mast cell contains potent mediators, including multifunctional cytokines, its potential contributions to the processes of inflammation and matrix degradation have recently become evident. Gamisopoonghwanghyul-tang (GSPHHT) has been used as a traditional Korean medicine for the treatment of RA. In this study, we investigated the effect of Gamisopoonghwanghyul-tang (GSPHHT) on the production of inflammatory cytokines by activated human mast cell line HMC-1 cells. When GSPHHT (1 mg/ mL) was added, the production of tumor necrosis factor (TNF)-alpha, interleukin (IL)-1beta, IL-6, IL-10 was inhibited by 36.3%, 36.3%, 30.8%, 48.7% respectively in phorbol 12-myristate 13-acetate plus calcium ionophore A23187 stimulated HMC-1 cells. However, the production of IL-4 was significantly increased at 0.01 mg/mL. GSPHHT had no effect on TNF-alpha mRNA expression. These results suggest that GSPHHT regulates production of inflammatory cytokines from activated mast cells. PMID- 15658608 TI - Anti-cancer and pro-apoptotic effects of an herbal medicine and Saccharomyces cerevisiae product (CKBM) on human hepatocellular carcinoma HepG2 cells in vitro and in vivo. AB - Hepatocellular carcinoma is a major health problem worldwide. Different treatment strategies have been developed to cope with this problem. Herbal medicine is now widely studied in both Eastern and Western countries. In this study, we used both in vitro and in vivo model to illustrate the anti-tumor effect of a product, CKBM, consisting of herbal medicine and specially processed Saccharomyces cerevisiae. Dose-dependent anti-proliferation effect was observed on in vitro growth of human hepatoma HepG2 cells after 48 hours incubation with CKBM. At the 50% inhibitory concentration (IC50) no significant toxic effect was observed on normal human fibroblasts Hs68 and human liver WRL-68 cells. The results of morphological changes, detection of DNA fragmentation, flow cytometric analysis and Western blot analysis indicated that this anti-tumor effect of CKBM was mediated via the process of apoptosis. In addition, HepG2 cells- bearing nude mice model was used for in vivo anti-tumor study. Our results showed that 14-day treatment with 0.8 ml daily dosage of CKBM could inhibit 54.1% of tumor growth. The plasma activities of enzymes specific for heart and liver, namely creatine kinase, lactate dehydrogenase, aspartate transaminase and alanine transaminase, remained at normal levels, indicated that CKBM did not produce toxicity to the host. PMID- 15658609 TI - M-2000, as a new anti-inflammatory molecule in treatment of experimental nephrosis. AB - The therapeutic effect of M-2000 (C6H10O7) molecule was tested in Adriamycin induced nephropathy. To induce experimental nephrosis, Adriamycin was given once by a single intravenous injection (7.5 mg/kg) through the tail vein. Six days after injection of Adriamycin, therapeutic protocol was developed by intraperitoneally (i.p) administration of 30 mg/kg M-2000 solution. Total of i.p. injections were 14, in which five injections were made every day and nine injections were carried out at regular 48-h intervals. Therapeutic protocol was terminated on day 28 and animals were killed on day 43. The treated patient rats showed a significant reduction in proteinuria, BUN, serum creatinine and serum cholesterol, as well as, administration of M-2000 could significantly diminish the serum level of interleukin-6 (IL-6) in treated animals compared to non treated controls. Moreover, treatment with M-2000 significantly reduced number of glomerular leukocytes, Hypercellularity and hydropic change in capillary network within the renal cortex and decreased tubular casts. These data suggest that M 2000 therapy can ameliorate proteinuria, and suppress the progression of glomerular lesions in experimental model of nephrosis. PMID- 15658610 TI - Stachys riederi inhibits mast cell-mediated acute and chronic allergic reactions. AB - The effect of aqueous extract of Stachys riederi var. japonica Miq. (Labiatae) (SRAE) on the mast cell-mediated allergic and inflammatory reactions were investigated. SRAE inhibited systemic allergic reaction induced by compound 48/80. SRAE dose-dependently inhibited passive cutaneous anaphylaxis (PCA) activated by anti-dinitrophenyl (DNP) IgE. SRAE also dose-dependently inhibited the histamine release from rat peritoneal mast cells (RPMC) activated by compound 48/80 or anti-DNP IgE. Moreover, SRAE inhibited the secretion of tumor necrosis factor-alpha (TNF-alpha) and interleukin (IL)-6 in phorbol 12-myristate 13 acetate (PMA) plus calcium ionophore A23187-stimulated human mast cells (HMC-1 cells). These results provide evidences that SRAE may be beneficial in the treatment of acute and chronic allergic diseases. PMID- 15658611 TI - In vitro immunomodulatory activity of Bo-yang-hwan-o-tang. AB - Bo-yang-hwan-o-tang (BHT), an herbal decoction has been mainly used for improvement of blood flow in oriental medicine. Its in vivo immunomodulation was recently demonstrated but the effective mechanisms have not been described. This study was carried out to evaluate in vitro immunomodulatory activity of BHT. Water extract of BHT significantly promoted in vitro proliferative responses of mouse spleen cells (SPC) and also further enhanced the proliferation of SPC stimulated with anti-CD3 antibody. Unexpectedly, addition of BHT extract did not affect proliferation of both resting and CD3-activated T cells, whereas it showed a strong mitogenic activity on B cells. Flow cytometric analysis of CFSE-stained SPC showed that BHT-mediated enhancement of CD3-activated SPC proliferation is due to T cell, but not B cell, division. Mixed culture experiment combining T and mitomycin C-treated B cells demonstrated that BHT-mediated enhancement of CD3 activated T cell proliferation was dependent on the presence of B cells. However, B cell-derived factors were not involved in BHT effect on T cell proliferation. In the presence of B cells, BHT treatment resulted in a great enhancement in IL-2 production of CD3-activated T cells, and BHT effect on T cell proliferation was completely abrogated by addition of exogenous IL-2, indicating that IL-2 plays a critical role in BHT-mediated enhancement of CD3-activated T cell proliferation. Taken together, our data revealed that BHT possesses a potent B cell mitogenic activity and also can enhance activated T cell response through B cell regulation. PMID- 15658612 TI - Mechanisms responsible for delayed and immediate hemolytic transfusion reactions in a patient with anti-E + Jk(b)+ Di(b) and anti-HLA alloantibodies. AB - Immediate hemolytic transfusion reactions (IHTR) occurred in the course of delayed hemolytic transfusion reactions (DHTR). An 84-year-old man had received a blood transfusion 20 years ago. Progressive anemia developed, because of continuous bleeding from a bladder tumor. He was transfused with concentrated red blood cells (CRC) which were Rh-E antigen negative, because he had anti-E antibodies (day 0). He received CRC on day 3, and underwent resection of bladder tumor on day 6. Although crossmatch-compatible CRCs were prepared for the operation, those were not required and were kept in a refrigerator in the ward. On day 9, when a CRC kept in the ward was transfused, he suddenly had a IHTR. In order to analyze a mechanism of IHTR, the anti-Jk(b) and anti-Di(b) antibodies, anti-HLA antibodies and the concentrations of inflammatory cytokines were measured in serum samples. The anti-Jk(b) and anti-Di(b) antibodies increased prior to IHTR experienced on day 9. The concentrations of IL-6 and IL-1beta increased from day 2, while the concentration of IL-8 increased from day 7. The anti-HLA class I antibody could be detected 2 days before IHTR. Thus, the anti Jk(b) and anti-Di(b) antibodies induced the production of inflammatory cytokines and symptoms of DHTR and IHTR. The anti-HLA class I antibody could be produced in spite of using the filer for removing leukocytes, and may take part in the induction of IHTR. Further, blood products should be transfused soon after completing a crossmatch test in patients with anti-RBC alloantibodies. PMID- 15658614 TI - Age at diagnosis, body mass index and physical morbidity in children and adults with the Prader-Willi syndrome. AB - Age at diagnosis, Body Mass Index and physical morbidity in children and adults with the Prader-Willi syndrome: The medical findings of a population of 54 Prader Willi patients with a molecular confirmed diagnosis are discussed. In the age group aged 18 or younger, a reasonably good control of weight as measured by Body Mass Index (BMI) is found. This is probably due to the fact that diagnosis was made at an early age and intensive diet management was started early. Despite their relatively low BMI, these children remain at high risk for developing scoliosis requiring active treatment (28% of the children). Adults (older than 18) diagnosed at the age of 10 or later have a high risk for developing obesity and obesity related health problems such as hypertension (38%), non-insulin dependant diabetes mellitus (11%) and cardio respiratory failure (16%). PMID- 15658613 TI - Interleukin-1 beta gene polymorphism related with allergic pathogenesis in Iris constitution. AB - Iridological constitution has a strong familial aggregation and is implicated in heredity. The aetiology of inflammatory bowel disease is still unknown. However, from genetic epidemiological studies there is considerable evidence that genetic factors are associated with both Crohn's disease and ulcerative colitis. We investigated the relationships between Iridological constitution and interleukin 1 beta (IL-1beta) gene polymorphism. IL-1beta is a major proinflammatiry cytokine, and the polymorphisms of this gene have been shown to be of importance in a number of diseases. Especially, IL-1 has been suspected of involvement in allergic pathogenesis. Also, IL-1beta genotype is one of the genetic markers of gastric cancer. Therefore, we classified 166 individuals according to Iris constitution, and determined IL-1beta genotype. The frequencies of Iris constitutions as follows: neurogenic type, 41 (24.7%); abdominal connective tissue weakness type, 53 (31.9%); cardio-renal connective tissue weakness type, 50 (30.1%); the others type, 22 (13.3%). Especially, the frequency of abdominal connective tissue weakness type was higher in C/T genotype than in the remaining constitutions although the statistical power was very weak. Furthermore, we first attempted to explore possible involvement of the IL-1beta polymorphism and the Iris constitution. PMID- 15658615 TI - Clinical and molecular cytogenetic studies in a case with partial trisomy 12p due to a de novo supernumerary ring chromosome. AB - Clinical and molecular cytogenetic studies in a case with partial trisomy 12p due to a de novo supernumerary ring chromosome: We report on a girl with a mosaic karyotype containing a supernumerary ring chromosome. Fluorescence in situ hybridization (FISH) studies showed that this marker chromosome was derived from chromosome 12, resulting in partial trisomy 12p13.1-->12q11. The girl showed developmental delay, cerebral visual impairment, obesity and mild dysmorphic features. Her clinical data at 6 months, 3 years, and 6 years of age were compared with the clinical data on other trisomy 12p patients. PMID- 15658616 TI - Long term follow-up of two sibs with an autosomal recessive form of chrondrodysplasia punctata and epilepsy. AB - Long term follow-up of two sibs with an autosomal recessive form of chrondrodysplasia punctata and epilepsy: A variety of osteodysplasias are referred to with the term chondrodysplasia punctata (CDP). Here we report on two sibs, a boy and a girl, with probable autosomal recessive form of CDP and epilepsy followed-up for 30 and 19 years, respectively. Family history was unremarkable but for consanguinity. Pregnancies and deliveries were uneventful. At birth, length was 46 (-3SD) and 45 (-4SD) cm, respectively. Craniofacial dysmorphism was noted: severe nasal hypoplasia, flat face, hypertelorism, a low nasal bridge, short stature. Skeletal abnormalities included epiphyseal stippling in the thoracic spine, bilateral proximal and distal humeri, femur, tibia and bilateral carpal and tarsal bones. The boy had a hemivertebrae T12, with absence of a rib. After the age of 6 years facial dysmorphism had improved. Final height was 154 cm (-3SD) in the boy and 158 cm (-0,5SD) in the girl. The boy was operated on for scoliosis. Both sibs had club feet, the girl had also genu valgum. IQ was evaluated to be 55 in the girl and 83 in the boy. The first non febrile generalized seizure appeared in the boy when he was 11 months of age, and in the girl when she was 25 months of age. Both had many other seizures and were taking antiepileptics. EEG were abnormal. Karyotypes were normal. Extensive screening for metabolic disorders was normal. Acquired in utero CDP were excluded. We suggest the sibs described in this report have yet another provisionally unique possibly autosomal recessive syndrome, with CDP and epilepsy as phenotypic traits. PMID- 15658617 TI - Brachydactyly type E in two sibs with increased bone density and mental retardation. A new autosomal recessive syndrome? AB - Brachydactyly type E in two sibs with increased bone density and mental retardation. A new autosomal recessive syndrome?: We report on two sibs, a boy and a girl, with syndromic brachydactyly type E. Parents were first cousins. Facial dysmorphia was characterized by a flat occiput, a large forehead, hypertelorism, a long triangular nose, an everted lower lip, downslanting palpebral fissures and strabismus. They had marked shortening of the third, fourth and fifth fingers and of the third, fourth, and fifth toes. IQ was 16 in the boy, 63 in the girl. In both sibs ophtalmologic examination showed strabismus, absence of cataract and normal fundus and radiological findings disclosed increased bone density involving the skull, the vertebrae and the corticalis of the long bones. Neither ectopic calcifications, nor exostosic, nor osteomalacia, and nor osteotis fibrosa cystica were present. Investigations revealed that plasma calcium, phosphate, vitamine D, parathyroid hormone (PTH), response to exogenous PTH, and Gs activity were normal as well as renal and thyroid function. Molecular genetic studies failed to identify mutations in the GNAS 1 gene, in the PTH receptor gene and in the HOX D13 gene. Analysis of 2q showed that there was no deletion 2q37. Other known syndromes with brachydactyly type E and mental retardation were excluded. In conclusion we suggest that these two sibs with a combination of brachydactyly, mental retardation and increased bone density have a specific autosomal recessive syndrome. PMID- 15658618 TI - Prenatal diagnosis of a true fetal tetraploidy in direct and cultured chorionic villi. AB - Prenatal diagnosis of a true fetal tetraploidy in direct and cultured chorionic villi: Tetraploidy is characterized by four complete sets of chromosomes (4n= 92). Although it has been frequently reported in spontaneous abortions, tetraploidy is extremely rare in term pregnancy. Most of late surviving patients are diploid/tetraploid mosaics and present severe mental and physical impairment. Up to date, only five tetraploidies were ascertained in the prenatal stage in amniocytes and/or fetal blood lymphocytes. No one has been reported in chorionic villi probably because tetraploidy is generally considered in this tissue as a false positive result due to confined placental mosaicism (CPM) or placental culture artefacts. We report here on a case of tetraploidy detected in chorionic villi because of fetal cystic hygroma. We discuss the reliability of this diagnosis and propose guidelines in the follow-up of tetraploidies detected after chorionic villus sampling (CVS). Thus a misdiagnosis of this poor condition will be avoided at best and an appropriate genetic counseling will be given to the parents. PMID- 15658619 TI - De novo satellited 21q associated with corpus callosum dysgenesis, colpocephaly, a concealed penis, congenital heart defects, and developmental delay. AB - De novo satellited 21q associated with corpus callosum dysgenesis, colpocephaly, a concealed penis, congenital heart defects, and developmental delay: We present clinical and cytogenetic data on an infant with de novo satellited 21 q. A 3 month-old boy was found to have microcephaly, developmental delay, hypertelorism, down-slanting palpebral fissures, large low-set ears, a prominent nose, a broad philtrum, a concealed penis, interventricular septal defects, corpus callosum dysgenesis, colpocephaly, ventriculomegaly, and a de novo karyotype of 46,XY,21qs. Standard Ag-NOR staining and FISH studies confirmed a satellite and a deletion on the long arm of a chromosome 21. Quantitative-fluorescent polymerase chain reaction using the polymorphic small tandem repeat markers specific for chromosome 21 determined a maternal origin of the deletion and the breakpoint between D21S156 (21q22.1) (present) and D21S53 (21q22.3) (absent), centromeric to the known minimal holoprosencephaly critical region, D21S13-21qter. The present case provides evidence of the correlation of a distal region of chromosome 21 to the phenotypic effects of monosomy 21. PMID- 15658620 TI - A new case of a severe clinical phenotype of the cat-eye syndrome. AB - A new case of severe clinical phenotype of the cat-eye syndrome: We report on a female infant with severe clinical phenotype of Cat-Eye Syndrome (CES). At birth, she had respiratory distress and marked hypotonia. Physical examination showed major craniofacial anomalies including microcephaly, bilateral total absence of the external ears, hypertelorism, bilateral ocular coloboma of iris and micrognathia. In addition, she had anal stenosis, a patent ductus arteriosus and intra- and extra- hepatic biliary atresia. She deteriorated with the development of bradycardia. She died at age one month of cardiac failure. Cytogenetic analysis of the proband showed an extra de novo small bisatelllited marker chromosome in all cells examined. Molecular cytogenetic analysis with fluorescence in situ hybridization (FISH) identified the marker as a CES chromosome. Thus, the patient's karyotype was: 47, XX, +idic(22)(pter-->q11.2 ::q11.2-->pter). The duplication breakpoints giving rise to the CES chromosome were distal to the DiGeorge Syndrome (DGS) locus 22q11.2. The marker could be classed as a type 11 symmetrical (10). According to a recent review of CES literature (1) only 41 % of the CES patients have the combination of iris coloboma, anal anomalies and preauricular anomalies. Almost 60% are hard to recognize by their phenotype alone. Only twelve patients showed a severe clinical phenotype leading to the death of the child. This phenotypic variability increases the difficulties of genetic counseling. PMID- 15658621 TI - Trisomy 1q43 syndrome: a consistent phenotype with macrocephaly, characteristic face, developmental delay and cardiac anomalies. AB - Trisomy 1q43 syndrome: a consistent phenotype with macrocephaly, characteristic face, developmental delay and cardiac anomalies: Patients with trisomy (1)(q42 qter) present with psychomotor retardation, macrocephaly, occasional presence of facial capillary naevi, cardio-vascular anomalies and small size for gestational age. We report on a girl with the same pattern of malformations, who has pure trisomy 1 q43: duplication of the region (1) (q43) and the translocation of the terminal region of the other chromosome 1 to the derivative 1, narrowing down the critical region for the characteristic traits of severe developmental delay, macrocephaly and congenital cardiac malformations. PMID- 15658622 TI - Complete achromatopsia associated with skeletal anomalies: a new autosomal recessive syndrome. AB - Complete achromatopsia associated with skeletal anomalies: a new autosomal recessive syndrome: Achromatopsia or rod monochromatism is the complete absence of color discrimination, with an estimated frequency of 1 in 100,000. To date the McKusick Catalogue includes more than 10 entities related to Achromatopsia. This paper describes four Mexican sibs with a stationary rod monochromatism, associated with long fingers and toes, hypothenar and thenar hypoplasia and pes planus, suggesting a new genetic entity probably inherited in an autosomal recessive mode. PMID- 15658623 TI - Complex facio-audio-symphalangism syndrome. An autosomal recessive type? AB - Complex facio-audio-symphalangism syndrome. an autosomal recessive type?: This report describes a new case of facio-audio-symphalangism syndrome in a 32-year old female patient from a consanguineous family. She had a severely mentally retarded and anophthalmic sister. These associations might be coincidental or demonstrate genetic heterogeneity in this syndrome We note the diagnostic features of the case, discuss the novel association with consanguinity and highlight the possible heterogeneity of the facio-audio-symphalangism syndrome. PMID- 15658624 TI - Report of a girl with Klippel-feil syndrome and Poland anomaly. AB - Report of a girl with Klippel-feil syndrome and Poland anomaly: Klippel-Feil syndrome, consisting of the triad of a short neck, low posterior hairline, and limitation of neck movement, is a congenital anomaly characterized by the fusion of cervical vertebrae, Poland anomaly consists of unilateral aplasia of the chest wall muscles and ipsilateral anomalies of upper extremity. We report a 7-year-old girl with typical findings of Klippel-Feil syndrome and Poland anomaly. To the best of our knowledge a case of Klippel-Feil syndrome and Poland anomaly has not been described before, although a combination of Poland, Klippel-Feil and Moebius anomalies has been reported in the literature. PMID- 15658625 TI - Discrepancy between the fetus and extra-embryonic tissues in prenatally detected mosaic distal 5p deletion. AB - Discrepancy between the fetus and extra-embryonic tissues in prenatally detected mosaic distal 5p deletion: We present clinical and cytogenetic data on a second trimester fetus with mosaic del(5)(p15.1) and the extra-embryonic tissues with a normal karyotype. A 34-year-old woman, gravida 2, para 0, underwent genetic amniocentesis at 20 weeks' gestation because of advanced maternal age. Cytogenetic analysis of the cultured amniocytes revealed mosaicism for a distal 5p deletion, mos 46,XY,del(5)(p15.1)[4]/46,XY[26]. The pregnancy was terminated subsequently. Postnatally, the fetus displayed a triangular face, hypertelorism, epicanthal folds, low-set ears, and micrognathia. A karyotype of mos 46,XY,del(5)(pl 5.1)/46,XY was found in the liver, lungs, skin, and cord blood, whereas, the placenta, amnion, and umbilical cord had a karyotype of 46,XY. Our observation of fetoplacental, fetoamniotic, and fetoumbilical discrepancies shows a limitation of using placenta, amnion, and umbilical cord as confirmatory tools for prenatally detected mosaic distal 5p deletion. Our case also reinforces the notion that amniocentesis offers a more reliable diagnosis, compared to chorionic villus sampling. PMID- 15658626 TI - Megalocornea-mental retardation (MMR or Neuhauser) syndrome: another case associated with cerebral cortical atrophy and Bifid uvula. PMID- 15658628 TI - Perinatal identification of caudal regression syndrome and alobar holoprosencephaly in pregnancies with poor maternal metabolic control. PMID- 15658627 TI - Partial craniosynostosis in a patient with deletion 22q11. PMID- 15658629 TI - 46, XY gonadal dysgenesis and chronic renal failure: first reported case with Frasier syndrome from Turkey. PMID- 15658630 TI - President's report. PMID- 15658631 TI - Incoming President's address. PMID- 15658632 TI - The Knud Jansen Lecture. Culture-sensitive innovations for quality living of lower limb amputees. PMID- 15658633 TI - The Brian Blatchford prize acceptance speech. PMID- 15658634 TI - Fibular segment bone bridging in trans-tibial amputation. AB - The purpose of this paper is to present an alternative method of achieving a synostosis between tibia and fibula in trans-tibial amputations by means of a fibular segment in place of tibial osteoperiosteal flaps as described by Ertl, in 1949. From May 1997 through to February 2003, 15 patients were submitted to the proposed procedure. As a result, all patients produced a solid synostosis between tibia and fibula and were rehabilitated with the use of prostheses. The capacity of these patients for distal weight-bearing on the stump was remarkable when compared to patients submitted to the conventional trans-tibial amputation technique. PMID- 15658635 TI - The Northern Ireland troubles and limb loss: a retrospective study. AB - The Troubles in Northern Ireland have now lasted 34 years. Divisions and strife between the opposing loyalist and republican communities, and between the communities and the security forces, have led to thousands of deaths and injuries. The violence has often been indiscriminate injuring and killing totally innocent people. Staff at the Regional Disablement Services at Musgrave Park Hospital, Belfast have had the responsibility for helping to rehabilitate those who have suffered limb loss, both civilians and security forces personnel. In this study the authors present patient demographics for those survivors, referred for prosthetic fitting, who have sustained limb amputations as a result of the Troubles from 1969 to 2003, with the cause of injury, resulting levels of amputation, associated injuries, time to first prosthetic fitting and reason for any delay in fitting identified. One hundred and twenty-nine (129) patients sustained amputations, 110 male and 19 female with an age range at the time of injury from 7 to 60 years. Seventy-two (72) were civilian. Ninety-three (93) underwent immediate amputation, the most frequent level of amputation being trans femoral. Delayed healing of deep wounds was the most common reason for delayed amputation; other causes were chronic osteomyelitis, malunited fractures and failed arthrodesis, often associated with chronic pain. Ninety-two (92) patients required amputation of one limb or part thereof, 35 required amputation of 2 limbs and 2 underwent triple amputation. Three (3) patients lost both hands. Sixty seven percent (67%) had other associated physical injuries. Thirty-two (32) patients had a delay of 6 months or more in fitting a prosthesis. The most common cause of injury was the car bomb. PMID- 15658636 TI - Clinical field follow-up of high density polyethylene (HDPE)-Jaipur prosthetic technology for trans-tibial amputees. AB - The purpose of this study was to examine the outcome of the application of the High Density Polyethylene (HDPE)-Jaipur prosthetic construction in fitting trans tibial amputees in a number of projects in the developing world. Projects in Honduras, Uganda and India were visited. Three hundred and twenty (320) patients had been provided with a HDPE-Jaipur prosthesis and of these 172 were seen for a technical and clinical follow-up after a median of 35 months. More than half the amputations were due to trauma, the remainder to disease. Fabrication and fitting in the three projects was carried out by individuals who had undertaken a twice week training course provided by Bhagwan Mahaveer Viklang Sahayata Samiti limb centre in Jaipur. The individuals involved had limited background training in prosthetics. Craftsmanship and fit were assessed as being poor in 56% of cases. The technical quality of the Jaipur foot was considered acceptable as its performance was better than previously observed results. Although there was patient satisfaction of 85% and compliance of 94% the HDPE-Jaipur trans-tibial system was not considered acceptable as 49% reported walking distances less than 1km and 36% discomfort. The major inadequacy in outcome relates to the use for fabrication and fitting of individuals with inadequate education and training. PMID- 15658638 TI - Orthotic design and foot impression procedures to control foot alignment. AB - The traditional theory on subtalar joint neutral position and intrinsic foot deformities for the evaluation and treatment of foot and ankle disorders has been the basis for foot orthotics for many years. Although clinical evaluations have suggested a relationship between subtalar pronation and a variety of lower limb problems, such as shin splints and anterior knee pain, recent research has raised serious concerns about the reliability and validity of the assessment and intervention methods. Results of recent studies in foot biomechanics suggest that the orthosis design to control foot alignment should stabilise the medial apical bony structure of the arch to control the first ray mobility and transmit load through the lateral support structures of the foot, locking the calcaneocuboid joint and decreasing strain in the plantar aponeurosis. The concept of "posting" according to a measured foot deformity is de-emphasised. Reliable foot impression procedures are required to provide appropriate orthotic design and thus management. A prone lying position manipulated foot impression method using polycaprolactone based low temperature thermoplastic material was introduced. Ten (10) subjects were recruited to participate in the reliability tests, which were conducted by 2 orthotists specialized in foot orthotics. Results showed high intrarater and interrater reliability of the measured forefoot width and the navicular height. The reliability of the forefoot-rearfoot relationship was demonstrated by the small variance of the root mean square calculation. Subsequently orthotic intervention can be done in a more consistent manner. PMID- 15658637 TI - The use of targeted muscle reinnervation for improved myoelectric prosthesis control in a bilateral shoulder disarticulation amputee. AB - A novel method for the control of a myoelectric upper limb prosthesis was achieved in a patient with bilateral amputations at the shoulder disarticulation level. Four independently controlled nerve-muscle units were created by surgically anastomosing residual brachial plexus nerves to dissected and divided aspects of the pectoralis major and minor muscles. The musculocutaneous nerve was anastomosed to the upper pectoralis major; the median nerve was transferred to the middle pectoralis major region; the radial nerve was anastomosed to the lower pectoralis major region; and the ulnar nerve was transferred to the pectoralis minor muscle which was moved out to the lateral chest wall. After five months, three nerve-muscle units were successful (the musculocutaneous, median and radial nerves) in that a contraction could be seen, felt and a surface electromyogram (EMG) could be recorded. Sensory reinnervation also occurred on the chest in an area where the subcutaneous fat was removed. The patient was fitted with a new myoelectric prosthesis using the targeted muscle reinnervation. The patient could simultaneously control two degrees-of-freedom with the experimental prosthesis, the elbow and either the terminal device or wrist. Objective testing showed a doubling of blocks moved with a box and blocks test and a 26% increase in speed with a clothes pin moving test. Subjectively the patient clearly preferred the new prosthesis. He reported that it was easier and faster to use, and felt more natural. PMID- 15658639 TI - Elbow orthoses: a review of literature. AB - The objective of this study is to review the available literature on elbow orthoses in patients with various diagnoses to assess the scientific base of the prescription of elbow orthoses. A search of literature in Medline, Embase, Cochrane and Recal was performed using the keyword "elbow" combined with "orthosis related keywords". Abstracts were read to select the papers dealing primarily with monoarticular elbow orthoses. References of the selected papers were also examined. A total of 57 papers were read of which 18 met the selection criteria. Both the quality and quantity of the studies appeared to be low, so in this study no definitive conctusions can be drawn about the efficacy of monoarticular elbow orthoses. Current prescriptions of elbow orthoses cannot be evidence based, because no scientific evidence on elbow orthoses is available. PMID- 15658640 TI - The short -term effects of an exercise programme as an adjunct to an orthosis in neuromuscular scoliosis. AB - The purpose of this study was to investigate the effects of a 4 week physiotherapy programme on patients who were given a spinal orthosis for neuromuscular scoliosis. This study was planned as a single group pre- and post intervention repeated measures design. All patients were given a polyethylene spinal orthosis with an anterior opening. Fifteen (15) patients with neuromuscular diseases and a mean age of 12.46 years were evaluated. An exercises programme consisting of postural training, muscle strengthening and stretching exercises with special emphasis on respiratory exercises was given as an adjunct to orthotic treatment. The degree of impairment in forced vital capacity was 17.56% upon wearing an orthosis, it decreased to 9.28% following therapy (p < 0.05). There was also a statistically significant increase in muscle strength, balance duration and a significant decrease in limitation of range of motion. The results of the study imply that the conservative treatment of neuromuscular scoliosis should include an exercise programme as an adjunct to an orthosis, both to reduce the compromising effect of an orthosis on respiratory function and to support the patient's physical capacities. PMID- 15658641 TI - Postoperative immobilisation orthosis for surgically corrected hallux valgus. AB - A postoperative immobilisation orthosis was developed at the School of Physical Therapy and Rehabilitation, Department of Orthotics and Prosthetics, and applied from the first postoperative day as an alternative device for ambulation of patients who have undergone hallux valgus surgery while maintaining the corrected position. The orthosis is constructed with polyethylene thermoplastic sheet of 3mm thickness and Velcro. During the ambulation period, there was no complication or complaint due to its application and the position of the great toe was maintained. The time of the orthotic application was six weeks. The orthosis has allowed patients to walk with full weight bearing from the postoperative first day without any complication or recurrence. PMID- 15658642 TI - [Working cooperatively--to advance the cause of Chinese medicine modernization and promote integrative medicine]. PMID- 15658643 TI - [Practice has proved the importance of integrating Chinese and Western medicine]. PMID- 15658644 TI - [The current status and trend of development of research on non-steroids anti inflammatory agents of Chinese and Western medicine]. PMID- 15658645 TI - [Effect of tongguan capsule on coagulant and fibrinolysis system in patients with coronary heart disease after percutaneous coronary intervention]. AB - OBJECTIVE: To explore the effect and mechanism of Tongguan capsule (TGC) on coagulant and fibrinolysis system in patients with coronary heart disease (CHD) after percutaneous coronary intervention (PCI). METHODS: Adopting the prospective, randomized controlled method to observe the coagulant-fibrinolysis related indexes, including tissue plasminogen activator (t-PA), plasminogen activator inhibitor 1 (PAI-1), von Willebrand factor (vWF), antithrombin II (AT III) and fibrinogen (FIB), in CHD patients after PCI, there were 26 patients in the treated group treated by TGC with Western medicine and 26 in the control group treated by Western medicine alone. RESULTS: After treatment, all the indexes were improved in both groups (P < 0.05). As comparison between the two groups, AT-III and t-PA were higher in the treated group than those in the control group; PAI-1 levels showed insignificant difference at 1 month after treatment, but at 3 months after treatment, it was significantly lower in the treated group (P < 0.05); vWF showed no significant difference either at 1 or at 3 months after treatment; and FIB was lower in the treated group both at 1 and 3 months after treatment (P < 0.01). CONCLUSION: TGC could improve the hypercoagulant status and adjust the balance between coagulant-fibrinolysis system of CHD patients after PCI by increasing AT-III and t-PA levels, and lowering FIB and PAI-1 levels in the body. PMID- 15658646 TI - [Change of peripheral blood monocytes derived macrophage scavenger receptors activity in patients with coronary heart disease, and the intervention effect of ginkgo biloba extract]. AB - OBJECTIVE: To observe the activity of peripheral blood monocytes (PBMs) derived macrophage scavenger receptors (MSR) and changes of serum inflammatory factor in peripheral blood in patients with coronary heart disease (CHD), and to evaluate the effect of Ginkgo biloba extract (GBE) on the MSR activity, to explore the relationship between inflammatory factor and scavenger receptors activity as well as the possible mechanism of GBE in stabilizing the atheromatous plaque. METHODS: Ninety-seven CHD patients with normal blood lipids were classified into the stable angina group, the unstable angina group and the acute myocardial infarction group, and 29 healthy persons were taken as control. Levels of C reactive protein (CRP), soluble intercellular adhesion molecule-1 (sICAM-1) and soluble vascular cell adhesion molecule-1 (sVCAM-1) in all subjects were determined. And their PBMs were isolated, cultured in vitro, and transferred into macrophage to observe the effect of GBE on the expression of scavenger receptors. RESULTS: The levels of MSR activity, CRP, sICAM-1 and sVCAM-1 in patients with acute myocardial infarction > unstable angina > stable angina > control. CONCLUSION: GBE could down-regulate the MSR activity in CHD patients, which was positively correlated with levels of CRP, sICAM-1 and sVCAM-1. MSR activity could be taken as a monitoring criteria for active degree of vulnerable atherosclerosis plaque. GBE has the effect of suppressing MSR activity. PMID- 15658647 TI - [Intervention of xuezhikang on patients of acute coronary syndrome with different levels of blood lipids]. AB - OBJECTIVE: To observe the intervention of Xuezhikang (XZK) on patients of acute coronary syndrome (ACS) with different levels of blood lipids. METHODS: Adopting the double blind randomized controlled method, 105 patients of ACS were divided into two groups. The 53 patients in the treated group, 26 with normal blood lipids (NBL) and 27 with hyperlipemia (HL) were treated with conventional western medicine plus XZK 1.2 g per day for 12 weeks. The 52 patients in the control group, 25 with NBL and 27 with HL, were treated with conventional western medicine alone. Besides, a healthy control group consisted of 40 subjects was set up. The changes of brachial arterial endothelium-dependent diastolic function (FMD) before and after treatment was observed, the changes of blood levels of nitric oxide (NO), endothelin-1 (ET-1), C-reactive protein (CRP) and lipids were also recorded. RESULTS: Before treatment, FMD value and serum NO level were lower and ET-1 and CRP levels in ACS patients were higher than those in the healthy subjects, and a significant correlation existed between CRP, NO and FMD with LDL C. After treated for 12 weeks, FMD value and serum NO level increased, levels of ET-1 and CRP decreased significantly in the treated group, showing significant difference to those in the control group (P < 0.05, P < 0.01). Serum levels of TC, TG and LDL-C in the treated group lowered significantly. HDL-C level in patients with HL increased significantly while in those with NBL, it showed a trend of increasing but with no statistical significance. CONCLUSION: Applying XZK in ACS patients in early stage, either with NBL or with HL, could improve the endothelial function, antagonize inflammatory response to stabilize the atheromatous plaque. PMID- 15658648 TI - [Effect of shexiang baoxin pill on the function of vascular endothelium in patients with diabetes mellitus type 2 complicated with angina pectoris]. AB - OBJECTIVE: To study the effect of Shexiang Baoxin pill (SBP) on the vascular endothelial function in patients with diabetes mellitus type 2 (DM2) complicated with angina pectoris. METHODS: Two weeks after runin, according to the randomizing table, 111 patients were divided into two groups, the XBP group (56 patients) and the control group (55 patients, treated with delayed-released isosorbide mononitrate, DRIM), they were treated for 6 months. In the treatment period, the episodes of angina attack and condition of rescue medication were recorded in the daily card, and brachial arterial changes of endothelium dependent relaxing function before and after treatment were measured by B ultrasonography. RESULTS: Comparison between the two groups in episodes of angina attack and rescue medication were insignificantly different. In the control group, the basal value of brachial arterial inner diameter before and after treatment was 3.68 +/- 0.56 mm and 3.70 +/- 0.58 mm respectively, those before and after responsive congestion was 5.44 +/- 0.81% vs 5.68 +/- 0.83%, and those before and after taking nitroglycerin was 19.8 +/- 4.9% vs 20. +/- 5.2%, all showed insignificant difference (P > 0.05). In the SBP group, the corresponding basal value was 3.73 +/- 0.62 mm vs 3.71 +/- 0.59 mm, and those after taking nitroglycerin 18.8 +/- 4.5 % vs 19.2 +/- 5.8%, also showed insignificant difference, but those before and after responsive congestion (5.69 +/- 0.79 % vs 9.56 +/- 3.82 %) did show significant difference (P < 0.01). CONCLUSION: XBP could improve the vascular endothelial function in patients with DM2 complicated with angina pectoris. PMID- 15658649 TI - [Effect of needling quchi and taichong points on blood levels of endothelin and angiotension converting enzyme in patients with hypertension]. AB - OBJECTIVE: To observe the effect and explore the mechanism of needling Quchi and Taichong points in treating hypertension patients and the influence on blood levels of angiotension converting enzyme (ACE) and endothelin (ET) levels. METHODS: Sixty hypertension patients were randomly divided into the Taichong needling group (A), Quchi needling group (B) and control group (C, treated by Captopril). Changes of plasma ET was determined by radioimmunoassay (RIA) and serum ACE content was measured by chemical colorimeter. RESULTS: The effect of lowering systolic pressure at 15 min after needling in Group B was better than that in Group A (P < 0.01), but it was inferior to the latter at 120 min after withdrawal of needle (P < 0.05), while after one course treatment, the effect in Group B and C was obviously better than that in Group A (P < 0.05 and P < 0.01). Content of serum ACE significantly increased in Group B and that of plasma ET significantly decreased in Group A, showing significant difference between the two groups, all P < 0.01. CONCLUSION: Needling Quchi and Taichong all show hypertensive effect, the former is obviously higher than that of the latter. They could regulate the blood level of ACE and ET, protect and repair vascular endothelial cells, but the key links of their mechanism might be different. PMID- 15658650 TI - [Clinical study on treatment of mid-advanced crescentic nephritis by qingre huoxue recipe]. AB - OBJECTIVE: To evaluate the therapeutic effect of integrative Chinese and Western medicine in treating mid-advanced crescentic nephritis (MACN). METHODS: Thirty two patients, their diagnosis was confirmed as MACN by renal biopsy, were divided, adopting randomized, controlled method, into two groups, the treated group and the control group, they were all, excepting one, treated with impact therapy of methyl-prednisolone followed with oral intake of prednisone, to part of them cyclophosphamide or mycophenolate mofetil was given in addition, to those with hypo-hemoglobin (< 90 g/L), subcutaneous injection of erythropoietin was administered. Decoction of Qingre Huoxue recipe (QHR), consisted of oldenlandia herb 30 g, honey-suckle stem 30 g, violet herb 30 g, red peony root 15 g, rehmannia root 15 g, solomonseal rhizome 15 g, asiabell root 30 g, red sage root 30 g, prepared rhubarb 12 g and giant-hyssop herb 12 g, were additionally given one dose per day to patients in the treated group. The renal function, improvement of anemia and immunosuppressive agents needed in patients were observed after 3 months treatment. RESULTS: After treatment, renal function was improved in both groups, but the effect in the treated group was better than that in the control group (P < 0.05). Anemia was partially alleviated in the two groups with no significant difference. The dosage of glucocorticoids used in the treated group was obviously lesser than that used in the control group (P < 0.01). CONCLUSION: Integrative Chinese and western medicine could treat crescentic nephritis to obtain good effect, and reduce the quantity of glucocorticoid necessity for treatment. PMID- 15658651 TI - [Effect of liuwei dihuang pill on erythrocyte aldose reductase activity in early diabetic nephropathy patients]. AB - OBJECTIVE: To observe the inhibitory effect of Liuwei Dihuang Pill (LDP) on erythrocyte aldose reductase (EAR) activity in early diabetic nephropathy (DN) patients and to explore the clinical significance of applying LDP in prevention and treatment of DN as an inhibitor of aldose reductase. METHODS: Seventy-two patients diagnosed as early DN with TCM Syndrome of both Qi-yin deficiency were randomly divided into the control group (31 patients) treated by conventional therapy (orally taken Gliquidone or injection of insulin) and the treated group (41 patients) treated by conventional therapy plus LDP for 3 months as one therapeutic course. Changes of symptoms, physical signs, fasting blood glucose (FBG), blood glucose 2 hrs post breakfast (2hPBG), blood total cholesterol (TC), triglyceride (TG), EAR activity, urinary albumin excretion rate (UAER), blood and urinary beta2-microglobulin (beta2-MG) in patients before and after treatment were observed. RESULTS: LDP could improve the symptoms and signs of patients with DN, it could significantly inhibit EAR activity, to make it significantly lower than that in the control group (P < 0.05), and the levels of UAER, beta2-MG in blood and urine in the treated group after treatment were obviously lower than those in the control group (P < 0.05), and LDP showed no apparent effect on blood glucose, lipids and mean arterial pressure (P < 0.05). CONCLUSION: LDP could obviously inhibit the activity of EAR in patients with early DN, improve various indexes of DN, so as to be helpful for its treatment. PMID- 15658652 TI - [Renal tubular protection of shenqi fuzheng injection in treating primary nephrotic syndrome]. AB - OBJECTIVE: To explore the effect and mechanism of Shenqi Fuzheng injection (SFI) on renal tubule in treating patients with primary nephrotic syndrome (PNS). METHODS: Eighty patients with PNS were randomly divided into two groups, 40 patients in the control group were treated by conventional therapy mainly with glucocorticoids; 40 patients in the treated group treated by conventional therapy plus intravenous injection of SFI additionally. The changes of serum albumin (sALB), urinary protein in 24 hrs (UP/24h), urinary levels of retinal binding protein (RBP), beta2 microglobulin (beta2-MG) and N-acetyl-D-glucosaminidase (NAG) as well as blood lipids before and after treatment were observed. RESULTS: After treatment, level of sALB increased and urinay levels of RBP, beta2-MG, NAG, UP/24h decreased in the treated group, as compared with those in the control group, showing significant difference (P < 0.05 or P < 0.01). Blood lipids level abated significantly after treatment in both groups (P < 0.01), comparison between them, levels of TC and LDL-C in the treated group were significantly lower than those in the control group (P < 0.01). CONCLUSION: SFI has protective effect on renal tubule in patients with PNS, also has effect on lowering blood lipids. PMID- 15658653 TI - [Experimental study on effect of epimedium flavonoids in protecting telomere length of senescence cells HU]. AB - OBJECTIVE: To investigate the mechanism of senescence delay of human diploid fibroblast (2BS) and protecting telomere length by epimedium flavonoids (EF). METHODS: The drug sera of EF were used to treat the 2BS. The population doublings of 2BS cells were observed, the mRNA expression of p16 gene were determined by fluorescence real-time quantitative RT-PCR, the telomerase activation of 2BS cells were determined by TRAP-Hyb, the total retinoblastoma (Rb) and phosphorated Rb protein content were detected by ELISA, the telomere length of 2BS cells were determined by telomere restriction fragment (TRF) Southern blot assay. RESULTS: EF could significantly extend the population doublings of 2BS cells, the expression of p16 mRNA was decreased and the content of phosphorated Rb protein were increased by EF. The telomere lengthening of 2BS cells were improved by EF, but the telomerase was not activated. CONCLUSION: In senescence human fibroblasts 2BS cells, p16 gene mRNA expression increased, content of phosphorated Rb protein decreased and the telomere length of 2BS shortened, EF might delay the aging of cells through inhibiting the p16 gene expression, promoting the production of phosphorated Rb protein and to protect the length of telomere, but not activating the telomerase. PMID- 15658654 TI - [Characteristics of oriented induction of xiangdan injection on differentiation of marrow mesenchymal stem cells into neurons and its influencing factors]. AB - OBJECTIVE: To study the effect of Xiangdan injection (XDI) in inducing adult SD rat marrow mesenchymal stem cells (rMSCs) orientedly differentiated into neuron like cells, its characteristics and influencing factors were explored. METHODS: The 5th generation of rMSCs cultured in vitro were pre-treated for 24 hrs by adding basic fibroblast growth factors (bFGF) into the medium, then the inducing liquid was replaced by XDI with different concentration to compare the rMSCs differentiation rate under different constitution of medium, different concentration of inducer and cell density of incubation. The induced cell survival rate under effects of above-mentioned factors was evaluated by trypan blue stain and MTT method. RESULTS: XDI in 1% - 5% concentration could induce rMSCs differentiated into neuron-like cells, the inducing rate reached 83.5 +/- 3.8% 6 - 12 hrs later, more than 90% cells, survival rate was over 36 hrs. The maximal inducing rate and cell survival rate could be obtained by treated with 3 % - 5% XDI, serum-free D/F12 + N2 + bFGF and with the cell density in 2.5 x 10(4)/cm2, when the other factors were the same. CONCLUSION: XDI of 3% - 5% concentration, serum-free D/ F12 + N2 + bFGF (10 microg/L) and with the cell density incubated of 2.5 x 10(4)/cm2 is the optimal condition for oriented induction of rMSCs differentiating to neuron-like cells. PMID- 15658655 TI - [Study on the pharmacological mechanism of sodium ferulate for anti-asthmatic effect in guinea pigs]. AB - OBJECTIVE: To study the anti-asthmatic effect of sodium ferulate (SF) and its mechanism in guinea pig asthmatic model. METHODS: Guinea pigs were sensitized with ovalbumin as animal asthmatic model and treated with 3 different concentration of SF for 8 days. Levels of endothelin (ET) and nitric oxide (NO) in blood and lung tissue, and eosinophil (EOS) in blood and bronchoalveolar lavage fluid (BLAF) was counted at the end of trial. RESULTS: SF could significantly lower the ET content and increase the NO concentration in serum and lung tissue, reduce the number of EOS in blood and BLAF (P < 0.05, P < 0.01). Stronger effect was showed in the high dose group. CONCLUSION: Mechanism of anti asthmatic action of SF might be to increase NO concentration, lower ET content, alleviate EOS infiltration. PMID- 15658656 TI - [Effect of emodin on motility signal transduction in colonic smooth muscle cells in rats with multiple organ dysfunction syndrome]. AB - OBJECTIVE: To observe the effect of emodin on motility signal transduction and calcium ion in colonic smooth muscle cells (SMC) in rats with bacterial peritonitis caused multiple organ dysfunction syndrome (MODS). METHODS: Observation was conducted in colon of MODS model rats on (1) effects of emodin on the contraction of muscular strip and cells of colonic smooth muscle, and influences of specific myoglobulin light chain kinase inhibitor (ML-7) and selective proteinkinase C inhibitor (Calphostin C) on these effects; and (2) effect of emodin on calcium ion in SMC. RESULTS: Emodin could directly contract the muscular strip and cells of smooth muscle; ML-7 and Calphostine could inhibit these contractile action to some extent. Under MODS condition, emodin could still increase the intracellular calcium ion concentration; this effect could be inhibited by heparin (inosamine triphosphate receptor inhibitor IP3 and ryanodine receptor inhibitor in MODS model but the calcium chelator EGTA and nifedipine (the specific cell membrane voltage dependent calcium channel blocker) showed no influence on it. CONCLUSION: Emodin could directly contract the colonic smooth muscle in MODS model rats, which is mediated by raise the signal path MLCK of calcium ion and the PKCa path for increase calcium sensibility. The mechanism of increasing calcium ion is mainly through IP3 and RyR the two calcium ion channel receptor in the sarcoplasm. PMID- 15658657 TI - [Protection of artesunate on activation and injury of vascular endothelial cells induced by lipopolysaccharide]. AB - OBJECTIVE: To investigate the protective effects and mechanism of artesunate (AR) on the activation and injury of human umbilical vein endothelial cells (HUVECs) induced by lipopolysaccharide (LPS). METHODS: After HUVECs were cultured and turned to fusion manner, LPS and different concentration of AR (0.04 mg/L, 0.2 mg/L, 1 mg/L, 5 mg/L and 20 mg/L) were added respectively and co-incubated for 24 hrs. The expression of von Willebrand factor (vWF) in the conditioned media was tested by ELISA, the expression of intercellular adhesion molecule (ICAM-1) protein was determined by Western blot method and the expression of tumor necrosis factor alpha (TNFalpha) mRNA was determined by in situ hybridization. RESULTS: After being exposed to 1 microg/ml LPS, vWF and ICAM-1 expression were higher than those in the control group. AR could significantly down-regulate the increased expressions concentration-dependently, significant difference showed as the concentration of AR reached 1 mg/L (P < 0.05). In situ hybridization showed that AR in 0.2 mg/L and 1 mg/L could markedly down-regulate the TNFalpha mRNA expression, showing significant difference as compared with that in LPS group (P < 0.05, P < 0.01). CONCLUSION: AR has protective effect on LPS induced HUVECs activation and injury, which might be related with its inhibition on TNFalpha mRNA expression. PMID- 15658658 TI - [Experimental study on xiaoliu decoction in treating tumor and modulating immune function]. AB - OBJECTIVE: To study the anti-tumor effect and immune function modulation of Xiaoliu decoction (XLD) and to explore the possible mechanism. METHODS: S180 tumor bearing mice were treated with large or small dose of XLD and cyclophosphamide (CTX) to observe the tumor suppressive rate, lymphocyte transformation rate, natural killer cell (NK) activity, tumor necrosis factor alpha (TNFalpha) level in the mice, as well as the tumor cell apoptosis in vitro. RESULTS: The tumor suppressive rate of XLD on tumor bearing mice was over 30%. XLD showed obvious trend in enhancing the tumor suppressing rate of chemotherapy, could increase the lymphocyte transformation rate, NK activity, and TNFalpha level. It also could accelerate the tumor cell apoptosis in vitro. CONCLUSION: XLD has significant anti-tumor effect, it could elevate the immune function or organism and alleviate the chemotherapy induced lowering of immune function. PMID- 15658659 TI - [Clinical observation on treatment of functional dyspepsia with weichangshu decoction]. PMID- 15658660 TI - [Effect of modified chaihu shugan powder in treating patients with functional dyspepsia accompanied with depression]. PMID- 15658661 TI - [Clinical observation on treatment of 90 patients with primary menalgia with guizhi fuling capsule]. PMID- 15658663 TI - [Principle and methods for English translation of TCM pulse-figure]. PMID- 15658662 TI - [Study of the therapeutic principle of scavenging evil-heat and removing toxin]. PMID- 15658664 TI - [Progress on the integrative Chinese and Western medicinal study of endometriosis]. PMID- 15658665 TI - Medullary thyroid carcinoma. AB - Medullary thyroid carcinoma (MTC) is subdivided into sporadic (75 %) and hereditary (25 %) forms. Several germline mutations in the RET proto-oncogene are the source of distinct clinical phenotypes in hereditary MTC including familial MTC (FMTC) and multiple endocrine neoplasia 2A (MEN 2A) and 2B (MEN 2B). The higher the penetrance of the MEN 2 phenotype the earlier the progression of MTC which forms the basis for the currently recommended codon-related concept of prophylactic thyroidectomy. In patients with sporadic MTC, routine calcitonin (CT) measurement in nodular goiter patients has been shown to reduce the frequency of advanced tumor stages. Patients with CT levels over 100 pg/ml after pentagastrin stimulation are recommended for total thyroidectomy. In patients with unexpected sporadic MTC after histological examination, completion thyroidectomy is currently only recommended when CT levels remain elevated. The extent of lymph node dissection in patients with MTC is controversial. However, with respect to lymphonodal micrometastases, systematic compartment-oriented microdissection has been shown to reduce the frequency of lymphonodal recurrence. On the other hand, to avoid unnecessary lymph node dissection, a more individualized concept is required in the future. New chemotherapeutic agents (tyrosine kinase inhibitors), therapeutic nuclids (90Yttrium-labeled octreotide), and chemoembolization of liver metastases are currently the most promising therapeutical concepts in patients with distant metastases. PMID- 15658666 TI - Well differentiated thyroid cancer. PMID- 15658668 TI - Benign multinodular goiter. PMID- 15658667 TI - Anaplastic giant cell thyroid carcinoma. AB - Anaplastic (giant cell) thyroid carcinoma (ATC), is one of the most aggressive malignancies in humans with a median survival time after diagnosis of 3-6 months. Death from ATC was earlier seen because of local growth and suffocation. ATC is uncommon, accounting for less than 5 % of all thyroid carcinomas. The diagnosis can be established by means of multiple fine needle aspiration biopsies, which are neither harmful nor troublesome for the patient. The cytological diagnosis of this high-grade malignant tumour is usually not difficult for a well trained cytologist. The intention to treat patients with ATC is cure, although only few of them survive. The majority of the patients are older than 60 years and treatment must be influenced by their high age. We have by using a combined modality regimen succeeded in achieving local control in most patients. Every effort should be made to control the primary tumour and thereby improve the quality of remaining life and it is important for patients, relatives and the personnel to know that cure is not impossible. Different treatment combinations have been used since 30 years including radiotherapy, cytostatic drugs and surgery, when feasible. In our latest combined regimen, 22 patients were treated with hyper fractionated radiotherapy 1.6Gy x 2 to a total target dose of 46 Gy given preoperatively, 20 mg doxorubicin was administered intravenously once weekly and surgery was carried out 2-3 weeks after the radiotherapy. 17 of these 22 patients were operated upon and none of these 17 patients got a local recurrence. In the future we are awaiting the development of new therapeutic approaches to this aggressive type of carcinoma. Inhibitors of angiogenesis might be useful. Combretastatin has displayed cytotoxicity against ATC cell lines and has had a positive effect on ATC in a patient. Sodium iodide symporter (NIS) genetherapy is also being currently considered for dedifferentiated thyroid carcinomas with the ultimate aim of making radio-iodine therapy possible. PMID- 15658669 TI - Primary hyperparathyroidism. Update on pathophysiology, clinical presentation and surgical treatment. AB - The most important step in calcium homeostasis is the regulation of parathyroid hormone (PTH) secretion. The discovery and characterization of the calcium sensing receptor (CaR) of the parathyroid cell has led to a better understanding not only of the physiology of the parathyroid glands, but also of the development of hyperparathyroidism. Drugs acting on CaR can now be designed to treat hyperparathyroidism and osteoporosis. The workshop on primary hyperparathyroidism held at the National Institutes of Health in 2002 has recommended new guidelines for the treatment of asymptomatic hyperparathyroidism. Controversy still exists regarding the treatment of patients with non-classical symptoms, such as weakness, fatigue and depression. Primary hyperparathyroidism as a risk factor for cardiovascular disease and mortality is also debated. Improved techniques for the preoperative localization of pathological parathyroid glands have led to a shift in surgical strategy: surgeons abandon the traditional bilateral neck exploration in favor of a more limited approach. This change of strategy has not been based on the results of prospective randomized studies and the long term results are not known. PMID- 15658670 TI - Medical and surgical treatment for secondary and tertiary hyperparathyroidism. AB - Prevention and treatment of secondary hyperparathyroidism (SHPT) in patients on chronic maintenance hemodialysis and of tertiary hyperparathyroidism (THPT) in patients after kidney transplantation is a challenge for the nephrologist and for the surgeon. Indication and results of medical and surgical therapy for SHPT and THPT have remained under discussion during the last decades. This review resumes the current medical and surgical strategies for patients with SHPT and THPT. PMID- 15658671 TI - Adrenal incidentaloma. AB - Adrenal incidentalomas are tumors that are serendipitously found by radiological examination. The incidence of adrenal incidentalomas in patients undergoing abdominal CT approaches 1 %. The evaluation of a patient with an adrenal incidentaloma requires, in addition to a clinical history and physical examination, a focused biochemical evaluation to investigate if there is excess secretion of catecholamines, glucocorticoids, or aldosterone. Some tumors have specific features on imaging that identify them as benign or malignant. The recommendations for management of adrenal incidentalomas include resection of all functioning tumors regardless of size, preferably by the laparoscopic approach. Large, non-functional tumors should also be removed. Biochemical and radiological surveillance is recommended for at least one year, if a tumor is left in place. PMID- 15658673 TI - Endocrine pancreatic tumors. AB - Neuroendocrine tumors of the pancreas are rare neoplasms that may arise sporadically or in association with a hereditary endocrine neoplasia syndrome. Effective management requires directed biochemical testing, careful choice of preoperative imaging tests, and complete pancreatic exploration by an experienced endocrine surgeon utilizing intraoperative ultrasound. Pancreatic endocrine tumors arising in the familial setting present unique diagnostic and therapeutic dilemmas. PMID- 15658672 TI - Pheochromocytoma. PMID- 15658674 TI - Gastroduodenal endocrine tumours. PMID- 15658677 TI - Sound equalization in a large region of a rectangular enclosure. PMID- 15658678 TI - Annoyance of bandpass-filtered noises in relation to the factor extracted from autocorrelation function. PMID- 15658679 TI - Eigenmodes of triaxial ellipsoidal acoustical cavities with mixed boundary conditions. AB - The linear acoustics problem of resonant vibrational modes in a triaxial ellipsoidal acoustic cavity with walls of arbitrary acoustic impedance has been quasi-analytically solved using the Frobenius power-series expansion method. Eigenmode results are presented for the lowest two eigenmodes in cases with pressure-release, rigid-wall, and lossy-wall boundary conditions. A mode crossing is obtained as a function of the specific acoustic impedance of the wall; the degeneracy is not symmetry related. Furthermore, the damping of the wave is found to be maximal near the crossing. PMID- 15658675 TI - An alternate approach to constructing distortion product otoacoustic emission (DPOAE) suppression tuning curves. PMID- 15658680 TI - Modeling three-dimensional elastic wave propagation in circular cylindrical structures using a finite-difference approach. AB - Wave propagation along circular cylindrical structures is important for nondestructive-testing applications and shocks in tubes. To simulate elastic wave propagation phenomena in such structures the governing equations in cylindrical coordinates are solved numerically. To reduce the required amount of computer memory and the computational time, the stress components are eliminated in the equilibrium equations. In the resulting coupled partial differential equations, in which only the three displacement components are involved, the derivatives with respect to spatial coordinates and time are approximated using second order central differences. This leads to the present new approach, which is both accurate and efficient. In order to obtain a stable scheme the displacements must be allocated on a staggered grid. The von Neumann stability analysis is performed and the result is compared with an existing empirical criterion. Mechanical energies are observed in order to validate the finite-difference code. Since no material damping or energy dissipation is taken into account in the equations of motion, the total energy must remain constant over time. Only negligible variations are observed during long-term simulations. Dispersion relations are used to check the physical behavior of the waves calculated with the proposed finite-difference method: Theoretically calculated curves are compared with values obtained by a spectrum estimation method, applied to the results of a simulation. PMID- 15658681 TI - Transient propagation in media with classical or power-law loss. AB - This paper addresses the problem of small-signal transient wave propagation in media whose absorption coefficient obeys power-law frequency dependence, i.e., alpha infinity omega n. Our approach makes use of previously derived relations between the absorption and dispersion based on the Kramers-Kronig relations. This, combined with a recently obtained solution to a causal convolution wave equation enable expressions to be obtained for one-dimensional transient propagation when n is in the range 0 < n < 3. For n = 2, corresponding to no dispersion, straightforward analytical expressions are obtained for a delta function and a sinusoidal step function sources and these are shown to correspond to relations previously derived. For other values of n, the effects of dispersion are accounted for by using Fourier transforms. Examples are used to illustrate the results for normal and anomalous dispersive media and to examine the question as to the conditions under which the effects of dispersion should be accounted for, especially for wideband ultrasound pulses of the type used in B-mode tissue imaging. It is shown that the product of the attenuation and total propagation path can be used as a criterion for judging whether dispersion needs to be accounted for. PMID- 15658682 TI - The complex equivalent source method for sound propagation over an impedance plane. AB - The sound field caused by a monopole source above an impedance plane can be calculated by using a superposition of equivalent point sources located along a line in the mirror space below the plane. Originally, such an approach for representing the half-space Green's function was described by Sommerfeld at the beginning of the last century, in order to treat half-space problems of heat conduction. However, the representation converges only for masslike impedances and cannot be used for the more important case of reflecting planes with springlike surface impedances. The singular part of the line integral can be transformed into a Hankel function, which shows that surface waves are contained in the whole solution. Unfortunately, this representation suffers from the lack of validity at certain receiver points and from restrictions on wave number and impedance range to ensure the necessary convergence. The main idea of the present method is to use also a superposition of equivalent point sources, but to allow that these sources can be located at complex source points. The corresponding form of the half-space Green's function is suitable for both masslike and springlike surface impedances, and can be used as a cornerstone for a boundary element method. PMID- 15658683 TI - Modifications of acoustic modes and coupling due to a leaning wall in a rectangular cavity. AB - Acoustic modes and the coupling characteristics of a rectangular-like cavity with a slight geometrical distortion introduced through a leaning wall are investigated in this paper. A pressure variation index is proposed to quantify the global changes in acoustic modes caused by the inclination of the wall. Effects on the coupling between acoustic modes and structural modes are investigated using coupling coefficients. Numerical results show a simple relationship between the distortion effect and the acoustic wavelength. The effect is most significant when the distortion approaches the half wavelength. Compared with a rectangular enclosure, the existence of the leaning wall gives rise to a much more effective coupling between the structure and the enclosure. PMID- 15658684 TI - Attenuation of ultrasonic waves in rolled metals. AB - Scattering of ultrasonic waves in polycrystals with texture is studied in this article. The attenuations of the three wave modes are determined as a function of dimensionless frequency and propagation direction, respectively, for given orientation distribution coefficients (ODCs). The calculation is done in the case of a statistically orthorhombic sample made up of cubic crystallites. The wave propagation and scattering model is formulated by the Dyson equation using an anisotropic Green's function approach. Within the limits of the first-order smoothing approximation, the Dyson equation is solved in the spatial Fourier transform domain. The results presented are shown to be directional dependent, frequency dependent, and especially dependent on the texture coefficients (ODCs) for the quasilongitudinal and two quasishear waves. The theoretical results presented may be used to improve the understanding of the microstructure during recrystallization processes. PMID- 15658685 TI - A theoretical study of special acoustic effects caused by the staircase of the El Castillo pyramid at the Maya ruins of Chichen-Itza in Mexico. AB - It is known that a handclap in front of the stairs of the great pyramid of Chichen Itza produces a chirp echo which sounds more or less like the sound of a Quetzal bird. The present work describes precise diffraction simulations and attempts to answer the critical question what physical effects cause the formation of the chirp echo. Comparison is made with experimental results obtained from David Lubman. Numerical simulations show that the echo shows a strong dependence on the kind of incident sound. Simulations are performed for a (delta function like) pulse and also for a real handclap. The effect of reflections on the ground in front of the pyramid is also discussed. The present work also explains why an observer seated on the lowest step of the pyramid hears the sound of raindrops falling in a water filled bucket instead of footstep sounds when people, situated higher up the pyramid, climb the stairs. PMID- 15658686 TI - On the acoustic modes in a cylindrical duct with an arbitrary wall impedance distribution. AB - The present paper considers the propagation of sound in a cylindrical duct, with a wall section of finite length covered by an acoustic liner whose impedance is an arbitrary function of position. The cases of (i) uniform wall impedance, and wall impedance varying along the (ii) circumference or (iii) axis of the duct, or (iv) both simultaneously, are explicitly considered. It is shown that a nonuniform wall impedance couples modes with distinct azimuthal l or axial m wave numbers, so that their radial wave numbers k can no longer be calculated separately for each pair (m,l). The radial wave numbers are the roots of an infinite determinant, in the case when the wall impedance varies either (i) circumferentially or (ii) radially. If the wall impedance varies (iv) both radially and circumferentially, then the radial wave numbers are the roots of a doubly infinite determinant, i.e., an infinite determinant in which each term is an infinite determinant. The infinite determinants specifying the radial wave numbers are written explicitly for sound in a cylindrical nozzle with a uniform axial flow, in which case the radial eigenfunctions are Bessel functions; the method of calculation of the radial wave numbers applies equally well to a cylindrical nozzle with shear flow and/or swirling flows, with the Bessel functions replaced by other eigenfunctions. The radial wave numbers are calculated by truncation of the infinite determinants, for several values of the aspect ratio, defined as the ratio of length to diameter. It is shown that a nonuniform wall impedance will give rise to additional modes compared with a uniform wall impedance. The radial wave numbers specify the eigenfrequencies for the acoustic modes in the duct; the imaginary parts of the eigenfrequencies specify the decay of the sound field with time, and thus the effectiveness of the acoustic liner. PMID- 15658687 TI - Wave interaction in acoustic resonators with and without hysteresis. AB - Nonlinear interaction of counterpropagating waves in solids is considered by using a general approach taking into account only the cumulative (resonant) nonlinear perturbations giving a nonzero contribution over the period and, consecutively, potentially able to significantly modify the linear solution. Different stress-strain relations are addressed, including those with hysteresis which serve as basic models for the recent acoustic experiments with rock and metals. An important case of the interaction of counterpropagating waves with close amplitudes in a high-Q resonator (bar) with hysteresis is specially addressed and compared with the case of a ring resonator. PMID- 15658688 TI - Nonlinear acoustic techniques for landmine detection. AB - Measurements of the top surface vibration of a buried (inert) VS 2.2 anti-tank plastic landmine reveal significant resonances in the frequency range between 80 and 650 Hz. Resonances from measurements of the normal component of the acoustically induced soil surface particle velocity (due to sufficient acoustic to-seismic coupling) have been used in detection schemes. Since the interface between the top plate and the soil responds nonlinearly to pressure fluctuations, characteristics of landmines, the soil, and the interface are rich in nonlinear physics and allow for a method of buried landmine detection not previously exploited. Tuning curve experiments (revealing "softening" and a back-bone curve linear in particle velocity amplitude versus frequency) help characterize the nonlinear resonant behavior of the soil-landmine oscillator. The results appear to exhibit the characteristics of nonlinear mesoscopic elastic behavior, which is explored. When two primary waves f1 and f2 drive the soil over the mine near resonance, a rich spectrum of nonlinearly generated tones is measured with a geophone on the surface over the buried landmine in agreement with Donskoy [SPIE Proc. 3392, 221-217 (1998); 3710, 239-246 (1999)]. In profiling, particular nonlinear tonals can improve the contrast ratio compared to using either primary tone in the spectrum. PMID- 15658689 TI - Multiple-order derivatives of a waveguide acoustic field with respect to sound speed, density, and frequency. AB - An adjoint perturbative method is used to derive expressions for the first- through third-order derivatives of a pressure field with respect to sound speed, density, and frequency, for the restricted case of a laterally homogenous waveguide in which environmental parameters are only a function of depth. By using a normal-mode Green's function, the three-dimensional spatial correlation required by the standard acoustic adjoint equation can be reduced to a set of one dimensional depth integrals. The resulting expressions for the first-order derivative are similar to those obtained by previous perturbative approaches based on the depth-separated wave equation, but the approach followed here permits straightforward extension to higher-order derivatives. Explicit evaluations of the expressions for a representative shallow-water waveguide model are in excellent agreement with numerical finite-difference computations. An analysis of the expressions as a function of source-receiver range finds the contributions to the mode amplitude derivatives to be non-negligible at ranges less than a few modal interference lengths, for parameters associated with the ocean bottom. Therefore, linear perturbative inversion methods that perturb only horizontal wavenumbers and not mode amplitudes should either be used with caution or modified to incorporate the expressions presented here. PMID- 15658690 TI - Development of an accelerometer-based underwater acoustic intensity sensor. AB - An underwater acoustic intensity sensor is described. This sensor derives acoustic intensity from simultaneous, co-located measurement of the acoustic pressure and one component of the acoustic particle acceleration vector. The sensor consists of a pressure transducer in the form of a hollow piezoceramic cylinder and a pair of miniature accelerometers mounted inside the cylinder. Since this sensor derives acoustic intensity from measurement of acoustic pressure and acoustic particle acceleration, it is called a p-a intensity probe. The sensor is ballasted to be nearly neutrally buoyant. It is desirable for the accelerometers to measure only the rigid body motion of the assembled probe and for the effective centers of the pressure sensor and accelerometer to be coincident. This is achieved by symmetric disposition of a pair of accelerometers inside the ceramic cylinder. The response of the intensity probe is determined by comparison with a reference hydrophone in a predominantly reactive acoustic field. PMID- 15658691 TI - Experimental demonstration of remote, passive acousto-optic sensing. AB - Passively detecting underwater sound from the air can allow aircraft and surface vessels to monitor the underwater acoustic environment. Experimental research into an optical hydrophone is being conducted for remote, aerial detection of underwater sound. A laser beam is directed onto the water surface to measure the velocity of the vibrations occurring as the underwater acoustic signal reaches the water surface. The acoustically generated surface vibrations modulate the phase of the laser beam. Sound detection occurs when the laser is reflected back towards the sensor. Therefore, laser alignment on the specularly reflecting water surface is critical. As the water surface moves, the laser beam is reflected away from the photodetector and no signal is obtained. One option to mitigate this problem is to continually steer the laser onto a spot on the water surface that provides a direct back-reflection. Results are presented from a laboratory test that investigates the feasibility of the acousto-optic sensor detection on hydrostatic and hydrodynamic surfaces using a laser Doppler vibrometer in combination with a laser-based, surface normal glint tracker for remotely detecting underwater sound. This paper outlines the acousto-optic sensor and tracker concepts and presents experimental results comparing sensor operation under various sea surface conditions. PMID- 15658692 TI - Analysis and modeling of broadband airgun data influenced by nonlinear internal waves. AB - To investigate acoustic effects of nonlinear internal waves, the two southwest tracks of the SWARM 95 experiment are considered. An airgun source produced broadband acoustic signals while a packet of large nonlinear internal waves passed between the source and two vertical linear arrays. The broadband data and its frequency range (10-180 Hz) distinguish this study from previous work. Models are developed for the internal wave environment, the geoacoustic parameters, and the airgun source signature. Parabolic equation simulations demonstrate that observed variations in intensity and wavelet time-frequency plots can be attributed to nonlinear internal waves. Empirical tests are provided of the internal wave-acoustic resonance condition that is the apparent theoretical mechanism responsible for the variations. Peaks of the effective internal wave spectrum are shown to coincide with differences in dominant acoustic wavenumbers comprising the airgun signal. The robustness of these relationships is investigated by simulations for a variety of geoacoustic and nonlinear internal wave model parameters. PMID- 15658693 TI - Bubble dynamics and size distributions during focused ultrasound insonation. AB - The deposition of ultrasonic energy in tissue can cause tissue damage due to local heating. For pressures above a critical threshold, cavitation will occur, inducing a much larger thermal energy deposition in a local region. The present work develops a nonlinear bubble dynamics model to numerically investigate bubble oscillations and bubble-enhanced heating during focused ultrasound (HIFU) insonation. The model is applied to calculate two threshold-dependent phenomena occurring for nonlinearly oscillating bubbles: Shape instability and growth by rectified diffusion. These instabilities in turn are shown to place physical boundaries on the time-dependent bubble size distribution, and thus the thermal energy deposition. PMID- 15658695 TI - An iterative effective medium approximation (IEMA) for wave dispersion and attenuation predictions in particulate composites, suspensions and emulsions. AB - In the present work we deal with the scattering dispersion and attenuation of elastic waves in different types of nonhomogeneous media. The iterative effective medium approximation based on a single scattering consideration, for the estimation of wave dispersion and attenuation, proposed in Tsinopoulos et al., [Adv. Compos. Lett. 9, 193-200 (2000)] is examined herein not only for solid components but for liquid suspensions as well. The iterations are conducted by means of the classical relation of Waterman and Truell, while the self-consistent condition proposed by Kim et al. [J. Acoust. Soc. Am. 97, 1380-1388 (1995)] is used for the convergence of the iterative procedure. The single scattering problem is solved using the Ying and Truell formulation, which with a minor modification can accommodate the solution of scattering on inclusions in liquid. Theoretical results for several different systems of particulates and suspensions are presented being in excellent agreement with experimental data taken from the literature. PMID- 15658696 TI - Insertion loss of an acoustic enclosure. AB - Acoustical enclosures are the common arrangements in reducing airborne noise from shipboard machinery such as engines and generators. In this paper the theoretical models, established based on statistical energy analysis, are presented for predicting the insertion loss of acoustical enclosures in different frequency ranges. In addition to the consideration of resonant modal coupling between internal sound field and enclosure structural vibration, the nonresonant transmission though and the interaction between enclosure walls in the models are also included. It is shown that the insertion loss of enclosures is mainly controlled by the nonresonant modes in the intermediate frequency range. At high frequencies, the insertion loss of enclosures can be improved by increasing the sound absorption at the internal boundaries of enclosures. Experiments were carried out on two enclosures made of different materials. The measured results are compared with the predicted values and the good agreement between them is the initial demonstration of the validity and feasibility of the theoretical models. PMID- 15658697 TI - Perception and annoyance due to wind turbine noise--a dose-response relationship. AB - Installed global wind power increased by 26% during 2003, with U.S and Europe accounting for 90% of the cumulative capacity. Little is known about wind turbines' impact on people living in their vicinity. The aims of this study were to evaluate the prevalence of annoyance due to wind turbine noise and to study dose-response relationships. Interrelationships between noise annoyance and sound characteristics, as well as the influence of subjective variables such as attitude and noise sensitivity, were also assessed. A cross-sectional study was performed in Sweden in 2000. Responses were obtained through questionnaires (n = 351; response rate 68.4%), and doses were calculated as A-weighted sound pressure levels for each respondent. A statistically significant dose-response relationship was found, showing higher proportion of people reporting perception and annoyance than expected from the present dose-response relationships for transportation noise. The unexpected high proportion of annoyance could be due to visual interference, influencing noise annoyance, as well as the presence of intrusive sound characteristics. The respondents' attitude to the visual impact of wind turbines on the landscape scenery was found to influence noise annoyance. PMID- 15658698 TI - The role of noise sensitivity in the noise-response relation: a comparison of three international airport studies. AB - In order to examine the role of noise sensitivity in response to environmental noise, this paper presents detailed comparisons of socio-acoustic studies conducted around international airports in Amsterdam, Sydney, and London. Earlier findings that noise sensitivity moderates the effect of noise on annoyance were examined to see if they could be replicated in each of the datasets, independent of the technique of measuring noise sensitivity. The relation between exposure to aircraft noise and noise annoyance was studied separately for groups of individuals with low, medium, and high noise sensitivity, with statistical adjustment for relevant confounders. Results support the previous findings that noise sensitivity is an independent predictor of annoyance and adds to the prediction of noise annoyance afforded by noise exposure level by up to 26% of explained variance. There is no evidence of a moderating effect, whereby the covariation between noise exposure level and annoyance is weak for people who score at the extreme high or low end of the sensitivity scale, and strong for people who score in the middle of the sensitivity scale. Generally, noise sensitivity appears to increase annoyance independently of the level of noise exposure after adjustment for relevant confounders. These findings were consistent across the three datasets. PMID- 15658699 TI - Measures for assessing architectural speech security (privacy) of closed offices and meeting rooms. AB - Objective measures were investigated as predictors of the speech security of closed offices and rooms. A new signal-to-noise type measure is shown to be a superior indicator for security than existing measures such as the Articulation Index, the Speech Intelligibility Index, the ratio of the loudness of speech to that of noise, and the A-weighted level difference of speech and noise. This new measure is a weighted sum of clipped one-third-octave-band signal-to-noise ratios; various weightings and clipping levels are explored. Listening tests had 19 subjects rate the audibility and intelligibility of 500 English sentences, filtered to simulate transmission through various wall constructions, and presented along with background noise. The results of the tests indicate that the new measure is highly correlated with sentence intelligibility scores and also with three security thresholds: the threshold of intelligibility (below which speech is unintelligible), the threshold of cadence (below which the cadence of speech is inaudible), and the threshold of audibility (below which speech is inaudible). The ratio of the loudness of speech to that of noise, and simple A weighted level differences are both shown to be well correlated with these latter two thresholds (cadence and audibility), but not well correlated with intelligibility. PMID- 15658700 TI - Robustness of spatial average equalization: a statistical reverberation model approach. AB - Traditionally, multiple listener room equalization is performed to improve sound quality at all listeners, during audio playback, in a multiple listener environment (e.g., movie theaters, automobiles, etc.). A typical way of doing multiple listener equalization is through spatial averaging, where the room responses are averaged spatially between positions and an inverse equalization filter is found from the spatially averaged result. However, the equalization performance, will be affected if there is a mismatch between the position of the microphones (which are used for measuring the room responses for designing the equalization filter) and the actual center of listener head position (during playback). In this paper, we will present results on the effects of microphone listener mismatch on spatial average equalization performance. The results indicate that, for the analyzed rectangular configuration, the region of effective equalization depends on (i) the distance of a listener from the source, (ii) the amount of mismatch between the responses, and (iii) the frequency of the audio signal. We also present some convergence analysis to interpret the results. PMID- 15658701 TI - Measurement of in-duct acoustic properties by using a single microphone with fixed position. AB - Acoustic properties of sound absorption materials and other acoustic structures can be measured in an impedance tube using the well-established two-microphone method to resolve the two traveling wave components of a standing wave pattern. The accuracy of such measurements depends crucially on the calibration of the two microphones placed in close proximity. To eliminate such calibration, the one microphone method [Chu, J. Acoust. Soc. Am. 80, 555-560 (1986)] uses the same microphone to probe at two positions sequentially using the voltage driving the loudspeaker as a reference signal. A variant of this method is introduced in this study in which the microphone is fixed at one position while a rigid end plate moves between two positions to resolve the standing wave. The sound source is installed as a side branch, and its driving signal is also used as a reference in the two-step measurement. Close agreement is found with the established two microphone method, and factors which might affect the accuracy of the new technique are discussed. As a demonstration of the robustness of the method, a low-budget electret microphone is used and the result also matches well with those obtained by the two-microphone method with high-quality condenser type microphones. PMID- 15658702 TI - Investigation of the validity of radiosity for sound-field prediction in cubic rooms. AB - This paper explores acoustical (or time-dependent) radiosity using predictions made in four cubic enclosures. The methods and algorithms used are those presented in a previous paper by the same authors [Nosal, Hodgson, and Ashdown, J. Acoust. Soc. Am. 116(2), 970-980 (2004)]. First, the algorithm, methods, and conditions for convergence are investigated by comparison of numerous predictions for the four cubic enclosures. Here, variables and parameters used in the predictions are varied to explore the effect of absorption distribution, the necessary conditions for convergence of the numerical solution to the analytical solution, form-factor prediction methods, and the computational requirements. The predictions are also used to investigate the effect of absorption distribution on sound fields in cubic enclosures with diffusely reflecting boundaries. Acoustical radiosity is then compared to predictions made in the four enclosures by a ray tracing model that can account for diffuse reflection. Comparisons are made of echograms, room-acoustical parameters, and discretized echograms. PMID- 15658703 TI - On the kinematics of broadband multipath scintillation and the approach to saturation. AB - Utilizing a simple model in which the acoustic wave function is a sum of independent Gaussian wave packets, the relative intensity variance or scintillation index (SI) is analytically calculated. The model has an unspecified probability density function (PDF) for wave packet amplitudes and Gaussian PDFs for travel-time-induced and non-travel-time-induced phase shifts; amplitudes and both phase shifts are assumed to be mutually uncorrelated. It is shown that a proper treatment of the mean field is required to obtain the saturation value, SI = 1, in the limit of a large number of interfering wave packets. The analytic formulas for SI allow identification of important wave packet parameters in the approach to saturation. Criteria are identified for both broadband and narrow band cases for which the approach to saturation is from above and below 1. It is demonstrated that the broadband approach to saturation is much slower than the narrow-band cases, since wave packets separated in time by more than an inverse bandwidth do not strongly contribute to interference. This effect is quantified by the time-bandwidth product. The model is also used to obtain an analytic expression for pulse time spread; it is shown that multipath conditions which favor a rapid approach to saturation do not favor large pulse spread. PMID- 15658704 TI - Development of a laser photothermoacoustic frequency-swept system for subsurface imaging: theory and experiment. AB - In conventional biomedical photoacoustic imaging systems, a pulsed laser is used to generate time-of-flight acoustic information of the subsurface features. This paper reports the theoretical and experimental development of a new frequency domain (FD) photo-thermo-acoustic (PTA) principle featuring frequency sweep (chirp) and heterodyne modulation and lock-in detection of a continuous-wave laser source at 1064 nm wavelength. PTA imaging is a promising new technique which is being developed to detect tumor masses in turbid biological tissue. Owing to the linear relationship between the depth of acoustic signal generation and the delay time of signal arrival to the transducer, information specific to a particular depth can be associated with a particular frequency in the chirp signal. Scanning laser modulation with a linear frequency sweep method preserves the depth-to-delay time linearity and recovers FD-PTA signals from a range of depths. Preliminary results performed on rubber samples and solid tissue phantoms indicate that the FD-PTA technique has the potential to be a reliable tool for biomedical depth-profilometric imaging. PMID- 15658705 TI - The temporal representation of speech in a nonlinear model of the guinea pig cochlea. AB - The temporal representation of speechlike stimuli in the auditory-nerve output of a guinea pig cochlea model is described. The model consists of a bank of dual resonance nonlinear filters that simulate the vibratory response of the basilar membrane followed by a model of the inner hair cell/auditory nerve complex. The model is evaluated by comparing its output with published physiological auditory nerve data in response to single and double vowels. The evaluation includes analyses of individual fibers, as well as ensemble responses over a wide range of best frequencies. In all cases the model response closely follows the patterns in the physiological data, particularly the tendency for the temporal firing pattern of each fiber to represent the frequency of a nearby formant of the speech sound. In the model this behavior is largely a consequence of filter shapes; nonlinear filtering has only a small contribution at low frequencies. The guinea pig cochlear model produces a useful simulation of the measured physiological response to simple speech sounds and is therefore suitable for use in more advanced applications including attempts to generalize these principles to the response of human auditory system, both normal and impaired. PMID- 15658706 TI - Age effects in the human middle ear: wideband acoustical measures. AB - Studies that have examined age effects in the human middle ear using either admittance measures at 220 or 660 Hz or multifrequency tympanometry from 200 to 2000 Hz have had conflicting results. Several studies have suggested an increase in admittance with age, while several others have suggested a decrease in admittance with age. A third group of studies found no significant age effect. This study examined 226 Hz tympanometry and wideband energy reflectance and impedance at ambient pressure in a group of 40 young adults and a group of 30 adults with age > or = 60 years. The groups did not differ in admittance measures of the middle ear at 226 Hz. However, significant age effects were found in wideband energy reflectance and impedance. In particular, in older adults there was a comparative decrease in reflectance from 800 to 2000 Hz but an increase near 4000 Hz. The results suggest a decrease in middle-ear stiffness with age. The findings of this study hold relevance for understanding the aging process in the auditory system, for the establishment of normative data for wideband energy reflectance, for the possibility of a conductive component to presbycusis, and for the interpretation of otoacoustic emission measurements. PMID- 15658707 TI - Cochlear compression: effects of low-frequency biasing on quadratic distortion product otoacoustic emission. AB - Distortion product otoacoustic emissions (DPOAEs) are generated from the nonlinear transduction n cochlear outer hair cells. The transducer function demonstrating a compressive nonlinearity can be estimated from low-frequency modulation of DPOAEs. Experimental results from the gerbils showed that the magnitude of quadratic difference tone (QDT, f2-f1) was either enhanced or suppressed depending on the phase of the low-frequency bias tone. Within one period of the bias tone, QDT magnitudes exhibited two similar modulation patterns, each resembling the absolute value of the second derivative of the transducer function. In the time domain, the center notches of the modulation patterns occurred around the zero crossings of the bias pressure, whereas peaks corresponded to the increase or decrease in bias pressure. Evaluated with respect to the bias pressure, modulated QDT magnitude displayed a double-modulation pattern marked by a separation of the center notches. Loading/unloading of the cochlear transducer or rise/fall in bias pressure shifted the center notch to positive or negative sound pressures, indicating a mechanical hysteresis. These results suggest that QDT arises from the compression that coexists with the active hysteresis in cochlear transduction. Modulation of QDT magnitude reflects the dynamic regulation of cochlear transducer gain and compression. PMID- 15658708 TI - Distortion product otoacoustic emission (2f1-f2) suppression in 3-month-old infants: evidence for postnatal maturation of human cochlear function? AB - The complete timeline for maturation of human cochlear function has not been defined. Distortion product otoacoustic emission (DPOAE)-based measures of cochlear function show non-adult-like responses from premature and term-born neonates at high f2 frequencies; however, older infants were not included in these studies. In the present experiment, previously collected DPOAE ipsilateral suppression data from premature neonates were combined with new data collected from adults, term-born neonates, and 3-month-old infants to further examine the time course for maturation of cochlear function. DPOAE suppression tuning curves (STC) and suppression growth patterns were measured in the three age groups at f2 = 6000 Hz, L1 = 65, L2 = 55 dB SPL, with an f2/f1 of 1.2. Results indicate that term-born neonates and 3-month-old infants have non-adult-like STC width, slope on the low-frequency flank, and tip features. However, the two infant groups are not significantly different from one another. Suppression growth patterns for low frequency suppressor tones show a clear developmental progression. In general, the younger the infant, the more shallow and compressive the suppression growth for the lowest suppressor frequencies. These findings suggest a high-frequency postnatal immaturity in cochlear function as measured by DPOAE suppression. Results may have been influenced by noncochlear factors, such as middle-ear immaturity. These factors are reviewed and considered. PMID- 15658709 TI - Human temporal auditory acuity as assessed by envelope following responses. AB - Temporal auditory acuity, the ability to discriminate rapid changes in the envelope of a sound, is essential for speech comprehension. Human envelope following responses (EFRs) recorded from scalp electrodes were evaluated as an objective measurement of temporal processing in the auditory nervous system. The temporal auditory acuity of older and younger participants was measured behaviorally using both gap and modulation detection tasks. These findings were then related to EFRs evoked by white noise that was amplitude modulated (25% modulation depth) with a sweep of modulation frequencies from 20 to 600 Hz. The frequency at which the EFR was no longer detectable was significantly correlated with behavioral measurements of gap detection (r = -0.43), and with the maximum perceptible modulation frequency (r = 0.72). The EFR techniques investigated here might be developed into a clinically useful objective estimate of temporal auditory acuity for subjects who cannot provide reliable behavioral responses. PMID- 15658710 TI - The bat head-related transfer function reveals binaural cues for sound localization in azimuth and elevation. AB - Directional properties of the sound transformation at the ear of four intact echolocating bats, Eptesicus fuscus, were investigated via measurements of the head-related transfer function (HRTF). Contributions of external ear structures to directional features of the transfer functions were examined by remeasuring the HRTF in the absence of the pinna and tragus. The investigation mainly focused on the interactions between the spatial and the spectral features in the bat HRTF. The pinna provides gain and shapes these features over a large frequency band (20-90 kHz), and the tragus contributes gain and directionality at the high frequencies (60 to 90 kHz). Analysis of the spatial and spectral characteristics of the bat HRTF reveals that both interaural level differences (ILD) and monaural spectral features are subject to changes in sound source azimuth and elevation. Consequently, localization cues for horizontal and vertical components of the sound source location interact. Availability of multiple cues about sound source azimuth and elevation should enhance information to support reliable sound localization. These findings stress the importance of the acoustic information received at the two ears for sound localization of sonar target position in both azimuth and elevation. PMID- 15658711 TI - Relative contributions of temporal and place pitch cues to fundamental frequency discrimination in cochlear implantees. AB - The effect of the filter bank on fundamental frequency (F0) discrimination was examined in four Nucleus CI24 cochlear implant subjects for synthetic stylized vowel-like stimuli. The four tested filter banks differed in cutoff frequencies, amount of overlap between filters, and shape of the filters. To assess the effects of temporal pitch cues on F0 discrimination, temporal fluctuations were removed above 10 Hz in one condition and above 200 Hz in another. Results indicate that F0 discrimination based upon place pitch cues is possible, but just noticeable differences exceed 1 octave or more depending on the filter bank used. Increasing the frequency resolution in the F0 range improves the F0 discrimination based upon place pitch cues. The results of F0 discrimination based upon place pitch agree with a model that compares the centroids of the electrical excitation pattern. The addition of temporal fluctuations up to 200 Hz significantly improves F0 discrimination. Just-noticeable differences using both place and temporal pitch cues range from 6% to 60%. Filter banks that do not resolve the higher harmonics provided the best temporal pitch cues, because temporal pitch cues are clearest when the fluctuation on all channels is at F0 and preferably in phase. PMID- 15658712 TI - Methodological aspects of an adaptive multidirectional pattern search to optimize speech perception using three hearing-aid algorithms. AB - In this study we investigated the reliability and convergence characteristics of an adaptive multidirectional pattern search procedure, relative to a nonadaptive multidirectional pattern search procedure. The procedure was designed to optimize three speech-processing strategies. These comprise noise reduction, spectral enhancement, and spectral lift. The search is based on a paired-comparison paradigm, in which subjects evaluated the listening comfort of speech-in-noise fragments. The procedural and nonprocedural factors that influence the reliability and convergence of the procedure are studied using various test conditions. The test conditions combine different tests, initial settings, background noise types, and step size configurations. Seven normal hearing subjects participated in this study. The results indicate that the reliability of the optimization strategy may benefit from the use of an adaptive step size. Decreasing the step size increases accuracy, while increasing the step size can be beneficial to create clear perceptual differences in the comparisons. The reliability also depends on starting point, stop criterion, step size constraints, background noise, algorithms used, as well as the presence of drifting cues and suboptimal settings. There appears to be a trade-off between reliability and convergence, i.e., when the step size is enlarged the reliability improves, but the convergence deteriorates. PMID- 15658713 TI - Auditory processing of real and illusory changes in frequency modulation (FM) phase. AB - Auditory processing of frequency modulation (FM) was explored. In experiment 1, detection of a tau-radians modulator phase shift deteriorated as modulation rate increased from 2.5 to 20 Hz, for 1- and 6-kHz carriers. In experiment 2, listeners discriminated between two 1-kHz carriers, where, mid-way through, the 10-Hz frequency modulator had either a phase shift or increased in depth by deltaD% for half a modulator period. Discrimination was poorer for deltaD = 4% than for smaller or larger increases. These results are consistent with instantaneous frequency being smoothed by a time window with a total duration of about 110 ms. In experiment 3, the central 200-ms of a 1-s 1-kHz carrier modulated at 5 Hz was replaced by noise, or by a faster FM applied to a more intense 1-kHz carrier. Listeners heard the 5-Hz FM continue at the same depth throughout the stimulus. Experiments 4 and 5 showed that, after an FM tone had been interrupted by a 200-ms noise, listeners were insensitive to the phase at which the FM resumed. It is argued that the auditory system explicitly encodes the presence, and possibly the rate and depth, of FM in a way that does not preserve information on FM phase. PMID- 15658714 TI - Direct measurement of onset and offset phonation threshold pressure in normal subjects. AB - Phonation threshold pressures were directly measured in five normal subjects in a variety of voicing conditions. The effects of fundamental frequency, intensity, closure speed of the vocal folds, and laryngeal airway resistance on phonation threshold pressures were determined. Subglottic air pressures were measured using percutaneous puncture of the cricothyroid membrane. Both onset and offset of phonation were studied to see if a hysteresis effect produced lower offset pressures than onset pressures. Univariate analysis showed that phonation threshold pressure was influenced most strongly by fundamental frequency and intensity. Multiple linear regression showed that these two variables, as well as laryngeal airway resistance, most strongly predicted phonation threshold pressure. Two of the five subjects demonstrated a significant hysteresis effect, but one subject actually had higher offset pressures than onset pressures. PMID- 15658715 TI - Rapid adaptation to foreign-accented English. AB - This study explored the perceptual benefits of brief exposure to non-native speech. Native English listeners were exposed to English sentences produced by non-native speakers. Perceptual processing speed was tracked by measuring reaction times to visual probe words following each sentence. Three experiments using Spanish- and Chinese-accented speech indicate that processing speed is initially slower for accented speech than for native speech but that this deficit diminishes within one minute of exposure. Control conditions rule out explanations for the adaptation effect based on practice with the task and general strategies for dealing with difficult speech. Further results suggest that adaptation can occur within as few as two to four sentence-length utterances. The findings emphasize the flexibility of human speech processing and require models of spoken word recognition that can rapidly accommodate significant acoustic-phonetic deviations from native language speech patterns. PMID- 15658716 TI - Enhancing Chinese tone recognition by manipulating amplitude envelope: implications for cochlear implants. AB - Tone recognition is important for speech understanding in tonal languages such as Mandarin Chinese. Cochlear implant patients are able to perceive some tonal information by using temporal cues such as periodicity-related amplitude fluctuations and similarities between the fundamental frequency (F0) contour and the amplitude envelope. The present study investigates whether modifying the amplitude envelope to better resemble the F0 contour can further improve tone recognition in multichannel cochlear implants. Chinese tone and vowel recognition were measured for six native Chinese normal-hearing subjects listening to a simulation of a four-channel cochlear implant speech processor with and without amplitude envelope enhancement. Two algorithms were proposed to modify the amplitude envelope to more closely resemble the F0 contour. In the first algorithm, the amplitude envelope as well as the modulation depth of periodicity fluctuations was adjusted for each spectral channel. In the second algorithm, the overall amplitude envelope was adjusted before multichannel speech processing, thus reducing any local distortions to the speech spectral envelope. The results showed that both algorithms significantly improved Chinese tone recognition. By adjusting the overall amplitude envelope to match the F0 contour before multichannel processing, vowel recognition was better preserved and less speech processing computation was required. The results suggest that modifying the amplitude envelope to more closely resemble the F0 contour may be a useful approach toward improving Chinese-speaking cochlear implant patients' tone recognition. PMID- 15658717 TI - Patterns of English phoneme confusions by native and non-native listeners. AB - Native American English and non-native (Dutch) listeners identified either the consonant or the vowel in all possible American English CV and VC syllables. The syllables were embedded in multispeaker babble at three signal-to-noise ratios (0, 8, and 16 dB). The phoneme identification performance of the non-native listeners was less accurate than that of the native listeners. All listeners were adversely affected by noise. With these isolated syllables, initial segments were harder to identify than final segments. Crucially, the effects of language background and noise did not interact; the performance asymmetry between the native and non-native groups was not significantly different across signal-to noise ratios. It is concluded that the frequently reported disproportionate difficulty of non-native listening under disadvantageous conditions is not due to a disproportionate increase in phoneme misidentifications. PMID- 15658719 TI - Robust and accurate fundamental frequency estimation based on dominant harmonic components. AB - This paper presents a new method for robust and accurate fundamental frequency (F0) estimation in the presence of background noise and spectral distortion. Degree of dominance and dominance spectrum are defined based on instantaneous frequencies. The degree of dominance allows one to evaluate the magnitude of individual harmonic components of the speech signals relative to background noise while reducing the influence of spectral distortion. The fundamental frequency is more accurately estimated from reliable harmonic components which are easy to select given the dominance spectra. Experiments are performed using white and babble background noise with and without spectral distortion as produced by a SRAEN filter. The results show that the present method is better than previously reported methods in terms of both gross and fine F0 errors. PMID- 15658718 TI - Analysis of speech-based Speech Transmission Index methods with implications for nonlinear operations. AB - The Speech Transmission Index (STI) is a physical metric that is well correlated with the intelligibility of speech degraded by additive noise and reverberation. The traditional STI uses modulated noise as a probe signal and is valid for assessing degradations that result from linear operations on the speech signal. Researchers have attempted to extend the STI to predict the intelligibility of nonlinearly processed speech by proposing variations that use speech as a probe signal. This work considers four previously proposed speech-based STI methods and four novel methods, studied under conditions of additive noise, reverberation, and two nonlinear operations (envelope thresholding and spectral subtraction). Analyzing intermediate metrics in the STI calculation reveals why some methods fail for nonlinear operations. Results indicate that none of the previously proposed methods is adequate for all of the conditions considered, while four proposed methods produce qualitatively reasonable results and warrant further study. The discussion considers the relevance of this work to predicting the intelligibility of cochlear-implant processed speech. PMID- 15658720 TI - A numerical study of the role of the tragus in the big brown bat. AB - A comprehensive characterization of the spatial sensitivity of an outer ear from a big brown bat (Eptesicus fuscus) has been obtained using numerical methods and visualization techniques. Pinna shape information was acquired through x-ray microtomography. It was used to set up a finite-element model of diffraction from which directivities were predicted by virtue of forward wave-field projections based on a Kirchhoff integral formulation. Digital shape manipulation was used to study the role of the tragus in detailed numerical experiments. The relative position between tragus and pinna aperture was found to control the strength of an extensive asymmetric sidelobe which points in a frequency-dependent direction. An upright tragus position resulted in the strongest sidelobe sensitivity. Using a bootstrap validation paradigm, the results were found to be robust against small perturbations of the finite-element mesh boundaries. Furthermore, it was established that a major aspect of the tragus effect (position dependence) can be studied in a simple shape model, an obliquely truncated horn augmented by a flap representing the tragus. In the simulated wave field around the outer-ear structure, strong correlates of the tragus rotation were identified, which provide a direct link to the underlying physical mechanism. PMID- 15658721 TI - Distortion product otoacoustic emissions provide clues hearing mechanisms in the frog ear. AB - 2f1-f2 and 2 f2-f1 distortion product otoacoustic emissions (DPOAEs) were recorded from both ears of male and female Rana pipiens pipiens and Rana catesbeiana. The input-output (I/O) curves obtained from the amphibian papilla (AP) of both frog species are analogous to I/O curves recorded from mammals suggesting that, similarly to the mammalian cochlea, there may be an amplification process present in the frog AP. DPOAE level dependence on L1-L2 is different from that in mammals and consistent with intermodulation distortion expectations. Therefore, if a mechanical structure in the frog inner ear is functioning analogously to the mammalian basilar membrane, it must be more broadly tuned. DPOAE audiograms were obtained for primary frequencies spanning the animals' hearing range and selected stimulus levels. The results confirm that DPOAEs are produced in both papillae, with R. catesbeiana producing stronger emissions than R. p. pipiens. Consistent with previously reported sexual dimorphism in the mammalian and anuran auditory systems, females of both species produce stronger emissions than males. Moreover, it appears that 2 f1-f2 in the frog is generated primarily at the DPOAE frequency place, while 2 f2-f1 is generated primarily at a frequency place around the primaries. Regardless of generation place, both emissions within the AP may be subject to the same filtering mechanism, possibly the tectorial membrane. PMID- 15658722 TI - Habitat-dependent ambient noise: consistent spectral profiles in two African forest types. AB - Many animal species use acoustic signals to attract mates, to defend territories, or to convey information that may contribute to their fitness in other ways. However, the natural environment is usually filled with competing sounds. Therefore, if ambient noise conditions are relatively constant, acoustic interference can drive evolutionary changes in animal signals. Furthermore, masking noise may cause acoustic divergence between populations of the same species if noise conditions differ consistently among habitats. In this study, ambient noise was sampled in a replicate set of sites in two habitat types in Cameroon: contiguous rainforest and ecotone forest patches north of the rainforest. The noise characteristics of the two forest types show significant and consistent differences. Multiple samples taken at two rainforest sites in different seasons vary little and remain distinct from those in ecotone forest. The rainforest recordings show many distinctive frequency bands, with a general increase in amplitude from low to high frequencies. Ecotone forest only shows a distinctive high-frequency band at some parts of the day. Habitat-dependent abiotic and biotic sound sources and to some extent habitat-dependent sound transmission are the likely causes of these habitat-dependent noise spectra. PMID- 15658723 TI - Measuring of viscoelastic properties of homogeneous soft solid using transient elastography: an inverse problem approach. AB - Two main questions are at the center of this paper. The first one concerns the choice of a rheological model in the frequency range of transient elastography, sonoelasticity or NMR elastography for soft solids (20-1000 Hz). Transient elastography experiments based on plane shear waves that propagate in an Agar gelatin phantom or in bovine muscles enable one to quantify their viscoelastic properties. The comparison of these experimental results to the prediction of the two simplest rheological models indicate clearly that Voigt's model is the better. The second question studied in the paper deals with the feasibility of quantitative viscosity mapping using inverse problem algorithm. In the ideal situation where plane shear waves propagate in a sample, a simple inverse problem based on the Helmholtz equation correctly retrieves both elasticity and viscosity. In a more realistic situation with nonplane shear waves, this simple approach fails. Nevertheless, it is shown that quantitative viscosity mapping is still possible if one uses an appropriate inverse problem that fully takes into account diffraction in solids. PMID- 15658724 TI - Theoretical gas body pulsation in relation to empirical gas-body destabilization and to cell membrane damage thresholds. AB - Contrast agent gas bodies attached to phagocytic monolayer cells pulsate in response to ultrasound exposure and damage the cells above thresholds, which increase in proportion to frequency. This study considered the physical basis for the thresholds and their frequency dependence. Theory for the pulsation was evaluated using empirical pulse waveforms acquired at thresholds for 1.0, 2.25, 3.5, 5.0, 7.5, and 10 MHz. For optimum-sized gas bodies, the amplitudes calculated at the thresholds were about 11% of the initial radii. At the cell membrane damage thresholds, theoretical negative shell stresses were approximately constant with frequency at about 50 MPa. This stress appears to be sufficient to induce failure of the shell, and gas body destabilization was confirmed by increases in transmission of ultrasound pulses through the monolayer and by microscopically-observed shrinkage of the gas bodies. A model of acoustic microstreaming was used to calculate the shear stress during the pulses. The maximum shear stress increased from about 1500 to 4500 Pa from 1 to 10 MHz, sufficient for the cell membrane damage. This theoretical analysis shows that both the gas body destabilization and the cell membrane damage could be expected at similar peak rarefactional pressure amplitudes, with thresholds having the observed proportionality to frequency. PMID- 15658725 TI - Vocal tract filtering and the "coo" of doves. AB - Ring doves (Streptopelia risoria) produce a "coo" vocalization that is essentially a pure-tone sound at a frequency of about 600 Hz and with a duration of about 1.5 s. While making this vocalization, the dove inflates the upper part of its esophagus to form a thin-walled sac structure that radiates sound to the surroundings. It is a reasonable assumption that the combined influence of the trachea, glottis and inflated upper esophagus acts as an effective band-pass filter to eliminate higher harmonics generated by the vibrating syringeal valve. Calculations reported here indicate that this is indeed the case. The tracheal tube, terminated by a glottal constriction, is the initial resonant structure, and subsequent resonant filtering takes place through the action of the inflated esophageal sac. The inflated esophagus proves to be a more efficient sound radiating mechanism than an open beak. The action of this sac is only moderately affected by the degree of inflation, although an uninflated esophagus is inactive as a sound radiator. These conclusions are supported by measurements and observations that have been reported in a companion paper. PMID- 15658726 TI - Geographic variations in the whistles of spinner dolphins (Stenella longirostris) of the Main Hawai'ian Islands. AB - Geographic variations in the whistles of Hawai'ian spinner dolphins are discussed by comparing 27 spinner dolphin pods recorded in waters off the Islands of Kaua'i, O'ahu, Lana'i, and Hawai'i. Three different behavioral states, the number of dolphins observed in each pod, and ten parameters extracted from each whistle contour were considered by using clustering and discriminant function analyses. The results suggest that spinner dolphin pods in the Main Hawai'ian Islands share characteristics in approximately 48% of their whistles. Spinner dolphin pods had similar whistle parameters regardless of the island, location, and date when they were sampled and the dolphins' behavioral state and pod size. The term "whistle specific subgroup" (WSS) was used to designate whistle groups with similar whistles parameters (which could have been produced in part by the same dolphins). The emission rate of whistles was higher when spinner dolphins were socializing than when they were traveling or resting, suggesting that whistles are mainly used during close-range interactions. Spinner dolphins also seem to vary whistle duration according to their general behavioral state. Whistle duration and the number of turns and steps of a whistle may be more important in delivering information at the individual level than whistle frequency parameters. PMID- 15658727 TI - Within-individual variation in bullfrog vocalizations: implications for a vocally mediated social recognition system. AB - Acoustic signals provide a basis for social recognition in a wide range of animals. Few studies, however, have attempted to relate the patterns of individual variation in signals to behavioral discrimination thresholds used by receivers to discriminate among individuals. North American bullfrogs (Rana catesbeiana) discriminate among familiar and unfamiliar individuals based on individual variation in advertisement calls. The sources, patterns, and magnitudes of variation in eight acoustic properties of multiple-note advertisement calls were examined to understand how patterns of within-individual variation might either constrain, or provide additional cues for, vocal recognition. Six of eight acoustic properties exhibited significant note-to-note variation within multiple-note calls. Despite this source of within-individual variation, all call properties varied significantly among individuals, and multivariate analyses indicated that call notes were individually distinct. Fine temporal and spectral call properties exhibited less within-individual variation compared to gross-temporal properties and contributed most toward statistically distinguishing among individuals. Among-individual differences in the patterns of within-individual variation in some properties suggest that within-individual variation could also function as a recognition cue. The distributions of among individual and within-individual differences were used to generate hypotheses about the expected behavioral discrimination thresholds of receivers. PMID- 15658728 TI - An echolocation model for the restoration of an acoustic image from a single emission echo. AB - Bats can form a fine acoustic image of an object using frequency-modulated echolocation sound. The acoustic image is an impulse response, known as a reflected-intensity distribution, which is composed of amplitude and phase spectra over a range of frequencies. However, bats detect only the amplitude spectrum due to the low-time resolution of their peripheral auditory system, and the frequency range of emission is restricted. It is therefore necessary to restore the acoustic image from limited information. The amplitude spectrum varies with the changes in the configuration of the reflected-intensity distribution, while the phase spectrum varies with the changes in its configuration and location. Here, by introducing some reasonable constraints, a method is proposed for restoring an acoustic image from the echo. The configuration is extrapolated from the amplitude spectrum of the restricted frequency range by using the continuity condition of the amplitude spectrum at the minimum frequency of the emission and the minimum phase condition. The determination of the location requires extracting the amplitude spectra, which vary with its location. For this purpose, the Gaussian chirplets with a carrier frequency compatible with bat emission sweep rates were used. The location is estimated from the temporal changes of the amplitude spectra. PMID- 15658729 TI - Noise emission during the first powerboat race in an Alpine lake and potential impact on fish communities. AB - In order to assess the effects of high-speed boating on fish communities, noise levels were measured during the first Class 1 powerboat race on the Austrian Lake Traunsee. The noise spectra were compared to natural ambient noise and hearing abilities of four native fish species. Sound pressure levels (SPLs) were significantly elevated during the training heats and the race compared with natural levels, reaching up to 128 dB re 1 microPa (instantaneous SPL) at a distance of 300 m to the powerboats. Continuous equivalent SPLs were significantly lower during training and the pole position race compared to the race itself because fewer boats were simultaneously on the lake. The hearing abilities of the native hearing specialists and generalists were investigated. While carp and roach (two cyprinids) showed enhanced auditory sensitivity typical for hearing specialists, perch and whitefish were much less sensitive to sounds. Comparisons between power boat noise spectra and audiograms showed that the cyprinids can detect the boats up to several hundred meters distance because the main noise energy is well within the most sensitive hearing range. The hearing generalists, however, probably only perceive the first harmonic of the boat noise at close distances. PMID- 15658730 TI - Impotence: my historical perspective. PMID- 15658731 TI - Making tough decisions. PMID- 15658732 TI - Sexually transmitted diseases: an update. AB - Sexually transmitted diseases (STDs) have reached epidemic numbers in this country and represent considerable costs to the health care system. Nurses, who see clients in a variety of acute and community settings, have a unique opportunity to discuss STDs and their prevention. However, nurses must retain knowledge that is up-to-date on each of these diseases. Nurses need to take the lead in evaluating their clients' risk of acquiring STDs and tailor specific preventative techniques to the individual needs uncovered. PMID- 15658733 TI - Management of urinary incontinence following radical prostatectomy. AB - INTRODUCTION: The impact of a urology oncology nurse specialist in the practice environment to assist with both pre and postoperative education needs of men undergoing genitourinary surgery for cancer was evaluated. OBJECTIVE: This study aimed to improve the preoperative process and teaching for men planning surgery for genitourinary cancers, including the delivery of pathology results. A secondary aim was to decrease the incidence of postoperative incontinence. METHODS: Male patients undergoing a radical prostatectomy were recruited. Patients who underwent surgery were evaluated both before and after the urology oncology nurse specialist role was established. Followup was via phone call interview and International Prostate Symptom Score questionnaire. RESULTS: Men who received the personalized education from the urology oncology nurse specialist had a higher return of continence postoperatively, as well as higher satisfaction scores with the outcome of their surgery, including higher quality of life scores. CONCLUSION: The urology oncology nurse specialist role had a significant impact on the postoperative incontinence rate and provided a missing patient education piece that improved the overall patient satisfaction rate with oncology-related care. PMID- 15658734 TI - Patient evaluation of a discharge program following a radical prostatectomy. AB - PURPOSE: The immediate postoperative period following radical retropubic prostatectomy (RP) can be stressful, especially if men perceive discharge teaching to be incomplete. In this study, using two telephone interviews, patient satisfaction with a revised teaching program on one urology unit was evaluated. METHODS: Men undergoing a RP were given an information letter about the study at the pre-admission clinic. At days 2 and 30 post discharge, they were asked to rate the value of the call, a patient education booklet, a discharge pack of incontinence supplies, and overall satisfaction with the discharge experience. They were also asked about the use of community resources after discharge (general practitioner, urologist, emergency, home care). RESULTS: One hundred men participated. Overall, they were very satisfied with the discharge teaching and felt it provided them necessary information for recovery. However, 25 subjects used community resources inappropriately, due to concerns about wound care, catheter care, or urine retention. CONCLUSION: Although men were satisfied with the information provided in discharge program, their overuse of community resources indicates additional reinforcement is needed for management in the first month after surgery. PMID- 15658735 TI - Pelvic floor exercises for treating post-micturition dribble in men with erectile dysfunction: a randomized controlled trial. AB - PURPOSE: The purpose of this trial was to compare the efficacy of pelvic floor muscle exercises and manometric biofeedback for post-micturition dribble (PMD) in men with erectile dysfunction. METHODS: Fifty-five men with erectile dysfunction (median age 59.2 years; range 22-78) were enrolled from a local urology clinic. Twenty-eight subjects were randomized to an intervention group and received pelvic floor muscle exercises including a strong post-void "squeeze out" pelvic floor muscle contraction, biofeedback, and suggestions for lifestyle changes. Twenty-seven control subjects were solely advised on lifestyle changes. The PMD status, anal manometry, and digital anal muscle grade were assessed at baseline, 3, and 6 months. After 3 months, the control group received the intervention with pelvic floor muscle exercises and biofeedback. Both groups were followed for an additional 3 months of home exercises. An independent assessor who was blinded to the grouping assessed the PMD status of subjects at 3 and 6 months. RESULTS: Thirty-six (65.5%) of the 55 subjects reported PMD at baseline. At 3 months, there was significant reduction in PMD after intervention (p=0.001) compared to the control subjects (p=0.102). In both groups combined after 3 months of pelvic floor muscle exercises and 3 months of home exercises, 27 (75%) subjects became asymptomatic of PMD, 3 (8.3%) improved, 5 (13.9%) dropped out, and 1 (2.8%) subject still reported PMD. PMD was not correlated to age, erectile function, anal manometric pressure, or digital anal muscle grade. CONCLUSION: Pelvic floor muscle exercises including a post-void "squeeze out" pelvic floor muscle contraction are an effective treatment for post-micturition dribble in men with erectile dysfunction. PMID- 15658736 TI - Report of a mail survey of women with bladder control disorders. AB - INTRODUCTION: Very little is known about a patients perception of the severity of incontinence, absorbent product use, and interaction with health care professionals despite the growing prevalence of bladder control disorders such as urinary incontinence (UI) and overactive bladder. PURPOSE: Using a simple mail-in questionnaire, this survey was meant to evaluate the limitations in intervention for UI in women due to communication shorffalls. It cites common resources utilized once a woman finds significance to her bladder control problem. The survey explores the adequacy of intervention relative to the effective availability of information, materials, and treatment. The objective was to understand the current patient-professional relationship in order to evaluate different vehicles that might effectively increase communication regarding incontinence issues. METHODS: In partnership with a mail order medical product company, the author formulated a questionnaire that was mailed by the company to 1,500 women. Half of the sample was derived from randomly selected "leads" from advertising sources and personal referrals who had identified themselves as incontinent but had never purchased incontinence supplies from them. The other half were randomly selected customers who had purchased the company's incontinence products. The 33-item multiple-choice questionnaire had 422 responses; of these, 405 women, aged 56 and older were evaluated. A research firm that conducts and analyzes consumer surveys using standard consumer research protocol tabulated results. FINDINGS: Of those suffering from UI, two groups emerged. The first felt that incontinence was related to aging, and were not embarrassed to discuss it with their physician. They had mixed feelings regarding impact on quality of life. The second group was reluctant to seek professional advice and was uncomfortable speaking about it. They felt strongly about its negative impact on daily life. Respondents from both groups who seek professional assistance still desired more information about the causes, treatments, and coping mechanisms. Most wanted more provider interaction and wanted their doctor or health care provider to proactively initiate discussion on incontinence and provide information, educational materials, and product samples. CONCLUSION: Respondents indicated that they want more information regarding incontinence. While they may not be equipped to fully understand the problem, they expect doctors, nurses, medical professionals, retail outlets, medical supply companies, and mail order houses to provide the information. Since respondents are receiving information through consumer advertising, which is usually commercially funded, a partnership between the health care community and these sources to produce this information would be ideal. PMID- 15658738 TI - The practice of informed consent. PMID- 15658737 TI - Evaluation of a new method to verify CE completion. AB - Methods of verifying completion of continuing education activities have changed recently as a result of adjustments in the criteria established by the American Nurses Credentialing Center's Commission on Accreditation. Evaluation of the new method of verifying completion of CE articles in Urologic Nursing in 2003 yielded findings that demonstrated that individuals who sent CE answer/evaluation forms for CE articles were thoughtful and insightful in their responses. The responders reported that they learned new information and planned to apply the new knowledge to their practice. PMID- 15658739 TI - Phenomenology as a qualitative research method. PMID- 15658740 TI - Update on CDC's global infectious disease strategy. PMID- 15658741 TI - Desmopressin (DDAVP) for enuresis, diabetes insipidus, and... PMID- 15658743 TI - The gift of laughter. PMID- 15658744 TI - Intonation development from five to thirteen. AB - Research undertaken to date suggests that important developments in the understanding and use of intonation may take place after the age of 5;0. The present study aims to provide a more comprehensive account of these developments. A specially designed battery of prosodic tasks was administered to four groups of thirty children, from London (U.K.), with mean ages of 5;6, 8;7, 10;10 and 13;9. The tasks tap comprehension and production of functional aspects of intonation, in four communicative areas: CHUNKING (i.e. prosodic phrasing), AFFECT, INTERACTION and FOCUS. Results indicate that there is considerable variability among children within each age band on most tasks. The ability to produce intonation functionally is largely established in five-year-olds, though some specific functional contrasts are not mastered until C.A. 8;7. Aspects of intonation comprehension continue to develop up to C.A. 10;10, correlating with measures of expressive and receptive language development. PMID- 15658745 TI - False belief understanding in Cantonese-speaking children. AB - The present study investigates the performance of 96 Cantonese-speaking three- to five-year-old preschoolers on three false belief tasks - a deceptive object, a change of location, and an unexpected contents task encompassing a variety of task factors. Most importantly, the research examines the possibility that false belief performance depends on specific linguistic factors such as the type of verb used in the test question--an explicitly false vs. a neutral belief verb. Cantonese was chosen as particularly useful for examining this question because it explicitly codes belief status as either neutral (nam5) or false (ji5wai4), and because it offers additional linguistic and cultural contrasts to research conducted on false belief with children learning English and other Indo-European languages. As expected, a strong age effect was found, as well as a significant advantage for children who received the explicit false belief (ji5wai4) wording and for those who were asked to explain rather than predict the protagonist's actions. Interestingly, there was also a strong task difference with children performing better on the deceptive object task than on the other two false belief tasks. We argue that these results point both to universal trajectories in theory of mind development and to interesting, but localized, effects of language and culture on children's false belief understanding. PMID- 15658746 TI - Expressing communicative intents in Estonian, Finnish, and Swedish mother adolescent interactions. AB - The present article focused on two types of communicative intent (directing behaviour vs. eliciting talk) expressed by mothers and teenagers during everyday family interactions in Estonian, Finnish, and Swedish mono- and bicultural families. Three monocultural groups consisted of 17 Estonian, 19 Swedish, and 18 Finnish families living in their country of origin; two bicultural and bilingual groups consisted of 18 Estonian and 18 Finnish families residing in Sweden. All the children were between 9;o and 13;0. The results revealed that the Estonian monocultural mothers were highly directive and direct: issuing behavioural directives most frequently and using the highest proportion of imperatives among all samples. Contrary to our expectations, the mothers who tended to be more concerned with controlling their children's behaviour also elicited conversation more frequently from teenagers by using a larger number of information requests. These requests seemed to serve as another form of attempted control over adolescents' behaviour--their communicative behaviour. The Swedish monocultural children were more active in controlling their mothers' behaviour than their counterparts, possibly illustrating the Swedish value of generational equality. PMID- 15658747 TI - Characterizing changes in parent labelling and gesturing and their relation to early communicative development. AB - In a longitudinal study, 17 parent-child dyads were observed during free-play when the children were 1;0, 1;6, and 2;0. Parents' labelling input in the verbal and gestural modalities was coded at each session, and parents completed a vocabulary checklist for their children at each visit. We analysed how the frequency of labelling in the verbal and gestural modalities changed across observation points and how changes in parental input related to children's vocabulary development. As a group, parents' verbal labelling remained constant across sessions, but gestural labelling declined at 2;0. However, there are notable individual differences in parental trajectories in both modalities. Parents whose verbal labelling frequency increased over time had children whose vocabulary grew more slowly than those whose labelling frequency decreased, remained constant, or peaked at 1;6. There were few systematic relations between patterns of parental gesturing and children's vocabulary development. Parents' verbal and gestural labelling patterns also appeared dissociable. However, parents' words and gestures were correlated when their children were 1;6, suggesting that gestures serve an important bootstrapping function at a critical point in children's vocabulary development. PMID- 15658748 TI - Perception of utterance relatedness during the first-word-period. AB - Interactions between six toddlers (aged 1;0 to 1;6) and adults were examined to ascertain adult perceptions of toddler utterance relatedness and to determine temporal and interactional features that underlie those perceptions. Five raters made judgments regarding relatedness of the child utterances to the previous adult utterances; 251 utterances were examined. Utterances judged by adults as related occurred within 4.25 seconds of the preceding adult utterance nearly 90% of the time. This study also points to the need for using interactional categories that go beyond describing utterance relatedness, and introduces terms (i.e. coparticipatory, initiation, narrowed focus) for doing so. PMID- 15658749 TI - Vocabulary competence in first- and secondborn siblings of the same chronological age. AB - We explored vocabulary competence in 55 firstborn and secondborn sibling pairs when each child reached 1;8 using multiple measures of maternal report, child speech, and experimenter assessment. Measures from each of the three sources were interrelated. Firstborns' vocabulary competence exceeded secondborns' only in maternal reports, not in child speech or in experimenter assessments. Firstborn girls outperformed boys on all vocabulary competence measures, and secondborn girls outperformed boys on most measures. Vocabulary competence was independent of the gender composition and, generally, of the age difference in sibling pairs. Vocabulary competence in firstborns and secondborns was only weakly related. PMID- 15658751 TI - What can be learned from positive data? Insights from an 'ideal learner'. Commentary on 'A multiple process solution to the logical problem of language acquisition' by Brian MacWhinney. PMID- 15658750 TI - A multiple process solution to the logical problem of language acquisition. AB - Many researchers believe that there is a logical problem at the centre of language acquisition theory. According to this analysis, the input to the learner is too inconsistent and incomplete to determine the acquisition of grammar. Moreover, when corrective feedback is provided, children tend to ignore it. As a result, language learning must rely on additional constraints from universal grammar. To solve this logical problem, theorists have proposed a series of constraints and parameterizations on the form of universal grammar. Plausible alternatives to these constraints include: conservatism, item-based learning, indirect negative evidence, competition, cue construction, and monitoring. Careful analysis of child language corpora has cast doubt on claims regarding the absence of positive exemplars. Using demonstrably available positive data, simple learning procedures can be formulated for each of the syntactic structures that have traditionally motivated invocation of the logical problem. Within the perspective of emergentist theory (MacWhinney, 2001), the operation of a set of mutually supportive processes is viewed as providing multiple buffering for developmental outcomes. However, the fact that some syntactic structures are more difficult to learn than others can be used to highlight areas of intense grammatical competition and processing load. PMID- 15658752 TI - What if Chomsky were right? PMID- 15658753 TI - Progress, but not a full solution to the logical problem of language acquisition. PMID- 15658754 TI - Bias, innateness and domain specificity. PMID- 15658755 TI - Overgeneralizations, competition, and recovery: solving the logical problem with positive evidence. PMID- 15658756 TI - Something old, something new. PMID- 15658757 TI - Comprehension matters: a commentary on 'A multiple process solution to the logical problem of language acquisition'. PMID- 15658758 TI - Comment on 'A multiple process solution...' (B Macwhinney). PMID- 15658759 TI - Inversion without grammar. PMID- 15658760 TI - Clarifying the logical problem of language acquisition. PMID- 15658761 TI - Learnability, stochastic input, and connectionist networks: a response to Brian MacWhinney's 'A multiple process solution to the logical problem of language acquisition'. PMID- 15658762 TI - Gold's theorems and the logical problem of language acquisition. PMID- 15658763 TI - Strategies for controlling intra- and intermicellar packing in block copolymer solutions: illustrating the flexibility of the self-assembly toolbox. AB - Block copolymers constitute a class of self-assembling macromolecules that offer remarkable flexibility for controlling nanostructure, both in discrete objects and in bulk materials. Block copolymer micelles may be formed with multiple compartments by judicious choice of ingredients in an ABC triblock copolymer. For example, we have shown that a poly(ethylene oxide-b-styrene-b-fluorinated butadiene) triblock assembles in dilute aqueous solution into large, flat core/shell/corona disks, with the fluorine containing block forming the core. In contrast, the unfluorinated precursor generates large spherical micelles. A numerical analysis suggests that the disk-like motif is characteristic of the so called superstrong segregation regime, whereby the interfacial tension becomes so large as to overwhelm the conformational entropy of the core blocks. For ABC miktoarm stars comprising polyethylene oxide, polyethylethylene, and polyhexafluoropropylene oxide arms, a much richer variety of micellar structures are observed. Prominent amongst these is a "segmented worm", in which alternating layers (5-7 nm thick) of hydrocarbon and fluorocarbon blocks form disks (6-10 nm in radius) that stack into cylindrical aggregates. The disk radii suggest almost fully stretched blocks, again consistent with the superstrong segregation regime. These structures are rationalized in terms of the constraints imposed by the star architecture, combined with the extremely strong interfacial tensions. In contrast, for lipids, surfactants, and aqueous diblock copolymers, increasing the interfacial tension can induce a transition from spheres to cylinders to flat bilayers; the disk-like motif is not usually seen. Interestingly, in aqueous diblocks both worm-like micelles and vesicles have been well-documented, whereas in "simple" organic systems they have not. We have shown that by suitable choice of block composition and solvent selectivity, the curvature sequence sphere/cylinder/vesicle can also be observed in poly(styrene-b-isoprene) diblocks in dialkyl phthalates. In more concentrated solutions such spherical micelles assemble onto body-centered or face-centered cubic lattices. In some cases a thermoreversible fcc/bcc transition has been noted. We have recently demonstrated that this transition corresponds to a particular aggregation number; higher aggregation numbers, found at lower temperatures, favor the "hard-sphere-like" fcc packing. All of these results are based on a combination of dynamic light scattering, small angle X-ray and neutron scattering, and cryogenic transmission electron microscopy. PMID- 15658764 TI - Aggregation across the length-scales in beta-lactoglobulin. AB - The protein beta-lactoglobulin (BLG) has been widely studied, in large part because of its importance to the food industry. Following denaturation during heating, under different conditions of pH it has been found to form either particulate (around the isoelectric point at pH 5.1) or fibrillar gels. The nature of the fibrils has recently been suggested to be the same as that identified with amyloid fibrils known for a wide-range of different proteins and implicated in many disease states. We confirm that the BLG fibrils show all the classical signatures of amyloid fibrils. In addition, the fibrils are capable themselves of aggregating further to form large-scale (many microns in size) spherulites. Polarized light microscopy and Environmental scanning electron microscopy (ESEM) have been used to explore the internal structure of these spherulites under conditions in which the solvent has not been dried off. The factors which determine whether or not the spherulites form have also been considered, together with implications for other amyloid-containing systems. PMID- 15658766 TI - Metal-ligand induced supramolecular polymerization: a route to responsive materials. AB - The use of metal-ligand binding as the driving force for the self-assembly polymerizations of a ditopic ligand offers a facile route to the preparation of organic/inorganic hybrid materials. Such metallo-supramolecular polymers potentially offer the functionality of the metal ion along with the processibility of a polymer. We report, herein, the preparation of gel-like metallo-supramolecular polymers prepared from a monomer unit, which consists of a 2,6-bis-(benzimidazolyl)-4-hydroxypyridine unit attached to either end of a polyether chain, mixed with a lanthanoid metal (e.g. La(III), Eu(III)) and a transition metal ion (e.g. Co(II) or Zn(II)). Such materials show dramatic reversible responses to a variety of stimuli, including thermal, mechanical, chemical and light. The nature of the response can be controlled by the nature of the combination of transition metal ion and lanthanoid metal ion used. PMID- 15658765 TI - Peptide mediated formation of hierarchically organized solution and solid state polymer nanostructures. AB - Biologically-inspired peptide sequences have been explored as auxiliaries to mediate self-assembly of synthetic macromolecules into hierarchically organized solution and solid state nanostructures. Peptide sequences inspired by the coiled coil motif and "switch" peptides, which can adopt both amphiphilic alpha-helical and beta-strand conformations, were conjugated to poly(ethylene glycol) (PEG). The solution and solid state self-assembly of these materials was investigated using a variety of spectroscopic, scattering and microscopic techniques. These experiments revealed that the folding and organization properties of the peptide sequences are retained upon conjugation of PEG and that they provide the driving force for the formation of the different nanoscale structures which were observed. The possibility of using defined peptide sequences to direct structure formation of synthetic polymers together with the potential of peptide sequences to induce a specific biological response offers interesting prospects for the development of novel self-assembled and biologically active materials. PMID- 15658767 TI - Responsive brushes and gels as components of soft nanotechnology. AB - Progress in the development of generic molecular devices based on responsive polymers is discussed. Characterisation of specially synthesised polyelectrolyte gels, "grafted from" brushes and triblock copolymers is reported. A Landolt pH oscillator, based on bromate/ sulfite/ferrocyanide, with a room temperature period of 20 min and a range of 3.1 < pH < 7.0, has been used to drive periodic oscillations in volume in a pH responsive hydrogel. The gel is coupled to the reaction and changes volume by a factor of at least 6. A continuously stirred, constant volume, tank reactor was set-up on an optical microscope and the reaction pH and gel size monitored. The cyclic force generation of this system has been measured directly in a modified JKR experiment. The responsive nature of polyelectrolyte brushes, grown by surface initiated ATRP, have been characterised by scanning force microscopy, neutron reflectometry and single molecule force measurements. Triblock copolymers, based on hydrophobic end-blocks and either polyacid or polybase mid-block, have been used to produce polymer gels where the deformation of the molecules can be followed directly by SAXS and a correlation between molecular shape change and macroscopic deformation has been established. The three systems studied allow both the macroscopic and a molecular response to be investigated independently for the crosslinked gels and the brushes. The triblock copolymers demonstrate that the individual response of the polyelectrolyte molecules scale-up to give the macroscopic response of the system in an oscillating chemical reaction. PMID- 15658768 TI - Molecular dynamics modeling of polymer crystallization; from simple polymers to helical ones. AB - Crystallization of helical polymers is a very big challenge for molecular simulation. It involves many significant issues, such as folding in biomolecules and molecular recognition during crystal growth. Though direct molecular simulations of the process still involve very difficult problems, we here report our recent efforts toward better understanding of the crystallization in helical polymers. We begin with a brief review of our former studies on simple polyethylene-like polymers, and then we introduce several helical polymer models which are systematically made more complicated. We have already reported that a simple polyethylene-like polymer crystallizes very fast into chain folded lamellae from the melt. A slight modification of this simple polymer model by introducing proper bond angle and dihedral angle potentials gives one of the present models of the helical polymer. This helical polymer model is devised to be relatively rigid but mobile, to show easy helix-reversals, and to have a definite preference for gauche bonds. We find that this highly mobile helical polymer shows quick chain folded crystallization and forms approximate 4/1 helical structure. The intra- and the intermolecular order grow quite simultaneously suggesting highly cooperative nature of the phenomena. Further elaboration of the helical model, giving pendant side groups and higher energy barrier to the helix reversals, leads us to a realistic united atom model of iPP. The conventional and the multi-canonical Monte Carlo simulations are applied to find probable modes of chain folding and the ground state conformations. Though a very short chain readily forms a regular 3/1 helix of alternating trans and gauche bonds, much longer chains of 30- and 50-propylene units are not found to have energetic ground states in the regularly folded conformations. PMID- 15658769 TI - The effect of molecular architecture on the grain growth kinetics of AnBn star block copolymers. AB - To investigate the effect of molecular architecture on the grain growth kinetics of star block copolymers, a series of AnBn miktoarm star block copolymers with different numbers of arms (n = 1, 2, 4 and 16) was studied. Across this entire series of materials, all the A arms are polystyrene (PS) blocks from the same anionically synthesized batch, and thus all the A arms are identical. Likewise, all the B arms are polyisoprene (PI) blocks from the same anionically synthesized batch, and thus all the B arms are identical. All the stars employed in this study are therefore composed of the same A and B arms liked together in symmetric numbers. The coarsening kinetics of grain growth was monitored in real space by transmission electron microscopy (TEM), followed by subsequent micrograph image analysis. It was found that the molecular architecture influenced the grain growth kinetics of these AnBn star copolymers dramatically. The grain coarsening kinetics was found to follow a scaling law as V approximately t(beta), where V is the characteristic grain volume and t is time. The exponent, beta, was found to be about 0.2 for the diblock copolymer (n = 1) and 0.4 for all three of the star block copolymers (n = 2, 4 and 16) in the series. It is postulated that the difference in grain growth rate between the diblock and the various stars is due to a reduction in molecular entanglements resulting from chain stretching near the junction points in the stars. PMID- 15658770 TI - On the theory of aggregation and micellization: PEO-PVP copolymer in water. AB - We develop a theoretical approach to micellization of the PEO-PVP block-copolymer in water. This copolymer is a weak polyelectrolyte due to protonation of VP blocks. The theory accounts for non-linear ion screening, and predicts strong position dependence of both ion concentration and the effective Debye length. We consider both the case when the local Debye length is small compared to the core radius and the case when it is large. We found that the effective (local) pH is not uniform even inside one micellar core, hence non-uniform protonation of the core with higher charge density near the surface. In many cases the core charge is concentrated in a relatively thin surface layer. Considering statistical weights of non-equilibrium micelles and their continuous evolution we show that kinetics of both formation and dissociation of typical block-copolymer or surfactant micelles can be extremely slow. Thus micelle formation at the genuine (equilibrium) critical micelle concentration (c.m.c.) is totally suppressed (involves astronomical time scales) if the micelles are big enough. An 'apparent' critical micelle concentration (c.m.c.*) is introduced to account for this effect. The apparent c.m.c.* could be much higher than the genuine equilibrium c.m.c., i.e. a significant hysteresis is inherent in these systems. We also determine the ranges of meta-stability of micelles depending on the experimental time-scales. PMID- 15658771 TI - Micellization in pH-sensitive amphiphilic block copolymers in aqueous media and the formation of metal nanoparticles. AB - Dynamic light scattering, potentiometric titration, transmission electron microscopy and atomic force microscopy have been used to investigate the micellar behaviour and metal-nanoparticle formation in poly(ethylene oxide)-block-poly(2 vinylpyridine), PEO-b-P2VP, poly(hexa(ethylene glycol) methacrylate)-block-poly(2 (diethylamino)ethyl methacrylate), PHEGMA-b-PDEAEMA, and PEO-b-PDEAEMA amphiphilic diblock copolymers in water. The hydrophobic block of these copolymers (P2VP or PDEAEMA) is pH-sensitive: at low pH it can be protonated and becomes partially or completely hydrophilic leading to molecular solubility whereas at higher pH micelles are formed. These micelles consist of a P2VP or PDEAEMA core and a PEO or PHEGMA corona, respectively, where the core forming amine units can incorporate metal compounds due to coordination. The metal compounds (e.g., H2PtCl6, K2PtCl6) can either be introduced in a micellar solution, where they are incorporated within the micelle core via coordination with functional groups, or can be added to a unimer solution at low pH, where they lead to a metal-induced micellization. In these micellar nanoreactors, metal nanoparticles nucleate and grow upon reduction with sizes in the range of a few nanometers as observed by TEM. The effect of the metal incorporation method on the characteristics of the micelles and of the synthesized nanoparticles is investigated. PMID- 15658772 TI - Control of pore hydrophilicity in ordered nanoporous polystyrene using an AB/AC block copolymer blending strategy. AB - Ordered nanoporous plastics with hydrophilic pore surfaces were prepared by the degradative removal of polylactide from a self-organised, multi-component composite containing two block copolymers: polystyrene-polylactide and polystyrene-polyethylene oxide. The solid-state characterization of blends containing up to 12 wt.% polyethylene oxide was consistent with nanoscopic cylinders of mixed polyethylene oxide and polylactide hexagonally packed in a polystyrene matrix. Orientation of these materials through simple channel die processing resulted in good cylinder alignment. Subsequent methanolysis/hydrolysis of the polylactide component gave nanoporous polystyrene with polyethylene oxide coated pores. The resulting nanoporous materials were able to imbibe water, in contrast to nanoporous polystyrene with no polyethylene oxide component. PMID- 15658773 TI - Vesicles made of PS-PI cyclic diblock copolymers: in situ freeze-drying cryo-TEM and dynamic light scattering experiments. AB - We have studied the morphology of self-assembled micelles made of linear and cyclic poly(styrene-b-isoprene) PS-b-PI block copolymers dispersed in selective solvents of the PI block (n-heptane, n-decane). Up to a copolymer concentration of 5 mg mL(-1), the micelles made from linear block copolymer chains adopt a spherical shape. Those arising from cyclic copolymer chains having exactly the same molar mass and volume fraction self-assemble into (i) planar sunflower shaped particles at low concentration (c < 0.1 mg mL(-1)), (ii) giant wormlike micelles at intermediate concentration (0.1 mg mL(-1) < c < 2 mg mL(-1)) and (iii) vesicles at higher concentration (2 mg mL(-1) < c < 5 mg mL(-1)). Those results were obtained using dynamic light scattering and in situ freeze-drying cryo-transmission electron microscopy. In this contribution, we discuss the effects of concentration and temperature on the morphology of the self-assembled particles made from both linear and cyclic PS-PI copolymers, and highlight the surprising vesicle formation in cyclic block copolymer solutions. PMID- 15658774 TI - Self-assembled nanostructures from peptide-synthetic hybrid block copolymers: complex, stimuli-responsive rod-coil architectures. AB - The synthesis, solution and solid state self-assembly properties of a series of polyisoprene-b-poly(epsilon-benzyloxycarbonyl-L-lysine) PI-b-PZLys and polyisoprene-b-poly(L-lysine) PI-b-PLys block copolymers have been examined. The formation of stimuli-responsive micelles in water has been studied as a function of pH and ionic strength using static and dynamic light scattering, UV-circular dichroism and transmission electron microscopy. The observed change in the micelles dimensions has been directly attributed to the conformational transition in the secondary structure of polypeptide chains. In bulk, these rod-coil copolymers form self-assembled structures that have been characterized using DSC, DMA, WAXS and SAXS techniques. Hexagonal in lamellar (HL) and more interestingly hexagonal in hexagonal (HH) morphologies have been evidenced as a function of the chemical composition. PMID- 15658775 TI - Tunable diblock copolymer micelles-adapting behaviour via subtle chemical modifications. AB - Selectively quaternising the PDMA block of poly(2-(dimethylamino)ethyl methacrylate)-block-poly(2-(diethylamino)ethyl methacrylate) (PDMA-PDEA) copolymers modifies both their solution and adsorption behaviour. These copolymers exist as free unimers in aqueous solution at low pH and form micelles, with PDMA coronas, at high pH. The critical micellisation pH, the hydrodynamic micelle diameter and electrophoretic behaviour are all affected by the degree of quaternisation of the PDMA block. Highly quaternised copolymers form smaller, more highly charged micelles at lower pH than weakly or non-quaternised copolymers. The adsorption of the copolymer micelles onto muscovite mica is studied by in situ atomic force microscopy. The adsorbed micelle monolayer becomes increasingly disordered as the degree of quaternisation increases. No micelle desorption occurs on removal of the bulk copolymer solution. Addition of acid to the overlying solution leads to different responses from the surface adsorbed micelles. Unquaternised micelles undergo a reversible change in morphology due to the formation of localised polymer brushes, whereas lightly quaternised micelles are characterised by irreversible changes. Highly quaternised micelle monolayers are disrupted by the addition of acid. Such differences can be rationalised by simple electrostatic arguments. This behaviour has been confirmed by quartz crystal microbalance studies, which show that the adsorbed mass decreases with increasing degrees of quaternisation. PMID- 15658776 TI - Self-nucleation and crystallization kinetics of double crystalline poly(p dioxanone)-b-poly(epsilon-caprolactone) diblock copolymers. AB - The crystallization kinetics of each constituent of poly(p-dioxanone)-b poly(epsilon-caprolactone) diblock copolymers (PPDX-b-PCL) has been determined in a wide composition range by differential scanning calorimetry and compared to that of the equivalent homopolymers. Spherulitic growth rates were also measured by polarized optical microscopy while atomic force microscopy was employed to reveal the morphology of one selected diblock copolymer. It was found that crystallization drives structure formation and both components form lamellae within mixed spherulitic superstructures. The overall isothermal crystallization kinetics of the PPDX block at high temperatures, where the PCL is molten, was determined by accelerating the kinetics through a previous self-nucleation procedure. The application of the Lauritzen and Hoffman theory to overall growth rate data yielded successful results for PPDX and the diblock copolymers. The theory was applied to isothermal overall crystallization of previously self nucleated PPDX (where growth should be the dominant factor if self-nucleation was effective) and the energetic parameters obtained were perfectly matched with those obtained from spherulitic growth rate data of neat PPDX. A quantitative estimate of the increase in the energy barrier for crystallization of the PPDX block, caused by the covalently bonded molten PCL as compared to homo-PPDX, was thus determined. This energy increase can dramatically reduce the crystallization rate of the PPDX block as compared to homo-PPDX. In the case of the PCL block, both the crystallization kinetics and the self-nucleation results indicate that the PPDX is able to nucleate the PCL within the copolymers and heterogeneous nucleation is always present regardless of composition. Finally, preliminary results on hydrolytic degradation showed that the presence of relatively small amounts of PCL within PPDX-b-PCL copolymers substantially retards hydrolytic degradation of the material in comparison to homo-PPDX. This increased resistance to hydrolysis is a complex function of composition and its knowledge may allow future prediction of the lifetime of the material for biomedical applications. PMID- 15658777 TI - Oriented primary crystal nucleation in lamellar diblock copolymer systems. AB - We report a numerical study of the primary crystal nucleation of diblock copolymers in a lamellar phase. Only one of two polymer blocks is crystallizable; the other is maintained in a glassy state. We find that crystals in this lamellar geometry tend to nucleate with the chain axis perpendicular to the lamellar plane. However, if in the same lamellar structure, we break the junction between the crystallizable and non-crystallizable blocks of the polymers, we find that crystallites tend to align parallel to the lamellar plane. This observation clarifies the molecular origin of the competition between parallel and perpendicular crystallite orientations in real block-copolymer systems. PMID- 15658778 TI - Isothermal reorganization of poly(ethylene terephthalate) revealed by fast calorimetry (1000 K s(-1); 5 ms). AB - Reorganization of semicrystalline polymers on heating is a fast process. For poly(ethylene terephthalate) (PET) heating rates of several thousand Kelvin per second are needed to prevent reorganization of unstable crystals. Utilizing a thin film vacuum gauge as a fast calorimeter we are able to extend the scanning rate range of commercial DSC's (microK s(-1) to 10 K s(-1)) to rates as high as 10000 K s(-1) on heating and cooling. Because of the fast equilibration time isothermal experiments can be performed after scanning at several thousand Kelvin per second. The dead time after such a quench is in the order of 10 ms and the time resolution is in the order of milliseconds. These ultra fast calorimeters allow us to study the kinetics of extremely fast processes in semicrystalline polymers like reorganization. For PET crystallized at 130 degrees C reorganization needs less than 40 ms between 150 degrees C and 200 degrees C. Isothermal reorganization at 223 degrees C is about two orders of magnitude faster than isothermal crystallization from the isotropic melt at the same temperature. The melt memory for the remaining structures needed for reorganization is removed 25 K above the equilibrium melting temperature of PET. PMID- 15658779 TI - The complex phase behaviour of suspensions of goethite (alpha-FeOOH) nanorods in a magnetic field. AB - In 1902, Majorana reported the magneto-optical properties of aqueous colloidal suspensions of mixed iron oxides. Oddly enough, the magnetic-field induced birefringence displayed a non-monotonic dependence upon field intensity. This behaviour was later interpreted as due to the existence in these sols of at least two different chemical species. During the course of our studies of mineral liquid crystals, we have revisited this problem by examining aqueous suspensions of pure goethite (alpha-FeOOH) nanorods. Although they are comprised of a single chemical species, these suspensions show the same odd behaviour reported by Majorana. Moreover, we show that, as the volume fraction increases, the suspensions have an isotropic liquid/nematic/rectangular columnar phase sequence, with first-order transitions between these phases. The non-monotonic dependence of the field-induced birefringence can be explained by the existence of a remanent magnetic moment of the nanorods and the negative anisotropy of their magnetic susceptibility. Therefore, the nanorods align parallel to a weak field but realign perpendicular to the field beyond Bc approximately 375 mT. In addition, other interesting phenomena appear upon application of a magnetic field: the disordered (i.e. isotropic in zero-field) phase becomes highly anisotropic and difficult to distinguish from the nematic phase. Both phases then acquire not only quadrupolar order but also dipolar order. The rectangular columnar phase is strongly stabilised versus the nematic one. Our experimental observations raise new theoretical questions about the phase diagram of these suspensions with respect to volume fraction and magnetic field intensity. PMID- 15658780 TI - Micro-convection, dissipative structure and pattern formation in polymer blend solutions under temperature gradients. AB - We report experimental results on time evolution of phase separation in a polymer blend solution under a temperature gradient. This study is aimed at making a self organized structure by utilizing convection in a polymer blend solution comprising polystyrene (PS) and polybutadiene (PB) as solute with dioctylphthalate (DOP) as solvent. The polymer blend solution (PS/PB/DOP) with various compositions (PS/PB = 20/80, 50/50 or 80/20) was prepared with a total polymer concentration (PS + PB) at 3.0 wt.%. The solution was confined in a layer (0.1 mm thick), which was further subjected to a temperature gradient by controlling top and bottom temperatures (Ttop and Tbot) independently with a 10 or 20 degrees C temperature difference (Ttop < Tbot). For all compositions, micro convection was generated due to the Soret effect, which is relevant to the fact that the temperature gradient induces a concentration gradient. An unusual phase separating process was observed as a result of dynamic coupling with convective flows. Especially, an interesting hierarchical structure was formed for a PS/PB = 50/50 solution. This structure is characterized by a doubly surrounding "sea island". Namely, the "sea" contains large "islands" which involve tiny "islands". The large "islands" are spatially arranged and embedded in polygonal convection cells. PMID- 15658781 TI - Shear-induced smectic ordering and crystallisation of isotactic polypropylene. AB - Shear-induced smectic ordering and crystallisation of isotactic polypropylene (iPP) has been studied with in-situ small- and wide-angle X-ray scattering. Shear induced smectic bundles with a periodicity of about 4 nm have been observed at temperatures below as well as above the melting point. This applies to iPP of different molecular weight and from different sources. An increase in the average molecular weight leads to a larger periodicity of the smectic layers. The smectic layers assemble in a fibrillar morphology with a length and a width up to 200 and 10 microm, respectively. After crystallisation, the smectic bundles show upon heating higher melting temperatures than their crystalline counterparts. In agreement with this behaviour, the correlation length along the smectic layer normal is of the order of tens of nanometers, much larger than the crystal thickness. We present an anisotropic drop model of smectic domains forming a conserved system in which the smectic layers can rotate. On the basis of this model we can explain the relative orientation of the smectic layers, the crystalline lamellae and the long axis of the drop, as well as the reversibility of the smectic periodicity during cooling and heating. In the supercooled melt, the smectic ordering is followed by crystallisation; during this process crystals grow epitaxially on the surface of the smectic bundles. This leads to a new picture of the shish-kebab structure in which smectic bundles rather than extended-chain crystals play the role of the shish. The crystallisation process of the smectic regions themselves indicates that the 'mesophase' previously reported in fast-quenched iPP, is a metastable state formed during the transition from the high-temperature smectic phase to a crystal. Moreover smectic domains rather than alpha crystallites form the nuclei for crystallisation of the beta phase. The high-temperature smectic phase presents an ideal model system to study the coupling between density and conformational ordering under shear flow. Our results support a nucleation and growth process for polymer crystallisation, in which smectic bundles or other mesophases are the primary nuclei. PMID- 15658782 TI - Self-organization of cationic dendrimers in polyanionic hydrogels. AB - Protonated poly(propylene imine) dendrimers (Astramol) of five generations: DAB dendr-(NH2)x (where x=4, 8, 16, 32 or 64) are sorbed by slightly cross-linked polyanionic hydrogels: poly(sodium acrylate) and poly(sodium 2-acrylamido-2 methylpropane sulfonate). As a result highly swollen original hydrogel transforms into compact cross-linked polyelectrolyte-dendrimer complexes. Sorption of dendrimers by the hydrogels is a chemically drawn frontal diffusion process. Driving force comes from the gain in the free energy of interpolyelectrolyte coupling reaction between the charged dendrimer molecules and the oppositely charged hydrogel network, accompanied with entropically favourable release of low molecular salt into environment. The amount of a simple salt released is equivalent to a number of intermolecular salt bonds, formed between protonated dendrimers and hydrogel networks. Apparently the mechanism of dendrimer uptake involves a "relay-race" transfer of dendrimer polycations from one fragment of polyelectrolyte network to the other via interpolyelectrolyte exchange reaction. As a result "core-shell" constructs consisting of outer weakly swollen complex shell and highly swollen hydrogel core are formed at intermediate stages of the process. The rate of sorption is determined by the rate of the interpolyelectrolyte exchange reaction that is the rate of the formation of free fragments of polyelectrolyte network (vacancies) on the inner complex-hydrogel boundary. The amount of vacancies depends on the area of this boundary. Consequently kinetics of dendrimer uptake could not be fitted in terms of Fickian diffusion (except DAB-dendr-(NH2)4), but expressed in terms of the kinetic equation derived for a frontal heterogeneous reaction. Sorbed dendrimers of all studied generations at pH values ensuring complete protonation of primary and tertiary amine groups are closely packed in hydrogel networks, so that all dendrimer cationic units form ion pairs with anionic units of hydrogels. In other words polyanionic network fragments are able to penetrate into the interior of fully protonated DAB-dendr-(NH2)x species as it was earlier shown for flexible linear polyanions. In such case the ultimate amount of sorbed dendrimer molecules is always determined by the condition n(a)/N- = 1, where n(a) is the total number of dendrimer amine groups, N- is the number of the anionic hydrogel units. The latter is also true for the complex shell composition in the heterogeneous reacting samples formed at intermediate stages of dendrimers uptake. Variation of pH and sorption extent is an effective tool to control dendrimer distribution, self-organization and the final structure of dendrimer-hydrogel constructs. PMID- 15658783 TI - Design of chimaeric polymersomes. AB - We discuss the development of hierarchical polymer particles, or variegated polymersome composites, in which at least two different components are phase separated within one polymersome chimaera. We briefly discuss the present status in experimental polymersome research, and then discuss a speculative design strategy, based on mesoscopic simulations with a dynamical variant of polymer self-consistent field theory (Mesodyn). The main conclusion is that the counter intuitive co-assembly of demixing block copolymers is the key in controlling hierarchical structures on a mesoscopic scale. This is the classical paradox of a chimaera: the constituents live in the same scaffold, but apart. Block copolymers beyond a certain length will always split the assembly, and without further precautions, polymer based chimaerae are intrinsically unstable. To this end, we propose the application of a branched block copolymer as composite compatibilizer, glueing the separate domains together, and thereby stabilizing the chimaeric polymersome. PMID- 15658784 TI - Organisation in two series of low-dimensional polymer electrolytes with high ambient lithium salt conductivity. AB - The amphiphilic polymers poly[2,5,8,11,14-pentaoxapentadecamethylene(5 hexadecyloxy-1,3-phenylene)] I (abbrev. C16O5), its homologues C16On, n = 1-4, and its copolymers with C16O1 have been synthesised by two Williamson procedures. Method X gives polyether-esters; method Y gives pure polyethers. DSC, optical microscopy, small-to-wide angle X-ray diffraction and complex impedance spectroscopy have been used to investigate organisation and ionic conductivities of the C16On and the C16O5-C16O1 copolymer series and their complexes with LiClO4 and LiBF4. Four-component mixtures of the latter systems (polymers I) with polyoxytetramethylene-decamethylene copolymer (II) and an octadecyl-terminated ABA block copolymer (III) (interfacial stabiliser) ('type D' systems) gave high ambient conductivities ( > or = 10(-4) S cm(-1)) with low temperature dependence. Highest conductivities (8 x 10(-4) S cm(-1) at 20 degrees C) were observed with a type D system based upon C12C18O5X. PMID- 15658785 TI - Light-sensitive fusion between polymer-coated liposomes following physical anchoring of polymerisable polymers onto lipid bilayers by self-assembly. AB - Delivery of therapeutics (drugs, radionuclides or genes) in vivo can be optimized when carried by a targeting delivery vehicle such as a surfactant vesicle, polymeric micelle or other polymer-coated colloidal particulate. In the present communication, we propose a general method based on self-assembly principles, to construct lipid-polymer bilayer vesicles whose featured characteristics may be altered according to the polymer molecule used, thus be easily designed along the needs of a particular delivery application. Polymer molecules containing non polymerizable (polypropylene) and polymerisable (methacrylate) hydrophobic groups were used to construct lipid-polymer vesicles by following two different methods of preparation. In accord with our previous findings, when both types of polymer molecules are added to pre-formed liposomes, only weak adsorption onto the lipid surface occurs. Preparation of the vesicles by pre-mixing the lipid and polymer molecules has proved essential in order to allow the hydrophobic blocks of the copolymers to participate as integral parts of the bilayer. Anchoring of a polymerisable polymer onto the lipid bilayer by hydrophobic interactions, resulted in steric stabilization of the vesicles. When UV polymerization of the bilayer-incorporated [Methyl(PEG)2000MA] polymer was induced, inter-vesicle fusion was triggered. Direct cryo-EM imaging of fusion between the PEG-coated liposomes has been observed. Such sterically stabilised fusogenic vesicles were constructed as potential triggered-release delivery systems, responsive to a variety of external stimuli depending on the type of polymerisable, hydrophobic group in the polymer molecule. By altering the properties of the incorporated hydrophobic group, liposomes able to fuse in response to initiators milder than UV light, such as green or red light, sound, temperature, oxygen or pH can be engineered. PMID- 15658786 TI - Polyelectrolyte-surfactant complex: phases of self-assembled structures. AB - We study the structure of complexes formed between ionic surfactants (SF) and a single oppositely charged polyelectrolyte (PE) chain. For our computer simulation we use the "primitive" electrolyte model: while the polyelectrolyte is modeled by a tethered chain of charged hard sphere beads, the surfactant molecules consist of a single charged head bead tethered to a tail of tethered hard spheres. A hydrophobic attraction between the tail beads is introduced by assuming a Lennard Jones potential outside the hard-sphere diameter. As a function of the strengths of both the electrostatic and the hydrophobic interactions, we find the following scenario: switching on and increasing the electrostatic forces first leads to a stretching of the PE and then by condensation of SF to the formation of a complex. For vanishing hydrophobic forces this complex has the architecture of a molecular bottle-brush cylindrically centered around the stretched PE molecule. Upon increasing the hydrophobic attraction between the SF tails, a transition occurs inverting this structure to a spherical micelle with a neutral core of SF tails and a charged corona of SF heads with the PE molecule wrapped around. At intermediate hydrophobicity there is a competition between the two structures indicated by a non-monotonic dependence of the shape as function of the Coulomb strength, favoring the cylindrical shape for weak and the spherical micellar complex for strong interaction. PMID- 15658787 TI - Self-organising polymers. PMID- 15658788 TI - Cognitive and behavioral correlates of tuberous sclerosis complex. AB - Tuberous sclerosis complex is a multisystem disorder in which neurologic problems cause the greatest disability. High rates of mental retardation and autism spectrum disorders are associated with the diagnosis. Early-onset seizures and increased tuber burden are risk factors for cognitive impairment. Early-onset seizures, particularly infantile spasms, are risk factors for autism. Tubers within the temporal lobe and cerebellum are often mentioned as risk factors for autism, although the findings are inconsistent. Seizure control is important for developmental outcome and quality of life. Early behavioral assessment and therapeutic intervention, as well as seizure control, are the most effective means of promoting neurodevelopmental outcome. PMID- 15658789 TI - T2-weighted hyperintensities (unidentified bright objects) in children with neurofibromatosis 1: their impact on cognitive function. AB - The impact of magnetic resonance imaging (MRI)-identified T2-weighted hyperintensities (unidentified bright objects) on the cognitive function of children with neurofibromatosis 1 is controversial. We recruited 32 right-handed children with neurofibromatosis 1 (22 boys, 10 girls) aged between 5 and 16 years (mean age 10.2 years) for magnetic resonance imaging examinations and neuropsychologic evaluation. Statistical analysis was performed to evaluate the significance of the hyperintensities. Twenty-four children had unidentified bright objects, whereas eight children did not. Using the t-test, thalamic lesions were associated with lower intellectual function (P = .031). Left globus pallidus hyperintensities were associated with a lower attention score (P = .04), and right middle cerebellar peduncle hyperintensities were associated with a lower sensorimotor score (P = .05). The size of the thalamic lesions correlated with cognitive function (P < .05). Among the group with unidentified bright objects, there was a significant association between more involved sites on the dominant hemisphere and impaired verbal function (r = -.55; P = .005). Unidentified bright objects in the thalamus, globus pallidus, and middle cerebellar peduncles and the laterality of the lesions had an impact on cognitive function. PMID- 15658791 TI - Diagnosis, treatment, and prognosis of Mycoplasma pneumoniae childhood encephalitis: systematic review of 58 cases. AB - Most of the knowledge on Mycoplasma pneumoniae encephalitis in children is based on case reports or small case series. The aim of this study was to assess the important features of the disease by a systematic review of previously published cases. Overall, 58 cases (22 female, 36 male; median age 10 years, range: 2-17 years), derived from 38 reports, were reviewed. Forty-five patients (76%) had flulike or respiratory symptoms prior to the onset of neurologic disease, and 23 patients (40%) presented with pulmonary infiltrates. Cerebrospinal fluid (CSF) pleocytosis (mostly mononuclear cells) and elevated cerebrospinal fluid protein counts were present in 34 (59%) and 21 (36%) patients, respectively. Cranial computed tomography (CT) or magnetic resonance imaging (MRI) abnormalities were seen in 18 (31%) and 17 (29%) patients, respectively, whereby various different presentations were observed. Forty-four patients (76%) received antimicrobial therapy active against mycoplasmas, most frequently erythromycin (n = 29) or minocycline (n = 6). Only in one patient, cure was directly attributed to antimicrobial therapy (with chloramphenicol) by the respective authors. Thirty three patients (57%) recovered without sequelae, 20 patients (34%) had minor to major sequelae, and 5 patients (9%) died. A high cerebrospinal fluid cell count, cerebrospinal fluid protein elevation, and higher age were associated with an unfavorable outcome. PMID- 15658790 TI - Respiratory syncytial virus infection and neurologic abnormalities: retrospective cohort study. AB - Respiratory syncytial virus is a common cause of acute respiratory infection in children. Previous reports have associated respiratory syncytial virus infection and acute neurologic symptoms, including apnea and seizures. This study examined the prevalence of acute neurologic symptoms associated with respiratory syncytial virus and non-respiratory syncytial virus respiratory infections in children requiring admission to a pediatric intensive care unit. We screened all admissions over a 4-year period at a single center, identifying children between birth and 2 years of age with bronchiolitis or acute upper respiratory infection. Children with a history of prematurity, preexisting medical conditions, or prior neurologic problems were excluded. Retrospective chart review of 236 admissions was conducted, identifying the etiology of respiratory infection and the presence or absence of acute neurologic signs or symptoms. This study demonstrated a prevalence of acute neurologic signs or symptoms in 36.4% of respiratory syncytial virus-negative patients (n = 115) and 39.1% in the respiratory syncytial virus-positive group (n = 121). Apnea, the most common symptom, was observed in 19.8% of respiratory syncytial virus-positive cases and 16.5% of negative patients. Seizures were reported in 6.6% of the positive cases and in 12.2% of the negative cohort. These differences showed no statistical significance. Although neurologic signs and symptoms are common in this population of children with acute respiratory infection, there was no significant difference in the prevalence of neurologic problems between respiratory syncytial virus-positive and -negative patients. PMID- 15658792 TI - Diffusion-weighted imaging and proton magnetic resonance spectroscopy in perinatal hypoxic-ischemic encephalopathy: association with neuromotor outcome at 18 months of age. AB - We evaluated early diffusion-weighted imaging findings, the quantitative apparent diffusion coefficient, and magnetic resonance spectroscopy (the presence of lactate and ratios of N-acetylaspartate to total creatine and choline to total creatine) in the prediction of the 18-month neuromotor outcome of term newborns with hypoxic-ischemic encephalopathy. Conventional T1- and T2-weighted and diffusion-weighted imaging was performed in 20 asphyxiated term newborns, with additional basal ganglia magnetic resonance spectroscopy in 15 newborns between 2 and 18 days of life (mean 7.3 days). Neuromotor outcome was dichotomized into normal and abnormal for statistical analysis. Statistically significant differences in the ratios of N-acetylaspartate to total creatine, but not apparent diffusion coefficient values and ratios of choline to total creatine, were found between infants with a normal and an abnormal outcome (Mann-Whitney U test, P = .010). There was a significant association between the presence of a lactate peak and an abnormal outcome (chi-square test, P = .017). The presence of a lactate peak for predicting an abnormal outcome had a sensitivity of 100% and a specificity of 80%, and the odds ratio was 37.4. Ischemic lesions were more conspicuous and/or extensive on diffusion-weighted imaging in all except one neonate. The presence of normal findings on both diffusion-weighted imaging and conventional magnetic resonance imaging is predictive of a normal neuromotor outcome, whereas lactate and a reduced ratio of N-acetylaspartate to total creatine in the basal ganglia, but not an apparent diffusion coefficient, are associated with an abnormal outcome at 18 months of age. PMID- 15658793 TI - Comparative audit of clinical research in pediatric neurology. AB - Clinical research involves direct observation or data collection on human subjects. This study was conducted to evaluate the profile of pediatric neurology clinical research over a decade. Trends in pediatric neurology clinical research were documented through a systematic comparative review of articles published in selected journals. Eleven journals (five pediatric neurology, three general neurology, three general pediatrics) were systematically reviewed for articles involving a majority of human subjects less than 18 years of age for the years 1990 and 2000. Three hundred thirty-five clinical research articles in pediatric neurology were identified in the 11 journals for 1990 and 398 for 2000, a 19% increase. A statistically significant increase in analytic design (21.8% vs 39.5%; P = .01), statistical support (6% vs 16.6%; P < .0001), and multidisciplinary team (69.9% vs 87%; P = .003) was observed. In terms of specific study design, a significant decline in case reports (34.3% vs 10.3%; P < .0001) and an increase in case-control studies (11.3% vs 22.9%; P = .02) were evident over the 10-year interval. This comparative audit revealed that there has been a discernible change in the methodology profile of clinical research in child neurology over a decade. Trends apparently suggest a more rigorous approach to study design and investigation in this field. PMID- 15658794 TI - Cerebral palsy: correlation of risk factors and functional performance using the Functional Independence Measure for Children (WeeFIM). AB - We studied the risk factors affecting the functional status of cerebral palsy. A cross-sectional study of 73 children with cerebral palsy was conducted with the Functional Independence Measure for Children (WeeFIM), which assesses functional skills in the domains of self-care, mobility, and cognition. The mean total Functional Independence Measure for Children quotient was 67.5%. The mean subquotients for self-care, mobility, and cognition were 68.3%, 62.7%, and 69.4%, respectively. The risk factors related to the degree of functional dependency were (1) mental retardation (P = .030), (2) epilepsy (P = .005), (3) type of cerebral palsy (P < .001), and (4) severity of cerebral palsy using the Gross Motor Function Classification System (P < .001) (using univariate analysis). However, when using multivariate analysis, only epilepsy (P = .02) and severity status according to the Gross Motor Function Classification System (P < .001) were significantly related. When the etiology was analyzed, only prematurity was significantly associated with better Functional Independence Measure for Children scores using both univariate (P = .022) and multivariate (P = .007) analyses. The functional status of children with cerebral palsy depends on the severity and the presence of epilepsy. Despite impairment, we found that most children with cerebral palsy could achieve functional independence. PMID- 15658795 TI - Intra- and interrater reliabilities of the Myotonometer when assessing the spastic condition of children with cerebral palsy. AB - The purposes of this study were to assess intra- and interrater reliabilities by novice users of the Myotonometer (Neurogenic Technologies, Inc., Missoula, MT), a portable electronic device that quantifies muscle tone (stiffness) and paresis, in assessing children with cerebral palsy. Two raters used the Myotonometer to assess the biceps brachii and medial gastrocnemius muscles of 10 children with spastic-type cerebral palsy. Muscles were measured in a relaxed state and during a voluntary isometric contraction. Intraclass correlation coefficients and repeatability coefficients were calculated for each muscle and for each condition (relaxed and contracted). Intrarater reliabilities ranged from 0.82 to 0.99 (biceps brachii muscles) and 0.88 to 0.99 (medial gastrocnemius muscles). Interrater reliabilities ranged from 0.74 to 0.99 (biceps brachii muscles) and 0.84 to 0.99 (medial gastrocnemius muscles). Repeatability coefficients indicated a 98% level of agreement between raters across all conditions. Novice users of the Myotonometer, with few exceptions, had high to very high intra- and interrater reliabilities for measurements of the biceps brachii and medial gastrocnemius muscles of children with spastic-type cerebral palsy. PMID- 15658796 TI - How can the Internet help parents of children with rare neurologic disorders? AB - The objective of this study was to determine the value of an e-mail listserv for parents of children with Rett syndrome, a rare neurologic disorder. This Web based survey was completed by parents and carers. The setting was an e-mail listserv established by the International Rett Syndrome Association for parents of children with Rett syndrome and other interested persons. The participants included members of the e-mail listserv Rettnet. The main outcome measures were the perceived advantages and disadvantages of the listserv, overall rating of usefulness, and reasons for satisfaction or dissatisfaction. Most (81.5%) of the participants felt that Rettnet provided helpful advice concerning their child's management. They also indicated that Rettnet was useful in dealing with their child's education and as a source of carer support. They rated it highly (mean 8.1 on a scale of 1 to 10), and the most common reason given for recommending the service to other parents was the emotional support provided. E-mail listservs can play an important role in disseminating information and providing networking and support to parents of children with rare disorders. Their impact and influence warrant attention from health professionals, including neurologists. PMID- 15658797 TI - A neurocutaneous disorder with a severe course: Wyburn-Mason's syndrome. AB - Wyburn-Mason's syndrome is a rare neurocutaneous disorder consisting mainly of unilateral arteriovenous malformations of the midbrain and retina with multiple cutaneous nevi. The authors report on the clinical presentation, neurologic phenotype, and long-term neurologic follow-up of two unrelated children. The first patient had recurrent epistaxis during early childhood. At the age of 7 years, he developed acute hemianopsia and right hemiplegia. Angiography revealed large bilateral arteriovenous malformations involving the midbrain thalamic area and the right optic nerve. During the following years, he had recurrent episodes of headache, right hemiplegia, and cognitive deterioration. The second patient had some episodes of epistaxis in the first years of life. At the age of 5 years, he presented with sudden onset of headache, followed by a loss of consciousness, vomiting, and, subsequently, visual disturbances. Angiography revealed deeply located arteriovenous malformations involving the right temporal, frontobasal, capsulonuclear, insular, and parietal areas and the right optic nerve. During the following years, he had an acute strokelike episode followed by transient hemiplegia and slow progressive signs, with mild worsening of cognitive abilities. Early onset of neurologic manifestations is a poor prognostic factor for long-term outcome. PMID- 15658798 TI - Acute cerebellitis complicated by hydrocephalus and impending cerebral herniation. AB - We present a 7-year-old boy with acute cerebellitis who required an emergency ventriculoperitoneal shunt for hydrocephalus caused by cerebellar swelling. This represents a very unusual, potentially life-threatening complication of a usually self-limiting condition. Early diagnosis of this complication is essential in view of the propensity to sudden and fatal deterioration. Magnetic resonance imaging (MRI) is useful in differentiating this unusual course of acute cerebellar ataxia from that of a posterior fossa tumor. In developing countries, however, computed tomography (CT) is often the only existing diagnostic modality, and access to MRI, when available, is limited. Our case demonstrates that the shape of the fourth ventricle on CT can be helpful in differentiating between a tumor and edema of the cerebellum and thus can assist in management. PMID- 15658799 TI - Extrapontine myelinolysis in infancy: report of a case. AB - Osmotic myelinolysis is a rare, acute, demyelinating process that involves the pons (central pontine myelinolysis) and other locations of the central nervous system (extrapontine myelinolysis). Central pontine myelinolysis is described in children, usually associated with rapid correction of hyponatremia. Other conditions, such as hypernatremia and hyperglycemia, have also been reported as being responsible for pontine myelinolysis. Extrapontine myelinolysis in childhood is very rare and presents in a wide variety of locations. We report a patient who developed extrapontine myelinolysis in the cerebellum during treatment of hyponatremic dehydration. This is the first case reported during infancy. PMID- 15658800 TI - A pH-stating mechanism in isolated wheat (Triticum aestivum) aleurone layers involves malic acid transport. AB - Acidification of the starchy endosperm by the aleurone layer following germination has been established; however, the physiological and metabolic responses of this tissue to external pH have been incompletely investigated. In this investigation, isolated wheat (Triticum aestivum) aleurone layers were incubated in different solutions at initial pH values of 3, 4 and 6 in the absence of phytohormones. After 24 h of incubation, the initial pH of all malate and succinate buffers shifted towards a value close to 4.2. These results suggest the existence of a pH-stating mechanism, instead of the simple acidification process reported previously. The rise of initial pH 3 by aleurone layers was accompanied by a high net uptake of external malic- or succinic acid. In contrast, incubation in glycyl-glycine buffer (a supposedly non-permeating cation at pH 3) partially prevented that pH rise in a pH-3 solution. The 14C-malate taken up from media at pH 3 was mostly broken down to CO2, indicating that an effective metabolic control of the intracellular malate level was operating. At pH 6, an uptake of 14C-malate and 14CO2 production occurred as well, but at slower rates than at pH 3. When buffer concentration was increased, at initial pH values of 3 or 6, a higher uptake or secretion of malic acid, respectively, was carried out by the aleurone layers. The pH of these buffers varied less than that of dilute ones, but always showed a tendency toward a pH near 4. These results suggest that a balance between secretion and uptake of malic acid, accompanied by the corresponding biosynthesis or degradation, is the basis of this pH-stating mechanism. PMID- 15658801 TI - Valine uptake in the tap root of sugar beet: a comparative analysis with sucrose uptake. AB - Given the lack of data on the absorption of amino acids in the tap root of Beta vulgaris, we studied the uptake of valine and compared it with that of sucrose at the same concentration (1 mM). The uptake of both substrates shared some similar characteristics. In particular, the absorption in both cases was controlled by an active process as evidenced by the inhibitory effect of CCCP and inhibitors of ATPases (DES, DCCD, orthovanadate). Both absorptions also involved the thiol and histidyl groups of protein carriers included in the plasmalemma as shown by treatment with specific compounds (PCMBS, mersalyl, NEM) inhibiting the transport of the nutrients in tissues and in purified PMV. However, it was shown that these uptakes present major differences. Firstly, unlike sucrose uptake, valine uptake was very sensitive to transmembrane electrical potential. Indeed, hyperpolarizing treatment with FC increased valine uptake but did not modify sucrose uptake. By contrast, treatment with high concentrations of KCl, which should result in depolarization of the cells, considerably decreased valine uptake and activated sucrose uptake. Secondly, ion mobilizations were different in the two types of transport. Unlike sucrose, application of valine to tissues strongly modified the time course of H+ influx. By contrast, sucrose uptake was controlled by K+ involvement as shown by effects either of modulators of K+ mobilization (LiCl, TEA) or of treatments inducing K+ starvation from the external medium. PMID- 15658802 TI - Paraheliotropism can protect water-stressed bean (Phaseolus vulgaris L.) plants against photoinhibition. AB - In order to estimate the importance of leaf movements on photosynthesis in well watered and water-stressed field grown bean cultivars (Arroz Tuscola (AT), Orfeo INIA (OI), Bayos Titan (BT), and Hallados Dorado (HD)), CO2 assimilation, leaf temperature, and capacity for the maximum quantum yield recovery, measured as Fv/Fm, were assessed. Leaf water potential was lower in water-stressed compared to control plants throughout the day. Water status determined a decrease in the CO2 assimilation and stomatal conductance as light intensity and temperature increased up to maximal intensities at midday. Both parameters were lower in stressed compared to control plants. Even though high light intensity and water stress induced stomatal closure is regarded as a photoinhibitory condition, the recovery of variable to maximal fluorescence (Fv/Fm) after 30min of darkness was nearly constant in both water regimes. In fact, higher values were observed in OI and AT when under stress. Photochemical and non-photochemical fluorescence quenching resulted in minor changes during the day and were similar between watered and stressed plants. It is concluded that paraheliotropism, present in the four bean cultivars, efficiently protects stressed plants from photoinhibition in the field and helps maintain leaf temperatures far below the ambient temperatures, however, it may also be responsible for low CO2 assimilation rates in watered plants. PMID- 15658803 TI - Light-induced proton slip and proton leak at the thylakoid membrane. AB - A treatment of leaves of Spinacia oleracea L. with light or with the thiol reagent dithiothreitol in the dark led to partly uncoupled thylakoids. After induction in intact leaves, the partial uncoupling was irreversible at the level of isolated thylakoids. We distinguish between uncoupling by proton slip, which means a decrease of the H+/e(-) -ratio due to less efficient proton pumping, and proton leak as defined by enhanced kinetics of proton efflux. Proton slip and proton leak made about equal contributions to the total uncoupling. The enhanced proton efflux kinetics corresponded to reduction of subunit CF1-gamma of the ATP synthase as shown by fluorescence labeling of thylakoid proteins with the sulfhydryl probe 5-iodoacetamido fluorescein. The maximum value of the fraction of reduced CF1-gamma was only 36%, which indicates that in vivo the reduction of CF1-gamma could be limited by fast reoxidation and/or restricted accessibility of CF1-gamma to thioredoxin. Measurements of the ratio ATP/2e indicated that only the uncoupling related to less efficient proton pumping led to a decrease in the ATP yield. PMID- 15658804 TI - Factors affecting determination of superoxide anion generated by mitochondria from barley roots after anaerobiosis. AB - During the post-hypoxic period, symptoms of oxidative stress and activation of enzymatic and non-enzymatic antioxidant systems were observed in several plant tissues. In the roots, mitochondrial respiratory chain is the main source of ROS. Superoxide anion radical is formed in the mitochondrial electron-transport chain at the level of Complexes I and III. The purpose of this work was to estimate superoxide anion production by the mitochondria isolated after a period of hypoxic treatment. Seedlings of barley (Hordeum vulgare L.) were grown on a nutrient medium flushed for 5d with air (control) or nitrogen (hypoxia) and then transferred for 24h to aerated medium (post-hypoxia). Production of superoxide anion by the mitochondria was measured by SOD-inhibitable oxidation of adrenaline to adrenochrome with NADH as a respiratory substrate. Hypoxic treatment increased mitochondrial activity but decreased mitochondrial superoxide anion appearance outside the mitochondrial membrane as compared to the mitochondria isolated from the roots continuously grown on aerated medium. The result of lower superoxide anion determination is attributed to increased antioxidants concentration during hypoxia. This was confirmed by inhibition of O2- production by exogenous GSH and stimulation by addition of 1-chloro-2,4-dinitrobenzene (CDNB), which depleted endogenous mitochondrial GSH. PMID- 15658805 TI - Hydrogen peroxide is necessary for abscisic acid-induced senescence of rice leaves. AB - The role of H2O2 in abscisic acid (ABA)-induced rice leaf senescence is investigated. ABA treatment resulted in H2O2 production in rice leaves, which preceded the occurrence of leaf senescence. Dimethylthiourea, a chemical trap for H2O2, was observed to be effective in inhibiting ABA-induced senescence, ABA increased matondialdehyde (MDA) content, ABA-increased antioxidative enzyme activities (superoxide dismutase, ascorbate peroxidase, glutathione reductase and catalase), and ABA-decreased antioxidant contents (ascorbic acid and reduced glutathione) in rice leaves. Diphenyteneiodonium chloride (DPI) and imidazole (IMD), inhibitors of NADPH oxidase, and KCN and NaN3, inhibitors of peroxidase, prevented ABA-induced H2O2 production, suggesting NADPH oxidase and peroxidase are H2O2-generating enzymes in ABA-treated rice leaves. DPI, IMD, KCN, and NaN3 also inhibited ABA-promoted senescence, ABA-increased MDA contents, ABA-increased antioxidative enzyme activities, and ABA-decreased antioxidants in rice leaves. These results suggest that H2O2 is involved in ABA-induced senescence of rice leaves. PMID- 15658806 TI - Seawater irrigation: antioxidant defence responses in leaves and roots of a sunflower (Helianthus annuus L.) ecotype. AB - Salinity is a widespread environmental stress for crop plants. It is common in arid, semiarid, and coast regions. In those environments, seawater infiltrations can occur or the sea provides the only source of water for irrigation. The effects of 10% and 20% seawater in nutrient solutions were studied in 30 day-old plants of sunflower (Helianthus annuus L.) ecotype Katharina Piacenza. Growth parameters, ascorbate and glutathione contents, and the activities of ascorbate peroxidase and glutathione reductase were determined in shoots and roots. The results showed antioxidative responses of the ecotype to both salt treatments. The different activity patterns of antioxidant molecules and enzymes in the leaves and roots suggested a different kind of reaction to the two seawater concentrations. PMID- 15658807 TI - Ultrastructural changes and location of beta-1, 3-glucanase in resistant and susceptible cotton callus cells in response to treatment with toxin of Verticillium dahliae and salicylic acid. AB - Calli from two cotton cultivars susceptible and resistant to Verticillium wilt, were treated with a crude toxin of Verticillium dahliae (VD-toxin) plus salicylic acid (SA). Cells treated with VD-toxin showed distinct ultrastructural changes. Cells from the susceptible cultivar displayed damage to plasma membrane and cytoplasm. The deleterious effect on cells of the resistant cultivar, with an accumulation of electron-dense precipitate in the vacuoles, was less noticeable. Exogenous SA protected callus cells from VD-toxin. We also report the localization of beta-1,3-glucanase in callus cells with immunofluorescence Labeling. Stronger fluorescence was observed in the extracellular space in resistant than in susceptible cotton; strongest in resistant cotton after 5 days of treatment with VD-toxin plus SA. The findings reported here indicate an important role of exogenous salicylic acid in the induction of resistance to VD toxin in cotton. Coupled with an increase in beta-1,3-glucanase, cellular integrity is maintained and damage to cell wall and plasma membrane is avoided. PMID- 15658808 TI - Rapid determination of fungal colonization and arbuscule formation in roots of Medicago truncatula using real-time (RT) PCR. AB - The quantifications of root colonization and symbiotic activity in the arbuscular mycorrhizal (AM) association of Medicago truncatula and Glomus intraradices were performed by quantitative polymerase chain reaction (real-time PCR). A strong correlation between fungal colonization of the root system and the amounts of fungal rDNA and rRNA were shown. In contrast, the transcript levels of the AM specific phosphate transporter 4 from M. truncatula (MtPT4) correlate with arbuscule formation rather than with fungal colonization. These results suggest (i) that real-time PCR assay is a rapid, useful, and accurate method for the determination of arbuscular mycorrhizal features, (ii) that the amount of fungal rDNA or rRNA is a good parameter to estimate fungal colonization, and (iii) that it is necessary to evaluate the amount of other transcripts-like the MtPT4 transcript-to obtain additional information about the symbiotic state of the colonized root system. PMID- 15658809 TI - Thermoperiodic effect on flowering and endogenous hormonal status in Dendrobium (Orchidaceae). AB - Two year-old cloned plants of Dendrobium Second Love were submitted to 25 degrees C (light) and 10 degrees C (dark) under a 12h photoperiod (60 micromol m(-1)s( 1)) for 30 days. The endogenous levels of IAA, ABA, and the cytokinins Z, [9R]Z, iP, and [9R]iP were measured 15, 22, and 30 days after the start of the thermoperiodic treatment in lateral buds and leaves. The endogenous levels of IAA and cytokinins, especially the zeatin-derived forms, increased significantly in buds after 15 days of treatment. On the other hand, the amount of ABA decreased progressively and significantly throughout the treatment. The treatment conspicuously accelerated flower-bud development. The found correlation suggests that hormones are involved in the signal transduction pathway of thermoperiodic flowering control. PMID- 15658810 TI - [Study on tissue culture of Ficus hirta]. AB - The tissue culture of Ficus hirta Vahl. was studied. The nodes were used as explants and 1/2MS media with different plant growth regulators were tested. The result showed that the adventitious bud differentiation medium was 1/2MS + BA1.0 mg/L + NAA1.0 mg/L and 1/2MS + BA1.5 mg/L + NAA0.5 mg/L, the media for multiplication was 1/2MS +6 - BA0.5 mg/L, and the medium for rooting was 1/2MS + IBA1 mg/L. The cause of nigrescence in the tissue culture and its preventive methods were also discussed in this paper. PMID- 15658811 TI - [Habit and control of pests in Santalum album]. AB - The habit of 5 species pests from South China Botanical Garden was reported in this paper, they are Delias aglaia Linni, Zenzera coffeae Nietner, Parlatoria pergandii Comstock, Scarab (grub), Agrotis ypsilon Rottemberg. Their control methods were presented. PMID- 15658812 TI - [Comparative analysis on effective compositions from tissue-cultured and wild Sophora tonkinensis]. AB - The qualitative analysis of the effective composition, matrine and oxymatrine in tissue-cultured and wild Sophora tonkinsis was done through thin-layer chromatography. The results showed that the tissue-cultured plants were almost as same as the wild plants in the ability to bio-synthesize matrine and oxymatrine. So, the tissue-cultured plants could be used as planting materials for artificial cultivation. PMID- 15658813 TI - [Resource investigation and exploitable foreground of Sarcandra glabra]. AB - The investigation showed the wild resource of Sarcandra glabra (Thumb.) Nakai in the part region of South China including Guangdong, Guangxi and Jiangxi was very plentiful and had a wonderful developmental foreground. PMID- 15658814 TI - [Studies on the fingerprint and quantitative analyzes of quercetin from Aster tataricus]. AB - OBJECTIVE: To establish a qualitative and quantitative method with RP-HPLC for controlling the quality of Aster tataricus L. f. METHODS: The experimental conditions of the RP-HPLC method were as follows: Ultrasphere ODS C18 columns (250 mm x 4.6 mm, 5 microm), and the mobile phase methanol-acetonitrile (1:2) 0.4% phosphoric acid, the flow rate was 1.0 ml/min, the detection wave-lengths was at 360 nm and the column temperature was 25 degrees C. The qualitative fingerprint of the samples dealt by ultrasonic bath and the quantitative measurement of quercetin were finished under the selected chromatographic conditions. RESULTS: Under the qualitative conditions, the 10 mutual peaks in RP HPLC fingerprint of the samples from 6 different Aster tataricus L. f. can be used as index peaks for qualitative identification. In RP-HPLC quantitative analysis, the contents of quercetin in Aster tataricus L. f. were 0.135% approximately 0.246%. CONCLUSION: The analytical method is simple, quick and accurate for controlling the quality of the medicinal material of Aster tataricus L. f. PMID- 15658815 TI - [Studies on constituents from Taxus mairei Bark]. AB - OBJECTIVE: To find the active constituents from Taxus chinensis var. mairei. METHODS: The constituents were isolated and purified by supercritical carbon dioxide extraction and column chromatography. Their structures were determined primarily in the basis of their 1H-NMR, 13C-NMR and DEPT, MS, IR. RESULTS AND CONCLUSIONS: Three compounds were isolated from Taxus mairei and identified as taxinine E, taxol and baccatin III. Taxinine E and baccatin III were found from Taxus chinensis var. mairei for the first time. PMID- 15658816 TI - [Studies on chemical constituents in essential oil from wild Atractylodes lancea in dabie mountains]. AB - The chemical constituents in essential oil of wild Atractylodes lancea (Thumb.) DC. from Dabie Mountains were studied. The essential oil was extracted by simultaneous distillation and extraction equipment and analyzed by gas chromatography-mass spectrometry method. 49 compounds were identified and they represent 92.49% of the total peak aeras. The extraction method and solvent selection of the essential oil were studied. The oil yields and major constituents from Atractylodes lancea (Thumb.) DC. from Dabie Mountains were contrasted to that had been reported. The results showed that Atractylodes lancea (Thumb.) DC. from Dabie Mountains had more essential oil (10.14%) and there were more constituents in it. The main constituents in the essential oil were hinesol, beta-eudesmol, 1H-cyclopropa (a) naphthalene, 1a,2,3,5,6,7,7a,7b-octaphdro 1,1,7,7a-tetramethyl-, [1aR-(1a. alpha, 7. alpha, 7a. alpha,7b. alpha-)] -and gamma-eudesmol. Both the major constituents and contents were different from that had been reported. PMID- 15658817 TI - [Comparative study on triterpenes in different Ganoderma species]. AB - The triterpene profiles of different Ganoderma products was analysed by HPLC. The results showed that the seven-day fermented mycelia had little triterpene and different growth stages fruiting bodies almost had changeless triterpene. Compared with the fruiting body, the triterpene level of spore was lower. Among different Ganoderma species, Ganoderma atrum had little triterpenes too. PMID- 15658818 TI - [Study on coumarin compounds from Exocarpium Citri Grandis]. AB - OBJECTIVE: To study the coumarin compounds from Exocarpium Citri Grandis. METHOD: The constituents were isolated by column chromatograph and identified by spectral data. RESULT: Two coumarin compounds were isolated from the non-volatile section of Exocarpium Citri Grandis and identified as isoimperator and bergapten. CONCLUSION: The two compounds were isolated from the plant for the first time. PMID- 15658819 TI - [Determination of alpha-cyperone in Rhizoma Cyperi oil by HPLC]. AB - OBJECTIVE: To establish a method deteminating alpha-cyperone in Rhizome Cyperi oil by HPLC. METHODS: The determination was carried out by Merck Lichrocart C18 column, with mobile phase consisted of methanol-water (68: 32), and detective wavelength 241 nm. RESULTS: The calibration curves was linear in the range of 0. 081 - 0. 81 microg/ml (r = 0.9998). The average recovery of loading sample was 99.1%, RSD =2.2%. CONCLUSION: The method is convenient for a good resolution. PMID- 15658820 TI - [Content determination of psoralen in root of Ficus hitra by HPLC method]. AB - OBJECTIVE: To investigate the contents of psoralen in Ficus hitra. METHODS: High Performance Liquid Chromatography was performed on Diamonsil C18. The chromatographic conditions were as follows: methanol-water (60:40) as mobile phase, flow rate being 1 ml/min and detecting wavelength at 245nm. RESULTS: A good linearity of psoralen was shown in range of 0. 356 - 2. 848 mg/ml (r = 0.9999), the average recovery is 103.99%, RSD = 3.07% (n = 5). The method has good reproducibility, RSD = 1.59% CONCLUSION: This method can supply evidence for the quality evaluation of Ficus hitra. PMID- 15658822 TI - [Effect of radix Rehmanniae preparata on the expression of c-fos and NGF in hippocampi and learning and memory in rats with damaged thalamic arcuate nucleus]. AB - OBJECTIVE: To study the effect of Radix Rehmanniae Preparata (RRP) on improving the function of learning and memory. METHOD: With injured thalamic arcuate nucleus rats induced by MSG, the improving function of RRP on learning and memory was observed by step down task and Morris water maze task, and the expression of c-fos and NGF in hippocampi was observed by immunohistochemical means. RESULT: RRP could decrease the times of mistakes and prolong the incubation period in step down task, shorten the incubation period of seeking the platform, and improve the rate through the platform position in Morris water maze task. RRP could also increase the expression of hippocampal NGF, c-fos. CONCLUSION: RRP could improve the function of learning and memory of MSG rats, and its mechanism may by related with the increase of the expression of hippocampal c-fos and NGF. PMID- 15658821 TI - [The study of committed differentiation from adult rMSCs into neuron-like cells induced by Xiangdan injection in vitro]. AB - OBJECTIVE: To study the induction of Xiangdan Injection (XDI) on the committed differentiation into neuron from rMSCs. METHODS: The 5th passage of rMSCs were pretreated with alpha-MEM and 10 microg/L basic fibroblast growth factor (bFGF) for 24h, and treated with the serum-free induction media (D/F12 + N2 + bFGF (10 microg/L) and 1% - 5% XDI). The differentiated cells were observed with phase contrast microscopy and detected the expression of several specific proteins, such as neurofilament-200 (NF-200), neuron-specific enolase (NSE), microtubule associated protein-2 (MAP-2) and glial fibrillary acidic protein (GFAP) with immuno-cytochemistry and semi-quantity reverse transcriptase-polymerase chain reaction (RT-PCR). RESULTS: At 3h of induction, rMSCs gradually turned into the neuron-like cells in the induction groups. The induction rate can reach 83.5%+/ 3. 8% at 6 - 8 h of induction. The percentages of responsive cells positive for NSE,NF-200 and GFAP were 82.9%+/-2.98%, 84.1%+/-4.45% and 3.49%+/-1.67% respectively. RT-PCR showed that the relative level of expression for NSE and MAP 2 was much higher than that of GFAP (over 33 fold) (P <0. 01). The unresponsive cells in the induction group and those cultured cells in the control were shown negative for all of the for- mer detected markers. CONCLUSIONS: The rMSCs could be differentiated into neuron-like cells in vitro with the induction of XDI in the free-serum media. PMID- 15658823 TI - [Effect of Yigu capsule on NO and NOS of rat osteoblasts in vitro]. AB - OBJECTIVE: The effects of Yigu Capsule on NO and NOS of osteoblasts were studied. METHODS: (1) Forty 12-month old Sprague-daweley female rats were divided into randomly four groups (Yigu Capsule high dose group, medium dose group and low dose group; saline group), the drug-containing serum and control serum were prepared. (2) The new-born Sprague-daweley rat Osteoblasts were cultured with different Yigu Capsule drug-containing serum concentration and at different exposure time. The mRNA levels of eNOS and iNOS were measured by RT-PCR; the activities of eNOS and iNOS were measured by test kit at different exposure time. NO secretion of osteoblasts was observed, too. RESULTS: The mRNA levels and the activities of eNOS and iNOS and NO secretion of osteoblasts were higher than the control groups respectively after 48,72 and 96 hours (P < 0.01 or P <0.05). CONCLUSIONS: Yigu Capsule drug-containing serum can promote NOS mRNA and NOS activities expressions and NO secretion of osteoblasts. These results may be parts of the mechanism of Yigu Capsule preventing and treating osteoporosis. PMID- 15658824 TI - [Protection of vascular endothelial cells from TNF-alpha induced injury by quercetin]. AB - OBJECTIVE: To investigate the effects of quercetin (Que) on the TNF-alpha injured human umbilical vein endothelial cell line (ECV-304). METHODS: The experiments were performed in culture of TNF-alpha-induced ECV-304 in vitro. Cell viability was assessed with MTT assay. Lipid peroxidation of ECV-304 was monitored as malonaldehyde (MDA) with a fluorometric assay. NO and SOD of ECV-304 were monitored with colorimetry. The expression of NF-kappaB was detected with immunocytochemistry. RESULTS: TNF-alpha inhibited the ECV-304 prolifteration. Preincubation of ECV-304 with Que for 24h before TNF-alpha exposure increased the cell viability. Que significantly decreased the level of NO and MDA and the expression of NF-kappaB, and increased the activity of SOD in TNF-alpha-induced ECV-304. CONCLUSION: These results demonstrate that Que can produce the protective action on TNF-alpha-induced cultured ECV-304 and its mechanism of action may be related to the decrease of NO, antioxidant effect of lipids. However, antioxidant effect might be exerted through NF-kappaB activation pathway. PMID- 15658825 TI - [The study on the effect of LBP on cleaning hydroxygen free radical by EPR technique]. AB - OBJECTIVE: To study the effect of Lycium barbarum polysaccharides (LBP) on cleaning hydroxygen free radical. METHODS: EPR. RESULT: When the concentrations of LBP were 0.25 mg/ml and 1.0 mg/ml, the cleaning rates of LBP to hydroxygen free radical were 18.64% and 87.29%. CONCLUSION: LBP in low concentration can obviously clean hydroxygen free radical, but its clearance ability in high concentration descends. PMID- 15658826 TI - [Effects of flavone from leaves of Diospyros kaki on rat vascular smooth muscle cells proliferation stimulated by native low-density lipoprotein in vitro]. AB - OBJECTIVE: To observe whether rat vascular smooth muscle cells (VSMCs) proliferation induced by native low-density lipoprotein (LDL) is affected by flavone from leaves of Diospyros kaki in vitro. METHODS: Rat aortic VSMCs were cultured in vitro and treated with LDL and flavone from leaves of Diospyros kaki, respectively, and were observed in comparison with the control group. The ratio of cell proliferation was determined by non-radioactive MTS/PES assay. RESULTS: Compared with control, flavone from leaves of Diospyros kaki can dose-dependently inhibit LDL-stimulated vascular smooth muscle cells proliferation (P < 0.05). CONCLUSIONS: The flavone from leaves of Diospyros kaki can inhibit proliferation of rat vascular smooth muscle cells exposed to high levels of native LDL. Flavone from leaves of Diospyros kaki may exert vascular protection by inhibiting vascular smooth muscle cell growth associated with hypercholesterolemia. PMID- 15658828 TI - Strong under pressure. Interview by Colin Parish. PMID- 15658829 TI - A new voice for employers. Interview by Colin Parish. PMID- 15658830 TI - Encouraging nature to act. PMID- 15658831 TI - Hands on care. Interview by Elizabeth Lynch. PMID- 15658832 TI - Maternal instinct. PMID- 15658833 TI - Good scents. PMID- 15658834 TI - Heart of the matter. PMID- 15658835 TI - Falling stars. PMID- 15658836 TI - Implementing a multidisciplinary approach to comprehensive critical care. AB - Following recent government recommendations on critical care services, the skills and training opportunities within the critical care environment should be shared with staff on the general wards. This article discusses a one-day study course offered at one NHS trust that was developed to address the educational needs of staff working on general wards. PMID- 15658837 TI - Estimating date of discharge at ward level: a pilot study. AB - BACKGROUND: Reimbursement is part of the government's strategy to reduce the level of delayed patient discharge from hospital. This article describes a pilot study, undertaken on one ward in a large NHS teaching trust, to involve clinicians in estimating a date of discharge for patients, to improve discharge practice and assist the reimbursement process. Since January 2004, if a patient is not fit for discharge within a day of being designated for discharge, social services has to reimburse the acute trust up to 120 pounds sterling per day for the delay, if the reasons for the delay were attributed to a delay in the provision of service. The barriers to implementing this initiative at ward level are discussed and suggestions made for a pragmatic way forward to enable a process for estimating a date of discharge to work in practice. CONCLUSION: Estimating a date for discharge requires two fundamental steps: a clinical process to estimate and/or document a date of predicted medical fitness, followed by a communication process to document an estimated date of discharge. Effective discharge planning leading to a reduction in delayed discharges will not occur without these two steps. PMID- 15658838 TI - Visual impairment in older people: the nurse's role. AB - This article provides an overview of the role of the nurse in the treatment and management of several common ocular conditions that cause visual impairment in older people. PMID- 15658839 TI - A&E target and PbR hit trusts with finance double whammy. PMID- 15658840 TI - DoH under pressure to tackle perverse finance incentives. PMID- 15658841 TI - Why the NHS is facing its bleakest outlook in years. PMID- 15658842 TI - Freedom fighter. Interview by Nick Edwards. PMID- 15658843 TI - Good management. Make boards work better. AB - Performance reviews for trust boards are rare, despite their strategic importance. Cohesion between executives and non-executives, and willingness for board members to challenge one another, are critical. Too much challenge and too little trust can suppress innovation. PMID- 15658844 TI - On clinical management. PMID- 15658845 TI - 2004 American Chemical Society Award for Computers in Chemical and Pharmaceutical Research. From diatomics to drugs and distributions. PMID- 15658846 TI - Studies toward the pharmacophore of salvinorin A, a potent kappa opioid receptor agonist. AB - Salvinorin A (1), from the sage Salvia divinorum, is a potent and selective kappa opioid receptor (KOR) agonist. We screened other salvinorins and derivatives for binding affinity and functional activity at opioid receptors. Our results suggest that the methyl ester and furan ring are required for activity but that the lactone and ketone functionalities are not. Other salvinorins showed negligible binding affinity at the KOR. None of the compounds bound to mu or delta opioid receptors. PMID- 15658847 TI - Generation of "neoheparin" from E. coli K5 capsular polysaccharide. AB - Heparin remains a major drug in prevention of thromboembolic disease. Concerns related to its animal source have prompted search for heparin analogues. The anticoagulant activity of heparin depends on a specific pentasaccharide sequence that binds antithrombin. We report the generation of a product with antithrombin binding, anticoagulant, and antithrombotic properties similar to those of heparin, through combined chemical and enzymatic modification of a bacterial (E. coli K5) polysaccharide. The process is readily applicable to large-scale production. PMID- 15658848 TI - Discovery of 5-arylsulfonamido-3-(pyrrolidin-2-ylmethyl)-1H-indole derivatives as potent, selective 5-HT6 receptor agonists and antagonists. AB - 5-Arylsulfonylamido-3-(pyrrolidin-2-ylmethyl)-1H-indoles have been identified as high-affinity 5-HT(6) receptor ligands. Within this class, several of the (R) enantiomers were potent agonists having EC(50) values of 1 nM or less and functioning as full agonists while the (S)-enantiomers displayed moderate antagonist activity. PMID- 15658849 TI - Synthesis, characterization, and plasma lipoprotein association of a nucleus targeted boronated porphyrin. AB - The efficacy of binary cancer therapies such as BNCT and PDT depends critically on the subcellular localization site of the sensitizer. This work presents the synthesis and plasma lipoprotein binding properties of the first reported binary conjugate of a boronated porphyrin with a peptide nuclear localization sequence. The porphyrin-NLS conjugate associates in vitro predominantly with low density lipoproteins. Such association provides a potentially selective entry pathway into malignant cells that overexpress the LDL receptor. PMID- 15658850 TI - Rational approach to discover multipotent anti-Alzheimer drugs. AB - The coupling of two different pharmacophores, each endowed with different biological properties, afforded the hybrid compound lipocrine (7), whose biological profile was markedly improved relative to those of prototypes tacrine and lipoic acid. Lipocrine is the first compound that inhibits the catalytic activity of AChE and AChE-induced amyloid-beta aggregation and protects against reactive oxygen species. Thus, it emerged as a valuable pharmacological tool to investigate Alzheimer's disease and as a promising lead compound for new anti Alzheimer drugs. PMID- 15658851 TI - Selective estrogen receptor modulators with conformationally restricted side chains. Synthesis and structure-activity relationship of ERalpha-selective tetrahydroisoquinoline ligands. AB - We disclose herein the discovery of estrogen receptor alpha (ERalpha) selective estrogen receptor modulators (SERMs) of the tetrahydroisoquinoline series that incorporate novel conformationally restricted side chains as replacement of the aminoethoxy residue typical of SERMs. Molecular modeling studies used in conjunction with the X-ray crystal structure of the ERalpha ligand binding domain (LBD) with raloxifene (7) suggested a diazadecaline moiety as a viable mimic of the SERM side chain. On the basis of this knowledge, the piperidinylethoxy moiety of our lead compound 60 was replaced by a diazadecaline subunit, providing the novel tetrahydroisoquinoline 29. In addition to exhibiting a binding affinity to ERalpha and antagonistic properties in the estrogen response element and MCF-7 assays similar to those of the parent compound 60, ligand 29 showed a reduced agonist behavior in the MCF-7 assay in the absence of 17beta-estradiol. These data point toward the fact that 29 may have a potential for breast cancer prevention/treatment in vivo, a feature which is particularly attractive in the quest for safe alternatives to hormone replacement therapy. In a pharmacokinetic experiment carried out in rats, 29 displayed an interesting profile, with a bioavailability of 49%. We also disclose the X-ray crystal structure of 29 in complex with ERalpha-LBD, which reveals the preferred configurations of 29 at the two chiral centers and the details of its interactions with the receptor. Finally, our structure-activity relationship studies show that other analogues bearing constrained side chains retain potency and antagonist activity and that a 3-OH substituted phenyl D-ring increases the selectivity of a set of piperazinyl containing ligands in favor of ERalpha over ERbeta. PMID- 15658852 TI - Stereoselective synthesis of [L-Arg-L/D-3-(2-naphthyl)alanine]-type (E)-alkene dipeptide isosteres and its application to the synthesis and biological evaluation of pseudopeptide analogues of the CXCR4 antagonist FC131. AB - L,L-Type and L,D-type (E)-alkene dipeptide isosteres (EADIs) that have unnatural side chains at the alpha-position were synthesized by the combination of stereoselective aziridinyl ring-opening reactions and organozinc-copper-mediated anti-S(N)2' reactions toward a single substrate of gamma,delta-cis-gamma,delta epimino (E)-alpha,beta-enoate. The utility of this methodology was demonstrated by the stereoselective synthesis of a set of diastereomeric EADIs of L-Arg-L/D-3 (2-naphthyl)alanine (Nal) that is contained in a small CXCR4 antagonist FC131 [cyclo(-D-Tyr-Arg-Arg-Nal-Gly-)]. Furthermore, a (Nal-Gly)-type EADI was synthesized by samarium diiodide (SmI(2))-induced reduction of a gamma-acetoxy alpha,beta-enoate. Several FC131 analogues, in which these EADIs were inserted for reduction of their peptide character, were synthesized with analogues containing reduced amide-type dipeptide isosteres to investigate the importance of these amide bonds for anti-HIV and CXCR4-antagonistic activity. PMID- 15658853 TI - Aminoimidazolylmethyluracil analogues as potent inhibitors of thymidine phosphorylase and their bioreductive nitroimidazolyl prodrugs. AB - Thymidine phosphorylase (TP) is an important target enzyme for cancer chemotherapy because it is expressed at high levels in the hypoxic regions of many tumors and inhibitors of TP have been shown in animal model studies to inhibit angiogenesis and metastasis, and to promote tumor cell apoptosis. The 5 halo-6-[(2'-aminoimidazol-1'-yl)methyl]uracils (3, X = Cl, Br) are very potent inhibitors of E. coli and human TP with IC(50) values of approximately 20 nM when the enzyme concentration is approximately 40 nM. Their 4'-aminoimidazol-1'-yl analogues (4, X = Cl, Br) are >350-fold less active with IC(50) values of approximately 7 microM. The 5-unsubstituted analogues (3 and 4, X = H) were both less active than their 5-halo derivatives. Determination of pK(a) values and molecular modeling studies of these compounds in the active site of human TP was used to rationalize their activities. The finding that 3, X = Br has a poor pharmacokinetic (PK) profile in mice, coupled with the desire for tumor selectivity, led us to design prodrugs. The corresponding 2'-nitroimidazol-1' ylmethyluracils (5, X = Cl, Br) are >1000-fold less active (IC(50) 22-24 microM) than their 2'-amino analogues and are reduced to the 2'-amino inhibitors (3, X = Cl, Br) by xanthine oxidase (XO). As XO is also highly expressed in many tumors, the 2'-nitro prodrugs have the potential to selectively deliver the potent 2' aminoimidazol-1'-yl TP inhibitors into hypoxic solid tumors. PMID- 15658854 TI - Fragment-based lead discovery using X-ray crystallography. AB - Fragment screening offers an alternative to traditional screening for discovering new leads in drug discovery programs. This paper describes a fragment screening methodology based on high throughput X-ray crystallography. The method is illustrated against five proteins (p38 MAP kinase, CDK2, thrombin, ribonuclease A, and PTP1B). The fragments identified have weak potency (>100 microM) but are efficient binders relative to their size and may therefore represent suitable starting points for evolution to good quality lead compounds. The examples illustrate that a range of molecular interactions (i.e., lipophilic, charge charge, neutral hydrogen bonds) can drive fragment binding and also that fragments can induce protein movement. We believe that the method has great potential for the discovery of novel lead compounds against a range of targets, and the companion paper illustrates how lead compounds have been identified for p38 MAP kinase starting from fragments such as those described in this paper. PMID- 15658855 TI - Identification of novel p38alpha MAP kinase inhibitors using fragment-based lead generation. AB - We describe the structure-guided optimization of the molecular fragments 2-amino 3-benzyloxypyridine 1 (IC(50) 1.3 mM) and 3-(2-(4-pyridyl)ethyl)indole 2 (IC(50) 35 microM) identified using X-ray crystallographic screening of p38alpha MAP kinase. Using two separate case studies, the article focuses on the key compounds synthesized, the structure-activity relationships and the binding mode observations made during this optimization process, resulting in two potent lead series that demonstrate significant increases in activity. We describe the process of compound elaboration either through the growing out from fragments into adjacent pockets or through the conjoining of overlapping fragments and demonstrate that we have exploited the mobile conserved activation loop, consisting in part of Asp168-Phe169-Gly170 (DFG), to generate significant improvements in potency and kinase selectivity. PMID- 15658857 TI - Quantitative structure-activity relationship analysis of inhibitors of the nicotine metabolizing CYP2A6 enzyme. AB - The purpose of this study was to develop screening and in silico modeling methods to obtain accurate information on the active center of CYP2A6, a nicotine oxidizing enzyme. The inhibitory potencies of 26 naphthalene and 16 non naphthalene derivatives were determined for human CYP2A6 and mouse CYP2A5 enzymes. Several comparative molecular field analysis (CoMFA) models were developed to find out what types of steric and electrostatic properties are required for potent inhibitors. The IC(50) values of the tested compounds varied from 0.55 to 35 400 microM for CYP2A6 and from 1 to 1500 microM for CYP2A5. The generated CoMFA models were able to accurately predict the inhibition potencies of an external test set of chemicals. Potent and specific inhibitors of the CYP2A6 enzyme can be used in the future to increase nicotine bioavailability and thus make oral nicotine administration feasible in smoking cessation therapy. PMID- 15658856 TI - Potent 4-aryl- or 4-arylalkyl-substituted 3-isoxazolol GABA(A) antagonists: synthesis, pharmacology, and molecular modeling. AB - We have previously described a series of competitive GABA(A) antagonists derived from the low-efficacy partial agonist 5-(4-piperidyl)-3-isoxazolol (4-PIOL, 4). The 2-naphthylmethyl analogue, 4-(2-naphthylmethyl)-5-(4-piperidyl)-3-isoxazolol (5), provided affinity for the GABA(A) receptor site higher than that of the standard GABA(A) receptor antagonist, SR 95531 (3). Molecular modeling studies of these compounds exposed a cavity at the receptor recognition site capable of accommodating aromatic groups of substantial size in the 4-position in the 3 isoxazolol ring. Here we present a series of analogues of 5, with various substituents in different positions in the naphthyl ring system (6a-k), and compounds with aromatic substituents directly attached to the 4-position of the 3 isoxazolol ring (7l-n). The compounds have been pharmacologically characterized using receptor-binding assays and electrophysiological whole-cell patch-clamp techniques. All of the tested compounds show affinity for the GABA(A) receptor site. While the 5-, 7-, and 8-bromo analogues, 6b-d, showed receptor affinities (K(i) = 45, 109, and 80 nM, respectively) comparable with that of 5 (K(i) = 49 nM), the 1-bromo analogue, 6a, provided the highest receptor affinity of the series (K(i) = 10 nM). Introduction of a series of different substituents in the 1-position in the 2-naphthyl ring system led to compounds, 6e-k, with retained high affinity for the GABA(A) receptor (K(i) = 16-250 nM). Introduction of a phenyl ring directly into the 4-position on the 3-isoxazolol ring gave a 41-fold increase in affinity relative to that of 4-PIOL. In whole-cell patch-clamp recordings from cultured cerebral cortical neurons, all of the tested compounds were able to inhibit the effect of the specific GABA(A) agonist isoguvacine, 6a showing antagonist potency (IC(50) = 42 nM) markedly higher than that of 3 (IC(50) = 240 nM). Molecular modeling studies, based on the compounds described, emphasized the importance of the distal ring in 5 for receptor affinity and the considerable dimensions of the proposed receptor cavity. Furthermore, the phenyl rings in 7l and in 6k were shown to represent highly favorable positions for an aromatic ring in previously unexplored receptor regions in terms of a pharmacophore model. PMID- 15658858 TI - Synthesis and antiviral evaluation of cis-substituted cyclohexenyl and cyclohexanyl nucleosides. AB - Starting from commercially available (rac)-3-cyclohexene-1-carboxylic acid, a series of purine and pyrimidine cis-substituted cyclohexenyl and cyclohexanyl nucleosides were synthesized through a key Mitsunobu reaction. Antiviral evaluations were performed on HIV, coxsackie B3, and herpes viruses (HSV-1, HSV 2, VZV, HCMV). Three compounds showed moderate activity against HSV-1 and coxsackie viruses. Specific computer modeling studies were performed on HSV-1 thymidine kinase in order to understand the enzyme activation of an analogue showing moderate antiviral activity. PMID- 15658859 TI - Quantitative structure-activity relationship (5D-QSAR) study of combretastatin like analogues as inhibitors of tubulin assembly. AB - A molecular modeling study was carried out to develop a predictive model for combretastatin-like analogues populating the colchicine-binding site of beta tubulin. A series of compounds built around a framework including two aromatic groups linked by various moieties such as alkenes (stilbenes), enones (chalcones), or ethers was selected for the study. The 5D-QSAR model was developed stepwise. First a model was generated for the chalcone series (19 compounds, 71 conformations), then for the stilbene series (18 compounds, 59 conformations), and finally for the combined dataset (47 ligands, 160 conformers). Although the models for the chalcone and stilbene series appeared slightly different when represented by QSAR colored surfaces, the combined model seems to reconcile the differences without compromise and represents a highly predictive model for compounds that bind to the colchicine-binding site of tubulin. PMID- 15658860 TI - Investigations on the effects of basic side chains on the hormonal profile of (4R,5S)/(4S,5R)-4,5-bis(4-hydroxyphenyl)-2-imidazolines. AB - Basic side chains determine the pharmacology of selective estrogen receptor modulators such as tamoxifen or raloxifene. In this study we tried to turn the hormonal profile of (4R,5S)/(4S,5R)-4,5-bis(4-hydroxyphenyl)-2-imidazolines from agonistic to antagonistic by introduction of a dimethylaminoethane, a piperidin-1 ylethane, or a pyrrolidin-1-ylethane side chain into one of the 4-hydroxyphenyl rings. The compounds were tested for agonistic and antagonistic activity on hormone sensitive, ERalpha-positive MCF7-2a cells, stably transfected with the plasmid ERE(wtc)luc and on U-2 OS cells transiently transfected with plasmids encoding for ERalpha (pSG5-ERalpha) or ERbeta (pSG5-ERbeta FL) as well as the reporter plasmid (ERE)(2)luc(+). Despite the presence of a basic side chain, the majority of the 4,5-diaryl-2-imidazolines showed agonistic effects. The most active compound, (4R,5S)/(4S,5R)-4-(2-chloro-4-(2-piperidin-1-ylethoxy)phenyl)-5 (2,6-dichloro-4-hydroxyphenyl)-2-imidazoline (5a), achieved at ERalpha an EC(50) value of 0.085 microM and at ERbeta an EC(50) = 0.40 microM. High antagonistic properties only possessed the C2 ethyl substituted compounds 2a and 4a. (4R,5S)/(4S,5R)-2-Ethyl-4-(4-hydroxyphenyl)-5-(4-(2-piperidin-1-ylethoxy)phenyl) 2-imidazoline (2a) reduced the effect of estradiol at ERalpha strongly with IC(50) = 0.038 microM, while its antagonistic properties at ERbeta were distinctly lower (IC(50) = 9.00 microM), probably due to the partial agonistic effects (EC(50) = 0.50 microM). PMID- 15658861 TI - The novel L- and D-amino acid derivatives of hydroxyurea and hydantoins: synthesis, X-ray crystal structure study, and cytostatic and antiviral activity evaluations. AB - The novel L- and D-amino acid derivatives of hydroxyurea 5a-o were prepared by aminolysis of N-(1-benzotriazolecarbonyl)amino acid amides 4a-o with hydroxylamine. The hydantoin derivatives 6a-e,m,p were synthesized by base catalyzed cyclization of amides 4, common precursors for 5 and 6. X-ray crystal structure analysis shows that the C5 atom in 6e possesses the S configuration, which is consistent with the configuration of the starting reagent, l-leucine. Among L-amino acid derivatives of hydroxyurea, 5h and 5i inhibited specifically murine leukemia and human T-lymphocytes (IC(50) = 10-19 microM) and showed selectivity with respect to normal human fibroblasts (WI 38). d-Amino acid derivatives of hydroxyurea 5m and 5o inhibited the growth of all tumor cell lines (IC(50) = 4.8-83.9 microM), but not the growth of normal fibroblasts (WI 38; IC(50) > 100 microM). Results on antiviral evaluations showed that N-(1 benzotriazolecarbonyl)amino acid amide 4m and hydantoin 6m had marked activity against the Davis strain of CMV (4m, EC(50) = 3.2 microg/mL; 6m, EC(50) = 4.0 microg/mL). However, these compounds showed also rather expressed cytotoxicity (4m, CC(50) = 43.4 microg/mL; 6m, CC(50) = 12.5 microg/mL(-1)). PMID- 15658862 TI - Endothelin-converting enzyme-1 inhibition and growth of human glioblastoma cells. AB - Endothelin-1 (ET-1) is mitogenic and/or antiapoptotic in human cancers, and antagonists to ET-1 receptors are under evaluation for cancer treatment. Inhibition of ET-1 activation by the endothelin-converting enzymes 1(a)(-)(d) (ECE-1(a)(-)(d); EC 3.4.24.71) represents another approach to block the ET-1 effect in cancer. To evaluate this potential, we synthesized and characterized a series of low nanomolar nonpeptidic thiol-containing ECE-1 inhibitors, and evaluated their effect, as well as the effect of inhibitors for the related metalloproteases neprilysin (NEP; EC 3.4.24.11) and angiotensin-converting enzyme (ACE; EC 3.4.15.1), on human glioblastoma cell growth. Only ECE-1 inhibitors inhibited DNA synthesis by human glioblastoma cells. Exogenous addition of ET-1 or bigET-1 to glioblastoma cells did not counterbalance the growth inhibition elicited by ECE-1 inhibitors, suggesting that ECE-1 inhibitors block the proliferation of human glioblastoma cells most likely via a mechanism not involving extracellular production of ET-1. This class of molecules may thus represent novel therapeutic agents for the potential treatment of human cancer. PMID- 15658864 TI - Somatostatin receptor 1 selective analogues: 2. N(alpha)-Methylated scan. AB - Des-AA(1,2,5)-[d-Trp(8)/d-Nal(8),IAmp(9)]SRIF (AA = amino acid, Nal = 3-(2 naphthyl)-alanine, IAmp = 4-(N-isopropyl)-aminomethylphenylalanine, SRIF = somatostatin), with or without a tyrosine or monoiodotyrosine, were scanned with the introduction of a backbone N-methyl group and tested for binding affinity at the five human somatostatin receptors (sst(1)(-)(5)). N(alpha)-Methylation resulted in loss of sst affinity (2- to >5-fold) when introduced at residues Lys(4) (6), Phe(6) (7), Phe(7) (8), Thr(10) (11), and Phe(11) (12) of the parent compound Des-AA(1,2,5)-[d-Nal(8),IAmp(9)]SRIF (4). N(alpha)-Methylation was tolerated at residues Cys(3) (5), d-Nal(8) (9), Thr(12) (13), and Cys(14) (15) with retention of binding sst affinity and selectivity and resulted in an increase in sst binding affinity at positions IAmp(9) (10) and Ser(13) (14). In these series, the d-Trp(8) substitution versus d-Nal(8) is clearly superior. C Terminally lysine-extended analogues (21-25) retained sst(1) selectivity and binding affinity when compared to their d-Nal(8)- (4) or d-Trp(8)- (3) containing parent. Des-AA(1,2,5)-[d-Trp(8), (N(alpha)Me)IAmp(9)]SRIF (17), Des-AA(1,2,5)-[d Trp(8),IAmp(9),(N(alpha)Me)Ser(13)]SRIF (19), Des-AA(1,2,5)-[d Trp(8),IAmp(9),(N(alpha)Me)Cys(14)]SRIF (20), Des-AA(1,2,5)-[d Trp(8),(N(alpha)Me)IAmp(9),Tyr(11)]SRIF (34), and Des-AA(1,2,5)-[d Agl(8)(N(beta)Me,2-naphthoyl),IAmp(9),Tyr(11)]SRIF (42) (Agl = aminoglycine) are sst(1) agonists in their ability to inhibit forskolin-induced cAMP production. PMID- 15658863 TI - Identification of 4-substituted 1,2,3-triazoles as novel oxazolidinone antibacterial agents with reduced activity against monoamine oxidase A. AB - Oxazolidinones represent a new and promising class of antibacterial agents. Current research in this area is mainly concentrated on improving the safety profile and the antibacterial spectrum. Many oxazolidinones, including linezolid (marketed as Zyvox), are inhibitors of monoamine oxidase A (MAO-A), which presents an undesired side effect. Recently, it was found that the 1,2,3-triazole is a good replacement for the conventional acetamide functionality found in oxazolidinones. We now disclose the finding that 1,2,3-triazoles bearing a substituent like methyl, small substituted methyl, bromo, or a linear (sp hybridized) group at the 4 position (compounds such as 5, 16, 19, and 21) are good antibacterials with reduced or no activity, within the detection limit of the assay, against MAO-A. The results are especially promising for the development of oxazolidinones with an improved safety profile. The MAO-A SAR can be rationalized on the basis of docking studies to a MAO-A/MAO-B homology model. PMID- 15658865 TI - Somatostatin receptor 1 selective analogues: 3. Dicyclic peptides. AB - The binding affinity of short chain somatostatin (SRIF) analogues at the five human SRIF receptors (sst) was determined to identify sterically constrained somatostatin receptor subtype 1 (sst(1)) selective scaffolds. Des-AA(1,2,4,13)-[d Trp(8)]SRIF (2) retained high binding affinity at all receptors but sst(1), Des AA(1,2,4,5)-[d-Trp(8)]SRIF (3) at sst(4) and sst(5), and Des-AA(1,2,4,5,13)-[d Trp(8)]SRIF (4) at sst(2) and sst(4) (AA = amino acid). Des-AA(1,2,4,12,13)-[d Trp(8)]SRIF (6) was potent and sst(4)-selective (>25-fold); Des-AA(1,2,5,12,13) [d-Trp(8)]SRIF (7) and Des-AA(1,2,4,5,12,13)-[d-Trp(8)]-SRIF (9, ODT-8) were most potent at sst(4) and moderately potent at all other receptors. Dicyclic SRIF agonists of the sst(1)-selective Des-AA(1,5)-[Tyr(2),d-Trp(8),IAmp(9)]SRIF, (14, sst(1) IC(50) = 14 nM) were prepared in which a lactam bridge introduced additional conformational constraint (IAmp = 4-(N-isopropyl) aminomethylphenylalanine). Cyclo(7-12)Des-AA(1,5)-[Tyr(2),Glu(7),d Trp(8),IAmp(9),hhLys(12)]SRIF (31) (sst(1) IC(50) = 16 nM) and cyclo(7-12) Des AA(1,2,5)-[Glu(7),d-Trp(8),IAmp(9),m-I-Tyr(11),hhLys(12)]SRIF (45) (sst(1) IC(50) = 6.1 nM) had equal or improved affinities over that of the parent 14. Binding affinity was decreased in all other cases with alternate bridging constraints such as cyclo (6-11), cyclo (6-12), and cyclo (7-11). Compound 45 is an agonist (EC(50) = 8.8 nM) in the adenylate cyclase assay. PMID- 15658866 TI - Somatostatin receptor 1 selective analogues: 4. Three-dimensional consensus structure by NMR. AB - The three-dimensional NMR structures of six analogues of somatostatin (SRIF) are described. These analogues with the amino acid 4-(N-isopropyl) aminomethylphenylalanine (IAmp) at position 9 exhibit potent and highly selective binding to human SRIF subtype 1 receptors (sst(1)). The conformations reveal that the backbones of these analogues have a hairpin-like structure similar to the sst(2)-subtype-selective analogues. This structure serves as a scaffold for retaining a unique arrangement of the side chains of d-Trp(8), IAmp(9), Phe(7), and Phe(11) or m-I-Tyr(11) (m-I-Tyr = mono-iodo-tyrosine). The conformational preferences and results from biological analyses of these analogues(1,2) allow a detailed study of the structure-activity relationship of SRIF. The proposed consensus pharmacophore of the sst(1)-selective analogues requires a unique set of distances between an indole/2-naphthyl ring, an IAmp side chain, and two aromatic rings. This motif is necessary and sufficient to explain the binding affinities of all of the analogues studied and is distinct from the existing models suggested for sst(4) as well as sst(2)/sst(5) selectivity. PMID- 15658867 TI - Synthesis and antiviral activity of helioxanthin analogues. AB - A series of natural product analogues based on helioxanthin (2), with particular attention to modification of the lactone ring and methylenedioxy group, were synthesized and evaluated for their antiviral activities. Among them, lactam derivative 18 and helioxanthin cyclic hydrazide 28 exhibited significant in vitro antiviral activity against hepatitis B virus (EC(50) = 0.08 and 0.03 microM, respectively). Compound 18 showed the most potent antiviral activity against hepatitis C virus (55% inhibition at 1.0 microM). Compound 12, an acid-hydrolyzed product of helioxanthin cyclic imide derivative 9, was found to exhibit broad spectrum antiviral activity against hepatitis B virus (EC(50) = 0.8 microM), herpes simplex virus type 1 (EC(50) = 0.15 microM) and type 2 (EC(50) < 0.1 microM), Epstein-Barr virus (EC(50) = 9.0 microM), and cytomegalovirus (EC(50) = 0.45 microM). Helioxanthin lactam derivative 18 also showed marked inhibition of herpes simplex virus type 1 (EC(50) = 0.29 microM) and type 2 (EC(50) = 0.16 microM). The cyclic hydrazide derivative of helioxanthin 28 and its brominated product 42 exhibited moderately potent activities against human immunodeficiency virus (EC(50) = 2.7 and 2.5 microM, respectively). Collectively, these molecules represent a novel set of antiviral compounds with unique structural features. PMID- 15658868 TI - Sulfonamide drugs binding to the colchicine site of tubulin: thermodynamic analysis of the drug-tubulin interactions by isothermal titration calorimetry. AB - The discovery of several sulfonamide drugs paved the way toward the synthesis of 6 (N-[2-[(4-hydroxyphenyl)amino]-3-pyridinyl]-4-methoxybenzenesulfonamide, E7010) and 7 (N-(3-fluoro-4-methoxyphenyl)pentafluorobenzenesulfonamide, T138067), both of which inhibit tubulin polymerization and are under clinical development. A series of diarylsulfonamides containing an indole scaffold was also found to have antimitotic properties, but their mode of interactions with tubulin has remained unidentified so far. In this study, we demonstrate that these sulfonamide drugs bind to the colchicine site of tubulin in a reversible manner. They quenched intrinsic tryptophan fluorescence of tubulin presumably due to drug-induced conformational changes in the protein, but were unable to modulate GTPase activity of tubulin in contrast to colchicine that enhances the same enzymatic activity. Further investigation using isothermal titration calorimetry (ITC) revealed that 5 (N-(5-chloro-7-indolyl)-4-methoxybenzenesulfonamide) afforded a large positive value of heat capacity change (DeltaC(p)() = +264 cal mol(-1) K( 1)) on binding to tubulin, suggesting a substantial conformational transition in the protein along with partial enthalpy-entropy compensation. On the other hand, the 2-chloro regioisomer 2 gave a large negative value of DeltaC(p)() (-589 cal mol(-1) K(-1)) along with complete enthalpy-entropy compensation. This thermodynamic profile was thought to be attributable to a prominent contribution of van der Waals interaction and hydrogen bonding between specific groups in the drug-tubulin complex. These results indicate that a mere alteration in the position of a single substituent chlorine on the indole scaffold has a great influence on the drug-tubulin binding thermodynamics. PMID- 15658869 TI - Further naphthylcombretastatins. An investigation on the role of the naphthalene moiety. AB - By synthesis and biological studies of new naphthalene analogues of combretastatins, we have found that the naphthalene is a good surrogate for the isovanillin moiety (3-hydroxy-4-methoxyphenyl) of combretastatin A-4, always generating highly cytotoxic analogues when combined with the 3,4,5 trimethoxyphenyl or related systems. On the other hand, when the naphthalene replaces the 3,4,5-trimethoxyphenyl moiety, the cytotoxic activity is largely decreased. The most cytotoxic naphthalene analogues of combretastatins, which also produce inhibition of tubulin polymerization, exerted their antimitotic effects through microtubule network disruption and subsequent G(2)/M arrest of the cell cycle in human cancer cells. PMID- 15658870 TI - Selective benzopyranone and pyrimido[2,1-a]isoquinolin-4-one inhibitors of DNA dependent protein kinase: synthesis, structure-activity studies, and radiosensitization of a human tumor cell line in vitro. AB - A diverse range of chromen-2-one, chromen-4-one and pyrimidoisoquinolin-4-one derivatives was synthesized and evaluated for inhibitory activity against the DNA repair enzyme DNA-dependent protein kinase (DNA-PK), with a view to elucidating structure-activity relationships for potency and kinase selectivity. DNA-PK inhibitory activity varied widely over the series of compounds evaluated (IC(50) values ranged from 0.19 to >10 microM), with excellent activity being observed for the 7,8-benzochromen-4-one and pyrimido[2,1-a]isoquinolin-4-one templates. By contrast, inhibitors based on the benzochromen-2-one (coumarin) or 2-aryl-7,8 benzochromen-4-one (flavone) scaffolds were less potent. Crucially, these studies revealed a very constrained structure-activity relationship at the 2-position of the benzopyranone and pyrimido[2,1-a]isoquinolin-4-one pharmacophore, with only a 2-morpholino or 2-(2'-methylmorpholino) group being tolerated at this position. More detailed biological studies conducted with the most potent inhibitor NU7163 (48; IC(50) = 0.19 microM) demonstrated ATP-competitive DNA-PK inhibition, with a K(i) value of 24 nM, and 48 exhibited selectivity for DNA-PK compared with the related enzymes ATM, ATR, mTOR, and PI 3-K (p110alpha). Compound 48 sensitized the HeLa human tumor cell line to the cytotoxic effects of ionizing radiation in vitro, a dose modification factor of 2.3 at 10% survival being observed with an inhibitor concentration of 5 microM. This study identified these structural classes as novel DNA-PK inhibitors and delineated initial structure-activity relationships against DNA-PK. PMID- 15658871 TI - Development of potent mu-opioid receptor ligands using unique tyrosine analogues of endomorphin-2. AB - Six analogues of tyrosine, which contained alkyl groups at positions 2', 3', and 6', either singly or in combination on the tyramine ring, were investigated for their effect on the opioid activity of [Xaa(1)]endomorphin-2 (EM-2). The opioid analogues displayed the following characteristics: (i) high mu-opioid receptor affinity [K(i)(mu) = 0.063-2.29 nM] with selectivity [K(i)(delta)/K(i)(mu)] ranging from 46 to 5347; (ii) potent functional mu-opioid agonism [GPI assay (IC(50) = 0.623-0.924 nM)] and with a correlation between delta-opioid receptor affinities and functional bioactivity using MVD; (iii) intracerebroventricular administration of [Dmt(1)]- (14) and [Det(1)]EM-2 (10) produced a dose-response antinociception in mice, with the former analogue more active than the latter; and (iv) a marked shift occurred from the trans-orientation at the Tyr(1)-Pro(2) bond to a cis-conformer compared to that observed previously with [Dmt(1)]EM-2 (14) (Okada et al. Bioorg. Med. Chem. 2003, 11, 1983-1984) except [Mmt(1)]EM-2 (7). The active profile of the [Xaa(1)]EM-2 analogues indicated that significant modifications on the tyramine ring are possible while high biological activity is maintained. PMID- 15658872 TI - Synthesis and biological study of a new series of 4'-demethylepipodophyllotoxin derivatives. AB - Etoposide (VP-16) is a potent human DNA topoisomerase II poison, derived from 4' demethylepipodophyllotoxin, widely used in cancer chemotherapy. Continuous efforts have driven to synthesize new related compounds, presenting decreased toxic side effects, metabolic inactivation, drug resistance, and increased water solubility. Identified structure-activity relationships have pointed out the importance of the 4beta-substitution and of the configuration of the D ring. Here we report the synthesis of two novel series of derivatives of 4' demethylepipodophyllotoxin. The first bears a carbamate chain in the 4 position (13a-f), whereas, in the second series, in addition to this chain, the lactone ring has been modified by shifting the carbonyl from position 13 to position 11 (27a-f). Moreover, an analogue of TOP-53 having this lactone modification has also been prepared (32). From this study, structure-activity relationships were established. Compounds 13a and 27a displayed potent cytotoxic activity against the L1210 cell line (10 to 20-fold higher than VP-16) and proved to be strong topoisomerase II poisons more potent than VP-16. From preliminary in vivo investigation of both compounds against P388 leukemia and orthotopically grafted human A549 lung carcinoma, it appeared that 13a and 27a constitute promising leads for a new class of antitumor agents. PMID- 15658873 TI - Exploring the role of different drug transport routes in permeability screening. AB - The influence of different drug transport routes in intestinal drug permeability screening assays was studied. Three experimental models were compared: the small intestine-like 2/4/A1 cell model, which has a leaky paracellular pathway, the Caco-2 cell model, which has a tighter paracellular pathway, and artificial hexadecane membranes (HDMs), which exclusively model the passive transcellular pathway. The models were investigated regarding their ability to divide passively and actively transported compounds into two permeability classes and to rank compounds according to human intestinal absorption. In silico permeability models based on two-dimensional (2D) and three-dimensional (3D) molecular descriptors were also developed and validated using external test sets. The cell-based models classified 80% of the acceptably absorbed compounds (FA >/= 30%) correctly, compared to 60% correct classifications using the HDM model. The best compound ranking was obtained with 2/4/A1 (r(s) = 0.74; r(s) = 0.95 after removing actively transported outliers). The in silico model based on 2/4/A1 permeability gave results of similar quality to those obtained when using experimental permeability, and it was also better than the experimental HDM model at compound ranking (r(s) = 0.85 and 0.47, respectively). We conclude that the paracellular transport pathway present in the cell models plays a significant role in models used for intestinal permeability screening and that 2/4/A1 in vitro and in silico models are promising alternatives for drug discovery permeability screening. PMID- 15658874 TI - 4,6-Disubstituted 2,2-dimethylchromans structurally related to the K(ATP) channel opener cromakalim: design, synthesis, and effect on insulin release and vascular tone. AB - Five series (ureas, thioureas, carbamates, sulfonylureas, and amides) of 4,6 disubstituted-2,2-dimethylchromans structurally related to cromakalim were prepared and evaluated, as putative ATP-sensitive potassium channel activators, on rat pancreatic islets and rat aorta rings. The biological data indicate that most compounds were, like the reference molecule cromakalim, more active on the vascular smooth muscle tissue (myorelaxant effect on 30 mM KCl induced contractions of rat aorta rings) than on the pancreatic tissue (inhibition of 16.7 mM glucose induced insulin release from rat pancreatic islets). However, some drugs (8h, 8i, 9f, 9g, 9h, and 9i) markedly inhibited insulin release and exhibited an activity equivalent or greater than that of diazoxide. Compounds 9h and 9i were also found to be more active on pancreatic beta-cells than on vascular smooth muscle cells. Last, the amide 6b was selected in order to examine its mechanism of action on vascular smooth muscle cells. Pharmacological results suggest that the compound acted as a K(ATP) channel opener. In conclusion, the present data indicate that appropriate structural modifications can generate dimethylchromans with pharmacological profiles different from that of cromakalim. PMID- 15658875 TI - Antitumor-active cobalt-alkyne complexes derived from acetylsalicylic acid: studies on the mode of drug action. AB - Cobalt-alkyne complexes are drugs with remarkable cytotoxicity. From the complexes tested up to now we selected the aspirin derivative [2-acetoxy-(2 propynyl)benzoate]hexacarbonyldicobalt (Co-ASS) as the lead compound. To get more insight into the mode of action, we systematically modified the alkyne ligand and determined the cytotoxic properties of the resulting cobalt complexes. Further investigations were performed on the drug lipophilicity, the cellular uptake into MCF-7 and MDA-MB 231 breast cancer cells, the DNA-binding efficacy, and the nuclear drug content. The ability to inhibit glutathione reductase and cyclooxygenase (COX) enzymes, the binding to the estrogen receptor, and the induction of apoptotic processes were examined for selected compounds. Interestingly, the most antitumor active compounds were potent COX inhibitors (COX-1 and COX-2). The presented results indicate that cobalt-alkyne complexes of the Co-ASS type, represent a new class of organometallic cytostatics with a mode of drug action in which COX inhibition probably plays a major role. PMID- 15658876 TI - Comparison of trace metal concentrations in malign and benign human prostate. AB - Imbalance in the composition of trace metals, recognized to be essential to normal human homeostasis, besides the accumulation of potentially toxic or nonessential trace metals, may cause disease. Thus, there is a need for their analysis in cancerous and noncancerous human tissues to examine the relationship between cancer and these elements. Trace metal concentrations including Cd, Ni, Zn, Cu, Fe, Mg, and Ca in both malign and benign prostate samples were determined by atomic absorption spectrophotometry. The tissues were digested by using microwave energy. In contrast to the literature data for zinc, the concentrations of calcium and zinc in the malign human prostate were found to be significantly higher than those in the benign human prostate (p < 0.05 for both metals). Similarly, the concentrations of iron, nickel, and magnesium in the malign prostate were also found to be higher than those in the benign prostate (p 0.05). Levels of apoptosis fluctuated as a function of dose, and modest increases above basal levels persisted throughout the 48-h period. The comparison of retained follicular structures revealed differences in the alpha components of the linear-quadratic dose-response curves (0.60 Gy(-1) for early passage and 0.71 Gy(-1) for late-passage cultures, P < 0.014). Cell cycle phase redistribution resulted in a G2 arrest (P < 0.001) for both early- and late passage cultures. In conclusion, the response of thyroid follicular cells to high LET radiation was not influenced by the presence of gap junctions or the proliferative status of the target cells. PMID- 15658894 TI - Genomic instability in human lymphocytes irradiated with individual charged particles: involvement of tumor necrosis factor alpha in irradiated cells but not bystander cells. AB - Exposure to ionizing radiation can increase the risk of cancer, which is often characterized by genomic instability. In environmental exposures to high-LET radiation (e.g. 222Ra), it is unlikely that many cells will be traversed or that any cell will be traversed by more than one alpha particle, resulting in an in vivo bystander situation, potentially involving inflammation. Here primary human lymphocytes were irradiated with precise numbers of 3He2+ ions delivered to defined cell population fractions, to as low as a single cell being traversed, resembling in vivo conditions. Also, we assessed the contribution to genomic instability of the pro-inflammatory cytokine tumor necrosis factor alpha (TNFA). Genomic instability was significantly elevated in irradiated groups (> or = two fold over controls) and was comparable whether cells were traversed by one or two 3He2+ ions. Interestingly, substantial heterogeneity in genomic instability between experiments was observed when only one cell was traversed. Genomic instability was significantly reduced (60%) in cultures in which all cells were irradiated in the presence of TNFA antibody, but not when fractions were irradiated under the same conditions, suggesting that TNFA may have a role in the initiation of genomic instability in irradiated cells but not bystander cells. These results have implications for low-dose exposure risks and cancer. PMID- 15658895 TI - "Modeled microgravity" affects cell response to ionizing radiation and increases genomic damage. AB - The aim of this work was to assess whether "modeled microgravity" affects cell response to ionizing radiation, increasing the risk associated with radiation exposure. Lymphoblastoid TK6 cells were irradiated with various doses of gamma rays and incubated for 24 h in a modeled microgravity environment obtained by the Rotating Wall Vessel bioreactor. Cell survival, induction of apoptosis and cell cycle alteration were compared in cells irradiated and then incubated in 1g or modeled microgravity conditions. Modulation of genomic damage induced by ionizing radiation was evaluated on the basis of HPRT mutant frequency and the micronucleus assay. A significant reduction in apoptotic cells was observed in cells incubated in modeled microgravity after gamma irradiation compared with cells maintained in 1g. Moreover, in irradiated cells, fewer G2-phase cells were found in modeled microgravity than in 1g, whereas more G1-phase cells were observed in modeled microgravity than in 1g. Genomic damage induced by ionizing radiation, i.e. frequency of HPRT mutants and micronucleated cells, increased more in cultures incubated in modeled microgravity than in 1g. Our results indicate that modeled microgravity incubation after irradiation affects cell response to ionizing radiation, reducing the level of radiation-induced apoptosis. As a consequence, modeled microgravity increases the frequency of damaged cells that survive after irradiation. PMID- 15658896 TI - Radiation-induced mutation at tandem repeat DNA Loci in the mouse germline: spectra and doubling doses. AB - The spectra and dose response for mutations at expanded simple tandem repeat (ESTR) loci in the germline of male mice acutely exposed to low-LET X or gamma rays at pre-meiotic stages of spermatogenesis were compared in five strains of laboratory mice. Most mutation events involved the gain or loss of a relatively small number of repeat units, and the distributions of length changes were indistinguishable between the exposed and control males. Overall, a significant bias toward gains of repeats was detected, with approximately 60% of mutants showing gains. The values for ESTR mutation induction did not differ substantially between strains. The highest values of doubling dose were obtained for two genetically related strains, BALB/c and C.B17 (mean value 0.98 Gy). The estimates of doubling dose for three other strains (CBA/H, C57BL/6 x CBA/H F1 and 129SVJ x C57BL/6) were lower, with a mean value of 0.44 Gy. The dose response for ESTR mutation across all five strains was very close to that for the specific loci (Russell 7-locus test). The mechanisms of ESTR mutation induction and applications of this system for monitoring radiation-induced mutation in the mouse germline are discussed. PMID- 15658897 TI - Radiobiological aspects of intraoperative radiotherapy (IORT) with isotropic low energy X rays for early-stage breast cancer. AB - The purpose of this study was to model the distribution of biological effect around a miniature isotropic X-ray source incorporating spherical applicators for single-dose or hypo-fractionated partial-breast intraoperative radiotherapy. A modification of the linear-quadratic formalism was used to calculate the relative biological effectiveness (RBE) of 50 kV X rays as a function of dose and irradiation time for late-reacting normal tissue and tumor cells. The response was modeled as a function of distance in the tissue based on the distribution of equivalent dose and published dose-response data for pneumonitis and subcutaneous fibrosis after single-dose conventional irradiation. Furthermore, the spatial distribution of tumor cell inactivation was assessed. The RBE for late reactions approached unity at the applicator surface but increased as the absorbed dose decreased with increasing distance from the applicator surface. The ED50 for pneumonitis was estimated to be reached at a depth of 6-11 mm in the tissue and that for subcutaneous fibrosis at 3-6 mm, depending on the applicator diameter and whether the effect of recovery was included. Thus lung tissue would be spared because of the thickness of the thorax wall. The RBE for tumor cells was higher than for late-reacting tissue. The applicator diameter is an important parameter in determining the range of tumor cell control in the irradiated tumor bed. PMID- 15658899 TI - A Monte Carlo code for the simulation of heavy-ion tracks in water. AB - TILDA, a new Monte Carlo track structure code for ions in gaseous water that is valid for both high-LET (approximately 10(4) keV/microm) and low-LET ions, is presented. It is specially designed for a comparison of the patterns of energy deposited by a large range of ions. Low-LET ions are described in a perturbative frame, whereas heavy ions with a very high stopping power are treated using the Lindhard local density approximation and the Russek and Meli statistical method. Ionization cross sections singly differential with energy compare well with the experiment. As an illustration of the non-perturbative interaction of high-LET ions, a comparison between the ion tracks of light and heavy ions with the same specific energy is presented (1.4 MeV/nucleon helium and uranium ions). The mean energy for ejected electrons was found to be approximately four times larger for uranium than for helium, leading to a much larger track radius in the first case. For electrons, except for the excitation cross sections that are deduced from experimental fits, cross sections are derived analytically. For any orientation of the target molecule, the code calculates multiple differential cross sections as a function of the ejection and scattering angles and of the energy transfer. The corresponding singly differential and total ionization cross sections are in good agreement with experimental data. The angular distribution of secondary electrons is shown to depend strongly on the orientation of the water molecule. PMID- 15658898 TI - Modeling multicellular response to nonuniform distributions of radioactivity: differences in cellular response to self-dose and cross-dose. AB - Radiopharmaceuticals are distributed nonuniformly in tissue. While distributions of radioactivity often appear uniform at the organ level, in fact, microscopic examination reveals that only a fraction of the cells in tissue are labeled. Labeled cells and unlabeled cells often receive different absorbed doses depending on the extent of the nonuniformity and the characteristics of the emitted radiations. The labeled cells receive an absorbed dose from radioactivity within the cell (self-dose) as well as an absorbed dose from radioactivity in surrounding labeled cells (cross-dose). Unlabeled cells receive only a cross dose. In recent communications, a multicellular cluster model was used to investigate the lethality of microscopic nonuniform distributions of 131I iododeoxyuridine (131IdU). For a given mean absorbed dose to the tissue, the dose response depended on the percentage of cells that were labeled. Specifically, when 1, 10 and 100% of the cells were labeled, a D37 of 6.4, 5.7 and 4.5 Gy, respectively, was observed. The reason for these differences was recently traced to differences in the cellular response to the self- and cross-doses delivered by 131IdU. Systematic isolation of the effects of self-dose resulted in a D37 of 1.2 +/- 0.3 Gy. The cross-dose component yielded a D37 of 6.4 +/- 0.5 Gy. In the present work, the overall survival of multicellular clusters containing 1, 10 and 100% labeled cells is modeled using a semi-empirical approach that uses the mean lethal self- and cross-doses and the fraction of cells labeled. There is excellent agreement between the theoretical model and the experimental data when the surviving fraction is greater than 1%. Therefore, when the distribution of 131I in tissue is nonuniform at the microscopic level, and the cellular response to self- and cross-doses differs, multicellular dosimetry can be used successfully to predict biological response, whereas the mean absorbed dose fails in this regard. PMID- 15658900 TI - Standardization of a fluorometric assay for measuring oxidative stress in irradiated cells. AB - The present study was undertaken to standardize a dichlorofluorescein (DCF) assay for measurement of radiation-induced oxidation of dichlorofluorescin (DCFH) substrate in MCF-10 cells. This assay was highly sensitive and capable of detecting increased DCFH oxidation in the cells exposed to gamma radiation at doses as low as 1.5 cGy with linear dose-response curves. However, the slope of the dose-response curves varied considerably from one experiment to another and was influenced by the fluorescent substrate concentration and cell density. To make the assay reproducible so that results obtained from different experiments could be compared, a series of conversion factors and equations have been established to normalize the data for these variables. The results demonstrate that the DCF assay, as standardized in the present study, is highly reproducible with acceptable assay precision. The normalized results can be compared from one experiment to another even when the experiments were performed using different fluorescent substrate concentrations and/or cell densities. Since changes in DCFH oxidation may be related to changes that are indicative of oxidative stress in cells, this assay can be useful to quantify radiation-induced oxidative stress and evaluate the efficacy of antioxidant agents in protection against radiation induced oxidative stress. PMID- 15658901 TI - Irradiation of mammalian cultured cells with a collimated heavy-ion microbeam. AB - As the first step for the analysis of the biological effect of heavy charged particle radiation, we established a method for the irradiation of individual cells with a heavy-ion microbeam apparatus at JAERI-Takasaki. CHO-K1 cells attached on a thin film of an ion track detector, CR-39, were automatically detected under a fluorescence microscope and irradiated individually with an 40Ar13+ ion (11.5 MeV/nucleon, LET 1260 keV/microm) microbeam. Without killing the irradiated cells, trajectories of irradiated ions were visualized as etch pits by treatment of the CR-39 with an alkaline-ethanol solution at 37 degrees C. The exact positions of ion hits were determined by overlaying images of both cells and etch pits. The cells that were irradiated with argon ions showed a reduced growth in postirradiation observations. Moreover, a single hit of an argon ion to the cell nucleus resulted in strong growth inhibition. These results tell us that our verified irradiation method enables us to start a precise study of the effects of high-LET radiation on cells. PMID- 15658902 TI - Thymoma. PMID- 15658903 TI - Recent changes and the future roles of esophageal cancer surgery. AB - An esophagectomy remains the mainstay treatment for esophageal cancer, and an R0 resection is the most important type of surgery performed with a curative intent. Although a transthoracic esophagectomy is thought to offer better chance for cure in comparison to a transhiatal esophagectomy, the superiority of the former procedure over the latter has not been demonstrated by randomized clinical trials (RCTs). An extended esophagectomy with a three-field lymphadenectomy is a type of esophagectomy with the highest quality of tumor clearance and the capability of prolonging patient survival, but it is contraindicated for patients with 5 or more positive nodes, with simultaneous metastasis to three anatomic compartments, with cervical metastasis from lower esophageal cancer, and with intramural metastasis, because of the absence of survival benefits in these cases. An esophagectomy performed by thoracoscopy and laparoscopy techniques is feasible, however, such an esophagectomy combined with a systematic lymphadenectomy may not qualify as minimally invasive because of the equivalent morbidity rates to an open radical esophagectomy. Whether adjuvant chemotherapy with or without radiotherapy can effectively improve patient survival remains controversial, because only two of 15 RCTs of such adjuvant therapy so far reported have demonstrated a positive survival impact in comparison to surgery alone. The recent increased use of definitive chemoradiotherapy suggests the potential need to perform a salvage esophagectomy because the presence of local persistent or recurrent disease is common after this treatment. The development of a safe and effective salvage esophagectomy should thus be urgently established. PMID- 15658904 TI - Review of preoperative functional evaluation for lung resection using the right ventricular hemodynamic functions. AB - Surgery for patients with lung cancer diminishes their lung functions, due to removal of their lung lobes. Therefore, thoracic surgeons have to consider postoperative lung function of patients. In this review, we explained recent approaches of estimation of postoperative lung function by spirometrical and also pulmonary circulatory measurement values. The most common and simple way to estimate postoperative conditions for patients who undergo lung resections is calculated by numbers of segments that are removed by surgery. However, these methods are not so accurate when the patients have chronic obstructive pulmonary disease. Another method for estimating postoperative conditions using right heart catheterization is a unilateral pulmonary arterial occlusion (UPAO) test. Applying this method, surgery related deaths have been decreased. Since, UPAO test mimics the postoperative state by occluding the pulmonary artery prior to lung surgery, it is supposed to be very accurate. Recently, a novel method to estimate postoperative right heart reserve functions was developed. Using this method, postoperative right heart failures can be anticipated prior to lung resections. In this review, we explain these kinds of methods to prevent impairment of postoperative quality of life. PMID- 15658905 TI - Surgical treatment of lung cancer combined with interstitial pneumonia: the effect of surgical approach on postoperative acute exacerbation. AB - Postoperative acute exacerbation of interstitial pneumonia (IP) is known to be a serious complication in the surgical treatment for primary lung cancer combined with IP. This retrospective study was conducted to investigate the influence of three different approaches to the thorax on postoperative acute exacerbation of IP in lung cancer patients. Forty-seven lung cancer patients who developed it underwent pulmonary resection between 1982 and 2003. Among them, approaches to the thorax consisted of posterolateral thoracotomy (PLT) (n=15), muscle-sparing thoracotomy (MST) (n=15), and video-assisted thoracic surgery (VATS) (n=17). Seven of 47 suffered from acute exacerbation of IP. Single variable analysis suggested that exertion dyspnea (Hugh-Jones classification), serum C-reactive protein, serum lactate dehydrogenase and total lung capacity were considered to be preoperative risk factors of acute exacerbation. As for the VATS patient, there was little frequency of postoperative complication in comparison with the other two approaches. However no significant difference was shown in the incidence of acute exacerbation between the three approaches. CONCLUSION: The use of VATS did not prevent acute exacerbation of IP. However, the incidence of postoperative complications in VATS seemed to be low, therefore further trials are required. PMID- 15658906 TI - Necessity of preoperative screening for brain metastasis in non-small cell lung cancer patients without lymph node metastasis. AB - BACKGROUND: The exclusion of brain metastasis is important to determine the optimal treatment plan in patients with non-small cell lung cancer (NSCLC). However, a routine examination using magnetic resonance imaging (MRI) for the brain remains controversial in preoperative patients with resectable disease. METHODS: To assess the necessity of routine brain MRI for preoperative patients, a retrospective analysis for a consecutive series of 338 patients with NSCLC was performed. Among the 338 patients, 141 patients who were considered to have potentially resectable diseases through an examination of the chest plus an upper abdominal computed tomography scan and bone radioisotope scan with no neurological symptoms received MRI for examination of brain metastasis. RESULTS: The incidence of brain metastasis detected by MRI was 2.1% (three of 141) in all patients, 0% (zero of 80) in patients with N0 disease, 5.2% (one of 19) in N1, and 4.7% (two of 42) in N2 cases. CONCLUSION: In patients with resectable NSCLC, a brain MRI is not considered to be useful due to the low incidence of asymptomatic brain metastasis. PMID- 15658907 TI - Effects of a left ventricular assist device on the myocardium in ischemia reperfusion injury. AB - OBJECTIVE: It is suggested that apoptosis plays a role in heart diseases. The effect of left ventricular assist device (LVAD) on apoptosis at ischemia reperfusion myocardial injury is unclear. We investigated the effect by assisting the ischemia-reperfusion myocardial injury models with LVAD. METHODS: Twelve swines were divided into two groups: the control group and LVAD group. The diagonal branch of the left coronary artery was occluded and released after 35 min. Reperfusion was performed, and observed for 3 hrs. The LVAD group was assisted by LVAD from 5 min before reperfusion to 3 hrs after it. Cardiac function, coronary flow, and cardiac tissue blood flow were measured. Pathologic assay was performed with terminal deoxynucleotidyl transferase-mediated dUTP in situ nick end labeling (TUNEL) and hematoxylin and eosin (HE) staining. mRNA of Bcl-xL and Bak were measured. RESULTS: Ejection fraction, cardiac output, and Emax in the LVAD group were improved (p<0.05 vs. the control group). There were less necrotic cells in the LVAD group than in the control group. There were more TUNEL positive cells in the LVAD group than in the control group. mRNA of Bcl-xL and Bak in LVAD group were high. CONCLUSION: The aggravation of cardiac dysfunction was limited to a minimum in the LVAD group. PMID- 15658908 TI - Non-occlusive condition with the Better-Header roller pump: impacts of flow dynamics and hemolysis. AB - PURPOSE: Impacts on hemolysis and backflow using a non-occlusive setting with the Better-Header (BH) roller pump were investigated. METHODS AND RESULTS: Pump flow of a non-occlusion setting was measured with a pump speed of 3 L/min and 5 L/min against various after-loads. With the non-occlusive setting (BH-NO350), backflow was less than 10% if the pump pressure head was <300 mmHg. When the outlet line is occluded, 80% of the set flow was shunted through the pressure relief valve and outlet pressure did not develop hazardous overpressure. During surgery with the BH-NO350, flow loss was <5% while the pump pressure was maintained at approximately 200 mmHg. An in vitro hemolysis test was conducted at 5 L/min against 350 mmHg, using the standard occlusion (BH-SO), the non-occlusion (BH NO350), and the centrifugal pump (CP). The CP demonstrated less hemolysis than the other two groups; the BH-SO and the BH-NO350 had similar hemolytic characteristics. During cardiopulmonary bypass, no significant differences in hemolysis were seen among the BH-SO, the BH-NO350, and the CP. CONCLUSION: Possible flow loss of the non-occlusion setting with the BH should be almost negligible in most clinical situations. The BH-NO350 demonstrated hemolytic characteristics similar to those of the BH-SO, but not as good as those of the CP. PMID- 15658909 TI - A prospective randomized study of sternal closure: comparison of Mersilene tape versus standard wire closure. AB - BACKGROUND: To compare in a prospective randomized study Mersilene tape and standard metal wire for complications and pain upon sternal wound closure. METHODS: Sixty-four patients scheduled for cardiac surgery were prospectively randomized to undergo sternal closure using either Mersilene tape (n=30) or standard metal wire (n=34). The intensity of postoperative pain from the chest wound was assessed using a visual analogue scale (VAS) on the second and seventh postoperative days and one month after surgery. The examinations with chest X-ray and computed tomography (CT) one month after surgery were performed to evaluate the chest wound and sternum. Follow-up data on sternal and wound healing was assessed for up to one year. RESULTS: No deaths, sternal dehiscence or infection occurred in either group. No wound complications were observed in either group during the year following surgery. A review of data revealed that there was no difference in the intensity of postoperative pain according to the VAS between the two groups. Chest CT demonstrated that no patients in either group had cuts in the sternum. CONCLUSION: This prospective randomized study showed Mersilene tape sternal closure not to be more closely associated with increased complications or patient discomfort due to sternal wound than the standard wire closure. PMID- 15658910 TI - Selective visceral perfusion during thoracoabdominal aortic aneurysm repair. AB - PURPOSE: To evaluate the effectiveness of selective visceral perfusion during repair of an thoracoabdominal aortic aneurysm (TAAA), we compared the postoperative renal and hepatic functions (blood urea nitrogen, serum creatinine, total bilirubin, glutamate pyruvate transaminase) between the two groups with and without perfusion. PATIENTS AND METHODS: We operated on 52 patients with TAAA. Among them, the visceral vessels were reconstructed in 22 patients with selective visceral perfusion and in 12 patients without perfusion. The average selective perfusion time was 49.5+/-25.5 min. in the celiac and superior mesenteric arteries and 32.8+/-18.8 min. in the renal arteries. The average perfusion flow rate per each visceral vessel was 155.4+/-97.4 ml/min. RESULTS: There were five hospital deaths. There was no significant difference between the groups in the postoperative value of four factors. The selective perfusion time for vessel reconstruction in the selective visceral perfusion group was significantly longer than the arterial clamp time for vessel reconstruction in the non-perfusion group (49.5+/-25.5 min. vs. 25.6+/-13.4 min.). CONCLUSION: Our selective visceral perfusion method is not only beneficial for organ protection, but also provides us with the necessary time to reimplant the visceral as well as intercostal or lumbar arteries. PMID- 15658911 TI - Retrospective assessment of vascular injuries: 23 years of experience. AB - PURPOSE: To analyze the operation methods, injury etiologies and localizations, post-operative complications and the reasons for mortality in patients who were admitted for peripheral vascular injuries to our clinics. METHODS: From January 1979 to February 2002, 410 patients were operated for peripheral vascular injuries. Three hundred and one of the patients were male (73.5%) and 109 of them were female (26.5%), and their ages ranged between 1-88 (mean 35.5 years). RESULTS: The most common etiological reason was firearm injuries in 163 patients (39.8%). The most common injured artery was the brachial artery (83 patients, 22.5%) among a total of 369 patients whereas the most commonly injured vein was the common femoral vein (60 patients, 23.4%) in a total of 256 patients. Isolated venous injuries were encountered in 41 patients whereas isolated arterial injuries were detected in 154 patients (37.5%). Hospital admission duration of the patients after trauma was approximately 3 hours. CONCLUSION: The extremity salvage rate in the group was 92.3%. The hospital stay period of the patients was 21.8 days. The mortality rate was 2.6% (11 patients). PMID- 15658912 TI - A tracheostomy case of aortic valve regurgitation successfully treated by a lower mini-sternotomy technique. AB - The performance of cardiac surgical procedures via median sternotomy in a patient with a tracheostomy can result in difficult problems, such as mediastinitis, stoma necrosis or inadequate operative exposure. We present a case of successful treatment for aortic valve regurgitation in a patient with a tracheostomy using a T-shaped sternotomy instead of a usual median sternotomy. This approach permitted adequate surgical exposure for cardiopulmonary bypass and aortic valve replacement. After surgery, the patient had a good clinical course without any complications and he was discharged on the 65th postoperative day. We consider this procedure to be applicable in consideration of the favorable results obtained. PMID- 15658913 TI - A case of early failure of the freestyle stentless bioprosthesis due to cuspal tear. AB - We report an unusual case of early structural failure of the Freestyle stentless bioprosthesis 22 months after initial implantation. A 71-year-old woman, who had undergone stentless valve implantation by the subcoronary method, presented with a new diastolic murmur and progressive congestive heart failure 20 months after the initial implantation. Reoperation was performed using a stented bioprosthesis. The explanted prosthesis showed a tear along the annulus in one cusp. There were no findings of bacterial endocarditis or other macroscopical changes including calcification, thrombus attachment and pannus formation. It is believed that the cause of the cuspal tear was primary tissue failure. Further study is needed to clarify the incidence of this event. PMID- 15658914 TI - Successful repair of a large pseudoaneurysm of the left ventricle late after mitral valve replacement due to rupture of the papillary muscle following acute myocardial infarction. AB - We present a rare case demonstrating a large pseudoaneurysm of the left ventricle late after mitral valve replacement due to rupture of the papillary muscle following acute myocardial infarction. A 52-year-old man, who had undergone mitral valve replacement 7 months previously, presented with severe congestive heart failure. Echocardiography and computed tomography of the chest demonstrated a large pseudoaneurysm of the left ventricle. The patch repair of the orifice of the pseudoaneurysm was successfully performed. PMID- 15658915 TI - Left subclavian artery to descending aorta bypass for coarctation physiology after descending aortic repair. AB - Stenosis of the aorta observed after descending aorta replacement for traumatic aortic injury was managed by a placement of a bypass between the left subclavian artery and the distal descending aorta with success. PMID- 15658920 TI - Do aberrations of single vision lenses really matter? PMID- 15658916 TI - Thoracoabdominal aortic replacement for abdominal aortic aneurysm with atypical coarctation of thoracoabdominal aorta following mitral valve plasty. AB - We successfully treated a case of a 65-year-old female with an abdominal aortic aneurysm coexisting with an atypical coarctation of thoracoabdominal aorta and celiac axis and superior mesenteric artery occlusion. A dilated inferior mesenteric artery was supplying the celiac artery and superior mesenteric artery regions. The patient also had mitral regurgitation. After a mitral valve plasty, we repaired the abdominal aortic aneurysm and the atypical coarctation of the thoracoabdominal aorta using partial extracorporeal circulation, segmental clamping, and a selective perfusion of both the bilateral renal artery and dilated inferior mesenteric artery. The patient had an uneventful hospital course and remains well. PMID- 15658921 TI - Is it any of our business if our patients smoke? PMID- 15658922 TI - Recent advances in measurement of monochromatic aberrations of human eyes. AB - The field of aberrations of the human eye is moving rapidly, being driven by the desire to monitor and optimise vision following refractive surgery. It is important for ophthalmologists and optometrists to have an understanding of the magnitude of various aberrations and how these are likely to be affected by refractive surgery and other corrections. In this paper, I consider methods used to measure aberrations, the magnitude of aberrations in general populations and how these are affected by various factors (for example, age, refractive error, accommodation and refractive surgery) and how aberrations and their correction affect spatial visual performance. PMID- 15658923 TI - Changes in ocular accommodation when shifting between global and local attention. AB - BACKGROUND: The link between cognitive changes in attentional focus and the physiological parameters of the eye is not well understood. The aim of the current work was to examine the role of ocular accommodation, that is, the process of changing the shape of the crystalline lens in order to focus an image onto the retina, in active shifts of visual attention between global and local information. METHOD: Ten adult participants (aged 19 to 27) viewed sequences of complex global/local figures presented at the same location. They were asked to identify either a global or local red target letter and to look for either a global or local letter x (probe) in the sequence following. Target and probe items were separated by a temporal gap of approximately one second. Refraction was measured using a Canon Autorefractor R1 at the time of target appearance and again at the time of probe presentation. Particular interest was paid to trials where participants were required to shift attention either from a local to global level or in the reverse direction. The difference between the two measurements of refraction gave a 'change score' which provided an indication of the change of accommodation. RESULTS: No significant change in refraction was observed when shifting from the local to the global condition. A mean change in refraction of 0.128 dioptres was observed for the global to local condition. The 95 per cent confidence interval for this difference did not overlap zero, indicating a significant change in refraction, which was attributed to an increase in accommodation. DISCUSSION: The results suggest that while a change in accommodation occurs in shifting attention from the global to the local aspect of the complex figures, a similar change is not observed in the reverse direction. These results are consistent with the hypothesis that it is more difficult to restrict attentional focus than it is to widen it. PMID- 15658924 TI - Is fluorescein pattern analysis a valid method of assessing the accuracy of reverse geometry lenses for orthokeratology? AB - BACKGROUND: The aims of this study were to investigate the relative frequencies of correct identifications of variations in the fit of conventional rigid gas permeable (RGP) lenses and reverse geometry lenses (RGL) from fluorescein pattern analysis by orthokeratology (ortho-k) practitioners and non-ortho-k practitioners and to determine whether fluorescein pattern analysis is sensitive for assessing ortho-k lens fittings. METHODS: Slides of fluorescein patterns of different lens fittings were shown to the practitioners, who were asked to identify the ideal, flatter, flattest, steeper and steepest lens fittings. RESULTS: Observed frequencies of correct identifications of most of the conventional RGP lens fittingss were not significantly different from the expected frequencies for both groups of practitioners. The observed frequencies of correct identifications of all of the RGL fittings were either not significantly different or were lower than the expected frequencies. CONCLUSION: The relative frequencies of correct identifications of fluorescein patterns of both conventional RGP lens and RGL fittings by experienced ortho-k practitioners were not different from those by non-experienced ortho-k practitioners. Practitioners from the two groups were not always able to diagnose conventional RGP lens and RGL fittings adequately from fluorescein pattern analysis alone. Fluorescein pattern analysis alone may not be sufficiently sensitive for assessing ortho-k lens fitting. PMID- 15658925 TI - Laughter abolishes binocular rivalry. AB - BACKGROUND: Binocular rivalry is an increasingly popular technique for the study of consciousness, which changes quasi-regularly during rivalry, despite the unchanging sensory stimuli presented to each eye. For example, if a small patch of horizontal stripes is presented constantly to the fovea of one eye and a small patch of vertical stripes is similarly presented constantly to the fovea of the other eye, most subjects experience an alternation between stimuli rather than a simultaneous mixed percept of both. METHODS: Binocular rivalry was induced, superimposed on normal viewing, using liquid crystal shutters and a short persistence monitor, which produced a one degree circular patch of horizontal gratings to the right eye and an identical patch of vertical gratings in the same location for the left eye. The subject signalled with key presses the three possible perceptual states that alternated with each other, namely horizontal, vertical and mixed percept (where horizontal and vertical were simultaneously visible). RESULTS: The present study builds on an incidental observation that laughter stopped the rivalry alternations between horizontal and vertical and induced the mixed percept instead. A physical explanation for this effect was ruled out by using stabilised imagery in the form of retinal after-images of the rivalling gratings. Under conditions of retinal stabilisation, laughter also produced the mixed percept. CONCLUSIONS: The results are discussed in the light of recent work that indicates the inadequacy of low-level explanation of rivalry, with laughter being another complex multi-level contribution to the neural basis of rivalry, along with other aspects of mood. The results are discussed in relation to the interesting literature on the neurology and postulated functions of laughter. PMID- 15658926 TI - Optometry and WebCT: a student survey of the value of web-based learning environments in optometric education. AB - PURPOSE: Improvements in information and communication technology and the need for off-campus delivery have led to the increased use of web-based learning tools in optometry schools around the world. This study compared student-reported preferences for traditional lecture-based learning with their preferences when using a web-based learning tool. METHODS: One hundred and thirteen second and third year students from the School of Optometry and Vision Science at the University of New South Wales were surveyed. All students had worked with WebCT for at least two years. Students were asked to rank a range of learning tools in perceived usefulness and also to state how often they used the particular learning tools. RESULTS: The students rated notes (lecture or WebCT delivered) and clinical laboratory sessions as their most useful learning tools. The use of specific learning tools was more diverse, with students reporting that they often used notes (lecture or WebCT delivered), the WebCT calendar tool and the WebCT discussion tool. This result highlights the valuable contribution of the communication aspect of WebCT to fostering learning communities. The least used learning tools were textbooks, websites mentioned in lectures and library print resources. Interestingly, the purchase of textbooks was high with 77 per cent of students on average reporting they had bought the recommended textbooks. CONCLUSION: Notes were the preferred learning tool of the optometry students at UNSW, suggesting that passive learning of content was the preferred learning style. It is hoped that the introduction of web-based learning environments may allow students and staff to reflect on their preferred teaching and learning styles. Web-based learning tools, such as WebCT, provide a powerful method to facilitate independent deeper learning in students with active learning styles. The current encouragement of student-based active learning methods should see increased use of independent learning platforms, such as WebCT, in optometry schools. PMID- 15658927 TI - Lenticular changes in Alport's syndrome. PMID- 15658928 TI - A bitoric rigid contact lens with base down prism to eliminate binocular vertical diplopia. AB - Prism base down can be incorporated into a rigid gas permeable contact lens to align the diplopic images in a patient who has a significant vertical phoria. A rigid lens may also incorporate both vertical base down prism and a toroidal back surface - the vertical prism to correct the heterophoria and the toric back surface to improve the physical fit of the lens on a highly toric cornea. This paper presents a case where vertical diplopia due to a post-surgical left hyperphoria of about four prism dioptres was eliminated by the incorporation of the appropriate amount of base down vertical prism into a bitoric rigid gas permeable contact lens. PMID- 15658929 TI - David A. Atchison. Keeping the flame of visual optics burning brightly. PMID- 15658930 TI - Fracture mechanics of cortical bone tissue: a hierarchical perspective. AB - The performance of bone tissue in the presence of flaws is a highly remarkable one. Bone tissue is the outcome of an adaptive evolutionary process; thus, insight into the mechanisms by which it fails would provide valuable information not only for development of mechanically superior biomimetic materials but also for development of treatment modalities to prevent debilitating bone fractures. Clinically, fractures of skeletal organs occur as a result of aging, disease, overuse, and trauma. Fracture mechanics, a sub-discipline of solid mechanics that investigates the performance of cracked materials, has been employed extensively in characterizing the mechanisms by which bone tissue fractures. At present the fracture mechanisms at the macroscale are better characterized than at the microscale. On the other hand, a mechanistic understanding of damage evolution at the submicroscopic scale is largely limited to postulations with little experimental insight. The challenge of skeletal fragility will be dealt with more efficiently with deeper understanding of the fracture process at each hierarchical size scale. The most recent review on this subject matter was a decade ago, and there have been numerous developments in the fracture mechanics of bone since then. This review recaps the existing literature with an emphasis on the hierarchical nature of the fracture process in bone, entailing the supramolecular, microscopic, and macroscopic scales. PMID- 15658931 TI - Gas plasma treatment: a new approach to surgery? AB - In this survey we analyse the status quo of gas plasma applications in medical sciences. Plasma is a partly ionized gas, which contains free charge carriers (electrons and ions), active radicals, and excited molecules. So-called nonthermal plasmas are particularly interesting, because they operate at relatively low temperatures and do not inflict thermal damage to nearby objects. In the past two decades nonthermal plasmas have made a revolutionary appearance in solid state processing technology. The recent trends focus on using plasmas in health care, for "processing" of medical equipment and even living tissues. The major goal of tissue treatment with plasmas is nondestructive surgery: controlled, high-precision removal of diseased sections with minimum damage to the organism. Furthermore, plasmas allow fast and efficient bacterial inactivation, which makes them suitable for sterilization of surgical tools and local disinfection of tissues. Much research effort must be undertaken before these techniques will become common in medicine, but it is expected that a novel approach to surgery will emerge from plasma science. PMID- 15658932 TI - The role of urea kinetic modeling in assessing the adequacy of dialysis. AB - The adequacy of dialysis based on urea kinetic modeling has more than 20 years of history. Its methodological approach has demonstrated a strong capacity to improve the outcomes of the dialysis therapy. However, recent results of clinical interventional studies and necessary advances in new models of dialysis schemes, particularly daily and nocturnal hemodialysis, have led to the question if whether that methodology is suitable to address current challenges. This work tries to reach an answer to that question. The major conclusion is that urea kinetic modeling provides an extraordinary starting point to both keeping the clinical support to physicians and building new pharmacokinetic and physiological models that throw light on the understanding of the basic mechanisms that underlie the chronic renal disease, in a synergetic manner. Despite this promising expectation, misuse and improper extensions of urea kinetic modeling may jeopardize its credibility to nephrologists. Our concern about this issue emerges mainly from the analysis of recent attempts to extend this methodology to more frequent renal replacement therapies. PMID- 15658933 TI - Orally fast disintegrating tablets: developments, technologies, taste-masking and clinical studies. AB - Fast disintegrating tablets (FDTs) have received ever-increasing demand during the last decade, and the field has become a rapidly growing area in the pharmaceutical industry. Upon introduction into the mouth, these tablets dissolve or disintegrate in the mouth in the absence of additional water for easy administration of active pharmaceutical ingredients. The popularity and usefulness of the formulation resulted in development of several FDT technologies. This review describes various formulations and technologies developed to achieve fast dissolution/dispersion of tablets in the oral cavity. In particular, this review describes in detail FDT technologies based on lyophilization, molding, sublimation, and compaction, as well as approaches to enhancing the FDT properties, such as spray-drying, moisture treatment, sintering, and use of sugar-based disintegrants. In addition, taste-masking technologies, experimental measurements of disintegration times, and clinical studies are also discussed. PMID- 15658934 TI - Osmotic pumps in drug delivery. AB - In recent years, novel drug delivery systems (NDDS) have been recognized as an attractive niche for the pharmaceutical and health industry. Among various NDDS, osmotic pumps have matured from their use with laboratory animals to the most reliable controlled release systems for humans. Osmotically controlled drug delivery systems use osmotic pressure for controlled delivery of active agent(s). Drug delivery from these systems, to a large extent, is independent of the physiological factors of the gastrointestinal tract. Because of their unique advantages over other types of dosage forms, osmotic pumps form a class of their own among the various drug delivery technologies, and a variety of products based on this technology are available on the market. This article is a review of different types of osmotic pumps and their role in drug delivery. PMID- 15658936 TI - Electrochemical studies on polysorbate-20 (Tween 20)-entrapped haemoglobin and its application in a hydrogen peroxide biosensor. AB - Haemoglobin (Hb) was entrapped in polysorbate 20 and then modified on a pyrolytic graphite electrode. Electrochemical studies revealed that a pair of stable and well-defined redox peaks attributed to the direct redox reaction of Hb could be observed in a phosphate buffer solution (pH 6.0). The anodic and cathodic peaks were located at -236 and -316 mV (versus a saturated calomel reference electrode) separately. The formal potential, E0', was linearly varied with pH in the range from 3.0 to 10.0 with a slope of -48.0 mV.pH-1. Moreover, the protein was capable of catalysing the reduction of H2O2. Accordingly, an unmediated biosensor for H2O2 was prepared with a linear range from 8.0x10(-7) to 1.0x10(-3) M. This biosensor exhibited good stability, sensitivity and reproducibility. PMID- 15658935 TI - Separation of native prion protein (PrP) glycoforms by copper-binding using immobilized metal affinity chromatography (IMAC). AB - The conformational conversion of the normal cellular prion protein (PrPC) into the pathology-associated PrPSc isoform is a key event in TSEs (transmissible spongiform encephalopathies). The host PrPC molecule contains two N-linked glycosylation sites and binds copper under physiological conditions. In contrast with PrPC, PrPSc is insoluble in non-ionic detergents and does not bind to Cu2+ ions. Hence, we utilized copper binding to separate and characterize both PrP isoforms. Infected and uninfected murine brain and bovine stem brain specimens were treated with the mild non-ionic detergent n-octyl-beta-D-glucopyranoside (octylglucoside) to maintain the native PrP conformations during isolation. The solubilized homogenates were loaded on to Cu2+-saturated IMAC (immobilized metal affinity chromatography) columns and eluted using the chelating agent EDTA. Fractions were separated by SDS/PAGE and analysed by immunoblotting using anti PrP monoclonal antibodies for glycosylation profiling. Whereas native PrPC and denatured PrPSc were retained by a Cu2+-loaded resin, native PrPSc and PrPres [PK (proteinase K)-resistant PrP] passed through the column. We demonstrate here that the IMAC technique is appropriate to isolate and partially purify PrPC from healthy brains in its native-like and biologically relevant glycosylated copper binding forms. The IMAC technique is also well suited for the separation of native PrPC from aggregated PrPSc in infected brains. Our results indicate that in contrast with PrPSc in uninfected as well as infected brains, PrPC is predominantly present in the glycosylated forms. PMID- 15658937 TI - A mammalian fatty acid hydroxylase responsible for the formation of alpha hydroxylated galactosylceramide in myelin. AB - Hydroxylation is an abundant modification of the ceramides in brain, skin, intestinal tract and kidney. Hydroxylation occurs at the sphingosine base at C-4 or within the amide-linked fatty acid. In myelin, hydroxylation of ceramide is exclusively found at the alpha-C atom of the fatty acid moiety. alpha Hydroxylated cerebrosides are the most abundant lipids in the myelin sheath. The functional role of this modification, however, is not known. On the basis of sequence similarity to a yeast C26 fatty acid hydroxylase, we have identified a murine cDNA encoding FA2H (fatty acid 2-hydroxylase). Transfection of FA2H cDNA in CHO cells (Chinese-hamster ovary cells) led to the formation of alpha hydroxylated fatty acid containing hexosylceramide. An EGFP (enhanced green fluorescent protein)-FA2H fusion protein co-localized with calnexin, indicating that the enzyme resides in the endoplasmic reticulum. FA2H is expressed in brain, stomach, skin, kidney and testis, i.e. in tissues known to synthesize fatty acid alpha-hydroxylated sphingolipids. The time course of its expression in brain closely follows the expression of myelin-specific genes, reaching a maximum at 2 3 weeks of age. This is in agreement with the reported time course of fatty acid alpha-hydroxylase activity in the developing brain. In situ hybridization of brain sections showed expression of FA2H in the white matter. Our results thus strongly suggest that FA2H is the enzyme responsible for the formation of alpha hydroxylated ceramide in oligodendrocytes of the mammalian brain. Its further characterization will provide insight into the functional role of alpha hydroxylation modification in myelin, skin and other organs. PMID- 15658941 TI - Molecular mechanisms of age-related regulation of genes. PMID- 15658943 TI - Bilateral paroxysmal hemicrania or bilateral paroxysmal cephalalgia, another novel indomethacin-responsive primary headache syndrome? PMID- 15658938 TI - Identification of three critical acidic residues of poly(ADP-ribose) glycohydrolase involved in catalysis: determining the PARG catalytic domain. AB - PARG [poly(ADP-ribose) glycohydrolase] catalyses the hydrolysis of alpha(1''- >2') or alpha(1'''-->2'') O-glycosidic linkages of ADP-ribose polymers to produce free ADP-ribose. We investigated possible mechanistic similarities between PARG and glycosidases, which also cleave O-glycosidic linkages. Glycosidases typically utilize two acidic residues for catalysis, thus we targeted acidic residues within a conserved region of bovine PARG that has been shown to contain an inhibitor-binding site. The targeted glutamate and aspartate residues were changed to asparagine in order to minimize structural alterations. Mutants were purified and assayed for catalytic activity, as well as binding, to an immobilized PARG inhibitor to determine ability to recognize substrate. Our investigation revealed residues essential for PARG catalytic activity. Two adjacent glutamic acid residues are found in the conserved sequence Gln755-Glu Glu757, and a third residue found in the conserved sequence Val737-Asp-Phe-Ala Asn741. Our functional characterization of PARG residues, along with recent identification of an inhibitor-binding residue Tyr796 and a glycine-rich region Gly745-Gly-Gly747 important for PARG function, allowed us to define a PARG 'signature sequence' [vDFA-X3-GGg-X6-8-vQEEIRF-X3-PE-X14-E-X12-YTGYa], which we used to identify putative PARG sequences across a range of organisms. Sequence alignments, along with our mapping of PARG functional residues, suggest the presence of a conserved catalytic domain of approx. 185 residues which spans residues 610-795 in bovine PARG. PMID- 15658944 TI - Vagus nerve stimulation relieves chronic refractory migraine and cluster headaches. AB - Anticonvulsant and antidepressant medications have demonstrated efficacy in migraine treatment. Vagus nerve stimulation (VNS) is an effective treatment for drug-refractory epilepsy and possibly depression and it also has documented analgesic effects. These observations suggested a possible role for VNS in treating severe refractory headaches, and led to a trial of VNS in patients with such headaches. VNS was implanted in four men and two women with disabling chronic cluster and migraine headaches. In one man and one woman with chronic migraines VNS produced dramatic improvement with restoration of ability to work. Two patients with chronic cluster headaches had significant improvement of their headaches. VNS was well tolerated in five patients, while one developed nausea even at the lowest current strength. In conclusion, VNS may be an effective therapy for intractable chronic migraine and cluster headaches and deserves further trials. PMID- 15658945 TI - An assessment of the burden of migraine using the willingness to pay model. AB - Willingness to pay methods measure treatment preferences and also measure the burden of illness in economic terms. We used a contingent valuation method to measure migraine sufferers' willingness to pay (WTP) for acute medication for their most severe headache attacks, based on various profiles of treatment benefits and the characteristics of the migraine sufferer. Subjects were identified from a population-based database of migraine sufferers, previously recruited by random digit dialing. Telephone interviews (n = 1428) were used to gather demographic and headache characteristics. Subjects who met the International Headache Society criteria for migraine with or without aura and satisfied the other inclusion criteria based on telephone interview (n = 312) were invited to participate in a mailed questionnaire study. The questionnaire was mailed to the 310 subjects who agreed to participate and 201 (65%) surveys were returned. The survey included questions on the demographics, the migraine characteristics, and the psychological disposition of the respondents. WTP for an acute migraine treatment with 14 different hypothetical treatment profiles was explored. Responders and non-responders to the survey were generally similar. The newly designed WTP questionnaire had high internal consistency (Cronbach's alpha 0.90) and test-retest reliability (Spearman's correlation coefficients 0.71 0.77). Study subjects were willing to pay a median price of US 5 dollars for a migraine treatment that provided complete relief in 30 min and worked 100% of the time, with no side-effects and no headache recurrence. Median WTP decreased as treatment attributes deviated from this ideal. For example, WTP declined to a median of US 1 dollar for complete relief in 2 h and to US 0.25 dollars for complete relief in 4 h. All of the medication attributes powerfully influenced WTP. Several variables predicted WTP including current payment for medication, MIDAS (Grade III), and those with headaches of long duration. Subjects who employed a greater number of coping skills were less willing to pay. Patient demographics and migraine severity predict WTP, but treatment attributes were also important. As treatment improves, WTP for migraine medications is likely to increase. PMID- 15658946 TI - Predictors of responsiveness to physiotherapy management of cervicogenic headache. AB - A recent randomized controlled trial tested the effectiveness of therapeutic exercise and manipulative therapy on 200 subjects with cervicogenic headache. Although treatments were efficacious, 25% of patients did not achieve a clinically acceptable outcome--50% reduction in headache frequency. This study aimed to identify predictors from variables in subjects' demographics and headache history which might identify those who did or did not achieve a 50-79% or 80-100% reduction in headache immediately after the active treatments and 12 months postintervention. The results revealed no consistent pattern of predictors, although the absence of light-headedness indicated higher odds of achieving either a 50-79% [odds ratio (OR) = 5.45) or 80-100% (OR = 5.7) reduction in headache frequency in the long term. Headaches of at least moderate intensity, the patient's age and chronicity of headache did not mitigate against a successful outcome from physiotherapy intervention. PMID- 15658947 TI - Long-term potentiation of orofacial sensorimotor processing by noxious input from the semispinal neck muscle in mice. AB - Tension-type headache is the most common type of primary headaches but no conclusive concept of pathophysiology exists. This may be due to a lack of an appropriate animal model. This study addressed the hypothesis that noxious neck muscle input induces central sensitization of orofacial sensorimotor processing. The effect of hypertonic saline injection into the semispinal neck muscle on the jaw-opening reflex (JOR) was investigated in anaesthetized mice (n = 11). Hypertonic saline injection into the neck muscle facilitated the JOR for at least one hour: integral (+94.5%) and duration (+18.7%) increased, latency decreased ( 7.5%). The reflex threshold decreased to 61% after injection. Isotonic saline injection into the neck muscle (n = 11) or hypertonic saline injection into a hindpaw muscle (n = 10) did neither change the reflex integral nor the threshold. Long-term potentiation of the JOR by noxious neck muscle input may be an appropriate model to investigate tension-type headache pathophysiology. PMID- 15658948 TI - Variability of reported headache symptoms and diagnosis of migraine at 12 months. AB - Assignment of a diagnosis of migraine has been formalized in diagnostic criteria proposed by the International Headache Society. The objective of the present study is to determine the reproductibility of the formal diagnosis of migraine in a cohort of headache sufferers over a one-year period. The study was performed in a community cohort taking part in a long-term prospective health survey, the GAZEL study. Two thousand five hundred individuals reporting headache in the GAZEL cohort were sent two postal questionnaires concerning headache symptoms and features at 12-monthly intervals. Replies to the questions allowed a migraine diagnosis to be attributed retrospectively using an algorithm based on the IHS classification scheme. The response rate was 82% for the first questionnaire and 69% for both questionnaires. Of the 1733 subjects providing information at both time-points, the agreement rate for the diagnosis of strict migraine (IHS categories 1.1 or 1.2) was 77.7% (kappa = 0.48), with 62.2% of the patients with this diagnosis (IHS categories 1.1 or 1.2) at Month 0 retaining the same diagnosis at Month 12. When diagnostic criteria were widened to include IHS category 1.7 (migrainous disorder), the agreement rate of the diagnosis was similar at 77.6% (kappa = 0.52), but 82% of the patients with this diagnosis (IHS categories 1.1 or 1.2 or 1.7) at Month 0 now retained the same diagnosis at Month 12. In conclusion, the one-year reproducibility of reporting of migraine headache symptoms is only moderate, varies between symptoms, and leads to instability in the formal assignment of a migraine headache diagnosis and to diagnostic drift between headache types. This finding is compatible with the continuum model of headache, where headache attacks can vary along a severity continuum from episodic tension-type headaches to full-blown migraine attacks. PMID- 15658949 TI - Placebo effects in oral triptan trials: the scientific and ethical rationale for continued use of placebo controls. AB - The aim of this study was to determine the characteristics of placebo effects in acute migraine treatment trials of triptans performed over 12 years and assess whether the use of placebo controls in trials of acute migraine treatment remains ethically and scientifically appropriate. We conducted a search for all controlled trials published in English between January 1991 and April 2002 in which adult subjects with migraine were randomly assigned to receive an oral triptan or placebo for the acute treatment of a migraine attack. Thirty-one trials met our criteria for inclusion. Placebo results for each study and pooled placebo results were calculated for the endpoints of headache response, pain-free response and adverse events. Heterogeneity was assessed using the Q statistic, and meta-regression using prespecified covariates was performed to investigate heterogeneity. The study results show a significant degree of heterogeneity. Efforts to explain heterogeneity with available data were not successful, with the exception of adverse event rates to placebo, for which study location (Europe vs. North America) partially explained differences in study results. AE rates were lower in European studies than in North American studies. Across all studies, the mean proportion of subjects who experienced a treatment response at two hours to placebo was 28.48 +/- 8.73% (range 17-50%). The mean proportion of subjects who experienced an adverse event to placebo was 23.40 +/- 14.05% (range 4.86-74%). The mean proportion of subjects who experienced a pain-free response to placebo at two hours was 6.08 +/- 4.43% (range 5-17%). Results of studies allowing use of prophylaxis did not differ significantly from those that did not allow prophylaxis. Placebo effects appear to be enhanced in studies involving children and adolescents. In contrast to an earlier, smaller review, our results do not suggest that randomization ratios influence placebo rates. We conclude that placebo effects in published trials of acute migraine medications are highly variable and often substantial. This variability in placebo response means that active control equivalence trials or the use of historical controls will not provide adequate proof of the safety or efficacy of new drugs, and will not differentiate between drugs that are active vs. placebo but of unknown efficacy relative to each other. The potential for approval of ineffective drugs, inability to compare results of studies performed in different locations, and poor characterization of the tolerability and safety profiles of new drugs represent a greater danger to migraineurs than does the limited-duration use of placebo in carefully monitored clinical trials of consenting subjects. These observations support the view that the inclusion of a placebo group remains of major scientific and ethical importance in trials of migraine medications. PMID- 15658950 TI - Chronic post-traumatic headache--a clinical analysis in relation to the International Headache Classification 2nd Edition. AB - The clinical presentation of chronic post-traumatic headache in 53 patients from a highly specialized headache clinic was analysed and classified according to the diagnostic criteria of the primary headaches in The International Headache Classification 2nd Edition, and compared with the 1st Edition. All patients fulfilled the criteria for both editions indicating that the restrictions in the 2nd Edition have no major influence on the prevalence in specialized clinics. We found the phenomenology to be very heterogeneous, but the vast majority (77%) of patients presented with headache resembling chronic tension-type headache, either as the only manifestation or in combination with migraine symptoms. For the first time episodic tension-type headache is described as occurring after head trauma. The prevalence of coexisting analgesic overuse was 42% and the treatment outcome for these patients was just as favourable as in primary headaches, whereas the time-consuming multidisciplinary treatment demonstrated only very modest results. PMID- 15658951 TI - The CGRP-antagonist, BIBN4096BS does not affect cerebral or systemic haemodynamics in healthy volunteers. AB - BIBN4096BS is a CGRP-antagonist effective in the treatment of migraine. Blocking the receptor of a strong vasodilator involves a theoretical risk of causing cerebral vasoconstriction, a probability not previously investigated with BIBN4096BS. Seven healthy volunteers completed this double-blinded placebo controlled crossover study. The volunteers received randomly 10 min infusions of either placebo, 2.5 mg or 10 mg of BIBN4096BS on 3 separate days. Transcranial Doppler was used to measure the middle cerebral artery blood flow velocity (V(MCA)); global and regional cerebral blood flow (rCBF(MCA)) was measured by 133 Xenon inhalation SPECT. The diameter of the temporal and radial artery was measured by high-resolution ultrasound. Systemic haemodynamics and partial pressure of CO(2) (P(et)CO(2)), and adverse events were monitored regularly. BIBN4096BS had no influence on global or regional cerebral blood flow, or on the blood flow velocity in the middle cerebral artery. There was no effect on systemic haemodynamics and adverse events were minor. We conclude that there is no effect of CGRP-receptor blockade on the cerebral or systemic circulation in humans. Circulating CGRP is therefore not likely to exert a vasodilatory activity in the resting state and the use of BIBN4096BS for acute migraine seems to be without risk of cerebral vasoactivity. These data suggest that BIBN4096BS is the first specific antimigraine drug without vasoactive effect. PMID- 15658952 TI - An unusual indomethacin-sensitive headache: a case of bilateral episodic paroxysmal hemicrania without autonomic symptoms? PMID- 15658953 TI - Ophthalmoplegic migraine: two patients, one at middle age with abducens palsy. PMID- 15658954 TI - Seeing the light: brainstem glioma causing visual auras and migraine. PMID- 15658955 TI - ICHD 2nd Edition: Some considerations on the application of criteria for primary headache. PMID- 15658956 TI - Comment on "ICHD 2nd Edition: Some considerations on the application of criteria for primary headache". PMID- 15658958 TI - What are the priorities for child health in 2004? PMID- 15658960 TI - Mothers, fathers and their children's health. PMID- 15658961 TI - Mind the gap! Helping the children of mothers with postnatal depression. PMID- 15658962 TI - The effects of maternal depression and parental conflict on children's peer play. AB - BACKGROUND: Little is known about the relative effects of exposure to postnatal depression and parental conflict on the social functioning of school-aged children. This is, in part, because of a lack of specificity in the measurement of child and parental behaviour and a reliance on children's reports of their hypothetical responses to conflict in play. METHODS: In the course of a prospective longitudinal study of children of postnatally depressed and well women, 5-year-old children were videotaped at home with a friend in a naturalistic dressing-up play setting. As well as examining possible associations between the occurrence of postnatal depression and the quality of the children's interactions, we investigated the influence of parental conflict and co operation, and the continuity of maternal depression. The quality of the current mother-child relationship was considered as a possible mediating factor. RESULTS: Exposure to postnatal depression was associated with increased likelihood, among boys, of displaying physical aggression in play with their friend. However, parental conflict mediated the effects of postnatal depression on active aggression during play, and was also associated with displays of autonomy and intense conflict. While there were no gender effects in terms of the degree or intensity of aggressive behaviours, girls were more likely to express aggression verbally using denigration and gloating whereas boys were more likely to display physical aggression via interpersonal and object struggles. CONCLUSIONS: The study provided evidence for the specificity of effects, with strong links between parental and child peer conflict. These effects appear to arise from direct exposure to parental conflict, rather than indirectly, through mother-child interactions. PMID- 15658963 TI - Strengthening the parent-child relationship: a review of family interventions and their use in medical settings. AB - The quality of family functioning is important for both psychological well-being and physical health. This review describes family interventions that aim to improve the strength of the parent-child relationship and considers ways in which these approaches can be applied to physical health care. This review first describes the development of family therapy in dealing with children's behavioural and emotional difficulties. As shown in psychotherapeutic settings, parenting skills can affect children's emotional well-being and ability to control their own conduct. Intervention strategies that focus on developing the ability of parents to provide a benign and nurturing parenting style are considered. The review then considers how the principles of family therapy can be applied to settings where physical health is the central issue. In medical settings, families are not only affected by medical interventions but they can also serve to facilitate or hinder clinical effectiveness. Illustrations of how these interventions can be applied in medical settings are provided. Although a practising clinician will need training in using family therapy techniques, it may be possible to recruit a family therapist to help in particular cases. PMID- 15658965 TI - Socially unbiased parenting support on the Internet: a cross-sectional study of users of a large Swedish parenting website. AB - BACKGROUND: The Internet is becoming increasingly recognized as a source of social support. Parents of children with cancer and autism have been shown to find social support online, and many parents of healthy infants and children seek information about parenting online. However, access and use of the Internet is greater among socio-economically advantaged groups, a phenomenon known as the 'digital divide'. Our aim was to investigate whether users of a Swedish general parenting website perceived support in the parenting role and, if so, whether this support was socially biased because of the digital divide phenomenon. METHODS: Users of the largest Swedish parenting website were asked to participate in the study. A total of 2221 users completed the anonymous survey posted on the website during a one-week period. RESULTS: Most respondents (95%) were female (mean age 30.6 years). Respondents' educational level was slightly, but not significantly, higher than that in the general population, whereas 68% had income levels at or under the national average, contradicting the intuitive hypothesis that users would be socio-economically privileged. Perceived social support, measured by the Interpersonal Support Evaluation List (ISEL) appraisal subscale, indicated high perceived support. Living without a partner and having lower levels of income and education increased perceived support. The perception that other parents' opinions are more valuable than the advice of experts was influential in the regression equation for ISEL scores, indicating that peer help is important in online social support. CONCLUSIONS: Internet use for general parenting issues in Sweden, mainly by women, does not seem to follow the digital divide phenomenon. Therefore, the internet provides an exciting opportunity for future infant and child public health work. The lack of fathers, however, was a surprising finding and introduces a gender bias into this seemingly socially unbiased medium. PMID- 15658964 TI - Preventing emotional and behavioural problems: the effectiveness of parenting programmes with children less than 3 years of age. AB - BACKGROUND: Emotional and behavioural problems in children under 3 years of age have a high prevalence, and parenting practices have been shown to be strongly associated with their development. A number of recent systematic reviews have shown that group-based parenting programmes can be effective in improving the emotional and behavioural adjustment of older children (aged 3-10 years). The aim of this review was to establish whether there is evidence from controlled trials that group-based parenting programmes are effective in improving the emotional and behavioural adjustment of children less than 3 years of age, and their role in the primary prevention of emotional and behavioural problems. METHODS: English and non-English language articles published between January 1970 and July 2001 were retrieved using a keyword search of a number of electronic databases. RESULTS: Five studies were included and two meta-analyses were conducted, the first combining data from parent reports and the second combining data from independent observations of children's behaviour. The combined parent reports showed a non-significant difference favouring the intervention group, while the combined independent observations showed a significant difference favouring the intervention group. CONCLUSION: It is concluded that this review points to the potential of parenting programmes to improve the emotional and behavioural adjustment of children less than 3 years of age, but that there is insufficient evidence from controlled trials to assess whether the short-term benefit is maintained over time, or the role that such programmes might play in the primary prevention of emotional and behavioural problems. This review points to the need for further primary preventive research on this important public health issue. PMID- 15658966 TI - Parents' perceptions of the value of the Webster-Stratton Parenting Programme: a qualitative study of a general practice based initiative. AB - BACKGROUND: Parenting styles and parent-child relationships are a determinant of emotional and behavioural problems in children. Controlled trials of parenting programmes have been shown to be effective in helping parents of children with clinical levels of behaviour problems, but there is little research on the impact of such programmes in families where children's development falls in the 'normal' range. Also, such trials do not shed light on why or how programmes do, or do not, work, or how they might be improved. METHODS: A qualitative study of the impact of the Webster-Stratton 'Parents and Children Series' programme on participants in a controlled trial of this programme, whose children's behaviour was below average, but, for the majority, in the normal range. Data were gathered in interviews, open-ended questions on a questionnaire and tape recordings of group leader supervision sessions. RESULTS: Parents reported increased confidence, better relationships with their children, successful use of new behaviour management techniques and improvements in their children's behaviour as a result of the programme. One parent found the programme unsuitable because she was already using the techniques that were taught, and another parent felt the programme was designed for parents of younger children. Many parents reported that additional sessions would have been useful to consolidate what they had learnt, and some parents felt the course would have been more effective if their partners had attended. CONCLUSIONS: The Webster-Stratton Parenting Programme is useful for parents of 'normal' children as well as for parents of children whose behaviour is in the clinical range. Follow-up sessions and attendance by both parents might increase effectiveness. The findings of this study suggest greater benefits to parents and children than were apparent in the controlled trial. PMID- 15658967 TI - The relationship between maternal self-efficacy and parenting practices: implications for parent training. AB - The present study examined the relationship between maternal self-efficacy, dysfunctional discipline practices and child conduct problems. Specifically, three levels of self-efficacy, global, domain and task-specific self-efficacy, were assessed in mothers of 2- to 8-year-old children with conduct problems (clinic group, n=45) and non-clinic mothers from the community (non-clinic group, n=79). Measures of global, domain and task-specific self-efficacy were completed by mothers. Clinic mothers reported significantly lower self-efficacy than non clinic mothers for all but one of the parenting tasks assessed. Both groups of mothers reported lowest self-efficacy for similar parenting tasks. In the sample as a whole self-efficacy measures were significant predictors of maternal discipline style after controlling for other parent, child and risk factors. Of the self-efficacy variables behavioural self-efficacy was the best predictor of mothers discipline style. The findings support the importance of developing parenting strategies that enable parents to generalize their parenting skills to a diverse range of diverse parenting contexts both in the home and in the community. PMID- 15658968 TI - One month after diagnosis: quality of life, coping and previous functioning in siblings of children with cancer. AB - BACKGROUND: The aim of the present study is to describe the quality of life (QoL) of siblings of children with cancer and to predict it according to their health before the diagnosis of cancer in the ill child and their ways of coping with the illness. METHODS: Participants were 83 siblings from 56 families-46 girls and 37 boys, aged 7-18. The assessment took place 1 month to 8 weeks after the diagnosis of cancer in the ill child. The siblings' QoL was assessed with the TNO-AZL children's quality of life questionnaire (TACQOL). Coping strategies were assessed with the Cognitive Coping Strategies Scale for siblings (CCSS-s). Physical problems and eating and sleeping problems that existed before the ill child was diagnosed were determined in a structured interview with the parents. RESULTS: A substantial number of siblings reported impaired cognitive and emotional QoL compared to the reference group. School-aged siblings (7-11 years) reported more trouble with motor functioning than peers. The coping strategy 'predictive control' (maintaining positive expectations regarding the illness) positively predicted siblings' QoL. The presence of health problems before diagnosis was negatively associated with siblings' QoL. Older siblings reported more negative emotions, while girls reported lower social QoL and reliance on 'interpretative control' (trying to understand the illness) was associated with fewer positive emotions. CONCLUSIONS: During the first 2 months after the diagnosis of cancer in a brother or sister, siblings have relatively lower QoL than peers. Health problems that existed before diagnosis may be a predictor of later adjustment problems. Positive expectations about the course of the illness appear to protect siblings from distress. Information about the illness is a delicate issue that requires parental guidance. PMID- 15658969 TI - Development of a questionnaire to collect public health data for school entrants in London: Child Health Assessment at School Entry (CHASE) project. AB - BACKGROUND: To develop a multiprofessional consensus about the relative contributions of the components of children's health and well-being and to develop a questionnaire that can be used to assess these in London's children. METHODS: Semi-structured interviews with health, education and social services professionals were used to identify areas to include in the questionnaire. These ideas were used as the basis for a wider Delphi consultation, with 79 experts in the area of child health. Round 1 of the Delphi asked panelists to rate 54 items as to whether they should be included in the questionnaire or not. Responses were divided into four categories: item to be included measurement method agreed, item to be included measurement method not agreed, no consensus, or excluded. In round 2, consensus was sought for the categories where there was none following round 1. RESULTS: Themes identified by the interviews were: economic factors, ethnicity, environment, nutrition, hygiene and physical activity, growth, suffers from chronic/serious illness, development, disability and learning, accidents and hospital attendances, self-regulation, psychological well-being, significant life events. After Delphi round 1, items included, where quality measurement method was agreed, were: deprivation index (from postcode), child's ethnicity, temporary accommodation, care status, registered with general practitioner, dental visits, height, weight, special educational needs status, baseline educational assessment result, immunization status, visual and hearing function. Following round 2, items relating to chronic illness, mental health, physical functioning, general health, self-esteem, family cohesion and accident status were agreed for inclusion with a measurement method also agreed. The questionnaire was acceptable to parents. CONCLUSION: The validity, reliability and feasibility of this questionnaire must now be examined. This data set, if collected across London, would substantially increase the public health data available and allow trends in health to be monitored. PMID- 15658970 TI - Ascertaining the prevalence of childhood disability. AB - OBJECTIVES: To reapply 1985 Office of Population Census and Surveys (OPCS) disability survey methods, modified as necessary, to a sample of children to ascertain presence of disability. To compare OPCS-based prevalence with prevalence based on carer's views and medical records. DESIGN: Analytical study. Setting Community Child Health Department in UK. PARTICIPANTS: Principal carers of 100 children aged 5-15, selected from a district special needs register. Main outcome measures Comparable information about disability from three sources and diagnosis from carers and medical records. RESULTS: Medical records of 46% contained a diagnosis. Carers were always aware of this, although a single question did not always elicit their knowledge. OPCS-derived threshold disability criteria in categories of Hand function, Personal care, Consciousness and Continence gave prevalence results similar to medical records and carers. OPCS criteria yielded higher prevalence of disability in the areas of Locomotion (8%), Communication (14%) and Hearing (18%). Carers, OPCS and medical records disagreed markedly about prevalence of disabilities of Vision, probably because of the use of differing definitions. OPCS learning criteria were judged unsuitable and standard attainment targets (SATs) were substituted. These provided similar prevalence figures to carers and medical records. OPCS behaviour criteria were also unsuitable and were replaced by the General Health and Behaviour Questionnaire (GHBQ). This found an increased prevalence of problems compared with carers and doctors. CONCLUSIONS: Diagnostic labels have limited use when collecting data about disabled children. Doubt is cast on the validity of some of the 1985 OPCS threshold criteria, and reassessment is suggested before their future use. Further work is needed on the use of SATs and GHBQ in the benchmarking of disability. To collect population data it would be easier and at least equally effective (with caution in the case of Vision) to ask carers directly rather than applying descriptive thresholds and external judgements. Similar information could be obtained from medical records, however, they are likely to be out of date. PMID- 15658971 TI - The impact of childhood chronic neurological diseases on Greek families. AB - BACKGROUND: Although the impact of childhood chronic neurological diseases (CND) on patients' psychological well-being has been increasingly addressed, little attention has been given to the influence of these conditions on family members and family functioning. The purpose of the present study was to investigate the family characteristics of Greek children suffering from CND. METHODS: A total of 52 parents of children with CND were studied by using the Family Environmental Scale (FES), the Family Burden Scale, the General Health Questionnaire (GHQ-28) and a questionnaire on the knowledge of their children's illness, their coping strategies and their satisfaction with our services. During the same period, 30 parents of hospitalized children for common paediatric illnesses completed the FES. In both groups social and demographic features were registered. Appropriate statistical processes were applied to compare the above-mentioned family groups and to study the differences between the families of children with epilepsy (n=37) and the families of children with other CND (n=15). RESULTS: Parents of children with CND discuss their problems less freely, talk less openly around home, score highly on FES subscale of Conflict and, pay more attention to ethical and religious issues and values. Furthermore, the families of children with other CND were more burdened regarding the financial state and the health status of other family members in comparison with families of children with epilepsy. In addition, families of children with epilepsy were more involved in social and recreational activities, appeared to be more knowledgeable on the availability of help in critical conditions and were more satisfied with rendered medical services, in comparison with families of children with other CND. CONCLUSION: These preliminary findings provide important information concerning the special characteristics of Greek families of children suffering from CND, which may prove especially helpful in organizing specific support services. PMID- 15658972 TI - Achieving early surgery for undescended testes. PMID- 15658982 TI - Life is not defined just in base pairs. PMID- 15658983 TI - Freshwater selenium-methylating bacterial thiopurine methyltransferases: diversity and molecular phylogeny. AB - The diversity of bacterial thiopurine methyltransferases (bTPMT) among five natural Se-methylating freshwaters was investigated by polymerase chain reaction (PCR) screenings and sequencings. DNA sequence analyses confirmed the cloned products' identity and revealed a broad diversity of freshwater TPMTs. Neighbour joining (NJ) phylogenetic analyses combining these sequences, all GenBank entries closely related to these sequences and deduced TPMTs obtained in this work from selected gamma-proteobacteria showed TPMTs to form a distinct radiation, closely related to UbiG methyltransferases. Inside the TPMT phylogenetic cluster, eukaryote sequences diverged early from the bacterial ones, and all the bacterial database entries belonged to a subgroup of gamma-proteobacteria, with an apparent lateral transfer of a particular allele to beta-proteobacteria of Bordetella. The NJ phylogenetic tree revealed 22 bTPMT lineages, 10 of which harboured freshwater sequences. All lineages showed deep and long branches indicative of major genetic drifts outside regions encoding highly conserved domains. Selected residues among these highly variable domains could reflect adaptations for particular ecological niches. PCR lineage-specific primers differentiated Se-methylating freshwaters according to their 'tpm lineage' signatures. Most freshwater tpm alleles were found to be distinct from those available in the databases, but a group of tpm was found encoding TPMTs identical to an Aeromonas veronii TPMT characterized in this work. PMID- 15658984 TI - Bacterial community composition determined by culture-independent and -dependent methods during propane-stimulated bioremediation in trichloroethene-contaminated groundwater. AB - An in situ co-metabolic air sparging (CAS) study was carried out at McClellan Air Force Base (MAFB), Sacramento, CA, USA, in a trichloroethene- (TCE) and cis dichloroethene (cis-DCE)-contaminated aquifer where one test zone received 2% propane in air and the other served as a control and received only air. As part of that study, bacterial population shifts were evaluated by length heterogeneity polymerase chain reaction (LH-PCR). The results showed that an organism(s) that had a fragment size of 385 bp was positively correlated with propane removal rates. The 385 bp fragment consisted of up to 83% of the total fragments in the analysis when propane removal rates peaked. A 16S rRNA clone library made from the bacteria sampled from the propane-sparged groundwater included clones of a TM7 division bacterium that had a 385 bp LH-PCR fragment; no other bacterial species with this fragment size were detected. Both propane removal rates and the 385 bp LH-PCR fragment decreased as nitrate levels in the groundwater decreased. Extinction culturing in natural unamended groundwater medium was used to assess the bacterial diversity of the culturable fraction of microorganisms in both CAS and air-sparged groundwater and to bring novel species into culture for further study. The dominant cultures acquired from the CAS groundwater were from the Herbaspirillum/Oxalobacter clade. The dominant cultures from the air-sparged groundwater were from a novel beta-Proteobacterial clade, which we named after isolate HTCC333. PMID- 15658985 TI - Proposed involvement of a soluble methane monooxygenase homologue in the cyclohexane-dependent growth of a new Brachymonas species. AB - High-throughput mRNA differential display (DD) was used to identify genes induced by cyclohexane in Brachymonas petroleovorans CHX, a recently isolated beta proteobacterium that grows on cyclohexane. Two metabolic gene clusters were identified multiple times in independent reverse transcription polymerase chain reactions (RT-PCR) in the course of this DD experiment. These clusters encode genes believed to be required for cyclohexane metabolism. One gene cluster (8 kb) encodes the subunits of a multicomponent hydroxylase related to the soluble butane of Pseudomonas butanovora and methane monooxygenases (sMMO) of methanotrophs. We propose that this butane monooxygenase homologue carries out the oxidation of cyclohexane into cyclohexanol during growth. A second gene cluster (11 kb) contains almost all the genes required for the oxidation of cyclohexanol to adipic acid. Real-time PCR experiments confirmed that genes from both clusters are induced by cyclohexane. The role of the Baeyer-Villiger cyclohexanone monooxygenase of the second cluster was confirmed by heterologous expression in Escherichia coli. PMID- 15658986 TI - Rhizodeposition and the enhanced mineralization of 2,4-dichlorophenoxyacetic acid in soil from the Trifolium pratense rhizosphere. AB - Enhanced biodegradation of organic xenobiotic compounds in the rhizosphere is frequently recorded although the specific mechanisms are poorly understood. We have shown that the mineralization of 2,4-dichlorophenoxyacetic acid (2,4-D) is enhanced in soil collected from the rhizosphere of Trifolium pratense[e.g. maximum mineralization rate=7.9 days-1 and time at maximum rate (t1)=16.7 days for 12-day-old T. pratense soil in comparison with 4.7 days-1 and 25.4 days, respectively, for non-planted controls). The purpose of this study was to gain a better understanding of the plant-microbe interactions involved in rhizosphere enhanced biodegradation by narrowing down the identity of the T. pratense rhizodeposit responsible for stimulating the microbial mineralization of 2,4-D. Specifically, we investigated the distribution of the stimulatory component(s) among rhizodeposit fractions (exudates or root debris) and the influence of soil properties and plant species on its production. Production of the stimulatory rhizodeposit was dependent on soil pH (e.g. t1 for roots grown at pH 6.5 was significantly lower than for those grown at pH 4.4) but independent of soil inorganic N concentration. Most strikingly, the stimulatory rhizodeposit was only produced by T. pratense grown in non-sterile soil and was present in both exudates and root debris. Comparison of the effect of root debris from plant species (three each) from the classes monocotyledon, dicotyledon (non-legume) and dicotyledon (legume) revealed that legumes had by far the greatest positive impact on 2,4-D mineralization kinetics. We discuss the significance of these findings with respect to legume-rhizobia interactions in the rhizosphere. PMID- 15658987 TI - Neorickettsia risticii is vertically transmitted in the trematode Acanthatrium oregonense and horizontally transmitted to bats. AB - Potomac horse fever is known to be transmitted through the ingestion of caddisflies parasitized with Neorickettsia (formerly Ehrlichia) risticii-infected metacercaria. However, the species of trematode involved and how N. risticii is maintained in nature are unknown. In this study, gravid trematodes were recovered from the intestines of 12 out of 15 Eptesicus fuscus big brown bats and eight out of nine Myotis lucifugus little brown bats from various sites in Pennsylvania, USA. Trematode specimens isolated from six E. fuscus bats contained N. risticii DNA. The trematode was identified as Acanthatrium oregonense. N. risticii was detected within individual trematode eggs by polymerase chain reaction as well as by immunofluorescence labelling with an anti-N. risticii antibody, indicating that N. risticii is vertically transmitted (from adult to egg) in A. oregonense. Furthermore, N. risticii DNA was detected in the blood, liver or spleen of 23 out of 53 E. fuscus and M. lucifugus bats, suggesting that N. risticii can also be transmitted horizontally from trematode to bat. These results indicate that A. oregonense is a natural reservoir and probably a vector of N. risticii. PMID- 15658988 TI - Rarity associated with specific ecological niches in the bacterial world: the 'Synergistes' example. AB - The 'Synergistes' group, which apparently represents an as yet unnamed division of the bacteria, was explored in 93 anaerobic environments (guts, soils, digestors, etc.). From 16S rDNA gene-targeted polymerase chain reaction (PCR) assays, this group appeared to be present in 90% of the anaerobic microbial ecosystems analysed. The phylogeny of 103 16S rDNA sequences from 30 ecosystems showed a strong link between 16S rDNA sequences and given ecosystems. 'Synergistes' 16S rDNA sequences from animal sources (termites, guinea pigs, pigs, birds, etc.) formed clustered phylogenetical groups. 'Synergistes' groups were also associated either with anaerobic digestors and soils or with thermophilic conditions. Sequences available from the DNA database were consistent with the results. These results show the wide diversity of the 'Synergistes' division as well as the specific ecological niche of each 16S rDNA sequences. PMID- 15658989 TI - REP code: defining bacterial identity in extragenic space. AB - Through the analysis of 57 bacterial genomes we have detected repetitive extragenic palindromic DNA sequences (REPs) in 11 species. For a sequence to be considered as REP, the following criteria should be met: (i) It should be extragenic, (ii) palindromic, (iii) of a length between 21 and 65 bases and (iv) should constitute more than 0.5% of the total extragenic space. Species-specific REPs have been found in human pathogens such as Escherichia coli, Salmonella enterica, Neisseria meningitidis, Mycobacterium tuberculosis, Rickettsia conorii and Pseudomonas aeruginosa, the plant pathogen Agrobacterium tumefaciens and the soil bacteria Deinococcus radiodurans, Pseudomonas putida and Sinorhizobium meliloti. PMID- 15658990 TI - Disruption of quorum sensing in seawater abolishes attraction of zoospores of the green alga Ulva to bacterial biofilms. AB - Zoospores of the eukaryotic green seaweed Ulva respond to bacterial N acylhomoserine lactone (AHL) quorum sensing signal molecules for the selection of surface sites for permanent attachment. In this study we have investigated the production and destruction of AHLs in biofilms of the AHL-producing marine bacterium, Vibrio anguillarum and their stability in seawater. While wild type V. anguillarum NB10 was a strong attractor of zoospores, inactivation of AHL production in this strain by either expressing the recombinant Bacillus lactonase coding gene aiiA, or by mutating the AHL biosynthetic genes, resulted in the abolition of zoospore attraction. In seawater, with a pH of 8.2, the degradation of AHL molecules was temperature-dependent, indicating that the AHLs produced by marine bacterial biofilms have short half-lives. The Ulva zoospores sensed a range of different AHL molecules and in particular more zoospores settled on surfaces releasing AHLs with longer (>six carbons) N-linked acyl chains. However, this finding is likely to be influenced by the differential diffusion rates of AHLs from the experimental surface matrix. Molecules with longer N-acyl chains, such as N-(3-oxodecanoyl)- L-homoserine lactone, diffused more slowly than those with shorter N-acyl chains such as N-(3-hydroxy-hexanoyl)- L-homoserine lactone. Image analysis using GFP-tagged V. anguillarum biofilms revealed that spores settle directly on bacterial cells and in particular on microcolonies which we show are sites of concentrated AHL production. PMID- 15658991 TI - Soil macropores and compaction control the leaching potential of Escherichia coli O157:H7. AB - The influence of soil structure in controlling leaching of Escherichia coli O157:H7 through soil was investigated under controlled conditions using both intact and repacked soil cores. Leaching rates of E. coli O157:H7 decreased with increasing dry bulk density and were significantly increased by the presence of earthworm (Lumbricus terrestris) burrows in repacked cores. For intact cores, the percentage of E. coli O157:H7 that leached through replicate cores within 72 h varied from 0.01% to 24%. In contrast, the dry bulk densities of intact cores varied only slightly and were not significantly correlated with leaching. Differences in the numbers of E. coli O157:H7 cells in the leachates were not related to variability in the flow volumes, which were relatively constant as a result of the experimental design, but were strongly correlated with the variations in concentrations of E. coli O157:H7 in the leachates. Relatively small variations in the internal structure of soil cores can therefore significantly affect the pathway that cells can take through soil. Factors such as compaction and the occurrence of pores providing preferential flow are prime determinants in the degree of leaching of E. coli O157:H7 through soil. PMID- 15658992 TI - Detection and quantification of the human-specific HF183 Bacteroides 16S rRNA genetic marker with real-time PCR for assessment of human faecal pollution in freshwater. AB - The human-specific HF183 Bacteriodes 16S rRNA genetic marker can be used to detect human faecal pollution in water environments. However, there is currently no method to quantify the prevalence of this marker in environmental samples. We developed a real-time polymerase chain reaction (PCR) assay using SYBR Green I detection to quantify this marker in faecal and environmental samples. To decrease the amplicon length to a suitable size for real-time PCR detection, a new reverse primer was designed and validated on human and animal faecal samples. The use of the newly developed reverse primer in combination with the human specific HF183 primer did not decrease the specificity of the real-time PCR assay but a melting curve analysis must always be included. This new assay was more sensitive than conventional PCR and highly reproducible with a coefficient of variation of less than 1% within an assay and 3% between assays. As the Bacteroides species that carries this human-specific marker has never been isolated, a bacteria real-time assay was used to determine the detection efficiency. The estimated detection efficiency in freshwater ranged from 78% to 91% of the true value with an average detection efficiency of 83+/-4% of the true value. Using a simple filtration method, the limit of quantification was 4.7+/ 0.3x10(5) human-specific Bacteroides markers per litre of freshwater. The aerobic incubation of the human-specific Bacteroides marker in freshwater for up to 24 days at 4 and 12 degrees C, and up to 8 days at 28 degrees C, indicated that the marker persisted up to the end of the incubation period for all incubation temperatures. PMID- 15658994 TI - Seasonal and substrate preferences of fungi colonizing leaves in streams: traditional versus molecular evidence. AB - Aquatic hyphomycetes are the main fungal decomposers of plant litter in streams. We compared the importance of substrate (three leaf species, wood) and season on fungal colonization. Substrates were exposed for 12 4-week periods. After recovery, mass loss, fungal biomass and release of conidia by aquatic hyphomycetes were measured. Fungal communities were characterized by counting and identifying released conidia and by extracting and amplifying fungal DNA (ITS2), which was subdivided into phylotypes by denaturing gradient gel electrophoresis (DGGE) and terminal-restriction fragment length polymorphism (T-RFLP). Mass loss, fungal biomass and reproduction were positively correlated with stream temperature. Conidial diversity was highest between May and September. Numbers of different phylotypes were more stable. Principal coordinate analyses (PCO) and canonical analyses of principal coordinates (CAP) of presence/absence data (DGGE bands, T-RFLP peaks and conidial species) showed a clear seasonal trend (Por=0.88). Season was also a significant factor when proportional similarities of conidial communities or relative intensities of DGGE bands were evaluated (P65-80% of the luminescence response. The reporters exhibited stable luminescence output during normal operations with maximum standard deviations of luminescence over time of c. 5-15% depending on the treatment compartment. Furthermore, deployment of the bioreporters over a 5-month period allowed the determination of an operational range (OR) for each reporter for effluent samples from each compartment. The OR allowed a convenient measure of toxicity effects between treatment compartments and accurately reflected a specific pollution event occurring within compartments of the treatment system. This work demonstrates the utility of genetic modification to provide ecologically relevant bioreporters, extends the sensing capabilities currently obtained through marine derived biosensors and significantly enhances the potential for in situ deployment of reporting agents. PMID- 15658995 TI - The microbial community structure of different permeable sandy sediments characterized by the investigation of bacterial fatty acids and fluorescence in situ hybridization. AB - This study describes the microbial community structure of three sandy sediment stations that differed with respect to median grain size and permeability in the German Bight of the Southern North Sea. The microbial community was investigated using lipid biomarker analyses and fluorescence in situ hybridization. For further characterization we determined the stable carbon isotope composition of the biomarkers. Biomarkers identified belong to different bacterial groups such as members of the Cytophaga-Flavobacterium cluster and sulfate-reducing bacteria (SRB). To support these findings, investigations using different fluorescent in situ hybridization probes were performed, specifically targeting Cytophaga Flavobacterium, gamma-Proteobacteria and different members of the SRB. Depth profiles of bacterial fatty acid relative abundances revealed elevated subsurface peaks for the fine sediment, whereas at the other sandy sediment stations the concentrations were less variable with depth. Although oxygen penetrates deeper into the coarser and more permeable sediments, the SRB biomarkers are similarly abundant, indicating suboxic to anoxic niches in these environments. We detected SRB in all sediment types as well as in the surface and at greater depth, which suggests that SRB play a more important role in oxygenated marine sediments than previously thought. PMID- 15658996 TI - Isolation and properties of a pure bacterial strain capable of fluorobenzene degradation as sole carbon and energy source. AB - A pure bacterial strain capable of aerobic biodegradation of fluorobenzene (FB) as the sole carbon and energy source was isolated by selective enrichment from sediments collected from a polluted site. 16S rRNA and fatty acid analyses support that strain F11 belongs to a novel genus within the alpha-2 subgroup of the Proteobacteria, possibly within a new clade related to the order Rhizobiales. In batch cultures, growth of strain F11 on FB led to stoichiometric release of fluoride ion. Maximum experimental growth rate of 0.04 h-1 was obtained at FB concentration of 0.4 mM. Growth kinetics were described by the Luong model. An inhibitory effect with increasing FB concentrations was observed, with no growth occurring at concentrations higher than 3.9 mM. Strain F11 was shown to be able to use a range of other organic compounds, including other fluorinated compounds such as 2-fluorobenzoate, 4-fluorobenzoate and 4-fluorophenol. To our knowledge, this is the first time biodegradation of FB, as the sole carbon and energy source, by a pure bacterium has been reported. PMID- 15658998 TI - Drug evaluation, drug development and pharmacogenetics: celebrating 30 years of progress. PMID- 15658999 TI - Appropriate dosing of antiarrhythmic drugs in Japan requires therapeutic drug monitoring. AB - OBJECTIVE: In general, drugs are used in accordance with an approved dosage regimen in expectation of an appropriate balance between efficacy and toxicity. However, dose control of drugs with a narrow therapeutic range and marked intersubject variability in pharmacokinetics should be established through individualization of dosing based on therapeutic drug monitoring (TDM). The purpose of this study was to examine differences between the approved dosage regimen and the doses of antiarrhythmic drugs and digoxin used in clinical practice and to examine the influence of TDM on dosing. METHODS: Prescription research of antiarrhythmic drugs was performed at five national hospitals in Japan. Prescriptions for antiarrhythmic drugs (cibenzoline, disopyramide, pirmenol, mexiletine, aprindine, flecainide, pilsicainide, amiodarone and digoxin) were counted for the study period. The mean dose and dose distribution of the drugs were determined in each hospital. Comparisons were made of mean dose obtained in the study with the dosage approved by the authority. In addition, the percentage of patients that received TDM was determined. RESULTS: A difference was seen between the approved dosage and the actual dose. For all drugs except flecainide, the mean dose was smaller than the approved dosage. For all drugs except digoxin, remarkable variations were seen in the dose distribution among the hospitals. Digoxin showed a similar dose distribution among the five hospitals. Overall, the percentage of patients that received TDM was low except for Hospital A. However, TDM of digoxin was relatively common at four of the hospitals. CONCLUSIONS: It is concluded that, with the exception of digoxin, the appropriate dosing regimen for antiarrhythmic drugs is not yet established. The establishment of appropriate dosing regimens for antiarrhythmic drugs requires the more widespread adoption of TDM. PMID- 15659000 TI - Pharmacoepidemiologic study of potential drug interactions in outpatients of a university hospital in Thailand. AB - BACKGROUND: Drug-drug interaction is a potential cause of adverse drug reactions. The incidence of such drug interactions in university hospitals in Thailand is unknown. PURPOSE: To estimate the rate of potential drug-drug interactions in outpatients of a typical Thai university hospital, and to identify risk factors for such interactions in Thai patients. METHODS: One-year outpatients' prescription data were retrieved from the hospital computer records. Potential drug interactions were identified using the existing drug-interaction database system. Potential interactions within a specific prescription and involving drugs prescribed 1-, 3- and 7-day earlier were searched for. Possible associations between occurrence of an interaction and a patient's age and gender and the number of items on the prescription were explored. RESULTS: The overall rate of potential drug interactions was 27.9% with a maximal value of 57.8% at the Department of Psychiatry. The rate of the most potentially significant interactions was 2.6%, being the highest in the Department of Medicine (6.0%), with isoniazid vs. rifampin as the most common interacting combination. The rate increased with the patient's age and prescription size (P=0.000). The odd's ratio of having at least one potential drug interaction was 1.8 (64.2%) when age increased by 20 years (P=0.000) and 2.8 (165.7%) when another drug was added (P=0.000). The rate of potential drug interactions was the same for both genders. The rate of potential drug interactions detected across prescriptions was higher than within prescriptions and was dependent on the time interval between prescriptions. CONCLUSIONS: Potential drug interactions were common in our sample of patients. The rate of such interactions increased with the number of drugs prescribed and the patient's age. PMID- 15659001 TI - A literature search on pharmacokinetic drug interactions of statins and analysis of how such interactions are reflected in package inserts in Japan. AB - BACKGROUND AND OBJECTIVES: Statins (HMG-CoA reductase inhibitors) are one of the most widely prescribed classes of drugs throughout the world, because of their excellent cholesterol-lowering effect and overall safety profile except for rare but fatal rhabdomyolysis arising either directly or indirectly by pharmacokinetic interactions with certain other drugs. As package inserts in pharmaceuticals are the primary source of information for health care providers, we carried out a literature search to examine how crucial information was provided in package inserts of five statins approved in Japan (simvastatin, atorvastatin, fluvastatin, pravastatin and pitavastatin). METHODS: A MEDLINE search from 1996 to June 2004 was carried out to identify studies on clinical pharmacokinetic drug interactions for the five statins. We mainly collected information on area under plasma concentration (AUC) following co-administration of statins with other drugs. The current package inserts used in Japan were obtained from the website of the Pharmaceutical and Medical Device Agency whereas USA package inserts were obtained from the Food and Drug Administration website. RESULTS: The majority of package inserts listed the drugs that interacted with statins with most describing the risk of rhabdomyolysis because of the possibility of increases in blood concentration. However, quantitative information such as change in AUC was provided in only a few cases. Instructions for dosage adjustment are seldom provided in the Japanese package inserts. USA package inserts list almost identical drug interactions as the Japanese package inserts, although they contain more quantitative data, especially for typical cytochrome P450 (CYP) inhibitors. CONCLUSION: All pharmacokinetic drug interactions including relevant quantitative data for potential effectors and details on mechanisms of interaction need to be given in package inserts as soon as the information becomes available, to ensure safe and proper use of the drugs concerned. Including such information in the package insert will be an extremely valuable aid for health care providers. PMID- 15659002 TI - Prevalence of over-the-counter drug-related overdoses at Accident and Emergency departments in Northern Ireland--a retrospective evaluation. AB - BACKGROUND AND OBJECTIVES: One major concern associated with misuse/abuse of over the-counter (OTC) products is the potential for over-dosage. The aim of this research study was to evaluate, over a 3-month period, OTC medicine-related overdoses (those involving OTC drugs only and OTC drugs in combination with other drugs) that led to patients presenting at the Accident and Emergency (A & E) departments in four Belfast hospitals. METHODS: A data collection sheet was designed to capture the information required from the A & E records in each hospital. A retrospective week-by-week data collection, reviewing A & E records, took place over a 3-month period (starting on 1 December 2002). All data related to cases presenting at the A & E departments because of drug overdoses (either accidental or deliberate according to Read Clinical Classification) were included in the study. Data were coded and entered into a custom designed SPSS database for analysis, using Chi square and Fisher exact tests. RESULTS: OTC drug-related overdoses comprised 40.1% of all overdoses, of which 24.0% were OTC-only overdoses. Those who overdosed on OTC drugs (solely or combined with other drugs) were mainly female (62.3%) and in the age category 31-50 years (44.9%; P <0.05). The majority (n=215) of OTC-related overdoses were intentional, whereas only 28 were accidental. Of those who attended the A & E departments and had an overdose history, one-third overdosed on OTC-related products and two-thirds overdosed on OTC drugs only. CONCLUSIONS: OTC drugs accounted for a significant proportion of overdose presentations at the A & E departments in Northern Ireland. Higher awareness of the potential of OTC product use in overdose cases (intentional or accidental) is recommended for both the public and health care professionals. PMID- 15659003 TI - Counselling quality in community pharmacies: implementation of the pseudo customer methodology in Germany. AB - OBJECTIVES: To investigate a new method for evaluating counselling performance of staff in community pharmacies and to assess the quality of patient counselling. METHOD: Trained pseudo customers, instructed to play their role according to two different self-medication scenarios, visited voluntarily participating community pharmacies in Berlin. After documenting the counselling process, immediately after each visit, outside the pharmacy on an assessment form, the pseudo customer re-entered the pharmacy and gave detailed performance feedback to the counsellor and the pharmacist in charge in order to provide support for improving counselling skills and practice behaviour, when appropriate. This was followed with a written summary of the general performance of all participating pharmacies and additional individual feedback and suggestions for improvement. Educational needs were identified for subsequent performance-based educational strategies such as group-workshops, team-training and on-site team-coaching. RESULTS: Forty nine community pharmacies in Berlin volunteered to participate in this pilot study. Ninety-eight per cent of the participating pharmacies offered advice. However, in 36% of the cases, advice was only given on request. The different types of scenarios--presentation of a symptom or request for a specific product- made a great difference to the spontaneity of questions and advice. At least one question to check on accuracy of self-diagnosis was asked in 95% of the cases of symptom presentation but in only 47% of the cases of specific product request. Information on appropriate self-medication was provided on at least one item in 74% of pseudo customer visits, but most of the time the information was not sufficient. Communication skills (nonverbal elements, comprehensibility etc.) were very good or good in 54% of the visits. Potential for improvement was mainly in relation to the use of open-ended questions to gain more information and on counselling about appropriate self-medication. Direct feedback was given in 96% of the pharmacies (one person refused to accept feedback and one feedback had to be postponed because of time shortage). All of the participants regarded counselling as an important subject in pharmacy practice. CONCLUSION: The pseudo customer method was successfully used in this study of German community pharmacies. It was shown that pseudo customer visits and performance feedback following the counselling process, were feasible in daily practice and well accepted by the participants. A training program, focussing on areas in most need of improvement, has been developed. The promising results have led to the Federal Chamber of Pharmacists in Germany adopting this method as part of a continuous quality improvement program in community pharmacies. PMID- 15659004 TI - Methylphenidate: prescribing patterns in a South African primary care patient population. AB - BACKGROUND AND OBJECTIVE: Sales figures have indicated that there has been a steady increase in the use of methylphenidate in South Africa. The primary aim of the study was to investigate the prescribing of methylphenidate (indicated for Attention Deficit Hyperactivity Disorder in children, and narcolepsy) in a South African primary care patient population. METHODS: A retrospective, exposure cohort drug utilization study was conducted in the South African private health care sector. Data were obtained from a medical aid administrator and were analysed for all patients who received one or more prescriptions for methylphenidate (N06B A04) during 2002. RESULTS AND DISCUSSION: A total of 106 patients received one or more prescriptions for methylphenidate during the year. Ninety of the 106 patients (84.9%) were under 25 years of age (average age 10.51 years, SD=3.92 years). These 90 patients were all single and the majority of them (75.6%) were males. The Lorenz curve was used to illustrate skewness in the prescribing of methylphenidate to the patient population. Patients under the age of 25 years received on average 3.46 prescriptions for methylphenidate during the year. The most common chronic diagnosis category was 'Hyperkinetic Syndrome of Childhood'. Most of the prescriptions were obtained from pharmacies, followed by dispensing doctors and rehabilitation services. CONCLUSION: The results were in agreement with a previous South African study conducted in 1996. Further studies are recommended, especially into the quality of life of patients before and after receiving methylphenidate. PMID- 15659005 TI - Adjunctive use of atypical antipsychotics and anticholinergic drugs among patients with schizophrenia. AB - BACKGROUND: Treatment of schizophrenia with antipsychotics is often associated with extrapyramidal symptoms (EPS), a disorder involving involuntary muscle movement. Because EPS are often associated with the use of antipsychotics, anticholinergic agents are often indicated. OBJECTIVE: In this observational, retrospective study, we examined whether the initiation of olanzapine or risperidone, the two most widely prescribed atypical antipsychotics, is related to the adjunctive use of anticholinergic agents. METHOD: We identified patients with schizophrenia from outpatient clinics in the Veterans Health Administration (VA) and defined initiation of olanzapine or risperidone as patients who were not on any antipsychotics for 6 months and subsequently initiated on the target drug between 1/4/1999 and 31/3/2000. The data were analysed using tests of means or chi-square tests. RESULTS: The study yielded two major findings. First, compared with risperidone initiators, there were significantly fewer olanzapine initiators who used at least one anticholinergic agent adjunctively. Secondly, among olanzapine or risperidone initiators, patients who used at least one anticholinergic agent adjunctively tended to stay on the target drug significantly longer than those who did not use any anticholinergic agent adjunctively with the target drug. CONCLUSION: As the use of anticholinergics is a proxy for the presence of EPS, these findings suggest that risperidone may be more associated with EPS than olanzapine. However, to assess the benefits and side effects associated with olanzapine or risperidone, future research needs to examine various patient outcomes resulting from the initiation of each drug. PMID- 15659006 TI - The efficacy of three desensitizing agents in treatment of dentine hypersensitivity. AB - PURPOSE: The purpose of this study was to evaluate the efficacy of three desensitizing agents vs. placebo. MATERIALS AND METHODS: One hundred and six hypersensitive teeth of 26 patients were included in this study, and the baseline hypersensitivity level of all teeth was established as 'moderate' by using Visual Analogue Scale (VAS). The teeth were divided into four groups: to the first group 5% potassium nitrate bio-adhesive gel, to the second 2% sodium fluoride bio adhesive gel and to the third one step adhesive system Prompt L-Pop were applied as desensitizing agents. Group 4 was the control group in which a desensitizer free bio-adhesive gel was used as placebo. Post treatment and eighth week control measurements were recorded on VAS. RESULTS: It was observed that the efficacy of three desensitizing agents did not differ from each other (P >0.05) and except for placebo all reduced moderate dentin hypersensitivity effectively (P <0.05). CLINICAL RELEVANCE: Five per cent potassium nitrate, 2% sodium fluoride bio adhesive gels and one-step bonding agent Prompt L-Pop were effective in reducing moderate dentine hypersensitivity. PMID- 15659007 TI - Factors influencing clinicians' decisions to prescribe medication to prevent coronary heart disease. AB - BACKGROUND AND OBJECTIVE: There are variations between individual clinicians as to the thresholds at which preventive treatment for coronary heart disease (CHD) should commence. Patients' decisions may be influenced by clinicians' recommendations. Free text comments added by respondents to closed questionnaires may identify areas which are of real concern to them about the topic being studied. The study aimed to identify issues voluntarily raised by clinicians surrounding the decision to prescribe preventive treatment for CHD. METHODS: An analysis was undertaken of the free text comments made by cardiologists, general practitioners and practice nurses who responded to a closed question postal questionnaire in which they were asked to identify at which level of pretreatment risk they would offer treatment. RESULTS AND DISCUSSION: A similar percentage of respondents in each professional group provided free text comments. Clinicians' concerns centred on five main themes around prescribing decisions: the risks and benefits of treatment, the patient's role in treatment decisions, patient characteristics, costs to patients, and costs to the health services. Different issues may be of more concern to some professional groups than others. CONCLUSION: In addition to the use of risk assessment tools and guidelines, clinicians' actual prescribing behaviour may be influenced by more subjective factors. Patients at similar risk may receive different advice depending on the individual clinician they consult. PMID- 15659008 TI - Assessing the bioequivalence of 4- and 8-mg benidipine tablets in healthy volunteers after a single oral dose. AB - OBJECTIVE: To assess the bioequivalence of a new tablet formulation of benidipine hydrochloride with reference to a marketed product. METHODS: Two groups, consisting of 24 healthy volunteers each, received a 4- or 8-mg (one or two tablets) reference benidipine hydrochloride tablet and one or two test tablets in a 2 x 2 cross-over study. There was a 6-day washout period between doses. The plasma benidipine concentration was monitored using LC/MS/MS for 8 h after the dose. The area under the plasma concentration-time curve from time 0 to the last sampling time (AUCt) was calculated using the linear-log trapezoidal rule. The maximum plasma drug concentration (Cmax) and the time to reach Cmax (Tmax) were compiled from the plasma concentration-time data. Analysis of variance was carried out using logarithmically transformed AUCt and Cmax, and untransformed Tmax. RESULTS: The geometric mean AUCt was 2.23 ng/mL/h (test medication) and 2.47 ng/mL/h (reference medication) for the 4-mg tablet, and 9.57 and 9.97 ng/mL/h for the 8-mg tablet, respectively. A Cmax of 1.94 and 2.01 ng/mL was achieved for the test and reference medication for the 4-mg tablet, and 5.94 and 6.53 ng/mL for the 8-mg tablet, respectively. The 90% confidence intervals for AUCt and Cmax were 0.8441-1.0481 and 0.8739-1.2037 for the 4-mg tablet, and 0.8559-1.1273 and 0.9926-1.2176 for the 8-mg tablet, respectively, satisfying the bioequivalence criteria of the US Food and Drug Administration Guidelines, and the Korea Food and Drug Administration Guidelines. These results indicate that the 4- and 8-mg tablets of benidipine are bioequivalent to the reference formulations. PMID- 15659009 TI - Myocarditis, pericarditis and cardiomyopathy in patients treated with clozapine. AB - Clozapine is known to cause cardiac side-effects, including myocarditis, pericarditis and cardiomyopathy. Prompted by a case of clozapine-related pericarditis in our hospital we undertook a review of the literature for reports of myocarditis, pericarditis and cardiomyopathy occurring in patients treated with clozapine. This is the first comprehensive review of the literature on this topic. PMID- 15659011 TI - Delivery of growth factors using gene therapy to enhance bone healing. AB - OBJECTIVE: To review the delivery of growth factors using gene therapy for enhancing long-bone fracture healing. STUDY DESIGN: Literature review. METHODS: MEDLINE and CAB Abstracts literature search (1980-2004). RESULTS: Non-union and infected non-union are relatively common complications of long-bone fractures in human and veterinary patients. Growth factors are cytokines that regulate many cell functions important in fracture healing. Exogenous growth factors can be delivered to the fracture site as recombinant proteins or using gene therapy. Recombinant human bone morphogenetic protein-2 and -7 (rhBMP-2 and -7), in particular, enhance fracture healing in numerous experimental and clinical studies. Some limitations with use of recombinant proteins may be overcome by use of gene therapy. Gene therapy involves delivery of the growth factor gene to cells at the fracture site using a viral or non-viral vector. The gene is then expressed (protein synthesis) by cells at the fracture site. Delivery of the BMP gene to the fracture site using gene therapy has been evaluated in laboratory animal models of non-union, with favorable results and without complications. CONCLUSION: Delivery of growth factors, particularly members of BMP family, to the fracture site using gene therapy may be a method to enhance fracture healing. Use of this technology may improve the prognosis for patients with long-bone fractures. CLINICAL RELEVANCE: Clinical application of gene therapy could improve the prognosis for human and veterinary patients with long-bone fractures, but has not been evaluated clinically. PMID- 15659012 TI - Cutaneous wound healing in the cat: a macroscopic description and comparison with cutaneous wound healing in the dog. AB - OBJECTIVE: To describe the macroscopic features of first and second intention cutaneous wound healing in the cat and compare with the dog. STUDY DESIGN: Experimental study. ANIMALS: Domestic shorthaired cats (6) and beagle dogs (6). METHODS: Square, open cutaneous wounds created on the dorsal aspect of the thorax were evaluated for 21 days for temporal and spatial development of granulation tissue, wound contraction, epithelialization, and total healing. To evaluate first intention healing, breaking strength of sutured linear cutaneous wounds was measured at 7 days post-wounding. Laser-Doppler perfusion imaging was used to measure cutaneous perfusion. RESULTS: First intention healing: sutured wounds in cats were only half as strong as those in dogs at day 7 (0.406 versus 0.818 kg breaking strength). Second intention healing: cats produced significantly less granulation tissue than dogs, with a peripheral, rather than central distribution. Wound epithelialization and total wound healing (total reduction in open wound area from contraction and epithelialization) were greater for dogs than for cats over 21 days. Wound contraction on day 7 was greater for dogs, but not on day 14 or 21. Cutaneous perfusion was initially greater for dogs than for cats, but no differences were detected after day 7. CONCLUSIONS: Significant, previously unreported differences in cutaneous wound healing exist between cats and dogs. In general, cutaneous wounds in cats are slower to heal. Cats and dogs also appear to use different mechanisms of second intention healing. In cats wounds close mainly by contraction of the wound edges, whereas in dogs wounds close more from central pull, and epithelialization. CLINICAL RELEVANCE: Surgeons should view the cat as a unique species, which presents its own special challenges in wound healing, and should take this into account when planning treatment of feline wounds, either by primary closure, or by second intention healing. PMID- 15659013 TI - Autologous osteochondral grafting (mosaic arthroplasty) for treatment of subchondral cystic lesions in the equine stifle and fetlock joints. AB - OBJECTIVE: To describe treatment of equine subchondral bone cysts (SBCs) by reconstruction of the articular surface with osteochondral grafts. STUDY DESIGN: Case series of horses with SBCs unresponsive to conservative therapy. ANIMALS: Eleven horses (1-12 years). METHODS: SBCs were identified in 4 locations: medial femoral condyle (5 horses), lateral femoral condyle (1), distal epiphysis of the metacarpus (4), or metatarsus (1). Osteochondral autograft transplantation (mosaic arthroplasty) was performed, taking grafts from the abaxial border of the medial femoral trochlea of the unaffected limb. Graft implantation was achieved through a small arthrotomy or by arthroscopy depending on SBC location. RESULTS: All horses improved postoperatively; 10 horses had successful outcomes with radiographic evidence of successful graft incorporation and 7 returned to a previous or higher activity level. On follow-up arthroscopy (5 horses) there was successful reconstitution of a functional gliding surface. One horse had delayed incorporation of a graft because of a technical error but became sound. One horse had recurrence after 4 years of work and soundness. One stallion was used for breeding and light riding because of medial meniscal injuries on the same limb. CONCLUSIONS: Implantation of osteochondral grafts should be considered for SBC when conservative management has not improved lameness and there is a risk of further joint injury and degeneration. CLINICAL RELEVANCE: Mosaic arthroplasty should be considered for treatment of subchondral bone cysts of the femoral condyle and distal articular surface of the metacarpus/tarsus in horses that are refractory to non-surgical management. PMID- 15659014 TI - Evaluation of the characteristics of venous occlusion after placement of an ameroid constrictor in dogs. AB - OBJECTIVES: To evaluate the mechanism of vascular occlusion after placement of an ameroid constrictor (AC) on a large intra-abdominal vein and document changes in blood flow. STUDY DESIGN: Experimental study. ANIMALS: Six adult dogs. METHODS: Six 6.5 mm ACs were digitally scanned to measure area and circumference of the ameroid and inner lumen before surgery. ACs were surgically positioned around the left common iliac vein (CIV) in each dog. Peri-vascular ultrasonic flow probes were positioned on the left CIV cranial to the AC, and on the right CIV as an internal control. Blood flow measurements were recorded daily until there was no, or prolonged minimal, blood flow, at which time the dogs were euthanatized. Left and right CIVs were removed for histologic evaluation, and the ACs re-scanned to evaluate degree and direction of expansion of the ameroid. RESULTS: Reduction in blood flow occurred within 10 days in all dogs. Three dogs had complete occlusion by day 10 from thrombus formation. One dog damaged the flow probes at 13 days when blood flow was approximately 50% of intra-operative values; the dog was euthanatized at 37 days; there was partial occlusion from thrombosis. Two dogs had persistent low blood flow and occlusion from thrombus formation when euthanatized at 59 and 98 days. There was a significant increase in total AC area and outer circumference, and decrease in luminal area and inner circumference when presurgical and postocclusion measurements were compared. CONCLUSIONS: Vascular occlusion after placement of an AC on a large intra-abdominal vein occurred from thrombus formation. CLINICAL RELEVANCE: A similar pattern of venous occlusion may occur after placement of an AC on a portosystemic shunt, which may lead to both short- and long-term complications associated with portal hypertension. PMID- 15659015 TI - Preservation of cadavers for surgical technique training. AB - OBJECTIVE: To evaluate a technique for preservation of organoleptic tissue characteristics (color, odor, texture, and flexibility) in cadavers used for surgical instruction. STUDY DESIGN: Experimental study. ANIMALS: Forty-three canine cadavers. METHODS: Cadavers were preserved with a modified Larssen solution of the Hospital Cochim, Paris and cryopreservation. Tissue handling qualities were evaluated in surgical laboratory sessions. RESULTS: All cadavers kept texture and tissues consistency, especially skin and muscle, similar to those of live animals. Some skin desquamation and pallor of the mucous membranes occurred with repetitive freeze-thaw cycles. CONCLUSIONS: This preservation technique provides acceptable cadaver quality and tissue handling for use in surgical instruction. CLINICAL RELEVANCE: Preparation of patient cadavers by intravascular injection of modified Larssen solution yielded suitable instructional models for surgical training. PMID- 15659016 TI - Increased sacral screw purchase minimizes screw loosening in canine triple pelvic osteotomy. AB - OBJECTIVE: To determine if screw loosening in triple pelvic osteotomies (TPO) is minimized when screws cranial to the ilial osteotomy had maximal sacral purchase. STUDY DESIGN: Prospective study. ANIMALS: Forty-six dogs with decreased acetabular coverage of the femoral head and minimal degenerative joint changes. METHODS: TPOs were performed where screws cranial to the ilial osteotomy were inserted to maximally engage sacral bone. Data collected were: use of ilial and ischial cerclage wire, screw length, ventrodorsal radiographic sacral width (most caudal aspect), pelvic canal diameter, and sacral penetration of the 3 cranial screws. On all subsequent radiographs, changes in screw position, pelvic canal diameter, and sacral purchase were noted. RESULTS: For 69 TPOs, 414 screws were used. Mean radiographic cranial screw length was 34.54 mm. Combined sacral depth of all 3 screws was 93.3% of sacral width. All osteotomies healed uneventfully. Twenty-four screws (6%) loosened with 12 being in the most cranial positions. Use of ischial or ilial cerclage wires did not statistically influence screw loosening. Pelvic diameter decreased by a mean of 7.79% from postoperative radiographs to the last radiographic recheck. CONCLUSIONS: By sufficiently engaging the sacrum with screws cranial to the ilial osteotomy, implant failures can be avoided and screw loosening minimized when a 6-hole TPO plate is used. CLINICAL RELEVANCE: To minimize screw-loosening in TPO, screws inserted cranial to the ilial osteotomy should be inserted to maximum sacral depth without penetrating the vertebral canal. PMID- 15659017 TI - Mechanical performance of a screw-type veterinary suture anchor subjected to single load to failure and cyclic loads. AB - OBJECTIVE: To characterize the mechanical performance of a veterinary bone anchor under static and cyclic loads. STUDY DESIGN: Mechanical testing study. ANIMALS: Cadaveric canine humeri. METHODS: Humeri (6 pairs) were collected from skeletally mature dogs (mean [+/-SD] age, 17.2+/-2.1 months; weight, 20.8+/-1.5 kg). Bone anchors were inserted in the proximal metaphysis using nylon, and were longitudinally extracted. For the opposite humerus, anchors were subjected to longitudinal cyclic load (50% of the load at failure of their pair) for 1200 cycles then longitudinally loaded to failure. Anchors were then installed in a similar and adjacent area of these 2(nd) humeri with nylon and cyclically tested perpendicular to the axis of anchor insertion (100% of the longitudinal holding power of their pair) for 1200 cycles, then perpendicularly loaded to failure. Paired t-tests were used to compare holding power before and after longitudinal cyclic testing. RESULTS: Longitudinal holding power of the screw-type anchor in the proximal humerus was 385+/-30 N. Anchor pullout was the only mode of failure. Anchors in the paired humeri did not fail after 1200 cycles of 50% longitudinal loading, and post-cycle holding strength was not different (335+/-87 N; P=.32). Perpendicularly loaded anchors did not fail after 1200 cycles of 100% of opposite longitudinal holding strength, and had post-cycle perpendicular holding strengths of 514+/-72 N. Suture breakage was the mode of failure. CLINICAL RELEVANCE: Bone anchor holding strength is dependent on orientation of suture load. Screw-type bone anchor holding strength was not affected by longitudinal cyclic loading, and holding strengths of approximately 385 N can be expected in metaphyseal bone of large-breed mature dogs. Perpendicularly loaded anchors have higher failure loads, and holding strength of approximately 514 N can be expected in metaphyseal bone of the proximal humerus. PMID- 15659018 TI - Thoracolumbar lateral corpectomy for treatment of chronic disk herniation: technique description and use in 15 dogs. AB - OBJECTIVE: To describe a technique for thoracolumbar lateral corpectomy and to evaluate its use for treatment of chronic thoracolumbar disk disease in dogs. STUDY DESIGN: Retrospective study. ANIMALS: Fifteen dogs with signs of chronic thoracolumbar disk herniation. METHODS: After a dorsal or lateral approach to the spine, a lateral slot was created in 2 adjacent vertebral bodies on either side of the herniated disk and extruded/protruded material was removed. Data collected included history, duration of clinical signs, presurgical assessment of neurologic status, postsurgical neurologic status, complications, and outcome. RESULTS: Ambulatory capacity was maintained or regained, and neurologic status improved by 1 grade (3 dogs), 2 grades (8), 3 grades (2), or 4 grades (2). Eleven dogs were considered free of disease. A seroma in 1 dog was the sole complication observed. CONCLUSIONS: Lateral corpectomy permits relatively easy removal of protruded-extruded disk material from within the vertebral canal in chronic disk disease without further iatrogenic injury to the spinal cord. CLINICAL RELEVANCE: Lateral corpectomy is an alternative to dorsal decompression for treatment of ventral and lateroventral thoracolumbar chronic disk disease in dogs. PMID- 15659019 TI - Use of a semitendinosus myocutaneous flap for soft-tissue reconstruction of a grade IIIB open tibial fracture in a dog. AB - OBJECTIVE: To describe the clinical use of a semitendinosus myocutaneous flap for soft-tissue reconstruction of a grade IIIB open tibial fracture. STUDY DESIGN: Case report. ANIMALS: A 5-year-old castrated male Labrador retriever with a grade IIIB open tibial fracture. METHODS: A myocutaneous flap was created by elevating the origin of the semitendinosus muscle and the associated overlying skin. The flap was rotated distally based on the distal vascular pedicle into a soft-tissue defect overlying the central and distal third of the tibia. RESULTS: The flap allowed one-stage reconstruction of a severe soft-tissue defect overlying denuded bone in a grade IIIB open tibial fracture. The muscular portion of the flap survived; however, there was necrosis of 3 cm of skin extending distally beyond the muscular portion of the flap. CONCLUSION: Distal rotation of the semitendinosus muscle and overlying skin can be used in the treatment of severe soft-tissue defects of the canine crus. CLINICAL RELEVANCE: Complex soft-tissue injuries to the canine crus can be treated with a myocutaneous flap as a reasonable alternative to other reconstructive techniques. PMID- 15659021 TI - Variables associated with outcome in dogs undergoing extrahepatic biliary surgery: 60 cases (1988-2002). AB - OBJECTIVE: To report clinical findings and define clinical variables associated with outcome in dogs undergoing extrahepatic biliary surgery. STUDY DESIGN: Retrospective study. ANIMALS: Sixty dogs that had extrahepatic biliary tract surgery. RESULTS: Primary diagnoses included necrotizing cholecystitis (36 dogs, 60%), pancreatitis (12 dogs, 20%), neoplasia (5 dogs, 8%), trauma (4 dogs, 7%), and gallbladder rupture from cholelithiasis without necrotizing cholecystitis (3 dogs, 5%). Bile peritonitis occurred in 19 (53%) dogs with necrotizing cholecystitis, 4 dogs with trauma, and 3 dogs with cholelithiasis without evidence of necrotizing cholecystitis. Cholecystectomy (37 dogs, 62%) and cholecystoduodenostomy (14 dogs, 23%) were the 2 most commonly performed procedures. Median hospitalization for survivors was 5 days (range, 1-15 days). There were 43 surviving dogs (72%) and 17 nonsurvivors (28%, 4 died, 13 euthanatized). Presence of septic bile peritonitis (P=.038), elevation in serum creatinine concentration (P=.003), prolonged partial thromboplastin times (PTTs; P=.003), and lower postoperative mean arterial pressures (P=.0001) were significantly associated with mortality. CONCLUSIONS: Extrahepatic biliary surgery is associated with high mortality and a relatively long hospitalization time for survivors. Cholecystectomy and cholecystoduodenostomy were the most common surgical procedures to treat the 4 major biliary problems (necrotizing cholecystitis, pancreatitis, neoplasia, and trauma) observed in this cohort of dogs. The relatively high mortality rate likely reflects the underlying diseases and their effects on the animal (septic bile peritonitis, higher serum creatinine, prolonged PTT, and lower postoperative mean arterial pressure) rather than complications of surgery. CLINICAL RELEVANCE: Septic bile peritonitis, preoperative elevated creatinine concentration, and immediate postoperative hypotension in dogs undergoing extrahepatic biliary tract surgery are associated with a poor clinical outcome. Adequate supportive care and monitoring in the perioperative period is critical to improve survival of dogs with extrahepatic biliary disease. PMID- 15659020 TI - A prospective comparison between stabilized glutaraldehyde and chlorhexidine gluconate for preoperative skin antisepsis in dogs. AB - OBJECTIVE: To compare the efficacy of 0.3% stabilized glutaraldehyde and alcohol (SG+A), 0.3% SG and water (SG+W), and 4% chlorhexidine gluconate tincture (CG+A), as skin disinfectants in dogs undergoing ovariohysterectomy. STUDY DESIGN: Prospective, blinded clinical study. ANIMALS: One hundred and twenty-one dogs. METHODS: Cutaneous bacterial colony forming units (CFU) from the perioperative site after skin preparation, after antisepsis, and after surgery (incisional and paramedian), were quantified. The influence of high initial bacterial counts (> or =150 CFU) and surgical time on antibacterial efficacy was examined and the proportion of dogs from which Staphylococcus intermedius was cultured, determined. Perioperative skin reactions and wound infections were documented. RESULTS: All 3 antiseptic solutions significantly and equally reduced CFU to all post-antisepsis sampling levels irrespective of surgical duration (mean surgical times 151.6, 136.2, and 149.6 minutes for CG+A, SG+A and SG+W, respectively). Median percentage reductions in CFU ranged between 99.3% and 100%. In dogs with initial high counts and disinfected with CG+A and SG+W, the incisional samples had significantly higher counts than the post-antisepsis samples. In the CG+A and SG+W groups, the proportion of post-surgery samples yielding S. intermedius was significantly higher at the incisional than the paramedian sites. Eight mild cutaneous reactions were recorded in equal proportions for the 3 solutions. There were no recorded infections. CONCLUSIONS: All 3 preparations had an equal ability to reduce and maintain low CFU counts, with minimal cutaneous reactions. CLINICAL RELEVANCE: SG solutions are safe and effective preoperative skin antiseptics for elective clean-contaminated surgical procedures. PMID- 15659023 TI - Caprine obstructive urolithiasis: requirement for 2nd surgical intervention and mortality after percutaneous tube cystostomy, surgical tube cystostomy, or urinary bladder marsupialization. AB - OBJECTIVES: To determine the requirement for 2nd surgical interventions and mortality after 3 different surgical techniques (percutaneous tube cystostomy [10 goats], surgical tube cystostomy [25 goats], urinary bladder marsupialization [10 goats]) for caprine obstructive urolithiasis, and to determine whether pre- or 24 hour postoperative physical examination findings or serum chemistry results could be used as predictors of mortality. STUDY DESIGN: Retrospective study. ANIMALS: Male goats (45) with obstructive urolithiasis. METHODS: Medical records for all male goats admitted and operated for obstructive urolithiasis between 1993 and 2003 were reviewed. Data retrieved included signalment, pre- and 24-hour postoperative values for temperature, pulse, respiratory rate, packed cell volume, serum K(+), serum creatinine, and blood urea nitrogen [BUN]. The type of initial surgical procedure, time to 2nd surgical intervention, time to death, and duration and cost of hospitalization were also obtained. RESULTS: Percutaneous tube cystostomy was associated with a significantly increased requirement for (5.6-fold increased hazard) and decreased time to a 2nd intervention (P=.002). There were no differences between the 3 procedures for time to mortality. Increased BUN and respiratory rate at admission were associated with increased mortality (hazards ratio of 4.8 and 5.0, respectively). Urinary bladder marsupialization was associated with significantly decreased hospitalization time (P=.02) and cost (P=.04) compared with surgical tube cystostomy and percutaneous tube cystostomy. CONCLUSION: Surgical tube cystostomy and bladder marsupialization are both acceptable surgical methods for treatment of caprine obstructive urolithiasis. Each procedure has inherent complications that should be discussed with the owner before choosing the surgical treatment. CLINICAL RELEVANCE: There are acceptable options for treatment of caprine obstructive urolithiasis; however, percutaneous tube cystostomy should be avoided. PMID- 15659022 TI - Analgesic comparison of meloxicam or ketoprofen for orthopedic surgery in dogs. AB - OBJECTIVE: To compare the safety and efficacy of 2 analgesic protocols (preoperative meloxicam or intraoperative ketoprofen administration) during the first 24 hours after orthopedic surgery in dogs. STUDY DESIGN: Double-blind, prospective randomized clinical trial. ANIMALS: Sixty client-owned dogs. METHODS: Dogs with surgical orthopedic disorders were randomly separated into 2 groups: 30 dogs were administered 0.2 mg/kg meloxicam intravenously (IV) immediately before induction and 30 dogs were administered 2 mg/kg ketoprofen IV, 30 minutes before the end of surgery. Pain was assessed with a visual analog scale (VAS) and a cumulative pain score (CPS) preoperatively and at 30 minutes, 1, 2, 3, 4, 6, 8, and 24 hours after extubation. Selected serum biochemical variables were measured before and 24 hours after surgery and, buccal mucosal bleeding time (BMBT) and whole blood clotting time (WBCT) were measured before and 8 hours after surgery. Dogs were anesthetized with propofol and maintained on halothane in oxygen. Any complications were documented for 7 days after surgery. Results were compared between the 2 groups for significant differences in VAS scores (2-sample t-test) and in CPS (Wilcoxon's 2-sample test). Moreover, results were analyzed for significant differences in area under the curve (AUC) for VAS (2-sample t-test) and CPS (Wilcoxon's 2-sample test) among groups. To assess the effects of treatments on biochemical and coagulation functions, pre- and postoperative mean values of BMBT and WBCT were compared within both treatment groups (paired t tests) and between both groups (2-sample t-test). RESULTS: No significant differences in pain response or coagulation were found between meloxicam- and ketoprofen-treated dogs. In both groups, alkaline phosphatase and alanine aminotransferase concentrations were significantly increased compared with baseline. No serious complications occurred. CONCLUSIONS: Preoperative administration of meloxicam is a safe and effective method of controlling postoperative pain for up to 24 hours in dogs undergoing orthopedic surgery. CLINICAL RELEVANCE: Analgesia after administration of preoperative meloxicam was comparable with administration of ketoprofen at the end of the surgery. PMID- 15659024 TI - Evaluation of a sheep laparoscopic uterine trauma model and repeat laparoscopy for evaluation of adhesion formation and prevention with sodium carboxymethylcellulose. AB - OBJECTIVE: To evaluate topical application of 1% sodium carboxymethyl cellulose (SCMC) for prevention of postoperative adhesions in a laparoscopic model of uterine trauma in sheep. STUDY DESIGN: Experimental study. ANIMALS: Fourteen non pregnant ewes. METHODS: Ewes were randomly assigned to 1 of 2 groups: control (saline solution) or 1% SCMC treatment. By left flank laparoscopy, traumatic forceps were used to create serosal trauma (1.5 x 5 cm) and hemorrhage on the left uterine horn. Either 30 mL saline solution or 30 mL 1% SCMC was applied topically to the traumatized uterine horn. Adhesion formation was evaluated by repeat laparoscopy at days 14 and 21. Sheep were euthanatized on day 28 for necropsy evaluation of adhesions. RESULTS: Five control sheep had adhesions of the uterine horn by day 14, but only 4 had adhesions at day 21, and 2 at day 28. Adhesions did not occur in SCMC-treated sheep. No adhesions occurred elsewhere in the abdomen. CONCLUSIONS: Laparoscopically created uterine trauma is an effective method for induction of uterine adhesions, and laparoscopy is an excellent method for serial evaluation of adhesion formation. SCMC (1%) was effective at preventing adhesion formation in sheep and no inflammatory response was noted. CLINICAL RELEVANCE: SCMC (1%) should be considered for prevention of adhesions in abdominal surgery in sheep. PMID- 15659025 TI - Two techniques for supplementing interlocking nail repair of fractures of the humerus, femur, and tibia: results in 12 dogs and cats. AB - OBJECTIVE: To describe 2 devices for improving stabilization of inadequately stabilized interlocking nail (ILN) repairs of the humerus, tibia, and femur in dogs and cats. STUDY DESIGN: Prospective study. ANIMALS: Twelve client-owned dogs and cats. METHODS: Two devices to further stabilize ILN repair of inadequately stabilized diaphyseal fractures were developed. Device 1 was an axial extension for the ILN that was connected to a conventional type I external skeletal fixator (ESF) with a short connecting bar. Device 2 had hybrid ILN bolt/ESF pins that were used to lock the ILN and serve as the pins for a type I ESF. Devices were used at the initial surgery when the stability of ILN repair was considered inadequate based on palpable fracture segment movement, insufficient medullary canal filling of the ILN at the fracture site, or when the ILN was used in a buttress mode. Outcome was obtained by recheck examinations, radiography, and telephone interview. RESULTS: Device 1 was applicable to fractures of the humerus and femur, but was not used for fractures of the tibia because the ILN extension would have interfered with the stifle. No gross loosening of the ILN/ESF extension connection to the ILN occurred. Device 2 was easily placed and used in the humerus, femur, and tibia. Device 2 allowed removal of the ILN interlock to one or both main fracture segments non-invasively. Clinically, both devices added stability compared with ILN repair alone. Both devices facilitated controlled destabilization of the fracture repair as healing progressed. Complications of pin tract infection, and premature hybrid bolt/ESF pin loosening resulting in premature ESF removal each occurred in 1 patient. Four of 28 hybrid ILN/ESF pins were grossly loose at 4- or 6-week postoperative recheck examinations. Outcomes were excellent (9), good (1), fair (1), and poor (1). CONCLUSIONS: Inadequately stabilized ILN repair of fractures can be stabilized by use of either device, both of which also permit controlled destabilization of the repair during healing. Device 2 can be used when non-invasive removal of the ILN interlock is desired during healing. CLINICAL RELEVANCE: These 2 devices should be considered as alternative methods for stabilization of inadequately stabilized ILN repairs in dogs and cats, or when controlled destabilization of an ILN fracture repair is desired. PMID- 15659026 TI - Re-evaluation of a portocaval venograft without an ameroid constrictor as a method for controlling portal hypertension after occlusion of intrahepatic portocaval shunts in dogs. AB - OBJECTIVE: To evaluate the use of a portocaval venograft without an ameroid constrictor in the surgical management of intrahepatic portosystemic shunts (PSS). STUDY DESIGN: Prospective clinical study. ANIMALS: Seven dogs with intrahepatic PSS. METHODS: Portal pressure was measured after temporary suture occlusion of the intrahepatic PSS. In dogs with an increase in portal pressure > or =8 mm Hg or signs of portal hypertension, a single extrahepatic portocaval shunt was created using a jugular vein. Clinical outcome and complications were recorded. RESULTS: The mean (+/-SD) portal pressure increased from 5.9+/-1.6 to 17.9+/-4.1 mm Hg with PSS occlusion. There were no intraoperative complications and, after creation of the portocaval shunt, the intrahepatic PSS could be completely ligated in all dogs. The final portal pressure was 9.6+/-1.9 mm Hg. Complications developed during postoperative hospitalization in 5 dogs and included incisional discharge (4 dogs), ascites (3), ventricular premature contractions (2), and melena, bloody diarrhea, neurologic signs, coagulopathy, and aspiration pneumonia (each in 1 dog). Six dogs died or were euthanatized with clinical signs related to depression, inappetance, abdominal pain, vomiting, melena, and abdominal distention, with a median survival of 82 days (range, 20 990 days). One dog was clinically normal at 33 months after surgery. CONCLUSIONS: Clinical signs observed in 6 dogs after surgery were consistent with portal hypertension. Use of a portocaval venograft without an ameroid constrictor may reduce the likelihood of hepatic vascular development, thereby increasing the risk of life-threatening portal hypertension should the venograft suddenly occlude. CLINICAL RELEVANCE: Use of a portocaval venograft without an ameroid constrictor to control portal hypertension after ligation of an intrahepatic PSS cannot be recommended. PMID- 15659027 TI - An in vitro biomechanical comparison of the insertion variables and pullout mechanical properties of ao 6.5-mm standard cancellous and 7.3-mm self-tapping, cannulated bone screws in foal femoral bone. AB - OBJECTIVE: To compare screw insertion variables and pullout mechanical properties between AO 6.5-mm cancellous and 7.3-mm cannulated bone screws in foal femoral bone. STUDY DESIGN: A paired, in vitro mechanical study. SAMPLE POPULATION: Seven pairs of femora from immature (1-7 months) foals. METHODS: The 6.5 cancellous and 7.3-mm cannulated screws were inserted at standardized proximal and distal metaphyseal, and mid-diaphyseal locations. Insertion torque, force, and time to drill, tap (6.5-mm cancellous), guide wire insertion (7.3-mm cannulated), and screw insertion were measured. Screw pullout properties (yield and failure load, displacement, and energy, and stiffness) were determined from mechanical tests. The effects of screw type and location on insertion variables and pullout properties were assessed with repeated measures ANOVA. Pairwise comparisons were examined with post hoc contrasts. Significance was set at P<.05 for all comparisons. RESULTS: Insertion torques for the 7.3-mm cannulated screws were significantly greater than for the 6.5-mm tap, but significantly lower than for the 6.5-mm cancellous screws. Total screw insertion times were similar. Pullout properties of both screws were similar at each femoral location. The holding power of both screws was significantly greater in the mid-diaphysis than in either metaphyseal location. Pullout failure occurred by bone shearing at the bone-screw interface in all specimens. CONCLUSIONS: The 6.5-mm cancellous and 7.3 mm cannulated screws vary in insertion properties, but have similar pullout properties in the mid-diaphysis, proximal, and distal metaphysis of foal femora. Both screw types have greater holding power at the mid-diaphyseal location compared with metaphyseal locations. Based on overall similar holding powers of 6.5-mm cancellous and 7.3-mm cannulated screws, it is unlikely that increasing the screw diameter beyond 6.5 mm will provide increased holding power in foal femoral bone. CLINICAL RELEVANCE: Use of the 7.3-mm cannulated screw should be considered for foal femoral fracture repair when greater accuracy is needed, or when bone threads for the 6.5-mm cancellous screw have been stripped. PMID- 15659030 TI - Tissue repair and regeneration in China: research focus and society. PMID- 15659032 TI - Annual costs of treatment for venous leg ulcers in Sweden and the United Kingdom. AB - The aim of this study was to estimate costs of treating venous leg ulcers in Sweden and the United Kingdom during 1 year and to quantify costs in different health states. The costs of treating four different types of venous leg ulcers were estimated for 52 weeks by a stochastic health economic model, which simulated resource use data obtained from prospectively collected patient data, expert panels in the two countries, and published scientific literature. The average cost of treating an ulcer varied between 1332 Euro and 2585 Euro in Sweden and from 814 Euro to 1994 Euro in the United Kingdom. Cost of treating large ulcers (>/= 10 cm(2)) of long duration (>/= 6 months) was highest in both countries. Frequency of dressing changes and duration of time for each dressing change were higher in Sweden than in the United Kingdom, resulting in higher total cost per patient in Sweden. An important factor for the total costs was time to heal. Other important variables influencing treatment costs were frequency and duration of dressing changes. Actions to reduce time used for dressing changes and the total time to healing are thus very important in reducing costs spent on treatment of venous leg ulcers in both countries. PMID- 15659031 TI - Tissue repair, contraction, and the myofibroblast. AB - After the first description of the myofibroblast in granulation tissue of an open wound by means of electron microscopy, as an intermediate cell between the fibroblast and the smooth muscle cell, the myofibroblast has been identified both in normal tissues, particularly in locations where there is a necessity of mechanical force development, and in pathological tissues, in relation with hypertrophic scarring, fibromatoses and fibrocontractive diseases as well as in the stroma reaction to epithelial tumors. It is now accepted that fibroblast/myofibroblast transition begins with the appearance of the protomyofibroblast, whose stress fibers contain only beta- and gamma-cytoplasmic actins and evolves, but not necessarily always, into the appearance of the differentiated myofibroblast, the most common variant of this cell, with stress fibers containing alpha-smooth muscle actin. Myofibroblast differentiation is a complex process, regulated by at least a cytokine (the transforming growth factor beta1), an extracellular matrix component (the ED-A splice variant of cellular fibronectin), as well as the presence of mechanical tension. The myofibroblast is a key cell for the connective tissue remodeling that takes place during wound healing and fibrosis development. On this basis, the myofibroblast may represent a new important target for improving the evolution of such diseases as hypertrophic scars, and liver, kidney or pulmonary fibrosis. PMID- 15659033 TI - Transcriptional events in a clinical model of oral mucosal tissue injury and repair. AB - Tissue injury in the oral mucosa activates a cascade of transcriptional events important during the healing process that are not yet clearly defined. To characterize these events and identify potential gene targets for future studies, we used cDNA expression arrays in a clinical model of tissue injury. Mucosal biopsies were taken before third molar extraction, 2-4 hours postoperatively, or at 48 hours. Hybridization patterns were analyzed and validated using real-time polymerase chain reaction. Prior to extraction, the biopsied mucosal tissues were characterized by a panoply of genes that were constitutively expressed. After injury, analysis revealed differential expression of genes involved in transcription, inflammation, and remodeling. At 2-4 hours after injury, genes such as Fos, Jun, and early growth response protein were up-regulated, while genes responsible for intercellular adhesion were down-regulated. At 48 hours after injury, the gene profile had shifted toward tissue remodeling. Here we identify genes constitutively expressed in normal oral mucosa and transcriptional events following mucosal tissue injury, which may be useful in identifying new therapeutic targets. PMID- 15659034 TI - Variable impairment of wound healing in the heterozygous collagenase-resistant mouse. AB - Collagen undergoes dramatic reorganization during wound repair. Matrix metalloproteinases degrade and remodel collagen in a tightly controlled process. The collagenase-resistant mouse, Col1a1(tm1Jae), produces type I collagen, which is resistant to degradation by human matrix metalloproteinase 1. These mice grow normally but develop thickened skin with age. We have previously reported that the early wound repair response in homozygous mutant (Col1a1(r/r)) mice is delayed compared to wild type (Col1a1(+/+)). However, the late-stage scar of Col1a1(r/r) wounds was not significantly altered compared to Col1a1(+/+). Here we have investigated the response of heterozygous mice (Col1a1(+/r)) to wounding, not previously reported. Wound reepithelialization was delayed to a similar degree to wounds in the Col1a1(r/r) mice. However, the recovery of impaired wound contraction was faster in Col1a1(+/r) than in Col1a1(r/r) mice, but still slower than in wild-type animals. Analysis of wound protein extracts showed expression of some matrix metalloproteinases was prolonged in both the Col1a1(r/r) and Col1a1(+/r) wounds compared to wild type. We suggest the partial resistance of collagen to collagenase-mediated degradation in the heterozygous animals causes equivalent impairment of keratinocyte migration compared to homozygous collagenase-resistant mice, but that wound contraction during late-stage healing is only partially retarded. PMID- 15659035 TI - Expression of different p63 variants in healing skin wounds suggests a role of p63 in reepithelialization and muscle repair. AB - Healing of skin wounds in mammals involves partial reconstruction of the dermis and coverage of the injured site by keratinocytes. The latter process is achieved by extensive migration and hyperproliferation of keratinocytes at the wound rim. Because the p53 protein family member p63 is expressed in human hyperproliferative epidermis, this study determined whether enhanced keratinocyte proliferation correlates with the expression of p63. Therefore, we investigated the temporal and spatial distribution of four major variants of the p63 transcription factor-TAp63alpha, TAp63gamma, DeltaNp63alpha and DeltaNp63gamma during normal skin wound healing in mice. Transcripts encoding amino-terminally truncated DeltaNp63 variants were found at high levels in basal and suprabasal keratinocytes of the hyperproliferative wound epithelium. Interestingly, TAp63 variants, which include the conserved transactivation domain TA at their amino terminus, were also expressed in wound keratinocytes as well as at the edge of the injured subcutaneous muscle panniculus carnosus. These findings suggest splice-variant specific functions of p63 in reepithelialization and muscle repair. PMID- 15659036 TI - In vivo gene delivery of Ad-VEGF121 to full-thickness wounds in aged pigs results in high levels of VEGF expression but not in accelerated healing. AB - We have previously reported that endogenous vascular endothelial growth factor (VEGF) concentration in older pig wounds peaked later and at one-fourth the level of young pigs. These data suggested that VEGF might play a major role in the healing of full-thickness wounds in the aged pig. By in vivo gene transfer using the microseeding technique, we treated full-thickness wounds with different doses of VEGF-expressing adenoviral vector (Ad-VEGF) varying from 1 x 10(7) to 2.7 x 10(11) particles per wound (ppw). We found that the VEGF expression in wound fluid followed a dose-response pattern. However, when wounds were microseeded with the highest concentration of Ad-VEGF (2.7 x 10(11) ppw), diminished healing rates were found. We then determined the minimal functional concentrations of Ad VEGF. We used five aged Yucatan minipigs, all retired breeders, to analyze the role of over-expression of 1 x 10(8) and 1 x 10(9) ppw of Ad-VEGF (n= 78) in terms of healing of full-thickness wounds, all 2.5 x 2.5 x 1 cm in size (n= 158). The Ad-VEGF solutions were delivered to the wound floor and borders by in vivo microseeding. Control wounds (n= 80) were microseeded with Ad-Lac-Z (n= 25), treated with saline (n= 49) or treated dry (n= 6). All wounds except for the dry treated ones were covered with a wound chamber and a wet environment was created by injecting 2.5 ml saline into the chamber. Peak VEGF expression (2300-4000 pg/ml) was detected on days 2 or 3 post gene delivery. This level of VEGF expression was not seen in the saline (n= 49) or Ad-null (n= 25) control groups. The VEGF expression in wounds treated with 1 x 10(8) and 3 x 10(8) ppw (n= 39) exhibited a slower onset with a peak concentration of 400-920 pg/ml on days 5-7. Although high levels of VEGF expression were achieved in the local wound environment, we could not show a significant increase in neovascularization as compared to saline-treated wounds. No significant differences were observed in the rate of reepithelialization and wound contraction among groups of full thickness wounds treated with Ad-VEGF, Ad-null mutant, or saline in the aged "wet wound healing" pig model. These results indicate that increased levels of VEGF in wounds produced by in vivo gene transfer have little effect on the healing of full-thickness wounds in the aged pig model. Moreover, significantly higher levels of VEGF expression by Ad-VEGF could lead to impaired wound healing. PMID- 15659037 TI - Transforming growth factor-beta and its effect on reepithelialization of partial thickness ear wounds in transgenic mice. AB - Transforming growth factor-beta (TGF-beta) is known to affect nearly every aspect of wound repair. Many of the effects have been extensively investigated; however, the primary effect of endogenously derived TGF-beta on wound reepithelialization is still not completely understood. To examine this, two types of wounds were made on a transgenic mouse over-expressing TGF-beta1. Full-thickness back wounds were made to compare the wound healing process in the presence of compensatory healing mechanisms. Superficial partial-thickness ear wounds involving only the epidermis were made to determine the effect of TGF-beta on reepithelialization. In the partial-thickness ear wounds, at later time points, the transgenic group had smaller epithelial gaps than the wild-type mice. A greater number of actively proliferating cells, as determined by bromodeoxyuridine incorporation, was also found in the transgenic mice at post-injury day 8. These results show that TGF beta1 stimulates the rate of reepithelialization at later time points in partial thickness wounds. However, in the full-thickness back wounds, the transgenic animals exhibited a slower reepithelialization rate at all time points and the number of bromodeoxyuridine-positive cells was fewer. Our findings would suggest that the overexpression of TGF-beta1 speeds the rate of wound closure in partial thickness wounds by promoting keratinocyte migration. In full-thickness wounds, however, the overexpression of TGF-beta1 slows the rate of wound reepithelialization. PMID- 15659038 TI - Ontogeny of expression of transforming growth factor-beta and its receptors and their possible relationship with scarless healing in human fetal skin. AB - Fetal cutaneous wounds that occur in early gestation heal without scar formation. Although much work has been done to characterize the role of transforming growth factor-beta (TGF-beta) isoforms and their receptors in the wound healing process, their roles in scarless wound repair observed in early gestation and their functions in human fetal skin development, and structural and functional maintenance are still not well understood. In this study, we explore the expression and distribution characteristics of three TGF-beta isoforms and their receptors, TGF-betaRI (TBRI) and TGF-betaRII (TBRII), in fetal and postnatal skins to understand the relevance of these five proteins to skin development and elucidate the mechanism(s) underlying the phenotypic transition from scarless to scar-forming healing observed during fetal gestation. Fetal skin biopsies of human embryo were obtained from spontaneous abortions at different gestational ages from 13 to 32 weeks and postnatal skin specimens were collected from patients undergoing plastic surgery. Gene expression and positive immunohistochemical signals of TGF-beta(1), TGF-beta(2), TGF-beta(3), TBRI, and TBRII could all be detected in fetal and postnatal skins. In early gestation, gene expression of TGF-beta(1), TBRI, and TBRII was weaker and protein contents were less compared with postnatal skins (p < 0.05). In contrast, more TGF-beta(2) mRNA transcript was found in early gestation than in late gestation and in postnatal skins, whereas protein content of this growth factor increased during gestation. Lastly, mRNA transcript and protein contents of TGF-beta(3) were apparently higher in early gestation compared to postnatal skin (p < 0.05). In postnatal skin, granules containing the three TGF-beta isoforms were mainly distributed in the cytoplasm and extracellular matrix of epidermal cells, interfollicular keratinocytes, and some fibroblasts. TBRI and TBRII were chiefly located in the cellular membrane of epidermal keratinocytes and some fibroblasts. The endogenous three TGF-beta isoforms and their receptors may be involved in the development of embryonic skin and in the maintenance of cutaneous structure and function, and also in postnatal wound healing. The differential levels of TGF beta isoforms may provide either a predominantly antiscarring or profibrotic signal upon wounding depending on the gestational period. Lower expression of their receptors in early gestational skins may be a reason for the reduced ability to perceive ligands, ultimately leading to scar-free healing. PMID- 15659039 TI - Cultured pressure ulcer fibroblasts show replicative senescence with elevated production of plasmin, plasminogen activator inhibitor-1, and transforming growth factor-beta1. AB - In a 16-patient study, cultured fibroblast populations from normal skin were able to replicate an average of 14.8 +/- 2.2 times before becoming senescent, while fibroblast populations from the ulcer bed reached the end of their replicative life span after 7.2 +/- 1.9 population doublings (p= 0.001). Fibroblast populations from 10 of 16 pressure ulcers became senescent after fewer than five population doublings, whereas when populations of fibroblasts from adjacent normal skin were studied, only 2 of 16 became senescent within this same time period. In addition, only an occasional fibroblast from normal skin stained positively for senescence-associated beta-galactosidase compared to approximately 50% of equally aged ulcer bed fibroblasts (p = 0.0060). Senescent ulcer bed fibroblasts secreted significantly more plasmin than early passage ulcer bed fibroblasts (p= 0.0237), nearly six times as much plasmin as early passage normal skin fibroblasts (p < 0.0001), three and a half times the level of normal skin fibroblasts of the same age (11.52 +/- 4.58 microg/mg protein; p= 0.0003), and more than one and a half times the level of senescent normal skin fibroblasts (p= 0.0525). Senescent pressure ulcer fibroblasts generated significantly more plasminogen activator inhibitor-1 (1179.27 +/- 25.37 ng/mg protein) than normal skin fibroblasts of the same age (132.16 +/- 16.20 ng/mg protein; p = 0.0357). Also, senescent ulcer bed fibroblasts produced higher levels of transforming growth factor-beta1, but these were not significantly different from senescent normal skin fibroblasts. Although senescent ulcer fibroblasts produce elevated levels of plasminogen activator inhibitor-1 and transforming growth factor-beta1, the ratio of these factors to plasmin levels suggests that this may have little influence on extracellular matrix synthesis or maintenance in the chronic wound. These data show that cultured fibroblasts from most patient pressure ulcers profile a wound environment that is associated with an increasing population of senescent fibroblasts; however, factors within the chronic wound environment that promote cellular senescence remain unclear. We have proposed that a prolonged inflammatory response may be a contributing factor to the chronic wound condition. PMID- 15659040 TI - Wound matrix attachment regulates actin content and organization in cells of the granulation tissue. AB - Actin cytoskeletal polymerization is associated with a pro-proliferative, pro survival state. We hypothesized that the actin polymerization of wound cells is increased in the presence of wound matrix attachment and is decreased after disruption of this attachment. Musculocutaneous flap and wound splinting models were used to investigate the effect of wound matrix attachment on the actin cytoskeleton. Disruption of wound matrix attachment was accomplished by incision of the wound matrix/dermis interface (wound matrix release) and/or desplinting. Polymerized actin was assayed with phalloidin labeling of wound specimens 24 hours after disruption of attachment and a method to quantify the content and organization of polymerized actin in granulation tissue was used. Disruption of wound matrix attachment decreased the content of polymerized actin, the actin staining intensity, and the actin fiber organization in the granulation tissue of both the flap and splint models. Disruption of wound matrix attachment decreased actin polymerization and fiber organization in the granulation tissue. Our data support the concept that the state of wound matrix attachment regulates the actin cytoskeleton of wound cells. PMID- 15659041 TI - Migration of keratinocytes is impaired on glycated collagen I. AB - Advanced glycation end products are the chemical modification of proteins induced by sugars in a hyperglycemic condition. Extracellular matrix proteins are prominent targets of nonenzymatic glycation because of their slow turnover rates. The aim of this study was to investigate the influence of nonenzymatic glycation of type I collagen on the migration of keratinocytes. The migration of keratinocytes was dramatically promoted on native type I collagen-coated dishes compared with that on uncoated dishes. When type I collagen was glycated with glycolaldehyde, large amounts of advanced glycation end products were produced; the glycated collagen I-coated dishes did not promote the migration of keratinocytes. Glycated collagen I did not affect the proliferative capacity of keratinocytes. However, the adhesion of keratinocytes to glycated collagen I was profoundly diminished in a glycation intensity-dependent manner. alpha2beta1 integrin is responsible for the migration and adhesion of keratinocytes to type I collagen. Pretreatment with glycated collagen I did not affect the expression level or functional activity of alpha2beta1 integrin on keratinocytes. These findings suggest that in the presence of glycated collagen I, keratinocytes lose their adhesive and migratory abilities. As the glycation did not modify the alpha2beta1 integrin on keratinocytes, it is suggested that glycation may diminish the binding capacity of type I collagen. PMID- 15659042 TI - Epidermal stem cells are the source of sweat glands in human fetal skin: evidence of synergetic development of stem cells, sweat glands, growth factors, and matrix metalloproteinases. AB - The development of sweat glands is a complex biological process, and the extent of cellular trafficking between epidermal stem cells and the development of sweat glands is uncertain. Therefore, we studied the synergetic development effects of stem cells, sweat glands, growth factors, and matrix metalloproteinases (MMPs) in human skin. Human fetal skin was obtained from spontaneously aborted fetuses at 11-31 weeks of gestation. Paraffin sections were cut and stained with hematoxylin and eosin or immunostained with antibodies against beta(1) integrin, keratin (K) 19 and K7, MMP-2 and -7, and epidermal growth factor. In situ hybridization was used along with semiquantitative analysis of the positive expression of these proteins to analyze for mRNA expression of MMP-2 and -7. Histological studies revealed the fetal epidermis began to form a primary epidermal ridge at gestational age 13-14 weeks and these primordial basal cells became tightly packed to take the form of multiple hillocks between 14 and 16 weeks. Furthermore, these cells gave rise to chord-like columnar buds in the embryonic epidermis, and these buds gradually migrated downward into the dermis to form juvenile sweat glands at 18-20 weeks. Mature sweat glands were found in the fetal epidermis at the end of 24 weeks. beta(1) integrin and K19 immunoreactivities were first detected in those cells that gathered together to form primary epidermal ridges, including sweat gland cords, buds, and immature sweat gland cells. The positive immunostaining for K7 appeared in early sweat gland buds at 14-16 weeks, and from then on K7 was concentrated in developing sweat gland cords or cells. At 14-16 weeks, positive epidermal growth factor, MMP-2, and MMP-7 expression was first observed weakly in developing sweat gland buds. The immunoreactivity of these proteins was then gradually increased in the developing sweat gland buds and extracellular stroma from 14 to 20 weeks. The intensity of the positive signal peaked at 20-22 weeks of gestational age. After that, the intensity of immunostaining for MMP-2 and MMP-7 proteins was gradually weakened. However, the expression of epidermal growth factor did not show an apparent decrease. These results suggest that epidermal stem cells are the source of sweat glands. Epidermal growth factor is one of the main inducers in the development and maturity of sweat gland buds or cells and the local activated MMPs may play an important role in cleaving the major matrix components in the basement membrane. PMID- 15659043 TI - Involvement of bone marrow-derived cells in healing of experimental colitis in rats. AB - Bone marrow is reported to contain hematopoietic stem cells and other adult somatic stem cells that have phenotypes of cells composing tissues other than bone marrow. To explore the implication of bone marrow-derived cells in the treatment of inflammatory bowel diseases, experimental colitis was induced in wild-type rats after transplantation of bone marrow from transgenic rats expressing green fluorescence protein (GFP). Chronic colitis was induced 21 days later using 30 mg 2,4,6-trinitrobenzenesulfonic acid (TNBS). Control rats received saline. At 28, 56, and 224 days after TNBS administration, rats were euthanized, and tissues were removed and processed for paraffin-embedded sections. Cells derived from bone marrow were identified by immunohistochemistry using anti-GFP antibody. To identify the phenotypes of the cells expressing GFP, we conducted serial-section analysis and double-staining analysis using antibodies against cytokeratin (epithelial cells) or vimentin (interstitial cells). In the present study, GFP-positive, bone marrow-derived cells occupied 37.6% and 4.25% of the colonic epithelium at 28 days and 56 days after the induction of TNBS-colitis, respectively. Also, significant amounts of mucosal and submucosal interstitial cells were derived from the bone marrow. These findings showed that a large amount of bone marrow-derived cells were involved in regeneration of the colon after experimental colitis in rats. PMID- 15659045 TI - Effects of selective dopamine receptor subtype agonists on cardiac contractility and regional haemodynamics in rats. AB - 1. Activation of dopamine (DA) receptors produces cardiovascular responses such as vasodilation and hypotension. However, knowledge of the role of specific dopamine receptor subtypes (especially D3 and D4) in the cardiovascular system is limited. The objective of the present study was to characterize the haemodynamic and cardiac responses to agonists with selectivity for D1, D2, D3 and D4 receptor subtypes. 2. Inactin-anaesthetized rats were instrumented to measure regional haemodynamic and cardiac contractility responses with slow intravenous infusion of agonists. 3. Fenoldopam (a D1 receptor agonist) decreased (P < 0.05) renal vascular resistance beginning at a dose of 3 micromol/kg. Infusion of PNU-95666E (a D2 receptor agonist) produced dose-dependent decreases (P < 0.05) in mean arterial pressure (MAP), heart rate (HR) and hindquarter vascular resistance (HQVR). Administration of BP897 (a partial D3 receptor agonist) decreased (P < 0.05) MAP and HQVR at 3 micromol/kg. PD168077 (a D4 receptor agonist) caused significant increases in HQVR at 1 micromol/kg. None of the compounds tested elicited significant changes in cardiac contractility. 4. Using selective agonists of dopamine receptor subtypes, the present studies characterize distinct cardiovascular effects in anaesthetized rats. Consistent with its well-defined effects as a D1 receptor agonist, fenoldopam administration resulted in renal vasodilation. Similar to earlier studies using the non-selective D2-like receptor agonist quinpirole, selective agonism at the D2 receptor using PNU-95666E resulted in bradycardia, hindquarter vasodilation and decreases in arterial pressure. Partial agonism at the D3 receptor with BP897 had no effect on heart rate, but did produce depressor responses driven by decreases in HQVR. Conversely, agonism of the D4 receptor using PD168077 resulted in modest hindquarter vasoconstriction that was not dose dependent. Hence, by comparison, agonism of the D4 receptor has little effect in the cardiovascular system of the rat relative to the other dopamine receptor subtype agonists tested. PMID- 15659046 TI - Inhibition of inducible nitric oxide synthase reduces lipopolysaccharide-induced renal injury in the rat. AB - 1. Gram-negative bacterial lipopolysaccharide (LPS) release and subsequent septic shock is a major cause of death in intensive care units. Lipopolysaccharide has been reported to increase the production of nitric oxide (NO) and the formation of oxygen-derived free radicals (OFR) in different organs. The aim of the present study was to evaluate the role of an inducible form of NO synthase (iNOS) and OFR production in LPS-induced renal impairment. 2. Measurement of vitamin E as the most important fat-soluble anti-oxidant was used as a marker of tissue oxidative stress. Lipopolysaccharide (10 mg/kg), L-iminoethyl lysine (L-Nil; 3 mg/kg, i.p.; a specific inhibitor of iNOS activity) and dimethyl thiourea (DMTU; 500 mg/kg i.p.; a well-known OFR scavenger) were used. Four groups of eight rats were studied. One group received LPS, whereas a second group received LPS + L-Nil. A third group received LPS + DMTU and the fourth group, receiving saline, acted as a control group. To evaluate renal function, plasma creatinine and blood urea nitrogen (BUN) were measured. High-pressure liquid chromatography and ultraviolet detection were used to measure plasma and tissue vitamin E levels. Light microscopy was used to examine histopathological changes in the four groups. 3. Lipopolysaccharide markedly decreased the vitamin E content of renal plasma and tissue (P < 0.05). Administration of L-Nil attenuated renal dysfunction and preserved vitamin E levels. However, DMTU failed to prevent renal injury, as indicated by plasma BUN levels and renal histology, despite the fact that it maintained renal vitamin E levels and increased plasma vitamin E levels. Thus, the overproduction of NO by iNOS may have a role in this model of LPS-induced renal impairment. PMID- 15659047 TI - T102C genetic polymorphism of the 5-HT2A receptor in Chinese hypertensive patients and healthy controls. AB - 1. The 5-HT(2A) receptor belongs to the G-protein superfamily. It plays an important role in vascular regulation. 2. Previous reports in the UK have indicated that there is an association of the T102C genetic polymorphism of the 5 HT(2A) receptor with hypertension, but no studies have been made on the T102C genetic polymorphism in Chinese hypertensive patients. In the present study, we investigated the T102C genetic polymorphism of 5-HT(2A) receptors in Chinese hypertensive patients to determine whether there is an association of this polymorphism with hypertension in Chinese. 3. The present study was conducted on 198 hypertensive patients and 164 healthy controls. Polymerase chain reaction restriction fragment length polymorphism was used to identify the T102C genetic polymorphism of the 5-HT(2A) receptor. 4. In the present study, the C allele frequency of the 5-HT(2A) receptor genetic polymorphism was 0.343 in hypertensive patients, which was not significantly different to that in healthy controls (0.393; chi(2) = 1.922; P = 0.166; odds ratio = 0.807, 95% confidence interval 0.596-1.093). In addition, no gender differences were observed. 5. In conclusion, to our knowledge, this is the first report on the T102C genetic polymorphism of the 5-HT(2A) receptor in Chinese hypertensive patients. We find that no correlation exists between the T102C genetic polymorphism and hypertension, which affords useful information on the pathogenesis of hypertension in the Chinese population. PMID- 15659048 TI - Conditions affecting hydrodynamics-based gene delivery into mouse liver in vivo. AB - 1. It has been demonstrated that the hydrodynamics-based procedure has high efficiency to deliver foreign genes into the liver. The widespread use of this procedure in gene function studies and as a treatment option for liver and other organ diseases puts considerable importance on the investigation of various conditions that affect hydrodynamics-based gene delivery into mouse liver in vivo. 2. Various conditions, including the volume, speed and solution of the injection and the state, gender and strain of the animal were manipulated to evaluate their effect on the expression levels in mice of human factor IX (hFIX) 8 h after tail vein injection of the plasmid pCMV-hFIX. 3. It was found that an injection volume of 2-2.5 mL and an injection speed of 5-7 s were very effective in delivering DNA into the mouse liver. Using Ringer's solution as an injection fluid increased the efficiency of hFIX expression. 4. Anaesthetized mice expressed higher hFIX than conscious mice. Males expressed higher hFIX than females. The ICR mouse strain demonstrated higher expression of the foreign gene than did the C57 strain. 5. The effects of these specific factors on hFIX expression may be caused by variations in hydrostatic pressure, the degree of liver damage and liver size. 6. It can be concluded that there are optimal conditions for hFIX expression in the liver. This information may be helpful for the application of hydrodynamics-based procedures in the investigation of gene expression and gene therapy. PMID- 15659049 TI - Cardiac DT-diaphorase contributes to the detoxification system against doxorubicin-induced positive inotropic effects in guinea-pig isolated atria. AB - 1. Doxorubicin (DOX), a standard chemotherapeutic anthracycline agent, causes a positive inotropic effect in guinea-pig isolated atria in a concentration dependent manner with an ED(50) of 3.6 micromol/L. This increase in contractility is strictly related to the generation of reactive oxygen species (ROS) as a consequence of quinone metabolism. The ED(50) of DOX is significantly increased (P < 0.05) in the presence of 150 U superoxide dismutase (SOD). In the heart, DOX may be subjected to one- or two-electron reductions catalysed by flavoenzymes in the presence of suitable electron donors. Two-electron reduction is catalysed by NAD(P)H quinone acceptor oxidoreductase (DT-diaphorase; DTD). Whether DOX will be activated or detoxified by two-electron reduction is important for the understanding of the mechanism of both the toxic and antitumour actions of DOX. 2. In order to assess the role of DTD in cardiac responses to DOX, we examined the effect of both a specific inhibitor (dicoumarol) and an inducer (3 methylcholanthrene; MCA) of the enzyme on the inotropic action of DOX. 3. In guinea-pig isolated left atria, 4 micromol/L dicoumarol significantly enhanced the positive inotropic effect of DOX, especially at lower concentrations of DOX. In atria isolated from guinea-pigs treated with MCA (44 mg/kg, i.p. for 4 days), DTD activity was enhanced (approximately twice that of the control; P < 0.01), whereas the activity of glutathione S-transferase (GST) was not significantly altered. In these preparations, DOX caused a significantly lower increase in force of contraction than in atria isolated from untreated animals. 4. These results demonstrate that cardiac DTD does not contribute to ROS generation, but represents a detoxification system. PMID- 15659050 TI - Role of non-selective adenosine receptor blockade and phosphodiesterase inhibition in cisplatin-induced nephrogonadal toxicity in rats. AB - 1. It is well documented that cisplatin (CDDP) treatment increases the expression of adenosine A(1) receptors in both kidney and testes. However, the effect of adenosine at these receptors is controversial. Adenosine A(1) receptors have been documented to be involved in either cytoprotection or aggravation of nephrotoxicity. The aim of the present study was to examine the effect of the non selective adenosine receptor inhibitor theophylline and the phosphodiesterase inhibitor pentoxifylline on CDDP-induced renal and testicular toxicity. 2. Male Wister rats were divided into six groups. Two control groups received plain drinking water and a third control group received theophylline 0.8 mg/mL in the drinking water for 2 weeks. One group of animals drinking plain water was injected intraperitoneally (i.p.) with pentoxifylline 50 mg/kg per day for 2 weeks. The remaining groups were treated in the same manner and received single dose of CDDP 7 mg/kg, i.p., 1 week after starting theophylline and pentoxifylline treatment and all animals were killed 1 week after CDDP treatment. 3. Rats treated with CDDP developed nephrotoxicity, as demonstrated by increased kidney and testes weight as a percentage of total bodyweight, blood urea nitrogen and serum creatinine levels and decreased serum calcium and albumin levels. In addition, CDDP treatment resulted in an increase in the production of malondialdehyde (MDA) and decreases in total nitrate/nitrite levels, as well as depletion of reduced glutathione (GSH) content and glutathione peroxidase (GPX) activity in both the kidney and testes. Administration of theophylline in the drinking water to CDDP-treated rats resulted in exacerbation of the indices of nephrotoxicity, depletion of GSH content and GPX activity levels, with increased MDA production and platinum accumulation in both the kidney and testes. However, pentoxifylline administration reduced CDDP-induced biochemical changes and reduced platinum accumulation in both organs. Histopathological examination of the kidney revealed that CDDP treatment produced multifocal tubular atrophy, atypical reparative changes of the tubular epithelium and marked tubular necrosis. Animals treated with the theophylline/CDDP combination showed extensive widespread damage with intratubular calcification. However, pentoxifylline treatment ameliorated the overt changes induced by CDDP treatment. 4. Theophylline exacerbates the deleterious effects of CDDP on rat kidney and testes. However, pentoxifylline alleviates CDDP-induced renal and testicular toxicity. PMID- 15659051 TI - Anti-infarct effect of magnesium is not mediated by adenosine A1 receptors in rat globally ischaemic isolated hearts. AB - 1. The aim of present study was to investigate the effects of magnesium (Mg) on cardiac function and infarct size and to compare it effects with those of adenosine. The mechanism of Mg-mediated cardioprotection was explored by combined use of Mg and a selective adenosine A(1) receptor antagonist. 2. Rat isolated hearts were used for Langendorff perfusion. Hearts were either non-preconditioned or preconditioned with Mg (6 mmol/L) or adenosine (1 mmol/L) before 30 min sustained ischaemia followed by 120 min reperfusion. Within each of these protocols, hearts were divided into two groups; one group was exposed to the A(1) receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX; 200 nmol/L). Infarct size was measured by the triphenyltetrazolium chloride method. Left ventricular function was assessed by left ventricular developed pressure (LVDP), the product of heart rate x LVDP and coronary flow (CF). 3. The administration of Mg had an anti-infarct effect independent of its effect on postischaemic functional recovery in rats. Both Mg and adenosine equipotently reduced infarct size, but this effect of Mg was not blocked by the simultaneous administration of DPCPX. Cardiac function was improved by both adenosine and Mg and blockade of adenosine A(1) receptors attenuated these effects for both agents. 4. In conclusion, the results of the present study indicate that stimulation of adenosine A(1) receptors is not responsible for the anti-infarct effect of Mg in ischaemic myocardium in rats, but that the Mg-mediated protection of postischaemic functional recovery in rats is mediated by these receptors. PMID- 15659052 TI - Granulocyte colony stimulating factor/macrophage colony stimulating factor improves postinfarct ventricular function by suppression of border zone remodelling in rats. AB - 1. The aim of the present study was to examine the effects of mobilization of bone marrow cells by granulocyte colony stimulating factor (G-CSF) and macrophage colony stimulating factor (M-CSF) on ventricular function after myocardial infarction (MI). 2. After ligation of the left coronary artery, rats were divided into a vehicle control group (MI group) and a CSF-treated group (MI-CSF group). Rats in the MI-CSF group received a combination of G-CSF (50 microg/kg per day) and M-CSF (10(6) IU/kg per day) for 5 days after MI. Two weeks after MI, hearts were isolated and perfused with a Krebs' buffer and their functional responses to step-wise elevation of left ventricular end-diastolic pressure (LVEDP) were assessed. In histological analysis, proliferating cells and bone marrow-derived cells were identified by antibodies against Ki-67 and c-kit and organization of collagen was examined by picrosirius red staining. The mRNA levels of transforming growth factor (TGF)-beta(1), collagen type I and collagen type III were measured by quantitative reverse transcription-polymerase chain reaction. 3. Numbers of Ki-67- and c-kit-positive cells in the infarct border zone after MI were increased by CSF treatment, but few of those cells were stained by anti alpha-sarcomeric actin. The levels in mRNA of TGF-beta1 and collagen type I in the infarct border zone were higher in the CSF-treated group compared with the MI group. Although CSF treatment did not reduce ventricular hypertrophy or infarct size at 2 weeks after MI, it did significantly improved the response of left ventricular developed pressure to step-wise elevation of LVEDP. This effect was mimicked by treatment with M-CSF alone. The functional improvement by CSF treatment was correlated with suppression of enlargement of the infarct-non infarct border associated with infarct expansion. Collagen fibres in the border zone were thicker and orientated more orderly in the CSF-treated group than in the untreated group. 4. The results suggest that G-CSF/M-CSF treatment improves contractile function of the ventricle after infarction, presumably by acceleration of infarct repair and suppression of remodelling in the border zone. PMID- 15659053 TI - Functional imaging: gaining new insight from biophotonic imaging. PMID- 15659054 TI - Development of low-affinity, membrane-targeted Ca2+ sensors suitable for measuring presynaptic Ca2+. AB - 1. Our aim is to measure near-membrane Ca(2+) flux within the presynaptic terminals of central neurons by modifying new genetically encoded Ca(2+) sensors to develop tools capable of measuring localized Ca(2+) signals. 2. We used standard recombinant DNA technologies to generate the DNA coding for a fusion construct of a modified fluorescent 'pericam' Ca(2+) biosensor with a presynaptic P2X7 receptor (P2X7R). The Ca(2+) sensitivity of the biosensor was modified by rational site-directed mutagenesis of the calmodulin portion of the pericam. 3. Biosensor-receptor fusions were transfected into expression systems for evaluation. Expression studies in HEK-293 cells showed that biosensor-receptor fusion construct-delivered protein was localized exclusively to the plasma membrane, confirming that fusion did not affect the ability of the receptor to undergo normal protein synthesis and trafficking. 4. The Ca(2+)-dependent fluorescence of the pericam portion of the fusion protein was also retained. Site direct mutagenesis within the calmodulin moiety of the pericam significantly reduced the Ca(2+) affinity of the complex. The dynamic range of the sensor following this modification is better matched to the higher Ca(2+) levels expected within presynaptic Ca(2+) microdomains. PMID- 15659055 TI - Functional imaging: new views on lens structure and function. AB - 1. We have developed an experimental imaging approach that allows the distribution of lens membrane proteins to be mapped with subcellular resolution over large distances as a function of fibre cell differentiation. 2. Using this approach in the rat lens, we have localized precisely histological sites of connexin 46 cleavage, quantitatively mapped changes in gap junction distribution and fibre cell morphology and correlated these changes to differences in intercellular dye transfer. 3. Profiling of glucose transporter isoform expression showed that lens epithelial cells express GLUT1, whereas deeper cortical fibre cells express the higher-affinity GLUT3 isoform. Near the lens periphery, GLUT3 was located in the cytoplasm of fibre cells, but it underwent a differentiation-dependent membrane insertion. 4. Similarly, the putative adhesion protein membrane protein 20 is inserted into fibre cell membranes at the stage when the cells lose their nuclei. This redistribution is strikingly rapid in terms of fibre cell differentiation and correlates with a barrier to extracellular diffusion. 5. Our imaging-orientated approach has facilitated new insights into the relationships between fibre cell differentiation and lens function. Taken together, our results indicate that a number of strategies are used by the lens during the course of normal differentiation to change the subcellular distribution, gross spatial location and functional properties of key membrane transport proteins. PMID- 15659056 TI - Quantitative phase microscopy: a new tool for investigating the structure and function of unstained live cells. AB - 1. The optical transparency of unstained live cell specimens limits the extent to which information can be recovered from bright-field microscopic images because these specimens generally lack visible amplitude-modulating components. However, visualization of the phase modulation that occurs when light traverses these specimens can provide additional information. 2. Optical phase microscopy and derivatives of this technique, such as differential interference contrast (DIC) and Hoffman modulation contrast (HMC), have been used widely in the study of cellular materials. With these techniques, enhanced contrast is achieved, which is useful in viewing specimens, but does not allow quantitative information to be extracted from the phase content available in the images. 3. An innovative computational approach to phase microscopy, which provides mathematically derived information about specimen phase-modulating characteristics, has been described recently. Known as quantitative phase microscopy (QPM), this method derives quantitative phase measurements from images captured using a bright-field microscope without phase- or interference-contrast optics. 4. The phase map generated from the bright-field images by the QPM method can be used to emulate other contrast image modes (including DIC and HMC) for qualitative viewing. Quantitative phase microscopy achieves improved discrimination of cellular detail, which permits more rigorous image analysis procedures to be undertaken compared with conventional optical methods. 5. The phase map contains information about cell thickness and refractive index and can allow quantification of cellular morphology under experimental conditions. As an example, the proliferative properties of smooth muscle cells have been evaluated using QPM to track growth and confluency of cell cultures. Quantitative phase microscopy has also been used to investigate erythrocyte cell volume and morphology in different osmotic environments. 6. Quantitative phase microscopy is a valuable, new, non destructive, non-interventional experimental tool for structural and functional cellular investigations. PMID- 15659057 TI - Microscopic imaging of extended tissue volumes. AB - 1. Detailed information about three-dimensional structure is key to understanding biological function. 2. Confocal laser microscopy has made it possible to reconstruct three-dimensional organization with exquisite resolution at cellular and subcellular levels. 3. There have been few attempts to acquire large image volumes using the confocal laser scanning microscope. 4. Previously, we have used manual techniques to construct extended volumes (several mm in extent, at 1.5 microm voxel size) of myocardial tissue. 5. We are now developing equipment and efficient automated methods for acquiring extended morphometric databases using confocal laser scanning microscopy. PMID- 15659058 TI - Inherited cardiac arrhythmia syndromes: what have they taught us about arrhythmias and anti-arrhythmic therapy? AB - 1. In recent years, the identification of the gene defects in a vast array of monogenic disorders has revolutionized our understanding of the basic mechanisms underlying numerous disease processes. 2. Mutations in cardiac ion channels have been identified as the basis of a wide range of inherited arrhythmia syndromes, including the congenital long QT syndromes, Brugada syndrome, Lenegre syndrome, Andersen's disease and familial atrial fibrillation. 3. Identification of mutations in the human-ether-a-go-go-related gene (HERG) K(+) channel as the molecular basis of congenital long QT syndrome type 2 also led to the discovery that HERG is the molecular target for the vast majority of drugs (both cardiac and non-cardiac) that cause drug-induced arrhythmias. This has had profound implications not only for the development of anti-arrhythmic agents, but also for drug development in general. 4. The sequencing of the human genome in a sense represents the pinnacle of the reductionist era of molecular medicine. The great challenge now is to re-integrate the information gathered during the 'reductionist era' to provide a better understanding of the intact organism. Computer modelling is likely to be a key component of that re-integration process. PMID- 15659059 TI - Cardiac structure and electrical activation: models and measurement. AB - 1. Our group has developed finite element models of ventricular anatomy that incorporate detailed structural information. These have been used to study normal electrical activation and re-entrant arrhythmia. 2. A model based on the actual three-dimensional microstructure of a transmural left ventricular (LV) segment predicts that cleavage planes between muscle layers may give rise to non-uniform, anisotropic electrical propagation and also provide a substrate for bulk resetting of the myocardium during defibrillation. 3. The model predictions are consistent with the results of preliminary experiments in which a novel fibre optic probe is used to record transmembrane potentials at multiple intramural sites in the intact heart. Extracellular potentials are recorded at adjacent LV sites in these studies. 4. We conclude that structural discontinuities in ventricular myocardium may play a role in the initiation of re-entrant arrhythmia and discuss future studies that address this hypothesis. PMID- 15659060 TI - Distinct functions of DC-SIGN and its homologues L-SIGN (DC-SIGNR) and mSIGNR1 in pathogen recognition and immune regulation. AB - Antigen presenting cells express C-type lectins that are involved in pathogen capture, processing and antigen presentation to induce immune responses against these pathogens. However, it is becoming clear that pathogens have evolved to subvert the function of some C-type lectins to escape immune surveillance. An important C-type lectin family is represented by DC-SIGN and its homologues in human and mouse. Here we discuss the structure in relation to the pathogen binding specificity of the SIGN receptors and the function of these receptors in mouse and humans. PMID- 15659061 TI - Listeria monocytogenes-infected bone marrow myeloid cells promote bacterial invasion of the central nervous system. AB - Listeria monocytogenes is a facultative intracellular pathogen that is able to invade the central nervous system causing meningoencephalitis and brain abscesses. The mechanisms allowing bacteria to cross the blood-brain barrier are poorly understood. In this work, we used an experimental model of acute listeriosis in the mouse inducing a reproducible invasion of the central nervous system. At the early phase of infection, we find that bacteria invade and rapidly grow in bone marrow cells identified as bone marrow myelomonocytic cells expressing the phenotype CD31pos:Ly-6Cpos:CD11b(pos):LY-6Glow. We demonstrate that central nervous system invasion is facilitated by injecting L. monocytogenes infected bone marrow cells in comparison with free bacteria or infected spleen cells. In mice transplanted with bone marrow cells from transgenic donor mice expressing the green fluorescent protein (GFP), we show that infected myeloid GFP+ cells adhere to activated brain endothelial cells, accumulate in brain vessels and participate to the pathogenesis of meningoencephalitis and brain abscesses. Our results demonstrate that bone marrow, the main haematopoietic tissue, is a previously unrecognized reservoir of L. monocytogenes-infected myeloid cells, which can play a crucial role in the pathophysiology of meningoencephalitis by releasing infected cells into the circulation that ultimately invade the central nervous system. PMID- 15659062 TI - Overexpression and altered nucleocytoplasmic distribution of Anopheles ovalbumin like SRPN10 serpins in Plasmodium-infected midgut cells. AB - The design of effective, vector-based malaria transmission blocking strategies relies on a thorough understanding of the molecular and cellular interactions that occur during the parasite sporogonic cycle in the mosquito. During Plasmodium berghei invasion, transcription from the SRPN10 locus, encoding four serine protease inhibitors of the ovalbumin family, is strongly induced in the mosquito midgut. Herein we demonstrate that intense induction as well as redistribution of SRPN10 occurs specifically in the parasite-invaded midgut epithelial cells. Quantitative analysis establishes that in response to epithelial invasion, SRPN10 translocates from the nucleus to the cytoplasm and this is followed by strong SRPN10 overexpression. The invaded cells exhibit signs of apoptosis, suggesting a link between this type of intracellular serpin and epithelial damage. The SRPN10 gene products constitute a novel, robust and cell autonomous marker of midgut invasion by ookinetes. The SRPN10 dynamics at the subcellular level confirm and further elaborate the 'time bomb' model of P. berghei invasion in both Anopheles stephensi and Anopheles gambiae. In contrast, this syndrome of responses is not elicited by mutant P. berghei ookinetes lacking the major ookinete surface proteins, P28 and P25. Molecular markers with defined expression patterns, in combination with mutant parasite strains, will facilitate dissection of the molecular mechanisms underlying vector competence and development of effective transmission blocking strategies. PMID- 15659063 TI - The Helicobacter pylori VacA cytotoxin activates RBL-2H3 cells by inducing cytosolic calcium oscillations. AB - Helicobacter pylori causes an acute inflammatory response followed by chronic infection of the human gastric mucosa. Identification of the bacterial molecules endowed with a pro-inflammatory activity is essential to a molecular understanding of the pathogenesis of H. pylori associated diseases. The vacuolating cytotoxin A (VacA) induces mast cells to release pro-inflammatory cytokines. Here, we show that VacA activates the mast cell line RBL-2H3 by rapidly inducing an oscillation of the level of cytosolic calcium with exocytosis of secretory granules. Cytosolic calcium derives mainly from intracellular stores. VacA also stimulates a calcium-dependent production of pro-inflammatory cytokines, including tumour necrosis factor alpha (TNF-alpha). These observations indicate that VacA may act as a pro-inflammatory factor of H. pylori at very early stages of the innate immune response. PMID- 15659064 TI - A Plasmodium sporozoite protein with a membrane attack complex domain is required for breaching the liver sinusoidal cell layer prior to hepatocyte infection. AB - Plasmodium sporozoites are injected into the mammalian host during mosquito blood feeding and carried by the blood stream to the liver, where they infect hepatocytes and develop into erythrocyte-invasive forms. To reach the hepatocytes, sporozoites must cross the liver sinusoidal cell layer, which separates the hepatocytes from the circulatory system. Little is known about the molecular mechanisms by which sporozoites breach this cellular barrier. Here we report that a protein with a membrane attack complex/perforin (MACPF)-related domain is involved in this step. This molecule is specifically expressed in liver infective sporozoites and localized in micronemes, organelles engaged in host cell invasion. Gene disruption experiments revealed that this protein is essential for the membrane-wounding activity of the sporozoite and is involved in its traversal of the sinusoidal cell layer prior to hepatocyte-infection. Disruptants failed to leave the circulation, and most of them were eliminated from the blood by liver perfusion. Our results suggest that rupture of the host plasma membrane by the pore-forming activity of this molecule is essential for cell passage of the sporozoite. This report is the first to demonstrate an important role of a MACPF-related protein in host cell invasion by a pathogenic microorganism. PMID- 15659065 TI - Expression and activity of a Xenorhabdus nematophila haemolysin required for full virulence towards Manduca sexta insects. AB - As an insect pathogen, the gamma-proteobacterium Xenorhabdus nematophila likely possesses an arsenal of virulence factors, one of which is described in this work. We present evidence that the X . nematophilahaemolysin XhlA is required for full virulence towards Manduca sexta larvae. Lrp (leucine-responsive regulatory protein), FlhDC (regulator of flagella synthesis), and iron (II) limitation positively influenced xhlA transcript levels, suggesting XhlA expression is linked with nutrient acquisition and motility regulons. To help understand the role of XhlA in virulence, we examined its cellular targets and found that XhlA was a cell-surface associated haemolysin that lysed the two most prevalent types of insect immune cells (granulocytes and plasmatocytes) as well as rabbit and horse erythrocytes. Taken together, the need for xhlA for full virulence and XhlA activity towards insect immune cells suggest this haemolysin functions in X. nematophila immune evasion during infection. Analysis of a gene located immediately upstream of the xhlA locus, hcp (haemolysin co-regulated protein) revealed that its transcript levels were elevated during iron (III) limitation and its expression was Lrp-dependent. Further characterization of xhlA, hcp, and lrp will clarify their regulatory and functional relationships and their individual roles during the infectious process. PMID- 15659066 TI - EhCP112 is an Entamoeba histolytica secreted cysteine protease that may be involved in the parasite-virulence. AB - EhCP112 is an Entamoeba histolytica protease that together with the EhADH112 protein forms the EhCPADH complex involved in trophozoite virulence. Here, we produced the recombinant EhCP112 and studied its relationships with extracellular matrix components and with target cells. A DNA fragment containing the pro peptide and the mature enzyme was expressed in bacteria as an active enzyme (rEhCP112), whereas the full gene containing the signal peptide, the pro-peptide and the mature enzyme expressed a non-active protein. The fragment only with the mature enzyme was not expressed. rEhCP112 purified by affinity columns digested azocasein and had a strong autoproteolytic activity. Four hours after purification the protein appeared degraded. Anti-tag antibodies, monoclonal antibodies against the EhCP112 and sera from human patients with amoebiasis recognized rEhCP112. rEhCP112 digested gelatin, collagen type I, fibronectin and haemoglobin; it destroyed MDCK cell monolayers and bound to red blood cells. The native EhCP112 was poorly expressed in a virulence-deficient mutant, and in the wild-type clone it was located in secreted vesicles, forming the EhCPADH complex. Altogether these results show that EhCP112 is a molecule able to disrupt cell monolayers and digest proteins of the extracellular matrix and haemoglobin, and it is secreted by the trophozoites. PMID- 15659067 TI - The Mycobacterium tuberculosis SigD sigma factor controls the expression of ribosome-associated gene products in stationary phase and is required for full virulence. AB - During infection Mycobacterium tuberculosis is exposed to several environmental conditions depending on the stage and severity of the disease. To survive, M. tuberculosis uses alternate sigma factors to regulate its gene expression in response to the changing host environment. In order to better understand the way in which stress response genes are regulated, the extracytoplasmic sigma factor gene sigD was deleted and subsequently complemented in the CDC1551 strain of M. tuberculosis. The DeltasigD mutant strain exhibited an in vitro growth rate in rich medium identical to that of both the sigD-complemented and wild-type CDC1551 strains. Additionally, no differences were observed in short-term intracellular growth between the mutant, complemented, and wild-type bacteria within the J774A.1 macrophage cell line. However, tumour necrosis factor (TNF)-alpha levels in macrophages infected with the DeltasigD mutant were decreased as compared to levels observed in macrophages infected with the wild-type bacteria. In time-to death studies, C3H mice infected with the DeltasigD mutant exhibited a mortality delay compared to those infected with either the complemented or wild-type strains. Although mice infected with the DeltasigD mutant died at a reduced rate, the bacillary loads in the lungs and spleen of these mice were comparable to those seen in mice infected with either the complemented or wild-type strains. Microarray analysis of the DeltasigD mutant relative to wild type revealed that SigD directs the expression of a small set of ribosomal genes and adenosine triphosphate transporters whose expression is normally induced during stationary phase growth in vitro. Altered expression of a subset of these genes was confirmed by quantitative reverse transcription polymerase chain reaction analysis. Promoter-like elements resembling the consensus sequence AGAAAG-N16-20 CGTTAA were found upstream of 19 of the genes underexpressed in the DeltasigD mutant suggesting this may be the recognition sequence for the M. tuberculosis SigD-holoenzyme, EsigmaD. These data indicate that the M. tuberculosis SigD sigma factor governs the expression of a small set of ribosomal genes typically expressed in stationary phase during in vitro growth and that loss of sigD reduces macrophage TNF-alpha secretion as well as the lethality of M. tuberculosis infection in mice. PMID- 15659068 TI - A Trichomonas vaginalis 120 kDa protein with identity to hydrogenosome pyruvate:ferredoxin oxidoreductase is a surface adhesin induced by iron. AB - Trichomonas vaginalis, a human sexually transmitted protozoan, relies on adherence to the vaginal epithelium for colonization and maintenance of infection in the host. Thus, adherence molecules play a fundamental role in the trichomonal infection. Here, we show the identification and characterization of a 120 kDa surface glycoprotein (AP120) induced by iron, which participates in cytoadherence. AP120 is synthesized by the parasite when grown in 250 microM iron medium. Antibodies to AP120 and the electro-eluted AP120 inhibited parasite adherence in a concentration-dependent manner, demonstrating its participation in cytoadherence. In addition, a protein of 130 kDa was detected on the surface of HeLa cells as the putative receptor for AP120. By peptide matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS), the AP120 adhesin showed homology with a hydrogenosomal enzyme, the pyruvate:ferredoxin oxidoreductase (PFO) encoded by the pfoa gene. This homology was confirmed by immunoblot and indirect immunofluorescence assays with an antibody to the carboxy terminus region of the Entamoeba histolytica PFO. Reverse transcription polymerase chain reaction (RT-PCR) assays showed that a pfoa-like gene was better transcribed in trichomonads grown in iron-rich medium. In conclusion, the homology of AP120 to PFO suggests that this novel adhesin induced by iron could be an example of moonlighting protein in T. vaginalis. PMID- 15659069 TI - Constitutive association of Mcm2-3-5 proteins with chromatin in Entamoeba histolytica. AB - Eukaryotic cells duplicate their genome once and only once per cell cycle. Our earlier studies with the protozoan parasite, Entamoeba histolytica, have shown that genome reduplication may occur several times without nuclear or cellular division. The Mcm2-7 protein complex is required for licensing of DNA replication. In an effort to understand whether genome reduplication occurs due to absence or failure of the DNA replication licensing system, we analysed the function of Mcm2-3-5 proteins in E. histolytica. In this study, we have cloned E. histolytica (Eh) MCM2 and Eh MCM5 genes, while Eh MCM3 was cloned earlier. The sequence of Eh MCM2-3-5 genes is well conserved with other eukaryotic homologues. We have shown that Eh Mcm2,3 proteins are functional in Saccharomyces cerevisiae. Our studies in E. histolytica showed that Eh Mcm2-3-5 proteins are associated with chromatin constitutively in cycling cells and during arrest of DNA synthesis induced by serum starvation. Alternation of genome duplication with mitosis is regulated by association-dissociation of Mcm2-7 proteins with chromatin in other eukaryotes. Our results suggest that constitutive association of Mcm proteins with chromatin could be one of the reasons why genome reduplication occurs in E. histolytica. PMID- 15659070 TI - Colonic short-chain fatty acids inhibit encystation of Entamoeba invadens. AB - Entamoeba parasites multiply as trophozoites in the layer of mucus that overlies the colonic epithelium. In response to stimuli that are not understood, trophozoites stop multiplying and differentiate into cysts that are released to infect another host. In the colon, Entamoeba trophozoites are exposed to the large variety of biochemicals that are carried into or are produced within this organ. The normal bacterial population of the colon releases large amounts of short-chain fatty acids (SCFAs). These compounds have effects on the growth, differentiation and repair of the colonic epithelium that correlate with de creased activity of a Class I/II histone deacetylase (HDAC). We found that the formation of cysts, but not the growth of trophozoite-stage Entamoeba invadens parasites, was inhibited by physiologic concentrations of SCFAs. Variable levels of cyst formation did occur if SCFA concentrations were lowered. Specific inhibitors of Class I/II-type HDACs also prevented encystation, and trophozoites exposed to these compounds had increased levels of acetylation of histone H4 and other nuclear proteins. These results suggest that production of the infectious cyst stage of Entamoeba parasites is regulated in part by the levels of SCFAs made by the bacterial population of the colon. PMID- 15659071 TI - Differentiation of human monocytic cell lines confers susceptibility to Bacillus anthracis lethal toxin. AB - Anthrax lethal toxin (LT) is comprised of protective antigen and lethal factor. Lethal factor enters mammalian cells in a protective antigen-dependent process and cleaves mitogen-activated protein kinase kinases. Although LT has no observable effect on many cell types, it causes necrosis in macrophages derived from certain mouse strains and apoptosis in activated mouse macrophages. In this study, we observed that LT treatment of three different human monocytic cell lines U-937, HL-60 and THP-1 did not induce cell death. Cells did become susceptible to the toxin, however, after differentiation into a macrophage-like state. Treatment with LT resulted in decreased phosphorylation of p38, ERK1/2 and JNK in both undifferentiated and differentiated HL-60 cells, suggesting that the change in susceptibility does not result from differences in toxin delivery or substrate cleavage. Death of differentiated HL-60 cells was accompanied by chromosome condensation and DNA fragmentation, but was not inhibited by the pan caspase inhibitor Z-VAD-FMK. In addition, we observed that the macrophage differentiation process could be inhibited by LT. Our results indicate that LT mediated death of mouse and human macrophages may occur through distinct processes and that the differentiation state of human cells can determine susceptibility or resistance to LT. PMID- 15659072 TI - Molecular and cellular profiles of insect bacteriocytes: mutualism and harm at the initial evolutionary step of symbiogenesis. AB - Intracellular symbiosis is considered to be a driving force in eukaryotic cell evolution. In insects, little is known about the molecular bases of the bacteria bearing host cells (bacteriocytes), particularly in the initial steps of symbiosis, where the bacterial genome has not experienced severe gene deletions because of evolutionary constraints associated with intracellular and vertical transmission. Here, we have applied polymerase chain reaction (PCR)-subtracted cDNA and reverse Northern analysis on the bacteriocytes of a recently established endosymbiosis, the weevil Sitophilus zeamais, to discover genes of potential relevance to bacteriocyte genetics. We provide a broad characterization of bacteriocyte transcriptional responses to intracellular bacteria, including pathways covering metabolism-transport-stress (MTS), cell signalling and trafficking, growth and apoptosis, as well as innate immunity. MTS genes show an intriguing diabetes-like pathogenic profile associated with increased stress, as indicated by high levels of upregulations of carbohydrate transporters, aldose reductases and stress-related genes. A high-performance liquid chromatography (HPLC) analysis of tissue carbohydrate contents highlighted an increased carbohydrate assimilation in symbiotic insects and the prevalence of a polyol biosynthetic pathway, as indicated by the accumulation of sorbitol, mannitol and fructose in the bacteriocytes. These findings provide the first genetic perspectives on the nature of the interaction between insect and cooperative bacteria. They unravel the profound insect bacteriocyte stress associated with increased metabolism and cell trafficking, and they shed light on the potential role of the innate immunity during the pathogeny-mutualism transition at the initial stage of insect symbiogenesis. PMID- 15659073 TI - Posturographic description of the regaining of postural stability following stroke. AB - BACKGROUND: Failing balance and increased liability to falling are common complaints among hemiplegic subjects. During rehabilitation much effort is put into regaining postural stability. PURPOSE: To describe the regaining of postural stability during rehabilitation for the first year following stroke in hemiplegic patients MATERIALS AND METHODS: Twenty-six patients were included within 5 days of suffering a stroke resulting from a CT-verified clearly defined thromboembolic lesion, localized in the vascular bed of middle cerebral artery on either side. Posturographic evaluation of sway was performed on a commercially available force plate system (Balance Master Pro). Measured parameters included per cent maximum stability (PMS), per cent ankle strategy (AST), and average angular velocity (AVE). An additional parameter was derived by calculating the slope (SLP) of the linear relationship between stability and ankle strategy. SLP reflects the predisposition of the stroke patient to keep using ankle strategy when faced with increasingly difficult balance tasks. The presence of familiarization to the measurement method was examined by including a preliminary measurement not otherwise included in the analysis of changes during the first year of follow-up. The temporal evolvement of sway was described by measuring sway parameters 1, 2, 4, 8 and 52 weeks following stroke. The proportions of the variation in sway caused by measurement error and between patients were compared. RESULTS: All parameters improved over time, as demonstrated by significant improvements from each of week 1, 2, 4 and 8 to week 52. There was evidence of familiarisation for PMS, AST and AVE, but not for SLP. Estimation of variance components showed that between patient variation accounts for between two-thirds and four-fifths of the total variation. A considerable part of this variation was caused by individual differences in the temporal improvement of sway over time. Patients considered the evaluation of postural sway demanding and time consuming resulting in a relatively large loss to follow-up: 15 patients completed the 8-week visit while seven completed the 1 year visit. CONCLUSION: Postural stability increases for the first year following stroke. There is evidence of familiarization towards the measurements process for the traditional posturographic parameters PMS, AST and AVE, but not for the derived parameter SLP, which might make the latter a candidate for a sway parameter reflecting basic mechanisms of upholding upright stance in hemiplegic patients. The estimated variance components stress the considerable between-patient variation and question the ability of the force plate method to monitor individual sway performance of such patients during the rehabilitation process. PMID- 15659074 TI - Hyperoxia-induced arterial compliance decrease in healthy man. AB - INTRODUCTION: Oxygen therapy is commonly used in emergency department and intensive care units without careful evaluation of its effects, especially on the haemodynamics and artery characteristics. MATERIALS AND METHODS: A prospective laboratory study evaluated brachial circulatory effects of normobaric hyperoxia using ultrasonography-Doppler. The study was set in a hospital research laboratory. The subjects were thirteen healthy volunteers. Investigations were performed under normal air ventilation and after 20 min of hyperoxic mixture ventilation using a high concentration mask. Two dimensional images and brachial blood flow velocities were recorded using ultrasonography and pulsed Doppler to study changes in cross sectional area, blood flow, resistance index, and cross sectional compliance coefficient. RESULTS: During hyperoxic exposure, mean PaO2 was 372 +/- 21 mmHg. A significant decrease of heart rate was observed. Arterial pressures (systolic and diastolic arterial pressures) were not modified. A decrease of cross sectional areas at end diastole and end systole was observed. Pulsed Doppler study showed a decrease of brachial artery blood flow and an increase of the resistance index. Furthermore, a decrease of the cross-sectional compliance coefficient was observed during hyperoxic exposure in all subjects. CONCLUSION: This study using two-dimensional ultrasonography and pulsed Doppler could demonstrate an increase in brachial arterial tone and a decrease in brachial blood flow under normobaric hyperoxia. PMID- 15659075 TI - Dizziness is associated with decreased vasoreactivity in right cerebral hemisphere for head-down manoeuvre--near-infrared spectroscopy study. AB - To investigate the vasoreactivity of cerebral hemisphere in patients with dizziness and syncope, we compared changes in total haemoglobin (THbl) and regional oxygen saturation (rSO2) of the right and left frontal lobes in response to head-down manoeuvre. Ninety-six right-handed subjects (aged 59 +/- 19 years) were asked to perform a head-down or a standing manoeuvre. Head-down manoeuvre produced a greater increase in right side THbl in subjects under 70 years of age (8.5 +/- 3.1) when compared with subjects older than 70 years (0.40 +/- 0.08). In contrast, the head-down manoeuvre had no effects on left side THbl, irrespective of patient age. Similarly, the head-down manoeuvre resulted in a greater decrease of right side rSO2 in subjects under 70 years of age (-5.2 +/- 2.1%) when compared with subjects older than 70 years (0.31 +/- 0.9%). In contrast, the head down manoeuvre had no effects on left side rSO2, irrespective of patient age. The head-down manoeuvre produced a smaller increase in right side THbl in subjects with dizziness (0.38 +/- 0.19) than in those without dizziness (9.4 +/- 3.5). A standing manoeuvre produced a smaller increase in right side THbl in subjects with syncope (-0.057 +/- 0.047) than in those without syncope (0.063 +/- 0.028). The head-down manoeuvre produced a decrease in right side rSO2 in subjects without dizziness (-6.4 +/- 2.4%) and a slight increase in right side rSO2 in subjects with dizziness (1.1 +/- 0.4%). Subjects with dizziness (67 +/- 2.1 years) were significantly older than those without dizziness (53 +/- 2.7 years) or those with syncope (44 +/- 4.2 years). These data indicate that reduced vasoreactivity to right hemispheric pressure changes is associated with dizziness in older subjects. Further, decreases in right hemispheric THbl during a standing manoeuvre are associated with syncope in relatively younger subjects. PMID- 15659076 TI - Bone infection in patients suspected of complicating osteomyelitis: the diagnostic value of dual isotope bone-granulocyte scintigraphy. AB - AIM: The purpose of this study was to evaluate the diagnostic value of dual isotope bone-granulocyte scintigraphy in patients with known bone pathology clinically suspected of osteomyelitis, i.e. complicating osteomyelitis, using per operative bacterial culture from bone as reference. METHODS: Simultaneous dual isotope bone-granulocyte scintigraphic images were obtained in 42 consecutive patients in whom conventional X-ray, erythrocyte sedimentation rate, and C reactive protein were also available. 99mTc MDP bone and 111In labelled granulocyte imaging was obtained simultaneously. The images were interpreted as positive for osteomyelitis if regions of interests of pathologic 111In granulocyte accumulation included 99mTc MDP activity on the bone images (except in the spine). RESULTS: The sensitivity, specificity, and accuracy were 84, 71 and 79%, respectively, for simultaneous, dual isotope bone-granulocyte scintigraphy, higher than the other diagnostic parameters. CONCLUSION: Simultaneous bone-granulocyte scintigraphy is a valuable diagnostic tool in diagnosing osteomyelitis complicating other bone pathology with or without soft tissue infection. PMID- 15659077 TI - Lower body positive pressure by anti-G garment inflation: a suitable method to increase pulmonary capillary wedge pressure in healthy elderly subjects. AB - In a study on non-invasive assessment of pulmonary capillary wedge pressure (PCWP), we sought a method to increase PCWP non-invasively. We hypothesized that inflation of an anti-G garment was suitable to increase PCWP non-invasively in healthy elderly subjects. In 20 subjects, aged 70 +/- 4 years (mean +/- SD), before, immediately after, and 4 min after anti-G garment inflation to 52 mmHg, PCWP and mean pulmonary artery pressure (MPAP) were measured with a Swan-Ganz catheter, and mean arterial blood pressure (MAP) with Finapres, in supine and semi-recumbent position. Supine, PCWP (mmHg, mean +/- SD) increased from 9.9 +/- 2.1 to 15.5 +/- 3.9** immediately after inflation and 13.4 +/- 3.7** at 4 min; semi-recumbent from 8.9 +/- 2.0 to 17.5 +/- 3.3** and 14.7 +/- 2.9** (*P<0.05, **P< 0.001 versus before inflation). MPAP (mmHg) increased after inflation: supine 16.9 +/- 2.3 to 22.3 +/- 4.6** and 20.6 +/- 3.9** and semi-recumbent 15.7 +/- 2.8 to 24.3 +/- 5.1** and 22.5 +/- 3.5**, suggesting that increased preload was the primary effect of anti-G garment inflation. Supine MAP (mmHg) increased from 96.0 +/- 11.3 to 101.4 +/- 13.4** and 100.5 +/- 12.7* and semi-recumbent from 102.0 +/- 8.9 to 108.3 +/- 11.4** and 106.0 +/- 11.3*, suggesting an effect of increased afterload as well. The latter was supported by an increase in total peripheral resistance (d s cm(-5)) from 1346 +/- 299 to 1441 +/- 384 after 4 min (P = 0.057) and from 1461 +/- 341 to 1532 +/- 406 (P = 0.054), supine and semi recumbent respectively, while cardiac output remained unchanged. Complications did not occur. We conclude that in healthy elderly subjects, anti-G garment inflation is a safe, non-invasive, method to induce a significant increase in PCWP. Our findings justify its application in future studies in which non invasive temporary increase in PCWP is required. PMID- 15659078 TI - Iodine 123-metaiodobenzylguanidine imaging reflect generalized sympathetic activation in patients with left ventricular dysfunction. AB - BACKGROUND: Iodine 123-metaiodobenzylguanidine (MIBG) imaging has been used to assess cardiac sympathetic nerve abnormalities. To determine the role of MIBG imaging as a measure of generalized sympathetic nerve activity, MIBG imaging was evaluated with muscle sympathetic nerve activity (MSNA) and plasma norepinephrine (noradrenaline) level in patients with old myocardial infarction. METHODS: Myocardial MIBG scintigraphy, MSNA and plasma norepinephrine measurement were performed within 3 days in 35 patients with old myocardial infarction. Patients were divided into three groups according to their ejection fraction (EF); preserved (EF > or = 50%, 12 patients), intermediate (35% < EF < 50%, 13 patients), or depressed (EF < or = 35%, 10 patients). The heart to mediastinum (H/M) ratio was obtained 4 h after tracer injection from the chest anterior view image. MIBG washout rate was calculated from the early and delayed images. MSNA was recorded by microneurography. RESULTS: Plasma norepinephrine level had weak negative correlations with early H/M ratio (r = 0.37, P<0.05) and delayed H/M ratio (r = 0.33, P<0.05), and a positive correlation with MIBG washout rate (r = 0.54, P<0.01). MSNA had weak negative correlations with early H/M ratio (r = 0.51, P<0.05) and delayed H/M ratio (r = 0.52, P<0.05). However, a strong correlation was found between MSNA and MIBG washout rate (r = 0.88, P<0.001). Despite no significant differences in plasma norepinephrine level and H/M ratio, patients with intermediate and depressed EF had higher MIBG washout rate and MSNA compared with those with preserved EF. CONCLUSIONS: Increased in cardiac sympathetic nerve activity was associated with augmented sympathetic outflow of the skeletal muscle and hence, MIBG washout rate allow the assessment of general sympathetic nerve activity. PMID- 15659079 TI - Forearm vascular reactivity is differentially influenced by gliclazide and glibenclamide in chronically treated type 2 diabetic patients. AB - Sulphonylureas (SUs) act by inhibition of beta-cell K(ATP) channels after binding to the sulphonylurea receptor SUR1. K(ATP) channels are also expressed in cardiac and vascular myocytes coupled to SUR2A and SUR2B involved into adaptations of vascular tone and myocardial contractility. Different influence of SUs on vascular function is based on different binding to the SUR family. Few data on the effect of different SUs, used in patients in therapeutic doses, on vascular function are currently available. We investigated possible effects of acute and chronic treatment with glibenclamide and gliclazide on forearm postischaemic reactive hyperaemia (RH) in type 2 diabetic patients. To that purpose a double blind, randomized, cross-over study with gliclazide (80 mg, b.i.d.) and glibenclamide (5 mg, b.i.d.) was performed in 15 type 2 diabetic patients. Forearm vascular reactivity was measured after 5 min of ischaemia by plethysmography before and after 4 weeks treatment. After acute administration of gliclazide (80 mg) or glibenclamide (5 mg) RH was not influenced. After 4 weeks of treatment, no influence of either drug was seen in the steady state before dosing. After dosing glibenclamide induced a significant (P = 0.004) reduction of RH from 26.4 +/- 6.9 to 21.9 +/- 7.6 ml min(-1)/100 ml after 4 h. Gliclazide, conversely, did not induce a reduction of RH (23.9 +/- 6.0 to 23.3 +/- 6.6 ml min(-1)/100 ml). No influence of HbA1c or actual glycaemia on RH was observed. Our results indicate that in chronically treated patients with type 2 diabetes ingestion of glibenclamide but not gliclazide results in sustained reduction of postischaemic RH. This difference is most probably based on different SUR binding. PMID- 15659080 TI - Comparison of stroke volume estimation for non-steady-state and steady-state graded exercise testing. AB - The stroke volume (SV) during exercise is an important index of the heart's functional capacity. A new method has been developed for the non-invasive estimation of exercise SV (SVex). It requires the determination of the slope for the oxygen uptake versus heart rate relationship in the steady state of graded exercise testing (GXT). The product of the slope and a constant (reciprocal of an assumed value of the arterial oxygen content) equals an estimated value for SVex. It was validated in a previous study using invasive measurements while subjects were performing steady-state GXT. However, currently the most commonly used GXT protocols are non-steady state, e.g. protocols with 1-min increment durations. We tested the hypothesis that SVex is the same for steady-state and non-steady-state GXT. A total of 30 subjects (15 males and 15 females) served as subjects for the study. Each subject performed two GXTs on different days with different increment durations - 1 and 4 min. Ventilation and gas exchange were measured with the Vacumed metabolic cart. For the male subjects, the mean (SD) SVex values for the 1- and 4-min GXTs were 155.4 (39.5) and 134.6 (27.5) ml, respectively. The corresponding values for the female subjects were 151.6 (37.6) and 134.3 (36.4) ml. Paired t-test analysis demonstrated that for both genders the mean SVex for the 1-min GXT was significantly larger than the 4-min GXT mean value (P<0.05). Hence, the commonly used 1-min GXT does not yield the same values for SVex as the steady-state GXT. PMID- 15659081 TI - Heart rate variability during sedentary work and sleep in normal and sleep deprived states. AB - The possibility of using heart rate variability (HRV) as an indicator of sleepiness was investigated by analysing heart rate (HR) activity and electroencephalography (EEG) recordings from 10 individuals who performed a monotonous attention task for 120 min in both sleep-deprived and rested states. In both conditions, measurements were collected during 60 min of sleep immediately following a 120 min of non-sleep (awake phase). Although HR decreased significantly in both the rested and the sleep-deprived states during the awake phase, HR significantly changed sooner when subjects were sleep-deprived than when they were rested. No significant changes in HRV were found during the awake phase; however, HRV correlated significantly with alpha and theta power densities when rested but not when sleep-deprived. During the sleep phase, the total HRV and very low and low frequency HRV components significantly decreased approximately 40 min after sleeping in the sleep-deprived condition. These HRV components were also significantly and negatively correlated with delta power densities. HRV does not seem to be a viable indicator of sleepiness; however, HRV may be useful for determining sleep stages. PMID- 15659082 TI - Limitations of non-invasive endothelial function assessment by brachial artery flow-mediated dilatation. AB - A non-invasive technique using high-frequency ultrasound brachial artery imaging to assess endothelium-dependent flow-mediated vasodilatation is a widely used test, but interpretation of results is not consistent. This study was designed to assess the method of non-invasive endothelial function determination of a brachial artery. Endothelial function was assessed by two physicians in 18 young, healthy volunteers. Each volunteer was examined by both physicians on the same day using an identical protocol; a second assessment was carried out at an interval of 6-7 days. When comparing arterial dilatation at first and second measurements by one physician, there were no statistically significant differences (first physician: 5.95 +/- 2.93% versus 7.63 +/- 4.3%; P = 0.21; second physician: 4.23 +/- 1.6% versus 4.94 +/- 2.69%; P = 0.22). Further, we found statistically significant differences in artery dilatation when comparing measurements made separately by both physicians on the same day (5.95 +/- 2.93% versus 4.23 +/- 1.6%; P = 0.03, and 7.63 +/- 4.3% versus 4.94 +/- 2.69%; P = 0.003). Our results suggest a large inter-individual variability of measurements within the whole group, if made on the same day and at the same time by two physicians. On the contrary, no significant differences were noted when comparing measurements of the whole group by the one physician at an interval of 1 week. It can be concluded that the degree of brachial artery flow-mediated dilatation is difficult to evaluate on the basis of a predefined cut-off point as a single measurement screening test. PMID- 15659083 TI - Assessment of lung volumes in children and adolescents: comparison of two plethysmographic techniques. AB - Thoracic gas volume (Vtg) can be measured with body plethysmography by either repetitive panting or one single inspiratory effort against a shutter occluding the airways. The panting technique is preferred, but may be demanding. We aimed to assess the precision of these two methods and the degree of agreement between them. Vtg and functional residual capacity (FRC) were measured in 155 subjects with a standard, commercially available plethysmograph, acting as a variable pressure, constant-volume device when Vtg is determined. Total lung capacity (TLC) and residual lung volume (RV) were calculated subsequent to a full vital capacity manoeuvre. For non-asthmatic healthy subjects, the standard deviations (SD) of the differences between two replicate measurements of FRC, TLC and RV were respectively 0.16, 0.13 and 0.14 litres with the panting technique, and 0.18, 0.18 and 0.23 litres with the single inspiratory effort technique. In percentage of the respective lung volumes, the corresponding 1.96 SDs were 20%, 8% and 40% with the panting technique and 23%, 12% and 67% with the single inspiratory effort technique. Between the two techniques, 95% limits of agreement were 21% for FRC, 11% for TLC and 58% for RV. The variability of Vtg and FRC accounted for most of the variability of TLC and RV. In conclusion, the panting and the single inspiratory effort technique produced results that were comparable in magnitude, however with a better precision with the panting technique. The single inspiratory effort technique can be used as an alternative if the panting technique fails. PMID- 15659085 TI - Towards an understanding of nursing as a response to human vulnerability. AB - It is not unusual for the adjective 'vulnerable' to be applied to those in receipt of nursing practice without making clear what it is that persons thus described are actually vulnerable to. In this paper I argue that the way nursing has adopted the idea of vulnerability tends to imply that some people are in some way invulnerable. This is conceptually unsustainable and renders the idea of the vulnerable patient (almost) meaningless. The paper explores the meaning of vulnerability both in general terms and in the context of nursing practice. It is argued that to be in receipt of nursing is to become, to a greater or lesser extent, more-than-ordinary vulnerable. Thus all patients are more-than-ordinarily vulnerable and this restricts their potential to flourish. Nurses are well placed to contribute to the flourishing of more-than-ordinarily vulnerable persons and my substantive claim is that this 'protective' function is indeed a legitimate and fundamental part of the role of nurses. PMID- 15659086 TI - Two models of mistake-making in professional practice: moving out of the closet. AB - Nurses make mistakes in practice despite the culturally based expectation of perfection. Such a disparity between reality and expectation calls members of the profession to question the current attitudes toward mistakes in practice. Two explanatory models of the origin of mistakes are presented. The Perfectibility Model holds that any error or harm is caused by an individual practitioner's lack of knowledge or motivation. The Faulty Systems Model offers a broader explanation of human error. I conclude that a Faulty Systems Model is more comprehensive and more effective for managing mistakes. Integrating the Faulty Systems Model into practice and education can result in more ethically fitting responses to errors and ultimately better outcomes for nurses, institutions and patients. PMID- 15659087 TI - Understanding end-of-life caring practices in the emergency department: developing Merleau-Ponty's notions of intentional arc and maximum grip through praxis and phronesis. AB - The emergency department (ED) is a fast-paced, highly stressful environment where clinicians function with little or suboptimal information and where time is measured in minutes and hours. In addition, death and dying are phenomena that are often experienced in the ED. Current end-of-life care models, based on chronic illness trajectories, may be difficult to apply in the ED. A philosophical approach examining end-of-life care may help us understand how core medical and nursing values are embodied as care practices and as ethical comportment. The integration of Aristotle's notions of phronesis and praxis with Merleau-Ponty's ontological notions of intentional arc and maximum grip in the context of the culture and practices of the ED offers a unique view of clinical and ethical practice at the end-of-life in the emergency setting. Caring for people at the end-of-life calls us to act virtuously based on previous experience, meanings and local practices. The maximum grip of the ultimate particulars of the situation combined with one's experiential and theoretical knowledge opens up situated possibilities for the expert clinician. PMID- 15659088 TI - The production of the psychiatric subject: power, knowledge and Michel Foucault. AB - The issue of power has become increasingly important within psychiatry, psychotherapy and mental health nursing generally. This paper will suggest that the work of Michel Foucault, the French philosopher and historian, has much to contribute to the discussion about the nature, existence and exercise of power within contemporary mental health care. As well as examining his original and challenging account of power, Foucault's emphasis on the intimate relationship between power and knowledge will be explored within the context of psychiatry and mental health nursing. This is to say that the paper will investigate Foucault's account of how power and knowledge are central to the process by which human beings are 'made subjects' and therefore how 'psychiatric identities' are produced. In doing so, it will be suggested that Foucault's work can not only make a valuable contribution to contemporary discussions about power and knowledge, but can also provide a significant critique and reconceptualization of the theoretical foundations and associated diagnostic and therapeutic practices of psychiatry and mental health nursing. PMID- 15659089 TI - Some remarks on the relevance of basic research in nursing inquiry. AB - The aim of this article was to illuminate the issue of basic research in nursing and to problematize its relevance for our discipline. First, we asked leading nursing scholars in the Nordic countries to share their views on basic research in nursing. Thereafter, the ideas, views and suggestions of the scholars were amalgamated with insights from the literature and from the discussions in our project team. Our two guiding questions were: What role can basic research be assigned? Which, if any, forms of basic research can be identified? We found that basic nursing research may be seen as a necessary basis for applied research, as a contribution to applied research, or that the whole issue is seen as a pseudo issue. We further found that basic nursing research can be seen as either contextual or general as well as either intradisciplinary or multidisciplinary in form. We conclude by the following three succinct remarks or assertions: that basic nursing research is not to be equated with biomedical conceptions of basic research; that basic nursing research may take place on several theoretical levels; and finally, that an indiscriminate dismissal of basic research in nursing may affect the cognitive autonomy of our discipline. PMID- 15659090 TI - Knowing the nurse practitioner: dominant discourses shaping our horizons. AB - The purpose of this paper is to examine the various discourses, particularly the dominant instrumental and economic discourses that have brought the phenomena of the nurse practitioner (NP) into being. It is proposed that NPs have been constituted as an object of nature and therefore understood metaphorically as a tool or instrument within the health care system to be used efficiently and effectively. Heidegger's philosophical analysis of the question concerning technology is used to argue that our current ways of knowing the NP through these discourses, with their emphasis on calculative logic, have resulted from our modern view of the essence of technology. It is also argued that there is now a need to shape our horizons concerning the NP in new and different ways. There is a need to engage in dialogical forms of research in order to evoke the richness and depth of what it means to be an NP, that is, to reveal the other modes of expression by which we define ourselves, understand others and nursing. PMID- 15659091 TI - Through the lens of Merleau-Ponty: advancing the phenomenological approach to nursing research. AB - Phenomenology has proved to be a popular methodology for nursing research. I argue, however, that phenomenological nursing research could be strengthened by greater attention to its philosophical underpinnings. Many research reports devote more page space to procedure than to the philosophy that purportedly guided it. The philosophy of Maurice Merleau-Ponty is an excellent fit for nursing, although his work has received less attention than that of Husserl and Heidegger. In this paper, I examine the life and thought of Merleau-Ponty, with emphasis on concepts, such as perception, intentionality and embodiment, which have particular relevance to the discipline of nursing. PMID- 15659092 TI - Personal identity. PMID- 15659093 TI - Cell-specific regulation of a Brassica napus CMS-associated gene by a nuclear restorer with related effects on a floral homeotic gene promoter. AB - Cytoplasmic male sterility (CMS) is a maternally inherited defect in pollen production specified by novel mitochondrial genes. It can be suppressed by nuclear restorer (Rf) genes which normally downregulate expression of a CMS associated novel mitochondrial gene. Two forms of Brassica napus CMS, nap and pol, are associated with related chimeric genes, orf222 and orf224, respectively. We show that in pol and nap CMS, anther locule development is asynchronous and asymmetric, that one or more locules within each anther may fail to develop entirely and that CMS anthers display polarity in locule development. We show, by in situ hybridization, that orf222 transcripts accumulate in sterile anthers prior to development of morphological differences between CMS and restored stamens, and remain preferentially localized to microsporangia. In fertility restored anthers, however, orf222 transcript levels remain low throughout development. Some sporogenous and meiotic cells differentiate within CMS anthers and form functional pollen despite retaining high orf222 transcript levels, suggesting that the effect of orf222 expression in blocking pollen development is limited to an early and specific stage. Transcripts of other mitochondrial genes, exemplified by atp6 and cob, and of the nuclear-encoded ATP synthase gamma subunit, accumulate preferentially in the microsporangia of both sterile and fertile anthers. Thus nuclear fertility restoration reduces orf222 transcript levels in a gene and tissue-specific manner. We observe differences between CMS and fertile plants in the timing and patterning of APETALA3 promoter activity that suggest a possible basis for the developmental abnormalities of CMS flowers. PMID- 15659094 TI - Histone deacetylation is required for progression through mitosis in tobacco cells. AB - Post-translational modifications of core histone proteins play a key role in chromatin structure and function. Here, we study histone post-translational modifications during reentry of protoplasts derived from tobacco mesophyll cells into the cell cycle and evaluate their significance for progression through mitosis. Methylation of histone H3 at lysine residues 4 and 9 persisted in chromosomes during all phases of the cell cycle. However, acetylation of H4 and H3 was dramatically reduced during mitosis in a stage-specific manner; while deacetylation of histone H4 commenced at prophase and persisted up to telophase, histone H3 remained acetylated up to metaphase but was deacetylated at anaphase and telophase. Phosphorylation of histone H3 at serine 10 was initiated at prophase, concomitantly with deacetylation of histone H4, and persisted up to telophase. Preventing histone deacetylation by the histone deacetylase inhibitor trichostatin A (TSA) led to accumulation of protoplasts at metaphase-anaphase, and reduced S10 phosphorylation during anaphase and telophase; in cultured tobacco cells, TSA significantly reduced the frequency of mitotic figures. Our results indicate that deacetylation of histone H4 and H3 in tobacco protoplasts occurs during mitosis in a phase-specific manner, and is important for progression through mitosis. PMID- 15659095 TI - ALS3 encodes a phloem-localized ABC transporter-like protein that is required for aluminum tolerance in Arabidopsis. AB - Aluminum (Al) toxicity in acid soils is a worldwide agricultural problem that severely limits crop productivity through inhibition of root growth. Previously, Arabidopsis mutants with increased Al sensitivity were isolated in order to identify genes important for Al tolerance in plants. One mutant, als3, exhibited extreme root growth inhibition in the presence of Al, suggesting that this mutation negatively impacts a gene required for Al tolerance. Map-based cloning of the als3-1 mutation resulted in the isolation of a novel gene that encodes a previously undescribed ABC transporter-like protein, which is highly homologous to a putative bacterial metal resistance protein, ybbM. Northern analysis for ALS3 expression revealed that it is found in all organs examined, which is consistent with the global nature of Al sensitivity displayed by als3, and that expression increases in roots following Al treatment. Based on GUS fusion and in situ hybridization analyses, ALS3 is primarily expressed in leaf hydathodes and the phloem throughout the plant, along with the root cortex following Al treatment. Immunolocalization indicates that ALS3 predominantly accumulates in the plasma membrane of cells that express ALS3. From our results, it appears that ALS3 encodes an ABC transporter-like protein that is required for Al resistance/tolerance and may function to redistribute accumulated Al away from sensitive tissues in order to protect the growing root from the toxic effects of Al. PMID- 15659096 TI - EGY1 encodes a membrane-associated and ATP-independent metalloprotease that is required for chloroplast development. AB - Chloroplast development requires coordinated expression of both nuclear- and chloroplast-encoded genes. To better understand the roles played by nuclear encoded chloroplast proteins in chloroplast biogenesis, we isolated an Arabidopsis mutant, egy1-1, which has a dual phenotype, reduced chlorophyll accumulation and abnormal hypocotyl gravicurvature. Subsequent map-based cloning and DNA sequencing of the mutant gene revealed a 10-bp deletion in an EGY1 gene, which encodes a 59-kDa metalloprotease that contains eight trans-membrane domains at its C-terminus, and carries out beta-casein degradation in an ATP-independent manner. EGY1 protein accumulation varies between tissue types, being most prominent in leaf and stem tissues, and is responsive to light and ethylene. EGY1 GFP hybrid proteins are localized in the chloroplast. egy1 mutant chloroplasts had reduced granal thylakoids and poorly developed lamellae networks. Furthermore, the accumulation of chlorophyll a/b binding proteins of the light harvesting complexes I and II (Lhca and Lhcb) are significantly decreased in three separate loss-of-function egy1 mutants. Taken together, these results suggest that EGY1 metalloprotease is required for chloroplast development and, hence, a defective EGY1 gene has pleiotropic effects both on chloroplast development and on ethylene-dependent gravitropism of light-grown hypocotyls. PMID- 15659097 TI - HUA2 is required for the expression of floral repressors in Arabidopsis thaliana. AB - The HUA2 gene acts as a repressor of floral transition. Lesions in hua2 were identified through a study of natural variation and through two mutant screens. An allele of HUA2 from Landsberg erecta (Ler) contains a premature stop codon and acts as an enhancer of early flowering 4 (elf4) mutants. hua2 single mutants, in the absence of the elf4 lesion, flower earlier than wild type under short days. hua2 mutations partially suppress late flowering in FRIGIDA (FRI )-containing lines, autonomous pathway mutants, and a photoperiod pathway mutant. hua2 mutations suppress late flowering by reducing the expression of several MADS genes that act as floral repressors including FLOWERING LOCUS C (FLC ) and FLOWERING LOCUS M (FLM ). PMID- 15659098 TI - Molecular and functional characterization of Arabidopsis Cullin 3A. AB - Cullin proteins, which belong to multigenic families in all eukaryotes, associate with other proteins to form ubiquitin protein ligases (E3s) that target substrates for proteolysis by the 26S proteasome. Here, we present the molecular and genetic characterization of a plant Cullin3. In contrast to fungi and animals, the genome of the model plant Arabidopsis thaliana contains two related CUL3 genes, called CUL3A and CUL3B. We found that CUL3A is ubiquitously expressed in plants and is able to interact with the ring-finger protein RBX1. A genomic search revealed the existence of at least 76 BTB-domain proteins in Arabidopsis belonging to 11 major families. Yeast two-hybrid experiments indicate that representative members of certain families are able to physically interact with both CUL3A and CUL3B, suggesting that Arabidopsis CUL3 forms E3 protein complexes with certain BTB domain proteins. In order to determine the function of CUL3A, we used a reverse genetic approach. The cul3a null mutant flowers slightly later than the control plants. Furthermore, this mutant exhibits a reduced sensitivity of the inhibition of hypocotyl growth in far-red light and miss-expresses COP1. The viability of the mutant plants suggests functional redundancy between the two CUL3 genes in Arabidopsis. PMID- 15659099 TI - X-ray diffraction structure of a plant glycosyl hydrolase family 32 protein: fructan 1-exohydrolase IIa of Cichorium intybus. AB - Fructan 1-exohydrolase, an enzyme involved in fructan degradation, belongs to the glycosyl hydrolase family 32. The structure of isoenzyme 1-FEH IIa from Cichorium intybus is described at a resolution of 2.35 A. The structure consists of an N terminal fivefold beta-propeller domain connected to two C-terminal beta-sheets. The putative active site is located entirely in the beta-propeller domain and is formed by amino acids which are highly conserved within glycosyl hydrolase family 32. The fructan-binding site is thought to be in the cleft formed between the two domains. The 1-FEH IIa structure is compared with the structures of two homologous but functionally different enzymes: a levansucrase from Bacillus subtilis (glycosyl hydrolase family 68) and an invertase from Thermotoga maritima (glycosyl hydrolase family 32). PMID- 15659100 TI - In vivo studies on the roles of Tic110, Tic40 and Hsp93 during chloroplast protein import. AB - A multisubunit translocon of the inner envelope membrane, termed Tic, mediates the late stages of protein import into chloroplasts. Membrane proteins, Tic110 and Tic40, and a stromal chaperone, Hsp93, have been proposed to function together within the Tic complex. In Arabidopsis, single genes, atTIC110 and atTIC40, encode the Tic proteins, and two homologous genes, atHSP93-V and atHSP93 III, encode Hsp93. These four genes exhibited relatively uniform patterns of expression, suggesting important roles for plastid biogenesis throughout development and in all tissues. To investigate the roles played by these proteins in vivo, we conducted a comparative study of T-DNA knockout mutants for each Tic gene, and for the most abundantly expressed Hsp93 gene, atHSP93-V. In the homozygous state, the tic110 mutation caused embryo lethality, implying an essential role for atTic110 during plastid biogenesis. Homozygous tic110 embryos exhibited retarded growth, developmental arrest at the globular stage and a 'raspberry-like' embryo-proper phenotype. Heterozygous tic110 plants, and plants homozygous for the tic40 and hsp93-V mutations, exhibited chlorosis, aberrant chloroplast biogenesis, and inefficient chloroplast-import of both photosynthetic and non-photosynthetic preproteins. Non-additive interactions amongst the mutations occurred in double mutants, suggesting that the three components may cooperate during chloroplast protein import. PMID- 15659101 TI - A unique family of proteins associated with internalized membranes in protein storage vacuoles of the Brassicaceae. AB - The protein storage vacuole (PSV) is a specialized organelle in plant seeds that accumulates storage proteins and phytate during seed development. In many plant species, such as tomato and tobacco, the PSV contains two types of microscopically visible intra-organellar inclusions: a large crystalline lattice of membranes and proteins, the crystalloid, and one or a few large phytate crystals, the globoids. In seeds of the family Brassicaceae, the PSVs lack visible crystalloids and have many small globoids dispersed throughout. We biochemically fractionated PSVs from Brassica napus and defined a crystalloid like fraction that contained integral membrane protein markers found in crystalloids of other plants. Protein analyses identified a previously undescribed family of proteins, the Brassicaceae PSV-embedded proteins (BPEPs), associated with 'crystalloid' and globoid fractions. The defining characteristics of the BPEPs are an N-terminal signal peptide and tandem MATH domains, which may mediate protein-protein interactions. Database analyses indicated that the BPEPs are unique to Brassicaceae. Immunofluorescence studies using anti-BPEP antibodies and antibodies to other biochemical markers to label B. napus and Arabidopsis thaliana seed sections localized the BPEPs to structures within the PSVs, whose appearance was consistent with a diffuse network of internalized membranes and globoids. These results demonstrate that Brassicaceae PSVs contain internalized membranes, and raise the possibility that BPEPs modify these internal membrane structures to yield a PSV morphology different from that of tomato or tobacco. PMID- 15659102 TI - A two-component high-affinity nitrate uptake system in barley. AB - The analysis of genome databases for many different plants has identified a group of genes that are related to one part of a two-component nitrate transport system found in algae. Earlier work using mutants and heterologous expression has shown that a high-affinity nitrate transport system from the unicellular green algae, Chlamydomonas reinhardtii required two gene products for function. One gene encoded a typical carrier-type structure with 12 putative trans-membrane (TM) domains and the other gene, nar2 encoded a much smaller protein that had only one TM domain. As both gene families occur in plants we investigated whether this transport model has more general relevance among plants. The screening for nitrate transporter activity was greatly helped by a novel assay using (15)N enriched nitrate uptake into Xenopus oocytes expressing the proteins. This assay enables many oocytes to be rapidly screened for nitrate transport activity. The functional activity of a barley nitrate transporter, HvNRT2.1, in oocytes required co-injection of a second mRNA. Although three very closely related nar2 like genes were cloned from barley, only one of these was able to give functional nitrate transport when co-injected into oocytes. The nitrate transport performed by this two-gene system was inhibited at more acidic external pH and by acidification of the cytoplasm. This specific requirement for two-gene products to give nitrate transport function has important implications for attempts to genetically manipulate this fundamental process in plants. PMID- 15659103 TI - Arabidopsis SHMT1, a serine hydroxymethyltransferase that functions in the photorespiratory pathway influences resistance to biotic and abiotic stress. AB - We found that a recessive mutation, shmt1-1, causes aberrant regulation of cell death resulting in chlorotic and necrotic lesion formation under a variety of environmental conditions. Salicylic acid-inducible genes and genes involved in H(2)O(2) detoxification were expressed constitutively in shmt1-1 plants in direct correlation with the severity of the lesions. The shmt1-1 mutants were more susceptible than control plants to infection with biotrophic and necrotrophic pathogens, developing severe infection symptoms in a high percentage of infected leaves. In addition, mutants carrying shmt1-1 or a loss-of-function shmt1-2 allele, were smaller and showed a greater loss of chlorophyll and greater accumulation of H(2)O(2) than wild-type plants when subjected to salt stress. SHMT1 was map-based cloned and found to encode a serine hydroxymetyltransferase (SHMT1) involved in the photorespiratory pathway. Our results indicate that this enzymatic activity plays a critical role in controlling the cell damage provoked by abiotic stresses such as high light and salt and in restricting pathogen induced cell death, supporting the notion that photorespiration forms part of the dissipatory mechanisms of plants to minimize production of reactive oxygen species (ROS) at the chloroplast and to mitigate oxidative damage. Moreover, results shown here indicate that whereas production of ROS is an essential component of the hypersensitive defense response, the excessive accumulation of these toxic compounds impairs cell death containment and counteracts the effectiveness of the plant defenses to restrict pathogen infection. PMID- 15659104 TI - Identification of Arabidopsis thaliana transformants without selection reveals a high occurrence of silenced T-DNA integrations. AB - Several recent investigations of T-DNA integration sites in Arabidopsis thaliana have reported 'cold spots' of integration, especially near centromeric regions. These observations have contributed to the ongoing debate over whether T-DNA integration is random or occurs preferentially in transcriptionally active regions. When transgenic plants are identified by selecting or screening for transgenic activity, transformants with integrations into genomic regions that suppress transcription, such as heterochromatin, may not be identified. This phenomenon, which we call selection bias, may explain the perceived non-random distribution of T-DNA integration in previous studies. In order to investigate this possibility, we have characterized the sites of T-DNA integration in the genomes of transgenic plants identified by pooled polymerase chain reaction (PCR), a procedure that does not require expression of the transgene, and is therefore free of selection bias. Over 100 transgenic Arabidopsis plants were identified by PCR and compared with kanamycin-selected transformants from the same T(1) seed pool. A higher perceived transformation efficiency and a higher frequency of transgene silencing were observed in the PCR-identified lines. Together, the data suggest approximately 30% of transformation events may result in non-expressing transgenes that would preclude identification by selection. Genomic integration sites in PCR-identified lines were compared with those in existing T-DNA integration databases. In PCR-identified lines with silenced transgenes, the integration sites mapped to regions significantly underrepresented by T-DNA integrations in studies where transformants were identified by selection. The data presented here suggest that selection bias can account for at least some of the observed non-random integration of T-DNA into the Arabidopsis genome. PMID- 15659105 TI - A study in scarlet: enzymes of ketocarotenoid biosynthesis in the flowers of Adonis aestivalis. AB - The red ketocarotenoid astaxanthin (3,3'-dihydroxy-4,4'-diketo-beta,beta carotene) is widely used as an additive in feed for the pigmentation of fish and crustaceans and is frequently included in human nutritional supplements as well. There is considerable interest in developing a plant-based biological production process for this valuable carotenoid. Adonis aestivalis (Ranunculaceae) is unusual among plants in synthesizing and accumulating large amounts of astaxanthin and other ketocarotenoids. The formation of astaxanthin requires only the addition of a carbonyl at the number 4 carbon of each beta-ring of zeaxanthin (3,3'-dihydroxy-beta,beta-carotene), a carotenoid typically present in the green tissues of higher plants. We screened an A. aestivalis flower library to identify cDNAs that might encode the enzyme that catalyzes the addition of the carbonyls. Two closely related cDNAs selected in this screen were found to specify polypeptides similar in sequence to plant beta-carotene 3-hydroxylases, enzymes that convert beta-carotene (beta,beta-carotene) into zeaxanthin. The Adonis enzymes, however, exhibited neither 4-ketolase nor 3-hydroxylase activity when presented with beta-carotene as the substrate in Escherichia coli. Instead, the products of the Adonis cDNAs were found to modify beta-rings in two distinctly different ways: desaturation at the 3,4 position and hydroxylation of the number 4 carbon. The 4-hydroxylated carotenoids formed in E. coli were slowly metabolized to yield compounds with ketocarotenoid-like absorption spectra. It is proposed that a 3,4-desaturation subsequent to 4-hydroxylation of the beta-ring leads to the formation of a 4-keto-beta-ring via an indirect and unexpected route: a keto-enol tautomerization. PMID- 15659106 TI - Quantification of allele-specific expression of a gene encoding strawberry polygalacturonase-inhibiting protein (PGIP) using Pyrosequencing. AB - Recent studies indicate that allele-specific differences in gene expression are a common phenomenon. The extent to which differential allelic expression exists might be underestimated, due to the limited accuracy of the methods used so far. To demonstrate allele-specific expression, we investigated the transcript abundance of six individual, highly homologous alleles of a polygalacturonase inhibiting protein gene (FaPGIP) from octoploid strawberry (Fragaria x ananassa). We applied the highly quantitative Pyrosequencing method which, for the gene under study, detected allele frequency differences as small as 4.0 +/- 2.8%. Pyrosequencing of RT-PCR products showed that one FaPGIP allele was preferentially expressed in leaf tissue, while two other alleles were expressed in a fruit-specific way. For fruits that were inoculated with Botrytis cinerea a strong increase in overall FaPGIP gene expression was observed. This upregulation was accompanied by a significant change in FaPGIP allele frequencies when compared with non-treated fruits. Remarkably, in the five cultivars tested, the allele frequency in cDNA from the inoculated fruits was similar to that in genomic DNA, suggesting uniform upregulation of all FaPGIP alleles present as a result of pathogenesis-related stress. The results demonstrate that when Pyrosequencing of RT-PCR products is performed, novel allele-specific gene regulation can be detected and quantified. PMID- 15659107 TI - Recombinant bispecific antibodies for cancer therapy. AB - Bispecific antibodies can serve as mediators to retarget effector mechanisms to disease-associated sites. Studies over the past two decades have revealed the potentials but also the limitations of conventional bispecific antibodies. The development of recombinant antibody formats has opened up the possibility of generating bispecific molecules with improved properties. This review summarizes recent developments in the field of recombinant bispecific antibodies and discusses further requirements for clinical development. PMID- 15659109 TI - Intracellular dopamine oxidation mediates rotenone-induced apoptosis in PC12 cells. AB - AIM: To study the role of dopamine (DA) in rotenone-induced neurotoxicity in PC12 cells. METHODS: Cell viability was assessed by detecting the leakage of lactate dehydrogenase (LDH) into the medium. Apoptosis rate was measured by flow cytometry. Caspase-3-like activity was measured by fluorescence assay using the probe Ac-DEVD-AMC. The level of intracellular hydrogen peroxide and other peroxides in PC12 cells were quantified by loading cells with 2'-7' Dichlorodihydrofluorescein diacetate (DCFH-DA) in fluorescence assay. Lactic acid was measured spectrophotometrically. The DA levels in PC12 cells were determined by HPLC-ECD. RESULTS: A 48-h incubation of PC12 cells with rotenone caused an apoptotic cell death and elevated intracellular reactive oxygen species (ROS) and lactic acid accumulation. Intracellular DA depletion with reserpine significantly attenuated rotenone-induced ROS accumulation and apoptotic cell death. No change was found in rotenone-induced ROS accumulation when cells were co-treated with deprenyl. Brief treatment with reserpine at the end of rotenone treatment had no effect on rotenone-induced neurotoxicity. However, when cells were first incubated with deprenyl, a monoamine oxidase-B inhibitor for 30 min then co incubated with rotenone plus deprenyl, a brief treatment with reserpine enhanced cell injury. CONCLUSION: Rotenone-induced apoptosis in PC12 cells was mediated by intracellular dopamine oxidation. PMID- 15659108 TI - Tonic activation of presynaptic GABA(B) receptors on rat pallidosubthalamic terminals. AB - AIM: The subthalamic nucleus plays a critical role in the regulation of movement, and abnormal activity of its neurons is associated with some basal ganglia motor symptoms. We examined the presence of functional presynaptic GABA(B) receptors on pallidosubthalamic terminals and tested whether they were tonically active in the in vitro subthalamic slices. METHODS: Whole-cell patch-clamp recordings were applied to acutely prepared rat subthalamic nucleus slices. The effects of specific GABA(B) agonist and antagonist on action potential-independent inhibitory postsynaptic currents (IPSCs), as well as holding current, were examined. RESULTS: Superfusion of baclofen, a GABA(B) receptor agonist, significantly reduced the frequency of GABA(A) receptor-mediated miniature IPSCs (mIPSCs), in a Cd2+-sensitive manner, with no effect on the amplitude, indicating presynaptic inhibition on GABA release. In addition, baclofen induced a weak outward current only in a minority of subthalamic neurons. Both the pre- and post synaptic effects of baclofen were prevented by the specific GABA(B) receptor antagonist, CGP55845. Furthermore, CGP55845 alone increased the frequency of mIPSCs, but had no effect on the holding current. CONCLUSION: These findings suggest the functional dominance of presynaptic GABA(B) receptors on the pallidosubthalamic terminals over the postsynaptic GABA(B) receptors on subthalamic neurons. Furthermore, the presynaptic, but not the postsynaptic, GABA(B) receptors are tonically active, suggesting that the presynaptic GABA(B) receptors in the subthalamic nucleus are potential therapeutic target for the treatment of Parkinson disease. PMID- 15659110 TI - Cloning, expression, and functional analysis of human dopamine D1 receptors. AB - AIM: To construct an HEK293 cell line stably expressing human dopamine D1 receptor (D1R). METHODS: cDNA was amplified by RT-PCR using total RNA from human embryo brain tissue as the template. The PCR products were subcloned into the plasmid pcDNA3 and cloned into the plasmid pcDNA3.1. The cloned D1R cDNA was sequenced and stably expressed in HEK293 cells. Expression of D1R in HEK293 cells was monitored by the [3H]SCH23390 binding assay. The function of D1R was studied by the cAMP accumulation assay, CRE-SEAP reporter gene activity assay, and intracellular calcium assay. RESULTS: An HEK293 cell line stably expressing human D1R was obtained. A saturation radioligand binding experiment with [3H]SCH23390 demonstrated that the Kd and Bmax values were 1.5+/-0.2 nmol/L and 2.94+/-0.15 nmol/g of protein, respectively. In the [3H]SCH23390 competition assay, D1R agonist SKF38393 displaced [3H]SCH23390 with an IC50 value of 2.0 (1.5-2.8) micromol/L. SKF38393 increased the intracellular cAMP level and CRE-SEAP activity through D1R expressed in HEK293 cells in a concentration-dependent manner with an EC50 value of 0.25 (0.12-0.53) micromol/L and 0.39 (0.27-0.57) micromol/L at 6 h/0.59 (0.22-1.58) micromol/L at 12 h, respectively. SKF38393 also increased the intracellular calcium level in a concentration-dependent manner with EC50 value of 27 (8.6-70) nmol/L. CONCLUSION: An HEK293 cell line stably expressing human D1R was obtained successfully. The study also demonstrated that the CRE-SEAP activity assay could be substituted for the cAMP accumulation assay for measuring increase in cAMP levels. Thus, both intracellular calcium measurements and the CRE-SEAP activity assay are suitable for high-throughput screening in drug research. PMID- 15659111 TI - Antiapoptotic effect both in vivo and in vitro of A20 gene when transfected into rat hippocampal neurons. AB - AIM: To evaluate the antiapoptotic effect of the A20 gene in primary hippocampal neurons both in vivo and in vitro. METHODS: Primary hippocampal neurons in embryonic day 18 (E18) rats were transfected with the A20 gene by using the new Nucleofector electroporation transfection method. We then examined, whether A20 neurons possessed anti-apoptotic abilities after TNF-alpha stimulation in vitro. A20-neurons and pcDNA3-neurons were transplanted into the penumbra of the brains of rats that had been subjected to 90-min of ischemia induced by left middle cerebral artery occlusion (MCAO). RESULTS: A20-neurons resisted TNF-alpha induced apoptosis in vitro. The apoptosis rate of neurons overexpressing A20 (28.46%+/ 3.87%) was lower than that in neurons transfected with pcDNA3 (53.06%+/-5.36%). More A20-neurons survived in the penumbra both 3-d and 7-d after transplantation than did sham pcDNA3 neurons. CONCLUSION: The novel function of A20 may make it a potential targets for the gene therapy for neurological diseases. PMID- 15659112 TI - Validation of a simple automated movement detection system for formalin test in rats. AB - AIM: To investigate the validity and sensitivity of an automatic movement detection system developed by our laboratory for the formalin test in rats. METHODS: The effects of systemic morphine and local anesthetic lidocaine on the nociceptive behaviors induced by formalin subcutaneously injected into the hindpaw were examined by using an automated movement detection system and manual measuring methods. RESULTS: Formalin subcutaneously injected into the hindpaw produced typical biphasic nociceptive behaviors (agitation). The mean agitation event rate during a 60-min observation period increased linearly following increases in the formalin concentration (0.0%, 0.5%, 1.5%, 2.5%, and 5%, 50 microL). Systemic application of morphine of different doses (1, 2, and 5 mg/kg) 10-min prior to formalin injection depressed the agitation responses induced by formalin injection in a dose-dependent manner, and the antinociceptive effect induced by the largest dose (5 mg/kg) of morphine was significantly antagonized by systemic application of the opioid receptor antagonist naloxone (1.25 mg/kg). Local anesthetic lidocaine (20 mg/kg) injected into the ipsilateral ankle subskin 5-min prior to formalin completely blocked the agitation response to formalin injection. These results were comparable to those obtained from manual measure of the incidence of flinching or the duration time of licking/biting of the injected paw. CONCLUSION: These data suggest that this automated movement detection system for formalin test is a simple, validated measure with good pharmacological sensitivity suitable for discovering novel analgesics or investigating central pain mechanisms. PMID- 15659113 TI - Intrathecal administration of roscovitine inhibits Cdk5 activity and attenuates formalin-induced nociceptive response in rats. AB - AIM: To investigate effects of the cyclin-dependent kinase5 (Cdk5) inhibitor roscovitine on formalin-induced nociceptive responses in rats. METHODS: The flinch response as a methood of pain threshold measurement and intrathecal injection techniques were used. Cdk5 and phosphorylation of its downstream target, DARPP-32 (dopamine- and cAMP-regulated phosphoprotein of M(r) 32 kDa), were investigated by Western blot analysis. RESULTS: Rats demonstrated a typical flinch response after formalin injection. Intrathecal roscovitine injections significantly suppressed the flinch response in a dose-dependent manner. Western blot analysis showed that phosphorylated DARPP-32 at Thr75 increased in concentration after formalin hyperalgesia, with this effect reduced by roscovitine administration. This antinociception was partially attenuated by administration of naloxone before the formalin test. CONCLUSION: DARPP-32 phosphorylation is involved in acute inflammatory pain response. Intrathecal roscovitine administration attenuates formalin-induced nociceptive responses and there is potential for further application. PMID- 15659114 TI - Construction of phospholamban antisense RNA recombinant adeno-associated virus vector and its effects in rat cardiomyocytes. AB - AIM: To construct a recombinant adeno-associated virus (rAAV) vector containing gene encoding phospholamban antisense RNA (asPLB), and analyse its effect on expression of PLB, expression and activity of sarco-endoplasmic reticulum Ca2+ ATPase (SERCA), and the change of intracellular free Ca2+ concentration ([Ca2+]i) in rat cardiomyocytes. METHODS: The target gene encoding PLB antisense RNA was inserted inversely into the adeno-associated virus plasmid pAAV-MCS digested by corresponding restricted endonuclease enzyme. The recombinant plasmid and pAAV-RC and pHelper were co-transfected into 293 cell. At the same time, a viral production positive control (rAAV-LacZ) and negative control were performed. The recombinant viruses were used to transfect the cultured rat cardiomyocytes. Site beta-Galactosidase staining were performed to observe the transfer efficiency. Reverse transcription-PCR and Western blot were used to determine the mRNA and protein expression of PLB and SERCA. The activity of SERCA and the [Ca2+]i were measured. RESULTS: The rAAV vectors were constructed successfully and were transfected into rat cardiomyocytes effectively. The PLB mRNA and protein expression were reduced in rat cardiomyocytes transfected by rAAV-asPLB compared with controls. The activity of SERCA was increased. In rest state, the level of [Ca2+]i in the rAAV-asPLB transfected group decreased. The level of [Ca2+]i increased when induced by isoproterenol. CONCLUSION: AAV-asPLB vector was constructed successfully, which disrupted the expression of PLB, enhanced the activity of SERCA, reduced the resting [Ca2+]i, and improved the cardiac function. PMID- 15659115 TI - Ginsenoside Rg1 reduces MPTP-induced substantia nigra neuron loss by suppressing oxidative stress. AB - AIM: To investigate the effect of ginsenoside Rg1, an effective ingredient from ginsenoside, on 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced substantia nigra neuron lesion. METHODS: C57-BL mice were given MPTP to prepare Parkinson disease mice model. Different doses of Rg1 (5, 10, and 20 mg.kg(-1).d( 1)) or N-acetylcystein (NAC) (300 mg.kg(-1).d(-1)) were given 3 d prior to MPTP in the pretreatment groups. Glutathione (GSH) level and total superoxide dismutase (T-SOD) activity in substantia nigra were determined by spectrophotometry. Nissl staining, tyrosine hydroxylase immunostaining, and TUNEL labeling were used to observe the damage and apoptosis of nigral neurons. Western blot analysis was used to detect the phospho-JNK and phospho-c-Jun levels in midbrain homogenates. RESULTS: Pretreatments of C57-BL mice with different doses of Rg1 or NAC were found to protect against MPTP-induced substantia nigra neurons loss. Rg1 or NAC prevented GSH reduction and T-SOD activation in substantia nigra, and attenuated the phosphorylations of JNK and c-Jun following MPTP treatment. CONCLUSION: The antioxidant property of Rg1 along with the blocking of JNK signaling cascade might contribute to the neuroprotective effect of ginsenoside Rg1 against MPTP. PMID- 15659116 TI - Neuropeptide Y expression in mouse hippocampus and its role in neuronal excitotoxicity. AB - AIM: To investigate neuropeptide Y (NPY) expression in mouse hippocampus within early stages of kainic acid (KA) treatment and to understand its role in neuronal excitotoxicity. METHODS: NPY expression in the hippocampus within early stages of KA intraperitoneal (ip) treatment was detected by immunohistochemistry (IHC) and in situ hybridization (ISH) methods. The role of NPY and Y5, Y2 receptors in excitotoxicity was analyzed by terminal deoxynucleotidyl transferase-mediated UTP nick end-labeling (TUNEL) assay. RESULTS: Using IHC assay, in granule cell layer of the dentate gyrus (DG), NPY positive signals appeared 4 h after KA injection, reached the peak at 8 h and leveled off at 16 and 24 h. In CA3, no positive signal was found within the first 4 h after KA injection, but strong signal appeared at 16 and 24 h. No noticeable signal was detected in CA1 at all time points after KA injection. Using the ISH method, positive signals were detected at 4, 8, and 16 h in CA3, CA1, and hilus. In DG, much stronger ISH signals were detected at 4 h, but leveled off at 8 and 16 h. TUNEL analysis showed that intracerebroventricularly (icv) infusion of NPY and Y5, Y2 receptor agonists within 8 h after KA insult with proper dose could remarkably rescue pyramidal neurons in CA3 and CA1 from apoptosis. CONCLUSION: NPY is an important anti epileptic agent. The preceding elevated expression of NPY in granule cell layer of DG after KA injection might partially explain its different excitotoxicity induced apoptotic responses in comparison with the pyramidal neurons from CA3 and CA1 regions. NPY can not only reduce neuronal excitability but also prevent excitotoxicity-induced neuronal apoptosis in a time- and dose-related way by activation of Y5 and Y2 receptors. PMID- 15659117 TI - Alteration of binding sites for [3H]P1075 and [3H]glibenclamide in renovascular hypertensive rat aorta. AB - AIM: The alterations of the binding sites for ATP-sensitive K+ channel (K(ATP)) openers and blockers in aortic strips were investigated in hypertensive rats. METHODS: Radioligand binding techniques were used to compare the specific binding properties of [3H]P1075 and [3H]glibenclamide (Gli) in normotensive (NWR) and reno-vascular hypertensive rat (RVHR) aortic strips. RESULTS: The KD values of [3H]P1075 binding were increased by 1.5-fold, while the Bmax values were unchanged in RVHR. The IC50 values of P1075 and pinacidil (Pin) for displacing the [3H]P1075 binding in RVHR were increased by 1.8- and 1.7-fold, respectively. The kinetic processes of association and dissociation of [3H]P1075 binding were slower in RVHR. Glibenclamide pretreatment slowed down the kinetic processes of the association and dissociation of [3H]P1075 binding in NWR, but failed to alter the kinetic processes of [3H]P1075 binding in RVHR. The IC50 values of Gli for displacing the [3H]Gli binding at high-affinity sites were increased by 3-fold, while those at low-affinity sites remained to be unchanged in RVHR. The kinetic processes of association of [3H]Gli binding were decreased and those of the dissociation were accelerated in RVHR. The treatment with Pin slowed down the association kinetic processes but accelerated the process of the dissociation of [3H]Gli binding in NWR, but did not alter the kinetics of [3H]Gli binding in RVHR. CONCLUSION: The affinity of binding sites for [3H]P1075 and of high affinity binding sites for [3H]Gli are decreased, and the negative allosteric interactions between the two binding sites are impaired in RVHR aorta. PMID- 15659118 TI - Ca2+ participates in alpha1B-adrenoceptor-mediated cAMP response in HEK293 cells. AB - AIM: To investigate the alpha1B-adrenoceptor (alpha1B-AR)-mediated cAMP response and underlying mechanisms in HEK293 cells. METHODS: Full-length cDNA encoding alpha1B-AR was transfected into HEK293 cells using the calcium phosphate precipitation method, and alpha1B-AR expression and cAMP accumulation were determined by using the saturation radioligand binding assay and ion-exchange chromatography, respectively. RESULTS: Under agonist stimulation, alpha1B-AR mediated cAMP synthesis in HEK293 cells, and blockade by PLC-PKC or tyrosine kinase did not reduce cAMP accumulation induced by NE. Pretreatment with pertussis toxin (PTX) had little effect on basal cAMP accumulation as well as norepinephrine (NE)-stimulated cAMP accumulation. In addition, pretreatment with cholera toxin (CTX) neither mimicked nor blocked the effect induced by NE. The extracellular Ca2+ chelator egtazic acid (EGTA), nonselective Ca2+ channel blocker CdCl2 and calmodulin (CaM) inhibitor W-7 significantly reduced NE-induced cAMP accumulation from 1.59%+/-0.47% to 1.00%+/-0.31%, 0.78%+/-0.23%, and 0.90%+/ 0.40%, respectively. CONCLUSION: By coupling with a PTX-insensitive G protein, alpha1BAR promotes Ca2+ influx via receptor-dependent Ca2+ channels, then Ca2+ is linked to CaM to form a Ca2+-CaM complex, which stimulates adenylyl cyclase (AC), thereby increasing the cAMP production in HEK293 cell lines. PMID- 15659119 TI - Signal pathways underlying homocysteine-induced production of MCP-1 and IL-8 in cultured human whole blood. AB - AIM: To elucidate the mechanisms underlying homocysteine (Hcy)-induced chemokine production. METHODS: Human whole blood was pretreated with inhibitors of calmodulin (CaM), protein kinase C (PKC), protein tyrosine kinase (PTK), mitogen activated protein kinase (MAPK), and NF-kappaB and activators of PPARgamma for 60 min followed by incubation with Hcy 100 micromol/L for 32 h. The levels of mitogen chemokine protein (MCP)-1 and interleukin-8 (IL-8) were determined by enzyme-linked immunosorbant assay (ELISA). RESULTS: Inhibitors of PKC (calphostin C, 50-500 nmol/L and RO-31-8220, 10-100 nmol/L), CaM (W7, 28-280 micromol/L), ERK1/2 MAPK (PD 98059, 2-20 micromol/L), p38 MAPK (SB 203580, 0.6-6 micromol/L), JNK MAPK (curcumin, 2-10 micromol/L), and NF-kappaB (PDTC, 10-100 nmol/L) markedly reduced Hcy 100 micromol/L-induced production of MCP-1 and IL-8 in human cultured whole blood, but the inhibitors of PTK (genistein, 2.6-26 micromol/L and tyrphostin, 0.5-5 micromol/L) had no obvious effect on MCP-1 and IL-8 production. PPARgamma activators (ciglitazone 30 micromol/L and troglitazone 10 micromol/L) depressed the Hcy-induced MCP-1 production but not IL-8 production in the cultured whole blood. CONCLUSION: Hcy-induced MCP-1 and IL-8 production is mediated by activated signaling pathways such as PKC, CaM, MAPK, and NF-kappaB. Our results not only provide clues for the signal transduction pathways mediating Hcy-induced chemokine production, but also offer a plausible explanation for a pathogenic role of hyperhomocysteinemia in these diseases. PMID- 15659120 TI - Effect of pravastatin on impaired endothelium-dependent relaxation induced by lysophosphatidylcholine in rat aorta. AB - AIM: To investigate the effects of pravastatin, a potent 3-hydroxy-3 methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor, on impaired endothelium dependent relaxation induced by lysophosphatidylcholine (LPC), the major component of oxidized low-density lipoprotein, in rat thoracic aorta. METHODS: Both the endothelium-dependent relaxation response to acetylcholine and the endothelium-independent relaxation response to sodium nitroprusside of aortic rings were measured by recording isometric tension after the rings were exposed to LPC in the absence or presence of pravastatin to estimate the injury effect of LPC and the protective effect of pravastatin on the aortic endothelium, respectively. RESULTS: Exposure of aortic rings to LPC (1-10 micromol/L) for 30 min induced a significant concentration-dependent inhibition of endothelium dependent relaxation to acetylcholine, but did not affect endothelium-independent relaxation in response to sodium nitroprusside. Pre-incubation of aortic rings with pravastatin (0.3-3 mmol/L) for 15 min and then co-incubation of the rings with LPC (3 micromol/L) for another 30 min significantly attenuated the inhibition of endothelium-dependent relaxation induced by LPC. This protective effect of pravastatin (1 mmol/L) was abolished by NG-nitro-L-arginine methyl ester (30 micromol/L), an inhibitor of nitric oxide synthase, but not by indomethacin (10 micromol/L), an inhibitor of cyclooxygenase. Moreover, protein kinase C inhibitor chelerythrine (1 micromol/L) the superoxide anion scavenger superoxide dismutase (200 kU/L), and the nitric oxide precursor L-arginine (3 mmol/L) also improved the impaired endothelium-dependent relaxation induced by LPC, similar to the effects of pravastatin. CONCLUSION: Pravastatin can protect the endothelium against functional injury induced by LPC in rat aorta, a fact which is related to increasing nitric oxide bioavailability. PMID- 15659122 TI - Using support vector classification for SAR of fentanyl derivatives. AB - AIM: To discriminate between fentanyl derivatives with high and low activities. METHODS: The support vector classification (SVC) method, a novel approach, was employed to investigate structure-activity relationship (SAR) of fentanyl derivatives based on the molecular descriptors, which were quantum parameters including DeltaE [energy difference between highest occupied molecular orbital energy (HOMO) and lowest empty molecular orbital energy (LUMO)], MR (molecular refractivity) and M(r) (molecular weight). RESULTS: By using leave-one-out cross validation test, the accuracies of prediction for activities of fentanyl derivatives in SVC, principal component analysis (PCA), artificial neural network (ANN) and K-nearest neighbor (KNN) models were 93%, 86%, 57%, and 71%, respectively. The results indicated that the performance of the SVC model was better than those of PCA, ANN, and KNN models for this data. CONCLUSION: SVC can be used to investigate SAR of fentanyl derivatives and could be a promising tool in the field of SAR research. PMID- 15659121 TI - Enzymatic activity characterization of SARS coronavirus 3C-like protease by fluorescence resonance energy transfer technique. AB - AIM: To characterize enzymatic activity of severe acute respiratory syndrome (SARS) coronavirus (CoV) 3C-like protease (3CL(pro)) and its four site-directed mutants. METHODS: Based on the fluorescence resonance energy transfer (FRET) principle using 5-[(2'-aminoethyl)-amino] naphthelenesulfonic acid (EDANS) and 4 [[4-(dimethylamino) phenyl] azo] benzoic acid (Dabcyl) as the energy transfer pair, one fluorogenic substrate was designed for the evaluation of SARS-CoV 3CL(pro) proteolytic activity. RESULTS: The kinetic parameters of the fluorogenic substrate have been determined as Km=404 micromol.L(-1), kcat=1.08 min(-1), and kcat/Km=2.7 mmol(-1).L.min(-1). SARS-CoV 3CL(pro) showed substantial pH and temperature-triggered activity switches, and site-directed mutagenesis analysis of SARS-CoV 3CL(pro) revealed that substitutions of His41, Cys145, and His163 resulted in complete loss of enzymatic activity, while replacement of Met162 with Ala caused strongly increased activity. CONCLUSION: This present work has provided valuable information for understanding the catalytic mechanism of SARS CoV 3CL(pro). This FRET-based assay might supply an ideal approach for the exploration SARS-CoV 3CL(pro) putative inhibitors. PMID- 15659123 TI - Evaluation of drug-muscarinic receptor affinities using cell membrane chromatography and radioligand binding assay in guinea pig jejunum membrane. AB - AIM: To study if cell membrane chromatography (CMC) could reflect drug-receptor interaction and evaluate the affinity and competitive binding to muscarinic acetylcholine receptor (mAChR). METHODS: The cell membrane stationary phase (CMSP) was prepared by immobilizing guinea pig jejunum cell membrane on the surface of a silica carrier, and was used for the rapid on-line chromatographic evaluation of ligand binding affinities to mAChR. The affinity to mAChR was also evaluated from radioligand binding assays (RBA) using the same jejunum membrane preparation. RESULTS: The capacity factor (k') profiles in guinea pig jejunum CMSP were: (-)QNB (15.4)>(+)QNB (11.5)>atropine (5.35)>pirenzepine (5.26)>4-DAMP (4.45)>AF-DX116 (4.18)>pilocarpine (3.93)>acetylcholine (1.31). These results compared with the affinity rank orders obtained from radioligand binding assays indicated that there was a positive correlation (r2= 0.8525, P<0.0001) between both data sets. CONCLUSION: The CMC method can be used to evaluate drug-receptor affinities for drug candidates. PMID- 15659124 TI - Melatonin attenuates 1-methyl-4-phenylpyridinium-induced PC12 cell death. AB - AIM: To explore the effect of melatonin on PC12 cell death induced by 1-methyl-4 phenylpyridinium (MPP+). METHODS: MTT assay, lactate dehydrogenase (LDH) efflux assay, and immunohistochemistry methods were used to measure neurotoxicity of PC12 cells treated acutely with MPP+ in low glucose and high glucose conditions, and to assess the neuroprotective effect of melatonin on PC12 cell death induced by MPP+. RESULTS: In a low glucose condition, MPP+ significantly induced PC12 cell death, which showed time and concentration dependence. In a serum-free low glucose condition, the percentages of viability of cells treated with MPP+ for 12, 24, 48, 72, and 96 h were 85.1%, 75.4%, 64.9%, 28.15%, and 9%, respectively. The level of LDH in the culture medium increased and tyrosine hydroxylase positive (TH+) cell count decreased. However, in a serum-free high glucose condition, MPP+ did not significantly induce PC12 cell death compared with control at various concentrations and time regimens. When the cells were preincubated with melatonin 250 micromol/L for 48, 72, and 96 h in a serum-free low glucose condition, cell survival rate significantly increased to 78.1%, 58.8%, and 31.6%, respectively. Melatonin abolished the LDH leakage of cells treated with MPP+ and increased TH+ cells count. CONCLUSION: MPP+ caused concentration-dependent PC12 cell death. The level of glucose was an important factor to MPP+ induced dopaminergic PC12 cell death. Low glucose level could potentiate MPP+ toxicity, while high glucose level could reduce the toxicity. In addition, melatonin attenuated PC12 cell death induced by MPP+. PMID- 15659126 TI - Objective measures of health-related quality of life over 24 months post-liver transplantation. AB - Many studies have reported improved health-related quality of life (HRQoL) from pre- to immediate post-orthotopic liver transplantation (OLT). However, few studies have evaluated longitudinal changes over the first 2 yr post-OLT and none have simultaneously examined objective measures of health-related fitness. A total of 50 OLT recipients (32 males,18 females; 51.4 +/- 11.8 yr) completed testing at 2, 6, 12, and 24 months post-OLT. Testing included assessment of exercise capacity (peak VO2), quadriceps muscle strength, body composition, physical activity participation, and self-reported functioning (SF-36). Repeated measures of analysis of variance (ANOVA) with post hoc contrasts was performed to determine differences over time and a second ANOVA assessed differences over time between genders. All patients increased peak VO2, quadriceps muscle strength, and percent body fat (p < 0.0001) from 2 to 24 months. Men and women differed in their changes of peak VO2 and percent body fat (p < 0.05). At 24 months, only 50% of the patients reported participating in regular physical activity. All SF-36 physical measures except general health, improved from 2 to 24 months (p < 0.0001). Measures of health-related fitness and QoL improve over the first 2 yr post-OLT with the greatest gains occurring in the first 6 months and all measures remain lower than recommended for cardiovascular and overall health. A randomized clinical trial of lifestyle modifications such as diet and exercise intervention is warranted to determine the impact of such modifications on HRQoL and fitness post-OLT. PMID- 15659125 TI - Pharmacokinetics of His-tag recombinant human endostatin in Rhesus monkeys. AB - AIM: To study the pharmacokinetics and accumulation of an Escherichia coli expressed His-tag fused recombinant human endostatin (rh-endostatin) in Rhesus monkeys. METHODS: Rh-endostatin was iv or sc injected in Rhesus monkeys, and the rh-endostatin concentration in serum samples was determined by an enzyme immunoassay (EIA) method. The serum drug concentration-time data were analyzed by compartmental analysis using the practical pharmacokinetic program 3p97. RESULTS: Following iv administration at a dose rate of 1.5, 4.5, and 13.5 mg/kg in rhesus monkeys, the concentration-time curves of rh-endostatin were best fitted to a three-compartment open model. AUC(0-infinity) linearly increased with dose, while Cls exhibited no significant difference among different dose groups. The terminal half-lives (lambda3) were 8+/-8, 3.1+/-1.4, and 20+/-14 h, respectively. After sc administration at a dose rate of 1.5 mg/kg, the concentration-time curve was best fitted to a two-compartment open model, with a terminal half-life (T(1/2beta)) of 8+/-3 h. Bioavailability following sc injection was approximately 70%+/-3%. After consecutive iv injection of rh-endostatin at a dose rate of 1.5 mg.kg(-1).d(-1) for 7 d, the AUC(0-24 h) substantially increased from 22+/-13 mg.h.L(-1) (d 1) to 50+/-29 mg.h.L(-1) (d 7), with an accumulation factor of 2.3+/-0.6 (P < 0.05). CONCLUSION: The pharmacokinetic behavior of rh-endostatin in Rhesus monkeys complies with linear kinetics within the examined dose range. It tends to be accumulated in bodies after repeated administration at a dose level of 1.5 mg.kg( 1).d(-1) for more than 7 consecutive days. PMID- 15659128 TI - A survey of dental care protocols among US organ transplant centers. AB - Untreated dental disease represents a potential risk for infection in transplant patients, but the vast transplantation literature has few references to this complication. There is also little information with regard to dental care protocols for patients before and after organ transplantation. To obtain more definitive documentation about the policies that deal with dental care and experience with dental infections, we conducted a survey of US transplant centers. The instrument consisted of eight questions that addressed pre transplant dental evaluation procedures, incidence of pre- and post-transplant dental infections, and recommendations for antibiotic prophylaxis with dental treatment after transplantation. Questionnaires were sent to 768 medical and/or surgical directors at all US transplant centers. Responses were received from 294 recipients (38%). Among the respondents, 80% routinely requested a pre-transplant dental evaluation, but 49% of these were only for specific organs. The occurrence of a dental infection prior to transplantation that resulted in a postponement or cancellation was reported by 38% of the respondents. Post-transplantation sepsis from a suspected dental source was acknowledged in 27% of the surveys. Prophylaxis with antibiotics prior to dental care was recommended by 83%; 77% indicated that it be used for all dental procedures, whether invasive or not. Most respondents (96%) recommended the 1997 American Heart Association endocarditis prevention regimen. A survey of organ transplant centers has provided some information with regard to pre-transplantation dental screening, dental infections, and the use of prophylactic antibiotics. Additional studies are needed in order to accrue more definitive data that will assist with the development of standardized and appropriate pre- and post-transplant dental care protocols. PMID- 15659127 TI - Angiotensinogen and plasminogen activator inhibitor-1 gene polymorphism in relation to chronic allograft dysfunction. AB - Chronic allograft dysfunction (CAD) is the most common cause of allograft failure in the long-term, and current immunologic strategies have little effect on this condition. The renin-angiotensin system (RAS) plays important roles progression of chronic renal disease. It is thought that plasminogen activator inhibitor-1 (PAI-1) functions in the RAS, in addition to involvement in thrombotic risk and fibrosis. This study investigated possible links between angiotensinogen (AGT) genotypes (M235T/MM, MT, TT) and PAI-1 genotypes (4G4G, 4G5G, 5G5G) and CAD assessments of both types of polymorphism were performed in 82 renal allograft recipients. One hundred healthy subjects were also investigated for AGT polymorphism, and 80 healthy subjects for PAI-1 polymorphism. Genotypes were determined using polymerase chain reaction (PCR) sequence-specific primers, and PCR followed by restriction fragment length polymorphism analysis. Kidney recipients with CAD had significantly lower frequencies of the MM genotype and the M allele than the recipients without CAD (p < 0.05 and <0.001). The transplant recipients with CAD also had significantly lower frequencies of the 5G5G genotype and the 5G allele than those without CAD (p < 0.001 and <0.05). Determination of AGT M235T and PAI-1 genotypes prior to transplantation may help identify patients who at risk for chronic renal transplant dysfunction. PMID- 15659129 TI - The need of mycophenolic acid monitoring in long-term renal transplants. AB - BACKGROUND: There is little information regarding the 12-h mycophenolic acid (MPA) pharmacokinetics (PK), a way to monitor the drug and the need of frequent monitoring, in stable patients. METHODS: A cohort of 35 adults, under long-term mycophenolate mofetil (MMF) therapy plus cyclosporin A (n = 12), TACimus (n = 12) or MMF only (n = 11); all with prednisone had a 12-h MPA-PK performed to ascertain the percentage of them within a defined therapeutic window. In 13 other patients, two PK studies undergone 1 wk apart were performed to evaluate the need for frequent measurements. RESULTS: Fourteen (40%) patients were within the defined therapeutic window (36-60 microg h/mL). Nine patients (26%) were overexposed while 12 (34%) were underexposed. A Cmax> or =10 microg/mL was seen in 20 (57%) of the patients. These percentages were equally distributed between the treatment groups both for AUC0-12 and Cmax. The equations using C0, C2 or both predict exposure, although the use of C2 seems to be more adequate in clinical practice. There were no differences in MPA exposure in patients with a repeated PK evaluated 1 wk later. CONCLUSION: The use of MMF without monitoring MPA blood levels may cause over-/underexposure to the drug in stable recipients. However, in patients under MMF for more than 1 yr, MPA levels are stable and there is no need for frequent measurements. PMID- 15659130 TI - A prospective 3-yr evaluation of tacrolimus-based immunosuppressive therapy in immunological high risk renal allograft recipients. AB - BACKGROUND: There have been no published data on use of the the newer immunosuppressants tacrolimus and mycophenolate mofetil (MMF) in high immunological risk renal transplantation. We therefore undertook a prospective study to systematically assess outcomes using these agents as part of an aggressive immunosuppressive regimen. METHODS: Fifty-nine high-risk renal allograft recipients were enrolled at 10 Canadian sites and given a regimen of: a biological induction agent, tacrolimus, MMF, and corticosteroids. Patients included 10 (17%) who had lost a previous graft to rejection <1 yr, 31 (53%) with a current panel reactive antibody (PRA) >30%, 47 (80%) with a historic PRA >50%, four (7%) who had a positive historical T-cell crossmatch with the current donor, and six (10%) with a current positive B-cell crossmatch. The mean peak PRA was 76 +/- 33%. RESULTS: The estimated 3-yr Kaplan-Meier patient and graft survival estimates were 89% and 75%, respectively. There were nine graft losses other than deaths with a functioning graft, of which six were preceded by delayed graft function (p = 0.01, chi2). Sixteen (27%) recipients experienced at least one episode of biopsy-confirmed acute rejection. Infections included cytomegalovirus in 16 patients, eight of whom had tissue-invasive disease. Only one malignancy occurred. CONCLUSIONS: The immunosuppressive strategy investigated is effective and displays a satisfactory safety profile in high immunological risk renal allograft recipients. PMID- 15659131 TI - Organ donation: a population-based study. AB - OBJECTIVE: Through a population-based study, it was identified the prevalence of people's willingness to donate their own organs and from their relatives, evaluating associated factors in an adult population. It was also identified their understanding of cerebral death. METHODOLOGY: Cross-sectional study, with people aged 20 yr or older in the urban area of Pelotas, State of Rio Grande do Sul, Brazil. The instrument used was a structured questionnaire, filled out in individual interviews. Chi-squared and linear trend test were used in the bivariated analysis. Multivariated analysis was conducted according to a hierarchical classification model using Poisson regression. It was considered meaningful the value for p < or = 0.05 two-sided. RESULTS: Amid 3159 participants, the prevalence to donate organs was 52%, amongst which 58% had expressed such willingness to a relative. Most respondents (80.1%) would authorize the donation of relative's organs who had previously declared their willingness to do so. When the subject had not been discussed, only a third of the total number of people interviewed would authorize the donation of a relative's organ. After adjustment to confusing factors, higher willingness was characterized among the youngest, the higher educated and those belonging to families with income over 10 minimum wages. The Evangelical and Jehovah's Witnesses practitioners showed to be less prone to donate. CONCLUSION: According to the study, when the peoples had not enough information regarding family member's donation wishes the rate of willingness to donate organs is lower. Sociodemographic characteristics influence the rate of public willingness to donate organs and campaigns educational should be directed to improve rates of donation the organs. PMID- 15659132 TI - Psychosocial profiling: a holistic management tool for non-compliance. AB - We introduce a new concept of psychosocial profiling as a tool that provides the transplant team with a psychosocial framework for identification, intervention and management of non-compliance. This will also increase our understanding of emotional problems experienced by patients before transplant, as a result of living with the uncertainty and medical side effects of chronic illness. Psychosocial profiling is adaptable throughout the transplant process and gives every patient an opportunity of psychosocial support to help him or her into a position of emotional stability and compliance with their medications and postoperative care. Implementation of this strategy will move health care professionals from being gatekeepers to managers and facilitators of holistic care in recipients of transplants. PMID- 15659133 TI - Incidence of serum creatine kinase elevation and its relation to medications used after heart transplantation. AB - BACKGROUND: Rhabdomyolysis after heart transplantation is well described as a drug-related phenomenon. The incidence of serum creatine kinase (CK) elevation after heart transplantation is not formally reported in previous clinical studies. This study sought to find the incidence of asymptomatic serum CK elevation after heart transplantation and assess its relation to medications that are commonly used after transplantation. METHODS: Data were collected in outpatient basis between August 2002 and August 2003. Patients with acute rejection, infection or muscle pain were excluded. All patients were followed monthly at a special clinic. Physical examinations and routine blood tests including serum CK were performed monthly. We evaluated the results of 106 asymptomatic patients and 765 serial data to determine the incidence of serum CK elevation after transplantation. Logistic regression was used to identify its risk factors. RESULTS: The incidence of serum CK elevation in asymptomatic heart transplant recipients was 16.2%. Risk factors of its elevation were diabetes mellitus and use of medications including cyclosporine, drugs for hypertriglyceridemia, antihypertensives, and prednisolone. Among the antihypertensives commonly used, angiotensin converting enzyme inhibitors, angiotensin receptor blockers and calcium antagonists were associated with a higher incidence of serum CK elevation, but beta-blockers protected patients from serum CK elevation. CONCLUSION: The incidence of serum CK elevation in asymptomatic heart transplant recipients was not low. Serum CK measurements should be part of the routine follow-up in transplant recipients, especially when patients had diabetes mellitus. Medications commonly used after heart transplantation significantly affected the incidence of serum CK elevation. PMID- 15659134 TI - Novel rescue therapy for C4d-positive acute humoral renal allograft rejection. AB - OBJECTIVE: To investigate the efficacy of immunoadsorption (IA) in combination with tacrolimus (TAC) and mycophenolate mofetil (MMF) rescue therapy for C4d positive acute humoral rejection (AHR) of renal transplants. METHODOLOGY: Six of 185 cadaveric renal allograft recipients transplanted at our institute developed AHR over a mean period of 4.8 +/- 0.8 d after operation. The ages ranged from 35 to 51 yr (mean 42.6 +/- 5.6 yr). C4d deposits in peritubular capillaries (PTC) and accumulation of granulocytes in PTC were observed. IA with staphylococcal protein A and TAC-MMF combination therapy were given. RESULTS: After subjected to IA for 6.3 +/- 1.03 sessions combined with TAC (0.14-0.16 mg/kg/d) and MMF (1.5 g/d) therapy, renal function recovered in all the patients. The mean duration of treatment when serum creatinine decreased was 14 +/- 2.9 d. The pre-IA panel reactive antibody reactivity was as high as 50.2 +/- 6.1%, and was significantly reduced to 8.3 +/- 2.9% after IA. Repeated allograft kidney biopsy in four of six patients revealed a favorable remission of AHR. With a mean follow-up of 18.8 +/- 5.46 months, patient and allograft survival are 100%, renal function remained stable with a mean serum creatinine of 1.2 +/- 0.22 mg/dL. CONCLUSION: The optimal treatment for alloantibody-mediated AHR remains undefined. Our findings suggest that a therapeutic approach combining IA and TAC-MMF rescue has excellence to improve the outcome of AHR. PMID- 15659135 TI - Standard chronic immunosuppression after kidney transplantation for systemic lupus erythematosus eliminates recurrence of disease. AB - BACKGROUND: There is only limited experience in patients with systemic lupus erythematosus (SLE) with drugs that have developed for immunosuppression after organ transplantation, namely calcineurin inhibitors (CI). The aim of this study is to determine the effect of these drugs on disease activity after kidney transplant in patients affected by SLE. METHODS: Between January 1990 to March 2003, 13 patients with end- stage renal disease secondary to SLE received 14 kidney transplants. The outcome variables assessed include graft and patient survival as well as clinical and serological lupus activity. RESULTS: All received CI-based immunosuppression (cyclosporine or tacrolimus). Actuarial patient and graft survivals at 5 yr were 100 and 93%, respectively. Recurrence of clinical or serological disease was never detected. CONCLUSIONS: To date, only anecdotal experience with CI in the treatment of SLE has been reported. The favorable response observed in our patients suggests that CI at low-doses are effective in preventing SLE-reactivation. Further studies focused on calcineurin inhibitor treatment in SLE patients who fail to respond to standard medical management should be conducted. PMID- 15659136 TI - Steroid avoidance in renal transplantation using basiliximab induction, cyclosporine-based immunosuppression and protocol biopsies. AB - BACKGROUND: Reducing chronic steroid exposure is important to minimize steroid related morbidity, particularly for susceptible renal transplant recipients. Steroid-free and steroid-sparing protocols have shown benefits, but safety has not been established for all populations. We investigated the safety of steroid avoidance (SA) in a population including African-Americans, using modern immunosuppression with protocol biopsy monitoring. METHODS: A randomized controlled SA trial (early discontinuation, days 2-7) was conducted in a population (n = 77) including African-Americans and cadaveric kidney recipients. Patients received basiliximab, cyclosporine (CsA), and mycophenolate mofetil (MMF). In controls, steroids were tapered to 5 mg prednisone/d by day 30. Protocol biopsies were performed (1, 6, 12 and 24 months) to evaluate subclinical acute rejection (SCAR) and chronic allograft nephropathy (CAN). RESULTS: The SA did not result in significantly higher incidences of graft loss, AR, SCAR, CAN, or renal fibrosis. SA patients experienced similar renal function, comparable serum lipid levels, and a trend toward fewer cases of new-onset diabetes. Clinical outcomes of African-American and non-African-American patients did not significantly differ. CONCLUSIONS: The SA is safe in the context of basiliximab induction and CsA-based immunosuppression. This protocol could minimize steroid related side effects in susceptible groups, including African-Americans, without increasing the risk of AR or graft failure. PMID- 15659137 TI - Factors modifying stress from adverse effects of immunosuppressive medication in kidney transplant recipients. AB - INTRODUCTION: The adverse effects of immunosuppression appear in the majority of patients with a negative impact on morbidity, mortality and quality of life. The group of adverse symptoms manifested as changes in appearance, mood and energy are often more stressful than serious metabolic changes because of their direct negative influence on patients' well-being. The aim of this study is to explore the adverse symptoms of immunosuppressive medication which are the most stressful for transplanted patients, and which are the modifying factors. PATIENTS AND METHODS: A total of 157 adult kidney transplant recipients from two transplant centres in Slovakia with a functioning graft transplanted <7 yr ago were examined. Patients participated in an interview focusing on stress from adverse effects, and their education and social support. Medical records were searched for information about immunosuppression protocols, dialysis treatment before transplantation, type of received organ and period after transplantation. The effect of the selected variables on the total score for stress from adverse effects was tested using ANOVA. The effect of the selected factors on stress from each single adverse effect was explored using t-test and ANOVA. RESULTS: The most stressful symptoms were pain, weakness, weight gain, facial changes, depression and anxiety. The mean value of the total score for stress from adverse effects was 8.03 +/- 6.53 (minimum 0, maximum 30, range: 0-64), indicating low stress. Women and patients with lower education significantly more often felt the adverse effects of immunosuppression as stressful (p < 0.001 and p < 0.05, respectively). Age, social support, dialysis modality before transplantation, time from transplantation and type of immunosuppressive treatment did not affect the total score for stress from adverse effects. However, variables that were not significant in the overall score reached significance in some symptoms. CONCLUSIONS: Women and patients with lower education significantly more often felt the adverse effects of immunosuppression as stressful; in a more detailed analysis the use of new drugs was connected with less stress in some symptoms. The use of these drugs can improve life quality for transplant recipients, decrease non-compliance, and thus prevent graft loss. PMID- 15659138 TI - Comparing renal transplant patients' adherence to free cyclosporine and free tacrolimus immunosuppressant therapy. AB - BACKGROUND: The purpose of this study is to determine if there is a difference in renal transplant patients' (RTPs) adherence to cyclosporine compared to tacrolimus when medications are supplied free to the RTPs. METHODS: Adult primary RTPs were included in the study if they received a renal transplant at the Medical College of Georgia (MCG) from June 1998 through August 2001 and received their first post-transplant year of follow-up care at MCG and free cyclosporine or free tacrolimus from the MCG outpatient pharmacy. Adherence was estimated by comparing each RTPs' tacrolimus or cyclosporine pharmacy refill records to the prescribed regimen for 12 months after transplant. Patients' cyclosporine and tacrolimus serum concentrations were used to validate adherence. Kaplan-Meier analysis was used to estimate the fraction of RTPs remaining adherent and to compare the mean time RTPs were adherent in each group (cyclosporine vs. tacrolimus). RESULTS: Thirty-three RTPs were included in the study, 25 (76%) received cyclosporine and eight received tacrolimus. The mean time to the first non-adherent month was 8 months post-transplant. At 12-months post-transplant, approximately 42% of the patients remained adherent. A greater percentage of the patients who received tacrolimus remained adherent compared with those who were taking cyclosporine (63% vs. 33%, p < 0.05). Approximately 75% of non-adherent patients were found to have subtarget drug concentrations, and only 24% of adherent patients had subtarget levels (p < 0.01). CONCLUSIONS: When provided free, patients are more adherent to tacrolimus than cyclosporine. Regardless of treatment, intensive efforts to increase adherence should be implemented. PMID- 15659139 TI - Induction therapy with either anti-CD25 monoclonal antibodies or rabbit antithymocyte globulins in liver transplantation for hepatitis C. AB - BACKGROUND: Hepatitis C virus (HCV)-related end-stage liver disease (ESLD) is the leading cause for orthotopic liver transplantation (OLT). The aim of our sequential study was to assess the safety and efficacy of induction therapy using either rabbit antithymocyte globulins (RATG) or anti-CD25 monoclonal antibodies. METHODS: From January 2000 to January 2003, 31 patients underwent OLT for HCV related ESLD, and survived more than 1 month post-transplantation. Up to July 2001, induction relied on RATG (Thymoglobulins; Sangstat, Lyon, France) which was given for a median of 5 d, i.e. a total dose of 406 +/- 45 mg (n = 16; group I). Thereafter, induction was based on either basiliximab or dacluzimab (n = 15; group II). Steroids and tacrolimus (from day 1 onwards) were given as maintenance therapy. The primary end-points were at 6 months post-transplantation. RESULTS: Patient and graft survivals were excellent and similar in both groups. The acute rejection rate was 37.5% in group I and 20% in group II (p = ns). The rates of serious bacterial and fungal infections as well as cytomegalovirus infections were similar in both groups. Even if the overall pattern of post-transplant liver enzymes did not differ between the two groups, the rate of HCV recurrence tended to be higher in group II (80%) compared with group I (56.25%; p = ns) patients. Both induction therapies were clinically and biologically well tolerated. CONCLUSION: RATG induction therapy is as efficient and as safe as induction with anti-CD25 monoclonal antibodies. PMID- 15659140 TI - Living donors' experiences 1 wk after donating a kidney. AB - In Norway living kidney donors account for approximately 40% of all renal transplants. There is a shortage of information about how living kidney donors experience the donation process during the initial recovery from surgery. The aim of this study was to explore physical and psychosocial issues related to the experiences of living kidney donors 1 wk after open donor nephrectomy. A total of 12 living kidney donors participated in the study. Data were collected by individual in-depth interviews and analysed using empirical phenomenological method. Being a living kidney donor is a complex experience. The informants expressed strong feelings of responsibility and obligation concerning the recipients and had a positive attitude towards the donation. On the other hand, the donors experienced it strange to be a fit individual and go through a major operation. Several of the donors reported that it was painful to go through donor surgery and regarded the recovery period as troublesome. Some donors also reported the double role of being both a patient and a relative to be a stressful experience. This study reveals the importance of being aware of the complex situation of living kidney donors. Health professionals need to understand the nature of the donation process and the donors' needs. It is essential to focus on physical, mental and interpersonal factors when counselling potential living kidney donors. PMID- 15659141 TI - Prevalence, characteristics, and outcome of BK virus nephropathy in Japanese renal transplant patients: analysis in protocol and episode biopsies. AB - BACKGROUND: BK virus nephropathy (BKN) is recognized as a cause of graft loss in renal transplant patients. This may be related to the introduction of new and potent immunosuppressive regimens. In Japan, our experience regarding its prevalence, clinical significance, and outcome is still limited. In this study, our primary purpose is to outline the prevalence, outcome, and clinical characteristics of BKN as observed at Osaka University Hospital. METHODS: We retrospectively analyzed 112 biopsy specimens from 87 renal transplant patients. All transplantations were from living donors. Of the 112 biopsy specimens, 71 were from protocol biopsies and 41 were from episode biopsies. Calcineurin inhibitors and corticosteroid were used in all patients (tacrolimus 32 and cyclosporin 55). In addition, azathioprine was used in 43 patients, mizoribine was used in 24 patients, and mycophenolate mofetil was used in 20 patients. BKN was diagnosed by light microscopic examination and a positive immunohistochemical staining of anti-SV40 antibody in a biopsy specimen. In order to investigate the outcome and potential risk factors of patients with different histological staging, we divided the patients into groups A (mild histological change) and B (moderate or severe histological change). RESULTS: Of the 87 patients, six were diagnosed with BKN. There were no significant differences between BKN patients and non-BKN patients, except for the number of patients with graft loss (p < 0.001). Of the six BKN patients, three were in group A, and three were in group B. We recognized a significant difference between group A and group B in terms of anti-rejection treatment including glucocorticoid, tacrolimus trough levels of over 8 ng/mL, episode of acute rejection within 1-month post-transplantation, and the time period between transplantation and BKN diagnosis. CONCLUSIONS: This is the first report of BKN in Japanese renal allograft recipients. In our hospital, the prevalence, risk factors, and outcome were similar to those previously for non-Japanese recipients. PMID- 15659142 TI - Laparoscopic donor nephrectomy vs. open live donor nephrectomy: a quality of life and functional study. AB - BACKGROUND: Few studies have compared the quality of life (QoL) and functional recuperation of laproscopic donor nephrectomy (LDN) vs. open donor nephrectomy (ODN) donors. This study utilized the SF-36 health survey, single-item health related quality of life (HRQOL) score, and a functional assessment questionnaire ('Donor Survey'). METHODS: Questionnaires were sent to 100 LDN and 50 ODN donors. These donors were patients whose procedures were performed at The University Hospital and The Christ Hospital in Cincinnati, Ohio. RESULTS: A total of 46 (46%) LDN and 21 (42%) ODN donors returned the completed surveys. The demographics of the two groups were similar. LDN patients reported a more rapid return to 100% normal health (69 vs. 116 d; p = 0.24), part-time work (21.9 vs. 23.2 d; p = 0.09), and necessitated fewer physician office visits post-operative (2.8 vs. 4.4; p = 0.01). ODN patients reported shorter duration of oral pain medication use (13.4 vs. 7.2 d; p = 0.02). However, a greater number of ODN patients reported post-surgical chronic pain (3 vs. 6; p < 0.05) and hernia (0 vs. 2; p = 0.19). The overall QoL for both groups was comparable with the general USA population. CONCLUSIONS: The results of this study support the decisions of many kidney transplant centers to adopt LDN programs as standard of care. PMID- 15659143 TI - Lobular damage caused by cellular and humoral immunity in liver allograft rejection. AB - BACKGROUND: Diagnosis of acute liver allograft rejection (ALAR) is usually performed by the estimation of changes in portal areas. In this study, changes in the hepatic lobules were investigated retrospectively by immunohistochemistry, and compared with changes in the portal areas. Humoral immunity in ALAR was also studied by C4d-staining. MATERIALS AND METHODS: In total, 35 biopsy specimens from 20 patients who had undergone living-related liver transplantation were included. Specimens had been graded as mild, moderate-to-severe acute rejection based on the Banff Schema. Changes in hepatic lobules were investigated by hematoxylin-eosin (H & E) staining, an immunohistochemical study using anti-CD3 (T cells), anti-GMP-17 (cytotoxic cells), anti-CD68 (macrophages) and C4d, and the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end labeling (TUNEL) method. RESULTS: Changes in hepatic lobules consisted of the infiltration of mononuclear cells, swelling and necrosis of hepatocytes, and hemorrhages. The degree of these changes increased with the severity of ALAR. Immunohistochemical analysis revealed that infiltrating cells included CD3+ T cells, GMP-17+ cytotoxic cells and CD68+ macrophages. The number of infiltrating cells increased with the severity of the ALAR. The TUNEL assay demonstrated apoptotic cells in ALAR and the number of apoptotic cells increased with the severity of the ALAR. In moderate-to-severe rejection, C4d depositions were observed in the hepatic sinusoids as well as the portal veins and hepatic arteries. CONCLUSIONS: Changes in the hepatic lobules were observed in ALAR and the severity increased with the severity of ALAR. Apoptosis was involved in the mechanism of cell death and humoral immunity plays a role in the mechanism of ALAR. PMID- 15659144 TI - Acitretin and skin cancer in kidney transplanted patients. Clinical and histological evaluation and immunohistochemical analysis of lymphocytes, natural killer cells and Langerhans' cells in sun exposed and sun protected skin. AB - BACKGROUND: Renal transplanted recipients have an increased incidence of actinic keratosis and skin cancer. METHODS: In order to examine the chemoprophylatic effects of low-dose acitretin on keratosis and skin cancer development we submitted 13 renal transplanted patients who presented actinic keratosis to acitretin therapy (20 mg/d) for 12 months. The patients were assessed at monthly intervals during the first 6 months and every 2 months until the 12th month for new skin lesions and for acitretin toxicity. Normal skin biopsies of sun exposed and sun protected areas were taken for histopathological examination and submitted to immunohistochemistry technique to demonstrate CD4+ and CD8+ T lymphocytes, natural killer (NK) cells and Langerhans' cells which were counted and compared before, after 6 and 12 months of the treatment. RESULTS: There was an improvement of actinic keratosis in all patients. Only one patient developed new skin cancer. Side-effects were well tolerated and no significant biochemical effects were observed. There were no differences in the microscopic aspects of the skin and in the number of CD4+ and CD8+ T lymphocytes and NK cells. There was a significant increase in the number of epidermal Langerhans' cells after 12 months of acitretin therapy. CONCLUSIONS: The data obtained permit us to conclude that low dose acitretin therapy is safe, well tolerated and partially effective in chemoprophylaxis of skin cancer in renal transplant recipients. The increase in epidermal Langerhans' cells observed may be an expression of the immunomodulatory effect of acitretin. PMID- 15659145 TI - Economic evaluation of everolimus vs. azathioprine at one year after de novo heart transplantation. AB - BACKGROUND: Everolimus decreases acute rejection and cardiac allograft vasculopathy after heart transplantation. We compared within-trial costs and resource use over 1 yr of follow-up in de novo heart transplant patients randomized to everolimus 1.5 mg/d (n = 209), everolimus 3.0 mg/d (n = 211), or azathioprine (n = 214). PATIENTS AND METHODS: Resource use data were collected prospectively for 634 patients from 14 countries. We used the nonparametric bootstrap method to test for differences in mean costs and to estimate confidence intervals for cost-effectiveness ratios. RESULTS: Everolimus patients had lower incidence of efficacy failure compared with azathioprine patients (41.6%, everolimus 1.5 mg; 32.2%, everolimus 3.0 mg; 52.8%, azathioprine). Compared with patients receiving azathioprine, everolimus patients spent more days in the hospital [36.3 d for everolimus 1.5 mg/d (p = 0.21); 38.4 d for everolimus 3.0 mg/d (p = 0.01); 32.2 d for azathioprine]. Mean total costs, excluding the study medications, were not significantly different among treatment groups ($72 065 for everolimus 1.5 mg; $72 631 for everolimus 3.0 mg; $70 815 for azathioprine). CONCLUSIONS: Over 1 yr of follow-up after heart transplantation, everolimus did not significantly increase treatment costs, excluding the costs of the study medications, while reducing efficacy failure. Longer follow-up and the cost of everolimus are required to fully evaluate the cost-effectiveness of everolimus vs. azathioprine in post-transplant maintenance. PMID- 15659146 TI - An open-label, pilot study evaluating the safety and efficacy of converting from calcineurin inhibitors to sirolimus in established renal allograft recipients with moderate renal insufficiency. AB - This pilot study was designed to evaluate the safety and efficacy of converting from a calcineurin inhibitor (CI) to a sirolimus (SRL)-based regimen in established renal transplant recipients with moderate renal insufficiency. Sixty renal transplant recipients on CI-based immuno-suppression with a serum creatinine (SCr) between 159 and 265 microM (1.8 and 3.0 mg/dL) and a glomerular filtration rate (GFR) between 30 and 70 mL/min were enrolled. SRL dosing was dependent upon concomitant immunosuppressive therapy. The mean patient age was 45 yr and the mean time from transplant to study enrollment was 60.8 months (range: 7-198). The median SCr was 168 microM (1.9 mg/dL) and the median GFR was 51 mL/min. Twelve months after conversion the patient and graft survival rates were 96.7% and 95%, respectively. The incidence of biopsy-proven acute rejection was 3.3% (two cases reported, Banff grades IA and IB). The median SCr and median creatinine clearance were 168 microM (1.9 mg/dL) and 53 mL/min, respectively. Hyperlipidemia, diarrhea, peripheral edema, rash, and anemia were the most commonly reported adverse events. Patients with moderate renal insufficiency can be converted from CI to SRL-based therapy and maintain renal function over a 1-yr period. PMID- 15659147 TI - Refractory acute kidney transplant rejection with CD20 graft infiltrates and successful therapy with rituximab. AB - Acute rejection is an expected event after transplantation and has been associated with poor long-term kidney transplant outcome. The presence of B cells in the kidney graft with acute rejection is thought to be an omnious sign, as it has been associated with poor graft outcome. There is no definitive treatment for acute rejection with B cells in the graft. Rituximab, a humanized monoclonal antibody against CD20, has been used in the treatment of B cell lymphoma. We present the case of a 49-yr-old Caucasian male with early acute kidney allograft rejection that was refractory to high doses of steroids and rabbit anti-thymocyte globulin (thymoglobulin). Repeat renal biopsy revealed T cell and B cells in the kidney graft and responded to the combination of rituximab and muromonab (a mouse monoclonal antibody to CD3 receptor). Over 9 months post-transplant, the patient remains rejection free with a serum creatinine of 1.7 mg/dL. PMID- 15659148 TI - Nephrotic syndrome after stem cell transplantation. AB - Nephrotic syndrome occurs rarely after bone marrow transplantation. We describe three patients with myeloid malignancy who developed nephrotic syndrome from 5, 22 and 25 months after allogeneic stem cell transplantation (SCT), confirmed by electron microscopy as membranous glomerulonephritis in two and minimal change glomerulonephritis in one. Proteinuria was initially severe in all and clinically distinct from prior graft-vs.-host disease in two patients. While all responded initially to prednisolone and cyclosporine therapy, two recipients with high-risk leukemia developed late solid organ and bone marrow relapse of their disease, which ultimately proved fatal. The third patient remains alive and disease-free with minimal proteinuria off immunosuppressive therapy. Hence, the onset of de novo high-grade proteinuria after allogeneic SCT should prompt renal histological confirmation, and a trial of immunosuppressive therapy after other causes of nephritic syndrome have been excluded. PMID- 15659150 TI - Influence of exposure to electromagnetic field on the cardiovascular system. AB - 1 We examined whether extremely low frequency electromagnetic fields (ELF-EMF) affect the basal level of cardiovascular parameters and influence of drugs acting on the sympathetic nervous system. 2 Male rats were exposed to sham control and EMF (60 Hz, 20 G) for 1 (MF-1) or 5 days (MF-5). We evaluated the alterations of blood pressure (BP), pulse pressure (PP), heart rate (HR), and the PR interval, QRS interval and QT interval on the electrocardiogram and dysrhythmic ratio in basal level and dysrhythmia induced by beta-adrenoceptor agonists. 3 In terms of the basal levels, there were no statistically significant differences among control, MF-1 and MF-5 in PR interval, QRS interval, mean BP, HR and PP. However, the QT interval, representing ventricular repolarization, was significantly reduced by MF-1 (P < 0.05). 4 (-)-Dobutamine (beta1-adrenoceptor-selective agonist)-induced tachycardia was significantly suppressed by ELF-EMF exposure in MF-1 for the increase in HR (DeltaHR), the decrease in QRS interval (DeltaQRS) and the decrease in QT (DeltaQT) interval. Adrenaline (nonselective beta-receptor agonist)-induced dysrhythmia was also significantly suppressed by ELF-EMF in MF-1 for the number of missing beats, the dysrhythmic ratio, and the increase in BP and PP. 5 These results indicated that 1-day exposure to ELF-EMF (60 Hz, 20 G) could suppress the increase in HR by affecting ventricular repolarization and may have a down-regulatory effect on responses of the cardiovascular system induced by sympathetic agonists. PMID- 15659149 TI - Fish somatostatin sst3 receptor: comparison of radioligand and GTPgammaS binding, adenylate cyclase and phospholipase C activities reveals different agonist dependent pharmacological signatures. AB - 1 The fish somatostatin receptor 3 (fsst3) is one of the few somatostatin (SRIF) receptors cloned from a non-mammalian species so far. Here we extended our earlier characterization of this receptor by investigating the guanine nucleotide sensitivity of agonist radioligand binding at the fsst3 receptor recombinantly expressed in CCL39 (Chinese hamster lung fibroblast) cells. Further, we measured somatostatin (SRIF) and cortistatin (CST) analogues stimulated GTPgammaS binding, inhibition of forskolin-stimulated adenylate cyclase (FSAC) and stimulation of phospholipase C (PLC) activities. The present transductional data were then compared with previous radioligand binding and/or second messenger features determined for fsst3 and/or human SRIF receptors (hsst2, hsst3 and hsst5). 2 The GTP analogue guanylylimidodiphosphate (GppNHp) inhibited binding of [125I]CGP 23996 and [125I][Tyr3octreotide by 72 and 83% suggesting preferential labelling of G-protein-coupled fsst3 receptors. By contrast, [125I]LTT-SRIF28 and [125I][Tyr10]CST14 binding was rather GppNHp insensitive (42 and 35% inhibition) suggesting labelling of both coupled and non-coupled receptor states. These results might explain the apparent higher receptor densities determined in saturation experiments with [125I]LTT-SRIF28 and [125I][Tyr10]CST14 (4470 and 4030 fmol mg(-1)) compared with [125I]CGP 23996 and [125I][Tyr3]octreotide (3420 and 1520 fmol mg(-1)). 3 SRIF14 (10 microm)-stimulated specific [35S]GTPgammaS binding by three-fold; SRIF28 and octreotide displayed full agonism, whereas most other ligands displayed 60-80% intrinsic activity compared with SRIF14. SRIF14 and SRIF28 inhibited forskolin-stimulated AC (FSAC) activity by 60%; all tested ligands except BIM 23056 inhibited FSAC with comparable high intrinsic activities. SRIF14 stimulated PLC activity five- to six-fold, as determined by measuring total [3H] IP(x) accumulation; it was rather insensitive to pertussis toxin (PTX, 100 ng ml(-1), 21% inhibition), which suggests the G(q)-family proteins couple to PLC activity. SRIF14, SRIF28 and [Tyr10]CST14 showed full agonism at PLC, whereas all other ligands behaved as partial agonists (20-70% intrinsic activity). BIM 23056, which showed weak partial or no agonism, antagonized SRIF14-induced total [3H]-IP(x) production (pK(B) = 6.83), but failed to block competitively agonist-stimulated [35S]GTPgammaS binding or agonist induced inhibition of FSAC activity. 4 Comparison of the pharmacological profiles of fsst3 receptors established in GTPgammaS binding, FSAC inhibition and PLC stimulation resulted in low correlations (r = 0.410-0.594). Both rank orders of potency and rank orders of relative efficacy varied in the three second messenger experiments. Significant, although variable correlations were obtained comparing GTPgammaS binding and inhibition of FSAC activity with previously reported affinity profiles of [125I]LTT-SRIF28, [125I][Tyr10]CST14, [125I]CGP 23996, [125I][Tyr3]octreotide (r = 0.75-0.83; 0.68-0.89). By contrast, the PLC stimulation and radioligand-binding profiles did not correlate. 5 Comparison of the functional data (GTPgammaS binding, FSAC inhibition, PLC stimulation) of fsst3 receptors with those of human sst2, sst3, sst5 receptors expressed in CCL39 cells resulted in highest correlation with the hsst5 receptor (r = 0.94, 0.97, 0.49) > hsst2 (0.80, 0.50, n.d.) > hsst3 (0.25, 0.19, 0.17). 6 In summary, fsst3 receptors expressed in CCL39 cells are involved in signalling cascades similar to those reported for mammalian SRIF receptors, suggesting SRIF receptors to be highly conserved in evolution. Binding and functional data showed highest similarity of fsst3 receptors with the human sst5 receptor subtype. Different affinities, receptor densities and GppNHp-sensitivities determined with the four radioligands (agonists) are assumed to results from ligand-specific states of the fsst3-ligand complex. The differences in the rank orders of potency and relative efficacy in the various signalling cascades may be explained by agonist-induced receptor trafficking. PMID- 15659151 TI - Is cyclic AMP formation desensitized in patients with end-stage renal failure? AB - 1 Cyclic AMP formation has consistently been reported to be desensitized in various tissues including heart of animal models of end-stage renal failure (ESRF). In contrast, reports on desensitization of cAMP formation in ESRF patients remain contradictory. Whether this discrepancy results from a difference between human ESRF and its animal models or from the use of circulating blood cells in the human and various solid tissues in the animal studies, remains unclear. Therefore, we performed three studies with heart and platelets of ESRF patients undergoing haemodialysis or continuous ambulatory peritoneal dialysis and age- and gender-matched controls with normal renal function (n = 11-13 each). 2 In platelets from haemodialysis patients adenylyl cyclase activity in response to receptor-dependent and -independent agonists was reduced by approximately 30%, and this could be explained by an alteration at the level of adenylyl cyclase itself. However, no such desensitization was seen in platelets from peritoneal dialysis patients. 3 In hearts from ESRF patients undergoing haemodialysis, beta adrenoceptor density and subtype distribution, cAMP formation in response to the beta-adrenoceptor agonist isoprenaline or various receptor-independent stimuli, were very similar to those in control patients but activity of G-protein-coupled receptor kinase was increased by approximately 20%. 4 We conclude that conflicting reports on the desensitization of cAMP formation between ESRF patients and ESRF animal models are not explained by the use of solid tissues in animal studies vs. circulating blood cells in patient studies. Rather desensitization of cAMP formation seems to be a less consistent feature of human ESRF than of its animal models. PMID- 15659153 TI - Raymond Dedonder 1920-2004. PMID- 15659154 TI - From fundamental studies of sporulation to applied spore research. AB - Sporulation in the Gram-positive bacterium, Bacillus subtilis, has been used as an excellent model system to study cell differentiation for almost half a century. This research has given us a detailed picture of the genetic, physiological and biochemical mechanisms that allow bacteria to survive harsh environmental conditions by forming highly robust spores. Although many basic aspects of this process are now understood in great detail, including the crystal and NMR structures of some of the key proteins and their complexes, bacterial sporulation still continues to be a highly attractive model for studying various cell processes at a molecular level. There are several reasons for such scientific interest. First, some of the complex steps in sporulation are not fully understood and/or are only described by 'controversial' models. Second, intensive research on unicellular development of a single microorganism, B. subtilis, left us largely unaware of the multitude of diverse sporulation mechanisms in many other Gram-positive endospore and exospore formers. This diversity would likely be increased if we were to include sporulation processes in the Gram-negative spore formers. Spore formers have great potential in applied research. They have been used for many years as biodosimeters and as natural insecticides, exploited in the industrial production of enzymes, antibiotics, used as probiotics and, more, exploited as possible vectors for drug delivery, vaccine antigens and other immunomodulating molecules. This report describes these and other aspects of current fundamental and applied spore research that were presented at European Spores Conference held in Smolenice Castle, Slovakia, June 2004. PMID- 15659155 TI - Recoding in archaea. AB - Standard decoding of the genetic information into polypeptides is performed by one of the most sophisticated cell machineries, the translating ribosome, which, by following the genetic code, ensures the correspondence between the mature mRNA and the protein sequence. However, the expression of a minority of genes requires programmed deviations from the standard decoding rules, globally named recoding. This includes ribosome programmed -/+1 frameshifting, ribosome hopping, and stop codon readthrough. Recoding in Archaea was unequivocally demonstrated only for the translation of the UGA stop codon into the amino acid selenocysteine. However, a new recoding event leading to the 22nd amino acid pyrrolysine and the preliminary reports on a gene regulated by programmed -1 frameshifting have been recently described in Archaea. Therefore, it appears that the study of this phenomenon in Archaea is still at its dawn and that most of the genes whose expression is regulated by recoding are still uncharacterized. PMID- 15659156 TI - Cell division protein DivIB influences the Spo0J/Soj system of chromosome segregation in Bacillus subtilis. AB - The initiation of the developmental process of sporulation in the rod-shaped bacterium Bacillus subtilis involves the activation of the Spo0A response regulator. Spo0A then drives the switch in the site of division septum formation from midcell to a polar position. Activated Spo0A is required for the transcription of key sporulation loci such as spoIIG, which are negatively regulated by the Soj protein. The transcriptional repressing activity of Soj is antagonized by Spo0J, and both proteins belong to the well-conserved Par family of partitioning proteins. Soj has been shown to jump from nucleoid to nucleoid via the cell pole. The dynamic behaviour of Soj is somehow controlled by Spo0J, which prevents the static association of Soj with the nucleoid, and presumably its transcriptional repression activity. Soj in turn is required for the proper condensation of Spo0J foci around the oriC region. The asymmetric partitioning of the sporangial cell requires DivIB and other proteins involved in vegetative (medial) division. We describe an allele of the cell division gene divIB (divIB80) that reduces the cellular levels of DivIB, and affects nucleoid structure and segregation in growing cells, yet has no major impact on cell division. In divIB80 cells Spo0J foci are not correctly condensed and Soj associates statically with the nucleoid. The divIB80 allele prevents transcription of spoIIG, and arrests sporulation prior to the formation of the asymmetric division septum. The defect in Spo0A-dependent gene expression, and the Spo- phenotype can be suppressed by expression of divIB in trans or by deletion of the soj-spo0J locus. However, deletion of the spo0J-soj region does not restore the normal cellular levels of DivIB. Therefore, the reduced levels of DivIB in the divIB80 mutant are sufficient for efficient cell division, but not to sustain a second, earlier function of DivIB related to the activity of the Spo0J/Soj system of chromosome segregation. PMID- 15659157 TI - A novel two-component system controls the expression of Pseudomonas aeruginosa fimbrial cup genes. AB - Biofilm formation by the opportunistic pathogen Pseudomonas aeruginosa requires the expression of a number of surface adhesive components. The expression of surface organelles facilitating biofilm formation is controlled by environmental signals acting through transcriptional regulatory networks. We analysed the expression of a family of P. aeruginosa adhesins encoded by three distinct fimbrial gene clusters (cupA, cupB and cupC). Using transposon mutagenesis, we have identified several regulatory loci that upregulated cupB and cupC transcription. One such locus contains three components, RocS1, RocR and RocA1, which represent a variant of a classical two-component signal transduction pathway. RocS1 is a sensor kinase, RocA1 is a DNA binding response regulator that activates cup genes, and RocR is an antagonist of RocA1 activity. Using a two hybrid assay, we have shown that RocS1 interacts with receiver domains of both RocA1 and RocR. Expression of the Cup system in response to environmental stimuli is accomplished by a novel mechanism in which the sensor kinase activates its cognate response regulator through a phosphorelay pathway, while an additional repressor protein modulates this interaction. PMID- 15659158 TI - Candida albicans protein kinase CaHsl1p regulates cell elongation and virulence. AB - Saccharomyces cerevisiae Hsl1p is a Ser/Thr protein kinase that regulates cell morphology. We identified Candida albicans CaHSL1 and analysed its function in C. albicans. Cells lacking CaHsl1p exhibited filamentous growth under yeast growth conditions with the filaments elongating more quickly than did those of the wild type under hyphal growth conditions, suggesting that it plays a role in the suppression of cell elongation. Green fluorescent protein-tagged CaHsl1p colocalized with a septin complex to the bud neck during yeast growth or to a potent septation site during hyphal growth, as expected from the localization in S. cerevisiae. However, the localization of the septin complex did not change in DeltaCahsl1, suggesting that CaHsl1p does not participate in septin organization. CaHsl1p was expressed in a cell cycle-dependent manner and, except for the G1 phase, phosphorylated throughout the cell cycle. In DeltaCahsl1 cells, the phosphorylation of a possible CaHsl1p target CaSwe1p decreased, while that of CaCdc28p at tyrosine18 increased. Either an extra copy of the tyrosine18-mutated CaCdc28p or deletion of CaSWE1 suppressed the cell elongation phenotype caused by CaHSL1 deletion. Furthermore, DeltaCahsl1 exhibited reduced virulence in the mouse systemic candidiasis model. Thus, the CaHsl1p-CaSwe1p-CaCdc28p pathway appears important in the cell elongation of both the yeast and hyphal forms and to the virulence of C. albicans. PMID- 15659159 TI - Natural merodiploidy involving duplicated rpoB alleles affects secondary metabolism in a producer actinomycete. AB - Actinomadura sp. ATCC 39727 produces the glycopeptide antibiotic A40926, structurally similar to teicoplanin. Production of A40926 is governed by the stringent response at the transcriptional level. In fact, addition of an amino acid pool prevented the transcription of dbv cluster genes involved in the A40926 biosynthesis and the antibiotic production in chemically defined media, and a thiostrepton-resistant relaxed mutant was severely impaired in its ability to produce the antibiotic. The derivative strain rif19, highly resistant to rifampicin (minimal inhibitory concentration, MIC > 200 microg ml(-1)), was isolated from the wild type strain that exhibited low resistance to rifampicin (MIC < 25 microg ml(-1)). In this strain A40926 production started earlier than in the wild type, and reached higher final levels. Moreover, the antibiotic production was not subjected to the stringent control. Molecular analysis led to the identification of two distinct rpoB alleles, rpoBS and rpoBR, in both the wild type and the rif19. rpoBR harboured the H426N missense which is responsible for rifampicin-resistance in bacteria, in addition to other nucleotide substitutions affecting the primary structure of the RNA polymerase beta-chain. Transcript analysis revealed that rpoBR was expressed at a very low level in the wild type strain during the pseudo-exponential growth phase, and that the amount of rpoBR mRNA increased during the transition to the stationary phase. In contrast, expression of rpoBR was constitutive in the rif19. The results of mRNA half-life analysis did not support the hypothesis that post-transcriptional events are responsible for the different rpoB expression patterns in the two strains, suggesting a role of transcriptional mechanisms. PMID- 15659160 TI - In vitro reconstitution of a trimeric complex of DivIB, DivIC and FtsL, and their transient co-localization at the division site in Streptococcus pneumoniae. AB - DivIB, DivIC and FtsL are bacterial proteins essential for cell division, which show interdependencies for their stabilities and localization. We have reconstituted in vitro a trimeric complex consisting of the recombinant extracellular domains of the three proteins from Streptococcus pneumoniae. The extracellular domain of DivIB was found to associate with a heterodimer of those of DivIC and FtsL. The heterodimerization of DivIC and FtsL was artificially constrained by fusion with interacting coiled-coils. Immunofluorescence experiments showed that DivIC is always localized at mid-cell, in contrast to DivIB and FtsL, which are co-localized with DivIC only during septation. Taken together, our data suggest that assembly of the trimeric complex DivIB/DivIC/FtsL is regulated during the cell cycle through controlled formation of the DivIC/FtsL heterodimer. PMID- 15659161 TI - Role of Mg2+ and pH in the modification of Salmonella lipid A after endocytosis by macrophage tumour cells. AB - Lipid A of Salmonella typhimurium is covalently modified with additional acyl and/or polar substituents in response to activation of the PhoP/PhoQ and/or PmrA/PmrB signalling systems, which are induced by growth at low Mg2+ concentrations and mild acid pH respectively. Although these conditions are thought to exist within macrophage phagolysosomes, no direct evidence for lipid A modification after endocytosis has been presented. To address this issue, we grew S. typhimurium inside RAW264.7 cells in the presence of 32Pi, and then isolated the labelled lipid A fraction, which was found to be extensively derivatized with phosphoethanolamine, aminoarabinose, 2-hydroxymyristate and/or palmitate moieties. S. typhimurium grown in tissue culture medium synthesized lipid A molecules lacking all these substituents with the exception of the 2 hydroxymyristate chain, which was still present. Using defined minimal media to simulate the intracellular pH and Mg2+ concentrations of endosomes, we found that lipid A of S. typhimurium grown in an acidic, low-Mg2+ medium closely resembled lipid A isolated from bacteria internalized by RAW264.7 cells. A subset of S. typhimurium lipid A modifications were induced by low Mg2+ alone. Escherichia coli K-12 W3110 modified its lipid A molecules in response to growth under acidic but not low-Mg2+ conditions. Growth in a high-Mg2+, mildly alkaline medium resulted in suppression of most lipid A modifications with the exception of the 2 hydroxymyristate in S. typhimurium. Although lpxO transcription was stimulated by growth on low Mg2+, the biosynthesis of lipid A species containing 2 hydroxymyristate was independent of PhoP/PhoQ and PmrA/PmrB in S. typhimurium. Our labelling methods should be applicable to studies of lipid A modifications induced by endocytosis of diverse bacteria. PMID- 15659162 TI - Receptor binding studies disclose a novel class of high-affinity inhibitors of the Escherichia coli FimH adhesin. AB - Mannose-binding type 1 pili are important virulence factors for the establishment of Escherichia coli urinary tract infections (UTIs). These infections are initiated by adhesion of uropathogenic E. coli to uroplakin receptors in the uroepithelium via the FimH adhesin located at the tips of type 1 pili. Blocking of bacterial adhesion is able to prevent infection. Here, we provide for the first time binding data of the molecular events underlying type 1 fimbrial adherence, by crystallographic analyses of the FimH receptor binding domains from a uropathogenic and a K-12 strain, and affinity measurements with mannose, common mono- and disaccharides, and a series of alkyl and aryl mannosides. Our results illustrate that the lectin domain of the FimH adhesin is a stable and functional entity and that an exogenous butyl alpha-D-mannoside, bound in the crystal structures, exhibits a significantly better affinity for FimH (Kd = 0.15 microM) than mannose (Kd = 2.3 microM). Exploration of the binding affinities of alpha- d mannosides with longer alkyl tails revealed affinities up to 5 nM. Aryl mannosides and fructose can also bind with high affinities to the FimH lectin domain, with a 100-fold improvement and 15-fold reduction in affinity, respectively, compared with mannose. Taken together, these relative FimH affinities correlate exceptionally well with the relative concentrations of the same glycans needed for the inhibition of adherence of type 1 piliated E. coli. We foresee that our findings will spark new ideas and initiatives for the development of UTI vaccines and anti-adhesive drugs to prevent anticipated and recurrent UTIs. PMID- 15659164 TI - Identification of novel non-coding RNAs as potential antisense regulators in the archaeon Sulfolobus solfataricus. AB - By generating a specialized cDNA library from the archaeon Sulfolobus solfataricus, we have identified 57 novel small non-coding RNA (ncRNA) candidates and confirmed their expression by Northern blot analysis. The majority was found to belong to one of two classes, either antisense or antisense-box RNAs, where the latter only exhibit partial complementarity to RNA targets. The most prominent group of antisense RNAs is transcribed in the opposite orientation to the transposase genes, encoded by insertion elements (transposons). Thus, these antisense RNAs may regulate transposition of insertion elements by inhibiting expression of the transposase mRNA. Surprisingly, the class of antisense RNAs also contained RNAs complementary to tRNAs or sRNAs (small-nucleolar-like RNAs). For the antisense-box ncRNAs, the majority could be assigned to the class of C/D sRNAs, which specify 2'-O-methylation sites on rRNAs or tRNAs. Five C/D sRNAs of this group are predicted to target methylation at six sites in 13 different tRNAs, thus pointing to the widespread role of these sRNA species in tRNA modification in Archaea. Another group of antisense-box RNAs, lacking typical C/D sRNA motifs, was predicted to target the 3'-untranslated regions of certain mRNAs. Furthermore, one of the ncRNAs that does not show antisense elements is transcribed from a repeat unit of a cluster of small regularly spaced repeats in S. solfataricus which is potentially involved in replicon partitioning. In conclusion, this is the first report of stably expressed antisense RNAs in an archaeal species and it raises the prospect that antisense-based mechanisms are also used widely in Archaea to regulate gene expression. PMID- 15659163 TI - Ca2+-dependent expression of the CIRCE regulon in Streptococcus pneumoniae. AB - DnaK and GroEL play a pivotal role in protein folding, and promote cell proliferation and survival. In Gram-positive and several Gram-negative bacteria, HrcA represses the transcription of dnaK and groE operons by binding to the highly conserved CIRCE (controlling inverted repeat of chaperone expression) operator sequence in the presence of GroEL. HrcA may respond to environmental stress and various other factors that modulate the transcription of the dnaK and groE operons. However, the mechanisms by which these factors modulate the activity of HrcA remain elusive. Here, we show that the thermoresistance of Streptococcus pneumoniae is significantly repressed in the presence of Ca2+. Furthermore, heat shock-induced expression of the CIRCE regulon in S. pneumoniae is repressed in the presence of Ca2+, although to a lesser degree than in the hrcA mutant, strongly suggesting that HrcA inhibits expression of the CIRCE regulon in a Ca2+-dependent manner. Although HrcA does not bind directly to Ca2+, its hydrophobicity is increased in the presence of the metal ion. Taken together, our observations suggest that Ca2+ induces conformational changes, such as exposure of the hydrophobic surfaces of HrcA, which facilitate binding to GroEL. Alternatively, the presence of Ca2+ may facilitate GroEL in interacting freely with HrcA. This, in turn, enhances access to CIRCE and leads to repression of the dnaK and groE operons in S. pneumoniae. PMID- 15659165 TI - A constitutively active GPCR retains its G protein specificity and the ability to form dimers. AB - G protein-coupled receptors (GPCRs) are cell surface proteins which help to regulate the physiology of all the major organ systems within higher eukaryotes. They are stimulated by multiple ligands and activate a range of effector molecules to bring about changes in cell behaviour. The use of constitutively active mutants (CAMs) of GPCRs has enabled a better understanding of receptor activation as CAMs exhibit ligand-independent signalling negating the use of ligands. Here we introduce the fission yeast Schizosaccharomyces pombe as a host for producing CAMs, by describing the isolation and characterization of constitutive mutants of the P-factor receptor (Mam2). One mutant Mam2[P261L] contained a single-amino-acid substitution (Pro261 to Leu) within a region of high homology in GPCRs. Substitution of this proline leads to an 18-fold increase in ligand-independent signalling. We utilized Mam2[P261L] to investigate CAM activity by demonstrating that Mam2[P261L] is efficiently trafficked to the cell surface where it can form fully functional oligomeric complexes with the native receptor. Mam2[P261L] also retains the G protein specificity (RG-profile) of the native receptor and only induces constitutive signalling in the same G proteins. Finally, evidence is provided to indicate that CAM activity results from a reduction in the kinetics of G protein binding. This is the first time that S. pombe has been utilized for isolating and characterizing CAMs and the techniques employed will complement the current systems available for studying these important receptors. PMID- 15659166 TI - Redox-sensitive transcriptional control by a thiol/disulphide switch in the global regulator, Spx. AB - The Spx protein is indispensable for survival of Bacillus subtilis under disulphide stress. Its interaction with the alpha-subunit of RNA polymerase is required for transcriptional induction of genes that function in thiol homeostasis, such as thioredoxin (trxA) and thioredoxin reductase (trxB). The N terminal end of Spx contains a Cys-X-X-Cys (CXXC) motif, which is a likely target for redox-sensitive control. We show here that Spx directly activates trxA and -B transcription by interacting with the RNA polymerase alpha-subunit, but it does so only under an oxidized condition. The transcriptional activation by Spx requires formation of an intramolecular disulphide bond between two cysteine residues that reside in the CXXC motif. The mechanism of Spx-dependent transcriptional activation is unique in that it does not involve initial Spx-DNA interaction. PMID- 15659167 TI - Probing plasmid partition with centromere-based incompatibility. AB - Low-copy number plasmids of bacteria rely on specific centromeres for regular partition into daughter cells. When also present on a second plasmid, the centromere can render the two plasmids incompatible, disrupting partition and causing plasmid loss. We have investigated the basis of incompatibility exerted by the F plasmid centromere, sopC, to probe the mechanism of partition. Measurements of the effects of sopC at various gene dosages on destabilization of mini-F, on repression of the sopAB operon and on occupancy of mini-F DNA by the centromere-binding protein, SopB, revealed that among mechanisms previously proposed, no single one fully explained incompatibility. sopC on multicopy plasmids depleted SopB by titration and by contributing to repression. The resulting SopB deficit is proposed to delay partition complex formation and facilitate pairing between mini-F and the centromere vector, thereby increasing randomization of segregation. Unexpectedly, sopC on mini-P1 exerted strong incompatibility if the P1 parABS locus was absent. A mutation preventing the P1 replication initiation protein from pairing (handcuffing) reduced this strong incompatibility to the level expected for random segregation. The results indicate the importance of kinetic considerations and suggest that mini-F handcuffing promotes pairing of SopB-sopC complexes that can subsequently segregate as intact aggregates. PMID- 15659168 TI - Carbapenem antibiotic biosynthesis in Erwinia carotovora is regulated by physiological and genetic factors modulating the quorum sensing-dependent control pathway. AB - Erwinia carotovora produces the beta-lactam antibiotic, carbapenem, in response to a quorum sensing signalling molecule, N-(3-oxohexanoyl)-L-homoserine lactone (OHHL). We have mapped the OHHL-dependent promoter upstream of the first of the biosynthetic genes, carA. We have also analysed the effect on this promoter of the known genetic regulators of carbapenem expression, carR, carI (encoding homologues of LuxR and LuxI respectively) and hor (encoding a SlyA/MarR-like transcriptional regulator). We describe a previously unknown promoter located within the carA-H operon. This promoter does not respond to CarR and is required for quorum sensing-independent expression of the carbapenem resistance determinants encoded by the carFG genes. We have mapped the carR, carI and hor transcription start points, shown that CarR is positively autoregulated in the presence of OHHL, and have demonstrated negative feedback affecting transcription of carI. In addition, various environmental and physiological factors were shown to impinge on the transcription of the car biosynthetic genes. The nature of the carbon source and the temperature of growth influence carbapenem production by modulating the level of the OHHL signalling molecule, and thereby physiologically fine-tune the quorum sensing regulatory system. PMID- 15659169 TI - PMT family of Candida albicans: five protein mannosyltransferase isoforms affect growth, morphogenesis and antifungal resistance. AB - Protein O-mannosyltransferases (Pmt proteins) initiate O-mannosylation of secretory proteins. The PMT gene family of the human fungal pathogen Candida albicans consists of PMT1 and PMT6, as well as three additional PMT genes encoding Pmt2, Pmt4 and Pmt5 isoforms described here. Both PMT2 alleles could not be deleted and growth of conditional strains, containing PMT2 controlled by the MET3- or tetOScHOP1-promoters, was blocked in non-permissive conditions, indicating that PMT2 is essential for growth. A homozygous pmt4 mutant was viable, but synthetic lethality of pmt4 was observed in combination with pmt1 mutations. Hyphal morphogenesis of a pmt4 mutant was defective under aerobic induction conditions, yet increased in embedded or hypoxic conditions, suggesting a role of Pmt4p-mediated O-glycosylation for environment-specific morphogenetic signalling. Although a PMT5 transcript was detected, a homozygous pmt5 mutant was phenotypically silent. All other pmt mutants showed variable degrees of supersensitivity to antifungals and to cell wall-destabilizing agents. Cell wall composition was markedly affected in pmt1 and pmt4 mutants, showing a significant decrease in wall mannoproteins. In a mouse model of haematogenously disseminated infection, PMT4 was required for full virulence of C. albicans. Functional analysis of the first complete PMT gene family in a fungal pathogen indicates that Pmt isoforms have variable and specific roles for in vitro and in vivo growth, morphogenesis and antifungal resistance. PMID- 15659170 TI - Characterization of a Mycobacterium tuberculosis proteasomal ATPase homologue. AB - A screen for Mycobacterium tuberculosis (Mtb) mutants sensitive to reactive nitrogen intermediates identified transposon insertions in the presumptive proteasomal ATPase gene mpa (mycobacterium proteasome ATPase; Rv2115c). mpa mutants are attenuated in both wild type and nitric oxide synthase 2 deficient mice. In this work, we show that attenuation of mpa mutants is severe, and that Mpa is an ATPase associated with various cellular activities (AAA) ATPase that forms hexameric rings resembling the eukaryotic complex p97/valosin-containing protein (VCP). Point mutations in the conserved Walker box ATPase motifs of Mpa greatly reduced or abolished ATPase activity in vitro and abrogated protection of Mtb against acidified nitrite. A mutant Mpa protein missing only its last two amino acids retained ATPase activity, yet failed to protect Mtb against nitrite. The corresponding strain was attenuated in mice. Thus, Mpa is an ATPase whose enzymatic activity is necessary but not sufficient to protect against reactive nitrogen intermediates. PMID- 15659171 TI - The polo-like kinase PLKA is required for initiation and progression through mitosis in the filamentous fungus Aspergillus nidulans. AB - Polo-like kinases (PLK) function during multiple stages of mitotic progression and in cytokinesis. We identified and cloned a PLK homologue in Aspergillus nidulans, plkA, which is the first PLK reported in a filamentous fungus and the largest member of the PLK family to date. As plkA was essential, the effects of overexpression and localization of protein in living cells were explored to determine PLKA function. Overexpression of PLKA permitted hyphal formation, but blocked nuclear division in interphase. In NIMA or NIMT temperature-sensitive backgrounds, overexpression of PLKA impaired normal entry into mitosis upon release from restrictive temperature, supporting a role for PLKA during G2/M. In the few mitotic cells present, spindles were monopolar or disorganized, and chromatin condensation and segregation were impaired, suggesting additional roles for PLKA in spindle formation and in chromosome dynamics. Consistent with this, green fluorescent protein (GFP)-tagged PLKA could localize to the spb during interphase, and to the spb and nucleus throughout mitosis. Intriguingly, PLKA remained on the spb during telophase and into G1, in contrast to other PLK. In addition, spb localization was independent of NIMA function, unlike that demonstrated in Schizosaccharomyces pombe where PLK localization to the spb required the NIMA homologue Fin1. PLKA was not detected at cortical, septation associated sites, and overexpression did not drive septum formation, also in contrast to that observed with other PLK. Therefore, PLKA is important for multiple events during mitosis, similar to PLK in higher organisms, but exhibits differences in size, localization and influence on septation/cytokinesis, suggesting additional novel regulatory features. PMID- 15659172 TI - Physiological characterization of Saccharomyces cerevisiae kha1 deletion mutants. AB - Maintenance of intracellular K+ homeostasis is one of the crucial requisites for the survival of yeast cells. In Saccharomyces cerevisiae, the high K+ content corresponds to a steady state between simultaneous influx and efflux across the plasma membrane. One of the transporters formerly believed to extrude K+ from the yeast cells (besides Ena1-4p and Nha1p) was named Kha1p and presumed as a putative plasma membrane K+/H+ antiporter. We prepared kha1 and tok1-kha1 deletion strains in the B31 and MAB 2d background. Both the strains contain the ena1-4 and nha1 deletions; that means they lack the main active sodium and potassium efflux systems. MAB 2d has additional trk1 and trk2 deletions, i.e. is impaired in active K+ uptake as well. We performed a large physiological study with these strains to specify the phenotype of kha1 deletion. In our experiments, no difference in K+ content or efflux was observed in strains lacking the KHA1 gene compared with control strains. Two main phenotype manifestations of the kha1 deletion were growth defect on high external pH and hygromycin sensitivity. The correlation between these phenotypes and the kha1 deletion was confirmed by plasmid complementation. Fluorescence microscopy of green fluorescent protein (GFP)-tagged Kha1p showed that this antiporter is localized preferentially intracellularly (in contrast to the plasma membrane Na+/H+ antiporter Nha1p). Based on these findings, Kha1p is probably not localized in plasma membrane and does not mediate efflux of alkali metal cations from cells, but is important for the regulation of intracellular cation homeostasis and optimal pH control, similarly as the Nhx1p. PMID- 15659173 TI - Topological insulators inhibit diffusion of transcription-induced positive supercoils in the chromosome of Escherichia coli. AB - The double helical nature of DNA implies that progression of transcription machinery that cannot rotate easily around the DNA axis creates waves of positive supercoils ahead of it and negative supercoils behind it. Using topological reporters that detect local variations in DNA supercoiling, we have characterized the diffusion of transcription-induced (TI) positive supercoils in plasmids or in the chromosome of wild type Escherichia coli cells. Transcription-induced positive supercoils were able to diffuse and affect local supercoiling several kilobases away from the site of origin. By testing the effect of various DNA sequences, these reporters enabled us to identify elements that impede supercoil diffusion, i.e. behave as topological insulators. All the elements tested correspond to DNA gyrase catalytic targets. These results correlate the ability of a DNA sequence to be cleaved by DNA gyrase with topological insulator activity. Implications of the asymmetry in supercoil diffusion for the control of DNA topology are discussed. PMID- 15659174 TI - The expression of a novel antisense gene mediates incompatibility within the large repABC family of alpha-proteobacterial plasmids. AB - Large extrachromosomal replicons in many members of the alpha-proteobacteria encode genes that are required for plant or animal pathogenesis or symbiosis. Most of these replicons encode repABC genes that control their replication and faithful segregation during cell division. In addition to its chromosome, the plant endosymbiont Sinorhizobium meliloti also maintains the 1.4 Mb pSymA and 1.7 Mb pSymB symbiotic megaplasmids both of which are repABC-type replicons. In all repABC loci that have been characterized, an apparently untranslated intergenic region between the repB and repC genes encodes a strong incompatibility determinant (referred to as incalpha). Here we report the isolation of mutations within the incalpha regions of pSymA and pSymB that eliminate incompatibility. These mutations map to and inactivate a promoter in the intergenic region that drives the expression of an approximately 56 nucleotide untranslated RNA molecule that mediates incompatibility. This gene, that we have named incA, is transcribed antisense to the repABC genes. Our analysis suggests that the incA gene is conserved in repABC loci from a diverse spectrum of bacteria. PMID- 15659175 TI - GlcP constitutes the major glucose uptake system of Streptomyces coelicolor A3(2). AB - We provide a functional and regulatory analysis of glcP, encoding the major glucose transporter of Streptomyces coelicolor A3(2). GlcP, a member of the Major Facilitator Superfamily (MFS) of bacterial and eucaryotic sugar permeases, was found to be encoded twice at two distinct loci, glcP1 and glcP2, located in the central core and in the variable right arm of the chromosome respectively. Heterologous expression of GlcP in Escherichia coli led to the full restoration of glucose fermentation to mutants lacking glucose transport activity. Biochemical analysis revealed an affinity constant in the low-micromolar range and substrate specificity for glucose and 2-deoxyglucose. Deletion of glcP1 but not glcP2 led to a drastic reduction in growth on glucose reflected by the loss of glucose uptake. This correlated with transcriptional analyses, which showed that glcP1 transcription was strongly inducible by glucose, while glcP2 transcripts were barely detectable. In conclusion, GlcP, which is the first glucose permease from high G+C Gram-positive bacteria characterized at the molecular level, represents the major glucose uptake system in S. coelicolor A3(2) that is indispensable for the high growth rate on glucose. It is anticipated that the activity of GlcP is linked to other glucose-mediated phenomena such as carbon catabolite repression, morphogenesis and antibiotic production. PMID- 15659176 TI - Differential expression of the NRG1 repressor controls species-specific regulation of chlamydospore development in Candida albicans and Candida dubliniensis. AB - Candida albicans and Candida dubliniensis are opportunistic fungal pathogens that are closely related but differ in their epidemiology and in some phenotypic characteristics, including certain virulence-related traits. A comparison of these two species at the molecular level could therefore provide new insights into the biology and pathogenicity of Candida. Both species share the ability to produce chlamydospores, but only C. dubliniensis forms pseudohyphae with abundant chlamydospores on Staib agar (syn. Guizotia abyssinica creatinine agar), on which C. albicans grows as a budding yeast. To understand the basis of this species specific, differential regulation of morphogenetic development, we set out to identify C. albicans genes that repress chlamydospore formation under these conditions. A C. albicans genomic library was integrated into the C. dubliniensis genome and transformants were screened for clones in which filamentation and/or chlamydospore production on Staib agar was suppressed. This screen identified two genes, CaNRG1 and CaPDE2, encoding a general transcriptional repressor and a high affinity cAMP phosphodiesterase, respectively. Expression of CaNRG1 in C. dubliniensis repressed pseudohyphae and chlamydospore formation, whereas expression of CaPDE2 only reduced the extent of filamentous growth but did not affect chlamydospore formation. We found that C. dubliniensis, but not C. albicans, specifically downregulates NRG1 expression on Staib medium to allow chlamydospore development. Artificial overexpression of CdNRG1 suppressed pseudohyphal growth and production of chlamydospores in C. dubliniensis. Conversely, deletion of CaNRG1 in C. albicans resulted in chlamydospore formation on Staib agar, confirming its central role in the regulation of this morphogenetic process. Our results demonstrate that differential regulation of a single gene, NRG1, in C. albicans and C. dubliniensis is responsible for their species-specific response to environmental signals that induce chlamydospore development. PMID- 15659179 TI - Inactivation of Staphylococcus aureus in raw milk cheese by combinations of high pressure treatments and bacteriocin-producing lactic acid bacteria. AB - AIMS: To investigate the combined effect of high-pressure treatments (HPT) and milk inoculation with bacteriocin-producing lactic acid bacteria (BP-LAB) on the survival of Staphylococcus aureus during ripening of raw milk cheese. METHODS AND RESULTS: Cheeses were manufactured from raw milk artificially contaminated with S. aureus at ca 5 log CFU ml(-1), a commercial starter culture and one of seven strains of BP-LAB, added as adjuncts at 0.1%. HPT of cheeses were performed on days 2 or 50 at 300 MPa (10 degrees C, 10 min) or 500 MPa (10 degrees C, 5 min). On day 3, S. aureus counts were 6.46 log CFU g(-1) in control cheese. Milk inoculation with different BP-LAB lowered S. aureus counts on day 3 when compared with control cheese by up to 0.46 log CFU g(-1), HPT at 300 MPa on day 2 by 0.45 log CFU g(-1) and HPT at 500 MPa on day 2 by 2.43 log CFU g(-1). Combinations of BP-LAB with HPT at 300 and 500 MPa on day 2 lowered S. aureus counts on day 3 by up to 1.02 and 4.00 log CFU g(-1) respectively. CONCLUSIONS: The combined effect of milk inoculation with some of the BP-LAB tested and HPT of cheese on S. aureus inactivation was synergistic. SIGNIFICANCE AND IMPACT OF THE STUDY: The combination of HPT at lower pressures with BP-LAB inoculation is a feasible system to improve cheese safety in case of deleterious effects on cheese quality caused by HPT at higher pressures. PMID- 15659180 TI - Identification of genes and proteins induced during the lag and early exponential phase of lager brewing yeasts. AB - AIMS: The aim of the present study is to identify genes and proteins whose expression is induced in lager brewing yeast during the lag phase and early exponential growth. METHODS AND RESULTS: Two-dimensional gel electrophoresis was used to identify proteins induced during the lag and early exponential phase of lager brewing yeast in minimal medium. The identified, early-induced proteins were Ade17p, Eno2p, Ilv5gp, Sam1p, Rps21p and Ssa2p. For most of these proteins, the patterns of induction differed from those of the corresponding genes. However, the genes had similar early expression patterns in minimal medium as observed during lager brewing conditions. The expression of previously identified early-induced genes in Saccharomyces cerevisiae grown in minimal medium, ADO1, ALD6, ASC1, ERG4, GPP1, RPL25, SSB1 and YKL056C, was also early induced in lager yeast under brewing conditions. CONCLUSIONS: The results indicate that the above mentioned genes in general are induced during the lag phase and early exponential growth in Saccharomyces yeasts. The processes in which these genes take part are likely to play an important role during growth initiation. SIGNIFICANCE AND IMPACT OF THE STUDY: Increased knowledge regarding the early growth phase of lager brewing yeast was obtained. Further, the universality of the identified expression patterns suggests new methodologies for optimization and control of growth initiation during brewing fermentations. PMID- 15659181 TI - Growth and exopolysaccharide production during free and immobilized cell chemostat culture of Lactobacillus rhamnosus RW-9595M. AB - AIMS: Biomass and exopolysaccharide (EPS) production were studied during chemostat cultures in whey permeate medium with Lactobacillus rhamnosus RW-9595M free cells and cells immobilized on solid porous supports (ImmobaSil). METHODS AND RESULTS: A continuous culture with free cells was conducted for 9 days at dilution rates (D) between 0.3 and 0.8 h(-1) in yeast extract (YE)/mineral supplemented whey permeate. Maximum EPS production (1808 mg l(-1)) and volumetric productivity (542.6 mg l(-1) h(-1)) were obtained for a low D of 0.3 h(-1). A continuous fermentation in a two-stage bioreactor system, composed of a first stage with immobilized cells and a second stage inoculated with free cells produced in the first reactor, was carried out for 32 days. The influence of YE concentration, temperature and dilution rate, and their interactions on biomass, EPS and lactic acid production was investigated. A statistically significant model was found only for lactic acid production. Marked cell morphological and physiological changes led to the formation of very large cell-containing aggregates and a low mean soluble EPS production (138 mg l(-1)). Aggregate volumetric productivity of the two-stage system varied between 5.7 and 49.5 g l( 1) h(-1) for different fermentation conditions and times. Aggregates contained a very high biomass concentration, estimated at 74% of aggregate dry weight by nitrogen analysis and 4.3 x 10(12) CFU g(-1) by a DNA extraction method and a high nonsoluble polysaccharide content (14.2%). At age 24 days, insoluble EPS concentration and volumetric productivity were 1250 mg l(-1) and 2240 mg l(-1) h( 1) respectively. The physiological changes were shown to be reversible when cells were incubated during three successive batch cultures. CONCLUSIONS: EPS production and volumetric productivity during continuous free-cell chemostat cultures with L. rhamnosus RW-9595M are among the highest values reported for lactobacilli in literature. Immobilization and continuous culture resulted in low soluble EPS production and large morphological and physiological changes of L. rhamnosus RW-9595M, with formation of macroscopical aggregates mainly composed of biomass and nonsoluble EPS. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first study on continuous EPS production by immobilized LAB. Immobilization and culture time-induced cell aggregation and could be used to produce new synbiotic products with very high viable cell and EPS concentrations. PMID- 15659182 TI - Importance of S-layer proteins in probiotic activity of Lactobacillus acidophilus M92. AB - AIMS: To investigate the functional role of surface layer proteins (S-layer) in probiotic strain Lactobacillus acidophilus M92, especially its influence on adhesiveness to mouse ileal epithelial cells. METHODS AND RESULTS: Sodium dodecyl sulphate polyacrylamide gel electrophoresis of cell surface proteins revealed the presence of potential surface layer (S-layer) proteins, ca at 45 kDa in L. acidophilus M92. Southern blot with pBK1 plasmid, containing slpA gene, gave a positive signal, suggesting that L. acidophilus M92 has a slpA gene coding for the S-layer proteins. S-layer proteins of this strain are present during all phases of growth. The S-layer proteins appeared when cells treated with 5 mol l( 1) LiCl were allowed to grow again. Removal of the S-layer proteins reduced adhesion of L. acidophilus M92 to mouse ileal epithelial cells. Furthermore, the viability of cells without S-layer were reduced in simulated gastric juice at low pH range (2, 2.5, 3) and simulated pancreatic juice with bile salts (1.5 and 3 g l(-1)). S-layer proteins of L. acidophilus M92 were resistant to pepsin and pancreatin, in contrast, the treatment with proteinase K led to a significant proteolysis of the S-layer proteins. CONCLUSIONS: These results demonstrated functional role of S-layer; it is responsible for adhesiveness of Lactobacillus acidophilus M92 to mouse ileal epithelial cells and has a protective role for this strain. SIGNIFICANCE AND IMPACT OF THE STUDY: S-layer proteins have an important role in the establishment of probiotic strain Lactobacillus acidophilus M92 in the gastrointestinal tract. PMID- 15659183 TI - Aerial release of bacteria from cot mattress materials and the sudden infant death syndrome. AB - AIM: To investigate aerial release of bacteria from used cot mattresses and to assess factors that may influence this process. METHODS AND RESULTS: Movement on used mattresses, simulating that of an infant's head, significantly enhanced aerial release of naturally acquired bacteria from the polyurethane foams (total count data, P = 0.008; Staphylococcus aureus, P = 0.004) or from polyvinyl chloride covers (total count data, P = 0.001). Aerial release of naturally acquired bacteria from used cot mattresses showed high variability and was poorly correlated (R2 < or = 0.294) with bacterial cell density within the materials. In experiments involving inoculation of S. aureus and Escherichia coli onto the polyurethane of unused cot mattresses, aerial release of the species correlated well (R2 > or = 0.950) with inoculation density when simulated infant head movement was applied. Aerial release of these bacterial species from the material decreased with increase in width or aqueous content of the material, and was lower from polyurethane foam of a used cot mattress. CONCLUSIONS: Simulated infant movement and mattress related factors influence aerial release of bacteria from cot mattress materials. With simulated infant movement on cot mattress polyurethane foam, levels of airborne bacteria above the material are proportional to bacterial population levels inoculated onto the material. SIGNIFICANCE AND IMPACT OF THE STUDY: Cot mattresses harbouring relatively high levels of naturally acquired toxigenic bacteria, such as S. aureus, could pose a relatively high risk of infection to the infant's respiratory tract through increased aerial contamination. This has impact in the context of recent findings on cot mattress related risk factors for sudden infant death syndrome. PMID- 15659184 TI - Analysis of the expression of some stress induced genes in several commercial wine yeast strains at the beginning of vinification. AB - AIMS: During fermentation yeast cells should cope with stress conditions. We pursue a better understanding of the stress response in wine yeasts at the beginning of vinification. METHODS AND RESULTS: We analyse by means of quantitative PCR the expression of several stress induced genes in 24 efficient commercial wine yeast strains at the beginning of vinifications performed under standard conditions or with small variations in pH and temperature. In all cases, high levels (with differences among strains) of GPD1 mRNA but quite low expression of other stress genes (TRX2, HSP104 and SSA3) were found. For all these genes, mRNA levels increase as temperature decreases or pH increases. CONCLUSIONS: Important levels of expression of GPD1 (but not of other stress genes) are required during the first hours of vinification, because of the need for glycerol production to counteract the hyperosmotic stress at this point. The differences among strains suggest that certain level of expression is enough to ensure the continuity of the process. Variations in the pH and temperature of the vinification can affect gene expression. SIGNIFICANCE AND IMPACT OF THE STUDY: A common pattern of stress response between efficient wine strains exists, which could be used as a criterion for selection. Studies of this kind can allow the establishment of connections between gene expression and physiological traits. PMID- 15659185 TI - Environmental inactivation of Cryptosporidium oocysts in catchment soils. AB - AIMS: To generate field-relevant inactivation rates for Cryptosporidium oocysts in soil that may serve as parameter values in models to predict the terrestrial fate and transport of oocysts in catchments. METHODS AND RESULTS: The inactivation of Cryptosporidium oocysts in closed soil microcosms over time was monitored using fluorescence in situ hybridization (FISH) as an estimate of oocyst 'viability'. Inactivation rates for Cryptosporidium in two soils were determined under a range of temperature, moisture and biotic status regimes. Temperature and soil type emerged as significantly influential factors (P < 0.05) for Cryptosporidium inactivation. In particular, temperatures as high as 35 degrees C may result in enhanced inactivation. CONCLUSIONS: When modelling the fate of Cryptosporidium oocysts in catchment soils, the use of inactivation rates that are appropriate for the specific catchment climate and soil types is essential. FISH was considered cost-effective and appropriate for determining oocyst inactivation rates in soil. SIGNIFICANCE AND IMPACT OF THE STUDY: Previous models for predicting the fate of pathogens in catchments have either made nonvalidated assumptions regarding inactivation of Cryptosporidium in the terrestrial environment or have not considered it at all. Field-relevant inactivation data are presented, with significant implications for the management of catchments in warm temperate and tropical environments. PMID- 15659186 TI - Novel mutation method for increased cellulase production. AB - AIM: Isolation of cellulase producing fungi and increasing cellulase production using novel mutations. METHODS AND RESULTS: Cellulase-producing fungi were isolated from different soil samples using enriched Mandels cellulose agar, which is a selective media and seven different fungi were selected in the screening programme. These organisms were tested for cellulase production and two potent strains were identified. Two methods of mutations for strain improvement were employed to these strains. (1) Germinating fungal spore suspension was treated with 0.1 and 0.2 mg ml(-1) of 1-methyl-3-nitro-1-nitrosoguanidine (MNNG), ethidium bromide (EtBr) and u.v. for 30 min and 1 h duration and plated on selective media with and with out amphotericin B. (2) Mutagens (EtBr and MNNG) were incorporated in the selective media in sublethal concentration (5 microg ml( 1)) along with antifungal antibiotic (amphotericin B 2 microg ml(-1)). Second method yielded maximum cellulase-producing mutants, which are also stable for cellulase production and are more potent than the mutants obtained by the first method. CONCLUSIONS: Mutations using sublethal concentrations of mutagen for a prolonged period of growth has yielded mutants, which can produce more cellulase. SIGNIFICANCE AND IMPACT OF THE STUDY: This new method could be applied to obtain potent fungal mutants for more enzymes production. PMID- 15659187 TI - Production of alternatives to fuel oil from organic waste by the alkane-producing bacterium, Vibrio furnissii M1. AB - AIMS: We investigated the production of alternatives to fuel oil through the bacterial metabolism of organic waste. The availability for this purpose of various sources of organic waste for hydrocarbon production by the alkane producing bacterium, Vibrio furnissii M1, was examined. METHODS AND RESULTS: We screened 17 authentic compounds which can generally be found in organic waste for their hydrocarbon production. Carbon (3 mmol) in a 50-ml culture with acetic acid, lactic acid, butyric acid, succinic acid, malic acid, pentanoic acid, hexanoic acid glucose, xylose, starch or sucrose yielded 10-27 mg of alkanes or alkenes. The chain length of these alkanes or alkenes varied according to the culture from C14 to C27. Varying the ratio of carbon to nitrogen in the culture had no effect on the hydrocarbon production. Crude blackstrap molasses were also converted into alkanes with a conversion ratio of 20% (half of that in an authentic sucrose medium) of the total carbon consumption. CONCLUSIONS: V. furnissii M1 could produce hydrocarbons corresponding to kerosene or light oil from volatile fatty acids and sugars. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first report on bacterial hydrocarbon production from organic waste. PMID- 15659188 TI - Characterization of a new tyrosinase from Pycnoporus species with high potential for food technological applications. AB - AIMS: Tyrosinase production by Pycnoporus cinnabarinus and Pycnoporus sanguineus was screened among 20 strains originating from various geographical areas, particularly from tropical environments. The tyrosinase from the most efficient strain was purified and characterized and tested for food additive applications. METHODS AND RESULTS: Monophenolase and diphenolase activities of tyrosinase were measured from cell lysate from the 20 Pycnoporus strains, for 8-10 days of cultivation. The strain P. sanguineus CBS 614.73 showed the highest productivity (45.4 and 163.6 U g(-1) protein per day for monophenolase and diphenolase respectively). P. sanguineus CBS 614.73 tyrosinase was purified from concentrated cell lysate, anion-exchange, size-exclusion and hydroxyapatite chromatography, with a final yield of 2% and a purification factor of 35-38. The pure enzyme was a monomere with a molecular mass of 45 kDa and it showed four isoforms or isoenzymes with pI between 4.5-5. No N-glycosylation was found. The N-terminal amino acid sequence was IVTGPVGGQTEGAPAPNR. The enzyme was shown to be almost fully active in a pH range of 6-7, in a large temperature range (30-70 degrees C), and was stable below 60 degrees C. The main kinetic constants were determined. The tyrosinase was able to convert p-tyrosol and p-coumaric acid into hydroxytyrosol and caffeic acid, respectively, and it could also catalyse the cross-linking formation of a model protein. CONCLUSIONS: Among the genus Pycnoporus, known for the production of laccase, the strain P. sanguineus CBS 614.73 was shown to produce one other phenoloxidase, a new monomeric tyrosinase with a specific activity of 30 and 84 U mg(-1) protein for monophenolase and diphenolase respectively. SIGNIFICANCE AND IMPACT OF THE STUDY: This study identified P. sanguineus CBS 614.73 as a potential producer of a tyrosinase which demonstrated effectiveness in the synthesis of antioxidant molecules and in protein cross-linking. PMID- 15659189 TI - Diversity of Staphylococcus aureus enterotoxin types within single samples of raw milk and raw milk products. AB - AIM: To find out if testing of up to 10 Staphylococcus aureus isolates from each sample from raw milk and raw milk products for staphylococcal enterotoxin (SE) might increase the chances of identifying potential sources of food intoxication. METHODS AND RESULTS: Altogether 386 S. aureus isolates were tested for the presence of SE by reversed passive latex agglutination (SET-RPLA), and SE genes (se) by a multiplex polymerase chain reaction (PCR). In 18 of 34 (53%) S. aureus positive samples a mixture of SE and/or se positive and negative isolates were identified. Multiplex PCR increased the number of potential SE producing strains, i.e. isolates that harboured se, with 51% among the product and 48% among the raw bovine milk isolates. Examination by pulsed-field gel electrophoresis mostly confirmed clonal similarity among isolates sharing SE/se profile, but did not further differentiate between them. CONCLUSIONS: Isolates of S. aureus collected from one sample may show great diversity in SE production and different plating media seem to suppress or favour different strains of S. aureus. SIGNIFICANCE AND IMPACT OF THE STUDY: Several isolates of S. aureus from each sample should be tested for enterotoxin production in cases with typical SE intoxication symptoms with methods that are able to reveal new SE/se. PMID- 15659190 TI - Relationship between physiochemical properties, aggregation and u.v. inactivation of isolated indigenous spores in water. AB - AIMS: The objective of the study was to compare ultraviolet (u.v.) inactivation kinetics of indigenous aerobic spores in surface water with their laboratory cultured spore isolates and to investigate the relationship between physicochemical characteristics and u.v. inactivation kinetics of spore isolates. METHODS AND RESULTS: Lake water samples were analysed for the presence of indigenous aerobic spores. Different bacterial isolates from the heterogeneous indigenous population were genetically characterized, resporulated and examined for hydrophobicity, surface charge, particle size distribution and survival at different u.v. 254 nm fluence levels. Cultured isolated spores exhibited a three stage inactivation curve consisting of shoulder, first order and tailing regions whereas indigenous spores exhibited only one stage of linear kinetics. Hydrophobicity of the Bacillus spore isolates was inversely related to the extent of u.v. inactivation before tailing occurred. CONCLUSIONS: Tailing in the u.v. inactivation curves results from aggregation of a portion of the spore population because of hydrophobic interactions, supporting the link between aggregation of spores, hydrophobicity and u.v. inactivation. SIGNIFICANCE AND IMPACT OF THE STUDY: Evidence of the link between spore physicochemical parameters and u.v. disinfection performance furthers the understanding of factors that affect inactivation of microbes in natural waters supplied to drinking water treatment plants. PMID- 15659191 TI - Characterization of Staphylococcus aureus mutants expressing reduced susceptibility to common house-cleaners. AB - AIMS: To characterize mutants of Staphylococcus aureus expressing reduced susceptibility to house cleaners (HC), assess the impact of the alternative sigma factor SigB on HC susceptibility, and determine the MIC of clinical methicillin resistant S. aureus (MRSA) to a HC. METHODS AND RESULTS: Susceptibility to HC, HC components, H2O2, vancomycin and oxacillin and physiological parameters were determined for HC-reduced susceptibility (HCRS) mutants, parent strain COL and COLsigB::kan. HCRS mutants selected with three HC expressed reduced susceptibility to multiple HC, HC components, H2O2 and vancomycin. Two unique HCRS mutants also lost the methicillin resistance determinant. In addition, all HCRS mutants exhibited better growth at two temperatures, and one HCRS mutant expressed reduced carotenoid production. COLsigB::kan demonstrated increased susceptibility to all HC and many HC components. sigB operon mutations were not detected in one HCRS mutant background. Of 76 clinical MRSA, 20 exhibited reduced susceptibility to a HC. CONCLUSIONS: HCRS mutants demonstrate altered susceptibility to multiple antimicrobials. While sigB is required for full HC resistance, one HCRS mechanism does not involve sigB operon mutations. Clinical MRSA expressing reduced susceptibility to a common HC were detected. SIGNIFICANCE AND IMPACT OF THE STUDY: This study suggests that HCRS mutants are not protected against, nor selected by, practical HC concentrations. PMID- 15659192 TI - Legionella waterline colonization: detection of Legionella species in domestic, hotel and hospital hot water systems. AB - AIMS: An evaluation was made of the prevalence of Legionella species in hot water distribution systems in the city of Bologna (Italy) and their possible association with bacterial contamination (total counts and Pseudomonadaceae) and the chemical characteristics of the water (pH, Ca, Mg, Fe, Mn, Cu, Zn and Total Organic Carbon, TOC). METHODS AND RESULTS: A total of 137 hot water samples were analysed: 59 from the same number of private apartments, 46 from 11 hotels and 32 from five hospitals, all using the same water supply. Legionella species were detected in 40.0% of the distribution systems, L. pneumophila in 33.3%. The highest colonization was found in the hot water systems of hospitals (93.7% of samples positive for L. pneumophila, geometric mean: 2.4 x 10(3) CFU l(-1)), followed by the hotels (60.9%, geometric mean: 127.3 CFU l(-1)) and the apartments with centralized heating (41.9%, geometric mean: 30.5 CFU l(-1)). The apartments with independent heating systems showed a lower level of colonization (3.6% for Legionella species), with no evidence of L. pneumophila. Correlation analysis suggests that copper exerts an inhibiting action, while the TOC tends to favour the development of L. pneumophila. No statistically significant association was seen with Pseudomonadaceae, which were found at lower water temperatures than legionellae and in individual distribution points rather than in the whole network. CONCLUSIONS: The water recirculation system used by centralized boilers enhances the spreading of legionellae throughout the whole network, both in terms of the number of colonized sites and in terms of CFU count. SIGNIFICANCE AND IMPACT OF THE STUDY: Differences in Legionella colonization between types of buildings are not due to a variation in water supply but to other factors. Besides the importance of water recirculation, the study demonstrates the inhibiting action of copper and the favourable action of TOC on the development of L. pneumophila. PMID- 15659193 TI - Land application of treated sewage sludge: quantifying pathogen risks from consumption of crops. AB - AIMS: To predict the number of humans in the UK infected through consumption of root crops grown on agricultural land to which treated sewage sludge has been applied in accordance with the current regulations and guidance (Safe Sludge Matrix). METHODS AND RESULTS: Quantitative risk assessments based on the source, pathway, receptor approach are developed for seven pathogens, namely salmonellas, Listeria monocytogenes, campylobacters, Escherichia coli O157, Cryptosporidium parvum, Giardia, and enteroviruses. Using laboratory data for pathogen destruction by mesophilic anaerobic digestion, and not extrapolating experimental data for pathogen decay in soil to the full 30-month harvest interval specified by the Matrix, predicts 50 Giardia infections per year, but less than one infection per year for the other six pathogens. Assuming linear decay in the soil, a 12-month harvest interval eliminates the risks from all seven pathogens; the highest predicted being one infection of C. parvum in the UK every 45 years. Computer simulations show that a protective effect from binding of pathogens to particulate matter could potentially exaggerate the observed rate of decay in experimental systems. CONCLUSIONS: The results confirm, assuming pathogens behave according to our current understanding, that the risks to humans from consumption of vegetable crops are remote. Furthermore the harvest intervals stipulated by the Safe Sludge Matrix compensate for potential lapses in the operational efficiency of sludge treatment. SIGNIFICANCE AND IMPACT OF THE STUDY: The models demonstrate the huge potential impact of decay in the soil over the 12/30-month intervals specified by the Matrix, although lack of knowledge on the exact nature of soil decay processes is a source of uncertainty. The models enable the sensitivity of the predicted risks to changes in the operational efficiency of sewage sludge treatment to be assessed. PMID- 15659194 TI - Estimation of bioaerosol risk of infection to residents adjacent to a land applied biosolids site using an empirically derived transport model. AB - AIM: The purpose of this study was to develop an empirically derived transport model, which could be used to predict downwind concentrations of viruses and bacteria during land application of liquid biosolids and subsequently assess microbial risk associated with this practice. METHODS AND RESULTS: To develop the model, coliphage MS-2 and Escherichia coli were aerosolized after addition to water within a biosolids spray application truck, and bioaerosols were collected at discrete downwind distances ranging from 2 to 70 m. Although coliphage were routinely detected, E. coli did not frequently survive aerosolization. Data on aerosolized coliphage was then used to generate a virus transport model. Risks of infection were calculated for various ranges of human virus concentrations that could be found in biosolids. CONCLUSIONS: A conservative estimate at 30.5 m (assumed to be nearest adjacent residences) downwind, resulted in risks of infection of 1 : 100,000, to the more realistic 1 : 10,000,000 per exposure. Conservative annual risks were calculated to be no more than 7 : 100,000 where as a more realistic risk was no greater than 7 : 10,000,000. Overall, the viral risk to residences adjacent to land application sites appears to be low, both for one time and annual probabilities of infection. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrated a simple approach towards modelling viral pathogens aerosolized from land applied liquid biosolids, and offers insight into the associated viral risk. PMID- 15659195 TI - Generating microbial survival curves during thermal processing in real time. AB - AIMS: To develop a method to calculate and record theoretical microbial survival curves during thermal processing of foods and pharmaceutical products simultaneously with the changing temperature. Moreover, to demonstrate that the method can be used to calculate nonisothermal survival curves, with widely available software such as Microsoft Excel. METHODS AND RESULTS: It has been assumed that the targeted organism's isothermal survival curves are not log linear and hence, the inactivation rate in nonisothermal processes is a function of the momentary temperature and the corresponding survival ratio. This could be expressed by a difference equation, which is an approximation to the continuous rate model. The concept was tested with the isothermal survival parameters of Clostridium botulinum and Bacillus sporothermodurans spores, and Salmonella enteritidis cells, using different kinds of survival models and under temperature profiles resembling those of commercial processes. As expected, there was an excellent agreement between the curves produced by solving the differential equation of the continuous model and by the incremental method, which has been posted on the web as freeware. CONCLUSIONS: It is possible to calculate nonisothermal survival curves, in real time, with an algorithm that can be written in the language of general purpose software, to follow the inactivation of one or more targeted organisms simultaneously and to simulate microbial survival patterns under existing or planned industrial thermal processes. SIGNIFICANCE AND IMPACT OF THE STUDY: Replacement of the traditional 'F0-value', which requires the log linearity of the organism's isothermal survival curves, by the more realistic theoretical survival ratio estimate as a measure of the thermal process efficacy. PMID- 15659196 TI - High level expression of a recombinant acid phytase gene in Pichia pastoris. AB - AIMS: To achieve high phytase yield with improved enzymatic activity in Pichia pastoris. METHODS AND RESULTS: The 1347-bp phytase gene of Aspergillus niger SK 57 was synthesized using a successive polymerase chain reaction and was altered by deleting intronic sequences, optimizing codon usage and replacing its original signal sequence with a synthetic signal peptide (designated MF4I) that is a codon modified Saccharomyces cerevisiae mating factor alpha-prepro-leader sequence. The gene constructs containing wild type or modified phytase gene coding sequences under the control of the highly-inducible alcohol oxidase gene promoter with the MF4I- or wild type alpha-signal sequence were used to transform Pichia pastoris. The P. pastoris strain that expressed the modified phytase gene (phyA-sh) with MF4I sequence produced 6.1 g purified phytase per litre of culture fluid, with the phytase activity of 865 U ml(-1). The expressed phytase varied in size (64, 67, 87, 110 and 120 kDa), but could be deglycosylated to produce a homogeneous 64 kDa protein. The recombinant phytase had two pH optima (pH 2.5 and pH 5.5) and an optimum temperature of 60 degrees C. CONCLUSIONS: The P. pastoris strain with the genetically engineered phytase gene produced 6.1 g l(-1) of phytase or 865 U ml( 1) phytase activity, a 14.5-fold increase compared with the P. pastoris strain with the wild type phytase gene. SIGNIFICANCE AND IMPACT OF THE STUDY: The P. pastoris strain expressing the modified phytase gene with the MF4I signal peptide showed great potential as a commercial phytase production system. PMID- 15659197 TI - Myxobacteria isolated in Israel as potential source of new anti-infectives. AB - AIMS: To evaluate the patterns of the production of antimicrobial compounds by Israeli myxobacteria newly isolated from soil samples and barks by a battery of isolation and purification methods. METHODS AND RESULTS: A total of 100 myxobacteria belonging to five of the 12 described genera, were isolated from 48 soil and 45 tree bark samples collected in different areas inside the State of Israel. Four isolation methods based on the peculiar metabolic and cell cycle aspects of myxobacteria, were combined with purification procedures and optimization of cultivation conditions. Ninety-seven strains were fermented and screened for antimicrobial activities. Production of antimicrobial activities was detected in 62 isolates. More than 50% of the collection (54 strains) was able to inhibit Escherichia coli growth. CONCLUSIONS: The results of this study support the idea that myxobacterial strains can be isolated from particular habitats and then cultivated and screened for their capacity to produce secondary metabolites endowed with antibacterial and antifungal activities. Myxovirescin, a typical poliketide myxobacterial antibiotic, has been identified in one Israeli isolate. Althiomycin, a thiazolyl peptide, which inhibits prokaryotic protein synthesis, usually produced by actinomycetes, was detected in three strains selected in this study. SIGNIFICANCE AND IMPACT OF THE STUDY: The results confirm that myxobacteria are prolific producers of a variety of bioactive secondary metabolites including antibacterial and antifungal compounds, being their high frequency of anti-Gram-negative activities particularly appealing for the current anti-infective research. So far their screening has often been hampered because their isolation is time-consuming and are quite difficult to handle and cultivate. In this paper we demonstrate that a proper combination of isolation, purification and cultivation methods allow their pharmaceutical exploitation. PMID- 15659198 TI - Comparison of the microbial population dynamics and phylogenetic characterization of a CANOXIS reactor and a UASB reactor degrading trichloroethene. AB - AIMS: To understand the microbial ecology underlying trichloethene (TCE) degradation in a coupled anaerobic/aerobic single stage (CANOXIS) reactor oxygenated with hydrogen peroxide (H2O2) and in an upflow anaerobic sludge bed (UASB) reactor. METHODS AND RESULTS: The molecular study of the microbial population dynamics and a phylogenetic characterization were conducted using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR-DGGE). In both reactors, TCE had a toxic effect on two uncultured bacterial populations whereas oxygen favoured the growth of aerobic species belonging to Rhizobiaceae and Dechloromonas. No methanotrophic bacteria were detected when targeting 16S rRNA gene with universal primers. Alternatively, pmo gene encoding the particulate methane monooxygenase of Methylomonas sp. LW21 could be detected in the coupled reactor when H2O2 was supplied at 0.7 g O2 l day(-1). CONCLUSIONS: Methylomonas sp. LW21 that could be responsible for the aerobic degradation of the TCE by-products is not among the predominant bacterial populations in the coupled reactor. It seems to have been outcompeted by heterotrophic bacteria (Rhizobiaceae and Dechloromonas sp.) for oxygen. SIGNIFICANCE AND IMPACT OF THE STUDY: The results obtained show the limitations of the coupled reactor examined in this study. Further investigations should focus on the operating conditions of this reactor in order to favour the growth of the methanotrophs. PMID- 15659199 TI - Evaluation of six commercial identification kits for the identification of Staphylococcus aureus isolated from bovine mastitis. AB - AIMS: Comparison of six commercially available in human medicine well-established slide agglutination systems for the identification of Staphylococcus aureus. METHODS AND RESULTS: Slide agglutination tests were compared with the conventional tube coagulase test, biochemical identification and with the molecular identification by polymerase chain reaction (PCR) amplification of species-specific parts of the gene encoding the 23S RNA. Systems evaluated included Masta-Staph (Mast Diagnostics), Staphylase-Test (Oxoid), Staphytect-Plus (Oxoid), Staphyloslide Latex (Becton Dickinson), Slidex Staph Plus (bioMerieux) and Dry Spot Staphytect Plus (Oxoid). A total of 141 staphylococcal strains isolated from cases of bovine mastitis including 90 S. aureus, 14 Staphylococcus epidermidis, 10 Staphylococcus warneri, 13 Staphylococcus xylosus, 11 Staphylococcus haemolyticus and three other coagulase-negative staphylococci were tested with each method. Staphylococcus aureus strains were selected by macrorestriction analysis with pulsed field gel electrophoresis (PFGE). Only genetically unrelated strains were included in the study. The sensitivities and specificities of the test were as follows: Masta-Staph 86.7 and 90.1%, Staphylase Test 78.4 and 85.1%, Staphytect-Plus 81.1 and 86.5%, Staphyloslide Latex 77.8 and 84.4%, Slidex Staph Plus 77.8 and 84.4%, Dry Spot Staphytect Plus 75.6 and 83.0%. CONCLUSIONS: The results of this evaluation suggest that the six slide agglutination methods tested can provide rapid identification of S. aureus also from bovine mastitis. The sensitivity and specificity seems to be less than those reported from human S. aureus isolates. SIGNIFICANCE AND IMPACT OF THE STUDY: This is one of the first comparative reported investigations about the applicability of different commercially available slide agglutination tests for the detection of S. aureus from bovine mastitis using PFGE selected clinical isolates. PMID- 15659200 TI - Production and characterization of a new bioemulsifier from Pseudomonas putida ML2. AB - AIMS: To characterize the bioemulsifier produced by a nonfluorescent strain of Pseudomonas putida isolated from a polluted sediment and to determine the influence of pH, temperature, media composition, and carbon and nitrogen source on growth and emulsifying activity. METHODS AND RESULTS: Different indexes were employed to determine the emulsifying properties of culture supernatants of P. putida ML2 in defined and complex media. Surface tension of cell-free supernatants was measured. Purification and chemical analysis of the emulsifier was performed. Confirmed results indicate that a polysaccharide with hexasaccharide repeating units is responsible for the emulsifying activity in a mineral medium with glucose as sole carbon source. Moreover, an emulsifier is produced when growing on naphthalene. CONCLUSIONS: Culture media composition influences the amount and the properties of the emulsifier produced by this P. putida strain. Under nitrogen limiting conditions, a polysaccharide is responsible for the emulsifying activity in defined mineral media. In complex nitrogen rich medium, a different kind of emulsifier is produced. The exopolymer may contribute to hydrocarbons solubilization. SIGNIFICANCE AND IMPACT OF THE STUDY: This is the first exopolysaccharide with emulsifying properties produced by a Pseudomonas strain reported to the present. Also chemical composition is significantly different from previous reports. This strain has potential use in bioremediation and the purified polysaccharide may be used in food and cosmetic industry. Moreover, the production of the exopolymer may play a role on biofilm formation. PMID- 15659201 TI - Effect of supplementing corn- or barley-based feedlot diets with canola oil on faecal shedding of Escherichia coli O157:H7 by steers. AB - AIMS: This study was conducted to evaluate the effect of supplementing barley- or corn-based diets with canola oil on faecal shedding of Escherichia coli O157:H7 by experimentally inoculated feedlot cattle. METHODS AND RESULTS: Four groups of yearling steers fed on barley- or corn-based feedlot diets containing 0% (BA; CO) or 6% canola oil (BA-O; CO-O) were inoculated with 10(10) CFU of a mixture of four nalidixic acid-resistant strains of E. coli O157:H7. The inoculated strains were tracked in oral (mouth swab) and environmental (water, water bowl interface, feed, faecal pat) samples by enrichment and immunomagnetic separation (IMS) for 12 weeks, and in rectally collected faecal samples for 23 weeks (enumeration by dilution plating for 12 weeks; detection by IMS for a further 11 weeks). Levels of E. coli O157:H7 shed in faecal samples over the course of the enumeration period were similar (P = 0.14) among treatments. Disappearance of the inoculated strains from faeces was more rapid (P = 0.009) with barley than with corn, but shedding levels at the end of the enumeration period were similar (P = 0.21) across grain types. Canola oil supplementation did not affect (P = 0.71) the rate of disappearance of E. coli O157:H7 from faeces. The numbers of steers culture positive for E. coli O157:H7 during the enumeration period were similar (P = 0.57) among treatments. During the 11-week detection period, however, more (P < 0.001) steers were E. coli O157:H7-positive in the BA group (15/64) than in BA-O (two of 64), CO (two of 56), or CO-O (one of 56). The organism was present in two of 48 water samples (both CO-O), one of 48 water bowl swabs (BA-O), four of 48 feed samples (two of 12 BA; two of 12 CO-O), 30 of 48 pen floor faecal pat samples, and 296 of 540 mouth swabs (81/144 BA, 80/144 BA-O, 74/126 CO and 61/126 CO-O). CONCLUSION: Supplementing corn or barley-based diets with canola oil did not affect shedding of E. coli O157:H7 by feedlot cattle. SIGNIFICANCE AND IMPACT OF THE STUDY: High-shedding individuals (i.e. 'super shedders') may be responsible for disseminating E. coli O157:H7 among penmates. Faeces on pen floors appears to be a more significant source of infection than are feed or drinking water. PMID- 15659202 TI - Link between spatial structure of microbial communities and degradation of a complex mixture of volatile organic compounds in peat biofilters. AB - AIMS: To investigate the relationships between the operation of the volatile organic compound (VOC) removal biofilter and the structure of microbial communities, and to study the impact on degradation activities and the structuring of microbial communities of biofilter malfunctions related to the qualitative composition of the polluted air. METHODS AND RESULTS: A microbiological study and a measurement of biodegradation activities were simultaneously carried out on two identical peat-packed columns, seeded with two different inocula, treating polluted air containing 11 VOCs. For both reactors, the spatial structure of the microbial communities was investigated by means of single-strand conformation polymorphism (SSCP) analysis. For both reactors, stratification of degradation activities in function of depth was observed. Oxygenated compounds were removed at the top of the column and aromatics at the bottom. Comparison of SSCP patterns clearly showed a shift in community structure in function of depth inside both biofilters. This distribution of biodegradation activities correlates with the spatialization of microbial density and diversity. Although the operating conditions of both reactors were identical and the biodegradation activities similar, the composition of microflora differed for biofilters A and B. Subdivision of biofilter B into two independent parts supplied with polluted air containing the complex VOC mixture showed that the microflora having colonized the bottom of biofilter B retained their potential for degrading oxygenated compounds. CONCLUSIONS: This work highlights the spatialization of biodegradation functions in a biofilter treating a complex mixture of VOCs. This distribution of biodegradation activities correlates with the spatialization of microbial density and diversity. SIGNIFICANCE AND IMPACT OF THE STUDY: This vertical structure of microbial communities must be taken into consideration when dealing with the malfunctioning of bioreactors. These results are also useful information about changes in microbial communities following natural or anthropogenic alterations in different ecosystems (soils and sediments) where structuring of microbial communities according to depth has been observed. PMID- 15659203 TI - Oxidant and SDS-stable alkaline protease from a halo-tolerant Bacillus clausii I 52: enhanced production and simple purification. AB - AIMS: An investigation was carried out on the enhancement of protease production and simple purification of an oxidant and SDS-stable alkaline protease produced by Bacillus clausii I-52 of industrial significance. METHODS AND RESULTS: The supplementation with 0.4% (w/v) NaCl and 0.05% (w/v) FeSO4.7H2O in a culture medium caused an increase in the protease production. The enzyme was purified to homogeneity with overall recovery of 79% and 10-fold purification from culture supernatant using Diaion HPA75, phenyl-Sepharose and DEAE-Sepharose column chromatographies. The protease was a halo-tolerant enzyme with apparent molecular mass of 28 kDa, and the Km and kcat values for N-Succinyl-Ala-Ala-Pro-Phe-pNA at 45 degrees C and pH 11.0 were determined to be 83.9 micromol l(-1) and 238.6 s( 1) respectively. CONCLUSIONS: Bacillus clausii I-52 was identified as a halo tolerant bacterium, and the extracellular alkaline protease produced by B. clausii I-52 also showed extreme halo-tolerance. The enzyme stability towards SDS and H2O2 could be increased by adding NaCl or propylene glycol to the enzyme solution. SIGNIFICANCE AND IMPACT OF THE STUDY: The alkaline protease secreted by B. clausii I-52 is significant from an industrial perspective because of its stability against surfactants and oxidants as well as its tolerance towards high salinity. These enzymatic properties suggest its suitable application for industrial purposes. PMID- 15659204 TI - Destruction and injury of Escherichia coli during microwave heating under vacuum. AB - AIMS: To study the effect of 2450 MHz microwave radiation under vacuum (vacuum microwave or VM) on survival and injury of Escherichia coli and to search for possible nonthermal effects associated with VM. METHODS AND RESULTS: Destruction kinetics of E. coli in peptone water were determined in a continuous-flow vacuum system, heated by convection heating in a water bath or with microwaves (VMs). Vacuum was used to control the boiling point of water and to maintain temperature in the bacterial suspensions at specified levels (49-64 degrees C). CONCLUSIONS: z-Value in the water bath treatment was 9.1 degrees C while for VM at 510 and 711 W it was 6.2 and 5.9 degrees C, suggesting that E. coli is more sensitive to temperature changes under microwave heating. Arrhenius calculations of the activation energies of the destruction reactions suggest that the mechanism of destruction in VM may be different from that of conventional heat. The number of injured micro-organisms showed no significant differences among treatments. SIGNIFICANCE AND IMPACT OF THE STUDY: The impact of temperature on E. coli destruction was different when microwaves were the medium of heat transfer, suggesting the existence of factors other than heat contributing to the lethal effect of VM. PMID- 15659205 TI - The effects of the presence of stored product pests on the microfauna of a flour community. AB - AIMS: To determine the effect that the presence of some beetles have on the species of bacteria found in their flour. METHODS AND RESULTS: Bacteria were isolated from flour that either did not contain beetles, contained Tribolium beetles in different environments, or contained either Stegobium paniceum or Lasioderma serricorne. These bacteria were tentatively identified by both the gas chromatography-fatty acid methyl esters (GC-FAME) method and partial sequencing of the 16S rRNA gene. All samples contained Bacillus species including the controls, but the non-Tribolium beetles and a Tribolium beetle line known to have low benzoquinones also contained Enterococcus and Enterobacter species. Additionally an unidentified bacteria isolate in the Enterobacteriaceae was also found in the L. serricorne sample. Our results also suggest incongruent identifications when using the GC-FAME method vs sequencing. CONCLUSIONS: Certain species of bacteria can be introduced by the presence of insect pests, but the diversity of species is far less in stocks of Tribolium beetles. SIGNIFICANCE AND IMPACT OF THE STUDY: Stored product pests can alter the bacterial community. Isolated species from this study show a strong genetic relationship to each other, suggesting an isolated evolving system. A unique bacteria was also isolated. GC and sequencing methods of identification are compared. PMID- 15659206 TI - Comparison of bacteriophage and enteric virus removal in pilot scale activated sludge plants. AB - AIMS: The aim of this experimental study was to determine comparatively the removal of two types of bacteriophages, a somatic coliphage and an F-specific RNA phage and of three types of enteric viruses, hepatitis A virus (HAV), poliovirus and rotavirus during sewage treatment by activated sludge using laboratory pilot plants. METHODS AND RESULTS: The cultivable simian rotavirus SA11, the HAV HM 175/18f cytopathic strain and poliovirus were quantified by cell culture. The bacteriophages were quantified by plaque formation on the host bacterium in agar medium. In each experiment, two pilots simulating full-scale activated sludge plants were inoculated with viruses at known concentrations, and mixed liquor and effluent samples were analysed regularly. In the mixed liquor, liquid and solid fractions were analysed separately. The viral behaviour in both the liquid and solid phases was similar between pilots of each experiment. Viral concentrations decreased rapidly following viral injection in the pilots. Ten minutes after the injections, viral concentrations in the liquid phase had decreased from 1.0 +/- 0.4 log to 2.2 +/- 0.3 log. Poliovirus and HAV were predominantly adsorbed on the solid matters of the mixed liquor while rotavirus was not detectable in the solid phase. In our model, the estimated mean log viral reductions after 3-day experiment were 9.2 +/- 0.4 for rotavirus, 6.6 +/- 2.4 for poliovirus, 5.9 +/- 3.5 for HAV, 3.2 +/- 1.2 for MS2 and 2.3 +/- 0.5 for PhiX174. SIGNIFICANCE AND IMPACT OF THE STUDY: This study demonstrates that the pilots are useful models to assess the removal of infectious enteric viruses and bacteriophages by activated sludge treatment. Our results show the efficacy of the activated sludge treatment on the five viruses and suggest that coliphages could be an acceptable indicator of viral removal in this treatment system. PMID- 15659207 TI - Flocculation onset in Saccharomyces cerevisiae: the role of nutrients. AB - AIMS: To examine the role of the nutrients on the onset of flocculation in an ale brewing strain, Saccharomyces cerevisiae NCYC 1195. METHODS AND RESULTS: Flocculation was evaluated using the method of Soares, E.V. and Vroman, A. [Journal of Applied Microbiology (2003) 95, 325]. For cells grown in chemically defined medium (yeast nitrogen base with glucose) or in rich medium (containing yeast extract, peptone and fermentable sugars: fructose or maltose), the onset of flocculation occurred after the end of exponential respiro-fermentative phase of growth being coincident with the attainment of the lower level of carbon source in the culture medium. Cells, in exponential respiro-fermentative phase of growth, transferred to a glucose-containing medium without nitrogen source, developed a flocculent phenotype, while these carbon source starved cells, in the presence of all other nutrients that support growth, did not flocculate. In addition, cells in exponential phase of growth, under catabolite repression, when transferred to a medium containing 0.2% (w/v) of fermentable sugar (fructose or maltose) or 2% (v/v) ethanol, showed a rapid triggering of flocculation, while when incubated in 2% (v/v) glycerol did not develop a flocculent phenotype. CONCLUSIONS: The onset of flocculation occurs when a low sugar and/or nitrogen concentration is reached in culture media. The triggering of flocculation is an energetic dependent process influenced by the carbon source metabolism. The presence of external nitrogen source is not necessary for developing a flocculent phenotype. SIGNIFICANCE AND IMPACT OF THE STUDY: This work contributes to the elucidation of the role of nutrients on the onset of flocculation in NewFlo phenotype yeast strains. This information might be useful to the brewing industry, in the control of yeast flocculation, as the time when the onset of flocculation occurs can determine the fermentation performance and the beer quality. PMID- 15659208 TI - Activation of the PI3K/Akt signal transduction pathway and increased levels of insulin receptor in protein repair-deficient mice. AB - Protein L-isoaspartate (D-aspartate) O-methyltransferase is an enzyme that catalyses the repair of isoaspartyl damage in proteins. Mice lacking this enzyme (Pcmt1-/- mice) have a progressive increase in brain size compared with wild-type mice (Pcmt1+/+ mice), a phenotype that can be associated with alterations in the PI3K/Akt signal transduction pathway. Here we show that components of this pathway, including Akt, GSK3beta and PDK-1, are more highly phosphorylated in the brains of Pcmt1-/- mice, particularly in cells of the hippocampus, in comparison with Pcmt1+/+ mice. Examination of upstream elements of this pathway in the hippocampus revealed that Pcmt1-/- mice have increased activation of insulin-like growth factor-I (IGF-I) receptor and/or insulin receptor. Western blot analysis revealed an approximate 200% increase in insulin receptor protein levels and an approximate 50% increase in IGF-I receptor protein levels in the hippocampus of Pcmt1-/- mice. Higher levels of the insulin receptor protein were also found in other regions of the adult brain and in whole tissue extracts of brain, liver, heart and testes of both juvenile and adult Pcmt1-/- mice. There were no significant differences in plasma insulin levels for adult Pcmt1-/- mice during glucose tolerance tests. However, they did show higher peak levels of blood glucose, suggesting a mild impairment in glucose tolerance. We propose that Pcmt1 /- mice have altered regulation of the insulin pathway, possibly as a compensatory response to altered glucose uptake or metabolism or as an adaptive response to a general accumulation of isoaspartyl protein damage in the brain and other tissues. PMID- 15659209 TI - Biodemographic analysis of male honey bee mortality. AB - Biodemographic studies of insects have significantly enhanced our understanding of the biology of aging. Eusocial insects have evolved to form different groups of colony members that are specialized for particular tasks and highly dependent on each other. These different groups (castes and sexes) also differ strongly in their life expectancy but relatively little is known about their mortality dynamics. In this study we present data on the age-specific flight activity and mortality of male honey bees from two different genetic lines that are exclusively dedicated to reproduction. We show that males initiating flight at a young age experience more flight events during their lifetime. No (negative) relation between the age at flight initiation and lifespan exists, as might be predicted on the basis of the antagonistic pleiotropy theory of aging. Furthermore, we fit our data to different aging models and conclude that overall a slight deceleration of the age-dependent mortality increase at advanced ages occurs. However, mortality risk increases according to the Gompertz-Makeham model when only days with flight activity (active days) are taken into account. Our interpretation of the latter is that two mortality components act on honey bee males during flight: increasing, age-dependent deaths (possibly from wear-and tear), and age-independent deaths (possibly due to predation). The overall mortality curve is caused by the interaction of the distribution of age at foraging initiation and the mortality function during the active (flight) lifespan. PMID- 15659210 TI - Telomere-independent cellular senescence in human fetal cardiomyocytes. AB - Fetal cardiomyocytes have been proposed as a potential source of cell-based therapy for heart failure. This study examined cellular senescence in cultured human fetal ventricular cardiomyocytes (HFCs). HFCs were isolated and identified by immunocytochemistry and RT-PCR. Cells were found to senesce after 20-25 population doublings, as determined by growth arrest, morphological changes and senescence-associated beta-galactosidase activity. Using the telomeric repeat amplification protocol assay, telomerase activity was undetectable in primary HFCs. Cells were transduced to express the human reverse transcriptase subunit (hTERT) of telomerase. This resulted in greatly increased telomerase activity, but no significant lifespan extension. Analysis of telomere length in primary HFCs revealed that the senescent phenotype was not accompanied by telomere shortening. Telomeres in hTERT-positive cells were elongated in comparison with primary cells, and elongation was retained in senescent cells. Levels of the tumor suppressor protein p16INK4A increased in all senescent cells whether telomerase-positive or -negative. Senescence was accompanied by a decline in transcript levels of the polycomb gene Bmi-1, Ets1 and Ets2 transcription factors, and Id1, Id2 and Id3 helix-loop-helix proteins, suggesting roles for these genes in maintenance of cardiomyocyte proliferative capacity. In addition to offering novel insights into the behavior of human fetal cardiomyocytes in culture, these findings have implications for the development of a cell-based therapy for cardiac injury using primary fetal heart tissue. PMID- 15659211 TI - Stochastic dietary restriction using a Markov-chain feeding protocol elicits complex, life history response in medflies. AB - Lifespan in individually housed medflies (virgins of both sexes) and daily reproduction for females were studied following one of 12 dietary restriction (DR) treatments in which the availability of high-quality food (yeast-sugar mixture) for each fly was based on a Markov chain feeding scheme--a stochastic dietary regime which specifies that the future dietary state depends only on the present dietary state and not on the path by which the present state was achieved. The stochastic treatments consisted of a combination of one of four values of a 'discovery' parameter and one of three values of a 'persistence' parameter. The results supported the hypotheses that: (i) longevity is extended in most medfly cohorts subject to stochastic DR; and (ii) longevity is more affected by the patch discovery than the patch persistence parameter. One of the main conclusions of the study is that, in combination with the results of earlier dietary restriction studies on the medfly, the results reinforce the concept that the details of the dietary restriction protocols have a profound impact on the sign and magnitude of the longevity extension relative to ad libitum cohorts and that a deeper understanding of the effect of food restriction on longevity is not possible without an understanding of its effect on reproduction. PMID- 15659212 TI - Overexpressed Sod1p acts either to reduce or to increase the lifespans and stress resistance of yeast, depending on whether it is Cu(2+)-deficient or an active Cu,Zn-superoxide dismutase. AB - Yeast overexpressing SOD1, the gene for Cu,Zn-superoxide dismutase (Cu,Zn-Sod), was used to determine how Sod1p overexpression influences the chronological lifespan [the survival of non-dividing stationary (G0) phase cells over time], the replicative lifespan (the number of buds produced by actively dividing yeast cells) and stress resistance. Increasing the level of active Cu,Zn-Sod in yeast was found to require either growth in the presence of high copper, or the simultaneous overexpression of both SOD1 and CCS1 (the latter being the gene that encodes the chaperone dedicated to Cu(2+)-loading of Sod1p in vivo). Dual SOD1 + CCS1 overexpression elevated the levels of Cu,Zn-Sod activity six- to eight-fold in vegetative cultures. It also increased the optimized survival of stationary cells up to two-fold, showing this chronological lifespan is ultimately limited by oxidative stress. In contrast, several detrimental effects resulted when the SOD1 gene was overexpressed in the absence of either high copper or a simultaneous overexpression of CCS1. Both the chronological and the replicative lifespans were shortened; the cells displayed an abnormally high level of endogenous oxidative stress, resulting in a high rate of spontaneous mutation. Such harmful effects were all reversed through the overexpression of CCS1. It is apparent therefore that they relate to the incomplete Cu(2+)-loading of the overexpressed Sod1p, most probably accumulation of a Cu(2+)-deficient Sod1p to appreciable levels in vivo. The same events may generate the detrimental effects that are frequently, though not universally, observed when Cu,Zn-Sod overexpression is attempted in metazoans. PMID- 15659213 TI - Drosophila longevity is not affected by heterochromatin-mediated gene silencing. AB - Two highly conserved histone deacetylases, Sir2 and Rpd3, have been linked to caloric restriction and the extension of longevity. Because the Drosophila forms of each protein can silence genes in either euchromatin or heterochromatin, we determined whether longevity extension is mediated by silencing in the latter domain. When silencing was increased and decreased using mutations that affect heterochromatin protein 1 (HP1), but have no direct effect upon Sir2 or Rpd3, lifespan was unaffected. Heterochromatin-mediated gene silencing was then modulated without directly influencing HP1 as well as the deacetylases, again yielding no effect on lifespan. Mortality rates were unchanged by all manipulations, indicating that euchromatic targets are likely to be the effectors of deacetylase-mediated longevity extension in Drosophila [corrected] PMID- 15659214 TI - Co-localization and functional interaction between adenosine A(2A) and metabotropic group 5 receptors in glutamatergic nerve terminals of the rat striatum. AB - The anti-Parkinsonian effect of glutamate metabotropic group 5 (mGluR5) and adenosine A(2A) receptor antagonists is believed to result from their ability to postsynaptically control the responsiveness of the indirect pathway that is hyperfunctioning in Parkinson's disease. mGluR5 and A(2A) antagonists are also neuroprotective in brain injury models involving glutamate excitotoxicity. Thus, we hypothesized that the anti-Parkinsonian and neuroprotective effects of A(2A) and mGluR5 receptors might be related to their control of striatal glutamate release that actually triggers the indirect pathway. The A(2A) agonist, CGS21680 (1-30 nM) facilitated glutamate release from striatal nerve terminals up to 57%, an effect prevented by the A(2A) antagonist, SCH58261 (50 nM). The mGluR5 agonist, CHPG (300-600 mum) also facilitated glutamate release up to 29%, an effect prevented by the mGluR5 antagonist, MPEP (10 microm). Both mGluR5 and A(2A) receptors were located in the active zone and 57 +/- 6% of striatal glutamatergic nerve terminals possessed both A(2A) and mGluR5 receptors, suggesting a presynaptic functional interaction. Indeed, submaximal concentrations of CGS21680 (1 nM) and CHPG (100 microm) synergistically facilitated glutamate release and the facilitation of glutamate release by 10 nM CGS21680 was prevented by 10 microm MPEP, whereas facilitation by 300 microm CHPG was prevented by 10 nM SCH58261. These results provide the first direct evidence that A(2A) and mGluR5 receptors are co-located in more than half of the striatal glutamatergic terminals where they facilitate glutamate release in a synergistic manner. This emphasizes the role of the modulation of glutamate release as a likely mechanism of action of these receptors both in striatal neuroprotection and in Parkinson's disease. PMID- 15659216 TI - Increased expression of phospholipase D1 in the brains of scrapie-infected mice. AB - Mitochondrial dysfunction and free radical-induced oxidative damage are critical factors in the pathogenesis of neurodegenerative diseases. Recently, phospholipid breakdown by phospholipase D (PLD) has been recognized as an important signalling pathway in the nervous system. Here, we examined the expression of PLD and alteration of membrane phospholipid in scrapie brain. We have found that protein expression and enzyme activity of PLD1 were increased in scrapie brains compared with controls; in particular, there was an increase in the mitochondrial fraction. PLD1 in mitochondrial membranes from scrapie brains, but not from control brains, was tyrosine phosphorylated. Furthermore, the concentration of mitochondrial phospholipids such as phosphatidylcholine and phosphatidylethanolamine was increased and the content of phosphatidic acid, a product of PLD activity, was up-regulated in the mitochondrial membrane fractions. Immunohistochemically, PLD1 immunoreactivity was significantly increased in activated astrocytes in both cerebral cortex and hippocampus of scrapie brains. Taken together, these results suggest that PLD activation might induce alterations in mitochondrial lipids and, in turn, mediate mitochondrial dysfunction in the brains of scrapie-infected mice. PMID- 15659215 TI - Neuronal calcium sensor-1 facilitates neuronal exocytosis through phosphatidylinositol 4-kinase. AB - This work tested the theory that neuronal calcium sensor-1 (NCS-1) has effects on neurotransmitter release beyond its actions on membrane channels. We used nerve ending preparations where membrane channels are bypassed through membrane permeabilization made by mechanical disruption or streptolysin-O. Nerve ending NCS-1 and phosphatidylinositol 4-kinase (PI4K) are largely or entirely particulate, so their concentrations in nerve endings remain constant after breaching the membrane. Exogenous, myristoylated NCS-1 stimulated nerve ending phosphatidylinositol 4-phosphate [PI(4)P] synthesis, but non-myristoylated-NCS-1 did not. The N-terminal peptide of NCS-1 interfered with PI(4)P synthesis, and with spontaneous and Ca(2+)-evoked release of both [(3)H]-norepinephrine (NA) and [(14)C]-glutamate (glu) in a concentration-dependent manner. An antibody raised against the N-terminal of NCS-1 inhibited perforated nerve ending PI(4)P synthesis, but the C-terminal antibody had no effects. Antibodies against the N- and C-termini of NCS-1 caused significant increases in mini/spontaneous/stimulation-independent release of [(3)H]-NA from perforated nerve endings, but had no effect on [(14)C]-glu release. These results support the idea that NCS-1 facilitates nerve ending neurotransmitter release and phosphoinositide production via PI4K and localizes these effects to the N terminal of NCS-1. Combined with previous work on the regulation of channels by NCS-1, the data are consistent with the hypothesis that a NCS-1-PI4K (NP, neuropotentiator) complex may serve as an essential linker between lipid and protein metabolism to regulate membrane traffic and co-ordinate it with ion fluxes and plasticity in the nerve ending. PMID- 15659217 TI - Rotenone-induced caspase 9/3-independent and -dependent cell death in undifferentiated and differentiated human neural stem cells. AB - We used human neural stem cells (hNSCs) and their differentiated cultures as a model system to evaluate the mechanism(s) involved in rotenone (RO)- and camptothecin (CA)-induced cytotoxicity. Results from ultrastructural damage and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining indicated that RO-induced cytotoxicity resembled CA-induced apoptosis more than H(2)O(2)-induced necrosis. However, unlike CA-induced, caspase 9/3 dependent apoptosis, there was no increased activity in caspase 9, caspase 3 or poly (ADP-ribose) polymerase (PARP) cleavage in RO-induced cytotoxicity, in spite of time-dependent release of cytochrome c and apoptosis-inducing factor (AIF) following mitochondrial membrane depolarization and a significant increase in reactive oxygen species generation. Equal doses of RO and CA used in hNSCs induced caspase 9/3-dependent apoptosis in differentiated cultures. Time dependent ATP depletion occurred earlier and to a greater extent in RO-treated hNSCs than in CA-treated hNSCs, or differentiated cultures treated with RO or CA. In conclusion, these results represent a unique ultrastructural and molecular characterization of RO- and CA-induced cytotoxicity in hNSCs and their differentiated cultures. Intracellular ATP levels may play an important role in determining whether neural progenitors or their differentiated cells follow a caspase 9/3-dependent or -independent pathway in response to acute insults from neuronal toxicants. PMID- 15659218 TI - Prostaglandin D2 mediates neuronal protection via the DP1 receptor. AB - Cyclo-oxygenases (COXs) catalyze the first committed step in the synthesis of the prostaglandins PGE(2), PGD(2), PGF(2alpha), PGI(2) and thomboxane A(2). Expression and enzymatic activity of COX-2, the inducible isoform of COX, are observed in several neurological diseases and result in significant neuronal injury. The neurotoxic effect of COX-2 is believed to occur through downstream effects of its prostaglandin products. In this study, we examined the function of PGD(2) and its two receptors DP1 and chemoattractant receptor-homologous molecule expressed on Th2 cells (CRTH2) (DP2) in neuronal survival. PGD(2) is the most abundant prostaglandin in brain and regulates sleep, temperature and nociception. It signals through two distinct G protein-coupled receptors, DP1 and DP2, that have opposing effects on cyclic AMP (cAMP) production. Physiological concentrations of PGD(2) potently and unexpectedly rescued neurons in paradigms of glutamate toxicity in cultured hippocampal neurons and organotypic slices. This effect was mimicked by the DP1-selective agonist BW245C but not by the PGD(2) metabolite 15d-PGJ(2), suggesting that neuroprotection was mediated by the DP1 receptor. Conversely, activation of the DP2 receptor promoted neuronal loss. The protein kinase A inhibitors H89 and KT5720 reversed the protective effect of PGD(2), indicating that PGD(2)-mediated neuroprotection was dependent on cAMP signaling. These studies indicate that activation of the PGD(2) DP1 receptor protects against excitotoxic injury in a cAMP-dependent manner, consistent with recent studies of PGE(2) receptors that also suggest a neuroprotective effect of prostaglandin receptors. Taken together, these data support an emerging and paradoxical neuroprotective role of prostaglandins in the CNS. PMID- 15659219 TI - Neuronal death in primary retinal cultures is related to nitric oxide production, and is inhibited by erythropoietin in a glucose-sensitive manner. AB - The aim of this work was to investigate the interrelated effects of glucose, nitric oxide (NO) and erythropoietin on neuronal survival in retinal cultures, thereby exploring the mechanism of neuronal death in the diabetic retina. Rat retinal cells were cultured in low (5 mM) or high (15 mM) glucose concentrations. After 9 days, cell viability was assessed by (3,4,5-dimethylthiazol-2-yl)-2,5 diphenyl tetrazolium bromide (MTT) assay and NO production was determined by the Griess reaction. Immunohistochemistry was used to quantify GABA-labelled neurones and cells staining for DNA breakdown. High or low glucose concentrations had no effect on basal NO production or the survival of neurones in culture, but treatment with N-nitro-L-arginine methyl ester reduced extracellular levels of NO and increased neuronal survival at both concentrations of glucose. Erythropoietin decreased cell death and NO levels, but only in cultures grown in low concentrations of glucose. It is concluded that erythropoietin's neurotrophic function in the retina is attenuated at glucose concentrations similar to those which occur in diabetes. PMID- 15659220 TI - Time-course of mitochondrial gene expressions in mice brains: implications for mitochondrial dysfunction, oxidative damage, and cytochrome c in aging. AB - The study of aging is critical for a better understanding of many age-related diseases. The free radical theory of aging, one of the prominent aging hypotheses, holds that during aging, increasing reactive oxygen species in mitochondria causes mutations in the mitochondrial DNA and damages mitochondrial components, resulting in senescence. Understanding a mitochondrial gene expression profile and its relationship to mitochondrial function becomes an important step in understanding aging. The objective of the present study was to determine mRNA expression of mitochondrial-encoded genes in brain slices from C57BL6 mice at four ages (2, 12, 18, and 24 months) and to determine how these altered mitochondrial genes influence age-related changes, including oxidative damage and cytochrome c in apoptosis. Using northern blot analysis, in situ hybridization, and immunofluorescence analyses, we analyzed changes in the expression of mitochondrial RNA encoding the mitochondrial genes, oxidative damage marker, 8-hydroxyguanosine (8-OHG), and cytochrome c in brain slices from the cortex of C57BL6 mice at each of the four ages. Our northern blot analysis revealed an increased expression of mitochondrial-encoded genes in complexes I, III, IV, and V of the respiratory chain in 12- and 18-month-old C57BL6 mice compared to 2-month-old mice, suggesting a compensatory mechanism that allows the production of proteins involved in the electron transport chain. In contrast to the up-regulation of mitochondrial genes in 12- and 18-month-old C57BL6 mice, mRNA expression in 24-month-old C57BL6 mice was decreased, suggesting that compensation maintained by the up-regulated genes cannot be sustained and that the down-regulation of expression results in the later stage of aging. Our in situ hybridization analyses of mitochondrial genes from the hippocampus and the cortex revealed that mitochondrial genes were over-expressed, suggesting that these brain areas are critical for mitochondrial functions. Our immunofluorescence analysis of 8-OHG and cytochrome c revealed increased 8-OHG and cytochrome c in 12-month-old C57BL6 mice, suggesting that age-related mitochondrial oxidative damage and apoptosis are associated with mitochondrial dysfunction. Our double-labeling analysis of in situ hybridization of ATPase 6 and our immunofluorescence analysis of 8-OHG suggest that specific neuronal populations undergo oxidative damage. Further, double-labeling analysis of in situ hybridization of ATPase 6 and immunofluorescence analysis of cytochrome c suggest cytochrome c release is related to mitochondrial dysfunction in the aging C57BL6 mouse brain. This study also suggests that these mitochondrial gene expression changes may relate to the role of mitochondrial dysfunction, oxidative damage, and cytochrome c in aging and in age-related diseases such as Alzheimer's disease and Parkinson's disease. PMID- 15659222 TI - C-terminal synaptic targeting elements for postsynaptic density proteins ProSAP1/Shank2 and ProSAP2/Shank3. AB - Synapses are specialized contact sites mediating communication between neurons. Synaptogenesis requires the specific assembly of protein clusters at both sides of the synaptic contact by mechanisms that are barely understood. We studied the synaptic targeting of multi-domain proteins of the ProSAP/Shank family thought to serve as master scaffolding molecules of the postsynaptic density. In contrast to Shank1, expression of green-fluorescent protein (GFP)-tagged ProSAP1/Shank2 and ProSAP2/Shank3 deletion constructs in hippocampal neurons revealed that their postsynaptic localization relies on the integrity of the C-termini. The shortest construct that was perfectly targeted to synaptic sites included the last 417 amino acids of ProSAP1/Shank2 and included the C-terminal sterile alpha motif (SAM) domain. Removal of 54 residues from the N-terminus of this construct resulted in a diffuse distribution in the cytoplasm. Altogether, our data delineate a hitherto unknown targeting signal in both ProSAP1/Shank2 and ProSAP2/Shank3 and provide evidence for an implication of these proteins and their close homologue, Shank1, in distinct molecular pathways. PMID- 15659221 TI - AG490 prevents cell death after exposure of rat astrocytes to hydrogen peroxide or proinflammatory cytokines: involvement of the Jak2/STAT pathway. AB - Janus kinases/STAT pathway mediates cellular responses to certain oxidative stress stimuli and cytokines. Here we examine the activation of Stat1 and Stat3 in rat astrocyte cultures and its involvement in cell death. H(2)O(2), interferon (INF)-gamma and interleukin (IL)-6 but not IL-10 caused cell death. Stat1 was phosphorylated on tyrosine (Tyr)-701 after exposure to H(2)O(2), INF-gamma or IL 6 but not IL-10. Tyr-705 pStat3 was observed after H(2)O(2), IL-6 and IL-10. Also, H(2)O(2) induced serine (Ser)-727 phosphorylation of Stat1 but not Stat3. The degree of Tyr-701 pStat1 by the different treatments positively correlated with the corresponding reduction of cell viability. AG490, a Jak2 inhibitor, prevented Tyr-701 but not Ser-727, Stat1 phosphorylation. Also, AG490 inhibited Tyr-705 Stat3 phosphorylation induced by H(2)O(2) and IL-6 but did not prevent that induced by IL-10. Furthermore, AG490 conferred strong protection against cell death induced by INF-gamma, IL-6 and H(2)O(2). These results suggest that Jak2/Stat1 activation mediates cell death induced by proinflammatory cytokines and peroxides. However, we found evidence suggesting that AG490 reduces oxidative stress induced by H(2)O(2), which further shows that H(2)O(2) and/or derived reactive oxygen species directly activate Jak2/Stat1, but masks the actual involvement of this pathway in H(2)O(2)-induced cell death. PMID- 15659223 TI - Bi-functional effects of ATP/P2 receptor activation on tumor necrosis factor alpha release in lipopolysaccharide-stimulated astrocytes. AB - Neuroinflammation is associated with a variety of CNS pathologies. Levels of tumor necrosis factor-alpha (TNF-alpha), a major proinflammatory cytokine, as well as extracellular ATP, are increased following various CNS insults. Here we report on the relationship between ATP/P2 purinergic receptor activation and lipopolysaccharide (LPS)-induced TNF-alpha release from primary cultures of rat cortical astrocytes. Using ELISA, we confirmed that treatment with LPS stimulated the release of TNF-alpha in a concentration and time dependent manner. ATP treatment alone had no effect on TNF-alpha release. LPS-induced TNF-alpha release was attenuated by 1 mm ATP, a concentration known to activate P2X7 receptors. Consistent with this, 3'-O-(4-Benzoyl)benzoyl-ATP (BzATP), a P2X7 receptor agonist, also attenuated LPS-induced TNF-alpha release. This reduction in TNF alpha release was not due to loss of cell viability. Adenosine and 2 chloroadenosine were ineffective, suggesting that attenuation of LPS-induced TNF alpha release by ATP was not due to ATP breakdown and subsequent activation of adenosine/P1 receptors. Interestingly, treatment of astrocyte cultures with 10 microm or 100 microm ATP potentiated TNF-alpha release induced by a submaximal concentration of LPS. UTP and 2methylthioADP (2-MeSADP), P2Y receptor agonists, also enhanced this LPS-induced TNF-alpha release. Our observations demonstrate opposing effects of ATP/P2 receptor activation on TNF-alpha release, i.e. P2X receptor activation attenuates, whereas P2Y receptor activation potentiates TNF alpha release in LPS-stimulated astrocytes. These observations suggest a mechanism whereby astrocytes can sense the severity of damage in the CNS via ATP release from damaged cells and can modulate the TNF-alpha mediated inflammatory response depending on the extracellular ATP concentration and corresponding type of astrocyte ATP/P2 receptor activated. PMID- 15659224 TI - Neuroadaptations of total levels of adenylate cyclase, protein kinase A, tyrosine hydroxylase, cdk5 and neurofilaments in the nucleus accumbens and ventral tegmental area do not correlate with expression of sensitized or tolerant locomotor responses to cocaine. AB - Neuroadaptations induced by high-dose cocaine treatment have been hypothesized to persist after the cessation of drug treatment and mediate the expression of sensitization and tolerance to cocaine. We looked for evidence of these neuroadaptations in rats receiving more modest behaviorally effective cocaine treatments. Rats were exposed to either a sensitizing regimen of seven once-daily injections of 15 mg/kg cocaine or a tolerance-producing regimen involving a continuous infusion of the same daily dose. We assessed enzyme activity levels of protein kinase A and adenylate cyclase, and protein levels of tyrosine hydroxylase, cdk5 and neurofilaments in the nucleus accumbens and ventral tegmental area. Only protein kinase A activity levels were altered by cocaine treatment, but this alteration persisted for only 7 days, whereas a sensitized locomotor response was still evident at 21 days. Although behavioral tolerance to cocaine was seen the day after the termination of treatment, none of the molecular measures was altered on this or any other day. Thus, although increased protein kinase A activity can temporarily modulate sensitized responses to cocaine, alterations in total levels of the molecules assessed in our study do not correlate with the expression of sensitized or tolerant locomotor responses to cocaine. PMID- 15659225 TI - S100B-induced microglial and neuronal IL-1 expression is mediated by cell type specific transcription factors. AB - Both the astrocytic cytokine S100B and the pro-inflammatory interleukin-1 (IL-1) are elevated in Alzheimer's disease, and each has been implicated in Alzheimer related neuropathology. We examined the gene-regulatory events through which S100B induces IL-1beta expression. In primary microglia, S100B activated the transcription factors Sp1 and NFkappaB, followed by an increase in IL-1beta mRNA levels. The latter was blocked by a peptide inhibitor of NFkappaB or by a double stranded oligonucleotide containing a NFkappaB-binding site to serve as "decoy" DNA and reduce available NFkappaB. But in primary cortical neurons, decoy and siRNA experiments indicated that the IL-1beta induction by S100B was mediated by Sp1 without evidence of a role for NFkappaB. Our results suggest that the elevation of S100B and IL-1 in Alzheimer brain and consequent neurodegenerative events are mediated through cell-type specific gene-regulatory events, providing mechanistic insight into connections between glial activation and neuronal dysfunction. PMID- 15659226 TI - Two distinct genes drive expression of seven tomosyn isoforms in the mammalian brain, sharing a conserved structure with a unique variable domain. AB - Tomosyn was previously identified as a syntaxin-binding protein that inhibits soluble NSF (n-ethylmaleimide-sensitive fusion protein) attachment protein receptor (SNARE)-mediated secretion. We set out to investigate the distribution of tomosyn mRNA in the mammalian brain and found evidence for the presence of two paralogous genes designated tomosyn-1 and -2. In a collection of tomosyn-2 cDNA clones, we observed four splice variants (named xb-, b-, m- and s-tomosyn-2) derived from the skipping of exons 19 and 21. This feature is conserved with tomosyn-1 that encodes three splice variants. To compare the expression pattern of tomosyn-1 and -2, we performed in situ hybridization experiments with gene specific probes. Both genes were expressed in the nervous system, clearly following distinct spatial and developmental expression patterns. Real-time quantitative PCR experiments indicated that tomosyn-1 expression was up-regulated less than threefold between developmental stages E10 and P12, whereas tomosyn-2 expression increased 31-fold. Not only the transcription level, but also the splice composition of tomosyn-2 mRNA shifted during development. We conclude that two distinct genes drive expression of seven tomosyn isoforms. Their expression patterns support a role in regulating neuronal secretion. All isoforms share conserved WD40 and SNARE domains separated by a hypervariable module, the function of which remains to be clarified. PMID- 15659227 TI - Age-dependent sensitivity of the developing brain to irradiation is correlated with the number and vulnerability of progenitor cells. AB - In a newly established model of unilateral, irradiation (IR)-induced injury we compared the outcome after IR to the immature and juvenile brain, using rats at postnatal days 9 or 23, respectively. We demonstrate that (i) the immature brains contained more progenitors in the subventricular zone (SVZ) and subgranular zone (SGZ) compared with the juvenile brains; (ii) cellular injury, as judged by activation of caspase 3 and p53, as well as nitrotyrosine formation, was more pronounced in the SVZ and SGZ in the immature brains 6 h after IR; (iii) the number of progenitor and immature cells in the SVZ and SGZ decreased 6 h and 7 days post-IR, corresponding to acute and subacute effects in humans, respectively, these effects were more pronounced in immature brains; (iv) myelination was impaired after IR at both ages, and much more pronounced after IR to immature brains; (v) the IR-induced changes remained significant for at least 10 weeks, corresponding to late effects in humans, and were most pronounced after IR to immature brains. It appears that IR induces both an acute loss of progenitors through apoptosis and a perturbed microenvironment incompatible with normal proliferation and differentiation, and that this is more pronounced in the immature brain. PMID- 15659229 TI - Synaptopodin, a molecule involved in the formation of the dendritic spine apparatus, is a dual actin/alpha-actinin binding protein. AB - Synaptopodin (SYNPO) is a cytoskeletal protein that is preferentially located in mature dendritic spines, where it accumulates in the spine neck and closely associates with the spine apparatus. Formation of the spine apparatus critically depends on SYNPO. To further determine its molecular action, we screened for cellular binding partners. Using the yeast two-hybrid system and biochemical assays, SYNPO was found to associate with both F-actin and alpha-actinin. Ectopic expression of SYNPO in neuronal and non-neuronal cells induced actin aggregates, thus confirming a cytoplasmic interaction with the actin cytoskeleton. Whereas F actin association is mediated by a central SYNPO motif, binding to alpha-actinin requires the C-terminal domain. Notably, the alpha-actinin binding domain is also essential for dendritic targeting and postsynaptic accumulation of SYNPO in primary neurons. Taken together, our data suggest that dendritic spine accumulation of SYNPO critically depends on its interaction with postsynaptic alpha-actinin and that SYNPO may regulate spine morphology, motility and function via its distinct modes of association with the actin cytoskeleton. PMID- 15659230 TI - Protective effects of insulin-like growth factor-I on the somatostatinergic system in the temporal cortex of beta-amyloid-treated rats. AB - Insulin-like growth factor-I (IGF-I) has protective effects against beta-amyloid (Abeta)-induced neuronal cell death. Because alterations of the somatostatinergic system have been described in Alzheimer's disease, we investigated the effects of the Abeta peptide and the possible protective role of IGF-I on the somatostatinergic system of the rat temporal cortex and on cell death and phosphorylated (p)-Akt levels in this area. Abeta25-35 was administered intracerebroventricularly to male rats via an osmotic minipump over 14 days (300 pmol/day). Another group received a subcutaneous IGF-I infusion (50 microg/kg/day), concomitant with Abeta25-35 administration, whereas a third group received IGF-I alone. Abeta25-35 significantly decreased the somatostatin (SRIF) like immunoreactive content and the SRIF receptor density, as a result of a decrease in the levels of the SRIF receptor subtype 2. The inhibitory effect of SRIF on adenylyl cyclase activity was significantly lower after Abeta25-35 infusion, whereas the levels of the inhibitory G protein subunit Gialpha1, Gialpha2 or Gialpha3 were unaltered. Cell death was increased and p-Akt levels decreased in Abeta25-35-treated animals. IGF-I administration increased immunoreactive IGF-I levels in the temporal cortex and restored all parameters affected by Abeta25-35 to baseline values. These findings suggest that IGF-I prevents the deleterious effect of Abeta25-35 on the somatostatinergic system. PMID- 15659228 TI - The basic helix-loop-helix transcription factor Nex-1/Math-2 promotes neuronal survival of PC12 cells by modulating the dynamic expression of anti-apoptotic and cell cycle regulators. AB - The basic helix-loop-helix transcription factor Nex1/Math-2 belongs to the NeuroD subfamily, which plays a critical role during neuronal differentiation and maintenance of the differentiated state. Previously, we demonstrated that Nex1 is a key regulatory component of the nerve growth factor (NGF) pathway. Further supporting this hypothesis, this study shows that Nex1 has survival-inducing properties similar to NGF, as Nex1-overexpressing PC12 cells survive in the absence of trophic factors. We dissected the molecular mechanism by which Nex1 confers neuroprotection upon serum removal and found that constitutive expression of Nex1 maintained the expression of specific G1 phase cyclin-dependent kinase inhibitors and concomitantly induced a dynamic expression profile of key anti apoptotic regulators. This study provides the first evidence of the underlying mechanism by which a member of the NeuroD-subfamily promotes an active anti apoptotic program essential to the survival of neurons. Our results suggest that the survival program may be viewed as an integral component of the intrinsic programming of the differentiated state. PMID- 15659231 TI - Effects of brain-derived neurotrophic factor (BDNF) on glial cells and serotonergic neurones during development. AB - Serotonergic neurones are among the first to develop in the central nervous system. Their survival and maturation is promoted by a variety of factors, including serotonin itself, brain-derived neurotrophic factor (BDNF) and S100beta, an astrocyte-specific Ca(2+) binding protein. Here, we used BDNF deficient mice and cell cultures of embryonic raphe neurones to determine whether or not BDNF effects on developing serotonergic raphe neurones are influenced by its action on glial cells. In BDNF-/- mice, the number of serotonin immunoreactive neuronal somata, the amount of the serotonin transporter, the serotonin content in the striatum and the hippocampus, and the content of 5 hydroxyindoleacetic acid in all brain regions analysed were increased. By contrast, reduced immunoreactivity was found for myelin basic protein (MBP) in all brain areas including the raphe and its target region, the hippocampus. Exogenously applied BDNF increased the number of MBP-immunopositive cells in the respective culture systems. The raphe area displayed selectively reduced immunoreactivity for S100beta. Accordingly, S100beta was increased in primary cultures of pure astrocytes by exogenous BDNF. In glia-free neuronal cultures prepared from the embryonic mouse raphe, addition of BDNF supported the survival of serotonergic neurones and increased the number of axon collaterals and primary dendrites. The latter effect was inhibited by the simultaneous addition of S100beta. These results suggest that the presence of BDNF is not a requirement for the survival and maturation of serotonergic neurones in vivo. BDNF is, however, required for the local expression of S100beta and production of MBP. Therefore BDNF might indirectly influence the development of the serotonergic system by stimulating the expression of S100beta in astrocytes and the production MBP in oligodendrocytes. PMID- 15659233 TI - Differential effects of melanin concentrating hormone on the central dopaminergic neurons induced by the cocaine- and amphetamine-regulated transcript peptide. AB - Stimulatory effects of cocaine- and amphetamine-regulated transcript (CART) peptide on central mesolimbic, nigrostriatal and mesocortical dopaminergic (DA) neurons were examined in female Sprague-Dawley rats. We also determined the different blocking effects of melanin concentrating hormone (MCH) on the stimulation by CART peptide in central DA systems. Intracerebroventricular administration of 1 microg CART peptide (55-102) produced increases in 3,4 dihydroxyphenylacetic acid (DOPAC) levels in the nucleus accumbens (NA) at 15 and 45 min, and in the striatum (ST) at 15 min, but not in the medial prefrontal cortex (MPFC). We found that the agonist of alpha-melanocyte stimulating hormone (alpha-MSH), MT II, at 10 microg had a stimulatory effect on the NA and ST DOPAC levels similar to the CART peptide. In contrast, 1 microg MCH and the antagonist of alpha-MSH, HS014, significantly decreased NA and ST DOPAC levels. However, only MCH prevented the stimulatory effect of CART peptide on DOPAC levels in the NA, but not in the ST. These results indicate that the stimulation of CART peptide on central DA neurons is region-specific, and that this effect can be blocked by MCH but not by the antagonist of alpha-MSH. PMID- 15659232 TI - Beta-site APP cleaving enzyme up-regulation induced by 4-hydroxynonenal is mediated by stress-activated protein kinases pathways. AB - 4-Hydroxynonenal (HNE), an aldehydic product of lipid peroxidation, up-regulates expression of the beta-site APP cleaving enzyme (BACE-1), an aspartyl protease responsible for the beta-secretase cleavage of amyloid precursor protein (AbetaPP), and results in increased levels of amyloid beta (Abeta) peptide. The mechanisms underlying this remain unclear but are of fundamental importance because prevention of BACE-1 up-regulation is viewed as an important therapeutic strategy. In this study, we exposed NT(2) neurons to a range of HNE concentrations (0.5-5 microm) that elicited an up-regulation of BACE-1 expression, a significant increase in intracellular and secreted levels of Abeta peptides as well as apoptosis involving poly-ADP ribose polymerase cleavage and activation of caspase 3. To delineate the molecular events involved in HNE mediated BACE-1 activation, we investigated the involvement of stress-activated protein kinases (SAPK), signal transducers and activators of transcription (STAT) and serine-threonine kinase B/phosphatidylinositol phosphate 3 kinase (Akt/PtdIns3K). Using specific pharmacological inhibitors, our results show that activation of c-Jun N-terminal kinases and p38(MAPK.), but not STAT or Akt/PtdIns3K, pathways mediate the HNE-dependent up-regulation of BACE-1 expression. Therefore, HNE, an oxidative stress mediator detected in vivo in the brains of Alzheimer's disease patients, may play a pathogenetic role in Alzheimer's disease by selectively activating SAPK pathways and BACE-1 that regulate the proteolytic processing of AbetaPP. PMID- 15659234 TI - BAALC 1-6-8 protein is targeted to postsynaptic lipid rafts by its N-terminal myristoylation and palmitoylation, and interacts with alpha, but not beta, subunit of Ca/calmodulin-dependent protein kinase II. AB - We cloned a rat BAALC 1-6-8 isoform cDNA (GenBank Accession No. AB073318) that encoded a 22-kDa protein, and identified endogenous BAALC 1-6-8 protein in the brain. The gene was expressed widely in the frontal part of the brain, and the protein was localized to the synaptic sites and was increased in parallel with synaptogenesis. The protein interacted with the alpha, but not beta, subunit of Ca(2+)/calmodulin-dependent protein kinase II (CaMKIIalpha). The interaction occurred between the N-terminal 35-amino-acid region of BAALC 1-6-8 protein and the C-terminal end of the regulatory domain of CaMKIIalpha, which contains alpha isoform-specific sequence. Thus, the interaction may be CaMKIIalpha-specific. We also found that BAALC 1-6-8 protein, as well as CaMKIIalpha, was localized to lipid rafts and that both myristoylation and palmitoylation of BAALC 1-6-8 N terminal portion were required for targeting of the protein into lipid rafts. These findings suggest that BAALC 1-6-8 protein play a synaptic role at the postsynaptic lipid raft possibly through interaction with CaMKIIalpha. PMID- 15659235 TI - The role of tissue plasminogen activator in methamphetamine-related reward and sensitization. AB - In the central nervous system, tissue plasminogen activator (tPA) plays a role in synaptic plasticity and remodeling. Our recent study has suggested that tPA participates in the rewarding effects of morphine by regulating dopamine release. In this study, we investigated the role of tPA in methamphetamine (METH)-related reward and sensitization. Repeated METH treatment dose-dependently induced tPA mRNA expression in the frontal cortex, nucleus accumbens, striatum and hippocampus, whereas single METH treatment did not affect tPA mRNA expression in these brain areas. The METH-induced increase in tPA mRNA expression in the nucleus accumbens was completely inhibited by pre-treatment with R(+)-SCH23390 and raclopride, dopamine D1 and D2 receptor antagonists, respectively. In addition, repeated METH treatment increased tPA activity in the nucleus accumbens. There was no difference in METH-induced hyperlocomotion between wild type and tPA-deficient (tPA-/-) mice. On the other hand, METH-induced conditioned place preference and behavioral sensitization after repeated METH treatment were significantly reduced in tPA-/- mice compared with wild-type mice. The defect of behavioral sensitization in tPA-/- mice was reversed by microinjections of exogenous tPA into the nucleus accumbens. Our findings suggest that tPA is involved in the rewarding effects as well as the sensitization of the locomotor stimulating effect of METH. PMID- 15659236 TI - Ampakine CX546 bolsters energetic response of astrocytes: a novel target for cognitive-enhancing drugs acting as alpha-amino-3-hydroxy-5-methyl-4 isoxazolepropionic acid (AMPA) receptor modulators. AB - Glutamate was previously shown to enhance aerobic glycolysis i.e. increase glucose utilization and lactate production with no change in oxygen levels, in mouse cortical astrocytes by a mechanism involving glutamate uptake. It is reported here that a similar response is produced in both hippocampal and cerebellar astrocytes. Application of the cognitive-enhancing drug CX546 promoted further enhancement of glucose utilization by astrocytes from each brain area following glutamate exposure. alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors represent the purported molecular target of cognitive enhancing drugs such as CX546, and the presence of AMPA receptor subunits GluR1-4 was evidenced in astrocytes from all three regions by immunocytochemistry. AMPA itself did not stimulate aerobic glycolysis, but in the presence of CX546, a strong enhancement of glucose utilization and lactate production was obtained in cortical, hippocampal and cerebellar astrocytes. The effect of CX546 was concentration-dependent, with an EC(50) of 93.2 microm in cortical astrocytes. AMPA-induced glucose utilization in the presence of CX546 was prevented by the AMPA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) and the negative modulator GYKI 52466. In addition, the metabolic effect of CX546 in the presence of AMPA was mimicked by the AMPA receptor modulator cyclothiazide. Our data suggest that astrocyte energetics represents a novel target for cognitive enhancing drugs acting as AMPA receptor modulators. PMID- 15659237 TI - Brain-derived neurotrophic factor prevents changes in Bcl-2 family members and caspase-3 activation induced by excitotoxicity in the striatum. AB - Brain-derived neurotrophic factor (BDNF) prevents the loss of striatal neurons caused by excitotoxicity. We examined whether these neuroprotective effects are mediated by changes in the regulation of Bcl-2 family members. We first analyzed the involvement of the phosphatidylinositol 3-kinase/Akt pathway in this regulation, showing a reduction in phosphorylated Akt (p-Akt) levels after both quinolinate (QUIN, an NMDA receptor agonist) and kainate (KA, a non-NMDA receptor agonist) intrastriatal injection. Our results also show that Bcl-2, Bcl-x(L) and Bax protein levels and heterodimerization are selectively regulated by NMDA and non-NMDA receptor stimulation. Striatal cell death induced by QUIN is mediated by an increase in Bax and a decrease in Bcl-2 protein levels, leading to reduced levels of Bax:Bcl-2 heterodimers. In contrast, changes in Bax protein levels are not required for KA-induced apoptotic cell death, but decreased levels of both Bax:Bcl-2 and Bax:Bcl-x(L) heterodimer levels are necessary. Furthermore, QUIN and KA injection activated caspase-3. Intrastriatal grafting of a BDNF-secreting cell line counter-regulated p-AKT, Bcl-2, Bcl-x(L) and Bax protein levels, prevented changes in the heterodimerization between Bax and pro-survival proteins, and blocked caspase-3 activation induced by excitotoxicity. These results provide a possible mechanism to explain the anti-apoptotic effect of BDNF against to excitotoxicity in the striatum through the regulation of Bcl-2 family members, which is probably mediated by Akt activation. PMID- 15659238 TI - Tournefolic acid B methyl ester attenuates glutamate-induced toxicity by blockade of ROS accumulation and abrogating the activation of caspases and JNK in rat cortical neurons. AB - The effects of nine polyphenolic compounds on glutamate-mediated toxicity were investigated. The underlying mechanisms by which a polyphenolic compound confers its effect were also elucidated. Treatment of cortical neurons with 50 microm glutamate for 24 h decreased cell viability by 45.8 +/- 7.9%, and 50 microm of tournefolic acid B methyl ester attenuated glutamate-induced cell death by 46.8 +/- 17.8%. Glutamate increased the activity of caspase 35.2-fold, and to a similar extent for caspase 2, 6, 8 and 9. Tournefolic acid B methyl ester abrogated glutamate-induced activation of caspase 2, 3, 6 and 9 by about 70%, and to a lesser extent for caspase 8. Treatment with glutamate for 1 h elevated reactive oxygen species (ROS) by 208.3 +/- 21.3%. Tournefolic acid B methyl ester eliminated the glutamate-induced accumulation of ROS. Glutamate increased the phosphorylation of p54-c-jun N-terminal kinase (JNK) concomitantly with activation of the endogenous antioxidant defense system. Tournefolic acid B methyl ester at 50 microm diminished the activity of p54-JNK in control and glutamate-treated cells, coinciding with the abolishment of the glutamate triggered antioxidant defense system. Therefore, tournefolic acid B methyl ester blocked the activation of the caspase cascade, eliminated ROS accumulation and abrogated the activation of JNK, thereby conferring a neuroprotective effect on glutamate-mediated neurotoxicity. PMID- 15659239 TI - Anesthesia and post-mortem interval profoundly influence the regulatory serine phosphorylation of glycogen synthase kinase-3 in mouse brain. AB - Glycogen synthase kinase-3 (GSK3) is a crucial enzyme contributing to the regulation of neuronal structure, plasticity and survival, is implicated as a contributory factor in prevalent diseases such as Alzheimer's disease and mood disorders and is regulated by a wide range of signaling systems and pharmacological agents. Therefore, factors regulating GSK3 in vivo are currently of much interest. GSK3 is inhibited by phosphorylation of serine-9 or serine-21 in GSK3beta and GSK3alpha, respectively. This study found that accurate measurements of phospho-Ser-GSK3 in brain are confounded by a rapid post-mortem dephosphorylation, with approximately 90% dephosphorylation of both GSK3 isoforms occurring within 2 min post-mortem. Furthermore, three anesthetics, pentobarbital, halothane and chloral hydrate, each caused large in vivo increases in the serine phosphorylation of both GSK3beta and GSK3alpha in several regions of mouse brain. Thus, studies of the phosphorylation state of GSK3 in brain, and perhaps in other tissues, need to take into account post-mortem changes and the effects of anesthetics and there is a direct correlation between anesthesia and high levels of serine-phosphorylated GSK3. PMID- 15659241 TI - Human resources: the Cinderella of health sector reform in Latin America. AB - Human resources are the most important assets of any health system, and health workforce problems have for decades limited the efficiency and quality of Latin America health systems. World Bank-led reforms aimed at increasing equity, efficiency, quality of care and user satisfaction did not attempt to resolve the human resources problems that had been identified in multiple health sector assessments. However, the two most important reform policies - decentralization and privatization - have had a negative impact on the conditions of employment and prompted opposition from organized professionals and unions. In several countries of the region, the workforce became the most important obstacle to successful reform.This article is based on fieldwork and a review of the literature. It discusses the reasons that led health workers to oppose reform; the institutional and legal constraints to implementing reform as originally designed; the mismatch between the types of personnel needed for reform and the availability of professionals; the deficiencies of the reform implementation process; and the regulatory weaknesses of the region.The discussion presents workforce strategies that the reforms could have included to achieve the intended goals, and the need to take into account the values and political realities of the countries. The authors suggest that autochthonous solutions are more likely to succeed than solutions imported from the outside. PMID- 15659240 TI - cAMP response element binding protein (CREB) activates transcription via two distinct genetic elements of the human glucose-6-phosphatase gene. AB - BACKGROUND: The enzyme glucose-6-phosphatase catalyzes the dephosphorylation of glucose-6-phosphatase to glucose, the final step in the gluconeogenic and glycogenolytic pathways. Expression of the glucose-6-phosphatase gene is induced by glucocorticoids and elevated levels of intracellular cAMP. The effect of cAMP in regulating glucose-6-phosphatase gene transcription was corroborated by the identification of two genetic motifs CRE1 and CRE2 in the human and murine glucose-6-phosphatase gene promoter that resemble cAMP response elements (CRE). RESULTS: The cAMP response element is a point of convergence for many extracellular and intracellular signals, including cAMP, calcium, and neurotrophins. The major CRE binding protein CREB, a member of the basic region leucine zipper (bZIP) family of transcription factors, requires phosphorylation to become a biologically active transcriptional activator. Since unphosphorylated CREB is transcriptionally silent simple overexpression studies cannot be performed to test the biological role of CRE-like sequences of the glucose-6 phosphatase gene. The use of a constitutively active CREB2/CREB fusion protein allowed us to uncouple the investigation of target genes of CREB from the variety of signaling pathways that lead to an activation of CREB. Here, we show that this constitutively active CREB2/CREB fusion protein strikingly enhanced reporter gene transcription mediated by either CRE1 or CRE2 derived from the glucose-6 phosphatase gene. Likewise, reporter gene transcription was enhanced following expression of the catalytic subunit of cAMP-dependent protein kinase (PKA) in the nucleus of transfected cells. In contrast, activating transcription factor 2 (ATF2), known to compete with CREB for binding to the canonical CRE sequence 5' TGACGTCA-3', did not transactivate reporter genes containing CRE1, CRE2, or both CREs derived from the glucose-6-phosphatase gene. CONCLUSIONS: Using a constitutively active CREB2/CREB fusion protein and a mutant of the PKA catalytic subunit that is targeted to the nucleus, we have shown that the glucose-6 phosphatase gene has two distinct genetic elements that function as bona fide CRE. This study further shows that the expression vectors encoding C2/CREB and catalytic subunit of PKA are valuable tools for the study of CREB-mediated gene transcription and the biological functions of CREB. PMID- 15659242 TI - Rivastigmine: an open-label, observational study of safety and effectiveness in treating patients with Alzheimer's disease for up to 5 years. AB - BACKGROUND: Rivastigmine, a butyl- and acetylcholinesterase inhibitor, is approved for symptomatic treatment of Alzheimer's disease (AD). Data supporting the safety and efficacy of second-generation cholinesterase inhibitors, such as rivastigmine, are available for treatment up to 1 year, with limited data up to 2 1/2 years. The purpose of this report is to present safety and effectiveness data for rivastigmine therapy in patients with mild to moderately severe AD receiving treatment for up to 5 years. METHODS: An observational approach was used to study 37 patients with originally mild to moderate AD receiving rivastigmine as a therapy for AD in an open-label extension (ENA713, B352 Study Group, 1998). RESULTS: The initial trial demonstrated rivastigmine was well-tolerated and effective in terms of cognition, global functioning and activities of daily living. In this open label extension, high-dose rivastigmine therapy was safe and well tolerated over a 5-year period. Two thirds of the participants still enrolled at week 234 were in the original high-dose rivastigmine group during the double-blind phase, suggesting that early therapy may confer some benefit in delaying long-term progression of symptoms. CONCLUSIONS: Long-term cholinesterase inhibition therapy with rivastigmine was well tolerated, with no dropouts due to adverse effects past the initial titration period. Early initiation of treatment, with titration to high-dose therapy, may have an advantage in delaying progression of the illness. PMID- 15659243 TI - Modelling the correlation between the activities of adjacent genes in Drosophila. AB - BACKGROUND: Correlation between the expression levels of genes which are located close to each other on the genome has been found in various organisms, including yeast, drosophila and humans. Since such a correlation could be explained by several biochemical, evolutionary, genetic and technological factors, there is a need for statistical models that correspond to specific biological models for the correlation structure. RESULTS: We modelled the pairwise correlation between the expressions of the genes in a Drosophila microarray experiment as a normal mixture under Fisher's z-transform, and fitted the model to the correlations of expressions of adjacent as well as non-adjacent genes. We also analyzed simulated data for comparison. The model provided a good fit to the data. Further, correlation between the activities of two genes could, in most cases, be attributed to either of two factors: the two genes both being active in the same age group (adult or embryo), or the two genes being in proximity of each other on the chromosome. The interaction between these two factors was weak. CONCLUSIONS: Correlation between the activities of adjacent genes is higher than between non adjacent genes. In the data we analyzed, this appeared, for the most part, to be a constant effect that applied to all pairs of adjacent genes. PMID- 15659244 TI - T cell responses against tumor associated antigens and prognosis in colorectal cancer patients. AB - INTRODUCTION: Spontaneous T cell responses against specific tumor-associated antigens (TAA) are frequently detected in peripheral blood of tumor patients of various histiotypes. However, little is known about whether these circulating, spontaneously occurring, TAA-reactive T cells influence the clinical course of disease. METHODS: Fifty-four HLA-A2 positive colorectal cancer patients had been analyzed for the presence of T cell responses against epitopes derived from the TAA Ep-CAM, her-2/neu, and CEA either by ELISPOT assay or by intracellular cytokine staining. Then, Kaplan-Meier survival analysis was performed comparing T cell-responders and T-cell-non-responders. For comparison, a group of T-cell-non responders was compiled stringently matched to T-cell-responders based on clinical criteria and also analyzed for survival. RESULTS: Sixteen out of 54 patients had a detectable T cell response against at least one of the three tested TAA. Two out of 21 patients (9.5%) with limited stage of disease (UICC I and II) and 14 out of 33 patients (42.4%) with advanced disease (UICC III and IV) were T cell response positive. Comparing all T-cell-responders (n = 16) and all T cell-non-responders (n = 38), no survival difference was found. In an attempt to reduce the influence of confounding clinical factors, we then compared 16 responders and 16 non-responders in a matched group survival analysis; and again no survival difference was found (p = 0.7). CONCLUSION: In summary, we found no evidence that spontaneous peripheral T cell responses against HLA-A2-binding epitopes of CEA, her-2/neu and Ep-CAM are a strong prognostic factor for survival. PMID- 15659245 TI - Free does not mean affordable: maternity patient expenditures in a public hospital in Bangladesh. AB - OBJECTIVE: This study investigated a) the amount and types of out-of-pocket expenditures by patients for nominally free services in a large public hospital in Bangladesh, b) the factors influencing these expenses, and c) the impact of these expenses on household income. METHODS: Eighty-one maternity patients were interviewed during their hospitalization in the Dhaka Medical College Hospital. Patients were selected by quota sample to match the distribution of maternity patient categories in the hospital. Patients were interviewed with a semi structured, in-depth questionnaire. RESULTS: All interviewees incurred substantial out-of-pocket expenditures for travel, hospital admission fees, medicine, tests, food, and tips. Only two of the expenditures, travel expenses and admission fees, were not supposed to be provided free of charge by the hospital. The median total per-patient expenditure was $65 (range $2-$350), equivalent to 7% (range 0.04%-225%) of annual household income. Half of all patients reported that their families had to borrow to pay for care at interest rates of 5%-30% per month. A third of these families reported selling jewelry, land or household items to moneylenders. The rural patients reported more difficulty in paying for care than the urban patients. Factors increasing the expenditures were duration of hospitalization, rural residence, and necessary (e.g. C-section, hysterectomy) and unnecessary (e.g. episiotomy) medical procedures. CONCLUSION: Free maternity services in Bangladesh impose large out-of pocket expenditures on patients. Authorities could reduce the burden by reducing the duration of hospital stays, limiting use of medical procedures, eliminating tips, and moving routine services closer to potential users. Fee for service could reduce unofficial expenditures if the fee were lower than and replaced typical unofficial expenditures, otherwise adding service fees without reform of current hospital practices would lead to even more burdensome expenditures and inequities. PMID- 15659246 TI - Using large-scale perturbations in gene network reconstruction. AB - BACKGROUND: Recent analysis of the yeast gene network shows that most genes have few inputs, indicating that enumerative gene reconstruction methods are both useful and computationally feasible. A simple enumerative reconstruction method based on a discrete dynamical system model is used to study how microarray experiments involving modulated global perturbations can be designed to obtain reasonably accurate reconstructions. The method is tested on artificial gene networks with biologically realistic in/out degree characteristics. RESULTS: It was found that a relatively small number of perturbations significantly improve inference accuracy, particularly for low-order inputs of one or two genes. The perturbations themselves should alter the expression level of approximately 50 60% of the genes in the network. CONCLUSIONS: Time-series obtained from perturbations are a common form of expression data. This study illustrates how gene networks can be significantly reconstructed from such time-series while requiring only a relatively small number of calibrated perturbations, even for large networks, thus reducing experimental costs. PMID- 15659247 TI - No influence of the P-glycoprotein polymorphisms MDR1 G2677T/A and C3435T on the virological and immunological response in treatment naive HIV-positive patients. AB - BACKGROUND: In a retrospective study of HIV-infected patients, we investigated the influence of the MDR1 genotype (G2677T/A and C3435T) on the virological and immunological response of treatment naive patients. METHODS: The MDR1 genotype was analysed from 72 patients in whom antiretroviral therapy was initiated between 1998 and 2004. Data were obtained at week 4, 12, 24 and 48 and were analysed by the Kruskal-Wallis test. RESULTS: During the first 48 weeks of antiretroviral therapy, there were no significant differences in the virological and immunological response with respect to the MDR1 2677 and 3435 genotypes and the 2677/3435 haplotype. CONCLUSIONS: In view of different results from several studies concerning the influence of MDR1 polymorphisms on the immunological and virological response to antiretroviral therapy, further studies with larger patient groups and longer follow-up are necessary in order to resolve conflicting issues. PMID- 15659248 TI - [Nursing as an art]. AB - It is more important now than ever to define the goals of nursing by using an aesthetic approach. In every day practice the nurse must explore and adopt these functions, which constitute the essence of nursing as an art. The aim of this paper is to firstly explore the notion of nursing as an art, underpinned by the philosophy of art and to secondly discuss the hidden connections and the criteria for nursing seen or thought to be an art. In this paper, the aesthetic values have been used to depict and to expose the invisible qualities of nursing. Through the written text the attempt to explore the nature of nursing by an aesthetic approach, reveals that the essence of nursing is constituted by the same origins, which define the essence of art. This exploration of the literature led to the recognition of the quality of imitation as an attempt of the artist to awaken individuals to understand what a piece of art represents. Respectively, a nurse by represents a picture of wholeness for the health, attempt to stimulate patient to move forwards regaining control and achieving the state of well-being. The search of the expression as one of the sources of the aesthetic values has brought to the surface that nursing is expressed as a whole of unique functions containing love, advocacy, calmness, care and empathy. The exploration of the combination of the elements, their articulation or their structure exposes the beauty itself. The beauty of a statue as Venus of Milo as well as, in terms of nursing, the beauty of caring as the heart of nursing. However it is difficult to capture the totality of nursing in this paper, which attempts more to portray this beauty of nursing rather than to seek for a clear answer of what nursing is. PMID- 15659249 TI - [Research in nursing domain: essential base for the development of nursing knowledge and professionalism]. AB - In this article the principles and reasons by which in the Italian Nursing field research is necessary, in a popular way, are discussed. The use of scientific inquiry to study its peculiar aspects through the use of the scientific method for the production of specific knowledge is faced. Phases of the quantitative method are illustrated It is also underlined the rationale and reasons why it is considered urgent to form aware and active professionals, able to use the results of the literature along with to be able to conduct or to participate in research studies with the purpose to nurturing the scientific debate and to care with consequential data from the scientific evidence. PMID- 15659250 TI - [The sign and the significance. Notes of sharing of meaning between nurse and patient]. AB - Attention to patient experience and words represents one of factors that contribute to outcome of care. Beginning from a preliminary analysis of characteristics of scientific and common language, this paper means to contribute to reflection about necessity, for nurse and patient, to share illness meaning in order to effectiveness of relationship. PMID- 15659251 TI - [When the patient asks for counselling, when the patient doesn't ask for counselling, when the patient refuses therapy]. AB - A large debate that has brought some depths changes is in action. It is on the relationship between patients and health professionals, physicians, nurses, psychologists etc. The path went through the medical paternalism to the phase of the autonomy, thin to reach the ethics of taking care. In these years, bioethics has tried to answer to urgent questions that primarily interest the private sphere of the people. Currently, by contributions of European and American philosophies, an ethics of the responsibility is delineating This, differentiates the ethics of the rights and the rules (autonomy, self-determination). The attitude of taking care postpones to a whole intellectual, affective, moral competences, that every health professional has to possess to establish relationship with people. This allows, beyond the autonomy, a fundamental moral principle that remains, to give a positive emotional answer and support the conditions and the situation of others. In this answer, the availability to be in the identification process is affirmed to, with their problems and, the desire to do the possible to assuage their suffering. If the person is set in a large net of relationships, is able to express autonomy mediating counselling process of health professionals. The client will freely choose to assume self-care responsibilities. PMID- 15659252 TI - [A survey on working satisfaction of nurses]. AB - This article details the results of a survey research undertaken by the Azienda Ospedaliera dell'Umbria to determine the knowledge of currently employed nurses on legislation change that has introduced the assumption of responsibility. Said legislation recognises the role of nursing as a true profession, where scientific methodology, on which its growth is based, should help to overcome out-dated perceptions of the role of the nursing assistant. 346 questionnaires were distributed to a representative sample (32.77%) of the total of 1056 nurses. Of these 346 questionnaires, 238 were completed, representing 68.79% of the total. PMID- 15659253 TI - [Nurses in specialistic surgery: centrality of a role]. AB - According international trend patients over 70 submitted surgery in our Thoracic Surgery Unit are increasing. Therefore these patients are at higher risk of complications in the post-operative period owing to advanced age and comorbidities; most important among them BPCO and cardiovascular diseases. In our study most frequent and best evaluable complication was pneumonia. However postoperative pneumonia percentage decreased since 2000, when a nurse dedicated for thoracic patients only was instituted. Since then in our unit assistance model consisted of two thoracic surgeons supported by a nurse in charge to General Surgery Paramedic Unit. Innovation did not request adjunctive costs and contributed to a significant increase in professional level of nursing standard. PMID- 15659254 TI - [Nurses presence in the sterilisation service according UNI EN directives]. AB - The aim of the sterilisation process is to inactivates the microbiological contamination, changing the status of contaminated product in sterile one. The exposition time at the correctly sterilisation is not the only factor which allow the process succeed: we need to pay attention at the microbiological charge that is present on the primary material, (bioburden), the place where the process occur, the assemblage, the pack and stockage. In the health services for a good successful of this passage, we need to identify an professional figure of reference as supervisors of the sterilisation process and as an expert adviser (consultant). This article represent an reflection of the nursing role. PMID- 15659255 TI - [Low back pain risk in nurses and its prevention]. AB - We present the results of a study on the prevalence of low back pain (LBP) in a group of hospital nursing personnel. In 180 subjects (89% women) a questionnaire on LBP was collected. The prevalence of LBP episodes during the last 12 months was 63,9%; no difference was observed according to the working area (medicine, surgery, emergency). The proportion of subjects with LBP tended to increase according to the previous duration of work, but not with age. The prevalence of LBP was significantly lower in nursing personnel working according the team nursing model, that is less repetitive, and involves less work load, compared to the functional nursing model. Furthermore, LBP prevalence was lower among nurses that received an adequate formation on the prevention of occupational LBP. The results of the study confirm the relevance of LBP in nursing personnel, but suggest also that an adequate preventive programme can significantly reduce the risk. PMID- 15659256 TI - Macrolide antibiotics as anti-inflammatory agents. PMID- 15659257 TI - Update on beta-lactam allergy diagnosis. AB - Allergic reactions to penicillin and cephalosporin antibiotics are common. The mechanisms involved are heterogeneous and not yet completely understood. However, clinical and immunologic studies suggest that type I (IgE-mediated) and type IV (cell-mediated) pathogenic mechanisms are involved in most immediate and non immediate reactions, respectively. The diagnostic procedure has recently been standardized under the aegis of the European Network for Drug Allergy and the European Academy of Allergy and Clinical Immunology interest group on drug hypersensitivity. In this review, we describe the general guidelines for evaluating subjects in whom there is suspicion of allergic reactions to beta lactams. PMID- 15659258 TI - Hypersensitivity reactions to fluoroquinolones. AB - Fluoroquinolone antibiotics cause immediate and delayed hypersensitivity reactions, and may also affect internal organs and circulating blood cells. The underlying pathomechanisms are only partly understood. The extent of cross reactivity among different quinolones depends on the type of clinical manifestation and its underlying mechanism. Despite recent advances, reliable diagnostic tests are still lacking. Recent studies have shown quinolone-specific IgE in vitro in more than 50% of patients with immediate-type reactions and a considerable cross-reactivity with related compounds. In maculopapular drug exanthems from ciprofloxacin, specific T-cell clones were identified, and cross reactivity to related compounds was detected in approximately 50% of the clones. From re-exposure studies in patients with exanthems, cross-reactivity appears to be lower. Cellular tests such as lymphocyte transformation tests are currently not very useful. For prick and intradermal skin tests, widely divergent nonirritant test concentrations have been recommended. Desensitization may be possible in selected patients. PMID- 15659259 TI - Immunoglobulin E-mediated reactions to corticosteroids. AB - Allergic reactions to steroids are rare, but are becoming more commonly recognized by clinicians. Although many reactions are thought to be IgE mediated, most are adverse drug reactions caused by nonallergic mechanisms, some related to additives in the steroid preparation being administered. Provocative challenge with the suspected offending agent, serologic tests for allergen-specific IgE, and skin testing are the most reliable methods for assessing hypersensitivity to steroids, although none is infallible. Therapeutic options for patients with documented steroid allergy include the use of alternative steroid preparations, desensitization, and, possibly, the use of monoclonal anti-IgE antibodies. PMID- 15659260 TI - Anaphylactoid reactions to radiocontrast media. AB - Adverse reactions to contrast material are a concern because iodinated contrast materials are commonly used drugs. The risk for adverse reaction is 4% to 12% with ionic contrast materials and 1% to 3% with nonionic contrast materials. The risk for severe adverse reaction is 0.16% with ionic contrast materials and 0.03% with nonionic contrast materials. The death rate, one to three per 100,000 contrast administrations, is similar for both ionic and nonionic agents. More than 90% of adverse reactions with nonionic contrast materials are anaphylactoid. The types of severe reactions seen with nonionic contrast administration were initially predominantly anaphylactoid. With the advent of helical CT angiography, the reactions are now predominantly attributable to cardiopulmonary decompensation. With the widespread use of nonionic contrast materials, adverse reactions are now seen less frequently. Skills involved in evaluating and treating adverse reactions are not as frequently used. Periodic reviews and updates of specific treatment plans for various reactions with the physicians and staff who use contrast material are very important to ensure optimal preparedness. The key to successful treatment is preparation and early intervention. PMID- 15659261 TI - CD25+ T cells and regulation of allergen-induced responses. AB - CD4 T helper 2 (Th2) cells, with the characteristic interleukin (IL)-4, IL-5, and IL-13 cytokine secretion profile, play an important role in the initiation and perpetuation of allergic airways disease. It is clear from recent studies that CD4+ T cells with distinct cytokine-producing abilities have regulatory functions that limit allergic inflammation. Studies of allergic airway inflammation in mice have identified different types of T regulatory cells (Tregs) that control the disease phenotype. The cytokines associated with the Treg phenotype in mice include both soluble and cell membrane-bound transforming growth factor (TGF) beta and IL-10. Both contact-dependent mechanisms involving membrane-bound TGF beta and contact-independent mechanisms involving soluble TGF-beta and IL-10 have been invoked to describe the function of these Tregs. In humans, studies of milk allergy show an association between circulating CD4+CD25+ Tregs and tolerance to the causative allergen, beta-lactoglobulin. The identification of Tregs as suppressors of allergic disease may promote the development of new therapeutic strategies. PMID- 15659263 TI - Regulatory cells and the control of respiratory infection. AB - The role of distinct CD4+ T-cell populations in regulating the nature and strength of immune responses is well documented, and has in the past principally focused on the mutual antagonism between Th1 and Th2 cells, which secrete interferon (IFN)-gamma and interleukin (IL)-4, respectively. However, the recent identification of T cells that secrete high levels of IL-10 and/or transforming growth factor-b, but not IFN-g or IL-4, called regulatory T (Tr) cells has prompted a paradigm shift in our understanding of the regulation of immune responses following infection. In this review, we focus on the role of antigen specific Tr cells in the lungs following infection with various respiratory pathogens and discuss the targeting of Tr in the development of new therapies for immune-mediated diseases, such as allergy. PMID- 15659262 TI - Helminth infections: protection from atopic disorders. AB - Westernized countries are suffering from an epidemic rise in immunologic disorders, such as childhood allergy. A popular explanation is that the increased prevalence in allergy is due to a diminished or altered exposure to gut-dwelling microbes, resulting in a disordered immunoregulation. Various population studies have provided a strong case for the involvement of helminth infections in this respect. Detailed analysis of helminth-induced immune responses showed that helminths not only prime for polarized Th2 responses but also potently induce T cell hyporesponsiveness. Recently, it has been demonstrated that helminths induce suppressed host immune responses by the priming for regulatory T cells. It is proposed that this regulatory T cell-inducing activity accounts for the protection observed in the development of allergic disorders. It would be interesting to define and characterize particular helminth molecules that have profound immunomodulatory capacities as a target for therapeutic application in the treatment or prophylaxis of allergic manifestations. PMID- 15659264 TI - Role of regulatory dendritic cells in allergy and asthma. AB - Dendritic cells (DCs) are the most efficient inducers of all immune responses, and are capable of either inducing productive immunity or maintaining the state of tolerance to self antigens and allergens. In this review, we summarize the emerging literature on DCs, with emphasis on the regulatory function of DCs in allergy and asthma. In particular, we summarize recent data regarding the relationship between DC subsets and TH1, TH2, and regulatory T (TReg) cells. The diverse functions of DCs have been attributed to distinct lineages of DCs, which arise from common immature precursor cells that differentiate in response to specific maturation-inducing or local microenvironment conditions. These subsets of DCs induce different lineages of T cells, such as TH1, TH2, and TReg cells, including Th1Reg and Th2Reg cells, which regulate allergic diseases and asthma. Subsets of DCs regulate the induction of a variety of T-cell subtypes, which suppress the development of allergy and asthma, thus providing anti-inflammatory responses and protective immunity. PMID- 15659267 TI - A genetic engineering strategy to eliminate peanut allergy. AB - Peanut allergy is an IgE-mediated hypersensitivity reaction with an increasing prevalence worldwide. Despite its seriousness, to date, there is no cure. Genetic engineering strategies can provide a solution. The post-transcriptional gene silencing (PTGS) model can be used effectively to knock out the production of allergenic proteins in peanut by specific degradation of the endogenous target messenger RNA (mRNA). Ara h 2, the most potent peanut allergenic protein, was selected as a model to demonstrate the feasibility of this concept. Transgenic peanut plants were produced via microprojectile-mediated transformation of peanut embryos using a plasmid construct, which contains a fragment of the coding region of Ara h 2 linked to an enhanced CaMV 35S constitutive promoter. Molecular analyses, including polymerase chain reaction and Southern blots, confirmed the presence of the stable integration of the Ara h 2 transgene into the peanut genome. Northern hybridization showed the expression of the Ara h 2 transgene in all vegetative tissues of the mature transgenic peanut plants, indicating the stable expression of the truncated Ara h 2 transgene throughout the development of the plants. It is, therefore, reasonable to expect that the truncated Ara h 2 transgene transcripts will be synthesized in the seeds and will trigger the specific degradation of endogenous Ara h 2 mRNA. The next step will be to grow the transgenic peanut plants to full maturity for seed production and to determine the level of allergen Ara h 2. PMID- 15659271 TI - Supplement roulette. PMID- 15659268 TI - Fish and shellfish allergy. AB - Fish and shellfish are important in the American diet and economy. Nearly $27 billion are spent each year in the United States on seafood products. Fish and shellfish are also important causes of food hypersensitivity. In fact, shellfish constitute the number one cause of food allergy in the American adult. During the past decade, much has been learned about allergens in fish and shellfish. The major allergens responsible for cross-reactivity among distinct species of fish and amphibians are parvalbumins. The major shellfish allergen has been identified as tropomyosin. Many new and important potential cross-reacting allergens have been identified within the fish family and between shellfish, arachnids, and insects. Extensive research is currently underway for the development of safer and more effective methods for the diagnosis and management of fish and shellfish hypersensitivity. PMID- 15659269 TI - Adult food allergy. AB - Adult food allergy is estimated at approximately 3.2% worldwide. The persistence of childhood food allergy is unusual, peanut allergies excepted. Once established in adults, food allergy is rarely cured. Factors favoring the acquisition of allergy could be sensitization to pollens, occupational sensitization by inhalation, drugs (such as tacrolimus), and sudden dietary changes. Severe anaphylaxis and oral allergy syndrome are frequent. The fatality risk is estimated at 1% in severe anaphylaxis. Risk factors for severe anaphylaxis are agents causing increased intestinal permeability, such as alcohol and aspirin. b blockers, angiotensin-converting enzyme (ACE) inhibitors, and exercise are other factors. Gastrointestinal food allergy remains, to a large extent, undiagnosed in adults. Food allergens are mainly fruit and vegetable, related to pollen sensitizations, or to latex allergy. Wheat flour allergy is increasing. The diagnosis relies on prick skin tests, detection of specific IgEs, and standardized oral challenges. Strict avoidance diets are necessary. Specific immunotherapy to pollens may be efficient for cross-reactive food allergies. PMID- 15659270 TI - The use of patch testing in the diagnosis of food allergy. AB - Diagnosing food allergies can be challenging to the practitioner. Our armamentarium includes standardized skin prick testing, radioallergoimmunosorbent (RAST) testing, and food challenges. These methods have certainly been helpful in the IgE-mediated disorders, including urticaria and anaphylaxis. However, diagnosing patients who have the non-IgE (cell-mediated) or mixed (IgE and cell mediated) disorders remains challenging with our current diagnostic methods. Recent studies have examined the use of patch testing for these food-allergic patients, specifically those with atopic dermatitis and eosinophilic esophagitis. In this article, we review literature regarding patch testing: its methods, its statistical usefulness, and its potential future role. PMID- 15659272 TI - The disposition of the concussed athlete: a conundrum and an all too common injury. PMID- 15659274 TI - Exercise and its effects on the central nervous system. AB - Exercise can have profound effects on numerous biologic systems within the human body, including the central nervous system (CNS). The inherent complexity of the CNS, and the methodologic difficulties in evaluating its in vivo neurochemistry in humans, provide challenges to investigators studying the impact of exercise on the CNS. As a result, our knowledge in this area of exercise science remains relatively limited. However, advances in research technology are allowing investigators to gain valuable insight into the neurobiologic mechanisms that contribute to the bidirectional communication that occurs between the periphery and the CNS during exercise. This article examines how exercise-induced alterations in the CNS contribute to central fatigue and the overtraining syndrome, and how exercise can influence psychologic wellbeing and cognitive function. PMID- 15659273 TI - Dental injuries in sports. AB - The expanding arena of sports and vigorous recreational activities is associated with an increased risk for sports-related injuries, including trauma to the teeth. Because dental professionals may not be present on site, in the best interest of athletes, it is suggested that sports medicine professionals who are present be provided with current evaluative, emergent, and referral protocols for proper management of sports-related dental injuries to address the immediate needs of the athlete and to enhance the long-term prognosis of the traumatized tooth. This article provides information related to some of the more common dental injuries encountered in sports, including crown fractures, root fractures, and traumatic tooth displacements. Use of properly fitted, custom-fabricated athletic mouth guards for the prevention of dental injuries is recommended strongly, as is the inclusion of a dentist on the sports medicine team roster. PMID- 15659275 TI - Acute disposition of neck injuries. AB - Neck injuries can be some of the most serious and anxiety-producing injuries that occur during sporting events. It is important for the team physician to be prepared for the care of these injuries and be able to identify some of the more serious injuries. Proper care of these injuries can be life saving and prevent further injury and permanent disability. This article reviews the principles of management and latest evidence for acute neck injuries. PMID- 15659276 TI - Current thinking: return to play and transient quadriplegia. AB - Athletes that participate in contact and collision sports assume risk of serious injury each time they take the field. For those athletes that have sustained an episode of transient quadriplegia, the decision of whether to return to competition can be a difficult one. Some athletes, realizing how close they may have come to permanent injury, may decide that further participation is not in their best interest. Others may be somewhat undecided, and some may want to return at all costs. As the treating physician, the goal is to identify those athletes who after a single episode of transient quadriplegia are at increased risk for further injury and consequently should discontinue participation in contact sports. Factors that may contribute to that determination include mechanism of injury, prior history of neurologic symptoms or injury, and anatomic features that may predispose to further injury such as disc herniation, fracture, or cervical stenosis. PMID- 15659277 TI - Bony healing in a patient with bilateral L5 spondylolysis. PMID- 15659278 TI - Beyond hypoestrogenism in amenorrheic athletes: energy deficiency as a contributing factor for bone loss. AB - The etiology of amenorrhea in exercising women is linked to a mismatch between caloric intake and high levels of exercise energy expenditure that results in a chronic energy deficit. This in turn stimulates compensatory mechanisms such as weight loss, metabolic hormone alterations, or energy conservation that subsequently causes a central suppression of reproductive function and concomitant hypoestrogenism. This suppression of reproductive function is associated with stress fractures, loss of bone mineral density, the failure to achieve peak bone mass, osteopenia, and osteoporosis. It has generally been accepted that the chronic hypoestrogenism is the major cause of bone loss in exercising women. However, the effects of food restriction and energy deficiency on bone mineral density likely represents an estrogen-independent mechanism for bone loss that involves some of the metabolic-related hormones altered with exercise-associated amenorrhea. These hormones (IGF-1 and leptin) play an important role in modulating bone turnover and bone mineral density in these women. PMID- 15659279 TI - Catastrophic spine injuries in sports. AB - Catastrophic spine injuries in sports are rare but tragic events. The sports with the highest risk of catastrophic spinal injuries are football, ice hockey, wrestling, diving, skiing and snowboarding, rugby, cheerleading, and baseball. A common mechanism of injury for all at-risk sports is an axial compression force to the top of the head with the neck slightly flexed. We review common mechanisms of injury and prevention strategies for spine injuries in the at-risk sports. PMID- 15659280 TI - Field-side and prehospital management of the spine-injured athlete. AB - Rapid on-field diagnosis and early stabilization can help to optimize the outcomes of spinal injury, which can have devastating consequences. Several basic principles will guide the rescuer through this process. Preinjury planning should include appointing a team leader, assessing the athletes' equipment, acquiring appropriate equipment to facilitate stabilization, and establishing lines of communication to emergency medical services (EMS). When an athlete is down, the team leader should proceed to quickly assess airway, breathing, circulation, level of consciousness, and activation of EMS. This should be followed by stabilization of the head and neck, a coordinated log roll, and ultimately complete spinal immobilization for transport. Specific techniques for stabilizing the cervical spine, removing the facemask, log rolling the athlete, and lifting the athlete, will improve outcome. The helmet and shoulder pads should remain in place during transport unless specific indications require their removal, in which case a specific protocol should be strictly followed. PMID- 15659281 TI - Validation study to evaluate the reproducibility of a candidate in vitro potency assay of newcastle disease vaccines and to establish the suitability of a candidate biological reference preparation. AB - A quantification assay for the Haemagglutinin-Neuraminidase (HN) protein of Newcastle Disease Virus (NDV) has been developed at CIDC-Lelystad as a candidate in vitro potency test for inactivated Newcastle disease (ND) vaccines. In studies performed at CIDC-Lelystad, a high correlation was demonstrated between the results of this candidate in vitro potency assay and the results of the serological potency assay (European Pharmacopoeia monograph 0870; test A). Furthermore, a high correlation between the serological data (Haemagglutination Inhibition-antibody titres) and clinical protection after challenge was demonstrated. Correlation between in vivo and in vitro potency assays was confirmed in a collaborative pre-validation study. In the pre-validation study three Official Medicines Control Laboratories (OMCLs) determined both the NDV-HN antigen content and the in vivo potency (vaccination-serology and vaccination challenge) of 6 vaccine batches. The conclusion of the pre-validation study was that a large-scale collaborative study should be organised to validate the in vitro method and the suitability of the reference preparation. This report describes the outcome of this study. In brief, 14 laboratories (8 OMCLs and 6 vaccine manufacturers) determined the NDV-HN antigen content of 9 different vaccines in 3 independent tests. The vaccine batches were produced by 5 different manufacturers and represent a quantitative range of ND antigen content. One vaccine batch with insufficient potency and one poultry vaccine not containing NDV were included. Statistical evaluation of the results indicated that the antigen content could be determined with high precision. A good repeatability as well as reproducibility was found. Furthermore all laboratories found a similar ranking of the vaccines, based on the antigen content. Comparison of the antigen content and the in vivo potency of a series of vaccines with relatively low potencies indicated that a threshold relative antigen level of 7.0 antigen units per dose would discriminate between vaccine batches with sufficient and insufficient potency. An in vitro assay with this threshold level for antigen content did not result in any false positive results and only a limited number of false negative results in the BSP055 study. We conclude that the in vitro measurement of the antigen content of inactivated ND-vaccines with the proposed method is a reliable alternative potency assay that could be included as a new method in monograph 0870 on ND-vaccines. PMID- 15659282 TI - Establishment of batch 4 of the Biological Reference Preparation (BRP) for rabies vaccine (inactivated) for veterinary use. AB - 9 laboratories from 7 countries including both laboratories from the public and private sector participated in a collaborative study organised under the aegis of the European Directorate for the Quality of Medicines Biological Standardisation Programme in order to establish batch 4 of the European Pharmacopoeia (Ph. Eur.) Biological Reference Preparation (BRP) for rabies vaccine (inactivated) for veterinary use. Establishment of Ph. Eur. BRP batch 4 was necessary in order to replace Ph. Eur. BRP batch 3, the stocks of which were dwindling. 8 laboratories provided results. Ph. Eur. BRP batch 4 was calibrated against the 5th International Standard for inactivated rabies vaccine in International Units (IU) using the vaccination challenge method of the Ph. Eur. monograph 0451. The International Standard (IS), Ph. Eur. BRP batch 4 and batch 3 are all freeze dried vaccines prepared by beta-propiolactone inactivation of the Pitman Moore strain of rabies. Based on the results of the study, a potency of 11 IU/vial was assigned to Ph. Eur. BRP batch 4 for rabies vaccine (inactivated) for veterinary use. Nevertheless, it was noted that the vaccination challenge assay used as the "golden standard" for potency determination of inactivated rabies vaccines for veterinary use is a crude assay requiring the use of a large number of animals. Evidence from this study and from the collaborative study to establish Ph. Eur. BRP batch 3 suggests that the assay is difficult to perform and provides highly variable results. The validation of a suitable in vitro alternative is therefore highly recommended, as is the possible improvement of the in vivo assay, which will most likely remain the "golden" standard. PMID- 15659283 TI - Collaborative study for the establishment of erythropoietin BRP batch 2. AB - The preparation and establishment of the 2nd European Pharmacopoeia (Ph. Eur.) Biological Reference Preparation (BRP) for erythropoietin was the goal of a project run within the framework of the European Biological Standardisation Programme. The project, coded BSP062, was carried out between October 2002 and July 2003. The candidate preparation (cBRP2) was prepared in a similar manner to the first BRP batch (BRP1), as follows: -50:50 (weight/weight) blending of the two erythropoietin preparations currently available on the European market (epoietin-alpha and epoietin-beta), -lyophilisation using a protein-free carrier formulation to allow use of the standard for both biological and physico-chemical assay methods, -each vial contains approximately 250 microg erythropoietin. The cBRP2 was analysed in a collaborative study, carried out with the following aims: -to calibrate cBRP2 by in vivo bioassay in terms of the International Standard for erythropoietin, and assign a unitage, -to demonstrate continuity of unitage with BRP1, -to evaluate the suitability of cBRP2 to serve as a reference material for physico-chemical tests of erythropoietin. The collaborative study involved 14 laboratories both from Europe, and from Australia, Canada, South-Korea and the United States of America. Participants carried out biological and physicochemical assays on the candidate BRP batch 2, using BRP 1 and the 2nd World Health Organization (WHO) International Standard (IS) for recombinant erythropoietin as the reference standards. It was demonstrated that: -an assigned potency of 32,500 U per vial would maintain continuity between BRP1 and BRP2 in terms of the IS for erythropoietin, -the replacement batch was appropriate for use as erythropoietin BRP in the context of the control of erythropoietin concentrated solutions according to the Ph. Eur. monograph 1316. In July 2003, the Ph. Eur. Commission established the proposed standard as 'Erythropoeitin BRP batch 2' for use as a reference preparation for the polycythaemic and normocythaemic mouse bioassay, with an assigned potency of 32,500 U/vial, the identification by capillary zone electrophoresis (CZE), by polyacrylamide gel electrophoresis, immunoblotting and peptide mapping and as a reference for checking the system suitability of size exclusion chromatographic procedures used in the test for 'Dimers and related substances of higher molecular mass'. PMID- 15659285 TI - Capillary electrophoresis for the control of impurities of rDNA somatropin. AB - In the European Pharmacopoeia, the monographs on somatropin, somatropin bulk solution and somatropin for injection prescribe a number of tests including "related substances", "dimer and related substances of higher molecular weight" and "isoform distribution" which are intended to control the levels of impurities in the substance. The aim of this study was to verify the robustness of a new method by capillary electrophoresis and to compare its performance with that of the existing test for "isoform distribution" by isoelectric focusing. It was demonstrated that the capillary electrophoresis method was superior to the method of isoelectric focusing. The interest of the CZE method consists in the resolution of related impurities that might be process specific and/or generated by the expression system. PMID- 15659284 TI - Somatropin and its variants: structural characterization and methods of analysis. AB - Human Growth hormone (hGH, somatotrophin) is a 22 kDa, 191 amino-acid single chain protein produced by somatroph cells of the anterior pituitary gland. It is the major physiological regulator of growth, and deficiencies in growth hormone levels have long been recognized as the underlying cause of growth disorders (dwarfism). The ability of exogenous hGH to restore normal rates of growth in both human and animal models of growth retardation has long been recognized and the use of hGH in therapy goes back several decades. Initial preparations were prepared by extraction and purification from cadaveric pituitary tissue, and since 1984, hGH has been prepared by recombinant Deoxyribosenucleic acid (rDNA) technology. As is usually the case with "biologicals", characterization of the drug substance depended on a combination of physico-chemical and biological methods, and the hGH molecule became well characterized fairly early in its life as a drug. Indeed, by 1980 the major degradation forms and structural variants of the hGH molecule had been described and reviewed. Little satisfactory progress had been made in refining biological assays for hGH, and, although in vitro assays were described, potency-defining assays remained dependant on the whole body growth response in rats, and were both invasive and imprecise. In the early 1990's a series of collaborative studies on analysis of recombinant hGH (somatropin) established that available bioassays were much less selective that physico-chemical methods in detecting and quantifying structural degradation, and 1994 saw an international consensus to replace the bioassays with physico chemical analytical methods for the routine batch release of somatropin. During the last decade in most markets somatropin has, unusually for a protein, been subject to batch release and control dependent entirely on physico-chemical analysis, without the routine use of any form of bioassay. During that time there has been a continuous development and refinement of methods, and the identification of a range of structural variants and degradation products of the molecule. The present review sets out to summarise the current knowledge on physico-chemical analytical methods for somatropin, and the structural variants that have been identified and characterized. A survey of available biological analytical methods is beyond the scope of this review, as is consideration of the earlier pituitary preparations and the recombinant 192 amino-acid methionyl form of the molecule (somatrem), although it is likely that many of the methods and variants described would be equally applicable to somatrem. PMID- 15659286 TI - Collaborative study to establish the Low-molecular-mass heparin for assay- European Pharmacopoeia Biological Reference Preparation. AB - Thirty laboratories participated in a collaborative study to calibrate replacements for the 1st International Standard for Low Molecular Weight Heparin and the European Pharmacopoeia Low-molecular-mass heparin for assay Biological Reference Preparation. Two freeze-dried materials and one liquid preparation were included in the study. All three samples gave excellent intra- and inter laboratory variations (majority of mean % geometric coefficient of variation < 10 %) when assayed against the 1st International Standard by both anti-Xa and anti IIa assays. There were no major differences found between potency estimates using all methods and that obtained using European Pharmacopoeia method only. Overall, this study showed that the differences between the candidates are marginal. Based on the results of the study Sample B, 01/608 was established as the 2nd International Standard for Low Molecular Weight Heparin. Sample A, 01/592 and sample C, the liquid preparation, were established as replacements for the European Pharmacopoeia 'Low-molecular-mass heparin for assay' Biological Reference Preparation. PMID- 15659287 TI - N6-cyclopentyladenosine inhibits proliferation of murine haematopoietic progenitor cells in vivo. AB - Effects of N6-cyclopentyladenosine (CPA), the selective adenosine A1 receptor agonist, on bone marrow haematopoietic progenitor cells for granulocytes and macrophages (CFC-GM) were investigated by utilizing the model of haematopoietic damage induced by 5-fluorouracil. Experiments were performed in vivo on B10CBAF1 mice. A single i.p. injection of CPA at the optimum dose of 200 nmol/kg administered 22 h before a single injection of 5-fluorouracil (100 mg/kg, i.p.) protected CFC-GM against the cytotoxic damage as determined 4 days later. Isomolar doses of the selective agonists for adenosine A2A receptors, i.e. 2-p-(2 carboxyethyl)-phenethylamino-5'-N-ethylcarboxamidoadenosine, and for adenosine A3 receptors, i.e. N6-(3-iodobenzyl)adenosine-5'-N-methyluronamide, did not induce such effects. Because 5-fluorouracil is a cell cycle-specific drug damaging mainly cells in the S-phase, protective effects of CPA can be explained by its inhibitory action on the cell cycling. This interpretation was confirmed by experiments demonstrating that repeated administration of CPA in the hyperproliferation phase of the recovering haematopoiesis after 5-fluorouracil treatment inhibited transiently restoration of CFC-GM counts. PMID- 15659288 TI - Enhancement of benzodiazepine binding sites following chronic treatment with flumazenil. AB - The aim of this study was to improve our knowledge of the mechanisms leading to adaptive changes in gamma-aminobutyric acid(A) (GABA(A)) receptors following chronic drug treatment. Exposure (48 h) of human embryonic kidney (HEK 293) cells stably expressing recombinant alpha1beta2gamma2S GABA(A) receptors to the antagonist of benzodiazepine binding sites, flumazenil (5 microM), enhanced the maximum number (B(max)) and the equilibrium dissociation constant (K(d)) of [3H]flunitrazepam binding sites. The flumazenil-induced enhancement in B(max) was potentiated by GABA (50 microM) and reduced by the GABA(A) receptor antagonist, bicuculline (100 microM). Flumazenil-induced enhancement in K(d) was affected by neither of these treatments. GABA (50 microM) enhanced the density of [3H]flunitrazepam binding sites, and this enhancement was greater in the presence of diazepam (1 microM). The results suggest that chronic flumazenil treatment up regulates in a bicuculline-sensitive manner benzodiazepine binding sites at stably expressed GABA(A) receptors. PMID- 15659289 TI - Low affinity block of native and cloned hyperpolarization-activated Ih channels by Ba2+ ions. AB - Ba2+ is commonly used to discriminate two classes of ion currents. The classical inward-rectifying K+ current, I(Kir), is blocked by low millimolar concentrations of Ba2+, whereas the hyperpolarization-activated cation current, I(h), is assumed not to be sensitive to Ba2+. Here we investigated the effects of Ba2+ on I(h) currents recorded from rat hippocampal CA1 pyramidal neurons, and on cloned I(h) channels composed of either HCN1 or HCN2 subunits transiently expressed in Human Embryonic Kidney (HEK) 293 cells. The results show that low millimolar concentrations of Ba2+ reduce the maximal I(h) conductance (IC50 approximately 3 5 mM) in both CA1 pyramidal neurons and in HEK 293 cells without specificity for HCN1 or HCN2 subunits. In addition, Ba2+ decreases the rate of activation and increases the rate of deactivation of I(h) currents. Neither the half-maximal voltage of activation, V(h), nor the reversal potential of the I(h) channels were affected by Ba2+. The combined results suggest that B2+, at concentrations commonly used to block I(Kir) currents, also reduces the conductance of I(h) channels without subunit specificity, and affects the kinetics of I(h) channel gating. PMID- 15659290 TI - Cortistatin modulates memory evocation in rats. AB - The neurochemical control of learning depends on several neurotransmitters, hormones, and neuropeptides. Cortistatin is a neuropeptide with sleep-modulating properties that regulates memory consolidation and evocation. Several reports have suggested that learning processes are expressed under diurnal variations; therefore, it seems that the efficiency to solve learning tasks is related to the arousal state. Although we know that cortistatin modulates learning, we do not know whether its effect is subjected to diurnal variations. Hence, we evaluated memory evocation and the sleep-waking cycle along the day. Additionally, we evaluated the effect of cortistatin on motor control and cyclic adenosine monophosphate (cAMP) concentration. Performance of rats was better at 01:00 h than at 13:00 h to solve the Barnes maze. Cortistatin impaired memory evocation, increased rapid-eye-movement (REM) sleep, and decreased wakefulness at 01:00 h, whereas increasing it at 13:00 h. Cortistatin blunts cAMP concentration and impairs motor control at 13:00 h. These results support further a cortistatin modulatory role in the memory process. PMID- 15659291 TI - A tachykinin NK1 receptor antagonist attenuates the 4 beta-phorbol-12-myristate 13-acetate-induced nociceptive behaviour in the rat. AB - Antinociceptive effect of a tachykinin NK1 receptor antagonist ezlopitant [(2S,3S cis)-2-(diphenylmethyl)-N-{(2-methoxy, 5-isopropylphenyl)methyl}-1 azabicyclo[2.2.2]octan-3-amine] was investigated in the 4beta-phorbol-12 myristate-13-acetate (PMA)-induced nociceptive test in the rat. Intraplantar injection of PMA-induced paw-licking and flinching behaviour lasted up to 120 min and was accompanied by inflammatory reactions, such as swelling and invasion of granulocytes. Pretreatment with resiniferatoxin [200 microg/kg, subcutaneous (s.c.)] blocked the PMA-induced nociceptive behaviour, suggesting that vanilloid VR1 receptor-expressing primary sensory neurons play a major role in this response. Subcutaneous pretreatment with ezlopitant (0.3-30 mg/kg) and morphine (0.3-6 mg/kg) caused a dose-dependent inhibition of the behaviour. Ezlopitant (3 30 mg/kg) given subcutaneously after PMA injection also significantly attenuated the behavioural response. When administered intrathecally, ezlopitant and a nonselective glutamate receptor antagonist MK-801 had an inhibitory effect, whereas CJ-12,191, an inactive isomer of ezlopitant, was unaffected. These results suggest that spinal tachykinin NK1 receptors contribute to processing of ongoing pain associated with peripheral inflammation. PMID- 15659292 TI - 3-Chloro,4-methoxyfendiline is a potent GABA(B) receptor potentiator in rat neocortical slices. AB - Using grease-gap recording from rat neocortical slices, the GABA(B) receptor agonist baclofen elicited reversible and concentration-dependent hyperpolarizing responses (EC50=18+/-2.3 microM). The hyperpolarizations were antagonised by the GABA(B) receptor antagonist Sch 50911 [(+)-(S)-5,5-dimethylmorpholinyl-2-acetic acid). (+)-N-1-(3-chloro-4-methoxyphenyl)ethyl-3,3-diphenylpropylamine (3 chloro,4-methoxyfendiline; 3-Cl,4-MeO-fendiline) reversibly potentiated baclofen induced hyperpolarizing responses, which were reduced by Sch 50911, producing leftward shifts of the baclofen concentration-response curves, with a marked increase in the maximal hyperpolarization (EC50=2+/-0.5 microM). In slices preincubated with either [3H]GABA or [3H]glutamic acid, 3-Cl,4-MeO-fendiline (1 microM) potentiated the inhibitory effect of baclofen (2 microM) on the electrically evoked release of [3H]GABA and had a similar effect on the release of [3H]glutamic acid at a concentration of 0.5 microM, without affecting the basal release. These effects were blocked by Sch 50911 (10 microM). Our findings suggest that 3-Cl,4-MeO-fendiline is a potent potentiator of pre- and postsynaptic GABA(B) receptor-mediated functions. PMID- 15659293 TI - Nociceptin/orphanin FQ prevents the antinociceptive action of paracetamol on the rat hot plate test. AB - Nociceptin/orphanin FQ (N/OFQ) is involved in many behavioural patterns; in particular, it exerts a modulating effect on nociception. Like other proposed antiopiates, nociceptin/orphanin FQ has been shown to have analgesic, hyperalgesic as well as antianalgesic properties. Among the various effects proposed on nociceptive sensitivity at supraspinal level, the antagonistic activity toward morphine analgesia seems to be of interest. Therefore, we decided to investigate whether nociceptin/orphanin FQ and [Arg14, Lys15] nociceptin/orphanin FQ (R-K, a nociceptin analogue) can have the same effect on the analgesia produced by nonopioid analgesics. In this study, we examined the antianalgesic effect of nociceptin/orphanin FQ and its analogue R-K on paracetamol-induced analgesia and evaluated by means of the hot plate test in rats. Nociceptin/orphanin FQ was intracerebroventricularly administered, and, after 5 min, a dose of 400 mg/kg paracetamol was injected intraperitoneally, 30 min before the hot plate test. Nociceptin/orphanin FQ and R-K showed a dose dependent antagonism on the antinociceptive effect of paracetamol, and the activity of both drugs was significantly reduced by the antagonist [Nphe1] Arg14, Lys15-N/OFQ-NH2 (UFP-101). These data indicate that nociceptin/orphanin FQ and R K have an antianalgesic effect on the analgesia produced by a nonopioid analgesic drug, like paracetamol, that seems to develop within the brain. PMID- 15659294 TI - Effects of nefiracetam on the levels of brain-derived neurotrophic factor and synapsin I mRNA and protein in the hippocampus of microsphere-embolized rats. AB - Our recent study demonstrated that nefiracetam, N-(2,6-dimethylphenyl)-2-(2-oxo-1 pyrrolidinyl) acetamide, prevented impairment of the cyclic AMP (cAMP)/cAMP responsive element binding (CREB) protein signaling pathway in sustained cerebral ischemia. The purpose of the present study was to determine whether nefiracetam has an effect on the expression of brain-derived neurotrophic factor (BDNF) and synapsin I mRNAs that are believed to be produced via CREB, and the alteration in their protein contents in the hippocampus after cerebral ischemia. Sustained cerebral ischemia was induced by injection of 700 microspheres into the right hemisphere of each rat. The rats were treated once daily with 10 mg/kg nefiracetam, p.o., from 15 h after the operation. Treatment with nefiracetam reduced the prolongation of the escape latency in the water maze test on days 7-9 after microsphere embolism-induced sustained cerebral ischemia, suggesting an improvement in the spatial learning function. Microsphere-embolized rats on day 5 showed decreases in BDNF and synapsin I mRNA levels and their protein contents in the ipsilateral hippocampus. Treatment with nefiracetam partially attenuated the decreases. These results suggest that enhancement of BDNF and synapsin I expression by nefiracetam treatment may be, at least in part, due to the improvement in the CREB binding activity, contributing to the prevention of learning and memory dysfunction after sustained cerebral ischemia. PMID- 15659295 TI - Methamphetamine sensitization in nociceptin receptor knockout mice: locomotor and c-fos expression. AB - The role of endogenous nociceptin in the development and expression of sensitization to repeated methamphetamine administration in a novel environment was studied in nociceptin receptor knockout mice. No differences in acute or sensitized locomotor responses were found in nociceptin receptor knockout mice. However, analysis of c-fos expression revealed significant interactions between chronic methamphetamine treatment and genotype in the nucleus accumbens and lateral septum. This was due to increased c-fos expression in chronically methamphetamine-treated nociceptin receptor knockout mice contrasted with reduced c-fos expression in chronically vehicle-treated nociceptin receptor knockout mice. Two further regions (nucleus accumbens core and ventromedial caudate putamen) showed significant interactions between genotype, chronic, and acute methamphetamine treatment due to accentuated c-fos expression in nociceptin receptor knockout mice sensitized and challenged with methamphetamine. These findings suggest endogenous nociceptin modulates the response of the central nervous system to repeated psychostimulant administration, although this is little reflected in locomotion. PMID- 15659296 TI - Celecoxib does not induce convulsions nor does it affect GABAA receptor binding activity in the presence of new quinolones in mice. AB - We sought to determine whether celecoxib would induce convulsions when coadministered with new quinolone antimicrobial agents in mice. The oral administration of celecoxib (500 mg/kg) alone or in combination with enoxacin (500 mg/kg), lomefloxacin (1000 mg/kg), ciprofloxacin (1000 mg/kg), or levofloxacin (1000 mg/kg) induced no convulsions in mice. In contrast, some nonsteroidal anti-inflammatory drugs (NSAIDs), fenbufen (200 mg/kg), indomethacin (500 mg/kg), and naproxen (500 mg/kg) induced convulsions in combination with the majority of the new quinolones tested. gamma-Aminobutyric acid (GABA)(A) receptor blockade-mediated neuronal excitation is assumed to be involved in these toxic convulsions. Enoxacin (100 microM) and lomefloxacin (100 microM) only slightly reduced [3H]muscimol binding to GABA(A) receptors in mouse whole brain membrane. However, these reductions were markedly enhanced by the addition of fenbufen (100 microM), indomethacin (100 microM), or naproxen (100 microM). Conversely, celecoxib (100 microM) had no apparent effect on [3H]muscimol binding when applied alone or in combination with enoxacin or lomefloxacin. These results suggest that celecoxib may be a more desirable anti-inflammatory agent with respect to drug interactions with new quinolones compared with some conventional NSAIDs. PMID- 15659297 TI - Estrogen attenuates hypoxic-ischemic brain injury in neonatal rats. AB - Estrogen is neuroprotective in adult animals. We wished to determine if estrogen protects against brain injury in the newborn. Four-day-old rat pups were treated with subcutaneously implanted pellets containing 0.05 mg (2.4 microg/day) of 17beta-estradiol or vehicle, designed to release the estrogen over 21 days. At 7 days old the pups had the right carotid artery ligated followed by 2.5 h of 8% oxygen. Brain damage was evaluated by weight deficit of the right hemisphere at 22 days following hypoxia. Estradiol treatments reduced brain weight loss from 17.4+/-2.8% S.E.M. in the vehicle group (n=32) to -9.3+/-2.7% in the treated group (n=32, P<0.05). Brain cortex thiobarbituric acid reacting substances and caspase activities were assessed 24 h after reoxygenation. Estradiol significantly reduced a hypoxia-induced increase in brain thiobarbituric acid reactive substances (P<0.05). Levels of caspase-3, -8 and -9 activity increased due to hypoxia-ischemia. Estradiol had no effect on caspase activity. Estradiol reduced brain injury in the neonatal rat. PMID- 15659298 TI - Broad analgesic profile of buprenorphine in rodent models of acute and chronic pain. AB - Buprenorphine is a potent opioid analgesic clinically used to treat moderate to severe pain. The present study assessed its analgesic efficacy in a broad range of rodent models of acute and chronic pain. In the phenylquinone writhing, hot plate, and tail flick mouse models of acute pain, full analgesic efficacy was obtained (ED50 values: 0.0084-0.16 mg/kg i.v.). Full analgesic efficacy was also obtained in yeast- and formalin-induced inflammatory pain (ED50 values: 0.0024 0.025 mg/kg i.v., rats and mice) and in mustard-oil-induced spontaneous pain, referred allodynia, and referred hyperalgesia in mice (ED50 values: 0.018-0.025 mg/kg i.v.). Buprenorphine strongly inhibited mechanical and cold allodynia in mononeuropathic rats, as well as mechanical hyperalgesia and cold allodynia in polyneuropathic rats (ED50 values: 0.055 and 0.036 mg/kg i.v. and 0.129 and 0.038 mg/kg i.p., respectively). It is concluded that buprenorphine shows a broad analgesic profile and offers the opportunity to treat different pain conditions, including neuropathic pain. PMID- 15659300 TI - Effect of cyclosporin A on morphine-induced place conditioning in mice: involvement of nitric oxide. AB - Cyclosporin A is shown to attenuate antinociceptive effects of morphine, development and expression of morphine-induced tolerance and dependency via nitric oxide (NO) pathway. In the present study, the effect of systemic cyclosporin A on morphine-induced conditioned place preference (CPP) and the probable involvement of nitric oxide were assessed in mice. Our data showed that administration of morphine (1, 2.5, 5, 7.5, 10 mg/kg) significantly increased the time spent in the drug-paired compartment in a dose-dependent manner. The maximum response was obtained with 5 mg/kg of morphine. Cyclosporin A (5, 10 mg/kg) and N(G)-nitro-L-arginine methyl ester (L-NAME; 2.5, 5, 10 mg/kg), a nonselective nitric oxide synthase (NOS) inhibitor, did not induce either conditioned place preference or conditioned place aversion (CPA), while cyclosporin A (20 mg/kg) induced CPA. Both cyclosporin A (10, 20 mg/kg) and L-NAME (5, 10 mg/kg), in combination with morphine (5 mg/kg) during conditioning, significantly suppressed acquisition of morphine-induced place preference. Lower and per se noneffective doses of Cyclosporin A (1, 2.5, 5 mg/kg) and L-NAME (2.5 mg/kg), when coadministered, exerted a significant potentiating effect on the attenuation of morphine-induced place preference. Aminoguanidine (50, 100 mg/kg), the specific inducible nitric oxide synthase (iNOS) inhibitor, whether alone or in combination with cyclosporin A failed to show this inhibitory effect on morphine-induced place preference. In conclusion, decreasing nitric oxide production through inhibiting constitutive nitric oxide synthase may be a mechanism through which cyclosporin A attenuates morphine-induced place preference. PMID- 15659301 TI - Effects of clonidine injections into the bed nucleus of the stria terminalis on fear and anxiety behavior in rats. AB - Emotions such as fear and anxiety are mediated by a neural network containing nuclei like the amygdala, the bed nucleus of the stria terminalis and the periaqueductal gray. Noradrenaline is a neurotransmitter closely connected with the processing of stimuli eliciting these emotions. The bed nucleus of the stria terminalis contains the highest density of noradrenaline within the brain. In the present study, we investigated effects of injections of the noradrenergic alpha2 adrenoceptor agonist clonidine into the bed nucleus of the stria terminalis on learned and unlearned fear (anxiety) in rats on different animal models of fear and anxiety: acquisition and expression of fear-potentiated startle, sensitization of the acoustic startle response by foot shocks and light-enhanced startle. Clonidine injections disrupted acquisition and expression of fear potentiated startle, as well as light-enhanced startle, whereas sensitization was not affected. These results indicate that noradrenaline within the bed nucleus of the stria terminalis mediates both fear and anxiety. We suggest that there is rather a neurochemical than a neuroanatomical dissociation between learned fear and anxiety as hypothesized by Walker and Davis (Walker, D.L. and M. Davis, 1997b, Double dissociation between the involvement of the bed nucleus of the stria terminalis and the central nucleus of the amygdala in startle increases produced by conditioned versus unconditioned fear, J. Neurosci. 17, 9375-9383.). PMID- 15659299 TI - Antidepressant-like effects of neurokinin receptor antagonists in the forced swim test in the rat. AB - Although a wide assortment of agents is currently available for the treatment of depression, this disorder remains poorly managed in a large proportion of patients. Traditional antidepressant treatments target the biogenic amine systems. However, a growing body of evidence is implicating the involvement of neuropeptides in depression, especially the neurokinin substance P. This study evaluated the effects of selective antagonists of the tachykinin NK1, NK2, and NK3 receptors in the forced swim test, a commonly used screen for antidepressants. Rats were given CP-96,345 (2S, 3S)-cis-2-(diphenylmethyl)-N-[(2 methoxyphenyl)-methyl]-1-azabicyclo[2.2.2]octan-3-amine, SR 48968 (S)-N-methyl N[4-(4-acetylamino-4-phenylpiperidino)-2-(3,4-dichlorophenyl)-butyl]benzamide, or SR 142801 (S)-(N)-(1-(3-(1-benzoyl-3-(3,4-dichlorophenyl) piperidin-3-yl) propyl) 4-phenylpiperidin-4-yl)-N-methylacetamide, antagonists of the NK1, NK2, and NK3 receptors, respectively, at doses of 2.5, 5, and 10 mg/kg, intraperitoneally (i.p.). The time of immobility during the forced swim test was used as an indicator of antidepressant activity of the antagonists. All antagonists decreased immobility times. CP-96,345 and SR 142801 showed dose-related effects; SR 48968 had its maximum effect at 2.5 mg/kg. The magnitude of the effects of the neurokinin receptor antagonists was approximately the same as that of amitriptyline and desipramine, two traditional antidepressants, both given at 10 mg/kg, i.p. This study provides comparative data on the relative effectiveness of NK1, NK2, and NK3 receptor antagonists in this screen for antidepressant drug activity. PMID- 15659302 TI - Characterization of the effects of corticotropin-releasing factor on prepulse inhibition of the acoustic startle response in Brown Norway and Wistar-Kyoto rats. AB - Sensori-motor gating, as assessed by prepulse inhibition of the startle response is diminished in patients with schizophrenia. We have previously shown that inbred Brown Norway (BN) rats display significantly less prepulse inhibition of the acoustic startle response than inbred Wistar-Kyoto (WKY) rats, and that prepulse inhibition is decreased by central administration of the neuropeptide, corticotropin-releasing factor (CRF) in both strains. The present study was conducted to establish whether peripheral administration of CRF alters prepulse inhibition, whether a low, threshold dose for decreasing prepulse inhibition is the same in the two rat strains, and whether central administration of a CRF receptor antagonist enhances prepulse inhibition in the BN strain. CRF-induced behavioral activation was also examined to determine whether the two rat strains are differentially sensitive to a behavioral effect of CRF that does not involve the startle response. In each experiment, BN rats showed significantly less prepulse inhibition than WKY rats. Subcutaneous administration of CRF had no affect on startle amplitude or prepulse inhibition of the startle response in either rat strain. In BN, but not in WKY rats, low-dose CRF (0.3 microg) decreased prepulse inhibition. However, doses of CRF that did not alter prepulse inhibition in the WKY strain, did result in behavioral activation. No dose of CRF tested affected baseline startle amplitude. Central administration of the CRF receptor antagonist, astressin had no effect on prepulse inhibition or startle amplitude in either rat strain. Central administration of the CRF receptor antagonist, D-Phe CRF (12-41) had no effect on prepulse inhibition in WKY rats, resulted in a only a small, non-significant increase in prepulse inhibition in BN rats, while it decreased startle amplitude. The results suggest that CRF reduces prepulse inhibition of the acoustic startle response independently of effects on the pituitary-adrenal axis, and that endogenous CRF has at most, a minor role in the low prepulse inhibition found in BN rats. PMID- 15659303 TI - Wortmannin inhibits the myofilament Ca2+ sensitization induced by endothelin-1. AB - Endothelin-1 induces a positive inotropic effect due to a combination of an increase in Ca2+ transients and myofilament Ca2+ sensitivity in rabbit ventricular myocardium. We carried out the experiments to examine the potential contribution of myosin light chain kinase to the Ca2+ sensitization induced by endothelin-1 by use of wortmannin that inhibits myosin light chain kinase at high concentrations (IC50=200 nM). Wortmannin at 3 microM suppressed the basal force of contraction, but did not affect the positive inotropic effect mediated by beta adrenoceptors. Wortmannin at 1 and 3 microM markedly inhibited the positive inotropic effect of endothelin-1, but did not affect the increase in Ca2+ transients elicited by endothelin-1. The present findings imply that the increase in myofilament Ca2+ sensitivity induced by endothelin-1 may be in part due to activation of myosin light chain kinase in rabbit ventricular myocardium. PMID- 15659304 TI - Intravenous administration of conivaptan hydrochloride improves cardiac hemodynamics in rats with myocardial infarction-induced congestive heart failure. AB - We investigated the effects of intravenously administered conivaptan hydrochloride, a dual vasopressin V1A and V2 receptor antagonist, on cardiac function in rats with congestive heart failure following myocardial infarction, and compared results with those for the selective vasopressin V2 receptor antagonist SR121463A. Rats were subjected to left coronary artery occlusion to induce myocardial infarction, which in turn led to congestive heart failure. At 4 weeks after coronary occlusion, conivaptan (0.03, 0.1 and 0.3 mg/kg i.v.) dose dependently increased urine volume and reduced urine osmolality in both myocardial infarction and sham-operated rats. SR121463A (0.3 mg/kg i.v.) also increased urine volume and decreased urine osmolality in myocardial infarction rats, to a degree comparable to that by conivaptan (0.3 mg/kg i.v.). At 6 weeks after surgery, myocardial infarction rats showed increases in right ventricular systolic pressure, right atrial pressure, left ventricular end-diastolic pressure and relative weights of the heart and the lungs, and a decrease in first derivative of left ventricular pressure (dP/dt(max))/left ventricular pressure, showing that congestive heart failure was well established. Conivaptan (0.3 mg/kg i.v.) significantly reduced right ventricular systolic pressure, left ventricular end-diastolic pressure, lung/body weight and right atrial pressure in myocardial infarction rats. Moreover, conivaptan (0.3 mg/kg i.v.) significantly increased dP/dt(max)/left ventricular pressure. SR121463A at a dose of 0.3 mg/kg i.v. significantly decreased left ventricular end-diastolic pressure and right atrial pressure, and tended to decrease right ventricular systolic pressure and relative lung weight in myocardial infarction rats. Although the aquaretic and preload reducing effects of SR121463A were similar to those of conivaptan, SR121463A failed to improve dP/dt(max)/left ventricular pressure. These results suggest that dual vasopressin V1A and V2 receptor antagonists provide greater benefit than selective vasopressin V2 receptor antagonists in the treatment of congestive heart failure. PMID- 15659306 TI - Pharmacokinetic and pharmacodynamic profiles of the antitussive principles of Glycyrrhizae radix (licorice), a main component of the Kampo preparation Bakumondo-to (Mai-men-dong-tang). AB - We examined the pharmacokinetic and pharmacodynamic properties of liquiritin apioside, a main antitussive component of Glycyrrhizae radix (licorice), with regard to its antitussive effect in guinea pigs. The peak plasma concentration of the unchanged compound was observed 15 min after the administration of liquiritin apiosaide. The plasma concentration then gradually decreased and was almost undetectable 4 h after administration. Liquiritigenin, a des-glycoside of liquiritin apioside, appeared in the plasma 2 h after the administration of liquiritin apioside and remained for more than 6 h after administration. The plasma concentration of unchanged liquiritigenin was observed 15 min after administration and then gradually increased for more than 6 h after administration. When the antitussive effects of liquiritin apioside, liquiritin and liquiritigenin, at respective doses of 30 mg/kg, p.o., were examined 1 h after administration, liquiritin apioside and liquiritigenin caused a significant reduction in the number of capsaicin-induced coughs. However, at the same dose, liquiritin had no significant effect on the number of capsaicin-induced coughs. On the other hand, when the antitussive effects of liquiritin apioside, liquiritin and liquiritigenin, at doses of 30 mg/kg, p.o., were examined 4 h after administration, each caused a more than 40% reduction in the number of capsaicin-induced coughs. The present results suggest that G. radix (licorice) may produce a persistent antitussive effect, and that liquiritin apioside plays an important role in the earlier phase, while liquiritigenin, which is a metabolite of liquiritin apioside and liquiritin, plays an important role in the late phase. PMID- 15659305 TI - Changes in production and metabolism of brain natriuretic peptide in rats with myocardial necrosis. AB - In this study, we employed rat model of acute myocardial necrosis induced by isoproterenol (ISO) to study the possible roles of corin, the protease uniquely distributing in myocardium to convert pro-brain natriuretic peptide (proBNP) to BNP, and neutral endopeptidase (NEP), the major enzyme to degrade BNP, in changing the levels of BNP. In rats with isoproterenol alone, the myocardium necrosis occurred and the cardiac function was inhibited; the BNP contents in plasma and myocardium were upregulated, so did the myocardial corin mRNA level; the NEP activity in plasma and myocardium were downregulated. Omapatrilat (OMA) treatment relieved myocardial lesions and improved cardiac function. In the plasma and myocardium, omapatrilat treatment increased BNP contents, reduced NEP activity; in myocardium, mRNA level of proBNP and corin decreased, but NEP mRNA expression increased. Our study confirmed that omapatrilat treated myocardial necrosis effectively and suggested that increased BNP in rats with myocardial necrosis could depend on increased production and conversion as well as decreased degradation. PMID- 15659308 TI - Inhibition of Rho-kinase stimulates nitric oxide-independent vasorelaxation. AB - Vasoconstrictor factors, like urotensin, angiotensin and catecholamines, activate Rho-dependent serine-threonine kinase (Rho-kinase) and inhibition of this pathway represents a novel therapy for cardiovascular diseases with hypertensive syndrome. The disbalance of relaxing endothelial nitric oxide (NO)-producing and vasoconstrictive pathways can be especially important in diseases where hypertension is accompanied by endothelial dysfunction that compromises NO generation. However, a recent study reported that the efficacy of the Rho-kinase inhibitor (R)-(+)-trans-N-(4-Pyridyl)-4-(1-aminoethyl)cyclohexanecarboxamide (Y27632) is dramatically attenuated upon removal of endothelium or inhibition of endothelial NO synthase (eNOS). This raises the question whether Rho-kinase inhibition could be an effective treatment in case of hypertension associated with endothelial dysfunction. The purpose of the present study was to determine whether the vasorelaxing effect of Rho-kinase inhibition is mediated through eNOS dependent mechanisms. We show here that in the models of genetically reduced endothelial NO production (eNOS-/- mice and spontaneous hypertensive rats (SHR)) or in models of pharmacologically reduced endogenous NO production (N(omega) nitro-L-arginine methyl ester (LNAME) treatment), Rho-kinase inhibition induced a strong vasodilation and reduction of blood pressure indicating independence of Rho-kinase pathway from eNOS. An additional important finding of our study is that Rho-kinase inhibitors induce a strong vasorelaxation and blood pressure reduction upon intravenous injection not only in hypertensive but in normotensive animals, as well. Inhibition of Rho-kinase represents a promising possibility to treat hypertension that is accompanied by endothelial dysfunction. PMID- 15659307 TI - Halothane sensitizes the canine heart to pharmacological IKr blockade. AB - The effects of halothane and pentobarbital on the cardiovascular system were compared using the in vivo canine models. The ventricular repolarization process was longer under the halothane-anesthesia than pentobarbital-anesthesia. Intravenous administration of a selective blocker of rapidly activating delayed rectifier K+ currents (I(Kr)) sematilide prolonged the ventricular repolarization period without affecting the intraventricular conduction under both anesthesia; however, the potency was about 1.5-folds greater under the halothane-anesthesia than pentobarbital-anesthesia. These results suggest that halothane can more effectively sensitize the heart to pharmacological I(Kr) blockade, resulting in the excessive QT interval prolongation. Thus, the halothane-anesthetized canine model can be useful for predicting the in vivo I(Kr) blocking property of new drugs. PMID- 15659309 TI - Involvement of serotonin1A receptors in cardiovascular responses to stress: a radio-telemetry study in four rat strains. AB - We studied the effect of treatment with the serotonin-1A (5-HT1A) receptor ligands buspirone, 8-hydroxy-di-propyl-aminotetralin (8-OH-DPAT), and (8-[2-(2,3 dihydro-1,4-benzodioxin-2-yl-methylamino)ethyl]-8-azaspiro[4,5]decane-7,9-dione methyl sulphonate (MDL73,005EF) on blood pressure and heart rate increases to open field stress. We compared Spontaneously Hypertensive Rats (SHR), Fawn-Hooded (FH) rats, Wistar-Kyoto (WKY) rats, and Sprague-Dawley (SD) rats instrumented with radio-telemetry probes. Buspirone treatment reduced the blood pressure increase in SHR, FH rats, and WKY rats and heart rate increase in FH rats and WKY rats. 8-OH-DPAT treatment reduced the blood pressure increase in FH rats and WKY rats, but had no effect in SHR and enhanced the pressor response in SD rats. This treatment reduced the heart rate increase in FH rats and WKY rats only. Similarly, MDL73,005EF treatment reduced the blood pressure increase in FH rats and WKY rats, but had no effect in SHR and enhanced this response in SD rats. Little effect of this treatment was seen on heart rate changes. For comparison, diazepam treatment abolished the pressor response in SD rats and reduced it in FH rats and WKY rats, but not SHR. Differential effects of the treatments were also seen between strains for locomotor activity in the open field, although behavioural changes could not explain the effects of the drugs on cardiovascular responses. These data suggest that 5-HT1A receptors are involved in cardiovascular stress responses; however, the extent of this involvement differs between rat strains and the drugs used. These results could be important for our understanding of possible anxiolytic properties of antipsychotic drugs with affinity for the 5-HT1A receptor. PMID- 15659310 TI - Contractile action of levosimendan and epinephrine during acidosis. AB - We evaluated the inotropic actions of levosimendan and epinephrine, both singly and in combination, under isohydric (pH 7.4) and acidotic (pH 7.0) conditions in isolated guinea-pig hearts. Acidosis depressed contractility and myocardial relaxation by 25-30%, and both inotropes were less efficacious at pH 7.0, while their potencies were unaffected. In combination experiments, the presence of levosimendan increased the potency of epinephrine approximately 17-fold (pH 7.4) and 11-fold (pH 7.0), and the presence of epinephrine increased the potency of levosimendan approximately 12-fold (pH 7.4) and approximately 21-fold (pH 7.0). At pH 7.0, both inotropes augmented papillary muscle contraction to a similar extent, but in contrast to epinephrine, levosimendan non-significantly [corrected] raised cAMP levels. In conclusion, combining levosimendan with epinephrine helps to overcome the depressed inotropic actions of epinephrine during acidosis, suggesting that additional studies which might justify clinical evaluation of the concurrent use of the two agents should be performed. PMID- 15659311 TI - Vascular effects of delta 9-tetrahydrocannabinol (THC), anandamide and N arachidonoyldopamine (NADA) in the rat isolated aorta. AB - The vascular effects of cannabinoids have been compared in the rat isolated aorta. Delta9-Tetrahydrocannabinol (THC), anandamide and N-arachidonoyl-dopamine (NADA) all caused vasorelaxation to similar degrees in pre-constricted aortae. Vasorelaxation to THC was inhibited by in vivo pre-treatment with pertussis toxin (10 microg/kg) or with the synthetic cannabinoid CP55,940 (((-)-cis-3-[2-hydroxy 4-(1,1-dimethylheptyl)phenyl]-trans-4-(3-hydroxypropyl)cyclohexanol), acutely or chronically), exposure to capsaicin in vitro (10 microM for 1 h), and de endothelialisation. Vasorelaxation to anandamide was only inhibited by pertussis toxin and chronic CP55,940 pre-treatment (0.4 mg/kg for 11 days). Vasorelaxation to NADA was inhibited by pertussis toxin and chronic CP55,940 pre-treatment, and by de-endothelialisation. The vasorelaxant effects of the cannabinoids were not inhibited by cannabinoid CB1 receptor antagonism; however, vasorelaxation to both CP55,940 and THC was inhibited by cannabinoid CB2 receptor antagonism. Vasorelaxation to all cannabinoids was enhanced in the presence of indomethacin (10 microM). THC also caused vasoconstriction of the aorta while anandamide, NADA, CP55,940 and WIN 55,212-2 (R(+)-[2,3-dihydro-5-methyl-3 [(morpholinyl)methyl]pyrrolo[1,2,3-de]-1,4benzoxazin-yl]-(1 naphthalenyl)methanone mesylate) did not. The vasoconstrictor effects of THC were inhibited by in vivo pre-treatment with pertussis toxin or CP55,940, acute exposure to CP55,940, cannabinoid CB1 receptor antagonism and cyclooxygenase inhibition. These results demonstrate the opposing vascular effects of cannabinoids in the rat aorta, and although vasorelaxation to each of the cannabinoids is of similar magnitude, it is mediated through different pathways. This gives further indication of the different vascular actions of cannabinoid compounds. PMID- 15659312 TI - Cigarette smoke increases mucosal permeability in guinea pig trachea via tachykinin NK2 receptor activation. AB - We investigated whether exposure to cigarette smoke increases the mucosal permeability in guinea pig trachea and if this effect could be mediated by tachykinin NK2 receptor activation. Guinea pigs were exposed to either three different doses of cigarette smoke or room air. Mucosal permeability was measured by monitoring the rate of appearance in the circulation of horseradish peroxidase (HRP) that had been instilled into the isolated tracheal segment. Exposure to 20 and 30 puffs but not 10 puffs of cigarette smoke increased the tracheal mucosal permeability. Pretreatment with the tachykinin NK2 receptor antagonist SR48,968 [(S)-N-methyl-N[4-(4-acetylamino-4-phenylpiperidino)-2-(3,4 dichlorophenyl)butyl]benzamide] completely inhibited the increase in the permeability of the tracheal mucosa induced by exposure to cigarette smoke, whereas the tachykinin NK1 receptor antagonist SSR240,600 [(R)-2-(1-{2-[4-{2-[3,5 bis(trifluoromethyl)phenyl]acetyl}-2-(3,4-dichlorophenyl)-2-morpholinyl]ethyl}-4 piperidinyl)-2-methylpropanamide] and the tachykinin NK3 receptor antagonist SR142,801 [(S)-(N)-(1-[3-(1-benzoyl-3(3,4-dichlorophenyl)piperidine-3-yl)propyl] 4-phenylpiperidin-4-yl)-N-methyl-acetamide] had no effect. It is concluded that endogenous tachykinins via NK2 receptor activation mediate the increase in the permeability of the tracheal mucosa induced by exposure to cigarette smoke in guinea pigs. PMID- 15659314 TI - Involvement of reactive oxygen species and stress-activated MAPKs in satratoxin H induced apoptosis. AB - Satratoxins, members of the trichothecene mycotoxin family, have been known to be harmful to health. However, the mechanisms underlying the toxicity still remain unclear. The present study is undertaken to elucidate the mechanisms of the satratoxin H-induced cytotoxicity in PC12 cells. Satratoxin H caused cytotoxicity, which was reflected from apoptosis determined by chromatin staining and flow cytometry. Satratoxin H stimulated the phosphorylation of extracellular signal-regulated kinase (ERK), p38 mitogen-activated protein kinase (MAPK) and c Jun N-terminal kinase (JNK). Pre-incubation with SB203580, a p38 MAPK inhibitor, or SP600125, a JNK inhibitor, but not PD98059, an ERK inhibitor, reduced satratoxin-induced cytotoxicity. Co-incubation of cells with glutathione, N acetyl-L-cysteine or glutathione reductase inhibited cytotoxicity and the phosphorylation of p38 MAPK induced by satratoxin H. Our data suggest that satratoxin H-induced apoptosis in PC12 cells is dependent on the activation of p38 MAPK/JNK and the increase in reactive oxygen species. PMID- 15659313 TI - Role of tumor necrosis factor-alpha in down-regulation of hepatic cytochrome P450 and P-glycoprotein by endotoxin. AB - We investigated the role of tumor necrosis factor-alpha (TNF-alpha) in the down regulation of hepatic P-glycoprotein and cytochrome P450 (CYP) by endotoxin, using TNF-alpha gene-deficient (TNF-alpha-/-) mice. In the case of P glycoprotein, endotoxin (10 mg/kg) significantly decreased the expression of hepatic P-glycoprotein in wild-type mice 6 h, but not 24 h, after intraperitoneal injection, with no significant differences in the constitutional expression of P glycoprotein between wild-type mice and TNF-alpha-/- mice. However, endotoxin had no effect on the expression of P-glycoprotein in TNF-alpha-/- mice either 6 or 24 h after injection. When doxorubicin was administered intravenously to TNF-alpha-/ mice treated 6 h earlier with and without endotoxin, no significant differences in the plasma concentrations of doxorubicin 3 h after injection were observed between endotoxin-treated and untreated TNF-alpha-/- mice. These results suggest that TNF-alpha plays a pivotal role in the down-regulation of P-glycoprotein by endotoxin. In the case of CYP, the constitutive expression of hepatic CYP3A2 and CYP2C11 had a tendency to decline in TNF-alpha-/- mice compared with that in wild type mice. Endotoxin significantly decreased the expression of hepatic CYP3A2 and CYP2C11 in wild-type mice 24 h after injection, and that decreased expression was significantly greater in TNF-alpha-/- mice than wild-type mice. When antipyrine was administered intravenously to wild-type mice and TNF-alpha-/- mice treated 24 h earlier with endotoxin, the plasma concentrations of antipyrine in TNF-alpha-/- mice 3 h after injection were significantly higher than those in wild-type mice. These findings suggest that TNF-alpha plays a key role in endotoxin-induced down regulation of hepatic P-glycoprotein, as well as plays a protective role in the regulation of hepatic CYP3A2 and CYP2C11 against endotoxin-induced acute inflammatory response. In TNF-alpha-/- mice, other cytokines appear to function as compensation for the lack of endogenous TNF-alpha. PMID- 15659315 TI - Decreased sensory neuropeptide release in isolated bronchi of rats with cisplatin induced neuropathy. AB - We studied if attenuated neurogenic bronchoconstriction was associated with a change in sensory neuropeptide release in preparations from rats with cisplatin induced neuropathy. Electrical field stimulation (100 stimuli, 20 V, 0.1 ms, 20 Hz) induced an increase in the release of somatostatin, calcitonin gene-related peptide (CGRP) and substance P determined by radioimmunoassay from baseline 0.18+/-0.01, 0.17+/-0.01 and 0.86+/-0.02, to 0.59+/-0.02, 1.77+/-0.04 and 5.96 fmol/mg wet tissue weight, respectively, in organ fluid of tracheal tubes from rats. This was significantly attenuated to post-stimulation values of 0.36+/ 0.02, 0.45+/-0.02, 4.68+/-0.24 fmol/mg wet tissue weight for somatostatin, CGRP, and substance P, respectively, with a significant decrease in field stimulation induced contraction of bronchial preparations from animals 11 days after a 5-day treatment period with cisplatin (1.5 mg/kg i.p. once a day). The cisplatin treated animals developed sensory neuropathy characterized by a 40% decrease in femoral nerve conduction velocity. The results show that a decrease in tracheo bronchial sensory neuropeptide release associates with feeble bronchomotor responses in rats with cisplatin-induced sensory neuropathy. PMID- 15659316 TI - Topical antiinflammatory effects of the ether extract from Protium kleinii and alpha-amyrin pentacyclic triterpene. AB - Protium kleinii (Burseraceae), a native Brazilian medicinal plant is claimed to be useful to treat some inflammatory states. Now we reported that topical application of either the ether extract or the main active constituent from P. kleinii the pentacyclic triterpene alpha-amyrin, all caused a dose-related inhibition of both ear oedema (ID50 values are 0.55 and 0.31 mg/ear, respectively) and influx of polymorphonuclear cells (ID50 values are 0.72 and 0.45 mg/ear, respectively) in response to topical application of 12-O tetradecanoylphorbol-acetate (TPA) in the of mice ear. In terms of the efficacy, the maximal obtained inhibition for both ear oedema and neutrophil influx was very similar to that produced by the topical application of the steroidal antiinflammatory drug dexamethasone (DE; with inhibition of 70+/-5%, 66+/-3%, and 87+/-4% for oedema and 83+/-6%, 73+/-5%, and 91+/-3% for neutrophil influx, for the ether extract, alpha-amyrin, and dexamethasone, respectively). Likewise, both the ether extract and alpha-amyrin given topically dose-dependently prevented the increase of the proinflammatory cytokine interleukin-1beta levels in response to topical application of TPA. The calculated mean ID50 values are 1.81 and 0.53 mg/ear, respectively. Again, the efficacy of the extract and alpha-amyrin was very similar to that produced by dexamethasone (63+/-6%, 61+/-5%, and 74+/-5%, respectively). In marked contrast to phenidone, a lipo and cyclooxygenase inhibitor, neither the ether extract nor the alpha-amyrin inhibited arachidonic acid-mediated ear oedema in mice. Collectively, these results indicate that the active constituents present in the ether extract of P. kleinii including the pentacyclic triterpene alpha-amyrin are good candidates to develop a skin permeable antiinflammatory drug. PMID- 15659317 TI - Effects of bemiparin on airway responses to antigen in sensitized Brown-Norway rats. AB - Heparins have demonstrated activity in asthma. The effects of bemiparin, a low molecular weight heparin, were examined on antigen-induced responses in sensitized Brown-Norway rats. Inhaled bemiparin (1 mg/ml) reduced the acute bronchospasm produced by aerosol antigen, prevented airway hyperresponsiveness to 5-hydroxytryptamine postantigen exposure, and reduced the eosinophil count (from 0.205+/-0.062 to 0.054+/-0.016 x 10(6) cells/ml in antigen and antigen+bemiparin groups, respectively; P<0.05), eosinophil peroxidase activity, and proteins in the bronchoalveolar lavage fluid (BALF), as well as the transiently augmented mucin Muc5ac expression. Hyperresponsiveness to adenosine was not affected by bemiparin. In similar experiments, inhaled fondaparinux (1 mg/ml) did not affect the antigen-induced responses, while a low-anticoagulant low molecular weight heparin was effective. In conclusion, bemiparin showed beneficial effects in experimental asthma, probably unrelated to its anticoagulant activity, which extends the previous positive findings obtained with other heparins. PMID- 15659318 TI - An examination of neurogenic mechanisms involved in mustard oil-induced inflammation in the mouse. AB - The mechanisms by which topical mustard oil causes vasodilatation in the mouse were investigated using the tachykinin NK1 receptor antagonist SR140333 and the calcitonin gene-related peptide (CGRP) antagonist BIBN4096BS, alongside alphaCGRP or NK1 receptor knockout mice. Blood flow was assessed by laser Doppler flowmetry and plasma extravasation by 125I-albumin accumulation. Mustard oil produced significant plasma extravasation and vasodilatation in wild type mice, although the plasma extravasation was less than that seen with capsaicin whilst the vasodilatation was greater. The plasma extravasation was abolished in tachykinin NK1 knockout mice, whilst the vasodilatation was enhanced. BIBN4096BS was unable to inhibit the vasodilatation in wild type mice but abolished it in the NK1 knockout mice. In alphaCGRP knockout mice, mustard oil also caused plasma extravasation and vasodilatation, which were both inhibited by treatment with SR140333. These data suggest that both a tachykinin NK1 receptor agonist and a CGRP agonist are active as vasodilators, producing redundancy, requiring blockade of both mediators to prevent vasodilatation. PMID- 15659319 TI - Beneficial effects of GW274150 treatment on the development of experimental colitis induced by dinitrobenzene sulfonic acid. AB - Inflammatory bowel disease is associated with inducible nitric oxide synthase (iNOS) expression, oxidative and nitrosative stress, and leukocyte infiltration in the colon. Here, we investigate the effects of the selective iNOS-inhibitor (S)-2-amino-(1-iminoethylamino)-5-thiopentanoic acid (GW274150) on the development of experimental colitis induced by dinitrobenzene sulfonic acid. When compared to dinitrobenzene sulfonic acid-treated mice, GW274150 (5 mg/kg i.p.) treated mice subjected to dinitrobenzene sulfonic ACID-induced colitis experienced a significantly lower rate of the extent and severity of the histological signs of colon injury. Dinitrobenzene sulfonic acid-treated mice experienced hemorrhagic diarrhoea and weight loss. At 4 days after the administration of dinitrobenzene sulfonic acid, the mucosa of the colon exhibited large areas of necrosis. Immunohistochemistry for nitrotyrosine and poly (ADP ribose) (PAR) showed an intense staining in the inflamed colon. Treatment of dinitrobenzene sulfonic acid-treated mice with GW274150 significantly reduced the degree of hemorrhagic diarrhoea and weight loss caused by administration of dinitrobenzene sulfonic acid. GW274150 also caused a substantial reduction of (i) the degree of colon injury, (ii) the rise in myeloperoxidase (MPO) activity (mucosa), (iii) the increase in staining (immunohistochemistry) for nitrotyrosine, as well as (iv) PARP activation caused by dinitrobenzene sulfonic acid in the colon. Thus, GW274150 treatment reduced the degree of colitis caused by dinitrobenzene sulfonic acid. We propose that selective inhibition of iNOS activity with GW274150 may be useful in the treatment of inflammatory bowel disease. PMID- 15659320 TI - The effects of selective cyclooxygenase-2 inhibitors, celecoxib and rofecoxib, on experimental colitis induced by acetic acid in rats. AB - Several mediators may be involved in the pathogenesis of inflammatory bowel disease, as well as in experimental colitis. The present work was conducted to investigate the effects of the two selective cyclooxygenase-2 inhibitors, celecoxib and rofecoxib, on experimentally induced colitis in rats. Rectal instillation of acetic acid was used to induce the colitis. Acetic acid treatment caused haemorrhagic diarrhoea and weight loss in rats. Celecoxib (5 mg/kg) or rofecoxib (2.5 mg/kg), when given twice daily by the oral route, reduced the degree of haemorrhagic diarrhoea and the weight loss produced. In addition, they produced a significant reduction in the degree of colonic injury, the rise in myeloperoxidase (MPO) levels, total nitric oxide synthetase (NOS) activity, platelet-activating factor (PAF), histamine levels and prostaglandin E2 levels. In contrast, there was a significant increase in the levels of reduced glutathione (GSH). Thus, the findings of the present study provide evidence that selective cyclooxygenase-2 inhibitors may be beneficial in patients with inflammatory bowel disease. PMID- 15659321 TI - The herbal medicine Sho-saiko-to selectively inhibits CD8+ T-cell proliferation. AB - Sho-saiko-to (SST), a Chinese/Japanese traditional herbal medicine, has been widely used to treat chronic hepatitis in Japan, and the immunomodulatory properties of SST are likely to mediate its beneficial effect. In the present study, we examined the effects of SST and its various ingredients on the count and proliferation of T-cell subsets in cultured splenocytes and hepatic mononuclear cells. SST, wogonin-7-O-glucuronoside (a major SST ingredient), and wogonin (an intestinal metabolite of wogonin-7-O-glucuronoside) increased CD4/CD8 ratio via a decrease of CD8+ T-cell counts with no effect on CD4+ T-cell counts. Flow cytometric analyses of viability, proliferation, and cell cycle revealed that wogonin suppressed CD8+ T-cell proliferation without inducing cell death. SST and wogonin administered to mice increased the CD4/CD8 ratio in hepatic mononuclear cells but not in splenocytes. These findings suggest that SST may modulate the CD4/CD8 ratio via the selective inhibition of CD8+ T-cell proliferation by the SST ingredient wogonin-7-O-glucuronoside or its metabolite wogonin. PMID- 15659323 TI - Rat model of the hypercalcaemia induced by parathyroid hormone-related protein: characteristics of three bisphosphonates. AB - In our preliminary experiment, we found that a constant infusion of a high dose of parathyroid hormone-related protein induced both hyperphosphataemia and hypocalcaemia, secondary to renal dysfunction. Therefore, in this study, we developed two types of parathyroid hormone-related protein-induced hypercalcaemia models. One is the hypercalcaemia model, which did not show renal-dysfunction induced hypocalcaemia. This model might be suitable for estimating hypocalcaemic activities of drugs, especially of those that act on bone resorption. The other is the model for estimating histological changes, which is associated with renal dysfunction. We then used these models to investigate the effects of three different bisphosphonates. Since the hypercalcaemic effect of parathyroid hormone related protein infusion plateaued at 20 pmol/h, and higher doses of parathyroid hormone-related protein caused an elevation of blood urea nitrogen, the parathyroid hormone-related protein infusion rate was fixed at 20 pmol/h to avoid renal dysfunction and at 40 pmol/h to elicit renal dysfunction. The hypocalcaemic efficiencies of clodronate and etidronate were almost the same but pamidronate was 17.9 times more potent than clodronate. Additionally, both clodronate and pamidronate decreased the plasma concentrations of blood urea nitrogen and the Ca2+ times inorganic P product, whereas etidronate lacked these effects. Clodronate suppressed renal calcification and tubular dilatation in the renal dysfunction model. These data indicated that clodronate and pamidronate not only decrease the plasma Ca2+ concentration but also improve the renal dysfunction induced by hypercalcaemia. PMID- 15659322 TI - Hypothyroidism changes adrenoceptor- and muscarinic receptor-mediated blood pressure responses. AB - Hypothyroidism was induced by the administration of 0.03% methimazole to drinking water for 1, 2 or 6 weeks to study whether there is a change in adrenoceptor- and muscarinic receptor-mediated blood pressure responses in hypothyroid rats. After 1, 2 and 6 weeks of treatment, the pressor response to norepinephrine was progressively suppressed, and after 6 weeks a significant suppression was observed as compared to control. The depressor response induced by isoprenaline, acetylcholine or sodium nitroprusside was not significantly different between control and hypothyroid rats at any time. The pressor response induced by N(G) nitro-L-arginine (L-NOARG), an inhibitor of nitric oxide (NO) synthase, was significantly reduced in hypothyroid rats after 1, 2 or 6 weeks of treatment, and the magnitude of the reduction was almost the same for three groups. These results indicated that hypothyroidism causes a time-dependent decrease in pressor responses mediated by alpha-adrenoceptors, but a time-independent decrease in those induced by L-NOARG, and suggest that a progressive decrease in alpha adrenoceptor-mediated pressor responses occurs in hypothyroidism; however, the decrease in basal NO production and/or release in the peripheral vasculature already occurs in hypothyroid rats at an early stage of the disease. PMID- 15659324 TI - Role of macrophage migration inhibitory factor (MIF) in the skin. AB - Macrophage migration inhibitory factor (MIF) functions as a pleiotropic protein, participating in both inflammation and immune responses. MIF was originally discovered as a lymphokine involved in delayed type hypersensitivity and various macrophage functions, including phagocytosis, and tumor surveillance. Recently, MIF has been re-evaluated as a pro-inflammatory cytokine and identified as a pituitary-derived hormone, potentiating endotoxemia. MIF is ubiquitously expressed in various tissues, including the skin. Clinical evidence of increased MIF expression in inflammatory diseases supports this potential role of MIF in inflammation. In addition to its role in inflammation, MIF has been shown to exhibit growth-promoting activity, with anti-MIF antibodies effectively suppressing tumor growth and tumor-associated angiogenesis. This review presents the latest findings on the roles of MIF in the skin with regard to inflammation, the immune response, skin disease, tumorigenesis and cutaneous wound healing, and discusses its potential functions in various pathophysiological states. PMID- 15659325 TI - Hepatocyte growth factor; expression, concentration and biological activity in chronic leg ulcers. AB - BACKGROUND: Hepatocyte growth factor (HGF) is a multifunctional cytokine that is involved in recovery process after organ injuries. OBJECTIVE: We studied HGF and the membrane bound receptor, c-met locally in patients who suffered from chronic leg ulcers (> or =1 year) caused by venous insufficiency. METHODS: Skin biopsies from the edge of the ulcers were taken from patients (n=13) and studied by immunohistochemical staining for detection of HGF and c-met. Skin biopsies from healthy volunteers (n=10) were used as the control material. Ulcer secretion from chronic ulcers (n=11) was examined for the presence of HGF by ELISA and the concentration of HGF was compared with acute ulcers in healthy controls (n=10) and in patients operated for a non-invasive breast cancer (n=12). RESULTS: We observed that c-met expression in the ulcer area increased significantly in chronic ulcers compared to controls (p=0.005). Concentration of ulcer-HGF in the patients with chronic ulcer was significantly higher than acute ulcers (p<0.01). The biological activity of HGF in ulcer secret was assessed in-vitro in transferred, mouse skin epithelial cell monolayer. Enhanced migration and morphologic changes were seen after adding ulcer secret from acute ulcers (> 1 ng/mL) that was inhibited by anti-HGF antibodies. No biological activity was observed by adding ulcer secret from chronic ulcers irrespective HGF concentration. CONCLUSION: We conclude that in chronic skin ulcers decreased biological activity of endogenous HGF and overexpression of c-met is seen which might explain fibrosis and delayed recovery. Administration of exogenous active HGF might contribute to accelerated healing in these patients. PMID- 15659326 TI - Plectin deficient epidermolysis bullosa simplex with 27-year-history of muscular dystrophy. AB - BACKGROUND: Epidermolysis bullosa simplex associated with muscular dystrophy is caused by plectin deficiency. OBJECTIVE: To report clinical, immunohistochemical, ultrastructural and molecular features of a 52-year-old Japanese patient affected with this disease, whose muscular disease had been followed-up for 27 years. METHODS: We performed histopathological study, immunofluorescence, electron microscopic study and mutation detection analysis for plectin. RESULTS: The patient developed blisters and erosions followed by nail deformity on the traumatized regions from birth. The skin lesions were continuously developed to date. The histopathological study showed subepidermal blister. Electron microscopic study showed blister formation inside the basal cells at the level just above the attachment plaque of hemidesmosome. Immunofluorescence showed complete loss of staining to plectin. The mutation analysis using protein truncation test and DNA sequencing revealed a C-to-T transition at nucleotide position 7006 of the plectin cDNA sequence, which lead a novel homozygous nonsense mutation (R2319X). CONCLUSION: From the above results, the diagnosis of epidermolysis bullosa simplex associated with muscular dystrophy was made. Slight muscular dystrophy was noticed at the age of 25 years. The muscular dystrophy gradually progressed and she could not walk at the age of 46 years. However, she can still breathe and swallow by herself. This is the patient of this disease with the longest follow-up, and may indicate the slow progress of muscular condition of this disease. PMID- 15659327 TI - A new arginine substitution mutation of DSRAD gene in a Chinese family with dyschromatosis symmetrica hereditaria. AB - BACKGROUND: Dyschromatosis symmetrica hereditaria (DSH) is a pigmentary genodermatosis of autosomal dominant inheritance characterized by a mixture of hyperpigmented and hypopigmented macules distributed on the dorsal aspects of the hands and feet. To date, only three articles testified that DSH is caused by the mutations of DSRAD gene (also called ADAR1) encoding for RNA-specific adenosine deaminase. OBJECTIVE: To identify mutations of DSRAD as the disease-causing gene and recognize different mutations giving a clue to insight into the mechanism of DSH. METHODS: We collected a Chinese DSH family consisting of a total of 11 individuals including five DSH patients (three males and two females). The whole coding region of DSRAD was amplified by polymerase chain reaction and products analyzed by direct sequencing. RESULTS: We detected a transition, 3463 C>T, leading to a missense mutation (R1155W) in genomic DNAs of five patients, and the point mutation was not found in normal individuals in this DSH family and in 100 unrelated, population-match control individuals. CONCLUSION: Our data suggests that R1155W missense mutation is a new mutation in exon 15 of DSRAD gene and further testify that DSRAD gene is the pathogenic gene of DSH. PMID- 15659328 TI - Up-regulation of cell surface Toll-like receptor 4-MD2 expression on dendritic epidermal T cells after the emigration from epidermis during cutaneous inflammation. AB - BACKGROUND: Mouse epidermis contains a population of gammadelta T cells, termed dendritic epidermal T cells (DETCs), which uniformly express the invariant Vgamma3 T cell receptor. Certain DETC lines were reported to respond to Gram negative bacteria in the presence of immobilized anti-CD3 monoclonal antibody or to lipopolysaccharide (LPS) in the presence of B cell lines. OBJECTIVE: To determine whether DETCs express the primary signaling receptor for LPS, Toll-like receptor (TLR) 4-MD2. METHODS: We analyzed expression of TLR4-MD2 in three independent DETC lines as well as in freshly isolated DETCs. RESULTS: All DETC lines expressed TLR4 and MD2 transcripts and TLR4-MD2 protein complex intracellularly, but none expressed TLR4-MD2 on the cell surface. By immunoblotting, only the immature form of TLR4 protein was detected in the DETC lines. The DETC lines did not respond to LPS even in the presence of immobilized anti-CD3 monoclonal antibody. Freshly isolated DETCs and their fetal thymic precursors also lacked cell surface expression of TLR4-MD2, but a small subpopulation of dermal Vgamma3 T cells isolated from croton oil-painted skin expressed TLR4-MD2 on the cell surface. Similarly, Vgamma3 T cells emigrated from organ-cultured epidermis expressed cell surface TLR4-MD2. CONCLUSIONS: These results demonstrate that DETCs do not constitutively express cell surface TLR4 MD2, but TLR4-MD2 expression may be up-regulated when DETCs emigrate from epidermis during cutaneous inflammation. PMID- 15659329 TI - Influence of genetic polymorphisms on interleukin-10 mRNA expression and psoriasis susceptibility. PMID- 15659331 TI - Up-regulation of activities of mitogen-activated protein kinase in psoriatic lesions. PMID- 15659330 TI - The role of heat shock proteins (HSPs) in vitiligo: deviation of cytotoxic response? PMID- 15659334 TI - eIF2 and the control of cell physiology. AB - Eukaryotic initiation factor eIF2 and its 'exchange factor' eIF2B play a key role in the regulation of protein synthesis in eukaryotes from yeast to mammals. Phosphorylation of eIF2 inhibits eIF2B and thus translation initiation. Four eIF2 kinases are now known in mammalian cells and these are activated in response to specific stress conditions. While phosphorylation of eIF2 serves to impair general protein synthesis, it causes upregulation of the translation of certain specific mRNAs that encode transcription factors. It can, therefore, exert effects on gene expression at multiple levels. The importance of correct control of eIF2 and eIF2B for normal physiology is exemplified by data from transgenic mice carrying knock-in or knock-out mutations and by the fact that mutations in the genes for the eIF2 kinase PERK or for eIF2B give rise to serious human diseases. PMID- 15659332 TI - Messenger RNA levels of melanogenesis-associated genes in lentigo senilis lesions. PMID- 15659335 TI - Translational control in virus-infected cells: models for cellular stress responses. AB - Protein synthesis is regulated at the translational level by a variety of mechanisms in virus-infected cells. Viruses often induce the shut-off of host translation in order to favour the expression of their own genetic information, but cells possess a number of strategies for counteracting such effects of infection. Important regulatory mechanisms include the phosphorylation of the alpha subunit of polypeptide chain initiation factor eIF2, RNA degradation mediated by the 2'5'-oligoadenylate/RNase L system, control of availability of the cap-binding protein eIF4E by its interaction with the 4E-binding proteins and specific proteolytic cleavage of several key initiation factors. Most of these mechanisms are also utilised in uninfected cells in response to a variety of physiological stresses and during the early stages of apoptosis. Thus, mechanisms of translational control during virus infection can provide models for the cellular stress responses observed in a wide range of other circumstances. PMID- 15659336 TI - Role of amino acids in the translational control of protein synthesis in mammals. AB - Amino acids, long considered simply substrates for protein synthesis, have been recently shown to act as modulators of intracellular signal transduction pathways typically associated with growth-promoting hormones such as insulin and insulin like growth factor-1. Many of the endpoints of the signaling pathways regulated by amino acids are proteins involved in mRNA translation. Thus, particular amino acids not only serve as substrates for protein synthesis but are also modulators of the process. The focus of this article is to review recent studies that have used intact animals as experimental models to examine the role of amino acids as modulators of signal transduction pathways. PMID- 15659337 TI - mTOR, translational control and human disease. AB - Many human diseases occur when the precise regulation of cell growth (cell mass/size) and proliferation (rates of cell division) is compromised. This review highlights those human disorders that occur as a result of inappropriate cellular signal transduction through the mammalian target of rapamycin (mTOR), a major pathway that coordinates proper cell growth and proliferation by regulating ribosomal biogenesis and protein translation. Recent studies reveal that the tuberous sclerosis complex (TSC)-1/2, PTEN, and LKB1 tumor suppressor proteins tightly control mTOR. Loss of these tumor suppressors leads to an array of hamartoma syndromes as a result of heightened mTOR signaling. Since mTOR plays a pivotal role in maintaining proper cell size and growth, dysregulation of mTOR signaling results in these benign tumor syndromes and an array of other human disorders. PMID- 15659338 TI - The implications of structured 5' untranslated regions on translation and disease. AB - Translational control is a key step in eukaryotic gene expression. The majority of translational control occurs at the level of initiation, thus implicating the 5' untranslated region as a major site of translational regulation. Many growth related mRNAs have atypical 5' UTRs, which are often long and GC-rich. Such features promote formation of stable secondary structure, and many mRNAs encoding proteins involved in cell growth, proliferation and apoptosis have structured 5' UTRs, which in many cases harbour internal ribosome entry sites (IRESs) and upstream open-reading frames (uORFs). In this review we discuss how secondary structural elements in the 5' UTR can regulate translation and how mutations that perturb these secondary structural elements can have implications for disease and tumourigenesis. PMID- 15659339 TI - Mechanisms of translational control by the 3' UTR in development and differentiation. AB - Translational control plays a major role in early development, differentiation and the cell cycle. In this review, we focus on the four main mechanisms of translational control by 3' untranslated regions: 1. Cytoplasmic polyadenylation and deadenylation; 2. Recruitment of 4E binding proteins; 3. Regulation of ribosomal subunit binding; 4. Post-initiation repression by microRNAs. Proteins with conserved functions in translational control during development include cytoplasmic polyadenylation element binding proteins (CPEB/Orb), Pumilio, Bruno, Fragile X mental retardation protein and RNA helicases. The translational regulation of the mRNAs encoding cyclin B1, Oskar, Nanos, Male specific lethal 2 (Msl-2), lipoxygenase and Lin-14 is discussed. PMID- 15659340 TI - The role of the AU-rich elements of mRNAs in controlling translation. AB - Adenosine- and uridine-rich elements (AREs) located in 3'-untranslated regions are the best-known determinants of RNA instability. These elements have also been shown to control translation in certain mRNAs, including mRNAs for prominent pro inflammatory and tumor growth-related proteins, and physiological anti inflammatory processes that target ARE-controlled translation of mRNAs coding for pro-inflammatory proteins have been described. A major research effort is now being made to understand the mechanisms by which the translation of these mRNAs is controlled and the signalling pathways involved. This review focuses on the role of ARE-containing gene translation in inflammation, and the disease models that have improved our understanding of ARE-mediated translational control. PMID- 15659342 TI - Regulation of GATA gene expression during vertebrate development. AB - GATA factors regulate critical events in hematopoietic lineages (GATA-1/2/3), the heart and gut (GATA-4/5/6) and various other tissues. Transgenic approaches have revealed that GATA genes are regulated in a modular fashion by sets of enhancers that govern distinct temporal and/or spatial facets of the overall expression patterns. Efforts are underway to resolve how these GATA gene enhancers are themselves regulated in order to elucidate the genetic and molecular hierarchies that govern GATA expression in particular developmental contexts. These enhancers also afford a raft of tools that can be used to selectively perturb and probe various developmental events in transgenic animals. PMID- 15659343 TI - The roles of GATA-4, -5 and -6 in vertebrate heart development. AB - The transcription factors GATA-4, -5 and -6 are expressed very early in heart tissue. Essential GATA sites have been detected in several cardiac genes and the cardiac GATA factors interact with a wide variety of cofactors which synergistically increase gene expression. These multi-protein transcriptional complexes confer promoter-specificity on the GATA factors and also on the more broadly expressed cofactors. Here we summarise the data on these interactions and represent the conclusions as a GATA factor-based genetic regulatory network for the heart. Of the three cardiac GATAs, GATA-4 is by far the most extensively studied, however, loss-of-function data question its presumed dominance during heart development as opposed to hypertrophy. PMID- 15659344 TI - Using the zebrafish model to study GATA transcription factors. AB - The zebrafish is an established animal model system that profits from the availability of strong experimental approaches in both genetics and embryology. As a vertebrate, zebrafish can be used to model many aspects of human development and disease. GATA transcription factors play important roles in the development of many organ systems, including those for hematopoietic, cardiovascular, reproductive, and gut-endoderm derived tissues. The six vertebrate GATA factors are highly conserved in zebrafish at the level of sequence, expression pattern, and function. The identification of mutants, establishment of transgenic GFP reporter fish, and the ease of performing loss- and gain-of-function experiments have all contributed new insight into our understanding of the regulation and function of GATA factors. We review recent advances toward this goal using the zebrafish system with a focus on hematopoiesis and cardiogenesis, and suggest how comparative genetics using the zebrafish genes might reveal core conserved properties, as well as changes in gene function that reflect different morphogenetic programs utilized by various vertebrate embryos. PMID- 15659345 TI - GATA factors in Drosophila heart and blood cell development. AB - GATA transcription factors comprise an evolutionarily conserved family of proteins that function in the specification and differentiation of various cell types during animal development. In this review, we examine current knowledge of the structure, expression, and function of the Pannier and Serpent GATA factors as they relate to cardiogenesis and hematopoiesis in the Drosophila system. We also assess the molecular and genetic characteristics of the Friend of GATA protein U-shaped, which serves as a regulator of Pannier and Serpent function in these two developmental processes. PMID- 15659346 TI - Coregulation of GATA factors by the Friend of GATA (FOG) family of multitype zinc finger proteins. AB - The Friend of GATA (FOG) family of proteins is an evolutionarily conserved class of large multitype zinc finger cofactors that bind to the amino zinc finger of GATA transcription factors and modulate their activity. Two FOG genes have been identified in mammals, both of which interact with each of the six known vertebrate GATA factors in vitro. Physical interaction between FOG and GATA proteins in vivo is essential for the development of a broad array of tissues, reflecting the overlapping expression patterns of these factors. In this review, we will discuss the identification and characterization of FOG proteins, their role in human disease, and recent studies that shed new light on their function and regulation. PMID- 15659347 TI - Gene expression regulation and domain function of hematopoietic GATA factors. AB - The hierarchical gene regulatory network in hematopoiesis is highly complex, making elucidation of the processes of specification and differentiation of hematopoietic cells a challenging task. Recent discoveries have divulged the GATA factors as central to the genetic control of hematopoiesis. In particular, hematopoietic development is subject to extensive and precise regulation of GATA 1 and GATA-2 at the molecular level. We wish to emphasize the regulatory relationships between GATA-1 and GATA-2 implicated in cell development. An advanced experimental genetic approach has provided evidence that abnormalities in this network may result in a variety of blood disorders. The most striking new finding is the novel pathogenesis arising from GATA-1 dysfunction that leads to leukemia. PMID- 15659348 TI - GATA1 in normal and malignant hematopoiesis. AB - In the late 1980s, several research groups independently discovered the founding member of the GATA family of transcription factors, GATA-1. Each group had evidence that GATA-1 played an important role in erythroid gene expression, but little did they know that it would turn out to be a key regulator of development of not only red blood cells, but of several other hematopoietic cell types as well. Furthermore, few would have guessed that missense mutations in GATA1 would cause inherited blood disorders, while acquired mutations would be found associated with essentially all cases of acute megakaryoblastic leukemia (AMKL) in children with Down syndrome (DS). With respect to the latter disorder, the presence of a GATA1 mutation is now arguably the defining feature of this leukemia. In this review, I will summarize our current knowledge of the role of GATA-1 in normal development, and discuss how mutations in GATA1 lead to abnormal and malignant hematopoiesis. PMID- 15659349 TI - Conceptual and methodological issues in testing the circumplex structure of data in personality and social psychology. AB - Circumplex representations of data have enjoyed widespread popularity in personality and social psychological research. In this article we review the conceptual assumptions implied by circumplex representations and we discuss the limitations of traditional statistical methods for testing these assumptions. A relatively nontechnical overview of a new covariance structure modeling approach to testing circumplex structure is provided. The use of this approach is illustrated with two published data sets. The advantages and disadvantages of this approach relative to more traditional statistical approaches are discussed. The conclusion is that the covariance structure modeling approach has significant advantages in that it provides a closer conceptual match to the theoretical assumptions of circumplex representations, supplies information more directly relevant to circumplex representations and permits more precise and flexible testing of hypotheses derived from circumplex representations. PMID- 15659350 TI - Accurate social perception at zero acquaintance: the affordances of a Gibsonian approach. AB - We review research on accurate social perception at zero acquaintance and apply a Gibsonian ecological approach to redress several shortcomings. We argue that recent use of Brunswik's lens model to determine what physical qualities accurately communicate psychological traits has limited utility because it fails to consider the structured information provided by configural physical qualities that is central to Gibson's (1979) theory. We elaborate a developmental model of relationships between physical and psychological qualities that highlights research needed to identify configural physical qualities that may inform accurate perceptions. This model and tenets of the ecological theory yield several hypotheses regarding such qualities. Finally, we advocate the value of studying perceived affordances (opportunities for acting, interacting, or being acted upon) because this will focus attention on the neglected issue of contextual influences on social perception accuracy, and because affordances may be perceived more accurately than global personality traits. PMID- 15659351 TI - Minding the close relationship. AB - In this theoretical analysis, we argue that a process referred to as minding is essential for a couple to feel mutually close and satisfied in a close relationship over a long period Minding represents a package of mutual self disclosure, other forms of goal-oriented behavior aimed at facilitating the relationship, and attributions about self's and other's motivations, intentions, and Mort in the relationship. Self-disclosure and attribution activities in minding are aimed at getting to know the other, trying to understand the other's motivations and deeper disposition as they pertain to the relationship, and showing respect and acceptance for knowledge gained about other. We link the concept of minding to other major ideas and literatures about how couples achieve closeness: self-disclosure and social penetration, intimacy, empathy and empathic accuracy, and love and self-expansion. We argue that the minding process articulated here has not previously been delineated and that it is a useful composite notion about essential steps in bonding among humans. We also argue that the minding concept stretches our understanding of the interface of attribution and close relationships. We present research possibilities and implications and consider possible alternative positions and counter arguments about the merits of the minding idea for close relationship satisfaction. PMID- 15659352 TI - The "golden section" and bias in perceptions of social consensus. AB - Meta-analytic examination of I28 false consensus effect issues supports the hypothesis that the "Golden Section" (61.8% group size) approximates the level of actual consensus that separates overestimation of consensus (group size < 61.8%) from underestimation (group size > 61.8%). Overestimation of the actual percentage of others who endorse one's own view increases as actual consensus decreases from 61.8%, and underestimation increases as it exceeds 61.8%. The form of the response (viz, a yes or no answer to a question) moderates this conclusion. The Golden Section holds for majorities and minorities defined by agreement with an issue. For majority and minority groups defined by disagreement, the inflection point is higher. Contrary to Mullen and Hu (1988), for agreeing majorities, the slope for consensus underestimation as a function of increased majority size does not differ from that of minority overestimation. PMID- 15659354 TI - A selenocysteine tRNA and SECIS element in Plasmodium falciparum. AB - The molecular machinery for incorporating selenocysteine into proteins is present in both prokaryotes and eukaryotes. Although selenocysteine insertion has been reported in animals, plants, and protozoans, known eukaryotic selenocysteine tRNA sequences and selenocysteine insertion sequences are limited to animals and plants. Here we present clear indications of the presence of selenocysteine-tRNA and a selenocysteine insertion sequence in Plasmodium falciparum. To our knowledge, this is the first report of an identification of protozoan selenocysteine insertion machinery at the sequence level. PMID- 15659355 TI - Bicistronic and fused monocistronic transcripts are derived from adjacent loci in the Arabidopsis genome. AB - Comparisons of full-length cDNAs and genomic DNAs available for Arabidopsis thaliana described here indicate that some adjacent loci are transcribed into extremely long RNAs spanning two annotated genes. Once expressed, some of these transcripts are post-transcriptionally spliced within their coding and intergenic sequences to generate bicistronic transcripts containing two complete open reading frames. Others are spliced to generate monocistronic transcripts coding for fusion proteins with sequences derived from both loci. RT-PCR of several P450 transcripts in this collection indicates that these extended transcripts exist side by side with shorter monocistronic transcripts derived from the individual loci in each pair. The existence of these unusual transcripts highlights variations in the processes of transcription and splicing that could not possibly have been predicted in the algorithms used for genome annotation and splice site predictions. PMID- 15659357 TI - NMR structure of the apoB mRNA stem-loop and its interaction with the C to U editing APOBEC1 complementary factor. AB - We have solved the NMR structure of the 31-nucleotide (nt) apoB mRNA stem-loop, a substrate of the cytidine deaminase APOBEC1. We found that the edited base located at the 5' end of the octa-loop is stacked between two adenosines in both the unedited (cytidine 6666) and the edited (uridine 6666) forms and that the rest of the loop is unstructured. The 11-nt "mooring" sequence essential for editing is partially flexible although it is mostly in the stem of the RNA. The octa-loop and the internal loop in the middle of the stem confer this flexibility. These findings shed light on why APOBEC1 alone cannot edit efficiently the cytidine 6666 under physiological conditions, the editing base being buried in the loop and not directly accessible. We also show that APOBEC1 does not specifically bind apoB mRNA and requires the auxiliary factor, APOBEC1 complementary factor (ACF), to edit specifically cytidine 6666. The binding of ACF to both the mooring sequence and APOBEC1 explains the specificity of the reaction. Our NMR study lead us to propose a mechanism in which ACF recognizes first the flexible nucleotides of the mooring sequence (the internal loop and the 3' end octa-loop) and subsequently melts the stem-loop, exposing the amino group of the cytidine 6666 to APOBEC1. Thus, the flexibility of the mooring sequence plays a central role in the RNA recognition by ACF. PMID- 15659356 TI - Linkage between proton binding and folding in RNA: a thermodynamic framework and its experimental application for investigating pKa shifting. AB - Perturbation of pKa values can change the favored protonation states of the nucleobases at biological pH and thereby modulate the function of RNA and DNA molecules. In an effort to understand the driving forces for pKa shifting specific to nucleic acids, we developed a thermodynamic framework that relates proton binding to the nucleobases and the helix-coil transition. Key features that emerge from the treatment are a comprehensive description of all the actions of proton binding on RNA folding: acid and alkaline denaturation of the helix and pKa shifting in the folded state. Practical experimental approaches for measuring pKas from thermal denaturation experiments are developed. Microscopic pka values (where ka is the acid dissociation constant) for the unfolded state were determined directly by experiments on unstructured oligonucleotides, which led to a macroscopic pKa for the ensemble of unfolded states shifted toward neutrality. The formalism was then applied to pH-dependent UV melting data for model DNA oligonucleotides. Folded-state pka) values were in good agreement with the outcome of pH titrations, and the acid and alkaline denaturation regions were well described. The formalism developed here is similar to that of Draper and coworkers for Mg2+ binding to RNA, except that the unfolded state is described explicitly owing to the presence of specific proton-binding sites on the bases. A principal conclusion is that it should be possible to attain large pKa shifts by designing RNA molecules that fold cooperatively. PMID- 15659358 TI - Characterization of a native hammerhead ribozyme derived from schistosomes. AB - A recent re-examination of the role of the helices surrounding the conserved core of the hammerhead ribozyme has identified putative loop-loop interactions between stems I and II in native hammerhead sequences. These extended hammerhead sequences are more active at low concentrations of divalent cations than are minimal hammerheads. The loop-loop interactions are proposed to stabilize a more active conformation of the conserved core. Here, a kinetic and thermodynamic characterization of an extended hammerhead sequence derived from Schistosoma mansoni is performed. Biphasic kinetics are observed, suggesting the presence of at least two conformers, one cleaving with a fast rate and the other with a slow rate. Replacing loop II with a poly(U) sequence designed to eliminate the interaction between the two loops results in greatly diminished activity, suggesting that the loop-loop interactions do aid in forming a more active conformation. Previous studies with minimal hammerheads have shown deleterious effects of Rp-phosphorothioate substitutions at the cleavage site and 5' to A9, both of which could be rescued with Cd2+. Here, phosphorothioate modifications at the cleavage site and 5' to A9 were made in the schistosome-derived sequence. In Mg2+, both phosphorothioate substitutions decreased the overall fraction cleaved without significantly affecting the observed rate of cleavage. The addition of Cd2+ rescued cleavage in both cases, suggesting that these are still putative metal binding sites in this native sequence. PMID- 15659359 TI - The snRNP 15.5K protein folds its cognate K-turn RNA: a combined theoretical and biochemical study. AB - The human 15.5K protein binds to the 5' stem-loop of U4 snRNA, promotes the assembly of the spliceosomal U4/U6 snRNP, and is required for the recruitment of the 61K protein and the 20/60/90K protein complex to the U4 snRNA. In the crystallographic structure of the 15.5K-U4 snRNA complex, the conformation of the RNA corresponds to the family of kink-turn (K-turn) structural motifs. We simulated the complex and the free RNA, showing how the protein binding and the intrinsic flexibility contribute to the RNA folding process. We found that the RNA is significantly more flexible in the absence of the 15.5K protein. Conformational transitions such as the interconversion between alternative purine stacking schemes, the loss of G-A base pairs, and the opening of the K-turn occur only in the free RNA. Furthermore, the stability of one canonical G-C base pair is influenced both by the binding of the 15.5K protein and the nature of the adjacent structural element in the RNA. We performed chemical RNA modification experiments and observed that the free RNA lacks secondary structure elements, a result in excellent agreement with the simulations. Based on these observations, we propose a protein-assisted RNA folding mechanism in which the RNA intrinsic flexibility functions as a catalyst. PMID- 15659360 TI - Number, position, and significance of the pseudouridines in the large subunit ribosomal RNA of Haloarcula marismortui and Deinococcus radiodurans. AB - The number and position of the pseudouridines of Haloarcula marismortui and Deinococcus radiodurans large subunit RNA have been determined by a combination of total nucleoside analysis by HPLC-mass spectrometry and pseudouridine sequencing by the reverse transcriptase method and by LC/MS/MS. Three pseudouridines were found in H. marismortui, located at positions 1956, 1958, and 2621 corresponding to Escherichia coli positions 1915, 1917, and 2586, respectively. The three pseudouridines are all in locations found in other organisms. Previous reports of a larger number of pseudouridines in this organism were incorrect. Three pseudouridines and one 3-methyl pseudouridine (m3Psi) were found in D. radiodurans 23S RNA at positions 1894, 1898 (m3Psi), 1900, and 2584, the m3Psi site being determined by a novel application of mass spectrometry. These positions correspond to E. coli positions 1911, 1915, 1917, and 2605, which are also pseudouridines in E. coli (1915 is m3Psi). The pseudouridines in the helix 69 loop, residues 1911, 1915, and 1917, are in positions highly conserved among all phyla. Pseudouridine 2584 in D. radiodurans is conserved in eubacteria and a chloroplast but is not found in archaea or eukaryotes, whereas pseudouridine 2621 in H. marismortui is more conserved in eukaryotes and is not found in eubacteria. All the pseudoridines are near, but not exactly at, nucleotides directly involved in various aspects of ribosome function. In addition, two D. radiodurans Psi synthases responsible for the four Psi were identified. PMID- 15659362 TI - Assessing predictions of protein-protein interaction: the CAPRI experiment. AB - The Critical Assessment of PRedicted Interactions (CAPRI) experiment was designed in 2000 to test protein docking algorithms in blind predictions of the structure of protein-protein complexes. In four years, 17 complexes offered by crystallographers as targets prior to publication, have been subjected to structure prediction by docking their two components. Models of these complexes were submitted by predictor groups and assessed by comparing their geometry to the X-ray structure and by evaluating the quality of the prediction of the regions of interaction and of the pair wise residue contacts. Prediction was successful on 12 of the 17 targets, most of the failures being due to large conformation changes that the algorithms could not cope with. Progress in the prediction quality observed in four years indicates that the experiment is a powerful incentive to develop new procedures that allow for flexibility during docking and incorporate nonstructural information. We therefore call upon structural biologists who study protein-protein complexes to provide targets for further rounds of CAPRI predictions. PMID- 15659363 TI - Analysis of allosteric signal transduction mechanisms in an engineered fluorescent maltose biosensor. AB - We previously reported the construction of a family of reagentless fluorescent biosensor proteins by the structure-based design of conjugation sites for a single, environmentally sensitive small molecule dye, thus providing a mechanism for the transduction of ligand-induced conformational changes into a macroscopic fluorescence observable. Here we investigate the microscopic mechanisms that may be responsible for the macroscopic fluorescent changes in such Fluorescent Allosteric Signal Transduction (FAST) proteins. As case studies, we selected three individual cysteine mutations (F92C, D95C, and S233C) of Escherichia coli maltose binding protein (MBP) covalently labeled with a single small molecule fluorescent probe, N-((2-iodoacetoxy)ethyl)-N-methyl)amino-7-nitrobenz-2-oxa-1,3 diazole (NBD), each giving rise to a robust FAST protein with a distinct maltose dependent fluorescence response. The fluorescence emission intensity, anisotropy, lifetime, and iodide-dependent fluorescence quenching were determined for each conjugate in the presence and absence of maltose. Structure-derived solvent accessible surface areas of the three FAST proteins are consistent with experimentally observed quenching data. The D95C protein exhibits the largest fluorescence change upon maltose binding. This mutant was selected for further characterization, and residues surrounding the fluorophore coupling site were mutagenized. Analysis of the resulting mutant FAST proteins suggests that specific hydrogen-bonding interactions between the fluorophore molecule and two tyrosine side-chains, Tyr171 and Tyr176, in the open state but not the closed, are responsible for the dramatic fluorescence response of this construct. Taken together these results provide insights that can be used in future design cycles to construct fluorescent biosensors that optimize signaling by engineering specific hydrogen bonds between a fluorophore and protein. PMID- 15659364 TI - Improving the alkalophilic performances of the Xyl1 xylanase from Streptomyces sp. S38: structural comparison and mutational analysis. AB - Endo-beta-1,4-xylanases of the family 11 glycosyl-hydrolases are catalytically active over a wide range of pH. Xyl1 from Streptomyces sp. S38 belongs to this family, and its optimum pH for enzymatic activity is 6. Xyn11 from Bacillus agaradhaerens and XylJ from Bacillus sp. 41M-1 share 85% sequence identity and have been described as highly alkalophilic enzymes. In an attempt to better understand the alkalophilic adaptation of xylanases, the three-dimensional structures of Xyn11 and Xyl1 were compared. This comparison highlighted an increased number of salt-bridges and the presence of more charged residues in the catalytic cleft as well as an eight-residue-longer loop in the alkalophilic xylanase Xyn11. Some of these charges were introduced in the structure of Xyl1 by site-directed mutagenesis with substitutions Y16D, S18E, G50R, N92D, A135Q, E139K, and Y186E. Furthermore, the eight additional loop residues of Xyn11 were introduced in the homologous loop of Xyl1. In addition, the coding sequence of the XylJ catalytic domain was synthesized by recursive PCR, expressed in a Streptomyces host, purified, and characterized together with the Xyl1 mutants. The Y186E substitution inactivated Xyl1, but the activity was restored when this mutation was combined with the G50R or S18E substitutions. Interestingly, the E139K mutation raised the optimum pH of Xyl1 from 6 to 7.5 but had no effect when combined with the N92D substitution. Modeling studies identified the possible formation of an interaction between the introduced lysine and the substrate, which could be eliminated by the formation of a putative salt-bridge in the N92D/E139K mutant. PMID- 15659365 TI - Stability of mutant serpin/furin complexes: dependence on pH and regulation at the deacylation step. AB - Furin proteolytically cleaves a wide variety of proprotein substrates mainly within the trans-Golgi network (TGN) but also at the cell membrane and in endosomal compartments where pH is more acidic. Incorporation of furin recognition sequences within the reactive site loop (RSL) of alpha(1)-antitrypsin (AT) leads to the production of furin inhibitors. In an attempt to design more stable, potent, and specific serpin-based inhibitors, we constructed a series of AT and alpha(1)-antichymotrypsin (ACT) mutants by modifying the P(7)-P(1) region of their RSLs. The biochemical properties of these variants were assessed by evaluating their propensity to establish SDS-resistant complexes with furin in a variety of conditions (pH 6.0-9.0) and by measuring their association rate constants. The effect of pH during the initial steps of complex formation was minimal, suggesting that the acylation step is not rate-limiting. The decrease in stoichiometry of inhibition (SI) values observed in AT variants at high pHs was a result of the reduced pH-dependent deacylation rate, which is rate-limiting in this mechanism and which suggests increased complex stability. Conversely, the SI values for ACT mutants had a tendency to be lower at acidic pH. Transiently transfecting HEK293 cells with these mutants abolished processing of the pro-von Willebrand factor precursor but, interestingly, only the ACT variants were secreted in the media as uncleaved forms. Our results suggest that reengineering the reactive site loops of serpins to accommodate and target furin or other serine proteases must take into account the intrinsic physicochemical properties of the serpin. PMID- 15659366 TI - Physicochemical and residue conservation calculations to improve the ranking of protein-protein docking solutions. AB - Many protein-protein docking algorithms generate numerous possible complex structures with only a few of them resembling the native structure. The major challenge is choosing the near-native structures from the generated set. Recently it has been observed that the density of conserved residue positions is higher at the interface regions of interacting protein surfaces, except for antibody antigen complexes, where a very low number of conserved positions is observed at the interface regions. In the present study we have used this observation to identify putative interacting regions on the surface of interacting partners. We studied 59 protein complexes, used previously as a benchmark data set for docking investigations. We computed conservation indices of residue positions on the surfaces of interacting proteins using available homologous sequences and used this information to filter out from 56% to 86% of generated docked models, retaining near-native structures for further evaluation. We used a reverse filter of conservation score to filter out the majority of nonnative antigen-antibody complex structures. For each docked model in the filtered subsets, we relaxed the conformation of the side chains by minimizing the energy with CHARMM, and then calculated the binding free energy using a generalized Born method and solvent accessible surface area calculations. Using the free energy along with conservation information and other descriptors used in the literature for ranking docking solutions, such as shape complementarity and pair potentials, we developed a global ranking procedure that significantly improves the docking results by giving top ranks to near-native complex structures. PMID- 15659368 TI - An engineered chaperonin caging a guest protein: Structural insights and potential as a protein expression tool. AB - The structure of a chaperonin caging a substrate protein is not quite clear. We made engineered group II chaperonins fused with a guest protein and analyzed their structural and functional features. Thermococcus sp. KS-1 chaperonin alpha subunit (TCP) which forms an eightfold symmetric double-ring structure was used. Expression plasmids were constructed which carried two or four TCP genes ligated head to tail in phase and a target protein gene at the 3' end of the linked TCP genes. Electron microscopy showed that the expressed gene products with the molecular sizes of ~120 kDa (di-TCP) and ~230 kDa (tetra-TCP) formed double-ring complexes similar to those of wild-type TCP. The tetra-TCP retained ATPase activity and its thermostability was significantly higher than that of the wild type. A 260-kDa fusion protein of tetra-TCP and green fluorescent protein (GFP, 27 kDa) was able to form the double-ring complexes with green fluorescence. Image analyses indicated that the GFP moiety of tetra-TCP/GFP fusion protein was accommodated in the central cavity, and tetra-TCP/GFP formed the closed-form similar to that crystallographically resolved in group II chaperonins. Furthermore, it was suggested that caging GFP expanded the cavity around the bottom. Using this tetra-TCP fusion strategy, two virus structural proteins (21 25 kDa) toxic to host cells or two antibody fragments (25-36 kDa) prone to aggregate were well expressed in the soluble fraction of Escherichia coli. These fusion products also assembled to double-ring complexes, suggesting encapsulation of the guest proteins. The antibody fragments liberated by site-specific protease digestion exhibited ligand-binding activities. PMID- 15659367 TI - Stability of HAMLET--a kinetically trapped alpha-lactalbumin oleic acid complex. AB - The stability toward thermal and urea denaturation was measured for HAMLET (human alpha-lactalbumin made lethal to tumor cells) and alpha-lactalbumin, using circular dichroism and fluorescence spectroscopy as well as differential scanning calorimetry. Under all conditions examined, HAMLET appears to have the same or lower stability than alpha-lactalbumin. The largest difference is seen for thermal denaturation of the calcium free (apo) forms, where the temperature at the transition midpoint is 15 degrees C lower for apo HAMLET than for apo alpha lactalbumin. The difference becomes progressively smaller as the calcium concentration increases. Denaturation of HAMLET was found to be irreversible. Samples of HAMLET that have been renatured after denaturation have lost the specific biological activity toward tumor cells. Three lines of evidence indicate that HAMLET is a kinetic trap: (1) It has lower stability than alpha-lactalbumin, although it is a complex of alpha-lactalbumin and oleic acid; (2) its denaturation is irreversible and HAMLET is lost after denaturation; (3) formation of HAMLET requires a specific conversion protocol. PMID- 15659369 TI - Spectral magnitude effects on the analyses of secondary structure from circular dichroism spectroscopic data. AB - The effects of spectral magnitude on the calculated secondary structures derived from circular dichroism (CD) spectra were examined for a number of the most commonly used algorithms and reference databases. Proteins with different secondary structures, ranging from mostly helical to mostly beta-sheet, but which were not components of existing reference databases, were used as test systems. These proteins had known crystal structures, so it was possible to ascertain the effects of magnitude on both the accuracy of determining the secondary structure and the goodness-of-fit of the calculated structures to the experimental data. It was found that most algorithms are highly sensitive to spectral magnitude, and that the goodness-of-fit parameter may be a useful tool in assessing the correct scaling of the data. This means that parameters that affect magnitude, including calibration of the instrument, the spectral cell pathlength, and the protein concentration, must be accurately determined to obtain correct secondary structural analyses of proteins from CD data using empirical methods. PMID- 15659370 TI - Helical structure determined by NMR of the HIV-1 (345-392)Gag sequence, surrounding p2: implications for particle assembly and RNA packaging. AB - Gag protein oligomerization, an essential step during virus assembly, results in budding of spherical virus particles. This process is critically dependent on the spacer p2, located between the capsid and the nucleocapsid proteins. P2 contributes also, in association with NCp7, to specific recognition of the HIV-1 packaging signal resulting in viral genome encapsidation. There is no structural information about the 20 last amino acids of the C-terminal part of capsid (CA[CTD]) and p2, in the molecular mechanism of Gag assembly. In this study the structure of a peptide encompassing the 14 residues of p2 with the upstream 21 residues and the downstream 13 residues was determined by (1)H NMR in 30% trifluoroethanol (TFE). The main structural motif is a well-defined amphipathic alpha-helix including p2, the seven last residues of the CA(CTD), and the two first residues of NCp7. Peptides containing the p2 domain have a strong tendency to aggregate in solution, as shown by gel filtration analyses in pure H(2)O. To take into account the aggregation phenomena, models of dimer and trimer formed through hydrophobic or hydrophilic interfaces were constructed by molecular dynamic simulations. Gel shift experiments demonstrate that the presence of at least p2 and the 13 first residues of NCp7 is required for RNA binding. A computer-generated model of the Gag polyprotein segment (282-434)Gag interacting with the packaging element SL3 is proposed, illustrating the importance of p2 and NCp7 in genomic encapsidation. PMID- 15659371 TI - The crystal structure of an eukaryotic iron superoxide dismutase suggests intersubunit cooperation during catalysis. AB - Superoxide dismutases (SODs) are a family of metalloenzymes that catalyze the dismutation of superoxide anion radicals into molecular oxygen and hydrogen peroxide. Iron superoxide dismutases (FeSODs) are only expressed in some prokaryotes and plants. A new and highly active FeSOD with an unusual subcellular localization has recently been isolated from the plant Vigna unguiculata (cowpea). This protein functions as a homodimer and, in contrast to the other members of the SOD family, is localized to the cytosol. The crystal structure of the recombinant enzyme has been solved and the model refined to 1.97 A resolution. The superoxide anion binding site is located in a cleft close to the dimer interface. The coordination geometry of the Fe site is a distorted trigonal bipyramidal arrangement, whose axial ligands are His43 and a solvent molecule, and whose in-plane ligands are His95, Asp195, and His199. A comparison of the structural features of cowpea FeSOD with those of homologous SODs reveals subtle differences in regard to the metal-protein interactions, and confirms the existence of two regions that may control the traffic of substrate and product: one located near the Fe binding site, and another in the dimer interface. The evolutionary conservation of reciprocal interactions of both monomers in neighboring active sites suggests possible subunit cooperation during catalysis. PMID- 15659372 TI - Crystal structure of the complex formed between a group I phospholipase A2 and a naturally occurring fatty acid at 2.7 A resolution. AB - This is the first evidence of a naturally bound fatty acid to a group I Phospholipase A(2) (PLA(2)) and also to a PLA(2) with Asp 49. The fatty acid identified as n-tridecanoic acid is observed at the substrate recognition site of PLA(2) hydrophobic channel. The complex was isolated from the venom of Bungarus caeruleus (Common Indian Krait). The primary sequence of the PLA(2) was determined using the cDNA method. Three-dimensional structure has been solved by the molecular replacement method and refined using the CNS package to a final R factor of 19.8% for the data in the resolution range from 20.0 to 2.7 A. The final refined model is comprised of 912 protein atoms, one sodium ion, one molecule of n-tridecanoic acid, and 60 water molecules. The sodium ion is located in the calcium-binding loop with a sevenfold coordination. A characteristic extra electron density was observed in the hydrophobic channel of the enzyme, into which a molecule of n-tridecanoic acid was clearly fitted. The MALDI-TOF measurements of the crystals had earlier indicated an increase in the molecular mass of PLA(2) by 212 Da over the native PLA(2). A major part of the ligand fits well in the binding pocket and interacts directly with His 48 and Asp 49. Although the overall structure of PLA(2) in the present complex is similar to the native structure reported earlier, it differs significantly in the folding of its calcium-binding loop. PMID- 15659373 TI - Folding and particle assembly are disrupted by single-point mutations near the autocatalytic cleavage site of Nudaurelia capensis omega virus capsid protein. AB - Protein subunits of several RNA viruses are known to undergo post-assembly, autocatalytic cleavage that is required for infectivity. Nudaurelia capensis omega virus (Nomega V) is one of the simplest viruses to undergo an autocatalytic cleavage, making it an excellent model to understand both assembly and the mechanism of autoproteolysis. Heterologous expression of the coat protein gene of Nomega V in a baculovirus system results in the spontaneous assembly of virus like particles (VLPs) that remain uncleaved when purified at neutral pH. After acidification to pH 5.0, the VLPs autocatalytically cleave at residue 570, providing an in vitro control of the cleavage. The crystal structure of Nomega V displays three residues near the scissile bond that were candidates for participation in the reaction. These were changed by site-directed mutagenesis to conservative and nonconservative residues and the products analyzed. Even conservative changes at the three residues dramatically reduced cleavage when the subunits assembled properly. Unexpectedly, we discovered that these residues are not only critical to the kinetics of Nomega V autoproteolysis, but are also necessary for proper folding of subunits and, ultimately, assembly of Nomega V VLPs. PMID- 15659374 TI - An avidin-like domain that does not bind biotin is adopted for oligomerization by the extracellular mosaic protein fibropellin. AB - The protein avidin found in egg white seems optimized for binding the small vitamin biotin as a stable homotetramer. Indeed, along with its streptavidin ortholog in the bacterium Streptomyces avidinii, this protein shows the strongest known noncovalent bond of a protein with a small ligand. A third known member of the avidin family, as similar to avidin as is streptavidin, is found at the C terminal ends of the multidomain fibropellin proteins found in sea urchin. The fibropellins form a layer known as the apical lamina that surrounds the sea urchin embryo throughout development. Based upon the structure of avidin, we deduced a structural model for the avidin-like domain of the fibropellins and found that computational modeling predicts a lack of biotin binding and the preservation of tetramerization. To test this prediction we expressed and purified the fibropellin avidin-like domain and found it indeed to be a homotetramer incapable of binding biotin. Several lines of evidence suggest that the avidin-like domain causes the entire fibropellin protein to tetramerize. We suggest that the presence of the avidin-like domain serves a structural (tetrameric form) rather than functional (biotin-binding) role and may therefore be a molecular instance of exaptation-the modification of an existing function toward a new function. Finally, based upon the oligomerization of the avidin-like domain, we propose a model for the overall structure of the apical lamina. PMID- 15659376 TI - Specific erythrocyte binding capacity and biological activity of Plasmodium falciparum erythrocyte binding ligand 1 (EBL-1)-derived peptides. AB - Erythrocyte binding ligand 1 (EBL-1) is a member of the ebl multigene family involved in Plasmodium falciparum invasion of erythrocytes. We found that five EBL-1 high-activity binding peptides (HABPs) bound specifically to erythrocytes: 29895 ((41)HKKKSGELNNNKSGILRSTY(60)), 29903 ((201)LYECGK-KIKEMKWICTDNQF(220)), 29923 ((601)CNAILGSYADIGDIVRGLDV(620)), 29924((621)WRDINTNKLSEK-FQKIFMGGY(640)), and 30018 ((2481)LEDIINLSKKKKKSINDTSFY(2500)). We also show that binding was saturable, not sialic acid-dependent, and that all peptides specifically bound to a 36-kDa protein on the erythrocyte membrane. The five HABPs inhibited in vitro merozoite invasion depending on the peptide concentration used, suggesting their possible role in the invasion process. PMID- 15659375 TI - Deamidation: Differentiation of aspartyl from isoaspartyl products in peptides by electron capture dissociation. AB - Deamidation of asparaginyl and isomerization of aspartyl residues in proteins proceed through a succinimide intermediate producing a mixture of aspartyl and isoaspartyl residues. Isoaspartic acid is an isomer of aspartic acid with the C(beta) incorporated into the backbone, thus increasing the length of the protein backbone by one methylene unit. This post-translation modification is suspected to contribute to the aging of proteins and to protein folding disorders such as Alzheimer's disease, so that differentiating the two isomers becomes important. This manuscript reports that distinguishing aspartyl from isoaspartyl residues in peptides has been accomplished by electron capture dissociation (ECD) using a Fourier transform mass spectrometer (FTMS). Model peptides with aspartyl residues and their isoaspartyl analogs were examined and unique peaks corresponding to c(n)*+58 and z(l-n)-57 fragment ions (n, position of Asp; l, total number of amino acids in the peptide) were found only in the spectra of the peptides with isoaspartyl residues. The proposed fragmentation mechanism involves cleavage of the C(alpha)-C(beta) backbone bond, therefore splitting the isoaspartyl residue between the two fragments. Also, a complementary feature observed specific to aspartyl residues was the neutral loss of the aspartic acid side chain from the charge reduced species. CAD spectra of the peptides from the same instrument demonstrated the improved method because previously published CAD methods rely on the comparison to the spectra of standards with aspartyl residues. The potential use of the top-down approach to detect and resolve products from the deamidation of asparaginyl and isomerization of aspartyl residues is discussed. PMID- 15659377 TI - Intramolecular disulphide bond arrangements in nonhomologous proteins. AB - The presence and location of intramolecular disulphide bonds are a key determinant of the structure and function of proteins. Intramolecular disulphide bonds in proteins have previously been analyzed under the assumption that there is no clear relationship between disulphide arrangement and disulphide concentration. To investigate this, a set of sequence nonhomologous protein chains containing one or more intramolecular disulphide bonds was extracted from the Protein Data Bank, and the arrangements of the bonds, Protein Data Bank header, and Structural Characterization of Proteins fold were analyzed as a function of intramolecular disulphide bond concentration. Two populations of intramolecular disulphide bond-containing proteins were identified, with a naturally occurring partition at 25 residues per bond. These populations were named intramolecular disulphide bond-rich and -poor. Benefits of partitioning were illustrated by three results: (1) rich chains most frequently contained three disulphides, explaining the plateaux in extant disulphide frequency distributions; (2) a positive relationship between median chain length and the number of disulphides, only seen when the data were partitioned; and (3) the most common bonding pattern for chains with three disulphide bonds was based on the most common for two, only when the data were partitioned. The two populations had different headers, folds, bond arrangements, and chain lengths. Associations between IDSB concentration, IDSB bonding pattern, loop sizes, SCOP fold, and PDB header were also found. From this, we found that intramolecular disulphide bond rich and -poor proteins follow different bonding rules, and must be considered separately to generate meaningful models of bond formation. PMID- 15659378 TI - Reversible self-association of ovalbumin at air-water interfaces and the consequences for the exerted surface pressure. AB - In this study the relation between the ability of protein self-association and the surface properties at air-water interfaces is investigated using a combination of spectroscopic techniques. Three forms of chicken egg ovalbumin were obtained with different self-associating behavior: native ovalbumin, heat treated ov-albumin-being a cluster of 12-16 predominantly noncovalently bound proteins, and succinylated ovalbumin, as a form with diminished aggregation properties due to increased electrostatic repulsion. While the bulk diffusion of aggregated protein is clearly slower compared to monomeric protein, the efficiency of transport to the interface is increased, just like the efficiency of sticking to rather than bouncing from the interface. On a timescale of hours, the aggregated protein dissociates and adopts a conformation comparable to that of native protein adsorbed to the interface. The exerted surface pressure is higher for aggregated material, most probably because the deformability of the particle is smaller. Aggregated protein has a lower ability to desorb from the interface upon compression of the surface layer, resulting in a steadily increasing surface pressure upon reducing the available area for the surface layer. This observation is opposite to what is observed for succinylated protein that may desorb more easily and thereby suppresses the buildup of a surface pressure. Generally, this work demonstrates that modulating the ability of proteins to self-associate offers a tool to control the rheological properties of interfaces. PMID- 15659379 TI - Identifying Plasmodium falciparum cytoadherence-linked asexual protein 3 (CLAG 3) sequences that specifically bind to C32 cells and erythrocytes. AB - Adhesion of mature asexual stage Plasmodium falciparum parasite-infected erythrocytes (iRBC) to the vascular endothelium is a critical event in the pathology of Plasmodium falciparum malaria. It has been suggested that the clag gene family is essential in cytoadherence to endothelial receptors. Primers used in PCR and RT-PCR assays allowed us to determine that the gene encoding CLAG 3 (GenBank accession no. NP_473155) is transcribed in the Plasmodium falciparum FCB2 strain. Western blot showed that antisera produced against polymerized synthetic peptides from this protein recognized a 142-kDa band in P. falciparum schizont lysate. Seventy-one 20-amino-acid-long nonoverlapping peptides, spanning the CLAG 3 (cytoadherence-linked asexual protein on chromosome 3) sequence were tested in C32 cell and erythrocyte binding assays. Twelve CLAG peptides specifically bound to C32 cells (which mainly express CD36) with high affinity, hereafter referred to as high-affinity binding peptides (HABPs). Five of them also bound to erythrocytes. HABP binding to C32 cells and erythrocytes was independent of peptide charge or peptide structure. Affinity constants were between 100 nM and 800 nM. Cross-linking and SDS-PAGE analysis allowed two erythrocyte binding proteins of around 26 kDa and 59 kDa to be identified, while proteins of around 53 kDa were identified as possible receptor sites for C-32 cells. The HABPs' role in Plasmodium falciparum invasion inhibition was determined. Such an approach analyzing various CLAG 3 regions may elucidate their functions and may help in the search for new antigens important for developing antimalarial vaccines. PMID- 15659380 TI - Mapping protein energy landscapes with amide hydrogen exchange and mass spectrometry: I. A generalized model for a two-state protein and comparison with experiment. AB - Protein amide hydrogen exchange (HDX) is a convoluted process, whose kinetics is determined by both dynamics of the protein and the intrinsic exchange rate of labile hydrogen atoms fully exposed to solvent. Both processes are influenced by a variety of intrinsic and extrinsic factors. A mathematical formalism initially developed to rationalize exchange kinetics of individual amide hydrogen atoms is now often used to interpret global exchange kinetics (e.g., as measured in HDX MS experiments). One particularly important advantage of HDX MS is direct visualization of various protein states by observing distinct protein ion populations with different levels of isotope labeling under conditions favoring correlated exchange (the so-called EX1 exchange mechanism). However, mildly denaturing conditions often lead to a situation where the overall HDX kinetics cannot be clearly classified as either EX1 or EX2. The goal of this work is to develop a framework for a generalized exchange model that takes into account multiple processes leading to amide hydrogen exchange, and does not require that the exchange proceed strictly via EX1 or EX2 kinetics. To achieve this goal, we use a probabilistic approach that assigns a transition probability and a residual protection to each equilibrium state of the protein. When applied to a small protein chymotrypsin inhibitor 2, the algorithm allows complex HDX patterns observed experimentally to be modeled with remarkably good fidelity. On the basis of the model we are now in a position to begin to extract quantitative dynamic information from convoluted exchange kinetics. PMID- 15659381 TI - Characterization of a conserved C-terminal motif (RSPRR) in ribosomal protein S6 kinase 1 required for its mammalian target of rapamycin-dependent regulation. AB - The mammalian target of rapamycin, mTOR, is a Ser/Thr kinase that promotes cell growth and proliferation by activating ribosomal protein S6 kinase 1 (S6K1). We previously identified a conserved TOR signaling (TOS) motif in the N terminus of S6K1 that is required for its mTOR-dependent activation. Furthermore, our data suggested that the TOS motif suppresses an inhibitory function associated with the C terminus of S6K1. Here, we have characterized the mTOR-regulated inhibitory region within the C terminus. We have identified a conserved C-terminal "RSPRR" sequence that is responsible for an mTOR-dependent suppression of S6K1 activation. Deletion or mutations within this RSPRR motif partially rescue the kinase activity of the S6K1 TOS motif mutant (S6K1-F5A), and this rescued activity is rapamycin resistant. Furthermore, we have shown that the RSPRR motif significantly suppresses S6K1 phosphorylation at two phosphorylation sites (Thr 389 and Thr-229) that are crucial for S6K1 activation. Importantly, introducing both the Thr-389 phosphomimetic and RSPRR motif mutations into the catalytically inactive S6K1 mutant S6K1-F5A completely rescues its activity and renders it fully rapamycin resistant. These data show that the N-terminal TOS motif suppresses an inhibitory function mediated by the C-terminal RSPRR motif. We propose that the RSPRR motif interacts with a negative regulator of S6K1 that is normally suppressed by mTOR. PMID- 15659382 TI - Epidermal growth factor-induced proliferation requires down-regulation of Pax6 in corneal epithelial cells. AB - Growth factors play important roles in regulating corneal epithelial cell proliferation/differentiation during wound healing. It is suggested that PAX6 involves corneal epithelium lineage-specific differentiation (Liu, J. J., Kao, W. W., and Wilson, S. E. (1999) Exp. Eye Res. 68, 295-301); however, the regulatory mechanism and function of Pax6 in growth factor-induced corneal epithelial responses is still unknown. In the present study, we found that the mitogenic effect of epidermal growth factor (EGF) in corneal epithelial cells required suppression of PAX6 activity through cellular mechanisms involving Erk-signaling pathway-mediated increase in CTCF expression. EGF-induced CCCTC binding factor (CTCF) activation subsequently inhibited Pax6 expression by interacting with a CTCF-specific region upstream of the pax6 P0 promoter. Suppression of EGF-induced Erk activation by specific inhibitor or by the dominant expression of a silent Erk mutant effectively abolished the effects of EGF stimulation on regulations of CTCF and pax6. Apparently, down-regulation of Pax6 expression induced by EGF is required for corneal epithelial proliferation, because overexpression of pax6 in these cells attenuated EGF-induced proliferation. In contrast, knockdown of mRNA expression with pax6- or CTCF-specific small interfering RNA in corneal epithelial cells significantly promoted or attenuated EGF-induced proliferation, respectively. Thus, our results revealed a new regulatory mechanism that involves cellular signaling events and pax6 transcription regulation in growth factor mediated proliferation. In corneal epithelial cells, this suggests that inhibition of pax6 expression is a prerequisite for EGF to elicit controls of cell growth and fate. PMID- 15659383 TI - Cytoskeleton-mediated death receptor and ligand concentration in lipid rafts forms apoptosis-promoting clusters in cancer chemotherapy. AB - While investigating the mechanism of action of the novel antitumor drug Aplidin, we have discovered a potent and novel cell-killing mechanism that involves the formation of Fas/CD95-driven scaffolds in membrane raft clusters housing death receptors and apoptosis-related molecules. Fas, tumor necrosis factor-receptor 1, and tumor necrosis factor-related apoptosis-inducing ligand receptor 2/death receptor 5 were clustered into lipid rafts in leukemic Jurkat cells following Aplidin treatment, the presence of Fas being essential for apoptosis. Preformed membrane-bound Fas ligand (FasL) as well as downstream signaling molecules, including Fas-associated death domain-containing protein, procaspase-8, procaspase-10, c-Jun amino-terminal kinase, and Bid, were also translocated into lipid rafts, connecting death receptor extrinsic and mitochondrial intrinsic apoptotic pathways. Blocking Fas/FasL interaction partially inhibited Aplidin induced apoptosis. Aplidin was rapidly incorporated into membrane rafts, and drug uptake was inhibited by lipid raft disruption. Actin-linking proteins ezrin, moesin, RhoA, and RhoGDI were conveyed into Fas-enriched rafts in drug-treated leukemic cells. Disruption of lipid rafts and interference with actin cytoskeleton prevented Fas clustering and apoptosis. Thus, Aplidin-induced apoptosis involves Fas activation in both a FasL-independent way and, following Fas/FasL interaction, an autocrine way through the concentration of Fas, membrane bound FasL, and signaling molecules in membrane rafts. These data indicate a major role of actin cytoskeleton in the formation of Fas caps and highlight the crucial role of the clusters of apoptotic signaling molecule-enriched rafts in apoptosis, acting as concentrators of death receptors and downstream signaling molecules and as the linchpin from which a potent death signal is launched. PMID- 15659384 TI - Prostaglandin E2 induces interleukin-8 gene transcription by activating C/EBP homologous protein in human T lymphocytes. AB - The effect of prostaglandin E(2) (PGE(2)) in regulating the synthesis of the pro inflammatory chemokine interleukin-8 (IL-8) in T lymphocytes is not yet defined, even though it may reduce or enhance IL-8 synthesis in other cell types. Here, we demonstrate that, in human T cells, PGE(2) induced IL-8 mRNA transcription through prostaglandin E(2) receptors 1- and 4-dependent signal transduction pathways leading to the activation of the transcription factor C/EBP homologous protein (CHOP), never before implicated in IL-8 transcription. Several kinases, including protein kinase C, Src family tyrosine kinases, phosphatidylinositol 3 kinase, and p38 MAPK, were involved in PGE(2)-induced CHOP activation and IL-8 production. The transactivation of the IL-8 promoter by CHOP was NF-kappaB independent. Our data suggest that PGE(2) acts as a potent pro-inflammatory mediator by inducing IL-8 gene transcription in activated T cells through different signal transduction pathways leading to CHOP activation. These findings show the complexity with which PGE(2) regulates IL-8 synthesis by inhibiting or enhancing its production depending on the cell types and environmental conditions. PMID- 15659386 TI - Mitochondrial malate-aspartate shuttle regulates mouse embryo nutrient consumption. AB - Pyruvate has been considered the sole substrate that can support development of the mouse zygote to the two-cell stage, with lactate able to support development from the two-cell stage. This study has determined for the first time that mitochondrial reducing equivalent shuttles regulate metabolism in the early embryo. Activity of the malate-aspartate shuttle was found to be essential for the metabolism of lactate in the two-cell embryo. Furthermore, the inability of the mouse zygote to use lactate as an energy source was a result of a lack of malate-aspartate shuttle activity. The mRNA for the four enzymes for shuttle activity were detected at all stages of development. It was determined that aspartate was a rate-limiting factor in the activity of the malate-aspartate shuttle in mouse zygotes probably due to the high K(m) of the cytoplasmic aspartate aminotransferase. Addition of high concentrations of exogenous aspartate to the culture medium enabled mouse zygotes to utilize lactate in the absence of pyruvate and develop normally to the blastocyst stage as well as produce normal viable offspring. This study determined that the malate-aspartate shuttle is a key regulator of embryo metabolism and therefore viability and is the first report that mouse zygotes can develop normally to term in the absence of pyruvate. PMID- 15659385 TI - Antiproliferative plant and synthetic polyphenolics are specific inhibitors of vertebrate inositol-1,4,5-trisphosphate 3-kinases and inositol polyphosphate multikinase. AB - Inositol-1,4,5-trisphosphate 3-kinases (IP3K) A, B, and C as well as inositol polyphosphate multikinase (IPMK) catalyze the first step in the formation of the higher phosphorylated inositols InsP5 and InsP6 by metabolizing Ins(1,4,5)P3 to Ins(1,3,4,5)P4. In order to clarify the special role of these InsP3 phosphorylating enzymes and of subsequent anabolic inositol phosphate reactions, a search was conducted for potent enzyme inhibitors starting with a fully active IP3K-A catalytic domain. Seven polyphenolic compounds could be identified as potent inhibitors with IC50 < 200 nM (IC50 given): ellagic acid (36 nM), gossypol (58 nM), (-)-epicatechin-3-gallate (94 nM), (-)-epigallocatechin-3-gallate (EGCG, 120 nM), aurintricarboxylic acid (ATA, 150 nM), hypericin (170 nM), and quercetin (180 nM). All inhibitors displayed a mixed-type inhibition with respect to ATP and a non-competitive inhibition with respect to Ins(1,4,5)P3. Examination of these inhibitors toward IP3K-A, -B, and -C and IPMK from mammals revealed that ATA potently inhibits all kinases while the other inhibitors do not markedly affect IPMK but differentially inhibit IP3K isoforms. We identified chlorogenic acid as a specific IPMK inhibitor whereas the flavonoids myricetin, 3',4',7,8 tetrahydroxyflavone and EGCG inhibit preferentially IP3K-A and IP3K-C. Mutagenesis studies revealed that both the calmodulin binding and the ATP [corrected] binding domain in IP3K are involved in inhibitor binding. Their absence in IPMK and the presence of a unique insertion in IPMK were found to be important for selectivity differences from IP3K. The fact that all identified IP3K and IPMK inhibitors have been reported as antiproliferative agents and that IP3Ks or IPMK often are the best binding targets deserves further investigation concerning their antitumor potential. PMID- 15659387 TI - Oxidative modifications and aggregation of Cu,Zn-superoxide dismutase associated with Alzheimer and Parkinson diseases. AB - Although oxidative stress has been strongly implicated in the pathogenesis of Alzheimer disease (AD) and Parkinson disease (PD), the identities of specific protein targets of oxidative damage remain largely unknown. Here, we report that Cu,Zn-superoxide dismutase (SOD1), a key antioxidant enzyme whose mutations have been linked to autosomal dominant neurodegenerative disorder familial amyotrophic lateral sclerosis (ALS), is a major target of oxidative damage in AD and PD brains. By using a combination of two-dimensional gel electrophoresis, immunoblot analysis, and mass spectrometry, we have identified four human brain SOD1 isoforms with pI values of 6.3, 6.0, 5.7, and 5.0, respectively. Of these, the SOD1 pI 6.0 isoform is oxidatively modified by carbonylation, and the pI 5.0 isoform is selectively accumulated in AD and PD. Moreover, Cys-146, a cysteine residue of SOD1 that is mutated in familial ALS, is oxidized to cysteic acid in AD and PD brains. Quantitative Western blot analyses demonstrate that the total level of SOD1 isoforms is significantly increased in both AD and PD. Furthermore, immunohistochemical and double fluorescence labeling studies reveal that SOD1 forms proteinaceous aggregates that are associated with amyloid senile plaques and neurofibrillary tangles in AD brains. These findings implicate, for the first time, the involvement of oxidative damage to SOD1 in the pathogenesis of sporadic AD and PD. This work suggests that AD, PD, and ALS may share a common or overlapping pathogenic mechanism(s) that could potentially be targeted by similar therapeutic strategies. PMID- 15659388 TI - Erbin regulates mitogen-activated protein (MAP) kinase activation and MAP kinase dependent interactions between Merlin and adherens junction protein complexes in Schwann cells. AB - Biallelic mutations in the neurofibromatosis 2 (NF2) gene are linked to schwannoma and meningioma tumorigenesis. Cells with NF2 mutations exhibit elevated levels of phosphorylated extracellular signal-regulated kinase (ERK) and aberrant cell-cell and cell-matrix contacts. The NF2 gene product, merlin, associates with adherens junction protein complexes, suggesting that part of its function as a tumor suppressor involves regulating cell junctions. Here, we find that a novel PDZ protein, called erbin, binds directly to the merlin-binding partner, EBP0, and regulates adherens junction dissociation through a MAP kinase dependent mechanism. Reducing erbin expression using a targeted siRNA in primary cultures of Schwann cells results in altered cell-cell interactions, disruption of E-cadherin adherens junctions, increased cell proliferation, and elevated levels of phosphorylated ERK, all phenotypes observed in cells that lack merlin. Reduction of erbin expression also results in the dissociation of merlin from adherens junction proteins and an increase in the levels of phosphorylated merlin. These phenotypes can be rescued if cells with reduced levels of erbin are treated with a pharmacological inhibitor of ERK kinase. Collectively, these data indicate that erbin regulates MAP kinase activation in Schwann cells and suggest that erbin links merlin to both adherens junction protein complexes and the MAP kinase signaling pathway. PMID- 15659389 TI - Site-directed mutagenesis, proteolytic cleavage, and activation of human proheparanase. AB - Heparanase is an endo-beta-D-glucuronidase that degrades heparan sulfate in the extracellular matrix and cell surfaces. Human proheparanase is produced as a latent 65-kDa polypeptide undergoing processing at two potential proteolytic cleavage sites, located at Glu109-Ser110 (site 1) and Gln157-Lys158 (site 2). Cleavage of proheparanase yields 8- and 50-kDa subunits that heterodimerize to form the active enzyme. The fate of the linker segment (Ser110-Gln157) residing between the two subunits, the mode of processing, and the protease(s) engaged in proheparanase processing are currently unknown. We applied multiple site-directed mutagenesis and deletions to study the nature of the potential cleavage sites and amino acids essential for processing of proheparanase in transfected human choriocarcinoma cells devoid of endogenous heparanase but possessing the enzymatic machinery for proper processing and activation of the proenzyme. Although mutagenesis at site 1 and its flanking sequences failed to identify critical residues for proteolytic cleavage, processing at site 2 required a bulky hydrophobic amino acid at position 156 (i.e. P2 of the cleavage site). Substitution of Tyr156 by Ala or Glu, but not Val, resulted in cleavage at an upstream site in the linker segment, yielding an improperly processed inactive enzyme. Processing of the latent 65-kDa proheparanase in transfected Jar cells was inhibited by a cell-permeable inhibitor of cathepsin L. Moreover, recombinant 65-kDa proheparanase was processed and activated by cathepsin L in a cell-free system. Altogether, these results suggest that proheparanase processing at site 2 is brought about by cathepsin L-like proteases. The involvement of other members of the cathepsin family with specificity to bulky hydrophobic residues cannot be excluded. Our results and a three-dimensional model of the enzyme are expected to accelerate the design of inhibitory molecules capable of suppressing heparanase mediated enhancement of tumor angiogenesis and metastasis. PMID- 15659390 TI - Strong in vivo maturation compensates for structurally restricted H3 loops in antibody repertoires. AB - A central paradigm in immunology states that successful generation of high affinity antibodies necessitates an immense primary repertoire of antigen combining sites. Much of the diversity of this repertoire is provided by varying one antigen binding loop, created by inserting randomly a D (diversity) gene out of a small pool between the V and J genes. It is therefore assumed that any particular D-encoded region surrounded by different V and J regions adopts a different conformation. We have solved the structure of two lysozyme-specific variable domains of heavy-chain antibodies isolated from two strictly unrelated dromedaries. These antibodies recombined identical D gene sequences to different V and J precursors with significant variance in their V(D)J junctions. Despite these large differences, the D-encoded loop segments adopt remarkably identical architectures, thus directing the antibodies toward identical epitopes. Furthermore, a striking convergent maturation process occurred in the V region, adapting both binders for their sub-nanomolar affinity association with lysozyme. Hence, on a structural level, humoral immunity may rely more on well developed maturation and selection systems than on the acquisition of large primary repertoires. PMID- 15659391 TI - Repression of an interleukin-4-responsive promoter requires cooperative BCL-6 function. AB - BCL-6 functions as a potent transcriptional repressor that binds with specificity to DNA elements bearing marked similarity to STAT recognition sequences. Previous studies have demonstrated that BCL-6 and Stat6 can both bind and regulate the Iepsilon promoter that controls immunoglobulin heavy chain class switching to IgE. Examination of BCL-6-/- and BCL-6-/-Stat6-/- mice has demonstrated that BCL 6 is a repressor of IgE and that Stat6 is still required for the interleukin-4 (IL-4) induction of class switching to IgE in B cells lacking BCL-6. To define the mechanisms by which BCL-6 represses IL-4 function, we analyzed the role of BCL-6 in repressing the Iepsilon promoter. There are three BCL-6-binding sites within this IL-4-responsive promoter. Analysis of Iepsilon promoters that have mutated BCL-6-binding sites demonstrates that at least two of these sites are required for maximal BCL-6 repression of this locus. Footprinting analysis demonstrates that BCL-6 binds cooperatively to the two upstream binding sites in the Iepsilon promoter. This cooperative binding requires the POZ domain of BCL-6. Furthermore, activated Stat6 molecules can displace BCL-6 from one of these binding sites. These data demonstrate that cooperative interaction between BCL-6 molecules is required for repression of the Iepsilon promoter. PMID- 15659393 TI - Seeding-dependent maturation of beta2-microglobulin amyloid fibrils at neutral pH. AB - Beta2-microglobulin (beta2-m) is a major component of amyloid fibrils deposited in patients with dialysis-related amyloidosis. Recent studies have focused on the mechanism by which amyloid fibrils are formed under physiological conditions, which had been difficult to reproduce quantitatively. Yamamoto et al. (Yamamoto, S., Hasegawa, K., Yamaguchi, I., Tsutsumi, S., Kardos, J., Goto, Y., Gejyo, F. & Naiki, H. (2004) Biochemistry 43, 11075-11082) showed that a combination of seed fibrils prepared under acidic conditions and a low concentration of sodium dodecyl sulfate below its critical micelle concentration enabled extensive fibril formation at pH 7.0. Here, we found that repeated self-seeding at pH 7.0 with fibrils formed at the same pH causes a marked acceleration of growth, indicating the maturation of fibrils. The observed maturation can be simulated by assuming the existence of two types of fibrils with different growth rates. Importantly, some mutations of beta2-m or the addition of a low concentration of urea, both destabilizing the native conformation, were not enough to extend the fibrils at pH 7.0, and a low concentration of sodium dodecyl sulfate (i.e. 0.5 mM) was essential. Thus, even though the first stage fibrils in patients are unstable and require stabilizing factors to remain at neutral pH, they can adapt to a neutral pH with repeated self-seeding, implying a mechanism of development of amyloid deposition after a long latent period in patients. PMID- 15659392 TI - Cell cycle and developmental regulations of replication factors in mouse embryonic stem cells. AB - Embryonic stem (ES) cells can grow rapidly and permanently while maintaining their differentiation capacity. To gain insight into how the cell cycle progression of undifferentiated murine ES cells is regulated, we have examined the expression patterns of various replication and cell cycle regulators. Most factors including cyclins, Cdc6, and geminin are rather constitutively expressed during the cell cycle of ES cells. Furthermore, the transcript levels of almost all the cell cycle regulators we investigated except for p21 and p27 are higher in undifferentiated ES cells than in murine embryonic fibroblasts (MEFs), and the increased stability of mRNA in ES cells may be partially responsible for this at least with some of the factors. More strikingly, the transcriptional levels of these factors are strongly correlated with the acetylated state of histone H3 at their promoter regions. However, the methylation state of histone or CpG methylation of the promoter region is not generally correlated significantly with the expression pattern of these factors in both cell types. On the protein level, Cdc6, ASK, cyclin A2, and cyclin B1 are extremely abundant in ES cells compared with MEFs. Furthermore, they are rapidly down-regulated upon induction of differentiation of ES cells. The significance of these findings is discussed in relation to the unusual proliferative properties of ES cells in an undifferentiated state. PMID- 15659394 TI - Trolox and 17beta-estradiol protect against amyloid beta-peptide neurotoxicity by a mechanism that involves modulation of the Wnt signaling pathway. AB - Oxidative stress is a key mechanism in amyloid beta-peptide (A beta)-mediated neurotoxicity; therefore, the protective roles of 17beta-estradiol (E2) and antioxidants (Trolox and vitamin C) were assayed on hippocampal neurons. Our results show the following: 1) E2 and Trolox attenuated the neurotoxicity mediated by A beta and H2O2 as measured by 3-(4,5-dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide reduction assays, quantification of apoptotic cells, and morphological studies of the integrity of the neurite network. 2) Vitamin C failed to protect neurons from A beta toxicity. 3) A beta-mediated endoperoxide production, reported to induce cell damage, was decreased in the presence of E2 and Trolox. 4) Two key Wnt signaling components were affected by E2 and Trolox; in fact, the enzyme glycogen synthase kinase 3beta was inhibited by both E2 and Trolox, and both compounds were able to stabilize cytoplasmic beta-catenin. 5) E2 activated the expression of the Wnt-5a and Wnt-7a ligands, and at the same time, E2, through the alpha-estrogen receptor, was able to prevent the excitotoxic A beta-induced rise in bulk-free Ca2+ as an alternative pathway to increase cell viability. 6) Finally, the Wnt-7a ligand protected against cytoplasmic calcium disturbances induced by A beta treatment. Our results suggest that control of oxidative stress, regulation of cytoplasmic calcium, and activation of Wnt signaling may prevent A beta neurotoxicity. PMID- 15659395 TI - Binding of two flaviolin substrate molecules, oxidative coupling, and crystal structure of Streptomyces coelicolor A3(2) cytochrome P450 158A2. AB - Cytochrome P450 158A2 (CYP158A2) is encoded within a three-gene operon (sco1206 sco1208) in the prototypic soil bacterium Streptomyces coelicolor A3(2). This operon is widely conserved among streptomycetes. CYP158A2 has been suggested to produce polymers of flaviolin, a pigment that may protect microbes from UV radiation, in combination with the adjacent rppA gene, which encodes the type III polyketide synthase, 1,3,6,8-tetrahydroxynaphthalene synthase. Following cloning, expression, and purification of this cytochrome P450, we have shown that it can produce dimer and trimer products from the substrate flaviolin and that the structures of two of the dimeric products were established using mass spectrometry and multiple NMR methods. A comparison of the x-ray structures of ligand-free (1.75 angstroms) and flaviolin-bound (1.62 angstroms) forms of CYP158A2 demonstrates a major conformational change upon ligand binding that closes the entry into the active site, partly due to repositioning of the F and G helices. Particularly interesting is the presence of two molecules of flaviolin in the closed active site. The flaviolin molecules form a quasi-planar three molecule stack including the heme of CYP158A2, suggesting that oxidative C-C coupling of these phenolic molecules leads to the production of flaviolin dimers. PMID- 15659396 TI - Crystal structure of human SCO1: implications for redox signaling by a mitochondrial cytochrome c oxidase "assembly" protein. AB - Human SCO1 and SCO2 are copper-binding proteins involved in the assembly of mitochondrial cytochrome c oxidase (COX). We have determined the crystal structure of the conserved, intermembrane space core portion of apo-hSCO1 to 2.8 A. It is similar to redox active proteins, including thioredoxins (Trx) and peroxiredoxins (Prx), with putative copper-binding ligands located at the same positions as the conserved catalytic residues in Trx and Prx. SCO1 does not have disulfide isomerization or peroxidase activity, but both hSCO1 and a sco1 null in yeast show extreme sensitivity to hydrogen peroxide. Of the six missense mutations in SCO1 and SCO2 associated with fatal mitochondrial disorders, one lies in a highly conserved exposed surface away from the copper-binding region, suggesting that this region is involved in protein-protein interactions. These data suggests that SCO functions not as a COX copper chaperone, but rather as a mitochondrial redox signaling molecule. PMID- 15659397 TI - Secretion of the human T cell leukemia virus type I transactivator protein tax. AB - Human T cell leukemia virus type I (HTLV-I) is the etiologic agent of adult T cell leukemia and HTLV-I-associated myelopathy/tropical spastic paraparesis. The HTLV-I protein Tax is well known as a transcriptional transactivator and inducer of cellular transformation. However, it is also known that extracellular Tax induces the production and release of cytokines, such as tumor necrosis factor alpha and interleukin-6, which have adverse effects on cells of the central nervous system. The cellular process by which Tax exits the cell into the extracellular environment is currently unknown. In most cell types, Tax has been shown to localize primarily to the nucleus. However, Tax has also been found to accumulate in the cytoplasm. The results contained herein begin to characterize the process of Tax secretion from the cell. Specifically, cytoplasmic Tax was demonstrated to localize to organelles associated with the cellular secretory process including the endoplasmic reticulum and Golgi complex. Additionally, it was demonstrated that full-length Tax was secreted from both baby hamster kidney cells and a human kidney tumor cell line, suggesting that Tax enters the secretory pathway in a leaderless manner. Tax secretion was partially inhibited by brefeldin A, suggesting that Tax migrated from the endoplasmic reticulum to the Golgi complex. In addition, combined treatment of Tax-transfected BHK-21 cells with phorbol myristate acetate and ionomycin resulted in a small increase in the amount of Tax secreted, suggesting that a fraction of cytoplasmic Tax was present in the regulated secretory pathway. These studies begin to provide a link between Tax localization to the cytoplasm, the detection of Tax in the extracellular environment, its possible role as an extracellular effector molecule, and a potential role in neurodegenerative disease associated with HTLV I infection. PMID- 15659398 TI - The role of mitochondria for Ca2+ refilling of the endoplasmic reticulum. AB - Endoplasmic reticulum (ER) Ca2+ refilling is an active process to ensure an appropriate ER Ca2+ content under basal conditions and to maintain or restore ER Ca2+ concentration during/after cell stimulation. The mechanisms to achieve successful ER Ca2+ refilling are multiple and built on a concerted action of processes that provide a suitable reservoir for Ca2+ sequestration into the ER. Despite mitochondria having been found to play an essential role in the maintenance of capacitative Ca2+ entry by buffering subplasmalemmal Ca2+, their contribution to ER Ca2+ refilling was not subjected to detailed analysis so far. Thus, this study was designed to elucidate the involvement of mitochondria in Ca2+ store refilling during and after cell stimulation. ER Ca2+ refilling was found to be accomplished even during continuous inositol 1,4,5-trisphosphate (IP3)-triggered ER Ca2+ release by an agonist. Basically, ER Ca2+ refilling depended on the presence of extracellular Ca2+ as the source and sarcoplasmic/endoplasmic reticulum Ca2+ ATPase (SERCA) activity. Interestingly, in the presence of an IP3-generating agonist, ER Ca2+ refilling was prevented by the inhibition of trans-mitochondrial Ca2+ flux by CGP 37157 (7-chloro-5-(2 chlorophenyl)-1,5-dihydro-4,1-benzothiazepin-2(3H)-one) that precludes the mitochondrial Na+/Ca2+ exchanger as well as by mitochondrial depolarization using a mixture of oligomycin and antimycin A. In contrast, after the removal of the agonist, ER refilling was found to be largely independent of trans-mitochondrial Ca2+ flux. Under these conditions, ER Ca2+ refilling took place even without an associated Ca2+ elevation in the deeper cytosol, thus, indicating that superficial ER domains mimic mitochondrial Ca2+ buffering and efficiently sequester subplasmalemmal Ca2+ and consequently facilitate capacitative Ca2+ entry. Hence, these data point to different contribution of mitochondria in the process of ER Ca2+ refilling based on the presence or absence of IP3, which represents the turning point for the dependence or autonomy of ER Ca2+ refilling from trans-mitochondrial Ca2+ flux. PMID- 15659399 TI - Identification of LAT4, a novel amino acid transporter with system L activity. AB - System L amino acid transporters mediate the movement of bulky neutral amino acids across cell membranes. Until now three proteins that induce system L activity have been identified: LAT1, LAT2, and LAT3. The former two proteins belong to the solute carrier family 7 (SLC7), whereas the latter belongs to SLC43. In the present study we present a new cDNA, designated LAT4, which also mediates system L activity when expressed in Xenopus laevis oocytes. Human LAT4 exhibits 57% identity to human LAT3. Like LAT3, the amino acid transport activity induced by LAT4 is sodium-, chloride- and pH-independent, is not trans stimulated, and shows two kinetic components. The low affinity component of LAT4 induced activity is sensitive to the sulfhydryl-specific reagent N-ethylmaleimide but not that with high affinity. Mutation in LAT4 of the SLC43 conserved serine 297 to alanine abolishes sensitivity to N-ethylmaleimide. LAT4 activity is detected at the basolateral membrane of PCT kidney cells. In situ hybridization experiments show that LAT4 mRNA is restricted to the epithelial cells of the distal tubule and the collecting duct in the kidney. In the intestine, LAT4 is mainly present in the cells of the crypt. PMID- 15659400 TI - Activation of the Trk signaling pathway by extracellular zinc. Role of metalloproteinases. AB - In certain brain regions, extracellular zinc concentrations can rise precipitously as intense neuronal activity releases large amounts of zinc from the nerve terminals. Although zinc release has been suggested to play a pathological role, its precise physiological effect is poorly understood. Here, we report that exposure to micromolar quantities of zinc for only a few minutes robustly and specifically activated tropomyosin-related kinase (Trk) receptors, most likely TrkB, in cultured cortical neurons. We further found that Trk activation by zinc is extracellularly mediated by activation of metalloproteinases, which release pro-BDNF from cells and convert pro-BDNF to mature BDNF. These results suggest that activity-dependent release of extracellular zinc leads to metalloproteinase activation, which plays a critically important role in Trk receptor activation at zinc-containing synapses. PMID- 15659402 TI - Novel symmetric and asymmetric DNA scission determinants for Streptococcus pneumoniae topoisomerase IV and gyrase are clustered at the DNA breakage site. AB - Topoisomerase (topo) IV and gyrase are bacterial type IIA DNA topoisomerases essential for DNA replication and chromosome segregation that act via a transient double-stranded DNA break involving a covalent enzyme-DNA "cleavage complex." Despite their mechanistic importance, the DNA breakage determinants are not understood for any bacterial type II enzyme. We investigated DNA cleavage by Streptococcus pneumoniae topo IV and gyrase stabilized by gemifloxacin and other antipneumococcal fluoroquinolones. Topo IV and gyrase induce distinct but overlapping repertoires of double-strand DNA breakage sites that were essentially identical for seven different quinolones and were augmented (in intensity) by positive or negative supercoiling. Sequence analysis of 180 topo IV and 126 gyrase sites promoted by gemifloxacin on pneumococcal DNA revealed the respective consensus sequences: G(G/c)(A/t)A*GNNCt(T/a)N(C/a) and GN4G(G/c)(A/c)G*GNNCtTN(C/a) (preferred bases are underlined; disfavored bases are in small capitals; N indicates no preference; and asterisk indicates DNA scission between -1 and +1 positions). Both enzymes show strong preferences for bases clustered symmetrically around the DNA scission site, i.e. +1G/+4C, 4G/+8C, and particularly the novel -2A/+6T, but with no preference at +2/+3 within the staggered 4-bp overhang. Asymmetric elements include -3G and several unfavored bases. These cleavage preferences, the first for Gram-positive type IIA topoisomerases, differ markedly from those reported for Escherichia coli topo IV (consensus (A/G)*T/A) and gyrase, which are based on fewer sites. However, both pneumococcal enzymes cleaved an E. coli gyrase site suggesting overlap in gyrase determinants. We propose a model for the cleavage complex of topo IV/gyrase that accommodates the unique -2A/+6T and other preferences. PMID- 15659401 TI - Characterization of the yeast trimeric-SAS acetyltransferase complex. AB - The yeast SAS2 (Something About Silencing 2) gene encodes a member of the MYST protein family of histone acetyltransferases (HATs) and is involved in transcriptional silencing at all silent loci (HML, HMR, telomeres, and rDNA) in Saccharomyces cerevisiae. Sas2 is the catalytic subunit of a yeast histone acetyltransferase complex termed SAS complex. The enzymatic activity of SAS complex on free histones has been reported, but nucleosomal HAT activity has not yet been documented. Here we show that the native yeast SAS complex is a small trimeric protein complex composed solely of Sas2, Sas4, and Sas5 with a molecular mass of about 125 kDa. It is capable of acetylating both free histones and nucleosomes, although the nucleosomal HAT activity of SAS complex is very weak when compared with that of NuA4, the other member of MYST HAT complex. We also demonstrate that the putative acetyl CoA binding motif in Sas2 is essential for both the in vivo silencing function and the enzymatic activity of SAS complex. Unlike NuA4, which acetylates all four available lysines at the N-terminal tail of histone H4, SAS complex exclusively acetylates lysine 16 of histone H4 in vitro and is required for the bulk of H4 lysine 16 acetylation in vivo. This specific lysine preference corresponds to the role of SAS complex in antagonizing the spreading of Sir proteins at silent loci in S. cerevisiae. PMID- 15659403 TI - Characterization of the protein dimerization domain responsible for assembly of functional selenodeiodinases. AB - Thyroid hormone metabolism is catalyzed by a small family of selenoenzymes. Type I deiodinase (D1) is the best characterized family member and is an integral membrane protein composed of two 27-kDa subunits that assemble to a functional holoenzyme after translation. To characterize the protein domain(s) responsible for this post-translational assembly event, we used deletion/truncation analysis coupled with immune depletion assays to map the dimerization domain of D1. The results of our studies show that a highly conserved sequence of 16 amino acids in the C-terminal half of the D1 subunit, -D148FL-YI-EAH-DGW163-, serves as the dimerization domain. Based on the high conservation of this domain, we synthesized a novel bait peptide-green fluorescent protein fusion probe (DDD(GFP)) to examine holoenzyme assembly of other family members. Overexpression of either the DDD(GFP) or an inert D1 subunit (M4) into SeD2 (accession number U53505)-expressing C6 cells specifically led to the loss of >90% of the catalytic activity. Catalytically inactive D2 heterodimers composed of SeD2: DDD(GFP) subunits were rescued by specific immune precipitation with anti-SeD2 IgG, suggesting that SeD2 requires two functional subunits to assemble a catalytically active holoenzyme. These findings identify and characterize the essential dimerization domain responsible for post-translational assembly of selenodeiodinases and show that family members can intermingle through this highly conserved protein domain. PMID- 15659404 TI - Maturation of bacteriophage T4 lagging strand fragments depends on interaction of T4 RNase H with T4 32 protein rather than the T4 gene 45 clamp. AB - In the bacteriophage T4 DNA replication system, T4 RNase H removes the RNA primers and some adjacent DNA before the lagging strand fragments are ligated. This 5'-nuclease has strong structural and functional similarity to the FEN1 nuclease family. We have shown previously that T4 32 protein binds DNA behind the nuclease and increases its processivity. Here we show that T4 RNase H with a C terminal deletion (residues 278-305) retains its exonuclease activity but is no longer affected by 32 protein. T4 gene 45 replication clamp stimulates T4 RNase H on nicked or gapped substrates, where it can be loaded behind the nuclease, but does not increase its processivity. An N-terminal deletion (residues 2-10) of a conserved clamp interaction motif eliminates stimulation by the clamp. In the crystal structure of T4 RNase H, the binding sites for the clamp at the N terminus and for 32 protein at the C terminus are located close together, away from the catalytic site of the enzyme. By using mutant T4 RNase H with deletions in the binding site for either the clamp or 32 protein, we show that it is the interaction of T4 RNase H with 32 protein, rather than the clamp, that most affects the maturation of lagging strand fragments in the T4 replication system in vitro and T4 phage production in vivo. PMID- 15659405 TI - CK2 phosphorylates SSRP1 and inhibits its DNA-binding activity. AB - We have previously shown that CK2 associates with the human high-mobility group protein SSRP1 and that this association increases in response to UV irradiation. CK2 also phosphorylates SSRP1 in vitro. Here we extend this work by investigating CK2 regulation of SSRP1 function through phosphorylation. Phosphorylation of SSRP1 by CK2 inhibited the nonspecific DNA-binding activity of SSRP1 and FACT (facilitating chromatin-mediated transcription) complex in vitro. Using a serine/threonine-scanning Auto-spot peptide array coupled with a filter-based kinase assay with synthetic peptides as substrates, we identified serines 510, 657, and 688 as phosphorylation targets of CK2 in vitro. Mutagenesis of the three serines revealed that serine 510 was more important for the regulation of SSRP1 DNA-binding activity. Furthermore, we found that SSRP1 was phosphorylated in cells in response to UV (but not gamma) irradiation. These results suggest that CK2 regulates the DNA-binding ability of SSRP1 and that this regulation may be responsive to specific cell stresses. PMID- 15659406 TI - Integrative genomics revealed RAI3 is a cell growth-promoting gene and a novel P53 transcriptional target. AB - In this study, differential gene expression between normal human mammary epithelial cells and their malignant counterparts (eight well established breast cancer cell lines) was studied using Incyte GeneAlbum 1-6, which contains 65,873 cDNA clones representing 33,515 individual genes. 3,152 cDNAs showed a > or =3.0 fold expression level change in at least one of the human breast cancer cell lines as compared with normal human mammary epithelial cells. Integration of breast tumor gene expression data with the genes in the tumor suppressor p53 signaling pathway yielded 128 genes whose expression is altered in breast tumor cell lines and in response to p53 expression. A hierarchical cluster analysis of the 128 genes revealed that a significant portion of genes demonstrate an opposing expression pattern, i.e. p53-activated genes are down-regulated in the breast tumor lines, whereas p53-repressed genes are up-regulated. Most of these genes are involved in cell cycle regulation and/or apoptosis, consistent with the tumor suppressor function of p53. Follow-up studies on one gene, RAI3, suggested that p53 interacts with the promoter of RAI3 and repressed its expression at the onset of apoptosis. The expression of RAI3 is elevated in most tumor cell lines expressing mutant p53, whereas RAI3 mRNA is relatively repressed in the tumor cell lines expressing wild-type p53. Furthermore, ectopic expression of RAI3 in 293 cells promotes anchorage-independent growth and small interfering RNA mediated depletion of RAI3 in AsPc-1 pancreatic tumor cells induces cell morphological change. Taken together, these data suggest a role for RAI3 in tumor growth and demonstrate the predictive power of integrative genomics. PMID- 15659408 TI - A systematic approach to evaluation of the rearfoot, ankle, and leg in reconstructive surgery. AB - The current literature shows that proper alignment of the lower extremity allows for greater function throughout the gait cycle. Therefore, realignment should be one of the primary goals in the surgical management of lower-extremity deformities and pathology. Multiplanar radiographic angular relationships should be critically evaluated to appropriately identify the level and extent of the deformity before performing realignment procedures. This article describes a systematic approach to deformity evaluation through a comprehensive radiographic assessment of the rearfoot, ankle, and lower leg. PMID- 15659409 TI - Realignment considerations in the triple arthrodesis. AB - Triple arthrodesis is indicated for pain, deformity, and instability of the tritarsal complex. Although osseous consolidation is important to outcome, success depends primarily on appropriate realignment. A poorly positioned triple arthrodesis may result in continued pain and disability, gait disturbances, excessive stress in adjacent joints, and footwear problems. We present a protocol for intraoperative realignment of the triple arthrodesis guided by image intensification. This protocol for alignment evaluation was followed in nine patients who underwent triple arthrodesis for a variety of pathologies. PMID- 15659410 TI - Gastrocnemius soleus recession: a simpler, more limited approach. AB - Multiple surgical procedures have been described for the correction of equinus deformity. We present a review of the anatomy, biomechanics, and clinical assessment of equinus. In addition, we provide a detailed surgical technique for gastrocnemius soleus recession and introduce an anatomical guide for surgical treatment. PMID- 15659411 TI - Static rearfoot alignment: a comparison of clinical and radiographic measures. AB - Foot structure is typically evaluated using static clinical and radiographic measures. To date, the literature is devoid of a correlation between rearfoot frontal plane radiographic parameters and clinical measures of alignment. In a repeated-measures study comparing radiographic and clinical rearfoot alignment in 24 healthy subjects, radiographic angular measurements were made from standard weightbearing anteroposterior, lateral, long leg calcaneal axial, and rearfoot alignment views. Clinical measurements were made using a jig and scanner to assess the malleolar valgus index and a goniometer to evaluate the resting and neutral calcaneal stance positions. There was a significant correlation between frontal plane radiographic angles (long leg calcaneal axial and rearfoot alignment views) (r = 0.814). Similarly, there was a significant correlation between clinical measures (resting calcaneal stance position and malleolar valgus index) (r = 0.714). A multivariate stepwise regression showed that resting calcaneal stance position can be accurately predicted from 3 of the 15 clinical and radiographic measurements collected: malleolar valgus index, rearfoot alignment view, and long leg calcaneal axial view (r = 0.829). In summary, a commonly used clinical measure of static rearfoot alignment, resting calcaneal stance position, was correlated closely with the malleolar valgus index and both frontal plane radiographic parameters. PMID- 15659412 TI - Subtalar joint arthrodesis. AB - Forty patients (12 men and 28 women) treated with isolated subtalar joint arthrodesis were retrospectively reviewed. The average patient age was 50 years (range, 21-76 years). Preoperative diagnoses included posterior tibial tendon dysfunction, post-traumatic arthritis, nontraumatic arthritis, and subtalar joint middle facet coalition. The average follow-up was 15 months (range, 12-74 months). Subjective postoperative questionnaire results were classified as satisfied (n = 32), satisfied but with reservations (n = 4), or dissatisfied (n = 4). Eighty-three percent of the patients (n = 33) stated that they would undergo the procedure again. Minor complications (those that resolved with nonoperative treatment) occurred in 55% of the patients. However, the major complication rate was only 12.5%. This study showed no statistical correlation between the preoperative diagnosis and the postoperative outcome. Our results also suggested that the prevalence of complications is slightly higher than in previous reports. Isolated subtalar joint arthrodesis is an effective treatment for pain and deformity of the rearfoot. PMID- 15659413 TI - Pes cavus. AB - Pes cavus is a complex deformity with various components. The etiology is multifaceted, which can result in changes to the rearfoot, forefoot, or both areas. Proper patient evaluation is crucial to understanding the deformity and its management. Various soft-tissue and osseous procedures may be used in the reconstruction of the symptomatic cavus foot deformity. This article discusses the clinical and radiographic findings of pes cavus, as well as the various etiologies of this challenging foot deformity and the surgical principles of its correction. PMID- 15659414 TI - Double calcaneal osteotomy: realignment considerations in eight patients. AB - Posterior calcaneal displacement and anterior calcaneal distraction osteotomies have been recommended for surgical management of flexible flatfoot deformity. We combined these procedures for surgical management of adolescent flatfoot and late stage II posterior tibial tendon dysfunction in the adult. Lateral column distraction has been shown to reduce rearfoot valgus and forefoot abduction by stabilizing the midtarsal joint. Posterior calcaneal displacement osteotomy, which results in medial translation of the tuberosity, converts the posterior muscle group from an everter to an inverter. A supinatory moment, therefore, is created about the subtalar joint axis, which results in supination during the midstance phase of gait. These procedures may be considered when calcaneal valgus, forefoot abduction, and midtarsal joint instability are clinically significant. We also include rearfoot alignment radiographs, long leg calcaneal axial radiographs, and malleolar valgus indices to further evaluate alignment of the foot, ankle, and leg. Ideal realignment is achieved when the heel is vertical in resting calcaneal stance position, the forefoot is parallel to the rearfoot in the frontal plane, and the medial column is stabilized, with elimination of forefoot abduction. PMID- 15659416 TI - Percutaneous supramalleolar osteotomy for distal tibial (near articular) ankle deformities. AB - Supramalleolar osteotomies are performed to realign the distal tibia and to improve foot and ankle function. This procedure requires a thorough preoperative and intraoperative understanding of limb-deformity parameters and of the center of rotation of angulation method of surgical planning. Radiographic, gait, and clinical analyses along with intraoperative fluoroscopic images are paramount to deformity correction. This article describes the radiographic and clinical analyses and technical considerations in performing a focal dome supramalleolar osteotomy. PMID- 15659415 TI - Realignment arthrodesis of the rearfoot and ankle: a comprehensive evaluation. AB - Ankle and tibiotalocalcaneal arthrodeses are performed for the treatment of painful, arthritic, unstable, and deformed rearfoot and ankle joints. Surgical complications are not uncommon (approximately 30%); some can be attributed to poor preoperative planning and inadequate intraoperative position. Several authors have attempted to define the optimal position for ankle arthrodesis without objective multiplanar radiographic analysis and consistent reference points. This investigation explored the effects of ankle and tibiotalocalcaneal realignment arthrodeses on static lower-extremity position in 20 patients. The most common preoperative diagnosis was severe degenerative joint disease following ankle fractures and ankle instability. Seven tibiotalocalcaneal arthrodeses and 13 isolated ankle arthrodeses were performed (mean follow-up, 22 months). Average time to radiographic osseous union of the isolated ankle and tibiotalocalcaneal arthrodeses was 11 and 7 weeks, respectively. Medical complications occurred in 2 patients (10%). There were no statistically significant differences between preoperative and postoperative angular relationships. This study objectively quantifies multiplanar foot-to-leg realignment and defines the optimal clinical and radiographic positions for ankle and tibiotalocalcaneal realignment arthrodeses. PMID- 15659417 TI - Technical considerations in tarsometatarsal joint arthrodesis. AB - Tarsometatarsal arthrodesis is indicated for degenerative joint disease after trauma. Deformity is often associated with post-traumatic arthropathy. Outcomes after surgical management are directly related to realignment. This article reviews the indications, preoperative evaluation, technical execution, and postoperative management of tarsometatarsal arthrodesis. Special emphasis is placed on realignment and restoration of normal foot architecture. PMID- 15659418 TI - Arthroscopy-assisted retrograde drilling of osteochondral lesions of the talar dome. AB - Traumatic ankle conditions can lead to long-term sequelae if a pathologic process is misdiagnosed. The clinical presentation of an osteochondral lesion of the talar dome requires the clinician to have a high index of suspicion, and advanced imaging is often necessary to make the final diagnosis. Treatment should be initiated once the lesion is appropriately staged by radiologic or magnetic resonance imaging. We discuss the use of arthroscopy-assisted retrograde drilling of the medial talar dome that spares the articular cartilage within the talotibial articulation. PMID- 15659419 TI - Topology of small-world networks of protein-protein complex structures. AB - The majority of real examples of small-world networks exhibit a power law distribution of edges among the nodes, therefore not fitting into the wiring model proposed by Watts and Strogatz. However, protein structures can be modeled as small-world networks, with a distribution of the number of links decaying exponentially as in the case of this wiring model. We approach the protein protein interaction mechanism by viewing it as a particular rewiring occurring in the system of two small-world networks represented by the monomers, where a re arrangement of links takes place upon dimerization leaving the small-world character in the dimer network. Due to this rewiring, the most central residues at the complex interfaces tend to form clusters, which are not homogenously distributed. We show that these highly central residues are strongly correlated with the presence of hot spots of binding free energy. CONTACT: ao mesa@fujirebio.co.jp SUPPLEMENTARY INFORMATION: http://www.fujirebio.co.jp/support/index.php (under construction). PMID- 15659420 TI - Sequential high-dose chemotherapy protocol for relapsed poor prognosis germ cell tumors combining two mobilization and cytoreductive treatments followed by three high-dose chemotherapy regimens supported by autologous stem cell transplantation. Results of the phase II multicentric TAXIF trial. AB - BACKGROUND: High-dose chemotherapy (HD-CT) is able to circumvent platinum resistance of resistant/refractory germ-cell tumors (GCTs), but expectancy of cure remains low. New strategies are needed with new drugs and a sequential approach. MATERIALS AND METHODS: Patients with relapsed poor-prognosis GCTs were scheduled to receive two cycles combining epirubicin and paclitaxel (Taxol) followed by three consecutive HD-CT supported by stem cell transplantation [one course combining cyclophosphamide, 3 g/m(2) + thiotepa, 400 mg/m(2), followed by two ICE regimens (ifosfamide, 10 g/m(2), carboplatin, AUC 20, etoposide, 1500 mg/m(2))]. RESULTS: From March 1998 to September 2001 (median follow-up, 31.8 months), 45 patients (median age, 28 years) were enrolled in this phase II study. Twenty-two patients received the complete course. Twenty-five patients died from progression and five from toxicity. The overall response rate was 37.7%, including an 8.9% complete response rate. The median overall survival was 11.8 months. The 3-year survival and progression-free survival rate was 23.5%. The 'Beyer' prognostic score predicted the outcome after HD-CT. CONCLUSION: Although our results warrant further studies on HD-CT in relapsed poor prognosis GCTs, patients with a Beyer score >2 did not benefit from this approach and should not be enrolled in HD-CT trials. Better selection criteria have to be fulfilled in forthcoming studies. PMID- 15659421 TI - The role of periventricular nodular heterotopia in epileptogenesis. AB - A temporal resection in patients with periventricular nodular heterotopia (PNH) and intractable focal seizures yields poor results. To define the role of heterotopic grey matter tissue in epileptogenesis and to improve outcome, we performed stereoencephalography (SEEG) recordings in eight patients with uni- or bilateral PNH and intractable focal epilepsy. The SEEG studies aimed to evaluate the most epileptogenic areas and included the allo- and neocortex and at least one nodule of grey matter. Interictal spiking activity was found in ectopic grey matter in three patients, in the cortex overlying the nodules in five and in the mesial temporal structures in all. At least one heterotopion was involved at seizure onset in six patients, synchronous with the overlying neocortex or ipsilateral hippocampus. Two patients had their seizures originating in the mesial temporal structures only. Six patients had surgery and the resected areas included the seizure onset, with follow-up from 1 to 8 years. An amygdalo hippocampectomy was performed in two (Engel class Id and III), an amygdalo hippocampectomy plus removal of an adjacent heterotopion in two (class Ia), and a resection of two contiguous nodules plus a small rim of overlying occipital cortex in one patient (class Id). One patient with bilateral PNH had three adjacent nodules resected and an ipsilateral amygdalo-hippocampectomy resulting in a reduction of the number of seizures by 25-50%. The best predictor of surgical outcome is the presence of a focal epileptic generator; this generator may or may not include the PNH. Invasive recording is required in patients with PNH; it improves localization and is the key to better outcome. PMID- 15659422 TI - Early and late HHV-6 gene transcripts in multiple sclerosis lesions and normal appearing white matter. AB - Multiple sclerosis is an inflammatory demyelinating disease of the CNS, the aetiology of which is believed to have both genetic and environmental components. We have investigated one of the candidate viruses for the environmental component of multiple sclerosis, the neurotropic human herpesvirus 6 (HHV-6). Utilizing fluorescent in situ hybridization (FISH) techniques, we have examined human post mortem tissues for the presence of immediate early and late viral gene expression in multiple sclerosis patient normal appearing white matter (NAWM), lesional tissue and normal control brain samples. HHV-6 gene transcription was detected in all tissue samples and was restricted to oligodendrocytes, as determined by double mRNA FISH analysis. Quantitative analysis of viral mRNA expression indicated that both NAWM and lesional multiple sclerosis samples exhibited significantly higher levels of HHV-6 expression compared with the normal control samples. Lesional samples exhibited the highest levels of viral gene expression, with NAWM exhibiting an intermediate level between lesional and control tissues. Immunofluorescence against early and late HHV-6 proteins verified active translation of HHV-6 viral mRNA in oligodendrocytes. Southern blot analysis of nested polymerase chain reactions using extracted genomic DNA and cDNA confirmed the presence of the HHV-6 genome in all individuals, with the active expression profile mirroring the FISH results. The frequent high level of HHV-6 infection in multiple sclerosis samples suggests a possible role in pathogenesis. PMID- 15659423 TI - The basal ganglia and rule-governed language use: evidence from vascular and degenerative conditions. AB - The Declarative/Procedural Model of Pinker, Ullman and colleagues claims that the basal ganglia are part of a fronto-striatal procedural memory system which applies grammatical rules to combine morphemes (the smallest meaningful units in language) into complex words (e.g. talk-ed, talk-ing). We tested this claim by investigating whether striatal damage or loss of its dopaminergic innervation is reliably associated with selective regular past tense deficits in patients with subcortical cerebrovascular damage, Parkinson's disease or Huntington's disease. We focused on past tense morphology since this allows us to contrast the regular past tense (jump-jumped), which is rule-based, with the irregular past tense (sleep-slept), which is not. We used elicitation and priming tasks to test patients' ability to comprehend and produce inflected forms. We found no evidence of a consistent association between striatal dysfunction and selective impairment of regular past tense morphology, suggesting that the basal ganglia are not essential for processing the regular past tense as a sequence of morphemes, either in comprehension or production, in contrast to the claims of the Declarative/Procedural Model. All patient groups showed normal activation of semantic and morphological representations in comprehension, despite difficulties suppressing semantically appropriate alternatives when trying to inflect novel verbs. This is consistent with previous reports that striatal dysfunction spares automatic activation of linguistic information, but disrupts later language processes that require inhibition of competing alternatives. PMID- 15659424 TI - Functional recovery of children and adolescents after cerebellar tumour resection. AB - This study examined whether lesions to the cerebellum obtained in early childhood are better compensated than lesions in middle childhood or adolescence. Since cerebellar lesions might affect motor as well a cognitive performance, posture, upper limb and working memory function were assessed in 22 patients after resection of a cerebellar tumour (age at surgery 1-17 years, minimum 3 years post surgery). Working memory was only impaired in those patients who had received chemo- or radiation therapy. Postural sway was enhanced in 64% of the patients during dynamic posturography conditions, which relied heavily on vestibular input for equilibrium control. Upper limb function was generally less impaired, but 54% of the patients revealed prolonged deceleration times in an arm pointing task, which probably does not reflect a genuine cerebellar deficit but rather the patients' adopted strategy to avoid overshooting. Age at surgery, time since surgery or lesion volume were poor predictors of motor or cognitive recovery. Brain imaging analysis revealed that lesions of all eight patients with abnormal posture who did not receive chemo- and/or radiation therapy included the fastigial and interposed nuclei (NF and NI). In patients with normal posture, NI and NF were spared. In 11 out of 12 patients with abnormal deceleration time, the region with the highest overlap included the NI and NF and dorsomedial portions of the dentate nuclei in 10 out of 12 patients. We conclude that cerebellar damage inflicted at a young age is not necessarily better compensated. The lesion site is critical for motor recovery, and lesions affecting the deep cerebellar nuclei are not fully compensated at any developmental age in humans. PMID- 15659425 TI - Clinical manifestation and natural course of late-onset Pompe's disease in 54 Dutch patients. AB - Late-onset Pompe's disease (acid maltase deficiency, glycogen storage disease type II) is a slowly progressive myopathy caused by deficiency of acid alpha glucosidase. Current developments in enzyme replacement therapy require detailed knowledge of the kind and severity of symptoms and the natural course of the disease in the patient population. A detailed questionnaire covering the patients' medical history and current situation was developed and information was gathered from 54 Dutch patients. The mean age of the participants was 48.6 +/- 15.6 years. The first complaints started at a mean age of 28.1 +/- 14.3 years and were mostly related to mobility problems and limb-girdle weakness. Fifty-eight percent of the adult patients indicated the presence of mild muscular symptoms during childhood. Twenty-eight percent of the patients waited >5 years for the final diagnosis after the first visit to a physician for disease-related complaints. At the time of questionnaire completion, 48% of the study population used a wheelchair and 37% used artificial ventilation. Movements such as rising from an armchair, taking stairs or getting upright after bending over were difficult or impossible for more than two-thirds of the respondents. The age at onset, the rate of disease progression and the sequence of respiratory and skeletal muscle involvement varied substantially between patients. Seventy-six percent of the participants indicated being troubled by fatigue and 46% by pain. This survey has mapped the age at onset, presenting symptoms, heterogeneity in progression and range of disease severity in a large group of Dutch patients. We conclude that early manifestations in childhood require proper attention to prevent unnecessary delay of the diagnosis. The follow-up of patients with late onset Pompe's disease should focus on respiratory and limb-girdle muscle function, the capacity to perform daily activities, and the presentation of fatigue and pain. PMID- 15659426 TI - Down-regulation of apoptosis mediators by RNAi inhibits axotomy-induced retinal ganglion cell death in vivo. AB - Transection of the optic nerve induces an apoptotic degeneration of retinal ganglion cells (RGC) in the rat retina. The immediate early gene c-Jun, the proapoptotic Bcl-2 family member Bax and the apoptosome constituent Apaf-1 have been shown previously to play major roles in the induction or execution of the apoptosis cascade. In this study we have designed and generated short interfering RNAs (siRNAs) against c-Jun, Bax and Apaf-1, which were injected into the optic nerve stump in order to inhibit axotomy-induced apoptosis. siRNAs were first tested in vitro to ensure silencing efficiency. In vivo, a clear neuronal localization of Cy3-labelled siRNA could be visualized in retinal flat mounts. Retinas that were injected with anti-Apaf-1- and anti-c-Jun-siRNA showed significantly more surviving RGC than non-injected or anti-EGFP-injected controls (approximately 2- to 3-fold, respectively). Anti-Bax-siRNA-injected retinas showed a trend towards an increased RGC number (not significant). Regulation of target proteins in situ could be visualized by immunohistochemical stainings. We conclude that (i) c-Jun and Apaf-1 play major roles in the apoptotic cascade of RGC and may represent useful targets for antiapoptotic strategies in RGC in vivo, and (ii) injection of siRNAs into the optic nerve stump is a new method to down regulate target genes specifically in RGC. PMID- 15659428 TI - Abeta-related angiitis: primary angiitis of the central nervous system associated with cerebral amyloid angiopathy. AB - Idiopathic or primary angiitis of the CNS (PACNS) and cerebral amyloid angiopathy (CAA) are unusual vasculopathies generally regarded as unrelated disorders. A few case reports have, however, described granulomatous angiitis in patients with sporadic, amyloid beta peptide (Abeta)-related CAA. Here we describe the clinical, neuroradiological and neuropathological features of nine patients with Abeta-related angiitis (ABRA). Combining these with the individual case reports drawn from the literature has allowed us to define ABRA as a clinical entity and to compare its features with those of PACNS. The mean age of presentation of ABRA (67 years) is higher than that of PACNS but lower than that of sporadic non inflammatory Abeta-related CAA. Alterations in mental status (59%), headaches (35%), seizures and focal neurological deficits (24%) are common. Hallucinations are a presenting manifestation in 12% of cases. Most patients have white matter hyperintensities on MRI but these are of similar appearance to those in PACNS. Cerebrospinal fluid usually shows modest elevation of protein and pleocytosis. Neuropathology reveals angiodestructive inflammation, often granulomatous, and meningeal lymphocytosis. Abeta is consistently present in abundance in affected blood vessels but usually scanty within the parenchyma of the cerebral cortex. However, the cortex includes numerous activated microglia, occasionally in a plaque-like distribution and containing cytoplasmic Abeta. The cerebral white matter shows patchy gliosis and rarefaction, in some cases marked. Our findings (i) help to dissect one separate clinicopathological entity from what is likely to be a spectrum of primary angiitides of the CNS; (ii) have important therapeutic implications for one category of patients with amyloid-related vasculopathy; and (iii) may provide valuable insights into the development of amyloid-associated inflammation, of relevance not only to ABRA but also to Abeta immunization-related encephalitis and to Alzheimer's disease. PMID- 15659427 TI - Widespread nuclear and cytoplasmic accumulation of mutant androgen receptor in SBMA patients. AB - Spinal and bulbar muscular atrophy (SBMA) is an inherited adult onset motor neuron disease caused by the expansion of a polyglutamine (polyQ) tract within the androgen receptor (AR), affecting only males. The characteristic pathological finding is nuclear inclusions (NIs) consisting of mutant AR with an expanded polyQ in residual motor neurons, and in certain visceral organs. We immunohistochemically examined 11 SBMA patients at autopsy with 1C2, an antibody that specifically recognizes expanded polyQ. Our study demonstrated that diffuse nuclear accumulation of mutant AR was far more frequent and extensive than NIs being distributed in a wide array of CNS nuclei, and in more visceral organs than thus far believed. Mutant AR accumulation was also present in the cytoplasm, particularly in the Golgi apparatus; nuclear or cytoplasmic predominance of accumulation was tissue specific. Furthermore, the extent of diffuse nuclear accumulation of mutant AR in motor and sensory neurons of the spinal cord was closely related to CAG repeat length. Thus, diffuse nuclear accumulation of mutant AR apparently is a cardinal pathogenetic process underlying neurological manifestations, as in SBMA transgenic mice, while cytoplasmic accumulation may also contribute to SBMA pathophysiology. PMID- 15659429 TI - Reversal of dyskinesias in an animal model of Parkinson's disease by continuous L DOPA delivery using rAAV vectors. AB - Dyskinesias are a major complication of long-term l-3,4-dihydroxyphenylalanine (L DOPA) treatment in Parkinson's disease, and are believed to result from the intermittent and pulsatile supply of L-DOPA. Daily injections of L-DOPA can prime similar abnormal involuntary movements of the limb, orolingual and axial muscles in rats rendered parkinsonian by destruction of the nigrostriatal dopamine (DA) neurons. In this study we used 33 rats with severe nigrostriatal dopamine depletion and showed that in vivo gene transfer of the DA-synthetic enzymes tyrosine hydroxylase (TH) and GTP cyclohydrolase 1 (GCH1) using recombinant adeno associated virus vectors can provide a constant source of DOPA production locally in the striatum, at a level that is effective in reducing L-DOPA-induced dyskinesias by >85%, and reverse lesion-induced motor impairments. Furthermore, the abnormal expression of DeltaFosB, prodynorphin and preproenkephalin mRNA within the striatal projection neurons normally seen in dyskinetic animals was completely reversed by TH-GCH1 gene transfer. These findings form a strong basis for replacing, or supplementing, conventional systemic L-DOPA therapy by continuous intrastriatal DOPA using in vivo gene transfer in the treatment of patients with advanced Parkinson's disease. PMID- 15659430 TI - Rice mutants and genes related to organ development, morphogenesis and physiological traits. AB - Recent advances in genomic studies and the sequenced genome information have made it possible to utilize phenotypic mutants for characterizing relevant genes at the molecular level and reveal their functions. Various mutants and strains expressing phenotypic and physiological variations provide an indispensable source for functional analysis of genes. In this review, we cover almost all of the rice mutants found to date and the variant strains that are important in developmental, physiological and agronomical studies. Mutants and genes showing defects in vegetative organs, i.e. leaf, culm and root, inflorescence reproductive organ and seeds with an embryo and endosperm are described with regards to their phenotypic and molecular characteristics. A variety of alleles detected by quantitative trait locus analysis, such as heading date, disease/insect resistance and stress tolerance, are also shown. PMID- 15659431 TI - Biological ontologies in rice databases. An introduction to the activities in Gramene and Oryzabase. AB - An enormous amount of information and materials in the field of biology has been accumulating, such as nucleotide and amino acid sequences, gene and protein functions, mutants and their phenotypes, and literature references, produced by the rapid development in this field. Effective use of the information may strongly promote biological studies, and may lead to many important findings. It is, however, time-consuming and laborious for individual researchers to collect information from individual original sites and to rearrange it for their own purpose. A concept, ontology, has been introduced in biology to support and encourage researchers to share and reuse information among biological databases. Ontology has a glossary, named dynamic controlled vocabulary, in which relationships between terms are defined. Since each term is strictly defined and identified with an ID number, a set of data represented in biological ontology is easily accessible to automated information processing, even if the data sets are across several databases and/or different organisms. In this mini-review, we introduce activities in Gramene and Oryzabase, which provide biological ontologies for Oryza sativa (rice). PMID- 15659432 TI - Genetics and evolution of inflorescence and flower development in grasses. AB - Inflorescences and flowers in the grass species have characteristic structures that are distinct from those in eudicots. Owing to the availability of genetic tools and their genome sequences, rice and maize have become model plants for the grasses and for the monocots in general. Recent studies have provided much insight into the genetic control of inflorescence and flower development in grasses, especially in rice and maize. Progress in elucidating the developmental mechanisms in each of these plants may contribute greatly to our understanding of the evolution of development in higher plants. PMID- 15659433 TI - From mapping to sequencing, post-sequencing and beyond. AB - The Rice Genome Research Program (RGP) in Japan has been collaborating with the international community in elucidating a complete high-quality sequence of the rice genome. As the pioneer in large-scale analysis of the rice genome, the RGP has successfully established the fundamental tools for genome research such as a genetic map, a yeast artificial chromosome (YAC)-based physical map, a transcript map and a phage P1 artificial chromosome (PAC)/bacterial artificial chromosome (BAC) sequence-ready physical map, which serve as common resources for genome sequencing. Among the 12 rice chromosomes, the RGP is in charge of sequencing six chromosomes covering 52% of the 390 Mb total length of the genome. The contribution of the RGP to the realization of decoding the rice genome sequence with high accuracy and deciphering the genetic information in the genome will have a great impact in understanding the biology of the rice plant that provides a major food source for almost half of the world's population. A high-quality draft sequence (phase 2) was completed in December 2002. Since then, much of the finished quality sequence (phase 3) has become available in public databases. With the completion of sequencing in December 2004, it is expected that the genome sequence would facilitate innovative research in functional and applied genomics. A map-based genome sequence is indispensable for further improvement of current rice varieties and for development of novel varieties carrying agronomically important traits such as high yield potential and tolerance to both biotic and abiotic stresses. In addition to genome sequencing, various related projects have been initiated to generate valuable resources, which could serve as indispensable tools in clarifying the structure and function of the rice genome. These resources have been made available to the scientific community through the Rice Genome Resource Center (RGRC) of the National Institute of Agrobiological Sciences (NIAS) to enable rapid progress in research that will lead to thorough understanding of the rice plant. As the next trend in rice genome research will focus on determining the function of about 40,000-50,000 genes predicted in the genome as well as applying various genomics tools in rice breeding, an unlimited access to rice DNA and seed stocks will provide a broad community of scientists with the necessary materials for formulating new concepts, developing innovative research and making new scientific discoveries in rice genomics. PMID- 15659434 TI - Molecular genetics using T-DNA in rice. AB - Now that sequencing of the rice genome is nearly completed, functional analysis of its large number of genes is the next challenge. Because rice is easy to transform, T-DNA has been used successfully to generate insertional mutant lines. Collectively, several laboratories throughout the world have established at least 200,000 T-DNA insertional lines. Some of those carry the GUS or GFP reporters for either gene or enhancer traps. Others are activation tagging lines for gain-of function mutagenesis when T-DNA is inserted in the intergenic region. A forward genetic approach showed limited success because of somaclonal variations induced during tissue culture. To utilize these resources more efficiently, tagged lines have been produced for reverse genetics approaches. DNA pools of the T-DNA-tagged lines have been prepared for polymerase chain reaction (PCR) screening of insertional mutants in a given gene. Appropriate T-DNA insertion sites are determined by sequencing the region flanking the T-DNA. This information is then used to make databases that are shared with the scientific community. International efforts on seed amplification and maintenance are needed to exploit these valuable materials efficiently. PMID- 15659435 TI - Rice plant development: from zygote to spikelet. AB - Rice is becoming a model plant in monocotyledons and a model cereal crop. For better understanding of the rice plant, it is essential to elucidate the developmental programs of the life cycle. To date, several attempts have been made in rice to categorize the developmental processes of some organs into substages. These studies are based exclusively on the morphological and anatomical viewpoints. Recent advancement in genetics and molecular biology has given us new aspects of developmental processes. In this review, we first describe the phasic development of the rice plant, and then describe in detail the developmental courses of major organs, leaf, root and spikelet, and specific organs/tissues. Also, for the facility of future studies, we propose a staging system for each organ. PMID- 15659436 TI - Suppression of tiller bud activity in tillering dwarf mutants of rice. AB - In this study, we analyzed five tillering dwarf mutants that exhibit reduction of plant stature and an increase in tiller numbers. We show that, in the mutants, axillary meristems are normally established but the suppression of tiller bud activity is weakened. The phenotypes of tillering dwarf mutants suggest that they play roles in the control of tiller bud dormancy to suppress bud activity. However, tillering dwarf mutants show the dependence of both node position and planting density on their growth, which implies that the functions of tillering dwarf genes are independent of the developmental and environmental control of bud activity. Map-based cloning of the D3 gene revealed that it encodes an F-box leucine-trich repeat (LRR) protein orthologous to Arabidopsis MAX2/ORE9. This indicates the conservation of mechanisms controlling axillary bud activity between monocots and eudicots. We suggest that tillering dwarf mutants are suitable for the study of bud activity control in rice and believe that future molecular and genetic studies using them may enable significant progress in understanding the control of tillering and shoot branching. PMID- 15659437 TI - Molecular characterization of a novel quinolizidine alkaloid O tigloyltransferase: cDNA cloning, catalytic activity of recombinant protein and expression analysis in Lupinus plants. AB - A novel acyltransferase committed to the final step of quinolizidine alkaloid biosynthesis, tigloyl-CoA:(-)-13alpha-hydroxymultiflorine/(+)-13alpha hydroxylupanine O-tigloyltransferase, has been purified from Lupinus albus. The internal amino acid sequences were determined with protease-digested fragments of 25 and 30 kDa bands, allowing design of primers for amplification of cDNA fragments by polymerase chain reaction. Using an amplified fragment as the probe, a full-length cDNA clone was isolated. Sequence analysis revealed that the cDNA encodes a protein of 453 amino acids with a molecular mass of 51.2 kDa. Phylogenetic analysis of the deduced amino acid sequences indicated that this alkaloid acyltransferase belongs to a unique subfamily of a plant acyl-CoA dependent acyltransferase gene family. The cDNA was expressed in bacterial cells as a recombinant protein fused to glutathione S-transferase. The fusion protein was affinity purified and cleaved to yield the recombinant enzyme for the study of catalytic properties. The recombinant enzyme catalyzed the acyltransfer reaction from tigloyl-CoA to (-)-13alpha-hydroxymultiflorine and (+)-13alpha hydroxylupanine. Benzoyl-CoA could also serve efficiently as an acyl donor for these hydroxylated alkaloids. RNA blot analysis suggested that the gene was expressed in roots and hypocotyls but not in cotyledons and leaves. These results indicated that this specialized acyltransferase, isolated for the first time as tigloyltransferase from nature, is committed to control the quinolizidine alkaloid patterns in a tissue-specific manner. PMID- 15659438 TI - Characterization of a member of the AN subfamily, IAN, from Ipomoea nil. AB - ANGUSTIFOLIA (AN) is the first C-terminal binding protein (CtBP) gene from plants and controls leaf width and pattern of trichome branching in Arabidopsis thaliana (L.) Heynh. We characterized an ortholog of AN from Ipomoea nil (L.) Roth (Japanese morning glory) and designated it Ipomoea nil's AN (IAN). IAN is a single-copy gene in the genome and is expressed ubiquitously in various organs of I. nil. IAN contains not only a D2-HDH motif, which is highly conserved within the CtBP family, but also LXCXE, NLS and PEST motifs, which are specific to the AN subfamily. The expression of IAN cDNA driven by the cauliflower mosaic virus 35S promoter restored a defect in leaf expansion in the leaf width direction in the angustifolia-1 (an-1) mutant of Arabidopsis, suggesting that IAN retains a common function with AN. In contrast, the complementation by IAN of a defect in the trichome branching pattern on the leaf surface of the an-1 mutant was less effective than that observed for leaf shape. These results suggest that the mechanisms by which AN regulates leaf width and trichome branching are separable. PMID- 15659439 TI - A novel vesicle derived directly from endoplasmic reticulum is involved in the transport of vacuolar storage proteins in rice endosperm. AB - We found novel vesicles derived from rough endoplasmic reticulum (ER) in rice endosperm. The novel vesicles had characteristic structures different from that of the ER-derived protein body type I and the Golgi-derived dense vesicles. Immunocytochemical analysis revealed that the novel vesicles are derived directly from the aggregates of vacuolar storage proteins in the rough ER. In addition, BiP, an ER-resident molecular chaperone, was localized in the novel vesicles, but also in protein storage vacuoles (PSVs). These results suggest that the novel vesicles mediate transport of vacuolar storage proteins directly from the ER to PSVs in rice endosperm. PMID- 15659440 TI - Involvement of extracellular dilignols in lignification during tracheary element differentiation of isolated Zinnia mesophyll cells. AB - During differentiation of isolated Zinnia mesophyll cells into tracheary elements (TEs), lignification on TEs progresses by supply of monolignols not only from TEs themselves but also from surrounding xylem parenchyma-like cells through the culture medium. However, how lignin polymerizes from the secreted monolignols has not been resolved. In this study, we analyzed phenol compounds in culture medium with reversed-phase HPLC, gas chromatography-mass spectrometry and nuclear magnetic resonance spectrometry, and found 12 phenolic compounds including coniferyl alcohol and four dilignols, i.e. erythro-guaiacylglycerol-beta coniferyl ether, threo-guaiacylglycerol-beta-coniferyl ether, dehydrodiconiferyl alcohol and pinoresinol, in the medium in which TEs were developing. Coniferyl alcohol applied to TE-inductive cultures during TE formation rapidly disappeared from the medium, and caused a sudden increase in dilignols. Addition of the dilignols promoted lignification of TEs in which monolignol biosynthesis was blocked by an inhibitor of phenylalanine anmmonia-lyase (PAL), L-alpha-aminooxy beta-phenylpropionic acid (AOPP). These results suggested that dilignols can act as intermediates of lignin polymerization. PMID- 15659441 TI - The different growth responses of the Arabidopsis thaliana leaf blade and the petiole during shade avoidance are regulated by photoreceptors and sugar. AB - During the shade-avoidance response, leaf blade expansion is inhibited and petiole elongation is enhanced. In this study, we examined the roles of photoreceptors and sugar on the differential growth of the leaf blade and petiole in shade conditions. Under the conditions examined, cell expansion, not cell division, played a major role in the differential leaf growth. The enhanced cell expansion in the leaf blade is associated with an increase in the ploidy level, whereas cell elongation was stimulated in the petiole in dark conditions without an increase in the ploidy level. Analysis of phytochrome, cryptochrome and phototropin mutants revealed that phytochromes and cryptochromes specifically regulate the contrasting growth patterns of the leaf blade and petiole in shade. Examination of the effects of photo-assimilated sucrose on the growth of the leaf blade and petiole revealed growth-promotional effects of sucrose that are highly dependent on the light conditions. The leaf blades of abscisic acid-deficient and sugar-insensitive mutants did not expand in blue light, but expanded normally in red light. These results suggest that both the regulation of light signals and the modulation of responses to sugar are important in the control of the differential photomorphogenesis of the leaf blade and petiole. PMID- 15659442 TI - Inhibition of contractile vacuole function by brefeldin A. AB - Brefeldin A (BFA) causes a block in the secretory system of eukaryotic cells. In the scaly green flagellate Scherffelia dubia, BFA also interfered with the function of the contractile vacuoles (CVs). The CV is an osmoregulatory organelle which periodically expels fluid from the cell in many freshwater protists. Fusion of the CV membrane with the plasma membrane is apparently blocked by BFA in S. dubia. The two CVs of S. dubia swell and finally form large central vacuoles (LCVs). BFA-induced formation of LCVs depends on V-ATPase activity, and can be reversed by hypertonic media, suggesting that water accumulation in the LCVs is driven by osmosis. We suggest that the BFA-induced formation of LCVs represents a prolonged diastole phase. A normal diastole phase takes about 20 s and is difficult to investigate. Therefore, BFA-induced formation of LCVs in S. dubia represents a unique model system to investigate the diastole phase of the CV cycle. PMID- 15659443 TI - Differential expression of AtXTH17, AtXTH18, AtXTH19 and AtXTH20 genes in Arabidopsis roots. Physiological roles in specification in cell wall construction. AB - Xyloglucan endotransglucosylase/hydrolases (XTHs) are a class of enzymes that are capable of splitting and reconnecting xyloglucan molecules, and are implicated in the construction and restructuring of the cellulose/xyloglucan framework. Thirty three members of the XTH gene family are found in the genome of Arabidopsis thaliana, but their roles remain unclear. Here, we describe the tissue-specific and growth stage-dependent expression profiles of promoter::GUS fusion constructs for four Arabidopsis XTH genes, AtXTH17, AtXTH18, AtXTH19 and AtXTH20, which are phylogenetically closely related to one another. AtXTH17 and AtXTH18 were expressed in all cell types in the elongating and differentiating region of the root, while AtXTH19 was expressed in the apical dividing and elongating regions, as well as in the differentiation zone, and was up-regulated by auxin. In contrast, AtXTH20 was expressed specifically in vascular tissues in the basal mature region of the root. This expression analysis also disclosed cis-regulatory sequences that are conserved among the four genes, and are responsible for the root-specific expression profile. These results indicate that the four XTH genes, which were generated by gene duplication, have diversified their expression profile within the root in such a way as to take responsibility for particular physiological roles in the cell wall dynamics. PMID- 15659444 TI - Light-induced wilting and its molecular mechanism in epicotyls of dark-germinated pea (Pisum sativum L.) seedlings. AB - Possible mechanisms behind the light-induced wilting of dark-germinated pea (Pisum sativum L.) epicotyls were studied. Illumination with photosynthetically active radiation caused a fast turgor loss and wilting in the middle segments of the epicotyls accompanied by accumulation of water in the intercellular cavities. During this process, room temperature fluorescence emission spectra showed gradual bleaching of porphyrin-type pigments, which was lessened by incubating the epicotyls with excess ascorbate before illumination. Detection of singlet oxygen and lipid peroxidation products in the illuminated epicotyls suggested the occurrence of porphyrin-photosenzitized membrane damage as a cause of disordered water status and sequential wilting. PMID- 15659445 TI - Phosphate starvation induces a determinate developmental program in the roots of Arabidopsis thaliana. AB - When growing under limiting phosphate (P) conditions, Arabidopsis thaliana plants show dramatic changes in root architecture, including a reduction in primary root length, increased formation of lateral roots and greater formation of root hairs. Here we report that primary root growth inhibition by low P is caused by a shift from an indeterminate to a determinate developmental program. In the primary root, the low P-induced determinate growth program initiates with a reduction of cell elongation followed by the progressive loss of meristematic cells. At later stages, cell proliferation ceases and cell differentiation takes place at the former cell elongation and meristematic regions of the primary root. In low P, not only the primary but also almost all mature lateral roots enter the determinate developmental program. Kinetic studies of expression of the cell cycle marker CycB1;1:uidA and the quiescent center (QC) identity marker QC46:GUS showed that in low P conditions, reduction in proliferation in the primary root was preceded by alterations in the QC. These results suggest that in Arabidopsis, P limitation can induce a determinate root developmental program that plays an important role in altering root system architecture and that the QC could act as a sensor of environmental signals. PMID- 15659447 TI - Cell cycle dependence of elicitor-induced signal transduction in tobacco BY-2 cells. AB - The molecular links between the cell cycle and defense responses in plants are largely unknown. Using synchronized tobacco BY-2 cells, we analyzed the cell cycle dependence of elicitor-induced defense responses. In synchronized cultured apoaequorin-expressing cells, the increase in cytosolic free Ca2+ induced by a proteinaceous elicitor, cryptogein, was greatly suppressed during the G2 and M phases in comparison with G1 or S phases. Treatment with cryptogein during the G1 or S phases also induced biphasic (rapid/transient and slow/prolonged) responses in activation of mitogen-activated protein kinases (MAPKs) and production of reactive oxygen species (ROS). In contrast, elicitor treatment during the G2 or M phases induced only a rapid and transient phase of MAPK activation and ROS production. Their slow and prolonged phases as well as expression of defense related genes, cell cycle arrest and cell death were induced only after the cell cycle progressed to the G1 phase; removal of the elicitor before the start of the G1 phase inhibited these responses. These results suggest that although cryptogein recognition occurred at all phases of the cell cycle, the recognition during the S or G1 phases, but not at the G2 or M phases, induces the prolonged activation of MAPKs and the prolonged production of ROS, followed by cell cycle arrest, accumulation of defense-related gene transcripts and cell death. Elicitor signal transduction depends on the cell cycle and is regulated differently at each phase. PMID- 15659446 TI - Involvement of the chloroplast signal recognition particle cpSRP43 in acclimation to conditions promoting photooxidative stress in Arabidopsis. AB - In this study, we have investigated the role of the CAO gene (coding for the chloroplast recognition particle cpSRP43) in the protection against and acclimation to environmental conditions that promote photooxidative stress. Deficiency of cpSRP43 in the Arabidopsis mutant chaos has been shown previously to lead to partial loss of a number of proteins of the photosystem II (PSII) antennae. In addition, as reported here, mutant plants have lower growth rates and reduced lignin contents under laboratory conditions. However, chaos seedlings showed significantly higher tolerance to photooxidative stress under both tightly controlled laboratory conditions and highly variable conditions in the field. This greater tolerance of chaos plants was manifested in less photooxidative damage together with faster growth recovery in young seedlings. It was also associated with a lower production of H2O2, lower ascorbate levels and less induction of ascorbate peroxidases. Under field conditions, chaos exhibited better overall photosynthetic performance and had higher survival rates. Expression of the CAO gene may be regulated by a light-dependent chloroplastic redox signalling pathway, and was inhibited during acclimation to high light and chilling temperatures, simultaneously with induction of ascorbate peroxidases. It is concluded that the presence/absence of the CAO gene has an impact on photo produced H2O2, lignification in the hypocotyls and on the plant's susceptibility to photooxidative stress. Therefore, regulation of the CAO gene may be part of the plant's system for acclimation to high light and chilling temperatures. PMID- 15659448 TI - Maize cyclin D2 expression, associated kinase activity and effect of phytohormones during germination. AB - A cDNA corresponding to 16 kDa of the maize cyclin D2 N-terminus was cloned and this polypeptide was overexpressed to produce homologous antibodies. This antibody recognized a 38 kDa protein in extracts from maize embryonic axes which corresponds to the predicted size for cyclin D2 protein. Expression of cyclin D2 was followed at the transcriptional and protein levels, and the effect of cytokinins and abscisic acid (ABA) was followed during maize germination. Cytokinins importantly stimulated cyclin D2 gene expression at late germination times and sucrose was necessary for stimulation, whereas the effect of ABA was not different from that in controls. However, cyclin D2 protein levels in control axes reached a peak at 6 h germination, declining thereafter, and neither cytokinins nor ABA modified this behavior. Two cyclic-dependent kinase A (Cdk-A) type proteins and proliferating cell nuclear antigen (PCNA) were found co immunoprecipitating with cyclin D2, and these immunoprecipitates were able to phosphorylate both histone H1 and the maize retinoblastoma-related protein (RBR). This protein kinase activity differed from the pattern of protein accumulation during germination, and the activity was not modified by either cytokinins or ABA. We discuss these findings in terms of the importance of the cell cycle for the germination process. PMID- 15659449 TI - Abundant expression in vascular tissue of plant TAF10, an orthologous gene for TATA box-binding protein-associated factor 10, in Flaveria trinervia and abnormal morphology of Arabidopsis thaliana transformants on its overexpression. AB - TAF10 is one of the TATA box-binding protein-associated factors (TAFs), which constitute the TFIID complex. We isolated a plant TAF10 ortholog from a Flaveria trinervia cDNA library, and named it ftTAF10. The ftTAF10 polypeptide contains a histone-fold motif, which is highly conserved among the TAF10s of other organisms. A transiently expressed green fluorescent protein (GFP) fusion protein was translocated into the nuclei of onion epidermal cells, suggesting that the ftTAF10 functions in nuclei. The transcript level was higher in stems and roots than in leaves, and in situ hybridization of F. trinervia seedlings revealed that the ftTAF10 transcript is accumulated abundantly in vascular tissues of hypocotyls, in the central cylinder of roots, and slightly in bundle sheath cells of leaves. Overexpression of ftTAF10 in Arabidopsis under the cauliflower mosaic virus 35S promoter caused two kinds of abnormal morphology, limitation of the indeterminate inflorescence and production of deformed leaves. These results indicate the possibility that ftTAF10 is a plant 'selective TAF' involved in the expression of a subset of vascular abundant genes, and that its appropriate gene expression is necessary for normal development. PMID- 15659450 TI - Possible involvement of mechanosensitive Ca2+ channels of plasma membrane in mechanoperception in Chara. AB - When an internodal cell of Chara corallina was stimulated with a mechanical pulse of various amplitudes lasting for 0.1 s (mechanical stimulus), the cell generated a receptor potential, which was highly dependent not only on the strength of the stimulus but also on the extracellular Cl- concentration. Extracellular Ca2+ was indispensable for generating receptor potential, since removal of Ca2+ reversibly inhibited generation of the receptor potential. The cytoplasmic Ca2+ level transiently rose upon mechanical stimulation. The stronger the mechanical stimulus, the larger was the increase in the cytoplasmic level of Ca2+. It is proposed that the first step of receptor potential is an activation of mechanosensitive Ca2+ channels at the plasma membrane. PMID- 15659451 TI - Spectroscopic analysis of the dark relaxation process of a photocycle in a sensor of blue light using FAD (BLUF) protein Slr1694 of the cyanobacterium Synechocystis sp. PCC6803. AB - Slr1694 is a BLUF (sensor of blue light using flavin adenine dinucleotide) protein and a putative photoreceptor in the cyanobacterium Synechocystis sp. PCC6803. Illumination of Slr1694 induced a signaling light state concurrent with a red shift in the UV-visible absorption of flavin, and formation of the bands from flavin and apo-protein in the light-minus-dark Fourier transform infrared (FTIR) difference spectrum. Replacement of Tyr8 with phenylalanine abolished these changes. The light state relaxed to the ground dark state, during which the FTIR bands decayed monophasically. These bands were classifiable into three groups according to their decay rates. The C4=O stretching bands of a flavin isoalloxazine ring had the highest decay rate, which corresponded to that of the absorption red shift. The result indicated that the hydrogen bonding at C4=O is responsible for the UV-visible red shift, consistent with the results of density functional calculation. All FTIR bands and the red shift decayed at the same slower rate in deuterated Slr1694. These results indicated that the dark relaxation from the light state is limited by proton transfer. In contrast, a constrained light state formed under dehydrated conditions decayed much more slowly with no deuteration effects. A photocycle mechanism involving the proton transfer was proposed. PMID- 15659452 TI - The rice nucellin gene ortholog OsAsp1 encodes an active aspartic protease without a plant-specific insert and is strongly expressed in early embryo. AB - The barley nucellin gene was reported to be nucellus specific in its expression and was hypothesized to play a role in the programmed cell death of the nucellus as an aspartic protease. Here we provide direct evidence that the rice ortholog encodes an active aspartic protease, but we prefer the name aspartic protease1 (OsAsp1) to nucellin after a detailed analysis of its expression pattern in rice and barley. Northern blots, RT-PCR and RNA in situ hybridization showed that OsAsp1 is expressed most abundantly in zygotic embryos 1-2 d after fertilization. It is also expressed in pollen, nucellus, ovary wall, shoot and root meristem, coleoptiles of immature seeds, and somatic embryos. A parallel study in barley showed that the barley nucellin gene was expressed not only in the nucellus but also strongly in embryos. Recombinant protein proOsAsp1 expressed in the bacterium Escherichia coli refolded and autolysed at acidic pH 3.5 in vitro, and the mature peptide displayed protease activity. Nucellin has three close homologs in rice on chromosomes 11 and 12 and in Arabidopsis on chromosomes 1 and 4. They lack the plant-specific insert that distinguishes the typical plant aspartic protease from aspartic proteases of other organisms. They constitute a new class of aspartic protease that is present in both monocots and dicots but whose function remains to be explored further. PMID- 15659453 TI - The value of health care information exchange and interoperability. AB - In this paper we assess the value of electronic health care information exchange and interoperability (HIEI) between providers (hospitals and medical group practices) and independent laboratories, radiology centers, pharmacies, payers, public health departments, and other providers. We have created an HIEI taxonomy and combined published evidence with expert opinion in a cost-benefit model. Fully standardized HIEI could yield a net value of dollar 77.8 billion per year once fully implemented. Nonstandardized HIEI offers smaller positive financial returns. The clinical impact of HIEI for which quantitative estimates cannot yet be made would likely add further value. A compelling business case exists for national implementation of fully standardized HIEI. PMID- 15659454 TI - Interoperability: the key to the future health care system. AB - The United States is building a point-of-care health information system to rival the worldwide network of electronic banking. Through health care information exchange and interoperability, clinicians will have access to a longitudinal medical record. This interoperability is a fundamental requirement for the health care system to derive the societal benefits promised by the adoption of electronic medical records (EMRs). The paper by Jan Walker and colleagues highlights some of these benefits. One critical question is whether the adoption of EMRs needs to wait for interoperability standards or whether it can proceed efficiently without them. PMID- 15659455 TI - Benefits of interoperability: a closer look at the estimates. AB - The paper by Jan Walker and colleagues provides an estimate of savings to be gained by increased health care information exchange and interoperability (HIEI). However, the assumptions on which their analysis was based seem very optimistic and could produce estimates that are not achievable. This commentary outlines some questions about their assumptions and suggests that less-aggressive assumptions could lead to more realistic expectations about the financial implications of achieving interoperability. PMID- 15659456 TI - E-health: steps on the road to interoperability. AB - Interoperable electronic medical records (EMRs) have the potential to produce better health outcomes while improving the efficiency of care delivery and reducing its costs. Implementation will require massive changes at all levels. In many instances, the costs of implementation could fall on one group, while savings will accrue to some other group. A successful transition strategy identifies a series of steps, where each step pays its own way, at the level of the local groups directly affected, and lays the foundation for the next step. Such a strategy implies an era in which large groups will likely play a critical role. PMID- 15659457 TI - Commentary: Using geographical data to monitor socioeconomic inequalities in mortality: experiences from Japanese studies. PMID- 15659458 TI - Commentary: Contrary findings from Guinea-Bissau and Papua New Guinea. PMID- 15659459 TI - Chronic hepatitis B virus infection and mortality from non-liver causes: results from the Haimen City cohort study. AB - BACKGROUND: Studies of chronic hepatitis B virus (HBV) infection in endemic populations have rarely documented causes of mortality other than liver disease and hepatocellular carcinoma (HCC). METHODS: We analysed all-cause mortality related to HBV infection, focusing on the deaths not related to liver disease in a prospective cohort of adults living in Haimen City, China, who were followed from 1992 to 2002. Death certificate data from 4590 deaths among 83 794 individuals were analysed. At cohort entry, 15.0% of the 58 454 male subjects and 10.7% of 25 340 female subjects were hepatitis B surface antigen positive [HbsAg(+)]. HCC and chronic liver disease were the major causes of death in both men and women in this population. The analysis focused on non-liver causes of death. RESULTS: When liver-related causes of death were excluded, there was still a significantly higher age-adjusted death rate among HBsAg(+) individuals. The relative risks (RRs) and 95% confidence intervals (CIs) for all non-liver deaths among HBsAg(+) subjects were 1.2 (1.1-1.3) in men and 1.4 (1.1-1.7) in women. Non liver causes were further subdivided into cancer and non-cancer groups. For all non-liver cancers, the RR was 1.2 (1.0-1.4) for males and 1.7 (1.2-2.3) for females. Non-liver, non-cancer deaths had RRs of 1.2 (1.1-1.4) and 1.2 (0.9-1.6) in males and females, respectively. CONCLUSIONS: HBV-infected individuals may be at increased mortality risk from non-liver causes. Possible reasons include a direct effect of HBV infection, changes in the host immune system as a cause or effect of chronic infection, and behavioural factors associated with HBV infection. Further studies are needed to confirm this finding. PMID- 15659460 TI - Educational inequalities in the metabolic syndrome and coronary heart disease among middle-aged men and women. AB - BACKGROUND: Previous studies have shown socioeconomic inequalities in the metabolic syndrome and coronary heart disease (CHD), but it is not known whether educational disparities in the metabolic syndrome explain educational inequalities in CHD. We investigated this question in a prospective study of middle-aged men and women. METHODS: Baseline data were collected in 1992 in Finland from 864 men and 1045 women aged 45-64 years without history of CHD. A total of 113 new CHD cases were identified by the end of 2001. Logistic and Cox regression models were used in data analysis. RESULTS: The metabolic syndrome defined by NCEP criteria was less prevalent in subjects with university education (21% in men and 14% in women) compared with basic level education (41% and 27%, respectively). Adjusting for health behavioural factors had only a slight effect on the educational gradient in the metabolic syndrome. An educational gradient in CHD incidence was clear [hazard ratio (HR) = 0.67 95% confidence interval (CI) 0.48-0.94, men and women combined]. Adjustment for the metabolic syndrome attenuated this gradient only slightly, but when individual components of the metabolic syndrome were included as covariates the attenuation was more substantial (HR = 0.73 95% CI 0.52-1.04). CONCLUSIONS: Educational differences in the metabolic syndrome and CHD incidence are clear. Metabolic risk factors explain the gradient in CHD incidence partly, but only when they are treated as independent risk factors. Screening for the metabolic syndrome alone is not sufficient to account for socioeconomic inequalities in cardiovascular disease. PMID- 15659461 TI - Commentary: is tooth loss good or bad for general health? PMID- 15659462 TI - Commentary: liver cancer and the epidemiological and cancer transition theories. PMID- 15659463 TI - Migration as a risk factor for measles after a mass vaccination campaign, Burkina Faso, 2002. AB - BACKGROUND: Shortly after a measles supplementary immunization activity (SIA) targeting children from 9 months to 14 years of age that achieved high coverage, Burkina Faso had a large, serologically confirmed measles outbreak. To investigate the causes of this first reported failure of a widely successful measles control strategy we conducted a case-control study. METHODS: Serologically confirmed measles cases aged > or =9 months at the time of the SIA in 6 heavily affected districts were frequency matched on age to 3 controls recruited from people frequenting health centres in the same districts. RESULTS: Between January and July 2002, 1287 measles cases were reported throughout Burkina Faso. Of the 707 cases that were serologically confirmed, 358 (51%) were from 9 months to 14 years of age and 265 (37%) were > or =15 years of age. Among cases and controls from 9 months to 14 years of age significant risk factors for measles were lack of measles vaccination and, in the unvaccinated, recent travel to Cote d'Ivoire. Of the recent measles cases in Cote d'Ivoire 54% were there when exposed to measles. Among adults, risk factors included non-vaccination and the lack of school attendance during childhood. Vaccine effectiveness was estimated to be 98%. CONCLUSIONS: Migration of children between Cote d'Ivoire and Burkina Faso played a major role in the failure of the SIA to interrupt measles transmission. Synchronization of measles control activities should be a high priority in countries with regions where much migration occurs. PMID- 15659464 TI - Prevalence of diarrhoea in the community in Australia, Canada, Ireland, and the United States. AB - BACKGROUND: Studies in several countries have estimated the prevalence of diarrhoea in the community. However, the use of different study designs and varying case definitions has made international comparisons difficult. METHODS: Similar cross-sectional telephone surveys were conducted in Australia, Canada, Ireland (including Northern Ireland), and the United States over 12 month periods between 2000 and 2002. Each survey asked about diarrhoea in the four weeks before the interview. For this comparative analysis, uniform definitions were used. RESULTS: Questionnaires were completed for 6087 respondents in Australia, 3496 in Canada, 9903 in Ireland, and 14,647 in the United States. In the four weeks prior to interview, at least one episode of diarrhoea was reported by 7.6% of respondents in Canada, 7.6% in the United States, 6.4% in Australia, and 3.4% in Ireland. The prevalence of diarrhoea was consistently higher in females. In all countries, the prevalence of diarrhoea was highest in children <5 years and lowest in persons > or =65 years of age. When diarrhoea and vomiting was considered, the prevalence was almost identical in the four studies (range: 2.0 2.6%). Despite different health care structures, a similar proportion of respondents sought medical care (approximately one in five). Antibiotic usage for the treatment of diarrhoea was reported by 8.3% of respondents in the United States, 5.6% in Ireland, 3.8% in Canada, and 3.6% in Australia. CONCLUSIONS: Diarrhoea is a common illness among persons in the community in Australia, Canada, Ireland, and the United States. With similar methodologies and a standard case definition, age and sex patterns and health care seeking behaviour were remarkably consistent between countries. PMID- 15659465 TI - Commentary: on form, comparability, and levels in the income and health relationship. PMID- 15659466 TI - Risks for Ross River virus disease in tropical Australia. AB - BACKGROUND: There are no analytical studies of individual risks for Ross River virus (RRV) disease. Therefore, we set out to determine individual risk and protective factors for RRV disease in a high incidence area and to assess the utility of the case-control design applied for this purpose to an arbovirus disease. METHODS: We used a prospective matched case-control study of new community cases of RRV disease in the local government areas of Cairns, Mareeba, Douglas, and Atherton, in tropical Queensland, from January 1 to May 31, 1998. RESULTS: Protective measures against mosquitoes reduced the risk for disease. Mosquito coils, repellents, and citronella candles each decreased risk by at least 2-fold, with a dose-response for the number of protective measures used. Light-coloured clothing decreased risk 3-fold. Camping increased the risk 8-fold. CONCLUSIONS: These risks were substantial and statistically significant, and provide a basis for educational programs on individual protection against RRV disease in Australia. Our study demonstrates the utility of the case-control method for investigating arbovirus risks. Such a risk analysis has not been done before for RRV infection, and is infrequently reported for other arbovirus infections. PMID- 15659467 TI - Prediction of coronary events in a low incidence population. Assessing accuracy of the CUORE Cohort Study prediction equation. AB - BACKGROUND: The aims of this paper are to derive a 10-year coronary risk predictive equation for adult Italian men, and to assess its accuracy in comparison with the Framingham Heart Study (FHS) and PROCAM study equations. METHODS: The CUORE study is a prospective fixed-cohort study. Eleven cohorts, from the north and the centre-south of Italy, had been investigated at baseline between 1982 and 1996, adopting MONICA methods to measure risk factors. Among this sample of 6865 men, aged 35-69 years and free of coronary heart disease (CHD) at baseline, 312 first fatal and non-fatal major coronary events occurred in 9.1 years median follow-up. Calibration, as the difference between 10-year predicted and actual risk, and discrimination, as the ability of the risk functions to separate high-risk from low-risk subjects, have been assessed to compare accuracy of the FHS, the PROCAM, and the CUORE study equations. RESULTS: The best CUORE equation includes age, total cholesterol, systolic blood pressure, cigarette smoking, HDL-cholesterol, diabetes mellitus, hypertension drug treatment, and family history of CHD (area under the ROC curve = 0.75). The uncalibrated estimates of the 10-year risk in this CUORE follow-up data were 0.093 and 0.109 higher (P < 0.05) from the Framingham and PROCAM risk scores, respectively, than the Kaplan-Meier estimate for CUORE, indicating risk overestimates for both equations. Standard recalibration techniques improved accuracy of the FHS equation only. PROCAM overestimates were prominent in the higher risk deciles. With an alternative method for recalibration better risk estimates were obtained, but a cohort study was needed to obtain a properly calibrated risk equation. CONCLUSIONS: The CUORE Project predictive equation showed better accuracy of the FHS and PROCAM equations, overcoming frequently reported risk overestimates. The CUORE equation may be adopted to identify men with high coronary risk in Italy. PMID- 15659468 TI - Prenatal risk factors for Caesarean section. Analyses of the ALSPAC cohort of 12,944 women in England. AB - BACKGROUND: There has been an escalation in Caesarean section rates globally. Numerous prenatal factors have been associated with elective and emergency Caesarean section, some of which may be amenable to change. METHODS: A population based cohort of 12,944 singleton, liveborn, term pregnancies were used to investigate risk factors for Caesarean section using multivariable logistic regression modelling. Numerous prenatal factors were investigated for their associations with the following outcomes: first, with Caesarean section (both elective and emergency) compared with vaginal delivery (spontaneous and assisted); second, for their associations with elective Caesarean section compared with attempted vaginal delivery; and finally emergency Caesarean section compared with spontaneous vaginal delivery. RESULTS: 11,791 women had vaginal delivery and 1153 had Caesarean section (685 emergency, 468 elective). Non cephalic (breech) presentation (all Caesareans odds ratio (OR) 36.6, 95% confidence interval (CI) 26.8-50.0; elective Caesarean OR 86.4, 95% CI 58.5 127.8; emergency Caesarean OR 9.58, 95% CI 6.06-15.1) and previous Caesarean section (all Caesareans OR 27.8, 95% CI 20.9-37.0, elective Caesarean OR 54.4, 95% CI 38.4-77.5; emergency Caesarean OR 13.0, 95% CI 7.76-21.7) were associated in all analyses with an increased risk of Caesarean section. Extremes of neonatal birthweight were associated with an increased risk of Caesarean section (all Caesareans and emergency section) compared with vaginal delivery as was increasing neonatal head circumferences. In all analyses increasing maternal age (OR 1.07 per year, 95 % CI 1.04-1.09; OR 1.04 per year, 95 % CI 1.01-1.08; OR 1.11 per year, 95% CI 1.08-1.15) was independently associated with increased odds of Caesarean section. Increasing parity was associated with a decrease in risk for all Caesareans and emergency section (OR 0.63, 95% CI 0.53-0.75 and OR 0.46, 95% CI 0.33-0.63, respectively), as was the outcome of the last pregnancy being a live child. Increasing gestation was independently associated with a decreased risk of both all Caesareans and elective Caesarean (OR 0.86, 95% CI 0.80-0.93 and OR 0.52, 95% CI 0.46-0.58 respectively), whereas diabetes mellitus was associated with increased risk. These variables were not associated with emergency section. However, epidural use was associated with an increased risk of emergency Caesarean (OR 6.49, 95% CI 4.78-8.82) while being in a preferred labour position decreased the risk (OR 0.59, 95% CI 0.49-0.73). CONCLUSIONS: A careful exploration of risk factors may allow us to identify reasons for the increasing rates of Caesarean section and the marked variation between institutions. PMID- 15659470 TI - Polyparasitism. PMID- 15659471 TI - The epidemiology and prevention of suicide by hanging: a systematic review. AB - BACKGROUND: Hanging is one of the most commonly used methods for suicide worldwide. In England and a number of other countries, its incidence has increased over the last 30 years. This review summarizes the published literature on suicide by hanging. The focus is on its epidemiology in England and on identifying potential means of prevention. METHODS: We searched Medline (1966 2003), Embase (1980-2003), CINAHL (1982-2003) and PsycINFO (1967-2003). As considerable research on suicides occurring in prisons and psychiatric hospitals in England and Wales has been carried out by the National Confidential Inquiry into Suicide and Homicide (Manchester) and the Prison Service's Safer Custody Group, we obtained additional information from these sources. RESULTS: Only a small proportion (around 10%) of hanging suicides occur in the controlled environments of hospitals, prisons, and police custody; the remainder occur in the community. The most commonly used ligatures (rope, belts, flex) and ligature points (beams, banisters, hooks, door knobs, and trees) are widely available; thus prevention strategies focused around restriction of access to means of hanging are of limited value. Around 50% of hanging suicides are not fully suspended--ligature points below head level are commonly used. Case fatality following attempted suicide by hanging is around 70%; the majority (80-90%) of those who reach hospital alive survive. CONCLUSION: Strategies to reduce suicide by hanging should focus on the prevention of suicide in controlled environments, the emergency management of 'near-hanging' and on the primary prevention of suicide in general. More research is required to better understand the recent rise in popularity of this method. PMID- 15659472 TI - Religious attendance as a predictor of survival in the EPESE cohorts. AB - BACKGROUND: Interest has arisen in recent years in the relationship between religious involvement and health outcomes. Although most of the early literature consists of studies with methodological flaws, some recent well-conducted reports show that religious attendance is associated with reduced mortality in selected subgroups and populations. METHODS: In this study, we investigated the relationship between religious attendance and mortality using the 14,456 participants in the National Institute of Aging-funded 'Established Populations for Epidemiologic Studies of the Elderly'. RESULTS: Our analyses show that after controlling for important prognostic factors, frequent religious attendance was associated with increased survival in the entire cohort [risk ratio (RR) = 0.78, 95% Confidence interval (CI) 0.70-0.88]. However, stratified analyses show that this association exists for only two of the four sites. CONCLUSIONS: We conclude that the association between religious attendance and survival is not robust and may depend upon unknown confounders and covariates. PMID- 15659473 TI - The contribution of childhood and adult socioeconomic position to adult obesity and smoking behaviour: an international comparison. AB - BACKGROUND: Our objective was to investigate the contribution of childhood and adult socioeconomic position (SEP) to adult obesity and smoking behaviour, in particular to establish the role of childhood circumstances across different studies in Europe and the US. METHODS: Seven population-based surveys in six Western countries (Britain, Denmark, Finland, Netherlands, Sweden, US) were examined, with participants aged 30-50 yr and born between 1910 and 1960. Adult smoking was analysed using three outcomes (ever, current, or ex-) and adult obesity was defined as body mass index (kg/m(2)) > or =30. RESULTS: A strong effect of adult social position was observed for smoking outcomes and obesity. For example, manual SEP in adulthood increased the risk of ever smoking (adjusted odds ratio (OR) 1.47-2.00 for men; 0.94-1.81 for women), and obesity (adjusted OR 1.06-2.24 for men, 1.21-3.26 for women). In most studies, childhood position was not associated with ever-smoking. For current smoking, manual childhood position was associated among women (adjusted OR 1.09-1.54), but no consistent pattern was seen for men. For ex-smoking, manual childhood origins lowered the chance of quitting among women (adjusted OR 0.64-0.81) except in the US (OR = 1.17); among men this association was seen in fewer studies (adjusted OR 0.74-1.09). For obesity, manual origins increased the risk for women (adjusted OR 0.96-2.50); effects were weaker among men but mostly in the same direction (adjusted OR 0.79 1.42). CONCLUSIONS: As expected, adult SEP was an important influence on smoking behaviour and obesity. In addition, factors related to disadvantaged social origins appeared to increase the risk of obesity and reduce the probability of quitting smoking in adulthood, particularly in women. PMID- 15659474 TI - BCG vaccination scar associated with better childhood survival in Guinea-Bissau. AB - BACKGROUND: Recent studies have suggested that Bacille Calmette-Guerin (BCG) vaccination may have a non-specific beneficial effect on infant survival and that a BCG scar may be associated with lower child mortality. No study has previously examined the influence of BCG vaccination on cause of death. METHODS: Two cohorts (A and B) were used to describe the mortality pattern for children with and without BCG scar and to determine specific causes of death. In cohort A (n = 1813), BCG scar was assessed at 6 months of age and as previously described children with a BCG scar had lower mortality over the next 12 months than children with no BCG scar. In cohort B, 1617 children aged 3 months to 5 years of age had their BCG scar status assessed in a household-based survey and mortality was assessed during a 12-month period. Causes of death were determined by verbal autopsy (VA) and related to BCG scar status in a cause-specific hazard function. RESULTS: Controlling for background factors associated with mortality, there was lower mortality for children with a BCG scar than without in cohort B, the mortality ratio (MR) being 0.45 (95% CI 0.21-0.96). Exclusion of children exposed to TB did not have any impact on the result. In a combined analysis of cohorts A and B, the MR was 0.43 (95% CI 0.28-0.65) controlling for background factors. There were no large differences in distribution of the five major causes of death (malaria, pneumonia, acute diarrhoea, chronic diarrhoea, and meningitis/encephalitis) according to BCG scar status in the two cohorts. Having a BCG scar significantly reduced the risk of death from malaria [MR 0.32 (95% CI 0.13-0.76)]. CONCLUSIONS: A BCG scar is a marker of better survival among children in countries with high child mortality. BCG vaccination may affect the response to several major infections including malaria. PMID- 15659475 TI - Socioeconomic status and health: is parasympathetic nervous system activity an intervening mechanism? AB - BACKGROUND: The link between socioeconomic status (SES) and health is widely recognized but the pathophysiologic mechanisms are not well understood. We tested the hypothesis that parasympathetic nervous system (PNS) regulation is one such mechanism. METHODS: In a cross-sectional study, electrocardiogram-derived RR interval variability (RRV), a non-invasive index of cardiac PNS regulation, and SES, measured as educational attainment and income, were collected in 756 subjects in the CARDIA study of heart disease in young adults. RESULTS: Relative to those with less than a high school education, those with high school to college and post-college education had 26% (beta = 0.233) and 43% (beta = 0.355) greater low frequency (LF) RRV, respectively, adjusted for age, sex, and race. For high frequency (HF) RRV, race interacted with income: relative to low income whites, intermediate and high income whites had 133 and 191% greater HF power, respectively, while intermediate and high income blacks had 32 and 44% greater HF RRV, respectively, relative to low income blacks. CONCLUSIONS: Numerous studies demonstrate that psychosocial stressors reduce cardiac parasympathetic regulation and that SES disparities are associated with increasing social stress proportional to the degree of disparity. Data from the current study suggest that PNS regulation may be a mechanism linking the stressful effects of low SES to increased morbidity and mortality. PMID- 15659476 TI - Tooth loss is associated with increased risk of total death and death from upper gastrointestinal cancer, heart disease, and stroke in a Chinese population-based cohort. AB - BACKGROUND: Tooth loss has previously been associated with a higher risk of cancer, heart disease, and stroke, but the role of confounding by smoking remains an issue. METHODS: We conducted a cohort study including 29,584 healthy, rural Chinese adults who were participants in a chemoprevention trial from 1986 through 1991 and who have been followed-up through 2001. We categorized tooth loss for each subject as less than or equal to or greater than the median number of teeth lost for other subjects of the same age at baseline. Mortality outcomes were categorized as follows: total death (n = 9362), upper gastrointestinal (GI) cancer death (n = 2625), other cancer death (n = 514), heart disease death (n = 1932), and fatal stroke (n = 2866). RESULTS: Individuals with greater than the age-specific median number of teeth lost had statistically significant 13% increased risk of total death [95% confidence interval (CI) 9-18%], 35% increased risk of upper GI cancer death (95% CI 14-59%), 28% increased risk of heart disease death (95% CI 17-40%), and 12% increased risk of stroke death (95% CI 2 23%), but no significantly increased risk of death from cancer at other sites. These elevated risks were present in male smokers, male non-smokers, and females, nearly all never-smokers. CONCLUSIONS: In this Asian population, tooth loss significantly increased the risk of total death and death from upper GI cancer, heart disease, and stroke. These associations were not limited to tobacco smokers. PMID- 15659477 TI - Area deprivation, social class, and quality of life among people aged 75 years and over in Britain. AB - BACKGROUND: There is a shortage of research studies that assess how selected characteristics of neighbourhood and personal social circumstances contribute towards health-related quality of life (QoL) among older people. METHODS: Analysis of baseline data for 5581 people aged > or =75 years and over from the Trial of Assessment and Management of Older People in the Community. The scores for four dimensions from the UK version of the Sickness Impact Profile and for the Philadelphia Geriatric Morale Scale were analysed in relation to individual social class and the Carstairs score of socioeconomic deprivation for the enumeration district of residence. RESULTS: In age and sex adjusted analyses, the proportion of participants of social class IV/V living in the most deprived areas who were in the quintile with worst QoL scores was more than double that among those from social class I/II living in the least deprived areas. Individual social class and area deprivation score contributed roughly equally to this doubling for home management, self-care and social interaction, whereas social class appeared a stronger determinant for mobility. Adjustment for living circumstances, health symptoms, and health behaviours substantially reduced the excess risk associated with social class and area deprivation. Being in a rural area was associated with lower risk of poor morale. CONCLUSION: Poor socioeconomic characteristics of both the area and the individual are associated with worse functioning (QoL) of older people in the community. This is not fully explained by health status. Policy should consider community-level interventions as well as those directed at individuals. PMID- 15659479 TI - A simple hint to improve Robinson and Dickersin's highly sensitive PubMed search strategy for controlled clinical trials. PMID- 15659480 TI - Use of inhaled corticosteroids during pregnancy and risk of pregnancy induced hypertension: nested case-control study. AB - OBJECTIVE: To determine whether the use of inhaled corticosteroids during pregnancy increases the risk of pregnancy induced hypertension and pre-eclampsia among asthmatic women. DESIGN: Nested case-control study. SETTING: Three administrative health databases from Quebec: RAMQ, MED-ECHO, and Fichier des evenements demographiques. PARTICIPANTS: 3505 women with asthma, totalling 4593 pregnancies, between 1990 and 2000. MAIN OUTCOME MEASURES: Pregnancy induced hypertension and pre-eclampsia. RESULTS: 302 cases of pregnancy induced hypertension and 165 cases of pre-eclampsia were identified. Use of inhaled corticosteroids from conception until date of outcome was not associated with an increased risk of pregnancy induced hypertension (adjusted odds ratio 1.02, 95% confidence interval 0.77 to 1.34) or pre-eclampsia (1.06, 0.74 to 1.53). No significant dose-response relation was observed between inhaled corticosteroids and pregnancy induced hypertension or pre-eclampsia. Oral corticosteroids were significantly associated with the risk of pregnancy induced hypertension (adjusted odds ratio 1.57, 1.02 to 2.41), and a trend was seen for pre-eclampsia (1.72, 0.98 to 3.02). CONCLUSION: No significant increase of the risk of pregnancy induced hypertension or pre-eclampsia was detected among users of inhaled corticosteroids during pregnancy, while markers of uncontrolled and severe asthma were found to significantly increase the risks of pregnancy induced hypertension and pre-eclampsia. PMID- 15659481 TI - The PRETTY FEW SEEDS2 gene encodes an Arabidopsis homeodomain protein that regulates ovule development. AB - The PRETTY FEW SEEDS2 gene encodes a homeodomain protein that regulates ovule development. In peptide alignments spanning the homeodomain and the WOX domain, PFS2 shared 95% amino acid identity with the PRESSED FLOWER and WUSCHEL proteins. In the pfs2-1 allele, the integuments display morphological abnormalities and 95% of the embryo sacs fail to develop properly, which results in reduced fecundity. PFS2 transcripts were most abundant in developing ovules, which accounts for the ovule phenotype in pfs2 mutants. In addition, PFS2 transcripts were present in developing primordia and differentiating organs, but, interestingly, they were absent during cell maturation. Ectopic PFS2 expression interfered with differentiation of primordia from meristems. For most plants, this resulted in fasciated stems, altered phyllotaxy, a cessation of primordia differentiation, or a combination of these. In the plants that made ovules, ectopic PFS2 expression blocked megaspore mother cell differentiation and often impeded polarized growth of the outer integument. PFS2 activity altered AGAMOUS expression, which accounts for some of the gain- and loss-of-function phenotypes. Based on analyses presented here, PFS2 affects either ovule patterning or differentiation. PMID- 15659482 TI - GATA4, 5 and 6 mediate TGFbeta maintenance of endodermal gene expression in Xenopus embryos. AB - The individual contributions of the three vertebrate GATA factors to endoderm formation have been unclear. Here we detail the early expression of GATA4, 5 and 6 in presumptive endoderm in Xenopus embryos and their induction of endodermal markers in presumptive ectoderm. Induction of HNF3beta by all three GATA factors was abolished when protein synthesis was inhibited, showing that these inductions are indirect. In contrast, whereas induction of Sox17alpha and HNF1beta by GATA4 and 5 was substantially reduced when protein synthesis was inhibited, induction by GATA6 was minimally affected, suggesting that GATA6 is a direct activator of these early endodermal genes. GATA4 induced GATA6 expression in the same assay and antisense morpholino oligonucleotides (MOs), designed to knock down translation of GATA6, blocked induction of Sox17alpha and HNF1beta by GATA4, suggesting that GATA4 induces these genes via GATA6 in this assay. All three GATA factors were induced by activin, although GATA4 and 6 required lower concentrations. GATA MOs inhibited Sox17alpha and HNF1beta induction by activin at low and high concentrations in the order: GATA6>GATA4>GATA5. Together with the timing of their expression and the effects of GATA MOs in vivo, these observations identify GATA6 as the predominant GATA factor in the maintenance of endodermal gene expression by TGFbeta signaling in gastrulating embryos. In addition, examination of gene expression and morphology in later embryos, revealed GATA5 and 6 as the most critical for the development of the gut and the liver. PMID- 15659483 TI - ceh-16/engrailed patterns the embryonic epidermis of Caenorhabditis elegans. AB - engrailed is a homeobox gene essential for developmental functions such as differentiation of cell populations and the onset of compartment boundaries in arthropods and vertebrates. We present the first functional study on engrailed in an unsegmented animal: the nematode Caenorhabditis elegans. In the developing worm embryo, ceh-16/engrailed is predominantly expressed in one bilateral row of epidermal cells (the seam cells). We show that ceh-16/engrailed primes a specification cascade through three mechanisms: (1) it suppresses fusion between seam cells and other epidermal cells by repressing eff-1/fusogen expression; (2) it triggers the differentiation of the seam cells through different factors, including the GATA factor elt-5; and (3) it segregates the seam cells into a distinct lateral cellular compartment, repressing cell migration toward dorsal and ventral compartments. PMID- 15659484 TI - Lysophosphatidic acid signaling controls cortical actin assembly and cytoarchitecture in Xenopus embryos. AB - The mechanisms that control shape and rigidity of early embryos are not well understood, and yet are required for all embryonic processes to take place. In the Xenopus blastula, the cortical actin network in each blastomere is required for the maintenance of overall embryonic shape and rigidity. However, the mechanism whereby each cell assembles the appropriate pattern and number of actin filament bundles is not known. The existence of a similar network in each blastomere suggests two possibilities: cell-autonomous inheritance of instructions from the egg; or mutual intercellular signaling mediated by cell contact or diffusible signals. We show that intercellular signaling is required for the correct pattern of cortical actin assembly in Xenopus embryos, and that lysophosphatidic acid (LPA) and its receptors, corresponding to LPA1 and LPA2 in mammals, are both necessary and sufficient for this function. PMID- 15659485 TI - Lineage analysis of the avian dermomyotome sheet reveals the existence of single cells with both dermal and muscle progenitor fates. AB - The dermomyotome develops into myotome and dermis. We previously showed that overall growth of the dermomyotome and myotome in the mediolateral direction occurs in a uniform pattern. While myofibers arise from all four dermomyotome lips, the dermis derives from both medial and lateral halves of the dermomyotome sheet. Here we mapped the fate of this epithelial sheet by analyzing cell types that arise from its central region. We found that these precursors give rise not only to dermis, as expected, but also to a population of proliferating progenitors in the myotome that maintain expression of PAX7, PAX3 and FREK. Given this dual fate, we asked whether single dermomyotome precursors generate both dermal and mitotic myoblast precursors, or alternatively, whether these cell types derive from distinct epithelial founders. Inovo clonal analysis revealed that single dermomyotome progenitors give rise to both derivatives. This is associated with a sharp change in the plane of cell division from the young epithelium, in which symmetrical divisions occur parallel to the mediolateral plane of the dermomyotome, to the dissociating dermomyotome, in which cell divisions become mostly perpendicular. Taken together with clonal analysis of the dermomyotome sheet, this suggests that a first stage of progenitor self-renewal, accounting for dermomyotomal expansion, is followed by fate segregation, which correlates with the observed shift in mitotic spindle orientation. PMID- 15659486 TI - Wnt1 regulates neurogenesis and mediates lateral inhibition of boundary cell specification in the zebrafish hindbrain. AB - The formation of localised signalling centres is essential for patterning of a number of tissues during development. Previous work has revealed that a distinct population of boundary cells forms at the interface of segments in the vertebrate hindbrain, but the role of these cells is not known. We have investigated the function of the Wnt1 signalling molecule that is expressed by boundary and roof plate cells in the zebrafish hindbrain. Knockdown of wnt1 or of tcf3b, a mediator of Wnt signalling, leads to ectopic expression of boundary cell markers, rfng and foxb1.2, in non-boundary regions of the hindbrain. Ectopic boundary marker expression also occurs following knockdown of rfng, a modulator of Notch signalling required for wnt1 expression at hindbrain boundaries. We show that the boundary and roof plate expression of wnt1 each contribute to upregulation of proneural and delta gene expression and neurogenesis in non-boundary regions, which in turn blocks ectopic boundary marker expression. Boundary cells therefore play a key role in the regulation of cell differentiation in the zebrafish hindbrain. The network of genes underlying the regulation of neurogenesis and lateral inhibition of boundary cell formation by Wnt1 has a striking similarity to mechanisms at the dorsoventral boundary in the Drosophila wing imaginal disc. PMID- 15659487 TI - Developmentally programmed remodeling of the Drosophila olfactory circuit. AB - Neural circuits are often remodeled after initial connections are established. The mechanisms by which remodeling occurs, in particular whether and how synaptically connected neurons coordinate their reorganization, are poorly understood. In Drosophila, olfactory projection neurons (PNs) receive input by synapsing with olfactory receptor neurons in the antennal lobe and relay information to the mushroom body (MB) calyx and lateral horn. Here we show that embryonic-born PNs participate in both the larval and adult olfactory circuits. In the larva, these neurons generally innervate a single glomerulus in the antennal lobe and one or two glomerulus-like substructures in the MB calyx. They persist in the adult olfactory circuit and are prespecified by birth order to innervate a subset of glomeruli distinct from larval-born PNs. Developmental studies indicate that these neurons undergo stereotyped pruning of their dendrites and axon terminal branches locally during early metamorphosis. Electron microscopy analysis reveals that these PNs synapse with MB gamma neurons in the larval calyx and that these synaptic profiles are engulfed by glia during early metamorphosis. As with MB gamma neurons, PN pruning requires cell-autonomous reception of the nuclear hormone ecdysone. Thus, these synaptic partners are independently programmed to prune their dendrites and axons. PMID- 15659488 TI - Lunatic fringe null female mice are infertile due to defects in meiotic maturation. AB - We have demonstrated that Notch genes are expressed in developing mammalian ovarian follicles. Lunatic fringe is an important regulator of Notch signaling. In this study, data are presented that demonstrate that radical fringe and lunatic fringe are expressed in the granulosa cells of developing follicles. Lunatic fringe null female mice were found to be infertile. Histological analysis of the lunatic fringe-deficient ovary demonstrated aberrant folliculogenesis. Furthermore, oocytes from these mutants did not complete meiotic maturation. This is a novel observation because this is the first report describing a meiotic defect that results from mutations in genes that are expressed in the somatic granulosa cells and not the oocytes. This represents a new role for the Notch signaling pathway and lunatic fringe in mammalian folliculogenesis. PMID- 15659489 TI - Prediction of docetaxel response in human breast cancer by gene expression profiling. AB - PURPOSE: Docetaxel is one of the most effective anticancer drugs available in the treatment of breast cancer. Nearly half of the treated patients, however, do not respond to chemotherapy and suffer from side effects. The ability to reliably predict a patient's response based on tumor gene expression will improve therapeutic decision making and save patients from unnecessary side effects. PATIENTS AND METHODS: A total of 44 breast tumor tissues were sampled by biopsy before treatment with docetaxel, and the response to therapy was clinically evaluated by the degree of reduction in tumor size. Gene expression profiling of the biopsy samples was performed with 2,453 genes using a high-throughput reverse transcriptase polymerase chain reaction technique. Using genes differentially expressed between responders and nonresponders, a diagnostic system based on the weighted-voting algorithm was constructed. RESULTS: This system predicted the clinical response of 26 previously unanalyzed samples with over 80% accuracy, a level promising for clinical applications. Diagnostic profiles in nonresponders were characterized by elevated expression of genes controlling the cellular redox environment (ie, redox genes, such as thioredoxin, glutathione-S-transferase, and peroxiredoxin). Overexpression of these genes protected cultured mammary tumor cells from docetaxel-induced cell death, suggesting that enhancement of the redox system plays a major role in docetaxel resistance. CONCLUSION: These results suggest that the clinical response to docetaxel can be predicted by gene expression patterns in biopsy samples. The results also suggest that one of the molecular mechanisms of the resistance is activation of a group of redox genes. PMID- 15659490 TI - Up-front tandem high-dose chemotherapy compared with standard chemotherapy with doxorubicin and paclitaxel in metastatic breast cancer: results of a randomized trial. AB - PURPOSE: The role of high-dose chemotherapy (HDCT) in metastatic breast cancer remains controversial. Trials with late intensification HDCT have failed to show an advantage in overall survival. This study was initiated to compare up-front tandem HDCT and standard combination therapy in patients with metastatic breast cancer. PATIENTS AND METHODS: Patients without prior chemotherapy for metastatic disease were randomly assigned to standard combination therapy with doxorubicin and paclitaxel (AT) or double HDCT with cyclophosphamide, mitoxantrone, and etoposide followed by peripheral-blood stem-cell transplantation. HDCT was repeated after 6 weeks. Patients were stratified by menopausal and hormone receptor status. The primary objective was to compare complete response (CR) rates. RESULTS: A total of 93 patients were enrolled onto the trial. Intent-to treat CR rates for patients randomized to HDCT and AT were 12.5% and 11.1%, respectively (P = .84). Objective response rates were 66.7% for patients in the high-dose group and 64.4% for patients in the AT arm (P = .82). In an intent-to treat analysis, there were no significant differences between the two treatments in median time to progression (HDCT, 11.1 months; AT, 10.6 months; P = .67), duration of response (HDCT, 13.9 months; AT, 14.3 months; P = .98), and overall survival (HDCT, 26.9 months; AT, 23.4 months; P = .60). HDCT was associated with significantly more myelosuppression, infection, diarrhea, stomatitis, and nausea and vomiting, whereas patients treated with AT developed more neurotoxicity. CONCLUSION: This trial failed to show a benefit for up-front tandem HDCT compared with standard combination therapy. HDCT was associated with more acute adverse effects. PMID- 15659491 TI - Randomized phase II study of two doses of gefitinib in hormone-refractory prostate cancer: a trial of the National Cancer Institute of Canada-Clinical Trials Group. AB - PURPOSE: Overexpression of the epidermal growth factor receptor has been demonstrated in advanced prostate cancer and is associated with a poor outcome. A multi-institutional, randomized, phase II study was undertaken by the National Cancer Institute of Canada-Clinical Trials Group to evaluate the efficacy and toxicity of two doses of oral gefitinib in patients with minimally symptomatic, hormone-refractory prostate cancer (HRPC). PATIENTS AND METHODS: Between July and November 2001, 40 patients with HRPC and increasing prostate-specific antigen (PSA) or progression in measurable disease who had not received prior chemotherapy were randomly assigned to 250 mg (n = 19) or 500 mg (n = 21) oral gefitinib daily continuously. The primary end points were PSA response rate and objective measurable response. Functional Assessment of Cancer Therapy Prostate Cancer Subscale (FACT-P) quality-of-life questionnaires were completed at baseline and during treatment. RESULTS: None of the patients demonstrated a PSA or objective measurable response. Five (14.3%) of 35 assessable patients had stable PSA (one patient at 250 mg and four patients at 500 mg), and five patients (14.3%) had a best response of stable disease (duration, 2.5 to 16.8 months). No significant effect on the rate of increase in PSA was seen. The most common drug related nonhematologic toxicities observed were grade 1 to 2 diarrhea (250 mg, 65%; 500 mg, 56%), fatigue (250 mg, 29%; 500 mg, 33%), and grade 1 to 2 skin rash (250 mg, 24%; 500 mg, 39%). FACT-P scores decreased during treatment, indicating worsening of symptoms compared with baseline. CONCLUSION: Gefitinib did not result in any responses in PSA or objective measurable disease at either dose level. Gefitinib has minimal single-agent activity in HRPC. PMID- 15659492 TI - Role of anti-hepatitis C virus (HCV) treatment in HCV-related, low-grade, B-cell, non-Hodgkin's lymphoma: a multicenter Italian experience. AB - PURPOSE: Hepatitis C virus (HCV) is endemic in some areas of Northwestern Europe and the United States. HCV has been shown to play a role in the development of both hepatocellular carcinoma and B-cell non-Hodgkin's lymphoma (B-NHL). The biologic mechanisms underlying the lymphomagenic activity of the virus so far are under investigation. In this study, the role of antiviral (anti-HCV) treatment in B-NHL associated with HCV infection is evaluated. PATIENTS AND METHODS: Thirteen patients with histologically proven low-grade B-NHL characterized by an indolent course (ie, doubling time no less than 1 year, no bulky disease) and carrying HCV infection were enrolled on the study. All patients underwent antiviral treatment alone with pegilated interferon and ribavirin. Response assessment took place at 6 and 12 months. RESULTS: Of the twelve assessable patients, seven (58%) achieved complete response and two (16%) partial hematologic response at 14.1 +/- 9.7 months (range, 2 to 24 months, median follow-up, 14 months), while two had stable disease with only one patient experiencing progression of disease. Hematologic responses (complete and partial, 75%) were highly significantly associated to clearance or decrease in serum HCV viral load following treatment (P = .005). Virologic response was more likely to be seen in HCV genotype 2 (P = .035), while hematologic response did not correlate with the viral genotype. Treatment-related toxicity did not cause discontinuation of therapy in all but two patients, one of whom, however, achieved complete response. CONCLUSION: This experience strongly provides a role for antiviral treatment in patients affected by HCV-related, low grade, B-cell NHL. PMID- 15659493 TI - Polymorphisms in FcgammaRIIIA (CD16) receptor expression are associated with clinical response to rituximab in Waldenstrom's macroglobulinemia. AB - PURPOSE: Rituximab is an important therapeutic for Waldenstrom's macroglobulinemia (WM). Polymorphisms in FcgammaRIIIA (CD16) receptor expression modulate human immunoglobulin G1 binding and antibody-dependent cell-mediated cytotoxicity, and may therefore influence responses to rituximab. PATIENTS AND METHODS: Sequence analysis of the entire coding region of FcgammaRIIIA was undertaken in 58 patients with WM whose outcomes after rituximab were known. RESULTS: Variations in five codons of FcgammaRIIIA were identified. Two were commonly observed (FcgammaRIIIA-48 and FcgammaRIIIA-158) and predicted for amino acid polymorphisms at FcgammaRIIIA-48: leucine/leucine (L/L), leucine/arginine (L/R), and leucine/histidine (L/H). Polymorphisms at FcgammaRIIIA-158 were phenylalanine/phenylalanine (F/F), phenylalanine/valine (F/V), and valine/valine (V/V). A clear linkage between these polymorphisms was detected and all patients with FcgammaRIIIA-158F/F were always FcgammaRIIIA-48L/L, and patients with either FcgammaRIIIA-L/R or -L/H always expressed at least one valine at FcgammaRIIIA-158 (P < or = .001). The response trend was higher for patients with FcgammaRIIIA 48L/H (38.5%) versus -48L/R (25.0%) and LL (22.0%), and was significantly higher for patients with FcgammaRIIIA-158V/V (40.0%) and -V/F (35%) versus -158F/F (9.0%; P = .030). Responses for patients with FcgammaRIIIA-48L/L were higher when at least one valine was present at FcgammaRIIIA-158 (P = .057), thereby supporting a primary role for FcgammaRIIIA-158 polymorphisms in predicting rituximab responses. With a median follow-up of 13 months, no significant differences in the median time to progression and progression-free survival were observed when patients were grouped according to their FcgammaRIIIA-48 and -158 polymorphisms. CONCLUSION: The results of these studies therefore support a predictive role for FcgammaRIIIA-158 polymorphisms and responses to rituximab in WM. PMID- 15659494 TI - Docetaxel and continuous-infusion fluorouracil versus epirubicin, cisplatin, and fluorouracil for advanced gastric adenocarcinoma: a randomized phase II study. AB - PURPOSE: A combination of docetaxel and fluorouracil (DF) was evaluated in an outpatient setting and compared with epirubicin, cisplatin, and fluorouracil (ECF), which served as an internal control arm to avoid selection bias. PATIENTS AND METHODS: Patients with metastatic or locally advanced gastric adenocarcinoma without prior chemotherapy were randomly assigned to receive either ECF (epirubicin 50 mg/m(2) day 1, cisplatin 60 mg/m(2) day 1, and fluorouracil 200 mg/m(2) days 1 through 21, every 3 weeks) or DF (docetaxel 75 mg/m(2) day 1, and fluorouracil 200 mg/m(2) days 1 through 21, every 3 weeks). RESULTS: Ninety patients were randomly assigned. Toxicity was rarely severe. Major toxic effects included diarrhea, stomatitis, and leukopenia in the DF arm and nausea, vomiting, and leukopenia in the ECF arm. Forty-three of 45 patients in each arm were assessable. In the DF arm, two patients (4.4%, intent to treat) experienced a confirmed complete tumor remission as best response, and 15 patients (33.3%) experienced a confirmed partial remission (overall response rate [ORR], 37.8%; 95% CI, 25.9% to 51.9%). Two patients (4.4%) in the ECF arm showed confirmed complete remission, and 14 (31.1%) showed confirmed partial remission (ORR, 35.6%; 95% CI, 24.8% to 48.7%). For the DF and ECF arms, the median survival was 9.5 and 9.7 months, and the median time to tumor progression 5.5 and 5.3 months, respectively. CONCLUSION: DF can be safely given in an ambulant setting. Compared with ECF, the dual combination of DF shows promising efficacy and may be an alternative treatment option that avoids cisplatin. PMID- 15659495 TI - Intensive systemic chemotherapy combined with surgery for metastatic colorectal cancer: results of a phase II study. AB - PURPOSE: To evaluate the efficacy and tolerability of the metastatic irinotecan plus oxaliplatin (MIROX) strategy (adjuvant FOLFOX-7 followed by FOLFIRI), in patients with resectable metastatic colorectal cancer. PATIENTS AND METHODS: Forty-seven patients with resectable metastases of colorectal cancer were prospectively enrolled onto this study. Treatment consisted of six cycles of leucovorin 400 mg/m(2), oxaliplatin 130 mg/m(2) in a 120-minute infusion, and fluorouracil (FU) 2,400 mg/m(2) in a 46-hour infusion, every 2 weeks (FOLFOX-7), followed by six cycles of leucovorin 400 mg/m(2), irinotecan 180 mg/m(2) in a 90 minute infusion, bolus FU 400 mg/m(2), and FU 2,400 mg/m(2) as a 46-hour infusion, every 2 weeks (FOLFIRI). Surgery was performed before chemotherapy in 25 patients and after six cycles of FOLFOX-7 in 22 patients (six cycles of FOLFIRI were administered after surgery). RESULTS: All but one of the patients underwent curative surgery. Two patients refused postoperative chemotherapy. Tolerability was generally good. The main toxicities were grade 3 to 4 neutropenia (13%) and thrombocytopenia (11%); no febrile neutropenia or bleeding occurred, and there were no deaths caused by toxicity. Two pathologically confirmed complete responses and 15 partial responses were obtained with FOLFOX-7 in the 22 patients who received this regimen before surgery (overall response rate, 77%; 95% CI, 68 to 86). The median disease-free survival time was 21 months; the median overall survival has not yet been reached. The 2-year overall and disease-free survival rates were 89% and 47%, respectively. CONCLUSION: The MIROX strategy is feasible and well tolerated by patients with resectable metastatic colorectal cancer. Progression-free and overall survival rates are promising, with a median of 38 months of follow-up. This strategy currently is being compared with the leucovorin and FU regimen in a phase III trial. PMID- 15659496 TI - Racial differences in surgical evaluation, treatment, and outcome of locoregional esophageal cancer: a population-based analysis of elderly patients. AB - PURPOSE: We investigated racial disparities in access to surgical evaluation, receipt of surgery, and survival among elderly patients with locoregional esophageal cancer. METHODS: We selected 2,946 white patients and 367 black patients who were older than 65 years and had clinically locoregional esophageal cancer in the Surveillance, Epidemiology, and End Results (SEER) registry (1991 to 1999). Treatment and outcome data were obtained from the linked SEER-Medicare databases. We used logistic regression analysis to estimate odds ratios (ORs) for being seen by a surgeon and for undergoing surgery. Cox proportional hazards analyses were performed to estimate hazard ratios (HRs) for survival adjusted for medical, nonmedical, and treatment characteristics. RESULTS: The rate of surgery for black patients was half that of white patients (25% v 46%; OR, 0.38; P < .001), which was caused by both a lower rate of seeing a surgeon (70% v 78%; OR, 0.66; P < .001) and a lower rate of surgery once seen (35% v 59%; OR, 0.38; P < .001). These racial disparities were only partly explained by differences in patient and cancer characteristics, and not by nonmedical factors, such as socioeconomic status. The 2-year survival rate was lower for black patients (18% v 25%; HR, 1.18; P = .004), but this racial difference disappeared when corrected for treatment received (adjusted HR, 1.02; P = .80). CONCLUSION: Underuse of potentially curative surgery is an important potential explanation for the poorer survival of black patients with locoregional esophageal cancer. Barriers to surgical evaluation and treatment need to be reduced, whether related to patient or healthcare system factors. PMID- 15659497 TI - Prognostic implications of multiple lymphatic basin drainage in patients with truncal melanoma. AB - PURPOSE: Lymphatic drainage to multiple basins (MLBD) is frequently observed in patients with truncal melanoma undergoing sentinel lymph node (SLN) mapping. Recently published data suggest that patients with MLBD are at increased risk of nodal metastases compared with those with single lymphatic basin drainage (SLBD). We studied the impact of MLBD on SLN positivity and survival. PATIENTS AND METHODS: We identified 266 patients with truncal melanoma undergoing SLN mapping and biopsy from 1995 to 2001. MLBD was defined as lymphoscintigraphic and intraoperative identification of an SLN in more than one nodal basin. Clinical and pathologic variables were recorded and analyzed for their impact on survival. RESULTS: MLBD occurred in 76 patients (29%), and SLBD occurred in 190 patients (71%). Clinical and pathologic variables were similar between the two groups, although there were more males in the MLBD group (78% v 64%; P = .034). Patients with MLBD did not have higher risk for positive SLNs compared with those with SLBD (22% v 21%, respectively). Five-year survival for patients with MLBD was less favorable than that of patients with SLBD (68% v 78%, respectively; log-rank P = .04). Multivariate analysis revealed that increasing Breslow thickness (P < .001), SLN metastasis (P < .001), and MLBD (P = .04) were independent predictors of poor overall and relapse-free survival. The negative prognostic implication of MLBD remained significant when only patients with negative SLNs were analyzed (relative risk, 2.7; P = .03). CONCLUSION: MLBD in patients with truncal melanoma undergoing SLN mapping is associated with a less favorable survival compared with patients with SLBD, independent of SLN status. PMID- 15659498 TI - Phase I clinical trial of mafosfamide in infants and children aged 3 years or younger with newly diagnosed embryonal tumors: a pediatric brain tumor consortium study (PBTC-001). AB - PURPOSE: A phase I trial of intrathecal (IT) mafosfamide was performed to determine the optimal dose, dose-limiting toxicities, and incidence and severity of other toxicities when administered in association with concomitant multiagent systemic chemotherapy to children younger than 3 years with newly diagnosed embryonal tumors. PATIENTS AND METHODS: Twenty-five assessable patients received IT mafosfamide at one of six dose levels ranging from 5 mg to 17 mg. Patients were premedicated with dexamethasone (0.15 mg/kg) and morphine (0.1 mg/kg) before receiving IT mafosfamide. Serial samples of CSF for pharmacokinetic studies were obtained in a subset of patients with Ommaya reservoirs. RESULTS: Irritability, presumably secondary to pain or headache during mafosfamide administration, was dose limiting in two of three patients at the 17-mg dose level. The maximum tolerated dose of IT mafosfamide following premedication with dexamethasone and morphine was 14 mg. CONCLUSION: The maximum tolerated dose and recommended phase II dose of IT mafosfamide in patients younger than 3 years with newly diagnosed embryonal CNS tumors is 14 mg. A trial to assess the efficacy of regional therapy with IT mafosfamide administered with intensive systemic chemotherapy in children younger than 3 years with primary intracranial embryonal tumors is now in progress. PMID- 15659499 TI - Results of induction chemotherapy in children older than 1 year with a stage 4 neuroblastoma treated with the NB 97 French Society of Pediatric Oncology (SFOP) protocol. AB - PURPOSE: To test the metastatic response rate in stage 4 neuroblastoma, using dose-intensive induction chemotherapy in a multi-institutional setting. PATIENTS AND METHODS: From 1998 to 1999, 47 consecutive children were treated according to N7 protocol. Children received cyclophosphamide 140 mg/kg, doxorubicin 75 mg/m(2), and vincristine 0.066 mg/kg (CAV) in cycles 1, 2, 4, and 6, and cisplatinum 200 mg/m(2) and etoposide 600 mg/m(2) (P/VP) in cycles 3, 5, and 7. The International Neuroblastoma Staging system was used with an emphasis on skeletal evaluation by 123-iodine-metaiodobenzylguanidine (MIBG) scintigraphy. A phase II study evaluating the metastasis complete response rate after induction chemotherapy was conducted in patients who had positive metastatic sites on MIBG scans at diagnosis. RESULTS: Forty-six patients were assessable for toxicity. Hematologic toxicity was the main toxicity observed. Neutropenia was more frequent after CAV than after P/VP (P < .001). A higher rate of thrombocytopenia was observed after P/VP (P = .03). Forty patients with positive MIBG were assessable for metastatic response, and complete regression of metastases was achieved in 17 patients (ie, 43%; 95% CI, 27% to 59%). Of all 47 patients, 21 achieved complete metastatic response. CONCLUSION: The N7 induction chemotherapy protocol was feasible in a multicentric setting. The observed metastasis complete response rate was similar to that obtained in our previous studies and significantly lower than that published in a previous series using the same regimen. In our hands, escalating doses of cyclophosphamide and prolonging conventional chemotherapy with the same drugs failed to improve the metastasis complete response rate. PMID- 15659500 TI - Advanced-stage large-cell lymphoma in children and adolescents: results of a randomized trial incorporating intermediate-dose methotrexate and high-dose cytarabine in the maintenance phase of the APO regimen: a Pediatric Oncology Group phase III trial. AB - PURPOSE: The Pediatric Oncology Group adopted a histology-based approach to non Hodgkin's lymphoma and treated patients with advanced large-cell lymphoma on a separate protocol (doxorubicin, vincristine, prednisone, 6-mercaptopurin, and methotrexate; APO regimen). In this study, we assessed the effects of an intense antimetabolite therapy alternating with APO on overall survival (OS) and event free survival (EFS) and looked into biologic correlates. PATIENTS AND METHODS: From December 1994 to April 2000, we enrolled 180 eligible pediatric patients with stage III/IV large-cell lymphoma (LCL); 90 patients were randomly assigned to the intermediate-dose methotrexate (IDM) and high-dose cytarabine (HiDAC) arm, 85 patients to the APO arm, and five patients directly to the APO arm by study design due to CNS involvement. Planned therapy duration was 12 months. RESULTS: The 4-year EFS for all patients was 67.4% (SE, 4.2%), and OS was 80.1% (SE, 3.6%) without any significant difference between the two arms. The 4-year EFS and OS were 71.8% (SE, 6.1%) and 88.1% (SE, 4.4%), respectively, for patients with anaplastic large-cell lymphoma, and 63.8% (SE, 10.3%) and 70.3% (SE, 9.0%), respectively, for patients with diffuse large B-cell lymphoma. Only 11 patients required radiation (due to unresponsive bulky disease or CNS involvement). The IDM/HiDAC arm was associated with more toxicity. CONCLUSION: The efficacy of incorporating IDM/HiDAC in the treatment plan of pediatric and adolescent patients with advanced-stage LCL was inconclusive as to its effect on EFS, regardless of the lymphoma phenotype. It cannot be excluded that with a higher number of patients, one treatment could prove superior and future studies will build on these data. PMID- 15659501 TI - Ewing sarcomas with p53 mutation or p16/p14ARF homozygous deletion: a highly lethal subset associated with poor chemoresponse. AB - PURPOSE: EWS-FLI1 fusion type, p53 mutation, and homozygous deletion of p16/p14ARF have each been shown to be prognostically significant in Ewing sarcoma (ES). We provide the first combined prognostic analysis of these three molecular parameters in ES. PATIENTS AND METHODS: We studied 60 patients with ES (stage: localized in 54, metastatic in six). All cases were confirmed to contain the EWS FLI1 (29 type 1, 12 type 2, 14 other types) or EWS-ERG fusions (five cases). Homozygous deletion of p16/p14ARF, and p53 mutations were determined by fluorescent in situ hybridization and Affymetrix (Santa Clara, CA) p53 GeneChip microarray hybridization, respectively. RESULTS: Eight cases (13.3%) contained point mutations of p53, and eight cases (13.3%) showed p16/p14ARF deletion, including one case with both alterations. Among 32 cases with data on histologic chemoresponse, all 10 with alterations in p53 or p16/p14ARF showed a poor chemoresponse (P = .03). Variables predicting poorer overall survival included p53 mutation alone (P < .001), either p53 or p16/p14ARF alteration (P < .001), and stage (P < .01). In multivariate analysis, alterations of p53 and/or p16/p14ARF as a single variable, was the most adverse prognostic factor (P < .001), followed by stage (P = .04). In a multivariate analysis with alterations of p53 and p16/p14ARF as separate variables, both were significant (P < .001 and P = .03, respectively). Six cases with p16/p14ARF deletion were also studied for co-deletion of the contiguous methylthioadenosine phosphorylase gene, and this was detected in four cases. CONCLUSION: Alterations in p53 or p16/p14ARF are found in a fourth of ES cases and define a subset with highly aggressive behavior and poor chemoresponse. PMID- 15659502 TI - Osteosarcoma relapse after combined modality therapy: an analysis of unselected patients in the Cooperative Osteosarcoma Study Group (COSS). AB - PURPOSE: To evaluate the impact of patient, tumor, and treatment-related factors on outcome in unselected patients with recurrent osteosarcoma. PATIENTS AND METHODS: Five hundred seventy-six consecutive patients who had achieved a first complete surgical remission (CR) during combined-modality therapy on neoadjuvant Cooperative Osteosarcoma Study Group (COSS) protocols and then developed recurrent osteosarcoma were analyzed (median time from biopsy to relapse, 1.6 years; range, 0.1 to 14.3 years). There were 501 patients with metastases, 44 with local recurrences, and 31 with both. Metastases involved lungs (469 patients), bones (90 patients), and/or other sites (54 patients). RESULTS: After a median follow-up of 1.2 years for all patients and 4.2 years for survivors, actuarial overall survival (OS) rates at 2, 5, and 10 years were 0.38, 0.23, and 0.18, respectively. Five-year OS was 0.39 for 339 patients with and 0.00 for 229 patients without a second surgical CR (P < .0001). A long time to relapse, a solitary lesion, and, in the case of pulmonary metastases, unilateral disease and the absence of pleural disruption, were of positive prognostic value in uni- and multivariate analyses, as were a second surgical CR and the use of second-line chemotherapy. Radiotherapy was associated with moderately prolonged survival in patients without a second CR. The very limited prognostic differences associated with the use of second-line chemotherapy appeared to be more pronounced with polychemotherapy. CONCLUSION: Time to relapse and tumor burden correlate with postrelapse outcome in osteosarcoma. Complete surgery is an essential component of curative second-line therapy. Chemotherapy, particularly chemotherapy with more than one agent, may contribute to limited improvements in outcome. PMID- 15659503 TI - Platinum-etoposide chemotherapy in elderly patients with small-cell lung cancer: results of a randomized multicenter phase II study assessing attenuated-dose or full-dose with lenograstim prophylaxis--a Forza Operativa Nazionale Italiana Carcinoma Polmonare and Gruppo Studio Tumori Polmonari Veneto (FONICAP-GSTPV) study. AB - PURPOSE: Small-cell lung cancer (SCLC) is increasingly diagnosed in elderly patients, who are at higher risk of treatment-related morbidity and mortality. We conducted a randomized two-stage phase II study to assess the therapeutic index of two different platinum/etoposide regimens, attenuated-dose (AD) and full-dose (FD) plus prophylactic lenograstim. PATIENTS AND METHODS: SCLC patients older than 70 years were randomized to receive four courses of cisplatin 25 mg/m(2) on days 1 and 2, and etoposide 60 mg/m(2) on days 1, 2, and 3 every 3 weeks (AD); or cisplatin 40 mg/m(2) on days 1 and 2, and etoposide 100 mg/m(2) on days 1, 2, and 3 every 3 weeks, plus lenograstim 5 mg/kg days 5 through 12, every 3 weeks (FD). A combined primary end point named therapeutic success (TS), which took into account activity, toxicity, and compliance, was used. RESULTS: Ninety-five patients were enrolled. Seventy-five percent and 72% of the patients in the AD and FD arms, respectively, completed the treatment as per protocol. Response rate was 39% and 69% in the AD and FD arms, respectively, and 1-year survival probability was 18% and 39%, respectively. Treatment was well tolerated in both groups, with no grade 3 to 4 myelotoxicity in the AD arm, and 12% myelotoxicity in the FD arm. Overall, the observed TSs were 10 (36%) of 28 patients and 42 (63%) of 67 patients for AD and FD treatments, respectively. CONCLUSION: In elderly patients with SCLC a full-dose cisplatin/etoposide regimen combined with prophylactic lenograstim is active and feasible, while attenuated doses of the same regimen are associated with a poor therapeutic outcome. PMID- 15659504 TI - Phase II study of ET-743 in advanced soft tissue sarcomas: a European Organisation for the Research and Treatment of Cancer (EORTC) soft tissue and bone sarcoma group trial. AB - PURPOSE: This nonrandomized multicenter phase II study was performed to evaluate the activity and safety of Ecteinascidin (ET-743) administered at a dose of 1.5 mg/m(2) as a 24-hour continuous infusion every 3 weeks in patients with pretreated advanced soft tissue sarcoma. PATIENTS AND METHODS: Patients with documented progressive advanced soft tissue sarcoma received ET-743 as second- or third-line chemotherapy. Antitumor activity was evaluated every 6 weeks until progression, excessive toxicity, or patient refusal. RESULTS: One hundred four patients from eight European institutions were included in the study (March 1999 to November 2000). A total of 410 cycles were administered in 99 assessable patients. Toxicity mainly involved reversible grade 3 to 4 asymptomatic elevation of transaminases in 40% of patients, and grade 3 to 4 neutropenia was observed in 52% of patients. There were eight partial responses (PR; objective regression rate, 8%), 45 no change (NC; > 6 months in 26% of patients), and 39 progressive disease. A progression arrest rate (PR + NC) of 56% was observed in leiomyosarcoma and 61% in synovialosarcoma. The median duration of the time to progression was 105 days, and the 6-month progression-free survival was 29%. The median duration of survival was 9.2 months. CONCLUSION: ET-743 seems to be a promising active agent in advanced soft tissue sarcoma, with no cumulative toxicities. The 6-months progression-free survival observed in advanced soft tissue sarcoma compares favorably with those obtained with other active drugs tested in second-line chemotherapy in previous European Organisation for the Research and Treatment of Cancer trials. The median overall survival was unusually long in these heavily pretreated patients mainly due to the high number of patients who benefit from the drug in terms of tumor control. PMID- 15659505 TI - Imatinib mesylate in patients with adenoid cystic cancers of the salivary glands expressing c-kit: a Princess Margaret Hospital phase II consortium study. AB - PURPOSE: This study aimed to assess the antitumor activity of imatinib in adenoid cystic carcinoma (ACC) of the salivary gland expressing c-kit. A high level of c kit expression has been identified in more than 90% of ACCs. Imatinib specifically inhibits autophosphorylation of the bcr-abl, platelet-derived growth factor receptor beta, and c-kit tyrosine kinases. PATIENTS AND METHODS: In a single-arm, two-stage, phase II clinical trial, adult patients with unresectable or metastatic ACC measurable by Response Evaluation Criteria in Solid Tumors Group criteria and expressing c-kit by immunohistochemistry were treated with imatinib 400 mg orally bid. Response was assessed every 8 weeks. RESULTS: Sixteen patients have been enrolled onto the study; 10 were female. Median age was 47 years (range, 31 to 69 years). Median Eastern Cooperative Oncology Group performance status was 1 (range, 0 to 2). Fourteen patients had lung metastases, 14 had prior radiotherapy, and six had prior chemotherapy. Toxicities occurring in at least 50% of patients included fatigue, nausea, vomiting, diarrhea, anorexia, edema, dyspnea, and/or headache, usually of mild to moderate severity. In 15 patients assessable for response, no objective responses have been observed. Nine patients had stable disease as best response. Six patients had progressive disease after two cycles. CONCLUSION: Because of the lack of activity, the study has been stopped after the first stage and additional evaluation of imatinib in this population is not warranted. Overexpression of wild-type c-kit was not sufficient for clinical benefit from imatinib in ACC. Accrual to this study was rapid for a relatively rare cancer, encouraging additional efforts to identify more effective systemic therapy for these patients. PMID- 15659506 TI - Quality-of-life assessment in the symptom management trials of the National Cancer Institute-supported Community Clinical Oncology Program. AB - PURPOSE: To examine how quality of life (QOL) is prospectively conceptualized, defined, and measured in the symptom management clinical trials supported by the National Cancer Institute Community Clinical Oncology Program (CCOP). METHODS: All QOL research objectives, rationales, assessment instruments, symptoms treated, and types of interventions from the CCOP symptom management portfolio of clinical trials were extracted and analyzed. RESULTS: QOL assessments were proposed in 68 (52%) of the 130 total CCOP symptom management trials initiated since 1987. A total of 22 global QOL instruments were identified. Both the frequency of symptom management trials and the frequency of QOL assessment have increased significantly over time. The Functional Assessment of Cancer Therapy and Uniscale instruments were the most widely used QOL instruments, included in 55% of trials assessing QOL. The conceptual framework for QOL inclusion was limited to univariate relationships between symptom relief and global improvements in QOL. No consistent associations were found between QOL assessment and either the symptoms targeted or types of interventions. CONCLUSION: To advance the state of the science, research protocols need to provide more explicit rationales for assessing QOL in symptom management trials and for the selection of the QOL instrument(s) to be used. Conceptual frameworks that specify the hypothesized links between the specific symptom(s) being managed, interactions with other symptoms, different domains of QOL, and global QOL also need to be more precisely described. Methodologic and conceptual advances in QOL symptom management trials are critical to fulfill the promise of alleviating suffering and improving the QOL of cancer patients. PMID- 15659507 TI - Long-term health-related quality of life, growth, and spiritual well-being after hematopoietic stem-cell transplantation. AB - PURPOSE: To examine health-related quality of life (HRQOL) and growth, and spiritual well-being in adult survivors of hematopoietic stem-cell transplantation (HSCT) for a malignant disease. METHODS: HSCT survivors (n = 662) were recruited through the International Bone Marrow Transplant Registry/Autologous Blood and Marrow Transplant Registry and were drawn from 40 transplantation centers. HSCT survivors completed a telephone interview and a set of questionnaires a mean of 7.0 years post-HSCT (range, 1.8 to 22.6 years). Study measures included a variety of standardized measures of HRQOL and growth and spiritual well-being. An age- and sex-matched healthy comparison (HC) group (n = 158) was recruited using a peer nomination method. The HC group completed a parallel telephone interview and set of questionnaires. RESULTS: Multivariate analysis of variance analyses found the HSCT survivor group reported poorer status relative to the HC group for all HRQOL outcome clusters including physical health, physical functioning, social functioning, psychological adjustment, and dyadic adjustment. In contrast, the HSCT survivor group reported more psychological and interpersonal growth. Mean effect size for the 24 outcome indices examined was 0.36 standard deviations, an effect size often considered clinically meaningful or important. The largest group differences were found for measures of general health, physical function and well-being, depression, cognitive function, and fatigue. CONCLUSION: The experience of HSCT for a malignant disease has a wide-ranging, longstanding, and profound impact on adult recipients. Relative to healthy controls, HSCT survivors reported poorer physical, psychological, and social functioning but, conversely, more psychological and interpersonal growth, differences that appeared to persist many years after HSCT. PMID- 15659508 TI - Systematic review of microsatellite instability and colorectal cancer prognosis. AB - PURPOSE: A number of studies have investigated the relationship between microsatellite instability (MSI) and colorectal cancer (CRC) prognosis. Although many have reported a better survival with MSI, estimates of the hazard ratio (HR) among studies differ. To derive a more precise estimate of the prognostic significance of MSI, we have reviewed and pooled data from published studies. METHODS: Studies stratifying survival in CRC patients by MSI status were eligible for analysis. The principal outcome measure was the HR. Data from eligible studies were pooled using standard techniques. RESULTS: Thirty-two eligible studies reported survival in a total of 7,642 cases, including 1,277 with MSI. There was no evidence of publication bias. The combined HR estimate for overall survival associated with MSI was 0.65 (95% CI, 0.59 to 0.71; heterogeneity P = .16; I(2) = 20%). This benefit was maintained restricting analyses to clinical trial patients (HR = 0.69; 95% CI, 0.56 to 0.85) and patients with locally advanced CRC (HR = 0.67; 95% CI, 0.58 to 0.78). In patients treated with adjuvant fluorouracil (FU) CRCs with MSI had a better prognosis (HR = 0.72; 95% CI, 0.61 to 0.84). However, while data are limited, tumors with MSI derived no benefit from adjuvant FU (HR = 1.24; 95% CI, 0.72 to 2.14). CONCLUSION: CRCs with MSI have a significantly better prognosis compared to those with intact mismatch repair. Additional studies are needed to further define the benefit of adjuvant chemotherapy in locally advanced tumors with MSI. PMID- 15659509 TI - Proteasome inhibition as a novel therapeutic target in human cancer. AB - The 26S proteasome is a large intracellular adenosine 5'-triphosphate-dependent protease that identifies and degrades proteins tagged for destruction by the ubiquitin system. The orderly degradation of cellular proteins is critical for normal cell cycling and function, and inhibition of the proteasome pathway results in cell-cycle arrest and apoptosis. Dysregulation of this enzymatic system may also play a role in tumor progression, drug resistance, and altered immune surveillance, making the proteasome an appropriate and novel therapeutic target in cancer. Bortezomib (formerly known as PS-341) is the first proteasome inhibitor to enter clinical practice. It is a boronic aid dipeptide that binds directly with and inhibits the enzymatic complex. Bortezomib has recently shown significant preclinical and clinical activity in several cancers, confirming the therapeutic value of proteasome inhibition in human malignancy. It was approved in 2003 for the treatment of advanced multiple myeloma (MM), with approximately one third of patients with relapsed and refractory MM showing significant clinical benefit in a large clinical trial. Its mechanism of action is partly mediated through nuclear factor-kappa B inhibition, resulting in apoptosis, decreased angiogenic cytokine expression, and inhibition of tumor cell adhesion to stroma. Additional mechanisms include c-Jun N-terminal kinase activation and effects on growth factor expression. Several clinical trials are currently ongoing in MM as well as several other malignancies. This article discusses proteasome inhibition as a novel therapeutic target in cancer and focuses on the development, mechanism of action, and current clinical experience with bortezomib. PMID- 15659511 TI - Benign lesions in cancer patients: Case 2. Unusual association of osteosarcoma and sarcoidosis. PMID- 15659510 TI - Benign lesions in cancer patients: Case 1. Sarcoidosis after chemoradiation for head and neck cancer. PMID- 15659512 TI - Benign lesions in cancer patients: Case 3. Vitiligo after radiotherapy for breast cancer in a woman with depigmentation disorder. PMID- 15659513 TI - Humor and oncology. PMID- 15659515 TI - Antiapoptotic effect of growth factors in leukemia. PMID- 15659517 TI - Relationship of serum bilirubin to toxicity in patients with metastatic colorectal cancer treated with single-agent high-dose irinotecan. PMID- 15659518 TI - Intravenous iron and recombinant human erythropoietin in cancer patients. PMID- 15659520 TI - Re: Pooled analysis of fluorouracil-based adjuvant therapy of stage II and III colon cancer: who benefits and by how much? PMID- 15659523 TI - Neoadjuvant chemotherapy followed by extrapleural pneumonectomy in malignant pleural mesothelioma: is it safe? PMID- 15659524 TI - On the division of cortical cells into simple and complex types: a comparative viewpoint. AB - Hubel and Weisel introduced the concept of cells in cat primary visual cortex being partitioned into two categories: simple and complex. Subsequent authors have developed a quantitative measure to distinguish the two cell types based on the ratio between modulated responses at the stimulus frequency (F1) and unmodulated (F0) components of the spiking responses to drifting sinusoidal gratings. It has been shown that cells in anesthetized cat and monkey cortex have bimodal distributions of F1/F0 ratios. A clear local minimum or dip exists in the distribution at a ratio close to unity. Here we present a comparison of the distributions of the F1/F0 ratios between cells in the primary visual cortex of the eutherian cat and marsupial Tammar wallaby, Macropus eugenii. This is the first quantitative description of any marsupial cortex using the F1/F0 ratio and follows earlier papers showing that cells in wallaby cortex are tightly oriented and spatial frequency tuned. The results reveal a bimodal distribution in the wallaby F1/F0 ratios that is very similar to that found in the rat, cat, and monkey. Discussion focuses on the mechanisms that could lead to such similar cell distributions in animals with diverse behaviors and phylogenies. PMID- 15659525 TI - Effect of chronic inflammation on dorsal horn nociceptive neurons in aged rats. AB - To elucidate the effect of chronic inflammation on spinal nociceptive neurons in the elderly, we compared nocifensive behavior, peripheral inflammatory responses, and spinal dorsal horn neuronal activities between the aged (29-34 mo) and adult (7-12 mo) male rats after injection of complete Freund's adjuvant (CFA) into the hind paw. Aged rats exhibited a significantly lower mechanical paw withdrawal threshold before inflammation. However, after CFA injection mechanical allodynia developed in both adult and aged rats after CFA injection. The changes of foot temperature and thickness after CFA injection were greater and lasted longer in aged than in adult rats. Sets of 124 wide dynamic range (WDR) neurons (aged: 59, adult: 65) and 26 nociceptive specific (NS) neurons (aged: 13, adult: 13) were recorded from the lumber spinal dorsal horn. NS neurons from the inflamed adult rats showed significantly higher responses to noxious mechanical stimulation than those in aged rats, whereas WDR neurons from inflamed adult and aged rats were similar. Background activity of WDR neurons from the adult rats increased after CFA, whereas WDR neurons of aged rats and NS neurons from either group were not. The afterdischarge followed by noxious mechanical stimulation was significantly greater for WDR neurons in both adult and aged rats, whereas no significant differences were observed in NS neurons. Two days after CFA injection, Fos expression increased similarly in aged and adult rats. Thus the aged rats showed enhanced peripheral inflammatory responses to CFA injection with only a slight change in dorsal horn neuronal activity. Together with our previous finding that nociceptive neurons in aged rats exhibit hyperexcitability, these results suggest that the dorsal horn nociceptive system becomes sensitized with advancing age and its excitability cannot be further increased by inflammation. PMID- 15659526 TI - Interaction of visual and proprioceptive feedback during adaptation of human reaching movements. AB - People tend to make straight and smooth hand movements when reaching for an object. These trajectory features are resistant to perturbation, and both proprioceptive as well as visual feedback may guide the adaptive updating of motor commands enforcing this regularity. How is information from the two senses combined to generate a coherent internal representation of how the arm moves? Here we show that eliminating visual feedback of hand-path deviations from the straight-line reach (constraining visual feedback of motion within a virtual, "visual channel") prevents compensation of initial direction errors induced by perturbations. Because adaptive reduction in direction errors occurred with proprioception alone, proprioceptive and visual information are not combined in this reaching task using a fixed, linear weighting scheme as reported for static tasks not requiring arm motion. A computer model can explain these findings, assuming that proprioceptive estimates of initial limb posture are used to select motor commands for a desired reach and visual feedback of hand-path errors brings proprioceptive estimates into registration with a visuocentric representation of limb position relative to its target. Simulations demonstrate that initial configuration estimation errors lead to movement direction errors as observed experimentally. Registration improves movement accuracy when veridical visual feedback is provided but is not invoked when hand-path errors are eliminated. However, the visual channel did not exclude adjustment of terminal movement features maximizing hand-path smoothness. Thus visual and proprioceptive feedback may be combined in fundamentally different ways during trajectory control and final position regulation of reaching movements. PMID- 15659527 TI - beta-Amyloid peptides impair PKC-dependent functions of metabotropic glutamate receptors in prefrontal cortical neurons. AB - The metabotropic glutamate receptors (mGluRs) have been implicated in cognition, memory, and some neurodegenerative disorders, including the Alzheimer's disease (AD). To understand how the dysfunction of mGluRs contributes to the pathophysiology of AD, we examined the beta-amyloid peptide (Abeta)-induced alterations in the physiological functions of mGluRs in prefrontal cortical pyramidal neurons. Two potential targets of mGluR signaling involved in cognition, the GABAergic system and the N-methyl-d-aspartate (NMDA) receptor, were examined. Activation of group I mGluRs with (S)-3,5-dihydroxyphenylglycine (DHPG) significantly increased the spontaneous inhibitory postsynaptic current (sIPSC) amplitude, and this effect was protein kinase C (PKC) sensitive. Treatment with Abeta abolished the DHPG-induced enhancement of sIPSC amplitude. On the other hand, activation of group II mGluRs with (2R,4R)-4-aminopyrrolidine 2,4-dicarboxylate (APDC) significantly increased the NMDA receptor (NMDAR) mediated currents via a PKC-dependent mechanism, and Abeta treatment also diminished the APDC-induced potentiation of NMDAR currents. In Abeta-treated slices, both DHPG and APDC failed to activate PKC. These results indicate that the mGluR regulation of GABA transmission and NMDAR currents is impaired by Abeta treatment probably due to the Abeta-mediated interference of mGluR activation of PKC. This study provides a framework within which the role of mGluRs in normal cognitive functions and AD can be better understood. PMID- 15659528 TI - Perceptual continuity and the emergence of perceptual persistence in the ventral visual pathway. AB - Perceptual continuity is an important aspect of our experience of the visual world. In this study, we focus on an example of perceptual continuity involving the maintenance of figure-ground segregation despite the removal of binding cues that initiated the segregation. Fragmented line drawings of objects were superimposed on a background of randomly oriented lines. Global forms could be discriminated from the background based on differences in motion or differences in color/brightness. Furthermore, perception of a global form persisted after the binding cue had been removed. A comparison between the persistence of forms constructed from motion or color demonstrated that both forms produced persistence after the object defining cues were removed. Functional imaging showed a gradual increase in the persistence of brain activity in the lower visual areas (V1, V2, VP), which reached significance in V4v and peaked in the lateral occipital area. There was no difference in the location of persistence for color- or motion-defined forms. These results suggest that the retention of a global percept is an emerging property of the ventral visual processing stream and the maintenance of grouped visual elements is independent of cue type. We postulated that perceptual persistence depends on a system of perceptual memory reflecting the state of perceptual organization. PMID- 15659529 TI - Degradation of temporal resolution in the auditory midbrain after prolonged deafness is reversed by electrical stimulation of the cochlea. AB - In an animal model of prelingual deafness, we examined the anatomical and physiological effects of prolonged deafness and chronic electrical stimulation on temporal resolution in the adult central auditory system. Maximum following frequencies (Fmax) and first spike latencies of single neurons responding to electrical pulse trains were evaluated in the inferior colliculus of two groups of neonatally deafened cats after prolonged periods of deafness (>2.5 yr): the first group was implanted with an intracochlear electrode and studied acutely (long-deafened unstimulated, LDU); the second group (LDS) received a chronic implant and several weeks of electrical stimulation (pulse rates > or =300 pps). Acutely deafened and implanted adult cats served as controls. Spiral ganglion cell density in all long-deafened animals was markedly reduced (mean <5.8% of normal). Both long-term deafness and chronic electrical stimulation altered temporal resolution of neurons in the central nucleus (ICC) but not in the external nucleus. Specifically, LDU animals exhibited significantly poorer temporal resolution of ICC neurons (lower Fmax, longer response latencies) as compared with control animals. In contrast, chronic stimulation in LDS animals led to a significant increase in temporal resolution. Changes in temporal resolution after long-term deafness and chronic stimulation occurred broadly across the entire ICC and were not correlated with its tonotopic gradient. These results indicate that chronic electrical stimulation can reverse the degradation in temporal resolution in the auditory midbrain after long-term deafness and suggest the importance of factors other than peripheral pathology on plastic changes in the temporal processing capabilities of the central auditory system. PMID- 15659530 TI - Wiener kernels of chinchilla auditory-nerve fibers: verification using responses to tones, clicks, and noise and comparison with basilar-membrane vibrations. AB - Responses to tones, clicks, and noise were recorded from chinchilla auditory nerve fibers (ANFs). The responses to noise were analyzed by computing the zeroth , first-, and second-order Wiener kernels (h0, h1, and h2). The h1s correctly predicted the frequency tuning and phases of responses to tones of ANFs with low characteristic frequency (CF). The h2s correctly predicted the frequency tuning and phases of responses to tones of all ANFs, regardless of CF. Also regardless of CF, the kernels jointly predicted about 77% of the features of ANF responses to "frozen" samples of noise. Near-CF group delays of kernels and signal-front delays of responses to intense rarefaction clicks exceeded by 1 ms the corresponding basilar-membrane delays at both apical and basal sites of the chinchilla cochlea. This result, confirming that synaptic and neural processes amount to 1 ms regardless of CF, permitted drawing a map of basilar-membrane delay as a function of position for the entire length of the chinchilla cochlea, a first for amniotic species. PMID- 15659531 TI - Generalization of motor learning based on multiple field exposures and local adaptation. AB - Previous studies have used transfer of learning over workspace locations as a means to determine whether subjects code information about dynamics in extrinsic or intrinsic coordinates. Transfer has been observed when the torque associated with joint displacement is similar between workspace locations-rather than when the mapping between hand displacement and force is preserved-which is consistent with muscle- or joint-based encoding. In the present study, we address the generality of an intrinsic coding of dynamics and examine how generalization occurs when the pattern of torques varies over the workspace. In two initial experiments, we examined transfer of learning when the direction of a force field was fixed relative to an external frame of reference. While there were no beneficial effects of transfer after training at a single location (experiments 1 and 2), excellent performance was observed at the center of the workspace after training at two lateral locations (experiment 2). Experiment 3 and associated simulations assessed the characteristics of this generalization. In these studies, we examined the patterns of transfer observed after adaptation to force fields that were composed of two subfields that acted in opposite directions. The experimental and simulated data are consistent with the idea that information about dynamics is encoded in intrinsic coordinates. The nervous system generalizes dynamics learning by interpolating between sets of control signals, each locally adapted to different patterns of torques. PMID- 15659532 TI - Wiener-kernel analysis of responses to noise of chinchilla auditory-nerve fibers. AB - Responses to broadband Gaussian white noise were recorded in auditory-nerve fibers of deeply anesthetized chinchillas and analyzed by computation of zeroth-, first-, and second-order Wiener kernels. The first-order kernels (similar to reverse correlations or "revcors") of fibers with characteristic frequency (CF) <2 kHz consisted of lightly damped transient oscillations with frequency equal to CF. Because of the decay of phase locking strength as a function of frequency, the signal-to-noise ratio of first-order kernels of fibers with CFs >2 kHz decreased with increasing CF at a rate of about -18 dB per octave. However, residual first-order kernels could be detected in fibers with CF as high as 12 kHz. Second-order kernels, 2-dimensional matrices, reveal prominent periodicity at the CF frequency, regardless of CF. Thus onset delays, frequency glides, and near-CF group delays could be estimated for auditory-nerve fibers innervating the entire length of the chinchilla cochlea. PMID- 15659533 TI - Spontaneous activity and properties of two types of principal neurons from the ventral tegmental area of rat. AB - We investigated the spontaneous activity and properties of freshly isolated ventral tegmental area (VTA) principal neurons by whole cell recording and single cell RT-PCR. The VTA principal neurons, which were tyrosine hydroxylase-positive and glutamic acid decarboxylase (GAD67)-negative, exhibited low firing frequency and a long action potential (AP) duration. The VTA principal neurons exhibited a calretinin-positive and parvalbumin-negative Ca2+-binding protein mRNA expression pattern. The VTA principal neurons were classified into two subpopulations based on their firing frequency coefficient of variation (CV) at room temperature (21 23 degrees C): irregular-type neurons with a large CV and tonic-type neurons with a small CV. These two firing patterns were also recorded at the temperature of 34 degrees C and in nystatin-perforated patch recording. In VTA principal neurons, the AP afterhyperpolarization (AHP) amplitude contributed to the firing regularity and AHP decay slope contributed to the firing frequency. The AHP amplitude in the irregular-type VTA principal neurons was smaller than that in the tonic-type VTA principal neurons. There was no significant difference in the AHP decay slope between the two-types of VTA principal neurons. Apamin-sensitive small-conductance Ca2+-activated K+ (SK) channels contributed to the AHP and the regular firing of the tonic-type neurons but contributed little to the AHP and firing of the irregular-type neurons. In voltage-clamp tail-current analysis, in both conventional and nystatin-perforated whole cell recording, the apamin sensitive AHP current density of the tonic-type neurons was significantly larger than that of the irregular-type neurons. We suggest that apamin-sensitive SK current contributes to intrinsic firing differences between the two subpopulations of VTA principal neurons. PMID- 15659534 TI - Encoding of virtual acoustic space stimuli by neurons in ferret primary auditory cortex. AB - Recent studies from our laboratory have indicated that the spatial response fields (SRFs) of neurons in the ferret primary auditory cortex (A1) with best frequencies > or =4 kHz may arise from a largely linear processing of binaural level and spectral localization cues. Here we extend this analysis to investigate how well the linear model can predict the SRFs of neurons with different binaural response properties and the manner in which SRFs change with increases in sound level. We also consider whether temporal features of the response (e.g., response latency) vary with sound direction and whether such variations can be explained by linear processing. In keeping with previous studies, we show that A1 SRFs, which we measured with individualized virtual acoustic space stimuli, expand and shift in direction with increasing sound level. We found that these changes are, in most cases, in good agreement with predictions from a linear threshold model. However, changes in spatial tuning with increasing sound level were generally less well predicted for neurons whose binaural frequency-time receptive field (FTRF) exhibited strong excitatory inputs from both ears than for those in which the binaural FTRF revealed either a predominantly inhibitory effect or no clear contribution from the ipsilateral ear. Finally, we found (in agreement with other authors) that many A1 neurons exhibit systematic response latency shifts as a function of sound-source direction, although these temporal details could usually not be predicted from the neuron's binaural FTRF. PMID- 15659535 TI - Identification and characterization of embryonic stem cell-derived pacemaker and atrial cardiomyocytes. AB - The aim of this study was to identify and functionally characterize cardiac subtypes during early stages of development. For this purpose, transgenic embryonic stem cells were generated using the alpha-myosin heavy chain promoter driving the expression of the enhanced green fluorescent protein (EGFP). EGFP positive clusters of cells were first observed as early as 7 days of development, thus, even before the initiation of the contractile activity. Flow cytometry and single-cell fluorescence measurements evidenced large diversities of EGFP intensity. Patch-clamp experiments showed EGFP expression exclusively in pacemaker and atrial but not ventricular cells. The highest fluorescence intensities were detected in pacemaker-like cardiomyocytes. In accordance, multielectrode-array recordings of whole embryoid bodies confirmed that the pacemaker center coincided with strongly EGFP-positive areas. The cardiac subtypes displayed already at this early stage differential characteristics of electrical activity and ion channel expression. Thus, quantitation of the alpha myosin heavy chain driven reporter gene expression allows identification and functional characterization of early cardiac subtypes. PMID- 15659536 TI - Novel mechanism of brain soluble epoxide hydrolase-mediated blood pressure regulation in the spontaneously hypertensive rat. AB - The role of soluble epoxide hydrolase (sEH) in the central control of blood pressure (BP) has not been elucidated in spite of peripheral sEH overexpression being linked to hypertension. Thus, our objective was to investigate the involvement of brain sEH in BP control. sEH expression in the hypothalamus and brain stem, two cardioregulatory brain areas, was increased in the spontaneously hypertensive rat (SHR) compared to the Wistar Kyoto (WKY) rat. Inhibition of the enzyme by intracerebroventricular (icv) delivery of AUDA further increased both BP and heart rate (HR) by 32 +/- 6 mmHg and 54 +/- 10 bpm, respectively, (P<0.05) in the SHR. Analysis of waveform telemetry data revealed a decrease in spontaneous baroreceptor reflex gain following sEH inhibition, indicating the sustained increase in BP may be due to a decrease in baroreceptor reflex function. The hypertensive effect of sEH inhibition is likely a result of an increase in epoxyeicosatrienoic acid (EET)-mediated generation of ROS. This view is supported by the following: 1) Inhibition of EET formation attenuates AUDA induced increase in BP; 2) delivery of an EET agonist increases BP and HR in the WKY rat, and 3) inhibition of NAD(P)H oxidase by gp91ds-tat prevents AUDA-induced increases in BP and HR. Finally, electrophysiological studies demonstrate that AUDA increased neuronal firing rate exclusively in the SHR, an effect completely abolished by gp91ds-tat. These observations suggest that EETs and sEH inhibition are involved in increasing BP in the SHR. We suggest that an increased expression of sEH is a futile central nervous system response in protection against hypertension. PMID- 15659537 TI - Tumor necrosis factor alpha and phorbol 12-myristate-13-acetate down-regulate human 11beta-hydroxysteroid dehydrogenase type 2 through p50/p50 NF-kappaB homodimers and Egr-1. AB - The 11beta-hydroxysteroid dehydrogenase type 2 (11beta-HSD2) regulates access of 11beta-hydroxyglucocorticoids to the mineralocorticoid receptor by reducing the hydroxyl group of these steroids at position 11. Previous cell culture studies revealed that tumor necrosis factor-alpha (TNF-alpha) down-regulates 11beta-HSD2 activity. Here, we demonstrate that transgenic mice overexpressing TNF-alpha have decreased mRNA abundance and activity of 11beta-HSD2 in kidney tissue, indicating that this effect may occur also in vivo. The analysis of the transcriptional regulation of 11beta-HSD2 by TNF-alpha and phorbol 12-myristate-13-acetate (PMA) with in vivo genomic footprinting in human colon SW620 cells revealed stimulus dependent protein-DNA interactions in three promoter regions, kappaB1, Sp1/Egr 1I, and Sp1/Egr-1II. Chromatin immunoprecipitation and electrophoretic mobility shift assays demonstrated the relevance of NF-kappaB binding to kappaB1 and of Egr-1 binding to Sp1/Egr-1 sites for the PMA and TNF-alpha effect. We observed a temporal switch of binding to kappaB1 site from active p65/p50 heterodimers to inactive p50/p50 homodimers. TNF-alpha or PMA treatment for 24 h resulted in accumulation of p50 and decrease of p65 nuclear proteins. Overexpression of p50 inhibited HSD11B2 promoter activity and overexpression of Egr-1 inhibited transactivation of the HSD11B2 promoter by p65/p50. In conclusion, TNF-alpha and PMA down-regulate expression and activity of 11beta-HSD2 most likely by a coordinate binding of p50/p50 and Egr-1 to the HSD11B2 promoter. PMID- 15659538 TI - Functional interactions between 5-hydroxytryptamine receptors and the serotonin transporter in pulmonary arteries. AB - Pulmonary arterial 5-hydroxytryptamine (serotonin) (5-HT) transporter (SERT)-, 5 HT receptor expression, and 5-HT-induced vasoconstriction can be increased in pulmonary hypertension. These variables were studied in normoxic and hypoxic Fawn Hooded (FH) and Sprague-Dawley (SD) rats. Furthermore, we compared the functional effects of SERT inhibitors and 5-HT receptor antagonists against 5-HT-induced vasoconstriction of pulmonary arteries. SERT and 5-HT(1B) expression was greater in FH rat lungs than in SD rats, as was 5-HT-mediated vasoconstriction. The 5 HT(2A) receptor antagonist ketanserin and the 5-HT(1B) receptor antagonist SB224289 (1'-methyl-5-[[2'-methyl-4'-(5-methyl-1,2,4-oxadiazol-3-yl)biphenyl-4 yl]carbonyl]-2,3,6,7-tetrahydro-spiro-[furo] 2, 3-f]indole-3,4'-piperidine]) inhibited responses to 5-HT in all vessels. The combined 5-HT(1B) receptor/SERT antagonist LY393558 (1-[2-[4-(6-fluoro-1H-indol-3-yl)-3,6-dihydro-1(2H) pyridinyl]ethyl]-3-isopropyl-6-(methylsulfonyl)-3,4-dihydro-1H-2,1,3 benzothiadiazine-2,2-dioxide) was the most potent inhibitor of constriction in all vessels. SERT inhibitors citalopram and fluoxetine inhibited responses to 5 HT in SD vessels. However, these inhibitors potentiated responses to 5-HT in FH vessels. After exposure of rats to 2 weeks of hypoxia, there was increased 5-HT mediated vasoconstriction and a profound decrease in SERT expression in both the FH and SD rat lung. Accordingly, citalopram had no effect on 5-HT-induced constriction in SD rat vessels and markedly less effect in FH rat vessels. Ketanserin, SB224289, and LY393558 inhibited responses to 5-HT in all hypoxic rat vessels. LY393558 was the most potent antagonist, and there was synergy between the effects of fluoxetine and SB224289 when given simultaneously. The results suggest that, in FH rats, SERT inhibitors may increase pulmonary vasoconstriction, but this can be inhibited by simultaneous 5-HT(1B) receptor antagonism. There is synergy between the inhibitory effects of 5-HT(1B) receptor antagonists and SERT inhibitors on 5-HT-induced pulmonary vasoconstriction. PMID- 15659539 TI - Clozapine potentiation of N-methyl-D-aspartate receptor currents in the nucleus accumbens: role of NR2B and protein kinase A/Src kinases. AB - Clozapine is an atypical antipsychotic that has a unique clinical profile that distinguishes it from other typical and atypical antipsychotics. At present, the underlying mechanisms of action of clozapine are unclear. Recent studies in the field of schizophrenia suggest that compounds that potentiate N-methyl-d aspartate (NMDA) receptor function in the appropriate brain regions might be an effective antipsychotic agent. One relevant region in which NMDA receptors play a key role in mediating neurotransmission is the nucleus accumbens. Therefore, we investigated the regulation of NMDA receptor currents and excitatory postsynaptic currents (EPSCs) by clozapine in nucleus accumbens neurons. Whole-cell patch clamp recordings were performed in rat brain slices. We demonstrate that bath application of clozapine but not haloperidol or the selective 5-hydroxytryptamine 2A antagonist MDL100907 [(R)-(+)-alpha-(2,3-dimethoxyphenyl)-1-[2-(4-fluoro phenyl)ethyl]-4-piperidine methanol] induces a robust potentiation of NMDA-evoked currents and of glutamatergic EPSCs and that this potentiation is dependent on dopamine release and postsynaptic activation of D1 receptors. Furthermore, the effect of clozapine is selective for NR2B subtype-containing NMDA receptors and is blocked by the selective Src family kinase inhibitor PP2 [4-amino-5-(4 chlorophenyl)-7-(t-butyl)pyrazolo[3,4-d]pyrimidine] and the protein kinase A selective inhibitor N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide but not by the protein kinase C-selective inhibitor bisindolylmaleimide I. This effect of clozapine in the nucleus accumbens might underlie the unique clinical profile of this atypical antipsychotic and provides a basis for novel treatment approaches. PMID- 15659540 TI - Effect of early phase adjuvant arthritis on hepatic P450 enzymes and pharmacokinetics of verapamil: an alternative approach to the use of an animal model of inflammation for pharmacokinetic studies. AB - The objective of this study was to evaluate the suitability of the early phase of adjuvant arthritis (pre-AA) as a model of inflammation for pharmacokinetic studies. Pre-AA is associated with little or no pain and discomfort as compared with fully developed adjuvant arthritis. Pre-AA was induced in male Sprague Dawley rats with a tail base injection of Mycobacterium butyricum. Animals were monitored for symptoms of arthritis and levels of the proinflammatory mediators, serum nitrite, C-reactive protein (CRP), and tumor necrosis factor alpha (TNFalpha). On day 6, rats were administered single i.v. (2 mg/kg) or oral (20 mg/kg) doses of racemic verapamil, and S- and R-verapamil concentrations were determined by high-performance liquid chromatography. Hepatic cytochrome P450 (P450) content and verapamil protein binding were also measured. All experiments were carried out in both pre-AA and control rats. Serum nitrite, CRP, and TNFalpha levels were significantly elevated in pre-AA rats while signs of pain and arthritis were absent. Pre-AA also significantly elevated plasma concentrations of S- and R-verapamil after both i.v. and oral doses, due, likely, to decreased drug clearance. This was accompanied by a significant reduction in hepatic cytochrome P450, CYP3A, and CYP1A content as well as significantly reduced verapamil free fraction in pre-AA. The early phase of AA is marked by increased proinflammatory mediators and reduced verapamil clearance, as well as decreased hepatic P450 enzymes. Hence, pre-AA is a suitable model of inflammation for pharmacokinetic studies that avoids unnecessary exposure of animals to the pain and distress of fully developed adjuvant arthritis. PMID- 15659541 TI - A specific peptide inhibitor of the class B metallo-beta-lactamase L-1 from Stenotrophomonas maltophilia identified using phage display. AB - OBJECTIVES: In Gram-negative bacteria, resistance to beta-lactam antibiotics and to known inhibitors mediated by metallo-beta-lactamases is a major concern and a serious threat to public health. Since no clinically useful inhibitors are available against class B metallo-beta-lactamases, the aim of the study was to identify peptides as inhibitors. METHODS: The L-1 metalloenzyme from Stenotrophomonas maltophilia was cloned, over-expressed, purified to homogeneity and used in screening of peptide libraries by phage display with a selective and competitive biopanning assay. This was based upon the high affinity of L-1 for cefoxitin and its slow hydrolysis. RESULTS: From six peptides, the consensus sequence Cys-Val-His-Ser-Pro-Asn-Arg-Glu-Cys was identified as a promising inhibitor of L-1 hydrolytic activity. This peptide showed a mixed inhibition of L 1 with a K(i competitive) of 16 +/- 4 microM and a K(i uncompetitive) of 9 +/- 1 microM. The same peptide was prepared without flanking Cys residues and demonstrated no detectable inhibition of L-1 hydrolytic activity with nitrocefin as a substrate. These data confirmed the importance of the peptide conformation for the inhibition of L-1 hydrolytic activity. Further analysis revealed rescue by Zn2+ ions. The mixed inhibition indicated peptide binding near the active site of L-1 and blocking of zinc atoms for optimal conformation in the pocket of the active site. CONCLUSION: This is the first report of a peptide inhibitor for Class B metallo-beta-lactamases. It will be used as a lead to identify more potent small molecule inhibitors via peptidomimetics. PMID- 15659542 TI - Daptomycin for treating infected diabetic foot ulcers: evidence from a randomized, controlled trial comparing daptomycin with vancomycin or semi synthetic penicillins for complicated skin and skin-structure infections. AB - OBJECTIVES: The predominant pathogens causing diabetic foot infections are Gram positive cocci, many of which are now resistant to commonly prescribed antibiotics. Daptomycin is a new agent that is active against most Gram-positive pathogens. To compare the effectiveness of daptomycin against semi-synthetic penicillins or vancomycin, we analysed the subset of diabetic patients with an infected ulcer enrolled in two randomized, controlled investigator-blind trials of patients with complicated skin and soft-tissue infections presumptively caused by Gram-positive organisms. PATIENTS AND METHODS: Patients with a diabetic ulcer infection were prospectively stratified to ensure they were equally represented in the treatment groups, then randomized to either daptomycin [4 mg/kg every 24 h intravenously (iv)] or a pre-selected comparator (vancomycin or a semi-synthetic penicillin) for 7-14 days. RESULTS: Among 133 patients with a diabetic ulcer infection, 103 were clinically evaluable; 47 received daptomycin and 56 received a comparator. Most infections were monomicrobial, and Staphylococcus aureus was the predominant pathogen. Success rates for patients treated with daptomycin or the comparators were not statistically different for clinical (66% versus 70%, respectively; 95% CI, -14.4, 21.8) or microbiological (overall or by pathogen) outcomes. Both treatments were generally well tolerated, with most adverse events of mild to moderate severity. CONCLUSIONS: The clinical and microbiological efficacy and safety of daptomycin were similar to those of commonly used comparator antibiotics for treating infected diabetic foot ulcers caused by Gram positive pathogens. Daptomycin should be considered for treating these infections, especially those caused by resistant Gram-positive pathogens. PMID- 15659543 TI - Moxifloxacin inhibits cytokine-induced MAP kinase and NF-kappaB activation as well as nitric oxide synthesis in a human respiratory epithelial cell line. AB - BACKGROUND: We previously demonstrated that the quinolone moxifloxacin prevents Candida albicans pneumonitis and epithelial nuclear factor kappaB (NF-kappaB) nuclear translocation in immunosuppressed mice. OBJECTIVES: To explore the anti inflammatory effects of moxifloxacin directly on a lung epithelial cell line. METHODS: We studied the effect of clinically relevant concentrations of moxifloxacin (2.5-10 mg/L) on cytokine-induced activation of nitric oxide (NO) secretion, inducible NO synthase (iNOS) expression and the activation of signal transduction pathways of inflammation, NF-kappaB and the mitogen-activated protein kinases [extracellular signal-regulated kinases (ERK1/2) and C-Jun N terminal kinase (JNK)], in the A549 lung epithelial cell line. RESULTS: Stimulation with the cytokines interleukin-1beta(IL-1beta)/interferon-gamma (IFN gamma) increased NO up to 3.3-fold and moxifloxacin inhibited this up to 68% (P < 0.05). Similarly, the increase in iNOS levels was inhibited in cells pre-treated with moxifloxacin by up to 62%. IL-1beta stimulated a rapid increase in the activities of early intracellular signalling molecules, ERK1/2 and JNK. Moxifloxacin inhibited ERK1/2 by up to 100% and p-JNK activation by 100%. NF kappaB, as measured by electrophoretic mobility shift assay, was inhibited up to 72% by moxifloxacin. Western-blot analysis revealed that IL-1beta enhanced NF kappaB p65 and p50 proteins by 1.7- and 3.6-fold, respectively, whereas moxifloxacin inhibited the proteins by up to 60%. CONCLUSIONS: Moxifloxacin inhibits intracellular signalling, iNOS expression and NO secretion in a lung epithelial cell line. Future studies may uncover a primary site of quinolone immunomodulation, either upstream or at the cell membrane. Eventually, this quinolone might become an important therapy for inflammatory lung diseases. PMID- 15659544 TI - Inhibition of cardiac lipoprotein utilization by transgenic overexpression of Angptl4 in the heart. AB - To investigate the role of Angptl4, an inhibitor of lipoprotein lipase that is induced by >3-fold in the heart after rosiglitazone treatment, we generated transgenic mice that overexpress Angptl4 in the heart (MHC-Angptl4). We show that MHC-Angptl4 mice exhibit cardiac-restricted expression of the transgene and inhibition of cardiac lipoprotein lipase (LPL) activity. However, LPL activities in other tissues or that released into plasma by heparin are not affected. In addition, MHC-Angptl4 mice also exhibit hypertriglyceridemia after 6 h of fasting. We use echocardiography to show that MHC-Angptl4 mice develop left ventricular dysfunction. Comparison of the metabolic profiles of isolated working hearts demonstrates that cardiac impairment in MHC-Angptl4 mice is positively associated with decreased triglyceride (TG) utilization. When bred to transgenic mice that overexpress acyl-CoA synthetase in the heart, a strain that exhibits elevated cardiac TG accumulation, cardiac TG content in double transgenic mice is reversed to that of wild-type mice. Taken together, our data support the hypothesis that induction of Angptl4 in the heart inhibits lipoprotein-derived fatty acid delivery. This mouse model will be useful to elucidate the role of reduced fatty acid supply in the pathogenesis of heart failure and related disorders. PMID- 15659545 TI - The Abl/Arg substrate ArgBP2/nArgBP2 coordinates the function of multiple regulatory mechanisms converging on the actin cytoskeleton. AB - ArgBP2, and its brain-specific splice variant, nArgBP2, are interactors and substrates of Abl/Arg tyrosine kinases and of the ubiquitin ligase Cbl. They are members of a family of adaptor proteins that colocalize with actin on stress fibers and at cell-adhesion sites, including neuronal synapses. We show here that their NH2-terminal region, which contains a sorbin homology domain domain, interacts with spectrin, and we identify binding proteins for their COOH-terminal SH3 domains. All these binding partners participate in the regulation of the actin cytoskeleton. These include dynamin, synaptojanin, and WAVE isoforms, as well as WAVE regulatory proteins. At least two of the ArgBP2/nArgBP2 binding partners, synaptojanin 2B and WAVE2, undergo ubiquitination and Abl-dependent tyrosine phosphorylation. ArgBP2/nArgBP2 knockdown in astrocytes produces a redistribution of focal adhesion proteins and an increase in peripheral actin ruffles, whereas nArgBP2 overexpression produces a collapse of the actin cytoskeleton. Thus, ArgBP2/nArgBP2 is a scaffold protein that control the balance between adhesion and motility by coordinating the function of multiple signaling pathways converging on the actin cytoskeleton. PMID- 15659546 TI - Cellular transformation by the MSP58 oncogene is inhibited by its physical interaction with the PTEN tumor suppressor. AB - The PTEN (phosphatase and tensin homologue) tumor suppressor protein contains a single catalytic domain with both lipid and protein phosphatase activities. The remaining C-terminal half of the PTEN protein plays a role in its stability and is mutated in many clinical cancer samples. Here, we report that the PTEN C terminal domain physically interacts with the forkhead-associated domain of the oncogenic MSP58 protein and that this interaction requires PTEN Thr-366. We further show that while MSP58 transforms Pten-/- mouse embryo fibroblasts (MEFs), concurrent introduction of wild-type PTEN causes a dramatic reduction in the number of MSP58-induced transformed foci. This PTEN-mediated inhibition of cellular transformation requires physical interaction as evidenced by the failure of PTEN(T366A) point mutation (residing within the MSP58 interaction domain) to suppress MSP-58-driven transformation. These observations, together with the capacity of catalytically inactive PTEN mutant (G129R) to suppress MSP58 oncogenicity, support the view that the C-terminal region of PTEN directly provides a previously uncharacterized biological function in its ability to regulate cellular transformation. PMID- 15659547 TI - Receptor editing in peripheral B cell tolerance. AB - Receptor editing or secondary Ig gene rearrangement occurs in immature, autoreactive B cells to maintain self-tolerance. Here we show that nonspontaneously autoimmune mice immunized with a peptide mimetope of DNA develop peptide- and DNA-reactive antibodies. Antigen-specific B cells display a follicular B cell phenotype. As these cells move into the memory compartment, many express RAG protein and acquire expression of both kappa and lambda light chains. Thus, this study provides evidence for receptor editing occurring in a mature, antigen-activated B cell population. Because the receptor editing observed here occurred in an autoreactive response to antigen, it may function to maintain peripheral tolerance. PMID- 15659548 TI - Relationship between retroviral DNA-integration-site selection and host cell transcription. AB - Retroviral DNA integration occurs throughout the genome; however, local "hot spots" exist where a strong preference for certain sites over others are seen, and more global preferences associated with genes have been reported. Previous data from our laboratory suggested that there are fewer integration events into a DNA template when it is undergoing active transcription than when it is not. Because these data were generated by using a stably transfected foreign gene that was only weakly inducible, we have extended this observation by comparing integration events into a highly inducible endogenous gene under both induced and uninduced transcriptional states. To examine the influence of transcription on site selection directly, we analyzed the frequency and distribution of integration of avian retrovirus DNA into the metallothionein gene, before and after its induction to a highly sustained level of expression by addition of ZnSO4. We found a 6-fold reduction in integration events after 100-fold induction of transcription. This result implies that, despite an apparent preference for integration of retroviral DNA into transcribed regions of host DNA, high-level transcription can be inhibitory to the integration process. Several possible models for our observation are as follows. First, when a DNA template is undergoing active transcription, integration might be blocked by the RNA polymerase II complex because of steric hindrance. Alternatively, the integrase complex may require DNA to be in a double-stranded conformation, which would not be the case during active transcription. Last, transcription might lead to remodeling of chromatin into a structure that is less favorable for integration. PMID- 15659549 TI - The trans-Golgi network-associated human ubiquitin-protein ligase POSH is essential for HIV type 1 production. AB - HIV type 1 (HIV-1) was shown to assemble either at the plasma membrane or in the membrane of late endosomes. Now, we report an essential role for human ubiquitin ligase POSH (Plenty of SH3s; hPOSH), a trans-Golgi network-associated protein, in the targeting of HIV-1 to the plasma membrane. Small inhibitory RNA-mediated silencing of hPOSH ablates virus secretion and Gag plasma membrane localization. Reintroduction of native, but not a RING finger mutant, hPOSH restores virus release and Gag plasma membrane localization in hPOSH-depleted cells. Furthermore, expression of the RING finger mutant hPOSH inhibits virus release and induces accumulation of intracellular Gag in normal cells. Together, our results identify a previously undescribed step in HIV biogenesis and suggest a direct function for hPOSH-mediated ubiquitination in protein sorting at the trans Golgi network. Consequently, hPOSH may be a useful host target for therapeutic intervention. PMID- 15659550 TI - A PKR-like eukaryotic initiation factor 2alpha kinase from zebrafish contains Z DNA binding domains instead of dsRNA binding domains. AB - The double-stranded RNA (dsRNA)-dependent protein kinase (PKR) is induced as part of the IFN response in mammals and acts to shut down protein synthesis by the phosphorylation of eukaryotic initiation factor 2alpha (eIF2alpha). In fish, a PKR-like kinase activity has been detected, but the enzyme responsible has eluded characterization. Here, we describe a PKR-like kinase from zebrafish. Phylogenetic analysis shows that the C-terminal kinase domain is more closely related to the kinase domain of PKR than to any of the other three known eIF2alpha kinases. Surprisingly, instead of the two dsRNA binding domains found at the N terminus of PKR, there are two Zalpha domains. Zalpha domains specifically bind dsDNA and RNA in the left-handed Z conformation, often with high affinity. They have been found previously in two other IFN-inducible proteins, the dsRNA editing enzyme, ADAR1, and Z-DNA binding protein 1 (ZBP1), as well as in the poxvirus virulence factor, E3L. This previously undescribed kinase, designated PKZ (protein kinase containing Z-DNA binding domains), is transcribed constitutively at low levels and is highly induced after injection of poly(inosinic)-poly(cytidylic) acid, which simulates viral infection. Binding of Z-DNA by the Zalpha domain of PKZ was demonstrated by circular dichroism. PKZ inhibits translation in transfected cells; site-directed mutagenesis indicates that this inhibition depends on its catalytic activity. Identification of a gene combining Zalpha domains with a PKR-like kinase domain strengthens the hypothesis that the ability to bind left-handed nucleic acid plays a role in the host response to viruses. PMID- 15659551 TI - The landscape of genetic complexity across 5,700 gene expression traits in yeast. AB - Many studies have identified quantitative trait loci (QTLs) that contribute to continuous variation in heritable traits of interest. However, general principles regarding the distribution of QTL numbers, effect sizes, and combined effects of multiple QTLs remain to be elucidated. Here, we characterize complex genetics underlying inheritance of thousands of transcript levels in a cross between two strains of Saccharomyces cerevisiae. Most detected QTLs have weak effects, with a median variance explained of 27% for highly heritable transcripts. Despite the high statistical power of the study, no QTLs were detected for 40% of highly heritable transcripts, indicating extensive genetic complexity. Modeling of QTL detection showed that only 3% of highly heritable transcripts are consistent with single-locus inheritance, 17-18% are consistent with control by one or two loci, and half require more than five loci under additive models. Strikingly, analysis of parent and progeny trait distributions showed that a majority of transcripts exhibit transgressive segregation. Sixteen percent of highly heritable transcripts exhibit evidence of interacting loci. Our results will aid design of future QTL mapping studies and may shed light on the evolution of quantitative traits. PMID- 15659553 TI - Role of SDF1 chemokine in the development of lateral line efferent and facial motor neurons. AB - Most sensory systems are innervated by efferent neurons as well as by afferent neurons. The efferent innervation modulates the sensitivity of the receptor cells or of the sensory terminals. In the posterior lateral line system of the zebrafish, two efferent nuclei have been described in the hindbrain. Here we examine the development of the efferent neurons. We show that their axons are guided toward the target organ along the lateral line nerve while their cell bodies migrate posteriorward across rhombomeres to achieve their final position in rhombomeres 6/7. This migration depends on the SDF1 chemokine. We show that the migration of motor neurons of the facial nucleus from rhombomere 4 to 6 is also affected in sdf1a morphants (embryos injected with morpholine-conjugated antisense oligonucleotides). We propose that SDF1/CXCR4-mediated cell migration is preferentially associated with movement along the anteroposterior axis of the animal. PMID- 15659552 TI - Mu-opioid receptors modulate the stability of dendritic spines. AB - Opioids classically regulate the excitability of neurons by suppressing synaptic GABA release from inhibitory neurons. Here, we report a role for opioids in modulating excitatory synaptic transmission. By activating ubiquitously clustered mu-opioid receptor (MOR) in excitatory synapses, morphine caused collapse of preexisting dendritic spines and decreased synaptic alpha-amino-3-hydroxy-5 methyl-4-isoxazolepropionic acid receptors. Meanwhile, the opioid antagonist naloxone increased the density of spines. Chronic treatment with morphine decreased the density of dendritic spines even in the presence of Tetrodotoxin, a sodium channel blocker, indicating that the morphine's effect was not caused by altered activity in neural network through suppression of GABA release. The effect of morphine on dendritic spines was absent in transgenic mice lacking MORs and was blocked by CTOP (D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-ThrNH2), a mu-receptor antagonist. These data together with others suggest that endogenous opioids and/or constitutive activity of MORs participate in maintaining normal morphology and function of spines, challenging the classical model of opioids. Abnormal alteration of spines may occur in drug addiction when opioid receptors are overactivated by exogenous opiates. PMID- 15659554 TI - Sequence turnover and tandem repeats in cis-regulatory modules in drosophila. AB - The path by which regulatory sequence can change, yet preserve function, is an important open question for both evolution and bioinformatics. The recent sequencing of two additional species of Drosophila plus the wealth of data on gene regulation in the fruit fly provides new means for addressing this question. For regulatory sequences, indels account for more base pairs (bp) of change than substitutions (between Drosophila melanogaster and Drosophila yakuba), though they are fewer in number. Using Drosophila pseudoobscura as an out-group, we can distinguish insertions from deletions (with maximum parsimony criteria), and find a ratio between 1 and 5 (insertions to deletions) that is species dependent and much larger than the ratio of 1/8 for neutral sequences (Petrov and Hartl 1998). Because neutral sequence is rapidly cleared from the genome, most noncoding regions which preserve their length between D. melanogaster-D. pseudoobscura and have an excess of insertions over deletions should be functional. A fraction of 15%-18% (i.e., more than 20 standard deviations from random expectation) of the regulatory sequence is covered by low copy number tandem repeats whose repeating unit has an average length of 5-10 bp and which occur preferentially (25%-45% coverage) in indels. All indels may be due to tandem repeats if we extrapolate the detection efficiency of the repeat-finding algorithms using the observed point mutation rate between the species we compare. Sequence creation by local duplication accords with the tendency for multiple copies of transcription factor binding sites to occur in regulatory modules. Thus, indel events and tandem repeats in particular need to be incorporated into models of regulatory evolution because they can alter the rate at which beneficial variants arise and should also influence bioinformatic algorithms that parse regulatory sequences into binding sites. PMID- 15659555 TI - The origin and evolution of porcine reproductive and respiratory syndrome viruses. AB - Porcine reproductive and respiratory syndrome viruses (PRRSV) are divided into North American and European types, which show about 40% difference in their amino acid sequences. The divergence time of these two types has been estimated to be about 1980 from epidemiological data. This suggested that PRRSV have evolved at a higher evolutionary rate (order of 10(-2)/site/year) compared with other RNA viruses of 10(-3) to 10(-5)/site/year. Here, to test the evolutionary history of PRRSV speculated by the epidemiological background, we estimated the divergence time and evolutionary rate of PRRSV with molecular evolutionary analysis. Estimated divergence time (1972-1988) corresponded well to that estimated by the epidemiological data, and the evolutionary rate (4.71-9.8) x 10(-2) of PRRSV was indeed the highest among RNA viruses so far reported. Furthermore, we inferred important sites for the adaptation in order to examine how PRRSV have adapted to swine since they emerged. The adaptive sites were located not only in the epitopes related to immunity but also in the transmembrane regions including a signal peptide. In particular, the adaptive sites in the transmembrane regions were considered to affect compatibility to the host cell membrane. We conclude that PRRSV were transmitted from another host species to swine in about 1980 and have adapted to swine by altering the transmembrane regions. PMID- 15659556 TI - High copy number in human endogenous retrovirus families is associated with copying mechanisms in addition to reinfection. AB - There are at least 31 families of human endogenous retroviruses (HERVs), each derived from an independent infection by an exogenous virus. Using evidence of purifying selection on HERV genes, we have shown previously that reinfection by replication-competent elements was the predominant mechanism of copying in some families. Here we analyze the evolution of 17 HERV families using d(N)/d(S) ratios and find a positive relationship between copy number and the use of additional copying mechanisms. All families with more than 200 elements have also used one or more of the following mechanisms: (1) complementation in trans (elements copied by other elements of the same family; HERV-H and ERV-9), (2) retrotransposition in cis (elements copying themselves) within germ-line cells (HERV-K(HML3)), and (3) being copied by non-HERV machinery (HERV-W). We discuss why these other mechanisms are rare in most families and suggest why complementation in trans is significant only in the larger families. PMID- 15659557 TI - Complex spliceosomal organization ancestral to extant eukaryotes. AB - In higher eukaryotes, introns are spliced out of protein-coding mRNAs by the spliceosome, a massive complex comprising five non-coding RNAs (ncRNAs) and about 200 proteins. By comparing the differences between spliceosomal proteins from many basal eukaryotic lineages, it is possible to infer properties of the splicing system in the last common ancestor of extant eukaryotes, the eukaryotic ancestor. We begin with the hypothesis that, similar to intron length (that appears to have increased in multicellular eukaryotes), the spliceosome has increased in complexity throughout eukaryotic evolution. However, examination of the distribution of spliceosomal components indicates that not only was a spliceosome present in the eukaryotic ancestor but it also contained most of the key components found in today's eukaryotes. All the small nuclear ribonucleoproteins (snRNPs) protein components are likely to have been present, as well as many splicing-related proteins. Both major and trans-splicing are likely to have been present, and the spliceosome had already formed links with other cellular processes such as transcription and capping. However, there is no evidence as yet to suggest that minor (U12-dependent) splicing was present in the eukaryotic ancestor. Although the last common ancestor of extant eukaryotes appears to show much of the molecular complexity seen today, we do not, from this work, infer anything of the properties of the earlier "first eukaryote." PMID- 15659558 TI - Phosphotyrosine proteomic study of interferon alpha signaling pathway using a combination of immunoprecipitation and immobilized metal affinity chromatography. AB - Tyrosine phosphorylation is a type of post-translational modification that plays a crucial role in signal transduction. Thus, the study of this modification at the proteomic level has great biological significance. However, because of the low abundance of tyrosine-phosphorylated proteins in total cell lysate, it is difficult to evaluate the dynamics of tyrosine phosphorylation at a global level. In this work, proteins carrying phosphotyrosine (pTyr) were first purified from whole cell lysate by immunoprecipitation using anti-pTyr monoclonal antibodies. After tryptic digestion, phosphopeptides were further enriched by IMAC and analyzed by LC-MS. Quantitative changes of tyrosine phosphorylation at the global level were evaluated using isotopic labeling (introduced at the methyl esterification step prior to IMAC). Using this double enrichment approach, we characterized interferon alpha (IFNalpha)-induced pTyr proteomic changes in Jurkat cells. We observed induced phosphorylation on several well documented as well as novel tyrosine phosphorylation sites on proteins involved in IFNalpha signal transduction, such as Tyk2, JAK1, and IFNAR subunits. A specific site on alpha-tubulin (Tyr-271) was observed to be phosphorylated upon treatment as well. Furthermore, our results suggest that LOC257106, a CDC42 GAP-like protein, is potentially involved in this pathway. PMID- 15659559 TI - Heterogenous nuclear ribonucleoprotein F modulates angiotensinogen gene expression in rat kidney proximal tubular cells. AB - An insulin-responsive element (IRE) in the rat angiotensinogen (ANG) gene promoter that binds to two nuclear proteins with apparent molecular weights of 48 and 70 kD was identified previously from rat immortalized renal proximal tubular cells (IRPTC). The present studies aimed to identify and clone the 48-kD nuclear protein and to define its action on ANG gene expression. Nuclear proteins were isolated from IRPTC and subjected to two-dimensional electrophoresis. The 48-kD nuclear protein was detected by Southwestern blotting and subsequently identified by mass spectrometry, revealing that it was identical to 46-kD heterogeneous nuclear ribonucleoprotein F (hnRNP F), a nuclear protein that binds to TATA binding protein and associates with RNA polymerase II and also interacts with nuclear cap-binding complex. The hnRNP F cDNA was cloned from IRPTC by reverse transcriptase-PCR. Bacterially expressed recombinant hnRNP F bound to the rat ANG IRE, as revealed by gel mobility shift assay. The addition of polyclonal antibodies against hnRNP F yielded a supershift in gel mobility. Transient transfer of sense and antisense hnRNP F cDNA in IRPTC inhibited and enhanced ANG gene expression, respectively. High glucose stimulated and insulin inhibited hnRNP F expression in IRPTC. Expression studies indicated that hnRNP F is present in the kidney, testis, liver, lung, and brain but not in the spleen. In conclusion, these studies demonstrate that hnRNP F binds to rANG-IRE and modulates renal ANG gene expression, implicating that dysregulation of hnRNP F might affect renin-angiotensin system activation and, subsequently, kidney injury in diabetes. PMID- 15659560 TI - Establishing the role of angiotensin-converting enzyme in renal function and blood pressure control through the analysis of genetically modified mice. PMID- 15659561 TI - Suicide in the United States end-stage renal disease program. AB - Although depression and dialysis withdrawal are relatively common among individuals with ESRD, there have been few systematic studies of suicide in this population. The goals of this study were to compare the incidence of suicide with national rates and to contrast the factors associated with suicide with those associated with withdrawal in persons with ESRD. All individuals who were aged 15 yr and older and initiated dialysis between April 1, 1995, and November 30, 2000, composed the analytic cohort. Patients were censored at the time of death, transplantation, or October 31, 2001. Death as a result of suicide in the ESRD population and the general US population was ascertained from the Death Notification Form and the Centers for Disease Control and Prevention, respectively. Standardized incidence ratios for suicide among patient subgroups were computed using national data from the year 2000 as the reference population. The crude suicide rate from 1995 to 2001 was 24.2 suicides per 100,000 patient years, and the overall standardized incidence ratio for suicide was 1.84 (95% confidence interval, 1.50 to 2.27). In multivariable models, age > or =75 yr, male gender, white or Asian race, geographic region, alcohol or drug dependence, and recent hospitalization with mental illness were significant independent predictors of death as a result of suicide. Persons with ESRD are significantly more likely to commit suicide than persons in the general population. Although relatively rare, risk assessment can be used to identify patients for whom counseling and other interventions might be beneficial. PMID- 15659562 TI - Predictive performance of the modification of diet in renal disease and Cockcroft Gault equations for estimating renal function. AB - Recent recommendations emphasize the need to assess kidney function using creatinine-based predictive equations to optimize the care of patients with chronic kidney disease. The most widely used equations are the Cockcroft-Gault (CG) and the simplified Modification of Diet in Renal Disease (MDRD) formulas. However, they still need to be validated in large samples of subjects, including large non-U.S. cohorts. Renal clearance of (51)Cr-EDTA was compared with GFR estimated using either the CG equation or the MDRD formula in a cohort of 2095 adult Europeans (863 female and 1232 male; median age, 53.2 yr; median measured GFR, 59.8 ml/min per 1.73 m(2)). When the entire study population was considered, the CG and MDRD equations showed very limited bias. They overestimated measured GFR by 1.94 ml/min per 1.73 m(2) and underestimated it by 0.99 ml/min per 1.73 m(2), respectively. However, analysis of subgroups defined by age, gender, body mass index, and GFR level showed that the biases of the two formulas could be much larger in selected populations. Furthermore, analysis of the SD of the mean difference between estimated and measured GFR showed that both formulas lacked precision; the CG formula was less precise than the MDRD one in most cases. In the whole study population, the SD was 15.1 and 13.5 ml/min per 1.73 m(2) for the CG and MDRD formulas, respectively. Finally, 29.2 and 32.4% of subjects were misclassified when the CG and MDRD formulas were used to categorize subjects according to the Kidney Disease Outcomes Quality Initiative chronic kidney disease classification, respectively. PMID- 15659563 TI - Nephrotic plasma alters slit diaphragm-dependent signaling and translocates nephrin, Podocin, and CD2 associated protein in cultured human podocytes. AB - Podocytes are critical in maintaining the filtration barrier of the glomerulus and are dependent on the slit diaphragm (SD) proteins nephrin, podocin, and CD2 associated protein (CD2AP) to function optimally. The effects of normal human plasma and nephrotic plasma on podocytes were tested, focusing particularly on the SD complex. With the use of a conditionally immortalized human podocyte cell line, it first was shown that exposure to normal and non-nephrotic human plasma leads to a concentration of nephrin, podocin, CD2AP, and actin at the cell surface. Next, the effects of plasma from patients with nephrotic conditions to non-nephrotic conditions were compared. When exposed to all nephrotic plasma samples (and a non-human serum control), nephrin podocin and CD2AP assumed a cytoplasmic distribution; nephrin and synaptopodin were selectively downregulated, and the relocation of nephrin induced by nephrotic plasma could be rescued back to the plasma membrane by co-incubation with non-nephrotic plasma. Furthermore, intracellular calcium signaling was altered by nephrotic plasma, which was mediated by tyrosine kinase phosphorylation. With the use of nephrin mutant human cell lines, it was shown that this signaling and translocation response to normal plasma is nephrin dependent. This work demonstrates that nephrotic plasma seems to be deficient in factors that act via the podocyte SD complex, which are essential in maintaining its physiologic function. PMID- 15659564 TI - Expression and phenotype analysis of the nephrocystin-1 and nephrocystin-4 homologs in Caenorhabditis elegans. AB - Nephronophthisis (NPHP), an autosomal-recessive cystic kidney disease, is the most frequent genetic cause of end-stage renal failure in children. NPHP types 1 and 4 are caused by mutations in NPHP1 and NPHP4, encoding the proteins nephrocystin-1 and nephrocystin-4, respectively. Nephrocystin-1 and nephrocystin 4 are expressed in primary cilia of renal epithelial cells. NPHP1 and NPHP4 are highly conserved in Caenorhabditis elegans. However, this species does not have a kidney but an excretory system that consists of an excretory cell, an excretory gland cell, a duct cell, and a pore cell. Therefore, cell type-specific expression pattern and function of the nephrocystin homologs in C. elegans were of interest. Expression of green fluorescence protein fusion constructs that contain the C. elegans promoter regions for nph-1 and nph-4 was not found in the excretory system but in ciliated sensory neurons of the head (amphid neurons) and the tail in hermaphrodites (phasmid neurons) and males (sensory ray neurons). As the knockout phenotype for the PKD homologs lov-1 and pkd-2 shows impaired male mating behavior, RNAi knockdown animals were analyzed for this phenotype. A similar phenotype was found in the nph-1 and nph-4 RNAi knockdown animals compared with the lov-1 and pkd-2 knockout phenotype. Thus, it is suggested that renal cyst-causing genes may be part of a shared functional module, highly conserved in evolution. The NPHP homologs may be necessary for initial assembly of the cilium, whereas the polycystic kidney disease homologs may function as sensory transducers. PMID- 15659565 TI - Neuroanatomical targets of the organophosphate chlorpyrifos by c-fos immunolabeling. AB - Chlorpyrifos (CPF) is an organophosphate widely used as an insecticide in agriculture which elicits short- and long-term neurobehavioral deficits after acute administration. Because little is known about the specific brain areas targeted by CPF, investigating for the location of its neuroanatomical targets could help to describe the brain systems involved in the neurobehavioral toxicity developed in CPF-exposed organisms. To meet this objective, in the present study we evaluated CPF-induced c-fos expression. In addition, locomotor behavior and cerebral cholinesterase level were evaluated. We found two main sets of results. First, no significant c-fos expression was found in cholinoceptive regions in CPF treated rats 2 h or 24 h post-administration, despite the fact that 41% and 62% acetylcholinesterase inhibition, respectively, were present in brain homogenates. These results are consistent with previous reports showing CPF-induced activation of adaptive neural mechanisms re-establishing cholinergic tone. Second, 24 h post intoxication CPF elicited c-fos expression in cytokine-related areas. Cytokines have been involved in anxiety-like responses and psychiatric stress syndromes. Taking into account that CPF triggers the synthesis of peripheral cytokines, the present data stress the need to further clarify functional relations between organophosphate-triggered peripheral cytokines and emotional disturbances reported in intoxicated organisms. PMID- 15659566 TI - Validation of human physiologically based pharmacokinetic model for vinyl acetate against human nasal dosimetry data. AB - Vinyl acetate has been shown to induce nasal lesions in rodents in inhalation bioassays. A physiologically based pharmacokinetic (PBPK) model for vinyl acetate has been used in human risk assessment, but previous in vivo validation was conducted only in rats. Controlled human exposures to vinyl acetate were conducted to provide validation data for the application of the model in humans. Five volunteers were exposed to 1, 5, and 10 ppm 13C1,13C2 vinyl acetate via inhalation. A probe inserted into the nasopharyngeal region sampled both 13C1,13C2 vinyl acetate and the major metabolite 13C1,13C2 acetaldehyde during rest and light exercise. Nasopharyngeal air concentrations were analyzed in real time by ion trap mass spectrometry (MS/MS). Experimental concentrations of both vinyl acetate and acetaldehyde were then compared to predicted concentrations calculated from the previously published human model. Model predictions of vinyl acetate nasal extraction compared favorably with measured values of vinyl acetate, as did predictions of nasopharyngeal acetaldehyde when compared to measured acetaldehyde. The results showed that the current PBPK model structure and parameterization are appropriate for vinyl acetate. These analyses were conducted from 1 to 10 ppm vinyl acetate, a range relevant to workplace exposure standards but which would not be expected to saturate vinyl acetate metabolism. Risk assessment based on this model further concluded that 24 h per day exposures up to 1 ppm do not present concern regarding cancer or non-cancer toxicity. Validation of the vinyl acetate human PBPK model provides support for these conclusions. PMID- 15659567 TI - Inhibition of human and rat CYP1A2 by TCDD and dioxin-like chemicals. AB - Dioxins have been shown to bind and induce rodent CYP1A2, producing a dose dependent hepatic sequestration in vivo. The induction of CYP1A2 activity has been used as a noninvasive biomarker for human exposure to dioxins; while there is a consistent relationship between exposure and hepatic CYP1A2 induction in rodents, this relationship has only been observed in some of the highest exposed human populations. This may be explained by inhibition of CYP1A2 activity by dioxins as some rodent studies demonstrate that rodent CYP1A2 activity can in fact be inhibited by dioxins in vitro. CYP1A2 activity was examined using a series of dioxins to inhibit human and rat CYP1A2 activity in species-specific CYP1A2 SUPERSOMES using three common CYP1A2 substrates. Methoxyresorufin was a more efficient substrate than acetanalide or caffeine in this in vitro system. Rat and human CYP1A2 enzymatic activity is inhibited by TCDD, PCDD, TCDF, 4 PeCDF, and PCBs 126, 169, 105, 118, and 156 in a concentration-dependent manner. These data demonstrate that the in vitro metabolism of prototype substrates is similar between the rat and human CYP1A2 SUPERSOME preparations and that dioxins inhibit CYP1A2 activity in both species. Because of the potential for inhibition of CYP1A2 activity by TCDD and other dioxins, studies examining CYP1A2 induction in dioxin-exposed populations using these substrates should be viewed cautiously. PMID- 15659568 TI - Cytochrome P450 1A1 and 1B1 in human blood lymphocytes are not suitable as biomarkers of exposure to dioxin-like compounds: polymorphisms and interindividual variation in expression and inducibility. AB - Cytochrome P450 1A1 (CYP1A1) and 1B1 (CYP1B1) are phase I enzymes, the expression of which can be affected by many environmental compounds, including dioxins and dioxin-like compounds. Because CYP1A1 and CYP1B1 expression can easily be determined in peripheral blood lymphocytes, it is often suggested as biomarker of exposure to these compounds. In this study we investigated the interindividual differences in constitutive and induced CYP1A1-catalyzed ethoxyresorufin-O deethylase (EROD) activity and CYP1A1 and CYP1B1 gene expression in human blood lymphocytes in a group of ten non-smoking females. Freshly isolated lymphocytes were cultured in medium containing the mitogen PHA and were exposed to the most potent dioxin, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) or the less potent dioxin-like polychlorinated biphenyl 126 (PCB126). In addition, we determined the occurrence of the CYP1A1 MspI and CYP1B1 Leu432Val polymorphisms. All individuals showed a concentration-dependent increase of EROD activity by TCDD, which was significantly correlated with an increase in CYP1A1, but not CYP1B1 expression. The maximum induced EROD activity by TCDD was very different among the individuals, but the EC50 values were about the same. PCB126 also caused a concentration-dependent increase of EROD activity, but was a factor 100-1000 less potent than TCDD among the individuals. The allele frequencies for CYP1A1 MspI and CYP1B1 Leu432Val reflected a normal Caucasian population and in this study the polymorphisms had no apparent effect on the expression and activity of these enzymes. Our study shows a large interindividual variability in constitutive and induced EROD activity, and CYP1A1 and CYP1B1 expression in human lymphocytes. In addition, dioxin concentrations at which effects were observed in our in vitro study are about 10-fold higher than the human blood levels found in vivo, indicating that EROD activity and CYP1A1 and CYP1B1 expression in human lymphocytes might not be applicable as biomarkers of exposure to dioxin and dioxin-like compounds. PMID- 15659569 TI - Bisphenol a accelerates terminal differentiation of 3T3-L1 cells into adipocytes through the phosphatidylinositol 3-kinase pathway. AB - In order to identify whether bisphenol A (BPA) acts as an adipogenic agent, following the hormonal induction of differentiation into adipocytes, 3T3-L1 cells were treated for six days with BPA alone. Treatment with BPA increased the triacylglycerol (TG) content of the cultures, increased the percentage of Oil Red O-staining cells in the cultures, and increased the levels of lipoprotein lipase (LPL) and adipocyte-specific fatty acid binding protein (aP2) mRNAs. These findings indicate that BPA was able to accelerate terminal differentiation of 3T3 L1 cells into adipocytes. LY294002, a chemical inhibitor of phosphatidylinositol 3-kinase (PI 3-kinase), blocked completely the increasing effect of BPA on TG accumulation and expression of LPL and aP2 mRNAs. Western blot analysis revealed that BPA increased the level of phosphorylated Akt kinase. Based on these findings, we concluded that BPA acted through the PI 3-kinase and Akt kinase pathway, resulting in increased TG accumulation and expression of adipocyte genes. The structure-activity relationship for BPA-related chemicals was examined. Eight derivatives of BPA (three diphenylalkanes with different substituents at the central carbon atom, three diphenylalkanes with ester bonds on hydroxyl groups in the phenolic rings, one bisphenol consisting of a sulphur atom at the central position, one chemical with cyanic groups, instead of hydroxyl groups, in the phenolic rings) accelerated terminal adipocyte differentiation and their potencies to increase TG accumulation were 73-97% of that of BPA. Two diphenylalkanes with ether bonds on hydroxyl groups and two alkylphenols (4-nonylphenol and 4-tert-octylphenol) did not have the ability to accelerate terminal adipocyte differentiation. PMID- 15659570 TI - Subchronic exposure to low concentrations of di-n-butyl phthalate disrupts spermatogenesis in Xenopus laevis frogs. AB - Due to its widespread use and production, di-n-butyl phthalate (DBP) has become an environmental contaminant. It has been detected in a variety of environmental strata worldwide, including air, water, and soil. Also, monobutyl phthalate, the major metabolite of DBP, has been detected in a variety of human matrices. As a proven endocrine disruptive compound, DBP may contribute to global amphibian declines at much lower concentrations than tested thus far. We evaluated the effects of low concentrations of DBP on spermatogenesis in Xenopus laevis, the African clawed frog. Xenopus tadpoles were exposed to 0, 0.1, 0.5, 1.0, 5.0, or 10.0 ppm DBP, beginning at sexual differentiation (Nieuwkoop and Faber stage 52; 3 weeks of age) and continuing until 100% of controls metamorphosed (stage 66; 8 weeks of age). Upon necropsy at 33 weeks, 4-6% of DBP-treated frogs had only one testis, and 2-4% had retained oviducts. In all DBP treatment groups, seminiferous tubule diameter and the average number of germ cell nests per tubule were lower, and the number of tubules with no germ cells was significantly higher (p < 0.05). The percent of secondary spermatogonial cell nests significantly decreased (p < 0.05) in 1.0, 5.0, and 10.0 ppm groups. Several lesions occurred in DBP-exposed testes including denudation of germ cells, vacuolization of Sertoli cell cytoplasm, thickening of lamina propria of seminiferous tubules, and focal lymphocytic infiltration. Entire sections of testes containing almost exclusively mature spermatozoa were found in 1.0, 5.0, and 10.0 ppm DBP-exposed testes, indicating impairment of spermiation. Testicular hypoplasia and seminiferous tubular dysgenesis were also evident in DBP-treated frogs. Thus, subchronic exposure to low concentrations of DBP impairs spermatogenesis in Xenopus laevis frogs. PMID- 15659571 TI - Mycotoxin fumonisin B1 alters the cytokine profile and decreases the vaccinal antibody titer in pigs. AB - Fumonisin B1 (FB1), a mycotoxin produced by Fusarium verticillioides, may contaminate feed and food. In the present study, we investigated the effect of FB1 on the modulation of the cytokine profile and on the establishment of a vaccinal antibody response. In vitro investigations on pig peripheral blood mononuclear cells (PBMC) indicate that FB1 decreased interleukin-4 (IL-4) and increased interferon-gamma (IFN-gamma) synthesis at both the protein and mRNA levels. A short in vivo exposure (7 days) of weanling piglets to 1.5 mg/kg body weight of purified FB1 altered the cytokine balance in mesenteric lymph nodes and spleen similarly to the in vitro PBMC results. We also investigated the effect of FB1 on the antibody response during a vaccination process. A prolonged in vivo exposure (28 days) of weanling piglets to feed contaminated with 8 mg FB1/kg significantly decreased the expression of IL-4 mRNA by porcine whole blood cells and diminished the specific antibody titer after vaccination against Mycoplasma agalactiae. By contrast, ingestion of the contaminated feed had no effect on the serum concentration of the immunoglobulin subset (IgG, IgA, and IgM). Taken together, our data suggest that FB1 alters the cytokine profile and decreases the specific antibody response built during a vaccination protocol. These results may have implications for humans or animals eating contaminated food or feed. PMID- 15659572 TI - Essential role of extracellular matrix (ECM) overlay in establishing the functional integrity of primary neonatal rat Sertoli cell/gonocyte co-cultures: an improved in vitro model for assessment of male reproductive toxicity. AB - The development of in vitro models for testicular toxicity may provide important tools for investigating specific mechanisms of toxicity in the testis. Although various systems have been reported, their application in toxicological studies has been limited by the poor ability to replicate the complex biochemical, molecular, and functional interactions observed in the testis. In the present study, we evaluated a significantly improved Sertoli cell/gonocyte co-culture (SGC) system that employs a 3-dimensional extracellular matrix Matrigel (ECM) applied as an overlay instead of a substratum. We explored the dose- and time dependent effects of the addition of such an ECM overlay on cytoskeletal and morphological changes in the SGC system, and the resulting effects on cellular integrity. Furthermore, we correlated the latter effects with the ECM-dependent modulation of stress and survival signaling pathways and, most critically, the expression levels of the spermatogonia-specific protein, c-Kit. Finally, we applied this co-culture system to investigate the dose- and time-dependent effects on the morphology and induction of apoptosis of cadmium. We observed that the dose-dependent addition of an ECM overlay led to an enhanced attachment of Sertoli cells and facilitated the establishment of SGC communication and cytoskeletal structure, with a dramatic improvement in cell viability. The latter was consistent with the observed dose- and time-dependent modulation of both stress signaling pathways (SAPK/JNK) and survival signaling pathways (ERK and AKT) in the presence of the ECM overlay. Furthermore, the dose-dependent stabilization of c-Kit protein expression confirmed the functional integrity of this co-culture system. We conclude that this modified SGC system will provide investigators with a simple, efficient, and highly reproducible alternative in the screen for testicular cell-specific cytotoxicity and the assessment of molecular mechanisms associated with both normal development and reproductive toxicity induced by environmental toxicants. PMID- 15659573 TI - A simple method for predicting the functional differentiation of duplicate genes and its application to MIKC-type MADS-box genes. AB - A simple statistical method for predicting the functional differentiation of duplicate genes was developed. This method is based on the premise that the extent of functional differentiation between duplicate genes is reflected in the difference in evolutionary rate because the functional change of genes is often caused by relaxation or intensification of functional constraints. With this idea in mind, we developed a window analysis of protein sequences to identify the protein regions in which the significant rate difference exists. We applied this method to MIKC-type MADS-box proteins that control flower development in plants. We examined 23 pairs of sequences of floral MADS-box proteins from petunia and found that the rate differences for 14 pairs are significant. The significant rate differences were observed mostly in the K domain, which is important for dimerization between MADS-box proteins. These results indicate that our statistical method may be useful for predicting protein regions that are likely to be functionally differentiated. These regions may be chosen for further experimental studies. PMID- 15659574 TI - A molecular beacon, bead-based assay for the detection of nucleic acids by flow cytometry. AB - Molecular beacons are dual-labelled probes that are typically used in real-time PCR assays, but have also been conjugated with solid matrices for use in microarrays or biosensors. We have developed a fluid array system using microsphere-conjugated molecular beacons and the flow cytometer for the specific, multiplexed detection of unlabelled nucleic acids in solution. For this array system, molecular beacons were conjugated with microspheres using a biotin streptavidin linkage. A bridged conjugation method using streptavidin increased the signal-to-noise ratio, allowing for further discrimination of target quantitation. Using beads of different sizes and molecular beacons in two fluorophore colours, synthetic nucleic acid control sequences were specifically detected for three respiratory pathogens, including the SARS coronavirus in proof of-concept experiments. Considering that routine flow cytometers are able to detect up to four fluorescent channels, this novel assay may allow for the specific multiplex detection of a nucleic acid panel in a single tube. PMID- 15659576 TI - SNP identification in unamplified human genomic DNA with gold nanoparticle probes. AB - Single nucleotide polymorphisms (SNPs) comprise the most abundant source of genetic variation in the human genome. SNPs may be linked to genetic predispositions, frank disorders or adverse drug responses, or they may serve as genetic markers in linkage disequilibrium analysis. Thus far, established SNP detection techniques have utilized enzymes to meet the sensitivity and specificity requirements needed to overcome the high complexity of the human genome. Herein, we present for the first time a microarray-based method that allows multiplex SNP genotyping in total human genomic DNA without the need for target amplification or complexity reduction. This direct SNP genotyping methodology requires no enzymes and relies on the high sensitivity of the gold nanoparticle probes. Specificity is derived from two sequential oligonucleotide hybridizations to the target by allele-specific surface-immobilized capture probes and gene-specific oligonucleotide-functionalized gold nanoparticle probes. Reproducible multiplex SNP detection is demonstrated with unamplified human genomic DNA samples representing all possible genotypes for three genes involved in thrombotic disorders. The assay format is simple, rapid and robust pointing to its suitability for multiplex SNP profiling at the 'point of care'. PMID- 15659575 TI - Gene trap and gene inversion methods for conditional gene inactivation in the mouse. AB - Conditional inactivation of individual genes in mice using site-specific recombinases is an extremely powerful method for determining the complex roles of mammalian genes in developmental and tissue-specific contexts, a major goal of post-genomic research. However, the process of generating mice with recombinase recognition sequences placed at specific locations within a gene, while maintaining a functional allele, is time consuming, expensive and technically challenging. We describe a system that combines gene trap and site-specific DNA inversion to generate mouse embryonic stem (ES) cell clones for the rapid production of conditional knockout mice, and the use of this system in an initial gene trap screen. Gene trapping should allow the selection of thousands of ES cell clones with defined insertions that can be used to generate conditional knockout mice, thereby providing extensive parallelism that eliminates the time consuming steps of targeting vector construction and homologous recombination for each gene. PMID- 15659577 TI - Facile FMR1 mRNA structure regulation by interruptions in CGG repeats. AB - RNA metabolism is a major contributor to the pathogenesis of clinical disorders associated with premutation size alleles of the fragile X mental retardation (FMR1) gene. Herein, we determined the structural properties of numerous FMR1 transcripts harboring different numbers of both CGG repeats and AGG interruptions. The stability of hairpins formed by uninterrupted repeat containing transcripts increased with the lengthening of the repeat tract. Even a single AGG interruption in the repeated sequence dramatically changed the folding of the 5'UTR fragments, typically resulting in branched hairpin structures. Transcripts containing different lengths of CGG repeats, but sharing a common AGG pattern, adopted similar types of secondary structures. We postulate that interruption-dependent structure variants of the FMR1 mRNA contribute to the phenotype diversity, observed in premutation carriers. PMID- 15659579 TI - Ribosomal protein L1 recognizes the same specific structural motif in its target sites on the autoregulatory mRNA and 23S rRNA. AB - The RNA-binding ability of ribosomal protein L1 is of profound interest since the protein has a dual function as a ribosomal protein binding rRNA and as a translational repressor binding its mRNA. Here, we report the crystal structure of ribosomal protein L1 in complex with a specific fragment of its mRNA and compare it with the structure of L1 in complex with a specific fragment of 23S rRNA determined earlier. In both complexes, a strongly conserved RNA structural motif is involved in L1 binding through a conserved network of RNA-protein H bonds inaccessible to the solvent. These interactions should be responsible for specific recognition between the protein and RNA. A large number of additional non-conserved RNA-protein H-bonds stabilizes both complexes. The added contribution of these non-conserved H-bonds makes the ribosomal complex much more stable than the regulatory one. PMID- 15659578 TI - Conserved and specific functions of mammalian ssu72. AB - We describe the cloning and characterization of a human homolog of the yeast transcription/RNA-processing factor Ssu72, following a yeast two-hybrid screen for pRb-binding factors in the prostate gland. Interaction between hSsu72 and pRb was observed in transfected mammalian cells and involved multiple domains in pRb; however, so far, mutual effects of these two factors could not be demonstrated. Like the yeast counterpart, mammalian Ssu72 associates with TFIIB and the yeast cleavage/polyadenylation factor Pta1, and exhibits intrinsic phosphatase activity. Mammals contain a single ssu72 gene and a few pseudogenes. During mouse embryogenesis, ssu72 was highly expressed in the nervous system and intestine; high expression in the nervous system persisted in adult mice and was also readily observed in multiple human tumor cell lines. Both endogenous and ectopically expressed mammalian Ssu72 proteins resided primarily in the cytoplasm and only partly in the nucleus. Interestingly, fusion to a strong nuclear localization signal conferred nuclear localization only in a fraction of transfected cells, suggesting active tethering in the cytoplasm. Suppression of ssu72 expression in mammalian cells by siRNA did not reduce proliferation/survival, and its over-expression did not affect transcription of candidate genes in transient reporter assays. Despite high conservation, hssu72 was unable to rescue an ssu72 lethal mutation in yeast. Together, our results highlight conserved and mammalian specific characteristics of mammalian ssu72. PMID- 15659582 TI - Descriptive research is the bench science of nursing. PMID- 15659580 TI - hnRNP A2, a potential ssDNA/RNA molecular adapter at the telomere. AB - The heterogeneous nuclear ribonucleoprotein (hnRNP) A2 is a multi-tasking protein that acts in the cytoplasm and nucleus. We have explored the possibility that this protein is associated with telomeres and participates in their maintenance. Rat brain hnRNP A2 was shown to have two nucleic acid binding sites. In the presence of heparin one site binds single-stranded oligodeoxyribonucleotides irrespective of sequence but not the corresponding oligoribonucleotides. Both the hnRNP A2-binding cis-acting element for the cytoplasmic RNA trafficking element, A2RE, and the ssDNA telomere repeat match a consensus sequence for binding to a second sequence-specific site identified by mutational analysis. hnRNP A2 protected the telomeric repeat sequence, but not the complementary sequence, against DNase digestion: the glycine-rich domain was found to be necessary, but not sufficient, for protection. The N-terminal RRM (RNA recognition motif) and tandem RRMs of hnRNP A2 also bind the single-stranded, template-containing segment of telomerase RNA. hnRNP A2 colocalizes with telomeric chromatin in the subset of PML bodies that are a hallmark of ALT cells, reinforcing the evidence for hnRNPs having a role in telomere maintenance. Our results support a model in which hnRNP A2 acts as a molecular adapter between single-stranded telomeric repeats, or telomerase RNA, and another segment of ssDNA. PMID- 15659581 TI - Preferential binding and structural distortion by Fe2+ at RGGG-containing DNA sequences correlates with enhanced oxidative cleavage at such sequences. AB - Certain DNA sequences are known to be unusually sensitive to nicking via the Fe2+ mediated Fenton reaction. Most notable are a purine nucleotide followed by three or more G residues, RGGG, and purine nucleotides flanking a TG combination, RTGR. Our laboratory previously demonstrated that nicking in the RGGG sequences occurs preferentially 5' to a G residue with the nicking probability decreasing from the 5' to 3'end of these sequences. Using 1H NMR to characterize Fe2+ binding within the duplex CGAGTTAGGGTAGC/GCTACCCTAACTCG and 7-deazaguanine-containing (Z) variants of it, we show that Fe2+ binds preferentially at the GGG sequence, most strongly towards its 5' end. Substitutions of individual guanines with Z indicate that the high affinity Fe2+ binding at AGGG involves two adjacent guanine N7 moieties. Binding is accompanied by large changes in specific imino, aromatic and methyl proton chemical shifts, indicating that a locally distorted structure forms at the binding site that affects the conformation of the two base pairs 3' to the GGG sequence. The binding of Fe2+ to RGGG contrasts with that previously observed for the RTGR sequence, which binds Fe2+ with negligible structural rearrangements. PMID- 15659583 TI - Biographic and psychobehavioral influences on body mass index in a nursing sample. AB - The purpose of this study was to investigate the direct influences of age, ethnicity, education, and number of children and the indirect influences of codependency and binge eating on increased body mass index (BMI). In this secondary analysis, data were collected from a convenience sample of 511 nurses who completed a codependency instrument and a personal inventory profile assessing the predictors included in the current path analysis study. The five predictors retained in the final BMI model demonstrated that binge eating, age, and Black ethnicity were positively correlated with BMI and that Asian ethnicity was negatively correlated with BMI. The relationship between binge eating and BMI was augmented by codependency but reduced by Black ethnicity. It was concluded that the path analysis supported hypotheses that Black nurses were overweight for reasons other than binge eating and that participants who scored high on the codependency instrument were more likely to binge eat. PMID- 15659585 TI - An example of a successful research proposal: part I. PMID- 15659584 TI - Condom use predictors for low-income African American women. AB - The purpose was to investigate whether an HIV-prevention curriculum and a health maintenance curriculum produced different rates of change in reported consistent condom use and to explore what mediating variables predicted reported consistent condom use over time. A longitudinal crossover research design with extended posttest observations was used with a sample of 279 African American women. After pretesting, women received the designated curriculum and were posttested at completion, 3, 6, and 9 months. After the 9-month posttest, the curriculum was switched, and the above procedure was repeated. Using SAS macro GLIMMIX, the data revealed that the HIV curriculum yielded higher consistent condom use than did the health maintenance curriculum and that the mediating variables that predicted consistent condom use over time were self-efficacy for low-risk HIV behavior, HIV related community behavior, and social norms. Enhancing consistent condom use over time may require the promotion and reinforcement of these mediating variables. PMID- 15659586 TI - Gendered elder care exchanges in a Caribbean village. AB - Caregiver gender research often translates into caregiver accounts of the care they provide for an elder parent or spouse, limiting an understanding of other community members'involvement and the actual gendered elder care activities performed and caregiver benefits received. This article describes women's and men's actual involvement in elder care. These findings are part of a four-phase ethnographic community study that includes in-depth network analysis of elder households. One-hundred and forty-three informal adult caregivers assisted 15 elders in obtaining the things they needed to live, providing 244 care activities. One-hundred and three women provided 194 care activities, and 40 men performed 50 care activities. In exchange, the frail elders gave the women caregivers 132 benefits and the men 54 benefits. Women and men differed in the amount and type of care involvement, as well as their motivational mechanisms for involvement in elder care. PMID- 15659587 TI - Effects of a physical activity intervention for women. AB - Physical activity is associated with health and reduced mortality risk, yet only 15% of U.S. adults achieve adequate activity. This study is an experimental repeated measures nested design randomizing two similar rural communities to investigate the effectiveness of the Heart and Soul Physical Activity Program (HSPAP) (Peterson, 2002) in promoting physical activity in midlife women (n=42) aged 35 to 65 years. The HSPAP, an innovative church-based health promotion intervention, is conceptualized in social support and designed to increase physical activity, energy expenditure (EE), and cardiorespiratory fitness (VO2 max), measured over time. A significant interaction (p<.001) was found for EE in one HSPAP group increasing their EE by 1,010 kcals/week. HSPAP participants increased their VO2 max level by 75% (p<.001) and 10%; comparison groups stayed the same or declined 16%. Study results provide preliminary support for the HSPAP intervention as an effective treatment to improve physical activity levels in sedentary, rural, midlife women. PMID- 15659588 TI - European nursing research and its contribution to nursing knowledge development. PMID- 15659589 TI - Synaptic targeting of retrogradely transported trophic factors in motoneurons: comparison of glial cell line-derived neurotrophic factor, brain-derived neurotrophic factor, and cardiotrophin-1 with tetanus toxin. AB - Glial cell line-derived neurotrophic factor (GDNF), brain-derived neurotrophic factor (BDNF), and cardiotrophin-1 (CT-1) are the most potent neurotrophic factors for motoneurons, but their fate after retrograde axonal transport is not known. Internalized trophic factors may be degraded, or they may be recycled and transferred to other neurons, similar to the known route of tetanus toxin. We tested whether neonatal rat hypoglossal motoneurons target retrogradely transported trophic factors to synaptic sites on their dendrites within the brainstem and subsequently transfer these trophins across the synaptic cleft to afferent synapses (transsynaptic transcytosis). Motoneurons retrogradely transport from the tongue radiolabeled GDNF, BDNF, and CT-1 as well as tetanus toxin. Quantitative autoradiographic electron microscopy showed that GDNF and BDNF were transported into motoneuron dendrites with labeling densities similar to those of tetanus toxin. Although tetanus toxin accumulated rapidly (within 8 h) at presynaptic sites, GDNF accumulated at synapses more slowly (within 15 h), and CT-1 never associated with synapses. Thus, some retrogradely transported neurotrophic factors are trafficked similarly but not identically to tetanus toxin. Both GDNF and BDNF accumulate at the external (limiting) membrane of multivesicular bodies within proximal dendrites. We conclude that tetanus toxin, GDNF, and BDNF are released from postsynaptic sites and are internalized by afferent presynaptic terminals, thus demonstrating transsynaptic transcytosis. CT 1, however, follows a strict degradation pathway after retrograde transport to the soma. Synaptic and transcytotic trafficking thus are restricted to particular neurotrophic factors such as GDNF and BDNF. PMID- 15659590 TI - Linking out-of-body experience and self processing to mental own-body imagery at the temporoparietal junction. AB - The spatial unity of self and body is challenged by various philosophical considerations and several phenomena, perhaps most notoriously the "out-of-body experience" (OBE) during which one's visual perspective and one's self are experienced to have departed from their habitual position within one's body. Although researchers started examining isolated aspects of the self, the neurocognitive processes of OBEs have not been investigated experimentally to further our understanding of the self. With the use of evoked potential mapping, we show the selective activation of the temporoparietal junction (TPJ) at 330-400 ms after stimulus onset when healthy volunteers imagined themselves in the position and visual perspective that generally are reported by people experiencing spontaneous OBEs. Interference with the TPJ by transcranial magnetic stimulation (TMS) at this time impaired mental transformation of one's own body in healthy volunteers relative to TMS over a control site. No such TMS effect was observed for imagined spatial transformations of external objects, suggesting the selective implication of the TPJ in mental imagery of one's own body. Finally, in an epileptic patient with OBEs originating from the TPJ, we show partial activation of the seizure focus during mental transformations of her body and visual perspective mimicking her OBE perceptions. These results suggest that the TPJ is a crucial structure for the conscious experience of the normal self, mediating spatial unity of self and body, and also suggest that impaired processing at the TPJ may lead to pathological selves such as OBEs. PMID- 15659591 TI - Endogenous Ca2+ buffer concentration and Ca2+ microdomains in hippocampal neurons. AB - Ca2+-binding proteins are ubiquitously expressed throughout the CNS and serve as valuable immunohistochemical markers for certain types of neurons. However, the functional role of most Ca2+-binding proteins has to date remained obscure because their concentration in central neurons is not known. In this study, we investigate the intracellular concentration of the widely expressed Ca2+-binding protein calbindin-D28k in adult hippocampal slices using patch-clamp recordings and immunohistochemistry. First, we show that calbindin-D28k freely exchanges between patch pipette and cytoplasm during whole cell patch-clamp recordings with a time constant of approximately 10 min. Substituting known concentrations of recombinant calbindin-D28k in patch pipettes enabled us to determine the endogenous calbindin-D28k concentration by postrecording immunohistochemistry. Using this calibration procedure, we find that mature granule cells (doublecortin ) contain approximately 40 microm, and newborn granule cells (doublecortin+) contain 0-20 microm calbindin-D28k. CA3 stratum radiatum interneurons and CA1 pyramidal cells enclose approximately 47 and approximately 45 microm calbindin D28k, respectively. Numerical simulations showed that 40 microm calbindin-D28k is capable of tuning Ca2+ microdomains associated with action potentials at the mouth of single or clustered Ca2+ channels: calbindin-D28k reduces the increment in free Ca2+ at a distance of 100 and 200 nm by 20 and 35%, respectively, and strongly accelerates the collapse of the Ca2+ gradient after cessation of Ca2+ influx. These data suggest that calbindin-D28k equips hippocampal neurons with approximately 160 microm mobile, high-affinity Ca2+-binding sites (kappa(S) approximately 200) that slow and reduce global Ca2+ signals while they enhance the spatiotemporal fidelity of submicroscopic Ca2+ signals. PMID- 15659592 TI - Deletion of connexin45 in mouse retinal neurons disrupts the rod/cone signaling pathway between AII amacrine and ON cone bipolar cells and leads to impaired visual transmission. AB - Connexin45 (Cx45) is known to be expressed in the retina, but its functional analysis was problematic because general deletion of Cx45 coding DNA resulted in cardiovascular defects and embryonic lethality at embryonic day 10.5. We generated mice with neuron-directed deletion of Cx45 and concomitant activation of the enhanced green fluorescent protein (EGFP). EGFP labeling was observed in bipolar, amacrine, and ganglion cell populations. Intracellular microinjection of fluorescent dyes in EGFP-labeled somata combined with immunohistological markers revealed Cx45 expression in both ON and OFF cone bipolar cells. The scotopic electroretinogram of mutant mice revealed a normal a-wave but a 40% reduction in the b-wave amplitude, similar to that found in Cx36-deficient animals, suggesting a possible defect in the rod pathway of visual transmission. Indeed, neurotransmitter coupling between AII amacrine cells and Cx45-expressing cone bipolar cells was disrupted in Cx45-deficient mice. These data suggest that both Cx45 and Cx36 participate in the formation of functional heterotypic electrical synapses between these two types of retinal neurons that make up the major rod pathway. PMID- 15659593 TI - Chronic cocaine administration switches corticotropin-releasing factor2 receptor mediated depression to facilitation of glutamatergic transmission in the lateral septum. AB - Corticotropin-releasing factor (CRF) and urocortin (Ucn I) are endogenous members among a family of CRF-related peptides that activate two different and synaptically localized G-protein-coupled receptors, CRF1 and CRF2. These peptides and their receptors have been implicated in stress responses and stress with cocaine abuse. In this study, we observed significant alterations in excitatory transmission and CRF-related peptide regulation of excitatory transmission in the lateral septum mediolateral nucleus (LSMLN) after chronic cocaine administration. In brain slice recordings from the LSMLN of control (saline-treated) rats, glutamatergic synaptic transmission was facilitated by activation of CRF1 receptors with CRF but was depressed after activation of CRF2 receptors with Ucn I. After acute withdrawal from a chronic cocaine administration regimen, CRF1 activation remained facilitatory, but CRF2 activation facilitated rather than depressed LSMLN EPSCs. These alterations in CRF2 effects occurred through both presynaptic and postsynaptic mechanisms. In saline-treated rats, CRF1 and CRF2 coupled predominantly to protein kinase A signaling pathways, whereas after cocaine withdrawal, protein kinase C activity was more prominent and likely contributed to the CRF2-mediated presynaptic facilitation. Neither CRF nor Ucn I altered monosynaptic GABA(A)-mediated IPSCs before or after chronic cocaine administration, suggesting that loss of GABAA-mediated inhibition could not account for the facilitation. This switch in polarity of Ucn I-mediated neuromodulation, from a negative to positive regulation of excitatory glutamatergic transmission after chronic cocaine administration, could generate an imbalance in the brain reward circuitry associated with the LSMLN. PMID- 15659594 TI - Membrane receptors involved in modulation of responses of spinal dorsal horn interneurons evoked by feline group II muscle afferents. AB - Modulatory actions of a metabotropic 5-HT1A&7 membrane receptor agonist and antagonist [(+/-)-8-hydroxy-2-(di-n-propylamino)-tetralin; N-[2-[4-(2 methoxyphenyl)-1-piperazinyl]ethyl]-N-(2-pyridinyl) cyclohexane-carboxamide] and an ionotropic 5-HT3 membrane receptor agonist and antagonist [2-methyl-serotonin (2-Me 5-HT); N-(1-azabicyclo[2.2.2]oct-3-yl)-6-chloro-4-methyl-3-oxo-3,4-dihydro 2H-1,4-benzoxazine-8-carboxamide hydrochloride] were investigated on dorsal horn interneurons mediating reflex actions of group II muscle afferents. All drugs were applied ionophoretically in deeply anesthetized cats. Effects of agonists were tested on extracellularly recorded responses of individual interneurons evoked by electrical stimulation of group II afferents in a muscle nerve. Effects of antagonists were tested against the depression of these responses after stimulation of raphe nuclei. The results show that both 5-HT1A&7 and 5-HT3 membrane receptors are involved in counteracting the activation of dorsal horn interneurons by group II afferents. Because only quantitative differences were found within the sample of the tested neurons, these results suggest that modulatory actions of 5-HT on excitatory and inhibitory interneurons might be similar. The relationship between 5-HT axons and axons immunoreactive for the 5 HT3A receptor subunit, which contact dorsal horn interneurons, was analyzed using immunofluorescence and confocal microscopy. Contacts from both types of axons were found on all interneurons, but their distribution and density varied, and there was no obvious relationship between them. In two of six interneurons, 5 HT3A-immunoreactive axons formed ring-like arrangements around the cell bodies. In previous studies, axons possessing 5-HT3 receptors were found to be excitatory, and as 2-Me 5-HT depressed transmission to dorsal horn interneurons, the results indicate that 5-HT operates at 5-HT3 receptors presynaptic to these neurons to depress excitatory transmission. PMID- 15659596 TI - Responses of human anterior cingulate cortex microdomains to error detection, conflict monitoring, stimulus-response mapping, familiarity, and orienting. AB - Human anterior cingulate cortex (ACC) activity modulation has been observed in numerous tasks, consistent with a wide variety of functions. However, previous recordings have not had sufficient spatial resolution to determine whether microdomains (approximately one to two columns) are involved in multiple tasks, how activity is distributed across cortical layers, or indeed whether modulation reflected neuronal excitation, inhibition, or both. In this study, linear arrays of 24 microelectrodes were used to estimate population synaptic currents and neuronal firing in different layers of ACC during simple/choice reaction time, delayed word recognition, rhyming, auditory oddball, and cued conditional letter discrimination tasks. Responses to all tasks, with differential responses to errors, familiarity, difficulty, and orienting, were recorded in single microdomains. The strongest responses occurred approximately 300-800 ms after stimulus onset and were usually a current source with inhibited firing, strongly suggesting active inhibition in superficial layers during the behavioral response period. This was usually followed by a sink from approximately 800 to 1400 ms, consistent with postresponse rebound activation. Transient phase locking of task related theta activity in superficial cingulate layers suggested extended interactions with medial and lateral frontal and temporal sites. These data suggest that each anterior cingulate microdomain participates in a multilobar cortical network after behavioral responses in a variety of tasks. PMID- 15659597 TI - Reorganization of visual processing in macular degeneration. AB - Macular degeneration (MD), the leading cause of visual impairment in the developed world, damages the central retina, often obliterating foveal vision and severely disrupting everyday tasks such as reading, driving, and face recognition. In such cases, the macular damage eliminates the normal retinal input to a large region of visual cortex, comprising tens of square centimeters of surface area in each hemisphere, which is normally responsive only to foveal stimuli. Using functional magnetic resonance imaging, we asked whether this deprived cortex simply becomes inactive in subjects with MD, or whether it takes on new functional properties. In two adult MD subjects with extensive bilateral central retinal lesions, we found that parts of visual cortex (including primary visual cortex) that normally respond only to central visual stimuli are strongly activated by peripheral stimuli. Such activation was not observed (1) with visual stimuli presented to the position of the former fovea and (2) in control subjects with visual stimuli presented to corresponding parts of peripheral retina. These results demonstrate large-scale reorganization of visual processing in MD and will likely prove important in any effort to develop new strategies for rehabilitation of MD subjects. PMID- 15659595 TI - Distinct gamma2 subunit domains mediate clustering and synaptic function of postsynaptic GABAA receptors and gephyrin. AB - Modulation of the concentration of postsynaptic GABA(A) receptors contributes to functional plasticity of inhibitory synapses. The gamma2 subunit of GABA(A) receptor is specifically required for clustering of these receptors, for recruitment of the submembrane scaffold protein gephyrin to postsynaptic sites, and for postsynaptic function of GABAergic inhibitory synapses. To elucidate this mechanism, we here have mapped the gamma2 subunit domains required for restoration of postsynaptic clustering and function of GABA(A) receptors in gamma2 subunit mutant neurons. Transfection of gamma2-/- neurons with the gamma2 subunit but not the alpha2 subunit rescues postsynaptic clustering of GABA(A) receptors, results in recruitment of gephyrin to postsynaptic sites, and restores the amplitude and frequency of miniature inhibitory postsynaptic currents to wild type levels. Analogous analyses of chimeric gamma2/alpha2 subunit constructs indicate, unexpectedly, that the fourth transmembrane domain of the gamma2 subunit is required and sufficient for postsynaptic clustering of GABA(A) receptors, whereas cytoplasmic gamma2 subunit domains are dispensable. In contrast, both the major cytoplasmic loop and the fourth transmembrane domain of the gamma2 subunit contribute to efficient recruitment of gephyrin to postsynaptic receptor clusters and are essential for restoration of miniature IPSCs. Our study points to a novel mechanism involved in targeting of GABA(A) receptors and gephyrin to inhibitory synapses. PMID- 15659599 TI - Exacerbation of cerebral amyloid angiopathy-associated microhemorrhage in amyloid precursor protein transgenic mice by immunotherapy is dependent on antibody recognition of deposited forms of amyloid beta. AB - Passive immunization with an antibody directed against the N terminus of amyloid beta (Abeta) has recently been reported to exacerbate cerebral amyloid angiopathy (CAA)-related microhemorrhage in a transgenic animal model. Although the mechanism responsible for the deleterious interaction is unclear, a direct binding event may be required. We characterized the binding properties of several monoclonal anti-Abeta antibodies to deposited Abeta in brain parenchyma and CAA. Biochemical analyses demonstrated that the 3D6 and 10D5, two N-terminally directed antibodies, bound with high affinity to deposited forms of Abeta, whereas 266, a central domain antibody, lacked affinity for deposited Abeta. To determine whether 266 or 3D6 would exacerbate CAA-associated microhemorrhage, we treated aged PDAPP mice with either antibody for 6 weeks. We observed an increase in both the incidence and severity of CAA-associated microhemorrhage when PDAPP transgenic mice were treated with the N-terminally directed 3D6 antibody, whereas mice treated with 266 were unaffected. These results may have important implications for future immune-based therapeutic strategies for Alzheimer's disease. PMID- 15659598 TI - Glial cell line-derived neurotrophic factor mediates the desirable actions of the anti-addiction drug ibogaine against alcohol consumption. AB - Alcohol addiction manifests as uncontrolled drinking despite negative consequences. Few medications are available to treat the disorder. Anecdotal reports suggest that ibogaine, a natural alkaloid, reverses behaviors associated with addiction including alcoholism; however, because of side effects, ibogaine is not used clinically. In this study, we first characterized the actions of ibogaine on ethanol self-administration in rodents. Ibogaine decreased ethanol intake by rats in two-bottle choice and operant self-administration paradigms. Ibogaine also reduced operant self-administration of ethanol in a relapse model. Next, we identified a molecular mechanism that mediates the desirable activities of ibogaine on ethanol intake. Microinjection of ibogaine into the ventral tegmental area (VTA), but not the substantia nigra, reduced self-administration of ethanol, and systemic administration of ibogaine increased the expression of glial cell line-derived neurotrophic factor (GDNF) in a midbrain region that includes the VTA. In dopaminergic neuron-like SHSY5Y cells, ibogaine treatment upregulated the GDNF pathway as indicated by increases in phosphorylation of the GDNF receptor, Ret, and the downstream kinase, ERK1 (extracellular signal regulated kinase 1). Finally, the ibogaine-mediated decrease in ethanol self administration was mimicked by intra-VTA microinjection of GDNF and was reduced by intra-VTA delivery of anti-GDNF neutralizing antibodies. Together, these results suggest that GDNF in the VTA mediates the action of ibogaine on ethanol consumption. These findings highlight the importance of GDNF as a new target for drug development for alcoholism that may mimic the effect of ibogaine against alcohol consumption but avoid the negative side effects. PMID- 15659600 TI - Anti-CD11d integrin antibody treatment restores normal serotonergic projections to the dorsal, intermediate, and ventral horns of the injured spinal cord. AB - Spinal serotonergic pathways provide inhibitory and excitatory modulation of sensory, autonomic, and motor processing. After spinal cord injury (SCI), the acute inflammatory response is one process that damages descending pathways. Increases in serotonergic fiber density in spinal segments rostral and decreases caudal to the lesion have been observed previously and may contribute to neuropathic pain and motor dysfunction associated with SCI. We investigated the effect of an acute anti-inflammatory treatment on the density of serotonergic fibers rostral and caudal to a thoracic SCI lesion. This treatment, a monoclonal antibody to the CD11d subunit of the leukocyte CD11d/CD18 integrin, limits the trafficking of neutrophils and macrophages into the SCI site. In the dorsal horn, after treatment, the typically increased serotonin immunoreactivity rostral to injury was reduced, whereas that caudal to the lesion increased toward normal. Coincidently, mechanical allodynia in the dorsal trunk and hindpaws was significantly reduced. Increased serotonergic fiber density below the lesion also occurred in the intermediolateral cell column and ventral horn of treated rats, relative to controls. Improved locomotor recovery paralleled this increased serotonin. The treatment increased compact myelin in and near the lesion epicenter and increased serotonergic fiber bundles coursing around part of the lesion but had no consistent effect on the number of raphe-spinal neurons retrogradely labeled by tracer injection below the injury. In conclusion, this anti-CD11d integrin antibody treatment is neuroprotective after SCI, corresponding with improved patterns of intraspinal serotonergic innervation. The improvement in serotonergic fiber projections paralleled reduced mechanical allodynia and enhanced locomotor recovery. PMID- 15659601 TI - How stress and fluoxetine modulate serotonin 2C receptor pre-mRNA editing. AB - In two inbred strains of mice, C57BL/6 and 129Sv, the majority of forebrain neocortical pre-mRNA encoding the serotonin 2C (5-HT2C) receptor is altered by adenosine-to-inosine editing. As a result, >60% of all mRNAs encode receptors with reduced constitutive and agonist-stimulated activity. However, in the BALB/c strain, a genetically distinct inbred strain with lower forebrain serotonin levels, spontaneously elevated anxiety, and increased stress reactivity, the majority of 5-HT2C mRNA is nonedited and encodes receptors with the highest constitutive activity and the highest agonist affinity and potency. Neither acute stress (the forced swim test) nor chronic treatment with the serotonin-selective reuptake inhibitor fluoxetine elicit significant changes in 5-HT2C pre-mRNA editing in C57BL/6 mice. In contrast, exposure of BALB/c mice to acute stress and chronic treatment of nonstressed BALB/c mice with fluoxetine elicit significant, site-specific increases in 5-HT2C pre-mRNA editing that increase the pool of mRNA encoding receptors with reduced function. These changes in 5-HT2C pre-mRNA editing resemble those detected previously in the prefrontal cortex of subjects with major depression. However, when chronic fluoxetine treatment is combined with stress exposure of BALB/c mice, these changes in 5-HT2C pre-mRNA editing are no longer detected. These findings illustrate that 5-HT2C pre-mRNA editing responses to stress and chronic fluoxetine are modulated by the genetic background, as well as the behavioral state of the animal. They suggest further that the changes in 5-HT2C pre-mRNA editing found in major depression reflect a previously unrecognized molecular response to stress that can be prevented by chronic antidepressant treatment. PMID- 15659602 TI - Ensemble coding of vocal control in birdsong. AB - Zebra finch song is represented in the high-level motor control nucleus high vocal center (HVC) (Reiner et al., 2004) as a sparse sequence of spike bursts. In contrast, the vocal organ is driven continuously by smoothly varying muscle control signals. To investigate how the sparse HVC code is transformed into continuous vocal patterns, we recorded in the singing zebra finch from populations of neurons in the robust nucleus of arcopallium (RA), a premotor area intermediate between HVC and the motor neurons. We found that highly similar song elements are typically produced by different RA ensembles. Furthermore, although the song is modulated on a wide range of time scales (10-100 ms), patterns of neural activity in RA change only on a short time scale (5-10 ms). We suggest that song is driven by a dynamic circuit that operates on a single underlying clock, and that the large convergence of RA neurons to vocal control muscles results in a many-to-one mapping of RA activity to song structure. This permits rapidly changing RA ensembles to drive both fast and slow acoustic modulations, thereby transforming the sparse HVC code into a continuous vocal pattern. PMID- 15659603 TI - Transient receptor potential vanilloid subtype 1 mediates cell death of mesencephalic dopaminergic neurons in vivo and in vitro. AB - Intranigral injection of the transient receptor potential vanilloid subtype 1 (TRPV1; also known as VR1) agonist capsaicin (CAP) into the rat brain, or treatment of rat mesencephalic cultures with CAP, resulted in cell death of dopaminergic (DA) neurons, as visualized by immunocytochemistry. This in vivo and in vitro effect was ameliorated by the TRPV1 antagonist capsazepine (CZP) or iodo resiniferatoxin, suggesting the direct involvement of TRPV1 in neurotoxicity. In cultures, both CAP and anandamide (AEA), an endogenous ligand for both TRPV1 and cannabinoid type 1 (CB1) receptors, induced degeneration of DA neurons, increases in intracellular Ca2+ ([Ca2+]i), and mitochondrial damage, which were inhibited by CZP, the CB1 antagonist N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4 dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (AM251) or the intracellular Ca2+ chelator BAPTA/AM. We also found that CAP or AEA increased mitochondrial cytochrome c release as well as immunoreactivity to cleaved caspase-3 and that the caspase-3 inhibitor z-Asp-Glu-Val-Asp-fmk protected DA neurons from CAP- or AEA-induced neurotoxicity. Additional studies demonstrated that treatment of mesencephalic cultures with CB1 receptor agonist (6aR)-trans 3-(1,1 dimethylheptyl)-6a,7,10,10a-tetrahydro-1-hydroxy-6,6-dimethyl-6H-dibenzo[b,d] pyran-9-methanol (HU210) also produced degeneration of DA neurons and increases in [Ca2+]i, which were inhibited by AM251 and BAPTA/AM. The CAP-, AEA-, or HU210 induced increases in [Ca2+]i were dependent on extracellular Ca2+, with significantly different patterns of Ca2+ influx. Surprisingly, CZP and AM251 reversed HU210- or CAP-induced neurotoxicity by inhibiting Ca2+ influx, respectively, suggesting the existence of functional cross talk between TRPV1 and CB1 receptors. To our knowledge, this study is the first to demonstrate that the activation of TRPV1 and/or CB1 receptors mediates cell death of DA neurons. Our findings suggest that these two types of receptors, TRPV1 and CB1, may contribute to neurodegeneration in response to endogenous ligands such as AEA. PMID- 15659604 TI - Copper-dependent inhibition of human cytochrome c oxidase by a dimeric conformer of amyloid-beta1-42. AB - In studies of Alzheimer's disease pathogenesis there is an increasing focus on mechanisms of intracellular amyloid-beta (Abeta) generation and toxicity. Here we investigated the inhibitory potential of the 42 amino acid Abeta peptide (Abeta1 42) on activity of electron transport chain enzyme complexes in human mitochondria. We found that synthetic Abeta1-42 specifically inhibited the terminal complex cytochrome c oxidase (COX) in a dose-dependent manner that was dependent on the presence of Cu2+ and specific "aging" of the Abeta1-42 solution. Maximal COX inhibition occurred when using Abeta1-42 solutions aged for 3-6 h at 30 degrees C. The level of Abeta1-42-mediated COX inhibition increased with aging time up to approximately 6 h and then declined progressively with continued aging to 48 h. Photo-induced cross-linking of unmodified proteins followed by SDS-PAGE analysis revealed dimeric Abeta as the only Abeta species to provide significant temporal correlation with the observed COX inhibition. Analysis of brain and liver from an Alzheimer's model mouse (Tg2576) revealed abundant Abeta immunoreactivity within the brain mitochondria fraction. Our data indicate that endogenous Abeta is associated with brain mitochondria and that Abeta1-42, possibly in its dimeric conformation, is a potent inhibitor of COX, but only when in the presence of Cu2+. We conclude that Cu2+-dependent Abeta-mediated inhibition of COX may be an important contributor to the neurodegeneration process in Alzheimer's disease. PMID- 15659606 TI - Altered neuronal mitochondrial coenzyme A synthesis in neurodegeneration with brain iron accumulation caused by abnormal processing, stability, and catalytic activity of mutant pantothenate kinase 2. AB - Mutations in the pantothenate kinase 2 (PANK2) gene have been identified in patients with neurodegeneration with brain iron accumulation (NBIA; formerly Hallervorden-Spatz disease). However, the mechanisms by which these mutations cause neurodegeneration are unclear, especially given the existence of multiple pantothenate kinase genes in humans and multiple PanK2 transcripts with potentially different subcellular localizations. We demonstrate that PanK2 protein is localized to mitochondria of neurons in human brain, distinguishing it from other pantothenate kinases that do not possess mitochondrial-targeting sequences. PanK2 protein translated from the most 5' start site is sequentially cleaved at two sites by the mitochondrial processing peptidase, generating a long lived 48 kDa mature protein identical to that found in human brain extracts. The mature protein catalyzes the initial step in coenzyme A (CoA) synthesis but displays feedback inhibition in response to species of acyl CoA rather than CoA itself. Some, but not all disease-associated point mutations result in significantly reduced catalytic activity. The most common mutation, G521R, results in marked instability of the intermediate PanK2 isoform and reduced production of the mature isoform. These results suggest that NBIA is caused by altered neuronal mitochondrial lipid metabolism caused by mutations disrupting PanK2 protein levels and catalytic activity. PMID- 15659607 TI - Enriched acoustic environment after noise trauma reduces hearing loss and prevents cortical map reorganization. AB - Exposure to sound of sufficient duration and level causes permanent damage to the peripheral auditory system, which results in the reorganization of the cortical tonotopic map. The changes are such that neurons with pre-exposure tuning to frequencies in the hearing loss range now become tuned to frequencies near the near-normal lower boundary of the hearing loss range, which thus becomes over represented. However, cats exposed to a traumatizing noise and immediately thereafter placed for a few weeks in an enriched acoustic environment presented a much-restricted hearing loss compared with similarly exposed cats that were placed for the same time in a quiet environment. The enriched environment spectrally matched the expected hearing loss range and was approximately 40 dB above the level of the expected hearing loss. The hearing loss in the quiet environment-reared cats ranged from 6 to 32 kHz with the largest loss (on average, 40 dB) ranging from 24 to 32 kHz. In contrast, the hearing loss in the enriched-environment cats was restricted to 6-8 kHz at a level of, on average, 35 dB and with 16-32 kHz having normal thresholds. Despite the remaining hearing loss for the enriched-environment cats in the 6-8 kHz range, plastic tonotopic map changes in primary auditory cortex could no longer be demonstrated, suggesting that the enriched acoustic environment prevents this reorganization. This finding has implications for the treatment of hearing disorders, such as tinnitus, that have been linked to cortical tonotopic map reorganization. PMID- 15659605 TI - Controlling bursting in cortical cultures with closed-loop multi-electrode stimulation. AB - One of the major modes of activity of high-density cultures of dissociated neurons is globally synchronized bursting. Unlike in vivo, neuronal ensembles in culture maintain activity patterns dominated by global bursts for the lifetime of the culture (up to 2 years). We hypothesize that persistence of bursting is caused by a lack of input from other brain areas. To study this hypothesis, we grew small but dense monolayer cultures of cortical neurons and glia from rat embryos on multi-electrode arrays and used electrical stimulation to substitute for afferents. We quantified the burstiness of the firing of the cultures in spontaneous activity and during several stimulation protocols. Although slow stimulation through individual electrodes increased burstiness as a result of burst entrainment, rapid stimulation reduced burstiness. Distributing stimuli across several electrodes, as well as continuously fine-tuning stimulus strength with closed-loop feedback, greatly enhanced burst control. We conclude that externally applied electrical stimulation can substitute for natural inputs to cortical neuronal ensembles in transforming burst-dominated activity to dispersed spiking, more reminiscent of the awake cortex in vivo. This nonpharmacological method of controlling bursts will be a critical tool for exploring the information processing capacities of neuronal ensembles in vitro and has potential applications for the treatment of epilepsy. PMID- 15659608 TI - Dissociating barrel development and lesion-induced plasticity in the mouse somatosensory cortex. AB - In the mouse somatosensory cortex, thalamocortical axons (TCAs) corresponding to individual whiskers cluster into restricted barrel domains during the first days of life. If whiskers are lesioned before that time, the cortical space devoted to the afferents from the damaged whisker shrinks and becomes occupied by thalamocortical afferents from neighboring unlesioned whiskers. This plasticity ends by postnatal day 3 (P3) to P4 when barrels emerge. To test whether TCA development and lesion-induced plasticity are linked, we used monoamine oxidase A knock-out (MAOA-KO) mice in which normal TCA development is halted by an excess of serotonin. Normal TCA development can be restored when serotonin levels are lowered by parachlorophenylalanine (PCPA). By varying the time of PCPA administration, we found that barrel development can be reinitiated until P11, although the emergence of TCA clusters becomes gradually slower and less complete. In mice in which barrels emerge 3 d later than the normal schedule, at P6 instead of P3, we examined lesion-induced plasticity. We find a progressive decline of the lesion-induced plasticity and a closure at P3, similar to normal mice, showing that this plasticity is not influenced by an excess of serotonin levels. Thus, in MAOA-KO mice, the emergence of barrel patterning can be delayed without a concomitant delay in lesion-induced plasticity, and the cortical space devoted to one whisker representation cannot be modified by the periphery once patterning is imprinted in the subcortical relays. We conclude that the closure of the lesion-induced plasticity period in the barrelfield is probably not determined at the cortical level. PMID- 15659609 TI - Correlation between speed perception and neural activity in the middle temporal visual area. AB - We conducted electrophysiological recording and microstimulation experiments to test the hypothesis that the middle temporal visual area (MT) plays a direct role in perception of the speed of moving visual stimuli. We trained rhesus monkeys on a speed discrimination task in which monkeys chose the faster speed of two moving random dot patterns presented simultaneously in spatially segregated apertures. In electrophysiological experiments, we analyzed the activity of speed-tuned MT neurons and multiunit clusters during the discrimination task. Neural activity was correlated with the monkeys' behavioral choices on a trial-to-trial basis (choice probability), and the correlation was predicted by the speed-tuning properties of each unit. In microstimulation experiments, we activated clusters of MT neurons with homogeneous speed-tuning properties during the same speed discrimination task. In one monkey, microstimulation biased speed judgments toward the preferred speed of the stimulated neurons. Together, evidence from these two experiments suggests that MT neurons play a direct role in the perception of visual speed. Comparison of psychometric and neurometric thresholds revealed that single and multineuronal signals were, on average, considerably less sensitive than were the monkeys perceptually, suggesting that signals must be pooled across neurons to account for performance. PMID- 15659610 TI - Neuroinflammation alters the hippocampal pattern of behaviorally induced Arc expression. AB - Neuroinflammation is associated with a variety of neurological and pathological diseases, such as Alzheimer's disease (AD), and is reliably detected by the presence of activated microglia. In early AD, the highest degree of activated microglia is observed in brain regions involved in learning and memory. To investigate whether neuroinflammation alters the pattern of rapid de novo gene expression associated with learning and memory, we studied the expression of the activity-induced immediate early gene Arc in the hippocampus of rats with experimental neuroinflammation. Rats were chronically infused with lipopolysaccharide (LPS) (0.25 mug/h) into the fourth ventricle for 28 d. On day 29, the rats explored twice a novel environment for 5 min, separated by 45 or 90 min. In the dentate gyrus and CA3 regions of LPS-infused rats, Arc and OX-6 (specific for major histocompatibility complex class II antigens) immunolabeling and Arc fluorescence in situ hybridization revealed both activated microglia (OX 6 immunoreactivity) and elevated exploration-induced Arc expression compared with control-infused rats. In contrast, in the CA1 of LPS-infused rats, where there was no OX-6 immunostaining, exploration-induced Arc mRNA and protein remained similar in both LPS- and control-infused rats. LPS-induced neuroinflammation did not affect basal levels of Arc expression. Behaviorally induced Arc expression was altered only within the regions showing activated microglia (OX-6 immunoreactivity), suggesting that neuroinflammation may alter the coupling of neural activity with macromolecular synthesis implicated in learning and plasticity. This activity-related alteration in Arc expression induced by neuroinflammation may contribute to the cognitive deficits found in diseases associated with inflammation, such as AD. PMID- 15659611 TI - A uniquely selective inhibitor of the mammalian fetal neuromuscular nicotinic acetylcholine receptor. AB - We have purified and characterized a novel conotoxin from the venom of Conus obscurus, which has the unique property of selectively and potently inhibiting the fetal form of the mammalian neuromuscular nicotinic acetylcholine receptor (nAChR) (alpha1beta1gammadelta-subunits). Although this conotoxin, alphaA conotoxin OIVB (alphaA-OIVB), is a high-affinity antagonist (IC50 of 56 nm) of the fetal muscle nAChR, it has >1800-fold lower affinity for the adult muscle nAChR (alpha1beta1epsilondelta-subunits) and virtually no inhibitory activity at a high concentration on various neuronal nAChRs (IC50 > 100 microm in all cases). The peptide (amino acid sequence, CCGVONAACPOCVCNKTCG), with three disulfide bonds, has been chemically synthesized in a biologically active form. Although the neuromuscular nAChRs are perhaps the most extensively characterized of the receptors/ion channels of the nervous system, the precise physiological roles of the fetal form of the muscle nAChR are essentially unknown.alphaA-OIVB is a potentially important tool for delineating the functional roles ofalpha1beta1gammadelta receptors in normal development, as well as in various adult tissues and in pathological states. In addition to its potential as a research tool, alphaA-OIVB may have some direct biomedical applications. PMID- 15659612 TI - Expression of stathmin, a developmentally controlled cytoskeleton-regulating molecule, in demyelinating disorders. AB - Understanding the biological relevance of reexpression of developmental molecules in pathological conditions is crucial for the development of new therapies. In this study, we report the increased expression of stathmin, a developmentally regulated tubulin-binding protein, in the brains of patients with multiple sclerosis (MS). In physiological conditions, stathmin immunoreactivity was observed in polysialic acid-neural cell adhesion molecule-positive migratory progenitors in the subventricular zone, and its expression progressively decreased as the cells matured into oligodendrocytes (OLs). In MS patients, however, stathmin levels were elevated in 2',3'-cyclic nucleotide 3' phosphodiesterase-positive OLs, in 10 of 10 bioptic samples analyzed. Increased levels of stathmin were confirmed by Western blot analysis of normal-appearing white matter samples from MS brains. In addition, using mass spectrometry, stathmin was identified as the main component of a specific myelin protein fraction consistently increased in MS preparations compared with controls. To test the biological relevance of increased stathmin levels, primary OL progenitors were transfected using a myc-tagged stathmin cDNA and were allowed to differentiate. Consistent with a distinct role played by this molecule in cells of the OL lineage at different developmental stages, transient transfection in progenitors favored the bipolar migratory phenotype but did not affect survival. However, sustained stathmin levels in differentiating OLs, because of overexpression, resulted in enhanced apoptotic susceptibility. We conclude that stathmin expression in demyelinating disorders could have a dual role. On one hand, by favoring the migratory phenotype of progenitors, it may promote myelin repair. On the other hand, stathmin in mature OLs may indicate cell stress and possibly affect survival. PMID- 15659613 TI - Mutations in PRPF31 inhibit pre-mRNA splicing of rhodopsin gene and cause apoptosis of retinal cells. AB - Mutations in human PRPF31 gene have been identified in patients with autosomal dominant retinitis pigmentosa (adRP). To begin to understand mechanisms by which defects in this general splicing factor cause retinal degeneration, we examined the relationship between PRPF31 and pre-mRNA splicing of photoreceptor-specific genes. We used a specific anti-PRPF31 antibody to immunoprecipitate splicing complexes from retinal cells and identified the transcript of rhodopsin gene (RHO) among RNA species associated with PRPF31-containing complexes. Mutant PRPF31 proteins significantly inhibited pre-mRNA splicing of intron 3 in RHO gene. In primary retinal cell cultures, expression of the mutant PRPF31 proteins reduced rhodopsin expression and caused apoptosis of rhodopsin-positive retinal cells. This primary retinal culture assay provides an in vitro model to study photoreceptor cell death caused by PRPF31 mutations. Our results demonstrate that mutations in PRPF31 gene affect RHO pre-mRNA splicing and reveal a link between PRPF31 and RHO, two major adRP genes. PMID- 15659614 TI - Neurotrophin-3 suppresses thermal hyperalgesia associated with neuropathic pain and attenuates transient receptor potential vanilloid receptor-1 expression in adult sensory neurons. AB - Neurotrophin-3 (NT-3) negatively modulates nerve growth factor (NGF) receptor expression and associated nociceptive phenotype in intact neurons, suggesting a beneficial role in treating aspects of neuropathic pain mediated by NGF. We report that NT-3 is effective at suppressing thermal hyperalgesia associated with chronic constriction injury (CCI); however, NT-3 does not alter the mechanical hypersensitivity that also develops with CCI. Thermal hyperalgesia is critically linked to expression and activation of the capsaicin receptor, transient receptor potential vanilloid receptor-1 (TRPV1). Thus, its modulation by NT-3 after CCI was examined. CCI results in elevated TRPV1 expression at both the mRNA and protein levels in predominantly small-to-medium neurons, with the percentage of neurons expressing TRPV1 remaining unchanged at approximately 56%. Attenuation of thermal hyperalgesia mediated by NT-3 correlates with decreased TRPV1 expression such that only approximately 26% of neurons ipsilateral to CCI expressed detectable TRPV1 mRNA. NT-3 effected a decrease in expression of the activated component of the signaling pathway linked to regulation of TRPV1 expression, phospho-p38 MAPK (Ji et al., 2002), in neurons ipsilateral to CCI. Exogenous NT-3 could both prevent the onset of thermal hyperalgesia and reverse established thermal hyperalgesia and elevated TRPV1 expression 1 week after CCI. Continuous infusion is required for suppression of both thermal hyperalgesia and TRPV1 expression, because removal of NT-3 resulted in a prompt reestablishment of the hyperalgesic state and corresponding CCI-associated TRPV1 phenotype. In conclusion, although NGF drives inflammation-associated thermal hyperalgesia via its regulation of TRPV1 expression, NT-3 is now identified as a potent negative modulator of this state. PMID- 15659615 TI - Interaction of Helicobacter pylori with sialylated carbohydrates: the dependence on different parts of the binding trisaccharide Neu5Ac{alpha}3Gal{beta}4GlcNAc. AB - We have recently shown that binding of Helicobacter pylori to sialylated carbohydrates is dependent on the presence of the carboxyl group and the glycerol chain of Neu5Ac. In this work we investigated the importance of GlcNAc in the binding trisaccharide Neu5Acalpha3Galbeta4GlcNAc and the role of the N-acetamido groups of both Neu5Ac and GlcNAc. An important part of the project was epitope dissection, that is chemical derivatizations of the active carbohydrate followed by binding studies. In addition we used a panel of various unmodified carbohydrate structures in the form of free oligosaccharides or glycolipids. These were tested for binding by hemagglutination inhibition assay, TLC overlay tests, and a new quantitative approach using radiolabeled neoglycoproteins. The studies showed that the N-acetamido group of Neu5Ac is important for binding by H. pylori, whereas the same group of GlcNAc is not. In addition, Fuc attached to GlcNAc, as tested with sialyl-Lewis x, did not affect the binding. Free Neu5Ac was inactive as inhibitor, and Neu5Acalpha3Gal turned out to be active. The binding preference for neolacto structures was confirmed, although one strain also was inhibited by lacto chains. The combined results revealed that an intact Neu5Ac is crucial for the interactions with H. pylori. Parts of Gal also seem to be necessary, whereas the role of the GlcNAc is secondary. GlcNAc does influence binding, however, primarily serving as a guiding carrier for the binding epitope rather than being a part of the binding structure. PMID- 15659616 TI - Testis-specific sulfoglycolipid, seminolipid, is essential for germ cell function in spermatogenesis. AB - More than 90% of the glycolipid in mammalian testis consists of a unique sulfated glyceroglycolipid, seminolipid. The sulfation of the molecule is catalyzed by a Golgi membrane-associated sulfotransferase, cerebroside sulfotransferase (CST). Disruption of the Cst gene in mice results in male infertility due to the arrest of spermatogenesis prior to the metaphase of the first meiosis. However, the issue of which side of the cell function-germ cells or Sertoli cells-is deteriorated in this mutant mouse remains unknown. Our findings show that the defect is in the germ cell side, as evidenced by a transplantation analysis, in which wild-type spermatogonia expressing the green fluorescent protein were injected into the seminiferous tubules of CST-null testis. The transplanted GFP positive cells generated colonies and spermatogenesis proceeded over meiosis in the mutant testis. The findings also clearly show that the seminolipid is expressed on the plasma membranes of spermatogonia, spermatocytes, spermatids, and spermatozoa, as evidenced by the immunostaining of wild-type testes using an anti-sulfogalactolipid antibody, Sulph-1 in comparison with CST-null testes as a negative control, and that seminolipid appears as early as day 8 of age, when Type B spermatogonia emerge. PMID- 15659617 TI - SCL/Tal1 and lymphoid versus myeloid lineage assignment. PMID- 15659619 TI - Reproducible measurements of AML blast p-glycoprotein function in 2 center analyses. PMID- 15659618 TI - Potential curability of newly diagnosed acute promyelocytic leukemia without use of chemotherapy: the example of liposomal all-trans retinoic acid. PMID- 15659620 TI - Is immune thrombocytopenic purpura less common among black Americans? PMID- 15659621 TI - Dissection of Arabidopsis ADP-RIBOSYLATION FACTOR 1 function in epidermal cell polarity. AB - Vesicle trafficking is essential for the generation of asymmetries, which are central to multicellular development. Core components of the vesicle transport machinery, such as ADP-ribosylation factor (ARF) GTPases, have been studied primarily at the single-cell level. Here, we analyze developmental functions of the ARF1 subclass of the Arabidopsis thaliana multigene ARF family. Six virtually identical ARF1 genes are ubiquitously expressed, and single loss-of-function mutants in these genes reveal no obvious developmental phenotypes. Fluorescence colocalization studies reveal that ARF1 is localized to the Golgi apparatus and endocytic organelles in both onion (Allium cepa) and Arabidopsis cells. Apical basal polarity of epidermal cells, reflected by the position of root hair outgrowth, is affected when ARF1 mutants are expressed at early stages of cell differentiation but after they exit mitosis. Genetic interactions during root hair tip growth and localization suggest that the ROP2 protein is a target of ARF1 action, but its localization is slowly affected upon ARF1 manipulation when compared with that of Golgi and endocytic markers. Localization of a second potential target of ARF1 action, PIN2, is also affected with slow kinetics. Although extreme redundancy precludes conventional genetic dissection of ARF1 functions, our approach separates different ARF1 downstream networks involved in local and specific aspects of cell polarity. PMID- 15659622 TI - The rb7 matrix attachment region increases the likelihood and magnitude of transgene expression in tobacco cells: a flow cytometric study. AB - Many studies in both plant and animal systems have shown that matrix attachment regions (MARs) can increase expression of transgenes in whole organisms or cells in culture. Because histochemical assays often indicate variegated transgene expression, a question arises: Do MARs increase transgene expression by increasing the percentage of cells expressing the transgene (likelihood), by increasing the level of expression in expressing cells (magnitude), or both? To address this question, we used flow cytometry to measure green fluorescent protein (GFP) expression in individual tobacco (Nicotiana tabacum) cells from lines transformed by Agrobacterium tumefaciens. We conclude that MAR-mediated overall increases in transgene expression involve both likelihood and magnitude. On average, cell lines transformed with the Rb7 MAR-containing vector expressed GFP at levels 2.0- to 3.7-fold higher than controls. MAR lines had fewer nonexpressing cells than control lines (10% versus 45%), and the magnitude of GFP expression in expressing cells was greater in MAR lines by 1.9- to 2.9-fold. We also show that flow cytometry measurements on cells from isogenic lines are consistent with those from populations of independently transformed cell lines. By obviating the need to establish isogenic lines, this use of flow cytometry could greatly simplify the evaluation of MARs or other sequence elements that affect transgene expression. PMID- 15659623 TI - Characterization of an Arabidopsis enzyme family that conjugates amino acids to indole-3-acetic acid. AB - Substantial evidence indicates that amino acid conjugates of indole-3-acetic acid (IAA) function in auxin homeostasis, yet the plant enzymes involved in their biosynthesis have not been identified. We tested whether several Arabidopsis thaliana enzymes that are related to the auxin-induced soybean (Glycine max) GH3 gene product synthesize IAA-amino acid conjugates. In vitro reactions with six recombinant GH3 enzymes produced IAA conjugates with several amino acids, based on thin layer chromatography. The identity of the Ala, Asp, Phe, and Trp conjugates was verified by gas chromatography-mass spectrometry. Insertional mutations in GH3.1, GH3.2, GH3.5, and GH3.17 resulted in modestly increased sensitivity to IAA in seedling root. Overexpression of GH3.6 in the activation tagged mutant dfl1-D did not significantly alter IAA level but resulted in 3.2- and 4.5-fold more IAA-Asp than in wild-type seedlings and mature leaves, respectively. In addition to IAA, dfl1-D was less sensitive to indole-3-butyric acid and naphthaleneacetic acid, consistent with the fact that GH3.6 was active on each of these auxins. By contrast, GH3.6 and the other five enzymes tested were inactive on halogenated auxins, and dfl1-D was not resistant to these. This evidence establishes that several GH3 genes encode IAA-amido synthetases, which help to maintain auxin homeostasis by conjugating excess IAA to amino acids. PMID- 15659624 TI - The role of KNOX genes in the evolution of morphological novelty in Streptocarpus. AB - The genus Streptocarpus comprises species with diverse body plans. Caulescent species produce leaves from a conventional shoot apical meristem (SAM), whereas acaulescent species lack a conventional SAM and produce only a single leaf (the unifoliate form) or clusters of leaves from the base of more mature leaves (the rosulate form). These distinct morphologies reflect fundamental differences in the role of the SAM and the process of leaf specification. A subfamily of KNOTTED like homeobox (KNOX) genes are known to be important in regulating meristem function and leaf development in model species with conventional morphologies. To test the involvement of KNOX genes in Streptocarpus evolution, two parologous KNOX genes (SSTM1 and SSTM2) were isolated from species with different growth forms. Their phylogenetic analysis suggested a gene duplication before the subgeneric split of Streptocarpus and resolved species relationships, supporting multiple evolutionary origins of the rosulate and unifoliate morphologies. In S. saxorum, a caulescent species with a conventional SAM, KNOX proteins were expressed in the SAM and transiently downregulated in incipient leaf primordia. The ability of acaulescent species to initiate leaves from existing leaves was found to correlate with SSTM1 expression and KNOX protein accumulation in leaves and to reflect genetic differences at two loci. Neither locus corresponded to SSTM1, suggesting that cis-acting differences in SSTM1 regulation were not responsible for evolution of the rosulate and unifoliate forms. However, the involvement of KNOX proteins in leaf formation in rosulate species suggests that they have played an indirect role in the development of morphological diversity in Streptocarpus. PMID- 15659625 TI - Characterization of the Arabidopsis clb6 mutant illustrates the importance of posttranscriptional regulation of the methyl-D-erythritol 4-phosphate pathway. AB - The biosynthesis of isopentenyl diphosphate and dimethylallyl diphosphate, the two building blocks for isoprenoid biosynthesis, occurs by two independent pathways in plants. The mevalonic pathway operates in the cytoplasm, and the methyl-d-erythritol 4-phosphate (MEP) pathway operates in plastids. Plastidic isoprenoids play essential roles in plant growth and development. Plants must regulate the biosynthesis of isoprenoids to fulfill metabolic requirements in specific tissues and developmental conditions. The regulatory events that modulate the plant MEP pathway are not well understood. In this article, we demonstrate that the CHLOROPLAST BIOGENESIS6 (CLB6) gene, previously shown to be required for chloroplast development, encodes 1-hydroxy-2-methyl-butenyl 4 diphosphate reductase, the last-acting enzyme of the MEP pathway. Comparative analysis of the expression levels of all MEP pathway gene transcripts and proteins in the clb6-1 mutant background revealed that posttranscriptional control modulates the levels of different proteins in this central pathway. Posttranscriptional regulation was also found during seedling development and during fosmidomycin inhibition of the pathway. Our results show that the first enzyme of the pathway, 1-deoxy-d-xylulose 5-phosphate synthase, is feedback regulated in response to the interruption of the flow of metabolites through the MEP pathway. PMID- 15659626 TI - Arabidopsis VILLIN1 generates actin filament cables that are resistant to depolymerization. AB - Dynamic cytoplasmic streaming, organelle positioning, and nuclear migration use molecular tracks generated from actin filaments arrayed into higher-order structures like actin cables and bundles. How these arrays are formed and stabilized against cellular depolymerizing forces remains an open question. Villin and fimbrin are the best characterized actin-filament bundling or cross linking proteins in plants and each is encoded by a multigene family of five members in Arabidopsis thaliana. The related villins and gelsolins are conserved proteins that are constructed from a core of six homologous gelsolin domains. Gelsolin is a calcium-regulated actin filament severing, nucleating and barbed end capping factor. Villin has a seventh domain at its C terminus, the villin headpiece, which can bind to an actin filament, conferring the ability to crosslink or bundle actin filaments. Many, but not all, villins retain the ability to sever, nucleate, and cap filaments. Here we have identified a putative calcium-insensitive villin isoform through comparison of sequence alignments between human gelsolin and plant villins with x-ray crystallography data for vertebrate gelsolin. VILLIN1 (VLN1) has the least well-conserved type 1 and type 2 calcium binding sites among the Arabidopsis VILLIN isoforms. Recombinant VLN1 binds to actin filaments with high affinity (K(d) approximately 1 microM) and generates bundled filament networks; both properties are independent of the free Ca(2+) concentration. Unlike human plasma gelsolin, VLN1 does not nucleate the assembly of filaments from monomer, does not block the polymerization of profilin actin onto barbed ends, and does not stimulate depolymerization or sever preexisting filaments. In kinetic assays with ADF/cofilin, villin appears to bind first to growing filaments and protects filaments against ADF-mediated depolymerization. We propose that VLN1 is a major regulator of the formation and stability of actin filament bundles in plant cells and that it functions to maintain the cable network even in the presence of stimuli that result in depolymerization of other actin arrays. PMID- 15659627 TI - The Arabidopsis WAVY GROWTH 2 protein modulates root bending in response to environmental stimuli. AB - To understand how the direction of root growth changes in response to obstacles, light, and gravity, we characterized an Arabidopsis thaliana mutant, wavy growth 2 (wav2), whose roots show a short-pitch pattern of wavy growth on inclined agar medium. The roots of the wav2 mutant bent with larger curvature than those of the wild-type seedlings in wavy growth and in gravitropic and phototropic responses. The cell file rotations of the root epidermis of wav2-1 in the wavy growth pattern were enhanced in both right-handed and left-handed rotations. WAV2 encodes a protein belonging to the BUD EMERGENCE 46 family with a transmembrane domain at the N terminus and an alpha/beta-hydrolase domain at the C terminus. Expression analyses showed that mRNA of WAV2 was expressed strongly in adult plant roots and seedlings, especially in the root tip, the cell elongation zone, and the stele. Our results suggest that WAV2 is not involved in sensing environmental stimuli but that it negatively regulates stimulus-induced root bending through inhibition of root tip rotation. PMID- 15659628 TI - Phosphoserines on maize CENTROMERIC HISTONE H3 and histone H3 demarcate the centromere and pericentromere during chromosome segregation. AB - We have identified and characterized a 17- to 18-kD Ser50-phosphorylated form of maize (Zea mays) CENTROMERIC HISTONE H3 (phCENH3-Ser50). Immunostaining in both mitosis and meiosis indicates that CENH3-Ser50 phosphorylation begins in prophase/diplotene, increases to a maximum at prometaphase-metaphase, and drops during anaphase. Dephosphorylation is precipitous (approximately sixfold) at the metaphase-anaphase transition, suggesting a role in the spindle checkpoint. Although phCENH3-Ser50 lies within a region that lacks homology to any other known histone, its closest counterpart is the phospho-Ser28 residue of histone H3 (phH3-Ser28). CENH3-Ser50 and H3-Ser28 are phosphorylated with nearly identical kinetics, but the former is restricted to centromeres and the latter to pericentromeres. Opposing centromeres separate in prometaphase, whereas the phH3 Ser28-marked pericentromeres remain attached and coalesce into a well-defined tether that binds the centromeres together. We propose that a centromere initiated wave of histone phosphorylation is an early step in defining the two major structural domains required for chromosome segregation: centromere (alignment, motility) and pericentromere (cohesion). PMID- 15659630 TI - The Arabidopsis HOMOLOGY-DEPENDENT GENE SILENCING1 gene codes for an S-adenosyl-L homocysteine hydrolase required for DNA methylation-dependent gene silencing. AB - Genes introduced into higher plant genomes can become silent (gene silencing) and/or cause silencing of homologous genes at unlinked sites (homology-dependent gene silencing or HDG silencing). Mutations of the HOMOLOGY-DEPENDENT GENE SILENCING1 (HOG1) locus relieve transcriptional gene silencing and methylation dependent HDG silencing and result in genome-wide demethylation. The hog1 mutant plants also grow slowly and have low fertility and reduced seed germination. Three independent mutants of HOG1 were each found to have point mutations at the 3' end of a gene coding for S-adenosyl-l-homocysteine (SAH) hydrolase, and hog1-1 plants show reduced SAH hydrolase activity. A transposon (hog1-4) and a T-DNA tag (hog1-5) in the HOG1 gene each behaved as zygotic embryo lethal mutants and could not be made homozygous. The results suggest that the homozygous hog1 point mutants are leaky and result in genome demethylation and poor growth and that homozygous insertion mutations result in zygotic lethality. Complementation of the hog1-1 point mutation with a T-DNA containing the gene coding for SAH hydrolase restored gene silencing, HDG silencing, DNA methylation, fast growth, and normal seed viability. The same T-DNA also complemented the zygotic embryo lethal phenotype of the hog1-4 tagged mutant. A model relating the HOG1 gene, DNA methylation, and methylation-dependent HDG silencing is presented. PMID- 15659631 TI - Functional genomic analysis of the AUXIN RESPONSE FACTOR gene family members in Arabidopsis thaliana: unique and overlapping functions of ARF7 and ARF19. AB - The AUXIN RESPONSE FACTOR (ARF) gene family products, together with the AUXIN/INDOLE-3-ACETIC ACID proteins, regulate auxin-mediated transcriptional activation/repression. The biological function(s) of most ARFs is poorly understood. Here, we report the identification and characterization of T-DNA insertion lines for 18 of the 23 ARF gene family members in Arabidopsis thaliana. Most of the lines fail to show an obvious growth phenotype except of the previously identified arf2/hss, arf3/ett, arf5/mp, and arf7/nph4 mutants, suggesting that there are functional redundancies among the ARF proteins. Subsequently, we generated double mutants. arf7 arf19 has a strong auxin-related phenotype not observed in the arf7 and arf19 single mutants, including severely impaired lateral root formation and abnormal gravitropism in both hypocotyl and root. Global gene expression analysis revealed that auxin-induced gene expression is severely impaired in the arf7 single and arf7 arf19 double mutants. For example, the expression of several genes, such as those encoding members of LATERAL ORGAN BOUNDARIES domain proteins and AUXIN-REGULATED GENE INVOLVED IN ORGAN SIZE, are disrupted in the double mutant. The data suggest that the ARF7 and ARF19 proteins play essential roles in auxin-mediated plant development by regulating both unique and partially overlapping sets of target genes. These observations provide molecular insight into the unique and overlapping functions of ARF gene family members in Arabidopsis. PMID- 15659632 TI - Large intraspecific haplotype variability at the Rph7 locus results from rapid and recent divergence in the barley genome. AB - To study genome evolution and diversity in barley (Hordeum vulgare), we have sequenced and compared more than 300 kb of sequence spanning the Rph7 leaf rust disease resistance gene in two barley cultivars. Colinearity was restricted to five genic and two intergenic regions representing <35% of the two sequences. In each interval separating the seven conserved regions, the number and type of repetitive elements were completely different between the two homologous sequences, and a single gene was absent in one cultivar. In both cultivars, the nonconserved regions consisted of approximately 53% repetitive sequences mainly represented by long-terminal repeat retrotransposons that have inserted <1 million years ago. PCR-based analysis of intergenic regions at the Rph7 locus and at three other independent loci in 41 H. vulgare lines indicated large haplotype variability in the cultivated barley gene pool. Together, our data indicate rapid and recent divergence at homologous loci in the genome of H. vulgare, possibly providing the molecular mechanism for the generation of high diversity in the barley gene pool. Finally, comparative analysis of the gene composition in barley, wheat (Triticum aestivum), rice (Oryza sativa), and sorghum (Sorghum bicolor) suggested massive gene movements at the Rph7 locus in the Triticeae lineage. PMID- 15659633 TI - Plus and minus sexual agglutinins from Chlamydomonas reinhardtii. AB - Gametes of the unicellular green alga Chlamydomonas reinhardtii undergo sexual adhesion via enormous chimeric Hyp-rich glycoproteins (HRGPs), the plus and minus sexual agglutinins, that are displayed on their flagellar membrane surfaces. We have previously purified the agglutinins and analyzed their structural organization using electron microscopy. We report here the cloning and sequencing of the Sag1 and Sad1 genes that encode the two agglutinins and relate their derived amino acid sequences and predicted secondary structure to the morphology of the purified proteins. Both agglutinin proteins are organized into three distinct domains: a head, a shaft in a polyproline II configuration, and an N terminal domain. The plus and minus heads are related in overall organization but poorly conserved in sequence except for two regions of predicted hydrophobic alpha-helix. The shafts contain numerous repeats of the PPSPX motif previously identified in Gp1, a cell wall HRGP. We propose that the head domains engage in autolectin associations with the distal termini of their own shafts and suggest ways that adhesion may involve head-head interactions, exolectin interactions between the heads and shafts of opposite type, and antiparallel shaft-shaft interactions mediated by carbohydrates displayed in polyproline II configurations. PMID- 15659634 TI - DISTORTED3/SCAR2 is a putative arabidopsis WAVE complex subunit that activates the Arp2/3 complex and is required for epidermal morphogenesis. AB - In a plant cell, a subset of actin filaments function as a scaffold that positions the endomembrane system and acts as a substrate on which organelle motility occurs. Other actin filament arrays appear to be more dynamic and reorganize in response to growth signals and external cues. The distorted group of trichome morphology mutants provides powerful genetic tools to study the control of actin filament nucleation in the context of morphogenesis. In this article, we report that DISTORTED3 (DIS3) encodes a plant-specific SCAR/WAVE homolog. Null alleles of DIS3, like those of other Arabidopsis thaliana WAVE and Actin-Related Protein (ARP) 2/3 subunit genes, cause trichome distortion, defects in cell-cell adhesion, and reduced hypocotyl growth in etiolated seedlings. DIS3 efficiently activates the actin filament nucleation and branching activity of vertebrate Arp2/3 and functions within a WAVE-ARP2/3 pathway in vivo. DIS3 may assemble into a WAVE complex via a physical interaction with a highly diverged Arabidopsis Abi-1-like bridging protein. These results demonstrate the utility of the Arabidopsis trichome system to understand how the WAVE and ARP2/3 complexes translate signaling inputs into a coordinated morphogenetic response. PMID- 15659635 TI - Mx-rMx, a family of interacting transposons in the growing hAT superfamily of maize. AB - More than half a century after the discovery of transposable elements, the number of genetically defined autonomous elements that have been isolated and characterized molecularly in any one species remains surprisingly small. Because of its rich genetic history, maize (Zea mays) is, by far, the plant with the largest number of such elements. Yet, even in maize, a maximum of only two autonomous elements have been characterized in any transposon superfamily. This article describes the isolation and molecular and genetic characterization of Mx (for mobile element induced by x-rays), a third autonomous member of the hAT transposon superfamily in maize. Mx is 3731 bp long, ends in 13-bp terminal inverted repeats (TIRs), and causes an 8-bp duplication of the target site. Mx and rMx (for responder to Mx), its 571-bp nonautonomous partner, define a classical family of interacting transposable elements. Surprisingly, the TIRs of Mx and rMx are only 73% identical, and the subterminal sequences are even less so, suggesting that Mx and rMx may represent diverging transposable elements still capable of mobilization by the same transposase. Sequences that are closer to the ends of either Mx or rMx are present in the maize genome. Mx is predicted to encode a 674-amino acid protein that is homologous to the Ac transposase. Although Mx and Ac are closely related, they do not interact. Other data suggest that maize may possess at least five families of hAT transposons that do not interact with each other. The possible origin of noninteracting transposon families within the same superfamily is discussed. PMID- 15659629 TI - Functional isolation of novel nuclear proteins showing a variety of subnuclear localizations. AB - Nuclear proteins play key roles in the fundamental regulation of genome instability, the phases of organ development, and physiological responsiveness through gene expression. Although nuclear proteins have been shown to account for approximately one-fourth of total proteins in yeast, no efficient method to identify novel nuclear proteins has been applied to plants. In this study, a trial to isolate nuclear proteins in rice was attempted, and several novel nuclear proteins showing a variety of subnuclear localizations were identified. The nuclear transportation trap (NTT) system, which is a modified two-hybrid system, isolated many nuclear proteins from rice (Oryza sativa) NTT cDNA libraries. Nuclear localization of the isolated proteins was confirmed by transient introduction of green fluorescent protein fusion constructs for a subset of protein genes into onion (Allium cepa) cells. The majority of these proteins, including novel proteins and proteins initially categorized as cytoplasmic proteins, were revealed to be localized in the nucleus. Detailed characterization of unknown proteins revealed various subnuclear localizations, indicating their possible association with chromatin and the nuclear matrix with a foci or speckle-like distribution. Some also showed dual distribution in the nucleus and cytoplasm. In the novel protein fraction, a protein was further identified for its chromatin-associated localization in a specific organ of rice by immunostaining. Thus, a variety of novel nuclear architectural proteins with chromatin or matrix associating abilities, which are important in nuclear organization by influencing certain organ developments or cell responsiveness, can be isolated using the NTT method. Because nuclear proteins other than transcription regulators have rarely been characterized in plants, such as matrix proteins and development-specific chromatin proteins, their identification and subsequent characterization could provide important information for genome-wide regulatory mechanisms controlled by nuclear organization. PMID- 15659636 TI - The Dof transcription factor OBP3 modulates phytochrome and cryptochrome signaling in Arabidopsis. AB - Plants perceive subtle changes in light quality and quantity through a set of photoreceptors, including phytochromes and cryptochromes. Upon perception, these photoreceptors initiate signal transduction pathways leading to photomorphogenic changes in development. Using activation-tagging mutagenesis to identify novel light-signaling components, we have isolated a gain-of-function mutant, sob1-D (suppressor of phytochrome B-4 [phyB-4] dominant), which suppresses the long hypocotyl phenotype of the phyB missense allele, phyB-4. The sob1-D mutant phenotype is caused by the overexpression of a Dof (DNA binding with one finger) transcription factor, OBF4 Binding Protein 3 (OBP3). A translational fusion between OBP3 and green fluorescent protein is nuclear localized in onion (Allium cepa) cells. Tissue-specific accumulation of an OBP3:OBP3-beta-glucuronidase translational fusion is regulated by light in Arabidopsis thaliana. Hypocotyls of transgenic lines with reduced OBP3 expression are less responsive to red light. This aberrant phenotype in red light requires functional phyB, suggesting that OBP3 is a positive regulator of phyB-mediated inhibition of hypocotyl elongation. Furthermore, these partial-loss-of-function lines have larger cotyledons. This light-dependent cotyledon phenotype is most dramatic in blue light and requires functional cryptochrome 1 (cry1), indicating that OBP3 is a negative regulator of cry1-mediated cotyledon expansion. These results suggest a model where OBP3 is a component in both phyB and cry1 signaling pathways, acting as a positive and negative regulator, respectively. An alternate, though not mutually exclusive, model places OBP3 as a general inhibitor of tissue expansion with phyB and cry1, differentially modulating OBP3's role in this response. PMID- 15659638 TI - Genetic analysis reveals domain interactions of Arabidopsis Hsp100/ClpB and cooperation with the small heat shock protein chaperone system. AB - We have defined amino acids important for function of the Arabidopsis thaliana Hsp100/ClpB chaperone (AtHsp101) in acquired thermotolerance by isolating recessive, loss-of-function mutations and a novel semidominant, gain-of-function allele [hot1-4 (A499T)]. The hot1-4 allele is unusual in that it not only fails to develop thermotolerance to 45 degrees C after acclimation at 38 degrees C, but also is sensitive to 38 degrees C, which is a permissive temperature for wild type and loss-of-function mutants. hot1-4 lies between nucleotide binding domain 1 (NBD1) and NBD2 in a coiled-coil domain that is characteristic of the Hsp100/ClpB proteins. We then isolated two classes of intragenic suppressor mutations of hot1-4: loss-of-function mutations (Class 1) that eliminated the 38 degrees C sensitivity, but did not restore thermotolerance function to hot1-4, and Class 2 suppressors that restored acquired thermotolerance function to hot1 4. Location of the hot1-4 Class 2 suppressors supports a functional link between the coiled-coil domain and both NBD1 and the axial channel of the Hsp100/ClpB hexamer. In addition, the strongest Class 2 suppressors restored solubility of aggregated small heat shock proteins (sHsps) after heat stress, revealing genetic interaction of the Hsp100/ClpB and sHsp chaperone systems. These results also demonstrate that quantitative phenotypes can be used for in vivo genetic dissection of protein mechanism in Arabidopsis. PMID- 15659637 TI - VANGUARD1 encodes a pectin methylesterase that enhances pollen tube growth in the Arabidopsis style and transmitting tract. AB - In flowering plants, penetration of the pollen tube through stigma, style, and transmitting tract is essential for delivery of sperm nuclei to the egg cells embedded deeply within female tissues. Despite its importance in plant reproduction, little is known about the underlying molecular mechanisms that regulate the navigation of the pollen tube through the stigma, style, and transmitting tract. Here, we report the identification and characterization of an Arabidopsis thaliana gene, VANGUARD1 (VGD1) that encodes a pectin methylesterase (PME)-homologous protein of 595 amino acids and is required for enhancing the growth of pollen tubes in the style and transmitting tract tissues. VGD1 was expressed specifically in pollen grain and the pollen tube. The VGD1 protein was distributed throughout the pollen grain and pollen tube, including the plasma membrane and cell wall. Functional interruption of VGD1 reduced PME activity in the pollen to 82% of the wild type and greatly retarded the growth of the pollen tube in the style and transmitting tract, resulting in a significant reduction of male fertility. In addition, the vgd1 pollen tubes were unstable and burst more frequently when germinated and grown on in vitro culture medium, compared with wild-type pollen tubes. Our study suggests that the VGD1 product is required for growth of the pollen tube, possibly via modifying the cell wall and enhancing the interaction of the pollen tube with the female style and transmitting tract tissues. PMID- 15659639 TI - The branching gene RAMOSUS1 mediates interactions among two novel signals and auxin in pea. AB - In Pisum sativum, the RAMOSUS genes RMS1, RMS2, and RMS5 regulate shoot branching via physiologically defined mobile signals. RMS1 is most likely a carotenoid cleavage enzyme and acts with RMS5 to control levels of an as yet unidentified mobile branching inhibitor required for auxin inhibition of branching. Our work provides molecular, genetic, and physiological evidence that RMS1 plays a central role in a shoot-to-root-to-shoot feedback system that regulates shoot branching in pea. Indole-3-acetic acid (IAA) positively regulates RMS1 transcript level, a potentially important mechanism for regulation of shoot branching by IAA. In addition, RMS1 transcript levels are dramatically elevated in rms3, rms4, and rms5 plants, which do not contain elevated IAA levels. This degree of upregulation of RMS1 expression cannot be achieved in wild-type plants by exogenous IAA application. Grafting studies indicate that an IAA-independent mobile feedback signal contributes to the elevated RMS1 transcript levels in rms4 plants. Therefore, the long-distance signaling network controlling branching in pea involves IAA, the RMS1 inhibitor, and an IAA-independent feedback signal. Consistent with physiological studies that predict an interaction between RMS2 and RMS1, rms2 mutations appear to disrupt this IAA-independent regulation of RMS1 expression. PMID- 15659641 TI - The functionally conserved nucleoporins Nup124p from fission yeast and the human Nup153 mediate nuclear import and activity of the Tf1 retrotransposon and HIV-1 Vpr. AB - We report that the fission yeast nucleoporin Nup124p is required for the nuclear import of both, retrotransposon Tf1-Gag as well as the retroviral HIV-1 Vpr. Failure to import Tf1-Gag into the nucleus in a nup124 null mutant resulted in complete loss of Tf1 transposition. Similarly, nuclear import of HIV-1 Vpr was impaired in nup124 null mutant strains and cells became resistant to Vpr's cell killing activity. On the basis of protein domain similarity, the human nucleoporin Nup153 was identified as a putative homolog of Nup124p. We demonstrate that in vitro-translated Nup124p and Nup153 coimmunoprecipitate Tf1 Gag or HIV-1 Vpr. Though full-length Nup153 was unable to complement the Tf1 transposition defect in a nup124 null mutant, we provide evidence that both nucleoporins share a unique N-terminal domain, Nup124p(AA264-454) and Nup153(AA448-634) that is absolutely essential for Tf1 transposition. Epigenetic overexpression of this domain in a wild-type (nup124(+)) background blocked Tf1 activity implying that sequences from Nup124p and the human Nup153 challenged the same pathway affecting Tf1 transposition. Our results establish a unique relationship between two analogous nucleoporins Nup124p and Nup153 wherein the function of a common domain in retrotransposition is conserved. PMID- 15659640 TI - Evolution of DNA sequence nonhomologies among maize inbreds. AB - Allelic chromosomal regions totaling more than 2.8 Mb and located on maize (Zea mays) chromosomes 1L, 2S, 7L, and 9S have been sequenced and compared over distances of 100 to 350 kb between the two maize inbred lines Mo17 and B73. The alleles contain extended regions of nonhomology. On average, more than 50% of the compared sequence is noncolinear, mainly because of the insertion of large numbers of long terminal repeat (LTR)-retrotransposons. Only 27 LTR-retroelements are shared between alleles, whereas 62 are allele specific. The insertion of LTR retrotransposons into the maize genome is statistically more recent for nonshared than shared ones. Most surprisingly, more than one-third of the genes (27/72) are absent in one of the inbreds at the loci examined. Such nonshared genes usually appear to be truncated and form clusters in which they are oriented in the same direction. However, the nonshared genome segments are gene-poor, relative to regions shared by both inbreds, with up to 12-fold difference in gene density. By contrast, miniature inverted terminal repeats (MITEs) occur at a similar frequency in the shared and nonshared fractions. Many times, MITES are present in an identical position in both LTRs of a retroelement, indicating that their insertion occurred before the replication of the retroelement in question. Maize ESTs and/or maize massively parallel signature sequencing tags were identified for the majority of the nonshared genes or homologs of them. In contrast with shared genes, which are usually conserved in gene order and location relative to rice (Oryza sativa), nonshared genes violate the maize colinearity with rice. Based on this, insertion by a yet unknown mechanism, rather than deletion events, seems to be the origin of the nonshared genes. The intergenic space between conserved genes is enlarged up to sixfold in maize compared with rice. Frequently, retroelement insertions create a different sequence environment adjacent to conserved genes. PMID- 15659642 TI - Mystique is a new insulin-like growth factor-I-regulated PDZ-LIM domain protein that promotes cell attachment and migration and suppresses Anchorage-independent growth. AB - By comparing differential gene expression in the insulin-like growth factor (IGF) IR null cell fibroblast cell line (R- cells) with cells overexpressing the IGF-IR (R+ cells), we identified the Mystique gene expressed as alternatively spliced variants. The human homologue of Mystique is located on chromosome 8p21.2 and encodes a PDZ LIM domain protein (PDLIM2). GFP-Mystique was colocalized at cytoskeleton focal contacts with alpha-actinin and beta1-integrin. Only one isoform of endogenous human Mystique protein, Mystique 2, was detected in cell lines. Mystique 2 was more abundant in nontransformed MCF10A breast epithelial cells than in MCF-7 breast carcinoma cells and was induced by IGF-I and cell adhesion. Overexpression of Mystique 2 in MCF-7 cells suppressed colony formation in soft agarose and enhanced cell adhesion to collagen and fibronectin. Point mutation of either the PDZ or LIM domain was sufficient to reverse suppression of colony formation, but mutation of the PDZ domain alone was sufficient to abolish enhanced adhesion. Knockdown of Mystique 2 with small interfering RNA abrogated both adhesion and migration in MCF10A and MCF-7 cells. The data indicate that Mystique is an IGF-IR-regulated adapter protein located at the actin cytoskeleton that is necessary for the migratory capacity of epithelial cells. PMID- 15659643 TI - Atg11 links cargo to the vesicle-forming machinery in the cytoplasm to vacuole targeting pathway. AB - Proteins are selectively packaged into vesicles at specific sites and then delivered correctly to the various organelles where they function, which is critical to the proper physiology of each organelle. The precursor form of the vacuolar hydrolase aminopeptidase I is a selective cargo molecule of the cytoplasm to vacuole targeting (Cvt) pathway and autophagy. Precursor Ape1 along with its receptor Atg19 forms the Cvt complex, which is transported to the pre autophagosomal structure (PAS), the putative site of Cvt vesicle formation, in a process dependent on Atg11. Here, we show that this interaction occurs through the Atg11 C terminus; subsequent recruitment of the Cvt complex to the PAS depends on central regions within Atg11. Atg11 was shown to physically link several proteins, although the timing of these interactions and their importance are unknown. Our mapping shows that the Atg11 coiled-coil domains are involved in self-assembly and the interaction with other proteins, including two previously unidentified partners, Atg17 and Atg20. Atg11 mutants defective in the transport of the Cvt complex to the PAS affect the localization of other Atg components, supporting the idea that the cargo facilitates the organization of the PAS in selective autophagy. These findings suggest that Atg11 plays an integral role in connecting cargo molecules with components of the vesicle-forming machinery. PMID- 15659644 TI - Fission yeast mto2p regulates microtubule nucleation by the centrosomin-related protein mto1p. AB - From an insertional mutagenesis screen, we isolated a novel gene, mto2+, involved in microtubule organization in fission yeast. mto2Delta strains are viable but exhibit defects in interphase microtubule nucleation and in formation of the postanaphase microtubule array at the end of mitosis. The mto2Delta defects represent a subset of the defects displayed by cells deleted for mto1+ (also known as mod20+ and mbo1+), a centrosomin-related protein required to recruit the gamma-tubulin complex to cytoplasmic microtubule-organizing centers (MTOCs). We show that mto2p colocalizes with mto1p at MTOCs throughout the cell cycle and that mto1p and mto2p coimmunoprecipitate from cytoplasmic extracts. In vitro studies suggest that mto2p binds directly to mto1p. In mto2Delta mutants, although some aspects of mto1p localization are perturbed, mto1p can still localize to spindle pole bodies and the cell division site and to "satellite" particles on interphase microtubules. In mto1Delta mutants, localization of mto2p to all of these MTOCs is strongly reduced or absent. We also find that in mto2Delta mutants, cytoplasmic forms of the gamma-tubulin complex are mislocalized, and the gamma-tubulin complex no longer coimmunoprecipitates with mto1p from cell extracts. These experiments establish mto2p as a major regulator of mto1p-mediated microtubule nucleation by the gamma-tubulin complex. PMID- 15659645 TI - Postprenylation CAAX processing is required for proper localization of Ras but not Rho GTPases. AB - The CAAX motif at the C terminus of most monomeric GTPases is required for membrane targeting because it signals for a series of three posttranslational modifications that include isoprenylation, endoproteolytic release of the C terminal- AAX amino acids, and carboxyl methylation of the newly exposed isoprenylcysteine. The individual contributions of these modifications to protein trafficking and function are unknown. To address this issue, we performed a series of experiments with mouse embryonic fibroblasts (MEFs) lacking Rce1 (responsible for removal of the -AAX sequence) or Icmt (responsible for carboxyl methylation of the isoprenylcysteine). In MEFs lacking Rce1 or Icmt, farnesylated Ras proteins were mislocalized. In contrast, the intracellular localizations of geranylgeranylated Rho GTPases were not perturbed. Consistent with the latter finding, RhoGDI binding and actin remodeling were normal in Rce1- and Icmt deficient cells. Swapping geranylgeranylation for farnesylation on Ras proteins or vice versa on Rho proteins reversed the differential sensitivities to Rce1 and Icmt deficiency. These results suggest that postprenylation CAAX processing is required for proper localization of farnesylated Ras but not geranygeranylated Rho proteins. PMID- 15659646 TI - A minus-end-directed kinesin with plus-end tracking protein activity is involved in spindle morphogenesis. AB - Diverse kinesin motor proteins are involved in spindle function; however, the mechanisms by which they are targeted to specific sites within spindles are not well understood. Here, we show that a fusion between yellow fluorescent protein (YFP) and a minus-end-directed Kinesin-14 (C-terminal family) from Arabidopsis, ATK5, localizes to mitotic spindle midzones and regions rich in growing plus-ends within phragmoplasts. Notably, in Arabidopsis interphase cells, YFP::ATK5 localizes to microtubules with a preferential enrichment at growing plus-ends; indicating ATK5 is a plus-end tracking protein (+TIP). This +TIP activity is conferred by regions outside of the C-terminal motor domain, which reveals the presence of independent plus-end tracking and minus-end motor activities within ATK5. Furthermore, mitotic spindles of atk5 null mutant plants are abnormally broadened. Based on these data, we propose a model in which ATK5 uses plus-end tracking to reach spindle midzones, where it then organizes microtubules via minus-end-directed motor activity. PMID- 15659647 TI - Yos1p is a novel subunit of the Yip1p-Yif1p complex and is required for transport between the endoplasmic reticulum and the Golgi complex. AB - Yeast Yip1p is a member of a conserved family of transmembrane proteins that interact with Rab GTPases. Previous studies also have indicated a role for Yip1p in the biogenesis of endoplasmic reticulum (ER)-derived COPII transport vesicles. In this report, we describe the identification and characterization of the uncharacterized open reading frame YER074W-A as a novel multicopy suppressor of the thermosensitive yip1-4 strain. We have termed this gene Yip One Suppressor 1 (YOS1). Yos1p is essential for growth and for function of the secretory pathway; depletion or inactivation of Yos1p blocks transport between the ER and the Golgi complex. YOS1 encodes an integral membrane protein of 87 amino acids that is conserved in eukaryotes. Yos1p localizes to ER and Golgi membranes and is efficiently packaged into ER-derived COPII transport vesicles. Yos1p associates with Yip1p and Yif1p, indicating Yos1p is a novel subunit of the Yip1p-Yif1p complex. PMID- 15659648 TI - Isoform-specific subcellular localization among 14-3-3 proteins in Arabidopsis seems to be driven by client interactions. AB - In most higher eukaryotes, the predominantly phosphoprotein-binding 14-3-3 proteins are the products of a multigene family, with many organisms having 10 or more family members. However, current models for 14-3-3/phosphopeptide interactions suggest that there is little specificity among 14-3-3s for diverse phosphopeptide clients. Therefore, the existence of sequence diversity among 14-3 3s within a single organism begs questions regarding the in vivo specificities of the interactions between the various 14-3-3s and their clients. Chief among those questions is, Do the different 14-3-3 isoforms interact with different clients within the same cell? Although the members of the Arabidopsis 14-3-3 family of proteins typically contain highly conserved regions of sequence, they also display distinctive variability with deep evolutionary roots. In the current study, a survey of several Arabidopsis 14-3-3/GFP fusions revealed that 14-3-3s demonstrate distinct and differential patterns of subcellular distribution, by using trichomes and stomate guard cells as in vivo experimental cellular contexts. The effects of client interaction on 14-3-3 localization were further analyzed by disrupting the partnering with peptide and chemical agents. Results indicate that 14-3-3 localization is both isoform specific and highly dependent upon interaction with cellular clients. PMID- 15659649 TI - Regulation of membrane trafficking by a novel Cdc42-related protein in Caenorhabditis elegans epithelial cells. AB - Rho GTPases are mainly known for their implication in cytoskeleton remodeling. They have also been recently shown to regulate various aspects of membrane trafficking. Here, we report the identification and the characterization of a novel Caenorhabditis elegans Cdc42-related protein, CRP-1, that shows atypical enzymatic characteristics in vitro. Expression in mouse fibroblasts revealed that, in contrast with CDC-42, CRP-1 was unable to reorganize the actin cytoskeleton and mainly localized to trans-Golgi network and recycling endosomes. This subcellular localization, as well as its expression profile restricted to a subset of epithelial-like cells in C. elegans, suggested a potential function for this protein in polarized membrane trafficking. Consistent with this hypothesis, alteration of CRP-1 expression affected the apical trafficking of CHE-14 in vulval and rectal epithelial cells and sphingolipids (C(6)-NBD-ceramide) uptake and/or trafficking in intestinal cells. However, it did not affect basolateral trafficking of myotactin in the pharynx and the targeting of IFB-2 and AJM-1, two cytosolic apical markers of intestine epithelial cells. Hence, our data demonstrate a function for CRP-1 in the regulation of membrane trafficking in a subset of cells with epithelial characteristics. PMID- 15659650 TI - p53 C-terminal phosphorylation by CHK1 and CHK2 participates in the regulation of DNA-damage-induced C-terminal acetylation. AB - The tumor suppressor protein p53 mediates stress-induced growth arrest or apoptosis and plays a major role in safeguarding genome integrity. In response to DNA damage, p53 can be modified at multiple sites by phosphorylation and acetylation. We report on the characterization of p53 C-terminal phosphorylation by CHK1 and CHK2, two serine/threonine (Ser/Thr) protein kinases, previously implicated in the phosphorylation of the p53 N terminus. Using tryptic phosphopeptide mapping, we have identified six additional CHK1 and CHK2 sites residing in the final 100 amino acids of p53. Phosphorylation of at least three of these sites, Ser366, Ser378, and Thr387, was induced by DNA damage, and the induction at Ser366 and Thr387 was abrogated by small interfering RNA targeting chk1 and chk2. Furthermore, mutation of these phosphorylation sites has a different impact on p53 C-terminal acetylation and on the activation of p53 targeted promoters. Our results demonstrate a possible interplay between p53 C terminal phosphorylation and acetylation, and they provide an additional mechanism for the control of the activity of p53 by CHK1 and CHK2. PMID- 15659651 TI - Dynamic recruitment of Nek2 kinase to the centrosome involves microtubules, PCM 1, and localized proteasomal degradation. AB - Centrosomes undergo dramatic changes in composition and activity during cell cycle progression. Yet mechanisms involved in recruiting centrosomal proteins are poorly understood. Nek2 is a cell cycle-regulated protein kinase required for regulation of centrosome structure at the G2/M transition. Here, we have addressed the processes involved in trafficking of Nek2 to the centrosome of human adult cells. We find that Nek2 exists in small, highly dynamic cytoplasmic particles that move to and from the centrosome. Many of these particles align along microtubules and a motif was identified in the Nek2 C-terminal noncatalytic domain that allows both microtubule binding and centrosome localization. FRAP experiments reveal that 70% of centrosomal Nek2 is rapidly turned over (t(1/2) approximately 3 s). Microtubules facilitate Nek2 trafficking to the centrosome but only over long distances. Cytoplasmic Nek2 particles colocalize in part with PCM-1 containing centriolar satellites and depletion of PCM-1 interferes with centrosomal recruitment of Nek2 and its substrate C-Nap1. Finally, we show that proteasomal degradation is necessary to allow rapid recruitment of new Nek2 molecules to the centrosome. Together, these data highlight multiple processes involved in regulating the abundance of Nek2 kinase at the centrosome including microtubule binding, the centriolar satellite component PCM-1, and localized protein degradation. PMID- 15659652 TI - A novel GTPase-activating protein for ARF6 directly interacts with clathrin and regulates clathrin-dependent endocytosis. AB - ADP-ribosylation factor 6 (Arf6) is a small-GTPase that regulates the membrane trafficking between the plasma membrane and endosome. It is also involved in the reorganization of the actin cytoskeleton. GTPase-activating protein (GAP) is a critical regulator of Arf function as it inactivates Arf. Here, we identified a novel species of GAP denoted as SMAP1 that preferentially acts on Arf6. Although overexpression of SMAP1 did not alter the subcellular distribution of the actin cytoskeleton, it did block the endocytosis of transferrin receptors. Knock down of endogenous SMAP1 also abolished transferrin internalization, which confirms that SMAP1 is needed for this endocytic process. SMAP1 overexpression had no effect on clathrin-independent endocytosis, however. Intriguingly, SMAP1 binds directly to the clathrin heavy chain via its clathrin-box and mutation studies revealed that its GAP domain and clathrin-box both contribute to the role SMAP1 plays in clathrin-dependent endocytosis. These observations suggest that SMAP1 may be an Arf6GAP that specifically regulates one of the multiple functions of Arf6, namely, clathrin-dependent endocytosis, and that it does so by binding directly to clathrin. PMID- 15659653 TI - The LIM/homeodomain protein Islet1 recruits Janus tyrosine kinases and signal transducer and activator of transcription 3 and stimulates their activities. AB - Islet1 (Isl1) belongs to the LIM homeodomain transcription factor family. Its roles in differentiation of motor neurons and organogenesis of pancreas and heart have been revealed. However, less is known about its regulatory mechanism and the target genes. In this study, we identified interactions between Isl1 and Janus tyrosine kinase (JAK), as well as signal transducer and activator of transcription (Stat)3, but not Stat1 and Stat5, in mammalian cells. We found that Isl1 not only forms a complex with Jak1 and Stat3 but also triggers the tyrosine phosphorylation of Jak1 and its kinase activity, thereby elevating the tyrosine phosphorylation, DNA binding activity, and target gene expression of Stat3. In vivo, the tyrosine-phosphorylated Stat3 was colocalized with Isl1 in the nucleus of the mouse motor neurons in spinal cord after nerve injury. Correspondingly, electroporation of Isl1 and Stat3 into the neural tube of chick embryos resulted in the activation of a reporter gene expression controlled by a Stat3 regulatory sequence, and cotransfection of Isl1 and Stat3 promoted the proliferation of the mouse motor neuron cells. Our data suggest a novel role of Isl1 as an adaptor for Jak1 and Stat3 and reveal a possible functional link between LIM homeodomain transcription factors and the Jak-Stat pathway. PMID- 15659655 TI - Multiple protein interactions involving proposed extracellular loop domains of the tight junction protein occludin. AB - Occludin is a tetraspan integral membrane protein in epithelial and endothelial tight junction (TJ) structures that is projected to have two extracellular loops. We have used peptides emulating central regions of human occludin's first and second loops, termed O-A:101-121 and O-B:210-228, respectively, to examine potential molecular interactions between these two regions of occludin and other TJ proteins. A superficial biophysical assessment of A:101-121 and O-B:210-228 showed them to have dissimilar solution conformation characteristics. Although O A:101-121 failed to strongly interact with protein components of the human epithelial intestinal cell line T84, O-B:210-228 selectively associated with occludin, claudin-one and the junctional adhesion molecule (JAM)-A. Further, the presence of O-B:210-228, but not O-A:101-121, impeded the recovery of functional TJ structures. A scrambled peptide sequences of O-B:210-228 failed to influence TJ assembly. These studies demonstrate distinct properties for these two extracellular segments of the occludin protein and provide an improved understanding of how specific domains of occludin may interact with proteins present at TJ structures. PMID- 15659654 TI - Transcriptional regulation by a DNA-associated form of cyclin D1. AB - Besides its function as a cell cycle regulator, cyclin D1 interacts with transcription factors to regulate gene activation. In this study, we show that cyclin D1 is recruited to the p21waf1 promoter by a STAT3-NcoA complex. The association of cyclin D1 with DNA prevented the recruitment of the CBP histone acetylase and RNA polymerase II, leading to an inhibition of the p21waf1 gene. Confirming the transcriptional function of the protein, the expression of the p21waf1 gene was enhanced in cyclin D1-/- fibroblasts or upon siRNA-mediated down regulation of the cyclin. Moreover, the STAT3-mediated activation of p21waf1 was also inhibited in breast cancer cells containing elevated levels of cyclin D1. Altogether, these results suggest that the transcriptional activities of cyclin D1 might play an important role in the regulation of cell-cycle regulatory genes and that these functions are probably involved in cell transformation. PMID- 15659656 TI - Isofrequency band-like zones of activation revealed by optical imaging of intrinsic signals in the cat primary auditory cortex. AB - Neurons of similar frequency preference are arranged in isofrequency bands (IFBs) across the primary auditory cortex (AI) of many mammals. Across the AI of the cat, one of the most frequently studied species for auditory anatomy and function, we demonstrate IFB-like responses using optical imaging of intrinsic signals (OIS). Optically defined activations were extensively elongated along the dorsoventral axis of AI (the ratio of the major and minor axes was approximately 2:1), and systematically shifted as a function of stimulus frequency. The elongation of this IFB-like zone was more conspicuous at higher frequencies. In the ventral sector of the imaged field, the IFB-like zones of activation evoked at different pure tone frequencies tended to overlap extensively. Electrophysiological recording from loci within the optically defined zones of activation revealed matched responses to the frequencies used for optical imaging at 65% of these loci. The dorsoventral orientation of these zones of activation was also closely matched with the orientation of tangentially spreading intrinsic axon terminals, as revealed anatomically. The visualization of IFB-like architecture and tonotopic organization by OIS provides a basic framework for investigating the relationships of different spectral channels and between multiple acoustic parameters at a neuronal population level. PMID- 15659657 TI - Coherent oscillatory activity in monkey area v4 predicts successful allocation of attention. AB - Attention serves to select objects from often complex scenes for enhanced processing and perception. In particular, the perception of shape depends critically on attention for integrating the various parts of the selected object into a coherent representation of object shape. To study whether oscillatory neuronal synchrony may serve as a mechanism of attention in shape perception, we introduced a novel shape-tracking task requiring sustained attention to a morphing shape. Attention was found to strongly increase oscillatory currents underlying the recorded field potentials in the gamma-frequency range, thus indicating enhanced neuronal synchrony within the population of V4 neurons representing the attended stimulus. Errors indicating a misdirection of attention to the distracter instead of the target were preceded by a corresponding shift of oscillatory activity from the target's neuronal representation to that of the distracter. No such effect was observed for errors unrelated to attention. Modulations of the attention-dependent enhancement of oscillatory activity occurred in correspondence with changing attentional demands during the course of a trial. The specificity of the effect of attentional errors together with the close coupling between attentional demand and oscillatory activity support the hypothesis that oscillatory neuronal synchrony serves as a mechanism of attention. PMID- 15659658 TI - Acetic acid induces expression of the Staphylococcus aureus cidABC and lrgAB murein hydrolase regulator operons. AB - The Staphylococcus aureus lrg and cid operons encode homologous proteins that regulate extracellular murein hydrolase activity and penicillin tolerance in a diametrically opposing manner. Although their specific regulatory functions remain unknown, it has been postulated that the functions of CidA and LrgA are analogous to those of bacteriophage holins and antiholins, respectively, and that these proteins serve as molecular control elements of bacterial programmed cell death. Although these studies demonstrated that cidBC transcription is abundant in sigmaB-proficient strains, cidABC transcription was only minimally expressed under standard growth conditions. In this study, we demonstrate that cidABC and lrgAB transcription in the clinical isolate UAMS-1 is induced by growth in the presence of 35 mM glucose and that this enhances murein hydrolase activity and decreases tolerance to vancomycin and rifampin. The effect of glucose on murein hydrolase activity was not observed in the cidA mutant, indicating that the induction of this activity was dependent on enhanced cidABC expression. Furthermore, we demonstrate that the effects of glucose on cidABC and lrgAB transcription are mediated by the generation of acetic acid produced by the metabolism of this and other carbon sources. These results shed new light on the control of the S. aureus cidABC and lrgAB genes and demonstrate that these operons, as well as murein hydrolase activity and antibiotic tolerance, are responsive to carbohydrate metabolism. PMID- 15659659 TI - Expression and functional analysis of the subtilin immunity genes spaIFEG in the subtilin-sensitive host Bacillus subtilis MO1099. AB - Bacillus subtilis ATCC 6633 produces the cationic pore-forming lantibiotic subtilin, which preferentially acts on gram-positive microorganisms; self protection of the producer cells is mediated by the four genes spaIFEG. To elucidate the mechanism of subtilin autoimmunity, we transferred different combinations of subtilin immunity genes under the control of an inducible promoter into the genome of subtilin-sensitive host strain B. subtilis MO1099. Recipient cells acquired subtilin tolerance through expression of either spaI or spaFEG, which shows that subtilin immunity is based on two independently acting systems. Cells coordinately expressing all four immunity genes acquired the strongest subtilin protection level. Quantitative in vivo peptide release assays demonstrated that SpaFEG diminished the quantity of cell-associated subtilin, suggesting that SpaFEG transports subtilin molecules from the membrane into the extracellular space. Homology and secondary structure analyses define SpaFEG as a prototype of lantibiotic immunity transporters that fall into the ABC-2 subfamily of multidrug resistance proteins. Membrane localization of the lipoprotein SpaI and specific interaction of SpaI with the cognate lantibiotic subtilin suggest a function of SpaI as a subtilin-intercepting protein. This interpretation was supported by hexahistidine-mediated 0-A cross-linking between hexahistidine tagged SpaI and subtilin. PMID- 15659660 TI - Disparate subcellular localization patterns of Pseudomonas aeruginosa Type IV pilus ATPases involved in twitching motility. AB - The opportunistic pathogen Pseudomonas aeruginosa expresses polar type IV pili (TFP), which are responsible for adhesion to various materials and twitching motility on surfaces. Twitching occurs by alternate extension and retraction of TFP, which arise from assembly and disassembly of pilin subunits at the base of the pilus. The ATPase PilB promotes pilin assembly, while the ATPase PilT or PilU or both promote pilin dissociation. Fluorescent fusions to two of the three ATPases (PilT and PilU) were functional, as shown by complementation of the corresponding mutants. PilB and PilT fusions localized to both poles, while PilU fusions localized only to the piliated pole. To identify the portion of the ATPases required for localization, sequential C-terminal deletions of PilT and PilU were generated. The conserved His and Walker B boxes were dispensable for polar localization but were required for twitching motility, showing that localization and function could be uncoupled. Truncated fusions that retained polar localization maintained their distinctive distribution patterns. To dissect the cellular factors involved in establishing polarity, fusion protein localization was monitored with a panel of TFP mutants. The localization of yellow fluorescent protein (YFP)-PilT and YFP-PilU was independent of the subunit PilA, other TFP ATPases, and TFP-associated proteins previously shown to be associated with the membrane or exhibiting polar localization. In contrast, YFP PilB exhibited diffuse cytoplasmic localization in a pilC mutant, suggesting that PilC is required for polar localization of PilB. Finally, localization studies performed with fluorescent ATPase chimeras of PilT and PilU demonstrated that information responsible for the characteristic localization patterns of the ATPases likely resides in their N termini. PMID- 15659661 TI - Mutants with temperature-sensitive defects in the Escherichia coli mismatch repair system: sensitivity to mispairs generated in vivo. AB - We have used direct selections to generate large numbers of mutants of Escherichia coli defective in the mismatch repair system and have screened these to identify mutants with temperature-sensitive defects. We detected and sequenced mutations that give rise to temperature-sensitive MutS, MutL, and MutH proteins. One mutation, mutS60, results in almost normal levels of spontaneous mutations at 37 degrees C but above this temperature gives rise to higher and higher levels of mutations, reaching the level of null mutations in mutS at 43 degrees C. However, at 37 degrees C the MutS60 protein can be much more easily titrated by mispairs than the wild-type MutS, as evidenced by the impaired ability to block homologous recombination in interspecies crosses and the increased levels of mutations from weak mutator alleles of mutD (dnaQ), mutC, and ndk. Strains with mutS60 can detect mispairs generated during replication that lead to mutation with much greater sensitivity than wild-type strains. The findings with ndk, lacking nucleotide diphosphate kinase, are striking. An ndk mutS60 strain yields four to five times the level of mutations seen in a full knockout of mutS. These results pose the question of whether similar altered Msh2 proteins result from presumed polymorphisms detected in tumor lines. The role of allele interactions in human disease susceptibility is discussed. PMID- 15659662 TI - Identification and characterization of genes involved in the downstream degradation pathway of gamma-hexachlorocyclohexane in Sphingomonas paucimobilis UT26. AB - Sphingomonas paucimobilis UT26 utilizes gamma-hexachlorocyclohexane (gamma-HCH) as a sole source of carbon and energy. In our previous study, we cloned and characterized genes that are involved in the conversion of gamma-HCH to maleylacetate (MA) via chlorohydroquinone (CHQ) in UT26. In this study, we identified and characterized an MA reductase gene, designated linF, that is essential for the utilization of gamma-HCH in UT26. A gene named linEb, whose deduced product showed significant identity to LinE (53%), was located close to linF. LinE is a novel type of ring cleavage dioxygenase that catalyzes the conversion of CHQ to MA. LinEb expressed in Escherichia coli transformed CHQ and 2,6-dichlorohydroquinone to MA and 2-chloromaleylacetate, respectively. Our previous and present results indicate that UT26 (i) has two gene clusters for degradation of chlorinated aromatic compounds via hydroquinone-type intermediates and (ii) uses at least parts of both clusters for gamma-HCH utilization. PMID- 15659663 TI - Characterization of a truncated lipoarabinomannan from the Actinomycete Turicella otitidis. AB - Lipoarabinomannan (LAM) lipoglycans have been characterized from a range of mycolic acid-containing actinomycetes and from the amycolate actinomycete Amycolatopsis sulphurea. To further understand the structural diversity of this family, we have characterized the lipoglycan of the otic commensal Turicella otitidis. T. otitidis LAM (TotLAM) has been determined to consist of a mannosyl phosphatidylinositol anchor unit carrying an (alpha 1-->6)-linked mannan core and substituted with terminal-arabinosyl branches. Thus, TotLAM has a novel truncated LAM structure. Using the human monocytic THP-1 cell line, it was found that TotLAM exhibited only minimal ability to induce tumor necrosis factor alpha. These findings contribute further to our understanding of actinomycete LAM diversity and allow further speculation as to the correlation between LAM structure and the immunomodulatory activities of these lipoglycans. PMID- 15659664 TI - Pyruvate:quinone oxidoreductase from Corynebacterium glutamicum: purification and biochemical characterization. AB - Pyruvate:quinone oxidoreductase catalyzes the oxidative decarboxylation of pyruvate to acetate and CO2 with a quinone as the physiological electron acceptor. So far, this enzyme activity has been found only in Escherichia coli. Using 2,6-dichloroindophenol as an artificial electron acceptor, we detected pyruvate:quinone oxidoreductase activity in cell extracts of the amino acid producer Corynebacterium glutamicum. The activity was highest (0.055 +/- 0.005 U/mg of protein) in cells grown on complex medium and about threefold lower when the cells were grown on medium containing glucose, pyruvate, or acetate as the carbon source. From wild-type C. glutamicum, the pyruvate:quinone oxidoreductase was purified about 180-fold to homogeneity in four steps and subjected to biochemical analysis. The enzyme is a flavoprotein, has a molecular mass of about 232 kDa, and consists of four identical subunits of about 62 kDa. It was activated by Triton X-100, phosphatidylglycerol, and dipalmitoyl phosphatidylglycerol, and the substrates were pyruvate (kcat=37.8 +/- 3 s(-1); Km=30 +/- 3 mM) and 2-oxobutyrate (kcat=33.2 +/- 3 s(-1); Km=90 +/- 8 mM). Thiamine pyrophosphate (Km=1 microM) and certain divalent metal ions such as Mg2+ (Km=29 microM), Mn2+ (Km=2 microM), and Co2+ (Km=11 microM) served as cofactors. In addition to several dyes (2,6-dichloroindophenol, p-iodonitrotetrazolium violet, and nitroblue tetrazolium), menadione (Km=106 microM) was efficiently reduced by the purified pyruvate:quinone oxidoreductase, indicating that a naphthoquinone may be the physiological electron acceptor of this enzyme in C. glutamicum. PMID- 15659665 TI - Different foreign genes incidentally integrated into the same locus of the Streptococcus suis genome. AB - Some strains of Streptococcus suis possess a type II restriction-modification (RM) system, whose genes are thought to be inserted into the genome between purH and purD from a foreign source by illegitimate recombination. In this study, we characterized the purHD locus of the S. suis genomes of 28 serotype reference strains by DNA sequencing. Four strains contained the RM genes in the locus, as described before, whereas 11 strains possessed other genetic regions of seven classes. The genetic regions contained a single gene or multiple genes that were either unknown or similar to hypothetical genes of other bacteria. The mutually exclusive localization of the genetic regions with the atypical G+C contents indicated that these regions were also acquired from foreign sources. No transposable element or long-repeat sequence was found in the neighboring regions. An alignment of the nucleotide sequences, including the RM gene regions, suggested that the foreign regions were integrated by illegitimate recombination via short stretches of nucleotide identity. By using a thermosensitive suicide plasmid, the RM genes were experimentally introduced into an S. suis strain that did not contain any foreign genes in that locus. Integration of the plasmid into the S. suis genome did not occur in the purHD locus but occurred at various chromosomal loci, where there were 2 to 10 bp of nucleotide identity between the chromosome and the plasmid. These results suggest that various foreign genes described here were incidentally integrated into the same locus of the S. suis genome. PMID- 15659666 TI - Heat shock proteome analysis of wild-type Corynebacterium glutamicum ATCC 13032 and a spontaneous mutant lacking GroEL1, a dispensable chaperone. AB - Proteome analysis of Corynebacterium glutamicum ATCC 13032 showed that levels of several proteins increased drastically in response to heat shock. These proteins were identified as DnaK, GroEL1, GroEL2, ClpB, GrpE, and PoxB, and their heat response was in agreement with previous transcriptomic results. A major heat induced protein was absent in the proteome of strain 13032B of C. glutamicum, used for genome sequencing in Germany, compared with the wild-type ATCC 13032 strain. The missing protein was identified as GroEL1 by matrix-assisted laser desorption ionization-time of flight peptide mass fingerprinting, and the mutation was found to be due to an insertion sequence, IsCg1, that was integrated at position 327 downstream of the translation start codon of the groEL1 gene, resulting in a truncated transcript of this gene, as shown by Northern analysis. The GroEL1 chaperone is, therefore, dispensable in C. glutamicum. On the other hand, GroEL2 appears to be essential for growth. Based on these results, the role of the duplicate groEL1 and groEL2 genes is analyzed. PMID- 15659667 TI - Characterization of the RokA and HexA broad-substrate-specificity hexokinases from Bacteroides fragilis and their role in hexose and N-acetylglucosamine utilization. AB - Bacteroides fragilis, a human gastrointestinal commensal and an opportunistic pathogen, utilizes simple and complex sugars and polysaccharides for growth in the large intestine and at sites of infection. Because B. fragilis lacks transport-linked sugar phosphorylation systems, cytoplasmic kinase(s) was expected to be required for the phosphorylation of hexoses and hexosamines. We have now identified two hexose kinases that are important for growth of B. fragilis on glucose, mannose, and other sugars. One kinase (RokA), a member of the ROK family of proteins, was found to be the sole kinase for activation of N acetyl-D-glucosamine (NAG). The other kinase (HexA) is responsible for the majority of the glucose kinase activity in the cell, although a hexA deletion mutant strain was not defective for growth on any substrate tested. Deletion of both the rokA and hexA kinase genes resulted in inability of the cell to use glucose, mannose, NAG, and many other sugars. We purified RokA and determined its approximate molecular mass to be 36.5 kDa. The purified RokA protein was shown to phosphorylate several substrates, including glucose, NAG, and mannose, but not N acetylmannosamine or N-acetylneuraminic acid. Phylogenetic analysis of RokA showed that it is most similar to kinases from the Cytophaga-Flavibacterium Bacteroides group, while HexA was most similar to other bacterial hexokinases and eukaryotic hexokinases. PMID- 15659668 TI - Trimeric structure of major outer membrane proteins homologous to OmpA in Porphyromonas gingivalis. AB - The major outer membrane proteins Pgm6 (41 kDa) and Pgm7 (40 kDa) of Porphyromonas gingivalis ATCC 33277 are encoded by open reading frames pg0695 and pg0694, respectively, which form a single operon. Pgm6 and Pgm7 (Pgm6/7) have a high degree of similarity to Escherichia coli OmpA in the C-terminal region and are predicted to form eight-stranded beta-barrels in the N-terminal region. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis, Pgm6/7 appear as bands with apparent molecular masses of 40 and 120 kDa, with and without a reducing agent, suggesting a monomer and trimer, respectively. To verify the predicted trimeric structure and function of Pgm6/7, we constructed three mutants with pg0695, pg0694, or both deleted. The double mutant produced no Pgm6/7. The single deletion mutants appeared to contain less Pgm7 and Pgm6 and to form homotrimers that migrated slightly faster (115 kDa) and slower (130 kDa), respectively, than wild-type Pgm6/7 under nonreducing conditions. N-terminal amino acid sequencing and mass spectrometry analysis of partially digested Pgm6/7 detected only fragments from Pgm6 and Pgm7. Two-dimensional, diagonal electrophoresis and chemical cross-linking experiments with or without a reducing agent clearly showed that Pgm6/7 mainly form stable heterotrimers via intermolecular disulfide bonds. Furthermore, growth retardation and arrest of the three mutants and increased permeability of their outer membranes indicated that Pgm6/7 play an important role in outer membrane integrity. Based on results of liposome swelling experiments, these proteins are likely to function as a stabilizer of the cell wall rather than as a major porin in this organism. PMID- 15659669 TI - Transcriptional regulation of sitABCD of Salmonella enterica serovar Typhimurium by MntR and Fur. AB - Salmonella enterica serovar Typhimurium has two manganese transport systems, MntH and SitABCD. MntH is a bacterial homolog of the eukaryotic natural resistance associated macrophage protein 1 (Nramp1), and SitABCD is an ABC-type transporter. Previously we showed that mntH is negatively controlled at the transcriptional level by the trans-acting regulatory factors, MntR and Fur. In this study, we examined the transcriptional regulation of sitABCD and compared it to the transcriptional regulation of mntH by constructing lacZ fusions to the promoter regions with and without mutations in putative MntR and/or Fur binding sites. The presence of Mn caused transcriptional repression of the sitABCD and mntH promoters primarily via MntR, but Fur was also capable of some repression in response to Mn. Likewise, Fe in the medium repressed transcription of both sit and mntH primarily via Fur, although MntR was also involved in this response. Transcriptional control by MntR and Fur was disrupted by site-specific mutations in the putative MntR and Fur binding sites, respectively. Transcription of the sit operon was also affected by the oxygen level and growth phase, but the increased expression observed under high oxygen conditions and higher cell densities is consistent with decreased availability of metals required for repression by the metalloregulatory proteins. PMID- 15659670 TI - Growth kinetics of extremely halophilic archaea (family halobacteriaceae) as revealed by arrhenius plots. AB - Members of the family Halobacteriaceae in the domain Archaea are obligate extreme halophiles. They occupy a variety of hypersaline environments, and their cellular biochemistry functions in a nearly saturated salty milieu. Despite extensive study, a detailed analysis of their growth kinetics is missing. To remedy this, Arrhenius plots for 14 type species of the family were generated. These organisms had maximum growth temperatures ranging from 49 to 58 degrees C. Nine of the organisms exhibited a single temperature optimum, while five grew optimally at more than one temperature. Generation times at these optimal temperatures ranged from 1.5 h (Haloterrigena turkmenica) to 3.0 h (Haloarcula vallismortis and Halorubrum saccharovorum). All shared an inflection point at 31 +/- 4 degrees C, and the temperature characteristics for 12 of the 14 type species were nearly parallel. The other two species (Natronomonas pharaonis and Natronorubrum bangense) had significantly different temperature characteristics, suggesting that the physiology of these strains is different. In addition, these data show that the type species for the family Halobacteriaceae share similar growth kinetics and are capable of much faster growth at higher temperatures than those previously reported. PMID- 15659671 TI - Conjugative transfer of the Lactococcus lactis chromosomal sex factor promotes dissemination of the Ll.LtrB group II intron. AB - The Ll.LtrB group II intron from the low-G+C gram-positive bacterium Lactococcus lactis was the first bacterial group II intron shown to splice and mobilize in vivo. This retroelement interrupts the relaxase gene (ltrB) of three L. lactis conjugative elements: plasmids pRS01 and pAH90 and the chromosomal sex factor. Conjugative transfer of a plasmid harboring a segment of the pRS01 conjugative plasmid including the Ll.LtrB intron allows dissemination of Ll.LtrB among L. lactis strains and lateral transfer of this retroelement from L. lactis to Enterococcus faecalis. Here we report the dissemination of the Ll.LtrB group II intron among L. lactis strains following conjugative transfer of the native chromosomally embedded L. lactis sex factor. We demonstrated that Ll.LtrB dissemination is highly variable and often more efficient from this integrative and conjugative element than from an engineered conjugative plasmid. Cotransfer among L. lactis strains of both Ll.LtrB-containing elements, the conjugative plasmid and the sex factor, was detected and shown to be synergistic. Moreover, following their concurrent transfer, both mobilizable elements supported the spread of their respective copies of the Ll.LtrB intron. Our findings explain the unusually high efficiency of Ll.LtrB mobility observed following conjugation of intron-containing plasmids. PMID- 15659672 TI - Pseudomonas-Saccharomyces interactions: influence of fungal metabolism on bacterial physiology and survival. AB - Fungal-bacterial interactions are ubiquitous, yet their molecular basis is only poorly understood. In this study, a novel beneficial interaction between a strain of Pseudomonas putida and the fungus Saccharomyces cerevisiae was identified. When the bacteria were incubated alone in grape juice or in synthetic medium containing various concentrations of glucose, they lost viability rapidly during stationary phase. However, when the bacteria were incubated in these media in the presence of the fungus, their stationary phase survival improved dramatically. On agar plates containing glucose, the beneficial effects of the fungus were manifested in robust bacterial growth and exopolysaccharide production that led to visible mucoidy. In contrast, bacteria grew poorly and were nonmucoid in such media in the absence of the fungus. By using the available S. cerevisiae deletion library, yeast mutants that were unable to mediate this beneficial interaction were identified. These mutants revealed that the beneficial effect on bacterial physiology and survival was mediated by the ability of the fungus to metabolize the available glucose and consequent effects on the medium's pH. In natural environments where the concentration of glucose is high, it is likely that the presence of fungi has had profound beneficial effects on the physiology and survival of certain P. putida strains throughout their natural history. PMID- 15659673 TI - Role of integration host factor in the transcriptional activation of flagellar gene expression in Caulobacter crescentus. AB - In the Caulobacter crescentus predivisional cell, class III and IV flagellar genes, encoding the extracytoplasmic components of the flagellum, are transcribed in the nascent swarmer compartment. This asymmetric expression pattern is attributable to the compartmentalized activity of the sigma54-dependent transcriptional activator FlbD. Additionally, these temporally transcribed flagellar promoters possess a consensus sequence for the DNA-binding protein integration host factor (IHF), located between the upstream FlbD binding site and the promoter sequences. Here, we deleted the C. crescentus gene encoding the beta subunit of the IHF, ihfB (himD), and examined the effect on flagellar gene expression. The DeltaihfB strain exhibited a mild defect in cell morphology and impaired motility. Using flagellar promoter reporter fusions, we observed that expression levels of a subset of class III flagellar promoters were decreased by the loss of IHF. However, one of these promoters, fliK-lacZ, exhibited a wild type cell cycle-regulated pattern of expression in the absence of IHF. Thus, IHF is required for maximal transcription of several late flagellar genes. The DeltaihfB strain was found to express significantly reduced amounts of the class IV flagellin, FljL, as a consequence of reduced transcriptional activity. Our results indicate that the motility defect exhibited by the DeltaihfB strain is most likely attributable to its failure to accumulate the class IV-encoded 27-kDa flagellin subunit, FljL. PMID- 15659674 TI - Substrate requirements for regulated intramembrane proteolysis of Bacillus subtilis pro-sigmaK. AB - During sporulation of Bacillus subtilis, pro-sigmaK is activated by regulated intramembrane proteolysis (RIP) in response to a signal from the forespore. RIP of pro-sigmaK removes its prosequence (amino acids 1 to 20), releasing sigmaK from the outer forespore membrane into the mother cell cytoplasm, in a reaction catalyzed by SpoIVFB, a metalloprotease in the S2P family of intramembrane cleaving proteases. The requirements for pro-sigmaK to serve as a substrate for RIP were investigated by producing C-terminally truncated pro-sigmaK fused at different points to the green fluorescent protein (GFP) or hexahistidine in sporulating B. subtilis or in Escherichia coli engineered to coexpress SpoIVFB. Nearly half of pro-sigmaK (amino acids 1 to 117), including part of sigma factor region 2.4, was required for RIP of pro-sigmaK-GFP chimeras in sporulating B. subtilis. Likewise, pro-sigmaK-hexahistidine chimeras demonstrated that the N terminal 117 amino acids of pro-sigma(K) are sufficient for RIP, although the N terminal 126 amino acids, which includes all of region 2.4, allowed much better accumulation of the chimeric protein in sporulating B. subtilis and more efficient processing by SpoIVFB in E. coli. In contrast to the requirements for RIP, a much smaller N-terminal segment (amino acids 1 to 27) was sufficient for membrane localization of a pro-sigmaK-GFP chimera. Addition or deletion of five amino acids near the N terminus allowed accurate processing of pro-sigmaK, ruling out a mechanism in which SpoIVFB measures the distance from the N terminus to the cleavage site. A charge reversal at position 13 (substituting glutamate for lysine) reduced accumulation of pro-sigmaK and prevented detectable RIP by SpoIVFB. These results elucidate substrate requirements for RIP of pro-sigmaK by SpoIVFB and may have implications for substrate recognition by other S2P family members. PMID- 15659675 TI - Markerless mutagenesis in Methanococcus maripaludis demonstrates roles for alanine dehydrogenase, alanine racemase, and alanine permease. AB - Among the archaea, Methanococcus maripaludis has the unusual ability to use L- or D-alanine as a nitrogen source. To understand how this occurs, we tested the roles of three adjacent genes encoding homologs of alanine dehydrogenase, alanine racemase, and alanine permease. To produce mutations in these genes, we devised a method for markerless mutagenesis that builds on previously established genetic tools for M. maripaludis. The technique uses a negative selection strategy that takes advantage of the ability of the M. maripaludis hpt gene encoding hypoxanthine phosphoribosyltransferase to confer sensitivity to the base analog 8 azahypoxanthine. In addition, we developed a negative selection method to stably incorporate constructs into the genome at the site of the upt gene encoding uracil phosphoribosyltransferase. Mutants with in-frame deletion mutations in the genes for alanine dehydrogenase and alanine permease lost the ability to grow on either isomer of alanine, while a mutant with an in-frame deletion mutation in the gene for alanine racemase lost only the ability to grow on D-alanine. The wild-type gene for alanine dehydrogenase, incorporated into the upt site, complemented the alanine dehydrogenase mutation. Hence, the permease is required for the transport of either isomer, the dehydrogenase is specific for the L isomer, and the racemase converts the D isomer to the L isomer. Phylogenetic analysis indicated that all three genes had been acquired by lateral gene transfer from the low-moles-percent G+C gram-positive bacteria. PMID- 15659676 TI - Functional interactions between the carbon and iron utilization regulators, Crp and Fur, in Escherichia coli. AB - In Escherichia coli, the ferric uptake regulator (Fur) controls expression of the iron regulon in response to iron availability while the cyclic AMP receptor protein (Crp) regulates expression of the carbon regulon in response to carbon availability. We here identify genes subject to significant changes in expression level in response to the loss of both Fur and Crp. Many iron transport genes and several carbon metabolic genes are subject to dual control, being repressed by the loss of Crp and activated by the loss of Fur. However, the sodB gene, encoding superoxide dismutase, and the aceBAK operon, encoding the glyoxalate shunt enzymes, show the opposite responses, being activated by the loss of Crp and repressed by the loss of Fur. Several other genes including the sdhA-D, sucA D, and fumA genes, encoding key constituents of the Krebs cycle, proved to be repressed by the loss of both transcription factors. Finally, the loss of both Crp and Fur activated a heterogeneous group of genes under sigmaS control encoding, for example, the cyclopropane fatty acid synthase, Cfa, the glycogen synthesis protein, GlgS, the 30S ribosomal protein, S22, and the mechanosensitive channel protein, YggB. Many genes appeared to be regulated by the two transcription factors in an apparently additive fashion, but apparent positive or negative cooperativity characterized several putative Crp/Fur interactions. Relevant published data were evaluated, putative Crp and Fur binding sites were identified, and representative results were confirmed by real-time PCR. Molecular explanations for some, but not all, of these effects are provided. PMID- 15659677 TI - Multiple regulators control expression of the Entner-Doudoroff aldolase (Eda) of Escherichia coli. AB - The Escherichia coli eda gene, which encodes the Entner-Doudoroff aldolase, is central to the catabolism of several sugar acids. Here, we show that Eda synthesis is induced by growth on gluconate, glucuronate, or methyl-beta-D glucuronide; phosphate limitation; and carbon starvation. Transcription of eda initiates from three promoters, designated P1, P2, and P4, each of which is responsible for induction under different growth conditions. P1 controls eda induction on gluconate and is regulated by GntR. P2 controls eda induction on glucuronate and galacturonate and is regulated by KdgR. P4 is active under conditions of phosphate starvation and is directly controlled by PhoB. In addition, CsrA activates Eda synthesis, apparently by an indirect mechanism that may be involved in the modest changes in expression level that are associated with carbon starvation. The complex regulation of eda is discussed with respect to its several physiological roles, which apparently accommodate not only sugar acid catabolism but also detoxification of metabolites that could accumulate during starvation-induced stress. PMID- 15659678 TI - Combined inactivation and expression strategy to study gene function under physiological conditions: application to identification of new Escherichia coli adhesins. AB - In bacteria, whereas disruption methods have been improved recently, the use of plasmid complementation strategies are still subject to limitations, such as cloning difficulties, nonphysiological levels of gene expression, or a requirement for antibiotics as plasmid selection pressure. Moreover, because of the pleiotropic modifications of cell physiology often induced by plasmid-based complementation, these strategies may introduce biases when biological process such as adhesion or biofilm formation are studied. We developed a plasmid-free approach that combines the lambda-red linear DNA recombination method with site directed insertion of a repression and expression (RExBAD) cassette which places a functional pBAD promoter upstream of a target gene. We showed that this method permits both inactivation and modulation of most Escherichia coli gene expression, including expression of toxin and essential genes. We used this strategy to study adhesion and bacterial biofilms by placing the RExBAD cassette in front of the tra operon, encoding the DNA transfer and pilus genes on the F conjugative plasmid, and in front of flu, the antigen 43 (Ag43) autotransporter adhesin-encoding gene. In silico analysis revealed the existence of 10 genes with homology to the Ag43 gene that were good candidates for genes that encode putative new adhesins in E. coli. We used the RExBAD strategy to study these genes and demonstrated that induction of expression of four of them is associated with adhesion of E. coli to abiotic surfaces. The potential use of the RExBAD approach to study the function of cryptic or uncharacterized genes in large-scale postgenomic functional analyses is discussed. PMID- 15659679 TI - Induction of rapid detachment in Shewanella oneidensis MR-1 biofilms. AB - Active detachment of cells from microbial biofilms is a critical yet poorly understood step in biofilm development. We discovered that detachment of cells from biofilms of Shewanella oneidensis MR-1 can be induced by arresting the medium flow in a hydrodynamic biofilm system. Induction of detachment was rapid, and substantial biofilm dispersal started as soon as 5 min after the stop of flow. We developed a confocal laser scanning microscopy-based assay to quantify detachment. The extent of biomass loss was found to be dependent on the time interval of flow stop and on the thickness of the biofilm. Up to 80% of the biomass of 16-h-old biofilms could be induced to detach. High-resolution microscopy studies revealed that detachment was associated with an overall loosening of the biofilm structure and a release of individual cells or small cell clusters. Swimming motility was not required for detachment. Although the loosening of cells from the biofilm structure was observed evenly throughout thin biofilms, the most pronounced detachment in thicker biofilms occurred in regions exposed to the flow of medium, suggesting a metabolic control of detachability. Deconvolution of the factors associated with the stop of medium flow revealed that a sudden decrease in oxygen tension is the predominant trigger for initiating detachment of individual cells. In contrast, carbon limitation did not trigger any substantial detachment, suggesting a physiological link between oxygen sensing or metabolism and detachment. In-frame deletions were introduced into genes encoding the known and putative global transcriptional regulators ArcA, CRP, and EtrA (FNR), which respond to changes in oxygen tension in S. oneidensis MR-1. Biofilms of null mutants in arcA and crp were severely impacted in the stop-of-flow-induced detachment response, suggesting a role for these genes in regulation of detachment. In contrast, an DeltaetrA mutant displayed a variable detachment phenotype. From this genetic evidence we conclude that detachment is a biologically controlled process and that a rapid change in oxygen concentration is a critical factor in detachment and, consequently, in dispersal of S. oneidensis cells from biofilms. Similar mechanisms might also operate in other bacteria. PMID- 15659680 TI - The Enterococcus faecalis sigV protein is an extracytoplasmic function sigma factor contributing to survival following heat, acid, and ethanol treatments. AB - Analysis of the genome sequence of Enterococcus faecalis allowed the identification of two genes whose protein products showed 33 and 34% identity with those of sigV and yrhM of Bacillus subtilis, respectively. These genes, named sigV and rsiV, are predicted to encode members of the extracytoplasmic function subfamily of eubacterial RNA polymerase sigma and anti-sigma factors, respectively. This group of sigma factors has been shown to regulate gene expression in response to stress conditions. sigV and rsiV were shown to be under the control of the same promoter. The transcriptional start site was determined, and the 1.5-kb mRNA transcript was shown to be overexpressed under glucose and complete starvation, as well as under physicochemical treatments. Three mutants, affected in sigV, rsiV, and both genes, were constructed by double-crossover recombination within the genome of E. faecalis strain JH2-2. Compared with the wild type and the rsiV mutant, the sigV mutants were more susceptible to heat shock, acid, and ethanol treatments and displayed decreased survival during long term starvation. A nisin-inducible sigV gene construction used in complementation assays restored the wild phenotype of the sigV mutants, confirming the involvement of SigV in the heat shock, ethanol, and acid stress responses. Northern blot analysis carried out with the three mutant strains revealed the inhibition of sigV expression by the related anti-sigma factor gene rsiV. In addition, putative candidates of the sigV regulon determined by computer search for the sigV promoter sequence were analyzed. PMID- 15659681 TI - Potassium transport in a halophilic member of the bacteria domain: identification and characterization of the K+ uptake systems TrkH and TrkI from Halomonas elongata DSM 2581T. AB - The halophilic bacterium Halomonas elongata accumulates K+, glutamate, and the compatible solute ectoine as osmoprotectants. By functional complementation of Escherichia coli mutants defective in K+ uptake, we cloned three genes that are required for K+ uptake in H. elongata. Two adjacent genes, named trkA (1,374 bp) and trkH (1,449 bp), were identified on an 8.5-kb DNA fragment, while a third gene, called trkI (1,479 bp), located at a different site in the H. elongata chromosome, was found on a second 8.5-kb fragment. The potential protein expressed by trkA is similar to the cytoplasmic NAD+/NADH binding protein TrkA from E. coli, which is required for the activity of the Trk K+ uptake system. The deduced amino acid sequences of trkH and trkI showed significant identity to the transmembrane protein of Trk transporters. K+ transport experiments with DeltatrkH and DeltatrkI mutants of H. elongata revealed that TrkI exhibits a Km value of 1.12 mM, while the TrkH system has a half-saturation constant of 3.36 mM. Strain KB12, relying on TrkH alone, accumulated K+ with a lower Vmax and required a higher K+ concentration for growth in highly saline medium than the wild type. Strain KB15, expressing only TrkI, showed the same phenotype and the same K+ transport kinetics as the wild type, proving that TrkI is the main K+ transport system in H. elongata. In the absence of both transporters TrkH and TrkI, K+ accumulation was not detectable. K+ transport was also abolished in a trkA deletion mutant, indicating that TrkI and TrkH depend on one type of TrkA protein. Reverse transcriptase PCR experiments and Northern hybridization analyses of the trkAH locus revealed cotranscription of trkAH as well as a monocistronic transcript with only trkA. PMID- 15659682 TI - Characterization of the pheromone response of the Enterococcus faecalis conjugative plasmid pCF10: complete sequence and comparative analysis of the transcriptional and phenotypic responses of pCF10-containing cells to pheromone induction. AB - The sex pheromone plasmids in Enterococcus faecalis are one of the most efficient conjugative plasmid transfer systems known in bacteria. Plasmid transfer rates can reach or exceed 10(-1) transconjugants per donor in vivo and under laboratory conditions. We report the completion of the DNA sequence of plasmid pCF10 and the analysis of the transcription profile of plasmid genes, relative to conjugative transfer ability following pheromone induction. These experiments employed a mini microarray containing all 57 open reading frames of pCF10 and a set of selected chromosomal genes. A clear peak of transcription activity was observed 30 to 60 min after pheromone addition, with transcription subsiding 2 h after pheromone induction. The transcript activity correlated with the ability of donor cells to transfer pCF10 to recipient cells. Remarkably, aggregation substance (Asc10, encoded by the prgB gene) was present on the cell surface for a long period of time after pheromone-induced transcription of prgB and plasmid transfer ability had ceased. This observation could have relevance for the virulence of E. faecalis. PMID- 15659683 TI - gpwac of the T4-type bacteriophages: structure, function, and evolution of a segmented coiled-coil protein that controls viral infectivity. AB - The wac gene product (gpwac) or fibritin of bacteriophage T4 forms the six fibers that radiate from the phage neck. During phage morphogenesis these whiskers bind the long tail fibers (LTFs) and facilitate their attachment to the phage baseplate. After the cell lysis, the gpwac fibers function as part of an environmental sensing device that retains the LTFs in a retracted configuration and thus prevents phage adsorption in unfavorable conditions. A comparative analysis of the sequences of 5 wac gene orthologs from various T4-type phages reveals that the approximately 50-amino-acid N-terminal domain is the only highly conserved segment of the protein. This sequence conservation is probably a direct consequence of the domain's strong and specific interactions with the neck proteins. The sequence of the central fibrous region of gpwac is highly plastic, with only the heptad periodicity of the coiled-coil structure being conserved. In the various gpwac sequences, the small C-terminal domain essential for initiation of the folding of T4 gpwac is replaced by unrelated sequences of unknown origin. When a distant T4-type phage has a novel C-terminal gpwac sequence, the phage's gp36 sequence that is located at the knee joint of the LTF invariably has a novel domain in its C terminus as well. The covariance of these two sequences is compatible with genetic data suggesting that the C termini of gpwac and gp36 engage in a protein-protein interaction that controls phage infectivity. These results add to the limited evidence for domain swapping in the evolution of phage structural proteins. PMID- 15659684 TI - Isolation and characterization of an aldehyde dehydrogenase encoded by the aldB gene of Escherichia coli. AB - An aldehyde dehydrogenase was detected in crude cell extracts of Escherichia coli DH5alpha. Growth studies indicated that the aldehyde dehydrogenase activity was growth phase dependent and increased in cells grown with ethanol. The N-terminal amino acid sequence of the purified enzyme identified the latter as an aldehyde dehydrogenase encoded by aldB, which was thought to play a role in the removal of aldehydes and alcohols in cells that were under stress. The purified enzyme showed an estimated molecular mass of 220 +/- 8 kDa, consisting of four identical subunits, and preferred to use NADP and acetaldehyde. MgCl2 increased the activity of the NADP-dependent enzyme with various substrates. A comparison of the effect of Mg2+ ions on the bacterial enzyme with the effect of Mg2+ ions on human liver mitochondrial aldehyde dehydrogenase revealed that the bacterial enzyme shared kinetic properties with the mammalian enzyme. An R197E mutant of the bacterial enzyme appeared to retain very little NADP-dependent activity on acetaldehyde. PMID- 15659685 TI - Sulfur and nitrogen limitation in Escherichia coli K-12: specific homeostatic responses. AB - We determined global transcriptional responses of Escherichia coli K-12 to sulfur (S)- or nitrogen (N)-limited growth in adapted batch cultures and cultures subjected to nutrient shifts. Using two limitations helped to distinguish between nutrient-specific changes in mRNA levels and common changes related to the growth rate. Both homeostatic and slow growth responses were amplified upon shifts. This made detection of these responses more reliable and increased the number of genes that were differentially expressed. We analyzed microarray data in several ways: by determining expression changes after use of a statistical normalization algorithm, by hierarchical and k-means clustering, and by visual inspection of aligned genome images. Using these tools, we confirmed known homeostatic responses to global S limitation, which are controlled by the activators CysB and Cbl, and found that S limitation propagated into methionine metabolism, synthesis of FeS clusters, and oxidative stress. In addition, we identified several open reading frames likely to respond specifically to S availability. As predicted from the fact that the ddp operon is activated by NtrC, synthesis of cross-links between diaminopimelate residues in the murein layer was increased under N limiting conditions, as was the proportion of tripeptides. Both of these effects may allow increased scavenging of N from the dipeptide D-alanine-D-alanine, the substrate of the Ddp system. PMID- 15659686 TI - The generalized transducing Salmonella bacteriophage ES18: complete genome sequence and DNA packaging strategy. AB - The generalized transducing double-stranded DNA bacteriophage ES18 has an icosahedral head and a long noncontractile tail, and it infects both rough and smooth Salmonella enterica strains. We report here the complete 46,900-bp genome nucleotide sequence and provide an analysis of the sequence. Its 79 genes and their organization clearly show that ES18 is a member of the lambda-like (lambdoid) phage group; however, it contains a novel set of genes that program assembly of the virion head. Most of its integration-excision, immunity, Nin region, and lysis genes are nearly identical to those of the short-tailed Salmonella phage P22, while other early genes are nearly identical to Escherichia coli phages lambda and HK97, S. enterica phage ST64T, or a Shigella flexneri prophage. Some of the ES18 late genes are novel, while others are most closely related to phages HK97, lambda, or N15. Thus, the ES18 genome is mosaically related to other lambdoid phages, as is typical for all group members. Analysis of virion DNA showed that it is circularly permuted and about 10% terminally redundant and that initiation of DNA packaging series occurs across an approximately 1-kbp region rather than at a precise location on the genome. This supports a model in which ES18 terminase can move substantial distances along the DNA between recognition and cleavage of DNA destined to be packaged. Bioinformatic analysis of large terminase subunits shows that the different functional classes of phage-encoded terminases can usually be predicted from their amino acid sequence. PMID- 15659687 TI - Regulation of virulence by a two-component system in group B streptococcus. AB - Group B Streptococcus (GBS) is frequently carried in the gastrointestinal or genitourinary tract as a commensal organism, yet it has the potential to cause life-threatening infection in newborn infants, pregnant women, and individuals with chronic illness. Regulation of virulence factor expression may affect whether GBS behaves as an asymptomatic colonizer or an invasive pathogen, but little is known about how such factors are controlled in GBS. We now report the characterization of a GBS locus that encodes a two-component regulatory system similar to CsrRS (or CovRS) in Streptococcus pyogenes. Inactivation of csrR, encoding the putative response regulator, in two unrelated wild-type strains of GBS resulted in a marked increase in production of beta-hemolysin/cytolysin and a striking decrease in production of CAMP factor, an unrelated cytolytic toxin. Quantitative RNA hybridization experiments revealed that these two phenotypes were associated with a marked increase and decrease in expression of the corresponding genes, cylE and cfb, respectively. The CsrR mutant strains also displayed increased expression of scpB encoding C5a peptidase. Similar, but less marked, changes in gene expression were observed in CsrS (putative sensor component) mutants, evidence that CsrR and CsrS constitute a functional two component system. Experimental infection studies in mice demonstrated reduced virulence of both CsrR and CsrS mutant strains relative to the wild type. Together, these results indicate that CsrRS regulates expression of multiple GBS virulence determinants and is likely to play an important role in GBS pathogenesis. PMID- 15659688 TI - Clustered genes required for the synthesis of heterocyst envelope polysaccharide in Anabaena sp. strain PCC 7120. AB - As demonstrated with alr2835 (hepA) and alr2834 (hepC) mutants, heterocysts of Anabaena sp. strain PCC 7120, a filamentous cyanobacterium, must have an envelope polysaccharide layer (the Hep+ phenotype) to fix dinitrogen in an oxygen containing milieu (the Fox+ phenotype). Transpositions presumptively responsible for a Fox- phenotype were localized in open reading frames (ORFs) near hepA and hepC. A mutation in each of nine of these ORFs was complemented by a clone bearing only that single, intact ORF. Heterocysts of the nine mutants were found to lack an envelope polysaccharide layer. Complementation of mutations in alr2832 and alr2840 may have resulted from recombination. However, alr2825, alr2827, alr2831, alr2833, alr2837, alr2839, and alr2841, like hepA and hepC, are required for a Hep+ Fox+ phenotype. PMID- 15659689 TI - Genome-wide transcriptional response of chemostat-cultured Escherichia coli to zinc. AB - Zinc is an essential trace metal ion for growth, but an excess of Zn is toxic and microorganisms express diverse resistance mechanisms. To understand global bacterial responses to excess Zn, we conducted transcriptome profiling experiments comparing Escherichia coli MG1655 grown under control conditions and cells grown with a toxic, sublethal ZnSO4 concentration (0.2 mM). Cultures were grown in a defined medium lacking inorganic phosphate, permitting maximum Zn bioavailability, and in glycerol-limited chemostats at a constant growth rate and pH. Sixty-four genes were significantly up-regulated by Zn stress, including genes known to be involved in Zn tolerance, particularly zntA, zraP, and hydG. Microarray transcriptome profiling was confirmed by real-time PCR determinations of cusF (involved in Ag and Cu efflux), ais (an Al-inducible gene), asr (encoding an acid shock-inducible periplasmic protein), cpxP (a periplasmic chaperone gene), and basR. Five up-regulated genes, basR and basS [encoding a sensor regulator implicated in Salmonella in Fe(III) sensing and antibiotic resistance], fliM (flagellar synthesis), and ycdM and yibD (both with unknown functions), are important for growth resistance to zinc, since mutants with mutations in these genes exhibited zinc sensitivity in liquid media and on metal gradient plates. Fifty-eight genes were significantly down-regulated by Zn stress; notably, several of these genes were involved in protection against acid stress. Since the mdt operon (encoding a multidrug resistance pump) was also up-regulated, these findings have important implications for understanding not only Zn homeostasis but also how bacterial antibiotic resistance is modulated by metal ions. PMID- 15659691 TI - Sinorhizobium meliloti dctA mutants with partial ability to transport dicarboxylic acids. AB - Sinorhizobium meliloti dctA encodes a transport protein needed for a successful nitrogen-fixing symbiosis between the bacteria and alfalfa. Using the toxicity of the DctA substrate fluoroorotic acid as a selective agent in an iterated selection procedure, four independent S. meliloti dctA mutants were isolated that retained some ability to transport dicarboxylates. Two mutations were located in a region called motif B located in a predicted transmembrane helix of the protein that has been shown in other members of the glutamate transporter family to be involved in cation binding. A G114D mutation was located in the third transmembrane helix, which had not previously been directly implicated in transport. Multiple sequence alignment of more than 60 members of the glutamate transporter family revealed a glycine at this position in nearly all members of the family. The fourth mutant was able to transport succinate at almost wild-type levels but was impaired in malate and fumarate transport. It contains two mutations: one in a periplasmic domain and the other predicted to be in the cytoplasm. Separation of the mutations showed that each contributed to the altered substrate preference. dctA deletion mutants that contain the mutant dctA alleles on a plasmid can proceed further in symbiotic development than null mutants of dctA, but none of the plasmids could support symbiotic nitrogen fixation, although they can transport dicarboxylates, some at relatively high levels. PMID- 15659690 TI - Genome-wide expression analysis indicates that FNR of Escherichia coli K-12 regulates a large number of genes of unknown function. AB - The major regulator controlling the physiological switch between aerobic and anaerobic growth conditions in Escherichia coli is the DNA binding protein FNR. To identify genes controlled by FNR, we used Affymetrix Antisense GeneChips to compare global gene expression profiles from isogenic MG1655 wild-type and Deltafnr strains grown in glucose minimal media under aerobic or anaerobic conditions. We found that 297 genes contained within 184 operons were regulated by FNR and/or by O2 levels. The expression of many genes known to be involved in anaerobic respiration and fermentation was increased under anaerobic growth conditions, while that of genes involved in aerobic respiration and the tricarboxylic acid cycle were repressed as expected. The expression of nine operons associated with acid resistance was also increased under anaerobic growth conditions, which may reflect the production of acidic fermentation products. Ninety-one genes with no presently defined function were also altered in expression, including seven of the most highly anaerobically induced genes, six of which we found to be directly regulated by FNR. Classification of the 297 genes into eight groups by k-means clustering analysis indicated that genes with common gene expression patterns also had a strong functional relationship, providing clues for studying the function of unknown genes in each group. Six of the eight groups showed regulation by FNR; while some expression groups represent genes that are simply activated or repressed by FNR, others, such as those encoding functions for chemotaxis and motility, showed a more complex pattern of regulation. A computer search for FNR DNA binding sites within predicted promoter regions identified 63 new sites for 54 genes. We suggest that E. coli MG1655 has a larger metabolic potential under anaerobic conditions than has been previously recognized. PMID- 15659693 TI - Specificity and polymorphism of the PlcR-PapR quorum-sensing system in the Bacillus cereus group. AB - The expression of extracellular virulence factors in various species of the Bacillus cereus group is controlled by the plcR and papR genes, which encode a transcriptional regulator and a cell-cell signaling peptide, respectively. A processed form of PapR, presumably a pentapeptide, specifically interacts with PlcR to facilitate its binding to its DNA targets. This activating mechanism is strain specific, with this specificity being determined by the first residue of the pentapeptide. We carried out in vivo complementation assays and compared the PlcR-PapR sequences of 29 strains from the B. cereus group. Our findings suggested that the fifth amino acid of the pentapeptide is also involved in the specificity of activation. We identified four classes of PlcR-PapR pairs, defining four distinct pherotypes in the B. cereus group. We used these findings to look at the evolution of the PlcR-PapR quorum-sensing system with regard to the phylogeny of the species forming the B. cereus group. PMID- 15659692 TI - NopB, a type III secreted protein of Rhizobium sp. strain NGR234, is associated with pilus-like surface appendages. AB - Rhizobium sp. strain NGR234 possesses a functional type three secretion system (TTSS), through which a number of proteins, called nodulation outer proteins (Nops), are delivered to the outside of the cell. A major constraint to the identification of Nops is their low abundance in the supernatants of NGR234 strains grown in culture. To overcome this limitation, a more sensitive proteomics-based strategy was developed. Secreted proteins from wild-type NGR234 were separated by two-dimensional gel electrophoresis, and the gel was compared to similar gels containing the proteins from a TTSS mutant (NGROmegarhcN). To identify the proteins, spots unique to the NGR234 gels were analyzed by matrix assisted laser desorption ionization-time of flight mass spectrometry and the data were compared to the sequence of the symbiotic plasmid of NGR234. A nonpolar mutant of one of these proteins was generated called NopB. NopB is required for Nop secretion but inhibits the interaction with Pachyrhizus tuberosus and augments nodulation of Tephrosia vogelii. Flavonoids and a functional TTSS are required for the formation of some surface appendages on NGR234. In situ immunogold labeling and isolation of these pili showed that they contain NopB. PMID- 15659694 TI - Site-directed mutagenesis analysis of amino acids critical for activity of the type I signal peptidase of the archaeon Methanococcus voltae. AB - Site-directed mutagenesis studies of the signal peptidase of the methanogenic archaeon Methanococcus voltae identified three conserved residues (Ser52, His122, and Asp148) critical for activity. The requirement for one conserved aspartic acid residue distinguishes the archaeal enzyme from both the Escherichia coli and yeast Sec11 enzymes. PMID- 15659695 TI - Characterization of phospholipase activity of the Pseudomonas aeruginosa type III cytotoxin, ExoU. AB - Recombinant ExoU (rExoU) and yeast extract were used to optimize an in vitro phospholipase assay as a basis for identifying the mechanism for enzyme activation and substrate specificity. Our results support a model in which a eukaryotic protein cofactor or complex facilitates the interaction of rExoU with phospholipid substrates. PMID- 15659696 TI - DNA methylation in lysogens of pathogenic Burkholderia spp. requires prophage induction and is restricted to excised phage DNA. AB - Burkholderia mallei-specific phage PhiE125 encodes DNA methyltransferases in both the lysogenic and replication modules within its genome. Characterization of DNA methylation in recombinant systems, specifically in PhiE125 lysogenic strains of B. mallei and Burkholderia thailandensis, revealed that, upon induction, cytosine methylation was targeted specifically to the phage episome but not the phage provirus or the host chromosome. PMID- 15659698 TI - Effects of 17-allylamino-17-demethoxygeldanamycin (17-AAG) on pediatric acute lymphoblastic leukemia (ALL) with respect to Bcr-Abl status and imatinib mesylate sensitivity. AB - As more and more effective targeted therapeutics have been developed to treat adults with cancer, it is of critical importance to devise appropriate in vitro experimental models to study their use in pediatric patients. Acute lymphoblastic leukemia (ALL) with Bcr-Abl translocation is one of the most difficult to treat and deadly diseases in children. The targeted kinase inhibitor imatinib mesylate has been shown to induce an initial response but resistance often develops. Recently, the geldanamycin family of antibiotics has been found to induce apoptosis in many malignant cells, including adult CML and AML. We describe experiments in which 17-allylamino-17-demethoxygeldanamycin (17-AAG) was evaluated in the context of Bcr-Abl and resistance to imatinib mesylate. Pediatric ALL cell lines with varying Bcr-Abl status and imatinib mesylate sensitivity were generated and their growth inhibition by 17-AAG was studied in vitro. Western blots were used to follow the changes in proteins that correlate with cell survival. Results show that apoptosis was induced in all lines with an increased 50% inhibitory concentration (IC50) for Bcr-Abl positive but imatinib mesylate-resistant cells. Addition of 17-AAG greatly increased imatinib sensitivity in vitro. A decrease in p53, survivin, Her2/neu, and WT1 was seen in cells that expressed these proteins. With some notable exceptions, when combined with 17-AAG, the IC50 of most of the common chemotherapeutic agents decreased. We describe an experimental approach to investigate the complex interaction between Bcr-Abl status, imatinib mesylate sensitivity, and 17-AAG in pediatric ALL. Information from such an approach will provide means to devise combined treatment approaches and to follow their effectiveness in vitro. PMID- 15659697 TI - Expression profiling of virulence and pathogenicity genes of Xanthomonas axonopodis pv. citri. AB - DNA macroarrays of 279 genes of Xanthomonas axonopodis pv. citri potentially associated with pathogenicity and virulence were used to compare the transcriptional alterations of this bacterium in response to two synthetic media. Data analysis indicated that 31 genes were up-regulated by synthetic medium XVM2, while only 7 genes were repressed. The results suggest that XVM2 could be used as an in vitro system to identify candidate genes involved in pathogenesis of X. axonopodis pv. citri. PMID- 15659699 TI - Bacteria and inflammatory cells in fetal membranes do not always cause preterm labor. AB - Intrauterine infection has been frequently linked with preterm labor before 30 wk of human pregnancy. Many different species of organisms have been detected, leading to the suggestion that infection-induced preterm labor is a generic inflammatory response to organisms rather than a specific response to a limited number of pathogens. The detection of organisms by microbiological culture is a laborious and unreliable process, so the aim of this study was to harness modern molecular techniques to detect organisms in tissues from human pregnancy. A DNA probe specific for conserved regions of bacterial 16S ribosomal RNA sequence was designed and labeled with fluorescein for fluorescence in situ hybridization. Organisms were detected in the great majority (>80%) of fetal membranes after prolonged premature rupture of the fetal membranes and after preterm labor, which was consistent with previous data. Organisms were also detected in fetal membranes after preterm delivery without labor and in term deliveries (with or without labour). Inflammatory cells were found frequently in the amnion or chorion of preterm fetal membranes but not in term tissues. Our primary finding is that fluorescence in situ hybridization is an appropriate method to detect organisms in human fetal membranes. In addition, our data show that bacteria may be present in fetal membranes without necessarily causing an inflammatory response, so the mere presence of bacteria may not be sufficient to cause preterm labor. PMID- 15659700 TI - Cerebral O2 supply thresholds for the preservation of electrocortical brain activity during hypotension in near-term-born lambs. AB - The fetal brain develops rapidly during the last trimester of pregnancy. Therefore, the brain of infants who are born preterm is vulnerable to changes in oxygen and nutrient supply in the neonatal period. The objective was to determine the effect of gestational age (GA) on the cerebral O2 supply threshold level for preservation of brain function during hypotension in near-term-born lambs. Lambs were delivered at 141 or 127 d of gestation. Hypotension was induced by stepwise withdrawal of blood. Mean arterial blood pressure (MABP) baseline levels were 63.2 (6.4) in 141-d and 54.4 (15.5) mm Hg in 127-d lambs. The MABP threshold below which MABP and blood flow in the left carotid artery were linearly related was 36.1 (13.1) mm Hg in 141-d lambs. In 127-d lambs, MABP and blood flow in the left carotid artery were linearly related over the whole range of recorded MABP values. Electrocortical brain activity (ECBA) was used as a measure of brain function. Thresholds of MABP for maintenance of ECBA were reached at, respectively, 31.6% (4.9%) of baseline in 141-d and 61.9% (13.0%) of baseline MABP in 127-d lambs. However, thresholds of cerebral O2 supply for maintenance of ECBA were similar in both GA groups. We conclude that thresholds of cerebral O2 supply for maintenance of brain cell function are independent of GA but are reached at higher MABP levels in 127-d than in 141-d lambs and therefore places the sick preterm infant easily at risk for ischemic cerebral injury. PMID- 15659701 TI - Omega-3 fatty acid supplementation prevents hepatic steatosis in a murine model of nonalcoholic fatty liver disease. AB - Prolonged use of total parenteral nutrition can lead to nonalcoholic fatty liver disease, ranging from hepatic steatosis to cirrhosis and liver failure. It has been demonstrated that omega-3 fatty acids are negative regulators of hepatic lipogenesis and that they can also modulate the inflammatory response in mice. Furthermore, they may attenuate hepatic steatosis even in leptin-deficient ob/ob mice. We hypothesized that omega-3 fatty acid supplementation may protect the liver against hepatic steatosis in a murine model of parenteral nutrition in which all animals develop steatosis and liver enzyme disturbances. For testing this hypothesis, groups of mice received a fat-free, high-carbohydrate liquid diet ad libitum for 19 d with enteral or i.v. supplementation of an omega-3 fatty acid emulsion or a standard i.v. lipid emulsion. Control mice received food alone or the fat-free, high-carbohydrate diet without lipid supplementation. Mice that received the fat-free, high-carbohydrate diet only or supplemented with a standard i.v. lipid emulsion developed severe liver damage as determined by histology and magnetic resonance spectroscopy as well as elevation of serum liver function tests. Animals that received an i.v. omega-3 fatty acid emulsion, however, showed only mild deposits of fat in the liver, whereas enteral omega-3 fatty acids prevented hepatic pathology and led to normalization of liver function tests. In conclusion, whereas standard i.v. lipid emulsions fail to improve dietary-induced steatotic injury to the liver, i.v. supplementation of omega-3 fatty acids partially and enteral supplementation completely protects the liver against such injury. PMID- 15659703 TI - Neonatal porcine Sertoli cells inhibit human natural antibody-mediated lysis. AB - Sertoli cells protect cotransplanted cells from allogeneic and xenogeneic rejection. Additionally, neonatal porcine Sertoli cells (NPSCs) survive long-term as xenografts in nonimmunosuppressed rodents. This has led to the hypothesis that NPSCs could be used to prevent cellular rejection in clinical transplantation, thereby eliminating the need for chronic immunosuppression. Prior to transplantation of NPSCs in humans it is necessary to determine whether they are also protected from humoral-mediated xenograft rejection. The presence of Gal alpha(1,3)Gal beta(1,4)GlcNAc-R (alphaGal epitope) as well as binding of human immunoglobulin G (IgG) and IgM to NPSCs was examined by immunocytochemical and fluorescence-activated cell sorter analysis. alphaGal was detected on 88.5% +/- 3.0% of NPSCs. Consistent with this, 71.7% +/- 1.0% and 65.4% +/- 5.2% of NPSCs were bound by IgG and IgM, respectively. When cultured NPSCs underwent an in vitro cytotoxicity assay by incubation with human AB serum plus complement, no increase in cellular lysis was observed, while controls--porcine aorta endothelial cells--were shown to contain > 60% dead cells. Finally, activation of the complement cascade was examined by immunohistochemistry. C3 and C4 were deposited on the surface of the NPSC membrane, indicating activation of complement. Although the complement cascade was activated, the membrane attack complex (MAC) was not formed. These data demonstrate that despite expression of alphaGal, binding of xenoreactive antibodies, and the activation of complement, NPSCs survive human antibody and complement-mediated lysis by preventing MAC formation. This suggests that NPSCs may be able to survive humoral-mediated rejection in a clinical situation. PMID- 15659702 TI - Follicle-stimulating hormone regulates both Sertoli cell and spermatogonial populations in the adult photoinhibited Djungarian hamster testis. AB - The hormones that regulate spermatogonial development are ill defined, in part due to lack of appropriate experimental models. The photoinhibited hamster model provides a rich source of spermatogonia, thus making it an ideal model to study their control. This study aimed to assess the effects of FSH, in the absence of testosterone, on the reinitiation of Sertoli cell and spermatogonial development in the photosensitive adult Djungarian hamster. Hamsters raised under long photoperiods (LD, 16L:8D) were exposed to short photoperiods (SD, 8L:16D) for 11 wk, leading to suppression of gonadotropins and regression of testicular function. Groups of 10 animals then received FSH alone or in combination with the antiandrogen, flutamide, for 7 days. Two control groups maintained either under long or short photoperiods were treated with vehicle. Sertoli and germ cell number were then determined using the optical disector (sic) stereological technique. The number of Sertoli cells, type A spermatogonia, type B spermatogonia/preleptotene spermatocytes, and leptotene/zygotene spermatocytes were suppressed in SD controls to 66%, 34%, 19%, and 10% (all P < 0.01) of long day control values, respectively. Later germ cell types were not detected. FSH treatment, with or without flutamide, increased Sertoli cell number (P < 0.01) to normal long-day values. Similarly, FSH treatment in the absence/presence of flutamide increased type A spermatogonia, type B spermatogonia/preleptotene spermatocytes, and leptotene/zygotene spermatocytes to approximately 85%, 69%, and 80% (all P < 0.01) of long-day controls, respectively. Our data demonstrate that the reinitiation of spermatogonial maturation in this model is dependent on FSH in the presence of an antiandrogen. Surprisingly, the adult Sertoli cell population in this model is also hormone dependent. This naturally occurring model provides a unique opportunity to understand the mechanisms (apoptotic and/or proliferative) by which FSH regulates Sertoli and germ cell development in the adult animal. PMID- 15659704 TI - Role of adenosine triphosphate, active mitochondria, and microtubules in the acquisition of developmental competence of parthenogenetically activated pig oocytes. AB - The purpose of this work was to determine the mechanisms regulating the acquisition of cytoplasmic maturation and embryonic developmental competence in pig oocytes. The presence or the absence of porcine follicular fluid (pff; 25% or 0%) in the maturation medium was used as a means to achieve complete nuclear maturation accompanied or not accompanied by cytoplasmic maturation. ATP content, active mitochondria relocation, and microtubule distribution were analyzed at different times during in vitro maturation (IVM). While nuclear maturation did not differ among the two groups, parthenogenetic embryonic development was significantly higher (41.5%) in the 25% pff group than in the 0% pff group (19.0%) with blastocysts that had a significantly higher number of blastomeres (76.1 +/- 6.3, and 47.2 +/- 6.5, respectively). Oocyte ATP content increased significantly during IVM, but at the end of maturation no significant differences were observed between high- and low-competence oocytes. An extensive relocation of mitochondria to the inner cytoplasm during IVM together with the formation of a well-developed mesh of cytoplasmic microtubules was observed only in the high competence oocyte group. However, no differences in the formation of microtubules associated with the meiotic spindles were observed between high- and low competence groups. We conclude that low developmental competence is associated with the lack of a microtubule cytoplasmic network, which prevents correct relocation of mitochondria and is likely to reflect a more generally altered compartmentalization of the ooplasm. This can be independent from the formation of the microtubule machinery required for the completion of chromosome disjunctions and does not affect the overall ATP content. PMID- 15659706 TI - Androgen regulation of stage-dependent cyclin D2 expression in Sertoli cells suggests a role in modulating androgen action on spermatogenesis. AB - Regulation of spermatogenesis involves stage-dependent androgen action on Sertoli cells, but the pathways involved are unclear. We assessed if cyclin D2 could play a role. In rats, Sertoli cell nuclear, stage-dependent immunoexpression of cyclin D2 switched on after Day 10 and persisted through Day 35, but disappeared by adulthood. However, ethane dimethane sulfonate (EDS)-induced testosterone withdrawal in adult rats for 6 days induced stage-dependent cyclin D2 immunoexpression in Sertoli cells, with highest expression at stages IX-XII and nondetectable at stages VI-VIII (opposite that for androgen receptor [AR] immunoexpression). In EDS-treated rats, a single injection of testosterone but not of estrogen reversed this change in 4 h, and testosterone administration from the time of EDS treatment prevented expression of cyclin D2 in Sertoli cells. The EDS-induced changes in cyclin D2 immunoexpression were matched by changes in expression of Ccnd2 (cyclin D2) mRNA in isolated stage-dissected tubules. Treatment of adult rats with flutamide induced stage-dependent cyclin D2 immunoexpression in Sertoli cells within 18 h, and confocal microscopy revealed that immunoexpression of AR and cyclin D2 were mutually exclusive within individual seminiferous tubules in these animals. Sertoli cell-selective ablation of the AR in mice using Cre/loxP technology also resulted in stage-dependent Sertoli cell cyclin D2 immunoexpression. Downstream from cyclin D2 action is retinoblastoma 1 (RB1), a tumor suppressor protein, immunoexpression of which paralleled stage-dependent AR expression in Sertoli cells; RB1 stage specificity disappeared after EDS treatment. These results point to a non-cell cycle role for cyclin D2 and RB1 in mature Sertoli cells in the stage-dependent mechanisms regulated by AR expression and androgen action. PMID- 15659705 TI - Regulated expression of ADAMTS family members in follicles and cumulus oocyte complexes: evidence for specific and redundant patterns during ovulation. AB - Protease cascades are essential for many biological events, including the LH induced process of ovulation. ADAMTS1 (a disintegrin and metalloproteinase with thrombospondin-like repeats-1) is expressed and hormonally regulated in the ovary by LH and the progesterone receptor. To determine whether other family members might be expressed and regulated in the rodent ovary, those closely related to ADAMTS1 (ADAMTS4 and ADAMTS5) were analyzed in the mouse ovary by reverse transcription-polymerase chain reaction as well as by Western blot, immunohistochemical, and immunocytochemical analyses using highly specific antibodies. Prior to ovulation, ADAMTS4 and ADAMTS5 were coexpressed in granulosa cells of most follicles, whereas ADAMTS5 was also present in granulosa cells of atretic follicles. Following ovulation, ADAMTS1 and ADAMTS4 (but not ADAMTS5) were expressed in multiple cell types, including those within the highly vascular ovulation cone that marks the site of follicle rupture, endothelial cells of newly forming corpora lutea, and cumulus cells within the ovulated cumulus cell oocyte complex (COC). Versican, a substrate for ADAMTS1 and ADAMTS4, colocalized with these proteases and hylauronan on the cumulus cell surface. To further characterize induction of these proteases and associated molecules, COCs and granulosa cells were isolated from preovulatory follicles and treated with FSH. In expanded COCs and differentiated granulosa cells, FSH induced expression of ADAMTS4 and versican message and protein, whereas increased levels of ADAMTS1 protein was observed in the media of granulosa cells where it was stabilized by heparin in this in vitro system. These studies provide the first evidence that ADAMTS1, ADAMTS4, and ADAMTS5 are expressed in spatiotemporal patterns that suggest distinct as well as some overlapping functions that relate to the broad expression pattern of versican in granulosa cells of small follicles, expanded COCs, and endothelial cells of the mouse ovary. PMID- 15659707 TI - Effect of cryopreservation on sea bass sperm proteins. AB - In the present study we used two-dimensional polyacrylamide gel electrophoresis (2-DE) and matrix-associated laser desorption/ionization time-of-flight (MALDI TOF) mass spectrometry to verify whether the protein expression of sea bass sperm was affected by the cryopreservation procedure. The protein profiles differed between fresh and frozen-thawed semen as revealed by visual inspection and by image analysis software. We identified 163 spots in fresh sperm; among these, 13 were significantly decreased and 8 were absent in two-dimensional gel obtained with cryopreserved sperm. Five of these spots were analyzed with MALDI-TOF, but only three showed a significant match in the databases used in bio-informatics analysis (PeptIdent, Mascot, and MS-Fit). In particular, spot 5 showed homology with a novel protein of zebrafish (similar to SKB1 of human and mouse), spot 13 showed homology with amphibian G1/S-specific cyclin E2, and spot 20 showed homology with the hypothetical protein DKFZp566A1524 of Brachidanio rerio. The present work shows that the use of the cryopreservation procedure causes the degradation of sperm proteins and among these, two could be at least partially responsible for the observed decrease in sperm motility duration and the lower hatching rate of eggs fertilized with cryopreserved sperm. PMID- 15659708 TI - Aged mouse oocytes fail to readjust intracellular adenosine triphosphates at fertilization. AB - Postovulatory aging of oocytes significantly affects embryonic development. Also, altered Ca2+ oscillation patterns can be observed in fertilized, aged mouse oocytes. Because Ca2+ oscillations depend on Ca2+ release and reuptake in the endoplasmic reticulum, and the latter relies on ATP availability, we simultaneously measured changes in intracellular ATP concentration ([ATP]i) and Ca2+ oscillations in fresh and aged mouse oocytes. We continuously assessed changes in [ATP]i from intracellular free Mg2+ concentration measured by fluorescent dye Magnesium Green (MgG) while intracellular Ca2+ concentration ([Ca2+]i) was monitored by Fura-PE3. At fertilization, MgG fluorescence was transiently increased concomitant with the first transient elevation in [Ca2+]i, indicating a relative decrease in [ATP]i. In fresh oocytes, it was quickly followed by a significant decrease below baseline, indicating a relative increase in [ATP]i. In contrast, in aged oocytes, such a decrease in MgG fluorescence was not observed. In a separate experiment, ATP content in fresh and aged oocytes was determined in vitro by the luciferin-luciferase assay. Intracellular ATP contents measured in vitro were comparable in unfertilized fresh and aged oocytes. Intracellular ATP content at 5 h after fertilization was increased in both oocytes, where fresh oocytes showed a significantly higher intracellular value than aged oocytes. These findings suggest that aged mouse oocytes fail to readjust the level of intracellular ATP at fertilization. Relative deficiencies of ATP at fertilization might lead to an altered Ca2+ oscillation pattern and poor developmental potency, which is commonly noted in aged oocytes. PMID- 15659709 TI - Uterine artery remodeling and reproductive performance are impaired in endothelial nitric oxide synthase-deficient mice. AB - The progressive rise in uterine blood flow during pregnancy is accompanied by outward hypertrophic remodeling of the uterine artery (UA). This process involves changes of the arterial smooth muscle cells and extracellular matrix. Acute increases in blood flow stimulate endothelial production of nitric oxide (NO). It remains to be established whether endothelial NO synthase (eNOS) is involved in pregnancy-related arterial remodeling. We tested the hypothesis that absence of eNOS results in a reduced remodeling capacity of the UA during pregnancy leading to a decline in neonatal outcome. UA of nonpregnant and pregnant wild-type (Nos3+/+) and eNOS-deficient (Nos3-/-) mice were collected and processed for standard morphometrical analyses. In addition, cross sections of UA were processed for cytological (smoothelin, smooth muscle alpha-actin) and proliferation (Ki-67) immunostaining. We compared the pregnancy-related changes longitudinally and, together with the data on pregnancy outcome, transversally by analysis of variance with Bonferroni correction. During pregnancy, the increases in radius and medial cross sectional area of Nos3-/- UA was significantly less than those of Nos3+/+ UA. Smooth muscle cell dedifferentiation and proliferation were impaired in gravid Nos3-/- mice as deduced from the lack of change in the expression of smoothelin and smooth muscle alpha-actin, and the reduced Ki-67 expression. Until 17 days of gestation, litter size did not differ between both genotypes, but at birth the number of viable newborn pups and their weights were smaller in Nos3-/- than in Nos3+/+ mice. We conclude that absence of eNOS adversely affects UA remodeling in pregnancy, which may explain the impaired pregnancy outcome observed in these mice. PMID- 15659710 TI - Cooperative expression of monocyte chemoattractant protein 1 within the bovine corpus luteum: evidence of immune cell-endothelial cell interactions in a coculture system. AB - Endothelial cells (EC) of the bovine corpus luteum (CL) are a known source of proinflammatory mediators, including monocyte chemoattractant protein 1 (CCL2) and endothelin 1 (EDN1). Here, a coculture system was devised to determine if immune cells and PGF 2alpha together affect CCL2 and EDN1 secretion by EC. Luteal EC were cultured either alone or together with peripheral blood mononuclear cells (PBMC), and treated without or with PGF 2alpha for 48 h (n = 6 experiments). Coculture of EC with PBMC increased CCL2 secretion an average of 5-fold higher compared with either cell type alone (P < 0.05). Basal secretion of EDN1 by EC was substantial (approximately 2 ng/ml), but was not affected by coculture with PBMC (P > 0.05). EC cocultured with concanavalin A-activated PBMC (ActPBMC) increased CCL2 secretion an average of 12-fold higher compared with controls (P < 0.05), but again, EDN1 secretion was unchanged (P > 0.05). Interestingly, PGF 2alpha did not alter either CCL2 or EDN1 secretion, regardless of culture conditions (P > 0.05). In a second series of experiments (n = 3 experiments), mixed luteal cells (MLC) were cultured alone or with PBMC as described above. Secretion of CCL2 and EDN1 was not affected by coculture or by PGF 2alpha (P > 0.05), but MLC produced less progesterone in the presence of ActPBMC (P < 0.05). Collectively, these results suggest that immune cells and EC can interact cooperatively to increase CCL2 secretion in the CL, but this interaction does not affect EDN1 secretion nor is it influenced by PGF 2alpha. Additionally, activated immune cells appear to produce a factor that impairs progesterone production by luteal steroidogenic cells. PMID- 15659711 TI - Cloning and characterization of the human soluble adenylyl cyclase. AB - We identified the human ortholog of soluble adenylyl cyclase (hsAC) in a locus linked to familial absorptive hypercalciuria and cloned it from a human cDNA library. hsAC transcripts were expressed in multiple tissues using RT-PCR and RNA blotting. RNA blot analysis revealed a predominant 5.1-kb band in a multiple human tissue blot, but three splice transcript variants were detected using RT PCR and confirmed by performing sequence analysis. Immunoblot analysis showed 190 and 80-kDa bands in multiple human cell lines from gut, renal, and bone origins in both cytosol and membrane fractions, including Caco-2 colorectal adenocarcinomas, HEK-293 cells, HOS cells, and primary human osteoblasts, as well as in vitro induced osteoclast-like cells. The specificity of the antiserum was verified by peptide blocking and reduction using sequence-specific small interfering RNA. Confocal immunofluorescence cytochemistry localized hsAC primarily in cytoplasm, but some labeling was observed in the nucleus and the plasma membrane. Cytoplasmic hsAC colocalized with microtubules but not with microfilaments. To test the function of hsAC, four constructs containing catalytic domains I and II (aa 1-802), catalytic domain II (aa 231-802), noncatalytic domain (aa 648-1,610), and full-length protein (aa 1-1,610) were expressed in Sf9 insect cells. Only catalytic domains I and II or full-length proteins showed adenylyl cyclase activity. Mg(2+), Mn(2+), and Ca(2+) all increased adenylyl cyclase activity in a dose-dependent manner. While hsAC had a minimal response to HCO(3)(-) in the absence of divalent cations, HCO(3)(-) robustly stimulated Mg(2+)-bound hsAC but inhibited Mn(2+)-bound hsAC in a dose dependent manner. In summary, hsAC is a divalent cation and HCO(3)(-) sensor, and its HCO(3)(-) sensitivity is modulated by divalent cations. PMID- 15659712 TI - Skeletal muscle contractile performance and ADP accumulation in adenylate kinase deficient mice. AB - The production of AMP by adenylate kinase (AK) and subsequent deamination by AMP deaminase limits ADP accumulation during conditions of high-energy demand in skeletal muscle. The goal of this study was to investigate the consequences of AK deficiency (-/-) on adenine nucleotide management and whole muscle function at high-energy demands. To do this, we examined isometric tetanic contractile performance of the gastrocnemius-plantaris-soleus (GPS) muscle group in situ in AK1(-/-) mice and wild-type (WT) controls over a range of contraction frequencies (30-120 tetani/min). We found that AK1(-/-) muscle exhibited a diminished inosine 5'-monophosphate formation rate (14% of WT) and an inordinate accumulation of ADP ( approximately 1.5 mM) at the highest energy demands, compared with WT controls. AK-deficient muscle exhibited similar initial contractile performance (521 +/- 9 and 521 +/- 10 g tension in WT and AK1(-/-) muscle, respectively), followed by a significant slowing of relaxation kinetics at the highest energy demands relative to WT controls. This is consistent with a depressed capacity to sequester calcium in the presence of high ADP. However, the overall pattern of fatigue in AK1(-/-) mice was similar to WT control muscle. Our findings directly demonstrate the importance of AMP formation and subsequent deamination in limiting ADP accumulation. Whole muscle contractile performance was, however, remarkably tolerant of ADP accumulation markedly in excess of what normally occurs in skeletal muscle. PMID- 15659713 TI - A rapid and efficient PCR-based mutagenesis method applicable to cell physiology study. AB - PCR-based mutagenesis is a cornerstone of molecular biology and protein engineering studies. Herein we describe a rapid and highly efficient mutagenesis method using type IIs restriction enzymes. A template gene is amplified into two separate PCR fragments using two pairs of anchor and mutagenic primers. Mutated sequences are located near the recognition site of a type IIs restriction enzyme. After digestion of two fragments with a type IIs enzyme, exposed cohesive ends that are complementary to each other are then ligated together to generate a mutated gene. We applied this method to introduce multiple site-directed mutations in EGFP and Bcl-2 family genes and observed perfect mutagenesis efficiency at the desired sites. This efficient and cost-effective mutagenesis method can be applied to a wide variety of structural and functional studies in cell physiology. PMID- 15659714 TI - Relative contribution of chloride channels and transporters to regulatory volume decrease in human glioma cells. AB - Primary brain tumors (gliomas) often present with peritumoral edema. Their ability to thrive in this osmotically altered environment prompted us to examine volume regulation in human glioma cells, specifically the relative contribution of Cl(-) channels and transporters to this process. After a hyposmotic challenge, cultured astrocytes, D54-MG glioma cells, and glioma cells from human patient biopsies exhibited a regulatory volume decrease (RVD). Although astrocytes were not able to completely reestablish their original prechallenge volumes, glioma cells exhibited complete volume recovery, sometimes recovering to a volume smaller than their original volumes (V(Post-RVD) < V(baseline)). In glioma cells, RVD was largely inhibited by treatment with a combination of Cl(-) channel inhibitors, 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) and Cd(2+) (V(Post RVD) > 1.4*V(baseline)). Volume regulation was also attenuated to a lesser degree by the addition of R-(+)-[(2-n-butyl-6,7-dichloro-2-cyclopentyl-2,3-dihydro-1-oxo 1H-inden-5-yl)oxy]acetic acid (DIOA), a known K(+)-Cl(-) cotransporter (KCC) inhibitor. To dissect the relative contribution of channels vs. transporters in RVD, we took advantage of the comparatively high temperature dependence of transport processes vs. channel-mediated diffusion. Cooling D54-MG glioma cells to 15 degrees C resulted in a loss of DIOA-sensitive volume regulation. Moreover, at 15 degrees C, the channel blockers NPPB + Cd(2+) completely inhibited RVD and cells behaved like perfect osmometers. The calculated osmolyte flux during RVD under these experimental conditions suggests that the relative contribution of Cl(-) channels vs. transporters to this process is approximately 60-70% and approximately 30-40%, respectively. Finally, we identified several candidate proteins that may be involved in RVD, including the Cl(-) channels ClC-2, ClC-3, ClC-5, ClC-6, and ClC-7 and the transporters KCC1 and KCC3a. PMID- 15659715 TI - Dual effects of n-alcohols on fluid secretion from guinea pig pancreatic ducts. AB - Ethanol strongly augments secretin-stimulated, but not acetylcholine (ACh) stimulated, fluid secretion from pancreatic duct cells. To understand its mechanism of action, we examined the effect of short-chain n-alcohols on fluid secretion and intracellular Ca(2+) concentration ([Ca(2+)](i)) in guinea pig pancreatic ducts. Fluid secretion was measured by monitoring the luminal volume of isolated interlobular ducts. [Ca(2+)](i) was estimated using fura-2 microfluorometry. Methanol and ethanol at 0.3-10 mM concentrations significantly augmented fluid secretion and induced a transient elevation of [Ca(2+)](i) in secretin- or dibutyryl adenosine 3',5'-cyclic monophosphate (DBcAMP)-stimulated ducts. However, they failed to affect fluid secretion and [Ca(2+)](i) in unstimulated and ACh-stimulated ducts. In contrast, propanol and butanol at 0.3 10 mM concentrations significantly reduced fluid secretion and decreased [Ca(2+)](i) in unstimulated ducts and in ducts stimulated with secretin, DBcAMP, or ACh. Both stimulatory and inhibitory effects of n-alcohols completely disappeared after their removal from the perfusate. Propanol and butanol inhibited the plateau phase, but not the initial peak, of [Ca(2+)](i) response to ACh as well as the [Ca(2+)](i) elevation induced by thapsigargin, suggesting that they inhibit Ca(2+) influx. Removal of extracellular Ca(2+) reduced [Ca(2+)](i) in duct cells and completely abolished secretin-stimulated fluid secretion. In conclusion, there is a distinct cutoff point between ethanol (C2) and propanol (C3) in their effects on fluid secretion and [Ca(2+)](i) in duct cells. Short chain n-alcohols appear to affect pancreatic ductal fluid secretion by activating or inhibiting the plasma membrane Ca(2+) channel. PMID- 15659716 TI - CaM kinase II and phospholamban contribute to caffeine-induced relaxation of murine gastric fundus smooth muscle. AB - Caffeine has been shown to increase the Ca(2+) release frequency (Ca(2+) sparks) from the sarcoplasmic reticulum (SR) through ryanodine-sensitive stores and relax gastric fundus smooth muscle. Increased Ca(2+) store refilling increases the frequency of Ca(2+) release events and store refilling is enhanced by CaM kinase II (CaMKII) phosphorylation of phospholamban (PLB). These findings suggest that transient, localized Ca(2+) release events from the SR may activate CaMKII and contribute to relaxation by enhancing store refilling due to PLB Thr17 phosphorylation. To investigate this possibility, we examined the effects of caffeine on CaMKII, muscle tone, and PLB phosphorylation in murine gastric fundus smooth muscle. Caffeine (1 mM) hyperpolarized and relaxed murine gastric fundus smooth muscle and activated CaMKII. Ryanodine, tetracaine, or cyclopiazonic acid each prevented CaMKII activation and significantly inhibited caffeine-induced relaxation. The large-conductance Ca(2+)-activated K(+) channel blocker iberiotoxin, but not apamin, partially inhibited caffeine-induced relaxation. Caffeine-induced CaMKII activation increased PLB Thr17, but not PLB Ser16 phosphorylation. 3-Isobutyl-1-methylxanthine increased PLB Ser16 phosphorylation, but not PLB Thr17 phosphorylation. The CaMKII inhibitor KN-93 inhibited caffeine induced relaxation and PLB Thr17 phosphorylation. These results show that caffeine-induced CaMKII activation and PLB phosphorylation play a role in the relaxation of gastric fundus smooth muscles. PMID- 15659718 TI - Pay at the pump. PMID- 15659717 TI - Sphingosine 1-phosphate protects mouse extensor digitorum longus skeletal muscle during fatigue. AB - Sphingomyelin derivatives exert various second messenger actions in numerous tissues. Sphingosine (SPH) and sphingosine 1-phosphate (S1P) are two major sphingomyelin derivatives present at high levels in blood. The aim of the present work was to investigate whether S1P and SPH exert relevant actions in mouse skeletal muscle contractility and fatigue. Exogenous S1P and SPH administration caused a significant reduction of tension decline during fatigue of extensor digitorum longus muscle. Final tension after the fatiguing protocol was 40% higher than in untreated muscle. Interestingly, N,N-dimethylsphingosine, an inhibitor of SPH kinase (SK), abolished the effect of supplemented SPH but not that of S1P, suggesting that SPH acts through its conversion to S1P. Moreover, SPH was not effective in Ca(2+)-free solutions, in agreement with the hypothesis that SPH action is dependent on its conversion to S1P by the Ca(2+)-requiring enzyme SK. In contrast to SPH, S1P produced its positive effects on fatigue in Ca(2+)-free conditions, indicating that S1P action does not require Ca(2+) entry and most likely is receptor mediated. The effects of S1P could be ascribed in part to its ability to prevent the reduction (-20 mV) of action potential amplitude caused by fatigue. In conclusion, these results indicate that extracellular S1P has protective effects during the development of muscle fatigue and that the extracellular conversion of SPH to S1P may represent a rheostat mechanism to protect skeletal muscle from possible cytotoxic actions of SPH. PMID- 15659719 TI - Diabetes and stem cell researchers turn to the lowly spleen. AB - The spleen gets no respect. Long seen as superfluous, the adult spleen of the mouse has recently been shown to hold stem cells that, in diabetic mice or genetically altered mice that lack a pancreas, effectively regenerate insulin producing islet cells of the pancreas. Stem cells of the spleen express Hox11, a highly conserved transcription factor that plays a major role in the development of organs in vertebrate and invertebrate embryos. Hox11 and other members of the Hox family of genes may give stem cells of the spleen the capacity to mature into cell types other than islet cells, including neurons and bone cells. Multilineage splenic stem cells may trace to the embryogenesis and possible persistence into adulthood of a fetal stem cell region called the aorta-gonad-mesonephros (AGM). This Perspective calls for reappraisal of the lowly spleen for treating diabetes and other diseases of aging. PMID- 15659720 TI - Beyond the Purple Heart--continuity of care for the wounded in Iraq. PMID- 15659721 TI - Medicare coverage of ICDs. PMID- 15659722 TI - Amiodarone or an implantable cardioverter-defibrillator for congestive heart failure. AB - BACKGROUND: Sudden death from cardiac causes remains a leading cause of death among patients with congestive heart failure (CHF). Treatment with amiodarone or an implantable cardioverter-defibrillator (ICD) has been proposed to improve the prognosis in such patients. METHODS: We randomly assigned 2521 patients with New York Heart Association (NYHA) class II or III CHF and a left ventricular ejection fraction (LVEF) of 35 percent or less to conventional therapy for CHF plus placebo (847 patients), conventional therapy plus amiodarone (845 patients), or conventional therapy plus a conservatively programmed, shock-only, single-lead ICD (829 patients). Placebo and amiodarone were administered in a double-blind fashion. The primary end point was death from any cause. RESULTS: The median LVEF in patients was 25 percent; 70 percent were in NYHA class II, and 30 percent were in class III CHF. The cause of CHF was ischemic in 52 percent and nonischemic in 48 percent. The median follow-up was 45.5 months. There were 244 deaths (29 percent) in the placebo group, 240 (28 percent) in the amiodarone group, and 182 (22 percent) in the ICD group. As compared with placebo, amiodarone was associated with a similar risk of death (hazard ratio, 1.06; 97.5 percent confidence interval, 0.86 to 1.30; P=0.53) and ICD therapy was associated with a decreased risk of death of 23 percent (0.77; 97.5 percent confidence interval, 0.62 to 0.96; P=0.007) and an absolute decrease in mortality of 7.2 percentage points after five years in the overall population. Results did not vary according to either ischemic or nonischemic causes of CHF, but they did vary according to the NYHA class. CONCLUSIONS: In patients with NYHA class II or III CHF and LVEF of 35 percent or less, amiodarone has no favorable effect on survival, whereas single-lead, shock-only ICD therapy reduces overall mortality by 23 percent. PMID- 15659723 TI - Clopidogrel versus aspirin and esomeprazole to prevent recurrent ulcer bleeding. AB - BACKGROUND: Concurrent therapy with a proton-pump inhibitor is a standard treatment for patients receiving aspirin who are at risk for ulcer. Current U.S. guidelines also recommend clopidrogel for patients who have major gastrointestinal intolerance of aspirin. We compared clopidogrel with aspirin plus esomeprazole for the prevention of recurrent bleeding from ulcers in high risk patients. METHODS: We studied patients who took aspirin to prevent vascular diseases and who presented with ulcer bleeding. After the ulcers had healed, we randomly assigned patients who were negative for Helicobacter pylori to receive either 75 mg of clopidogrel daily plus esomeprazole placebo twice daily or 80 mg of aspirin daily plus 20 mg of esomeprazole twice daily for 12 months. The end point was recurrent ulcer bleeding. RESULTS: We enrolled 320 patients (161 patients assigned to receive clopidogrel and 159 to receive aspirin plus esomeprazole). Recurrent ulcer bleeding occurred in 13 patients receiving clopidogrel and 1 receiving aspirin plus esomeprazole. The cumulative incidence of recurrent bleeding during the 12-month period was 8.6 percent (95 percent confidence interval, 4.1 to 13.1 percent) among patients who received clopidogrel and 0.7 percent (95 percent confidence interval, 0 to 2.0 percent) among those who received aspirin plus esomeprazole (difference, 7.9 percentage points; 95 percent confidence interval for the difference, 3.4 to 12.4; P=0.001). CONCLUSIONS: Among patients with a history of aspirin-induced ulcer bleeding whose ulcers had healed before they received the study treatment, aspirin plus esomeprazole was superior to clopidogrel in the prevention of recurrent ulcer bleeding. Our finding does not support the current recommendation that patients with major gastrointestinal intolerance of aspirin be given clopidogrel. PMID- 15659724 TI - Effects of moderate alcohol consumption on cognitive function in women. AB - BACKGROUND: The adverse effects of excess alcohol intake on cognitive function are well established, but the effect of moderate consumption is uncertain. METHODS: Between 1995 and 2001, we evaluated cognitive function in 12,480 participants in the Nurses' Health Study who were 70 to 81 years old, with follow up assessments in 11,102 two years later. The level of alcohol consumption was ascertained regularly beginning in 1980. We calculated multivariate-adjusted mean cognitive scores and multivariate-adjusted risks of cognitive impairment (defined as the lowest 10 percent of the scores) and a substantial decline in cognitive function over time (defined as a change that was in the worst 10 percent of the distribution of the decline). We also stratified analyses according to the apolipoprotein E genotype in a subgroup of women. RESULTS: After multivariate adjustment, moderate drinkers (those who consumed less than 15.0 g of alcohol per day [about one drink]) had better mean cognitive scores than nondrinkers. Among moderate drinkers, as compared with nondrinkers, the relative risk of impairment was 0.77 on our test of general cognition (95 percent confidence interval, 0.67 to 0.88) and 0.81 on the basis of a global cognitive score combining the results of all tests (95 percent confidence interval, 0.70 to 0.93). The results for cognitive decline were similar; for example, on our test of general cognition, the relative risk of a substantial decline in performance over a two-year period was 0.85 (95 percent confidence interval, 0.74 to 0.98) among moderate drinkers, as compared with nondrinkers. There were no significant associations between higher levels of drinking (15.0 to 30.0 g per day) and the risk of cognitive impairment or decline. There were no significant differences in risks according to the beverage (e.g., wine or beer) and no interaction with the apolipoprotein E genotype. CONCLUSIONS: Our data suggest that in women, up to one drink per day does not impair cognitive function and may actually decrease the risk of cognitive decline. PMID- 15659727 TI - Images in clinical medicine. A medical mystery. PMID- 15659726 TI - Shattuck Lecture: health care in the 21st century. PMID- 15659725 TI - Cytoplasmic nucleophosmin in acute myelogenous leukemia with a normal karyotype. AB - BACKGROUND: Nucleophosmin (NPM), a nucleocytoplasmic shuttling protein with prominent nucleolar localization, regulates the ARF-p53 tumor-suppressor pathway. Translocations involving the NPM gene cause cytoplasmic dislocation of the NPM protein. METHODS: We used immunohistochemical methods to study the subcellular localization of NPM in bone marrow-biopsy specimens from 591 patients with primary acute myelogenous leukemia (AML). We then correlated the presence of cytoplasmic NPM with clinical and biologic features of the disease. RESULTS: Cytoplasmic NPM was detected in 208 (35.2 percent) of the 591 specimens from patients with primary AML but not in 135 secondary AML specimens or in 980 hematopoietic or extrahematopoietic neoplasms other than AML. It was associated with a wide spectrum of morphologic subtypes of the disease, a normal karyotype, and responsiveness to induction chemotherapy, but not with recurrent genetic abnormalities. There was a high frequency of FLT3 internal tandem duplications and absence of CD34 and CD133 in AML specimens with a normal karyotype and cytoplasmic dislocation of NPM, but not in those in which the protein was restricted to the nucleus. AML specimens with cytoplasmic NPM carried mutations of the NPM gene that were predicted to alter the protein at its C-terminal; this mutant gene caused cytoplasmic localization of NPM in transfected cells. CONCLUSIONS: Cytoplasmic NPM is a characteristic feature of a large subgroup of patients with AML who have a normal karyotype, NPM gene mutations, and responsiveness to induction chemotherapy. PMID- 15659728 TI - Case records of the Massachusetts General Hospital. Weekly clinicopathological exercises. Case 2-2005. A 39-year-old woman with headache, stiff neck, and photophobia. PMID- 15659729 TI - Prophylactic defibrillator implantation--toward an evidence-based approach. PMID- 15659730 TI - Reducing the risks of gastrointestinal bleeding with antiplatelet therapies. PMID- 15659731 TI - Alcohol consumption and cognition. PMID- 15659732 TI - NPM mutations in acute myelogenous leukemia. PMID- 15659733 TI - Today's FDA. PMID- 15659734 TI - Topical microbicides become topical. PMID- 15659735 TI - Oral erythromycin and the risk of sudden death. PMID- 15659736 TI - Birth weight and breast cancer. PMID- 15659737 TI - Autoimmune lymphoproliferative syndrome and perforin. PMID- 15659738 TI - Diastolic heart failure. PMID- 15659739 TI - Federal funding for stem-cell research. PMID- 15659740 TI - Treatment of pulmonary arterial hypertension. PMID- 15659741 TI - Case 29-2004: a woman with acute onset of chest pain and fever. PMID- 15659742 TI - Adefovir dipivoxil for chemotherapy-induced activation of hepatitis B virus infection. PMID- 15659743 TI - Type-specific reactivity of anti-glycoprotein G antibodies from herpes simplex virus-infected individuals is maintained by single or dual type-specific residues. AB - Glycoprotein G-1 (gG-1) of herpes simplex virus type 1 (HSV-1) and gG-2 of HSV-2 are the only known HSV proteins that induce type-specific human antibody responses. Recently, it was shown that purified human anti-gG-1 and anti-gG-2 antibodies presented a type-specific reactivity to immunogenic stretches with high similarity between gG-1 and gG-2. In this study, the molecular basis for this type-specific recognition was investigated employing synthetic peptides covering the indicated regions, including substitutions of the type-specific residues. The results revealed that single or dual type-specific residues localized within regions of high similarity could induce significant structural differences, explaining the type-specific recognition of the human antibody response to the gG proteins. PMID- 15659744 TI - The cytoplasmic tail of herpes simplex virus envelope glycoprotein D binds to the tegument protein VP22 and to capsids. AB - Herpes simplex virus (HSV) capsids assemble, mature and package their viral genome in the nucleoplasm. They then exit the nucleus into the cytoplasm, where they acquire their final tegument and envelope. The molecular mechanism of cytoplasmic envelopment is unclear, but evidence suggests that the viral glycoprotein tails play an important role in the recruitment of tegument and capsids at the final envelopment site. However, due to redundancy in protein protein interactions among the viral glycoproteins, genetic analysis of the role of individual glycoproteins in assembly has been difficult. To overcome this problem, a glutathione S-transferase fusion protein-binding assay was used in this study to test the interaction between the cytoplasmic tail of one specific viral glycoprotein, gD, and tegument proteins. The study demonstrated that the 38 kDa tegument protein VP22 bound specifically to the gD tail. This association was dependent on arginine and lysine residues at positions 5 and 6 in the gD tail. In addition, HSV-1 capsids bound the gD tail and exhibited a similar sequence dependence. It is concluded that VP22 may serve as a linker protein, mediating the interaction of the HSV capsid with gD. PMID- 15659745 TI - The English strain of rat cytomegalovirus (CMV) contains a novel captured CD200 (vOX2) gene and a spliced CC chemokine upstream from the major immediate-early region: further evidence for a separate evolutionary lineage from that of rat CMV Maastricht. AB - Sequence data for eight genes, together with time-course Northern blotting and 3' and 5'-RACE (rapid amplification of cDNA ends) analysis for some mRNAs from a 12 kb region upstream from the major immediate-early (MIE) genes of the English isolate of rat cytomegalovirus (RCMV), are presented. The results identified important differences compared to both murine cytomegalovirus (MCMV) and the Maastricht isolate of RCMV. A striking finding is the presence of a highly conserved, rightwards-oriented homologue of the rat cellular CD200 (OX2) gene immediately to the right of the MIE region, which replaces either the leftwards oriented AAV REP gene of RCMV (Maastricht) or the upstream spliced portions of the immediate-early 2 gene (ie2) in MCMV. From the presence of other homologues of MCMV- and RCMV-specific genes, such as the beta-chemokine MCK-2, SGG1 and an Fcgamma receptor gene, as reported here, the basic architecture of the MIE region (reported previously) and the level of IE2 and DNA polymerase (POL) protein conservation in phylogenetic analyses, it is clear that the English strain of RCMV is also a member of the genus Muromegalovirus, but is a beta-herpesvirus species that is very distinct from both MCMV and RCMV (Maastricht). Both the lack of a CD200 homologue in the other two rodent viruses and the depth of sequence divergence of the rodent CMV IE2 and POL proteins suggest that these three viruses have evolved as separate species in the genus Muromegalovirus since very early in the host rodent lineage. PMID- 15659746 TI - Dendritic-cell infection by human cytomegalovirus is restricted to strains carrying functional UL131-128 genes and mediates efficient viral antigen presentation to CD8+ T cells. AB - Human cytomegalovirus (HCMV) genetic determinants of endothelial-cell tropism and virus transfer to leukocytes (both polymorphonuclear and monocyte) have been recently identified in the UL131-128 genes. Here it is documented that the same genetic determinants of HCMV are responsible for monocyte-derived dendritic-cell (DC) tropism, i.e. all endotheliotropic and leukotropic strains of HCMV are also DC-tropic (or dendrotropic). In fact, all recent clinical HCMV isolates and deletion mutants sparing the UL131-128 locus as well as the endotheliotropic revertants AD169 and Towne were able to productively infect DC following co culture with infected endothelial cells. On the contrary, the same clinical isolates extensively propagated in human fibroblasts, the UL131-128 deletion mutants and the reference laboratory strains were not. Peak extracellular virus titres in DC were reached 4-7 days post-infection (p.i.). Viral proteins pp65 and p72 were detected 1-3 h p.i., involving the great majority of DC 24 h p.i., while gB was abundantly detected 96 h p.i., when a cytopathic effect first appeared. Infection of DC with cell-free virus released into the medium could only be achieved with HCMV strains extensively adapted to growth in endothelial cells, reaching the peak titres 10 days p.i. DC infected for 24 h with cell-free virus and incubated for 16 h with autologous peripheral blood mononuclear cells were found to act as a potent stimulator of both HCMV-specific CD4+- and CD8+-mediated immune responses, as determined by cytokine flow cytometry. DC incubated with inactivated crude whole viral antigen preparations were only capable of eliciting a significant CD4+-mediated immune response. PMID- 15659747 TI - NF-kappaB activation can mediate inhibition of human cytomegalovirus replication. AB - The activation of NF-kappaB has long been considered a positive factor for human cytomegalovirus (HCMV) replication. The HCMV immediate-early promoter, the initial transcriptional element in the HCMV replication cycle, is activated by the transcription factor NF-kappaB, and several HCMV gene products have been demonstrated to activate this transcription factor. However, the role of NF kappaB in the full replication cycle of the virus has not been carefully examined. A series of experiments that demonstrate an important inhibitory role of NF-kappaB for HCMV replication in fibroblasts is presented here. Using both genetic and pharmaceutical methods, it was shown that blocking NF-kappaB activation in cell culture does not inhibit HCMV replication, but rather leads to a modest increase in replication. Two cytokines inhibitory for HCMV, tumour necrosis factor-alpha and interferon-gamma, no longer inhibit HCMV when NF-kappaB activation is blocked. Furthermore, forced expression of the NF-kappaB activating IkappaB kinase beta (IKKbeta), but not a kinase inactive mutant, also inhibits HCMV replication. In addition, it was shown that NF-kappaB signalling is essential for the production of an anti-viral factor in the supernatant of HCMV infected fibroblasts, and identified interferon-beta as this factor. Thus, the role of NF-kappaB in fibroblasts is to activate a host defence against HCMV. PMID- 15659748 TI - The human cytomegalovirus UL78 gene is highly conserved among clinical isolates, but is dispensable for replication in fibroblasts and a renal artery organ culture system. AB - The human cytomegalovirus (HCMV) UL78 ORF is considered to encode a seven transmembrane receptor. However, neither the gene nor the UL78 protein has been characterized so far. The objective of this study was to investigate the UL78 gene and to clarify whether it is essential for replication. UL78 transcription was activated early after infection, was inhibited by cycloheximide but not by phosphonoacetic acid, and resulted in a 1.7 kb mRNA. Later in the replication cycle, a second mRNA of 4 kb evolved, comprising the UL77 and UL78 ORFs. The 5' end of the UL78 mRNA initiated 48 bp upstream of the translation start and the polyadenylated tail started 268 bp downstream of the UL78 translation stop codon within the UL79 ORF. By using bacterial artificial chromosome technology, a recombinant HCMV lacking most of the UL78 coding region was constructed. Successful reconstitution of the UL78-deficient virus proved that the gene was not essential for virus replication in fibroblasts. The deletion also did not reduce virus replication in ex vivo-cultured sections of human renal arteries. Analysis of viral proteins at different stages of the replication cycle confirmed these results. Among clinical HCMV isolates, the predicted UL78 protein was highly conserved. However, an accumulation of different single mutations could be found in the N-terminal region and at the very end of the C terminus. Due to the absence of an in vivo HCMV model, the role of UL78 in the pathogenesis of HCMV infection in humans remains unclear. PMID- 15659749 TI - On phylogenetic relationships among major lineages of the Gammaherpesvirinae. AB - Phylogenetic relationships within the subfamily Gammaherpesvirinae of the family Herpesviridae were investigated for three species in the genus Lymphocryptovirus (or gamma1 group) and nine in the genus Rhadinovirus (or gamma2 group). Alignments of amino acid sequences from up to 28 genes were used to derive trees by maximum-likelihood and Bayesian Monte Carlo Markov chain methods. Two problem areas were identified involving an unresolvable multifurcation for a clade within the gamma2 group, and a high divergence for Murid herpesvirus 4 (MHV4). A robust final tree was obtained, which was valid for genes from across the virus genomes and was rooted by reference to previous analyses of the whole family Herpesviridae. This tree comprised four major lineages: the gamma1 group of primate viruses; a clade of artiodactyl gamma2 viruses; a clade of perissodactyl gamma2 viruses; and a clade of gamma2 viruses with a multifurcation at its base and containing Old World and New World primate viruses, Bovine herpesvirus 4 and MHV4. Developing previous work it was proposed, on the basis of similarities between the gammaherpesvirus tree and the tree of corresponding mammalian hosts, that the first three of these major viral lineages arose in a coevolutionary manner with host lineages, while the fourth had its origin in an ancient interspecies transfer. Transfer of dates from mammalian palaeontology then allowed estimation of dates for nodes in the gammaherpesvirus tree. PMID- 15659750 TI - Reactivation of Epstein-Barr virus can be triggered by an Rta protein mutated at the nuclear localization signal. AB - Rta, an immediate-early protein of Epstein-Barr virus (EBV), is a transcriptional activator that induces lytic gene expression and triggers virus reactivation. Being located predominantly in the nucleus, Rta can exert its transactivation function through either direct DNA binding or certain indirect mechanisms mediated by cellular signalling and other transcriptional factors. This study examined whether the subcellular localization of Rta was critical for the induction of target genes. First, 410KRKK413 was identified as a nuclear localization signal (NLS) of Rta. An Rta mutant with the NLS converted to 410AAAA413 showed cytoplasmic localization and failed to activate the promoter of BGLF5. Interestingly, ectopic expression of the Rta mutant still disrupted EBV latency in an epithelial cell line. Reporter gene assays revealed that the NLS mutated Rta retained the ability to activate two lytic promoters, Zp and Rp, at a considerable level. Thus, the cytoplasmic Rta mutant could induce expression of endogenous Zta and Rta, triggering reactivation of EBV. PMID- 15659751 TI - Immunostimulatory potential of hepatitis B nucleocapsid preparations: lipopolysaccharide contamination should not be overlooked. AB - The nucleocapsid of hepatitis B virus (HBV) allows insertions of heterologous peptides and even complete proteins. Because of its outstanding capacity to induce B-cell, T-helper and cytotoxic T-cell responses, this structure is considered to be an important instrument for future vaccine development. Most of the evidence for the unique immunogenic qualities of nucleocapsids has been generated in mice, which are not natural hosts of HBV. Moreover, most nucleocapsid preparations used in these studies were produced in a recombinant manner in Escherichia coli. Such preparations have been shown to contain lipopolysaccharide (LPS). Not unexpectedly, it is shown here that contaminating LPS, rather than the nucleocapsid structure itself, is responsible for the activation of human antigen-presenting cells. Careful examination of the literature dealing with the immunogenicity of HBV nucleocapsids suggests that the possible presence of LPS has been largely ignored or underestimated in several studies. This raises doubts on some of the underlying mechanisms that have been proposed to explain the unique immunogenicity of the HBV nucleocapsid. PMID- 15659752 TI - Frequent infection of Hylobates pileatus (pileated gibbon) with species associated variants of hepatitis B virus in Cambodia. AB - As well as being distributed widely in human populations, hepatitis B virus (HBV) infections occur frequently in chimpanzee, gibbon and other ape populations in sub-Saharan Africa and South-East Asia. To investigate the frequency and genetic relationships of HBV infecting gibbons in Cambodia, pileated gibbons (Hylobates pileatus) that were originally wild-caught were screened for surface antigen. Twelve of 26 (46 %) were positive, of which 11 were positive for HBV DNA. Phylogenetic analysis of complete genome sequences revealed two distinct genetic groups in the gibbon/orangutan clade. Three were similar to previously described variants infecting H. pileatus in Thailand and eight formed a distinct clade, potentially representing distinct strains of HBV circulating in geographically separated populations in South-East Asia. Because of the ability of HBV to cross species barriers, large reservoirs of infection in gibbons may hamper ongoing attempts at permanent eradication of HBV infection from human populations in South-East Asia through immunization. PMID- 15659753 TI - Predictive value of anti-cell and anti-human immunodeficiency virus (HIV) humoral responses in HIV-1-exposed seronegative cohorts of European and Asian origin. AB - Unconventional immune responses have been demonstrated in individuals who, despite repeated exposure to human immunodeficiency virus (HIV) infection, remain seronegative. As environmental exposure to pathogens and genetic background may modulate immune responses differentially, one Italian and two Asian populations of HIV-1-exposed seronegative individuals were studied. In serum samples from each group, IgG to CCR5, IgG to CD4 and IgA to gp41 were measured, which were previously described as markers of unconventional immunity in HIV-exposed seronegative Caucasians. Given the importance of conformational epitopes in virus cell interactions, IgG to CD4-gp120 complex was also measured. It was found that markers of HIV exposure were present in all populations studied. HIV-specific humoral responses (IgA to gp41 and IgG to CD4-gp120 complex) were extremely significant predictors of HIV exposure (P<0.0001 in both cases), whereas the predictive values of anti-cell antibodies (anti-CCR5 and anti-CD4) varied between populations. Evidence is provided for the correlation of these differences with route of exposure to HIV and level of natural antibodies to cross-reactive microbial antigens. In conclusion, exposed seronegative individuals of ethnically different origins display similar signs of HIV-dependent unconventional immunity. A specific relevance must be attributed to different innate and acquired factors. PMID- 15659754 TI - Enhanced immunogenicity using an alphavirus replicon DNA vaccine against human immunodeficiency virus type 1. AB - With the human immunodeficiency virus type 1 (HIV-1) epidemic expanding at increasing speed, development of a safe and effective vaccine remains a high priority. One of the most central vaccine platforms considered is plasmid DNA. However, high doses of DNA and several immunizations are typically needed to achieve detectable T-cell responses. In this study, a Semliki Forest virus replicon DNA vaccine designed for human clinical trials, DREP.HIVA, encoding an antigen that is currently being used in human trials in the context of a conventional DNA plasmid, pTHr.HIVA, was generated. It was shown that a single immunization of DREP.HIVA stimulated HIV-1-specific T-cell responses in mice, suggesting that the poor immunogenicity of conventional DNA vaccines may be enhanced by using viral replicon-based plasmid systems. The results presented here support the evaluation of Semliki Forest virus replicon DNA vaccines in non human primates and in clinical studies. PMID- 15659756 TI - Probing neutralizing-antibody responses against emerging measles viruses (MVs): immune selection of MV by H protein-specific antibodies? AB - Measles virus (MV) infection and vaccination induce long-lasting immunity and neutralizing-antibody responses that are directed against the MV haemagglutinin (H) and the fusion (F) protein. A new MV genotype, D7, emerged recently in western Germany and rapidly replaced the long-term endemically circulating genotypes C2 and D6. Analysis of the H gene of C2, D6, D7 and vaccine viruses revealed uniform sequences for each genotype. Interestingly, a consistent exchange of seven distinct amino acids in the D7 H was observed when compared with residues shared between C2, D6 and vaccine viruses, and one exchange (D416- >N) in the D7 H was associated with an additional N-linked glycosylation. In contrast, the F gene is highly conserved between MVs of these genotypes. To test whether the D7 H protein escapes from antibody responses that were raised against earlier circulating or vaccine viruses, the neutralizing capacity of mAbs recognizing seven distinct domains on the H of an Edmonston-related MV was compared. The mAbs revealed a selective and complete loss of two neutralizing epitopes on the D7 H when compared with C2, D6 and vaccine viruses. To assess whether these alterations of the D7 H affect the neutralizing capacity of polyclonal B-cell responses, genotype-specific antisera were produced in cotton rats. However, no significant genotype-dependent difference was found. Likewise, human sera obtained from vaccinees (n=7) and convalescents (n=6) did not distinguish between the MV genotypes. Although the hypothesis of selection of D7 viruses by pre-existing neutralizing antibodies is compatible with the differing pattern of neutralizing epitopes on the H protein, it was not confirmed by the results of MV neutralization with polyclonal sera. PMID- 15659755 TI - A subset of human immunodeficiency virus type 1 long-term non-progressors is characterized by the unique presence of ancestral sequences in the viral population. AB - Within human immunodeficiency virus type 1 (HIV-1)-infected patients, there are those who have been infected for more than 10 years with a CD4+ cell count of >500 cells microl(-1) and who remain asymptomatic without antiretroviral therapy; these patients are designated long-term non-progressors (LTNPs). In a set of 16 LTNPs, viral dating, DNA viral load, quasispecies heterogeneity and antibody (Ab) titres against gp160 and beta2 microglobulin (beta2m) were determined. Plasma viral RNA and CD4+ and CD8+ T-cell numbers were estimated in more than three samples per patient. Host genetic characteristics, such as Delta32-CCR5 genotype and human leukocyte antigen (HLA) genotype and supertypes, and clinical epidemiological factors were evaluated. Dating of global populations and of DNA and RNA viral quasispecies identified two subsets of patients: one displaying only ancestral sequences and the other displaying predominantly modern sequences. The ancestral patients displayed a significant reduction in RNA and DNA viral loads, quasispecies heterogeneity, CD8+ cell number, anti-gp160 Ab titres and beta2m level, and they were also associated with better use of safe-sex practices and higher presence of the HLA sB58 supertype than the modern subset. Viral dating has therefore permitted the segregation of LTNPs into two subsets that show very different virological, immunological, host and clinical-epidemiological characteristics. Moreover, whereas the modern subset displayed low levels of virus replication, the ancestral group displayed not only a very limited virus replication, often to undetectable levels, but also very slow or arrested viral evolution, maintaining the close relationship of the viral population to the transmitted virus. PMID- 15659757 TI - The respiratory syncytial virus small hydrophobic protein is phosphorylated via a mitogen-activated protein kinase p38-dependent tyrosine kinase activity during virus infection. AB - The phosphorylation status of the small hydrophobic (SH) protein of respiratory syncytial virus (RSV) was examined in virus-infected Vero cells. The SH protein was isolated from [35S]methionine- and [33P]orthophosphate-labelled RSV-infected cells and analysed by SDS-PAGE. In each case, a protein product of the expected size for the SH protein was observed. Phosphoamino acid analysis and reactivity with the phosphotyrosine specific antibody PY20 showed that the SH protein was modified by tyrosine phosphorylation. The role of tyrosine kinase activity in SH protein phosphorylation was confirmed by the use of genistein, a broad-spectrum tyrosine kinase inhibitor, to inhibit SH protein phosphorylation. Further analysis showed that the different glycosylated forms of the SH protein were phosphorylated, as was the oligomeric form of the protein. Phosphorylation of the SH protein was specifically inhibited by the mitogen-activated protein kinase (MAPK) p38 inhibitor SB203580, suggesting that SH protein phosphorylation occurs via a MAPK p38-dependent pathway. Analysis of virus-infected cells using fluorescence microscopy showed that, although the SH protein was distributed throughout the cytoplasm, it appeared to accumulate, at low levels, in the endoplasmic reticulum/Golgi complex, confirming recent observations. However, in the presence of SB203580, an increased accumulation of the SH protein in the Golgi complex was observed, although other virus structures, such as virus filaments and inclusion bodies, remained largely unaffected. These results showed that during RSV infection, the SH protein is modified by an MAPK p38-dependent tyrosine kinase activity and that this modification influences its cellular distribution. PMID- 15659758 TI - Genetic clustering of Borna disease virus natural animal isolates, laboratory and vaccine strains strongly reflects their regional geographical origin. AB - The aim of this study was to gain more detailed insights into the genetic evolution and variability of Borna disease virus (BDV). Phylogenetic analyses were performed on field viruses originating from naturally infected animals, the BDV vaccine strain 'Dessau', four widely used laboratory strains and the novel BDV subtype No/98. Four regions of the BDV genome were analysed: the complete p40, p10 and p24 genes and the 5'-untranslated region of the X/P transcript. BDV isolates from the same geographical area exhibited a clearly higher degree of identity to each other than to BDV isolates from other regions, independent of host species and year of isolation. Five different clusters could be established within endemic areas, corresponding to the geographical regions from which the viruses originated: (i) a Swiss, Austrian and Liechtenstein Rhine valley group, related closely to the geographically bordering Baden-Wurttemberg and Bavaria II group (ii) in the western part of Germany; (iii) a third group, called Bavaria I group, limited in occurrence to Bavaria; (iv) a southern Saxony-Anhalt and bordering northern Saxony group, bound to the territories of these federal states in the eastern part of Germany; and (v) a mixed group, consisting of samples from different areas of Germany; however, these were mainly from the federal states of Thuringia and Lower Saxony. The laboratory strains and the vaccine strain clustered within these groups according to their geographical origins. All field and laboratory strains, as well as the vaccine strain, clearly segregated from the recently described and highly divergent BDV strain No/98, which originated from an area in Austria where Borna disease is not endemic. PMID- 15659759 TI - Vaccine-induced protection against Borna disease in wild-type and perforin deficient mice. AB - Borna disease virus (BDV) can persistently infect the central nervous system and induce CD8+ T-cell-mediated neurological disease in MRL mice. To determine whether specific immune priming would prevent disease, a prime-boost immunization protocol was established in which intramuscular injection of a recombinant parapoxvirus expressing BDV nucleoprotein (BDV-N) was followed by intraperitoneal infection with vaccinia virus expressing BDV-N. Immunized wild-type and perforin deficient mice remained healthy after intracerebral infection with BDV and contained almost no virus in the brain at 5 weeks post-challenge. Immunization failed to induce resistance against BDV in mice lacking mature CD8+ T cells. Immunization of perforin-deficient mice with a poxvirus vector expressing mutant BDV-N lacking the known CD8+ T-cell epitope did not efficiently block multiplication of BDV in the brain and did not prevent neurological disease, indicating that vaccine-induced immunity to BDV in wild-type and perforin deficient mice resulted from the action of CD8+ T cells. PMID- 15659760 TI - Inhibition of West Nile virus entry by using a recombinant domain III from the envelope glycoprotein. AB - The envelope glycoprotein located at the outermost surface of the flavivirus particle mediates entry of virus into host cells. In this study, the involvement of domain III of West Nile virus (WNV-DIII) envelope protein in binding to host cell surface was investigated. WNV-DIII was first expressed as a recombinant protein and purified after a solubilization and refolding procedure. The refolded WNV-DIII protein displays a content of beta-sheets consistent with known homologous structures of other flavivirus envelope DIII, shown by using circular dichroism analysis. Purified recombinant WNV-DIII protein was able to inhibit WNV entry into Vero cells and C6/36 mosquito cells. Recombinant WNV-DIII only partially blocked the entry of dengue-2 (Den 2) virus into Vero cells. However, entry of Den 2 virus into C6/36 was blocked effectively by recombinant WNV-DIII. Murine polyclonal serum produced against recombinant WNV-DIII protein inhibited infection with WNV and to a much lesser extent with Den 2 virus, as demonstrated by plaque neutralization assays. Together these results provided strong evidence that immunoglobulin-like DIII of WNV envelope protein is responsible for binding to receptor on the surface of host cells. The data also suggest that similar attachment molecule(s) or receptor(s) were used by WNV and Den 2 virus for entry into C6/36 mosquito cells. PMID- 15659761 TI - Neuroblastoma cell-adapted yellow fever virus: mutagenesis of the E protein locus involved in persistent infection and its effects on virus penetration and spread. AB - Persistent infection of mouse neuroblastoma NB41A3 cells with yellow fever 17D virus generates viral variants which exhibit defective cell penetration, poor cell-to-cell spread, small plaque size and reduced growth efficiency, caused by substitution of glycine for aspartic acid or glutamic acid at positions 360 and 362 in the envelope protein. These positions occur within a charge cluster, Asp360-Asp361-Glu362, located in domain III, near its interface with domain I. To characterize further the molecular basis for the variant phenotype, a series of mutant viruses containing substitutions at position 360, 361 and 362, were studied for effects on the cell culture properties typical of the neuroblastoma adapted variant. Most substitutions at position 360 gave rise to viruses that were very defective in cell penetration, growth efficiency and cell-to-cell spread, whereas substitution with glutamic acid yielded a virus indistinguishable from parental yellow fever 17D. Substitution with lysine was not tolerated and substitution with asparagine resulted in frequent wild-type revertants. A glycine residue was not tolerated at position 361, but substitution at 362 yielded a small plaque virus, similar to the effect of substitution at position 360. These data indicate that the yellow fever virus E protein contains a locus within domain III where a negative-charge cluster is important for optimal function of this domain in virus-cell interactions beyond the stage of virus attachment. Modelling predictions suggest that the mutations alter the local properties of the loop within domain III, and may compromise interactions of this domain with an adjacent region of domain I during conformational changes that occur in the E protein in association with virus entry. PMID- 15659762 TI - Molecular characterization of the complete genome of human influenza H5N1 virus isolates from Thailand. AB - The complete genomes of three human H5N1 influenza isolates were characterized, together with the haemagglutinin (HA) and neuraminidase (NA) genes from two additional human isolates and one chicken isolate. These six influenza isolates were obtained from four different provinces of Thailand during the avian influenza outbreak in Asia from late 2003 to May 2004. All six Thailand isolates contained multiple basic amino acids at the cleavage site in the HA gene. Amino acid residues at the receptor-binding site of the five human viruses were similar to those of the chicken virus and other H5N1 viruses from Hong Kong. The presence of amantadine resistance in the Thailand viruses isolated during this outbreak was suggested by a fixed mutation in M2 and confirmed by a phenotypic assay. All genomic segments of the Thailand viruses clustered with the recently described genotype Z. The Thailand viruses contained more avian-specific residues than the 1997 Hong Kong H5N1 viruses, suggesting that the virus may have adapted to allow a more efficient spread in avian species. PMID- 15659763 TI - Following the very initial growth of biological RNA viral clones. AB - Due to their extremely high genetic diversity, which is a direct consequence of high mutation rates, RNA viruses are often described as molecular quasispecies. According to this theory, RNA virus populations cannot be understood in terms of individual viral clones, as they are clouds of interconnected mutants, but this prediction has not yet been demonstrated experimentally. The goal of this study was to determine the fitness of individual clones sampled from a given RNA virus population, a necessary previous step to test the above prediction. To do so, limiting dilutions of a vesicular stomatitis virus population were employed to isolate single viral clones and their initial growth dynamics were followed, corresponding to the release of the first few hundred viral particles. This technique is useful for estimating basic fitness parameters, such as intracellular growth rate, viral yield per cell, rate at which cells are infected and time spent in cell-to-cell transmission. A combination of these parameters allows estimation of the fitness of individual clones, which seems to be determined mainly by their ability to complete infection cycles more quickly. Interestingly, fitness was systematically higher for initial clones than for their derived populations. In addition to environmental changes, such as cellular defence mechanisms, these differences are attributable to high RNA virus mutation rates. PMID- 15659764 TI - Enteroviruses 76, 89, 90 and 91 represent a novel group within the species Human enterovirus A. AB - Molecular methods have enabled the rapid identification of new enterovirus (EV) serotypes that would have been untypable using existing neutralizing antisera. Nineteen strains of four new EV types termed EV76 (11 isolates), EV89 (two isolates), EV90 (four isolates) and EV91 (two isolates), isolated from clinical specimens from patients in France (one isolate) and Bangladesh (18 isolates), are described. Nucleotide sequences encoding the VP1 capsid protein (882-888 nt) are less than 65 % identical to the homologous sequences of the recognized human EV serotypes, but within each group the sequences are more than 78 % identical. The deduced amino acid sequences of the complete capsid (P1) region are more than 94 % identical within type but less than 76 % identical to those of the recognized serotypes. For both VP1 and P1, the 19 isolates are monophyletic by type with respect to all other EV serotypes. Using the proposed molecular typing scheme, these data support their identification as four new types within the species Human enterovirus A (HEV-A). In almost all cases, the VP1 sequences were more similar to those of some simian EVs than to the human EVs. Partial 3D sequences of all 19 isolates also clustered within HEV-A; they were monophyletic as a group, but not by type, suggesting that recombination has occurred among viruses of these four types. Partial 3D sequences were more closely related to those of simian EVs than to human viruses in HEV-A. These results suggest that the four new types may represent a new subgroup within HEV-A, in addition to the existing human and simian subgroups. PMID- 15659765 TI - Studies on overwintering of bluetongue viruses in insects. AB - Bluetongue viruses (BTVs) are economically important arboviruses that affect sheep and cattle. The overwintering mechanism of BTVs in temperate climates has eluded researchers for many years. Many arboviruses overwinter in their invertebrate vectors. To test the hypothesis that BTVs overwinter in their vertically infected insect vectors, Culicoides sonorensis larvae were collected from long-term study sites in northern Colorado, USA, and assayed for the presence of BTV RNA by nested RT-PCR. Sequences from BTV RNA segment 7 were detected in 30 % (17/56) of pools composed of larvae and pupae collected in 1998 and in 10 % (31/319) of pools composed of adults reared from larvae collected in 1996. BTV was not isolated from the insects. Additionally, Culicoides cell culture lines derived from material collected at one of the sites, or derived from insect samples collected during a BTV outbreak, contained BTV RNA segment 7. In contrast, segment 2 RNA was detected at half the rate of segment 7 RNA in the field-collected larvae and was only detected in the Culicoides cell lines with one of two primer sets. These data suggest that BTVs could overwinter in the insect vector and that there is reduced expression of the outer capsid genes during persistent infection. PMID- 15659766 TI - Use of a Beet necrotic yellow vein virus RNA-5-derived replicon as a new tool for gene expression. AB - A new gene-expression system based on RNA-5 of Beet necrotic yellow vein virus (BNYVV) was constructed to allow the expression of recombinant proteins in virally infected cells. Replication and expression levels of the RNA-5-based replicon containing the green fluorescence protein (GFP) gene were compared with those obtained with the well-characterized RNA-3-derived replicon (Rep-3). When RNA-3 and/or RNA-4 BNYVV RNAs were added to the inoculum, the expression levels of RNA-5-encoded GFP were considerably reduced. To a lesser extent, RNA-3-derived GFP expression was also affected by the presence of RNA-4 and -5. Both RNA-3- and RNA-5-derived molecules were able to express proteins within the same infected cells. Together with Rep-3, the RNA-5-derived replicon thus provides a new tool for the co-expression of different recombinant proteins. In Beta macrocarpa, Rep 5-GFP was able to move in systemic tissues in the presence of RNA-3 and thus provides a new expression system that is not restricted to the inoculated leaves. PMID- 15659767 TI - Intracellular salivation is the aphid activity associated with inoculation of non persistently transmitted viruses. AB - Approximately 75 % of aphid-vectored viruses are transmitted in a non-persistent (non-circulative) manner. Localization studies indicate that such viruses are acquired via ingestion and retained in the food canal of the maxillary stylets, but the inoculation mechanism has remained unresolved. Electrical recording of stylet penetration activities reveals that inoculation is associated with the first intracellular activity (subphase II-1) following maxillary puncture of an epidermal cell. Subphase II-1 may represent virus inoculation via egestion (regurgitation of virions with food-canal contents) or salivation (saliva mediated release of virions from the common food-salivary duct at the tips of the maxillary stylets). Here, inoculation of the circulatively transmitted Pea enation mosaic virus was used as a marker for intracellular salivation during epidermal cell punctures. The results confirmed that inoculation of non persistently transmitted viruses (subphase II-1) is associated with active injection of saliva directly into the cytoplasm. PMID- 15659768 TI - Host-directed processing of Citrus exocortis viroid. AB - Prolonged infection of tomato hybrid (Lycopersicon esculentum x Lycopersicon peruvianum) by Citrus exocortis viroid (CEVd) resulted in viroid-like enlarged structures, detected by gel electrophoresis. This population included two new enlarged variants or D-variants, D-87 and D-76, and three transient species or D forms, D-38, D-40 and D-43. Sequence analyses exposed a locus near the terminal repeat region where major changes appeared consistently. In transmission tests to CEVd hosts, a variety of progeny populations were recovered, including progeny enlargements of and reversions to CEVd, as well as sequence fidelity to the inoculum. Transmission tests to citrus hosts of the genera Citrus, Poncirus or Fortunella were unsuccessful. The importance of host specificity to the recovery and processing of the various CEVd-related structures, as well as the temporal variability of progeny populations, was demonstrated. PMID- 15659769 TI - Localization of Poa semilatent virus cysteine-rich protein in peroxisomes is dispensable for its ability to suppress RNA silencing. AB - Subcellular localization of the Poa semilatent virus cysteine-rich gammab protein was studied by using different approaches. In infected tissue, gammab was detected mainly in the P30 fraction as monomers, dimers and oligomers. Green fluorescent protein-fused gammab was found to localize in punctate bodies in the cytoplasm. Colocalization with marker proteins demonstrated that these bodies represent peroxisomes. Immunoelectron microscopy revealed that gammab was localized in the peroxisomal matrix and that localization of gammab in peroxisomes required the C-terminal signal tripeptide SKL. An SKL-deletion mutant exhibited a diffuse localization, but retained the protein's ability to suppress RNA silencing, determine infection phenotype and support virus systemic spread. These data indicate that gammab functions are not associated with the protein's localization to peroxisomes. PMID- 15659770 TI - Taro vein chlorosis virus: characterization and variability of a new nucleorhabdovirus. AB - Sequencing of the monopartite RNA genome of a Fijian isolate of Taro vein chlorosis virus (TaVCV) confirmed that it is a definitive rhabdovirus with most similarity to members of the genus Nucleorhabdovirus. The TaVCV 12 020 nt negative-sense RNA genome contained six ORFs in the antigenomic sequence, equivalent to the N, P, 3, M, G and L genes that have been identified in other rhabdoviruses. The putative gene products had highest similarity to those of the nucleorhabdovirus Maize mosaic virus. A characteristic 3'-AAUUCUUUUUGGGUUGU/A-5' sequence was identified in each of the intergenic regions and the TaVCV leader and trailer sequences comprised 140 and 61 nt, respectively. Assignment of TaVCV to the genus Nucleorhabdovirus was supported by thin-section electron microscopy of TaVCV-infected taro leaves, which identified virions budding from nuclear membranes into the perinuclear space. Variability studies identified high levels of TaVCV sequence diversity. Within the L gene of 20 TaVCV isolates from Fiji, the Federated States of Micronesia, New Caledonia, Papua New Guinea, Solomon Islands and Vanuatu, maximum variability at the nucleotide level was 27.4 %. Within the N gene, maximum variability among 15 isolates at the nucleotide level was 19.3 %. The high level of TaVCV variability observed suggested that the introduction of TaVCV to the Pacific Islands was not a recent occurrence. PMID- 15659771 TI - Mutations in Turnip mosaic virus genomes that have adapted to Raphanus sativus. AB - The genetic basis for virulence in potyviruses is largely unknown. Earlier studies showed that there are two host types of Turnip mosaic virus (TuMV); the Brassica/Raphanus (BR)-host type infects both Brassica and Raphanus systemically, whereas the Brassica (B)-host type infects Brassica fully and systemically, but not Raphanus. The genetic basis of this difference has been explored by using the progeny of an infectious clone, p35Tunos; this clone is derived from the UK1 isolate, which is of the B-host type, but rarely infects Raphanus systemically and then only asymptomatically. Two inocula from one such infection were adapted to Raphanus by passaging, during which the infectivity and concentration of the virions of successive infections increased. The variant genomes in the samples, 16 in total, were sequenced fully. Four of the 39 nucleotide substitutions that were detected among the Raphanus sativus-adapted variant genomes were probably crucial for adaptation, as they were found in several variants with independent passage histories. These four were found in the protein 1 (P1), protein 3 (P3), cylindrical inclusion protein (CI) and genome-liked viral protein (VPg) genes. One of four 'parallel evolution' substitutions, 3430G-->A, resulted in a 1100Met- >Ile amino acid change in the C terminus of P3. It seems likely that this site is important in the initial stages of adaptation to R. sativus. Other independent substitutions were mostly found in the P3, CI and VPg genes. PMID- 15659772 TI - Banana contains a diverse array of endogenous badnaviruses. AB - Banana streak disease is caused by several distinct badnavirus species, one of which is Banana streak Obino l'Ewai virus. Banana streak Obino l'Ewai virus has severely hindered international banana (Musa spp.) breeding programmes, as new hybrids are frequently infected with this virus, curtailing any further exploitation. This infection is thought to arise from viral DNA integrated in the nuclear genome of Musa balbisiana (B genome), one of the wild species contributing to many of the banana cultivars currently grown. In order to determine whether the DNA of other badnavirus species is integrated in the Musa genome, PCR-amplified DNA fragments from Musa acuminata, M. balbisiana and Musa schizocarpa, as well as cultivars 'Obino l'Ewai' and 'Klue Tiparot', were cloned. In total, 103 clones were sequenced and all had similarity to open reading frame III in the badnavirus genome, although there was remarkable variation, with 36 distinct sequences being recognized with less than 85 % nucleotide identity to each other. There was no commonality in the sequences amplified from M. acuminata and M. balbisiana, suggesting that integration occurred following the separation of these species. Analysis of rates of non-synonymous and synonymous substitution suggested that the integrated sequences evolved under a high degree of selective constraint as might be expected for a living badnavirus, and that each distinct sequence resulted from an independent integration event. PMID- 15659773 TI - Back-priming mode of phi6 RNA-dependent RNA polymerase. AB - The RNA-dependent RNA polymerase of the double-stranded RNA bacteriophage phi6 is capable of primer-independent initiation, as are many RNA polymerases. The structure of this polymerase revealed an initiation platform, composed of a loop in the C-terminal domain (QYKW, aa 629-632), that was essential for de novo initiation. A similar element has been identified in hepatitis C virus RNA dependent RNA polymerase. Biochemical studies have addressed the role of this platform, revealing that a mutant version can utilize a back-priming initiation mechanism, where the 3' terminus of the template adopts a hairpin-like conformation. Here, the mechanism of back-primed initiation is studied further by biochemical and structural methods. PMID- 15659775 TI - A beginner's guide to NF-kappaB signaling pathways. AB - Nuclear factor kappaB (NF-kappaB) belongs to a family of heterodimeric transcription factors that play a key role in inflammatory and stress responses as well as in tumor cell resistance to apoptosis. These effects are due to the NF kappaB-dependent transcription of many proinflammatory and antiapoptotic genes, whose products ensure various cell responses to environmental conditions. The signal transduction pathways leading to NF-kappaB activation are well characterized, and the different steps implicated in these pathways involve proteins that could constitute targets for NF-kappaB inhibition. Several inhibitors aiming to prevent NF-kappaB activity and thus the transcription of target genes are studied, and a few compounds seem particularly promising. We try here to summarize the advantages that can issue from various studies on NF kappaB. PMID- 15659774 TI - A multifunctional aromatic residue in the external pore vestibule of Na+ channels contributes to the local anesthetic receptor. AB - Voltage-gated Na(+) (Na(v)) channels are responsible for initiating action potentials in excitable cells and are the targets of local anesthetics (LA). The LA receptor is localized to the cytoplasmic pore mouth formed by the S6 segments from all four domains (DI-DIV) but several outer pore-lining residues have also been shown to influence LA block (albeit somewhat modestly). Many of the reported amino acid substitutions, however, also disrupt the inactivated conformations that favor LA binding, complicating the interpretation of their specific effects on drug block. In this article, we report that an externally accessible aromatic residue in the Na(v) channel pore, DIV-Trp1531, when substituted with cysteine, completely abolished LA block (e.g., 300 microM mexiletine induced a use dependent block with 65.0 +/- 2.9% remaining current and -11.0 +/- 0.6 mV of steady-state inactivation shift of wild-type (WT) channels versus 97.4 +/- 0.7% and -2.4 +/- 2.1 mV of W1531C, respectively; p < 0.05) without destabilizing fast inactivation (complete inactivation at 20 ms at -20 mV; V(1/2) = -70.0 +/- 1.6 mV versus -48.6 +/- 0.5 mV of WT). W1531C also abolished internal QX-222 block (200 microM; 98.4 +/- 3.4% versus 54.0 +/- 3.2% of WT) without altering drug access. It is interesting that W1531Y restored WT blocking behavior, whereas W1531A channels exhibited an intermediate phenotype. Together, our results provide novel insights into the mechanism of drug action, and the structural relationship between the LA receptor and the outer pore vestibule. PMID- 15659776 TI - The vascular endothelial growth factor-induced disruption of gap junctions is relayed by an autocrine communication via ATP release in coronary capillary endothelium. AB - Little is known concerning how the coordination of Ca(2+) signaling aids in capillary endothelial cell (CEC) functions, such as microvascular permeability and angiogenesis. Previous reports support the major involvement of gap junction (GJ) channels. However, the cell-to-cell communication may not be straightforward, especially if we consider the participation of active molecules released by CEC. In this study, short-term effects of vascular endothelial growth factor (VEGF-165) were compared with those of bradykinin (BK) on gap junction coupling (GJC) and remodeling of connexin-43 (Cx43) and then analyzed for intercellular Ca(2+) signal in primary cultures of coronary CEC. Dye-coupling experiments revealed that BK or VEGF completely blocked GJC. These effects correlated with the rapid internalization of Cx43 and its tyrosine phosphorylation in part via the phosphatidylinositol 3-kinase/Akt pathway. GJC slowly recovered with BK but not with VEGF in the following hour. In control conditions, mechanical stimulation of a single cell within a confluent monolayer triggered an intercellular Ca(2+) wave that was partially inhibited by GJC blockers or purinergic inhibitors. No wave propagation was observed after blockage of both GJC and purinergic receptors. Cell treatment with VEGF also reduced propagation of the Ca(2+) wave, which was totally prevented by applying a purinergic receptor antagonist but not with a GJC blocker. That excludes purine efflux through Cx hemichannels. We conclude that VEGF-induced disruption of GJC via Cx43 remodeling is relayed by an autocrine communication via secretion of ATP to preserve intercellular Ca(2+) signaling in capillary endothelium. PMID- 15659777 TI - Exogenous tissue inhibitor of metalloproteinase-1 promotes endothelial cell survival through activation of the phosphatidylinositol 3-kinase/Akt pathway. AB - Control of molecular targets and signaling pathways which improve endothelial cell survival may be an attractive concept for interfering with dysregulated vascular injury and remodeling, a key mechanism for transplant arteriosclerosis and chronic allograft rejection. In addition to inhibiting matrix metalloproteinase activity, it has been suggested by recent studies that the tissue inhibitor of metalloproteinase (TIMP)-1 may inhibit apoptosis in various cell types. The present work examines the possibility that TIMP-1 belongs to a protective pathway via antiapoptotic properties and investigates the signaling pathway mediated by TIMP-1 in human ECs. We demonstrate that exogenous, recombinant, TIMP-1 efficiently prevents apoptosis induced by TNFalpha in cycloheximide-sensitized ECs. The antiapoptotic effect of TIMP-1 was dose dependent and a maximal effect of TIMP-1 (30% protection) was reached using 250 ng/mL of recombinant TIMP-1. We present evidence that TIMP-1 induces activation of PI3-kinase but not NFkappaB pathway in ECs. Our findings further indicate that TIMP-1-induced EC survival is mediated through activation of PI3-kinase pathway and the downstream phosphorylation of Akt kinase. Blocking the PI3-kinase pathway with wortmannin or LY294002 restores TNFalpha-mediated EC death. In conclusion, our findings suggest that TIMP-1, generated upon inflammation, acts as an antiapoptotic molecule that can prevent EC apoptosis through activation of the PI3-kinase and phosphorylation of the Akt kinase. PMID- 15659778 TI - The cyclooxygenase inhibitor flurbiprofen reduces radiation-induced angiogenic growth factor secretion of squamous cell carcinoma cell lines. AB - Surgical intervention and radiotherapy still represent the gold standard in the therapy of head and neck squamous cell carcinoma (SCC), although often with unsatisfactory results. Radiation might induce the expression and secretion of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) by unknown mechanisms. These two highly active proangiogenic and cytoprotective factors might contribute to a limited therapeutic success by promoting revascularization and cytoprotection of the radiated tumor. The aim of the present study was to analyze the potential of the cyclooxygenase inhibitor flurbiprofen to reduce radiation-induced increase of VEGF and bFGF secretion of tumor cells. We analyzed the expression of VEGF and bFGF at 72 h after radiation with 30 Gy in four SCC cell lines (De-pt, Hun, Lau, and A549) in cell culture with or without added flurbiprofen. Controls were not exposed to radiation and were analyzed at the same time after culture in the same media. We observed increased VEGF levels in all and increased bFGF levels in three of four lines after radiation. In irradiated cultures with flurbiprofen, VEGF was reduced between 13% and 26% and bFGF was reduced between 84% and 93% compared with radiated cultures without flurbiprofen. We found no reduction of VEGF and bFGF secretion in the unirradiated cultures despite added flurbiprofen. We conclude that flurbiprofen is able to alter the radiation-induced secretion of these two growth factors and might be useful in decreasing the resistance of SCC to radiation. PMID- 15659780 TI - Design and characterization of a new class of inhibitors of ras activation. AB - The Ras proteins, which regulate intracellular signaling by a cyclic process involving interconversion between active GTP-bound and inactive GDP-bound states, play an essential role in controlling the activity of several crucial signaling pathways regulating normal cellular proliferation. Mutational activation of RAS genes can induce cancer in humans and other mammals. About 30% of human tumors contain an altered oncogenic Ras; therefore, inhibitors of Ras activation are potentially antineoplastic drugs. In this work we describe original molecules acting as Ras inhibitors. Recently a new class of inhibitors of the Ras nucleotide exchange process was described by Taveras et al. These molecules are able to form a noncovalent complex with Ras-GDP, inhibiting the GDP-GTP nucleotide exchange. We synthesized molecule SCH-53870 and we found that it inhibits p21-hRas nucleotide exchange in vitro, but it has very low solubility in water and undergoes rapid degradation at room temperature when dissolved in water DMSO mixtures. This chemical instability could prejudice pharmacological activity in vivo. With the aim to improve solubility and chemical stability, we designed and synthesized other original bioactive molecules that have been characterized in vitro using purified human and yeast Ras proteins and in vivo using suitable Saccharomyces cerevisiae strains. In the long term we hope that the knowledge we derive from these compounds will help in the development of an alternative therapy targeting Ras for a specific inhibition of transformed cell proliferation. PMID- 15659779 TI - Spontaneous generation of reactive oxygen species in the mixture of cyanide and glycerol. AB - Reactive oxygen species are involved in tumor promotion or apoptosis. In assaying prooxidant or antioxidant activities, cyanide has been commonly used as an inhibitor of mitochondrial oxidases, peroxidases, or Cu,Zn-superoxide dismutase, which have an influence on intracellular levels of reactive oxygen species. It has also been used to chemically mimic hypoxia. On the other hand, glycerol has been widely used as a stabilizer of various enzymes. In particular, glycerol is required to maintain the enzymatic activities of membrane-bound NAD(P)H oxidases extracted from surrounding phospholipids. Since both cyanide and glycerol are relatively inert, they have been used concomitantly regardless of any mutual interference. In this study, we demonstrate that a mixture of glycerol and cyanide reduced cytochrome c and nitroblue tetrazolium, both of which are superoxide anion indicators. The mixture also enhanced the production of superoxide anion in the presence of redox-cycling compounds. Superoxide production by the mixture was confirmed by electron spin resonance spectra. Moreover, the mixture induced lipid peroxidation and hemolysis in human erythrocytes. These results suggest that cyanide and glycerol should be used carefully in reaction systems used to measure superoxide production or antioxidant activity. However, sucrose and sodium azide in combination do not produce such artifacts and thus may be used as an alternative. PMID- 15659782 TI - Epidermal growth factor receptor as a novel therapeutic target in anaplastic thyroid carcinomas. AB - Anaplastic thyroid carcinoma (ATC) is one of the most aggressive human malignancies, with a median survival of up to 6 months. Such a bad prognosis under the present treatment procedures suggests the need for novel approaches in the management of this disease. Since some epidermal growth factor receptor (EGFR) inhibitors are now in clinical trials and few data are available concerning EGFR expression in anaplastic thyroid carcinomas, we tried to estimate a possible overexpression of this receptor in a larger tumor series. Twenty-five ATCs, including 3 ATCs with poorly differentiated thyroid carcinoma (PDTC) parts, were immunohistochemically investigated with a mouse monoclonal antibody directed against EGFR (EGFR pharmDX kit). The tumors revealed primarily a distinct membranous staining pattern, and in several tumor cells an additional cytoplasmic reactivity could be observed. The anaplastic carcinomas presented with 5 of 25 (20%) without EGFR reaction, 10 of 25 (40%) with reactivity, and 10 of 25 (40%) with overexpression of the receptor. All ATCs with PDTC parts (100%) showed EGFR overexpression. Cytoplasmic reactivity was observed in 56% of all ATCs. A significant correlation was calculated for EGFR overexpression and cytoplasmic staining (P = 0.036). Concerning receptor overexpression, ATCs were significantly different from ATCs with PDTC parts (P = 0.023). For the first time, we present EGFR overexpression in ATC in a larger tumor series, demonstrating that EGFR overexpression is a common finding in ATC. For at least one-third of all anaplastic thyroid carcinomas, EGFR seems to be a promising agent for the targeted molecular therapy of these extraordinarily aggressive tumors. PMID- 15659781 TI - Endogenous oxidative stress induces distinct redox forms of tumor necrosis factor receptor-1 in melanoma cells. AB - Receptors of the TNFR superfamily possess abundant thiols in their extracellular domains, which makes them susceptible to redox modulation by prooxidant agents and processes. Previous studies from our laboratory have documented that membrane gamma-glutamyltransferase (GGT) activity can originate reactive oxygen species in the extracellular milieu, during the GGT-mediated metabolism of extracellular glutathione. The present study was aimed thus to verify a possible redox modulating effect of GGT activity on TNFR1 receptors. The thiol-specific probe maleimide-polyethylene glycol was used to selectively label the reduced thiol groups in proteins of cell lysates; fractions corresponding to TNFR1 were then identified by immunoblot. In human melanoma Me665/2 cells, expressing varying GGT levels, at least five distinct forms of TNFR1 have been thus identified. The more oxidized forms appear to be prevalent in the 2/60 clone, expressing higher GGT levels, as compared to clone 2/21. Stimulation of GGT activity in the latter induced an increase of the oxidized TNFR1 forms. It is conceivable that different redox states of TNFR1 may correspond to different binding affinity and/or changes in the transducing function of the receptor. As GGT is frequently expressed by malignant tumors, the described phenomena might concur to alter the sensitivity of cancer cells to agents targeted on activation of TNF-alpha-dependent signaling pathways. PMID- 15659783 TI - Analysis of the effects of different alcohols on MCF-7 human breast cancer cells. AB - Alcohol consumption is known to be an increased risk factor for breast cancer, but the underlying molecular mechanisms are not well understood. We have recently shown that the exposure of MCF-7 breast cancer cells to 0.1% ethanol enhanced their proliferation and increased their content in both estrogen receptor-alpha (ERalpha) and aromatase. The aim of the present work was to determine if the effects of ethanol could be mimicked by other short-chain aliphatic alcohols such as methanol and 1-butanol. Our results show that these compounds do not stimulate MCF-7 cell proliferation. An increase in ERalpha content was observed by Western blot in methanol-treated cells, but this parameter was not affected in butanol treated cells. Neither of these two alcohols induced an increase in aromatase mRNA level. So despite a similarity in molecular structure, these primary alcohols do not exert the same effects. Taken together, these results suggest that the increase in aromatase expression might be a key event required for the enhanced proliferation observed in the presence of ethanol. PMID- 15659784 TI - Redox signaling by ionizing radiation in mouse liver. AB - Since radiation treatment has been reappraised in the treatment of hepatic tumors, radiation response in the liver is emerging as an interesting new area of investigation. In this study, identification of the repertoire of signaling proteins was performed using a proteomics approach involving cellular responses of liver tissue to ionizing radiation. Approximately 800 protein spots were detected. Among them, at least 28 proteins showed significant quantitative alterations after radiation. The significantly altered proteins were categorized as those related to reactive oxygen species (ROS) metabolism, metabolic pathway proteins, and G-type proteins. Particularly, the expression levels of proteins related to ROS metabolism, including cytochrome c, glutathione S-transferase Pi, NADH dehydrogenase, and peroxiredoxin VI, were increased after radiation. It is suggested that although radiation initiates cytotoxic effects, it can also induce a radioprotective antioxidant system. PMID- 15659785 TI - Enhancement of tumor response by farnesyltransferase inhibitor in C3H/HeJ hepatocarcinoma. AB - Farnesyltransferase inhibitor (FTI) acts on ras, which can ultimately enhance radiosensitivity. The objective of this study was to explore whether FTI could potentiate the antitumor efficacy of radiation in vivo, particularly in radio resistant hepatocarcinomas (HCa-I) syngeneic to C3H/HeJ mice. The presence of ras mutations was examined by PCR and DNA sequencing. C3H/HeJ mice, bearing HCa-I, were treated with FTI, LB42907, and 25 Gy radiation. FTI was orally administered, 60 mg/kg, twice daily for 30 days. The expression of regulating molecules was analyzed by Western blotting for p53, p21(WAF1/CIP1), and the Bcl-2 family, such as Bcl-2, Bax, and Bcl-X(L/s). In HCa-I, no ras mutations were detected. Downregulation of ras by FTI was most prominent at 4 h after treatment. In a tumor growth delay assay, FTI increased the effect of the tumor's radioresponse, with an enhancement factor of 1.32. Combined irradiation and FTI increased radiation-induced apoptosis; the peak apoptotic index was 3.6% with irradiation alone and with the drug alone but 7.1% in the combined treatment group. The analysis of apoptosis-regulating molecules by Western blotting showed upregulation of p53 and p21(WAF1/CIP1) in the combined treatment group compared with those in either of the single treatment groups, but the Bcl-2 family remained unchanged. FTI, in combination with radiation therapy, may have potential benefits in cancer treatment even if there are no ras mutations. FTI could inhibit ras activity but may also affect any protein that requires farnesylation for its activity. PMID- 15659786 TI - Three-dimensional bioreactor cultures: a useful dynamic model for the study of cellular interactions. AB - The ex vivo expansion of hematopoietic cells is a developing area with emphasis on bioreactor systems for amelioration of culture conditions. A rational design of bioreactors, especially those allowing microgravity, could permit the production of stem cells and will offer new approaches for studying the mechanisms of proliferation, differentiation, and signal transduction of cultured cells. The efficacy of two commercially available bioreactors (rotating-vessel miniPERM and static INTEGRA CL 350) to support long-term bone marrow cell cultures (LTBMCC) and three-dimensional growth of Hodgkin's lymphoma HD-MY-Z cells was investigated. In the miniPERM system, the growth of LTBMCC spheroids (containing 30-40 cells) was obtained. An essentially higher content of hematopoietic precursor cells (colony-forming units-granulocyte macrophage) was registered in the rotating-vessel system. In this bioreactor, a growth of large HD-MY-Z spheroids (containing 100-200 cells) was achieved. The composed mathematical models of the physicomechanical behavior of spheroids enabled the evaluation of the revolution frequency increase schedule. The differential equations took into account all inertial effects caused by the production module rotation movement as well as those caused by the relative movement of the spheroid in the fluid. The models aimed at the optimization of the rotation frequency increase schedule for different types of cells to reduce shear stress, augment productivity, and tolerate the growth of large spheroids. The models were numerically tested using MATLAB-SIMULINK software, and the trajectories of prestained HD-MY-Z spheroids were filmed. The coincidence of the theoretical and experimental trajectories was sufficient. PMID- 15659787 TI - Platelet-activating factor receptor signals in rat osteoblasts during spaceflight. AB - The platelet-activating factor (PAF) is a lipid mediator. The G-protein-coupled receptor of PAF (PAF-R) is activated by inflammatory and stressful conditions in numerous cell types. PAF/PAF-R is involved in apoptotic and antiapoptotic processes. We examined microgravity effects on the expression of PAF-R and second messengers in rat osteoblasts. The PAF-R signals are transmitted via arachidonic acid, phospholipase C (PLC), protein kinase C (PKC), and mitogen-activated protein kinase. Rat osteoblasts were cultured for 4 and 5 days aboard a space shuttle and solubilized on board. PAF-R gene expression in flight cultures increased to 2-6-fold higher than in ground controls. Gene expression of the G protein alpha subunit Galphaq in flight cultures increased to 3-fold and higher than in ground controls. It is known that Galphaq stimulates the effecter PLCbeta, activating PKC. The mRNA levels of PKCdelta and PKCtheta in flight cultures were increased to 2-5-fold higher than in ground controls. The PKCalpha mRNA level in flight cultures was increased to 3-fold higher than in ground controls on the 4th day. Gene expression of catalytic and regulatory subunits of protein kinase A was suppressed in flight cultures. PKCdelta and PKCtheta are novel PKCs that can be target substrates of caspases. The PAF-R gene may act as a mechano-sensitive gene that is involved in the apoptotic and antiapoptotic processes of osteoblasts under microgravity. PMID- 15659788 TI - Osteoblast responsiveness to 1alpha,25-dihydroxyvitamin D3 during spaceflight. AB - Rat osteoblasts were cultured aboard a space shuttle for 4 and 5 days. Cells were treated with 1 nM 1alpha,25-dihydroxyvitamin D(3) (VD) for the last 1 day. The conditioned media were harvested. Cells were solubilized with guanidine solution on board. We examined microgravity effects on the production/expression of osteocalcin, bone sialoprotein (BSP), and VD receptor (VD-R) in osteoblasts. Under VD treatment, the osteocalcin protein level was 243 +/- 117 and 1,718 +/- 534 pg/microg cellular DNA in flight cultures and ground controls, respectively. Without VD treatment, the osteocalcin protein level was not different between flight cultures and ground controls. The osteocalcin mRNA level in the VD-treated flight cultures was as low as 16% of that in ground controls. The VD-R mRNA level in the VD-treated flight cultures was also decreased to 16% of that in ground controls. Microgravity would suppress the VD-inducible production of osteocalcin but not the basal productivity. The BSP mRNA level was increased by microgravity. VD/VD-R binds to the vitamin D-responsive element (VDRE) on the target genes. The rat osteocalcin gene is positively regulated via "enhancer" VDRE, whereas the rat BSP gene is negatively regulated via "repressor " VDRE. Microgravity might modulate osteoblast responsiveness to VD through the suppression of VD-R. PMID- 15659789 TI - Low-grade inflammation with aging has consequences for T-lymphocyte signaling. AB - T lymphocytes are key players of immune responses that are associated with immune senescence because their functions are the most affected with aging. Defects in signal delivery to the nucleus have been demonstrated, by our group and others, in human aging and may explain the dysfunctions that occur in T cells with aging. Although aging has been related to a decline in several biological functions, it is also associated with a basal low-grade proinflammatory state and an increase in oxidative stress that lead to changes and damage. However, there are no clear data concerning the basal state of activation of T lymphocytes and its putative link with the low-grade inflammation observed with aging. Since membrane microdomains (lipid rafts) are specialized plasma membrane structures involved in T-lymphocyte activation, we studied the effect of aging on the phosphorylation state of signaling molecules associated with lipid rafts. We found that the signaling molecules in T lymphocytes from elderly donors were hyperphosphorylated and that the high basal state of phosphorylation did not allow activation exposure to a T-cell stimulus. We found that the cholesterol composition is changed in lipid rafts of resting T cells. We analyzed lipid raft distribution in situ using confocal microscopy and found a disorganization of these microdomains in T cells from aged donors. In conclusion, we show a link between aging, T lymphocyte lipid rafts, immune senescence, and low-grade inflammation. PMID- 15659790 TI - Role of the hedgehog/patched signaling pathway in oncogenesis: a new polymorphism in the PTCH gene in ovarian fibroma. AB - We compared the expression of target genes of Hedgehog/Patched signaling in ovarian fibromas and ovarian dermoids. We noted that high levels of SHH appear almost regularly, especially in dermoids, usually accompanied by increased expression of SMO. GLI overexpression does not coincide with that of PTCH. Loss of heterozygosity findings in the PTCH locus and increased expression of several genes in the pathway strongly suggest that the pathway is involved in both ovarian fibroma and dermoids. PMID- 15659791 TI - Effects of oxalate on IMCD cells: a line of mouse inner medullary collecting duct cells. AB - Oxalate, a metabolic end product and a major constituent of the majority of renal stones, has been shown to be toxic to renal epithelial cells of cortical origin. However, it is unknown whether inner medullary collecting duct (IMCD) cells that are physiologically exposed to higher concentrations of oxalate also behave in a similar manner. In the present study, we examined the effects of oxalate on IMCD cells. IMCD cells from the mouse were maintained in DMEM/F12 media supplemented with fetal bovine serum and antibiotics. Exposure of IMCD cells to oxalate produced time- and concentration-dependent changes in the light microscopic appearance of the cells. Long-term exposure to oxalate resulted in alterations in cell viability, with net cell loss after exposure to concentrations of 2 mM or greater. The production of free radicals was directly related to the exposure time and the concentration of oxalate. Crystal formation occurred in less than 1 h and cells in proximity to crystals would lose membrane integrity. Compared with IMCD cells, LLC-PK1 cells as well as HK-2 cells showed significant toxicity starting at lower oxalate concentrations (0.4 mM or greater). These results provide the first direct demonstration of toxic effects of oxalate in IMCD cells, a line of renal epithelial cells of the inner medullary collecting duct, and suggest that the cells lining the collecting duct are relatively resistant to oxalate toxicity. PMID- 15659792 TI - Supervision of multiple signaling protein kinases by the CK2-Cdc37 couple, a possible novel cancer therapeutic target. AB - Overexpression of a pleiotropic Ser/Thr kinase CK2 (casein kinase II), or of a kinase-targeting molecular chaperone Cdc37, induces neoplastic cell growth in animals. Recent genetic and biochemical evidence from several laboratories has revealed an unexpected direct link between CK2 and Cdc37. In this short review, we describe the basic characteristics of CK2 and Cdc37 and introduce recent findings on the interaction between CK2 and Cdc37. Cdc37 was identified as a multicopy suppressor of a temperature-sensitive allele of CK2 in Saccharomyces cerevisiae. CK2 phosphorylates a conserved serine residue in the N-terminal extremity of Cdc37 in vitro and in yeast as well as mammalian cells, and this is the unique phosphorylation site of Cdc37 under normal conditions. Mutations in the CK2-mediated phosphorylation site abolish the association of Cdc37 with various protein kinases. The same mutations in yeast cause severe growth and morphological defects. Specific inhibition of CK2 activity decreases intracellular levels of Cdc37-dependent protein kinases. Altogether, this evidence clearly indicates that the CK2-dependent phosphorylation is essential for the proper function of Cdc37 to bind and stabilize signaling protein kinases. In contrast, CK2 activity is enhanced by Cdc37 both in vitro and in vivo; thus, the CK2-Cdc37 couple seems to constitute a positive feedback control mechanism that may govern the activity of multiple protein kinases. Cdc37-dependent protein kinases include important signaling molecules whose disregulations are intimately related to neoplastic cell growth; hence, inhibition of the CK2-Cdc37 system may simultaneously suppress various cancer-promoting signal cascades. We propose that the CK2-Cdc37 couple can be a novel and efficient pharmacological target for cancer chemotherapy. PMID- 15659793 TI - Hypergravity stimulates osteoblast phenotype expression: a therapeutic hint for disuse bone atrophy. AB - Physiological actions of osteoblasts are disordered by gravity unloading. We investigated the possibility that the appropriate level of hypergravity could improve osteoblast functions that are susceptible to mechanical unloading. We evaluated hypergravity effects on the 1alpha,25-dihydroxyvitamin D(3) (VD) inducible osteocalcin expression of primary rat osteoblasts. Cell culture plates were centrifuged for 24 h at 3, 6, 12, 24, and 48 g in a 37 degrees C incubator. The mRNA levels were analyzed by quantitative RT-PCR. The mRNA levels for osteocalcin and vitamin D receptor (VD-R) at 12 g were enhanced to 187% and 228% of the 1 g control, respectively. However, the excess hypergravity conversely decreased osteocalcin expression. Osteocalcin gene expression was enhanced by VD/VD-R through the vitamin D-responsive element in the promoter. The increased osteocalcin expression might reflect the augmented VD-R expression. Alternatively, Runx2, a master gene of osteoblast differentiation, might be responsible for the osteocalcin induction, since the Runx2 mRNA levels were also increased to 247% of control at 12 g. Another VD-inducible osteoblast phenotype, alkaline phosphatase, was also upregulated at 12 g and 24 g. The appropriate level of hypergravity enhanced the VD-inducible expression of osteocalcin, a typical phenotype of osteoblast differentiation. These data suggest molecular features to prevent disuse bone atrophy of long-term bed-rest patients. PMID- 15659794 TI - CXCR4 activation induces epidermal growth factor receptor transactivation in an ovarian cancer cell line. AB - Chemokines are a family of proteins that have pleiotropic biological effects. They are well known to regulate the recruitment and trafficking of leukocytes to sites of inflammation. Chemokines are grouped into four classes based on the positions of key cysteine residues: C, CC, CXC, and CX3C. Stromal cell-derived factor-1 (SDF-1), the ligand of the CXCR4 receptor, is a CXC chemokine and is a highly conserved gene. Ovarian cancer typically disseminates widely in the abdomen, a characteristic that limits curative therapy. The mechanisms that promote ovarian cancer proliferation are incompletely understood. We studied a human ovarian adenocarcinoma cell line (OC 314) and investigated the role of CXCR4 activation by SDF-1 in human ovarian cancer. We demonstrate that CXCR4 and SDF-1 mRNA are expressed in OC 314. We show that SDF-1alpha induces proliferation in ovarian cancer cells in a dose-dependent manner. Moreover, we demonstrate that the SDF-1-dependent proliferation correlates to the phosphorylation and activation of extracellular signal-regulated kinases (ERK)1/2, which in turn are correlated to epidermal growth factor (EGF) receptor transactivation. In fact, AG1478, a specific inhibitor of the EGF receptor kinase, blocked both SDF-1alpha dependent proliferation and ERK1/2 activation. PMID- 15659795 TI - Downregulation of angiogenic factors in Ewing tumor xenografts by the combination of human interferon-alpha or interferon-beta with ifosfamide. AB - Ewing sarcoma is the second most common bone tumor in childhood. Despite aggressive chemotherapy and radiotherapy, the prognosis of metastatic disease remains poor. In a nude mouse model of Ewing tumor xenografts, we recently showed that human type I interferons (IFNs) inhibit the growth of established xenografts. Combined therapy with human IFNs and ifosfamide (IFO), an alkylating agent widely used in high-dose chemotherapy of Ewing tumors, results in a strong synergistic antitumor effect. We have investigated the effect of IFNs/IFO treatment on the expression of vascular endothelial growth factor (VEGF), matrix metalloproteinase 9 (MMP-9), and urokinase plasminogen activator receptor (uPAR), three key mediators of tumor growth and angiogenesis, in tumor xenografts generated either from a primary tumor (EW7) or from a metastatic tumor (COH). COH tumors expressed 5-fold higher levels of VEGF than EW7 tumors. IFNs/IFO treatment reduced by >70% the amount of VEGF in COH and EW7 tumors. We did not detect constitutive MMP-9 activity in EW7 tumors. In contrast, the metastasis-derived COH tumor expressed very high levels of active MMP-9. Although the total amount of MMP-9 remained unchanged, active MMP-9 was reduced by up to 75% in IFNs/IFO treated COH tumors. IFNs/IFO treatment triggered in both COH and EW7 tumors the downregulation of uPAR expression, a molecule involved in vascularization and endothelial cell migration. Our results partly explain the mechanism of tumor growth inhibition by IFNs/IFO therapy and provide a rational foundation for the development of a new therapeutic approach to Ewing tumors resistant to conventional chemotherapy. PMID- 15659796 TI - Radiation-induced alteration of pain-related signals in an animal model with bone invasion from cancer. AB - Although radiotherapy is highly effective in relieving bone pain from cancer invasion, the mechanism of pain relief remains unclear. To explore the mechanism of radiotherapy-induced analgesia, we have developed an animal model of bone pain resulting from cancer invasion. Using this animal model system, radiation-induced pain response and pain-related signals in the spinal cord were analyzed. The hind paw model of bone pain from cancer invasion was developed by injecting transplantable hepatocellular carcinoma, HCa-1, into the periosteal membrane of the foot dorsum in C3H/HeJ mice. Bony invasion from HCa-1 cells was confirmed by histopathological examinations. We also measured the development of pain associated behaviors. In this model, changes in the objective level of pain response after irradiation of the tumor were analyzed. Expression of pain-related host signals in the spinal cord, such as calcitonin gene-related peptide (CGRP), substance P, and c-fos, was investigated with immunohistochemical staining. In the histopathological examinations, bone invasion from HCa-1 cells was seen from day 7 and was evident at day 14 after injection. Measurable pain-associated behaviors were developed from day 7. In this model, mice treated with radiotherapy showed decreased objective levels of pain with a higher threshold to graded mechanical stimulation than did control mice from day 3 after irradiation. After irradiation of tumors, significant decreases in the expression of CGRP were shown in the spinal cord, whereas neither substance P nor c-fos showed any alteration. We developed a novel hind paw model of bone pain from cancer invasion that was confirmed by histopathological examination and measurable pain associated behaviors. Radiotherapy decreased the objective level of pain and the underlying mechanism involved in the alteration of pain-related host signal, CGRP, in the spinal cord. PMID- 15659797 TI - Analysis of the signal transduction pathway leading to human immunodeficiency virus-1-induced interferon regulatory factor-1 upregulation. AB - Interferon (IFN) regulatory factors (IRFs) constitute a family of transcriptional activators and repressors involved in the regulation of immune system, host defense, and cell growth. All members share conserved DNA-binding domains that recognize DNA sequences termed IRF-binding elements/IFN-stimulated response elements (IRF-E/ISRE) present on the promoter of IFN-alpha/beta and IFN stimulated genes. An ISRE has been identified downstream of the transcription start site of the long terminal repeat (LTR) of human immunodeficiency virus-1 (HIV-1). Our previous results showed that among the IRF factors, IRF-1 is able to stimulate HIV-1 LTR transcription and its expression is induced by HIV-1, early, upon infection and before the expression of Tat. In this study we investigated the signal transduction pathway leading to HIV-1-induced IRF-1 expression. Key IRF-1 promoter elements that mediate the activation of transcription upon induction by inflammatory cytokines are IFN-gamma-activated sequences that bind members of the signal transducer and activator of transcription (STAT) family and binding sites for nuclear factor kappaB (NF-kappaB). Both STAT-1 and NF-kappaB activation were examined to determine putative molecular targets whose inhibition resulted in the inhibition of HIV-1 replication. The results show that at early time points after HIV-1 infection, NF-kappaB but not STAT-1 is activated. Moreover, a significant decrease in HIV-1 replication was observed upon de novo infection of Jurkat T cells expressing an NF-kappaB super-repressor (IkappaB alpha 2NDelta4). These results suggest that in early phases of HIV-1 infection, before detectable cytokine production, NF-kappaB seems responsible for HIV-1 induced IRF-1 expression. PMID- 15659798 TI - Macrophage hemoglobin scavenger receptor and ferritin accumulation in human atherosclerotic lesions. AB - We previously proposed that erythrophagocytosis and iron metabolism by macrophages may contribute to iron-driven oxidative stress in atherogenesis. Recent studies have indicated that the macrophage hemoglobin scavenger receptor (HbSR/CD163) is a key molecule in the process of removing hemoglobin released from senescent erythrocytes. In this study we investigated erythrophagocytosis and its relation to ferritin accumulation and the involvement of CD163 in ferritin induction in human atheroma lesions. Normal and atherosclerotic human arterial segments obtained at autopsy and surgery were collected for iron histochemistry, hemoglobin and ferritin immunohistochemistry, and computerized image analysis. The lesion-dependent accumulation of ferritin and hemoglobin was seen in atherosclerotic carotid and coronary arteries. The immunoreactivity of hemoglobin was significantly correlated to the same regions of ferritin immunoreactivity on serial sections. The staining intensity of hemoglobin and ferritin was also significantly correlated. Hemoglobin deposition is often associated with microvessels adjacent to the lipid core areas in advanced lesions, where most CD68-positive macrophages were. CD163 expression appeared in both early and advanced lesions. The accumulation of tissue iron and ferritin also frequently occurs in CD163-positive and vessel-rich regions in the advanced atheroma. Although they were not always correspondingly positive on the serial sections, tissue iron and ferritin were significantly correlated. We conclude that erythrophagocytosis and hemoglobin catabolism by macrophages contribute to iron deposition and ferritin induction in human atheroma. The involvement of CD163 during ferritin induction may play an important role in modulating inflammatory processes in atherogenesis. PMID- 15659799 TI - Enhanced expression of natural resistance-associated macrophage protein 1 in atherosclerotic lesions may be associated with oxidized lipid-induced apoptosis. AB - The natural resistance-associated macrophage proteins (Nramps) can modulate inflammatory reactions. Nramps are not only responsible for intracellular divalent metal transport but also determine the macrophage functions in inflammatory processes. In the present study we tested whether Nramp1 is involved in macrophage apoptosis induced by oxidized lipids in atherogenesis. Arterial segments of Watanabe heritable hyperlipidemic rabbits were used for an examination of Nramp1 mRNA by in situ RT-PCR and macrophage immunohistochemistry. Annexin V/PI staining and terminal dUTP nick-end labeling (TUNEL) techniques were used for apoptosis detection. We found that, in macrophage-rich areas (positive to RMA-11) of the rabbit atherosclerotic aorta, there were lesion-dependent increases in Nramp1 mRNA, which are mainly apoptotic foamy macrophages that are positive to TUNEL staining. U937 cells were treated with 7beta-hydroxycholesterol (7beta-OH) in the presence or absence of the redox-active iron chelator desferrioxamine (DFO) or 1,10-phenanthroline. The cellular iron chelators considerably reduced, whereas iron compounds enhanced, 7beta-OH-induced apoptosis and necrosis. DFO also decreased mRNA levels of Nramp1, whereas both iron compounds and 7beta-OH dramatically enhanced the expression of Nramp1 mRNA, particularly among 7beta-OH-induced apoptotic cells. We conclude that the enhanced expression of Nramp1 in macrophage regions of atherosclerotic lesions may be associated with ferrous iron-enhanced, oxidized lipid-induced apoptosis. This finding reveals a novel function of Nramp1 in atherogenesis. PMID- 15659800 TI - Regulation of tumor cell motility by ERK mitogen-activated protein kinases. AB - Metastasis is the main cause of human cancer deaths, and cell motility is one key component of the process of metastasis. The ERK mitogen-activated protein kinase (MAPK) signaling pathway is probably one of the most commonly activated in malignant human cancers. We describe the molecular mechanisms by which ERK MAPK signaling regulates cell motility and its importance for cancer cell dissemination. PMID- 15659801 TI - Role of constitutively activated and insulin-like growth factor-stimulated ERK1/2 signaling in human hepatoma cell proliferation and apoptosis: evidence for heterogeneity of tumor cell lines. AB - Enhanced insulin-like growth factor II (IGF-II) and type I IGF receptor (IGF-IR) gene expression in liver tumors and the development of liver tumors in transgenic mice overexpressing IGF-II in the liver suggest that the IGFs and underlying signaling cascades may play auto/paracrine roles in the control of hepatocarcinoma (HCC) cell proliferation and in their protection against apoptosis. We have focused on the role of mitogen-activated protein kinase (ERK1/2) signaling on human HepG2 and Huh-7 hepatoma cell proliferation and on the protection of these cells against drug-induced apoptosis. Physiological concentrations of IGF-I stimulated DNA replication in HepG2 cells (1.5-fold) but not in Huh-7 cells, and this effect was abolished by PD98059 (MEK-1 inhibitor). Doxorubicin or cisplatin treatment induced apoptosis (caspase-dependent poly[ADP ribose]polymerase cleavage) in both cell lines, but dose-dependent reversion of drug-induced apoptosis (57-84%) by IGF-I was only observed in HepG2 cells. The very low level of IGF-IR at the plasma membrane of Huh-7 cells may account for their unresponsiveness to IGF-I. We have shown that drug treatment enhanced (17 fold) or did not modify constitutive ERK1/2 activity in cultured HepG2 or Huh-7 cells, respectively. In both cell lines, inhibition of constitutive and drug induced ERK1/2 activity by PD98059 yielded a complete inhibition of drug-induced apoptosis. Altogether, our data demonstrate the heterogeneous response of human hepatoma cells to an IGF stimulus and suggest (1) that auto/paracrine effects of IGF-I/-II might contribute to the proliferation of HCC cells and to their protection against apoptosis in vivo and (2) that drug-induced activation of ERK1/2 plays a role in drug-induced apoptosis in human hepatoma cells. PMID- 15659802 TI - Cooperative effects between protein kinase A and p44/p42 mitogen-activated protein kinase to promote cAMP-responsive element binding protein activation after beta cell stimulation by glucose and its alteration due to glucotoxicity. AB - Long-term hyperglycemia, a major characteristic of the diabetic state, contributes to the deterioration of the beta cell function, a concept known as beta cell glucotoxicity. We used the MIN6 beta cell line and isolated rat islets to clarify the signaling mechanism(s) used by glucose to activate cAMP-responsive element binding protein (CREB), a transcription factor crucial for beta cell biology, and to evaluate the possible downregulation of this mechanism mediated by long-term hyperglycemia. We report that glucose (10 mM) induces an increase in cytosolic calcium concentration that leads to cAMP-induced protein kinase A (PKA) activation, promoting nuclear translocation of activated ERK1/2. The observation that glucose-induced CREB phosphorylation was totally inhibited by the PKA inhibitor H89 (2 microM) and reduced by 50% with the ERK1/2 inhibitor PD98059 (20 microM) indicates that ERK1/2, located downstream of PKA, cooperates with PKA and is responsible for half of the PKA-mediated CREB phosphorylation elicited by glucose in MIN6 beta cells. We also found that exposure of mu cells for 24 h to high glucose (25 mM) induced a 70% decrease in cellular ERK1/2 and a 50% decrease in CREB content. In high-glucose-treated, ERK1/2- and CREB-downregulated beta cells, there was a loss of glucose (10 mM, 5 min)-stimulated ERK1/2 and CREB phosphorylation that was associated with nuclear apoptotic characteristics. Since we have shown that activation of ERK1/2 is crucial for CREB phosphorylation, loss of the ERK1/2-CREB signaling pathway in beta cells due to long-term hyperglycemia is likely to exacerbate beta cell failure in diabetic states by affecting physiologically relevant gene expression and by inducing apoptosis. PMID- 15659803 TI - p42/p44 Mitogen-activated protein kinase signal transduction pathway: a novel target for the treatment of hormone-resistant prostate cancer? AB - Prostate cancer is the second leading cause of cancer deaths in men. Conventional therapies produce a high rate of cure for patients with localized prostate cancer, but there is no cure once the disease has spread beyond the prostate. Androgen withdrawal remains the only treatment for these men with clinically advanced disease; however, most of these men, who initially respond to hormone ablation therapy, fail and the disease progresses. There is at present no effective treatment for hormone-independent prostate cancer. Several lines of evidence suggest a role of p42/p44 mitogen-activated protein kinase (p42/p44 MAP kinase) signal transduction pathways in prostate cancer. At the molecular level, a variety of genetic alterations lead to an epigenetic mechanism by which a feedback autocrine loop between membrane receptors and associated ligands serves as an essential component of the growth, proliferation, and metastasis of prostate cancer at an advanced and androgen-independent stage. Peptide growth factors are known to exert their effects by a complex array of mechanisms primarily mediated by the p42/p44 MAP kinase signal transduction pathway. Thus, we hypothesized that MAP kinase signal transduction pathways could serve as new and novel targets in prostate cancer therapy. In this article we provide an overview of the role played by MAP kinase signal transduction in the prostate. PMID- 15659804 TI - p42/p44 Mitogen-activated protein kinase signal transduction pathway regulates interleukin-6 expression in PC3 cells, a line of hormone-refractory prostate cancer cells. AB - The mechanisms that regulate prostate cancer growth and proliferation are not fully understood. IL-6, a multifunctional cytokine, has been shown to play an important role in prostate cancer biology. Functional role of MAP-kinase signal transduction pathways in prostate biology has not been evaluated in detail. In the present study we evaluated the effects of modulation of p42/44 MAP kinase signal transduction pathway on IL-6 expression and secretion by PC3 cells, a line of hormone refractory prostate cancer cells. Results presented, herein, demonstrate that modulation of p42/44 MAP kinase activity results in partial inhibition of synthesis and secretion of IL-6. These data suggest that modulation of p42/p44 may result in regulation of other survival pathways as well. PMID- 15659805 TI - p38 is a key signaling molecule for H-ras-induced inhibition of gap junction intercellular communication in rat liver epithelial cells. AB - Multiple lines of evidence indicate that inhibition of gap junction intercellular communication (GJIC) is a major carcinogenic process. Several reports suggest that activation of extracellular signal-regulated protein kinases 1 and 2 (ERK1/2) plays a key role in the disrupted GJIC by hydrogen peroxide, 12-O tetradecanoylphorbol-13-acetate, and quinones in rat liver epithelial cells. Recently, we reported that p38 mitogen-activated protein kinase (MAPK) is also involved in the inhibition of GJIC by hydrogen peroxide in WB-F344 rat liver epithelial cells (WB cells). The present study investigated the role of ERK1/2 and p38 MAPK in H-ras-induced inhibition of GJIC in WB cells. H-ras induces complete inhibition of GJIC and unphosphorylation of connexin 43 (Cx43) in WB cells. SB203580, an inhibitor of p38, restored inhibition of GJIC and blocked unphosphorylation of Cx43 in H-ras-transformed WB cells. However, PD98059, an inhibitor of ERK1/2, had no effect. Our results suggest that the disruption of GJIC induced by H-ras may be strongly related to the unphosphorylation of Cx43 via the activation of p38 but not ERK1/2. Thus, p38 is a key signaling molecule for H-ras-induced inhibition of GJIC in WB cells. PMID- 15659806 TI - The phosphotyrosine phosphatase eta mediates somatostatin inhibition of glioma proliferation via the dephosphorylation of ERK1/2. AB - Somatostatin (SST) controls the proliferation of a variety of cell types. Its effects are mediated by five G protein-coupled receptors (SSTR1-SSTR5), variably expressed in normal and cancer tissues. SST inhibition of cell proliferation can be exploited by both direct and indirect mechanisms: the main direct pathway involves the modulation of phosphotyrosine phosphatase (PTP) activity. Here we show that SST cytostatic activity is mediated by the activation of a receptor like PTP, named PTPeta. The role of this PTP in the antiproliferative activity of SST in five glioma cell lines (C6, U87MG, U373MG, DBTRG05MG, and CAS1) and in four postsurgical human glioblastoma specimens, has been studied. SST inhibited growth only in C6 and U87MG that express PTPeta. In C6 cells, SST antiproliferative effects were reverted by pretreatment with pertussis toxin and vanadate, indicating the involvement of G proteins and PTPs. The role of PTPeta in the SST inhibitory effects was demonstrated by testing the PTPeta activity: it was increased by SST treatment and paralleled by inhibition of ERK1/2 activation. Since basic fibroblast growth factor-dependent MEK phosphorylation was not affected by SST, we propose a direct effect of SST-activated PTPeta on ERK1/2 phosphorylation. Finally, the SSTR mRNAs were identified in all of the 36 gliomas analyzed, whereas PTPeta expression was found in 33% of cases. Culturing four gliomas, a precise correlation between the expression of PTPeta and the SST antiproliferative effects was identified. In conclusion, in glioma cells, SST antiproliferative activity requires the expression and activation of PTPeta, which directly dephosphorylates ERK1/2. PMID- 15659808 TI - Functional characterization of amyloid beta precursor protein regulatory elements: rationale for the identification of genetic polymorphism. AB - Alzheimer's disease (AD) is characterized by the formation of senile plaques of the amyloid peptide (Abeta) derived from a large Abeta precursor protein (APP). Autosomally inherited or "familial" AD has only been previously demonstrated in connection with coding sequence missense mutations. Abnormal regulation of APP gene expression has been demonstrated to play a role in AD. Genome screen and linkage analysis suggest that the APP locus may predispose to AD. The aim is to characterize genetic variability in the APP gene within its upstream regulatory region and to determine whether that variability is associated with AD and affects the expression of APP. This article describes the rationale and strategy for identifying genetic polymorphisms in the APP regulatory region, including its promoter, to associate any variability with the disease. PMID- 15659807 TI - Blockade by SB203580 of Cyp1a1 induction by 2,3,7,8-tetrachlorodibenzo-p-dioxin, and the possible mechanism: possible involvement of the p38 mitogen-activated protein kinase pathway in shuttling of Ah receptor overexpressed in COS-7 cells. AB - A transiently overexpressed aryl hydrocarbon receptor (AhR) became translocated into the nucleus of COS-7 cells without treatment with any ligand, such as 2,3,7,8-tetrachlorodibenzo-p-dioxin. This spontaneous AhR translocation into the nucleus was reduced by pretreatment of the recipient cells with the p38 mitogen activated protein (MAP) kinase inhibitor SB203580. Immunofluorescent microscopic analysis revealed that SB203580 treatment increased the fluorescence intensity of AhR within the cytoplasm. An analogue compound, SB202474, which does not inhibit p38 MAP kinase, did not reduce AhR translocation into the nucleus. Moreover, a reporter gene assay showed that the AhR spontaneously translocated into the nucleus activated reporter gene transcription and that SB203580 suppressed its transcriptional activity. From these data we conclude that the p38 MAP kinase pathway is involved in determining AhR cellular localization in COS-7 cells overexpressing AhR. PMID- 15659809 TI - Increased intracellular ionic content is correlated with a decreased perichromatin granule density in old neurons. AB - The intracellular content of monovalent ions (Na(+), Cl(-), and K(+)) in brain cortical cells from young, adult, and old rats was measured by X-ray microanalysis. By referring the peak of each ion to the respective value of the dry mass, we obtained the intracellular content as a percentage of dry mass. A significant increase of the intracellular content of Na(+), Cl(-), and K(+) was found in old animals. In the same type of cells from rats of different ages, we estimated the density of perichromatin granules (PGs), the RNA structural correlates containing ribonucleoproteins (RNPs). PGs were discovered by Bernhard's method, which allows the preferential staining of RNPs. The PG density (i.e., the number of PGs/microm2 of nuclear area) was significantly lower ( 27.8%) in the neurons of old animals. Taken together, the present findings document that in the nuclei of brain cortical cells of old animals the ionic strength is markedly increased and that this alteration may affect chromatin functions (e.g., DNA template activity). The ionic content of the nucleoplasm is reported to modulate the structure of chromatin molecules and regulate gene expression; thus, the increased content of the three ions found by us in old animals, by increasing the viscosity of the nucleoplasm, may affect gene expression by disturbing the inhibition (negative regulation) or potentiation (positive regulation) of RNA polymerase binding. In physiological aging, increased ionic strength may lead to age-related DNA dysfunction as well as to alterations in the processing of RNA structural constituents, such as PGs. PMID- 15659810 TI - Lack of N1L gene expression results in a significant decrease of vaccinia virus replication in mouse brain. AB - Vaccinia virus encodes secretory proteins termed virokines. One of the major virokines encoded by the N1L open reading frame is the 13.8 kDa protein. A recombinant virus, termed vGK5, lacking this protein when injected intracranially into mice, has one of the highest levels of in vivo attenuation achieved by deletion of any single open reading frame of vaccinia virus. Here we show that the 13.8 kDa protein significantly enhances viral replication within brain tissue; however, analysis of histology, neutrophil infiltrate, and nitric oxide synthase activity of brain tissue shows no significant differences between wild type vaccinia virus and vGK5. Since there is poor growth of vGK5 virus in the brain, the possibility of postvaccinial encephalitis is significantly diminished. Mice injected with vGK5 became resistant to the lethal effects of vaccinia virus, indicating that vGK5 is immunogenic in the brain without being virulent and therefore is a vaccine candidate. This suggests that should vGK5 reach the brain it will not replicate efficiently but still serve as a live vaccine. PMID- 15659811 TI - Increased mitochondrial and nuclear gene expression of cytochrome oxidase subunits I and IV in neuronal aging. AB - To assess the role of mitochondrial metabolic competence (MMC) in neuronal aging, quantitative immunohistochemistry of cytochrome oxidase (COX) subunits I (mitochondrial-encoded) and IV (nuclear-encoded) was carried out in the cerebellar cortex of adult and old rats. The optical density (OD) values of the immunostained COX subunits I and IV were measured on an overall area of 75,000 microm(2) in the granular and molecular layers of the cerebellar cortex of each animal. In old animals, OD values of subunit I were increased by 35.5 and 34.2% in the molecular and granular layers, respectively, but only the difference found in the latter cerebellar zone was statistically significant (p < 0.05%). As regards subunit IV, old animals showed higher, not significant, densitometric values in the molecular (120.6%) and granular (126.8%) layers. The present findings sustain that gene expression of COX subunits I and IV appears not to be involved in the well-documented time-related mitochondrial decay. The proper functioning of COX depends on several factors that can affect MMC in the aging cell. In the fully assembled holoenzyme, both the subunits I and IV span the inner mitochondrial membrane. On the basis of these molecular biology data, it is reasonable to suppose that any alteration of the physicochemical features and chemical composition of the mitochondrial membranes reported to occur in aging (e.g., decreased membrane fluidity and cardiolipin content, increased cholesterol/phospholipid molar ratio and free-radical damage, etc.) may significantly affect the proper assembling of the enzyme and, in turn, its activity. Considering the reported significant decline of COX activity with advancing age, our findings further support that an adequate mitochondrial metabolic competence, while including proper nuclear and mitochondrial gene expression of subunits of the respiratory chain, relies on the overall balance among various determinants that can be differently damaged by aging and represent critical causative events responsible for the age-related functional decline of selected mitochondrial populations. PMID- 15659812 TI - Role of the APP promoter in Alzheimer's disease: cell type-specific expression of the beta-amyloid precursor protein. AB - One of the major hallmarks in Alzheimer's disease (AD) is amyloid deposition in the brain of afflicted subjects. This tissue-specific deposition of the amyloid beta-protein (Abeta) is the major characteristic of AD. Abeta is proteolytically derived from a large Abeta precursor protein (APP). An apparent overexpression of the APP gene in certain areas of the AD brain indicates that abnormalities in gene regulation might be an important factor in AD pathology. The mechanism of expression of APP in different cell types is poorly understood. To understand the contribution of different cell types, such as neuronal, glial, and epithelial cells, APP expression was studied at the message and protein levels. Levels of APP expression, both message and protein, were greater in human neuroblastoma (NB) and PC12 cells than in glial and HeLa cells. DNA transfection experiments suggest that the relative activities of different promoter regions varied according to cell type. Although the upstream regulatory element in the promoter region is necessary for activity in PC12 and HeLa cells, this is not the case for NB cells. A 30-bp proximal promoter region was found to be important for cell type-specific APP gene expression. PMID- 15659813 TI - Protective effects of oligomers of grape seed polyphenols against beta-amyloid induced oxidative cell death. AB - beta-Amyloid (Abeta) is considered to be responsible for the formation of senile plaques that accumulate in the brains of patients with Alzheimer's disease (AD). There is compelling evidence supporting the notion that Abeta-induced cytotoxicity is mediated through the generation of reactive oxygen species (ROS). Recently, considerable attention has been focused on a wide array of non-vitamin antioxidants present in edible plants that are able to scavenge ROS, thereby protecting against oxidative damage. In this study, we have investigated the possible protective effects of formulated polyphenol oligomers (Oligonol) derived from grape seed extracts on Abeta-induced oxidative cell death. Rat pheochromocytoma (PC12) cells treated with Abeta exhibited increased accumulation of intracellular ROS and underwent apoptosis, as determined by positive in situ terminal end labeling, decreased mitochondrial membrane potential, and the cleavage of poly(ADP-ribose)polymerase. Oligonal attenuated Abeta-induced cytotoxicity, apoptotic features, intracellular ROS accumulation, and lipid peroxidation and increased the cellular glutathione pool. Moreover, Abeta transiently induced the activation of nuclear factor kappaB in PC12 cells, which was suppressed by pretreatment with Oligonol. PMID- 15659815 TI - Apoptotic cell death and amyloid precursor protein signaling in neuroblastoma SH SY5Y cells. AB - We have recently shown that the amyloid precursor protein (APP) and a subset of its C-terminal fragments (CTFs) are tyrosine phosphorylated in human brain and in cultured cells. Tyrosine phosphorylation generates a substrate that is sequentially bound by the adaptor proteins ShcA and Grb2, and this interaction is significantly enhanced in Alzheimer's disease brains. Here we have studied the APP/CTFs phosphorylation and ShcA activation in a human neuroblastoma cell line, SH-SY5Y, under basal and apoptotic conditions. To commit these cells to apoptosis, we used staurosporin, a well-known apoptotic inducer and protein kinase C blocker. Our data suggest the following: (1) in normally proliferating SH-SY5Y cells, full-length APP is complexed with Grb2[Q3], likely through its SH2 domain; (2) upon induction of apoptosis, APP is degraded and ShcA-Grb2 coimmunoprecipitates with CTFs recognized by anti-APP antibodies; and (3) caspase inhibitors partially block the degradation of APP and the coprecipitation of CTFs with ShcA-Grb2 adaptors. In summary, our data suggest that in SH-SY5Y cells, tyrosine-phosphorylated APP is involved in a complex with ShcA-Grb2 adaptors that is disrupted during apoptosis. The abnormal degradation of APP and consequent increased levels of CTFs (as has been observed in Alzheimer's disease and Down's syndrome) generate a complex between tyrosine-phosphorylated CTFs and intracellular adaptors. The signaling through APP and its CTFs may have significant relevance for apoptotic cell death in Alzheimer's disease. PMID- 15659814 TI - BACE1 overexpression regulates amyloid precursor protein cleavage and interaction with the ShcA adapter. AB - The amyloid precursor protein (APP) is a cell surface protein with a large extracellular N-terminal domain, a single transmembrane segment, and a short cytoplasmic tail. Its location and structural features are characteristic of a receptor for signal transduction. Yet, the physiological function of APP is unclear, although it is well documented that APP's proteolytic processing, through the formation of membrane-bound C-terminal fragments (CTFs) and of beta amyloid peptides, likely influences the development of Alzheimer's disease (AD). There is evidence that BACE1 is the enzyme responsible for beta-site cleavage of the APP and for the generation of CTFs. BACE1 expression is upregulated in AD brain, and we have recently shown in human brain and in vitro that BACE product CTFs, when phosphorylated in tyrosine residues, interact with the adaptor proteins ShcA and Grb2, which usually are involved in signal transduction pathways. We investigated the interaction between ShcA, APP, and CTFs in the H4 human cell line that overexpresses BACE1 to clarify the significance of such interactions in vitro and for AD generation. Our result show that the APP, CTF, and ShcA interaction is induced only upon overexpression of BACE1 either transiently or in stable cell lines. In particular, although BACE1 drives the formation of C99 and C89 CTFs, only C99 interacts with the ShcA adaptor protein. Therefore, our data suggest that BACE1 activity influences APP processing and its intracellular signaling through the ShcA adaptor protein. PMID- 15659816 TI - Effect of metal complexes on thioredoxin reductase and the regulation of mitochondrial permeability conditions. AB - Gold(I) compounds such as auranofin, chloro(triethylphosphine) gold(I), and aurothiomalate act on mitochondrial functional parameters by determining an extensive permeability transition and a decrease of membrane potential. On the contrary, pyridine nucleotides and glutathione are not modified, whereas a slight but significant decrease of total thiols is apparent. The effect of gold(I) compounds is essentially referable to the inhibition, in the nanomolar range, of thioredoxin reductase activity and to an increase of hydrogen peroxide production. Metal ions and metal complexes (zinc and cadmium acetate, cisplatin, tributyltin) are also good inhibitors of thioredoxin reductase, although in the micromolar range, and in addition, they act as inducers of permeability transition and of membrane potential decrease. At variance with gold(I) compounds, which appear to work almost exclusively on thioredoxin reductase, metal ions and complexes are less specific, since they are active on different mitochondrial targets, including the respiratory chain. PMID- 15659817 TI - The antiapoptotic protein ICBP90 is a target for protein kinase 2. AB - Protein kinase 2 (casein kinase 2 [CK2]) is a protein serine/threonine kinase involved in cell proliferation with an expression that is dysregulated in tumors. ICBP90, a transcription factor exhibiting antiapoptotic properties, has several putative CK2 phosphorylation sites. The aim of the present study was to investigate whether ICBP90 could behave as a CK2 substrate. We observed that ICBP90 was more efficiently phosphorylated by the free CK2a subunit than by the heterotetrameric CK2 (alpha(2), beta(2)). Our results suggest that CK2 is an important regulator of the transcriptional activity of ICBP90 and therefore of the antiapoptotic properties of ICBP90. We propose that the "ICBP90 family" members may be substrates for CK2. PMID- 15659818 TI - Docosahexaenoic acid from a cultured microalga inhibits cell growth and induces apoptosis by upregulating Bax/Bcl-2 ratio in human breast carcinoma MCF-7 cells. AB - Docosahexaenoic acid (DHA) is an omega-3 fatty acid that comprises 22 carbons and 6 alternative double bonds in its hydrocarbon chain (22:6omega3). Previous studies have shown that DHA from fish oil controls the growth and development of different cancers; however, safety issues have been raised repeatedly about contamination of toxins in fish oil that makes it no longer a clean and safe source of the fatty acid. We investigated the cell growth inhibition of DHA from the cultured microalga Crypthecodinium cohnii (algal DHA [aDHA]) in human breast carcinoma MCF-7 cells. aDHA exhibited growth inhibition on breast cancer cells dose-dependently by 16.0% to 59.0% of the control level after 72-h incubations with 40 to 160 microM of the fatty acid. DNA flow cytometry shows that aDHA induced sub-G(1) cells, or apoptotic cells, by 64.4% to 171.3% of the control levels after incubations with 80 mM of the fatty acid for 24, 48, and 72 h. Western blot studies further show that aDHA did not modulate the expression of proapoptotic Bax protein but induced the downregulation of anti-apoptotic Bcl-2 expression time-dependently, causing increases of Bax/Bcl-2 ratio by 303.4% and 386.5% after 48- and 72-h incubations respectively with the fatty acid. Results from this study suggest that DHA from the cultured microalga is also effective in controlling cancer cell growth and that downregulation of antiapoptotic Bcl-2 is an important step in the induced apoptosis. PMID- 15659819 TI - Plasma membrane alteration is an early signaling event in doxorubicin-induced apoptosis in the yeast Candida utilis. AB - Signaling pathways such as increased ceramide, mitochondrial dysfunction, and P3 and caspase activation are produced by anticancer drugs and lead to apoptosis. In this research we show that the first event after culturing the yeast Candida utilis in the presence of low doses of doxorubicin (25 microg/mL) is the morphological alteration of the plasma membrane. In the presence of higher doxorubicin doses (>/=50 microg/mL), in addition to profound alterations in the plasma membrane, changes in mitochondrial shape and cristae organization were observed. Concomitantly, increases in respiration, substrate oxidation, and cytochrome biosynthesis were observed at low doxorubicin doses (up to 25 microg/mL), whereas a progressive decrease was observed at higher doses. [(3)H]Leu incorporation into proteins increased by 40% in the mitochondrial fraction and by 19% in the cytosol in the presence of 25 microg/mL doxorubicin; it decreased to 80% of the control in the cytosol in the presence of 1 mg/mL doxorubicin. Morphologically, doxorubicin doses of up to 200 microg/mL produced apoptosis, whereas higher doxorubicin doses produced necrosis. PMID- 15659820 TI - Expression of the insulin-like growth factor-I receptor and proapoptotic Bax and Bak proteins in human colorectal cancer. AB - Insulin-like growth factor-I (IGF-I) and IGF-I receptor (IGF-IR), despite their well-known roles in cell survival and proliferation, can also weakly enhance apoptosis. To study the relationships between the IGF-IR and Bax as well as Bak, 144 cases of colorectal cancer were examined by immunohistochemistry, using the avidin-biotin-peroxidase method. The results were correlated with selected clinicopathological features of colorectal cancer and with the expression of IGF IR, Bax, and Bak in normal colon mucosa. In Bax-, Bak-, or IGF-IR-positive cancers, the adjacent colorectal mucosa revealed positive immunostaining for these proteins. In the majority of Bax-, Bak-, or IGF-IR-negative tumors, we observed no staining for these proteins in adjacent mucosa. The strong immunostaining for IGF-IR, Bax, and Bak was noted in 50.8, 55.5, and 49.3% of tumors, respectively. We observed positive correlations between IGF-IR and Bax (P < 0.002, r = 0.302), between IGF-IR and Bak (P < 0.0001, r = 0.407), and between Bax and Bak (P < 0.0001, r = 0.474). No relationship was noted between IGF-IR expression and tumor grade, stage, or lymph node status. We found negative associations between Bax, Bak, and tumor grade (P < 0.01 and P < 0.003, respectively), but no relationships between Bax and Bak and tumor stage or between Bax and Bak and lymph node status. Our results demonstrate that, in addition to overexpressed IGF-IR, there are relationships between IGF-IR and proapoptotic proteins in colorectal carcinomas that could contribute to increased cell turnover and the progression of colorectal cancer. PMID- 15659822 TI - Alterations in protein expression in HL-60 cells during etoposide-induced apoptosis modulated by the caspase inhibitor ZVAD.fmk. AB - DNA topoisomerase inhibitors induce a specific signaling cascade that promotes an active apoptotic caspase-dependent cell death process. However, little is known about the initial signals elicited by these agents. In the present study, we compared apoptosis in HL-60 cells treated either with the chemotherapeutic drug etoposide (VP16) alone or combined with the broad caspase inhibitor ZVAD.fmk. Apoptosis was assessed by changes in cell morphology and agarose gel electrophoresis of extracted cell DNA. We found that ZVAD.fmk prevents VP16 induced DNA fragmentation and the appearance of an increased number of apoptotic cells in the culture. We also compared the effects of etoposide alone or together with the pan-caspase inhibitor ZVAD.fmk on proliferating cell nuclear antigen, Bcl-2, and actin expression in human promyelocytic leukemia HL-60 cells. In addition, we screened for proteins that were initially upregulated in a caspase dependent manner. Indeed, some proteins were induced in the cytoplasm and subsequently accumulated in the nuclei after etoposide treatment. This process was slightly inhibited by the caspase inhibitor ZVAD.fmk. We suggest that these proteins are associated with the induction of specific signaling cascades that characterize the apoptotic cell death process. PMID- 15659821 TI - Inhibition of B16 mouse melanoma cell growth and induction of apoptotic cell death with 8-chloroadenosine-3',5'-monophosphate and tiazofurin. AB - Novel antineoplastic agents, 8-chloroadenosine 3',5'-monophosphate (8-Cl-cAMP) and tiazofurin (TR), have been shown to be effective against different malignant cells. Through specific mechanisms of action they modulate the cellular signal transduction pathway, thereby causing growth inhibition, cell differentiation, and apoptosis. The aim of this work was the in vitro study of either 8-Cl-cAMP or TR effects on B16/F10 and B16/C3 mouse melanoma cell growth and cell death. Significant cell growth inhibition was obtained after the application of 8-Cl cAMP or TR. The presence and number of apoptotic cells was evaluated using agarose gel electrophoresis and flow cytometry. The number of apoptotic nuclei, after treatment with antineoplastic agents, did not significantly change in B16/F10 cells, although it did show a significant increase in B16/C3 cells. The expression of c-myc did not significantly change in B16/F10 cells after treatment with 8-Cl-cAMP or TR. The same results were obtained in B16/C3 cells after treatment with 8-Cl-cAMP. The level of c-myc expression showed a significant increase in B16/C3 cells after treatment with TR. Concerning the effects that the analyzed agents exhibited on melanoma cells and other cancer cells, further preclinical studies of these drugs will potentially lead to better understanding of the molecular mechanisms of their action and finally more efficient therapeutic approaches to malignant diseases. PMID- 15659823 TI - Mitochondria of human spermatozoa are preferentially susceptible to apoptosis. AB - Although elements of type I and II apoptosis have been demonstrated in human spermatozoa, their functionality has not been evaluated. The first dynamic studies revealed no type I apoptosis signal transduction via CD95. The aim of our study was to clarify whether type II apoptosis can be induced in human spermatozoa. Betulinic acid, a cytotoxic agent and highly specific inductor of type II apoptosis, acts by a direct effect on mitochondria. Motility, mitochondrial transmembrane potential, and active caspase-9 and -3 were examined in human ejaculated spermatozoa of 33 semen samples from healthy volunteers after incubation with 60 microg/mL betulinic acid for 10 and 60 min, respectively. Untreated aliquots of each sample served as negative controls. Treatment with betulinic acid resulted in the induction of type II apoptosis measured by disruption of mitochondrial transmembrane potential and activation of caspase-9 and -3. The loss of mitochondrial energy supply resulted in a significant decrease of spermatozoal motility and velocity. In spermatozoa, mitochondria are tightly packed and located exclusively in the midpiece region. This might contribute to their susceptibility against the induction of type II apoptosis and should be considered for therapeutic interventions and might have a future in the development of advanced birth control methods. PMID- 15659824 TI - Proapoptotic potentials of genistein under growth stimulation by estrogen. AB - In mammary carcinogenesis, hormonal effects have been reported to be important factors. Estrogens are known to regulate the proliferation of breast cancer cells, whereas genistein has been shown to induce apoptosis in mammary tumor cells. This study examined genistein-induced apoptosis through the regulation of bcl-2 and bax expression in the presence of estrogen. MCF-7 cells were treated with either genistein (25, 50, and 100 microM) or in the presence of 17beta estradiol (12.5, 25, and 50 nM) for 48 h. DNA ladder analysis and Western blot analysis of bcl-2, bax, cyclin B(1), p21, and p53 were carried out. For comparison, the in vivo system was employed using estrogen-deficient and estrogen sufficient female rats at two different concentrations of genistein. In MCF-7 cells, DNA fragmentation was evident by the treatment of genistein in the absence and presence of estrogen. Downregulation of bcl-2 and upregulation of bax by genistein were observed. However, genistein showed no proapoptotic properties in the presence of estrogen except with the lowest concentration of estrogen. In the presence of estrogen, p21 and p53 protein expression were upregulated by high concentrations of genistein. Bcl-2/bax ratios were decreased by genistein treatment in the presence or absence of estrogen in female rats. These results demonstrate that the proapoptotic property of genistein might be influenced greatly by the concentration of estrogen in vitro, but that this influence by estrogen is not evident in vivo. PMID- 15659825 TI - Expression of cell proliferation and apoptosis markers in papillomas and cancers of conjunctiva and eyelid. AB - Cell proliferation and programmed cell death are considered to be important events in carcinogenesis. The object of our study was to evaluate the expression of the Bcl-2 protein family (Bcl-2, Bak, Bax), p53, proliferating cell nuclear antigen (PCNA), and Ki-67 protein immunoexpression as well as the correlation between the examined markers and some clinicopathological features in papillomas and cancers of conjunctiva and eyelid. Forty-five squamous cell papillomas (SCP), 11 squamous cell cancers (SCC), and 27 basal cell cancers (BCC) were estimated. In the SCP group, p53 protein expression was observed in 30 cases (66.6%), Ki-67 in 14 (31.1%), PCNA in 44 (97.8%), Bcl-2 in 24 (53.3%), Bak in 28 (62.2%), Bax in 31 (68.9%), and Bcl-xl in 11 (100%). In the SCC group, p53 protein expression was evaluated in 8 cases (72.8%), Ki-67 in 2 (18.2%), PCNA in 8 (72.7%), Bcl-2 in 5 (45.4%), Bax and Bak both in 10 (90.9%), and Bcl-xl in 100%. In the BCC group, p53 protein expression was estimated in 23 cases (85.1%), Ki-67 in 13 (48.1%), PCNA in 26 (96.2%), Bcl-2 in 13 (48.1%), Bak in 21 (77.8%), Bax in 22 (81.5%), and Bcl-xl in 23 (85.2%). We observed a correlation between some clinicopathological features and the examined markers of apoptosis and cell proliferation, which seemed to be important events in cancer development. PMID- 15659826 TI - Increased expression and translocation of lysosomal cathepsins contribute to macrophage apoptosis in atherogenesis. AB - It has been recently reported that atherosclerotic lesions in both humans and mice express several lysosomal proteases, including cathepsins B, D, L, and S, which may affect plaque development and stability. The mechanisms responsible for the extralysosomal expression of lysosomal cathepsins and the related atherogenic implications remain unknown. We find that the lesion-dependent expression of cathepsins B and L is mainly in macrophage-infiltrated areas of human carotid atheroma. These enzymes appear in the cytoplasm and nuclei of apoptotic macrophages (normally confined to the lysosomal compartment) and in the extracellular areas. After exposure to oxidized low-density lipoprotein (oxLDL) or 7beta-hydroxycholesterol (7beta-OH), macrophages initially transform into foam cells and then undergo apoptotic cell death. The oxidized lipids induce lysosomal destabilization, with leakage to the cytosol of lysosomal enzymes (cathepsins B, D, and L), as detected by cytochemistry and immunocytochemistry. A remarkable increase in cathepsin D mRNA levels was observed after 7beta-OH exposure. Like macrophages within atheroma, intralysosomal cathepsins B and L are translocated to the cytoplasm and nuclei of 7beta-OH-exposed cells. Our results suggest that endocytosed oxLDL and oxysterols not only destabilize the acidic vacuolar compartment but also cause the upregulation and translocation of lysosomal cathepsins, which may act as cleaving enzymes during the apoptotic process. The increased macrophage apoptosis and nuclear and matrix degradation by lysosomal enzymes in atheroma may play important roles in plaque development and rupture. PMID- 15659827 TI - Suppression of the nuclear factor-kappaB activation pathway by spice-derived phytochemicals: reasoning for seasoning. AB - The activation of nuclear transcription factor kappaB has now been linked with a variety of inflammatory diseases, including cancer, atherosclerosis, myocardial infarction, diabetes, allergy, asthma, arthritis, Crohn's disease, multiple sclerosis, Alzheimer's disease, osteoporosis, psoriasis, septic shock, and AIDS. Extensive research in the last few years has shown that the pathway that activates this transcription factor can be interrupted by phytochemicals derived from spices such as turmeric (curcumin), red pepper (capsaicin), cloves (eugenol), ginger (gingerol), cumin, anise, and fennel (anethol), basil and rosemary (ursolic acid), garlic (diallyl sulfide, S-allylmercaptocysteine, ajoene), and pomegranate (ellagic acid). For the first time, therefore, research provides "reasoning for seasoning." PMID- 15659828 TI - Curcumin stability and its effect on glutathione S-transferase P1-1 mRNA expression in K562 cells. AB - To investigate the stability of curcumin in physiological media, the absorption variation of a curcumin solution was measured in 0.1% and 10% FCS. Under daylight conditions, curcumin degraded very rapidly in 0.1% FCS and was found to be more stable in higher serum concentrations. Under dark conditions, almost no decomposition could be observed after 2 h, whether the measurements were performed in 0.1% or 10% FCS. Furthermore, depending on the medium concentration, differential glutathione S-transferase P1-1 mRNA expression could be observed in K562 cells after incubation with curcumin. Indeed, incubation in 0.1% FCS led to a decrease of mRNA expression, whereas incubation in 10% FCS induced an increase of mRNA production. PMID- 15659829 TI - Hypoxia/anoxia as signaling for increased alcohol dehydrogenase activity in saffron (Crocus sativus L.) corm. AB - Alcohol dehydrogenase, NAD-dependent lactate dehydrogenase, and NAD-independent lactate dehydrogenase activities were investigated in corms cultivated in normoxic and hypoxic/anoxic conditions. Depending on the developmental stage, hypoxia/anoxia was a signal for increase in either alcohol dehydrogenase or NAD dependent lactate dehydrogenase. NAD-independent lactate dehydrogenase contributed to the recycling of lactate, thus preventing acidosis. PMID- 15659830 TI - Prevention against oxidative stress of eukaryotic cell membranes by selenium compounds of the rat. AB - By a combination of trace techniques and various biochemical methods, information about the characteristics of a 15-kDa selenoprotein was obtained. After labeling of rats in vivo with [(75)Se]selenite, subcellular fractionation of the homogenates of the prostate, lung, brain, thyroid gland, and large intestine, and gel electrophoretic separation of the proteins and subcellular fractionation, 15 kDa (75)Se was found in the cytosols of the tissues prostate > brain > lung > thyroid gland > large intestine after autoradiography. After coelectrophoresis of the separated 15-kDa labeled band obtained from each cytosolic fraction, the 15 kDa (75)Se band migrated in the same way as the combined bands isolated from the five tissue cytosols. After proteolytic cleavage in the gel of the 15-kDa labeled band obtained from the cytosol of each tissue and re-electrophoresis, the same labeled peptide pattern was found in each gel slice after autoradiography. By means of reversed-phase HPLC, we characterized a selenocysteine-containing protein that has enzymatic activity like that of glutathione peroxidase. PMID- 15659831 TI - Effect of dietary restriction on DNA synthesis in vitamin E-deficient rats. AB - To assess the effect of dietary restriction on increased oxidative stress conditions, we measured the proliferative response of spleen lymphocytes from the following groups of adult rats: (1) control fed ad libitum (14 months of age); (2) vitamin E-deficient (12 months of age); and (3) vitamin E-deficient maintained on dietary restricted paradigm, that is, every other day schedule (12 months of age) animals. No significant change was observed among the three groups investigated at 24 h. At 48 h, [(3)H]thymidine incorporation was significantly lower in vitamin E-deficient rats vs. the other groups at Con A concentrations of 1 and 5 mug/mL, while at Con A concentration of 10 mug/mL the incorporation of the labeled compound in lymphocytes was significantly lower than only the vitamin E-deficient rats vs. controls. At 72 h: nonstimulated lymphocytes from ad libitum fed control rats showed significant higher values of [(3)H]thymidine incorporation vs. the other groups; no significant difference was found among the three groups investigated at 1 and 10 mug/mL Con A concentrations, while at 5 mug/mL Con A concentration, the lymphocytes from vitamin E-deficient rats showed a significant lower value of [(3)H]thymidine incorporation vs. the other groups. These data support that vitamin E-deficiency impairs the proliferative response of spleen lymphocytes from adult rats, while dietary restriction appears to be able to reverse this alteration. Although the mechanism(s) of action of dietary restriction in prolonging the life span and ameliorating health conditions are not know, it is currently supported that a reduced food intake results in a better control of free radical attacks to biological molecules as well as to several cellular and system functions. With specific reference to the present findings, dietary restriction may help the mitotic process dynamics to be accomplished in a condition of low rate of free radical damage, thus representing a physiological intervention capable of modulating positively the proliferative capacity of spleen lymphocytes and, in turn, the immune system, even in adverse conditions such as increased oxidative stress. PMID- 15659832 TI - Alternatively spliced protein variants as potential therapeutic targets for male infertility and contraception. AB - Mammalian sperm were previously shown to express the PP1gamma2 isoform of protein phosphatase 1 (PP1) as well as its regulatory proteins inhibitor 2 and glycogen synthase kinase 3. Furthermore, the development of sperm motility during transit through the epididymis correlates with changes in PP1 activity. Thus, since PP1 cellular activity is determined by the partners it binds, we embarked on a study aimed at defining the specific interactomes of PP1gamma1 and PP1gamma2 (the two known alternatively spliced variants of PP1gamma). To this end, exhaustive screens were performed on a human testis cDNA library using the yeast two-hybrid method. Among the various proteins detected, the most abundant interactors with PP1gamma2 were Nek2A and R15B. Closer sequence analysis revealed novel alternatively spliced variants of Nek2A and NIPP1, which we designated Nek2A-T and NIPP1-T, respectively. They were shown to be highly expressed in rat and human testis by Northern analysis and to result from alternative splicing events by RT-PCR. Thus, both the previously known Nek2A isoform and the novel Nek2A-T and NIPP1-T variants appear to bind PP1gamma2 in vitro (blot overlays) and in vivo by coexpression in yeast. The usefulness of testis-specific alternatively spliced proteins as targets for the development of novel therapeutic strategies for male infertility and contraception is discussed. PP1gamma2, Nek2A-T, and NIPP1-T are currently being investigated as alternatively spliced targets for signal transduction therapeutics. PMID- 15659833 TI - Relaxin and phosphodiesterases collaborate during decidualization. AB - During the evolution of mammals, the endometrium has developed for one reason only: to implant an embryo in the uterus. In higher primates, should an oocyte fail to be fertilized, then the endometrial layer is sloughed off during menses and the menstrual cycle starts again with a new round of endometrial differentiation. This stromal differentiation process is called decidualization and is accompanied in vivo by sustained high levels of intracellular cAMP. The present study was conducted to determine whether manipulation of cAMP phosphodiesterase (PDE) activities in cultured human endometrial stromal cells could positively influence the decidualization process. The combination of relaxin treatment with inhibition of PDE4 by the specific inhibitor rolipram induced a strong increase in relaxin-mediated cAMP production, both acutely, after 20 min, and after long-term treatment for 3 days, to promote a sustained intracellular cAMP concentration. Moreover, there was a dramatic synergistic effect on the decidualization phenotype, characterized both morphologically and by increased production of prolactin and insulin-like growth factor binding protein-1 gene transcripts. The observations that expression of PDE4D transcripts were selectively increased by cAMP and that inhibition of protein kinase A by H89 to potentially block negative feedback regulation enhanced the relaxin/rolipram mediated cAMP accumulation lead to a complex picture of cAMP regulation in these cells. There appears to be a coordinated contribution by relaxin and PDE4 at different levels to promote a sustained increased cAMP concentration during decidualization, and thus to provide an adequate maternal interface for the implanting blastocyst. PMID- 15659834 TI - Effects of cyclopentenone prostaglandins on the expression of heme oxygenase-1 in MCF-7 cells. AB - Cyclopentenone prostaglandins (cyPGs) are produced by dehydration of precursor molecules. The cyPGs are reported to have proapoptotic effects in a variety of cell types. However, cyPGs, particularly 15-deoxy-Delta(12,14)-prostaglandin J(2) (15d-PGJ(2)), can also exert cytoprotective effects at relatively low concentrations. The cytoprotective activity of cyPGs appears to be mediated by the reactive alpha,beta-unsaturated carbonyl group located in the cyclopentene ring. In this study, we investigated the effect of cyPGs on the expression of heme oxygenase-1 (HO-1), a ubiquitous stress-responsive enzyme that catalyzes oxidative cleavage of heme to form iron, carbon monoxide, and biliverdin. Treatment of the human breast cancer cell line (MCF-7) with 15d-PGJ(2) resulted in a concentration- and time-dependent increase in the expression of HO-1, whereas prostaglandin A(2) (PGA(2)) and the non-PG derivative 2-cyclopenten-1-one failed to induce HO-1 expression at the protein level. RT-PCR revealed that the expression of HO-1 mRNA was induced at 6 h by 15d-PGJ(2) at 10 microM. However, PGA(2) induced HO-1 mRNA expression at a higher concentration (30 microM). 2 Cyclopenten-1-one did not induce the expression of HO-1 mRNA at all. Likewise, 15d-PGJ(2) treatment for 6 h led to phosphorylation of Akt/protein kinase B (PKB) to a greater extent than that achieved with PGA(2). Thus, the induction of HO-1 expression and the activation of Akt/PKB by 15d-PGJ(2) and PGA(2) are likely to confer cytoprotective or antiapoptotic effects exerted by these cyPGs. PMID- 15659835 TI - Inhibition of cyclooxygenase-2 expression and restoration of gap junction intercellular communication in H-ras-transformed rat liver epithelial cells by caffeic acid phenethyl ester. AB - One of the most frequent defects in human cancers is the uncontrolled activation of the ras signaling pathways. Increased expression of cyclooxygenase-2 (COX-2) and inhibition of gap junction intercellular communication (GJIC) have been frequently observed in several forms of human malignancies. The present study investigated the effects of caffeic acid phenethyl ester (CAPE), a chemopreventive phytochemical derived from honey propolis, on COX-2 expression and GJIC in Harvey-ras-transformed WB-F344 rat liver epithelial cells (H-ras WB cells). H-ras induced COX-2 expression in WB-F344 rat liver epithelial cells (WB cells). H-ras WB cells also exhibited complete inhibition of GJIC and predominant unphosphorylation of connexin 43 (Cx43), a major protein modulating GJIC. CAPE significantly inhibited the constitutive expression of COX-2 and restored the disrupted GJIC through the phosphorylation of Cx43 at a concentration of 12.5 microM in H-ras WB cells. Although the molecular basis for the cancer chemopreventive activity of CAPE is not completely understood, several studies suggest that CAPE is a potent and specific inhibitor of the transcription factor nuclear factor kappaB (NF-kappaB) activation. We also found that CAPE significantly inhibited H-ras-induced NF-kappaB DNA-binding activity without affecting the activation of extracellular signal-regulated kinase and p38 mitogen activated protein kinase, which are major intracellular molecules involved in the Ras signaling pathways. In conclusion, CAPE may exert cancer chemopreventive effects through the inhibition of COX-2 expression and the restoration of disrupted GJIC induced by H-ras, possibly by targeting NF-kappaB. PMID- 15659836 TI - Comparison of gene expression in intra-abdominal and subcutaneous fat: a study of men with morbid obesity and nonobese men using microarray and proteomics. AB - Extent of intra-abdominal fat had significant linear relations with six metabolic coronary risk factors: systolic and diastolic blood pressure, fasting blood concentrations of glucose, high density lipoprotein (HDL) cholesterol, triglyceride, and plasminogen activator inhibitor-1. Tumor necrosis factor-alpha and adiponectin can be biological mediators from the intra-abdominal fat to the metabolic coronary risk factors. Complementarily, we describe a new study that will analyze the gene expression in intra-abdominal and subcutaneous fat on mRNA and protein level using high throughput methods. The study will elucidate further whether intra-abdominal obesity is the common denominator for the different components of the metabolic syndrome. PMID- 15659837 TI - GATA-1: friends, brothers, and coworkers. AB - GATA-1 is the founding member of the GATA family of transcription factors. GATA-1 and GATA family member GATA-2 are expressed in erythroid and megakaryocytic lineages, in which they play a crucial role in cell maturation and differentiation. GATA-1 regulates the transcription of many specific and nonspecific erythroid genes by binding to DNA at the consensus sequence WGATAR, which is recognized by all of the GATA family of transcription factors. However, it was identified in eosinophilic cells and also in Sertoli cells in testis. Its activity depends on close cooperation with a functional network of cofactors, among them Friend of GATA, PU.1, and CBP/p300. The GATA-1 protein structure has been well described and includes two zinc fingers that are directly involved in the interaction with DNA and other proteins in vivo. GATA-1 mutations in the zinc fingers can cause deregulation of required interactions and lead to severe dysfunction in the hematopoietic system. PMID- 15659838 TI - Tetramethoxy hydroxyflavone p7F downregulates inflammatory mediators via the inhibition of nuclear factor kappaB. AB - Artemisia has been traditionally used in Korean herbal medicine to clear damp heat and to treat uteritis and jaundice. Flavonoids isolated from Artemisia are also known to possess anti-inflammatory activities. In this study, 5,6,3',5' tetramethoxy 7,4'-hydroxyflavone (p7F) was isolated from Artemisia absinthium. We examined in vitro and in vivo regulatory functions of p7F on the production of nitric oxide (NO), prostaglandin E(2) (PGE(2)), and tumor necrosis factor-alpha (TNF-alpha) as well as the expression of inducible NO synthase (iNOS), cyclooxygenase-2 (COX-2), and collagen-induced arthritis. p7F inhibited the expression or production of proinflammatory mediators such as COX-2/PGE(2) and iNOS/NO in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. p7F also suppressed the serum level of TNF-alpha in mice treated with collagen and inhibited nuclear factor-kappaB (NF-kappaB) activation as well as NF-kappaB promoter activity in RAW 264.7 cells stimulated with LPS. This compound directly inhibited the intracellular accumulation of reactive oxygen species in hydrogen peroxide-stimulated RAW 264.7 cells. p7F has antioxidant activity and inhibits NF kappaB activation. Taken together, these results suggest that p7F can be clinically applied to the treatment of inflammatory diseases. PMID- 15659839 TI - Sp1 and NF-kappaB transcription factor activity in the regulation of the p21 and FasL promoters during promyelocytic leukemia cell monocytic differentiation and its associated apoptosis. AB - Treatment of human acute promyelocytic leukemia cells with phorbol 12-myristate 13-acetate (PMA) results in growth arrest and differentiation toward monocytes, which subsequently die by apoptosis. However, the relationship between terminal differentiation and apoptosis remains unclear. Here we have studied Sp1 and nuclear factor kappaB (NF-kappaB) transcription factor activity in controlling promoters of cell cycle-regulating (p21/WAF1/CIP1) and cell death (FasL) genes during monocytic differentiation and apoptosis of the human acute promyelocytic leukemia cell lines NB4 and HL-60. Using the electrophoretic mobility shift assay, we observed that PMA treatment of NB4 cells caused an early response in Sp1 binding to the p21 and FasL promoters at 8 h. The firmly adherent cell phenotype, characteristic of differentiated cells, retained Sp1-binding activity to either promoter, but it was often lost completely in detached, apoptotic cells. The association of Sp1 with the p21 promoter during monocytic differentiation correlated with the levels of expressed p21 in the cytoplasmic fraction, as detected by immunoblotting. In HL-60 cells, very weak or no Sp1 binding to either promoter was observed. Low NF-kappaB affinity for its consensus sites and to the FasL promoter was characteristic of apoptotic cells. The results of this study suggest a positive role of Sp1 and NF-kappaB, as regulators of p21 and FasL genes, in leukemic cell survival and monocytic differentiation and a negative role in apoptotic cell death. PMID- 15659841 TI - The hinge region of the androgen receptor plays a role in proteasome-mediated transcriptional activation. AB - To investigate the function of the hinge region in transcriptional activation by the androgen receptor, we compared the actions of the wild-type receptor with a mutant receptor, deleted of amino acids 628-646 of the hinge. The role of the proteasome on the expression and activity of these two proteins was investigated. The deletion mutant demonstrated a threefold increase in transcriptional activity when compared to the wild-type receptor protein. Furthermore, we found that hormone-dependent stabilization of the receptor protein was more enhanced for the deletion mutant. In addition, experiments using the proteasome inhibitor, MG132, demonstrated that the deletion mutant is more sensitive to proteasome-mediated degradation than the wild-type receptor. However, inhibition of the proteasome had a negative effect on the transcriptional activity of the deletion mutant. Taken together, our results suggest that the hinge region not only plays an important role in controlling the transactivation potential of the androgen receptor but also in determining the influence of the proteasome on androgen receptor-mediated transcriptional activation. PMID- 15659840 TI - Effects of curcumin (diferuloylmethane) on nuclear factor kappaB signaling in interleukin-1beta-stimulated chondrocytes. AB - Curcumin (diferuloylmethane) is a nontoxic dietary pigment in tumeric and curry and a potent inhibitor of the common transcription factor Nuclear Factor kappaB (NF-kappaB) in several cell types. It is well established that some of the catabolic effects of the proinflammatory cytokines interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha in osteoarthritis are regulated by the activation of NF-kappaB. Therefore, the aim of this study was to determine whether curcumin modifies the catabolic response of chondrocytes to IL-1beta. Human articular chondrocytes were prestimulated with 10 ng/mL IL-1beta for 0, 4, 8, 12, or 24 h and then cotreated with 50 microM curcumin for 0, 12, 24, 36, or 48 h. Synthesis of the cartilage-specific collagen type II and matrix-degrading enzyme matrix metalloproteinase-3 (MMP-3) was investigated in chondrocytes by Western blot analysis. Activation and nuclear translocation of NF-kappaB were observed by immunofluorescence microscopy. IL-1beta induced a decrease in collagen type II and upregulation of MMP-3 in a time-dependent manner. Upregulation of MMP-3 was inhibited by curcumin in a time-dependent manner. In addition, IL-1beta-induced a decrease in type II collagen, which was relieved by curcumin treatment. In response to IL-1beta, NF-kappaB translocated to the nucleus, but translocation was inhibited by curcumin, as revealed by immunofluorescence microscopy. Taken together, these results confirmed an IL-1beta-mediated upregulation of proinflammatory MMP-3 in chondrocytes via an NF-kappaB activation mechanism. Curcumin protected chondrocytes from the catabolic effects of IL-1beta, such as MMP-3 upregulation, and interestingly also relieved cytokine-induced suppression of matrix protein synthesis. Therefore, curcumin antagonizes crucial catabolic effects of IL-1beta signaling that are known to contribute to the pathogenesis of osteoarthritis. PMID- 15659842 TI - Polynucleosomal synthesis of poly(ADP-ribose) causes chromatin unfolding as determined by micrococcal nuclease digestion. AB - We have evaluated the influence of protein poly(ADP-ribosyl)ation in the relaxation of chromatin by exposing a rat liver polynucleosomal extract to micrococcal nuclease (MNase) digestion. The kinetic susceptibility of polynucleosomes to endonuclease digestion was determined as a function of the time of incubation as well as endonuclease concentration. To validate our assay, we also ran control experiments with protein-free calf thymus DNA as the opposite of polynucleosomal DNA. Rat liver chromatin was also incubated in the absence or presence of exogenously added 200 microM betaNAD(+), the poly(ADP-ribosyl)ation substrate, before MNase digestion. For incubations in the presence of betaNAD(+), the synthesis of polynucleosomal poly(ADP-ribose) was stopped with 1 mM benzamide. After addition of MNase, endonuclease digestion was blocked with EDTA to chelate the Mg(2+) ions needed for enzymatic activation, and the samples were subjected to electrophoresis through 1.5% agarose gels. As expected, a faster degradation of chromatin into oligonucleosomal DNA ladders was observed upon protein poly(ADP-ribosyl)ation when the chromatin extract was preincubated with 200 microM betaNAD(+). Thus, our results are consistent with the conclusion that the covalent poly(ADP-ribosyl)ation of polynucleosomal proteins favors a more "relaxed" or "open" structure, which renders chromatin more susceptible to MNase digestion. PMID- 15659843 TI - From electrophysiology to chromatin: a bottom-up approach to Angelman syndrome. AB - Angelman syndrome is one of the most studied human diseases related to a gene that is expressed on the maternal chromosome only in at least some brain cells. It is caused by inactivation of the UBE3A gene in the brain due to various abnormalities of the 15q11-q13 chromosome inherited from the mother. It is characterized by severe developmental delay, seizures, virtual absence of speech, motor impairment, and a particular behavioral phenotype. Studies of cortical, electromyographic and cerebellar electrophysiology in patients with Angelman syndrome and a mouse model revealed unique rhythmic neurophysiological activities in the cerebral cortex, cerebellar cortex, and muscles. The oscillatory patterns may be linked to molecular pathophysiology of the syndrome involving dysregulation of synaptic neurotransmission through UBE3A-related modulation of functional GABAA receptor complexes. PMID- 15659844 TI - Repetitive treatments of colon HT-29 cells with diallyl disulfide induce a prolonged hyperacetylation of histone H3 K14. AB - Diallyl disulfide (DADS) is a sulfur compound derived from garlic. Several studies carried out in rodents have revealed protective effects of DADS against colon carcinogenesis. The antipromoting effects of DADS may be partly related to its ability to inhibit tumoral cell proliferation. In a previous study, we have shown that in two human colon tumor cell lines (HT-29 and Caco-2) seeded at a low density (0.2 x 10(6) cells/100-mm petri dish), DADS antiproliferative effects were associated with a transient increase of histone H3 K14 acetylation. Moreover, DADS could inhibit nuclear histone deacetylase activity. Therefore, in the present study, we examined the possible effects of different experimental conditions (HT-29 cells at high density, repetitive treatments with DADS) on the pattern of DADS-induced histone hyperacetylation. Using HT-29 cells seeded at a higher density (5 x 10(6) cells/100-mm petri dish), we found that DADS induced histone H3 K14 hyperacetylation rapidly (3 h). When administrated as single treatments, the DADS effect on histone H3 K14 remained transient. In contrast, repetitive treatment with DADS resulted in a prolonged hyperacetylation of histone H3 K14. Whatever the cell culture conditions were, DADS had no effect on histone H4 acetylation. Thus, in vitro, the cell density and pattern of DADS treatment influenced the HT-29 nuclear response to DADS. DADS belongs to food borne molecules that may play a role in chromatin remodeling and contribute to the nutritional modulation of gene expression. PMID- 15659845 TI - Thymocyte apoptosis induced by various compounds including YO-2 is accompanied by a change in chromatin structure. AB - To elucidate the role of chromatin structure in DNA fragmentation during apoptosis, we have examined whether chromatin structural change is observed after treatment with proapoptotic compounds. Analysis of the circular dichroism (CD) spectrum of the soluble chromatin from dexamethasone-treated thymocytes revealed a decrease in alpha-helical content. Mifepristone, an antagonist of glucocorticoid receptor, prevented both the change in chromatin structure and DNA fragmentation induced by dexamethasone. The effect of YO-2 [trans-4 aminomethylcyclohexanecarbonyl-l-(O-picolyl)tyrosine-n-octylamide], which possesses proapoptotic action, on chromatin structure was also examined. Judging from the CD spectrum of the soluble chromatin from YO-2-treated thymocytes, the structure was changed by this compound as well as by dexamethasone. A decrease in alpha-helical content was also observed in cells treated with etoposide, which is used clinically as an anticancer agent. These results suggest that the change in chromatin structure is likely to be an important process in DNA fragmentation of cells undergoing apoptosis. PMID- 15659846 TI - Defining the chemotherapeutic targets of histone deacetylase inhibitors. AB - The use of many conventional chemotherapeutic drugs is often severely restricted due to dose-limiting toxicities, as these drugs target the destruction of the proliferating fraction of cells, often with little specificity for tumor cells over proliferating normal body tissue. Many newer drugs attempt to overcome this shortcoming by targeting defective gene products or cellular mechanisms that are specific to the tumor, thereby minimizing the toxicity to normal tissue. Histone deacetylase inhibitors are an example of this type of tumor-directed drug, having significant toxicity for tumors but minimal effects on normal tissue. These drugs can affect the transcriptional program by modifying chromatin structure, but it is not yet clear whether specific transcriptional changes are directly responsible for their tumor-selective toxicity. Recent evidence suggests that transcriptional changes underlie their cytostatic activity, although this is not tumor-selective and affects all proliferating cells. Here we present evidence that supports an alternative mechanism for the tumor-selective cytotoxicity of histone deacetylase inhibitors. The target is still likely to be the chromatin histones, but rather than transcriptional changes due to modification of the transcriptionally active euchromatin, we propose that hyperacetylation and disruption of the transcriptionally inactive heterochromatin, particularly the centromeric heterochromatin, and the inability of tumor cells to cell cycle arrest in response to a specific checkpoint, underlies the tumor-selective cytotoxicity of these drugs. PMID- 15659847 TI - Role of acetylases and deacetylase inhibitors in IRF-1-mediated HIV-1 long terminal repeat transcription. AB - There is strong evidence that both transcriptional activation and silencing are mediated through the recruitment of enzymes that control reversible protein acetylation: histone acetylase (HAT) and histone deacetylase proteins. Acetylation is also a critical post-translational modification of general and tissue-specific transcription factors. In HIV-1-infected cells, the long terminal repeat (LTR) promoter, once organized into chromatin, is transcriptionally inactive in the absence of stimulation. LTR transcription is regulated by protein acetylation, since treatment with deacetylase inhibitors markedly induces transcriptional activity of the LTR. Besides cellular transcription factors involved in LTR activation, early in infection, and during reactivation from latency, we have previously shown that proteins of the IRF family play an important role. In particular, IRF-1 is able per se to stimulate HIV-1 LTR transcription even in the absence of Tat. IRF-1 is also acetylated and associates with HATs such as p300/CBP and PCAF to form a multiprotein complex that assembles on the promoter of target genes. Here we show that CBP can be recruited by IRF-1 to the HIV-1 LTR promoter even in the absence of Tat and that treatment with deacetylase inhibitors, such as trichostatin A (TSA), increases LTR transactivation in response to both IRF-1 and Tat. These results help to define the architecture of interactions between transcription factors binding HIV-1 LTR and confirm the possibility that deacetylase inhibitors, such as TSA, combined with antiviral therapy may represent a valuable approach to control HIV-1 infection. PMID- 15659848 TI - Histone structures: targets for modifications by molecular assemblies. AB - The core histone proteins contain modification sites that are key elements in the regulation of the cell cycle, DNA replication and repair with histone assembly, control of gene expression, and transcriptional elongation. Much work has been done on the various molecular assemblies that remodel nucleosomes, methylate, ubiquitinate, and cause ADP-ribosylation of histones, and acetylate and phosphorylate core histone tails. The core histones are the final targets of the enzymes in the molecular assemblies. What structural changes in the histones are correlated with these modifications? This paper considers the high-resolution structure of the histone octamer and stresses the importance of histone docking sequences in the binding of the two (H2A-H2B) dimers to the (H3-H4)(2) tetramer. There is an extensive acid-base area of interaction between histone octamers in crystals at high salt, which may have implications for nucleosome remodeling. We show that there are regions of high alpha-helix probability in all core histone N terminal tails in regions where lysine acetylation occurs. There are also consensus sequences spanning up to eight amino acid residues between some histone tail regions. Circular dichroism studies using synchrotron radiation at wavelengths as low as 130 nm are promising for the accurate measurement of changes of histone secondary structure related to function. PMID- 15659849 TI - HDAC-3 participates in the repression of e2f-dependent gene transcription in primary differentiated neurons. AB - Activation of e2f-1 gene expression is an event that has been now established in many models of neuronal apoptosis. Accumulated E2F-1 protein has also been observed in post mortem brains obtained from patients suffering from different neurodegenerative diseases. We have previously shown in primary neuronal cultures that e2f-1 gene transcription was actively repressed in neuroprotective conditions through HDAC-dependent regulation on the E2F-responsive elements (E2F REs) located in the e2f-1 gene promoter. Here, we further investigated the protein complex bound to these sites by gel shift analysis. We found that the specific protein binding to E2F-REs is altered in apoptotic conditions compared to neuroprotective conditions, suggesting that the proteic constituents of the complex are likely to be modified upon apoptosis onset. Indeed, Western blot analysis showed a time-dependent degradation of the Rb/E2F binding protein HDAC-3 during apoptosis, a degradation that is caspase-dependent. Altogether, these data point to HDAC-3 as a good candidate involved in the active e2f-1 repression necessary for neuroprotection. PMID- 15659850 TI - Plants contain a high number of proteins showing sequence similarity to the animal SUV39H family of histone methyltransferases. AB - The SET domain, first identified within and named after proteins encoded by three Drosophila genes [Su(var)3-9, E(z), and Trithorax], is recognized as a signature motif for histone methyltransferases that are involved in epigenetic processes. The SUV39H family of SET domain proteins methylate specifically the residue lysine 9 of histone H3, creating a code for gene silencing. This family of proteins contain at their C termini a unique catalytic domain consisting of pre SET, SET, and post-SET domains. Sequence homology-based searches identified 15 Arabidopsis, 14 maize, and 12 rice proteins that can be assigned to the SUV39H family. These high numbers in plants are in marked contrast to the situation in animals, in which each species appears to contain only two to three proteins of this family. Our phylogenetic analyses revealed that plant proteins can be classified into seven orthology groups. Representative members of each group can be found in single plant species, suggesting that different group members are evolutionarily conserved to perform specific functions. PMID- 15659851 TI - Anti-KSbF6 structure of CaTbF6 and CdTbF6: a confirmation of the singular crystal chemistry of Tb4+ in fluorides. AB - The crystal structures of two new tetravalent terbium fluorides, CaTbF6 and CdTbF6, have been determined from X-ray and neutron powder diffraction data. The title compounds exhibit an anti-KSbF6 structure, the three-dimensional framework of which is built of [TbF6]2- chains of edge-sharing dodecahedra further linked, by sharing corners, to isolated [MF6]4- octahedra (M=Ca, Cd). The mechanism of the anionic sublattice rearrangement when going from KSbF6 to CaTbF6 is described and related to a simple cubic fluoride-ion packing. Comparison with the crystal structures of beta-BaTbF6 and other representatives of the M(II)M('IV)F6 family allows the singular crystal-chemical properties of some fluoroterbates to be emphasized. PMID- 15659852 TI - Structural characterization of SmMn2GeO7 single microcrystals by electron microscopy. AB - Single microcrystals of the new compound samarium dimanganese germanium oxide, SmMn2GeO7, were grown using the flux method in a double spherical mirror furnace (DSMF). The micrometric crystals were observed and chemically analysed with scanning electron microscopy (SEM) and X-ray energy dispersive spectroscopy (EDX). The structural characterization and chemical analysis of these crystals were also carried out using transmission electron microscopy (TEM) and high resolution transmission electron microscopy (HRTEM), together with electron energy-loss spectroscopy (EELS). We found that the new quaternary compound crystallizes in the orthorhombic system with the point group mmm (D2h), space group Immm (No. 71) and cell parameters a=8.30 (10), b=8.18 (10), c=8.22 (10) A and V=558.76 A3. PMID- 15659853 TI - Crystal structure study of a beta'-copper vanadium bronze, Cu(x)V2O5 (x = 0.63), by X-ray and convergent beam electron diffraction. AB - The single-crystal structure of a beta'-copper vanadium bronze, Cu0.63V2O5, has been studied at room temperature and 9.6 K, and compared with that of the beta sodium vanadium bronze, Na0.33V2O5, structure. No convincing evidence to oppose an assignment of centrosymmetric C2/m symmetry to the structure was identified using the X-ray data. A subsequent convergent beam electron diffraction (CBED) experiment was performed and confirmed the C2/m space group. The oxygen-vanadium atom framework of Cu0.63V2O5 is close to that of Na0.33V2O5. However, in the copper compound the Cu atoms are located in two positions: Cu1 in the 4(i) position with x=0.541, y=0 and z=0.345, and Cu2 in the 8(j) position with x=0.529, y=0.038 and z=0.357. The crystal structure changes little with temperature. Disorder of the Cu ion over two sites is seen at 9.6 K. This suggests that distribution of the Cu atoms over two sites is of a more static than dynamic nature. PMID- 15659854 TI - Photo-induced spin-transition: the role of the iron(II) environment distortion. AB - The [FeLn(NCS)2] iron(II) spin-crossover complexes cover a wide range of magnetic behaviour. Owing to the large number of known structural and magnetic data, this series is perfectly adapted to the investigation of the structure-magnetic properties relationship. In this paper we propose a new structural parameter, denoted Theta, which is used to correlate the features of the spin-crossover phenomena with the distortion of the iron environment. In particular, this parameter has shed light on the role of such distortion on the limiting temperature of photo-inscription, known as T(LIESST). A strong dependence of T(LIESST) on Theta is clearly demonstrated. The stronger the distortion the higher the T(LIESST) value. This structure-property dependence represents, for instance, a powerful tool to estimate the highest potential T(LIESST) value for a series of complexes. This limit in the [FeLn(NCS)2] series is estimated to be around 120 K, which probably prevents their use in any industrial application. PMID- 15659855 TI - Method for the computational comparison of crystal structures. AB - A new method for assessing the similarity of crystal structures is described. A similarity measure is important in classification and clustering problems in which the crystal structures are the source of information. Classification is particularly important for the understanding of properties of crystals, while clustering can be used as a data reduction step in polymorph prediction. The method described uses a radial distribution function that combines atomic coordinates with partial atomic charges. The descriptor is validated using experimental data from a classification study of clathrate structures of cephalosporins and data from a polymorph prediction run. In both cases, excellent results were obtained. PMID- 15659857 TI - Formation of quinol co-crystals with hydrogen-bond acceptors. AB - The crystal structures of eight new co-crystals of quinol with pyrazine, piperazine, morpholine, pyridine, piperidine, 4,4'-bipyridine, N-methylmorpholine and N,N'-dimethylpiperazine are reported. Quinol forms 1:1 co-crystals with pyrazine, piperazine and N,N'-dimethylpiperazine, but 1:2 co-crystals with morpholine, 4,4'-bipyridine, N-methylmorpholine, pyridine and piperidine. This difference can be rationalized in most cases by the presence of, respectively, two or one strong hydrogen-bond acceptor(s) in the guest molecule. The exception to this generalization is 4,4'-bipyridine, which forms a 1:2 co-crystal, possibly to optimize crystal packing. All structures are dominated by hydrogen bonding between quinol and the guest molecules. A doubly bridging motif, which connects pairs of quinol and guest molecules via NH...O or CH...O interactions, is present in all but the sterically hindered N,N'-dimethylpiperazine and N-methylmorpholine co-crystals. PMID- 15659856 TI - Determination of the structure of the violet pigment C22H12Cl2N6O4 from a non indexed X-ray powder diagram. AB - The violet pigment methylbenzimidazolonodioxazine, C22H12Cl2N6O4 (systematic name: 6,14-dichloro-3,11-dimethyl-1,3,9,11-tetrahydro-5,13-dioxa-7,15 diazadiimidazo-[4,5-b:4',5'-m]pentacene-2,10-dione), shows an X-ray powder diagram consisting of only ca 12 broad peaks. Indexing was not possible. The structure was solved by global lattice energy minimizations. The program CRYSCA [Schmidt & Kalkhof (1999), CRYSCA. Clariant GmbH, Pigments Research, Frankfurt am Main, Germany] was used to predict the possible crystal structures in different space groups. By comparing simulated and experimental powder diagrams, the correct structure was identified among the predicted structures. Owing to the low quality of the experimental powder diagram the Rietveld refinements gave no distinctive results and it was difficult to prove the correctness of the crystal structure. Finally, the structure was confirmed to be correct by refining the crystal structure of an isostructural mixed crystal having a better X-ray powder diagram. The compound crystallizes in P1, Z=1. The crystal structure consists of a very dense packing of molecules, which are connected by hydrogen bridges of the type N-H...O=C. This packing explains the observed insolubility. The work shows that crystal structures of molecular compounds may be solved by lattice energy minimization from diffraction data of limited quality, even when indexing is not possible. PMID- 15659858 TI - Effect of pressure on the crystal structure of L-serine-I and the crystal structure of L-serine-II at 5.4 GPa. AB - The crystal structure of L-serine has been determined at room temperature at pressures between 0.3 and 4.8 GPa. The structure of this phase (hereafter termed L-serine-I), which consists of the molecules in their zwitterionic tautomer, is orthorhombic, space group P212121. The least compressible cell dimension (c), corresponds to chains of head-to-tail NH...carboxylate hydrogen bonds. The most compressible direction is along b, and the pressure-induced distortion in this direction takes the form of closing up voids in the middle of R-type hydrogen bonded ring motifs. This occurs by a change in the geometry of hydrogen-bonded chains connecting the hydroxyl groups of the -CH2OH side chains. These hydrogen bonds are the longest conventional hydrogen bonds in the system at ambient pressure, having an O...O separation of 2.918 (4) A and an O...O...O angle of 148.5 (2) degrees ; at 4.8 GPa these parameters are 2.781 (11) and 158.5 (7) degrees . Elsewhere in the structure one NH...O interaction reaches an N...O separation of 2.691 (13) A at 4.8 GPa. This is amongst the shortest of this type of interaction to have been observed in an amino acid crystal structure. Above 4.8 GPa the structure undergoes a single-crystal-to-single-crystal phase transition to a hitherto uncharacterized polymorph, which we designate L-serine II. The OH...OH hydrogen-bonded chains of L-serine-I are replaced in L-serine-II by shorter OH...carboxyl interactions, which have an O...O separation of 2.62 (2) A. This phase transition occurs via a change from a gauche to an anti conformation of the OH group, and a change in the NCalphaCO torsion angle from 178.1 (2) degrees at 4.8 GPa to -156.3 (10) degrees at 5.4 GPa. Thus, the same topology appears in both crystal forms, which explains why it occurs from one single-crystal form to another. The transition to L-serine-II is also characterized by the closing-up of voids which occur in the centres of other R type motifs elsewhere in the structure. There is a marked increase in CH...O hydrogen bonding in both phases relative to L-serine-I at ambient pressure. PMID- 15659859 TI - Structures of the monofluoro- and monochlorophenols at low temperature and high pressure. AB - 2-Fluorophenol, 3-fluorophenol and 3-chlorophenol were recrystallized from frozen solids at 260, 263 and 283 K. All compounds were also crystallized by the application of high pressure (0.36, 0.12 and 0.10 GPa). While 3-fluorophenol and 3-chlorophenol yielded the same phases under both conditions, different polymorphs were obtained for 2-fluorophenol. 4-Chlorophenol was crystallized both from the melt and from benzene to yield two different ambient-pressure polymorphs; crystallization from the melt at 0.02 GPa yielded the same phase as from benzene at ambient pressure. 3-Fluorophenol is unusual in forming a hydrogen bonded chain along a 2(1) screw axis. Such behaviour is usually only observed for small alcohols, but here it appears to be stabilized by intermolecular C-H...F hydrogen-bond formation. 3-Chlorophenol is a more typical large alcohol and emulates a fourfold screw axis with two independent molecules positioned about a 2(1) axis, although there are significant distortions from this ideal geometry. The two phases of 4-chlorophenol consist of chains or rings connected by C-Cl...H interactions. The low-temperature and high-pressure polymorphs of 2-fluorophenol consist of chains of molecules connected through OH...OH hydrogen bonds; while inter-chain C-H...F interactions are significant at high pressure, there are none in the low-temperature form. PMID- 15659860 TI - Structure determination of forms I and II of phenobarbital from X-ray powder diffraction. AB - From pure powders of forms I and II of phenobarbital, X-ray diffraction patterns were recorded at room temperature. The starting crystal structural models were found by a Monte-Carlo simulated annealing method. The structures of the two forms were obtained through Rietveld refinements. Soft restraints were applied on bond lengths and bond angles, all H-atom positions were calculated. The cell of form I is monoclinic with the space group P2(1)/n, Z=12, Z'=3. Form II has a triclinic cell, with the space group P1 , Z=6, Z'=3. For both forms, the crystal cohesion is achieved by networks of N-H...O hydrogen bonds along [101]. The broadening of the Bragg peak profiles is interpreted in terms of isotropic strain effects and anisotropic size effects. PMID- 15659861 TI - X-ray studies on crystalline complexes involving amino acids and peptides. XLII. Adipic acid complexes of L- and DL-arginine and supramolecular association in arginine-dicarboxylic acid complexes. AB - The adipic acid complexes of DL-arginine and L-arginine are made up of zwitterionic, singularly positively charged arginium ions and doubly negatively charged adipate ions, with a 2:1 stoichiometry. One of the two crystallographically independent arginium ions in the L-arginine complex has a conformation hitherto unobserved in crystal structures containing the amino acid. In the present study the structural data on arginine complexes of saturated dicarboxylic acids with 0-5 C atoms separating the two carboxyl functions are given. In terms of molecular aggregation, formic and acetic acid complexes behave in a similar way to those involving fairly long carboxylic acids such as adipic acid. By and large, the supramolecular assembly in complexes involving dicarboxylic acids with 3 or more C atoms separating the carboxyl groups (glutaric, adipic and pimelic acids), and those involving formic and acetic acids, have common features. The aggregation patterns in complexes involving oxalic, malonic and maleic acids do not share striking features among themselves (except for the mode of hydrogen-bonded dimerization of arginium ions) or with those involving larger dicarboxylic acids. Complexes of succinic acid, the shortest linear dicarboxylic acid, share features with those involving shorter as well as longer dicarboxylic acids. The difference in the behaviour of long and short dicarboxylic acids and the ambiguous behaviour of succinic acid can be broadly related to their lengths. PMID- 15659862 TI - Temperature effects on the hydrogen-bond patterns in 4-piperidinecarboxylic acid. AB - A temperature-controlled X-ray powder diffraction experiment, complemented with TGA and DSC analysis, allowed us to follow changes in the molecular conformation and hydrogen-bond patterns of 4-piperidinecarboxylic acid. The presence of three phases is confirmed. Phase 1 represents the monohydrated form of 4 piperidinecarboxylic acid, which exists from room temperature to 359 K, where dehydration occurs. Phase 2 measured at 363 K corresponds to an anhydrous form of the acid. At ca 458 K the onset of a second, more gradual transition is observed, which ends at around 543 K. Phase 3 measured at 543 K is a high-temperature anhydrous form of the acid. The structures of phases 2 and 3 were solved from synchrotron powder diffraction data by simulated annealing using the DASH program and refined by the Rietveld method. The phase changes are accompanied by modification of the hydrogen-bond patterns and of the torsional orientation of the terminal carboxylate group. This group makes a 49 degrees rotation about the C1-C2 bond during the first transition. PMID- 15659863 TI - Supramolecular structures of 1-phenylethylammonium tartrates. AB - The structures of six 1-phenylethylammonium tartrates have been determined and in each of them a distinctive hydrogen-bonded anion substructure can be identified. (S)-1-Phenylethylammonium (R,R)-hydrogen tartrate [(I), P21, Z'=1] contains anion sheets built from a single type of R4(4)(22) ring with cations pendent, via three N-H...O hydrogen bonds, from just one face of the sheet. (S)-1 Phenylethylammonium rac-hydrogen tartrate [(II), P21, Z'=2] and its enantiomorph (R)-1-phenylethylammonium rac-hydrogen tartrate [(III), P21, Z'=2] contain anion sheets built from four types of ring, R2(2)(10), R2(2)(12), R2(4)(14) and R4(4)(20), and there are cations pendent from both faces of the sheet. The anion substructure in bis[(S)-1-phenylethylammonium] (R,R)-tartrate [(IV), P2(1), Z'=1] consists of simple C(5) chains, which are linked into sheets by the cations, while in bis(rac-1-phenylethylammonium) (R,R)-tartrate [(V), P2(1), Z'=2] there are anion sheets containing two distinct types of R4(4)(22) ring, with equal numbers of (R) and (S) cations pendent from each face of the anion sheet. Bis[(R) 1-phenylethylammonium] rac-tartrate methanol hemisolvate [(VI), P1, Z'=4, with 14 independent components in the asymmetric unit] contains anion sheets built from two types of R2(2)(12) ring and two types of R6(6)(32) ring; half of the cations and half of the methanol molecules are pendent from each face of the sheet. PMID- 15659864 TI - Charge density of (-)-strychnine from 100 to 15 K, a comparison of four data sets. AB - Strychnine has an interesting oligocyclic structure of seven condensed rings. It is easy to crystallize and gives crystals of excellent quality which diffract nicely to high regions in reciprocal space. It was thus chosen for a comparative charge-density study based on four high-resolution data sets (sin theta;/lambda >or= 1.15 A-1) that were measured with different experimental setups in the temperature range 100-15 K. In addition, a theoretical charge density was derived from a B3LYP/6-311++G(3df,3pd) calculation. The agreement expressed in bond topological parameters among the four experimental charge densities is better than between experiment and theory. PMID- 15659866 TI - A review of association of dietary factors in gallbladder cancer. AB - Gallbladder cancer (GBC) is the prominent malignancy of hepato-biliary tract, being the fifth most common carcinoma for gastrointestinal tract in United States. Epidemiological studies world wide have implicated dietary factors in the development of gallbladder cancer. The ecological evidences indicate considerable geographic variation in the incidence of gallbladder cancer. However the variations in GBC incidence of different populations might be partly determined by their dietary variations. Higher intake of energy and carbohydrate possibly increase the risk of gallbladder cancer. Obesity plays an important role in the causation of GBC. Adequate intake of fruits and vegetables probably reduce the risk of GBC. This nutritional preventive effect against GBC could be attributed to high content of vitamins, carotenes and fibers. They can not be too emphatically stated as the sole determinants of GBC. It is apparently clear that a variety of essential nutrients can significantly modify the carcinogenic process. Furthermore, an attempt has been made to establish an association between dietary factors and the occurrence of gallbladder cancer. PMID- 15659868 TI - Pancreatic involvement in Von Hippel-Lindau disease. AB - BACKGROUND: Involvement of the pancreas in Von Hippel-Lindau disease that is a tumor predisposing syndrome mentioned in literature with some morbid and mortal progression. AIMS: For evaluation the faith of the pancreatic involvement in VHL disease we analysed our patient population with VHL disease. MATERIALS AND METHODS: 12 of the 56 patients that were evaluated in our institute with the diagnosis of Von Hippel-Lindau disease had pancreatic involvement. They are periodically examined for 5 years follow up period. Their retrospective analysis was accomplished. RESULTS AND CONCLUSIONS: Pancreatic involvement in our patient population disclosed lesions that were multicysts or serous cystadenomas. During follow up period, we did not observe significant morbidity related to pancreatic involvement. Repeated radiological examination of pancreatic lesions disclosed insignificant modifications such as slight increase or decrease in size. Whereas we considered morbidity and mortality related to renal and central nervous system pathologies in VHL disease. Shortly, even pancreatic involvement in VHL disease requires close clinical follow up, morbidity and mortality in this case is not severe as in renal and the central nervous system involvement. PMID- 15659867 TI - FISH for HER-2/neu in breast cancer: standardization makes the difference! AB - CONTEXT: Overexpression of HER-2/neu oncogene in breast cancer patients is correlated with disease free survival (DFS) and overall survival (OS). The most commonly used methods for the detection of HER-2/neu status are immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH). However, therse is a lot of controversy with regard to the best method. Most of the FISH studies chose arbitrary cut-off levels for positive results (10%) and had no validation. AIM: In order to address these issues, we designed a pilot study of 38 samples with known IHC status representing all 4 categories. SETTINGS AND DESIGN: FISH was performed using Vysis Pathvysion probe. For validation, 5 cases of reduction mammoplasty were analyzed using same protocols. RESULTS: Our results showed significant discordance between FISH and IHC. The rate of discordance was much higher in the 0, 1+, and 2+ categories compared to published literature. This could be due to the lower cut-off rates for positive amplification established by validation in our study (5.7% vs 10%). Our analysis showed that FISH positive and IHC negative patients have a poor prognosis in terms of DFS and OS compared to FISH negative and IHC negative patients. Further, our results also showed that IHC in comparison to FISH has a comparable specificity (98%), but has a very low sensitivity (46%). CONCLUSION: Based on these results, we consider FISH to be the gold standard for detecting HER-2/neu status in breast cancer. PMID- 15659869 TI - Tobacco use among school personnel in Rajasthan, India. AB - BACKGROUND: A very little information about tobacco use among school personnel is available. This is a step to have cross country and within country data using standardized methodology. AIMS: To obtain baseline information about tobacco use prevalence, knowledge and attitude among school personnel. STUDY DESIGN: Two stage cluster sample. SETTING: A state of Rajasthan. MATERIALS AND MATERIALS AND METHODS: A cross-sectional study, using anonymous self-administered questionnaire. A sample of schools with probability proportional to the enrollment in grades 8-10. All school personnel in sampled schools were eligible to participate. STATISTICAL ANALYSIS: Percentage, 95% confidence interval. RESULTS: School response rate was 97.4% (75/77) and school personnel response rate was 67.2% (909/1352). Majority of school personnel (men 69%, women 31%) were school teachers (78.3%). The prevalence of ever any tobacco use was reported by 35.9%, more among men than women (46.2% vs. 13.0%). The prevalence of current daily smoking was reported by 14.4% (men 20.6%, women 0.8%) and occasional by 7.3%, where as current daily smokeless tobacco use was 11.7% (men 16.4%, women 1.1%) and occasional 13.6%. Current daily tobacco use was significantly more among men than women. Four out of nine reported their schools have a tobacco prohibiting policy for both students (48.4%) as well as for school personnel (44.4%) and about same (47.2%) reported their schools enforce its tobacco policy or rule. Over 85% of all school personnel strongly support the tobacco control policies and wanted training in tobacco cessation and prevention. CONCLUSION: First study from Rajasthan to report tobacco use among school personnel. School personnel not only strongly support the tobacco control policies but also ready to work for its successful implementation with proper training. PMID- 15659870 TI - Omentoplasty versus no omentoplasty for esophagogastrostomy after surgery for cancer of cardia and esophagus. AB - BACKGROUND: The standard of care of patients with cancer of cardia and esophagus still remains surgery in early stage. One of the most feared complications after such procedure is anastomotic leak. AIM: We present our experience with omental wrapping of anastomosis (omentoplasty) to decrease the anastomotic leak. SETTINGS AND DESIGN: Retrospective study. MATERIALS AND METHODS: An analysis of 50 consecutive patients, who underwent surgical resection for cancer of cardia and esophagus at BPKMCH, is done. For cancer of esophagus, a 10 cm proximal tumor free margin and for lesions of cardia, at least 5 cm margin was achieved. A 5 cm distal tumor free margin was achieved in each case. A subset of patients was considered for omentoplasty after completion of anastomosis. RESULTS: There were 29 male and 21 female with a mean age of 56.3 years. The average postoperative stay was 13.14 days. The stomach was the organ of substitute in 48 and jejunum in 2 cases. Omentoplasty was done in 37 cases, whereas in 13 cases, no omental wrapping was done. The rate of anastomotic leak was 6%. There was no leak from anastomosis placed at chest, whereas three cases of leak was observed in the anastomosis at the level of neck (P=.013). Overall, there was no leak in omentoplasty group, whereas there were three cases with leak in the group without omentoplasty (P=0.003). CONCLUSION: Omentoplasty should be considered in every case after surgical resection for cancer of cardia and esophagus. PMID- 15659871 TI - Brainstem gliomas--a clinicopathological study of 45 cases with p53 immunohistochemistry. AB - BACKGROUND: Brainstem tumors represent 10% of central nervous system tumors, accounting for 30% of pediatric posterior fossa tumors. AIMS: The aim of this study was to clinicopathologically correlate 45 cases of brain stem gliomas and determine the occurrence and prognostic significance of p53 expression. MATERIALS AND METHOD: 45 cases of brain stem gliomas encountered during a 19-year period. 30 were diagnosed by surgical biopsy and 15 at autopsy. In 25 cases p53 immunohistochemistry (Avidin Biotinylated technique) was performed. The WHO brain tumor classification and Stroink's CT classification were applied. STATISTICAL ANALYSIS USED: Chi square test. RESULTS AND CONCLUSIONS: 51 % of gliomas were observed in the first decade of life. The female to male ratio was 1.04: 1. The commonest presenting features were cranial nerve palsies (33%) and cerebellar signs (29.8%). 55.55% of cases were located in the pons, 31.01% in the medulla and 13.33% in the midbrain. Diffuse astrocytomas were seen in 40 cases (5% were Grade I, 47.5%Grade II, 32.5% Grade III and 15% Grade IV) and pilocytic astrocytomas in 5 cases. Grade IV patients had 2- 3 mitoses /10 high power fields and had a poorer survival rate. Grade II astrocytomas were treated with excision and radiotherapy, while grade III and IV tumors were treated with radiotherapy and chemotherapy (CCNU). Improvement was noted in 20% of patients postoperatively. The outcome was better in patients who were treated surgically. p53 is a frequently mutated gene in brain stem astrocytomas. It was found in 50 % of glioblastoma multiforme, 28.57% of grade III astrocytoma and 12.5% of grade II astrocytoma, while grade 1 astrocytomas failed to express p53 protein. p53 positivity was more in high grade lesions, decreasing significantly in lower grade lesions. PMID- 15659872 TI - Gastrointestinal stromal tumour of the colon presenting with intestinal obstruction. AB - Gastrointestinal Stromal Tumours are the commonest mesenchymal tumors of the gastrointestinal tract, the stomach and small intestine being the favored sites. They rarely occur in the colon and rectum and esophagus. The diagnosis is difficult, especially in the rarer sites, since there are no pathognomic features to suggest GIST on preoperative clinical examination and investigations, and only a detailed histopathological analysis of the specimen reveals their true nature. The case of a young female patient who presented with intestinal obstruction due a GIST of the transverse colon is reported. The relevant literature is briefly reviewed. PMID- 15659873 TI - Early occurrence of acute myeloid leukemia following adjuvant radiotherapy and higher cumulative dose of cyclophosphamide in carcinoma breast. AB - We report a case of cancer breast developing acute myeloid leukemia (AML) within a relatively short interval of two and a half years of her primary treatment. This could be attributed to post operative radiotherapy and a higher cumulative dose of cyclophosphamide (14.4 gm) which had to be given as a part of her combination chemotherapy regimen, initially as adjuvant and then later as salvage chemotherapy. The successful salvage therapy for secondary AML instituted in this case is also discussed. PMID- 15659874 TI - Hemangioma of base of tongue. AB - Although vascular malformations of the tongue comprise a significant portion of head and neck angiodysplastic lesions, hemangioma of base of tongue is rare. We report a case of hemangioma of base of tongue extending to the supraglottis, which necessitated an extended supraglottic laryngectomy. Patient had an uneventful recovery and at three year, follow-up has a normal speech and no difficulty in swallowing or aspiration. More importantly, there was no recurrence of hemangioma or bleeding. Although hemangiomas may be treated by various conservative methods, occasionally patient may require surgical excision as in the present case due to the repeated bleeding episode and difficult access. A high index of suspicion and radiological investigations should be performed if the clinical presentation is atypical for malignancy, as in our case. PMID- 15659875 TI - Combination therapies in spondyloarthropathies. AB - OBJECTIVES: To compare the efficacy and tolerability of single-agent sulfasalazine (SSZ) with combination therapies composed of SSZ and methotrexate (MTX) and SSZ, MTX and hydroxychloroquine (HCQ) in active spondyloarthropathy (SpA) patients with peripheral arthritis. METHODS: One hundred and fifty SpA patients with peripheral arthritis (male/ female: 92/58) who were treated in our clinic between 1994 and 1998 were enrolled in this trial. Patients treated with SSZ alone (1-3 gr/day) were included in Group I, patients treated with combination of SSZ (1-3 gr/day) and MTX (7.5-15 mg/week) in Group II, and patients treated with combination of SSZ (1-3 gr/day), MTX (7.5-15 mg/week) and HCQ (200 mg/day) in Group III. Forty-eight patients in Group I, 45 patients in Group II and III were eligible for statistical analysis at the end of study. RESULTS: The combination of MTX, SSZ, and HCQ, and the combination of MTX and SSZ were more effective regarding the clinical and laboratory parameters than SSZ alone (p<0.05). Moreover, the combination of MTX, SSZ, and HCQ was more effective than the combination of MTX and SSZ (p<0.05). CONCLUSION: Combination therapies seem to be more effective and no more toxic than monotherapy in SpA patients with peripheral arthritis. PMID- 15659876 TI - Direct effects of tumor necrosis factor alpha (TNF-alpha) on L6 myotubes. AB - Tumor necrosis factor (TNF)-alpha is a pleiotropic cytokine responsible for a diverse range of signaling events within cells. We studied direct effects of TNF alpha on skeletal muscle protein content. Incubation of the L6 myotubes with 1 to 10 U/ml of TNF-alpha resulted in a significant increase of total and myofibrillar protein contents as compared with the control. However, incubation with lower (0.001-0.1 U/ ml) or higher (100-300 U/ml) doses of TNF-alpha resulted in a decrease of protein content. These findings suggest that TNF-alpha may elicit both anabolic and catabolic effects on L6 myotubes in a dose dependent manner. The anabolic effect of TNF-alpha was mediated at least in part by mitogen activated protein kinase (MAPK), especially by an extracellular-regulated kinases (ERK). This divergent effect of TNF-alpha may be crucial to elucidate the complexity of TNF-alpha action on the skeletal muscle. PMID- 15659877 TI - Fission yeast epsin, Ent1p is required for endocytosis and involved in actin organization. AB - In this study, we have characterized an essential gene ent1+ encoding fission yeast epsin, which is similar to mammalian and budding yeast endocytic protein epsins. The S. pombe Ent1p contains ENTH (epsin amino-terminal homology) domain at its amino terminus, two copies of a ubiquitin-interacting motif (UIM) immediately carboxyl-terminal to the ENTH domain, three NPF motifs in the carboxyl-terminal half, and the clathrin-binding motif at the carboxyl terminal. When repressed the expression of ent1+ gene, the conditional ent1 gene knockout cells showed a marked defect in internalization of fluorescent dyes, suggesting that Ent1p is essential for endocytosis. Changes in conserved amino acid residues within ENTH domain in ent1 mutant cells revealed temperature-sensitive defect in actin organization and cell morphology. The Ent1p bound PI(4,5)P2 and PI(3,5)P2 immobilized onto nitrocellulose in vitro and also weakly bound PI(3,4)P2, PI(3,4,5)P3, PI4P and PI5P. Surprisingly, the localization of Ent1p-GFP was not affected even in the its3-1 cells, in which the level of PI(4,5)P2 was severely reduced, suggesting that PI(4,5)P2 may not be essential for proper localization of Ent1p at endocytic sites. Our findings indicate that S. pombe Ent1p is an essential component in endocytic process, and involved in actin organization and cell morphogenesis. PMID- 15659878 TI - Immunoglobulin levels and prediction of progression to AIDS in HIV-infected injection drug users. PMID- 15659879 TI - Rhabdomyolysis after simvastatin therapy in an HIV-infected patient with chronic renal failure. AB - We encountered a 49-year-old HIV-infected man with chronic renal insufficiency who developed rhabdomyolysis after treatment with simvastatin. He recovered after initiating hemodialysis and discontinuing oral medications. Rhabdomyolysis most likely resulted from an excessive blood concentration of simvastatin caused by concomitant use of fluconazole in the presence of renal insufficiency. PMID- 15659880 TI - Challenges for HIV vaccine dissemination and clinical trial recruitment: if we build it, will they come? AB - HIV vaccine availability does not guarantee uptake. Given suboptimal uptake of highly efficacious and already accessible vaccines in the United States, low vaccine coverage in the developing world, and the expectation that initial HIV vaccines will be only partially efficacious, the public health community will face formidable challenges in disseminating U.S. Food and Drug Administration (FDA)-approved HIV vaccines. HIV/AIDS stigma, fear of vaccine- induced HIV infection, social side effects of testing HIV-positive, and mistrust of government and research present additional obstacles to HIV vaccine dissemination. Increased risk behaviors because of HIV vaccine availability can undermine the effectiveness of partially efficacious vaccines in reducing HIV incidence. HIV vaccine efficacy trials also face significant challenges in recruitment of sufficient volunteers and possible increases in risk behaviors due to trial participation. Planning and designing interventions to facilitate successful recruitment for large-scale phase 3 efficacy trials is a vital step towards U.S. FDA-approved HIV vaccines. Rather than despair in the face of momentous HIV vaccine dissemination challenges, or presume unrealistically that vaccine uptake will ensue automatically and that risk behavior increases will not occur, let us deem the estimated 10-year window to an approved HIV vaccine as an opportunity to investigate and confront these challenges. A consumer research agenda founded on social marketing principles is needed to facilitate the design of empirically-based interventions tailored to the unique needs and preferences of specific segments of consumers. Social marketing interventions may increase future HIV vaccine uptake and clinical trial participation, and mitigate increases in HIV risk behaviors. PMID- 15659882 TI - Attitudes toward HIV Health Care Providers scale: development and validation. AB - Patient attitudes toward their health care providers can play an important role in determining health behavior change. The frequency of contact with health care professionals and disease stigma makes assessing patients' perception of this relationship of particular interest in an HIV medical population. While past general satisfaction and attitude tools have been used to assess this construct, there is a need for an assessment tool specific to patient attitudes in an HIV setting. This study was designed to validate the Attitudes toward HIV Health Care Provider scale (AHHCP) in an HIV medical population. Principal components analysis of the AHHCP yielded a two-factor structure accounting for 53.3% of the total variance in attitudes toward health care providers. The two factors represented items concerning Professionalism and Emotional Support. The AHHCP was found to have good internal consistency (0.92) and convergent validity with a measure of patient satisfaction (r = 0.59). The results of the present study suggest that the AHHCP is a reliable and valid instrument for use in assessing patient attitudes toward their health care providers. PMID- 15659881 TI - Initiation of regular marijuana use among a cohort of women infected with or at risk for HIV in the Women's Interagency HIV Study (WIHS). AB - Our study sought to determine the incidence of weekly marijuana use among HIV infected and uninfected women, to identify correlates of weekly marijuana use, and to test its association with stage of HIV disease and type of HIV treatment received. A total of 2059 HIV-positive and 569 HIV-negative women from 6 sites were recruited between 1994 and 1995 and followed through 2000. After excluding women who reported weekly marijuana use at baseline, 2050 women were included in the analysis. The incidence rate for initiating marijuana was calculated and survival analysis was performed to determine the correlates of initiating weekly marijuana use. Three hundred and three women initiated weekly marijuana use within 5.5 years of the baseline visit, yielding a cumulative incidence (CI) of 14.8%. There was no significant difference in weekly marijuana use initiation between HIV-infected (CI = 14.5%) and HIV-uninfected women (CI = 16.0%). Younger age and having more sex partners was associated with incident weekly marijuana use among both infected and uninfected women. While undetectable viral load was associated with lower incidence rate (p < 0.001, RH = 0.44) and wasting syndrome with higher incidence (p < 0.01, relative hazard [RH] = 3.1), CD4 count was not. Compared to receiving no AIDS treatment at all, women who received basic combination antiretroviral therapy had significantly higher incidence of weekly marijuana use (p < 0.001, RH = 1.93), while highly active antiretroviral therapy (HAART) receivers had significantly lower incidence (p < 0.001, RH = 0.24). In summary, among HIV-infected women, the incidence of weekly marijuana use was associated with only one marker of HIV disease stage and HAART was associated with lower initiation rate of weekly marijuana use. PMID- 15659883 TI - Factors related to and consequences of adherence to antiretroviral therapy in an ambulatory HIV-infected patient cohort. AB - In a confidential medication adherence questionnaire completed by 255 participants in the HIV Outpatient Study (HOPS) between March and November 1999, 33% reported skipping antiretroviral doses within the previous 3 days. The respondents, with a median age of 41, were predominantly male (86%), white (62%), and highly educated (33% had some post-high school training but no college degree and 39% had a college degree; only 11% had less than a high school diploma). Twenty-one percent had a history of injection drug use, 12% were unemployed, and 18% had Medicaid insurance. Questions about difficulty taking antiretroviral medications or drug holidays identified an additional 16% of patients experiencing adherence problems and explained significantly more of the failure to achieve undetectable viral loads than simply querying about skipped doses. PMID- 15659884 TI - Multidisciplinary HIV adherence intervention: a randomized study. AB - Maintaining greater than 95% adherence to antiretroviral medication is necessary in order to have the greatest therapeutic impact on HIV infection. Furthermore, evidence suggests that adherence rates of between 70% and 89% are significantly associated with viral rebound and the development of drug resistance. Adherence rates at and above the 95% level are difficult for patients to achieve and maintain. Our aim was to determine if an adherence intervention could improve adherence among patients attending an ambulatory care clinic at a large public hospital. The intervention was delivered by a multidisciplinary team of health care professionals and consisted of education coupled with the provision of devices designed to assist patient memory and adherence. A crucial component of the intervention consisted of the identification of patient specific barriers to adherence and the development of strategies to circumvent these problems. Adherence was assessed using patient self-report over the past 4, 7, and 28 days and by calculation of the Morisky score. The study was conducted as a randomised controlled trial using the stepped wedge design with a total of 68 subjects randomised to receive the intervention over a 20-week period. Adherence before and after the intervention formed the analysis. There was a significant decrease in the number of missed doses over the past 4 (1.9 to 1.0, p < 0.001), 7 (3.0 to 1.8, p < 0.001) and 28 (7.4 to 4.2, p < 0.001) days and a decrease in the Morisky score, indicating an improvement in medication taking behaviour (1.3 to 0.5 p < 0.001). PMID- 15659885 TI - Formal approval for Fuzeon. PMID- 15659886 TI - TB and AIDS cases rise in NYC. PMID- 15659890 TI - Factors affecting blood loss during percutaneous nephrolithotomy: prospective study. AB - BACKGROUND AND PURPOSE: Bleeding is a major concern during percutaneous nephrolithotomy (PCNL), especially with the use of multiple tracts. This prospective study aimed to identify factors affecting blood loss during PCNL. PATIENTS AND METHODS: Data were collected prospectively from 236 patients undergoing 301 PCNL procedures at our institute since June 2002. Blood loss was estimated by the postoperative drop in hemoglobin factored by the quantity of any blood transfusion. Various patient-related and intraoperative factors were assessed for association with total blood loss or blood transfusion requirement using stepwise multivariate regression analysis. RESULTS: The average hemoglobin drop was 1.68 +/- 1.23 g/dL. Stepwise multivariate regression analysis showed that the occurrence of operative complications (P < 0.0001), mature nephrostomy tract (P < 0.0001), operative time (P < 0.0001), method of access guidance (fluoroscopy v ultrasound) (P = 0.0001), method of tract dilatation (P = 0.0001), multiple (> or =2) tracts (P = 0.003), size of the tract (P = 0.001), renal parenchymal thickness (P = 0.05), and diabetes (P = 0.05) were significant predictors of blood loss. The overall blood transfusion rate for all patients was 7.9%. Preoperative hemoglobin, multiple tracts, stone size, and total blood loss were significant in predicting perioperative blood transfusion requirement. Factors such as age, hypertension, renal insufficiency, urinary infection, the degree of hydronephrosis, stone bulk, and the function of the ipsilateral renal unit did not have any effect on the blood loss. Technical factors such as the operating surgeon and the calix of entry also did not affect the blood loss. CONCLUSIONS: Diabetes, multiple-tract procedures, prolonged operative time, and the occurrence of intraoperative complications are associated with significantly increased blood loss. Atrophic parenchyma and past ipsilateral intervention are associated with reduced blood loss. On the basis of this evidence, maneuvers that may reduce blood loss and transfusion rate include ultrasound-guided access, use of Amplatz or balloon dilatation systems, reducing the operative time, and staging the procedure in cases of a large stone burden or intraoperative complications. Reducing the tract size in pediatric cases, nonhydronephrotic systems and those with a narrow infundibulum, and secondary tracts in a multiple tract procedure may also reduce blood loss during PCNL. PMID- 15659892 TI - Laparoscopic radical prostatectomy: is intact organ removal attainable? Study of margin status. AB - PURPOSE: To determine the initial oncologic results (pathology) of specimens removed by laparoscopic radical prostatectomy (LRP) by examining the surgical margins. PATIENTS AND METHODS: The 70 consecutive LRP procedures performed for clinically localized prostate cancer at Eastern Virginia Medical School from April 2001 to November 2002 were reviewed for preoperative and important intraoperative variables. The initial histopathology report and a prospective review by a single genitourinary pathologist for margin status as well parenchymal exposure of benign glands were assessed. Postoperative prostate specific antigen (PSA) levels were noted. RESULTS: The clinical stage distribution was as follows: T1c = 59, T2a = 10, and T2b = 1. The preoperative median PSA value was 6.96 ng/mL. The mean operative time was 307 minutes. The mean estimated blood loss was 298 mL. The prospective pathologic review results were as follows: 1 pT0 (1.4%), 60 pT2 (85.7%), and 9 pT3 (12.8%). The overall positive surgical-margin rate was 15.7%. Of those patients with pT2 disease; 8 specimens (13.3%) had a positive margin, whereas 33% of patients with pT3 disease had a positive surgical margin. Parenchymal exposure of benign glands on the inked surface was recognized in 8 patients (13.3%). Data from PSA assays 1 month postoperatively were available in 69 patients. Serum PSA was undetectable (< or =0.1 ng/mL) in 67 men (97%). CONCLUSION: An LRP can offer surgical margins comparable to those of open procedures series reported in the literature. Long term progression and survival outcome data are necessary before this procedure should be offered as a replacement for open prostatectomy. PMID- 15659893 TI - Performance and durability of leading flexible ureteroscopes. AB - PURPOSE: The role of ureteronephroscopy in the diagnosis and treatment of upper tract disease has expanded dramatically in recent years with technological advances. In a study involving 14 urologists, we measured the practical performance and durability of the most common commercially available flexible ureteroscopes. PATIENTS AND METHODS: Six commercially available flexible ureteroscopes (Circon-ACMI DUR 8, ACMI DUR 8-Elite, Storz 11274AA, Wolf 7325.172 7.5Fr, Wolf 7330.072-9.0 Fr, Olympus URF-P3) were used consecutively 102 times over a 7-month period. Comparative data, including preoperative and postoperative maximal deflection angles and luminosity, were accumulated, as well as a survey of insertion methods, irrigation methods, instruments used in the working channel, visibility, maneuverability, and overall satisfaction. Maintenance records were analyzed to evaluate the need or reason for repair. RESULTS: Overall satisfaction, visibility, luminosity, and maneuverability were similar for all instruments. The ACMI DUR-8 Elite scored slightly higher than the others, but the difference was not statistically significant. Ureteroscopes were used an average of 10 to 34 times between breakages. The ACMI DUR-8 and DUR-8 Elite instruments had a higher durability score than the others, but with significantly more uses. Channel perforation by a laser fiber was the predominant cause of ureteroscope breakage. CONCLUSIONS: All ureteroscopes performed similarly by objective and subjective criteria. The largest and newest instruments were significantly more durable than the smaller ones, even in the hands of numerous endoscopists. This finding has significant implications for the cost-effectiveness of the various ureteroscopes. PMID- 15659891 TI - Double-blind randomized controlled trial assessing the safety and efficacy of intravesical agents for ureteral stent symptoms after extracorporeal shockwave lithotripsy. AB - BACKGROUND AND PURPOSE: Ureteral stents are a significant source of pain and discomfort for many urologic patients. A novel approach to addressing this problem is the intravesical instillation of a selected pharmacologic agent after stent insertion. The purpose of this study was to assess the safety and efficacy of intravesical instillation of various agents in reducing ureteral stent associated discomfort in patients requiring a stent after extracorporeal shockwave lithotripsy (SWL). PATIENTS AND METHODS: In this double-blind prospective trial, 42 patients were randomized to receive intravesical instillation of one of three agents (oxybutynin, alkalinized lidocaine, or ketorolac) or a control solution (0.9% sodium chloride) immediately after stent insertion at time of SWL. The four groups of patients were demographically similar. Preoperative, intraoperative, and postoperative data were collected prospectively and analyzed statistically. The primary outcome measure was reduction in ureteral stent symptoms, and the secondary outcome measure was the safety of intravesical instillation of each agent through assessment of drug related adverse events. RESULTS: There were no intraoperative or postoperative complications, nor were there any serious side effects attributable to any of the intravesically instilled agents. There was a statistically significant decrease in stent-related discomfort at the 1-hour time point in the group of patients who received intravesical ketorolac compared with the control group. CONCLUSIONS: Intravesical instillation represents a novel approach to the problem of ureteral stent-related discomfort. From our results, ketorolac appears to be the most effective intravesical agent in reducing stent-related patient discomfort, and we have established that intravesical instillation of ketorolac is safe in humans. PMID- 15659894 TI - Fluid absorption during ureterorenoscopy. AB - BACKGROUND AND PURPOSE: Ureterorenoscopy (URS) is a common minimally invasive diagnostic and therapeutic modality for ureteral and renal pathology. Fluid absorption during routine URS has not been studied prospectively, despite the fact that fluid absorption during other endoscopic urologic procedures can be substantial. PATIENTS AND METHODS: During URS in 15 male and 8 female patients with a mean age of 54 years (range 19-81 years), volumetric balance was performed by measuring all fluids instilled into the urinary tract (irrigation fluid and contrast medium) and fluids collected from the urinary tract (irrigation fluid, contrast medium, and urine output) and by estimating urine output from creatinine concentration in the urine and in the fluids collected from the urinary tract. Fluids from the urinary tract were assessed by measuring drainage fluid and the preoperative and postoperative weights of the drapes and bedsheets. Of the procedures, 11 were right-sided and 12 were left-sided. The indications for URS were urolithiasis (N = 18) and diagnosis (hematuria in 2, ureteral or renal filling defect in 2, flank pain and hydronephrosis in 1). RESULTS: The mean total operative time was 55 minutes (range 20-95 minutes), and the mean URS time was 37 minutes (range 8-83 minutes). The mean volume of irrigation fluid used was 2531 mL (range 552-5580 mL). The mean estimated urine output during the procedure was 62 mL (range 7-201 mL). The mean estimated systemic fluid absorption during URS was 54 mL (range 4-137 mL). There were two intraoperative complications (ureteral perforations) but no postoperative complications. CONCLUSIONS: Routine URS is associated with minimal systemic fluid absorption, even if ureteral perforation occurs. Estimated absorption of as much as 137 mL was seen; however, evaporative losses and unaccounted-for losses of fluid likely account for a substantial portion of this fluid discrepancy. This result suggests that irrigation with fluids other than normal saline, such as sterile water, during URS is likely safe. PMID- 15659895 TI - No panacea for drainage after percutaneous nephrolithotomy. AB - BACKGROUND AND PURPOSE: The optimal postoperative drainage system for patients undergoing percutaneous nephrolithotomy (PCNL) has not yet been determined. Each of the different tubes available has theoretical advantages and disadvantages. The objective of this study was to determine if any one system of post-PCNL drainage has any significant advantage over the others. PATIENTS AND METHODS: Sixty patients were randomized to either a 24F Re-entry tube (Group 1), an 8F pigtail catheter (Group 2), or a double-J stent (Group 3) (N = 20 for all groups). The double-J stent patients also had an 18F Councill catheter as a nephrostomy tube, which was removed on the morning of postoperative day 1. The three groups were well-matched for age, stone size, operative time, and mean number of percutaneous tracts (P > 0.05 for all). Data collected included intraoperative findings, postoperative analog pain scores and narcotic usage, presence of extravasation on postoperative nephrostogram, presence of perinephric fluid on postoperative ultrasound scans, length of stay, and whether leakage from the nephrostomy site persisted more than 48 hours after tube removal. RESULTS: There were no statistically significant differences among the three groups in terms of change in hematocrit or number of patients requiring blood transfusion; in the incidence of tube blockage, extravasation, or presence of perinephritic fluid; or in complication rates, length of hospital stay, or persistent leakage after tube removal. There was a strong trend to less postoperative leakage with the 8F pigtail than the Reentry tube (P = 0.05). Postoperative analog pain scores and narcotic usage were equivalent for all three groups. Patients Groups 1 and 2 were free of all tubes sooner than patients in Group 3 (P < 0.001). CONCLUSIONS: This study did not demonstrate an overwhelming advantage of any one drainage system over the others. All three systems were equally tolerated by patients. There may be less chance of prolonged nephrostomy-site leakage with an 8F pigtail catheter. PMID- 15659896 TI - Laparoscopic pyeloplasty: history, evolution, and future. AB - Ureteropelvic junction (UPJ) obstruction is characterized by a functionally significant impairment of urinary transport caused by obstruction in the area where the ureter joins the renal pelvis. The majority of cases are congenital; however, acquired conditions at the level of the UPJ may also present with symptoms and signs of obstruction. Until recently, open pyeloplasty and endoscopic techniques have been the main surgical options with the intent of complete excision or incision of the obstruction. The introduction of laparoscopy has allowed minimally invasive reconstructive surgery that mirrors open surgical techniques. In the hands of experienced surgeons, laparoscopic pyeloplasty offers a less invasive alternative to open surgery with decreased morbidity, shorter hospital stay, and faster convalescence. During the last decade, laparoscopic pyeloplasty for the treatment of congenital or acquired UPJ obstruction has garnered much interest, but, as this procedure is technically challenging, it is being performed only at selected medical centers by surgeons with advanced laparoscopic training. This review describes the early results, ongoing evaluation, and future role for this novel surgical procedure. PMID- 15659897 TI - Application of strict radiologic criteria to define success in laparoscopic pyeloplasty. AB - PURPOSE: To review our experience and utilize rigorous radiologic criteria to establish success in laparoscopic pyeloplasty. PATIENTS AND METHODS: The hospital records of consecutive adult patients who underwent laparoscopic pyeloplasty for ureteropelvic junction (UPJ) obstruction during a 5-year period at our institution were reviewed. We identified 47 candidates for study inclusion. With one open conversion secondary to excessive fibrosis and one patient requiring repeat laparoscopic pyeloplasty for recurrence, 46 patients with 47 renal units were included for analysis. Thirteen renal units had prior surgery for UPJ obstruction. Preoperative and postoperative symptomatology were compared to determine subjective outcome. Mercaptoacetyltriglycine (MAG3) nuclear renography was performed preoperatively and postoperatively to assess differential renal function (DRF) and obstruction, defined as a T1/2 -20 minutes, and evaluate objective success. There were 34 dismembered and 13 Y-V pyeloplasties. RESULTS: The mean operative time was 341.6 minutes (range 200-717 minutes). The mean preoperative T1/2 was 48.4 minutes (range 14 minutes-xc), with a mean DRF of 39.8% (range 22%-60%). The mean postoperative T1/2 was 9.32 minutes (range 2-27 minutes), and the mean DRF was 39.5% (range 7%-57%). The average length of hospital stay was 2.25 days (range 1-3 days). At a mean follow-up of 19.93 months (range 2-55 months), the subjective success rate was 95.7%, and the objective success rate was 93.6%. CONCLUSION: Even when using strict radiologic criteria to define success, laparoscopic pyeloplasty is found to be similar to open surgery in its efficacy for the correction of adult UPJ obstruction. PMID- 15659898 TI - Ureteropelvic junction obstruction: endourologic relocation of laterally inserting ureters. AB - PURPOSE: To describe a technique of endourologic management for ureteropelvic junction (UPJ) obstruction associated with lateral insertion of the ureter. PATIENTS AND METHODS: Among 57 patients with primary UPJ obstruction selected to undergo retrograde endopyelotomy, 2 patients (3.5%) demonstrated a nondilated ureter inserting laterally along a hydronephrotic renal pelvis on retrograde pyelography. Using the hot-wire balloon catheter in one and a holmium laser inserted ureteroscopically in one, retrograde endopyelotomy was performed with the incision directed medially in order to marsupialize the ureter into the renal pelvis and to relocate the UPJ to the dependent portion of the medial aspect of the renal pelvis. RESULTS: The hospital stay for both patients was 1 night, and there were no complications. The stent was removed 4 or 6 weeks postoperatively. Intravenous urography 6 weeks thereafter demonstrated relocation of the UPJ to a medial and dependent position, associated with prompt drainage of contrast medium and decreased pelvocaliectasis compared with preoperative studies. Both patients are asymptomatic. CONCLUSIONS: In cases of primary UPJ obstruction associated with lateral insertion of the ureter, the UPJ can be relocated to the medial and dependent portion of the renal pelvis using minimally invasive endourologic techniques. The dictum of performing a endopyelotomy incision laterally does not hold true in all patients and may compromise functional outcomes in cases where the ureteral insertion is nondependent and laterally positioned. PMID- 15659899 TI - Laparoscopic dismembered pyeloureteroplasty: a single institution's 3-year experience. AB - PURPOSE: To review our most recent experience with laparoscopic dismembered pyeloureteroplasty (LDP) as treatment for ureteropelvic junction (UPJ) obstruction. PATIENTS AND METHODS: All patient charts from March 1, 2000, through March 1, 2003, were reviewed to identify individuals who had undergone LDP. Twenty-two consecutive procedures were performed on 21 patients. The average patient age was 37 years (range 6-77 years). Multiple data points were collected, including patient age, presenting symptoms, degree of hydronephrosis, presence of crossing vessels, operative time (cystoscopy, retrograde pyelography, stent placement, and LDP), estimated blood loss (EBL), length of hospital stay (time to discharge home after preoperative admission), parenteral analgesic use, complications, and results of follow-up imaging studies and symptoms (including an analog pain scale). Mean and range values for quantitative data are reported. RESULTS: Twelve right and ten left renal units were operated on; in 12 UPJs, a crossing vessel was causative of the obstruction. The mean operative time was 291 minutes (range 180-500 minutes) and the mean intraoperative EBL was 63 mL (range 20-100 mL). There were no conversions to open surgery. The mean hospital stay was 40 hours (range 24-120 hours). The average number of parenteral analgesic doses before conversion to oral narcotics was 5 (range 1-10). Three patients had successful pyelolithotomies at the time of reconstruction with a 100% stone-free rate on postoperative imaging studies. The average time of follow-up evaluation was 199 days (range 31-705 days). All patients either had serial ultrasound examinations, diuretic intravenous urograms, or both for follow-up evaluation, which demonstrated improvement in hydronephrosis and renal drainage. All patients with pain preoperatively had resolution of their symptoms postoperatively. There were no intraoperative complications, seven early postoperative complications (three urinary tract infections, two hospital readmissions for ileus, one urine leak, and one delayed hemorrhage, presumably from a trocar site). CONCLUSIONS: Laparoscopic dismembered pyeloureteroplasty is safe and effective and associated with minimal morbidity. PMID- 15659900 TI - Hand-assisted laparoscopic nephrectomy for inflammatory renal conditions. AB - PURPOSE: Laparoscopic surgery for large renal lesion or kidneys with chronic inflammation has proved to be technically challenging. Hand-assisted laparoscopic surgery might be useful in these complex cases, as it provides surgeons the benefits of tactile feedback, digital retraction, and facilitated dissection of the renal hilar vessels. PATIENTS AND METHODS: Twenty-two patients undergoing hand-assisted laparoscopic (HAL) nephrectomy for benign conditions were compared with patients who underwent HAL radical nephrectomy during the same period. The demographic data, laterality, operative time, estimated blood loss, conversion rate, length of stay, histopathology findings, morbidity, and mortality were reviewed. RESULTS: The main indications for surgery were chronic inflammation and xanthogranulomatous pyelonephritis. Twenty patients had unilateral nephrectomy (10 each on the right and left), and two patients had bilateral nephrectomy. The mean operative times for unilateral and bilateral nephrectomy were 163 minutes (range 55-261 minutes) and 265 minutes (range, 238-291 minutes), respectively. Nine patients (45%) with inflammation had complications (15% major and 30% minor). The mean length of hospitalization for patients undergoing HAL nephrectomy was 7.2 days (range 2-35 days). The patients with inflammatory pathology had longer mean operative times, higher estimated blood loss, longer hospital stay, and higher morbidity than patients undergoing radical nephrectomy. CONCLUSION: Compared with standard laparoscopy, the hand-assisted approach has been reported to reduce operative times and increase safety. The advantages of minimally invasive surgery, such as reduced analgesia, shorter hospital stay, and faster return to normal activity, appear to be similar to those in patients undergoing a pure laparoscopic nephrectomy. Compared with radical nephrectomy for renal tumor, HAL simple nephrectomy can often be more challenging and associated with greater morbidity. For both the community urologist as well as an experienced laparoscopist, this approach is useful in handling these challenging cases. PMID- 15659901 TI - Hand-assisted laparoscopic renal surgery: hand-port incision complications. AB - BACKGROUND AND PURPOSE: Hand-assisted laparoscopic (HAL) renal surgery is an accepted standard of practice. We report our complications at the hand port-site incision in various HAL urologic procedures. PATIENTS AND METHODS: Data and follow-up were complete for 54 patients who underwent HAL nephrectomy. HAL partial nephrectomy, or HAL nephroureterectomy at our institution from October 2001 to April 2003. A retrospective review of the patients' charts was performed to identify the incidence of postoperative hand port-site complications. The mean patient age was 59 years, and the mean follow-up was 5.4 months. RESULTS: Five complications were observed in four patients (9.3%). One patient had a superficial wound infection (1.9%) and an incisional hernia (1.9%) that was surgically repaired. One patient had a superficial incisional breakdown that healed well after conservative measures. One obese patient with severe chronic obstructive pulmonary disease eviscerated through his hand-port incision after a coughing spell. This was repaired operatively, and the patient healed well with the aid of an abdominal binder. One patient developed an enterocutaneous fistula at her hand-port site that healed well after both operative and conservative management. CONCLUSIONS: The technique of HAL renal surgery is safe and effective. Complications associated with the hand-port incision exist, and as they become better defined, the surgeon may take extra steps toward their recognition and prevention. PMID- 15659902 TI - Holmium laser percutaneous nephrolithotomy using a unique suction device. AB - BACKGROUND AND PURPOSE: Most percutaneous nephrolithotomies are performed with electrohydraulic, ballistic, or ultrasonic lithotripsy, with holmium laser use reserved for ureteroscopy or ureterorenoscopy. We evaluated the feasibility of holmium laser use in percutaneous nephrolithotomy with the assistance of a unique suction device. PATIENTS AND METHODS: The charts of 71 consecutive patients undergoing 90 percutaneous procedures were reviewed. A unique suction device and the holmium laser were used in nearly half the patients. The average stone burden was 3.25 cm. Use of holmium laser, energy settings, stone burden, operative time, complications, and stone-free rate were evaluated. RESULTS: The average operative time was 167 minutes with a complication rate of 14%. A stone-free rate of 83% was achieved with the laser. CONCLUSIONS: With the assistance of a unique suction device, the holmium laser can, in the appropriate situation, be an efficient and successful lithotrite with percutaneous nephrolithotomy. PMID- 15659903 TI - Laparoscopic nephron-sparing surgery for a renal mass: 1-year minimum follow-up. AB - BACKGROUND AND PURPOSE: Because of the explosion of laparoscopy in urology coinciding with the excellent results of open nephron-sparing surgery (NSS) for small renal masses, laparoscopic NSS has become an alternative to an open surgical approach. We report our results with laparoscopic NSS in patients who have had a minimum of 1 year of follow-up. PATIENTS AND METHODS: All consecutive laparoscopic partial nephrectomies from November 1998 through February 2002 were assessed. The mean patient age, body mass index, and American Society of Anesthesiology score were 57.1 years, 28.5 cm/kg2, and 2.0, respectively. The procedures were performed using hand-assisted (N = 28) or standard (N = 12) laparoscopic techniques. Hospital records were reviewed in order to obtain operative, perioperative, and follow-up data. RESULTS: The median operating room time, estimated blood loss, and hospital stay were 184 minutes, 300 mL, and 2.0 days, respectively. No patients were converted to an open surgical procedure. Four patients (10%) required a blood transfusion, and one (2.5%) had a postoperative urinoma. The mean tumor size was 2.3 cm. Twenty-nine lesions were renal-cell carcinoma, and 11 were benign. With a mean CT scan follow-up of 100.0 weeks, there has not been any recurrence of renal-cell carcinoma. CONCLUSION: Laparoscopic NSS can be performed with acceptable complication rates, which will continue to decrease as newer methods of controlling hemostasis are developed. Although follow-up is fairly short, no renal-cell carcinoma recurrences have appeared. At this point in time, the oncologic efficacy of a laparoscopic approach appears to mirror that of the open surgical technique. PMID- 15659904 TI - Endoscopic treatment of bladder calculi in post-renal transplant patients: a 10 year experience. AB - BACKGROUND AND PURPOSE: Urinary calculus formation following renal transplantation is an uncommon phenomenon. As a result of the growing number of renal transplants performed and the greater graft survival, there has been increased awareness of transplant-related complications, one of which is calculus formation. We report our experience in the management of bladder calculi after renal transplantation. PATIENTS AND METHODS: We retrospectively reviewed the charts of 500 consecutive renal transplant patients from 1992 through 2002 and encountered 7 who had bladder calculi postoperatively. Ureteroneocystostomy had been performed using polyglactic acid suture. Bladder calculi were treated endoscopically by litholapaxy, electrohydraulic lithotripsy (EHL), or holmium:YAG laser lithotripsy. RESULTS: Three calculi were found incidentally at the time of stent removal, and the others were associated with hematuria (43%), urinary tract infection (14%), or irritative voiding symptoms (14%). Eighty-six percent of the calculi were close to the allograft ureteral orifice. CONCLUSION: While various forms of lithotripsy were employed in treating bladder calculi, Hol:YAG laser lithotripsy appeared to be both efficacious and safe. Both EHL and litholapaxy were complicated by mucosal bleeding necessitating Bugbee fulguration. Holmium:YAG laser lithotripsy was not associated with mucosal bleeding, and fulguration was not required near the allograft ureteral orifice. In addition, ureteral stenting is not required. Bladder calculi may form over both absorbable and nonabsorbable suture material, and the ideal suture for the ureteroneocystostomy has yet to be found. Until then, the Hol:YAG laser should be the lithotrite of choice for bladder calculi following renal transplantation. PMID- 15659905 TI - Holmium:YAG laser enucleation of the prostate: multimedia description from a trainee's perspective. AB - Adoption of holmium laser enucleation of the prostate has been limited by its difficult learning curve. To enhance training, digital video clips from four procedures were captured in MPEG-2 format and compiled into a PowerPoint file with professional narration. The two surgical techniques used--the two-lobe and three-lobe methods--are described with a focus on the technical details and common mistakes. PMID- 15659906 TI - Holmium laser enucleation of the prostate in critically ill patients with technique modification. AB - BACKGROUND AND PURPOSE: The use of the holmium:YAG laser for treating benign prostatic hyperplasia has developed substantially over the last decade. We present our experience with holmium laser enucleation of the prostate (HoLEP) in critically ill patients. PATIENTS AND METHODS: From January 1999 to May 2003, 40 critically ill patients with a mean age of 64.7 years (range 48-87 years) underwent HoLEP. Critically ill was defined as major comorbidity, including coronary and peripheral vascular disease, renal transplantation/immunosuppression, postradiation bleeding and sloughing, end stage renal disease, advanced diabetes, severe anemia (hemoglobin <9.0 g/L), anticoagulant dependency, or bleeding diathesis. Preoperative and postoperative changes in hemoglobin (Hb) and sodium (Na), transfusion rates, and urethral catheter time were evaluated. This is a one-time instrument-insertion technique with the use of a large fiber and modification of the median lobe resection. RESULTS: There were no significant changes in Hb (8.7 g/L to 8.5 g/L) or Na (135 mmol/L to 137 mmol/L). One patient was returned to the operating room for bleeding, and this and one other patient required postoperative blood transfusions. The urethral catheter was removed within 36 hours in 34 patients, and all patients were catheter free within 4 weeks of surgery. CONCLUSION: In our experience, HoLEP can be performed safely and effectively in patients who are critically ill. PMID- 15659907 TI - Recognized Fellowship Programs in Endourology. PMID- 15659908 TI - Corporate-Sponsored Programs in Endourology. PMID- 15659909 TI - Upcoming Meetings and World Congress Site. PMID- 15659910 TI - Management of caliceal diverticula. AB - Although most caliceal diverticula are asymptomatic, they can be associated with pain, and concomitant calculi are common. When treatment is indicated, a variety of surgical options are available, although the management is often a challenge even to the most experienced surgeon. We review the indications, surgical options, and outcomes for the treatment of caliceal diverticula. PMID- 15659911 TI - Microlaparoscopy. AB - Laparoscopic surgeons have made significant efforts to minimize the trauma induced by their procedures. One of the newest methods is the use of mini and micro instruments. This work is reviewed here. PMID- 15659912 TI - Review of laser fibers: a practical guide for urologists. AB - Holmium:YAG lithotripsy requires transmission of optical energy using laser fibers. Optical fibers may be subjected to severe angular deflection and bending during flexible nephroscopy or flexible ureteronephroscopy. Irradiation of a deflected fiber in a tight bending radius may produce fiber failure and consequent irradiation of the ureteroscope, with risk of instrument damage and patient injury. We review the physics and engineering aspects of optical fibers to explain why fibers fail. PMID- 15659913 TI - Erbium:YAG laser lithotripsy using hybrid germanium/silica optical fibers. AB - BACKGROUND AND PURPOSE: Previous studies have demonstrated that the erbium:YAG laser is two to three times more efficient for laser lithotripsy than the holmium:YAG laser. However, the lack of a suitable optical fiber delivery system remains a major obstacle to clinical application of Er:YAG laser lithotripsy. This paper describes the initial testing of a hybrid germanium oxide/silica optical fiber for potential endoscopic use with the Er:YAG laser. MATERIALS AND METHODS: Er:YAG laser radiation with a wavelength of 2.94 microm, a pulse energy of 10 to 600 mJ, a pulse length of 220 microsec, and pulse-repetition rates of 3 to 10 Hz was focused into either 350- or 425- microm-core hybrid germanium/silica fibers in contact with human uric acid or calcium oxalate monohydrate stones. RESULTS: Average Er:YAG pulse energies of 157 +/- 46 mJ (66 J/cm(2)) (N = 8) were delivered at 10 Hz through the 425-microm hybrid fibers in contact with urinary stones before fiber damage was observed. A maximum pulse energy of 233 mJ (98 J/cm(2)) was also measured through the hybrid fiber in contact with the stones. These values are significantly greater than the stone ablation thresholds of 15 to 23 mJ (6-10 J/cm(2)) and the fiber damage thresholds measured for germanium oxide, 18 +/- 1 mJ (13 J/cm(2)), and sapphire, 73 mJ (51 J/cm(2)), optical fibers during Er:YAG laser lithotripsy (P < 0.05). CONCLUSIONS: A prototype hybrid germanium/silica optical fiber demonstrated better performance than both germanium oxide and sapphire fibers for transmission of Er:YAG laser radiation during in vitro lithotripsy. PMID- 15659914 TI - Feedback from operative performance to improve training program of laparoscopic radical prostatectomy. AB - PURPOSE: We report our preliminary experience identifying factors that contribute to differences in operative performance in laparoscopic radical prostatectomy (LRP) between trainees and experienced laparoscopic urologists based on objective and subjective feedback. MATERIALS AND METHODS: Between December 2001 and May 2002, six trainees at the Institut Montsouris and Ecole Europeenne de Chirurgie, Paris, were evaluated to identify the factors contributing to differences in operative performance in LRP between them and experienced laparoscopic urologists, both objectively and subjectively. Objective evaluation was done by an independent observer who was well versed in laparoscopic surgery, while all the trainees evaluated themselves subjectively. All the trainees had in vitro, in vivo, and didactic training sessions during the evaluation period. RESULTS: The following deficiencies were noted objectively in the trainees compared with experienced laparoscopic urologists: (1) lack of perfect knowledge of each step, (2) lack of synchronized movements of the nondominant hand; and (3) easy physical fatigue. The trainees took a long time to dissect the vas deferens, seminal vesicles, and bladder neck because they did not have perfect knowledge of each step. It was difficult for them to pass the needle for ligation of dorsal venous complex because they tried to hold the needle far away from the needle tip at the wrong angle, and the direction of movement was not correct. For the urethrovesical anastomosis, they struggled to find the right angle to pass the needle on the urethral side. CONCLUSION: Lack of synchronized movement of the nondominant hand, easy physical fatigue, and lack of perfect knowledge of operative steps are crucial limitations for live operative performance of trainees in LRP. Incorporation of intensive well-planned in vitro training into the curriculum to correct these deficiencies and feedback from the operative performance can make a significant contribute to shortening the learning curve. PMID- 15659915 TI - Hand-assisted laparoscopic nephrectomy: the transfer of experience to a new academic center. AB - BACKGROUND AND PURPOSE: Hand-assisted laparoscopic nephrectomy (HALN) has become widely used for the management of localized renal masses and for simple nephrectomy [corrected] Centers of excellence have slowly disseminated this surgical approach throughout academic institutions and private practices. The transfer of this technique to inexperienced surgeons and centers has not been well studied. We examined our outcomes for HALN with an experienced surgeon (DMA) [corrected] at a new academic center. We also examined the effectiveness of the transfer of these techniques as trainees go out into practice [corrected] PATIENTS AND METHODS: A total of 85 hand-assisted laparoscopy procedures were performed between September 2001 and August 2003 of which 61 were HALN. Four fellows and eight chief residents, under the guidance of one attending surgeon (DMA), performed all HALN procedures. Parameters measured included patient age, ASA score, body mass index, operative time, estimated blood loss, number of trocars used, time to oral intake, analgesics required, length of stay, complications, and tumor size. The average patient age was 57.4 years (range 26 87 years) and the mean ASA score 2.5 (1-4). The mean BMI was 28.3 (range 20-46) [corrected] There was a slight predominance of right-sided lesions. In addition to evaluating our early results with HALN, a questionnaire was sent to all graduates of our program starting 2 years prior to the arrival of DMA to assess the application of laparoscopy to their practices [corrected] RESULTS: All cases were completed without open conversion. The total operative time averaged 184 [corrected] minutes (range 67-257 [corrected] minutes), with 80% of patients requiring two trocars. The average blood loss was 136 [corrected] ml (range 25 700 mL), but only one patient required transfusion postoperatively [corrected] The mean time to oral intake was 17.1 hours (range 1.5-240 hours), the average length of stay was 4.3 days (range 1-28 days), and total narcotic requirements averaged 111 mg of morphine sulfate equivalents (range 6.7-519 mg). Sixty-six percent of the procedures were performed for malignancy. The average tumor size in these cases was 3.9 cm (range 1-12 cm). There was one death, in an 80-year-old patient who had a bowel injury necessitating re-exploration and bowel resection. This patient had a postoperative myocardial infarction and died. Two patients developed postoperative hernias at their hand-port site. Other significant [corrected] complications included diaphragmatic [corrected] injury (repaired laparoscopically), one [corrected] pulmonary embolus, two cases of pancreatitis, and one case of pneumonia. Three patients experienced postoperative ileus. Of the 20 graduates of this program since 2000, 4 were laparoscopic/endourology fellows, and 2 of the residents pursued fellowship training after graduating. Graduates of the year 2000 and 2001 represent surgeons who graduated prior to the arrival of DMA. Of those resident graduates who did not pursue fellowship, two of the seven surgeons who graduated prior to the arrival of DMA are performing laparoscopy. Both of these surgeons pursued formal postgraduate laparoscopic training. Six of the seven non-fellowship-trained residents who graduated since DMA's arrival are performing laparoscopy; the other is early in practice and intends to do so. None of these surgeons has pursued postgraduate training prior to performing laparoscopy in their practices [corrected] CONCLUSIONS: The HALN techniques can be transferred quickly and efficiently between [corrected]one center and [corrected] another under the guidance of an experienced surgeon. Operative times are acceptable, with complication rates comparable to [corrected] previously reported series. Our data show that exposure during residency markedly increases the likelihood that surgeons will carry the techniques into their practices [corrected] PMID- 15659916 TI - "Pass the ball!" Simultaneous flexible nephroscopy and retrograde intrarenal surgery for large residual upper-pole staghorn stone. AB - Complex staghorn calculi have conventionally been treated with percutaneous nephrolithotomy, extracorporeal lithotripsy, or, occasionally, open surgery. Access to branched calculi is difficult through a single percutaneous track. We tested and make a case for a synchronous bidirectional technique combining the use of flexible instruments through percutaneous and retrograde approaches. We present a case of "pass the ball," which may have a place in tackling large, complex, or branched staghorn calculi without the need for multiple percutaneous punctures, removing larger fragments than is possible through the ureter and avoiding multiple passes through the ureter, saving time and decreasing the possibility of iatrogenic ureteral injury. PMID- 15659917 TI - Controversial cases in endourology. PMID- 15659918 TI - Stone disease in the Hmong of Minnesota: initial description of a high-risk population. AB - BACKGROUND AND PURPOSE: The Hmong are an ethically distinct refugee population from the highlands of Laos. Metropolitan Minneapolis-St. Paul, Minnesota is home to the largest population of Hmong ( 40,000) outside Asia. A clinical impression that the Hmong have a high rate of uric acid stone disease was evaluated. PATIENTS AND METHODS: A retrospective chart review was performed. All adult Hmong patients seen between January 1, 2000, and December 31, 2001, by a large urology group practice (N = 205) were compared with a similar number of non-Hmong patients (N = 204). RESULTS: Of these patients, 94 Hmong (46%) and 23 non-Hmong (11%) patients had stone disease (P < 0.001). After addition of 75 non-Hmong stone patients to create a suitable comparison group, a total of 86 Hmong and 88 non-Hmong stone patients had complete records. Sex (57% v 60% male) and mean age (47.9 v 47.3 years) were similar. Staghorn calculi were found in 21 Hmong (24%) and 0 non-Hmong patients (P < 0.001). Surgical treatments differed between Hmong and non-Hmong (P = 0.004): SWL (16 v 35), ureteroscopy (24 v 29), percutaneous nephrolithotomy (19 v 7), and nephrectomy (5 v 0). Treatment for staghorn calculi was refused by 9 Hmong patients (43%). Stone composition was available in 40 Hmong and 39 non-Hmong. Uric acid (>10%) was found in 50% of Hmong and 10% of non Hmong stones (P < 0.001). CONCLUSIONS: Hmong patients who come to urologic attention tend to do so for stone disease and tend to have uric acid calculi. The frequent requirement for invasive therapy, including nephrectomy, in the Hmong is of concern. PMID- 15659919 TI - Laparoscopy-assisted transperitoneal percutaneous nephrolithotomy for recurrent calculus in isthmic calix of horseshoe kidney. AB - BACKGROUND AND PURPOSE: Management of urolithiasis in a horseshoe kidney (HSK) poses a unique challenge. Although most patients can be managed by a combination of percutaneous nephrolithotomy (PCNL) and extracorporeal shockwave lithotripsy (SWL), calculi in the isthmic calix remain difficult to treat, as this area is out of reach during rigid PCNL, and, owing to the poor evacuation of the fragments, the results of SWL are suboptimal. CASE REPORT: A 59-year-old man known to have an HSK presented with right-sided lower-abdominal pain and episodes of urinary infection. In the past, he had undergone pyelolithotomy and lithotripsy for lithiasis in the kidney. Imaging studies identified a large recurrent calculus in the anteriorly directed isthmic calix. This was treated successfully by a laparoscopy-assisted transperitoneal PCNL. The laparoscopic view allowed the bowel to be retracted away from the site of the HSK, and PCNL guided by fluoroscopy and laparoscopy was performed. Complete stone clearance was achieved in a single stage. The patient remains free of symptoms and recurrence 3 months later. CONCLUSION: We believe this to be the first report describing this novel approach to lithiasis in an HSK. PMID- 15659920 TI - Efficacy of the Stone Cone for treatment of proximal ureteral stones. AB - BACKGROUND AND PURPOSE: Whereas ureteroscopic lithotripsy is more efficacious than SWL in treating lower-ureteral stones, the same has not been universally said of its ability to treat proximal ureteral stones. Because failed proximal lithotripsy is often attributable to the complications associated with stone migration into the renal pelvis and calices, an instrument that can prevent this migration is a potentially important tool in the ureteroscopic armamentarium. This study sought to assess the role of just such an instrument, the Stone Cone, in proximal-ureteral lithotripsy. PATIENTS AND METHODS: We treated 19 consecutive patients having proximal-ureteral stones using semirigid ureteroscopy, a Stone Cone Nitinol urologic retrieval coil, and holmium:YAG laser lithotripsy with a 200- or 365-microm fiber. In all patients, both the Stone Cone and the laser fiber were utilized under direct visual guidance through the working channel(s) of the ureteroscope. RESULTS: All 19 patients were rendered stone free after Ho:YAG laser lithotripsy in conjunction with a Stone Cone. No stone fragments were noted to migrate into the renal pelvis, and the Stone Cone did not break or become entrapped in any of the 19 cases. CONCLUSION: The Stone Cone is a powerful new tool for proximal-ureteral lithotripsy and will likely revolutionize the treatment of proximal-ureteral stones. The savings in morbidity, time, and money associated with not having to chase stone fragments using flexible ureteroscopy are considerable. PMID- 15659921 TI - Endoscopic management of renal candidiasis. AB - Renal candidial infections have been described in the literature in susceptible populations of all age groups. Systemic antifungal agents have been successful in treating these patients, but in the presence of an obstructed kidney, antegrade (perurethral) or retrograde (percutaneous) drainage of the pelvicaliceal system is mandatory to salvage the kidney. An aggressive percutaneous endoscopic approach for the management of such a case in a young diabetic patient with multiple comorbidities is described here with initial success. PMID- 15659922 TI - Laparoscopy-assisted transvaginal total exenteration for locally advanced cervical cancer with bladder invasion after radiotherapy. AB - When invasive cervical cancer involves the urinary bladder or rectum, exenteration can be curative treatment. However, this operation, particularly by an open approach, carries significant morbidity, both physically and psychologically. Laparoscopic surgery has been documented to be a reasonable alternative to the open counterpart for a variety of pelvic operative procedures, including such advanced procedures as laparoscopy-assisted vaginal hysterectomy, total laparoscopic hysterectomy, and laparoscopy radical hysterectomy. With improving surgical technology and increasing surgical experience, exenteration is a logical extension of current laparoscopic practice. However, it raises skepticism regarding the feasibility and justification for the complicated surgery. We herein describe our experience in a patient undergoing total exenteration assisted by laparoscopic technology for advanced recurrent cervical cancer after extensive radiotherapy. Transperitoneal laparoscopic total exenteration with ureterosigmoidstomy and end-sigmoidostomy was accomplished in 6 hours. The whole specimen was removed en bloc transvaginally. The patient tolerated the procedure well. The only complication was a wound infection 50 days postoperatively that was controlled with debridement and antibiotics. No episodes of pyelonephritis occurred. After 1 year of follow-up, the patient is free of cancer by imaging studies and lives without associated morbidity of this extensive palliative operation except the care of the sigmoid colostomy. PMID- 15659923 TI - Laparoscopic nephrectomy for renal-cell carcinoma during pregnancy. AB - Renal-cell carcinoma (RCC) during pregnancy is rare. Laparoscopic nephrectomy has been used effectively and safely in nonpregnant patients with RCC. We report a case of a 34-year-old pregnant woman with RCC, which we believe to be the first such case to be managed by laparoscopic nephrectomy. PMID- 15659924 TI - Comparison of dismembered and nondismembered laparoscopic pyeloplasty in the pediatric patient. AB - BACKGROUND AND PURPOSE: Laparoscopic dismembered pyeloplasty is an acceptable option for ureteropelvic junction (UPS) obstruction in the pediatric population. We compared our results with dismembered and nondismembered laparoscopic pyeloplasty. PATIENTS AND METHODS: A series of 26 children between the ages of 8 months and 15 years (mean age 5 years) underwent transperitoneal laparoscopic pyeloplasty for an obstruction not caused by a crossing vessel. Nineteen had an Anderson-Hynes dismembered pyeloplasty (AH), while the remaining seven had a nondismembered pyeloplasty in a Heineke-Mikulicz fashion (HM). The outcome measures were operative time, length of hospital stay, and resolution of obstruction by ultrasonography and diuretic radionuclide imaging. RESULTS: The mean operative time was 3.1 hours and 2.5 hours for AH and HM, respectively. No difference in hospital stay was noted, with a mean of 3 days. The stent was removed 6 weeks later. Four of the seven patients having nondismembered procedures presented with acute flank pain within 3 days of stent removal. The AH pyeloplasty produced a 94% rate of resolution of UPJ obstruction, while the HM patients did poorly, with a success rate of only 43% (P = 0.002; Fisher's exact test). CONCLUSIONS: We believe that for UPJ obstructions in children not involving a crossing vessel, laparoscopic dismembered (AH) pyeloplasty may be considered a safe alternative. PMID- 15659926 TI - Hand-assisted laparoscopic partial nephrectomy without hilar vascular clamping using a saline-cooled, high-density monopolar radiofrequency device. AB - BACKGROUND AND PURPOSE: Nephron-sparing surgery is now accepted as an alternative treatment option for small renal tumors. However, hemostasis during laparoscopic partial nephrectomy can be technically challenging, especially without hilar vascular clamping. The aim of our study was to evaluate the technique of hand assisted laparoscopic partial nephrectomy using the TissueLink (TissueLink Medical, Dover, NH), a saline-cooled monopolar radiofrequency device, without hilar vascular clamping. PATIENTS AND METHODS: Using the hand-assisted laparoscopic approach, the kidney is mobilized transperitoneally, and the renal tumor with overlying perinephric fat is exposed. The tumor is excised with a 1-cm margin using a combination of the TissueLink device and endoscopic scissors. The tumor and a biopsy of the base of the tumor bed are sent for frozen-section examination. The bleeding vessels are controlled with digital compression and the TissueLink device. At the end of procedure, the tumor bed is covered with a hemostatic agent. Three female and four male patients ages 52 to 76 years (mean 66 years) were treated with this new device for incidental tumors detected during imaging studies (N = 6) or during work-up for gross hematuria (N = 1). Preoperative imaging studies included CT in six patients and MRI in three. The average tumor size was 2.2 cm (range 1.3-3 cm). Only peripheral tumors that did not approach the hilum or the collecting system were selected. RESULTS: All of the patients underwent a hand-assisted laparoscopic partial nephrectomy using the TissueLink device without hilar vascular clamping. There were no intraoperative complications or conversions to open surgery. The mean operative time was 175 minutes, with an estimated blood loss of 186 mL (range 100-300 mL). Histologic examination demonstrated renal-cell carcinoma in five cases, oncocytoma in one, and an angiomyolipoma in one. The dimensions of the normal tissue around the tumor ranged from 1 to 4 mm, and frozen-section analysis showed tumor-free margins in all cases. Postoperatively, all patients recovered well except one patient who developed transient atrial fibrillation, which was treated medically in the immediate postoperative period. All patients were discharged in good condition at an average of 3 days (range 2-6 days). CONCLUSION: Hand-assisted laparoscopic partial nephrectomy without vascular clamping using the TissueLink device is a safe and feasible technique for exclusion of small exophytic renal tissues. PMID- 15659925 TI - Retroperitoneal-assisted laparoscopic pyeloplasty in children: initial experience. AB - PURPOSE: To demonstrate a novel application of laparoscopy to assist in the correction of ureteropelvic junction (UPJ) obstruction. PATIENTS AND METHODS: A retroperitoneal space is created, and the UPJ is identified and dissected from the surrounding tissue with a 1 x 10-mm 0 degrees telescope and 2 x 5-mm endoscopic instruments. Using a 5-0 Prolene stay suture, the UPJ is brought up through the 10-mm trocar site to the skin level, and the pyeloplasty is performed ex vivo over a double-J stent with loop magnification. Between July 2002 and March 2003, nine male and two female patients with UPJ obstruction underwent a pyeloplasty using this retroperitoneal-assisted laparoscopic pyeloplasty (RALP). The operative time, hospital stay, pain management, and follow-up radiologic studies were reviewed. RESULTS: In nine patients, RALP (bilateral in one patient) was performed (nine on the left side and one on the right). In the remaining two patients, mobilization of the UPJ was inadequate, and open conversion was performed. The average RALP operative time was 160 minutes (range 121-193 minutes). Postoperative pain management was optimal using oral codeine in eight patients, while one patient (RALP and pyelolithotomy) received parenteral analgesic postoperatively. The average length of stay in the hospital was 2 days (range 1-3 days). Follow-up (5-14 months) radiologic studies after stent removal showed improvement in the hydronephrosis in all except one patient. Furosemide renal scan at 3 to 6 months postoperatively was done in all patients and showed no obstruction. CONCLUSION: We consider RALP as the technique of choice for surgeons who want to increase their reconstructive laparoscopic skills. It provides an additional technique when skilled laparoscopists are faced with technically difficult pyeloplasties. PMID- 15659927 TI - Visual impairment after laparoscopic donor nephrectomy. AB - Loss of vision after surgery is rare and has never been reported after a laparoscopic procedure. We describe a case of visual deficits secondary to posterior ischemic optic neuropathy after a laparoscopic donor nephrectomy. The potential etiologies of postoperative visual loss are reviewed, and recommendations for avoiding this complication are discussed. PMID- 15659928 TI - Bacteriology of urinary tract infection associated with indwelling J ureteral stents. AB - PURPOSE: To investigate the microorganisms responsible for urinary tract infection (UTI) and stent colonization in patients with indwelling J ureteral stents and to compare the antimicrobial susceptibility pattern of the isolates from urine and J stents in order to establish the etiologic agents of bacteriuria and colonized stents in such patients and provide baseline data on an antibiotic policy for the urology unit. PATIENTS AND METHODS: Midstream urine from 250 patients requiring J stent insertion was investigated microbiologically prior to stent insertion and on the day of stent removal. After stent removal, 3 to 5 cm of the tip located in the bladder was also sent for culture. Patients' bio-data and underlying diseases were documented. Those with no known systemic diseases ("normal patients") were also studied as controls. Of the 250 patients studied, 152 (61%) were normal, while 27 (11%), 53 (21%), and 18 (7%) had diabetes mellitus (DM), chronic renal failure (CRF), and diabetic nephropathy (DN), respectively. The mean duration of stent retention was 27 days. All microbial isolates were tested for their susceptibility to a panel of 10 antibiotics. RESULTS: Twelve patients (5%) before stent insertion and 42 patients (17%; P < 0.001) on the day of stent removal had positive urine cultures. One hundred four stents (42%) were culture positive. Of the 104 patients with positive stent cultures, in 62 patients (60%), urine culture was sterile. The commonest isolates were Escherichia coli, Enterococcus spp., Staphylococcus spp., Pseudomonas, and Candida spp. On the day of stent removal, urine culture was positive in 28% of the normal patients compared with 57% (P = 0.11), 78% (P < 0.001), and 62% (P < 0.001) of patients with CRF, DM, and DN, respectively. Stent isolates were more resistant to antibiotics than the organism isolated before stent insertion. CONCLUSION: An indwelling J ureteral stent carries a significant risk of bacteriuria and stent colonization. The sensitivity of urine culture to stent colonization is low, and therefore, a negative culture does not rule out a colonized stent. Bacteria cultured from urine after stent insertion and from the stents are more resistant to antibiotics than are those cultured from urine before stent insertion. Norfloxacin or ciprofloxacin is recommended as prophylaxis prior to stent insertion, and an aminoglycoside can be added to treat symptomatic patients with severe infections. PMID- 15659929 TI - Randomized comparison of loops for transurethral resection of the prostate: preliminary results. AB - PURPOSE: To evaluate the comparative efficacy and morbidity of standard transurethral resection of prostate (TURP) and transurethral vaporesection (TUVRP) using four types of loops. PATIENTS AND METHODS: In a one-to-one randomized study, 50 patients with lower urinary-tract symptoms suggestive of bladder outlet obstruction and benign prostatic enlargement underwent TURP. Clinical data were collected using digital rectal examination, transrectal ultrasonography for evaluation of prostate volume, IPSS and IIEF-5 questionnaires, and serum prostate specific antigen concentrations. Intraoperative blood loss and fluid absorption were evaluated by measuring serum hemoglobin and respiratory alcohol concentration. Patients were followed at 3 and 18 months with evaluation of clinical symptoms, flow rates, residual urine volumes, and complications. RESULTS: There were no significant differences in blood loss, intraoperative fluid absorption, procedure time, or weight of the resected tissue between standard TURP and TUVRP with the various loops. No significant complications (infections, urethral stricture, reintervention) were seen. CONCLUSIONS: In this comparison of the clinical outcome and morbidity of standard TURP versus different loops for TUVRP, there were no significant differences in any of the parameters evaluated. PMID- 15659930 TI - Minimally invasive surgery for female urinary incontinence: experience with periurethral microballoon implantation. AB - PURPOSE: To assess the effectiveness of periurethral microballoon implantation for management of female urinary incontinence. PATIENTS AND METHODS: A total of 45 women with urinary incontinence (age range 47-88 years) were treated between May 2000 and June 2001. Microballoons were placed in the proximal periurethral tissue using endoscopic instrumentation under local anesthesia. Follow-up assessment including urodynamic study and quality- of-life assessment was performed at 1, 3, and 6 months. RESULTS: Implantation was successful in all cases with no significant adverse effects. Cure was achieved in 60% and improvement in 5%. The failure rate was 35%. Microballoon implantation significantly increased the mean urethral closure pressure, from 35.2 cm H(2)O (SD 14.9; range 15-72 cm H(2)O) to 46.5 cm H(2)O (SD 19.7; range 16-87 cm H(2)O) (P = 0.001). The best predictors of successful outcome in univariate analysis were initial urethral closure pressure (P = 0.047) and degree of incontinence (P < 0.001). CONCLUSIONS: Periurethral microballoon implantation is a useful technique for the management of female urinary incontinence, especially in elderly and inoperable patients. The microballoons do not present the adverse effects associated with other bulking agents. PMID- 15659931 TI - Treatment of intrinsic sphincter deficiency using the Macroplastique Implantation System: two-year follow-up. AB - PURPOSE: To evaluate the 24-month outcome of the Macroplastique Implantation System (MIS) for the treatment of stress urinary incontinence (SUI) using objective and subjective measures, including quality-of-life (QoL) impact. PATIENTS AND METHODS: Twenty-one consecutive women with a mean age of 47.4 years and a mean body mass index 26.16 kg/m(2) having urodynamically diagnosed intrinsic sphincter deficiency (ISD) were enrolled. Patients were assessed preoperatively and postoperatively by physical examination, Stamey grading of incontinence, pad usage, pad weight test, and urodynamic measurements. Quality of life was assessed by the King's Health Questionnaire. Patients underwent transurethral injection of the MIS under local anesthesia on an outpatient basis. The mean volume injected was 6.3 mL. The data presented are based on 24-month followup from the last injection, and the patients were considered their own controls. RESULTS: Patient QoL improved in all domains. Ten patients (47%) considered themselves cured; three (14.3%) thought they were improved, and eight (38.1%) believed the procedure had failed. According to the subjective surgeon Stamey grading, 8 patients (38.1%) were considered cured, 5 (23.8%) improved, and 8 (38.1%) failed. Daily pad usage was reduced from a mean of 4.38 to 1.47. According to the pad weight test, 12 patients (57.1%) were dry and two (9.5%) improved, while in 7 (33.3%), MIS failed. Urodynamic testing demonstrated 10 patients (50%) dry and 5 (25%) improved. CONCLUSION: The MIS proved to have an outcome acceptable to patients and surgeons at 24-month followup. The procedure can be performed easily under local anesthesia. PMID- 15659932 TI - Abnormal selective cytology results predict recurrence of upper-tract transitional-cell carcinoma treated with ureteroscopic laser ablation. AB - BACKGROUND AND PURPOSE: Endoscopic management of transitional-cell carcinoma (TCC) of the upper urinary tract remains associated with a significant rate of recurrence. We evaluated the impact of selective upper-tract cytology findings on tumor recurrence and renal salvage rate after ureteroscopic laser tumor ablation. PATIENTS AND METHODS: From 1993 though 2003, 38 patients with upper-tract TCC underwent ureteroscopic laser tumor ablation. Cytology specimens were collected from the upper urinary tract prior to ablation. "Abnormal cytology" was defined as the presence of malignant or atypical cells. Patients with abnormal cytology results were compared with patients with those having negative findings for tumor recurrence and renal salvage rates using the X (2) test. RESULTS: Of the 38 patients, 26 (68.4%) experienced at least one recurrence at a mean follow-up of 37.2 months. Pretreatment upper-tract cytology results were available in 34 of these patients: 17 (50%) were negative, and 17 were abnormal. Sixteen of the patients (94.1%) with abnormal cytology results had tumor recurrence after ablation, compared with 8 of the 17 (47.1%) with negative cytology findings (P = 0.0026). Twelve patients (31.5%) underwent nephroureterectomy during follow-up: 8 of the 17 (47.1%) with abnormal cytology, and 4 of the 17 (23.5%) with negative cytology (P = 0.15). CONCLUSION: Abnormal selective cytology results were associated with a significantly higher rate of tumor recurrence and a trend toward increased renal loss in patients with upper-tract TCC treated with ureteroscopic ablation. These findings suggest a prognostic value for upper-tract cytology analysis in patients undergoing endoscopic therapy. PMID- 15659933 TI - High-intensity focused ultrasound for ex vivo kidney tissue ablation: influence of generator power and pulse duration. AB - BACKGROUND AND PURPOSE: The therapeutic application of noninvasive tissue ablation by high-intensity focused ultrasound (HIFU) requires precise physical definition of the focal size and determination of control parameters. The objective of this study was to measure the extent of ex-vivo porcine kidney tissue ablation at variable generator parameters and to identify parameters to control lesion size. MATERIALS AND METHODS: The ultrasound waves generated by a cylindrical piezoceramic element (1.04 MHz) were focused at a depth of 100 mm using a parabolic reflector (diameter 100 mm). A needle hydrophone was used to measure the field distribution of the sound pressure. The morphology and extent of tissue necrosis were examined at generator powers of up to 400 W (P(el)) and single pulse durations of as long as 8 seconds. RESULTS: The two-dimensional field distribution resulted in an approximately ellipsoidal focus of 32 x 4 mm ( 6 dB). A sharp demarcation between coagulation necrosis and intact tissue was observed. Lesion size was controlled by both the variation of generator power and the pulse duration. At a constant pulse duration of 2 seconds, a generator power of 100 W remained below the threshold doses for inducing a reproducible lesion. An increase in power to as high as 400 W induced lesions with average dimensions of as much as 11.2 x 3 mm. At constant total energy (generator power x pulse duration), lesion size increased at higher generator power. CONCLUSIONS: This ultrasound generator can induce defined and reproducible necrosis in ex-vivo kidney tissue. Lesion size can be controlled by adjusting the generator power and pulse duration. Generator power, in particular, turned out to be a suitable control parameter for obtaining a lesion of a defined size. PMID- 15659934 TI - Lithotripter shockwave-induced enhancement of mouse melanoma lung metastasis: dependence on cavitation nucleation. AB - PURPOSE: To confirm a previous report of metastasis enhancement by lithotripter shockwaves (LSW) and to test the hypothesis that this effect is attributable to cavitation. MATERIALS AND METHODS: The metastatic B16-D5 melanoma cell line was implanted on the hind legs of female C57/b16 mice 12 days before tumor treatment. The tumors were treated with 500 LSW in a waterbath arrangement. The effect of augmented cavitation nucleation was tested by intratumor injection of air bubbles or ultrasound contrast agent gas bodies (UCAGB). The primary tumor was surgically removed on day 1 after treatment. The six groups of mice were sham, LSW, sham + air bubbles, LSW + air bubbles, sham + UCAGB, and LSW + UCAGB. Data were collected for the 113 mice that survived at least 25 days. Lung evaluations were performed blind after 2 weeks of bleaching in Fekete's solution. RESULTS: The outcomes of the three sham groups were very similar and indicated that the simple injection of material into the tumor did not increase metastasis. In comparison with the pooled shams, both the LSW + air bubbles and LSW + UCAGB groups had statistically significant increases in metastasis counts. Only the LSW + UCAGB group had a significant increase in incidence of metastasis relative to the pooled shams. The LSW + UCAGB also had significantly reduced survival. CONCLUSION: Shockwaves can enhance metastasis from tumors, and this effect is attributable to cavitation. PMID- 15659939 TI - Is loaded breathing an inflammatory stimulus? AB - PURPOSE OF REVIEW: To summarize recent data indicating that loaded breathing generates an inflammatory response. RECENT FINDINGS: Loaded breathing initiates an inflammatory response consisting of elevation of plasma cytokines and recruitment and activation of lymphocyte subpopulations. These cytokines do not originate from monocytes but are instead produced within the diaphragm secondary to the increased muscle activation. Oxidative stress is a major stimulus for the cytokine induction secondary to loaded breathing. The production of cytokines within the diaphragm may mediate the diaphragm muscle fiber injury that occurs with strenuous contractions, or contribute to the expected repair process. These cytokines may also compromise diaphragmatic contractility or contribute to the development of muscle cachexia. They may also have systemic effects, mobilizing glucose from the liver and free fatty acids from the adipose tissue to the strenuously working respiratory muscles. At the same time, they stimulate the hypothalamic-pituitary-adrenal axis, leading to the production of adrenocorticotropic hormone and beta-endorphins. The adrenocorticotropic hormone response may represent an attempt of the organism to reduce the injury occurring in the respiratory muscles through the production of glucocorticoids and the induction of the acute-phase response proteins. The beta-endorphin response would decrease the activation of the respiratory muscles and change the pattern of breathing, which becomes more rapid and shallow, possibly in an attempt to reduce and/or prevent further injury to the respiratory muscles. SUMMARY: Loaded breathing is an immune challenge for the body, initiating an inflammatory response. Further studies are needed to elucidate the role of this response in the development of ventilatory failure. PMID- 15659940 TI - Pulmonary and extrapulmonary acute respiratory distress syndrome: are they different? AB - PURPOSE OF REVIEW: Acute respiratory distress syndrome has been considered a morphologic and functional expression of lung injury caused by a variety of insults. Two distinct forms of acute respiratory distress syndrome/acute lung injury are described, because there are differences between pulmonary acute respiratory distress syndrome (direct effects on lung cells) and extrapulmonary acute respiratory distress syndrome (reflecting lung involvement in a more distant systemic inflammatory response). This article will focus on the differences in lung histology and morphology, respiratory mechanics, and response to ventilatory strategies and pharmacologic therapies in pulmonary and extrapulmonary acute respiratory distress syndrome. RECENT FINDINGS: Many researchers recognize that experimental pulmonary and extrapulmonary acute respiratory distress syndrome are not identical. In addition, clinical studies have described the detection of differences radiographically, functionally, and by analysis of the responses to therapeutic interventions (ventilatory strategies, positive end-expiratory pressure, prone position, drugs). However, there are contradictions among the different studies addressing these issues, which could be attributed to the fact that the distinction between pulmonary and extrapulmonary acute respiratory distress syndrome is not always clear and simple. Furthermore, there may be frequent overlapping in pathogenetic mechanisms and morphologic alterations. SUMMARY: The understanding of acute respiratory distress syndrome needs to take into account its origin. If each pathogenetic mechanism were to be considered, clinical management would be more precise, and probably the outcome could include real amelioration. PMID- 15659941 TI - Mechanical ventilation in acute respiratory failure: recruitment and high positive end-expiratory pressure are necessary. AB - PURPOSE OF REVIEW: To review as best the critical care clinicians can recruit the acute respiratory distress syndrome (ARDS) lungs and keep the lungs opened, assuring homogeneous ventilation, and to present the experimental and clinical results of these mechanical ventilation strategies, along with possible improvements in patient outcome based on selected published medical literature from 1972 to 2004 (highlighting the period from June 2003 to June 2004 and recent results of the authors' group research). RECENT FINDINGS: In the experimental setting, repeated derecruitments accentuate lung injury during mechanical ventilation, whereas open lung concept strategies can attenuate lung injury. In the clinical setting, recruitment maneuvers improve short-term oxygenation in ARDS patients. A recent prospective clinical trial showed that low versus intermediate positive end-expiratory pressure (PEEP) levels (8 vs 13 cm H2O) associated with low tidal ventilation had the same effect on ARDS patient survival. Nevertheless, both conventional and electrical impedance thoracic tomography studies indicate that stepwise PEEP recruitment maneuvers increase lung volume and the recruitment percentage of lung tissue, and higher levels of PEEP (18-26 cm H2O) are necessary to keep the ARDS lungs opened and assure a more homogeneous low tidal ventilation. SUMMARY: Stepwise PEEP recruitment maneuvers can open collapsed ARDS lungs. Higher levels of PEEP are necessary to maintain the lungs open and assure homogenous ventilation in ARDS. In the near future, thoracic CT associated with high-performance monitoring of regional ventilation (electrical impedance tomography) may be used at the bedside to determine the optimal mechanical ventilation of ARDS patients. PMID- 15659942 TI - New insights into the pathology of acute respiratory failure. AB - PURPOSE OF REVIEW: The purpose of this review is to provide a historical perspective and to analyze the recent advances in the understanding of the cellular and tissue pathology of acute respiratory failure, specifically of the acute respiratory distress syndrome. The scope of mechanisms involved in acute lung injury and acute respiratory distress syndrome is far too great to do it justice in a single review. Therefore, this review will focus only on recent advances in the understanding of the morphologic changes that occur in acute lung injury, acute respiratory distress syndrome, and ventilator-induced lung injury. RECENT FINDINGS: The use of fluorescent labels brought a novel method to identify and quantify cell wounding in the whole organ animal model of ventilator-induced lung injury. Real-time in vivo microscopy demonstrated the injurious effects of alveolar instability in the pathogenesis of ventilator-induced lung injury. Lipid tether mechanics, using laser tweezers, have advanced the understanding of the mechanical properties of the plasma membrane in response to mechanical stress. New animal injury models have brought forward new insights into the pathogenesis and structural abnormalities seen in acute respiratory distress syndrome. Apoptosis and epithelial wounding and repair have been examined in novel methods, and new mechanisms in lung edema formation have been proposed. SUMMARY: New mechanisms in the pathology of acute respiratory failure have shifted the focus to lung mechanics, tissue damage, remodeling, and the systemic effects derived from the mechanical stress imposed by the ventilator in patients with adult respiratory distress syndrome. PMID- 15659943 TI - An integrated approach to prevent and treat respiratory failure in brain-injured patients. AB - PURPOSE OF REVIEW: Brain-injured patients are at increased risk of extracerebral organ dysfunction, in particular ventilator-associated pneumonia. The purpose of this review is to discuss functional abnormalities, clinical treatment, and possible prevention of respiratory function abnormalities in brain-injured patients. RECENT FINDINGS: Ventilator-associated pneumonia worsens the neurologic outcome and increases the intensive care unit and hospital stay, costs, and risk of death. The respiratory dysfunction can be due to several causes, but atelectasis and/or consolidation of the lower lobes predominates in the most severe cases. Strategies should be implemented to prevent lung infections and accelerate weaning from mechanical ventilation to reduce the incidence of respiratory dysfunction and ventilator-associated pneumonia. SUMMARY: An integrated approach including appropriate ventilatory, antibiotic, and fluid management could be extremely useful, not only to prevent and more rapidly treat respiratory failure but also to improve neurologic outcome and reduce hospital stay. Further studies are warranted to better elucidate the pathophysiology and clinical treatment of respiratory dysfunction in brain-injured patients. PMID- 15659944 TI - Epidemiology of acute lung injury. AB - PURPOSE OF REVIEW: Acute lung injury and its extreme manifestation, acute respiratory distress syndrome, complicate a wide variety of serious medical and surgical conditions, only some of which affect the lung directly. Despite recent evidence-based advances in clinical management, acute lung injury and acute respiratory distress syndrome are associated with significant mortality. Detailed epidemiology is essential in guiding the recruitment of patients into trials of new therapeutic interventions, thereby improving outcome and allowing directed allocation of scarce resources. RECENT FINDINGS: The incidence of acute lung injury in the United States overall (17-64 per 100,000 person-years) seems to be higher than in Europe, Australia, and other developed countries (17-34 per 100, 000 person-years). The mortality rates for patients with acute respiratory distress syndrome range from 34 to 58%. The hypothesis that pulmonary and extrapulmonary acute respiratory distress syndromes are different disease entities continues to gain momentum. A genetic predisposition to acute respiratory distress syndrome may contribute to its pathogenesis and outcome. SUMMARY: Recent epidemiologic studies of the incidence of acute lung injury and acute respiratory distress syndrome have indicated a similar incidence in developed societies, and they confirm that mortality is falling in comparison with a decade ago. The awaited publication of new consensus guidelines for the definition of acute lung injury and acute respiratory distress syndrome may render new studies necessary. PMID- 15659945 TI - How to choose a mechanical ventilator. AB - PURPOSE OF REVIEW: To provide some practical and clinical considerations that may guide users through the decision process when choosing mechanical ventilators RECENT FINDINGS: Although the complexity of mechanical ventilators is steadily increasing, the importance of many devices developed over the course of the technical evolution is still a matter of discussion. Recent data demonstrate that the technical performance of equivalent ventilators (ie, machines of the same generation and category) is pretty similar, suggesting that the different manufacturers keep in step with new developments. Thus, other factors than technical limitations will probably influence the choice of ventilators. Among them the ability of the staff to understand the rationale of the different devices and controls as well as deal with the complexity of the ventilator may be particularly important. SUMMARY: Choosing mechanical ventilators should begin by defining the algorithms of how to ventilate a patient. Once this is done, a ventilator should allow the transformation of specific strategies into practice and the adaptation of the mechanical support to the needs of the individual patient. This procedure is crucially important, because ventilator therapy should always be determined by the physician and based on solid physiologic rationales rather than by the technical features of the machine. PMID- 15659946 TI - Permissive hypercapnia: role in protective lung ventilatory strategies. AB - PURPOSE OF REVIEW: Hypercapnia is a central component of current protective ventilatory strategies. This review aims to present and interpret data from recent clinical and experimental studies relating to hypercapnia and its role in protective ventilatory strategies. RECENT FINDINGS: Increasing clinical evidence supports the use of permissive hypercapnia, particularly in acute lung injury/acute respiratory distress syndrome, status asthmaticus, and neonatal respiratory failure. However, there are no clinical data examining the contribution of hypercapnia per se to protective ventilatory strategies. Recent experimental studies provide further support for the concept of therapeutic hypercapnia, whereby deliberately elevated PaCO2 may attenuate lung and systemic organ injury. CO2 administration attenuates experimental acute lung injury because of adverse ventilatory strategies, mesenteric ischemia reperfusion, and pulmonary endotoxin instillation. Hypercapnic acidosis attenuates key effectors of the inflammatory response and reduces lung neutrophil infiltration. At the genomic level, hypercapnic acidosis attenuates the activation of nuclear factor kappaB, a key regulator of the expression of multiple genes involved in the inflammatory response. The physiologic effects of hypercapnia, both beneficial and potentially deleterious, are increasingly well understood. In addition, reports suggest that humans can tolerate extreme levels of hypercapnia for relatively prolonged periods without adverse effects. SUMMARY: The potential for hypercapnia to contribute to the beneficial effects of protective lung ventilatory strategies is clear from experimental studies. However, the optimal ventilatory strategy and the precise contribution of hypercapnia to this strategy remain unclear. A clearer understanding of its effects and mechanisms of action is central to determining the safety and therapeutic utility of hypercapnia in protective lung ventilatory strategies. PMID- 15659947 TI - Assisted breathing is better in acute respiratory failure. AB - PURPOSE OF REVIEW: Mechanical ventilation is usually provided in acute lung injury to ensure alveolar ventilation and reduce the patients' work of breathing without further damaging the lungs by the treatment itself. Although partial ventilatory support modalities were initially developed for weaning from mechanical ventilation, they are increasingly used as primary modes of ventilation, even in patients in the acute phase of pulmonary dysfunction. The aim of this paper is to review the role of spontaneous breathing ventilatory modalities with respect to their physiologic or clinical evidence. RECENT FINDINGS: By allowing patients with acute lung injury to breathe spontaneously, one can expect improvement in gas exchange and in systemic blood flow, on the basis of both experimental and clinical trials. In addition, by increasing end expiratory lung volume, as will occur when airway pressure release ventilation is used, recruitment of collapsed or consolidated lung is likely to occur, especially in juxtadiaphragmatic lung regions. Until recently, traditional approaches to mechanical ventilatory support of patients with acute lung injury have called for adaptation of the patient to the mechanical ventilator using heavy sedation and administration of neuromuscular blocking agents. Recent investigations have questioned the utility of sedation, muscle paralysis, and mechanical control of ventilation. Further, evidence exists that lowering sedation levels will decrease the duration of mechanical ventilatory support, the length of stay in the intensive care unit, and the overall costs of hospitalization. SUMMARY: On the basis of currently available data, the authors suggest the use of techniques of mechanical ventilatory support that maintain, rather than suppress, spontaneous ventilatory effort, especially in patients with severe pulmonary dysfunction. PMID- 15659948 TI - How to ventilate patients with acute lung injury and acute respiratory distress syndrome. AB - PURPOSE OF REVIEW: The purpose of this paper is to review the mechanisms of ventilator-induced lung injury as a basis for providing the less damaging mechanical ventilation in patients with acute respiratory failure. RECENT FINDINGS: In normal lungs, high tidal volume causes an immediate gene upregulation and downregulation. Although the importance of alveolar inflammatory reaction is well known, recent findings suggest the potential role of airway distension in causing ventilator-induced lung injury. The initial activation has been shown to occur in the airways, accounting for the damages induced by high peak flow. The healthier lung regions are more exposed to the injury, since they may be subjected to strain. Challenge with endotoxin enhances in a synergistic manner the pulmonary inflammation induced by mechanical ventilation. However, mechanical strain and endotoxin seem to trigger lung inflammation through two different pathways. Despite convincing experimental and clinical evidences of lung injury, the clinical implementation of low tidal volume ventilation is still limited and has not yet become part of standard clinical practice. Setting positive end-expiratory pressure remains an open problem because the ALVEOLI study did not provide any exhaustive answers, likely because of methodologic problems and, unphysiologic design. SUMMARY: Gentle lung ventilation must be standard practice. Because stress and strain are the triggers of ventilator induced lung injury, their clinical equivalents should be measured (transpulmonary pressure and the ratio between tidal volume and end-expiratory lung volume). For a rational application of positive end-expiratory pressure, the potential for recruitment in any single patient should be estimated. PMID- 15659949 TI - Noninvasive ventilation for acute respiratory failure. AB - PURPOSE OF REVIEW: This review critically examines recent literature related to applications of noninvasive ventilation in the acute setting. RECENT FINDINGS: Recent articles have strengthened the evidence supporting the use of noninvasive ventilation for patients with cardiogenic pulmonary edema and exacerbation of severe chronic pulmonary obstructive disease. In the former, however, it remains unclear whether noninvasive ventilation offers any significant advantages over continuous positive airway pressure. The rate of myocardial infarction seems to be no higher when patients with cardiogenic pulmonary edema are treated with noninvasive ventilation rather than continuous positive airway pressure, although caution is still advised in patients with acute coronary syndromes. Noninvasive ventilation also does not seem to increase the risk of dissemination of severe acute respiratory syndrome to health care workers as long as strict isolation procedures are used. Noninvasive ventilation facilitates weaning in patients with chronic obstructive pulmonary disease but should not be used routinely to treat extubation failure, and necessary intubation should not be delayed. Guidelines for the use of noninvasive ventilation can alter caregivers' behavior but have not been clearly shown to improve outcomes. Outcomes do seem to improve, however, as caregivers acquire experience with the technique. SUMMARY: The recent literature has refined some of the current indications for noninvasive ventilation in the acute-care setting, including chronic pulmonary obstructive disease and cardiogenic pulmonary edema. Guidelines for use are now being developed, and outcomes seem to be improving, partly as a consequence of greater caregiver experience and possibly related to technologic advances. PMID- 15659950 TI - Overview of ventilator-induced lung injury mechanisms. AB - PURPOSE OF REVIEW: Mechanical ventilation is the main supportive therapy for patients with acute respiratory distress syndrome. As with any therapy, mechanical ventilation has side effects and may induce lung injury. This review will focus on stretch-dependent activation of alveolar epithelial and endothelial cells and polymorphonuclear leukocytes, and apoptosis/necrosis balance. RECENT FINDINGS: The past year has seen important research in the area of mechanotransduction and lung native immunity, suggesting further mechanisms of lung inflammation and injury in ventilator-induced lung injury. Research in the past year has also stressed the importance of inflammatory response by alveolar cells and role of polymorphonuclear leukocytes in stretch-induced lung injury and has suggested a role for apoptosis in the maintenance of the alveolar epithelium. SUMMARY: The proportion of patients receiving protective ventilatory strategies remains modest. If efforts to minimize the iatrogenic consequences of mechanical ventilation are to succeed, there must be a greater understanding of the signal transduction mechanisms and the development of potential pharmacologic targets to modulate the molecular and cellular effects of lung stretch. PMID- 15659951 TI - Extracorporeal membrane oxygenation. AB - PURPOSE OF REVIEW: Extracorporeal membrane oxygenation (ECMO) has become a more or less accepted standard in the algorithm of advanced acute respiratory distress syndrome therapy in adult patients when all other treatment options have failed. This article reviews the current status of ECMO therapy with particular focus on new technical developments and their potential implications for performance and indications for ECMO therapy. RECENT FINDINGS: A recently published review on a single-center experience in 255 adult ECMO patients identified using multivariate logistic regression analysis age, sex, initial pH 7.10 or lower and PaO2/FiO2 ratio, and days of mechanical ventilation before ECMO as a significant predictors of survival. Additionally, a careful cost-effectiveness study for neonatal ECMO relating a 4-year base to the UK neonatal ECMO trial has clearly demonstrated cost-effectiveness. SUMMARY: Over the years, the technique for ECMO therapy underwent substantial changes in indications and the materials used. Impressive technical progress has been made in pumps, oxygenators, and coating of artificial surfaces, leading to a higher biocompatibility and to a lower rate of procedure related complications. The potential of new inline pumps in combination with a decreasing rate of procedure-related complications might lead to a re-evaluation of the role of extracorporeal lung support in acute respiratory distress syndrome therapy. A very recent development is the use of spontaneous arteriovenous devices for carbon dioxide removal, allowing significant reduction of ventilator settings at decreased carbon dioxide partial pressures and at increased pH values. Ongoing studies are looking at the potential of this approach to reduce side effects of mechanical ventilation further. PMID- 15659952 TI - Bibliography. Current world literature. Respiratory system. PMID- 15659954 TI - Hematology and oncology. PMID- 15659955 TI - Hemophilia management: optimizing treatment based on patient needs. AB - PURPOSE OF REVIEW: Progress in hemophilia management over the past year has focused on improving understanding of the most common complications of genetic bleeding disorders and building upon available therapeutic strategies. RECENT FINDINGS: Research continued to link factor VIII structure with immune recognition and inhibitor formation. Clinical regimens of immune tolerance induction confirmed and expanded basic understanding. Barriers to optimal prevention using prophylaxis were explored allowing future refinements to address unmet needs. Outcome tools to assess joint health and overall quality of life were developed and validated. The inclusion of standardized instruments in assessment of outcome will allow meaningful comparison of available therapies. Use and complications of central venous access devices (CVAD), needed to deliver aggressive infusion regimens, were exhaustively reviewed. Finally, continued progress was achieved in development of improved vectors for future gene therapy of the hemophilias. SUMMARY: A general theme of recent progress in hemophilia management is harmonization in definitions and assessments of complications and outcomes, facilitating more rigorous and ultimately more useful interpretation of laboratory and clinical research. PMID- 15659956 TI - The biologic basis for neuroblastoma heterogeneity and risk stratification. AB - PURPOSE OF REVIEW: Neuroblastoma serves as the paradigm for the clinical utility of tumor-specific biologic data for prognostication. This review will describe the genetic and biologic basis for the diverse clinical phenotypes observed in neuroblastoma patients. It will also discuss the current approach to risk classification and how this may change in the future. RECENT FINDINGS: The biologic basis of neuroblastoma has come into clearer focus. PHOX2B is the first bona fide neuroblastoma predisposition gene identified, but is mutated in only a small subset of cases. Somatically acquired alterations at chromosome arms 3p and 11q are highly correlated with acquisition of metastases in the absence of MYCN amplification and may be useful as prognostic markers. The Children's Oncology Group risk classification system has been validated, with current emphasis on further refinement such as reevaluation of the age cutoff used to stratify therapy, and incorporation of additional molecular genetic markers is being studied prospectively. High-throughput genome scale analyses of neuroblastomas are further clarifying the genetic basis of this heterogeneous disease. SUMMARY: Neuroblastoma remains a significant challenge as high-risk patients are treated with intensive multimodal therapies but cure rates remain suboptimal. There is remarkable heterogeneity observed in tumor phenotype, ranging from spontaneous regression to relentless progression. There are literally dozens of clinical and biologic markers that have been proposed as being predictive of disease outcome, but large clinical correlative studies are sharpening the focus of which markers can be used by the clinician to optimize therapy for an individual patient. PMID- 15659957 TI - Pediatric malignancies provide unique cancer therapy targets. AB - PURPOSE OF REVIEW: Improving overall survival and reducing morbidity are major goals of childhood cancer research. This review explores an old idea that increased survival in childhood cancer can be achieved by inhibiting specific cancer targets. Specific therapeutic targeting would theoretically cause reduced morbidity as well as increased survival. Tumor-specific translocation-generated fusion proteins appear to be ideal tumor-specific therapeutic targets. This review will describe advances in aspects of target identification, potential for small molecule screening, and the evolution of clinical resistance to this new generation of pharmaceuticals. RECENT FINDINGS: Advances in molecular biology have identified new protein targets along with increased understanding of the biologic role of these proteins. Ewing sarcoma family of tumors research has benefited from new target discovery and enhanced biologic understanding of the EWS-FLI1 fusion protein. Congenital (infantile) fibrosarcoma and cellular mesoblastic nephroma have been grouped based on the presence of a common translocation fusion protein, ETV6-NTRK3. Functional knowledge of ETV6-NTRK3 has advanced so that strategies for screening small molecule inhibitors can proceed. Patients with chronic myeloid leukemia have benefited from the discovery of the BCR-ABL kinase inhibitor imatinib mesylate (Gleevec), thus showing how a molecular therapeutic target can be inactivated for improved therapy. This review will describe challenges raised by clinical resistance to imatinib mesylate as a paradigm for how resistance might evolve in other disease models. This review also describes how patients with synovial sarcoma might benefit from future therapy directed towards the SYT-SSX family of fusion proteins. SUMMARY: The increased utilization of small molecules to disrupt or inactivate tumor-specific molecular targets is rapidly evolving. The use of these small molecules to probe biology and treat disease is advancing towards a new generation of anticancer therapies. PMID- 15659959 TI - Traumatic shoulder dislocation in the adolescent athlete: advances in surgical treatment. AB - PURPOSE OF REVIEW: The shoulder joint has the greatest range of motion of any joint in the body and as a result is particularly susceptible to dislocation and subluxation. Recurrent instability is a common complication after traumatic shoulder dislocation in young people, with rates as high as 100% in skeletally immature patients and 96% in for adolescents. Treatment for shoulder dislocation has traditionally involved immobilization followed by a rehabilitation program. Recent studies have reported decreased rates of recurrent instability and improved outcomes in patients treated with surgical stabilization of acute, traumatic shoulder dislocation. The purpose of this review is to review recent publications concerning the treatment of traumatic shoulder dislocations in adolescents. RECENT FINDINGS: Lawton et al. retrospectively reviewed 70 shoulders in 66 patients 16 years old or younger treated for shoulder instability with follow-up more than 2 years. Forty-two shoulders were successfully treated with physical therapy, whereas 28 eventually required surgery. Subsequently, Deitch et al. retrospectively identified 32 patients between 11 and 18 years of age with radiographically documented traumatic anterior shoulder dislocation. Instability recurred in 75% of patients and 50% eventually required surgical stabilization. Bottoni et al. reported results of a prospective randomized trial comparing arthroscopic stabilization to nonoperative treatment of acute, traumatic shoulder dislocations in patients aged 18 to 26 years. Recurrent instability developed in 75% of patients treated conservatively versus 11% in those treated with surgery. DeBerardino et al. prospectively evaluated arthroscopic stabilization of acute shoulder dislocations in 48 young athletes with an average follow-up of 37 months and reported a 12% rate of recurrent instability. All patients with stable shoulders were able to return to their previous levels of activity. SUMMARY: Conservative management of traumatic shoulder dislocations in young patients is associated with high rates of recurrent instability. Recent studies have demonstrated improved results and significant reduction in recurrent instability in patients treated with surgical stabilization when compared with nonoperative treatment. PMID- 15659958 TI - Informed consent in pediatric clinical trials. AB - PURPOSE OF REVIEW: Clinical research in children is increasing. Concerns have been raised about both the inclusion and the exclusion of children in such research. Corresponding to these concerns, issues in informed consent for pediatric trials have become more pressing. This review discusses informed consent in pediatric trials and characterizes the latest literature. RECENT FINDINGS: Research into the consent process has shown that central concepts in pediatric research such as randomization and the distinctions between phases of clinical trials are not uniformly understood by parents or older pediatric patients. Newer approaches that are being developed to help remedy these deficits include the use of staged informed consent, the incorporation of interactive computer technologies to convey complex ideas, and variations in approaches to assent of the child based on multifactorial assessments of competence. Other variables in the consent process that are being studied for their impact on the process include individual life experiences and cultural background. Gaps in federal regulation and oversight of "informed consent" are emerging as new cases test established assumptions. SUMMARY: Lessons learned from recent studies regarding oversight of the consent process in pediatric clinical trials, the complex nature of assent, the impact of cultural variables, and more effective means of communicating what is involved in a clinical trial will shape future studies in consent and help to improve the process. PMID- 15659960 TI - Ultrasound for screening developmental dysplasia of the hip: a European perspective. AB - PURPOSE OF REVIEW: For the past 20 years, sonography of the hip has become the imaging procedure of choice to diagnose hip maturation disorders in German speaking countries. In Austria, hip sonography is a mandatory and essential feature of preventive public health measures; the "Mutter-Kind-Pass" has been the official medical identity card for mother and child since 1992 in Austria. This review summarizes the most recent literature dealing with ultrasound screening for developmental dysplasia of the hip in Europe. RECENT FINDINGS: This article focuses on the results in recent articles about sonography screening of the hip joint in the first year of life. SUMMARY: The continued development and refinement of the method, which is now standardized in German-speaking countries and hence can be taught in uniformly structured courses, has made sonography of the infant hip a gold standard for screening for developmental dysplasia of the hip in these countries. PMID- 15659961 TI - Foot and ankle injuries in the adolescent runner. AB - PURPOSE OF REVIEW: To evaluate the increase in overuse injuries in the adolescent athlete, specifically sports-related injuries to the foot and ankle of the adolescent runner. Factors affecting these injuries include anatomic considerations, gender, rate of development, growth, training errors, shoe wear, and running surface. RECENT FINDINGS: Most injuries in adolescent runners are overuse injuries rather than sudden traumatic events. Adolescent athletes are as prone as adults to poor form, poor habits, and poor training patterns. In addition, anatomic variants in the hindfoot, midfoot, and forefoot may predispose an athlete to specific injury. SUMMARY: Adolescent running injuries are common and becoming more frequent as trainers and athletes place increasing demands on the growing body. There is no evidence that this increased demand produces long term adverse effects; however, a significant amount of time can be lost to injuries unless training patterns are constructed to allow for repair of the adolescent athlete. PMID- 15659962 TI - Pediatric knee fractures. AB - PURPOSE OF REVIEW: Fractures about the knee are a common occurrence in children, and characteristics of the growing skeleton make children susceptible to specific fractures that do not occur in adults. The purpose of this review is to help the clinician to recognize, understand, and appropriately treat these injuries. RECENT FINDINGS: Pediatric knee fractures are diagnosed by a comprehensive history and physical examination supplemented with appropriate imaging modalities. Depending on the injury, treatment may include immobilization, arthroscopic treatment, or open reduction and internal fixation. SUMMARY: A thorough understanding of pediatric knee fractures will enable clinicians to appropriately manage these injuries and provide patients with a rapid return to preinjury activities. PMID- 15659964 TI - The immunization schedule confronts adolescence. PMID- 15659963 TI - Fusionless scoliosis surgery. AB - PURPOSE OF REVIEW: Severe spinal deformity in young children is a formidable challenge for optimal treatment. Standard interventions for adolescents and adults, including spinal deformity correction and fusion, may not be appropriate for young patients with considerable growth remaining. Alternate surgical options that provide deformity correction and protect the growth remaining in the spine are needed to treat this population of patients best. RECENT FINDINGS: Several groups have reported very exciting advances in the field of deformity spine surgery. Updated findings concerning the successful implementation of growing rods have revived this technique as a viable option for preserving near normal growth of the spine. New techniques have also been recently described, including vertebral stapling that produces asymmetric and corrective growth of the concavity of a deformity, and vertical expandable prosthetic titanium rib instrumentation that indirectly corrects spine deformity and protects spine growth remaining to treat an associated thoracic insufficiency syndrome. SUMMARY: New techniques and instrumentation at the disposal of spine surgeons allow the treatment of this challenging patient population to approach the goals of deformity correction and maintenance with preservation of potential growth. Preliminary outcomes from the different techniques are promising, but further investigation, including long-term follow-up, is necessary. PMID- 15659965 TI - Fever in the new millennium: a review of recent studies of markers of serious bacterial infection in febrile children. AB - PURPOSE OF REVIEW: Evaluation of a febrile infant or child for serious bacterial infections (SBI) can be a challenging task; there is no single reliable predictor of SBI in infants. This review examines some of the recent work evaluating the usefulness of indicators for SBI, such as white blood cell count (WBC), C reactive protein (CRP), procalcitonin (PCT), and interleukin-6 (IL-6). RECENT FINDINGS: While WBC is traditionally used as an indicator of serious infection, it appears to be the least specific and sensitive test in children. CRP and PCT are the most promising, but neither is an ideal single indicator by itself, especially in infants. There has been very limited experience with PCT in this country, however. IL-6 is more useful than WBC but less accurate than either CRP or PCT. SUMMARY: Much progress has been made in recent years in finding more accurate indicators of SBI than WBC. However, while recent developments have given clinicians some new tools in evaluating febrile infants and children, it remains a formidable undertaking. In the especially vulnerable infant population, the holy grail of a single ideal SBI indicator remains elusive. PMID- 15659966 TI - Controversies in the treatment of bronchiolitis. AB - PURPOSE OF REVIEW: Bronchiolitis is a very common and potentially serious respiratory disease of young children. To date, there is not a single, widely practiced, evidence-driven treatment approach. This review summarizes important recently published studies on the treatment of acute bronchiolitis for both outpatients and hospitalized children. RECENT FINDINGS: Bronchodilators, epinephrine, and corticosteroids have all been used in the treatment of bronchiolitis. As with older studies, most recently published randomized clinical trials have failed to demonstrate clinical efficacy in the use of these medications to treat either outpatients or infants hospitalized with bronchiolitis. Further, several meta-analyses and systematic reviews on this subject have been published in the last year or 2. Once again, most fail to provide convincing evidence to support the routine use of these medications to treat bronchiolitis. SUMMARY: The routine and repetitive use of bronchodilators, epinephrine, or corticosteroids to treat bronchiolitis in the absence of demonstrated clinical benefits for individual patients is not justified. PMID- 15659967 TI - Community-acquired methicillin-resistant Staphylococcus aureus in pediatrics. AB - PURPOSE OF REVIEW: Community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) is an emerging problem in pediatrics, with clinical and microbiologic characteristics that differentiate it from hospital-acquired MRSA (HA-MRSA). RECENT FINDINGS: Relative to HA-MRSA, CA-MRSA tends to cause localized disease (although serious illness occurs), is susceptible to more antibiotics, and has the same risk factors for acquisition/disease as methicillin-susceptible S. aureus (MSSA). At the gene level, CA-MRSA is more similar to MSSA than HA-MRSA: its emergence is apparently due to acquisition by an MSSA of the Staphylococcal Cassette Chromosome that bears mecA: the gene that encodes the methicillin resistant penicillin binding protein. Carriage of recognized staphylococcal virulence factors, particularly Panton-Valentine leukocidin, is common in CA MRSA, emphasizing its potential for causing serious illness. CA-MRSA is usually susceptible to clindamycin, trimethoprim-sulfamethoxazole, and rifampin, but inducible macrolide-lincosamide-streptogramin resistance in a subset of CA-MRSA could be problematic when clindamycin is used. SUMMARY: The appearance and spread of CA-MRSA represents a new challenge in pediatric medicine. A high level of clinical suspicion and development of rapid methods for its identification are needs for the future. PMID- 15659968 TI - Newer antistaphylococcal agents. AB - PURPOSE OF REVIEW: As antibiotic resistance in staphylococci continues to evolve, the ability to treat infections in children with confidence using first generation cephalosporins, penicillins, and macrolides is decreasing. Knowledge of the local trends in resistance is important in making decisions of empiric antibiotic therapy. The antibiotic resistance pattern for the child's pathogen should be assessed whenever possible, to allow the practitioner to properly judge the risks and benefits of alternative antibiotic agents, should they be required for definitive therapy. RECENT FINDINGS: Options for therapy of most methicillin resistant and macrolide-resistant community-acquired strains of Staphylococcus aureus include vancomycin, linezolid, and, in communities with a high proportion of susceptible strains, clindamycin. Daptomycin, a lipopeptide antibiotic with activity against virtually all strains of S. aureus, was recently approved by the United States Food and Drug Administration for adults. Second-generation glycopeptide antibiotics similar to vancomycin are in clinical trials in adults, including dalbavancin and oritavancin. Several new compounds, including cephalosporins active against methicillin-resistant S. aureus, are in preclinical development as well. SUMMARY: The recent worldwide emergence and rapid spread of community-acquired methicillin-resistant S. aureus has prompted a change in the approach to therapy of staphylococcal infections in both the outpatient clinic and the hospital. Newer agents active against methicillin-resistant S. aureus such as linezolid have been recently approved for children and other agents recently approved for adults are under investigation in children. Older agents for which relatively few data from prospective, controlled, comparative studies exist in the treatment of staphylococcal infections may also offer effective and less costly options for therapy. PMID- 15659969 TI - New agents for invasive mycoses in children. AB - PURPOSE OF REVIEW: Invasive fungal infections are an important cause of morbidity and mortality in immunocompromised children of all ages. This review summarizes information on new antifungal agents, including current data on their clinical use in children, as well as alternative strategies such as antifungal combination and immunomodulation therapy. RECENT FINDINGS: Novel antifungal agents, such as the echinocandins and the second-generation triazoles, were recently introduced that exhibit promising efficacy against Candida spp., Aspergillus spp., and other opportunistic fungal pathogens. These compounds are generally well tolerated and show substantial efficacy as salvage treatment and equal or even superior efficacy compared with older azoles or amphotericin B as first-line or empiric therapy for fungal infections. Clinical studies of pharmacokinetics and efficacy of the new agents in the pediatric population are, however, limited. SUMMARY: The response rates observed with the recently introduced drugs, although superior in some cases compared with older antifungal agents, are still far from satisfactory. The development of new antifungal compounds as well as the use of alternative approaches of combination therapy and immunomodulation should be pursued through well-designed laboratory and clinical studies in pediatric patients. PMID- 15659970 TI - Rotavirus vaccines: an update. AB - PURPOSE OF REVIEW: Rotavirus infection is the most common cause of severe diarrhea disease in infants and young children worldwide and has a major global impact on childhood morbidity and mortality. Vaccination is the only control measure likely to have a significant impact on the incidence of severe dehydrating rotavirus disease. RECENT FINDINGS: Rotavirus disease prevention efforts suffered a great setback in 1999 with the withdrawal of the RRV-TV vaccine less than a year after its introduction. Several new rotavirus vaccine candidates have now been developed and are undergoing clinical trials. SUMMARY: New safe and effective rotavirus vaccines offer the best hope of reducing the toll of acute rotavirus gastroenteritis in both developed and developing countries. PMID- 15659971 TI - Fever without apparent source on clinical examination. AB - PURPOSE: This review discusses recent literature that has focused on the epidemiology, clinical and laboratory evaluation and treatment of episodes of acute illnesses associated with fever and also of prolonged episodes of fever in children. RECENT FINDINGS: Articles addressed the epidemiology of invasive pneumococcal disease in children in other countries that have not yet initiated vaccination with the conjugated pneumococcal vaccine. From the United States there was a report of the decreased occurrence of invasive pneumococcal disease in patients being provided primary care who had been vaccinated with conjugated pneumococcal vaccine. Another report outlined the experience at children's hospitals with invasive pneumococcal disease in the years pre- and post introduction of pneumococcal vaccine. One of the studies found that there was a slight increase in pneumococcal disease caused by non-vaccine serotypes. Another group of articles focused on serious bacterial infections in infants with fever who are positive for respiratory syncytial virus (RSV). All studies found a significant occurrence of urinary tract infections in these patients. One report found that bacteremia may occur in such patients if less than 28 days of age. Finally articles from Turkey, Thailand, and Italy give excellent discussions about the range of diagnoses and key clinical findings that may be seen in children with prolonged fever. SUMMARY: In the review period, there was a particular emphasis on invasive disease caused by S. pneumoniae and the impact of vaccination with conjugated pneumococcal vaccine, on the occurrence of serious bacterial infection in febrile infants with RSV infections, and on the broad spectrum of diagnoses in children with prolonged fever in varying geographic locales. PMID- 15659972 TI - Office pediatrics: current issues in lower respiratory infections in children. AB - PURPOSE OF REVIEW: Recent estimates indicate that 1.9 million children worldwide die each year from acute respiratory illnesses, many of which are lower respiratory infections (LRIs). Global threats from indoor and outdoor air pollution, urban crowding, biologic weapons, and worldwide rapid transportation complicate our efforts to reduce the impact of LRI disease in children. As new strategies are developed to limit the spread of LRI disease in children, existing pathogens become more complex to treat and newer pathogens emerge as causes of clinical disease. Most LRI disease is managed in outpatient settings and remains a challenge to those who care for children. RECENT FINDINGS: Atypical pathogens (notably mycoplasma pneumoniae) are now known to cause a wide spectrum of disease, and with more extensive complications than previously recognized. Available data suggest a rapid increase in S. pneumoniae resistance to penicillin and macrolides in many cases of community acquired pneumonia, which has resulted in shifts in outpatient (and inpatient) antibiotic treatment regimens. Human metapneumovirus (HMPV) is now recognized to cause LRI illness similar to respiratory syncytial virus (RSV) in infants and children. General strategies for the prevention of influenza infection are expanded to include many infants and young children, and are projected to significantly reduce morbidity and mortality from this key LRI pathogen. SUMMARY: The spectrum of LRI pathogens--and the clinical illnesses caused by them--continues to change. Innovations in immunotherapy and vaccines will be instrumental to reducing the burden of LRI morbidity in infants and children worldwide in the future. PMID- 15659973 TI - Bibliography. Current world literature. Hematology and oncology. PMID- 15659975 TI - Resolutions nurses can keep: cleaning house in the new year. PMID- 15659976 TI - What if nurses said no to the war? PMID- 15659977 TI - Nursing and torture. PMID- 15659978 TI - Nursing and torture. PMID- 15659979 TI - Gerontologic nursing. PMID- 15659981 TI - Experiencing avlimil. PMID- 15659982 TI - Nursing aides and assault. PMID- 15659983 TI - A new year's update: three significant health stories of 2004. PMID- 15659990 TI - The public defibrillator: mandated in airports but not health clubs and workplaces. PMID- 15659992 TI - The COMFORT Behavior Scale: a tool for assessing pain and sedation in infants. PMID- 15659993 TI - Who scratched the end table?: On observation and imagination. PMID- 15659994 TI - Breastfeeding in nonmaternity settings. PMID- 15659996 TI - The state of the science: maternal-child health research. PMID- 15659998 TI - Disruptive behavior and clinical outcomes: perceptions of nurses and physicians. AB - Providing safe, error-free care is the number-one priority of all health care professionals. Excellent outcomes have been associated with procedural efficiency, the implementation of evidence-based standards, and the use of tools designed to reduce the likelihood of medical error (such as computerized medication orders and bar-coded patient identification). But the impact of work relationships on clinical outcomes isn't as well documented. The current survey was designed as a follow-up to a previous VHA West Coast survey that examined the prevalence and impact of physicians' disruptive behavior on the job satisfaction and retention of nurses (see "Nurse-Physician Relationships: Impact on Nurse Satisfaction and Retention," June 2002). Based on the findings of that survey and subsequent comments on it, the follow-up survey examined the disruptive behavior of both physicians and nurses, as well as both groups' and administrators' perceptions of its effects on providers and its impact on clinical outcomes. Surveys were distributed to 50 VHA hospitals across the country, and results from more than 1,500 survey participants were evaluated. Nurses were reported to have behaved disruptively almost as frequently as physicians. Most respondents perceived disruptive behavior as having negative or worsening effects, in both nurses and physicians, on stress, frustration, concentration, communication, collaboration, information transfer, and workplace relationships. Even more disturbing was the respondents' perceptions of negative or worsening effects of disruptive behavior on adverse events, medical errors, patient safety, patient mortality, the quality of care, and patient satisfaction. These findings suggest that the consequences of disruptive behavior go far beyond nurses' job satisfaction and morale, affecting communication and collaboration among clinicians, which may well, in turn, have a negative impact on clinical outcomes. Strategies aimed at reducing the incidence and impact of disruptive behavior are recommended. PMID- 15660000 TI - Sternal intraosseous infusions: emergency vascular access in adults. PMID- 15660001 TI - Pertussis on the rise. PMID- 15660005 TI - Germ warfare: The ANA and nurses work to prevent the misuse of antimicrobials in agriculture. PMID- 15660010 TI - Keeping PACE with older adults: the program of all-inclusive care to the elderly. PMID- 15660012 TI - A long way from home: two American RNs begin humanitarian work in Sudan. PMID- 15660013 TI - Preventing exposure to influenza: steps health care workers can take. PMID- 15660014 TI - Case definitions for chemical poisoning. AB - When human illness results from an unintentional or intentional release of a toxin (chemicals produced by metabolism in an organism [e.g., ricin]) or a toxicant (natural or synthetic chemicals not metabolically produced by an organism [e.g., nerve agents]) into the environment, uniform reporting is necessary to direct appropriate resources, assess the extent of morbidity and mortality, track poisoned persons, and monitor response to intervention. In this report, CDC presents case definitions to facilitate uniform reporting among local, state, and federal public health agencies of illness resulting from a chemical release. The report also explains the rationale for the structure of the case definitions, the audience for whom it is intended, the setting in which the case definitions might be used, and reasons each chemical presented in the report was selected. Clinical knowledge and diagnostic tools (e.g., biologic laboratory tests) for detecting chemical poisoning are likely to improve over time. CDC will create new case definitions and revise existing definitions to meet the needs related to emerging threats and to enhance case definition sensitivity and specificity, when possible, with developing clinical information. PMID- 15660015 TI - Antiretroviral postexposure prophylaxis after sexual, injection-drug use, or other nonoccupational exposure to HIV in the United States: recommendations from the U.S. Department of Health and Human Services. AB - The most effective means of preventing human immunodeficiency virus (HIV) infection is preventing exposure. The provision of antiretroviral drugs to prevent HIV infection after unanticipated sexual or injection-drug--use exposure might be beneficial. The U.S. Department of Health and Human Services (DHHS) Working Group on Nonoccupational Postexposure Prophylaxis (nPEP) made the following recommendations for the United States. For persons seeking care < or =72 hours after nonoccupational exposure to blood, genital secretions, or other potentially infectious body fluids of a person known to be HIV infected, when that exposure represents a substantial risk for transmission, a 28-day course of highly active antiretroviral therapy (HAART) is recommended. Antiretroviral medications should be initiated as soon as possible after exposure. For persons seeking care < or =72 hours after nonoccupational exposure to blood, genital secretions, or other potentially infectious body fluids of a person of unknown HIV status, when such exposure would represent a substantial risk for transmission if the source were HIV infected, no recommendations are made for the use of nPEP. Clinicians should evaluate risks and benefits of nPEP on a case-by case basis. For persons with exposure histories that represent no substantial risk for HIV transmission or who seek care >72 hours after exposure, DHHS does not recommend the use of nPEP. Clinicians might consider prescribing nPEP for exposures conferring a serious risk for transmission, even if the person seeks care >72 hours after exposure if, in their judgment, the diminished potential benefit of nPEP outweighs the risks for transmission and adverse events. For all exposures, other health risks resulting from the exposure should be considered and prophylaxis administered when indicated. Risk-reduction counseling and indicated intervention services should be provided to reduce the risk for recurrent exposures. PMID- 15660016 TI - Increase in poisoning deaths caused by non-illicit drugs--Utah, 1991-2003. AB - Deaths caused by drug poisoning of unintentional and undetermined intent are an increasing problem in Utah and elsewhere in the United States. To characterize the trend in drug-poisoning deaths in Utah, CDC and the Utah Department of Health analyzed medical examiner (ME) data for 1991-1998 and 1999-2003. This report summarizes the results of that analysis, which determined that, during 1991-2003, the number of Utah residents dying from all drug poisoning increased nearly fivefold, from 79 deaths in 1991 (rate: 4.4 per 100,000 population) to 391 deaths in 2003 (rate: 16.6). This increase has been largely the result of the tripling of the rate (from 1.5 during 1991-1998 to 4.4 during 1999-2003) in poisoning deaths of unintentional or undetermined intent caused by non-illicit drugs (i.e., medications that can be legally prescribed). Further study is needed to understand these trends and to develop strategies to prevent deaths of unintentional or undetermined intent from non-illicit drug poisoning. PMID- 15660017 TI - Unintentional non-fire-related carbon monoxide exposures--United States, 2001 2003. AB - Carbon monoxide (CO) is a colorless, odorless, poisonous gas that results from incomplete combustion of fuels (e.g., natural or liquefied petroleum gas, oil, wood, coal, or other fuels). CO sources (e.g., furnaces, generators, gas heaters, and motor vehicles) are common in homes or work environments and can put persons at risk for CO exposure and poisoning. Most signs and symptoms of CO exposure are nonspecific (e.g., headache or nausea) and can be mistakenly attributed to other causes, such as viral illnesses. Undetected or unsuspected CO exposure can result in death. To examine fatal and nonfatal unintentional, non-fire-related CO exposures, CDC analyzed 2001-2003 data on emergency department (ED) visits from the National Electronic Injury Surveillance System All Injury Program (NEISS-AIP) and 2001-2002 death certificate data from the National Vital Statistics System (NVSS). During 2001-2003, an estimated 15,200 persons with confirmed or possible non-fire-related CO exposure were treated annually in hospital EDs. In addition, during 2001-2002, an average of 480 persons died annually from non-fire-related CO poisoning. Although males and females were equally likely to visit an ED for CO exposure, males were 2.3 times more likely to die from CO poisoning. Most (64%) of the nonfatal CO exposures occurred in homes. Efforts are needed to educate the public about preventing CO exposure. PMID- 15660018 TI - Escherichia coli O157:H7 infections associated with ground beef from a U.S. military installation--Okinawa, Japan, February 2004. AB - In February 2004, the Okinawa Prefectural Chubu Health Center (OCHC) and the Okinawa Prefectural Institute of Health and Environment (OIHE), Japan, investigated three cases of Escherichia coli O157:H7 infection in a Japanese family associated with eating ground beef. Public health officials from multiple agencies in Japan and the United States collaborated on this investigation, which resulted in a voluntary recall of approximately 90,000 pounds of frozen ground beef in the United States and at U.S. military bases in the Far East. This was the first reported instance in which Japanese public health officials identified contaminated, commercially distributed ground beef that was produced in the United States. This report summarizes epidemiologic and laboratory investigations conducted by OCHC and OIHE. The results underscore the importance of using standardized molecular subtyping methods throughout the world to facilitate international public health communication and intervention. PMID- 15660019 TI - Elevated blood lead levels in refugee children--New Hampshire, 2003-2004. AB - As a result of reductions in lead hazards and improved screening practices, blood lead levels (BLLs) in children aged 1-5 years are decreasing in the United States. However, the risk for elevated BLLs (> or =10 microg/dL) remains high for certain populations, including refugees. After the death of a Sudanese refugee child from lead poisoning in New Hampshire in 2000, the New Hampshire Department of Health and Human Services (NHDHHS) developed lead testing guidelines to screen and monitor refugee children. These guidelines recommend 1) capillary blood lead testing for refugee children aged 6 months-15 years within 3 months after arrival in New Hampshire, 2) follow-up venous testing of children aged <6 years within 3 6 months after initial screening, and 3) notation of refugee status on laboratory slips for first tests. In 2004, routine laboratory telephone reports of elevated BLLs to the New Hampshire Childhood Lead Poisoning Prevention Program (NHCLPPP) called attention to a pattern of elevated BLLs among refugee children. To develop prevention strategies, NHDHHS analyzed NHCLPPP and Manchester Health Department (MHD) data, focusing on the 37 African refugee children with elevated BLLs on follow-up for whom complete data were available. This report describes the results of that analysis, which indicated that 1) follow-up blood lead testing is useful to identify lead exposure that occurs after resettlement and 2) refugee children in New Hampshire older than those routinely tested might have elevated BLLs. Refugee children in all states should be tested for lead poisoning on arrival and several months after initial screening to assess exposure after resettlement. PMID- 15660020 TI - The effects of amniotic membrane on retinal pigment epithelial cell differentiation. AB - PURPOSE: To examine the characteristics of retinal pigment epithelial (RPE) cells cultured on amniotic membrane (AM). The present study examined how AM modulates RPE cell differentiation. METHODS: Human RPE cells were cultured on the basement membrane side of dispase treated AM. After one week of cellular confluence, cultures were terminated, conditioned medium was collected, and total RNA was extracted. The expression pattern of several genes considered to participate in the function of differentiated RPE was evaluated. Ultrastructural changes were evaluated by transmission electron microscopy. RESULTS: Morphologically, RPE cells cultured on AM exhibited ultrastructural epithelial features such as microvilli of the apical membrane and intercellular junctions. Gene expression of RPE65, CRALBP, bestrophin, and tyrosinase related protein (TRP)-2 was upregulated in RPE cells cultured on AM compared to cells cultured on plastic. In addition, protein production of vascular endothelial growth factor, thrombospondin-1, and pigment epithelium derived factor was markedly increased in cells cultivated on AM. Gene expression of cathepsin D, brain derived neurotrophic factor, and basic fibroblast growth factor, however, did not differ between RPE cells cultured on plastic or AM. CONCLUSIONS: RPE cells cultured on AM demonstrated an epithelial phenotype morphologically and several growth factors important for maintaining retinal homeostasis were upregulated. AM might be a useful matrix substrate to retain the differentiated and epithelial phenotype of RPE for subretinal transplantation. PMID- 15660021 TI - Detailed histopathologic characterization of the retinopathy, globe enlarged (rge) chick phenotype. AB - PURPOSE: The purpose of this study was to characterize the morphological abnormalities in the retinas of chicks (Gallus gallus) suffering from the autosomal recessive disease, retinopathy, globe enlarged (rge/rge). METHODS: rge/rge affected and age matched control retinas were examined from hatch up to 730 days of age. Thickness of retinal layers at six retinal regions was measured from plastic embedded sections. Morphological features were examined on semi-thin sections by light microscopy and on ultra-thin sections by transmission electron microscopy. Immunohistochemistry was performed using a panel of several different antibodies. Additionally, comparative counting of rod outer segments, rows of cells in the inner nuclear layer, and ganglion cells per unit length was performed. RESULTS: The earliest changes observed in rge/rge retinas were disorganization of the outer plexiform layer and abnormal location of the endoplasmic reticulum of the photoreceptors. In rge/rge retinas, cone pedicles were larger, irregular in shape, and usually contained multivesicular bodies. In addition, synaptic ribbons of the cone pedicles and rod spherules in rge/rge retinas were less numerous compared to controls. Large glycogen deposits progressively accumulated in the perinuclear cytoplasm associated with the abnormally located endoplasmic reticuli in accessory cones and rods. Total retinal thickness progressively decreased with age in rge/rge birds. This was accompanied by a decrease in the number of cells in the inner nuclear layer and a decrease in the number of rod outer segments (OSs). Several changes were detected in the rge/rge retinas using immunohistochemistry, including mislocalized opsin immunoreactivity of rod photoreceptors, a decrease in number and disorganization of opsin positive rod OSs (especially in the peripheral regions), a decrease in number of tyrosine hydroxylase positive neurites in the distal inner plexiform layer, and activation of macroglial and microglial cells. CONCLUSIONS: As we previously reported, the rge/rge chick has vision loss that is not the result of photoreceptor loss and is unusual in that electroretinographic responses, although abnormal, are maintained until well after vision loss has developed. The phenotype is associated with a developmental disruption of both rod and cone photoreceptor synaptic terminals that progresses with age. It is possible that these changes may be indicative of abnormal circuitry within the outer plexiform layer, and that they underlie the progressive loss of vision in rge/rge birds. Other early changes suggesting photoreceptor abnormality are dilation of photoreceptor cell bodies, abnormal positioning of endoplasmic reticulum in the perinuclear region that is associated with abnormal glycogen deposition, and mislocalization of opsin immunoreactivity in rods. The rge/rge birds develop globe enlargement after the morphological and electroretinographic abnormalities. Globe enlargement in chicks can be induced by a number of different environmental factors. It is possible that abnormal signaling of photoreceptors to inner retinal cells could induce excessive ocular growth in the rge/rge birds. Many of the morphological changes such as retinal thinning seen in older rge/rge birds may be partly the result of the considerable globe enlargement that occurs later in the disease process. Molecular genetic studies to identify the causal gene mutation should help explain the morphological features of the rge/rge phenotype and clarify their association with vision loss and electroretinographic abnormalities. PMID- 15660022 TI - Inhibition of corneal angiogenesis by local application of vasostatin. AB - PURPOSE: This study was designed to investigate the effects of the locally supplied endogenous angiogenesis inhibitor vasostatin (VS) on corneal angiogenesis. METHODS: Recombinant VS was expressed and purified. The effects of VS on the proliferation of endothelial cells were investigated using the methyl thiazolyl tetrazolium (MTT) assay in the absence or presence of angiogenic factors such as basic fibroblast growth factor (bFGF) or vascular endothelial growth factor (VEGF). Corneal neovascularization was induced by implantation of hydron pellets containing bFGF in rat corneal micropockets. The potency of VS to inhibit corneal angiogenesis was investigated by incorporation of VS with bFGF in hydron pellets or topical application of VS containing eye drops to rat eyes implanted with bFGF pellets. The extent of corneal neovascularization was evaluated by microscopic and histological analyses. RESULTS: VS potently inhibited the growth of endothelial cells in the absence or presence of angiogenic factors such as bFGF or VEGF. In the rat corneal micropocket assay, concurrent incorporation of VS abolished the bFGF induced neovascularization. When formulated in a methylcellulose eye drop, VS remained intact and functional in a 4 degrees C solution for more than 7 days. Topical application of VS eye drops potently inhibited bFGF induced neovascularization in rat corneas. CONCLUSIONS: The present study effectively demonstrated the potential feasibility of local application of VS for treatment of corneal angiogenesis. PMID- 15660023 TI - Neonatal aphakia retards ocular growth and alters scleral gene expression in rhesus monkeys. AB - PURPOSE: We hypothesize that remodeling of the scleral extracellular matrix, involving collagen and proteoglycan synthesis and turnover, is a key process involved in ocular growth. Decreased axial elongation is observed following neonatal removal of the crystalline lens in a rhesus monkey model of congenital cataract. We wanted to determine changes in gene expression in the operated and companion eye following lensectomy, especially for extracellular matrix in the sclera. METHODS: Between 4 and 7 days of age, infant monkeys underwent surgical removal of the lens from the right eye. Axial lengths of the operated and unmanipulated fellow eyes were measured and when interocular differences of >0.4 mm were achieved, monkeys were sacrificed and RNA was isolated from sclera. In order to determine changes in scleral gene expression in aphakic versus control eyes, we used Clontech's Atlas Gene Array (Human Cancer Array version 1.2) hybridized with total RNA from three monkeys. RESULTS: Atlas Gene Array analysis demonstrated differential expression of several genes in the operated versus the unmanipulated eye. Most notably, there was a statistically significant increase in expression of several extracellular matrix (ECM) genes including: aggrecan, decorin, biglycan, several collagens, and tenascin in the RNA from the sclera of the aphakic eyes when compared to the unmanipulated eyes. Genes for several matrix metalloproteinases (MMPs) showed no significant change following lens removal although there was a trend towards decreased expression. There were also statistically significant changes in the pattern of gene expression in the operated eye relative to the unmanipulated eye for cell adhesion, cell cycle, apoptosis, and cytoskeleton transcripts. CONCLUSIONS: Our results suggest that removal of the crystalline lens alters gene expression in the sclera with a prominent upregulation of ECM transcripts. These data support recent evidence that remodeling of the ECM composition of the sclera may be an important regulator of ocular growth. PMID- 15660024 TI - Comparison of the efficacy and safety of atorvastatin initiated at different starting doses in patients with dyslipidemia. AB - BACKGROUND: The NASDAC study was designed to evaluate the safety and efficacy of atorvastatin at starting doses of 10, 20, 40, and 80 mg. METHODS: After an 8-week placebo washout period, 919 patients who were candidates for lipid-lowering therapy according to the National Cholesterol Education Program's Adult Treatment Panel III guidelines were randomized to 1 of 4 atorvastatin treatment groups: 10 mg (n = 229), 20 mg (n = 228), 40 mg (n = 231), and 80 mg (n = 231). RESULTS: Atorvastatin reduced low-density lipoprotein cholesterol (LDL-C) levels dose dependently across the 10- to 80-mg-dose range (35.7%-52.2%). Each of the 20-, 40 , and 80-mg doses provided significantly greater decreases in LDL-C than all lower doses (P < .01). All doses also reduced total cholesterol, the LDL-C/high density lipoprotein cholesterol ratio, apolipoprotein B, and triglycerides from baseline. An increase in high-density lipoprotein cholesterol was observed in all dose groups. Most participants, regardless of their level of coronary heart disease risk, attained their National Cholesterol Education Program's Adult Treatment Panel III LDL-C goal by the end of the study. Patients in all risk groups were more likely to achieve the NCEP LDL-C goal at higher starting doses. Atorvastatin was well tolerated at all dose levels. CONCLUSIONS: Atorvastatin initiated at doses of 10, 20, 40, and 80 mg is effective and safe for the treatment of patients with dyslipidemia. Depending on the percentage reduction needed to achieve an LDL-C goal, patients with or at risk of coronary heart disease may benefit from starting therapy at a higher dose of atorvastatin. PMID- 15660025 TI - The angiotensin II receptor blocker candesartan cilexetil reduces neointima proliferation after coronary stent implantation: a prospective randomized study under intravascular ultrasound guidance. AB - OBJECTIVES: The purpose of this study was to evaluate whether an angiotensin II receptor blocker, candesartan cilexetil, reduced neointima formation after coronary stent implantation by way of serial intravascular ultrasound analysis. BACKGROUND: Previous experimental studies have suggested that angiotensin II receptor blocker reduced neointima formation after vascular injury. However, it is unclear whether candesartan cilexetil has a similar effect on human coronary artery. METHODS: We recruited 50 consecutive patients with stable angina pectoris and 60 stenotic lesions. Patients were prospectively randomized into 2 groups: (1) 25 patients with 31 lesions received candesartan cilexetil (4-12 mg/d), and (2) 25 patients with 29 lesions did not receive the drug. Follow-up intravascular ultrasound was performed 6 m after the stent implantation. Cross-sectional images were obtained at 1-mm intervals within the stent, and the stent volume (SV), lumen volume (LV), and neointima volume (NV = SV - LV) were calculated using Simpson's rule. The percentage neointima volume obstruction (%NV) was calculated as NV/SV x 100. RESULTS: Clinical and angiographic backgrounds were comparable between the 2 groups. At follow-up, the candesartan group had smaller SV and larger LV (SV, 156.3 +/- 53.7 vs 165.4 +/- 61.8 mm3 , ns; LV, 122.2 +/- 49.0 vs 113.1 +/- 45.5 mm3 , ns), and significantly smaller NV and significantly smaller %NV than the control group (NV, 34.2 +/- 16.6 vs 52.3 +/- 32.6 mm3 , P < .01; %NV, 22.7 +/- 10.9% vs 31.3 +/- 13.4%, P < .01). CONCLUSIONS: Candesartan treatment decreases neointima formation and hence may reduce in-stent restenosis. PMID- 15660026 TI - Clopidogrel-aspirin combination for atrial fibrillation: CLAAF is not CLEAR. PMID- 15660027 TI - Conflict of interest for the physician-inventor using a device in human subjects. PMID- 15660028 TI - Atrial fibrillation and quality of life: clarity or evidence-based confusion? PMID- 15660029 TI - Diagnostic approach to the patient with cardiomyopathy: whom to biopsy. AB - The prevalence of congestive heart failure is increasing and the prognosis remains poor. Cardiomyopathy is one of the most frequent causes of congestive heart failure and is the most common etiology of patients submitted to heart transplant. Determination of the etiology of cardiomyopathy has both prognostic and therapeutic implications. History, physical examination, transthoracic echocardiogram, selected laboratory studies, and coronary angiography can often define the cause of cardiomyopathy, however, the etiology occasionally remains unknown despite this initial evaluation. The indications for endomyocardial biopsy (EMBx) in patients with cardiomyopathy and a negative initial evaluation remains uncertain. The search for Dallas histological criteria proven myocarditis prompted the performance of EMBx in patients with unexplained cardiomyopathy in hopes of identifying an etiology for which treatment would result in improvement in left ventricular function. The "negative" results of the Myocarditis Treatment Trial, with treated and controlled patients improving equally, dampened enthusiasm for this diagnostic procedure. However, our experience and recent evidence suggests that EMBx may be a valuable diagnostic modality and should be included in the evaluation of patients with initially unexplained cardiomyopathy. PMID- 15660031 TI - Metabolic syndrome: evaluation of pathological and therapeutic outcomes. PMID- 15660030 TI - The Coronary Artery Revascularisation in Diabetes (CARDia) trial: background, aims, and design. AB - BACKGROUND: Patients with diabetes have an increased incidence and severity of ischemic heart disease, which leads to an increased requirement for coronary revascularization. Comparative information regarding mode of revascularization- coronary artery bypass graft surgery surgery (CABG) or percutaneous coronary intervention (PCI)--is limited, mainly confined to a subanalysis of the Bypass Angioplasty Revascularization (BARI) trial, suggesting a mortality benefit of CABG over PCI. No prospective trial has specifically compared these modes of revascularization in patients with diabetes. OBJECTIVE: The Coronary Artery Revascularisation in Diabetes (CARDia) trial is designed to address the hypothesis that optimal PCI is not inferior to modern CABG as a revascularization strategy for diabetics with multivessel or complex single-vessel coronary disease. The primary end point is a composite of death, nonfatal myocardial infarction, and cerebrovascular accident at 1 year. METHOD: A total of 600 patients with diabetes are to be randomized to either PCI or CABG, with few protocol restrictions on operative techniques or use of new technology. This gives a power of 80% to detect non-inferiority of PCI assuming that the PCI 1 year event rate is 9%. A cardiac surgeon and a cardiologist must agree that a patient is suitable for revascularization by either technique prior to recruitment into the study. Twenty-one centers in the United Kingdom and Ireland are recruiting patients. Data on cost effectiveness, quality of life, and neurocognitive function are being collected. Long-term (3-5 year) follow-up data will also be collected. PMID- 15660032 TI - Metabolic syndrome: definition, pathophysiology, and mechanisms. PMID- 15660033 TI - Relationship between adolescent fitness and fatness and cardiovascular disease risk factors in adulthood: the Aerobics Center Longitudinal Study (ACLS). AB - PURPOSE: To examine the relationship between cardiorespiratory fitness and body fatness during adolescence and cardiovascular disease (CVD) risk factors in adulthood. METHODS: The sample was 48 participants from the Aerobics Center Longitudinal Study (ACLS) who had one clinical visit during adolescence (mean age = 15.8 years) and a follow-up visit during adulthood (mean age = 26.6 years). Values for treadmill time (TM), body fatness (body mass index [BMI], percent body fat [%BF], and waist circumference [WC]), traditional CVD risk factors (blood pressure [BP], fasting total cholesterol [TC], high-density lipoprotein cholesterol [HDL-C], triglycerides [TG], and glucose), and a composite metabolic syndrome risk factor score were adjusted for age and sex. The relationships between adolescent TM and body fatness variables and adult CVD risk factors were examined by partial correlations, controlling for length of follow-up (mean = 11 years). The correlation between changes in TM and body fatness (DeltaTM and Deltabody fatness, respectively) and changes in CVD risk factors between adolescence and adulthood (DeltaCVD risk factors) were also examined. Adolescent cardiorespiratory fitness and body fatness variables also were separated into 2 categories based on the median split: low (below median) and high (above median). RESULTS: TM and body fatness variables showed moderate stability between adolescence and adulthood ( r = 0.44-0.78). Adolescent TM and DeltaTM showed moderate correlations with adult body fatness indicators (BMI, WC, and %BF, r = 0.34 to -0.47) and Deltabody fatness ( r = -0.24 to -0.46), respectively. Adolescent TM was not significantly related to traditional CVD risk factors in adulthood. Adolescent WC was significantly related to adult BP ( r = 0.33 to 0.45), and body fatness variables during adolescence were significantly related to adult TM ( r = -0.32 to -0.44). The DeltaWC was significantly related to DeltaTM ( r = -0.46) and DeltaHDL-C ( r = -0.51), and DeltaBMI was significantly related to DeltaBP ( r = 0.45) and DeltaHDL-C ( r = -0.34). Subjects with a high TM during adolescence showed lower levels of body fatness as adults, and those with high levels of WC during adolescence showed higher BP during adulthood. CONCLUSIONS: The results indicate a significant relationship between adolescent cardiorespiratory fitness and adult body fatness and a lack of an association between adolescent cardiorespiratory fitness and adult cholesterol, BP, and glucose levels. Adolescent body fatness is moderately related to selected adult CVD risk factors. PMID- 15660034 TI - Prognostic impact of body weight and abdominal obesity in women and men with cardiovascular disease. AB - BACKGROUND: Increased body mass index (BMI) and abdominal adiposity increase the risk of cardiovascular disease (CVD) in persons free of these diseases, but their independent prognostic impact in persons with CVD has not been well defined. METHODS: BMI, waist-to-hip ratio (WHR), and waist circumference (WC) were measured in 6620 men and 2182 women with a mean age of 66 and stable CVD without congestive heart failure (CHF) participating in the Heart Outcomes Prevention Evaluation (HOPE) study. The main outcomes were CVD death, myocardial infarction, stroke, hospitalization for CHF, and all-cause mortality. RESULTS: During the 4.5 year follow-up, 658 had a CVD death, 1018 a myocardial infarction, 364 a stroke, 297 a CHF event, and 1034 died. When compared with the first tertile, the third tertile of BMI increased the adjusted relative risk (RR) of myocardial infarction by 20% (P < .02). Patients in the third tertile of WC had an increased adjusted RR of 23% for myocardial infarction (P < .01), 38% for heart failure (P < .03), and 17% for total mortality (P < .05). For WHR, there was an increased adjusted RR of 24% for CVD death (P < .03), 20% for myocardial infarction (P < .01), and 32% for total mortality (P < .001). CONCLUSIONS: Obesity, particularly abdominal adiposity, worsens the prognosis of patients with CVD; weight reduction program should be integrated in the active management of these patients. PMID- 15660035 TI - Factors associated with an increased chance of survival among patients suffering from an out-of-hospital cardiac arrest in a national perspective in Sweden. AB - AIM: To describe factors associated with an increased chance of survival among patients suffering from an out-of-hospital cardiac arrest in Sweden. PATIENTS AND METHODS: All patients suffering from an out-of-hospital cardiac arrest, which were not crew witnessed, in Sweden and in whom cardiopulmonary resuscitation (CPR) was attempted and who were registered in the Swedish Cardiac Arrest Registry. This registry covers about 85% of the Swedish population and has been running since 1990. RESULTS: In all, 33,453 patients, 71% of whom had a cardiac etiology, were included in the survey. The following were independent predictors for an increased chance of survival in order of magnitude: (1) patients found in ventricular fibrillation (odds ratio [OR] 5.3, 95% confidence limits [CL] 4.2 6.8), (2) the interval between call for and arrival of the ambulance less than or equal to the median (OR 3.6, 95% CL 2.9-4.6), (3) cardiac arrest occurred outside the home (OR 2.2, 95% CL 1.9-2.7), (4) cardiac arrest was witnessed (OR 2.0, 95% CL 1.6-2.7), (5) bystanders performing CPR before the arrival of the ambulance (OR 2.0, 95% CL 1.7-2.4), and (6) age less than or equal to the median (OR 1.6, 95% CL 1.4-2.0). When none of these factors were present, survival to 1 m was 0.4%; when all factors were present, survival was 23.8%. CONCLUSION: Among patients suffering from an out-of-hospital cardiac arrest, which were not crew witnessed, in Sweden and in whom CPR was attempted, 6 factors for an increased chance of survival could be defined. These include (1) initial rhythm, (2) delay to arrival of the rescue team, (3) place of arrest, (4) witnessed status, (5) bystander CPR, and (6) age. PMID- 15660036 TI - Impact of age on management and outcome of acute coronary syndrome: observations from the Global Registry of Acute Coronary Events (GRACE). AB - BACKGROUND: Evidence-based cardiac therapies are underutilized in elderly patients. We assessed differences in practice patterns, comorbidities, and in hospital event rates, by age and type of acute coronary syndrome (ACS). METHODS: We studied 24165 ACS patients in 102 hospitals in 14 countries stratified by age. RESULTS: Approximately two-thirds of patients were men, but this proportion decreased with age. In elderly patients (> or = 65 years), history of angina, transient ischemic attack/stroke, myocardial infarction(MI), congestive heart failure, coronary artery bypass graft (CABG) surgery, hypertension or atrial fibrillation were more common, and delay in seeking medical attention and non-ST segment elevation MI were significantly higher. Aspirin, beta-blockers, thrombolytic therapy, statins and glycoprotein IIb/IIIa inhibitors were prescribed less, while calcium antagonists and angiotensin-converting enzyme inhibitors were prescribed more often to elderly patients. Unfractionated heparin was prescribed more often in young patients, while low-molecular-weight heparins were similarly prescribed across all age groups. Coronary angiography and percutaneous intervention rates significantly decreased with age. The rate of CABG surgery was highest among patients aged 65-74 years (8.1%) and 55-64 years (7.7%), but reduced in the youngest (4.7%) and oldest (2.7%) groups. Major bleeding rates were 2-3% among patients aged < 65 years, and > 6% in those > or = 85 years. Hospital-mortality rates, adjusted for baseline risk differences, increased with age (odds ratio: 15.7 in patients > or = 85 years compared with those < 45 years). CONCLUSIONS: Many elderly ACS patients do not receive evidence based therapies, highlighting the need for clinical trials targeted specifically at elderly cohorts, and quality-of-care programs that reinforce the use of such therapies among these individuals. PMID- 15660037 TI - Prediction of missed myocardial infarction among symptomatic outpatients without coronary heart disease. AB - BACKGROUND: Missed diagnoses of acute myocardial infarction (AMI) in the ambulatory setting can cause patient suffering and malpractice litigation. Multiple algorithms have been developed to detect the presence of coronary heart disease (CHD) or acute coronary ischemia. METHODS: We performed a case-control study of patients with no prior history of CHD presenting to outpatient practices with potential cardiac ischemia. Malpractice claims files were used to identify 18 cases of patients with missed AMIs. For each case, we identified 3 control patients who had office visits for chest pain during the same month and assessed the association of 4 different prediction tools with missed AMI. RESULTS: The 18 cases of missed AMI had a 39% 1-month mortality rate. Cases were more likely than controls to be men (67% vs 26%, P = .001), to be smokers (88% vs 39%, P < .001), and to have low HDL cholesterol (39 mg/dL vs 59 mg/dL, P < .001) and elevated total cholesterol (236 mg/dL vs 213 mg/dL, P = .01). A Framingham risk score predicting a 10-year risk of CHD > or =10% and a positive score using the Goldman risk predictor were associated with an increased risk of missed AMI (odds ratio 5.7, 95% CI 1.8-18.4 for Framingham risk score; odds ratio 7.2, 95% CI 1.4-36.8 for Goldman risk predictor). CONCLUSIONS: Among ambulatory patients with possible cardiac ischemia and no prior CHD, multiple algorithms may be useful for improvement of risk stratification. PMID- 15660038 TI - Hospital therapy traditions influence long-term survival in patients with acute myocardial infarction. AB - BACKGROUND: Differences in therapy and outcome among hospitals have been reported, but these studies have seldom used adjustment for differences in patient characteristics. The objective was to investigate the differences in treatment of acute myocardial infarction (AMI) among different hospitals within 1 country and the possible causes and outcomes of these differences. METHODS: Prospective cohort study using data from the Register of Information and Knowledge about Swedish Heart Intensive care Admissions (RIKS-HIA) on 32,954 consecutive primary admissions of patients with AMI admitted to the intensive coronary care units of 67 Swedish hospitals in 1999-2000. An activity index was calculated for each hospital based on the hospital's ranking regarding proportion of performed examination or given therapy among the AMI patients. RESULTS: After adjustment for 24 background characteristics, there were few significant deviations among hospitals in the proportion treated with acute reperfusion, antiplatelets, beta-blockers, or angiotensin-converting enzyme (ACE) inhibitors. However, 3- to 10-fold differences existed among hospitals in the proportion of patients treated with intravenous beta-blockers, intravenous nitroglycerin, intravenous or subcutaneous anticoagulants, and lipid-lowering medication, and even larger discrepancies in echocardiography and revascularization within 14 days. There was a strong (r = 0.69, P < .001) correlation between hospital activity index between the years and a correlation between the hospital's activity index and 1-year mortality (r = -0.30, P = .014). There was no correlation between hospital size and activity index. CONCLUSIONS: Even after adjustment for differences in patient characteristics, there are differences between the hospital treatment cultures for patients with AMI that persists over time. Concerning everywhere-available treatment options, the treatment activity is independent of the size of the center. A more active treatment tradition is associated with a lower short- and long-term mortality in AMI patients. PMID- 15660039 TI - ST resolution in a single electrocardiographic lead: a simple and accurate predictor of cardiac mortality in patients with fibrinolytic therapy for acute ST elevation myocardial infarction. AB - BACKGROUND: The extent of ST-segment resolution in the 12-lead electrocardiograph (ECG) obtained early after reperfusion therapy in patients with ST-elevation myocardial infarction (MI) has been shown to predict short- and long-term mortalities. To improve the ease of this method in clinical practice, we sought to evaluate the optimal cutoffs and the prognostic value of ST resolution (STR) measured in a single ECG lead. METHODS: In conjunction with the Intravenous nPA for the Treatment of Infarcting Myocardium Early (InTIME)-2 study, in which patients with an ST-elevation MI of <6 hours' duration were treated with alteplase or lanoteplase, 12-lead ECGs were obtained at baseline and 90 minutes after the start of fibrinolytic therapy in 3030 patients. RESULTS: There was a close correlation between the extent of the sum STR and single-lead ST-elevation resolution ( r = 0.94). The optimal cutoffs for definition of single-lead complete, partial, and no-STR groups were 70% and 50% for anterior infarcts and 70% and 20% for inferior infarcts. The cardiac 30-day mortality rates for the 2 sets of risk groups by sum or single-lead STR were as follows: no resolution, 9.5% vs 10.3%; partial resolution, 5.0% vs 3.6%; complete resolution, 2.0% vs 1.2%. The predictive power was significantly better for single-lead STR. CONCLUSIONS: ST resolution obtained in a single lead is an easy and accurate prognosticator of cardiac 30-day mortality in patients with ST-elevation MI. It is therefore useful for early identification of low- and high-risk subgroups after fibrinolysis and as a surrogate end point in clinical trials. PMID- 15660040 TI - Does current treatment of cardiogenic shock complicating the acute coronary syndromes comply with guidelines? AB - BACKGROUND: The purpose of our study was to evaluate the implementation of guidelines for the treatment of cardiogenic shock (CS) complicating the acute coronary syndromes (ACS). METHODS AND RESULTS: Of the 10 136 patients in the Euro Heart-Survey-ACS with complete data, CS occurred in 549 (5.4%), of whom 28.6% had CS upon presentation. We examined the use of coronary angiography (CA), percutaneous (PCI) and surgical (CABG) revascularization, and intra-aortic balloon counterpulsation (IABP) among ACS patients with and without CS. During the hospital course, there were no significant differences between patients with and without CS in referral to CA (52.4% vs 53.3%, respectively) or CABG (4.4% vs 4.5%), but CS patients were more likely to undergo IABP (17.7% vs 0.8%, P < .001) and PCI (40.8% vs 31.8%, P < .001), especially younger (<75 years) patients (52.2% vs 31.8%, P < .001). A similar trend was observed when comparing ST elevation-ACS patients with (368 [8.5%]) and without CS (3945): CA (58.1% vs 56.2%), CABG (3.6% vs 3.3%), IABP (20.0% vs 0.9%, P < .01), and PCI (47.3% vs 40.6%, P < .01; 54.4% vs. 44.6% for patients <75 years, P < .003). Of the 94 ST elevation-ACS patients presenting with CS, only 39 (41.4%) received any reperfusion treatment, more often fibrinolysis (64.1%). The in-hospital mortality was 52.1% for all CS pts vs 2.0% for all others ( P < .001). CONCLUSIONS: Our contemporary survey demonstrates prohibitively-high mortality rates among ACS patients complicated by CS and poor implementation of recent guidelines advocating an aggressive invasive approach, including low rates of revascularization and IABP. Improved adherence to the guidelines pertaining to ACS patients developing CS may hopefully improve outcomes. PMID- 15660041 TI - Impact of physicians' beliefs and practices on cholesterol levels in patients with type 2 diabetes: a longitudinal assessment. AB - BACKGROUND: Clinical trials demonstrate significant benefit from cholesterol management for patients with type 2 diabetes. The aim of this work was to explore the correlates of lipid management in patients with type 2 diabetes, including the subjective beliefs of physicians, setting of care, and patient-related factors. METHODS: This longitudinal outcomes research study involved 2359 patients with type 2 diabetes recruited by 111 general practitioners and 214 physicians practicing in diabetes clinics. Physicians' beliefs were assessed through a questionnaire administered when the study started in 1998. Main outcome measures were total cholesterol (TC) and LDL cholesterol (LDL-C) levels over 3 years and the proportion of patients treated with lipid-lowering drugs (LLDs). RESULTS: Less than one-third of the physicians (27%) stated that they routinely started pharmacologic therapy for TC values > or =200 mg/dL (more aggressive), whereas 46% considered a TC level > or =240 mg/dL as the threshold for the initiation of treatment (less aggressive). During 3 years of observation, mean TC and LDL-C levels decreased from 215 +/- 40 mg/dL to 203 +/- 37 mg/dL and from 135 +/- 36 mg/dL to 126 +/- 35 mg/dL respectively, while the proportion of patients treated with LLDs increased from 13.2% to 24.6%; in particular, among individuals cared for by the more aggressive physicians, 30.0% were taking LLDs after 3 years, while only 17.7% of those followed by the less aggressive physicians and 18.1% of those followed by >1 physician were being treated with LLDs. Multilevel analysis showed that physicians' beliefs were an independent predictor of TC levels over the 3-year period. In patients treated with LLDs, TC levels decreased on average by 14%, and LDL-C levels decreased by 20%. CONCLUSION: Our data show that physicians' beliefs in more aggressive management strategies will result in better mean TC values over a 3-year period. PMID- 15660042 TI - Quality of life in atrial fibrillation: the Atrial Fibrillation Follow-up Investigation of Rhythm Management (AFFIRM) study. AB - OBJECTIVE: The objective of this prespecified substudy of the AFFIRM study, in which no differences in survival or event rates were found in patients with atrial fibrillation (AF) randomized to either rate control or rhythm control, was to test the null hypothesis that quality of life (QoL) is equal with rate- versus rhythm-control treatment strategies in AF. METHODS: Fifty-six (25%) of AFFIRM sites were randomly selected to recruit AFFIRM patients for the QoL substudy. Instruments used in the QoL assessment were (1) Perceived Health; (2) the Cantril Ladder of Life; (3) the Short Form 36 survey; (4) the QoL Index; and (5) the Symptom Checklist: Frequency and Severity. Data were collected at baseline, 2 months, 12 months, and annually; data are reported through 4 years of follow-up. RESULTS: Baseline characteristics of the AFFIRM QoL patients (n = 716) were generally similar to those of the rest of AFFIRM patients. Quality-of-life scores were similar in rate- and rhythm-control assignment groups at all time points. Quality-of-life scores were similar whether the actual rhythm was sinus or AF. Scores increased from baseline to subsequent time points similarly for both groups; these improvements were not additive over time. CONCLUSIONS: Quality of life was comparable between rate- and rhythm-control treatment strategies. In addition, QoL was similar with sinus rhythm versus AF. Attempts to improve QoL by restoring sinus rhythm will usually be unsuccessful. PMID- 15660044 TI - Collaterals and the recovery of left ventricular function after recanalization of a chronic total coronary occlusion. AB - BACKGROUND: A good collateral function in patients with regional myocardial dysfunction may indicate viability with the potential for left ventricular (LV) recovery after revascularization of a chronic total coronary occlusion (CTO). METHODS: A CTO (duration > 2 weeks) was successfully recanalized in 126 patients. During this procedure, the collateral function was assessed before the first balloon inflation by intracoronary Doppler and pressure wires. Collateral function indexes were calculated. Left ventricular function was assessed by the LV ejection fraction (LVEF) and the wall motion severity index (WMSI [SD/chords]). A repeat angiography was available in 119 patients after 4.9 +/- 1.4 m. An improvement of WMSI > or =1 SD/chord was considered significant. RESULTS: Left ventricular function was normal in 42%, regional dysfunction with LVEF > or = 0.60 was observed in 16%, and regional dysfunction with LVEF < 0.60 in 42%. The former had a better collateral function than patients with LV dysfunction. In 39% of patients with LV dysfunction, a significant myocardial recovery was observed at follow-up. The collateral function was similar in patients with and without recovery. However, patients with recovery had a lower peripheral resistance as an indicator of a better preserved microvascular integrity. CONCLUSIONS: Recovery of impaired LV function after revascularization of a CTO is not directly related to the quality of collateral function, as collateral development does not appear to require the presence of viable myocardium. However, a preserved microvascular integrity may be of relevance for myocardial recovery. PMID- 15660043 TI - Sex, quality of care, and outcomes of elderly patients hospitalized with heart failure: findings from the National Heart Failure Project. AB - BACKGROUND: Previous studies have demonstrated that women hospitalized for heart failure receive poorer quality of care and have worse outcomes than men. However, these studies were based upon selected patient populations and lacked quality of care measures. METHODS: We used data from the National Heart Failure Project, a national sample of fee-for-service Medicare patients hospitalized with heart failure in the United States in 1998-1999, to evaluate differences in quality of care and patient outcomes between men and women. Multivariable hierarchical logistic regression models and chi2 analyses were used to examine sex differences in the documentation of left ventricular systolic function (LVSF), prescription of angiotensin-converting enzyme (ACE) inhibitors or angiotensin receptor blockers (ARBs) for patients with left ventricular dysfunction, and mortality within 30 days and 1 year of admission in the study cohort (n = 30,996). RESULTS: Women had lower overall rates of LVSF assessment than men (64.9% vs 69.5%, P < .001). Among patients classified as candidates for ACE inhibitor prescription, women had lower crude rates of ACE inhibitor prescription than men (70.1% vs 74.2%, P = .015), but treatment rates were similar when evaluating the prescription of ACE inhibitors or ARBs (78.9% women vs 81.3% men, P = .11). Despite lower rates of treatment, women had lower mortality rates than men at 30 days (9.2% vs 11.4%, P < .001) and 1 year (36.2% vs 43.0%, P < .001) after admission. Results were similar after multivariable adjustment. CONCLUSIONS: There were small sex differences in the quality of care provided to fee-for service Medicare patients hospitalized with heart failure, although women had higher rates of survival than men up to 1 year after hospitalization. PMID- 15660045 TI - Elective percutaneous coronary intervention using broad-spectrum antiplatelet therapy (eptifibatide, clopidogrel, and aspirin) alone, without scheduled unfractionated heparin or other antithrombin therapy. AB - BACKGROUND: Adjunctive pharmacotherapy during percutaneous coronary intervention (PCI) has historically consisted of a regimen of antiplatelet agents accompanied by an antithrombin agent, typically unfractionated heparin. Paradoxically, unfractionated heparin may activate platelets, induce other pro-thrombotic activities, increase bleeding complications, and cause thrombocytopenia. To optimize patient care and avoid the potential risks of unfractionated heparin in patients undergoing elective PCI, one of the authors began to use adjunctive pharmacotherapy consisting of broad-spectrum antiplatelet therapy alone, without scheduled unfractionated heparin or other antithrombin therapy. METHODS: Five hundred consecutive patients undergoing scheduled, elective PCI (stent deployment, cutting balloon atherotomy, conventional balloon angioplasty, or high speed rotational atherectomy) received adjunctive pharmacotherapy consisting of eptifibatide, clopidogrel, and aspirin. RESULTS: The technical success rate was 100%. During the first 24 hours, there were no major adverse clinical events. Non Q-wave myocardial infarction occurred in 1.6% of patients, major and minor bleeding complications in 0.2% and 0.6%, respectively, and thrombocytopenia in 0.6%. During the first 30 days, there was 1 (0.2%) major adverse clinical event. CONCLUSIONS: For elective PCI, adjunctive pharmacotherapy consisting of broad spectrum antiplatelet therapy alone, without scheduled unfractionated heparin or other antithrombin therapy, appears to be safe and may prove to be efficacious. PMID- 15660046 TI - Incremental prognostic value of exercise single-photon emission computed tomographic (SPECT) thallium 201 imaging in patients with ST-T abnormalities on their resting electrocardiograms. AB - BACKGROUND: The incremental prognostic value of thallium 201 imaging in patients with nonspecific ST-T abnormalities on the resting electrocardiogram (ECG) may be different from those with a normal resting ECG. METHODS: Nine hundred thirty-nine consecutive patients with nonspecific ST-T abnormalities on their resting ECG who had undergone exercise 201 Tl imaging were followed for a median duration of 7.0 y (94% complete). The Cox proportional hazards regression model was used in a stepwise fashion to generate (1) a clinical (Cl) model, (2) a clinical and exercise (Cl + Ex) model, (3) and a clinical, exercise, and thallium (Cl + Ex + Tl) model, for the prediction of cardiac death. RESULTS: Age, sex, and diabetes composed the Cl model (chi2 = 63, P < .0001). The Duke treadmill score added to the Cl + Ex model (chi2 = 71, P < .0001). Increased lung uptake (P < .0001) added significantly and summed reversibility score ( P = .03) added modestly to the Cl + Ex + Tl model (chi2 = 96, P < .0001). On the basis of the Cl + Ex + Tl model, the low-, intermediate-, and high-risk groups had a 7-y survival free of cardiac death of 99%, 88%, and 58%, respectively (P < .0001). Using the Cl + Ex + Tl model, only a small number of low-risk and high-risk patients by the Cl + Ex model were reclassified. However, 48% of the 230 patients in the intermediate risk group by the Cl + Ex model were reclassified as low risk or high risk. CONCLUSIONS: 201 Tl imaging has incremental prognostic value in patients with nonspecific abnormalities on their resting ECG. However, patients classified as low risk or high risk by exercise testing using the Cl + Ex model do not require 201 Tl imaging. Intermediate-risk patients should be further risk-stratified by 201 Tl imaging. PMID- 15660047 TI - Determinants of functional capacity in patients with chronic heart failure: role of filling pressure and systolic and diastolic function. AB - BACKGROUND: Previous work suggesting a better correlation of diastolic than systolic function with exercise capacity in heart failure may reflect the relative insensitivity and load-dependence of ejection fraction (EF). We sought the correlation of new and more sensitive methods of quantifying systolic and diastolic function and filling pressure with functional capacity. METHODS: We studied 155 consecutive exercise tests on 95 patients with congestive heart failure (81 male, aged 62 +/- 10 years), who underwent resting 2-dimensional echocardiography and tissue Doppler imaging before and after measurement of maximum oxygen uptake (peak VO2). RESULTS: The resting EF was 31% +/- 10% and a peak VO2 was 13 +/- 5 mL/kg.min; the majority of these patients (80%) had an ischemic cardiomyopathy. Resting EF (r = 0.14, P = .09) correlated poorly with peak VO2 and mean systolic (r = 0.23, P = .004) and diastolic tissue velocities (r = 0.18, P = .02). Peak EF was weakly correlated with the mean systolic (r = 0.18, P = .02) and diastolic velocities (r = 0.16, P < .04). The mean sum of systolic and diastolic velocities in both annuli (r = 0.30, P < .001) and E/Ea ratio (r = -0.31, P < .001) were better correlated with peak VO2 . Prediction of peak VO2 was similar with models based on models of filling pressure (R = 0.61), systolic factors (R = 0.63), and diastolic factors (R = 0.59), although a composite model of filling pressure, systolic and diastolic function was a superior predictor of peak VO2 (R = 0.69; all P < .001). CONCLUSIONS: The reported association of diastolic rather than systolic function with functional capacity may have reflected the limitations of EF. Functional capacity appears related not only to diastolic function, but also to systolic function and filling pressure, and is most closely associated with a combination of these factors. PMID- 15660048 TI - Efficacy, safety and tolerability of metoprolol CR/XL in patients with diabetes and chronic heart failure: experiences from MERIT-HF. AB - BACKGROUND: The objective of the current study was to examine the efficacy and tolerability of the beta-blocker metoprolol succinate controlled release/extended release (CR/XL) in patients with diabetes in the Metoprolol CR/XL Randomized Intervention Trial in Chronic Heart Failure (MERIT-HF). METHODS: The Cox proportional hazards model was used to calculate hazard ratios (HR) for convenience expressed as relative risks (risk reduction = 1-HR), and 95% confidence intervals (CI). RESULTS: The risk of hospitalization for heart failure was 76% higher in diabetics compared to non-diabetics (95% CI 38% to 123%). Metoprolol CR/XL was well tolerated and reduced the risk of hospitalization for heart failure by 37% in the diabetic group (95% CI 53% to 15%), and by 35% in the non-diabetic group (95% CI 48% to 19%). Pooling of mortality data from the Cardiac Insufficiency Bisoprolol Study II (CIBIS II), MERIT-HF, and the Carvedilol Prospective Randomized Cumulative Survival Study (COPERNICUS) showed similar survival benefits in patients with diabetes (25%; 95% CI 40% to 4%) and without diabetes (36%; 95% CI 44% to 27%); test of diabetes by treatment interaction was non-significant. Adverse events were reported more often on placebo than on metoprolol CR/XL. CONCLUSIONS: Patients with heart failure and diabetes have a much higher risk of hospitalization than patients without diabetes. Regardless of diabetic status, a highly significant reduction in hospitalizations for heart failure was observed with metoprolol CR/XL therapy, which was very well tolerated also by patients with diabetes. Furthermore, the pooled data showed a statistically significant survival benefit in patients with diabetes. PMID- 15660049 TI - Insulin-treated diabetes is associated with a marked increase in mortality in patients with advanced heart failure. AB - OBJECTIVES: This study aimed to investigate the effect of diabetes and insulin use on survival in a large cohort of patients with advanced heart failure (HF) of multiple etiologies. BACKGROUND: Although diabetes is a well-known risk factor for both systolic and diastolic dysfunction, the impact of diabetes and insulin treatment on prognosis of patients with HF has not been well studied. METHODS: History of diabetes and insulin treatment was assessed in 554 consecutive patients with advanced systolic HF who presented to a single center for HF management and/or transplant evaluation (mean age 52.0 +/- 13.1 years, ejection fraction 24.6 +/- 7.4). Patients were stratified into 3 groups based on presence or absence of diabetes and insulin use. Differences in patient characteristics and survival were evaluated. RESULTS: There were 132 patients (23.8%) with diabetes; 43 patients (7.8%) were insulin treated and 89 patients (16.1%) were non-insulin-treated patients with diabetes. The groups were similar in sex, smoking history, medication profile, ejection fraction, body mass index, and serum sodium. Survival at 1 year was 89.7% for nondiabetic patients, 85.8% for non-insulin-treated diabetic patients, and 62.1% for insulin-treated diabetic patients (P < .000 01). After Cox multivariate analysis, insulin-treated diabetes was found to be an independent predictor of mortality (hazard ratio 4.30, 95% CI 1.69-10.94) whereas non-insulin-treated diabetes was not (hazard ratio 0.95, 95% CI 0.31-2.93). Similar findings were seen in clinically relevant subgroups. CONCLUSIONS: Insulin-treated diabetes is associated with a significantly worse prognosis in patients with advanced HF. Further investigations into mechanisms for the association of insulin treatment and mortality in patients with HF are warranted. PMID- 15660050 TI - Oxygen uptake efficiency slope, a new submaximal parameter in evaluating exercise capacity in chronic heart failure patients. AB - BACKGROUND: The oxygen uptake efficiency slope (OUES) is a new submaximal parameter which objectively predicts the maximal exercise capacity in children and healthy subjects. However, the usefulness of OUES in adult patients with and without advanced heart failure remains undetermined. The present study investigates the stability and the usefulness of OUES in adult cardiac patients with and without heart failure. METHODS: Forty-five patients with advanced heart failure (group A) and 35 patients with ischemic heart disease but normal left ventricular ejection fraction (group B) performed a maximal exercise test. PeakVO2 and percentage of predicted peakVO2 were markers of maximal exercise capacity, whereas OUES, ventilatory anaerobic threshold (VAT), and slope VE/VCO2 were calculated as parameters of submaximal exercise. RESULTS: Group A patients had lower peakVO2 (P < .001), lower percentage of predicted peakVO2 (P = .001), lower VAT (P < .05), steeper slope VE/VCO2 (P < .001), and lower OUES (P < .02). Within group A, significant differences were found for VAT, slope VE/VCO2, and OUES (all P < .01) between patients with peakVO2 above and below 14 mL O2/kg/min. Of all the submaximal parameters, VAT correlated best with peakVO2 (r =.814, P < .01) followed by OUES/kg (r = .781, P < .01), and slope VE/VCO2 (r = -.492, P < .001). However, VAT could not be determined in 18 (23%) patients. CONCLUSIONS: OUES remains stable over the entire exercise duration and is significantly correlated with peakVO2 in adult cardiac patients with and without impaired LVEF. Therefore, OUES could be helpful to assess exercise performance in advanced heart failure patients unable to perform a maximal exercise test. Further studies are needed to confirm our hypothesis. PMID- 15660051 TI - Additional impact of electrocardiographic over echocardiographic diagnosis of left ventricular hypertrophy for predicting the risk of ischemic stroke. AB - BACKGROUND: Patients with left ventricular hypertrophy (LVH) have an increased risk of ischemic stroke. Although echocardiography is commonly used for the diagnosis of LVH, there is little information about the potential role of electrocardiography in providing additional prognostic information. The purpose of this study is to determine if electrocardiographically derived criteria for LVH provide additional prognostic value over echocardiography for predicting ischemic stroke in a multiethnic population. METHODS: A population-based, case control study was conducted in 177 patients who had had a first ischemic stroke and in 246 control patients matched for age, gender, and race or ethnicity. Left ventricular mass was measured by using 2-dimensional transthoracic echocardiography. Logistic regression analysis was performed to assess the risk of stroke associated with the presence of LVH diagnosed by electrocardiography (defined by 4 established criteria) after adjustment for the presence of other stroke risk factors and for echocardiographically determined LVH. RESULTS: After adjustment for the presence of other established stroke risk factors, ECG-LVH was associated with ischemic stroke, using Sokolow-Lyon (odds ratio [OR] 2.12, 95% CI 1.05-4.30), Cornell voltage (OR 2.06, 95% CI, 1.26-3.35), and Cornell product criteria (OR 2.12, 95% CI, 1.13-3.97). Cornell voltage criterion (men, >2.8 mV; women, >2.0 mV) was associated with ischemic stroke even after adjustment for echocardiographically determined LVH (OR 1.73, 95% CI, 1.04-2.88). The combination of echo-LVH and a positive Cornell voltage criterion was associated with a 3.5-fold increase in stroke risk. CONCLUSIONS: Our study indicates that the presence of ECG-LVH is associated with an increased risk of ischemic stroke after adjustment for other stroke risk factors. For Cornell voltage criteria, this relationship persisted even after adjustment for echocardiographic LVH. Electrocardiographic results can provide independent information for left ventricular myocardial changes and should be considered together with echocardiographic results to fully assess the risk of ischemic stroke. PMID- 15660052 TI - "Substantially equivalent": the FDA, orthopedic device manufacturer, and you. PMID- 15660053 TI - Cementless porous-coated sockets without holes implanted with pure press-fit technique. AB - We evaluated the use of a pure press-fit technique in 220 porous-coated acetabular components without screw holes at mean follow-up of 6.1 years (4-7.5 years). The socket had a 14 degrees enhanced gradient at the periphery and was underreamed by 1 to 2 mm depending on bone quality. The Harris Hip average score was 44.3 (26-70) preoperatively and 93.1 (66-100) at final follow-up. There were no postoperative gaps in any zone. Nonprogressive radiolucent lines were detected in 4 hips. There was no movement or migration of the acetabular component on the follow-up x-rays. Pelvic osteolysis was not observed. Whereas 5 cups (2.3%) had revisions, none was revised for aseptic loosening. The press-fit technique provides stability in appropriate cases avoiding the theoretical complications related to screws or screw holes. PMID- 15660054 TI - Range of motion and stability in total hip arthroplasty with 28-, 32-, 38-, and 44-mm femoral head sizes. AB - The purpose of this study was to evaluate, via experimental models, the effect of larger head sizes for total hip arthroplasty on the type of impingement, range of motion (ROM), and joint stability. Testing was conducted using an anatomic full size hip model (anatomic goniometer) and a novel anatomic dislocation simulator with 28-, 32-, 38-, and 44-mm diameter femoral heads within a 61-mm acetabular shell. Femoral heads >32-mm provided greater ROM and virtually complete elimination of component-to-component impingement. A significant increase in both flexion before dislocation and displacement between the femoral head and acetabulum to produce dislocation occurred with femoral heads >32-mm in diameter. These data indicate that larger femoral heads offer potential in providing greater hip ROM and joint stability. PMID- 15660055 TI - Sensitivity and specificity of plain radiographs for detection of medial-wall perforation secondary to osteolysis. AB - Medial-wall perforation secondary to pelvic osteolysis impacts planning of acetabular revision surgery and may result in pelvic fracture. We compared commonly used radiographic signs for detecting medial-wall perforation (ballooning or discontinuity of Kohler's line or the iliopubic line) to findings from computed tomography (CT) in 27 cementless total hip arthroplasty patients with pelvic osteolysis. Used alone, none of the radiographic signs examined were reliable for detecting medial-wall perforation. When assessed together, however, the development of a discontinuity of Kohler's line, the iliopubic line, or both on anteroposterior pelvic radiographs was a reliable indicator for the presence of medial-wall perforation. Thus, the authors recommend a CT scan to evaluate the integrity of the medial wall when a discontinuity of Kohler's line or the iliopubic line has occurred and a revision surgery is planned. PMID- 15660056 TI - The variability of intramedullary alignment of the femoral component during total knee arthroplasty. AB - Intramedullary instrumentation for femoral component alignment during total knee arthroplasty is readily used. Newer alignment techniques using computer navigation are now available. This study assesses the difference in the sagittal and coronal plane alignments using a cadaveric model with 3 different entry points for intramedullary alignment compared with a navigation system. Seven cadaveric limb's results show that the anterior starting point resulted in recurvatum (-2.2 degrees +/- 1.4 degrees ), the middle starting point resulted in 1.9 degrees +/- 2.2 degrees of flexion, and the posterior starting point in 3.8 degrees +/- 2.6 degrees of flexion compared with the calculated femoral axis by the computer navigation system. When comparing the valgus angle, no statistical difference between any methods resulted (average 5.2 degrees +/- 0.9 degrees valgus). The anterior and posterior starting points were significantly different in the sagittal plane. These data suggest that alignment can be significantly affected by the starting point chosen for intramedullary instrumentation. PMID- 15660057 TI - The use of computer-assisted surgical navigation to prevent malalignment in unicompartmental knee arthroplasty. AB - We reviewed the outcome of 30 consecutive primary unicompartmental knee arthroplasties (UKAs) performed by a single surgeon over a 26-month period. All operations were performed to treat osteoarthritis of the medial compartment of the knee. Fifteen Allegretto (Sulzer, Winterthur, Switzerland) UKAs were implanted without computer navigation whereas 15 EIUS (Stryker-Howmedica, Allendale, NJ) UKAs were implanted using navigation. The patients were assessed clinically using the Oxford knee score and radiologically using long-leg weight bearing films and non-weight-bearing computed tomography leg alignment films. No patients operated on were lost to follow-up. Unicompartmental knee arthroplasty performed with computer-assisted surgical navigation resulted in a more accurate and reproducible limb alignment than UKA performed without surgical navigation. PMID- 15660058 TI - Augments and allografts in revision total knee arthroplasty: usage and outcome using one modular revision prosthesis. AB - Sixty-five consecutive Coordinate (DePuy, Warsaw, Ind) revision total knee arthroplasties were eligible for minimum 5-year follow-up. Nine patients died and 2 patients were lost. Therefore, 54 knees (51 patients) had a known outcome. Nine knees failed and required either revision or component removal. Eight additional knees were considered clinical failures. Despite the use of metallic augmentation in 89% of the knees, large structural allografts were required in 48% of the knees. Revisions with bone loss that required bulk allograft failed less often (19.2%) than revisions managed without bulk allografts (42.9%). Of 24 knees originally revised for osteolysis/polyethylene wear, only 1 required rerevision for the same mechanism of failure. Modular augments did not effectively address the bone loss and instability encountered in many instances at revision surgery. Survivorship of this implant was 79.4% +/- 13.7% at 8 years. PMID- 15660059 TI - The effect of intraoperative heparin administered during total hip arthroplasty on the incidence of proximal deep vein thrombosis assessed by magnetic resonance venography. AB - Intraoperative, unfractionated heparin, administered intravenously before the femoral work, has demonstrated to be effective in reducing the strong thrombotic stimulus that occurs during total hip arthroplasty (THA) surgery. This randomized, double-blind, prospective study included only THA patients with significant comorbidities predisposing them to deep vein thrombosis (DVT). The 2 groups consisted of study patients who received a single dose of intravenous, intraoperative, unfractionated heparin and control patients who received a single dose of intravenous, intraoperative saline. Magnetic resonance venography was used as the DVT diagnostic tool. The overall prevalence of proximal femoral vein clots was 2.2% (3 of 134), whereas pelvic thrombosis was detected in 10.4% (14 of 134). This study demonstrated that pelvic thrombi may form following THA and that a single dose of intraoperative heparin does not prevent their formation, but may be effective at preventing ipsilateral femoral thrombi. This study strongly supports a multimodal approach to DVT prophylaxis following THA. PMID- 15660060 TI - The precision and usefulness of preoperative planning for cemented and hybrid primary total hip arthroplasty. AB - We evaluated the utility of a preoperative planning technique with a review of preoperative radiographs, templates, plans, charts and 6-week postoperative radiographs of 139 total hip arthroplasties (THAs) (116 cemented and 23 hybrid) to determine size, orientation of the implants, and cement column mantle, location of the planned and achieved center of rotation, and limb-length discrepancy. The acetabular component size was predicted exactly in 116 hips (83%) (within +/-1 size in 138 hips [99%]); the femoral component size was predicted exactly in 108 hips (78%) (within +/-1 size in 138 [99%]). In 75 arthroplasties (45%), the center of rotation was within 2 mm of horizontal and vertical distance from the plan, and in 127 (91%) arthroplasties, it was within 4 mm. The inclination of the cup averaged 44 degrees (range, 30 degrees -58 degrees ). The stem was in a neutral alignment in 122 hips (88%), varus in 11 hips (8%), and in 2 degrees of valgus in 6 hips (4%). In 103 arthroplasties with a normal contralateral hip or a THA, the average limb-length discrepancy was 1.71 mm. Preoperative planning is useful to predict the implant size, position, and alignment, to restore the center of rotation, and to equalize limb length. PMID- 15660061 TI - Evaluation of the anterior cruciate ligament integrity and degenerative arthritic patterns in patients undergoing total knee arthroplasty. AB - We prospectively reviewed 107 consecutive primary total knee arthroplasties performed over a 1-year period. Intraoperatively, the integrity of the anterior cruciate ligament (ACL), the characteristics of the intercondylar notch, and the patterns of cartilage wear were evaluated. The ACL was found to be deficient in 41 knees (39%) at the time of surgery. The ACL-deficient knee had significantly narrower intercondylar notch widths compared with knees with an intact ACL (average, 9.75 vs 16 mm, P < .01). Furthermore, patients with ACL deficiency were found to have a higher percentage of Outerbridge grade IV changes at the lateral femoral condyle, lateral tibial plateau, and patellar surfaces when compared to the ACL-intact group. An intact ACL appeared to be protective against severe patellar degeneration. In conclusion, intercondylar notch narrowing from the arthritic process can lead to attrition and rupture of the ACL. An ACL deficiency appears to be associated increased wear of the lateral femorotibial and patellofemoral joints. PMID- 15660062 TI - Efficacy of cementless total hip arthroplasty in patients on long-term hemodialysis. AB - We retrospectively reviewed 11 hip arthroplasties in 7 patients receiving long term (15 years) hemodialysis for chronic renal failure. Osteonecrosis of the femoral head, osteoarthritis, and amyloid arthropathies were treated by total hip arthroplasty (THA) or bipolar hemiarthroplasty (BHA) using cementless anatomic medullary locking stem. The average follow-up period was 8 years and 3 months (range, 3-13 years). The D'Aubigne and Postel hip score was restored to 14.5 postoperatively from 7.7 preoperatively. All femoral cementless components achieved bone-ingrown fixation. Although 1 BHA resulted in central migration, there was no loosening of the cementless acetabular component. We are encouraged by the predictable long-term stability of the bone-implant interface achieved by cementless fixation with an extensively coated implant. Cementless THA remains a useful treatment option for patients on long-term hemodialysis. PMID- 15660063 TI - Intramedullary compression arthrodesis of the knee: early experience with a new device and technique. AB - The irretrievably failed total knee arthroplasty is the primary indication for knee arthrodesis. Because this difficult condition is relatively rare, an intramedullary arthrodesis system was developed which requires minimal surgeon experience for successful use. The new system called the Wichita Fusion Nail was implanted by a single surgeon in 13 consecutive patients: 11 for arthrodesis alone, 1 for stabilization of a supracondylar fracture nonunion, and 1 for arthrodesis coupled with a supracondylar fracture nonunion. All arthrodesis attempts were successful. The average fusion time was 15.2 weeks except for 2 infected delayed arthrodeses. Both fracture nonunions persisted and went on to amputation. The WFN provides a simple arthrodesis system with minimal technique dependence and a high potential for success. PMID- 15660064 TI - Extended femoral osteotomy for revision of hip arthroplasty: results and complications. AB - Seventy-five revision total hip arthroplasties (73 patients) performed using an extended greater trochanteric osteotomy for exposure and an extensively porous coated uncemented stem were reviewed after 2 years (range, 1 to 5). The mean osteotomy length was 14 cm (range, 7 to 19 cm) and fixation was with at least 2 cables. One stem was revised at 4 months for stem subsidence and loosening. Of the remaining 74 osteotomies, 73 healed without further procedures. A single nonunion was reoperated at 4 months and healed. Sixty-eight of 73 osteotomies healed with no migration and 5 did with less than 5 mm of proximal migration of the osteotomy fragment. Complications included 3 intraoperative and 1 postoperative osteotomy fragment fractures. Extended trochanteric osteotomy, executed preserving the fragment's vascularity and fixed with rigid fixation, can be performed with a low rate of complications. PMID- 15660065 TI - Analysis of 16 retrieved proximally cemented femoral stems. AB - Sixteen proximally cemented, collared, and distally splined, Bridge Hip femoral stems with a matte proximal surface and smooth distal surface were retrieved because of loosening. Electron microscopy, with correlated elemental analysis, identified titanium particulate embedded in the internal surface of the cement mantle. Data supported the observations that loosening of the femoral stems was related to proximal debonding at the cement-implant interface, loosening at the proximal cement-bone interface, and inherent rotational instability. Cement implant interface debonding resulted in the proximally matte femoral stem surface abrading with the opposing cement mantle, resulting in particulate and osteolysis in some cases. Careful consideration of implant design and clinically relevant biomechanical testing protocols should be considered before the clinical introduction of future proximally cemented femoral stems. PMID- 15660066 TI - Comparison between triple-tapered and double-tapered cemented femoral stems in total hip arthroplasty: a prospective study comparing the C-Stem versus the Exeter Universal early results after 5 years of clinical experience. AB - A prospective study was conducted comparing the early clinical and radiological results between the triple-tapered C-Stem (DePuy International, Leeds, UK) and the double-tapered Exeter Universal (Stryker Howmedica Osteonics, Mahwah, NJ) in cemented total hip arthroplasty. A total of 192 patients (200 hips) received the C-Stem and 189 patients (205 hips) received the Exeter Universal. Average follow up was 29 and 31 months in the C-Stem and Exeter groups, respectively. From the short-term results, we found similar mean improvements in clinical scores and no difference was noted in the incidence of complications. Average subsidence was comparable (C-Stem, 0.77 mm +/- 0.56; Exeter, 0.82 +/- 0.63). Radiolucencies in the cement-stem and cement-bone interfaces were seen primarily in the proximal zones in both groups. No stems were considered at risk of aseptic loosening and the extent of proximal femoral resorption was comparable. PMID- 15660068 TI - Radiological findings and clinical results of 102 thrust-plate femoral hip prostheses: a follow-up of 2 to 8 years. AB - We report the radiological and clinical outcome of 102 consecutive femoral hip arthroplasty we prospectively followed up in 84 patients using the third generation of the thrust-plate prosthesis with a mean period of follow-up of 58 (range, 26-100) months. Four implants were revised: 2 because of an infection and 2 because of aseptic loosening. The thrust-plate prosthesis, which allows preservation of part of the femoral neck, was used in younger patients, 80% were younger than 60 years. In 95 implants, contact was maintained between thrust plate and underlying bone, and in only 3 instances, without any clinical manifestation, did the bone retract from the thrust plate to the extent that a gap appeared. The extent of radiologically evident bone contact with the flat surface of the thrust plate, as a consequence of the bone remodeling behavior, is described and retrospectively classified. The average Harris hip score increased from 51 points preoperatively to 96 points postoperatively at the last follow-up. Survivorship analysis according to Kaplan-Meier showed a survival rate of 98% after 6 years, with no further losses up to the end of the 8-year follow-up period. PMID- 15660067 TI - Retrieval study of tibial baseplate fracture after total knee arthroplasty. AB - Twenty-five cases of fractured metal tibial baseplates were reviewed from retrieved knee arthroplasty. A total of 74 cases have been reported, including the 25 in this series. Baseplate fracture is an avoidable problem with careful choice of prosthesis, attention to surgical detail, and satisfactory postoperative follow-up. There is a relatively short duration of symptoms before fracture, (mean, 10.8 months; range, 1-36 months). Patients should also be advised to self-report if there is sudden onset of pain or any symptoms of instability or mechanical failure, such as squeaks, clicks, or swelling. Early revision should be considered if there is concern about potential baseplate fracture as delay may compromise the ultimate result. A classification for these baseplate fractures is proposed, and the multifactorial etiology is discussed. PMID- 15660069 TI - Outcome following stabilization of type B1 periprosthetic femoral fractures. AB - We present results of internal stabilization of 16 type B1 periprosthetic femoral fractures following total or hemiarthroplasty of the hip. Seven patients had cemented stems, and the rest had cementless, extensively hydroxyapatite-coated stems. Fourteen were managed by a cable-grip plating system, 1 by Dynamic compression plate, and 1 had insertion of cables only. Minimum follow-up was 1 year. Four patients had a major complication: 2 had deep infection; 1 had nonunion requiring amputation; and 1 had persistent hip pain requiring removal of all metalwork, including the hip prosthesis. The average time to healing in the remaining patients was 16.3 weeks. Harris Hip Scores dropped from an average of 86.8 preoperatively to 73.4 on last follow-up. Periprosthetic fractures are a significant injury, with a high risk of complications. PMID- 15660070 TI - Bony ankylosis following total knee arthroplasty: a case report. AB - While bony ankylosis secondary to heterotopic ossification (HO) following total hip arthroplasty (THA) is well known to produce disability in the hip, much less has been written about the development of heterotopic ossification about the knee after total knee arthroplasty (TKA). Additionally, to our knowledge, there has been no published case of complete bony ankylosis of the knee following total knee arthroplasty. We present such a case in a 53 year old male who underwent a primary total knee arthroplasty after resolution of septic arthritis. PMID- 15660071 TI - Ginkgo biloba: persistent bleeding after total hip arthroplasty caused by herbal self-medication. AB - We describe a case of persistent postoperative bleeding following total hip arthroplasty in which, initially, no obvious cause was identified. The patient was taking ginkgo biloba extract, an unlicensed herbal medicine taken to improve mental alertness. Ginkgo biloba is an anticoagulant that inhibits platelet activating factor and is contraindicated with aspirin. Four weeks' postoperatively, the ginkgo was stopped, and the bleeding stopped 6 weeks later. PMID- 15660072 TI - Limb salvage and abductor reattachment using a custom prosthesis with porous tantalum components. AB - Segmental bone loss secondary to trauma and infection often requires prosthetic components to provide for limb salvage and functional restoration. Current limb salvage techniques and implants are associated with up to a 46% rate of loosening and eventual failure [Clin Orthop Rel Res 1999;358:8]. This case report describes the use of a custom prosthesis for salvage total hip arthroplasty after a proximal femoral resection, for a patient with a history of a comminuted hip fracture and subsequent postoperative infection. Two component parts of the salvage prosthesis were a modular porous tantalum sleeve (Trabecular Metal, Zimmer TMT, Allendale, NJ) and a porous tantalum inset and washer intended for reattachment of the abductors. Throughout the 3-year follow-up of this patient, functional recovery and pain relief were obtained with few limitations. PMID- 15660073 TI - The correction of severe varus deformity in total knee arthroplasty by tibial component downsizing and resection of uncapped proximal medial bone. PMID- 15660074 TI - Total knee arthroplasty in hemophilic arthropathy. PMID- 15660075 TI - Preemptive rofecoxib and dexamethasone for prevention of pain and trismus following third molar surgery *. AB - Objective The goal of this preliminary randomized prospective clinical trial was to compare the analgesic efficacy and the reduction in trismus of preoperative rofecoxib, intraoperative dexamethasone, and both rofecoxib and dexamethasone following third molar extraction surgery. Study design Thirty-five subjects requiring surgical removal of at least 1 partial bony impacted mandibular third molar were invited to participate in this double-blind and double-dummy placebo controlled clinical trial. Subjects were randomly assigned into 1 of 4 treatment groups: (1) placebo po preoperatively and placebo IV intraoperatively; (2) rofecoxib 50 mg po preoperatively and placebo IV intraoperatively; (3) placebo po preoperatively and dexamethasone10 mg IV intraoperatively; and (4) rofecoxib 50 mg po preoperatively and dexamethasone 10 mg IV intraoperatively. Subjects completed a diary assessing postoperative pain onset and intensity using categorical and visual analogue scales. Interincisal opening was assessed 1, 2, 3, and 7 days postoperatively using a Therabite ruler. Results This randomized controlled clinical trial enrolled 35 subjects. Two subjects did not meet the inclusion criteria and 4 did not return completed diaries. The mean age of the remaining 29 subjects (11 males, 18 females) was 22.8 years (+/- 0.6 year). The active treatments tended to delay the need for initial pain medication. When compared to other active treatments and to placebo, the combination of preoperative rofecoxib and intraoperative dexamethasone significantly reduced initial pain intensity ( P < .05). Baseline interincisal opening was 52.6 mm (+/- 6.2). The greatest decrease in interincisal opening was 43.3% for the placebo group at 24 hours. Preoperative rofecoxib alone showed a decrease in interincisal opening of 42.3% ( P = ns) at 24 hours. Intraoperative dexamethasone alone showed a decrease in the interincisal opening of 24.1% of baseline ( P < .05 vs placebo). The group receiving the combination of rofecoxib and dexamethasone showed a decrease in interincisal opening of 23.7% of baseline ( P < .05 vs placebo). Conclusions The results of this trial indicate that the use of intraoperative dexamethasone is an effective therapeutic strategy for limiting trismus following surgical removal of impacted third molars. The combination of preoperative rofecoxib 50 mg and intraoperative dexamethasone 10 mg was most effective in minimizing pain and trismus following third molar surgery. PMID- 15660076 TI - Hematogenous infection of the human temporomandibular joint. AB - Objective This study was designed to discover the relationship between bacteremia and the presence of specific bacterial species in the synovial fluid of the human temporomandibular joint (TMJ). Study design Sixteen volunteers (female to male, 1:2.2; average age, 30.00 +/- 9.93 years) who received operations via intraoral incision participated in this study. Samples from the blood and TMJ synovial fluid of the patients were taken preoperatively and postoperatively and analyzed by PCR assays with specific primers for the clinically important bacteria Staphylococcus aureus , Streptococcus mitis , and beta-hemolytic Streptococcus . Results PCR-based assays revealed the occurrence of S aureus in 68.8% and 50% of the total blood and synovial fluid samples collected immediately after the operations, which statistically differed from the preoperative samples ( P < .05). However, the frequency of the other bacteria, S mitis and beta-hemolytic Streptococcus , in pre- and postoperative blood samples had no such a difference ( P > .05). The chi-square test showed a significant association between the presence of S aureus in the blood and in TMJ synovial fluid (chi 2 = 6.409, P < .05), and the probability of hematogenous infection of the TMJ was estimated as 55.5%. Conclusion Hence, the data obtained provided evidence that the invasion of S aureus into TMJ synovial fluid was due to the presence of these bacteria in the blood. To our knowledge, this is the first report of the bacteremia-related mechanism of S aureus invasion in the human TMJ. PMID- 15660077 TI - A complication of lingual piercing: A case report. AB - Piercing is popular among young people, who view this practice as a sign of marginality, beauty, or group identity. The complications of lingual piercing include infection, pain, bleeding, edema, inhalation, dental trauma, contact lesions, and oral interferences. The objective of this case report is to present a case of lingual piercing in which the piercing had become partially embedded in the interior of the tongue. PMID- 15660078 TI - Anesthetic management of a patient with celiac disease. AB - A 22-year-old female with an extensive medical and surgical history that included mental retardation presented for dental treatment under conscious sedation at a special needs clinic. The patient's history revealed a delayed diagnosis of celiac disease (CD) that resulted in growth retardation with poor muscle development. Because of the patient's delayed diagnosis of CD and history of difficult airway management, special considerations were necessary for proper anesthesia administration. PMID- 15660079 TI - Improving dental treatment for the medically complicated patient. PMID- 15660080 TI - Bisphosphonate-associated oral osteonecrosis. PMID- 15660081 TI - Management of parotid duct injuries. AB - Injuries to the buccal region of the face can carry multiple complications due to the complex anatomy that lies within. The facial nerve and the parotid duct can be easily injured by sharp or penetrating trauma to the cheek. The purpose of this paper is to present the full spectrum of current treatment modalities available to manage these injuries. The anatomy of the parotid gland and duct are described, and surgical techniques and therapeutic alternatives for the immediate and delayed treatment of the parotid duct injuries are reviewed. Clinical cases are presented to illustrate the treatment options outlined. PMID- 15660082 TI - Lag screw osteosynthesis of fractures of the mandibular condyle: potential benefit of preoperative planning using multiplanar CT reconstruction. AB - OBJECTIVE: The purpose of the study was to develop a method of multiplanar CT reconstruction in fractures of the mandibular condyle in order to evaluate the feasibility of lag screw osteosynthesis. STUDY DESIGN: Angulated axial and sagittal images of the mandibular ramus were calculated using collected CT data. In 183 mandibular rami the optimal position of the screw axis was determined retrospectively. The thickness of the covering bone as well as the degree of screw exposure were measured. RESULTS: Application of the method led to medial (28.4% of cases) or lateral (3.8% of cases) correction of the usual screw axis. In 1.6% of cases an extraosseous position of screws could not be avoided by this correction. CONCLUSIONS: Evaluation of the feasibility of lag screw osteosynthesis by this method is possible. Medial or lateral change of the axis ensures the optimal intraosseous screw position. Intraosseous screw placement is almost always possible. PMID- 15660083 TI - In vivo experimental study on bone regeneration in critical bone defects using an injectable biodegradable PLA/PGA copolymer. AB - OBJECTIVES: An assessment was done of the bone-healing rate after implantation of a polylactide/polyglycolide copolymer (PLA-PGA) 50/50 dispersed in aqueous solution of PGA and dextran, used as bone substitutes in an animal model. STUDY DESIGN: Two groups of 5 rabbits each were used. In both the femoral condyles, a critical size defect of 6x10 mm was made. On the right side PLA/PGA was inserted; the left side remained empty. Thirty and 90 days after surgery the animals were killed. RESULTS: Defects left unfilled showed no spontaneous healing after 30 and 90 days. Sites filled with experimental materials showed new bone ranging between 11.46% and 76.82% after 30 days, and 75.98% and 95.34% after 90 days. Histomorphometry showed an increase in bone maturation between day 30 and 90 in experimental sites. At day 90, no statistical difference was seen as compared to normal bone. CONCLUSION: PLA/PGA copolymer dispersed in hydrosoluble matrix seems to be suitable as osteoconductive material in critical size defects. PMID- 15660085 TI - Desmoplastic ameloblastoma featuring basal cell ameloblastoma: a case report. AB - The desmoplastic ameloblastoma is a histological variant of ameloblastoma. The neoplastic epithelial islands seen in desmoplastic ameloblastoma are small and ameloblastic cells are rare. Basal cell ameloblastoma is also a rare variant of ameloblastoma, in which the tumor is composed of more primitive cells and has even fewer features of peripheral palisading. This report describes the case of a 17-year-old female with an ameloblastoma in the right anterior maxilla. Orthopantomography and computed tomography showed a well-defined lesion in the right maxilla. A partial maxillectomy for tumor resection was performed under general anesthesia. Histologically, ameloblastic tumor cells were seen with dense collagenous stroma and the tumor cells showed primarily basal cell variants of ameloblastoma. After 7 years of follow-up, clinical and radiographic examinations have revealed no evidences of recurrence. PMID- 15660084 TI - Prevention of alveolar osteitis with chlorhexidine: a meta-analytic review. AB - OBJECTIVE: The objective of this study was to assess if chlorhexidine (CHX) rinse decreases the occurrence of alveolar osteitis (AO) following third molar removal. STUDY DESIGN: A literature search identified 7 randomized prospective clinical trials reporting incidence of AO following removal of mandibular third molars. Studies were combined into 2 groups and summary relative risks were calculated for each group. One group of studies evaluated rinsing on the day of surgery only and the second group of studies rinsed at least on the day of surgery and several days after surgery. RESULTS: The relative risk for the single rinse group was 1.36 (95% confidence interval [CI] 0.80, 2.33), P>.05, whereas for the multiple rinse group, the relative risk was 1.90 (95% CI 1.46, 2.47), P<.05. CONCLUSION: Rinsing with CHX on the day of surgery and several days after may reduce the incidence of AO. PMID- 15660086 TI - Flight-associated inferior alveolar nerve paresthesia: a case report. PMID- 15660087 TI - Fatigue and fatigue-related symptoms in an orofacial pain population. AB - OBJECTIVES: The aims of this study were to investigate the presence and magnitude of self-reported fatigue and fatigue-related symptoms and to determine whether fatigue can be distinguished as a unique clinical symptom in a sample of patients diagnosed with chronic temporomandibular joint or masticatory muscle pain. STUDY DESIGN: Fifty-five chronic TMD patients and 55 age-, sex-, and education-matched healthy volunteers completed a battery of 4 different fatigue measures as well as the SCL90-R, MPI, and PSQI. RESULTS: Fatigue and fatigue-related symptoms were reported significantly more often by chronic TMD patients than by healthy volunteers. MANCOVAs with somatization, depression, anxiety, general activity level, and sleep disturbances as covariates eliminated the differences between patients and controls. Stepwise regression showed that fatigue and fatigue related symptoms did not appear to be a unique clinical symptom of TMD patients, but merely emerged as somatic symptoms accounted for by somatization and depression. CONCLUSION: Fatigue and fatigue related symptoms may be symptoms of somatization and depression in this sample of chronic TMD patients. PMID- 15660088 TI - Multiple suppurative cystic lesions of the lips and buccal mucosa: a case of suppurative stomatitis glandularis. AB - Cheilitis glandularis (CG) is a rare inflammatory salivary gland disease that usually affects the lips. Although the etiology of CG is still unknown, it is believed to be a hereditary disease with an autosomal dominant pattern of inheritance. Three clinical presentations of CG are described in the literature: simple, superficial suppurative, and deep suppurative. A case of deep suppurative CG that extended to the buccal mucosa has been previously reported as suppurative stomatitis glandularis (SSG). Here we report a case of SSG in a 64-year-old white female with a history of bilateral renal transplants for adult polycystic kidney disease, who presented with painful swollen lips and bilateral buccal mucosal lesions. The diagnosis and management of the case is discussed. To the best of our knowledge, this is the second report of SSG, a rare condition affecting the minor salivary glands in the oral cavity. PMID- 15660090 TI - Morphologic characteristics of masseter muscle in cleidocranial dysplasia: a report of 3 cases. AB - Cleidocranial dysplasia (CCD) is a rare autosomal dominant skeletal dysplasia. In order to define the morphologic characteristics of the masseter muscle incidental to bone abnormalities, we present 3 cases of CCD with the masseter muscle thickness and maxillofacial bone abnormalities, using computed tomography (CT) and panoramic radiographs. In CCD patients (a) the masseter muscles were less thick than in age- and sex-matched control subjects, (b) the zygomatic arch was discontinuous with the hypoplastic zygomatic bone, (c) the ascending ramus of the mandible had parallel-sided borders, and (d) the coronoid process pointed upwards and/or posteriorly. We have concluded the masseter muscles are less thick than normal, alongside the maxillofacial bone abnormalities in CCD patients. PMID- 15660089 TI - Mutans streptococci in xerostomic cancer patients after pilocarpine therapy: a pilot study. AB - OBJECTIVES: Opiod- and/or radiation-induced xerostomia in cancer patients is frequently associated with elevated levels of cariogenic mutans streptococci (MS). STUDY DESIGN: In a single-center, single blind 8-week clinical trial at The University of Texas M. D. Anderson Cancer Center, and from an initial sample of 32 patients, we evaluated MS counts in 28 cancer patients receiving chronic analgesic treatment for cancer pain. All patients received escalating doses of pilocarpine (Salagen) tablets, either 2.5 mg to 5 mg or 5 mg to 7.5 mg qid for 6 weeks, followed by placebo qid for a 2-week washout period. Whole resting saliva flow rates (g/5 min) and MS counts were evaluated at pretreatment, 3 weeks, 6 weeks, and 8 weeks. MS samples were obtained by 5-mL saline rinse (15 sec) at each visit prior to sialometry. RESULTS: In 19 patients (59%), MS counts exceeded 10(5) CFU/mL. At the end of the 6-week trial, 96% of patients showed a positive response to pilocarpine following a 30-minute postdosing evaluation (P=.001). MS counts were lower in 17 patients, higher in 6 patients, and nondetectable before and after pilocarpine in 5 patients (P=.03). CONCLUSION: The reduced MS counts associated with improved saliva flow rates following pilocarpine therapy in this short-term pilot study are encouraging, but further investigation in a larger group of patients over a longer study period is indicated. PMID- 15660091 TI - Odontogenic tumors: a review of 319 cases in a Nigerian teaching hospital. AB - OBJECTIVE: This study sought to determine the relative frequency of odontogenic tumors in a Nigerian population and to compare these data with previous reports. STUDY DESIGN: Records of patients seen at the Lagos University Teaching Hospital between January 1980 and December 2003, with histologic diagnosis of odontogenic tumors (based on World Health Organisation classification, 1992), were analyzed. RESULTS: Odontogenic tumors constituted 9.6% of all the biopsies of oral and jaw lesions seen within the period under study. Three hundred and eight (96.6%) were intraosseous, and 11 (3.4%) were peripheral (peripheral odontogenic fibroma=7; peripheral myxoma=3; peripheral ameloblastoma=1). The mean age of patients was 29.9+/-15.6 years (range, 4-85 years). Among these cases, 96.6% of the tumors were benign and 3.4% were malignant. Ameloblastoma with predilection for the mandible was the most frequent odontogenic tumor (63%), followed by adenomatoid odontogenic tumor (AOT) (7.5%), myxoma (6.5%), calcifying epithelial odontogenic cyst (5.3%), and odontogenic fibroma (5.3%). More cases of malignant odontogenic tumors were seen than cases of calcifying epithelial odontogenic tumor and odontomas. The mean ages of patients with AOT, ameloblastic fibroma, and odontoma were significantly lower than those with ameloblastoma ( P<.05). No significant difference was found between the mean ages of patients with benign odontogenic tumors and those with malignant odontogenic tumors ( P=.058). CONCLUSIONS: Odontogenic tumors, especially ameloblastoma, are not considered rare among Nigerians, whereas odontoma, regarded as the most frequent odontogenic tumor in North and South America, is rare. PMID- 15660092 TI - Myoepithelial carcinoma ex pleomorphic adenoma of salivary glands: a problematic diagnosis. AB - We report the case of a 77-year-old woman who presented with a 6-year history of pleomorphic adenoma with multiple recurrences. Despite the clinical suspicion of a malignant process, the detection of myoepithelial carcinoma ex pleomorphic adenoma was not achieved by means of a preoperative biopsy or a frozen section removed intraoperatively, as the lesion only showed mild cytological atypia with no frank malignant features. Only a thorough evaluation of the resected tumor enabled the diagnosis of a malignancy. We discuss the difficulties that may be encountered in attempting to diagnose these rare myoepithelial lesions by means of small biopsy specimens. PMID- 15660093 TI - Fibrolipomatous hamartoma in a patient with tuberous sclerosis: report of a case. AB - Tuberous sclerosis is a rare congenital disorder with an incidence of 1 in 6000 births. The classic triad is seizure, mental deficiency, and angiofibromas. Orofacial manifestations include fibrous hyperplasia, hemangioma, bifid uvula, cleft lip and palate, macroglossia, high arched palate, and enamel defects. Benign tumors of the jaws including desmoplastic fibroma, calcifying odontogenic tumor, and odontogenic myxoma have been recently reported in tuberous sclerosis. This case report adds fibrolipomatous hamartoma of the mandible to this list. PMID- 15660094 TI - A previously unreported association between Nance-Horan syndrome and spontaneous dental abscesses. AB - Atypical dentofacial structures may be the first indicator of other anomalies linked to a syndrome. This case describes the management of a 9-year-old girl referred for the routine management of supernumerary teeth. The anomalous form of her teeth, together with multiple supernumerary units and a history of congenital cataracts, were suggestive of a diagnosis of Nance-Horan syndrome. This is an X linked disorder, in which females usually demonstrate mild expression; this case was unusual in respect to the marked phenotype expressed. Unusually, the girl developed 2 spontaneous abscesses of her noncarious upper incisor teeth; a feature never previously described in this syndrome. This report details the patient's dental management and discusses the possible pathogenesis of the dental abscesses, together with the genetic implications of this syndrome. PMID- 15660095 TI - 3D-CT evaluation of facial asymmetry. AB - OBJECTIVE: Recently, 3-dimensional-computed tomography (3D-CT) imaging has been used in the diagnosis and surgical treatment planning of patients with craniofacial deformities. The present authors have developed a 3D-CT imaging procedure for a 3-dimensional coordinate point evaluation system to assess and diagnose patients with facial asymmetry. STUDY DESIGN: The CT data of 16 subjects was selected retrospectively as the control group from patients who had undergone CT examinations to diagnose conditions other than maxillofacial deformities. Anatomical landmarks modified from orthodontic craniometric (cephalometric) points were defined on the 3D-CT images and the asymmetry index of each point was calculated in millimeters. A diagrammatic chart with a baseline indicating the mean asymmetry indices plus the standard deviation in the control group was designed. The resulting diagrammatic chart was used to evaluate the degree of deformity in facial asymmetry patients. RESULTS AND CONCLUSIONS: The topography of facial asymmetry was assessed. The 3D-CT imaging technique as described herein is a practical method of evaluating the morphology of facial asymmetry. PMID- 15660096 TI - MRI of nasoalveolar cyst: case report. AB - Nasoalveolar cyst is a rare nonodontogenic cyst arising in the nasoalar region of the face. In this case, a 39-year-old female had a nontender, fluctuant, soft, round, and bluish lesion in the right anterior gingiva of the right maxilla. Panoramic and magnetic resonance imaging examinatioins were performed. No abnormality was evident on the radiograph. The lesion was hypointense signal intensity on T1-weighted images and hyperintense on T2-weighted images. On sagittal T2-weighted images, a septum could be seen within the lesion. After injection of contrast medium and acquisition of T1-weighted images, enhancement was not observed in the lesion, but was evident at the periphery. The lesion did not enhance with contrast on T1-weighted images, but the periphery was enhanced. The lesion was surgically excised. Histopathologic examination showed the lesion to be a nasoalveolar cyst. PMID- 15660097 TI - Triticeous cartilage: prevalence on panoramic radiographs and diagnostic criteria. AB - OBJECTIVES: To determine (1) the prevalence of calcified triticeous cartilage on panoramic radiographs of subjects older than 40 years of age, (2) the age and sex distribution of the subjects who have calcified triticeous cartilage, and (3) if the shape of triticeous cartilage differs from that of calcified carotid atheroma. STUDY DESIGN: In this retrospective study, triticeous cartilages were diagnosed from 847 panoramic radiographs. The shapes of triticeous cartilages and calcified carotid atheromas were determined using SigmaScan software. RESULTS: Prevalence of triticeous cartilage was 5.0% in males and 12.0% in females. The prevalence was 9.3% in the 40- to 60-year-old group (n=464) and 7.8% in the group older than 60 years (n=383). The triticeous cartilages were oval with smooth borders, whereas calcified carotid atheromas were mostly linear. CONCLUSIONS: Calcified triticeous cartilage was more prevalent in females. The prevalence was not age dependent. Shape, outline, and location of the triticeous cartilage can help in differentiating it from a carotid atheroma. PMID- 15660098 TI - Microbial status of apical root canal system of human mandibular first molars with primary apical periodontitis after "one-visit" endodontic treatment. AB - OBJECTIVE: To assess the in vivo intracanal microbial status of apical root canal system of mesial roots of human mandibular first molars with primary apical periodontitis immediately after one-visit endodontic treatment. The residual intracanal infection was confirmed by correlative light and transmission electron microscopy. STUDY DESIGN: Sixteen diseased mesial roots of mandibular first molars were treated endodontically, each in one visit. Mesio-buccal canals were instrumented using stainless steel hand files and mesio-lingual canals with a nickel-titanium rotary system. The canals were irrigated with 5.25% sodium hypochlorite (NaOCl) during the instrumentation procedures, rinsed with 10 mL of 17% ethylenediamine tetraacetic acid (EDTA), and obturated with gutta-percha and zinc oxide eugenol cement. Thereafter, the apical portion of the root of each tooth was removed by flap-surgery. The specimens were fixed, decalcified, subdivided in horizontal plane, embedded in plastic, processed, and evaluated by correlative light and transmission electron microscopy. RESULTS: Fourteen of the 16 endodontically treated teeth revealed residual intracanal infection after instrumentation, antimicrobial irrigation, and obturation. The microbes were located in inaccessible recesses and diverticula of instrumented main canals, the intercanal isthmus, and accessory canals, mostly as biofilms. CONCLUSIONS: The results show (1) the anatomical complexity of the root canal system of mandibular first molar roots and (2) the organization of the flora as biofilms in inaccessible areas of the canal system that cannot be removed by contemporary instruments and irrigation alone in one-visit treatment. These findings demonstrate the importance of stringent application of all nonantibiotic chemo mechanical measures to treat teeth with infected and necrotic root canals so as to disrupt the biofilms and reduce the intraradicular microbial load to the lowest possible level so as to expect a highly favorable long-term prognosis of the root canal treatment. PMID- 15660099 TI - Increased plasma glutamate in stroke patients might be linked to altered platelet release and uptake. AB - Experimental studies have shown the role of excitotoxicity in the pathogenesis of ischemic brain lesions, and glutamate levels have been found to be elevated in CSF and plasma from patients, early after stroke. In this study, we investigated whether platelets could be involved in the mechanism of altered plasma glutamate levels after stroke. Forty four patients, from 6 hours to 9 months after ischemic stroke, 15 age-related healthy controls and 15 controls with stroke risk factors or previous transient ischemic attack were enrolled. Glutamate plasma levels, platelet glutamate release after aggregation and platelet glutamate uptake were assessed. Plasma glutamate levels were increased up to 15 days after the ischemic event in stroke patients, and the levels at day 3 were inversely correlated with the neurologic improvement between day 3 and 15. Ex vivo platelet glutamate release was decreased by 70% in stroke patients, suggesting previous in vivo platelet activation. Moreover, platelet glutamate uptake in these patients was decreased by 75% up to 15 days and was still reduced 90 days after stroke. Our data show a prolonged increase of glutamate in plasma after stroke, which might presumably be linked to altered platelet functions, such as excessive release of the amino acid or impaired uptake. PMID- 15660100 TI - Nitric oxide inhibits caspase activation and apoptotic morphology but does not rescue neuronal death. AB - Nitric oxide (NO) has been shown to inhibit apoptotic cell death by S nitrosylation of the catalytic-site cysteine residue of caspases. However, it is not clear whether in neurons NO-mediated caspase inactivation leads to improved cell survival. To address this issue, we studied the effect of NO donors on caspase activity and cell survival in cortical neuronal culture treated with the apoptosis inducer staurosporine (STS) and camptothecin. In parallel, cell viability was assessed by the MTS assay and MAP2 staining. We found that NO donors ((+/-)-S-nitroso-N-acetylpenicillamine, S-nitrosoglutathione, and NONOates) dose-dependently inhibited caspase-3 and -9 activity induced by STS and camptothecin. The reduction in caspase-3 activity was, in large part, because of the blockage of the proteolytic conversion of pro-caspase-3 to active caspase-3. NO donors also inhibited the appearance of the classical apoptotic nuclear morphology. However, inhibition of both caspase activity and apoptotic morphology was not associated with enhancement of cell viability. Thus, inhibition of caspase and apoptotic morphology by NO donors does not improve neuronal survival. The data suggest that inhibition of caspase by NO unmasks a caspase-independent form of cell death. A better understanding of this form of cell death may provide new strategies for neuroprotection in neuropathologies, such as ischemic brain injury, associated with apoptosis. PMID- 15660101 TI - G-CSF reduces infarct volume and improves functional outcome after transient focal cerebral ischemia in mice. AB - Growth factors possess neuroprotective and neurotrophic properties in vitro, but few have been extensively studied in vivo after stroke. In the present study, we investigated the potential functional benefits of granulocyte colony-stimulating factor (G-CSF) administration after focal cerebral ischemia. Male mice underwent 60-minute middle cerebral artery occlusion (MCAO) and received G-CSF (50 microg/kg, subcutaneously) or vehicle (saline) at the onset of reperfusion. Granulocyte colony-stimulating factor-treated mice killed at 48 hours after MCAO revealed a >45% reduction (P<0.05) in lesion volume. In terms of body weight recovery, and in tests of motor (grid test and rotarod) and cognitive ability (water maze), MCAO significantly worsened the outcome in vehicle-treated mice as compared with shams (P<0.05). However, G-CSF treatment was beneficial as, compared with vehicle, this significantly improved weight recovery and motor ability. This effect was most apparent on the water maze where G-CSF-treated mice were indistinguishable from shams in terms of acquiring the task. These results indicate long-term beneficial effects of a single dose of G-CSF administered on reperfusion, and illustrate the need to further investigate the mechanisms of G CSF action. PMID- 15660102 TI - Association of 14-3-3gamma and phosphorylated bad attenuates injury in ischemic astrocytes. AB - Our recent findings indicate an induced upregulation of 14-3-3gamma mRNA and protein in ischemic cortical astrocytes. Despite being brain-specific, the functional role of 14-3-3gamma in the brain still remains largely unknown. In this study, we show that among all the 14-3-3 isoforms, only the gamma isoform is inducible under ischemia in astrocytes. Furthermore, this upregulation of 14-3 3gamma may play a specific protective role in astrocytes under ischemia. Overexpression experiments and antisense treatment show that an elevation of 14-3 3gamma protein in astrocytes promotes survival, while a decrease in 14-3-3gamma enhances apoptosis in astrocytes under ischemia. Under ischemia, endogenous 14-3 3gamma binds p-Bad, thus preventing Bad from entering mitochondria to initiate apoptosis. Therefore, 14-3-3gamma is selectively induced during ischemia to protect astrocytes from apoptosis through p-Bad-related signaling. PMID- 15660104 TI - Hypotension in the newborn: who needs hydrocortisone? PMID- 15660103 TI - Changes in cerebral blood flow and cerebral oxygen metabolism during neural activation measured by positron emission tomography: comparison with blood oxygenation level-dependent contrast measured by functional magnetic resonance imaging. AB - The discrepancy between the increases in cerebral blood flow (CBF) and CMRO2 during neural activation causes an increase in venous blood oxygenation and, therefore, a decrease in paramagnetic deoxyhemoglobin concentration in venous blood. This can be detected by functional magnetic resonance imaging (fMRI) as blood oxygenation level-dependent (BOLD) contrast. In the present study, changes in the cerebral oxygen extraction fraction (OEF) that corresponds to the ratio of CMRO2 to CBF, and in the BOLD signal during neural activation, were measured by both positron emission tomography (PET) and fMRI in the same human subjects. C15O, 15O2, and H2(15)O PET studies were performed in each subject at rest (baseline) and during performance of a right-hand motor task. Functional magnetic resonance imaging studies were then performed to measure the BOLD signal under the two conditions. During performance of the motor task, a significant increase in CBF and a significant decrease in OEF were observed in the left precentral gyrus, left superior frontal gyrus, right precentral gyrus, right cingulate gyrus, and right cerebellum. A significant positive correlation was observed between changes in the CBF and the BOLD signal, and a significant negative correlation was observed between changes in the OEF and the BOLD signal. This supports the assumption on which BOLD contrast studies during neural activation are based. PMID- 15660105 TI - An infant with Kasabach-Merritt syndrome with associated renal hematoma and intussusception. AB - A neonate that had a forearm hemangioma, gross hematuria, and a renal parenchymal hematoma was found to have Kasabach-Merritt syndrome. He subsequently returned to our hospital and was found to have an intussusception secondary to an intestinal hemangioma. PMID- 15660106 TI - McKusick-Kaufman Syndrome (MK catalogue #236700) presenting prenatally as fetal abdominal mass. PMID- 15660107 TI - A universal trend of amino acid gain and loss in protein evolution. AB - Amino acid composition of proteins varies substantially between taxa and, thus, can evolve. For example, proteins from organisms with (G + C)-rich (or (A + T) rich) genomes contain more (or fewer) amino acids encoded by (G + C)-rich codons. However, no universal trends in ongoing changes of amino acid frequencies have been reported. We compared sets of orthologous proteins encoded by triplets of closely related genomes from 15 taxa representing all three domains of life (Bacteria, Archaea and Eukaryota), and used phylogenies to polarize amino acid substitutions. Cys, Met, His, Ser and Phe accrue in at least 14 taxa, whereas Pro, Ala, Glu and Gly are consistently lost. The same nine amino acids are currently accrued or lost in human proteins, as shown by analysis of non synonymous single-nucleotide polymorphisms. All amino acids with declining frequencies are thought to be among the first incorporated into the genetic code; conversely, all amino acids with increasing frequencies, except Ser, were probably recruited late. Thus, expansion of initially under-represented amino acids, which began over 3,400 million years ago, apparently continues to this day. PMID- 15660108 TI - Functional imaging with cellular resolution reveals precise micro-architecture in visual cortex. AB - Neurons in the cerebral cortex are organized into anatomical columns, with ensembles of cells arranged from the surface to the white matter. Within a column, neurons often share functional properties, such as selectivity for stimulus orientation; columns with distinct properties, such as different preferred orientations, tile the cortical surface in orderly patterns. This functional architecture was discovered with the relatively sparse sampling of microelectrode recordings. Optical imaging of membrane voltage or metabolic activity elucidated the overall geometry of functional maps, but is averaged over many cells (resolution >100 microm). Consequently, the purity of functional domains and the precision of the borders between them could not be resolved. Here, we labelled thousands of neurons of the visual cortex with a calcium sensitive indicator in vivo. We then imaged the activity of neuronal populations at single-cell resolution with two-photon microscopy up to a depth of 400 microm. In rat primary visual cortex, neurons had robust orientation selectivity but there was no discernible local structure; neighbouring neurons often responded to different orientations. In area 18 of cat visual cortex, functional maps were organized at a fine scale. Neurons with opposite preferences for stimulus direction were segregated with extraordinary spatial precision in three dimensions, with columnar borders one to two cells wide. These results indicate that cortical maps can be built with single-cell precision. PMID- 15660109 TI - Control of DNA replication and its potential clinical exploitation. AB - Multiple conserved mechanisms limit DNA replication to once per cell cycle. One vital level of control focuses on the loading of the heterohexameric ring of minichromosome maintenance proteins (MCMs) onto chromatin in the hierarchical assembly of the pre-replication complex at origins of replication. An essential role in proliferation for MCMs and their regulators makes them potentially important biomarkers for routine clinical use in cancer detection and prognosis. PMID- 15660110 TI - Nature and nurture - lessons from chemical carcinogenesis. AB - The roles of genetic constitution versus environmental factors in cancer development have been a matter of debate even long before the discovery of 'oncogenes'. Evidence from epidemiological, occupational and migration studies has consistently pointed to environmental factors as the major contributing factors to cancer, so it seems reasonable to discuss the importance of chemical carcinogenesis in the present 'age of cancer genetics'. PMID- 15660112 TI - Fas-ligand gene silencing in basal cell carcinoma tissue with small interfering RNA. AB - Basal cell carcinoma (BCC) is the most frequent cancer in the Caucasian population. Cells of BCC strongly express Fas-ligand (FasL), a member of the tumor necrosis family, which induces apoptosis in Fas receptor-expressing cells. It has been suggested that by expression of FasL, BCC cells may evade the attack of Fas-positive immune effector cells allowing the tumor to expand. Thus, downregulation of FasL should prime BCC to the assault of immune effector cells. Recently, it has been shown that RNA interference is a highly successful approach to specifically silence a gene of interest in single cells and some animal models. However, RNAi in human tissues has not been shown so far. Here, we provide evidence that small interfering RNAs (siRNAs) efficiently transfect tumor tissue ex vivo and silence the gene of interest. We demonstrate that a specific siRNA efficiently downregulates FasL not only in FasL-positive indicator cells but also in surgically excised BCC tissue at both the protein and the mRNA level. The successful transfection of tumor tissues with siRNAs now allows to test the function of the molecule under study and opens up the investigation of other target genes in the tumor. PMID- 15660111 TI - Long-term preservation of retinal function in the RCS rat model of retinitis pigmentosa following lentivirus-mediated gene therapy. AB - The Royal College of Surgeons (RCS) rat is a well-characterized model of autosomal recessive retinitis pigmentosa (RP) due to a defect in the retinal pigment epithelium (RPE). It is homozygous for a null mutation in the gene encoding , a receptor tyrosine kinase found in RPE cells, that is required for phagocytosis of shed photoreceptor outer segments. The absence of Mertk results in accumulation of outer segment debris. This subsequently leads to progressive loss of photoreceptor cells. In order to evaluate the efficacy of lentiviral mediated gene replacement therapy in the RCS rat, we produced recombinant VSV-G pseudotyped HIV-1-based lentiviruses containing a murine Mertk cDNA driven by a spleen focus forming virus (SFFV) promoter. The vector was subretinally injected into the right eye of 10-day-old RCS rats; the left eye was left untreated as an internal control. Here, we present a detailed assessment of the duration and extent of the morphological rescue and the resulting functional benefits. We examined animals at various time points over a period of 7 months by light and electron microscopy, and electroretinography. We observed correction of the phagocytic defect, slowing of photoreceptor cell loss and preservation of retinal function for up to 7 months. This study demonstrates the potential of gene therapy approaches for the treatment of retinal degenerations caused by defects specific to the RPE and supports the use of lentiviral vectors for the treatment of such disorders. PMID- 15660114 TI - New complex Ad vectors incorporating both rtTA and tTS deliver tightly regulated transgene expression both in vitro and in vivo. AB - Regulation of transgene expression is a major goal of gene therapy research. Previously, we have developed a complex adenovirus (Ad) vector with tetracycline regulated expression of a Fas ligand (FasL)-green fluorescent protein (GFP) fusion protein. This vector delivered high levels of activity that was regulated by doxycycline. However, this regulation was limited by the low but significant background activity of the TRE promoter. Recently, the Tet-regulated transcriptional silencer, tTS, was reported to suppress efficiently basal TRE activity without affecting induced expression levels. Here, we report development of Ad vectors that incorporate tTS in combination with that of reverse transactivator (rtTA) coupled with TRE promoter driving transgene expression. Incorporation of tTS improved control of transgene expression in vitro, so that an induction range of over three orders of magnitude was achieved in some cell lines. Effective regulation of transgene expression was also seen in a mouse model in vivo, following systemic vector delivery. In the case of FasL-GFP expression, significant improvement in the control of apoptotic activity both in vitro and in a mouse hepatotoxicity model was demonstrated when using rtTA-tTS vectors. In conclusion, a highly effective transgene regulation system, deliverable by a single adenoviral vector, is now available. PMID- 15660115 TI - Association analysis of the Gln223Arg polymorphism in the human leptin receptor gene, and traits related to obesity in Mexican adolescents. AB - Polymorphisms of leptin receptor (LEPR) may contribute to a common form of obesity and, as a consequence, obesity-related diseases. We evaluated the potential role of genetic variation at the LEPR gene in heart sympathetic activity and other traits related to obesity in Mexican adolescents. Adolescents aged between 12 and 17 years, with steady body weight for the last 3 months were included. We evaluated anthropometric measurements, blood pressure, seric glucose, insulin, leptin levels, heart sympathetic activity (by electrocardiograph monitoring at rest), and the Gln223Arg and Pro1019Pro LEPR polymorphisms in each subject. In total, 103 adolescents (55 obese and 48 nonobese) were included. The group of obese adolescents showed higher sympathetic activity, blood pressure, glucose, insulin, and leptin levels. The genotype frequencies for the two polymorphisms were found to be in Hardy-Weinberg equilibrium. There was no difference in the genotype frequencies for Gln223Arg or Pro1019Pro polymorphisms between obese and nonobese adolescents. However, there was a higher prevalence of Gln223 allele among subjects with higher insulin levels (0.72 vs 0.57; P = 0.04 for adolescents with insulin levels higher and lower than 100 pmol/l, respectively). According to Gln223Arg polymorphism, those with Gln allele (Gln/Gln and Gln/Arg) had higher heart sympathetic activity, body fat percentage, and leptin levels. To conclude, our results support the hypothesis that Gln223Arg polymorphism of LEPR in Mexican adolescents is associated with haemodynamic and metabolic disturbances related to obesity. PMID- 15660116 TI - How many antihypertensives do patients need to achieve a target blood pressure? PMID- 15660117 TI - Glucocorticoid remediable aldosteronism (GRA) screening in hypertensive patients from a primary care setting. PMID- 15660118 TI - Hypertension--still an important cause of heart failure? AB - Hypertension has been the single most important risk factor for heart failure until the last few decades. Now, it is frequently claimed that atherosclerotic coronary artery disease dominates as the major underlying cause, and hypertension is of lesser importance. We here review evidence regarding the contribution of hypertension to heart failure in the recent decades. It is not possible, in our view, to be confident of the relative importance of hypertension and coronary artery disease since there are significant limitations in the available data. The often-questionable diagnostic criteria used in defining heart failure is one such limitation. The absence or inadequacy of blood pressure recordings over the years prior to a diagnosis of heart failure seriously hinders the reaching of firm conclusions in many reports. Extrapolations from aetiological observations in one racial group to those in other racial groups, and from highly selected study groups in tertiary referral centres to patients with heart failure in primary and secondary care, may not be justified. Finally, the situation of heart failure primarily due to impaired left ventricular diastolic function, where hypertension is a frequent precursor, is often ignored in discussions of aetiology. Our view is that hypertension remains and probably is the single most, important modifiable risk factor for cardiac failure in some races and countries, where the dominant cardiac abnormality is left ventricular diastolic dysfunction. The situation is less clear for patients with heart failure primarily due to left ventricular systolic dysfunction. PMID- 15660119 TI - Different effects of blood pressure on mortality from stroke subtypes depending on BMI levels: a 19-year cohort study in the Japanese general population--NIPPON DATA80. AB - To evaluate the relationship between blood pressure and cerebrovascular death depending on body mass index (BMI) levels, we analysed a database of 9338 subjects from the National Integrated Project for Prospective Observation of Non communicable Disease and its Trends in the Aged, which was originally conducted a baseline survey in 1980 and followed up in 1999. Relative risk (RR) and a 95% confidence interval (CI) of death from total stroke, cerebral infarction, and intracerebral haemorrhage after adjusting for age, sex, serum cholesterol, albumin, glucose, the use of antihypertensive agents, a past history of diabetes, BMI, smoking, and drinking were estimated with the Cox-proportional hazard model in the BMI tertile groups of a representative Japanese population. Cutoff points of BMI tertiles are 21.2 and 23.8 kg/m2. The results indicated that a 10 mmHg systolic blood pressure (SBP) increase was associated with mortality from intracerebral haemorrhage at low and middle BMI groups (RR= 1.38 and 1.23; 95% CI=1.17-1.62 and 1.03-1.47, respectively). SBP was positively associated with mortality from cerebral infarction in middle and high BMI groups (RR=1.19 and 1.21; 95% CI=1.06-1.33 and 1.06-1.38, respectively). The effects of diastolic blood pressure on intracerebral haemorrhage and infarction had the same tendency as those of SBP. These results suggested that the causal effect of blood pressure on stroke subtypes might be modified by BMI. PMID- 15660121 TI - Ultrafiltration improves aortic compliance in haemodialysis patients. AB - An elevated pulse pressure leads to an increased pulsatile cardiac load, and results from arterial stiffening. The aim of our study was to test whether a reduction in volume overload by ultrafiltration (UF) during haemodialysis (HD) leads to an improvement of aortic compliance. In 18 patients, aortic compliance was estimated noninvasively before and after HD with UF using a pulse pressure method based on the Windkessel model. This technique has not been applied before in a dialysis population, and combines carotid pulse contour analysis by applanation tonometry with aortic outflow measurements by Doppler echocardiography. The median UF volume was 2450 ml (range 1000-4000 ml). The aortic outflow volume after HD (39 ml; 32-53 ml) was lower (P=0.01) than before (46 ml; 29-60 ml). Carotid pulse pressure after HD (42 mmHg; 25-85 mmHg) was lower (P=0.01) than before (46 mmHg; 35-93 mmHg). Carotid augmentation index after HD (22%; 3-30%) was lower (P=0.001) than before (31%; 7-53%). Carotid femoral pulse wave velocity was not different after HD (8.7 m/s; 5.6-28.9 m/s vs 7.7 m/s; 4.7-36.8 m/s). Aortic compliance after HD (1.10 ml/mmHg; 0.60-2.43 ml/mmHg) was higher (P=0.02) than before (1.05 ml/mmHg; 0.45-1.69 ml/mmHg). The increase in aortic stiffness in HD patients is partly caused by a reversible reduction of aortic compliance due to volume expansion. Volume withdrawal by HD moves the arterial wall characteristics back to a more favourable position on the nonlinear pressure-volume curve, reflected in a concomitant decrease in arterial pressure and improved aortic compliance. PMID- 15660120 TI - Usefulness of QT-interval parameters for cardiovascular risk stratification in type 2 diabetic patients with arterial hypertension. AB - QT-interval parameters are potential indicators of increased cardiovascular risk. We evaluated prospectively their prognostic value, in relation to other risk markers, for cardiovascular fatal and nonfatal events in a cohort of 271 hypertensive type 2 diabetic outpatients. QT intervals were measured from 12-lead standard ECGs obtained on admission and maximum rate-corrected QT-interval duration and QT-interval dispersion (QTd) calculated. Clinical and laboratory data and 2-D echocardiograms (available in 126 patients) were recorded. Survival analyses included Kaplan-Meier survival curves, uni and multivariate Cox proportional-hazards models. After a median follow-up of 55 months (range 2-84), 68 total fatal or nonfatal cardiovascular events and 34 cardiovascular deaths (24 of them from cardiac causes) were observed. In multivariate Cox analysis, QTd was an independent predictor for total cardiovascular events (HR: 1.16, 95% CI: 1.01 1.34, for each 10 ms increments) and for cardiac deaths (HR: 1.28, 95% CI: 1.01 1.60). Other independent risk indicators for cardiovascular morbidity and mortality were echocardiographic left ventricular hypertrophy (Echo-LVH), serum triglycerides, presence of pre-existing cardiac and peripheral arterial disease, age, diabetes duration, heart rate and the presence of frequent ventricular premature contractions on ECG. The combination of QTd and Echo-LVH improved cardiovascular risk stratification compared with either alone, the presence of both prolonged QTd (>65 ms) and Echo-LVH was associated with a 3.2-fold (95% CI: 1.7-6.1) increased risk of a first cardiovascular event and a 5.9-fold (95% CI: 2.1-16.4) increased risk of cardiovascular death. Thus, QT provided additive prognostic information for cardiovascular morbidity and mortality beyond that obtained from conventional risk markers, including Echo-LVH, in type 2 diabetic patients with arterial hypertension. PMID- 15660122 TI - Current status of angiotensin receptor blockers for the treatment of cardiovascular diseases: focus on telmisartan. AB - Poorly controlled hypertension is a major risk for cardiovascular morbidity and mortality, strokes, heart failure and renal failure. Despite these devastating complications, blood pressure control of or = 12%. RESULTS: The ratio of FEV1 to forced vital capacity (FVC) was less than 70% in 21 (65.6%) patients, but EFL as demonstrated by NEP (EFL-N) was present in only 10 patients (31.3%). The baseline EFL-N was only weakly correlated with FEV1 (r = 0.36, p = 0.04), but not with Rint or FEV1/FVC. Among the 14 patients with positive BDT results, the change in FEV1 was correlated with the change in Rint (r = -0.69, p < 0.01), but not with the change in EFL-N. CONCLUSIONS: These data suggest that, compared with spirometry and the interrupter method, NEP applied in the sitting position is not sensitive in the assessment of bronchodilator response in patients with obstructive airway disorders. PMID- 15660172 TI - Role of open lung biopsy in patients with diffuse lung infiltrates and acute respiratory failure. AB - BACKGROUND AND PURPOSE: Open lung biopsy (OLB) is the standard procedure for the diagnosis of specific parenchymal lung diseases. The purpose of this study was to investigate the influence of OLB on subsequent treatment strategy and outcome in patients with diffuse lung infiltrates and acute respiratory failure. METHODS: This retrospective review included 32 patients (aged 50.6 +/- 21.7 years) with acute respiratory failure and diffuse pulmonary infiltrates who underwent OLB from 1990-2002. Data analyzed included diagnoses, treatment alterations, 30-day survival, oxygenation status, and histologic results. RESULTS: Specific diagnoses were made in 53.1% of patients (17/32), 23 (71.9%) of whom had acute respiratory distress syndrome (ARDS). Diagnostic yields did not differ with immunity status or ARDS. OLB led to specific decisions of treatment in 46.9% of patients (15/32), and only 7 of these 32 patients (21.8%) survived. Overall mortality was 56.2% (18/32) and was not influenced by pre-OLB oxygenation or histologic results. Although perioperative complications affected 40.6% of patients (13/32), none of the deaths were surgery-related. Complication rates were significantly higher in patients with ARDS (p = 0.04). CONCLUSIONS: OLB is associated with a low perioperative mortality rate and acceptable morbidity rate in patients with diffuse lung infiltrates and acute respiratory failure, including those patients with ARDS. In this study, a specific diagnosis was obtained by OLB in more than half of patients with diffuse pulmonary infiltrates and ARDS. In addition, OLB resulted in either use of a new therapeutic strategy or elimination of unnecessary treatment in nearly one-half of patients (46.9%). PMID- 15660173 TI - Effects of conversion from sirolimus oral solution to tablets in stable Taiwanese renal transplant recipients. AB - BACKGROUND AND PURPOSE: Sirolimus (SRL) has a considerable inter- and intra- individual variability in clearance. Steady-state trough concentration (C(0)) is a reliable index of SRL exposure. This study assessed the effect of conversion of SRL oral solution to tablet form on C(0) in stable renal transplant recipients. METHODS: Twenty two stable renal transplant recipients who had received calcineurin inhibitor (CNI)/SRL solution/ steroid for more than 3 months before conversion from SRL solution to tablets were included. C(0) values of SRL were compared for the periods of use of each dosage form. The relation between liver function and SRL levels was also assessed. RESULTS: With a dose of 0.03 mg/kg/day, SRL solution and tablets achieved a similar dose-adjusted C(0) (mean +/- SEM, 2.9 +/- 0.3 ng/mL/mg) upon conversion. Similar results were found when multiple SRL C(0) values from different dosage form periods were compared. Four patients with persistent liver enzyme elevation had significantly higher dose adjusted SRL C(0) values with both the solution (mean +/- SEM, 4.5 +/- 0.7 vs 2.3 +/- 0.1 ng/mL/mg; p < 0.01) and the tablet formulation (4.0 +/- 0.5 vs 2.6 +/- 0.2 ng/mL/mg; p < 0.05). CONCLUSIONS: Conversion from SRL solution to SRL tablets did not significantly affect the dose-adjusted SRL C(0). The dose-adjusted C(0) of SRL in patients with persistent liver enzyme elevation was significantly higher than in those with normal liver function. PMID- 15660174 TI - Characteristics of primary osteomyelitis among children in a medical center in Taipei, 1984-2002. AB - BACKGROUND AND PURPOSE: The presentation and sequelae of osteomyelitis are variable. This study evaluated the clinical manifestations and outcome of osteomyelitis in children in different age groups, and in different periods before and after the implementation of National Health Insurance (NHI). METHODS: The records of pediatric patients with osteomyelitis treated at a medical center in Taipei from 1984 to 2002 were reviewed. Clinical features, pathogens, laboratory and imaging findings, treatment, and outcome were analyzed. The patients were stratified into 3 groups based on age: infants (< or = 3 months of age), young children (between 4 months and 5 years), and older children and adolescents (> 5 years). Based on the date of implementation of the NHI program, the study period was divided into 2 stages: prior to implementation, from January 1984 to February 1995; and after implementation, from March 1995 to December 2002. RESULTS: The records of 209 patients were reviewed, including 45 infants, 77 young children, and 87 older children. The most common presenting findings were local tenderness (79%), local swelling (72%), and fever (57%). The lower limbs were the most commonly involved sites (65%). Staphylococcus aureus (34%), Mycobacterium tuberculosis (10%), Salmonella species (7%), and Pseudomonas aeruginosa (6%) were the most frequently isolated pathogens. At the time of diagnosis, abnormalities were found on radiographs in 83% of patients and on radionuclide bone scans in 86%. Surgical intervention with local debridement and curettage was performed in 102 patients (49%). Sixty five patients (31%) developed complications, including specific bony sequelae in 26 (12%). Sepsis and septic arthritis were more common in infants (p < 0.01). A history of trauma, protracted course of osteomyelitis, and surgical intervention were more common in older children and adolescents (p < 0.01). After the implementation of NHI, a larger proportion of patients had negative cultures (p < 0.01), and the mean duration of antibiotic therapy was shorter (p = 0.01). CONCLUSIONS: The clinical characteristics of osteomyelitis associated with sepsis or septic arthritis, chronic changes, and the need for surgery may differ depending on the age of the child. S. aureus, M. tuberculosis, and salmonellae were the most common pathogens in this Taiwanese series. Implementation of NHI in the more recent decade of the study period was associated with a shorter duration of intravenous antibiotic administration. PMID- 15660175 TI - Increased frequency of genital human papillomavirus infection in human immunodeficiency virus-seropositive Taiwanese women. AB - BACKGROUND AND PURPOSE: Human papillomavirus (HPV) infection is associated with increased incidence and severity of HPV-related cervical dysplasia and cervical cancer in women with human immunodeficiency virus (HIV) infection. This study examined the incidence of genital HPV infection in HIV-infected Taiwanese women and its relationship with cervical neoplasia. METHODS: This hospital-based, case control study enrolled 31 consecutive HIV-seropositive women and 124 age-matched women who were free from HIV infection. Polymerase chain reaction (PCR) was used to distinguish high-risk (types 16, 18, 31, 33, 52 and 58) and low-risk HPV (types 6 and 11). The occurrence of genital HPV infection was compared between women with and without HIV infection. In addition, CD4 lymphocyte counts were determined by flow cytometry and Papanicolaou test was done in women with HIV infection. RESULTS: HPV and Papanicolaou test were done soon after the diagnosis of HIV infection. HIV seropositive women had a significantly greater high-risk HPV infection rate (48.4%; 15/31) than women without HIV infection (20.2%; 25/124; odds ratio, 3.71; p = 0.001). However, the prevalence of cervical intraepithelial neoplasia was similar between women with and without HIV infection. The CD4 lymphocyte counts in HIV-seropositive women were similar between those with and without genital HPV infection. CONCLUSIONS: The risk of genital HPV infection was significantly increased in HIV-infected women. Due to the association between high-risk HPV infection and the development of cervical dysplasia and cervical cancer, regular follow-up of Papanicolaou test is necessary in these women. PMID- 15660176 TI - Cryptococcus neoformans peritonitis in two patients with liver cirrhosis. AB - Cryptococcus neoformans is an important pathogen in immunocompromised patients. We report 2 cases of spontaneous C. neoformans peritonitis in patients with liver cirrhosis, a condition not previously reported in Taiwan. Patient 1, a 59-year old man with alcoholic liver cirrhosis, had primary C. neoformans peritonitis with fungemia. The patient recovered completely after prolonged fluconazole therapy without relapse. Patient 2, a 51-year-old woman with liver cirrhosis due to Budd-Chiari syndrome, had C. neoformans isolated from ascites, cerebrospinal fluid, and blood culture. In spite of adequate antifungal treatment, the patient died of fulminant sepsis. Information about the interaction and relation between liver cirrhosis and cryptococcal peritonitis is rare in the literature. The experience of these cases may help facilitate the diagnosis and treatment of cryptococcal peritonitis. PMID- 15660177 TI - Diabetic mastopathy in type II diabetes mellitus. AB - Diabetic mastopathy can mimic cancer. We report 2 cases of diabetic mastopathy in patients with long-standing type II diabetes. One was insulin-dependent, and the other had never been treated with insulin. These 2 patients had classical acoustical shadow on ultrasonograms. Breast core biopsies showed constellations of morphological features resembling diabetic mastopathy, including sclerotic changes of the fibrous stroma with keloid-like collagen fibers, few epithelioid fibroblasts, perivascular and interlobular mononuclear cell infiltrates, and focal atrophic changes of the ductal-lobular units. Both patients were free of malignancy at 3 and 4 years of follow-up, respectively. There are limited data on diabetic mastopathy in insulin-naive type II diabetes mellitus patients. Better awareness of this entity and its sonographic features may allow more patients to be spared from excisional biopsy. PMID- 15660178 TI - Endogenous endophthalmitis with subretinal abscess after dental procedures. AB - Endogenous endophthalmitis with subretinal abscess is an extremely uncommon complication following dental procedures. An unsuspected source of infection or misdiagnosis of the condition as iridocyclitis may result in inadequate or delayed treatment leading to severe visual impairment. A high index of suspicion is required for accurate diagnosis and prompt treatment. A 49-year-old man was found to have endophthalmitis in his right eye after treatment of gingival abscess. Ophthalmoscopic examination revealed multiple septic emboli and a 4-disc diameter sized subretinal abscess in his retina. No other infectious origin could be found on systemic work-up. Cultures from blood, urine, aqueous and vitreous were all negative. Pars plana vitrectomy was performed 8 days after the initial presentation because of deterioration of visual acuity to finger counting despite intravitreal and systemic antibiotics administration. Visual acuity recovered to 20/25 one month postoperatively and the retina remained attached during follow-up for 1 year. Ophthalmologists should be aware of the possibility of transient septicemia-induced endophthalmitis and subretinal abscess after dental procedures. Prompt intervention with systemic and intravitreal antibiotics combined with pars plana vitrectomy can achieve successful visual and anatomic outcome. PMID- 15660179 TI - Leptospirosis presenting with fever and pulmonary hemorrhage. AB - Reports of leptospirosis have recently been increasing in Taiwan. We report a case of leptospirosis with the unusual initial manifestation of pulmonary hemorrhage. The patient presented with cough for 1 week and was admitted. After admission, fever, hemoptysis and severe dyspnea developed suddenly. Chest radiograph showed bilateral diffuse pulmonary infiltrates and he was transferred to the emergency room of our hospital. Oxygen saturation was 86% under room air and respiratory rate was 30 per minute. After admission to the thoracic ward on the third morning, parenteral penicillin and trimethoprim-sulfamethoxazole were given empirically, and a dramatic recovery ensued. Microscopic agglutination test showed an increased titer of 1:6400 against Leptospira interrogans serogroup shermani on the fourth day of hospitalization. Neither jaundice nor renal insufficiency occurred during treatment. Pulmonary hemorrhage may be an under recognized manifestation of leptospirosis in Taiwanese patients. Leptospirosis should be taken into consideration in the differential diagnosis of pulmonary hemorrhage. Early treatment can lead to cure with reduced morbidity. PMID- 15660180 TI - Retrograde intramedullary nailing for nonunions of supracondylar femur fracture of osteoporotic bones. AB - Nonunion of supracondylar femur fracture remains a challenging problem because of limited treatment options. The situation is more complex when it occurs in elderly patients with osteoporotic bones. We report the treatment of 3 elderly patients with supracondylar femur fracture nonunion after open reduction and internal fixation with various plate-screw internal fixation systems. Two of these patients had traumatic fractures and the third had a periprosthetic fracture after primary total knee arthroplasty (TKA). After revision surgeries using retrograde nailing techniques, all fractures united eventually. When combined with indirect reduction, these techniques provide superior biomechanical properties and reduce the need for soft tissue dissection. PMID- 15660181 TI - Effects of continuous intravenous epoprostenol therapy on advanced primary pulmonary hypertension in Taiwanese patients. AB - Primary pulmonary hypertension (PPH) is a progressive disease for which there is no cure. Continuous intravenous infusion of epoprostenol in selected patients with advanced PPH improves symptoms and survival, but the long-term impact has not been reported in Taiwanese patients. Four patients with advanced PPH treated with epoprostenol therapy were enrolled in this study from July 2000 to September 2001. The basic hemodynamic status, cardiothoracic ratio, 6-minute-walk test results, maximum oxygen consumption, New York Heart Association functional class and survival were re-evaluated before and after 12 months of epoprostenol treatment. One of the patients died after 10 months of epoprostenol therapy due to heart failure, and the other 3 were still alive after a mean of 28 months. The surviving patients had pronounced symptomatic, hemodynamic, functional and survival benefit from epoprostenol therapy. Our results suggest that chronic infusion of epoprostenol can improve functional status and survival of Taiwanese patients with advanced PPH disease. PMID- 15660182 TI - Severity of pelvic organ prolapse associated with measurements of pelvic floor function. AB - This study tested the hypothesis that clinical measurements of the superficial perineum and of pelvic floor muscle (PFM) function correlate with the severity of pelvic organ prolapse. This retrospective cross-sectional study assessed 1037 women in an academic urogynecologic practice. Greatest descent of prolapse, by the Pelvic Organ Prolapse Quantification system, was correlated with two assessments of levator function--the Oxford grading scale and levator hiatus (LH) size measured by digital examination. Correlations were calculated using Pearson's correlation for continuous variables and Kendall's tau-b. Severity of prolapse correlated moderately with genital hiatus (GH) (r = 0.5, p<0.0001) and with LH (transverse r = 0.4, p < 0.0001; longitudinal r = 0.5, p < 0.0001), but weakly with the Oxford grading scale (r = -0.16, p < 0.0001). LH correlated with GH (r = 0.5, p < 0.0001) but not with perineal body (r = 0.06, p = 0.06). Both GH and LH size are associated with the severity of prolapse. LH size correlates more strongly to prolapse severity than assessment of PFM function by the Oxford grading scale. PMID- 15660183 TI - Determinants of the length of episiotomy or spontaneous posterior perineal lacerations during vaginal birth. AB - The objective of this study was to measure the length of episiotomy or spontaneous posterior perineal laceration and their relationship to perineal measurements and obstetric variables. The length of the perineum and genital hiatus and vertical length of episiotomy or posterior perineal tears were measured in 114 consecutive parturients with spontaneous singleton term deliveries. Seventy-four (65%) women underwent episiotomy while 40 (35%) sustained spontaneous posterior tears. Perineal or genital hiatus length was significantly correlated to episiotomy (r=0.34, p=0.003) or laceration (r=0.37, p=0.02) length, respectively. This association was significant (p=0.001) in a generalized linear model with duration of second stage of labor (p=0.005), degree of tear (p=0), and parity (p=0). Perineal length was significantly related to maternal age (p=0.036) and weight (p=0.037) and hiatal length (p=0). Short perineum and genital hiatus, long second stage of labor, and low parity are associated with longer posterior perineal injury. PMID- 15660184 TI - Paula method of circular muscle exercises for urinary stress incontinence--a clinical trial. AB - The aim of this study was to determine the efficacy of the Paula method of circular muscle training in the management of stress incontinence (SI). The theory behind this method states that activity of distant sphincters affects other muscles. In a pilot study, 59 women, mainly hospital employees, were randomly assigned to participate in exercises according to the Paula method or pelvic floor training. Efficacy was measured by reports of incontinence, quality of life (I-QOL), pad test, and pelvic floor muscle strength (assessed by perineometer and digital examination). Both the Paula exercises and pelvic floor training produced significant changes in urinary leakage compared to baseline as measured by the pad test [mean decrease of 5.4 g (p=0.002) and 9.5 g (p=0.003), respectively]. Women randomized to the Paula method reported improvement in I-QOL scores. The Paula method was found to be efficacious for SI in a population of Israeli women. Larger community-based studies will be required to confirm these results and enable evaluation of between-group differences. PMID- 15660185 TI - Evidence of the hexagonal columnar liquid-crystal phase of hard colloidal platelets by high-resolution SAXS. AB - We report Small-Angle X-ray Scattering (SAXS) measurements of the columnar phase of hard colloidal gibbsite platelets. We have been able to create large oriented domains of the columnar phase both perpendicular and parallel to the sample wall, varying the volume fraction of platelets and adding non-adsorbing polymer to the dispersion. In conjunction with the increased resolution of the SAXS setup, this allowed a detailed analysis of the columnar phase, providing unambiguous evidence for the hexagonal nature of the phase. PMID- 15660186 TI - Electrically induced microflows probed by fluorescence correlation spectroscopy. AB - We report on the experimental characterisation of electrically induced flows at the micrometer scale through Fluorescence Correlation Spectroscopy (FCS) measurements. We stress the potential of FCS as a useful characterisation technique in microfluidics devices for transport properties cartography. The experimental results obtained in a model situation are in agreement with previous calculations (F. Nadal, F. Argoul, P. Kestener, B. Pouligny, C. Ybert, A. Ajdari, Eur. Phys. J. E 9, 387 (2002)) predicting the structure and electric-field dependency of the induced flow. Additionally, the present study evidences a complex behaviour of the probe nanobeads under electric field whose precise understanding might prove relevant for situations where nano-objects interact with an external electric field. PMID- 15660187 TI - Partial breast irradiation after breast-conserving surgery. AB - BACKGROUND: The standard technique of postoperative radiotherapy after breast conserving surgery is percutaneous irradiation of the entire breast to a total dose of 45-50 Gy which is usually followed by a tumor bed boost. Since the majority of local recurrences in selected patients occur close to the former tumor bed, the question arises whether a sole tumor bed irradiation might be a therapeutic alternative to total breast irradiation. METHODS: A systematic review of relevant literature concerning partial breast irradiation (PBI) up to November 2004 was undertaken. Studies of any design were included for comparison and discussion. RESULTS: Nine unique brachytherapy studies using the multi-catheter technique, one the balloon technique (MammoSite), and eight particular intraoperative radiotherapy (IORT) trials were located of which only one was a randomized trial. Only minor postoperative complications were reported. Preliminary results are similar in terms of local tumor control, disease-free and overall survival. However, the current evidence base of IORT studies is poor. CONCLUSION: Despite controversies regarding PBI after breast-conserving surgery, results of phase I-II trials suggest that sole tumor bed irradiation might be an appropriate therapeutic alternative for selected breast cancer patients. However, more experience and data from ongoing phase III trials are required to define these new methods to be an appropriate treatment option. Therefore, total breast irradiation still remains the standard irradiation modality even in the treatment of early breast cancer, and PBI should be considered investigational. PMID- 15660188 TI - Waiting for radiotherapy a national call for ethical discourse on waiting lists in radiotherapy: findings from a preliminary survey. AB - BACKGROUND AND PURPOSE: Growing incidence of cancer and the demographic changes of the society can cause severe shortage in radiotherapy services. This can lead to the use of waiting lists, which can serve as an indicator model for shortage of medical service. To investigate whether and how waiting lists are used in Germany, a survey was undertaken among German radio-oncologists. Their opinion concerning the use of waiting lists and their attitude toward the involved ethical problems were the central topics of this study. MATERIAL AND METHODS: A questionnaire was developed which consisted of 18 items: four items covered general aspects, four were about the management of waiting lists, seven concerned problems of waiting lists, and three were about the future aspects. From all items, at least ten touched ethical aspects such as patient information or the doctor-patient relationship. 19 consecutive radio-oncologists answered the questionnaire via telephone. The main interest of this study was to generate further hypotheses for future research. RESULTS: From all doctors (n = 19), most did not use waiting lists (n = 12) in their practice, whereas the remaining did use them (n = 7). Limited resources were seen as the central underlying problem (n = 16). Two guiding principles were most often quoted for managing waiting lists: patients with complaints such as pain before those without (n = 18) and curative before palliative radiotherapy (n = 15). Concerning the information given to patients about the use of waiting lists, some doctors voted for detailed information (n = 7), which covers negative side effects of waiting lists such as tumor progression during waiting time, others voted against (n = 11). There was a profound disagreement on the question of whether curative and palliative radiotherapy should be differently treated in the context of waiting lists. 70% of the group, who used waiting lists, could give a moral justification for waiting lists, whereas only 28% of the group, who did not use them, could offer a justification. CONCLUSION: The survey showed that doctors were most concerned about the ethical aspects of waiting lists. Open questions are: (1) What is the best principle for a fair distribution of limited radiotherapy places? (2) How should patients be informed about waiting lists? (3) What moral justifications can be given for waiting lists? These issues must therefore become topics of future research and national dialogue, because the answers to these questions are necessary for the clinical use of waiting lists in radiotherapy. PMID- 15660189 TI - Phase I/II trial of simultaneous whole-brain irradiation and dose-escalating topotecan for brain metastases. AB - BACKGROUND AND PURPOSE: Topotecan penetrates the blood-brain barrier and sensitizes tumor cells against radiation. A phase I/II dose-escalating trial of repetitive daily i. v. topotecan application simultaneously with whole-brain irradiation (WBRT) was conducted to estimate toxicity, maximum tolerated dose and survival in patients with inoperable brain metastases. PATIENTS AND METHODS: In 47 patients suffering from previously untreated brain metastases, topotecan was applied on a daily i. v. schedule simultaneously with WBRT (36 Gy/3-Gy fractions). The infusion schedule started at the beginning of WBRT and was discontinued during weekends. Each infusion was completed within 1-2 h before irradiation. In a dose-finding study, topotecan was escalated from 5 x 0.5 mg/m(2), 8 x 0.5 mg/m(2), 12 x 0.5 mg/m(2) to 12 x 0.6 mg/m(2). RESULTS: Altogether, 38/47 patients (81%) completed the prescribed schedule. Leukopenia and thrombocytopenia were dose-limiting. Grade 3/4 hematologic toxicity occurred in 5/32 chemonaive patients (16%) and 7/15 patients (47%) with previous chemotherapy. At 12 x 0.6 mg/m(2), 2/4 patients experienced grade 4 leukopenia/thrombopenia. Nonhematologic toxicities were generally mild to moderate and unrelated to topotecan. Response evaluation was possible in 26/47 patients, overall response rate was 58% (CR [complete remission] 5/26, PR [partial remission] 10/26, NC [no change] 8/26). Median survival amounted to 5.1 months. In 15/42 patients (36%), brain metastases were the dominant cause of death. CONCLUSION: For a daily topotecan schedule simultaneous to WBRT, the maximum tolerated dose is 12 x 0.5 mg/m(2) in chemonaive patients. For chemo pretreated patients, daily doses should be reduced to 0.4 mg/m(2). A phase III trial has now been started to find out whether WBRT + topotecan increases survival compared to WBRT alone. PMID- 15660190 TI - Multimodality treatment including postoperative radiation and concurrent chemotherapy with weekly docetaxel is feasible and effective in patients with oral and oropharyngeal cancer. AB - BACKGROUND: To examine the feasibility and efficacy of weekly docetaxel with concurrent radiation as postoperative treatment in a multimodality approach to oral and oropharyngeal cancer. PATIENTS AND METHODS: 94 patients (Table 1) with primary resectable squamous cell carcinoma of the oral cavity and oropharynx (UICC stage I 14%, II 15%, III 18%, IV 53%; Table 2) were treated with a multimodality therapy program consisting of neoadjuvant intra-arterial high-dose chemotherapy (cisplatin 150 mg/m(2) with parallel systemic sodium thiosulfate 9 g/m(2) for neutralization), followed by surgery of the primary and neck, and postoperative concurrent radiation and chemotherapy with weekly docetaxel (20-30 mg/m(2); Table 3). Chronic toxicities were followed over a period of 5 years. RESULTS: At a median follow-up of 4 years, the 5-year survival rate for all 94 patients was 80%, and disease-free survival was 73% (Figures 1 and 2). Among patients with advanced disease (stage III and IV), survival was 83 and 59%, respectively (Figure 4). Grade 3 and 4 mucositis was the main acute toxicity necessitating supportive care. Long-term toxicity appears to be moderate (Table 4). The maximum tolerated dose of weekly docetaxel was 25 mg/m(2). CONCLUSIONS: Concurrent radiation and chemotherapy with weekly docetaxel is a feasible postoperative treatment in a multimodality approach to oral and oropharyngeal cancer, resulting in high overall and disease-free survival. This approach warrants further evaluation in prospective randomized trials. PMID- 15660191 TI - Regional hyperthermia in conjunction with definitive radiotherapy against recurrent or locally advanced prostate cancer T3 pN0 M0. AB - BACKGROUND AND PURPOSE: Since long-term results of the standard treatment of locally advanced or recurrent prostatic carcinoma are unsatisfactory, the role for additional regional hyperthermia was evaluated in a phase I/II study. PATIENTS AND METHODS: From 08/1996 to 03/2000, 22 patients were treated by a standard irradiation regimen (68.4 Gy) in combination with regional hyperthermia (weekly, five to six times), and five of 22 patients received short-term (neoadjuvant) hormonal treatment. Of these, 15 patients had primary prostatic carcinoma T3 pN0 M0 and seven a histologically confirmed local recurrence after radical prostatectomy. Feasibility of hyperthermia, and acute/late toxicity as well as long-term follow-up (prostate- specific antigen [PSA] control, overall survival) were analyzed. Clinical endpoints were correlated with thermal parameters. RESULTS: Mean maximum temperatures along the urethra of 41.4 degrees C (41.0 degrees C for the recurrences), and mean T(90) values of 40.7 degrees C could be achieved. Severe acute toxicity of grade 3 occurred at the rectum in three, at the urethra in four, at the intestine in one, and a burn induced by hyperthermia in one of 22 patients. Late toxicity was only observed rectally in one patient (grade 3) and at the urethra in two patients (grade 2). There was no correlation between thermal parameters and any toxicity. The survival curves showed a PSA control for primary prostatic carcinoma > 50% after 6 years, but no long-term PSA control for the recurrences. Overall survival after 6 years was 95% for primary carcinoma, and 60% for the recurrences. There was a clear correlation between higher temperatures or thermal doses with long-term PSA control. CONCLUSION: Regional hyperthermia might be a low-toxicity approach to increase PSA control of common treatment schedules. Further evaluation, in particular employing improved hyperthermia technology, is worthwhile. PMID- 15660192 TI - Radiobiological investigation of dose-rate effects in intensity-modulated radiation therapy. AB - BACKGROUND AND PURPOSE: Intensity-modulated radiation therapy (IMRT) has proven extraordinary capability in physical terms such as target conformity, dose escalation in the target volume, and sparing of neighboring organs at risk. The radiobiological consequences of the protracted dose delivery for cell survival and cell cycle progression are still unclear and shall be examined in this study. MATERIAL AND METHODS: Human lymphoblasts (TK6) and human melanoma cells (MeWo) were irradiated with protocols of increasing dose protraction. In addition, a new biophysical phantom was developed and used to transfer clinical IMRT plans to experimental cell irradiation. Clonogenic cell survival and cell cycle analysis were performed after various irradiation experiments. RESULTS: In a first series of experiments, melanoma cells showed a highly significant increase of survival of 6.0% after protracted dose delivery of 2 Gy compared to conventional fast application with the same dose. Lymphoblastoid cells also showed a significant increase of survival of 2.2%. Experiments with patient plans in the phantom confirmed the trend of increased cell survival after protracted dose delivery. Cells were irradiated at 13 points in four different IMRT plans. In comparison to irradiation with application of the same dose in a classic four-field box, a significantly increased survival of 5.1% (mean value) was determined. CONCLUSION: Even at fraction times of 15-30 min the protracted dose delivery increases the survival rates in cell culture. The altered survival rates indicate the importance of the dose rate in the effectivity of IMRT. Besides physical parameters the consideration of biological factors might contribute to the optimization of IMRT in the future. PMID- 15660193 TI - Donut-shaped high-dose configuration for proton beam radiation therapy. AB - BACKGROUND: The authors report on the conception and first clinical application of a donut-shaped high-dose configuration for proton therapy (PT). This approach allows one to intensify target volume dose coverage for targets encompassing a critical, dose-limiting structure--like here, the cauda equina--, whilst delivering minimal dose to other healthy structures surrounding the target, thereby reducing the integral dose. METHODS AND RESULTS: Intensity-modulated PT methods (IMPT) for spot scanning were applied to create and deliver a donut shaped high-dose configuration with protons, allowing treating > 75% of the target with at least 95% of the prescribed dose of 72.8 CGE, whilst restricting dose to the cauda equina to 60-65 CGE. Integral dose was lower by a factor of 3.3 as compared to intensity-modulated radiotherapy with photons (IMXT). CONCLUSION: IMPT and spot scanning technology allow a potentially clinically useful approach which is also applicable to spare other critical structures passing through a target volume, including spinal cord, optic nerves, chiasm, brain stem, or urethra. PMID- 15660194 TI - J. Jassem on A. Mayer, et al. Primary radiotherapy of stage IIA/B-IIIB cervical carcinoma. A comparison of continuous versus sequential regimens. Strahlenther Onkol 2004;180:209-15 (No. 4). PMID- 15660196 TI - Sibutramine in the treatment of obesity in type 2 diabetic patients and in nondiabetic subjects. AB - Obesity is considered a chronic disease requiring treatment. The effect of sibutramine combined with hypocaloric diet and exercise on body weight, body fat mass, lipids, glycemic control, insulin secretion and insulin resistance was evaluated in a randomized, controlled, open-label study. A total of 44 obese type 2 diabetic patients (aged 45.2+/-5.2 years, BMI 33.62+/-2.2 kg/m(2)) and 49 obese nondiabetic subjects (aged 41.9+/-5.7 years, BMI 34.3+/-2.6 kg/m(2)) were treated with sibutramine for 3 months. Moreover, 39 age-matched obese type 2 diabetic patients and 41 obese nondiabetic subjects only on hypocaloric diet and exercise served as control groups. Insulin secretion was estimated during intravenous glucose tolerance test; insulin resistance was assessed by the HOMA index. There was a significant reduction in body weight in both sibutramine-treated diabetic patients (7.1%) and nondiabetic subjects (9.1%), accompanied by a significant reduction in body fat mass. HbA1c decreased significantly in the diabetic patients after sibutramine treatment. There was a significant improvement of lipid parameters in the two groups. Insulin resistance decreased by 21.9% in the sibutramine-treated diabetic patients and by 38.5% in the nondiabetic group. Weight loss was accompanied by an increase of 43.8% in first phase insulin secretion in the sibutramine-treated diabetic group; in the treated nondiabetic subjects there was a decrease in first and second phase insulin secretion and the area under the curve for total insulin secretion. In conclusion, sibutramine leads to a significant reduction in body weight, body fat mass and waist and hip circumferences; it improves insulin sensitivity, insulin secretion, glycaemic control and lipid parameters in both diabetic and nondiabetic obese subjects. PMID- 15660195 TI - Scanning for MODY5 gene mutations in Chinese early onset or multiple affected diabetes pedigrees. AB - Mutation of HNF-1beta gene has been reported in early onset diabetes or MODY families and this gene has been defined as MODY5 gene. The aim of our study was to examine whether HNF-1beta mutation contribute to early onset or multiple affected diabetes pedigrees in Chinese. Molecular scanning of HNF-1beta gene promoter region, nine exons and flanking introns was performed in 154 unrelated probands from early onset and multiple affected diabetes Chinese pedigrees. The family members of probands with mutations or variants and 58 nondiabetics were also examined. Clinical examinations of renal morphology, renal function and beta cell function were performed in the HNF-1beta gene mutation carriers and family members. Mutation of HNF-1beta gene causing the substitution S36F was found in two subjects of an early onset diabetic family. One carrier has early onset diabetes, renal function impairment and renal cyst, while the other has impaired glucose tolerance only. This is the first case of MODY5 gene mutation diabetes found in the Chinese. Three HNF-1beta variants were identified and no significant differences in allele frequencies for these variants were detected between the nondiabetic and diabetic groups. Nucleotide 66 of intron 8 of HNF-1beta gene was G in the Chinese population rather than A as noted in the GenBank sequence. These results suggest that HNF-1beta gene mutations may be associated with nondiabetic renal dysfunction and diabetes in Chinese, but they are responsible for only a small percentage of early onset or multiple affected diabetes pedigrees including MODY. PMID- 15660197 TI - Is selective screening for gestational diabetes mellitus worthwhile everywhere? AB - We assessed if selective screening for gestational diabetes mellitus (GDM) as recommended by the Fourth Workshop on GDM is worthwhile in our centre. Detection is performed using universal screening in three pregnancy periods using the tests recommended by the first three Workshops. We have analysed the prevalence of low risk characteristics for GDM in the 917 women delivering in the centre in 1992 and in the whole cohort of 1635 women with GDM delivering between 1986 and 1998. The rate of women with all low risk characteristics was 7.0% among the general pregnant population and 1.3% in the cohort of women with GDM (p<0.001). We conclude that in our population, selective screening of GDM is reliable in identifying women at low risk of GDM, but since only a negligible subset of the pregnant population would not need to be screened, adherence to these guidelines would make the screening policy unnecessarily complicated. PMID- 15660198 TI - Advanced glycation endproducts in peripheral nerve in type 2 diabetes with neuropathy. AB - Advanced glycation endproducts (AGE) accumulate over proteins as a consequence of diabetic hyperglycemia, and thus contribute to the pathogenesis of diabetic complications. To improve the understanding of the pathology of diabetic neuropathy, AGE accumulation was analyzed in sural and/or femoral nerves obtained under spinal anesthesia from 8 type 2 diabetic patients with both distal symmetrical polyneuropathy and proximal neuropathy. Pronounced AGE immunoreactivity was detected on axons and myelin sheaths in 90% of diabetic peripheral nerves but not in the control specimen. The intensity of axonal AGE immunopositivity significantly correlated with the severity of morphological alterations (p<0.005). AGE localization, demonstrated by immunohistochemical methods, was also present in the endoneurium, perineurium and microvessels. Morphometric analysis of the diabetic peripheral nerve showed perineurial thickening (diabetic vs. control, 15.5+/-4.9 vs. 6.6+/-2.1 microm, p<0.001), narrowing of the microvessel lumina (66.6+/-50.5 vs. 579.5+/-38.4 x10(3) microm(2), p<0.001) and significant reduction in the number of preserved axons (3.6+/-3 vs. 8.9+/-2.3 per 10(5) microm(2) per area, p<0.037). The sera of diabetic patients contained epitope(s) of AGE structure and soluble immune complexes containing AGE moiety. In conclusion, to the best of our knowledge, this is the first study providing evidence for excessive AGE formation on peripheral nerve components, primarily axons, and a significantly higher level of circulating AGE-immune complexes in patients with both distal diabetic polyneuropathy and proximal neuropathy. Humoral immune mechanisms, including the production of anti-AGE autoantibody, may potentially be involved in the development of structural abnormalities described in this report. PMID- 15660199 TI - T-cell receptor repertoire and function in umbilical cord blood lymphocytes from newborns of type 1 diabetic mothers. AB - Recent observations show that the development of the human fetal immune system is susceptible to conditions of the maternal immune system and vice versa. To investigate the impact of type 1 diabetes mellitus in pregnant women on the maturation of the immune system of their newborn infants, the umbilical cord blood (UCB) T-cell repertoire at birth was analysed for clonal or oligoclonal expansions and TCRBV gene usage. Quantitative PCR using TCRBV family-specific probes and the spectratyping method were used. The extent of oligoclonal expansions observed in cord blood was markedly lower than in peripheral blood of the diabetic mothers and healthy adults. Functional analysis revealed that UCB T cells from newborns of both type 1 diabetic and gestational diabetic mothers are mature and can be readily stimulated by superantigens and phytohemagglutinin in vitro. No significant differences of the clonal contingent and the TCRBV usage were found in newborns of type 1 diabetic mothers when compared to newborns of gestational diabetic mothers, independent from the presence or absence of islet autoantigens. Our findings indicate that the immune system of type 1 diabetic mothers has no major effects on the TCR repertoire of the newborn infants. In that respect, no evidence was found for superantigen-activated T cells, nor was chronic hyperglycaemia per se found to alter either T-cell repertoire or functional activity. PMID- 15660200 TI - Effect of the pyridoindole antioxidant stobadine on sodium handling of renal Na,K ATPase in rats with streptozotocin-induced diabetes. AB - Overload of reactive oxygen species during diabetes is known to impair cellular homeostasis and to promote deterioration of membrane function in the organism. The aim of the present study was to examine the effect of dietary supplementation with the pyridoindole atioxidant stobadine on functional properties of the renal Na, K-ATPase in diabetic rats. After 16 weeks of streptozotocin-induced diabetes (single intravenous dose of streptozotocin; 55 mg/kg), a significant inhibition (by 35%-42%) of the enzyme was observed throughout the range of NaCl 2-100 mmol/l, probably as an event of altered functional properties of Na,K-ATPase, suggested by the 42% decrease of the V(max) value. Administration of 0.05% (w/w) stobadine in the diet dramatically improved the function of renal Na,K-ATPase in diabetic rats with regard to sodium handling, as suggested by significant stimulation (by 104%-77% in accordance with increasing concentration of NaCl) of the enzyme over the whole NaCl concentration range investigated. This stimulatory effect was accompanied by an increase of V(max) value to the level of nondiabetic rats on standard diet. In conclusion, stobadine was found to antagonise the negative effects of diabetes on the renal Na,K-ATPase, preserving its normal function in regulation of intracellular homeostasis of Na(+) and K(+) ions. PMID- 15660201 TI - A pilot study of mitochondrial DNA point mutation A3243G in a sample of Croatian patients having type 2 diabetes mellitus associated with maternal inheritance. AB - In this work, patients having type 2 diabetes mellitus and diabetic mothers were tested for the presence of mitochondrial DNA point mutation A3243G. This mutation is associated with the MELAS syndrome (mitochondrial myopathy, encephalopathy, lactic acidosis and stroke-like episodes), diabetes and deafness. Twenty-two diabetic persons were screened. DNA was isolated from peripheral blood lymphocytes and from swabs of oral mucosa. The mitochondrial DNA point mutation A3243G was detected using PCR-RFLP test. The mutation was detected in oral mucosal DNA of two patients (but not from lymphocyte DNA). One patient was a man with hearing and visual impairments and proteinuria; the other was a woman having proteinuria but no hearing impairment. The mutation was not detectable in oral mucosal DNA from the control persons: 20 diabetic patients having diabetic fathers and 22 healthy, nondiabetic volunteers. The incidence of mitochondrial DNA point mutation A3243G in this study of Croatian diabetic patients is in line with data in the literature. PMID- 15660202 TI - Functional identification and monitoring of individual alpha and beta cells in cultured mouse islets of Langerhans. AB - The aim of the present study was to evaluate, by use of fluorescence microscopy and immunofluorescence stainings, the use of a fluorescent membrane potential sensitive probe as a means to identify and monitor changes in membrane potential of individual cell types in whole islets of Langerhans over time. Our work supports the use of the fluorescent probe bis-(1,3 dibutylbarbituric acid) trimethine oxonol (diBAC(4)(3)), in identification of single alpha and beta cells in the periphery of mouse pancreatic islets cultured on extracellular matrix. At a low extracellular glucose concentration (3 mM), heterogeneous staining of the islets was observed. Approximately 97% of the peripheral cells that stained brightly with diBAC(4)(3) were glucagon positive. Additional diBAC(4)(3) studies, demonstrated that an increase in glucose concentration from 3 to 10 mM is paralleled by repolarization of alpha cells and depolarization of beta cells. This suggests that reciprocity of glucagon and insulin release upon glucose stimulation is coupled to divergent changes in membrane potential of these cell types and supports the use of diBAC(4)(3) as a means to detect changes in secretion in both cell types. PMID- 15660203 TI - Re-institution of good metabolic control in diabetic rats and activation of caspase-3 and nuclear transcriptional factor (NF-kappaB) in the retina. AB - Hyperglycemia is one of the major underlying factors in the development of retinopathy in diabetes. Retinal microvascular cells undergo accelerated apoptosis before other histopathological changes are detectable in diabetes. We examined the effect of re-institution of good metabolic control (GC) on the activation of retinal apoptosis executor enzyme, caspase-3, and nuclear transcriptional factor NF-kB. In streptozotocin diabetic rats, two or six months of poor metabolic control (PC) with glycated hemoglobin >11.0% was followed by seven additional months of GC (glycated hemoglobin <5.5%). Caspase-3 activity in retina was measured by the cleavage of its substrate, the expression of active 17 kD subunit, and cleavage of poly(ADP ribosyl) polymerase. NF-kB activation was determined by electrophoretic shift assay and by western blots for P65 subunit. Caspase-3 activity in diabetic rats kept in PC for 13 months was 175% that in normal rats. Re-institution of GC after two months of PC partially normalized the hyperglycemia-induced activation of caspase-3 (to 140% of normal values) while re institution of GC after six months of PC had no significant effect on the activation of caspase-3 NF-kB activity was 2.5-fold higher in diabetic rats kept in PC than in normal rats. Re-institution of GC after 2 months of PC partially reversed this increase (X-fold over normal), but GC after 6 months of PC had no effect. Initiation of GC soon after induction of diabetes in rats prevented activation of retinal caspase-3 and NF-kB. These results suggest that the process of activation of apoptosis execution enzyme and NF-kB in retina that starts before appearance of histopathological changes is not easily reversed by re institution of GC. Characterization of the abnormalities responsible for the resistance of retinopathy to halt after re-institution of GC will help identify potential therapies for inhibition of progression of diabetic retinopathy. PMID- 15660205 TI - Unfolded protein response followed by induction of cell death in cultured tobacco cells treated with tunicamycin. AB - When correct folding of protein in the endoplasmic reticulum (ER) is prevented, cells respond to overcome the accumulation of unfolded proteins. This cellular response, which includes the induction of ER chaperones, is called an unfolded protein response (UPR). Although a link between the UPR and apoptosis has been reported in mammalian cells, little is known about this mechanism in plant cells. Asparagine (N)-linked glycosylation of proteins is critical for protein folding in the ER; and tunicamycin, a potent inhibitor of N-linked glycosylation, induces UPR. Growth arrest was observed in cultured tobacco cells treated with tunicamycin. Cell death and induction of Hsr203J, a marker for programmed cell death, were observed in the 24-h period after addition of tunicamycin, following UPR that started within 2 h. These results indicate a strong link between UPR and programmed cell death in plant cells. PMID- 15660206 TI - Gravitropic moss cells default to spiral growth on the clinostat and in microgravity during spaceflight. AB - In addition to shoots and roots, the gravity (g)-vector orients the growth of specialized cells such as the apical cell of dark-grown moss protonemata. Each apical cell of the moss Ceratodon purpureus senses the g-vector and adjusts polar growth accordingly producing entire cultures of upright protonemata (negative gravitropism). The effect of withdrawing a constant gravity stimulus on moss growth was studied on two NASA Space Shuttle (STS) missions as well as during clinostat rotation on earth. Cultures grown in microgravity (spaceflight) on the STS-87 mission exhibited two successive phases of non-random growth and patterning, a radial outgrowth followed by the formation of net clockwise spiral growth. Also, cultures pre-aligned by unilateral light developed clockwise hooks during the subsequent dark period. The second spaceflight experiment flew on STS 107 which disintegrated during its descent on 1 February 2003. However, most of the moss experimental hardware was recovered on the ground, and most cultures, which had been chemically fixed during spaceflight, were retrieved. Almost all intact STS-107 cultures displayed strong spiral growth. Non-random culture growth including clockwise spiral growth was also observed after clinostat rotation. Together these data demonstrate the existence of default non-random growth patterns that develop at a population level in microgravity, a response that must normally be overridden and masked by a constant g-vector on earth. PMID- 15660208 TI - Analog number representations in mongoose lemurs (Eulemur mongoz): evidence from a search task. AB - A wealth of data demonstrating that monkeys and apes represent number have been interpreted as suggesting that sensitivity to number emerged early in primate evolution, if not before. Here we examine the numerical capacities of the mongoose lemur (Eulemur mongoz), a member of the prosimian suborder of primates that split from the common ancestor of monkeys, apes and humans approximately 47 54 million years ago. Subjects observed as an experimenter sequentially placed grapes into an opaque bucket. On half of the trials the experimenter placed a subset of the grapes into a false bottom such that they were inaccessible to the lemur. The critical question was whether lemurs would spend more time searching the bucket when food should have remained in the bucket, compared to when they had retrieved all of the food. We found that the amount of time lemurs spent searching was indicative of whether grapes should have remained in the bucket, and furthermore that lemur search time reliably differentiated numerosities that differed by a 1:2 ratio, but not those that differed by a 2:3 or 3:4 ratio. Finally, two control conditions determined that lemurs represented the number of food items, and neither the odor of the grapes, nor the amount of grape (e.g., area) in the bucket. These results suggest that mongoose lemurs have numerical representations that are modulated by Weber's Law. PMID- 15660207 TI - The inositol oxygenase gene family of Arabidopsis is involved in the biosynthesis of nucleotide sugar precursors for cell-wall matrix polysaccharides. AB - The nucleotide sugar UDP-glucuronic acid (UDP-GlcA) is the principal precursor for galacturonic acid, xylose, apiose and arabinose residues of the plant cell wall polymers. UDP-GlcA can be synthesized by two different functional pathways in Arabidopsis involving either UDP-glucose dehydrogenase or inositol oxygenase as the initial enzyme reaction to channel carbohydrates into a pool of UDP sugars used for cell-wall biosynthesis. The genes for the enzyme myo-inositol oxygenase (MIOX) were analyzed in Arabidopsis. They represent a small gene family containing four members. The transcription of all those members indicates a transient and organ-specific gene expression pattern in growing plant tissues as analyzed by RT-PCR and in promoter::GUS reporter gene lines. Two isoforms (MIOX1, MIOX2) are expressed in almost all tissues of the plant, whereas the expression of MIOX4 and MIOX5 is largely restricted to flowers, particularly maturing pollen. T-DNA insertion lines in MIOX genes were isolated; however, single knock outs show growth phenotypes similar to the wild type. The monosaccharide composition of the cell wall in these mutants is not significantly changed compared to wild type plants. However, the incorporation of 3H-inositol into wall polymers of seedlings is greatly impaired in the mutant lines (Delta)MIOX1 and (Delta)MIOX2, which are the only isoforms that are expressed in seedlings. PMID- 15660209 TI - Do responses of galliform birds vary adaptively with predator size? AB - Past studies of galliform anti-predator behavior show that they discriminate between aerial and ground predators, producing distinctive, functionally referential vocalizations to each class. Within the category of aerial predators, however, studies using overhead models, video images and observations of natural encounters with birds of prey report little evidence that galliforms discriminate between different raptor species. This pattern suggests that the aerial alarm response may be triggered by general features of objects moving in the air. To test whether these birds are also sensitive to more detailed differences between raptor species, adult chickens with young were presented with variously sized trained raptors (small, intermediate, large) under controlled conditions. In response to the small hawk, there was a decline in anti-predator aggression and in aerial alarm calling as the young grew older and less vulnerable to attack by a hawk of this size. During the same developmental period, responses to the largest hawk, which posed the smallest threat to the young at all stages, did not change; there were intermediate changes at this time in response to the middle sized hawk. Thus the anti-predator behavior of the adult birds varied in an adaptive fashion, changing as a function of both chick age and risk. We discuss these results in light of current issues concerning the cognitive mechanisms underlying alarm calling behavior in animals. PMID- 15660210 TI - Sucrose utilisation in bacteria: genetic organisation and regulation. AB - Sucrose is the most abundant disaccharide in the environment because of its origin in higher plant tissues, and many Eubacteria possess catalytic enzymes, such as the sucrose-6-phosphate hydrolases and sucrose phosphorylases, that enable them to metabolise this carbohydrate in a regulated manner. This review describes the range of gene architecture, uptake systems, catabolic activity and regulation of the sucrose-utilisation regulons that have been reported in the Eubacteria to date. Evidence is presented that, although there are many common features to these gene clusters and high conservation of the proteins involved, there has been a certain degree of gene shuffling. Phylogenetic analyses of these proteins supports the hypothesis that these clusters have been acquired through horizontal gene transfer via mobile elements and transposons, and this may have enabled the recipient bacteria to colonise sucrose-rich environmental niches. PMID- 15660211 TI - Morphological engineering of Streptomyces hygroscopicus var. geldanus: regulation of pellet morphology through manipulation of broth viscosity. AB - Actinomycetes, especially members of the genus Streptomyces, are responsible for producing the majority of known antibiotics. The production of antibiotics by filamentous organisms is often dependent on the morphology and size distribution of the pellet population within the culture. Particle interaction and subsequent pellet formation are primarily dependent on the rate of collision of particles in culture, which is in turn, a function of fluid turbulence. The microbial polysaccharide xanthan gum was used to artificially regulate the apparent viscosity (mu(a)) of S. hygroscopicus fermentation broths with the aim of controlling particle interaction, aggregation and hence pellet formation. An increase in both pellet count and biomass concentration from approximately 2,000 to 8,000 pellets ml(-1) and 0.9-2.1 g l(-1) dry weight of biomass, as well a decrease in the mean pellet volume from 0.014 to 0.004 mm(3) was observed in cultures supplemented with 3 g l(-1) xanthan gum. The addition of xanthan gum significantly alters fluid rheology by increasing the mu(a). Counter-intuitively, an increase in the mu(a) within the experimental range examined resulted in an increase in the rate of gas-liquid mass transfer. This was attributed to the predominantly diffusive nature of oxygen transfer in shake flask cultures. PMID- 15660212 TI - Influenza vaccines: recent advances in production technologies. AB - In spite of ongoing annual vaccination programs, the seasonal influenza epidemics remain a major cause of high morbidity and mortality. The currently used "inactivated" vaccines provide very short-term and highly specific humoral immunity due to the frequent antigenic variations in the influenza virion. These intra-muscularly administered vaccines also fail to induce protective mucosal immunity at the portal of viral entry and destruction of the virally infected cells by induction of cytotoxic T lymphocytes. Therefore, it is necessary to develop immunologically superior vaccines. This article highlights some of the recent developments in investigational influenza vaccines. The most notable recent developments of interest include the use of immunopotentiators, development of DNA vaccines, use of reverse genetics, and the feasibility of mammalian cell-based production processes. Presently, due to their safety and efficacy, the cold-adapted "live attenuated" vaccines are seen as viable alternatives to the "inactivated vaccines". The DNA vaccines are gaining importance due to the induction of broad-spectrum immunity. In addition, recent advances in recombinant technologies have shown the possibility of constructing pre-made libraries of vaccine strains, so that adequately preparations can be made for epidemics and pandemics. PMID- 15660213 TI - Biotransformations for the production of the chiral drug (S)-Duloxetine catalyzed by a novel isolate of Candida tropicalis. AB - A yeast strain, Candida tropicalis PBR-2, isolated from soil, is capable of carrying out the enantioselective reduction of N,N-dimethyl-3-keto-3-(2-thienyl) 1-propanamine to (S)-N,N-dimethyl-3-hydroxy-3-(2-thienyl)-1-propanamine, a key intermediate in the synthesis of the chiral drug (S)-Duloxetine. The organism produced the enantiopure (S)-alcohol with a good yield (> 80%) and almost absolute enantioselectivity, with an enantiomeric excess (ee) > 99%. Parameters of the bioreduction reaction were optimized and the optimal temperature and pH for the reduction were found to be 30 degrees C and 7.0, respectively. The optimized substrate and the resting cell concentration were 1 g/l and 250 g/l, respectively. The preparative-scale reaction using resting cells of C. tropicalis yielded the (S)-alcohol at 84-88% conversion and ee > 99%. PMID- 15660214 TI - Expression of SARS-coronavirus nucleocapsid protein in Escherichia coli and Lactococcus lactis for serodiagnosis and mucosal vaccination. AB - The nucleocapsid (N) protein of the severe acute respiratory syndrome (SARS) associated coronavirus (SARS-CoV) is an important antigen for the early diagnosis of SARS and the development of vaccines. It was expressed in Escherichia coli as a fusion with human glutathione S-transferase (hGST) and was confirmed by Western blotting analysis. This recombinant N protein (hGST-N) was purified and used to measure the SARS-CoV N-specific antibody in the sera of eight SARS patients by enzyme-linked immunosorbent assay. Specific antibody response to this purified recombinant N protein was 100% positive in the SARS patients' sera, while none of the control sera from 30 healthy people gave a positive reaction in the same assay. The SARS-CoV N protein was also expressed in Lactococcus lactis in the cytoplasm or secreted into the medium. The N-producing strain MG1363/pSECN and the purified hGST-N protein were respectively administered to mice, either orally or intranasally. Results indicated that orally delivered MG1363/pSECN induced significant N-specific IgG in the sera. In conclusion, our work provides a novel strategy to produce the SARS-CoV N protein for serodiagnosis and for L. lactis based mucosal vaccines. PMID- 15660215 TI - Bioconversion of lutein using a microbial mixture--maximizing the production of tobacco aroma compounds by manipulation of culture medium. AB - The generation of aroma compounds by carotenoid cleavage in the 9-10 position was studied, due to the importance of these compounds in the flavor and fragrance industry. The bioconversion of the carotenoid lutein to C(13) norisoprenoids utilizing a microbial mixture composed of Trichosporon asahii and Paenibacillus amylolyticus was carried out by a fermentation process. Applying an experimental design methodology, the effects of nutritional factors on the production of aroma compounds present in the tobacco profile were studied. After an assessment of the significance of each nutritional factor, the levels of the variables yielding the maximum response were calculated. Glucose, tryptone, and yeast extract exerted a strong negative effect over the objective function, with glucose being the strongest. Lutein possessed a positive effect over the tobacco aroma production, while sodium chloride and trace elements showed no influence over the process. The yield attained after culture medium manipulation was almost ten-fold higher, compared with the base medium; and the aroma mixture was characterized as: 7,8 dihydro-beta-ionol (95.2%), 7,8-dihydro-beta-ionone (3.7%), and beta-ionone (1.1%). PMID- 15660216 TI - Optimization of extracellular production of recombinant asparaginase in Escherichia coli in shake-flask and bioreactor. AB - Various host-vector combinations were tested to maximize the extracellular production of recombinant asparaginase in Escherichia coli. Expression of recombinant asparaginase fused to pelB leader sequence under the inducible T7lac promoter in BLR (DE3) host cells resulted in optimum extracellular production in shake-flasks. Fed-batch studies were carried out using this recombinant strain and an exponential feeding strategy was used to maintain a specific growth rate of 0.3 h(-1). To check the effect of the time of induction on expression, cultures were induced with 1 mM isopropyl-beta-D-thiogalactopyranoside at varying cell optical densities (OD(600): 33, 60, 90, 135). Although the specific product formation rates declined with increasing OD of induction, a maximum volumetric activity of 8.7 x 10(5) units l(-1), corresponding to approximately 5.24 g l(-1) of recombinant asparaginase, was obtained when induction was done at an OD(600) of 90. The recombinant protein was purified directly from the culture medium, using a rapid two-step purification strategy, which resulted in a recovery of approximately 70% and a specific activity of approximately 80% of that of the native enzyme. PMID- 15660217 TI - Experimental determination of viability loss of Penicillium bilaiae conidia during convective air-drying. AB - A study was conducted on the drying of Penicillium bilaiae, a fungal micro organism used to promote soil-bound phosphorous uptake in several crop species, such as wheat, canola and pulse crops. A wet pellet formed from a mixture of the inoculant and a starch-based carrier was air-dried to the appropriate water activity to extend the shelf-life of the viable fungal conidia. Convective air drying was examined as a low-energy alternative to the more expensive freeze drying technology that is currently in use. Experiments were conducted to measure the loss of conidia viability during drying in a fixed-bed, thin-layer convective dryer. The dryer air inlet temperature and relative humidity were controlled in experiments to determine the effect of thermal and dessicative stresses on conidial viability. The measured survivor fraction was determined to be dependent on solids temperature, moisture content and drying rate. Thermal stresses became significant for process temperatures above 30 degrees C, while the survivor fraction fell sharply below a dry basis moisture ratio of 30%. Slower drying kinetics associated with high inlet air relative humidity were found to significantly improve the recovery of viable conidia. By minimising environmental stresses, survivor fractions of up to 75% could be achieved, but this result fell dramatically with the introduction of more severe conditions. A general linear statistical model is used to quantify experimental error and the significance level of each factor. PMID- 15660218 TI - Exopolysaccharide biosynthesis by Lactobacillus helveticus ATCC 15807. AB - Exopolysaccharide (EPS) production and the activities of the enzymes involved in sugar nucleotide biosynthesis in Lactobacillus helveticus ATCC 15807 under controlled pH conditions were investigated. Batch fermentations using lactose as energy source showed higher EPS synthesis by L. helveticus ATCC 15807 at pH 4.5 with respect to pH 6.2, the enzyme alpha-phosphoglucomutase (alpha-PGM) being correlated with both total and specific EPS production. When glucose was used as carbon source instead of lactose, the lower EPS synthesis obtained was linked to a decrease in alpha-PGM and galactose 1-phosphate-uridyltransferase (GalT) activities, the reduction of the latter being more pronounced. Higher EPS production by L. helveticus ATCC 15807 at the acidic constant pH of 4.5 requires that both alpha-PGM and GalT activities are high. These enzymes are needed to synthesize UDP-glucose and UDP-galactose for supplying the corresponding monomers for EPS biosynthesis. Although differences are observed in EPS production by this strain regarding the energy source (lactose or glucose), the monomeric composition of the polymers produced is independent of the carbohydrate used. The obtained results contribute to a better understanding of the physiological factors that affect EPS biosynthesis by lactobacilli, which could help in the correct handling of the fermentation parameters within the fermented dairy industry. PMID- 15660219 TI - Heterologous expression of enterocin A, a bacteriocin from Enterococcus faecium, fused to a cellulose-binding domain in Escherichia coli results in a functional protein with inhibitory activity against Listeria. AB - The genes for the bacteriocins enterocin A and B were isolated from Enterococcus faecium ATB 197a. Using the pET37b(+) vector, the enterocin genes were fused to an Escherichia coli specific export signal sequence, a cellulose-binding domain (CBD(cenA)) and a S-tag under the control of a T7lac promotor. The constructs were subsequently cloned into E. coli host cells. The expression of the recombinant enterocins had different effects on both the host cells and other Gram-positive bacteria. The expression of entA in Esc. coli led to the synthesis and secretion of functional active enterocin A fusion proteins, which were active against some Gram-positive indicator bacteria, but did not influence the viability of the host cells. In contrast, the expression of enterocin B fusion proteins led to a reduced viability of the host cells, indicating a misfolding of the protein or interference with the cellular metabolism of Esc. coli. Indicator strains of Gram-positive bacteria were not inhibited by purified enterocin B fusion proteins. However, recombinant enterocin B displayed inhibitory activity after the proteolytic cleavage of the fused peptides. PMID- 15660220 TI - Engineering of pyranose 2-oxidase from Peniophora gigantea towards improved thermostability and catalytic efficiency. AB - To improve the stability and catalytic efficiency of pyranose 2-oxidase (P2Ox) by molecular enzyme evolution, we cloned P2Ox cDNA by RACE-PCR from a cDNA library derived from the basidiomycete Peniophora gigantea. The P2Ox gene was expressed in Escherichia coli BL21(DE3), yielding an intracellular and enzymatically active P2OxB with a volumetric yield of 500 units/l. Site-directed mutagenesis was employed to construct the P2Ox variant E540K (termed P2OxB1), which exhibited increased thermo- and pH-stability compared with the wild type, concomitantly with increased catalytic efficiencies (k(cat)/K(m)) for D-xylose and L-sorbose. P2OxB1 was provided with a C-terminal His(6)-tag (termed P2OxB1H) and subjected to directed evolution using error-prone PCR. Screening based on a chromogenic assay yielded the new P2Ox variant K312E (termed P2OxB2H) that showed significant improvements with respect to k(cat)/K(m) for D-glucose (5.3-fold), methyl-beta-D glucoside (2.0-fold), D-galactose (4.8-fold), D-xylose (59.9-fold), and L-sorbose (69.0-fold), compared with wild-type P2Ox. The improved catalytic performance of P2OxB2H was demonstrated by bioconversions of L-sorbose that initially was a poor substrate for wild-type P2Ox. This is the first report on the improvement of a pyranose 2-oxidase by a dual approach of site-directed mutagenesis and directed evolution, and the application of the engineered P2Ox in bioconversions. PMID- 15660221 TI - Stable isotope dilution analysis of wine fermentation products by HS-SPME-GC-MS. AB - The aim of this study was to quantify, in a single analysis, 31 volatile fermentation-derived products that contribute to the aroma of red and white wine. We developed a multi-component method based on headspace solid-phase microextraction coupled with gas chromatography mass spectrometry (HS-SPME-GC MS). The 31 volatile compounds analysed include ethyl esters, acetates, acids and alcohols. Although these compounds have a range of functional groups, chemical properties, volatilities, affinities for the SPME fibre, and are found in wine at various concentrations, the accuracy of the analysis was achieved with the use of polydeuterated internal standards for stable isotope dilution analyses (SIDA). Nine of the labelled standards were commercially available, while 22 were synthesised. The method was validated by a series of duplicate spiked standard additions to model, white and red wine matrices over the concentration range relevant for each compound in wine. This demonstrated that the appropriate use of SIDA helped to account for matrix effects, for instance potential sources of variation such as the relative response to the MS detector, ionic strength, ethanol content and pH of different wine matrices. The resultant calibration functions had correlation coefficients (R(2)) ranging from 0.995 to 1.000. Each compound could be quantified at levels below its aroma threshold in wine. Relative standard deviations were all <5%. The method was optimised for the best compromise (over the 31 compounds) of wine dilution factor, level of sodium chloride addition, SPME fibre, SPME temperature, SPME time, GC column and MS conditions. Confirmation of identity was achieved by retention time and peak shape, and measurement of at least three ions for each analyte and internal standard with the MS operating in selected ion monitoring mode to facilitate more precise quantitation with a high sampling rate. The method is a valuable research tool with many relevant applications. A novel method for the combined chiral separation and SIDA quantification of 2- and 3-methylbutanoic acid is also demonstrated. PMID- 15660222 TI - Electrochemical characteristics of a platinum electrode modified with a matrix of polyvinyl butyral and colloidal Ag containing immobilized horseradish peroxidase. AB - A new hydrogen peroxide biosensor was constructed, which consisted of a platinum electrode modified by a matrix of polyvinyl butyral (PVB) and nanometer-sized Ag colloid containing immobilized horseradish peroxidase (HRP), and using Co(bpy)3(3+) as mediator in the hydrogen peroxide solution. The electrochemical characteristics of the biosensor were studied by cyclic voltammetry and chronoamperometry. The modified process was characterized by electrochemical impedance spectroscopy and cyclic voltammetry. The HRP immobilized on colloidal Ag was stable and retained its biological activity. The sensor displays excellent electrocatalytic response to the reduction of H2O2. Analytical parameters such as pH and temperature were also studied. Linear calibration for H2O2 was obtained in the range of 1x10(-5) to 1x10(-2) M under optimized conditions. The sensor was highly sensitive to H2O2, with a detection limit of 2x10(-6) M, and the sensor achieved 95% of steady-state current within 10 s. The sensor exhibited high sensitivity, selectivity and stability. PMID- 15660223 TI - Assessing the biocompatibility of NiTi shape memory alloys used for medical applications. AB - The present paper reviews aspects related to the biocompatibility of NiTi shape memory alloys used for medical applications. These smart metallic materials, which are characterised by outstanding mechanical properties, have been gaining increasing importance over the last two decades in many minimal invasive surgery and diagnostic applications, as well as for other uses, such as in orthodontic appliances. Due to the presence of high amounts of Ni, the cytotoxicity of such alloys is under scrutiny. In this review paper we analyse work published on the biocompatibility of NiTi alloys, considering aspects related to: (1) corrosion properties and the different methods used to test them, as well as specimen surface states; (2) biocompatibility tests in vitro and in vivo; (3) the release of Ni ions. It is shown that NiTi shape memory alloys are generally characterised by good corrosion properties, in most cases superior to those of conventional stainless steel or Co-Cr-Mo-based biomedical materials. The majority of biocompatibility studies suggest that these alloys have low cytotoxicity (both in vitro and in vivo) as well as low genotoxicity. The release of Ni ions depends on the surface state and the surface chemistry. Smooth surfaces with well-controlled structures and chemistries of the outermost protective TiO2 layer lead to negligible release of Ni ions, with concentrations below the normal human daily intake. PMID- 15660224 TI - Marital and sexual satisfaction in testicular cancer survivors and their spouses. AB - GOAL: To compare marital and sexual satisfaction of men who survived testicular cancer (TC) and their spouses to a reference group, and to compare marital and sexual satisfaction of couples who had a relationship at time of diagnosis (couples during TC) to couples who developed a relationship after completion of treatment (couples after TC). PATIENTS AND METHODS: Two hundred and nineteen couples during TC and 40 couples after TC completed the Maudsley Marital Questionnaire, a validated instrument to measure marital and sexual satisfaction. RESULTS: Survivors and spouses of both couple groups reported similar marital satisfaction as men and women of the reference group. Survivors (t=2.9, p<0.01) and spouses (t=2.9, p<0.01) of couples during TC and survivors of couples after TC (t=1.9, p=0.05) reported less sexual satisfaction than the reference groups. Survivors of couples after TC reported less sexual satisfaction than survivors of couples during TC (F=4.0, p<0.05). Correlations between sexual satisfaction of survivors and spouses in couples during TC (r=0.76, p<0.001) and couples after TC (r=0.77, p<0.001) were high. CONCLUSION: Testicular cancer did not appear to have a negative effect on marital satisfaction in couples during TC, although TC survivors and their spouses reported less sexual satisfaction than men and women of the reference group. Survivors who developed a relationship after completion of treatment seemed to form a vulnerable group: their sexual satisfaction was lower than that of men in the reference group and of TC survivors with a longer relationship. Besides that, they more often reported marital problems than their spouses did. PMID- 15660225 TI - Motives for becoming and remaining member of patient associations: a study of 1,810 Swedish individuals with cancer associations. AB - Patient associations for cancer patients (PACPs) are increasing in number as well as in the number of members. We utilised a questionnaire to investigate how members of 13 PACPs motivated their memberships. The study included 1,810 individuals who had been treated for breast cancer, gynaecological cancer, or prostate cancer. Through questionnaires these individuals were asked to articulate why they became and chose to remain members. Descriptive statistics and content analyses were used to analyse the open and structured questions. Motives for membership reflected both benefits for the individuals and the welfare of others; themes such as 'needs related to having cancer' (reported by 33% as motives for becoming members; 14% for remaining members), 'wanted to use the PACP's information and activities' (24%; 38%) and 'wanted to support the PACP and its possibilities to have an impact' (9%; 20%) were dominant. The theme 'needs and experiences related to having cancer' was more common among members with breast cancer (38%) and ovarian cancer (36%) than among members with prostate cancer (25%), whereas 53% of men with prostate cancer reported 'wanted to use the PACP's information and activities' compared to 19-9% among female members. The motives showed that needs related to having cancer and that activities and information offered by the PACPs were important to the members, as were their beliefs that the PACP prompted issues that were important to the members. PMID- 15660226 TI - Characterization of Usher syndrome type I gene mutations in an Usher syndrome patient population. AB - Usher syndrome type I (USH1), the most severe form of this syndrome, is characterized by profound congenital sensorineural deafness, vestibular dysfunction, and retinitis pigmentosa. At least seven USH1 loci, USH1A-G, have been mapped to the chromosome regions 14q32, 11q13.5, 11p15, 10q21-q22, 21q21, 10q21-q22, and 17q24-25, respectively. Mutations in five genes, including MYO7A, USH1C, CDH23, PCDH15 and SANS, have been shown to be the cause of Usher syndrome type 1B, type 1C, type 1D, type 1F and type 1G, respectively. In the present study, we carried out a systematic mutation screening of these genes in USH1 patients from USA and from UK. We identified a total of 27 different mutations; of these, 19 are novel, including nine missense, two nonsense, four deletions, one insertion and three splicing defects. Approximatelly 35-39% of the observed mutations involved the USH1B and USH1D genes, followed by 11% for USH1F and 7% for USH1C in non-Acadian alleles and 7% for USH1G. Two of the 12 MYO7A mutations, R666X and IVS40-1G > T accounted for 38% of the mutations at that locus. A 193delC mutation accounted for 26% of CDH23 (USH1D) mutations, confirming its high frequency. The most common PCDH15 (USH1F) mutation in this study, 5601 5603delAAC, accounts for 33% of mutant alleles. Interestingly, a novel SANS mutation, W38X, was observed only in the USA cohort. The present study suggests that mutations in MYO7A and CDH23 are the two major components of causes for USH1, while PCDH15, USH1C, and SANS are less frequent causes. PMID- 15660227 TI - Signature of recent historical events in the European Y-chromosomal STR haplotype distribution. AB - Previous studies of human Y-chromosomal single-nucleotide polymorphisms (Y-SNPs) established a link between the extant Y-SNP haplogroup distribution and the prehistoric demography of Europe. By contrast, our analysis of seven rapidly evolving Y-chromosomal short tandem repeat loci (Y-STRs) in over 12,700 samples from 91 different locations in Europe reveals a signature of more recent historic events, not previously detected by other genetic markers. Cluster analysis based upon molecular variance yields two clearly identifiable sub-clusters of Western and Eastern European Y-STR haplotypes, and a diverse transition zone in central Europe, where haplotype spectra change more rapidly with longitude than with latitude. This and other observed patterns of Y-STR similarity may plausibly be related to particular historical incidents, including, for example, the expansion of the Franconian and Ottoman Empires. We conclude that Y-STRs may be capable of resolving male genealogies to an unparalleled degree and could therefore provide a useful means to study local population structure and recent demographic history. PMID- 15660228 TI - Relative ototoxicity of 21 aromatic solvents. AB - Some aromatic solvents (e.g. toluene, p-xylene, styrene, and ethylbenzene) show, in the rat, striking ototoxicity characterized by an irreversible hearing loss, as measured by behavioural or electrophysiological methods, associated with damage to outer hair cells in the cochlea of the exposed animals. To broaden the range of aromatic solvents studied concerning their potential ototoxicity and to compare their ototoxicity quantitatively, 21 aromatic solvents were administered orally by gastric intubation to Sprague-Dawley rats for 5 days/week for a 2-week period. The dose used was 8.47 mmol kg(-1) body weight day(-1). The possible ototoxicity of the aromatic solvents was evaluated by morphological investigation of the cochlea. Whole-mount surface preparations of the organ of Corti were made to quantify the number of missing hair cells (cytocochleogram). Among the 21 solvents studied, eight (toluene, p-xylene, ethylbenzene, n-propylbenzene, styrene, alpha-methylstyrene, trans-beta-methylstyrene, and allylbenzene) caused histological lesions of the organ of Corti. They differed widely in their potency. The least ototoxic solvents caused outer hair cell (OHC) loss in the middle turn of the organ of Corti. The OHC loss was slight in the first row, and greater in the second and third rows. The most ototoxic solvents caused high losses in the three rows of the outer hair cells along the entire length of the basilar membrane. There were also occasional inner hair cell (ICH) losses in the most affected animals. Although no measurements were made of the chemical concentrations reached in the blood or the brain, tentative ranking of an increasing ototoxicity of the eight aromatic solvents could be proposed on the basis of the histological losses observed-alpha-methylstyrene 0.05). Lumbar and femur neck BMD scores improved significantly at the end of treatment in all three groups (p < 0.05). There was no statistically-significant difference in BMD scores between groups at final (p > 0.05). Urinary DPD/Cre levels decreased significantly in all three groups by the end of the year (p < 0.05). There was no statistically-significant difference in urinary DPD/Cre final levels between groups (p > 0.05). According to the results of the present study, consecutive 10 days therapy with SCT, which is the first in the literature to our knowledge, is as effective as the other two regimens in the treatment of osteoporosis. Both cyclic regimens in our study (alternating 15 days and 10 consecutive days each month for 1 year) do appear to offer some advantages, especially economically and clinically, as compared to continuous treatment. PMID- 15660236 TI - A case of idiopathic hypertrophic cranial pachymeningitis manifested only by positive rheumatoid factor and abnormal findings of the anterior falx. AB - In this report, we present a rare case of a 52-year-old man with a unique form of hypertrophic pachymeningitis involving the anterior part of the falx and who was positive for rheumatoid factor. The clinical symptom was only headache, without any cranial nerve palsies or ataxia. Diagnosis was made by gallium scintigraphy and magnet resonance imaging but was not confirmed by dural biopsy. Treatment with corticosteroid alone was extremely effective for him, while in most cases hypertrophic pachymeningitis recurs or progresses despite the treatment. PMID- 15660237 TI - Distinct post-transcriptional modifications result into seven alternative transcripts of the CC-NBS-LRR gene JA1tr of Phaseolus vulgaris. AB - The generation of splice variants has been reported for various plant resistance (R) genes, suggesting that these variants play an important role in disease resistance. Most of the time these R genes belong to the Toll and mammalian IL-1 receptor-nucleotide-binding site-leucine-rich repeat (TIR-NBS-LRR) class of R genes. In Phaseolus vulgaris, a resistance gene cluster (referred to as the B4 R gene cluster) has been identified at the end of linkage group B4. At this complex resistance cluster, three R specificities (Co-9, Co-y and Co-z) and two R QTLs effective against the fungal pathogen Colletotrichum lindemuthianum, the causal agent of anthracnose, have been identified. At the molecular level, four resistance gene candidates encoding putative full-length, coiled-coil (CC)-NBS LRR R-like proteins, with LRR numbers ranging from 18 to 20, have been previously characterized. In the present study, seven cDNA corresponding to truncated R-like transcripts, belonging to the CC-NBS-LRR class of plant disease R genes, have been identified. These seven transcripts correspond to a single gene named JA1tr, which encodes, at most, only five LRRs. The seven JA1tr transcript variants result from distinct post-transcriptional modifications of JA1tr, corresponding to alternative splicing events of two introns, exon skipping and multiple 'aberrant splicing' events in the open reading frame (ORF). JA1tr was mapped at the B4 R-gene cluster identified in common bean. These post-transcriptional modifications of the single gene JA1tr could constitute an efficient source of diversity. The present results provide one of the few reports of transcript variants with truncated ORFs resulting from a CC-NBS-LRR gene. PMID- 15660238 TI - Development of a co-dominant, PCR-based marker for allelic selection of the pink trait in onions (Allium cepa), based on the insertion mutation in the promoter of the anthocyanidin synthase gene. AB - Bulb color in onions (Allium cepa) is an important trait and is inherited in a complex manner. However, the mechanism of color inheritance is poorly understood at the molecular level. A previous study showed that pink bulb color in onions is inherited as a single recessive trait. This trait is attributable to a significantly reduced transcription of the anthocyanidin synthase (ANS) gene. In this study, we developed a PCR-based marker for an allelic selection of the ANS gene to avoid the laborious progeny tests traditionally employed. To identify polymorphisms between pink and red alleles of the ANS gene, promoter sequences of both alleles were isolated. There was 97% nucleotide sequence identity between the promoter sequences of the two alleles. A 390-bp insertion was identified 632 bp upstream from the putative transcription start site in the pink allele. A pair of primers was designed on the flanking sequences of the inserted region and utilized as a PCR-based marker for allelic selection of the ANS gene. The reliability of the marker was tested using parents, F1 hybrids, and F3 lines whose genotypes had been identified by progeny tests. The marker was also used to evaluate the distribution of the pink allele in white and yellow breeding lines. The results indicated that a majority of the breeding lines tested were homozygous recessive. PMID- 15660239 TI - TM2, a novel strong matrix attachment region isolated from tobacco, increases transgene expression in transgenic rice calli and plants. AB - Nuclear matrix attachment regions (MARs) are thought to influence the expression of the flanking genes. TM2, a new DNA fragment isolated from tobacco, can bind with the rice nuclear matrix in vitro. In this study, we investigated the effect of TM2 on transgene expression under the control of three different promoters in stably transformed rice calli and plants. The presence of TM2 flanking the transgene increased the expression of constructs based on the constitutive CaMV 35S and maize ubiquitin gene promoters in both resistant calli and transformed plants. The GUS expression directed by the photosynthetic-tissue-specific PNZIP promoter was also increased in photosynthetic tissues of transformants. However, TM2 did not change the gene expression pattern controlled by the PNZIP promoter. The effect of TM2 in transgenic plants was stronger than that in transgenic calli based on all three promoters. Our results indicate that TM2, as a novel strong MAR, can be used to increase the transgene expression levels in the whole plant or in particular tissues of monocotyledons. PMID- 15660240 TI - [Ocular tilt reaction. A rare complication after acoustic neuroma surgery]. AB - Visual disturbances after acoustic neuroma surgery are very rare complications. We present a patient with diplopia after a middle fossa approach for resection of an acoustic neuroma. The patient presented ocular torsion and tilts of subjective visual vertical which is typical for the ocular tilt reaction. It is generally agreed that this reaction is secondary to a dysfunction of the tonic bilateral vestibular inputs that stabilize the eyes and head in a normal upright position in the roll plane. It has also been described after denervation of the utricle and injury of the brain stem. PMID- 15660241 TI - [Minimally invasive substitute tissue in head and neck surgery]. PMID- 15660242 TI - Effects of combined lamotrigine and valproate on basal and stimulated extracellular amino acids and monoamines in the hippocampus of freely moving rats. AB - The antiepileptic drugs sodium valproate (VPA) and lamotrigine (LTG) are increasingly used in combination in patients in whom monotherapy has failed to control seizures. Although these drugs are known to interact pharmacokinetically, several authors have proposed a pharmacodynamic interaction between the two. In order to investigate this we have studied the effects of combined treatment with LTG and VPA on basal and stimulated extracellular aspartate (ASP), glutamate (GLU), taurine (TAU), gamma amino butyric acid (GABA), 5-hydroxytryptamine (5-HT) and dopamine (DA) release in the hippocampus of freely moving rats using microdialysis. Additionally, we measured the possible effect of VPA on LTG in plasma, whole brain and dialysates. Neither LTG (10 mg/kg) nor VPA (300 mg/kg) given alone significantly altered basal levels of ASP, GLU or TAU. When given together, however, the two drugs significantly reduced extracellular ASP and GLU while increasing TAU levels. In the case of GABA, LTG was without effect on basal levels of the transmitter, but these increased following VPA and this persisted with both drugs. When transmitter release was stimulated by 50 muM veratridine, marked increases in the release of all amino acids occurred and this was decreased by LTG in all cases. VPA alone only altered GABA release, increasing it by approximately the same extent as basal GABA. For all of the amino acids studied, however, VPA reversed the decreases in release seen after LTG. VPA and LTG increased and decreased respectively basal 5-HT and DA. When given together the increase in extracellular 5-HT was greatly prolonged, but no effect on DA release was seen. When 5-HT release was evoked by veratridine this was increased by VPA and no other treatment. With DA, however, neither drug alone altered evoked release, but the two combined led to a marked increase. Co-administration of VPA with LTG showed no significant effect of this combination on LTG in any of the three compartments studied indicating that in this case a significant pharmacokinetic contribution to our findings is unlikely, which suggests that there is a probable pharmacodynamic interaction of the two drugs. PMID- 15660243 TI - Search for an endogenous cannabinoid-mediated effect in the sympathetic nervous system. AB - Activation of CB(1) cannabinoid receptors by exogenous agonists causes presynaptic inhibition of neurotransmitter release from axon terminals. In the central nervous system, presynaptic CB(1) receptors can also be activated by endogenous cannabinoids (endocannabinoids) released from postsynaptic neurons. Except in the vas deferens, there is no indication of endocannabinoid-mediated presynaptic inhibition in the sympathetic nervous system. The aim of the present study was to search for such inhibition in pithed rats. Artificial sympathetic tone was established by continuous electrical stimulation of preganglionic sympathetic axons. The CB(1) cannabinoid receptor antagonist rimonabant (0.5 and 2 mg kg(-1) i.v.) did not change blood pressure, heart rate or plasma noradrenaline concentration. Since activation of Galpha(q/11) protein-coupled receptors enhances endocannabinoid synthesis in the central nervous system, we attempted to stimulate endocannabinoid production by infusion of arginine vasopressin and phenylephrine (both activate Galpha(q/11) protein-coupled receptors). Rimonabant (2 mg kg(-1) i.v.) did not change blood pressure, heart rate or plasma noradrenaline concentration during infusion of phenylephrine or vasopressin. In the final series of experiments we verified that an exogenous cannabinoid agonist produces sympathoinhibition. The synthetic CB(1)/CB(2) receptor agonist WIN55212-2 (0.1 and 1 mg kg(-1) i.v.) markedly lowered blood pressure and plasma noradrenaline concentration in pithed rats with electrically stimulated sympathetic outflow. In contrast, in pithed rats with a pressor infusion of noradrenaline, WIN55212-2 did not change blood pressure or heart rate. The results verify that activation of peripheral presynaptic CB(1) receptors inhibits noradrenaline release from sympathetic nerve terminals. The lack of effect of the CB(1) receptor antagonist rimonabant indicates that, even under conditions favouring endocannabinoid synthesis, endocannabinoid-mediated presynaptic inhibition is not operating in the sympathetic nervous system of the pithed rat. PMID- 15660245 TI - Collateral damage or apocalypse now for European academic research. PMID- 15660244 TI - Breath-to-breath analysis of abdominal and rib cage motion in surfactant-depleted piglets during high-frequency oscillatory ventilation. AB - OBJECTIVE: To assess the value of monitoring abdominal and rib cage tidal displacement as an indicator of optimal mean airway pressure (Paw) during high frequency oscillatory ventilation (HFOV). DESIGN AND SETTING: Prospective observational study in a university research laboratory. ANIMALS: Eight piglets weighing 12.0+/-0.5 kg, surfactant depleted by lung lavage. INTERVENTIONS: Compliance of the respiratory system (C(rs)) was calculated from a quasistatic pressure volume loop. After initiation of HFOV lung volume was recruited by increasing Paw to 40 cmH(2)O. Then mean Paw was decreased in steps until PaO(2)/FIO(2) was below 100 mmHg. Proximal pressure amplitude remained constant. MEASUREMENTS AND RESULTS: Abdominal and rib cage tidal displacement was determined using respiratory inductive plethysmography. During HFOV there was maximum in tidal volume (Vt) in seven of eight piglets. At maximal mean Paw abdominal and rib cage displacement were in phase. Phase difference between abdominal and rib cage displacement increased to a maximum of 178+/-28 degrees at minimum mean Paw. A minimum in abdominal displacement and a maximum of Vt was found near the optimal mean Paw, defined as the lowest mean Paw where shunt fraction is below 0.1. CONCLUSIONS: During HFOV abdominal and rib cage displacement displayed mean Paw dependent asynchrony. Maximal Vt and minimal abdominal displacement coincided with optimal C(rs), oxygenation, and ventilation, suggesting potential clinical relevance of monitoring Vt and abdominal displacement during HFOV. PMID- 15660247 TI - Overlapping stent-supported coil embolization of wide-neck basilar tip aneurysm: technical case reports. AB - We report two cases in which wide-neck basilar aneurysms were treated with overlapping stent-supported coil embolization. This overlapping technique ensures complete stent interface with the aneurysm neck and likely improves long-term outcomes. We demonstrated the feasibility of this novel technique and suggest its application for other bifurcation aneurysms requiring stent-supported coil embolization. PMID- 15660246 TI - Regulation of natural killer cell function: a role for the NK cell's own MHC class I molecules. PMID- 15660248 TI - Magnetization transfer ratio in neuro-Behcet disease. AB - The aim of this study was to determine the contribution of magnetization transfer ratios (MTRs) in detecting disease in normal-appearing brain regions of patients with neuro-Behcet (NB) disease. Thirty-two patients with NB disease were assessed. Fifteen healthy volunteers were examined as the control group. Magnetic resonance (MR) imaging of the head was performed without and with magnetization transfer (MT) contrast. Signal intensity measurements were obtained from ten anatomical regions (centrum semiovale, corona radiata, internal capsule, forceps major, forceps minor, thalamus, substantia nigra pars compacta, substantia nigra pars grisea, inferior pons and middle cerebellar peduncle) in both groups. Also measured in the NB group were parenchymal lesions in the brain stem, basal ganglia and cerebral deep white matter. MTR was calculated for each measurement. Statistical analysis was performed with Mann-Whitney U and independent t-tests with computer-based SPSS 11.0 for Windows software. A P value below 0.05 was considered statistically significant. The mean MTR of the parenchymal lesions in the NB group was lower than the mean MTR of the normal-appearing parenchyma in both the NB patients and the normal group. For the normal-appearing parenchyma the mean MTR in the NB group was higher than that for the controls for all regions except the corona radiata; however, the difference was statistically significant only for the thalamus. The MRI-visible parenchymal involvement of Behcet's disease causes a decrease in MTR. For the normal-appearing brain, although lacking statistical significance for the most regions studied, the tendency for higher MTR in NB patients compared with controls may offer an insight into the pathophysiology of Behcet's disease. PMID- 15660249 TI - Recognition of a familiar place by the honeybee (Apis mellifera). AB - Recent work shows that at any one place bees detect a limited variety of simple cues in parallel. At each choice point, they recognize a few cues in the range of positions where the cues occurred during the learning process. There is no need to postulate that they re-assemble the surrounding panorama in memory; only that they retain memories of the coincidences of cues in the expected retinotopic directions. The cues could be stimuli that excite groups of peripheral visual neurons. All the experimentally known cues are described, including modulation of the receptors, the locations of areas of black or colour, the nearness, size, averaged edge orientation, and radial and tangential edges. Cues of each type are separately summed within large fields, the size of which varies with the cue. Local orientation cues from edges at right angles cancel each other within each field, which also suggests that the discrimination of shape and texture is limited. Resolution depends on lateral interactions and the number of ommatidia required for each cue. To identify a new place, a few sparse cues, together with their directions, are learned in orientation flights. When the bee returns, the cues in the panorama are progressively matched as they coincide with the cues in memory. The limited number of cues, though economical for memory, may restrict the foraging behaviour and lead to flower constancy. This kind of a visual system is a candidate model for other animals or machines with economical processing systems. PMID- 15660250 TI - The structure of the USP/RXR of Xenos pecki indicates that Strepsiptera are not closely related to Diptera. AB - The receptor for the insect molting hormone, ecdysone, is a heterodimer consisting of the Ecdysone Receptor and Ultraspiracle (USP) proteins. The ligand binding domain sequences of arthropod USPs divide into two distinct groups. One group consists of sequences from members of the holometabolous Lepidoptera and Diptera, while the other arthropod sequences group with vertebrate retinoid-X receptors (RXRs). We therefore wondered whether USP/RXR structure could be used to clarify the contentious phylogenetic position of the order Strepsiptera, which has proposed affinities with either Diptera or Coleoptera. We have cloned and sequenced the USP/RXR from the strepsipteran Xenos pecki. Phylogenetic analyses are not consistent with a close affinity between Strepsiptera and Diptera. PMID- 15660251 TI - Characterization of SpAPETALA3 and SpPISTILLATA, B class floral identity genes in Spinacia oleracea, and their relationship to sexual dimorphism. AB - Floral organ identity B class genes are generally recognized as being required for development of petals and stamens in angiosperm flowers. Spinach flowers are distinguished in their complete absence of petals in both sexes, and the absence of a developed stamen whorl in female flowers. As such, we hypothesized that differential expression of B class floral identity genes is integral to the sexual dimorphism in spinach flowers. We isolated two spinach orthologs of Arabidopsis B class genes by 3' and 5' RACE. Homology assignments were tested by comparisons of percent amino acid identities, searches for diagnostic consensus amino acid residues, conserved motifs, and phylogenetic groupings. In situ hybridization studies demonstrate that both spinach B class genes are expressed throughout the male floral meristem in early stages, and continue to be expressed in sepal primordia in reduced amounts at later stages of development. They are also highly expressed in the third whorl primordia when they arise and continue to be expressed in these tissues through the development of mature anthers. In contrast, neither gene can be detected in any stage in female flowers by in situ analyses, although northern blot experiments indicate low levels of SpAP3 within the inflorescence. The early, strong expressions of both B class floral identity genes in male floral primordia and their absence in female flowers demonstrate that B class gene expression precedes the origination of third whorl primordia (stamen) in males and is associated with the establishment of sexual floral dimorphism as it initiates in the first (sepal) whorl. These observations suggest that regulation of B class floral identity genes has a role in the development of sexual dimorphism and dioecy in spinach rather than being a secondary result of organ abortion. PMID- 15660252 TI - A reliable method to detect bacterial contamination of blood components using an automated blood culture system. PMID- 15660253 TI - Staphylococcus aureus carriage among participants at the 13th European Congress of Clinical Microbiology and Infectious Diseases. AB - The aim of this study was to measure the rate of Staphylococcus aureus nasal colonization among attendees of the 13th European Congress of Clinical Microbiology and Infectious Diseases (ECCMID), particularly with regard to methicillin-resistant (MRSA) strains. The 31.4% rate of Staphylococcus aureus colonization detected among the participants was in line with colonization rates reported previously for healthcare workers. A statistical difference was found between the rates of Staphylococcus aureus carriage in physicians (37.4%) and non physicians (21.7%) but not between males (35.0%) and females (28.9%). Only one participant (a Belgian physician) was found to carry MRSA. Surprisingly, the rate of methicillin-susceptible Staphylococcus aureus carriage was significantly higher among participants from countries with a low prevalence of MRSA. PMID- 15660254 TI - Emergence of endocarditis due to group D streptococci: findings derived from the merged database of the International Collaboration on Endocarditis. AB - The aim of the present study was to compare the epidemiological and clinical characteristics of Streptococcus bovis endocarditis with those of endocarditis caused by oral streptococci, using data obtained from a large international database of uniformly defined cases of infective endocarditis. S. bovis, a well known cause of infective endocarditis, remains the common name used to designate group D nonenterococcal streptococci. In some countries, the frequency of S. bovis endocarditis has increased significantly in recent years. Data from the International Collaboration on Endocarditis merged database was used to identify the main characteristics of S. bovis endocarditis and compared them with those of infective endocarditis (IE) due to oral streptococci. The database contained 136 cases of S. bovis IE and 511 cases of IE due to oral streptococci. Patients with S. bovis IE were significantly older those with IE due to oral streptococci (63+/ 16 vs. 55+/-18 years, P<0.00001). The proportion of streptococcal IE due to S. bovis increased from 10.9% before 1989 to 23.3% after 1989 (P=0.0007) and was 56.7% in France as compared with 9.4% in the rest of Europe and 6.0% in the USA (P<0.00001). Patients with S. bovis IE had more comorbidity and never used intravenous drugs. Complication rates, rates of valve replacement, and mortality rates were similar in the two groups. In conclusion, this study confirmed that S. bovis IE has unique characteristics when compared to endocarditis due to oral streptococci and that it emerged in the 1990s, mainly in France, a finding that is yet unexplained. PMID- 15660255 TI - Prevalence and risk factors of extended-spectrum beta-lactamase-producing Escherichia coli and Klebsiella pneumoniae in an Israeli long-term care facility. AB - The purpose of this study was to ascertain the prevalence of extended-spectrum beta-lactamases (ESBLs) among Escherichia coli and Klebsiella pneumoniae strains obtained from urine samples of residents of a long-term care facility and to determine the risk factors for acquisition of ESBL-producing strains. All urine samples collected from January 2003 to October 2003 that were positive for E. coli or K. pneumoniae were tested for the presence of ESBL. Records of patients with ESBL-positive (ESBL-P) samples were analyzed for clinical and demographic data. The records of a matched control group of patients whose urine samples were positive for E. coli or K. pneumoniae but were ESBL-negative (ESBL-N) were also analyzed. The overall rate of ESBLs among the E. coli and K. pneumoniae samples was 25.6%. Of 350 urine samples that grew E. coli, 77 (22%) were positive for ESBL; 34 of 84 (40.5%) samples that grew K. pneumoniae were ESBL-P. Male sex, treatment in the subacute care unit, recent antimicrobial treatment, pressure sores, (percutaneous endoscopic gastrostomy) PEG tube, anemia, hypoalbuminemia, permanent urinary catheter, and any recent invasive procedure were all associated with ESBL-P bacteria in the univariate analysis. The multivariate analysis revealed three independent risk factors for the presence of an ESBL-producing strain: anemia, permanent urinary catheter, and previous antibiotic use. Fluoroquinolones were most strongly associated with the development of ESBL producing bacteria. The prevalence of ESBL-producing E. coli and K. pneumoniae in the long-term care facility investigated was unexpectedly high and corroborates the notion that long-term care facilities could be important reservoirs of resistant bacteria. Identification of the risk factors for ESBLs is the first step in formulating an effective strategy to curtail the spread of ESBL resistance in long-term care facilities. PMID- 15660256 TI - Capillary haemangioma arising from the anterior choroidal artery. AB - BACKGROUND: Capillary haemangioma of the central nervous system is extremely rare. Histologically proven cases developed in the dura mater and choroid plexus, or were typically intracranial extensions of an extra-cranial lesion. FEATURES: This report details a case that developed in the anterior choroidal artery of a newborn infant and manifested as a lethal intra-cerebral haemorrhage. Pathological criteria for the diagnosis of vascular malformations should be carefully investigated and the differential diagnoses of the present case are discussed. PROGNOSIS: Intracranial haemangioma presents a diagnostic challenge and the treatment of deep lesions remains problematic. PMID- 15660257 TI - Certification in nuclear cardiology in Europe. PMID- 15660258 TI - [methyl-3H]Choline incorporation into MCF7 tumour cells: correlation with proliferation. AB - PURPOSE: The aim of the study was to investigate the intracellular location of [methyl-(3)H]choline in MCF7 tumour cells and to determine the relationship between [methyl-(3)H]choline incorporation and proliferation. METHODS: Tumour cells were incubated with [methyl-(3)H]choline for 10 min, and then in cold medium to simulate the rapid blood clearance of [methyl-(11)C]choline. Labelled metabolites were then extracted from cells by treating them with organic and aqueous solvents to determine the distribution of tracer between phospholipid and water-soluble metabolite pools. Aqueous extracts were subjected to thin-layer chromatography, ion exchange chromatography and a choline extraction procedure to identify (3)H-containing metabolites. Procedures were carried out on fast- and slow-growing populations of MCF7 cells to determine the relationship between choline incorporation and proliferation. RESULTS: Only about 5% of [methyl (3)H]choline was present as phospholipid. [methyl-(3)H]choline incorporation was found to be related to S-phase fraction. In another experiment, [methyl (14)C]choline incorporation was found to be correlated with [methyl (3)H]thymidine incorporation. The V(max) of choline uptake was found to be increased whilst K(m) was decreased in populations of MCF7 cells with higher proliferative fractions, compared with populations having lower proliferative fractions. CONCLUSION: Choline incorporation into tumour cells under conditions that simulate rapid blood clearance of [methyl-(11)C]choline is correlated with proliferation. Most of the activity (about 95%) was in the non-lipid fraction of the cell. PMID- 15660259 TI - Regulation of the voltage-gated potassium channel KCNQ4 in the auditory pathway. AB - The potassium channel KCNQ4, expressed in the mammalian cochlea, has been associated tentatively with an outer hair cell (OHC) potassium current, I(K,n), a current distinguished by an activation curve shifted to exceptionally negative potentials. Using CHO cells as a mammalian expression system, we have examined the properties of KCNQ4 channels under different phosphorylation conditions. The expressed current showed the typical KCNQ4 voltage-dependence, with a voltage for half-maximal activation (V(1/2)) of -25 mV, and was blocked almost completely by 200 microM linopirdine. Application of 8-bromo-cAMP or the catalytic sub-unit of PKA shifted V(1/2) by approximately -10 and -20 mV, respectively. Co-expression of KCNQ4 and prestin, the OHC motor protein, altered the voltage activation by a further -15 mV. Currents recorded with less than 1 nM Ca(2+) in the pipette ran down slowly (12% over 5 min). Buffering the pipette Ca(2+) to 100 nM increased the run-down rate sevenfold. Exogenous PKA in the pipette prevented the effect of elevated [Ca(2+)](i) on run-down. Inhibition of the calcium binding proteins calmodulin or calcineurin by W-7 or cyclosporin A, respectively, also prevented the calcium-dependent rapid run-down. We suggest that KCNQ4 phosphorylation via PKA and coupling to a complex that may include prestin can lead to the negative activation and the negative resting potential found in adult OHCs. PMID- 15660260 TI - Glycation of low-density lipoproteins by methylglyoxal and glycolaldehyde gives rise to the in vitro formation of lipid-laden cells. AB - AIMS/HYPOTHESIS: Previous studies have implicated the glycoxidative modification of low-density lipoprotein (LDL) by glucose and aldehydes (apparently comprising both glycation and oxidation), as a causative factor in the elevated levels of atherosclerosis observed in diabetic patients. Such LDL modification can result in unregulated cellular accumulation of lipids. In previous studies we have characterized the formation of glycated, but nonoxidized, LDL by glucose and aldehydes; in this study we examine whether glycation of LDL, in the absence of oxidation, gives rise to lipid accumulation in arterial wall cell types. METHODS: Glycated LDLs were incubated with macrophage, smooth muscle, or endothelial cells. Lipid loading was assessed by HPLC analysis of cholesterol and individual esters. Oxidation was assessed by cholesterol ester loss and 7-ketocholesterol formation. Cell viability was assessed by lactate dehydrogenase release and cell protein levels. RESULTS: Glycation of LDL by glycolaldehyde and methylglyoxal, but not glucose (in either the presence or absence of copper ions), resulted in cholesterol and cholesterol ester accumulation in macrophage cells, but not smooth muscle or endothelial cells. The extent of lipid accumulation depends on the degree of glycation, with increasing aldehyde concentration or incubation time, giving rise to greater extents of particle modification and lipid accumulation. Modification of lysine residues appears to be a key determinant of cellular uptake. CONCLUSIONS/INTERPRETATION: These results are consistent with LDL glycation, in the absence of oxidation, being sufficient for rapid lipid accumulation by macrophage cells. Aldehyde-mediated "carbonyl-stress" may therefore facilitate the formation of lipid-laden (foam) cells in the artery wall. PMID- 15660261 TI - Circulating endothelial cells are elevated in patients with type 2 diabetes mellitus independently of HbA(1)c. AB - AIMS/HYPOTHESIS: Patients with diabetes mellitus are well known to be at high risk for vascular disease. Circulating endothelial cells (CECs) have been reported to be an ex vivo indicator of vascular injury. We investigated the presence of CECs in the peripheral blood of 25 patients with diabetes mellitus and in nine non-diabetic control donors. METHODS: Endothelial cells were isolated from peripheral blood with anti-CD-146-coated immunomagnetic Dynabeads, and were stained with acridine orange dye and counted by fluorescence microscopy. The cells were also stained for von Willebrand factor and Ulex europaeus lectin 1. RESULTS: Patients with diabetes mellitus had an elevated number of CECs (mean 69+/-30 cells/ml, range 35-126) compared with healthy controls (mean 10+/-5 cells/ml, range 3-18) (p<0.001). The increase in CECs did not correlate with the levels of HbA(1)c. Circulating endothelial cell numbers were elevated regardless of glucose levels, suggesting that, even with control of glucose levels, there is increased endothelial cell sloughing. CONCLUSIONS: Our study suggests that the higher number of CECs in patients with type 2 diabetes may reflect ongoing vascular injury that is not directly dependent on glucose control. PMID- 15660262 TI - Low subcutaneous thigh fat is a risk factor for unfavourable glucose and lipid levels, independently of high abdominal fat. The Health ABC Study. AB - AIMS: We investigated whether low subcutaneous thigh fat is an independent risk factor for unfavourable glucose and lipid levels, and whether these associations differ between sexes, and between white and black adults. Our secondary aim was to investigate which body composition characteristics (lean tissue, fat tissue) are reflected by anthropometric measures (waist and thigh circumference). METHODS: Anthropometric measurements and computed tomography of the abdomen and of the thigh were performed for all participants of the Health, Aging and Body Composition Study, who were aged 70-79 years. Fasting glucose, triglycerides and HDL-cholesterol, and 2-h postload glucose were determined. RESULTS: After excluding those already diagnosed with diabetes or dyslipidaemia, we analysed data from 2,106 participants. After adjustment for abdominal subcutaneous and visceral fat, and intermuscular thigh fat, larger thigh subcutaneous fat area was statistically significantly associated with lower ln-transformed triglycerides [standardised beta (95% CI) -0.12 (-0.20 to -0.04) in men and -0.13 (-0.21 to 0.05) in women] and higher ln-HDL-cholesterol [0.10 (0.02 to 0.19) and 0.09 (0.01 to 0.18), respectively]. The associations with lower glucose levels were strong in men [-0.11 (-0.20 to -0.02) for fasting and -0.14 (-0.23 to -0.05) for postload glucose], but not statistically significant in women [-0.02 (-0.10 to 0.07) and -0.04 (-0.13 to 0.05), respectively]. There were no differences in the associations between white and black persons. Waist circumference was more strongly associated with abdominal subcutaneous fat, and this association became stronger with increasing BMI, whereas the association with visceral fat became weaker. Thigh circumference was equally dependent on thigh fat and thigh muscle in men, whereas in women the fat component was the main contributor. CONCLUSION: Larger subcutaneous thigh fat is independently associated with more favourable glucose (in men) and lipid levels (in both sexes) after accounting for abdominal fat depots, which are associated with unfavourable glucose and lipid levels. Anthropometric measures reflect different fat depots at different levels of BMI at the abdomen, and reflect both fat and lean tissue at the thigh. These results emphasise the importance of accurate measures of regional body composition when investigating potential health risks. PMID- 15660263 TI - Genetic testing for maturity onset diabetes of the young: uptake, attitudes and comparison with hereditary non-polyposis colorectal cancer. AB - AIMS/HYPOTHESIS: Mutations in hepatic nuclear factor 1alpha cause a monogenic form of diabetes, maturity onset diabetes of the young type 3 (MODY3). Our aim was (1) to assess the uptake of genetic testing for MODY3 and to determine factors affecting it, and (2) to compare attitudes to predictive genetic testing between families with MODY3 and a previously studied group at risk of hereditary non-polyposis colorectal cancer (HNPCC). METHODS: Adult members of two extended MODY3 pedigrees, either with diabetes or a 50% risk of having inherited the mutation (n=144, age 18-60 years), were invited to an educational counselling session followed by a possibility to obtain the gene test result. Data were collected through questionnaires before counselling and 1 month after the test disclosure. RESULTS: Eighty-nine out of 144 (62%) participated in counselling, and all but one wanted the test result disclosed. No significant sociodemographic differences were observed between the participants and non-participants. The counselling uptake was similar among diabetic and non-diabetic subjects. Uncertainty about the future and the risk for the children were the most common reasons to take the gene test. At follow-up, most subjects in both MODY3 (100%) and HNPCC (99%) families were satisfied with their decision to take the test and trusted the result. The majority of both diabetic and non-diabetic subjects considered that the MODY3 gene test should be offered either in childhood (50 and 37%) or as a teenager (30 and 37%). CONCLUSIONS: Genetic testing for MODY3 was well accepted among both diabetic and non-diabetic participants. The subjects found the gene test reliable and they were satisfied with their decision regarding the predictive test. PMID- 15660264 TI - Labor analgesia for the parturient with pregnancy-induced hypertension: what does an obstetrician need to know? AB - INTRODUCTION: Pregnancy-induced hypertension (PIH), also known as preeclampsia, remains one of the leading causes of maternal death worldwide. The term preeclampsia describes the development of hypertension with proteinuria and/or pathologic edema after the 20th week of gestation. The parturient with PIH usually has multiple organ alterations, which may affect the selection of analgesia and anesthesia for labor and delivery. DISCUSSION: This article reviews the special concerns posed by PIH, one of the most common medical conditions encountered in pregnancy. PMID- 15660265 TI - Prognostic value of some ovarian reserve tests in poor responders. AB - BACKGROUND: The objective of this study was to determine the prognostic value of the basal estradiol (E2) and inhibin-B levels, the antral follicle count (AFC), and the clomiphene citrate challenge test (CCCT) of ovarian response in controlled ovarian hyperstimulation (COH), in an outcome with normal follicle stimulating hormone (FSH) concentration in the early follicular phase of the menstrual cycle. METHODS: Fifty-two patients undergoing IVF treatment were included in the study. Blood samples were collected for assessment of basal E2, FSH, and inhibin-B levels. Transvaginal ultrasound of an unstimulated cycle was performed to determine the mean antral follicle count (AFC). Serum FSH concentration was measured again on day 10 for CCCT performance. RESULTS: The mean values of women's age, and basal and day 10 FSH levels were significantly higher in cancelled cycles than in the control group, whereas basal inhibin-B and AFC were significantly higher in the latter. The mean basal E2 concentration was similar in both groups. The results from the logistic regression analysis show that CCCT (cut-off point FSH > 12.5 mIU/ml; AUCROC = 0.90) was a better single predictor of poor ovarian response than AFC (AUCROC = 0.85) and inhibin-B (AUCROC = 0.79) with a correct prediction for CCCT (86.5%), antral follicle count (84.6%), and for inhibin-B (82.7%). CONCLUSIONS: In women with normal basal FSH level, the determination of E2 has no prognostic value for the outcome of poor responders. However, CCCT, AFC, and inhibin-B tests, when applied separately, produce good prognostic values. CCCT is the best single predictor of poor ovarian response, followed by antral follicle count and basal inhibin-B values. In spite of that, CCCT does not add significantly to the simpler AFC ultrasound test in the prediction of poor ovarian response. PMID- 15660267 TI - Ganglioneuroma and adenocarcinoma associated with neurofibromatosis type 1 in the colorectal region. PMID- 15660268 TI - Biochemical and immunohistochemical analysis of glycosaminoglycans in inflamed and non-inflamed intestinal mucosa of patients with Crohn's disease. AB - OBJECTIVES: Changes in extracellular matrix glycosaminoglycans (GAGs) of the intestinal mucosa have been associated with inflammatory bowel disease. The aim of the present study was to follow the changes in GAGs metabolism during the progression from non-inflamed to inflamed intestinal colon of patients with Crohn's disease (CD), using direct biochemical analysis and specific immunohistochemistry against chondroitin/dermatan sulfate and heparan sulfate. DESIGN AND METHODS: The content of GAGs from inflamed and non-inflamed colon of eight patients with active CD was estimated by uronic acid per dry weight of tissue and analyzed by agarose gel electrophoresis and ion-exchange chromatography. Intestinal sections were stained using antibodies against dermatan sulfate/chondroitin 4-sulfate (DS/CS), heparan sulfate (HS), and ICAM-1 (CD54), and analyzed by confocal microscopy. RESULTS: There was a reduction in the amount of GAGs in the non-inflamed colon of patients with CD. In the inflamed colon, HS, CS and DS showed increased concentrations compared with the non inflamed colon. GAGs showed a diffuse distribution in the lamina propria and in the basement membrane of both inflamed and non-inflamed mucosa of patients with CD. CONCLUSION: We observed a marked reduction in GAGs with altered patterns of distribution in the non-inflamed colon of patients with CD. The increase in the synthesis of GAGs observed in the inflamed colon may be a compensatory mechanism for the restoration of the integrity of the intestinal mucosa. PMID- 15660269 TI - Genetic transformation of Tylophora indica with Agrobacterium rhizogenes A4: growth and tylophorine productivity in different transformed root clones. AB - We have developed an efficient transformation system for Tylophora indica, an important medicinal plant in India, using Agrobacterium rhizogenes strains LBA9402 and A4 to infect excised leaf and stem explants and intact shoots at different sites. The induction of callus and transformed roots was dependent on the bacterial strain, explant type and inoculation site used. Transformed roots were induced only in explants infected with A. rhizogenes strain A4, while an optimal transformation frequency of up to 60% was obtained with intact shoots inoculated at the nodes. The presence of the left-hand transferred DNA (T(L)-DNA) in the genome of T. indica roots induced by A. rhizogenes was confirmed by PCR amplification of the rooting locus genes of A. rhizogenes. Root growth and the production of tylophorine, the major alkaloid of the plant, varied substantially among the nine root clones studied. Both parameters increased over time in liquid cultures, with maximum biomass and tylophorine accumulation occurring within 4-6 weeks of growth in fresh medium. Interestingly, in liquid culture, the culture medium also accumulated tylophorine up to concentrations of 9.78+/-0.21 mg l(-1). PMID- 15660270 TI - High prevalence of cytochrome P450 2A6*1A alleles in a black African population of Ghana. AB - OBJECTIVE: We investigated the frequencies of the functionally important variants of the CYP2A6 gene in black African populations. METHODS: Using genomic DNA sequencing, polymerase chain reaction (PCR)-restriction fragment length polymorphism and allele-specific PCR, the allele frequencies of CYP2A6 *1A, *1B, *2, *4A, *5, *6, *7, *8, *9, *10 and * 11 among 120 black Africans- including 105 Ghanaians, 12 Nigerians, 2 Ivorians and 1 Ugandan-were determined. RESULTS: The allele frequencies were 80.5% for CYP2A6*1A, 11.9% for CYP2A6*1B, 1.9% for CYP2A6*4A and 5.7% for CYP2A6*9 in the Ghanaian subjects. No subject homozygous for the CYP2A6*4A allele, a whole gene deletion type of polymorphism prevalent among Orientals, was found. Furthermore, CYP2A6 variants such as *2, *5, *6, *7, *8, *10 and *11 were absent in these black African populations. CONCLUSIONS: This study provides, for the first time, the results of the analysis of CYP2A6 allele frequency in black African populations and confirms large ethnic differences in the polymorphic CYP2A6 gene. PMID- 15660272 TI - Phase I trial of weekly docetaxel with a 4-weekly cisplatin administration in patients with advanced gastric carcinoma. AB - The docetaxel-cisplatin combination is active against several tumors including gastric cancer but it is followed by severe myelosuppression. Recent experience with weekly taxanes has demonstrated a mild myelotoxicity with high dose intensity. We investigated in a phase I study a weekly schedule of docetaxel on days 1, 8 and 15 and cisplatin on day 1 every 4 weeks in 19 patients with advanced gastric cancer with no prior chemotherapy. Cohorts of patients were treated with escalating doses of docetaxel (starting dose 30 mg/m(2) per week and increments of 10 mg/m(2) per week) and cisplatin (starting dose 70 mg/m(2) and increments of 5 mg/m(2)). Febrile neutropenia was the only dose-limiting event occurring in four (20%) patients; the dose-limiting toxicity was reached at dose level three (docetaxel 40 mg/m(2) per week and cisplatin 75 mg/m(2)). The maximum tolerated dose was 40 mg/m(2) per week for docetaxel and 70 mg/m(2) every 4 weeks for cisplatin. Grade 3/4 neutropenia occurred in six patients (30%); early death occurred in one patient with septic shock because of neutropenia and another with acute coronary ischemia. Two (11%) complete and two (11%) partial responses were documented (ORR 22%; 95% CI 3-39%), with a median response duration of 5 months and median time to progression of 7 months. In conclusion, the combination of weekly docetaxel plus cisplatin is feasible with moderate toxicity and merits further investigation in phase II studies in advanced gastric cancer. PMID- 15660271 TI - Continuing clozapine treatment despite neutropenia. AB - RATIONALE: Approximately 1-2% of patients treated with the atypical antipsychotic clozapine develop severe neutropenia and agranulocytosis. The usual recommendation is to discontinue treatment with the drug when the peripheral neutrophil count drops below 1,500/mm3. METHODS: We have reviewed several reports describing procedures that allowed the patients to continue clozapine treatment despite the occurrence of these haematological side effects. RESULTS: The therapeutic procedures described (symptomatic treatment of neutropenia by co administration of lithium or granulopoiesis-stimulating factors, management of the adjunctive medication) seem to be efficient strategies that allow continuation of clozapine treatment despite the occurrence of neutropenia. However, these types of therapy have only been used in a limited number of cases, and the evidence supporting their use remains anecdotal. CONCLUSION: Although the procedures adopted in the cases described in this review are uncommon, they potentially provide an alternative to the discontinuation of clozapine treatment in patients with complex symptomatologies for whom treatment with other antipsychotic medication is insufficient. PMID- 15660273 TI - Segmental cement extraction system (SEG-CES) and the Ilizarov method in limb salvage procedure after total knee cemented prosthesis removal in a former osteosarcoma patient. AB - Survival of osteosarcoma has greatly improved in the past few decades. Knee prosthesis is a well-recognized limb salvage procedure for osteosarcoma of the distal end of the femur. One drawback is that prostheses have a limited life and prosthetic failure with the inherent high rate of reoperations remains a serious long-term problem for former osteosarcoma patients. The segmental cement extraction system (SEG-CES) is a technique to remove cement in arthroplasty revision, based on a cement-bone interface with a lower strength compared to the old cement-new cement interface. We report the case of a 32-year-old former osteosarcoma patient in whom the SEG-CES was applied to remove a long-stemmed total knee cemented prosthesis. The prosthesis was placed 17 years before for a recurrent telangiectatic osteosarcoma of the left femur. Thirteen years after the prosthesis implantation, the patient complained of knee instability, pain, and complete failure of the extensor apparatus. The extraction of the prosthesis was performed using cylindrical batters of diameter corresponding to the diameter of the axle, two hammer extractors clamping the prosthesis components between two jaws. Extraction of the periprosthetic cement in the femoral and tibial components was done using the SEG-CES technique. The successful prosthesis removal performed in this patient allowed us to perform an external fixation with bone lengthening and reconstruction by the Ilizarov method. PMID- 15660274 TI - Comparing in vivo kinematics of unicondylar and bi-unicondylar knee replacements. AB - Preserving both cruciate ligaments in unicondylar knee arthroplasty likely provides more normal knee mechanics and contributes to enhanced patient function. It follows that preserving both cruciate ligaments with total knee arthroplasty should provide functional benefit compared to arthroplasty sacrificing one or both cruciates. The purpose of this study was to compare knee kinematics in patients with optimally functioning cruciate-preserving medial unicondylar and bi unicondylar arthroplasty to determine if knee motions differed. Eight consenting patients with seven medial unicondylar and five bi-unicondylar arthroplasties were studied using lateral fluoroscopy during treadmill gait, stair stepping, and maximum flexion activities. Patient-specific geometric models based on CT and CAD data were used for shape matching to determine the three-dimensional knee kinematics. Tibiofemoral contact locations were computed for the replaced compartments. Maximum flexion in kneeling was 135 degrees +/-14 degrees for unicondylar knees and 123 degrees +/-14 degrees for bi-unicondylar knees (p=0.22). For 0 degrees -30 degrees flexion during the stair activity, the medial condyle translated posterior 3.5+/-2.5 mm in unicondylar knees and 4.7+/-1.9 mm in bi-unicondylar knees (p>0.05). Lateral posterior translation was 5.0+/-2.3 mm in bi-unicondylar knees for 0 degrees -30 degrees flexion. From heel-strike to mid-stance phase, there was little tibial rotation, but unicondylar knees showed 1.5+/-1.6 mm posterior translation of the medial condyle, while bi-unicondylar knees showed 5.1+/-2.2 mm (p<<0.05). The bi-unicondylar knees showed 3.8+/-3.4 mm posterior lateral condylar translation. Preserving both cruciate ligaments in knee arthroplasty appears to maintain some basic features of normal knee kinematics. Knees with bi-unicondylar arthroplasty showed kinematics closer to motions observed in total knee arthroplasty, slightly less weight-bearing flexion, and greater dynamic laxity in gait than unicondylar knees. Despite kinematic differences, knees with unicondylar and bi-unicondylar arthroplasty can provide excellent functional outcomes in appropriately selected patients. PMID- 15660275 TI - [Conventional thorax diagnostics in neonates and children]. AB - Advances in neonatal medicine have led to an increasing number of premature infant patients and to an improved survival rate of these children. Chest X-rays of premature infants, newborns, and older children with respiratory difficulties provide information leading to diagnosis and help decide on further clinical management. Diagnostic findings and their appreciation by the radiologist require knowledge of the specific anatomy of the infant chest, and also of common diseases in early and later childhood, congenital or acquired. Radiologic findings in pediatric patients may differ significantly from those in adults. Close collaboration between the neonatologist and radiologist is essential for reaching the right diagnosis. PMID- 15660277 TI - The influence of defocus on multifocal visual evoked potentials. AB - BACKGROUND: In order to assess the influence of optical factors on the multifocal visual evoked potential (mfVEP), we obtained mfVEPs with optimal refraction and compared them to recordings with various degrees of dioptrical defocus. METHODS: Monocular mfVEPs were recorded from the right eye in eight normal subjects. Dartboard stimuli with 60 sectors arranged in six concentric annuli spanning 60 degrees were generated with a VERIS system and presented on a computer monitor. Two pairs of electrodes were placed 3 cm above and below and 3 cm to the right and left of the inion. Two sets of mfVEP records per subject were obtained, one with best-corrected visual acuity and another when the stimulus was defocused by +1.0, +2.0 or +3.0 D. A signal-to-noise ratio (SNR) measure was calculated for every response from the two channels. RESULTS: The effect of defocus depended on eccentricity: when defocus was at +2.0 D and higher, reducing visual acuity to <0.3, the central mfVEP responses were reduced to approximately 60%, while defocus had no marked effect at eccentricities >7 degrees. CONCLUSIONS: The results suggest that, in contrast to the mfERG, the mfVEP requires optimal refraction to correctly assess the cortical responses. PMID- 15660279 TI - Photodynamic therapy and indocyanine green guided feeder vessel photocoagulation of choroidal neovascularization secondary to choroid rupture after blunt trauma. AB - PURPOSE: To describe photodynamic therapy (PDT) and additional indocyanine green (ICG) guided feeder vessel photocoagulation as a treatment of choroidal neovascularization (CNV) secondary to choroidal rupture in case of blunt head trauma. DESIGN: Interventional case report. METHODS: A 61-year-old woman developed subfoveal CNV originating from a choroid tear 8 years after blunt head trauma. Four sessions of PDT were applied and an additional two consecutive sessions of selective ICG-guided feeder vessel photocoagulation conducted. RESULTS: Transient reduction of leakage and closure of feeder vessels could not prevent further growth of the CNV. CONCLUSIONS: PDT reduced leakage temporarily and additional ICG-guided feeder vessel photocoagulation closed treated feeder vessel and CNV. New feeder vessel formation and growth of CNV in case of traumatic choroid rupture could not be treated effectively by these two laser treatment modalities to prevent severe deterioration of visual acuity. PMID- 15660278 TI - Evaluation of a human capsular bag model for secondary cataract determination after intraocular lens implantation. AB - PURPOSE: To investigate the effect of different intraocular lenses (IOLs) on lens epithelial cells (LECs) and on human capsular bags in vitro and to evaluate the transferability of this model to clinical situations. METHODS: Sham cataract surgery, including IOL-into-the-bag implantation, was performed on 38 donor eyes after removal of the cornea. The capsular bag including the IOL was removed, pinned on a culture dish, covered with medium, and incubated. Two different IOLs were compared per pair of donor eyes. In each pair of donor eyes, the two different IOLs to be compared were implanted individually into capsular bags. The time required for complete coverage of the posterior capsule by a confluent monolayer of LECs was documented. The following IOLs were compared: three-piece acrylic IOLs of different sizes and single-piece polymethylmethacrylate (PMMA) versus acrylic IOLs. RESULTS: A complete monolayer of LECs on the posterior capsule was observed to form at times varying from 8 days (PMMA IOLs) to more than 60 days (three-piece hydrophobic acrylic IOLs). A significant difference between PMMA IOLs and hydrophobic acrylic IOLs was found. CONCLUSIONS: The human capsular bag model employed allows short-term evaluation of secondary cataract formation for different IOLs. This model's correlation with clinical results is good. PMID- 15660280 TI - STRATUS optical coherence tomography in unilateral colobomatous excavation of the optic disc and secondary retinoschisis. AB - PURPOSE: To report the STRATUS optical coherence tomography (STRATUSOCT) findings in a patient with unilateral coloboma-like excavation of the optic disc without pit but secondary retinoschisis, as well as to discuss the possible involved pathophysiologic mechanisms. METHODS: Observational case report. STRATUSOCT findings in a 66-year-old woman with a coloboma-like excavation of the optic disc without pit but secondary retinoschisis encompassing the macular region, along with evidence of a mild epiretinal membrane superonasal to the disc were evaluated. RESULTS: STRATUSOCT showed signs of a connection between the perineural space and the inner retinal layers on the temporal optic disc border, as well as schisis-like changes extending from the disc to the macula, with cystoid degeneration and two lamellar holes in their nasal portion. CONCLUSION: The use of third generation OCT afforded an enhanced visualization of retinal structures, revealing signs of fluid at several distinct levels, as well as deep and superficial inner breaks apart from the schisis cavity. We are unaware of such previous reports, and could find no reference to them in a computerized search using MEDLINE. In addition, our study supports a common pathomechanism for the development of macular complications in optic pits and colobomas. PMID- 15660282 TI - Expression of stem cell markers and transcription factors during the remodeling of the rat pancreas after duct ligation. AB - Ligation of the pancreatic duct has been shown to induce islet cell neogenesis from duct cells in the adult rat pancreas. The transcription factors that regulate islet cell neogenesis and the phenotype of putative precursor cells involved in neogenesis are unknown. We, therefore, studied the expression of the transcription factors Pdx1, Pbx1, Meis2, Nkx2.2 and the putative stem cell markers c-Kit and nestin in rat pancreata 3, 5 and 7 days after duct ligation. Immunocytochemical staining revealed a subpopulation of cells in the ligated portion of the pancreas that was positive for the putative stem cell markers c Kit and nestin. The c-Kit immunoreactivity was upregulated, reaching a peak at day 3, while nestin expression peaked at day 7. The c-Kit-positive cells were located among the duct and islet cells, while nestin-expressing cells were found scattered in the duct epithelium at day 3 and around the ducts at day 7. Both c Kit- and nestin-positive cells showed high proliferative activity, as determined by BrdU labeling. Pdx1 and Nkx2.2 were found predominantly in the duct cells of the ligated pancreas. There were significant changes in the expression patterns of Pbx1 and Meis2 in the ductular complexes. The findings indicate that the stem cell markers c-Kit and nestin as well as the transcription factors Pdx1 and Nkx2.2 are upregulated in compartments of the pancreas that are involved in islet cell neogenesis after duct ligation. PMID- 15660283 TI - Adult sclerosing rhabdomyosarcoma: cytogenetic link with embryonal rhabdomyosarcoma. AB - Rhabdomyosarcomas are classified into three well-defined categories: embryonal, alveolar and pleomorphic rhabdomyosarcoma. Recently, seven cases of an unusual adult type of rhabdomyosarcoma with a prominent hyaline sclerosis have been described. We report the hitherto unreported cytogenetic changes of an adult sclerosing rhabdomyosarcoma. A 79-year-old woman underwent an amputation for a rapidly growing soft tissue mass in the anterior compartment of the right lower leg. The tumor infiltrated the tibia. On histology, a fascicular spindle to round cell proliferation, embedded in a prominent hyaline matrix, was seen. Immunohistochemistry showed focal desmin, myogenin and MyOD1 expression, and electron microscopy revealed Z-band material. Cytogenetic analysis disclosed a 44 49,XX,+del(1)(p22)[2],+11,+16[5],+18[12],+21[3],-22 [cp13] karyotype. Using fluorescent in situ hybridization (FISH) analysis, the tumor cells were negative for FOXO1A-disrupting translocations specific for alveolar rhabdomyosarcoma. The chromosomal composition of malignant cells resembled the pattern of numerical changes frequently observed in embryonal rhabdomyosarcoma, suggesting a close relationship of an adult sclerosing rhabdomyosarcoma with this entity. PMID- 15660281 TI - Immunophenotype of lymphocytic infiltration in medullary carcinoma of the breast. AB - Medullary carcinoma (MC) of the breast is characterized by large anaplastic cells and infiltration by benign lymphocytes. Patients with this pattern of breast carcinoma are considered to have a better prognosis than those with other histological subtypes. We reviewed cases of primary breast carcinoma that were surgically resected between 1990 and 2004. Of these, 13 cases of medullary carcinoma of the breast with lymphocyte infiltration were reported. Tests for CD3, CD4, CD8, CD20, CD56, TIA-1, and granzyme B were performed on paraffin sections. We found that the MC contained very few NK cells, as assessed by their reactivity with the CD56 antibodies. However, MC had a significantly greater percentage of CD3, CD8, TIA-1, and granzyme B lymphocytes infiltrating the stroma of the tumor. Furthermore, more CD8-positive than CD4-positive T-cell lymphocytes were present within the tumor cell nests in MC, as opposed to the proportion in usual ductal carcinoma. The infiltrating cytotoxic/suppressor T cells in MC represent host resistance against cancer, and the high grading of the T-cell infiltration could explain, in part, a key mechanism controlling the good prognosis for this type of tumor and solve the pathological paradox of MC. PMID- 15660284 TI - Lung adenocarcinoma associated with familial adenomatous polyposis. Clear cell carcinoma with beta-catenin accumulation accompanied by atypical adenomatous hyperplasia. AB - A 46-year-old man presented with a lung tumor 17 years after a subtotal colectomy and 13 years after a partial duodenectomy for familial adenomatous polyposis (FAP). There had been no malignant transformation in the specimens from his colectomy and duodenectomy, and a current gastrointestinal investigation revealed no evidence of malignancy. Pathological analysis of the lung tumor demonstrated adenocarcinoma with clear cells and a papillary structure, accompanied by tiny tumorous nodules in the background lung parenchyma. Many of the nodules were multifocal adenocarcinoma; however, some of the nodules demonstrated atypical adenomatous hyperplasia (AAH). This is the first case report of a lung adenocarcinoma accompanied by AAH in a FAP patient. Immunohistochemical and loss of heterozygosity studies revealed unique features of the lesions reflecting a disruption of the adenomatous poliposis coli-beta-catenin pathway. PMID- 15660286 TI - Primary epithelioid hemangioendothelioma of the peritoneum--a diagnostic dilemma. PMID- 15660285 TI - Multiple calcifying fibrous tumors of the pleura. AB - Calcifying fibrous tumor (CFT) is a rare lesion characterized histologically by hypocellular hyalinized collagenous tissue with psammomatous and/or dystrophic calcifications and patchy lymphoplasmacytic infiltrates. CFT usually occurs in the somatic soft tissue of children and young adults but is rarely found in the pleura. We describe here an unusual case of multiple small CFTs in the right mediastinal pleura of a 54-year-old man who had a history of renal cell carcinoma. Suspecting pulmonary and pleural metastases, we performed wedge resection of the right middle lobe and local excision of two nodules in the right pleura. Light microscopy revealed metastatic lesions of renal cell carcinoma in the resected wedge. The pleural nodules were well circumscribed and composed of hypocellular, dense, hyalinized, collagenous tissue with scant lymphoplasmacytic infiltration and characteristic psammoma bodies. Immunohistochemical staining revealed that most spindle cells were positive for vimentin, CD34 and factor XIIIa, and negative for epithelial membrane antigen, keratin, smooth-muscle actin, desmin, S-100 protein and anaplastic lymphoma kinase. We made a histological diagnosis of CFT of the pleura, and the patient remains well 6 months after the wedge resection. PMID- 15660287 TI - Myofibroblastoma-like changes in fibro(stromo)-epithelial lesions of the breast: report of two cases. PMID- 15660288 TI - Reactive arthritis following BCG immunotherapy for bladder carcinoma. AB - Intravesical instillation of bacillus Calmette-Guerin (BCG) is used in the treatment of patients with intermediate and high-risk superficial bladder carcinoma with efficacy and safety. The vast majority of patients do not present any side effects and only 5% of patients have mild and short-lived clinical manifestations such as malaise, low-grade fever, cystitis, and hematuria. Arthralgia and/or arthritis is one of the rare severe complications following intravesical BCG immunotherapy. We report here the case of a patient with reactive arthritis successfully treated with nonsteroidal anti-inflammatory drugs (NSAIDs) after the discontinuation of BCG immunotherapy. PMID- 15660289 TI - Camurati-Engelmann disease: failure of response to bisphosphonates: report of two cases. AB - Camurati-Engelmann disease is a rare bone disorder characterized by cortical thickening of the diaphysis of tubular bones, with sparing of the epiphysis. It has variable degrees of penetrance and expression, but may be very disabling for the affected individuals who manifest the painful symptoms. The authors report on two women with typical presentation of severe Camurati-Engelmann disease whose treatment with bisphosphonates failed to add any improvement beyond that elicited by corticosteroids alone. PMID- 15660290 TI - Multiple avascular necrosis in a patient with systemic lupus erythematosus/systemic sclerosis overlap syndrome. AB - A white female patient developed overlapping features of systemic lupus erythematosus (SLE) and systemic sclerosis (SSc) with severe pulmonary compromise. She was treated with steroids and azathioprine, which improved her clinical condition and spirometric status. In May 2002 she presented with continuous pain in her left ankle that continued even during rest and under treatment with nonsteroidal anti-inflammatory drugs (NSAIDs). Magnetic resonance imaging (MRI) showed multiple avascular necrosis (AVN). Rest and kinesitherapy were indicated for 1 year, and gradually an orthosis was introduced allowing the patient to walk normally. PMID- 15660291 TI - Population structure, admixture, and aging-related phenotypes in African American adults: the Cardiovascular Health Study. AB - U.S. populations are genetically admixed, but surprisingly little empirical data exists documenting the impact of such heterogeneity on type I and type II error in genetic-association studies of unrelated individuals. By applying several complementary analytical techniques, we characterize genetic background heterogeneity among 810 self-identified African American subjects sampled as part of a multisite cohort study of cardiovascular disease in older adults. On the basis of the typing of 24 ancestry-informative biallelic single-nucleotide polymorphism markers, there was evidence of substantial population substructure and admixture. We used an allele-sharing-based clustering algorithm to infer evidence for four genetically distinct subpopulations. Using multivariable regression models, we demonstrate the complex interplay of genetic and socioeconomic factors on quantitative phenotypes related to cardiovascular disease and aging. Blood glucose level correlated with individual African ancestry, whereas body mass index was associated more strongly with genetic similarity. Blood pressure, HDL cholesterol level, C-reactive protein level, and carotid wall thickness were not associated with genetic background. Blood pressure and HDL cholesterol level varied by geographic site, whereas C-reactive protein level differed by occupation. Both ancestry and genetic similarity predicted the number and quality of years lived during follow-up, but socioeconomic factors largely accounted for these associations. When the 24 genetic markers were tested individually, there were an excess number of marker trait associations, most of which were attenuated by adjustment for genetic ancestry. We conclude that the genetic demography underlying older individuals who self identify as African American is complex, and that controlling for both genetic admixture and socioeconomic characteristics will be required in assessing genetic associations with chronic-disease-related traits in African Americans. Complementary methods that identify discrete subgroups on the basis of genetic similarity may help to further characterize the complex biodemographic structure of human populations. PMID- 15660292 TI - Accuracy of haplotype reconstruction from haplotype-tagging single-nucleotide polymorphisms. AB - Many investigators are now using haplotype-tagging single-nucleotide polymorphism (htSNPs) as a way of screening regions of the genome for association with disease. A common approach is to genotype htSNPs in a study population and to use this information to draw inferences about each individual's haplotypic makeup, including SNPs that were not directly genotyped. To test the validity of this approach, we simulated the exercise of typing htSNPs in a large sample of individuals and compared the true and inferred haplotypes. The accuracy of haplotype inference varied, depending on the method of selecting htSNPs, the linkage-disequilibrium structure of the region, and the amount of missing data. At the stage of selection of htSNPs, haplotype-block-based methods required a larger number of htSNPs than did unstructured methods but gave lower levels of error in haplotype inference, particularly when there was a significant amount of missing data. We present a Web-based utility that allows investigators to compare the likely error rates of different sets of htSNPs and to arrive at an economical set of htSNPs that provides acceptable levels of accuracy in haplotype inference. PMID- 15660293 TI - No evidence of association or interaction between the IL4RA, IL4, and IL13 genes in type 1 diabetes. AB - Attempts to identify susceptibility loci that, on their own, have marginal main effects by use of gene-gene interaction tests have increased in popularity. The results obtained from analyses of epistasis are, however, difficult to interpret. Gene-gene interaction, albeit only marginally significant, has recently been reported for the interleukin-4 and interleukin-13 genes (IL4 and IL13) with the interleukin-4 receptor A gene (IL4RA), contributing to the susceptibility of type 1 diabetes (T1D). We aimed to replicate these findings by genotyping both large family and case-control data sets and by using previously published data. Gene gene interaction tests were performed using linear regression models in cases only. We did not find any single-locus associations with T1D and did not obtain evidence of gene-gene interaction. Additional support from independent samples will be even more important in the study of gene-gene interactions and other subgroup analyses. PMID- 15660295 TI - To tell or not to tell? Professional and lay perspectives on the disclosure of personal health information in community-based dementia care. AB - Developments in information technology and the ongoing restructuring of health services to increase provision in community settings militate in favour of a streamlining of communications and the exchange of information about patients among health and social care providers. Yet the principles of confidentiality and privacy appear to inhibit this process. In order to explore the practical, ethical, and legal imperatives attendant upon personal health information exchange, we conducted a series of interviews with professional care providers, persons with early-stage dementia, and their family caregivers. The findings indicate some degree of discordance. Professionals reported valuing disclosure both to colleagues and family caregivers on the basis of its being in the patients' best interests. Patients also valued inter-professional exchange, but sought strong control over disclosure to family members. Family caregivers valued being kept informed of the patient's condition, even without the latter's consent. Implications for research and policy are discussed. PMID- 15660294 TI - "Doing ethics" in the context of sharing patients' personal health information. AB - There are at present two inconsistencies with respect to the sharing of personal health information (PHI) among health care professionals caring for a patient whom the information concerns. First, there is an inconsistency between what is in theory the ethics and law governing the confidentiality and privacy of this information--it may only be disclosed with informed consent--and what is the actual practice of health care professionals--they share it without such consent. Second, there is an inconsistency between what ethics and law demand in theory and what all parties want: They all approve of the current practice. Ethics and law can be brought into line with what is needed to act in the patient's best interests and with what people want, without opening up any serious potential for abuse of privacy and confidentiality, by establishing a safeguarded, provision-of care exception that allows co-caring health care professionals to share patients' PHI. The requirements for a system establishing such safeguards are proposed. The basic governing presumption is respect for the person and for rights to autonomy, self-determination, privacy, and confidentiality. Therefore, disclosure may only be made with the informed consent of the competent person to whom the information pertains, unless a defence of necessity applies. Where there is doubt about someone's competence, there should likewise be doubt about disclosure without that person's informed consent. Where the person is incompetent, such a disclosure can be made to the patient's substitute decision makers, most often the family, if that is necessary for the care of the patient and in the patient's best interests. To the extent possible, consistent with the best interests of the patient, the wishes of incompetent people should be respected. PMID- 15660296 TI - Geriatric day hospital: who improves the most? AB - This study compared the changes in some bio-psychosocial variables (functional independence, nutritional risk, pain, balance and walking, grip strength, general well-being, psychiatric profile, perception of social support, leisure satisfaction, and caregivers' feeling of burden) in four categories of clients during their program at a geriatric day hospital (GDH). The study also evaluated whether or not improvements, if any, were maintained 3 months after discharge. One-hundred-and-fifty-one people, categorized by primary reason for admission, were assessed at the GDH with reliable and valid tools, at admission and at discharge. Three months after discharge, they were reassessed with the same tools. Overall, two categories of clients, stroke / neurological diseases and musculoskeletal disorders / amputations, improved the most. For the gait disorders and falls group, only the functional independence score improved, but not at a clinically significant level. Finally, clients in the cognitive function disorders / psychopathologies group improved the most on their well-being scores and caregivers' burden decreased the most. All gains were maintained up to 3 months after discharge, except for leisure satisfaction. With the exception of clients who attended the GDH because of gait disorders and falls, the improvements and maintenance achieved in each category occurred in the domains where improvement had been hoped for, because of the particular disabilities in question and because of the nature of the GDH services offered. PMID- 15660297 TI - Quasi-experimental study of the effectiveness of an integrated service delivery network for the frail elderly. AB - The aim of this study was to examine the effectiveness of a new, integrated service delivery (ISD) network of health and social services for frail elderly living in a semi-urban community. A quasi-experimental study was conducted from 1997 to 2000, with measures taken before implementation (T0) and every 12 months after implementation for a 3-year period (T1, T2, T3); 482 people aged 75 years or older from 2 communities, 272 in the experimental and 210 in the control group, were followed. Analyses were conducted using parametric and non-parametric statistics, a generalized linear model, and Cox regression for survival analysis. A declining trend in institutionalization was observed and the desire to be institutionalized was lower in the study group. When absence of deterioration on follow-up was analysed in terms of the level of autonomy, the ISD network produced positive effects on the frailer clientele at T1, effects that tended to be maintained at T2. In regard to caregivers' burden, the ISD network seemed to have been effective, with caregivers' burden being less at T1 and T2. The ISD network did not have a marked effect on the utilization of services. The ISD network did, in fact, produce some effects that should be seen as important for the elderly population. PMID- 15660298 TI - Predictors and sequelae of fractures in the elderly: the Canadian Study of Health and Aging (CSHA). AB - The objective of this study was to describe the incidence, type, risk factors, and sequelae of fractures experienced by community-dwelling elderly Canadians. Data are from the Canadian Study of Health and Aging (CSHA), a longitudinal cohort study, collected in three waves: baseline (1991), wave 2 (1996), and wave 3 (2001). In CSHA-2 (1996), fractures were reported by 3.7 per cent of men and 7.5 per cent of women; in CSHA-3 (2001), by 2.9 per cent of men and 7.3 per cent of women. The distribution of fracture types was similar for men and women. Risk factors for women were increased age, history of fracture, and regular alcohol use; for men, stroke, arthritis, and lack of independent bathing. Walkers or wheelchairs were needed by over 30 per cent of those with any lower body fracture. One third of subjects with any fracture and over 50 per cent with femur fractures reported moderate or severe pain. It is concluded that elderly Canadians are at high risk for fractures, and older women are at most risk. Pain and mobility problems were common sequelae. The need for primary and secondary prevention of these age-related events is paramount. PMID- 15660299 TI - The effect of age upon care and outcomes in patients hospitalized for congestive heart failure in Alberta, Canada. AB - We describe the age-specific outcomes for patients hospitalized with newly diagnosed congestive heart failure using administrative hospital abstracts from Alberta, Canada, from April 1, 1994, to March 31, 2000. Seniors (aged 65 years and older) constituted about 85 per cent of the 16,162 patients. Both co morbidity and severity of illness tended to increase with age. The use of special care unit admissions, coronary artery diagnostic services (cardiac catheterization), and revascularization procedures (percutaneous transluminal coronary angioplasty/stenting, coronary artery bypass surgery) peaked in the 50 to 64-year age group and decreased with increasing age. Specialist/sub-specialist care, prescriptions of beta blockers and angiotensin-converting enzyme inhibitors / angiotensin receptor blockers decreased with age in seniors. Adjusted in hospital, 1-year mortality and crude, age-specific 5-year mortality were significantly greater in those 75 years and older. Outcomes and process of care in patients with newly diagnosed congestive heart failure were not uniformly distributed with age. The elderly had greater mortality but received less therapy. PMID- 15660300 TI - Barriers and facilitators in pain management in long-term care institutions: a qualitative study. AB - The purpose of this study was to identify barriers to the management of pain in long-term care institutions. Formal caregivers practising in four long-term care institutions in Hamilton, Ontario participated in eight focus groups. Participants included 6 physicians, 19 registered nurses, 8 registered practical nurses, 13 health care aides and 8 occupational therapists or physiotherapists. Three types of barriers were found: caregiver-related, patient-related and systems-related. Four themes for facilitating pain management were identified: (a) caregivers knowing the patient, (b) family support, (c) caregivers demonstrating understanding and compassion, and (d) teamwork by caregivers. Formal caregivers practising in long-term care institutions encounter a multitude of barriers that hinder the adequate assessment and treatment of pain, such as caregiver beliefs and knowledge and cognitive impairment in patients. Innovative strategies and strong political will are needed to overcome them. PMID- 15660301 TI - An examination of the health profile, service use and care needs of older adults in residential care facilities. AB - Private, unregulated residential care facilities have become an increasingly important component of the continuum of housing and care for frail older adults in Canada. To date, this growing segment of the older population has received very little research attention. This study involved an in-depth examination of the functional/health profile, patterns of service use, and medical/care needs of a representative sample of 178 older adults in residential care facilities in the City of Ottawa. The results indicate great diversity in resident and facility profiles in this setting and confirm earlier impressions that special care units in the residential care sector have become increasingly close to being unlicensed pseudo-nursing homes. Despite the heavy burden of care, the evidence suggests that the care needs of the majority of residents are adequately met in the residential care environment. The results can inform future research, case finding, educational, and policy planning initiatives in this setting. PMID- 15660302 TI - Publicly funded and family-friend care in the case of long-term illness: the role of the spouse. AB - In recent years, considerable attention has been devoted to the value of unpaid work in the economy. One very important aspect of unpaid work is caregiving for chronically ill or disabled people and the question of whether or not family-and friend caregiving eases the burden on the publicly funded system. Using data from the 1996 General Social Survey, this paper investigates the extent to which the presence of a spouse of a senior aged 55 and over with a long-term illness reduces the amount of publicly funded care received. The findings from the multivariate Tobit analysis show that the presence of the spouse does significantly reduce the amount of publicly funded care used. This result is not gender-specific. The total hours of caregiving by the spouse are valued, as well as the savings generated for the publicly funded system, and the policy issue of tax relief for spouses who are engaged in long-term caregiving is explored. PMID- 15660303 TI - Geographic dimensions of aging in Canada, 1991-2001. AB - Although population aging on a national scale has received much attention in Canada, its geographical dimensions have not. This paper examines the demographic processes that underlie population aging at the provincial and metropolitan scales for the periods 1991 to 1996 and 1996 to 2001. We differentiate between the effects of aging-in-place and net migration on population aging. We also examine the relationships between the various measures of aging and the social and economic characteristics of metropolitan areas over the two periods. While aging-in-place is the dominant force in population aging, net migration is more volatile and more likely to produce shifts in the trajectory of aging. Out migration of the young from areas with weak local economies is a particularly important contributor to aging. British Columbia exhibited a sharp increase in aging in the latter part of the 1990s as net in-migration of younger populations declined, while a number of smaller, resource-based, urban communities also exhibited aging, as out-migration of the young accelerated. PMID- 15660305 TI - Help to older parents and parents-in-law: does paid employment constrain women's helping behaviour? AB - While concern has been expressed for some time about the impact of rising female employment on informal help to older adults, few studies have directly compared employed and not-employed women and only rarely has research utilized national, population-based samples. This article examines whether paid employment reduces the provision and/or the intensity of specific types of help offered by women to older parents and parents-in-law. Data were drawn from the 1996 General Social Survey of Canada and consist of a sub-sample of women aged 40 to 64 who had at least one parent or parent-in-law alive. The independent variable is employment status (full-time, part-time, not employed). Dependent variables are type of help (meal preparation, cleaning, home maintenance, grocery shopping, transportation, banking and money management, personal care) and intensity of help. Two additional types of help examined are emotional support and checking up or monitoring. The analysis shows that employed daughters and daughters-in-law are less likely than those who are not employed to provide help with banking or bill paying but no differences are found on other types of help. Employment status is not significantly related to the total amount of time spent helping older parents and parents-in-law, although not-employed women spend more time helping with meal preparation. Women employed part-time are less likely to provide emotional support to a parent or parent-in-law. There are no differences by employment status in the likelihood of checking up on or monitoring the parent's or parent in-law's situation. It is concluded that, on the whole, in the domains measured in this study, employment does not reduce women's provision of help to parents and parents-in-law. PMID- 15660306 TI - Helping to build and rebuild secure lives and futures: financial transfers from parents to adult children and grandchildren. AB - This paper explores intergenerational financial transfers from parents to adult children and grandchildren within a life course perspective. Research on intergenerational financial transfers has been sparse, and little is known about the financial support older Canadians provide to family members or about the meaning of that assistance. Survey data from a convenience sample of older Canadians was analysed to determine the types of financial assistance older persons provide, what motivates them to provide this assistance, and what meanings such transfers have for the older persons themselves. Findings suggest that it is often events and transitions in the lives of adult children that shape the financial assistance that is needed and given within these families. Older parents demonstrate a strong desire to help their children and grandchildren through important or difficult transitions to "build or rebuild secure lives and futures". Parents' assistance is also influenced by their own family history of assistance and their desire to pass on an early inheritance during their lifetime. PMID- 15660307 TI - A life course perspective on the relationship between socio-economic status and health: testing the divergence hypothesis. AB - While adults from all socio-economic status (SES) levels generally encounter a decline in health as they grow older, research shows that health status is tied to SES at all stages of life. The dynamics of the relationship between SES and health over the life course of adult Canadians, however, remain largely unexplored. This paper tests the divergence hypothesis, which postulates that the SES-based gap in health widens with age, using a representative sample of Canadians aged 25 to 79 from the 1994-1995 National Population Health Survey. Multiple linear regression analyses show support for this assumption; that is, the relationship between SES (measured by years of education and annual household income) and health (measured by self-rated and functional health indexes) strengthens with age. The results of this study provide insight into and answers about healthy aging among Canadians. PMID- 15660309 TI - Life cycle theory and the residential mobility of older Canadians. AB - This paper outlines a debate in the economics literature about the role of housing wealth in post-retirement consumption choices and provides a description of the patterns of the residential mobility, tenure, and dwelling-type transitions of older Canadians, using the newly available Statistics Canada Survey of Labour and Income Dynamics (SLID). The paper also presents estimation results for a panel data model that accounts for individual heterogeneity. The patterns of residential mobility of older adults in Canada appear to be similar to those in the United States and some European countries and do not seem to be motivated by the desire to use housing wealth for general consumption. PMID- 15660308 TI - Healthy aging at older ages: are income and education important? AB - Being higher on the socio-economic scale is correlated with being in better health, but is there is a causal relationship? Using 3 years of longitudinal data for individuals aged 50 and older from the Canadian Survey of Labour and Income Dynamics, we study the health transitions for those who were in good health in the first year, focusing especially on income and education. The initial good health restriction removes from the sample those whose incomes may have been affected by a previous history of poor health, thus avoiding a well-known problem of econometric endogeneity. We then ask, for those in good health, whether later transitions in health status are related to socio-economic status. We find that they are-that changes in health status over the subsequent 2 years are related, in particular, to income and education. PMID- 15660310 TI - Understanding the influence of the complex relationships among informal and formal supports on the well-being of caregivers of persons with dementia. AB - This study examined the direct and indirect relationships between caring for a person with dementia and caregiver health. A conceptual model of the caregiver stress process considered informal caregiver characteristics, sources of caregiver stress, and the influence of informal and formal support on the well being of the caregivers of persons with dementia. Structural equation modeling was used to test specific hypotheses in a sample of 327 caregivers of people with dementia who were living in the community, as part of the first wave of the Canadian Study of Health and Aging. Our findings show that the negative impact of the increasing disability of the care recipient on the psychological well-being of the caregiver was mediated mostly by the use of informal support systems and marginally by the use of formal support systems. The use of informal support by caregivers was associated with a decreased use of formal supports. The relationship between caregiver characteristics and their psychological health was mediated by the use of formal support but not by the use of informal support systems. PMID- 15660311 TI - Examining the healthy immigrant effect in mid- to later life: findings from the Canadian Community Health Survey. AB - Recent studies have established that a healthy immigrant effect operates in Canada-immigrants are generally healthier than Canadian-born persons-but that this effect tends to diminish over time, as the health of immigrants converges to the Canadian norm. Although this effect has been examined by place of birth, language, marital status, socio-economic status, charter-language ability, and category of immigrant status in Canada, less is known about the healthy immigrant effect at different stages of the life course, particularly in mid- to later adulthood, stages at which there is an increased likelihood of decline in physical and mental health status. This study examines how age at immigration affects the health of mid- to later-life immigrants, compared to Canadian-born persons, using data from the 2000-2001 Canadian Community Health Survey. These data indicate that the healthy immigrant effect applies to later mid-life immigrants; that is, new immigrants-those who immigrated less than 10 years ago aged 45 to 64 have better health than their longer-term counterparts-those who immigrated 10 or more years ago-whose health status is similar to that of Canadian-born persons. Interestingly, a different picture emerges in old age (65 years and over), where recent immigrants have poorer overall health compared to Canadian-born persons. When a number of socio-demographic, socio-economic, and health behaviour factors are controlled, however, this disadvantage largely disappears. The findings are discussed in terms of their implications for Canadian health care policy and program planning for immigrants in the latter stages of the life course. PMID- 15660312 TI - Reflexive planning for later life. AB - Informed by Giddens' (1991) concept of reflexive life planning and the notion of later life as a time of increasing social and financial risk, this research explores the idea of reflexive planning for later life. We utilize a conceptual model that incorporates three types of planning for later life: public protection, self-insurance, and self-protection. Drawing on qualitative, life history data from a study of 51 mid-life and older Canadians, we examined whether individuals recognized the risks associated with later life, and if so, how far these recognitions entered into the preparations people made for their futures. We also considered how social circumstances facilitate and/or constrain an individual's planning for later life. Overall, most participants recognized risks and engaged in reflexive planning. On the other hand, there was a small group of non-planners, or day-by-dayers who were getting by with little preparation. We suggest that what distinguishes these groups is that the former have a future time perspective, which is associated with certain socio-demographic characteristics, including high household incomes. PMID- 15660313 TI - The economic legacy of divorce and separation for women in old age. AB - Although progress has been made over the last 20 years, the burden of a low income in old age is still carried by unattached women. Few researchers, however, have examined exactly where the burden of poverty falls within the category of unattached older women or what the nature of this poverty is. Like any other group of older Canadians, unattached women are not a homogeneous population. The category of unattached includes the separated, divorced, widowed, and ever single, all of whom face different circumstances in old age because of differences over the life course. Using Survey of Labour and Income Dynamics (SLID) data, we examine income and sources of income from 1993 to 1999 to identify differences among these groups. The findings indicate that the separated and divorced are the poorest of all older unattached women in Canada. A key source of the difference is the differential growth in private pension incomes. PMID- 15660314 TI - The interplay between women's life course work patterns and financial planning for later life. AB - In order to gain a comprehensive understanding of the interplay between women's life course work patterns and their financial planning for later life, we examined data from semi-structured interviews with retired women (n = 28) aged 59 to 92. The majority of women disrupted their careers at some point in time, for an average of 14 years, primarily for child-rearing responsibilities. We found that financial preparedness and income security in later life are structured by women's life course work patterns. However, individuals also have the ability to shape their own lives and many of the women took the initiative to acquire financial knowledge irrespective of their work situation. Financial-planning advice that participants gave to future generations of older women was also explored and centred on the importance of saving, avoiding debt, maintaining financial independence, and planning ahead. PMID- 15660317 TI - Molecular genetics of Hirschsprung's disease. PMID- 15660318 TI - Primary versus staged pull-through for the treatment of Hirschsprung disease. PMID- 15660319 TI - Laparoscopic-assisted approaches for the definitive surgery for Hirschsprung's disease. AB - The surgical management of Hirschsprung's disease has progressed from a two- or three-stage procedure to a primary operation over the last 25 years. More recently, definitive surgery for Hirschsprung's disease has been performed using minimally invasive techniques. The Swenson, Duhamel and Soave endorectal pull through procedures have all been reported using minimally invasive approaches. The endorectal dissection has become the dominant minimal access procedure because of the ease and reliability in performing this technique and the excellent results obtained. Although a transanal endorectal pull-through can be performed without laparoscopy, the laparoscopic-assisted transanal endorectal pull-through is a much more versatile technique and allows early biopsies to determine the extent of aganglionic and dysfunctional bowel before ablation of the rectum and mesocolon. The authors use a laparoscopic-assisted transanal pull through for aganglionosis of the left and transverse colon. Total colon aganglionosis or aganglionosis of the ascending colon is managed by a laparoscopic-assisted Duhamel procedure which provides a better reservoir in patients with a short or absent colon. PMID- 15660316 TI - Developmental biology of the enteric nervous system: pathogenesis of Hirschsprung's disease and other congenital dysmotilities. PMID- 15660320 TI - Enterocolitis complicating Hirschsprung's disease. AB - Enterocolitis remains a relatively common complication of Hirschsprung's disease with significant morbidity and mortality. The etiology of Hirschsprung's enterocolitis (HEC) is multifactorial and remains poorly understood. Preventative measures and better treatment modalities will evolve out of a better understanding of the underlying pathophysiology. Prompt recognition of HEC allows early intervention and a potential reduction in disease severity and mortality. This review of HEC describes the epidemiology, clinical and pathological features and current best practice in management. Some of the areas of research into etiology and treatment are discussed. PMID- 15660321 TI - Long-term results of treatment of Hirschsprung's disease. AB - Many children with Hirschsprung's disease (HD) have a good outcome following surgical treatment, but long-term follow-up studies have identified a number of concerns. Analysis of long-term function in children after surgical management is difficult. The most commonly encountered problems include constipation, incontinence, enterocolitis and the overall impact of the disease on lifestyle (quality of life). Other complications are less frequent. Each of these problems will be discussed. PMID- 15660322 TI - Intestinal transplantation for total/near-total aganglionosis and intestinal pseudo-obstruction. AB - Whether from anatomical short gut (such as after resection of extensive intestinal aganglionosis) or from a functional cause (such as intestinal pseudoobstruction), intestinal failure is a devastating disease process with profound morbidity and mortality. These patients require total parenteral nutrition (TPN) and are at risk of developing complications such as liver failure, catheter-related sepsis and loss of venous access. Intestinal transplantation, which has advanced markedly over the last 14 years, is now the accepted standard of care for patients failing TPN. Survival outcomes have improved significantly, infectious complications are better controlled, and new immunosuppressive therapies offer great hope for the future. In particular, the results of intestinal transplantation achieved with the motility disorders are equivalent to those experienced with other causes of intestinal failure. In themselves, the motility disorders present their own set of complicating factors, including determining the extent of the disease process (which may involve any part of the gastrointestinal tract), associated urological anomalies, and the type of organ transplantation required. Extensive workup and careful consideration is required before transplantation is undertaken. However, early referral is desirable once complications arise if these patients are to be offered optimal medical care before the chance of transplantation is lost. PMID- 15660323 TI - Variant Hirschsprung's disease. AB - Conditions that clinically resemble HD despite the presence of ganglion cells on suction rectal biopsy results, can be diagnosed by providing an adequate biopsy and employing a variety of histological techniques. Intestinal neuronal dysplasia is a distinct clinical entity that can be clearly proven histologically. Patients with IND not only have abnormalities of submucosal and myenteric plexuses but also defective innervation of the muscle. Internal sphincter achalasia, which is histologically characterized by nitrergic nerve depletion, can be diagnosed on anorectal manometry and successfully treated by internal sphincter myectomy. The outcome of smooth muscle disorders is generally fatal. The need for surgical intervention should be weighed carefully and individualized because most explorations have not been helpful and are probably not necessary. PMID- 15660324 TI - Diagnosis and management of children with intractable constipation. PMID- 15660328 TI - Uric acid, evolution and primitive cultures. AB - Hypertension is epidemic and currently affects 25% of the world's population and is a major cause of stroke, congestive heart failure, and end-stage renal disease. Interestingly, there is evidence that the increased frequency of hypertension is a recent event in human history and correlates with dietary changes associated with Westernization. In this article, we review the evidence that links uric acid to the cause and epidemiology of hypertension. Specifically, we review the evidence that the mutation of uricase that occurred in the Miocene that resulted in a higher serum uric acid in humans compared with most other mammals may have occurred as a means to increase blood pressure in early hominoids in response to a low-sodium and low-purine diet. We then review the evidence that the epidemic of hypertension that evolved with Westernization was associated with an increase in the intake of red meat with a marked increase in serum uric acid levels. Indeed, gout and hyperuricemia should be considered a part of the obesity, type 2 diabetes, and hypertension epidemic that is occurring worldwide. Although other mechanisms certainly contribute to the pathogenesis of hypertension, the possibility that serum uric acid level may have a major role is suggested by these studies. PMID- 15660329 TI - Hereditary hyperuricemia and renal disease. AB - Hyperuricemia and gout have long been known to run in families. As well as an apparently multifactorial genetic component to classic gout itself, 2 rather unusual sex-linked single-gene disorders of purine biosynthesis or recycling have been defined: deficiency of the enzyme hypoxanthine-guaninephosphoribosyl transferase (HPRT), and overactivity of PPriboseP synthase. Both result in overproduction of urate, hyperuricemia, and secondary overexcretion that may lead to acute or chronic renal damage. Familial juvenile hyperuricemic nephropathy (FJHN) and autosomal-dominant medullary cystic kidney disease (ADMCKD) are more common but less well-defined hyperuricemic conditions resulting from a decrease in the fractional excretion of filtered urate, with normal urate production. Although having features in common, ADMCKD is distinguished in particular by the presence of medullary cysts. One major group of both disorders is associated with mutations in the gene for uromodulin, but this accounts for only about one third of cases, and genetic heterogeneity is present. Whether the genes involved in these latter disorders contribute to the polygenic hyperuricemia and urate underexcretion of classic gout remains unexplored. PMID- 15660330 TI - Hemodynamics of hyperuricemia. AB - Prolonged hyperuricemia is associated with the development of hypertension, renal arteriolosclerosis, glomerulosclerosis, and tubulointerstitial injury. It confers a greater risk than proteinuria for developing chronic renal disease and is associated with the development of hypertension. Mild chronic hyperuricemia without intrarenal crystal deposition was induced in rats by inhibiting uricase with oxonic acid. Hyperuricemic rats developed hypertension, afferent arteriolar thickening, and mild renal interstitial fibrosis. Additionally, hyperuricemia accelerated renal damage and vascular disease in rats undergoing renal ablation. To better understand the role of hyperuricemia in the kidney, micropuncture studies were performed. Hyperuricemia resulted in renal cortical vasoconstriction (single nephron glomerular filtration rate (SNGFR) 35%, P < .05) and glomerular hypertension (P < .05). The possibility that hyperuricemia could modify renal hemodynamic disturbances during progression of renal disease was tested in rats with 5/6 nephrectomy. Hyperuricemia accentuated the renal vascular damage and caused cortical vasoconstriction (SNGFR 40%, P < .05) and persistent glomerular hypertension. In conclusion, hyperuricemia impairs the autoregulatory response of preglomerular vessels, resulting in glomerular hypertension. Lumen obliteration induced by vascular wall thickening results in severe vasoconstriction. The resulting ischemia is a potent stimulus that induces tubulointerstitial inflammation and fibrosis as well as arterial hypertension. PMID- 15660331 TI - Clinical evidence for the influence of uric acid on hypertension, cardiovascular disease, and kidney disease: a statistical modeling perspective. AB - This article critically evaluates the clinical evidence regarding the influence of uric acid on hypertension, cardiovascular disease, and kidney disease. Data on these relationships are largely observational and exceedingly complex. The complexity is owing to indirect and direct relations, and bidirectional influences, simultaneously operating on multiple outcomes. Limitations of previous analyses include inadequate statistical methods using only bivariate correlations or poorly specified multiple regression models. As a result, great controversy developed as to whether uric acid is an independent predictor of important outcomes. An example of such analytic limitations is including hypertension as an independent variable, together with uric acid, in a multivariate model for predicting cardiovascular disease. Hypertension may predict significant variance in cardiovascular disease, but the contribution of uric acid may not be recognized if uric acid exerts its influence indirectly through hypertension. Path analysis, which can model direct and indirect influences on outcomes simultaneously, would address this substantive question. Studies of uric acid in relation to hypertension, cardiovascular disease, and kidney disease using a path-analytic approach would help specify such conditions as well as optimize design of clinical trials to determine if decreasing uric acid levels improves outcomes. PMID- 15660332 TI - Uric acid and hypertension in adolescents. AB - Hypertension is the most common form of cardiovascular disease. Although it is less common in adolescents than in adults, hypertension and the associated organ damage can and often does begin early in life. Consequently, for many with high blood pressure (BP), measures directed at the long-term prevention of cardiovascular morbidity may need to be started in adolescence to achieve maximal effectiveness. This article discusses some of the problems unique to hypertension in the young, as well as recent discoveries regarding the likely role played by increases in serum uric acid level in the development of adolescent-onset essential hypertension. PMID- 15660333 TI - Uric acid as a mediator of endothelial dysfunction, inflammation, and vascular disease. AB - Recent experimental findings have led to renewed interest in the possible role of uric acid in the pathogenesis of both hypertension and vascular disease. Often considered an antioxidant, biochemical and in vitro data indicate that noncrystalline, soluble uric acid also can react to form radicals, increase lipid oxidation, and induce various pro-oxidant effects in vascular cells. In vitro and in vivo findings suggest that uric acid may contribute to endothelial dysfunction by inducing antiproliferative effects on endothelium and impairing nitric oxide production. Proinflammatory and proliferative effects of soluble uric acid have been described on vascular smooth muscle cells (VSMCs), and in animal models of mild hyperuricemia, hypertension develops in association with intrarenal vascular disease. Possible adverse effects of uric acid on the vasculature have been linked to increased chemokine and cytokine expression, induction of the renin angiotensin system, and to increased vascular C-reactive protein (CRP) expression. Experimental evidence suggests a complex but potentially direct causal role for uric acid in the pathogenesis of hypertension and atherosclerosis. PMID- 15660334 TI - Uric acid and chronic renal disease: possible implication of hyperuricemia on progression of renal disease. AB - Although hyperuricemia has long been associated with renal disease, uric acid has not been considered as a true mediator of progression of renal disease. The observation that hyperuricemia commonly is associated with other risk factors of cardiovascular and renal disease, especially hypertension, has made it difficult to dissect the effect of uric acid itself. However, recent epidemiologic evidence suggests a significant and independent association between the level of serum uric acid and renal disease progression with beneficial effect of decreasing uric acid levels. Furthermore, our experimental data using hyperuricemic animals and cultured cells have provided robust evidence regarding the role of uric acid on progression of renal disease. Hyperuricemia increased systemic blood pressure, proteinuria, renal dysfunction, vascular disease, and progressive renal scarring in rats. Recent data also suggest hyperuricemia may be one of the key and previously unknown mechanisms for the activation of the renin-angiotensin and cyclooxygenase-2 (COX-2) systems in progressive renal disease. Although we must be cautious in the interpretation of animal models to human disease, these studies provide a mechanism to explain epidemiologic data that show uric acid is an independent risk factor for renal progression. Although there is no concrete evidence yet that uric acid bears a causal or reversible relationship to progressive renal disease in humans, it is time to reevaluate the implication of hyperuricemia as an important player for progression of renal disease and to try to find safe and reasonable therapeutic modalities in individual patients based on their clinical data, medication history, and the presence of cardiovascular complications. PMID- 15660335 TI - Uric acid and transplantation. AB - Hyperuricemia is a common complication in organ transplant recipients, and frequently is associated with chronic cyclosporine immunosuppressive therapy. Kidney and heart transplant recipients are prone to develop posttransplant hyperuricemia. Risk factors for hyperuricemia include decreased glomerular filtration rate (GFR), diuretic use, and preexistent history of hyperuricemia. The influence of hyperuricemia in patient and graft survival is unclear because uric acid is not usually considered a common risk factor for cardiovascular disease that affects graft and patient survival. However, there have been small studies that have suggested that control of uric acid levels contributes to recovery of renal function (in heart and liver transplant recipients) and in an improvement in GFR in renal transplant recipients. Despite controversies in the need for hyperuricemia treatment in transplant patients, strategies to decrease uric acid levels includes a decrease or avoidance of cyclosporine treatment, adequacy of antihypertension treatment, avoidance of diuretics, nutritional management, and use of uric acid-decreasing agents. In this article we review the incidence and risk factors for the development of posttransplant hyperuricemia, discuss the influence of different immunosuppressive agents on uric acid metabolism, and suggest some alternative treatments for posttransplant hyperuricemia. We also consider that uric acid should be considered as a potential risk factor for renal allograft nephropathy or for renal dysfunction in nonrenal transplant recipients, as well as a comorbid factor for a decrease in patient and graft survival. PMID- 15660336 TI - Uric acid and preeclampsia. AB - Increased uric acid level is a key clinical feature of preeclampsia; higher levels correlate with significant maternal and fetal morbidity and mortality. The cause of hyperuricemia and its specific role in the pathogenesis of preeclampsia, however, remain unclear. Although uric acid has been shown to roughly parallel the severity of the maternal syndrome, it has not been useful in predicting the development of preeclampsia. Nevertheless, there have been recent data supporting a pathogenic role potentially in the hypertension and endothelial cell dysfunction of preeclampsia. This article reviews our current understanding of hyperuricemia in the setting of preeclampsia, and highlights the hypothesis that hyperuricemia may contribute to vascular damage in preeclampsia. PMID- 15660337 TI - Uric acid in chronic heart failure. AB - The pathophysiologic understanding of chronic heart failure (CHF) has shifted from a mere hemodynamic disorder to a much more complex approach including changes and imbalances in neurohormonal, immune, and metabolic functions. Among metabolic abnormalities, hyperuricemia is a constant finding in CHF. The xanthine oxidase metabolic pathway increasingly is appreciated as an important contributor to both symptoms of CHF as well as progression of the disease. Recent data suggest hyperuricemia to be an independent marker of impaired prognosis in CHF. In this article, the significance of the xanthine oxidase metabolic pathway in CHF is discussed. Data on xanthine oxidase inhibition are reviewed, which suggest a beneficial effect of therapeutically targeting this enzymatic pathway. PMID- 15660339 TI - The Journal of Electrocardiology and the 21st century electrocardiogram. PMID- 15660340 TI - Modern morphologic electrocardiographic interpretation--a valuable tool for rapid clinical decision making in acute ischemic coronary syndromes. PMID- 15660341 TI - A farewell to QT dispersion. Are the alternatives any better? PMID- 15660342 TI - Drug-induced QT dispersion: does it predict the risk of torsade de pointes? AB - Drug-induced delay in ventricular repolarization and proarrhythmias have attracted considerable regulatory attention. The measure of delayed ventricular repolarization most frequently used clinically is the ability of the new chemical entity (NCE) to prolong the QTc interval on surface electrocardiogram. Before they can be approved, new chemical entities with systemic bioavailability require characterization for their potential to prolong the QTc interval. Inevitably, QTc interval prolongation has come to be recognized as a surrogate marker of the risk of torsade de pointes (TdP)--a unique form of potentially fatal polymorphic ventricular tachycardia. Although it is the best and the simplest clinical measure that is available at present, QTc interval is not a reliable surrogate of TdP. Intramyocardial dispersion of repolarization appears to play a more important role both in electrical stability of the ventricles and in arrhythmogenesis. The potential importance of myocardial dispersion of refractoriness in arrhythmogenesis has led to a number of attempts to assess it from the surface electrocardiogram. This review summarizes the evidence for and against the predictive value of one of these attempts-measurement of the so called QT dispersion. Although the concept of QT dispersion is the best known and most widely investigated, it has also proved to be the least successful in predicting the risks of drug-induced TdP. PMID- 15660343 TI - QT dispersion failed to estimate the global dispersion of ventricular repolarization measured using monophasic action potential mapping technique in swine and patients. AB - The aim of this study was to evaluate whether the QT dispersion measured from 12 lead electrocardiogram (ECG) can estimate the global dispersion of ventricular repolarization (DVR) measured using a monophasic action potential (MAP) mapping technique. Monophasic action potentials were recorded from 75 +/- 12 left ventricular sites in 10 pigs and from 48 +/- 16 left or right ventricular sites in 15 patients using the CARTO mapping system. The maximum DVRs in both end-of repolarization and MAP duration among all the mapped sites were calculated and termed as global DVR for each measurement. QT intervals, QT peak and QT end , were measured from the 12-lead ECG, and QT dispersions; namely the differences between the maximum and the minimum of the QT peak and QT end were calculated. We found that QT dispersions were significantly smaller than (P < .05) and poorly correlated with the global DVRs both in pigs and patients. Bland-Altman agreement analysis demonstrated a marked variation of the differences and an obvious lack of agreement between the results obtained using the ECG and the MAP methods. In our patients, the global DVR increased markedly during ventricular tachycardia as compared with that during sinus rhythm (P < .05), whereas there was no significant difference in QT dispersion between these 2 subgroups. In conclusion, QT dispersion on the surface ECG could not estimate the global DVR measured using the MAP mapping technique. These findings are not consistent with some previously reported observations, suggesting the need for reappraisal of the electrophysiological implications of QT dispersion. PMID- 15660344 TI - Activation recovery time measurements in evaluation of global sequence and dispersion of ventricular repolarization. AB - INTRODUCTION: Activation recovery time (ART), defined as the time from the earliest ventricular activation time to the end of T wave on unipolar electrograms, has been used as an index of myocardial repolarization time. However, it is unknown whether the ART can be used to estimate the global sequence and dispersion of ventricular repolarization as determined by the monophasic action potential (MAP) mapping technique. METHODS AND RESULTS: Endocardial MAPs and unipolar electrograms were simultaneously recorded using the CARTO system from 34 +/- 12 left (n = 6) or right (n = 9) ventricular sites in 12 patients. End-of-repolarization (EOR) times from the MAPs and ARTs from the unipolar electrograms were calculated, based on which 15 sets of 3-dimensional maps of global EOR sequence and ART sequence were reconstructed. The ART sequence was consistent with the EOR sequence in 14 of 15 maps. In the 473 paired measurements obtained, the differences between the ART and the EOR time were 2 +/ 22 milliseconds (NS). A significant positive correlation between the ART and the EOR time was found in all the maps (r = 0.58 +/- 0.22). Agreement analyses showed that the differences between these 2 measurements were almost all within the range of mean difference +/- 2 SD for each individual map and for all the 473 recordings. The global dispersion of ART was 79 +/- 35 milliseconds, as compared with that of EOR time of 78 +/- 35 milliseconds (NS). CONCLUSION: The ART from unipolar electrograms is a good estimate of EOR time measured from MAPs, suggesting the usefulness of the former in evaluation of global sequence and dispersion of ventricular repolarization. PMID- 15660345 TI - Circadian rhythm of the QT interval dispersion in healthy subjects. Correlation with heart rate variability circadian pattern. AB - Circadian variation of QT interval dispersion (QTd) and heart rate variability spectral indices was evaluated in healthy persons in 24-hour 3-lead electrocardiogram. Mean values, SD, and SD/mean were evaluated for 24 hours, each hour separately and in night, day, and morning periods. Table Curve 2D and multiple regression were applied to find correlations between parameters. In 50% of subjects, a significant negative correlation was revealed between QTd and HF. Also, in 50% of persons, a significant positive correlation was found between QTd and low frequency/high frequency. After adjustment for periods, correlations were only observed during morning hours. With Table Curve 2D, 2 models of correlations between QTd and HF were found. Multiple regression analysis revealed relations between mean QTd and R-R as well as mean QTd and HF. It is possible that it is sympathovagal balance, as reflected in heart rate variability, and not the tone of both autonomic components that affects QTd variability. PMID- 15660346 TI - Increased P-wave dispersion predicts recurrent atrial fibrillation after cardioversion. AB - Atrial fibrillation (AF) will recur in a number of patients treated with cardioversion. Being able to identify reliable risk factors would be useful for making management decisions. P-wave dispersion (PWD) is an electrocardiographic measurement, which reflects a disparity in atrial conduction. P-wave dispersion has been shown to be increased in patients with paroxysmal AF. This pilot study aims to determine the role of PWD in predicting AF recurrence in patients who underwent elective cardioversion. Forty-five patients who were successfully cardioverted for persistent AF were included for study. Eighteen patients had a PWD greater than 80 ms; of these 13 had AF recurrence. Of the 25 patients with PWD less than 80 milliseconds, 12 had recurrent AF. P-wave dispersion values greater than 80 milliseconds were found more frequently in patients with AF recurrence (P = .05), supporting the use of this parameter in predicting return of AF. Larger studies are needed for further evaluation. PMID- 15660347 TI - Different methods of heart rate variability analysis reveal different correlations of heart rate variability spectrum with average heart rate. AB - Heart rate variability (HRV) spectrum can be calculated from the R-R or the sequence of instantaneous heart rates (IHRs). Because these signals are reciprocal, their use in the determination of the correlation of HRV with average HR may yield opposite results. AIM: The aim of this study is to reveal how HRV parameters correlate with HR depending on the kind of signal used and whether the normalization procedure or use of corrected signals affect the correlations. METHODS: Electrocardiographic recordings (512 beats) of 55 patients were considered in the analysis. RR and IHRs were calculated. Both signals were divided by their average values yielding the corrected RR and IHR. Heart rate variability spectra were estimated from these 4 kinds of signals. Total power (TP), low frequency (LF) and high frequency (HF) components, LF/HF ratio, and normalized values (ie, nLF, nHF) were calculated. RESULTS: Total power and LF estimated from RR correlated negatively with HR, but those calculated from IHR revealed a trend toward a positive correlation (respective correlation coefficients significantly differed, P < .01). The sign of correlation between HF and HR was the same regardless of the type of signal applied. The application of corrected signals made the results almost identical despite different signal origins (RR or IHR). The correlations of LF/HF, nLF, and nHF with HR were similar in all cases. CONCLUSIONS: The type of signal determines the sign of correlation among TP, LF, and HR. The parameters obtained from corrected signals, normalized quantities, and LF/HF reveal a consistent relationship with HR. PMID- 15660348 TI - On the mechanism of augmentation of electrocardiogram QRS complexes in patients with congestive heart failure responding to diuresis. AB - Patients with congestive heart failure (CHF) responding to diuresis reveal marked augmentation of the QRS complexes (AUG-QRS) in their electrocardiograms (ECGs). Recently, such change in the ECG has been observed in patients with anasarca (AN) of varying etiology commensurate with partial alleviation of the volume overload; similar ECG change has been noted in patients with end-stage renal failure after hemodialysis. The mechanism for the AUG-QRS in patients with CHF has been debated, and many have ascribed this ECG change to the "Brody effect," linking the AUG-QRS to reduction of intracardiac blood volumes resulting from diuresis. However, the Brody effect (a theoretical formulation not fully validated by experimentation and associated with controversy in its clinical implementation) has not provided a satisfactory explanation for the AUG-QRS in patients with CHF. In contrast, the described association between amelioration of AN in a diverse patient population and AUG-QRS suggests that this ECG change in patients with CHF is due to an increased electrical resistance of the passive body volume conductor, resulting from water loss effected by diuresis. This thesis is supported by theoretical work, animal experimentation, and clinical evidence. PMID- 15660349 TI - Distance correction in the electrocardiographic estimation of left ventricular mass. AB - Assessment of the left ventricular mass (LVM) from electrocardiograms may be improved by the addition of clinical variables into a multivariate equation. As the heart-thorax distance may affect the results, its relationships with electrocardiographic and clinical data have been evaluated in a group of 220 subjects (53 +/- 15 years, 126 female, 175 without demonstrated heart disease) who were assessed for echocardiographic LVM and heart-thorax distance. Sokolow, Cornell, and total QRS voltage indexes were obtained. Multiple regression equations with LVM as the dependent variable were fit, with an ECG index, body mass index (BMI), age, and gender as the independent predictors. Each of the 3 ECG indexes, BMI, age, and sex was shown to be independent predictors of LVM, with the ECG and BMI contributing with most of the explanatory power. When added to the model, the distance from the interventricular septum to the precordium (septal-LVD) was not a predictor of LVM, but when BMI was withdrawn, septal-LVD became an independent predictor of LVM (P < .001). This was not observed when septal-LVD was substituted for any other clinical or ECG variable, thus suggesting that septal-LVD accounts for information contained in BMI but not in the remaining variables. In addition, the distance from the center of LV to the precordium (mid-LVD) achieved significance as an independent LVM predictor, although the coefficient of multiple determination (R) practically did not change. Almost identical results are obtained when LVM is indexed for body surface area. Body mass index supplies virtually all the information contained in the heart-thorax distance. PMID- 15660350 TI - Prognostic value of ambulatory electrocardiography monitoring in patients with dilated cardiomyopathy. AB - BACKGROUND: Performance of ambulatory electrocardiography (AECG) may provide data useful for counseling patients regarding prognosis and for selecting potential patients for defibrillator implantation, but this practice remains controversial. METHODS: We reviewed clinical and AECG data on 355 patients diagnosed with dilated cardiomyopathy (DCM). Predictors of survival were identified in a multivariable analysis using a Cox proportional hazard model. Ability of the derived model to predict outcomes was tested using a second cohort of 144 patients. RESULTS: Nonsustained ventricular tachycardia (NSVT) was present on AECG in 31% of the subjects. Ambulatory electrocardiography provided 3 independent predictors of mortality: NSVT (relative risk [RR], 1.63; 95% confidence interval [CI], 1.06-2.51; P = .02), mean heart rate (RR, 1.03; 95% CI, 1.02-1.04; P = .0001), and heart rate range (RR, 0.990; 95% CI, 0.982-0.997; P = .008). Performance of the multivariable model was validated (area under the curve = 0.83) on a second cohort of patients. CONCLUSION: Ambulatory electrocardiography provides useful prognostic information in patients with DCM, identifying 3 independent predictors of mortality. PMID- 15660351 TI - Characteristics and radiofrequency ablation in posteroseptal and left free-wall subepicardial accessory pathways. AB - Accessory pathways (APs) that can only be ablated from the coronary sinus are likely to be located subepicardially. The electrocardiographic (ECG) and electrophysiological characteristics as well as the immediate radiofrequency ablation success rate and the recurrence rate were compared in 15 patients (11 posteroseptal and 4 left free-wall) with subepicardial APs and in 31 control patients with posteroseptal (15) and left free-wall (16) APs matched with age, sex, and AP location during the same study period in whom APs were successfully ablated from the endocardial approach. Patients with posteroseptal subepicardial APs had a longer tachycardia cycle length (355 +/- 32 vs 286 +/- 49 milliseconds, P < .05), a lower success rate (9 /11 vs 15/15, P = .09), and a higher recurrence rate (3/9 vs 0/15, P < .05) as compared with control patients. A negative delta wave with QS or QR pattern in lead II was present in all 4 patients with a manifest posteroseptal subepicardial AP located in the middle cardiac vein as compared with none of the 5 control patients with posteroseptal APs located in the proximal coronary sinus and 1 of the 9 control patients (P < .01). A positive delta wave in lead I along with an R/S of less than 1 in lead V 1 , and a negative delta wave in lead II, was noted in 1 of the 2 patients with left free wall subepicardial APs and none of the 7 controls (P = .047). The local activation time is significantly shorter in the 4 patients with left free-wall subepicardial AP than in the 16 control patients (31 +/- 9 vs 89 +/- milliseconds, P = .044). CONCLUSIONS: Some ECG characteristics are suggestive of APs located in the middle cardiac vein and left free-wall subepicardial site, while a longer local activation time is characteristic of left free-wall APs. The success rate is lower and the recurrence rate higher with radiofrequency ablation in patients with subepicardial AP. PMID- 15660353 TI - Autonomic antecedents to variant angina exacerbation after beta-blockade withdrawal. AB - We describe a patient with nonsignificant coronary artery disease who experienced variant angina after beta -blockade withdrawal. Standard therapy with nifedipine and nitrates aimed at suppressing symptoms and typical transient ST-segment elevations was superseded by the reinstitution of metoprolol. The autonomic alternations before and after readministration of metoprolol were analyzed by time and spectral indices of heart rate variability (HRV). Metoprolol reduced the HRV and reversed the low-frequency/high-frequency power ratio toward a more physiological autonomic balance. We conclude that the reinstitution of beta blocker acted protectively by preventing surges of sympathetic activity on an underlying basis of parasympathetic predominance. PMID- 15660352 TI - Electrocardiographic changes after head trauma. AB - We report the case of a patient who developed, a few days after a closed head injury, marked electrocardiographic changes mimicking an acute coronary event, in the absence of actual cardiac damage. The electrocardiographic changes were fully reversible, paralleling the neurologic status. Neuroimaging examinations excluded subarachnoid hemorrhage or space-occupying hematoma, but demonstrated diffuse axonal injury using susceptibility-weighted magnetic resonance techniques. This kind of traumatic brain injury thus may be responsible for a pseudo-acute myocardial ischemic syndrome. PMID- 15660354 TI - Controllable growth of chains and grids from polyoxomolybdate building blocks linked by silver(I) dimers. AB - Molecular growth processes utilizing a beta-octamolybdate synthon and {Ag2} dimers are described and the directing influence of "encapsulating" cations and coordinating solvent is also demonstrated. The growth of two 1D chains, (nBu4N)2n[Ag2Mo8O26]n (1) and (nBu4N)2n[Ag2Mo8O26(CH3CN)2]n (2), is achieved when nBu4N+ ions are used, and the diameter of the chains can be expanded by the coordination of CH3CN solvent (2). The formation of a type of gridlike structure in which 1D chains are crossed-over each other in alternatively packed layers is achieved in DMSO as the solvent; DMSO acts as a linking group to give (nBu4N)2n[Ag2Mo8O26(dmso)2]n (3), which, similar to 1 and 2, still incorporates the Bu4N+ ions that exert an "encapsulating" influence. However, in (HDMF)n[Ag3(Mo8O26)(dmf)4]n (4) the relatively bulky Bu4N+ ions are exchanged for protonated DMF cations, thereby allowing the chains to condense to a 2D array. The building block concept is further enforced by the isolation of a "monomeric" unit (Ph4P)2[Ag2Mo8O26(dmso)4] (5), which is isolated when the Ph4P+ ions are so "encapsulating" as to prevent aggregation of the {Ag-Mo8-Ag} building blocks. The nature of the AgAg dimers in each of the compounds 1-4 is examined by DFT calculations and the interplay between these Ag-Ag interactions and the structure types is described. PMID- 15660355 TI - Development of PPO inhibitor-resistant cultures and crops. AB - Recent progress in the development of protoporphyrinogen oxidase (PPO, Protox) inhibitor-resistant plant cell cultures and crops is reviewed, with emphasis on the molecular and cellular aspects of this topic. PPO herbicide-resistant maize plants have been reported, along with the isolation of plant PPO genes and the isolation of herbicide-resistant mutants. At the same time, PPO inhibitor resistant rice plants have been developed by expression of the Bacillus subtilis PPO gene via targeting the gene into either chloroplast or cytoplasm. Other attempts to develop PPO herbicide-resistant plants include conventional tissue culture methods, expression of modified co-factors of the protoporphyrin IX binding subunit proteins, over-expression of wild-type plant PPO gene, and engineering of P-450 monooxygenases to degrade the PPO inhibitor. PMID- 15660356 TI - Herbicide resistance in transgenic plants with mammalian P450 monooxygenase genes. AB - Transgenic potato and rice plants were generated by the introduction of human P450 species, CYP1A1, CYP2B6, CYP2C9 and CYP2C19, which metabolized a number of herbicides, insecticides and industrial chemicals. The transgenic potato plant T1977 co-expressing CYP1A1, CYP2B6 and CYP2C19 genes showed remarkable cross resistance to several herbicides with different structures and modes of action due to metabolism of these herbicides by the P450 species expressed. The transgenic rice plant 2C9-57R2 expressing CYP2C9 gene showed resistance to sulfonylureas, and the transgenic rice plant 2C19-12R1 expressing CYP2C19 gene showed cross-resistance to certain herbicides with different structures and modes of action. These transgenic plants appear to be useful for herbicide resistance as well as phytoremediation of environmental contaminants. PMID- 15660357 TI - Insect saliva: an integrative approach. PMID- 15660358 TI - The role of salivary lipocalins in blood feeding by Rhodnius prolixus. AB - In order to overcome host mechanisms that prevent blood loss, the blood-sucking bug Rhodnius prolixus has evolved a complex salivary secretion containing dozens of different proteins. A number of these have been characterized and found to have roles in inhibiting various hemostatic or inflammatory systems. Interestingly, many of these biologically active salivary proteins belong to the lipocalin protein family. A proliferation of lipocalin genes has occurred via gene duplication and subsequent divergence. Functional genomic, proteomic, and functional studies have been performed to probe the role of salivary lipocalins in blood feeding. In the course of these investigations, anticoagulant, antiplatelet, antiinflammatory, and vasodilatory molecules have been described. PMID- 15660359 TI - The host plant as a factor in the synthesis and secretion of salivary glucose oxidase in larval Helicoverpa zea. AB - We investigated the effect of the host plant on the synthesis and secretion of the elicitor glucose oxidase in the salivary glands of larval Helicoverpa zea. Glucose oxidase catalyses the oxidation of d-glucose to produce d-gluconic acid and hydrogen peroxide. Previous studies have found that the product hydrogen peroxide is primarily responsible for suppressing the wound-inducible defenses of the host plant. Using an antibody specific for glucose oxidase, we determined the effect of the host plant on the rate of secretion of glucose oxidase. Larval H. zea secrete microgram amounts of the enzyme glucose oxidase from their principal salivary glands, the labial glands. Larvae reared on different host plants produce varying amounts of glucose oxidase in their labial glands. We used a tissue printing procedure with our antibody to determine if larvae secrete glucose oxidase directly at the feeding or wound sites. Significant amounts of the enzyme are deposited at the feeding site, although some is deposited outside the feeding margins. PMID- 15660360 TI - Evidence that caterpillar labial saliva suppresses infectivity of potential bacterial pathogens. AB - Salivary enzyme, glucose oxidase (GOX) from the caterpillar Helicoverpa zea, catalyzes the conversion of glucose to gluconic acid and hydrogen peroxide. Because hydrogen peroxide has well-known antimicrobial properties, we examined whether caterpillar labial saliva could reduce the infectivity of bacterial pathogens. We examined the effects of caterpillar saliva on the growth of two bacteria species Serratia marcescens and Pseudomonas aeruginosa. Wells formed in LB agar contained a solution of salivary gland extract (Sx) and glucose, GOX and glucose, Sx only, GOX only, or glucose only. After 18 h of incubation, the diameter of cleared bacteria was measured. Wells treated with only GOX, Sx, or glucose showed no measurable area of clearing, while wells treated with GOX with glucose or Sx with glucose had considerable clearing. To determine if saliva could provide protection to caterpillars in vivo, a surgery was performed on caterpillars that prevented the secretion of labial saliva. Caterpillars were fed a diet containing either no added bacteria or treated with high levels of S. marcescens or P. aeruginosa. Caterpillars that could not secrete saliva had significantly higher levels of mortality when feeding on diet treated with either bacterium than caterpillars that could secrete saliva when feeding on equal levels of bacteria-treated diet. Our evidence demonstrates for the first time that insect saliva in situ can provide protection against bacterial pathogens and that the salivary enzyme GOX appears to provide the antimicrobial properties. PMID- 15660361 TI - Biosynthesis of fatty acid amide elicitors of plant volatiles by insect herbivores. AB - Larvae of several species of Lepidoptera produce fatty acid amide elicitors that induce the plants on which they feed to synthesize and release volatile organic compounds. The volatiles released by the plants act as cues that aid in host location by natural enemies of the herbivorous larvae. The elicitors are synthesized in the larvae by enzymes embedded in the membranes of the crop and anterior midgut tissues. The fatty acid precursors of the elicitors are obtained from the plants on which the caterpillars feed, while the amino acid moieties appear to be obtained from pools within the insects. The fatty acid amide elicitors are rapidly hydrolyzed in the midgut and hindgut by enzymes in the gut lumen. The role of these fatty acid amides in caterpillar metabolism is not yet understood. PMID- 15660362 TI - Lepidopteran herbivory and oral factors induce transcripts encoding novel terpene synthases in Medicago truncatula. AB - Terpenes are an important class of defense compounds that accumulate in plants after pathogen infection or arthropod injury. Sequences predicted to encode terpene synthases were selected from an expressed sequence tag (EST) database of Medicago truncatula. Four putative terpene synthase clones (MtTps1-MtTps4), originating from a chewing insect-damaged M. truncatula leaf cDNA library, were isolated. Transcript levels of each gene examined increased in response to artificial wounding, Spodoptera exigua herbivory, and treatment with volatile methyl jasmonate (meJA). Addition of S. exigua regurgitant to wound sites triggered transcript accumulation of MtTps1 and levels increased with higher concentrations of regurgitant. Furthermore, induction of MtTps1 occurred after application of N-linolenoyl-glutamate or N-linoleoyl-glutamate, factors found in lepidopteran regurgitant. Genomic DNA blots indicate that each of the putative proteins is encoded by a single-copy gene or a small gene family. Proteins encoded by MtTps3 and MtTps4 are imported into the soluble fraction of chloroplasts in in vitro assays, whereas proteins encoded by MtTps1 and MtTps2 are not imported into chloroplasts. Combined with sequence comparisons of multiple plant terpene synthases, the import data indicate that MtTps1 and MtTps2 likely encode sesquiterpene synthases and that MtTps3 and MtTps4 encode mono- or di-terpene synthases. In addition to serving as a valuable model legume species for genomic studies, M. truncatula should prove a valuable source of novel terpene-producing enzymes. Induction of wound-responsive genes by insect oral factors suggests that M. truncatula senses biotic damage through the presence of elicitors originating in the herbivore. PMID- 15660363 TI - Evidence that the caterpillar salivary enzyme glucose oxidase provides herbivore offense in solanaceous plants. AB - The insect salivary enzyme glucose oxidase (GOX) can inhibit wound-inducible nicotine production in tobacco, Nicotiana tabacum. We examined whether salivary gland extracts of Helicoverpa zea lacking active GOX could still suppress nicotine in tobacco, Nicotiana tabacum, and whether GOX could suppress wound inducible defenses of another Solanaceous plant, tomato Lycopersicon esculentum. Tobacco leaves were wounded with a cork borer and treated with water, salivary gland extracts with active GOX (SxG), or salivary gland extracts with inactive GOX (SxI). After three days, leaves treated with SxG had significantly less nicotine than all other wounded treatments. Neonates that fed on the terminal leaves of tobacco plants treated with SxG had significantly higher survival than neonates that fed on leaves treated with either SxI or water. This evidence supports the assertion that GOX is the salivary factor responsible for the suppression of tobacco plant nicotine production by H. zea saliva. Results for the NahG tobacco plants, which lack salicylic acid (SA) due to a transgene for bacterial SA hydroxylase, indicate that suppression of nicotine by GOX does not require SA. However, tobacco leaves that were wounded and treated with SxG had significantly higher levels of the SA-mediated PR-1a protein than leaves treated with SxI or water. Leaves of tomato plants wounded with scissors and then treated with SxG had trypsin inhibitor levels that were moderately lower than plants wounded and treated with purified GOX, water, or SxI. However, all the wounded tomato leaves irrespective of treatment resulted in lower caterpillar growth rates than the non-wounded tomato leaves. Glucose oxidase is the first insect salivary enzyme shown to suppress wound-inducible herbivore defenses of plants. PMID- 15660364 TI - Micro-injection of Lygus salivary gland proteins to simulate feeding damage in alfalfa and cotton flowers. AB - Alfalfa and cotton flowers were pierced with small glass capillaries of an overall size and shape similar to that of Lygus stylets, and injected with small quantities (6 to 100 nL) of solutions that contained Lygus salivary enzymes. Crude and partially purified protein solutions from Lygus heads and isolated salivary glands showed substantial polygalacturonase (PG) activity, as has been previously reported. Following injection with both crude and partially purified protein solutions, as well as with pure fungal and bacterial PGs, flowers of both alfalfa and cotton exhibited damage similar to that caused by Lygus feeding. Injection with the same volume of a buffer control as well as a buffer control containing BSA at a comparable protein concentration (approximately 6 microg/mL) showed no symptoms. These results are consistent with a previously suggested hypothesis that the extensive tissue damage caused by Lygus feeding is primarily due to the action of the PG enzyme on the host tissue, rather than to mechanical damage caused by the insect stylet. Substantial genotypic variation for a PG inhibiting protein (PGIP) exists in alfalfa and cotton. We, therefore, suggest that breeding and selection for increased native PGIP levels, or transformation with genes encoding PGIP from other plant species, may be of value in obtaining alfalfa and cotton varieties that are more resistant to Lygus feeding damage. PMID- 15660365 TI - The piercing-sucking herbivores Lygus hesperus and Nezara viridula induce volatile emissions in plants. AB - Plant volatiles induced by herbivory are often used as olfactory cues by foraging herbivores and their natural enemies, and thus have potential for control of agricultural pests. Compared to chewing insects and mites, little is known about plant volatile production following herbivory by insects with piercing-sucking mouthparts. Here, we studied factors (insect life stage, gender, the role of salivary glands, and type of bioassay used for volatile induction) that influence the induction of plant volatiles by two agriculturally important hemipterans, Lygus hesperus and Nezara viridula. Feeding on intact cotton by virgin females of L. hesperus induced 2.6-fold greater volatile response compared to that induced by mated females, possibly due to increased feeding activity by virgin females. This plant volatile response was associated with elicitors present in the insect's salivary glands as well as to the degree of mechanical injury. Feeding injury by N. viridula females also increased volatile emissions in intact maize by approximately 2-fold compared to control plants. Maize seedlings injured by N. viridula emitted higher amounts of the monoterpene linalool, the sesquiterpenes (E)-beta-caryophyllene, alpha-trans-bergamotene, and (E,E)-beta-farnesene, and the homoterpene (E,E)-4,8,12-trimethyl-1,3,7,11-tridecatetraene, but not amounts of green leaf volatiles, compared to uninjured plants. Emissions from intact maize injured by adult males were lower than those emitted by adult females of the same age and did not differ from those emitted by uninjured plants. Similarly, feeding by virgin female N. viridula followed by excision led to 64% higher quantities of volatiles compared to untreated plants. Volatile emission in excised plants, however, was considerably greater than in intact plants, suggesting that careful consideration must be given to bioassay design in studies of herbivore-induced plant volatiles. Salivary gland extracts of N. viridula led to sesquiterpene emissions approximately 2.5-fold higher than for controls, although no significant differences were observed for green leaf volatiles, monoterpenes, and homoterpenes. These results indicate that L. hesperus and female N. viridula feeding induce volatile production in plants, and that volatile production is affected by gender and life stage of the bug. Although oviposition and mechanical injury by stylets may increase release of volatiles, elicitors from salivary glands of L. hesperus and N. viridula also seem to play a role in the emission of plant volatiles. PMID- 15660366 TI - Right colectomy with isoperistaltic side-to-side stapled ileocolic anastomosis. AB - We describe here a novel technique of isoperistaltic side-to-side ileocolic anastomosis following right hemicolectomy with minimal use of linear cutter (GIA 80) in 16 of our patients. Compared to the traditional standard technique, we found our new technique to be simple, safe, and cost-effective. It probably also helped in faster and uncomplicated recovery of our patients. PMID- 15660368 TI - Laparoscopic resection of a huge intraluminal gastric submucosal tumor located in the anterior wall: eversion method. AB - Although numerous operative approaches have been utilized for gastric submucosal tumors, laparoscopic wedge resection has been regarded as the treatment of choice in recent years. As widespread use of diagnostic endoscopy has exposed a number of gastric submucosal tumors, the laparoscopic wedge resections are being performed with increased frequency. Many reports have been published which describe the technique of laparoscopic wedge resection of gastric submucosal tumors, and by far most of them were about the techniques for masses located at the posterior wall or esophagogastric junction. Generally, wedge resection for tumors on the anterior wall is regarded as an easy procedure especially when the mass is extraluminal mass. However, when the tumor is huge and intraluminal, it is very difficult to resect without compromising the gastric lumen as wedge resection of a huge intraluminal mass in situ inevitably includes a wide portion of normal gastric wall. In this article, we describe a successfully performed laparoscopic wedge resection of a huge intraluminal gastric submucosal tumor while preserving the volume of residual stomach without compromising the gastric lumen using the Eversion method through gastrotomy made with laparoscopic ultrasound guidance. PMID- 15660369 TI - Simple technique of securing intraoral skin grafts. AB - Small defects following intraoral resection are often resurfaced by skin grafts. Skin grafting has the advantage of ease of harvest with minimal additional operating time and post-operative hospital stay, an acceptable functional cosmetic result, and the ability to survive post-operative radiation 1. In addition to adequate vascularity of the recipient area, the most important aspect for graft survival is immobilization and adherence of the graft to the defect. However, in the oral cavity due to the uneven wound bed and constant mobility of the cheek, the graft is not completely immobilized. In addition, the salivary secretions tend to accumulate beneath the graft, separating the graft from the bed. Graft-failure can be prevented by immobilizing the graft and closing up any potential dead space that might lead to separation 2. A variety of methods have been described for immobilization and bolstering the graft to the wound. Many types of stents have been used varying from the simple cotton balls, resin molds, and foam pads, to complex stents like metal, plastic, and dental liner 34. The traditional tie over bolster technique described by Schramm and Myers involves fixation of the skin graft to the raw area, followed by placement of non absorbable silk sutures from the adjacent mucosa, which are then tied over the bolster 1. However, the placement of this tie over sutures requires adjacent normal mucosa for anchorage, which may not be sufficient especially in the gingivo-buccal sulcus. Although external fixation of the stents to the cheek has been described, this results in ugly scarring of the cheek 2. We describe a simple technique of fixation of the skin graft in the oral cavity, which avoids the placement of additional tie over sutures and in our opinion results in better anchorage. PMID- 15660367 TI - Characterization of zebrafish PSD-95 gene family members. AB - The PSD-95 family of membrane- associated guanylate kinases (MAGUKs) are thought to act as molecular scaffolds that regulate the assembly and function of the multiprotein signaling complex found at the postsynaptic density of excitatory synapses. Genetic analysis of PSD-95 family members in the mammalian nervous system has so far been difficult, but the zebrafish is emerging as an ideal vertebrate system for studying the role of particular genes in the developing and mature nervous system. Here we describe the cloning of the zebrafish orthologs of PSD-95, PSD-93, and two isoforms of SAP-97. Using in situ hybridization analysis we show that these zebrafish MAGUKs have overlapping but distinct patterns of expression in the developing nervous system and craniofacial skeleton. Using a pan-MAGUK antibody we show that MAGUK proteins localize to neurons within the developing hindbrain, cerebellum, visual and olfactory systems, and to skin epithelial cells. In the olfactory and visual systems MAGUK proteins are expressed strongly in synaptic regions, and the onset of expression in these areas coincides with periods of synapse formation. These data are consistent with the idea that PSD-95 family members are involved in synapse assembly and function, and provide a platform for future functional studies in vivo in a highly tractable model organism. PMID- 15660370 TI - Full-term gestation and transvaginal delivery after wide resection of an abdominal desmoid tumor during pregnancy. AB - A 29-year-old female was diagnosed with a symptomatic, extra-abdominal desmoid tumor during the first trimester of pregnancy. Computerized tomography (CT) and transabdominal ultrasound (US) noted a mass within the left rectus sheath measuring up to 15 cm in greatest diameter, with mild compression of the uterus. Preoperative diagnosis was confirmed by core-needle biopsy of the lesion. At 20 weeks gestation, wide local resection of the tumor with disease-free margins, as well as abdominal wall reconstruction with polytetrafluoroethylene (PTFE) mesh was successfully undertaken. Histological examination of the tumor ex vivo confirmed that the lesion was a desmoid tumor consisting of spindle cells with dense infiltrating collagenous fibers. Subsequent to her resection, the patient completed a full-term pregnancy without complication, and proceeded with a complication-free transvaginal delivery at 39 weeks. This case illustrates the probable contribution of estrogens towards desmoid tumor development, the durability of abdominal wall reconstruction when subjected to the extraordinary strain of both a gravid uterus and labor, as well as the safety and efficacy of aggressive surgical therapy during pregnancy. PMID- 15660371 TI - Tikhoff-Linberg procedure and chest wall resection for recurrent sarcoma of the shoulder girdle involving the chest wall. AB - Since the early 1980s, limb-sparing procedures have become the standard of care for the treatment of sarcoma involving the extremities. The Tikhoff-Linberg procedure has been described as a limb-sparing option for the treatment of neoplasms of the shoulder girdle. Historically, the involvement of the chest wall with tumor has been considered an absolute contraindication for this operation. We describe a case in which a recurrent sarcoma of the shoulder girdle with chest wall involvement was successfully treated with the Tikhoff-Linberg procedure modified to include chest wall resection, along with a simple reconstructive technique. PMID- 15660373 TI - Protease-activated receptor-2 regulates cell proliferation and enhances cyclooxygenase-2 mRNA expression in human pancreatic cancer cells. AB - BACKGROUND AND OBJECTIVES: Protease-activated receptor-2 (PAR-2) is a G protein coupled receptor that is activated by trypsin. Recent studies have suggested that PAR-2 activity correlates with inflammatory processes and cell proliferation and that PAR-2 activation in non-neoplastic cells induces expression of cyclooxygenase-2 (COX-2). In the present study, we examined whether PAR-2 activation regulates cell proliferation and COX-2 expression by pancreatic cancer cells. METHODS: We analyzed PAR-2 expression immunohistochemically in 40 intraductal papillary-mucinous neoplasms (IPMNs) and 73 invasive ductal carcinomas (IDCs) of the pancreas. We used four pancreatic cancer cell lines (Panc1, T3M4, BxPC3, and MIApaca2) to measure cell proliferation and COX-2 mRNA expression after PAR-2 activation. RESULTS: PAR-2 protein was detected immunohistochemically in 85.0% of IPMNs and 65.8% of IDCs. Trypsin and a PAR-2 agonist peptide, SLIGKV, stimulated proliferation of each cell line in a dose dependent manner. Exposure of cells to anti-PAR-2 neutralizing antibody prior to PAR-2 activation suppressed cell proliferation. In COX-2-positive cell lines (T3M4 and BxPC3), PAR-2 activation significantly increased COX-2 mRNA expression. CONCLUSIONS: Our results suggest that PAR-2 activation is associated with cell proliferation and COX-2 expression in pancreatic cancer cells. Blockade of the PAR-2 signaling pathway may be a novel strategy for suppressing pancreatic tumor growth. PMID- 15660374 TI - Decreasing role of lymphatic system surgery in surgical oncology. PMID- 15660372 TI - The role of liquid-based cytology in the investigation of breast lesions using fine-needle aspiration: a cytohistopathological evaluation. AB - INTRODUCTION: Liquid-based cytology processing of breast fine-needle aspiration (FNA) specimens has been used in our institution since 1999. The aim of this study was to investigate the role of liquid-based cytology (Thin-Prep(R)) in the evaluation of breast FNA specimens. PATIENTS AND METHODS: This study was carried out on 352 cases corresponding to equal number of women (134 malignant and 218 benign) examined from September 1999 to June 2003. A direct to vial study was planned. Aspirated material was directly immersed to cytolytR vial (Cytyc Corporation, Boxborough, MA). The specimens were transported to the laboratory non-refrigerated within 24 hr. Thin-Prep slides were made using the Thin-Prep 2000 processor. The remaining material was embedded in paraffin. RESULTS: Altogether, 352 cases were examined and unsatisfactory smears were observed in 14 (3.9 %) out of all cases. The histologic examination verified that 128 out of 131 total malignant cases and 203 out of 207 total benign cases were correctly diagnosed with the cytological technique. False positives were found in four cases and false negative in three cases. Liquid-based technique presented a sensitivity of 97.7%, a specificity of 98%, a positive predictive value (PPV) of 96.9%, a PNV of 98.5%, and an over all accuracy (OA) of 97.9%. CONCLUSIONS: The processing of FNA breast specimens using liquid-based cytology offers an accurate diagnostic tool. Liquid-based cytology could reduce the uncertainty based in the cytological evaluation of FNA, because it offers the possibility to embed material in paraffin and take tissue sections with histological appearance identical to those observed in routine histological sections. Moreover it offers the potential advantage of retraining extra material for the purpose of ploidy analysis and immunohistochemical study in order to offer information concerning the expression of prognostic markers such as p53, MIB-1, steroid hormone receptors, and EGFR. PMID- 15660375 TI - Clinical outcome of esophageal cancer patients with history of gastrectomy. AB - BACKGROUND: Surgery for thoracic esophageal cancer after gastrectomy involves a complicated reconstruction procedure. A surgeon's hesitation is further increased because the clinical outcome of surgical treatment of these patients has not been elucidated. OBJECTIVES: Among 948 thoracic esophageal cancer patients who underwent curative operation, 72 (7.6%) had a history of gastrectomy. Their clinico-pathological features and survival (follow-up average 881 days) were compared with those without gastrectomy. RESULTS: Esophagectomy for patients after gastrectomy was performed via right thoracotomy (66), left thoracotomy (4), and transhiatal resection (2), and reconstruction was done using the right-side colon (57) or jejunum (15). Compared to non-gastrectomized patients, gastrectomized patients were exposed to longer operation time (523 min vs. 460 min), but no significant difference was observed in operative mortality (4.2% vs. 2.5%) or blood loss (1,189 ml vs. 990 ml). Pathological examination showed no significant difference in depth of tumor invasion, lymph node metastasis, and TNM staging between gastrectomized and non-gastrectomized patients, while tumors were located at lower position in the gastrectomized patients (P = 0.046). The overall and cause-specific 5-year survival rates were 56% and 65% for gastrectomized esophageal cancer patients, which were significantly better than for non gastrectomized patients (36% and 44%, P = 0.0235 and 0.024, respectively). Multivariate analysis showed gastrectomy as a marginally independent factor for a favorable prognosis (hazard ratio 1.832, P = 0.0324). With respect to tumor recurrence, hematogenic metastasis tended to be less frequent in gastrectomized patients than in non-gastrectomized patients. In gastrectomized patients, neither disease (peptic ulcer or gastric cancer) nor reconstruction (Billroth-I, Billroth II, and Roux-Y) for gastrectomy affected the clinicopathological findings or post operable survival. CONCLUSIONS: Surgical treatment of esophageal cancer patients after gastrectomy was complicated but tolerable, and should be considered as a reliable therapeutic modality because of favorable patient prognosis. PMID- 15660376 TI - Nemaline rods in chorea-acanthocytosis. AB - In chorea-acanthocytosis, a neurological disorder associated with multisystem degeneration, amyotrophy and peripheral neuropathy are sometimes conspicuous. We describe a patient with chorea-acanthocytosis who showed distributed nemaline rods in biopsied muscle. It has been suggested that in chorea-acanthocytosis, the muscle membranous structures are disordered, and our finding may be attributable to this underlying myopathic condition. PMID- 15660377 TI - Lymph node staging surgery and breast cancer: potholes in the fast lane from more to less. PMID- 15660378 TI - Peer victimization and the development of obsessive-compulsive disorder in adolescence. AB - We describe an adolescent boy's experience of peer victimization and its relation with his development of obsessive-compulsive disorder (OCD). Subsequent to being peer victimized, this boy was seen for 20 sessions of cognitive-behavioral therapy over the course of 4 weeks that followed the protocol outlined by March and Mulle in 1998. Standardized post-treatment assessment indicated significant reductions in OCD, depressive, and anxious symptomatology as compared to baseline. This case illustrates how negative peer experiences may be related to the development and maintenance of OCD. PMID- 15660379 TI - Parallel synthesis and biological evaluation of different sizes of bicyclo[2,3] Leu-enkephalin analogues. AB - Parallel synthesis of peptides and peptidomimetics has been an important approach to search for biologically active ligands. A novel systematic synthesis of different size bicyclic dipeptide mimetics was developed on solid-phase supports. By taking advantage of the enantioselective synthesis of omega-unsaturated amino acids and their N-methylated derivatives, the hemiaminal problem was prevented in the pathway to thiazolidine formation. The bicyclic dipeptide was generated on the solid-phase support in three steps by an unconventional method. By inserting this bicyclic scaffold into the synthesis of a larger bioactive peptide, 11 different sizes of bicyclo[2,3]-Leu-enkephalin analogues were synthesized in a fast and efficient way. Modeling studies show that a reversed turn structure at positions 2-3 was favored when an L- and L-bicyclic scaffold was used, and that an extended conformation at the N-terminal was favored when a D- and L-bicyclic scaffold was inserted. Binding affinities and bioassay studies show ligands with micromolar binding affinities and antagonist bioactivities for the [6,5]- and [7,5]-bicyclo-Leu-enkephalin analogues. PMID- 15660380 TI - NMR structures and orientation of the fourth transmembrane domain of the rat divalent metal transporter (DMT1) with G185D mutation in SDS micelles. AB - DMT1, also known as Nramp2, is an iron transporter, and belongs to the family of Nramp proteins. Disease-causing mutations both in Nramp1 and Nramp2 occurring at the conserved two adjacent glycine residues located within the fourth transmembrane domain (TM4) suggest that TM4 may serve an important biological function. In the present study, we have determined the high-resolution structures of a synthetic peptide, corresponding to the sequence of the fourth transmembrane domain of rat DMT1 with G185D mutation, in membrane-mimetic environments (e.g., SDS micelles) using NMR spectroscopy and distance-geometry/simulated annealing calculations. The spatial structures showed alpha-helices without a kink in the middle portion of the peptide, with a highly flexible and poorly defined N terminus. Both the N-terminus and the helical core of the peptide were embedded into the SDS micelles. Interestingly, the folding and membrane location of the C terminus was pH dependent, being well-folded and inserted into SDS micelles only at a low pH value (4.0). The peptide exhibited amphipathic characteristics, with hydrophilic residues (Asp7, Thr11, Asp14, and Thr15) lying in one side of the helix, which provide a basis for the formation of water-filled channel architectures through self-associations. The significant broadening of the resonances of the hydrophilic residues Asp7, Thr11, and Asp14, which are buried inside SDS micelles, upon addition of Mn2+ further verified the possibility of the formation of a channel through which metal ions pass. The substitution of Gly7 by an aspartate residue neither significantly altered the structure and membrane location of the peptide nor abolished its properties of channel forming and metal permeation compared with the wild-type peptide. PMID- 15660381 TI - Small nonphosphorylated Grb2-SH2 domain antagonists evaluated by surface plasmon resonance technology. AB - The growth factor receptor-binding protein-Src homology 2 (Grb2-SH2) domain plays an important role in the oncogenic Ras signal transduction pathway, which involves cell proliferation and differentiation. Therefore, the Grb2-SH2 domain has been chosen as our target for development of potential antiproliferative agents. Herein, we report the study of the inhibitory effects of small nonphosphorylated peptide analogs interacting with the Grb2-SH2 domain protein by surface plasmon resonance (SPR) technology. A set of 8 related peptide analogs were synthesized, purified, and characterized. Their inhibitory effects on Grb2 SH2 were evaluated by the SPR technology developed with the BIACORE X instrument. The lead peptide, Fmoc-Glu-Tyr-Aib-Asn-NH2 (Fmoc-E-Y-Aib-N; Fmoc: 9 fluorenylmethyoxycarbonyl; Aib=alpha-amino isobutyric acid) inhibited Grb2-SH2 domain function with an IC50 value of 8.7 microM. A molecular modeling study of the lead peptide indicated that the glutamate in the Fmoc peptide is ideally positioned to form a strong salt bridge to Arg 67 in the Grb2-SH2 domain, using both its backbone carbonyl and its acidic group. Residue Glu 89 in Grb2-SH2 flips inward to fill the binding site and partially replace the phosphate group as a hydrogen-bond acceptor. Results of these studies provide important information for further development of potent nonphosphorylated peptide inhibitors of the Grb2-SH2 domain. PMID- 15660382 TI - Gefitinib, an EGFR inhibitor, prevents hepatocellular carcinoma development in the rat liver with cirrhosis. AB - Epidermal growth factor receptor (EGFR) binds transforming growth factor alpha (TGF-alpha) which is mitogenic for hepatocytes. Diverse lines of evidence suggest that activation of the TGF-alpha /EGFR pathway contributes to hepatocellular carcinoma (HCC) formation. Herein, we developed an experimental model of cirrhosis giving rise to HCC and tested the antitumoral effect of gefitinib, a selective EGFR tyrosine kinase inhibitor, in this model. Rats received weekly intraperitoneal injections of diethylnitrosamine (DEN) followed by a 2-week wash out period that caused cirrhosis in 14 weeks and multifocal HCC in 18 weeks. Hepatocyte proliferation was increased in diseased tissue at 14 weeks compared with control liver and at even higher levels in HCC nodules compared with surrounding diseased tissues at 18 weeks. Increased proliferation was paralleled by upregulation of TGF-alpha messenger RNA expression. A group of DEN-treated rats received daily intraperitoneal injections of gefitinib between weeks 12 and 18. In rats treated with gefitinib, the number of HCC nodules was significantly lower than in untreated rats (18.1 +/- 2.4 vs. 3.7 +/- 0.45; P < .05), while EGFR was activated to a lesser extent in the diseased and tumoral tissues of these animals compared with untreated rats. HCC nodules from both untreated and gefitinib-treated animals displayed insulin-like growth factor 2 overexpression that contributed to tumor formation in treated animals. In conclusion, the blockade of EGFR activity by gefitinib has an antitumoral effect on the development of HCC in DEN-exposed rats, suggesting that it may provide benefit for the chemoprevention of HCC. PMID- 15660383 TI - MELD accurately predicts mortality in patients with alcoholic hepatitis. AB - Assessing severity of disease in patients with alcoholic hepatitis (AH) is useful for predicting mortality, guiding treatment decisions, and stratifying patients for therapeutic trials. The traditional disease-specific prognostic model used for this purpose is the Maddrey discriminant function (DF). The model for end stage liver disease (MELD) is a more recently developed scoring system that has been validated as an independent predictor of patient survival in candidates for liver transplantation. The aim of the present study was to examine the ability of MELD to predict mortality in patients with AH. A retrospective cohort study of 73 patients diagnosed with AH between 1995 and 2001 was performed at the Mayo Clinic in Rochester, Minnesota. MELD was the only independent predictor of mortality in patients with AH. MELD was comparable to DF in predicting 30-day mortality (c statistic and 95% CI: 0.83 [0.71-0.96] and 0.74 [0.62-0.87] for MELD and DF, respectively, not significant) and 90-day mortality (c-statistic and 95% CI: 0.86 [0.77-0.96] and 0.83 [0.74-0.92] for MELD and DF, respectively, not significant). A MELD score of 21 had a sensitivity of 75% and a specificity of 75% in predicting 90-day mortality in AH. In conclusion, MELD is useful for predicting 30-day and 90-day mortality in patients with AH and maintains some practical and statistical advantages over DF in predicting mortality rate in these patients. MELD is a useful clinical tool for gauging mortality and guiding treatment decisions in patients with AH, particularly those complicated by ascites and/or encephalopathy. PMID- 15660384 TI - Hepatitis B virus mutations associated with fulminant hepatitis induce apoptosis in primary Tupaia hepatocytes. AB - Hepatitis B virus (HBV) core promoter mutations have been implicated in the pathogenesis of fulminant hepatitis B. Due to the limited availability of primary human hepatocytes, the functional characterization of HBV mutants has been performed predominantly in transformed cells, which may not represent ideal model systems for studying virus-cell interactions. We and others have shown that primary hepatocytes of the tree shrew Tupaia belangeri support HBV infection and replication. In this study, we used primary Tupaia hepatocytes to analyze the phenotype of two HBV core promoter mutations that have been associated with a clinical outbreak of fatal fulminant hepatitis. Similar to previous findings in human hepatoma cells, the HBV core promoter mutations resulted in enhanced viral replication and core expression. Surprisingly, however, the presence of the mutations had a marked effect on hepatocyte viability not previously observed in hepatoma cells. Reduced cell viability was found to be due to the induction of apoptosis, as evidenced by caspase-3 activation and nuclear fragmentation. In conclusion, HBV mutants exhibit a novel phenotype in primary hepatocytes distinctly different from previous findings in hepatoma cell lines. This phenotype may have important implications for the understanding of the fulminant clinical course associated with HBV mutations. PMID- 15660385 TI - Incidence, natural history, and risk factors for biliary sludge and stones during pregnancy. AB - Gallstones are strongly associated with higher parity in women. This study prospectively assessed the incidence, natural history, and risk factors for biliary sludge and stones during pregnancy and the postpartum in 3,254 women at an army medical center. Women with a prior cholecystectomy or with stones at their first study ultrasound were excluded. Gallbladder ultrasound and subject questionnaires were obtained in each trimester and at 4 to 6 weeks postpartum. Serum glucose, lipids, insulin, leptin, estradiol, and progesterone were measured at 26 to 28 weeks' gestation. A nested case-control study was done to examine the effects of serum leptin and insulin on incident gallbladder disease. At least two study ultrasounds were available for 3,254 women. Sludge or stones had been found on at least one study ultrasound in 5.1% by the second trimester, 7.9% by the third trimester, and 10.2% by 4 to 6 weeks postpartum. Regression of sludge and stones was common, such that overall 4.2% had new sludge or stones on the postpartum ultrasound. Twenty-eight women (0.8%) underwent cholecystectomy within the first year postpartum. Prepregnancy body mass index was a strong predictor of incident gallbladder disease (P < .001). Serum leptin was independently associated with gallbladder disease (odds ratio per 1 ng/dL increase, 1.05; 95% CI, 1.01, 1.11), even after adjusting for body mass index. In conclusion, incident gallbladder sludge and stones are common in pregnancy and the postpartum, and cholecystectomy is frequently done within the first year postpartum. Prepregnancy obesity and serum leptin are strong risk factors for pregnancy-associated gallbladder disease. PMID- 15660386 TI - Outcome in adulthood of biliary atresia: a study of 63 patients who survived for over 20 years with their native liver. AB - To define the long-term prognosis of children undergoing the Kasai operation for biliary atresia, a retrospective study was undertaken comprising 271 patients operated between 1968 and 1983. Twenty years after surgery, 63 (23%) were alive with their native liver. Serum bilirubin was normal in 21 of these patients, 12 also had normal serum aminotransferase and gamma-glutamyltransferase activities, all but 2 had signs of cirrhosis, 44 had signs of portal hypertension, 19 had late bacterial cholangitis, and 6 had gallstones. Seven female patients gave birth to 9 children, and 3 male patients fathered 6 children. After age 20, 2 patients died of liver failure and 14 underwent or are awaiting liver transplantation. Twenty-year survival with native liver was significantly better in children with biliary atresia restricted to the hepatic ducts or with cysts at the porta hepatis. In conclusion, in the long term, less than 18% of infants with biliary atresia who are treated with corrective surgery may avoid liver transplantation, but even these patients require assiduous lifelong care. PMID- 15660387 TI - Cytochrome P450 CYP2E1, but not nicotinamide adenine dinucleotide phosphate oxidase, is required for ethanol-induced oxidative DNA damage in rodent liver. AB - The occurrence of malignant tumors of the upper gastrointestinal tract and liver is, based largely on epidemiological evidence, causally related to the consumption of ethanol. It is widely recognized that oxidants play a key role in alcohol-induced liver injury; however, it is unclear how oxidants may be involved in DNA damage. We asked whether nicotinamide adenine dinucleotide phosphate oxidase, cytochrome P450 CYP2E1, or both are responsible for the production of DNA damage. The rodent Tsukamoto-French model of intragastric ethanol infusion was used. Wistar rats, Cyp2e1-, p47(phox)-null, and hCyp2e1 transgenic mice were used. The abundance of oxidative DNA adducts, mutagenic apurinic/apyrimidinic sites, and expression of base excision DNA repair genes was determined. In rats and wild-type mice, ethanol treatment for 4 weeks led to an increase in oxidative DNA damage and induction of expression of the base excision DNA repair genes that are known to remove oxidative DNA lesions. No increase in either of the endpoints was observed in ethanol-treated Cyp2e1-null mice, whereas the magnitude of response in p47(phox)-null mice and transgenic hCyp2e1 was identical to that in wild types. The increase in expression of DNA repair genes was completely abolished by treatment with the P450 inhibitor 1-aminobenzotriazole. In conclusion, the data support the hypothesis that oxidative stress to DNA is induced in liver by ethanol. Furthermore, although it was shown that nicotinamide adenine dinucleotide phosphate oxidase-derived oxidants are critical for the development of ethanol-induced liver injury, CYP2E1 is required for the induction of oxidative stress to DNA, and thus may play a key role in ethanol-associated hepatocarcinogenesis. PMID- 15660388 TI - Serum bilirubin levels and mortality after myeloablative allogeneic hematopoietic cell transplantation. AB - Many patients who undergo hematopoietic cell transplantation experience liver injury. We examined the association of serum bilirubin levels with nonrelapse mortality by day +200, testing the hypothesis that the duration of jaundice up to a given point in time provides more prognostic information than either the maximum bilirubin value or the value at that point in time. We studied 1,419 consecutive patients transplanted from allogeneic donors. Total serum bilirubin values up to day +100, death, or relapse were retrieved-along with nonrelapse mortality by day +200 as an outcome measure--using Cox regression models with each bilirubin measure modeled as a time-dependent covariate. The bilirubin value at a particular point in time provided the best fit to the model for mortality. With bilirubin at a point in time modeled as an 8th-degree polynomial, an increase in bilirubin from 1 to 3 mg/dL is associated with a mortality hazard ratio of 6.42. An increase from 4 to 6 mg/dL yields a hazard ratio of 2.05, and an increase from 10 to 12 mg/dL yields a hazard ratio of 1.17. Among patients who were deeply jaundiced, survival was related to the absence of multiorgan failure and to higher platelet counts. In conclusion, the value of total serum bilirubin at a particular point in time after transplant carries more informative prognostic information than does the maximum or average value up to that point in time. The increase in mortality for a given increase in bilirubin value is larger when the starting value is lower. PMID- 15660389 TI - Sources of variability in histological scoring of chronic viral hepatitis. AB - Inter-observer agreement on activity and fibrosis scores used in chronic viral hepatitis has only been studied under selected conditions. The aim of this study was to identify the sources of variability due to specimen characteristics and observers. This study included 254 liver specimens and 15 pathologists and used the Metavir score. In 44 specimens scored by 4 academic pathologists, agreement of Metavir score was good overall, but better for fibrosis (kappa = 0.59) than for activity (kappa = 0.43) and poor for lobular necrosis (kappa = 0.15). The mean agreement was better for senior (0.60 +/- 0.24) than junior pathologists (0.52 +/- 0.30, P < .05). Mean intrabserver agreement was better than inter observer agreement (0.77 +/- 0.18 vs. 0.58 +/- 0.26, P < .01). In 157 specimens scored by 2 expert pathologists (one senior, one junior), agreement of Metavir score was only good but greatly improved after consensus reading (fibrosis: kappa = 0.48 and 0.77, activity: kappa = 0.44 and 0.70, respectively, before and after consensus). Several causes of disagreement were identified: specimen length, fibrosis class number, observer bias, and putative causes related to Metavir score or specimen. In an intercenter evaluation involving 59 specimens, 1 expert and 10 nonacademic pathologists, agreement was very poor and did not improve over 5 years for activity (kappa = 0.22-0.25) or fibrosis (kappa = 0.13-0.18). In conclusion, the level of experience (specialization, duration, and location of practice) has more influence on agreement than the characteristics of the specimen (length, fibrosis class number, miscellaneous factors). Agreement can be improved by experienced pathologist or consensus reading. PMID- 15660390 TI - Differential lymphotoxin-beta and interferon gamma signaling during mouse liver regeneration induced by chronic and acute injury. AB - The liver regenerates after acute injury via hepatocyte cell division; during chronic injury, when hepatocyte replication is impaired or blocked, liver progenitor oval cells mediate liver regeneration. If both regeneration options are blocked in animal models, then liver failure and death ensues. The mechanisms underlying oval cell induction, proliferation, and subsequent liver regeneration remain poorly characterized. In particular, cell-signaling pathways that distinguish the alternative pathways are unknown. This study shows that in a mouse model, hepatic expression of lymphotoxin-beta (LTbeta) and interferon gamma (IFNgamma) transcripts is increased in response to the choline-deficient, ethionine-supplemented (CDE) diet, which induces oval cell-mediated liver regeneration. Oval cells express LTbeta and IFNgamma transcripts, contributing to the increased expression in the liver of mice fed the CDE diet. An attenuated oval cell response to such a diet was observed in LTbeta receptor-, LTbeta-, and IFNgamma-gene targeted mice. Loss of LTbeta and LTbeta receptor signaling reduced the number of oval cells expressing A6 and muscle pyruvate kinase. The lack of IFNgamma signaling reduced muscle pyruvate kinase(+), but not A6(+), oval cells. In contrast, partial hepatectomy suppressed LTbeta and IFNgamma transcripts. We also show that IFNgamma induces STAT-3 phosphorylation in an oval cell line. In conclusion, LTbeta, LTbeta receptor, and IFNgamma are involved in oval cell mediated, but not hepatocyte-mediated, liver regeneration, and the absence of these pathways impairs the oval cell-dependent regenerative response. PMID- 15660391 TI - NFkappaB-mediated upregulation of bcl-xl restrains TRAIL-mediated apoptosis in murine viral hepatitis. AB - Inhibition of NFkappaB enhances the susceptibility of cancer to TRAIL-mediated apoptosis and is suggested as a strategy for cancer therapy. Because the role of NFkappaB in TRAIL-mediated apoptosis of hepatocytes is unknown, we investigated the influence of NFkappaB-inhibition in death ligand-mediated apoptosis in hepatitis. Adenoviral hepatitis resulted in upregulation of NFkappaB-activity, which could be inhibited by expression of IkappaBalpha-superrepressor. We treated mice after the onset of adenoviral hepatitis with adenoviruses expressing FasL (AdFasL), TRAIL (AdTRAIL), or GFP (AdGFP). In contrast to apoptosis induced by AdFasL, NFkappaB inhibition strongly enhanced AdTRAIL-mediated apoptosis of hepatocytes. Expression of IkappaBalpha inhibits adenoviral infection-mediated overexpression of bcl-xl, providing a molecular mechanism for TRAIL sensitization. In agreement with this hypothesis, downregulation of bcl-xl by siRNA enhanced susceptibility of hepatocytes to TRAIL, but not to FasL-mediated apoptosis, resulting in TRAIL-mediated severe liver damage after AdTRAIL application. Our data demonstrate that inhibition of NFkappaB in adenoviral hepatitis strongly sensitizes hepatocytes to TRAIL-mediated apoptosis. Bcl-xl, in contrast to bcl-2 and c-FLIP, is strongly upregulated after viral infection and represents an essential NFkappaB-dependent survival factor against TRAIL-mediated apoptosis. In conclusion, inhibition of NFkappaB or bcl-xl during TRAIL therapy may harbor a risk of liver damage in patients with viral hepatitis. PMID- 15660392 TI - Peginterferon alfa-2a for hepatitis C after liver transplantation: two randomized, controlled trials. AB - There is currently no effective treatment for recurrent hepatitis C after orthotopic liver transplantation (OLT). We therefore performed two randomized, controlled trials--a prophylaxis trial and a treatment trial--to evaluate the safety and efficacy of peginterferon alfa-2a in patients who had undergone OLT. The prophylaxis trial enrolled 54 patients within 3 weeks after OLT, and the treatment trial enrolled 67 patients 6 to 60 months after OLT. In each trial, patients were randomized to treatment with once weekly injections of 180 microg peginterferon alfa-2a or no antiviral treatment for 48 weeks and were followed up for 24 weeks thereafter. Peginterferon alfa-2a treated patients had significantly lower hepatitis C virus RNA levels and more favorable changes in hepatic histological features compared with untreated controls. However, only 2 treated patients in the prophylaxis trial (8%) and 3 in the treatment trial (12%) achieved a sustained virological response. In the prophylaxis trial, 8 patients (31%) in the peginterferon alfa-2a group and 9 (32%) in the untreated group were withdrawn prematurely; whereas in the treatment trial, 10 patients (30%) in the peginterferon alfa-2a group and 6 (19%) in the untreated group were withdrawn prematurely. The incidence of acute rejection was similar in the treated and untreated groups in both the prophylaxis (12% vs. 21%; P = .5) and treatment (12% vs. 0%; P = .1) trials. In conclusion, peginterferon alfa-2a treatment for 48 weeks is safe and tolerable and offers some efficacy in the post-OLT setting. Randomized controlled studies are needed to establish the efficacy of pegylated interferon and ribavirin in patients who have undergone OLT. PMID- 15660393 TI - High-dose ribavirin in combination with standard dose peginterferon for treatment of patients with chronic hepatitis C. AB - Improved treatment regimens for patients with chronic hepatitis C, genotype 1 and high viral load are needed. Increasing the dose of ribavirin has increased the response rate, but experience with doses of more than 1,200 mg/day is limited. The aim of this study was to investigate the safety and tolerance to treatment with a high and individualized dose of ribavirin in combination with peginterferon. Ten patients with chronic hepatitis C, genotype 1 and high viral load were treated with peginterferon alfa-2a and ribavirin for 48 weeks in a prospective trial. The initial ribavirin dose was individualized and calculated from a pharmacokinetic formula based mainly on renal function. Ribavirin plasma concentrations were monitored, and the dose was adjusted to reach the target concentration. Hemoglobin was monitored, and patients were treated with erythropoietin and blood transfusions when indicated. After dose adjustments, the mean dose of ribavirin was 2,540 mg/day (range, 1,600-3,600) at week 24. The main side effect was anemia, which was controlled with erythropoietin. Two patients required blood transfusions. One patient was withdrawn at week 24 because of a lack of viral response, and one patient at week 39 because of side effects, primarily interferon associated. At follow-up (>or=24 weeks posttreatment), nine of ten patients had undetectable HCV RNA and thus were cured by standard definitions. In conclusion, a high dose of ribavirin according to an individualized schedule is feasible but associated with more frequent and serious side effects such as anemia. The viral response merits further evaluation. PMID- 15660394 TI - Fluorescence resonance energy transfer analysis of proapoptotic CD95-EGF receptor interactions in Huh7 cells. AB - Hyperosmolarity- and CD95 ligand (CD95L)-induced interactions between CD95 (Fas/APO-1) and the epidermal growth factor receptor (EGFR) involve EGFR catalyzed CD95 tyrosine phosphorylation. Such interactions were studied by means of fluorescence resonance energy transfer (FRET) and CD95 receptor mutagenesis in Huh7 hepatoma cells. In cells cotransfected with EGFR-cyan fluorescent protein and CD95-yellow fluorescent protein, FRET studies showed a rapid, hyperosmolarity induced, c-Jun-N-terminal kinase-dependent CD95-EGFR association in the cytosol with subsequent microtubule-dependent translocation of the protein complex to the plasma membrane. Inhibition of EGFR tyrosine kinase activity by AG1478 and cyclic adenosine monophosphate had no effect on hyperosmotic CD95-EGFR association in the cytosol but prevented CD95 tyrosine phosphorylation, targeting of the protein complex to the plasma membrane, and formation of the death-inducing signaling complex (DISC). The requirement of EGFR-mediated CD95 tyrosine phosphorylation for hyperosmotic and CD95L-induced CD95 membrane targeting and DISC formation was also shown in CD95 mutagenesis experiments. CD95 mutants with tyrosine phenylalanine exchanges at positions 232 and 291 failed to translocate to the plasma membrane and to recruit Fas-associated death domain and caspase 8, although these mutants still associated with the EGFR in the cytosol in response to hyperosmolarity and CD95L. Cells transfected with these mutants were also resistant to CD95L-induced apoptosis. Single mutations of tyrosine 91, 232, and 291 failed to inhibit CD95 membrane targeting, DISC formation, or CD95L-induced apoptosis. In conclusion, we identify EGFR-CD95 interaction and phosphorylation of critical CD95 tyrosine residues as important early events in hyperosmotic and CD95L-induced CD95 activation and apoptosis induction. PMID- 15660395 TI - In vitro leakage susceptibility of tracheoesophageal shunt prostheses in the absence and presence of a biofilm. AB - Although leakage through a tracheoesophageal shunt prosthesis is the main cause of prosthesis failure in a laryngectomy patient, this has never been the subject of in vitro evaluation. The aim of this study was to compare three commercially available voice prostheses by comparison of their in vitro leakage patterns, in absence or presence of a biofilm. To compare in vitro leakage patterns, a model comprised of an artificial throat equipped with a single prosthesis coupled to a water reservoir was developed. By varying the height of the water reservoir, different pressures on the voice prosthesis can be obtained. Both in absence and presence of a biofilm, the Blom Singer voice prosthesis demonstrated the lowest leakage, followed by Groningen Low Resistance. The Provox2 showed significantly the most leakage, however, in presence of a biofilm the leakage of the Provox2 significantly decreased. Regular airflow during biofilm formation significantly increased leakage through the Provox2. Out of 746 clinical replacements, Provox2 showed 76% and Groningen Low Resistance 57% replacements due to leakage. The model used in this study showed significant differences in leakage of the three types of voice prostheses used. Leakage occurred more readily through Provox2 than through Groningen Low Resistance and Blom Singer prostheses, which is in line with clinical observations and enforces the model. PMID- 15660396 TI - Characterization of host-range and cell entry properties of the major genotypes and subtypes of hepatitis C virus. AB - Because of the lack of a robust cell culture system, relatively little is known about the molecular details of the cell entry mechanism for hepatitis C virus (HCV). Recently, we described infectious HCV pseudo-particles (HCVpp) that were generated by incorporating unmodified HCV E1E2 glycoproteins into the membrane of retroviral core particles. These initial studies, performed with E1E2 glycoproteins of genotype 1, noted that HCVpp closely mimic the cell entry and neutralization properties of parental HCV. Because sequence variations in E1 and E2 may account for differences in tropism, replication properties, neutralization, and response to treatment in patients infected with different genotypes, we investigated the functional properties of HCV envelope glycoproteins from different genotypes/subtypes. Our studies indicate that hepatocytes were preferential targets of infection in vitro, although HCV replication in extrahepatic sites has been reported in vivo. Receptor competition assays using antibodies against the CD81 ectodomain as well as ectopic expression of CD81 in CD81-deficient HepG2 cells indicated that CD81 is used by all the different genotypes/subtypes analyzed to enter the cells. However, by silencing RNA (siRNA) interference assays, our results show that the level of Scavenger Receptor Class-B Type-I (SR-BI) needed for efficient infection varies between genotypes and subtypes. Finally, sera from chronic HCV carriers were found to exhibit broadly reactive activities that inhibited HCVpp cell entry, but failed to neutralize all the different genotypes. In conclusion, we characterize common steps in the cell entry pathways of the major HCV genotypes that should provide clues for the development of cell entry inhibitors and vaccines. PMID- 15660397 TI - The feasibility of adjuvant interferon alpha-2b in children with high-risk melanoma. AB - BACKGROUND: It has been shown that induction high-dose interferon alpha-2b (IFN alpha-2b) followed by maintenance therapy improves recurrence-free survival in adults with high-risk, resected melanoma. In this study, the feasibility and toxicity of this regimen were evaluated in newly diagnosed pediatric patients with Stage III melanoma involving regional lymph nodes. METHODS: Fifteen patients age or= 4 hours of sleep during normal waking hours) in 9 patients (35%) and Grade 2 somnolence (800 mg if it was delivered at bedtime. Grade 3-4 skin reactions were observed in 20% of patients, and exfoliative dermatitis was observed in 1 responding patient. The overall median survival was 6.8 months. CONCLUSIONS: With a 5% PR rate, a 5% MR rate, and a 31% SD rate, the results indicate that thalidomide mostly may offer HCC patients disease stabilization. It is possible that, at a different dosage, or combined with other chemotherapy agents, or with the use of a different thalidomide analogue, longer patient survival may be achieved. However, in view of the significant neurologic toxicity encountered among these commonly cirrhotic HCC patients, thalidomide monotherapy at the high doses studied cannot be recommended for the treatment of HCC. PMID- 15660402 TI - The liver has a body--a Cook's tour. PMID- 15660403 TI - Trends in suicide by drowning in the elderly in England and Wales 1979-2001. AB - BACKGROUND: Suicide by drowning increases with age but its rates vary between countries and among communities. Drowning suicide rates in some of the available studies may have been over reported or under reported because of misclassification. OBJECTIVE: This study presents data on the time trends, age/sex mortality rates from death by drowning in the elderly in England and Wales between 1979 and 2001. All coroners' verdicts in death by drowning; suicide, deaths undetermined whether accidentally or purposely inflicted were examined. METHODS: Counts of suicide due to drowning and submersion [ICD 9 codes; E954] and undetermined injury deaths [E984] (WHO, 1977), reported in England and Wales between 1979 and 2001 were obtained from National Statistics (ONS). RESULTS: There has been a gradual reduction in suicide by drowning in men and women by more than a third the observed count in 1979 (p < 0.01). However, this decline was less evident in the elderly particularly those over the age of 75. Elderly drownings appear to attract more verdicts of suicide compared to younger age groups (Odds Ratio 4.3 95% CI 2.3-8.3). Women, particularly elderly, are more likely to have a suicide verdict returned in drowning compared to men (Odds Ratio 1.5 95% CI 1.1-1.6). CONCLUSIONS: The high rate of open verdicts in elderly drowning over the study period and compared to any other method of fatal self harm in England and Wales confirms the difficulties in reaching a firm conclusion in drowning death. Therefore combining suicide and all undetermined deaths in drowning as a matter of course, in nationally collected statistics, may result in grossly exaggerated rates and misleading trends in suicidal drowning. Suicide by drowning is probably not amenable to prevention and although the elderly are often thought to benefit more from suicide prevention than younger adults, the study findings seem to suggest that this is not likely to be the case in drowning. PMID- 15660404 TI - Social distance towards a person with Alzheimer's disease. AB - OBJECTIVES: This study examined determinants of behavioral discrimination (i.e. social distance) against persons with Alzheimer's disease (AD). Face-to-face interviews were conducted with a total of 206 Jewish Israeli adults (mean age = 59.7) using an experimental vignette methodology, varying in the severity of the disease. METHODS: Information regarding participants' preferences for social distance from a person with AD was elicited. The effects of emotional reactions, perceptions of dangerousness of the person depicted in the vignette, personal experience with the disease, and perceptions about the susceptibility of developing AD were examined. RESULTS: Results of the study provide partial support for an extended version of the attribution model, in that prosocial feelings were found to decrease the behavioral discrimination against the person described, whereas feelings of rejection increased the discrimination. A direct, unmediated effect of the severity of the disease on social distance was found, as well as a direct effect of familiarity with someone with the disease. CONCLUSION: Findings of this study stress the importance of increasing government and advocacy initiatives that would provide the lay public with accurate knowledge about the disease, its symptoms, and treatments. These campaigns should be aimed at clarifying misconceptions and communicating empathetic feelings towards people diagnosed with AD. PMID- 15660405 TI - The role of stigma in the quality of life of older adults with severe mental illness. AB - BACKGROUND: Stigma and discrimination against older people with mental illness is a seriously neglected problem. OBJECTIVES: (1) To investigate whether stigmatisation of older adults with mental disorder is associated with the type of residential institution they live in or the type of disorder they suffer and (2) to assess the role of stigma experiences in their quality of life. METHODS: A cross-sectional study was carried out of 131 older adults with severe mental illness, recruited in 18 elder care homes operating supported living programmes and in eight psychiatric hospitals throughout the Netherlands. Stigmatisation was assessed with an 11-item questionnaire on stigma experiences associated with mental illness. Quality of life was assessed with the Manchester Short Assessment of Quality of Life (MANSA). To better ascertain the role of stigma, we also assessed in comparison the relationship of social participation to quality of life. RESULTS: Some 57% of the respondents had experienced stigmatisation. No association emerged between residential type or disorder type and the extent of stigma experiences. Stigmatisation did show a negative association with quality of life, a connection stronger than that between social participation and quality of life. CONCLUSION: A feeling of belonging, as contrasted with being excluded, is at least as important for the quality of life of older people with severe mental illness as their actual participation in the community. PMID- 15660406 TI - Physical health, depression and cognitive function as correlates of disability in an older Korean population. AB - BACKGROUND: The World Health Organisation Disability Assessment Schedule II (WHODAS II) measures functioning and disability in concordance with the bio psycho-social model of the WHO's International Classification of Functioning, Disability, and Health. Our objectives were to investigate the independent associations of physical health, depression and cognitive function with WHODAS II disability in an older Korean population. METHODS: The WHODAS II was administered to a community sample of 1204 residents age 65 or over. Data on demographic characteristics (age, gender, living area, marital state, and religion), socio economic state (education, type of accommodation, number of rooms, previous occupation, current employment, monthly income), social support, number of physical illness, depression (GMS), and cognitive function (MMSE) was gathered. A subsample (n = 746) received a clinical examination for dementia. RESULTS: Scores on the WHODAS II were significantly and independently associated with worse physical health, depression, and lower cognitive function. Associations with other factors were no longer apparent after adjustment for these. These associations persisted in the absence of dementia. In participants with dementia (n = 110), WHODAS II scores were principally associated with physical health and accommodation type. CONCLUSIONS: Level of disability, as measured by the WHODAS II, was principally associated with physical health, depression and cognitive function, rather than socio-demographic factors. PMID- 15660407 TI - Antidepressant drug prescribing among elderly subjects: a population-based study. AB - BACKGROUND: The patterns of antidepressant drug prescribing have rarely been studied in large and geographically defined populations of elderly subjects. In the present study we examined the prevalence and distribution of antidepressant prescribing in Lombardy, a northern Italy region with more than one and a half million elderly inhabitants. METHODS: We used the Regional Administrative Database of Lombardy. This database includes all prescriptions reimbursed by the National Health System in the population living in this region. All antidepressant prescriptions dispensed to subjects aged 65 years or above during 2001 were extracted and prevalence data calculated by dividing antidepressant users by the total number of male and female residents in each age group. RESULTS: During the 12 months surveyed 153,706 subjects were dispensed one or more prescriptions of antidepressants, yielding a prevalence of use of 9.49 subjects per 100 inhabitants (95% confidence interval 9.44, 9.53). Although the proportion of chronic users slightly decreased with age, more than 35% of those older that 85 years were moderate or chronic antidepressant users. General practitioners issued the majority of antidepressant prescriptions, and most antidepressant users were also dispensed agents for medical disorders. CONCLUSIONS: The very high rates of antidepressant drug prescribing detected in late life suggest the need of characterising these subjects in terms of medical and psychiatric characteristics, needs and quality of life. It also suggests the need for pragmatic clinical trials, carried out in the general practice, with the aim of assessing whether antidepressants are effective in these conditions. PMID- 15660408 TI - Object naming and semantic fluency among individuals with genetic risk for Alzheimer's disease. AB - OBJECTIVES: This study longitudinally examined the object naming and semantic fluency of individuals who are at risk for developing Alzheimer's disease (AD) by virtue of having APOE-4 or a family history of AD. METHODS: A total of 108 participants (40 with a family history of AD and 43 with APOE-4) completed the Boston Naming Test and the Animal Naming task at initial assessment and after two years. RESULTS: At baseline, object naming was significantly lower for those with both risk factors, F(2, 99) = 5.72, p < 0.01, but those with either risk factor had significantly lower scores at follow-up, F(2, 99) = 3.41, p < 0.05. Semantic fluency (animal naming) was reduced among subjects with the APOE-4 allele at baseline, F(1, 100) = 4.02, p < 0.05, but it was not associated with either risk factor at follow-up. CONCLUSIONS: These deficits may be associated with a prodromal risk for AD and may serve as pre-symptomatic markers for the development of AD. PMID- 15660410 TI - Biomarkers for apoptosis in Alzheimer's disease. AB - Alzheimer's disease (AD) affects millions of people worldwide and the number of AD cases will increase with increased life expectancy. Today there is no cure for this devastating and always lethal disease and therefore it is of great interest for patients, relatives and societies to find new drugs that can hinder the disease process. During the progression of AD a substantial amount of neurons degenerate in the brain. The mechanisms of cell death involved in AD have not been fully elucidated. However, there are several reports showing that neurons die partly by apoptosis in the AD brain. Drugs blocking apoptosis could therefore be potentially useful for early prevention of neuronal cell death. Biomarkers for apoptosis should be important tools in the evaluation of drug effects and in the diagnostics of AD. Here we review the current knowledge in the field and discuss potential biomarkers for apoptosis in AD. PMID- 15660409 TI - Predictors of high level of burden and distress in caregivers of demented patients: results of an Italian multicenter study. AB - BACKGROUND: Considerable suffering is experienced by carers of patients with dementia. Most existing studies do not consider the coexistence of subjective and objective aspects that cause, interacting to each other, this suffering. OBJECTIVES: In this study we: (1) define the high-risk group of caregivers on the bases of the scores obtained on the four scales evaluating burden, distress, depression and anxiety (BDDA) taken into account simultaneously and (2) evaluate risk factors related to the high level of BDDA. SUBJECTS AND METHODS: 419 elderly outpatients with dementia and their caregivers were enrolled. Patients were evaluated for their cognitive, neuropsychological and functional impairment and for comorbidity. Caregivers were evaluated with four scales for the assessment of burden, distress related to neuropsychological disturbances, depression and anxiety. Cluster analysis was used to identify the group with the High level of BDDA (HBDDA). RESULTS: By multiple logistic analysis, disability, specific behavioural disturbances of the patients as well as caregiver's age, type of relationship and living in the south of Italy were observed to be a major risk factor for HBDDA. CONCLUSION: The targeted use of scales specifically assessing BDDA of the caregiver and the identification of particular patient and caregiver characteristics are able to allow a precise and early definition of caregivers at high risk of burden and distress. This might be helpful in planning the correct social/clinical/rehabilitative approach. PMID- 15660411 TI - Components of behavioral pathology in dementia. AB - OBJECTIVE: The purpose of this study was to examine the occurence of the noncognitive behavioral and psychological symptoms and signs of dementia in a geriatric chronic-care hospital and to separate agitated and affective components of behavioral pathology using factor analysis. METHODS: The frequency and severity of Alzheimer's disease, vascular dementia, mixed dementia and Lewy Body dementia was assessed in 145 consecutive residents of a chronic-care hospital. The presence of noncognitive behavioral symptoms was evaluated with the Behavioral Pathology in Alzheimer's Disease Rating Scale (BEHAVE-AD) and the Cohen-Mansfield Agitation Inventory (CMAI). A Factor analysis on the BEHAVE-AD subscores was performed to create symptom clusters. Analysis of covariance and post hoc tests were used to compare means of factor variables between different types of dementia. RESULTS: Statistical analysis showed a significant correlation between severity of dementia and BEHAVE-AD total score and between severity of dementia and CMAI total score. Factor analysis with Varimax rotation revealed the presence of three behavioral subsyndromes: agitation, affectivity and day/night disturbances. CONCLUSIONS: The finding of three factors of behavioral pathology in demented patients reflects the possibility that different etiological mechanisms could explain the expression of the symptoms and signs of psychosis in demented patients. PMID- 15660412 TI - Plasma insulin growth factor-1 and incident delirium in older people. AB - BACKGROUND: A variety of demographic and clinical variables are acknowledged as risk factors for delirium; a syndrome thought to be mediated by abnormalities in a wide range of neurotransmitters. However, little research has been conducted in this field and the role of neuro-immunological factors as a mechanism of medication has received very little attention. AIMS: To determine if low base line (on admission) IGF-1 levels (a protective cytokine released by brain cells in response to insult) is a risk factor for incident delirium in patients aged 75 and over admitted to an acute medical ward. METHOD: Base line demographic and clinical variables and serum IGF-1 levels were measured in a consecutive series of 100 non-delirious subjects on inpatient admission. Subjects were assessed daily regarding the development of delirium during the inpatient episode. RESULTS: Twelve patients developed incident delirium. IGF-1 (OR: 0.822, CI: 0.69, 0.97, p = 0.027), pre-admission cognitive deterioration (assessed by IQCODE) (OR; 3.26, CI: 1.18, 9.04, p = 0.023) and depression (GDS four item: cut-off score > or = 3) (OR; 8.99, CI 1.59,50.76, p = 0.013) were identified as risk factors for developing subsequent delirium. CONCLUSIONS: Despite the small size of this study our findings suggest that low, pre-morbid IGF-1 is a risk factor for subsequent delirium in this population, emphasizing the potential protective role of this anabolic cytokine and the need for replication of these findings. PMID- 15660413 TI - New molecular markers of early and progressive CJD brain infection. AB - Transmissible spongiform encephalopathies (TSEs), including human Creutzfeldt Jakob disease (CJD), are caused by a related group of infectious agents that can be transmitted to many mammalian species. Because the infectious component of TSE agents has not been identified, we examined myeloid cell linked inflammatory pathways to find if they were activated early in CJD infection. We here identify a specific set of transcripts in CJD infected mouse brains that define early and later stages of progressive disease. Serum amyloid A3 and L-selectin mRNAs were elevated as early as 20 days after intracerebral inoculation. Transcripts of myeloid cell recruitment factors such as MIP-1alpha, MIP-1beta, and MCP1, as well as IL1alpha and TNFalpha were upregulated > 10 fold between 30 and 40 days, well before prion protein (PrP) abnormalities that begin only after 80 days. At later stages of symptomatic neurodegenerative disease (100-110 days), a selected set of transcripts rose by as much as 100 fold. In contrast, normal brain inoculated controls showed no similar sequential changes. In sum, rapid and simple PCR tests defined progressive stages of CJD brain infection. These markers may also facilitate early diagnosis of CJD in accessible peripheral tissues such as spleen and blood. Because some TSE strains can differentially target particular cell types such as microglia, several of these molecular changes may also distinguish specific agent strains. The many host responses to the CJD agent challenge the assumption that the immune system does not recognize TSE infections because these agents are composed only of the host's own PrP. PMID- 15660414 TI - Modeled microgravity disrupts collagen I/integrin signaling during osteoblastic differentiation of human mesenchymal stem cells. AB - Spaceflight leads to reduced bone mineral density in weight bearing bones that is primarily attributed to a reduction in bone formation. We have previously demonstrated severely reduced osteoblastogenesis of human mesenchymal stem cells (hMSC) following 7 days culture in modeled microgravity (MMG). One potential mechanism for reduced osteoblastic differentiation is disruption of type I collagen (Col I)-integrin interactions and reduced integrin signaling. Integrins are heterodimeric transmembrane receptors that bind extracellular matrix (ECM) proteins and produce signals essential for proper cellular function, survival, and differentiation. Therefore, we investigated the effects of MMG on integrin expression and function in hMSC. We demonstrate that 7 days of culture in MMG leads to reduced expression of the ECM protein, Col I. Conversely, MMG consistently increases Col I-specific alpha2 and beta1 integrin protein expression. Despite this increase in integrin subunit expression, autophosphorylation of adhesion-dependent kinases, focal adhesion kinase (FAK) and proline-rich tyrosine kinase 2 (PYK2), is significantly reduced. Activation of Akt protein kinase (Akt) is unaffected by the reduction in FAK activation. However, reduced downstream signaling via the Ras-mitogen activated protein kinase (MAPK) pathway is evidenced by a reduction in Ras and extracellular signal related protein kinase (ERK) activation. Taken together, our findings indicate that MMG decreases integrin/MAPK signaling, which likely contributes to the observed reduction in osteoblastogenesis. PMID- 15660415 TI - Abnormal cell cycle regulation in primary human uveal melanoma cultures. AB - Uveal malignant melanoma is the most frequent primary intraocular tumor in adult humans. The cellular events leading to neoplasic transformation of normal uveal melanocytes are not well known when compared to other cancers. In this study, we investigated the role of G1 and G1/S regulatory proteins of the cell cycle in human uveal melanoma (UM) primary cell cultures, since these proteins are common targets in tumor development. Further, freshly established and characterized tumor cells are a better model for in vitro studies when compared to cell lines established long ago. Human primary cell cultures from eight different UM were established, as well as one primary culture from rhesus uveal normal melanocytes (UNM). Primary human UM cultures were characterized by a low establishment and growing rate. From four successful cultures, three showed a high expression of cyclin D1, cyclin E, p16NK4A, and p27KIP1 with no variations in cyclin A, cyclin dependent kinase 2 (CDK2), and CDK4. Interestingly, in one of the cultured tumors, tumor suppressor protein retinoblastoma (Rb) did not bind E2F despite the fact that Rb was found in its hypophosphorylated form. No mutations in either RB1 or the Rb-binding pocket of E2F-1 were detected. Furthermore, we identified seven proteins co-immunoprecipitating with Rb in this tumor, including Lamin A/C and six proteins not previously reported to bind Rb: Hsc70, high mobility group protein 1 (HMG-1), hnRPN, glyceraldehyde 3 phosphate dehydrogenase (G3PDH), EF-1, and EF-2. Our results indicate that the overexpression of cyclins D1/E and CDKIs p16 and p27, together with a deregulation of the Rb/E2F pathway, may be implicated in the development of human UM. PMID- 15660416 TI - Cytoskeletal organization of human mesenchymal stem cells (MSC) changes during their osteogenic differentiation. AB - Human MSCs have been studied to define the mechanisms involved in normal bone remodeling and the regulation of osteogenesis. During osteogenic differentiation, MSCs change from their characteristic fibroblast-like phenotype to near spherical shape. In this study, we analyzed the correlation between the organization of cytoskeleton of MSCs, changes in cell morphology, and the expression of specific markers (alkaline phosphatase activity and calcium deposition) of osteogenic differentiation. For osteoblastic differentiation, cells were cultured in a culture medium supplemented with 100 nM dexamethasone, 10 mM beta- glycerophosphate, and 50 microg/ml ascorbic acid. The organization of microfilaments and microtubules was examined by inmunofluorescence using Alexa fluor 594 phalloidin and anti alpha-tubulin monoclonal antibody. Cytochalasin D and nocodazole were used to alter reversibly the cytoskeleton dynamic. A remarkable change in cytoskeleton organization was observed in human MSCs during osteogenic differentiation. Actin cytoskeleton changed from a large number of thin, parallel microfilament bundles extending across the entire cytoplasm in undifferentiated MSCs to a few thick actin filament bundles located at the outermost periphery in differentiated cells. Under osteogenic culture conditions, a reversible reorganization of microfilaments induced by an initial treatment with cytochalasin D but not with nocodazole reduced the expression of differentiation markers, without affecting the final morphology of the cells. The results indicate that changes in the assembly and disassembly kinetics of microfilaments dynamic of actin network formation may be critical in supporting the osteogenic differentiation of human MSCs; also indicated that the organization of microtubules appears to have a regulatory role on the kinetic of this process. PMID- 15660417 TI - Proteomic analyzes of copper metabolism in an in vitro model of Wilson disease using surface enhanced laser desorption/ionization-time of flight-mass spectrometry. AB - In Wilson disease, mutations in the ATP7B-gene lead to hepatic accumulation of copper that becomes toxic when the hepatic binding capacity is exceeded, leading to oxidative stress and acute liver failure. Several proteins are probably involved in dealing with the excess copper and oxidative stress. As a first step towards biomarker discovery and analyzes of copper metabolism in Wilson disease patients we characterized copper-induced changes in protein expression in cell lysates and culture media from an in vitro copper-overload model using surface enhanced laser desorption/ionization (SELDI) proteomics technology. HepG2 cells were cultured for 48 h with a physiological (0.5 microM) or a pathological (100 microM) copper concentration. Samples were applied to weak cation exchange (WCX) proteinchip arrays and chips were analyzed by time of flight (TOF)-mass spectrometry. Copper-coated IMAC chips were used to detect copper-binding proteins in cell lysate of copper depleted cells using buffers with increasing imidazole concentrations. Data from the 2 to 50 kDa range indicate that high extra-cellular copper substantially altered both intra-cellular protein expression as well as the composition of the secretome. In the lysate 15 proteins were found up-regulated, while 6 proteins were down-regulated. In culture media 21 proteins were increased while 4 proteins were decreased in abundance. Copper coated protein chips revealed the presence of 18 high-affinity copper-binding proteins. Further identification is necessary to determine the exact cellular roles of the discovered proteins. PMID- 15660418 TI - Differential regulation of platelet-derived growth factor stimulated migration and proliferation in osteoblastic cells. AB - Osteoblastic migration and proliferation in response to growth factors are essential for skeletal development, bone remodeling, and fracture repair, as well as pathologic processes, such as metastasis. We studied migration in response to platelet-derived growth factor (PDGF, 10 ng/ml) in a wounding model. PDGF stimulated a twofold increase in migration of osteoblastic MC3T3-E1 cells and murine calvarial osteoblasts over 24-48 h. PDGF also stimulated a tenfold increase in 3H-thymidine (3H-TdR) incorporation in MC3T3-E1 cells. Migration and DNA replication, as measured by BrdU incorporation, could be stimulated in the same cell. Blocking DNA replication with aphidicolin did not reduce the distance migrated. To examine the role of mitogen-activated protein (MAP) kinases in migration and proliferation, we used specific inhibitors of p38 MAP kinase, extracellular signal regulated kinase (ERK), and c-Jun N-terminal kinase (JNK). For these signaling studies, proliferation was measured by carboxyfluorescein diacetate succinimidyl ester (CFSE) using flow cytometry. Inhibition of the p38 MAP kinase pathway by SB203580 and SB202190 blocked PDGF-stimulated migration but had no effect on proliferation. Inhibition of the ERK pathway by PD98059 and U0126 inhibited proliferation but did not inhibit migration. Inhibition of JNK activity by SP600125 inhibited both migration and proliferation. Hence, the stimulation of migration and proliferation by PDGF occurred by both overlapping and independent pathways. The JNK pathway was involved in both migration and proliferation, whereas the p38 pathway was predominantly involved in migration and the ERK pathway predominantly involved in proliferation. PMID- 15660419 TI - Constitutive association of cell surface CCR5 and CXCR4 in the presence of CD4. AB - Chemokine receptors CCR5 and CXCR4 are the major coreceptors of HIV-1 infection and also play fundamental roles in leukocyte trafficking, metastasis, angiogenesis, and embyogenesis. Here, we show that transfection of CCR5 into CXCR4 and CD4 expressing 3T3 cells enhances the cell surface level of CXCR4. In CCR5 high expressing cells, cell surface level of CXCR4 was incompletely modulated in the presence of the CXCR4 ligand CXCL12/SDF-1alpha. CCR5 was resistant to ligand-dependent modulation with the CCR5 ligand CCL5/RANTES. Confocal laser microscopy revealed that CCR5 was colocalized with CXCR4 on the cell surface. In CD4 expressing CCR5 and CXCR4 double positive NIH 3T3 cells, immunoprecipitation followed by Western blot analysis revealed that CCR5 was associated with CXCR4 and CD4. CXCR4 and CCR5 were not co-immunoprecipitated in cells expressing CCR5 and CXCR4 but without CD4 expression. Compared to NIH 3T3CD4 cells expressing CXCR4, the entry of an HIV-1 X4 isolate (HCF) into NIH 3T3CD4 expressing both CXCR4 and CCR5 was reduced. Our data indicate that chemokine receptors interact with each other, which may modulate chemokine chemokine receptor interactions and HIV-1 coreceptor functions. PMID- 15660420 TI - The tyrosine phosphatase, OST-PTP, is expressed in mesenchymal progenitor cells early during skeletogenesis in the mouse. AB - Osteotesticular protein tyrosine phosphatase (OST-PTP; OST), is a signaling molecule which catalyzes the removal of phosphates from tyrosine residues. It is known to be highly regulated in bone cells and has been shown to be important for the in vitro progression from a preosteoblast to a mature, mineralizing cell. However, the in vivo expression of this phosphatase during skeletogenesis has not been examined. Using Northern analysis and in situ hybridization (ISH), we have observed that this gene is strongly expressed early during the formation of the mouse skeleton. By 12.5 days postcoitum (dpc), expression of OST mRNA transcripts increases and is localized within the mesenchyme of craniofacial bones, ribs, limbs, and Meckel's cartilage. Following initiation of chondrogenesis, OST mRNA becomes restricted to the perichondrium of all endochondral elements. With ossification, this gene is also expressed by cells, presumably osteoblasts, at the chondro-osseous border and along cortical and trabecular bone surfaces. Unlike other bone markers examined such as Osterix and type II collagen, OST transcripts do not appear to be expressed by chondrocytes of epiphyseal cartilage or by non-hypertrophic or hypertrophic chondrocytes. Because the temporal expression patterns of OST and Runx2 were similar suggesting a potential interrelationship in bone regulation and function, OST expression was examined in transgenic mice lacking a functional Runx2/Cbfal protein (Runx2/Cbfal delta C (deltaC)) and possessing a cartilaginous skeleton. Interestingly, the OST gene was expressed with localization similar in wild-type, homozygous, and heterozygous embryos. These studies suggest that the expression of the OST gene may be important during skeletogenesis, potentially from commitment of mesenchymal cells to the ossification of new bones. Early in embryogenesis, regulation of OST expression may be independent of Runx2/Cbfa1. PMID- 15660421 TI - In vivo activated caspase-3 cleaves PARP-1 in rat liver after administration of the hepatocarcinogen N-nitrosomorpholine (NNM) generating the 85 kDa fragment. AB - We reported previously that treatment of rats with the hepatocarcinogen N nitrosomorpholine (NNM) caused severe hepatotoxicity associated with apoptosis of hepatocytes beginning 12 h after administration of NNM. We observed that poly(ADP ribose) polymerase 1 (PARP-1), one of the major nuclear targets for caspases, was proteolytically degraded generating primarily 64 and 54 kDa fragments. Interestingly, at 20, 30, and 40 h post-treatment a 85 kDa cleavage product of PARP-1 resembling that generated by caspase-3 appeared additionally in hepatocytes. More detailed analysis performed in the present study revealed that the 85 kDa fragment of PARP-1 was generated in the liver in 10 of 17 (60%) animals examined between 20 and 40 h after NNM administration. The caspase-3 generated 85 kDa fragment was detected solely in hepatocytes undergoing apoptosis as evidenced by immunostaining performed with the antibody recognizing exclusively PARP-1 cleaved at position 214/215. The appearance of the 85 kDa fragment of PARP-1 in the liver nuclei coincided temporally with an significant increase of caspase-3 activity in hepatocytes. In contrast, in testis samples obtained from the same animals, no changes characteristic for apoptosis such as induction of caspases activity or degradation of nuclear PARP-1 could be detected. Our results evidence unequivocally that PARP-1 in liver is not resistant to caspases and can be processed in vivo by activated caspase-3 producing the p85 kDa fragment. Moreover, the caspase-3 induced PARP-1 fragmentation coinciding with the increase of caspase-3 activity was detected solely in the target organ and exclusively in hepatocytes undergoing apoptosis. Considering the fact that the caspase-3 mediated PARP-1 cleavage occurred only in 60% of animals tested between 20 and 40 h, it becomes obvious that the cellular response in vivo to the same trigger(s) strongly varies and may depend on a variety of intrinsic factors. It remains to elucidate which additional factors may be involved in the modulation of cellular response to the strong insults thereby activating different pathways and generating distinct outcomes. PMID- 15660422 TI - ABT1-associated protein (ABTAP), a novel nuclear protein conserved from yeast to mammals, represses transcriptional activation by ABT1. AB - Various TATA-binding protein (TBP)-associated proteins are involved in the regulation of gene expression through control of basal transcription directed by RNA polymerase (Pol) II. We recently identified a novel nuclear protein, activator of basal transcription 1 (ABT1), which binds TBP and DNA, and enhances Pol II-directed basal transcription. To better understand regulatory mechanisms for ABT1, we searched for ABT1-binding proteins using a yeast two-hybrid screening and isolated a cDNA clone encoding a novel protein termed ABT1 associated protein (ABTAP). ABTAP formed a complex with ABT1 and suppressed the ABT1-induced activation of Pol II-directed transcription in mammalian cells. Furthermore, ABTAP directly bound to ABT1, disrupted the interaction between ABT1 and TBP, and suppressed the ABT1-induced activation of Pol II-directed basal transcription in vitro. These two proteins colocalized in the nucleolus and nucleoplasm and were concomitantly relocalized into discrete nuclear bodies at higher expression of ABTAP. Taken together, these results suggest that ABTAP binds and negatively regulates ABT1. The ABT1/ABTAP complex is evolutionarily conserved and may constitute a novel regulatory system for basal transcription. PMID- 15660423 TI - Deliberate self-harm (DSH) among older people: a retrospective study in Barnet, North London. AB - BACKGROUND: Rates of suicide remain high among older people and those who deliberately self harm are believed to be at an increased risk of killing themselves in the future. If older people who deliberately harm themselves are to be helped by developments in services we need to understand what currently happens to them in terms of service provision and outcome. METHODS: A retrospective paper and electronic case note survey was carried out on all older people living in the London Borough of Barnet who presented to Accident & Emergency Departments with DSH over a two-year period. Ensuing actions and events were then tracked. RESULTS: Forty-three older people with DSH were identified. 18/43 (42%) had previous contact with local psychiatric services. The main method of DSH was overdose of medication (36/43 or 84%). Compared to the general population there were more women and widows. There were similar levels of physical ill-health. Thirty-seven of 43 (86%) received documented psychiatric input outside of hospital following the DSH. The mean follow-up period was 789.0 days (SD 419.8) and during this time 8/43 (19%) had a further documented episode of DSH, and 18/43 (40%) died from natural causes. CONCLUSIONS: This study confirms the need for improved documentation of DSH and its coding; this needs to be reviewed at local and national level. The vast majority of older people who attempt suicide do have subsequent contact with psychiatric services. There is a strong likelihood of repeat DSH and a higher risk of death by natural causes, emphasising the need to conceptualise DSH as a risk factor relevant to all medical specialities. PMID- 15660424 TI - Clinical application of operationalized criteria for 'Depression of Alzheimer's Disease'. AB - OBJECTIVES: 'Depression of Alzheimer's Disease' (dAD) is a common complication of Alzheimer's disease and is increasingly recognized as a syndrome with a clinical presentation differing from major depression. Criteria for the diagnosis of dAD have been proposed previously. METHODS: This paper presents these criteria in operationalized format designed to be accessible for clinical use. Four cases are discussed that demonstrate the use of these criteria and illustrate important differences between dAD and major depression. RESULTS: The dAD criteria are broader than DSM-IV criteria for Major Depressive Episode and incorporate caregiver input. CONCLUSIONS: Given the differences between dAD and major depression diagnoses, it is important to assess the efficacy of treatments for dAD. Depression in Alzheimer's Disease-2 (DIADS-2) is a controlled trial of dAD treatments that will also assess the validity of these criteria. PMID- 15660426 TI - Measurement of hepatic venous pressure gradient in patients with active variceal bleeding. PMID- 15660428 TI - p18(INK4c) expression in hepatocellular carcinoma. PMID- 15660429 TI - Reduced plasma adiponectin in NASH: central obesity as an underestimated causative risk factor. PMID- 15660430 TI - A challenge on the use of the words embryonic and perinatal in the context of biliary atresia. PMID- 15660431 TI - Preprocedure coagulation tests are unnecessary before abdominal paracentesis in emergency departments. PMID- 15660434 TI - The role of transjugular intrahepatic portosystemic shunt in the management of portal hypertension. PMID- 15660433 TI - Serum alanine aminotransferase in skeletal muscle diseases. AB - Although elevation of the levels of serum alanine aminotransferase (ALT) following liver injury is well known, confusion exists concerning skeletal muscle injury as the cause of this rise. We reviewed the records of 16 patients who had muscle necrosis without evidence of liver disease. The patients were divided into three groups: extreme exercise, polymyositis, and seizures. All patients exhibited markedly elevated creatine kinase and lactate dehydrogenase levels consistent with muscle injury. In acute cases, aspartate aminotransferase (AST) and ALT were both elevated, and the AST/ALT ratio was greater than 3, but this ratio approached 1 after a few days because of a faster decline in AST. In conclusion, this difference in half-life accounts for the comparable AST and ALT levels in our cases with chronic muscle injury. PMID- 15660435 TI - An integrated mBAND and submegabase resolution tiling set (SMRT) CGH array analysis of focal amplification, microdeletions, and ladder structures consistent with breakage-fusion-bridge cycle events in osteosarcoma. AB - Osteosarcoma (OS) is characterized by chromosomal instability and high-copy number gene amplification. The breakage-fusion-bridge (BFB) cycle is a well established mechanism of genomic instability in tumors and in vitro models used to study the origins of complex chromosomal rearrangements and cancer genome amplification. However, until now, there have been no high-resolution cytogenetic or genomic array studies of BFB events in OS. In the present study, multicolor banding (mBAND) FISH and submegabase resolution tiling set (SMRT) array comparative genomic hybridization (CGH) were used to identify and map genomic signatures of BFB events in four OS cell lines and one patient tumor. The expected intermediates associated with BFB-dicentric chromosomes, inverted duplications, and intra- and interchromosomal amplifications-were identified. mBAND analysis provided detailed mapping of rearrangements in 1p, 6p, and 8q and showed that translocation junctions were often in close proximity to fragile sites. More detailed mBAND studies of OS cell line MG-63 revealed ladderlike FISH signals of equally spaced interchromosomal coamplifications of 6p21, 8q24, and 9p21-p22 in a homogeneously staining region (hsr). Focal amplifications that concordantly mapped to the hsr were localized to discrete genomic intervals by SMRT array CGH. The complex amplicon structure in this hsr suggests focal amplifications immediately adjacent to microdeletions. Moreover, the genomic regions in which there was deletion/amplification had a preponderance of fragile sites. In summary, this study has provided further support for the role of the BFB mechanism and fragile sites in facilitating gene amplification and chromosomal rearrangement in OS. PMID- 15660437 TI - Bishop score and transvaginal ultrasound for preinduction cervical assessment: a randomized clinical trial. AB - OBJECTIVE: To compare transvaginal ultrasound with the Bishop score in assessment of cervical ripening for choice of induction agent. METHODS: Eighty women were randomized to have preinduction cervical assessment for choice of induction agent based on either Bishop score or transvaginal ultrasound. The primary outcome measure was the percentage of women who were administered prostaglandin as a preinduction agent. The criteria for considering the cervix as unripe and thus for using prostaglandin were either a Bishop score < 6 or a cervical length > 30 mm with cervical wedging of < 30% of the total cervical length. Secondary outcome measures included interval to active phase, interval to delivery and rate of Cesarean section. RESULTS: While 85% of women received prostaglandin in the Bishop score group, only 50% of them did in the transvaginal ultrasound group (P = 0.001). The interval to active phase, interval to delivery and rate of Cesarean section were similar in both groups. CONCLUSIONS: With the suggested cut-off values of a Bishop score < 6 or a cervical length > 30 mm and wedging < 30%, the use of transvaginal ultrasound instead of Bishop score for preinduction cervical assessment to choose induction agent significantly reduces the need for intracervical prostaglandin treatment without adversely affecting the success of induction. PMID- 15660436 TI - Oral administration of sildenafil restores learning ability in rats with hyperammonemia and with portacaval shunts. AB - Patients with liver disease with overt or minimal hepatic encephalopathy show impaired intellectual capacity. The underlying molecular mechanism remains unknown. Rats with portacaval anastomosis or with hyperammonemia without liver failure also show impaired learning ability and impaired function of the glutamate-nitric oxide-cyclic guanine monophosphate (glutamate-NO-cGMP) pathway in brain. We hypothesized that pharmacological manipulation of the pathway in order to increase cGMP content could restore learning ability. We show by in vivo brain microdialysis that chronic oral administration of sildenafil, an inhibitor of the phosphodiesterase that degrades cGMP, normalizes the function of the glutamate-NO-cGMP pathway and extracellular cGMP in brain in vivo in rats with portacaval anastomosis or with hyperammonemia. Moreover, sildenafil restored the ability of rats with hyperammonemia or with portacaval shunts to learn a conditional discrimination task. In conclusion, impairment of learning ability in rats with chronic liver failure or with hyperammonemia is the result of impairment of the glutamate-NO-cGMP pathway. Moreover, chronic treatment with sildenafil normalizes the function of the pathway and restores learning ability in rats with portacaval shunts or with hyperammonemia. Pharmacological manipulation of the pathway may be useful for the clinical treatment of patients with overt or minimal hepatic encephalopathy. PMID- 15660438 TI - A comparative study of the in vitro corrosion behavior and cytotoxicity of a superferritic stainless steel, a Ti-13Nb-13Zr alloy, and an austenitic stainless steel in Hank's solution. AB - In this study, the in vitro corrosion resistance of a superferritic stainless steel in naturally aerated Hank's solution at 37 degrees C has been determined to evaluate the steel for use as a biomaterial. The potentiodynamic polarization method and electrochemical impedance spectroscopy (EIS) were used to determine the corrosion resistance. The polarization results showed very low current densities at the corrosion potential and electrochemical behavior typical of passive metals. At potentials above 0.75 V (SCE), and up to that of the oxygen evolution reaction, the superferritic steel exhibited transpassive behavior followed by secondary passivation. The superferritic stainless steel exhibited high pitting resistance in Hank's solution. This steel did not reveal pits even after polarization to 3000 mV (SCE). The EIS results indicated high impedance values at low frequencies, supporting the results obtained from the polarization measurements. The results obtained for the superferritic steel have been compared with those of the Ti-13Nb-13Zr alloy and an austenitic stainless steel, as Ti alloys are well known for their high corrosion resistance and biocompatibility, and the austenitic stainless steel is widely used as an implant material. The cytotoxicity tests indicated that the superferritic steel, the austenitic steel, and the Ti-13Nb-13Zr alloy were not toxic. Based on corrosion resistance and cytotoxicity results, the superferritic stainless steel can be considered as a potential biomaterial. PMID- 15660439 TI - Rigid nonproteinogenic cyclic amino acids as ligands for glutamate receptors: trans-tris(homoglutamic) acids. AB - The second-generation asymmetric synthesis of the trans-tris(homoglutamic) acids reported herein proceeds via Strecker reaction of chiral ketimines, obtained from condensation of racemic 2-ethoxycarbonylmethylcyclopentanone and commercially available (S)- and (R)-1-phenylethylamine, respectively. In the key stereodifferentiating step, the cyanide addition leads to mixtures of diastereomeric alpha-amino nitrile-esters, the composition of which is independent of the reaction temperature and the type of the solvent, respectively. Hydrolysis of the alpha-amino nitrile-esters with concentrated H(2)SO(4) yielded diastereomeric mixtures of secondary alpha-amino amido-esters, which after separation were hydrogenolyzed and hydrolyzed each to the enantiomeric trans-1-amino-2-carboxymethylcyclopentanecarboxylic acids. Their configuration was completely established by NMR methods, CD spectra, and X-ray analysis of the trans-1S,2R-configured secondary alpha-amino amido-ester. In receptor binding assays and functional tests, trans-1S,2R-1-amino-2 carboxymethylcyclopentanecarboxylic acid hydrochloride was found to behave as a selective mGluR(2)-antagonist without relevant binding properties at iGluRs. PMID- 15660440 TI - Synthesis, absolute configuration, and application of enantiopure trans-1 aminobenz[f]indan-2-ol. AB - Both novel enantiopure trans-1-aminobenz[f]indan-2-ols (4) were obtained from the racemate by the diastereomeric salt formation with (+)- and (-)-dibenzoyltartaric acids (8), respectively, and the absolute configuration of the enantiomer 4 in the less-soluble diastereomeric salt of racemic 4 with (+)-8 was determined to be (1S,2S) by an X-ray crystallographic analysis. The chiral recognition ability of the enantiopure amino alcohol was examined for the enantioseparation of racemic 2 arylalkanoic acids by the diastereomeric salt formation. The role of the naphthylene group of the amino alcohol was found to be closely associated with the stabilization of the crystal by CH/pi interactions on the basis of an X-ray crystallographic study. PMID- 15660441 TI - The effects of serial intravascular transfusions in ascitic/hydropic RhD alloimmunized fetuses. AB - OBJECTIVE: To evaluate the effects of serial intravascular transfusions on RhD alloimmunized fetuses with ascites/hydrops at the time of the first transfusion by measuring multiple hematological/biochemical blood variables. METHODS: Thirty one singleton pregnancies were referred for management of RhD alloimmunization. Seven fetuses had hydrops on presentation and were transfused immediately. The remainder underwent weekly ultrasound examinations, and fetal blood sampling and transfusion were performed on development of ascites. In the 104 samples collected overall from the 31 fetuses, glucose, uric acid, urea, creatinine, total protein, total and direct bilirubin, aspartate aminotransferase (AST), alanine aminotransferase (ALT), gamma-glutamyltransferase, alkaline phosphatase, lactic dehydrogenase, amylase, pseudocholinesterase (PCHE), creatine kinase, triglycerides and cholesterol were measured and compared with a reference range for non-anemic fetuses. RESULTS: The median gestational age at first transfusion was 26 (range, 18-34) weeks. There were three fetal losses after the first transfusion, two of which were due to procedure-related complications; one further loss occurred. At the first transfusion fetal hematocrit, pO2, total protein, PCHE, creatinine and urea concentrations were significantly decreased compared to reference data, while total and direct bilirubin, AST, ALT, amylase, triglyceride and uric acid concentrations were increased. In all surviving fetuses ascites/hydrops had disappeared by the second transfusion. Fetal pO2, total protein, AST, ALT and PCHE concentrations had normalized by the third transfusion. Correction of fetal anemia did not affect the other variables. CONCLUSIONS: RhD-alloimmunized fetuses with ascites/hydrops at the time of the first transfusion had a survival rate of 87%. Alterations of several biochemical fetal blood indices are present at the first sampling/transfusion, but most variables normalize with intravascular transfusions. PMID- 15660442 TI - Structural accelerated failure time models for survival analysis in studies with time-varying treatments. AB - BACKGROUND: In the absence of unmeasured confounding factors and model misspecification, standard methods for estimating the causal effect of time varying treatments on survival are biased when (i) there exists a time-dependent risk factor for survival that also predicts subsequent treatment and (ii) past treatment history predicts subsequent risk factor level. In contrast, structural models provide consistent estimates of causal effects when unmeasured confounding and model misspecification are absent. The parameters of nested structural models are estimated by g-estimation and those of marginal structural models by inverse probability weighting. METHODS: We describe a nested structural accelerated failure time model and use it to estimate the total causal effect of highly active antiretroviral therapy (HAART) on the time to AIDS or death among human immunodeficiency virus (HIV)-infected participants of the Multicenter AIDS Cohort and Women's Interagency HIV Studies. The Appendix describes g-estimation and methods to deal with censoring. RESULTS: Comparing the regime 'always treated' to 'never treated,' the AIDS-free survival time ratio was 2.5 (95% confidence interval [CI]: 1.7, 3.3). CONCLUSIONS: Our finding of a strongly beneficial effect is consistent with results from randomized trials and from a previous analysis of the same data using a marginal structural Cox model. In contrast, a previous analysis using a standard (non-structural) model did not find an effect of treatment on survival. PMID- 15660443 TI - Oxidative stress and uremia. AB - Oxidative stress is a pathogenic element of great importance in uremic patients, with a great impact on their survival. The cause of oxidative stress in patients on hemodialysis is traditionally attributed to the recurrent activation of polymorphonucleate neutrophils and monocytes. The effects of oxidative stress are evident on all biochemical components of biological tissues: lipids, proteins, carbohydrates, and nucleic acids. This study briefly reviews the effects of different dialytic techniques and of kidney transplant on several parameters of oxidative stress. Many different modalities of pharmaceutical intervention are then analyzed, and the clinical evidences reported. PMID- 15660444 TI - Stimulation of neurite outgrowth in a human nerve scaffold designed for peripheral nerve reconstruction. AB - The clinical outcome of microsurgical reconstruction of large peripheral nerve lesions depends on the availability of suitable graft material. Allogenic nerve grafts are rejected by the immune system. Extracellular-matrix proteins, in contrast to the resident cells, are of low immunogenicity in allografts. Here, human tibial nerve segments were extracted with lysophosphatidyl choline. The obtained cell-free and myelin-free scaffold consisted of empty endoneural tubes with maintained extracellular matrix architecture. The nerve scaffold had mechanical properties comparable to intact nerve, making it suitable for microsurgical reconstruction. Sections of the nerve scaffold were tested as a substrate for the adhesion and neuronal differentiation of human neuroblastoma derived LAN-5 cells. Nerve extraction removed laminin-2, an isoform of laminin important for peripheral nerve regeneration. Laminin-2 reloading of the nerve scaffold did not improve cell adhesion and axon growth. Chemical crosslinking of heparan sulfate, on the other hand, increased the percentage of adherent cells with outgrowing neurites from 48 to 85%. Combined laminin-2 reloading and heparan sulfate crosslinking reduced the percentage of neurite-forming cells to 22% of the number of adherent cells. Implantation of the nerve scaffold into the peritoneal cavity of mice was not cytotoxic, and neovascularization of the graft material was observed within weeks. In conclusion, extraction of human nerve with detergents revealed a biocompatible nerve scaffold supporting neuronal cell adhesion. Heparan sulfate crosslinking to the scaffold surface improved neurite outgrowth, presumably mediated by midkine, a member of the neurokine family of growth factors, which is secreted by neuroblastoma-derived cells and binds to heparan sulfate. PMID- 15660445 TI - Simple ultrasound evaluation of the anal sphincter in female patients using a transvaginal transducer. AB - OBJECTIVE: Fecal incontinence affects 0.2% of women aged 15-64 years and about 1.3% of women over 64 years. Most cases are related to instrumental deliveries affecting the anal sphincter complex. We propose a simple technique using the generally available transvaginal transducer to evaluate the anal sphincter complex. METHODS: Ninety-two patients underwent ultrasound examination. Group I consisted of 53 nulliparous patients. In Group II there were six patients with normal spontaneous vaginal deliveries without episiotomies. In Group III there were 14 patients with vaginal deliveries and one to three episiotomies but no lacerations. In Group IV there were nine postpartum patients with recently repaired (48 h to 3 weeks) third- and fourth-degree lacerations. All women in Groups I-IV were asymptomatic. Group V consisted of 10 patients symptomatic for fecal incontinence. We used a vaginal probe (5-9-MHz) with the footprint placed in the fourchette pointing towards the anus in a transverse and then in a median (sagittal) plane. If seen, the combined internal and external anal sphincter thickness at the 12 o'clock location was measured. We visualized normal star shaped mucosal folds on the transverse section and described the sonographic anatomy in both planes. RESULTS: The mean sphincter thickness measured at 12 o'clock in Group I was 2.3 (range, 1.0-4.7) mm, in Group II it was 2.9 (range, 2.4-3.4) mm, and in Group III it was 2.3 (range, 1.0-3.7) mm. The differences between these three groups were not significant. Patients from Group IV showed thinning or discontinuous sphincter anatomy at the 12 o'clock position. All symptomatic patients from Group V showed abnormal sphincter anatomy, and the normal star-like appearance of the anal mucosa on the transverse section was deformed, radiating from the point of the sphincter damage. Four of the 10 patients in this group underwent surgical repair. In these patients the sonographic findings were confirmed. CONCLUSIONS: The images obtained using this imaging modality show the sphincter muscle anatomy as well as the possible pathology. Due to its simplicity the technique can be applied in any place where a vaginal transducer is available. PMID- 15660446 TI - Reliability of the lung-to-head ratio in predicting outcome and neonatal ventilation parameters in fetuses with congenital diaphragmatic hernia. AB - OBJECTIVES: The ratio of the lung area (on the contralateral side of the hernia) to the head circumference, the lung-to-head ratio (LHR), has been proposed as a reliable tool in the assessment of the prognosis of congenital diaphragmatic hernia (CDH). An LHR < 0.6 has been associated with poor outcome whereas one > 1.4 has been associated with survival. We aimed to analyze the role of LHR in predicting fetal outcome and ventilation parameters in cases of isolated CDH in our center. METHODS: During the 40-month study period, 22 fetuses with an isolated CDH were delivered alive under optimized conditions at our perinatal center. LHR was measured at the time of diagnosis (median, 27 weeks' gestation). In addition to survival, postnatal ventilation parameters including pCO2, pO2, inspiratory O2 partial pressure, inspiratory pressure and oxygenation index were determined, as was the occurrence of pulmonary hypertension. RESULTS: The overall survival rate was 59%. The LHR was not able to predict fetal outcome reliably. There was no correlation between the liver herniation, LHR and fetal outcome. The prenatally determined lung size reflected in the LHR did not show any significant association with individual ventilation parameters. Eleven of 17 infants examined had signs of pulmonary hypertension and the LHR did not predict this condition. CONCLUSION: Our study cannot support the optimistic results reported by other groups on the use of LHR as a reliable predictor of outcome in fetuses with CDH. The LHR, as a reflection of lung size, correlates neither with survival patterns nor with various postnatal ventilation parameters. PMID- 15660447 TI - Congenital renal tract anomalies: outcome and follow-up of 402 cases detected antenatally between 1986 and 2001. AB - OBJECTIVE: To determine the long-term prognosis of antenatally detected renal tract anomalies in order to optimize parental counseling. METHODS: This was a follow-up study of all renal tract abnormalities detected antenatally in a Level 3 ultrasound department between 1986 and 2001. Follow-up data (median age, 8 years) were retrieved from the records of the Paediatric Urology Department or the attending pediatrician. RESULTS: A urinary tract anomaly was detected in 408 fetuses. There were four false-positive diagnoses. From two children follow-up data were incomplete, leaving 402 cases for analysis. A chromosomal abnormality was present in 7/81 (8.6%) fetuses that had karyotyping. Termination of pregnancy was performed in 55 (13.7%) cases and a further 66 (16.4%) children died during the perinatal period and up to 1 year of age. In 106/121 (26.4% of all fetuses) deceased children the cause of death was directly related to the renal tract anomaly. In the 281 surviving children a total of 545 renal tract anomalies were diagnosed postnatally, requiring a total of 381 surgical interventions in 156 infants. Outcome in survivors was generally good, with impaired renal function in nine infants and hypertension in three (4% of the survivors). CONCLUSIONS: Congenital renal tract anomalies are associated with a high mortality rate, especially when they are structural developmental anomalies of the kidneys. Survivors require multiple operations, but the outcome is generally favorable. Ultrasound diagnosis, especially when made early, of non-lethal urinary tract anomalies may prevent additional renal damage by timing of delivery and early postnatal treatment. PMID- 15660448 TI - Improvement of arthroscopic cartilage stiffness probe using amorphous diamond coating. AB - During arthroscopic evaluation of articular cartilage unstable contact and even slipping of the measurement instrument on the tissue surface may degrade the reproducibility of the measurement. The main aim of the present study was to achieve more stable contact by controlling the friction between articular cartilage surface and the arthroscopic cartilage stiffness probe (Artscan 200, Artscan Oy, Helsinki, Finland) using amorphous diamond (AD) coating. In order to obtain surfaces with different average roughnesses (R(a)), polished stainless steel disks were coated with AD by using the filtered pulsed arc-discharge (FPAD) method. Dynamic coefficient of friction (mu) between the articular cartilage (n = 8) and the coated plates along one non-coated plate was then determined. The friction between AD and cartilage could be controlled over a wide range (mu = 0.027-0.728, p < 0.05, Wilcoxon test) by altering the roughness. Possible deterioration of cartilage was investigated by measuring surface roughness after friction tests and comparing it with the roughness of the adjacent, untested samples (n = 8). Importantly, even testing with the roughest AD (R(a) = 1250 nm) did not damage articular surface. On the basis of the friction measurements, a proper AD coating was selected for the stiffness probe. The performance of coated and non-coated probe was compared by measuring bovine osteochondral samples (n = 22) with both instruments. The reproducibility of the stiffness measurements was significantly better with the AD-coated probe (CV% = 4.7) than with the uncoated probe (CV% = 8.2). To conclude, AD coating can be used to safely control dynamic friction with articular surface. Sufficient friction between articular surface and reference plate of the arthroscopic probe improves significantly reproducibility of the stiffness measurements. PMID- 15660449 TI - Molecular dynamics (MD) investigations of preformed structures of the transmembrane domain of the oncogenic Neu receptor dimer in a DMPC bilayer. AB - The critical Val/Glu mutation in the membrane spanning domain of the rat Neu receptor confers the ability for ligand-independent signaling and leads to increased dimerization and transforming ability. There is evidence that the two transmembrane interacting helices play a role in receptor activation by imposing orientation constraints to the intracellular tyrosine kinase domains. By using MD simulations we have attempted to discriminate between correct and improper helix helix packing by examining the structural and energetic properties of preformed left-handed and right-handed structures in a fully hydrated DMPC bilayer. The best energetic balance between the residues at the helix-helix interface and the residues exposed to the lipids is obtained for helices in symmetrical left-handed interactions packed together via Glu side chain/Ala backbone interhelical hydrogen bonds. Analyses demonstrate the importance of the ATVEG motif in helix helix packing and point to additional contacting residues necessary for association. Our findings, all consistent with experimental data, suggest that a symmetrical left-handed structure of the helices could be the transmembrane domain configuration that promotes receptor activation and transformation. The present study may provide further insight into signal transduction mechanisms of the ErbB/Neu receptors. PMID- 15660450 TI - Time-domain semi-parametric estimation based on a metabolite basis set. AB - A novel and fast time-domain quantitation algorithm--quantitation based on semi parametric quantum estimation (QUEST)--invoking optimal prior knowledge is proposed and tested. This nonlinear least-squares algorithm fits a time-domain model function, made up from a basis set of quantum-mechanically simulated whole metabolite signals, to low-SNR in vivo data. A basis set of in vitro measured signals can be used too. The simulated basis set was created with the software package NMR-SCOPE which can invoke various experimental protocols. Quantitation of 1H short echo-time signals is often hampered by a background signal originating mainly from macromolecules and lipids. Here, we propose and compare three novel semi-parametric approaches to handle such signals in terms of bias variance trade-off. The performances of our methods are evaluated through extensive Monte-Carlo studies. Uncertainty caused by the background is accounted for in the Cramer-Rao lower bounds calculation. Valuable insight about quantitation precision is obtained from the correlation matrices. Quantitation with QUEST of 1H in vitro data, 1H in vivo short echo-time and 31P human brain signals at 1.5 T, as well as 1H spectroscopic imaging data of human brain at 1.5 T, is demonstrated. PMID- 15660451 TI - Reactions of (di)manganese carbonyl ions with 1,4,7-trimethyl-1,4,7 triazacyclononane (Me3TACN) in the gas phase. AB - The gas-phase reactions of a series of (di)manganese carbonyl positive ions with 1,4,7-trimethyl-1,4,7-triazacyclononane (Me(3)TACN) have been examined with the aid of Fourier transform ion cyclotron resonance (FTICR) mass spectrometry. The monomanganese carbonyl ions, [Mn(CO)(n)](+) (n = 2-5), react predominantly by ligand exchange and to a minor extent by electron transfer with the formation of the radical cation of Me(3)TACN. For the [Mn(CO)(n)](+) (n = 2-4) ions, the ligand exchange results in the exclusive formation of a [Mn(Me(3)TACN)](+) complex, whereas small amounts of [Mn(CO)(Me(3)TACN)](+) ions are also generated in the reactions of the [Mn(CO)(5)](+) ion. The [Mn(2)(CO)(n)](+) ions (n = 2, 4 and 5) react also by competing electron transfer and ligand exchange. The reaction of the [Mn(2)(CO)(2)](+) and [Mn(2)(CO)(4)](+) ions is associated with cleavage of the Mn--Mn bond as evidenced by the pronounced formation of [Mn(Me(3)TACN)](+) ions. For [Mn(2)(CO)(5)](+), the ligand exchange leads mainly to the formation of [Mn(2)(CO)(n)(Me(3)TACN)](+) (n = 1-3) ions. These primary product ions react subsequently by the incorporation of a second Me(3)TACN molecule to afford [Mn(2)(CO)(Me(3)TACN)(2)](+) and [Mn(2)(CO)(2)(Me(3)TACN)(2)](+) ions. Both of these latter species incorporate an oxygen molecule with formation of ions with the assigned composition of [Mn(2)(O(2))(CO)(Me(3)TACN)(2)](+) and [Mn(2)(O(2))(CO)(2)(Me(3)TACN)(2)](+). PMID- 15660452 TI - Taking stock of herbicide-resistant crops ten years after introduction. AB - Since transgenic, bromoxynil-resistant cotton and glufosinate-resistant canola were introduced in 1995, planting of transgenic herbicide-resistant crops has grown substantially, revolutionizing weed management where they have been available. Before 1995, several commercial herbicide-resistant crops were produced by biotechnology through selection for resistance in tissue culture. However, non-transgenic herbicide-resistant crops have had less commercial impact. Since the introduction of glyphosate-resistant soybean in 1996, and the subsequent introduction of other glyphosate-resistant crops, where available, they have taken a commanding share of the herbicide-resistant crop market, especially in soybean, cotton and canola. The high level of adoption of glyphosate-resistant crops by North American farmers has helped to significantly reduce the value of the remaining herbicide market. This has resulted in reduced investment in herbicide discovery, which may be problematic for addressing future weed-management problems. Introduction of herbicide-resistant crops that can be used with selective herbicides has apparently been hindered by the great success of glyphosate-resistant crops. Evolution of glyphosate-resistant weeds and movement of naturally resistant weed species into glyphosate-resistant crop fields will require increases in the use of other herbicides, but the speed with which these processes compromise the use of glyphosate alone is uncertain. The future of herbicide-resistant crops will be influenced by many factors, including alternative technologies, public opinion and weed resistance. Considering the relatively few recent approvals for field testing new herbicide-resistant crops and recent decisions not to grow glyphosate-resistant sugarbeet and wheat, the introduction and adoption of herbicide-resistant crops during the next 10 years is not likely to be as dramatic as in the past 10 years. PMID- 15660453 TI - Frontiers in inflammatory joint diseases. Proceedings of a Canadian arthritis research forum. May 7-9, 2004. Toronto, Canada. PMID- 15660454 TI - Common denominators of inflammatory joint diseases. AB - Inflammatory joint diseases (IJD) are a heterogeneous group of disorders whose primary pathologic target is articular and periarticular tissue. Although each form of IJD is clinically distinct, these disorders share a number of common clinical, epidemiological, and pathogenetic elements. In all cases, there is a complex interaction between genetic and environmental factors that serves to initiate the process, and likely, there is a different set of interactions to perpetuate the arthritis, as well as determine the destructiveness of the process. There are a number of intervention checkpoints common to all forms of IJD. These include pri-mary prevention by identifying populations at risk and environmental triggers; prevention of persistent synovitis, articular damage, and functional disability; and restoration of function. Research and innovation directed at each of these key intervention checkpoints will improve the care of these disorders, and ultimately save valuable health care resources. PMID- 15660455 TI - Animal models of arthritis. What have we learned? AB - Animal models of arthritis can be used to understand elements of the arthritic process in patients. Recent therapeutic approaches in patients with rheumatoid arthritis (RA) with biologics are based on initial findings in murine models of experimental arthritis, although final proof of concept must come from clinical studies. Animal models are powerful tools for studying pathologic changes in articular cartilage and bone in great detail, and can be used to evaluate mechanisms of erosive processes. Although in general more inflammation drives more destruction, the uncoupling of inflammation and erosion can be seen as well, and different mediators are involved in these processes. PMID- 15660456 TI - Genetic studies in the rheumatic diseases: present status and implications for the future. AB - Recent breakthroughs in genetic methodology have greatly augmented our understanding of the contribution of genetics to susceptibility to the rheumatic diseases. Disorders in which familial aggregation has been best documented include rheumatoid arthritis (RA), ankylosing spondylitis (AS), systemic lupus erythematosus (SLE), and systemic sclerosis (SSc). Much of the genetic contribution to these diseases lies in the MHC, including HLA-DR4 (RA), HLA-B27 (AS), HLA-DRB1*0301, DRB1*1501/*1503, DRB1*08, and C4 null alleles (SLE), and HLA DRB1*11 and DRB1*1502 (SSc). Genome-wide scans have provided inconsistent data in RA, although consistent regions have been observed in scans from different groups in AS and SLE. No consistent non-MHC candidate gene has been identified in RA. There is active investigation in AS in this area. In SLE the Fc gamma RIIa and Fc gamma IIIa genes have been most thoroughly described, and in SSc fibrillin and SPARC. Newer techniques being developed presently, such as high density single nucleotide polymorphism genome-wide scanning, show promise to bring these analyses to the next level, which will hopefully result not only in better screening of individuals at highest risk, but also in novel treatments. PMID- 15660458 TI - Gene therapy: what have we accomplished and where do we go from here? AB - As a potential treatment for arthritis, gene transfer should be viewed within the context of biological therapy. Its particular strengths include the ability to deliver therapeutic gene products, both RNA and protein, to specific cells or tissues in a targeted, sustained, and potentially regulated, cost-effective fashion. An expanded definition of gene therapy includes the delivery of noncoding nucleotide sequences that act, for example, as decoy molecules. Considerable experimental progress has been made in the preclinical development of gene therapies for arthritis. Indeed, there is overwhelming proof of principle in animal models of rheumatoid arthritis (RA) and accumulating evidence of efficacy in animal models of osteoarthritis (OA). Early-phase human clinical trials have been successfully conducted and others are in progress. Additional research is necessary to optimize gene transfer technologies and achieve regulated transgene expression. However, the most urgent need is for interventional studies in human disease and the funding with which to implement them. PMID- 15660457 TI - Rheumatoid arthritis initial therapy: unanswered questions. AB - Recent changes in the treatment of patients with rheumatoid arthritis (RA) are high lighted in this article,with emphasis on initial therapy in RA. Critical new concepts and developments are discussed: the importance of early treatment, increased use of combination disease modifying antirheumatic therapy as initial treatment and after failure of monotherapy, the role of the newer biological therapies, and the influence of comorbidity on morbidity and mortality. PMID- 15660459 TI - What else can I do but take drugs? The future of research in nonpharmacological treatment in early inflammatory arthritis. AB - Nonpharmacological treatments, including physiotherapy and occupational therapy, have assumed a complementary role to drug therapy in managing inflammatory arthritis. Clinicians and researchers are facing 3 major challenges concerning the use of these treatments. First, strong evidence is only present in a few nonpharmacological interventions, such as exercise, patient education, and low level laser in the treatment of rheumatoid arthritis. The evidence on the majority of interventions is, however, weak or inconclusive. Second, knowledge is lacking on the elements associated with models of nonpharmacological care. The multidisciplinary team approach has been viewed as the standard for arthritis treatment; however, the team structure and the communication style among team members vary around the world. The influence of these elements on treatment success remains unclear. Finally, disparities in knowledge management and translation in nonpharmacological research have hindered the clinical use of these treatments and the growth of research in the field. To address the challenges, the author is recommending 4 research priorities for nonpharmacological treatments: 1. Evaluation of less well-studied interventions; 2. Understanding the relationships among rehabilitation-related variables and disability; 3. Development and evaluation of innovative care models; and 4. Design and evaluation of knowledge transfer innovations. PMID- 15660460 TI - Analyzing synovial tissue samples. What can we learn about early rheumatoid arthritis, the heterogeneity of the disease, and the effects of treatment? AB - The synovium is the key target of the disease process in rheumatoid arthritis (RA). Examination of synovial tissue samples may provide insight into the events that take place in different phases of the disease and may help to decipher the mechanism of action of antirheumatic treatment. This review describes the features of synovitis in early RA, which clearly represent chronic inflammation. There is marked interindividual variability, suggesting that RA consists of different pathogenetic subsets. Evaluation of serial synovial tissue samples has shown that effective treatment is associated with a reduction in synovial macrophages, independent of the specific mechanism of action of the compound. This suggests that these are key effector cells in the pathogenesis. In addition, it provides a biomarker that can be used in clinical trials. PMID- 15660461 TI - Clinical research and the new public partnership--a view from the South. AB - The clinical research enterprise in the United States is in a state of considerable transition. Many, including the US National Institutes of Health through its Roadmap Initiative, have called for a reengineering of the enterprise in ways that will build transparency, public disclosure of trial results, and enhanced public trust. This presentation will set forth a premise for a new public partnership model that will define the clinical research enterprise, in terms of both the relationship between the participant and the investigator in each clinical trial, and the relationship between the public and the clinical research enterprise as a whole. Ten guiding principles to direct this new partnership relationship are suggested. PMID- 15660462 TI - Longitudinal cohort studies. AB - Rheumatological disorders are complex conditions characterized by a variety of clinical manifestations and courses in different patients with the same condition. The best way to understand the course and prognosis of these patients is through longitudinal observational cohort studies. Such studies have been facilitated by computer technology that allows tracking of large numbers of patients and visits and analysis of a large amount of data. There are requirements for such datasets to be informative: internal and external consistency; clearly defined methods of observation and measurement; complete followup; consideration of potential confounders. We describe the usefulness of an observational cohort analysis using the University of Toronto Psoriatic Arthritis Clinic Database as a model. Analysis issues are highlighted and proposed mechanisms offered for keeping patients involved to ensure complete followup is maintained. PMID- 15660463 TI - Treating children with arthritis: towards an evidence-based culture. AB - We live in a culture of evidence-based medicine. Many areas of medicine have embraced this culture. However, for unusual diseases, like the childhood arthritides, there is little evidence. To provide this evidence a culture change must occur in pediatric rheumatology. The most convincing reason to make this change comes from the field of childhood oncology. Through successive clinical trials, collaborative oncology study groups have discovered cures for many childhood cancers. The most convincing studies are randomized trials; however, these are difficult to do. Collaborative trial groups and innovative designs are needed for an acceptable culture change in childhood arthritis. Recently a number of collaborations have been developed to help further the study of pediatric rheumatology. The best known are the Pediatric Rheumatology Collaborative Study Group in North America, and the Paediatric Rheumatology International Trials Organization in Europe, South America, and Asia. A new North American collaborative study group has formed--the Childhood Arthritis and Rheumatology Research Alliance (CARRA)--to undertake investigator-initiated clinical trials. These groups might potentially lead the way to a new evidence-based culture for childhood arthritis. PMID- 15660464 TI - Is early intervention worth it? AB - Historically, the largest components of costs associated with rheumatoid arthritis (RA), the most common inflammatory rheumatic disease, were hospitalizations, principally for joint replacement surgery, and work loss. Thus, for expensive interventions such as biological agents to be "worthwhile," they must reduce the prevalence of joint replacement and assist persons with RA in maintaining employment. However, joint replacement surgery and work losses tend to occur at least several years after onset of disease, even in severe cases. Assessing the cost-effectiveness of expenditures becomes computationally and politically difficult when the expenditure and the outcome are separated in time. The computational issue concerns the translation of future benefits--surgeries avoided and jobs held onto years from now--into present monetary values. The computational issue may be even more complex when the benefits are less tangible than surgery and wages; for example, when measured by quality-adjusted life years. The political issue concerns the disjuncture between the agents making the expenditures--provincial health insurance in Canada or an employer's health plan in the US--and the agents reaping the benefits, a private disability insurance company or provincial or state workers' compensation fund. In addition, there is an ethical dilemma. In the US, many of the advances in the care for RA such as the biological agents derive, at least in part, from federal research expenditures. Such expenditures are financed by increasingly regressive taxes. Yet the individuals bearing an increasing share of the tax burden find themselves relegated to more restrictive health insurance plans less likely to provide access to those agents. Thus, whether expenditures for early interventions are worthwhile may turn on such issues as how long the expenditure and the benefits are separated in time, how well the interests of the agent making the expenditure and the agent reaping the rewards are aligned, and how equitable the financing of the benefit and the access to it. PMID- 15660465 TI - Arthritis in Canada: what do we know and what should we know? AB - Doctors' visits for inflammatory arthritis reportedly represent only 6% of the overall visit rates for all arthritis and related conditions (163 per 1000), with about 40% of these patients seeing a specialist. Data from provincial drug plan databases show that although the proportion of the population aged 65 years and older with prescriptions for disease modifying antirheumatic drugs increased to 1% in 2000, this is only half the expected prevalence of rheumatoid arthritis in this age group. There are large provincial variations. Despite data on the efficacy and importance of treatment of early inflammatory arthritis, research is lacking on: the experience of arthritis, decision-making about seeking medical care, and factors affecting access to, and payment for, treatment, including drugs and rehabilitation; primary care decision-making about referral and treatment; organizational aspects of specialist care and access to drugs; and new ways of delivering services to reach patients in underserviced or remote areas. Monitoring the population impact of arthritis, including economic costs, is a priority for research, as are epidemiological studies on risk factors. PMID- 15660466 TI - Arthritis and employment research: where are we? Where do we need to go? AB - Studies of work disability among individuals with arthritis reveal that loss of employment is a common, important, and costly problem. Arthritis and musculoskeletal conditions are the leading cause of longterm work disability in Canada and the US, with an estimated yearly cost of 13.7 billion dollars in Canada. In rheumatoid arthritis, reported rates of work disability are remarkably high, ranging from 32% to 50% 10 years after RA onset, and increasing to 50% to 90% after 30 years. Studies have shown that work disability starts early in the course of RA, emphasizing the need for early intervention. To date, research in the area of arthritis and employment has mostly focused on measuring the extent of the problem and on identifying predictors of work loss. Despite the importance of the problem, there has been little intervention research assessing the effectiveness of medical treatment and few interventions specifically aimed at employment, reducing work loss, or improving ability to work. Research needed includes evaluating the effect of current therapies on employment outcomes, and studying interventions specifically aimed at employment, as well as addressing methodological issues in employment research. PMID- 15660467 TI - The anti-tumor necrosis factor agents are a major advance in the treatment of rheumatoid arthritis. AB - Despite optimization of the use of the traditional disease modifying antirheumatic drugs (DMARD) as mono- or combination therapy, between 25% and 50% of patients with rheumatoid arthritis (RA) still have clinically active synovitis with progressive structural articular damage. Several clinical trials with the anti-tumor necrosis factor (TNF) agents etanercept, infliximab, and adalimumab, used alone or in combination with methotrexate, in patients with early or established RA have shown consistent improvement in signs and symptoms as well as function, with a dramatic slowing of radiographic damage. The anti-TNF agents combined with optimal doses of MTX currently constitute the best therapeutic regimen for the management of patients with RA failing to adequately respond to traditional DMARD. PMID- 15660468 TI - Assessing the safety of biologic therapies in rheumatoid arthritis: the challenges of study design. AB - Clinical trials have shown the anti-tumor necrosis factor-a (TNF-a) drugs to be safe and efficacious for the treatment of rheumatoid arthritis (RA). However, since their release for general use, reports have raised concerns about potentially serious complications including tuberculosis, lymphoma, and cardiac failure. It must be remembered that patients with RA are already at increased risk of many of these complications,due both to their underlying inflammatory disease activity and the immunosuppressing effects of many conventional disease modifying antirheumatic drugs. It is unknown whether anti-TNF-a therapies are putting patients at increased risk of adverse events above what might already be expected. Data on the frequency of these adverse events have come predominantly from 3 sources: followup of subjects recruited to clinical trials, spontaneous adverse event reporting to national pharmacovigilance systems, and surveillance of patients treated in routine practice. Each of these study designs plays an important role in assessment of new drugs. However, each also has limitations, which must be considered when interpreting adverse event rates. PMID- 15660469 TI - Cost-effectiveness estimates reported for tumor necrosis factor blocking agents in rheumatoid arthritis refractory to methotrexate--a brief summary. AB - In the climate of rising healthcare expenditures the economic evaluation of new therapies becomes increasingly important in decision-making by health authorities. This article highlights some of the considerations regarding the economic assessment of drug treatments as they relate to rheumatic diseases, with emphasis on new biologic therapies such as tumor necrosis factor inhibitors. PMID- 15660470 TI - Integrating biologic therapy into the comprehensive care of patients with rheumatoid arthritis. AB - The advent of biologic therapy has not only provided the opportunity for better care of patients with rheumatoid arthritis (RA), but also has permitted a better understanding of the pathogenesis of this autoimmune/inflammatory disease. The capacity of these agents to suppress signs and symptoms as well as radiographic progression of RA strongly indicates that they can alter the course of the disease. Appropriate analysis of the effect of biologics should provide new insight into the role of the specific targeted molecules in rheumatoid inflammation, and provide information about means to optimize therapy with these highly potent therapeutics. PMID- 15660471 TI - Setting research priorities for arthritis: the environmental perspective. AB - Recent enthusiasm for genetic advances in prevention is out of keeping with the etiology of most common diseases of the industrialized world, including the major inflammatory arthritides. These conditions have a genetically "complex" causation, involving many genes, and strong influences of the environment, acting on our individual genetic endowments over the entire life course. Lines of evidence that this is so are reviewed--especially migrant epidemiological cohort studies, which are stronger etiological evidence that "twins reared apart" studies, since they tend to involve massive cultural and environmental changes, while "holding genetic factors constant." More such studies would better inform preventive strategies for the inflammatory arthritides, which lag behind cardiovascular disease in understanding causation, and therefore primary prevention. Finally, factors are briefly reviewed that affect risks, benefits, and costs of single-locus genetic tests to predict lifelong risk of chronic diseases with complex and multifactorial determination. Both negative and positive predictive values of such tests for predicting lifetime disease occurrence are generally unacceptable for use in the general population. Expert genetic counseling is therefore important before such testing, to ensure that an appropriate family and personal history justifies these expensive tests, the "labeling" effects of which can last a lifetime. PMID- 15660473 TI - The economics of follow-on drug research and development: trends in entry rates and the timing of development. AB - OBJECTIVES: The development of so-called 'me-too', or 'follow-on', drugs by the pharmaceutical industry has been viewed by some as duplicative and wasteful, while others have argued that these drugs often provide needed therapeutic options and inject some price competition into the marketplace. This study examines data on the trends in the speed with which competitive entry has occurred in the pharmaceutical marketplace and the competitive nature of the industry's development of these drugs. DATA AND METHODS: We examined data on the entry rates of drugs in a large number of therapeutic classes over time, as well as detailed survey information on the relative timing of the development of drugs in the classes. Classes were defined according to chemical structure or pharmacologic mode of action and similarity of clinical use. We determined average times to initial and subsequent entry in drug classes by period and examined the timing of development milestones achieved by what have turned out to be follow-on drugs in relation to the development and approval of the first drug in a class to be approved. RESULTS: We found that the period of marketing exclusivity that the breakthrough drug in a new class enjoys has fallen dramatically over time (a median of 10.2 years in the 1970s to 1.2 years for the late 1990 s). Approximately one-third of follow-on new drugs received a priority rating from the US FDA. The vast majority of the follow-on drugs for drug classes that were created in the last decade were in clinical development prior to the approval of the class breakthrough drug. CONCLUSIONS: The data suggest that entry barriers have fallen over time for new drug introductions. The increased competitiveness of the pharmaceutical marketplace was likely fueled by changes over time on both the supply and demand sides. The development histories of entrants to new drug classes suggest that development races better characterise new drug development than does a model of post hoc imitation. Thus, the usual distinctions drawn between breakthrough and 'me-too' drugs may not be very meaningful. PMID- 15660474 TI - Are the economics of pharmaceutical research and development changing?: productivity, patents and political pressures. AB - Pharmaceutical research and development (R&D) competition in the 1980s and 1990 s was characterised by rising R&D expenditures, favourable returns to innovators and the introduction of many new classes of drugs with high social benefits. However, in the past 3 years, the number of new drug introductions has been well below the historical trend, while the cost per new drug continues to increase. In addition to lagging R&D productivity, the industry has been characterised by other economic and policy uncertainties. These include a wave of early patent challenges and growing political pressure to contain pharmaceutical expenditures. This paper examines the consequences of these developments. PMID- 15660475 TI - The effects of pharmaceutical consumption and obesity on the quality of life in the organization of economic cooperation and development (OECD) countries. AB - OBJECTIVE: The objective of this study is to validate our earlier work on life expectancy with more recent data and, more importantly, to extend it to examine quality of life, not only the length of life. DESIGN AND SETTING: The analysis focuses on the production of health, disaggregating healthcare into pharmaceutical consumption and other healthcare. Going beyond our earlier work, measures of health include life expectancy and disability-adjusted life expectancy (DALE). Also, we consider the impact of obesity. The sample was 18 Organization of Economic Cooperation and Development (OECD) countries. The measure of pharmaceutical consumption is the best that is available for these countries. MAIN OUTCOME MEASURES AND RESULTS: Confirming our earlier work, pharmaceutical consumption has a positive and statistically significant effect on life expectancy at 40 and 60 years (significant at the 0.05 level, based on a two tailed test). The effects are slightly larger than in the earlier work. Turning to DALE, pharmaceutical consumption has a positive and statistically significant effect at birth and at 60 years (significant at the 0.05 and 0.01 levels, respectively), based on a two-tailed test. The effects on DALE are larger than the effects on life expectancy. CONCLUSIONS: Increased pharmaceutical consumption helps improve quality of life, as well as life expectancy. PMID- 15660476 TI - Changes in the costs of treating mental health disorders: an overview of recent research findings. AB - This article reviews recent evidence on changes over time in the direct medical costs of treating three of the more common mental health disorders in the US: the acute (16-week) phase of major depressive disorder, the ongoing treatment of schizophrenia, and the ongoing treatment of bipolar I disorders. The three studies discussed in this article cover various time intervals over the decade from 1991 through 2000, and encompass both private sector and governmental funding sources. Although there has been a shift over time away from intensive psychosocial/psychotherapy and towards increasingly expensive psychopharmacotherapy for all three disorders, total direct medical costs of treatment for each of these three mental health disorders have been declining over time. However, a substantial portion of treatment is not supported by clinical evidence. PMID- 15660477 TI - Impediments to the diffusion of innovative medicines in Europe. AB - The pharmaceutical industry has developed many innovative medicines, which are able to extend the life expectancy of patients to increase their quality of life and often to reduce expenditures in the health care sector as a whole. Although these medicines are available in principle for all eligible patients throughout Europe, not everyone receives adequate treatment. There is a huge difference between a possible optimal treatment and the treatment delivered to the patient. In some cases patients are not treated at all; in some cases they only receive outdated medicines (e.g. with lower effectiveness and/or more severe side effects); and in some cases the prescribed dosages of the innovative drugs are too low to be effective. This study gives an overview of the shortfalls in provision of state-of-the-art medicines in selected European countries for about 20 of the most relevant diseases. The following five different groups of factors can be identified as leading to this insufficient diffusion of medicines and are discussed in this text: (i) patient-related factors; (ii) healthcare professional related factors; (iii) industry-related factors; (iv) system-related (long-term) factors; and (v) policy-related (short-term) factors. It must be clear that these shortages are not isolated cases but general trends in Europe, which have to be discussed in public. PMID- 15660478 TI - The impact of healthcare reform in the Netherlands. AB - In 1987, the Dekker committee proposed managed competition as the dominant principle for reforming healthcare in The Netherlands. Considerable progress has been made in implementing the Dekker proposal, such as risk-adjusted capitation of Sick Funds, yearly open enrollment and selective contracting with providers. The centre-right government, which came to power in 2003, plans to implement the final steps in the coming years, which will drastically change pharmaceutical policies and shift power from central government to regional actors in healthcare. The current price reference system, which was initiated in 1991, has failed in containing expenditure or providing incentives towards efficiency. For new drugs, which can not be clustered in the price reference system or for which the producer wants a premium price, a pharmacoeconomic study and budget impact analysis is formally required from 2005 onwards. This fourth hurdle may limit access to new drugs, which contribute considerably to expenditure or for which relative efficiency is above a certain cost-effectiveness threshold. In line with the Dekker principle, insurance companies are expected to step into the market and initiate policies to reduce costs. However, the government seems somewhat reluctant to abandon instruments belonging to old supply-side regulation. Given the current healthcare cost explosion, which coincides with an economic recession, it remains to be seen whether politicians will really shift the power to the insurers and not return to traditional cost-containment policies. PMID- 15660479 TI - Global principles for better healthcare. AB - This article provides an overview of a report by NERA Economic Consulting based on the outcomes of the health reform experiences of over 25 countries. The report is organised around four widely accepted 'global principles' governing the planning, funding and provision of healthcare services: fair access, efficiency, responsiveness to society and innovation. These principles, in turn, provide guidance for addressing problems that commonly plague national healthcare systems, such as balancing funding constraints against demand pressures and maintaining fair access for all to healthcare whilst responding to ever more demanding patients and rising expectations. Whilst the global principles cannot provide a formula for dealing with the array of problems facing various countries, they can help healthcare decision-makers recognise the necessary trade offs they must face, take the long-term view towards problem-solving and work toward addressing disparate needs in a consistent manner. PMID- 15660480 TI - The role of pharmacoepidemiology and pharmacoeconomics in promoting access and stimulating innovation. AB - Answering the demanding questions asked of pharmacotherapy in the 21st century will require more reliance on the disciplines of pharmacoepidemiology and pharmacoeconomics. Pharmacoepidemiology can help assess patterns and appropriateness of drug utilisation, provide explanations for poor compliance, quantify the frequency and severity of side effects, and aid in the design and evaluation of interventions to improve drug use and outcomes. Pharmacoeconomics can help determine whether a new costlier product offers sufficient clinical advantage over its predecessors to justify the increased cost. Taken together, pharmacoepidemiology and pharmacoeconomics represent the next logical step in the evolution of medication assessment; their judicious deployment can help ensure both access to new medicines and innovation. PMID- 15660481 TI - The policy implications of gender mainstreaming for healthcare research in the EU. AB - Scientific and technological progress in the life sciences and modern biotechnology is continuing at a breathtaking pace. At the same time, the potential implications for individuals and society are giving rise to intense debate. This article gives a background to the European Institute of Women's Health (EIWH) and the work of the organisation in contributing to and developing policy at a European level. We will identify the policy issues related to gender based healthcare research arising from the EU 6th Framework Programme for Research, which was introduced this year, and the implications for the EU Public Health Strategy. Additionally, the issues of personalized medicine and the promise of biotechnology for improved medicines and healthcare will be addressed from a gender perspective. What are the prospects in the medium and long term? How can we best address the ethical and socio-economic implications? PMID- 15660488 TI - Raising the standards in education and nursing. PMID- 15660489 TI - Understanding the culture of Chinese children and families. AB - Providing appropriate health care to a client can be accomplished only in an environment that is sensitive to the cultural values and beliefs of the client. As the population of first- and second-generation Chinese immigrants increases in the United States, the need to develop culturally sensitive health care becomes significant. Chinese immigrants and their families have become an important part of American society, including the school setting. The school nurse, who regularly works with students and families, should work in a manner that allows Chinese immigrants to maintain their cultural values and beliefs, while providing appropriate care for the student. The Chinese culture is unique and holds values and beliefs that contrast with those of the Western culture. A school nurse who understands and incorporates the Chinese culture will be better able to develop a positive interaction with the family and make arrangements for culturally appropriate care. PMID- 15660490 TI - The significance of herpes simplex for school nurses. AB - Herpes simplex is a common recurrent viral infection caused by the herpes simplex virus. The two closely related but distinct viruses that cause herpes simplex infections are herpes simplex 1 (HSV-1) and herpes simplex 2 (HSV-2). HSV-1 is commonly associated with infections around the oral mucosa and is the cause of herpes labialis, often referred to as a fever blister or cold sore. HSV-2 infections are usually acquired sexually. Genital herpes is a sexually transmitted disease with the highest prevalence among adolescents and young adults. Knowledge of viral activity, disease management, and community resources will assist the school nurse in developing and implementing strategies to prevent and manage this chronic disease. PMID- 15660492 TI - Increasing minority representation in the health professions. AB - Research indicates that health disparities may be reduced by increasing the number of ethnic minorities working in health occupations. Establishing health career pathway programs for immigrant and ethnic minority students is one way to address this problem. One such program, Cross-Cultural Education in Public Health (CCEPH), was developed, implemented, and evaluated to determine whether participants expressed greater interest in pursuing health care careers after program completion. A sample of 72 immigrant students in two high schools participated in the program, which was based on Bandura's theory of self efficacy. Data were gathered using pre- and postprogram surveys that measured academic self-efficacy and career consideration. Results for academic efficacy were not statistically significant, but interest level in health care careers rose substantially. Although further research needs to be conducted to determine whether such programs increase self-efficacy, programs such as CCEPH can increase the consideration of health careers among immigrant and ethnic minority students. PMID- 15660491 TI - The school nurse role in asthma management: can the action plan help? AB - Asthma is the most common chronic disorder in American schoolchildren, and school nurses play a valuable role in its management. A study was conducted in which school nurses were asked to describe their role in caring for students with asthma and their use of Asthma Action Plans (AAPs). The nurses indicated that they frequently provided direct care and education. They were comfortable with providing care to students with asthma and familiar with AAPs. Having an AAP increased their confidence in managing students with asthma. This emphasizes the need for continued education regarding the AAP and the development of policies that direct care and encourage use of an AAP at school. PMID- 15660493 TI - Coping and social support for parents of children with autism. AB - Autism in children has increased significantly in the past 15 years. The challenges and stressors associated with providing services and caring for a child with autism affect families, educators, and health professionals. This descriptive study used a survey to collect data on parents' perceptions of coping strategies and social support. Instruments included the Social Support Index and the Family Crisis Oriented Personal Evaluation Scales. One half of the families identified serious stressors in addition to autism. Acquiring social support and reframing were the most frequently used coping strategies. The school nurse is in a position to identify needs and refer families to local support groups and agencies, facilitating social support and development of coping strategies. PMID- 15660494 TI - Section 504 of the Rehabilitation Act: determining eligibility and implications for school districts. AB - Today, school districts are challenged in meeting the health and educational needs of students with chronic health conditions. One of the challenges school districts face is determining when students with health-related disabilities are eligible for services under Section 504 of the Rehabilitation Act of 1973. This article reviews Section 504 and its regulations as they apply to public schools, particularly with respect to eligibility criteria for students with special health care needs. The article also reviews recent case law and explores the implications of these legal standards for school district practice, including the need for clear policies and procedures, consistent Section 504 teams, training, and alternatives for meeting the needs of students who are found not to be eligible for services under Section 504. PMID- 15660496 TI - Data integrity: backup. AB - School nurses and health office staff have an ethical and legal obligation to protect the integrity of electronic student health records. Various sources, both accidental and malicious, can threaten student health data. School nurses must employ regular and reliable methods to back up student health data. PMID- 15660497 TI - Barriers identified by Swedish school nurses in giving information about testicular cancer and testicular self-examination to adolescent males. AB - The purpose of this study was to investigate to what extent school nurses in Sweden inform adolescent men about testicular cancer (TC) and testicular self examination (TSE). A questionnaire was completed by 129 school nurses from 29 randomly selected municipalities. All respondents were women, with a mean age of 42 years. The results showed that about 6% of the school nurses had provided information on TC and 9% on TSE; however, the majority was open to presenting such information. The major reason reported for not providing this guidance was insufficient knowledge about the subject. The nurses who had given information were knowledgeable about both TC and TSE and more often informed young men about TC and TSE, whereas those who had only received information about TC were not as likely to provide information about TC or TSE. The results of this study highlight the need to educate school nurses about TC and TSE so they can include this information in their health teaching to adolescent males. PMID- 15660498 TI - Myelodysplastic syndrome. AB - During the past 15 years, important progress has been made in the understanding of the biology and prognosis of myelodysplastic syndrome (MDS). MDS is a clonal disorder characterized by ineffective hematopoiesis, which can lead to either fatal cytopenias or acute myelogenous leukemia (AML). Risk-adapted treatment strategies were established because of the high median age (60-75 years) of the MDS patients and the individual history of the disease (number of cytopenias, cytogenetic changes, transfusion requirements). Allogeneic bone marrow transplantation currently offers the only potentially curative treatment, but this form of therapy is not available for the typical MDS patient, who is >60 years of age. Therapy with erythropoietin and G-CSF has improved the quality of life of selected patients. The development of small molecules directed against specific molecular targets with minimal adverse effects is the hope for the future. Innovative uses of immunomodulatory agents and the optimizing of cytotoxic treatment should continue to help in the treatment of MDS. PMID- 15660499 TI - G protein polymorphisms in hypertension, atherosclerosis, and diabetes. AB - A common C825T polymorphism in the gene GNB3, which encodes the beta 3 subunit of heterotrimeric G proteins, was identified in cell lines from patients with hypertension. The 825T allele is associated with increased intracellular signal transduction. Many population-based and case-control studies in different ethnicities have investigated an association between this polymorphism and hypertension, obesity, and atherosclerosis. A critical assessment of published studies suggests that 825T allele carriers have an increased risk for hypertension combined with features of the metabolic syndrome, such as dyslipidemia, hypercholesterolemia, insulin resistance, and obesity. It is anticipated that this polymorphism will be used in clinical practice to better characterize hypertension and for individualized treatment regimens. PMID- 15660501 TI - Metabolic syndrome: a clinical and molecular perspective. AB - The metabolic syndrome is a cluster of interrelated common clinical disorders, including obesity, insulin resistance, glucose intolerance, hypertension, and dyslipidemia (hypertriglyceridemia and low HDL cholesterol levels). According to recently defined criteria, the metabolic syndrome is prevalent and is associated with a greater risk of atherosclerotic cardiovascular disease than any of its individual components. Primary defects in energy balance that produce obesity (and visceral adiposity in particular) are sufficient to drive all aspects of the syndrome. Increased free fatty acids and lipid accumulation in certain organs are mediators of insulin resistance. Obesity also leads to a proinflammatory and prothrombotic state that potentiates atherosclerosis. Pathways leading directly from adiposity to the genesis of dyslipidemia and hypertension have been elucidated. Recent knowledge implies a role for fat-derived "adipokines," including TNF alpha and adiponectin, as pathogenic contributors or protective factors. Current therapies include diet and exercise as well as agents indicated for the treatment of individual components of the syndrome. Future therapies may accrue from the aggressive pursuit of newer molecular drug targets that have the potential to prevent or treat multiple aspects of the metabolic syndrome. PMID- 15660500 TI - Post-transplant lymphoproliferative disorders. AB - Post-transplant lymphoproliferative disorder (PTLD) is a life-threatening complication after hematopoietic stem cell or solid organ transplantation. The majority of PTLD is of B-cell origin and associated with Epstein-Barr virus (EBV). During the past decade progress has been made in better understanding the pathogenesis of PTLD, and early detection strategies, such as serial measurement of EBV-DNA load in peripheral blood samples, have assisted in the identification of high-risk patients. In addition, novel immunotherapies have been developed, including the use of monoclonal antibodies and adoptive transfer of EBV-specific T cells. Despite these advances, it remains a major challenge to define indications for preemptive therapies for PTLD and to integrate novel therapeutic approaches with conventional therapies. PMID- 15660502 TI - New anticoagulant therapy. AB - The development of new anticoagulants is expanding the list of drugs that can be used to prevent and treat venous and arterial thrombosis. New parenteral anticoagulants have been developed to overcome the limitations of heparin and low molecular-weight heparin, whereas novel orally active anticoagulants have been designed to provide more streamlined therapy than vitamin K antagonists. This review identifies the molecular targets of new anticoagulants, describes the results of clinical trials, and provides clinical perspective on the opportunities for new anticoagulants. PMID- 15660503 TI - Endothelial progenitor cells. AB - Endothelial progenitor cells are a circulating, bone marrow-derived cell population that appears to participate in both vasculogenesis and vascular homeostasis. Questions persist regarding their functional characteristics, as well as the precise panel of cell surface markers that uniquely define this newly described progenitor cell population. We review experimental results obtained from both animal studies and recent clinical trials that suggest this cell type may have tremendous therapeutic potential for a wide range of human diseases. PMID- 15660504 TI - Aromatase inhibitors: rationale and use in breast cancer. AB - Considerable data implicate estrogens in breast cancer carcinogenesis and progression. In the postmenopausal woman, estrogens are produced in breast tissues and many other sites throughout the body when androgen precursors are converted into estrogens via the enzyme aromatase. Inhibition of this enzyme with aromatase inhibitors (AIs) has demonstrated reductions in systemic as well as intratumoral estrogens. These drugs have now been utilized in large phase 3 randomized trials and have led to greater improved clinical benefit than the "gold standard," tamoxifen. Questions remain about the long-term side effects and safety profile of AIs. They are associated with increasing incidence of osteoporosis and bone fractures. Nevertheless, AIs add to our armamentarium for therapy and possible prevention of breast cancer. PMID- 15660505 TI - Andropause: is androgen replacement therapy indicated for the aging male? AB - The number of men in the United States > or =65 years of age is projected to increase from 14,452,000 in 2000 to 31,343,000 in 2030. Approximately 30% of men 60-70 years of age and 70% of men 70-80 years of age have low bioavailable or free testosterone levels. Symptoms and findings of testosterone deficiency are similar to those associated with aging. They include loss of energy, depressed mood, decreased libido, erectile dysfunction, decreased muscle mass and strength, increased fat mass, frailty, osteopenia, and osteoporosis. Several small clinical trials indicate that testosterone replacement therapy can improve many of these findings; however, the studies have not been powered to assess potential risks, such as the need for invasive treatment of benign prostatic hyperplasia, development of a clinical prostate cancer, or cardiovascular events. Thus, the benefit/risk ratio of testosterone replacement therapy in aging men is not known. PMID- 15660506 TI - Surgical therapy for metastatic disease to the liver. AB - The liver is a common site of hematogenous metastasis. In the past, patients with liver metastases were often deemed inoperable, and medical therapy conferred only minor survival benefit. However, advances in surgical techniques and chemotherapeutic agents during the past two decades have led to effective treatments for selected patients with metastases to the liver. Up to approximately 80% of the liver can be resected, and partial hepatectomy is now routinely performed with a perioperative mortality rate of <5%. Surgical resection of colorectal cancer metastatic to the liver results in a 5-year survival rate of 40%. These results are expected to improve even further with multimodality approaches that include newer chemotherapy regimens. Liver metastases from other primary tumors, such as neuroendocrine carcinoma and genitourinary tumors, are also treated effectively with liver resection. The indications for surgical treatment of liver metastases are broadening as a variety of novel therapies are being developed, including hepatic artery embolization, hepatic artery infusion of chemotherapy, and radiofrequency ablation. PMID- 15660507 TI - New strategies in the treatment of the thalassemias. AB - In addition to the severe beta thalassemias, hematologists have begun to recognize the more severe forms of alpha thalassemia, namely hemoglobin (Hb) H disease and Hb H/Hb Constant Spring, as well as the beta compound heterozygote, beta thalassemia/HbE. Clinically, variably severe anemia becomes apparent in the first year accompanied by occasionally massive expansion of erythropoiesis. The most anemic patients require regular red blood cell transfusions to avoid death from cardiac failure. However, the inevitable iron accumulation leads to dysfunction, primarily involving the heart, liver, and endocrine system; thus, regularly transfused patients require iron chelation. A major discovery was that allogeneic bone marrow (stem cell) transplantation in severely affected subjects with both alpha and beta thalassemia could result in cure. Current work deals with specific complications, such as iron overload and endocrine, cardiopulmonary, thrombophilic, and osteopenic problems. The thalassemias are likely to benefit in the future from specific gene therapy. There are also important advances in genetic counseling based on results of early fetal diagnosis. PMID- 15660508 TI - New concepts in von Willebrand disease. AB - Von Willebrand factor (VWF) behaves as an extracellular adapter molecule, linking platelets to the extracellular matrix at sites of vascular injury. These interactions are crucial for hemostasis. Too little platelet adhesion causes bleeding that is typical of von Willebrand disease, whereas too much platelet adhesion may cause thrombotic thrombocytopenic purpura. Mutations in VWF or platelet glycoprotein Ib can either reduce or increase the affinity of platelet binding. Paradoxically, affinity changes in either direction cause bleeding. Crystallographic studies now suggest molecular explanations for all of these phenotypes. Clinical investigations of von Willebrand disease type 1 are defining the relationship between plasma VWF level and the risk of bleeding or thrombosis. Emerging data suggest that VWF level is a useful biomarker for the risk of either bleeding or thrombosis and could be incorporated into a comprehensive approach to treat or prevent these adverse events. PMID- 15660509 TI - Genetics of longevity and aging. AB - Longevity, i.e., the property of being long-lived, has its natural limitation in the aging process. Longevity has a strong genetic component, as has become apparent from studies with a variety of organisms, from yeast to humans. Genetic screening efforts with invertebrates have unraveled multiple genetic pathways that suggest longevity is promoted through the manipulation of metabolism and the resistance to oxidative stress. To some extent, these same mechanisms appear to act in mammals also, despite considerable divergence during evolution. Thus far, evidence from population-based studies with humans suggests the importance of genes involved in cardiovascular disease as important determinants of longevity. The challenge is to test if the candidate longevity genes that have emerged from studies with model organisms exhibit genetic variation for life span in human populations. Future investigations are likely to involve large-scale case-control studies, in which large numbers of genes, corresponding to entire gene functional modules, will be assessed for all possible sequence variation and associated with detailed phenotypic information on each individual over extended periods of time. This should eventually unravel the genetic factors that contribute to each particular aging phenotype. PMID- 15660511 TI - Therapeutic intervention and targets for sepsis. AB - Sepsis syndrome, a systemic response to infection, can beget devastating outcomes even in previously normal individuals. Recent research in septic patients has led to the discovery that early goal-directed resuscitation guided by continuous monitoring of mixed venous hemoglobin saturation, along with moderate doses of corticosteroids, can reduce mortality. An improved understanding of the complex interaction between the inflammatory and coagulant systems in sepsis pathophysiology has resulted in novel treatments, such as recombinant human activated protein C, which improves survival in patients with severe sepsis and a high risk of death. However, despite an increased understanding of the complex pathophysiology of this syndrome and the discovery of new, effective treatments, severe sepsis still results in significant morbidity and mortality. Consequently, investigations continue into additional therapeutic agents directed against novel targets. Following a review of recent advances in sepsis treatment, we briefly discuss a few of the new, promising therapeutic strategies currently being investigated. PMID- 15660510 TI - Progress toward an HIV vaccine. AB - The development of an HIV vaccine is proving to be an unprecedented challenge. The difficulty in creating this vaccine arises from the enormous genetic variation of the virus and the unusual importance of cytotoxic T lymphocytes (CTL) in controlling its spread. Whereas traditional vaccine strategies are unlikely to confer safe and effective HIV protection, novel strategies for eliciting CTL have provided substantial clinical benefits in nonhuman primate model systems. These vaccine strategies, including plasmid DNA and live recombinant vectors, are currently being evaluated in human clinical trials. PMID- 15660512 TI - Management of peripheral vascular disease. AB - The management of patients with peripheral arterial occlusive disease (PAD) has to be planned in the context of natural history, epidemiology, and apparent risk factors that predict deterioration. The ankle-brachial index to date has proved to be the most effective, accurate, and practical method of PAD detection. Given that PAD is a powerful indicator of systemic atherosclerosis and (independent of symptoms) is associated with an increased risk of myocardial infarction and stroke, as well as a six times greater likelihood of death, the prevalence and demographic distribution of measurable PAD becomes particularly relevant. Reliable information on interventions to confer symptom relief is much weaker and reflects discrepancies between published reports from centers of excellence and the experience of patients routinely treated in communities around the world. The impact of newer treatment modalities, such as complex endovascular procedures and therapeutic angiogenesis, has been a subject of recent controversy. PMID- 15660513 TI - Role of magnetic resonance imaging and immunotherapy in treating multiple sclerosis. AB - Significant advances in magnetic resonance imaging (MRI) technology and treatment of multiple sclerosis (MS) have been made during the past decade. These advances have revealed evidence of profound heterogeneity in MS. There is a clear need to revisit the key issues in MS pathogenesis and treatment strategies, taking new data into consideration. This paper provides an overview of recent progress in MS research, including (a) a review of clinical, pathologic, and immunologic aspects of MS, (b) a discussion of the mechanism of action of currently available disease modifying drugs for MS, (c) an account of the role of MRI in clinical management and clinical trials in MS, and (d) an overview of some emerging treatments for MS. PMID- 15660514 TI - Definition and clinical importance of haplotypes. AB - Advances in genotyping and sequencing technologies, coupled with the development of sophisticated statistical methods, have afforded investigators novel opportunities to define the role of sequence variation in the development of common human diseases. At the forefront of these investigations is the use of dense maps of single-nucleotide polymorphisms (SNPs) and the haplotypes derived from these polymorphisms. Here we review basic concepts of high-density genetic maps of SNPs and haplotypes and how they are typically generated and used in human genetic research. We also provide useful examples and tools available for researchers interested in incorporating haplotypes into their studies. Finally, we discuss the latest concepts for the analysis of haplotypes related to human disease, including haplotype blocks, the International HapMap Project, and the future directions of these resources. PMID- 15660515 TI - Approaches to therapy of prion diseases. AB - Devising approaches to the therapy of transmissible spongiform encephalopathies, or prion diseases, is beset by many difficulties. For one, the nature of the infectious agent, the prion, is understood only in outline, and its composition, structure, and mode of replication are still shrouded in mystery. In addition, the mechanism of pathogenesis is not well understood. Because clinical disease affects mainly the brain parenchyme, therapeutic agents must be able to traverse the brain-blood barrier (BBB) or have to be introduced directly into the cerebrospinal fluid or brain tissue. And finally, because the disease is usually recognized only after onset of severe clinical symptoms, the question arises as to whether the neurodegenerative processes can be reversed to any extent after a successful eradication of the agent. PMID- 15660516 TI - Endometriosis: new genetic approaches and therapy. AB - Endometriosis is a relatively common condition in women of reproductive age and is associated with considerable morbidity. Despite an extensive literature describing its multiple clinical manifestations, their management, and many aspects of the biology of endometriotic lesions, the pathophysiological mechanisms involved remain poorly understood. A genetic component in endometriosis is now recognized, and several groups have taken up the challenge of using genetic techniques to identify the aberrant molecular and cellular mechanisms in endometriosis with the intention of providing much-needed insights that might, in turn, lead to new therapies. The techniques that have been applied include expression profiling, tumor genetic studies, functional candidate gene studies, and linkage studies that can adopt a hypothesis-free approach. This review describes the current status of these studies and explores the prospects for new therapies. PMID- 15660517 TI - Severe acute respiratory syndrome (SARS): a year in review. AB - Severe acute respiratory syndrome (SARS) emerged from China as an untreatable and rapidly spreading respiratory illness of unknown etiology. Following point source exposure in February 2003, more than a dozen guests infected at a Hong Kong hotel seeded multi-country outbreaks that persisted through the spring of 2003. The World Health Organization responded by invoking traditional public health measures and advanced technologies to control the illness and contain the cause. A novel coronavirus was implicated and its entire genome was sequenced by mid April 2003. The urgency of responding to this threat focused scientific endeavor and stimulated global collaboration. Through real-time application of accumulating knowledge, the world proved capable of arresting the first pandemic threat of the twenty-first century, despite early respiratory-borne spread and global susceptibility. This review synthesizes lessons learned from this remarkable achievement. These lessons can be applied to re-emergence of SARS or to the next pandemic threat to arise. PMID- 15660518 TI - Gene-environment interactions in asthma and other respiratory diseases. AB - It is generally agreed that many lung diseases such as asthma and chronic obstructive pulmonary disease (COPD) have polygenic inheritance, and that the association of a specific genotype or genotypes with the disease is likely to vary between populations. Furthermore, it is recognized that the etiology of many lung diseases involves a complex interplay between genetic background and exposure to multiple environmental stimuli, and understanding the mechanisms through which genes and environment interact represents a major challenge for pulmonary researchers. We discuss experimental approaches and challenges that must be overcome to identify disease genes for asthma, COPD and chronic bronchitis, and occupational lung diseases. In particular, common polymorphisms in CD14, glutathione S-transferase, and tumor necrosis factor alpha have been found to be important in gene-environment interaction and asthma pathogenesis. An understanding of gene-environment interactions in complex lung diseases is essential to the development of new strategies for lung disease prevention and treatment. PMID- 15660519 TI - The silent revolution: RNA interference as basic biology, research tool, and therapeutic. AB - RNA interference (RNAi) is an evolutionarily conserved mechanism for silencing gene expression. In primitive organisms, RNAi protects the genome from viruses and other insertable genetic elements and regulates gene expression during development. The antisense (guide) strand of short double-stranded RNAs is incorporated into an RNA-induced silencing complex that can either suppress protein expression or direct degradation of messenger RNAs that contain homologous sequence(s). The discovery that RNAi works in mammalian cells has sparked intense investigation into its role in normal mammalian cell function, its use as a tool to understand or screen for genes functioning in cellular pathways in healthy and diseased cells and animals, and its potential for therapeutic gene silencing. RNAi may provide an important new therapeutic modality for treating infection, cancer, neurodegenerative disease, and other illnesses, although in vivo delivery of small interfering RNAs into cells remains a significant obstacle. PMID- 15660520 TI - Management of adult idiopathic thrombocytopenic purpura. AB - Idiopathic thrombocytopenic purpura (ITP) is a common hematologic disorder manifested by immune-mediated thrombocytopenia. The diagnosis remains one of exclusion, after other thrombocytopenic disorders are ruled out based on history, physical examination, and laboratory evaluation. The goal of treatment is to raise the platelet count into a hemostatically safe range. The disorder is usually chronic, although there is considerable variation in the clinical course and most patients eventually attain safe platelet counts off treatment. However, a subset of patients has severe disease refractory to all treatment modalities, which is associated with considerable morbidity and mortality. This article focuses on the management of primary ITP in adults. We discuss criteria for treatment, the roles of splenectomy and other treatment options along with their side effects, and the management of ITP during pregnancy. PMID- 15660521 TI - Monogenic obesity in humans. AB - Until relatively recently, the small number of identifiable inherited human diseases associated with marked obesity were complex, pleiotropic developmental disorders, the molecular basis for which were entirely obscure. The molecular basis for many of these complex syndromes, such as Bardet Beidl syndrome, has been revealed, providing novel insights into processes essential for human hypothalamic function and energy balance. In addition to these discoveries, which were the fruits of positional cloning, the molecular constituents of the signaling pathways responsible for the control of mammalian energy homeostasis have been identified, largely through the study of natural or artificial mutations in mice. We discuss the increasing number of human disorders that result from genetic disruption of the leptin-melanocortin pathways that have been identified. Practical implications of these findings for genetic counseling, prognostication, and even therapy have already emerged. PMID- 15660522 TI - Application of microbial genomic science to advanced therapeutics. AB - Since the publication of the first complete microbial genome sequence of Haemophilus influenzae in 1995, more than 200 additional microbial genome sequences have become available in the public domain. Approximately 40% of these represent important human pathogens. Comparative in silico methods, along with large-scale approaches such as transcriptomics and proteomics, are beginning to reveal insights into new virulence genes, pathogen-host interactions, and the molecular basis of host specificity. Sequence data are also starting to accumulate from multiple isolates or strains of a single pathogen, and this type of data has proven to be quite valuable in providing new insights into the genetic variability that is present in a particular species as well as in facilitating correlations between genotype and phenotype. Ultimately, a major goal of genome-enabled infectious disease research is the development of novel diagnostics, therapeutics, and vaccines. PMID- 15660523 TI - Atrial fibrillation: modern concepts and management. AB - Atrial fibrillation (AF) is a common cardiac arrhythmia responsible for significant morbidity and mortality. In recent years, progress has been made in determining the genetic abnormalities that may lead to AF. New trials have shown that rate control and anticoagulation are acceptable as a primary treatment strategy in many patients who have a high risk of recurrence. Newer and safer antiarrhythmics are now available. Pacemaker and implantable cardiac defibrillator technology is rapidly evolving and may play a significant role in future treatment and prevention of AF. Direct thrombin inhibitors are likely to add a user-friendly option to the current standard therapy for stroke prevention. PMID- 15660524 TI - DNA repair defects in stem cell function and aging. AB - Cellular DNA is under constant challenge by exogenous and endogenous genotoxic stress, which results in both transient and accumulated DNA damage and genomic instability. All cells are equipped with DNA damage response pathways that trigger DNA repair, cell cycle arrest, and, if need be, apoptosis, to eliminate DNA damage or damaged cells. The consequences of these processes for stem cells can be profound: diminution in stem cell pools, or, because of altered gene expression, an increased chance for stem cell differentiation or malignant transformation. Furthermore, a number of DNA repair abnormalities are linked to premature aging syndromes, and these are associated with defects in the stem cell population. The specific DNA repair systems for which there are data regarding the impact of repair defects on stem cell function include O(6)-alkylguanine DNA alkyltransferase, nucleotide excision repair, base excision repair, mismatch repair, non-homologous DNA end-joining Fanconi's anemia protein complex, and homologous recombination. It has recently become clear that deficiencies of these processes are associated not only with cancer and/or aging but also with stem cell defects. This discovery raises the possibility of a link between aging and stem cell dysfunction. In this review, we provide evidence for a link between DNA repair systems and the maintenance and longevity of stem cells. PMID- 15660525 TI - Hematopoietic stem and progenitor cells: clinical and preclinical regeneration of the hematolymphoid system. AB - A vast literature exists on the biology of blood formation and regeneration under experimental and clinical conditions. The field of hematopoiesis was recently advanced by the capacity to purify to homogeneity primitive hematopoietic stem and progenitor cells. Isolation of cells at defined maturational stages has enhanced the understanding of the fundamental nature of stem cells, including how cell fate decisions are made, and this understanding is relevant to the development of other normal as well as malignant tissues. This review updates the basic biology of hematopoietic stem cells (HSC) and progenitors, the evolving use of purified HSC as grafts for clinical hematopoietic cell transplantation (HCT) including immune tolerance induction, and the application of HSC biology to other stem cell fields. PMID- 15660526 TI - Inherited susceptibility to colorectal cancer. AB - The principal Mendelian disorders predisposing to colorectal cancer are familial adenomatous polyposis (FAP) and hereditary nonpolyposis colorectal cancer (HNPCC). FAP is due to mutations in the APC gene. HNPCC is due to a mutation in one of at least five mismatch repair genes. Identification of individuals with these conditions is important because colon cancer will occur in approximately 80% and onset is early. For FAP, protein truncation testing will identify the vast majority of mutations. For HNPCC, 80%-95% can be identified by microsatellite instability testing. A current U.S. study reports that 12% of consecutive colorectal cancers have high microsatellite instability and that, of this 12%, 25% have detectable mutations of MLH1, MSH2, or MSH6. Potential benefits of identification include improved compliance with recommended surveillance, early detection of polyps, reduction in cancer mortality, offering of testing to relatives, and reassurance for relatives found to be negative with attendant savings in the time and expense of surveillance. PMID- 15660527 TI - Aptamers: an emerging class of therapeutics. AB - Numerous nucleic acid ligands, also termed decoys or aptamers, have been developed during the past 15 years that can inhibit the activity of many pathogenic proteins. Two of them, Macugen and E2F decoy, are in phase III clinical trials. Several properties of aptamers make them an attractive class of therapeutic compounds. Their affinity and specificity for a given protein make it possible to isolate a ligand to virtually any target, and adjusting their bioavailability expands their clinical utility. The ability to develop aptamers that retain activity in multiple organisms facilitates preclinical development. Antidote control of aptamer activity enables safe, tightly controlled therapeutics. Aptamers may prove useful in the treatment of a wide variety of human maladies, including infectious diseases, cancer, and cardiovascular disease. We review the observations that facilitated the development of this emerging class of therapeutics, summarize progress to date, and speculate on the eventual utility of such agents in the clinic. PMID- 15660528 TI - Gene therapy for severe combined immunodeficiency. AB - Studies of severe combined immunodeficiency (SCID), a group of rare monogenic disorders, have provided key findings about the physiology of immune system development. The common characteristic of these diseases is the occurrence of a block in T cell differentiation, always associated with a direct or indirect impairment of B cell immunity. The resulting combined immunodeficiency is responsible for the clinical severity of SCID, which, without treatment, leads to death within the first year of life. Eleven distinct SCID phenotypes have been identified to date. Mutations of ten genes have been found to cause SCID. Identifying the pathophysiological basis of most SCID conditions has led to the possibility of molecular therapy as an alternative to allogeneic hematopoietic stem cell transplantation. This review discusses recent developments in SCID identification and treatment. PMID- 15660529 TI - Interaction of U-box E3 ligase SNEV with PSMB4, the beta7 subunit of the 20 S proteasome. AB - Recognition of specific substrates for degradation by the ubiquitin-proteasome pathway is ensured by a cascade of ubiquitin transferases E1, E2 and E3. The mechanism by which the target proteins are transported to the proteasome is not clear, but two yeast E3s and one mammalian E3 ligase seem to be involved in the delivery of targets to the proteasome, by escorting them and by binding to the 19 S regulatory particle of the proteasome. In the present study, we show that SNEV (senescence evasion factor), a protein with in vitro E3 ligase activity, which is also involved in DNA repair and splicing, associates with the proteasome by directly binding to the beta7 subunit of the 20 S proteasome. Upon inhibition of proteasome activity, SNEV does not accumulate within the cells although its co localization with the proteasome increases significantly. Since immunofluorescence microscopy also shows increased co-localization of SNEV with ubiquitin after proteasome inhibition, without SNEV being ubiquitinated by itself, we suggest that SNEV shows E3 ligase activity not only in vitro but also in vivo and escorts its substrate to the proteasome. Since the yeast homologue of SNEV, Prp19, also interacts with the yeast beta7 subunit of the proteasome, this mechanism seems to be conserved during evolution. Therefore these results support the hypothesis that E3 ligases might generally be involved in substrate transport to the proteasome. Additionally, our results provide the first evidence for a physical link between components of the ubiquitin-proteasome system and the spliceosome. PMID- 15660531 TI - Life on the exponential curve--time to rattle the academic cage? A view from the street. AB - The history of health service research has been characterized by an overwhelming volume of literature that has little impact on those who actually get on and do the work. The focus has been on an examination of why evidence is not accommodated into practice and how the barriers to implementation can be reduced. The fact that the evidence-based product may not be relevant to those at whom it is directed had not until recently been considered a possibility. Over the past 20 years there has been a consolidation of two cultures in the National Health Service--the academic/researcher and the clinician/practitioner. This paper sets the two cultures within the context of a community of practice framework and argues that the emphasis should move away from managing the interface to a fundamental reappraisal of the health service academic community. PMID- 15660532 TI - Development and psychometric properties of the Individualized Care Scale. AB - RATIONALE, AIMS AND OBJECTIVES: In this study we describe the development of the Individualized Care Scale (ICS) and evaluate its validity, psychometric properties and feasibility. The ICS was designed to measure patients' views on how individuality is supported through specific nursing interventions (ICA) and how they perceive individuality in their own care (ICB) during hospitalization. METHOD: Three different data sets were collected among patients being discharged from hospital (n1 = 203, n2 = 279, n3 = 454). This bipartite 38-item ICS promises to be a brief, timely, easy to administer and useful self-completion measure for evaluating clinical nursing practice from the patient's point of view. RESULTS: The findings supported the internal consistency reliability of the ICS (alpha 0.94 for ICA and ICB 0.93) and the three subscales (alphas 0.85-0.90). Item analysis supported the item construction of each scale. Content validity was furthered by a critical literature review and four expert analyses. Principal component analysis (Promax with Kaiser normalization) among earlier factor analyses supported construct validity by generating a three-factor solution which accounted for 65% of the variance in the ICA and 61% in the ICB. Pearson's correlation coefficients were at least 0.88 between the subscales and the total domain ICA or ICB. CONCLUSIONS: The ICS has demonstrated promise as a tool for measuring patients' evaluations of their hospital experience and individuality in care. PMID- 15660533 TI - Misrepresentations about palliative options and prognosis in motor neurone disease: some legal considerations. AB - If euthanasia were legalized, clinicians would be under a duty to explain to patients requesting euthanasia what the prognosis and palliative options were. Anecdotal evidence suggests that some patients suffering from motor neurone disease may request euthanasia because of a fear of choking to death. The literature indicates that with competent palliative care this is unlikely to occur. It is assumed, for the purposes of argument, that responsible clinicians would accordingly reassure patients that such a fear was unwarranted, and that such a reassurance would cause patients for whom choking was the main concern either to withdraw a request for euthanasia or not to make it in the first place. The legal consequences of both negligent and deliberate failure to represent the true prognostic and palliative situation are discussed. In the case of a deliberate failure, with the intention to induce the patient to consent to euthanasia, it is suggested that a verdict of murder would be ethically right. It is argued that such a failure is best regarded as an omission. The English law currently does not countenance the possibility of murder by omission. It is suggested that the distinction between acts and omissions, while sometimes convenient, can sometimes produce injustice, and that the distinction should not be allowed to prevent conviction for murder where this is clearly appropriate. PMID- 15660534 TI - Breast cancer incidence: what do the figures mean? AB - ABSTRACT RATIONALE, AIMS AND OBJECTIVES: Against the background of a general rise in the incidence of breast cancer temporal variations appear in different age groups, including changes attributable to the introduction of breast screening. This paper attempts to distinguish the influence of breast screening and to examine any underlying patterns. METHODS: The incidence data published by the Office of National Statistics for England and Wales have been examined using the method of least squares regression. These data have been subjected to major revision: a new system was introduced in 1971. That year, therefore, has been taken as the starting point for the analyses. Regression lines were fitted from 1971, 1984 and 1988 (when breast screening was started). Results Between 1971 and 1984 breast cancer incidence was unchanged in women under 50 years. In all other age groups and time periods the incidence has risen throughout 1971-1999. The rate of increase remained constant for women over 64 but changed in 1984 for all women under 65 years. The rate of increase is rising exponentially with age since 1984 in the women under 65. The superimposed increases due to screening have been separated from the underlying rates. CONCLUSIONS: In the period surveyed there appears to have been a change in the rates of increase in incidence of breast cancer in women under 65 that dates from 1984. This change can be separated from the spurious increase due to breast screening. The rates in women over 64 have remained constant. PMID- 15660530 TI - Homodimerization of calpain 3 penta-EF-hand domain. AB - Calpains 1 and 2 are heterodimeric proteases in which large (relative molecular mass M(r) 80000) and small (M(r) 28000) subunits are linked through their respective PEF (penta-EF-hand) domains. The skeletal muscle-specific calpain 3 is believed not to form a heterodimer with the small subunit but might homodimerize through its PEF domain. Size-exclusion chromatography and analytical ultracentrifugation of the recombinant PEF domain of calpain 3 show that it forms a stable homodimer that does not dissociate on dilution. Molecular modelling suggests that there would be no barriers to the dimerization of the whole enzyme through the PEF domains. This orientation would place the catalytic centres at opposite ends of the dimer. PMID- 15660535 TI - Demographic and clinical characteristics of patients admitted to medical departments. AB - INTRODUCTION: The significant changes, which have affected departments of medicine over the last two decades, have been much editorialized. Surprisingly few data have been published that document these changes. We describe the demographic and clinical characteristics of patients admitted to one hospital's departments of internal medicine. METHODS: During a 3 months prospective study, 1039 patients (46% of all 2277 new admissions to the departments of medicine, consisting of +/- 175 beds) were randomized for inclusion. Information was extracted from the medical records regarding demography; functional capacity; clinical data and outcome. RESULTS: The mean age of the patients was 72 +/- 16 years (mean +/- SD); 51% were male, 56% were married, and 83% lived at home. Although 293 patients (28%) were defined as dependent, the majority lived at home (170, 58%). Cognition was normal in only 74%. The four most common disease categories leading to admission were infections (383 patients, 37%), cardiovascular disorders (372, 36%), respiratory conditions (284, 27%) and genito urinary problems (90, 9%). Overall, 98 persons died (9%). Independent risk factors for death were: mechanical ventilation; a 'do not resuscitate' order; a high APACHE-II score; a low serum albumin level; higher age; and not being married (P < 0.01). CONCLUSIONS: Geriatric patients occupy a central position in our medical wards. These data are relevant for allocation of special resources for departments with high proportions of geriatric patients; for the design of employment conditions that ascertains continuing job satisfaction; as well as for the planning of teaching opportunities for residents and students. PMID- 15660536 TI - Selecting a subjective health status measure for optimum utility in everyday orthopaedic practice. AB - BACKGROUND: The time required to complete patient outcome questionnaires such as the Western Ontario and McMaster Universities Osteoarthritis Index (WOMAC), the Short Form General Health Survey (SF-36) and the Musculoskeletal Function Assessment (MFA) can sometimes threaten elderly, compromised patients sufficiently to compromise compliance with follow-up clinics. Incomplete questionnaires can also present problems of data bias. A little used (in the USA), patient-friendly questionnaire, the Nottingham Health Profile (NHP) has the potential to reduce the statistical and practical problems associated with the more generally used instruments. We hypothesized that NHP will produce similar results to WOMAC and SF-36 and is more sensitive to small changes in patient outcomes than SF-36 and MFA. METHOD: Twenty-three patients blindly completed WOMAC, SF-36 and NHP questionnaires. Spearman's Rank Order Correlation was used to compare the component scores of each instrument. Simulation of the before and after results of 10 fictitious patient comparing MFA and NHP was conducted using the related sample t-test. RESULTS: Seven of nine correlation coefficients were statistically significant and ranged from 0.711 to 0.901. The significance of the before and after difference on the five-point scale response was P = 0.05 when the NHP was used and P = 0.07 when MFA was used. The before and after difference on 'yes-no' response questions was P < 0.001 when NHP was used but showed no difference when the MFA was used. CONCLUSIONS: Our hypothesis was supported and we suggest that NHP can be used with confidence as an alternative to other patient outcome instruments. PMID- 15660537 TI - Why the need to reduce medical errors is not obvious. AB - According to Leape & Berwick (2000) the need to reduce medical errors is 'obvious and the mandate is clear'. My article questions this assertion. I go beyond the unknown incidence of medical errors in a general medical population to suggest that the meaning of medical errors is itself equivocal. I contest the assumption that the 'wrongness' of medical errors is always problematic, arguing instead for a distinction between desirable errors and undesirable errors. This distinction takes into account the consequences of errors, and why they may occur. Reasons include the inappropriateness of two cultural contexts--evidence-based medicine and continuous quality improvement--within which patient safety standards can be constructed and hence, medical errors can be defined. PMID- 15660538 TI - Regression to the mean: treatment effect without the intervention. AB - Regression to the mean (RTM) is a widespread statistical phenomenon. It is a group phenomenon that occurs whenever an extreme group is selected from a population based on the measurement of a particular variable. If a second measurement is then taken for the same group, the second mean will be closer to the population mean than the first measurement. This decrease (or increase) can be mistakenly attributed to a treatment effect; the conclusion can be drawn that an effect results from treatment when it in fact results from chance. Any intervention that is aimed at a group or characteristic that is very different from the average will appear to be successful because of RTM. It is therefore important that any genuine reductions because of the treatment are separated out from the effect of RTM. If the problem is ignored then this will lead to errors in the interpretation of results and, potentially, decisions made on the evidence of those results. This paper highlights the importance of the issue and its effects on many common clinical, public health and managerial decisions. PMID- 15660539 TI - Hand-held miniaturized cardiac ultrasound instruments for rapid and effective bedside diagnosis and patient screening. AB - RATIONALE, AIMS AND OBJECTIVES: Technological progress in recent years has made it possible that ultrasound industry can now offer affordable, portable and battery-operated ultrasound systems the size of a laptop computer. The purpose of this study was to compare these hand-carried ultrasound instruments with standard echocardiography in order to investigate the facility of a rapid bedside diagnosis in patients with suspected or known cardiovascular disease. METHODS: Fifty consecutive patients were studied with miniaturized ultrasound equipment (SonoHeart) and a conventional scanner (Acuson Sequoia) in a blinded manner. All studies were performed by three board-certified cardiologists skilled and experienced in echocardiographic practice. Investigators were not aware of any previous medical reports. RESULTS: With the new system, adequate images could be obtained in all patients. Left ventricular and left atrial diameters measured with the hand-held system correlated well with those obtained with conventional scanning: r = 0.87, mean difference 3.12 +/- 2.7 mm and r = 0.84, mean difference 2.8 + 2.4 mm, respectively. The presence of left ventricular dysfunction, regional wall motion abnormalities, relevant valvular regurgitation (moderate or more) or valve stenosis was correctly diagnosed in all patients. However, there was a tendency towards underestimating the extent of wall motion abnormalities particularly in patients difficult to image. Discrepancies also frequently occurred in patients with trivial or mild regurgitation, where false-positive and false-negative findings were described. CONCLUSION: Currently available hand-held echocardiography systems can facilitate rapid bedside diagnosis and patient screening. However, this recent development in echocardiography also raises a number of questions and its actual impact on general clinical practice still remains to be evaluated. PMID- 15660540 TI - Evaluation of a service development to increase detection of urinary tract infections in children. AB - RATIONALE, AIMS AND OBJECTIVES: It is suspected that childhood urinary tract infection (UTI) remains under-diagnosed in primary care, and is consequently the cause of subsequent morbidity from renal scarring, hypertension and eventual renal failure. Practice-based education and service developments were undertaken to try to improve the detection of childhood UTI. METHODS: A controlled before and-after intervention study was conducted. The educational and service developments promoted awareness of and greater testing for UTI among children less than two years of age presenting with febrile illness or other potentially relevant symptoms or signs. Appropriate diagnostic equipment was provided. RESULTS AND CONCLUSIONS: More urine samples were sent by the intervention practices but without a concomitant increase in detection of UTIs. This may indicate that current practice is approaching near maximal detection of UTI in young children. PMID- 15660541 TI - Patient expectations in placebo-controlled randomized clinical trials. AB - OBJECTIVE: To explore participants' experience in placebo-controlled randomized clinical trials (RCTs) specifically in relationship to their expectations. BACKGROUND: Aspects of being in RCTs, such as informed consent, perception of benefit and understanding of randomization, have been examined. In contrast, little is known concerning the formation of patient expectations before and during trials. METHODS: Qualitative methods using in-depth interviews with a semi structured interview guide of nine patients from four different RCTs. Data analysis was conducted using a codebook format arranging participant responses under broad analytical headings. The interviewer used a semi-structured interview guide to direct the conversation from one broad topic to the next within the context of the ongoing conversation. A checklist of topics encouraged participants to describe their experiences in RCTs. Narratives concerning expectation, blinding and placebo were compared to identify common themes. RESULTS: Patient anticipatory processes were influenced and modified both before and during the trial from multiple inputs. Such factors as past experiences in RCTs, past experiences of ineffective treatment, stress of being off regular medications, fear of being a 'placebo responder', input of non-study doctors or other health professionals, the experience of other participants, measurements of health parameters made during the trial and the presence or absence of side effects all affected patient expectation. CONCLUSION: Expectations in RCTs are not fixed and instead may be viewed as continuously shaped by multiple inputs that include experience and information received both before and during the trial. Variability in placebo response observed in previous studies may be related to the fluid nature of expectations. Trying to control and equalize expectations in RCTs may be more difficult than previously assumed. PMID- 15660542 TI - The epidemiology of peripheral vein complications: evaluation of the efficiency of differing methods for the maintenance of catheter patency and thrombophlebitis prevention. AB - BACKGROUND AND AIMS: Peripheral vein catheter patency and infusion thrombophlebitis remains a significant problem in everyday clinical practice. The aim of the study was to investigate the epidemiology of peripheral vein complications and to evaluate three different methods for the maintenance of peripheral vein catheter patency and the prevention of vein thrombophlebitis. METHODS: A total of 300 post-operative patients undergoing elective orthopaedic surgery were prospectively studied. Patients were divided into three groups: controls--catheters not flushed following drug administration; saline group--the catheters flushed with 3 mL of normal saline 0.9% after each catheter use; heparin group--the catheters flushed with 3 mL of 100-U/mL heparin in normal saline 0.9% after each catheter use. RESULTS: Complications occurred in 36% of the patients and the incidence of thrombophlebitis was 8% and only 4% in the control group. In the normal saline group there was a significant increase in total complications and obstruction together with thrombophlebitis as compared with the control group. Kaplan-Meier curves demonstrate that the control group had a significantly higher proportion of catheters without complications. CONCLUSIONS: The use of normal saline solutions in post-operative patients thus should be avoided for catheter maintenance. In patients receiving low molecular weight heparin, no intravenous flushing should be used for preventing catheter obstruction or thrombophlebitis in order to reduce costs and nursing workload. PMID- 15660543 TI - Peri-operative MRSA policy for orthopaedic patients. PMID- 15660544 TI - Pre-operative planning and the role of templating in total knee arthroplasty. PMID- 15660545 TI - Audit of the management of facial lacerations in accident and emergency department: wound closure without appropriate training or guidelines. PMID- 15660547 TI - Music improves sleep quality in older adults. AB - AIM: The aim of this paper is to report an investigation of the effects of soft music on sleep quality in older community-dwelling men and women in Taiwan. BACKGROUND: Sleep is a complex rhythmic state that may be affected by the ageing process. Few studies have focused on the effects of music, a non-pharmacological method of improving the quality of sleep in older adults. METHOD: A randomized controlled trial was used with a two-group repeated measures design. Sixty people aged 60-83 years with difficulty in sleeping were recruited through community leaders and screened using the Pittsburgh Sleep Quality Index (PSQI) and Epworth Sleepiness Scale. Those reporting depression, cognitive impairment, medical or environmental problems that might interfere with sleep; and those who used sleeping medications, meditation, or caffeine at bedtime were excluded. Participants listened to their choice among six 45-minute sedative music tapes at bedtime for 3 weeks. There were five types of Western and one of Chinese music. Sleep quality was measured with the PSQI before the study and at three weekly post-tests. Groups were comparable on demographic variables, anxiety, depressive symptoms, physical activity, bedtime routine, herbal tea use, napping, pain, and pretest overall sleep quality. RESULTS: Music resulted in significantly better sleep quality in the experimental group, as well as significantly better components of sleep quality: better perceived sleep quality, longer sleep duration, greater sleep efficiency, shorter sleep latency, less sleep disturbance and less daytime dysfunction (P = 0.04-0.001). Sleep improved weekly, indicating a cumulative dose effect. CONCLUSION: The findings provide evidence for the use of soothing music as an empirically-based intervention for sleep in older people. PMID- 15660548 TI - Risk factors for chronic insomnia following hospitalization. AB - AIM: This paper reports a study whose purpose was to determine whether there is an increase in the incidence of chronic insomnia following hospitalization and, if so, to identify patients at risk. BACKGROUND: The consequences of difficulty sleeping in hospital have received scant attention from clinicians or researchers. Implicit in this lack of interest is the assumption that difficulty in sleeping is a transient reaction to hospitalization that will resolve on discharge, an assumption not empirically supported. It has been argued that in susceptible people this type of temporary disruption to sleep can be the catalyst for the development of chronic insomnia. METHOD: Established sleep and depression rating instruments were used to monitor the sleep of 57 cardiac and 29 orthopaedic patients after elective surgery (n = 86), recruited through a hospital preadmission clinic. RESULTS: Preadmission chronic insomnia of 10% was consistent with general population prevalence estimates of 6-12%. Three months after discharge the incidence had almost doubled to 19%. Sixty-one per cent of this variance could be explained by hyperarousal, sleep hygiene issues, and dysfunctional cognitions about sleep. Depression was found to be a salient predictor but not an independent risk factor. Age, sex, and hospital-related data, such as score for difficulty sleeping in hospital, proved to be statistically insignificant. CONCLUSIONS: The results support the role of hyperarousal and dysfunctional sleep attitudes and behaviours as stronger predictors of chronic insomnia than patient demographics or environmental issues. Given that most of the patients were ambivalent about how they slept in hospital, with high satisfaction (71%) in the presence of significant disruption (63%), preadmission sleep education given to these patients prior to admission potentially contributed to the development of more realistic expectations of the quality of in-hospital sleep. PMID- 15660549 TI - Understanding personal narratives: an approach to practice. AB - AIM: This paper explores the need for and nature of personal narratives and their relevance to nursing practice. It proposes that the co-creative aesthetic process can be used to understand and co-create personal narratives through an emphasis on self-defining memories and metaphor. BACKGROUND: Many authors in nursing and other human sciences have recognized the need for and importance of personal narrative, its relationship to aesthetic knowing and its value in qualitative research and in practice. The role of memory and metaphor in the creation of meaning in personal narratives, however, has not been sufficiently explored in nursing literature. DISCUSSION: The nature of personal narrative is explored, focusing on the way meaning is created from self-defining memories using metaphor. Then, the importance of personal narratives in nursing practice is considered, followed by discussion about how meaning in personal narratives may be co-created between clients and nurses using an aesthetic process developed by the author. CONCLUSION: The co-creative aesthetic process is an example of nursing as art and can be used to co-create personal narratives in practice. The experience of co-creating a self story with a nurse can be healing, as the self story is heard by a caring person, memories are understood in new ways, and the self story is both confirmed and recreated. PMID- 15660550 TI - Spiritual care nursing: what cancer patients and family caregivers want. AB - AIM: This paper presents findings from a study that was designed to understand, from the perspective of cancer patients and their family caregivers, what spiritual care is wanted from nurses. BACKGROUND: Distressing and transformative spiritual responses to living with cancer have been documented. Although there is momentum for providing spiritual care, previous research provides scanty and conflicting evidence about what are the clients' wishes or preferences with regard to receiving spiritual care from nurses. METHODS: A convenience sample of 156 adult cancer patients and 68 primary family caregivers, most of whom were Christians, independently completed the Spiritual Interests Related to Illness Scale and a demographic form, both of which were self-completed questionnaires. RESULTS: A variation in responses to items about nurses providing spiritual care therapeutics was observed; means and medians for these items mostly fell between 2 (disagree) and 3 (agree) on a scale of 1-4. Generally, therapeutics that were less intimate, commonly used, and not overtly religious were most welcomed. No significant differences were found between patient and caregiver preferences. A modest, direct correlation was observed between frequency of attendance at religious services and increased preference for nurse spiritual care. CONCLUSION: For both patients and caregivers, nurses must be sensitive to providing spiritual nurture in ways that are welcomed. PMID- 15660551 TI - Smoking among Japanese nursing students: nationwide survey. AB - AIM: This paper reports a study estimating the nationwide prevalence of and attitudes towards smoking among Japanese nursing students. BACKGROUND: The World Health Organization established "World No Tobacco Day" in 1987, and has been promoting antismoking measures worldwide since then, with annual themes. It has emphasized that health care professionals, including nurses, as role models for healthy living, should not smoke, and that as promoters of health education they should not seem to justify or condone their patients' smoking. To promote antismoking measures among nurses, it will be necessary to scrutinize the smoking habits and behavior of nursing students and associated factors, and to conduct effective antismoking education and health education before they acquire the smoking habit. METHODS: A cross-sectional survey was carried out through self administered, closed-ended, structured questionnaires. Questionnaires were mailed to 4169 nursing students at 27 randomly selected vocational nursing schools nationwide. Smoking status, history, and attitudes towards smoking were examined. RESULTS: Smoking prevalence among female nursing students was 23.5%, which was higher than that among the Japanese general female population aged 20-29 (21.9%). Smoker-students were significantly more positive toward smoking than non-smokers in all opinions about health care professionals' smoking. Multiple logistic regression analysis showed that smoking statuses of the people around the participants, dissatisfaction with being a nursing student, and living alone were associated with participants' smoking behaviour. CONCLUSIONS: The results of the present study suggest an urgent necessity to provide effective antismoking measures for nursing students. PMID- 15660553 TI - Non-somatic effects of patient aggression on nurses: a systematic review. AB - AIM: This paper describes a systematic review of the predominant non-somatic effects of patient assault on nurses. Background. Patient aggression towards nurses is a longstanding problem in most nursing domains. Although reports on the consequences of physical aggression are more numerous, the non-physical effects create much suffering. METHOD: A systematic review of literature from 1983 to May 2003 was conducted using the Medline, CINAHL, PsychINFO and PSYINDEX databases. Articles from international journals in English or German and reporting at least three non-somatic responses to patient aggression were included. FINDINGS: The electronic search produced 6616 articles. After application of the inclusion and exclusion criteria, 25 texts from eight countries and four domains of nursing remained. Twenty-eight main effects were found, and these were categorized using a system suggested by Lanza and including bio-physiological, emotional, cognitive, and social dimensions. The predominant responses were anger, fear or anxiety, post-traumatic stress disorder symptoms, guilt, self-blame, and shame. These main effects occurred across most countries and nursing domains. CONCLUSION: Despite differing countries, cultures, research designs and settings, nurses' responses to patient aggression are similar. Standardized questionnaires could help improve estimations of the real prevalence of non-somatic effects. Given the suffering caused by non-somatic effects, research should be aimed at preventing patient aggression and at developing better ways to prepare nurses to cope with this problem. PMID- 15660552 TI - NHS cadet schemes: do they widen access to professional healthcare education? AB - AIM: The aim of this paper is to report a study investigating the extent to which National Health Service cadet schemes widen access to professional health care education. BACKGROUND: Cadet schemes have been reintroduced in the United Kingdom to increase recruitment of nurses and other health care staff to the National Health Service and also to widen access and increase participation in professional health care education by groups poorly represented in such education, including minority ethnic groups. METHODS: A questionnaire survey of all cadet schemes (n = 62) in England at the time of the study was carried out, and the respondents were cadet scheme leaders (n = 62) and cadet students (n = 411). The questionnaires to scheme leaders enquired about when the schemes were established, what the schemes were preparing cadets for, modes of delivery and entry qualifications. The questionnaires to cadets enquired about age, gender, family circumstances, prior experience and ethnic background. FINDINGS: The majority of schemes had been established since the health service reforms of 1999 and most were preparing cadets to enter professional nurse education programmes. Very few provided opportunities for part-time study and some asked for entry qualifications. Cadets were younger on entry than a comparator group of student nurses, fewer were married, fewer had previous employment or health-related employment and a lower percentage of cadets were white. CONCLUSION: Cadet schemes have the potential to widen access to professional health care study, but there is only limited evidence that they are doing so. In particular there was a lack of mature entrants to health care professional education via the schemes. However, the majority of schemes offered a route into professional education for students who did not hold sufficient educational qualifications for direct entry to professional health care education. It is encouraging that cadet schemes appear to be attracting a significantly greater proportion of students from Black and minority ethnic groups than preregistration nursing programmes overall. PMID- 15660554 TI - Respite care for caregivers and people with severe mental illness: literature review. AB - AIM: The aim of this study was to review research literature over the past 10 years on respite care for people affected by severe mental illness; and identify key implications for nursing practice in provision of respite care for family caregivers of people with severe mental illness. BACKGROUND: Family caregivers play an important role in health care, but need regular breaks to maintain their own health and well-being. Respite care is one of the few services available with a primary focus on supporting family caregivers. In most developed countries the notion of respite care as an extension of the health care service has been embraced, evidenced by a growing body of literature in health and health-related disciplines. METHODS: An initial literature search was undertaken using the key words "respite", "short-term care", "shared care" and "day care" in major electronic databases for nursing, psychiatry, psychology and sociology literature between 1967 and 2002, identifying 704 articles. Closer examination of the literature from 1993 to 2002 on gaps and trends in respite care for people affected by severe mental illness was conducted. This is discussed in the context of the broader literature, particularly on dementia, where the mainstream research on respite care is found. RESULTS: The majority of family caregiving studies identified a need for greater quality, quantity, variety and flexibility in respite provision, and the literature has remained largely silent in relation to those affected by severe mental illness. There are contradictory findings on outcomes of respite care services and a lack of controlled empirical studies and evaluative research on effectiveness. CONCLUSIONS: Respite care is beneficial for caregivers, there is significant unmet need in provision of services for the mentally ill, and greater flexibility and the needs of caregivers should be recognised and addressed. PMID- 15660555 TI - Changes in community nursing in Australia: 1995-2000. AB - AIM: This paper reports a study whose aim was to examine the congruence between community nurses' perceptions and the realities of changes in their work. BACKGROUND: There have been increasing challenges to the delivery of community nursing services in Australia over the past decade. Acute care sector changes and the recent focus on health promotion, prevention, early identification and intervention adds pressure and creates tensions for community nurses, which are well-documented in the literature. There is, however, a lack of empirical evidence of actual changes in community nurses' workloads and the focus of their work. Validation of nurses' perceptions would enable them to have a stronger voice in the future development of community health care. METHODS: Four sources of data were used: community health client administrative data 1995-2000; occasions of service data 1995-2000; staffing numbers 1998-2001; and interviews with 14 community nurses in late 2001. RESULTS: Documentary evidence shows that there has been a large increase in the number of adult clients, and all clients are increasingly receiving a shorter, more intensive, clinically focussed service and are then discharged from care, rather than receiving a lower intensity service over a longer period of time. Staffing numbers have not increased to match this higher acuity and intensity. These changes were echoed by the nurses, who reported that expanded acute care roles were impacting on their workload and resulting in a loss of holistic primary health care focus. There has been a lack of leadership and proactive planning by community nurses in response to these changes. CONCLUSION: Community health care in Australia is shifting from primary to short-term clinical care. Greater opportunities for community nurses to engage proactively in defining and promoting their role in the health care system are needed in order to ensure an appropriate balance of acute clinical and holistic primary health care in the community. PMID- 15660557 TI - AIDS epidemic poses serious threat to Europe. PMID- 15660556 TI - Towards a typology of nursing turnover: the role of shocks in nurses' decisions to leave. AB - AIMS: The paper reports a study to explore the decision process nurses go through before leaving, focusing on leaving decisions that are precipitated by a single, jarring event or shock. BACKGROUND: Nursing turnover is a significant problem. Although a range of initiatives has been adopted to improve retention, recent insights from the academic literature on labour turnover have additional implications for how this problem might be managed. METHOD: A structured questionnaire, with some open-ended items, was used to collect data. For respondents who reported a shock (n = 153), responses were cluster analysed (hierarchical, agglomerative clustering generated a solution and k-means clustering enhanced the solution). Clusters were validated using responses to open items. RESULTS: There were three broad clusters of nursing turnover: cluster 1 described nurses whose decision to leave was precipitated by a shock that was work-related, negative and unexpected; cluster 2 described those whose decision was precipitated by a shock that was personal, positive and expected; cluster 3 describes those whose decision unfolded more gradually. Cluster 3 described the conventional picture of how turnover occurs (i.e where there is no shock), whereas clusters 1 and 2 were evidence of different types, where a shock prompts the quitting. CONCLUSION: In many cases of nurse turnover, a single, jarring event, or shock, initiates thoughts of quitting. Understanding the role of shocks has implications for a range of management activities. Allocation of education, promotion and distribution of other benefits should be managed in such a way as to minimize the likelihood of shocks. Profiling of nurse leavers should be undertaken so that managers have an accurate and detailed picture of turnover. PMID- 15660564 TI - Relationship between outdoor and indoor air quality in eight French schools. AB - In the frame of the French national research program PRIMEQUAL (inter-ministry program for better air quality in urban environments), measurements of outdoor and indoor pollution have been carried out in eight schools in La Rochelle (France) and its suburbs. The buildings were naturally ventilated by opening the windows, or mechanically ventilated, and showed various air permeabilities. Ozone, nitrogen oxides (NO and NO(2)), and airborne particle (particle counts within 15 size intervals ranging from 0.3 to 15 mum) concentrations were continuously monitored indoors and outdoors for two 2-week periods. The indoor humidity, temperature, CO(2) concentration (an indicator of occupancy), window openings and building permeability were also measured. The temporal profiles of indoor and outdoor concentrations show ozone and nitrogen oxides behave differently: NO and NO(2) indoor/outdoor concentration ratios (I/O) were found to vary in a range from 0.5 to 1, and from 0.88 to 1, respectively, but no correlation with building permeability was observed. On the contrary, I/O ratios of ozone vary in a range from 0 to 0.45 and seem to be strongly influenced by the building air-tightness: the more airtight the building envelope, the lower the ratio. Occupancy, through re-suspension of previously deposited particles and possible particle generation, strongly influences the indoor concentration level of airborne particles. However, this influence decreases with particle size, reflecting the way deposition velocities vary as a function of size. The influence of particle size on deposition and penetration across the building envelope is also discussed by analyzing the I/O ratios measured when the buildings were unoccupied, by comparing the indoor concentrations measured when the buildings were occupied and when they were not (O/U ratios), and by referring to previously published studies focussing on this topic. Except one case, I/O were found to vary in the range from 0.03 to 1.79. All O/U are greater than one and increase up to 100 with particle size. PRACTICAL IMPLICATIONS: Assessing children's total exposure requires the knowledge of outdoor and indoor air contaminant concentrations. The study presented here provides data on compared outdoor and indoor concentration levels in school buildings, as well as information on the parameters influencing the relationship between outdoor and indoor air quality. It may be used as a basis for estimating indoor concentrations from outdoor concentrations data, or as a first step in designing buildings sheltering children against atmospheric pollution. PMID- 15660565 TI - Correlation between temperature satisfaction and unsolicited complaint rates in commercial buildings. AB - This paper analyzes the relation between temperature satisfaction ratings expressed on a questionnaire and unsolicited complaint rates recorded in a maintenance database. The key findings are as follows: (i) the satisfaction ratings and complaint rates are negatively correlated with a moderate magnitude (r(s) = -0.31 to -0.36), and the correlation is statistically significant (P = 0.01-0.005), and (ii) the percent dissatisfied with temperature and the complaint rate are positively correlated with moderate magnitude (r(s) = 0.31-0.36), and the correlation is statistically significant (P = 0.01-0.004). Both data sets contain 'real-world' measures of temperature satisfaction, with the complaints contributing directly to the cost of operations and maintenance. The relationship between two validates a new method of assessing the economic cost of thermal discomfort in commercial buildings. PRACTICAL IMPLICATIONS: Complaints in commercial buildings indicate occupants' dissatisfaction to their environments. It not only deteriorates occupants' performance and organization productivity, but also increases building maintenance and operating cost. Nailing economic consequences of complaints will enable monetary comparison of discomfort cost with building and operating costs. This comparison may be desirable for building owners and tenants to make well-informed decisions on construction, rental, and retrofit. It may also be used to evaluate complaint diagnostic and eliminating techniques. PMID- 15660566 TI - Changing microbial concentrations are associated with ventilation performance in Taiwan's air-conditioned office buildings. AB - Our study conducted serial environmental measurements in 12 large office buildings with two different ventilation designs to obtain airborne microbial concentrations in typical office buildings, and to examine the effects of occupant density, ventilation type and air exchange efficiency on indoor microbial concentrations. Duplicate samples of airborne fungi and bacteria, a total of 2477 measurements, were collected based on a scheme of conducting sampling three times a day for at least seven consecutive days at every study building. Air change rates (ACHs) were also estimated by tracer gas concentration decay method, and measured by continuous Multi-Gas monitor for each building. Most sampling sites were with total fungal and bacteria concentrations higher than 1000 CFU/m(3), an often-quoted guideline in earlier research. Significantly higher concentrations of fungi and bacteria, as well as higher indoor/outdoor (I/O) ratios across most groups of airborne microbes, were identified in buildings with fan coil unit (FCU) system than those with air-handling unit (AHU) system (Student's t test, P < 0.0001). Older buildings and higher air exchange rates were statistically associated with greater indoor bacteria levels in FCU ventilated buildings (R(2) = 0.452); a pattern not found in AHU buildings. Increasing ACH seemed to be the determinant factor for rising indoor fungal and Cladosporium concentrations in those FCU buildings (R(2) = 0.346; 0.518). Our data indicated that FCU ventilated buildings might have provided more outdoor matters into indoor environments through direct penetration of outdoor air. Results also demonstrated a quantitative association between rising numbers of occupants and increasing indoor levels of yeast in both FCU and AHU ventilated buildings. The regression model identified in this study may be considered a reference value for proposing an optimal ACH, while with adequate filtration of fresh air, as an effective strategy in lowering indoor microbial concentrations in air-conditioned buildings. PRACTICAL IMPLICATIONS: As control of indoor microbial contamination has become an increasing concern around the world, feasibility and effectiveness of adopting ventilation approach has attracted a significant interest. This field investigation demonstrated, quantitatively, critical variables to be taken into consideration while applying such a measure, including the kinds of microbes to be removed and the types of ventilation system already in place. PMID- 15660567 TI - Do indoor pollutants and thermal conditions in schools influence student performance? A critical review of the literature. AB - To assess whether school environments can adversely affect academic performance, we review scientific evidence relating indoor pollutants and thermal conditions, in schools or other indoor environments, to human performance or attendance. We critically review evidence for direct associations between these aspects of indoor environmental quality (IEQ) and performance or attendance. Secondarily, we summarize, without critique, evidence on indirect connections potentially linking IEQ to performance or attendance. Regarding direct associations, little strongly designed research was available. Persuasive evidence links higher indoor concentrations of NO(2) to reduced school attendance, and suggestive evidence links low ventilation rates to reduced performance. Regarding indirect associations, many studies link indoor dampness and microbiologic pollutants (primarily in homes) to asthma exacerbations and respiratory infections, which in turn have been related to reduced performance and attendance. Also, much evidence links poor IEQ (e.g. low ventilation rate, excess moisture, or formaldehyde) with adverse health effects in children and adults and documents dampness problems and inadequate ventilation as common in schools. Overall, evidence suggests that poor IEQ in schools is common and adversely influences the performance and attendance of students, primarily through health effects from indoor pollutants. Evidence is available to justify (i) immediate actions to assess and improve IEQ in schools and (ii) focused research to guide IEQ improvements in schools. PRACTICAL IMPLICATIONS: There is more justification now for improving IEQ in schools to reduce health risks to students than to reduce performance or attendance risks. However, as IEQ-performance links are likely to operate largely through effects of IEQ on health, IEQ improvements that benefit the health of students are likely to have performance and attendance benefits as well. Immediate actions are warranted in schools to prevent dampness problems, inadequate ventilation, and excess indoor exposures to substances such as NO(2) and formaldehyde. Also, siting of new schools in areas with lower outdoor pollutant levels is preferable. PMID- 15660569 TI - Association of air-conditioning with respiratory symptoms in office workers in tropical climate. AB - To evaluate the association of heating, ventilation and air-conditioning systems (HVAC) and respiratory symptoms in a tropical city, self-administered questionnaires were given to 2000 individuals working in air-conditioned office buildings and to 500 control workers in naturally ventilated buildings. Reported symptoms from the two populations were analyzed using chi-square tests, univariate and multiple logistic regressions models. Symptoms were the outcome variable and the odds ratios were adjusted by gender, age, accumulated work time, smoking habits and atopic background. There was a 79.8% response rate and there was a positive association of nasal symptoms (odds ratio, OR = 1.59, 95% confidence interval, CI = 1.11-2.28), naso-ocular symptoms (OR = 1.58, 95% CI = 1.05-2.38), persistent cough (OR = 3.04, 95% CI = 2.00-4.63) sinusitis symptoms (OR = 1.85, 95% CI = 1.27-2.71) and building-related worsening of the symptoms (OR = 4.92, 95% CI = 2.93-8.27) with working in air-conditioned buildings. In conclusion, our study suggests that artificial air-conditioning is a matter of concern for respiratory symptoms in cities with hot and humid climate. PRACTICAL IMPLICATIONS: This study suggests that indoor air-related respiratory symptoms are a matter of concern in places with hot and humid climate. The regression models were adjusted by confounders that could be used in further reanalysis of indoor air quality related symptoms and ventilation systems with expanded variety of climatic conditions. PMID- 15660568 TI - Functional group characterization of indoor, outdoor, and personal PM: results from RIOPA. AB - Fourier transform infrared (FTIR) spectra of outdoor, indoor, and personal fine particulate matter (PM(2.5)) samples were collected during the Relationship of Indoor, Outdoor, and Personal Air (RIOPA) study. FTIR spectroscopy provides functional group information about the entire PM(2.5) sample without any chemical preparation. It is particularly important to characterizing the poorly understood organic fraction of PM(2.5). To our knowledge this is the first time that FTIR spectroscopy has been applied to a PM(2.5) exposure study. The results were used to chemically characterize indoor air and personal exposure. Sulfate was strongest in outdoor samples, which is consistent with the generally accepted understanding that sulfate is of outdoor origin. Absorbances attributed to soil dust were also seen in many outdoor and some indoor and personal samples. Inorganic nitrate absorbances were a common feature of many California and some New Jersey samples. Carbonyl absorbances showed substantial variation in strength, number of peaks, and wave number shift between samples, indicating variability in composition and sources. Absorbances attributed to aliphatic hydrocarbon and amide functional groups were enhanced in many personal and indoor samples, which suggested the influence of indoor sources in these homes. We speculate that meat cooking is one possible source of particulate amides. PRACTICAL IMPLICATIONS: To our knowledge this is the first time that FTIR spectroscopy has been used to characterize the composition of indoor and personal PM(2.5). The presence of sulfate, nitrate, ammonium, soil dust and a number of organic functional groups are all detected in one analysis on filter samples without extraction or other sample preparation. Differences between indoor and outdoor spectra are used to identify spectral features due to indoor-generated PM(2.5). Particularly interesting are the much larger aliphatic absorbances, shifts in carbonyl absorbances, and occasional small amide absorbances found in indoor and personal spectra but rarely in outdoor spectra. These observations are important because organics make up a large portion of PM(2.5) mass and their composition and properties are poorly characterized. The properties and behavior of organic compounds in airborne particles are often predicted based on their functional group composition. This analysis begins the development of a better understanding of the functional group composition of indoor and personal PM(2.5) and how it differs from that of outdoor PM(2.5). Eventually this will lead to an improved understanding of the properties, behavior and effects of PM(2.5) of indoor and outdoor origin. PMID- 15660572 TI - The evidence against malnutrition as a prominent problem for chronic dialysis patients. AB - A large number of dialysis patients have been classified as being malnourished because of anthropometric measurements and laboratory values that are similar to those in patients with protein-energy malnutrition. Malnutrition implies that abnormalities can be reversed with better nutrition. However, this approach of providing more nutrients in the diet has been unsuccessful in reversing these changes, suggesting that a diagnosis of malnutrition is generally a misdiagnosis for dialysis patients In this review we discuss mechanisms that cause a loss of protein stores, muscle mass, albumin, and other plasma proteins in dialysis patients. We also review the shortcomings of techniques that are used to measure the nutritional status of these patients. PMID- 15660573 TI - Serum albumin: relationship to inflammation and nutrition. AB - Hypoalbuminemia is the result of the combined effects of inflammation and inadequate protein and caloric intake in patients with chronic disease such as chronic renal failure. Inflammation and malnutrition both reduce albumin concentration by decreasing its rate of synthesis, while inflammation alone is associated with a greater fractional catabolic rate (FCR) and, when extreme, increased transfer of albumin out of the vascular compartment. A vicious cascade of events ensues in which inflammation induces anorexia and reduces the effective use of dietary protein and energy intake and augments catabolism of the key somatic protein, albumin. Hypoalbuminemia is a powerful predictor of mortality in patients with chronic renal failure, and the major cause of death in this population is due to cardiovascular events. Inflammation is associated with vascular disease and likely causes injury to the vascular endothelium, and hypoalbuminemia as two separate expressions of the inflammatory process. Albumin has a myriad of important physiologic effects that are essential for normal health. However, simply administering albumin to critically ill patients with hypoalbuminemia has not been shown to improve survival or reduce morbidity. Thus the inference from these clinical studies suggests that the cause of hypoalbuminemia, rather than low albumin levels specifically, is responsible for morbidity and mortality. PMID- 15660574 TI - C-reactive protein and end-stage renal disease. AB - The significance of CRP and inflammation has increased over time, especially in the end-stage renal disease (ESRD) population. From a simple marker it now appears that CRP is an active participant in pro-atherosclerotic phenomenon including local pro-inflammatory and thrombotic events. Studies in the general population indicate the usefulness of CRP in prognostication and in monitoring response to therapy. The clinical usefulness of CRP monitoring in chronic kidney disease (CKD) and especially in ESRD deserves closer study. In the meantime, the utility of CRP measurements for monitoring and treatment is on a case-by-case basis. Management of traditional cardiovascular risk factors should be considered. In the interest of optimizing therapy it is prudent to use biocompatible membranes and ultrapure water. A careful search for infectious processes in dialysis patients is recommended, with special attention to vascular access sites, periodontitis, gastritis, and other potentially chronic or covert infections. ACE-inhibitor use should be maximized in all eligible CKD patients. The data on the use of statins in ESRD have been generally positive but await further validation. Individualized use for selected patients is probably beneficial. PMID- 15660575 TI - Linking oxidative stress and inflammation in kidney disease: which is the chicken and which is the egg? AB - For end-stage renal disease (ESRD) patients, cardiovascular disease remains the single most common cause of excess morbidity and mortality. Furthermore, although the prevalence of traditional cardiovascular risk factors is high in the dialysis population, the extent and severity of associated cardiovascular morbidity and mortality remain disproportionate to traditional risk factor profiles. Consequently, considerable effort has been focused on "nontraditional" risk factors for cardiovascular events in this patient population. Among the examined nontraditional risk factors, increased oxidative stress as well as increased acute phase inflammation are postulated to be important contributors to uremic cardiovascular risk. Additional important uremic cardiovascular risk factors include malnutrition and endothelial dysfunction, both of which may be directly linked to the processes that cause increased oxidative stress and inflammation in uremia. In this context I review available data linking the pathogenesis of oxidative stress to acute phase inflammation and uremia. I also review data suggesting that oxidative stress in uremia directly contributes to the development of acute phase inflammation and that patients with higher levels of inflammation have higher levels of oxidative stress biomarkers. Similarly I review emerging data on the potential effects of antioxidant therapy on inflammatory biomarkers, as well as data suggesting that strategies to lower acute phase inflammation may also improve biomarkers of oxidative stress. Theoretical constructs evaluating the linkage of oxidative stress and inflammation in uremia and their contribution to the pathogenesis of atherosclerosis are suggested. PMID- 15660576 TI - Metabolic acidosis and malnutrition-inflammation complex syndrome in chronic renal failure. AB - Metabolic acidosis, a common condition in patients with renal failure, may be linked to protein-energy malnutrition (PEM) and inflammation, together also known as malnutrition-inflammation complex syndrome (MICS). Methods of serum bicarbonate measurement may misrepresent the true bicarbonate level, since the total serum carbon dioxide measurement usually overestimates the serum bicarbonate concentration. Moreover, the air transportation of blood samples to distant laboratories may lead to erroneous readings. In patients with chronic kidney disease (CKD) or end-stage renal disease (ESRD), a significant number of endocrine, musculoskeletal, and metabolic abnormalities are believed to result from acidemia. Metabolic acidosis may be related to PEM and MICS due to an increased protein catabolism, decreased protein synthesis, endocrine abnormalities including insulin resistance, decreased serum leptin level, and inflammation among individuals with renal failure. Evidence suggests that the catabolic effects of metabolic acidosis may result from an increased activity of the adenosine triphosphate (ATP)-dependent ubiquitin-proteasome and branched chain keto acid dehydrogenase. In contrast to the metabolic studies, many epidemiologic studies in maintenance dialysis patients have indicated a paradoxically inverse association between mildly decreased serum bicarbonate and improved markers of protein-energy nutritional state. Hence metabolic acidosis may be considered as yet another element of the reverse epidemiology in ESRD patients. Interventional studies have yielded inconsistent results in CKD and ESRD patients, although in peritoneal dialysis patients, mitigating acidemia appears to more consistently improve nutritional status and reduce hospitalizations. Large-scale, prospective randomized interventional studies are needed to ascertain the potential benefits of correcting acidemia in malnourished and/or inflamed CKD and maintenance hemodialysis patients. Until then, all attempts should be made to adhere to the National Kidney Foundation Kidney Disease and Dialysis Outcome Quality Initiative guidelines to maintain a serum bicarbonate level in ESRD patients of at least 22 mEq/L. PMID- 15660577 TI - Anthropometric and body composition assessment in dialysis patients. AB - Anthropometric and body composition assessments provide important information about the nutritional status of dialysis patients. Anthropometric measurements describe body size, fatness, and leanness in dialysis patients and have been collected in the Modification of Diet in Renal Disease (MDRD) and HEMO studies. Dialysis patients present special problems for anthropometry, including decreased functional status and increased comorbidity, that challenge nutrition assessment methodology. Recumbent anthropometric techniques are recommended and stature is estimated from knee height. Measures of weight, stature, calf circumference, arm circumference, and triceps and subscapular skinfolds have recently been reported for dialysis patients, who tend to be shorter, lighter, and have less adipose tissue than healthy persons of the same age. The HEMO study anthropometric data provide a clinical reference for assessing the nutritional status of dialysis patients. The most common body composition methods used with dialysis patients are dual energy X-ray absorptiometry (DEXA), bioelectrical impedance, total body water (TBW), and prediction equations, but they are not recommended for assessment of predialysis patients, as estimates are best obtained postdialysis. The TBW volume used in calculating the dose of dialysis has commonly been predicted from the limited, out-of-date equations of Watson, based on nonrepresentative samples. New prediction equations are available for white, black, and Mexican American children and adults. Watson's data are not representative of the TBW of U.S. men and women. The greater TBW in non-Hispanic black men and women and Mexican American women reflects the greater levels of obesity in the U.S. population. PMID- 15660578 TI - Protein and energy: recommended intake and nutrient supplementation in chronic dialysis patients. AB - Nutritional status is an important predictor of clinical outcome in end-stage renal disease (ESRD) patients, especially in patients on chronic hemodialysis. Uremic malnutrition is strongly associated with increased risk of death and hospitalization events in this patient population, and decreased muscle mass is the most significant predictor of these outcomes. Several factors that influence protein metabolism predispose chronic hemodialysis patients to increased catabolism and loss of lean body mass. The available evidence suggests that low protein and energy intake associated with advanced uremia along with catabolic consequences of dialytic therapies can lead to the development of uremic malnutrition. Recent studies show that the hemodialysis procedure induces a net protein catabolic state at the whole-body level as well as skeletal muscle. There is evidence to suggest that these undesirable effects are due to decreased protein synthesis and increased proteolysis. Provision of nutrients, either in the form of intradialytic parenteral nutrition or oral feeding during hemodialysis, can adequately compensate for the catabolic effects of the hemodialysis procedure. While the mechanisms of these effects are not studied in detail, changes in extracellular amino acid concentrations, along with certain anabolic hormones such as insulin, are important mediators of these actions. PMID- 15660579 TI - The dose of dialysis in hemodialysis patients: impact on nutrition. AB - Multiple lines of evidence have indicated that the dose of hemodialysis impacts upon patient outcome. Among these outcome measures, nutrition is inextricably linked to the adequacy of the treatment. All of the methods of determining dialysis adequacy are based on assessing the removal of toxic substances retained in renal failure, the majority of which are derivatives of protein metabolism. Urea kinetics, employing urea as a surrogate for quantifying the elimination of small molecular weight nitrogenous substances, is the method that has been most thoroughly validated to date as defining a dose range for thrice-weekly hemodialysis: Both inadequate and optimal levels of hemodialysis dose have been identified by prospective, randomized clinic trials utilizing Kt/V(urea) as the index of adequacy. The impact of urea kinetics on nutritional status during thrice-weekly hemodialysis is discussed. Recently, in an attempt to improve outcome beyond that achievable with thrice-weekly hemodialysis, alternative regimens, consisting of daily treatments, have received increasing interest. In order to compare the dose of hemodialysis associated with these regimens with conventional thrice-weekly regimens in terms of removal of small molecular weight substances, standard Kt/V(urea), a parameter that combines treatment dose with treatment frequency, and thus allows for various intermittent therapies to be compared to continuous therapy, must be used. In addition, membrane flux and middle molecule removal, factors that have not yet been well defined as parameters of adequacy during thrice-weekly regimens, may be shown to be important indices with longer hemodialysis treatments, particularly daily nocturnal hemodialysis. The impact that these alternative regimens have had on nutritional status in hemodialysis patients and how they compare to conventional therapy are important considerations. PMID- 15660580 TI - Ultrapure dialysate. AB - To prevent pyrogenic reactions during hemodialysis, it is recommended that bacteria and endotoxin in dialysate not exceed 100-200 colony forming units (CFU)/ml and 0.25-2 endotoxin units (EU)/ml, respectively. While these limits are adequate to prevent acute pyrogenic reactions, data are accumulating to suggest they may not prevent stimulation of chronic inflammation in hemodialysis patients. Fragments of endotoxin and other bacterial products capable of stimulating immune cells cross low-flux and high-flux membranes in vitro. In clinical studies, use of ultrapure dialysate (bacteria < 0.1 CFU/ml and endotoxin < 0.03 EU/ml) is associated with lower concentrations of inflammatory markers and acute phase reactants than are observed with dialysate meeting current quality recommendations. Moreover, observational studies suggest a link between clinical outcomes and dialysate purity. Treatment of patients with ultrapure dialysate is reported to improve nutritional status, increase responsiveness to erythropoietin, slow the decline in residual renal function, lessen cardiovascular morbidity, and decrease the incidence of beta(2)-microglobulin amyloidosis. To date, however, none of these studies has shown a cause-and-effect relationship between dialysate purity and outcome. Further, there are no data defining the concentration dependence of outcomes on dialysate purity and the relative importance of dialysate purity as a trigger of inflammation remains unclear. While the technology exists to routinely provide ultrapure dialysate, controlled clinical trials are still needed to answer the question of whether or not introducing ultrapure dialysate into routine clinical practice represents an efficient use of limited resources in terms of decreasing inflammation and improving outcomes in hemodialysis patients. PMID- 15660581 TI - Metabolic consequences of peritoneal dialysis. AB - Optimization of the peritoneal dialysis (PD) prescription includes attempts to normalize the patient's blood pressure and extracellular volume. To do so, one must utilize crystalloid or colloid osmotic agents to achieve ultrafiltration. These osmotic agents are systemically absorbed and thus have both potential benefits and adverse effects. With glucose-based dialysate solutions, the average patient absorbs 300-450 kcal of glucose per day on either continuous ambulatory peritoneal dialysis (CAPD) or the cycler. The amount of glucose absorbed varies based on peritoneal transport characteristics, prescription, and tonicity of fluids used. Alternative osmotic agents such as amino acids and macromolecular solutions, including polypeptides and polyglucose (icodextrin) solutions, have a different rate of systemic absorption and thus a different caloric load profile. In addition, there are protein losses that average about 10 g/day with glucose based solutions and glucose losses with either amino acid or icodextrin dialysate solutions. There are also potential advantages of these alternative solutions with regard to ultrafiltration. Glucose-based solutions require the development of significant crystalloid osmotic forces, which are dissipated as glucose is absorbed systemically. In contrast, macromolecular solutions achieve ultrafiltration via differences in colloid osmotic pressure, and the absorption of these agents is of a lesser magnitude than glucose-based solutions. Further research is needed to determine other potential risks and benefits of these alternative dialysate solutions. PMID- 15660582 TI - Novel approaches in an integrated therapy of inflammatory-associated wasting in end-stage renal disease. AB - It is increasingly apparent that end-stage renal disease (ESRD) patients carry an inflammatory burden, which may play a pivotal role in the evolution of not only wasting, but also the massive increase in the relative risk of cardiovascular disease (CVD). Thus wasting is strongly associated with a persistent systemic inflammatory response, CVD, and impaired patient survival in end-stage renal disease (ESRD), as well as in other chronic diseases. Evidence suggests that a facilitative interaction between inflammatory cytokines and other factors such as poor appetite, comorbidity, acidosis, anemia, and hormonal derangements may cause wasting in this patient group. Clearly, isolated interventions in the form of nutritional energy and protein supplementation have seldom proven to be very effective in improving nutritional status and outcome in ESRD patients, presumably because of the need to attack other causative factors. Therefore, new treatment strategies must be evaluated. Strategies such as multiple appetite stimulants, various "anti-inflammatory diets," and new potentially useful anti inflammatory pharmacologic agents may be tested alone, or in combination, to evaluate if these new therapeutic modalities can improve the outcome of ESRD patients. As the etiology of wasting in ESRD is multifactorial, we propose that its treatment must include not one, but a number of concomitant measures to provide an integrated therapy against this devastating complication. PMID- 15660583 TI - An interesting case of early arteriovenous fistula failure: how I would code it. AB - Treatment of early fistula failure can present a challenge as it relates to the evaluation and treatments required for successful management. It can also present a challenge related to accurate and correct coding of the procedures performed. A clinical case is presented in order to discuss the coding of a case with early fistula failure. The procedure is analyzed in detail in order to determine the appropriate codes to apply to the case. The coding is done in accordance with the principles adopted by the American Society of Diagnostic and Interventional Nephrology (ASDIN). PMID- 15660584 TI - Cutting balloon angioplasty for resistant venous anastomotic stenoses. AB - Conventional angioplasty of stenoses at the venous anastomosis has been demonstrated to be an important endovascular adjunct to mechanical thrombectomy of clotted arteriovenous dialysis grafts. In some cases, however, severe venous anastomosis stenosis is resistant to angioplasty. Cutting balloon angioplasty may have an advantageous role in these difficult situations in order to avoid surgical revision. In this series of 350 patients receiving percutaneous, endovascular declotting procedures, 9 patients had high-grade venous anastomotic stenoses (opening less than 2 mm) that could not be remedied with either conventional or high-pressure noncompliant peripheral angioplasty balloons. These nine patients had the lesions opened with the use of 4 mm x 10 mm cutting balloons and placement of self-expanding nitinol stents at the venous anastomosis during the same angiography procedure. Patients were followed for patency and functionality of the graft. In all cases, immediate technical success occurred. Among these patients, the vessels have remained patent for as long as 20 months of follow-up and grafts have remained functional. Cutting balloon angioplasty may have a potential therapeutic role in resistant venous anastomotic stenoses. PMID- 15660585 TI - Hemodialysis vascular access: percutaneous interventions by nephrologists. AB - Traditionally hemodialysis vascular access-related procedures have been almost exclusively performed by surgeons and interventional radiologists. In recent years, nephrologists have taken the initiative of performing these procedures themselves. Because of their unique clinical perspective on dialysis access and better understanding of the intricacies of renal replacement therapy, nephrologists are ideally suited for this activity. This approach has minimized delays, decreased hospitalizations, and decreased the use of temporary catheters, thereby improving medical care, decreasing costs, and increasing patient convenience. Vascular access-related procedures commonly performed by nephrologists include percutaneous balloon angioplasty, thrombectomy, and tunneled hemodialysis catheter-related procedures. In addition, using vein obliteration and percutaneous balloon angioplasty techniques, nephrologists have recently documented successful salvage of arteriovenous fistulas that had failed to mature, whereas traditionally these fistulas have frequently been abandoned. While the performance of these procedures by nephrologists offers many advantages, appropriate training in order to develop the necessary procedural skills is critical. Recent data have emphasized that a nephrologist can be successfully trained to become a competent interventionalist. In addition to documenting excellent outcome data, multiple reports have demonstrated the safety and success of interventional nephrology. This review focuses on hemodialysis access-related procedures performed by nephrologists and calls for a proactive approach in optimizing this aspect of patient care. PMID- 15660587 TI - Mental health nursing: a changing landscape. PMID- 15660588 TI - Standardized routine outcome measurement: pot holes in the road to recovery. AB - Routine 'outcome measurement' is currently being introduced across Australian mental health services. This paper asserts that routine standardized outcome measurement in its current form can only provide a crude and narrow lens through which to witness recovery. It has only a limited capacity to capture the richness of people's recovery journeys or provide information that can usefully inform care. Indeed, in its implementation nurses may be required to collude in practices or account for practice in ways which run counter to the personal recovery paradigm. Nurses should view a focus on outcomes as an opportunity for critical reflection as well as to seek ways to account for recovery stories in meaningful ways. PMID- 15660589 TI - Linguistic sensitivity, indigenous peoples and the mental health system in Wales. AB - This paper presents findings from a pilot research project to explore the significance and availability of mental health services in the medium of Welsh in Wales, UK. Based on small-scale research with Welsh-speaking mental health service users this article argues that being bilingual can be a significant factor in the complex biopsychosocial matrix that underpins mental health problems amongst Welsh speakers. It also argues that the destructive effects of linguistic oppression, and the difficulties of second language communication for mental health service users, are such that an appropriate health and social care response in Wales involves providing services in a user's preferred language. Service users' views about the current state of bilingual service provision in Wales are presented, which suggests that insufficient attention is being paid to the linguistic needs of Welsh speakers. Eight principles are proposed for mental health service policy and practice in Wales. PMID- 15660590 TI - Effective collaboration enhances rural preceptorship training. AB - Preceptorship is a valuable strategy largely underestimated in its ability to influence nursing students' attitudes and beliefs about mental health nursing. As a model, it has the potential to influence nursing practice, enhance clinical learning, promote recruitment and retention, and generate a more collaborative approach for nursing student supervision. The relationship is usually for a fixed and limited timeframe where the preceptor inspires and supports the growth and development of the student nurse, and encourages role socialization into the profession (Morton-Cooper & Palmer 2000). The challenge for mental health services is to achieve success in the provision of effective preceptorship, ensure positive and rewarding clinical experiences for nursing students, and improve recruitment and retention rates for the service. These aims are substantially more difficult to achieve in the absence of quality education, training and support for preceptors. This paper describes the use of preceptorship training to address recruitment issues in a rural service, particularly the need for effective preceptorship at undergraduate level, which has been shown to have an impact on the willingness of new nursing graduates to enter the mental health nursing field. The design and delivery of the training programme is outlined, including details of the program evaluation and its subsequent impact on psychiatric nurses' approach to and practice of preceptorship. Finally, the broader issue of the value of training nurses in larger groups, enlisting a critical mass of preceptors within an individual mental health service, is discussed. PMID- 15660591 TI - Clients and facilitators' experiences of participating in a Hong Kong self-help group for people recovering from mental illness. AB - The purpose of this study was to examine the effectiveness of self-help groups in the rehabilitation of people recovering from mental illness. A qualitative approach was adopted, with individual interviews of 12 clients and four focus group interviews of 22 professionals and volunteers, in a Hong Kong self-help organization. Three major categories emerged from the client interviews: the meaning of self-help; experience of self-help group involvement; and changes in life. The major categories which emerged from the focus groups were: the meaning of self-help; therapeutic factors; therapeutic process; facilitators' expectation; and difficulties encountered. Results indicated that self-help group involvement provided positive experiences for the members and led to some changes in their lives which contribute to the rehabilitation of their illness. Meanwhile, the professionals' and volunteers' view of the use of self-help groups was found to coincide with previous literature. Moreover, a spiritual dimension was expressed by all respondents as one of the factors that enhanced the group cohesiveness. It was concluded that self-help group involvement is beneficial to psychiatric rehabilitation. Findings have implications for further utilization of self-help groups for the people recovering from mental illness, as an alternative form of health care to complement the inadequacies in the present health care system. PMID- 15660592 TI - Wanting to be heard: mental health consumers' experiences of information about medication. AB - The complexities accompanying the prescription of antipsychotic medication for people experiencing a mental illness have been extensively documented in the literature. The views and experiences of consumers of mental health services, however, are almost entirely absent. This paper describes the findings of a qualitative study undertaken to examine the experiences of consumers, specifically in relation to education and decision making with regards to medication. The findings from a focus group conducted with consumers (n = 9) revealed an overall dissatisfaction with information provided and the opportunity to participate in decision making. Data analysis revealed four major themes: information to consumers; acknowledgement and recognition of consumers; roles of health professionals; and the experience of wellness and adherence. The findings suggest the need for significant change if the goals of the Third National Mental Health Plan are to be realized. PMID- 15660593 TI - Mental health services responding to men and their anger. AB - Due to their propensity for acting out with antisocial behaviours, angry men are often regarded as having antisocial personality disorder with little hope for treatment success. Whilst not denying the difficulties in working with angry men, this paper looks to challenge some of the received wisdom in this area. Antisocial personality disorder is compared with borderline personality disorder and the many similarities, including the association with childhood trauma, course of illness, symptoms and rates of occurrence, are discussed. Differences between the two groups may be associated with the internalization or externalization of anger. Some issues related to bias in diagnosis are raised, as is the universal hopelessness implied in the literature related to treatment of antisocial personality disorder. Some issues related to engaging traditional men are discussed. In this paper the term 'traditional men' is used to describe men who value stoicism, self-reliance, strength, work, status and aggression highly whilst denying any vulnerability and exhibiting restricted emotionality. There is a need to reframe the diagnosis in order to recognize the associated behaviours as being an adult manifestation of complex childhood trauma. Further, there is a need for treatment directed towards altering ongoing patterns of retraumatization which characterize the lives of these men. A case study is included to demonstrate a number of the issues involved in working with angry men. These include issues of connection, boundaries, safety and utilization of the general practitioner in setting up a containing structure. The case study is used to illustrate that when issues of connection and anger containment are positively addressed, good therapeutic outcomes are possible. PMID- 15660594 TI - Relaxation training methods for nurse managers in Hong Kong: a controlled study. AB - Nurse managers are under increased stress because of excessive workloads and hospitals' restructuring which is affecting their work tasks. High levels of stress could affect their mental health. Yet, few stress management training programmes are provided for this population. The purpose of this study was to apply stretch-release relaxation and cognitive relaxation training to enhance the mental health for nurse managers. A total of 65 nurse managers in Hong Kong were randomly assigned to stretch-release relaxation (n = 17), cognitive relaxation (n = 18), and a test control group (n = 35). Mental health status was assessed using the Chinese version of State-Trait Anxiety Inventory and the Chinese version of the General Health Questionnaire. Participants were assessed at the pretreatment session, the fourth posttreatment session, and at the 1-month follow-up session. The results revealed both the stretch-release and cognitive relaxation training enhanced mental health in nurse managers in Hong Kong. The application of relaxation training in enhancing mental health status for nurses and health professionals is discussed. PMID- 15660595 TI - Research involving mental health consumers and carers: a reference group approach. AB - Policy changes within the mental health system are creating an increasing expectation that service providers increase the opportunities for consumer and carer participation in the planning, delivery and evaluation of mental health services. If they are to reflect this change in philosophy, researchers have an obligation to ensure the involvement of consumers and carers, above and beyond involvement as a participant in all relevant mental health research. This paper describes the establishment and function of a reference group, established to guide and assist with the conduct of a research project examining the experiences of consumers and carers with psychiatric disability support services. The formation and operation of, and the valuable contribution made by, the reference group is discussed. PMID- 15660596 TI - Issues and dynamics of sexually assaulted adolescents and their families. AB - Interpersonal violence such as sexual assault creates a variety of traumatic responses. Adolescents encounter a significantly high rate of exposure to sexual assault. In the aftermath of sexual assault, issues and dynamics related to traumatic responses include ongoing fear and threats to personal safety, stability, and structure of the family and environment. Each issue is of concern for community and health care practitioners. Sexual assault has a detrimental effect on adolescent intrapsychic development and interpersonal relationships. Symptoms are disturbing and disruptive to daily routines, negatively affect adolescent normal growth and development, and can result in post-traumatic stress disorder. Issues and dynamics regarding sexual assault are explored, with suggestions on how to help adolescents avoid developing a negative world view and long-term negative health consequences. PMID- 15660597 TI - Managing increased demand for mental health services in a public hospital emergency department: a trial of 'Hospital-in-the-Home' for mental health consumers. AB - Increasing demand from mental health consumers for crisis assessment and intervention in public Emergency Departments (ED) has placed considerable strain on the resources of the ED and long delays awaiting admission are experienced. At Flinders Medical Centre in South Australia, the Psychiatry Department trialled a 'hospital-in-the-home' service to relieve the pressure on the ED and enhance inpatient capacity. The trial has been successful in diverting mental health consumers directly to intensive home-based services from the ED and freeing up beds in the inpatient unit. Evaluation showed that both consumers and their carers were highly satisfied with the hospital-at-home service. PMID- 15660598 TI - Neurocognitive functioning and quality of life among dually diagnosed and non substance abusing schizophrenia inpatients. AB - The aim of this study was to compare the domains of intellectual, memory and executive functions of persons with schizophrenia who concurrently have substance abuse disorders (the dually diagnosed) with a group of non-substance-abusing patients with schizophrenia and to ascertain if there were differences between the two groups in their perceptions of quality of life. Neuropsychological and quality of life data of 46 dually diagnosed and 43 non-substance-abusing patients with schizophrenia was analysed retrospectively. All subjects were inpatients of a state psychiatric hospital. Selected subtests of the Wechsler Adult Intelligence Scale-III and the Wechsler Memory Scale-III constituted the intellectual and memory measures whilst the measures of executive functioning were the Stroop Color Word Test, the FAS version of the Controlled Oral Word Association Test, and the Trail Making Test (Trails A & B). Perceptions of quality of life were evaluated using the World Health Organization Quality of Life measure. The two groups did not differentiate on intellectual and memory domains, however, the dually diagnosed showed a significantly better facility with tasks of executive functions. In addition, the dually diagnosed expressed higher levels of satisfaction with their quality of life compared to the non substance-abusing patients with schizophrenia. These results have implications for interventions. PMID- 15660599 TI - Comment on "Probabilistic risk analysis for a high-level radioactive waste repository" by B. L. Cohen in Risk Analysis, volume 23, 909-915. PMID- 15660601 TI - On the limitations of redundancies in the improvement of system reliability. AB - Some program managers share a common belief that adding a redundant component to a system reduces the probability of failure by half. This is true only if the failures of the redundant components are independent events, which is rarely the case. For example, the redundant components may be subjected to the same external loads. There is, however, in general a decrease in the failure probability of the system. Nonetheless, the redundant element comes at a cost, even if it is less than that of developing the first one when both are based on the same design. Identical parts save the most in terms of design costs, but are subjected to common failure modes from possible design errors that limit the effectiveness of the redundancy. In the development of critical systems, managers thus need to decide if the costs of a parallel system are justified by the increase in the system's reliability. NASA, for example, has used redundant spacecraft to increase the chances of mission success, which worked well in the cases of the Viking and Voyager missions. These two successes, however, do not guarantee future ones. We present here a risk analysis framework accounting for dependencies to support the decision to launch at the same time a twin mission of identical spacecraft, given incremental costs and risk-reduction benefits of the second one. We illustrate this analytical approach with the case of the Mars Exploration Rovers launched by NASA in 2003, for which we had performed this assessment in 2001. PMID- 15660602 TI - Modeling the demand reduction input-output (I-O) inoperability due to terrorism of interconnected infrastructures. AB - Interdependency analysis in the context of this article is a process of assessing and managing risks inherent in a system of interconnected entities (e.g., infrastructures or industry sectors). Invoking the principles of input-output (I O) and decomposition analysis, the article offers a framework for describing how terrorism-induced perturbations can propagate due to interconnectedness. Data published by the Bureau of Economic Analysis Division of the U.S. Department of Commerce is utilized to present applications to serve as test beds for the proposed framework. Specifically, a case study estimating the economic impact of airline demand perturbations to national-level U.S. sectors is made possible using I-O matrices. A ranking of the affected sectors according to their vulnerability to perturbations originating from a primary sector (e.g., air transportation) can serve as important input to risk management. For example, limited resources can be prioritized for the "top-n" sectors that are perceived to suffer the greatest economic losses due to terrorism. In addition, regional decomposition via location quotients enables the analysis of local-level terrorism events. The Regional I-O Multiplier System II (RIMS II) Division of the U.S. Department of Commerce is the agency responsible for releasing the regional multipliers for various geographical resolutions (economic areas, states, and counties). A regional-level case study demonstrates a process of estimating the economic impact of transportation-related scenarios on industry sectors within Economic Area 010 (the New York metropolitan region and vicinities). PMID- 15660603 TI - Acceptance sampling based on truncated life tests in the Birnbaum Saunders model. AB - We develop acceptance sampling plans assuming that the life test is truncated at a preassigned time. The lifetimes of the test units are assumed to follow the Birnbaum Saunders distribution. The minimum sample size necessary to ensure the specified average life is obtained and the operating characteristic values of the sampling plans and producer's risk are presented. An illustrative example is given. PMID- 15660604 TI - Application of human reliability analysis to nursing errors in hospitals. AB - Adverse events in hospitals, such as in surgery, anesthesia, radiology, intensive care, internal medicine, and pharmacy, are of worldwide concern and it is important, therefore, to learn from such incidents. There are currently no appropriate tools based on state-of-the art models available for the analysis of large bodies of medical incident reports. In this study, a new model was developed to facilitate medical error analysis in combination with quantitative risk assessment. This model enables detection of the organizational factors that underlie medical errors, and the expedition of decision making in terms of necessary action. Furthermore, it determines medical tasks as module practices and uses a unique coding system to describe incidents. This coding system has seven vectors for error classification: patient category, working shift, module practice, linkage chain (error type, direct threat, and indirect threat), medication, severity, and potential hazard. Such mathematical formulation permitted us to derive two parameters: error rates for module practices and weights for the aforementioned seven elements. The error rate of each module practice was calculated by dividing the annual number of incident reports of each module practice by the annual number of the corresponding module practice. The weight of a given element was calculated by the summation of incident report error rates for an element of interest. This model was applied specifically to nursing practices in six hospitals over a year; 5,339 incident reports with a total of 63,294,144 module practices conducted were analyzed. Quality assurance (QA) of our model was introduced by checking the records of quantities of practices and reproducibility of analysis of medical incident reports. For both items, QA guaranteed legitimacy of our model. Error rates for all module practices were approximately of the order 10(-4) in all hospitals. Three major organizational factors were found to underlie medical errors: "violation of rules" with a weight of 826 x 10(-4), "failure of labor management" with a weight of 661 x 10(-4), and "defects in the standardization of nursing practices" with a weight of 495 x 10(-4). PMID- 15660605 TI - Trust, the asymmetry principle, and the role of prior beliefs. AB - Within the risk literature there is an ongoing debate on whether trust is vulnerable or enduring. Previous research on nuclear energy by Slovic in 1993 has shown that negative events have much greater impact on self-reported trust than do positive events. Slovic attributes this to the asymmetry principle: specifically, that trust is much easier to destroy than to create. In a questionnaire survey concerning genetically modified (GM) food in Britain (n= 396) we similarly find that negative events have a greater impact on trust than positive events. Because public opinion in Britain is skewed in the direction of opposition toward GM food, the pattern of results could either be caused by the fact that negative information is more informative than positive information (a negativity bias) or reflect the influence of people's prior attitudes toward the issue (a confirmatory bias). The results were largely in line with the confirmatory bias hypothesis: participants with clear positive or negative beliefs interpreted events in line with their existing attitude position. However, for participants with intermediate attitudes, negative items still had greater impact than the positive. This latter finding suggests that, congruent with the negativity bias hypothesis, negative information may still be more informative than positive information for undecided people. The study also identified the labeling of GM products, consulting the public, making biotechnology companies liable for any damage, and making a test available to detect GM produce as being particularly important preconditions for maintaining trust in the regulation of agricultural biotechnology. PMID- 15660606 TI - Managing potential health risks from electric powerlines: a decision analysis caught in controversy. AB - Over the past 20 years, several epidemiological studies have found an association between exposure to electromagnetic fields (EMFs) and health effects, including childhood leukemia and adult brain cancer. However, experts strongly disagree about whether this association is causal and, if so, how strong it is. In this article, we examine several alternatives to reduce EMFs from sources of the California power grid, including undergrounding distribution and transmission lines and reconfiguring or rephasing lines. The alternatives were evaluated in terms of the potential health risk reduction, cost, impacts on service reliability, property values, and many other consequences. Because of the uncertainty about an EMF-health link, the main effort was to determine the sensitivity of the decisions to the probability and seriousness of an EMF hazard. User-friendly computer models were developed to allow stakeholders to change the model assumptions and parameters to analyze the impacts of their own assumptions and estimates on the decision. The analysis clearly demonstrated that only four of the many concerns raised by the stakeholders could make a difference in the decision: health risks, costs, service reliability, and property values. Whether undergrounding, moderate alternatives for EMF reduction, or no change was the best decision depended on a few key factors, including the probability that EMF exposure is a hazard, the severity of this hazard, how the EMF reduction measures are financed, and the impacts on property values. While the analysis did not resolve the EMF issues, it showed that even in the most controversial settings, a little analysis goes a long way to clarifying the issues and to focus the debate. PMID- 15660607 TI - Toward improved communication about wildland fire: mental models research to identify information needs for natural resource management. AB - The management of natural- and human-induced wildland fires is an intricate process that must balance two considerations: that of fire as a necessary natural disturbance and that of the risks that fire poses. Reconciling tradeoffs between these risks and benefits is contingent upon informed, directed, and two-way communication between wildland fire managers and stakeholders. In an effort to aid with the design of such a communication effort, this study used mental models research to determine the unique wildland fire information and decision-making needs of stakeholders living at the wildland-urban interface of a fire-prone area. While the analysis revealed many similarities in how stakeholders conceptualized the risks and benefits of wildland fire, many misconceptions and important gaps in understanding on the part of both expert and nonexpert respondents were identified. PMID- 15660608 TI - Elicitation of expert judgments of uncertainty in the risk assessment of herbicide-tolerant oilseed crops. AB - One of the lay public's concerns about genetically modified (GM) organisms (GMO) and related emerging technologies is that not all the important risks are evaluated or even identified yet--and that ignorance of the unanticipated risks could lead to severe environmental or public health consequences. To some degree, even the scientists who participated in the analysis of the risks from GMOs (arguably the people most qualified to critique these analyses) share some of this concern. To formally explore the uncertainty in the risk assessment of a GM crop, we conducted detailed interviews of seven leading experts on GM oilseed crops to obtain qualitative and quantitative information on their understanding of the uncertainties associated with the risks to agriculture from GM oilseed crops (canola or rapeseed). The results of these elicitations revealed three issues of potential concern that are currently left outside the scope of risk assessments. These are (1) the potential loss of the agronomic and environmental benefits of glyphosate (a herbicide widely used in no-till agriculture) due to the combined problems of glyphosate-tolerant canola and wheat volunteer plants, (2) the growing problem of seed lot contamination, and (3) the potential market impacts. The elicitations also identified two areas where knowledge is insufficient. These are: the occurrence of hybridization between canola and wild relatives and the ability of the hybrids to perpetuate themselves in nature, and the fate of the herbicide-tolerance genes in soil and their interaction with soil microfauna and -flora. The methodological contribution of this work is a formal approach to analyzing the uncertainty surrounding complex problems. PMID- 15660609 TI - Site-specific updating and aggregation of Bayesian belief network models for multiple experts. AB - A method for combining multiple expert opinions that are encoded in a Bayesian Belief Network (BBN) model is presented and applied to a problem involving the cleanup of hazardous chemicals at a site with contaminated groundwater. The method uses Bayes Rule to update each expert model with the observed evidence, then uses it again to compute posterior probability weights for each model. The weights reflect the consistency of each model with the observed evidence, allowing the aggregate model to be tailored to the particular conditions observed in the site-specific application of the risk model. The Bayesian update is easy to implement, since the likelihood for the set of evidence (observations for selected nodes of the BBN model) is readily computed by sequential execution of the BBN model. The method is demonstrated using a simple pedagogical example and subsequently applied to a groundwater contamination problem using an expert knowledge BBN model. The BBN model in this application predicts the probability that reductive dechlorination of the contaminant trichlorethene (TCE) is occurring at a site--a critical step in the demonstration of the feasibility of monitored natural attenuation for site cleanup--given information on 14 measurable antecedent and descendant conditions. The predictions for the BBN models for 21 experts are weighted and aggregated using examples of hypothetical and actual site data. The method allows more weight for those expert models that are more reflective of the site conditions, and is shown to yield an aggregate prediction that differs from that of simple model averaging in a potentially significant manner. PMID- 15660610 TI - The role of risk and future land use in cleanup decisions at the Department Of Energy. AB - As a result of the legacy of the Cold War, the Departments of Energy and Defense are involved in massive cleanup and remediation projects. While health risk to humans and ecological receptors is perceived to be the basis for remediation, this assumption is rarely examined. In this article, we examine the role of risk and future land-use designations in cleanup decisions, using the Department of Energy's self-assessment of 36 sites. We then discuss the risk-related tools that might be required to address the cleanup challenge. Much of the current cleanup program is driven by compliance with federal and state statutes and regulations, presumably to protect human health and the environment. Compliance, however, is not synonymous with cleanup. Although some of these laws and regulations take risk into account, the lack of site-specific data on exposures and risk scenarios, and the lack of attention to future land use or end states, has often resulted in disconnects between risk and cleanup goals, risk and final end states, and cleanup levels and end state or subsequent land use. Partly, these disconnects result from the need for a range of technical, economic, sociological, and public policy tools to address the issues. A better transfer of information among and within Department of Energy facilities, operations offices, and DOE headquarters is required. Further, linking cleanup decisions and goals with the final end state involves a number of risk tradeoffs, including (1) ecological versus human health, (2) worker versus public health, (3) among competing contaminated areas, (4) among temporal patterns of cleanup, (5) among different ecological receptors (plants vs. animals, one animal vs. another), and (6) among the sites across the DOE complex. For the nation, balancing among risks is essential within sites and among Department of Energy sites, as well as among other remediation sites (such as those of Department of Defense and Superfund sites). PMID- 15660611 TI - Characterizing environmental harm: developments in an approach to strategic risk assessment and risk management. AB - Environmental policymakers and regulators are often in the position of having to prioritize their actions across a diverse range of environmental pressures to secure environmental protection and improvements. Information on environmental issues to inform this type of strategic analysis can be disparate; it may be too voluminous or even absent. Data on a range of issues are rarely presented in a common format that allows easy analysis and comparison. Nevertheless, judgments are required on the significance of various environmental pressures and on the inherent uncertainties to inform strategic assessments such as "state of the environment" reports. How can decisionmakers go about this type of strategic and comparative risk analysis? In an attempt to provide practical tools for the analysis of environmental risks at a strategic level, the Environment Agency of England and Wales has conducted a program of developmental research on strategic risk assessment since 1996. The tools developed under this program use the concept of "environmental harm" as a common metric, viewed from technical, social, and economic perspectives, to analyze impacts from a range of environmental pressures. Critical to an informed debate on the relative importance of these perspectives is an understanding and analysis of the various characteristics of harm (spatial and temporal extent, reversibility, latency, etc.) and of the social response to actual or potential environmental harm from a range of hazards. Recent developments in our approach, described herein, allow a presentation of the analysis in a structured fashion so as to better inform risk management decisions. PMID- 15660612 TI - Examining localized patterns of air quality perception in Texas: a spatial and statistical analysis. AB - Environmental and human health issues associated with outdoor air pollution, such as ozone, sulfur dioxide, and other pollutants in metropolitan regions, are an area of growing concern for both policy officials and the general public. Increasing attention from the news media, new health data, and public debate over the effectiveness of clean air regulations have raised the importance of air quality in the public consciousness. While public perceptions of air quality have been studied thoroughly dating back to the 1960s, little empirical research has been conducted to explain the spatial aspects of these perceptions, particularly at the local level. Although recent studies suggest characteristics of local setting are important in shaping perceptions of air quality, the roles of proximity, neighborhood characteristics, and location have not been clarified. This study seeks to improve understanding of the major factors shaping public perceptions of air quality by examining the spatial pattern of local risk perception, the role of socioeconomic characteristics in forming these perceptions, and the relationship between perceived and scientifically measured air pollution. First, we map the spatial pattern of local air quality perceptions using Geographic Information Systems (GIS) across the Dallas and Houston metropolitan areas. Next, we explain these perceptions through local contextual factors using both bivariate correlations and multivariate regression analysis. Results indicate that perceptions of air quality in the study areas are not significantly correlated with air quality based on readings of air monitoring stations. Instead, perceptions appear to be influenced by setting (urban vs. rural), state identification, access to information, and socioeconomic characteristics such as age, race, and political identification. We discuss the implications of the findings and provide direction on how further research can provide a deeper understanding of the local contextual factors influencing public perceptions. PMID- 15660613 TI - Major psychological factors affecting acceptance of gene-recombination technology. AB - The purpose of this study was to verify the validity of a causal model that was made to predict the acceptance of gene-recombination technology. A structural equation model was used as a causal model. First of all, based on preceding studies, the factors of perceived risk, perceived benefit, and trust were set up as important psychological factors determining acceptance of gene-recombination technology in the structural equation model. An additional factor, "sense of bioethics," which I consider to be important for acceptance of biotechnology, was added to the model. Based on previous studies, trust was set up to have an indirect influence on the acceptance of gene-recombination technology through perceived risk and perceived benefit in the model. Participants were 231 undergraduate students in Japan who answered a questionnaire with a 5-point bipolar scale. The results indicated that the proposed model fits the data well, and showed that acceptance of gene-recombination technology is explained largely by four factors, that is, perceived risk, perceived benefit, trust, and sense of bioethics, whether the technology is applied to plants, animals, or human beings. However, the relative importance of the four factors was found to vary depending on whether the gene-recombination technology was applied to plants, animals, or human beings. Specifically, the factor of sense of bioethics is the most important factor in acceptance of plant gene-recombination technology and animal gene-recombination technology, and the factors of trust and perceived risk are the most important factors in acceptance of human being gene-recombination technology. PMID- 15660614 TI - Aviation risk perception: a comparison between experts and novices. AB - This article describes an exploratory investigation of the risk perceptions of experts and novices in relation to helicopter operations, under conditions where the participants are matched on various characteristics previously found to affect perceptions, such as demographic, gender, and background factors. The study reports considerable evidence of perceptual differences between the two participant groups (i.e., expert pilots and candidate pilots). We find that the experts' perceptions of relative risks are more veridical, in terms of their higher correlation with the true relative frequencies. A significant positive correlation between the flight hours and the contextual risk-taking tendency is also shown, leading the experienced pilots' choices toward risky alternatives in scenarios--a potential result of their overconfidence based on superior task performance. Possible explanations are offered for the findings and potential avenues for future research are identified. PMID- 15660615 TI - Direct data manipulation for local decision analysis as applied to the problem of arsenic in drinking water from tube wells in Bangladesh. AB - A wide variety of tools are available, both parametric and nonparametric, for analyzing spatial data. However, it is not always clear how to translate statistical inferences into decision recommendations. This article explores the possibilities of estimating the effects of decision options using very direct manipulation of data, bypassing formal statistical analysis. We illustrate with the application that motivated this research, a study of arsenic in drinking water in nearly 5,000 wells in a small area in rural Bangladesh. We estimate the potential benefits of two possible remedial actions: (1) recommendations that people switch to nearby wells with lower arsenic levels; and (2) drilling new community wells. We use simple nonparametric clustering methods and estimate uncertainties using cross-validation. PMID- 15660616 TI - Integrated analysis: combining risk and economic assessments while preserving the separation of powers. AB - This article presents a process for an integrated policy analysis that combines risk assessment and benefit-cost analysis. This concept, which explicitly combines the two types of related analyses, seems to contradict the long-accepted risk analysis paradigm of separating risk assessment and risk management since benefit-cost analysis is generally considered to be a part of risk management. Yet that separation has become a problem because benefit-cost analysis uses risk assessment results as a starting point and considerable debate over the last several years focused on the incompatibility of the use of upper bounds or "safe" point estimates in many risk assessments with benefit-cost analysis. The problem with these risk assessments is that they ignore probabilistic information. As advanced probabilistic techniques for risk assessment emerge and economic analysts receive distributions of risks instead of point estimates, the artificial separation between risk analysts and the economic/decision analysts complicates the overall analysis. In addition, recent developments in countervailing risk theory suggest that combining the risk and benefit-cost analyses is required to fully understand the complexity of choices and tradeoffs faced by the decisionmaker. This article also argues that the separation of analysis and management is important, but that benefit-cost analysis has been wrongly classified into the risk management category and that the analytical effort associated with understanding the economic impacts of risk reduction actions need to be part of a broader risk assessment process. PMID- 15660617 TI - Tiered chemical testing: a value of information approach. AB - In December 2000 the EPA initiated the Voluntary Children's Chemical Evaluation Program (VCCEP) by asking manufacturers to voluntarily sponsor toxicological testing in a tiered process for 23 chemicals selected for the pilot phase. The tiered nature of the VCCEP pilot program creates the need for clearly defined criteria for determining when information is sufficient to assess the potential risks to children. This raises questions about how to determine the "adequacy" of the existing information and assess the need to undertake efforts to reduce uncertainty (through further testing). This article applies a value of information analysis approach to determine adequacy by modeling how toxicological and exposure data collected through the VCCEP may be used to inform risk management decisions. The analysis demonstrates the importance of information about the exposure level and control costs in making decisions regarding further toxicological testing. This article accounts for the cost of delaying control action and identifies the optimal testing strategy for a constrained decisionmaker who, absent applicable human data, cannot regulate without bioassay data on a specific chemical. It also quantifies the differences in optimal testing strategy for three decision criteria: maximizing societal net benefits, ensuring maximum exposure control while net benefits are positive (i.e., benefits outweigh costs), and controlling to the maximum extent technologically feasible while the lifetime risk of cancer exceeds a specific level of risk. Finally, this article shows the large differences that exist in net benefits between the three criteria for the range of exposure levels where the optimal actions differ. PMID- 15660618 TI - Promoting transparency of long-term environmental decisions: the Hanford Decision Mapping System pilot project. AB - Nuclear waste cleanup is a challenging and complex problem that requires both scientific analysis and dialogue among a variety of stakeholders. This article describes an effort to develop an online information system that supports this analytic-deliberative dialogue by integrating cleanup information for the Hanford Site, and making it more "transparent." A framework for understanding and evaluating transparency guided system development. Working directly with stakeholders, we identified information needs and developed new ways to organize and present the information so that it would be more transparent to interested parties, with the ultimate aim of fostering greater participation in decision dialogues and processes. The complexity of the information needed for dialogue suggested that several types of communication devices ("information structures") were warranted. Five information structures were developed for the pilot Decision Mapping System (http://nalu.geog.washington.edu/dms). Decision maps hyperlinked decision information to maps of Hanford. Background Information provided context in a narrative format. Decision Paths organized decision process information on a timeline and provided direct hyperlinks to online documentation. The Geographic Library hyperlinked decision documents to maps. Finally, a Discussion Forum allowed users to make comments and view remarks from others. Early lessons from this work suggest that transparency is integral to long-term management, a participatory design process contributed greatly to its perceived success, and better data integration to support decision making is needed. This work has broad implications for risk communicators and risk managers because it speaks to the design of information systems to support "analytic-deliberative" decision processes (i.e., those that rely upon both risk science and public dialogue). PMID- 15660619 TI - Developmental disruption of thyroid hormone: correlations with hearing dysfunction in rats. AB - A wide variety of environmental contaminants adversely affect thyroid hormone (TH) homeostasis. Hypothyroidism and/or hypothyroxinemia during the early postnatal period in the rat leads to permanent structural damage and loss of function in the cochlea. A major uncertainty in assessing the risks of developmental exposure to thyroid-disrupting chemicals (TDCs) is the lack of a clear characterization of the dose-response relationship, especially in the lower region, between disruption of hormones and adverse consequences. The current work correlated early postnatal hypothyroxinemia with hearing loss in the adult rat. Linear regression was performed on the log transform for total serum thyroxine (T4) concentrations on postnatal day 14 or 21 versus dB(SPL) of hearing loss in adult animals developmentally exposed to TDCs. Regression analyses revealed a highly significant correlation between T4 concentration and hearing loss. In the rat, a 50-60% decrease in circulating T4 was needed to significantly impact hearing function. This correlation suggests that T4 serum concentrations at 14 or 21 days of postnatal age may be a good predictive biomarker in rodents of the adverse consequence of developmental exposure to TDCs. PMID- 15660620 TI - Quantitative risk assessment for developmental neurotoxic effects. AB - Developmental neurotoxicity concerns the adverse health effects of exogenous agents acting on neurodevelopment. Because human brain development is a delicate process involving many cellular events, the developing fetus is rather susceptible to compounds that can alter the structure and function of the brain. Today, there is clear evidence that early exposure to many neurotoxicants can severely damage the developing nervous system. Although in recent years, there has been much attention given to model development and risk assessment procedures for developmental toxicants, the area of developmental neurotoxicity has been largely ignored. Here, we consider the problem of risk estimation for developmental neurotoxicants from animal bioassay data. Since most responses from developmental neurotoxicity experiments are nonquantal in nature, an adverse health effect will be defined as a response that occurs with very small probability in unexposed animals. Using a two-stage hierarchical normal dose response model, upper confidence limits on the excess risk due to a given level of added exposure are derived. Equivalently, the model is used to obtain lower confidence limits on dose for a small negligible level of risk. Our method is based on the asymptotic distribution of the likelihood ratio statistic (cf. Crump, 1995). An example is used to provide further illustration. PMID- 15660621 TI - Role of the standard deviation in the estimation of benchmark doses with continuous data. AB - For continuous data, risk is defined here as the proportion of animals with values above a large percentile, e.g., the 99th percentile or below the 1st percentile, for the distribution of values among control animals. It is known that reducing the standard deviation of measurements through improved experimental techniques will result in less stringent (higher) doses for the lower confidence limit on the benchmark dose that is estimated to produce a specified risk of animals with abnormal levels for a biological effect. Thus, a somewhat larger (less stringent) lower confidence limit is obtained that may be used as a point of departure for low-dose risk assessment. It is shown in this article that it is important for the benchmark dose to be based primarily on the standard deviation among animals, s(a), apart from the standard deviation of measurement errors, s(m), within animals. If the benchmark dose is incorrectly based on the overall standard deviation among average values for animals, which includes measurement error variation, the benchmark dose will be overestimated and the risk will be underestimated. The bias increases as s(m) increases relative to s(a). The bias is relatively small if s(m) is less than one-third of s(a), a condition achieved in most experimental designs. PMID- 15660622 TI - Effects of exposure imprecision on estimation of the benchmark dose. AB - In regression analysis failure to adjust for imprecision in the exposure variable is likely to lead to underestimation of the exposure effect. However, the consequences of exposure error for determination of safe doses of toxic substances have so far not received much attention. The benchmark approach is one of the most widely used methods for development of exposure limits. An important advantage of this approach is that it can be applied to observational data. However, in this type of data, exposure markers are seldom measured without error. It is shown that, if the exposure error is ignored, then the benchmark approach produces results that are biased toward higher and less protective levels. It is therefore important to take exposure measurement error into account when calculating benchmark doses. Methods that allow this adjustment are described and illustrated in data from an epidemiological study on the health effects of prenatal mercury exposure. PMID- 15660623 TI - Framework for evaluation of physiologically-based pharmacokinetic models for use in safety or risk assessment. AB - Proposed applications of increasingly sophisticated biologically-based computational models, such as physiologically-based pharmacokinetic models, raise the issue of how to evaluate whether the models are adequate for proposed uses, including safety or risk assessment. A six-step process for model evaluation is described. It relies on multidisciplinary expertise to address the biological, toxicological, mathematical, statistical, and risk assessment aspects of the modeling and its application. The first step is to have a clear definition of the purpose(s) of the model in the particular assessment; this provides critical perspectives on all subsequent steps. The second step is to evaluate the biological characterization described by the model structure based on the intended uses of the model and available information on the compound being modeled or related compounds. The next two steps review the mathematical equations used to describe the biology and their implementation in an appropriate computer program. At this point, the values selected for the model parameters (i.e., model calibration) must be evaluated. Thus, the fifth step is a combination of evaluating the model parameterization and calibration against data and evaluating the uncertainty in the model outputs. The final step is to evaluate specialized analyses that were done using the model, such as modeling of population distributions of parameters leading to population estimates for model outcomes or inclusion of early pharmacodynamic events. The process also helps to define the kinds of documentation that would be needed for a model to facilitate its evaluation and implementation. PMID- 15660624 TI - MORM--a Petri net based model for assessing OH&S risks in industrial processes: modeling qualitative aspects. AB - Because of the increase in workplace automation and the diversification of industrial processes, workplaces have become more and more complex. The classical approaches used to address workplace hazard concerns, such as checklists or sequence models, are, therefore, of limited use in such complex systems. Moreover, because of the multifaceted nature of workplaces, the use of single oriented methods, such as AEA (man oriented), FMEA (system oriented), or HAZOP (process oriented), is not satisfactory. The use of a dynamic modeling approach in order to allow multiple-oriented analyses may constitute an alternative to overcome this limitation. The qualitative modeling aspects of the MORM (man machine occupational risk modeling) model are discussed in this article. The model, realized on an object-oriented Petri net tool (CO-OPN), has been developed to simulate and analyze industrial processes in an OH&S perspective. The industrial process is modeled as a set of interconnected subnets (state spaces), which describe its constitutive machines. Process-related factors are introduced, in an explicit way, through machine interconnections and flow properties. While man-machine interactions are modeled as triggering events for the state spaces of the machines, the CREAM cognitive behavior model is used in order to establish the relevant triggering events. In the CO-OPN formalism, the model is expressed as a set of interconnected CO-OPN objects defined over data types expressing the measure attached to the flow of entities transiting through the machines. Constraints on the measures assigned to these entities are used to determine the state changes in each machine. Interconnecting machines implies the composition of such flow and consequently the interconnection of the measure constraints. This is reflected by the construction of constraint enrichment hierarchies, which can be used for simulation and analysis optimization in a clear mathematical framework. The use of Petri nets to perform multiple-oriented analysis opens perspectives in the field of industrial risk management. It may significantly reduce the duration of the assessment process. But, most of all, it opens perspectives in the field of risk comparisons and integrated risk management. Moreover, because of the generic nature of the model and tool used, the same concepts and patterns may be used to model a wide range of systems and application fields. PMID- 15660625 TI - The use of risk assessment to decide the control strategy for bluetongue in Italian ruminant populations. AB - The affiliation, assessment and management of risks is a traditional part of veterinary medicine. In the past, veterinary services involved in this type of activity usually assessed risks qualitatively. However, since the 1990s, quantitative methods have become increasingly important. The establishment of the World Trade Organization in 1994, and the promulgation of its Agreement on the Application of Sanitary and Phytosanitary Measures (the "SPS Agreement") led to an increased application of import risk analysis and to significant improvements in the methodology of risk analysis as applied to international trade policy for animals and animal products. However, there was very little development of risk analysis in veterinary fields other than international trade and management of health risks to consumers of animal products and little has been published on its use in the choice and definition of control or prophylaxis strategies for animal diseases. This article describes a quantitative risk assessment, which was undertaken in Italy to help choose an appropriate national response strategy following an incursion of bluetongue, an infectious disease of sheep and goats. The results obtained in this study support the use of risk analysis as a tool to assist in choosing an appropriate animal disease management strategy. The use of risk analysis in the evaluation of disease management strategies also offers advantages in international trade. It makes easier the comparison of different strategies applied in the various countries, and thus facilitates the assessment of equivalence of the guarantees provided by different strategies. PMID- 15660628 TI - Connections between experience, beliefs, scientific knowledge, and self-evaluated expertise among investigators of child sexual abuse in Finland. AB - The purpose of this study was to investigate whether clinicians investigating child sexual abuse (CSA) rely more on scientific knowledge or on clinical experience when evaluating their own expertise. Another goal was to check what kind of pre-trial beliefs the clinicians had. The connections between these different factors were investigated. A questionnaire covering items concerning demographic data, experience, knowledge about CSA, self-evaluated expertise and beliefs about CSA was given to 126 social workers, 60 child psychiatrists and 134 psychologists. The results showed that the clinicians relied more on their clinical experience than on scientific knowledge when evaluating their expertise as investigators of CSA. Furthermore, social workers possessed stronger attitudes in favor of children than the other groups, while child psychiatrists had more negative attitudes towards the criminal justice system. Male participants had less strong beliefs than did the female participants. The findings indicate that the education of CSA investigators should focus more on theoretical knowledge and decision-making processes as well as the role of pre-trial beliefs. PMID- 15660629 TI - Memory for action events: findings in neurological patients. AB - Encoding action phrases by enacting leads normally to better memory performance than verbal encoding. In order to gain additional insight into the representational basis of the enactment effect, neurological patients are contrasted with healthy participants. Persons suffering from Parkinson's disease, which primarily impairs the motor system, and patients suffering from Frontal Lobe Syndrome, which primarily affects action-related planning processes, were involved. We investigated whether the enactment effect would be differentially affected by these disorders. In addition, the characteristics of information processing after encoding by enacting was analyzed by varying memory material (unrelated versus clusterable actions) and by adding an encoding condition that included obligatory action planning (director condition). The findings indicate that the impact of motor information for the enactment effect is not dominant compared to the role of action-related cognitive and motivational processes, in particular planning processes. The findings of the two experiments are explained within traditional conceptual memory theories. PMID- 15660630 TI - Crossfostering in mice selectively bred for high and low levels of open-field thigmotaxis. AB - The main purpose of this research was to investigate whether the difference in open-field (OF) thigmotaxis between mice selectively bred for high and low levels of wall-seeking behavior originated from genetic or acquired sources. Unfostered, infostered, and crossfostered mice were compared in two experiments in which the effects of strain, sex, and fostering on ambulation, defecation, exploration, grooming, latency to move, radial latency, rearing, thigmotaxis, and urination were studied. These experiments revealed that OF thigmotaxis was unaffected by the foster condition and thus genetically determined. The selected strains of mice also diverged repeatedly with regard to exploration and rearing. The findings are in line with the previously described existence of an inverse relationship between emotionality and exploration. PMID- 15660631 TI - Construct validation of the Swedish Scholastic Aptitude Test by means of the Swedish Enlistment Battery. AB - The construct validity of the Swedish Scholastic Test (SweSAT), used for admissions to higher education, was studied through relating it to the Computerized Enlistment Battery (CAT-SEB) used at conscription to military service. Out of three male age cohorts tested at 18 years with the CAT-SEB in 1997, 1998, or 1999 six groups of men that had taken their first SweSAT in the Autumn (n= 3,163; 2,938; 2,820) the same year, or in the Spring (n= 3,238; 2,587; 2,447) the next year were used for the analyses. Four types of models were fitted to the tests: (1) a nested factor model with the latent variables general intelligence (g), crystallized intelligence (Gc) and general visualization; (2) a higher order model with g as an apex factor; (3) a multivariate regression model where the SeSAT subtests were regressed on the SEB factors; and (4) separate models of SweSAT and CAT-SEB with covariances between latent variables. The results show that the SweSAT is dominated by Gc factors and that the g factor has a smaller influence. PMID- 15660632 TI - Responsibility assignment at the workplace: a Finnish and Ghanaian perspective. AB - The study sought to explore management's responsibility in accident causation as perceived by Ghanaian and Finnish industrial workers. It was anticipated that the Ghanaian industrial workers would externalize accident causality more than their Finnish counterparts, and would assign only marginal responsibility to the management for industrial accidents. The predicted difference was based on the premise that the difference in these two cultures on the power-distance cultural dimensions would have an impact on their responsibility assignments. The results confirmed our expectation. PMID- 15660633 TI - Interviewing children with the cognitive interview: assessing the reliability of statements based on observed and imagined events. AB - This paper investigated whether criteria stemming from the Reality Monitoring (RM) framework could be trusted to assess the reliability of statements obtained by the use of a cognitive interview (CI). Fifty-eight children, aged 10-11, participated. One-third watched a film about a fakir and were then interviewed according to a CI (n= 19). The remaining two-thirds made up a story about a fakir and were then interviewed according to either a CI (n= 21), or a structured interview (SI) (n= 18). The CI statements based on observed events contained more visual, affective, spatial and temporal information compared to CI statements based on imagined events. The CI statements based on imagined events did not differ from the SI statements based on imagined events. Considerable developmental work is recommended to turn the RM technique to a reliable test that could be used by practitioners. PMID- 15660634 TI - What's measured is not necessarily managed: cognitive contingencies of organizational measurement. AB - Organizational measurement is necessary for managing incremental improvements or change efforts. Some have argued that indicators of financial performance measure the past and should be joined by "lead" measures such as management development and customer relations, assumed to be drivers of performance. Approaches of "lead" measures have met criticism for not contributing to operational management. The present is an empirical case study where measures from a customer satisfaction survey and a management assessment instrument were analyzed and compared to actual measures of financial performance. The effects of measures obtained seem to depend on the interpretations and reflection taking place in the organization. The pursuit of "lead" indicators did not seem to drive results. Counter-intuitively, higher scores on these measures were related to poorer financial performance. The explication of and reflection on the underlying dynamic seemed to turn the development in a profitable direction. PMID- 15660635 TI - Extracting a maximum of useful information from statistical research data. AB - The practice of statistical inference in psychological research is critically reviewed. Particular emphasis is put on the fast pace of change from the sole reliance on null hypothesis significance testing (NHST) to the inclusion of effect size estimates, confidence intervals, and an interest in the Bayesian approach. We conclude that these developments are helpful for psychologists seeking to extract a maximum of useful information from statistical research data, and that seven decades of criticism against NHST is finally having an effect. PMID- 15660636 TI - Significance testing, interval estimation or Bayesian inference: comments to "Extracting a maximum of useful information from statistical research data" by S. Sohlberg and G. Andersson. AB - Statistical inference plays an important part in the formation of scientific knowledge in psychology. Starting from a paper by Sohlberg and Andersson (2005; Scandinavian Journal of Psychology, 46, 69-77) these issues are discussed. It is argued that interval estimates are easy to understand and that they are more suitable than significance testing for most problems. Bayesian inference is a coherent description of the information building process. With some examples it is shown that null hypothesis significance testing is full of contradictions. Finally, some other important issues like convenience sampling and model selection are shortly mentioned. PMID- 15660637 TI - The ecological validity of the "who said what?" technique: an examination of the role of self-involvement, cognitive interference and acquaintanceship. AB - Questions are raised about the possible ecological invalidity of the traditional "who said what?" (WSW) procedure. Three studies are reported which use a modified, quasi-naturalistic version of the WSW technique, in which rather than being passive recipients of presented information, participants instead play an active role in the production of such information in an interactive context. Data are presented that demonstrate that the social categorization effect typically obtained through the traditional WSW procedure generalizes beyond the circumstances in which it was initially demonstrated. Thus, in Study 1 it was established that social categories (gender, in this case) are cognitively used in real interactions between participants in order to organize information. Moreover, Study 2 demonstrated that the use of categories is accentuated when, as is typical in real world situations, recollections are not made immediately after the perception of the relevant stimuli but following a period of intervening cognitive activity. The findings of Study 3, however, suggest that the WSW technique may not inevitably lead to social categorization effects: typical effects did not appear when participants knew each other well, and therefore saw one another as unique, idiosyncratic individuals, rather than as category members. PMID- 15660638 TI - Worries in restaurant managers. AB - This paper explores the concept of job specific worries (JS worries) in managers in the restaurant industry. The focus of interest in the present explorative study was the degree and content of restaurant managers' worries. Of a total of 58 eligible managers in a Norwegian (fast food) restaurant chain, 57 (representing an equal number of restaurants) were surveyed for JS worries and personal worries. These managers also judged aspects of their relation to the organization they work for, such as perceived organizational hierarchy, job commitment and perceived risk at work. The results indicate that the concept of JS worry is state dependent as opposed to personal worry, which may be regarded as a trait. In addition, the observed correlations indicate a connection between characteristics of perceived job environment and the respondents' degree of JS worry, whereas no such correlations were observed between trait worry and job environment. PMID- 15660640 TI - The development of arithmetical abilities. AB - BACKGROUND: Arithmetical skills are essential to the effective exercise of citizenship in a numerate society. How these skills are acquired, or fail to be acquired, is of great importance not only to individual children but to the organisation of formal education and its role in society. METHOD: The evidence on the normal and abnormal developmental progression of arithmetical abilities is reviewed; in particular, evidence for arithmetical ability arising from innate specific cognitive skills (innate numerosity) vs. general cognitive abilities (the Piagetian view) is compared. RESULTS: These include evidence from infancy research, neuropsychological studies of developmental dyscalculia, neuroimaging and genetics. The development of arithmetical abilities can be described in terms of the idea of numerosity -- the number of objects in a set. Early arithmetic is usually thought of as the effects on numerosity of operations on sets such as set union. The child's concept of numerosity appears to be innate, as infants, even in the first week of life, seem to discriminate visual arrays on the basis of numerosity. Development can be seen in terms of an increasingly sophisticated understanding of numerosity and its implications, and in increasing skill in manipulating numerosities. The impairment in the capacity to learn arithmetic -- dyscalculia -- can be interpreted in many cases as a deficit in the concept in the child's concept of numerosity. The neuroanatomical bases of arithmetical development and other outstanding issues are discussed. CONCLUSIONS: The evidence broadly supports the idea of an innate specific capacity for acquiring arithmetical skills, but the effects of the content of learning, and the timing of learning in the course of development, requires further investigation. PMID- 15660641 TI - Pathways to conscience: early mother-child mutually responsive orientation and children's moral emotion, conduct, and cognition. AB - BACKGROUND: Associations between early mother-child mutually responsive orientation (MRO) and children's conscience have been previously established, but the mechanisms accounting for those links are not understood. We examined three such mediational mechanisms: (a) the child's enhanced enjoyment of interactions with the mother, (b) increased committed, self-regulated compliance with the mother, and/or (c) a decreased need for maternal use of power assertion. Children's conscience was seen as a complex system encompassing moral emotion (guilt), conduct, and cognition. METHODS: In a longitudinal design, MRO was observed in mothers' and children's multiple naturalistic interactions at 9, 14, and 22 months. The mediators were observed at 33 months. Children's conscience was observed at 45 months (moral emotion) and at 56 months (moral conduct and cognition). RESULTS: The mediating paths were different for the three components of conscience. MRO had a direct, unmediated effect on moral emotion. MRO influenced moral conduct through two mediational paths: by promoting the child's enjoyment of interactions with the mother and by enhancing committed compliance. MRO influenced moral cognition by promoting the child's enjoyment of mother-child interactions. Maternal power assertion did not mediate the relation between MRO and conscience once the influence of the other mediators was considered. CONCLUSIONS: The impact of the early mother-child relationship on future conscience appears to be a complex process that progresses along distinct paths. PMID- 15660642 TI - The influence of perinatal complications and environmental adversity on boys' antisocial behavior. AB - BACKGROUND: The purpose of the present study was to test components of Raine's (2002) biosocial model, specifically the interactive effects of perinatal complications, rejecting parenting, and family adversity on the development of early-onset antisocial behavior (ASB). Boys' internalizing problems were also tested to investigate the specificity of the model. METHODS: Birth records in addition to longitudinal data were collected on 310 low-income boys followed from birth until 10 years of age. RESULTS: Findings demonstrated support for a biosocial framework in predicting ASB but not internalizing problems. Family adversity, and to some extent rejecting parenting, consistently predicted youth outcome. Perinatal complications emerged as a predictor of ASB but only in the context of other family risk factors. According to maternal report, boys experiencing high levels of perinatal complications, rejecting parenting, and family adversity showed significantly higher levels of ASB than boys with lower levels of these risk factors. This finding was partially corroborated by youth self-report, such that boys experiencing high levels of perinatal complications and family adversity reported more antisocial activity than boys experiencing no risk or risk in only one domain. CONCLUSIONS: This study supports the specific prediction of ASB in middle childhood from a biosocial model. Findings also highlight the salience of a negative psychosocial environment on childhood maladjustment. Intervention efforts including parenting skills and coping strategies for mothers of children from multiple risk environments are advocated. PMID- 15660643 TI - Risk factors of parents abused as children: a mediational analysis of the intergenerational continuity of child maltreatment (Part I). AB - BACKGROUND: This study provides an exploration of factors implicated in the intergenerational cycle of child maltreatment. Families with newborns where at least one of the parents was physically and/or sexually abused as a child (AP families) were compared in terms of risk factors to families where the parents had no childhood history of victimization (NAP families). The mediational properties of risk factors in the intergenerational cycle of maltreatment were then explored. METHODS: Information was collected by community nurses as a part of the 'health visiting' service. Data was collated across 4351 families, of which 135 (3.1%) had a parent who self-reported a history of abuse in childhood. The health visitor visited each family at home when the child was 4 to 6 weeks of age to assess the presence of risk factors. RESULTS: Within 13 months after birth, 9 (6.7%) AP families were referred for maltreating their own child in comparison to 18 (.4%) NAP families. Assessments found a significantly higher number of risk factors for AP families. Mediational analysis demonstrated that the presence of three significant risk factors (parenting under 21 years, history of mental illness or depression, residing with a violent adult) provided partial mediation of the intergenerational continuity of child maltreatment, explaining 53% of the total effect. CONCLUSION: Prevention may be possible, once a history of parental childhood abuse has been identified, by offering services in priority to those families where a parent is under 21 years, has a history of mental illness/depression and/or there is a violent adult residing in the household. However, it must also be acknowledged that these factors do not provide a full causal account of the intergenerational transmission and consideration should be given to additional factors, such as parenting styles (see Part II of this mediational model, Dixon, Hamilton-Giachritsis, and Browne, 2004). PMID- 15660644 TI - Attributions and behaviours of parents abused as children: a mediational analysis of the intergenerational continuity of child maltreatment (Part II). AB - BACKGROUND: This study extends previous research (Dixon, Browne, & Hamilton Giachritsis, 2004) by exploring the mediational properties of parenting styles and their relation to risk factors in the intergenerational cycle of child maltreatment. Families with newborns where at least one of the parents was physically and/or sexually abused as a child (AP families) were compared, in terms of parents' attributions and behaviour, to families where the parents had no childhood history of victimization (NAP families). METHODS: Information was collected from 4351 families (135 AP families) by community nurses as part of the 'health visiting' service. The same health visitor visited each family twice at home when the child was 4 to 6 weeks and 3 to 5 months of age, to assess behavioural indicators of positive parenting. RESULTS: Within 13 months after birth, 9 (6.7%) AP families were referred for maltreating their own child in comparison to 18 (.4%) NAP families. Assessments found a significantly higher number of risk factors and measures indicating poor parenting for AP families. Mediational analysis found that intergenerational continuity of child maltreatment was explained to a larger extent (62% of the total effect) by the presence of poor parenting styles together with the three significant risk factors (parenting under 21 years, history of mental illness or depression, residing with a violent adult). The three risk factors alone were less explanatory (53% of the total effect). CONCLUSION: This study provides an explanation for why a minority of parents abused in childhood go on to maltreat their own infant, evidencing poor parenting styles and mediating risk factors. Hence, prevention may be enhanced in AP families by the promotion of 'positive parenting' in addition to providing additional support to young parents, tackling mental illness/depression and domestic violence problems. PMID- 15660645 TI - Psychiatric disorders in parents of children with autism: a meta-analysis. AB - BACKGROUND: The genetic basis of autism has received great attention during the last few years. The psychiatric status of parents of persons with autism has been studied as part of the broad phenotype of autism. METHODS: In the current study we examined all studies in which psychiatric difficulties of parents of children with autism were compared to those of parents of other children, as part of the broad phenotype of autism. RESULTS: Overall, when parents of children with autism were compared to all other parents taken together, i.e., parents of typically developing children grouped together with parents of children with diagnoses other than autism, they were found to have more psychiatric difficulties, but this finding was inconsistent. Once type of comparison group (e.g., Down Syndrome, learning disabilities) and method of assessing psychiatric difficulties (e.g., self-report, clinical measures) were taken into account, different findings were yielded for the various comparison groups and methods of assessing psychiatric difficulties in parents, suggesting that parents of children with autism present more psychiatric difficulties, mostly in comparisons involving samples of parents of children with no known genetic risk (e.g., Down Syndrome). CONCLUSIONS: These findings suggest that in future studies, researchers should carefully consider the advantages and disadvantages of various comparison groups and other characteristics, such as method of assessment, gender, and level of functioning, in attempting to delineate the specificity and universality of the broad phenotype of autism. PMID- 15660646 TI - Attention and memory biases in the offspring of parents with bipolar disorder: indications from a pilot study. AB - BACKGROUND: Although children of bipolar parents are at heightened risk for developing emotional disorders, the processes underlying this vulnerability are not well understood. This study examined biases in the processing of emotional stimuli as a potential vulnerability marker of bipolar disorder. METHODS: Sixteen children of bipolar parents who did not show any indication of having an emotional disorder at the time of testing and ten children of never-disordered control parents underwent a negative mood induction designed to activate cognitive schemas and were then administered an emotion Stroop task and a self referent encoding task. RESULTS: Children of bipolar parents were found to exhibit an attentional bias towards social-threat and manic-irritable words. Furthermore, although high- and low-risk children did not differ in their endorsement of positive and negative words as self-descriptive, the high-risk children demonstrated better recall of negative words than did the low-risk children. CONCLUSIONS: Thus, children without a mood disorder who are at high risk for developing a mood disorder were found to exhibit biases in attention and memory that are similar to those found for bipolar and unipolar depressed adults, suggesting that children at increased risk for affective disorder are characterized by potentially pathogenic cognitive structures that can be activated by sad mood. These findings offer insights into mechanisms of cognitive vulnerability for bipolar disorders. PMID- 15660647 TI - Larger deficits in brain networks for response inhibition than for visual selective attention in attention deficit hyperactivity disorder (ADHD). AB - BACKGROUND: Brain activation differences between 12 control and 12 attention deficit hyperactivity disorder (ADHD) children (9- to 12-year-olds) were examined on two cognitive tasks during functional magnetic resonance imaging (fMRI). METHOD: Visual selective attention was measured with the visual search of a conjunction target (red triangle) in a field of distracters and response inhibition was measured with a go/no-go task. RESULTS: There were limited group differences in the selective attention task, with control children showing significantly greater intensity of activation in a small area of the superior parietal lobule region of interest. There were large group differences in the response inhibition task, with control children showing significantly greater intensity of activation in fronto-striatal regions of interest including the inferior, middle, superior and medial frontal gyri as well as the caudate nucleus and globus pallidus. CONCLUSION: The widespread hypoactivity for the ADHD children on the go/no-go task is consistent with the hypothesis that response inhibition is a specific deficit in attention deficit hyperactivity disorder. PMID- 15660649 TI - Cutaneous carcinosarcoma: adnexal vs. epidermal types define high- and low-risk tumors. Results of a meta-analysis. AB - OBJECTIVE: We report four cases of cutaneous carcinosarcoma (CS) and perform a meta-analysis of the cutaneous CS literature. RESULTS: CS occurred in elderly patients (mean of 80 years) on sun-damaged skin, and were keratotic papules of short duration. They did not recur after excision. CS exhibited basal cell carcinoma mixed with atypical fibroxanthoma cell populations. Immunophenotyping revealed vimentin+/keratin- spindle cells and vimentin-/keratin+ epithelial cells. Three cases exhibited p53 protein expression of both carcinomatous and sarcomatous components. Literature review identified 38 cases of cutaneous CS that could be broadly classified into two distinct groups. Epidermal-derived (basal or squamous cell carcinoma epithelial component) CS arose on the sun damaged skin of the head and neck of elderly males (mean age 72 years) and had a 70% 5-year disease-free survival. In contrast, adnexal CS (spiradenocarcinoma, porocarcinoma, proliferating tricholemmal cystic carcinoma, or matrical carcinoma) occurred in younger patients (mean age 58 years), showed recent growth in a long-standing nodule and had a 25% 5-year disease-free survival. Age less than 65 years, recent growth, long-standing skin tumor, and tumor size greater than 2 cm significantly correlated with poor outcome. CONCLUSIONS: Cutaneous CS is an aggressive skin cancer with high risk for advanced disease. Significant risk factors exist whose identification will allow for better management of CS patients. PMID- 15660650 TI - The application of a monoclonal antibody to CD62L on paraffin-embedded tissue samples in the assessment of the cutaneous T-cell infiltrates. AB - BACKGROUND: A reduction in the expression of the pan T-cell markers CD7 and CD62L supports an endogenous T-cell dyscrasia. Previously, clone availability for CD62L restricted its application to frozen tissue sections. MATERIALS AND METHODS: A nonavidin/biotin technique to examine CD3, CD62L, and CD7 in paraffin formalin fixed tissue in non-neoplastic and neoplastic T-cell infiltrates. RESULTS: In the reactive group, CD62L manifested a 15 and 22% reduction in epidermal and dermal staining, respectively; there was a 42 and 31% reduction in epidermal and dermal CD7 staining. In lymphomatoid hypersensitivity, CD62L showed a 24 and 9% reduction in epidermal and dermal staining, respectively; CD7 staining demonstrated reduced staining by 70 and 66% in the epidermis and dermis. In the non-lymphomatous endogenous T-cell dyscrasia and lymphoma categories, an 80% diminution in CD62L and CD7 expression was seen. CONCLUSIONS: CD62L can be successfully applied in formalin-fixed tissue and exhibits enhanced specificity compared to CD7 in the evaluation of cutaneous T-cell infiltrates. Both CD62L and CD7 in paraffin-embedded, formalin-fixed tissue are useful diagnostic adjuncts, especially in regard to the discrimination of lymphomatoid hypersensitivity reactions from true endogenous T-cell dyscrasia. PMID- 15660651 TI - HHV-8/KSHV during the development of Kaposi's sarcoma: evaluation by polymerase chain reaction and immunohistochemistry. AB - The human gamma-herpes virus-8 (HHV-8) was first described in AIDS-related Kaposi's sarcoma (KS) tumour samples. In this study, we report comparative studies on paraffin-embedded biopsies of AIDS-related KS (AKS) and endemic KS (EKS) with regard to HHV-8 content as evaluated using polymerase chain reaction (PCR) and immunohistochemistry. DNA was extracted either using Chelex-100 or using Qia-gene kit and was evaluated with the help of a semiquantitative PCR assay. The PCR detection of HHV-8 was more sensitive to the Chelex method than to Qia-gene. The threshold for PCR test sensitivity with the help of serial dilution of DNA was at the level of five plasmid ORF-26 regions, and DNA from 25 body cavity-based lymphoma-1 cells. The results expressed as virus load/actin unit showed progressively higher HHV-8 levels in late (nodular) cases, compared to those in early (patch/plaque) stages. Evaluation of HHV-8 DNA levels in tumour tissues, thus, indicates a correlation between virus load and KS stage. Double immunostaining of spindle cells (SC) in KS biopsies for CD34 and HHV-8/latency associated nuclear antigen (LANA) showed an increase in double-positive SC in the lesions of nodular AKS and EKS cases, compared to that in plaque and patch stages. However, 10-15% of CD34+/LANA- SC cells were observed during the development from patch to nodular cases of AKS and EKS. Our results indicate that PCR analysis is a simple and sensitive diagnostic method for HHV-8 evaluation in KS tissues, processed for conventional histopathology. PMID- 15660652 TI - Monkeypox virus: histologic, immunohistochemical and electron-microscopic findings. AB - BACKGROUND: Human monkeypox, an emerging viral zoonosis first recognized in Africa, has recently emerged in the mid-western US. Initially, it presents with skin eruptions and fevers with diaphoresis and rigors. Clinically, the skin lesions progress from papules to vesiculopustules to resolving eschars. METHODS: Three cutaneous biopsy specimens from two patients with polymerase chain reaction (PCR)-proven monkeypox were available for review. The histologic, immunohistochemical and electron-microscopic features were identified. RESULTS: The clinical progression of lesions is mirrored histologically with ballooning degeneration of basal keratinocytes and spongiosis of a mildly acanthotic epidermis progressing to full thickness necrosis of a markedly acanthotic epidermis containing few viable keratinocytes. A lichenoid-mixed inflammatory cell infiltrate is present, which exhibits progressive exocytosis with the keratinocyte necrosis. Inflammation of the superficial and deep vascular plexes, eccrine units and follicles is also present. Viral cytopathic effect is manifest by multinucleated syncytial keratinocytes. Immunohistochemically, viral antigen is detected within keratinocytes of the lesional epidermis, follicular and eccrine epithelium and few dermal mononuclear cells. Electron microscopy reveals virions at various stages of assembly within the keratinocyte cytoplasm. CONCLUSIONS: The histologic differential diagnosis includes herpes simplex virus, varicella and other pox viruses, such as smallpox. The first one may be differentiated histologically, immunohistochemically and electron microscopically. The last two may be differentiated using PCR assay for the monkeypox extracellular-envelope virus protein gene. PMID- 15660653 TI - Epstein-Barr virus- and human herpesvirus 8-associated primary cutaneous plasmablastic lymphoma in the setting of renal transplantation. AB - BACKGROUND: Plasmablastic lymphoma (PBL) is a recently recognized entity most often reported in the oral cavity, mainly in the setting of underlying human immunodeficiency viral infection whereby a role for Epstein-Barr virus (EBV) and more recently human herpesvirus 8 (HHV8) has been described. Although EBV has been implicated in a variety of lymphoproliferative lesions, until recently HHV8 has only been associated with primary effusion lymphoma, multicentric Castleman's disease, and Kaposi's sarcoma. We describe a case of PBL occurring in the setting of renal transplantation. METHODS: We encountered a case of PBL occurring in the setting of renal transplantation. We characterized the tumor by routine immunohistochemistry and evaluated for the presence of immunoglobulin light chain restriction and EBV RNA by in situ hybridization. We assessed for the presence of HHV8 RNA by reverse transcriptase in situ hybridization. RESULTS: The tumor showed a histomorphology compatible with a PBL. In addition, there was strong RNA expression in the neoplastic cells for EBER-1, EBER-2, and HHV8. CONCLUSION: This case suggests a possible role of both viruses in the pathogenesis of PBL in sites other than the oral cavity and expands the spectrum of post-transplantation lymphoproliferative disease to include PBL. PMID- 15660654 TI - The characteristic histopathologic features of nevi on and around the ear. AB - BACKGROUND: Nevi on certain areas of the body such as the acral, genital, and flexural regions may exhibit uncommon but characteristic histopathologic features. The purpose of this study was to characterize the distinctive features of nevi with a junctional component located on and around the ear. MATERIALS AND METHODS: A total of 101 compound and junctional nevi of the ear received at the Yale Dermatopathology Laboratory during a 10-year period were examined in this study. The most characteristic feature of the majority of these nevi was irregularity of nesting pattern, with nests, which varied in size and shape and which were sometimes located between rete ridges. RESULTS: Forty-two (42%) of nevi on and around the ear showed poor circumscription, lateral extension of the junctional component beyond the dermal component, and elongation of rete ridges with bridging between them. A subset of these nevi (26 cases) showed uniformly large melanocytes with large vesicular nuclei without prominent nucleoli, and abundant pale, finely granular cytoplasm. These lesions did not show a tendency to recur. CONCLUSIONS: This study supports the existence of a subset of nevi on or near the ear that, like certain nevi located on other special sites, exhibit unusual but characteristic features, which may be misinterpreted as atypical or malignant. PMID- 15660655 TI - Cluster designation 5 staining of normal and non-lymphoid neoplastic skin. AB - BACKGROUND: Immunohistochemical staining for cluster designation 5 (CD5) has been found to label a variety of non-lymphoid tumors. METHODS: A variety of eccrine, apocrine, follicular, epithelial, and pagetoid lesions were selected and stained with an anti-CD5 monoclonal antibody (Novocastra Labs, Newcastle upon Tyne, UK, clone 4C7) by immunohistochemistry. The intensity of positive cytoplasmic staining was graded semiquantitatively (1+ weak staining, 2+ strong staining). Additionally, the percentage of positive lesional cells was placed in one of four categories: >75%, 25-75%, 1-25%, and <1%. RESULTS: Within normal skin, CD5 labeled lymphocytes, apocrine glands, deep dermal eccrine glands, and smooth muscle (weak). The majority of benign and malignant apocrine lesions demonstrated strong focal (36%, n=11)-to-diffuse (64%, n=16) staining. In contrast, labeling of benign eccrine tumors was more focal, tending to localize around ducts (79%, n=19). Microcystic adnexal carcinoma demonstrated focal staining of deeper ductal structures (71%, n=7), whereas desmoplastic trichoepithelioma and basal cell carcinoma showed only rare positive cells. All cases of mammary (n=7) and extramammary (n=8) Paget's disease labeled diffusely for CD5. Pagetoid Bowen's disease (n=6), intraepidermal sebaceous carcinoma (n=3), nor melanoma in situ (n=6) showed any CD5 staining. CONCLUSIONS: Immunohistochemical staining for CD5 is extremely useful in the differential diagnosis of pagetoid epidermal lesions and will mark mammary and extramammary Paget's disease, but not pagetoid Bowen's disease, melanoma in situ, or sebaceous carcinoma. PMID- 15660657 TI - Congenital panfollicular nevus: report of a new entity. AB - The various forms of non-melanocytic nevi (hamartomas) are usually encountered in pediatric patients, and nevus sebaceous of Jadassohn is the most common to have undifferentiated pilosebaceous units. We report a unique congenital follicular nevus that fails to meet the criteria of any previously described follicular neoplasm, despite the plethora of alternatives. Clinically considered a syringocystadenoma papilliferum, the excised lesion contained multiple dermal nodules that exhibited nearly all stages of follicular differentiation. The periodicity of the follicular proliferations was akin to normal terminal hair, and a prominent perifollicular sheath surrounded each. This benign lesion of abortive hair follicles was unassociated with any established genodermatous syndrome or other adnexal neoplasm. PMID- 15660656 TI - Cyclooxygenase-2 expression in primary Merkel cell carcinoma. AB - BACKGROUND: Merkel cell carcinoma (MCC) is a highly malignant neuroendocrine skin tumor. The typical course of MCC is rapid progression of the primary tumor and metastatic dissemination to the regional lymph nodes. Thus far, no biological, prognostic marker has been established for this aggressive neoplasm. Cyclooxygenase-2 (Cox-2) is undetectable in most normal tissues, but it is induced in various cell types by inflammation and carcinogenesis. Although the expression and function of Cox-2 have been studied extensively in several carcinomas, little is known about Cox-2 expression in neuroendocrine carcinomas. The aim of the present report was to study Cox-2 expression in MCC and find out whether this expression correlates with outcome. METHODS: Immunohistochemical analysis for Cox-2 was performed on 22 primary MCC samples. RESULTS: Almost 70% of the samples showed positive staining. Protein expression of Cox-2 was sparse and low in intensity. We found a tendency for enhanced Cox-2 expression in tumors located in sun-exposed areas. Cox-2 expression had no significant statistical correlation with clinical parameters. CONCLUSIONS: MCC expresses Cox-2 in low levels, and the expression did not prove to be a prognostic factor. Furthermore, the low expression suggests that the primary treatment option for MCC is not therapeutic inhibition of Cox-2. PMID- 15660658 TI - Longitudinal melanonychia of the toenails with presence of Medlar bodies on biopsy. AB - A 9-year-old girl presented with a 2-year history of pigmented streaks on her second right toenail as well as on her fourth and fifth left toenails. The patient was otherwise asymptomatic with no other physical findings. Owing to parental concern, a biopsy was performed, which revealed numerous bacteria as well as Medlar bodies overlying the nail bed with no evidence of a nevomelanocytic lesion. To our knowledge, this is the first report of Medlar bodies causing pigmented streaks in the toenails. PMID- 15660660 TI - Proliferation, apoptosis, and survivin expression in a spectrum of melanocytic nevi. AB - BACKGROUND: Apoptosis is important for maintenance of tissue homeostasis and often dysregulated in cutaneous neoplasms. The apoptosis inhibitor survivin is expressed in melanoma and non-melanoma skin cancers and benign keratinocytic lesions. Its expression has not been studied in melanocytic nevi. OBJECTIVE: We determined the expression pattern of survivin in benign melanocytic nevi in comparison to markers of proliferation and apoptosis. METHODS: Six cases of each of the following melanocytic nevi were retrieved from a dermatopathology archive: compound dysplastic nevus, intradermal nevus, compound nevus, neurotized intradermal nevus, and Spitz nevus. Survivin expression was evaluated by in situ hybridization. Apoptotic and proliferation indices were calculated by counting immunoreactive cells in terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling and proliferating cell nuclear antigen immunostained sections, respectively. RESULTS: All nevi, regardless of histologic type, expressed survivin. Compound melanocytic lesions expressed survivin in both epidermal and dermal compartments. The apoptotic rate was low for dysplastic, compound, and Spitz nevi, and apoptotic cells were not identified in any neurotized nevus. The proliferative index was highest for Spitz nevi, while all other nevi demonstrated rare positive cells. CONCLUSIONS: Survivin is consistently expressed in benign melanocytic lesions, while apoptotic cells are rarely identified, suggesting the dysregulation of apoptotic pathways with the accumulation of cells in these neoplasms. PMID- 15660663 TI - Genetic study of the myelin oligodendrocyte glycoprotein (MOG) gene in schizophrenia. AB - Schizophrenia (SCZ) is a neuropsychiatric disorder that affects approximately 1% of the general population. The human leukocyte antigen (HLA) system has been implicated in several genetic studies of SCZ. The myelin oligodendrocyte glycoprotein (MOG) gene, which is located close to the HLA region, is considered a candidate for SCZ due to its association with white matter abnormalities and its importance in mediating the complement cascade. Four polymorphisms in the MOG gene (CA)n (TAAA)n, and two intronic polymorphisms, C1334T and C10991T, were investigated for the possibility of association with SCZ using 111 SCZ proband and their families. We examined the transmission of the alleles of each of these polymorphisms with the transmission disequilibrium test. We did not observe significant evidence for biased transmission of alleles at the (CA)n (chi2=2.430, 6 df, P=0.876) (TAAA)n (chi2=3.550, 5 df, P=0.616), C1334T (chi2=0.040, 1 df, P=0.841) and C10991T (chi2=0.154, 1 df, P=0.695) polymorphisms. Overall haplotype analysis using the TRANSMIT program was also not significant (chi2=7.954, 9 df, P=0.539). Furthermore, our results comparing mean age at onset in the genotype groups using the Kruskal-Wallis Test were not significant. Our case-control analyses (182 cases age-, sex- and ethnicity-matched with healthy controls) and combined z-score [(CA)n: z-score=-1.126, P=0.130; (TAAA)n: z-score=-0.233, P=0.408; C1334T: z-score=0.703, P=0.241; C10991T: z-score=0.551, P=0.291] were also not significant. Although our data are negative, the intriguing hypothesis for MOG in SCZ may warrant further investigation of this gene. PMID- 15660664 TI - A gain-of-function mutation in the GABA receptor produces synaptic and behavioral abnormalities in the mouse. AB - In mammalian species, inhibition in the brain is mediated predominantly by the activation of GABAA receptors. We report here changes in inhibitory synaptic function and behavior in a mouse line harboring a gain-of-function mutation at Serine 270 (S270) in the GABAA receptor alpha1 subunit. In recombinant alpha1beta2gamma2 receptors, replacement of S270 by Histidine (H) results in an increase in sensitivity to gamma-aminobutyric acid (GABA), and slowing of deactivation following transient activation by saturating concentrations of GABA. Heterozygous mice expressing the S270H mutation are hyper-responsive to human contact, exhibit intention tremor, smaller body size and reduced viability. These mice also displayed reduced motor coordination, were hypoactive in the home cage, but paradoxically were hyperactive in a novel open field environment. Heterozygous knockin mice of both sexes were fertile but females failed to care for offspring. This deficit in maternal behavior prevented production of homozygous animals. Recordings from brain slices prepared from these animals revealed a substantial prolongation of miniature inhibitory postsynaptic currents (IPSCs) and a loss of sensitivity to the anesthetic isoflurane, in neurons that express a substantial amount of the alpha1 subunit. The results suggest that the biophysical properties of GABAA receptors are important in determining the time course of inhibition in vivo, and suggest that the duration of synaptic inhibition is a critical determinant that influences a variety of behaviors in the mouse. PMID- 15660665 TI - Gene expression in the hippocampus of inbred alcohol-preferring and nonpreferring rats. AB - The hippocampus is sensitive to the effects of ethanol and appears to have a role in the development of alcohol tolerance. The objective of this study was to test the hypothesis that there are innate differences in gene expression in the hippocampus of inbred alcohol-preferring (iP) and -nonpreferring (iNP) rats that may contribute to differences in sensitivity to ethanol and/or in the development of tolerance. Affymetrix microarrays were used to measure gene expression in the hippocampus of alcohol-naive male iP and iNP rats in two experiments (n=4 and 6 per strain in the two experiments). Combining data from the two experiments, there were 137 probesets representing 129 genes that significantly differed (P < or = 0.01); 62 probesets differed at P < or = 0.001. Among the 36% of the genes that were expressed more in the iP than iNP rat at this level of significance, many were involved in cell growth and adhesion, cellular stress reduction and anti-oxidation, protein trafficking, regulation of gene expression, synaptic function and metabolism. Among the 64% of the genes that had lower expression in the hippocampus of iP than iNP rats were genes involved in metabolic pathways, cellular signaling systems, protein trafficking, cell death and neurotransmission. Overall, the data indicate that there are significant innate differences in gene expression in the hippocampus between iP and iNP rats, some of which might contribute to the differences observed in the development of alcohol tolerance between the selectively bred P and NP lines. PMID- 15660666 TI - Changes in gene expression within the nucleus accumbens and striatum following sexual experience. AB - Sexual experience, like repeated drug use, produces long-term changes including sensitization in the nucleus accumbens and dorsal striatum. To better understand the molecular mechanisms underlying the neuroadaptations following sexual experience, we employed a DNA microarray approach to identify genes differentially expressed between sexually experienced and sexually naive female hamsters within the nucleus accumbens and dorsal striatum. For 6 weeks, a stimulus male was placed in the home cage of one-half of the hormonally primed, ovariectomized female hamsters. On the seventh week, the two experimental groups were subdivided, with one half paired with a stimulus male. In comparison with sexually naive animals, sexually experienced hamsters receiving a stimulus male on week 7 exhibited an increase in a large number of genes. Conversely, sexually experienced female hamsters not receiving a stimulus male on week 7 exhibited a reduction in the expression of many genes. For directional changes and the categories of genes regulated by the experimental conditions, data were consistent across the nucleus accumbens and dorsal striatum. However, the specific genes exhibiting changes in expression were disparate. These experiments, among the first to profile genes regulated by female sexual behavior, will provide insight into the mechanisms by which both motivated behaviors and drugs of abuse induce long-term changes in the mesolimbic and nigrostriatal dopamine pathways. PMID- 15660667 TI - Association and linkage analysis of RGS4 polymorphisms with schizophrenia and bipolar disorder in Brazil. AB - Linkage and association studies in five independently ascertained samples have suggested that polymorphisms of the regulator of G-protein signaling 4 (RGS4) may confer risk for schizophrenia (SCZ). Suggestive evidence for association with bipolar disorder (BD) has also been presented. However, the associated alleles and haplotypes have differed among the samples. Data from other independent samples may clarify the putative associations. Hence, we investigated an independent, ethnically diverse Brazilian population comprising patients with SCZ (n=271) or BD1 (n=306), who were contrasted with 576 community-based controls. Parents of 49 SCZ cases and 44 BD cases were available for transmission disequilibrium tests (TDTs). Four RGS4 single-nucleotide polymorphisms (SNPs) 1, 4, 7 and 18 putatively associated with SCZ were investigated. In the SCZ samples, significant case-control differences were not observed for individual SNPs or haplotypes, though the TDT suggested transmission distortion similar to that observed in the initial report. For the BD sample, case-control comparisons revealed no significant differences for individual SNPs, but an omnibus test suggested differences in the overall distribution of haplotypes bearing all four SNPs (SNP-EM Omnibus likelihood ratio test; P=0.003). The TDT revealed over transmission of allele A at SNP7 (P=0.016), as well as haplotypes incorporating this allele. However, global tests incorporating all haplotypes yielded only suggestive trends for association (P=0.19). In conclusion, association with SCZ was not detected in the present analyses. The failure to detect an association may be related to inadequate power or to confounds related to ethnic admixture. Suggestive associations with BD detected here require further investigation in a larger sample. PMID- 15660668 TI - Tetrachlorodibenzo-p-dioxin exposure alters radial arm maze performance and hippocampal morphology in female AhR mice. AB - Perinatal exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) has been reported to alter spatial learning in rats tested on a radial arm maze (RAM). TCDD is believed to exert most of its effects through binding to the aryl hydrocarbon receptor (AhR). To determine whether the AhR mediates TCDD-induced alterations in spatial learning, we tested male and female AhR-knockout (AhR-/-), heterozygous (AhR+/-) and wild-type (AhR+/+) mice on the RAM. AhR+/- male and female mice were time mated, and treated dams were dosed with 5 microg TCDD/kg body weight on day 13 of gestation. When offspring reached adulthood, male and female AhR+/+, AhR+/- and AhR-/- mice from TCDD-exposed and unexposed litters were tested on the eight-arm RAM. After testing, we examined hippocampal morphology as visualized by the Timm's silver sulfide stain. TCDD-exposed female AhR+/- mice made more errors than their respective controls on the RAM and exhibited a decrease in the size of the intra- and infrapyramidal mossy fiber (IIP-MF) field of the hippocampus. None of the other TCDD-exposed groups differed from their respective control groups with regard to maze performance or hippocampal morphology. The reduction of IIP-MF field indicates a possible morphological basis for the learning deficit that was observed in the female AhR+/- mice. It is hypothesized that the effect of TCDD exposure is AhR dependent and that TCDD may alter GABAergic activity in the hippocampus of female mice during development. PMID- 15660669 TI - Different levels of gamma-synuclein mRNA in the cerebral cortex of dominant, neutral and submissive rats selected in the competition test. AB - Synucleins are small proteins regulating the filamentous network that in turn influences the release of dopamine and glutamate neurotransmitters involved in mood and motivation processes. We have studied the pattern of synuclein expression in animal models for mania and depression. Dominant behavior, as defined in a food competition test with dyads of rats, can serve as a model of mania and submissive behavior as a model of depression. The expression of alpha-, beta- and gamma-synuclein was analyzed in four regions of cortex from dominant, neutral and submissive rats using TaqMan reverse transcription-polymerase chain reaction technology. The expression levels of gamma-synuclein were elevated consistently in all regions of cerebral cortex of dominant rats (P <0.05; 23.5 +/ 1.1, normalized units) in contrast to the submissive rat group (10.3 +/- 1.2). Neutral rats had intermediate cerebral cortex levels of gamma-synuclein expression (15.7 +/- 1.4) that were significantly lower than that in dominant rats (P <0.05). No changes in alpha- or beta-synuclein expression were observed among the groups. These studies indicate that gamma-synuclein levels in the cerebral cortex were differentially associated with dominant and submissive behavior. PMID- 15660671 TI - Psychiatric patient and informant reports of patient behavior. AB - Informant reports of psychiatric patient behavior are collected routinely during intake interviews and to monitor therapeutic interventions. We investigated agreement between informant and adult psychiatric patient reports of patient behaviors (N=110). Behaviors were about substance use, physical and mental health, employment, illegal behavior, and recreational and social activities. Informant agreement with patients regarding whether or not a particular behavior occurred recently and also behavior frequency ratings were moderate overall (M r=.47 for frequency) and highly variable (rs=-.09 to .80). No informant, patient, or relationship characteristics were found consistently to moderate agreement. Thus, informants can report very accurately about the presence and frequency of some, but not all, patient behaviors. Low patient-informant agreement may be due to informants having relatively less detailed knowledge of patient behavior over time or patients not reporting accurately about certain behaviors. PMID- 15660672 TI - The role of mothers' use of control in children's perfectionism: implications for the development of children's depressive symptoms. AB - The central aim of this research was to investigate the possibility that when parents use heightened control with children, children develop perfectionistic concerns, which may foster depressive symptoms. Mothers' use of control with their elementary school children (N=104) was observed in the laboratory along with their affective expression toward their children; children's behavior (e.g., task engagement) that might influence mothers' use of control was also observed. Self-oriented and socially prescribed perfectionism and depression were assessed in children through self-report. Mothers using heightened control had children with perfectionistic concerns, particularly socially prescribed ones. This was not due to mothers' affective expression or children's behavior. Children's socially prescribed perfectionism mediated the link between mothers' use of heightened control and children's heightened depressive symptoms. PMID- 15660673 TI - Stress and mood: the moderating role of activity inhibition. AB - The present research tested the hypothesis that activity inhibition (as measured by the picture story exercise) moderates the relationship between stress and mood. Based on prior research that shows that (a) individuals high in activity inhibition restrain emotional and motivational impulses and (b) inhibiting negative emotion may lead to further emotional impairments, we assumed that individuals high in activity inhibition show evidence of increased negative mood when they are confronted with stressful events. Study 1 found evidence of increased negative mood-relevant thought among individuals high in activity inhibition who were confronted with the threat of social rejection. Study 2 found high negative mood among individuals high in activity inhibition who experienced many daily hassles. Also, participants high in activity inhibition reported a disproportionally higher amount of daily hassles in the middle (but not at the beginning) of a university semester. We suggest that this pattern of results can be accounted for by Wegner's (1994) theory of ironic effects of the suppression of thought and emotion. PMID- 15660674 TI - Appreciation: individual differences in finding value and meaning as a unique predictor of subjective well-being. AB - Adler (2002; Adler & Fagley, 2001) argued that being appreciative facilitates and enhances feelings of well-being and life satisfaction, as well as feelings of connection to what we have, to what we experience, and to life itself. In addition, expressing appreciation to others is believed to build social bonds. Although appreciation is viewed as a disposition, it is also viewed as something people can learn over time, making it an especially valuable construct to measure. Appreciating something (e.g. an event, a person, a behavior, an object) involves noticing and acknowledging its value and meaning and feeling a positive emotional connection to it. We defined eight aspects of appreciation and developed scales to measure them: a focus on what one has ("Have" Focus), Awe, Ritual, Present Moment, Self/Social Comparison, Gratitude, Loss/Adversity, Interpersonal. Scores on the subscales may be totaled to yield a score representing one's overall degree of appreciation (or level of appreciativeness) (coefficient alpha=.94). We also developed an 18-item short form (coefficient alpha=.91) that correlates .95 with scores on the long form. The scales correlated in predicted ways with measures of life satisfaction, positive affect, and negative affect. More importantly, appreciation was significantly related to life satisfaction and positive affect, even after the effects of optimism, spirituality, and emotional self-awareness had been statistically controlled. PMID- 15660675 TI - The socioemotional costs and benefits of social-evaluative concerns: do girls care too much? AB - This research investigated the hypothesis that girls' heightened concerns about social evaluation contribute to sex differences in depression and interpersonal competence during early adolescence. A short-term longitudinal study was conducted with 474 adolescents to examine the consequences of heightened social evaluative concerns. Adolescents reported on their levels of social-evaluative concerns and depressive symptoms. Teachers provided ratings of adolescents' competence with peers (displays of prosocial and aggressive behavior). As anticipated, girls demonstrated higher levels of social-evaluative concerns, depressive symptoms, and interpersonal competence than did boys. Moreover, path analysis confirmed that heightened social-evaluative concerns were associated both concurrently and over time with higher levels of depression, as well as with higher levels of interpersonal competence. Notably, social-evaluative concerns accounted fully for the sex difference in depression and partially for the sex difference in interpersonal competence. These findings highlight the need to consider both the socioemotional costs and benefits of sex-linked relational orientations. PMID- 15660676 TI - Searching for a developmental typology of personality and its relations to antisocial behavior: a longitudinal study of a representative sample of men. AB - The search for an empirically based personality typology has regained the interest of researchers. To date, however, the empirical inquiries were mainly cross-sectional. In this study, an empirically based developmental typology of personality was identified using data from a prospective longitudinal study of a representative sample of men assessed on four occasions from adolescence to midlife. Cluster analyses were performed on measures of Disinhibition, Negative Emotionality, and Extraversion. Four developmental types of personality were identified. The first was characterized by average scores in the three traits in adolescence that decreased rapidly until midlife (38%). The second was similar, but, instead, displayed increases in Extraversion during adulthood (32%). The third type showed high scores in Disinhibition and Negative Emotionality in adolescence that decreased progressively until midlife (23%). The fourth type was characterized by average scores in the three traits during adolescence followed by important increases in Negative Emotionality and decreases in Extraversion during adulthood (7%). These four developmental types of personality seemed to be related to known antisocial behavior trajectories. PMID- 15660677 TI - Forgivingness, vengeful rumination, and affective traits. AB - Trait forgivingness is the disposition to forgive interpersonal transgressions over time and across situations. We define forgiveness as the replacement of negative unforgiving emotions with positive, other-oriented emotions. Rumination has been suggested as a mediator between forgivingness and emotional outcomes; however, we suggest that different content of rumination leads to different outcomes after transgressions. In four studies of 179, 233, 80, and 66 undergraduate students, trait forgivingness was negatively correlated with trait anger, hostility, neuroticism, fear, and vengeful rumination and was positively correlated with agreeableness, extraversion, and trait empathy. The disposition to ruminate vengefully mediated the relationship between trait forgivingness and (1) anger-related traits and (2) both revenge motivations and state anger following a specific recent transgression, but it did not mediate between forgivingness and (1) fearfulness and (2) avoidance motivations following a specific transgression. Self-hate statements, a proxy for depressive rumination, mediated the relationship between forgivingness and both depression and fearfulness but not the relationship between forgivingness and trait anger. Future research should distinguish the contents of mental rumination following interpersonal transgressions. PMID- 15660678 TI - Nonlinear personality-performance relationships and the spurious moderating effects of traitedness. AB - This article evaluated the validity of two traditional traitedness measures (i.e., standard deviation traitedness measures and global, single-item traitedness measures) and three new traitedness measures (i.e., multi-item, ranking, and facet traitedness measures). Overall, there was poor construct validity for both the traditional and new measures of traitedness. Furthermore, traitedness was a spurious moderator of the relationship between openness to experience and academic performance. Finally, evidence of nonlinear personality academic performance relationships was discovered. PMID- 15660679 TI - Psychological responses to outbreak of severe acute respiratory syndrome: a prospective, multiple time-point study. AB - This study examined anxiety and coping responses to the outbreak of severe acute respiratory syndrome (SARS) in Hong Kong. Trait anxiety, situation-specific coping strategies, and coping flexibility were proposed as predictors of state anxiety during the early phase of the SARS epidemic. Seventy-two Hong Kong undergraduates (31 men, 41 women) participated in a prospective, multiple time point study. Participants' trait anxiety and coping flexibility had been assessed in an earlier study. Five months later, they reported their anxiety and coping at each of the four time points during the outbreak. Results revealed fluctuations in state anxiety across time points. Results from hierarchical linear modeling showed that trait anxiety as well as the situation-appropriate coping strategies of avoidance and personal hygiene practice accounted for changes in state anxiety. PMID- 15660680 TI - Comparison of coronary flow velocities between patients with obstructive and nonobstructive type hypertrophic cardiomyopathy: noninvasive assessment by transthoracic Doppler echocardiography. AB - BACKGROUND: We aimed to compare coronary flow velocity (CFV) measurements of patients with nonobstructive (NHCM) and obstructive hypertrophic cardiomyopathy (HOCM) by using transthoracic Doppler echocardiography (TTDE). METHODS AND RESULTS: In 11 patients with NHCM and 26 with HOCM, CFV in the distal left anterior descending (LAD) coronary was measured by TTDE (3.5 MHz) under the guidance of color Doppler flow mapping in addition to standard 2D and Doppler echocardiography. The results were compared with 24 normal participants who had no evidence of cardiac disease. Peak diastolic velocity of LAD was also higher in NHCM and HOCM than controls (52 +/- 14 cm/sec and 54 +/- 20 cm/sec vs 41 +/- 11 cm/sec, respectively, P < 0.01). The analysis of systolic velocities revealed abnormal flow patterns in 16 (61%) patients with HOCM (12 systolic-reversal flow and 4 no systolic flow) and 6 (54%) (5 reversal flow and 1 zero flow) patients with NHCM (-11 +/- 30 cm/sec and -13 +/- 38 cm/sec, vs 24 +/- 9 cm/sec, respectively, P < 0.001). Linear regression analysis demonstrated no correlation between intraventricular pressure gradient and coronary flow velocities in HOCM patients. However, there were significant positive and negative correlations between septal thickness and diastolic and systolic velocities, respectively (r = 0.50, P < 0.002, and r =-0.43, P < 0.005). CONCLUSION: We conclude that the coronary flow velocity abnormalities are independent from the type of hypertrophic cardiomyopathy. PMID- 15660681 TI - Role of transthoracic Doppler pulmonary venous flow pattern in large atrial septal defects. AB - OBJECTIVE: Large uncomplicated atrial septal defect (ASD) alters the pulmonary venous flow (PVF) pattern. We aimed to study the role of transthoracic echocardiography (TTE) in estimating the PVF Doppler abnormalities in ASD. By repeating the study soon after ASD closure, we correlated the hemodynamics of atrial shunting with PVF patterns. METHODS: This study was performed in a tertiary care referral teaching hospital in 2003. The TTE PVF patterns of 34 patients with ASD were studied. TTE study was reported by two blinded investigators independently. Surgical closure of ASD was done on eleven patients and the early postoperative PVF Doppler pattern was also studied with TTE. RESULTS: PVF patterns were adequately recorded in 34 of 38 (90%) subjects with ASD with equal male: female ratio (n = 17 each). The mean age of the study group was 21.4 +/- 8.7 years. ASD ranged from 10 to 38 mm in diameter with a mean of 18 +/- 4.2 mm. Continuous antegrade wave (CAW, mean 68.45 +/- 13.6 cm/s) replaced normally occurring S and D waves in all ASD patients. The atrial reversal wave was reduced or absent (mean 20.18 +/- 3.28 cm/sec). After ASD closure, the CAW was replaced by the S (46.18 +/- 7.5 cm/sec) and D waves (57.72 +/- 9.7 cm/sec) with increase in atrial reversal wave to 27.81 +/- 5.1 cm/sec. CONCLUSIONS: The S and D antegrade waves normally seen in PVF are replaced by a continuous antegrade wave in ASD. Atrial reversal wave is also reduced. PVF waveform becomes normal after ASD closure. TTE PVF Doppler pattern can help estimate ASD hemodynamics. PMID- 15660682 TI - Assessment of the tricuspid valve morphology by transthoracic real-time-3D echocardiography. AB - AIM: To demonstrate the feasibility of transthoracic three-dimensional real-time echocardiography (3D-TTE) supplemental to routine assessments of the tricuspid valve and to analyze interrater agreement. METHODS: Twenty healthy subjects and 74 patients with right ventricular failure were examined with conventional 2D and additionally 3D-TTE (SONOS 7500, Philips, Netherlands). The 3D exams were performed and recorded by one of two raters. The recordings were evaluated offline and independently by both raters for visualization of morphological and functional features of the tricuspid valve according to a subjective 3-point scale. Statistical analyses were performed for interrater agreement and for comparison of imaging quality between the two study groups. In addition, we present an illustrative case report. RESULTS: Visualization of the spatial relationship between the tricuspid valve and vicinal structures, of the commissures, the orifice, and entirety of valve depiction were better in the ventricular failure group as compared to the control group. Annular dimensions were equally assessable in both groups, leaflet thickness and mobility were not significantly different. Interrater agreement on assessability was slight for leaflet thickness, fair for leaflet mobility and orifice area, and good for the remaining features. The 3D-TTE exam including offline evaluation took 6.5 minutes on average and maximally 14 minutes. CONCLUSION: 3D-TTE of the tricuspid valve can be performed in addition to routine 2D echocardiography within a reasonable time and with high assessability of important features in patients with right ventricular failure. Interrater agreement was fair to good overall. Thus, its feasibility may encourage prospective studies on its potential for more detailed noninvasive diagnosis and preoperative planning. PMID- 15660684 TI - Traumatic aortic transection. AB - We report three cases of patients with blunt thoracic trauma, who underwent transesophageal echocardiography (TEE) because of high index of clinical suspicion for acute traumatic aortic syndrome. TEE revealed three different locations of aortic injury one of which was not diagnosed with aortography. PMID- 15660685 TI - Subaortic stenosis produced by an accessory mitral valve: the role of echocardiography. AB - Subaortic stenosis caused by an accessory mitral valve is an exceedingly rare finding. We report the case of an asymptomatic 14-year-old patient, in whom transthoracic echocardiography revealed an accessory mitral valve in the left ventricular outflow tract, producing mild subaortic stenosis. Except for an aneurysm of the interventricular septum, with no shunt, there were no other anomalies. Transesophageal echocardiography provided details about the morphology and location of the accessory valve. Being asymptomatic and having only a mild gradient, antibiotic prophylaxis for infective endocarditis and follow-up were recommended. After 2 years the patient is asymptomatic, with a similar echocardiographic gradient. PMID- 15660683 TI - Histological characterization of microlesions induced by myocardial contrast echocardiography. AB - BACKGROUND: Myocardial contrast echocardiography (MCE) has been shown to have a potential for apparently reversible side effects related to the interaction of ultrasound with the contrast microbubbles, including premature ventricular contractions and microvascular leakage. We investigated the potential for high dose MCE to induce histologically definable microlesions. METHODS: Myocardial contrast echocardiography with 1:4 end-systolic triggering was performed at 1.5 MHz and 1.7 mechanical index in a short axis view of the left ventricle in rats. Two high doses (500 microl/kg) of Optison agent were given 5 minutes apart during 10 minutes of echocardiography. For histology, the hearts were perfused and fixed in 10% neutral-buffered formalin. Slides from rats sacrificed 1 day after MCE were scored blind by a pathologist, and, in addition, photomicrographs in the anterior half were evaluated by digital image analysis. RESULTS: In rats sacrificed 10 minutes after MCE, microvascular leakage and petechiae were highly significant. However, lesions displaying necrotic debris associated with inflammatory infiltrates were not histologically evident at this time. Heart samples 24 hours after MCE showed microlesions with inflammatory infiltrates scattered primarily over the anterior half of the sections. Pathologically, there was inflammatory cell infiltration in areas of 0.6 +/- 0.5% for shams and 3.6 +/- 3.6% for MCE (P < 0.01). Analysis of photographs from the anterior wall found microlesion areas of 0.5 +/- 0.8% for shams and 7.4 +/- 5.0% for MCE (P < 0.02). For rats sacrificed 1 week and 6 weeks after MCE, the microlesions healed to form small fibrous regions interspersed with normal myocytes. CONCLUSION: High-dose MCE has a potential for causing microscale lesions in the myocardium and the possibility of therapeutic applications. PMID- 15660686 TI - Transesophageal echocardiographic diagnosis of aortic arch-left innominate vein fistula. AB - We present a patient with aortic arch-left innominate vein (LIV) fistula diagnosed by transesophageal echocardiography. Also, an interrupted linear echo was noted within the dilated LIV with flow signals moving across the area of interruption, suggestive of left innominate vein pseudo-aneurysm or dissection. The patient subsequently underwent successful surgical repair of the fistula. PMID- 15660687 TI - Severe intraventricular diastolic gradient due to hypertrophic cardiomyopathy and systolic left ventricular midcavitary obstruction. PMID- 15660688 TI - Giant cystic left atrial myxoma as a cause of secondary pulmonary hypertension. PMID- 15660689 TI - Isolated ventricular noncompaction with left ventricular thrombus. PMID- 15660690 TI - How not to miss a bicuspid aortic valve in the echocardiography laboratory. PMID- 15660691 TI - Assessing myocardial viability in ischemic cardiomyopathy. PMID- 15660692 TI - Viable myocardium: how much is enough? AB - Left ventricular systolic dysfunction is mainly a result of coronary artery disease (CAD). Decrease in myocardial contractility results as a response to a chronic hypoperfusion state that produces a change in cardiac myocyte metabolism, resulting in a perfusion-contraction mismatch in which function is sacrificed for survival. If revascularization is performed in a timely fashion, metabolism can be restored leading to recovery of function. Through the use of noninvasive imaging modalities, assessing myocardial viability can be easily performed and will aid in selecting those patients who will benefit from revascularization. Viable myocardium can be identified by nuclear modalities that have a high sensitivity but a lower specificity, such as thallium-201 single photon emission computed tomography and positron emission tomography (PET); or by the use of dobutamine stress echocardiogram (DSE), which has a decreased sensitivity but a better specificity. A modality that is increasingly being used with an overall good sensitivity and specificity is contrast-enhanced magnetic resonance imaging. The purpose of this review is to explore the amount of myocardial viability that is relevant to pursue revascularization, since as myocardial function improves there is a decrease in morbidity and mortality from heart failure and arrhythmias. PMID- 15660693 TI - Assessment of myocardial viability with dobutamine stress echocardiography in patients with ischemic left ventricular dysfunction. AB - The noninvasive assessment of myocardial viability has proved clinically useful for distinguishing hibernating and/or stunned myocardium from irreversibly injured myocardium in patients with chronic ischemic heart disease or recent myocardial infarction, with marked regional and/or global left ventricular (LV) dysfunction. Noninvasive techniques utilized for the detection of viability in asynergic myocardial regions include positron emission tomographic imaging of residual metabolic activity, single photon emission tomography (SPECT) of radioisotope uptake with thallium-201, low-dose dobutamine echocardiography assessment of inotropic reserve and myocardial contrast echocardiography for evaluation of microvascular integrity. Of these techniques, dobutamine stress echocardiography is a safe, widely available and relatively inexpensive modality for the identification of myocardial viability for risk stratification and prognosis. Low-dose dobutamine response can accurately predict improvement of dysfunctional yet viable myocardial regions, and thus identify a subset of patients whose LV function will improve following successful coronary revascularization. PMID- 15660694 TI - Assessment of myocardial viability using myocardial contrast echocardiography. AB - Myocardial contrast echocardiography (MCE) is a technique that uses microbubbles as a tracer during simultaneous ultrasound of the heart. The microbubbles can be used to provide quantitative information regarding the adequacy of myocardial blood flow (MBF), as well as the spatial extent of microvascular integrity. In acute myocardial infarction, MCE can identify the presence of collateral flow within the risk area, and can therefore predict preservation of myocardial viability and ultimate infarct size even prior to reperfusion. After reperfusion, the extent of microvascular no-reflow can be determined, and has significant implications for recovery of left ventricular function. In chronic ischemic heart disease, MCE has also been shown to successfully differentiate viable from necrotic myocardium. This technique can accurately predict recovery of function after revascularization. More importantly, MCE can be used to identify viable segments that may help to prevent infarct expansion and remodeling, and thus improve patient outcomes. PMID- 15660695 TI - Live three-dimensional transthoracic echocardiography: case study world atlas. PMID- 15660696 TI - Expression of E-cadherin and catenins in meningioma: ubiquitous expression and its irrelevance to malignancy. AB - The expression of cell adhesion molecules in 107 meningiomas was analyzed with immunohistochemical methods using antibodies to epithelial (E)-cadherin and catenins (alpha, beta and gamma). According to the provided World Health Organization (WHO) grading, 84, 18 and five cases were classified as grade I, II and III, respectively. In addition, hemangioblastoma (15 cases) and hemangiopericytoma (four cases) were also evaluated. In most meningiomas, E cadherin, alpha- and beta-catenins were expressed along the cell membrane or inside the cytoplasm. The tumor cells constituting whorls and glandular structures of secretory type showed a strong immunoreactivity. gamma-Catenin expression tended to be weak and infrequent in fibrous meningiomas, while other types exhibited diffuse stainings. Even in meningiomas of more than grade II, the expressions of cell adhesion molecules were detected in all cases. Hemangiopericytoma was positive for alpha- and beta-catenins, and hemangioblastomas were positive for beta-catenin alone, which was distinct from the expression pattern in meningiomas. Quantitatively, there were no correlations between the histological variants, Ki-67 indexes, or grades of meningiomas and the immunoreactive scores except for gamma-catenin scores of fibrous meningiomas. The present study demonstrates that cell adhesion molecules are ubiquitously expressed in all variants of meningioma and may be involved in the tumor morphogenesis. This result suggests that the expression of cell adhesion molecules is not a reliable indicator of malignancy in meningiomas. The present study also suggests that these markers may be useful for the differential diagnosis of meningioma. PMID- 15660697 TI - Reproducibility of the diagnosis of small adenocarcinoma of the lung and usefulness of an educational program for the diagnostic criteria. AB - Using 32 small adenocarcinomas of the lung including bronchioloalveolar carcinoma (BAC), the reproducibility of diagnosis by the modified diagnostic criteria for small adenocarcinoma (Cancer 75; 2844, 1995) and the effectiveness of an educational program for 27 volunteer general pathologists were examined. The average coincidence rate of the diagnosis before and after the program was 42.4% and 56.6%, respectively. The coincidence rate after the program was significantly higher than that before the program (P < 0.05). In contrast, the average coincidence rate of six lung cancer specialists was 71.4%, and this was significantly higher than that for general pathologists after the program (P < 0.05). When the cases were divided into two groups (in situ adenocarcinoma (BAC and BAC with alveolar collapse) and early invasive adenocarcinoma), the average coincidence rate for the general pathologists after the program increased to 85.3%, which was significantly higher than that before the program (80.3%; P < 0.05). The rate for the specialists was 89%, which was higher than that for the general pathologists after the program but not significantly so. This trial was thought to provide a theoretical background for the histological diagnosis of peripheral type adenocarcinoma of the lung and to justify the existing diagnostic criteria. PMID- 15660698 TI - Loss of expression of E-cadherin and beta-catenin is associated with progression of pulmonary adenocarcinoma. AB - The aim of the present study was to determine the association of loss of membranous expression of epithelial (E)-cadherin and beta-catenin with the progression of pulmonary adenocarcinoma. The expression of E-cadherin and beta catenin was examined in 154 cases of pulmonary adenocarcinoma, including 49 cases of atypical adenomatous hyperplasia (AAH), 40 cases of bronchioloalveolar carcinoma (BAC), 42 cases of BAC-dominant type of adenocarcinoma with mixed subtypes (early MX) and 23 cases of BAC-recessive type of adenocarcinoma with mixed subtypes (overt MX), by immunohistochemistry. E-cadherin expression was positive in all cases of AAH, in 37 cases (92.5%) of BAC and in 34 cases (81.0%) of early MX, while it was positive in three cases (13.0%) of overt MX. beta Catenin expression was positive in 47 cases (95.9%) of AAH, in 28 cases (70%) of BAC, in 32 cases (76.2%) of early MX and in 11 cases (47.8%) of overt MX. The rates of expression of E-cadherin and beta-catenin among cases of AAH, BAC, early MX and overt MX were significantly reduced. Loss of expression of E-cadherin and beta-catenin may play an important role in the progression of pulmonary adenocarcinoma, and these events occur before structural destruction of the alveolar wall by invasion of carcinoma cell. PMID- 15660700 TI - Pulmonary tumor thrombotic microangiopathy caused by a gastric carcinoma expressing vascular endothelial growth factor and tissue factor. AB - Pulmonary tumor thrombotic microangiopathy (PTTM) is a rare clinicopathological entity causing severe pulmonary hypertension. Its histological features include widespread tumor emboli along with fibrocellular intimal proliferation and thrombus formation in the small arteries and arterioles of the lungs. The result is occlusion or stenosis of the pulmonary vasculature, but the detailed pathogenesis has yet to be clarified in spite of the serious clinical manifestations. Herein is described the case of a 62-year-old man with a gastric adenocarcinoma who died of sudden cardiopulmonary arrest. The autopsy revealed advanced cancer disease as well as findings of PTTM, which seemed to be the cause of his unexpected death. The carcinoma cells were immunohistochemically positive for vascular endothelial growth factor (VEGF) and also for tissue factor (TF). There is another report suggesting that TF might play an important role in the pathogenesis of PTTM. Also, VEGF has been reported to be involved in a variety of forms of pulmonary hypertension and to be upregulated by TF. These findings suggest that VEGF and TF may be involved in the pathogenesis of PTTM. The present PTTM case, in which the tumor cells demonstrate the coexpression of VEGF and TF, is important in facilitating understanding of the lethal disorder in the future. PMID- 15660699 TI - Uninephrectomy induces progressive glomerulosclerosis and apoptosis in anti-Thy1 glomerulonephritis. AB - Administration of the anti-Thy1 antibody in rats induces reversible glomerulonephritis resembling human mesangiolytic and mesangioproliferative diseases. The purpose of the present study was to design a model of irreversible glomerulosclerosis, using the anti-Thy1 antibody injection after uninephrectomy, and examine it, focusing on apoptosis in the process of progressive sclerotic changes. Wistar rats were divided into three groups: one-kidney groups (group I and III) and a two-kidney group (group II). All groups were injected with the anti-Thy1 antibody (OX-7) at day 0, and group I and III were uninephrectomized at day -6. Only group III rats were given a half dose of OX-7 as compared with group I and II. Rats were killed for histological examinations at days 7, 14 and 30. In group I, progressive glomerular lesions, such as glomerular adhesion to Bowman's capsule, crescent formation, and collapse of capillary tufts were observed at days 14 and 30. No significant differences were observed in the pathological findings between group I and III. There was a significantly higher number of glomerular terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling positive cells in group I as compared to group II at days 7 and 14. Moreover, the glomerular expression of transforming growth factor-beta, heparan sulfate proteoglycan and chondroitin sulfate proteoglycan significantly increased in group I as compared to group II at days 7 and 14. Progressive glomerulosclerosis can be induced in the rat by a single injection of the anti-Thy1 antibody after unilateral nephrectomy. It is suggested that apoptosis and extracellular matrix accumulation play an important role in the development of glomerulosclerosis. PMID- 15660702 TI - Royal Australian and New Zealand College of Psychiatrists clinical practice guidelines for the treatment of schizophrenia and related disorders. AB - BACKGROUND: The Royal Australian and New Zealand College of Psychiatrists is co ordinating the development of clinical practice guidelines (CPGs) in psychiatry, funded under the National Mental Health Strategy (Australia) and the New Zealand Health Funding Authority. This paper presents CPGs for schizophrenia and related disorders. Over the past decade schizophrenia has become more treatable than ever before. A new generation of drug therapies, a renaissance of psychological and psychosocial interventions and a first generation of reform within the specialist mental health system have combined to create an evidence-based climate of realistic optimism. Progressive neuroscientific advances hold out the strong possibility of more definitive biological treatments in the near future. However, this improved potential for better outcomes and quality of life for people with schizophrenia has not been translated into reality in Australia. The efficacy effectiveness gap is wider for schizophrenia than any other serious medical disorder. Therapeutic nihilism, under-resourcing of services and a stalling of the service reform process, poor morale within specialist mental health services, a lack of broad-based recovery and life support programs, and a climate of tenacious stigma and consequent lack of concern for people with schizophrenia are the contributory causes for this failure to effectively treat. These guidelines therefore tackle only one element in the endeavour to reduce the impact of schizophrenia. They distil the current evidence-base and make recommendations based on the best available knowledge. METHOD: A comprehensive literature review (1990-2003) was conducted, including all Cochrane schizophrenia reviews and all relevant meta-analyses, and a number of recent international clinical practice guidelines were consulted. A series of drafts were refined by the expert committee and enhanced through a bi-national consultation process. TREATMENT RECOMMENDATIONS: This guideline provides evidence-based recommendations for the management of schizophrenia by treatment type and by phase of illness. The essential features of the guidelines are: (i) Early detection and comprehensive treatment of first episode cases is a priority since the psychosocial and possibly the biological impact of illness can be minimized and outcome improved. An optimistic attitude on the part of health professionals is an essential ingredient from the outset and across all phases of illness. (ii) Comprehensive and sustained intervention should be assured during the initial 3-5 years following diagnosis since course of illness is strongly influenced by what occurs in this 'critical period'. Patients should not have to 'prove chronicity' before they gain consistent access and tenure to specialist mental health services. (iii) Antipsychotic medication is the cornerstone of treatment. These medicines have improved in quality and tolerability, yet should be used cautiously and in a more targeted manner than in the past. The treatment of choice for most patients is now the novel antipsychotic medications because of their superior tolerability and, in particular, the reduced risk of tardive dyskinesia. This is particularly so for the first episode patient where, due to superior tolerability, novel agents are the first, second and third line choice. These novel agents are nevertheless associated with potentially serious medium to long-term side-effects of their own for which patients must be carefully monitored. Conventional antipsychotic medications in low dosage may still have a role in a small proportion of patients, where there has been full remission and good tolerability; however, the indications are shrinking progressively. These principles are now accepted in most developed countries. (vi) Clozapine should be used early in the course, as soon as treatment resistance to at least two antipsychotics has been demonstrated. This usually means incomplete remission of positive symptomatology, but clozapine may also be considered where there are pervasive negative symptoms or significant or persistent suicidal risk is present. (v) Comprehensive psychosocial interventions should be routinely available to all patients and their families, and provided by appropriately trained mental health professionals with time to devote to the task. This includes family interventions, cognitive-behaviour therapy, vocational rehabilitation and other forms of therapy, especially for comorbid conditions, such as substance abuse, depression and anxiety. (vi) The social and cultural environment of people with schizophrenia is an essential arena for intervention. Adequate shelter, financial security, access to meaningful social roles and availability of social support are essential components of recovery and quality of life. (vii) Interventions should be carefully tailored to phase and stage of illness, and to gender and cultural background. (viii) Genuine involvement of consumers and relatives in service development and provision should be standard. (ix) Maintenance of good physical health and prevention and early treatment of serious medical illness has been seriously neglected in the management of schizophrenia, and results in premature death and widespread morbidity. Quality of medical care for people with schizophrenia should be equivalent to the general community standard. (x) General practitioners (GPs)s should always be closely involved in the care of people with schizophrenia. However, this should be truly shared care, and sole care by a GP with minimal or no specialist involvement, while very common, is not regarded as an acceptable standard of care. Optimal treatment of schizophrenia requires a multidisciplinary team approach with a consultant psychiatrist centrally involved. PMID- 15660701 TI - Autopsy case of CD4/CD8 cutaneous T-cell lymphoma presenting disseminated pagetoid reticulosis with aggressive granulomatous invasion to the lungs and pancreas. AB - Pagetoid reticulosis is a rare cutaneous T-cell lymphoma with striking epidermotropism similar to that present in Paget's disease. There are two forms of pagetoid reticulosis: localized and disseminated. Reported herein is an autopsy case of disseminated pagetoid reticulosis with CD4(-)/CD8(-) phenotype T cells and massive invasion of the lungs and pancreas. The abnormal cells in the epidermis expressed a protein derived from a rearranged T-cell receptor beta gene, and this feature was used to confirm the monoclonality of these cells by polymerase chain reaction. At present, the World Health Organization (WHO) classification system considers pagetoid reticulosis to be an indolent form of primary cutaneous T-cell lymphoma and a variant of mycosis fungoides/Sezary syndrome with prominent epidermotropism. Some differences have been observed between pagetoid reticulosis and mycosis fungoides in terms of clinical course, tumor cell phenotype, and genetic findings; and these differences are highlighted in the present case. The relation between disseminated pagetoid reticulosis, CD4( )/CD8(-) cutaneous T-cell lymphoma, and gammadelta T-cell lymphoma, including whether pagetoid reticulosis is a variant of mycosis fungoides, remains unclear. PMID- 15660703 TI - Mental health research and philanthropy: possible partnerships? AB - Mental health research has received relatively little philanthropic support in Australia compared with other areas of health research. Philanthropic trusts do not generally provide recurrent funding or make grants for that perceived to be the responsibility of the state or the market. The emergence of 'strategic philanthropy' however, provides potential for mental health researchers to form partnerships with philanthropic foundations, particularly on initiatives that are focused on prevention and innovative and sustainable models with the capacity to 'go to scale' across the service system. PMID- 15660704 TI - Clinical profiles, scope and general findings of the Western Sydney First Episode Psychosis Project. AB - OBJECTIVE: To examine the clinical profile, treatment and social functioning of a community-based sample of young people presenting with their first episode of psychosis. METHODS: Over a 2-year period, young people with their first episode of psychosis referred to early intervention services in two area mental health services in western Sydney were assessed with a battery of clinical, neuropsychological, psychophysiological and neuroanatomical measures. This paper reports the clinical results of the baseline section of the study. RESULTS: Of the 224 referrals to the project, 94 subjects meet inclusion criteria and agreed to take part. Subjects were divided into three diagnostic groups- 'Schizophrenia', 'Mood Disorders' and 'Mixed Psychosis', the latter principally comprised of substance induced psychotic disorders. Subjects from the 'Schizophrenia' group differed significantly from the other two groups in that they had higher levels of negative symptoms and general psychopathology, and were less likely to be employed or engaged in study. They had poorer overall social functioning. Subjects with 'Mixed Psychosis' were similar to those from the 'Schizophrenia' group in that they were older and male, but they did not have the same burden of negative symptoms as the 'Schizophrenia' group. The 'Mood Disorders' group was younger, female and had overall a higher level of psychosocial functioning than the other two groups. Subjects from the 'Mood Disorders' group were more likely to be managed with mood stabilisers and multiple drug therapies. The use of atypical antipsychotic medication was almost universal. CONCLUSIONS: Even shortly after the time of presentation to mental health services young people with a schizophrenia spectrum diagnosis have a heavier burden of symptoms and are significantly more impaired by them than young people with other psychotic illnesses. This and their symptom profile differentiated them from young people with other psychotic disorders. PMID- 15660705 TI - Assessing cost-effectiveness of drug interventions for schizophrenia. AB - OBJECTIVE: To assess from a health sector perspective the incremental cost effectiveness of eight drug treatment scenarios for established schizophrenia. METHOD: Using a standardized methodology, costs and outcomes are modelled over the lifetime of prevalent cases of schizophrenia in Australia in 2000. A two stage approach to assessment of health benefit is used. The first stage involves a quantitative analysis based on disability-adjusted life years (DALYs) averted, using best available evidence. The robustness of results is tested using probabilistic uncertainty analysis. The second stage involves application of 'second filter' criteria (equity, strength of evidence, feasibility and acceptability) to allow broader concepts of benefit to be considered. RESULTS: Replacing oral typicals with risperidone or olanzapine has an incremental cost effectiveness ratio (ICER) of 48,000 Australian dollars and 92,000 Australian dollars/DALY respectively. Switching from low-dose typicals to risperidone has an ICER of 80,000 Australian dollars. Giving risperidone to people experiencing side effects on typicals is more cost-effective at 20,000 Australian dollars. Giving clozapine to people taking typicals, with the worst course of the disorder and either little or clear deterioration, is cost-effective at 42,000 Australian dollars or 23,000 Australian dollars/DALY respectively. The least cost-effective intervention is to replace risperidone with olanzapine at 160,000 Australian dollars/DALY. CONCLUSIONS: Based on an 50,000 Australian dollars/DALY threshold, low-dose typical neuroleptics are indicated as the treatment of choice for established schizophrenia, with risperidone being reserved for those experiencing moderate to severe side-effects on typicals. The more expensive olanzapine should only be prescribed when risperidone is not clinically indicated. The high cost of risperidone and olanzapine relative to modest health gains underlie this conclusion. Earlier introduction of clozapine however, would be cost-effective. This work is limited by weaknesses in trials (lack of long-term efficacy data, quality of life and consumer satisfaction evidence) and the translation of effect size into a DALY change. Some stakeholders, including SANE Australia, argue the modest health gains reported in the literature do not adequately reflect perceptions by patients, clinicians and carers, of improved quality of life with these atypicals. PMID- 15660706 TI - Disability, employment and work performance among people with ICD-10 anxiety disorders. AB - OBJECTIVE: To ascertain at a population level, patterns of disability, labour force participation, employment and work performance among people with ICD-10 anxiety disorders in comparison to people without disability or long-term health conditions. METHOD: A secondary analysis was conducted of a probability sample of 42 664 individuals collected in an Australian Bureau of Statistics (ABS) national survey in 1998. Trained lay interviewers using ICD-10 computer-assisted interviews identified household residents with anxiety disorders. RESULTS: Anxiety disorders were associated with: reduced labour force participation, degraded employment trajectories and impaired work performance compared to people without disabilities or long-term health conditions. CONCLUSION: People with anxiety disorders may need more effective treatments and assistance with completing education and training, joining and rejoining the workforce, developing career pathways, remaining in the workforce and sustaining work performance. A whole-of-government approach appears needed to reduce the burden of disease and increase community labour resources. Implications for clinicians, vocational professionals and policy makers are discussed. PMID- 15660707 TI - Public and private psychiatry: can they work together and is it worth the effort? AB - OBJECTIVE: Partnerships in mental health care, particularly between public and private psychiatric services, are being increasingly recognized as important for optimizing patient management and the efficient organization of services. However, public sector mental health services and private psychiatrists do not always work well together and there seem to be a number of barriers to effective collaboration. This study set out to investigate the extent of collaborative 'shared care' arrangements between a public mental health service and private psychiatrists practising nearby. It also examined possible barriers to collaboration and some possible solutions to the identified problems. METHOD: A questionnaire examining the above factors was sent to all public sector mental health clinicians and all private psychiatrists in the area. RESULTS: One hundred and five of the 154 (68.2%) public sector clinicians and 103 of the 194 (53.1%) private psychiatrists returned surveys. The main barriers to successful collaboration identified by members of both sectors were: 'Difficulty communicating' endorsed by 71.4% of public clinicians and 72% of private psychiatrists, 'Confusion of roles and responsibilities' endorsed by 62.9% and 66%, respectively, and 'Different treatment approach' by 47.6% and 45.6%, respectively. Over 60% of private psychiatrists identified problems with access to the public system as a barrier to successful shared care arrangements. It also emerged, as hypothesized, that the public and private systems tend to manage different patient populations and that public clinicians in particular are not fully aware of the private psychiatrists' range of expertise. This would result in fewer referrals for shared care across the sectors. CONCLUSIONS: A number of barriers to public sector clinicians and private psychiatrists collaborating in shared care arrangements were identified. The two groups surveyed identified similar barriers. Some of these can potentially be addressed by changes to service systems. Others require cultural shifts in both sectors. Improved communications including more opportunities for formal and informal meetings between people working in the two sectors would be likely to improve the understanding of the complementary sector's perspective and practice. Further changes would be expected to require careful work between the sectors on training, employment and practice protocols and initiatives, to allow better use of the existing services and resources. PMID- 15660708 TI - Changing demand for mental health services in the emergency department of a public hospital. AB - OBJECTIVE: Deinstitutionalization and mainstreaming may have contributed to increased attendance in public emergency departments by people with mental health problems. This study describes changing patterns of attendances by patients with mental health problems to the emergency department (ED) of a public teaching hospital in Adelaide, South Australia. METHOD: Records from a 10-year period from the ED were examined to identify changes in the number of, and diagnoses for, patients attending for primarily mental health concerns. Admission rates, detention and length of stay (LOS) were also examined in an attempt to identify trends. RESULTS: A tenfold increase in the number of patients attending the ED with primarily mental health problems has occurred over the 10-year period. This is within the context of relatively stable total ED presentations. The increase has been observed in all diagnostic categories although the greatest increase, by percentage, has been for psychotic disorders. A lesser increase was observed for patients presenting with overdose. People presenting with psychotic disorders are also more likely to be detained and admitted. LOS in the ED has also increased along with increasing demand. CONCLUSIONS: Reasons for the increased demand are likely multifactorial. While deinstitutionalization and mainstreaming have contributed, the closure of the ED at the local psychiatric hospital does not account entirely for the change. Insufficient community-based mental health services may also contribute to the reasons why people present to the ED and lack of inpatient beds contributes to the increasing LOS experienced in the ED. PMID- 15660709 TI - Psychosocial support for war-traumatized child and adolescent refugees: evaluation of a short-term treatment program. AB - OBJECTIVE: The purpose of this study was to evaluate a newly designed psychosocial treatment program for war traumatized child and adolescent refugees. The program was designed to reduce emotional distress and improve psychosocial functioning. METHOD: Ten young Kosovan refugees (mean age 13.3 years) residing in Germany participated in the manual based intervention program. This multimodal program consists of individual, family and group sessions using a psychoeducational approach beside trauma and grief focusing activities, creative techniques and relaxation. Kind and severity of traumatic experiences were gathered by interviewing the child and their caretakers using the Harvard Trauma Questionnaire (HTQ). Psychiatric diagnoses (Schedule for Affective Disorders and Schizophrenia for School-Age Children, K-SADS) were assessed prior to the intervention. Post-traumatic symptomatology (HTQ), emotional problems (Diagnostic System for Psychological Disorders, DYSIPS) and the overall psychosocial functioning (Child Global Assessment Scale, CGAS) were assessed before and after the 12-week intervention. RESULTS: Following the intervention the degree of overall psychosocial functioning increased substantially in 9 of 10 participants. Furthermore, post-traumatic, anxiety and depressive symptoms were reduced significantly. The rate of post-traumatic stress disorder (PTSD) diagnoses fell from 60% to 30%. The number of patients with PTSD and a high rate of comorbid symptoms (depression and anxiety) as well as a history of severe traumatization remained at 30%. CONCLUSIONS: This study suggests that the psychosocial treatment program specified for war traumatized adolescents may be useful for the relief of psychiatric sequelae and for an improvement in overall psychosocial functions, but not for the subgroup of severely traumatized patients with complex psychiatric disturbances. PMID- 15660710 TI - Grief experiences of parents whose children suffer from mental illness. AB - OBJECTIVE: To examine the grief experience of parents of adult children with a mental illness and its relationship to parental health and well-being and parent child attachment and affective relationship. METHODS: Participants were recruited from a variety of organizations throughout Australia that provide support services for sufferers of mental illness and/or for their families. Seventy-one participants (62 mothers and nine fathers) all of whom had a child diagnosed with mental illness volunteered to take part in the study. All completed measures of grief, health status and parent-child relationship. RESULTS: Parents reported experiencing grief in relation to their child's illness as evidenced by intrusive thoughts and feelings and avoidance of behaviour as well as difficulties adapting to and distress associated with reminders of the illness. Parental grief appears to reduce over time, but only in some aspects of grief and after an extended period. Increased parental grief was related to lowered psychological well-being and health status and associated with an anxious/ambivalent and a negative affective parent-child relationship. CONCLUSION: The study provides important insights into the grief experiences of parents following their child's diagnosis with mental illness. The significant relationship between parental grief and parental psychological well-being and health status as well as to parent-child relationship has important implications for health professionals. Foremost amongst these are the need to validate the distress and grief of parents and to better understand how to provide interventions that promote grief work and family bonds while reducing emotional distress and life disruption. PMID- 15660711 TI - Mortality and further suicidal behaviour after an index suicide attempt: a 10 year study. AB - OBJECTIVE: To examine further suicide attempts and mortality in the 10 years after a suicide attempt requiring hospital admission. METHOD: Participants were a consecutive series of 3690 individuals admitted to Christchurch Hospital for attempted suicide during the 10-year period 1993-2002. Data were obtained on admissions to Christchurch Hospital for attempted suicide during the study period. Mortality subsequent to the index suicide attempt was established from the National Mortality Database. The influence of age, gender and method of index suicide attempt on mortality and further suicide attempts requiring hospitalization were examined. RESULTS: Within 10 years, 28.1% of those who had been admitted for an index suicide attempt were readmitted for a further non fatal suicide attempt, and 4.6% died by suicide. Risks and rates of readmission were higher in: females; those under 55; and those whose index attempt involved a method of low lethality. Risks and rates of suicide were higher in: males; those aged 25 and over; and those using an index suicide attempt method of high lethality. Risks and rates of readmission and mortality from suicide were highest in the first 2 years after the index attempt, although deaths and readmissions occurred throughout the 10 years study period. CONCLUSIONS: Those making suicide attempts requiring hospital admission are at high risk of further hospitalization for suicide attempt and of death from suicide. These findings suggest a need for ongoing support and monitoring, and for enhanced treatment and management of all those making suicide attempts which require hospital admission in an effort to reduce risks of further suicidal behaviour. PMID- 15660712 TI - Non-suicidal deaths following hospital-treated self-poisoning. AB - OBJECTIVE: To quantify the non-suicidal mortality subsequent to hospital treated self-poisoning, and to identify risk factors for non-suicidal death. METHOD: A prospective longitudinal cohort design was used, with data-linkage between the Hunter Area Toxicology Service database and the National Death Index. All patients with deliberate self-poisoning for a 10-year period (1991-2000) were studied and the first episode in the period was used as the index episode. The outcomes were: accidental, 'natural' and non-suicidal death, with follow-up for the study duration. RESULTS: There were 4044 patients studied, and 170 (4.2%) of these had non-suicidal death; 64 were accidental and 106 were 'natural' cause deaths. The standardized mortality ratio (95% CI) for non-suicidal death for males, females and combined were 4.98 (4.08-6.07), 3.78 (3.0-4.75) and 4.20 (3.62 4.88), respectively. The increased mortality was apparent for both males and females, and was more marked in the younger age groups. For non-suicidal death the adjusted hazard ratio (95% CI) for increased risk were: increasing age 1.07 (1.06-1.08), male gender 1.77 (1.24-2.52), psychiatric diagnosis of substance related disorder 1.49 (1.03-2.16), prescription of a respiratory drug 2.69 (1.31 5.55), and prescription of an anti-diabetic drug 1.95 (0.93-4.07), while psychiatric diagnosis of adjustment disorder 0.64 (0.38-1.053) was associated with decreased risk. CONCLUSIONS: Patients who present with self-poisoning have increased mortality from accidental and 'natural' causes. Long-term treatment goals for these patients need to address non-suicide mortality in addition to suicide mortality. PMID- 15660713 TI - Olanzapine in the treatment of resistant depression. PMID- 15660714 TI - Lithium augmentation in milnacipran-refractory depression for the prevention of relapse following electroconvulsive therapy. PMID- 15660715 TI - Dissociative identity disorder. PMID- 15660717 TI - Body dysmorphic disorder treated with bupropion: cases report. PMID- 15660718 TI - Olanzapine augmentation of fluoxetine in trichotillomania: two cases. PMID- 15660719 TI - Neuroleptic malignant syndrome with promethazine. PMID- 15660726 TI - Is the 'Accelerator Hypothesis' worthy of our attention? PMID- 15660727 TI - Insulin treatment and cardiovascular disease; friend or foe? A point of view. AB - BACKGROUND: Several observational studies have shown that higher insulin levels are associated with an increased risk of cardiovascular disease. If higher endogenous insulin levels are causally related to cardiovascular disease, one might expect an increased risk of cardiovascular disease in patients treated with insulin, as this results in high circulating insulin levels. Such risk elevation might counteract the benefits of tight glucose control. Our objective was to explore the relationship between insulin therapy and cardiovascular disease in Type 1 and Type 2 diabetes mellitus using information from available literature. SUMMARY OF COMMENT: Several experimental studies in animals and humans support the presence of a harmful effect of insulin on the vascular endothelium. In prospective follow-up studies increased insulin dosage was associated with increased risks of cardiovascular disease, although confounding by indication could not be excluded. Randomized controlled trials in diabetic patients, comparing conventional with intensive glucose-lowering treatment, although showing a reduction in microvascular disease, showed no significant difference in the incidence of cardiovascular disease. The results with respect to exposure to insulin are, however, difficult to interpret due to insufficient information on exposure to insulin levels as well as confounding by glycaemic control and body mass index. In addition, these studies were not designed to address the question whether higher insulin use relates to increased cardiovascular risk. CONCLUSION: Published research provides conflicting evidence as to whether exposure to high levels of exogenous insulin in diabetes mellitus affects the risk of cardiovascular disease. The currently available studies have a number of serious methodological restraints that limit accurate interpretation and conclusions in this area. PMID- 15660728 TI - Creation of a multidisciplinary, evidence based, clinical guideline for the assessment, investigation and management of acute diabetes related foot complications. AB - AIMS: To design a multidisciplinary, evidenced-based, clinical guideline for the assessment, investigation and management of inpatients with acute diabetes related foot complications. METHODS: A systematic search of both published (identified by searching all major electronic databases and hand searching key journals) and unpublished literature (derived from national and internationally recognized experts) identified 266 articles specific to diabetes related foot complications. Of these, 126 (47%) were assessed to be methodologically sound and clinically relevant. A narrative summary with the articles tabulated according to their level of evidence was prepared. A multidisciplinary expert group of health professionals, with a known interest and recognized expertise in diabetes related foot complications, was established to assess the evidence. RESULTS: The multidisciplinary expert group used the identified literature and clinical experience to create a comprehensive, evidence-based, clinical guideline for the assessment, investigation and management of acute diabetes related foot complications. Included within the guideline is a novel, diabetes specific classification system, which codes for the presence/absence and severity of the four principle causative factors (Neuropathy, Vascular compromise, Ulceration and Infection) in the development of acute diabetes related foot complications. CONCLUSION: Through the creation and implementation of this evidence-based clinical guideline, specific for acute diabetes related foot complications, it is hoped that health professionals will be better equipped to make informed decisions for this patient population. This may benefit the individual and health system through reductions in amputation rate, length of hospital stay and health expenditure. PMID- 15660729 TI - Clinical characteristics, beta-cell function, HLA class II and mutations in MODY genes in non-paediatric subjects with Type 1 diabetes without pancreatic autoantibodies. AB - OBJECTIVE: To study clinical characteristics, beta-cell function, HLA typing and mutations in the hepatocyte nuclear factor (HNF)-1alpha and HNF-4alpha genes in Type 1 diabetes mellitus (T1D) patients without pancreatic autoantibodies. DESIGN AND METHODS: Twenty patients without pancreatic autoantibodies (Ab neg) and 20 with autoantibodies (Ab pos), age/gender matched, were included (age 17-34 years). Islet cell, glutamic acid decarboxylase, tyrosine phosphatase and insulin autoantibodies, basal and stimulated C-peptide were measured. HLA-DRB1-DQA1-DQB1 typing and screening for mutations in the HNF-1alpha and HNF-4alpha genes were performed. RESULTS: No differences were found in clinical presentation, metabolic control and beta-cell function in the two groups (onset or after 12 months). DRB1*0301-DQA1*0501-DQB1*0201 was the most frequent haplotype in both groups but we found a higher proportion of protective T1D haplotypes and Asp(beta57) in the Ab neg group, but in all the cases in combination with susceptible T1D haplotypes. We found two previously reported polymorphisms (HNF-1alpha, Ala98Val; HNF-4alpha, Thr130Ile) in Ab neg and a new variant (Ser165Gly) in the HNF-4alpha gene in an Ab pos subject. Conclusions In a non-paediatric population with newly diagnosed T1D, the absence of islet antibodies does not imply clinical or metabolic differences when compared with those cases with islet antibodies. Despite a similar HLA-DR/DQ typing, the presence of protective alleles and molecular properties in a higher proportion in the Ab neg group suggests that these factors could modulate the presence or absence of islet antibodies. Variants in HNF-1alpha and HNF-4alpha are unlikely to be major contributors to the pathogenesis of diabetes in antibody-negative T1D. PMID- 15660730 TI - Increasing body weight predicts the earlier onset of insulin-dependant diabetes in childhood: testing the 'accelerator hypothesis' (2). AB - AIMS AND METHODS: It has recently been hypothesized that weight gain in childhood accelerates the onset of Type 1 diabetes, as well as increasing its risk, and that Type 1 diabetes and Type 2 diabetes may be one and the same disorder of insulin resistance. An explanation is needed for the rising incidence of childhood diabetes and, to test the Accelerator Hypothesis, we examined the anthropometric measurements recorded from birth in 168 young people presenting with Type 1 diabetes between 1980 and 2002. Pre-onset as well as peri- and post onset measurements of height and weight were available, and waist circumference was recorded at various intervals after onset. RESULTS: The mean birth weight of the children and their height, weight and body mass index (BMI) at diagnosis lay close to the population mean. However, pre-onset and post-onset BMI were both well above the population mean, were closely correlated with each other (r = 0.79, P < 0.001) and (inversely) with age at onset (r = -0.30, P < 0.001). A significant correlation was also found between BMI standard deviation scores (sds) and year of diagnosis (r = 0.27, P < 0.001) and, importantly, waist circumference sds in the children with Type 1 diabetes was found to be substantially greater than average for the population [boys: +0.96 (sd 1.04), girls: +1.30 (sd 0.89)]. CONCLUSIONS: The data suggest that children with Type 1 diabetes have become progressively heavier at diagnosis over the past 20 years, and that the heavier child develops it earlier. Waist circumference, a proxy for visceral fat mass and insulin resistance, is substantially greater in children with Type 1 diabetes. Weight centile crossing appears to be an important environmental accelerator which may contribute to or account for the striking increase in both Type 1 diabetes and Type 2 diabetes in childhood. A reduction of body weight and improved lifestyle might reverse this trend in both types of diabetes. PMID- 15660731 TI - Psychiatric morbidity and health outcome in Type 1 diabetes--perspectives from a prospective longitudinal study. AB - AIMS: To describe psychiatric status and relationship to metabolic control in adolescents with Type 1 diabetes studied prospectively from diagnosis. METHODS: Adolescents (n = 41) completed a self-report measure of psychiatric status 10 years after disease onset. Metabolic control information was recorded prospectively from diagnosis. The rate and type of psychiatric disorder were determined and the relationship between mental health status and metabolic control history examined. RESULTS: Thirty-seven per cent of the adolescents met criteria for a DSM-IV psychiatric disorder, two to three times higher than community levels of psychiatric morbidity. Females were significantly more likely to receive a diagnoses (chi2 = 4.98, P < 0.05). Two thirds of participants had experienced at least one serious hypoglycaemic episode and one third had a history of chronic poor metabolic control. DSM-IV diagnoses were present in half of those with a history of chronic poor control, one third of the hypoglycaemia group and one quarter of well controlled participants. Adolescents with a current Mood (t = -2.83, P < 0. 01), Anxiety (t = -3.77, P = 0.001) or Behaviour (t = 2.56, P < 0.05) disorder and those with a history of poorly controlled diabetes (F (2,29) = 5.4, P = 0.01) had higher externalizing behaviour problem scores at diagnosis than those without current disorder. CONCLUSIONS: Adolescents with Type 1 diabetes are at high risk for psychiatric disorder. Poorly controlled diabetes over the first 10 years of illness was associated with pre-existing behaviour problems at diagnosis and there was a trend for an association with current psychiatric status. PMID- 15660732 TI - Effects of nateglinide on myocardial microvascular reactivity in Type 2 diabetes mellitus--a randomized study using positron emission tomography. AB - AIMS: To evaluate effects of the oral antidiabetic insulinotropic agent nateglinide on myocardial blood flow (MBF) and microvascular reactivity in Type 2 diabetic patients. METHODS: Forty-seven Type 2 diabetic patients were randomly assigned 2 : 1 to nateglinide 120 mg (t.i.d., n = 33) or placebo (n = 14). At baseline and after 16 weeks of treatment, MBF was quantified using positron emission tomography with N-13 ammonia at rest, during endothelial-dependent stimulation by cold pressor test and during adenosine-mediated vasodilation. Additional blood samples were taken to assess glycaemic control and lipid profile. RESULTS: MBF at rest and during adenosine did not change during the study. The percentage of flow increase from rest during cold pressor test did not improve significantly in the nateglinide group vs. placebo (from 26.1 +/- 37.2% to 29.1 +/- 27.8% between week 0 to week 16 for nateglinide vs. 14.9 +/- 37.1% to 18.1 +/- 28.4% for placebo; P = 0.07 for nateglinide when adjusted for higher baseline values). Nateglinide decreased HbA1c by 0.4% (from 7.6 +/- 0.9% to 7.2 +/- 1.3%) compared to an increase of 0.5% in the placebo group (from 7.9 +/- 0.8% to 8.4 +/- 1.7%; P = 0.02 for nateglinide). No differences between the two groups were observed in insulin levels and lipid status. CONCLUSIONS: Nateglinide neither improved, nor impaired myocardial blood flow in Type 2 diabetic patients. Potential effects on endothelial-dependent myocardial blood flow remain to be investigated further. Positron emission tomography is a sensitive approach to assess the effects of therapeutic agents on myocardial blood flow in patients with diabetes. PMID- 15660733 TI - Predicting vascular risk in Type 1 diabetes: stratification in a hospital based population in Scotland. AB - AIMS: To estimate the absolute cardiovascular risk of patients with Type 1 diabetes attending hospital diabetes clinics in Scotland and to develop a method for identifying those at highest risk, thus enabling therapy to be targeted. METHODS: Baseline information was collected for 2136 patients with Type 1 diabetes using the Royal College of Physicians of Edinburgh Diabetes Register. These records were then linked to diagnoses of macrovascular disease in databases of the Information Statistics Division of the Common Services Agency. RESULTS: During six to nine years of follow up 110 patients (5%) developed macrovascular disease. There were significant associations between baseline age (P < 0.00001), blood pressure (P < 0.00001), albuminuria (P < 0.0002), HbA1c (P < 0.001), cholesterol (P < 0.00001) and smoking status (P < 0.00001) with the development of macrovascular disease. A scoring system for future macrovascular risk was developed from a multivariate analysis of this data. CONCLUSIONS: The data confirm the high vascular risk of patients with Type 1 diabetes. The relationship with age is such that those patients above 50 years require only one additional risk factor to reach such a high vascular risk that intervention is indicated. Using these data many patients between the age of 40 and 49 years are also likely to be identified to be at high risk. PMID- 15660734 TI - Healing diabetic neuropathic foot ulcers: are we getting better? AB - AIM: To benchmark by year the likelihood that an individual with a diabetic neuropathic foot ulcer will heal over more than a 10-year period. PATIENTS AND METHODS: A cohort study within a multicentre wound care network of individuals with a diabetic neuropathic foot ulcer who were treated by a standard wound care algorithm. The main outcome was a healed wound by the 20th week of care stratified by calendar year. RESULTS: We evaluated 27 193 individuals with a neuropathic foot ulcer. Between 1988 and 1990 approximately 66% of patients did not heal. By 1999 this percentage had decreased to 49%. The change in the rate of failure to heal is very closely associated with an increase over time in the proportion of patients seen with wounds identified as prognostically favourable using a previously published prognostic model (i.e. individuals with wounds < or = 2 cm2, wounds < or = 2 months old, and wounds of grade < or = 2). Nevertheless, even among those most likely to heal, the likelihood of failing to heal went from 62% prior to 1991 to 32% in 2000. CONCLUSIONS: We have shown that individuals with a diabetic neuropathic foot ulcer seeking care are more likely to heal today than 10 years ago. The primary reason for this improvement is that individuals are seeking care when their wounds are most easily treated and these are now more likely to heal. PMID- 15660735 TI - In-hospital breast feeding rates among women with gestational diabetes and pregestational Type 2 diabetes in South Auckland. AB - AIM: To describe the uptake of breast feeding in mothers with either Type 2 diabetes or gestational diabetes (GDM) in a hospital serving a multiethnic community in South Auckland, New Zealand. RESEARCH DESIGN AND METHODS: A retrospective study of all women attending the Diabetes in Pregnancy clinic over a 4-year period was undertaken: 30 women had Type 2 diabetes and 373 GDM. RESULTS: Compared with mothers with GDM, mothers with Type 2 diabetes were less likely to breast feed in any way as the first feed (41.4% vs. 68.0%, P = 0.011) or at discharge (69.0% vs. 84.0%, P = 0.039). In the combined group, there were no differences in uptake of breast feeding by ethnicity, age, parity, body mass index, smoking or antenatal glycaemia, use of insulin or presence of hypertension. Breast feeding on discharge was associated with a higher APGAR score, breast feeding as the first feed (78.2% vs. 19.4%, P < 0.001) and lower rates of delivery by Caesarean section (17.0% vs. 31.8%, P = 0.006). Logistic regression showed breast feeding as the first feed, the major determinant for breast feeding on discharge. CONCLUSIONS: Factors delaying breast feeding as the first feed are the major determinant of breast feeding on discharge. Strategies to increase breast feeding as the first feed among women with Type 2 diabetes, and those having a Caesarean section, may be useful in increasing the uptake of breast feeding in the longer term. PMID- 15660736 TI - Unchanged incidence of diabetic nephropathy in Type 1 diabetes: a nation-wide study in Iceland. AB - AIMS: Diabetic nephropathy is an uncommon cause of end-stage renal disease in Iceland in contrast to most industrialized countries. The aim of this study was to examine the incidence of diabetic nephropathy in Iceland. METHODS: All patients diagnosed with Type 1 diabetes in Iceland before 1992 were studied retrospectively. Patients diagnosed before age 30, who were insulin dependent from the onset, were defined as having Type 1 diabetes. Diabetic nephropathy was defined as persistent proteinuria measured with a dipstick test (Albustix) on three consecutive clinic visits at least 2 months apart. Patients were followed to the end of year 1998, to their last recorded outpatient visit, or until death. The cumulative incidence of diabetic nephropathy was calculated with the Kaplan Meier method and presented according to the duration of diabetes divided into 5 year intervals. RESULTS: A total of 343 patients with Type 1 diabetes were identified. The mean follow-up period was 20.2 +/- 11.4 (mean +/- sd) years. Only 9.3% of patients were lost to follow-up. Sixty-five patients developed diabetic nephropathy. The cumulative incidence was 22.6% at 20 years and levelled off at 40.3% after approximately 35 years of diabetes duration. No significant changes in cumulative incidence were observed over time. Mean glycated haemoglobin was 8.4% in patients with proteinuria and 7.8% in a group of patients without proteinuria that was matched for age, gender and duration of diabetes (P = 0.04). CONCLUSIONS: The cumulative incidence of diabetic nephropathy in Iceland is comparable with previously reported cumulative incidence rates and has remained unchanged. Glycaemic control was significantly better in patients without proteinuria. PMID- 15660737 TI - NT-proBNP in normoalbuminuric patients with Type 2 diabetes mellitus. AB - OBJECTIVE: To examine levels of NT-proBNP and its relation to hypertension, as well as diastolic function in normoalbuminuric patients with Type 2 diabetes. RESEARCH DESIGN AND METHODS: The study comprised 60 Type 2 diabetic patients without albuminuria. Thirty patients were normotensive and 30 had hypertension. Exclusion criteria were cardiac symptoms and an ejection fraction < 55%. Thirty age- and sex-matched normal subjects served as controls. Diastolic dysfunction was assessed with echocardiography, by means of mitral inflow and colour M-Mode flow propagation recordings. RESULTS: Overall NT-proBNP was significantly elevated in the Type 2 diabetes group, compared with the controls [54.5 pg/ml (5 162) vs. 32.7 pg/ml (5-74.3) P = 0.02]. NT-proBNP was significantly higher among hypertensive patients compared with both normotensive patients and controls but no difference was found between the normotensive patients and the controls [58.0 pg/ml (8.5-162), P < 0.05 vs. 50.8 pg/ml (5-131) P = 0.4]. Patients with concentric and eccentric hypertrophy had significantly higher NT-proBNP levels compared with the control group [81.0 pg/ml (5-147), P < 0.001 and 66.8 pg/ml (42 128), P < 0.001], whereas patients with left ventricular remodelling (enlarged relative wall diameter but normal left ventricular mass) were comparable with the control group [42.3 pg/ml (8.3-142) P = 0.55]. Patients with left atrial enlargement also had incremental NT-proBNP values. NT-proBNP was only moderately correlated to age (r = 0.33, P < 0.05) and left ventricular diastolic diameter (r = 0.41, P < 0.05), but unrelated to diastolic function. CONCLUSIONS: NT-proBNP is significantly increased in hypertensive, normoalbuminuric patients with Type 2 diabetes. These findings were related to left ventricular hypertrophy and increased left atrial and ventricular diameters. PMID- 15660738 TI - Interclinician variation in diabetes foot assessment--a national lottery? AB - AIM: The aim was to evaluate variation among clinicians in the outcome of assessments of foot health status and risk status in patients with diabetes. METHODS: Seventeen clinicians assessed three patients with diabetes using a standardized assessment form and risk classification system. RESULTS: There was variation among clinicians in all aspects of the assessment; recording basic demographic information; taking a medical history; vascular and neurological assessments. Variation was also evident in the risk categories allocated to each of the three patients. CONCLUSIONS: As a consequence of the variation among clinicians in the foot assessment the same patient would have received different care pathways to monitor and manage their foot health depending upon which clinician undertook their initial assessment. We therefore recommend that more attention is placed on training for objective clinical testing, at both pre- and postgraduate levels. PMID- 15660739 TI - Parental age at delivery, birth order, birth weight and gestational age are associated with the risk of childhood Type 1 diabetes: a UK regional retrospective cohort study. AB - AIMS: To investigate perinatal risk factors for childhood Type 1 diabetes in a UK population cohort. METHODS: Perinatal data have been routinely recorded in Northern Ireland for all births in the period 1971-86 (n = 447 663). Diabetes status at the age of 15 years was ascertained in this cohort by identifying 991 children from 1079 registered with Type 1 diabetes diagnosed from 1971 to 2001 and date of birth in the period 1971-86. RESULTS: Increased Type 1 diabetes risk was associated with higher maternal age, paternal age, birth weight and birth weight for gestational and lower gestational age. After adjustment for maternal age, the association between Type 1 diabetes and paternal age remained significant [relative risk (RR) = 1.52 (1.10, 2.09) comparing father's age 35 years or more to less than 25 years] but not vice versa [RR = 1.11 (0.80, 1.54) comparing mother's age 35 years or more to less than 25 years]. Increased birth order was associated with a significant decrease in the risk of Type 1 diabetes [adjusted RR = 0.75 (0.62, 0.90) comparing birth order three or more with firstborn], but this only became apparent when adjustment was made for maternal age. Furthermore this association with birth order was significant only for diabetes diagnosed under the age of 5 years. CONCLUSIONS: Our analysis demonstrates, for the first time in a UK regional cohort setting, that maternal age and paternal age at delivery, birth order, birth weight and gestational age are significantly associated with Type 1 diabetes risk. PMID- 15660740 TI - Controlling the diabetes epidemic: how should we screen for undiagnosed diabetes and dysglycaemia? AB - AIMS: To compare the detection of undiagnosed diabetes and dysglycaemia (impaired glucose tolerance, impaired fasting glucose, diabetes) using risk factors and laboratory measures of glycaemia. METHODS: Casual blood glucose samples were taken from 1899 (69.4% of 2737 invited) European, Maori and Pacific Islands subjects aged 40-79 years from randomly selected households in South Auckland, New Zealand. Of these, 534 attended for a 75-g oral glucose tolerance test (OGTT) if an elevated result was identified [327/478 (68.4%)] or if randomly selected with a 'normal' screening result [207/308 (67.2%)]. RESULTS: Several Europeans with undiagnosed diabetes (25.0%) and dysglycaemia (31.4%) had no diabetes risk factors. Most Maori and Pacific Islanders had at least one risk factor. The area under the receiver operating curve (ROC) for the detection of undiagnosed diabetes was 0.92 (0.89-0.95) using fasting glucose, 0.86 (0.82-0.90) using HbA1c, 0.75 (0.69-0.80) using random glucose, but 0.60 (0.55-0.66) using risk factor screening. The ROC for detecting any dysglycaemia was 0.88 (0.85-0.90), 0.68 (0.64-0.71), 0.72 (0.69-0.75), 0.61 (0.58-0.65), respectively. Screening using fasting glucose (the best test) detected 90.4% of new diabetes and 78.4% of dysglycaemia; risk factor screening followed by fasting glucose detected significantly less cases [88 (82-93)% and 86 (82-89)%, respectively] with 9.2% less OGTTs. CONCLUSIONS: Using risk factors for the identification of who should receive a blood test for dysglycaemia adds little to direct screening with the risk of missing some with significant hyperglycaemia. Screening for dysglycaemia may best be undertaken using blood tests without initial risk factor symptom screening. PMID- 15660741 TI - Glycaemic and insulin responses of six traditional Sudanese carbohydrate-rich meals in subjects with Type 2 diabetes mellitus. AB - AIMS: The objective of this study was to investigate the influence of six Sudanese traditional carbohydrate-rich meals on glucose and insulin responses and to formulate appropriate dietary guidelines based on glycaemic excursions of traditional foods. METHODS: On six occasions with 1-week intervals, 10 Type 2 diabetic subjects consumed six Sudanese traditional carbohydrate-rich meals. The following meals were tested and also analysed for their composition: wheat gorasa (pancakes), sorghum kisra (flat bread) and sorghum acida (porridge), millet kisra and millet acida and maize acida. Blood samples were collected before and after meal ingestion at 0, 30, 60, 120 and 240 min and analysed for plasma glucose and plasma insulin and incremental areas under the curves (AUC) were calculated. RESULTS: A significant variation in AUC for glucose and insulin responses were found between meals, the over all differences in incremental AUCs between the six meals were significant for both plasma glucose (P = 0.0092) and insulin (P = 0.0001). The 2-h glucose values were 10.5 +/- 2.7 for sorghum flatbread, 9.5 +/- 3.1 for sorghum porridge, 10.3 +/- 3.4 for millet flatbread, 10.6 +/- 3.6 for millet porridge, 11.4 +/- 2.7 for maize porridge and 8.7 +/- 2.4 for the wheat pancakes. The comparison between the AUCs of the meals showed that millet acida (porridge) followed by wheat gorasa (pancakes) displayed significantly lower post prandial glucose and insulin responses, whereas maize acida induced a higher post prandial glucose and insulin response. CONCLUSIONS: The comparison of glycaemic and insulin responses to six traditional Sudanese meals show differences of clinical importance, and could form a basis for dietary advice to diabetic subjects in Sudan and countries sharing similar food traditions. PMID- 15660742 TI - 'Disappearing diabetes'--resolution of apparent Type 1 diabetes in a patient with AIDS and cytomegalovirus (CMV) infection. AB - A 30-year-old African female with established acquired immunodeficiency syndrome (AIDS) and no history of diabetes, presented in severe diabetic ketoacidosis (DKA). Blood pH was 6.96, serum bicarbonate 5 mmol/l, plasma glucose (PG) 33.0 mmol/l, and urine heavily positive for ketones. She responded to standard treatment and was established on twice-daily subcutaneous insulin. Four months later her insulin was stopped because of hypoglycaemic attacks on small doses. A glucose tolerance test (GTT) at 6 months postdiagnosis was normal (fasting PG 4.4 mmol/l and 2 h PG 7.5 mmol/l), and at 12 months random PG was 4.1 mmol/l and HbA1c 4.3%. The onset of her apparent Type 1 diabetes coincided with an HIV associated cytomegalovirus (CMV) infection, and a reversible 'CMV insulitis' may be an explanation. Alternatively, the patient may have had what has recently been described as 'atypical diabetes' in African or Afro-Caribbean diabetic patients. Here resolution of diabetes may occur after presentation, though complete return to normoglycaemia after true DKA is very unusual. PMID- 15660743 TI - Near patient blood ketone measurements and their utility in predicting diabetic ketoacidosis. AB - AIM: To assess the utility of near patient blood ketone measurements in predicting diabetic ketoacidosis (DKA) among a group of hyperglycaemic unwell patients presenting to a hospital emergency department. METHODS: Near patient blood beta-hydroxybutyrate (beta-OHB) testing has recently been introduced as a new tool in our hospital Accident and Emergency department (A&E) for patients with a finger-prick glucose of > 11 mmol/l. We reviewed the records of the first 50 patients to have a beta-OHB measurement to establish if they developed DKA or received treatment with intravenous insulin within 48 h of presentation. We then compared the diagnostic power of beta-OHB measurements with other clinical, physiological and biochemical markers of DKA. RESULTS: Nine patients had DKA, eight had a compensated metabolic acidosis secondary to raised serum ketones, and 33 had no evidence of DKA during the following 48 h. The median (range) beta-OHB levels in each group were 6.0 (3.1-6.0) mmol/l, 3.4 (1.2-5.7) mmol/l, and 0.1 (0.0-1.2) mmol/l, respectively. A beta-OHB level of > or = 3.0 mmol/l had a sensitivity of 100% and specificity of 88% for DKA. All those with beta-OHB level > 3.0 mmol/l required treatment with intravenous insulin. CONCLUSION: Measuring beta-OHB when a hyperglycaemic patient is identified could offer a simple method of identifying at an early stage those patients at highest risk of DKA (beta-OHB > 3.0 mmol/l), and redirecting the search for a diagnosis in others (beta-OHB < 1.0 mmol/l). PMID- 15660744 TI - Insulin resistance determines efficacy of glimepiride in Type 2 diabetic patients not well controlled by diet alone. PMID- 15660745 TI - Regarding the use of HOMA to aid drug selection. PMID- 15660746 TI - A deletion polymorphism of the alpha2B-adrenergic receptor gene is not associated with late complications in Type 1 diabetic patients. PMID- 15660747 TI - Framingham risk equations underestimate coronary heart disease risk in diabetes. PMID- 15660748 TI - Apexification: a review. AB - This paper reviews the rationale and techniques for treatment of the non-vital immature tooth. The importance of careful case assessment and accurate pulpal diagnosis in the treatment of immature teeth with pulpal injury cannot be overemphasized. The treatment of choice for necrotic teeth is apexification, which is induction of apical closure to produce more favorable conditions for conventional root canal filling. The most commonly advocated medicament is calcium hydroxide, although recently considerable interest has been expressed in the use of mineral trioxide aggregate. Introduction of techniques for one-visit apexification provide an alternative treatment option in these cases. Success rates for calcium hydroxide apexification are high although risks such as reinfection and tooth fracture exist. Prospective clinical trials comparing this and one-visit apexification techniques are required. PMID- 15660749 TI - Prevalence of oral trauma in children with bilateral clefts. AB - The main object of this study is to analyze the prevalence of oral trauma in subjects with complete bilateral clefts, with anterior projection of the premaxilla. A total of 106 children aging 6 months to 9 years were analyzed. The caretakers answered a specific questionnaire, in order to report the presence or absence of trauma to the soft and/or hard tissues of the child's mouth. Whenever there was a history of trauma, the patients were submitted to clinical examination. The prevalence of oral trauma was 53%, being 91% of soft tissue lesions, 8.9% of avulsion, 7% of luxation and 1.8% of intrusion. For the males, the prevalence was 56% and for the females it was 47%, with no statistical significance. Regarding the following aspects:period of time spent with the parents and at school, and presence or absence of siblings, no statistical difference could be found. Among the traumatized individuals, 80% aged less than 3 years by the moment of the trauma, 89% suffered the trauma at home, 75% presented lesions in the soft tissue at the premaxilla, 16% in the maxillary incisors, and 8.9% presented lesions in both structures. It was noticed that 45% of the permanent incisors that succeeded the traumatized deciduous teeth presented alterations, being 48% of structure and 52% of structure and position. The prevalence of trauma in this sample was superior to that observed in the literature, without any positive associations between the evaluated aspects. These results suggest that the projection of the premaxilla brings about a higher risk of oral trauma around this area. PMID- 15660750 TI - Have the causes of maxillofacial fractures changed over the last 16 years in Finland? An epidemiological study of 725 fractures. AB - A retrospective study was undertaken to assess causes of maxillofacial fractures in Helsinki in 1981 and 1997. Hospital records of 725 patients were analyzed according to several factors including age, sex, cause of fracture and time of the injury. The time intervals between the accident and hospital examination were also evaluated. Number of maxillofacial fractures was 318 in 1981 and 407 in 1997 (27.9% increase) and most patients were men. The male to female ratio was 2.8:1 in 1981, 3:1 in 1997. In 1981, most affected patients were in the age group of 31 40 years (33.2% of men, 28.9% of women). Sixteen years later the most affected age group was 41-50 years (23.3% of men, 30.4% of women). Assault was the cause of the injury in 42% of patients followed by traffic accidents (26%) and fall (17%). During the study period violence had become more severe in nature. Kicking as the cause of maxillofacial fracture increased by 7.3% and use of a weapon by 5.7% between the years studied. Bicycle accidents increased by 19.3% but motor vehicle accidents decreased by 31.6% between the years. Falls, and bicycle and pedestrian accidents were the causes that accounted for most of the increase in maxillofacial fracture. In 1997, maxillofacial fractures were slightly more common from June to August and from Friday to Sunday than at other times (45.2 and 50%, respectively). PMID- 15660751 TI - Traumatic injuries of the permanent incisors in children in southern Turkey: a retrospective study. AB - The aim of this study is to investigate the incidence, etiologic factors and results of dental trauma and the effects of age and gender on the trauma in permanent incisors. Over a 3-year period, 514 permanent incisor teeth in 317 patients with trauma history, who applied to Suleyman Demirel University School of Dentistry, Department of Pedodontics from the southern cities of Turkey, were evaluated. Standardized trauma forms were filled for each patient. In all age groups, the most frequent cause of trauma was found to be unspecified falls (47.6%). Maxillary teeth (88.5%) and central incisors (87.5%) were the most affected teeth from dental trauma. Ellis class II crown fracture was the most frequently seen type of injury (43.8%). The percent of the patients who applied to a dental clinic in the first 3 days after the trauma occurred (22.8%) was less than the percent of the patients who applied after 3 months and more time period (45.1%). It reveals that it is important to inform the public about dental trauma and the importance of time in these cases. PMID- 15660753 TI - Dental trauma and mouthguard usage among ice hockey players in Turkey premier league. AB - An epidemiological survey was carried out among the ice hockey players of Turkish Premier Ice Hockey League. The purpose of the present study was to evaluate the prevalence of dental trauma in youth and adult ice hockey players, as well as to check if the players were aware of mouthguards and had used any. The players were interviewed to determine the occurrence of dental trauma during ice hockey and mouthguard usage level. Results revealed that awareness of Turkish ice hockey players to dental trauma is neglected while the majority of players also demonstrated limited utilization of mouthguard. PMID- 15660752 TI - Factors affecting coronal fracture of anterior teeth in North Jordanian children. AB - The aims of this study were to investigate the incidence of coronal fracture of the anterior teeth in North Jordanian schoolchildren and to study the main predisposing factors and the factors that may affect the severity of this fracture. A study group (958) comprising schoolchildren aged 13-15 years was chosen by a simple random method from five geographical areas in Irbid Governate, Jordan. All children completed a questionnaire related to history of trauma to their anterior teeth before they had a clinical examination for lip competence, lip line and amount of fracture. Overjet was recorded from a study cast made for each student. Statistical analysis was performed using chi-square test. The results showed a prevalence of 11% of coronal fracture with female-male ratio of 1:1. An increase in the overjet more than 3 mm doubled the incidence of coronal fracture while overjet more than 6 mm increased the incidence fourfold. There was higher incidence of coronal fracture associated with lip incompetence and low lip line (P <0.01). The severity of fracture increased in children with a larger overjet (P <0.001). It was concluded that overjet, lip competence and lip line were important predisposing factors to coronal fracture of the anterior teeth while the severity of the fracture was mainly determined by overjet. PMID- 15660754 TI - High resolution charge-coupled device sensor vs. medium resolution photostimulable phosphor plate digital receptors for detection of root fractures in vitro. AB - The aim of this study was to compare the diagnostic accuracy of a high resolution charge-coupled device (CCD) sensor and a medium resolution photostimulable phosphor (PSP) plate for detecting experimentally induced root fractures and further, to evaluate differences between images taken with various horizontal and vertical angles. Forty-seven extracted single-rooted human teeth mounted in a dry human skull were used in the experiment. The teeth were radiographed, before and after root fractures were induced, with two digital receptors: the Digora PSP system (approx. 8 lp mm(-1)) and the RVG-ui(TM), a CCD sensor with a high resolution mode (15-20 lp mm(-1)). Four images were taken with each of the receptors of each tooth: one orthogonal exposure (O-images), one exposure with a vertical angle of 15 degrees by which the root was imaged elongated (L-images), and two eccentric exposures with a horizontal angle of 15 degrees mesially and distally. Three observers marked a fracture line if detected, in each image. Three sessions were held, one assessing the O-images, one the L-images, and one in which all four images of the same tooth were displayed simultaneously (X images). The RVG-ui images obtained higher sensitivities than the Digora PSP images (P <0.05). Sensitivity was statistically significantly higher for the X images than for both the O-images and the L-images (P <0.05). Based on the observed means, specificities were significantly different neither between the angles, nor between the images from the two digital systems taken with the same angle (P >0.05). It may be speculated that the difference in spatial resolution between the two digital systems accounts for the differences in their sensitivity. PMID- 15660755 TI - Possible application of transmitted laser light for the assessment of human pulp vitality. Part 2. Increased laser power for enhanced detection of pulpal blood flow. AB - The purpose of this study was to investigate the effect of an increase in laser power on the transmitted laser signals from vital and non-vital teeth, in the hope of achieving a better assessment of human pulp vitality with the transmitted laser-light flowmeter. The experiments were carried out on total of 61 vital teeth with no restoration (19 upper central incisors, 16 upper lateral incisors, 16 upper canines, and 10 first premolars) and five non-vital upper central incisors (the root canals of which were filled with gutta-percha) in 15 subjects aged 22-28 years. For use with transmitted laser light, the fibers within the probe of a conventional laser Doppler flowmeter (LDF) apparatus were used, one for transmitting light onto the labial surface, the other for receiving it at the palatal surface of the same tooth, as reported previously. Laser output power was set at the original 2 mW and also at 5, 7, and 10 mW. The number of vital teeth displaying a blood flow (BF) signal at each laser power setting was: 1) 12/19 central incisors at 2 mW, 19/19 at 5, 7, and 10 mW, 2) 19/19 lateral incisors at 2, 5, 7, and 10 mW, 3) 0/16 canines at 2 mW, but eight, 12, and 14 at 5, 7, and 10 mW, 4) 0/10 first premolars at 2, 5, 7, and 10 mW. Thus, an increase in laser power increased BF detection from the thicker teeth (but not from premolars). In addition, clearer BF signals synchronized with heart rate, and greater passive BF changes secondary to blood pressure (BP) changes were observed at higher laser settings. In non-vital teeth, no signals synchronized with heart rate or BP changes were observed, indicating that no BF signal of non-pulpal origin was ever monitored with this ballistic light even when the laser power was increased. These results indicate that high-powered transmitted laser light could be a useful tool both for monitoring pulpal BF and for the assessment of tooth-pulp vitality. PMID- 15660756 TI - Effect of electrophoretically activated calcium hydroxide on bacterial viability in dentinal tubules--in vitro. AB - To evaluate the ability of electrophoretically activated calcium hydroxide (CH) to eliminate bacteria in dentinal tubules. In an in vitro model of dentinal tubule infection, 18 cylindrical root specimens prepared from freshly extracted bovine teeth were used. After removal of the smear layer, intracanal dentinal tubules were infected with Enterococcus faecalis for 21 days. In 12 specimens, CH paste was placed in the root canals for 7 days. In six of these, an electrophoretic current (10 mA per 10 min), using two electrodes, was applied after placing the medicament in the canal. Powder dentin samples obtained from within the canal lumina using ISO 025, 027, 029, 031 and 033 burs were examined for the presence of vital bacteria by inoculating agar plates and counting colony forming units. anova with repeated measures was used to analyze results. A significant difference was found between experimental groups and the positive control group. CH and electrophoretically activated CH significantly (P <0.001) reduced bacterial viabilities in dentinal tubules to a depth of 200 microm. Treatment with electrophoresis was significantly (P <0.001) more effective than pure CH in depths of 200-500 microm. Specimens treated with electrophoretically activated CH showed no viable bacteria in dentinal tubules to a depth of 500 microm from the root canal space within 7 days. The time required for treatment of pulpal infection root resorption may be decreased, thus minimizing the risk of coronal fractures in young patients with traumatized teeth. PMID- 15660757 TI - Emdogain does not prevent progressive root resorption after replantation of avulsed teeth: a clinical study. AB - Emdogain has been shown in clinical and experimental studies to promote regeneration of all periodontal tissues: cementum with anchoring fibres, a functional, periodontal ligament and alveolar bone in connection with treatment of marginal periodontitis. The intention of this study was to analyse whether this regenerative capacity upon the periodontal ligament also worked in a trauma situation where a significant number of PDL cells have been eliminated because of unphysiologic storage or actual damage during avulsion or replantation. Furthermore if ankylosis sites already established because of earlier replantation after avulsion could be surgical removed and application of Emdogain could revert the ankylosis stage to a normal PDL situation. The first treatment situation was tested in seven patients with a total of 16 avulsed teeth with varying time of extra oral storage. The teeth were extra-orally endodontically treated and the root and socket covered with Emdogain before replantation. All teeth demonstrated subsequent ankylosis, primarily diagnosed by a percussion test. The second treatment situation where an ankylosis was already established constituted of seven patients with a total of 11 teeth because of previous replantation after avulsion. These teeth were all extracted, the ankylosis sites removed and the root and socket treated with Emdogain. After 6 months all teeth showed recurrence of ankylosis. It is concluded that Emdogain was not able to prevent or cure ankylosis. PMID- 15660758 TI - Treatment of avulsed teeth with Emdogain--a case report. AB - The present case report describes the reimplantation of avulsed teeth with the treatment of Emdogain. Case was avulsed right maxillary permanent central and lateral incisor in a 9-year-old girl suffering from a traumatic injury. After pretreatment of avulsed teeth, Emdogain was applied to the root surface and into the extraction socket with subsequent replantation of the tooth. Evaluation parameters included horizontal and vertical percussion sound and periapical radiographs. At 1-2-6-12-month follow-up period, the clinical and radiographic appearance of the teeth showed resolution of mobility and no signs of replacement resorbption. PMID- 15660759 TI - Treatment of a severe dental lateral luxation associated with extrusion in an 8 month-old baby: a conservative approach. AB - This case report documents the trauma and follow-up care of lateral luxation associated with extrusion of the lower central incisors in an 8-month-old patient. The teeth were repositioned by digital pressure and stabilized using proximal sutures. Clinical and radiographic follow-up 40 months after the injury showed alterations in both incisors, but both remained functional and free of pathology. PMID- 15660760 TI - Conservative treatment of paediatric mandibular fracture by the use of orthodontic appliance and rubber elastics: report of a case. AB - Treatment principles of paediatric mandibular fractures may differ from the treatment of the adult population in that a conservative approach is in most cases advocated before the use of internal rigid fixation with plates and screws. This is because of a relative high risk of disturbed facial skeletal growth and risk of damaging unerupted teeth. Knowledge of conservative treatment options is essential in order to minimize these risks and one option is presented in this paper. This case report describes a 5-year-old girl that sustained an open fracture of the mandible and who was successfully treated by the means of applying orthodontic brackets and an arch bar combined with ligatures and rubber elastics. PMID- 15660761 TI - Adhesive fragment reattachment after orthodontic extrusion: a case report. AB - In the treatment of crown fractures, adhesive fragment reattachment provides a good alternative to other restorative techniques, offering several advantages. The present paper reports a case in which the treatment of a cervical crown fracture was accomplished by reattaching the tooth fragment with a flowable resin composite. Orthodontic root extrusion was performed with a modified Hawley appliance prior to fragment reattachment. The clinical and radiographic results after 2.5 years were successful. PMID- 15660762 TI - Preemptive low-frequency stimulation decreases the incidence of amygdala-kindled seizures. AB - PURPOSE: The use of electrical stimulation as a therapy for epilepsy is currently being studied in experimental animals and in patients with epilepsy. This study examined the effect of preemptive, low-frequency, 1-Hz sine wave stimulation (LFS) on the incidence of amygdala-kindled seizures in the rat. METHODS: Electrodes were implanted into the basolateral amygdalae of adult male rats. All animals received a kindling stimulus of 60-Hz, 400-microA, sine wave for 1 s twice a day. Experimental animals received an additional LFS consisting of 1 Hz, 50 microA for 30 s immediately before the kindling stimulus. Afterdischarge (AD) duration, behavioral seizure score, the number of stimulations required to elicit the first stage five seizure and to become fully kindled were measured. After 20 stimulations, a crossover procedure was performed. Fully kindled rats from each group were switched, so that the original controls received LFS plus the kindling stimulus, and the original experimental rats received only the kindling stimulus. RESULTS: During kindling acquisition, LFS induced a significant decrease in AD duration. A significant increase in the number of times the kindling stimulus failed to elicit an AD was noted. Control rats exhibited an AD 99% of the time compared with 70% in experimental rats (p < 0.0001; Fisher's Exact test). In fully kindled animals, the incidence of stage five seizures in the original controls significantly decreased from 98% to 42% (p < 0.0001) when the LFS was added to the kindling paradigm. CONCLUSIONS: The dramatic decrease in the incidence of stage 5 seizures in fully kindled animals after preemptive LFS strongly suggests that LFS may be an effective therapy for the prevention of seizures in patients with epilepsy. PMID- 15660763 TI - Use of chronic epilepsy models in antiepileptic drug discovery: the effect of topiramate on spontaneous motor seizures in rats with kainate-induced epilepsy. AB - PURPOSE: Potential antiepileptic drugs (AEDs) are typically screened on acute seizures in normal animals, such as those induced in the maximal electroshock and pentylenetet-razole models. As a proof-of-principle test, the present experiments used spontaneous epileptic seizures in kainate-treated rats to examine the efficacy of topiramate (TPM) with a repeated-measures, crossover protocol. METHODS: Kainic acid was administered in repeated low doses (5 mg/kg) every hour until each Sprague-Dawley rat experienced convulsive status epilepticus for >3 h. Six 1-month trials (n = 6-10 rats) assessed the effects of 0.3-100 mg/kg TPM on spontaneous seizures. Each trial involved six pairs of TPM and saline-control treatments administered as intraperitoneal injections on alternate days with a recovery day between each treatment day. Data analysis included a log transformation to compensate for the asymmetric distribution of values and the heterogeneous variances, which appeared to arise from clustering of seizures. RESULTS: A significant effect of TPM was observed for 12 h (i.e., two 6-h periods) after a 30-mg/kg injection, and full recovery from the drug effect was complete within 43 h. TPM exerted a significant effect at doses of 10, 30, and 100 mg/kg, and the effects of TPM (0.3-100 mg/kg) were dose dependent. CONCLUSIONS: These data suggest that animal models with spontaneous seizures, such as kainate- and pilocarpine-treated rats, can be used efficiently for rapid testing of AEDs with a repeated-measures, crossover protocol. Furthermore, the results indicate that this design allows both dose-effect and time-course-of recovery studies. PMID- 15660764 TI - Anticonvulsant activity of ginseng on seizures induced by chemical convulsants. AB - PURPOSE: To test the anticonvulsant activity of three preparations of American ginseng: whole root extract, whole leaves/stems extract, and a partially purified extract that concentrates the Rb ginsenosides (Rb extract). METHODS: One hour after treatment with normal saline, or one of the three ginseng preparations, seizures were induced in adult, male, Sprague-Dawley rats with kainic acid (KA; 10 mg/kg), pilocarpine (300 mg/kg, preceded by methylscopolamine, 1 mg/kg, s.c.), or pentylenetetrazol (PTZ, 50 mg/kg). Time to onset of seizure activity, duration of seizure activity for PTZ, seizure severity, and weight change for KA and pilocarpine were determined for each animal. The brains from animals who had received KA or pilocarpine were examined for severe neuronal stress, by using immunoreactivity for heat-shock protein (HSP)72. RESULTS: The Rb extract had a dose-dependent anticonvulsant effect in all three models of chemically induced seizures: increasing the latency to the seizures; decreasing the seizure score, weight loss, and subsequent neuronal damage after pilocarpine; and shortening the seizure duration and reducing mortality after PTZ. The Rb extract also significantly reduced the effects of KA, including completely blocking behavioral seizures. The root preparation increased the mortality rate after administration of pilocarpine, but had no other significant effects. The leaves/stems preparation, at 120 mg/kg, reduced the weight loss after pilocarpine, but had no other significant effects. CONCLUSIONS: Ginseng extract made from either the root or leaves/stems is ineffective against chemically induced seizures. A partial purification of the whole extract that concentrates the Rb1 and Rb3 ginsenosides has significant anticonvulsant properties. PMID- 15660765 TI - Quantitative neuropathology of the entorhinal cortex region in patients with hippocampal sclerosis and temporal lobe epilepsy. AB - PURPOSE: Clinical, radiologic, and experimental evidence indicates that the entorhinal cortex (EC) region may be linked to the pathophysiology of hippocampal sclerosis (HS) in patients with temporal lobe epilepsy. Few neuropathologic studies of this region have been undertaken in patients with HS undergoing surgery, some suggesting preferential loss of layer III neurones. METHODS: We carried out a quantitative analysis in 26 patients with HS, nine patients with lesional temporal lobe epilepsy (LTLE), and eight postmortem controls. We measured neuronal densities in EC by using a three-dimensional cell-counting technique on NeuN immunostained and Nissl-stained sections. We also quantified the density of calretinin-positive interneurones in this region and the density of neurones in adjacent subiculum and CA1 subfields. We also assessed the patterns of gliosis in the EC in the patient groups and the presence of any neocortical neurone loss. RESULTS: No significant difference was found in the mean neuronal densities in the EC region between HS and LTLE groups or postmortem controls. Laminar gliosis in midcortical layers was seen in a proportion of HS cases but also in the LTLE group. No significant difference was seen in the density of calretinin interneurones and no correlation between the presence of neocortical neuronal loss and EC neuronal densities. CONCLUSIONS: A stereotypical pattern of neuronal loss and gliosis in the EC region in patients with HS is not confirmed that distinguishes this pathologic process from that in patients with lesional TLE. PMID- 15660766 TI - Dose-dependent safety and efficacy of zonisamide: a randomized, double-blind, placebo-controlled study in patients with refractory partial seizures. AB - PURPOSE: To evaluate the safety and efficacy of zonisamide (ZNS) as adjunctive treatment in patients with refractory localization-related epilepsy. METHODS: This was a double-blind, placebo-controlled study of adjunctive ZNS in 351 patients with refractory partial seizures receiving a stable regimen of one to three antiepileptic drugs (AEDs). Patients were randomized to placebo or ZNS, 100 mg, 300 mg, or 500 mg/day (2:1:1:2) after a 12-week baseline. Dose titration was undertaken over a 6-week titration phase, which was followed by an 18-week fixed dose assessment phase. Primary efficacy parameters were the differences between ZNS, 500 mg/day, and placebo in the change from baseline in frequency of complex partial (CP) seizures during the fixed-dose assessment phase and in the proportion of CP responders (> or =50% decrease from baseline in seizure frequency). Safety and tolerability also were assessed. RESULTS: Compared with placebo, the highest dose of ZNS (500 mg/day) resulted in a significantly greater decrease in CP seizure frequency from baseline (51.2% vs. 16.3%; p < 0.0001) and a significantly higher proportion of CP responders (52.3% vs. 21.3%; p < 0.001). Both ZNS, 500 mg/day, and 300 mg/day were statistically superior to placebo in reducing the frequency of "all seizures" and simple partial (SP) + CP seizures. For all seizures, a significant dose-response relation was observed (p < 0.0001). The most common adverse events were somnolence, headache, dizziness, and nausea during the titration phase and headache and pharyngitis during the fixed-dose assessment phase. CONCLUSIONS: ZNS provides dose-dependent, effective, and generally well-tolerated adjunctive therapy in patients with partial seizures. PMID- 15660767 TI - Hypothalamic hamartomas and seizures: distinct natural history of isolated and Pallister-Hall syndrome cases. AB - PURPOSE: Hypothalamic hamartomas (HHs) have been associated with uncontrolled seizures, and aggressive therapy including surgery is often recommended. However, some patients, particularly those with other findings associated with Pallister Hall syndrome (PHS), have a more benign course. METHODS: Thirty-seven of 40 PHS patients and 16 of 16 patients with isolated HH had a lesion confirmed on magnetic resonance imaging (MRI). Records for all patients were reviewed for the following information: presence of seizures, age at seizure onset, seizure type, seizure frequency, number of antiepileptic medications (AEDs) at the time of evaluation, past AEDs, MRI characteristics of the HH, presence of endocrine dysfunction, and presence of developmental and behavioral problems. RESULTS: All isolated HH patients had a history of seizures, compared with 13 of 40 PHS patients (all PHS patients with seizures had hamartomas). In isolated HH, seizures started earlier in life, occurred more frequently, and were harder to control than those in patients with PHS. Isolated HH patients were more likely to have behavioral and developmental problems than were PHS patients. The T2 signal of the hamartoma was isointense to gray matter in the majority of PHS patients, but showed a significant increase in all but one patient with isolated HH. CONCLUSIONS: Patients with isolated HH have a distinct clinical phenotype, showing more severe seizures and neurologic dysfunction, HH showing increased T2 signal, and are more likely to have precocious puberty. In contrast, PHS patients usually have well-controlled seizures and other endocrine disturbances than precocious puberty. Patients with HH with or without seizures should be evaluated carefully for other clinical manifestations of PHS, particularly before surgery is considered. PMID- 15660768 TI - Pattern-sensitive epilepsy: electroclinical characteristics, natural history, and delineation of the epileptic syndrome. AB - PURPOSE: To elucidate the electroclinical features and long-term outcome of patients with pattern-sensitive epilepsy. METHODS: We reviewed the clinical and electroencephalographic (EEG) findings of 73 (43 female and 30 male) patients in whom pattern-sensitive epilepsy was diagnosed at Mayo Clinic (Rochester, Minnesota, U.S.A.) from 1950 through 1999. We contacted patients and their relatives by letter or telephone to obtain the latest seizure and quality-of-life outcomes. RESULTS: The median age at onset of seizures was 12.8 years (range, 0.6 32.9 years). Most patients had absence, myoclonic, or generalized tonic-clonic seizures. Interictal epileptiform discharges in the EEG were detected in 61 (83.6%) patients and were generalized in 54 (74%). Paroxysmal epileptiform discharges in the EEG elicited with standard patterns were all generalized in two thirds of patients but were restricted to the posterior head region in one-third. Eight (11%) patients did not exhibit photosensitivity. Television was the most common precipitant [30 patients (41%)]. Twenty-nine patients gave a clear history of one or more seizures precipitated while viewing environmental patterns such as window screens, garments, tablecloths, and ceiling tiles; the rest of the patients admitted that they preferred to avoid looking at patterned objects because these objects made them uncomfortable. The electroclinical features suggested juvenile myoclonic epilepsy in 14 patients, progressive myoclonus epilepsy in three, progressive familial cerebellar ataxia with myoclonus in two, and severe myoclonic epilepsy of infancy in one. During a median follow-up period of 15.7 years, 25 (45.5%) of 55 patients who were followed up for > or =5 years achieved complete seizure remission. The median age at remission was 24.4 years. The absence of progressive neurologic disease was correlated significantly with remission; a family history of seizures showed a trend in favor of remission. More than two thirds of the patients did not consider the seizures an impediment to their family life or to educational and occupational achievements. CONCLUSIONS: Although pattern sensitivity as a trait occurs in various epileptic syndromes, pattern-sensitive epilepsy is a readily distinguishable subtype of the visually provoked reflex epilepsies. In our opinion, the location and extent of the excitable region or regions within the visual cortex concerned with different attributes of visual function dictate susceptibility to a specific trigger (intermittent light, pattern, or color) or closely related multiple triggers and the resultant electroclinical phenomenon. PMID- 15660769 TI - Detection of epileptiform activity by human interpreters: blinded comparison between electroencephalography and magnetoencephalography. AB - PURPOSE: Objectively to evaluate whether independent spike detection by human interpreters is clinically valid in magnetoencephalography (MEG) and to characterize detection differences between MEG and scalp electroencephalography (EEG). METHODS: We simultaneously recorded scalp EEG and MEG data from 43 patients with intractable focal epilepsy. Raw EEG and MEG waveforms were reviewed independently by two experienced epileptologists, one for EEG and one for MEG, blinded to the other modality and to the clinical information. The number and localization of spikes detected by EEG and/or MEG were compared in relation to clinical diagnosis based on postoperative seizure freedom. RESULTS: Interictal spikes were captured in both EEG and MEG in 31, in MEG alone in eight, in EEG alone in one, and in neither modality in three patients. The number of detections ranged widely with no statistical difference between modalities. A median of 25.7% of total spikes was detectable by both modalities. Spike localization was similarly consistent with the epilepsy diagnosis in 85.2% (EEG) and 78.1% (MEG) of the patients. Inaccurate localization occurred only in those cases with very few spikes detected, especially when the detections were in one modality alone. CONCLUSIONS: Interictal epileptiform discharges are easily perceived in MEG. Independent spike identification in MEG can provide clinical results comparable, but not superior, to EEG. Many spikes were seen in only one modality or the other; therefore the use of both EEG and MEG may provide additional information. PMID- 15660770 TI - Hyperventilation revisited: physiological effects and efficacy on focal seizure activation in the era of video-EEG monitoring. AB - PURPOSE: Hyperventilation is an activation method that provokes physiological slowing of brain rhythms, interictal discharges, and seizures, especially in generalized idiopathic epilepsies. In this study we assessed its effectiveness in inducing focal seizures during video-EEG monitoring. METHODS: We analyzed the effects of hyperventilation (HV) during video-EEG monitoring (video-EEG) of patients with medically intractable focal epilepsies. We excluded children younger than 10 years, mentally retarded patients, and individuals with frequent seizures. RESULTS: We analyzed 97 patients; 24 had positive seizure activation (PSA), and 73 had negative seizure activation (NSA). No differences were found between groups regarding sex, age, age at epilepsy onset, duration of epilepsy, frequency of seizures, and etiology. Temporal lobe epilepsies were significantly more activated than frontal lobe epilepsies. Spontaneous and activated seizures did not differ in terms of their clinical characteristics, and the activation did not affect the performance of ictal single-photon emission computed tomography (SPECT). CONCLUSIONS: HV is a safe and effective method of seizure activation during monitoring. It does not modify any of the characteristics of the seizures and allows the obtaining of valuable ictal SPECTs. This observation is clinically relevant and suggests the effectiveness and the potential of HV in shortening the presurgical evaluation, especially of temporal lobe epilepsy patients, consequently reducing its costs and increasing the number of candidates for epilepsy surgery. PMID- 15660771 TI - Cortical resection with electrocorticography for intractable porencephaly-related partial epilepsy. AB - PURPOSE: We evaluated the results of cortical resection of epileptogenic tissue for treatment of intractable porencephaly-related epilepsy. METHODS: We examined clinical features, electrophysiological data, surgical findings, and seizure outcomes after cortical resection in eight patients with intractable epilepsy related to porencephalic cysts. RESULTS: All eight patients had hemiparesis. Five retained motor function in the hemiparetic extremities; six retained visual fields. All had partial seizures, six with secondary generalization. Seven patients had simple and three had complex partial seizures (CPSs); two also had drop attacks. Four patients had multiple seizure types. Long-term scalp video-EEG (LVEEG) localized interictal epileptic abnormalities that anatomically corresponded to the cyst location in three patients. LVEEG recorded ictal-onset zones in five; these anatomically corresponded to the cyst location in three of the five. EEG recorded generalized seizures in two patients, hemispheric in one, and multifocal in two. Intraoperative electrocorticography (ECoG) revealed interictal epileptic areas extending beyond the margins of the cyst in seven patients. We resected ECoG-localized interictal epileptic areas completely in five patients and partially in two. Cortical resection was based on seizure semiology and LVEEG in one patient whose ECoG showed no epileptiform discharges. After a minimum follow-up of 1 year, six patients had excellent seizure outcome (Engel class I), and two had a >90% seizure reduction (Engel class III) without complications. CONCLUSIONS: Cortical resection guided by ECoG allows preservation of motor function and visual field and provides an effective surgical procedure for treatment of intractable epilepsy secondary to porencephaly. PMID- 15660772 TI - Electrographic neonatal seizures after infant heart surgery. AB - PURPOSE: Neonatal seizures are relatively common and an important early sign of acute encephalopathy in those who survive infant heart surgery. The contemporary occurrence of seizures in this setting is not fully known, and their electrographic characteristics are incompletely described. This study describes the characteristics of electrographic neonatal seizures (ENSs) in contemporary infants with congenital heart disease (CHD) surgically repaired by using cardiopulmonary bypass, with or without deep hypothermic circulatory arrest. METHODS: Consecutive infants undergoing heart surgery were monitored by video-EEG for 48 h postoperatively to establish the time of first seizure, total number of ENSs, site(s) of ENS(s) origin and other characteristics. RESULTS: ENSs occurred in 21 (11.5%) of 183 infants. None had clinically visible seizures. The mean time to the first ENS was 21 h (range, 10-36 h). The total number of ENSs among the entire cohort was 1,429. Mean total number of ENSs per patient over a 48-h period was 72 (range, 1-217). Phenobarbital administration was associated with a > or =50% reduction in seizure counts in five (41.7%) of 12 subjects. CONCLUSIONS: ENSs were relatively common in a large, contemporary cohort of infants after infant heart surgery. A wide variation was noted in seizure burden, but many experienced numerous seizures. Electrographic neonatal seizures are a candidate outcome end point in future neuroprotection trials in this patient population. PMID- 15660773 TI - Right-sided vagus nerve stimulation as a treatment for refractory epilepsy in humans. AB - PURPOSE: We present three children who underwent right-sided vagus nerve stimulation (R-VNS). This treatment option for people with refractory epilepsy has not been described in children. METHODS: We reviewed our database of >350 patients implanted with vagus nerve stimulators and now describe our experience in three patients with R-VNS for the treatment of intractable seizures. All three patients improved dramatically with left-sided vagus nerve stimulation (L-VNS), but the devices had to be removed because of infection. The patients were thought to be at high risk for nerve injury if they were reapproached for L-VNSs; therefore R-VNSs were implanted. RESULTS: All three patients with an R-VNS had a reduction in seizures. Our first patient has had an R-VNS for 5 years; he has been seizure free for >2 years on R-VNS monotherapy. The second patient had an R VNS for 8 months. His seizure control improved slightly, but not as dramatically as with L-VNS. The third child has had an R-VNS for >7 months and has cessation of his most disabling seizure type (generalized tonic-clonic seizures). None of the patients had cardiac side effects from therapeutic R-VNS. However, two of the three patients had respiratory events with R-VNS. CONCLUSIONS: VNS is known to be an effective treatment in pharmacoresistant epilepsy. R-VNS should be considered if a patient has significant benefit from L-VNS but is unable to continue with L VNS. R-VNS appears also to have antiepilepsy effects. Additionally, our case report suggests that in some patients, a differential response is found regarding seizure control with R-VNS or L-VNS, raising the question whether L-VNS failures should pursue a trial of R-VNS. Patients should be cautioned and monitored for reactive airway disease if they undergo R-VNS. More research is needed to compare the effects of right- and left-sided VNS on cardiac and pulmonary function in humans and to determine which has the best antiseizure effect. PMID- 15660774 TI - Prediction of verbal memory decline after epilepsy surgery in children: effectiveness of Wada memory asymmetries. AB - PURPOSE: Differences in Wada memory performance after left and right amobarbital injection are powerful predictors of pre- to postoperative memory change among adult epilepsy patients after anterior temporal lobectomy. It is unknown, however, whether these Wada memory asymmetries apply to children who undergo focal cortical resection or to epilepsy surgery patients who undergo resection outside the temporal lobes. METHODS: To investigate these issues, Wada memory asymmetries and pre- to postoperative neuropsychological memory test performances were examined in 132 children who underwent some form of resective epilepsy surgery. Ninety-three (70%) children showed Wada memory asymmetries in the predicted direction (memory after injection ipsilateral to side of surgery better than memory after contralateral injection), and 39 (30%) did not. RESULTS: Children with Wada memory asymmetries showed significant improvement in verbal memory after surgery as compared with children without Wada memory asymmetries who showed significant verbal memory decline. This result was also obtained when individual cases were examined: 77% of children with Wada memory asymmetries in predicted direction showed no verbal memory decline after surgery, whereas 80% of children without asymmetries had lower postoperative verbal memory (passage recall) test scores. Wada memory asymmetries had no value in predicting postoperative changes in visual-spatial memory. CONCLUSIONS: Wada memory asymmetries may be used as one of the factors to assess risk for verbal memory decline after epilepsy surgery in children. PMID- 15660775 TI - Sleep architecture in children with idiopathic generalized epilepsy. AB - PURPOSE: Children with epilepsy experience sleep disturbances, behavioral and attentional problems at higher rates than their peers. However, the relation between sleep disturbances and the observed behavioral and attentional abnormalities is poorly defined. METHODS: Children with primary generalized epilepsy who were seizure free and between the ages of 5 and 18 years were matched with age- and gender-matched healthy controls and underwent two consecutive nights of nocturnal polysomnography with extended electroencephalography. Connor's Continuous Performance Test (CPT) was administered to assess daytime attentional function. Parents completed the Child Behavior Checklist (CBCL) to assess their emotional-behavioral status. Two sample t tests were used to examine group differences. Spearman correlations were used to examine the relation between sleep variables and behavior and attention variables. Multiple regression analysis was used to identify independent predictors of abnormal behavior and attention among patients. RESULTS: Eleven children with primary generalized epilepsy and eight age- and sex-matched controls participated in the study. Children with epilepsy had longer stage 1 sleep percentage (7.19 +/- 3.2 vs. 4.8 +/- 3.5; p = 0.05) and latency to rapid eye-movement (REM) sleep (123.5 +/- 40.1 vs. 101.75 +/- 24.3; p = 0.018) compared with controls. Children with epilepsy had worse attention (CPT index, 10.94 +/- 6.55 vs. 3.42 +/- 4.04; p = 0.004) and exhibited significantly higher CBCL Total Behavior and Internalizing Behavior Problem scales. Whereas regression analysis showed no independent predictors of abnormal behavior and attention, a tendency toward association between CBCL total behavior scale and REM percentage (r= 0.55; p = 0.07), and between CPT overall index and stage 1 sleep percentage (r= 0.40; p = 0.10) was noted. CONCLUSIONS: Sleep architecture is abnormal in children with primary generalized epilepsy. Further studies are needed to determine whether abnormalities in sleep architecture contribute to poor daytime behavior and attention. PMID- 15660776 TI - ApoE epsilon4 allele and disease duration affect verbal learning in mild temporal lobe epilepsy. AB - PURPOSE: To clarify the possible role of other factors including the ApoE epsilon4 allele for memory decline in temporal lobe epilepsy (TLE). METHODS: We conducted a neuropsychological and molecular study in 138 consecutive patients (78 female patients; mean age, 50.2 years, SD +/- 17.9; range, 14 to 87 years) with mild nonlesional TLE, who rarely or never had seizures at long-term follow up. The mean age at seizure onset was 33.0 years (SD, +/-21.7), and the mean duration of epilepsy was 17.1 years (SD, +/-15.7). RESULTS: Thirty-four (25%) of 138 patients had test scores indicating verbal learning deficit (VLD). The presence of an ApoE epsilon4 allele was associated with an increased risk of VLD (OR, 4.18; 95% CI, 1.66-10.55). The effect of the ApoE genotype was independent of both the age at epilepsy onset and disease duration as well as of a low educational level, which were separately associated with VLD (p values = 0.045, 0.001, and 0.001, respectively). A significant linear trend (p = 0.005) was seen in the relation between disease duration and cognitive impairment, with the highest risk being in patients with an epilepsy duration > or =25.5 years (OR, 7.06; 95% CI, 1.67-29.85), especially if they carried the epsilon4 allele (OR, 32.29; 95% CI, 5.23-195.72). CONCLUSIONS: These results provide evidence for an alteration in cognitive performance as a function of the presence of the ApoE epsilon4 allele and point to the critical role of disease duration itself for cognitive impairment in TLE. PMID- 15660777 TI - Abnormal phonologic processing in familial lateral temporal lobe epilepsy due to a new LGI1 mutation. AB - PURPOSE: Autosomal dominant lateral temporal lobe epilepsy (ADLTLE) is a rare familial epilepsy with onset in adolescence or early adulthood, associated with mutations of LGI1 in most families. We describe the clinical, neuropsychological, and molecular genetic study of a new ADLTLE Italian family. METHODS: A four generation family from Sardinia was studied. Clinical, neuropsychological, and genetic analysis were performed in eight living affected family members. RESULTS: Nine family members had seizures over four generations; four of them had auditory auras and aphasia followed by secondarily generalized tonic-clonic seizures (SGTCs). One individual in addition had visual symptoms, and one family member had only vertigo followed by SGTCs. The side of seizure onset could not be determined in these five patients with focal seizures. The proband had febrile and afebrile tonic-clonic seizures. Two family members had only febrile seizures. Inheritance was autosomal dominant with 59% penetrance. Genetic molecular analysis showed a new LGI1 missense mutation causing a Leu154Pro substitution in six affected and one unaffected individuals. Dichotic listening performance was abnormal in four affected individuals compared with controls. Fluency and lexical abilities also were pathological in three patients. These findings showed that in patients, the left temporal lobe was less specialized in the auditory processing function than in controls. CONCLUSIONS: In this ADLTLE family, both seizure semiology and neuropsychological findings point to a lateral temporal lobe dysfunction. The newly identified LGI1 mutation might underlie both the seizure disorder and the neuropsychological deficits. PMID- 15660778 TI - Prevalence, incidence, and etiology of epilepsies in rural Honduras: the Salama Study. AB - PURPOSE: Determination of epilepsy etiology in population-based studies is difficult because of the high cost of diagnostic tests. However, cost effectiveness may be proven if preventive public-health strategies can be established from the test results. We report an epilepsy population-based study using clinical and laboratory techniques. METHODS: A medical team administered an epilepsy survey to 88% of the residents by census in the rural county of Salama, Honduras. Ninety of 100 participants identified with active epilepsy underwent a neurologic examination, video-electroencephalography (video-EEG), brain computed tomography (CT) scan, and serum enzyme-linked immunoelectrotransfer blot (EITB) for cysticercosis. Final diagnoses were based on the International League Against Epilepsy classifications for seizures and epilepsy syndromes. Combined epidemiologic, clinical, video-EEG, neuroimaging, and serum EITB assays were used for the diagnosis of epilepsy etiologies. RESULTS: Among 6,473 residents surveyed, 151 persons with epilepsy (prevalence rate, 23.3/1,000) were identified, 100 of whom had active epilepsy (15.4/1,000) on the prevalence day. Incidence was determined to be 92.7/100,000. Partial seizures with or without secondary generalization were common (92.2%). Symptomatic epilepsy (62%) was primarily due to neurocysticercosis (37%), perinatal brain damage (8%), post traumatic (3%), and poststroke (2%). Eight percent were idiopathic, and 30% were cryptogenic (unknown cause). CONCLUSIONS: Symptomatic epilepsies primarily explained the high prevalence and incidence of epilepsy in Salama. Integration of video-EEG and brain CT scan with clinical-epidemiologic evaluation was critical for determination of epilepsy etiology. Establishment of specific programs for continuation of epidemiologic surveillance, education, intervention, and long term follow-up will benefit the Salama region. PMID- 15660779 TI - Quality of life of people with epilepsy in Iran, the Gulf, and Near East. AB - PURPOSE: To study the impact of epilepsy and its treatment on the quality of life of people living in Iran and in countries in the Gulf and Near East. METHODS: Clinical, demographic, and psychosocial details were collected through the use of a self-completed questionnaire distributed to patients attending hospital outpatient clinics. RESULTS: Data were collected from 3,889 people with epilepsy from 10 countries. More than 40% of all respondents had frequent seizures, and reported levels of side effects from medication were high, the most commonly reported being nervousness, headaches, and tiredness. A significant number of respondents reported changing their medications because of side effects or poor seizure control. Respondents reported that epilepsy and its treatment had a significant impact on a number of different aspects of their daily lives. A significant number of respondents felt stigmatized by their epilepsy. Reported health status was reduced when compared with that of people without epilepsy, particularly for physical and social functioning and energy and vitality, as assessed by using a generic health status measure, the SF-36. CONCLUSIONS: This is the largest study to date documenting the impact of epilepsy and its treatment in Iran, the Gulf, and Near East regions. Differences were found between the quality-of-life profiles of respondents in this study and those who participated in an earlier parallel study in Europe. PMID- 15660780 TI - Evidence for a role of the parafascicular nucleus of the thalamus in the control of epileptic seizures by the superior colliculus. AB - PURPOSE: The aim of this study was to investigate whether the nucleus parafascicularis (Pf) of the thalamus could be a relay of the control of epileptic seizures by the superior colliculus (SC). The Pf is one of the main ascending projections of the SC, the disinhibition of which has been shown to suppress seizures in different animal models and has been proposed as the main relay of the nigral control of epilepsy. METHODS: Rats with genetic absence seizures (generalized absence epilepsy rat from Strasbourg or GAERS) were used in this study. The effect of bilateral microinjection of picrotoxin, a gamma aminobutyric acid (GABA) antagonist, in the SC on the glutamate and GABA extracellular concentration within the Pf was first investigated by using microdialysis. In a second experiment, the effect of direct activation of Pf neurons on the occurrence of absence seizures was examined with microinjection of low doses of kainate, a glutamate agonist. RESULTS: Bilateral injection of picrotoxin (33 pmol/side) in the SC suppressed spike-and-wave discharges for 20 min. This treatment resulted in an increase of glutamate but not GABA levels in the Pf during the same time course. Bilateral injection of kainate (35 pmol/side) into the Pf significantly suppressed spike-and-wave discharges for 20 min, whereas such injections were without effects when at least one site was located outside the Pf. CONCLUSIONS: These data suggest that glutamatergic projections to the Pf could be involved in the control of seizures by the SC. Disinhibition of these neurons could lead to seizure suppression and may be involved in the nigral control of epilepsy. PMID- 15660781 TI - Seizure clustering: risks and outcomes. AB - PURPOSE: Patients with epilepsy often experience seizures in clusters. In this preliminary study, we examined the prevalence of self-reported clustering and identified potential risk factors and outcomes associated with reported clustering for further analysis in an ongoing prospective diary study. METHODS: Detailed intake questionnaires were administered. Subjects reporting their typical seizure pattern to be three or more seizures within a 24-h period were classified as clusterers. Magnetic resonance imaging (MRI) and EEG data were obtained, and epilepsy was classified by International League Against Epilepsy (ILAE) criteria. All subjects completed Beck Anxiety and Depression Inventories, and Quality of Life in Epilepsy testing. RESULTS: Overall, 29% of subjects reported typical seizure clustering. Extratemporal lobe epilepsy [p = 0.02; OR, 3.0 (1.1-7.8)] was significantly associated with reported seizure clustering. Remote symptomatic epilepsy was associated with clustering [p = 0.03; OR, 2.3 (1.1-4.3)], particularly in association with a history of head trauma with loss of consciousness before epilepsy onset [p = 0.003; OR, 4.2 (1.6-11.1)], although this analysis was limited. Reported clustering was significantly associated with a history of convulsive status epilepticus (SE) [p = 0.029; OR, 3.0 (1.1-8.3)], other seizure-related hospitalization [p = 0.006; OR, 5.3 (1.5-17.6)], and worse seizure control (p = 0.004). Quality-of-life measures were not significantly associated with reported clustering. CONCLUSIONS: These preliminary results identify extratemporal epilepsy and possibly head trauma as potential risk factors for reported seizure clustering and indicate a significant association between reported clustering and convulsive SE. Additionally, seizure clustering appears to be a marker for more intractable epilepsy. The ongoing diary study will refine the definition of seizure clustering and further evaluate specific predictors of poor and favorable outcomes of clustering. PMID- 15660782 TI - Epilepsy in Wolf-Hirschhorn syndrome (4p-). AB - PURPOSE: We investigated the evolution of epilepsy, seizure types, and effective drugs in Wolf-Hirschhorn syndrome, which is a malformation syndrome often with refractory seizures and status epilepticus. METHODS: We reviewed 11 cases of Wolf Hirschhorn syndrome (age range, 2-25 years; SD, 7.2 years) and who were treated in Osaka University or Osaka Medical Center of Research Institute for Maternal and Child Health. RESULTS: In all patients, febrile or afebrile convulsions had developed. Epileptic seizures included alternative hemiconvulsions, generalized tonic-clonic seizures, focal clonic seizures, tonic seizures, and epileptic spasms. Seizures were often induced by a high fever or a hot bath. Status epilepticus occurred in all patients, including one patient who died at the first status epilepticus. In some cases, intratracheal intubation was needed because of respiratory insufficiency. The effective antiepileptic drugs for long-term use were sodium bromide (four of four), followed by clorazepate (CLP; one of two), and nitrazepam (NZP; two of four). Sodium bromide was particularly effective for preventing status epilepticus. The mean age of last status epilepticus in patients receiving sodium bromide (1 year 8 months) was significantly younger than that in those not treated with sodium bromide (3 year 4 months). CONCLUSIONS: We identified that, in most patients of Wolf-Hirschhorn syndrome, the frequency of both seizures and status epilepticus decreased gradually after age 5 years. However, during infancy, status epilepticus sometimes resulted in permanent disability or even death. We propose that sodium bromide should be used as the initial treatment for the prevention of the development of status epilepticus associated with Wolf-Hirschhorn syndrome. PMID- 15660783 TI - Gabapentin-induced myoclonus in end-stage renal disease. AB - PURPOSE: We analyzed the occurrence and clinical features of myoclonus in patients with end-stage renal disease (ESRD) who were treated with gabapentin (GBP). METHODS: We reviewed the medical records of patients with ESRD who were treated with GBP and hospitalized during an 18-month period and analyzed clinical details such as type of myoclonus, doses of GBP, electroencephalographic (EEG) findings, and relation of symptoms to GBP exposure and dosage. RESULTS: Three of 71 patients had myoclonus with GBP doses ranging from 9 mg/kg to 20 mg/kg and within 4 months of treatment onset. Myoclonus was characterized as multifocal, involving all extremities in the three patients. EEG did not show epileptiform discharges with the myoclonus. Myoclonus resolved in the three individuals within 4-15 days after GBP was discontinued. CONCLUSIONS: GBP increases the risk of myoclonus in ESRD. Myoclonus in these individuals was more disabling than that in patients with normal renal function, and discontinuation of GBP is required to restore normal function. PMID- 15660784 TI - Typical absence seizures triggered by photosensitivity. AB - PURPOSE: To describe the characteristics of patients with typical absence seizures (TASs), consistently triggered by photosensitivity. METHODS: Consecutive patients having TAS induced by intermittent photic stimulation were included in the study. All clinical parameters, EEG, and video-EEG data were assessed during the long-term follow-up. Statistical analyses were performed with SPSS 10.0 software. RESULTS: Nine female and two male patients with a mean age at onset of 14 +/- 5.9 years (range, 7-27 years) and with a mean follow-up of 9 +/- 7.56 years had photosensitive TASs. They constituted 7.64% of absence epilepsies and 0.4% of all patients seen in our tertiary center. The seizures were usually subtle and had a reported frequency of 1 to 9 times daily. Seven patients were clinically photosensitive and reported that some of their TASs were induced by photic stimuli in daily life. All patients also had spontaneous TASs, and four of them had generalized tonic-clonic seizures. EEG results did not show any distinctive features when compared with those of other cases with TASs. Remission could not be achieved in five patients with antiepileptic drug treatments, and we always observed relapses after drug discontinuation or dose reduction in the remaining six cases in remission. Spontaneous remission did not occur even in the five patients older than 30 years. CONCLUSIONS: TASs triggered by photosensitivity are a rare and heterogeneous clinical condition with a marked female preponderance. It is notable that TASs do not remit in these cases. PMID- 15660785 TI - Paroxysmal dyskinesia with secondary generalization of tonic-clonic seizures in pseudohypoparathyroidism. PMID- 15660786 TI - Erectile dysfunction with topiramate. PMID- 15660789 TI - Vaccine-associated granulomatous inflammation and melanin accumulation in Atlantic salmon, Salmo salar L., white muscle. AB - The purpose of this study was to investigate the nature of variably sized pigmented foci encountered in fillets of farmed Atlantic salmon, Salmo salar L. The material was sampled on the fillet production line and on salmon farms from fish with an average size of 3 kg from various producers. The fish had been routinely vaccinated by injection. Gross pathology, histology, immunohistochemistry using antisera against major histocompatibility complex (MHC) class II beta chain and transmission electron microscopy (TEM) were used to characterize the changes. Macroscopically, melanized foci were seen penetrating from the peritoneum deep into the abdominal wall, sometimes right through to the skin, and also embedded in the caudal musculature. Histological investigation revealed muscle degeneration and necrosis, fibrosis and granulomatous inflammation containing varying numbers of melano-macrophages. Vacuoles, either empty or containing heterogeneous material, were frequently seen. The presence of abundant MHC class II+ cells indicated an active inflammatory condition. TEM showed large extracellular vacuoles and leucocytes containing homogeneous material of lipid-like appearance. The results showed that the melanized foci in Atlantic salmon fillet resulted from an inflammatory condition probably induced by vaccination. The described condition is not known in wild salmon and in farmed salmon where injection vaccination is not applied. PMID- 15660788 TI - The pathology of chronic erosive dermatopathy in Murray cod, Maccullochella peelii peelii (Mitchell). AB - Chronic erosive dermatopathy (CED) is a disease of intensively farmed Murray cod in Australia that has been reported in association with the use of groundwater (mechanically extracted from shallow boreholes) supplies. CED results in focal ulceration of the skin overlying sensory canals of the head and flanks. Trials were conducted at an affected fish farm to study the development of the condition, both in Murray cod and in goldfish, and also to assess the reported recovery of lesions when affected fish were transferred to river water. Grossly, lesions began after 2-3 weeks with degeneration of tissue at the periphery of pores communicating with the sensory canals. Widening of these pores along the axis of the canals resulted from a loss of tissue covering the canal. Histopathologically, hyperplasia of the canal epithelial lining was seen after 3 weeks in borehole water and subsequent necrosis and sloughing of this tissue resulted in the loss of the canal roof. Canal regeneration occurred when fish were transferred from borehole water into river water. The lack of lesions in other organs and the pattern of lesion development support exposure to waterborne factors as the most likely aetiology. PMID- 15660790 TI - White tail disease of the giant freshwater prawn, Macrobrachium rosenbergii: separation of the associated virions and characterization of MrNV as a new type of nodavirus. AB - White tail disease of the farmed freshwater prawn, Macrobrachium rosenbergii, is the cause of mortalities in the French West Indies, China and India. Two different sized particles, both developing in the cytoplasm of target cells, are found associated with diseased animals. These two viruses were separated, purified and subsequently characterized. The larger one, called MrNV, is icosahedral in shape and 27 nm in diameter. Its genome is composed of two fragments of linear single-stranded RNA (ss-RNA), of 2.9 and 1.3 kb, respectively and its capsids exhibited a single polypeptide of 43 kDa. These characteristics and the partial sequence of a cloned fragment of RNA-1 suggest this agent is a member of the family Nodaviridae, but with differences from both the genera Alphanodavirus and Betanodavirus. The smaller virus, named XSV, is icosahedral in shape, 15 nm in diameter, possesses a linear ss-RNA genome of about 0.9 kb, and its capsid exhibits two polypeptides of 16 and 17 kDa, respectively. The relationships between these two viruses remain unknown. PMID- 15660791 TI - Effectiveness of a divalent vaccine for sole, Solea senegalensis (Kaup), against Vibrio harveyi and Photobacterium damselae subsp. piscicida. AB - The protection of cultured sole, Solea senegalensis, against Vibrio harveyi and Photobacterium damselae subsp. piscicida was evaluated following the use of a divalent vaccine prepared with formalized whole cells and extracellular products of virulent strains of both pathogenic microorganisms and administered by the immersion route. Two prolonged immersions of 5-10 g fish in the divalent bacterin at a 1-month interval gave high levels of protection similar to those obtained when the respective monovalent vaccines were administered by the intraperitoneal route [relative percentage of survival (RPS) values >70%], which indicates that the former procedure can be a useful strategy with small fish. The high protection afforded by the divalent vaccine in sole lasted for 4 months after which the RPS values against both pathogens decreased significantly. PMID- 15660792 TI - Flavobacterium psychrophilum in rainbow trout, Oncorhynchus mykiss (Walbaum), hatcheries: studies on broodstock, eggs, fry and environment. AB - The occurrence of Flavobacterium psychrophilum at four rainbow trout hatcheries was investigated to provide more knowledge about the reservoirs and transmission of this bacterium. Broodstock were sampled at stripping (including both unfertilized and fertilized eggs), and the offspring were then sampled at the eyed egg and fry stages. Water and surface samples (e.g. hatchery trays) were also sampled. Flavobacterium psychrophilum was found in ovarian fluid and milt, indicating that broodstock may serve as a reservoir and are latent carriers of the pathogen. Flavobacterium psychrophilum was not found on or inside eggs, but further egg studies will be necessary to elucidate the possibility of vertical transmission of the pathogen. Flavobacterium psychrophilum was isolated from water samples, but only from water that had been in close contact with farmed rainbow trout or eggs. Flavobacterium psychrophilum isolates were characterized and compared with well-characterized strains, using degradation of elastin, serotype and ribotype profiles. Different ribotypes of F. psychrophilum were found between hatcheries, but a common ribotype A was found at all four hatcheries. Different ribotypes were found in broodstock without clinical disease, whereas only a few ribotypes (mostly ribotype A) were found in diseased fry. The same ribotype A was found in broodstock, in water samples from hatchery trays and in fry, which suggests the possibility of transmission of F. psychrophilum between broodstock and offspring. PMID- 15660793 TI - Neoparamoeba branchiphila n. sp., and related species of the genus Neoparamoeba Page, 1987: morphological and molecular characterization of selected strains. AB - A total of 18 Neoparamoeba strains were characterized both morphologically and using the SSU rRNA gene sequences as molecular markers. Nine were isolated from gills of farmed Atlantic salmon, Salmo salar L., six from sediments sampled in areas of sea-cage farms and three from net material of sea-cages. The newly obtained sequences extended substantially the dataset of Neoparamoeba strains available for phylogenetic analyses, which were used to infer taxonomic relatedness among 32 strains morphologically assigned to this genus. In addition to the N. pemaquidensis and N. aestuarina clades, phylogenetic analyses clearly distinguished a third clade with sequences from six strains. Members of this clade are characterized as representatives of a new species, N. branchiphila n. sp. The diagnostic primers for the identification of this species are introduced. PMID- 15660794 TI - Transthoracic direct current cardioversion of atrial fibrillation: one biphasic cardioverter is better than two monophasic machines. PMID- 15660795 TI - Repeated dual external direct cardioversions using two simultaneous 360-J shocks for refractory atrial fibrillation are safe and effective. AB - Failure of cardioversion of atrial fibrillation (AF) to sinus rhythm (SR) by standard external direct current cardioversion (DCC) may be due to failure of delivery of enough defibrillating energy rather than to the true refractoriness of AF. Ninety-nine patients with persistent AF (76 male; age 63.7 +/- 0.4 years; weight 113.1 +/- 25.1 kg) who failed standard DCC were included in this report. Under anesthesia, QRS synchronous shocks were delivered across anteroposterior electrodes in the following sequence: (1) a single 360-J shock; (2) another single 360-J shock within 2 minutes; (3) 30 minutes of rest, reinduction of anesthesia and delivery of two simultaneous monophasic 360-J shocks. All patients underwent all three DCC steps. Sixty-six (67%) patients converted to SR following the first dual simultaneous shock. Fourteen patients (14%) required more than one dual shock to achieve SR. This increased the overall success rate of resuming SR to 81%. Except for minor skin burns in three patients there were no procedure related complications. On follow-up at 1 month, 55 (56%) patients were still in SR, whereas 50 (51%) patients maintained SR at 12 months. This was similar to our general DCC population (55% of the 1698 patients were in SR 6 months post-DCC, P = ns). In conclusion, dual external monophasic 360-J DCC is an effective rescue technique for restoration of SR in patients with AF refractory to standard DCC. AF in these patients seems to be as amenable to chronic suppression as AF in the general population of DCC patients. PMID- 15660796 TI - The value of magnetocardiography in patients with and without relevant stenoses of the coronary arteries using an unshielded system. AB - BACKGROUND: The diagnostic management of patients with chest pain remains a clinical challenge. Magnetocardiography (MCG) is a noninvasive method for the recording of cardiac electromagnetic signals at multiple sites above the chest cage. Contrary to electrocardiogram (ECG) the magnetic field is unaltered by surrounding tissues. The present study aimed to analyze the diagnostic value of an unshielded four-channel MCG for the detection of coronary artery disease (CAD) in patients with chest pain. METHODS: The study included 417 subjects: 177 patients with angiographically documented CAD (stenoses > or =50%), 123 symptomatic patients without hemodynamically relevant stenosis (nCAD) and 117 healthy subjects. Twelve-lead ECG was obtained in all subjects. The magnetocardiography recordings were taken from 36 positions at rest. From these current density vector maps were generated during the ST-T interval. Each map was classified using a classification system with a scale from 0 (normal) to 4 (grossly abnormal). RESULTS: While the ECG was normal in all subjects the MCG revealed typical differences. In normals most maps were classified as category 0, 1 or 2, in nCAD and more so in CAD patients the categories 3 and 4 prevailed. Using a cut-off value of 39.2% for the discrimination between normals and CAD patients sensitivity was 73.3%, specificity 70.1%. CONCLUSION: Contrary to ECG, unshielded MCG reveals significant differences between normals and symptomatic patients with and without relevant stenoses using current density reconstruction during repolarization at rest. This method might be a suitable noninvasive tool for the management of patients with chest pain. PMID- 15660797 TI - Individual optimization of pacing sensors improves exercise capacity without influencing quality of life. AB - INTRODUCTION: Programmable pacemaker sensor features are frequently used in default setting. Limited data are available about the effect of sensor optimization on exercise capacity and quality of life (QOL). Influence of individual optimization of sensors on QOL and exercise tolerance was investigated in a randomized, single blind study in patients with VVIR, DDDR, or AAIR pacemakers. METHODS: Patients with > or =75% pacing were randomized to optimized sensor settings (OSS) or default sensor setting (DSS). Standardized optimization was performed using three different exercise tests. QOL questionnaires (QOL-q: Hacettepe, Karolinska, and RAND-36) were used for evaluation of the sensor optimization. One month before and after optimization, exercise capacity using chronotropic assessment exercise protocol and the three QOL-q were assessed. RESULTS: Fifty-four patients (26 male, 28 female) with a mean age of 65 +/- 16 years were enrolled in the study. In each group (OSS and DSS) 27 patients were included. One month after sensor optimization, the achieved maximal heart rate (HR) and metabolic workload (METS) were significantly higher in OSS when compared with DSS (124 +/- 28 bpm vs 108 +/- 20 bpm, P = 0.036; 7.3 +/- 4 METS vs 4.9 +/- 4 METS, P = 0.045). Highest HR and METS were achieved in patients with pacemakers with accessible sensor algorithms. In patients with automatic slope settings (33%), exercise capacity did not improve after sensor optimization. QOL did not improve in OSS compared with DSS. CONCLUSION: After 1 month of individual optimization of rate response pacemakers, exercise capacity was improved and maximum HR increased, although QOL remained unchanged. Accessible pacemaker sensor algorithms are mandatory for individual optimization. PMID- 15660798 TI - Electrocardiographic characteristics of fasciculoventricular pathways. AB - INTRODUCTION: Fasciculoventricular (FV) pathways are rare variants of preexcitation, and their ECGs may be misinterpreted as Wolff-Parkinson-White syndrome with anteroseptal accessory pathways (WPW-AS). We analyzed the electrocardiographic characteristics of the patients with FV pathways to find out the different findings from WPW-AS. METHODS AND RESULTS: Five patients with FV pathways and four patients with WPW-AS who underwent electrophysiologic studies were evaluated. Intervals and amplitudes of each wave and QRS morphologies were analyzed in standard 12-lead ECGs of these patients by two independent cardiologists without the information of the electrophysiologic findings. PR intervals were longer in FV pathways (122 +/- 11.0 vs 83 +/- 21 ms, FV pathways vs WPW-AS, P = 0.017). In lead V1, narrower width of R waves (25 +/- 6 vs 45 +/- 13 ms, P = 0.037) and smaller amplitude of S waves (12.8 +/- 8.3 vs 26.6 +/- 7.4 mm, P = 0.037) were observed in FV pathways. The polarity of delta waves in V1 was flat or negative in contrast with the cases of WPW-AS in which the polarity was positive. Three of five patients had notching in the descending limb of S waves in V1, which was not observed in WPW-AS. CONCLUSION: FV pathways have different ECG characteristics from WPW-AS in PR interval and morphology of QRS complexes in lead V1 of the standard 12-lead ECG. PMID- 15660799 TI - Diagnostic value of nitrate stimulated tilt testing without preceding passive tilt in patients with suspected vasovagal syncope and a healthy control group. AB - BACKGROUND: The contribution of the passive tilt phase, in a combined nitrate stimulated tilt protocol may be low yielding. In order to develop a shortened, and thus easier to perform test, we investigated the optimal duration of a nitrate tilt protocol without a preceding passive phase. METHODS: Thirty-eight consecutive patients (18F/20M; mean age 46 +/- 16) with clinically suspected vasovagal syncope and 31 control subjects (15F/16M; mean age 40 +/- 18) were tested. The subjects were tilted to 70 degrees for a maximum period of 30 minutes, and sublingual nitroglycerin 0.4 mg spray was administrated directly after attaining erect posture. Receiver operator characteristics (ROC) analysis was done to determine the optimal test duration. RESULTS: In the patient group 31 (82%) and in controls 5 (16%) had a positive test. Sensitivity, specificity, and accuracy at test end were 82, 84, and 83%, respectively. ROC analysis revealed that a maximum accuracy of 83% was attained at 14 minutes, with a sensitivity and specificity of 79 and 87%, respectively. CONCLUSION: Nitrate stimulated tilt testing, without a preceding passive tilt phase, and limited to a test duration of 15 minutes, provides an accurate, sensitive, and specific method to provoke vasovagal reactions in subjects with clinically suspected vasovagal syncope. PMID- 15660800 TI - Editorial comment: Tilt testing made easy? PMID- 15660801 TI - Site specificity of transverse crista terminalis conduction in patients with atrial flutter. AB - INTRODUCTION: The causes of transcristal conduction (TC) in patients with atrial flutter (AFL) are unknown. METHODS AND RESULTS: In two groups of patients referred for AFL ablation, 36 had cavotricuspid isthmus (CTI) dependent flutter (Group I) and 24 had lower (n = 21) or upper loop reentry (n = 5) (Group II). After ablation, isthmus block was evaluated by pacing from the coronary sinus (CS) and low lateral right atrium and by alternative techniques, including mapping with electrodes spanning the CTI or electroanatomic mapping. After bidirectional CTI block was verified, 21/36 (58%) in Group I showed TC with CS pacing, including low TC in 16 (including 11 showing "pseudo" CTI conduction), higher TC in 6 and multiple breaks in 3. However, 8 with low TC during CS pacing showed unidirectional block by pacing outside of the CS os. Twelve (50%) in Group II had TC during CS pacing after bidirectional CTI block, with low TC in 5 (2 mimicking residual CTI conduction) and higher breaks in 9. There was no significant difference in the incidence of TC during CS pacing after CTI block between groups. In seven autopsied hearts, the muscle orientation between the proximal CS musculature and Eustachian ridge were examined. Muscular connections between the CS and Eustachian ridge coursing toward the orifice of inferior vena cava were found in one of the hearts. CONCLUSIONS: It is concluded that in patients with bidirectional CTI block, pacing from the CS may be associated with TC mimicking a conduction leak through the isthmus. Pacing just outside the CS os helps distinguish pseudo from true isthmus block. PMID- 15660802 TI - Prevalence and management of inappropriate detection and therapies in patients with first-generation biventricular pacemaker-defibrillators. AB - BACKGROUND: Tachycardia detection in first-generation biventricular pacemaker implantable cardioverter defibrillators (BiV ICD) occurs through both the right ventricular (RV) and left ventricular (LV) leads, creating the potential for inappropriate detection and therapies. Little is known regarding the prevalence and management of patients with BiV ICDs and inappropriate detection. METHODS AND RESULTS: A transvenous, first-generation BiV ICD was implanted in 77 consecutive patients (age 61 +/- 11 years) for drug-refractory heart failure. The mean New York Heart Association class, QRS duration, and ejection fraction were 3.1 +/- 0.4, 168 +/- 24 ms, and 0.19 +/- 0.07, respectively. Among the 77 patients, 17 (22%) experienced inappropriate detection at a mean of 154 +/- 140 days after implantation. Fifteen of the 17 patients (88%) experienced inappropriate ICD therapy. In 16 of the 17 (94%) patients, the cause of inappropriate detection was double counting during sinus (8) or atrial rhythm (3), and nonsustained ventricular tachycardia (5). Despite reprogramming of the ICD, 9 patients (53%) required an additional procedure because of inappropriate therapies, including an upgrade to a dedicated BiV ICD (5), revision of the LV lead (2), ablation of the atrioventricular junction (1), and repeat defibrillation threshold testing (2). CONCLUSIONS: Inappropriate detection in patients with a first-generation BiV ICD is common and often results in inappropriate ICD therapy. The most common mechanism of inappropriate detection is double counting that often creates the need for additional procedures. Although devices in which tachycardia detection occurs only through the RV lead now are available, close follow-up of the many patients who received a first-generation BiV ICD is necessary. PMID- 15660803 TI - Coronary sinus lead extraction in the era of cardiac resynchronization therapy: single center experience. AB - As the number of coronary sinus (CS) lead implantations for cardiac resynchronization therapy increases so will the need for extraction of these leads. The safety of extraction of leads from the branches of the CS has not been reported. We reviewed our database of patients undergoing pacemaker lead extraction from January 2002 through February 2004 at our institution. Of 149 patients referred for lead extraction, 14 (9%) had a biventricular device. The indications for lead extraction were infection, lead malfunction, and exit block. The duration of CS lead implants ranged between 2 and 43 months (mean 17 months). All 14 CS leads were removed successfully using nonsurgical lead extraction techniques. Three leads that were in place the longest (> or =27 months) were removed via the femoral vein approach due to fibrous attachment of the CS lead body to the other pacemaker leads. The leads were structurally intact and without any significant fibrosis of their tips upon visual inspection. There were no major complications of CS laceration, hypotension, pericardial effusion, or excessive blood loss associated with any of the extraction procedures. CS leads were removed safely, successfully and with relative ease based on our experience in this small cohort of patients. PMID- 15660804 TI - QTc interval in patients with changing edematous states: implications on interpreting repeat QTc interval measurements in patients with anasarca of varying etiology and those undergoing hemodialysis. AB - Associations have been described among weight, amplitude of QRS complexes, and QRS duration (QRSd) in patients with anasarca (AN), and changes in the amplitude of the QRS complexes, QRSd, and QTc after hemodialysis (HD) and in patients with heart failure with associated peripheral edema congestive heart failure. The objective of this study was to evaluate the hypothesis that changes in QTc in patients with AN and after HD are at least partially apparent, due to changing edematous states, and not totally due to altered electrophysiology. QTc was measured in patients with AN on admission, at peak weight (N = 28), and at their subsequent lowest weight (N = 12), in 28 control patients without change in weight during hospitalization, and in one patient before and after 26 HD sessions. In the patients with AN, the QTc was 451 +/- 36 ms on admission and dropped to 423 +/- 46 ms at peak weight (P = 0.005). QTc was 421 +/- 44 ms at peak weight and raised to 434 +/- 30 at subsequent lowest weight (P = 0.32). In the controls, QTc on admission and at discharge were 435 +/- 34 and 428 +/- 23 ms, correspondingly (P = 0.18). QTc increased from 472 +/- 18 ms before to 489 +/ 36 ms after HD (P = 0.017). Alterations in QTc in AN, or HD suggest that the changes in the QTc may be partially only apparent, and due to the electrocardiogram machine-based measurement of the attenuated/augmented QRST complexes resulting from fluid shifts. PMID- 15660805 TI - Inhibition of Unnecessary RV Pacing with AV Search Hysteresis in ICDs (INTRINSIC RV): design and clinical protocol. AB - Implantable cardioverter defibrillators (ICDs) reduce sudden arrhythmic death risk but when these devices are programmed DDD and pace in the right ventricle (RV), they can be associated with increased mortality and heart failure morbidity compared to an ICD programmed to back-up RV. An ideal ICD would provide effective treatment for life-threatening tachyarrhythmias, reduce unnecessary RV pacing and maintain AV synchrony. The Inhibition of Unnecessary RV Pacing with AV Search Hysteresis (AVSH) in ICDs (INTRINSIC RV) study will assess whether an ICD programmed to DDDR with AVSH is equal to an ICD programmed to VVI with regard to mortality, heart failure hospitalizations, and several predefined secondary enpoints. AVSH allows intrinsic AV conduction beyond the programmed AV delay to help minimize ventricular pacing. INTRINSIC RV, a multi-center, randomized, prospective trial will enroll >1,200 participants who receive a Guidant VITALITY AVT ICD. ICDs are programmed initially to DDDR AVSH 60-130. Then, after a week, if the %RV pacing <20%, patients are randomized to VVI-40 or DDDR 60-130 with AVSH. Those with RV pacing > or =20% are placed in an obvservational arm and programmed ad libitum by the treating physician. Patients are followed for one year. This large, randomized, controlled, clinical trial will address whether DDDR with AVSH programming is equivalent to VVI programming in an ICD with regard to mortality and heart failure hospitalization. PMID- 15660806 TI - An elderly woman with AV block in sinus rhythm and conducted atrial tachycardia. PMID- 15660807 TI - Dancing with the crab--for Debbi. PMID- 15660808 TI - Radiofrequency ablation of a concealed right anterior accessory pathway associated with right atrial diverticulum in a child. AB - A 9-year-old boy had refractory supraventricular tachycardia and a right atrial (RA) diverticulum as diagnosed by ultrafast computed tomography. An electrophysiological study under transesophageal echocardiography guidance revealed a concealed right anterior accessory pathway passing through the body of the RA diverticulum. Radiofrequency ablation was performed to eliminate retrograde ventriculoatrial conduction. PMID- 15660809 TI - Surgical open-chest ventricular defibrillation: triphasic waveforms are superior to biphasic waveforms. PMID- 15660810 TI - Leads frozen in the header: a real problem or just many solutions? PMID- 15660811 TI - Delayed detection of ventricular tachycardia in a dual chamber rate adaptive pacing implantable cardioverter defibrillator: a case of intradevice interaction. PMID- 15660813 TI - Comparison of the specificity of implantable dual chamber defibrillator detection algorithms. PMID- 15660815 TI - Intramural radiofrequency ablation: effects of electrode temperature and length. PMID- 15660816 TI - Catheter inversion: a technique to complete isthmus ablation and cure atrial flutter. PMID- 15660817 TI - Transfusion medicine illustrated. Massive hemolysis secondary to alpha-toxin release. PMID- 15660818 TI - The "crisis" of blood donation . . . science. PMID- 15660819 TI - Predonation testing of potential blood donors in resource-restricted settings. PMID- 15660820 TI - Predonation screening of blood donors with rapid tests: implementation and efficacy of a novel approach to blood safety in resource-poor settings. AB - BACKGROUND: In sub-Saharan Africa, the percentage of screened blood is limited to approximately 75 percent for human immunodeficiency virus antibodies (anti-HIV), 50 percent for hepatitis B surface antigen, and 19 percent for hepatitis C virus antibodies (anti-HCV), mainly because of costs. STUDY DESIGN AND METHODS: In 2002 to 2003, candidate blood donors were screened before donation for HIV, HCV, and hepatitis B virus (HBV) serologic markers with rapid tests. The efficacy of this screening was assessed by nucleic acid testing (NAT) applied to pools of 10 plasma samples from donated units with a virus specific triplex assay. NAT reactive pools were resolved by viral genome identification in individual plasma sample. Deferred candidate donors were referred to a donor-care program. RESULTS: A total of 9372 people were screened and 1534 (16.4%) were deferred. No HIV or HCV RNA-containing samples remained undetected by rapid tests unless a human testing error was involved. In contrast, 1.3 and 3.0 percent of HBV DNA containing blood units were negative with rapid tests but were detected in individual donations with enzyme immunoassay and genomic amplification, respectively. Only half of these units were detectable in pools of 10 samples. One-third of deferred candidate donors attended the donor-care program and were informed and counseled. CONCLUSIONS: Predonation viral screening of blood donors is effective in high endemic areas, and the savings it generates may improve the safety and limit the cost of blood. Communication with deferred donors may contribute to public health. A new screening strategy associating serologic rapid test before donation and NAT on pools of 10 plasma samples after donation is proposed. PMID- 15660821 TI - Blood collection and transfusion in the United States in 1999. AB - BACKGROUND: Collection, processing, and transfusion of blood and blood components in the US in 1999 were measured and compared with prior years. STUDY DESIGN AND METHODS: Questionnaires were completed by 2040 blood centers and hospitals. Statistical procedures were used to verify the representativeness of the sample and to estimate national totals. RESULTS: The total US blood supply in 1999 was 13,876,000 units (before testing), 10.1 percent greater than in 1997. It included 13,109,000 allogeneic units, 651,000 autologous units, and 116,000 red cell (RBC) units collected by apheresis. Transfusion of whole blood and RBCs increased by 7.6 percent to 12,389,000 units. Platelet (PLT) transfusions totaled 9,052,000 PLT concentrate equivalent units, of which 66.5 percent were PLTs from apheresis. In comparison with 1997, the total number of PLT units transfused was unchanged, whereas single-donor PLT units transfused increased by 6.7 percent and the transfusion of PLTs from whole blood (PLT concentrates) declined by 10.6 percent (a difference of approximately 400,000 units in each case). CONCLUSIONS: The margin between transfusion demand and the total allogeneic supply in 1999 was 1,203,000 units, 9.1 percent of the supply. By comparison, the margin in 1997 was 7.2 percent, whereas in 1989 it was 13.8 percent. Similarly, the rate of blood collection in 1999 per 1000 population was 11.9 percent higher than the 1997 rate. During the same period, however, the rate of transfusion per 1000 population increased by 5.8 percent. Risk in the future lies primarily in the increasing demand for RBCs and further shrinkage of the supply-and-demand margin. PMID- 15660822 TI - Understanding Canadian student motivations and beliefs about giving blood. AB - BACKGROUND: A better understanding of Canadian blood donor beliefs and motivations is needed to develop targeted interventions. Recruiters must know how motivation variables and correlation patterns differ with donor experience and sex. STUDY DESIGN AND METHODS: Data addressing reasons for donating, statements about the blood supply, beliefs about donation consequences, and reasons for avoiding donation were collected from 450 undergraduates. Principal components analysis was used to investigate the underlying factorial structure of each domain. Men-women and donor-nondonor differences were explored with multivariate analysis of variance techniques. RESULTS: A bivariate model better represented donor beliefs than did a bipolar conceptualization. Negative beliefs distinguished donors and nondonors more so than did positive factors. Altruism dominated reasons for donating, whereas logistic factors accounted for the most variance in donation avoidance. Women were more concerned about adverse physical consequences, and nondonors expressed higher levels of groundless donation related fears. CONCLUSION: Recruiters should consider the sex and donation experience of targets when they develop recruitment and retention strategies. Education programs aimed at overcoming fears and heightening awareness of need are recommended, as are operational improvements aimed at reducing barriers posed by time and inconvenience. PMID- 15660823 TI - Predicting blood donor arrival. AB - BACKGROUND: Keeping waiting time at blood donation short is important for making donation a good experience for the donors and hence to motivate for repeat donations. At the Blood Bank of Oslo, fixed appointments are used, and few donors arrive without appointments. On average, 59 percent of scheduled donors arrive, but day-to-day variations are large. Methods for predicting the number of donors that will arrive on a given day would be valuable in reducing waiting times. STUDY DESIGN AND METHODS: Information about candidate explanatory variables was collected for all appointments made in a 971-day period (179,121 appointments). A logistic regression model for the prediction of blood donor arrival was fitted. RESULTS: Among 18 explanatory variables, the most important were the time from appointment making to appointment date; the contact medium used; the donor age and total number of donations; and the number of no-shows, arrivals, and deferrals during the preceding 2 years. Compared to taking only the average arrival rate into account, prediction intervals were reduced by 43 percent. CONCLUSION: Statistical modeling can provide useful estimates of blood donor arrival, allowing for better planning of donation sessions. PMID- 15660824 TI - Audiovisual touch-screen computer-assisted self-interviewing for donor health histories: results from two years experience with the system. AB - BACKGROUND: The donor history interview is an important aspect of blood safety, in part designed to identify unsuitable donors who may present a risk to blood recipients. There is evidence from behavioral science literature that use of computer-assisted interviewing may be superior to face-to-face (FTF) and paper techniques in eliciting sensitive behavioral information of interest to blood collection facilities. STUDY DESIGN AND METHODS: Audiovisual touch-screen computer-assisted donor self-interviewing with the AABB Uniform Donor History Questionnaire was deployed for routine use in a regional blood center replacing FTF interviews. Donor and staff perception and satisfaction surveys were performed to assess acceptance of the system. Time studies of automated and manual methods were conducted. Rates of deferral of first-time donors for high risk behaviors and rates of errors and omissions on donor interviewing for the two systems were tabulated and compared. RESULTS: Donors and staff strongly preferred the automated system in all dimensions assessed. Donor time increased by 4 minutes but staff time declined by 5 minutes per interview. Identification of high-risk behaviors among first-time donors significantly increased. Rates of errors and omissions on donor history forms identified at audit were reduced. CONCLUSIONS: Both blood donors and collections staff enthusiastically accepted the automated donor interviewing system. A well-designed audiovisual touch-screen donor self-interviewing system is superior to face-to-face interviewing and most likely more effective than paper interviewing. PMID- 15660825 TI - Induction of immune tolerance across major histocompatibility complex barrier by transfusion of ultraviolet B-irradiated immature dendritic cells. AB - BACKGROUND: Transfusion of ultraviolet B (UVB)-irradiated peripheral blood mononuclear cells (PBMNCs) induces immunologic tolerance across the major histocompatibility complex (MHC) barrier in a murine model. It is necessary, however, to reduce contaminating platelets (PLTs) to a minimum. Ex vivo preparation of dendritic cells (DCs) offers an opportunity to rid the contaminating PLTs. The use of immature and mature DCs (iDCs and mDCs) with and without UVB irradiation for tolerance induction was therefore investigated. STUDY DESIGN AND METHODS: iDCs and mDCs were prepared by culture of Balb/c (H-2d) mouse marrow cells in the presence of granulocyte-macrophage-colony-stimulating factor and interleukin-4. Different dose schedules of iDCs were tested for tolerance induction in CBA (H-2k) mice. Tolerance induction by UVB-irradiated iDCs and mDCs was compared. Tolerance induction in two additional strains of mice (C3H and C57BL/6) was also studied. The induction of tolerance was tested by challenging transfusions of treated mice with PBMNCs from the donor strain. The induction of negative regulatory CD4+ T cells in tolerized mice was examined. RESULTS: Four weekly transfusions of 5 x 10(4) UVB-irradiated iDCs from Balb/c mice efficiently induced immune tolerance in CBA and C3H mice. iDCs and mDCs without UVB irradiation could not induce immune tolerance. Tolerance induced by transfusions of UVB-irradiated iDCs was associated with the development of CD4+-regulatory T cells as demonstrated by an adoptive transfer study. CONCLUSION: Transfusion of UVB-iDCs induces immune tolerance across the MHC barrier in certain combinations of mouse strains. The results support the idea that iDCs irradiated with UVB may be applied to induce immune tolerance across the MHC barrier. PMID- 15660826 TI - Impact of donor- and collection-related variables on product quality in ex utero cord blood banking. AB - BACKGROUND: Optimizing product quality is a current focus in cord blood banking. This study evaluates the role of selected donor- and collection-related variables. STUDY DESIGN AND METHODS: Retrospective review was performed of cord blood units (CBUs) collected ex utero between February 1, 2000, and February 28, 2002. Preprocessing volume and total nucleated cell (TNC) counts and postprocessing CD34 cell counts were used as product quality indicators. RESULTS: Of 2084 CBUs, volume determinations and TNC counts were performed on 1628 and CD34+ counts on 1124 CBUs. Mean volume and TNC and CD34+ counts were 85.2 mL, 118.9 x 10(7), and 5.2 x 10(6), respectively. In univariate analysis, placental weight of greater than 500 g and meconium in amniotic fluid correlated with better volume and TNC and CD34+ counts. Greater than 40 weeks' gestation predicted enhanced volume and TNC count. Cesarean section, two- versus one-person collection, and not greater than 5 minutes between placental delivery and collection produced superior volume. Increased TNC count was also seen in Caucasian women, primigravidae, female newborns, and collection duration of more than 5 minutes. A time between delivery of newborn and placenta of not greater than 10 minutes predicted better volume and CD34+ count. By regression analysis, collection within not greater than 5 minutes of placental delivery produced superior volume and TNC count. CONCLUSION: Donor selection and collection technique modifications may improve product quality. TNC count appears to be more affected by different variables than CD34+ count. PMID- 15660827 TI - The influence of human platelet antigen match on the success of allogeneic peripheral blood progenitor cell transplantation following a reduced-intensity conditioning regimen. AB - BACKGROUND: Allogeneic transplantation in elderly patients requires a dose reduced conditioning regimen. Owing to reduced-intensity conditioning, host- and donor-type immune responses may affect the early posttransplant period, whereas only later on donor-derived reactions may ensue. Mismatches in the HLA system are known to be detrimental for the outcome of transplantation. Mismatches between donor and recipient for human platelet antigens (HPAs) may also affect the success of transplantation owing to serving as minor histocompatibility antigens and therefore rendering recipients at risk for graft-versus-host disease (GVHD) or graft rejection and inhibition of thrombopoiesis attributed to platelet (PLT) antibodies. PATIENTS AND METHODS: Therefore, the occurrence of GVHD, incidence of relapse, need of PLT support, and outcome by analysis of 45 donor-recipient pairs for HPA-1, -2, -3, and -5 allotypes and screening for PLT antibodies were evaluated before transplantation and again 1 year thereafter. RESULTS: Mismatches within the HPA system were not associated with an increased occurrence of transplant-related mortality or GVHD, the onset of thrombopoiesis, the frequency of PLT transfusions, or the incidence of relapse. Neither were settings of homozygous donors versus heterozygous recipients (graft-vs.-host direction) nor homozygous recipients versus heterozygous donors (host-vs.-graft direction) associated with any adverse effects on the outcome of the transplantation. CONCLUSION: Thus, the HPA match does not affect the outcome of transplantation after reduced-intensity conditioning. PMID- 15660828 TI - Composition of the blood sampled from surgical drainage after joint arthroplasty: quality of return. AB - BACKGROUND: The quality of blood obtained with a device for postoperative blood drainage and autotransfusion (Bellovac-ABT, Astra Tech AB) was investigated in 50 patients who underwent total hip replacement (THR) or total knee replacement (TKR) surgeries. STUDY DESIGN AND METHODS: The Bellovac-ABT drainage set was inserted and blood collection began after skin closure (THR) or 15 minutes after tourniquet deflation (TKR). A collecting bag stayed in place for less than 6 hours. The product was then returned after sedimentation and discard of supernatant, without anticoagulants. RESULTS: Blood returned with the Bellovac ABT had optimal hemoglobin levels and red blood cell (RBC) counts; the low platelet count reduced the risk of disseminated intravascular coagulation. Inertia of materials maintained acceptable values of C-reactive protein, whereas cytokines and complement split products rose rapidly. Higher concentrations of adenosine triphosphate and 2,3-diphosphoglycerate than in circulating blood suggested a normal RBC metabolism. No adverse reactions were observed in any participants. CONCLUSION: Because of its simplicity and safety, the Bellovac-ABT autologous blood transfusion system is recommended in THR or TKR without joint infection or malignancy. PMID- 15660829 TI - Red blood cell exchange transfusion in two patients with advanced erythropoietic protoporphyria. AB - BACKGROUND: Erythropoietic protoporphyria (EPP) is a rare, autosomal dominant genetic disorder caused by the decreased or absent activity of ferrochelatase, the final enzyme in the heme biosynthetic pathway. This enzyme defect in peripheral blood progenitor cells leads to the accumulation of protoporphyrin deposits in multiple tissues. Plasmapheresis has been previously reported as an adjunctive therapy for patients with advanced hepatic EPP. Because the concentration of protoporphyrins is severalfold higher inside the red blood cell (RBC) compared to plasma, it was hypothesized that RBC exchange therapy might absorb excess protoporphyrins from the plasma and serve as an effective therapy to reduce protoporphyrin load in patients with advanced hepatic EPP. The effectiveness of RBC exchange plus hematin versus plasmapheresis plus hematin in two patients with advanced hepatic EPP is reported. STUDY DESIGN AND METHODS: Two patients with advanced hepatic EPP were treated with RBC exchange and plasmapheresis in the setting of recurrent disease in the graft (Patient 1) or preparation for liver transplantation (Patient 2). In vitro studies were performed to test transport of protoporphyrins from patients' plasma to normal RBCs. RESULTS: Compared with plasmapheresis, RBC exchange was more effective, for the duration of the therapy, in reducing blood levels of protoporphyrins. Liver function tests, however, showed only a modest improvement during therapy. In vitro extracellular protoporphyrin were rapidly adsorbed into normal RBCs. CONCLUSION: Neither RBC exchange nor plasmapheresis prevented progressive hepatic deterioration in advanced hepatic EPP despite a significant decrease in protoporphyrin levels. PMID- 15660830 TI - Prolonged platelet preservation by transient metabolic suppression. AB - BACKGROUND: In this study whether metabolic suppression can be used to preserve platelet (PLT) function during prolonged storage was investigated. STUDY DESIGN AND METHODS: Washed human PLTs were incubated without glucose and with antimycin A to block energy generation. Metabolic suppressed PLTs (MSPs) were stored for 72 hours at different temperatures to find the optimal storage temperature. Controls were incubated with 5 mmol per L glucose and stored at 22 and 4 degrees C. RESULTS: Following metabolic recovery with glucose, MSPs stored at 37, 22, and 4 degrees C showed an increase in basal P-selectin expression (PSE) reaching greater than 40 percent after about 2, 20, and 48 hours; a decrease in thrombin receptor-activating peptide SFLLRN (TRAP)-induced PSE inversely related to the increase in basal PSE; and a decrease in TRAP-induced aggregation reaching less than 30 percent after about 4, 24, and more than 72 hours. When compared with control suspensions, MSPs stored at 4 degrees C better preserved a low basal PSE and in addition showed a better adhesion to surface coated-von Willebrand factor and fibrinogen in a flow chamber. CONCLUSION: Metabolic suppression before storage at 4 degrees C contributes to better preservation of PLT function. PMID- 15660831 TI - Preparation and storage of white blood cell-reduced split apheresis platelet concentrates for pediatric use. AB - BACKGROUND: White blood cell (WBC) reduction and bacterial screening induce unacceptable product loss when platelet (PLT) concentrates (PCs) for pediatric transfusion are prepared from whole blood. The aim was to investigate PCs, WBC reduced and bacterially screened, from single-donor apheresis procedures, divided in 3 or 4 pediatric units and stored up to 5 days. STUDY DESIGN AND METHODS: PCs were collected with an apheresis machine and WBC reduced by in-process filtration. The PCs were sampled for bacterial screening and subsequently divided in 70-mL products. Initially, storage characteristics of split units in 400-mL polyvinylchloride (PVC) bags with 17 split PCs originating from five apheresis donations were studied. When a 600-mL container made of the more gas-permeable polyolefin became available, a paired comparison was performed with 9 split PCs from nine donations and with a higher-yield PLT collection procedure. RESULTS: Split PCs contained 69 x 10(9) +/- 14 x 10(9) PLTs in 69 +/- 1 mL of plasma, and storage in the PVC containers gave a pH value of 6.86 +/- 0.10 on Day 6 (mean +/- SD, n = 17). When comparing the containers, the PVC bag contained 98 x 10(9) +/- 15 x 10(9) PLTs in 72 +/- 4 mL versus 102 x 10(9) +/- 18 x 10(9) PLTs in 74 +/- 8 mL for the polyolefin bag (n = 9, not significant). This gave pH values on Day 6 of 6.12 +/- 0.50 in the PVC container, whereas pH remained acceptable in the polyolefin container: 6.85 +/- 0.10 on Day 6 (p < 0.01). CONCLUSION: PCs for pediatric use from split single-donor apheresis concentrates, WBC reduced and bacterially screened, can be stored for up to 5 days in a 600-mL polyolefin container with maintenance of good in vitro storage variables. PMID- 15660832 TI - Low propidium iodide intensity in flow cytometric white blood cell counting as a marker of cell destruction? AB - BACKGROUND: Residual white blood cells (WBC) in filtered blood products were investigated with flow cytometry. Frequently two distinct populations with different propidium iodide (PI) intensities can be found. The aim of this study was to specify a population with low PI intensity and discuss it as a marker of ongoing cell destruction and their possible impact on cytomegalovirus safety. STUDY DESIGN AND METHODS: Buffy coat-depleted red blood cells were filtered with an in-line filtration set (LCR5, MacoPharma) after 4 hours (LCR5/4 hr) and 16 hours (LCR5/16 hr) of storage, and whole blood was filtered with a whole-blood filtration set (LST1, MacoPharma [LST1/4 hr]). The population with low PI intensity was sorted with a flow cytometer and prepared for transmission electron microscopy. RESULTS: The absolute count obtained in the low-PI-intensity area before filtration was significantly different comparing LCR5/4 hr (11.5 x 10(6) +/- 6.84 x 10(6) and 0.12 x 10(6) +/- 0.1 x 10(6)/unit) and LCR5/16 hr (69.3 +/- 42.12 and 0.06 +/- 0.05; p < 0.002). By use of LST1/4 hr no difference was found compared to LCR5/4 hr after filtration (0.12 +/- 0.09 vs. 0.12 +/- 0.1), but a significant difference was found when comparing the results before filtration (1.25 +/- 0.41 vs. 11.5 +/- 6.84; p < 0.02). Electron microscopy revealed that the sorted population consisted of predominantly cell and nuclear fragments. CONCLUSIONS: Events found in the low-PI-intensity area are not WBCs but partially degraded DNA coming from ongoing cell destruction during extended storage. Our results provide evidence that the absolute count of events found in the low-PI intensity area can be used as a semiquantitative marker of WBC destruction. PMID- 15660833 TI - Characterization of complement receptor 1 domains for prevention of complement mediated red cell destruction. AB - BACKGROUND: Complement activation resulting in intravascular hemolysis can cause transfusion-associated mortality. We recently showed that a recombinant soluble form of complement receptor 1 (CR1) effectively reduces complement-mediated red blood cell (RBC) destruction in vitro and more importantly prolongs the survival of transfused human RBCs in mice. To determine CR1-active sites that prevent RBC destruction, structure-function analysis of its extracellular 1930-amino-acid domain has been performed. STUDY DESIGN AND METHODS: Several CR1-truncated soluble proteins were prepared and tested for their ability to prevent complement mediated RBC destruction in vitro and in mice. RESULTS: A 250-amino-acid region in CR1 that possesses antihemolytic activity and is effective in prolonging survival of transfused RBCs in vivo was identified. Mutation of two critical residues (D109N and E116K) in this 250-amino-acid domain, previously shown to improve complement-inhibitory functions of CR1 derivatives, resulted in a more potent inhibition of complement activation in vitro. In vivo, however, the activity of mutant proteins was comparable to the wild-type molecules. CONCLUSION: Our structure-function studies have characterized smaller CR1-based complement inhibitors for future development of structure-derived transfusion therapeutics. Our studies underscore the importance of testing CR1 inhibitors in vivo. PMID- 15660834 TI - STAR: a novel high-prevalence antigen in the Scianna blood group system. AB - BACKGROUND: More than 20 years ago, a proband was described whose red blood cells (RBCs) typed Sc:1,-2,3. His serum sample contained an immunoglobulin G alloantibody that reacted with all RBCs tested except his own, his brother's, and those with the Sc:-1,-2 phenotype. Cloning of the SC gene allowed determination of the molecular basis associated with this novel high-prevalence antigen. STUDY DESIGN AND METHODS: Samples from frozen storage were obtained from the proband, his serologically matched brother, and 15 serologically mismatched family members. DNA was extracted, and amplified products from all 11 SC (ERMAP) exons and their flanking regions of the proband were sequenced. RESULTS: A single nucleotide mutation was detected (139G>A) in Exon 3 that is predicted to encode a change of Amino Acid 47 from glutamic acid to lysine. The sequence analyses on samples from family members were as expected. CONCLUSIONS: The absence of the high-prevalence antigen STAR detected by the proband's antibody is likely associated with lysine at Position 47 of the Sc glycoprotein. This amino acid change is located on the extracellular portion of HERMAP, 10 residues upstream from the polymorphism associated with Sc1 and Sc2 (Gly57Arg). STAR expands the Sc blood group system to five antigens and has been assigned the ISBT Number 013005 (SC5). PMID- 15660835 TI - Large-volume leukapheresis yields more viable CD34+ cells and colony-forming units than normal-volume leukapheresis, especially in patients who mobilize low numbers of CD34+ cells. AB - BACKGROUND: Large-volume leukapheresis (LVL) differs from normal-volume leukapheresis (NVL) by increased blood flow and altered anticoagulation regimen. LVL is now regarded as a safe procedure for collection of peripheral blood progenitor cells (PBPCs), but it is not known whether the procedure will alter CD34+ cell quality or will be useful for patients who mobilize few CD34+ cells into peripheral blood. STUDY DESIGN AND METHODS: The results from 82 LVL and 125 NVL (4.0-5.3 and 2.7-3.5 times the patients' blood volumes processed, respectively) were retrospectively analyzed in altogether 112 consecutive patients with malignant diseases. RESULTS: The LVL yielded significantly more CD34+ cells (4.2 x 10(6) vs. 3.1 x 10(6)/kg, p = 0.006, all patients; and 1.8 x 10(6) vs. 1.3 x 10(6)/kg, p = 0.004, bad mobilizers) and significantly higher colony-forming units (77 x 10(4) vs. 33 x 10(4)/kg; all patients and 33 x 10(4) vs. 20 x 10(4)/kg, p < 0.001, both groups). Significantly fewer leukapheresis procedures were required to obtain 2 x 10(6) CD34+ cells per kg (one vs. two, p = 0.001, all patients; and two vs. three, p = 0.009, bad mobilizers). No significant differences in CD34+ cell viability and time to hematologic recovery were observed between the patients who received PBPCs harvested by NVL and LVL. CONCLUSION: Although a median platelet loss of 36 percent can be expected, LVL can be recommended as the standard apheresis method for PBPC collections in patients with malignant diseases. LVL is particularly useful in patients who mobilize a low number of CD34+ cells into the peripheral blood. PMID- 15660836 TI - A new strategy for estimating risks of transfusion-transmitted viral infections based on rates of detection of recently infected donors. AB - BACKGROUND: Estimates for human immunodeficiency virus (HIV)-1 and hepatitis C virus (HCV) transfusion-transmitted risks have relied on incidence derived from repeat donor histories and imprecise estimates for infectious, preseroconversion window periods (WPs). STUDY DESIGN AND METHODS: By use of novel approaches, WPs were estimated by back-extrapolation of acute viral replication dynamics. Incidence was derived from the yield of viremic, antibody-negative donations detected by routine minipool nucleic acid testing (MP-NAT) of 37 million US donations (1999-2002) or from sensitive/less-sensitive HIV-1 enzyme immunoassay (S/LS-EIA) results for seropositive samples from 6.5 million donations (1999). Incidences and WPs were combined to calculate risks and project yield of individual donation (ID)-NAT. RESULTS: The HIV-1 WP from presumed infectivity (1 copy/20 mL) to ID-NAT detection was estimated at 5.6 days, and the periods from ID to MP-NAT detection and from MP-NAT to p24 detection at 3.4 and 6.0 days, respectively; corresponding estimates for HCV were 4.9, 2.5, and 50.9 days (the latter represents period from MP-NAT to HCV antibody detection). The HIV-1 incidence projected from MP-NAT yield or from S/LS-EIA data was 1.8 per 100,000 person-years, resulting in a corresponding HIV-1 transfusion-transmitted risk of 1 in 2.3 million. The HCV incidence from MP-NAT yield was 2.70 per 100,000 person years with a corresponding risk of 1 in 1.8 million donations. Conversion from MP NAT to ID-NAT was projected to detect two to three additional HIV-1 and HCV infectious units annually. CONCLUSIONS: MP-NAT yield and S/LS-EIA rates can accurately project transfusion risks. HCV and HIV-1 risks, currently estimated at 1 per 2 million units, could be reduced to 1 in 3 to 4 million units by ID-NAT screening. PMID- 15660837 TI - Evaluation of the Scansystem method for detection of bacterially contaminated platelets. AB - BACKGROUND: Platelet (PLT) bacterial contamination occurs in approximately 1 in 2000 PLT units. The College of American Pathologists recommends and AABB requires procedures to detect PLT bacterial contamination. Although two methods, BacT/ALERT (bioMerieux) and Pall BDS (Pall Corporation), have FDA approval for quality control testing, additional methods are in development. One such method was evaluated, the Scansystem (Hemosystem), which has been developed for use on leukoreduced PLT components between 30 and 72 hours after collection. STUDY DESIGN AND METHODS: Leukoreduced, single-donor apheresis PLT units (LR-SDPs) were inoculated with 10 bacterial species (low and high inocula) associated with PLT contamination. Bacterial detection was compared with the Scansystem and BacT/ALERT. Testing was initiated (10 replicates performed) when LR-SDPs were experimentally inoculated with bacteria. The Scansystem was evaluated 30 hours later, the shortest manufacturer recommended time after PLT collection. RESULTS: All replicates were positive with the Scansystem at 30 hours and with the BacT/ALERT, at 9.3 to 24.0 hours after inoculation. The Scansystem detected bacteria in 83 of 200 replicates (42%) at the time of inoculation indicating a potential for earlier application. CONCLUSIONS: The Scansystem, used to test LR SDPs 30 hours after bacterial inoculation, detected all 20 replicates with a sensitivity equal to the BacT/ALERT system. Based on use of Scansystem with LR SDPs 30 hours after collection and the BacT/ALERT being inoculated 24 hours after collection and incubated for an additional 24 hours before being determined to be negative, the Scansystem will potentially provide results at an earlier time point (32 hr) than provided by the BacT/ALERT system (48 hr). PMID- 15660838 TI - Arteriovenous fistula following needle puncture for blood donation. PMID- 15660839 TI - On the mechanism of staphylococcal protein A immunomodulation. PMID- 15660840 TI - Protein C, protein S, and antithrombin III in twice-frozen plasma. PMID- 15660841 TI - Toxoplasma gondii antibodies in Mexican blood donors. PMID- 15660843 TI - Functional fixedness in a technologically sparse culture. AB - Problem solving can be inefficient when the solution requires subjects to generate an atypical function for an object and the object's typical function has been primed. Subjects become "fixed" on the design function of the object, and problem solving suffers relative to control conditions in which the object's function is not demonstrated. In the current study, such functional fixedness was demonstrated in a sample of adolescents (mean age of 16 years) among the Shuar of Ecuadorian Amazonia, whose technologically sparse culture provides limited access to large numbers of artifacts with highly specialized functions. This result suggests that design function may universally be the core property of artifact concepts in human semantic memory. PMID- 15660844 TI - Constraining theories of embodied cognition. AB - Influences of perceptual and motor activity on evaluation have led to theories of embodied cognition suggesting that putatively complex judgments can be carried out using only perceptual and motor representations. We present an experiment that revisited a movement-compatibility effect in which people are faster to respond to positive words by pulling a lever than by pushing a lever and are faster to respond to negative words by pushing than by pulling. We demonstrate that the compatibility effect depends on people's representation of their selves in space rather than on their physical location. These data suggest that accounting for embodied phenomena requires understanding the complex interplay between perceptual and motor representations and people's representations of their selves in space. PMID- 15660845 TI - Superior performance of blind compared with sighted individuals on bimanual estimations of object size. AB - Five preliminary experiments on sighted individuals revealed marked overestimation on an object size-estimation task using a bimanual response. These experiments ruled out the possibility that overestimation was due to the mode of visual presentation (whether two-dimensional or three-dimensional), the input modality (visual or kinesthetic), or the influence of other visual cues. The main experiment then investigated whether these distortions are due to visual experience by using a variant of the same task to test 24 blind and 24 sighted control participants. Remarkably, the sighted control participants overestimated object size, on average, but the blind participants did not. A follow-up experiment demonstrated that visual memory was the primary influence causing the size over-estimations. We conclude that blind individuals are more accurate than sighted individuals in representing the size of familiar objects because they rely on manual representations, which are less influenced by visual experience than are visual memory representations. PMID- 15660846 TI - The left-to-right bias in inhibition of return is due to the direction of reading. AB - Previous research has shown a left-to-right bias in the inhibition-of-return effect. This bias was found in a sample of English-speaking participants who read in a predominantly left-to-right manner. The goal of the current study was to examine the role that the direction of text reading plays in this bias. The findings replicated the left-to-right bias with an English sample, but showed the opposite bias in an Arabic sample, who read text from right to left. Thus, the regularity of shifting attention in a particular way during text reading seems to be the cause of the bias observed. PMID- 15660847 TI - Mismaking memories: neural precursors of memory illusions in electrical brain activity. AB - Memory illusions--vivid experiences of events that never occurred--could result from inaccuracies either in retrieving memories or in initially storing them. In two experiments, people studied lists of associated words that either did or did not induce later illusory (false) memories of associated but nonpresented lure words. The amplitude of the electrical brain activity during study of words (approximately 500-1,300 ms) that were themselves later correctly remembered reliably distinguished list words that led to such illusory memories from those that did not. This encoding difference associated with subsequent illusory memory (referred to as a DIM)--presumably reflecting item-specific encoding differences- is a neural precursor of memory illusions. PMID- 15660848 TI - Emotion, memory, and attention in the taboo Stroop paradigm. AB - This study tested the binding hypothesis: that emotional reactions trigger binding mechanisms that link an emotional event to salient contextual features such as event location, a frequently recalled aspect of naturally occurring flash bulb memories. Our emotional events were taboo words in a Stroop color-naming task, and event location was manipulated by presenting the words in different task-irrelevant screen locations. Seventy-two participants named the font color of taboo and neutral words, with instructions to ignore word meaning; in one condition, several words were location consistent (i.e., always occupied the same screen location), whereas in another condition, several colors were location consistent. Then, in a surprise recognition memory test, participants recalled the locations of location-consistent words or colors. Although attention enhanced overall location memory for colors (the attended dimension during color naming), emotion (taboo vs. neutral words) enhanced location memory for words but not colors. These results support the binding hypothesis but contradict the hypothesis that emotional events induce image-like memories more often than non emotional events. PMID- 15660849 TI - Traumatic impact predicts long-term memory for documented child sexual abuse. AB - Prospective studies of adults' memories of documented child sexual abuse (CSA) reveal that the majority of individuals remember their victimization. However, the accuracy of these memories has rarely been investigated scientifically. The present study examined predictors of memory accuracy and errors 12 to 21 years after abuse ended for individuals with legal experiences resulting from documented CSA. Severity of posttraumatic stress disorder (PTSD) symptomatology was positively associated with memory accuracy. However, individuals nominating CSA as their most traumatic life event exhibited relatively accurate memory regardless of indicators of PTSD. Predictors of memory errors were also identified (e.g., less maternal support). These results indicate that, in addition to understanding the role of traditional cognitive factors, understanding an event's traumatic impact is important for predicting the accuracy of long-term memory for reported CSA. PMID- 15660850 TI - Developments in declarative memory. AB - The second year of life is marked by changes in the robustness of recall memory. Both retrieval and storage processes have been implicated as the major source of age-related improvements in recall. Children 13 to 20 months of age were matched for levels of learning of laboratory events (thereby eliminating encoding as a source of developmental difference) and tested for recall after delays as long as 6 months. In Experiment 1, 16-month-olds evidenced less loss of information and more relearning than 13-month-olds. In Experiment 2, 20-month-olds evidenced less loss of information and more relearning than 16-month-olds. Patterns of performance across test trials and in relearning implicate a decline in susceptibility to storage failure as the primary source of the observed developmental trend. PMID- 15660851 TI - Metrical categories in infancy and adulthood. AB - Intrinsic perceptual biases for simple duration ratios are thought to constrain the organization of rhythmic patterns in music. We tested that hypothesis by exposing listeners to folk melodies differing in metrical structure (simple or complex duration ratios), then testing them on alterations that preserved or violated the original metrical structure. Simple meters predominate in North American music, but complex meters are common in many other musical cultures. In Experiment 1, North American adults rated structure-violating alterations as less similar to the original version than structure-preserving alterations for simple meter patterns but not for complex-meter patterns. In Experiment 2, adults of Bulgarian or Macedonian origin provided differential ratings to structure violating and structure-preserving alterations in complex- as well as simple meter contexts. In Experiment 3, 6-month-old infants responded differentially to structure-violating and structure-preserving alterations in both metrical contexts. These findings imply that the metrical biases of North American adults reflect enculturation processes rather than processing predispositions for simple meters. PMID- 15660852 TI - Controlling racial prejudice: social-cognitive goals affect amygdala and stereotype activation. AB - The malleability of stereotyping matters in social psychology and in society. Previous work indicates rapid amygdala and cognitive responses to racial out groups, leading some researchers to view these responses as inevitable. In this study, the methods of social-cognitive neuroscience were used to investigate how social goals control prejudiced responses. Participants viewed photographs of unfamiliar Black and White faces, under each of three social goals: social categorization (by age), social individuation (vegetable preference), and simple visual inspection (detecting a dot). One study recorded brain activity in the amygdala using functional magnetic resonance imaging, and another measured cognitive activation of stereotypes by lexical priming. Neither response to photos of the racial out-group was inevitable; instead, both responses depended on perceivers' current social-cognitive goal. PMID- 15660853 TI - Reaction time explains IQ's association with death. AB - Lower IQ is associated with earlier death, but the cause of the relationship is unknown. In the present study, psychometric intelligence and reaction times were both significantly related to all-cause mortality in a representative sample of 898 people aged 56 years who were followed up with respect to survival until age 70. The association between IQ and mortality remained significant after adjusting for education, occupational social class, and smoking, all of which have been hypothesized as confounding variables. The effect of IQ on mortality was not significant after adjusting for reaction time, suggesting that reduced efficiency of information processing might link lower mental ability and earlier death. This new field of cognitive epidemiology provides arguably the strongest evidence for the importance of psychological factors in physical health and human survival. Finding the mechanisms that relate psychometric intelligence to mortality might help in formulating effective interventions to reduce inequalities in health. PMID- 15660854 TI - How many variables can humans process? AB - The conceptual complexity of problems was manipulated to probe the limits of human information processing capacity. Participants were asked to interpret graphically displayed statistical interactions. In such problems, all independent variables need to be considered together, so that decomposition into smaller subtasks is constrained, and thus the order of the interaction directly determines conceptual complexity. As the order of the interaction increases, the number of variables increases. Results showed a significant decline in accuracy and speed of solution from three-way to four-way interactions. Furthermore, performance on a five-way interaction was at chance level. These findings suggest that a structure defined on four variables is at the limit of human processing capacity. PMID- 15660855 TI - Relational and arelational confidence intervals: a comment on Fidler, Thomason, Cumming, Finch, and Leeman (2004). PMID- 15660856 TI - Blindness for unchanging targets in the absence of motion filtering: a response to Theeuwes (2004). PMID- 15660857 TI - Do dogs resemble their owners? A reanalysis of Roy and Christenfeld (2004). PMID- 15660858 TI - Patients with benign rolandic epilepsy have a longer duration of somatosensory evoked high-frequency oscillations. AB - BACKGROUND: High-frequency oscillations (HFO) ranging between 300-900 Hz have been shown to be superimposed on an early component of somatosensory evoked potentials (SEP) to median nerve stimulation in humans. Although the HFO are speculated to be a localized activity of the GABAergic inhibitory interneurons, the significance in the epileptogenicity remains unclear. The authors of this study analyzed HFO using magnetoencephalography in patients with benign rolandic epilepsy (BRE) to clarify the neurophysio-logical basis of rolandic discharges (RD). METHODS: Nine patients with BRE and six patients with other epileptic syndrome (non-BRE) participated in the study. Somatosensory evoked fields (SEF) including HFO to median nerve stimulation were measured in a magnetically shielded room with a 37-channel neuromagnetometer. RESULTS: Two kinds of HFO, 300 Hz- and 600 Hz-HFO, were identified and the duration of the HFO in patients with BRE was significantly longer than that in patients with non-BRE. CONCLUSIONS: The results suggest that the longer part of HFO (P30m-related) is closely related to the pathogenesis of RD and that the longer HFO in patients with BRE might be mediated by altered GABAergic inhibition modulated by the cholinergic system. PMID- 15660859 TI - Oral iron supplementation in preterm infants treated with erythropoietin. AB - BACKGROUND: It is not known whether a moderate dose of oral iron supplementation would further enhance erythropoiesis in recombinant human erythropoietin (EPO) treated very low-birthweight (VLBW) infants. METHODS: In total, 24 preterm infants with birthweights 750-1499 g were enrolled at the age of 14-28 days to receive 400 IU/kg per week EPO subcutaneously for 8 weeks. The infants were randomly allocated either to receive oral iron supplementation 4 mg/kg per day or to serve as controls. RESULTS: Hemoglobin and the absolute reticulocyte count in the iron supplementation and the control groups remained identical throughout the study period, whereas serum ferritin was significantly lower in the control group at study exit and follow up. Rates of treatment success (no need for transfusion and hemoglobin never below 8 g/dL) also did not differ between the groups. CONCLUSIONS: In this study we did not find a clear advantage in a moderate dose of oral iron supplementation on erythropoiesis in EPO-treated VLBW infants. Whether a higher dose would lead to enhanced erythropoiesis remains to be answered. PMID- 15660860 TI - Change of bilirubin photoisomers in the urine and serum before and after phototherapy compared with light source. AB - BACKGROUND: The clinical effect of phototherapy for neonatal hyperbilirubinemia is based on the production and elimination of cyclobilirubin. Generally, the clinical effect of light sources is estimated by the reduction in the total serum bilirubin level. One procedure with less invasiveness than blood collecting is urine collection. Whether the effectiveness of light sources used for phototherapy could be assessed using measurements of bilirubin photoisomers in urine was studied. METHODS: This study was a retrospective analysis of 38 term infants with hyperbilirubinemia who underwent phototherapy. Bilirubin fractions in serum and urine before and 24 h after the phototherapy were measured by high performance liquid chromatography. The light sources used for the phototherapy were blue-white light (n = 11), Biliblanket plus high output (n = 13) or green light (n = 14). The relationships between serum and urine bilirubin photoisomers after phototherapy and whether the levels of urine bilirubin photoisomer are affected by the light sources with different wavelength characteristic were analyzed. RESULTS: There was no correlation between serum (ZE)-bilirubin and urine configurational isomers, but a weak positive correlation between serum (EZ) cyclobilirubin and urine structural isomers after phototherapy. Although serum (ZE)-bilirubin levels depended on the wavelength characteristic of each light source during phototherapy, the urine configurational isomer levels did not depend on it. The increase in serum (EZ)-cyclobilirubin levels and the urine structural isomer levels were mostly in agreement. CONCLUSIONS: The urine bilirubin structural isomers may be used to estimate the serum (EZ) cyclobilirubin levels and to evaluate the clinical effects of light sources. PMID- 15660861 TI - Dosage regimen of arbekacin for methicillin-resistant Staphylococcus aureus infection in newborns and infants. AB - BACKGROUND: Arbekacin (ABK) is an aminoglycoside antibiotic that has a dose dependent bactericidal action. Because it inhibits the production of toxic shock syndrome toxin-1 (TSST-1) by methicillin-resistant Staphylococcus aureus (MRSA), it has attracted attention as a therapeutic drug for MRSA infection. In this study, the authors investigated the pharmacokinetics of ABK based on therapeutic drug monitoring (TDM), in order to establish an effective dosage regimen with minimal adverse reactions in MRSA infected newborns and infants. METHODS: Arbekacin was administered to nine MRSA infected newborns and infants between October 2000 and March 2002. Following the initial ABK administration, blood was collected and the blood concentration of ABK was measured. The blood concentrations of ABK were analyzed by the two-compartment model or by a model independent method in order to elucidate the pharmacokinetics of ABK. Pharmacokinetic analysis was performed using WinNonlin Professional V3.1. RESULTS: The mean age at initial ABK administration was 24.0 +/- 26.0 days (postconceptional age: 39.2 +/- 3.9 weeks). The increase in the peak blood concentration of ABK was 2.40 +/- 0.20 microg/mL per mg ABK per kg bodyweight, showing great consistency among cases. The elimination half-life of ABK was 0.22 3.52 h in the alpha phase (T(1/2alpha)) and 2.42-33.44 h in the beta phase (T(1/2beta)), showing great variation among cases. The distribution volume was 0.75 +/- 0.13 L/kg, and systemic clearance was 0.054 +/- 0.012 L/h/kg. ABK alleviated clinical symptoms and inflammations in all cases. CONCLUSION: Nine newborns and infants with MRSA infection and various underlying diseases were successfully treated with TDM-based administration of ABK with no severe adverse reactions. PMID- 15660862 TI - Safety and efficacy of acellular pertussis vaccine in Japan, evaluated by 23 years of its use for routine immunization. AB - BACKGROUND: Real evaluation of any vaccine can only be done after the vaccine has been in routine use for a substantially long period of time. In Japan, acellular pertussis vaccine was introduced and totally replaced whole cell pertussis vaccine in 1981. From 1982 to 1988 40.3 million doses of acellular pertussis vaccine were given to 2-year-olds and from 1989 to 2001 59.3 million doses of acellular pertussis vaccine were given to 3-month-olds. It is now time to evaluate the efficacy and safety of acellular pertussis vaccine by the use of national data officially supplied by the Government. METHODS: Government national surveillance of pertussis, which began in 1981, was used to analyze epidemiology of pertussis. Official Government reports on acceptance rates of pertussis were analyzed. A peer review has been made on all severe neurological illnesses/death occurring after pertussis immunization which have been applied for through the Vaccine Injury Compensation System, Ministry of Health Labor and Welfare, Japan. RESULTS: High acceptance rates of acellular pertussis vaccine combined with diphtheria and tetanus toxoids (DTaP) has been maintained and a dramatic decrease in pertussis was noted over the past 23 years. Neurological illnesses temporally associated both with whole cell and with acellular pertussis vaccination has been a rare phenomenon. However, incidences of encephalopathy/encephalitis and status epileptics/frequent convulsions, febrile seizures/provocation of convulsions, and sudden deaths were significantly lower with acellular pertussis vaccination than with whole cell pertussis vaccination. CONCLUSION: With the use of acellular pertussis vaccine which has been accepted by the public, pertussis has been well controlled in Japan. PMID- 15660863 TI - Intestinal parasitic infection in children in post-disaster situations years after earthquake. AB - BACKGROUND: Two earthquakes in the north-west region of Turkey destroyed 80% of the houses and schools in Duzce in 1999. This study was conducted to determine the parasitic infection rate associated with the post-earthquake unhealty living conditions and related epidemiological risk factors. METHODS: Two populations living and studying in different socioeconomic conditions as a result of the earthquake were compared: Group 1 (study group) consisted of 326 children living and studying in transitory houses and classes. Group 2 (control) consisted of 127 children living in normal houses and studying in normal school classes. Fecal samples were obtained from both groups and examined for intestinal parasites' eggs and trofozoid forms. In addition, selotype procedure was applied to both populations. Epidemiological data determining the socioeconomic status of the populations were collected by questionnaire. RESULTS: In group 1, Giardia lamblia (G. lamblia) cysts were observed in 10.4% of the fecal samples and Enterobius vermicularis (E. vermicularis) eggs were observed in 13.5% of selotype samples. In group 2, Giardia cysts were observed in 3.1% of the samples and E. vermicularis eggs were observed in 5.5% of selotype samples. The rate of Giardiasis and Enterobiasis was found to be significantly higher in children still living and studying in temporary houses and schools years after the earthquakes (P < 0.05). The following pidemiological and social factors arising after the earthquakes were associated with increased rate of G. lamblia and E. Vermicularis infections: number of communal toilets per child at school, socioeconomic level of the children, dimensions of the classrooms where the children are studying and living and frequency of hand-washing at school. These parameters are significantly different between the groups (P < 0.05). CONCLUSION: Giardiasis should be considered as an emerging disease in postdisaster situations and adequate prevention measures should be implemented in these circumstances. It should also be known that the rate of Enterobiasis is increased in populations living in crowded unhealthy conditions. PMID- 15660864 TI - Complementary findings in clinical and epidemiologic features of mumps and mumps meningoencephalitis in children without mumps vaccination. AB - BACKGROUND: Mumps vaccine has not yet been included in the routine vaccination programme, for this reason mumps is still one of the most common infections for children in Turkey. One of the major complication of mumps is meningoencephalitis, which although usually heals spontaneously, it may cause neurologic complications. METHODS: This study was undertaken to investigate epidemiologic and demographic characteristics in children with mumps and mumps meningoencephalitis and clinical/laboratory findings in children with mumps meningoencephalitis diagnosed over a 11 year period. A total of 2422 mumps and 135 mumps meningoencephalitis cases were covered in this study which constitutes one of the largest series of mumps meningoencephalitis in the literature. The mean age of mumps and mumps meningoencephalitis cases were 6.6 +/- 2.7 and 7.6 +/ 2.6 years, respectively. RESULTS: There was a male predominance both among the cases of mumps and mumps meningoencephalitis. The age and seasonal distributions were similar in the mumps and mumps meningoencephalitis groups. The most common symptoms of mumps meningoencephalitis were fever (97%), vomiting (94%) and headache (88.8%). The mean cerebrospinal fluid (CSF) total cell count and lymphocyte count were 540 +/- 460/mm(3) and 300 +/- 330/mm(3), respectively. The mean CSF protein and glucose levels were found to be 56.97 +/- 27.94 mg/dL and 53.67 +/- 15.46 mg/dL, respectively. The mean of CSF/blood glucose ratio was 0.53 +/- 0.16. The mean duration of hospitalization in mumps meningoencephalitis cases was found to be 5.1 +/- 2.4 days and this was longer in boys (P < 0.05). CONCLUSION: In patients with mumps meningoencephalitis, higher CSF protein levels and lower CSF glucose/blood glucose ratio were associated with longer hospitalization periods. There was no death. PMID- 15660865 TI - Changes in seroprevalence of hepatitis A in children and adolescents in Manisa, Turkey. AB - BACKGROUND: Hepatitis A virus (HAV), being an enteric transmitting virus wide world, occurs mostly in children of developing countries. However, the virus has recently been seen in adolescents and young adults worldwide. The aim of the present study was to evaluate the seroprevalence of the HAV infection in children and adolescents in Manisa, Turkey, and to verify whether the increased incidence of HAV infection in other parts of the world and Turkey generally is also true for the area of Manisa. METHODS: The authors studied blood samples of 1395 adolescents and children aged between 6 months and 17 years from the Manisa area in order to evaluate the existence of anti-HAV antibodies using micro-ELISA test. RESULTS: Total seropositivity was 44.6% while age related values were as followed: 6-23 months, 47.8%; 2-6 years, 23.7%; 7-10 years, 43.4%; 11-14 years, 52.4%; and 15-17 years, 76.6%. These results suggest that the seropositivity increases significantly among children at school age and also increases parallel to age. CONCLUSIONS: According to these results, the authors concluded that there has been a shift of seropositivity from children to adolescents especially in families with an average or high socio-economical level. PMID- 15660866 TI - Long-term expressed human alpha-galactosidase A in tissues of HalphaG transgenic mice. AB - BACKGROUND: Human alpha-galactosidase A (halphaG) is an essential lysosomal enzyme in catalyzing the hydrolysis of ceramide trihexoside in humans. Effects have been directed to develop effective gene and replacement therapies for the deficiency of halphaG, Fabry disease. In recent years, halphaG transgenic mice (TGM) have been established, and the expression of halphaG in their general organs has been reported. However, detailed distribution of the cells expressing halphaG have not yet been defined. METHODS: The distribution of halphaG in organs of the halphaG-TGM was studied by means of immunohistochemistry and enzyme assay. RESULTS: Immunohistochemical analysis revealed a systematic halphaG expression in the TGM, including endothelial cells of the bone marrow, liver, spleen, pancreas, lungs, uriniferous tubules in the kidneys, and choroids plexus in the brain. Enzyme assay demonstrated a persistent expression of halphaG in the TGM during 14 20 months after birth. CONCLUSION: A long-term expression of halphaG in organs may indicate halphaG-TGM as a useful tool in the research of gene and replacement therapies for Fabry disease. PMID- 15660867 TI - Improved diabetes control by using 'close adjustment algorithms'. AB - BACKGROUND: Intensive insulin therapy increases the frequency of severe hypoglycemia despite markedly improved glycemic control in patients with type 1 diabetes mellitus. To determine the optimal dose of insulin, the authors designed algorithms based on self-monitored blood glucose levels. METHODS: Each dose of insulin was composed of two components: a basal dose determined on the basis of blood glucose levels over the previous two days and an additional dose determined on the basis of blood glucose level just before insulin injection. The patients were instructed to adjust each dose according to the algorithms. The authors investigated the effects of using algorithms on glycemic control, anthropometric data, body composition, and lipid profile in seven females with type 1 diabetes 12-20 years old. RESULTS: After 3 months, the daily dose of insulin increased significantly from 0.93 +/- 0.18 to 1.16 +/- 0.26 units/kg of body weight, and haemoglobin A(1C) decreased significantly from 8.27 +/- 1.33 to 6.50 +/- 0.64%. Severe hypoglycemia, however, did not occur. Body mass index increased significantly from 21.7 +/- 2.7 to 22.7 +/- 2.9 kg/m(2) with no increase in the percentage of body fat. All lipid-profile data showed a decreasing trend. CONCLUSIONS: Algorithms developed on the basis of self-monitored blood glucose levels are useful in determining the optimal dose of insulin and can improve glycemic control and lipid metabolism. PMID- 15660868 TI - Dynamic movement of cytochrome c from mitochondria into cytosol and peripheral circulation in massive hepatic cell injury. AB - BACKGROUND: In the process of apoptosis, it is known that the transition of cytochrome c from mitochondria into the cytosol occurs, and tumor necrosis factor (TNF)-alpha is one of the molecules responsible for this event. But in the state of hypercytokine induced by D-galactosamine (D-GaIN)/Lipopolysaccharide (LPS), the localization of cytochrome c is little known. METHODS: Rats were administrated with D-GaIN(700 mg/kg)/LPS(200 microg/kg). Blood and tissue samples were collected and examined for levels of pro-inflammatory cytokines, the apoptosis of liver cells, and the localization of cytochrome c. RESULT: Before administration of D-GaIN/LPS, cytochrome c was definitely localized in the mitochondria. At 2 h after simultaneous administration of D-GaIN/LPS, cytochrome c had accumulated in the cytosol following abrupt increases of plasma TNF-alpha. Massive cell destruction due to apoptosis proved by Terminal deoxynucleo-tidyl transferase-mediated dUTP nick end labeling staining was observed in liver tissue 4 h later and markedly increased levels of cytochrome c were detected in the plasma 12 h after D-GaIN/LPS administration. CONCLUSION: Liver injury induced by simultaneous administration of D-GaIN/LPS was closely associated with the production of TNF-alpha, and also with the dynamic movement of cytochrome c from the mitochondria into the cytosol, and then into the systemic circulation. The detection of plasma cytochrome c levels may be a useful clinical tool for the detection of apoptosis in vivo. PMID- 15660869 TI - Celiac disease with various presentations. AB - BACKGROUND: Celiac disease (CD) has a wide clinical spectrum from malabsorption syndrome to extra intestinal presentations. A total of 45 children with CD presented with mainly chronic diarrhea (n :23), anemia (n: 12), and short stature (n: 10) were evaluated in this study. The aim was to find common parameters of CD with various presentations. METHODS: Basic anthropometric, biochemical and hematological parameters in cases with CD with various presentations were compared. RESULTS: It was found that children with CD presenting with chronic diarrhea were younger. There was no significant difference in hemoglobin levels in children with CD presenting with anemia. Children with CD with short stature had significantly lower serum vitamin B(12) levels and lower levels of height standard deviation scores, bone age delay, and alkaline phosphatase. CONCLUSIONS: It was concluded that children, especially infants with chronic diarrhea with CD, may not be affected with generalized malabsorption. Anemia and short stature are frequent findings in cases with CD whether they are main presenting symptoms or not. Children with CD presenting with short stature may have lower levels of vitamin B(12) than other presentations. PMID- 15660870 TI - Leptin and the metabolic syndrome in Korean adolescents: factor analysis. AB - BACKGROUND: The objectives of this study were to evaluate the relationships of leptin with the metabolic syndrome and to examine leptin's role in clustering of the metabolic components among Korean adolescents. METHODS: A cross sectional study was carried out in 68 male and 80 female adolescents aged 13-18 years in an urban area of South Korea. Anthropometric variables were measured and blood pressure, fasting plasma glucose, low-density lipoprotein cholesterol, triglyceride, high-density lipoprotein cholesterol, and serum leptin were obtained. RESULTS: As more metabolic components were clustered, body mass index, body fat, waist circumference, and serum leptin levels were significantly increased. Principal components factor analysis revealed three factors in males and females that explained 70% and 65%, respectively, of the observed variance of the 10 measured variables. These were obesity-leptin-lipid factor, blood pressure factor, and glucose-cholesterol factor in males and obesity-leptin-glucose factor, blood pressure factor, and cholesterol factor in females. Leptin loaded on only one factor in both genders. CONCLUSION: Leptin did not appear to have a major role linking various components of the metabolic syndrome, even though it was strongly associated with obesity indices. Gender difference of linking of leptin with glucose or lipid was observed. There seems to be more than one pathophysiological mechanism which might underlie full expression of the metabolic syndrome among Korean adolescents. PMID- 15660871 TI - Usefulness of electron beam computed tomography for quantitative estimation of myocardial ischemia in patients. AB - BACKGROUND: The electron beam computed tomography (EBT) was used as non-invasive methods for estimating mycardial characteristics in patients after onset of Kawasaki disease. METHODS: All of the 22 coronary branches with localized stenosis (LS), and 22 branches without coronary artery lesion after Kawasaki disease were examined for changes of myocardial CT number. In this study, each myocardial region was divided into three parts (endocardial, central, epicardial parts) and the myocardium/lumen (M/L) ratio was calculated. RESULTS: The mean M/L ratio in the endocardial part of early image with LS was significantly lower than the ratios in the other parts of early images with LS (P = 0.0286, P = 0.0069, respectively). and only the mean M/L ratio in the endocardial part was significantly different on images from patients with LS and images from patients without LS (P = 0.0296). Inverse correlations were found in the LS group between coronary artery stenosis rate determined by coronary angiography and M/L ratio in the endocardial part on early images (P = 0.0018, r = -0.614, y = 0.56 -0.605x). CONCLUSIONS: It was possible to quantitatively evalutate myocardial characteristics using EBT. It is suggested that myocardial ischemia progressed from the endocardial region and EBT is useful to detect the endocardial ischemia. We suggested that the rate of coronary stenosis in patients with LS can be estimated by calculating the M/L rations in the endocardial parts of early EBT images. EBT enables accurate evaluation of myocardial characteristics non invasively. PMID- 15660872 TI - Tc-99m-HMPAO SPECT in the diagnosis of brain death in children. AB - BACKGROUND: The diagnosis of brain death is based on both clinical and laboratory findings. However, diagnosis of brain death is still contentious and reliable tests are required. Early recognition and declaration of the diagnosis is the main goal, which is important for discontinuation of life support and organ donation for transplantation. In order to achieve this goal, competent diagnostic procedures should be performed. In this paper the authors review the diagnosis of brain death in eight children from different age groups, with an emphasis on factors concerning the reliability, use and appropriate application time of Tc 99m-HMPAO single photon emission computed tomography (SPECT) in early diagnosis in infants and newborns. METHODS: Eight patients who fulfilled the clinical criteria of brain death underwent Tc-99m HMPAO SPECT and electroencephalogram (EEG) monitoring. RESULTS: All patients had electrocerebral silence on EEG recordings. Six patients showed lack of perfusion in cerebrum in their first SPECT, however, newborns needed a second image for a confirmed diagnosis. CONCLUSION: In infants, SPECT has a high reliability for confirmed diagnosis of brain death; however, in newborns the application time is important for an accurate diagnosis. PMID- 15660873 TI - Environmental mercury exposure in children: South China's experience. AB - BACKGROUND: Environmental mercury levels significantly increased in the past decades following its increase in industrial applications. In spite of an increasing concern on the potential harmful effects of mercury on children, there is no reported data for the Chinese population. The relationship between dietary habit and environmental mercury exposure in Chinese children was studied. METHODS: The hair and blood mercury levels of Chinese children aged above 3 years in 2000 March to September, were studied. Sociodemographic data, dietary habits of the past 6 months, and other risk factors for environmental mercury exposure were collected. Those children with blood mercury levels above the toxic range (i.e. > 45 nmol/L) and their family members were further evaluated and their blood and hair mercury levels were monitored before and after Fishing-Moratorium period (June to August 2000) in South China Sea. RESULTS: Altogether, 137 Chinese children (mean age, 7.2 years) were recruited. The mean hair mercury level was 2.2 p.p.m and the mean blood mercury level was 17.6 nmol/L. There was a strong correlation (r = 0.88) between hair and blood mercury levels in our cohort. Frequency of fish consumption correlated with hair (r = 0.51) and blood (r = 0.54) mercury levels. For those children who consumed fish more than 3 times/week, hair and blood mercury levels were twice as high as those who consumed fish l-3 times/week and threefold of those who never consumed fish. Five children and 12 family members had toxic blood mercury levels. Their blood (P < 0.0001) and hair (P = 0.02) mercury levels dropped significantly after reducing fish consumption during Fishing-Moratorium period. CONCLUSION: Both blood and hair (i.e. Tissue) mercury levels of children in Hong Kong was elevated and correlated with the frequency of fish consumption. PMID- 15660874 TI - Spontaneous closure of an unroofed coronary sinus. PMID- 15660875 TI - Hemolytic crisis with fulminant hepatic failure in Wilson disease without consanguinity. PMID- 15660876 TI - Human herpesvirus 6 fulminant hepatic failure treated by living donor liver transplantation. PMID- 15660877 TI - Multiple fungal splenic abscesses in a patient with T-acute lymphoblastic leukemia undergoing chemotherapy. PMID- 15660878 TI - Primary tuberculous osteomyelitis of the mandible. PMID- 15660879 TI - Subacute vertebral osteomyelitis in a child with diabetes mellitus associated with Fusarium. PMID- 15660880 TI - Severe hyperlipidemia in a child with acute lymphoblastic leukemia treated with L asparaginase and prednisone. PMID- 15660881 TI - Unusual cranial magnetic resonance imaging findings in a case with Sydenham's chorea. PMID- 15660882 TI - Transthoracic echocardiography-assisted valve perforation in pulmonary atresia. PMID- 15660883 TI - A high incidence of scoliosis in children with tension-type headache. PMID- 15660885 TI - Could Vitamin C deficiency have a role in shaken baby syndrome? PMID- 15660887 TI - Contact sensitization in 1094 children undergoing patch testing over a 7-year period. AB - Contact sensitization in children is frequent. However, because exposure to sensitizing agents varies rapidly, it is of utmost importance to perform a periodic evaluation of patch test results. Our purpose was to compare our data on contact sensitization in children during the past 7 years to our previous 1988 1994 findings, in order to identify emerging allergens and update our pediatric series. From 1995 to 2001, 1094 consecutive children were examined. Of these, 997 patients were patch tested with our pediatric series, which includes 30 allergens, whereas 97 underwent patch testing with 46 allergens. A total of 570 children proved allergic (52.1%). The highest sensitization rate was observed in children under 3 years of age. No differences between atopic dermatitis patients and nonatopic ones were observed in the sensitization rate. Neomycin, nickel, wool alcohols, thimerosal, and ammoniated mercury gave most of the positive responses. With respect to 1988-1995 data, allergy to substances such as neomycin, nickel, wool alcohols, thimerosal, ammoniated mercury, propolis, potassium dichromate, and thiuram mix proved more frequent. In conclusion, as sensitization rates to different allergens show great variations over time, periodic evaluations of patch test results in children is necessary in order to update the test trays. PMID- 15660888 TI - Prevalence of skin disorders in school children in Ibadan, Nigeria. AB - Although skin diseases are common in children, there are very few population based studies in West Africa. Recently, there has been considerable emphasis on hygiene and socio-economic impact on the incidence of allergic disorders in children. We determined the prevalence of skin diseases in a public primary school in Ibadan (southwest Nigeria). A questionnaire for assessing factors associated with the prevalence of diseases was completed, and a complete physical examination was carried out on 1066 students. The study included 529 (49.6%) boys and 537 (50.4%) girls with a mean age of 8.8 +/- 2.5 years. The mean family size of the subjects was 6.7 +/- 2.3 while the mean number of rooms in their homes was 2.6 +/- 1.45. Infectious dermatosis was commonly observed. Of 375 children with a skin lesion, 162 (15.2%) had dermatophytosis, most often tinea capitis, 50 (4.7%) had pityriasis versicolor, and 50 (4.7%) had scabies. Other dermatoses observed included papular urticaria in 35 (3.3%) and angular cheilitis in 27 (2.5%) children. One or more melanocytic nevi were found in 40 (3.8%) children while 138 (12.9%) and 77 (7.25%) had tribal and scarification marks, respectively. Atopic eczema and viral warts were virtually absent. We concluded that fungal infections and scabies were the most common skin diseases in our study population, whereas allergic illnesses were nearly absent. PMID- 15660889 TI - Alagille syndrome: cutaneous manifestations in 38 children. AB - The Alagille syndrome is one of the most common inherited disorders causing chronic liver disease during childhood. During the 1990s, 38 children with Alagille syndrome were evaluated at two pediatric centers in Buenos Aires, Argentina. Characteristic clinical, humoral, and cutaneous features were analyzed. The average age of diagnosis was 29 months old (range of between 2 months and 15 years). Cholestasis was evident in 92% of patients during the neonatal period. Family antecedents related to the syndrome were found in 18.5% of the patients. Peculiar facies developed in 85% of patients. Chronic cholestasis and pruritus were observed in all of the patients and jaundice was evident in 78%. Eighty-four percent of the patients had heart disease (pulmonary stenosis, intraauricular communication, intraventricular communication), 76% of them showed growth retardation, and vertebrae abnormalities were found in 63%. Embryotoxon appeared in 76% of patients, and renal disturbances in 21%. Eleven children (28%) had xanthomas, in the neck, elbows, palms, helixes, inguinal area, gluteus, and knees. The earliest findings appeared in the first months of life, and the latest at 5 years of age. The xanthomas located in the folds had a stony aspect. Cholesterol levels ranging from 220 to 1600 mg percentage (mg%) were demonstrated in all of the children with xanthomas. Liver transplantation was performed in seven of the patients (18.4%). Two of them died after this operation. The disappearance of xanthomas after transplantation was remarkable in all of the patients. PMID- 15660890 TI - Primary annular plaque-type psoriasis. AB - We described two adolescent girls with untreated, consistently annular, plaque type psoriasis without pustules, a presentation that is to our knowledge, not previously described. No typical confluent plaque-type lesions were present. The plaques in our patients resembled other entities such as tinea corporis and erythema annulare centrifugum, given the erythematous, scaling borders and central clearing. Biopsy specimens from our patients showed features characteristic of psoriasis vulgaris. Both patients responded to combination therapy with calcipotriene and a mid-potency steroid. We conclude that primary annular plaque-type psoriasis shares features of both typical plaque-type and annular pustular psoriasis, suggesting that these entities represent a spectrum of psoriatic disease. PMID- 15660891 TI - Cutaneous tumors as the initial presentation of non-T, non-B, nonmyeloid CD4+ CD56+ hematolymphoid malignancy in an adolescent boy. AB - We describe a 14-year-old Hispanic boy who presented with a 2-month history of enlarging plum-colored cutaneous tumors on his face, trunk, and proximal extremities. Histopathologic examination showed nodular infiltrates of malignant mononuclear cells extending from the superficial dermis to the deep subcutis. Immunohistochemical staining of the biopsy specimen and flow cytometry studies on a bone marrow aspirate revealed a CD4+, CD56+ hematolymphoid tumor that was negative for all other myeloid and lymphoid markers. Based on this information, the patient was diagnosed with the recently described, rare non-T, non-B, nonmyeloid CD4+ CD56+ hematolymphoid malignancy. To our knowledge, this is the youngest patient reported in the literature. PMID- 15660892 TI - Concurrent mycosis fungoides and precursor B cell lymphoblastic lymphoma in a 6 year-old child. AB - We report a 6-year-old boy with mycosis fungoides on the dorsal surface of his left hand, a diagnosis supported by positive T cell receptor gamma gene rearrangement findings. Precursor B cell lymphoblastic lymphoma of the left orbit developed subsequently. He was treated with chemotherapy and remained under control. PMID- 15660893 TI - Spontaneous regression of aleukemia congenital leukemia cutis. AB - A full-term 2-week-old boy was referred to the pediatric dermatology clinic with numerous blue to violaceous nodules present since birth. TORCH titers (against toxoplasmosis, cytomegalovirus, herpes simplex virus, rubella, and syphilis) were negative. Complete blood count and peripheral smear were normal. A skin biopsy specimen showed an atypical cellular infiltrate suspicious for leukemia or lymphoma. A bone marrow biopsy specimen demonstrated acute myelogenous leukemia (M4 subtype). Following consultation with pediatric oncology and the recognition of the potential for spontaneous regression, chemotherapy for the infant's condition was not recommended. He remained otherwise healthy and was followed-up with biweekly to monthly complete blood counts and physical examinations, which were repeatedly normal. By 3 months of age, the nodules had completely resolved and there was no evidence of recurrence at 8 months of follow-up. We report this instance of aleukemic congenital leukemia with spontaneous regression of leukemia cutis without therapeutic intervention. PMID- 15660894 TI - Dyskeratosis congenita in an adolescent girl with associated choanal atresia. AB - Dyskeratosis congenita is a rare, progressive, degenerative disorder characterized by cutaneous and mucosal involvement in the first decade of life with malignant changes and bone marrow failure in the second and third decades. The primary inheritance pattern is X-linked recessive, with the majority of cases presenting in boys. We report dyskeratosis congenita in an adolescent girl with choanal atresia, a previously unreported association. PMID- 15660895 TI - Scleredema adultorum associated with hyperkeratosis. AB - Scleredema adultorum is a rare skin disorder reported to occur usually following streptococcal infections. It has characteristic clinical and histopathologic findings; however, associated hyperkeratosis is not known. We report a 6-year-old girl with scleredema adultorum and hyperkeratosis. The patient's disease had a benign course. To our knowledge, this patient is the first reported with scleredema adultorum associated with hyperkeratosis. PMID- 15660896 TI - Is disseminated juvenile xanthogranulomatosis benign cephalic histiocytosis? AB - We describe an infant with a congenital form of non-Langerhans cell histiocytosis with clinical and pathologic features of both disseminated juvenile xanthogranulomatosis and benign cephalic histiocytosis. The findings in this case support the concept of these non-Langerhans cell histiocytoses forming part of a spectrum of disease rather than being separate pathologic entities. PMID- 15660897 TI - Phacomatosis pigmentokeratotica: a 20-year follow-up with malignant degeneration of both nevus components. AB - Phacomatosis pigmentokeratotica is a rare syndrome defined by the association of an organoid nevus occasionally with sebaceous differentiation, a speckled lentiginous nevus, and other extracutaneous anomalies. The disorder is a consequence of the so-called twin spot genetic mechanism. We describe the first occurrence involving malignant degeneration of both nevus components, giving rise to three basal cell carcinomas over the sebaceous nevus and a malignant melanoma of the superficial spreading type over the speckled lentiginous nevus. This observation, in concert with the other instances reported in the literature, points to the need for adequate patient follow-up to ensure early detection and treatment of any possible associated malignant degeneration. PMID- 15660898 TI - Epidermal nevus syndrome associated with adnexal tumors, spitz nevus, and hypophosphatemic vitamin D-resistant rickets. AB - The epidermal nevus syndrome is the association of epidermal nevi with abnormalities in other organ systems, most commonly the central nervous system, the skeletal system, and the eyes. We present a patient with epidermal nevus syndrome associated with hypophosphatemic vitamin D-resistant rickets and multiple adnexal and spindle cell tumors. PMID- 15660899 TI - Congenital erosive and vesicular dermatosis healing with reticulated supple scarring. AB - Congenital erosive and vesicular dermatosis healing with reticulated supple scarring is a rare disorder of unknown etiology first reported in three patients in 1985. Nine patients have subsequently been reported, helping to further characterize this unique dermatosis. We describe another patient and further describe the histologic and electron microscopic findings of this entity. This case is unique in that histologic distinction is made between vesicular and scarred lesions. PMID- 15660900 TI - Phrynoderma: a manifestation of vitamin A deficiency?... The rest of the story. AB - Phrynoderma is a distinctive form of follicular hyperkeratosis associated with nutritional deficiency. Although originally thought to represent vitamin A deficiency, several studies have demonstrated multiple etiologies. Characteristic skin lesions are hyperkeratotic papules that first appear on the extensor surfaces of the extremities, shoulders, and buttocks. We report a 14-month-old boy with malnourishment and hyperkeratotic papules and plaques with histologic changes typical of phrynoderma. Despite an extensive evaluation, a specific nutritional deficiency was not identified. Phrynoderma is believed to be a manifestation of severe malnutrition, not necessarily accompanying low vitamin A levels. While the literature supports a link between phrynoderma and vitamins E, B, A, and essential fatty acids general malnutrition seems to be the strongest association. The clinical picture typically improves with enhanced nutritional status. Phrynoderma must be considered in the differential diagnosis in patients with extensor surface hyperkeratotic papules and plaques in the setting of malnourishment and should prompt the clinician to evaluate cell markers of nutritional status, not just vitamin A. We believe this patient exemplifies the conundrum that faces clinicians in evaluating patients with extensor surface predominant hyperkeratotic papules and plaques in the setting of malnourishment. PMID- 15660901 TI - Atopic dermatitis exclusively localized on nipples and areolas. AB - We present an 11-year-old girl, with celiac disease, and a 9- month history of itchy and erythemato-edematous lesions with vesicles and exudation on her nipples and areolas. No other lesions or signs of scratching were present on her face or folds. She had no specific lesions of atopic dermatitis in typical sites nor in other body surface during her life. Patch tests showed a positive reaction to nickel and thimerosal that was not significantly related to the clinical appearance. This presentation documents the clinical relevance of atopic dermatitis minor diagnostic criteria. We discuss the importance of nipple eczema in AD and its differential diagnosis. PMID- 15660902 TI - Treatment of multiple trichoepitheliomas with topical imiquimod and tretinoin. AB - An 11-year-old white girl presented multiple flesh-colored dome-shaped papules on the forehead, nose, scalp, and posterior neck. The lesions had been present for 2 years and were asymptomatic. Family history was negative for skin diseases. A skin biopsy specimen revealed histopathology consistent with trichoepitheliomas. Treatment of multiple trichoepitheliomas is usually difficult. Methods reported in the literature include laser treatment, surgery, cryotherapy, electrodessication, and radiation. The parents were concerned about the risk of scarring and wanted to pursue a nonscarring treatment. The patient was initially treated with topical imiquimod three times a week. She progressively increased the frequency of application to twice daily and added topical tretinoin gel once daily to her regimen for more resistant lesions. After 3 years of treatment, the patient experienced approximately 80% clearing of lesions without scarring. The advantage of using this nonsurgical treatment is no scarring, painless, and no need for other invasive procedures such as injection of local anesthetic. PMID- 15660903 TI - Drug-induced hypersensitivity syndrome in a premature infant. AB - Fever, skin reactions, and limb edema because of drug-induced hypersensitivity have been reported in children because of various drugs, mainly aromatic antiepileptic drugs such as phenytoin, phenobarbital, carbamazepin, and primidone. The skin reactions differ in severity and range from mild maculopapular erythema to exfoliative dermatitis. They have been described in older children but have not been reported in newborn infants. We report a premature newborn infant who developed fever, skin reactions, and edema because of phenytoin while receiving anticonvulsant therapy. PMID- 15660904 TI - What syndrome is this? Wiedemann-Rautenstrauch syndrome. PMID- 15660905 TI - Generalized bullous eruption in an infant. PMID- 15660906 TI - Alopecia areata in infants: is it uncommon? PMID- 15660907 TI - Hypopigmented mycosis fungoides in a child successfully treated with UVA1-light. PMID- 15660908 TI - Geographic tongue in two children with nonpustular psoriasis. PMID- 15660909 TI - Methotrexate for generalized pustular psoriasis in a 2-year-old child. PMID- 15660910 TI - Focal hypertrichosis during topical tacrolimus therapy for childhood vitiligo. PMID- 15660911 TI - Cutaneous mastocytosis in children: an Indian experience. PMID- 15660912 TI - Fox fordyce disease in a prepubertal girl. PMID- 15660914 TI - Directing stem cells into the keratinocyte lineage in vitro. AB - A major area of research in regenerative medicine is the potential application of stem cells in skin grafting and tissue engineering. This would require well defined and efficient protocols for directing the commitment and differentiation of stem cells into the keratinocyte lineage, together with their selective purification and proliferation in vitro. The development of such protocols would reduce the likelihood of spontaneous differentiation of stem cells into divergent lineages upon transplantation, as well as reduce the risk of teratoma formation in the case of embryonic stem cells. Additionally, such protocols could provide useful in vitro models for studying skin tissue biology, as well as facilitate the genetic manipulation of stem cells for therapeutic applications. The development of pharmacokinetic and cytotoxicity/genotoxicity screening tests for skin-related biomaterials and drugs could also utilize protocols developed for the commitment and differentiation of stem cells into the keratinocyte lineage. Hence, this review critically examines the various strategies that could be employed to direct the commitment and differentiation of stem cells into the keratinocyte lineage in vitro. PMID- 15660915 TI - Deficient translocation of c-Rel is associated with impaired Th1 cytokine production in T cells from atopic dermatitis patients. AB - Decreased production of T helper type 1 (Th1) cytokines, such as interferon-gamma (IFN-gamma) or interleukin-2 (IL-2), is a hallmark of atopic diseases. While accessory signals from antigen-presenting cells may be missing, T cells themselves may be suppressed in their ability to produce substantial amounts of Th1 cytokines. We show, in this study, that T cell receptor (TCR)-activated T cells from atopic dermatitis (AD) patients proliferate less than control T cells and produce lower amounts of IFN-gamma and IL-2, but comparable amounts of IL-4. Because mice lacking the nuclear factor kappa B (NF-kappaB) transcription factors - p65 or c-Rel - show reduced Th1, but undisturbed Th2 responses, we investigated the role of c-Rel and p65 for Th1 cytokine production in T cells from healthy and severe AD patients. TCR-activated primary T cells from healthy donors treated with c-Rel antisense oligonucleotides produced lower levels of IL-2 and IFN-gamma and proliferated less efficiently than the corresponding control T cells. Moreover, transfection of primary T cells with c-Rel or p65 enhanced proliferation and production of IL-2 and IFN-gamma. Nuclear extracts of activated primary T cells from AD donors bound weakly to NF-kappaB-specific oligonucleotides, compared to extracts from healthy control T cells. Western blotting studies revealed that nuclear, but not cytosolic, extracts from T cells of AD patients lacked significant amounts of c-Rel and p65. T cell clones derived from AD patients failed to sufficiently translocate c-Rel and p65 into the nucleus following activation. Thus, impaired nuclear translocation of c-Rel and p65 may determine an impaired Th1 cytokine response in AD. PMID- 15660916 TI - Increased mitochondrial respiratory chain enzyme activities correlate with minor extent of liver damage in mice suffering from erythropoietic protoporphyria. AB - Mitochondrial dysfunction might play a role in the pathogenesis of liver damage in erythropoietic protoporphyria (EPP). Changes in mitochondrial respiratory chain activities were evaluated in the Fech(m1pas)/Fech(m1pas) mouse model for EPP. Mice from different strains congenic for the same ferrochelatase germline mutation manifest variable degrees of hepatobiliary injury. Protoporphyric animals bred into the C57BL/6J background showed a higher degree of hepatomegaly and liver damage as well as higher protoporphyrin (PP) accumulation than those bred into the SJL/J and BALB/cJ backgrounds. Whereas mitochondrial respiratory chain activities remained unchanged in the liver of protoporphyric mice C57BL/6J, they were increased in protoporphyric mice from both SJL/J and BALB/cJ backgrounds, when compared to wild-type animals. Mitochondrial respiratory chain activities were increased in Hep G2 cell line after accumulation of PP following addition of aminolevulinic acid. As a direct effect of these elevated mitochondrial activities, in both hepatic cells from mutant mouse strains and Hep G2 cells, adenosine 5'-triphosphate (ATP) levels significantly increased as the intracellular PP concentration was reduced. These results indicate that PP modifies intracellular ATP requirements as well as hepatic mitochondrial respiratory chain enzymatic activities and further suggest that an increase of these activities may provide a certain degree of protection against liver damage in protoporphyric mice. PMID- 15660917 TI - Effects of glycolic acid on desquamation-regulating proteinases in human stratum corneum. AB - In order to investigate the mechanism of glycolic acid (GA) function in human stratum corneum, we monitored changes in cathepsin D-like (CD) and chymotrypsin like (SCCE) proteinases for 3 weeks following topical GA application (50% w/v, pH 0.9) for 30 min to human skin. In the early phase, weakened stratum corneum cohesion in the lower layers was observed on day 2 and the amount of active CD in the upper layer of the stratum corneum was significantly decreased from 30 min until day 2, whereas that in the lower layer remained normal. In contrast, the amount of active SCCE showed no change during the experimental period. The surface pH of the stratum corneum drastically decreased to pH 2 at 30 min and slightly recovered to around pH 3 until 1 day after treatment. From 9 to 19 days, a decrease in corneocyte cell area and a remarkable long-term increase in the amount of active CD in the upper layer were observed. In an in vitro study, the activities of desquamation-regulating proteinases were shown to have remarkably increased at around pH 3, due to activation of CD at its optimal pH. These results suggest that GA functions via at least two different mechanisms, acute activation of CD in the lower layer by acidification around pH 3, along with inactivation of CD in the upper layer, and long-term enhancement of de novo CD production in the few weeks following GA treatment. PMID- 15660918 TI - Human herpesvirus-8 infection of umbilical cord-blood-derived CD34+ stem cells enhances the immunostimulatory function of their dendritic cell progeny. AB - CD34(+) progenitor cells carrying human herpesvirus-8, Kaposi's sarcoma associated herpesvirus (HHV-8/KSHV), have been described in the peripheral blood of AIDS patients suffering from Kaposi's sarcoma (KS). In this study, we investigated the influence of HHV-8 on the differentiation of CD34(+) progenitor cells. Native CD34(+) cells derived from cord blood could be infected by a laboratory strain of HHV-8, as shown by immunofluorescence staining and polymerase chain reaction, but no significant initial maturation/differentiation effects were observed. In addition, these infected cells were differentiated into immature and mature dendritic cells (DCs) using cytokine induction with recombinant human granulocyte-macrophage colony-stimulating factor (rhGm-CSF), recombinant human tumor necrosis factor (rhTNF-alpha) and recombinant human stem cell factor (rhSCF). Double immunofluorescence and flow cytometry studies demonstrated that virus infection did not impair the development of immature and mature DC populations. Subsequently, the immunostimulating capacity of DC populations was tested in a mixed lymphocyte reaction using allogeneic T-cells. The HHV-8-infected CD34(+) progenitor cell-derived mature DC population showed a significantly enhanced antigen-presenting capacity, compared to non-infected DCs, which was not observed with the immature DCs. This suggests stimulation of DC function by HHV-8 infection. Because there are only a small percentage of HHV-8 positive DCs in the preparations and because it is not clear whether infection is abortive or productive to some extent, this seems to be most likely due to an indirect viral effect. PMID- 15660920 TI - Possible histopathologic correlates of dermoscopic features in pigmented melanocytic lesions identified by means of optical coherence tomography. AB - Optical coherence tomography (OCT) is a novel non-invasive imaging technique for in vivo histologic characterization of tissues. Besides pure morphology, additional functional parameters of the tissue investigated can be evaluated at the same time, such as the refractive index and the scattering coefficient. The purpose of our study is to correlate in vivo the dermoscopic structures that first appear in the melanocytic pigmented lesion (pigment network and brown globules) using this new method, with the histopathologic correlates, in order to improve their characterization and achieve easier recognition and inter-observer reproducibility. We concentrated in particular on the pigment network and the brown globules, as these are dermoscopic parameters of great diagnostic importance in melanocytic lesions. Moreover, as these parameters are the histopathologic equivalents of structures located at the level of the dermo epidermal junction, they enable a correct evaluation to be made using OCT, that at present has only a few millimetres penetration power. The results of our trial, performed using the histopathological preparation as an evaluation gold standard, show that in selected cases OCT allows an in vivo correlation to be made between surface dermoscopic parameters and histopathologic correlates, in particular the pigment network and brown globules. The resolution is not high enough to reveal the morphology of the single cells, but it is possible to evaluate the architecture of a lesion. PMID- 15660919 TI - Overrepresentation of the founder PPOX gene mutation R59W in a South African patient with severe clinical manifestation of porphyria. AB - A patient, who presented with abdominal pain and severe photosensitivity that resulted in scarring and mutilation of the fingers, nose and ears, was referred for biochemical assessment of porphyria and DNA screening. Although these clinical manifestations were suggestive of both acute porphyria and congenital erythropoietic porphyria, the biochemical profile was consistent with variegate porphyria (VP). Analysis of the protoporphyrinogen oxidase (PPOX) gene underlying VP resulted in the identification of the founder mutation R59W in a heterozygous state in this patient. Despite extensive mutation analysis, no other potential disease-causing genetic alterations could be detected in the PPOX gene or the uroporphyrinogen III synthase gene. Slight overrepresentation of the mutant PPOX allele was however, observed repeatedly in DNA of the proband compared to other R59W heterozygotes, including his mother who also tested positive for mutation R59W using restriction enzyme analysis and direct DNA sequencing. Confirmation of this phenomenon by real-time polymerase chain reaction analysis and microsatellite analysis, using highly informative markers flanking the PPOX gene, raised the possibility of partial homozygosity for VP in this patient. This study represents the first report of overrepresentation of mutation R59W in a patient with a severe form of VP. A homozygote for the R59W mutation has never been detected, and the severe clinical manifestation observed in our patient is consistent with the hypothesis that such a genotype will not be compatible with life. PMID- 15660921 TI - Optimization of a transplant model to assess skin reconstitution from stem cell enriched primary human keratinocyte populations. AB - Given that an important functional attribute of stem cells in vivo is their ability to sustain tissue regeneration, we set out to establish a simple and easy technique to assess this property from candidate populations of human keratinocyte stem cells in an in vivo setting. Keratinocytes were inoculated into devitalized rat tracheas and transplanted subcutaneously into SCID mice, and the epithelial lining regenerated characterized to establish the validity of this heterotypic model. Furthermore, the rate and quality of epidermal tissue reconstitution obtained from freshly isolated unfractionated vs. keratinocyte stem cell-enriched populations was tested as a function of (a) cell numbers inoculated; and (b) the inclusion of irradiated support keratinocytes and dermal cells. Rapid and sustained epidermal tissue regeneration from small numbers of freshly isolated human keratinocyte stem cells validates the utilization of this simple and reliable model system to assay for enrichment of epidermal tissue reconstituting cells. PMID- 15660923 TI - Leukocyte extravasation as a target for anti-inflammatory therapy - Which molecule to choose? AB - In view of the central pathogenic importance of leukocyte extravasation in inflammatory skin diseases, therapeutic interference with this - surprisingly complex - process is clearly a promising new approach for treating these dermatoses. Despite some disappointments during the clinical use of these agents and despite their crippling price tag, the recent incorporation of biologicals that target defined molecular controls of leukocyte extravasation into dermatological and rheumatological practise, consequently, has greatly enriched our therapeutic options for battling major, chronic, inflammatory dermatoses such as psoriasis. However, the - as yet unresolved and still rather controversially discussed - critical question is: Which of the multiple steps that control leukocyte extravasation in the human system really offer the most promising, most pragmatic, and safest molecular targets for therapeutic intervention for which disease entity? The current debate intends to stimulate public and rational debate of this crucial issue, beyond the evident commercial interests that are touched by whatever stand one takes. PMID- 15660930 TI - Integrating QTL mapping and genome scans towards the characterization of candidate loci under parallel selection in the lake whitefish (Coregonus clupeaformis). AB - As natural selection must act on underlying genetic variation, discovering the number and location of loci under the influence of selection is imperative towards understanding adaptive divergence in evolving populations. Studies employing genome scans have hypothesized that the action of divergent selection should reduce gene flow at the genomic locations implicated in adaptation and speciation among natural populations, yet once 'outlier' patterns of variation have been identified the function and role of such loci needs to be confirmed. We integrated adaptive QTL mapping and genomic scans among diverging sympatric pairs of the lake whitefish (Coregonus clupeaformis) species complex in order to test the hypothesis that differentiation between dwarf and normal ecotypes at growth associated QTL was maintained by directional selection. We found evidence of significantly high levels of molecular divergence among eight growth QTL where two of the strongest candidate loci under the influence of directional selection exhibited parallel reductions of gene flow over multiple populations. PMID- 15660931 TI - Quantitative trait locus mapping in natural populations: progress, caveats and future directions. AB - Over the last 15 years quantitative trait locus (QTL) mapping has become a popular method for understanding the genetic basis of continuous variation in a variety of systems. For example, the technique is now an integral tool in medical genetics, livestock production, plant breeding and population genetics of model organisms. Ten years ago, it was suggested that the method could be used to understand continuous variation in natural populations. In this review I: (i) clarify what is meant by natural population in the QTL context, (ii) discuss whether evolutionary biologists have successfully mapped QTL in natural populations, (iii) highlight some of the questions that have been addressed by QTL mapping in natural populations, (iv) describe how QTL mapping can be conducted in unmanipulated natural populations, (v) highlight some of the limitations of QTL mapping and (vi) try to predict some future directions for QTL mapping in natural populations. PMID- 15660932 TI - Local adaptation and species segregation in two mussel (Mytilus edulis x Mytilus trossulus) hybrid zones. AB - Few marine hybrid zones have been studied extensively, the major exception being the hybrid zone between the mussels Mytilus edulis and Mytilus galloprovincialis in southwestern Europe. Here, we focus on two less studied hybrid zones that also involve Mytilus spp.; Mytilus edulis and Mytilus trossulus are sympatric and hybridize on both western and eastern coasts of the Atlantic Ocean. We review the dynamics of hybridization in these two hybrid zones and evaluate the role of local adaptation for maintaining species boundaries. In Scandinavia, hybridization and gene introgression is so extensive that no individuals with pure M. trossulus genotypes have been found. However, M. trossulus alleles are maintained at high frequencies in the extremely low salinity Baltic Sea for some allozyme genes. A synthesis of reciprocal transplantation experiments between different salinity regimes shows that unlinked Gpi and Pgm alleles change frequency following transplantation, such that post-transplantation allelic composition resembles native populations found in the same salinity. These experiments provide strong evidence for salinity adaptation at Gpi and Pgm (or genes linked to them). In the Canadian Maritimes, pure M. edulis and M. trossulus individuals are abundant, and limited data suggest that M. edulis predominates in low salinity and sheltered conditions, whereas M. trossulus are more abundant on the wave-exposed open coasts. We suggest that these conflicting patterns of species segregation are, in part, caused by local adaptation of Scandinavian M. trossulus to the extremely low salinity Baltic Sea environment. PMID- 15660933 TI - Phylogeography and population genetics of the endangered Amazonian manatee, Trichechus inunguis Natterer, 1883 (Mammalia, Sirenia). AB - We used mitochondrial DNA control region sequences to examine phylogeography and population differentiation of the endangered Amazonian manatee Trichechus inunguis. We observe lack of molecular differentiation among localities and we find weak association between geographical and genetic distances. However, nested clade analysis supports restricted gene flow and/or dispersal with some long distance dispersal. Although this species has a history of extensive hunting, genetic diversity and effective population sizes are relatively high when compared to the West Indian manatee Trichechus manatus. Patterns of mtDNA haplotype diversity in T. inunguis suggest a genetic disequilibrium most likely explained by demographic expansion resulting from secession of hunting and enforcement of conservation and protective measures. Phylogenetic analysis of T. manatus and T. inunguis haplotypes suggests that T. inunguis is nested within T. manatus, effectively making T. manatus a paraphyletic entity. Paraphyly of T. manatus and recent divergence times of T. inunguis and the three main T. manatus lineages suggest a possible need for a taxonomic re-evaluation of the western Atlantic Trichechus. PMID- 15660934 TI - Similarity coefficients for molecular markers in studies of genetic relationships between individuals for haploid, diploid, and polyploid species. AB - Determining true genetic dissimilarity between individuals is an important and decisive point for clustering and analysing diversity within and among populations, because different dissimilarity indices may yield conflicting outcomes. We show that there are no acceptable universal approaches to assessing the dissimilarity between individuals with molecular markers. Different measures are relevant to dominant and codominant DNA markers depending on the ploidy of organisms. The Dice coefficient is the suitable measure for haploids with codominant markers and it can be applied directly to (0,1)-vectors representing banding profiles of individuals. None of the common measures, Dice, Jaccard, simple mismatch coefficient (or the squared Euclidean distance), is appropriate for diploids with codominant markers. By transforming multiallelic banding patterns at each locus into the corresponding homozygous or heterozygous states, a new measure of dissimilarity within locus was developed and expanded to assess dissimilarity between multilocus states of two individuals by averaging across all codominant loci tested. There is no rigorous well-founded solution in the case of dominant markers. The simple mismatch coefficient is the most suitable measure of dissimilarity between banding patterns of closely related haploid forms. For distantly related haploid individuals, the Jaccard dissimilarity is recommended. In general, no suitable method for measuring genetic dissimilarity between diploids with dominant markers can be proposed. Banding patterns of diploids with dominant markers and polyploids with codominant markers represent individuals' phenotypes rather than genotypes. All dissimilarity measures proposed and developed herein are metrics. PMID- 15660935 TI - Genetic diversity and differentiation of central European freshwater pearl mussel (Margaritifera margaritifera L.) populations: implications for conservation and management. AB - Despite the fact that mollusc species play an important role in many aquatic ecosystems, little is known about their biodiversity and conservation genetics. Freshwater pearl mussel (Margaritifera margaritifera L.) populations are seriously declining all over Europe and a variety of conservation programs are being established to support the remaining endangered central European populations. In order to provide guidelines for conservation strategies and management programs, we investigated the genetic structure of 24 freshwater pearl mussel populations originating from five major central European drainages including Elbe, Danube, Rhine, Maas and Weser, representing the last and most important populations in this area. We present a nondestructive sampling method of haemolymph for DNA analyses, which is applicable for endangered bivalves. The analyses of nine microsatellite loci with different levels of polymorphism revealed a high degree of fragmented population structure and very different levels of genetic diversity within populations. These patterns can be explained by historical and demographic effects and have been enforced by anthropogenic activities. Even within drainages, distinct conservation units were detected, as revealed from high F(ST) values, private alleles and genetic distance measures. Populations sampled close to contact zones between main drainage systems showed lowest levels of correct assignment to present-day drainage systems. Populations with high priority for conservation should not only be selected by means of census population size and geographical distance to other populations. Instead, detailed genetic analyses are mandatory for revealing differentiation and diversity parameters, which should be combined with ecological criteria for sustainable conservation and recovery programs. PMID- 15660936 TI - Patterns of genetic diversity and migration in increasingly fragmented and declining orang-utan (Pongo pygmaeus) populations from Sabah, Malaysia. AB - We investigated the genetic structure within and among Bornean orang-utans (Pongo pygmaeus) in forest fragments of the Lower Kinabatangan flood plain in Sabah, Malaysia. DNA was extracted from hair and faecal samples for 200 wild individuals collected during boat surveys on the Kinabatangan River. Fourteen microsatellite loci were used to characterize patterns of genetic diversity. We found that genetic diversity was high in the set of samples (mean H(E) = 0.74) and that genetic differentiation was significant between the samples (average F(ST) = 0.04, P < 0.001) with F(ST) values ranging from low (0.01) to moderately large (0.12) values. Pairwise F(ST) values were significantly higher across the Kinabatangan River than between samples from the same river side, thereby confirming the role of the river as a natural barrier to gene flow. The correlation between genetic and geographical distance was tested by means of a series of Mantel tests based on different measures of geographical distance. We used a Bayesian method to estimate immigration rates. The results indicate that migration is unlikely across the river but cannot be completely ruled out because of the limited F(ST) values. Assignment tests confirm the overall picture that gene flow is limited across the river. We found that migration between samples from the same side of the river had a high probability indicating that orang utans used to move relatively freely between neighbouring areas. This strongly suggests that there is a need to maintain migration between isolated forest fragments. This could be done by restoring forest corridors alongside the river banks and between patches. PMID- 15660937 TI - Microsatellites reveal regional population differentiation and isolation in Lobaria pulmonaria, an epiphytic lichen. AB - Many lichen species produce both sexual and asexual propagules, but, aside from being minute, these diaspores lack special adaptations for long-distance dispersal. So far, molecular studies have not directly addressed isolation and genetic differentiation of lichen populations, both being affected by gene flow, at a regional scale. We used six mycobiont-specific microsatellite loci to investigate the population genetic structure of the epiphytic lichen Lobaria pulmonaria in two regions that strongly differed with respect to anthropogenic impact. In British Columbia, L. pulmonaria grows in continuous old-growth forests, while its populations in the old cultural landscape of Switzerland are comparably small and fragmented. Populations from both British Columbia and Switzerland were genetically diverse at the loci. Geographically restricted alleles, low historical gene flow, and analyses of genetic distance (upgma tree) and of differentiation (amova) indicated that populations from Vancouver Island and from the Canadian mainland were separated from each other, except for one, geographically intermediate population. This differentiation was attributed to different glacial and postglacial histories of coastal and inland populations in British Columbia. In contrast to expectations, the three investigated Swiss populations were genetically neither isolated nor differentiated from each other despite the long-lasting negative human impact on the lichen's range size in Central Europe. We propose that detailed studies integrating local landscape and regional scales are now needed to understand the processes of dispersal and gene flow in lichens. PMID- 15660938 TI - Geographical distance and physical barriers shape the genetic structure of Eurasian red squirrels (Sciurus vulgaris) in the Italian Alps. AB - Red squirrels (Sciurus vulgaris) are widely distributed throughout Eurasia, occurring in many types of coniferous and mixed-deciduous forests. In fragmented landscapes, small and partly isolated populations with low immigration rates show reduced genetic diversity, but reforestation can increase gene flow and restore levels of genetic variation in a few decades. No studies have so far investigated the genetic structure of red squirrel in large, continuous forests. The Italian Alps are presently characterized by almost continuous, recently reconnected forest habitats, that were affected by deep landscape changes during last glaciations but remained mostly unchanged between 10 000 and 200 years bp, when forest cover was heavily reduced. In this study we analyse patterns of genetic variability of red squirrels in and between seven sites distributed over 250 km of Alpine habitat, using mitochondrial DNA (mtDNA) and microsatellites. We use isolation-by-distance (IBD) models to investigate the relative importance that past (Pleistocene glaciations) and recent (fragmentation, bottlenecks) events had on the present genetic situation. Both nuclear and mtDNA data indicate a significant differentiation among study sites and a significant correlation between genetic and geographical distance only over a large scale. No recent bottlenecks are recorded through microsatellites and demographic models strongly support equilibrium between gene flow and drift; however, mtDNA suggests that there may have been local demographic crashes, probably in correspondence with the 19th-century forest fragmentation. These findings indicate that local landscape factors other than geographical distance per se, such as barriers of unsuitable habitat, affect gene flow and determine differentiation. PMID- 15660939 TI - Population structure of Columbia spotted frogs (Rana luteiventris) is strongly affected by the landscape. AB - Landscape features such as mountains, rivers, and ecological gradients may strongly affect patterns of dispersal and gene flow among populations and thereby shape population dynamics and evolutionary trajectories. The landscape may have a particularly strong effect on patterns of dispersal and gene flow in amphibians because amphibians are thought to have poor dispersal abilities. We examined genetic variation at six microsatellite loci in Columbia spotted frogs (Rana luteiventris) from 28 breeding ponds in western Montana and Idaho, USA, in order to investigate the effects of landscape structure on patterns of gene flow. We were particularly interested in addressing three questions: (i) do ridges act as barriers to gene flow? (ii) is gene flow restricted between low and high elevation ponds? (iii) does a pond equal a 'randomly mating population' (a deme)? We found that mountain ridges and elevational differences were associated with increased genetic differentiation among sites, suggesting that gene flow is restricted by ridges and elevation in this species. We also found that populations of Columbia spotted frogs generally include more than a single pond except for very isolated ponds. There was also evidence for surprisingly high levels of gene flow among low elevation sites separated by large distances. Moreover, genetic variation within populations was strongly negatively correlated with elevation, suggesting effective population sizes are much smaller at high elevation than at low elevation. Our results show that landscape features have a profound effect on patterns of genetic variation in Columbia spotted frogs. PMID- 15660940 TI - Do farmers reduce genetic diversity when they domesticate tropical trees? A case study from Amazonia. AB - Agroforestry ecosystems may be an important resource for conservation and sustainable use of tropical trees, but little is known of the genetic diversity they contain. Inga edulis, a widespread indigenous fruit tree in South America, is used as a model to assess the maintenance of genetic diversity in five planted vs. five natural stands in the Peruvian Amazon. Analysis of five SSR (simple sequence repeat) loci indicated lower allelic variation in planted stands [mean corrected allelic richness 31.3 (planted) and 39.3 (natural), P = 0.009]. Concerns regarding genetic erosion in planted Amazonian tree stands appear valid, although allelic variation on-farm is still relatively high. PMID- 15660941 TI - SNPs in ecological and conservation studies: a test in the Scandinavian wolf population. AB - Single nucleotide polymorphisms (SNPs) have the potential to become the genetic marker of choice in studies of the ecology and conservation of natural populations because of their capacity to access variability across the genome. In this study, we provide one of the first demonstrations of SNP discovery in a wild population in order to address typical issues of importance in ecology and conservation in the recolonized Scandinavian and neighbouring Finnish wolf Canis lupus populations. Using end sequence from BAC (bacterial artificial chromosome) clones specific for dogs, we designed assays for 24 SNP loci, 20 sites of which had previously been shown to be polymorphic in domestic dogs and four sites were newly identified as polymorphic in wolves. Of the 24 assayed loci, 22 SNPs were found to be variable within the Scandinavian population and, importantly, these were able to distinguish individual wolves from one another (unbiased probability of identity of 4.33 x 10(-8)), providing equivalent results to that derived from 12 variable microsatellites genotyped in the same population. An assignment test shows differentiation between the Scandinavian and neighbouring Finnish wolf populations, although not all known immigrants are accurately identified. An exploration of the misclassification rates in the identification of relationships shows that neither 22 SNP nor 20 microsatellite loci are able to discriminate across single order relationships. Despite the remaining obstacle of SNP discovery in nonmodel organisms, the use of SNPs in ecological and conservation studies is encouraged by the advent of large scale screening methods. Furthermore, the ability to amplify extremely small fragments makes SNPs of particular use for population monitoring, where faecal and other noninvasive samples are routinely used. PMID- 15660942 TI - Chloroplast DNA variation of Quercus rubra L. in North America and comparison with other Fagaceae. AB - Quercus rubra is one of the most important timber and ornamental tree species from eastern North America. It is a widespread species growing under variable ecological conditions. Chloroplast DNA variation was studied by PCR-RFLP (polymerase chain reaction-restriction fragment length polymorphism) in 290 individuals from 66 populations sampled throughout the natural range. A total of 12 haplotypes were detected, with one found in 75% of the trees. Population differentiation is relatively low (G(ST) = 0.46), even when similarities between haplotypes are taken into account (N(ST) = 0.50), pointing to a weak phylogeographical structure. Furthermore, no spatial structure of genetic diversity could be detected. The genetic differentiation increased northwards, reflecting the postglacial history of Q. rubra. The unusual aspect of this study was the low level of chloroplast DNA genetic differentiation in Q. rubra compared to that typically observed in other oak species. Palynological evidence indicates that during the last glacial maximum, Q. rubra had one major distribution range with populations located relatively far to the north, resulting in only modest movement northwards when climate improved, whereas European white oaks were largely restricted to the southern European peninsulas and experienced extensive movements during the postglacial period. The contrasted geographical features and levels of tree species richness of both continents might further explain why congeneric species sharing similar life history traits have genetic structures that are so different. PMID- 15660943 TI - Geographic and phylogenetic evidence for dispersed nuclear introgression in a daphniid with sexual propagules. AB - The role of among-species gene flow in eukaryotic evolution remains controversial. Putative hybrid lineages are common in water fleas, but their ecological success is often associated with polyploidy and the production of asexual propagules. Advanced hybrid lineages with sexual propagules are expected to be geographically restricted because their successful dispersal is contingent on overcoming fertility complications, assimilation by parent taxa, and competition with parent taxa. Here we provide evidence that a diploid lineage of Daphnia has been formed by introgression between distantly related species and attained a broad distribution (Nearctic) despite its requirement for sexual propagules. The evidence is based on geographical discordance, phylogenetic discordance, recombinant genotypes and additive genotypes of the nuclear internal transcribed spacer regions (ITS) and mitochondrial DNA. Additive genotypes also provided evidence of hybridization between introduced European Daphnia and North American Daphnia. We argue that the unique biology of Holarctic lacustrine water fleas and the spatial separation of lineages during Pleistocene glaciation have promoted hybridization and its evolutionary consequences. PMID- 15660944 TI - Genetic variation and phylogeographic analyses of two species of Carpobrotus and their hybrids in California. AB - Despite the commonality and study of hybridization in plants, there are few studies between invasive and noninvasive species that examine the genetic variability and gene flow of cytoplasmic DNA. We describe the phylogeographical structure of chloroplast DNA (cpDNA) variation within and among several interspecific populations of the putative native, Carpobrotus chilensis and the introduced, Carpobrotus edulis (Aizoaceae). These species co-occur throughout much of coastal California and form several 'geographical hybrid populations'. Two hundred and thirty-seven individuals were analysed for variation in an approximate 7.0 kb region of the chloroplast genome using PCR-RFLP (polymerase chain reaction - restriction fragment length polymorphism) data. Phylogenetic analyses and cpDNA population differentiation were conducted for all morphotypes. Historic geographical dispersion and the coefficient of ancestry of the haplotypes were determined using nested clade analyses. Two haplotypic groupings (I and II) were represented in C. chilensis and C. edulis, respectively. The variation in cpDNA data is in agreement with the previously reported allozyme and morphological data; this supports relatively limited variation and high population differentiation among C. chilensis and hybrids and more wide-ranging variation in C. edulis and C. edulis populations backcrossed with C. chilensis. C. chilensis disproportionately contributes to the creation of hybrids with the direction of gene flow from C. chilensis into C. edulis. The cpDNA data support C. chilensis as the maternal contributor to the hybrid populations. PMID- 15660945 TI - Evidence for shared ancestral polymorphism rather than recurrent gene flow at microsatellite loci differentiating two hybridizing oaks (Quercus spp.). AB - Quercus petraea and Quercus robur are two closely related oak species, considered to hybridize. Genetic markers, however, indicate that despite sharing most alleles, the two species remain separate genetic units. Analysis of 20 microsatellite loci in multiple populations from both species suggested a genome wide differentiation. Thus, the allele sharing between both species could be explained either by low rates of gene flow or shared ancestral variation. We performed further analyses of population differentiation in a biogeographical setting and an admixture analysis in mixed oak stands to distinguish between both hypotheses. Based on our results we propose that the low genetic differentiation among these species results from shared ancestry rather than high rates of gene flow. PMID- 15660946 TI - Population structure in African Drosophila melanogaster revealed by microsatellite analysis. AB - Tropical sub-Saharan regions are considered to be the geographical origin of Drosophila melanogaster. Starting from there, the species colonized the rest of the world after the last glaciation about 10 000 years ago. Consistent with this demographic scenario, African populations have been shown to harbour higher levels of microsatellite and sequence variation than cosmopolitan populations. Nevertheless, limited information is available on the genetic structure of African populations. We used X chromosomal microsatellite variation to study the population structure of D. melanogaster populations using 13 sampling sites in North, West and East Africa. These populations were compared to six European and one North American population. Significant population structure was found among African D. melanogaster populations. Using a Bayesian method for inferring population structure we detected two distinct groups of populations among African D. melanogaster. Interestingly, the comparison to cosmopolitan D. melanogaster populations indicated that one of the divergent African groups is closely related to cosmopolitan flies. Low, but significant levels of differentiation were observed for sub-Saharan D. melanogaster populations from West and East Africa. PMID- 15660947 TI - Phylogeography of the Italian wall lizard, Podarcis sicula, as revealed by mitochondrial DNA sequences. AB - In a phylogeographical survey of the Italian wall lizard, Podarcis sicula, DNA sequence variation along an 887-bp segment of the cytochrome b gene was examined in 96 specimens from 86 localities covering the distribution range of the species. In addition, parts of the 12S rRNA and 16S rRNA genes from 12 selected specimens as representatives of more divergent cytochrome b haploclades were sequenced (together about 950 bp). Six phylogeographical main groups were found, three representing samples of the nominate subspecies Podarcis sicula sicula and closely related subspecies and the other three comprising Podarcis sicula campestris as well as all subspecies described from northern and eastern Adriatic islands. In southern Italy a population group with morphological characters of P. s. sicula but with the mitochondrial DNA features of P. s. campestris was detected indicating a probably recent hybridization zone. The present distribution patterns were interpreted as the consequence of natural events like retreats to glacial refuges and postglacial area expansions, but also as the results of multiple introductions by man. PMID- 15660948 TI - Multilevel genetic analyses of two European supercolonies of the Argentine ant, Linepithema humile. AB - Some ants have an extraordinary unicolonial social organization, whereby individuals mix freely among physically separated nests. Recently, it was shown that the European population of Linepithema humile consisted of two enormous unicolonial supercolonies. Workers of the same supercolony are never aggressive to each other. In contrast, aggressiveness is invariably high between workers from different supercolonies. Here we investigated whether gene flow occurs between two supercolonies. We identified a contact zone in which we sampled 46 nests. For each nest, aggression tests were conducted against workers from reference nests from both supercolonies. Workers were always very aggressive towards workers of one of the supercolonies but not to workers of the other. Thus, all nests could be clearly assigned to one of the two supercolonies. For 22 of the 46 nests, we genotyped 15-16 workers at five microsatellite loci. A four level hierarchical analysis of variance revealed very strong genetic differentiation between the two supercolonies (F(SUPERCOLONY-TOTAL) = 0.541) and low differentiation between sectors (i.e. group of nests connected together with trails) within supercolonies (F(SECTOR-SUPERCOLONY) = 0.064). The very high differentiation between the two supercolonies indicates a lack of ongoing gene flow, a conclusion further bolstered by the finding that the two supercolonies share no common alleles at two of the five microsatellite loci. A Bayesian clustering method also revealed the occurrence of two distinct clusters. These clusters exactly match the grouping obtained by aggression tests. None of the 332 genotyped individuals were admixed despite the fact that some nests of the two supercolonies were separated by less than 30 m. These results demonstrate that the two supercolonies have completely separate gene pools. PMID- 15660949 TI - Microsatellite genotyping errors: detection approaches, common sources and consequences for paternal exclusion. AB - Microsatellite genotyping errors will be present in all but the smallest data sets and have the potential to undermine the conclusions of most downstream analyses. Despite this, little rigorous effort has been made to quantify the size of the problem and to identify the commonest sources of error. Here, we use a large data set comprising almost 2000 Antarctic fur seals Arctocephalus gazella genotyped at nine hypervariable microsatellite loci to explore error detection methods, common sources of error and the consequences of errors on paternal exclusion. We found good concordance among a range of contrasting approaches to error-rate estimation, our range being 0.0013 to 0.0074 per single locus PCR (polymerase chain reaction). The best approach probably involves blind repeat genotyping, but this is also the most labour-intensive. We show that several other approaches are also effective at detecting errors, although the most convenient alternative, namely mother-offspring comparisons, yielded the lowest estimate of the error rate. In total, we found 75 errors, emphasizing their ubiquitous presence. The most common errors involved the misinterpretation of allele banding patterns (n = 60, 80%) and of these, over a third (n = 22, 36.7%) were due to confusion between homozygote and adjacent allele heterozygote genotypes. A specific test for whether a data set contains the expected number of adjacent allele heterozygotes could provide a useful tool with which workers can assess the likely size of the problem. Error rates are also positively correlated with both locus polymorphism and product size, again indicating aspects where extra effort at error reduction should be directed. Finally, we conducted simulations to explore the potential impact of genotyping errors on paternity exclusion. Error rates as low as 0.01 per allele resulted in a rate of false paternity exclusion exceeding 20%. Errors also led to reduced estimates of male reproductive skew and increases in the numbers of pups that matched more than one candidate male. Because even modest error rates can be strongly influential, we recommend that error rates should be routinely published and that researchers make an attempt to calculate how robust their analyses are to errors. PMID- 15660950 TI - High specificity generally characterizes mycorrhizal association in rare lady's slipper orchids, genus Cypripedium. AB - Lady's slipper orchids (Cypripedium spp.) are rare terrestrial plants that grow throughout the temperate Northern Hemisphere. Like all orchids, they require mycorrhizal fungi for germination and seedling nutrition. The nutritional relationships of adult Cypripedium mycorrhizae are unclear; however, Cypripedium distribution may be limited by mycorrhizal specificity, whether this specificity occurs only during the seedling stage or carries on into adulthood. We attempted to identify the primary mycorrhizal symbionts for 100 Cypripedium plants, and successfully did so with two Cypripedium calceolus, 10 Cypripedium californicum, six Cypripedium candidum, 16 Cypripedium fasciculatum, two Cypripedium guttatum, 12 Cypripedium montanum, and 11 Cypripedium parviflorum plants from a total of 44 populations in Europe and North America, yielding fungal nuclear large subunit and mitochondrial large subunit sequence and RFLP (restriction fragment length polymorphism) data for 59 plants. Because orchid mycorrhizal fungi are typically observed without fruiting structures, we assessed fungal identity through direct PCR (polymerase chain reaction) amplification of fungal genes from mycorrhizally colonized root tissue. Phylogenetic analysis revealed that the great majority of Cypripedium mycorrhizal fungi are members of narrow clades within the fungal family Tulasnellaceae. Rarely occurring root endophytes include members of the Sebacinaceae, Ceratobasidiaceae, and the ascomycetous genus, Phialophora. C. californicum was the only orchid species with apparently low specificity, as it associated with tulasnelloid, ceratobasidioid, and sebacinoid fungi in roughly equal proportion. Our results add support to the growing literature showing that high specificity is not limited to nonphotosynthetic plants, but also occurs in photosynthetic ones. PMID- 15660951 TI - Super-infections of Wolbachia in byturid beetles and evidence for genetic transfer between A and B super-groups of Wolbachia. AB - Wolbachia are maternally inherited bacteria responsible for altering host reproduction. The two main groups found in insects, A and B, are based on molecular characterization using ribosomal, ftsZ, wsp (Wolbachia surface protein) or groE genes. We have used the wsp and ftsZ genes to study Wolbachia in byturid beetles. Byturus affinis contained a single copy of the ftsZ gene which grouped with A ftsZ sequences and a single copy of the wsp gene which grouped with B wsp sequences. This suggests that genetic exchange between A and B groups has occurred in the Wolbachia of this beetle. FtsZ and wsp sequences that were identical or nearly identical to those of B. affinis were found in B. tomentosus, suggesting that it also contains the same recombinant Wolbachia genotype. Most other byturids had more than one wsp sequence with at least one from the A and B groups, suggesting multiple copies of bacterial genes or multiple infections. B. ochraceus and B. unicolor both had four distinct wsp gene sequences. All the byturids had a closely related A wsp sequence and most a closely related B wsp sequence. Therefore, there appears to be an association between specific A and B wsp types. PMID- 15660952 TI - Brood size modifications affect plumage bacterial assemblages of European starlings. AB - During reproduction, birds face trade-offs between time and energy devoted to parental effort and traits associated with self-maintenance. We manipulated brood sizes to investigate the effects of such trade-offs on feather bacterial densities and the structure of bacterial assemblages on feathers in adult European starlings, Sturnus vulgaris, and in vitro feather degradation. As predicted by a trade-off between parental effort and self-maintenance, we found that birds with enlarged broods had more free-living bacteria on their feathers than birds with reduced broods. Furthermore, we found a significant interaction between brood manipulation and original brood size on free-living bacterial densities suggesting that the trade-off is mediated by the adults' initial reproductive investment. In contrast, brood size manipulations had no significant effect on densities of attached bacteria. Using ribosomal intergenic spacer analysis (RISA), we demonstrated that brood manipulations significantly modified the structure (band pattern) of feather-degrading bacterial assemblages, but had no significant effect on their richness (number of bands) or the in vitro feather degradation. In vitro feather degradation varied in relation to the premanipulation brood size and positively with the richness of the feather degrading bacterial community. Besides brood manipulation effect, we found that ecological factors and individual traits, such as the age, the nest location or the capture date, shaped bacterial assemblages and feather degradation capacities. PMID- 15660953 TI - Intriguing asexual life in marginal populations of the brown seaweed Fucus vesiculosus. AB - Reproduction of attached large brown algae is known to occur only by sexual zygotes. Using microsatellites we show evolution of asexual reproduction in the bladder wrack promoting population persistence in the brackish water Baltic Sea (< 6 psu). Here a dwarf morph of Fucus vesiculosus is dominated by a single clone but clonal reproduction is also present in the common form of the species. We describe a possible mechanism for vegetative reproduction of attached algae, and conclude that clonality plays an important role in persistence and dispersal of these marginal populations, in which sexual reproduction is impaired by low salinity. PMID- 15660954 TI - Male nest site fidelity and female serial polyandry in lingcod (Ophiodon elongatus, Hexagrammidae). AB - Nest site fidelity and serial polyandry were examined in lingcod, Ophiodon elongatus, a teleost fish in which the nest-guarding male parent invests more heavily in parental care than the elusive female parent. Lingcod parental and progeny genotypes were established for fish spawning on a 200 m(2) section of Snake Island reef, British Columbia in two successive years to evaluate male and female mate choice (monogamy or polygamy) and nest site reuse by the same parents (nest site fidelity) and/or different parents (nest site affinity). Thirteen nests (egg masses) guarded by nine males and 14 nests guarded by seven males were observed in 2002 and 2003, respectively. No female laid more than one nest per season or spawned in the study area in both years. In contrast, at least six (86%) and possibly all seven (100%) of the 2003 guardian males had been guardian or auxiliary males in 2002. Both nest site affinity and extreme male nest site fidelity were observed, with at least four males reusing the exact same nest site. Serial polyandry resulting from the high male and low female nest site fidelity is consistent with predictions based on a low female parental investment and high rate of progeny loss to predation and cannibalism. Male polygyny, achieved primarily by cuckoldry within seasons, was enhanced by the lack of female fidelity between seasons. Polygamy in both sexes of nest-tending marine fish may minimize reproductive skew and maximize genetic diversity within populations. PMID- 15660955 TI - Nonshivering thermogenesis capacity associated to mitochondrial DNA haplotypes and gender in the greater white-toothed shrew, Crocidura russula. AB - A selection gradient was recently suggested as one possible cause for a clinal distribution of mitochondrial DNA (mtDNA) haplotypes along an altitudinal transect in the greater white-toothed shrew, Crocidura russula (Ehinger et al. 2002). One mtDNA haplotype (H1) rare in lowland, became widespread when approaching the altitudinal margin of the distribution. As H1 differs from the main lowland haplotype by several nonsynonymous mutations (including on ATP6), and as mitochondria play a crucial role in metabolism and thermogenesis, distribution patterns might stem from differences in the thermogenic capacity of different mtDNA haplotypes. In order to test this hypothesis, we measured the nonshivering thermogenesis (NST) associated with different mtDNA haplotypes. Sixty-two shrews, half of which had the H1 haplotype, were acclimated in November at semioutdoor conditions and measured for NST throughout winter. Our results showed the crucial role of NST for winter survival in C. russula. The individuals that survived winter displayed a higher significant increase in NST during acclimation, associated with a significant gain in body mass, presumably from brown fat accumulation. The NST capacity (ratio of NST to basal metabolic rate) was exceptionally high for such a small species. NST was significantly affected by a gender x haplotype interaction after winter-acclimation: females bearing the H1 haplotype displayed a better thermogenesis at the onset of the breeding season, while the reverse was true for males. Altogether, our results suggest a sexually antagonistic cyto-nuclear selection on thermogenesis. PMID- 15660956 TI - Herb-drug interactions: an overview of the clinical evidence. AB - Herbal medicines are mixtures of more than one active ingredient. The multitude of pharmacologically active compounds obviously increases the likelihood of interactions taking place. Hence, the likelihood of herb-drug interactions is theoretically higher than drug-drug interactions, if only because synthetic drugs usually contain single chemical entities. Case reports and clinical studies have highlighted the existence of a number of clinically important interactions, although cause-and-effect relationships have not always been established. Herbs and drugs may interact either pharmacokinetically or pharmacodynamically. Through induction of cytochrome P450 enzymes and/or P-glycoprotein, some herbal products (e.g. St John's wort) have been shown to lower the plasma concentration (and/or the pharmacological effect) of a number of conventional drugs, including cyclosporine, indinavir, irinotecan, nevirapine, oral contraceptives and digoxin. The majority of such interactions involves medicines that require regular monitoring of blood levels. To date there is less evidence relating to the pharmacodynamic interaction. However, for many of the interactions discussed here, the understanding of the mechanisms involved is incomplete. Taking herbal agents may represent a potential risk to patients under conventional pharmacotherapy. PMID- 15660957 TI - Immunomodulatory therapy for chronic hepatitis B virus infection. AB - Hepatitis B virus (HBV) is one of the most prevalent viral pathogens of man with around 350 million chronically infected patients. It has been postulated that in persistently infected individuals the HBV-specific immune response is too weak to eliminate HBV from all infected hepatocytes, but sufficiently strong to continuously destroy HBV-infected hepatocytes and to induce chronic inflammatory liver disease. The primary aim in the treatment of chronic hepatitis B is to induce sustained disease remission and prevent serious complications like liver failure and/or hepatocellular carcinoma. The recent emergence of drug-resistant HBV mutants and post-treatment relapse as a consequence of nucleoside analogue monotherapy emphasizes that the principal goal should be to stimulate a successful immune response. In this paper we will focus on the immune response to HBV and we will review reported data on immunotherapeutic strategies like immunomodulatory drugs (cytokines and Thymic derivates) and vaccine therapies using currently available recombinant anti-HBV vaccines, lipopeptide-based T cell vaccine and newly developed genetic vaccines. PMID- 15660958 TI - Pharmacoepidemiology of anabolic androgenic steroids: a review. AB - Non-prescribed use of anabolic androgenic steroids (AAS) has been associated with a number of physical and psychiatric/behavioural complications, some of which are potentially lethal. Here, we review both observational and experimental studies on human subjects concerned with such side-effects. The only physical complication of AAS use that receives definitive support from such investigations is unfavourable changes in blood lipid profiles. Support for various psychiatric complications has also been provided by a number of cross-sectional studies, most involving comparisons between weight-training individuals who use or do not use AAS. Certain of these complications, in particular hypomania and increased aggressiveness, have been confirmed in some, but not all, randomized controlled studies. Epidemiological attempts to determine whether AAS use triggers violent behaviour have failed, primarily because of high rates of non-participation. Studies regarding the prevalence of AAS use in different populations typically report life-time prevalences of 1-5% among adolescents. However, the life-time prevalence (i.e. use on at least one occasion) is of doubtful relevance in attempting to estimate the number of individuals at risk for side-effects, as most of these complications appear to develop during prolonged use of AAS. Furthermore, it is reasonable to assume that the symptoms and signs of AAS use are often overlooked by healthcare professionals, so that the number of cases of possible AAS-related complications is virtually unknown. These limitations, together with an apparently low prevalence of prolonged AAS use among the general population, indicate that future epidemiological research in this area should focus on retrospective case-control studies and, perhaps, also on prospective cohort studies of populations selected for a high prevalence of AAS use, rather than attempting to perform large-scale population-based studies. PMID- 15660959 TI - G protein-coupled receptors: a count of 1001 conformations. AB - G protein-coupled receptors (GPCRs) were initially regarded to adopt an inactive and an active conformation and to activate a single type of G protein. Studies with recombinant cell systems have led to a more complex picture. First, GPCRs can activate distinct G protein species. Second, GPCR multistate models have been invoked to explain their complex behaviour in the presence of agonists, antagonists and other binding partners. The occurrence of intermediate receptor conformational states during GPCR activation and antagonist binding is suggested by fluorescence measurements and studies with constitutively active receptor mutants and insurmountable antagonists. Different agonists may trigger distinct effector pathways through a single receptor by dictating its preference for certain G proteins (i.e. 'agonist trafficking'). Structural modification and exogenous and endogenous (e.g. other cellular proteins, lipids) allosteric modulators also affect ligand-GPCR interaction and receptor activation. These new developments in GPCR research could lead to the development of more selective therapeutic drugs. PMID- 15660960 TI - Time course of oxidative stress, lesion and edema after intrastriatal injection of malonate in rat: effect of alpha-phenyl-N-tert-butylnitrone. AB - The aim of this study was to characterize the model of oxidative stress consisting in the infection of malonate (3 mumol), an inhibitor of mitochondrial complex II, in the rat striatum. The striatal concentrations of both the reduced and oxidized forms of glutathione (the major endogenous antioxidant) were determined at various times after malonate injection (1-4 h) in order to evaluate the evolution of oxidative stress. The progression of lesion size and edema was also determined up to 24 h after malonate administration. Finally, the effect of alpha-phenyl-N-tert-butylnitrone (PBN), an antioxidant nitrone, was studied. The levels of reduced glutathione (GSH) progressively decreased after malonate injection up to 40% of those of sham animals at 4 h. An increase in the concentrations of oxidized glutathione (GSSG) was also observed as early as 1 h after malonate administration which was maintained up to 4 h. The size of the lesion was maximal within 2 h of malonate injection, whereas edema continued to increase between 2 and 24 h. Injection of PBN at 100 mg/kg i.p. 30 min before and 2 h after malonate administration abolished the GSSG increase caused by malonate but did not modify the drop in GSH. This moderate antioxidant effect of PBN was associated with a slight decrease of the lesion area at two levels (10.7 and 10.2 mm anterior to the interaural line), but the lesion volume remained unchanged. By contrast, PBN reduced edema by 30%. Taken together, these results show that malonate induced a severe oxidative stress leading to the rapid development of the lesion. PBN demonstrates anti-edematous properties that are not sufficient to reduce the lesion. PMID- 15660961 TI - On the relation of nitric oxide to nifedipine-induced gingival hyperplasia and impaired submandibular glands function in rats in vivo. AB - Calcium-channel blockers such as nifedipine could be associated with gingival overgrowth. The aim of this study was to examine the role of nitric oxide (NO) on nifedipine-induced gingival hyperplasia along with submandibular secretory function in rats. Animals in divided groups received nifedipine (250 mg/kg diet) alone and in combination with L-arginine (2.25% w/v) or N(omega)-nitro-L-arginine methyl ester (L-NAME) (0.7% w/v) in drinking water for 20 days. Controls received only tap water. Pure submandibular saliva was collected intraorally by micropolyethylene cannula and the mandibular gingiva was examined by means of dissecting microscope for signs of redness, thickness, inflammation and exuda. Twenty-day nifedipine treatment induced gingival hyperplasia accompanied with reduced salivary flow rate and concentrations of total protein, epidermal growth factor (EGF) and calcium in comparison with controls. Co-treatment of animals with nifedipine and L-arginine protected from gingival hyperplasia and retained flow rate, and concentrations of total protein, EGF and calcium in normal levels. Co-treatment of animals with nifedipine and L-NAME potentiated nifedipine-induced gingival hyperplasia and reductions in flow rate and concentrations of total protein, EGF, and calcium. It is concluded that nifedipine-induced gingival hyperplasia is associated with salivary dysfunction. Activation of cGMP-dependent positive signal-transduction mechanisms in salivary glands might be the mechanism for protective effects of NO against nifedipine-induced gingival hyperplasia. PMID- 15660963 TI - The effect of methotrexate and glucocorticosteroids on apoptosis of phythemaglutinin-stimulated mononuclear cells from peripheral blood. AB - Immunosuppressive drugs are widely used in the therapy of autoimmune disorders. Induction of apoptosis is a key mechanism to destroy autoreactive T cells and attenuate immune responses. The aim of the study was to evaluate the apoptotic properties of methotrexate (MTX) and glucocorticosteroids (GS), dexamethasone, methylprednisolone, cortisone in cultured phythemaglutinin-stimulated mononuclear cells (MNC) from healthy subjects. Apoptosis of MNC was measured using two methods: Annexin V and TUNEL. The increase in MTX and GS concentrations led to progressive increase in the percentage of apoptotic cells. There was no statistically significant difference in percentage of apoptotic cells between cultures with MTX + GS and MTX alone. The results suggest that MTX and GS did not act synergistically, the addition of GS to MTX did not enhance the pro-apoptotic properties of MTX. PMID- 15660962 TI - Reactive oxygen species mediate abnormal contractile response to sympathetic nerve stimulation and noradrenaline in the vas deferens of chronically diabetic rats: effects of in vivo treatment with antioxidants. AB - Previous studies suggest that a link exists between increased oxidative stress and diabetic neuropathy. Moreover, antioxidants may protect neurones from the degenerative effects of reactive oxygen species. In our study, we used streptozotocin (STZ)-diabetic rats in a 8-month chronic diabetes model to study the effects of in vivo treatment with stobadine (ST), a pyridoindole antioxidant, and vitamin E. STZ-diabetic rats were treated with ST (24.7 mg/kg/day), vitamin E (D,L-alpha-tocopheryl acetate, 400-500 IU/kg/day) or ST plus vitamin E through an intra-oral catheter for a 8-month period beginning 10 days after STZ injection. Blood glucose and HbA1c levels were increased in diabetic rats by about 400 and 100%, respectively. Antioxidant treatment significantly decreased haemoglobin glycosylation (P < 0.05). We also determined the effects of chronic diabetes on sympathetic neurotransmission by measuring the contractility of isolated vas deferens. Furthermore, we investigated contractions elicited by electrical field stimulation (EFS) (1-64 Hz) which were significantly decreased in diabetic rats when compared with control rats. Treatment with ST or vitamin E alone partly enhanced the amplitude of the contractions induced by EFS, but a combination of ST and vitamin E treatment showed no additional effects. Contractile response of the vas deferens to exogenous noradrenaline, was increased in diabetic rats when compared with control rats. While the addition of vitamin E alone had no effect, ST completely returned noradrenaline-induced contractions to basal levels. The tension induced by 120 mm KCl was not statistically different among the experimental groups. In normal rats, EFS-induced contractions were significantly inhibited by pyrogallol (10(-4) m), a free-radical generator. Percentage inhibition of pyrogallol on EFS (32 Hz)-induced contractions in ring sections was 48 +/- 5.8 in control, 75 +/- 5.5 in untreated-diabetic, 54 +/- 2.7 in ST-treated diabetic, and 58 +/- 4.7 in vitamin E-treated diabetic rats. Combining both ST and vitamin E treatment had the same effects as each antioxidant alone with a percent inhibition of 48 +/- 6.8. These results are consistent with the degenerative changes seen in sympathetic nerves and the abnormal function observed in chronically diabetic rats, leading to a decrease in EFS response and an increase in response to adrenergic agonists in the vas deferens. Furthermore, we demonstrated that reactive oxygen species are responsible for impaired sympathetic neurotransmission and abnormal function of diabetic vas deferens, and that a combination of antioxidants may be better for the therapy of reproductive system disabilities in male diabetics. PMID- 15660964 TI - Experimental evidence for in vitro fluid transport in the presence of a traditional medicinal fruit extract across rat everted intestinal sacs. AB - The present study was designed to investigate the effects of aqueous fruit extract of Momordica charantia (MC), a traditional medicinal plant, on the transport of fluid in vitro. Everted intestinal sacs from rats were mounted in an organ bath containing Krebs solution. We compared the effect of MC extract on water transport with increasing inorganic phosphate concentration with or without D-glucose in the buffer. In the control experiments, fluid uptake was enhanced significantly (P < 0.05) at high inorganic phosphate concentration (8-10 mM) in the presence of 5.5 mM D-glucose. Addition of 3.0 mg/mL MC extract to the serosal side inhibits the uptake of fluid significantly (P < 0.05). At high inorganic phosphate concentration (8-10 mM), fluid uptake was not inhibited (P > 0.05) when incubated with 3.0 mg/mL MC fruit extract. It is hypothesized that an increase in inorganic phosphate enhances oxidative phosphorylation thereby increasing the fluid uptake across everted intestinal sacs of rat. These findings seem to indicate that the MC-induced reduction on intestinal fluid absorption capacity could be mainly the result of an interference with the carrier-mediated coupled entrance of glucose and Na(+) across the brush-border membrane. PMID- 15660965 TI - Protective effect of taurine against alendronate-induced gastric damage in rats. AB - Alendronate (ALD) causes serious gastrointestinal adverse effects. The aim of this study was to investigate whether taurine (TAU), a semi-essential amino acid and an antioxidant, improves the alendronate-induced gastric injury. Rats were administered 20 mg/kg ALD by gavage for 4 days, either alone or following treatment with TAU (50 mg/kg, i.p.). On the last day of treatment, following drug administration, pylorus ligation was performed and 2 h later, rats were killed and stomachs were removed. Gastric acidity and tissue ulcer index values, lipid peroxidation and glutathione (GSH) levels, myeloperoxidase (MPO) activity as well as the histologic appearance of the stomach tissues were determined. Chronic oral administration of ALD induced significant gastric damage, increasing lipid peroxidation, MPO activity and collagen content, as well as decreasing tissue GSH levels. Treatment with TAU prevented the damage and also the changes in biochemical parameters. Findings of the present study suggest that ALD induces oxidative gastric damage by a local irritant effect, and that TAU ameliorates this damage by its antioxidant and/or membrane-stabilizing effects. PMID- 15660966 TI - CYP2C9 and CYP2C19 genetic polymorphisms: frequencies in the south Indian population. AB - The aim of the study was to establish the frequencies of CYP2C9*1, *2, *3 and CYP2C19*1, *2 and *3 in the south Indian population and to compare them with the inter-racial distribution of the CYP2C9 and CYP2C19 genetic polymorphisms. Genotyping analyses of CYP2C9 and CYP2C19 were conducted in unrelated, healthy volunteers from the three south Indian states of Andhra Pradesh, Karnataka and Kerala, by the polymerase chain reaction-restriction fragment-length polymorphism (PCR-RFLP). The allele frequencies of the populations of these three states were then pooled with our previous genotyping data of Tamilians (also in south India), to arrive at the distribution of CYP2C9 and CYP2C19 alleles in the south Indian population. Frequencies of CYP2C9 and CYP2C19 alleles and genotypes among various populations were compared using the two-tailed Fisher's exact test. The frequencies of CYP2C9*1, *2 and *3 in the south Indian population were 0.88 (95% CI 0.85-0.91), 0.04 (95% CI 0.02-0.06) and 0.08 (95% CI 0.06-0.11), respectively. The frequencies of CYP2C9 genotypes *1/*1, *1/*2, *1/*3, *2/*2, *2/*3 and *3/*3 were 0.78 (95% CI 0.74-0.82), 0.05 (95% CI 0.03-0.07), 0.15 (95% CI 0.12-0.18), 0.01 (95% CI 0.0-0.02), 0.01 (95% CI 0.0-0.02) and 0.0, respectively. CYP2C19*1, *2 and *3 frequencies were 0.64 (95% CI 0.60-0.68), 0.35 (95% CI 0.31-0.39) and 0.01 (95% CI 0.0-0.03), respectively. As a result of a significant heterogeneity, the data on CYP2C19 genotype frequencies were not pooled. The frequency of CYP2C9*2 mutant alleles in south Indians was higher than in Chinese and Caucasians, while CYP2C9*3 was similar to Caucasians. CYP2C19*2 was higher than in other major populations reported so far. The relatively high CYP2C19 poor metabolizer genotype frequency of 12.6% indicates that over 28 million south Indians are poor metabolizers of CYP2C19 substrates. PMID- 15660967 TI - How appropriate is asthma therapy in general practice? AB - High association between burden of asthma and inadequate disease control make asthma management a major public health issue. We studied asthma management practices of general practitioners (GPs) in France to describe drug therapy and more specifically, to identify correlates of antibiotic prescriptions, a marker of inappropriate asthma management. Patients with persistent asthma aged 17-50 years were evaluated in a 12-month retrospective study using a computerized GPs database (Thales) and a patient survey, in which patients reported hospital contacts, use of oral corticosteroids and recent asthma symptoms. Therapy was described and the correlates of antibiotic prescriptions in the previous year were identified using multivariate logistic regression. During the study period, 16.4% of 1038 patients received one or more prescriptions of theophylline, 31.3% long-acting beta-agonists and 61.6% inhaled corticosteroids. Rates of prescription of antibiotics, expectorants, antihistamines, antitussives and nasal corticosteroids were 57.6, 42.0, 33.0, 19.9, and 14.4%, respectively. In parallel, 15% of patients reported at least one hospital contact for asthma and 43.1% used oral corticosteroids. Antibiotic prescriptions were more likely co prescribed in patients using expectorants [odds ratio (OR) = 13.0, 95% confidence interval (CI) = 8.5-19.8] and antitussives (OR = 6.5, 95% CI = 3.7-11.6). Moreover, patients using courses of oral corticosteroids, and often visiting their GP (more than four times) during the study period were more likely to receive antibiotics. The results were unchanged when analyses were restricted to non-smokers and younger patients (< or = 40 years). Asthma management was sub optimal among asthma patients treated by general practitioners in France. Antibiotics, expectorants, antihistamines, antitussives and nasal corticosteroids were commonly prescribed while asthma controllers were under-used. PMID- 15660968 TI - Chemical, pharmacokinetic and pharmacodynamic properties of statins: an update. AB - Statins are the treatment of choice for the management of hypercholesterolaemia because of their proven efficacy and safety profile. They also have an increasing role in managing cardiovascular risk in patients with relatively normal levels of plasma cholesterol. Although all statins share a common mechanism of action, they differ in terms of their chemical structures, pharmacokinetic profiles, and lipid modifying efficacy. The chemical structures of statins govern their water solubility, which in turn influences their absorption, distribution, metabolism and excretion. Lovastatin, pravastatin and simvastatin are derived from fungal metabolites and have elimination half-lives of 1-3 h. Atorvastatin, cerivastatin (withdrawn from clinical use in 2001), fluvastatin, pitavastatin and rosuvastatin are fully synthetic compounds, with elimination half-lives ranging from 1 h for fluvastatin to 19 h for rosuvastatin. Atorvastatin, simvastatin, lovastatin, fluvastatin, cerivastatin and pitavastatin are relatively lipophilic compounds. Lipophilic statins are more susceptible to metabolism by the cytochrome P(450) system, except for pitavastatin, which undergoes limited metabolism via this pathway. Pravastatin and rosuvastatin are relatively hydrophilic and not significantly metabolized by cytochrome P(450) enzymes. All statins are selective for effect in the liver, largely because of efficient first-pass uptake; passive diffusion through hepatocyte cell membranes is primarily responsible for hepatic uptake of lipophilic statins, while hydrophilic agents are taken up by active carrier-mediated processes. Pravastatin and rosuvastatin show greater hepatoselectivity than lipophilic agents, as well as a reduced potential for uptake by peripheral cells. The bioavailability of the statins differs greatly, from 5% for lovastatin and simvastatin to 60% or greater for cerivastatin and pitavastatin. Clinical studies have demonstrated rosuvastatin to be the most effective for reducing low-density lipoprotein cholesterol, followed by atorvastatin, simvastatin and pravastatin. As a class, statins are generally well tolerated and serious adverse events, including muscle toxicity leading to rhabdomyolysis, are rare. Consideration of the differences between the statins helps to provide a rational basis for their use in clinical practice. PMID- 15660969 TI - The incidence of aneuploidy in human oocytes assessed by conventional cytogenetic analysis. AB - Human oocytes failing to fertilize during assisted reproduction are an important source of information for assessing incidence and causal mechanisms of maternal aneuploidy. This review describes the techniques of conventional oocyte chromosome analysis and evaluates the results of 59 studies comprising a total of>10,000 female gametes. The mean rate of aneuploidy (hypohaploidy + hyperhaploidy) amounts to approximately 20%, but this incidence is reduced as soon as possible artifacts introduced by the fixation technique are taken into consideration. It is therefore concluded that a realistic value for numerical abnormalities arising during first meiotic division lies between 12 and 15%. All chromosome groups are affected by aneuploidy but the actually observed frequencies exceed the expected frequencies in groups D, E, and G. Two aneuploidy causing mechanisms have been identified in human oocytes: nondisjunction, resulting in the loss or gain of whole chromosomes, and predivision, resulting in the loss or gain of single chromatids. A brief analysis including only aneuploid complements with one extra or missing chromosome/chromatid shows a slight increase in predivision (52.9%) compared with nondisjunction (47.1%). Finally, suggestions for future studies are given since, for instance, the presentation of results and the use of cytogenetic nomenclature have not been uniform. PMID- 15660970 TI - Heterosis studies for yield and its components in bread wheat over environments. AB - A set of diallel crosses involving 10 parents was made to have information on the extent of heterosis over mid-parent and better parent and inbreeding depression for yield and yield contributing characters under three different environments. Marked heterobeltiosis for grain yield and its important components were observed. For grain yield, 83 crosses showed significant positive heterobeltiosis in all the three sowing dates, however, twenty crosses showed significant consistent heterobeltiosis for grain yield per plant over all the three environments. The maximum heterobeltiosis for grain yield per plant observed was 50.94% (Raj 3765 x HD 2285), 121.08% (PBW 373 x HD 2329) and 93.96% (PBW 373 x HD 2329) under early, normal and late sowing conditions, respectively. Cross PBW 373 x HD 2329 in both early and normal plantings and cross Raj 3765 x HD 2285 under late planting were observed most heterotic for grain yield. The crosses showing heterosis for grain yield were not heterotic for all the characters. Heterosis for grain yield per spike followed by tillers per plant and 1000-grain weight was independently associated with heterosis for grain yield in early and normal plantings. However, heterosis for grain yield per spike, dwarf plant height and tillers per plant contributed maximum towards yield heterosis. Significant inbreeding depression was recorded frequently for yield and yield contributing traits, however, in a few traits it was observed significant negative indicated that F(2) was superior to F(1) considered desirable combination for trait(s). The study reveals good scope for commercial exploitation of heterosis as well as isolation of pure lines among the progenies of heterotic F(1) for improvement of yield levels in bread wheat. PMID- 15660971 TI - Combining ability in the F1 and F2 generations of diallel cross in hexaploid wheat (Triticum aestivum L. em. Thell). AB - The F(1) and F(2) progenies of a ten-parent diallel cross (excluding reciprocals) of hexaploid wheat (Triticum aestivum L. em. Thell) were analyzed for combining ability for quantitative and quality traits. The results indicated significant differences among the parents for general combining ability (gca) and crosses for specific combining ability (sca) for all the characters studied. The gca and sca components of variance were significant for all the traits. However, the gca component of variance was predominant indicating the predominance of additive gene effects for the traits studied. Among the parents Durgapura 65, HD 2285, Lok 1, Raj 1972 and HD 2329 were the best general combiners for grain yield and average to high combiners for tillers per plant, grain yield per spike, grains per spike and 1000-grain weight. The best specific crosses for grain yield were Sonalika x WH 157, HD 2428 x Durgapura 65, Durgapura 65 x Sonalika, HD 2428 x Lok 1 and CPAN 3004 x Raj 1972. The parent Raj 1972, Lok-1 and HD 2285 were the best general combiners for grain yield and protein content, however, Raj 3077 was the best general combiner for protein content. The most suitable specific crosses for protein content were HD 2329 x HD 2285, HD 2428 x Raj 1972 and CPAN 3004 x WH 157. Most of the specific crosses for grain yield as well as protein content involved high x average, average x average and average x poor general combiners. To ensure further increase in grain yield along with high protein, combinations of desirable yield components is advocated. Inclusion of F(1) hybrids showing high sca and having parents with good gca, into multiple crosses and/or bi parental mating, or diallel selective mating could prove a worthwhile approach for further improvement of grain yield in bread wheat. PMID- 15660972 TI - Chromosomal distribution of telomeric and telomeric-associated sequences in Hordeum chilense by in situ hybridization. AB - The chromosomal distribution of telomeric repeat pAtT4 from Arabidopsis thaliana and telomeric associated repetitive sequence HvT01 from Hordeum vulgare have been studied by FISH (fluorescence in situ hybridization) in two accessions (H1 and H7) of Hordeum chilense. The telomeric sequence pAtT4 is present at the end of all chromosome arms in H1 and H7 accessions. In contrast, the telomeric associated sequence homologous to HvT01 showed variability for size, intensity and position of the signals for each line. In H1, HvT01 was present in every chromosome whereas only four chromosomes were labeled in H7 accession. Physical distribution of GAA-satellite sequence on both H1 and H7 metaphase chromosomes was also studied. Polymorphism for hybridization signals between the two accessions for GAA-banding pattern was also found. Based on differences in position and intensity of the hybridization signals found for both GAA and HvT01 homologous sequences, karyotypes for the in situ hybridization patterns are presented for H1 and H7 accessions of H. chilense. PMID- 15660973 TI - Genetic analysis of non-essential amino acid contents in rice (Oryza sativa L.) across environments. AB - Genetic main effects and genotype x environment (GE) interaction effects for 7 non-essential amino acids in milled rice were analyzed for two year data by using the genetic models based on mixed linear model approaches for quantitative traits of triploid endosperm. Nine cytoplasmic, male sterile lines as females and five restoring lines as males were introduced in a diallel cross in two environments. It was found that the content of non-essential amino acids including Asp, Ser, Glu, Gly and Tyr were mainly controlled by genetic main effects, whereas the content of Ala or Pro was mainly affected by GE effects. In genetic main effects, the cytoplasmic and maternal genetic effects were preponderant for all traits of non-essential amino acids, indicating that selection for improving these traits based on the maternal plant would be more effective than on seeds. The total narrow-sense heritabilities for non-essential amino acids were 70.9-85.9%. By predicating the genetic effects of parents, the total genetic effects from Xieqingzao, V20, Zuo 5 and Zhenshan 97 were mainly negative and these parents would decrease the content of most essential amino acids. Since parents of Zhenan 3, Yinchao 1, T49, 26715, 102 and 1391 had possessed a positive value of most total genetic effects, these parents could be chosen as optimal parents for increasing the content of most non-essential amino acids. PMID- 15660974 TI - Meiotic pairing in the hybrid (Zea diploperennis x Zea perennis) x Zea mays and its reciprocal. AB - Genomic formulae, fertility, chromosome pairing, and the cryptic intergenomic pairing (induced by using diluted colchicine solution) were analysed in the tri hybrid (MDP), obtained by crossing DP40 (2n=40, which was inferred in previous studies to have originated from the fusion of an unreduced gamete of Zea diploperennis with a normal gamete of Z. perennis) with the maize inbred line Zm40 (2n=40). MDP (2n=40) showed a higher fertility (90% of the seeds are viable) than Zm40 (60%) and DP40 (80%). A regular migration of 20 chromosomes to each pole occurred in 92% of the cells in anaphase I, while bridges were observed in the other 8% of the cells. When Zm40 was used as female of the crossing (Zm40 x DP40), ears were similar to corn. Conversely, ears resembled those of the wild species when cytoplasm was donoured by Zd. Then, it can be stated the existence of cytoplasmic influence on MDP ear type. MDP had almost no I or III, with an average of 0.04I + 10.90II + 0.01III + 4.50IV. The most frequent meiotic configuration was 10II + 5IV (43.93% of the cells). On average, 33.81 chiasmata/cell were observed (17.34, 0.05 and 16.42 average numbers of chiasmata/cell in bivalents, trivalents and tetravalents, respectively). It can be inferred that the 5IV were the product of homoeologous chromosome pairing of A genomes from the three species. On the other hand, the 10II configuration suggests separate pairing of the 5 homologous B chromosomes from maize and the 5 homoeologous B chromosomes from Zp and Zd. PMID- 15660975 TI - Influence of climatic conditions on the mutations in pollen mother cells of Tradescantia clone 4430 and implications for the Trad-MCN bioassay protocol. AB - The present was study aimed at investigating the influence of relative humidity and temperature on spontaneous and pollution-induced mutation rates during exposure and recovery periods in the Trad-MCN test. Cuttings of Tradescantia clone 4430 were exposed to a negative control, to 4 mM maleic hydrazide (MH), and to a polluted water sample under varying conditions of air temperature and humidity in climatic chamber experiments. The relative humidity did not affect the spontaneous mutation rate in the clone investigated, but was negatively correlated with the frequency of pollution-induced mutations. Low temperature caused an increase in the number of micronuclei in the negative control, but no comparable response in polluted samples. At an extremely high temperature, signs of strong physiological damage and/or of a meiotic delay of pollen maturation were detected. When the temperature increased gradually and the extreme value was maintained only for short time, such detrimental effects were not observed. Subsequent treatment with high and low temperatures, by contrast, resulted in the highest MCN rates of all experiments. Our studies point to the possibility of producing irregular results of the Trad-MCN test if the influence of climatic factors has not sufficiently been considered. PMID- 15660976 TI - Analysis of genetic marker-phenotype relationships by jack-knifed partial least squares regression (PLSR). AB - The utility of a relatively new multivariate method, bi-linear modelling by cross validated partial least squares regression (PLSR), was investigated in the analysis of QTL. The distinguishing feature of PLSR is to reveal reliable covariance structures in data of different types with regard to the same set objects. Two matrices X (here: genetic markers) and Y (here: phenotypes) are interactively decomposed into latent variables (PLS components, or PCs) in a way which facilitates statistically reliable and graphically interpretable model building. Natural collinearities between input variables are utilized actively to stabilise the modelling, instead of being treated as a statistical problem. The importance of cross-validation/jack-knifing as an intuitively appealing way to avoid overfitting, is emphasized. Two datasets from chromosomal mapping studies of different complexity were chosen for illustration (QTL for tomato yield and for oat heading date). Results from PLSR analysis were compared to published results and to results using the package PLABQTL in these data sets. In all cases PLSR gave at least similar explained validation variances as the reported studies. An attractive feature is that PLSR allows the analysis of several traits/replicates in one analysis, and the direct visual identification of individuals with desirable marker genotypes. It is suggested that PLSR may be useful in structural and functional genomics and in marker assisted selection, particularly in cases with limited number of objects. PMID- 15660977 TI - Resistance to barley scald (Rhynchosporium secalis) in the Ethiopian donor lines 'Steudelli' and 'Jet', analyzed by partial least squares regression and interval mapping. AB - The resistance of barley (Hordeum vulgare L.) to Rhynchosporium secalis (scald) has been investigated in two crosses between the susceptible cv. 'Ingrid' and two resistant Ethiopian landraces, 'Steudelli' and 'Jet'. Doubled haploids were inoculated in replicated tests using two isolates of R. secalis, '4004' and 'WRS1872'. Expression of resistance differed widely between replicated tests. AFLP, SSR and RFLP markers were used to develop chromosome maps. Results have been analysed using partial least squares regression (PLSR) and interval mapping. In PLSR the major covariance structures or 'latent variables' between X (markers) and Y (isolates, tests) are modelled as principal components and their optimal number determined by cross-validation. In 'Steudelli' two QTL were detected, one on each of chromosomes 3H and 7H, in 4 out of 5 tests, while in 'Jet' only one (different) allele at the 3H locus was found. The validated R(2) varied between 11.0% and 64.9% in the replicated tests with '4004'.With isolate 'WRS1872' the 7H locus and another 3H locus were detected. By interval mapping the QTL detected were less stable and generally gave lower R(2) values than PLSR. PLSR does not depend on maps, but interval mapping based on values predicted by PLSR had R(2) around 90%. It is suggested that PLSR may be a useful tool in QTL analysis. PMID- 15660978 TI - Interaction among different heterocromatic variants in the grasshopper Dichroplus elongatus. AB - Simultaneous chromosome polymorphisms for supernumerary elements allow us to analyse the relationships among different forms of heterochromatic variation in nature. We report simultaneous variation patterns for supernumerary segments in chromosomes S10 (SS10), S9 (SS9) and S6 (SS6) and B chromosomes in nine populations of the grasshopper Dichroplus elongatus from two biogeographic provinces from east Argentina. Our results show spatial chromosome differentiation for three out of four supernumerary heterochromatic variants (B chromosomes, SS6 and SS10). The incidence of B chromosomes was negatively correlated with the SS10 frequency. The distribution pattern analysis shows different degree of differentiation among populations for each supernumerary heterochromatic variant suggesting that the detected chromosome variation cannot be explained by interaction between migration and genetic drift. Moreover, the observed population chromosome differentiation was not in agreement with the hierarchical analysis of molecular of heterogeneity at mitochondrial DNA level (mtDNA). The present results point out the importance of the interaction among heterochromatic variants in the chromosome intraspecific variation in east Argentina natural populations of the grasshopper D. elongatus. PMID- 15660979 TI - Inter simple sequence repeat analysis of genetic diversity and relationships in cultivated barley of Nordic and Baltic origin. AB - This study evaluates putative changes of genetic diversity and relationships of barley in the Nordic and Baltic countries that might have taken place during the last century as a result of commercial breeding. Four ISSR primers were used to analyse 227 accessions, yielding a total of 47 polymorphic loci. Shannon-Weaver diversity values for each locus ranged from 0.012 to 0.693. Overall, there were no significant changes of genetic diversity observed over time. A significant decrease of diversity was, however, observed in material from the southern parts of the Nordic and Baltic countries. In material from the northern parts no decrease of diversity was observed. The genetic diversity of six-rowed barley bred in the middle of the 20th century was low, but there was no significant difference between modern accessions and landraces or old cultivars. The magnitude in changes of genetic diversity differed also in material from different countries of origin. A cluster analysis clearly separated the material into two groups. The first cluster included 86.5% of all six-rowed accessions, whereas the second cluster contained 97.4% of all two-rowed accessions. PMID- 15660981 TI - Management of oral bleedings with recombinant factor VIIa in children with haemophilia A and inhibitor. AB - Dental extraction in patients with haemophilia A and high-titre inhibitor is always a high-risk procedure, which often presents a lot of problems associated with bleeding. Prothrombin complex concentrates or recombinant activated factor VII (rFVIIa) has been used to control bleeding. rFVIIa was administered to five boys with severe haemophilia A complicated with inhibitor, who underwent seven dental extractions. The age of the patients ranged between 8 and 13 years (median 10 years). The concentrate was administered in doses of 90-100 microg kg(-1) body weight. Duration in the therapy and intervals between rFVIIa doses depended on the severity of bleeding. rFVIIa was proven to be highly effective and no side effects of the product were observed. PMID- 15660982 TI - A Bayesian approach to the assessment of inhibitor risk in studies of factor VIII concentrates. AB - The assessment of inhibitor risk is a crucial component in the clinical development of new and modified factor VIII (FVIII) preparations. There has been a recent discussion about the design of studies and the assessment of inhibitors and inhibitor risk in such studies at a recent FDA-sponsored FVIII Inhibitor Workshop, and new requirements for the success of these trials have been proposed to evaluate inhibitor data based on the use of an upper 95% confidence bound. We review the consequences of these requirements and demonstrate that for any product to succeed, it must have an extremely low underlying risk of inhibitor development. Furthermore, several existing commercially available FVIII products with an excellent safety record would not necessarily pass these endpoints. As a result, we propose an alternative set of acceptance criteria based on a Bayesian statistical paradigm. This approach is based on the determination of a probability that the product in question actually has an inhibitor risk below some pre-set limit, a concept that we believe is more intuitive than the traditional confidence interval method. We show that all existing products would pass this approach, but a product (Bisinact) with known inhibitor risk would not. PMID- 15660983 TI - Rituximab for autoimmune haemophilia: a proposed treatment algorithm. AB - We previously reported durable complete responses following brief courses of rituximab and prednisone with or without cyclophosphamide in four patients with autoimmune haemophilia and inhibitor titres of 5-60 BU. We report here responses to this monoclonal anti-CD20 antibody in four additional patients, including two patients with inhibitor titres >200 BU. Factor VIII levels became normal 2 35weeks after 4 or 8 weekly doses of rituximab, brief courses of prednisone and in one patient immunoglobulin. Complete responses are ongoing at 10 months in two patients. Two patients relapsed: a patient whose initial inhibitor titre was 525 BU relapsed at 3.5 months and a long-term prednisone-dependent patient at 8.5 months. Both responded to second courses of rituximab and prednisone and are in remission. Our experience suggests that rituximab is a safe and effective addition to immunosuppression with prednisone and cyclophosphamide to treat autoimmune haemophilia, and may permit early discontinuation or even avoidance of these potentially toxic agents. High-titre inhibitor patients, however, may require multiple courses of rituximab or the addition of cyclophosphamide. Pending randomized studies, we propose an algorithm based on our experience and other reports for incorporating rituximab in the treatment of this rare disorder. PMID- 15660984 TI - Treatment of factor XI inhibitor using recombinant activated factor VIIa. AB - A 30-year-old female with severe factor XI deficiency of 0-2% acquired factor XI inhibitor following many infusions for fresh frozen plasma (FFP) for surgical procedures starting at 4 years of age. Seven months before this inhibitor was diagnosed, surgery was complicated by prolonged bleeding resistant to FFP, requiring epsilon aminocaproic acid (EACA) and surgical packing. The inhibitor was measured at 2.2 Bethesda units, 7 months since the last FFP. The inhibitor was confirmed as specific anti-XI and anti-XIa binding by patient's IgG to immobilized factor XI and factor XIa from whole plasma and purified IgG. For repair of a painful anterior cruciate ligament (ACL) defect she was given recombinant factor VIIa (rVIIa) at 90 mug kg(-1), starting one-half hour preoperatively and continued every 2 h for 8 h when haemostasis was complete. Thereafter the rVIIa was given every 3 h for two doses, and then every 4 h for four doses at which time she was discharged on EACA which was continued for 6 days. There was excellent haemostasis during and following the surgery. There was no evidence of consumptive coagulopathy, with no change in the fibrinogen, platelet count, or D-D dimer; and no increase of platelet factor 4, beta thromboglobulin, or prothrombin fragment F 1.2. The thrombin-antithrombin complex increased over baseline after 24 h. There was no postoperative deep vein thrombosis or pulmonary embolus. In this patient with a factor XI inhibitor, the recombinant factor VIIa was effective and safe, ensuring adequate haemostasis with no thrombotic complications. This product which was designed for patients with inhibitors to factor VIII or factor IX, and factor VII deficiency, has now been given successfully to four patients with factor XI inhibitors. PMID- 15660985 TI - Pregnancy and oral contraceptives in factor V deficiency: a study of 22 patients (five homozygotes and 17 heterozygotes) and review of the literature. AB - Information on the effect of pregnancy or oral contraceptives (OC) in congenital factor V (FV) deficiency is scanty. The personal investigation of five homozygous and 17 female heterozygous showed that patients with severe deficiency bleed considerably at the time of delivery. However, bleeding can be controlled properly by administration of fresh frozen plasma with excellent foetal outcome. The safe level for adequate haemostasis seems around 25% of normal. On the contrary, heterozygote patients show no significant postpartum bleeding and therefore need no substitution therapy. Oral contraceptives were taken and well tolerated by four of our homozygous patients and appear to be beneficial because they cause a decrease in menometrorrhagies thereby improving the anaemia and decreasing transfusional needs. One patient took hormonal replacement therapy with no undue effects. No thrombosis was noted in the propositae during oral contraceptive therapy. The review of the literature has allowed the gathering of information on 20 additional pregnancies. The foetal outcome was satisfactory in every instance. Excessive bleeding was noted in 11 pregnancies. In seven of the remaining pregnancies, no undue bleeding was noted thanks to appropriate substitution therapy. In the remaining two pregnancies no bleeding was noted and no substitution therapy was given. No data are apparently available in the literature about the use of OCs in FV deficiency. PMID- 15660986 TI - Molecular characterization of two novel mutations causing factor X deficiency in a Chinese pedigree. AB - Factor X (FX) deficiency is a rare bleeding disorder inherited as an autosomal recessive trait. In this study, we investigated the molecular basis of FX deficiency in a Chinese pedigree. The proposita showed a markedly prolonged activated partial thromboplastin time and a mild prolongation of prothrombin time. The levels of FX antigen and FX activity were 58.6% and 2.5%, respectively. Molecular analysis revealed that the proposita was compound heterozygous for two novel mutations: IVS1 + 1G > A and G1185A (Arg347His). The aberrant transcripts from the IVS1 + 1G > A mutant allele were not detected by analyzing the splicing pattern of ectopic transcripts in leukocytes of the patient with nested polymerase chain reaction after reverse transcription. We thus hypothesize that the mRNA molecules originating from the IVS1 + 1G > A mutation were rapidly destroyed in vivo. Site-directed mutagenesis of FX cDNA was used to introduce FXG1185A mutation, and wild-type as well as mutant FX proteins were expressed by transient transfection in HEK 293 cells. Normal FX antigen levels both in the conditioned media of cells expressing the mutant and in cell lysates were detected by an enzyme-linked immunoadsorbent assay. Evaluation of wild-type and mutant coagulant activity demonstrated that the FX molecules carrying the Arg347His mutation have dramatically decreased activity. PMID- 15660987 TI - Identification of new dinucleotide-repeat polymorphisms in factor VIII gene using fluorescent PCR. AB - Haemophilia A is an X-linked inherited bleeding disorder. Linkage diagnosis using polymorphic markers in the factor VIII gene is used to archive the carrier detection and prenatal diagnosis. The objective of this study was to establish the allele frequency and heterozygosity rate (HR) of two new intragenic markers (Intron 1 and 24) and other markers (Intron 13 and 22) using fluorescent PCR. Five hundred unrelated healthy women were screened and haemophilic family was studied for carrier detection and prenatal diagnosis. We observed five different alleles of Intron 1, 10 of Intron 24, nine of Intron 13 and six of Intron 22. The observed HR for Intron 1, 24, 13 and 22 were 34.0, 35.2, 53.0 and 42.6%, while the expected HR were 33.6, 36.3, 50.1 and 44.3%, respectively. Heterozygosity rate with the combined use of all four intragenic markers was 76.6% (383/500). In prenatal diagnosis of a haemophilic family, a pregnant woman was heterozygous with three intragenic (Intron 1, 13 and 22) and one extragenic St14 VNTR (DXS52) markers. She was considered to be a carrier, and she carried a male foetus by AMXY PCR and chromosome analysis of amniocytes. Foetus did not have mutant haplotype as his uncle, suggesting a normal male status. Our study demonstrates the utility of two new intragenic markers in FVIII gene for carrier detection and prenatal diagnosis of haemophilic families. PMID- 15660988 TI - Effects of haemophilic arthropathy on health-related quality of life and socio economic parameters. AB - Although prophylactic treatment is advised for all children with severe haemophilia, the optimal regimen is still under discussion. Should all joint bleeds be prevented, or can a limited amount of arthropathy be tolerated in adulthood without loss of quality of life? To answer this question, the effect of haemophilic arthropathy on health-related quality of life (HRQoL) needs to be quantified. In a retrospective study, the effect of arthropathy on HRQoL and socio-economic parameters was assessed in a single-centre cohort of 96 patients with severe and moderate haemophilia with a minimum age of 13 years. Arthropathy was measured by the radiological Pettersson score of the elbows, knees and ankles (maximum: 78 points). HRQoL was assessed by the Short Form 36 (SF36), measuring eight domains of health. Labourforce participation and medical consumption were assessed using a separate questionnaire. Patients were studied at a mean age of 28.6 years (range: 13-54), the mean time between evaluation and the last Pettersson score was 0.4 years (SD: 1.1). The overall median Pettersson score was 13 (range: 0-78). There was a trend towards lower quality of life with increasing Pettersson scores and age, especially in the physical domains of the SF36. An age adjusted analysis showed that arthropathy had a small but significant effect on HRQoL in the domain of 'physical function' of the SF36, but not on its other domains, or on labourforce participation and medical consumption. Thus suggesting that the SF36 can be used to assess the effects of haemophilic arthropathy, especially in the domain of 'physical function'. PMID- 15660989 TI - Measuring health state preferences for hemophilia: development of a disease specific utility instrument. AB - Generic and disease-specific instruments have been used to assess health-related quality of life (HRQoL) in hemophilia. However, HRQoL measures also need to reflect patient preferences for various hemophilia health states. The goal of this project was to develop a disease-specific utility instrument that measures patient preferences for various health states unique to hemophilia. The visual analog scale (VAS) and the standard gamble (SG) methods were used. Study participants (n = 128) were tested and stratified into paediatric and adult groups. Test-retest reliability was demonstrated for both instruments, with r = 0.91 for the VAS and r = 0.79 for the SG. When comparing results by age group, statistically significant differences were observed between paediatric and adult participants with the SG instrument (P = 0.045), with older participants taking more risk overall. However, no significant differences based on age were seen when using the VAS (P = 0.636). Statistically significant differences were observed between the VAS and SG instruments within both the paediatric and adult groups (P < 0.0001). In general, the SG yielded higher preference scores than the VAS for the majority of health states. Results derived from the SG instrument indicate that age can influence patients' preferences regarding their state of health. This can have implications for considering treatment options based on the mean age of the population under consideration. Both instruments demonstrated reliability and validity indicating that they could be used to assess patient preferences in hemophilia. However, preference score differences indicate that the two measures may not be interchangeable. PMID- 15660990 TI - Proceedings of the International Haemophilia Prophylaxis Study Group Meeting, November 2003, Montreal, PQ, Canada. PMID- 15660991 TI - Economic challenges in haemophilia. PMID- 15660992 TI - Intracranial haemorrhage as initial presentation of severe haemophilia B: case report and review of Mayo Clinic Comprehensive Hemophilia Center experience. AB - A neonate who had intracranial haemorrhage (ICH) at birth received a diagnosis of severe haemophilia B at 6 months of age. ICH had been the initial presentation of his bleeding disorder. His family history was negative for haemophilia. Review of our institutional experience as well as the literature indicates that intracranial bleeding as the initial presentation of haemophilia is rare. PMID- 15660993 TI - Who's talking to whom? Epithelial-bacterial pathogen interactions. AB - Our perception that host-bacterial interactions lead to disease comes from rare, unsuccessful interactions resulting in the development of detectable symptoms. In contrast, the majority of host-bacterial interactions go unnoticed as the host and bacteria perceive each other to be no threat. In July 2004, a focused international symposium on epithelial-bacterial pathogen interactions was held in Newcastle upon Tyne (UK). The symposium concentrated on recent advances in our understanding of bacterial interactions at respiratory and gastrointestinal mucosal epithelial layers. For the host these epithelial tissues represent a first line of defence against invading bacterial pathogens. Through the discovery that the innate immune system plays a pivotal role during host-bacterial interactions, it has become clear that epithelia are being utilized by the host to monitor or communicate with both pathogenic and commensal bacteria. Interest in understanding the bacterial perspective of these interactions has lead researchers to realize that the bacteria utilize the same factors associated with disease to establish successful long-term interactions. Here we discuss several common themes and concepts that emerged from recent studies that have allowed physiologists and microbiologists to interact at a common interface similar to their counterparts -- epithelia and bacterial pathogens. These studies highlight the need for further multidisciplinary studies into how the host differentiates between pathogenic and commensal bacteria. PMID- 15660994 TI - Population density-dependent regulation of exopolysaccharide formation in the hyperthermophilic bacterium Thermotoga maritima. AB - Co-cultivation of the hyperthermophiles Thermotoga maritima and Methanococcus jannaschii resulted in fivefold higher T. maritima cell densities when compared with monoculture as well as concomitant formation of exopolysaccharide and flocculation of heterotroph-methanogen cellular aggregates. Transcriptional analysis of T. maritima cells from these aggregates using a whole genome cDNA microarray revealed the induction of a putative exopolysaccharide synthesis pathway, regulated by intracellular levels of cyclic diguanosine 3',5' (cyclic)phosphate (cyclic di-GMP) and mediated by the action of several GGDEF proteins, including a putative diguanylate cyclase (TM1163) and a putative phosphodiesterase (TM1184). Transcriptional analysis also showed that TM0504, which encodes a polypeptide containing a motif common to known peptide-signalling molecules in mesophilic bacteria, was strongly upregulated in the co-culture. Indeed, when a synthetically produced peptide based on TM0504 was dosed into the culture at ecologically relevant levels, the production of exopolysaccharide was induced at significantly lower cell densities than was observed in cultures lacking added peptide. In addition to identifying a pathway for polysaccharide formation in T. maritima, these results point to the existence of peptide-based quorum sensing in hyperthermophilic bacteria and indicate that cellular communication should be considered as a component of the microbial ecology within hydrothermal habitats. PMID- 15660995 TI - Regulation of the hetero-octameric ATP phosphoribosyl transferase complex from Thermotoga maritima by a tRNA synthetase-like subunit. AB - The molecular structure of the ATP phosphoribosyl transferase from the hyperthermophile Thermotoga maritima is composed of a 220 kDa hetero-octameric complex comprising four catalytic subunits (HisGS) and four regulatory subunits (HisZ). Steady-state kinetics indicate that only the complete octameric complex is active and non-competitively inhibited by the pathway product histidine. The rationale for these findings is provided by the crystal structure revealing a total of eight histidine binding sites that are located within each of the four HisGS-HisZ subunit interfaces formed by the ATP phosphoribosyl transferase complex. While the structure of the catalytic HisGS subunit is related to the catalytic domain of another family of (HisGL)2 ATP phosphoribosyl transferases that is functional in the absence of additional regulatory subunits, the structure of the regulatory HisZ subunit is distantly related to class II aminoacyl-tRNA synthetases. However, neither the mode of the oligomeric subunit arrangement nor the type of histidine binding pockets is found in these structural relatives. Common ancestry of the regulatory HisZ subunit and class II aminoacyl-tRNA synthetase may reflect the balanced need of regulated amounts of a cognate amino acid (histidine) in the translation apparatus, ultimately linking amino acid biosynthesis and protein biosynthesis in terms of function, structure and evolution. PMID- 15660996 TI - Neisseria meningitidis NadA is a new invasin which promotes bacterial adhesion to and penetration into human epithelial cells. AB - Neisseria meningitidis is a human pathogen, which is a major cause of sepsis and meningitis. The bacterium colonizes the upper respiratory tract of approximately 10% of humans where it lives as a commensal. On rare occasions, it crosses the epithelium and reaches the bloodstream causing sepsis. From the bloodstream it translocates the blood-brain barrier, causing meningitis. Although all strains have the potential to cause disease, a subset of them, which belongs to hypervirulent lineages, causes disease more frequently than others. Recently, we described NadA, a novel antigen of N. meningitidis, present in three of the four known hypervirulent lineages. Here we show that NadA is a novel bacterial invasin which, when expressed on the surface of Escherichia coli, promotes adhesion to and invasion into Chang epithelial cells. Deletion of the N-terminal globular domain of recombinant NadA or pronase treatment of human cells abrogated the adhesive phenotype. A hypervirulent strain of N. meningitidis where the nad A gene was inactivated had a reduced ability to adhere to and invade into epithelial cells in vitro. NadA is likely to improve the fitness of N. meningitidis contributing to the increased virulence of strains that belong to the hypervirulent lineages. PMID- 15660997 TI - Cell division in cocci: localization and properties of the Streptococcus pneumoniae FtsA protein. AB - We studied the cytological and biochemical properties of the FtsA protein of Streptococcus pneumoniae. FtsA is a widespread bacterial cell division protein that belongs to the actin superfamily. In Escherichia coli and Bacillus subtilis, FtsA localizes to the septal ring after FtsZ, but its exact role in septation is not known. In S. pneumoniae, we found that, during exponential growth, the protein localizes to the nascent septa, at the equatorial zones of the dividing cells, where an average of 2200 FtsA molecules per cell are present. Likewise, FtsZ was found to localize with the same pattern and to be present at an average of 3000 molecules per cell. Consistent with the colocalization, FtsA was found to interact with FtsZ and with itself. Purified FtsA is able to bind several nucleotides, the affinity being highest for adenosine triphosphate (ATP), and lower for other triphosphates and diphosphates. The protein polymerizes in vitro, in a nucleotide-dependent manner, forming long corkscrew-like helixes, composed of 2 + 2 paired protofilaments. No nucleotide hydrolytic activity was detected. Consistent with the absence of an ATPase activity, the polymers are highly stable and not dynamic. These results suggest that the FtsA protein could also polymerize in vivo and the polymers participate in septation. PMID- 15660998 TI - Directed evolution of Vibrio fischeri LuxR for increased sensitivity to a broad spectrum of acyl-homoserine lactones. AB - LuxR-type transcriptional regulators play key roles in quorum-sensing systems that employ acyl-homoserine lactones (acyl-HSLs) as signal molecules. These proteins mediate quorum control by changing their interactions with RNA polymerase and DNA in response to binding their cognate acyl-HSL. The evolutionarily related LuxR-type proteins exhibit considerable diversity in primary sequence and in their response to acyl-HSLs having acyl groups of differing length and composition. Little is known about which residues determine acyl-HSL specificity, and less about the evolutionary time scales required to forge new ones. To begin to examine such issues, we have focused on the LuxR protein from Vibrio fischeri, which activates gene transcription in response to binding its cognate quorum signal, 3-oxohexanoyl-homoserine lactone (3OC6HSL). Libraries of luxR mutants were screened for variants exhibiting increased gene activation in response to octanoyl-HSL (C8HSL), with which wild-type LuxR interacts only weakly. Eight LuxR variants were identified that showed a 100-fold increase in sensitivity to C8HSL; these variants also displayed increased sensitivities to pentanoyl-HSL and tetradecanoyl-HSL, while maintaining a wild type or greater response to 3OC6HSL. The most sensitive variants activated gene transcription as strongly with C8HSL as the wild type did with 3OC6HSL. With one exception, the amino acid residues involved were restricted to the N-terminal, 'signal-binding' domain of LuxR. These residue positions differed from critical positions previously identified via 'loss-of-function' mutagenesis. We have demonstrated that acyl-HSL-dependent quorum-sensing systems can evolve rapidly to respond to new acyl-HSLs, suggesting that there may be an evolutionary advantage to maintaining such plasticity. PMID- 15660999 TI - Environmental and genetic regulation of the phosphorylcholine epitope of Haemophilus influenzae lipooligosaccharide. AB - In response to environmental signals in the host, bacterial pathogens express factors required during infection and repress those that interfere with specific stages of this process. Signalling pathways controlling virulence factors of the human respiratory pathogen, Haemophilus influenzae, are predominantly unknown. The lipooligosaccharide (LOS) outer core represents a prototypical virulence trait of H. influenzae that enhances virulence but also provides targets for innate and adaptive immunity. We report regulation of the display of the virulence-associated phosphorylcholine (PC) epitope on the LOS in response to environmental conditions. PC display is optimal under microaerobic conditions and markedly decreased under conditions of high culture aeration. Gene expression analysis using a DNA microarray was performed to begin to define the metabolic state of the cell under these conditions and to identify genes potentially involved in PC epitope modulation. Global gene expression profiling detected changes in redox responsive genes and in genes of carbohydrate metabolism. The effects on carbohydrate metabolism led us to examine the role of the putative H. influenzae homologue of csrA, a regulator of glycolysis and gluconeogenesis in Escherichia coli. A mutant containing an in-frame deletion of the H. influenzae csrA gene showed increased PC epitope levels under aerobic conditions. Furthermore, deletion of csrA elevated mRNA expression of galU, an essential virulence gene that is critical in generating sugar precursors needed for polysaccharide formation and LOS outer core synthesis. Growth conditions predicted to alter the redox state of the culture modulated the PC epitope and galU expression as well. The results are consistent with a multifactorial mechanism of control of LOS-PC epitope display involving csrA and environmental signals that coordinately regulate biosynthetic and metabolic genes controlling the LOS structure. PMID- 15661000 TI - A master regulator for biofilm formation by Bacillus subtilis. AB - Wild strains of Bacillus subtilis are capable of forming architecturally complex communities of cells known as biofilms. Critical to biofilm formation is the eps operon, which is believed to be responsible for the biosynthesis of an exopolysaccharide that binds chains of cells together in bundles. We report that transcription of eps is under the negative regulation of SinR, a repressor that was found to bind to multiple sites in the regulatory region of the operon. Mutations in sinR bypassed the requirement in biofilm formation of two genes of unknown function, ylbF and ymcA, and sinI, which is known to encode an antagonist of SinR. We propose that these genes are members of a pathway that is responsible for counteracting SinR-mediated repression. We further propose that SinR is a master regulator that governs the transition between a planktonic state in which the bacteria swim as single cells in liquid or swarm in small groups over surfaces, and a sessile state in which the bacteria adhere to each other to form bundled chains and assemble into multicellular communities. PMID- 15661001 TI - Mechanism of lipid-body formation in prokaryotes: how bacteria fatten up. AB - Neutral lipid accumulation is frequently observed in some Gram-negative prokaryotes like Acinetobacter sp. and most actinomycetes, including the pathogenic Mycobacterium tuberculosis and antibiotic producing streptomycetes. We examined the formation of wax ester- and triacylglycerol (TAG)-bodies in Acinetobacter calcoaceticus and Rhodococcus opacus using microscopic, immunological and biophysical methods. A general model for prokaryotic lipid-body formation is proposed, clearly differing from the current models for the formation of lipid inclusions in eukaryotes and of poly(hydroxyalkanoic acid) (PHA) inclusions in prokaryotes. Formation of lipid-bodies starts with the docking of wax ester synthase/acyl-CoA:diacylglycerol acyltransferase (WS/DGAT) to the cytoplasm membrane. Both, analyses of in vivo and in vitro lipid-body synthesis, demonstrated the formation of small lipid droplets (SLDs), which remain bound to the membrane-associated enzyme. SLDs conglomerated subsequently to membrane-bound lipid-prebodies which are then released into the cytoplasm. The formation of matured lipid-bodies in the cytoplasm occurred by means of coalescence of SLDs inside the lipid prebodies, which are surrounded by a half unit membrane of phospholipids. PMID- 15661002 TI - The tale of two RNA polymerases: transcription profiling and gene expression strategy of bacteriophage Xp10. AB - Bacteriophage Xp10 infects rice pathogen Xanthomonas oryzae. Xp10 encodes its own single-subunit RNA polymerase (RNAP), similar to that found in phages of the T7 family. On the other hand, most of Xp10 genes are organized in a manner typical of lambdoid phages that are known to rely only on host RNAP for their development. To better understand the temporal pattern of viral transcription during Xp10 development, we performed global transcription profiling, primer extension, chemical kinetic modelling and bioinformatic analyses of Xp10 gene expression. Our results indicate that true to its mosaic nature, Xp10 relies on both host and viral RNAPs for expression of genes coding for virion components and host lysis. The joint transcription of the same set of genes by two types of RNA polymerases is unprecedented for a bacteriophage. Curiously, such a situation is realized in chloroplasts. PMID- 15661003 TI - Why are pathogenic staphylococci so lysozyme resistant? The peptidoglycan O acetyltransferase OatA is the major determinant for lysozyme resistance of Staphylococcus aureus. AB - Staphylococcus species belong to one of the few bacterial genera that are completely lysozyme resistant, which greatly contributes to their persistence and success in colonizing the skin and mucosal areas of humans and animals. In an attempt to discover the cause of lysozyme resistance, we identified a gene, oatA, in Staphylococcus aureus. The corresponding oatA deletion mutant had an increased sensitivity to lysozyme. HPLC and electrospray ionization tandem mass spectrometry analyses of the cell wall revealed that the muramic acid of peptidoglycan of the wild-type strain was O-acetylated at C6-OH, whereas the muramic acid of the oatA mutant lacked this modification. The complemented oatA mutant was lysozyme resistant. We identified the first bacterial peptidoglycan specific O-acetyltransferase in S. aureus and showed that OatA, an integral membrane protein, is the molecular basis for the high lysozyme resistance in staphylococci. PMID- 15661004 TI - Demonstration of differential virulence gene promoter activation in vivo in Bordetella pertussis using RIVET. AB - Bordetella pertussis, the etiologic agent of whooping cough, causes disease by employing an array of virulence factors controlled by the BvgA-BvgS two-component signal transduction system. Regulation by this system has been extensively characterized in vitro, where bvg-activated genes are repressed in a process known as phenotypic modulation. Differential regulation of these genes by the response regulator BvgA results in promoters that are activated early, middle, or late after being released from modulation. However, the in vivo environmental signal and regulation pattern has not been described. In order to investigate BvgAS-mediated regulation of B. pertussis virulence factors in vivo using the mouse aerosol challenge model, we have adapted the recombinase-based in vivo technology (RIVET) system for use in B. pertussis. We have demonstrated that these strains show resolution during in vitro growth under non-modulating conditions. In addition, we have demonstrated that modulating strains by growth on media containing MgSO4 does not affect virulence in the mouse aerosol challenge model. We have therefore used the RIVET system to reveal the time course of gene expression in vivo for selected B. pertussis virulence factors (cya, fha, prn and ptx). Our data indicate that this method can be effectively used to monitor and compare in vivo and in vitro gene expression in B. pertussis, and that temporal regulation patterns previously observed in vitro are mirrored in vivo. PMID- 15661005 TI - Recruitment of penicillin-binding protein PBP2 to the division site of Staphylococcus aureus is dependent on its transpeptidation substrates. AB - Staphylococcus aureus penicillin-binding protein PBP2 is an enzyme involved in the last stages of peptidoglycan assembly and is an important player in the mechanism of methicillin resistance of this pathogen. PBP2 localized to the division site but its recruitment to the forming division septum was prevented after acylation by oxacillin. The presence of the antibiotic did not affect FtsZ ring maintenance nor the localization of externalized peptidoglycan precursors. Delocalization of PBP2 was also observed when its pentapeptide substrate was eliminated by addition of d-cycloserine or blocked by addition of vancomycin. Taken together these observations suggest that PBP2 is recruited to the division site by binding to its substrate, which is localized at that place. In methicillin-resistant S. aureus, addition of oxacillin does not result in delocalization of PBP2 indicating that acylated PBP2 can be maintained in place by functional PBP2A, the central element of this resistance mechanism. PMID- 15661006 TI - A truncated H-NS-like protein from enteropathogenic Escherichia coli acts as an H NS antagonist. AB - The H-NS nucleoid-associated protein of Escherichia coli is the founder member of a widespread family of gene regulatory proteins which have a bipartite structure, consisting of an N-terminal coiled-coil oligomerization domain and a C-terminal DNA-binding domain. Here we characterize a family of naturally occurring truncated H-NS derivatives lacking the DNA-binding domain, which we term the H NST family. H-NST proteins are found in large genomic islands in pathogenic E. coli strains, which are absent from the corresponding positions in the E. coli K 12 genome. Detailed analysis of the H-NST proteins from enteropathogenic E. coli (EPEC) and uropathogenic E. coli (UPEC) shows that the EPEC protein (H-NST(EPEC)) has a potent anti-H-NS function at the classical H-NS-repressed operon proU. This correlates with the ability of H-NST(EPEC) to co-purify with H-NS in vitro, and can be abolished by a mutation of leucine 30 to proline which is predicted to prevent the N-terminal region from forming a coiled-coil structure. In contrast, despite being 90% identical to H-NST(EPEC) at the protein level, the UPEC homologue (H-NST(UPEC)) has only a weak anti-H-NS activity, correlating with a much-reduced ability to interact with H-NS during column chromatography. A single amino acid difference at residue 16 appears to account for these different properties. The hnsT(EPEC) gene is transcribed monocistronically and expressed throughout the exponential growth phase in DMEM medium. Our data suggest that a truncated derivative of H-NS encoded by an ancestral mobile DNA element can interact with the endogenous H-NS regulatory network of a bacterial pathogen. PMID- 15661007 TI - The Neurospora crassa gene responsible for the cut and ovc phenotypes encodes a protein of the haloacid dehalogenase family. AB - Light stimulation of carotenogenesis in Neurospora crassa, mediated by the White Collar proteins, is enhanced in some regulatory mutants, such as vivid and ovc. The gene responsible for the vivid mutation has been identified, but not the one responsible for the ovc phenotype. The ovc mutant is sensitive to high osmotic conditions and allelic with another mutant, cut, also osmosensitive but not affected in carotenogenesis. A phenotypic characterization of both strains is presented. Light induction of mRNA levels of the carotenoid genes al-1 and al-2, the regulatory gene wc-1 or the conidiation-specific gene con-10 is not significantly changed in the ovc mutant when compared with the wild type. We have identified the gene affected in the ovc mutant by complementation of osmosensitivity with a cosmid library. This gene, which we call cut-1, codes for an enzyme of the haloacid dehalogenase family, which includes different classes of phosphatases. cut-1 is able to restore the wild-type phenotype of the ovc and cut strains, confirming that they are affected in the same gene. DNA sequence analysis identified a point mutation in the cut mutant, leading to a truncated protein. The ovc mutant represents a deletion encompassing the entire gene and surrounding sequences. The cut-1 promoter contains putative regulatory elements involved in osmotic or thermal stress. We show that cut-1 transcription is low in illuminated or dark-grown cultures, and is induced by high osmotic conditions or by heat shock. PMID- 15661008 TI - Degradation of the HilC and HilD regulator proteins by ATP-dependent Lon protease leads to downregulation of Salmonella pathogenicity island 1 gene expression. AB - Salmonella pathogenicity island 1 (SPI1) enables infecting Salmonella to cross the small intestinal barrier and to escape phagocytosis by inducing apoptosis. Several environmental signals and transcriptional regulators modulate the expression of hilA, which encodes a protein playing a central role in the regulatory hierarchy of SPI1 gene expression. We have previously shown that Lon, a stress-induced ATP-dependent protease, is a negative regulator of hilA, suggesting that it targets factors required for activating hilA expression. To elucidate the mechanisms by which Lon protease negatively regulates SPI1 transcription, we looked for its substrate proteins. We found that HilC and HilD, which are positive regulators of hilA expression, accumulate in Lon-depleted cells, and that the enhancement of SPI1 expression that occurs in a lon-disrupted mutant is not observed in the lon hilC hilD triple null mutant. Furthermore, we demonstrated that the half-lives of HilC and HilD are, respectively, about 12 times and three times longer in the Lon-depleted mutant, than in the Lon+ cells, suggesting that Lon targets both of HilC and HilD. In view of these findings, we suggest that the regulation of SPI1 expression is negatively controlled through degradation of the HilC and HilD transcriptional regulators by Lon. PMID- 15661009 TI - Contact with host cells induces a DNA repair system in pathogenic Neisseriae. AB - DNA repair systems play a major role in maintaining the integrity of bacterial genomes. Neisseria meningitidis, a human pathogen capable of colonizing the human nasopharynx, possesses numerous DNA repair genes but lacks inducible DNA repair systems such as the SOS response, present in most bacteria species. We recently identified a set of genes upregulated by contact with host cells. An open reading frame having high homology with the small subunit of Escherichia coli exonuclease VII (xseB) belongs to this regulon. The increased sensitivity of a mutant in this coding sequence to UV irradiation, alkylating agent and nalidixic acid demonstrates the participation of this gene in meningococcal DNA repair. In addition, the upregulation of the transcription of this open reading frame upon interaction of N. meningitidis with host cells increased not only the bacterial ability to repair its DNA but also the rate of phase variation by frameshifting. Together these data demonstrate that N. meningitidis possesses an inducible DNA repair system that might be used by the bacteria to adapt to its niches when it is colonizing a new host. PMID- 15661010 TI - PKA and Sch9 control a molecular switch important for the proper adaptation to nutrient availability. AB - In the yeast Saccharomyces cerevisiae, PKA and Sch9 exert similar physiological roles in response to nutrient availability. However, their functional redundancy complicates to distinguish properly the target genes for both kinases. In this article, we analysed different phenotypic read-outs. The data unequivocally showed that both kinases act through separate signalling cascades. In addition, genome-wide expression analysis under conditions and with strains in which either PKA and/or Sch9 signalling was specifically affected, demonstrated that both kinases synergistically or oppositely regulate given gene targets. Unlike PKA, which negatively regulates stress-responsive element (STRE)- and post-diauxic shift (PDS)-driven gene expression, Sch9 appears to exert additional positive control on the Rim15-effector Gis1 to regulate PDS-driven gene expression. The data presented are consistent with a cyclic AMP (cAMP)-gating phenomenon recognized in higher eukaryotes consisting of a main gatekeeper, the protein kinase PKA, switching on or off the activities and signals transmitted through primary pathways such as, in case of yeast, the Sch9-controlled signalling route. This mechanism allows fine-tuning various nutritional responses in yeast cells, allowing them to adapt metabolism and growth appropriately. PMID- 15661011 TI - Biogenesis of a putative channel protein, ComEC, required for DNA uptake: membrane topology, oligomerization and formation of disulphide bonds. AB - ComEC is a putative channel protein for DNA uptake in Bacillus subtilis and other genetically transformable bacteria. Membrane topology studies suggest a model of ComEC as a multispanning membrane protein with seven transmembrane segments (TMSs), and possibly with one laterally inserted amphipathic helix. We show that ComEC contains an intramolecular disulphide bond in its N-terminal extracellular loop (between the residues C131 and C172), which is required for the stability of the protein, and is probably introduced by BdbDC, a pair of competence-induced oxidoreductase proteins. By in vitro cross-linking using native cysteine residues we show that ComEC forms an oligomer. The oligomerization surface includes a transmembrane segment, TMS-G, near the cytoplasmic C-terminus of ComEC. PMID- 15661012 TI - The sigma54-dependent transcriptional activator SfnR regulates the expression of the Pseudomonas putida sfnFG operon responsible for dimethyl sulphone utilization. AB - Pseudomonas putida DS1 is able to utilize dimethyl sulphide through dimethyl sulphoxide, dimethyl sulphone (DMSO2), methanesulphonate (MSA) and sulphite as a sulphur source. We previously demonstrated that sfnR encoding a sigma54-dependent transcriptional regulator is essential for DMSO2 utilization by P. putida DS1. To identify the target genes of SfnR, we carried out transposon mutagenesis on an sfnR disruptant (DMSO2-utilization-defective phenotype) using mini-Tn5, which contains two outward-facing constitutively active promoters; as a result, we obtained a mutant that restored the ability to utilize DMSO2. The DMSO2-positive mutant carried a mini-Tn5 insertion in the intergenic region between two opposite facing operons, sfnAB and sfnFG. Both sfnA and sfnB products were similar to acyl CoA dehydrogenase family proteins, whereas sfnF and sfnG encoded a putative NADH dependent FMN reductase (SfnF) and an FMNH2-dependent monooxygenase (SfnG). Disruption and complementation of the sfn genes indicated that the sfnG product is essential for DMSO2 utilization by P. putida DS1. Furthermore, an enzyme assay demonstrated that SfnG is an FMNH2-dependent DMSO2 monooxygenase that converts DMSO2 to MSA. It was revealed that the expression of the sfnFG operon is directly activated by the binding of SfnR at its upstream region. Site-directed mutagenesis of the SfnR binding sequences allowed us to define a potential recognition sequence for SfnR. These results provided insight into regulation of sulphate starvation-induced genes in bacteria. PMID- 15661013 TI - The Legionella IcmS-IcmW protein complex is important for Dot/Icm-mediated protein translocation. AB - The intracellular pathogen Legionella pneumophila can infect and replicate within macrophages of a human host. To establish infection, Legionella require the Dot/Icm secretion system to inject protein substrates directly into the host cell cytoplasm. The mechanism by which substrate proteins are engaged and translocated by the Dot/Icm system is not well understood. Here we show that two cytosolic components of the Dot/Icm secretion machinery, the proteins IcmS and IcmW, play an important role in substrate translocation. Biochemical analysis indicates that IcmS and IcmW form a stable protein complex. In Legionella, the IcmW protein is rapidly degraded in the absence of the IcmS protein. Substrate proteins translocated into mammalian host cells by the Dot/Icm system were identified using the IcmW protein as bait in a yeast two-hybrid screen. It was determined that the IcmS-IcmW complex interacts with these substrates and plays an important role in translocation of these proteins into mammalian cells. These data are consistent with the IcmS-IcmW complex being involved in the recognition and Dot/Icm-dependent translocation of substrate proteins during Legionella infection of host cells. PMID- 15661014 TI - The svpA-srtB locus of Listeria monocytogenes: fur-mediated iron regulation and effect on virulence. AB - In Listeria monocytogenes the promoter region of the svpA-srtB locus contains a well-conserved Fur box. We characterized the iron-regulation of this locus: real time polymerase chain reaction analyses and anti-SvpA immunoblots showed that, in response to iron deprivation svpA transcription and SvpA production markedly increased (80-fold and 10-fold respectively), when initiated by either the addition of the iron chelator 2,2'-bipyridyl to BHI media, or by growth in iron restricted minimal media. Green fluorescent protein (GFP) reporter constructs also showed increased activity of the svpA-srtB promoter in Escherichia coli (37 fold) and in L. monocytogenes (two- to threefold) when the bacteria were grown in iron-deficient conditions. A Deltafur mutant of L. monocytogenes constitutively synthesized SvpA, as well as GFP fused to the svpA-srtB promoter. Cellular fractionation data revealed that in iron-rich media wild-type SvpA was exclusively secreted to the culture supernatant. However, both the Deltafur derivative and wild-type L. monocytogenes grown in iron-deficient media anchored a fraction of the SvpA proteins (approximately 5%) to peptidoglycan, and produced a lower-molecular weight, wholly secreted form of SvpA. Together these data establish that iron availability controls transcription of the svpA-srtB locus (through Fur-mediated regulation), and attachment of SvpA to the cell wall (through SrtB-mediated covalent linkage). SvpA bears homology to IsdC, a haemin binding protein of Staphylococcus aureus, and haemin bound to SvpA in solution. However, site-directed deletions of four structural genes and the promoter of the svpA-srtB locus did not impair haemin, haemoglobin or ferrichrome utilization in nutrition tests. We did not find strong evidence to support the notion that the svpA-srtB locus participates in haemin acquisition, as was reported for the homologous isd operon of S. aureus. Furthermore, the svpA-srtB mutant strains showed no significant attenuation of virulence in an intravenous mouse model system, but we found that the mutations reduced the persistence of L. monocytogenes in murine liver, spleen and intestines after oral administration. PMID- 15661015 TI - Type III secretion chaperones of Pseudomonas syringae protect effectors from Lon associated degradation. AB - The hrp type III secretion system (TTSS) of Pseudomonas syringae translocates effector proteins into the cytoplasm of host cells. Proteolysis of HrpR by Lon has been shown to negatively regulate the hrp TTSS. The inability to bypass Lon associated effects on the regulatory system by ectopic expression of the known regulators suggested a second site of action for Lon in TTSS-dependent effector secretion. In this study we report that TTSS-dependent effectors are subject to the proteolytic degradation that appears to be rate-limiting to secretion. The half-lives of the effectors AvrPto, AvrRpt2, HopPsyA, HopPsyB1, HopPtoB2, HopPsyV1, HopPtoG and HopPtoM were substantially higher in bacteria lacking Lon. TTSS-dependent secretion of several effectors was enhanced from Lon mutants. A primary role for chaperones appears to be protection of effectors from Lon associated degradation prior to secretion. When coexpressed with their cognate chaperone, HopPsyB1, HopPsyV1 and HopPtoM were at least 10 times more stable in strains expressing Lon. Distinct Lon-targeting and chaperone-binding domains were identified in HopPtoM. The results imply that Lon is involved at two distinct levels in the regulation of the P. syringae TTSS: regulation of assembly of the secreton and modulation of effector secretion. PMID- 15661016 TI - Mig-14 is an inner membrane-associated protein that promotes Salmonella typhimurium resistance to CRAMP, survival within activated macrophages and persistent infection. AB - Salmonella enterica serovar Typhimurium (S. typhimurium) infects a wide variety of mammalian hosts and in rodents causes a typhoid-like systemic disease involving replication of bacteria inside macrophages within reticuloendothelial tissues. Previous studies demonstrated that the mig-14 and virK genes of Salmonella enterica are important in bacterial resistance to anti-microbial peptides and are necessary for continued replication of S. typhimurium in the liver and spleen of susceptible mice after orogastric inoculation. In this work we report that inflammatory signalling via interferon-gamma (IFN-gamma) is crucial to controlling replication of mig-14 mutant bacteria within the liver and spleen of mice after oral infection. Using a Salmonella persistence model recently developed in our laboratory, we further demonstrate that mig-14 contributes to long-term persistence of Salmonella in the spleen and mesenteric lymph nodes of chronically infected mice. Both mig-14 and virK contribute to the survival of Salmonella in macrophages treated with IFN-gamma and are necessary for resistance to cathelin-related anti-microbial peptide (CRAMP), an anti microbial peptide expressed at high levels in activated mouse macrophages. We also show that both Mig-14 and VirK inhibit the binding of CRAMP to Salmonella, and demonstrate that Mig-14 is an inner membrane-associated protein. We further demonstrate by transmission electron microscopy that the primary locus of CRAMP activity appears to be intracytoplasmic, rather than at the outer membrane, suggesting that Mig-14 may prevent the penetration of the inner membrane by CRAMP. Together, these data indicate an important role for mig-14 in anti microbial peptide resistance in vivo, and show that this resistance is important to the survival of Salmonella in systemic sites during both acute and persistent infection. PMID- 15661019 TI - Human disease resulting from gene mutations that interfere with appropriate nuclear factor-kappaB activation. AB - The nuclear factor (NF)-kappaB family of transcription factors serves vital roles in a wide array of cell functions. An increasing number of human genetic lesions that result in defined disease entities are linked to inappropriate activation of NF-kappaB. The resulting aberrant NF-kappaB function can lead to cellular defects that ultimately impair normal developmental processes, host immune defenses, or both. Molecular defects that lie upstream in cell-signaling pathways and rely upon NF-kappaB activation tend to give a more specific phenotype, whereas those closer to the actual NF-kappaB proteins have broader defects. A detailed study of these diseases can provide insight into the biochemistry of NF-kappaB activation as well as the role of NF-kappaB in human health. PMID- 15661018 TI - Inherited disorders of human Toll-like receptor signaling: immunological implications. AB - In vitro nine of 10 known human Toll-like receptors (TLRs) are engaged by well defined chemical agonists that mimic microbial compounds, raising the possibility that human TLRs play a critical role in protective immunity in vivo. We thus review here the recently described human primary immunodeficiencies caused by germline mutations in genes encoding molecules involved in cell signaling downstream from TLRs. Subjects with anhidrotic ectodermal dysplasia with immunodeficiency (EDA-ID) carry either X-linked recessive hypomorphic mutations in NEMO or autosomal dominant hypermorphic mutations in IKBA. Their cells show a broad defect in nuclear factor-kappaB (NF-kappaB) activation, with an impaired, but not abolished response to a large variety of stimuli including TLR agonists. EDA-ID patients show developmental anomalies of skin appendages and a broad spectrum of infectious diseases. Patients with autosomal recessive amorphic mutations in IRAK4 present a purely immunological syndrome and more restricted defects, with specific impairment of the Toll and interleukin-1 receptor (TIR) interleukin-1 receptor-associated kinase (IRAK) signaling pathway. In these subjects, the NF-kappaB- and mitogen-activated protein kinase-mediated induction of inflammatory cytokines in response to TIR agonists is impaired. The patients present a narrow range of pyogenic bacterial infections that become increasingly rare with age. Altogether, these data suggest that human TLRs play a critical role in host defense. However, they do not provide compelling evidence, as even the infectious phenotype of patients with mutations in IRAK4 may result from impaired signaling via receptors other than TLRs. Paradoxically, these experiments of nature raise the possibility that the entire set of human TLRs is largely redundant in protective immunity in vivo. PMID- 15661020 TI - Defects in the interferon-gamma and interleukin-12 pathways. AB - The interferon-gamma (IFN-gamma)/interleukin-12 (IL-12) pathway is a pivotal player in the immune system and is central to controlling mycobacterial infections. We highlight the most recent and relevant advances in understanding this pathway and their repercussions on basic and clinical science. Human mutations in IFN-gamma receptor-1 (IFN-gammaR1), IFN-gammaR2, IL-12p40, IL-12 receptor-beta1, signal transducer and activator of transcription-1, and nuclear factor-kappaB essential modulator are analyzed in the context of genetic susceptibility to mycobacterial diseases. A diagnostic and therapeutic approach is described. The IFN-gamma/IL-12 pathway is central in immune control of both environmental and autochthonous challenges, as reflected in human mutations and animal models. Besides being crucial for mycobacterial control, the IFN-gamma/IL 12 pathway is also involved in the pathogenesis of autoimmune disease as well as tumor development and control. Genotype-phenotype correlations have been established for certain genes in this pathway, some of which have therapeutic implications. PMID- 15661021 TI - Hyper immunoglobulin M syndrome due to CD40 deficiency: clinical, molecular, and immunological features. AB - CD40 is a member of the tumor necrosis factor receptor family, which is expressed by a variety of cells including B cells, macrophages, dendritic cells, and other nonimmune cell types. CD40 activation is critical for B-cell proliferation, immunoglobulin (Ig)-isotype switching, and germinal center formation. In physiological conditions, the activation of CD40 occurs by binding to its natural ligand, CD154, which is expressed on activated T cells. The in vivo critical role of CD40-CD154 interaction on B-cell differentiation and isotype switching is provided by the discovery that mutations in either CD40 or CD154 gene cause the hyper IgM syndrome, termed HIGM3 or HIGM1, respectively, characterized by very low levels of serum IgG, IgA, and IgE, with normal or elevated IgM, associated with a defective germinal center formation. Originally considered humoral primary immunodeficiencies, the clinical features and the defect of T-cell priming, resulting from a defective T-B cell or dendritic cell interaction, is now considered as combined immunodeficiencies. In this article, we present a comprehensive overview of the clinical, genetic, and immunological features of patients with hyper IgM syndrome due to CD40 mutations. PMID- 15661022 TI - Hyper-immunoglobulin M syndromes caused by intrinsic B-lymphocyte defects. AB - Hyper-immunoglobulin M (IgM) syndromes are primary immunodeficiencies characterized by normal or elevated serum IgM levels with the absence of other isotypes, pinpointing to a defect in the Ig class switch recombination (CSR). The delineation of hyper-IgM syndromes made it possible to better define the mechanisms underlying the two major events of antibody maturation in humans, CSR and introduction of somatic hypermutation (SHM) in the variable region of immunoglobulins. The description of the activation-induced cytidine deaminase (AID) deficiency, characterized by a defect in both CSR and SHM, demonstrated for the first time that this molecule acts as a master player in the antigen-induced Ig gene-modification events responsible for both CSR and SHM. However, deleterious mutations located in the C-terminus lead to a CSR defect without affecting SHM, providing evidence for a role of AID in CSR distinct from the cytidine deaminase activity, likely by binding to a specific CSR cofactor. Molecular causes of two other hyper-IgM conditions have not yet been defined. However, they may be caused by either a defect in AID targeting on S regions or a CSR-specific DNA-repair defect. The mechanism of action of AID remains somewhat debated, but the observation that uracil-DNA-glycosylase deficiency leads to a severe hyper-IgM syndrome strongly argues in favor of a DNA-editing activity of AID. PMID- 15661023 TI - DNA deamination in immunity. AB - A functional immune system is one of the prerequisites for the survival of a species. Humans have one of the most complicated immune systems, with the ability to learn from and adapt to pathogens. At first, a primary repertoire of antibodies is generated, which, upon antigen encounter, will diversify and adapt to produce a highly specific and potent secondary response, part of which is kept in memory to fight off future infections. In this review, the mechanism as well as the specificities of the key protein in the secondary immune response, activation-induced cytidine deaminase (AID), are highlighted, as well as its role in the DNA deamination model of immunoglobulin diversification. The review also highlights aspects of AID's regulation on both the transcriptional as well as post-translational level and its potential molecular mechanism and specificity. Furthermore, it expands outside the involvement of AID in somatic hypermutation, class switching, and gene conversion to discuss the implications of DNA deamination in epigenetic modifications of DNA (as a potential demethylase), the induction of mutations during oncogenesis, and includes an evolutionary comparison to the DNA deaminase family member APOBEC3G, a key protein in human immunodeficiency virus pathogenesis. PMID- 15661024 TI - Severe combined immunodeficiency. A model disease for molecular immunology and therapy. AB - Severe combined immunodeficiencies (SCIDs) consist of genetically determined arrest of T-cell differentiation. Ten different molecular defects have now been identified, which all lead to early death in the absence of therapy. Transplantation of allogeneic hematopoietic stem cells (HSCT) can restore T-cell development, thus saving the lives of SCID patients. In this review, the different characteristics of HSCT are discussed along with the available data regarding the long-term outcome. Transient thymopoiesis caused by an exhaustion of donor progenitor cells and possibly a progressive loss of thymus function can lead to a progressive decline in T-cell functions. The preliminary results of gene therapy show the correction of two SCID conditions. Based on the assumption that long-lasting pluripotent progenitor cells are transduced, these data suggest that gene therapy could overcome the long-term recurrence of the T-cell immunodeficiency. SCID is thus a disease model for experimental therapy in the hematopoietic system. PMID- 15661025 TI - Interleukin-7 receptor alpha (IL-7Ralpha) deficiency: cellular and molecular bases. Analysis of clinical, immunological, and molecular features in 16 novel patients. AB - Analysis of gene-targeted mice and patients with severe combined immunodeficiency due to mutations of the alpha chain of the interleukin-7 receptor (IL-7Ralpha) has shown important differences between mice and humans in the role played by IL 7 in lymphoid development. More recently, it has been shown that IL-7Ralpha is also shared by the receptor for another cytokine, thymic stromal lymphopoietin (TSLP). In this review, we discuss recent advances in IL-7- and TSLP-mediated signaling. We also report on the clinical and immunological features of 16 novel patients with IL-7Ralpha deficiency and discuss the results of hematopoietic stem cell transplantation. PMID- 15661026 TI - Jak3, severe combined immunodeficiency, and a new class of immunosuppressive drugs. AB - The recent elucidation of the multiple molecular mechanisms underlying severe combined immunodeficiency (SCID) is an impressive example of the power of molecular medicine. Analysis of patients and the concomitant generation of animal models mimicking these disorders have quickly provided great insights into the pathophysiology of these potentially devastating illnesses. In this review, we summarize the discoveries that led to the understanding of the role of cytokine receptors and a specific tyrosine kinase, Janus kinase 3 (Jak3), in the pathogenesis of SCID. We discuss how the identification of mutations of Jak3 in autosomal recessive SCID has facilitated the diagnosis of these disorders, offered new insights into the biology of this kinase, permitted new avenues for therapy, and provided the rationale for a generation of a new class of immunosuppressants. PMID- 15661027 TI - Studies of patients' thymi aid in the discovery and characterization of immunodeficiency in humans. AB - Studying the molecular and genetic bases of primary immunodeficiency is valuable at several levels. First, such information directly benefits patients in both short- and long-term management. Sophisticated diagnostic tools based on these studies can be used early and lead to appropriate treatment before potentially fatal infections and complications arise. Genotyping is also critical for future development and implementation of gene therapy. Secondly, investigating primary immunodeficiency helps understand the normal immune system in humans. As described in this report, the roles of zeta-associated protein of 70 kDa (Zap 70), CD25, and CD3delta are substantially different in humans when compared with the roles of homologous molecules in other species. Last, information obtained from these studies can be applied to other fields of investigation. Prominent examples for such applications include the intensive effort to design and produce specific inhibitors of Zap-70 and Janus kinase 3 as specific immunosuppressive agents. Most types of primary immunodeficiency in general and severe combined immunodeficiency in particular are rare and therefore cannot be easily studied by using traditional genetic methodology. Instead, biochemical methods were used to explore for candidate genes as was the case in the discovery of Zap-70 deficiency. Critical to the success of these discoveries was the careful analysis of patients' thymus glands. Detection of abnormalities in the thymus in these patients, which preceded identification of the genetic defect, aided in the assessment of the severity and nature of the immune disorder (primary versus secondary). Such assessment is critical before high-risk bone marrow transplantation. Equally important was the contribution of studies of the thymus to the description of novel phenotype of immunodeficiency as clearly demonstrated in defining CD8 lymphocytopenia, Zap-70 deficiency, and CD25 deficiency. Indeed, analysis of the thymus directly pointed to CD25 as candidate gene. Recently, the study of thymocyte-derived transcripts using DNA microarrays was key to discovering CD3delta deficiency. Finally, immunohistochemical analysis of the thymus was critical in pinpointing the roles of Zap-70, CD25, and CD3delta in the development of human T cells. PMID- 15661028 TI - FOXP3 acts as a rheostat of the immune response. AB - The study of a rare human X-linked disease resulting in a characteristic clinical phenotype of multiple autoimmune disorders and the in-depth exploration of a spontaneous mouse model, scurfy (sf), have contributed to a better understanding of the regulation of immunologic responses, particularly to self. Forkhead box P3 (FOXP3), the gene responsible for IPEX (immune dysregulation, polyendocrinopathy, enteropathy, X-linked) and sf is located on the X chromosome and is of crucial importance for the generation of CD4+ CD25+ regulatory T cells. Loss of FOXP3 function and the resultant lack of regulatory T cells result in lethal auto aggressive lymphoproliferation, whereas overexpression of this modulator results in severe immunodeficiency. The in-depth analysis of FOXP3 regulation and elucidation of the precise mechanisms by which FOXP3 exerts its regulatory effect will provide important insights into the understanding of autoimmunity and should predictably result in new therapeutic possibilities. PMID- 15661029 TI - Primary hemophagocytic syndromes point to a direct link between lymphocyte cytotoxicity and homeostasis. AB - Hemophagocytic syndrome (HS) is a severe and often fatal syndrome resulting from potent and uncontrolled activation and proliferation of T-lymphocytes, leading to excessive macrophage activation and multiple deleterious effects. The onset of HS characterizes several inherited disorders in humans. In each condition, the molecular defect impairs the granule-dependent cytotoxic activity of lymphocytes, thus highlighting the determinant role of this function in driving the immune system to a state of equilibrium following infection. It has also been shown that some of the proteins required for lytic granule secretion are required for melanocyte function, leading to associated hypopigmentation in these conditions. This review focuses on several effectors of this secretory pathway, recently identified, because their defects cause these disorders, and discusses their role and molecular interactions in granule-dependent cytotoxic activity. PMID- 15661030 TI - Molecular and cellular pathogenesis of X-linked lymphoproliferative disease. AB - X-linked lymphoproliferative disease (XLP) is an inherited immune defect caused by mutations in the Src homology 2 domain-containing gene 1A, which encodes the adapter protein, signaling lymphocytic activation molecule (SLAM)-associated protein (SAP). SAP is expressed in T cells, natural killer (NK) cells, and NKT cells, where it binds to the cytoplasmic domain of the surface receptor SLAM (CD150) and the related receptors, 2B4 (CD244), CD84, Ly9 (CD229), NK-T-B antigen, and CD2-like receptor-activating cytotoxic T cells. SAP also binds to the Src family tyrosine kinase Fyn and recruits it to SLAM, which leads to the generation of downstream phosphotyrosine signals. While the roles of the SLAM family receptors are only beginning to be understood, experiments suggest that these molecules regulate important aspects of lymphocyte function, such as proliferation, cytokine secretion, cytotoxicity, and antibody production. Thus, in XLP patients who lack functional SAP, the SLAM family receptors may not signal properly. This property likely contributes to the phenotypes of XLP, including fulminant infectious mononucleosis, lymphoma, and hypogammaglobulinemia. Further studies of SAP and the SLAM family receptors will provide insights into XLP and elucidate the signaling events regulating lymphocyte ontogeny and function. PMID- 15661032 TI - Genetic analysis of patients with defects in early B-cell development. AB - Approximately 85% of patients with defects in early B-cell development have X linked agammaglobulinemia (XLA), a disorder caused by mutations in the cytoplasmic Bruton's tyrosine kinase (Btk). Although Btk is activated by cross linking of a variety of cell-surface receptors, the most critical signal transduction pathway is the one initiated by the pre-B cell and B-cell antigen receptor complex. Mutations in Btk are highly diverse, and no single mutation accounts for more than 3% of patients. Although there is no strong genotype/phenotype correlation in XLA, the specific mutation in Btk is one of the factors that influences the severity of disease. Mutations in the components of the pre-B cell and B-cell antigen receptor complex account for an additional 5-7% of patients with defects in early B-cell development. Patients with defects in these proteins are clinically indistinguishable from those with XLA. However, they tend to be younger at the time of diagnosis, and whereas most patients with XLA have a small number of B cells in the peripheral circulation, these cells are not found in patients with defects in micro heavy chain or Igalpha. Polymorphic variants in the components of the pre-B cell and B-cell receptor complex, particularly micro heavy chain and lambda5, may contribute to the severity of XLA. PMID- 15661031 TI - Bruton's tyrosine kinase: cell biology, sequence conservation, mutation spectrum, siRNA modifications, and expression profiling. AB - Bruton's tyrosine kinase (Btk) is encoded by the gene that when mutated causes the primary immunodeficiency disease X-linked agammaglobulinemia (XLA) in humans and X-linked immunodeficiency (Xid) in mice. Btk is a member of the Tec family of protein tyrosine kinases (PTKs) and plays a vital, but diverse, modulatory role in many cellular processes. Mutations affecting Btk block B-lymphocyte development. Btk is conserved among species, and in this review, we present the sequence of the full-length rat Btk and find it to be analogous to the mouse Btk sequence. We have also analyzed the wealth of information compiled in the mutation database for XLA (BTKbase), representing 554 unique molecular events in 823 families and demonstrate that only selected amino acids are sensitive to replacement (P < 0.001). Although genotype-phenotype correlations have not been established in XLA, based on these findings, we hypothesize that this relationship indeed exists. Using short interfering-RNA technology, we have previously generated active constructs downregulating Btk expression. However, application of recently established guidelines to enhance or decrease the activity was not successful, demonstrating the importance of the primary sequence. We also review the outcome of expression profiling, comparing B lymphocytes from XLA-, Xid-, and Btk-knockout (KO) donors to healthy controls. Finally, in spite of a few genes differing in expression between Xid- and Btk-KO mice, in vivo competition between cells expressing either mutation shows that there is no selective survival advantage of cells carrying one genetic defect over the other. We conclusively demonstrate that for the R28C-missense mutant (Xid), there is no biologically relevant residual activity or any dominant negative effect versus other proteins. PMID- 15661033 TI - CXCR4 mutations in WHIM syndrome: a misguided immune system? AB - Chemokines and their receptors are key molecules in the development and function of immune cell populations and the organization of lymphoid organs. Despite their central role in immunologic function, genetic studies exploring the intersection of chemokines or their receptors and human health have revealed few associations of unambiguous significance. The best-characterized examples have revealed striking selective advantage conferred by loss of receptors used as portals of entry by pathogens. Recently, mutations in the CXCR4 chemokine receptor gene were identified in a dominantly inherited immunodeficiency disease, WHIM syndrome. Genetic and biochemical evidences suggest that the loss of the receptor cytoplasmic tail domain results in aberrant signaling. Analyses of mutant cell responses to the receptor ligand CXCL12 have revealed enhanced chemotaxis, confirming the gain-of-function effect of the truncation mutations. The clinical features and potential mechanism of immunodeficiency in WHIM syndrome patients are discussed in this review. PMID- 15661034 TI - Hyper-IgE syndromes. AB - The hyper-immunoglobulin E (IgE) syndromes (HIES) are primary immunodeficiencies characterized by the clinical triad of recurrent staphylococcal abscesses, recurrent cyst-forming pneumonia, and an elevated serum IgE level of >2000 IU/ml. Most cases are sporadic; however, multiplex families displaying autosomal dominant (AD) and autosomal recessive (AR) inheritance have been described. In most sporadic and AD cases, the HIES clinical triad is part of a multisystem disorder including abnormalities of the soft tissue, skeletal, and dental systems. In contrast, those with AR-HIES have severe molluscum contagiosum and other viral infections and may develop severe neurological complications. Unlike patients with sporadic HIES and AD-HIES, those with AR-HIES lack skeletal or dental involvement and do not develop lung cysts. Additional variants of HIES are discussed in this review. The etiology of HIES is still unresolved. Recent research points toward a skewed T helper 1 (Th1) cell/Th2 cell ratio and the involvement of chemokines. Therapy for HIES is directed at prevention and management of infections by using sustained systemic antibiotics and antifungals along with topical therapy for eczema and drainage of abscesses. Anti staphylococcal antibiotic prophylaxis is useful. Interferons, immunoglobulin supplementation, or low-dose cyclosporine A have been reported to benefit selected patients, but they are not generally indicated. PMID- 15661035 TI - Structural and functional aspects of the Ly49 natural killer cell receptors. AB - Natural killer cells are part of the first line of innate immune defence against virus-infected cells and cancer cells in the vertebrate immune system. They are called 'natural' killers because, unlike cytotoxic T cells, they do not require a previous challenge and preactivation to become active. The Ly49 NK receptors are type II transmembrane glycoproteins, structurally characterized as disulphide linked homodimers. They share extensive homology with C-type lectins, and they are encoded by a multigene family that in mice maps on chromosome 6. A fine balance between inhibitory and activating signals regulates the function of NK cells. Inhibitory Ly49 molecules bind primarily MHC class I ligands, whereas the ligands for activating Ly49 molecules may include MHC class I, but also interestingly MHC class I-like molecules expressed by viruses, as is the case for Ly49H, which binds the m157 gene product of murine cytomegalovirus. In this study, we review the function and X-ray crystal structure of the Ly49 NK cell receptors hitherto determined (Ly49A, Ly49C and Ly49I), and the structural features of the Ly49/MHC class I interaction as revealed by the X-ray crystal structures of Ly49A/H-2Dd and the recently determined Ly49C/H-2Kb. PMID- 15661036 TI - Immune dysregulation and self-reactivity in schizophrenia: do some cases of schizophrenia have an autoimmune basis? AB - Schizophrenia affects 1% of the world's population, but its cause remains obscure. Numerous theories have been proposed regarding the cause of schizophrenia, ranging from developmental or neurodegenerative processes or neurotransmitter abnormalities to infectious or autoimmune processes. In this review, findings suggestive of immune dysregulation and reactivity to self in patients with schizophrenia are examined with reference to criteria for defining whether or not a human disease is autoimmune in origin. Associations with other autoimmune diseases and particular MHC haplotypes, increased serum levels of autoantibodies, and in vivo and in vitro replication of some of the functional and ultrastructural abnormalities of schizophrenia by transfer of autoantibodies from the sera of patients with schizophrenia suggest that, in some patients at least, autoimmune mechanisms could play a role in the development of disease. Recent findings regarding specific autoimmune responses directed against neurotransmitter receptors in the brain in patients with schizophrenia will also be reviewed. PMID- 15661037 TI - IFN-gamma and TNF-alpha are involved in urushiol-induced contact hypersensitivity in mice. AB - Contact hypersensitivity (CHS) is a cutaneous T-cell-mediated immunological reaction to applied haptens. Activated antigen-specific T cells release several cytokines and chemokines followed by the recruitment of inflammatory cells and skin damage. CD8+ T cells and CD4+ T cells have been involved in the establishment of previously described CHS. In this study, we investigated the induction of CHS by urushiol in mice. Maximum swelling in mouse ears was elicited 24 h after challenge with urushiol on day 9 of sensitization. IFN-gamma, TNF alpha and IFN-gamma-inducible protein 10 (IP-10) mRNA were expressed after challenge of the antigen in urushiol-sensitized mice, but not in unsensitized mice. IFN-gamma knockout (KO) mice and TNF-alpha KO mice failed to elicit CHS with urushiol. Contact hypersensitivity and expressions of IFN-gamma, TNF-alpha and IP-10 mRNA were markedly suppressed in CD4+ and CD8+ cell-depleted mice. These results suggest that IFN-gamma, TNF-alpha, and possibly IP-10, play a critical role in CHS induced by urushiol, depending on both CD4+ T cells and CD8+ T cells. PMID- 15661038 TI - RGDS peptide inhibits activation of lymphocytes and adhesion of activated lymphocytes to human umbilical vein endothelial cells in vitro. AB - The Arg-Gly-Asp (RGD) motif is known to mediate cell adhesion to several extracellular matrix components as well as cell-cell interactions. In the present study, we investigated whether the RGDS peptide interferes with cell-cell recognition-based events such as allogeneic activation of PBMC and PBMC adhesion to human umbilical vein endothelial cells (HUVEC). We show here for the first time, to our knowledge, that RGDS significantly inhibits adhesion of activated PBMC to HUVEC; in addition, RGDS inhibits PBMC allogenenic activation in human mixed lymphocyte reaction assays. Caspases played a pivotal role in both events, because preventing their activation abolished or strongly reduced the observed inhibitory effect. The RGDS antirecognition effect was strongly increased by pretreatment of HUVEC with RGDS, which affected mostly T lymphocyte adhesion to HUVEC. These results indicate that PBMC allogeneic activation, as well as reciprocal recognition between activated PBMC and endothelial cells, are RGDS dependent events that occur through a dual effect involving anti-adhesive and caspase-dependent mechanisms. These data suggest a potential role of RGDS in cell mediated immunity, inflammation and organ transplantation. PMID- 15661039 TI - SAP and SLAM expression in anti-CD3 activated lymphocytes correlates with cytotoxic activity. AB - Signalling lymphocyte activation molecule (SLAM)-associated protein (SAP) is a small protein that is mutant in humans with X-linked lymphoproliferative (XLP) disease. Patients with XLP disease are affected by fatal EBV infection and malignant B-cell lymphomas. The increased risk for B-cell lymphomas is suggested to result from impaired immunosurveillance of B-cell proliferation by T cells. In this study, we investigated the role of SLAM and SAP in activation of effector cells with cytotoxic activity, cytokine-induced killer (CIK) cells, which are generated by non-specific stimulation of the TCR and addition of exogenous IL-2. Agonistic TCR activation 1 day after preparation (day +1) resulted in cell activation, with a peak of SLAM on day +6 visible at both the protein and mRNA level as well as membrane detectable SLAM. This increase in SLAM expression correlated significantly with SAP expression at the mRNA level as well as at the protein level. Cytotoxic activity peaked 1 day after the observed SAP and SLAM peaks. At that point in time, IL-10 secretion, which was high during the early days of culture, decreased. In conclusion, activation of peripheral blood cells with agonistic anti-CD3 antibody and exogenous IL-2, as used for generation of CIK cells, results in significant SLAM and SAP activation 5 days after TCR stimulation. This peak correlates with cytotoxic activity against tumour cells. Expression of SLAM and SAP seems to be important in the activation of cytotoxic effector cells. PMID- 15661040 TI - Secretory products from infective forms of Nippostrongylus brasiliensis induce a rapid allergic airway inflammatory response. AB - Allergic asthma is responsible for widespread morbidity and mortality and its incidence has increased dramatically in industrialized countries over the past two decades. Here, we describe a new murine model of allergic asthma utilizing a novel allergen with intrinsic enzymatic activity similar to that reported for many environmental allergens. The allergen, NES, is excreted and secreted from the nematode Nippostrongylus brasiliensis, and can readily be isolated from in vitro parasite cultures. When NES is administered intranasally to presensitized mice, allergic airway disease develops, including airway hyper-responsiveness, airway eosinophilia, IgE antibody production and Th2 cytokine production. This disease is characteristic of atopic asthma and can be induced within 11 days, thus providing a platform for the rapid analysis of allergic disease and high throughput testing of immunomodulatory factors. PMID- 15661041 TI - Imatinib inhibits the functional capacity of cultured human monocytes. AB - Imatinib is a tyrosine kinase inhibitor that has been reported to specifically inhibit the growth of bcr-abl expressing chronic myeloid leukaemia progenitors. This drug functions by blocking the ATP-binding site of the kinase domain of bcr abl, and has also been found to inhibit the c-abl, platelet-derived growth factor receptor, ARG and stem cell factor receptor tyrosine kinases. Reports have recently emerged demonstrating that imatinib also inhibits the growth of non malignant haemopoietic cells. Here, we demonstrate that concentrations of imatinib within the therapeutic dose range inhibit the function of cultured monocytes (CM) from normal donors. A decrease in the response of CM to LPS was observed morphologically and functionally, with CM grown in the presence of imatinib showing decreased pseudopodia formation and inhibition of IL-6 and TNF alpha production following LPS stimulation. Imatinib also reduced the ability of M-CSF and GM-CSF stimulated CM to phagocytose zymosan particles, with uptake of non-opsonized zymosan by M-CSF stimulated CM (M-CM) being most affected. M-CM that had been cultured in the presence of imatinib were also impaired in their ability to stimulate responder cells in a mixed lymphocyte reaction. These results demonstrate that human monocytes cultured in the presence of imatinib are functionally impaired, and suggest that imatinib displays inhibitory activity against other kinase(s) that play a role in monocyte/macrophage development. PMID- 15661042 TI - Vaccination of brushtail possums, Trichosurus vulpecula, with Bacille Calmette Guerin induces T lymphocytes that reduce Mycobacterium bovis replication in alveolar macrophages via a contact-dependent/nitric oxide-independent mechanism. AB - The permissiveness of alveolar macrophages from brushtail possums for the replication of Mycobacterium bovis was examined. Mycobacterium bovis replication was indirectly measured by assessing bacterial metabolism via the incorporation of [3-H]-uracil by bacilli released from lysed macrophages previously infected with mycobacteria. Alveolar macrophages allowed substantial replication of virulent M. bovis, in contrast to Bacille Calmette-Guerin (BCG) Pasteur, which replicated poorly. The addition of crude lymphokines enhanced the metabolic activity of phagocytosed M. bovis in possum macrophages. Possum lymphokines enhanced the ability of possum macrophages to generate reactive oxygen intermediates, measured by the reduction of nitroblue tetrazolium, which is indicative of an activation process. Similarly, the addition of recombinant possum TNF-alpha enhanced the permissiveness of alveolar macrophages for M. bovis. In contrast to mouse peritoneal macrophages, possum alveolar macrophages did not release significant levels of nitric oxide (NO) after stimulation with M. bovis and/or lymphokines. However, the uptake of virulent M. bovis by possum macrophages was associated with an enhanced ability of cells to release TNF alpha, whereas very low levels of TNF-alpha were released after infection with BCG. The addition of a selective inhibitor of inducible NO synthase had no impact on the replication of M. bovis or BCG in possum macrophages in the presence or absence of lymphokines. Co-culturing infected possum alveolar macrophages with autologous blood mononuclear cells from BCG-vaccinated possums led to a significant decrease in the metabolic activity of intracellular M. bovis. This effect was contact dependent and NO independent and was mediated by a population of CD3+ cells. In addition, adding scavengers of reactive oxygen intermediates did not abrogate this phenomenon. PMID- 15661043 TI - Herpesvirus saimiri immortalization of Aotus T lymphocytes specific for an immunogenically modified peptide of Plasmodium falciparum merozoite surface antigen 2. AB - The Plasmodium merozoite surface antigen 2 (MSA2) is one of several candidates for a protective vaccine against malaria. Previous studies have shown that antibodies directed against the MSA2 variable region are not protective and that constant regions are non-immunogenic. However, modified peptides derived from constant regions can be rendered immunogenic and partially protective in Aotus monkeys. In this study, we reveal the establishment, using in vitro Herpesvirus samiri (HVS) infection, of an Aotus monkey T-cell line (AnTMSA2) specific for a modified immunogenic and partially protective peptide derived from a constant and highly conserved region of MSA2 (SKYSNTFINNAYNMSIRRSM). AnTMSA2 is a CD4 T lymphocyte expressing high levels of MHC class II molecules, CD58 and CD2, which are important for proliferation and growth. AnTMSA2 proliferates specifically in response to the modified monomeric MSA2 peptide sequence. It is also capable of specific antigen recognition after glycine-cysteine-polymerized sequence processing and presentation by autologous APC. Interestingly, AnTMSA2 presents cross-reactivity with D-peptide analogues in which residues in positions 8 and 9 were changed for NDID residues. Therefore, at least for this particular sequence, polymerized D-peptides could be used for immunizing animals without losing the immunogenic epitope. AnTMSA2 presents a cytokine profile corresponding to a Th0 like pattern, which suggests that as a result of HVS immortalization AnTMSA2 is in transit from a Th2 to a Th1 pattern. Taken together our results suggest that Th2 T-cell induction and/or T-cell cross-reactivity generation by the modified peptide could be responsible for the immunogenic conversion observed in Aotus monkeys and that D-peptide analogues with longer half-lives could provide an alternative for inducing protective immunity. PMID- 15661044 TI - Age-related changes in the development of experimental autoimmune encephalomyelitis. AB - A prominent feature of multiple sclerosis is its high incidence of onset in the third decade of life and its relatively rare onset in persons older than 50 years. In order to study age-related restriction of clinical expression, a comparative biochemical, immunological and histological study was undertaken during development of experimental autoimmune encephalomyelitis (EAE) in young (7 weeks) and middle-aged (15 months) Wistar rats. Young rats showed characteristic clinical signs 12-16 days postinduction, and then they spontaneously recuperated. In middle-aged rats, the incidence of clinical signs was significantly reduced, with a later onset of the disease. Similar biochemical and histological alterations were detected in both age groups, but they were present in a later stage in middle-aged animals. However, cellular and humoral immune responses to myelin basic protein (MBP) were observed 15 days postinduction in all EAE animals. The study of anti-MBP IgG isotype pattern in 7-week-old animals indicated a predominant Th1-type immune response during the acute stage of EAE, with antibodies predominantly recognizing the MBP 96-128 peptide. In contrast, 15 month-old animals showed a less prominent Th1 response, without any epitope dominance. The changes in immune function found in middle-aged animals may account for the different susceptibility and expression of EAE, and may also be relevant to the different clinical expression observed in multiple sclerosis with maturation. PMID- 15661045 TI - Activation of dendritic cells by human papillomavirus-like particles through TLR4 and NF-kappaB-mediated signalling, moderated by TGF-beta. AB - Human papillomavirus-like particles (HPV-VLP) are a candidate vaccine for prevention of HPV infection, and also are a candidate for an immunogenic delivery system for incorporated antigen. VLP activate in vitro generated dendritic cells (DC) but not Langerhans cells (LC); however, the mechanism of this activation is unknown. We have shown that uptake and activation of DC by VLP involves proteoglycan receptors and can be inhibited by heparin. Heparin has been shown to activate DC by signalling through Toll-like receptor 4 (TLR4) and nuclear factor (NF)-kappaB. The pathway of DC activation by VLP was further investigated in the present study. Exposure to VLP induced costimulatory molecule expression, RelB translocation and IL-10 production by DC but not by LC. The lack of LC activation was reversible when TGF-beta was removed from the LC medium. VLP-induced induction of costimulatory molecule expression, RelB activation and cytokine secretion by DC was blocked by inhibition of NF-kappaB activation, heparin or TLR4 mAb. The data provide evidence that HPV-VLP signal DC through a pathway involving proteoglycan receptors, TLR4 and NF-kappaB, and shed light on the mechanism by which VLP stimulate immunity in the absence of adjuvants in vivo. LC may resist activation in normal epithelium abundant in TGF-beta, but not in situations in which TGF-beta concentrations are reduced. PMID- 15661046 TI - IFN-gamma overcomes low responsiveness of myeloid dendritic cells to CpG DNA. AB - Dendritic cells (DC) are professional APC that have an extraordinary capacity to prime naive T cells. It has been reported that human DC subsets express distinct toll-like receptor (TLR), which influences their function. In mice, we observed that plasmocytoid DC (pDC) express a higher level of TLR9 compared with myeloid DC (mDC) cultured with GM-CSF. However, we demonstrated that stimulation with IFN gamma is capable of upregulating TLR9 expression in mDC to a level comparable with expression in pDC. Consistent with this observation, IL-12 p40 and IL-6 mRNA expression and IL-12 p70 secretion in response to CpG-oligodeoxynucleotides are enhanced in mDC pretreated with IFN-gamma compared with untreated cells. Therefore, TLR-mediated responses of DC subsets may be influenced not only by signals delivered by pathogens but also by regulatory signals from cytokines such as IFN-gamma. PMID- 15661047 TI - Learning curve of hand-assisted retroperitoneoscopic nephrectomy in less experienced laparoscopic surgeons. AB - AIM: To evaluate the learning curve of hand-assisted retroperitoneoscopic nephrectomy (HALS) performed by less-experienced surgeons. METHODS: The operative records of 166 patients, including 103 with renal tumors and 63 with renal pelvic or ureteral tumors, who underwent HALS performed by 18 less-experienced urologists were reviewed. RESULTS: The insufflation time in the first four cases was significantly longer than that in the sixteenth and later cases. The insufflation time in cases 5-10 was 14-24 min longer than that in the cases 16 onward, although the differences were not significant. The estimated blood loss did not differ in each group of cases. The complication rate in early cases, in which the operators' experience was five cases or less, was 6% (4/71), while that in later cases was also 7% (7/95). In the analysis of the learning curve of a single surgeon who performed 57 procedures, the insufflation time in cases 1-5 was significantly longer than in cases 41-57. The insufflation times in cases 5 10 were 45 min longer than those in cases 41-57, although the difference was not significant. The estimated blood loss did not differ in each group of cases. Complications did not seem related to operation experience. CONCLUSION: In HALS, 5-10 cases were necessary for less-experienced urologists to gain average operating skills for this procedure. It may be reasonable for less-experienced surgeons to begin standard laparoscopic procedures after experiencing 10 cases of the present procedure. PMID- 15661048 TI - Therapeutic effects of long-term administration of an oral adsorbent in patients with chronic renal failure: two-year study. AB - BACKGROUND: Kremezin is an oral adsorbent that attenuates the progression of chronic renal failure by removing uremic toxins and their precursors from the gastrointestinal tract. Previously two clinical studies based on reciprocal serum creatinine levels (1/Scr) have confirmed the efficacy of Kremezin (Kureha Chemical, Tokyo, Japan) in undialyzed patients who had been followed up for 6 months or 1 year. This is the first report to evaluate the therapeutic effects of long-term administration (2 years.) of Kremezin in undialyzed patients. METHODS: Kremezin was given to 48 enrolled undialyzed patients with a median Scr level of 4.3 mg/dL. Rates of decline of 1/Scr, as well as the time for Scr level to reach 10 mg/dL, the critical value requiring dialysis, were compared before and after administration of Kremezin. RESULTS: During the 2-year therapeutic period, 1/Scr gradients were significantly attenuated (P = 0.0083), and the estimated time to dialysis was prolonged from 16.3 +/- 16.3 months to 29.8 +/- 24.1 months (P = 0.002). When the patients were divided into two groups, based on of systolic blood pressure (SBP), defined by the World Health Organization (WHO) classification, a significantly smaller number of patients in the low blood pressure group (SBP < 160 mmHg) were introduced to dialysis (P = 0.0005), and the estimated time to dialysis was significantly extended in the low blood pressure group (P = 0.0125). CONCLUSION: In addition to the control of blood pressure in undialyzed patients, Kremezin has additive salutary effects to halt the progressive loss of renal function, resulting in the delay of dialysis. PMID- 15661049 TI - Spectrum of stone composition: structural analysis of 1050 upper urinary tract calculi from northern India. AB - BACKGROUND: The purpose of the present paper was to study the spectrum of stone composition of upper urinary tract calculi by X-ray diffraction crystallography technique, in patients managed at All India Institute of Medical Sciences. METHODS: Between 30 April 1998 and 31 March 2003, a total of 1050 urinary calculi (900 renal, 150 ureteric) were analyzed. The stone fragments were collected after extracorporeal shock-wave lithotripsy, or retrieval by endoscopic (percutaneous nephrolithotomy, ureterorenoscopy), laparoscopic and various open surgical procedures. The structural analysis of the stones was done using X-ray diffraction crystallography. RESULTS: Four types of primary and three secondary X ray diffraction patterns were obtained. The primary patterns were as follows. Pattern A, well organized crystalline structure; pattern B, moderately organized crystalline structure; pattern C, poorly organized crystalline structure; pattern D, very poorly organized crystalline structure. The three secondary patterns mainly highlighted the mixed variety of stones. These patterns were further analyzed and compared with standard X-ray diffraction (powder) photographs. Of the 1050 stones analyzed, 977 (93.04%) were calcium oxalate stones, out of which 80% were calcium oxalate monohydrate (COM) and 20% were calcium oxalate dihydrate (COD). Fifteen were struvite (1.42%) and 19 were apatite (1.80%). Ten were uric acid stones (0.95%) and the remaining 29 (2.76%) were mixed stones (COM + COD and calcium oxalate + uric acid, calcium oxalate + calcium phosphate, and calcium phosphate + magnesium ammonium phosphate). A total of 89.98% of staghorn stones were made of oxalates (COM/+COD) and only 4.02% were struvite. CONCLUSION: Urinary stone disease in the Indian population is different from that in Western countries, with a larger percentage of patients having calcium oxalate stones, predominantly COM. Also, the majority of staghorn stones (89.98%) were made of oxalates. PMID- 15661050 TI - Altered messenger RNA and protein expressions for insulin-like growth factor family members in clear cell and papillary renal cell carcinomas. AB - BACKGROUND: The purpose of the present paper was to describe the pattern of expression of insulin-like growth factor (IGF-I) and its regulatory binding proteins (IGFBP) in renal cell carcinoma (RCC). METHODS: The expressions of mRNA and protein for various IGF members were assessed in 24 paired normal and malignant human renal tissues (16 clear cell and 8 papillary RCC) using semiquantitative reverse transcription-polymerase chain reaction and immunohistochemistry. Paired tissue samples were also obtained from six patients with oncocytoma in order to compare the specificity of changes in IGF/IGFBP expression between tumors derived from proximal (RCC) and distal (oncocytoma) tubular epithelium. RESULTS: Clear cell RCC were characterized by significant increases in the mRNA expression of IGF-I, IGFBP-3 and IGFBP-6 while papillary RCC exhibited down-regulated expression of IGF-I, IGFBP-4 and IGFBP-5. The IGFBP 2, IGFBP-4 and IGFBP-5 mRNA were down-regulated in oncocytomas. Semiquantitative assessment of immunohistochemical staining demonstrated significant increases in epithelial associated IGF-I and IGFBP-3 in clear cell RCC, increased IGFBP-5 protein in papillary RCC and no significant changes in IGF/IGFBP protein expression in oncocytoma. CONCLUSIONS: The expression of IGF-I and certain IGFBP is significantly altered in RCC compared with normal renal tissue and oncocytomas. This altered expression is differentially regulated according to the histologic subtype of RCC, and suggests that the IGF/IGFBP axis may play an important role in determining the malignant phenotype of RCC. PMID- 15661051 TI - Expression of thymidine phosphorylase in human superficial bladder cancer. AB - BACKGROUND: The purpose of the present paper was to investigate the expression level of thymidine phosphorylase (TPase) in superficial bladder cancer tissues obtained by transurethral resection, and determine whether its expression correlates with tumor recurrence. METHODS: From March 1998 to December 2001, 99 patients with superficial bladder cancer were diagnosed and treated at eight affiliated hospitals. Tissue specimens obtained by transurethral resection of superficial bladder cancer (TURBT) were applied to immunohistochemical study using anti-TPase antibody as well as pathological diagnosis. The data were subjected to statistical analysis. RESULTS: Using MoAb 654-1 as the primary antibody, TPase was clearly stained in human bladder cancer tissues. The maximum TPase level measured by enzyme-linked immunosorbent assay (ELISA) method in normal bladder tissues was 18.7 U/mg protein. The TPase activity was 2.8-fold higher in tumors than in normal bladder samples (P = 0.037). The TPase positivity rates determined by immunohistochemical and ELISA methods were distinctly correlated (P = 0.046). For the recurrence-free rates in pT1 tumors treated by TURBT alone (n = 46), there were no statistically significant differences between Tpase-positive or -negative cases. CONCLUSIONS: The TPase expression determined by ELISA and immunohistochemistry is significantly up-regulated in superficial bladder tumors compared with normal bladder samples. However, TPase expression by immunohistochemistry is not a predictive index of recurrence-free rate for superficial bladder cancer treated with TURBT alone. PMID- 15661052 TI - Sexual function before and after mesh repair of inguinal hernia. AB - AIM: Several factors having an influence on the quality of life after an inguinal hernia repair have been studied, yet little has been reported on sexual function before and after this operation. METHODS: In a prospective follow-up study from January 1999 to July 2002, 210 men and 14 women were asked to answer an anonymous questionnaire of 40 questions before elective inguinal hernia repair to assess pre- and postoperative sexual function (preoperatively, 3 months postoperatively and every 6 months afterwards). Inguinal hernia repair was performed using a standardized Plug and Patch mesh technique. RESULTS: Fifty-two (23.2%) patients mentioned preoperative sexual dysfunction related to the groin hernia. Postoperatively, the surgical repair had a positive influence on the sexual function in these patients. Thirty-six (16%) patients complained of postoperative sexual dysfunctions, which improved or disappeared over the next few months. All patients suffered surgical complications, which were surgically treated in six cases. Symptoms specifically associated with inguinal hernias can cause certain limitations in the sexual life. CONCLUSIONS: In most cases, and in the absence of postoperative complications, the operative repair can lead to a recovery of the sexual life in patients with preoperative sexual dysfunction, while, in most cases, it does not affect patients with a preoperative normal sexual life. PMID- 15661053 TI - Construction of the Mandarin version of the International Prostate Symptom Score inventory in assessing lower urinary tract symptoms in a Malaysian population. AB - BACKGROUND: The purpose of the present study was to validate the Mandarin version of the International Prostate Symptom Score (Mand-IPSS) in a Malaysian population. METHODS: The validity and reliability were studied in patients with lower urinary tract symptoms (LUTS; benign prostatic hyperplasia [BPH] group) and without LUTS (control group). Test-retest methodology was used to assess the reliability while Cronbach alpha was used to assess the internal consistency. Sensitivity to change was used to express the effect size index in the preintervention versus post-intervention score in patients with LUTS who underwent transurethral resection of the prostate. RESULTS: For the control group and BPH group, the internal consistency was excellent and a high degree of internal consistency was observed for all seven items (Cronbach alpha = 0.86-0.98 and 0.90-0.98, respectively). Test-retest correlation coefficients for all items were highly significant. Intraclass correlation coefficient (ICC) was high for the control (ICC = 0.93-0.99) and BPH group (ICC = 0.91-0.99). The sensitivity and specificity showed a high degree of sensitivity and specificity to the effects of treatment. A high degree of significance between baseline and post treatment scores was observed across all seven items in the BPH group but not in the control group. CONCLUSIONS: The Mand-IPSS is a suitable, reliable, valid and sensitive instrument to measure clinical change in the Malaysian population. PMID- 15661054 TI - Clinicopathological statistics on registered prostate cancer patients in Japan: 2000 report from the Japanese Urological Association. AB - BACKGROUND: The purpose of the present paper was to investigate etiology, diagnosis, initial treatment, pathological findings and final outcomes for prostate cancer in Japan. METHODS: From 2001, the Japanese Urological Association initiated computer-based registration of prostate cancer patients in Japan to estimate etiology, diagnosis, initial planed treatment, pathological findings and final outcome. RESULTS: A total of 173 institutions responded and 4529 patients who were diagnosed with prostate cancer in 2000 were registered. In the first year, background factors, diagnostic procedures and initially planned treatment were recorded. The analysis of these registered cases is presented here. Nearly 30% of the cases demonstrated <10 ng/mL of prostate specific antigen. The six core biopsy was the most commonly used procedure. The clinical T staging distribution was as follows: T1c, 20.3%; T2a, 21.8%; T2b, 17.3%; T3a, 15.8%; T3b, 11.0%; and T4, 8%. More than 70% of cases were diagnosed as M0. Hormone therapy alone was the initial treatment plan in more than half of the cases. CONCLUSION: This is the first report on prostate cancer patients in Japan based on multi institutional registration. Pathological findings and final outcome will be surveyed later by the Japanese Urological Association. After 5 years, adopted treatment, pathological results and final outcome will be registered. PMID- 15661055 TI - The effectiveness of early primary realignment in children with posterior urethral injury. AB - AIM: The aim of this retrospective study was to compare the results of delayed repair and early primary realignments in patients with posterior urethral injury. METHODS: From 1990 to 2003, 20 children were admitted to the Medical Faculty of Uluday University, Bursa, Turkey, for posterior urethral injuries. Traffic accidents were the most common cause of injury (n = 17). Twelve patients (60%) who were referred early (1-10 days) underwent early realignment over a urethral tube. A total of eight patients (40%) underwent delayed repair using transpubic route. In these patients, surgical repair of the urethra was performed 5-6 months later. RESULTS: Of the 12 patients who underwent early urethral realignment, six required at least one visual internal urethrotomy following the removal of the urethral catheter. Urethral stricture developed in two of 12 patients (16.6%) who underwent early urethral realignment. Of the eight patients who underwent delayed repair, six required at least one visual internal urethrotomy following removal of the urethral catheter. Urethral stricture developed in three of eight patients (37.5%) who underwent delayed repair. This difference was statistically significant (P < 0.05). CONCLUSION: The urethral stricture in patients who underwent early primary realignment was less developed than the stricture that developed in those who underwent delayed management. According to these results we recommend early primary realignment in children with posterior urethral injury. PMID- 15661056 TI - In vitro chemosensitivity test for human genito-urinary tumors using collagen gel matrix. AB - BACKGROUND: Although the aim of chemosensitivity tests is to predict the efficacy of anticancer agents for individual patients, no generally accepted assay has been established. METHODS: A chemosensitivity test was conducted for solid tumors with an organ culture system using collagen gel matrix (CGM). Seventy-five samples of transitional cell carcinoma (TCC), 20 of germ cell tumor (GCT) and 13 of renal cell carcinoma (RCC) were used for the chemosensitivity test, and 20 patients were treated with anticancer drugs on the basis of the test results. RESULTS: Positive rates of anticancer drugs for the 75 TCC samples were 64.9% for carboplatin, 63.4% for cisplatin, 32.1% for etoposide, 19.7% for THP-adriamycin, 16.7% for vinblastine, and 12.3% for methotrexate, indicating that positive rates of the latter three agents consisting of an MVAC regimen were unexpectedly low. The GCT had higher positive rates than the other cancers while RCC had the lowest. In 20 eligible patients (seven patients with bladder tumors and 13 with GCT), the true positive and true negative rates were 42% (5/12) and 75% (6/8), respectively, and the sensitivity and specificity were 71% (5/7) and 46% (6/13), resulting in a 55% (11/20) accurate predictive value. CONCLUSION: Although predictive accuracy was moderate when combination chemotherapy was used, information about chemosensitivity may have some beneficial effect on the treatment of patients with invasive bladder cancer or advanced GCT, because insensitive drugs detected by the test could be deleted or replaced with more sensitive ones. PMID- 15661057 TI - Genistein induces cell growth inhibition in prostate cancer through the suppression of telomerase activity. AB - AIM: To clarify the mechanism of the anticancer effect of genistein, we examined the effect of genistein on telomerase activity in prostate cancer cells. We hypothesized that genistein may exert its anticancer effect by modifying telomerase activity in prostate cancer cells. METHODS: Prostate cancer (LNCaP) cells were cultured with genistein and the number of viable cells was counted. Growth medium was also collected to measure prostate-specific antigen (PSA) concentration. Polymerase chain reaction (PCR)-based telomeric repeat amplification protocol (TRAP) assay and reverse transcriptase (RT)-PCR analysis were performed to investigate telomerase activity and the expression of human telomerase reverse transcriptase (hTERT), c-myc and p21 mRNA. To examine the possibility that hTERT transcriptional activity is modulated by genistein, transient cell transfection studies were performed by using luciferase reporter assay. Telomere repeat amplification protocol (TRAP) assay and PCR analysis of hTERT were performed in androgen independent cells, DU-145. RESULTS: Cell growth of LNCaP was inhibited by genistein and PSA secretion was similarly reduced. In TRAP assay, the telomerase activity of LNCaP cells was reduced by genistein. Reverse transcriptase-PCR analysis revealed that the expression of hTERT and c myc mRNA was down-regulated by genistein, whereas p21 mRNA increased in response to genistein. Luciferase reporter assay revealed that genistein reduced the transcriptional activity of hTERT. In DU-145 cells, telomerase activity and the expression of hTERT mRNA were also reduced by genistein. CONCLUSION: The current study elucidated the molecular mechanism of cell growth inhibition by genistein. The antiproliferative effects of genistein seem to be exerted on the hTERT transcriptional activity via different molecular pathways. PMID- 15661058 TI - Significance of determining the point of reperfusion failure in experimental torsion of testis. AB - BACKGROUND: Experimental studies of the use of free radical scavengers in ischemic/reperfusion (I/R) injury following detorsion of the torted testis have yielded conflicting results due to differences in the period of ischemia used. The authors studied I/R injury in the rabbit model, to define the point beyond which there is reperfusion failure. METHODS: Ischemia/reperfusion injury of the testis was created in 3-6-month-old male New Zealand white rabbits by cross clamping the left spermatic cord for periods of ischemia lasting 0, 15, 30, 60, 90, 120 and 180 min. There were eight animals per experimental group. The right testis served as internal control. Both testes were harvested after 24 h of reperfusion in four animals and after 3 months in the remaining four animals for each group. Testicular malondialdehyde (MDA), a measure of free radical damage, was determined by using the thiobarbituric acid reaction on testicular homogenates. Johnsen score was used to assess morphological damage caused by the ischemia. RESULTS: After 24 h of reperfusion, the mean testicular MDA in the control right testes at 0, 15, 30, 60, 90, 120 and 180 min was 2.1, 2.5, 2.9, 2.4, 2.1 and 1.9 nmol/mg protein, respectively. The mean left testicular MDA at corresponding ischemic periods was 1.6, 2.0, 3.9, 10.0, 4.4, 6.1 and 1.0 nmol/mg protein, respectively. The maximum left testicular MDA was at 60 min (10.0 nmol/mg protein), following which the level dropped significantly to 1.0 nmol/mg protein at 180 min. At 3 months, the mean Johnsen scores for left testes subjected to 0, 60, 120 and 180 min ischemia were 9.4, 8.8, 2.3, 3.5, respectively. CONCLUSION: The results suggest that following ischemia of up to 60 min in the rabbit testis, adequate reperfusion is possible, but ischemia lasting beyond 60 min results in inadequate reperfusion leading to irreversible damage. Thus, in experiments for assessing the effect of antioxidants on I/R injury of the testis in rabbits, periods up to 60 min of ischemia should be regarded as optimum to observe an effect. PMID- 15661059 TI - Splenogonadal fusion: report of a case and review of the literature. AB - We report a case of splenogonadal fusion masquerading as a testicular tumor and review the etiology, the pathogenesis and the management of this rare disease. A 20-year-old male patient presented with clinical and radiological findings of a left testicular mass for which he underwent a left radical inguinal orchidectomy. Histopathology of the removed testicle revealed no tumor. The gonad was fused to ectopic splenic tissue, the later forming a distinct encapsulated mass attached to the lower pole of the testicle inside the tunica vaginalis. Splenogonadal fusion, although a rare condition, may account for a non-malignant testicular mass and should be suspected in young patients with other congenital anomalies. PMID- 15661060 TI - Renal papillae metastasis of sigmoid colon adenocarcinoma. AB - Colonic carcinoma metastatic to the kidney is very rare. The usual anatomical localization for secondary renal neoplasms is the renal cortex. We report a case of sigmoid colon carcinoma with unilateral kidney metastasis localized only in the renal papillae without obvious metastatic disease. PMID- 15661062 TI - Bilateral ureteral fibroepithelial polyps and review of the literature. AB - Despite being the most common forms of benign ureteral neoplasms, ureteral fibroepithelial polyps are rare and their etiology is still unknown. To our knowledge, we report the fourth case of bilateral fibroepithelial polyps in the English literature with a discussion of clinical features and etiology. PMID- 15661061 TI - Solitary psoas muscle metastasis after radical nephrectomy for renal cell carcinoma. AB - Skeletal muscle is a very rare location for the metastasis of renal cell carcinoma (RCC) and only one case of solitary metastasis to the psoas muscle has been reported. We present a 63-year-old male patient with late recurrence (14 years) after left side radical nephrectomy for RCC. He first visited Chikushi Hospital, Fukuoka University, Japan in January 2000 for a postoperative follow-up because he had shifted residence to the area. Follow-up was by abdominal computed tomography (CT) and chest X-ray. In December 2001, a CT scan showed a 1.5 cm enhanced mass in the right psoas muscle without any other metastasis. The mass was resected that month and histological study showed RCC metastasis. PMID- 15661063 TI - Can epithelized neobladder-cutaneous fistula provide urinary continence? AB - The paper presents a case of a boy born with a bladder exstrophy, who underwent several surgical procedures, including bladder augmentation. During the last procedure, the patient received a suprapubic catheter that accidentally slipped out. The epithelized neobladder-cutaneous fistula resulted in a continent and easily catheterizable abdominal stoma. PMID- 15661064 TI - Bladder cancer discovered by ovarian metastasis: cytokeratin expression is useful when making differential diagnosis. AB - A 49-year-old woman underwent hysterectomy and bilateral adnexectomy after the diagnosis of a right ovarian tumor with paraaortic and pelvic lymph node metastases. The pathological diagnosis was undifferentiated carcinoma of the ovary. After the operation, a bladder tumor was discovered during the evaluation for microscopic hematuria. The bladder tumor was pathologically diagnosed as transitional cell carcinoma, pT1b, G3. Although the pathological findings of the bladder cancer and ovarian cancer were very similar, we could diagnose primary bladder cancer with ovary and lymph node metastases according to the immunohistochemical staining pattern of cytokeratins 7 and 20. Herein, the clinical usefulness of immunohistochemical staining using cytokeratins for making a differential diagnosis of the origin of a tumor in the pelvic cavity is demonstrated. PMID- 15661065 TI - Small cell carcinoma of the prostate with hypercalcemia. AB - We present a case of small cell prostate carcinoma with hypercalcemia in a 75 year-old man. He was diagnosed as having stage T3bN1M0 adenocarcinoma of the prostate. His serum prostate-specific antigen level was reduced to below the normal range after a combination treatment of a luteinizing hormone-releasing hormone agonist and flutamide for prostate carcinoma. He subsequently experienced increasing fatigue, poor appetite, short time loss of consciousness and pain in his lower abdomen. His serum calcium level and carcinoembryonic antigen were increased. He died 5 months from the start of treatment. The autopsy revealed small cell carcinoma of the prostate and multiple metastasis of the lung, liver, pancreas, lymph nodes and spine. PMID- 15661066 TI - Torsion of a communicating hydrocele in a child. AB - Among the diseases presenting as acute scrotum, torsion of a communicating hydrocele is extremely rare. A 5-year-old boy was referred with discomfort and swelling of the left scrotum. Operative findings revealed a torsion of a communicating hydrocele. We report a case of torsion with preoperative findings of ultrasonography and magnetic resonance imaging. We also emphasize the awareness of this disease as a differential diagnosis of the acute scrotum. PMID- 15661067 TI - Asthenozoospermia: possible association with long-term exposure to an anti epileptic drug of carbamazepine. AB - Little attention has been paid to infertility in men with epilepsy and little information exists about the mechanisms by which anti-epileptic drugs affect spermatogenesis or sperm function. We report a case of a male infertility patient with asthenozoospermia during long-term treatment with anti-epileptic drugs. A 29 year-old man had continued treatment with anti-epileptic drugs under the diagnosis of epilepsy for 13 years. He and his wife had been examined and treated as an infertile couple for 3 years. The patient was found to have no motile sperm with a normal sperm count, while taking a dose of 400 mg/day of carbamazepine. On suspicion of an adverse effect of carbamazepine, he was switched to phenytoin monotherapy. One month after that, sperm motility was vastly improved (65%) and they conceived a child 5 months after that. One must be cautious in extrapolating from a case report, but these findings strongly suggest a direct effect of carbamazepine on spermatic function. PMID- 15661068 TI - Retroperitoneal seminoma with 'burned out' phenomenon in the testis. AB - The rare 'burned out' phenomenon in germ cell tumors is known as the presence of an extragonadal germ cell tumor without traces of neoplasm in the testis. This condition is different and less common from the primary extragonadal germ cell malignancies. These malignancies are treated surgically with or without adjuvant chemotherapy or radiotherapy and their prognosis is better than that of other types of primary extragonadal tumors. PMID- 15661069 TI - Female para-urethral adenocarcinoma: histological and immunohistochemical study. AB - BACKGROUND: Female urethral cancer with a diverticular form is assumed to originate from the para-urethral duct, which is embryologically homologous to the male prostate gland. The purpose of the present paper was to investigate the female para-urethral adenocarcinomas histologically and immunohistochemically. METHODS: Surgical specimens obtained from six female patients with para-urethral adenocarcinomas were examined histologically, and an immunohistochemical study using antibodies against carcinoembryonic antigen (CEA), prostate specific antigen (PSA), and chromogranin A was performed. RESULTS: On histologic examination, the female para-urethral cancers were divided into five cases of mucin-producing-type adenocarcinoma and one case of clear cell-type adenocarcinoma. All five mucin-producing-type adenocarcinomas were positive with anti-CEA, and two of them showed neuroendicrine differentiation. One of them showed a focally positive area with anti-PSA. The clear cell-type adenocarcinoma had no positive reactions to these antibodies. CONCLUSIONS: On the basis of histologic structure, positive CEA staining, and the presence of focal neuroendocrine differentiation, mucin-producing-type adenocarcinomas may arise from the proximal part of the para-urethral duct. PMID- 15661070 TI - Increased polyclonal immunoglobulin reactivity toward human and bacterial proteins is associated with clinical protection in human Plasmodium infection. AB - BACKGROUND: Polyclonal B-cell activation is well known to occur in Plasmodium infections, but its role in pathogenesis or protection remains unclear. However, protective properties of natural antibodies have previously been demonstrated in other contexts. METHODS: Sera from asymptomatic and symptomatic Plasmodium infected subjects locally detected in a survey study in the Brazilian Amazon, and from unexposed and exposed but presently uninfected control subjects, were assayed by a standardized quantitative immunoblot method allowing simultaneous detection of IgG or IgM reactivity to a large number of parasite-unrelated proteins. RESULTS: In subjects free of coinfection with hepatitis B virus, IgG reactivity to human brain antigens and Escherichia coli proteins was strikingly enhanced in asymptomatic Plasmodium-infected individuals when compared to such with clinical malaria symptoms, or to uninfected control subjects. This difference was most characteristic for limited exposure times (less than ten years locally, or 20 years in endemic areas). It was more significant than a similar trend found for IgG to Plasmodium falciparum antigens, and unrelated to parasitaemia levels. Asymptomatic subjects with comparatively short exposure characteristically showed relatively elevated IgG versus IgM reactivity. Polyclonal IgG reactivity appears triggered by previous P. falciparum but not Plasmodium vivax malaria. CONCLUSION: The observed difference in polyclonal antibody production seems related to intrinsic activation states of infected individuals, rather than to parasite-antigen specific immune responses. However, it appears influenced by preceding stimuli. This supports the idea that acquired clinical immunity may not exclusively depend on antigen-specific responses, but also on the individual polyclonal reaction. PMID- 15661072 TI - Non-linear mapping for exploratory data analysis in functional genomics. AB - BACKGROUND: Several supervised and unsupervised learning tools are available to classify functional genomics data. However, relatively less attention has been given to exploratory, visualisation-driven approaches. Such approaches should satisfy the following factors: Support for intuitive cluster visualisation, user friendly and robust application, computational efficiency and generation of biologically meaningful outcomes. This research assesses a relaxation method for non-linear mapping that addresses these concerns. Its applications to gene expression and protein-protein interaction data analyses are investigated. RESULTS: Publicly available expression data originating from leukaemia, round blue-cell tumours and Parkinson disease studies were analysed. The method distinguished relevant clusters and critical analysis areas. The system does not require assumptions about the inherent class structure of the data, its mapping process is controlled by only one parameter and the resulting transformations offer intuitive, meaningful visual displays. Comparisons with traditional mapping models are presented. As a way of promoting potential, alternative applications of the methodology presented, an example of exploratory data analysis of interactome networks is illustrated. Data from the C. elegans interactome were analysed. Results suggest that this method might represent an effective solution for detecting key network hubs and for clustering biologically meaningful groups of proteins. CONCLUSION: A relaxation method for non-linear mapping provided the basis for visualisation-driven analyses using different types of data. This study indicates that such a system may represent a user-friendly and robust approach to exploratory data analysis. It may allow users to gain better insights into the underlying data structure, detect potential outliers and assess assumptions about the cluster composition of the data. PMID- 15661071 TI - Evaluating the evidence for models of life course socioeconomic factors and cardiovascular outcomes: a systematic review. AB - BACKGROUND: A relatively consistent body of research supports an inverse graded relationship between socioeconomic status (SES) and cardiovascular disease (CVD). More recently, researchers have proposed various life course SES hypotheses, which posit that the combination, accumulation, and/or interactions of different environments and experiences throughout life can affect adult risk of CVD. Different life course designs have been utilized to examine the impact of SES throughout the life course. This systematic review describes the four most common life course hypotheses, categorizes the studies that have examined the associations between life course SES and CVD according to their life course design, discusses the strengths and weaknesses of the different designs, and summarizes the studies' findings. METHODS: This research reviewed 49 observational studies in the biomedical literature that included socioeconomic measures at a time other than adulthood as independent variables, and assessed subclinical CHD, incident CVD morbidity and/or mortality, and/or the prevalence of traditional CVD risk factors as their outcomes. Studies were categorized into four groups based upon life course design and analytic approach. The study authors' conclusions and statistical tests were considered in summarizing study results. RESULTS: Study results suggest that low SES throughout the life course modestly impacts CVD risk factors and CVD risk. Specifically, studies reviewed provided moderate support for the role of low early-life SES and elevated levels of CVD risk factors and CVD morbidity and mortality, little support for a unique influence of social mobility on CVD, and consistent support for the detrimental impact of the accumulation of negative SES experiences/conditions across the life course on CVD risk. CONCLUSIONS: While the basic life course SES study designs have various methodologic and conceptual limitations, they provide an important approach from which to examine the influence of social factors on CVD development. Some limitations may be addressed through the analysis of study cohorts followed from childhood, the evaluation of CVD risk factors in early and middle adulthood, and the use of multiple SES measures and multiple life course analysis approaches in each life course study. PMID- 15661073 TI - A7DB: a relational database for mutational, physiological and pharmacological data related to the alpha7 nicotinic acetylcholine receptor. AB - BACKGROUND: Nicotinic acetylcholine receptors (nAChRs) are pentameric proteins that are important drug targets for a variety of diseases including Alzheimer's, schizophrenia and various forms of epilepsy. One of the most intensively studied nAChR subunits in recent years has been alpha7. This subunit can form functional homomeric pentamers (alpha7)5, which can make interpretation of physiological and structural data much simpler. The growing amount of structural, pharmacological and physiological data for these receptors indicates the need for a dedicated and accurate database to provide a means to access this information in a coherent manner. DESCRIPTION: A7DB http://www.lgics.org/a7db/ is a new relational database of manually curated experimental physiological data associated with the alpha7 nAChR. It aims to store as much of the pharmacology, physiology and structural data pertaining to the alpha7 nAChR. The data is accessed via web interface that allows a user to search the data in multiple ways: 1) a simple text query 2) an incremental query builder 3) an interactive query builder and 4) a file-based uploadable query. It currently holds more than 460 separately reported experiments on over 85 mutations. CONCLUSIONS: A7DB will be a useful tool to molecular biologists and bioinformaticians not only working on the alpha7 receptor family of proteins but also in the more general context of nicotinic receptor modelling. Furthermore it sets a precedent for expansion with the inclusion of all nicotinic receptor families and eventually all cys-loop receptor families. PMID- 15661074 TI - Functional promoter upstream p53 regulatory sequence of IGFBP3 that is silenced by tumor specific methylation. AB - BACKGROUND: Insulin-like growth factor binding protein (IGFBP)-3 functions as a carrier of insulin-like growth factors (IGFs) in circulation and a mediator of the growth suppression signal in cells. There are two reported p53 regulatory regions in the IGFBP3 gene; one upstream of the promoter and one intronic. We previously reported a hot spot of promoter hypermethylation of IGFBP-3 in human hepatocellular carcinomas and derivative cell lines. As the hot spot locates at the putative upstream p53 consensus sequences, these p53 consensus sequences are really functional is a question to be answered. METHODS: In this study, we examined the p53 consensus sequences upstream of the IGFBP-3 promoter for the p53 induced expression of IGFBP-3. Deletion, mutagenesis, and methylation constructs of IGFBP-3 promoter were assessed in the human hepatoblastoma cell line HepG2 for promoter activity. RESULTS: Deletions and mutations of these sequences completely abolished the expression of IGFBP-3 in the presence of p53 overexpression. In vitro methylation of these p53 consensus sequences also suppressed IGFBP-3 expression. In contrast, the expression of IGFBP-3 was not affected in the absence of p53 overexpression. Further, we observed by electrophoresis mobility shift assay that p53 binding to the promoter region was diminished when methylated. CONCLUSION: From these observations, we conclude that four out of eleven p53 consensus sequences upstream of the IGFBP-3 promoter are essential for the p53 induced expression of IGFBP-3, and hypermethylation of these sequences selectively suppresses p53 induced IGFBP-3 expression in HepG2 cells. PMID- 15661076 TI - Examination of Cholesterol oxidase attachment to magnetic nanoparticles. AB - Magnetic nanoparticles (Fe3O4) were synthesized by thermal co-precipitation of ferric and ferrous chlorides. The sizes and structure of the particles were characterized using transmission electron microscopy (TEM). The size of the particles was in the range between 9.7 and 56.4 nm. Cholesterol oxidase (CHO) was successfully bound to the particles via carbodiimide activation. FTIR spectroscopy was used to confirm the binding of CHO to the particles. The binding efficiency was between 98 and 100% irrespective of the amount of particles used. Kinetic studies of the free and bound CHO revealed that the stability and activity of the enzyme were significantly improved upon binding to the nanoparticles. Furthermore, the bound enzyme exhibited a better tolerance to pH, temperature and substrate concentration. The activation energy for free and bound CHO was 13.6 and 9.3 kJ/mol, respectively. This indicated that the energy barrier of CHO activity was reduced upon binding onto Fe3O4 nanoparticles. The improvements observed in activity, stability, and functionality of CHO resulted from structural and conformational changes of the bound enzyme. The study indicates that the stability and activity of CHO could be enhanced via attachment to magnetic nanoparticles and subsequently will contribute to better uses of this enzyme in various biological and clinical applications. PMID- 15661075 TI - Common variants of the beta and gamma subunits of the epithelial sodium channel and their relation to plasma renin and aldosterone levels in essential hypertension. AB - BACKGROUND: Rare mutations of the epithelial sodium channel (ENaC) result in the monogenic hypertension form of Liddle's syndrome. We decided to screen for common variants in the ENaC beta and gamma subunits in patients with essential hypertension and to relate their occurrence to the activity of circulating renin angiotensin-aldosterone system. METHODS: Initially, DNA samples from 27 patients with low renin/low aldosterone hypertension were examined. The DNA variants were subsequently screened for in 347 patients with treatment-resistant hypertension, 175 male subjects with documented long-lasting normotension and 301 healthy Plasma renin and aldosterone levels were measured under baseline conditions and during postural and captopril challenge tests. RESULTS: Two commonly occurring betaENaC variants (G589S and a novel intronic i12-17CT substitution) and one novel gammaENaC variant (V546I) were detected. One of these variants occurred in a heterozygous form in 32 patients, a prevalence (9.2%) significantly higher than that in normotensive males (2.9%, p = 0.007) and blood donors (3.0%, p = 0.001). betaENaC i12-17CT was significantly more prevalent in the hypertension group than in the two control groups combined (4.6% vs. 1.1%, p = 0.001). When expressed in Xenopus oocytes, neither of the two ENaC amino acid-changing variants showed a significant difference in activity compared with ENaC wild-type. No direct evidence for a mRNA splicing defect could be obtained for the betaENaC intronic variant. The ratio of daily urinary potassium excretion to upright and mean (of supine and upright values) plasma renin activity was higher in variant allele carriers than in non-carriers (p = 0.034 and p = 0.048). CONCLUSIONS: At least 9% of Finnish patients with hypertension admitted to a specialized center carry genetic variants of beta and gammaENaC, a three times higher prevalence than in the normotensive individuals or in random healthy controls. Patients with the variant alleles showed an increased urinary potassium excretion rate in relation to their renin levels. PMID- 15661078 TI - MILANO--custom annotation of microarray results using automatic literature searches. AB - BACKGROUND: High-throughput genomic research tools are becoming standard in the biologist's toolbox. After processing the genomic data with one of the many available statistical algorithms to identify statistically significant genes, these genes need to be further analyzed for biological significance in light of all the existing knowledge. Literature mining--the process of representing literature data in a fashion that is easy to relate to genomic data--is one solution to this problem. RESULTS: We present a web-based tool, MILANO (Microarray Literature-based Annotation), that allows annotation of lists of genes derived from microarray results by user defined terms. Our annotation strategy is based on counting the number of literature co-occurrences of each gene on the list with a user defined term. This strategy allows the customization of the annotation procedure and thus overcomes one of the major limitations of the functional annotations usually provided with microarray results. MILANO expands the gene names to include all their informative synonyms while filtering out gene symbols that are likely to be less informative as literature searching terms. MILANO supports searching two literature databases: GeneRIF and Medline (through PubMed), allowing retrieval of both quick and comprehensive results. We demonstrate MILANO's ability to improve microarray analysis by analyzing a list of 150 genes that were affected by p53 overproduction. This analysis reveals that MILANO enables immediate identification of known p53 target genes on this list and assists in sorting the list into genes known to be involved in p53 related pathways, apoptosis and cell cycle arrest. CONCLUSIONS: MILANO provides a useful tool for the automatic custom annotation of microarray results which is based on all the available literature. MILANO has two major advances over similar tools: the ability to expand gene names to include all their informative synonyms while removing synonyms that are not informative and access to the GeneRIF database which provides short summaries of curated articles relevant to known genes. MILANO is available at http://milano.md.huji.ac.il. PMID- 15661077 TI - The effect of IL-13 and IL-13R130Q, a naturally occurring IL-13 polymorphism, on the gene expression of human airway smooth muscle cells. AB - BACKGROUND: Growing evidence shows that interleukin 13 (IL-13) may play an essential role in the development of airway inflammation and bronchial hyper responsiveness (BHR), two defining features of asthma. Although the underlying mechanisms remain unknown, a number of reports have shown that IL-13 may exert its deleterious effects in asthma by directly acting on airway resident cells, including epithelial cells and airway smooth muscle cells. In this report, we hypothesize that IL-13 may participate in the pathogenesis of asthma by activating a set of "pro-asthmatic" genes in airway smooth muscle (ASM) cells. METHODS: Microarray technology was used to study the modulation of gene expression of airway smooth muscle by IL-13 and IL-13R130Q. TaqMan Real Time PCR and flow cytometry was used to validate the gene array data. RESULTS: IL-13 and the IL-13 polymorphism IL-13R130Q (Arg130Gln), recently associated with allergic asthma, seem to modulate the same set of genes, which encode many potentially interesting proteins including vascular cellular adhesion molecule (VCAM)-1, IL 13Ralpha2, Tenascin C and Histamine Receptor H1, that may be relevant for the pathogenesis of asthma. CONCLUSIONS: The data supports the hypothesis that gene modulation by IL-13 in ASM may be essential for the events leading to the development of allergic asthma. PMID- 15661079 TI - Reflections of physiotherapy students in the United Arab Emirates during their clinical placements: a qualitative study. AB - BACKGROUND: Although Western models of education are being used to establish health professional programs in non-Western countries, little is known about how students in these countries perceive their learning experiences. The purpose of this qualitative study was to describe the reflections of physiotherapy students from a Middle East culture during their clinical placements and to compare them to reflections of physiotherapy students from a Western culture. METHODS: Subjects were six senior students (3 females, 3 males, mean age 22.6 years) and 15 junior, female students (mean age 20.1 years) in the baccalaureate physiotherapy program at a university in the United Arab Emirates (UAE). They wrote weekly entries in a journal while in their clinical placements. They described an event, their reaction to it, and how it might affect their future behavior. Two evaluators independently read and coded the content of all the journals, and then worked together to categorize the data and develop themes. A third evaluator, an UAE national, independently read the journals to validate the content analysis. A feedback session with students was used to further validate the data interpretation. The themes were compared to those derived from a similar study of Canadian physiotherapy students. RESULTS: The content of the students' reflections were grouped into 4 themes: professional behavior, awareness of learning, self-development and shift to a patient orientation, and identification and analysis of ethical issues. Although the events were different, students from the UAE considered many of the same issues reflected on by Canadian students. CONCLUSION: Physiotherapy students from a Middle East culture consider many of the same issues as students from a Western culture when asked to reflect on their clinical experience. They reflect on their personal growth, on how they learn in a clinical setting, and on the ethical and professional behaviors of themselves and others. PMID- 15661080 TI - Public appraisal of government efforts and participation intent in medico-ethical policymaking in Japan: a large scale national survey concerning brain death and organ transplant. AB - BACKGROUND: Public satisfaction with policy process influences the legitimacy and acceptance of policies, and conditions the future political process, especially when contending ethical value judgments are involved. On the other hand, public involvement is required if effective policy is to be developed and accepted. METHODS: Using the data from a large-scale national opinion survey, this study evaluates public appraisal of past government efforts to legalize organ transplant from brain-dead bodies in Japan, and examines the public's intent to participate in future policy. RESULTS: A relatively large percentage of people became aware of the issue when government actions were initiated, and many increasingly formed their own opinions on the policy in question. However, a significant number (43.3%) remained unaware of any legislative efforts, and only 26.3% of those who were aware provided positive appraisals of the policymaking process. Furthermore, a majority of respondents (61.8%) indicated unwillingness to participate in future policy discussions of bioethical issues. Multivariate analysis revealed the following factors are associated with positive appraisals of policy development: greater age; earlier opinion formation; and familiarity with donor cards. Factors associated with likelihood of future participation in policy discussion include younger age, earlier attention to the issue, and knowledge of past government efforts. Those unwilling to participate cited as their reasons that experts are more knowledgeable and that the issues are too complex. CONCLUSIONS: Results of an opinion survey in Japan were presented, and a set of factors statistically associated with them were discussed. Further efforts to improve policy making process on bioethical issues are desirable. PMID- 15661081 TI - Red maca (Lepidium meyenii) reduced prostate size in rats. AB - BACKGROUND: Epidemiological studies have found that consumption of cruciferous vegetables is associated with a reduced risk of prostate cancer. This effect seems to be due to aromatic glucosinolate content. Glucosinolates are known for have both antiproliferative and proapoptotic actions. Maca is a cruciferous cultivated in the highlands of Peru. The absolute content of glucosinolates in Maca hypocotyls is relatively higher than that reported in other cruciferous crops. Therefore, Maca may have proapoptotic and anti-proliferative effects in the prostate. METHODS: Male rats treated with or without aqueous extracts of three ecotypes of Maca (Yellow, Black and Red) were analyzed to determine the effect on ventral prostate weight, epithelial height and duct luminal area. Effects on serum testosterone (T) and estradiol (E2) levels were also assessed. Besides, the effect of Red Maca on prostate was analyzed in rats treated with testosterone enanthate (TE). RESULTS: Red Maca but neither Yellow nor Black Maca reduced significantly ventral prostate size in rats. Serum T or E2 levels were not affected by any of the ecotypes of Maca assessed. Red Maca also prevented the prostate weight increase induced by TE treatment. Red Maca administered for 42 days reduced ventral prostatic epithelial height. TE increased ventral prostatic epithelial height and duct luminal area. These increases by TE were reduced after treatment with Red Maca for 42 days. Histology pictures in rats treated with Red Maca plus TE were similar to controls. Phytochemical screening showed that aqueous extract of Red Maca has alkaloids, steroids, tannins, saponins, and cardiotonic glycosides. The IR spectra of the three ecotypes of Maca in 3800-650 cm (-1) region had 7 peaks representing 7 functional chemical groups. Highest peak values were observed for Red Maca, intermediate values for Yellow Maca and low values for Black Maca. These functional groups correspond among others to benzyl glucosinolate. CONCLUSIONS: Red Maca, a cruciferous plant from the highland of Peru, reduced ventral prostate size in normal and TE treated rats. PMID- 15661085 TI - Clinical features and diagnosis of multiple sclerosis. PMID- 15661082 TI - Molecular mechanisms of severe acute respiratory syndrome (SARS). AB - Severe acute respiratory syndrome (SARS) is a new infectious disease caused by a novel coronavirus that leads to deleterious pulmonary pathological features. Due to its high morbidity and mortality and widespread occurrence, SARS has evolved as an important respiratory disease which may be encountered everywhere in the world. The virus was identified as the causative agent of SARS due to the efforts of a WHO-led laboratory network. The potential mutability of the SARS-CoV genome may lead to new SARS outbreaks and several regions of the viral genomes open reading frames have been identified which may contribute to the severe virulence of the virus. With regard to the pathogenesis of SARS, several mechanisms involving both direct effects on target cells and indirect effects via the immune system may exist. Vaccination would offer the most attractive approach to prevent new epidemics of SARS, but the development of vaccines is difficult due to missing data on the role of immune system-virus interactions and the potential mutability of the virus. Even in a situation of no new infections, SARS remains a major health hazard, as new epidemics may arise. Therefore, further experimental and clinical research is required to control the disease. PMID- 15661086 TI - Natural history of multiple sclerosis. PMID- 15661083 TI - Extragonadal aromatization increases with time after ovariectomy in rats. AB - BACKGROUND: The circulating estrogen concentration elevated gradually along with time after ovariectomy in rats. To explore the source of the increased circulation estrogen, the extragonadal aromatization as well as the synthesis of androgen in the adrenal cortex of the ovariectomized rats was evaluated. METHODS: Female rats were divided into twelve groups: 1 month after ovariectomy (OVX1M), OVX2M, OVX3M, OVX4M, OVX5M, OVX6M; intact 1 month (INT1M), INT2M, INT3M, INT4M, INT5M, INT6M. The blood concentration of testosterone (T) was measured by radioimmunoassay. The mRNA expressions of P450 aromatase in the liver and subcutaneous abdominal (SA) adipose as well as the adrenal cytochrome P450 17 alpha hydroxylase/lyase (P450c17) were semiquantified by RT-PCR. The P450 aromatase protein expressions in the liver and SA adipose were detected by Western blot. RESULTS: The blood E2 concentrations increased gradually along with time after ovariectomy in the rats. The 58-kDa aromatase protein and mRNA expressions normalized to beta-actin in the OVX6M rats' SA adipose tissues showed higher levels than those from corresponding tissues in the INT6M (p < 0.05). And the ratios of aromatase mRNA and protein to beta-actin in the OVX6M rats' liver tissues increased significantly compared with those in the OVX1M rats (p < 0.05). The ratio of adrenal P450c17 to beta-actin in the OVX6M increased markedly, and was higher than OVX1M (p < 0.05), though the blood concentration of T decreased significantly in all the ovariectomized rats (p < 0.05). CONCLUSION: Both the subcutaneous abdominal adipose tissues and the liver tissues contributed to the extragonadal aromatisation to promote the circulating E2 levels in the rats along with time after ovariectomy; the adrenal compensation might also be activated naturally. PMID- 15661087 TI - Gender issues. PMID- 15661088 TI - Genetics of multiple sclerosis. PMID- 15661089 TI - The pathology of multiple sclerosis. PMID- 15661090 TI - Neuropathobiology of multiple sclerosis. PMID- 15661091 TI - Neuroimaging in multiple sclerosis. PMID- 15661092 TI - Immunology of multiple sclerosis. PMID- 15661093 TI - Managing symptoms of multiple sclerosis. PMID- 15661094 TI - Interferons in multiple sclerosis. PMID- 15661095 TI - Glatiramer acetate in the treatment of multiple sclerosis. PMID- 15661096 TI - Natalizumab and other monoclonal antibodies. PMID- 15661097 TI - Immunosuppressive therapy for multiple sclerosis. PMID- 15661098 TI - Immunoablative therapy as a treatment aggressive multiple sclerosis. PMID- 15661099 TI - One patient, two carpal tunnels: statistical and clinical analysis--by hand or by patient? PMID- 15661100 TI - Motor cortex dysfunction revealed by cortical excitability studies in Parkinson's disease: influence of antiparkinsonian treatment and cortical stimulation. AB - Single or paired pulse paradigms of transcranial magnetic stimulation (TMS) provide several parameters to test motor cortex excitability, such as motor threshold (MT), motor evoked potential (MEP) amplitude, electromyographic silent period to cortical stimulation (CSP) and intracortical facilitation (ICF) or inhibition (ICI). Various changes in TMS parameters, revealing motor cortex dysfunction, were found in patients with Parkinson's disease (PD). For instance, low MT and increased MEP size disclosed an enhanced corticospinal motor output at rest, while reduced ICF and failure of MEP size increase during contraction suggested defective facilitatory cortical inputs, particularly for movement execution. Inhibitory cortical pathways were also found less excitable at rest (reduced ICI) and sometimes during contraction (shortened CSP). By restoring cortical inhibition, dopaminergic drugs and deep brain stimulation probably overcome the difficulty to focus neuronal activity onto the appropriate network required for a specific motor task. The application of repetitive TMS trains over motor cortical areas also showed some effect on cortical excitability, opening perspectives to consider the motor cortex as a target for therapeutic neuromodulation in PD. However, systematic studies of cortical excitability remained to be performed in large series of patients with PD, taking into account disease stage, clinical symptoms and medication influence. PMID- 15661101 TI - Strength-duration properties and glycemic control in human diabetic motor nerves. AB - OBJECTIVE: To investigate the influences of hyperglycemia on axonal excitability in human diabetic nerves. Hyperglycemia results in decreased Na+-K+ pump function, presumably leading to intra-axonal Na+ accumulation and thereby, reduced Na+ currents. METHODS: The strength-duration time constant (tau(SD)), which partly depends on persistent Na+ conductance active at the resting membrane potential, was measured in median motor axons of 79 diabetic patients. The relationship of tau(SD) with the state of glycemic control (hemoglobin A1c [HbA1c] levels) was analyzed. RESULTS: The mean tau(SD) was longer for diabetic patients than for normal controls, but the difference was not significant. Among diabetic patients, the subgroup of patients with good glycemic control (HbA1c<7%) had significantly longer tau(SD) than the patient group with poor control (HbA1c>9%; P=0.04). The mean tau(SD) was longest at the HbA1c level of 5-6%, gradually decreasing and reaching a plateau around the HbA1c level of 9%. There was an inverse relationship between HbA1c levels and tau(SD), when the HbA1c levels ranged from 5 to 9% (P=0.04). CONCLUSIONS: In diabetic nerves, tau(SD) is generally longer than normal, but hyperglycemia is associated with paradoxically shortened tau(SD), because of a decrease in axonal persistent Na+ conductance, possibly related to reduced membranous Na+ gradient, tissue acidosis, or other metabolic factors. SIGNIFICANCE: Measurements of tau(SD) could provide a new insight into changes in ionic conductance in human diabetic nerves. PMID- 15661102 TI - Frequency of neurogenic thoracic outlet syndrome in patients with definite carpal tunnel syndrome: an electrophysiological evaluation in 100 women. AB - OBJECTIVE: The aim of this work was to evaluate the frequency of lower brachial plexus lesions as true neurogenic thoracic outlet syndrome (NTOS) in women with carpal tunnel syndrome (CTS). METHODS: This was a prospective and descriptive study. The CTS was clinically and electrodiagnostically defined in each patient. The conduction of ulnar and medial antebrachial cutaneous nerves (MABCN) was bilaterally studied to evaluate the function of lower brachial plexus. One hundred women with 176 median nerve lesions at wrist were studied. Patients with ulnar nerve lesion at elbow or wrist were excluded. The sensory nerve action potential (SNAP) of ulnar nerve was abnormal when the interside amplitude ratio was greater than 1.66 or when its amplitude was smaller than 8 microV. The SNAP of MABCN was abnormal when the interside amplitude ratio was greater than 1.66 with both techniques (antidromic and orthodromic) or when its amplitude was smaller than 8 and 6 microV for non-obese patients, respectively, less and more than 60 years old. RESULTS: The mean SNAP amplitude of ulnar and MABC nerves was normal compared with control subjects, and none of the 100 women with CTS had an abnormal interside amplitude ratio for one or both nerves. In 7-10 cases, all of which were either obese or elderly patients, SNAP amplitude of MABCN was decreased, but the interside amplitude ratio remained normal. CONCLUSIONS: On the basis of tests considered quite sensitive (70%) and specific (100%), the occurrence of true NTOS is lower than 1/100 in woman with definite CTS. SIGNIFICANCE: These results demonstrate that there is no appreciable link between CTS and true NTOS. There is no need for the systematic MABCN testing had in CTS patients. PMID- 15661103 TI - Comparison of automated and manual F-wave latency measurements. AB - OBJECTIVE: F-waves are well-established clinical neurophysiological studies. F wave analysis is now cumbersome limiting the usefulness of F-waves. This study evaluates the accuracy and reliability of an automated analysis method for F-wave latencies. METHODS: F-waves following 20 supramaximal stimuli recorded from the extensor digitorum brevis muscle of 80 limbs (55 subjects) were analyzed. F-wave latencies were determined using a computer program developed by NEUROMetrix (Waltham, MA). These results were compared in a blinded fashion with manual measurements of the same datasets by a clinical neurophysiologist with established expertise in F-waves. The manual measurements were repeated once. RESULTS: The yield rate of automated median F-wave latencies was 100% with a correlation coefficient (CC) of 0.996 when compared with manual assignment results. For individual F-wave latency measurements, comparable values were 90% and 0.977, respectively. The repeated manual measurements revealed a yield rate and CC for median latencies of 100% and 0.998, respectively, with comparable values for individual latency measurements of 95% and 0.992. CONCLUSIONS: These results indicate the feasibility of a reliable computerized automated analysis of F-wave latencies. SIGNIFICANCE: A reliable automated analysis of F-waves should add meaningfully to the value of these responses in clinical neurophysiology. PMID- 15661104 TI - The acute effects of glycemic control on nerve conduction in human diabetics. AB - OBJECTIVE: To investigate acute changes in nerve conduction associated with glycemic control. In diabetes, nerve dysfunction can result from reversible metabolic factors associated with hyperglycemia, as well as structural changes. METHODS: Multiple nerve conduction parameters including F-wave latencies were measured in 47 diabetic patients with prominent hyperglycemia before and after intensive insulin treatment. RESULTS: Four weeks after the start of treatment, there was a significant improvement in minimal F-wave latencies of the median (P<0.001) and tibial (P<0.001) nerves, and in distal latencies (P=0.01) and sensory nerve conduction velocities (P<0.001) of the median nerves. Amplitudes of motor and sensory responses did not change significantly. These findings were similar for patients with type 1 (n=8) and those with type 2 (n=39) diabetes. Patients with poorer glycemic control or milder neuropathy tended to show greater changes after treatment. CONCLUSIONS: Glycemic control quickly alters the speed of nerve conduction. F-wave latencies and conduction times across the carpal tunnel are very sensitive parameters. SIGNIFICANCE: Serial nerve conduction studies can detect reversible slowing of nerve conduction presumably caused by metabolic factors, such as decreased Na+/K+-ATPase activity, the altered polyol pathway, and tissue acidosis. PMID- 15661105 TI - What symptoms are truly caused by median nerve compression in carpal tunnel syndrome? AB - OBJECTIVE: This study aims to identify the symptoms, signs and distributions which are associated with neurophysiological carpal tunnel syndrome (N-CTS), defined by the finding of a median nerve lesion at the wrist through neurophysiologic studies, and to compare them with those of patients with sensory or motor complaints in the upper limbs whose electrophysiologic investigation did not show evidence of this syndrome. METHODS: A cross-sectional study was performed, with prospective gathering of data, following a predetermined protocol. We included all patients older than 12 years who were referred for nerve conduction studies and electromyography of at least one of the upper limbs between August 2001 and January 2003. The patients answered a clinicoepidemiologic questionnaire and painted the areas in which they felt pain and those in which they felt paresthesia, and were also examined to test for classical signs CTS. RESULTS: A neurophysiological diagnosis of CTS was reached in 1549 upper limbs (39%). Approximately 6% of the upper limbs with N-CTS and 16% of those without median nerve lesion do not have any hand or wrist symptoms (P<0.001). Paresthesia, pain, hand weakness and cramps were the symptoms statistically associated to N-CTS, with paresthesia being the one with the greatest power of association. This is usually felt in median innervation's territory, and is frequently extended to the whole hand, but without association with N-CTS when felt in proximal region. CONCLUSIONS: One concludes that the most characteristic manifestation of carpal tunnel syndrome (CTS) is paresthesia. Pain also occurs very frequently but it is less specific, and weakness, meanwhile, seems to have a low frequency. We believe that the variability in the clinical presentation of CTS is largely due to the presence of associated diseases and our results provide information which could help to better define the clinical criteria used in the diagnosis of this syndrome. PMID- 15661106 TI - The effects of mexiletine on excitability properties of human median motor axons. AB - OBJECTIVE: To investigate the effects of mexiletine, an analog of lidocaine, on excitability of human axons in vivo. METHODS: Threshold tracking was used to measure multiple excitability indices (strength-duration time constant, rheobase, refractoriness, supernormality, and threshold electrotonus) in median motor axons of 20 patients with neuropathic pain or muscle cramping, before and 3 months after treatment with oral 300 mg mexiletine per day. RESULTS: After treatment, there was a reduction in pain/muscle cramps, associated with decreased strength duration time constants (P=0.01), increased rheobasic currents (P=0.06), and lower refractoriness (P=0.02), all of which were consistent with reduced nodal Na+ currents. Supernormality and threshold electrotonus did not change significantly. The changes in strength-duration properties suggest a decrease in persistent Na+ conductance. The lowered refractoriness after treatment might result from reduced transient Na+ currents, but the lack of change in supernormality and threshold electrotonus was not consistent with this hypothesis. CONCLUSIONS: Oral mexiletine in a dosage of 300 mg daily suppresses persistent Na+ currents in human motor axons. SIGNIFICANCE: Measurements of the excitability indices can be used for non-invasive assessment and monitoring of the effects of mexiletine in patients with neuropathic pain or muscle cramps. PMID- 15661107 TI - Electrode position and size in electrical impedance myography. AB - OBJECTIVE: Linear-electrical impedance myography (EIM) is a non-invasive technique for the evaluation of muscle, in which high-frequency alternating current is injected into the body via two surface electrodes, and the resulting voltage pattern over a selected muscle is measured using a second, larger set of electrodes. The precise location and size of the electrodes can be critical to the data obtained, and in this study the effects of variation in these factors were evaluated. METHODS: Linear-EIM was performed in 5 subjects while varying the location of the current injecting electrodes and in an additional 8 subjects while varying the position of the voltage electrodes. RESULTS: The major outcome variable, the 'spatially averaged phase' (theta(avg)), decreased as the ipsilateral current injecting electrode was moved farther from the voltage electrodes, reaching a plateau 15-20 cm distant. As for the voltage electrode array, distal-proximal shifts resulted in the greatest changes, with variation in theta(avg) being as high as 14% per cm; circumferential shifts around the limb had more modest effects. CONCLUSIONS: Linear-EIM results depend systematically on current and voltage electrode positions, but with reasonable care variation can be minimized. SIGNIFICANCE: With proper attention to electrode placement, linear EIM has sufficient reproducibility to become an important clinical tool in neuromuscular disease evaluation. PMID- 15661108 TI - Ipsi- and contralateral exteroceptive EMG modulation in uni- and bilaterally activated thenar muscles. AB - OBJECTIVE: The cutaneous silent period (CSP) is a spinal inhibitory reflex mediated by A-delta fibers. The exact underlying neural pathway, however, is unknown. This study was undertaken to investigate whether the neural circuitry mediating CSPs is wired unilaterally or whether there is evidence of influence from or upon the contralateral side. METHODS: Fifteen healthy subjects underwent bilateral CSP testing following unilateral nociceptive digit II stimulation. Surface electromyographic (EMG) recordings were obtained bilaterally from thenar muscles following unilateral recurrent nociceptive digit II stimulation while activating the ipsilateral or the contralateral or both thenar muscles against resistance. RESULTS: Nociceptive digit II stimulation evoked consistent CSPs in ipsilateral thenar muscles during voluntary contraction, while there was no consistent influence on EMG activity in contralateral thenar muscles at rest. Furthermore, nociceptive digit II stimulation did neither consistently affect EMG activity in ipsilateral thenar muscles at rest nor in contralateral thenar muscles during voluntary contraction. Finally, there was no significant difference between any CSP parameters obtained during unilateral versus bilateral muscle contraction. Occasional late excitatory EMG activity in relaxed or contracted thenar muscles resembled startle reflexes, which seem to contribute to the post-inhibition EMG-rebound. CONCLUSIONS: The present findings are consistent with unilateral wiring of the spinal circuitry mediating CSPs. SIGNIFICANCE: The essential lack of a crossed inhibitory influence of nociceptive digit II stimulation underscores the utility of CSP testing in the assessment of the A delta fiber system in healthy subjects and patients with various pathologies. PMID- 15661110 TI - Role of cerebral cortex in human postural control: an EEG study. AB - OBJECTIVE: It was our primary objective to provide evidence supporting the existence of neural detectors for postural instability that could trigger the compensatory adjustments to avoid falls. METHODS: Twelve young healthy subjects performed self-initiated oscillatory and discrete postural movements in the anterior-posterior (AP) directions with maximal range of motion predominantly at ankle joint. Movements were recorded by the system and included force plate and EMG, and EEG measures from 25 electrode sites. The center of pressure dynamics and stability index were calculated, and EEG potentials both in voltage and frequency domains were extracted by averaging and Morlet wavelet techniques, respectively. RESULTS: The initiation of self-paced postural movement was preceded by slow negative DC shift, similar to movement-related cortical potentials (MRCP) accompanying voluntary limb movement. A burst of gamma activity preceded the initiation of compensatory backward postural movement when balance was in danger. This was evident for both oscillatory and discrete AP postural movements. The spatial distribution of EEG patterns in postural actions approximated that previously observed during the postural perceptual tasks. CONCLUSIONS: The results suggest an important role of the higher cortical structures in regulation of posture equilibrium in dynamic stances. Postural reactions to prevent falls may be triggered by central command mechanisms identified by a burst of EEG gamma activity. SIGNIFICANCE: The results from this study contribute to our understanding of neurophysiological mechanisms underlying the cortical control of human upright posture in normal subjects. PMID- 15661109 TI - Caffeine has no effect on measures of cortical excitability. AB - OBJECTIVE: To assess the effect of caffeine on motor thresholds, short interval intra-cortical inhibition (SICI), intra-cortical facilitation (ICF) and cortical silent periods in a placebo controlled double-blinded trial. METHODS: In eleven healthy non-smoking subjects the following parameters were measured using transcranial magnetic stimulation (TMS): motor thresholds (rest, RMT and active, AMT), SICI and ICF at different conditioning stimulus intensities (60, 70, 80, 90% AMT), cortical silent periods at 130, 150 and 175% AMT, and size of motor evoked potential at rest at 110, 125 and 150% RMT. Measurements were repeated after one cup of decaffeinated coffee. On another day, measurements were obtained before and after one cup of decaffeinated coffee that contained caffeine (3 mg/kg bodyweight). Caffeine concentrations were measured in serum before and after experiments. Experiments were conducted and data were evaluated blinded to the experimental condition. RESULTS: The results of repeated measurements of all parameters were similar comparing experiments on each day, or when comparing the caffeine arm of the study with the placebo arm. CONCLUSIONS: Caffeine in a concentration similar to that in a strong cup of coffee does not have a major effect on TMS measures of motor cortex excitability. SIGNIFICANCE: In healthy controls, the design of TMS experiments that investigate the parameters assessed in this TMS study does not need to control for caffeine. PMID- 15661111 TI - Stimulant drug action in attention deficit hyperactivity disorder (ADHD): inference of neurophysiological mechanisms via quantitative modelling. AB - OBJECTIVE: To infer the neural mechanisms underlying tonic transitions in the electroencephalogram (EEG) in 11 adolescents diagnosed with attention deficit hyperactivity disorder (ADHD) before and after treatment with stimulant medication. METHODS: A biophysical model was used to analyse electroencephalographic (EEG) measures of tonic brain activity at multiple scalp sites before and after treatment with medication. RESULTS: It was observed that stimulants had the affect of significantly reducing the parameter controlling activation in the intrathalamic pathway involving the thalamic reticular nucleus (TRN) and the parameter controlling excitatory cortical activity. The effect of stimulant medication was also found to be preferentially localized within subcortical nuclei projecting towards frontal and central scalp sites. CONCLUSIONS: It is suggested that the action of stimulant medication occurs via suppression of the locus coeruleus, which in turn reduces stimulation of the TRN, and improves cortical arousal. The effects localized to frontal and central sites are consistent with the occurrence of frontal delta-theta EEG abnormalities in ADHD, and existing theories of hypoarousal. SIGNIFICANCE: To our knowledge, this is the first study where a detailed biophysical model of the brain has been used to estimate changes in neurophysiological parameters underlying the effects of stimulant medication in ADHD. PMID- 15661112 TI - The 'F-complex' and MMN tap different aspects of deviance. AB - OBJECTIVE: To compare the 'F(fusion)-complex' with the Mismatch negativity (MMN), both components associated with automatic detection of changes in the acoustic stimulus flow. METHODS: Ten right-handed adult native Hebrew speakers discriminated vowel-consonant-vowel (V-C-V) sequences /ada/ (deviant) and /aga/ (standard) in an active auditory 'Oddball' task, and the brain potentials associated with performance of the task were recorded from 21 electrodes. Stimuli were generated by fusing the acoustic elements of the V-C-V sequences as follows: base was always presented in front of the subject, and formant transitions were presented to the front, left or right in a virtual reality room. An illusion of a lateralized echo (duplex sensation) accompanied base fusion with the lateralized formant locations. Source current density estimates were derived for the net response to the fusion of the speech elements (F-complex) and for the MMN, using low-resolution electromagnetic tomography (LORETA). Statistical non-parametric mapping was used to estimate the current density differences between the brain sources of the F-complex and the MMN. RESULTS: Occipito-parietal regions and prefrontal regions were associated with the F-complex in all formant locations, whereas the vicinity of the supratemporal plane was bilaterally associated with the MMN, but only in case of front-fusion (no duplex effect). CONCLUSIONS: MMN is sensitive to the novelty of the auditory object in relation to other stimuli in a sequence, whereas the F-complex is sensitive to the acoustic features of the auditory object and reflects a process of matching them with target categories. SIGNIFICANCE: The F-complex and MMN reflect different aspects of auditory processing in a stimulus-rich and changing environment: content analysis of the stimulus and novelty detection, respectively. PMID- 15661113 TI - Deviance-related electrophysiological activity in mice: is there mismatch negativity in mice? AB - OBJECTIVE: Mismatch negativity (MMN) is an auditory event-related potential (ERP) that provides an index of auditory sensory memory and has become an important tool to investigate auditory sensory memory in cognitive neuroscience and disorders such as schizophrenia and dyslexia. The development of a mouse model of human MMN would permit to investigate the molecular biology of normal and dysfunctional MMN generation. However, the presence of MMN-like electrophysiological activity in mice has not been demonstrated. METHODS: Deviance-related ERPs were recorded in awake mice using 3 frequency deviance paradigms and one duration deviance paradigm. These paradigms were modelled after paradigms used in human studies to characterize MMN. RESULTS: Significant deviance-related activity was observed in all paradigms. However, in all frequency deviance paradigms this activity manifested as an enhancement of similar activity to the standard due to differences in stimulation rate between deviant and standard stimuli rather than qualitatively different MMN-like activity. In the duration deviance paradigm negative deflections were observed that showed characteristics typical of human MMN. CONCLUSIONS: MMN-like activity can be observed in mice in duration deviance paradigms. In frequency deviance paradigms effects of different stimulation rates of deviant and standard stimuli seem to be the main determinants of deviance-related activity. SIGNIFICANCE: Investigations of MMN-like ERPs in mice may permit to investigate the molecular basis for normal and abnormal MMN generation in neuropsychiatric disorders and dyslexia. PMID- 15661114 TI - Resting SPECT-neuropsychology correlation in very mild Alzheimer's disease. AB - OBJECTIVE: To investigate the relationships between brain function and some of the most frequently impaired cognitive domains in the first stages of Alzheimer's disease (AD), we searched for correlation between the scores on 3 neuropsychological tests and brain perfusion, assessed by single photon emission computed tomography (SPECT) in patients with very mild AD. METHODS: Twenty-nine consecutive outpatients (mean age 78.2+/-5.5) affected by probable AD in the very mild phase (i.e. with a score > or =20 on the mini-mental state examination, MMSE) underwent brain SPECT with (99m)Tc-ethylcisteinate dimer. For correlative purposes, word list learning (by the selective reminding test, SRT), constructional praxis test (CPT) and visual search test (VST) were chosen a priori out of an extended battery employed to diagnose AD at first patient evaluation. Voxel-based correlation analysis was achieved by statistical parametric mapping (SPM99) with a height threshold of P=0.005. Age, years of education and the MMSE score were inserted in the correlative analysis as confounding variables. RESULTS: The SRT score showed correlation with brain perfusion in 3 clusters of the left hemisphere, including the post-central gyrus, the parietal precuneus, the inferior parietal lobule and the middle temporal gyrus, and in one cluster in the right hemisphere including the middle temporal gyrus and the middle occipital gyrus. The CPT score was significantly correlated with brain perfusion in the parietal precuneus and the posterior cingulate gyrus in the left hemisphere, whereas the VST score gave a significant correlation with brain perfusion in a left cluster including the parietal precuneus and the superior temporal gyrus. CONCLUSIONS: Cognitive impairment in very mild AD is reflected by brain dysfunction in posterior associative areas, with peculiar topographical differences proper of each domain. The parietal precuneus was a common site of correlation of all 3 neuropsychological tests. This region, together with the posterior cingulate and the superficial posterior temporal parietal cortex, is thought to be affected by disconnection from the mesial temporal lobe, besides being directly affected by increased oxidative stress and by atrophy as well. The impairment of these areas is thought to contribute to cognitive decline in verbal memory, constructional praxis and visual sustained attention which are indeed among the earliest signs of cognitive impairment in AD. SIGNIFICANCE: Assessing the relationships between neuropsychology and brain functional imaging is a key approach to clarify the pathophysiology of cognitive failure in AD; the specificity of these findings in AD remains to be proven through comparison with correlation achieved in matched controls. PMID- 15661115 TI - Chronic effects of social drinking in a card-sorting task: an event related potential study. AB - OBJECTIVE: The Wisconsin Card Sorting Task (WCST) is one of the most widely used neuropsychological tests of frontal lobe function, which is thought to be affected by regular alcohol use. The present study used a computer-adapted version of the WCST to assess the effects of chronic alcohol consumption on the brain. METHODS: Participants (N=59) sorted cards according to an initially unknown sorting rule, which referred to shape, number, or color. The correctness of the chosen sorting rule was indicated by a feedback stimulus. This correct sorting rule had to be followed for a number of stimuli, and when it changed participants had to find out which rule had to be followed next. A distinction was made between early (correct sorting rule is unknown) and late trials (correct sorting rule is known and applied). To measure brain activity related during the task event related potentials (ERPs) were recorded to the target and feedback stimulus in light (N=14), moderate (N=16) and heavy (N=19) social drinkers and excessive alcohol users (N=10). RESULTS: No differences in number of series completed or the reaction time in each trial, were found between the four groups. In contrast, a mid-frontal N1 component in reaction to the feedback stimuli did reveal differences between the four groups. In the light and moderate drinkers, on early feedback trials the N1 was larger relative to late feedback trials, but this effect was absent in the heavy social drinkers and excessive drinkers. CONCLUSIONS: The reduced N1 effect with increasing alcohol intake could reflect abnormal allocation of attention or impaired conflict monitoring, possibly based on activity in the anterior cingulate cortex. SIGNIFICANCE: Heavy social drinking and excessive drinking leads to changes in the mid-frontal N1 during feedback trials of the WCST. PMID- 15661116 TI - Effect of electroconvulsive therapy on cortical excitability in patients with major depression: a transcranial magnetic stimulation study. AB - OBJECTIVE: The antidepressant action of electro-convulsive therapy (ECT) and repetitive transcranial magnetic stimulation (rTMS) may be related to their ability to modulate cortical excitability. The aim of this study was to investigate changes in cortical excitability following ECT in patients with major depression (MD) and to compare therapeutic efficacy of ECT combined with rTMS to that of ECT alone. METHODS: Twenty-two patients with MD were assigned to receive ECT and right prefrontal 1 Hz rTMS (n=12) or ECT with sham rTMS (n=10). ECT was given twice weekly and rTMS was applied on the remaining 4 days, throughout 3 weeks. The resting motor threshold (rMT) and motor evoked potential (MEP)/M-wave area ratio were evaluated before and 6 h after the first, third and sixth ECT session. The active motor threshold (aMT), intra-cortical inhibition (ICI) and intra-cortical facilitation (ICF) were measured at baseline and 24 h after the last ECT. RESULTS: There were no significant differences in the degree of clinical improvement and measures of cortical excitability in the ECT+active rTMS group as compared to the ECT+sham rTMS group. Marked clinical improvement observed in 19 out of the 22 patients was associated with a significant increase of the MEP/M-wave area ratio, decrease of the aMT and reduction of the ICI in the left hemisphere. CONCLUSIONS: The antidepressant effect of ECT was associated with an enhancement of left hemispheric excitability. rTMS did not add to the beneficial effect of ECT. However, the small sample size and the robust effect of ECT might have obscured a potential therapeutic effect of rTMS. SIGNIFICANCE: Measures of cortical excitability may provide insight to our understanding of the mechanism of action of ECT and might be useful for the assessment of treatment response. PMID- 15661117 TI - Left hemisphere specialization for rapid temporal processing: a study with auditory 40 Hz steady-state responses. AB - OBJECTIVE: To investigate rapid temporal processing in the auditory cortex by using auditory 40 Hz steady-state responses (SSRs). METHODS: A 40 Hz tone-burst at 500 Hz spectral frequency was presented monaurally to record SSRs in 10 normal subjects. The recording electrodes were placed over C1, C2, C3, C4, C5, C6, T3, T4, Fz, Cz and Pz, referring to an electrode at the 7th cervical spinous process. For comparison, unstimulated SSRs were recorded. A total of 200 responses of 1s epoch were averaged and subjected to discrete fast Fourier transforms to yield the amplitude and phase of the 40 Hz component. The coherence (Coh) values of the 40 Hz component between homologous electrodes were also calculated. RESULTS: At the temporal electrodes contralateral to the stimulated ear, the amplitude was significantly larger and its phase was significantly smaller than those of the ipsilateral side. The interhemispheric Coh between T3 and T4 in response to right ear stimulation was significantly greater than those of left ear stimulation or the unstimulated condition. CONCLUSIONS: Our results suggest that 40 Hz auditory information is predominantly processed in the left auditory cortex, interacting with the right hemisphere. This finding is consistent with the fact that the left auditory cortex plays an important role in rapid temporal processing. SIGNIFICANCE: Auditory 40 Hz SSRs with Coh analysis are useful for investigating the left hemisphere specialization for rapid temporal processing. PMID- 15661118 TI - Sound-evoked neurogenic responses with short latency of vestibular origin. AB - OBJECTIVE: In ABR recording, a large negative deflection with a latency of 3 ms (N3) has been recorded in patients with peripheral profound deafness. It has been suggested that N3 might be of vestibular origin. So far, N3 has been recorded only in patients with peripheral profound deafness. If we can record N3 potentials in subjects with preserved hearing, recording N3 potentials might be a new clinical test of the vestibular system. To record neurogenic potentials (N3) of vestibular origin in healthy volunteers and patients with vestibular disorders. METHODS: Twelve healthy volunteers (10 men and two women, aged 23-37 years) and 12 patients with vestibular disorders (6 men and 6 women, aged 29-71 years) were enrolled in this study. To record responses, surface electrodes were placed on the ipsilateral mastoid and the vertex. An electrode on the nasion served as the ground. Recording was performed using an auditory evoked potential recording system with a mini-mixer and a stereo-amplifier. Signals at the vertex to the ispilateral mastoid were amplified and bandpass filtered (100-3000 Hz). One thousand-hertz short tone bursts (1 kHz STB; rise/fall time=0.5 ms, plateau time=1 ms) were presented to either ear through a headphone with or without white noise (WN) ipsilateral to the stimulated ear. The stimulation rate was 10 Hz, and the analysis time was 10 ms. The responses to 500 stimuli were averaged twice. RESULTS: When 1 kHz STB (95 dBnHL, equivalent to 130 dBSPL) were presented with 100 dBSPL WN (ipsilateral to the stimulated ear), a negative peak with 3-4 ms latency (N3) was observed in 23 of the 24 ears (95.8%) with reproducibility in healthy subjects. Without WN, N3 was observed in 17 of the 24 ears (70.8%). The threshold of N3 was 90.2 dBnHL on the average. The presence of N3 in the patients was in agreement with the presence of the VEMP, which were also recorded. CONCLUSIONS: Using techniques of WN exposure ipsilateral to the stimulated ear, we recorded N3 in healthy subjects and in vestibular disorder patients with preserved hearing. This negative peak is likely to be of vestibular origin. SIGNIFICANCE: N3 may be measured from subjects who cannot contract neck muscles due to their ages, mental states, or consciousness disorders. In other words, N3 may be measured from subjects from whom VEMP cannot be recorded. In combination with VEMP, N3 may be useful for the detection of lesion sites. PMID- 15661119 TI - General vestibular testing. AB - A dysfunction of the vestibular system is commonly characterized by a combination of phenomena involving perceptual, ocular motor, postural, and autonomic manifestations: vertigo/dizziness, nystagmus, ataxia, and nausea. These 4 manifestations correlate with different aspects of vestibular function and emanate from different sites within the central nervous system. The diagnosis of vestibular syndromes always requires interdisciplinary thinking. A detailed history allows early differentiation into 9 categories that serve as a practical guide for differential diagnosis: (1) dizziness and lightheadedness; (2) single or recurrent attacks of vertigo; (3) sustained vertigo; (4) positional/positioning vertigo; (5) oscillopsia; (6) vertigo associated with auditory dysfunction; (7) vertigo associated with brainstem or cerebellar symptoms; (8) vertigo associated with headache; and (9) dizziness or to-and-fro vertigo with postural imbalance. A careful and systematic neuro-ophthalmological and neuro-otological examination is also mandatory, especially to differentiate between central and peripheral vestibular disorders. Important signs are nystagmus, ocular tilt reaction, other central or peripheral ocular motor dysfunctions, or a unilateral or bilateral peripheral vestibular deficit. This deficit can be easily detected by the head-impulse test, the most relevant bedside test for the vestibulo-ocular reflex. Laboratory examinations are used to measure eye movements, to test semicircular canal, otolith, and spatial perceptional function and to determine postural control. It must, however, be kept in mind that all signs and ocular motor and vestibular findings have to be interpreted within the context of the patient's history and a complete neurological examination. PMID- 15661120 TI - A system to detect the onset of epileptic seizures in scalp EEG. AB - OBJECTIVE: A new method for automatic seizure detection and onset warning is proposed. The system is based on determining the seizure probability of a section of EEG. Operation features a user-tuneable threshold to exploit the trade-off between sensitivity and detection delay and an acceptable false detection rate. METHODS: The system was designed using 652 h of scalp EEG, including 126 seizures in 28 patients. Wavelet decomposition, feature extraction and data segmentation were employed to compute the a priori probabilities required for the Bayesian formulation used in training, testing and operation. RESULTS: Results based on the analysis of separate testing data (360 h of scalp EEG, including 69 seizures in 16 patients) initially show a sensitivity of 77.9%, a false detection rate of 0.86/h and a median detection delay of 9.8 s. Results after use of the tuning mechanism show a sensitivity of 76.0%, a false detection rate of 0.34/h and a median detection delay of 10 s. Missed seizures are characterized mainly by subtle or focal activity, mixed frequencies, short duration or some combination of these traits. False detections are mainly caused by short bursts of rhythmic activity, rapid eye blinking and EMG artifact caused by chewing. Evaluation of the traditional seizure detection method of using both data sets shows a sensitivity of 50.1%, a false detection rate of 0.5/h and a median detection delay of 14.3 s. CONCLUSIONS: The system performed well enough to be considered for use within a clinical setting. In patients having an unacceptable level of false detection, the tuning mechanism provided an important reduction in false detections with minimal loss of detection sensitivity and detection delay. SIGNIFICANCE: During prolonged EEG monitoring of epileptic patients, the continuous recording may be marked where seizures are likely to have taken place. Several methods of automatic seizure detection exist, but few can operate as an on-line seizure alert system. We propose a seizure detection system that can alert medical staff to the onset of a seizure and hence improve clinical diagnosis. PMID- 15661121 TI - A method to identify reproducible subsets of co-activated structures during interictal spikes. Application to intracerebral EEG in temporal lobe epilepsy. AB - OBJECTIVE: We present a novel quantitative method to statistically analyze the distribution of multichannel intracerebral interictal spikes (multi-IIS) in stereoelectroencephalographic (SEEG) recordings. The method automatically extracts groups of brain structures conjointly and frequently involved in the generation of interictal activity. These groups are referred to as 'subsets of co activated structures' (SCAS). We applied the method to long duration interictal recordings in patients with mesial temporal lobe epilepsy (MTLE) and analyzed the reproducibility of subsets of structures involved in the generation of multi-IIS for each patient and among patients. METHODS: Fifteen patients underwent long term intracerebral EEG recording (SEEG technique) using depth electrodes. A 1 h period of continuous interictal EEG recording was selected for each patient with precautions regarding the time after anesthesia pre-SEEG, the temporal distance with respect to seizures, the vigilance state of the patient, and the anti epileptic drug withdrawal. A research of SCAS was conducted on each recording using the developed method that includes 3 steps: (i) automatic detection of monochannel intracerebral interictal spikes (mono-IIS), (ii) formation of multi IIS using a temporal sliding window, and (iii) extraction of SCAS. In the third step, statistical tests are used to evaluate the frequency of multi-IIS as well as their significance (with respect to the 'random distribution of mono-IIS' case). RESULTS: In each patient, several thousands of multi-IIS (mean+/-SD, 3322+/-2190) were formed and several SCAS (mean+/-SD, 3.80+/-1.47) were automatically extracted. Results show that reproducible subsets of brain structures are involved in the generation of interictal activity. Although SCAS were found to be variable from one patient to another, some invariant information was pointed up. In all patients, multi-IIS distribute over two distinct groups of structures: mesial structures (15/15) and lateral structures (7/15). Moreover, two particular structures, the internal temporal pole and the temporo-basal cortex, may be conjointly involved with either the first or the second group. Finally, some extracted SCAS seem to match well-defined anatomo-functional circuits of the temporal lobe. CONCLUSIONS AND SIGNIFICANCE: During interictal activity in MTLE, similar subsets of temporal lobe structures are involved in the generation of spikes. This paper brings statistical evidence for the existence of these subsets and presents a method to automatically extract them from SEEG recordings. Interictal activity is spatially organized in the temporal lobe and preferentially involves two functional systems of the temporal lobe (either mesial or lateral). PMID- 15661122 TI - Estimation of in vivo human brain-to-skull conductivity ratio from simultaneous extra- and intra-cranial electrical potential recordings. AB - OBJECTIVE: The present study aims to accurately estimate the in vivo brain-to skull conductivity ratio by means of cortical imaging technique. Simultaneous extra- and intra-cranial potential recordings induced by subdural current stimulation were analyzed to get the estimation. METHODS: The effective brain-to skull conductivity ratio was estimated in vivo for 5 epilepsy patients. The estimation was performed using multi-channel simultaneously recorded scalp and cortical electrical potentials during subdural electrical stimulation. The cortical imaging technique was used to compute the inverse cortical potential distribution from the scalp recorded potentials using a 3-shell head volume conductor model. The brain-to-skull conductivity ratio, which leads to the most consistent cortical potential estimates with respect to the direct intra-cranial measurements, is considered to be the effective brain-to-skull conductivity ratio. RESULTS: The present estimation provided consistent results in 5 human subjects studied. The in vivo effective brain-to-skull conductivity ratio ranged from 18 to 34 in the 5 epilepsy patients. CONCLUSIONS: The effective brain-to skull conductivity ratio can be estimated from simultaneous intra- and extra cranial potential recordings and the averaged value/standard deviation is 25+/-7. SIGNIFICANCE: The present results provide important experimental data on the brain-to-skull conductivity ratio, which is of significance for accurate brain source localization using piece-wise homogeneous head models. PMID- 15661123 TI - Mind the gap: from rhetoric to action. PMID- 15661124 TI - Health status and the challenges for prevention in The Netherlands. AB - In 2002, the third Public Health Status and Forecasts report was published, containing up-to-date information about Dutch public health and health care. A striking finding was that although life expectancy (LE) in The Netherlands increased between 1980 and 2000, the LE of men is rising less rapidly than the European Union (EU) average. The LE of Dutch women is stagnating and has now fallen below the EU average. These and many other unfavourable trends in the health status of the Dutch population were found to be largely due to unhealthy behaviour. One of the policy recommendations therefore was to strengthen the investments in prevention in order to reverse the stagnation in health status. In response to the findings, the Ministry of Health, Welfare and Sport published the National Prevention Paper. This Paper states that the Ministry, within the existing prevention policy, will pay more attention to healthy behaviour, stressing the responsibility of citizens as well as the societal responsibility of other parties, such as business communities, schools, health insurers and care suppliers. The prevention of specific diseases (diabetes, psychosocial problems, heart diseases, cancer, musculosceletal disorders, asthma and chronic obstructive pulmonary disease) has been given priority status. In this article, we present the major findings regarding the health status of the Dutch population and discuss the implications for prevention policy. PMID- 15661125 TI - Developing a speciality: regearing the specialist public health workforce. AB - OBJECTIVES: To identify issues surrounding the future training needs of the specialist public health workforce following the most recent restructuring of the National Health Service (NHS) in England. METHODS: All directors of public health (DsPH) based in strategic health authorities and nine senior staff working in public health at the regional level were invited to participate in a semi structured telephone interview. RESULTS: Twenty-six people were interviewed. Many interviewees expressed concern that because consultants and specialists in public health will be working in much smaller teams than hitherto, they will have to generalize their skills to cover a much wider range of functions (including board level duties). This may result in a loss of specialist expertise. Successful public health practice in the new structures will require new ways of interorganizational working that will add an administrative burden to specialists in public health. Also, the creation of a board-level post in each primary care trust (PCT) has resulted in more time spent on corporate responsibilities and less on public health for DsPH, who are often the only fully trained specialist in public health in their PCT. Furthermore, interviewees expressed their anxiety about the lack of diversity in the posts available to specialists in public health and particularly to those newly completing their specialist training. Generally, interviewees felt that traditional public health roles and responsibilities were being eclipsed by corporate and managerial ones. Professional development activities were being carried out, but in a rather ad hoc fashion. Interviewees were hopeful that public health networks would lead professional development initiatives once they were more established. CONCLUSIONS: It is important that excellence in public health is maintained through a set of accreditable standards, whilst corporate skills, essential to successful public health practice in the new UK NHS, are developed among specialists in public health. PMID- 15661126 TI - Building postgraduate capacity in medical and public health research in Vietnam: an in-service training model. AB - In Vietnam, needs to update medical and public health research capacity, but advanced training in research methodology in universities is still very limited. The first university-certified research training course aimed at physicians was conducted in Ho Chi Minh City in 2000. This paper describes the processes, outcomes and lessons learned from the project, to enable the training course to be repeated to increase research capacity among health professionals in Vietnam and other developing countries. Based on needs assessment, the 9-month, part-time course offered advanced training in practical research skills, including epidemiology, biostatistics and qualitative research methods. Thirty-seven trainees (teaching staff, interns and clinicians) completed the course, which included four group research projects. With active support from university management, the course was rigorous and participation was excellent. Trainees's knowledge of research methodologies increased, and their self-evaluation of achievement of the training objectives was high. The total score of the epidemiology and biostatistics test improved from 56% (median) in the pre-course test to 83% in the post-course test. The group research project results were disseminated through a conference and publication in peer-reviewed journals. The course has since been repeated twice as a 1-month intensive course to accommodate trainees from distant regions. This type of rigorous in-service research training at universities and teaching hospitals in developing countries appears to be an effective, sustainable approach. It also holds promise for producing research data that are responsive to a country's own needs and which are likely to be applied in health policy and practice. Further evaluation of the impact and cost of such models is indicated. PMID- 15661127 TI - Emerging epidemic in a growing industry: cigarette smoking among female micro electronics workers in Taiwan. AB - OBJECTIVE: To explore the emerging tobacco epidemic in female workers in the growing micro-electronics industry of Taiwan. METHODS: Workers were surveyed regarding their smoking status, sociodemographics and work characteristics. In total, 1950 female employees in two large micro-electronics companies in Taiwan completed the survey. RESULTS: Approximately 9.3% of the female employees were occasional or daily smokers at the time of the survey. The prevalence of smoking was higher in those aged 16-19 years (20.9%), those not married (12.9%), those with a high school education or less (11.7%), those employed by Company A (11.7%), shift workers (14.3%), and those who had been in their present employment for 1 year or less (13.6%). Results of multivariate adjusted logistic regression indicated that younger age, lower level of education, shorter periods of employment with the company and shift working were the important factors in determining cigarette smoking among the study participants. The odds ratio of being a daily smoker was similar to that of being a current smoker. Marital status was the only significant variable when comparing former smokers with current smokers. CONCLUSIONS: Smoking prevalence in female workers in the two micro-electronics companies studied was much higher than previous reports have suggested about female smoking prevalence in Taiwan and China. We suggest that smoking is no longer a 'male problem' in Taiwan. Future smoking cessation and prevention programmes should target young working women as well as men. PMID- 15661128 TI - Social and behavioural determinants of body mass index among adolescent females in Croatia. PMID- 15661129 TI - Health policy analysis: a simple tool for policy makers. AB - Policy analysis is not an easy task. Its scope is broad and can include both the analysis of policy process and the analysis of policy content. This paper is concerned with the analysis of policy content and offers some practical guidance regarding how to analyse health policy and link it to health outcomes. An eight step framework for policy analysis is proposed that public health policy makers and public health practitioners may find especially useful due to its simplicity. PMID- 15661130 TI - Health fair in a mosque: putting policy into practice. AB - BACKGROUND: Towards the end of 2001, the Scottish Executive Health Department published a unique report, 'Fair for All', on the issue of addressing the health needs of ethnic minorities. One of the recommendations contained within the report was that different groups and organizations within communities should work together to provide services to minority groups. This descriptive paper is an illustration of how a settled community, in this case a Muslim community, was encouraged to become involved in its own health care by holding a health fair at its mosque. PLANNING: The health fair was organized by the Minority Ethnic Health Inclusion Project, in collaboration with the Local Health Care Co-operative, and in co-operation with Edinburgh Central Mosque. The health fair was held over two consecutive Fridays in May 2003 and focused primarily on diabetes, high blood pressure, healthy eating and oral hygiene, as well as providing information on cancer and local community services. OUTCOMES: The health fair was well received on both days, with around 200 men and 120 women participating. In total, there were 99 recorded blood pressure measurements and 81 blood glucose checks, and the community dental service was very well received. EVALUATION: Strong participation by the community shows that people from ethnic minorities are interested in their health and will participate in health-related activities arranged for them. The success of this health fair shows that, as proposed by the Scottish Executive, different groups and organizations within the community can work together to provide services to minority ethnic groups. PMID- 15661131 TI - Malaria: knowledge and behaviour in an endemic rural area of Turkey. AB - This cross-sectional study was conducted to determine the knowledge and behaviour of people in the Sanliurfa province of Turkey regarding the prevention of malaria. A modified 30-cluster sampling method based on the traditional Expanded Programme for Immunization coverage surveys was employed to select a representative sample from 210 households. A questionnaire that focused on sociodemographic characteristics, knowledge and behaviour of malaria prevention, treatment-seeking behaviour and the use of antimalarials was applied. Eighty-nine percent of respondents knew at least one of the classical symptoms of malaria, and fever and chills were the most commonly reported symptoms (78.6%). Of the people interviewed, 33% believed that malaria can be acquired from dirty water, by changing place of residence, by working in cotton or tomato fields, or from malaria patients' belongings. None of the respondents knew how mosquitoes acquire the parasite. Twenty-five percent of respondents believed that elimination of breeding sites was one way to prevent malaria, and 8% identified the use of bednets. Fifty-five percent of respondents reported protective behaviours that are not directly associated with malaria transmission. Almost 47% of respondents reported that they completed their antimalarials, and only 21% of respondents indicated that they would seek treatment for febrile disease from physicians or a malaria unit. Understanding community perceptions of aetiology, symptom identification and treatment of malaria is an important step towards disease control. PMID- 15661133 TI - An enquiry into the respiratory health effects on a rural community of a soil mound erected close to residential property. AB - The health concerns of a rural community were investigated following the erection of a soil mound in close proximity to residential property. Retrospective comparisons were made of respiratory and non-respiratory consultations with general practitioners between the exposed population and a sociodemographically similar comparison population. A 2-year period was examined, 1 year before and 1 year after the mound was erected. In the 1-year period prior to erection of the mound, similar consultation rates for both respiratory and non-respiratory conditions were observed in both populations. In the 1-year period following erection of the mound, the exposed population was more likely to consult for respiratory conditions than the comparison population (OR=4.10, 95% CI 2.26 7.44). No differences were observed for non-respiratory conditions. We identified a significant increase in respiratory consultations in the exposed population following erection of the soil mound. Limitations associated with this type of study should be considered when interpreting the results. PMID- 15661132 TI - Effectiveness of Bacillus Calmette Guerin (BCG) vaccination in the prevention of leprosy: a population-based case-control study in Yavatmal District, India. AB - OBJECTIVE: To estimate the effectiveness of Bacillus Calmette Guerin (BCG) vaccination in the prevention of leprosy. Study design. Population-based case control study. METHODS: The study was carried out in Yavatmal District, Maharashtra, India. It included 364 cases of leprosy (diagnosed by the World Health Organization's criteria), born since 1962, that were detected during a leprosy survey conducted by the Government of Maharashtra in 2,175,514 people. Each case was pair-matched with one neighbourhood control for age, sex and socio economic status. Exclusion criteria for controls included past or current history of tuberculosis or leprosy. BCG vaccination status was assessed by examination for the presence of a BCG scar, immunization records if available and information from subjects/parents of children. Subjects who were uncertain about BCG vaccination were not included. RESULTS: A significant protective association between BCG and leprosy was observed [odds ratio=0.46, 95% confidence intervals (CI) 0.34-0.61]. Overall vaccine effectiveness (VE) was 54% (95% CI 39-66). BCG effectiveness against multibacillary, paucibacillary and single skin lesion leprosy was 68% (95% CI 26-86), 57% (95% CI 29-74) and 48% (95% CI 22-65), respectively. Analysis of linear trend revealed a significant linear association between the protective effect of BCG and the type of leprosy. The BCG vaccine was more effective in those aged < or =20 years compared with those aged >20 years (VE 61%, 95% CI), among females compared with males (VE 60%, 95% CI), in lower socio-economic strata compared with upper and middle strata (VE 57%, 95% CI), and in subjects who had a BCG scar size < or =5 mm compared with those with a BCG scar size >5 mm (VE 61%, 95% CI). However, these differences were not statistically significant, as reflected by the overlapping 95% CIs. The overall prevented fraction was 35% (95% CI 22-46). CONCLUSION: The current study identified a beneficial role of BCG vaccination in the prevention of leprosy in the study population. PMID- 15661134 TI - In vitro dendritic cell infection by pseudotyped adenoviral vectors does not correlate with their in vivo immunogenicity. AB - Expression of antigens in dendritic cells (DC) can stimulate protective immunity against both viral infection and tumor growth, making them important targets for gene therapy. In-vitro-generated DC are commonly used in gene delivery studies with the assumption that the results will correlate with in vivo activity. Adenovirus Type 5 (Ad5) vectors have been widely used with DC, but these cells lack the primary receptor (CAR) used by Ad5 and are poorly infected. We investigated the use of Ad5 vector particles pseudotyped with fibers from other Ad serotypes in DC targeting. Several fiber proteins, including those from Ad16 (Subgroup B) and Ad37 (Subgroup D), conferred dramatically increased in vitro infection. Surprisingly, neither dendritic cell infection nor the immune response to an Ad-delivered antigen was improved when the modified viruses were tested in vivo. These results underscore the importance of using appropriate animal models in gene delivery studies. PMID- 15661135 TI - Inhibition of native hepatitis C virus replicase by nucleotide and non-nucleoside inhibitors. AB - A number of nucleotide and non-nucleoside inhibitors of HCV polymerase are currently under investigation as potential antiviral agents to treat HCV-infected patients. HCV polymerase is part of a replicase complex including the polymerase subunit NS5B together with other viral and host proteins and viral RNA. The RNA synthesis activity of the native replicase complex was inhibited by 3'-deoxy-CTP, a chain-terminating nucleotide analog, but not inhibited by non-nucleoside NS5B polymerase inhibitors of three different structural classes. The HCV replicase was also resistant to heparin, a broad-spectrum, RNA-competitive polymerase inhibitor. Prebinding of the recombinant NS5B protein with a RNA template rendered the polymerase largely resistant to the inhibition by heparin and the non-nucleoside inhibitors, but did not affect the inhibitory potency of 3'-deoxy CTP. Therefore, the HCV replicase showed a similar pattern of inhibitor sensitivity as compared to RNA-bound NS5B. These results suggest that the native HCV replicase complex represents a stable and productive polymerase-RNA complex. The allosteric non-nucleoside NS5B polymerase inhibitors are inactive against established HCV replicase but may function antagonistically with the formation of a productive enzyme-template complex. PMID- 15661136 TI - Decreased perforin and granzyme B expression in senescent HIV-1-specific cytotoxic T lymphocytes. AB - Cytotoxic T lymphocyte (CTL) senescence may be an important mechanism of immune failure in HIV-1 infection. We find that senescence of HIV-1-specific CTL clones causes loss of killing activity, preventable by transduction with telomerase. Furthermore, senescence is associated with reduced expression of the effector molecules granzyme and perforin, suggesting CTL "exhaustion" can result in hypofunction. These results agree with other studies showing that HIV-1-specific CTL exhibit abnormal phenotypes in vivo, and suggest the possibility that chronic turnover is an important mechanism of antiviral failure in HIV-1 infection. PMID- 15661137 TI - Generation of eGFP expressing recombinant Zaire ebolavirus for analysis of early pathogenesis events and high-throughput antiviral drug screening. AB - Zaire ebolavirus causes large outbreaks of severe and usually fatal hemorrhagic disease in humans for which there is no effective treatment or cure. To facilitate examination of early critical events in viral pathogenesis and to identify antiviral compounds, a recombinant Zaire ebolavirus was engineered to express a foreign protein, eGFP, to provide a rapid and sensitive means to monitor virus replication in infected cells. This genetically engineered virus represents the first insertion of a foreign gene into ebolavirus. We show that Ebola-eGFP virus (EboZ-eGFP) infects known early targets of human infections and serves as an ideal model to screen antiviral compounds in less time than any previously published assay. PMID- 15661138 TI - In vitro simian virus 40 large tumor antigen expression correlates with differential immune responses following DNA immunization. AB - Simian virus 40 (SV40) contains an essential protein, large tumor antigen (Tag), which assists in viral replication and causes cell transformation and immortalization. Our laboratory has examined plasmid DNA, expressing SV40 Tag under two different promoters, for use in potential cancer vaccination strategies. One plasmid, pSV3-neo, failed to induce SV40 Tag antibody, produced a weak cell-mediated response, and only partial protection in murine experimental tumor challenge systems. The second plasmid, pCMV-Tag, induced antibodies to SV40 Tag, produced a robust cell-mediated response, and invoked complete tumor immunity in vivo. The induction of CD4+ and CD8+ T cell responses following plasmid DNA immunization and tumor cell challenge reflected a type 1 cytokine secretion profile. Our hypothesis for this differential immune response is that pCMV-Tag exhibits a higher level of transgene expression due to a more efficient promoter. We determined that pCMV-Tag levels of SV40 Tag mRNA and protein expression were higher when compared to pSV3-neo. A threshold amount of SV40 Tag may be required to stimulate antibody production and provide complete systemic tumor immunity. PMID- 15661139 TI - Genetic characterization of Yokose virus, a flavivirus isolated from the bat in Japan. AB - Yokose virus (strain Oita-36) was isolated from the bat in Japan in 1971. In the present study, we determined complete nucleotide sequences of Yokose virus using RT-PCR and RACE techniques. Yokose virus genome consists of 10,857 nucleotides in length (accession no. AB114858), containing a single open reading frame (3425 amino acids) encoding 11 viral proteins. We deduced the boundaries of each protein in the polyprotein sequence according to the protein cleavage sites of other flaviviruses. The nucleotide sequences of the 5' and 3' nontranslated region (NTR) and amino acid sequences of individual proteins of the virus were compared with those of six other flaviviruses including Japanese encephalitis virus, dengue-2 virus, yellow fever virus, West Nile virus, tick-borne encephalitis virus, and Rio Bravo virus or Modoc virus. Yokose virus demonstrated the highest similarity to yellow fever virus. Yokose virus also has CS1 motif, which are well-conserved specifically in mosquito-born flaviviruses, in its 3' NTR. When a part of the NS5 amino acid sequence (345 amino acids) was compared with those of other four flaviviruses, Entebbe bat virus, Sokuluk virus, Sepik virus, and yellow fever virus, the three former viruses are more closely related to Yokose virus than yellow fever virus. Human sera from dengue-virus-infected case and yellow fever vaccine reacted with the viral proteins. Moreover, human serum from a yellow fever vaccine weakly neutralized Yokose virus. Our results suggest that there are cross-reactive antigenicities among Yokose virus and other flaviviruses. PMID- 15661140 TI - Complete genome sequence of bacteriophage T5. AB - The 121,752-bp genome sequence of bacteriophage T5 was determined; the linear, double-stranded DNA is nicked in one of the strands and has large direct terminal repeats of 10,139 bp (8.3%) at both ends. The genome structure is consistently arranged according to its lytic life cycle. Of the 168 potential open reading frames (ORFs), 61 were annotated; these annotated ORFs are mainly enzymes involved in phage DNA replication, repair, and nucleotide metabolism. At least five endonucleases that believed to help inducing nicks in T5 genomic DNA, and a DNA ligase gene was found to be split into two separate ORFs. Analysis of T5 early promoters suggests a probable motif AAA{3, 4 T}nTTGCTT{17, 18 n}TATAATA{12, 13 W}{10 R} for strong promoters that may strengthen the step modification of host RNA polymerase, and thus control transcription of phage DNA. The distinct protein domain profile and a mosaic genome structure suggest an origin from the common genetic pool. PMID- 15661141 TI - West Nile premembrane-envelope genetic vaccine encoded as a chimera containing the transmembrane and cytoplasmic domains of a lysosome-associated membrane protein: increased cellular concentration of the transgene product, targeting to the MHC II compartment, and enhanced neutralizing antibody response. AB - A genetic vaccine for West Nile virus (WN) has been synthesized with the WN premembrane-envelope (WN preM-E) gene sequences encoded as a chimera with the transmembrane and carboxyl terminal domains of the lysosome-associated membrane protein (LAMP). The LAMP sequences are used to direct the antigen protein to the major histocompatibility class II (MHC II) vesicular compartment of transfected professional antigen-presenting cells (APCs). Vaccine constructs encoding the native WN preM-E and WN preM-E/LAMP chimera were synthesized in pVAX1 and pITR plasmid backbones. Extracts of human fibroblast 293 and monkey kidney COS-7 cells transfected with the WN preM-E/LAMP chimera constructs contained much greater amounts of E than did the cells transfected with constructs encoding the native WN preM-E. This difference in the concentration of native E and the E/LAMP chimera in transfected cells is attributed to the secretion of native E. The amount of preM protein in cell extracts, in contrast to the E protein, and the levels of DNA and RNA transcripts, did not differ between WN preM-E- and WN preM E/LAMP-transfected cells. Additionally, confocal and immunoelectron microscopic analyses of transfected B cells showed localization of the WN preM-E/LAMP chimera in vesicular compartments containing endogenous LAMP, MHC II, and H2-M, whereas native viral preM-E lacking the LAMP sequences was distributed within the cellular vesicular network with little LAMP or MHC II association. Mice immunized with a DNA construct expressing the WN preM-E/LAMP antigen induced significant antibody and long-term neutralization titers in contrast to the minimal and short lived neutralization titer of mice vaccinated with a plasmid expressing the untargeted antigen. These results underscore the utility of LAMP targeting of the WN envelope to the MHC II compartments in the design of a genetic WN vaccine. PMID- 15661142 TI - An acidic amphipathic helix in the Bovine Papillomavirus E2 protein is critical for DNA replication and interaction with the E1 protein. AB - The papillomavirus E2 proteins function in viral transcriptional regulation, and genome replication and episomal maintenance. The transactivation domain is essential for these activities. To identify functional regions, a structural model of the BPV1 E2 transactivation domain was used to target surface residues for mutation. Mutation of several previously uncharacterized regions yielded proteins specifically disrupted in the replication activity of E2. Mutations in an amino-terminal acidic amphipathic helix disrupted the interaction of the E1 and E2 proteins and a peptide derived from this helix blocked cooperative origin binding of E1 and E2. Mutation of clusters of charged residues, R47, K48, K49, R58, and H61 or R172, D175, E176, and R179, or residue R68 in the previously described putative E1 interaction region, specifically disrupted replication while retaining the ability to bind to the E1 protein. Thus, this approach has identified novel regions that are required for the replication function of E2. PMID- 15661143 TI - Multiple domains of the SIV Env protein determine virus replication efficiency and neutralization sensitivity. AB - SIVmac239 and SIVmac1A11 are wild-type viruses encoding Env proteins with full length or truncated cytoplasmic tails (CTs), respectively. A mutant designated SIVmac239T has a site-specific mutation which introduces a stop codon in the env gene resulting a truncated protein of similar length to SIVmac1A11 Env. To investigate the role of specific sequence differences in these Env proteins, we constructed SIV mutants encoding 1A11 or 239 Env proteins with reciprocal exchanges of the CT or exchanges of both the surface unit (SU) and CT sequences. A truncated CT in the context of the 1A11 SU subunit was found to significantly enhance replication in CEMx174 (human T-cell line) and rhesus PBMCs. However, similar Env CT truncation did not enhance replication of SIVmac239 in human or monkey cells. SIVmac1A11 with a full-length SIVmac239 CT did not replicate in human T-cell lines, but truncation of the CT by a stop codon resulted in replication. We also observed that these viruses differed significantly in sensitivity to neutralization by antibody. Taken together, the results indicated that the length of the CT domain as well as specific sequence differences in the SU domain affect viral replication capacity as well as sensitivity to neutralization. PMID- 15661144 TI - Are transforming properties of the bovine papillomavirus E5 protein shared by E5 from high-risk human papillomavirus type 16? AB - The E5 proteins of bovine papillomavirus type 1 (BPV-1) and human papillomavirus type 16 (HPV-16) are small (44-83 amino acids), hydrophobic polypeptides that localize to membranes of the Golgi apparatus and endoplasmic reticulum, respectively. While the oncogenic properties of BPV-1 E5 have been characterized in detail, less is known about HPV-16 E5 due to its low expression in mammalian cells. Using codon-optimized HPV-16 E5 DNA, we have generated stable fibroblast cell lines that express equivalent levels of epitope-tagged BPV-1 and HPV-16 E5 proteins. In contrast to BPV-1 E5, HPV-16 E5 does not activate growth factor receptors, phosphoinositide 3-kinase or c-Src, and fails to induce focus formation, although it does promote anchorage-independent growth in soft agar. These variant activities are apparently unrelated to differences in intracellular localization of the E5 proteins since retargeting HPV-16 E5 to the Golgi apparatus does not induce focus formation. PMID- 15661145 TI - Genomic and bioinformatics analysis of HAdV-7, a human adenovirus of species B1 that causes acute respiratory disease: implications for vector development in human gene therapy. AB - Human adenovirus serotype 7 (HAdV-7) is a reemerging pathogen identified in acute respiratory disease (ARD), particularly in epidemics affecting basic military trainee populations of otherwise healthy young adults. The genome has been sequenced and annotated (GenBank accession no. ). Comparative genomics and bioinformatics analyses of the HAdV-7 genome sequence provide insight into its natural history and phylogenetic relationships. A putative origin of HAdV-7 from a chimpanzee host is observed. This has implications within the current biotechnological interest of using chimpanzee adenoviruses as vectors for human gene therapy and DNA vaccine delivery. Rapid genome sequencing and analyses of this species B1 member provide an example of exploiting accurate low-pass DNA sequencing technology in pathogen characterization and epidemic outbreak surveillance through the identification, validation, and application of unique pathogen genome signatures. PMID- 15661146 TI - Molecular profiling of early stage liver fibrosis in patients with chronic hepatitis C virus infection. AB - The molecular mechanisms of acute hepatitis C virus (HCV) infection, end-stage hepatitis (cirrhosis), and hepatocellular carcinoma have been extensively studied, but little is known of the changes in liver gene expression during the early stages of liver fibrosis associated with chronic HCV infection, that is, the transition from normal liver (NL) of uninfected patients to the first stage of liver fibrosis (F1-CH-C). To obtain insight into the molecular pathogenesis of F1-CH-C, we used real-time quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) to study the mRNA expression of 240 selected genes in liver tissue with F1-CH-C, in comparison with NL. The expression of 54 (22.5%) of the 240 genes was significantly different between F1-CH-C and NL; 46 genes were upregulated and 8 were downregulated in F1-CH-C. The most noteworthy changes in gene expression mainly affected the transcriptional network regulated by interferons (IFNs), including both IFN-alpha/beta-inducible genes (STAT1, STAT2, ISGF3G/IRF9, IFI27, G1P3, G1P2, OAS2, MX1) and IFN-gamma-inducible genes (CXCL9, CXCL10, CXCL11). Interesting, upregulation of IFN-alpha/beta-inducible genes (but not IFN-gamma-inducible genes) was independent of histological scores (grade and stage of fibrosis) and HCV characteristics (hepatic HCV mRNA levels and the HCV genotype), and was specific to HCV (as compared to hepatitis B virus (HBV)). Other genes dysregulated in F1-CH-C, albeit less markedly than IFN-alpha/beta- and IFN-gamma-inducible genes, were mainly involved in the activation of lymphocytes infiltrating the liver (IFNG, TNF, CXCL6, IL6, CCL8, CXCR3, CXCR4, CCR2), cell proliferation (p16/CDKN2A, MKI67, p14/ARF), extracellular matrix remodeling (MMP9, ITGA2), lymphangiogenesis (XLKD1/LYVE), oxidative stress (CYP2E1), and cytoskeleton microtubule organization (STMN2/SCG10). Thus, a limited number of signaling pathways, and particularly the transcriptional network regulated by interferons, are dysregulated in the first stage of HCV induced liver fibrosis. Some of the genes identified here could form the basis for new approaches aimed at refining IFN-based therapies for chronic HCV infection. PMID- 15661147 TI - Selective recognition of oligomeric HIV-1 primary isolate envelope glycoproteins by potently neutralizing ligands requires efficient precursor cleavage. AB - A critical component of an effective HIV vaccine will be the induction of broadly neutralizing antibodies. Comprising the HIV spike, the exterior envelope glycoprotein gp120 and the transmembrane glycoprotein gp41 mediate receptor binding, viral entry, and are the targets for neutralizing antibodies. The gp120 and gp41 glycoproteins are derived from the gp160 precursor glycoprotein and following gp160 glycosylation, oligomerization and cleavage in the endoplasmic reticulum and Golgi, remain as non-covalently associated trimers of heterodimers. Previously, using cell-surface envelope glycoproteins derived from infection of a laboratory-adapted HIV-1 strain, a correlation had been established between the binding of gp120-directed antibodies to the viral glycoprotein and the ability of the antibodies to neutralize laboratory-adapted isolates. However, this has been more difficult to demonstrate for glycoproteins derived from primary patient isolates. Here, using a FACS-based method, we report that only gp120-directed neutralizing antibodies and the neutralizing ligand soluble CD4 efficiently bind to glycoproteins derived from the JR-FL primary isolate provided that the gp160 precursor protein is efficiently cleaved. Precursor cleavage was demonstrated by cell-surface biotinylation and Western blotting. In stark contrast, both non neutralizing and neutralizing antibodies bind non-cleaved envelope glycoproteins from JR-FL and YU2 isolates. These data imply that significant changes in Env spike structure are dependent upon precursor gp160 cleavage and are consistent with a restricted-binding-to-Env model of neutralization. The data also have implications in regards to the use and design of non-cleaved envelope glycoprotein trimeric immunogens as a means to selectively and preferentially present neutralizing epitopes to the host immune system. PMID- 15661148 TI - Porcine adenovirus serotype 3 internalization is independent of CAR and alphavbeta3 or alphavbeta5 integrin. AB - Nonhuman adenoviruses including porcine adenovirus serotype 3 (PAd3) are emerging vectors for gene delivery. PAd3 efficiently transduces human and murine cells in culture, and circumvents preexisting humoral immunity in humans. The coxsackievirus-adenovirus receptor (CAR) serves as a primary receptor and alphavbeta3 or alphavbeta5 integrin as a secondary receptor for several human adenovirus (HAd) subtypes including HAd5. In this study, we deduced the role of CAR, alphavbeta3 or alphavbeta5 integrin in PAd3 internalization. Transduction experiments were conducted in human mammary epithelial (MCF-10A) cells using replication-defective PAd-GFP (PAd3 vector expressing green fluorescent protein [GFP]) and HAd-GFP (HAd5 vector expressing GFP). MCF-10A cells were treated with or without anti-human CAR, or anti-alphavbeta3 or anti-alphavbeta5 integrin antibodies prior to infection with HAd-GFP or PAd-GFP. Significant (P <0.05) inhibition in transduction by HAd-GFP was observed in antibody-treated cells as compared to untreated cells, whereas transduction by PAd-GFP remained to similar levels irrespective of the treatment. To study the adenoviral fiber knob-mediated virus interference, MCF-10A cells were treated with or without the recombinant HAd5 or PAd3 knob followed by infection with HAd-GFP or PAd-GFP. Significant (P <0.05) inhibition was observed only in transduction of the homologous vector. These results suggested that PAd3 internalization was CAR- as well as alphavbeta3 or alphavbeta5 integrin-independent and the primary receptor for HAd5 and PAd3 were distinct. CAR- and alphavbeta3 or alphavbeta5 integrin-independent entry of PAd3 vectors may have implications in targeting cell types that are not efficiently transduced by other adenoviral vectors. PMID- 15661149 TI - Characterization of H5N1 influenza A viruses isolated during the 2003-2004 influenza outbreaks in Japan. AB - In Japan, between the end of December 2003 and March 2004, four outbreaks of acute, highly transmissible and lethal disease occurred in birds in three prefectures separated by 150-450 km, involving three chicken farms and a group of chickens raised as pets. The cause of each outbreak was an H5N1 influenza A virus the first highly pathogenic virus to be isolated from the outbreaks in Japan since 1925. The H5N1 virus was also isolated from dead crows, apparently infected by contact with virus-contaminated material. These H5N1 viruses were antigenically similar to each other, but could be differentiated from other H5 viruses, including those isolated from Hong Kong in 1997 and 2003, by use of a panel of monoclonal antibodies in hemagglutination inhibition assays. Genetically, the H5N1 viruses in Japan were closely related to each other in all genes and were genetically closely related to a single isolate of genotype V that was isolated in 2003 in the Guandong Province of mainland China (A/chicken/Shantou/4231/2003). The virulence of the index isolate (A/chicken/Yamaguchi/7/2004) was studied in chickens and mice. Chickens intravenously or intranasally inoculated with the isolate died within 1 or 3 days of inoculation, respectively. In mice, although this virus replicated well in the lung without prior adaptation and spread to the brain, the dose lethal to 50% of the mice was 5 x 10(5) 50% egg infectious doses (EID50), which is less pathogenic than the Hong Kong 1997 H5N1 viruses isolated from humans. Our findings indicate that the H5N1 viruses associated with the influenza outbreaks in chickens in Japan were genotypically closely related to an H5N1 virus isolated from chicken in China in 2003 (genotype V), but were different from those prevalent in southeastern Asia in 2003-2004 (i.e., genotype Z) and that these highly pathogenic viruses can be transmitted to crows, which are highly susceptible to these viruses. PMID- 15661150 TI - A porcine G9 rotavirus strain shares neutralization and VP7 phylogenetic sequence lineage 3 characteristics with contemporary human G9 rotavirus strains. AB - Of five globally important VP7 (G) serotypes (G1-4 and 9) of group A rotaviruses (the single most important etiologic agents of infantile diarrhea worldwide), G9 continues to attract considerable attention because of its unique natural history. Serotype G9 rotavirus was isolated from a child with diarrhea first in the United States in 1983 and subsequently in Japan in 1985. Curiously, soon after their detection, G9 rotaviruses were not detected for about a decade in both countries and then reemerged in both countries in the mid-1990s. Unexpectedly, however, such reemerged G9 strains were distinct genetically and molecularly from those isolated in the 1980s. Thus, the origin of the reemerged G9 viruses remains an enigma. Sequence analysis has demonstrated that the G9 rotavirus VP7 gene belongs to one of at least three phylogenetic lineages: lineage 1 (strains isolated in the 1980s in the United States and Japan), lineage 2 (strains first isolated in 1986 and exclusively in India thus far), and lineage 3 (strains that emerged/reemerged in the mid-1990s). Currently, lineage 3 G9 viruses are the most frequently detected G9 strains globally. We characterized a porcine rotavirus (A2 strain) isolated in the United States that was known to belong to the P[7] genotype but had not been serotyped by neutralization. The A2 strain was found to bear serotype G9 and P9 specificities as well as NSP4 [B] and subgroup I characteristics. By VP7-specific neutralization, the porcine G9 strain was more closely related to lineage 3 viruses than to lineage 1 or 2 viruses. Furthermore, by sequence analysis, the A2 VP7 was shown to belong to lineage 3 G9. These findings raise intriguing questions regarding possible explanations for the emergence of variations among the G9 strains. PMID- 15661151 TI - Parvovirus B19 does not bind to membrane-associated globoside in vitro. AB - The glycosphingolipid globoside (globotetraosylceramide, Gb4Cer) has been proposed to be the cellular receptor of human parvovirus B19. Quantitative measurements of the binding of parvovirus B19 to Gb4Cer were performed to explore the molecular basis of the virus tropism. Solid-phase assays with fluorescence labeled liposomes or 125iodine-labeled empty capsids were used to characterize the specificity of binding. In addition, surface plasmon resonance on lipid layers, as well as isothermal titration microcalorimetry, was utilized for real time analysis of the virus-receptor interaction. These studies did not confirm binding of Gb4Cer to recombinant B19 VP2 capsids, suggesting that Gb4Cer does not function on its own as the cellular receptor of human parvovirus B19, but might be involved in a more complex recognition event. The biochemical results were further confirmed by cryo-electron microscopy image reconstructions at 10 A resolution, in which the structures of empty capsids were compared with empty capsids incubated with Gb4Cer. PMID- 15661152 TI - Self-interaction of ORF II protein through the leucine zipper is essential for Soybean chlorotic mottle virus infectivity. AB - The ORF II protein (PII) of Soybean chlorotic mottle virus (SbCMV) is essential for the virus life cycle. We investigated the interactions of SbCMV PII with itself and with other essential virus proteins using a Gal4-based yeast two hybrid system. PII interacted only with itself and not with any other virus proteins. The PII-PII interaction was confirmed by a Sos-based yeast two-hybrid system and a far-western analysis. Deletion mutagenesis mapped the self interacting domain to the C-terminal 48 amino acids (amino acids 154-201), which contain two putative leucine zipper motifs. Introduction of amino acid substitutions to leucine/isoleucine in zipper sequences prevented the PII-PII interaction and abolished the infectivity of SbCMV. These results revealed that the self-interaction of PII through a leucine zipper is necessary for virus infection. PMID- 15661153 TI - In vitro assembled, recombinant infectious bronchitis viruses demonstrate that the 5a open reading frame is not essential for replication. AB - Molecular clones of infectious bronchitis virus (IBV), derived from the Vero cell adapted Beaudette strain, were constructed, using an in vitro assembly method. In vitro transcribed RNA from a cDNA template that had been constructed from seven cDNA fragments, encompassing the entire genome of IBV, was electroporated into BHK-21 cells. The cells were overlaid onto the susceptible Vero cells and viable virus was recovered from the molecular clone. The molecularly cloned IBV (MIBV) demonstrated growth kinetics, and plaque size and morphology that resembled the parental Beaudette strain IBV. The recombinant virus was further manipulated to express enhanced green fluorescent protein (EGFP) by replacing an open reading frame (ORF) of the group-specific gene, ORF 5a, with the EGFP ORF. The rescued recombinant virus, expressing EGFP (GIBV), replicated to lower viral titers and formed smaller plaques compared to the parental virus and the MIBV. After six passages of GIBV, a minority of plaques were observed that had reverted to the larger plaque size and virus from these plaques no longer expressed EGFP. Direct sequencing of RT-PCR products derived from cells infected with the plaque purified virus, which had lost expression of EGFP, confirmed loss of the EGFP ORF. The loss of EGFP expression (Delta5a IBV) was also accompanied by reversion to growth kinetics resembling the standard virus and intact recombinant virus. This study demonstrates that the 5a ORF is not essential for viral multiplication in Vero cells. PMID- 15661155 TI - The polymerase (L) protein of rinderpest virus interacts with the host cell protein striatin. AB - Rinderpest virus (RPV) is a morbillivirus that causes a highly contagious disease affecting members of the order Artiodactyla. The viral L protein is the catalytic subunit of the RNA-dependent RNA polymerase. To search for host cell proteins with which L interacts, a library screen was performed using the yeast two-hybrid system. Several host cell proteins were recovered from the library screen as putative L-interactors; one of these was identified as striatin. A direct interaction between RPV L and striatin was confirmed using both co immunoprecipitation assays and co-localisation studies using confocal microscopy. Striatin was also shown to co-localise with the RPV L protein in infected cells. The L proteins of morbilliviruses consist of three long highly conserved domains separated by short unconserved stretches of amino acids. The L domain with which striatin interacts was investigated by co-immunoprecipitation and striatin was shown to interact primarily with the central conserved domain. PMID- 15661154 TI - Effect of mutating the two cysteines required for HBe antigenicity on hepatitis B virus DNA replication and virion secretion. AB - Hepatitis B virus (HBV) variants with impaired expression of e antigen (HBeAg) frequently arise at the chronic stage of infection, as exemplified by precore and core promoter mutants. Since an intramolecular disulfide bond maintains the secondary structure of HBeAg, we explored effect of missense mutations of either cysteine codon. Consistent with earlier reports, substitution of each cysteine rendered HBeAg nearly undetectable. With underlying nucleotide changes at the loop of pregenome encapsidation signal, the C-7 mutants were severely impaired in pregenomic RNA packaging and hence DNA replication. Although none of the missense mutations at C61 reduced DNA replication, replacement with arginine, but not alanine, aspartic acid, phenylalanine, or serine, blocked virion secretion. Consistent with the detection of C61R genome from a patient serum, secretion block of the C61R mutant could be overcome by co-expression of wild-type core protein. In conclusion, point mutations of the C61 codon may generate viable HBeAg-negative variants. PMID- 15661156 TI - Myxoma virus infection of primary human fibroblasts varies with cellular age and is regulated by host interferon responses. AB - Recent studies have indicated a critical role for interferon (IFN)-mediated antiviral responses in the host range of myxoma virus (MV), a pathogenic poxvirus of rabbits. To investigate the contribution of IFN to MV tropism in nonleporine cells, primary human dermal fibroblasts (HDFs) were tested for permissiveness to MV infection. Low-passage HDFs that underwent fewer than 25 population doublings (PD) were fully permissive for MV infection, supporting productive virus replication and cell-to-cell spread. In contrast, early and late viral gene expression was detectable in high-passage HDF (>75 PD), but MV failed to generate infectious progeny and could not form foci in these cells. Vesicular stomatitis virus (VSV) plaque reduction assays confirmed that constitutive IFN production progressively increased as HDFs were passaged, concurrent with an increase in the expression of transcripts for type I IFN and IFN-responsive genes involved in antiviral responses. These findings correlated with the enhanced sensitivity of higher-passage HDF to inducers of type I IFN responses, such as dsRNA. Furthermore, pretreatment of low-passage HDF with type I IFN abrogated MV spread and replication while treatment of mature HDF with neutralizing antibodies to IFN beta, but not IFN-alpha, restored the capacity to form foci. These findings emphasize the importance of post-entry events in determining the permissiveness of human cells to MV infection and support a critical role for innate type I IFN responses as key determinants of poxvirus host range and species restriction. PMID- 15661157 TI - p21WAF1 modulates NF-kappaB signaling and induces anti-apoptotic protein Bcl-2 in Tax-expressing rat fibroblast. AB - Of the cell cycle-associated genes regulated by human T-cell leukemia virus type 1 (HTLV-1) Tax, cyclin-dependent kinase (CDK) inhibitor p21WAF1 is upregulated in HTLV-1-infected cells. Previously, we reported that p21WAF1 stimulated Tax dependent NF-kappaB activation which influences a variety of cellular processes, including proliferation, differentiation, and apoptosis. In HTLV-1-infected cells, Tax is primarily involved in the constitutive activation of NF-kappaB signaling. Here, we demonstrate that p21WAF1 affects Tax-dependent NF-kappaB signaling by inducing p100/52, an NF-kappaB-related protein. W4, a Tax transformed rat fibroblast cell line, exhibits the constitutive activation of NF kappaB signaling, potentially mediated by overexpression of RelB. Ectopic expression of p21WAF1 in W4 cells, which lack endogenous expression due to methylation of the p21WAF1 promoter, induces the expression of p100/52. Bcl-2 expression was also upregulated by ectopic p21WAF1 in this cell line, suggesting that p21WAF1 plays an important role in the regulation of apoptosis by modulating NF-kappaB signaling in Tax-expressing rat fibroblasts. We also address the expression of NF-kappaB-related proteins in HTLV-1-infected cells. PMID- 15661158 TI - Spleen necrosis virus-based vector delivery of anti-HIV-1 genes potently protects human hematopoietic cells from HIV-1 infection. AB - In this study, we report on the efficacy of using a spleen necrosis virus (SNV) based vector delivery system to block human immunodeficiency virus type I (HIV-1) replication in human hematopoietic cells. These efforts were directed towards the development of human immune system cell resistance to HIV-1 infection, based on the strategy of "intracellular immunization" via generation of a series of anti HIV-1 therapeutic constructs carrying scFvs, single-chain variable fragments, against HIV-1 integrase and reverse transcriptase in combination with the trans dominant mutant of HIV-1 Rev, RevM10. The efficiency of the anti-HIV-1 constructs were tested in viral challenge assays with different doses of HIV-1 NL4-3, Bal, 89.6 and R7-GFP strains. These experiments demonstrated the reduction of HIV-1 replication by these retroviral vector constructs in a range of 4- to 10-fold in CD4+ T-lymphocytes, human peripheral blood mononuclear cells (PBMCs), and primary human macrophages. We observed selective efficiency of SNV-based therapeutics in H9, C8166 and Jurkat T-lymphocytic cell lines, demonstrating the most efficient inhibition of HIV-1 replication in Jurkat T-cells. Thus, these data are the first demonstration of the ability of SNV-based retroviral vectors with select transgenes, which may have certain molecular advantages over other retroviral vector systems, to combat HIV-1 replication in human hematopoietic cells and support the potential for using SNV-expressed constructs in anti-HIV-1 molecular therapeutics. PMID- 15661160 TI - Cooperative interactions between bacteriophage P2 integrase and its accessory factors IHF and Cox. AB - Bacteriophage P2 integrase (Int) mediates site-specific recombination leading to integration or excision of the phage genome in or out of the bacterial chromosome. Int belongs to the large family of tyrosine recombinases that have two different DNA recognition motifs binding to the arm and core sites, respectively, which are located within the phage attachment sites (attP). In addition to the P2 integrase, the accessory proteins Escherichia coli IHF and P2 Cox are needed for recombination. IHF is a structural protein needed for integration and excision by bending the DNA. As opposed to lambda, only one IHF site is found in P2 attP. P2 Cox controls the direction of recombination by inhibiting integration but being required for excision. In this work, the effects of accessory proteins on the capacity of Int to bind to its DNA recognition sequences are analyzed using electromobility shifts. P2 Int binds with low affinity to the arm site, and this binding is greatly enhanced by IHF. The arm binding domain of Int is located at the N-terminus. P2 Int binds with high affinity to the core site, and this binding is also enhanced by IHF. The fact that the cooperative binding of Int and IHF is strongly reduced by lengthening the distance between the IHF and core binding sites indicates that the distance between these sites may be important for cooperative binding. The Int and Cox proteins also bind cooperatively to attP. PMID- 15661159 TI - Re-examination of feline leukemia virus: host relationships using real-time PCR. AB - The mechanisms responsible for effective vs. ineffective viral containment are central to immunoprevention and therapies of retroviral infections. Feline leukemia virus (FeLV) infection is unique as a naturally occurring, diametric example of effective vs. ineffective retroviral containment by the host. We developed a sensitive quantitative real-time DNA PCR assay specific for exogenous FeLV to further explore the FeLV-host relationship. By assaying p27 capsid antigen in blood and FeLV DNA in blood and tissues of successfully vaccinated, unsuccessfully vaccinated, and unvaccinated pathogen-free cats, we defined four statistically separable classes of FeLV infection, provisionally designated as abortive, regressive, latent, and progressive. These host-virus relationships were established by 8 weeks post-challenge and could be maintained for years. Real-time PCR methods offer promise in gaining deeper insight into the mechanisms of FeLV infection and immunity. PMID- 15661161 TI - Differential effects of virulent and avirulent equine infectious anemia virus on macrophage cytokine expression. AB - Equine infectious anemia virus (EIAV) causes rapid development of acute disease followed by recurring episodes of fever, thrombocytopenia, and viremia. Most infected equid eventually bring the virus under immunological control. We recently reported the development of an equine-specific ribonuclease protection assay (RPA) to quantitate mRNA levels of 10 cytokines. Using this newly developed RPA, we now show significant differences in cytokine induction in equine monocyte derived macrophages (EMDM) exposed to virulent and avirulent EIAV. Virulent EIAV17 induced significant increases in interleukin (IL)-1alpha, IL-1beta, IL-6, IL-10, and tumor necrosis factor (TNF)-alpha by 0.5-1 h postinfection (hpi). In contrast, the avirulent virus failed to induce any of the tested cytokines above that of control levels. These data show a direct correlation between cytokine dysregulation and EIAV pathogenesis. PMID- 15661162 TI - Requirement of the vesicular system for membrane permeabilization by Sindbis virus. AB - The vast majority of animal viruses enhance membrane permeability at two moments of infection. Herein, we describe that the entry of Sindbis virus (SV) in BHK cells promotes the co-entry of the macromolecule alpha-sarcin into the cytoplasm, thereby blocking translation. At a later stage, this protein toxin cannot enter the cell, while low molecular weight compounds, such as hygromycin B, readily pass through the plasma membrane of Sindbis virus-infected BHK cells. To unveil the participation of the different Sindbis virus structural proteins in late permeabilization, transfection experiments with each late gene by separate have been carried out. Our findings indicate that 6K is the main determinant that enhances membrane permeabilization. The co-expression of both viral glycoproteins employing a Sindbis virus variant that lacks the entire 6K gene partly modifies membrane permeability. Brefeldin A, a macrolide antibiotic that interferes with the proper functioning of the vesicular system, hampers the induction of membrane leakiness without significantly affecting viral protein synthesis. On the other hand, the flavone compound Ro-090179 also diminishes the entry of hygromycin B, while bafilomycin A1 or nocodazole have no effect. These data reveal the requirement of the vesicular system for late viral membrane permeabilization. PMID- 15661163 TI - Human immunodeficiency virus type 1 Tat protein induces an intracellular calcium increase in human monocytes that requires DHP receptors: involvement in TNF-alpha production. AB - HIV-1 Tat protein, acting at the cell membrane, stimulates the production by human monocytes of TNF-alpha, a cytokine implicated in both HIV-1 replication and pathogenesis. Here, we analyze, in primary human monocytes, the mechanisms involved in Tat-stimulated calcium mobilization and its relationship with TNF alpha production. We show that the Tat protein induces a calcium signal by mobilizing calcium from extracellular stores. This calcium signal is totally blocked when cells are stimulated in the presence of DHP receptor inhibitors such as nimodipine or calcicludine, thus suggesting the implication of this L-type calcium channel. By using RT-PCR amplification, Western blot with antibodies directed against the alpha1D subunit, binding assays with specific agonists or antagonists, and inhibition with specific antisense oligonucleotides, we show that DHP receptors are expressed and functional in primary human monocytes. Interestingly, we demonstrate that Tat-induced calcium mobilization is tightly linked to TNF-alpha production, thus indicating that Tat-induced mobilization and TNF-alpha production are entirely mediated by DHP receptors, as shown by their total inhibition by nimodipine, calcicludine, or anti-alpha1D antisense oligonucleotides. PMID- 15661164 TI - Interaction of influenza virus proteins with nucleosomes. AB - During influenza virus infection, transcription and replication of the viral RNA take place in the cell nucleus. Directly after entry in the nucleus the viral ribonucleoproteins (RNPs, the viral subunits containing vRNA, nucleoprotein and the viral polymerase) are tightly associated with the nuclear matrix. Here, we have analysed the binding of RNPs, M1 and NS2/NEP proteins to purified nucleosomes, reconstituted histone octamers and purified single histones. RNPs and M1 both bind to the chromatin components but at two different sites, RNP to the histone tails and M1 to the globular domain of the histone octamer. NS2/NEP did not bind to nucleosomes at all. The possible consequences of these findings for nuclear release of newly made RNPs and for other processes during the infection cycle are discussed. PMID- 15661165 TI - Analysis of varicella zoster virus attenuation by evaluation of chimeric parent Oka/vaccine Oka recombinant viruses in skin xenografts in the SCIDhu mouse model. AB - Varicella-zoster virus (VZV) is the only human herpes virus for which a vaccine has been licensed. A clinical VZV isolate, designated the parent Oka (pOka) strain was passed in human and non-human fibroblasts to produce vaccine Oka (vOka). The pOka and vOka viruses exhibit similar infectivity in cultured cells but healthy susceptible individuals given vaccines derived from vOka rarely develop the cutaneous vesicular lesions characteristic of varicella. Inoculation of skin xenografts in the SCIDhu mouse model of VZV pathogenesis demonstrated that vOka had a reduced capacity to replicate in differentiated human epidermal cells in vivo (Moffat, J.F., Zerboni, L., Kinchington, P.R., Grose, C., Kaneshima, H., Arvin A.M., 1998a. Attenuation of the vaccine Oka strain of varicella-zoster virus and role of glycoprotein C in alphaherpesvirus virulence demonstrated in the SCID-hu mouse. J Virol. 72:965-74). In order to investigate the attenuation of vOka in skin, we made chimeric pOka and vOka recombinant viruses from VZV cosmids. Six chimeric pOka/vOka viruses were generated using cosmid sets that incorporate linear overlapping fragments of VZV DNA from cells infected with pOka or vOka. The cosmid sets consist of pOka and vOka DNA segments that have identical restriction sites. As expected, the growth kinetics and plaque morphologies of the six chimeric pOka/vOka viruses were indistinguishable in vitro. However, the chimeric viruses exhibited varying capacities to replicate when evaluated in skin xenografts in vivo. The presence of ORFs 30-55 from the pOka genome was sufficient to maintain wild-type infectivity in skin. Chimeric viruses containing different vOka components retained the attenuation phenotype, suggesting that vOka attenuation is multi-factorial and can be produced by genes from different regions of the vOka genome. PMID- 15661166 TI - Role of HIV-2 envelope in Lv2-mediated restriction. AB - We have characterized envelope protein pseudotyped HIV-2 particles derived from two HIV-2 isolates termed prCBL23 and CBL23 in order to define the role of the envelope protein for the Lv2-mediated restriction to infection. Previously, it has been described that the primary isolate prCBL23 is restricted to infection of several human cell types, whereas the T cell line adapted isolate CBL23 is not restricted in these cell types. Molecular cloning of the two isolates revealed that the env and the gag gene are responsible for the observed phenotype and that this restriction is mediated by Lv2, which is distinct from Ref1/Lv1 (Schmitz, C., Marchant, D., Neil, S.J., Aubin, K., Reuter, S., Dittmar, M.T., McKnight, A., Kizhatil, K., Albritton, L.M., 2004. Lv2, a novel postentry restriction, is mediated by both capsid and envelope. J. Virol. 78 (4), 2006-2016). We generated pseudotyped viruses consisting of HIV-2 (ROD-ADeltaenv-GFP, ROD-ADeltaenv-RFP, or ROD-ADeltaenv-REN) and the prCBL23 or CBL23 envelope proteins as well as chimeric proteins between these envelopes. We demonstrate that a single amino acid exchange at position 74 in the surface unit of CBL23-Env confers restriction to infection. This single point mutation causes tighter CD4 binding, resulting in a less efficient fusion into the cytosol of the restricted cell line. Prevention of endosome formation and prevention of endosome acidification enhance infectivity of the restricted particles for GHOST/X4 cells indicating a degradative lysosomal pathway as a cause for the reduced cytosolic entry. The described restriction to infection of the primary isolate prCBL23 is therefore largely caused by an entry defect. A remaining restriction to infection (19-fold) is preserved when endosomal acidification is prevented. This restriction to infection is also dependent on the presence of the point mutation at position 74 (G74E). PMID- 15661167 TI - Role of recombination in the evolution of natural populations of Cucumber mosaic virus, a tripartite RNA plant virus. AB - The role of recombination in the evolution of Cucumber mosaic virus (CMV) was analyzed in a collection of Spanish isolates from 1989 to 2002. Isolates were characterized by ribonuclease protection assay using six RNA probes, two for each of the three genomic RNAs, which allowed the identification of the analyzed regions as belonging to CMV isolates in subgroups IA, IB, and II. Most isolates belonged to subgroups IA (64%) and IB (12%), 5% were reassortants among subgroups IA, IB, or II, and 17% were recombinants between these groups. Recombinants at RNA3 were significantly more frequent than recombinants at RNAs 1 and 2. One IB IA recombinant RNA3 was as frequent in central Spain as the IA RNA3. The genetic structure of the virus population suggested that reassortants and most recombinant genotypes were selected against and was consistent with a higher biological cost of reassortment than recombination. Data also suggest that recombinants that encode hybrid proteins are at a higher disadvantage than recombinants that exchange whole ORFs. PMID- 15661168 TI - Inter-subunit disulfide bonds in soluble HIV-1 envelope glycoprotein trimers. AB - Soluble forms of the trimeric human immunodeficiency virus (HIV-1) envelope glycoproteins are important tools for structural studies and in the construction of improved immunogens. We found that a substantial fraction of soluble envelope glycoprotein trimers contain inter-subunit disulfide bonds (inter-S-S bonds) that render the trimers resistant to heat and denaturing agents. These inter-S-S bonds can be reduced without disrupting the trimers by treatment with a low concentration of beta-mercaptoethanol or DTT. Antibody mapping studies suggest that the soluble HIV-1 envelope glycoprotein trimers lacking the inter-S-S bonds exhibit a conformation closer to that of the native HIV-1 envelope glycoprotein complex. However, reducing these inter-S-S bonds had only modest effects on the inefficient elicitation of neutralizing antibodies by the soluble trimers. These studies provide guidance in improving the resemblance of tractable, soluble forms of the HIV-1 envelope glycoproteins to the native virion spikes. PMID- 15661169 TI - Foot-and-mouth disease virus (FMDV) causes an acute disease that can be lethal for adult laboratory mice. AB - Foot-and-mouth disease virus (FMDV) is a picornavirus that causes an acute vesicular disease of cloven-hoofed animals. This virus continues to be threat to livestock worldwide with outbreaks causing severe economic losses. However, very little is known about FMDV pathogenesis, partially due to the inconveniences of working with cattle and swine, the main natural hosts of the virus. Here we demonstrate that C57BL/6 and BALB/C adult mice are highly susceptible to FMDV infection when the virus is administered subcutaneously or intraperitoneally. The first clinical signs are ruffled fur, apathy, humped posture, and wasting, which are followed by neurological signs such as hind-limb paralysis. Within 2-3 days of disease onset, the animals die. Virus is found in all major organs, indicating a systemic infection. Mice developed microvesicles near the basal layer of the epithelium, event that precedes the vesiculation characteristics of FMD. In addition, a lymphoid depletion in spleen and thymus and severe lymphopenia is observed in the infected mice. When these mice were immunized with conventional inactivated FMDV vaccine, they were protected (100% of vaccinated animals) against challenge with a lethal dose of FMDV. The data indicate that this mouse model may facilitate the study of FMDV pathogenesis, and the development of new effective vaccines for FMD. PMID- 15661170 TI - Virion stability and aphid vector transmissibility of Cucumber mosaic virus mutants. AB - The physical stability of virions of Cucumber mosaic virus (CMV) mutants was investigated to determine if relative stability correlated with efficiency of aphid transmission. Virion stability was evaluated by a urea disruption assay and by testing the infectivity of virus following purifications. All viruses were infectious when purified using a low salt buffer without organic solvent, whereas two of seven viruses were less stable and inactivated following purification with a high salt buffer and chloroform. These two viruses were both reassortants derived from the spontaneous transmission-defective mutant CMV-M (F1F2M3 and F1F2M3-L129P). F1F2M3 was relatively unstable, being disrupted between 0 and 1 M urea versus the wild-type CMV-Fny (F1F2M3) that was destabilized at 3-4 M urea. Modifications of F1F2M3 at three amino acid positions (129, 162, 168), singly or in combination, increased the relative stability of virions. A second class of transmission-defective CMVs with engineered mutations in the betaH-betaI surface loop of the CMV-Fny capsid protein (CP) exhibited near wild-type levels of stability. Lastly, a single Pro to Leu substitution at CP position 129 of CMV-Fny (F1F2M3-P129L) conferred the induction of necrosis in tobacco plants and reduced aphid transmissibility, but did not markedly alter the physical stability of virions. Thus, only among CMV-M derivatives harboring the CP mutation of Thr to Ala at position 162 were increases in stability correlated with restoration of transmissibility by the aphid Aphis gossypii. PMID- 15661171 TI - A reconstituted replication and transcription system for Ebola virus Reston and comparison with Ebola virus Zaire. AB - The only known filovirus, which presumably is not pathogenic for humans, is Ebola virus (EBOV) Reston. When EBOV Reston and the highly pathogenic EBOV Zaire were grown in cell culture, comparison of the replication kinetics showed a clear growth impairment of EBOV Reston, indicating that the replication cycle of EBOV Reston might be delayed. In addition, the cytopathic effect caused by the virus was much milder with EBOV Reston than with EBOV Zaire. To compare replication and transcription of EBOV Reston and Zaire, a reconstituted minigenomic replication and transcription system based on reverse genetics has been established for EBOV Reston. This system was used to exchange the EBOV Zaire and EBOV Reston nucleocapsid (NC) proteins NP, VP35, VP30, and L, which catalyze replication and transcription. Furthermore, chimeric minigenomes were constructed containing the cis-acting replication signals of EBOV Zaire combined with those of EBOV Reston. Surprisingly, the cis-acting signals as well as almost all NC proteins could be exchanged between EBOV Reston and Zaire, suggesting a high degree of functional homology of the replication/transcription complexes of EBOV Zaire and EBOV Reston. Only the combination of EBOV Zaire VP35 and EBOV Reston L did not result in replication and transcription activity. Although these two proteins did not constitute an active polymerase complex, it was shown by immunofluorescence analysis that they were still able to interact. PMID- 15661172 TI - Effects of virion surface gp120 density on infection by HIV-1 and viral production by infected cells. AB - The quantity of envelope glycoprotein molecules (Env) on HIV-1 particles is still an issue of debate and, depending on the strain of virus and the nature of the producer cells, it can vary greatly. Here, we have attempted to address how Env density influences HIV-1 fitness. To this aim, we have produced HIV-1-derived viral particles with various amounts of R5 Env (low Env: Envlo; high Env: Envhi), using a regulatable expression system. The infectivity was assayed on human cells, engineered to express the HIV receptor CD4 and the co-receptor CCR5, as well as on peripheral blood lymphocytes and macrophages. In these experiments, low levels of Env were sufficient for cell infection, albeit at low efficiency. Increasing the amount of Env resulted in cooperatively improved infectivity, but a threshold was rapidly attained, indicating that only a fraction of Env was required for efficient infection. Unexpectedly, Env incorporation beyond what gives maximal infection transiently stimulated the expression of proviral genes, as well as retrovirus production, in newly infected cells. This was likely a consequence of induced NF-kappaB activity, as this transcription factor is triggered by Envhi, but not by Envlo, virions. Thus, our data suggest that one major effect of high Env density on the surface of HIV may not be better infection yields but rather improved viral production by newly infected cells. PMID- 15661173 TI - The conserved glycine residues in the transmembrane domain of the Semliki Forest virus fusion protein are not required for assembly and fusion. AB - The alphavirus Semliki Forest virus (SFV) infects cells via a low pH-triggered fusion reaction mediated by the viral E1 protein. Both the E1 fusion peptide and transmembrane (TM) domain are essential for membrane fusion, but the functional requirements for the TM domain are poorly understood. Here we explored the role of the five TM domain glycine residues, including the highly conserved glycine pair at E1 residues 415/416. SFV mutants with alanine substitutions for individual or all five glycine residues (5G/A) showed growth kinetics and fusion pH dependence similar to those of wild-type SFV. Mutants with increasing substitution of glycine residues showed an increasingly more stringent requirement for cholesterol during fusion. The 5G/A mutant showed decreased fusion kinetics and extent in fluorescent lipid mixing assays. TM domain glycine residues thus are not required for efficient SFV fusion or assembly but can cause subtle effects on the properties of membrane fusion. PMID- 15661174 TI - Folding and dimerization of hepatitis C virus E1 and E2 glycoproteins in stably transfected CHO cells. AB - The recombinant E1E2 heterodimer of the hepatitis C virus is a candidate for a subunit vaccine. Folding analysis of E1 and E2 glycoproteins, stably expressed in CHO cells, showed that E1 folding was faster and more efficient than E2. The oxidized DTT-resistant conformation of E1 was completed within 2 h post synthesis, while E2 not only required up to 6 h but also generated non-native species. Calnexin was found to assist E1 folding, whereas no chaperone association was found with E2. The assembly of E1 and E2 was assessed by co immunoprecipitation and sedimentation velocity analysis. We found that the formation of native E1E2 heterodimers paralleled E2 oxidation kinetics, suggesting that E2 completed its folding process after association with E1. Once formed, sedimentation of the native E1E2 heterodimers was consistent with the absence of additional associated factors. Taken together, our data strongly suggest that productive folding of the major HCV spike protein E2 is assisted by E1. PMID- 15661175 TI - Calcium ions affect the hepatitis B virus core assembly. AB - Previous report showed that cytosolic Ca2+ induced by hepatitis B virus X protein (HBx) promotes HBV replication. In this study, in vitro experiments showed that (i) HBV core assembly in vitro was promoted by Ca2+ through the sucrose density gradient and the analytical ultracentrifuge analysis. Also, (ii) transmission electron microscope analysis demonstrated these assembled HBV core particles were the capsids. Ex vivo experiments showed that the treatment of BAPTA-AM and cyclosporine A (CsA) reduced HBV capsids in the transfected HepG2 cells. In addition to that, the treatment of Thapsigargin (TG) increased HBV capsids in the transfected HepG2 cells. Furthermore, we investigated the increased HBV core assembly by HBx. The results show that the increased cytosolic calcium ions by HBx promote the HBV core assembly. PMID- 15661176 TI - Creating Coordination in the Cerebellum. Proceedings of a meeting in honor of Constantino Sotelo. October 2-4, 2003, Catania, Italy. PMID- 15661177 TI - Development of the olivocerebellar system: migration and formation of cerebellar maps. PMID- 15661178 TI - The genetics of early cerebellar development: networks not pathways. PMID- 15661179 TI - Bcl-2 protection of axotomized Purkinje cells in organotypic culture is age dependent and not associated with an enhancement of axonal regeneration. PMID- 15661180 TI - Axonal and synaptic remodeling in the mature cerebellar cortex. AB - By blocking electrical activity in the cerebellar cortex the Purkinje cell dendrites become a uniform territory with a high density of spines all bearing the glutamate receptor delta2 subunit (GluRdelta2) and being mainly innervated by parallel fibers. Such a subunit, which is constitutively targeted specifically to the parallel fiber synapses, appears in the spines contacted by the climbing fibers before they disconnect from the target. A similar pattern of hyperspiny transformation and innervation occurs a few days after a subtotal lesion of the inferior olive, the source of climbing fibers. During the climbing fiber reinnervation process which follows the removal of the electrical block or by collateral sprouting of surviving inferior olive neurons, the new active climbing fibers establish synaptic contacts with proximal dendritic spines that bear GluRdelta2s. After, they repress these subunits and displace the parallel fibers to the distal dendritic territory. These findings suggest the following operational principle in the axonal competition for a common target. The Purkinje cells have an intrinsic phenotypic profile which is compatible with the parallel fiber innervation, this mode being operational in targets innervated by a single neuronal population, like the neuromuscular system. An additional input, the climbing fibers, in order to achieve its own territory on the proximal dendrite needs the ability to displace the competitor. Such an inhibition is activity dependent and the activity needs to be present in order to allow the climbing fiber to maintain its territory, even when the developmental period is over. PMID- 15661181 TI - Fate restriction and developmental potential of cerebellar progenitors. Transplantation studies in the developing CNS. AB - The generation of cell diversity from undifferentiated progenitors is regulated by interdependent mechanisms, including cell intrinsic programs and environmental cues. This interaction can be investigated by means of heterochronic/heterotopic transplantation, which allows to examine the behaviour of precursor cells in an unusual environment. The cerebellum provides an ideal model to study cell specification, because its neurons originate according to a well-defined timetable and they can be are readily recognised by morphological features and specific markers. Cerebellar progenitors transplanted to the embryonic cerebellum develop fully mature cerebellar neurons, which often integrate in the host circuitry in a highly specific manner. In extracerebellar locations, cerebellar progenitors preferentially settle in caudal CNS regions where they exclusively acquire cerebellar identities. By contrast, neocortical precursors preferentially settle in rostral regions and fail to develop hindbrain phenotypes. The phenotypic repertoire generated by transplanted cerebellar progenitors is strictly dependent on their age. Embryonic progenitors originate all mature cerebellar cells, whereas postnatal ones exclusively generate later-born types, such as molecular layer interneurons and granule cells. Together, these observations foster the hypothesis that neural progenitors are first specified towards region-specific phenotypes along the rostro-caudal axis of the neural tube. Thereafter, the developmental potential of progenitor cells is progressively restricted towards later generated types. Such a progressive specification of precursor cells in space and time is stably transmitted to their progeny and it cannot be modified by local cues, when these cells are confronted with heterotopic and/or heterochronic environments. PMID- 15661182 TI - Long-term potentiation of synaptic transmission at the mossy fiber-granule cell relay of cerebellum. AB - In the last decade, the physiology of cerebellar neurons and synapses has been extended to a considerable extent. We have found that the mossy fiber-granule cell relay can generate a complex form of long-term potentiation (mf-GrC LTP) following high-frequency mf discharge. Induction. Mf-GrC LTP depends on NMDA and mGlu receptor activation, intracellular Ca(2+) increase, PKC activation, and NO production. The preventative action of intracellular agents (BAPTA, PKC inhibitors) and of membrane hyperpolarization, and the correlated increase in intracellular Ca(2+) observed using fluorescent dyes, indicate that induction occurs postsynaptically. Expression. Expression includes three components: (a) an increase of synaptic currents, (b) an increase of intrinsic excitability in GrC, and (c) an increase of intrinsic excitability in mf terminals. Based on quantal analysis, the EPSC increase is mostly explained by enhanced neurotransmitter release. NO is a candidate retrograde neurotransmitter which could determine both presynaptic current changes and LTP. NO cascade blockers inhibit both presynaptic current changes and LTP. The increase in intrinsic excitability involves a raise in apparent input resistance in the subthreshold region and a spike threshold reduction. Together with other forms of cerebellar plasticity, mf-GrC LTP opens new hypothesis on how the cerebellum processes incoming information. PMID- 15661183 TI - Climbing fiber synaptic plasticity and modifications in Purkinje cell excitability. PMID- 15661184 TI - Bases and implications of learning in the cerebellum--adaptive control and internal model mechanism. AB - The cerebellum has a fine compartment structure, which represents various discrete functions. Evolutionarily old, medial parts of the cerebellum are involved in the adaptive control of various brainstem and spinal cord functions, and long-term depression (LTD) plays a key role in this adaptive control. To extend these views to evolutionarily newer, lateral parts of the cerebellum involved in cerebral cortical functions such as voluntary movement, perception and language, the internal model hypothesis is instrumental. This hypothesis explains not only the characteristic cerebellar symptom, dysmetria, but also a number of otherwise unexplainable phenomena displayed in movements and mental actions. PMID- 15661185 TI - Synaptic transmission and long-term depression in Purkinje cells in an in vitro block preparation of the cerebellum isolated from neonatal rats. PMID- 15661186 TI - Optical imaging of cerebellar functional architectures: parallel fiber beams, parasagittal bands and spreading acidification. PMID- 15661187 TI - Imaging cerebellum activity in real time with magnetoencephalographic data. AB - The cerebellum has traditionally been associated with motor movements but recent studies suggest its involvement with fine timing, sensory analysis and cognition. Much of the new data comes from neuroimaging techniques such as fMRI and PET, which have high spatial resolution and show that for even simple stimuli many cerebellar and cortical areas are involved. We use examples from recent studies to demonstrate that magnetic field tomography (MFT) offers a new and powerful tool for studying cerebellar function through real time localization of cortical, brainstem and cerebellar activations over timescales ranging from a fraction of a millisecond to seconds, minutes and hours. The examples include demonstration of cerebellar activations along well-established anatomical pathways during saccades and the visualization of the ascending medullar volley after median nerve stimulation. MFT analysis of single trial MEG signals elicited by the presentation of faces in emotion and object recognition tasks, show changes in cerebellar activation between schizophrenics and normal subjects in agreement with proposals for disturbed cerebellar function in schizophrenia. The ability of MFT to identify cerebellar, brainstem and cortical activations in real time can add new insights about dynamics of brain activity to the recent findings about cerebellar function from PET and fMRI. PMID- 15661188 TI - The cerebellum in the cerebro-cerebellar network for the control of eye and hand movements--an fMRI study. AB - The coordination of optical information and manipulation of objects in space by eye and hand movements is controlled by a cerebro-cerebellar network. The differential influence of prefrontal, motor, or parietal areas in combination with cerebellar areas, especially within the posterior hemispheres, on the control of eye and hand movements is not very well defined. Using fMRI we investigated the functional representation of isolated or combined eye and hand movements within the cerebellum and the impact of differential cognitive preload on the activation patterns. Each task consisted of the performance of saccades or hand movements triggered by a cue presented on a screen in front of the scanner. Saccades were tested for visually guided saccades, triple step saccades, and for visuospatial memory. Sequential finger opposition movements were tested for predictive and nonpredictive movements. Combined and isolated eye-hand reaching movements were tested toward a target presented in 5 different horizontal positions. Visually guided saccades activated the cerebellar vermis lobuli VI VII, triple step saccades, including visuospatial memorization, in addition the cerebellar hemispheres lobuli VII-VIII. Sequential finger movements and reaching movements activated a cerebellar network consisting of the lobuli IV-VI, the vermis, and the lobuli VII-VIII with broader areas and additional regions especially within the lobus VII for more complex movements. The combined in contrast to the isolated performance of eye and hand movements demonstrated specialized activation foci within the cerebellar vermis and posterior hemispheres. We could demonstrate a differential representation of eye and hand movements within the cerebellum. Additional "cognitive" preload within a given task leads to additional activation of the posterior cerebellar hemispheres, with a subspecialization corresponding to premotor and parietal area connections. PMID- 15661189 TI - Fast oscillation in the cerebellar cortex of calcium binding protein-deficient mice: a new sensorimotor arrest rhythm. AB - Fast oscillations (>100 Hz) may serve physiological roles when regulated properly. They may also appear in pathological conditions. In cerebellum, 160 Hz oscillation emerge in mice lacking calbindin and/or calretinin, two proteins devoted to calcium buffering in Purkinje and granule cells, respectively. Here, we review the pharmacological and spatiotemporal properties of this fast cerebellar oscillation and the related Purkinje cell firing behaviour in alert mice. We show that this oscillation is highly synchronized along the parallel fiber beam and reversibly inhibited by gap junctions, GABA(A) and NMDA receptors blockers. Cutaneous stimulation of the whisker region transiently suppressed the oscillation which shows in some aspects similarities with cerebral "resting" rhythmic activities of wakefulness arresting to sensory or motor information such as alpha and mu rhythms. The Purkinje cells of these mutants present an increased simple spike firing rate, rhythmicity and synchronicity, and a decreased complex spike duration and subsequent pause. Both simple and complex spikes may be tightly phase-locked with the oscillation. Contrastingly, on slice recordings, the intrinsic membrane properties of Purkinje cell are similar in wild type mice and in mice lacking calbindin. The role played by this fast cerebellar oscillation in the emergence of ataxia is yet to be solved. PMID- 15661190 TI - Oscillations in the cerebellar cortex: a prediction of their frequency bands. AB - Local recurrent connections endow the cerebellar cortex with an intrinsic dynamics. We performed computer simulations to predict the frequency bands of the oscillations that will most likely emerge. Feedback inhibition from the Golgi to the granule cells induced 10-50 Hz oscillations, the period at resonance being approximately equal to four times the maximum conduction delay generated along the parallel-fiber connections from granule to Golgi cells. In the molecular layer, the interneurons tended to induce fast oscillations (100-250 Hz), having a period equal to about four times the delay over their reciprocal synaptic connections. Finally, although the presence of lateral inhibition among the Purkinje cells has not been firmly established, reciprocal Purkinje-cell synapses are predicted to transform the cerebellar cortex into a potential temporal integrator. PMID- 15661191 TI - Gap junctions synchronize synaptic input rather than spike output of olivary neurons. AB - Electronic coupling in the inferior olive is supposed to underlie the synchrony of complex spike activities of Purkinje cells in the cerebellar cortex. Here we show a computational model which suggests that the olivary gap junctions may synchronize the input rather than the neuronal output. As such, coupling may influence the absolute moment in time of the complex spike activity rather than their synchrony. PMID- 15661192 TI - Is the cerebellum ready for navigation? AB - Spatial navigation required the acquisition of at least two complementary processes: the organization of the spatial representation of the environment (declarative learning) and the acquisition of a motor behaviour adapted to the specific context (procedural learning). The potential role of the cerebellum in spatial navigation is part of the debate concerning its role in cognitive function. Experiments ranging from cerebellar patients to animal models have indicated that cerebellar damage affects the processing of spatial information. The main unresolved issue concern the interpretation of these deficits. Is the cerebellum involved in both declarative and procedural components of navigation? Could all deficits in navigation paradigms be interpreted by a deficit in a motor dependant process? The purpose of this review is to examine different results coming from anatomical data, experimental paradigms and models in order to give a critical answer to this question. PMID- 15661193 TI - The lateral cerebellum and visuomotor control. AB - The lateral cerebellum receives an abundance of visual input providing the link between visual and motor control centers. In this review we discuss experiments designed to increase our understanding of how visual inputs to the cerebellum are arranged in relation to the zonal organization of the cerebellar cortex, and how visual inputs are utilized to assist in the regulation of a visually guided movement. On the basis of anatomical and physiological characteristics our findings indicate that the medial-most folium in crus I of the cat lateral cerebellum can be subdivided into at least three functionally distinct zones; from lateral to medial along the length of the folium these correspond to zones D(1), lateral C(3) and C(2). Each zone displays clear differences in olivo cortico-nuclear connectivity and in the anesthetized animal zones D(1) and C(2) both receive powerful visual inputs relayed via the climbing fiber system. Complementary experiments in awake behaving cats found that Purkinje cells located in the D(1) and D(2) zones of crus I exhibit changes in simple spike discharge time locked to target motion during a visually guided reaching task. These changes were unaffected by temporary visual denial of the target, raising the possibility that internally generated feedforward visuomotor control mechanisms are operating, in which a predictive model of the target's motion has been constructed by the CNS. PMID- 15661194 TI - Coupling of hand and foot voluntary oscillations in patients suffering cerebellar ataxia: different effect of lateral or medial lesions on coordination. AB - Motor coordination has been investigated in seven ataxic patients who underwent surgery of the cerebellar hemisphere (4) or of the vermis-paravermis region (3). Subjects, tested ipsilaterally to the lesion, were asked to couple in-phase rhythmic oscillations of the prone hand and the ipsilateral foot for at least 10 s. The oscillation frequency, paced by a metronome, ranged 0.8-3 Hz. Hand and foot angular displacements were measured by a potentiometric technique; EMG from Extensor Carpi Radialis and Tibialis Anterior was recorded by surface electrodes. The phase-relations between the hand and foot movements, as well as between the onsets of motor commands, were calculated. For each of the limbs the frequency response curve was estimated by plotting the mean phase values between the onset of the motor command and the onset of the related movement. The experiment was repeated with the same schedule after a strong artificial increase of the hand inertial momentum (15 g m(2)). In the unloaded condition, all patients failed to achieve a hand-foot synchrony (0 degrees ), the hand movement showing a net phase lag. In four hemispheric and one vermian patients (group 1) this lag progressively grew with frequency up to 110 degrees , in the other two vermian patients (group 2) the hand lag kept almost constant ( approximately 45 degrees ). Group 1 subjects were unable to adequate the delay between the motor commands to the increase in frequency, as instead did group 2 subjects, although this was insufficient to produce movement synchrony. Subjects reacted to hand loading with different strategies. In group 1, due to the net increase of hand inertia, movement synchrony required a strong advance of the hand motor command. Patients succeeded in this, but because of their inability to compensate for changes in frequency, they still produced a progressive lag between movements. In group 2, loading strongly increased the hand dynamic stiffness while it slightly lowered that of the foot, resulting in a rather small difference between mechanical properties of the limbs. Thus, compensation required only a slight anticipatory activation of the hand motor command. Patients failed to do so, however they were able to adjust the command delay to the required frequency and produced a constant hand lag. Their main motor handicap was found to to be the incapability of judging the hand lag as a lack of synchrony. These results seems to indicate that the cerebellum must be involved both in measuring the time difference between hand and foot movements and in weighting this delay in function of the oscillation frequency. These two processes may be confined to the vermis paravermis region and to the hemisphere, respectively. PMID- 15661195 TI - Modulation of cutaneous reflexes in hindlimb muscles during locomotion in the freely walking rat: A model for studying cerebellar involvement in the adaptive control of reflexes during rhythmic movements. AB - This study aims to demonstrate stepphase-dependent modulation in the gain of cutaneously triggered reflexes in the freely locomoting rat. Electromyographic recordings of biceps femoris (mainly involved in knee flexion) and gastrocnemius (mainly involved in ankle extension) muscles were continuously monitored during locomotion and cutaneous reflexes were induced by subcutaneously placed stimulation electrodes in the lateral malleolal region. The results show that the reflex responses in both muscles during locomotion were generally reduced compared to reflexes induces in rest. For the biceps femoris reduction of reflex gain was highest during the stance phase whereas for the gastrocnemius the period of highest depression was found during the swing phase. We conclude that stepphase-dependent modulation of peripheral reflexes can be measured in freely locomoting rats and generally concur with previous studies in cat and man that this type of modulation may be functionally important for maintaining and adjusting gait. Moreover, although the mechanism of inducing and maintaining this modulation is not fully known, it is now open to experimental investigation in rodents. PMID- 15661196 TI - The basilar pontine nuclei and the nucleus reticularis tegmenti pontis subserve distinct cerebrocerebellar pathways. AB - Previous studies often considered the basilar pontine nuclei (BPN) and the nucleus reticularis tegmenti pontis (NRTP) as relays of a single cerebro-(ponto) cerebellar pathway. Conversely, the different cortical afferences to the BPN and the NRTP, as well as the anatomical and functional features of the cerebellopetal projections from these pontine nuclei, support the different, and for some aspect, complementary arrangement of the cerebrocerebellar pathways relayed by the BPN or NRTP. Both the BPN and the NRTP are innervated from the cerebral cortex, but with regional prevalence. The NRTP is principally innervated from motor or sensori-motor areas while the BPN are principally innervated from sensory, mainly teloceptive, and associative area. Projections from sensory-motor areas were also traced to the BPN. The BPN and NRTP project to all parts of the cerebellar cortex with a similar pattern. In fact, from single areas of them projections were traced to set of sagittal stripes of the cerebellar cortex. In variance to such analogies, the projections to the cerebellar nuclei differed between those traced from the NRTP and from BPN. In fact, BPN and NRTP have private terminal areas in the cerebellar nuclei with relatively little overlaps. The BPN innervated the lateroventral part of the nucleus lateralis and the caudoventral aspect of the nucleus interpositalis posterioris. The NRTP principally innervated the mediodorsal part of the nucleus lateralis, the nucleus interpositalis anterioris, the nucleus medialis. Since the single cerebellar nuclei have their specific targets in the extracerebellar brain areas, it follows that the BPN and the NRTP, passing through their cerebellar nuclei relays, are devoted to control different brain areas and thus likely to play different functional roles. From single pontine regions (of both BPN and NRTP) projections were traced to the cerebellar cortex and to the cerebellar nuclei. In some cases these projections reached areas which are likely anatomically connected (by Purkinje axons). This pattern of the pontine projections was termed as coupled projection. In some other cases, the projections reached areas of the cerebellar cortex but not the nuclear regions innervated by them. We termed this as uncoupled projection. The existence of both coupled and uncoupled projections, open new vistas on the functional architecture of the pontocerebellar pathway. More in detail, this study showed the different quantitative and topographic distribution of the coupled and uncoupled projections visualized in the cerebellar projections from BPN and NRTP. All these evidences strongly support the anatomical and the functional differences that characterise the cerebrocerebellar pathways relayed by the BPN and the NRTP. PMID- 15661197 TI - Conservation of the architecture of the anterior lobe vermis of the cerebellum across mammalian species. PMID- 15661198 TI - Pharmacology of the metabotropic glutamate receptor mediated current at the climbing fiber to Purkinje cell synapse. AB - Different forms of synaptic plasticity in the cerebellum are mediated by metabotropic glutamate receptors (mGluRs). At parallel fiber (PF) to Purkinje cell (PC) synapses activation of mGluR gives rise to a well known slow synaptic current inhibited by antagonists of mGluR1. The distribution of mGluR types in the climbing fiber (CF) to PC synapses is not well known. However, a mGluR1alpha mediated all-or-none postsynaptic current was also demonstrated at the CF-PC synapse (Dzubay and Otis, Neuron 36, 1159, 2002). Using whole cell patch-clamp recording from PCs in rat cerebellar slices with AMPA receptors blocked and glutamate uptake impaired we demonstrate a more complex pharmacology of a current obtained by single or train CF stimulation. The mGluR1 specific antagonist CPCCOEt in a group of cells suppressed this response while in a similar number of other cells it induced a potentiating effect. The antagonists of mGluR groups II and III (LY341495 and MSOP, respectively) predominantly suppressed the current. The ambiguous effect of CPCCOEt was checked by measuring the paired-pulse depression of the CF EPSC, which was not changed with the antagonist in normal as well as in low (0.5 mM) external Ca(2+) (used to prevent saturation of AMPARs), thus excluding a presynaptic effect. However, CPCCOEt induced a rise in the amplitude (by approximately 50%) as well as a prolongation (p<0.05) of the decay time of CF EPSCs at normal 2 mM Ca(2+), i.e. under conditions of AMPAR saturation, thus indicating an effect of postsynaptic origin. In 0.5 mM Ca(2+) the decay of CF EPSCs was longer but it was also significantly prolonged (p?0.01) by CPCCOEt. However, the CF EPSC amplitude was not significantly affected indicating an underlying Ca(2+)-dependent mechanism. Thus, the pharmacology of the PC mGluR-mediated response points to a dual postsynaptic role of mGluR1 giving rise to a slow postsynaptic current but also regulating other presumably mGluR-dependent currents via second messenger molecules and Ca(2+). The additional electrophysiological role of mGluR II & III types was also indicated. Such a complex regulatory mechanism may have an important role in the mGluR dependent forms of homosynaptic plasticity and motor learning at the CF-PC synapse. PMID- 15661199 TI - Nicotinic receptor modulation of neurotransmitter release in the cerebellum. AB - Nicotinic ACh receptors (nAChRs) are formed by pentameric combinations of alpha and beta subunits, differentially expressed throughout the central nervous system (CNS), where they have been shown to play a role in the modulation of neurotransmitter release. nAChRs are also important during neuronal differentiation, regulating gene expression and contributing to neuronal pathfinding. The cerebellum, which is involved in the maintenance of balance and orientation as well as refinement of motor action, in motor memory and in some aspects of cognition, undergoes a significant process of development and maturation of its neuronal networks during the first three postnatal weeks in the rat. Autoradiographic as well as in situ hybridization and immunocytochemical studies have shown that several nicotinic receptor binding sites and subunits are expressed in the rat cerebellum from embryonic stage through to adulthood, with the highest expression levels seen during the development of the cerebellar cortex. A diffuse cholinergic afferent projection to all lobules of the cerebellar cortex has been described, with the uvulanodulus, flocculus and lobules I and II of the anterior vermis regions receiving a particularly dense projection. Low levels of nAChR subunit transcripts and immunoreactivity, particularly during adulthood, and the scattered distribution of immunoreactivity between neurons in the cerebellar cortex, can explain the difficulty in assessing electrophysiologically the presence of functional nAChRs in the cerebellar cortex and some contradictory results reported in the early-published papers. In recent years, several groups have shown that also in the cerebellum different nAChR subtypes modulate release of glutamate and GABA at different synapses. The possible role of these mechanisms in synaptic consolidation during development, as well as on plasticity phenomena and network activity at mature synapses, are discussed. PMID- 15661200 TI - Role of calcium binding proteins in the control of cerebellar granule cell neuronal excitability: experimental and modeling studies. AB - Calcium binding proteins, such as calretinin, are abundantly expressed in distinctive patterns in the central nervous system but their physiological function remains poorly understood. Calretinin is expressed in cerebellar granule cells which provide the major excitatory input to Purkinje cells through parallel fibers. Calretinin deficient mice exhibit dramatic alterations in motor coordination and in Purkinje cell firing recorded in vivo through unknown mechanisms. In the present paper, we review the results obtained with the patch clamp recording techniques in acute slice preparation. This data allow us to investigate the effect of a null mutation of the calretinin gene on the intrinsic electroresponsiveness of cerebellar granule cells at a mature developmental stage. Calretinin deficient granule cells exhibit faster action potentials and generate repetitive spike discharge showing an enhanced frequency increase with injected currents. These alterations disappear when 0.15 mM of the exogenous fast calcium buffer BAPTA is infused in the cytosol to restore the calcium buffering capacity. Furthermore, we propose a mathematical model demonstrating that the observed alterations of granule cell excitability can be explained by a decreased cytosolic calcium buffering capacity due to the absence of calretinin. We suggest that calcium binding proteins modulate intrinsic neuronal excitability and may therefore play a role in the information processing in the central nervous system. PMID- 15661201 TI - Between in and out: linking morphology and physiology of cerebellar cortical interneurons. AB - We used the juxtacellular recording and labeling technique of Pinault (1996) in the uvula/nodulus of the ketamine anesthetized rat in an attempt to link different patterns of spontaneous activity with different types of morphologically identified cerebellar cortical interneurons. Cells displaying a somewhat irregular, syncopated cadence of spontaneous activity averaging 4-10 Hz could, upon successful entrainment and visualization, be morphologically identified as Golgi cells. Spontaneously firing cells with a highly or fairly regular firing rate of 10-35 Hz turned out to be unipolar brush cells. We also found indications that other types of cerebellar cortical neurons might also be distinguished on the basis of the characteristics of their spontaneous firing. Comparison of the interspike interval histograms of spontaneous activity obtained in the anaesthetized rat with those obtained in the awake rabbit points to a way whereby the behaviorally related modulation of specific types of interneurons can be studied. In particular, the spontaneous activity signatures of Golgi cells and unipolar brush cells anatomically identified in the uvula/nodulus of the anaesthetized rat are remarkably similar to the spontaneous activity patterns of some units we have recorded in the flocculus of the awake rabbit. The spontaneous activity patterns of at least some types of cerebellar interneurons clearly have the potential to serve as identifying signatures in behaving animals. PMID- 15661202 TI - Sexual dimorphism in cerebellar structure, function, and response to environmental perturbations. AB - Sexual dimorphism of CNS structure and function has been observed in humans and animals, but remains relatively unrecognized in the context of the cerebellum. Recent research in our laboratory has examined whether these gender differences extend to cerebellar structure and function, as well as the impact of environmental factors on the developing cerebellum. Perinatal exposure to both chemical and physical perturbations in the environment (in our experiments, PCBs or hypergravity) affects growth, neurodevelopment, and motor coordination differently in males and females. These neurodevelopmental and behavioral effects are accompanied by sex-related changes in cerebellar mass and cerebellar protein expression. Exposure to chemical toxins (PCBs) resulted in more dramatic neurodevelopmental and behavioral changes in male neonates. It is possible that gender-related differences in male and female cerebellar structure and function are related to sex-specific development of the cerebellum and sex-specific distribution of specific receptors, local synthesis of trophic factors, and maturation of the pituitary hypophesial axis. These sex-related differences may underlie the sex-specific preponderance of certain neuropsychiatric disorders, and must be incorporated in the design of future basic and clinical investigations. PMID- 15661203 TI - Cerebellar dysfunction in multiple sclerosis: evidence for an acquired channelopathy. AB - Cerebellar dysfunction in multiple sclerosis (MS) is a significant contributor to disability, is relatively refractory to symptomatic therapy, and often progresses despite treatment with disease-modifying agents. Thus, there is a need for better understanding of its pathophysiology. This chapter reviews a growing body of evidence which suggests that mis-tuning of Purkinje cells, due to expression of an abnormal repertoire of sodium channels, contributes to cerebellar deficits in MS. Within the normal nervous system, sodium channel Na(v)1.8 is expressed in a highly specific manner within spinal sensory and trigeminal neurons, and is not present within Purkinje cells, Na(v)1.8 mRNA and protein are, however, expressed within Purkinje cells both in models of MS (experimenal autoimmume encephalomyelitis; EAE), and in postmortem tissue from humans with MS. Expression of Na(v)1.8 within Purkinje cells in vitro alters electrogenesis in these cells in several ways: first, by increasing duration and amplitude of action potentials; second, by decreasing the proportion of action potentials that are conglomerate and the number of spikes per conglomerate action potential; and third, by supporting sustained, pacemaker-like impulse trains in response to depolarization, which are not seen in the absence of Na(v)1.8. Similar changes are observed in recordings from Purkinje cells in vivo from mice with EAE. Taken together, these results suggest that expression of Na(v)1.8 within Purkinje cells distorts their pattern of firing in MS. PMID- 15661204 TI - Don't get too excited: mechanisms of glutamate-mediated Purkinje cell death. AB - Purkinje cells (PCs) present a unique cellular profile in both the cerebellum and the brain. Because they represent the only output cell of the cerebellar cortex, they play a vital role in the normal function of the cerebellum. Interestingly, PCs are highly susceptible to a variety of pathological conditions that may involve glutamate-mediated 'excitotoxicity', a term coined to describe an excessive release of glutamate, and a subsequent over-activation of excitatory amino acid (NMDA, AMPA, and kainite) receptors. Mature PCs, however, lack functional NMDA receptors, the means by which Ca(2+) enters the cell in classic hippocampal and cortical models of excitotoxicity. In PCs, glutamate predominantly mediates its effects, first via a rapid influx of Ca(2+)through voltage-gated calcium channels, caused by the depolarization of the membrane after AMPA receptor activation (and through Ca(2+)-permeable AMPA receptors themselves), and second, via a delayed release of Ca(2+) from intracellular stores. Although physiological levels of intracellular free Ca(2+) initiate vital second messenger signaling pathways in PCs, excessive Ca(2+) influx can detrimentally alter dendritic spine morphology via interactions with the neuronal cytoskeleton, and thus can perturb normal synaptic function. PCs possess various calcium-binding proteins, such as calbindin-D28K and parvalbumin, and glutamate transporters, in order to prevent glutamate from exerting deleterious effects. Bergmann glia are gaining recognition as key players in the clearance of extracellular glutamate; these cells are also high in S-100beta, a protein with both neurodegenerative and neuroprotective abilities. In this review, we discuss PC-specific mechanisms of glutamate-mediated excitotoxic cell death, the relationship between Ca(2+) and cytoskeleton, and the implications of glutamate, and S-100beta for pathological conditions, such as traumatic brain injury. PMID- 15661206 TI - Untreated cervical cancer. PMID- 15661207 TI - Untreated cervical cancer in the United States. AB - OBJECTIVE: To evaluate treatment patterns, including lack of treatment, among women diagnosed with cervical cancer in the United States. METHODS: Using the National Cancer Institute's (NCI's) Surveillance, Epidemiology, and End Results (SEER) program, we identified 13,715 women diagnosed with invasive cervical cancer between 1992 and 1999 and eligible for inclusion in the study. RESULTS: Nearly 9% of women diagnosed with invasive cervical cancer received no therapy for their disease. Lack of therapy was associated with a later stage of disease at diagnosis, older age, and unmarried status. More than 16% of women aged 65 and older with stage IIB/IV cervical cancer received no therapy for their disease. CONCLUSION: We must educate women diagnosed with cervical cancer and their families about the importance of treatment for potential cure and control of symptoms. We must identify and overcome obstacles that may prevent adherence to treatment recommendations. These may include comorbidity, access to cancer treatment, inability to pay for treatment, and inadequate social support. PMID- 15661208 TI - Vascular anomalies in the paraaortic region diagnosed by laparoscopy in patients with gynaecologic malignancies. AB - OBJECTIVE: Paraaortic infrarenal lymphadenectomy is indicated in patients with gynecologic tumors of high metastasising potential and can be done successfully by laparoscopic approach. Vascular anomalies in this region are incidental findings during these approaches and may increase the surgical complication rate. In this study, we have documented the frequency and pattern of the vascular anomalies in paraaortic region intraoperatively and on cadavers in an attempt to increase surgical safety. METHODS: A total of 86 consecutive patients underwent laparoscopic infrarenal paraaortic lymphadenectomy by a standardised technique between 1st of January 2002 and 1st of March 2004. Of the 86, 52 were primary cervical, 5 recurrent cervical, 14 endometrial, 14 early ovarian and 1 vulvar tumor with positive groin and pelvic lymph nodes. In the same time, anatomical dissections of the paraaortic region on 18 cadavers were performed at the Institute of Anatomy. RESULTS: Arterial or venous abnormalities were identified in 30.2% (26/86) of patients by laparoscopy. The most frequent anomalies were related to atypical renal arteries (pole arteries-9 patients) and an abnormal course of lumbar veins directly draining in the left renal vein (15 patients). In one of the patients, the complete left renal vein went retroaortic to the inferior vena cava. In cadaveric dissections, vascular anomalies were noted in 44.4% (8/18) which included variations in renal and lumbar vessels and ovarian vessels. Duplicated inferior vena cava was the least common anomaly and was detected in only one case. CONCLUSION: During laparoscopic paraaortic inframesenteric and infrarenal lymphadenectomy, care must be taken because of possible abnormalities in arterially and venous system to avoid massive hemorrhage, transfusion and conversion to laparotomy. PMID- 15661209 TI - Does radical trachelectomy influence uterine blood supply? AB - OBJECTIVE: Radical trachelectomy in combination with pelvic and parametric lymphadenectomy is indicated in young patients with early cervical cancer and planned pregnancy. If pregnancy occurs, premature delivery is a known problem in these patients. We evaluated if uterine blood supply is decreased after radical trachelectomy as one of various possible causes of preterm birth. METHODS: Between October 2003 and April 2004, 14 consecutive patients with early cervical cancer underwent radical trachelectomy with pelvic and parametric lymphadenectomy. The uterine blood supply was measured as resistance index (RI) by Doppler sonography pre- and postoperatively. Doppler sonography of the uterine artery was also performed in 14 healthy students as a control cohort. RESULTS: Fourteen patients with histologically confirmed adenocarcinoma or squamous carcinoma of the cervix uteri stage Ia1 L1 to 1b1 underwent radical trachelectomy. Mean age of patients was 33.4 years (31-37). On average, 11.5 mm (5-23) of cervical length and 24.6 (14-35) tumor-free lymph nodes were removed. Decrease of RI of the uterine artery was 0.06 on the right side (0.76-0.70) and 0.07 (0.75-0.68) on the left side. The absolute RI values after radical trachelectomy were not different compared to the values in the control group (0.76 versus 0.70 right side, 0.74 versus 0.68 left side). CONCLUSION: Uterine perfusion after radical trachelectomy with pelvic and parametric lymphadenectomy remains unchanged. PMID- 15661210 TI - Expression of Bcl-xL in ovarian carcinoma is associated with chemoresistance and recurrent disease. AB - OBJECTIVE: The long-term survival of patients with epithelial ovarian cancer is limited by the emergence of tumor cells that are resistant to chemotherapy. We hypothesized that expression of Bcl-x(L), a homologue of Bcl-2 that confers protection from chemotherapy-induced apoptosis, may be predictive of patients' clinical response to treatment, and that treatment with chemotherapy may result in the selection of tumor cells that overexpress this protein. METHODS: We determined the expression of Bcl-x(L) in epithelial ovarian cancers from 28 patients at the time of initial staging laparotomy and in recurrent tumors in the same patients following treatment with platinum-based chemotherapy. The data were analyzed to determine whether Bcl-x(L) expression was predictive of clinical outcome. A2780 ovarian cancer cells were stably transfected with Bcl-x(L) or control plasmid. Chemotherapy-induced apoptosis in these cell lines was determined in vitro and in a xenograft model. RESULTS: Bcl-x(L) expression in primary tumors was associated with a significantly shorter disease-free interval as compared to patients whose tumors did not express Bcl-x(L) (1.6 months as compared to 7.7 months). We found that Bcl-x(L) expression conferred resistance to chemotherapy-induced apoptosis resulting from treatment with cisplatin, paclitaxel, topotecan, and gemcitabine in vitro. In a xenograft model, Bcl-x(L) expressing tumors continued to grow following treatment with cisplatin, paclitaxel, topotecan, and gemcitabine, in contrast to control tumors, which disappeared. CONCLUSION: These results portray an important role for Bcl-x(L) as a key factor associated with chemotherapy failure in the treatment of ovarian cancer. PMID- 15661212 TI - Role of CT and MR imaging in predicting optimal cytoreduction of newly diagnosed primary epithelial ovarian cancer. AB - OBJECTIVE: To determine the relative accuracy of CT or MR imaging in the detection of inoperable tumor sites prior to cytoreductive surgery in a large series of patients with newly diagnosed primary epithelial ovarian cancer. METHODS: One hundred thirty-seven women with newly diagnosed primary epithelial ovarian cancer underwent CT (n = 91) or MR imaging (n = 46) prior to cytoreductive surgery. The following imaging criteria were used to identify inoperable tumor sites: (1) peritoneal implants greater than 2 cm in maximum diameter in the porta hepatis, intersegmental fissure, gall bladder fossa, subphrenic space, gastrohepatic ligament, gastrosplenic ligament, lesser sac, or root of the small bowel mesentery; (2) retroperitoneal adenopathy greater than 2 cm in maximum diameter above the renal hila; (3) hepatic metastases or abdominal wall invasion. Imaging results were compared with operability at surgery. RESULTS: Cytoreductive surgery was suboptimal in 21 of the 137 (15%) patients. Sixteen of these patients had inoperable tumor on preoperative imaging, while one additional patient had apparently inoperable tumor on imaging but was optimally debulked at surgery. The sensitivity, specificity, positive predictive value, and negative predictive value of preoperative imaging for the prediction of suboptimal debulking were 76% (16/21), 99% (115/116), 94% (16/17), and 96% (115/120), respectively. CT and MR imaging were equally effective (P = 1.0) in the detection of inoperable tumor. CONCLUSION: Preoperative CT and MR imaging are equally accurate in the detection of inoperable tumor and the prediction of suboptimal debulking in newly diagnosed epithelial ovarian cancer. This suggests imaging may help select patients who might be more appropriately managed by neoadjuvant chemotherapy. PMID- 15661211 TI - A phase II trial of weekly paclitaxel and every 3 weeks of carboplatin in potentially platinum-sensitive ovarian and peritoneal carcinoma. AB - OBJECTIVE: Paclitaxel administered weekly in equal cumulative doses is associated with less hematologic and non-hematologic toxicity than an every 3-week administration. We studied weekly paclitaxel and 3-week carboplatin in potentially platinum-sensitive recurrent ovarian and peritoneal carcinoma. METHODS: Paclitaxel at a dose of 80 mg/m(2) over 1 h in combination with carboplatin at an AUC of 5 was administered on day 1. Subsequent paclitaxel doses, modified based on the day of treatment ANC, were administered on days 8 and 15. Paclitaxel dose reductions to 75% of prior dose were performed for chemotherapy delays or toxicity. RESULTS: Twenty-eight patients were studied. The median age was 59 (range 42-80). The median platinum-free interval was 12 months (range 7-129 months). A median of six courses (range 1-13) was administered. Paclitaxel dose reductions to 60 mg/m(2) were required in 85% of the patients. Grades 3 and 4 thrombocytopenia were seen in 5 and 0 patients, respectively. Grades 3 and 4 neutropenia were seen in 14 and 1 patients, respectively. One patient was hospitalized for neutropenic fever. Twenty of 26 (77%) evaluable patients have responded with 15 patients (58%) achieving a complete response. CONCLUSIONS: Weekly paclitaxel at a dose of 60 mg/m(2) in combination with carboplatin at an AUC of 5 is well tolerated and active in potentially platinum sensitive recurrent ovarian and peritoneal carcinoma. PMID- 15661213 TI - Twenty-five year outcome of sequential abdominal radiotherapy and melphalan:implications for future management of epithelial carcinoma of the ovary. AB - OBJECTIVE: The purpose of the present study is evaluation of the long-term efficacy of sequential abdominopelvic radiotherapy and melphalan in the management of ovarian carcinoma. METHODS: From 1970 to 1976, 94 women with stages I-III epithelial ovarian carcinoma enrolled in a prospective nonrandomized clinical trial were prescribed 20 Gy to the upper abdomen and 50 Gy to the pelvis followed by courses of melphalan (1 mg/kg/course). Primary endpoints were survival, recurrence, and toxicity. RESULTS: There were 19 stage I, 25 stage II, and 50 stage III patients. For all stages, overall survival was 42% at 5 years, 30% at 10 years, and 17% at 25 years. Median follow-up of the survivors was 24 years. Disease-free survival was 48% at 5 years and remained at 45% from 10 to 25 years. All but two recurrences occurred within the first 27 months. No recurrence or treatment-related death occurred after 8 years. No recurrence was salvaged. All but one initial recurrence was within the peritoneal cavity. Of the 31 patients undergoing a second-look surgical procedure, 84% were free of tumor. Only 8% of patients recurred after a negative second look. Stage and the presence of palpable postoperative disease were significant prognostic factors. Disease free survivals were 95% from 5 to 25 years for stage I, 70% at 5 years and 60% at 25 years for stage II, and 20% from 5 to 25 years for stage III (P < 0.0001). Although no patient with postoperative palpable tumor was cured, 25% lived beyond 2 years. Stage III patients without postoperative palpable tumor achieved a 47% 25-year disease-free survival. Acute toxicity was acceptable, and 98% of patients completed radiation therapy. Chronic toxicity included a 12% small bowel obstruction rate and a 3% fatal second malignancy/hematological toxicity rate (two cases of acute myelocytic leukemia, one case of thrombocytopenia). CONCLUSIONS: The long-term disease-free survival obtained with abdominopelvic radiotherapy followed by single alkylating agent chemotherapy has not been exceeded by three subsequent decades of multiagent chemotherapy trials. Abdominal radiotherapy may be useful to consolidate complete responses following therapy multiagent chemotherapy, particularly with the upper abdominal dose escalation provided by intensity modulated radiation therapy and possibly in conjunction with chemotherapy. PMID- 15661214 TI - Matrix metalloproteinase 1 gene polymorphism as a prognostic predictor of invasive cervical cancer. AB - OBJECTIVES: Whereas human papillomavirus (HPV) infection is the major determinant of cervical carcinogenesis, host genetic factors may confer individual susceptibility and prognosis. Matrix metalloproteinase 1 (MMP-1) is an important modulator of carcinogenesis. A guanine insertion (2G) polymorphism at nucleotide 1607 of the MMP-1 gene promoter creates an Ets-1-binding site, which increases transcription activity. The present study investigates the association between MMP-1 polymorphism and cervical neoplasia, and their prognostic significance. METHODS: In this study, the MMP-1 polymorphism was assessed in 135 high-grade squamous intraepithelial lesions (HSILs) and 197 invasive squamous cell carcinomas (SCCs), and in age-matched controls, by capillary electrophoresis. The association of clinicopathological factors and HPV status with MMP-1 genotypes was tested. RESULTS: Frequencies of the 2G allele in HSIL and SCC were 64% and 65%, respectively, which did not differ significantly from control values (66% and 64%, respectively). The 2G allele was associated with advanced stages of disease (P = 0.03), whereas the G allele was more common in patients with regional lymph node metastases (P = 0.02). The survival time in patients with the heterozygous genotype G/2G (median, 55.3 months) was significantly longer than those with either the G/G (50.3 months) or 2G/2G genotype (43.9 months) (P = 0.02). No significant correlation between HPV status and MMP-1 genotype was identified. CONCLUSIONS: The genetic polymorphisms of MMP-1 are not associated with the risk of HSIL and SCC, but with the invasiveness and prognosis of SCC. The heterozygous genotype of MMP-1 can be used as a prognostic marker in patients with invasive cervical cancer. PMID- 15661215 TI - Electrosurgical bipolar vessel sealing for radical abdominal hysterectomy. AB - OBJECTIVE: We analyzed the use of an electrosurgical bipolar vessel sealing system for radical abdominal hysterectomy. METHODS: We compared operating time, transfusion requirements and other surgical parameters in 52 patients undergoing radical abdominal hysterectomy with systematic pelvic +/- paraaortic lymphadenectomy for stage IB1-IIB cervical cancer between January 2001 and June 2004. At 21 operations between January 2001 and March 2002, the parametrium, paracolpos and vaginal cuff were resected with clamps and the pedicles suture ligated; 31 operations between November 2001 and June 2004 were done with a bipolar vessel sealing system (LigaSure Vessel Sealing System, Valleylab, Boulder, CO). Four surgeons did all operations. Data were compared with the t test. RESULTS: Patients operated with the LigaSure system received fewer packed RBC transfusions than those operated with clamps (mean, 0.61 +/- 1.1 vs. 2.14 +/- 2.6 units, respectively; P = 0.01), even when looking at patients who underwent only pelvic (without paraaortic) lymphadenectomy (mean, 0.52 +/- 1.1 vs. 1.29 +/- 1.2 units, respectively; P = 0.02). The transfusion rate in the two groups was 26% (8/31) and 67% (14/21), respectively (P < 0.05). Operating time did not differ significantly (199 +/- 33 vs. 213 +/- 45 min, respectively). There were no significant differences between the groups in number of pelvic nodes removed, febrile morbidity, postoperative stay, or days until residual urine <100 ml. One patient in the LigaSure group developed a ureterovaginal fistula that resolved with stenting. CONCLUSION: The LigaSure system appears useful to reduce blood loss at radical abdominal hysterectomy. PMID- 15661216 TI - Evaluation of weekly low-dose paclitaxel and carboplatin treatment for patients with platinum-sensitive relapsed ovarian cancer. AB - OBJECTIVE: Although paclitaxel and carboplatin therapy (TC) is an established effective standard regimen for patients with ovarian cancer, both treatment delay for hematologic toxicity and discontinuation of treatment due to neurotoxicity have occasionally been reported. To achieve therapeutic density, we evaluated the usefulness of weekly low-dose TC therapy (WTC) in patients with platinum sensitive (median PFI was 11.4 +/- 2.7 months) recurrent epithelial ovarian cancer. METHODS: A total of 25 patients were treated with paclitaxel at 60 mg/m(2) and carboplatin at AUC = 2 using 3 weekly courses with a 1-week break schedule. Eighteen patients had assessable tumors for response, and the other seven patients were evaluated by CA-125-based response. All of the patients were assessable for toxicity. RESULTS: The overall response rate (OR) based on WHO criteria was 84.2% (95% CI; 0.65-0.98), including nine complete responses (CR); OR based on CA-125 was 85.7% (95% CI; 0.42-0.99), including 3 CR. The total response rate was 88.0% (95% C.I.; 0.68-0.97). The median progression-free survival of the patients was 13.5 months during the mean follow-up period of 21.9 +/- 9.2 months. No patients had grade 1 or higher thrombocytopenia, and although 44% of the patients developed neurotoxicity, all cases remained grade 1. Treatment delay of over 7 days due to toxicity was observed in only two patients (16.0%) and in six cycles (1.3%) in a total of 451 cycles. CONCLUSION: WTC combination, as used in this study, produced a high response rate with acceptable toxicity, and the optimal combination in a weekly regimen remains to be determined. PMID- 15661217 TI - Percutaneous endoscopic gastrostomy tube placement in patients with malignant bowel obstruction due to ovarian carcinoma. AB - OBJECTIVES: To analyze the feasibility of using percutaneous endoscopic gastrostomy (PEG) tube placement in ovarian cancer patients with malignant bowel obstruction and to analyze the outcome of these patients. METHODS: We performed a retrospective review of all patients with ovarian carcinoma who underwent PEG tube placement between 1995 and 2002 at our institution. Abstracted data included patient demographics, procedure information, symptom resolution, diet tolerated, complications, further treatment, and survival. RESULTS: Ninety-four patients with ovarian carcinoma requiring PEG tube placement for malignant bowel obstruction were identified. The mean age at the time of PEG tube placement was 56 years. The mean interval from the initial cancer diagnosis to the placement of the PEG tube was 3.1 years. Twenty-two of 77 patients who had a computed tomography (CT) scan prior to PEG tube placement had tumor encasing the stomach. Fifty-nine (63%) of 94 patients had ascites, 25 of whom underwent a pre-PEG paracentesis (mean, 2845 ml). Ninety-four patients had a successful PEG tube placement under conscious sedation by the gastroenterology service (92) and/or by interventional radiology (2). Symptomatic relief, defined as no nausea or vomiting within 7 days of PEG tube placement, was noted in 86 (91%) of 94 patients undergoing PEG tube placement. Diets tolerated with and without the PEG tube being clamped were as follows: none, 3; sips, 9; liquids, 40; soft/regular, 40; and unknown, 2. The mean hospital stay after the procedure was 6 days. Eighteen patients had one or more of the following complications: leakage, 8; peristomal infection, 3; obstruction, 3; PEG tube migration, 2; catheter malfunction, 2; hemorrhage, 2; and peritonitis, 1. Nine patients required PEG tube revision due to complications. Chemotherapy after PEG tube placement was administered in 29 (31%) of the 94 patients, with resolution of obstruction and removal of the PEG tube in 4. In addition, 14 (15%) received limited total parental nutrition (TPN) after PEG tube insertion. Seventy-five (85%) of 88 patients died at home or under hospice care. The median overall survival for the 94 patients undergoing PEG tube placement was 8 weeks (95% CI, 6-10). Multivariate survival analysis revealed the presence of liver metastases (P < 0.001) and older age (P = 0.01) to be statistically significant predictors of shorter survival. The use of TPN after PEG tube placement was not a statistically significant prognostic factor in this model (P = 0.09). CONCLUSIONS: PEG tube placement in ovarian carcinoma is technically feasible and safe in the palliative setting. In addition, PEG tube placement allowed the majority of patients to have end-of-life care either at home or in an inpatient hospice. For the total population, no benefit was seen regarding survival with the use of TPN in this setting. Selected patients (younger age and without liver metastasis) may benefit from chemotherapy after PEG tube placement. PMID- 15661218 TI - Postoperative pelvic radiotherapy for cervical cancer patients with positive parametrial invasion. AB - OBJECTIVE: To evaluate patterns of failure in cervical cancer patients with histopathologic parametrial invasion treated with postoperative pelvic radiation therapy. METHODS: Records of 117 stages IB-IIB cervical cancer patients with parametrial invasion treated with postoperative radiation therapy from 1985 to 2002 were retrospectively reviewed. Patients were divided into two groups based on status of pelvic lymph nodes. Patterns of recurrence and prognosis by status of pelvic lymph nodes were statistically analyzed. RESULTS: Status of pelvic lymph nodes had significant impact on both recurrence and survival. Extrapelvic recurrence was observed in 23 of 66 node-positive patients compared with 6 of 51 node-negative patients (P = 0.005). Of 66 patients with a positive pelvic lymph node, 18 developed visceral metastases, whereas only three visceral metastases were noted in the 51 node-negative patients (P = 0.003). Five-year overall survival in node-positive and -negative patients was 52% and 89%, respectively (P = 0.0005). Corresponding rates for recurrence-free survival were 44% and 83%, respectively (P = 0.0002). The correlation between nodal metastasis and prognosis was enhanced when node-positive patients were stratified into two groups based on number of positive nodes (n = 1 and n > or = 2). Five-year recurrence-free survival rates for patients with negative, one positive, and two or more positive nodes were 83%, 61%, and 31%, respectively (P = 0.0001). CONCLUSIONS: Extrapelvic recurrence was uncommon in node-negative patients with parametrial invasion. These findings do not support use of systemic therapy for cervical cancer patients with parametrial invasion if pelvic lymph node metastasis is negative. PMID- 15661219 TI - Adenoviruses with an RGD-4C modification of the fiber knob elicit a neutralizing antibody response but continue to allow enhanced gene delivery. AB - OBJECTIVE: The purpose of this study was to investigate the effect of preexisting neutralizing antibody (NAbs) in naive mice and the effect of induced NAbs in mice immunized with either an RGD or nonmodified Ad5 vector on the transduction efficiency of adenoviral vectors. METHODS: BALB/c mice were immunized with Ad5LucRGD, with the unmodified Ad5Luc1, or with Opti-MEM intraperitoneally (ip) from one to three times. Sera were collected on day 27 and serially diluted to block Ad5Luc1 or Ad5LucRGD prior to infection of SKOV3.ip1 human ovarian carcinoma cells with these same vectors. Forty-eight hours post Ad infection, a luciferase assay was performed to determine the titer of NAbs. RESULTS: Luciferase assay data showed that the gene transfer efficacy of Ad5LucRGD was 1.56-fold higher than Ad5Luc1 in the presence of serum from naive mice. In the presence of serum from Ad5Luc1-challenged mice, the transduction efficiency of Ad5LucRGD was 3.27-fold higher (single challenge) and 4.2-fold higher (triple challenge) than Ad5Luc1. In the presence of serum from Ad5LucRGD-challenged mice, the transduction efficiency of Ad5LucRGD was 2.24-fold higher (single challenge) and 2.53-fold higher (triple challenge) than Ad5Luc1. CONCLUSION: The RGD modified human Ad vectors appear to be less recognizable than unmodified Ad to preexisting NAbs in mouse models. RGD-modified Ad vectors also appear to elicit a relatively lower level of NAbs that may also contribute to the higher gene transduction efficiency of these modified vectors. Therefore, RGD-modified Ad vectors may be reagents of clinical utility in the context of preformed anti-Ad immunity and in the setting of repetitive dosing. PMID- 15661220 TI - HLA-DQB1 and cervical cancer in Venezuelan women. AB - BACKGROUND: Cervical cancer represents a major health problem in Venezuela as well as in other Latin American countries. High-risk human papillomavirus (HR HPV) infection is known as the major risk factor of cervical cancer. However, whether or not a HR-HPV-infected woman progresses to cervical cancer may depend on the immune system effectors induced by viral antigens presented by her specific human leukocyte antigens (HLA) alleles. The role of the HLA system in presenting peptides to antigen-specific T-cells may be critical for genetic susceptibility and genetic resistance to cervical carcinoma. OBJECTIVE: We aimed to investigate the relationship between HLA-DQB1, HPV infection, and cervical cancer in Venezuelan women. METHODS: Blood samples and cervical swabs were obtained from 36 patients and 79 healthy controls; additional cervical biopsies were obtained from all the patients. HPV DNA was detected by PCR and HLA-DQB1 genotyping was performed using a PCR-SSP protocol. RESULTS.: A positive association with cervical cancer was observed for HLA-DQB1*0201-0202 and *0402 alleles, however after Bonferroni correction only HLA-DQB1*0402 remained statistically significant (P value = 0.004, RR = 5.067). CONCLUSION: This is the first report of HLA-DQB1 alleles associated with cervical carcinoma in Venezuelan women. Larger studies are needed to assess whether these HLA-DQB1*0201-0202 and *0402 alleles have a direct effect on disease susceptibility. PMID- 15661221 TI - High insulin-like growth factor-2 (IGF-2) gene expression is an independent predictor of poor survival for patients with advanced stage serous epithelial ovarian cancer. AB - OBJECTIVE: Epithelial ovarian cancer is the deadliest gynecologic malignancy, yet its molecular etiology remains poorly understood. Evidence is accumulating to support a role for the insulin-like growth factor family in human carcinogenesis, and recently using microarray expression analysis, we demonstrated over expression of the insulin-like growth factor-2 (IGF-2) gene in advanced stage epithelial ovarian cancers. The purpose of the current study is to further elucidate the role of the IGF-2 gene in ovarian cancer development and progression. METHODS: Relative expression of IGF-2 was measured in 109 epithelial ovarian cancers and eight normal ovarian surface epithelial (NOSE) samples, using quantitative real-time polymerase chain reaction. Associations with clinicopathological parameters were examined. RESULTS: Expression of the IGF-2 gene was more than 300-fold higher in ovarian cancers compared with normal ovarian surface epithelium samples (P <0.001). High IGF-2 expression was associated with advanced stage disease at diagnosis (P <0.001), high-grade cancers (P <0.05) and sub-optimal surgical cytoreduction (P = 0.08). In multivariate analysis, relative IGF-2 expression was an independent predictor of poor survival. CONCLUSIONS: Expression of the IGF-2 gene is significantly higher in ovarian cancers relative to normal ovarian surface epithelium. Further, high IGF-2 gene expression is associated with high grade, advanced stage disease, and is an independent predictor of poor survival in patients with epithelial ovarian cancer. As such, IGF-2 is a molecular marker and potential therapeutic target for the most aggressive epithelial ovarian cancers. PMID- 15661222 TI - Vaginal hysterectomy and abdominal hysterectomy for treatment of endometrial cancer in the elderly. AB - OBJECTIVE: The purpose of this study was to analyze the outcome of vaginal and abdominal hysterectomy for treatment of endometrial cancer in elderly patients. METHODS: In a retrospective series of 171 patients with age > or =70 years and at stages I-III, we evaluated operative and hospitalization data, as well as morbidity, mortality, and long-term survival associated with vaginal and abdominal hysterectomy. A total of 128 patients were operated on with vaginal hysterectomy and 43 cases underwent abdominal hysterectomy. RESULTS: Medically compromised patients were significantly more frequent in the vaginal surgery group (P = 0.01). Overall, the 10-year disease-specific survival rates after vaginal and abdominal hysterectomy were 80% and 78%, respectively (P = n.s.). Limiting the analysis to stage I (130 patients), 10-year disease-specific survival was 83% in 95 women operated on by the vaginal route and 84% in 35 patients operated by the abdominal approach (P = n.s.). Patients in the vaginal surgery group had a significantly shorter operative time (P = 0.01), less blood loss (P < 0.05), and were discharged earlier (P < 0.05). Severe complications occurred in 5.4% of the vaginal and in 7.0% of the abdominal procedures. Perioperative mortality was zero after vaginal hysterectomy and 2.3% after abdominal hysterectomy, respectively. CONCLUSIONS: Vaginal hysterectomy showed a high cure rate, shorter operative time, less blood loss, reduced morbidity, and no mortality and therefore may be considered the elective approach for treatment of elderly patients with endometrial cancer. PMID- 15661223 TI - Overexpression of the TGF-beta antagonist Smad7 in endometrial cancer. AB - OBJECTIVE: We have shown that HER2/Neu may activate the Smad7 promoter in endometrial, ovarian, and breast cancer cell lines. Elevated Smad7 levels could then antagonize the TGF-beta pathway, leading to a reduction in tumor surveillance and potential cancer formation. Our aim was to determine if Smad7 was in fact overexpressed in endometrial cancers and whether Smad7 RNA levels correlated with tumor grade or clinical endpoints. METHODS: Snap-frozen endometrial cancer specimens from 16 patients with grade 1 disease and 23 patients with grade 3 disease were obtained. Additionally, the endometrium from 18 patients who underwent hysterectomy for benign indications was collected as a control. RNA was extracted and subjected to quantitative real-time PCR to determine the degree of Smad7 RNA expression. Clinical outcomes including time to recurrence were recorded through retrospective chart review. RESULTS: Smad7 transcripts in the tumors were over 11-fold elevated on average than in controls (P < 0.001). There was no significant difference in Smad7 RNA between grades 1 and 3 tumors. For the 19 patients who recurred, median time to recurrence was 56.3 months for those with low Smad7 expression versus 30 months for those with high Smad7 expression (P < 0.004). CONCLUSION: Smad7 appears to be upregulated in endometrial cancers compared to normal endometrium. Furthermore, high Smad7 gene expression was associated with a shorter time to recurrence. Given that many endometrial cancers have been shown to be TGF-beta-unresponsive, Smad7 should be investigated as a potential target to restore TGF-beta responsiveness and limit tumor growth. PMID- 15661224 TI - A phase II study of gemcitabine plus carboplatin in platinum-sensitive, recurrent ovarian carcinoma. AB - OBJECTIVES: Gemcitabine and carboplatin each have demonstrated effectiveness without increased neurotoxicity in pretreated patients with ovarian cancer. We evaluated the efficacy and safety of gemcitabine plus carboplatin in patients with recurrent ovarian cancer in a multicenter phase II study. METHODS: Women with histologically proven measurable or evaluable epithelial ovarian cancer (any FIGO) who relapsed > or =6 months after discontinuation of first-line, platinum containing therapy received gemcitabine 1000 mg/m(2) on days 1 and 8 and carboplatin AUC 4 on day 1 (after gemcitabine) every 21 days for up to six cycles. RESULTS: Of the 40 enrolled/evaluable patients, 6 (15%) had complete response and 19 (47.5%) had partial response (PR), including one patient with PR in nonmeasurable disease (PRNM), for an overall response rate of 62.5% (95% CI, 45.8-77.3%). The median duration of response was 7.8 months (95% CI, 6.7-10.0), the median time to progressive disease was 9.6 months (95% CI, 8.5-11.0), and the median time to treatment failure was 9.3 months (95% CI, 8.2-10.4). The main grade 3/4 toxicities were neutropenia (78% of patients), leukopenia (30%), thrombocytopenia (18%), and anemia (15%); no grade 4 nonhematologic toxicities occurred, and grade 3 nonhematologic toxicities were mild. CONCLUSIONS: The combination of gemcitabine and carboplatin is active and feasible in platinum sensitive patients with recurrent ovarian cancer. This regimen is undergoing further evaluation in a large phase III trial. PMID- 15661225 TI - 17-Allyamino-17-demethoxygeldanamycin and 17-NN-dimethyl ethylene diamine geldanamycin have cytotoxic activity against multiple gynecologic cancer cell types. AB - OBJECTIVE: HSP90 is a cellular chaperone that is overexpressed in many cancers. HSP90 assists in proper folding of a variety of clients, many of which are oncoproteins. HSP90 has been shown to be elevated in endometrial, ovarian, and breast cancer. Furthermore, HSP90 is known to stabilize the oncoprotein Akt; disruptions of the Akt pathway are common in gynecologic malignancies. We sought to evaluate the effectiveness of HSP90 inhibitors in gynecologic cancer. METHODS: We tested two HSP90 inhibitors, 17-AAG and 17-DMAG, against gynecologic cancer cell lines (four endometrial, one cervical, one ovarian, and one breast cancer line). We performed Western blots to determine effects of treatment on levels of HSP90 client proteins and PARP cleavage. 3-(4,5-Dimethylthiazol-2-yl)-2,5 diphenyltetrazolium bromide (MTT) assays were used to assess cell viability, and flow cytometry to quantitate cell-cycle distribution and apoptosis. RESULTS: After treatment with 17-AAG or 17-DMAG, we detected no decrease in HSP90 levels. Levels of other oncoproteins did decrease with treatment: phosphorylated and total Akt, and Met. One cell line underwent G(1) arrest, and five showed G(2) arrest. All showed some level of apoptotic cell death, which was confirmed by detection of PARP cleavage. Sensitivity to the drugs varied among cell lines, ranging from 20% to 90% apoptosis after treatment. Our data suggest that 17-DMAG may be more potent than 17-AAG. CONCLUSIONS: HSP90 inhibitors are effective cytotoxic agents in gynecologic cancer cells. Further testing in in vivo model systems is warranted, with the goal of eventual translation to clinical trials in gynecologic oncology patients. PMID- 15661226 TI - Salvage whole-abdominal radiation therapy after second-look laparotomy or secondary debulking surgery in patients with ovarian cancer. AB - OBJECTIVES: Our aim was to determine the outcomes associated with use of whole abdominal radiation therapy (WART) in women with ovarian cancer, to identify predictors of response, and to assess associated toxicity. METHODS: From 1981 through 2000, 171 women received WART at our institution after ovarian cancer surgery. Relevant clinical information was extracted through retrospective chart review. RESULTS: One hundred nine patients received WART after positive second look laparotomy (SLL), and 62 were treated after secondary debulking (SD) for recurrent disease. The median dose to the whole abdomen was 25.5 Gy (range, 1.0 30.5 Gy). Therapy included a pelvic boost in 120 patients (70%) and a para-aortic boost in 21 patients (12%). The planned radiation course was completed in 123 patients (72%). In the SLL group, 5-year survival was 29% with a median follow-up of 98.4 months. The 5-year progression-free survival (PFS) was 41% in those with microscopic disease. There was one treatment-related death (1%). For the SD group, median PFS was 11 months and associated with treatment-related mortality in 5%. Overall, treatment-related small bowel obstruction occurred in 26 patients (15%). CONCLUSIONS: In patients with a positive SLL, WART should be considered only for those with microscopic residual disease. Treatment-related small bowel obstruction can be expected in 15% of these patients. Use of WART for recurrent disease appears to be related to serious bowel toxicity in 5% with an associated short disease-free interval; the therapeutic index of WART may not be acceptable in patients with recurrent disease regardless of the degree of cytoreduction. PMID- 15661227 TI - Benign ovarian serous tumors: a re-evaluation and proposed reclassification of serous "cystadenomas" and "cystadenofibromas". AB - OBJECTIVE: Serous cystadenomas and cystadenofibromas of the ovaries are currently regarded as neoplasms and are considered the most common ovarian neoplasms. The purpose of this study is to determine what proportion of benign serous tumors contain an epithelial proliferation (the hallmark of a neoplastic process in nearly all other sites) that can be considered neoplastic as opposed to reactive in nature. METHODS: An unselected series of 113 ovarian serous tumors (76 serous cystadenomas and 37 serous cystadenofibromas) were histologically evaluated. A 1 mm in diameter area of epithelial proliferation was considered potentially neoplastic. RESULTS: Eight tumors (7%) displayed at least 1 mm of epithelial proliferation (1% of serous cystadenomas and 19% of serous cystadenofibromas). CONCLUSION: The vast majority of benign serous tumors may not be bona fide epithelial neoplasms, but rather, may represent cystically dilated glandular inclusions (cystadenomas) and fibromas with epithelial inclusions (cystadenofibromas). A recently published study evaluating clonality in serous cystadenomas found that the vast majority are polyclonal and thus supports this hypothesis. These findings have important implications for the pathogenesis of ovarian cancer, for the distribution of ovarian neoplasms, and for the interpretation of molecular biological studies of ovarian tumors. PMID- 15661228 TI - Lymph node metastases in low-grade endometrial stromal sarcoma. AB - OBJECTIVES: The objective of this study was to review our experience on lymphatic dissemination in patients with low-grade endometrial stromal sarcoma. METHODS: All cases diagnosed as low-grade endometrial stromal sarcoma or endolymphatic stromal myosis before October 2003 and who had lymph node sampling at some point in their evolution were retrieved from the files of the pathology and gynecologic oncology departments of l'Hotel-Dieu de Quebec University Hospital (HDQ). RESULTS: Fifteen patients with either limited lymph node biopsies or a complete lymph node dissection at some point in the course of their disease were found. Five of these patients (33%) presented lymph node metastases either at the initial hysterectomy, during a subsequent staging procedure, or at the time of a recurrence. CONCLUSION: These findings suggest that the incidence of lymph node involvement in low-grade endometrial stromal sarcoma is higher than expected. More extensive sampling of lymph nodes in a larger number of patients may allow a better understanding of the frequency and prognostic significance of these metastases. PMID- 15661229 TI - Post-hysterectomy radiotherapy in FIGO stage IB-IIB uterine cervical carcinoma. AB - OBJECTIVE: This study is a retrospective analysis of stage IB-IIB cervical carcinoma patients who had received postoperative radiotherapy (PORT). METHODS: Eight hundred patients with stage IB-IIB cervical carcinomas who received PORT after radical hysterectomy and bilateral pelvic lymph node dissection (PLND) between February 1979 and March 2000 were analyzed. RESULTS: The median follow-up duration was 100 months. The 5-year overall survival (OS) and disease-free survival (DFS) rates were 88% and 81%, respectively. One hundred forty-six patients (18%) failed, and 103 of these had distant metastases. Multivariate analysis revealed that pelvic lymph node (LN) metastasis significantly compromised OS, DFS, pelvic failure-free survival (PFFS), and distant failure free survival (DFFS) (P < 0.05). Patients with age <50 years, deep stromal invasion (DSI), and lymphovascular space invasion (LVSI) were significantly associated with a higher risk of distant metastasis after PORT. The incidences of late rectal, urinary, and small bowel complications of grade 3 or higher were 1.6%, 1.4%, and 1.0%, respectively. CONCLUSIONS: PORT achieved good OS and DFS in the patients with risk factors after radical hysterectomy for stage IB-IIB cervical carcinomas. Distant metastasis was the major pattern of treatment failure after PORT. Effective systemic chemotherapy might be a breakthrough in improving the outcome of PORT in patients with cervical carcinomas. PMID- 15661230 TI - Soyasaponin-I-modified invasive behavior of cancer by changing cell surface sialic acids. AB - OBJECTIVE: Sialylation involving tumor formation and invasive behavior goes along with altered sialyltransferase (ST) activity. A potent ST inhibitor, soyasaponin I (SsaI), was discovered to selectively inhibit the cellular alpha2,3 sialyltranserase activity. In this study, we further test the effects of SsaI on modifying the metastatic and invasive behaviors of cancer cell lines. METHODS: Nonmetastatic breast cancer cell line, MCF-7, and highly metastastic breast cancer cell line, MDA-MB-231, were used to investigate the effects of SsaI on tumor cells. RESULTS: SsaI did not affect cell growth cycle and also failed to inhibit cell growth in this study (the concentration of SsaI < or=100 muM). SsaI was as predicted to successfully inhibit cellular alpha2,3-ST activity and depressed the dose-dependent tumor cell surface alpha2,3-sialic acid expression. In addition, different concentrations of SsaI did stimulate MCF-7 cell adhesion to collagen type I linearly and significantly enhanced cell adhesion to the Matrigel-matrix. Furthermore, SsaI significantly decreased MDA-MB-231 cell migration. Reverse transcriptase polymerase chain reaction for evaluating mRNA expression of ST3Gal I, III and IV showed that SsaI also down-regulated the expression of ST3Gal IV but did not affect the other two. CONCLUSIONS: The results showed that SsaI was implicated in the invasive behavior of tumor cells, suggesting that altered alpha2,3-sialylation pathway played a crucial role in the adhesion and tumor metastases. SsaI is a good candidate for studying the biological roles of ST, and might provide a new preventive strategy in tumor metastasis. PMID- 15661231 TI - Association of the NAD(P)H: quinone oxidoreductase C609T polymorphism and the risk of cervical cancer in Japanese subjects. AB - OBJECTIVE: In this study, genetic polymorphisms, NQO1 C609T, GSTM1 positive/null, and GSTT1 positive/null, were examined with reference to cervical cancer risk in a population-based incident case-control study in Japanese. METHODS: The cases comprised 131 cervical cancer patients: 87 cases with squamous cell carcinoma (SCC) and 44 with adenocarcinoma (ADC) or adenosquamous carcinoma (ADSC). Controls were sampled from 320 healthy women who underwent a health checkup. RESULTS: The cervical cancer risk was substantially elevated with smoking for all cases, SCC cases, and ADC/ADSC cases (OR = 4.50, 95% CI = 2.48-8.17, P < 0.001; OR = 5.68, 95% CI = 2.99-10.78, P < 0.001; and OR = 2.57, 95% CI = 1.09-6.08, P = 0.032; respectively). The frequency of the NQO1 609TT genotype, reported to be associated with null enzyme activity, was higher in individuals with all cases and SCC than in the healthy controls (OR = 1.97, 95% CI = 1.06-3.66, P = 0.032; and OR = 2.42, 95% CI = 1.21-4.82, P = 0.012; respectively), but not in ADC/ADSC cases. Analysis of polymorphisms for GSTM1 and GSTT1 showed no significant differences between cervical cancer patients and controls. In stratification analysis, significant elevated risk of all cases and SCC was associated with the NQO1 609TT genotype among nonsmokers (OR = 2.15, 95% CI = 1.08-4.30, P = 0.030; and OR = 2.83, 95% CI = 1.21-6.31, P = 0.011; respectively), but not smokers. No gene-gene interaction was observed in our case subjects. CONCLUSION: This is the first report that the NQO1 gene might be important in relation to the risk of squamous cell carcinoma of the cervix. PMID- 15661233 TI - Additional salpingectomy after previous prophylactic oophorectomy in high-risk women: sense or nonsense? AB - OBJECTIVES: Since BRCA1/2 germ line mutation carriers are also at a higher risk of developing fallopian tube carcinoma, resection of the fallopian tubes is currently included at the time of risk reducing surgery. In this study, we comment on the need of additional bilateral prophylactic salpingectomy (BPS) following previous bilateral prophylactic oophorectomy (BPO) in women at high risk of ovarian cancer. METHODS: Retrospectively, the medical files of 42 high risk women, who had undergone BPO only, were reviewed. RESULTS: In our center, risk-reducing surgery consisted of BPO only for 42 women. Twenty-seven women received an informative letter in which counseling for additional BPS was offered. In total, 15 women opted for additional BPS. Surgery was performed with a mean interval of 65 months (range 6-101) in 10 BRCA1 carriers, one BRCA2 carrier, one BRCA1 and 2 carrier, and three women with non-informative test results. The procedure was readily done by laparoscopy in 13 women and two needed a laparotomy. No post-operative complications had occurred. Histopathological examination revealed no malignancy. CONCLUSIONS: We believe that additional risk reduction of cancer necessitates BPS in BRCA1/2 carriers after previous BPO. BPS after previous BPO was easily performed. Today, physicians should include resection of the fallopian tube at prophylactic surgery in high-risk women and should consider additional BPS in women who have undergone BPO only. PMID- 15661232 TI - Malignant ascites increases the antioxidant ability of human ovarian (SKOV-3) and gastric adenocarcinoma (KATO-III) cells. AB - OBJECTIVES: The antioxidant status of cancer cells is an important factor in tumor invasion and metastases. This study investigated whether metastatic cancer cells derive beneficial antioxidant protection from ascitic fluid and are rendered resistant to oxidative stress in the form of a chemically generated free radical insult. METHODS: Human gastric carcinoma (KATO-III) and ovarian adenocarcinoma (SKOV-3) cell lines were cultured and incubated for 24 h with (1) M199 medium; (2) M199 + 20% fetal calf serum (FCS); (3) malignant ascites. All cells were exposed to a hydroxyl radical-generating system for 1 h. Cellular lipid peroxidation was assessed by measuring malondialdehyde (MDA) in cell suspensions. Glutathione (GSH) levels in cell pellet were measured in SKOV-3 cells after 0, 24, 48, and 72 h of incubation with buthionine sulphoximine (BSO). CD44 gene expression of cancer cells was analyzed by Northern blotting. RESULTS: The results showed that the cancer cells were rendered resistant to oxidative stress and with upregulated CD44 gene expression by components of malignant ascites. CONCLUSIONS: These findings suggest that malignant ascites increases the antioxidant ability of cancer cells and the potential of adhesion and invasion. Thus, determination of the nature of these putative tumor-protective components of ascites may provide targets for therapeutic intervention. PMID- 15661234 TI - A phase II study of paclitaxel, carboplatin, and gemcitabine in previously untreated patients with epithelial ovarian cancer FIGO stage IC-IV (AGO-OVAR protocol OVAR-8). AB - PURPOSE: A multicenter, nonrandomized, phase II study was initiated to evaluate the tolerability, toxicity, and activity of paclitaxel, carboplatin, and gemcitabine combination in previously untreated ovarian cancer. PATIENTS AND METHODS: Chemonaive patients who had radical debulking surgery for primary epithelial ovarian cancer International Federation of Gynecology and Obstetrics (FIGO) IC-IV received sequentially paclitaxel 175 mg/m(2), carboplatin AUC 5, and gemcitabine 800 mg/m(2) on day 1 and gemcitabine 800 mg/m(2) on day 8, every 3 weeks. RESULTS: From October 2001 to July 2002, 55 patients were treated and evaluated. Main toxicities were hematological with NCI-CTC grade 3/4 anemia 12.7%, leukopenia 70.9%, neutropenia 76.3%, and thrombocytopenia 45.5. However, febrile neutropenia occurred only in 1.8%. Grade 3/4 nonhematological toxicities were rare and occurred in less than 10% of patients. Toxicity-induced treatment delays occurred in 3.1% of cycles and resulted in early treatment cessation in four patients. Dose intensity reached 90.8% for carboplatin and paclitaxel, and 73.3% for gemcitabine. Objective response was observed in 10 of 14 patients with measurable disease. CONCLUSIONS: The triplet combination of paclitaxel carboplatin-gemcitabine is feasible and active, with manageable hematological toxicity and no unexpected nonhematological toxicity. This regimen has proceeded to phase III evaluation. PMID- 15661235 TI - Immune responses to human papillomavirus in genital tract of women with cervical cancer. AB - OBJECTIVES: To address a question whether immune responses to HPV infection play a role in control of cervical cancer, we analyzed systemic and mucosal immune responses to HPV in women who underwent radical hysterectomy for cervical cancer (HCC) or loop conization due to cervical dysplasia (LOOP), or had hysterectomy for other reasons (HNN). METHODS: HPV-specific antibodies in sera and vaginal washes were determined by ELISA using recombinant HPV 16 E7 oncoprotein. Cytokines in vaginal washes were assayed by Linco cytokine multiplex method using Luminex technology. Differential gene expression profiling in cervical tumor was determined by microarray analysis and Real-time RT-PCR. RESULTS: While levels of HPV-16 E7-specific IgG in vaginal wash were significantly higher in women undergoing HCC and HNN, the levels of the HPV-16 E7-specific IgA in vaginal wash of women with cervical cancer and cervical dysplasia were lower as compared to patients in HNN. Proinflammatory cytokines, such as IL-6 and IL-8, were dominant in vaginal washes of all subjects studied. However, no pattern of Th1-type and Th2-type cytokine induction was observed as demonstrated by protein analysis as well as differential gene expression profiling in cervical tumor. CONCLUSIONS: These results suggest a selective down-regulation of local HPV-specific IgA responses in women with cervical cancer. PMID- 15661236 TI - Translocation of Fas by LPA prevents ovarian cancer cells from anti-Fas-induced apoptosis. AB - OBJECTIVES: Alterations in the expression of Fas have been demonstrated in various cancers as a mechanism for tumor cells to escape from immune surveillance. In this study, we observed the effect of lysophosphatidic acid (LPA) on Fas expression and function in ovarian cancer cells. METHODS: Ovarian cancer cell lines were incubated with or without LPA and Fas cell surface presentations were detected by flow cytometry. Anti-Fas IgM was added for induction and analysis of apoptosis by flow cytometry. Cell lysis and subcellular fractions were probed for protein expression by Western blot. Cells were also stained with human anti-Fas Ab, followed with Rhodamine red-X-conjugated goat anti-mouse IgG, and immunofluorescence images were acquired on a Nikon digital camera. RESULTS: Following treatment with LPA, ovarian cancer cells showed significant rapid reduction of Fas presentation on the cell surface. LPA protected ovarian cancer cells from anti-Fas-induced apoptosis. Cell lysis and subcellular fractionations proved that LPA treatment induced a translocation of Fas receptors, along with phosphorylated ezrin, from the membrane anchored to the actin cytoskeleton, to the cytosol. Translocation of the Fas receptor reduced Fas concentration in the membrane and may inhibit its clustering and internalization during early apoptosis induced by anti-Fas. DISC staining proved that LPA inhibited Fas receptor aggregation and caspase-8 activation at the membrane, which further inhibited caspase-3 and 7 activation in the cytosol. CONCLUSIONS: Our studies suggest that LPA induces translocation of Fas from the cell membrane to the cytosol, which may provide a mechanism by which ovarian cancer cells evade FasL-bearing immune cells. PMID- 15661237 TI - Thromboembolic events in patients treated with definitive chemotherapy and radiation therapy for invasive cervical cancer. AB - OBJECTIVES: Determine the incidence of and risk factors for thromboembolic events (TE) in patients treated with definitive chemoradiation for cervical cancer at our institution. METHODS: A retrospective chart review was performed of all patients with a diagnosis of invasive carcinoma of the cervix (FIGO Stage IB-IVA) treated with definitive chemoradiation at University of Iowa Hospitals and Clinics (UIHC) from July 2002 to December 2003. Forty-eight patients met these criteria. All but one patient received 45 Gy to the pelvis followed by brachytherapy, IMRT, or conformal boost. One patient received 39.6 Gy to the pelvis. Cisplatin chemotherapy, 40 mg/m squared, was given weekly for 6 weeks. Data were collected for FIGO stage, age, body mass index (BMI), and smoking history. TE were confirmed by Doppler ultrasound or pulmonary imaging. Log-rank tests were used to examine the association between time to TE and the variables FIGO stage and smoking status. The association between time to TE and the continuous variables age and BMI was examined with Cox proportional hazards regression. All tests were two-sided and carried out to the 5% level of significance using the SAS statistical software package. RESULTS: Minimum follow up was 8 months. Eight patients (16.7%) developed a TE. The associations were not statistically significant for stage (P = 0.72), smoking status (P = 0.72), age (P = 0.63) or BMI (P = 0.86). Risk factors were similar in both groups. Data review suggests that the entire group had risk factors for TE. CONCLUSIONS: We noted a high incidence of TE (16.7%) in patients treated at UIHC with chemoradiation for invasive cervical cancer. We did not find a statistical association between age, stage, smoking history, or BMI and risk of TE in this group. Patients with and without TE had multiple risk factors for TE. PMID- 15661238 TI - Human papillomavirus type 16 E7 peptide(38-61) linked with an immunoglobulin G fragment provides protective immunity in mice. AB - OBJECTIVE: To explore whether the recombinant protein (Human papillomavirus (HPV) type16 E7 peptide(38-61) linked with an immunoglobulin G fragment) will generate protective immunity in mouse model. METHODS: In our study, we combined the HPV16 E7 peptide(38-61) with a murine IgG heavy chain constant region to construct a chimeric protein compound, which was highly expressed as inclusion bodies in a bacterial expression system with Escherichia coli. The purified chimeric protein was injected into C57BL/6 mice and the efficiency of the chimeric vaccine candidate was evaluated by antibody response assay, T cell proliferation assay, CTL assay, tumor challenge assay and therapeutic experiment. RESULTS: The chimeric vaccine candidate was able to induce anti-HPV antibodies as well as to elicit HPV16 E7-specific CTLs and T cell proliferation in a pre-clinical mouse model. It was also able to effectively protect mice against the challenge of HPV16-positive tumor cells, and to eradicate HPV16-expressing tumors in mice. CONCLUSIONS: The chimeric protein vaccine can induce E7-specific immune responses and protect mice against challenge of HPV16-positive tumor, even eradicate developed tumor. The results indicated a possibility to use the chimeric protein vaccine to protect human against HPV infection. PMID- 15661239 TI - Laparoscopic modified radical hysterectomy: a strategy for a clinical dilemma. AB - OBJECTIVE: To investigate the role of laparoscopic modified radical (type 2) hysterectomy when cervical cancer cannot be excluded or documented preoperatively. METHODS: Between 1996 and 2004, 50 patients with cervical intraepithelial neoplasia (CIN III) or adenocarcinoma in situ (AIS) involvement of cone endocervical margins and/or endocervical curettings, who were not candidates for observation or repeat conization, underwent laparoscopy to perform a modified radical hysterectomy. RESULTS: Forty-nine (98.0%) modified radical hysterectomies were completed laparoscopically and one (2.0%) patient required a laparotomy. Of the overall group, 35 (70.0%) had residual pathology; 26 (52.0%) were precancerous lesions, and 9 (18.0%) had invasive disease (5 adenocarcinomas, 3 squamous lesions, and 1 adenosquamous carcinoma). Of the nine with cancer, one had stage IA1 disease, three had stage IA2 disease, and five had stage IB1 disease. Five (55.6%) invasive lesions were diagnosed intraoperatively (frozen section), and a laparoscopic pelvic and lower aortic lymph node dissection was performed. The median operative time was 96 min (range 58-185), blood loss 100 ml (50-450), and postoperative hospital stay 2.5 days (range 1-14). There were no incidences of prolonged urinary retention fistulas, or other serious complications. All patients with cancer remain disease-free (median follow-up 44.2 months, range 1-88.7 months). CONCLUSIONS: Laparoscopic modified radical hysterectomy is a treatment option for patients for whom cervical cancer cannot be definitively excluded, and can be completed with acceptable operative time, blood loss, and hospitalization. PMID- 15661240 TI - Postoperative adjuvant concurrent chemoradiotherapy improves survival rates for high-risk, early stage cervical cancer patients. AB - OBJECTIVE: This study was undertaken to evaluate the efficacy of postoperative concurrent chemoradiotherapy (CCRT) and to investigate the recurrence and survival rates after adjuvant CCRT in high-risk early cervical cancer (stages IA2, IB, IIA) patients who were treated by radical hysterectomy and pelvic lymphadenectomy. METHODS: From July 1994 to June 2001, we retrospectively reviewed the medical records of 151 patients who had undergone radical abdominal hysterectomy with pelvic lymphadenectomy and paraaortic lymph nodes dissection at Ajou University Hospital for early cervical cancer (stages IA2, IB, IIA). CCRT was performed in 30 patients with high-risk factors such as positive pelvic lymph nodes, parametrial involvement, or positive surgical margins. Adjuvant chemotherapy consisted of cisplatin (70 mg/m(2) on day 1) and 5-fluorouracil (5 FU; 1000 mg/m(2) on days 2-5) for four cycles every 4 weeks beginning 2-3 weeks after surgery. Pelvic radiotherapy was started concurrently at the second and third cycle of chemotherapy. We also analyzed the recurrence pattern and survival rates of 114 patients (control group) who received no adjuvant therapy after surgery. The mean follow-up period was 49 months (24-98 months). RESULTS: There were recurrences in three patients after CCRT (10%) and in five patients in the control group (4.4%). The actuarial 5-year overall survival rates for patients in CCRT and control group were 96.7% vs. 97.7%, respectively. The progression-free survival rates were 88.7% for the high-risk group and 95.4% for the non-high-risk group. CONCLUSIONS: This study confirms good local control and 5-year overall and progression-free survival rates in high-risk cervical cancer patients after CCRT, which is comparable with the results of the control group. Our results indicate that adjuvant concurrent chemoradiotherapy seems to be effective in stages IA2 IIA cervical cancer patients with high-risk factors. PMID- 15661241 TI - Are germinal inclusion cysts markers of ovulation? AB - OBJECTIVES: Germinal inclusion cysts (GICs) of the ovary are possible precursor lesions for epithelial ovarian cancer. These cysts have been postulated to form as stigmata of ovulation. It is known that oral contraceptives, breastfeeding, and pregnancy, which decrease lifetime ovulations, confer protection against epithelial ovarian cancer. This study was undertaken to determine if ovarian density of epithelial inclusion cysts corresponds to a woman's frequency of ovulation. METHODS: One hundred two cases of women undergoing incidental oophorectomy at the time of surgery for benign gynecological conditions were interviewed for information relating to ovulation. Review of multiple sections of ovarian tissue was performed to count numbers of GICs. RESULTS: There was a modest correlation between the average number of germinal inclusion cysts and ovulatory age (Spearman's rho = 0.2, P = 0.06). There were fewer GICs in current smokers and ever-users of oral contraceptives; however these associations were not significant (P > 0.1). CONCLUSIONS: This study showed increased numbers of GICs in women of greater ovulatory age, and those who were not current smokers or ever used oral contraceptives. This provides some evidence that GICs may be caused by ovulation. The small number of subjects limited our ability to evaluate associations. PMID- 15661242 TI - The influence of time intervals between loop electrosurgical excision and subsequent hysterectomy on the morbidity of patients with cervical neoplasia. AB - OBJECTIVE: To improve clinical prospects by reducing intraoperative or postoperative complications, subsequent hysterectomy is generally conducted within 48 h or 6 weeks after cervical cold-knife conization. The loop electrosurgical excision procedure (LEEP) is widely used for cervical conization. However, no study has ever been undertaken on the relation between postoperative sequelae and the time between LEEP and hysterectomy. Therefore, this study was undertaken to evaluate the correlations between postoperative sequelae and the interval between LEEP and hysterectomy. METHODS: The medical records of 338 patients, who underwent type 1 extended hysterectomy after LEEP at the Department of Obstetrics and Gynecology, Yonsei University College of Medicine, were retrospectively reviewed. The subjects were divided into three groups according to time from LEEP to hysterectomy: group 1 (within 48 h, n = 210), group 2 (between 48 h and 6 weeks, n = 88), and group 3 (>6 weeks, n = 40). RESULTS: The three groups showed no significant differences with respect to patient characteristics (age, delivery history, body mass index, and a history of surgery). Postoperative complications such as fever, dysuria, and surgical region complications (effraction, infection, and rubefaction) were not significantly different among the three groups. Other complications, namely, ureter injury and abdominal wall hematoma, were found in one case in each group 1. CONCLUSION: The postoperative clinical courses were not significantly different regardless of time interval between LEEP and subsequent hysterectomy. Therefore, hysterectomies can be conducted at any time when the patient is in an appropriate condition, i.e., not precisely within 48 h or >6 weeks after LEEP. PMID- 15661243 TI - Role of sentinel lymph node biopsy procedure in cervical cancer: a critical point of view. AB - OBJECTIVE: To report our experience about the role of sentinel node biopsy in cervical cancer patients while debating provocatory arguments concerning this procedure. METHODS: From June 2001 to February 2003, patients affected by stage IB(1) cervical cancer were submitted to the sentinel node biopsy procedure. Patients were submitted to lymphoscintigraphy and, subsequently, to laparoscopy in order to locate the sentinel lymph node. RESULTS: Thirty-seven patients were enrolled in the study. Sentinel node(s) was(were) identified with preoperative lymphoscintigraphy in 89% of the patients. Intraoperative detection rate was 70%. During surgery in 31% of the patients, sentinel node was detected bilaterally; in 15%, two sentinel nodes on the same side of the lymphatic vessels were detected. The sentinel node was located at the level of superficial common iliac vessels (26%), external iliac vessels (69%), and superficial obturator vessels (49%). In 77% of the patients, the histologic specimen sent by the surgeon as unique sentinel node contained two or more nodes. Metastatic sentinel nodes were found in 23% of the patients. There was no case with a positive nonsentinel node in the presence of a negative sentinel node. CONCLUSION: Sentinel node detection is a feasible procedure in cervical cancer patients. However, a high percentage of patients is found with bilateral and/or more than one sentinel lymph node. Improvements in detection rate and pathological analysis are needed prior to consider the sentinel node biopsy a routine procedure in cervical cancer patients. PMID- 15661244 TI - Clinicopathologic significance of loss of heterozygosity on chromosome 1 in cervical cancer. AB - OBJECTIVES: We studied the loss of heterozygosity (LOH) in chromosome 1 in squamous cell carcinoma (SCC) of the uterine cervix and evaluated its clinical and pathological significance. METHODS: Sixty-three highly polymorphic markers were used to study the LOH in 84 SCC. Microdissection was performed to enrich the tumor cells population before the alleotyping study. The findings were correlated with clinicopathologic findings. RESULTS: LOH was detected in all but one SCC. The number of loci showing LOH in each case ranged from 0 to 41. Five loci showed LOH in > or =30% SCC and 28 other loci had an LOH frequency between 20% and 30%. Six of the eight markers located at 1p36.21 to 1p36.33 had a frequency of LOH >20%. Shortened total survival was associated with LOH at 14 loci and shortened disease-free survival was associated with LOH at 11 loci while LOH at nine loci were associated with both. A high frequency of LOH was associated with stage as well as shortened total and disease-free survival. CONCLUSIONS: LOH is a common and early event in the development of cervical SCC. Tumor suppressor genes may be present at 1p36. The incidence of LOH increases as the tumor progresses but a high frequency of LOH is not an independent prognostic factor. PMID- 15661245 TI - Use of serum secretory leukocyte protease inhibitor levels in patients to improve specificity of ovarian cancer diagnosis. AB - OBJECTIVE: To assess the clinical relevance of serum secretory leukocyte protease inhibitor (SLPI) levels in distinguishing patients with ovarian cancers from those with benign ovarian cysts, we determined concentrations with reference to the FIGO stage and other clinical characteristics. METHODS: Preoperative serum SLPI levels were measured in women with invasive epithelial ovarian cancer (n = 55), benign ovarian cysts (n = 25), or normal controls (n = 38) using an enzyme linked immunosorbent assay (ELISA). RESULTS: The serum SLPI concentration was significantly elevated in the ovarian cancer patients (median 67 ng/ml, interquartile range 26-124 ng/ml) as compared to the benign cyst patients (37 and 25-66 ng/ml) or healthy women (32 and 25-43 ng/ml). Using an SLPI cutoff of 50 ng/ml and a CA125 cutoff of 30 units/ml, with both markers elevated the sensitivity was 95%, the specificity was 100%, the positive predictive value was 100%, and the negative predictive value was 89% between the malignant and benign cyst patients. CONCLUSION: Serum SLPI levels could be useful for differentiating benign ovarian cysts from malignancies and to improve the specificity of diagnosis. PMID- 15661246 TI - Histologic types of epithelial ovarian cancer: have they different risk factors? AB - OBJECTIVES: The histologic types of epithelial ovarian cancer differ in clinical behavior, descriptive epidemiology, and genetic origins. The goals of the current study were to characterize further the relation of histologic-specific ovarian cancer risks to reproductive and lifestyle attributes. METHODS: The authors conducted a pooled analysis of 10 case-control studies of ovarian cancer in US White women, involving 1834 patients with invasive epithelial ovarian cancer (1067 serous, 254 mucinous, 373 endometrioid, and 140 clear cell) and 7484 control women. RESULTS: Risks of all four histological types were inversely associated with parity and oral contraceptive use, but the histologic types showed different associations with nonreproductive factors. Unique associations include an inverse relation of serous cancer risk to body mass index, a positive relation of mucinous cancer risk to cigarette smoking, and a weakly positive relation of endometrioid cancer risk to body mass index. Risk of all histologic types was unassociated with age at menarche, age at menopause, a history of infertility, noncontraceptive estrogen use, and alcohol consumption. CONCLUSIONS: The most important modifiers of ovarian cancer risk (parity and oral contraceptive use) showed similar associations across the histologies. Nevertheless, the unique associations seen for other modifiers support the conjecture that the histologic types of epithelial ovarian cancer have different etiologies, which should be addressed in future investigations of the molecular basis of ovarian cancers and their responses to therapies. PMID- 15661247 TI - Up-regulation of DNA methyltransferase 3B expression in endometrial cancers. AB - OBJECTIVE: To understand the role of epigenetic regulation in the pathogenesis of endometrial cancer, we have characterized DNA methyltransferase 3B (DNMT3B) gene expression in normal, Grade I and Grade III endometrioid cancers, and examined DNMT3B promoter activities in endometrial cancer cell lines. METHODS: DNMT3B expression was measured in normal, Grade I, and Grade III endometrioid cancer samples. Real-time PCR and Western blot analysis were performed to compare DNMT3B mRNA and protein levels. DNMT3B levels were also compared among endometrial cell lines including those for Ishikawa, KLE, AN3, RL-95, HEC-1A, and HEC-1B. DNMT3B promoter reporter plasmids were constructed. Promoter activities in well and poorly differentiated cell lines were compared by in vitro reporter gene transfection. RESULTS: DNMT3B was significantly up-regulated in both Grade I and Grade III cancers as compared to normal controls. Western blot analysis confirmed the increased DNMT3B protein expression in cancer tissues. It was also found that the well-differentiated endometrial cell line, Ishikawa, expressed lower levels of DNMT3B than the poorly differentiated KLE cells, the expression patterns similar to those observed in tumor specimens. CONCLUSION: The results suggest that DNMT3B overexpression may play a significant role in endometrial cancer development. In addition, the transfection experiments indicated that DNMT3B promoters are more active in the poorly differentiated endometrial cancer cell lines, suggesting that the in vitro assay provides a useful model for studying the DNMT3B transactivation mechanism related to tumor transformation. PMID- 15661248 TI - Port-site metastasis after laparoscopic surgery for endometrial carcinoma: two case reports. AB - BACKGROUND: Although studies have reported good results with laparoscopic assisted vaginal hysterectomy (LAVH) to treat endometrial cancer, it has been associated with recurrent disease at trocar insertion sites. Long-term follow-up is necessary to detect possible adverse effects of this technique. CASES: We present two case reports of stage IIB endometrial cancer with port-site metastasis 39 and 48 months after initial surgery with LAVH. CONCLUSION: Although LAVH is a good technique to treat patients with endometrial cancer, port-site metastasis is a possible complication and should be taken into consideration until a randomized study shows the long-term benefits and risks of laparoscopic over standard treatment. PMID- 15661249 TI - Metastases of breast carcinoma to the uterus. Report of two cases, one harboring a primary endometrioid carcinoma, with review of the literature. AB - BACKGROUND: Metastases to the uterus are rare, accounting for less than 10% of all cases of metastases to the female genital tract from extragenital cancers. The endometrium is even less frequently affected by metastases. Lobular carcinoma is the most common type of breast cancer that metastasizes to the uterus. CASES: Two cases of infiltrating lobular carcinoma of the breast metastatic to endometrium and myometrium, one of them harboring an endometrioid adenocarcinoma, are reported. Both patients were on tamoxifen therapy and presented with uterine bleeding. CONCLUSION: To the best of our knowledge, uterine carcinoma serving as recipient of metastatic carcinoma from the breast has not been previously documented. This possibility should be considered when an unusual bimorphic pattern appears in a tumor until proven otherwise. PMID- 15661250 TI - Myxoid leiomyosarcoma of the vulva. AB - BACKGROUND: Myxoid leiomyosarcoma (MLMS) of the vulva is a mesenchymal tumor with only five reported cases in the literature. CASE: We report an 85-year-old woman with a unilateral nonulcerating, painless vulvar mass. According to the patient, the mass slowly enlarged. Initial biopsies were benign. However, clinically, this lesion was suspicious for a soft tissue tumor. Therefore, the mass was removed by a wide local excision. Definitive histology revealed a myxoid leiomyosarcoma of the vulva. At present, 25 months after the operation, the patient is well with no sign of recurrence. CONCLUSIONS: Vulvar myxoid leiomyosarcomas are rare and can be confused with other benign or malignant tumors. It is important to be aware of this rare tumor variant, in order to plan appropriate treatment. PMID- 15661251 TI - Chemoresistant placental site trophoblastic tumor with hilar lymph node metastasis: an unusual site of involvement. AB - BACKGROUND: Placental site trophoblastic tumor (PSTT) is an uncommon variant of gestational trophoblastic diseases. In most cases, disease is confined to the uterus and treated with a simple hysterectomy. However, 30% of these patients will present with metastatic disease. Patients with metastases frequently have progression of disease and die despite aggressive multiagent chemotherapy. CASE: We present a case of 33-year-old female with PSTT and metastases to the hilar lymph nodes of the right lung. Primary surgical treatment consisting of abdominal hysterectomy and unilateral salpingo-oophorectomy was followed by six cycles of EMA/CO (etoposide, methotrexate, actinomycin D, cyclophosphamide, and vincristine) chemotherapy. After the completion of chemotherapy, betahCG titers stayed within normal range, but a repeated CT scan of chest revealed enlargement of the hilar lymph nodes. The patient underwent right thoracotomy with hilar lymph nodes resection. The resected nodules were pathologically consistent with primary PSTT. CONCLUSION: In this case report, we have determined a PSTT with hilar region metastasis other than parenchyma of lung and confirmed the chemoresistant nature of tumor with the guidance of the previous reports. PMID- 15661252 TI - Tubulo-villous adenoma of the vagina. AB - BACKGROUND: Tubulo-villous adenomas are common in the colon and rectum, but extremely rare in the vagina. As far as we know, only two cases of tubulo-villous adenoma have ever been reported. We report the third case of enteric-type tubulo villous adenoma of the vagina. CASE: A 61-year-old woman presented a symptom of intermittent vaginal spotting and a tumor located outside the vaginal introitus near the urethra. The tumor was excised and pathologically confirmed as a tubulo villous adenoma. CONCLUSION: Because some tubulo-villous adenomas in the colon and rectum can progress to the cancer, this case requires long-term follow-up to detect recurrence and malignant transformation. PMID- 15661254 TI - Re: "Selenium as an element in the treatment of ovarian cancer in women receiving chemotherapy". PMID- 15661256 TI - Is tamoxifen an option for patients with endometrial stromal sarcoma? PMID- 15661257 TI - Is antibody therapy of tumor compromised by infusion-related reactions? A case for inhibiting the activity of cyclooxygenase-2. AB - Evidence suggests that amelioration of childhood immune thrombocytopenic purpura and some other autoimmune states by intravenous normal IgG is due to the following chain of events: (1) cross-linking of Fcgamma-receptors on blood effector cells; (2) release of mediators from these cells, often yielding an infusion-related reaction; (3) mediator-induced development of a cytokine field characterized by a mutually stabilizing Th2 polarization of CD4 T lymphocytes and alternative activation of macrophages; (4) selective quiescence of these macrophages towards targets coated with IgG autoantibody, due to increased expression of the macrophage Fcgamma-receptor IIB. In this paper it is postulated that in the field of antibody therapy of tumor, an undesirable delayed or absent subsidence of antibody-coated tumor is due to immunomodulation of the same type as yields amelioration of autoimmunity, and arising from a similar chain of events. If the postulate is correct the chain could usefully be broken at the level of mediator action, possibly by blocking that increased synthesis of prostaglandin E(2) which is catalyzed by the enzyme cyclooxygenase-2. PMID- 15661258 TI - Care versus cure at the end of life. PMID- 15661259 TI - New anti-T cell immunotoxins for the clinic. PMID- 15661260 TI - Fluorescent-labeled DNA probes applied to novel biological aspects of B-cell chronic lymphocytic leukemia. AB - Fluorescent-labeled DNA probes were used to study 52 chronic lymphocytic leukemia (B-CLL) patients for (1) disease progression, (2) angiogenesis genes, (3) T-cell leukemia 1 gene (TCL1), (4) immunoglobulin heavy chain variable region (IGHv) and (5) chromosome 6q. Compared to stable disease, more patients with progressive disease had > or =2 anomalies and a high percentage of neoplastic nuclei. Anomalies of genes for basic fibroblast growth factor, interleukin 4, vascular endothelial growth factor or TCL1 were not detected. Deletions in IGHv occurred in 25% of patients and correlated with IGHv gene expression. Probes for 6q23 detected more deletions in 6q than probes for 6q21. PMID- 15661261 TI - A phase I and pharmacologic study of idarubicin, cytarabine, etoposide, and the multidrug resistance protein (MDR1/Pgp) inhibitor PSC-833 in patients with refractory leukemia. AB - This study was conducted to define the maximum tolerated dose (MTD), dose limiting toxicity (DLT), and pharmacokinetics of idarubicin when administered with and without the P-glycoprotein inhibitor PSC-833 in combination with cytarabine, and etoposide. Fifteen patients with relapsed and refractory acute leukemia were enrolled and received cytarabine as a 7-day continuous infusion, with etoposide and idarubicin administered for any three consecutive days during the cytarabine infusion. Two hours prior to the second dose of idarubicin, PSC 833 administration was initiated. The pharmacokinetics of idarubicin alone and with PSC-833 was assessed at three idarubicin dose levels (6, 8 and 10 mg/m(2)). The MTD of idarubicin in this combination was 8 mg/(m(2) day) with a DLT of oral mucositis. The complete remission rate (on an intent-to-treat basis) for this regimen was 33%, with a median duration of 6 months. The clearance of idarubicin was 140 +/- 200 and 181 +/- 94.3 l/h for idarubicin alone and with PSC-833, respectively. The volume of distribution of the central compartment was 423 +/- 443 and 337 +/- 394 l for idarubicin alone and in combination with PSC-833, respectively. This combination including PSC-833 was well tolerated. Although a pharmacokinetic interaction might have been expected, PSC-833 did not significantly alter the disposition of idarubicin. PMID- 15661262 TI - Prognostic value of structural chromosomal rearrangements and small cell clones with high hyperdiploidy in children with acute lymphoblastic leukemia. AB - In this study, 107 children with acute lymphoblastic leukemia (ALL) were analysed for the presence of hyperdiploidy by cytogenetics and interphase fluorescence in situ hybridisation (I-FISH). Structural aberrations in hyperdiploid cells were investigated by multiple colour FISH (mFISH). Clones with high hyperdiploidy (>50 chromosomes) (HeH) were found in 46 patients (43%). In nine of these (20%), the abnormal clone was present in <20% of the total cell population. There was no significant difference in EFS between those patients with HeH in 2.5-20% or >20% of cells. Structural rearrangements in the HeH clone were found in 10 patients (22%). In this study, HeH karyotypes containing structural aberrations were an indication of a poor prognosis in childhood ALL. PMID- 15661263 TI - Expression of IFITM1 in chronic myeloid leukemia patients. AB - We investigated the peripheral blood gene expression profile of interferon induced transmembrane protein 1 (IFITM1) in sixty chronic myeloid leukemia (CML) patients classified according to new prognostic score (NPS). IFITM1 is a component of a multimeric complex involved in the trunsduction of antiproliferative and cell adhesion signals. Expression level of IFITM1 was found significantly different between the high- and low-risk groups (P = 9.7976 x 10( 11)) by real-time reverse transcription polymerase chain reaction (RT-PCR). Higher IFITM1 expression correlated with improved survival (P = 0.01). These results indicate that IFITM1 expression profiling could be used for molecular classification of CML, which may also predict survival. PMID- 15661264 TI - Elderly patients with Ph+ chronic myelogenous leukemia (CML): results of imatinib mesylate treatment. AB - Thirty-five patients with Ph+ CML aged more than 60 years were treated with imatinib. Twenty-four patients (group A) were in late chronic phase (CP) and eleven patients (group B) were in accelerated/blastic phase (AP/BP). In group A, complete haematological response (CHR) was achieved by all patients; seventeen patients (70.8%) attained a complete cytogenetic response (CCR), one (4.1%) attained a partial CR, one (4.1%) a minor CR (Ph+ 70%) and five (21%) were resistant (Ph+ 100%), toxicity was mild: seven patients had a transient cytopenia, three a skin reaction, one a moderate oedema and one muscular pain. After a median follow-up of 15 months, 1 patient died in progression and 23 patients are alive (2 in BP and 21 in persisting response). In group B, one patient died after 3 months in aplastic phase from sepsis, three patients were resistant and seven patients (63.7%) achieved CHR; of these, four obtained CCR. After a median follow-up of 17 months, 4 patients have died from progressive disease, 6 are alive; 1 in AP and 5 in CHR (4 of them being in CCR). Present data indicate that imatinib is safe also in elderly with clinical results as good as in younger patients. PMID- 15661265 TI - Nurses' perception of adequacy of care for leukemia patients with distress during the incurable phase and related factors. AB - Leukemia patients experience distress during the incurable phase of illness. Adequacy of care (i.e., extent to which care relieves the patients' suffering) was described from the nurses' viewpoint, and institutional factors related to adequacy of care were explored. A self-administered questionnaire was completed by 425 nurses working in 26 hematology wards. Nurses tended to rate the care as inadequate. In particular, care for social distress needs to be improved. Our findings suggest that communication among health professionals, utilization of hospital resources, and palliative care education for staff were important in palliative care for leukemia patients during the incurable phase. PMID- 15661266 TI - Specific targeting of CD33(+) leukemia cells by a natural killer cell line modified with a chimeric receptor. AB - We directed the human natural killer (NK) cell line YT by gene transfer of a humanized chimeric immunoglobulin T cell receptor to CD33, a marker on myeloid leukemias. The chimeric receptor was generated using a CD33 specific single-chain Fv (scFv) fragment based on the humanized antibody HuM195, the human IgG1 Fc domains and the human CD3 zeta signal chain. YT cells transfected by electroporation with the chimeric receptor gene specifically lysed the acute myeloid leukemia (AML) cell line KG1. This gene-modified NK cell line available in unlimited source could be an attractive tool in immunotherapy. PMID- 15661267 TI - DNA-dependent protein kinase enhances DNA damage-induced apoptosis in association with Friend gp70. AB - Friend leukemia virus (FLV) infection strongly enhances gamma-irradiation-induced apoptosis of hematopoietic cells of C3H hosts leading to a lethal anemia. Experiments using p53 knockout mice with the C3H background have clarified that the apoptosis is p53-dependent and would not be associated with changes of cell populations caused by the infection with FLV. In bone marrow cells of FLV + total body irradiation (TBI)-treated C3H mice, the p53 protein was prominently activated to overexpress p21 and bax suggesting that apoptosis-enhancing mechanisms lay upstream of p53 protein in the signaling pathway. Neither of DNA dependent protein kinase (DNA-PK)-deficient SCID mice nor ataxia telangiectasia mutated (ATM) gene knockout mice with the C3H background exhibited a remarkable enhancement of apoptosis or p53 activation on FLV + TBI-treatment indicating that DNA-PK and ATM were both essential. ATM appeared necessary for introducing DNA damage-induced apoptosis, while DNA-PK enhanced p53-dependent apoptosis under FLV infection. Surprisingly, viral envelope protein, gp70, was co-precipitated with DNA-PK but not with ATM in FLV + TBI-treated C3H mice. These results indicated that FLV-infection enhances DNA damage-induced apoptosis via p53 activation and that DNA-PK, in association with gp70, might play critical roles in modulating the signaling pathway. PMID- 15661268 TI - Potential and origin of the hematopoietic population in human skeletal muscle. AB - While mononuclear cells isolated from murine skeletal muscle were shown to be capable of hematopoietic activity, similar hematopoietic cells (HC) recently were reported to exist in primate muscle. We investigated muscle-derived HC from young and adult human subjects. Although hematopoietic stem cells were rare in muscle, their frequency nonetheless was approximately four times greater than in peripheral blood. These cells in muscle appeared to originate from CD45(+) bone marrow cells. Our results suggested an additional function of human skeletal muscle as a reservoir of HC. PMID- 15661269 TI - Up-regulation of DNA methyltransferases DNMT1, 3A, and 3B in myelodysplastic syndrome. AB - Analyzing bone marrow trephines from (myelodysplastic syndrome) MDS patients we show for the first time strong over-expression of the DNA methyltransferases 1, and 3A (and 3B to a much lesser extent) in the myelodysplastic syndrome FAB subtypes refractory anaemia (RA) and refractory anaemia with excess of blasts (RAEB). The increase in mRNA expression was much less pronounced in refractory anaemia with ringed sideroblasts (RARS). Classification according to the new WHO guidelines revealed distinct differences between RCMD and RARS. This elevated mRNA expression most probably contributes to the frequently found aberrant hypermethylation in MDS and might explain the promising clinical response of MDS patients treated with DNMT inhibitors. PMID- 15661270 TI - Molecular modification of a recombinant, bivalent anti-human CD3 immunotoxin (Bic3) results in reduced in vivo toxicity in mice. AB - A novel bivalent single chain fusion protein, Bic3, was assembled consisting of the catalytic and translocation domains of diphtheria toxin (DT(390)) fused to two repeating sFv molecules recognizing human CD3 epsilon of the human T-cell receptor. Historically, problems with these constructs include low yield, toxicity, and reduced efficacy. Instead of using conventional Gly(4)Ser linkers to connect heavy/light chains, aggregation reducing linkers (ARL) were used which when combined with a new SLS-based refolding method reduced aggregation and enhanced the yield of final product. Toxicity was reduced at least 25-fold by repeating the two sFv molecules and adding a portion of the hinge-CH2-CH3 human constant regions. The resulting Bic3 was just as cytotoxic to HPB-MLT.UM T leukemia cells in vitro (IC(50)=4 pmol) as a monovalent construct made with the same DT and sFv. In vivo, Bic3 was effective in a new and aggressive therapy model in which it significantly prolonged survival of scid mice with established human T-cell leukemia (p<0.0001 compared to controls). Importantly, no toxicity measured by weight loss, enzyme function, or histology was observed at the highest dose of Bic3 tested (2000 ug/kg). Bic3 warrants investigation as a new drug for treating T-cell malignancy and other T-cell related disorders. PMID- 15661271 TI - Quantitative RT-PCR of Wilms tumor gene transcripts (WT1) for the molecular monitoring of patients with accelerated phase bcr/abl + CML. AB - The tyrosine kinase inhibitor imatinib inhibits the activity of the bcr/abl fusion protein present in patients with chronic myeloid leukemia. Although in chronic phase patients response to therapy can be monitored by quantitative RT PCR for bcr/abl mRNA transcripts, in advanced disease (accelerated phase or blast crisis) only few patients respond on a molecular level. We investigated Wilms tumor gene (WT1) and bcr/abl mRNA transcripts in 16 accelerated phase CML patients by quantitative real time PCR. In contrast to the bcr/abl mRNA levels the WT1 mRNA levels were indicative for hematologic relapse (n = 6) versus response (n = 10). PMID- 15661272 TI - Meeting report. Acute promyelocytic leukemia-associated coagulopathy, 21 January 2004, London, United Kingdom. AB - Despite successful treatment with all-trans retinoic acid and chemotherapy, life threatening bleeding remains a challenging complication of acute promyelocytic leukemia (APL). Indeed, bleeding and thrombosis are major complications of APL that lead to early death in approximately 10% of patients despite the success of current treatment. This condition may be attributed, in part, to the diffuse activation of coagulation, hyperfibrinolysis, and non-specific proteolytic activity that is observed in patients with APL. Therapeutic agents that induce the differentiation of leukemia cells improve outcomes compared with those observed using chemotherapy alone. They also correct the hyperactivity of the coagulation and fibrinolytic systems, thereby reducing early death from bleeding. Prophylactic therapy with newer anticoagulants may prove beneficial in patients with APL, but this must be confirmed in well-designed, randomized, controlled trials. A workshop was convened 21 January 2004 in London, England, to discuss the clinical and biological aspects of the APL-associated coagulopathy and the application of recent findings to the management of patients with APL. Eight speakers participated in the workshop. This meeting report provides synopses of their presentations and a summary of highlights from the meeting. PMID- 15661273 TI - Chronic myeloid leukemia following kidney transplantation. AB - Immunosuppressed renal recipients are at an increased risk of developing cancer. Leukemias are less frequent than other hematopoietic tumours and development of CML after immunosuppression is rare. We describe a 37-year-old male who presented with left-shifted leukocytosis, hypercellular bone marrow 32 months after the kidney transplant. G-banding karyotype revealed 46,XY,t(9;22)(q34;q11) and the diagnosis of chronic myeloid leukemia was made. This is the 13th case of CML after kidney transplant reported. Whether this CML appeared as a random phenomenon or chemically induced is a matter of debate. Some individuals might have an increased susceptibility to the effects of azathioprine. PMID- 15661275 TI - Degradation of polycyclic aromatic hydrocarbons by a bacterial consortium enriched from mangrove sediments. AB - The biodegradability of a polycyclic aromatic hydrocarbons (PAHs) mixture consisted of fluorene (Fl), phenanthrene (Phe) and pyrene (Pyr) by a bacterial consortium enriched from mangrove sediments under sediment-free and sediment slurry conditions was investigated. The enriched consortium made up of three bacterial strains, namely Rhodococcus sp., Acinetobacter sp. and Pseudomonas sp., had a good PAH degradation capability with 100% degradation of Fl and Phe in sediment-free liquid medium after 4 weeks of growth. The Fl and Phe degradation percentages in sediment slurry were higher than that in liquid medium. Autochthonous microorganisms in sediments also possessed satisfactory PAH degradation capability and all three PAHs were almost completely degraded after 4 weeks of growth. Bioaugumentation (inoculation of the enriched consortium to sediments) showed a positive effect on PAH biodegradation after 1 week of growth. Complete biodegradation of pyrene took longer time than that for Fl and Phe, indicating the enriched bacterial consortium had preference to utilize low molecular weight PAHs. PMID- 15661276 TI - Effect of biosurfactants on crude oil desorption and mobilization in a soil system. AB - Microbially produced biosurfactants were studied to enhance crude oil desorption and mobilization in model soil column systems. The ability of biosurfactants from Rhodococcus ruber to remove the oil from the soil core was 1.4-2.3 times greater than that of a synthetic surfactant of suitable properties, Tween 60. Biosurfactant-enhanced oil mobilization was temperature-related, and it was slower at 15 degrees C than at 22-28 degrees C. Mathematical modelling using a one-dimensional filtration model was applied to simulate the process of oil penetration through a soil column in the presence of (bio)surfactants. A strong positive correlation (R(2)=0.99) was found between surfactant penetration through oil-contaminated soil and oil removal activity. Biosurfactant was less adsorbed to soil components than synthetic surfactant, thus rapidly penetrating through the soil column and effectively removing 65-82% of crude oil. Chemical analysis showed that crude oil removed by biosurfactant contained a lower proportion of high-molecular-weight paraffins and asphaltenes, the most nonbiodegradable compounds, compared to initial oil composition. This result suggests that oil mobilized by biosurfactants could be easily biodegraded by soil bacteria. Rhodococcus biosurfactants can be used for in situ remediation of oil contaminated soils. PMID- 15661277 TI - Application of biopreparation "Rhoder" for remediation of oil polluted polar marshy wetlands in Komi Republic. AB - This paper describes the testing and corresponding results of the preparation "Rhoder" in comparison with several other bioremediation variants during the field trials in Komi Republic throughout 2002-2003. All bioremediation trials were performed on one vast polar marshy wetland polluted by accidental crude oil spill and uncovered by grass. After application of the "Rhoder" at the site, with an area of approximately 2000 m(2), during the cold and rainy summer of 2002 (1.5 months), the level of oil contamination decreased by 20-51%, depending on initial oil pollution (458-738 g/kg dry weight of soil). In the middle of September 2002, the treated site was covered by 70-85% with green grass. Though, during 2003, the "Rhoder" treatment was not practiced, at the end of August 2003, the site was already covered by 85-95% with green grass and the level of oil contamination further decreased by 54-79% from the initial level of oil pollution at the beginning of 2002. These results were much better compared to those from other bioremediation variants applied at this spill. PMID- 15661278 TI - Naphthalene biodegradation kinetics in an aerobic slurry-phase bioreactor. AB - The research was focused on the slurry-phase biodegradation of naphthalene in soil. Among ex situ techniques, the slurry phase offers the advantage of increased availability of contaminants to bacteria. From naphthalene contaminated soil, a Pseudomonas putida M8 strain capable to degrade naphthalene was selected. Experiments were performed in a stirred and oxygenated reactor. In this study, the influence of air flow rate and agitation rate on volatilisation and biodegradation of naphthalene was investigated. The hydrocarbon disappearance, the carbon dioxide production, and the ratio of total heterotrophic and naphthalene-degrading bacteria was monitored. The results obtained confirm that the selected bioremediation technology is successful in the treatment of contaminated soils. PMID- 15661279 TI - In-vessel composting--bioremediation of aged coal tar soil: effect of temperature and soil/green waste amendment ratio. AB - The biodegradation of 16 United States Environmental Protection Agency (USEPA) listed polycyclic aromatic hydrocarbons (PAHs) present in contaminated soil from a manufactured gas plant site was investigated using laboratory-scale in-vessel composting-bioremediation reactors over 8 weeks. The influence of temperature (T, 38, 55, and 70 degrees C) and soil/green waste ratio (S:GW, 0.6:1, 0.7:1, 0.8:1, and 0.9:1) was investigated. A comparative study using a temperature profile during in-vessel composting-bioremediation to meet current regulatory requirements was also investigated. Temperature and amendment ratio were found to be important operating parameters for PAH removal for in-vessel composting bioremediation of aged coal tar-contaminated soil. After 8 weeks of continuous treatment, the highest removal of 16 USEPA PAHs was observed at T=38 degrees C and S:GW=0.8:1 (75.2%). Lower removal of 16 USEPA PAHs was observed for temperature profile treatment (60.8%). We recommend that when conventional composting processes using temperature profiles to meet regulatory requirements for pathogen control need to be used, these should start with a prolonged mesophilic stage (6 weeks in this investigation) followed by thermophilic, cooling, and maturation stages. PMID- 15661280 TI - An insight into soil bioremediation through respirometry. AB - Respirometric tests on a soil contaminated by crude oil were performed. Continuous measurements of oxygen and carbon dioxide concentrations and temperature in the soil atmosphere resulted in a large volume of data. Time series and system identification theories were used to analyze data as a biological signal, allowing us to detect some particularities related to daily cycles of the studied variables as well as its time relationships through autocorrelation and cross-correlation functions. Using system identification techniques, it was possible to build black box models, namely autoregressive moving average models which enable to predict oxygen concentration at the outlet in a good agreement with measured data. PMID- 15661281 TI - Assessment of natural or enhanced in situ bioremediation at a chlorinated solvent contaminated aquifer in Italy: a microcosm study. AB - A microcosm study was used to assess the potential for in situ natural or enhanced bioremediation at a chloroethane- (i.e., tetrachloroethane, TeCA) and chloroethene-contaminated (i.e., tetrachloroethene, PCE; trichloroethene, TCE) groundwater in Northern Italy. All the live microcosms were positive for dechlorination, indicating the presence of an active native dechlorinating population in the subsurface. All the tested electron donors (i.e., yeast extract, lactate, butyrate, hydrogen) promoted enhanced dechlorination of chlorinated contaminants. Lactate- and butyrate-amended microcosms performed the best, and also dechlorinated the solvents past cis-dichloroethene (cis-DCE). The microcosm bioaugmented with a PCE-dechlorinating mixed culture containing Dehalococcoides spp. dechlorinated groundwater contaminants to DCE, vinyl chloride (VC), and ethene (ETH). In conclusion, results from this microcosm study indicate the potential for enhancing full dechlorination at the contaminated site, through a proper addition of a suitable electron donor (e.g., lactate or butyrate) and/or through bioaugmentation with a Dehalococcoides-containing culture. PMID- 15661282 TI - Biodegradation of lindane by Pleurotus ostreatus via central composite design. AB - The degradation of lindane was studied in liquid-agitated cultures using a commercial strain of the fungus Pleurotus ostreatus as the biodegrading organism. The biodegradation was accomplished with the action of extracellular oxidative enzymes, produced by the fungus to decompose woody substrates. Enzyme activities of manganese peroxidase and laccase were measured in a liquid mineral medium. An orthogonal Central Composite Design of experiments was used to construct second order response surfaces with the fungus growth, final pH and the lindane biodegradation as optimization parameters. The initial lindane concentration, the nitrogen content, the incubation time and the temperature were used as design factors. Optimal conditions found for all these parameters will be used for the continuation of this project aiming at the bioremediation of contaminated sites with persistent organic pollutants such as lindane. PMID- 15661283 TI - Colonization of surfaces by phenolic compounds utilizing microorganisms. AB - The aim of the present study is to determine optimal adhesive interaction of phenolic compounds utilizing Candida maltosa and Rhodococcus erythropolis when adhering to kaolin, silicone, synthetic foil (Steriking R40) and fluorinated silicones, comparing cell and support surface hydrophobicity. In parallel, the interfering effect of detergents was investigated. Data obtained show that the less hydrophobic supports display high initial cell adhesion when contacted with the cell type with a lower surface hydrophobicity (yeast cell) but most stable yeast biofilms are those formed on highly hydrophobic fluorinated silicones. On the other hand, support hydrophobicity has no effect on bacterial cell detachment; however, bacterial biofilms are denser when growing on more hydrophobic supports. Both detergents interfere (independently on the cell type) with the early and late phases of biofilm development. PMID- 15661284 TI - Azo dye biodegradation by microbial cultures immobilized in alginate beads. AB - Microbial degradation of azo dyes usually starts in anaerobic conditions with a reductive cleavage of the azo bond, followed by an aerobic step necessary for the degradation of the aromatic amines formed. Because some reductive processes take place also in presence of molecular oxygen, a one-step azo dye degrading process has been investigated. A microbial consortium able to degrade ethyl orange in aerobic conditions has been selected and immobilized in alginate beads coated with polyacrylamide resin. Different concentrations of ethyl orange have been completely degraded in the presence of 1% glucose or starch as cosubstrates, and different beads preparation procedures have been studied to determine the best condition for microbial degradation. The catalytic activity of the immobilized consortium improved during five serial processes carried out for 30 days at room temperature. Three pure cultures were then isolated from the consortium. The one with the greatest degrading activity, a filamentous fungus, had a degradative capacity similar to that of the whole consortium. PMID- 15661285 TI - Extent of sonochemical degradation and change of toxicity of a pharmaceutical precursor (triphenylphosphine oxide) in water as a function of treatment conditions. AB - The degradation of triphenylphosphine oxide (TPPO) in water, a toxic compound typically found in effluents from the pharmaceutical industry, by means of ultrasonic irradiation at 20 kHz has been investigated with emphasis on the effect of various parameters on conversion and acute toxicity. Experiments were carried out at liquid volumes of 50 and 80 ml, electric power outputs of 125, 187.5 and 250 W, initial TPPO concentrations of 10, 100 and 350 mg/L and temperatures of 5, 20, 35, 50 and 70 degrees C. TPPO conversion was found to increase with increasing power output and decreasing initial concentration and temperature. Measurements of dissolved total carbon showed that liquid-phase degradation by-products were more stable to ultrasonic irradiation than TPPO. Addition of t-butanol as a radical scavenger at a concentration of 1000 mg/L nearly completely suppressed TPPO degradation. Conversely, addition of radical promoters (Fe(2+) ions or H(2)O(2)) had a positive effect on degradation. Acute toxicity to marine bacteria vibrio fischeri was measured before and after ultrasonic irradiation. At the conditions employed in this study, irradiated TPPO samples were always more toxic than TPPO itself with toxicity levels being a function of treatment conditions. PMID- 15661286 TI - Recent advances in the bioremediation of arsenic-contaminated groundwater. AB - The biological treatment of groundwater is used primarily to remove electron donors from water sources, providing (biologically) stable drinking water, which preclude bacterial regrowth during subsequent water distribution. To the electron donors belong also the dissolved metal cations of ferrous iron and manganese, which are common contaminants found in most (anaerobic) groundwater. The removal of iron and manganese is usually accomplished by the application of chemical oxidation and filtration. However, biological oxidation has recently gained increased importance and application due to the existence of certain advantages, over the conventional physicochemical treatment. The oxidation of iron and manganese is accelerated by the presence of certain indigenous bacteria, the so called "iron and manganese oxidizing bacteria." In the present paper, selected long-term experimental results will be presented, regarding the bioremediation of natural groundwater, containing elevated concentrations of iron and arsenic. Arsenic is considered as a primary pollutant in drinking water due to its high toxicity. Therefore, its efficient removal from natural waters intended for drinking water is considered of great importance. The application of biological processes for the oxidation and removal of dissolved iron was found to be an efficient treatment technique for the simultaneous removal of arsenic, from initial concentrations between 60 and 80 microg/l to residual (effluent) arsenic concentrations lower than the limit of 10 microg/l. The paper was focused on the removal of As(III) as the most common species in anaerobic groundwater and generally is removed less efficiently than the oxidized form of As(V). To obtain information for the mechanism of As(III) removal, X-ray photoelectron spectroscopy (XPS) analyses were applied and it was found that As(III) was partially oxidized to As(V), which enabled the high arsenic removal efficiency over a treatment period of 10 months. PMID- 15661287 TI - Effect of indigenous bacterial activity on arsenic mobilization under anaerobic conditions. AB - Batch biochemical leaching tests were carried out to investigate the mobility of arsenic from a contaminated soil collected from a French gold mining site. The specific objective of this research was to examine the effect of indigenous bacterial activity on arsenic mobilization under anaerobic conditions. In a first step, physical and chemical characterizations were performed to provide data concerning the liquid-solid partitioning and mobility of arsenic and other inorganic constituents. In a second step, batch bioleaching tests were conducted in shaker flasks to determine the effect of indigenous bacterial activity under different anaerobic conditions (i.e., addition of mineral nutrients and carbon sources) on arsenic mobilization. Results indicated that arsenic release during contact with deionized water was limited by its very low solubility in the interstitial solution and by the stability of the different arsenic compounds formed with the amorphous solid phases of the soil (mainly iron (oxy)hydroxides). However, an increased mobilization potential was observed over the long term under anaerobic conditions with indigenous bacterial activity enhanced by the addition of carbon sources. PMID- 15661288 TI - Comparison of differences between copper bioaccumulation and biosorption. AB - Biological methods for wastewater treatment are becoming more accepted all over the world. The method, which makes use of accumulating abilities of living cells, is called bioaccumulation. In contrast with it, biosorption takes advantage of sequestering capabilities of dry or dead cells, which is technologically advantageous. The differences between bioaccumulation and biosorption of copper from model solutions were studied in this work. Application of living algal cells has some disadvantages. Copper significantly damages the surface of living cells, which results in partial loss of cell-binding abilities and release of accumulated copper back into solution. The binding capacity of living cells is significantly lower than that of dead cells. There is also a possibility of desorption and reuse of biomass in case of biosorption. PMID- 15661289 TI - Selenite precipitation by a rhizospheric strain of Stenotrophomonas sp. isolated from the root system of Astragalus bisulcatus: a biotechnological perspective. AB - A bacterial strain (SeITE02), related to the species Stenotrophomonas maltophilia and resistant to selenite (SeIV) up to 50 mM in the growth medium, was isolated from rhizospheric soil of a selenium hyperaccumulator plant, the legume Astragalus bisulcatus. The influence of SeIV on the active growth of this Se tolerant bacterial strain has been investigated in oxic conditions, along with the isolate's ability to reduce selenite to elemental selenium (Se(0)). Interestingly, concentrations of 0.5 mM SeIV were wholly reduced by strain SeITE02 in liquid culture within 52 h. Moreover, 87% of SeIV added to the growth medium at the initial concentration of 2.0 mM underwent again reduction in 120 h. Actually, a selenite-mediated induction of a sort of adaptive response to detrimental SeIV effects magnified the efficiency of SeITE02 in reducing this toxic oxyanion. Furthermore, the SeIV influence on cell morphology of strain SeITE02 was evidenced by phase-contrast and electron microscopy analyses. In particular, transmission electron microscopy (TEM)-energy-dispersive X-ray (EDX) analysis of S. maltophilia strain SeITE02, grown in presence of SeIV, showed electron-dense Se(0) granules either in the cell cytoplasm or in the extracellular space. Therefore, the capability of strain SeITE02 to quickly reduce soluble and harmful SeIV to insoluble and unavailable Se(0) may be looked at as a promising exploitable option for the setup of low-cost biological treatments tailored to manage contamination in selenium-laden effluents. PMID- 15661290 TI - Photosynthesis and growth responses of giant reed (Arundo donax L.) to the heavy metals Cd and Ni. AB - Giant reed (Arundo donax L.) was grown on surface soil and irrigated with mixed heavy metal solutions of Cd(II) and Ni(II) to study the impact of these heavy metals on its growth and photosynthesis. The tested concentrations were 5, 50, and 100 ppm for each heavy metal against the control and resulted in high cadmium and nickel (DTPA extractable) concentrations in the top zone of the pot soil. The examined parameters, namely, stem height and diameter, number of nodes, fresh and dry weight of leaves, and net photosynthesis (Pn) were not affected, indicating that plants tolerate the high concentrations of Cd and Ni. As giant reed plants are very promising energy plants, they can be cultivated in contaminated soils to provide biomass for energy production purposes. PMID- 15661291 TI - Ability of transgenic poplars with elevated glutathione content to tolerate zinc(2+) stress. AB - Phytoremediation potentials of four poplar lines, Populus nigra (N-SL clone), Populus canescens, and two transgenic P. canescens clones were investigated using in vitro leaf discs cultures. The transgenic poplars overexpressed a bacterial gene encoding gamma-glutamylcysteine synthetase in the cytosol (11ggs) or in the chlopoplasts (6LgI), and therefore, they contained an elevated level of glutathione. Leaf discs of poplar clones were exposed to different concentrations of ZnSO(4) for 21 days. Zinc(2+) was phytotoxic only at high concentrations (10( 2) to 10(-1) M) at all P. canescens lines, but P. nigra was more sensitive. Transgenic poplars showed elevated heavy metal uptake as compared to the nontransformed clones. Treatments with zinc(2+) strongly induced the activity of glutathione S-transferase enzyme in untransformed poplar lines but to a lesser extent in the transgenic clones. These results suggest that transgenic poplars are more suitable for phytoremediation of soils contaminated with zinc(2+) than wild-type plants. PMID- 15661292 TI - A whole-plant mathematical model for the phytoextraction of lead (Pb) by maize. AB - Phytoextraction is a technology that uses plants to remove heavy metals from contaminated soils. Although it is economically attractive compared to other methods, little attention has been paid to the mathematical modeling of the mechanisms involved. In this work, we simulate the phytoextraction of Pb using a mechanistic system dynamics modelling approach and the physiology model of maize (Zea mays) as a model system as it is a good Pb accumulator and translocator. Simulation results showed that precipitation is the most important mechanism related to the uptake of Pb from the ground. The most important model parameters have been identified through sensitivity analysis. PMID- 15661293 TI - Possible involvement of plant ABC transporters in cadmium detoxification: a cDNA sub-microarray approach. AB - As a nontolerant plant to a large number of toxic compounds, Arabidopsis thaliana is a suitable model to study regulation of genes involved in response to heavy metals. Using a cDNA-microarray approach, we identified some ABC transporters that are differentially regulated after cadmium treatments, making them putative candidates for being involved in Cd sequestration and redistribution in plants. Regarding yeast and fission yeast, in which Cd is able to form complexes either with glutathione (GSH) or phytochelatins (PC) subsequently transported into vacuoles via ABC transporters, it is also very likely that some plant ABC transporters are able to transport GS(2)-Cd or PC-Cd complexes into subcellular compartments or outside of the cell. The characterization of such transporters is of great interest for developing molecular biology approaches in phytoremediation. PMID- 15661294 TI - Acute and sublethal toxicity tests to monitor the impact of leachate on an aquatic environment. AB - In this study, a specific landfill leachate (1200 mg l(-1) COD and 600 mg l(-1) BOD(5)) was used to develop a standardised short-term acute and longer-term sublethal ex-situ toxicity testing programme, in order to determine the potential ecological implications of leaching contaminants reaching the water table. Bioassays were undertaken with juvenile Gammarus pulex and Asellus aquaticus macro-invertebrates. Preliminary acute test variables included static and static renewed flow rates for 96-h, starved and fed specimens, and aerobic and oxygen depleting conditions. However, regardless of any test variable, the lethal concentration (LC(50)) for A. aquaticus remained at 12.3% v/v leachate in deionized water, whilst that for G. pulex was only 1%. Sublethal toxicity was judged on the basis of frequency of births and the growth rate of newly born individuals. Tests showed that even a dilution as high as 1:66- would influence the fecundity of a Gammarus population, whilst a dilution of 1:20 would affect the size of an Asellus breeding colony. PMID- 15661295 TI - Measuring the antioxidant activity of olive oil mill wastewater using chemiluminescence. AB - A sensitive and simple procedure is described for determining the total phenolic/antioxidant levels of olive oil mill wastewater (OMW), using for the first time Co(II)/ethylenediaminetetracetic acid (EDTA)-induced luminol chemiluminescence. Olive oil wastewater samples were tested for their composition in simple phenolic compounds as a function of the extraction system (two- and three-phase centrifugation systems). The results revealed that the three-phase system had a stronger antioxidant activity and a higher total phenolic content than the two-phase system. The relationship between antioxidant values and total phenolic content is also discussed. PMID- 15661296 TI - Sonochemical reduction of the antioxidant activity of olive mill wastewater. AB - The use of ultrasonic irradiation to reduce the antioxidant activity of olive oil mill wastewaters (OMWs) originating from two-phase and three-phase decanters was examined. Sonication of diluted OMW samples was conducted at ultrasonic frequencies of 24 and 80 kHz, an applied power varying between 75 and 150 W, and liquid bulk temperatures varying between 25 and 60 degrees C. At the conditions in question, the reduction in antioxidant activity was found to increase with decreasing temperature and increasing power and frequency. Addition of NaCl in the samples also appeared to enhance reduction. Antioxidant activity of OMW samples was assessed using the recently developed Co(II)/ethylenediaminetetraacetic acid (EDTA)-induced luminol chemiluminescence analytical protocol, while the total phenolic load was measured according to the Folin-Ciocalteu method. PMID- 15661297 TI - Treatment of olive mill effluents Part I. Organic matter degradation by chemical and biological processes--an overview. AB - Olive mill effluents constitute a serious environmental problem in the Mediterranean Sea region due to the unique features associated with this type of agro-waste (i.e. seasonal and localized production, high and diverse organic load, low flow rates). Therefore, it is not surprising that research efforts have been directed towards the development of efficient treatment technologies including various physico-chemical and biological processes. This work reviews recent advances regarding olive mill effluent treatment with emphasis given on biological and chemical degradation processes. PMID- 15661298 TI - Treatment of olive mill effluents Part II. Complete removal of solids by direct flocculation with poly-electrolytes. AB - The pre-treatment of three different olive oil processing effluents by means of direct flocculation (i.e. without prior coagulation) was investigated. Four cationic and two anionic poly-electrolytes were tested and most of them were found capable of removing nearly completely total suspended solids (TSS) as well as reducing considerably the concentration of chemical (COD) and biochemical oxygen demand (BOD(5)) without altering solution pH. Flocculant dosage was crucial to achieve effective separation. For three cationic and one anionic poly electrolytes, the minimum dosage required to initiate separation was about 2.5-3 g/L. The remaining two poly-electrolytes failed to cause separation even at dosages as high as 7 g/L. Lime and ferric chloride were also tested as reference coagulants and found quite effective in terms of TSS removal although the degree of COD reduction was generally lower than that with poly-electrolytes. However, lime treatment would require greater dosages and longer treatment times than that with poly-electrolytes and would also increase considerably solution pH. A preliminary cost analysis showed that lime treatment for complete solids removal was generally less costly than that with poly-electrolytes presumably due to its low market price. Nonetheless, cost-benefits may be defied by several drawbacks associated with the use of lime. PMID- 15661299 TI - Influence of olive mill wastewater in composting and impact of the compost on a Swiss chard crop and soil properties. AB - The suitability of olive mill wastewater (OMW) for composting was studied by the addition of this liquid waste to a mixture of cotton gin waste and sewage sludge, and its composting was compared with that of another pile of similar composition, but without olive mill wastewater. Both piles were composted by the Rutgers static pile system in a pilot plant. To study the effects of both composts on plant yield and soil properties, a plot experiment was carried out with Swiss chard (Beta vulgaris L. var. cicla). Five treatments were applied: mineral fertiliser and two doses (30 and 60 tons ha(-1)) of both composts. The olive mill wastewater addition produced a compost with lower organic matter and nitrate concentrations, higher electrical conductivity, and a stabilised and humified organic matter similar to that of the compost produced without olive mill wastewater. The olive mill wastewater compost application to soil did not injure plants, producing a similar plant yield to both compost without olive mill wastewater and inorganic fertiliser. Also, the accumulation of potentially toxic heavy metals in plants cultivated with organic or mineral fertilisers did not reveal significant differences. The olive mill wastewater compost application to soil also improved the chemical and physicochemical properties of the soil. PMID- 15661300 TI - Electrical stimulation of excitable tissue: design of efficacious and safe protocols. AB - The physical basis for electrical stimulation of excitable tissue, as used by electrophysiological researchers and clinicians in functional electrical stimulation, is presented with emphasis on the fundamental mechanisms of charge injection at the electrode/tissue interface. Faradaic and non-Faradaic charge transfer mechanisms are presented and contrasted. An electrical model of the electrode/tissue interface is given. The physical basis for the origin of electrode potentials is given. Various methods of controlling charge delivery during pulsing are presented. Electrochemical reversibility is discussed. Commonly used electrode materials and stimulation protocols are reviewed in terms of stimulation efficacy and safety. Principles of stimulation of excitable tissue are reviewed with emphasis on efficacy and safety. Mechanisms of damage to tissue and the electrode are reviewed. PMID- 15661301 TI - A microcapsule technique for long-term conduction block of the sciatic nerve by tetrodotoxin. AB - Tetrodotoxin (TTX) is a selective blocker of voltage-gated Na+ channels that is used to block action potentials in vitro and in vivo. Maintaining a sufficiently high local concentration of TTX in vivo to block conduction in a peripheral nerve is technically demanding and carries a risk of systemic toxicity. We report that slow diffusion of TTX out of a microcapsule (glass capillary) inserted beneath the epineurium of the sciatic nerve, with a loose cuff around the nerve, combines high blocking efficacy with low systemic toxicity in rats and mice. The local anaesthesia and motor paralysis was stable for at least 4-6 weeks. The conduction block was reversible and did not cause any obvious nerve injury. Low cost and simple surgical implementation make this new system an interesting alternative to existing long-term drug delivery methods. PMID- 15661302 TI - A new device and protocol for combining TMS and online recordings of EEG and evoked potentials. AB - We describe an electroencephalographic (EEG) device and protocol that allows recording of electrophysiological signals generated by the human brain during transcranial magnetic stimulation (TMS) despite the TMS-induced high-voltage artifacts. The key hardware components include slew-rate limited preamplifiers to prevent saturation of the EEG system due to TMS. The protocol involves artifact subtraction to isolate the electrophysiological signals from residual TMS-induced contaminations. The TMS compatibility of the protocol is illustrated with examples of two data sets demonstrating the feasibility of the approach in the single-pulse TMS design, as well as during repetitive TMS. Our data show that both high-amplitude potentials evoked by visual checkerboard stimulation and low amplitude steady-state oscillations induced by auditory click-trains can be retrieved with the present protocol. The signals recorded during TMS perfectly matched control EEG responses to the same visual and auditory stimuli. The main field of application of the present protocol is in cognitive neuroscience complementing behavioral studies that use TMS to induce transient, 'virtual lesions'. Combined EEG-TMS techniques provide neuroscientists with a unique method to test hypothesis on functional connectivity, as well as on mechanisms of functional orchestration, reorganization, and plasticity. PMID- 15661303 TI - Monitoring thiobarbituric acid-reactive substances (TBARs) as an assay for oxidative damage in neuronal cultures and central nervous system. AB - Oxidative stress is a pivotal factor in neuronal degeneration. A simple method to quantify oxidative damage in culture and in situ is therefore important for studies of neurodegeneration. We present herein modifications of the standard assay for thiobarbituric acid-reactive substances (TBARs) for analyses of both cell cultures and brain tissue homogenates. Since the TBAR assay measures end point oxidative damage, it is useful to assess the overall impact of oxidative stress-inducing and neuroprotective agents; interpretation is not potentially confounded by the presence or absence of transient products of oxidative damage. PMID- 15661304 TI - Extraction of the average and differential dynamical response in stimulus-locked experimental data. AB - In optical imaging experiments of primary visual cortex, visual stimuli evoke a complicated dynamics. Typically, any stimulus with sufficient contrast evokes a response. Much of the response is the same regardless of which stimulus is presented. For instance, when oriented drifting gratings are presented to the visual system, over 90% of the response is the same from orientation to orientation. Small differences may be seen, however, between the responses to different orientations. A problem in the analysis of optical measurements of the response to stimulus in cortical tissue is the distinction of the 'global' or 'non-specific' response from the 'differential' or 'stimulus-specific' response. This problem arises whenever the signal of interest is the difference in response to various stimuli and is evident in many kinds of uni- and multivariate data. To this end, we present enhancements to a frequency-based method that we previously introduced called the periodic stacking method. These enhancements allow us to separately estimate the dynamics of both the average signal across all stimuli (the 'global' response) and deviations from the average amongst the various stimuli (the 'stimulus-specific' response) evoked in response to a set of stimuli. We also discuss improvements in the signal-to-noise ratio, relative to standard trial averaging methods, that result from the data-adaptive smoothing in our method. PMID- 15661305 TI - A new stabilizing craniotomy-duratomy technique for single-cell anatomo electrophysiological exploration of living intact brain networks. AB - Standard large craniotomies induce undesirable brain motions during intracellular recordings in whole animal preparations. Practically all of the papers available in the literature outline a number of specific methodological approaches designed to avoid this inconvenience. Our study describes a new craniotomy-duratomy, which consists of the maintenance of a thin bone membrane and dura mater surrounding the small hole opened for lowering the recording micropipette. This new surgical preparation avoids brain movements by keeping the brain's volume constant within the cranial cavity and does not require additional technical procedures. It is an all-purpose surgical technique, although it was developed in anaesthetized rats while studying spatio-temporal dynamics of cellular interactions associated with thalamocortical oscillations. It significantly improves both the precision of stereotaxic approaches and the success rate of single-cell recordings (e.g., current-clamp intracellular and paired recordings) compared to standard craniotomy/electrophysiology techniques. PMID- 15661306 TI - Pulser: user-friendly, graphical user-interface based software for controlling stimuli during data acquisition with Spike2 for Windows. AB - This paper describes software that runs in the Spike2 for Windows environment and provides a versatile tool for generating stimuli during data acquisition from the 1401 family of interfaces (CED, UK). A graphical user interface (GUI) is used to provide dynamic control of stimulus timing. Both single stimuli and trains of stimuli can be generated. The pulse generation routines make use of programmable variables within the interface and allow these to be rapidly changed during an experiment. The routines therefore provide the ease-of-use associated with external, stand-alone pulse generators. Complex stimulus protocols can be loaded from an external text file and facilities are included to create these files through the GUI. The software consists of a Spike2 script that runs in the host PC, and accompanying routines written in the 1401 sequencer control code, that run in the 1401 interface. Handshaking between the PC and the interface card are built into the routines and provides for full integration of sampling, analysis and stimulus generation during an experiment. Control of the 1401 digital-to analogue converters is also provided; this allows control of stimulus amplitude as well as timing and also provides a sample-hold feature that may be used to remove DC offsets and drift from recorded data. PMID- 15661307 TI - Analysis of behavioral asymmetries in the elevated plus-maze and in the T-maze. AB - When studying functional asymmetries in normal laboratory rats, several behavioral tests have been applied and proven their utility, including turning in rotometers or open-fields, handedness in paw usage, T-maze alternation, and others. Here, we analyzed male Wistar rats in two tests, namely the elevated plus maze and the T-maze. In these tests, behavioral asymmetries are rather likely to occur, since the animals have to show several types of turns towards the left or right when ambulating through these environments. In a first study using the plus maze, we provide detailed data on (A) the types of turns which the animals showed when changing their direction within arms (i.e., 180 degrees turns), and (B) the types of turns when proceeding from one arm to an adjacent one (i.e., 90 degrees turns). With respect to asymmetry, we found moderate biases in favor of the right. On the 1st day of plus-maze testing, there was a trend for more rightward turns within arms. On the 2nd day of testing, there was a trend for turns towards the right when alternating between arms of the plus-maze. In a 2nd study, we asked for asymmetries in the plus-maze in animals, which had been treated acutely with the psychostimulatory amphetamine analogue 3,4-methylene dioxymethamphetamine (MDMA). Psychostimulants drugs, especially amphetamine, have repeatedly been used before in work on functional asymmetry, since they can enhance or reveal asymmetries in normal rats. MDMA had dose-dependent effects on activity, which affected turns within arms, and turns between arms; however, there was only sparse evidence with respect to asymmetry. Interestingly, and if at all, asymmetry was in favor of the right. Finally, we present data for behavior in the T-maze, where we used a spontaneous test version, that is, the animals could explore the maze but had no task to solve. Asymmetries were measured as turns within the start arm (180 degrees), and as left- or rightward turns between arms (90 degrees ) at the T-point of the maze. In both measures, we again obtained evidence for asymmetries in favor of the right. This work supports previous studies showing that the T-maze is suitable to analyze behavioral asymmetries in rats. In addition, it provides new evidence with respect to the elevated plus-maze, indicating that this standard tool of anxiety research may also be useful in research on behavioral asymmetries and their underlying brain mechanisms. Behavioral biases in favor of the right, as shown here, have often been reported before, especially with Wistar rats. Such biases should be taken into account, since they can serve as an approach to study brain/behavior relationships, and since they may affect the outcome of physiological manipulations or behavioral trainings. PMID- 15661308 TI - Computerized analysis of audible and ultrasonic vocalizations of rats as a standardized measure of pain-related behavior. AB - The behavioral assessment of experimental pain is essential for the analysis of pain mechanisms and the validation of therapeutic targets. Arthritic pain, in particular, is significantly associated with negative affective states and disorders. Here we present a standardized method for the quantitative analysis of audible and ultrasonic (25 +/- 4 kHz) vocalizations in awake rats as a measure of higher integrated behavior in a model of arthritic pain. A bat detector and a condenser microphone were used to record ultrasonic and audible vocalizations, respectively, in response to innocuous and noxious mechanical stimulation of the knee before and after induction of acute arthritis in one knee. A computerized system was used to analyze number and duration of the filtered signals. For the behavioral tests, the animal was placed in a customized recording chamber to ensure consistent stimulus application and stable recordings and to eliminate any movement-induced noise. Noxious stimuli produced stronger vocalizations than innocuous stimuli. Both audible and ultrasonic vocalizations to innocuous (allodynia) and noxious (hyperalgesia) stimuli increased after the induction of acute arthritis. These changes were accompanied by increased knee joint circumference, lowered hind limb withdrawal thresholds and reduced exploratory behavior in the same animals. The computerized analysis of audible and ultrasonic vocalizations is a valid, quantitative, reliable and convenient method to measure pain-related behavior. PMID- 15661309 TI - A simple method to study cellular migration. AB - We describe here a simple and fast method for the characterisation of cell motion. By projecting on a single plane different positions of the cell a ribbon is generated, whose characteristics can be related to the type of motion. The proposed method allows both to determine, very quickly, the motility of a population of cells and to investigate and characterise properties of a single cell's motion. The methodology presented here can be applied to a large range of cell movement and also adapted and extended to other problems involving biological motion. PMID- 15661310 TI - Design of a head fixation device for experiments in behaving monkeys. AB - We have designed a new device for head fixation of behaving monkeys. The fixation device consists of a duralumin head ring mounted with four screw holders. It is firmly fixed to the animal's skull with four stainless steel screw pins. The head ring is then attached to a primate chair in any desirable position and angle using a set of adjustable plates. The device has been used for behavioral training that requires accurate gaze monitoring and for recording single-unit activity over a several-month period. The advantage of our device is that it is simple to use; it can be attached readily without major surgical procedures and it can be quickly removed when experiments are not running. This head fixation system is suitable for behavioral experiments and single-unit recording studies. It may also be applicable for studies on functional imaging of the macaque brain, by constructing it of non-magnetic materials. PMID- 15661311 TI - Visual landmark orientation by flying bats at a large-scale touch and walk screen for bats, birds and rodents. AB - Orientation depends on multi-modal information about the locally perceptible environment (local view) in many situations. We developed a behavioural paradigm for investigating visual orientation of flying bats based on a large-scale touch screen (1.2 m x 1.8 m). It functions by a grid of rows and columns of infra-red beams just in front of a screen with back-projected visual stimuli. Approaching animals interrupt the beams and thus permit automatic recording of the time and place of an animal's locational choice. We used it as a vertical touch surface. Installed as a horizontal walk surface, it may also serve as a more natural 'firm ground', circular arena analogue to the 'Morris water maze' for investigating orientation behaviour and spatial cognition from rodents to birds while offering automatic real-time recording of paths, times and latencies with enhanced possibilities to score details of motor behaviour and to control stimuli interactively. Bats offer a unique possibility to investigate the use of both echo-acoustic and visual information processing pathways for the process of self localization and orientation. In our first experiment, a bat was presented with five identical targets, one central and four peripheral and had to choose the central target. After task acquisition, the array was shifted by the distance between targets, so that a formerly peripheral landmark was now in the absolute location of the formerly central target. At small inter-target distances, the bat 'went with' the array, and chose the new central target (at a new absolute location). With 30 cm or more of inter-target distance (60 cm across the landmark configuration), however, the bat went with absolute location, and chose a peripheral target. In experiment 2, the bat was presented with two landmarks 30 cm apart and an unmarked target located at midline beneath them. On tests, the landmarks either maintained training distance or were expanded to 50 cm apart. On such expansion tests, the bat chose most the location at the correct vector from the right landmark. This showed that the bat first identified a single landmark by the configuration and then applied a previously learnt vector (angle and distance) to locate the target. Glossophaga did not orient by pure angular geometry between landmarks and target. PMID- 15661312 TI - CoCoDat: a database system for organizing and selecting quantitative data on single neurons and neuronal microcircuitry. AB - We present a novel database system for organizing and selecting quantitative experimental data on single neurons and neuronal microcircuitry that has proven useful for reference-keeping, experimental planning and computational modelling. Building on our previous experience with large neuroscientific databases, the system takes into account the diversity and method-dependence of single cell and microcircuitry data and provides tools for entering and retrieving published data without a priori interpretation or summarizing. Data representation is based on the framework suggested by biophysical theory and enables flexible combinations of data on membrane conductances, ionic and synaptic currents, morphology, connectivity and firing patterns. Innovative tools have been implemented for data retrieval with optional relaxation of search criteria along the conceptual dimensions of brain region, cortical layer, cell type and subcellular compartment. The relaxation procedures help to overcome the traditional trade-off between exact, non-interpreted data representation in the original nomenclature and convenient data retrieval. We demonstrate the use of these tools for the construction, tuning and validation of a multicompartmental model of a layer V pyramidal cell from the rat barrel cortex. CoCoDat is freely available at . Its application is scalable from offline use by individual researchers via local laboratory networks to a federation of distributed web sites in platform independent XML format using Axiope tools. PMID- 15661313 TI - Optical assessment of motoneuron function in a "twenty-four-hour" acute spinal cord slice model from fetal rats. AB - In acute slice preparations of most brain regions, neuronal functions are preserved for only few hours. Since the effects of growth factors or neurotoxic agents are often manifested beyond this time scale, corresponding studies are typically performed on cultured cells. However, cell cultures are generated and maintained under vastly different conditions that can grossly alter neuronal properties. For example, glutamate application to motoneuronal cultures has been reported to modulate neurite formation in some studies while in others it has been reported to kill cells. Here, we have examined whether acute spinal cord slices from rat fetuses can be used within a time window of 24 h for assessment of long-term effects of neuromodulators. In these slices, we have studied the action of glutamate on lumbar motoneurons loaded with fura-2 and rhodamine-123 to monitor intracellular Ca2+ ([Ca2+]i) and mitochondrial potential (Deltapsi), respectively. Further, loading with fura-2 or propidium iodide allowed for morphological assessment of cell viability and death, respectively. Pulses (15 s) or 1 h application of glutamate (300 microM) evoked a moderate (approximately 500 nM) [Ca2+]i rise, but no change of Deltapsi. Even after 24 h, no glutamate induced cell death was observed and glutamate pulse-evoked [Ca2+]i transients were comparable to controls. The data demonstrate that glutamate does not deregulate [Ca2+]i homeostasis in fetal motoneurons in situ. We propose that acute spinal cord slices from perinatal rodents are a robust model that allows for analysis of neuronal properties and cell viability within a time window of at least 24 h. PMID- 15661314 TI - A statistically based density map method for identification and quantification of regional differences in microcolumnarity in the monkey brain. AB - We present a statistical density map method derived from condensed matter physics to quantify microcolumns, the fundamental computational unit of the cerebral cortex. This method provides measures for microcolumnar strength, width, spacing, length, and periodicity. We applied this method to Nissl-stained 30 microm thick frozen sections from areas 46, TE, and TL of rhesus monkey brains, areas that differ visually in microcolumnarity and are associated with different cognitive functions. Our results indicate that microcolumns in these areas are similar in width, spacing, and periodicity, but are stronger (possess a higher neuronal density) in area TE, as compared to areas TL and 46. We modeled the effect of section orientation on microcolumnar spacing and demonstrated that this method provides an adequate estimate of spacing. We also modeled disruption of microcolumnarity by performing simulations that randomly displace neurons and demonstrated that displacements of only one neuronal diameter effectively eliminate microcolumnar organization. These results indicate that our density map method is sensitive enough to detect and quantify subtle differences in microcolumnar organization that may occur in the context of development, aging, and neuropathology, as well as between areas and species. PMID- 15661315 TI - Affective semantic priming in patients with schizophrenia. AB - Affective semantic priming was examined in normal controls and patients with schizophrenia using a lexical decision task with four affective categories of related word pairs: neutral, happy, fearful and sad. Results demonstrated a striking and reliable effect of affective category upon semantic priming. Neutral and happy prime targets yielded significant semantic priming. Fearful and sad pairs showed no or modest semantic priming facilitation. Schizophrenia patients did not differ from normal controls on their priming scores to any of the four affective categories. These results support the notion that the associative mechanisms that bind negative valence words are distinct in nature; they also clarify that patients with schizophrenia do not show increased sensitivity to affect, and more specifically negative affective context during priming paradigms. This study further indicates the importance of replicating novel findings and reporting negative results. PMID- 15661316 TI - Stability of set-shifting and planning abilities in patients with schizophrenia. AB - Patients with schizophrenia have deficits in executive function that involve attentional set-shifting and planning ability. It is unclear, however, whether such deficits are stable during the course of the illness or if they fluctuate in response to medication effects or symptom changes. Patients with a DSM-IV diagnosis of schizophrenia (n=28) and healthy control subjects (n=17) were tested on computerised measures of attentional set-shifting and planning at baseline and 9-month follow-up. The measures used were the Intra/Extradimensional Shift test (ID/ED) and the Stockings of Cambridge test (SoC) from the Cambridge Automated Neuropsychological Testing Battery (CANTAB). On both tests, the patients were poorer than controls at baseline; however, there was no evidence of change over the 9-month follow-up. Additionally, there was little evidence of a relationship between executive test performance and medication or length of illness. This study accords with the presence of executive processing deficits in schizophrenia that are stable across time. PMID- 15661317 TI - Test-retest stability of neuropsychological testing and individual differences in variability in schizophrenia outpatients. AB - The accurate measurement of neurocognitive function requires stable and reliable instruments. These assessments are critically important as correlates, predictors, and outcome measures of psychopathology, neuropathology, and treatment. Particularly, in studies that evaluate the impact of treatments on neurocognitive function, it is essential that their reliability as repeated measures be quantified. In an attempt to measure within- and between-subject variance on a battery of neuropsychological tests commonly used in schizophrenia research, baseline measures were administered twice, approximately 10 weeks apart, to 54 stable outpatients with schizophrenia or schizoaffective disorder. Instruments were judged for their stability, and individuals were assessed for individual differences in test-retest variability. The majority of the instruments administered were highly stable. Group means for several tests showed improvement on retest, which may indicate a practice effect, although many tests did not reach our criteria for "change." Of 962 test scores, 21 (2%) showed significant change. Most subjects were stable, with only 7% of subjects accounting for 38% of significant test-retest changes. Results indicate that the instruments assessed have sufficient reliability and stability to be used as repeated measures in clinical trials and that a small number of patients may have instability of performance as an individual characteristic. PMID- 15661318 TI - High CSF-insulin in violent suicide attempters. AB - Several studies have investigated a connection between diabetes and major depressive disorder (MDD). Whether these associations are mediated by changes in insulin is not known. Insulin seems to play a role in violent behaviour. To further elucidate the role of insulin in MDD and violent, aggressive, or impulsive behaviour, we measured insulin in cerebrospinal fluid (CSF) in 74 patients with a recent suicide attempt. Patients were divided into those with and without MDD, and they were also subgrouped by whether the suicide attempt was considered to be violent or not. It was found that patients with a violent suicide attempt had significantly higher CSF-insulin (5.9+/-1.0 pmol/l) than those with a nonviolent attempt (5.3+/-0.7 pmol/l). In contrast, there were no significant differences in CSF-insulin between patients with MDD and patients without. Our findings support the hypothesis that CSF-insulin is involved in violent behaviour, but not connected to MDD as such. PMID- 15661319 TI - Internalized stigma predicts erosion of morale among psychiatric outpatients. AB - Stigma in society causes harm to people with severe mental illness (SMI) and internalized stigma represents its psychological point of impact. We evaluated the extent of internalized stigma in a sample of outpatients with SMI, using the Internalized Stigma of Mental Illness (ISMI) Scale, developed with consumer input. About a third of the sample reported high levels of internalized stigma. We tested whether internalized stigma predicted increased depressive symptoms and reduced self-esteem at 4-month follow-up, controlling for baseline levels. Depression was predicted by Alienation, Stereotype Endorsement, Social Withdrawal Scales and total ISMI score. Reduced self-esteem was predicted by Alienation. ISMI results were stronger than those for the widely used Devaluation Discrimination Scale. The finding that alienation further reduces morale speaks to the difficulty of pulling oneself out of this type of vicious cycle without assistance. PMID- 15661320 TI - T-helper types 1, 2, and 3 cytokine interactions in symptomatic manic patients. AB - It has been reported that the balance between T-helper type 1 (Th1) cytokines and T-helper type 2 (Th2) cytokines plays a role in psychiatric disorders such as bipolar disorder. The T-helper type 3 (Th3) cytokine, which transforming growth factor beta-1 (TGF-beta1), has been shown to modulate the production of Th1 and Th2 cytokines. However, the role of TGF-beta1 in bipolar disorder has not yet been explored. A total of 70 manic patients with bipolar disorder and 96 normal controls was recruited. The plasma levels of IFN-gamma, IL-4, and TGF-beta1 were studied at the time of admission and 8 weeks after mood stabilizer treatment. The detection rate and plasma concentrations of IFN-gamma and IL-4 and the IFN gamma/TGF-beta1 and IL-4/TGF-beta1 ratios were significantly higher in patients than in controls, while the TGF-beta1 level was significantly lower. The TGF beta1 level increased significantly after treatment and the IFN-gamma/TGF-beta1 and IL-4/TGF-beta1 ratios returned to control values. TGF-beta1 may play a role in the pathophysiology of bipolar disorder through the action of TGF-beta1 in modulating the IL-4/TGF-beta1 ratio. PMID- 15661321 TI - Anxiety sensitivity as a predictor of panic attacks. AB - Anxiety sensitivity (AS) is the fear of physical symptoms of anxiety and related sensations believed to have harmful consequences. AS may play a central role in the nature and etiology of panic disorder (PD) and the genesis of panic attacks. We collected Anxiety Sensitivity Index (ASI) scores from PD patients and controls to determine if AS accurately predicts panic. ASIs were completed prior to panic induction using the modified Read rebreathing test in both hypoxic and hyperoxic conditions. Total scores first-order factors, and individual item ASI scores were correlated with panic presence (Spearman correlation) for each of the hypoxic and hyperoxic rebreathing tests for both study populations. Control subjects' data correlated significantly for items 4, 8, and 11 of the ASI for the hyperoxic (n=9; r(S)=0.63, 0.70, and 0.63, respectively) and items 4 and 8 for the hypoxic rebreathing tests (n = 9; r(S) = 0.63 and 0.70, respectively). Panic patients' data correlated significantly for item 1 of the ASI for hyperoxic tests (n=8; r(S)=0.76) and item 5 for the hypoxic tests (n = 8; r(S) = 0.95). Total ASI scores or first-order factors (physical, social concerns, and mental incapacitation) scores of either study group did not correlate significantly with panic presence. AS may not be a reliable predictor of panicogenic responses to CO2-induced panic in either PD or normal control populations. AS may not be an ultimate causal element in eliciting panic attacks. PMID- 15661322 TI - An examination of the psychometric properties of the Hospital Anxiety and Depression Scale in Chinese patients with acute coronary syndrome. AB - The psychometric properties of the Hospital Anxiety and Depression Scale (HADS) as a screening instrument for patients with acute coronary syndrome (ACS) were investigated in a translated Chinese version of the instrument. A confirmatory factor analysis (CFA) was conducted on the HADS to establish its psychometric properties in 138 ACS patients over two observation points (within 1 week and 6 months post-admission for ACS). Internal and test-retest reliability values for the HADS total and HADS anxiety sub-scales were found to be acceptable. The HADS depression sub-scale lacked acceptable internal reliability. The underlying factor structure of the HADS comprised three distinct factors, though inconsistency between the best three-factor model fit was observed between observation points. The HADS was confirmed to be a useful screening instrument to assess symptoms of psychological distress in ACS patients. Further research is required to determine the most appropriate use of HADS sub-scale structures in clinical populations. PMID- 15661323 TI - Lack of depression effect on platelet activation in patients with heart failure. AB - P-selectin is a marker of platelet activation. Previous studies have reported elevated P-selectin in patients with congestive heart failure (CHF) and depression as separate disorders. We examined if comorbid depression had an effect on platelet activation in CHF patients. Soluble (s)P-selectin was measured in 108 CHF patients; 24 with comorbid depression. There were no significant differences in age, cardiac parameters or (s)P-selectin levels between CHF-only patients and patients with comorbid depression. Our data show no group differences in P-selectin values, which suggests that comorbid depression has no additive effect on platelet activation in CHF patient. PMID- 15661325 TI - 'Alternative' cures for depression--how safe are web sites? AB - Complementary and alternative medicine (CAM) continue to be promoted for various health conditions, recently also via the Internet. This study assessed whether patients suffering from depression might be put at risk through the advice offered by these websites. Some websites assessed offered advice with a potential to harm patients suffering from depression. PMID- 15661324 TI - Relationship between somatization and remission with ECT. AB - Patients treated with electroconvulsive therapy (ECT) were divided into those with less severe depression and those with more severe depression. In the less severely depressed group, high somatic anxiety and hypochondriasis predicted a low likelihood of sustained remission with ECT. In the more severely depressed group, these traits were not predictive of ECT outcome. PMID- 15661326 TI - The development of methods for immunophenotypic and lymphocyte function analyzes for assessment of Southern sea otter (Enhydra lutris nereis) health. AB - The Southern sea otter (Enhydra lutris nereis) is listed as threatened under the Endangered Species Act. The population began a pattern of slow decline in 1995. The decline was attributed to high adult mortality rates with infectious disease being the major cause of death. Multiple pathogens were implicated in these deaths including opportunistic pathogens such as Coccidiodes immitis and Toxoplasma sp. These findings suggested that the immunological health of mature animals in this population might be compromised. The primary goal of this study was to establish techniques for assessing phenotypic and functional baseline data for peripheral blood mononuclear cells (PBMC) in free-ranging sea otters. Standard total and differential white blood cell counts were augmented by emumeration of T and B lymphocyte subsets. Lymphocyte function was determined by both mitogen-induced proliferation and expression of IL-2 receptors. In addition to establishing normal ranges for adult animals, age-related changes were identified in B lymphocyte numbers and cell-surface density of major histocompatability complex class II (MHC II) proteins. The predominant lymphocyte subpopulation in Southern sea otters is the T lymphocyte. Substantial variation among individual animals was observed within the B lymphocyte population both in cell number and density of MHC II expression. Pups had greater numbers of T and B lymphocyte, as well as, greater MHC II expression on B lymphocytes than adults. Mitogen-induced proliferation of peripheral blood mononuclear cells (PBMC) was variable among individual animals with no significant difference in cell response between age class and gender. Concanavalin (ConA) was a more effective mitogen in stimulating proliferation and interleukin (IL)-2 receptor expression than pokeweed. This data can be used to augment routine hematology profiles and aid in the identification of animals with immunologic perturbations. PMID- 15661327 TI - Expression and function of Toll-like receptor 2 in canine blood phagocytes. AB - Toll-like receptors (TLRs) are a family of highly conserved pattern recognition receptors (PRR) of mammals that participate in the activation of innate immune responses against microbial infections. Among these receptors, TLR2 is essential for the recognition of conserved structural components of bacteria, protozoa and fungi. Until now, expression of TLR2 in dogs has not been investigated. In this work we describe a partial sequence of the gene coding for canine TLR2 and show that TLR2 mRNA is constitutively expressed in canine blood PMNs. We also show that stimulation of purified PMNs with lipoteichoic acid (LTA), a ligand of TLR2, leads to the release of proinflammatory chemokine IL-8. Furthermore, TLR2 protein is easily detectable by flow cytometry on the canine peripheral blood granulocyte and monocyte cell surface, and slightly on lymphocytes. These findings suggest that, also in dogs as in humans the initial antibacterial response of PMNs could be elicited through engagement of TLR2. PMID- 15661328 TI - Kinetics of local and systemic isoforms of serum amyloid A in bovine mastitic milk. AB - The aim of the present study was to characterise the serum amyloid A (SAA) response to intramammary inoculation of Escherichia coli and to examine the distribution of hepatically and extrahepatically produced SAA isoforms in plasma and milk from cows with mastitis. Milk and plasma SAA concentrations were determined before and after experimental induction of E. coli mastitis in six dairy cows. The milk SAA response was characterised by low or undetectable levels before inoculation, very rapid and large increases in concentration after inoculation, and rapid decline towards baseline levels after resolution of disease. In plasma from cows with experimentally induced E. coli mastitis, four hepatically derived SAA isoforms with apparent isoelectric point (pI) values of 5.8, 6.2, 6.8 and 7.4 were demonstrated by denaturing isoelectric focusing. In milk three highly alkaline isoforms with apparent pI values above 9.3 appeared 12 h post-inoculation. These isoforms were not present in any of the plasma samples, and it therefore seems likely that they were locally produced, tissue-specific isoforms. At 24-36 h post-inoculation one or more acidic isoforms corresponding to those found in plasma appeared in the milk samples. The isoforms demonstrated in plasma from cows with E. coli mastitis were also present in serum obtained from three cows with clinical Streptococcus uberis mastitis. In conclusion, experimentally induced E. coli mastitis is accompanied by a prominent SAA response. The results of the present study indicate that SAA accumulation in mastitic milk is the result of both local synthesis of SAA and of hepatically derived SAA gaining access to the milk due to increased permeability of the blood milk barrier. PMID- 15661329 TI - Canine distemper virus-induced depletion of uninfected lymphocytes is associated with apoptosis. AB - Canine distemper virus (CDV), a negative stranded RNA morbillivirus, causes a multisystemic disease in dogs, which is associated with a severe immune suppression. The aim of the study was to examine the influence of early CDV infection on leukocyte depletion, lymphopenia and virus-induced cell death in dogs infected with a virulent CDV strain. From 10 infected dogs, peripheral blood leukocytes were harvested periodically, phenotyped and analyzed for CDV antigen content and apoptosis using Annexin V-FITC and propidium iodide labeling. CDV infection induced a severe CD3+ T cell and CD21+ B cell depletion in all animals at 3 days post-infection (d.p.i.). For dogs with severe distemper, developing virus persistence in the lymphoid tissue and central nervous system, this lymphopenia lasted until the end of the experiment. Increased levels of lymphocyte apoptosis were found at 3 d.p.i., and monocyte apoptosis at 6 d.p.i. This was more prominent in the group of animals with severe distemper. At 3 d.p.i. no leukocyte infection was detectable indicating that the early lymphocyte depletion and apoptosis was not a direct consequence of virus infection. Taken together, our results demonstrate that CDV-induced lymphopenia is an early event and that the degree of lymphocyte depletion correlates with the severity of disease and virus persistence in the lymphoid tissue and central nervous system. PMID- 15661330 TI - Evaluation of bovine cutaneous delayed-type hypersensitivity (DTH) to various test antigens and a mitogen using several adjuvants. AB - The Bacillus Calmette Guerin (BCG)-induced/purified protein derivative (PPD) elicited tuberculin skin test is a reliable measure of cell-mediated immune response (CMIR), specifically delayed-type hypersensitivity (DTH); however, its use in livestock may confound diagnosis of Mycobacterium tuberculosis. Therefore, various alternative antigen/adjuvant combinations were evaluated as inducers of DTH that were compared to the BCG/PPD test system with the purpose of finding a skin DTH protocol that does not cross-react with the tuberculin test and allows identification of high and low CMIR responder phenotypes. Specifically, 30 non lactating cows (five/treatment) were sensitized on day 0 with mycobacteria [BCG, M. tuberculosis or Mycobacterium phlei cell wall extract (MCWE)], and ovalbumin (OVA) emulsified in Freund's complete adjuvant (FCA), non-ulcerative Freund's adjuvant (NUFA), complete NUFA or MCWE. On day 21, cows were injected intradermally with various test antigens including PPD tuberculin, phlein, and OVA. Phosphate buffered saline was included as the negative control and the T cell mitogen phytohemagglutinin (PHA) was also administered. Double skin-fold thickness was evaluated before and at 6, 24, and 48 h post-injection. Skin biopsies were taken at 24 and 48 h to assess oedema, necrosis, and inflammatory cell infiltration. BCG/PPD and M. phlei/phlein treatments when given with a Freund's adjuvant induced equivalent DTH with peak reactions at 24-48 h after antigen injection. Cows receiving NUFA had fewer injection site granulomas than FCA or CNUFA treatments. The change in skin thickness response to PHA peaked at 6 h. Only cows receiving mycobacteria in NUFA had skin response to OVA, which peaked 6-24 h post-injection. Only sites tested with PPD or phlein had significantly higher lymphocyte infiltration than control, whereas neutrophils were significantly higher at PHA test sites and eosinophils predominated at the PHA test sites. Macrophages were significantly more numerous at the PPD and/or phlein test sites in treatment groups that received killed mycobacteria in a Freund's adjuvant and/or with BCG, and at the PHA test sites in all treatment groups. It was concluded that the M. phlei/phlein system did induce DTH and was similar to the DTH induced by the BCG/PPD system when MCWE was administered with a Freund's adjuvant. Therefore, this protocol is suitable for detecting high/low CMIR responders in research herds. However, cross-reaction to PPD was evident following induction of DTH using M. phlei. Hence, this protocol does not alleviate the problem of artificial induction of DTH cross-reactivity and would not be suitable for commercial herds where tuberculin testing is required. PMID- 15661331 TI - Epitope specificity of monoclonal antibodies to the N-protein of porcine reproductive and respiratory syndrome virus determined by ELISA with synthetic peptides. AB - In an attempt to develop an alternate to ELISAs using recombinant N-proteins as antigen for the sero-diagnosis of porcine reproductive and respiratory syndrome virus (PRRSV) infections of pigs I have measured the binding of nine anti-N protein mAbs, which had been previously generated by various investigators, to overlapping peptides encompassing amino acids 19-70 of the N-proteins of the North American prototype (VR2332) and the European prototype (Lelystad virus, LV) of PRRSV. I also measured the binding of the mAbs to HerdChek ELISA plates coated with recombinant N-protein. All mAbs bound in an indirect ELISA to some of the peptides whether the mAbs had previously been reported to recognize continuous or discontinuous epitopes, but with different specificity and titer. Three mAbs bound with high titer to different linear epitopes located in amino acid segments 23-33, 31-50 and 43-56 and also with similar high titers to HerdChek plates. mAb SDOW17 bound with high titer to HerdChek plates but poorly to any of the peptides. In contrast, four mAbs bound with broad specificity to peptides containing an epitope(s) in amino acid segment 30-48, but poorly, or not at all, to HerdChek ELISA plates. Thus, this epitope is missing on the antigens of the HerdChek ELISA or is destroyed during immobilization of the antigens on the plate. A mAb to the N-protein of the closely related mouse arterivirus lactate dehydrogenase-elevating virus bound to the same epitope. Abs that bound with broad specificity to an epitope(s) in the 30-50 amino acid segment were also detected by the peptide ELISA in sera of 25 field sera that were sero-positive in the HerdChek ELISA, but also in sera of pigs from two out of three herds tested that were sero-negative by this test. PMID- 15661332 TI - Total and nematode-specific IgE responses in intestinal lymph of genetically resistant and susceptible sheep during infection with Trichostrongylus colubriformis. AB - Total and antigen-specific IgE responses in afferent (AIL) and efferent (EIL) intestinal lymph of sheep with a nematode resistant (R) or susceptible (S) genotype during challenge infection with the intestinal nematode parasite Trichostrongylus colubriformis were examined. Within each sheep line, lambs with a nematode naive or nematode field-primed pre-challenge status were used. Total IgE level in AIL and EIL was dependent on nematode infection and was further influenced by genotype or the immune phenotype (nematode immune mean FEC+/ SDM=77+/-179 or non-immune mean FEC+/-SDM=4016+/-4318) of the animal. During T. colubriformis challenge immune animals had higher levels of total IgE in lymph than non-immune sheep, R line sheep had higher concentrations of total IgE than S line sheep, and field-primed animals had higher total IgE levels than nematode naive animals. Concentrations of total IgE were consistently higher in AIL than EIL or serum and were higher in lymph draining the proximal than the distal jejunum demonstrating that polyclonal IgE in AIL was largely derived from the intestinal mucosa of the anatomical compartment where the nematodes reside. The consistently higher concentration of total IgE in AIL was dependent on phenotype or genotype and in S genotype sheep also on the pre-challenge status. Concentrations of nematode specific IgE were significantly higher in EIL than AIL indicating a preference for the production of IgE reacting with excretory secretory products of the infective T. colubriformis larvae in the regional lymph node. PMID- 15661333 TI - Cloning and expression of interferon-alpha/gamma from a domestic porcine breed and its effect on classical swine fever virus. AB - To further evaluate the clinical impact of recombinant PoIFN-alpha/gamma, PoIFN alpha/gamma genes from a Chinese domestic big-white porcine breed were cloned using PCR, and expressed in a high-level prokaryotic system. The antiviral activities of rPoIFN-alpha/gamma on vesicular stomatitis virus (VSV), porcine reproductive and respiratory syndrome virus (PRRSV), and classical swine fever virus (CSFV) were investigated in different cell lines. The cloned PoIFN-alpha gene encodes a protein of 166 amino acids and has been named PoIFN-alphac. In a comparison of PoIFN-alphac with reported PoIFN-alphaI genes, eight amino acid substitutions at positions 43 (F to L), 78 (N to D), 86 (Y to C), 104 (A to V), 118 (R to L), 128 (T to P), 151 (S to V), and 156 (R to T) were observed, and resulted in no potential N-glycosylation site in the deduced PoIFN-alpha amino acid sequences. In contrast to PoIFN-alphac, one nucleotide substitution was found at position 462 (A to G), hence 0.1% synonymity is specific for the PoIFN gamma gene. Both PoIFN-alphac and PoIFN-gamma genes were inserted into a prokaryotic vector pQE30, and expressed in E. coli M15 (pREP4) or SC11103 (pREP4) with the N-terminal six consecutive histidine residues, respectively. rPoIFN alphac and rPoIFN-gamma proteins were detected by SDS-PAGE and Western blotting analysis at 20.7 and 18.0 kDa, respectively. In addition, the rPoIFN-alphac and rPoIFN-gamma protein were purified using Ni-NTA metal-affinity chromatography, and their anti-VSV, anti-PRRSV, and anti-CSFV activities were surveyed in homologous and heterologous cell lines. The results suggested that rPoIFN-alpha and rPoIFN-gamma could inhibit classical swine fever virus and other important viral pathogens in different cell lines. PMID- 15661335 TI - Adhesion molecule and homing receptor expression on blood and milk polymorphonuclear leukocytes during the periparturient period of dairy cattle. AB - Adhesion molecule and homing receptor expression on blood and milk polymorphonuclear leukocytes (PMN) from periparturient dairy cattle was studied. Both percentages and the mean fluorescence intensity (MFI) of PMN expressing CD11a, CD44, CD62L, and LPAM-1 (alpha4 beta7) were evaluated at seven time points during the twenty-one day period post calving. CD11a and CD62L were expressed on 94-100% of PMN in both blood and milk and there were no significant differences in these percentages at any time point. LPAM-1 was expressed on 3-10% of the PMN in the blood and 13-45% in the milk and the percentage of cells expressing LPAM-1 in milk was significantly (P<0.05) greater than in blood at 0, 4, 10, 14, 18 and 21 days after calving. CD44 was expressed on 11-39% of the PMN in blood and 33 69% in the milk and the percentage of cells expressing CD44 in milk was significantly (P<0.05) greater than in blood at all time points. The MFI of CD11a on milk PMN was consistently higher than that of blood PMN throughout the study period and significantly (P<0.05) higher at days 4, 10 and 18 after calving. PMID- 15661334 TI - Cytokine profiles of peripheral blood and airway CD4 and CD8 T lymphocytes in horses with recurrent airway obstruction. AB - Equine recurrent airway obstruction (RAO) is thought to result from an aberrant immune response to inhaled antigens, modulated by T lymphocytes via the secretion of pro-inflammatory cytokines. However data relating to the phenotypes of the T lymphocytes present in peripheral blood and bronchoalveolar lavage fluid of RAO horses and their cytokine profiles are contradictory. The aim of this study was to further investigate the cytokine (IL-4, IL-5, IL-13 and INF-gamma) mRNA expression profile in peripheral blood lymphocytes and bronchoalveolar lavage lymphocytes from RAO and control horses, before and at 48 h after horses were exposed to hay/straw. In contrast to previous studies, cytokine expression was quantified in populations of CD4 and CD8 T lymphocytes which were purified using magnetic bead antibody cell separation. Hay/straw exposure induced clinical airway obstruction, airway neutrophilia and airway lymphocytosis in RAO horses, and, induced a mild, but significant, airway neutrophilia in controls. However, hay/straw exposure had no significant effect on peripheral blood lymphocyte or bronchoalveolar lavage lymphocyte cytokine expression in either group. In conclusion, RAO was not associated with alterations in lymphocyte cytokine expression that are consistent with Th1 or Th2 responses, but rather with a general down-regulation in expression of the measured cytokines in peripheral blood lymphocytes and bronchoalveolar lavage lymphocytes. PMID- 15661336 TI - Characterisation and quantification of equine interferon gamma. AB - Interferon-gamma (IFN-gamma) is a key cytokine in cell-mediated immunity. To measure IFN-gamma production of equine lymphocytes (eqIFN-gamma), we developed a quantitative ELISA. Monoclonal antibodies (mAb) were produced against bacterially derived eqIFN-gamma. The mAbs recognised recombinant and lymphocyte-derived eqIFN gamma in ELISA, Western blotting, as well as flow cytometric and microscopic analysis. In contrast to bacterially derived material, mammalian and insect cell derived eqIFN-gamma was biologically active but could be neutralised by one of the monoclonal antibodies. Unexpectedly, glycosylation seemed to be required for antiviral activity of eqIFN-gamma. PMID- 15661337 TI - Expression patterns of chicken Toll-like receptor mRNA in tissues, immune cell subsets and cell lines. AB - The Toll-like receptor (TLR) family of cell surface molecules represent a major component of the pattern recognition system, which enables both vertebrates and invertebrates to detect invading microorganisms and mount an anti-microbial response. The TLR repertoire of mouse and man has been intensively studied and in this manuscript we report the identification of ESTs with homology to chTLR5 and chTLR7, and independently confirm the identification of chTLR 1/6/10 and 3 in the EST databases. We have determined the mRNA expression patterns for seven chicken TLRs (chTLR) in a wide range of chicken tissues, isolated immune cell types and cultured cells. Some of the chTLR were expressed in most tissues (chTLR1/6/10, chTLR3, chTLR4 and chTLR5), whereas others exhibited more restricted expression patterns (chTLR2 type 1, type 2 and chTLR7). Similarly distinct patterns of chTLR expression were seen with innate and adaptive immune cell types isolated from peripheral blood or spleen and with cultured cells of somatic or immunological origin. An understanding of the TLR repertoire for different tissues, immune cell subsets and cultured cell types allows more refined interpretation of immune induction in response to chicken pathogens. PMID- 15661338 TI - Chaperones and slow death--a recipe for tumor immunotherapy. AB - In an exciting recent paper Richard Vile and coworkers describe a novel approach to tumor immunotherapy based on the killing of a population of normal cells within an inflammatory environment. This approach builds on recent studies showing that the molecular chaperone Hsp70 can act at multiple stages during tumor antigen presentation to enhance the generation of CD8(+) T lymphocyte mediated immunity and lead to regression of primary and metastatic tumors. PMID- 15661339 TI - Papa's got a brand new tag: advances in identification of proteases and their substrates. AB - Characterization of proteolytic enzymes and their substrates presents a formidable challenge in the context of biological systems. Despite the fact that an estimated 2% of the human genome codes for proteases, only a small fraction of these enzymes have well-characterized functions. Much of the difficulty in understanding protease biology is a direct result of the complexity of regulation, localization and activation exhibited by this class of enzymes. Here, we focus on several recently developed techniques representing crucial advances toward identification of proteases and their natural substrates. PMID- 15661340 TI - The growing importance of fat in regenerative medicine. AB - A recent publication by Michael Longaker and colleagues represents a landmark for the use of adipose tissue as a source of cells for tissue regeneration. The authors investigated the ability of adipose tissue-derived cells (ADCs) to regenerate critical size calvarial (superior portion of the skull) defects in mice by using a novel osteoconducive apatite-coated Poly-lactic-co-glycolic acid (PLGA) scaffold for cell delivery. Direct comparison of this osteogenic ability was performed with bone marrow stromal cells and juvenile calvarial-derived osteoblasts. PMID- 15661341 TI - Protease inhibitor MG132 in cloning: no end to the nightmare. PMID- 15661342 TI - Nitric oxide detoxification--a new era for bacterial globins in biotechnology? AB - For more than a decade, the expression of Vitreoscilla hemoglobin (VHb) has been used to improve the growth and/or productivity of various organisms that are important for the production of valuable metabolites and recombinant proteins by biotechnological processes. Extensive experimental data have shown that VHb enhances the energy status of the cell under oxygen-limited conditions, presumably by improving the supply of intracellular oxygen. Recently, bacterial globin proteins have gained more attention in research because of their ability to detoxify nitric oxide (NO) in vivo. These new results have increased our knowledge, encouraging us to reconsider the role of VHb in vivo. The expression of heterologous globins might improve cellular protection against nitrosative stress under oxygen-limited conditions. PMID- 15661343 TI - Perspectives and vision for strain selection in bioaugmentation. AB - Notwithstanding the phenomenally large and ever-increasing resource of pollutant degrading microbial isolates in laboratories around the globe, inoculum survival remains the 'Achilles' heel' for bioaugmentation of contaminated land. Considerable effort has been invested into inoculum strain selection to facilitate pollutant biodegradation, ranging from the isolation of 'superbugs,' which are microorganisms highly resilient to environmental stresses, harboring catabolically superior pollutant-degrading enzymes, to the other extreme in 'priming', where pollutant degradation is carried out through the addition of soil enriched with an undefined consortium of pollutant-degrading microorganisms. PMID- 15661344 TI - In situ stem cell therapy: novel targets, familiar challenges. AB - Tissue engineering approaches for expanding, differentiating and engrafting embryonic or adult stem cells have significant potential for tissue repair but harnessing endogenous stem cell populations offers numerous advantages over these approaches. There has been rapid basic biological progress in the identification of stem cell niches throughout the body and the molecular factors that regulate their function. These niches represent novel therapeutic targets and efforts to use them involve the familiar challenges of delivering molecular medicines in vivo. Here we review recent progress in the use of genes, proteins and small molecules for in situ stem cell control and manipulation, with a focus on using stem cells of the central nervous system for neuroregeneration. PMID- 15661345 TI - The impact of genomics on vaccine design. AB - After 200 years of practice, vaccinology has gained new perspectives for preventing infectious diseases. Sequencing of complete bacterial genomes led to the development of new large-scale technologies, such as bioinformatics, proteomics and DNA microarrays. By examining genetic content, as well as transcription and expression profiles, a more detailed understanding of bacterial pathogenesis can be reached. Moreover, the whole-genome perspective is expected to provide an instrumental contribution to vaccine development, particularly to target those pathogens for which the traditional approaches have failed so far. In this review, we describe how genomic approaches can be used to identify novel vaccine candidates or create safer live-attenuated vaccines. PMID- 15661347 TI - Homologous recombination and RecA protein: towards a new generation of tools for genome manipulations. AB - Homologous recombination (HR) is one of the central processes of DNA metabolism, combining roles in both cell housekeeping and the evolution of genomes. In eukaryotes, HR underlies meiosis and ensures genome stability. The complete sequencing of numerous bacterial genomes has shown that HR has a substantial role in the evolution of microorganisms, especially pathogens. HR systems from different species and their isolated components are finding an expanding field of applications in modern genetic engineering and bio- and nanotechnologies. Recently, much progress has been made in our understanding of HR mechanisms in eukaryotes and the practical applications of HR systems. PMID- 15661346 TI - Proofreading genotyping assays mediated by high fidelity exo+ DNA polymerases. AB - DNA polymerases with 3'-5' proofreading function mediate high fidelity DNA replication but their application for mutation detection was almost completely neglected before 1998. The obstacle facing the use of exo(+) polymerases for mutation detection could be overcome by primer-3'-termini modification, which has been tested using allele-specific primers with 3' labeling, 3' exonuclease resistance and 3' dehydroxylation modifications. Accordingly, three new types of single nucleotide polymorphism (SNP) assays have been developed to carry out genome-wide genotyping making use of the fidelity advantage of exo(+) polymerases. Such SNP assays might also provide a novel approach for re sequencing and de novo sequencing. These new mutation detection assays are widely adaptable to a variety of platforms, including real-time PCR, multi-well plate and microarray technologies. Application of exo(+) polymerases to genetic analysis could accelerate the pace of personalized medicine. PMID- 15661348 TI - Structural genomics of GPCRs. AB - G protein-coupled receptors (GPCRs) are targets for 60-70% of drugs in development today. Traditionally, the drug discovery process has relied on screening of chemical compounds to identify novel and more-efficient drug molecules. Structure-based drug design, however, provides a targeted approach but has been severely hampered by limited knowledge of high-resolution structures of GPCRs owing to the difficulties encountered in their expression, purification and crystallization. In addition to individual laboratories studying specific GPCRs, structural genomics initiatives have been established as large networks with a wide range of expertise in protein expression, purification and crystallography. Several of these national and international consortia have included GPCRs in their programs. Milligram quantities of GPCRs can now be expressed in several expression systems and purified to high homogeneity. However, success in crystallization still requires major technological improvement. PMID- 15661349 TI - Genetic instability of C. elegans comes naturally. AB - There have been many attempts to measure the genome-wide mutation rate for spontaneous mutations, using measurements of traits in inbred lines in which mutations have accumulated. However, these are likely to miss many small-effect mutations that are important for evolutionary processes. Recently, the genome wide spontaneous mutation rate in inbred lines of Caenorhabditis elegans was estimated, using DNA sequencing. The results imply that the mutation rate is surprisingly high, and that insertion-deletion mutations are unexpectedly common. Phenotypic assays of the same lines detected only a small proportion of mutations that were predicted to have evolutionarily significant fitness effects. PMID- 15661350 TI - Getting to know your neighbours; a new mechanism for cell intercalation. AB - As an embryo gastrulates or makes neural tissue it shortens across the dorso ventral axis and extends dramatically along the perpendicular, antero-posterior axis, resulting in the embryo doubling in length. This process is known as convergent extension and it is so powerful in remodelling tissue that it is used time and again during development. New research in Drosophila melanogaster and other model organisms is shedding fresh light on how it happens. PMID- 15661351 TI - Alternative splicing of conserved exons is frequently species-specific in human and mouse. AB - In this article, we provide evidence that a frequent source of diversity between mammalian transcripts occurs as a consequence of species-specific alternative splicing (AS) of conserved exons. Using a highly predictive computational method, we estimate that >11% of human and mouse cassette alternative exons undergo skipping in one species but constitutively splicing in the other. These species specific AS events are predicted to modify conserved domains in proteins more frequently than other classes of AS events. The results thus provide evidence that species-specific AS of conserved exons constitutes an additional potential source of complexity and species-specific differences between mammals. PMID- 15661352 TI - Is abundant A-to-I RNA editing primate-specific? AB - A-to-I RNA editing is common in all eukaryotes, and is associated with various neurological functions. Recently, A-to-I editing was found to occur frequently in the human transcriptome. In this article, we show that the frequency of A-to-I editing in humans is at least an order of magnitude higher than in the mouse, rat, chicken or fly genomes. The extraordinary frequency of RNA editing in human is explained by the dominance of the primate-specific Alu element in the human transcriptome, which increases the number of double-stranded RNA substrates. PMID- 15661353 TI - Cryptic causation of human disease: reading between the (germ) lines. AB - Most cases of complex human diseases arise sporadically. However, usually there is a significant level of familial aggregation of risk and genetic mapping has identified the responsible gene in a few mendelian cases. Although a disease can be causally genetic, intensified mapping efforts have so far been unable to identify genes that account for more than a small fraction of the familial risk, perhaps because the responsible variation arises by somatic mutation (SM). SM explains the kind of epidemiological pattern seen in cancer, and might have a comparable role in many other diseases. For example, in epilepsy, which has largely defied mapping analysis, the underlying disease pathology, undamped neuronal signaling, is closely connected to gene function. Better technologies to detect and characterize SM are becoming available. However, until it is studied directly, SM will remain a cryptic etiological force, even for diseases that are essentially "genetic". PMID- 15661354 TI - Molecular dating: ape bones agree with chicken entrails. AB - Molecular time estimates, especially those that employed the 310 million years ago (Mya) date of mammal-bird divergence as the calibration point, were criticized in recent publications. In this article, we estimate the divergence time of primates and rodents, primates and artiodactyls and the different great ape species by using two independent calibration-time ranges and maximally conservative error estimates. We observed a variation of approximately +/-15-20% for most of the molecular time estimates in the 10-100 Mya range. The estimated range of the primate-rodent divergence time, 84-121 Mya, includes the date obtained with the 310 million years calibration point (110 Mya). We conclude that molecular time estimates remain useful tools of evolutionary biology, although utmost caution is required when interpreting the results. PMID- 15661355 TI - Dark matter in the genome: evidence of widespread transcription detected by microarray tiling experiments. AB - Microarrays provide the opportunity to measure transcription from regions of the genome without bias towards the location of known genes. This technology thus offers an important source of genomic sequence annotation that is complementary to cDNA sequencing and computational gene-finding methods. Recent "tiling" microarray experiments that assay transcription at regular intervals throughout the genome have shown evidence of large amounts of transcription outside the boundaries of known genes. This transcription is observed in polyadenylated RNA samples and appears to be derived from intergenic regions, from introns of known genes and from sequences antisense to known transcripts. In this article, we discuss different explanations for this phenomenon. PMID- 15661356 TI - Light in retinitis pigmentosa. AB - Retinitis pigmentosa (RP) is one of the most genetically heterogeneous inherited disorders. Twelve genes have now been identified in the autosomal dominant form of the disease, including some recently characterized genes that show unprecedented and fascinating traits in both their function and in their expression profiles. These include many widely expressed genes encoding components of the spliceosome and a guanine nucleotide synthesis gene. Intriguingly, the most recently identified dominant gene does not appear to be expressed in the neuronal retina but is expressed in the capillaries of the choroid. In attempting to understand the effects of mutations in these genes, investigators are forced to re-evaluate their thinking on the molecular mechanisms of genetic blindness and to undertake an increasingly inter disciplinary approach in their analysis of this disease. Recently, this has resulted in significant developments in the elucidation of the molecular pathogenesis of RP. PMID- 15661357 TI - The natural history of group I introns. AB - There are four major classes of introns: self-splicing group I and group II introns, tRNA and/or archaeal introns and spliceosomal introns in nuclear pre mRNA. Group I introns are widely distributed in protists, bacteria and bacteriophages. Group II introns are found in fungal and land plant mitochondria, algal plastids, bacteria and Archaea. Group II and spliceosomal introns share a common splicing pathway and might be related to each other. The tRNA and/or archaeal introns are found in the nuclear tRNA of eukaryotes and in archaeal tRNA, rRNA and mRNA. The mechanisms underlying the self-splicing and mobility of a few model group I introns are well understood. By contrast, the role of these highly distinct processes in the evolution of the 1500 group I introns found thus far in nature (e.g. in algae and fungi) has only recently been clarified. The explosion of new sequence data has facilitated the use of comparative methods to understand group I intron evolution in a broader context and to generate hypotheses about intron insertion, splicing and spread that can be tested experimentally. PMID- 15661358 TI - X chromosome imprinting and inactivation in preimplantation mammalian embryos. AB - Dosage compensation for the mammalian X chromosome involves the silencing of one X chromosome to achieve equal X-linked gene expression between males and females. X chromosome inactivation (XCI) is controlled by a complex set of genetic elements located in a region known as the X chromosome inactivation center, and is regulated by a combination of genomic imprinting, cell lineage-dependent erasure of imprinting, an unidentified mechanism of X chromosome counting, an incompletely understood means of selection of one X chromosome for inactivation and developmentally regulated changes in X chromosome chromatin. A detailed understanding of when and how these components of XCI occur is essential for elucidating the operative mechanisms. A model accounting for early events related to XCI, including observations in uniparental and aneuploid embryos, is presented. PMID- 15661359 TI - Metalloproteinase ADAMTS-1 but not ADAMTS-5 is manifold overexpressed in neurodegenerative disorders as Down syndrome, Alzheimer's and Pick's disease. AB - ADAMTS-1 is a disintegrin and metalloproteinase with thrombospondin 1 (TSP1)-like motifs with ubiquitous though variable expression. Natural substrates of this protease are proteoglycans as aggrecan and versican and null mutant mice propose a role for growth, fertility, organ structure and function. As the gene for this protein is encoded on chromosome 21 and maybe overexpressed due to the gene dosage hypothesis based upon the presence of a third chromosome in trisomy 21, we decided to study expression in Down syndrome (DS) brain and used brains of patients with Alzheimer's (AD) and Pick's disease (PD) as controls. Frontal cortex of controls, DS, AD and PD were homogenized and extracted proteins were used for immunoblotting using antibodies against ADAMTS-1 and ADAMTS-5. ADAMTS-1 immunoreactivity was manifold increased in brain with DS and neurodegeneration, whereas ADAMTS-5 levels were comparable. Overexpression of this metalloproteinase maybe specifically involved in proteoglycan degradation and handling in brain of patients with neurodegenerative disease which in turn may lead to or reflect pathological lesions in DS, AD and PD brain. The manifold overexpression of ADAMTS-1 may be used as marker protein for neurodegeneration. PMID- 15661360 TI - Quantitative analysis of binding parameters of [3H]N-methylscopolamine in central nervous system of muscarinic acetylcholine receptor knockout mice. AB - We have studied binding parameters (Kd, Bmax) of [3H]N-methylscopolamine ([3H]NMS) in various brain regions and spinal cord of wild-type (WT) and muscarinic acetylcholine receptor (mAChR) subtype (M1-M5) knockout (KO) mice. In the M1-M4 KO mice, the number of [3H]NMS binding sites (Bmax) was decreased throughout the central nervous system (CNS) with significant regional differences. Our results collectively suggest that M1 receptor was present in a relatively high density in the cerebral cortex and hippocampus, and the densities of M1 and M4 subtypes were highest in the corpus striatum. M2 receptor appeared to be the major subtype in the thalamus, hypothalamus, midbrain, pons-medulla, cerebellum and spinal cord. These findings may contribute significantly not only to the further understanding of the physiological roles of mAChR subtypes in the central cholinergic functions, but also to the development of selective therapeutic agents targeting specific subtype. PMID- 15661361 TI - Differential functional expression of cation-Cl- cotransporter mRNAs (KCC1, KCC2, and NKCC1) in rat trigeminal nervous system. AB - GABA is the main inhibitory neurotransmitter in the adult brain, which causes Cl- influx into the cell via GABAA receptors. The direction of Cl- inflow is dependent on the Cl- gradient across the membrane. Cation-Cl- cotransporters have been considered to play pivotal roles in controlling intracellular Cl- concentration ([Cl-]i) of neurons; hence, they modulate the GABAergic function. To elucidate how these cotransporters are distributed in the trigeminal nuclei, we investigated the expressions of K+-Cl- cotransporters (KCC1 and KCC2) and Na+ K+-2Cl- cotransporter (NKCC1) mRNAs by using in situ hybridization histochemistry. KCC2 mRNA was expressed in the motor trigeminal nucleus (Mo5), the principal trigeminal nucleus (Pr5), and the spinal trigeminal nucleus (Sp5), but not in the trigeminal ganglion (TG) and the mesencephalic trigeminal nucleus (Me5). On the other hand, KCC1 and NKCC1 mRNAs were expressed in all the trigeminal nuclei. The resting [Cl-]i of Me5 neurons was significantly higher than that of Mo5 neurons. Thus, in primary sensory neurons such as the TG and the Me5, [Cl-]i would be higher than those in the other trigeminal nuclei because of the lack of KCC2 mRNA expression. Since Me5 neurons, but not Mo5 neurons, responded to GABA by depolarization, GABA would have differential physiological functions among trigeminal nuclei and TG. PMID- 15661362 TI - Assessment of differential gene expression in vestibular epithelial cell types using microarray analysis. AB - Current global gene expression techniques allow the evaluation and comparison of the expression of thousands of genes in a single experiment, providing a tremendous amount of information. However, the data generated by these techniques are context-dependent, and minor differences in the individual biological samples, methodologies for RNA acquisition, amplification, hybridization protocol and gene chip preparation, as well as hardware and analysis software, lead to poor correlation between the results. One of the significant difficulties presently faced is the standardization of the protocols for the meaningful comparison of results. In the inner ear, the acquisition of RNA from individual cell populations remains a challenge due to the high density of the different cell types and the paucity of tissue. Consequently, laser capture microdissection was used to selectively collect individual cells and regions of cells from cristae ampullares followed by extraction of total RNA and amplification to amounts sufficient for high throughput analysis. To demonstrate hair cell specific gene expression, myosin VIIA, calmodulin and alpha9 nicotinic acetylcholine receptor subunit mRNAs were amplified using reverse transcription polymerase chain reaction (RT-PCR). To demonstrate supporting cell-specific gene expression, cyclin-dependent kinase inhibitor p27kip1 mRNA was amplified using RT PCR. Subsequent experiments with alpha9 RT-PCR demonstrated phenotypic differences between type I and type II hair cells, with expression only in type II hair cells. Using the laser capture microdissection technique, microarray expression profiling demonstrated 408 genes with more than a five-fold difference in expression between the hair cells and supporting cells, of these 175 were well annotated. There were 97 annotated genes with greater than a five-fold expression difference in the hair cells relative to the supporting cells, and 78 annotated genes with greater than a five-fold expression difference in the supporting cells relative to the hair cells. PMID- 15661363 TI - Differential regulation of nuclear receptors, neuropeptides and peptide hormones in the hypothalamus and pituitary of food restricted rats. AB - Food restriction is associated with a number of endocrine disturbances. We validated the experimental conditions for several house-keeping genes and determined the effects of 12 day 50% food restriction on hypothalamic and pituitary transcription of genes involved in different neuroendocrine systems, using real-time quantitative polymerase chain reaction (PCR). A total of 7 nuclear receptors and 12 neuropeptides and peptide hormones were investigated in the dorsal and ventral hypothalamus and the pituitary gland in rats. In the hypothalamus, food restriction reduced mRNA levels of estrogen receptor alpha (ERalpha), progesterone receptor, glucocorticoid receptor, thyroid hormone receptor alpha and beta, pro-opiomelanocortin (POMC), growth hormone-releasing factor (GHRF), corticotropin-releasing factor (CRF), thyrotropin-releasing factor (TRF), somatostatin, and increased that of neuropeptide Y (NPY). In the pituitary, the treatment reduced growth hormone (GH), luteinizing hormone beta (LHbeta) and thyrotropin beta, but increased ERalpha mRNA levels. The study provides a map of how food restriction affects the regulation of a number of transcripts involved in neuroendocrine control. PMID- 15661364 TI - Purkinje cell expression of the mouse aldolase C gene in transgenic mice is directed by an upstream regulatory element. AB - We have sought to understand the regulation of the expression pattern of aldolase C (Zebrin II) in cerebellar Purkinje cells. Normally, aldolase C is expressed in a series of sagittal stripes of Purkinje cells interrupted by stripes of little or no expression. Genomic aldolase C:LacZ fusion genes with 1.8 kb of sequence 5' to the transcription start site drive CNS expression of LacZ only in astrocytes and cells of the pia mater. If the 5' portion of the transgene is extended to a full 5.0 kb, expression is reliably observed in Purkinje cells, yet none of the astrocyte expression is lost. We broke the additional 3.0 kb into 1.0 kb fragments and tested each for Purkinje cell enhancer activity when appended to the original 1.8 kb construct. We show that the 886 bp region from nucleotide 2796 to -3682 (relative to the start of transcription) contains virtually all of the Purkinje cell enhancer activity. However, neither the full 5.0 kb nor the 886 bp region directed a striped expression pattern, as is seen for the endogenous gene. Taken together, our study localizes a Purkinje cell enhancer to a small 5' region of the aldolase C gene and illustrates that the element(s) responsible for the normal anatomically complex pattern of aldolase C expression are separate from those conferring cell-type specificity. The relationship of these findings to previous work in other laboratories is discussed. PMID- 15661365 TI - Proteomic analysis of phosphotyrosyl proteins in morphine-dependent rat brains. AB - Morphine has been used as a potent analgesic, having a high propensity to induce tolerance and physical dependence following their repeated administration. Although the mechanisms that underlie the development of dependence on morphine remain unclear, previous studies suggested that phosphorylations of diverse types of cellular proteins are crucial determinants of the neuroadaptive mechanisms associated with morphine dependence. Thus, understanding global phosphorylation events induced by chronic morphine administration is essential for understanding the complex signaling mechanisms of morphine dependence. This study characterized the alteration of tyrosine phosphorylation of frontal cortical proteins in morphine-dependent rat brains using a proteomic approach. Dependence was produced by continuous intracerebroventricular (i.c.v.) infusion of morphine (26 nmol/microl/h) for 72 h via osmotic minipumps in rats. Phosphotyrosyl (p-Tyr) protein spots in brain frontal cortical regions were detected by two-dimensional electrophoresis (2-DE) and immunoblotting with anti-p-Tyr-specific antibodies. The protein spots showing significant changes in tyrosine phosphorylation were identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS). Similar patterns of protein expression were detected by 2-DE gels in morphine-dependent and saline-treated control rat brains. However, phosphotyrosine 2-DE images of the frontal cortical proteins from saline treated control and morphine-dependent rat brains were apparently different. The densities of most matched p-Tyr protein spots were increased in morphine dependent rat brains compared with that of control samples. Additional p-Tyr protein spots were detected in 2-DE image of morphine-dependent rat brains. Fifty of p-Tyr protein spots, corresponding to 40 different proteins, were identified from 2-DE gels of morphine-dependent rat brains. The identified proteins include enzymes, cytoskeletal proteins, cell signaling molecules, and other proteins. In conclusion, the first available phosphotyrosine proteomic resources of morphine dependence were established using an animal model. The findings illustrate the potential of proteomics as an effective technique for studying phosphorylation events of morphine dependence in brains. PMID- 15661366 TI - Expression of glia maturation factor beta after cryogenic brain injury. AB - Glia maturation factor beta (GMFB) was identified as a growth and differentiation factor acting on neurons as well as glia. We investigated the expression of GMFB during 56 days after cryogenic brain injury, using immunohistochemistry, reverse transcriptase polymerase chain reaction (RT-PCR), Western blotting and enzyme immunoassay. Immunohistochemical analysis demonstrated that the GFAP-positive astrocytes around the lesion expressed GMFB protein, peaking 14 days after injury. Weak astrocytic expression of GMFB-immunoreactivity was seen in sham operated animal brains. Cryogenic injury (CI) induced GMFB mRNA in the lesioned side after 7 days with a maximum at 14 days. Western blotting revealed the induction of GMFB protein starting 1 day after injury, and continuing until 14 days after injury. In the enzyme immunoassay, GMFB protein concentration peaked 14 days after injury in extracts from the injured side of the brain, whereas in serum it peaked 1 day after injury. These data indicate that the expression of GMFB increased in the astrocytes around the lesioned area after cortical cryogenic brain injury. These findings may provide new insight into GMFB function in pathological conditions following brain injury. PMID- 15661367 TI - Effects of a 14-day period of hindpaw sensory restriction on mRNA and protein levels of NGF and BDNF in the hindpaw primary somatosensory cortex. AB - Neurotrophins have been reported to play an important role in neuronal plasticity and to be regulated by neuronal activity and/or neurotransmitters. Recently, we have shown that hindpaw sensory restriction induces a cortical reorganisation in the hindpaw primary somatosensory cortex, and that acetylcholine plays a significant role in this process. Sensory restriction was obtained by hindlimb suspension for 14 days. In this study, we examined the effects of a long period of hindpaw sensory restriction on the NGF and BDNF mRNA and protein expressions in the hindpaw somatosensory cortex. mRNA and protein levels were assessed by RT PCR and ELISA, respectively. First, we found that NGF and BDNF mRNA relative levels increased after hindpaw sensory restriction. Second, the level of NGF protein increased, whereas that of BDNF remained unchanged. This differential response of NGF and BDNF proteins to sensory restriction suggested different levels of gene regulation, i.e., at pretranslational or posttranslational states. Moreover, inasmuch as our results differ from other models of sensory restriction (dark rearing, whisker removal, etc.), we hypothesized that the regulation of neurotrophin expression is dependent on the type and duration of the sensory restriction. In conclusion, we argue that neuronal plasticity induced by hindpaw sensory restriction requires neurotrophin expression. PMID- 15661368 TI - Activation of ATP-sensitive potassium channels prevents the cleavage of cytosolic mu-calpain and abrogates the elevation of nuclear c-Fos and c-Jun expressions after hypoxic-ischemia in neonatal rat brain. AB - The purpose of this study was to determine whether activation of ATP-sensitive K+ (KATP) channels with diazoxide (DIZ) is able to prevent the cleavage of cytosolic mu-calpain and abrogate the elevation of nuclear c-Fos and c-Jun protein (c-Fos, c-Jun) expressions after hypoxic-ischemia (HI) in brain. The model of hypoxic ischemic brain injury (HIBI) was made in the 7-day-old Sprague-Dawley (SD) rats by left carotid arterial ligation and hypoxia (8% oxygen). DIZ was injected into the left lateral ventricle (5 microl, 1 mg/ml) before or post-hypoxic-ischemia (HI) insults. Western blot and computer image processing were used to detect the integrated density of nuclear c-Fos and c-Jun at 4 h and cleavage of cytosolic mu calpain at 24 h after HI insults from cerebral cortical and hippocampal samples. Compared with HI controls (c-Fos=30.37+/-7.39 from cortical samples, 58.61+/-3.64 from hippocampal samples; c-Jun=52.48+/-14.23 from cortical samples, 35.55+/-4.73 from hippocampal samples), there was a significant down-regulation of c-Fos and c Jun expressions from cortical and hippocampal samples in rats treated with DIZ before (c-Fos=11.10+/-4.64 from cortical samples, 4.82+/-3.38 from hippocampal samples; c-Jun=19.01+/-5.29 from cortical samples, 35.55+/-4.73 from hippocampal samples) or post- (c-Fos=18.81+/-7.93 from cortical samples, 11.33+/-7.05 from hippocampal samples; c-Jun=24.64+/-10.01 from cortical samples, 19.75+/-3.47 from hippocampal samples) HI insults. Furthermore, the ratio of 76 kD/80 kD of mu calpain was down-regulated from cortical and hippocampal samples in rats treated with DIZ before or post-HI insults, demonstrating a significant difference compared with that observed in HI controls. Finally, the increase in DNA fragments caused by the HI injury was decreased or eliminated by the treatment with DIZ. These data suggests that activation of KATP channels by DIZ reduces the degree of mu-calpain proteolysis, and c-Fos and c-Jun expressions in immature brain may contribute to the neuroprotection of K(ATP) channel openers against HIBI. PMID- 15661370 TI - Expression of neural connexins and pannexin1 in the hippocampus and inferior olive: a quantitative approach. AB - Electrical synapses (or neuronal gap junctions) are thought to be essential for the generation of synchronous oscillatory activities in various areas of the brain. In this study, we quantified the steady state mRNA expression levels of two neuronal gap junction proteins, connexin36 (Cx36) and connexin45 (Cx45), as well as of pannexin1, a member of a novel class of communicative junction forming proteins, and of connexin47 (Cx47) which is expressed in oligodendrocytes. The expression levels of these genes were compared in two regions known for oscillatory activity and which are equipped with electrically coupled neurons. Assessment of the levels of mRNA expression in the hippocampus and the nuclear complex of the inferior olive (IO) was achieved by means of laser microdissection (LMM) in combination with real time RT-PCR. Our results demonstrate the differential expression of Cx36, Cx45, pannexin1 and Cx47 in the hippocampus, with pannexin1 showing the highest level of expression followed by Cx36, Cx47, and Cx45. In the IO, pannexin1 showed a comparable expression level as in the hippocampus, but connexin expression levels were increased. Upon direct comparison, the combination of LMM and real time RT-PCR data generated specific, robust and reproducible results consistent with recent data reported about connexin expression in the nervous system. We conclude that the analytical strategy shown here provides a technological solution to overcome the less sensitive and notoriously less specific analysis of connexin expression by in situ hybridization. PMID- 15661369 TI - Platelet-activating factor enhances the expression of nerve growth factor in normal human astrocytes under hypoxia. AB - Nerve growth factor (NGF) is required for the survival of neurons. We have addressed the effect of platelet-activating factor (PAF), one of the mediators of ischemic injury of the brain, on NGF expression in astrocytes. Normal human astrocytes in culture were stimulated with PAF, and levels of NGF mRNA and protein were analyzed by reverse transcription-polymerase chain reaction (RT PCR), real-time quantitative PCR and enzyme-linked immunosorbent assay (ELISA). PAF increased the expressions of NGF mRNA and protein in astrocytes in time- and concentration-dependent manners. After 48-h stimulation, 10 nmol/L PAF increased the levels of NGF protein in astrocyte-conditioned medium by 1.4-fold. The PAF induced stimulation of NGF expression was further enhanced (2.1-fold of the control) in the cells under hypoxic culture condition. BN52021 (Ginkgolide B), an antagonist for PAF binding sites, suppressed the effect of PAF. We conclude that PAF enhances NGF gene expression in human astrocytes, and the PAF-induced increase in the expression of NGF under hypoxia may benefit the protection of the nervous tissue by promoting neuronal survival. PMID- 15661371 TI - Dendritic BC1 RNA: modulation by kindling-induced afterdischarges. AB - Local protein synthesis in dendrites is thought to provide a mechanism for long lasting modifications of synapses in response to physiological activity and behavioral experience. New synthesis of dendritic proteins may be triggered by various paradigms, including induction of epileptiform activity. Prerequisite for such modulated synthesis is a mechanism that limits translation of synaptodendritic mRNAs to times of demand. Recently identified as a translational repressor that is localized to dendrites, small untranslated BC1 RNA has been implicated in the regulation of postsynaptic protein synthesis. Here we show that translational repressor BC1 RNA is itself undergoing modulation as a result of neuronal stimulation. Induction of hippocampal epileptiform activity resulted in a significant decrease of BC1 RNA in the CA3 region over several hours after excitation. The observed decrease was cell-wide, thus indicating reduced expression rather than intracellular redistribution. We suggest that a downregulation of the translational repressor BC1 RNA serves to modulate postsynaptic protein complements in response to the induction of epileptiform activity. Such increased protein synthesis in dendrites may be required for the consolidation of enduring epileptogenic mechanisms. PMID- 15661372 TI - The differential expression patterns of messenger RNAs encoding Nogo-A and Nogo receptor in the rat central nervous system. AB - Nogo-A and Nogo-receptor have been considered to play pivotal roles in controlling axonal regeneration and neuronal plasticity. We investigated the total distribution of Nogo-A and Nogo-receptor mRNAs in the adult rat central nervous system using in situ hybridization histochemistry. Nogo-A is abundantly expressed in both neurons and oligodendrocytes throughout the central nervous system. Interestingly, we could not find any neuron which lacks Nogo-A mRNA expression, indicating that Nogo-A mRNA is universally expressed in all neurons. In contrast, Nogo-R mRNA expression was very restricted. Nogo-R mRNA was expressed in the olfactory bulb, hippocampus, tentia tecta, some amygdala nuclei, cerebral cortex, some thalamic nuclei, medial habenular, whereas we could not detect it in the other regions. Interestingly, we did not detect Nogo-R mRNA in monoaminergic neurons, which are known to have high regenerative capacity, in the substantia nigra, ventral tegmental area, locus caeruleus, and raphe nuclei. In addition, although neurons in the reticular thalamus and cerebellar nuclei are also known to show high capacity for regeneration, Nogo-R mRNA was not detected there. These data indicate that Nogo-A and Nogo-R mRNAs were differentially expressed in the central nervous system, and suggest that the lack of Nogo-R expression in a given neuron might be necessary to keep its high regenerative capacity. PMID- 15661373 TI - 3,4-Methylenedioxymethamphetamine increases neuropeptide messenger RNA expression in rat striatum. AB - The amphetamine analog 3,4-methylenedioxymethamphetamine (MDMA) is also known as the recreational drug of abuse, Ecstasy. Several neuropeptides are found in striatal neurons postsynaptic to dopamine and serotonin nerve terminals, and changes in neuropeptide neurotransmission may be important for behavioral effects of 3,4-methylenedioxymethamphetamine. This study used in situ hybridization to characterize the effects of 3,4-methylenedioxymethamphetamine on four neuropeptide mRNAs: preprodynorphin, preprotachykinin, neurotensin/neuromedin N, and preproenkephalin. Male, Sprague-Dawley rats received a single administration of 10 mg/kg 3,4-methylenedioxymethamphetamine and were sacrificed 30 min or 3 h later. Three hours after administration, 3,4-methylenedioxymethamphetamine increased preprodynorphin, preprotachykinin, and neurotensin/neuromedin N mRNAs. These increases were most prominent in ventral and medial aspects of the rostral middle striatum, and then became more dorsally restricted in the caudal striatum. At the 30-minute time point, MDMA significantly decreased the signal for preproenkephalin mRNA in a general manner but did not affect the signal for the other neuropeptide precursors. These data suggest that 3,4 methylenedioxymethamphetamine has a generalized, transient, inhibitory effect on striatopallidal neuron gene expression, and then preferentially influences striatonigral neuropeptide systems at the later time point in a regionally selective manner. PMID- 15661374 TI - Analyses of murine postsynaptic density-95 identify novel isoforms and potential translational control elements. AB - Postsynaptic density-95 (PSD-95) is an evolutionarily conserved synaptic adaptor protein that is known to bind many proteins including the NMDA receptor. This observation has implicated it in many NMDA receptor-dependent processes including spatial learning and synaptic plasticity. We have cloned and characterised the murine PSD-95 gene. In addition, we have identified two previously uncharacterised splice variants of the major murine PSD-95 transcript (PSD 95alpha): PSD-95alpha-2b results from an extension of exon 2 and PSD-95alpha Delta18 from the temporal exclusion of exon 18. The presence of PSD-95alpha-2b sequences in other PSD-95 family members implicates this peptide stretch as functionally significant. Another potential transcript (PSD-95gamma) was also identified based on examination of EST databases. Immunoprecipitation assays demonstrate that proteins corresponding in size to PSD-95alpha-Delta18 and PSD 95gamma interact with the NMDA receptor, suggesting an important biological role for these isoforms. Finally, we have performed bioinformatics analyses of the PSD 95 mRNA untranslated regions, identifying multiple translational control elements that suggest protein production could be regulated post-transcriptionally. The variety of mRNA isoforms and regulatory elements identified provides for a high degree of diversity in the structure and function of PSD-95 proteins. PMID- 15661375 TI - CAA insertion polymorphism in the 3'UTR of Nogo gene on 2p14 is not associated with schizophrenia. AB - The Nogo gene was putatively implicated in schizophrenia based on gene expression and genetic association data. In this study, we attempt to replicate the possible association of the CAA insertion and a nearby TATC deletion with schizophrenia in 204 complete and incomplete triads and in a sample of 462 unrelated cases and 153 controls, all of Caucasian origin. Our genotyping results indicated that neither the trinucleotide insertion polymorphism (CAAins; 43.4% vs. 41.8%, p>0.5) nor the polymorphism-TATC deletion (TATCdel; 49.8% vs. 49.3%, p>0.1) allele frequency is significantly different in patients compared to controls. The homozygous CAAins frequency is not significantly different between patients and controls either (18.0% vs. 15.0%, chi2=0.985, p>0.1). Furthermore, neither CAAins/TATCdel individually, nor the haplotype carrying both CAAins and TATCdel is preferentially transmitted to affected offspring. PMID- 15661376 TI - Multiple 5'-splice variants of the rat glutamate transporter-1. AB - In most brain areas, uptake of extracellular glutamate predominantly occurs through the glutamate transporter subtype, glutamate transporter-1 (GLT-1), which is enriched in astroglia. Here, we report the identification of five splice variants of the 5'-leader sequence of rat GLT-1 which contain varying numbers of upstream open reading frames and encode putative GLT-1 proteins with two distinct N-terminal modifications. We further demonstrate that the identified rat 5'-GLT-1 splice variants are expressed in a brain region-specific manner. Our findings point to potential influences of RNA splicing on glial glutamate transport in the intact and injured rat brain. PMID- 15661377 TI - Rgs4 mRNA expression is decreased in the brain of Fmr1 knockout mouse. AB - In the Fmr1 knockout mice, a model for fragile X mental retardation syndrome, the levels of regulator of G-protein signaling (Rgs) 4 but not Rgs2 mRNA were considerably reduced (65% from control) in the cerebral cortex and hippocampal CA1 region. The expression of Rgs4 was normal in animals lacking a related protein, FXR2P, indicating that the decrease in Rgs4 expression was specific for the absence of FMRP, and suggests a role for FMRP in G-protein signaling. PMID- 15661378 TI - Reconstituted, refrigerated and stored tissue culture rabies vaccines retain immunogenicity for longer than 1 week. PMID- 15661379 TI - Adverse reactions to human rabies immune globulin manufactured by the Thai Red Cross Society. AB - Rabies Immunoglobulins (RIG) are an essential part of optimal management of rabies exposures [World Health Organization (WHO)]. They provide protection against the virus during the critical initial 7 days before adequate endogenous vaccine induced antibodies are formed. Even though Human Rabies Immune Globulin (HRIG) is the ideal product for post-exposure rabies treatment it, like most biological, can cause very rare adverse reactions. A retrospective review of a series of 8737 patients, who received HRIG at one institution, revealed that only 15 (0.183 %) reported transient mild adverse reactions. PMID- 15661380 TI - Schistosoma mansoni infection reduces the protective efficacy of BCG vaccination against virulent Mycobacterium tuberculosis. AB - We hypothesized that the ability of BCG vaccination to protect against Mycobacterium tuberculosis is less in hosts exposed to chronic helminthes infection compared to unexposed individuals. To test this hypothesis we evaluated the efficacy of BCG vaccination in protecting against M. tuberculosis challenge in Schistosoma mansoni pre-infected mice by analyzing their ability to limit the replication of TB bacilli in the lung and liver and the histology of lung sections. The results show that BCG vaccinated mice with prior S. mansoni infection show significantly higher number of colony forming units of TB bacilli as well as significant reduction in air exchange area in the lung compared to controls. In addition, spleen cells from S. mansoni infected mice were found to produce significantly less IFN-gamma and nitric oxide when stimulated in vitro with PPD and several fold higher soluble egg antigen (SEA) and Concanavalin A induced IL-4 and IL-5 secretion. Taken together, our data show that S. mansoni infection reduces the protective efficacy of BCG vaccination against M. tuberculosis possibly by attenuation of protective immune responses to mycobacterial antigens and/or by polarizing the general immune responses to the Th2 profile. PMID- 15661381 TI - Intragastric administration of Lactobacillus casei expressing transmissible gastroentritis coronavirus spike glycoprotein induced specific antibody production. AB - Lactobacillus casei strain Shirota was selected as a bacterial carrier for the development of live mucosal vaccines against coronavirus. A 75 kDa fragment of transmissible gastroenteritis coronavirus (TGEV) spike glycoprotein S was used as the model coronavirus antigen. The S glycoprotein was cloned into a Lactobacillus/E. coli shuttle vector (pLP500) where expression and secretion of the glycoprotein S from the recombinant lactobacilli was detected via immunoblotting. Oral immunization of BALB/c mice with recombinant LcS that constitutively expresses the 75 kDa fragment of the glycoprotein S, induced both local mucosal and systemic immune responses against TGEV. Maximum titers of IgG (8.38+/-0.19 ng/ml of serum) and IgA (64.82+/-2.9 ng/ml of intestinal water) were attained 32 days post oral inturbation. The induced antibodies demonstrated neutralizing effects on TGEV infection. PMID- 15661382 TI - Immunisation with virion-loaded plasmacytoid or myeloid dendritic cells induces primary Th-1 immune responses. AB - Dendritic cells (DCs) induce different types of immune responses depending on their lineage and activation signals. When exposed to inactivated pseudorabiesvirus (iPRV), plasmacytoid but not myeloid DCs released IFN-alpha and IL-12. Remarkably, both iPRV-pulsed DC types were able to induce primary IFN gamma producing T cells and IgG isotype switching in vivo. In contrast, tetanus toxoid pulsed DCs did not induce detectable primary immune responses. The efficacy of antiviral T and B cell priming proved dependent on the recipient's genotype. We conclude that either plasmacytoid or myeloid DCs pulsed with inactivated virus suffice to induce primary Th1-polarised immune responses. PMID- 15661383 TI - Correlates of HIV vaccine trial participation: an Indian perspective. AB - Successful conduct of HIV vaccine trials in a population of great cultural diversity like India could be a challenge. Concerns, knowledge gaps and willingness to participate in future HIV vaccine trials were studied among 349 patients attending three sexually transmitted infections clinics and one reproductive tract infections clinic. Overall willingness to volunteer for HIV vaccine trials was 48%. Women and men at risk of HIV infection were willing to participate in the HIV vaccine trials. Factors associated with increased willingness to participate in these trials were awareness of current HIV vaccine efforts, realization of importance of vaccine for self, concern about adverse events and altruism. PMID- 15661384 TI - Aluminium assay and evaluation of the local reaction at several time points after intramuscular administration of aluminium containing vaccines in the Cynomolgus monkey. AB - Aluminium hydroxide and aluminium phosphate have been widely used as vaccine adjuvants with a good safety record for several decades. The recent observation in human deltoid muscle of macrophage aggregates containing aluminium hydroxide spicules and termed Macrophagic Myofasciitis (MMF) has encouraged research on aluminium salts. This study was conducted in order to further investigate the clearance of aluminium at the vaccine injection site and the features of induced histopathological lesions. Two groups of 12 monkeys were immunised in the quadriceps muscle with Diphtheria-Tetanus vaccines, which were adjuvanted with either aluminium hydroxide or aluminium phosphate. Three, six or twelve months after vaccination, four monkeys from each group were sacrificed and histopathological examination and aluminium assays were performed on quadriceps muscle sections. Histopathological lesions, similar to the MMF described in humans, were observed and were still present 3 months after aluminium phosphate and 12 months after aluminium hydroxide adjuvanted vaccine administration. An increase in aluminium concentration, more marked in the area of the lesions, was also observed at the 3- and 6-month time points. These findings were localised at the injection site and no similar changes were observed in the distal or proximal muscle fragments. We conclude from this study that aluminium adjuvanted vaccines administered by the intramuscular route trigger histopathological changes restricted to the area around the injection site which persist for several months but are not associated with abnormal clinical signs. PMID- 15661385 TI - Validation of serotyping of Streptococcus pneumoniae in Europe. AB - INTRODUCTION: Serotyping of pneumococci has become increasingly important as new pneumococcal vaccines are introduced and place emphasis on knowledge of national serotype-distributions and their development over time. AIM: The aim of this study was to evaluate the quality of serotyping of pneumococci in Europe, and to focus on possible problems with methods, procedures, etc. that may lead to wrong serotypings. METHODS: This study was part of a larger EU-project. Eleven reference laboratories in Europe participated in the validation of pneumococcal serotyping. The Streptococcus Unit at SSI functioned as the gold standard for use of the Neufeld test. Each laboratory was asked to type 70 blinded pneumococcal strains by use of their normal serotyping procedure and to answer to a questionnaire regarding their experience and serotyping procedure for pneumococci. The 70 strains were chosen to represent all the 23 pneumococcal types included in the 23-valent pneumococcal vaccine plus a number of other less common types. RESULTS: A total of 735 serotypings was performed. Five laboratories performed complete serotyping whereas the remaining six laboratories performed partial or variable serotyping into groups or types that did not belong to groups. In general, a high degree of consensus appeared between the 11 European reference laboratories. Of 735 serotypings, 39 erroneous serotypings were made (5% of all). Most serotyping errors included a wrong serotype within the correct serogroup, where especially types 9N, 18C and 19F were mistyped. Furthermore, misidentification of noncapsular pneumococci like S. mitis and S. oralis was also a frequent error. For 22 strains (30%) of pneumococci, serotyping mistakes were made. The erroneous serotypings were neither correlated to the use of other methods than the Neufeld test, nor to the serotyping routine of the laboratories. A number of errors may be due to a serotyping result based on a negative reaction with a specific factor serum, instead of a positive reaction with another factor serum. This may be chosen in order to simplify the serotyping procedure. Thus, all the necessary factor sera must be used in order to assure correct serotyping. CONCLUSIONS: Overall, the quality of serotyping of pneumococci was high, and a high degree of consensus was found between the eleven laboratories. It is important to use all the necessary factor sera for serotyping, to perform all the necessary tests and to base a serotyping result always on one or more positive reactions and not on a negative reaction alone. More focus on serotyping of serotypes within groups seems to be warranted. PMID- 15661386 TI - A novel intranasal Protollin-based measles vaccine induces mucosal and systemic neutralizing antibody responses and cell-mediated immunity in mice. AB - Protollin-MV is a vaccine produced by mixing split measles virus (MV) antigen with the novel adjuvant Protollin (Neisseria meningitidis outer membrane proteins non-covalently complexed with Shigella flexneri 2a lipopolysaccharide). Intranasal immunization of mice with two or three doses of Protollin-MV induces both serum IgG and mucosal IgA with strong neutralizing activity. There is a dose dependent shift towards lower IgG1:IgG2a ratios and MV-specific IFNgamma production in splenocytes. Intranasal Protollin-MV can therefore induce systemic and mucosal neutralizing antibody responses as well as elicit a balanced TH1/TH2 type response. PMID- 15661387 TI - Induction of gp120-specific protective immune responses by genetic vaccination with linear polyethylenimine-plasmid complex. AB - The induction of IFN-gamma-secreting CD8+ T cells and neutralizing antibodies to HIV-1 are both key requirements for prevention of viral transmission and clearance of pathogenic HIV. Although DNA vaccination has been shown to induce both humoral and cellular immune responses against HIV antigens, the magnitude of the immune responses has always been disappointing. In this report, we analyze the ability of polyethylenimine (PEI)-DNA complex expressing an HIV-glycoprotein 120 (gp120) antigen (PEI-pgp120) to induce systemic CD8+ T cell and humoral responses to the gp120 antigen. The administration of PEI-plasmid complex resulted in rapid elevation of serum levels of IL-12 and IFN-gamma. Furthermore, a single administration of PEI-pgp120 complex elicits a number of gp120-specific CD8+ T cells 20 times higher than that elicited by three intramuscular injections of naked DNA. Interestingly, we found that systemic vaccination with PEI-pgp120 induced protective immune responses against both systemic and mucosal challenges with a recombinant vaccinia virus expressing a gp120 antigen. The data also demonstrated that the depletion of macrophages with liposome-encapsulated clodronate completely abolished gp120-specific cellular response. Overall, our results showed that a single administration of PEI-pgp120 complexes, eliciting strong immune responses, is an effective vaccination approach to generate protection against systemic and mucosal viral infections. PMID- 15661388 TI - Immunotherapy with fragmented Mycobacterium tuberculosis cells increases the effectiveness of chemotherapy against a chronical infection in a murine model of tuberculosis. AB - Reduction of colony forming units by rifampicin-isoniazid therapy given 9-17 weeks post-infection was made more pronounced by immunotherapy with a vaccine made of fragmented Mycobacterium tuberculosis cells detoxified and liposomed (RUTI), given on weeks 17, 19 and 21 post-infection, in the murine model of tuberculosis in C57BL/6 and DBA/2 inbred strains. RUTI triggered a Th1/Th2 response, as demonstrated by the production of IgG1, IgG2a and IgG3 antibodies against a wide range of peptides. The histological analysis did not show neither eosinophilia nor necrosis, and granulomatous infiltration was only slightly increased in C57BL/6 mice when RUTI was administered intranasally. PMID- 15661389 TI - Tat-neutralizing versus Tat-protecting antibodies in rhesus macaques vaccinated with Tat peptides. AB - The human immunodeficiency virus (HIV) regulatory protein Tat represents an attractive target for developing vaccine strategies. Both humoral and cellular responses against Tat might reduce disease progression by interfering with the deleterious functions of extracellularly secreted protein and by reducing viral replication. We have immunized Rhesus macaques intramuscularly and intranasally with a cocktail of three Tat peptides encompassing residues 1-20, 1-61 and 44-61 administrated in the presence of Montanide ISA 720 as adjuvant. The monkeys were challenged by the intrarectal route with 10 MID50 of SHIV BX08. All immunized macaques but one gave a good cross-reactive antibody response to Tat but the proliferative response and levels of IL-2, IFN-gamma and TNF-alpha secretion of peripheral blood mononuclear cells (PBMCs) recalled ex vivo with active Tat protein were weak. After viral challenge one peptide-vaccinated macaque only remained free of virus. The presence in the serum of vaccinated animals of neutralizing antibodies able to inhibit Tat transactivation activity or Tat induced apoptosis was not correlated to protection. PMID- 15661390 TI - The FAI protein of group C streptococci targets B-cells and exhibits adjuvant activity. AB - We have demonstrated that the fibrinogen-albumin-IgG receptor of group C streptococci (FAI) targets B-cells in vivo. We exploited the targeting properties of FAI to improve the immune responses stimulated by soluble antigens administered by the mucosal route. Enhanced systemic and mucosal immune responses were observed in mice after intranasal immunization when ovalbumin was fused to FAI or truncated derivatives. The FAI fragment encompassing the IgG- and fibrinogen-binding regions was the minimal domain exhibiting optimal carrier/adjuvant properties. The obtained results suggest that the FAI protein represents a useful tool to improve the immunogenicity of vaccine antigens. PMID- 15661391 TI - Amount and avidity of serum antibodies against native glycoproteins and denatured virus after repeated influenza whole-virus vaccination. AB - There is still uncertainty on the correlates of protection by influenza vaccine. To determine the relationship between hemagglutination-inhibition (HI) titer and the specificity and avidity of serum antibodies, we analyzed serum from a longitudinal trial (1983-1987) of influenza vaccine efficacy [Keitel WA, Cate TR, Couch RB, Huggins LL, Hess KR. Efficacy of repeated annual immunization with inactivated influenza virus vaccines over a five year period. Vaccine 1997;15(10):1114-22 ]. We captured native virus particles with fetuin and separately measured relative antibody levels and avidities of antibodies against native glycoproteins and antibodies against denatured viral proteins. Most subjects had pre-existing antibodies against A/Victoria/75 and, although 70% had >two-fold increased antibodies against A/Philippines/82 after vaccination, only 30% showed increased antibodies to A/Victoria/75 indicating no dominance of original antigenic sin. There was variation in the levels of antibodies to unfolded antigens compared to native, but antibodies against denatured proteins never exceeded those against native virus. In some cases, the avidity increased without a significant increase in antibody concentration, which might explain why some vaccinees with low HI titer demonstrate adequate protection. We found that the negative correlation between pre-vaccination HI titer and the increase after vaccination is also seen when antibodies are measured directly, but that there is little relationship between HI titer and antibodies against native glycoproteins, either in amount or avidity. Our assay, which has also been adapted for recent influenza viruses that do not bind to fetuin, may be useful for vaccine evaluation. PMID- 15661392 TI - Kinetics and symmetry of divisions of hematopoietic stem cells. AB - To fulfill the dual abilities to self-renew and to differentiate into cells of multiple lineages, stem cells must undergo, at some stage, asymmetric divisions to generate cells to sustain the stem cell pool as well as the various progeny cells of the distinct lineages. A central question in developmental biology is how a single cell can divide to produce two progeny cells that adopt different fates. Different daughter cells can theoretically arise by uneven distribution of determinants upon cell division, i.e., due to intrinsic factors, or become different upon subsequent exposure to environmental signals, i.e., due to extrinsic factors. Recent advances in the understanding of stem cell biology in Drosophila and murine models have served as a model for hematopoietic stem cell (HSC) development. Provided with advances in molecular and cellular biology, we have gained insight into the mechanisms governing self-renewing asymmetric divisions of primitive HSC. Direct contact with cellular determinants in the niche has been shown to play an essential role in the balance between self renewing asymmetric division versus differentiation. Identification of the molecular interactions between stem cells and their niche will lead to an understanding of the mechanisms controlling the long-term destiny of stem cells. Ultimately, molecular signals triggered by adhesion and junction complexes are probably responsible for the specific adoption of differentiation pathways. PMID- 15661393 TI - Perspective: fundamental and clinical concepts on stem cell homing and engraftment: a journey to niches and beyond. AB - In many ways, the homing of hematopoietic stem cells to bone marrow and other tissues defines these cells and their immediate and long-term fates Once homed, an inevitable series of proliferative and differentiative events presumptively follows. These comments, of course, hold for both homing to marrow, or alternatively, to other nonmarrow tissues. In this review, we will specifically focus on homing and engraftment to bone marrow because this is the best-studied and clinically applicable system. PMID- 15661394 TI - Evidence of similar effects of short-term culture on the initial repopulating activity of mobilized peripheral blood transplants assessed in NOD/SCID beta2microglobulin(null) mice and in autografted patients. AB - OBJECTIVE: Human mobilized peripheral blood (mPB) is known to contain high numbers of cells with rapid but short-term repopulating activity in NOD/SCID beta2microglobulin(-/-) mice. Here we assessed the effect of short-term culture on these cells and compared the levels of retained activity with the pace of hematologic recovery in myeloablated patients transplanted with similarly cultured autografts of the same cells. PATIENTS AND METHODS: In a phase 1 clinical study, mPB cells were collected from 6 advanced cancer patients. CD34(+) cells were then harvested, cultured for 3 days in the presence of early-acting growth factors, and transplanted, and posttransplant recovery of blood cell parameters monitored. Assays for primitive hematopoietic activity using both in vivo (in NOD/SCID-beta2microglobulin(-/-) mice) and in vitro (CFC and LTC-IC) endpoints were also performed on the cells pre- and posttransplant. RESULTS: All patients showed event-free, timely leukocyte recoveries but slightly delayed platelet recoveries in some cases. During the 3-day period of culture, the CFCs doubled but the LTC-IC activity decreased (twofold), as did the short-term repopulating activity in NOD/SCID-beta2microglobulin(-/-) mice. CONCLUSION: Patients can be transplanted with 3-day cultured autografts with minimal effects on hematologic recovery. This is associated with a variable but, on average, modest loss of short-term repopulating activity detectable in NOD/SCID beta2microglobulin(-/-) mice. PMID- 15661395 TI - Abnormal erythroid differentiation in neonatal bcl-6-deficient mice. AB - OBJECTIVE: The bcl-6 proto-oncogene is ubiquitously expressed in various tissues. Since we found out the smaller number of TER119(+) cells in the spleen of neonatal bcl-6-deficient (bcl-6(-/-)) mice compared with that of control (bcl 6(+/+)) littermates, we studied functions of bcl-6 in differentiation of erythroid lineage cells. MATERIALS AND METHODS: Erythroblasts in the definitive erythropoiesis were separated into four subsets using anti-TER119 and anti-CD71 mAbs. The cell number and property of these four subsets in spleens of neonatal bcl-6(+/+) and bcl-6(-/-) mice were examined using a flow cytometry. RESULTS: bcl 6 mRNA expression was detected in the TER119(high)CD71(high) subset, which is morphologically equivalent to basophilic erythroblasts, by reverse-transcribed polymerase chain reaction. High percentages of cells in the TER119(low)CD71(high) and TER119(high)CD71(high) subsets were in the cell cycle. The cell number of the TER119(high)CD71(high) subset in the spleen and the percentage of reticulocytes in the peripheral blood of neonatal bcl-6(-/-) mice were significantly lower than those of neonatal bcl-6(+/+) mice. However, the percentage of apoptotic cells and that of cells in the cell cycle in the TER119(high)CD71(high) subset of bcl-6(-/ ) mice were similar to those of bcl-6(+/+) mice. CONCLUSION: bcl-6 detected in the TER119(high)CD71(high) subset of erythroblasts in the spleen of neonatal mice may be required to retain the erythroblasts in the cell proliferation stage. PMID- 15661396 TI - An in vivo assay for retrovirally transduced human peripheral T lymphocytes using nonobese diabetic/severe combined immunodeficiency mice. AB - OBJECTIVE: Availability of a mouse model to analyze human peripheral lymphocytes genetically modified with retroviral vectors would be useful in T-cell-directed gene transfer studies. To address this issue, we assessed the ability of nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice to maintain such cells in their peripheral blood. MATERIALS AND METHODS: Human peripheral lymphocytes stimulated with recombinant human interleukin-2 (rhIL-2) and anti-CD3 and CD28 antibodies were transduced with the enhanced green fluorescent protein (EGFP) gene using the retroviral vector GCsap(MSCV) and then transplanted into NOD/SCID mice at 1 x 10(8) cells per mouse. RESULTS: Transplanted human peripheral lymphocytes survived and expressed EGFP in the mice over the 6- to 8-week posttransplant period without any signs of graft-vs-host disease. Of importance was that these cells remained at the G(0)/G(1) stage and again proliferated in response to cytokines when cultured in vitro. Interestingly, the mice in which the transduced T lymphocytes remained at the resting stage clearly elucidated the superiority of the murine stem cell virus (MSCV) LTR to maintain the transgene expression by nonproliferating T lymphocytes over the Moloney murine leukemia virus (MoMLV)- and myeloproliferative sarcoma virus (MPSV)-derived LTRs, which was obscure in in vitro culture where the transduced lymphocytes was being stimulated with rhIL-2. CONCLUSIONS: The mouse model and GCsap(MSCV) vector described herein comprise a simple and reliable in vivo assay system for studies of gene and cell therapies employing human peripheral lymphocytes. PMID- 15661397 TI - Human neutrophil collagenase expression is C/EBP-dependent during myeloid development. AB - OBJECTIVE: Human neutrophil collagenase (HNC) is one of several secondary granule proteins (SGP) expressed late in the myeloid maturation pathway. SGPs are encoded by unlinked and functionally diverse genes that are hypothesized to be coordinately regulated at the transcriptional level and demonstrate uniform dysregulation in leukemic cells. In support of the hypothesis that tissue and stage-specific expression of SGP genes is regulated by shared factor(s), we sought to identify factors responsible for positive regulation of the SGP genes. METHODS: Using 5' deletion analysis, we identified a minimal HNC promoter located within the first 193 bp upstream of the transcription start site. Three CCAAT enhancer binding protein (C/EBP) sites were identified within this region and their functional importance was confirmed by mutational analysis, gel retardation, and oligonucleotide pulldown assays. Using chromatin immunoprecipitation (ChIP), we demonstrated that C/EBPalpha binds to the SGP gene promoters lactoferrin and HNC in nonexpressing cells. Upon induction of maturation, C/EBPalpha binds to these promoters and this binding correlates with the expression of both SGP genes. CONCLUSION: We conclude that in the later stages of myeloid development, SGP genes are coordinately upregulated, and that members of the C/EBP family of transcription factors, in particular C/EBPalpha and C/EBPepsilon, play specific and unique roles in upregulating their expression. PMID- 15661398 TI - Histone deacetylase inhibitors profoundly decrease proliferation of human lymphoid cancer cell lines. AB - Methylation of tumor suppressor genes is frequently observed in human cancers. These genes are silenced by histone deacetylase (HDAC) recruited by methylated DNA in their promoter regions. HDAC removes acetyl groups from histones and prevents the basic transcriptional machinary access to the target gene, leading to transcriptional repression. HDAC inhibitors (HDACIs) can restore the expression of the tumor suppressor and/or cell cycle regulatory genes in cancer cells and block the cellular proliferation of these cells. In this study, we investigated the in vitro antiproliferative activities of the HDACIs, suberoylanilide hydroxamic acid (SAHA), and valproic acid against 14 human lymphoid cancer cell lines. All of these cell lines were sensitive to the antiproliferative effects of the HDACI. SAHA induced either G1 or G2-M arrest as well as apoptosis. SAHA downregulated cyclin D1 and D2, and upregulated p53, p21, and p27. Chromatin immunoprecipitation analysis revealed a remarkable increase in the level of acetylated histones associated with the p21 promoter after SAHA treatment. In nude mice, SAHA significantly inhibited growth of a mantle cell lymphoma without major toxic side effects. In summary, HDACIs are promising therapeutic agents for human lymphoid cancers. PMID- 15661399 TI - P-glycoprotein is implicated in the inhibition of ceramide-induced apoptosis in TF-1 acute myeloid leukemia cells by modulation of the glucosylceramide synthase pathway. AB - OBJECTIVE: Ceramide, an intermediate of apoptosis induction in response to chemotherapy, can be detoxified by glycosylation at the cytoplasmic surface of the Golgi membrane. P-glycoprotein (p-gp) might augment ceramide glycosylation by translocating glucosylceramide (GC) across the Golgi membrane. We aimed to show that glucosylceramide synthase (GCS) activity is linked to p-gp expression and resistance to ceramide-induced apoptosis in acute myeloid leukemia (AML). METHODS: Apoptosis and cell-cycle analysis were measured using propidium iodide staining and flow cytometry. Fluorescent microscopy assessed p-gp expression in, and rhodamine 123 uptake by, the Golgi. P-gp interaction with GC was assessed by modulation of rhodamine accumulation. The GCS activity assay was based upon the transfer of UDP-(3)H-glucose to C8-ceramide to form radiolabeled GC, by rate limiting cell-derived GCS. TLC and fluorimetry were used to measure the metabolites of fluorescent ceramide. Cell viability was measured using 7-amino actinomycin D staining and flow cytometry with an internal standard for cell enumeration. RESULTS: P-gp(+) cell lines (KG1a, TF-1) were resistant to C8 ceramide-induced apoptosis compared to p-gp(-) cell lines (HL-60, U937). P-gp inhibitors GF120918 and cyclosporin A enhanced ceramide-induced apoptosis in the p-gp expressing cells. P-gp expression was identified in the Golgi of these cells. Pgp's efflux function in TF-1 but not KG1a cells was inhibited by glucosylceramide. In the presence of p-gp inhibitors, R123 accumulation in the Golgi of TF-1 cells was lost, and GCS activity and lactosylceramide formation were downregulated. Intact cells were necessary for the involvement of p-gp in the regulation of GCS activity. CONCLUSION: Our data suggests that ceramide induces apoptosis in AML cells and that p-gp confers resistance to ceramide induced apoptosis, with modulation of the ceramide-glucosylceramide pathway making a marked contribution to this resistance in TF-1 cells. PMID- 15661400 TI - Human platelets exhibit chemotaxis using functional N-formyl peptide receptors. AB - OBJECTIVE: Activated platelets participate in inflammatory and microbicidal processes by upregulation of surface selectins, shedding of CD40 ligand, and release of platelet microbicidal proteins and microparticles. Given their myeloid lineage, we hypothesized that platelets express functional N-formyl peptide receptors and respond to the bacterially derived chemotactic peptide N-formyl peptide with gradient-driven chemotaxis. METHODS AND RESULTS: Here we show specific binding of N-formyl peptides to the surface of activated platelets. Platelet expression and function of the formyl peptide receptor, FPR, was verified by RT-PCR of the differentiated megakaryocyte MEG-01 cell line, immunoblotting of platelet proteins, and calcium mobilization in platelets with formyl peptide binding. Furthermore, we demonstrate gradient-driven chemotaxis of platelets by video microscopy and transwell migration toward formyl peptides. We also show that endogenous formyl peptides, released by eukaryotic mitochondria from necrotic cells, induce chemotaxis using formyl peptide receptors expressed by thrombin-activated platelets. Conversely, supernatants from cells undergoing apoptotic cell death do not induce platelet chemotaxis. Platelet chemotaxis to formyl peptides was blocked with FPR-specific antibody as well as by pertussis toxin inhibition of the formyl peptide G-coupled receptor. CONCLUSION: These data establish a new role for platelets in host defense and suggest reexamination of their active function in microbicidal and other host defense activities. PMID- 15661401 TI - Discovery and characterization of a selective, nonpeptidyl thrombopoietin receptor agonist. AB - OBJECTIVE: Peptide and other small molecule agonists have been described for several cytokines and growth factors. Hydrazone compounds described here as thrombopoietin receptor agonists were identified as activating STAT proteins in a Tpo responsive cell line. METHODS: STAT activation and analysis of signal transduction pathways in cell lines and normal human platelets was elucidated by Western blot and electrophoretic mobility shift assays. Proliferation assays in cell types responsive to other cytokines determined specificity for Tpo receptor. Flow cytometry quantified differentiation of CD34(+) cells into CD41(+) megakaryocytes and platelet production in vitro. RESULTS: Activation of STAT5, mitogen-activated protein kinase, p38, and early response genes by SB 394725 was similar to that induced by Tpo. SB 394725 induced a reporter gene response under a STAT activation promoter as well as the megakaryocyte-specific gpIIb promoter. The compound induced proliferation of Tpo responsive lines but demonstrated no activity in cell lines responding to other cytokines, i.e., erythropoietin, granulocyte-colony stimulating factor, interleukin-3, interferon-gamma. The response of normal human Tpo receptors was elucidated by measuring growth and differentiation of human bone marrow in vitro. Activation of endogenous Tpo receptors by SB 394725 was demonstrated in human and chimp platelets, but not in platelets of other species including mouse, dog, rabbit, or cynomolgus monkey. CONCLUSIONS: SB 394725, a small molecule with a molecular weight of 452 Da, is capable of activating Tpo-specific signal transduction, proliferation, and differentiation responses similar to the responses and functions of the protein growth factor, Tpo. PMID- 15661402 TI - Engagement of integrin alpha4beta1 enhances thrombopoietin-induced megakaryopoiesis. AB - OBJECTIVE: Studies in numerous adherent cell systems have indicated that engagement of integrins is required for cell survival and proliferation. Although not classically thought of as an adherent cell type, megakaryocytes in the marrow develop in juxtaposition to endothelial cells which display a number of integrin counter-receptors. Moreover, a number of other hematopoietic cell types, including stem cells and erythroid progenitors, have been shown to engage and be affected by integrin ligands. METHODS: The role of beta1 integrins in thrombopoietin-mediated megakaryopoiesis was studied using both gain-of-function and loss-of-function strategies. RESULTS: We found that pan-blockade of integrins with a relatively nonspecific disintegrin blocked TPO-induced MK growth, but that an alpha5beta1 disintegrin, and a function-blocking monoclonal antibody, failed to affect megakaryopoiesis in vitro. In contrast, a neutralizing alpha4beta1 monoclonal antibody blocked TPO-induced MK growth, and an integrin alpha4beta1 ligand, the H296 fragment of fibronectin, enhanced MK growth at all concentrations of TPO. CONCLUSIONS: These findings have important implications for thrombopoiesis in general, and potentially for the enhanced platelet production found in states of systemic inflammation and following the use of therapeutic strategies designed to block alpha4beta1 integrin engagement in states of chronic inflammation and autoimmunity. PMID- 15661403 TI - Clock gene expression in purified mouse hematopoietic stem cells. AB - OBJECTIVE: Circadian genes have recently been characterized in many tissues, but not in hematopoietic stem cells. These cells are rare in the bone marrow (BM), which makes it difficult to collect enough cells for detailed molecular analysis in a short period of time without reduced RNA quality. The aim was to improve methodology and reliability of clock gene expression analysis in purified mouse hematopoietic stem cells. METHODS: Stem cells were highly enriched by high-speed flow cytometric cell sorting of the side population (SP) cells from Hoechst 33342 (Hoechst)-stained mouse BM. Total RNA was isolated from sorted SP and whole BM cells and exposed to DNase treatment. The relative mRNA levels of major clock genes mPer1, mPer2, mBmal1, mCry1, mClock, and mRev-erb alpha were measured with real-time quantitative reverse transcription polymerase chain reaction (Q-RT-PCR) and normalized to m36B4, used as a reference gene. The clonogenity of sorted SP cells and whole BM; cells taken before and after sorting, were tested in colony formation assay. RESULTS: Clock gene activity in sorted SP cells showed pronounced relative differences compared with whole BM for mPer1 and mCry1. The high-speed sorting procedure did not influence clock gene expression or cell clonogenity, even when this was performed with a delay period up to 24 hours. CONCLUSIONS: We demonstrated expression of six clock genes in mouse hematopoietic stem cells. A combination of high-speed flow cytometric sorting and Q-RT-PCR was shown to be useful and reliable for analysis of clock gene activity in small stem cell fractions. PMID- 15661404 TI - G-CSF-primed hematopoietic stem cells or G-CSF per se accelerate recovery and improve survival after liver injury, predominantly by promoting endogenous repair programs. AB - OBJECTIVE: On the basis of the recently recognized potential of bone marrow (BM) cells to give rise to hepatocytes, we investigated the possibility that granulocyte colony-stimulating factor (G-CSF)-mobilized BM cells could home to the injured liver and promote tissue repair. We also examined the origin of cells (endogenous or BM) reconstituting liver after damage. METHODS: Acute and chronic liver injury models were generated by injecting CCl4 in C57Bl6 mice and G-CSF was administered in hematopoietic stem cell (HSC) mobilization doses. After sex mismatched BM transplantation into lethally irradiated recipients and treatment with CCl4 +/- G-CSF, sry (sex-determining region for Y chromosome) protein was detected by immunohistochemistry in liver sections. Double immunohistochemistry for sry and ki-67 protein was used to define the origin of proliferating cells reconstituting liver after injury. RESULTS: In both acute and chronic liver injury model, G-CSF administration ameliorated the histological damage and accelerated the regeneration process. This was accompanied by a strong survival benefit in G-CSF-treated group vs CCl4 group. Quantitative analysis showed higher percentage of BM-origin hepatocytes in the CCl4+G-CSF group compared with the CCl4 group, although the liver engraftment rate still remained rather low. Double staining for ki-67 and sry demonstrated that the recovery acceleration after chemical injury and G-CSF treatment was mainly mediated by increased proliferation of host hepatocytes (ki-67(+)/sry(-)) with less support from BM origin cells (ki-67(+)/sry(+)). CONCLUSION: G-CSF treatment significantly improved survival and liver histology in chemically injured mice, predominantly by promoting endogenous repair mechanisms. Therefore, mobilization with G-CSF might offer a novel therapeutic approach for the treatment of acute and chronic liver diseases in humans. PMID- 15661405 TI - Purging of peripheral blood stem cell transplants in AML: a predictive model based on minimal residual disease burden. AB - OBJECTIVE: Minimal residual disease (MRD) present in peripheral blood stem cell (PBSC) products of AML patients may contribute to relapse. Our goal was to 1) predict leukemia recurrence based on the frequency of MRD present in PBSC products, 2) establish the efficacy of different purging procedures, and 3) integrate this into a model that enables to predict whether or not to purge. METHODS: Minimal residual disease was measured with flow cytometry using leukemia associated phenotypes as established at diagnosis. Toxicity of purging procedures was established using clonogenic assays. Purging procedures used were cryopreservation, hyperthermia, ether lipid ET-18-OCH3, and combinations. RESULTS: Minimal residual disease in PBSC products correlated significantly with relapse-free survival (n=24, p=0.003). At a cut-off value of 0.05% MRD the relative risk of relapse was 4.6 times lower in the group with less than 0.05% MRD. As measured in 54 PBSC products, the MRD level was less than 0.05% in 17 of 54 cases, between 0.05% and 0.5% in 19 of 54 cases, and higher than 0.5% in 18 of 54 cases. Based on the MRD cut-off of 0.05%, the log tumor reduction needed to achieve this threshold is zero for the 17 of 54 cases in which MRD was below 0.05%, less than or equal to 1 log in 19 of 54 cases, and greater than 1-2 log in 18 of 54 cases. When applying purging with 25 mug/mL ET-18-OCH3 combined with cryopreservation at 10% DMSO and hyperthermia at 42 degrees C combined with cryopreservation at 10% or 4% DMSO, there was greater than or equal to 1 log depletion of AML blasts. CONCLUSION: This study establishes (1) a threshold level for MRD above which prognosis is worse, (2) that stem cell products from 69% of patients have higher than this "safe" MRD level, and (3) that ET-18-OCH3 and hyperthermia may be used to purge products in part of these patients. PMID- 15661406 TI - Intraoperative frozen section diagnosis in urological oncology. AB - The intraoperative frozen sections are indicated if the pathological findings change the surgical procedure. In urological oncology is not recommended, as a general attitude, in the tumor diagnosis/staging during the surgery. The assessment of the surgical margins is recommended in partial surgical resections but the literature discourages its systematic use in the radical surgical resections. The assessment of the lymph nodes is specially indicated in the penile cancer with intermediate or high risk and non-palpable nodes, and is debated its utility in non-palpable lymph nodes of cystectomies and prostatectomies. PMID- 15661407 TI - The current status of therapy for symptomatic late-onset hypogonadism with transdermal testosterone gel. AB - For over 50 years, testosterone therapy has been used for the treatment of hypogonadism. In recent years, there has been an increase in the use of testosterone therapy for men with late-onset hypogonadism, as more convenient and effective modes of application are developed. Testosterone therapy in these men can significantly improve their sense of well-being, and lead to increases in muscle and bone mass, upper body strength, virility and libido [Gruenewald, Matsumoto. J Am Geriatr Soc 2003;51:101; Morales. Aging Male 2004; in press]. However, ensuring that optimal testosterone therapy is achieved in men with hypogonadism remains challenging. Oral delivery of unmodified testosterone is not possible, due to rapid first-pass metabolism and its short half-life. Therefore, different derivatives and formulations of testosterone have been developed to enhance potency, prolong duration of action or improve bioavailability. In addition, several different routes of administration have now been evaluated, including intramuscular injections, oral formulations, transdermal patches, transbuccal systems and transdermal testosterone gel. Despite the broad range of testosterone therapy on offer, each form has its benefits and limitations, and some will suit one patient more than another. An important concern among clinicians is that testosterone therapy may cause or promote prostate cancer. While current evidence supports the safety of testosterone therapy, androgens are growth factors for pre-existing prostate cancer. Therefore, before therapy is initiated, careful digital rectal examination and determination of prostate specific antigen (PSA) in serum should be performed, in order to exclude evident or suspected prostate cancer. The first 3-6 months after initiating testosterone therapy is the most critical time for monitoring effects on the prostate. Therefore, it is important to monitor PSA levels every 3 months for the first year of treatment; thereafter, regular monitoring (mostly for prostate safety but also for cardiovascular and haematological safety) during therapy is mandatory. PMID- 15661408 TI - Neuroendocrine differentiation in prostate cancer: implications for new treatment modalities. AB - OBJECTIVES: This review aims to provide practising clinicians with the most recent knowledge of the biological nature of prostate cancer (PC) to facilitate investigation of new treatment modalities for patients with PC, especially the hormone-refractory state of the disease. METHODS: Review of the literature using PubMed search and scientific journal publications. RESULTS: Much progress has been made towards an understanding of the development and progression of PC, and the factors which drive the development of androgen independence. Neuroendocrine (NE) cells may provide an intriguing link between NE cell differentiation and tumour progression in PC with the genetically supported formation of androgen independent clones which regulate the proliferation of neighbouring non-NE phenotype cancer cells in a paracrine manner by secretion of NE products. Various NE peptides stimulate proliferation of androgen-independent PC through transactivation of the androgen receptors (AR). Therefore, cancerous epithelial cells that increase their responsiveness to NE factors or induce NE cells to release trophic factors may have a survival advantage over their siblings. CONCLUSION: This review shows the need to improve our understanding of the biological nature of PC, especially cancer cells of the NE phenotype and their regulatory products to develop new therapeutic protocols and trials based on NE hormones and their agonists/antagonists. PMID- 15661409 TI - EAU guidelines on renal transplantation. AB - OBJECTIVES: To produce a guidelines text, on behalf of the European Association of Urology, providing insights in the issues surrounding renal transplantation. METHOD: A group of international experts in renal transplantation carried out a non-structured literature review on available medical databases and urological literature. RESULT: A guideline text is presented providing an overview of key issues involved in the patients' management such as assessment of donors, pre transplant evaluation, techniques, management, post-transplant care, etc. CONCLUSION: The current text represents a consensus statement developed by a group of international experts in renal transplantation. PMID- 15661410 TI - Laparoscopic radical prostatectomy: prospective evaluation of the learning curve. AB - OBJECTIVE: We prospectively evaluated the learning curve (LC) of laparoscopic radical prostatectomy (L-RPE) regarding the improvements in operative times (OT) and technical difficulty in one-operator-practice as it compares with open RPE. METHODS: Over 18 months, 50 L-RPE were performed by an inexperienced surgeon in laparoscopy but skilled in open surgery. Difficulty scores were obtained at the completion of each L-RPE comparing L-RPE to open RPE. OT, estimated blood loss (EBL), length of stay, and catheterization time were also obtained. RESULTS: In the ablative part of L-RPE the median difficulty score was significantly higher (p<0.001) for the first 10 cases, decreased dramatically by case 11 becoming equivalent (p=0.3) to open RPE and by case 31 the L-RPE becomes significantly easier than open RPE (p=0.002). The difficulty scores for the urethrovesical anastomosis performance is always uniformly higher for the whole patient series (p<0.001). Median OT decreased significantly from 293 minutes in the first 10 cases to 114 minutes in the last 10 cases (p<0.001). Catheterization time and length of hospitalization, decreased significantly with the progression of the LC. EBL remained stable throughout the patient cases. Obesity, prior surgery, and extension of the procedure (lymphadenectomy, nerve-sparing) significantly increased the OT. CONCLUSION: Although the ablative part of L-RPE has a relatively short LC for a skilled open surgeon reflected by the rapid decrease in difficulty scores and OT by case 21, the performance of anastomosis shows a longer LC. Intensive training on anastomosis may be necessary to master this skill. PMID- 15661411 TI - Influence of nerve-sparing (NS) procedure during radical prostatectomy (RP) on margin status and biochemical failure. AB - PURPOSE: To evaluate whether nerve-sparing procedure itself is a risk factor for biochemical recurrence in carefully selected patients. MATERIAL AND METHODS: We compared patients of our historical series who in retrospect were candidates for nerve-sparing (NS) procedure with a contemporary cohort of patients. With respect to stage migration and selection bias between these two groups we performed a multivariate analysis adjusting for all explanatory variables in the model. NS was performed in n = 723 patients (bilateral n = 359, unilateral n = 364) in comparison to n = 620 patients undergoing non-NS RP, comprising n = 756 patients within the favorable pT2 category. We examined the association of clinical and histopathological parameters in relation to PSA recurrence in uni- and multivariate analyses including NS as a variable. Furthermore, for each prostate lobe separately we determined whether surgical procedure (nerve-sparing vs. non nerve-sparing RP) resulted in a positive margin. RESULTS: In univariate analysis there was no difference in pT2 (log rank p = 0.091), pT3a (log rank p = 0.171) and pT3b (log rank p = 0.110) cancers between patients treated with NS compared to non-NS surgery. The 3- and 5-year recurrence free survival rate for patients with pT2, pT3a and pT3b cancers treated by NS vs. non-NS were 96.3/94.9 vs. 94.9/90.8, 75.0/61.8 vs. 73.4/55.0 and 46/30 vs. 38/23. Multivariate regression analysis showed no association with PSA failure (p = 0.798) for patients who underwent NS. Capsular penetration (p < 0.001), lymph-node status (p < 0.001), seminal vesicle invasion (p < 0.001), surgical margin status (p = 0.0130), Gleason score (p < 0.001) and preoperative PSA (p = 0.005) were significantly associated with risk of failure. The positive margin rate per each prostate lobe in pT2 cancers was 6.5% vs. 5.1% in NS and non-NS cases, 10.3% vs. 17.3% in patients with extracapsular extension and 15.0% vs. 25.1% in cases with seminal vesicle invasion respectively. CONCLUSION: NS RP is an oncologically safe procedure provided that appropriate preoperative selection of patients by means of a validated nomogram is performed. Moreover, evaluation of positive margins in patients undergoing NS and non-NS RP revealed no evidence that adequacy of tumor excision is compromised by NS procedure. PMID- 15661412 TI - Validation of 2001 Partin tables in Turkey: a multicenter study. AB - OBJECTIVE: Although Partin tables were developed in United States to predict the stage of prostate cancer preoperatively, they are used by many clinics throughout the world assuming that these figures apply to their population as well. However the predictive value of current Partin tables, which was updated in 2001, has not been validated in most of the countries as well as in Turkey. Therefore, we evaluated the validity of 2001 Partin tables, for the ability to predict the pathological stage in Turkish patients. PATIENTS AND METHODS: The clinical and pathological findings of 1043 patients who have had radical prostatectomy were assessed. Serum PSA values, clinical stage, biopsy Gleason score and the pathological features of the radical prostatectomy specimens were collected from each clinic and evaluated. The predictive value of Partin nomogram and pathological findings of prostatectomy specimens were compared and analyzed according to Receiver Operating Characteristics (ROC) analysis. RESULTS: Median age of the patients was 60 (45-74). In the presented study, percentage of patients with clinical stage T1c was 43%. Patients with Gleason score of 2-4 in biopsy constituted 23.4% of the study group. In the present study, the ratio of the patients with serum PSA higher than 10 ng/ml was 39.6%. Organ confined disease, seminal vesicle involvement, lymph node metastases ratios were 64.7%, 10.3%, 1.8% respectively. Area Under Curve (AUC) values for organ confined disease, seminal vesicle involvement and lymph node involvement were calculated as 0.665, 0.733 and 0.759 respectively. CONCLUSION: It appears that Partin tables have a reasonable predictive value for the final pathological features like organ confined disease, seminal vesicle and lymph node involvement in Turkish patients. This multicenter study showed that current Partin tables could also be used in Turkish patients with comparable accuracy. PMID- 15661413 TI - Feasibility and limitations of comorbidity measurement in patients undergoing radical prostatectomy. AB - OBJECTIVE: To compare different comorbidity classifications as predictors of survival after radical prostatectomy (RPE). METHODS: 444 consecutive RPE patients (mean follow-up 6.9 years) were stratified according to age, Charlson score, American Society of Anesthesiologists Physical Status classification (ASA), New York Heart Association classification of cardiac insufficiency, classification of angina pectoris of the Canadian Cardiovascular Society and a combination of both cardiac risk scores. Comorbid and overall mortality were the study endpoints. Mantel-Haenszel hazard ratios, p values and 8-year survival probabilities were used for comparison. A modified Charlson score was created by a restriction to the five individual conditions significantly associated with comorbid mortality. RESULTS: When three strata (low, intermediate, high risk) were used, all stratifications displayed dose-response patterns and reached statistical significance as predictors of survival at least for the high-risk group. Only the ASA classification, however, discriminated three significantly different risk groups. Only the modified (restricted) Charlson score reached statistical significance as predictor of comorbid mortality in the age group of 70 or more years. CONCLUSION: Although all investigated comorbidity classifications had some prognostic relevance in patients selected for RPE, their clinical applicability appears to be limited beyond the 70th year of life. The results of this study might, nevertheless, assist the treatment decision in patients with low-risk tumors eligible for modern watchful waiting strategies who mainly belong to the age group between 60-69 years where comorbidity seems to be of distinct prognostic value. PMID- 15661414 TI - Trends in reporting Gleason score 1991 to 2001: changes in the pathologist's practice. AB - OBJECTIVE: The prostate specific antigen (PSA) era has been associated with a grade migration towards moderately-differentiated (Gleason 5-7) prostate cancer. We investigated whether changes in interpretation of the Gleason system could be a contributing factor by reviewing the Gleason scores for prostate cancer in our region. PATIENTS AND METHODS: Records of patients with prostate cancer assigned a Gleason score between 1991-2001 were retrospectively reviewed. We analysed trends in Gleason score, method of diagnosis and age at diagnosis. Following this, 50 cases from the dataset were randomly selected (stratified to contain half Gleason 2-4 reports) and reviewed in a blinded manner by an uropathologist and given a new Gleason score. RESULTS: 2737 patients were diagnosed and given a Gleason score; 1484 by prostate biopsy (PB) and 1172 by transurethral resection of prostate (TURP). 273 radical prostatectomy (RP) specimens were received, although the results of pre-operative biopsies were available in only 192 of these patients. Over time, there was an increase in the proportion of patients with Gleason 5-7, and a significant decrease in reporting of Gleason 2-4 cancer (r2 = 0.81, p < 0.0001). In 1991, 24% of cancers were Gleason 2-4; in 2001 this had decreased to 2.4%. TURP was associated with more Gleason 2-4 reports (23%) compared with PB (13.2%) and RP (9.2%). On blinded review, all Gleason 2-4 reports were upgraded to Gleason 5-7 cancer (p < 0.001). CONCLUSION: Over time, the proportion of Gleason 2-4 prostate cancer reported has significantly decreased. Our study suggests that a change in practice by the pathologist is a significant factor in this grade migration. PMID- 15661415 TI - Pfannenstiel versus vertical laparotomy in patients undergoing radical retropubic prostatectomy with spinal anesthesia: results of a prospective, randomized trial. AB - OBJECTIVE: To evaluate the impact of a standard vertical laparotomy versus a Pfannenstiel transverse laparotomy on intra-, peri-operative, and 6-month follow up outcome in patients undergoing radical retropubic prostatectomy with pelvic lymphadenectomy with spinal anesthesia. METHODS: Between January 2003 and June 2003, 69 age-matched consecutive patients with clinically localized prostate cancer underwent radical retropubic prostatectomy with pelvic lymphadenectomy with spinal anesthesia and were randomized into Group 1 (vertical laparotomy: 35 patients) and Group 2 (Pfannenstiel laparotomy: 34 patients). An extensive analysis of the critical intra-, peri-operative, and 6-month follow-up clinical parameters was performed. RESULTS: Both the hemodynamics and the biochemical balance were not significantly different between the two groups. Overall blood loss (p = 0.78), autologous (p = 0.88) and homologous (p = 0.36) blood transfusions were similar regardless of the type of laparotomy. Surgical time was not significantly (p = 0.27) different between the two groups. Similarly, the two forms of laparotomy did not differ regarding the length of the surgical incision (p = 0.21), as measured at the end of the procedure. Post-operative oxygen saturation percentage by pulse oximetry, as well as post-op sedation score, were not significantly different (p = 0.06 and p = 0.97, respectively). Waiting time in the post-operative holding area (p = 0.15), and pain score in the post operative holding area (p = 0.9) as well as on post-operative day 1 (p = 0.1) were not significantly different between the two groups. The rate of first flatus passage and of unassisted ambulation were similar regardless of the type of laparotomy during post-operative day day 1. The two types of incision made it possible to remove a similar (p = 0.34) number of pelvic lymph nodes and were associated to a similar rate of positive surgical margins among pT2 patients. At the 6-month follow-up the occurrence of a pelvic lymphocele and of deep venous thrombosis was similar in the two groups (p = 0.6 and p = 0.16, respectively). Complete urinary continence and spontaneous erectile function recovery was reported in a similar number of patients regardless of the type of surgical incision (p = 0.59 and p = 0.40, respectively). CONCLUSIONS: These results suggest that a Pfannenstiel transverse suprapubic laparotomy does not result in a significantly different outcome from a standard vertical laparotomy in patients undergoing a radical retropubic prostatectomy with pelvic lymphadenectomy with L2 L3 spinal anesthesia for clinically localized prostate cancer. PMID- 15661416 TI - The comparison of artificial urinary sphincter implantation and endourethral macroplastique injection for the treatment of postprostatectomy incontinence. AB - OBJECTIVES: To compare the effectiveness of macroplastique injection with artificial urinary sphincter implantation (AUS) for treatment of postprostatectomy incontinence (PPI). METHODS: A prospective randomized clinical trial including 45 patients with PPI was performed secondary to radical retropubic prostatectomy (RRP), transvesical prostatectomy (TVP), transurethral prostatectomy (TURP), and TURP with TVP, in 12, 16, 16, 1 patients respectively. Patients were divided into two groups as minimal (group I) and total incontinence (group II) according to the severity of incontinence. Respectively, Group I (n = 21) and group II (n = 24) patients were randomized as AUS implantation (n = 11, n = 11) and macroplastique injection (n = 10, n = 13). Follow-up period was 48 (6 84) months in patients with macroplastique injection and 60 (8-120) months in AUS implantation. The success of the treatment was evaluated by calculating the average number of pads used by the patient per day, the weight of the pads and score of quality of life survey scale for each group both in the preoperative and in the postoperative period. RESULTS: There were statistically significant differences between preoperative and postoperative average pad weight, average number of pads and quality of life scores, both in patients with minimal and total incontinence. In group I there was no statistically significant difference between the two techniques. However, in group II there was a significant difference favoring AUS implantation. CONCLUSIONS: Endourethral injection should be the treatment of choice for patients with minimal incontinence, whereas AUS implantation as the first choice for patients with total incontinence. PMID- 15661418 TI - Soft prosthesis implant and relaxing albugineal incision with saphenous grafting for surgical therapy of Peyronie's disease: a 5-year experience and long-term follow-up on 145 operated patients. AB - INTRODUCTION: Most surgical therapies used to date for Peyronie's disease produce good results but cause penis retraction. The authors propose a personal surgical technique that makes use of an intracavernous cylinder, soft but axially rigid in consistency, in association with tunical relaxing incisions and saphenous vein grafting, in this exact sequence. METHODS: Between March 1997 and March 2003, 145 patients presenting with Peyronie's disease underwent this 3-phase personal surgical procedure. Complete plaque excision was necessary only for 13 patients (8.9%); in the remaining 132, a single incision was placed in 113 cases (77.9%) and double incisions were used for 19 patients (13.1%). Penile length was measured intraoperatively, after drug-hydraulic erection, then at the later control visits. Distance follow-up ranged from a minimum of 6 months to a maximum period of 3 years (mean 13 months); the latter follow-up period was possible for 38 patients. All patients and partners with at least one year follow-up were asked to answer a questionnaire administered over the telephone at 12 months post surgery. RESULTS: Average penile lengthening was approximately 1.5 cm. 80 patients answered the questionnaire (70.75%): 76 expressed complete satisfaction, while 4 patients reported glans paresthesia. No major complications were noted; for 8 patients circumcision was repeated because of preputial edema. In 6 cases there was a local, postoperative infection, that was treated conservatively with a prolonged antibiotic therapy. CONCLUSIONS: This technique allows to successfully solve penile retraction, caused by traditional surgery for Peyronie's disease. The technique is easy to carry out, has low risks of complications and ensures very high rates of patient satisfaction. PMID- 15661417 TI - Chronic treatment with tadalafil improves endothelial function in men with increased cardiovascular risk. AB - OBJECTIVE: Erectile dysfunction (ED) is often associated with a cluster of risk factors for coronary artery disease and reduced endothelial function. Acute and chronic administration of oral sildenafil, a phosphodiesterase type 5 (PDE5) inhibitor, improves endothelial function in patients with ED. Tadalafil (TAD) is a new PDE5 inhibitor with a long half life that allows alternate day administration. Aim of the study was to evaluate whether chronic therapy (4 weeks) with TAD improves endothelial function in patients with increased cardiovascular risk and whether this effect is sustained after discontinuation of therapy. METHODS: We randomized 32 patients with increased cardiovascular risk to receive either TAD 20 mg on alternate days or matching placebo (PLB) for 4 weeks. Patients underwent evaluation of brachial artery flow-mediated dilation (FMD), nitrite/nitrate and endothelin-1 plasma levels at baseline, at the end of treatment period and after two-weeks follow-up. RESULTS: At 4 weeks, FMD was significantly improved by TAD (from 4.2+/-3.2 to 9.3+/-3.7%, p<0.01 vs. baseline), but was not modified by PLB (from 4.1+/-2.8 to 4.0+/-3.4%, p=NS). At 6 weeks the benefit in FMD was sustained in patients that received TAD (9.1+/-3.9% vs. 4.2+/-3.2%, p=0.01 vs. baseline; 9.1+/-3.9% vs. 9.3+/-3.7%, vs. 4 weeks, p=NS) while no changes in FMD were observed in patients randomized to PLB. Also, compared to baseline, a net increase in nitrite/nitrate levels (38.2+/-12.3 vs. 52.6+/-11.7 and 51.1+/-3.1, p<0.05) and a decrease in endothelin-1 levels (3.3+/ 0.9 vs. 2.9.+/-0.7 and 2.9+/-0.9, p<0.05) was found both at four and six-weeks after TAD; these changes were inversely correlated as shown by regression analysis (adjusted R2=0.81, p<0.0001). CONCLUSIONS: Chronic therapy with TAD improves endothelial function in patients with increased cardiovascular risk regardless their degree of ED. The benefit of this therapy is sustained for at least two weeks after the discontinuation of therapy. Larger studies are needed in order to assess the possible clinical implications of chronic therapy with TAD. PMID- 15661419 TI - Laparoscopic sacral colpopexy approach for genito-urinary prolapse: experience with 363 cases. AB - OBJECTIVE: To evaluate the surgical outcome, complications and benefits of laparoscopic double promonto-fixation for patients with pelvic prolapse. METHODS: Women with genito-urinary prolapse underwent a transperitoneal placement of a 100% polyester mesh on the anterior vaginal wall and a posterior mesh on the levator ani muscle. Both of these were anchored to the sacral promontory. A TVT was placed simultaneously in patients who had concurrent stress urinary incontinence. RESULTS: A total of 363 patients were operated upon between 1996 and 2002. Their mean age was 63 (range 35-78), average follow-up was 14.6 months, the mean operating time was 97 minutes. There were 8 conversions due to anesthetic or surgical difficulties. Follow up was done by a postal questionnaire and physical examination at 6 months and then yearly. 96% were satisfied with the results of their operation and no patients complained of sexual dysfunction. There was a 4% recurrence rate of prolapse, 3 vaginal erosions, 2 urinary retentions that required TVT section, 1 bowel incarcerations, 1 spondylitis and 2 mesh infection. CONCLUSIONS: Laparoscopic promonto-fixation is feasible and highly effective technique that offers good long-term results with complication rates similar to open surgery, with the added benefits of minimally invasive surgery. PMID- 15661420 TI - Patient satisfaction and clinical efficacy of the new perineal bone-anchored male sling. AB - OBJECTIVE: We retrospectively evaluated the safety, efficacy and patient satisfaction following the male sling procedure for stress urinary incontinence. MATERIALS AND METHODS: Forty-six patients, with a median age of 67 years, underwent the perineal male sling procedure for stress urinary incontinence. Radical retropubic prostatectomy was the commonest etiology of SUI. The male sling was placed over the bulbar urethra, through a perineal incision and anchored with the help of bone screws inserted in the pubic rami. Efficacy and treatment satisfaction were evaluated with the help of the validated UCLA/RAND questionnaire and questions addressing treatment satisfaction. Patients were defined as "cured" if they were dry or "improved" if utilizing 1-2 pads per day (based on response to question 1 of the UCLA/RAND questionnaire. Responses to questions 4 and 5 of the UCLA/RAND were utilized to determine the impact of the procedure on urinary leakage and urinary function postoperatively. Complications if any were noted. RESULTS: One patient developed infection and two developed short-lasting perineal/buttock pain. Significantly, no patient developed urethral erosion. At a mean follow-up of 24 months, 34 patients were either dry (17/46, 37%) or utilizing only 1-2 pads per day (17/46, 37%), giving a cure/improvement rate of 74%. On evaluating the response to questions 4 and 5 of the UCLA/RAND questionnaire, 72% patients stated that urinary leakage and function were a "small to no problem". Moreover, 59% patients were moderately or completely satisfied with the procedure and 11% stated they were halfway satisfied with the procedure giving an overall satisfaction of 70%. A similar percentage felt that the treatment halfway or completely met their expectations. CONCLUSIONS: At a mean follow-up of 24 months, the male sling procedure appears to be effective in the management of male SUI with a success rate of 74%. Moreover, it is safe as evidenced by the absence of any major complications such as urethral erosion. Finally, it is associated with a 70% patient satisfaction. PMID- 15661421 TI - Effects of local estrogen therapy on recurrent urinary tract infections in young females under oral contraceptives. AB - BACKGROUND: Previous studies have demonstrated the efficacy of local application of estrogen in treating postmenopausal women with recurrent urinary tract infections (RUTI) and urinary incontinence. Younger women under oral contraceptives (OC) can suffer from similar symptoms. The aim of this pilot study was to evaluate the effectiveness of local estrogens on RUTI and the impact of local hormonal supplementation on bladder neck vascularization. METHODS: 30 women (mean age 22.7 years) with a longstanding history of RUTI were included. Pre treatment investigation included complete clinical history, urinalysis, urine culture and cystoscopy. All subjects completed a questionnaire about onset and duration of disease and quality of life before and after treatment. Local (vaginal) estrogen therapy consisted of 1mg estriol (E3) 7 times a week for two weeks and twice a week for two additional weeks. Sonographic examination of bladder vascularization was performed before and after treatment using transperineal color Doppler ultrasound (6 MHz, Acuson Sequoia 512, Mountain View, CA, USA) with a filled bladder. After angle correction, peak systolic blood flow velocity (PSBFV) and end diastolic blood flow velocity (EDBFV) were measured in 2 bladder arteries; and the Resistive Index (RI) was calculated. Flow velocity in each vessel was measured at least four times and the mean value determined. RESULTS: All patients completed the therapy course without severe side effects. Patients had a mean history of RUTI over 2.3 years; the mean period under OC was 3.2 years. In the follow-up period of 11 months after treatment, 24/30 patients reported no symptoms of cystitis and used no additional medication. Normal bladder epithelium in control cystoscopy after E3 therapy was seen in all patients with trigonal metaplasia and vulnerable, highly vascularized urothelium at the initial investigation. RI decreased from 0.945 to 0.705 after treatment (p<0.001), concomitantly the mean EDBFV increased highly significantly from 0.82 cm/sec to 4.45 cm/sec after estrogen treatment (p<0.001). INTERPRETATION: In a majority of young patients under OC and a longstanding history of RUTI, a considerable infection-free period was achieved after local application of estrogen. Decreased RI and increased EDBFV indicate vasodilatation and less peripheral vascular resistance. Responsiveness to local E3 may correspond to improved cystoscopic findings as a consequence of increased bladder perfusion. PMID- 15661422 TI - Application of targeted radiotherapy/gene therapy to bladder cancer cell lines. AB - OBJECTIVES: A targeted radiotherapy/gene therapy strategy for transitional cell carcinoma of bladder is described, using [131I]meta-iodobenzylguanidine ([131I]MIBG), a radionuclide combined with a tumour-seeking drug. The aim is to decrease side effects from radiation toxicity, while increasing radiation dose to tumour. This tumour cell kill approach is augmented by radiological bystander effects. METHODS: The bladder cancer cell line EJ138 was transfected with a gene encoding the noradrenaline transporter (NAT) under the control of tumour-specific telomerase promoters. Resulting uptake of [131I]MIBG was assessed by gamma counting of cell lysates, and NAT transgene expression by real-time RT-PCR. Cell kill of monolayers and disaggregated spheroids, dosed with [131I]MIBG, was assessed by clonogenic assay. RESULTS: NAT gene transfected cells exhibited a significantly increased active uptake of [131I]MIBG, leading to dose-dependent cell kill. Clonogenic assay of disaggregated spheroids, a three-dimensional model, suggested cell kill via bystander effects. CONCLUSIONS: Expression of a functional NAT after in vitro transfection of bladder cancer cells with the NAT gene under the control of telomerase promoters leads to active uptake of [131I]MIBG and dose-dependent cell kill. This strategy could produce a promising new treatment option for bladder cancer. PMID- 15661423 TI - In vivo retroviral mediated gene transfer into bladder urothelium results in preferential transduction of tumoral cells. AB - OBJECTIVES: Superficial bladder tumours are at high risk for recurrence, relapse after resection, escape to intravesical immunotherapy and they may become invasive. New therapeutics are therefore needed to achieve cure. Thus, gene therapy is an attractive new treatment modality for malignant bladder tumours. The purpose of this study was to evaluate the feasibility and the efficiency of retroviral mediated reporter gene transfer into malignant urothelial cells both in vitro and in vivo. METHODS: We evaluated the feasibility of the transfection of bladder tumour with direct intravesical instillation of a defective retrovirus. The vector was derived from LXSN. The efficiency of transduction with the Moloney Leukaemia Murine virus-based vector, amphotrophic retroviral vector, was monitored through the expression of two marker genes (nls-LacZ and NeoR). The canine animal was chosen since it can present with spontaneous bladder carcinomas mimicking human pathology. Primary cultures of two normal canine bladder urothelium and two canine primary bladder tumours were first studied. We then investigated in vivo, in two normal and two spontaneous tumour bearing dogs, the transduction of urothelial cells following direct intravesical instillation of 2.10(4) to 3.10(6) of the retroviral vector. RESULTS: Transduced cells were evidenced in all primary cultures of canine normal urothelium and transitional cell carcinoma. Bladder biopsies from sound dogs instilled with the viral solution showed long lasting transduction up to 60 days long. Bladder cryosections from tumour-bearing dogs displayed transduction of superficial layers of urothelial cancer cells without passing through lamina propria. In vivo transduction was evidenced in 1 to 15% (mean 5%) of the cells in the tumours and preferentially addressed malignant cells. Normal epithelium either originating from sound or tumour-bearing animals was not transduced. CONCLUSION: These results demonstrate for the first time the feasibility of in vivo retroviral transduction of bladder cancer using a clinically relevant procedure. PMID- 15661425 TI - Re: Schrier BPh, Hollander MP, van Rhijn BWG, Kiemeney LALM, Witjes JA. Prognosis of muscle-invasive bladder cancer: difference between primary and progressive tumours and implications for therapy. Eur Urol 2004;45:292-6. PMID- 15661426 TI - Re: Puppo P, Introini C, Calvi P, Naselli A. Long term results of excision of small renal cancer surrounded by a minimal layer of grossly normal parenchyma: review of 94 cases. Eur Urol 2004;46:477-81. PMID- 15661427 TI - Re: Steiner T, Knels R, Schubert J. Prognostic significance of tumor size in patients after tumour nephrectomy for localised renal cell carcinoma. Eur Urol 46;2004:327-30. PMID- 15661429 TI - Active self-management of chronic pain in the community. PMID- 15661430 TI - Pharmacological treatment of peripheral neuropathic pain conditions based on shared commonalities despite multiple etiologies. PMID- 15661431 TI - The effects of racemic ketamine on painful stimulation of skin and viscera in human subjects. AB - Evidence suggests that NMDA receptors may have a differential role in the modulation of visceral and somatic pain. Specifically, animal data indicate an analgesic role of NMDA-R antagonists in acute visceral but not acute somatic pain. In humans analgesic effects are documented in acute somatic pain, while the role of NMDA-R antagonists in acute visceral pain is still questionable. We, therefore, conducted a study in humans comparing the analgesic effects of ketamine in an experimental model of visceral and cutaneous pain. In a double blind, randomized, cross-over study, 11 healthy volunteers (3M, 8F) participated in two experimental sessions in which they evaluated perceptions induced by balloon distention of the distal esophagus and contact heat on the upper chest during continuous computer-controlled i.v. infusion of either ketamine (60 and 120 ng/mL) or saline. Two stimulus intensities producing non-painful and painful sensation were used for each stimulus modality. Subjects reported maximum pain intensity and unpleasantness on visual analog scales (VAS). For noxious visceral stimulation, low dose ketamine produced significant attenuation of both pain intensity and unpleasantness. In contrast, for noxious cutaneous stimulation, ketamine reduced pain unpleasantness, but not perceived intensity. In addition, ketamine did not alter the perception of innocuous stimuli in either modality. Our results confirm the analgesic effects of low-dose ketamine, with minimal side effects, on acute visceral pain and indicate a similar but smaller effect on acute cutaneous pain. A decrease in the unpleasantness but not in the intensity of cutaneous pain may reflect the differential effect of NMDA-R antagonists for the two pain states observed in animal models. PMID- 15661432 TI - The effect on mechanical pain threshold over human muscles by oral administration of granisetron and diclofenac-sodium. AB - Previous studies indicate that plasma levels of serotonin (5-HT) and intramuscular prostaglandin E2 (PGE2) participate in determining the mechanical pain threshold and tolerance level to pressure applied on the skin over healthy muscles. Other studies reported gender differences regarding responses to noxious stimuli. The present study aimed to determine whether the mechanical pain threshold of healthy muscles is influenced by oral administration of 5-HT3 or PGE2-inhibitors and if there are any gender differences in this respect. Ten healthy female subjects and 10 age-matched healthy male subjects participated in the study, which was randomized and double blind with crossover design. Granisetron (5-HT3-antagonist), diclofenac-sodium (PGE2-antagonist) and placebo were administered for 3 days. The pressure pain threshold (PPT) was recorded bilaterally with an algometer over certain orofacial, trunk, and limb muscles before and after administration of the antagonists. The PPT over all muscles combined increased after administration of granisetron. There was no change after administration of placebo. The difference between granisetron and placebo was significant for the trapezius and tibialis anterior muscles. Diclofenac-sodium did not influence the PPT and there was no difference compared to placebo. Although the basal PPT values were lower in females, the PPT response to granisetron differed significantly between genders only in the tibialis anterior muscle. In conclusion, the results of this study showed that oral administration of the 5-HT3-antagonist granisetron increased the PPT over healthy trunk and limb muscles but not over orofacial muscles, and that the response in the limb muscles was greater in males. PMID- 15661433 TI - Spinal noradrenaline transporter inhibition by reboxetine and Xen2174 reduces tactile hypersensitivity after surgery in rats. AB - Spinal noradrenaline (NA) released in response to noxious stimuli may play an important role in suppression of nociceptive transmission. Here, we investigated the efficacy of a competitive NA transporter inhibitor (reboxetine) and a noncompetitive NA transporter inhibitor peptide, Xen2174, isolated from the Pacific cone snail, to treat tactile hypersensitivity following paw incisional surgery. Male Sprague-Dawley rats were anesthetized, an incision of the plantar aspect of the hind paw was performed, and withdrawal threshold to von Frey filaments near the surgical site determined. Reboxetine (0.5-5 microg) and Xen2174 (0.3-100 microg) increased withdrawal threshold when injected 24h after paw incision, with a peak effect at 15-60 min, for Xen2174, an ED50 value of 0.64 microg. Administration of Xen2174 (3-30 microg) 15 min before incision also reduced hypersensitivity in a dose-dependent manner. Withdrawal threshold after the single 30 microg dose was greater than vehicle control even at 2, 3, and 5 days after incision. Doses or=6 months. The patients' maintenance methadone dosage and urine tests for drug abuse during the month before and of the survey were recorded. Of the 170 patients, 94 (55.3%) experienced chronic pain. They had a significantly higher proportion of chronic illness (74.5%) compared to non-pain patients (44.7%) (Fisher's Exact Test P<0.0005). Twelve (12.8%) of the chronic pain patients scored their pain as mild, 38 (40.4%) as moderate, 22 (23.4%) as severe and 22 (23.4%) as very severe. Pain duration significantly correlated with pain severity (Pearson R=0.3, P>0.0005) and was significantly associated with methadone daily dosage: patients with pain duration of >or=10 years (n=26) were receiving the highest methadone dosage (182.1+/-59.2 mg/day), those with pain duration from 1 to 10 years (n=59) 160.9+/-56.2 mg/day, and those with pain duration of <1 year (n=9) 134.2+/-73.2 mg/day. Patients in the non-pain group (n=76) were receiving 147.1+/-52.8 mg/day of methadone (ANOVA, F=3.1, P=0.03). We conclude that pain duration and severity significantly correlated. Although methadone was not prescribed for the treatment of pain but rather for opiate addiction, the patients in the MMT clinic with prolonged pain were prescribed a significantly higher methadone dosage compared to patients with short pain duration, and non pain patients. PMID- 15661443 TI - Long-term effects of neonatal surgery on adulthood pain behavior. AB - The long-term consequences of neonatal noxious stimulation on adulthood pain behavior were investigated in male and female mice. On the day of birth, mouse pups were exposed to a laparotomy under cold anesthesia followed by an analgesic dose of morphine (10 mg/kg) post-operatively, or a saline control. An additional group of subjects was exposed to the non-noxious aspects of the surgical procedure (cold exposure, separation from the dam, injection) comprising a 'sham' surgery control group, whereas another group of control subjects was administered an injection of saline or morphine, but was otherwise undisturbed. Behavioral observations of the pups immediately following the procedure indicated that the laparotomy produced increased distress vocalizations in the ultrasonic range (40 kHz) compared to both groups of control subjects. During 90 min observations periods following the surgery and 1-week later, maternal care did not vary among treatment conditions. In adulthood, offspring were tested for nociceptive sensitivity on the hot-plate (HP; 53 degrees C), tail-withdrawal (TW; 50 degrees C) and acetic acid abdominal constriction test (AC). On both the TW and the AC tests, neonatal surgery decreased pain behavior relative to both groups of control subjects, an effect that was reversed by post-operative morphine treatment. On the HP test, both groups of subjects exposed to the stressful aspects of neonatal surgery (laparotomy or sham surgery) exhibited decreased pain behavior in adulthood. These findings suggest that early exposure to noxious and/or stressful stimuli may induce long-lasting changes in pain behavior, perhaps mediated by alterations in the stress-axis and antinociceptive circuitry. PMID- 15661444 TI - Vagal stomach afferents inhibit somatic pain perception. AB - Vagal stimulation inhibits systemic pain perception in animals, probably via the nucleus tractus solitarius and its connections with descending nuclei in the brainstem which inhibit pain. Pain-inhibiting effects of such stimulation in humans, obtained from epileptic patients treated by vagal stimulation, are controversial. The aim of our study was to evaluate whether vagal stomach afferent activation inhibits pain perception in healthy humans. Pain thresholds, magnitude of tonic heat pain at 46 degrees C stimulation, pain temporal summation and laser pain evoked potentials were measured at the hand before and immediately after rapid drinking of 1500 ml water in 31 volunteers. We found an increase in heat pain threshold from 43.3+/-2.6 to 44.7+/-2.2 degrees C, P<0.0001, a decrease of peak pain magnitude to tonic heat from 56.3+/-26.2 to 43.7+/-25.8 (on 0-100 VAS), P<0.0001, a lowering of area under the curve during tonic noxious heat stimulus from 1962+/-984 to 1411+/-934, P<0.001. Additionally, we observed a decrease in the peak to peak evoked potential amplitude from 19.2 microV+/-1.2 to 15.6 microV+/-1.2 (P=0.005) together with a decrease in the estimation of mean laser induced pain from 52.28+/-18.00 to 48.14+/-20.18 (P=0.025). Mechanical pain thresholds and temporal summation did not change significantly. We conclude that vagal stomach afferents exert an inhibitory effect on somatic pain perception in humans. PMID- 15661446 TI - Confirmatory factor analysis of the Pain Catastrophizing Scale in African American and Caucasian Workers' Compensation claimants with low back injuries. AB - Pain catastrophizing is an important cognitive construct that has been linked with many aspects of the pain experience, including pain intensity, emotional distress, pain-related disability, and pain behavior. The Pain Catastrophizing Scale (PCS), an instrument often used to assess this construct, reflects three aspects of catastrophizing: Rumination, Magnification, and Helplessness. Its factor structure, however, has never been examined in clinical samples of persons with occupational injury or as a function of race. In this study, exploratory and confirmatory factor analyses were used to examine the factor structure of the PCS in a large, racially diverse sample of Workers' Compensation claimants with low back injuries. Results indicated that a two-factor model of the PCS (Rumination and 'Powerlessness,' the latter a combination of the PCS Magnification and Helplessness scores) was the most parsimonious fit to the data, particularly in the African American sample. Future research in other clinical samples that include African Americans is needed to examine the stability of the results reported here. PMID- 15661445 TI - Intravenous dextromethorphan to human volunteers: relationship between pharmacokinetics and anti-hyperalgesic effect. AB - The aim of this study was to investigate the effect of dextromethorphan (DM) 0.5 mg/kg administered intravenously (i.v.) on hyperalgesia and pain after a tissue injury in human volunteers, and to describe the relationship between pharmacokinetic and pharmacodynamic data. The heat-capsaicin sensitisation model, a well-established experimental hyperalgesia model was induced in 24 healthy, male volunteers aged 21-35 years. The subjects received i.v. DM 0.5 mg/kg or isotonic saline on two separate study sessions. The primary outcome measure from 0 to 3 h was reduction in area of established secondary hyperalgesia. Secondary outcome measures were reduction in area of secondary hyperalgesia in response to brief thermal stimulation, heat pain detection thresholds and painfulness after tonic heat pain. Blood samples were collected throughout the study to describe the relationship between pharmacokinetic and pharmacodynamic data. Intravenous DM 0.5 mg/kg significantly reduced areas of established secondary hyperalgesia with an average of 39% (P<0.05). Development of secondary hyperalgesia was substantially prevented by DM (P<0.05). No significant effect was seen on either heat pain detection thresholds or after tonic heat pain. The pharmacokinetic pharmacodynamic relationship showed a large inter-subject variation with a mean delay in effect of nearly 2 h in relation to peak serum concentration. The results strongly indicate that DM is an anti-hyperalgesic drug. The delay in effect may be explained by several mechanisms and suggests that timing of DM administration is an essential factor for using the drug in clinical settings. PMID- 15661447 TI - Intrathecal morphine and ketorolac analgesia after surgery: comparison of spontaneous and elicited responses in rats. AB - Pain after surgery results in significant morbidity, and systemic opioids often fail to provide adequate analgesia without marked sedation and respiratory depression. Intrathecal morphine provides better analgesia, but is limited by delayed respiratory depression. Intrathecal injection of the cyclooxygenase inhibitor, ketorolac, has recently entered clinical trials, and the current study examined the interaction between intrathecal morphine and ketorolac to treat postoperative pain. We also sought to compare these treatments on a commonly used assessment of withdrawal threshold and a new assessment of spontaneous behavior after surgery. Male Sprague Dawley rats and underwent hind paw incision or subcostal laparotomy surgery. Intrathecal morphine, ketorolac, or their combination were injected on the first postoperative day, with outcome measure being return to pre-surgery withdrawal threshold with von Frey filament testing of the paw after paw incision, or return to pre-surgery exploratory activity after laparotomy. Intrathecal morphine completely reversed the effects of surgery in both models, but intrathecal ketorolac only partially reversed them. Ketorolac enhanced the potency of morphine several fold in both models, and did so synergistically after paw incision. In all cases drug potency was greater for spontaneous than elicited responses. These data confirm that spinal opioid receptor and cyclooxygenase enzyme inhibition diminish elicited tactile hypersensitivity after surgery, and that they similarly return spontaneous behavior to normal. Differences in drug potency could reflect fundamental differences in outcome measures or in the surgical procedures themselves. These data support combination study of intrathecal morphine and ketorolac for postoperative pain. PMID- 15661448 TI - Medium and large injured dorsal root ganglion cells increase TRPV-1, accompanied by increased alpha2C-adrenoceptor co-expression and functional inhibition by clonidine. AB - Some electrophysiologic studies demonstrate new, excitatory alpha2-adrenoceptors on peripheral nociceptors and their dorsal root ganglion (DRG) cell bodies after nerve injury, yet administration of alpha2-adrenoceptor agonists at these sites reduces hypersensitivity rather than worsens it. Since TRPV-1 expressing nociceptor afferents are important in many pain states, we examined the expression of this channel and its co-expression with alpha2C-adrenoceptors in injured DRG cell bodies and the ability of alpha2-adrenoceptors to inhibit responses to stimulation. Rats underwent tight ligation of the left L5 and L6 spinal nerves, followed by behavioral testing, removal of L5 and L6 DRGs, and either immunostaining for TRPV-1 channels and alpha2C-adrenoceptors or intracellular calcium videomicroscopy in response to electrical field stimulation before and after perfusion with clonidine and capsaicin. Spinal nerve ligation produced tactile allodynia. In normal and sham controls, about one-third of DRG neurons were TRPV-1-immunoreactive (IR), one half were alpha2C-adrenoceptor-IR and one-fourth co-expressed both. After nerve ligation there was a reduction in the number of small, strongly TRPV-1-IR or alpha2C-adrenoceptor-IR neurons, but an increase in medium and large, lightly stained cells and in their co expression. The proportion of clonidine inhibited cells which responded to capsaicin increased 5 fold after injury. We conclude that TRPV-1 and alpha2C adrenoceptors are up-regulated in some injured medium and large size neurons after nerve ligation. Increased co-expression by immunocytochemistry, and increased proportion of cells inhibited by clonidine and expressing functional TRPV-1 channels suggest that these cells may play an important role in the analgesic effects of alpha2-adrenoceptor agonists in neuropathic pain. PMID- 15661449 TI - Increased nociceptive response in mice lacking the adenosine A1 receptor. AB - The role of the adenosine A1 receptor in nociception was assessed using mice lacking the A1 receptor (A1R-/-) and in rats. Under normal conditions, the A1R-/- mice exhibited moderate heat hyperalgesia in comparison to the wild-type mice (A1R+/+). The mechanical and cold sensitivity were unchanged. The antinociceptive effect of morphine given intrathecally (i.t.), but not systemically, was reduced in A1R-/- mice and this reduction in the spinal effect of morphine was not associated with a decrease in binding of the mu-opioid ligand DAMGO in the spinal cord. A1R-/- mice also exhibited hypersensitivity to heat, but not mechanical stimuli, after localized inflammation induced by carrageenan. In mice with photochemically induced partial sciatic nerve injury, the neuropathic pain-like behavioral response to heat or cold stimulation were significantly increased in the A1R-/-mice. Peripheral nerve injury did not change the level of adenosine A1 receptor in the dorsal spinal cord in rats and i.t. administration of R-PIA effectively alleviated pain-like behaviors after partial nerve injury in rats and in C57/BL/6 mice. Taken together, these data suggest that the adenosine A1 receptor plays a physiological role in inhibiting nociceptive input at the spinal level in mice. The C-fiber input mediating noxious heat is inhibited more than other inputs. A1 receptors also contribute to the antinociceptive effect of spinal morphine. Selective A1 receptor agonists may be tested clinically as analgesics, particularly under conditions of neuropathic pain. PMID- 15661451 TI - Do high TSH values protect against chronic musculoskeletal complaints? The Nord Trondelag Health Study (HUNT). AB - The aim of this large cross-sectional population-based study was to examine a possible positive or negative association between thyroid dysfunction and chronic musculoskeletal complaints (MSC). Between 1995 and 97, all 94,197 adults in Nord Trondelag County in Norway were invited to participate in a health survey. A total of 64,787 (69%) responded to questions related to MSC, whereof thyroid stimulating hormone (TSH) was measured in 34,960 individuals. These included a 5% random sample of women and men 20-40 years of age (n=2165), nearly all women above 40 (n=19,308), a random sample which included 50% of men older than 40 years (n=9983), and 3504 (97%) with self-reported thyroid dysfunction. Among the 64,787 participants, 30,158 (47%) who reported MSC continuously for at least 3 months during the past year where defined as having chronic MSC. Associations between thyroid dysfunction and chronic MSC were assessed in multivariate analyses, estimating prevalence odds ratios (ORs) with 95% confidence intervals (CIs). High TSH values were associated with lower prevalence of chronic MSC at ten anatomical sites among women with no history of thyroid dysfunction. Among these, chronic MSC was less likely (OR=0.6, 95% CI 0.4-0.8) if TSH >or=10 mU/L than in women with normal TSH (0.2-4 mU/L). Chronic MSC was less likely among women with high TSH values. The mechanism is unclear and, theoretically, may reflect a fundamental gender-specific relationship between TSH and pain perception in the central nervous system. PMID- 15661450 TI - Activation of 5-HT1A and 5-HT7 receptors in the parafascicular nucleus suppresses the affective reaction of rats to noxious stimulation. AB - The antinociceptive effects of the serotonin (5-HT)1A/7 receptor agonist 8 hydroxy-dipropylaminotetralin (8-OH-DPAT) administered into the medial thalamus were evaluated. Pain behaviors organized at spinal (spinal motor reflexes, SMRs), medullary (vocalizations during shock, VDSs), and forebrain (vocalization after discharges, VADs) levels of the neuraxis were elicited by tailshock. Administration of 8-OH-DPAT (5, 10, and 20 microg/side) into nucleus parafascicularis (nPf) produced dose-dependent increases in VDS and VAD thresholds, but failed to elevate SMR threshold. The increase in VAD threshold was significantly greater than that of VDS threshold. Similar effects were observed with administration of 8-OH-DPAT (20 microg/side) into the rostral portion of the central lateral thalamic nucleus. The bilateral or unilateral administration of 8-OH-DPAT (20 microg) into other thalamic nuclei, or into sites dorsal to nPf, did not elevate vocalization thresholds. Increases in vocalization thresholds produced by nPf-administered 8-OH-DPAT were mediated by both 5-HT1A and 5-HT7 receptors. Intra-nPf administration of the 5-HT1A receptor antagonist WAY-100635 (0.05 or 0.5 microg/side), or the 5-HT7 receptor antagonist SB-269970 (1 or 2 microg/side), but not the dopamine D2 receptor antagonist raclopride (10 microg/side), reversed 8-OH-DPAT induced elevations in vocalization thresholds. These results provide the first reported evidence of behavioral antinociception following the administration of a 5-HT agonist into the medial thalamus. PMID- 15661453 TI - Extensive abscess formation after repeated paravertebral injections for the treatment of chronic back pain. AB - Paravertebral injections are commonly used for the treatment of chronic back pain despite the lack of evidence for their efficacy. Here, we report a case in which repeated paravertebral injections of local anesthetics, corticosteroids and botulinum toxin resulted in extensive abscess formation involving the entire paravertebral musculature descending to the level of the midthighs. These abscesses induced a systemic inflammatory response syndrome and required surgical drainage as well as broad-spectrum antibiotic therapy. PMID- 15661452 TI - SUNCT: bilateral hypothalamic activation during headache attacks and resolving of symptoms after trigeminal decompression. AB - Short-lasting unilateral neuralgiform headache with conjunctival injection and tearing (SUNCT) is a primary head-pain syndrome, which is often refractory to any medical treatment. Concerning the pathophysiology of SUNCT, hypothalamic involvement ipsilaterally to the pain has been suggested based on the clinical features and one functional imaging case report. Here we now report a new case with SUNCT and the concomitant cerebral activation pattern (fMRI) during the pain attacks. In addition to an activation of several brain structures known to be generally involved in pain processing, bilateral hypothalamic activation occurred during the pain attacks, arguing for a central origin of the headache. Interestingly, this patient became completely pain free after surgical decompression of the ipsilateral trigeminal nerve. We hypothesize that in this case with a central predisposition for trigeminal autonomic cephalgias, a peripheral trigger with ectopic excitation might have contributed to the clinical picture of SUNCT. PMID- 15661454 TI - Pain. PMID- 15661455 TI - [Management of a medical disaster or cosmetic actions]. PMID- 15661456 TI - [Which place for the anaesthesiologist/intensivist in the surgical ward: a folding seat or an orchestra stall?]. PMID- 15661457 TI - [Governance]. PMID- 15661458 TI - [Iodine allergy: myth or reality?]. PMID- 15661459 TI - [Analgesic effect of continuous intravenous nefopam after urological surgery]. AB - OBJECTIVES: To evaluate the efficacy of continuous infusion of nefopam. Indeed this analgesic is commonly used by continuous infusion by many anaesthetists to reduce its adverse effects. However whether the analgesic effect of an intermittent administration of nefopam has been proven, the efficacy of continuous infusion has not been established. STUDY DESIGN: Double-blind placebo controlled prospective randomised study. PATIENTS AND METHODS: Sixty patients ASA 1 to 3 undergoing planned urological surgery with laparotomy were included. At the end of surgery, bolus doses of placebo (Group 3) or nefopam 20 mg (Group 1 and 2) were administered to all the patients. Placebo (Group 3), nefopam 80 mg (Group 1) or 120 mg (Group 2) was thereafter continuously infused over 24 hours. All patients received additional analgesia with PCA morphine. We measured pain at rest and on cough with VAS. Adverse side effects such as nausea and vomiting, sedation and respiratory depression were evaluated. Mental performance was measured with mini mental status tests. RESULTS: Patients were older in the placebo group by approximately six years but anesthetic and surgical variables were not different between groups. Pain at rest and on cough was not statistically different between groups. In the placebo group, the median (interquartile range) morphine consumption reached 29 mg (13-53) whereas in patients receiving 80 and 120 mg nefopam, it levelled to 44 mg (11-54) and 35 mg (9-82) respectively (p > 0.05). Patients needed morphine during the same time period whether they received nefopam or not. Patients suffering from adverse effects were similar between groups. CONCLUSION: In this study, continuous administration of nefopam did not reduce morphine consumption nor ameliorate analgesia and thus may not be recommended in urological surgery. Nefopam pharmacokinetics when used with continuous infusion as well as surgery types and differences in age between groups may explain these results. PMID- 15661460 TI - [Surgical antimicrobial prophylaxis: compliance to guidelines and impact of targeted information program]. AB - OBJECTIVES: Surgical antimicrobial prophylaxis is used to decrease postoperative wound infection. We assessed the compliance to surgical antimicrobial prophylaxis guidelines in our hospital and the impact of an information program. PATIENTS AND METHODS: Observational study of clean or clean contamined surgery, during two 3 week periods, separated by a targeted information period. The following data were collected prospectively: prophylaxis indication, antimicrobial agent, timing, dose, route and duration of prophylaxis. Chi square test was used for analysis or Fischer test when available. RESULTS: Four hundred seventy-seven patients were enrolled - 270 and 207 for each period respectively. For both periods only 49% of prophylaxis was appropriated. When prophylaxis should be administered - 15 and 13% of patients for each period - it was antibioprophylaxy was strictly adequate with recommandations. The most common error was administration timing. Only the choice of antimicrobial agent was optimized after information period. DISCUSSION: These results are consistent with previous studies. Information program alone have no effect on the good use of antimicrobial for surgical prophylaxis. Only a policy associating organization, restriction and education could improve practices. PMID- 15661461 TI - [Complications during the intrahospital transport in critically ill patients]. AB - INTRODUCTION: Intrahospital transport (IHT) of mechanically ventilated critically ill patients is associated with an important risk of complications. OBJECTIVES: The purpose of this study was to assess the incidence of complications occurring during the IHT and to analyse the causes and the consequences of such complications. PATIENTS AND METHODS: All the IHT performed in mechanically ventilated patients, hospitalised in medical and surgical intensive care units of a university hospital were prospectively included during a three-month period. Complications were defined as follows: patient related problems (desaturation, restlessness, haemodynamic instability, extubation) and ventilator related problems (breakdown or defect of the material). RESULTS: 123 IHT concerning 64 patients were analysed, with 64 IHT were realised for diagnostic procedure (computed tomography) and 59 for therapeutic procedure (surgical procedure or interventional radiology). At least one patient related problem occurred during 41 IHT (33%) (desaturation n = 11, agitation n = 21, haemodynamic alterations n = 19, extubation n = 0). In two patients, these complications led to cardiac arrest. Patient related problems were observed more frequently in sedated patient (66 %, p = 0.0001) as well as during IHT for diagnostic procedure (p = 0.03). A ventilator problem occurred in 26 transports (21 %) and was more frequently reported when a turbine ventilator was used (p = 0.0056). CONCLUSION: This study supports the fact that IHT of mechanically ventilated critically ill patients, is a high-risk procedure associated with potentially severe complications. This finding emphasises the need of standardised procedures and medical surveillance during IHT. PMID- 15661462 TI - [Peripheral venous placement: evaluation of guidelines respect in prehospital setting]. AB - INTRODUCTION: Respect of guidelines for peripheral venous placement has never been studied in prehospital setting. OBJECTIVE: Describe prehospital practice in peripheral venous placement and perform comparison with guidelines. MATERIAL AND METHODS: Prospective study, in mobile intensive care unit from a university hospital, October 2002 to January 2003. Study of prehospital practice for peripheral venous placement in prehospital setting and comparison with guidelines. Guideline from Anaes: wearing a cap and a mask, washing hands, washing insertion site with water and soap, wearing sterile gloves, using sterile gauze, applying iodine solution to the insertion site, using catheter with a protective system and applying occlusive dressing. RESULTS: Six hundred seventy three reports were studied. A cap and a mask wearing and washing insertion site with water and soap were never respected. Washing hand with antiseptic was reported in 56% of the cases, washing insertion site with iodine solution in 4%, wearing sterile gloves in 21%, using catheter with a protective system in 89%, using sterile gauze in 1% and applying occlusive dressing in 99%. For each criterion, respect of guidelines was significantly more frequent in nurse practice than in physician practice. CONCLUSION: Guidelines for peripheral venous placement were not respected in prehospital setting. Education and formation are necessary to optimize infectious risk prevention in prehospital care. PMID- 15661463 TI - [Intensive care management of patients with severe head traumatism in Benin from 1998 to 2002]. AB - OBJECTIVE: To study in Cotonou (Benin) in tropical environment, the specific problems of severe head injury. STUDY DESIGN: Five years retrospective study in Cotonou University National and Teaching Hospital. PATIENTS AND METHODS: All patients admitted with severe head injury between January 1998 and December 2002 were included. The following parameters were studied: age, sex, traumatism cause, temperature, blood pressure, Glasgow coma scale. RESULTS: Two Hundred thirty six patients (mean age 32 +/- 18 years) essentially men, were admitted. The cause of the injury was road traffic accidents in 86% of cases. Prehospital medicalisation was achieved in only 9% of cases. A head CT-scan was performed in 5% of the cases. Respiratory resuscitation essentially used the association intubation and oxygenation (59%), 19% of the patients were ventilated and 21% were sedated. Mortality was closely related to the initial Glasgow coma scale score (GCS): GCS 3-4 = 97%; GCS 5-6 = 72% GCS 7-8 = 36%. Overall mortality was 70%. CONCLUSION: Severe head injury is a major cause of morbidity and mortality in developing countries. Limited access to highly specialised care explain a high mortality rate. An active policy of prevention from administrative authorities and a greater effectiveness of medical management may improve outcome. PMID- 15661465 TI - [Clinical governance]. AB - The direction of health organisations requires the coordination of diversified competencies, notably in administration, management and care. In France, for university hospitals, the law (decree) of 1958 is pivotal. Other approaches have been developed in other countries, depending on their history, traditions, and the organisation of their respective health care systems. These models can enlighten our current reflexions at a time when in France, the question of a new definition of governance for university hospitals is raised. PMID- 15661464 TI - ["Iodine allergy": point of view]. AB - OBJECTIVE: The aim of this literature review is to suggest a diagnostic and a preventive attitude in patients having presented an immediate hypersensitivity reaction due to an iodinated drug. DATA SOURCES: Literature review. Data were searched in the Medline database from 1967 to 2004 in English and French language. Complementary references were selected from the bibliography of selected references or from authors' personal databases. The following key-words were used separately or combined: Hypersensitivity, Immediate; Allergy; Contrast Media; Povidone-Iodine; Iodine; Iodine Compounds; Iodides; Amiodarone; Seafood, Parvalbumins; Tropomyosin. STUDY SELECTION: Randomized studies, epidemiological studies, original articles, clinical cases, and letters to the editor were selected. DATA SYNTHESIS: The implication of iodine has never been demonstrated during allergic hypersensitivity reactions due to iodinated drugs. However, IgE mediated allergic hypersensitivity reactions have been published with contrast media or iodinated antiseptics and will be described in this development. In a wider sense, allergic hypersensitivity reactions due to seafood are evoked because often improperly considered as a risk factor of allergic reaction to iodinated drugs. The allergenic determinant responsible of patient sensitization is not known for iodinated contrast media, but is probably due to povidone in case of iodine povidone. In fish, the allergen is described as the protein M. There has also been strong immunological evidence that tropomyosin is a cross reactive allergen among crustaceans and molluscs (shellfishs). In case of hypersensitivity reaction occurring with iodinated drug, an allergological assessment is required to confirm the immune mechanism, to identify the culprit drug or substance and to identify cross-reactivity especially with iodinated contrast media. CONCLUSION: Asking a patient if he/she is "allergic to iodine" is a question that should be avoided because its significance is null. A diagnosis of drug allergy, essentially relying on clinical symptoms, biological tests and cutaneous tests, is required to take adequate preventive measures. PMID- 15661466 TI - [Diagnosis and management of a haemorrhagic emergency revealing an acquired haemophilia]. AB - We relate the case of a 22-year-old woman, without any particular medical history, apart from a previous full-term pregnancy with no complication, for whom the discovery of prolongated APTT (Activated Partial Thromboplastin Time) postponed an emergency operation in spite of intracystic ovarian haemorrhage. The complementary check-up revealed the existence of an autoantibody antifactor VIII (antithromboplastinogen), showing acquired postpartum haemophilia. PMID- 15661468 TI - [Involvement of anaesthesiologists in surgery offices: point of view of anaesthesiologists residents from Lyon]. PMID- 15661467 TI - [Cerebral gas embolism after a cough fit]. AB - An 81-year-old man, with a history of pulmonary tuberculosis presented with coma after coughing associated to haemoptysis. The CT-scan revealed several scattered gas bubbles, while the thoracic scan disclosed an old tubercular parenchymatous cave. In spite of several sessions of hyperbaric oxygen treatment, the patient died with multivisceral organ failure. This case report reviews the pathophysiology of gas embolism due to coughing. PMID- 15661469 TI - [Anaphylactic shock and uterine atony: must we use sulprostone]. PMID- 15661470 TI - Diagnostic lumbar puncture in the parturient with history of previous post-lumbar puncture headache. PMID- 15661471 TI - The passing of an era and the birth of a new one. PMID- 15661472 TI - Nutritional disorders in Africa: the triple burden. PMID- 15661473 TI - Secondary anthropometric data analysis of the National Food Consumption Survey in South Africa: the double burden. AB - OBJECTIVE: There is an increase in the prevalence of overweight and obesity in children worldwide, including South Africa. We investigated the prevalences of overweight, obesity, and stunting in a current generation of children (ages 12 to 108 mo), which has a high prevalence of stunting, and evaluated the determinants of both nutritional disorders. METHODS: Secondary data analysis of the weight and height measurements of 12- to 108-mo-old children (weighted n = 2200, non weighted n = 2894) during the 1999 National Food Consumption Survey in South Africa is reported. The body mass index reference percentiles recommended for use in children by the International Obesity Task Force were used to determine the prevalence of overweight and obesity, and the National Center for Health Statistics (NCHS) percentiles were used to determine the prevalence of stunting. RESULTS: Nationally, the prevalence of stunting (height-for-age < or = -2 standard deviations, NCHS 50th percentile) in these children was 19.3% (95% confidence interval [CI] = 17.49 to 21.16) and was highest in 1- to 3-y-old children (24.4%) and in children of farm workers on commercial farms (25.6%). The prevalence of combined overweight and obesity (body mass index > or = 25 kg/m(2) in 17.1%, 95% CI = 15.00 to 19.23) at the national level was nearly as high as that for stunting. Further, the types of determinants for stunting and overweight were generally similar (although directionally opposite in degree of risk conferred) and included type of housing, type of toilet in the home, fuel used in cooking, presence of a refrigerator or stove, presence of a television in the house, educational level of the caregiver, and maternal education level. An example of the directionally opposite degree of risk is exemplified by the use of paraffin as a fuel being protective against being overweight (odds ratio = 0.78, 95% CI = 0.63 to 0.97) but predictive of an increased risk for stunting (odds ratio = 1.24, 95% CI = 1.04 to 1.48). Stunting itself conferred an increased risk (odds ratio = 1.80, 95% CI = 1.48 to 2.20) of being overweight. CONCLUSION: Certain defined determinants appear to play important roles in children's nutritional outcomes in relation to stunting and to overweight and obesity. PMID- 15661474 TI - Iodine status as determined by urinary iodine excretion in Lesotho two years after introducing legislation on universal salt iodization. AB - OBJECTIVE: Mild to severe iodine deficiency has been documented in Lesotho since 1960. To eliminate the persisting iodine deficiency, legislation on universal salt iodization was introduced in 2000 as a long-term public health intervention strategy. We assessed the urinary iodine status of school children and women of child-bearing age in Lesotho 2 y after the introduction of legislation on universal salt iodization. METHODS: A 31-cluster national survey was conducted in 2002 by using the proportion to population size method. In each cluster, 30 women ages 15 to 30 y and 30 primary school children ages 8 to 12 y were randomly selected to provide urine samples for urinary iodine analysis. Data were interpreted according to criteria of the World Health Organization, United Nations Children's Fund, and International Council for Control of Iodine Deficiency Disorders (2001). Statistical analysis was performed using SAS. RESULTS: We analyzed 912 urine samples from children and 924 from women. The median urinary iodine concentrations were 214.7 microg/L for children and 280.1 microg/L for women, indicating more than adequate iodine intake. Median iodine concentration was higher in the lowlands (256.0 microg/L in children and 329.9 microg/L in women) than in the mountains (99.30 microg/L in children and 182.6 microg/L in women). Analysis of the distribution of the data showed values below 50 microg/L in 10.1% of children and in 9.8% of women. In addition, 21.5% of children and 17.9% of women had urinary iodine excretion values below 100 microg/L. In contrast, 36% of children and 47.2% of women had urinary iodine concentrations in excess of 300 microg/L. CONCLUSION: Results of urinary iodine excretion measurements indicated that iodine deficiency has been eliminated as a public health problem in Lesotho. However, the high median urinary iodine concentration of women in the lowlands indicated the possibility of a risk of iodine-induced hyperthyroidism in vulnerable people. PMID- 15661475 TI - Effect of periodic vitamin A supplementation on mortality and morbidity of human immunodeficiency virus-infected children in Uganda: A controlled clinical trial. AB - OBJECTIVE: We investigated whether vitamin A supplementation would decrease mortality and morbidity rates in children infected with the human immunodeficiency virus (HIV). METHODS: We conducted a randomized, double-blind, placebo-controlled clinical trial at Mulago Hospital, a large hospital that serves the urban and semiurban populations of Kampala, Uganda. One hundred eighty one HIV-infected children were enrolled at 6 mo and randomized to receive vitamin A supplementation, 60 mg retinol equivalent, or placebo every 3 mo from ages 15 to 36 mo. Morbidity was assessed through a 7-d morbidity history every 3 mo, and vital events were measured. Children received daily trimethoprim-sulfamethoxazole prophylactic therapy. RESULTS: After age 15 mo, children were followed for a median of 17.8 mo (interquartile range = 11.1 to 21.0 mo). The trial was stopped when there was a new policy to implement a program of mass supplementation of vitamin A in the country. Mortality rates among 87 children in the vitamin A group and 94 children in the control group were 20.6% and 32.9%, respectively, yielding a relative risk of 0.54 (95% confidence interval, 0.30 to 0.98; P = 0.044) after adjusting for baseline weight-for-height Z score. Children who received vitamin A had lower modified point prevalences of persistent cough (odds ratio, 0.47; 95% confidence interval, 0.23 to 0.96; P = 0.038) and chronic diarrhea (odds ratio, 0.48; 95% confidence interval, 0.19 to 1.18; P = 0.11) and a shorter duration of ear discharge (P = 0.03). Vitamin A supplementation had no significant effect on modified point prevalences of fever, ear discharge, bloody stools, or hospitalizations. CONCLUSIONS: Vitamin A supplementation decreases mortality rate in HIV-infected children and should be considered in the care for these children in developing countries. PMID- 15661476 TI - Nutritional status and birth outcomes of adolescent pregnant girls in Morogoro, Coast, and Dar es Salaam regions, Tanzania. AB - OBJECTIVE: Studies that link adolescence pregnancies, nutritional status, and birth outcomes in Tanzania are scarce. We examined the nutritional status and birth outcomes of pregnant adolescent girls from rural and urban areas of three regions in Tanzania. METHODS: The study was carried out in the regions of Dar es Salaam (Chamazi and Gezaulole dispensaries and Round Table Maternity Home), Coast (Tumbi Regional Hospital and Mlandizi Health Center), and Morogoro (Regional Hospital, Uhuru Clinic, and Mlali Health Center). One hundred eighty pregnant adolescent girls ages 15 to 19 y were recruited and interviewed, and their nutritional status measurements were taken at the seven health facilities. Information concerning date of birth, marital status, educational status, sex education, and income status was collected with a structured questionnaire. Height, weight, and mid-upper arm circumference were measured according to standard techniques. Hemoglobin concentration was measured with a hemoglobinometer and the HemoCue technique. Nutritional status was assessed by body mass index, and hemoglobin concentration was determined by cutoff points of the World Health Organization. Suitable statistical analysis was done with SPSS 9.0. Weekly weight gain during pregnancy was measured in 123 subjects who kept their appointments and reported back after 2 wk. Fifty-seven subjects did not keep their appointments and were lost to follow-up. Records of infants' birth weights and mode of delivery were obtained from 50 subjects who delivered at the study sites. RESULTS: The height of about 54% of the subjects was shorter than 151 cm, suggestive of short maternal height. Severe wasting was observed in 27% of subjects. Mean weekly weight gain during pregnancy was 317 +/-110 g (-500 to 500 g). No significant differences were observed between rural and urban settings. Mean infant birth weight was 2600 +/- 480 g. About 48% of infants had low birth weight (<2500 g) and only 14% of infants had birth weight greater than 3000 g. About 14% of infants were born by cesarean section. Nearly 86% of the pregnant adolescent girls were anemic. A hemoglobin concentration below 7 g/dL was observed in 5% of subjects. Most subjects (55%) had hemoglobin concentrations from 7 to lower than 10 g/dL. There was a weak correlation between infant birth weight and weekly weight gain of the girls during pregnancy (r = 0.36, P < or = 0.01). However, a strong correlation was observed between birth weight and hemoglobin level of adolescent girls during pregnancy (r = 0.67, P = 0.01). Short stature was observed to contribute toward cesarean delivery (P = 0.05) because more cesarean deliveries were performed in short girls (<151 cm tall). CONCLUSIONS: The nutritional status of pregnant adolescent girls in the study areas was poor and resulted in poor pregnancy outcome. Girls should be educated about reproductive health at the primary level of education. PMID- 15661477 TI - Diet and blood pressure in South Africa: Intake of foods containing sodium, potassium, calcium, and magnesium in three ethnic groups. AB - OBJECTIVES: We investigated whether habitual intakes of sodium (Na), potassium, magnesium, and calcium differ across South African ethnic groups, assessed the proportion of Na intake, which is discretionary, and identified which food sources were the major contributors to Na intake. METHODS: This was a cross sectional study of 325 black, white, and mixed ancestry hypertensive and normotensive subjects. Three repeated 24-h urine samples were collected for assessment of urinary Na, and three corresponding 24-h dietary recalls were administered by trained fieldworkers. Blood pressure and weight were measured at each visit. Secondary analyses were performed on existing dietary databases obtained from four regional surveys undertaken in South African adults. RESULTS: Mean urinary Na excretion values equated to daily salt (NaCl) intakes of 7.8, 8.5, and 9.5 g in black, mixed ancestry, and white subjects, respectively (P < 0.05). Between 33% and 46% of total Na intake was discretionary, and, of the non discretionary sources, bread was the single greatest contributor to Na intake in all groups. Ethnic differences in calcium intake were evident, with black subjects having particularly low intakes. Urban versus rural differences existed with respect to sources of dietary Na, with greater than 70% of total non discretionary Na being provided by bread and cereals in rural black South Africans compared with 49% to 54% in urban dwellers. CONCLUSION: White South Africans have higher habitual intakes of Na, but also higher calcium intakes, than their black and mixed ancestry counterparts. All ethnic groups had Na intakes in excess of 6 g/d of salt, whereas potassium intakes in all groups were below the recommended level of 90 mM/d. Dietary differences may contribute to ethnically related differences in blood pressure. PMID- 15661478 TI - Development and validation of a questionnaire to test knowledge and practices of dietitians regarding dietary supplements. AB - OBJECTIVE: We developed a questionnaire to evaluate dietitians' knowledge and practices about the use of dietary supplements in light of new draft regulations in South Africa that will allow dietitians to prescribe supplements for the management of nutrition-related diseases. METHODS: A cross-sectional validation study was carried out in two phases: 1) questionnaire planning and development and 2) determination of reliability (internal consistency). The study sample comprised 48 nurses, 105 dietetic interns, and 367 registered dietitians. An expert academic group comprising nine dietitians with research and teaching experience rated the academic relevance and importance of knowledge questions in an item pool of 355 questions to ensure face and content validity. Criterion validity was measured by testing the knowledge questions (KQs) on 48 nurses and 105 dietetic interns. Thereafter questions were deleted based on their difficulty, discrimination, and Cronbach's alpha values. The final questionnaire, which at this point included a test of 50 KQs, was mailed to registered dietitians (n = 1450) in South Africa to determine internal consistency of the final KQs. RESULTS: Criterion validity of the KQs was indicated by a significant difference (P = 0.000) in the mean test scores between nurses (36.9, 95% confidence interval 24.2 to 39.6) and dietetic interns (40.2, 95% confidence interval 38.8 to 41.7). Item discrimination was attained by the deletion of NQs having a minimum correlation of 0.20 with overall test score. Further, item difficulty was controlled for by removal of questions correctly answered by more than 80% (too easy) or fewer than 20% (too difficult). Internal consistency of the KQs, after completion by 367 dietitians, was 0.87 (Cronbach's alpha). The mean knowledge score of dietitians ranged from 56.5% to 62.5%, with the oldest group (> or =40 y) having the highest score (P = 0.018). Thirty-eight percent of dietitians recommended dietary supplements at least 3 times a week; 51% consumed dietary supplements themselves; and 17% sold them to clients. CONCLUSIONS: A valid and reliable questionnaire that tests knowledge and practices of dietary supplements was developed for dietitians. This tool can be used by academic institutions to evaluate the knowledge and practices of final-year students. It can also be used on a representative sample of dietitians in South Africa and possibly elsewhere within the profession. PMID- 15661479 TI - Nutritional status and functional ability of the elderly aged 60 to 90 years in the Mpigi district of central Uganda. AB - OBJECTIVES: This study assessed the nutritional status of the elderly and their functional ability because poor nutritional status in the elderly is associated with poor functional ability. METHODS: Anthropometric measurements, demographic and socioeconomic data, dietary assessment by a food frequency list, and activities of daily life data were collected cross sectionally in 2002. Participants (n = 100) were randomly selected and the response rate was 95.2%. RESULTS: The overall prevalences of undernutrition were 33.3% based on body mass index (<18.5 kg/m(2)) and 52% based on mid-upper arm circumference (<24 cm). There was a large, significant difference between prevalences of malnutrition by sex: 68% of women were undernourished (body mass index < 18.5 kg/m(2)) compared with 32.4% of men. Dietary assessment showed that intake of fish, cereals, vegetables, tubers, and legumes was moderate (three to six times/wk). Evaluation of the ability of elderly people to perform basic activities of daily living showed that 33% of subjects were independent in all activities of daily living, except for mobility and feeding. The relation between body mass index and variables associated with functional ability were significant with regard to mobility, continence, and feeding (P < 0.05). CONCLUSION: This study found that a large percentage of older men and women are malnourished. This influenced their daily activities, especially mobility and feeding. The elderly need to be incorporated into health programs and policy. PMID- 15661480 TI - Substitution of high monounsaturated fatty acid avocado for mixed dietary fats during an energy-restricted diet: effects on weight loss, serum lipids, fibrinogen, and vascular function. AB - OBJECTIVE: First, we wanted to dispel the myth that avocados are fattening and therefore should be avoided in energy-restricted diets. Second, we examined the effects of avocados, a rich source of monounsaturated fatty acids, as part of an energy-restricted diet on weight loss, serum lipids, fibrinogen, and vascular function in overweight and obese subjects. METHODS: Sixty-one free-living volunteers (13 men and 48 women), with body mass index of 32 +/- 3.9 kg/m(2) (mean +/- standard deviation) participated in this randomized, controlled, parallel study. Subjects were paired and randomly assigned to one of two groups. The experimental group consumed 200 g/d of avocado (30.6 g of fat), which substituted for 30 g of other mixed dietary fats such as margarine or oil, and the control group excluded avocado from their energy-restricted diet for 6 wk. Seven-day isoenergetic menus were planned according to mean energy requirements of both sexes to provide total energy intakes consisting of 30% fat, 55% carbohydrates, and 15% protein. Anthropometric measurements, physical activity, blood pressure, and arterial compliance were measured with standard methods at the beginning and end of the intervention. Fasting blood samples were drawn at the beginning and end of the intervention. RESULTS: Fifty-five subjects completed the study. The compliance rate to avocado intake in the experimental group was 94.6%. The percentage of plasma oleic acid increased significantly with the consumption of avocado in the experimental group, whereas a decrease was seen in the percentage of myristic acid from baseline to the end of the intervention in both groups but was significant only in the experimental group. Anthropometric measurements (body mass, body mass index, and percentage of body fat) decreased significantly in both groups during the study (P < 0.001), and the change was similar in both groups. Serum lipid concentrations (total cholesterol, low density lipoprotein cholesterol, high-density lipoprotein cholesterol, and triacylglycerols), fibrinogen, blood pressure, and arterial compliance did not change significantly within or between groups. CONCLUSION: The consumption of 200 g/d of avocado within an energy-restricted diet does not compromise weight loss when substituted for 30 g of mixed dietary fat. Serum lipid concentrations, plasma fibrinogen, arterial compliance, and systolic and diastolic blood pressures were not affected by weight loss or avocado intake. PMID- 15661481 TI - Development of a reliable and valid nutritional knowledge questionnaire for urban South African adolescents. AB - OBJECTIVES: We wanted to develop a valid and reliable nutritional knowledge test for urban South African adolescents who were participating in the Birth-to-Twenty cohort study. The questionnaire was intended for use every second year, from ages 13 to 14 y until age 20 y. METHODS: The initial steps involved the development of a conceptual framework and identification of nutritional concepts in collaboration with nutritional experts, and this defined the construct of the questionnaire. The South African national teaching curriculum objectives for nutritional education and other relevant issues were selected as the desired concepts, and most items were phrased in accordance with the recently developed South African Food-Based Dietary Guidelines. Thereafter, 140 items (questions) were developed and in turn assessed by an expert panel, and the result was that only 88 items remained. This was done to ensure content and face validity of the items. The 88 items were constructed into a questionnaire and piloted for appropriateness and understanding by adolescents, ages 13 to 14 y, as a means of assessing face validity by non-experts. The edited preliminary questionnaire (still with 88 items) was administered to a nutrition expert group (n = 71) versus a non-expert group (n = 82), referred to as sample 1, for the purposes of performing item analysis and assessing construct validity of the questionnaire. The result of the analysis, a 63-item questionnaire, was administered to adolescents at three school grade levels, 8 (n = 128), 10 (n = 143), and 12 (n = 98), referred to as sample 2, which was representative of the grades in which the Birth-to-Twenty group will be in when the questionnaire is administered. The questionnaire was administered to the sample to assess its content validity and internal consistency reliability. The final questionnaire had 60 items, and its construct, content, and internal consistency reliability were reassessed. RESULTS: The final 60-item questionnaire displayed a significant difference (P < 0.0001) in the mean scores of the expert and non-expert groups tested. It had internal consistencies (Cronbach's alpha) of 0.71, 0.79, and 0.82 for grades 8, 10, and 12 respectively, and an overall value of 0.77 for all groups combined. However, it was less than 0.7 for most grade 8 pupils and for all grades at a historically disadvantaged school. CONCLUSION: A nutritional knowledge questionnaire with construct, face, and content validities and internal consistency was developed for use in South African adolescents to evaluate their nutritional knowledge. Internal consistency was low in children at a disadvantaged school and those in grade 8 compared with multiracial groups at a multiracial school. It is recommended that pupils at disadvantaged schools be assisted by trained interviewers when taking the test. PMID- 15661482 TI - Predictors of food variety and dietary diversity among older persons in Botswana. AB - OBJECTIVE: We investigated whether food variety and diversity are associated with physical and cognitive functioning in older adults in Botswana and designed a simple set of screening questions that predict food variety in this population. METHODS: Data were collected (1998) as a national household survey of 1085 subjects 60 y and older. A food variety score, based on a food frequency checklist, was calculated by summing the frequency of weekly intakes of 16 food items (0 to 66). A dietary diversity score was calculated as the number of food groups consumed weekly (0 to 5). A representative subsample (n = 393) was randomly selected for the clinical component of the survey, and measurements on dependency and cognitive function were conducted. RESULTS: Low food variety was found: 35.2%, 59.3%, and 22.4% of subjects consumed no dairy products, fruits, and vegetables, respectively. A higher food variety score was associated with urban residence, ownership of cattle, higher education, and more frequent meals, and these indicators were used to construct a nutritional risk indicator. Higher food variety score was associated with better self-reported health and better cognitive function. Similarly, a higher score on the nutritional risk indicator screening tool was associated with desirable health outcomes. CONCLUSION: A limited number of foods is consumed, leading to an overall pattern of poor food variety. Higher food variety was associated with improved physical and cognitive functions. A screening tool that predicts food variety in this population has been developed and is recommended to be incorporated at a primary care level to identify older adults most at risk of a poor quality diet. PMID- 15661483 TI - Nutrition and HIV/AIDS in sub-Saharan Africa: an overview. AB - HIV/AIDS is most prevalent in Sub-Saharan Africa where, exacerbated by the presence of other common conditions such as malnutrition and opportunistic infections, it is wreaking devastation on families, communities, and nations. Just as epidemics vary by country, so do national responses to this complex emergency. This is illustrated by the cases of Botswana, South Africa, and Uganda. Nutritional and micronutrient deficiencies play an important additive role in immune degradation and impaired development in children. Careful implementation of antiretroviral drugs, complemented by simultaneous efforts to ensure proper nutrition among HIV-infected children and adults are essential components of an effective response to the HIV/AIDS pandemic in Africa and elsewhere. PMID- 15661484 TI - Review of the South African nutrition policy 1994-2002 and targets for 2007: achievements and challenges. AB - Primary and secondary nutrition interventions are essential in South Africa, a country with mortality rates of 45.2 deaths per 1000 live births and 61 per 1000 for children younger than 5 y and an estimated prevalence of 8.3% for low birth weight. In addition, the National Food Consumption Survey has recently reported that approximately one in five children 1 to 9 y of age in South Africa are stunted (21.6%) and 1 in 10 (10.3%) is underweight for age. The prevalence of stunting was highest in children 1 to 3 y old (25.5%) and lowest in those 7 to 9 y old (13%). In terms of overnutrition, the survey also reported overweight and obesity in 17.1% of the sample at the national level. In this overview, the focus areas of the Integrated Nutrition Program are examined and critically discussed to determine whether targets planned for 2002 were met and how they are envisioned to change by 2007. PMID- 15661485 TI - Enteral nutrition support. PMID- 15661486 TI - What can we learn from atomic force microscopy adhesion measurements with single drug particles? AB - Frequently solid dosage form formulation manufacture and delivery depend critically on the control and exploitation of interparticulate interactions. Traditional approaches to understand such interactions rely on indirect assessments of adhesion or consider the behaviour of large numbers of particles. In recent years, the possibility of characterizing and perhaps quantifying forces of adhesion between individual micron and sub-micron sized particles has become viable using the atomic force microscope. This has significant potential in formulation development, particularly in the optimization of inhalation and other solid-dosage form based therapies. However, before a widespread acceptance of this approach by pharmaceutical scientists and industry can proceed a number of issues remain to be considered. These include how can single particle events be mapped on to bulk behaviour, the need to understand the sometimes wide variations in adhesion data observed and can formulations be compared quantitatively and perhaps be screened by this approach? PMID- 15661487 TI - Synthesis and in vitro evaluation of macromolecular antitumour derivatives based on phenylenediamine mustard. AB - Poly-[N-(2-hydroxyethyl)-L-glutamine] (PHEG) and poly(ethylene glycol) (PEG) grafted PHEG conjugates of N,N-di(2-chloroethyl)-4-phenylenediamine mustard (PDM) were synthetised. A collagenase-sensitive oligopeptide spacer was selected to link the cytotoxic agent PDM onto the polymeric carrier. First, the oligopeptide drug conjugate, L-pro-L-leu-gly-L-pro-gly-PDM, was prepared. In a second step, the low molecular weight PDM derivative and PEG-NH(2) were coupled to a N,N disuccinimidylcarbonate activated PHEG. Dynamic laser light scattering measurements indicated the formation of aggregates. The presence of human serum albumin had no significant effect on the diameter of the conjugates. The hydrolytic stability of the conjugates was investigated in buffer solutions. The conjugates showed an improved stability compared to the parent nitrogen mustard. The enzymatic degradation studies of the polymeric conjugates were performed in the presence of collagenase type IV (Clostridiopeptidase A; EC 3.4.24.3), cathepsin B (EC 3.4.22.1), cathepsin D (EC 3.4.23.5) and tritosomes. Only the bacterial collagenase type IV was able to cleave the spacer releasing free PDM and its peptidyl derivative, gly-L-pro-gly-PDM. The in vitro cytotoxicity of the conjugates was evaluated against HT1080 fibrosarcoma cells and MDA adenocarcinoma cells. All conjugates showed low toxicity towards these cell lines. PMID- 15661488 TI - Compact size and mechanical strength of pharmaceutical diluents. AB - The effects of compact size and powder particle size on the axial compressive (sigma(C)) and diametral tensile (sigma(T)) strength were evaluated, for three compressed pharmaceutical diluents (microcrystalline cellulose, MCC, calcium hydrogen phosphate dihydrate, CHPD, and pregelatinized starch, PGS) differing in deformational behaviour during compaction and testing. Cylindrical specimens of similar aspect ratio ( approximately 1) but different diameter (13, 10 and 5mm) and total porosity (0.35-0.05) were employed and the extrapolated and interpolated strength values, at zero and 0.15 porosity (sigma(C0) or sigma(T0) and sigma(C0.15) or sigma(T0.15)), were predicted by applying an exponential relation. The ratio of sigma(C0.15)/sigma(T0.15) was correlated to the corresponding percent radial elastic recovery (%RR). It was found that the changes of compressive and tensile strength with compact size were not characteristic and general. Specimen's size affected both extrapolated and interpolated values of sigma(C) and sigma(T) most significantly in the case of MCC, for which sigma(T) increased while sigma(C) decreased, as the compact size became smaller. Less significant was the effect of compact size on the interpolated value of compressive strength (sigma(C)) and the extrapolated value of tensile strength (sigma(T)) for PGS. For CHPD, no significant and characteristic effect of compact size was observed. Particle size significantly affected the interpolated values of sigma(T) for MCC and both sigma(C) and sigma(T) for CHPD; they all increased with decreasing particle size. The sigma(C0.15)/sigma(T0.15) ratio was affected significantly by the compact size only in the case of MCC and seems to be an indication of material elasto plasticity, since PGS has by far the highest ratio, followed by MCC and CHPD. A correlation between sigma(C0.15)/sigma(T0.15) and %RR was established as was its dependence upon deformational behaviour. PMID- 15661489 TI - Clinical study of solid dispersions of itraconazole prepared by hot-stage extrusion. AB - The aim of this study was to investigate the performance of three new solid dispersion formulations of itraconazole in human volunteers in comparison with Sporanox, the marketed form. Solid dispersions made up of itraconazole (40%, w/w) and HPMC 2910, Eudragit E100 or a mixture of Eudragit E100-PVPVA64 were manufactured by hot-stage extrusion and filled in gelatin capsules. The formulations were tested in eight human volunteers in a double blind, single dose, and cross-over study. Concentrations of the drug and its metabolite hydroxyitraconazole in the plasma were determined using HPLC. The in vivo performance was evaluated by comparing the mean area under the plasma concentration-time curves (AUC), the mean maximum plasma concentration (C(max)), and the mean time to reach C(max) (T(max)). The mean bioavailability of itraconazole was comparable after administration of the HPMC solid dispersion, compared to Sporanox, while it was lower after administration of the Eudragit E100 or Eudragit E100-PVPVA64 dispersions. Due to high variability, a significant decrease in AUC and C(max) was only observed for the Eudragit E100-PVPVA formulation. Although the solid dispersions showed different in vitro dissolution behaviour, T(max) values were comparable. The same observations with respect to AUC, C(max) and T(max) could be made for hydroxyitraconazole. The present results indicate that hot-stage extrusion can be considered as a valuable alternative for manufacturing solid dispersions of itraconazole. PMID- 15661490 TI - Coating alginate microspheres with a serum albumin-alginate membrane: application to the encapsulation of a peptide. AB - Calcium alginate gel microspheres coated with a human serum albumin (HSA) alginate membrane were prepared adapting a transacylation method previously applied to large beads. The procedure involved emulsification of an aqueous solution of sodium alginate and propylene glycol alginate (PGA) in an oily phase, followed by addition of CaCl(2). The resulting gel microspheres were transferred in an aqueous solution of HSA. The addition of 0.5 M NaOH started the reaction between PGA and HSA, producing amide bonds and forming a membrane around the particles. An optimization study was conducted, notably exploring the addition of HSA to the internal phase. The microcapsules were studied with respect to morphology (optical and scanning electron microscopy) and size (laser granulometry), in comparison with uncoated gel microspheres. Biocompatibility was checked in osteoblast cultures. Lysine-arginine-phenylalanine-lysine (KRFK) was encapsulated and the release kinetics was studied in vitro. The method provided stable microspheres (size around 60 microm), with a membrane surviving a treatment with citrate and resisting lyophilization. The microcapsules were shown biocompatible. The release of KRFK was slower (release time>8 days) than that of uncoated microspheres. These microcapsules might be useful as peptide containers to be combined with prosthetic materials for improving osteointegration. PMID- 15661491 TI - Preparation and characterization of solid lipid nanoparticles (SLN) made of cacao butter and curdlan. AB - Solid lipid nanoparticles (SLN) were prepared using cacao butter, as the lipid core, and curdlan, as the shell material. Tween 80 was used as a co-surfactant in order to prevent aggregation and gelling of the curdlan. Mannitol was used as a cryoprotectant in order to prevent aggregation during redispersion. No significant change in the size of the SLN was observed up to a lipid concentration of 1.0%, and the particle size ranged from 140 to 200 nm with a unimodal distribution. When an alternating pH between 7 and 11 was used to test the physical stability of an SLN solution, the change in the particle size remained within a narrow range up to a lipid concentration of 0.5%. Above 0.5%, the particles began to aggregate due to the insufficient amount of the coating material, curdlan and Tween 80. The critical aggregation concentration at pH 7.4 was found to be 6.95 x 10(-4) mg/ml. Pyrene was used as a fluorescence probe. As the temperature increased, pyrene was gradually released from the SLN. The loading efficiency was >75% when the verapamil to lipid ratios were 1:10 and 1:5 and decreased significantly as the ratio became 1:1. The release rate was significantly delayed when verapamil was loaded into the SLN. PMID- 15661492 TI - Production of enteric capsules by means of hot-melt extrusion. AB - The aim of this study was to develop an alternative technique for enteric coating consisting of the hot-melt extrusion of coating polymers. An enteric coating polymer (PVAP or HPMC AS), premixed with a plasticizer, was extruded into hollow cylinders. The hollow pipes were filled with a model drug and both open ends of the cylinders were closed, yielding hot-melt extruded enteric capsules. Main advantages of this new technology are the continuity of the process and its application for the formulation of moisture sensitive active ingredients. The enteric capsules showed excellent gastro-resistance, since no drug release was observed after 2 h 0.1N HCl. The influence of wall thickness (0.15, 0.3, 0.5, 0.8, and 1.0 mm) of the capsules on drug release was investigated. Enteric capsules with a wall thickness of 1.0 mm were subjected to a pH gradient dissolution method, simulating passage through the gastro-intestinal tract, in order to evaluate their suitability for ileal or colonic drug targeting. Storing the capsules for 1 month at high relative humidity (RH) (60 and 75% RH) revealed that the HPMC AS capsules were superior to the PVAP capsules. It can be concluded that hot-melt extruded capsules seem suitable as an alternative for enteric coating. PMID- 15661493 TI - Application of graph theory: prediction of glycogen synthase kinase-3 beta inhibitory activity of thiadiazolidinones as potential drugs for the treatment of Alzheimer's disease. AB - The relationship of Wiener's index, a distance-based topological descriptor; Zagreb group parameter, M(1), an adjacency-based topological descriptor and eccentric connectivity index,an adjacency-cum-distance based topological descriptor with the glycogen synthase kinase-3 beta inhibitory activity of thiadiazolidinones has been investigated. A training set comprising 28 analogues of substituted thiadiazolidinones was selected for the present investigations. The values of the Wiener's index, Zagreb group parameter, and eccentric connectivity index for each of 28 analogues comprising the data set were computed. Resulting data was analyzed and suitable models developed after identification of active ranges. Subsequently, biological activity was predicted for each analogue involved in the data set using these models, which was then compared with the reported glycogen synthase kinase-3 beta inhibitory activities. Accuracy of prediction was found to vary from a minimum of approximately 83% for model based on Zagreb group parameter to a maximum of approximately 87% for model based on Wiener's index. PMID- 15661494 TI - 2,3-Diarylimidazo[1,2-a]pyridines as potential inhibitors of UV-induced keratinocytes apoptosis: synthesis, pharmacological properties and interactions with model membranes and oligonucleotides by NMR. AB - Four 2,3-diarylimidazo[1,2-a]pyridines (I, 1a-c) were synthesized as inhibitors of UV-induced apoptosis and showed quite different properties. First, only the pyridinyl derivative I showed protection in molt cells. From the supposed intracellular target, phospholipid membrane models were studied by (1)H, (2)H and (31)P NMR spectroscopy. All these molecules can incorporate the membrane bilayer of small unilamellar vesicles of lecithin (SUV). However, I is clearly closed to the external polar head of the lipids, and is relatively mobile in the layer. Conversely, the other molecules are strongly immobilized in the deep part of the external layer. (31)P solid-state NMR spectra recorded on phospholipid dispersions (multilayers vesicles (MLV)) completely excluded any detergent effect or any modification of temperature transition. The only structural or dynamic effect observed was a homogeneous, but limited, reduction in the chemical shift anisotropy in the presence of I, in agreement with its superficial location. (2)H NMR experiment performed on the same model using perdeuterated phospholipids showed no significant fluidity reduction at the level of terminal CD(3) groups in the presence of 1a-c, according to their deep location. Finally, their interactions with synthetic oligonucleotide, d(CGATCG)(2) was studied showing non specific interactions of 1a on the external GC pair, while no interaction was observed with the other derivatives. PMID- 15661495 TI - A new method to determine the partial solubility parameters of polymers from intrinsic viscosity. AB - A modification of the extended Hansen method, formerly used to determine the partial solubility parameters of drugs and non-polymeric excipients is tested with a polymer for the first time. The proposed method relates the logarithm of the intrinsic viscosities of the polymer in a series of solvents and solvent mixtures with the Hansen (three parameter model) and Karger (four parameter model) partial solubility parameters. The viscosity of diluted solutions of hydroxypropyl methylcellulose (HPMC) was determined in pure solvents and binary mixtures of varying polarity. The intrinsic viscosity was obtained from the common intercept of the Huggins and Kraemer relationships. The intrinsic viscosity tends to increase with increasing the solubility parameter of the medium. The results show that hydrogen bonding and polarity of the polymer largely determine polymer-solvent interactions. The models proposed provided reasonable partial and total solubility parameters for the polymer and enable one to quantitatively characterize, for the first time, the Lewis acid-base ability of a polymer thus, providing a more realistic picture of hydrogen bonding for solvent selection/compatibility and to predict drug-polymer interactions. Combination of the dispersion and polar parameters into a single non-specific solubility parameter was also tested. The results extend earlier findings and suggest that the models are quite versatile and may be applied to drugs, non polymeric and polymeric excipients. PMID- 15661496 TI - Percutaneous absorption of cyclizine and its alkyl analogues. AB - Cyclizine (I) alkyl analogues (II-IV) were synthesized and their skin permeation parameters evaluated in vitro. It was hoped that these compounds would possess physicochemical properties more favourable for percutaneous delivery than (I). The identification and levels of purity for the compounds were confirmed by mass spectrometry (MS), nuclear magnetic resonance (NMR) spectrometry, and infrared spectrometry (IR) while melting points were determined by an electrothermal digital Bupsilonchi melting point apparatus. Aqueous solubilities (25 degrees C) and partition coefficients were determined and in vitro permeation studies were performed in buffer (37 degrees C) at pH 7.4 over a period of 24 h, using Franz diffusion cells fitted with human epidermal membranes. Generally, the analogues were more lipophilic, but nevertheless possessed higher aqueous solubilities as compared to (I). (II) and (IV) exhibited two- to three-fold increase in aqueous solubility and their melting temperatures dropped by more than 55 degrees C. Compound (III) had similar aqueous solubility to (I), but its melting point dropped by about 35 degrees C. Measured steady-state fluxes indicated that (II) is a far better penetrant (J=6.95 microg/cm(2)/h) of human epidermis than (I). Although fluxes of (III) and (IV) drop off markedly from that of (II), they remained above the flux of (I), which is (0.132 microg/cm(2)/h). In conclusion, (II) was the best skin permeant and also exhibited the highest aqueous solubility and lowest level of crystallinity as compared to (I) and other analogues. (III) and (IV) were more lipophilic. The overall permeation data of this series indicated that the more water-soluble and the lowest melting point compound was the best skin permeant. PMID- 15661497 TI - Spray-drying of proteins: effects of sorbitol and trehalose on aggregation and FT IR amide I spectrum of an immunoglobulin G. AB - An immunoglobulin G (IgG) was spray-dried on a Buchi 190 laboratory spray-dryer at inlet and outlet air temperatures of 130 and 190 degrees C, respectively. The IgG solution contains initially 115 mg/ml IgG plus 50 mg/ml sorbitol. After dialysis, at least 80% of low molecular weight component was removed. After spray drying the dialyzed IgG and immediate redissolution of the powder, an increase in aggregates from 1 to 17% occurred. A major shift towards increase beta-sheet structure was detected in the spray-dried solid, which, however, reverted to native structure on redissolution of the powder. A correlation between aggregation determined by size exclusion chromatography and alterations in secondary structure determined by Fourier transformation infra-red spectroscopy could not therefore be established. On spray-drying a non-dialyzed, sorbitol containing IgG only some 0.7% aggregates were formed. The sorbitol is therefore evidently able to stabilize partially the IgG during the process of spray-drying. Addition of trehalose to the liquid feed produced quantitatively the same stabilizing action on the IgG during spray-drying as did the sorbitol. This finding again points towards a water replacement stabilization mechanism. The IgG spray-dried powder prepared from the dialyzed liquid feed showed continued substantial aggregation on dry storage at 25 degrees C. This was substantially less in the non-dialyzed, sorbitol-containing spray-dried powder. Addition of trehalose to both dialyzed and non-dialyzed system produced substantial improvement in storage stability and reduction in aggregate formation in storage. The quantitative stabilizing effect of the trehalose was only slightly higher than that of the sorbitol. Taken together, these results indicate that both the sorbitol and trehalose stabilize the IgG primarily by a water replacement mechanism rather than by glassy immobilization. The relevance of this work is its questioning of the importance of the usually considered dominance of glassy stabilization of protein in dried systems of high glass transition temperature, such as trehalose. The low glass transition temperature sorbitol produces almost equal process and storage stability in this case. PMID- 15661498 TI - Novel D-penicillamine carrying nanoparticles for metal chelation therapy in Alzheimer's and other CNS diseases. AB - Metal ions accumulate in the brain with aging and in several neurodegenerative diseases. Aside from the copper storage disease, Wilson's disease, recent attention has focused on the accumulation of zinc, copper and iron in the Alzheimer's disease (AD) brain and the accumulation of iron in Parkinson's disease. In particular, the parenchymal deposition of beta-amyloid (Abeta) and its interaction with metal ions has been postulated to play a role in the progression of AD. Thus, the strategy of lowering brain metal ions and targeting the interaction of Abeta peptide and metal ions through the administration of chelators has merit. Our recent finding that nanoparticle delivery systems can cross the blood-brain barrier has led us to investigate whether chelators delivered conjugated to nanoparticles could act to reverse metal ion induced protein precipitation. In the present studies, the Cu (I) chelator D penicillamine was covalently conjugated to nanoparticles via a disulfide bond or a thioether bond. Nanoparticle-chelator conjugates were stable between pH 6-8 in aqueous suspension if stored at 4 degrees C, and did not aggregate when challenged with salts and serum. Release of D-penicillamine from the nanoparticles was achieved using reducing agents such as dithiothreitol (as a model for glutathione). Nanoparticles treated only under reducing conditions that released the conjugated D-penicillamine were able to effectively resolubilize copper-Abeta (1-42) aggregates. These results indicate that nanoparticles have potential to deliver D-penicillamine to the brain for the prevention of Abeta (1 42) accumulation, as well as to reduce metal ion accumulation in other CNS diseases. PMID- 15661499 TI - Development of an enteric-coated pellet formulation of F4 fimbriae for oral vaccination of suckling piglets against enterotoxigenic Escherichia coli infections. AB - A multi-particulate formulation of F4 fimbriae was developed for oral vaccination of suckling piglets against enterotoxigenic Escherichia coli infections. A feasibility test showed that incorporation of F4 fimbriae in a disintegrating pellet formulation consisting of 87.5% Pharmatose 200 M, 2.5% Avicel CL 611 and 10% Explotab by extrusion/spheronisation and subsequent fluid bed drying resulted in the maintenance of 69+/-12% of the biological activity. But subsequent coating resulted in pellets with poor enteric properties, although good in vivo immunising results were obtained after administration to piglets. From the economical point of view, a pellet formulation was optimised to decrease vaccine dose and dosing frequency. After disintegration testing, pellets consisting of lactose (alpha-lactose monohydrate 90 mesh/beta-lactose 75/25 (w/w)) and microcrystalline cellulose in a ratio of 80/20 (w/w) showed a sponge-like structure from which F4 fimbriae could be released. Coating of these pellets resulted in good enteric properties. To improve disintegrating properties of the pellets, the lactose concentration was increased or sodium carboxymethyl starch was added. But this resulted in poor enteric properties after coating. Dissolution test showed that F4 fimbriae were released from the optimised enteric coated pellets but interaction between F4 fimbriae and the coating polymer was seen. This incompatibility leads to unpredictable in vitro quantification of F4 biological activity. PMID- 15661500 TI - Studies on a new device for drug delivery to the eye. AB - Delivery of drugs to the anterior side of the eye is routinely done with eye drops, but this method results in low bioavailability and low patient compliance. Herein, we describe a new device for the delivery of drugs to the eye. The device, called the OphthaCoil, consists of a drug-loaded adherent hydrogel coating on a thin metallic wire, which is coiled. The drug release rates of the dye fluorescein and the antibiotic chloramphenicol have already been evaluated in vitro. In this report the drug release rate of the anti-infective pradofloxacin was evaluated in vitro and in vivo. The data show that the OphthaCoil is capable of sustained drug delivery to the tear film in dogs. Drug levels in the tear fluid of the dogs were well above the MIC-values of relevant bacteria after 16 h, but it should be noted that pradofloxacin has an exceptionally high antimicrobial activity. The study indicates that the OphthaCoil holds promise as a platform for sustained release of drugs to the eye. The device was well tolerated, but the devices were lost when left overnight. Most probably, this is due to the third eyelid pushing the device out of the conjunctival sac during sleep. It should be noted that this complication has no immediate implication for extended wear of the OphthaCoil in humans, as humans do not have third eyelids. PMID- 15661501 TI - Application of an electronic nose system for evaluation of unpleasant odor in coated tablets. AB - The purpose of this study was to apply an electronic nose system for evaluation of unpleasant odor in tablets containing L-cysteine, an unpleasant odor drug, and demonstrate the odor masking ability of thin-layer sugarless coated tablets, which we have newly developed, by both electronic nose system and sensory evaluations. We demonstrated the qualitative evaluation of the unpleasant odor using air as a reference indicator and the quantitative evaluation of the unpleasant odor using the distances between air and samples in the electronic nose system evaluation. The electronic nose system evaluation was positively and well-correlated with the sensory evaluation by volunteers. We suggest that the electronic nose system evaluation is appropriate as an alternative or a support method for sensory evaluation by volunteers. As the results of both electronic nose system and sensory evaluations, we demonstrated that the thin-layer sugarless coated tablets have excellent masking ability of the unpleasant odor, equivalent to that of sugar-coated tablets due to the dense coating layers. PMID- 15661502 TI - Quantitative evaluation of polymer concentration profile during swelling of hydrophilic matrix tablets using 1H NMR and MRI methods. AB - Many pharmaceutical tablets are based on hydrophilic polymers, which, after exposure to water, form a gel layer around the tablet that limits the dissolution and diffusion of the drug and provides a mechanism for controlled drug release. Our aim was to determine the thickness of the swollen gel layer of matrix tablets and to develop a method for calculating the polymer concentration profile across the gel layer. MR imaging has been used to investigate the in situ swelling behaviour of cellulose ether matrix tablets and NMR spectroscopy experiments were performed on homogeneous hydrogels with known polymer concentration. The MRI results show that the thickest gel layer was observed for hydroxyethylcellulose tablets, followed by definitely thinner but almost equal gel layer for hydroxypropylcellulose and hydroxypropylmethylcellulose of both molecular weights. The water proton NMR relaxation parameters were combined with the MRI data to obtain a quantitative description of the swelling process on the basis of the concentrations and mobilities of water and polymer as functions of time and distance. The different concentration profiles observed after the same swelling time are the consequence of the different polymer characteristics. The procedure developed here could be used as a general method for calculating polymer concentration profiles on other similar polymeric systems. PMID- 15661503 TI - pH- and temperature-sensitive release behaviors from polyelectrolyte complex films composed of chitosan and PAOMA copolymer. AB - This paper describes the pH and temperature effects on drug release from polyelectrolyte complex (PEC) films composed of a cationic polymer, chitosan, and an anionic polymer, polyalkyleneoxide-maleic acid copolymer (PAOMA). In this study, we prepared and investigated PEC films in terms of the drug release properties as pH- and temperature-sensitive drug carriers. Drug release rates were tested at pH 3.8 and 7.2, and at 25 and 50 degrees C. Salicylic acid and phenol were selected as model drugs. An increase in pH from 3.8 to 7.2 resulted in an increase in the rate of drug release because of the repulsive forces between carboxyl groups in PAOMA and anionic groups in model drugs. When the hydrophobic PAOMA was used as a polyanion, the drug release rate increased at 50 degrees C. This is attributed to the increase of release area due to the phase transition of PAOMA and the increase of repulsive forces between carboxyl groups in PAOMA and anionic groups in model drugs. PMID- 15661504 TI - Understanding agglomeration of indomethacin during the dissolution of micronised indomethacin mixtures through dissolution and de-agglomeration modeling approaches. AB - The purpose of this research was to correlate the state of agglomeration determined by the modeling of dissolution and de-agglomeration profiles, using mixtures of micronised indomethacin designed to have different dissolution rates and extents of particle agglomeration in dissolution media. Dissolution profiles were determined using the USP paddle method. De-agglomeration profiles were obtained from laser diffraction particle sizing of mixtures of indomethacin in dissolution media under non-sink conditions. Data were modeled and key parameters estimated using a non-linear least squares estimation algorithm. The key parameters of initial apparent volume concentrations as dispersed and agglomerated particles, and dissolution rate constants (for dissolution modeling), and the apparent volume concentrations of dispersible and non dispersible agglomerates and the de-agglomeration rate constant (for de agglomeration modeling) were related to indomethacin and sodium lauryl sulphate concentrations in the lactose-povidone mixtures. Micronised sodium lauryl sulphate added to the mixture was more effective in de-agglomeration than equivalent concentrations in the dissolution media. An excellent correlation existed between the total initial apparent volume concentration of agglomerates determined by dissolution and de-agglomeration (P=0.98). The use of key parameters estimated from the modeling of dissolution and de-agglomeration profiles provides a useful tool in dosage form development of formulations of poorly water soluble drugs. PMID- 15661505 TI - Isothermal titration calorimetry method for determination of cyclodextrin complexation thermodynamics between artemisinin and naproxen under varying environmental conditions. AB - A novel isothermal titration calorimetry method was used to determine the complexation thermodynamics for hydroxypropyl-beta-cyclodextrin with artemisinin and naproxen at varying temperature and pH. The new method is very useful for studying complexation reactions between cyclodextrin and drugs with poor solubility and all the thermodynamic parameters of the cyclodextrin complexation were determined. The analysis of the thermodynamic data reveals involvement of hydrophobic bonding in the cyclodextrin complexes studied. The data also reveals the presence of enthalpy-entropy compensation in the system and provide information as to the orientation of the drug molecule inside the cyclodextrin cavity. From the thermodynamic parameters for dissociation of HPBCD complexes of artemisinin and naproxen at pH 2 it is concluded that the complexation is primarily driven by enthalpy with entropic assistance at all temperatures studied. From the dissociation studies of HPBCD complexes of naproxen at pH 10 it is concluded that the complexation is predominantly driven by entropy and moderately by enthalpy at lower temperatures and by enthalpy with entropic assistance at higher temperatures. PMID- 15661506 TI - Viscoelastic and structural properties of pharmaceutical hydrogels containing monocaprin. AB - The purpose of this study was to characterize the rheological and structural properties of a pharmaceutical multicomponent hydrogel formulation. This formulation consists of a hydrogel-forming poly(acrylic acid) polymer (Carbopol 974P), microbicide (monocaprin), non-ionic surfactant (Tween 20 or Tween 40), and preservatives. The effects of surfactant addition, monocaprin concentration, and pH on the formulation are investigated with the aid of various rheological methods and small angle neutron scattering (SANS). A change of pH from 4 (reduced electrostatic interactions) to higher pH values (prominent electrostatic forces) resulted in a marked impact on both the structural and rheological characteristics, with higher values of the dynamic moduli. At pH 4, the rheological features were strongly influenced by the addition of surfactant and monocaprin concentration, whereas at higher values of pH the effect of additives was modest. The picture that emerges from SANS and rheology is that enhanced association structures evolve at pH 4, while at higher pH the tendency to form associations is inhibited. At all the conditions, the rheological results suggest a viscoelastic solid behavior, which is typical for many gels. PMID- 15661507 TI - Physical solid-state properties and dissolution of sustained-release matrices of polyvinylacetate. AB - Solid-state compatibility and in vitro dissolution of direct-compressed sustained release matrices of polyvinylacetate (PVAc) and polyvinylpyrrolidone (PVP) containing ibuprofen as a model drug were studied. Polyvinylalcohol (PVA) was used as an alternative water-soluble polymer to PVP. Differential scanning calorimetry (DSC) and powder X-ray diffractometry (PXRD) were used for characterizing solid-state polymer-polymer and drug-polymer interactions. The mechanical treatment for preparing physical mixtures of polyvinyl polymers and the drug (i.e. simple blending or stressed cogrinding) was shown not to affect the physical state of the drug and the polymers. With the drug-polymer mixtures the endothermic effect due to drug melting was always evident, but a considerable modification of the melting point of the drug in physical binary mixtures (drug:PVP) was observed, suggesting some interaction between the two. On the other hand, the lack of a significant shift of the melting endothermic peak of the drug in physical tertiary drug-polymer mixtures revealed no evidence of solid state interaction between the drug and the present polymers. Sustained-release dissolution profiles were achieved from the direct-compressed matrices made from powder mixtures of the drug and PVAc combined with PVP, and the proportion of PVAc in the mixture clearly altered the drug release profiles in vitro. The drug release from the present matrix systems is controlled by both diffusion of the drug through the hydrate matrix and the erosion of the matrix itself. PMID- 15661508 TI - Use of roughness maps in visualisation of surfaces. AB - In this study we will present a new method to describe surface roughness. This method builds a roughness map of the studied area. The roughness map can give information of localised roughness. The test surfaces used in the evaluation of the method were tablets, which were made of lactose monohydrate, theophylline anhydrate, sodium chloride and potassium chloride. The roughness determinations were made by a laser profilometer. The new matrix method gives detailed roughness maps, which are able to show local variations in surface roughness values and provide an illustrative picture of the heterogeneity of surface roughness of various materials. PMID- 15661509 TI - Targeting of sterically stabilised pH-sensitive liposomes to human T-leukaemia cells. AB - The main aim of this work was to develop novel targeted sterically stabilised pH sensitive liposomes tailored to promote efficient intracellular delivery of therapeutic molecules into human T-leukaemia cells. Our results indicate that the targeting moiety (thiolated transferrin) was successfully coupled to the distal reactive maleimide terminus of poly(ethylene glycol)-phospholipid conjugates incorporated in the liposomal bilayer. Results from atomic force microscopy studies, performed to characterise vesicle surface topology, indicated that, to a certain extent, thiolated transferrin has the ability to associate in a non specific manner with the lipid membrane of pegylated liposomes. This is an issue not commonly reported in the literature but which is crucial to demonstrate the targeting proof of principle. Nevertheless, fluorimetric studies together with confocal microscopy clearly demonstrate that liposomes bearing covalently coupled transferrin associate more extensively to human T-leukaemia cells in vitro than non-targeted liposomes. Cell mechanistic studies indicate that targeted liposomes bind specifically to transferrin receptors and are internalised via receptor dependent endocytotic pathway. In addition, the biophysical features exhibited by the developed liposomes, namely their ability to promote pH-triggered cytoplasmic delivery of loaded material, make them promising delivery systems for in vivo targeting of therapeutic molecules to tumours. PMID- 15661510 TI - Observations in simultaneous microencapsulation of 5-fluorouracil and leucovorin for combined pH-dependent release. AB - 5-Fluorouracil (5-FU) in combination with leucovorin (LV) is nowadays the standard treatment in colon cancer and would be a candidate to be delivered orally to the colon. Eudragit P-4135F or Eudragit RS100 were used separately to prepare microspheres by an oil/oil emulsification process trapping 5-FU and LV simultaneously. Scanning electron microscopy permitted a structural analysis, process parameters were analyzed and drug loading and release profiles were recorded. Particle size varied between 123 (RS100) and 146 microm (P-4135F). Generally, higher encapsulation rates were found with RS100 (5-FU, 60.3+/-9.7%; LV, 81.4+/-8.6%) compared to P-4135F (5-FU, 48.3+/-2.0%; LV, 55.4+/-2.7%). Microparticles made from Eudragit RS100 released the incorporated drug combination within 8 h not exhibiting general differences between the kinetics of both drugs. P-4135F was found to maintain the undesired 5-FU release at pH 6.8 lower than 25% within 4 h while at pH 7.4, a nearly immediate release (within 15 min) was observed. Although the release was similar at pH 7.4, at pH 6.8 LV showed a distinct initial drug loss of about 60% and a complete release within 2 h. SEM analyses revealed a substantial presence of LV crystals on the particle surface provoking a distinct burst effect of LV. These observations were concluded to be related to the high lipophilicity of P-4135F provoking a separation between P-4135F and LV during the preparation process. PMID- 15661512 TI - RNAi as a tool to study cell biology: building the genome-phenome bridge. AB - In the few short years since its discovery, RNA interference (RNAi) has revolutionized the functional analysis of genomes: both technical and conceptual approaches to the investigation of gene function are being transformed as a result of this new technology. Genome-scale RNAi analyses have already been performed in the model organisms Caenorhabditis elegans (in vivo) and Drosophila melanogaster (in cell lines), ushering in a new era of RNAi-based approaches to probing the inner workings of the cell. The transformation of complex phenotypic data into mineable 'digitized' formats is fostering the emergence of a new area of bioinformatics related to the phenome. PMID- 15661513 TI - A reductionist's systems biology: opinion. AB - To tackle the complexity inherent in understanding large networks of interacting biomolecules, systems biology emphasizes cybernetic and systems theoretical approaches. The resulting focus on organization independent of physical manifestation threatens to throw away all that has been learned from molecular studies and ignores the reality that biologists are drawn together more by a shared interest in mechanism and structure than anything else. The field of reaction engineering suggests a reductionist approach to systems biology that fits easily within existing molecular paradigms but that can nonetheless be integrated into expansive physiological perspectives through the use of multi scale modeling. PMID- 15661514 TI - Dissecting cell biology with chemical scalpels. AB - Understanding complex cellular processes requires methods for specifically perturbing protein function in a temporally defined fashion. In recent years a variety of chemical tools have been used to study the cytoskeleton and cell division, protein trafficking, and the destruction of proteins by the ubiquitin proteasome system. The ability to use combinations of reversible inhibitors is proving to be especially helpful in dissecting complex cellular events. Furthermore, the identification of novel inhibitors through unbiased screening approaches is revealing novel drugable steps in important cellular pathways. PMID- 15661515 TI - Oscillations in cell biology. AB - Oscillations play an important role in many dynamic cellular processes. They can emerge as the collective dynamic behavior of an ensemble of interacting components in the cell. Examples include oscillations in cytoskeletal structures such as the axonemes of cilia. Spontaneous oscillations of mechano-sensitive hair bundles have been shown to give frequency selectivity and amplification to mechano-sensation. In some bacteria, oscillations of Min proteins are important for division site selection. Genetic oscillators form the basis of circadian clocks. All these oscillations share many general features. Models and theoretical approaches are essential for an understanding of the principles underlying these dynamic cellular processes. PMID- 15661516 TI - Quantitative models of nuclear transport. AB - Nuclear pore complexes mediate the rapid trafficking of target macromolecules between the nucleus and the cytoplasm but exclude non-targets. Mathematical modeling helps to define the physical properties of a transport medium that can selectively enhance the permeation of some molecules but block others. Recent pioneering work has established a basis for quantitative modeling of nuclear translocation, and we expect this field to expand rapidly. A second area where modeling of nucleocytoplasmic transport has been prominently employed is in efforts to understand the regulatory networks by which signals pass between the nuclear and cytoplasmic compartments. Recent evidence suggests that the distinctive kinetics and spatial organization of nuclear transport processes can be used to efficiently propagate signals by new and unexpected pathways. PMID- 15661517 TI - Kinetochore-spindle microtubule interactions during mitosis. AB - The kinetochore is a proteinaceous structure that assembles onto centromeric DNA and mediates chromosome attachment to microtubules during mitosis. This description is deceivingly simple: recent proteomic studies suggest that the diminutive kinetochores of Saccharomyces cerevisiae are comprised of at least 60 proteins organized into as many as 14 different subcomplexes. Many of these proteins, such as the centromeric histone variant CENP-A, and entire subcomplexes, such as the Ndc80(Hec1) complex, are conserved from yeast to humans despite the diverse nature of the DNA sequences on which they assemble. There have recently been advances in our understanding of the molecular basis of how kinetochores establish dynamic attachments to spindle microtubules, and how these attachments are correctly oriented to ensure segregation of sister chromatids to daughter cells. PMID- 15661518 TI - Microtubule plus-end-tracking proteins: mechanisms and functions. AB - Microtubule plus-end-tracking proteins (+TIPs) are a diverse group of molecules that display dynamic accumulation at the distal ends of growing microtubules. Specific binding to the growing microtubule tip coupled with quick detachment from the older lattice, plus-end-directed transport, and association with other +TIPs can all contribute to this protein localisation. +TIPs act mainly as microtubule-stabilising factors and at the same time often link microtubule ends to various cellular structures, such as the cell cortex or kinetochores. Regulation of the activity of +TIPs has profound effects on the shape of the microtubule network and plays an essential role in cell division, motility and morphogenesis. PMID- 15661519 TI - When size matters: the dynamic regulation of stereocilia lengths. AB - Stereocilia, the mechanosensitive protrusions in hair cells, are organized into rows of graded heights forming precisely uniform staircase patterns. The actin turnover process in stereocilia follows a treadmill model in which the rate of treadmilling is scaled to the stereocilium's length. Myosin XVa, which is present at the site of actin polymerization at concentrations proportional to the length of the actin filament bundles, plays a combined role with the treadmill machinery in regulating the steady state length of these actin protrusions, together with other myosins localized alongside the actin bundles. PMID- 15661520 TI - The actin slingshot. AB - Actin polymerization generates the force that deforms the cell membrane, pulls the cell forward and propels endosomes and bacteria within the cell. The mechanism of force generation has been probed using experimental biomimetic systems where force generation and movement occur by the same actin polymerization processes observed in cells. The advantage of such systems over living cells is that their physical properties can be changed, such as the size of the load, its composition and its deformability, in order to respond to specific questions. Recent experimental developments and associated theoretical models have provided us with a better understanding of motility based on actin polymerization. This paves the way towards a better comprehension of cell motility. PMID- 15661521 TI - Force generation by dynamic microtubules. AB - The assembly and disassembly of microtubules can generate pushing and pulling forces that, together with motor proteins, contribute to the correct positioning of chromosomes, mitotic spindles and nuclei in cells. In vitro experiments combined with modeling have shed light on the intrinsic capability of dynamic microtubules to generate force, and various observations of positioning processes in cells and model systems have shown how pushing and pulling forces are used in different situations. A sophisticated set of microtubule-end-binding proteins is responsible for steering dynamic microtubules toward their cellular target and regulating the pushing and/or pulling forces that are generated once contact is established. PMID- 15661522 TI - Increasing complexity of the bacterial cytoskeleton. AB - Bacteria contain cytoskeletal elements involved in major cellular processes including DNA segregation and cell morphogenesis and division. Distant bacterial homologues of tubulin (FtsZ) and actin (MreB and ParM) not only resemble their eukaryotic counterparts structurally but also show similar functional characteristics, assembling into filamentous structures in a nucleotide-dependent fashion. Recent advances in fluorescence microscopic imaging have revealed that FtsZ and MreB form highly dynamic helical structures that encircle the cells along the inside of the cell membrane. With the discovery of crescentin, a cell shape-determining protein that resembles eukaryotic intermediate filament proteins, the third major cytoskeletal element has now been identified in bacteria as well. PMID- 15661523 TI - Microtubule-depolymerizing kinesins. AB - The fact that some kinesin-related proteins can destabilize microtubules is now a well-established fact. However, the contribution that these kinesins make to cellular function is just coming into focus. Key structural and kinetic studies on the mechanism of microtubule depolymerization by these kinesins have provided a framework for understanding their cellular regulation and function. Completion of some of the genome sequences and recent technological advances enabling the rapid depletion of cellular proteins in metazoans have clarified the functional role and level of cooperation between members of the depolymerizing kinesin families. Recent studies utilizing these technologies have revealed how these kinesins play an integral role in the mechanics of mitotic spindle assembly, chromosome segregation and the shaping of connections in the brain. PMID- 15661524 TI - Kinesin: world's tiniest biped. AB - Kinesin, an essential motor protein that moves intracellular cargo along microtubules, walks like a person. When we walk, our feet exchange roles with each step, one moving and one remaining stationary. The moving foot travels twice as far as our torso during a single step, and our body alternates between two configurations (left vs. right leg leading). Recent work shows that kinesin shares all three of these hallmarks of bipedal walking. The challenge now is to determine how the gait of this lilliputian biped is coordinated. PMID- 15661525 TI - Recent progress in dynein structure and mechanism. AB - Dynein is a minus-end-directed microtubule motor crucial to diverse cellular processes. The unwieldy size of the molecule and the difficulty of expressing and purifying mutants have hampered mechanistic studies of dynein. Recent progress sheds light on key unsolved questions concerning how the molecule is really organized, what conformational changes accompany ATP hydrolysis and whether two or three motor domains are coordinated in their motion. PMID- 15661527 TI - Mutational analysis of gene families in human cancer. AB - The completion of the human genome project has marked a new beginning in biomedical sciences. Human cancer is a genetic disease and, accordingly, the field of oncology has been one of the first to be impacted by this historic revolution. Knowledge of the sequence and organization of the human genome facilitates the systematic analysis of the genetic alterations underlying the origin and evolution of tumors. Recent mutational analyses in colorectal and other cancers have focused on examination of gene families involved in signal transduction, such as kinases and phosphatases. This approach has been successful in identifying mutations in a variety of different genes, including the identification of PI3KCA as one of the most commonly mutated oncogenes in human cancer. Such genomic analyses have already demonstrated their utility in basic and clinical cancer research, and are expected to have an important impact on future diagnostic and therapeutic strategies. PMID- 15661528 TI - Understanding transformation: progress and gaps. AB - Cancer is a collection of complex genetic diseases characterized by multiple defects in the homeostatic mechanisms that regulate cell growth, proliferation and differentiation. Although the analysis of human tumor specimens has allowed the identification of many molecules and pathways important for the malignant phenotype, we still lack a complete understanding of the events that conspire to program any specific type of cancer. Recent advances in developing human experimental models of cancer have provided new insights into the pathways whose perturbation is necessary to achieve cell transformation. These studies indicate that many combinations of genetic mutations confer tumorigenicity on human cells and that both cell-type and tumor-stromal interactions play critical roles in dictating the tumor phenotype. PMID- 15661529 TI - DNA tumor viruses -- the spies who lyse us. AB - Identifying the molecular lesions that are 'mission critical' for tumorigenesis and maintenance is one of the burning questions in contemporary cancer biology. In addition, therapeutic strategies that trigger the lytic and selective death of tumor cells are the unfulfilled promise of cancer research. Fortunately, viruses can provide not only the necessary 'intelligence' to identify the critical players in the cancer cell program but also have great potential as lytic agents for tumor therapy. Recent studies with DNA viruses have contributed to our understanding of critical tumor targets (such as EGFR, PP2A, Rb and p53) and have an impact on the development of novel therapies, including oncolytic viral agents, for the treatment of cancer. PMID- 15661530 TI - p53: a heavily dictated dictator of life and death. AB - In 2003, a p53-expressing adenovirus was approved as a cancer therapy drug in China. Consequently, there has been a surge in the need to understand the regulation of wild type p53 function in vivo. The majority of the progress made during the past two years has focused on the cellular factors and post translational modifications that regulate the expression levels and activities of p53 in response to stress signals. PMID- 15661531 TI - PP2A: the expected tumor suppressor. AB - PP2A is one of the few serine/threonine-specific phosphatases in the cell, and its complex structure and regulation guarantees its many different functions. Some viruses have chosen to target this enzyme system in order to manage the host cell machinery for their own profit and to program cells into a malignant state. Suppression of PR61/B'gamma, a specific third regulatory subunit of PP2A, can substitute for the viral SV40 protein small t antigen in causing tumorigenic transformation of several human cell lines -- provided that telomerase, SV40 large T antigen and oncogenic Ras are also present. Accumulation of c-Myc seems to be the common denominator. PMID- 15661532 TI - New and unexpected: forkhead meets ARF. AB - Recent genetic studies demonstrate that mice deficient in the forkhead box m1b (Foxm1b) transcription factor are highly resistant to developing hepatocellular carcinoma, which is among the most lethal cancers worldwide. In addition, the Foxm1b transcription factor was identified as a novel inhibitory target of the p19ARF tumor suppressor during early stages of liver tumorigenesis, but p19ARF expression is extinguished in hepatic tumors that develop at later stages. Structure-function studies demonstrate that amino acids 26-46 of the p19ARF protein are sufficient to bind Foxm1b and reduce Foxm1b transcriptional activity by targeting it to the nucleolus. A peptide containing amino acids 24-46 of p19ARF, which was modified to enhance cellular uptake, is an effective inhibitor of Foxm1b transcriptional activity and prevents Foxm1b stimulation of anchorage independent growth of cells on soft agar. Thus, the p19ARF peptide is an effective inhibitor of Foxm1b and represents a potential therapy for hepatocellular carcinoma. PMID- 15661533 TI - OVCA1: tumor suppressor gene. AB - OVCA1, also known as DPH2L1, is a tumor suppressor gene associated with ovarian carcinoma and other tumors. Ovca1 homozygous mutant mice die at birth with developmental delay and cell-autonomous proliferation defects. Ovca1 heterozygous mutant mice are tumor-prone but rarely develop ovarian tumors. OVCA1 appears to be the homolog of yeast DPH2, which participates in the first biosynthetic step of diphthamide, by modification of histidine on translation elongation factor 2 (EF-2). Yeast dph2 mutants are resistant to diphtheria toxin, which catalyses ADP ribosylation of EF-2 at diphthamide. Thus, there appears to be growing evidence implicating alterations in protein translation with tumorigenesis. PMID- 15661534 TI - Hsp90 and Cdc37 -- a chaperone cancer conspiracy. AB - The Hsp90 molecular chaperone system is involved in the activation of an important set of cell regulatory proteins, including many whose disregulation drives cancer. Recruitment of protein kinases to the Hsp90 system is mediated by the co-chaperone adaptor Cdc37 -- an essential protein whose overexpression is itself, oncogenic. Current structural, biochemical and biological studies of Cdc37 are beginning to unravel the nature of its interactions with Hsp90 and protein kinase clients, and implicate it as a key permissive factor in cell transformation by disregulated protein kinases. The central role of the Hsp90 Cdc37 chaperone complex makes it an important target for future anti-cancer drug development. PMID- 15661535 TI - Targeting Ras and Rho GTPases as opportunities for cancer therapeutics. AB - The Ras and Rho GTPases contribute to the initiation and progression of cancer by subverting the normal regulation of specific intracellular signalling pathways. As a result, Ras and Rho play significant roles in the development of numerous aspects of the malignant phenotype by promoting cell cycle progression, resistance to apoptotic stimuli, neo-vascularisation and tumour cell motility, invasiveness and metastasis. With these GTPases contributing at so many levels, they are appealing targets for the development of cancer chemotherapeutic agents. PMID- 15661536 TI - TSC1-2 tumour suppressor and regulation of mTOR signalling: linking cell growth and proliferation? AB - Understanding the relationship between growth and proliferation in multicellular organisms requires identification of the key regulators of growth control, and an understanding of how they regulate growth and how growth is linked to cell proliferation. Recent progress in understanding the mechanisms of growth control indicates that the tuberous sclerosis complex tumour-suppressor TSC1-2 serves as a point of integration between growth-stimulatory and growth-suppressive signalling upstream of a small GTPase, Rheb. However, Rheb-induced growth might not explain the additional effects of TSC1-2 upon cell proliferation. PMID- 15661537 TI - Targeting key steps in metastatic tumour progression. AB - The key determinants of tumour progression and discriminators of benign and malignant lesions include neoangiogenesis (the induction of a new blood supply) and the capacity of malignant cells to invade and metastasise. It is now recognized that these processes can be co-ordinately regulated by the activity of specific genes -- often distinct from those involved in early oncogenesis -- and involve common signalling pathways. Cell motility and chemotaxis (the ability to respond to gradients of chemoattractants) are implicated in both tumour-cell invasion and response of activated endothelial cells to angiogenic cytokines, and provide interesting and novel points for therapeutic intervention. PMID- 15661538 TI - Mechanisms of cancer cell invasion. AB - The movement of cancer cells into tissue surrounding the tumour and the vasculature is the first step in the spread of metastatic cancers. Recent advances in imaging, the use of 3D model systems and the application of microarray technologies have yielded new insights into these processes. This work has challenged our views about what causes cancer cells to become motile in the first place, and has demonstrated that cancer cells can move in many different ways. PMID- 15661539 TI - Tumor-stroma interactions. AB - The importance of stromal cells and the factors that they express during cancer initiation and progression has been highlighted by recent literature. The cellular components of the stroma of epithelial tissues are well-recognized as having a supportive role in carcinogenesis, where the initiating mutations of a tumor originate in the epithelial cells. The use of mouse models and xenografts suggests that mutations in the stromal fibroblasts can also initiate epithelial tumors. Many of these tumors result from the alteration of paracrine growth factor pathways that act on the epithelia. However, the tissue specificity of the responses to the growth factors is a mystery not yet solved. PMID- 15661540 TI - Cellular abnormalities of blood vessels as targets in cancer. AB - Tumor blood vessels have multiple structural and functional abnormalities. They are unusually dynamic, and naturally undergo sprouting, proliferation, remodeling or regression. The vessels are irregularly shaped, tortuous, and lack the normal hierarchical arrangement of arterioles, capillaries and venules. Endothelial cells in tumors have abnormalities in gene expression, require growth factors for survival and have defective barrier function to plasma proteins. Pericytes on tumor vessels are also abnormal. Aberrant endothelial cells and pericytes generate defective basement membrane. Angiogenesis inhibitors can stop the growth of tumor vessels, prune existing vessels and normalize surviving vessels. Loss of endothelial cells is not necessarily accompanied by simultaneous loss of pericytes and surrounding basement membrane, which together can then provide a scaffold for regrowth of tumor vessels. Rapid vascular regrowth reflects the ongoing drive for angiogenesis and bizarre microenvironment in tumors that promote vascular abnormalities and thereby create therapeutic targets. PMID- 15661541 TI - Phase II trials of drug combinations: fairytales or fact? PMID- 15661542 TI - Local recurrence in the axilla after sentinel node biopsy. PMID- 15661543 TI - Chemotherapy-induced nausea and vomiting: current and new standards in the antiemetic prophylaxis and treatment. AB - Nausea and vomiting are considered as two of the most distressing side-effects of chemotherapy. Chemotherapy-induced nausea and vomiting have been classified into acute, delayed and anticipatory based on the time of onset. The frequency of nausea and vomiting depends primarily on the emetogenic potential of the chemotherapeutic agents used. With the introduction of the 5-HT3 receptor antagonists in combination with dexamethasone in the early 1990s approximately 70% of patients receiving highly emetogenic chemotherapy were protected from acute emesis. However, 40% of patients have symptoms in the delayed phase. Another group of antiemetics, the neurokinin-1-receptor-antagonists, have recently been introduced. The addition of neurokinin receptor (NK1 receptor) antagonists to standard therapy significantly improves emesis protection in the acute and in particular in the delayed phase by approximately 20%. Due to these new developments, revised antiemetic guidelines have been set. Here, the most recent developments in antiemetic therapy, including these guidelines, are reviewed. PMID- 15661544 TI - Intracellular signalling and cancer: complex pathways lead to multiple targets. AB - Normal cells proliferate, die and differentiate as and when they should for the proper functioning of any particular tissue type. These processes are governed by a complex series of intracellular pathways that have many internal checkpoints and safety nets. These ensuring a fine, but tight, balance on overall tissue growth and distribution. A series of key aberrations, resulting in the disruption of these intracellular pathways, can lead to the development of a malignancy. The nature of these alterations is often not only tumour-specific, but also different between individuals with the same tumour type. As a result, these pathways have to be carefully dissected and functionally assessed to identify valid targets with therapeutic potential in a wide range of tumour types. PMID- 15661545 TI - Evaluation and outcome of behavioural changes in the rehabilitation of cancer patients: a review. AB - The global increase in the number of newly diagnosed cancers has led in most affected countries to increased numbers of cancer survivors, who have specific needs for physical and psychosocial rehabilitation. In spite of recent progress, little is known about the specific rehabilitation measures that could increase the quality of life for cancer survivors. We reviewed published interventions that focussed on changing known risk factors for cancer recurrence and improving physical well-being; those we selected were exercise, smoking, alcohol consumption, diet and the use of sun screens. The published trials varied in the quality of the methods used, often had inadequate sample sizes and showed difficulty in validating outcomes. We conclude that there is still insufficient evidence to assess the importance of these behavioural risk factors in the rehabilitation of cancer patients. Future interventions should be designed to assess the separate effects of dietary changes, exercise and psychosocial interventions. PMID- 15661546 TI - Doxorubicin and cisplatin chemotherapy in high-grade spindle cell sarcomas of the bone, other than osteosarcoma or malignant fibrous histiocytoma: a European Osteosarcoma Intergroup Study. AB - There are limited data that define the role of chemotherapy in the treatment of high-grade spindle cell sarcomas of bone, other than osteosarcoma or malignant fibrous histiocytoma (MFH-B). This prospective study evaluates the effect of doxorubicin and cisplatin on these tumours. Thirty-seven patients, age 65 years, with spindle cell sarcoma of bone, except osteosarcoma or MFH-B, were included. Chemotherapy consisted of doxorubicin and cisplatin every 3 weeks for six cycles. Resection was performed after three cycles. In 15 patients with metastases, response assessment showed three complete responses (CR), four stable disease (SD), five progression; three were not evaluable. Median time to progression was 30 months (95% Confidence Interval (CI), 8-51 months) for the operable non metastatic patients; median survival 41 months (95% CI, 16-82 months). Median time to progression in the metastatic group was 10 months (95% CI, 0-18 months) and median survival was 14 months (95% CI, 4-45 months). This study suggests a limited role for doxorubicin and cisplatin in metastatic high-grade spindle cell sarcoma of bone, other than osteosarcoma or MFH-B cases. PMID- 15661547 TI - Sentinel node biopsy in breast cancer: early results in 953 patients with negative sentinel node biopsy and no axillary dissection. AB - Sentinel node biopsy in patients with breast carcinoma accurately predicts the axillary nodal status. However, in some 6% of patients with negative sentinel nodes the remaining axillary nodes harbour metastases. Our purpose was to observe a large number of patients who did not undergo an axillary dissection after a negative sentinel node biopsy for the appearance of overt axillary metastases. 953 patients treated from 1996 to 2000, with negative sentinel nodes not submitted to axillary dissection, were followed-up to 7 years, with a median follow-up of 38 months. Fifty-five unfavourable events occurred among the 953 patients, 37 (4%) related to the primary breast carcinoma. Three cases of overt axillary metastases were found: they received total axillary dissection and are presently alive and well. The 5 year overall survival rate of the whole series was 98%. Patients with negative sentinel node biopsies not submitted to axillary dissection show during follow-up a rate of overt axillary metastases that is lower than that expected. PMID- 15661548 TI - Role of routine preoperative lymphoscintigraphy in sentinel node biopsy for breast cancer. AB - Sentinel node biopsy (SNB) is rapidly emerging as the preferred technique for nodal staging in breast cancer. When radioactive colloid is used, a preoperative lymphoscintiscan is obtained to ease sentinel lymph node (SN) identification. This study evaluates whether preoperative lymphoscintigraphy adds diagnostic accuracy to offset the additional time and cost required. 823 breast cancer patients underwent SNB based on lymphoscintigraphy, intraoperative gamma probe detection, and blue dye mapping using 99 mTc-nanocolloid and Patent Blue V injected peritumourally. The SNB was followed by standard axillary treatment at the same operation. Preoperative lymphoscintigraphy was performed around 3 h after the radioisotope injection. Preoperative lymphoscintigraphy revealed SNs in 593 (72%) of the 823 patients imaged. SN visualisation on lymphoscintigraphy was less successful in large tumours and tumours involving the upper outer quadrant of the breast (P=0.046, P<0.001, respectively). Lymphoscintigraphy showed internal mammary sentinel nodes in 9% (62/707) patients. The SN was identified intraoperatively in 98% (581) patients who had SN visualised on preoperative lymphoscintigraphy, with a false-negative rate of 7%. In patients who did not have SN visualised on preoperative lymphoscintigraphy, the SN was identified at operation in 90% (204) patients, with a false-negative rate of 7%. The SN identification rate was significantly higher in patients with SN visualised on preoperative lymphoscintigraphy (P<0.001). SN identification rate intraoperatively using the gamma probe was significantly higher in the SN visualised group compared with the SN non-visualised group (95% vs. 68%; chi square (1 degrees of freedom (df)) P<0.001. There was no statistically significant difference in the false-negative rate and the operative time between the two groups. A mean of 2.3 (standard deviation (SD) 1.3) SNs per patient were removed in patients with SN visualised on preoperative lymphoscintigraphy compared with 1.8 (SD 1.2) in patients with no SN visualised on lymphoscintigraphy (P<0.001). Although SN visualisation on preoperative lymphoscintigraphy significantly improved the intraoperative SN localisation rate, SN was successfully identified in 90% of patients with no SN visualisation on lymphoscintigraphy. Given the time and cost required to perform routine preoperative lymphoscintigraphy, these data suggest that it may not be necessary in all cases. It may be valuable for surgeons in the learning phase to shorten the learning curve and in patients who have increased risk of intraoperative failed localisation (obese or old patients). A negative preoperative lymphoscintiscan predicts the inability to localise with the hand-held gamma probe. Therefore, if the SN is not visualised on lymphoscintigraphy then the addition of intraoperative blue dye is recommended to increase the likelihood of SN identification. PMID- 15661549 TI - Minimal risk of macrometastases in the non-sentinel axillary lymph nodes in breast cancer patients with micrometastatic sentinel lymph nodes and preoperatively ultrasonically uninvolved axillary lymph nodes. AB - Micrometastases in the sentinel lymph node (SLN) carry a considerable risk of macrometastases in the non-sentinel lymph nodes (NSLN), resulting in axillary lymph node dissection (ALND). Preoperative ultrasound (US) examination of the axillary lymph nodes combined with a fine-needle aspiration biopsy (FNAB) has been proved to discover metastases in the axillary lymph nodes. The aim of our study was to assess the risk of macrometastases in NSLN in patients with micrometastatic SLN after a preoperative US examination of the axillary lymph nodes. The study included 36 patients in whom, after preoperative axillary US, micrometastases in the SLN were revealed and ALND was subsequently performed. At final histopathology, no macrometastases were discovered in the NSLN. In four patients, additional micrometastases were discovered in the NSLN. In conclusion, the risk of macrometastases in the NSLN in patients with preoperatively ultrasonically uninvolved axillary lymph nodes is minimal. PMID- 15661550 TI - Combination regimen of epirubicin, vinorelbine and 5-fluorouracil continuous infusion as first-line chemotherapy in anthracycline-naive metastatic breast cancer patients. AB - We investigated the activity and toxicity of a combination of vinorelbine 25 mg/m2 on days 1 and 15; epirubicin 25 mg/m2 on days 1, 8, 15; and 5-fluorouracil continuous infusion at 200 mg/m2 every day, administered as first-line chemotherapy in anthracycline-naive metastatic breast cancer patients. Fifty three patients entered the study. Cycles were repeated every 28 days. Objective response was 60% by World Health Organisation (WHO) criteria and 63% by Response Evaluation Criteria in Solid Tumours (RECIST). The median time to progression was 12.7 months (17.6 months in responders) and the median survival duration was 32.9 months. The dose-limiting toxicity was leucopenia (grade 3/4 in 36% of patients). Grade 3/4 non-haematological toxicities included mucositis in 11% of patients, skin and cardiac toxicity in 4% and 2%, respectively. The combination of vinorelbine, epirubicin and 5-fluorouracil continuous infusion was found to be an active and manageable first-line regimen for metastatic breast cancer patients. PMID- 15661551 TI - Two years of adjuvant tamoxifen in premenopausal patients with breast cancer: a randomised, controlled trial with long-term follow-up. AB - Adjuvant tamoxifen treatment increases recurrence-free survival (RFS) and overall survival (OS) in early breast cancer, although in premenopausal patients the number of studies comparing tamoxifen vs no treatment are limited. We report herein the effect on RFS of adjuvant tamoxifen treatment in a multicentre trial of premenopausal patients with stage II breast cancer patients randomised between 1986 and 1991 to 2 years of tamoxifen treatment (n=276) or no treatment (n=288). The receptor status of the tumour was known for 541 (96%) of the patients included. Tamoxifen treatment significantly increased RFS in patients with hormone receptor-positive (oestrogen receptor-positive (ER+) and/or progesterone receptor-positive (PR+)) tumours (Relative Risk (RR) 0.65; 95% Confidence Interval (CI): 0.48-0.89, P=0.006), and the beneficial effect of tamoxifen was extended to patients with indicators of poor prognosis, such as young age and nodal-positivity. PR status was a significant predictor of response to tamoxifen in multivariate models with testing of interactions of hormone receptor status and adjuvant therapy. PMID- 15661552 TI - The role of early expression of inducible nitric oxide synthase in human breast cancer. AB - Nitric oxide synthases are expressed in breast cancer. To elucidate the clinical role of the inducible NOS (i-NOS) in human breast cancer, 161 primary breast cancer tissues were stained immunohistochemically. Staining patterns for i-NOS were correlated with classical prognostic factors such as lymph node status, age, hormonal receptor status, tumour size and tumour differentiation. With classical prognostic factors such as lymph node status, age, hormonal receptor status, tumour size and tumour differentiation. Patients survival was also analysed. Sixty-one percent of the tumours stained positively for i-NOS. Detection of i-NOS was positively correlated with increasing tumour size and decreasing tumour differentiation (P=0.018 and P=0.039, respectively). However, in the 50 year age group, i-NOS staining also correlated with lymph node status. Patients with i-NOS positive breast carcinomas had a significantly worse overall survival rate versus those with negative stains (5-year survival rate 84.8% versus 67.1%; P=0.049; log rank test). To date, this is the largest analysis of i-NOS expression in breast cancer patients and the only study to assess survival. PMID- 15661553 TI - Number of lymph nodes examined and prognosis of TNM stage II colorectal cancer. AB - The diagnosis of a lymph node-negative colorectal carcinoma should imply a good prognosis; however, the outcomes for TNM stage II patients remain variable. Few studies have examined the relationship of the number of lymph nodes examined to the prognosis of this stage. The aim of this study was to determine whether the number of lymph nodes examined has an effect on prognosis of a relatively large sample of patients undergoing curative surgery for stage II colorectal cancer at a single institution. Data on patients who underwent surgery for colorectal cancer between January 1980 and April 2000 were prospectively collected in a database. Patients with TNM stage II or stage III tumours who were treated with curative intent were removed. Patients over 80 years of age were excluded from the survival analysis. Survival comparisons were made using Kaplan-Meier curves and the log-rank test. Multivariate analysis was performed using a Cox regression model. A total of 625 cases of TNM stage II cases and, for comparison purposes, 415 stage III cases, were analysed. Lymph node retrieval in stage II cases was affected by the patient's age (P=0.04) and gender (P=0.02), tumour grade (P<0.0001), tumour site (P<0.0001), and necessity to carry out extended resection (P<0.0001). In stage III cases, lymph node retrieval was affected by patient age (P<0.0001), tumour grade (P=0.02), and tumour site (P=0.002). Decreased lymph node detection was associated with increasing hazard ratios among the 480 TNM stage II patients under 80 years of age, but not among the 345 patients with TNM stage III tumours. Five year survival rate for patients with stage III tumours with only 1-3 positive lymph nodes (52.6%) was similar to that of patients with stage II tumour who had nine or fewer lymph nodes examined (51.3%). These results demonstrate that the prognosis of TNM stage II colorectal cancer is dependent on the number of lymph nodes examined. Patients with few nodes examined have a poorer prognosis. It is possible that a smaller number of lymph nodes examined reflects a diminished immune response. It can be presumed that those patients with stage II tumour with only a few nodes examined should be offered postoperative chemotherapy on a routine basis. PMID- 15661554 TI - Analysis and interpretation of health-related quality-of-life data from clinical trials: basic approach of The National Cancer Institute of Canada Clinical Trials Group. AB - Clinicians are being confronted with increasing amounts of health-related quality of life (HRQOL) data. Thus, there is a need for a greater understanding of the analysis and interpretation of HRQOL so that the data can be reported in an appropriate manner. The approach of the Clinical Trials Group of the National Cancer Institute of Canada emphasises the clinical meaning of the results, while avoiding complex statistical modelling. It consists of four steps: calculating the questionnaire completion rates, calculating the baseline scores, determining the individual change in scores over time for the domains specified in the trial hypothesis, and culminates in determining the proportions of patients who have reported clinically meaningful changes in scores since baseline. A rationale supporting each step is given. This approach is presented as a simple and practical aid to the analysis, interpretation and reporting of HRQOL results. PMID- 15661555 TI - Stress response symptoms in adolescent and young adult children of parents diagnosed with cancer. AB - The aim of this study was to assess stress response symptoms in children of parents diagnosed with cancer 1-5 year prior to study entry. The impact of event scale was used to measure stress response symptoms in terms of intrusion and avoidance; the youth self-report assessed emotional and behavioural functioning; the state-trait anxiety inventory for children measured trait-anxiety. Participants included 220 adolescents (aged 11-18 years) and 64 young adults (aged 19-23 years) from 169 families. Twenty-one percent of the sons and 35% of the daughters reported clinically elevated stress response symptoms. Daughters, particularly those whose mothers were ill, reported significantly more intrusion and avoidance than did sons. Intrusion among daughters was positively related to age. Stress response symptoms in both sons and daughters were significantly associated with trait anxiety, but not with intensity of treatment or time since diagnosis. Daughters whose parents suffered from recurrent illness reported more symptoms than did daughters whose parents had a primary disease. Children (daughters in particular) with clinically elevated stress response symptoms reported significantly more problems of internalising and cognition than did their norm group peers. One-fifth of the sons and more than one-third of the daughters expressed clinically elevated stress response symptoms. These children also reported internalising and cognitive problems. Daughters appeared to be more at risk than sons. PMID- 15661556 TI - Prevalence of human papillomavirus infection in women in Turin, Italy. AB - Human papillomavirus (HPV) is recognised as necessary for the development of cervical cancer. An age-stratified random sample of 1013 women, aged 25-70 years, participating in the organised cervical screening programme in Turin, Italy was tested for 36 HPV types using polymerase chain reaction (PCR) with the general primers GP5+/GP6+. The overall HPV prevalence was 8.8%. High-risk types were found in 7.1% of women and multiple infections in 1.1%. HPV-16 was the most common type (32.6% of HPV-positive women). HPV prevalence (any type) was 13-14% at ages 25-39 years, 11.5% at age 40-44 years, and approximately 5% among older women. After age-adjustment, HPV prevalence was significantly increased in single vs married, (Odds Ratio (OR)=2.23; 95% Confidence Interval (CI): 1.28-3.89) and decreased in parous vs nulliparous women (OR=0.49; 95% CI: 0.31-0.78). However, the association with marital status and parity was restricted to women less than 45 years of age. In conclusion, overall, the female population of Turin showed an HPV prevalence that is intermediate compared with worldwide levels. PMID- 15661557 TI - Association of CK19 mRNA detection of occult cancer cells in mediastinal lymph nodes in non-small cell lung carcinoma and high risk of early recurrence. AB - This study was designed to screen occult cancer cells by CK19 mRNA detection using reverse transcriptase-polymerase chain reaction (RT-PCR) in mediastinal lymph nodes stations (MLNS) in non-small cell lung carcinoma (NSCLC). In 49 NSCLC patients free of mediastinal adenopathy on computed tomograph, 254 MLNS were evaluated by histopathology, immunohistochemistry (IHC) and RT-PCR. Of 225 non tumoral MLNS on histopathology, 32 (14.2%) were positive by RT-PCR. IHC did not provide significant additional results. Seventeen patients were without mediastinal tumoral extension on histopathology and RT-PCR (Group 1), 16 were upgraded by RT-PCR (Group 2) and 16 pN2 on histopathology (Group 3). The two-year cancer-related death survival in Groups 1 (100%) and 2 (64.5%) was significantly different (P=0.04). The relative risk of recurrence in Group 2 compared with Group 1, evaluated by the Cox model multivariate analysis, was 5.61 (P=0.02). In conclusion, CK19 mRNA detected by RT-PCR in MLNS was significantly associated with an increased risk of rapid recurrence. PMID- 15661558 TI - Combination of mutated herpes simplex virus type 1 (G207 virus) with radiation for the treatment of squamous cell carcinoma of the head and neck. AB - G207 is an oncolytic herpes simplex virus (HSV) with deletions at both gamma134.5 loci and a LacZ gene insertion inactivating the HSV ribonucleotide reductase gene. Ionising radiation induces the growth arrest-inducible gene, GADD34, and ribonucleotide reductase. GADD34 is a protein that correlates with apoptosis following radiation and has homology with the G207 gamma134.5 gene. We hypothesised that the combination of radiotherapy with G207 may have a potentiating effect on viral replication and anti-tumour efficacy. The purpose of this study was therefore to evaluate the combination of G207 with radiation therapy to treat head and neck tumours. The cytotoxicity of G207 was tested in six head and neck squamous carcinoma cell lines, in the presence or absence of irradiation. For in vivo experiments, flank tumours in C3H/HeJ mice or in nude mice were treated with direct injections of G207, with or without radiation. All head and neck squamous cancer cell lines tested demonstrated significantly increased antitumour effects with the combination of G207 virus and radiation therapy compared with each individual modality (P<0.01). Furthermore, the combination treatment effect was better than the expected additive effect of the two therapies in combination. Even the radiation-resistant cell lines (SCC25, MSKQLL2, SCCVII) were susceptible. The combination of direct G207 injection with radiation therapy suppressed human and murine squamous cell carcinoma growth significantly (P<0.05 and P<0.001) compared with controls or single modality therapy. G207 enhanced the effectiveness of radiation therapy and low-dose radiation potentiated the effectiveness of G207 viral therapy in head and neck cancer. These findings suggest a potential clinical application for this combined therapy as treatment for radiation-resistant head and neck cancers. PMID- 15661560 TI - The new dimension of oncology. Teleoncology ante portas. AB - Telemedicine is defined as the practice of medicine from a distance, without the doctor's physical presence. Teleoncology consist of telediagnosis, teleconsultation, telepathology, telesurgery, distant help in computer-aided radiotherapy planning and telefollow-up. In this paper, we present the wide range of possibilities in providing cancer care at a distance. The theoretical and practical aspects of teleoncology and its current as well as future role in a multidisciplinary approach to cancer patients are discussed. PMID- 15661559 TI - Molecular characterisation of two human cancer cell lines selected in vitro for their chemotherapeutic drug resistance to ET-743. AB - ET-743 (Yondelis(TM), Trabectedin) isolated from the tunicate Ecteinascidia turbinata, is being tested in phase II clinical trials in Europe and the United States of America (USA). Studies with different solid tumours have shown antitumour activity in advanced, pre-treated sarcomas as well as in drug resistant breast and ovarian cancer. The primary mechanism of action for ET-743 has not been fully elucidated and different models have been suggested to explain its molecular mechanism of action. ET-743 binds tightly to the minor groove of DNA and previous data have suggested that ET-743 acts by interfering with RNA transcription. To further investigate the mechanism of in vitro drug resistance, we evaluated the gene expression profile in ovarian and chondrosarcoma cell lines selected for resistance to ET-743. We found 70 genes whose expression was modulated in both drug-resistant cell lines when compared with their respective parental drug-sensitive cell lines. This pattern of gene expression seems to be selective for ET-743-resistant cells, since ovarian cancer cells resistant to paclitaxel did not share the same gene expression changes. Data presented in this study reveal different molecular pathways that could be involved in the cellular mechanism of ET-743 resistance. PMID- 15661561 TI - The dilemma of the strive for apoptosis in oncology: mind the heart. AB - In recent years, apoptosis has increasingly drawn the attention of both oncologists and cardiologists alike. Anticancer treatment is possible by induction of apoptosis in cancer cells, and targeted anticancer drugs are being developed to promote this. However, since these drugs usually are not selective for malignant cells, side effects on non-cancerous tissue, such as the myocardium must be anticipated. Since apoptosis is a pathophysiological mechanism in cardiac diseases leading to heart failure, cardiologists in contrast to oncologists, aim at preventing apoptosis in the heart. The purpose of this review is to describe new insights in mechanisms of cardiomyocyte apoptosis. In addition to the mitochondrial and death receptor apoptotic pathways, apoptosis through lack or inhibition of growth factor receptor-mediated signalling is discussed. Exploration of the apoptotic pathways in the heart can contribute to the safer use of new anticancer drugs and to the development of new therapies for heart failure. PMID- 15661562 TI - When hypoxia signalling meets the ubiquitin-proteasomal pathway, new targets for cancer therapy. AB - The ubiquitin-proteasomal pathway of degradation of proteins is activated or repressed in response to a number of environmental stresses and thereby plays an essential role in cell function and survival. Hypoxic stress, resulting from a decrease in the concentration of oxygen in tissues, is encountered in both physiological and pathological situations, in particular in cancer. The transcriptional complex hypoxia-inducible factor (HIF) is the key player in the signalling pathway that controls the hypoxic response of mammalian cells. Under hypoxic conditions it transactivates an impressive number of genes involved in a multitude of cellular functions. Tight regulation of this response in part involves the ubiquitin-proteasomal system where oxygen-dependent prolyl-4 hydroxylation of the alpha subunit of HIF triggers a cascade of events that leads to its degradation by the 26S proteasome. Inhibition of the proteasome in conjunction with topoisomerase inhibition has shown some promise in the treatment of experimental cancer. Such treatment may impact on the hypoxic adaptation of tumour cells. PMID- 15661563 TI - Incidence of febrile neutropenia and neutropenic infections in elderly patients receiving anthracycline-based chemotherapy for breast cancer without primary prophylaxis with colony-stimulating factors. AB - There is concern about the potential increase of hematological toxicity in elderly patients treated with chemotherapy. Recently, primary prophylaxis with colony-stimulating factors (CSFs) was proposed for elderly patients receiving moderately toxic chemotherapy. However, evidence for the benefits of this primary prophylaxis for elderly breast cancer patients is currently lacking. We retrospectively analyzed the incidence of febrile neutropenia (FN) and neutropenic infections in elderly breast cancer patients receiving anthracycline based chemotherapy without primary prophylaxis with colony-stimulating factors. In addition, we assessed the direct costs of hospitalization for these complications. Febrile neutropenia or neutropenic infection occurred in 13% of the 46 patients. Further studies are needed to adequately evaluate the risk of neutropenic complications (NC) in elderly patients receiving standard-dose chemotherapy for breast cancer and the potential benefits of primary prophylaxis with colony-stimulating factors. PMID- 15661564 TI - A tailored regimen including capecitabine and oxaliplatin for treating elderly patients with metastatic colorectal carcinoma Southern Italy Cooperative Oncology Group trial 0108. AB - From September 2001 to November 2002, 35 patients aged 70-81 (median, 75) years, with measurable metastatic lesions from colorectal carcinoma, were treated with a combination of oxaliplatin (OXA) infused i.v. over 2 h on day 1, and capecitabine, assumed orally twice a day (12-h apart) from day 2 to day 15. An alternated dose escalation for both drugs was planned over the first three cycles for each patient, in the absence of WHO grade > or =2 toxicity on previous cycle: starting doses were 85 mg/m2 for OXA, and 2000 mg/m2 (day) for capecitabine on first cycle; on second cycle, OXA was planned at 100mg/m2, while capecitabine was planned at 2500 mg/(m2 day) on third cycle. Treatment was repeated every 3 weeks until progression, or for a maximum of 12 cycles. A total of 212 cycles were administered, with a median of 6 (range, 1-12) cycles/patient. Dose escalation was performed in 18 (51%) patients for OXA, and in 4 (11%) patients for capecitabine. No grade 4, and 10 (29%) cases of grade 3 toxicity of any type were reported. Abdominal symptoms (pain, nausea, or vomiting) affected 66% of patients, but they were of grade 3 in only 2 (6%) patients. Grade 3 diarrhoea occurred in 3 (9%) patients. Two complete and 12 partial responses (PR) were reported, for an overall response rate of 40% (95% CI, 24-58%). Progression of disease occurred in 23 (66%) patients, and 18 (51%) died. The actuarial median progression-free and survival time were 6.9 and 14.1 months, respectively. PMID- 15661565 TI - Germ cell tumours of the testis. AB - Cancer of the testis is a relatively rare disease, accounting for about 1% of all cancers in men. Cryptorchidism is the only confirmed risk factor for testicular germ cell tumour. The majority of GCT are clinically detectable at initial presentation. Any nodular, hard, or fixed area discovered in the testis, must be considered neoplastic until proved otherwise. The appropriate surgical procedure to make the diagnosis is a radical orchidectomy through an inguinal incision. Many GCT produce tumoural markers (AFP, HCG, LDH), who are useful in the diagnosis and staging of disease; to monitor the therapeutic response and to detect tumour recurrence. In 1997 a prognostic factor-based classification for the metastatic germ cell tumours was developed by the IGCCCG: good, intermediate and poor prognosis, with 5-year survival of 91, 79 and 48%, respectively. GCT of the testis is a highly table, often curable, cancer. Germ cell testicular cancers are divided into seminoma and non-seminoma types for treatment planning because seminomatous testicular cancers are more sensitive to radiotherapy. Seminoma (all stages combined) has a cure rate of greater than 90%. For patients with low-stage disease, the cure approaches 100%. For patients with non-seminoma tumours, the cure rate is >95% in stages I and II; it is approximately 70% with standard chemotherapy and resection of residual disease, if necessary, in stages III and IV. Minimum guidelines for clinical, biochemical, and radiological follow-up have been reported by ESMO in 2001. PMID- 15661566 TI - Cancer of the penis. AB - Cancer of the penis is rare in Europe, accounting for less than 0.5% of all cancers. Phimosis and poor hygiene are strong risk factors whereas neonatal circumcision is a contributing factor in the prevention of this disease. More than 95% of penile carcinomas are squamous cell carcinomas. Early disease (stage I-II) is curable in most patients, who can be treated by conventional penile amputation or, in selected cases, by organ preserving techniques, including Moh's micrographic surgery, laser ablation or radiation therapy (external-beam, brachytherapy). For more advanced primary tumours, penile amputation is required. Survival of patients with penile cancer is strongly related to the presence and extent of nodal metastases. Bilateral inguinal lymphadenectomy is recommended for palpable lymph nodes that persist 3 or more weeks after removal of the primary tumour and a course of antibiotic therapy. In patients with proven inguinal lymph node metastases, bilateral ileoinguinal dissection should be performed. When the nodes are clinically negative, "prophylactic" inguinal lymphadenectomy may be a reasonable approach in patients with invasive tumours (T2 or greater), high grade tumours, or tumours exhibiting vascular invasion. The role of chemotherapy, as adjuvant and neoadjuvant or primary treatment in metastatic disease, needs to be further explored in prospective clinical trials. PMID- 15661567 TI - Novel effect of Y-24180, a presumed specific platelet-activating factor receptor antagonist, on Ca2+ levels and growth of human osteosarcoma cells. AB - In human osteosarcoma MG63 cells, the effect of Y-24180, a presumed specific platelet-activating factor (PAF) receptor antagonist, on intracellular Ca(2+) concentration ([Ca(2+)](i)) and proliferation was measured by using fura-2 and tetrazolium as fluorescent dyes, respectively. Y-24180 (1-5 microM) caused a rapid and sustained [Ca(2+)](i) rise in a concentration-dependent manner. The [Ca(2+)](i) rise was inhibited by 35% by dihydropyridines or removal of extracellular Ca(2+), but was not altered by verapamil and diltiazem. In Ca(2+) free medium, thapsigargin, an inhibitor of the endoplasmic reticulum Ca(2+) ATPase, caused a monophasic [Ca(2+)](i) rise, after which 5 microM Y-24180 failed to increase [Ca(2+)](i); conversely, depletion of Ca(2+) stores with 5 microM Y 24180 abolished thapsigargin-induced [Ca(2+)](i) rise. U73122, an inhibitor of phoispholipase C, inhibited histamine-induced, but not 5 microM Y-24180-induced [Ca(2+)](i) rise. Overnight treatment with 0.1-5 microM Y-24180 inhibited cell proliferation in a concentration-dependent manner. Together, these findings suggest that Y-24180 acts as a potent and cytotoxic Ca(2+) mobilizer in human osteosarcoma cells, by inducing both extracellular Ca(2+) influx and intracellular Ca(2+) release. Alterations in cytosolic Ca(2+) regulation may lead to interferences of various cellular functions; thus, attention should be exercised in using Y-24180 as a selective PAF receptor antagonist. PMID- 15661568 TI - Adenosine receptor mediates nicotine-induced antinociception in formalin test. AB - In this study, the effect of adenosine receptor agents on nicotine induced antinociception, in formalin test, has been investigated. Intraperitoneal (i.p.) administration of different doses of nicotine (0.1, 1, 10 and 100 microgkg(-1)) induced a dose-dependent antinociception in mice, in the both first and second phases of the test. Adenosine receptor antagonist, theophylline (5, 10, 20 and 80 mgkg(-1), i.p.) also induced antinociception in the both phases, while a dose of the drug (40 mgkg(-1), i.p.) did not induce any response. Theophylline reduced antinociception induced by nicotine in both phases of formalin test. The A(2) receptor agonist, 5'-N-ethylcarboxamide adenosine (NECA; 1 and 5 microgkg(-1), i.p.) also produced antinociception, which was reversed with different doses of theophylline (5, 10, 20 and 40 mgkg(-1), i.p.). But administration of the adenosine receptor agonist, NECA did not potentiate the response of nicotine. It is concluded that adenosine system may be involved in modulation of antinociception induced by nicotine. PMID- 15661569 TI - Effect of scutellarin on nitric oxide production in early stages of neuron damage induced by hydrogen peroxide. AB - The aims of the present study were to investigate the regulatory function of scutellarin on production of nitric oxide (NO) as well as activities of constitutive NO synthase (cNOS) and inducible NO synthase (iNOS) in early stages of neuron damage induced by hydrogen peroxide. Direct detection of NO production was performed on primary cultures of living rat neuronal cells with an electrochemical sensor. Hydrogen peroxide significantly increased culture supernatant levels of NO, the total integral value of the defined areas (500-6500 sxpA) reached 3.68 x 10(6). Pre-treatment with scutellarin, caused the total integral value to decrease in a dose-dependent fashion (3.24 x 10(6), 2.15 x 10(6), 1.84 x 10(6) for groups 10, 50, and 100 uM scutellarin, respectively). After exposure to 2.0mM hydrogen peroxide for 2h, malondialdehyde (MDA) level, a marker of lipid peroxidation, was remarkably increased. The elevation can be suppressed by scutellarin. Hydrogen peroxide also caused significant loss of neuron viability. In comparison with the control group, scutellarin significant attenuated the loss. Results also showed that hydrogen peroxide increased activity of cNOS, which was markedly inhibited by scutellarin. However, exposure of neuronal cells to hydrogen peroxide did not lead to an increase in iNOS activity. In conclusion, our results suggest that NO production, which increased in early stages of neuron damage induced by hydrogen peroxide can be effectively inhibited by scutellarin. Moreover, our results indicate that increase in NO production is mediated by cNOS. PMID- 15661570 TI - Neuropsychiatric reactions to drugs: an analysis of spontaneous reports from general practitioners in Italy. AB - Limited information is available on drug-induced neuropsychiatric disorders in general practice. The spontaneous reports of both neurological and psychiatric adverse drug reactions (ADRs) collected during a 2-year period in PharmaSearch database, an Italian database recording reports of ADRs from general practitioners (GPs), were examined. Between January 2002 and December 2003, 171 general practitioners sent to PharmaSearch a total of 1131 reports corresponding to 1892 ADRs. Of overall reports, 310 (27.4%) involved the central nervous system resulting in 440 neuropsychiatric reactions (specifically, 241 neurological and 199 psychiatric). In our survey, 40 reports were excluded because they were incomplete or contradictory and thus classified as 'unlikely' or 'unclassifiable'. Therefore, the present analysis was carried out on 270 reports with 391 neuropsychiatric reactions (213 neurological and 178 psychiatric, respectively). Vertigo (16.4%), confusion (10.7%) and headache (10.0%) were the reactions more commonly reported. Drugs indicated for the treatment of nervous system disorders (ATC 1 code=N) accounted only for 38.4% of neuropsychiatric reactions, while most of these reactions were related to drugs indicated for other than nervous system diseases. Non-steroidal anti-inflammatory drugs (NSAIDs), fluoroquinolones, antidepressant drugs, opioids, and drugs for peptic ulcer were the categories most frequently suspected for neuropsychiatric reactions. Of 391 neuropsychiatric reactions, 78 (19.9%) were unlabeled and 41 (10.5%) were serious. In conclusion, the present study carried out in general practice underlines the importance of neuropsychiatric ADRs and reminds GPs to pay attention to this kind of toxicity when they prescribe pharmacological agents to their patients. PMID- 15661571 TI - Differential effects of echistatin and thrombin on collagen production and prolidase activity in human dermal fibroblasts and their possible implication in beta1-integrin-mediated signaling. AB - Prolidase [E.C. 3.4.13.9] is a cytosolic imidodipeptidase that plays an important role in collagen biosynthesis. The enzyme contributes to the recovery of proline from protein degradation products (mainly collagen) for collagen resynthesis. Prolidase activity and collagen biosynthesis are supposed to be regulated by beta(1)-integrins, which initiate a signaling pathway in which several kinases and intracellular proteins are involved, including focal adhesion kinase pp125(FAK) (FAK), Src, Shc, growth factor receptor bound protein 2 (Grb-2), son of sevenless protein (SOS), Ras, Raf and mitogen-activated protein kinases (MAPK), extracellular-signal regulated kinase 1 (ERK(1)) and kinase 2 (ERK(2)). We studied the effects of echistatin, a well-known disintegrin and thrombin, a serine protease capable of activation of platelet integrin alpha(2)beta(1) receptor on collagen production, prolidase activity, expression of prolidase, beta(1)-integrin receptor, FAK, SOS-protein and phosphorylated MAP-kinases (ERK(1) and ERK(2)) in confluent human dermal fibroblasts. It has been found that treatment of the cells with 100nM echistatin contributes to inhibition of collagen production, as well as prolidase activity and expression compared to control cells. These phenomena were accompanied by a decrease in the expression of FAK, SOS-protein and phosphorylated MAP-kinases, ERK(1) and ERK(2). An opposite phenomenon was observed in fibroblasts treated with 0.1IU thrombin. In this case, a significant increase in collagen production and prolidase activity, accompanied by a distinct raise in the expression of prolidase, FAK and phosphorylated MAP-kinases and a slight increase in expression of SOS compared to controls were found. The results suggest that regulation of prolidase activity and collagen biosynthesis in human dermal fibroblasts may involve beta(1) integrin-dependent signaling. PMID- 15661572 TI - Study of daunorubicin cardiotoxicity prevention with pyridoxal isonicotinoyl hydrazone in rabbits. AB - Risk of cardiotoxicity is the most serious drawback of the clinical usefulness of anthracycline antineoplastic antibiotics, which however, remain among the most powerful and widely employed anticancer drugs. In this study we have used daunorubicin-induced cardiomyopathy in rabbits as a model to investigate possible cardioprotective effects of pyridoxal isonicotinoyl hydrazone (PIH)-a principal representative of a novel group of aroylhydrazone iron chelators. Three groups of animals were used: a control group (n=11; i.v. saline), daunorubicin-treated animals (n=11; 3mg/kg, i.v.), and animals pretreated with PIH (n=9, 25 mg/kg, i.p.) 60 min before daunorubicin administration. All substances were administered once weekly for 10 weeks. Repeated administration of daunorubicin caused premature death in four animals and induced conspicuous histopathological changes in the myocardium, progressive and significant impairment of systolic heart function (a decrease in left ventricular dP/dt(max), ejection fraction, an increase in the pre-ejection period/left ventricular ejection time index), and a gradual increase in cardiac troponin T plasma concentrations. On the contrary, all the PIH-treated animals have survived all daunorubicin applications. Furthermore, in this group, the daunorubicin-induced cardiac changes were in most functional, biochemical as well as morphological parameters less pronounced than in the group receiving daunorubicin alone. Hence, PIH and other aroylhydrazones merit further investigation as potentially protective agents against anthracycline-induced cardiotoxicity. PMID- 15661573 TI - Development of a novel prolonged-release nicotine transdermal patch. AB - A transdermal patch for delivering nicotine for periods of 12-48h was designed. An inclusion complex formed between the nicotine and beta-cyclodextrine (beta-CD) was used in drug depot. The usefulness of a specially cross-linked polyvinyl alcohol (cross-PVA) membrane was investigated as a rate controlling membrane. The influence of carbopol polymers, type C-934P and C-940 and propylene glycol on transdermal permeation of nicotine through the rat skin was investigated. The results indicated a maximum flux of 42 microgcm(-2)h(-1) after 48 h from the patches made from C-934P when the propylene glycol concentration was 15% and the nicotine-beta-CD mole ratio in the inclusion complex was 3:1. These nicotine transdermal patches can be fabricated to obtain a controlled release, zero order systems. PMID- 15661574 TI - St John's wort (Hypericum perforatum) diminishes cognitive impairment caused by the chronic restraint stress in rats. AB - In this study we tested the hypothesis that St John's wort (Hypericum perforatum) may counteract stress-induced memory impairment. Object recognition test and Morris water maze were used to determine whether administration of H. perforatum (350 mg kg(-1) for 21 days), standardized to 0.3% hypericin content, protects against non-spatial and/or spatial memory impairments due to chronic restraint stress (2h daily for 21 days). A group of rats administered the exogenous corticosterone at the dose of 5 mg kg(-1) daily for 21 days, yielding its similar plasma levels as these observed in stress was run in parallel. In the first experiment all rats were tested for recognition memory in the object recognition test. On the following day, the animals were tested in open field and elevated "plus" maze to control for the contribution of respectively, motor and emotional effects of our treatments to the memory tests. In the second experiment, new group of stressed animals was tested for spatial memory in the water maze. We observed that H. perforatum prevented the deleterious effects of both chronic restraint stress and long-term corticosterone on learning and memory as measured in both, the object recognition and the water maze tests. The herb not only prevented stress- and corticosterone-induced memory impairments, but it significantly improved recognition memory (p<0.01) in comparison to control. These results suggest that H. perforatum has a potential to prevent stress memory disorders. PMID- 15661575 TI - Involvement of endothelin and nitric oxide in cyclosporine A-induced contractions in guinea pig isolated gallbladder strips. AB - The involvement of endothelin (ET), ET(A) receptors and nitric oxide (NO) in the contractions induced by cyclosporine A (CyA) were investigated in guinea pig isolated gallbladder strips. Both BQ-123, a selective ET(A) receptor antagonist, and phosphoramidon, an ET converting enzyme inhibitor, inhibited the contractile responses to the parenteral and oral CyA preparations, whereas l-NOARG, a NO synthase inhibitor, potentiated these contractions. Additionally, the pattern of the concentration-dependent contractions in response to ET-1 was similar to that of CyA preparations in gallbladder strips. Both bosentan, a non-selective ET receptor antagonist, and BQ-123 inhibited the ET-1-induced contractions. These findings suggest that an ET-1-mediated mechanism contributes to the contractile response to CyA preparations in guinea pig isolated gallbladder strips. ET(A) receptor activation is likely to be involved in this process. We also speculate that CyA-induced stimulation of NO production might act as a counter-regulatory mechanism in the effect of CyA preparations in this tissue. PMID- 15661576 TI - Ondansetron, a selective 5-HT3 antagonist, antagonizes methamphetamine-induced anorexia in mice. AB - Effects of some selective serotonergic (5-HT) antagonists on methamphetamine induced anorexia were investigated in male mice. The least possible dose of methamphetamine alone that caused significant anorectic activity was 11 micromolkg(-1), i.p. (2 mgkg(-1)). Various doses of some selective serotonergic receptor antagonists were administered half an hour before the above mentioned dose of methamphetamine. Methiothepin potentiated, whereas NAN-190, methysergide, mianserin and ondansetron antagonized methamphetamine-induced anorectic activity. The least possible doses of these antagonists which modified methamphetamine induced anorexia were as follows: methiothepin (1.1 micromolkg(-1), i.p.), NAN 190 (4.2 micromolkg(-1), i.p.), methysergide (2.1 micromolkg(-1), i.p.), mianserin (3.3 micromolkg(-1), i.p.) and ondansetron (0.003 micromolkg(-1), i.p.). The serotonergic antagonists at the above mentioned doses did not modify the food intake of animals not treated with methamphetamine, except for methiothepin, which produced a significant reduction, and mianserin, which produced a significant increase in food intake. The results of the present study indicated that the anorectic activity induced by methamphetamine is related to the interactions of methamphetamine with 5-HT receptor. Since a very small dose (0.003 micromolkg(-1)) of ondansetron (the 5-HT(3) antagonist), as compared with the other antagonists used in this study, antagonized the anorexia induced by methamphetamine, the 5-HT(3) receptor is likely to be the site for this interaction. PMID- 15661577 TI - Stimulation of 5-HT1A receptor with 8-OH-DPAT inhibits hydrogen peroxide-induced neurotoxicity in cultured rat cortical cells. AB - We investigated the effect of 8-hydroxy-2-(N,N-dipropylamino)tetralin (8-OH DPAT), a specific 5-HT(1A) receptor agonist, on H(2)O(2)-induced neuronal cell death in cultured rat cortical cells. H(2)O(2) produced a concentration-dependent reduction of cell viability, which was significantly reduced by (5R,10S)-(+)-5 methyl-10,11-dihydro-5H-dibenzo[a,d] cyclohepten-5,10-imine (MK-801), an N-methyl d-aspartate (NMDA) receptor antagonist. Pretreatment of 8-OH-DPAT over the concentration range of 1-100 microM significantly inhibited the H(2)O(2) (100 microM)-induced neuronal cell death as assessed by a MTT assay and the number of apoptotic nuclei, evidenced by Hoechst 33342 staining. The protective effect of 8 OH-DPAT (100 microM) was completely blocked by the simultaneous treatment of 1-(2 methoxyphenyl)-4-[4-(2-phthalimideo)butyl]piperazine (NAN-190, 10muM), a selective 5-HT(1A) receptor antagonist, but not in the presence of the dopamine receptor blocker spiperone (10 microM), indicating that the protective effect of 8-OH-DPAT was mediated via 5-HT(1A) receptors. In addition, 8-OH-DPAT inhibited the H(2)O(2)-induced elevation of glutamate release into the medium and cytosolic Ca(2+) concentration ([Ca(2+)](c)), generation of reactive oxygen species (ROS), and caspase-3 activity. These results suggest that the activation of 5-HT(1A) receptor with 8-OH-DPAT may ameliorate an oxydative stress-induced apoptosis of neuronal cell by interfering with the increase of [Ca(2+)](c), and then by inhibiting glutamate release, generation of ROS and caspase activity. PMID- 15661578 TI - Hospital-based intensive monitoring of antibiotic-induced adverse events in a university hospital. AB - Hospital-based monitoring is one of the methods used to collect data about drug prescriptions and adverse events. The aim of this 20-day observational prospective study was to evaluate the frequency and type of adverse reaction to antibiotics, and predisposing risk factors in inpatients in six departments of a university hospital (ophthalmology, paediatrics, internal medicine, general surgery, infectious diseases, anaesthesiology and intensive care). The data on all inpatients undergoing antibiotic treatment were collected by physicians trained by our team and validated by an expert panel. Data were recorded on pre formatted confidential cards (MIO-card). In the 171 inpatients evaluated (125 adults: 39.5% male, mean age 61.6 years, range 21-93; and 46 children: 50% male; mean age 4.75 years, range 3 months-12 years), cefazolin (19.9%), chloramphenicol (18.6%), ceftriaxone (15.4%) and netilmicin (12.9%) were the most frequently used antibiotics. Adverse events occurred in four adults and three children: one had leucopenia (trimethoprim/sulfamethoxazole), one nephrotoxicity (netilmicin+teicoplanin) and one nephrotoxicity (cefotaxime), one diarrhoea (ceftriaxone), one neurotoxicity (isoniazid), one angioneurotic oedema (piperacillin) and one skin rashes (ceftriaxone). A number of strategies (educative and persuasive, facilitative and restrictive) have been proposed to improve antibiotic use. Our study suggests that hospital-based monitoring is a good method with which to detect links between drug exposure and adverse drug reactions in children and adults. PMID- 15661580 TI - Increased urinary coproporphyrin excretion observed in patients with differently staged Hodgkin's disease treated with chemotherapy. AB - It has been reported that patients with Hodgkin's disease (HD) show altered porphyrin metabolism, and suggested that the cause is the neoplastic process itself. If this is true, disease progression should be associated with higher levels of porphyrin excretion. The aim of this study was to evaluate urinary coproporphyrin levels in patients with Hodgkin's disease at different stages. As many of the patients received chemotherapy, another aim was to verify experimentally whether chemotherapeutic agents might increase porphyrin levels in rabbits. All of the patients had above-normal urinary coproporphyrin levels. On the other hand, rabbits receiving the porphyrin precursor 5-aminolevulinic acid (5-ALA), and also treated with doxorubicin, showed very high plasma porphyrin levels. The increased levels of urinary coproporphyrins seem to be due to the disease itself, since the patients in stages III and IV had higher excretion values, presumably due to biochemical heme synthesis lesions that lead to the availability of the porphyrin precursor, as well as coproporphyrin accumulation and excretion. The altered porphyrin synthesis may be attributable to the cytotoxic oxygen species generated in the presence of NADH and iron. As the patients also received extensive chemotherapy regimes, the altered porphyrin metabolism may be affected by antineoplastic treatment generating oxygen reactive radicals. The alterations in porphyrin metabolism induced by chemotherapeutic agents appear to be demonstrated in rabbits in which doxorubicin increases porphyrin synthesis after porphyrin precursor treatment. PMID- 15661579 TI - Effects of YM218, a nonpeptide vasopressin V1A receptor-selective antagonist, on human vasopressin and oxytocin receptors. AB - The binding and signal transduction characteristics of YM218 ((Z)-4'-{4,4 difluoro-5-[2-oxo-2-(4-piperidinopiperidino)ethylidene]-2,3,4,5-tetrahydro-1H-1 benzoazepine-1-carbonyl}-2-methyl-3-furanilide hemifumarate), a newly synthesized, potent arginine vasopressin (AVP) V(1A) receptor-selective antagonist, were examined using cloned human AVP receptors (V(1A), V(1B) and V(2)) stably expressed in Chinese hamster ovary (CHO) cells and human uterine smooth muscle cells (USMCs) expressing oxytocin receptors. YM218 potently inhibited specific binding of [(3)H] AVP to V(1A) receptors, exhibiting a K(i) value of 0.30 nM. In contrast, YM218 exhibited much lower affinity for V(1B), V(2) and oxytocin receptors, exhibiting K(i) values of 25,500 nM, 381 nM and 71.0 nM, respectively. In CHO cells expressing V(1A) receptors, YM218 potently inhibited the AVP-induced increase in intracellular Ca(2+) concentration ([Ca(2+)](i)), exhibiting an IC(50) value of 0.25 nM. However, in human USMCs expressing oxytocin receptors, YM218 exhibited a much lower potency in inhibiting the oxytocin-induced [Ca(2+)](i) increase, showing an IC(50) value of 607 nM, and had no effect on the AVP-induced [Ca(2+)](i) increase in CHO cells expressing V(1B) receptors. Furthermore, in CHO cells expressing V(2) receptors, YM218 did not potently inhibit the production of cAMP stimulated by AVP, showing an IC(50) value of 62.2 nM. In all assays used, YM218 did not exhibit any agonistic activity. These results demonstrate that YM218 is a potent, nonpeptide human V(1A) receptor-selective antagonist, and that YM218 will be a valuable new tool to gain further insight into the physiologic and pharmacologic actions of AVP. PMID- 15661581 TI - Are women better off because of the new Medicare drug legislation? AB - The passage of the Medicare Prescription Drug, Improvement and Modernization Act of 2003 will help to reduce the out-of-pocket burdens women will face in 2006 once the full drug benefit is introduced. Nonetheless, the legislation is less than ideal and creates a number of issues that should be improved to meet women's needs. Three key elements of the legislation that were essential in gaining its passage stand in the way of such improvements: limits on the amount spent on the benefit, requirements to rely on the private sector, and a failure to adequately arrange for future financing. A major overhaul is unlikely, but it is possible that modest improvements to aid Medicare beneficiaries will be considered in the future. Several of those improvements are described here. PMID- 15661582 TI - Use of complementary and alternative medicine among American women. AB - INTRODUCTION AND BACKGROUND: Although women are increasingly using complementary and alternative (CAM) therapies, a national profile characterizing women who use CAM has yet to be described. The purpose of this study is to provide prevalence estimates of recent CAM use among American women and to examine sociodemographic and other characteristics associated with use. METHODS: Data from the 1999 National Health Interview Survey are used for the analysis. Three operational definitions of recent CAM use are employed (any CAM, type-specific, and domain specific use). All proportions and prevalence estimates are weighted and standard errors are adjusted to account for complex sample design; weighted logistic regression (with coefficient variance adjustment) is also used. RESULTS: Overall, 33.5% of American women used CAM in the past 12 months. Spiritual healing/prayer and herbal medicine are the most commonly used, and hypnosis, biofeedback, and energy healing are the least common. Multivariate results show that women who are older, have more education, poorer health, or live in the west or midwest (versus south) are more likely to use CAM. Compared to whites, blacks, Hispanics, and Asians are less likely to use CAM. Foreign-born women, those with lower income, or who live in the Northeast are also less likely to use CAM. Insurance status is not independently associated with CAM use. CONCLUSIONS AND DISCUSSION: This study provides one of the first comprehensive investigations of CAM use among American women. Future research examining the determinants of CAM use, incorporating attitudinal and health conditions, as well as clinical efficacy, effectiveness, and health outcome studies of specific CAM therapies are warranted. PMID- 15661583 TI - Stress and women with physical disabilities: identifying correlates. AB - INTRODUCTION: We examined correlates of perceived stress among women with physical disabilities to identify variables that may be amenable to change through psychosocial interventions. METHOD: The method for this investigation involved a correlational analysis of data gathered from 415 women living with physical disabilities on abuse and other health concerns. The women were recruited in outpatient clinics where they each participated in a face-to-face, semistructured interview. RESULTS: Based on multiple regression analyses, the findings indicate that demographic (age, income) and disability (mobility, level of assistance needed) variables explained a small but significant proportion of the variance in perceived stress. Variables judged to be potentially amenable to change through psychosocial interventions (i.e., social support, pain interference, and abuse) contributed significantly to stress over and above the demographic and disability variables. CONCLUSION: Women with physical disabilities reported high levels of perceived stress. Particularly at high risk are women who are limited by pain, lack social support, and/or have experience with recent abuse. Stress management interventions for this population of women should consider incorporating components addressing pain, social support, and abuse. PMID- 15661584 TI - Health care interventions for intimate partner violence: what women want. AB - OBJECTIVE: We sought to determine what women want from health care interventions for intimate partner violence (IPV) and understand why they found certain interventions useful or not useful. METHODS: We conducted interviews with 21 women who have a past or current history of intimate partner violence. Participants were given cards describing various IPV interventions and asked to perform a pile sort by placing cards into three categories ("definitely yes," "maybe," and "definitely no") indicating whether they would want that resource available. They were then asked to explain their categorizations. RESULTS: The pile sort identified that the majority of participants supported informational interventions and individual counseling. Only 9 of 17, however, felt couple's counseling was a good idea with seven reporting it was definitely not useful. Half wanted help with substance use and treatment for depression. Interventions not well regarded included "Receiving a follow-up telephone call from the doctor's office/clinic" and "Go stay at shelter" with only 7 and 5 of the 21 women placing these cards in the "definitely yes" pile. "Health provider reporting to police" was the intervention most often placed in the "definitely no" pile, with 9 of 19 women doing so. The women described several elements that affected their likelihood of using particular IPV interventions. One theme related stages of "readiness" for change. Another theme dealt with the complexity of many women's lives. Interventions that could accommodate various stages of "readiness" and helped address concomitant issues were deemed more useful. Characteristics of such interventions included: 1) not requiring disclosure or identification as IPV victims, 2) presenting multiple options, and 3) preserving respect for autonomy. CONCLUSIONS: Women who had experienced IPV described not only what they wanted from IPV interventions but how they wished to receive these services and why they would chose to use certain resources. They advised providing a variety of options to allow individualizing according to different needs and readiness to seek help. They emphasized interventions that protected safety, privacy, and autonomy. PMID- 15661585 TI - Understanding physicians' attitudes towards hormone therapy. AB - OBJECTIVE: We sought to understand the relationship among components of residency education about hormone therapy (HT), knowledge about HT, and provider attitudes toward HT during a time of rapidly changing practice guidelines. METHODS: We surveyed residents in the University of Pittsburgh Internal Medicine residency programs between February to April 2002 (after the release of the Heart Estrogen/Progestin Replacement Study and prior to the release of preliminary Women's Health Initiative data) regarding demographics, educational (didactic and experiential) exposures to HT and menopause management, knowledge about HT, and attitudes toward HT. RESULTS: Sixty-nine of 92 (75%) eligible residents completed the survey; 38% were women. The race and gender of responders did not differ from nonresponders. Residents had significant didactic exposure to HT and menopause management with 80% reporting more than one didactic exposure. Despite this, HT knowledge was low (mean knowledge score 47 +/- 16%) and only 26% of residents felt prepared to counsel patients about HT. We identified four factors related to provider attitudes toward HT: "persistence" in universally recommending HT, confidence in "HT benefits," concern about "HT cardiac risks," and concern about "HT noncardiac risks." More appropriate attitudes were associated with attending a lecture, having a rotation with a discussion of menopause management (i.e., Women's Health), and a continuity practice including more than 30% women. Pharmaceutical detailing and self-directed study were associated with less appropriate attitudes. Knowledge did not influence attitudes. Strongly held beliefs about the benefits of HT, appropriate or inappropriate, were associated with increasing "persistence." CONCLUSIONS: In an area of rapidly changing information, such as the risks and benefits of HT, knowledge is low. Experiential learning appropriately influences attitudes, while pharmaceutical detailing was associated with inappropriate attitudes toward HT risks. PMID- 15661589 TI - Adolescent's concept of drug experimentation. PMID- 15661586 TI - Willing and able? Provision of medication for abortion by future internists. AB - INTRODUCTION AND BACKGROUND: The development of medications such as mifepristone (RU486) has created the opportunity to introduce medication abortion as a component of office practice. METHODS: Two hundred twelve residents training in internal medicine, family practice, and gynecology at 11 residency programs completed anonymous surveys assessing willingness to provide medication for abortion and perceived barriers to future provision of mifepristone. RESULTS: Residents training in internal medicine knew less about mifepristone and preabortion screening than other primary care trainees. Forty-two percent of internists, 84% of family practitioners, and 83% of gynecologists were willing to prescribe mifepristone (p < .001). Many internists were concerned about lacking adequate "backup" access to vacuum aspiration services (84% of internists, 74% of family practitioners, 35% of gynecologists; p < .001). In multivariable analysis, the training-related factors most predictive of whether an internist was willing to provide medication for abortion were feeling that mifepristone is very safe, abortion services are needed by the patients served, knowing to check an ultrasound before inducing abortion, and having no concern of how to manage bleeding or of lacking adequate backup should vacuum aspiration be needed. CONCLUSIONS AND DISCUSSION: Many (42%) future internists are willing to provide mifepristone, but most lack adequate knowledge of mifepristone and preabortion screening. As access to abortion services is limited in many U.S. counties, internists who are willing to provide mifepristone should be offered the necessary training to do so safely. PMID- 15661590 TI - High school off-campus lunch policies and adolescent motor vehicle crash risks. AB - PURPOSE: To examine differences in motor vehicle crash involvement for teenagers in communities with and without school policies enabling teens to drive off campus during lunchtime. METHODS: Comparison of lunchtime motor vehicle crashes involving teen drivers in two North Carolina counties having open-lunch policies with a third county without an open-lunch policy. We also compare crash rates during the before-school period and at all times of the day in the three counties. Data were analyzed by computing rate ratios of teens' involvement in a crash during the three time periods and comparing them among the three counties. RESULTS: Crash rates over the lunch hours were significantly higher for teenagers in the counties with open-lunch policies, despite these counties having no elevated crash risk during other time periods. This resulted in a relative risk of lunchtime crash involvement of 3.10 and 2.98 (95% CI 1.97-4.89 and 1.87-4.74, respectively) compared with the county without an open-lunch policy. Number of vehicle occupants also increased during the lunch hours in the counties with open lunch policies. CONCLUSIONS: Open-lunch policies contribute to motor vehicle crashes in teenagers and encourage a situation where there are multiple occupants per vehicle, a known risk factor for teenage motor vehicle crashes. PMID- 15661591 TI - Depressive symptomatology, youth Internet use, and online interactions: A national survey. AB - PURPOSE: To investigate the online communications and self-disclosure practices of youth reporting depressive symptomatology. METHOD: The Youth Internet Safety Survey was a nationally representative telephone survey of 1501 Internet-using youth between the ages of 10 and 17 years, and one caregiver in their household. Fifty-three percent of youth participants were male and 73% were white race. The purpose of the survey was to obtain prevalence rates for unwanted sexual solicitation, harassment, and unwanted exposure to sexual material among young people online. Questions about current depressive symptomatology were also queried; this variable was defined based upon the DSM-IV definition of a major depressive episode: major depressive-like symptomatology (5+ symptoms of depression and functional impairment in at least one area); minor depressive-like symptomatology (3+ symptoms of depression); mild or no depressive symptomatology (<3 symptoms of depression). Data were cross-sectional and collected between the fall of 1999 and spring 2000. Multinomial logistic regression was used to estimate the conditional odds of reporting DSM-IV-like major or minor depressive symptomatology vs. mild/no symptomatology given the indication of self-disclosure practices and interactions with others online. Males and females were assessed separately. RESULTS: Talking with strangers online, using the Internet most frequently for e-mailing others, and intensity of Internet use differentiated youth reporting depressive symptoms from asymptomatic peers. Report of depressive symptomatology was not related to most measures of general Internet use nor gender differences. Personal disclosure was significantly more likely to be reported by both young men and young women who reported major depressive symptomatology vs. mild or no symptomatology. Differences were observed for how adolescents choose to self-disclose; females posted pictures of themselves, whereas males were more likely to provide personally identifiable information. Finally, most gender-related variation reflected differences in the magnitude of Internet associations with depressive symptoms rather than the types of Internet use, access, or online communications. CONCLUSIONS: Youth-reported depressive symptomatology is associated with differences in online interactions and self disclosure practices. PMID- 15661592 TI - Attention deficit disorder and hospitalization owing to intra- and interpersonal violence among children and young adolescents. AB - PURPOSE: To investigate the associations between intra-and interpersonal violence and related injuries and the diagnosis of attention deficit disorder (ADD) among children and young adolescents. METHODS: This was a population-based epidemiological study that analyzed data routinely collected on hospitalized patients owing to injuries. Data were obtained from the routinely collected inpatient statistics. Information included patients' demographics, diagnostic classifications of admitting problem, classification of external causes of injury, length of stay, and outcome of hospitalization. Patients with comorbidity of ADD were identified by the ICD-9CM diagnosis code. Data were analyzed univirately using Pearson Chi-square tests. Logistic regression analyses were also applied to calculate the adjusted odds ratio and their corresponding 95% confidence intervals. RESULTS: Significant associations between suicide and self harm, injuries owing to assault, and diagnosis of ADD were found. Patients diagnosed with ADD stayed in the hospital longer than others, disregarding the cause of their injury. CONCLUSION: Children and adolescents with ADD are at risk of being victims of assaults, as well as suicide and self harm. Assessment for ADD can be considered as part of school-age childhood screening programs. PMID- 15661593 TI - Adolescent help-seeking for dating violence: prevalence, sociodemographic correlates, and sources of help. AB - PURPOSE: To examine the prevalence and sociodemographic correlates of help seeking and helping sources used by adolescent victims and perpetrators of dating violence. METHODS: Data are from the victims (n = 225) and perpetrators (n = 140) of dating violence identified from a longitudinal study of adolescent dating violence conducted in the public school system of a primarily rural North Carolina county. Logistic regression was used to examine bivariate and multivariate relationships between study variables and help-seeking or sources of help. RESULTS: Sixty percent of victims and 79% of perpetrators did not seek help for dating violence. Male perpetrators were more likely to seek help than female perpetrators. The odds of seeking help increased with perpetrators' age (OR = 1.79 per year; 95% CI = 1.05, 5.76). Most victims and perpetrators who sought help chose friends and family members rather than professionals. Male victims and perpetrators who sought help were more likely than female victims and perpetrators to choose professional sources of help. CONCLUSIONS: Most adolescent dating violence victims and perpetrators do not seek help. Male perpetrators and older perpetrators were more likely to seek help than female perpetrators or younger perpetrators. Friends and family members are more common sources for help seeking than professionals, but males were more likely to seek help from professionals than females. PMID- 15661594 TI - Nonspecific low-back pain in Kuwaiti children and adolescents: associated factors. AB - PURPOSE: To investigate factors associated with low-back pain among Kuwaiti children and adolescents; and study their relation with social activities and school performance. METHOD: A cross-sectional population of 400 schoolchildren aged 10 to 18 years were selected from junior and high schools in one of the governorates in Kuwait. Data were collected through personal interviews using a questionnaire and analyzed using SPSS software. RESULTS: Older adolescent females who practice strenuous physical activities and spend a significant amount of time watching TV, and smokers were found to be significantly more likely to have low back pain. Also, high grade-point average students were found to experience more low-back pain. Methods of carrying school materials were not significantly associated with low-back pain. Low-back pain did not significantly affect the number of absent days from school. CONCLUSIONS: The study suggests that back pain in Kuwaiti schoolchildren and adolescents is associated with older age, female gender, increase in physical activity, and time spent watching television. PMID- 15661595 TI - Social environmental factors and preteen health-related behaviors. AB - PURPOSE: To examine associations among risk and protective factors with negative (substance use, delinquent behavior, sedentary recreation, empty calorie consumption, suicidal behavior) and positive behaviors (prosocial recreation, productive behavior, exercise, nutrition behavior, academic achievement, seatbelt use). METHODS: Data were from sixth-grade public school students (n = 133,629) in 2001. Factor analysis produced five risks, five protectors, seven negative and six positive behaviors. Associations were tested among single and cumulative risks and protectors with behaviors (linear, logit regression) and combinations of high and low risks and protectors with behaviors (analysis of variance, Chi square). RESULTS: Individual and cumulative risks were related to higher and protectors were related to lower negative behaviors. Protectors were associated with higher positive behaviors, with some exceptions. Risks and their sum were associated with lower academic achievement and seatbelt use, but were linked to higher, rather than lower productive behavior. Being bullied or victimized was correlated with higher levels of exercise, good nutrition, and prosocial recreation. The high risk/low protection combination had the highest negative behaviors and low risk/high protection the lowest, but for positive behaviors, high protectors with either high or low risks showed higher positive behaviors. CONCLUSIONS: These findings provide new information about how (a) risks and protectors are associated with negative behaviors besides substance use and delinquency, (b) cumulative protectors, as well as risks, are related to negative and positive behaviors, and (c) combinations of high and low risks and protectors are related to behaviors. The unusual findings for positive behaviors merit further exploration. PMID- 15661596 TI - Overweight status and depressive symptoms during adolescence. AB - PURPOSE: To: (a) extend previous research on the association between overweight status and depressive symptoms among adults to adolescents, (b) consider whether this association varies across social structural contexts and school context, and (c) explore additional mechanisms linking overweight status to depressive symptoms. METHODS: We used survey regression procedures to analyze data from the first wave of the National Longitudinal Study of Adolescent Health. Degree of overweight was indicated by body mass index (BMI), which we calculated using self reported height and weight information, whereas depressive symptoms were assessed with the Center for Epidemiological Studies Depression Scale (CES-D). Data were analyzed to determine (a) the social groups in which being overweight was least common, (b) the association between overweight status and depressive symptoms, and (c) potential mediators of the association between relative weight and symptoms of depression, including dieting and self-rated health. The analytic sample contained 18,924 adolescents aged 11 to 21 years (mean age was 15.68). Approximately half the sample consisted of females (n = 9634). RESULTS: Adjusting for exercise and sociodemographic characteristics, we found that relative weight was associated with depressive symptoms for girls but not boys. For both, the association between overweight status and symptoms of depression was stronger among adolescents in lower grades. Dieting explained the positive association between relative weight and depressive symptoms for girls, whereas self-rated health mediated the association between relative weight and symptoms of depression for adolescents in lower grades. CONCLUSION: To fully understand both the physical and mental health consequences of adolescent obesity, the social dimension of weight must be examined. PMID- 15661598 TI - Adolescent pregnancy prevention: An abstinence-centered randomized controlled intervention in a Chilean public high school. AB - PURPOSE: To evaluate the efficacy of an abstinence-centered sex education program in adolescent pregnancy prevention, the TeenSTAR Program was applied in a high school in Santiago, Chile. METHODS: A total of 1259 girls from a Santiago high school were divided into three cohorts depending on the year they started high school: the 1996 cohort of 425 students, which received no intervention; the 1997 cohort, in which 210 students received an intervention and 213 (control group) did not; and the 1998 cohort, in which 328 students received an intervention and 83 (control group) did not. Students were randomly assigned to control and intervention groups in these cohorts, before starting with the program. We conducted a prospective, randomized study using the application of the TeenSTAR sex education program during the first year of high school to the intervention groups in the 1997 and 1998 cohorts. All cohorts were followed up for 4 years; pregnancy rates were recorded and subsequently contrasted in the intervention and control groups. Pregnancy rates were measured and Risk Ratio with 95% confidence interval were calculated for intervention and control groups in each cohort. RESULTS: Pregnancy rates for the intervention and control groups in the 1997 cohort were 3.3% and 18.9%, respectively (RR: 0.176, CI: 0.076-0.408). Pregnancy rates for the intervention and control groups in the 1998 cohort were 4.4% and 22.6%, respectively (RR 0.195, CI: 0.099-0.384). CONCLUSIONS: The abstinence centered TeenSTAR sex education intervention was effective in the prevention of unintended adolescent pregnancy. PMID- 15661597 TI - Influence of prior sexual risk experience on response to intervention targeting multiple risk behaviors among adolescents. AB - PURPOSE: To identify correlates of sexual risk variations among African-American adolescents, and to examine the influence of prior sexual experience on response to a HIV risk-reduction intervention. METHODS: Eight hundred seventeen African American youth aged 13 to 16 years living in and around urban public housing in Baltimore were recruited to participate in a HIV risk-reduction intervention targeting multiple risk behaviors. An instrument designed to measure three levels of sexual risk ("abstinent," "protected sex" [having sex with a condom], and "unprotected sex" [having sex without a condom]) was administered at baseline, 6 months and 12 months postintervention. Multiple regression analyses were conducted to identify predictors of the degree of sexual risk using longitudinal data. Repeated measure analyses were conducted to assess behavioral changes over time among the three groups. RESULTS: Data confirmed the co-variation of sexual risk behavior and other problem behaviors among adolescents, cross-sectionally and longitudinally. After exposure to an 8-session risk-reduction intervention, youth engaging in the highest degree of sexual risk demonstrated the greatest reduction in both sexual risk and other risks. These improvements were seen at both 6 months and 12 months postintervention. Youth who were abstinent at baseline maintained the lowest levels in risk involvement throughout the study period when compared with sexually active youth. However, abstinent youth risk involvement significantly increased at 6 months and 12 months after baseline. Youth engaging in protected sex at baseline demonstrated a significant increase in non-condom use and a significant decrease in multiple risk involvement over time. CONCLUSIONS: Results support HIV risk-reduction intervention efforts that target multiple risk behaviors. Response of adolescents to the intervention is directly related to the sexual risk behavior at baseline. These data may suggest that the response to risk behavior intervention depends in part on the risk behavior profile of the population to which it is being applied. PMID- 15661599 TI - Emotional and behavioral outcomes among adolescents with mild developmental deficits in early childhood. AB - PURPOSES: To evaluate emotional and behavioral outcomes in adolescents who in early childhood were treated for mild developmental deficits; and to identify predictive factors in early childhood, for future emotional and social competence abnormalities. METHODS: The records of children referred to Hanna Khoushi Child Development Center in Haifa for mild developmental delay were reviewed. Parents and adolescents were requested to complete the Child Behavior Checklist (CBCL) and the Youth Self-Report (YSR) 12 to 16 years after discharge from the Child Development Center. Analysis of the two questionnaires and identification of predictive variables in early childhood for emotional problems and social competence difficulties during adolescence were performed, using ANOVA Student's T-test, Chi-square, and multiple regression. RESULTS: The most frequent developmental diagnoses on admission to the Child Development Center were mild motor (27.3%) or language (23.2%) deficits. Of the treated children, 53.4% were discharged without any developmental deficit. All CBCL and YSR T-scores were within the nonpsychopathology range. No notable differences were found between the study scores and the scores regarding typical Israeli and American youths. Significant differences were, however, observed in self-perception of internalizing emotional problems between male and female adolescents: T-scores of 51.9 +/- 8.0 vs. 47.4 +/- 10.8, respectively (p < .05). Motor and language deficits were associated with lower general competence than general developmental delay and emotional developmental disturbances (T-scores: 47.9 vs. 49.1 and 50.9, p< .05). Admission to the Center after the age of 18 months was associated with higher T-scores for general as well as externalizing problems when compared with scores associated with admission before the age of 18 months (by 5.75 points for both parameters, p < .001). CONCLUSION: The perception of parents and children with minor developmental deficits observed in early childhood regarding their emotional and social competence during adolescence is similar to typical youths. PMID- 15661600 TI - Assessment of trauma symptoms among adolescent assault victims. AB - PURPOSE: To identify and evaluate the effectiveness of an assessment tool that could be used to assess the psychological needs of youth injured by community violence. METHODS: The Trauma Symptom Checklist for Children (TSCC) was administered to 120 adolescents participating in Project Ujima, a hospital-based program providing Emergency Department support and home-based psychosocial follow up to victims of violent crime (mean age = 14.8 years; 72% male; 70% African American, 19% white, 8% Latino). Participants' TSCC scores were compared with normative data using one-sample, two-tailed, Student's t-tests. Comparisons of TSCC scale scores were also made based on participant age, gender, ethnicity, and injury type using one-way multivariate analysis of variance. RESULTS: Seventeen percent of the participants scored in the clinical range on the Underreporting Scale of the TSCC, reflecting a tendency to deny common thoughts, emotions, and behaviors. Elevations on all clinical scales were modest. Males showed elevations on Underreporting and decreased trauma symptoms, in comparison to normative data and to females in the sample. CONCLUSIONS: The reported low levels of symptomatology among this sample of youth may be due, in part, to a defensive response style. PMID- 15661601 TI - Internet-administered adolescent health questionnaires compared with a paper version in a randomized study. AB - PURPOSE: To assess whether the scores of an Internet-administered adolescent health questionnaire (using two different interfaces) are equivalent to those obtained via paper and pencil (P&P). Furthermore, it compares adolescents' evaluations of modes of administration. METHODS: We randomly assigned 591 adolescents (aged 13-17 years) from five secondary schools within their classes to one of the two Internet interfaces (multiple items vs. one item per screen) or P&P. Adolescents completed questionnaires on psychosocial well-being (KIVPA), self-reported problems, health care utilization, and health-related behavior and supplementary evaluation surveys (on the given health questionnaire mode) in the computer classrooms. Differences in questionnaire scores among administration modes were analyzed by the Student's t-test and Wald test. RESULTS: Response rate was 96% (n = 565). Adolescents in the Internet one-item mode more frequently reported satisfaction with appearance compared with the Internet multiple-items mode (p 0.05) in DNA damage in control (9.0 +/- 5.7 AU) versus exposed (8.5 +/- 4.8 AU) groups were found. These results suggest that low doses of 137Cs internal contamination are not able to produce detectable DNA damage under the conditions used for the comet assay in this study. Further studies considering effects of high exposure should be performed on chronically exposed people using this assay. PMID- 15661619 TI - Genetic disposition to addictive disorders--current knowledge and future perspectives. AB - It is well established that addictive disorders have a strong genetic background. Multiple, and in part interacting, genes are likely to be responsible for the disease phenotype, making the search for underlying alleles a challenging and complicated task. Linkage analyses and association studies have failed to unequivocally identify underlying genes. Conversely, genome sequencing and the systematic search for polymorphic marker loci have yielded dense chromosome maps so that, along with automated genotyping, the identification of individual genes will soon become possible. Initial results provide hints that regulators of gene expression might play an important role in addiction. PMID- 15661620 TI - Neurobiology of craving, conditioned reward and relapse. AB - Chronic vulnerability to relapse is a formidable challenge for the treatment of drug addiction. The neurobiological basis of relapse and its prevention has, therefore, attracted major attention in addiction research. Current conceptualizations of addiction recognize craving as a central driving force for ongoing drug use, as well as for relapse following abstinence. Progress has been made in understanding experiential factors, neurocircuitry components and signaling mechanisms that mediate conditioned drug-seeking behaviour, craving and long-lasting susceptibility to relapse. Importantly, stress contributes to drug craving, and there is evidence for overlap between the neural and neuroendocrine mechanisms implicated in drug desire evoked by drug cues and stress. Recent research has substantially advanced our understanding of the neurobiological factors responsible for drug craving and relapse, with promising therapeutic implications. PMID- 15661621 TI - Synaptic plasticity and drug addiction. AB - Recent studies have suggested that the development of addictive behaviours shares common features with traditional learning models. Synaptic plasticity, a possible substrate for learning, has been demonstrated in neural reward circuits and might contribute to the learning of addictive behaviours. Changes in the strength of synaptic connections have been investigated in dopaminergic cells of the ventral tegmental area in response to several addictive drugs. Rapid and persistent forms of synaptic plasticity (specifically, long-lasting synaptic potentiation) have been demonstrated to accompany some of the behavioural effects of addictive drugs. We hypothesize that drug-induced synaptic plasticity might play a role in reward-related learning and addiction by modifying the fine tuning of dopaminergic cell firing. PMID- 15661622 TI - Gene expression induced by drugs of abuse. AB - The transition from infrequent drug use to addiction (i.e. the loss of control over consumption of a drug) probably involves changes in gene expression that restructure neural circuits in the brain. The number of genes that have been demonstrated to change expression in response to drugs has increased rapidly in recent years owing to microarray technology, which allows measurement of thousands of genes at one time. It is now important to identify which of these changes are causally related to the compulsive behavior associated with drug addiction, and which are non-specific changes related to general features of arousal or other physiological responses (e.g. stress, altered body temperature or energy metabolism). PMID- 15661623 TI - Beyond the reward hypothesis: alternative functions of nucleus accumbens dopamine. AB - According to the dopamine (DA) hypothesis of reward, DA systems in the brain, particularly in the nucleus accumbens, are thought to directly mediate the rewarding or primary motivational characteristics of natural stimuli such as food, water and sex, as well as various drugs of abuse. However, there are numerous problems associated with this hypothesis. Interference with accumbens DA transmission does not substantially blunt primary motivation for natural rewards such as food, but it does disrupt the propensity of animals to engage in effortful responding to obtain food. Electrophysiological and voltammetric studies indicate that novel stimuli, conditioned stimuli that predict reward, and instrumental behaviors that deliver natural rewards all act to stimulate DA activity. Accumbens DA acts as a modulator of several functions related to motivated behavior, and can influence normal and pathological cognitive function, activational aspects of motivation, anergia or psychomotor slowing in depression, the impact of conditioned stimuli, plasticity and a variety of sensorimotor functions. PMID- 15661624 TI - Human brain imaging and substance abuse. AB - Over the past decade, functional neuroimaging has contributed greatly to our knowledge about the neuropharmacology of substance misuse in man. Techniques such as functional magnetic resonance imaging, positron emission tomography (PET) or single photon emission tomography (SPET) can measure changes in regional cerebral activity, whereas changes in neuropharmacological parameters (e.g. receptor number and neurotransmitter levels) can be directly measured only with PET and SPET. Recently, a series of studies have shown common effects of substances of misuse on the brain, such as an acute increase in dopamine release (followed by hypofunction after chronic use) and cue exposure-induced activation of the frontal cortex, as well as specific effects, such as ecstasy-induced decreases in serotonergic transporters and occupancy of opiate receptors by substitute drugs (e.g. methadone and buprenorphine) in opiate addicts. PMID- 15661625 TI - GABAA receptor subtypes: any clues to the mechanism of benzodiazepine dependence? AB - Chronic use of benzodiazepines for the treatment of anxiety has revealed that these drugs can lead to dependence as indicated by withdrawal symptoms following cessation and tolerance to the drugs effects. Together with their reinforcing properties, this has led to them being labelled as scheduled drugs. Our new knowledge regarding the molecular structure of the benzodiazepine binding site and the growing ability to differentiate GABA(A) receptor subtypes, either by genetic manipulation or subtype selective compounds, have begun to facilitate our understanding of what underlies the mechanism of benzodiazepine dependence. In addition, the involvement of GABA(A) receptors in this phenomenon is leading to a greater understanding of other drugs such as alcohol and opiates. PMID- 15661626 TI - Nicotine: from molecular mechanisms to behaviour. AB - The addictive potential of nicotine is clearly recognized by the tenacity of tobacco smoking for most users, and has prompted extensive psychopharmacological studies in animals. In parallel, the interaction of nicotine with the many subtypes of its eponymous receptor has been the focus of molecular and cellular investigations. More recently, a convergence of these approaches has been stimulated by the generation of transgenic animals, which facilitates analysis of the impact of molecular changes on behaviour. Nicotine, like other addictive drugs including psychomotor stimulants, promotes dopamine release in the nucleus accumbens. This transmitter system has been a major focus of both neurochemical and behavioural investigations, although recently the pre-eminence of this system in nicotine dependence has been challenged. Complexities in the brain circuitry (including the subdivisions of the nucleus accumbens) and differences between behavioural models help to rationalise the current controversy. PMID- 15661627 TI - Opioids: cellular mechanisms of tolerance and physical dependence. AB - Morphine and other opioids are used and abused for their analgesic and rewarding properties. Tolerance to these effects develops over hours/days to weeks, as can physical and psychological dependence. Despite much investigation, the precise cellular mechanisms underlying opioid tolerance and dependence remain elusive. Recent studies examining mu-opioid receptor desensitization and trafficking have revealed several potential mechanisms for acute receptor regulation. Other studies have reported changes in many other proteins that develop during chronic opioid treatment or withdrawal and such changes may be partly responsible for the cellular and synaptic adaptations to prolonged opioid exposure. While these studies have added to our knowledge of the cellular processes participating in opioid tolerance and dependence, the challenge remains to integrate these observations into a coherent explanation of the complex changes observed in whole animals chronically exposed to opioids. PMID- 15661628 TI - Long-term effects of exposure to cannabis. AB - The long-term use of cannabis, particularly at high intake levels, is associated with several adverse psychosocial features, including lower educational achievement and, in some instances, psychiatric illness. There is little evidence, however, that long-term cannabis use causes permanent cognitive impairment, nor is there is any clear cause and effect relationship to explain the psychosocial associations. There are some physical health risks, particularly the possibility of damage to the airways in cannabis smokers. Overall, by comparison with other drugs used mainly for 'recreational' purposes, cannabis could be rated to be a relatively safe drug. PMID- 15661629 TI - Ethanol and brain damage. AB - It is now well established that even uncomplicated alcoholics who have no specific neurological or hepatic problems show signs of regional brain damage and cognitive dysfunction. Improvements in neuroimaging technology, magnetic resonance imaging, magnetic resonance spectroscopy and positron emission tomography have contributed significantly, revealing alcoholic-specific changes in the CNS associated with neuropsychological abnormalities. Although greater efforts are needed, a human brain bank specifically targeting alcohol cases is now able to provide fresh and frozen tissue for alcohol researchers. These tissues can be used to test hypotheses developed using animal models and/or in vitro studies. The aim is to delineate mechanisms underlying alcohol-related brain damage in humans. The development of high-throughput, non-hypothesis-driven approaches using DNA microarrays and proteomics might also provide clues to this important problem. PMID- 15661630 TI - Ecstasy: pharmacology and neurotoxicity. AB - In part because it is amphetamine derived, ecstasy has inherited some of its parent compound's reputation for being neurotoxic. However, whereas amphetamine and methamphetamine undoubtedly cause irreversible brain damage with long-term use, the jury is still out on the party drug ecstasy. The deadly reputation of ecstasy has been fuelled by the tragic fates of healthy young clubbers who have died after taking the drug. However, compared with other recreational drugs, there have been very few ecstasy-related deaths. Further, there is little evidence for short-term neurotoxicity of ecstasy at recreational doses. That is not to say that ecstasy leaves the user neutral. Chronic ecstasy use causes depletion of serotonin, which has subtle but important long-term effects on cognition and mood. Although it seems unlikely that we will be faced with a generation of party goers who suffer from premature Parkinson's disease, so little is known about the long-term effects of ecstasy on mood, emotional states and cognitive function that at present we cannot predict what impact their use of ecstasy will have on the middle-age of the average ecstasy user. PMID- 15661631 TI - Stimulants: use and abuse in the treatment of attention deficit hyperactivity disorder. AB - Attention deficit hyperactivity disorder (ADHD) is the most prevalent childhood developmental disorder and is also of unclear neurobiological aetiology. However, recent advances in molecular genetics and brain imaging implicate dopaminergic hypofunction in the frontal lobes and basal ganglia in ADHD. Psychostimulants (e.g. methylphenidate and amphetamine, which are potent inhibitors of the dopamine transporter) are the first choice medication for ADHD and have a good acute efficacy and safety profile when used for this disorder. Whether long-term psychostimulant administration to adolescents alters neural development and behaviour or increases the risk of substance abuse is less certain. The precise molecular mechanism of action of psychostimulants is beginning to be established. Furthermore, preclinical studies have begun to use lower clinically relevant doses and oral administration of psychostimulants to determine their long-term effect on development, behaviour and neurochemistry, which is an important public health issue associated with chronic medication of adolescents with ADHD. PMID- 15661632 TI - Psychostimulants and monoamine transporters: upsetting the balance. AB - Monoamine transporters were originally associated simply with the termination of synaptic monoamine function. In addition to amine reuptake, however, the transporters can act as ion channels that affect exocytotic neurotransmitter release and can operate in reverse mode, mediating non-exocytotic amine release. Activity at the plasma membrane is controlled by trafficking, which is modulated by interaction with both substrates and inhibitors and by cytosolic kinases and phosphatases. Monoamine transporters also constitute the principal sites of action of many psychoactive drugs, including amphetamines and cocaine, as well as therapeutic drugs for the treatment of depression, addiction and attention deficit hyperactivity disorder, each modifying the balance of presynaptic neurotransmitter function. PMID- 15661633 TI - PCP: from pharmacology to modelling schizophrenia. AB - Phencyclidine has attracted the attention of neuroscientists for many years because of its ability to produce, in humans, a range of symptoms remarkably similar to those of patients suffering from schizophrenia. The main action of phencyclidine is as a non-competitive antagonist of the NMDA class of glutamate receptor. In the past few years, dramatic advances have been made in our understanding of the neuroanatomical and pathological basis of schizophrenia. In turn, these have allowed assessment of the ability of phencyclidine to produce equivalent changes in the rodent CNS. It has now become clear that chronic intermittent low doses of phencyclidine produce a pattern of metabolic and neurochemical changes in the rodent brain that mirror those observed in the brains of schizophrenic patients with impressive precision. This should be of enormous benefit in the search for new anti-psychotic drugs with improved efficacy against the full range of schizophrenic symptoms. PMID- 15661634 TI - Novel pharmacotherapeutic approaches for the treatment of drug addiction and craving. AB - Pharmacological agents have shown limited efficacy and consistency in the treatment of drug addiction. Hence, the development of new medications with improved long-term efficacy and reduced side effects should be given a high priority given the costs to society associated with drug abuse and drug-related pathologies. Neurochemical systems can be significantly altered by repeated exposure to drugs of abuse. These long-term molecular and neurochemical changes might, in turn, explain the core features of addiction--the compulsive seeking and taking of the drug--as well as the risk of relapse. PMID- 15661635 TI - Developmentally regulated expression of the regulator of G-protein signaling gene 2 (Rgs2) in the embryonic mouse pituitary. AB - During the development of the anterior pituitary gland, five distinct hormone producing cell types emerge in a spatially and temporally regulated pattern from an invagination of oral ectoderm termed Rathke's Pouch. Evidence from mouse knockout and ectopic expression studies indicates that 12.5 days post coitum (dpc) to 14.5 dpc is a critical period for the expansion of the progenitor cell pool and the determination of most hormone-secreting cell types. While signaling proteins and transcription factors have been identified as having key roles in pituitary cell differentiation, little is known about the identity and function of proteins that mediate signal transduction in progenitor cells. To identify genes that are enriched in the embryonic pituitary gland, we compared gene expression in 14.5 dpc pituitary and 14.5 dpc embryo minus pituitary tissues using the NIA 15K microarray. Analysis of the data using the R program revealed that the Regulator of G Protein Signaling 2 (Rgs2) gene was 3.9-fold more abundant in the 14.5 dpc pituitary. In situ hybridisation confirmed this finding, and showed that Rgs2 expression in midline tissues was restricted to the pituitary and discrete regions of the nervous system. Within the pituitary, Rgs2 was expressed in undifferentiated cells, and was downregulated at the completion of the hormone cell differentiation. To investigate Rgs2 function in the pituitary, we examined hormone cell differentiation in Rgs2 null neonate mice. Pituitary cell differentiation and morphology appeared normal in the Rgs2 mutant animals, suggesting that other Rgs family members with similar activities may be present in the developing pituitary. PMID- 15661636 TI - The amphioxus FoxQ1 gene is expressed in the developing endostyle. AB - The FoxQ1 genes form a distinct group within the Fox (also known as forkhead) gene family. We have isolated a gene from the amphioxus Branchiostoma floridae that encodes a forkhead domain with high identity to FoxQ1 genes in other chordates. Molecular phylogenetic analysis places AmphiFoxQ1 in a robust grouping with vertebrate FoxQ1 genes and with Ciona intestinalis Ci-FoxQ1. This group is separate from that containing AmphiFoxQ2, which instead groups with other invertebrate Fox genes. The expression of AmphiFoxQ1 was analysed by whole mount in situ hybridisation. The results show that AmphiFoxQ1 expression is confined to the developing endoderm, and specifically marks the endostyle and associated peripharyngeal bands of amphioxus larvae. Ci-FoxQ1 is also expressed in the endostyle, highlighting this as a conserved site of FoxQ1 gene expression in basal chordates. PMID- 15661637 TI - Stromal cell-derived factor-1 (SDF-1) expression in embryonic mouse cerebral cortex starts in the intermediate zone close to the pallial-subpallial boundary and extends progressively towards the cortical hem. AB - We describe the onset and the expansion of stromal cell-derived factor 1 (SDF-1) expression in the intermediate zone of embryonic mouse cerebral cortex between embryonic days (E)11.5 and 18.5, and on postnatal day 1. Using in situ hybridisation with a digoxigenin-labeled probe, SDF-1 mRNA was detectable by E 12.5 in a small area of the intermediate zone just dorsal to the pallial subpallial boundary. During the following days, SDF-1 expression extended towards the dorso-lateral pallium, and then the hippocampus and cortical hem. The position of the SDF-1 positive cells within the intermediate zone was closely correlated with the stream of tangentially migrating cells carrying the polysialylated form of neural cell adhesion molecule (PSA-NCAM). However, whereas these cells form a ventro-dorsal stream passing from the subpallium into the pallium, SDF-1 was not detectable on the ventral side of the pallial-subpallial boundary at any of the developmental stages studied. By E 16.5, the intensity of SDF-1 hybridisation signal in the intermediate zone decreased, to become undetectable by E 18.5. PMID- 15661638 TI - Comparative analysis of the development of the mandibular salivary glands and the labial silk glands in the mulberry silkworm, Bombyx mori. AB - The mulberry silkworm, Bombyx mori has a pair of salivary glands arising from the mandibular segment, in addition to the labial silk glands which are generally considered as modified salivary glands. Here we report the characterization of salivary glands and the comparative gene expression profiling of the silk and salivary glands. The two independent salivary glands made up by 330 cells, grow about 1000 fold during larval development. These individual glands extend up to the T(1) thoracic segment unlike silk glands with fused anterior ends and extending up to the caudal region. The salivary glands also undergo endomitosis resembling the silk glands. The B. mori homologue of the homeotic gene Deformed (BmDfd) was expressed in the mandibular and maxillary segments in stage 17 embryo and got localized to the centre of the mandibular segment at stage 18 to form the salivary gland placodes. The expression was also seen in the distal ends of the leg appendages after blastokinesis (stage 22). Only low variations in BmDfd expression ranging from 1.6 to 2.1 fold were apparent during embryonic development. BmDfd expression was observed in the salivary glands all through the larval instars but not in the silk glands. The transcription factor, Forkhead and the segment polarity gene, Wingless were expressed throughout the salivary glands, the latter confirming the absence of physiological compartmentation within these glands unlike the silk glands. The expression of Amylase and Fibrohexamerin was restricted to the salivary and silk glands, respectively and therefore, served as molecular markers for these tissues. PMID- 15661639 TI - Prominent transcription of zebrafish N-myc (nmyc1) in tectal and retinal growth zones during embryonic and early larval development. AB - Because of its oncogenic capacity and expression restricted to embryonic and newborn tissues, the N-myc proto-oncogene is suggested to play a key role in vertebrate organogenesis as well as in the control of cell proliferation and differentiation. To further approach the developmental function of N-myc, we cloned full-length zebrafish N-myc (nmyc1) and analyzed its expression in the embryo and early larva. nmyc1 transcription is initiated at the mid-blastula stage. At somitogenesis stages, its expression was detected in the retina, midbrain, posterior hindbrain and presumptive spinal cord. nmyc1 was also transcribed in the endoderm and its derivatives as well as in branchial arches. At later developmental stages, posterior neural expression of nmyc1 was switched off, but expression remained intense in the brain, mainly in the optic tectum, cerebellar plate and dorsal rhombomere 2. Comparison of nmyc1 transcription with proliferation zones using a M phase mitotic marker revealed that nmyc1 expression is specifically associated with mitosis in the optic tectum and the retina. This result contrasts with previous studies in other vertebrates where N-myc expression can persist in differentiating cells. PMID- 15661640 TI - Pattern of expression of the podocalyxin gene in the mouse brain during development. AB - We studied the expression pattern of the major renal protein Podocalyxin during the development of mouse brain using in situ hybridization. Podocalyxin mRNA was widely expressed at least from E14, the first age we studied, and expression remained high until adulthood. The highest levels of expression were postnatal. Podocalyxin expression was particularly elevated in the cortical plate, the hippocampus and cerebellum, and in several basal forebrain nuclei. PMID- 15661641 TI - The expression of plexins during mouse embryogenesis. AB - Plexins are large transmembrane proteins that are receptors for semaphorins, either alone or in a complex with neuropilin-1 or -2. Nine different mouse plexins have been found: Plexin-A1-4, -B1-3, -C1 and -D1. The expression and function of plexins in non-neuronal tissues has been poorly characterized, although Plexin-A1 has been shown to have a role during lung and cardiac morphogenesis. We have done an extensive non-radioactive in situ hybridisation survey of Plxna1-a4, Plxnb1 -b3 and Plxnc1 in E14 mouse embryo. At E14, Plxnb3 expression could not be detected by in situ hybridisation. All other plexins studied are widely expressed both in neuronal and non-neuronal tissues. We have also followed the expression patterns of plexins during the development of the kidney, tooth and testis. Plxnb1 and Plxnb2 are expressed in the immature glomeruli and mesenchyme of the developing kidney. In the tooth bud, Plxna1 and Plxnb1 are expressed in the oral epithelium, enamel knot and in both the inner and outer enamel epithelium, whereas the expression of Plxnb2 is more restricted to the inner enamel epithelium. In the testis, Plxna1, Plxnb1 and Plxnc1 are expressed in the developing sex chords. This study shows that during development, plexins are expressed in specific and distinct patterns also in non-neuronal tissues. PMID- 15661643 TI - Differential expression of the duplicated cellular retinoic acid-binding protein 2 genes (crabp2a and crabp2b) during zebrafish embryonic development. AB - The cellular retinoic acid-binding protein 2 (CRABP2) is believed to be involved in regulating access of retinoic acid to nuclear retinoic acid receptors. We have determined the cDNA sequence and the genomic organization of the duplicated crabp2 gene (crabp2b) in zebrafish. The crabp2b cDNA was 522bp in length and encodes a polypeptide consisting of 146 amino acids. Radiation hybrid mapping assigned the crabp2b gene to zebrafish linkage group 19. The comparison of the mapped human CRABP2 gene, zebrafish crabp2a and zebrafish crabp2b genes revealed that human chromosome 1 has a syntenic relationship to zebrafish linkage groups 16 and 19. Reverse transcription-polymerase chain reaction (RT-PCR) detected crabp2b mRNA in total RNA extracted from whole adult zebrafish, but not in any of the adult zebrafish tissues examined. The crabp2a mRNA was detected in total RNA extracted from whole adult zebrafish, adult zebrafish muscle, testes, and skin and to a lesser extent in heart, ovary and brain. No crabp2a mRNA-specific product was detected in kidney, liver or intestine of the adult zebrafish. Whole mount in situ hybridization detected crabp2b and crabp2a mRNA in a number of structures known to require retinoic acid signaling during embryonic development. The crabp2b mRNA was detected in the central nervous system, branchial arches, pectoral fins, retina (dorsal to the lens), epidermis and otic vesicle of the developing zebrafish. The crabp2a transcripts were detected by whole mount in situ hybridization in the central nervous system, epidermis, proliferative zone of the retina, intestinal bulb, oesophagus, pectoral fins and branchial arches during zebrafish embryonic development. PMID- 15661642 TI - Expression of cyp26b1 during zebrafish early development. AB - We have cloned the zebrafish ortholog of the mammalian cyp26b1 gene. The predicted zebrafish cyp26b1 protein shares greater than 73% identity with mammalian homologues. cDNA transfection assays showed that like human cyp26b1, zebrafish cyp26b1 is involved in limiting the activity of retinoic acid. Reverse transcription-polymerase chain reaction (RT-PCR) analysis of embryonic RNAs suggested that no maternal cyp26b1 message is detectable. Zygotic cyp26b1 message could be detected at 75% epiboly by RT-PCR and localized to presumptive rhombomere 3 and rhombomere 4 at the early two-somite (2S) stage (10.5 hpf: hour post fertilization) by whole mount in situ hybridization. As development proceeds expression expands anteriorly to include rhombomere 2 at the 10S stage (14hpf). By 14S (16hpf) expression in the hindbrain has also expanded posteriorly and encompasses rhombomere 2 through rhombomere 6. At later stages, 24 through 48 hpf, additional expression was found in the eyes, diencephalon, midbrain hindbrain boundary, cerebellum, pectoral fin and the pharyngeal arch primordia. PMID- 15661644 TI - Expression of AmHNF6, a sea star orthologue of a transcription factor with multiple distinct roles in sea urchin development. AB - The sea urchin transcription factor SpHNF6 is an early activator of differentiation genes in skeletogenic lineages and regulatory genes in the oral ectoderm. We report here the cloning and the expression of an orthologue of this gene, AmHNF6, from the sea star Asterina miniata. The vertebrate and the echinoderm hnf6 and onecut genes belong to the novel ONECUT homeo domain class of transcription factors. In blastula stage sea star embryos, AmHNF6 is expressed everywhere except around the vegetal pole. As is observed in sea urchin, by the end of gastrulation, the expression of AmHNF6 is distinctly localized to the ciliary bands. This terminal phase of expression has remained unchanged since the divergence of these two taxa half a billion years ago. PMID- 15661645 TI - NeuroM and MyoD are expressed in separate subpopulations of cells in the pregastrulating epiblast. AB - Epiblast cells form skeletal muscle and neurons in culture and some express mRNA for the skeletal muscle specific transcription factor MyoD in vivo. The following experiments were designed to determine whether the neurogenic transcription factor NeuroM is expressed in the epiblast and if NeuroM and MyoD are present in separate subpopulations of epiblast cells that can differentiate into neurons and muscle, respectively. In situ hybridization revealed that NeuroM was present in the anterior region of the pregastrulating epiblast. Some cells with NeuroM were proliferating and expressed two molecules present in neurogenic cells, NCAM and the Zn-12/HNK-1 carbohydrate. The G8 antibody labeled cells with MyoD but not NeuroM. When G8 positive cells were isolated by magnetic cell sorting and placed in culture, nearly all differentiated into skeletal muscle in serum free medium. A subpopulation of cells isolated with antibodies that bound to cells expressing NeuroM formed neurons when cultured in medium supplemented with sera and embryo extract. These experiments demonstrate that NeuroM and MyoD are present in separate subpopulations of cells in the pregastrulating epiblast. Epiblast cells with NeuroM are more dependent on exogenous factors to differentiate than those with MyoD. PMID- 15661646 TI - HvJNK, a Hydra member of the c-Jun NH2-terminal kinase gene family, is expressed during nematocyte differentiation. AB - C-jun NH(2)-terminal kinases (JNKs) represent a subgroup of mitogen-activated protein kinases (MAPKs). MAPK pathways are important regulators of cell proliferation, apoptosis, and gene expression throughout higher metazoans. We report here the characterization of a highly conserved Hydra JNK orthologue (HvJNK) that exhibits amino acid sequence identity of 61% as compared with human JNK1alpha. Phylogenetic analysis places HvJNK in a cluster with other metazoan JNKs. HvJNK is expressed in the nematocyte differentiation pathway. Double in situ hybridizations demonstrate overlapping expression with two other genes specifically activated during nematocyte differentiation, HyZic and Nowa, and restrict the phase of HvJNK expression to late proliferating nematoblasts and early differentiating nematocytes. Our results indicate that JNKs might have acted in cell differentiation in simple, pre-bilaterian animals. PMID- 15661647 TI - Dynamic localisation of KR-H during an ecdysone response in Drosophila. AB - The propagation of a hormonal response following an increase in titre involves intensive cross-talk between the products of the key regulatory genes. The Kr-h gene of Drosophila is a modulator of both the embryonic and metamorphic hierarchies of ecdysone responsive genes, but its mode of action is puzzling as mutants have both quantitative and qualitative (timing) effects on the ecdysone responses. We have used an antibody against KR-H to follow its distribution in larval tissues as they prepare for metamorphosis. While in most tissues protein levels remain stable, its distribution within salivary gland cells changes throughout the late larval ecdysone response and the ensuing prepupal period. We show that, at the chromosomal level, KR-H localisation is dynamic and that the protein is recruited to, and released from, loci harbouring an important subset of the known regulatory genes as the response advances. Such behaviour is most likely a conserved characteristic of hormonal responses. PMID- 15661648 TI - Distinct developmental expression of Drosophila retinoblastoma factors. AB - Retinoblastoma (RB) tumor suppressor proteins are important regulators of the cell cycle and are implicated in a wide variety of human tumors. Genetic analysis of RB mutations in humans and in model systems indicates that individual RB proteins also have distinct functions in development. Specific target genes or mechanisms of action of individual RB proteins in developmental contexts are not well understood, however. To better understand the developmental activities of the two RB family members in Drosophila, we have characterized endogenous expression patterns of Rbf1 and Rbf2 proteins and transcripts in embryos and imaginal discs. These gene products are coexpressed at several stages of development, however, spatial and temporal differences are evident, including partly complementary patterns of expression in the embryonic central nervous system. PMID- 15661649 TI - Molecular cloning and expression of Ena/Vasp-like (Evl) during Xenopus development. AB - Ena/VASP proteins are actin-binding proteins implicated in the regulation of axon guidance, platelet aggregation, cell motility, and cell adhesion. The vertebrate Ena/VASP family is comprised of three genes: Ena (Enabled), VASP (Vasodilator Stimulated Phosphoprotein), and Evl (Ena/VASP-Like). We have cloned and characterized cDNAs encoding three alternatively spliced isoforms of Xenopus laevis Evl, designated Xevl, Xevl-I and Xevl-H. Analysis of the temporal expression of Xevl, Xevl-I and Xevl-H demonstrates that transcripts for each isoform are first detectable at low levels at stage 18, show increased abundance by stage 23, and persist throughout the remainder of embryogenesis. In situ hybridization analyses using a probe that detects all three Xevl isoforms or a probe that specifically detects the Xevl-H isoform revealed expression in the cement gland, brain, neural tube, myotome, and neural placodes, including the otic, lateral line, and olfactory placodes. These results suggest roles for Xevl in regulating actin dynamics and cell adhesion in neural and mesodermal tissues during later stages of Xenopus development. PMID- 15661651 TI - cfm is a novel gene uniquely expressed in developing forebrain and midbrain, but its null mutant exhibits no obvious phenotype. AB - A novel gene, cfm, that is expressed uniquely during early forebrain and midbrain development was isolated, and its null mutant was generated. cfm does not have any known functional domains, but is conserved in human, chick, Xenopus and zebrafish; a site of phosphorylation by MAP kinase exists in one of the domains conserved among them. Its expression was initially found at the 5-somite stage in the future midbrain and caudal forebrain region. The expression in mesencephalon subsequently decreased, was found in a stripe in the mid mesencephalon at E9.0. The expression in diencephalon was restricted to the dorsal thalamic region by E9.5 and to epiphysis at E12.5. In mouse a cognate, cfm2, exists that is expressed uniquely in the somite just formed and the presomite to be segmented, but not in forebrain or midbrain during early development. However, the cfm null mutant was live-born without any apparent defects. PMID- 15661650 TI - Macho-1 functions as transcriptional activator for muscle formation in embryos of the ascidian Halocynthia roretzi. AB - Various kinds of maternal factor that play crucial roles in embryogenesis are present and localized in the ascidian egg cytoplasm. Localized maternal mRNA of the macho-1 gene is a muscle determinant in the embryo of the ascidian Halocynthia roretzi. The macho-1 protein has a zinc-finger domain and accumulates in nuclei, being expected to function as a DNA-binding transcription factor. In the present study, we show that macho-1 is, indeed, a DNA-binding transcriptional activator, and directly or indirectly regulates the expression of six downstream genes. Macho-1 was required and sufficient for expression of the muscle actin, myosin, calcium transporter, myogenic factor, Tbx6, and snail genes, whose expression is initiated in muscle blastomeres at the cleavage stages in normal embryos. Furthermore, when macho-1 conjugated with a transcription-repression domain of Drosophila engrailed (En(R)) was expressed in embryos, it repressed expression of these downstream genes. In contrast, expression of macho-1 fused with a transcription-activation domain of VP16 caused ectopic expression of these muscle genes in non-muscle blastomeres. PCR-assisted binding-site selection and gel-retardation assay showed that macho-1 protein binds to the consensus target sequence (TGGGTGGTC) for GLI/Zic-family proteins, and that three guanine residues with underlines are crucial for the specificity. The 5' promoter region of the muscle actin gene supported expression of the reporter gene only in muscle cells at late stage. By contrast, when the target sequence was added to the promoter, it well reproduced early expression of muscle actin at the cleavage stage, indicating that macho-1 can recognize the target sequence in vivo. In conclusion, the maternal muscle determinant macho-1 functions as a transcription factor that positively regulates gene expression for muscle formation in ascidian embryos. PMID- 15661652 TI - Secretin, a known gastrointestinal peptide, is widely expressed during mouse embryonic development. AB - The gastrointestinal functions of the 27-amino acid secretin peptide have been well established. In previous prenatal studies, secretin expression in the rat duodenum was reported after day 17 of gestation while its expression in other organs and its functions in the developing embryos are still unknown. By in situ hybridization and immunohistochemical staining, secretin transcripts and peptides were found to be widely expressed in mouse embryos. Consistent with the idea that secretin is a brain-gut peptide, its expressions are present in several developing brain regions such as cephalic mesenchyme, cerebellar primordium and choroid plexus as well as the epithelial villi lining and inner circular muscle of the developing intestine. Other than these organs, secretin was also detected in the developing heart including the ventricular epicardium and myocardium and certain structures of the developing kidney like ureteric bud, collecting duct and glomerulus. These observations strongly suggest for a functional role of secretin during mouse embryonic development. PMID- 15661654 TI - The role of SMC proteins in the responses to DNA damage. AB - The SMC proteins form the cores of three protein complexes in eukaryotes, cohesin, condensin and the Smc5-6 complex. Cohesin holds sister chromatids together after DNA replication and is involved in both the repair of double strand breaks by homologous recombination and the intra-S-phase checkpoint. Condensin assists in the condensation of chromosomes at mitosis and also has a role in checkpoint control pathways. The Smc5-6 complex is involved in a variety of DNA repair and damage response pathways by as yet unknown mechanisms, but is also associated with repair by homologous recombination. PMID- 15661655 TI - Insight into the functional consequences of hMYH variants associated with colorectal cancer: distinct differences in the adenine glycosylase activity and the response to AP endonucleases of Y150C and G365D murine MYH. AB - Escherichia coli MutY and its eukaryotic homologues play an important role in preventing mutations by removing adenine from 7,8-dihydro-8-oxo-2'-deoxyguanosine (OG):A mismatches. It has recently been demonstrated that inherited biallelic mutations in the genes encoding the human homologue of MutY (hMYH) are correlated with a genetic predisposition for multiple colorectal adenomas and carcinomas. The two most common hMYH variants found in patients with colorectal cancer are Y165C and G382D. In this study, we examined the equivalent variants in the murine MutY homologue (mMYH), Y150C and G365D. The Y150C mMYH enzyme showed a large decrease in the rate of adenine removal from both OG:A- and G:A-containing substrates, while G365D mMYH showed a decrease in the ability to catalyze adenine removal only with a G:A-containing substrate. Both mMYH variants exhibit a significantly decreased affinity for duplexes containing noncleavable 2' deoxyadenosine analogues. In addition, the human apurinic/apyrimidinic endonuclease (Ape1) stimulated product formation by wild-type and G365D mMYH with an OG:A substrate under conditions of multiple-turnover ([E]<[S]). In contrast, the presence of Ape1 nearly completely inhibited adenine removal by Y150C mMYH from the OG:A mismatch substrate. The more deleterious effect of Ape1 on the glycosylase activity of Y150C relative to G365D mMYH correlated with the more compromised binding affinity of Y150C to substrate analogue duplexes. These results suggest that the equivalent hMYH variants may be significantly compromised in substrate targeting in vivo due to a decrease in binding to substrate DNA; moreover, competition with other DNA binding proteins may further reduce the effective adenine glycosylase activity in vivo. PMID- 15661656 TI - Solution-state NMR investigation of DNA binding interactions in Escherichia coli formamidopyrimidine-DNA glycosylase (Fpg): a dynamic description of the DNA/protein interface. AB - Formamidopyrimidine-DNA glycosylase (Fpg) is a base excision repair (BER) protein that removes oxidative DNA lesions. Recent crystal structures of Fpg bound to DNA revealed residues involved in damage recognition and enzyme catalysis, but failed to shed light on the dynamic nature of the processes. To examine the structural and dynamic changes that occur in solution when Fpg binds DNA, NMR spectroscopy was used to study Escherichia coli Fpg free in solution and bound to a double stranded DNA oligomer containing 1,3-propanediol (13-PD), a non-hydrolyzable abasic-site analogue. Only 209 out of a possible 251 (83%) free-precession 15N/1H HSQC cross peaks were observed and 180 of these were assignable, indicating that approximately 30% of the residues undergo intermediate motion on the NMR timescale, broadening the resonances beyond detection or making them intractable in backbone assignment experiments. The majority of these affected residues were in the polypeptide linker region and the interface between the N- and C-terminal domains. DNA titration experiments revealed line broadening and chemical shift perturbations for backbone amides nearby and distant from the DNA binding surface, but failed to quench the intermediate timescale motion observed for free Fpg, including those residues directly involved in DNA binding, notwithstanding a nanomolar dissociation constant for 13-PD binding. Indeed, after binding to 13 PD, at least approximately 40% of the Fpg residues undergo intermediate timescale motion even though all other residues exhibit tight DNA binding characteristic of slow exchange. CPMG-HSQC experiments revealed millisecond to microsecond motion for the backbone amides of D91 and H92 that were quenched upon binding 13-PD. In free Fpg, heteronuclear 1H-15N NOE experiments detected picosecond timescale backbone motion in the alphaF-beta9 loop, the region primarily responsible for chemically discriminating 8-oxoguanine (8-oxoG) over normal guanine, that was quenched after binding 13-PD. Collectively, these observations reveal that, in solution, Fpg is a very dynamic molecule even after binding damaged DNA. Such motion, especially at the DNA binding surface, may be key to its processive search for DNA damage and its catalytic functions once it recognizes damaged DNA. PMID- 15661657 TI - XP-A cells complemented with Arg228Gln and Val234Leu polymorphic XPA alleles repair BPDE-induced DNA damage better than cells complemented with the wild type allele. AB - Functional effects of Arg228Gln and Val2343Leu XPA polymorphisms on benzo[a]pyrene-r-7,t-8-dihydrodiol-t-9,10-epoxide-(+/-)-anti (BPDE) survival and repair were investigated in SV40 immortalized XP12RO cells complemented with wild type and polymorphic XPA cDNAs in an inducible cDNA expression system. In contrast to previous studies showing little impact of XPA polymorphisms on UV survival and repair, cells complemented with polymorphic XPAs displayed improved BPDE survival and repair as compared to wild type XPA-complemented cells. Survival after BPDE treatment was measured using AlamarBlue reduction and colony forming ability. Cells expressing low levels of either polymorphic XPA had equivalent or improved survival compared to wild type XPA-complemented cells (XPAwt cells). XPA induction improved BPDE survival in Arg228Gln (R228Q cells) and Val234Leu (V234L cells) complemented cells, but not XPAwt cells. BPDE-induced DNA damage repair was measured both by reactivation after transfection of a luciferase reporter plasmid reacted with BPDE in vitro, and by removal of adducts from genomic DNA of BPDE-treated cells. BPDE-induced DNA damage repair in R228Q and V234L cells expressing XPA at very low levels was similar to repair in XPAwt cells expressing XPA at normal levels. XPA induction improved repair in R228Q and V234L cells but not in XPAwt cells. Our findings suggest that both Arg228Gln and Val234Leu XPAs function better than wild type XPA for BPDE adduct removal. These observations differ from UV repair results suggesting that the differences are lesion specific. The location of the polymorphisms within the putative poly(ADP ribose) binding domain suggests that poly(ADP-ribose) interaction is important in repair. PMID- 15661658 TI - Severe growth retardation and short life span of double-mutant mice lacking Xpa and exon 15 of Xpg. AB - In addition to xeroderma pigmentosum (XP), mutations in the human XPG gene cause an early onset of Cockayne syndrome (CS) in some patients (XP-G/CS) with characteristics, such as growth retardation and a short life span. In the previous studies, we generated four Xpg mutant mice with two different C-terminal truncations, null, or a base substitution mutation to identify the protein region that causes the onset of CS, and found that the CS-causing mutations, null or a deletion of the last 360 amino acids, completely inhibited the NER activity of mouse XPG (Xpg), but the non-CS-causing mutations, XpgD811A (base substitution that eliminates the nuclease activity of Xpg) or XpgDeltaex15 (deletion of the exon 15 corresponding to the last 183 amino acids), resulted in the retention of residual NER activity. To understand why mutations that completely eliminate the NER activity of Xpg cause CS but those that abolish the nuclease activity without totally eliminating the NER activity of Xpg do not result in CS, we made a series of Xpg mutant mice with Xpa-null mutant allele and found that mice with the non CS-causing deletion mutation (XpgDeltaex15) exhibited the CS phenotype when XPA was also absent but the base substitution mutation (XpgD811A) that eliminated the Xpg nuclease activity did not. These results indicate that Xpg has a second function, beside NER, and that the disruption of this second function (deletion of the last 183 amino acids) when combined with an NER defect causes CS. When we compared amino acid sequences corresponding to the exon 15 of Xpg, a significant homology was conserved among vertebrates, but not in Drosophila and Saccharomyces cerevisiae. These observations suggest that the second function of XPG may be conserved only in vertebrates and CS symptoms may occur in its absence. PMID- 15661659 TI - Differential incorporation of halogenated deoxyuridines during UV-induced DNA repair synthesis in human cells. AB - Double labeling of interphase and metaphase chromosomes by 5-chlorodeoxyuridine (CldU) and 5-iododeoxyuridine (IdU) has been used in studies of the dynamics of DNA replication. Here, we have used this approach and confocal microscopy to analyze sites of DNA repair synthesis during nucleotide excision repair (NER) in quiescent human fibroblasts. Surprisingly, we have found that when both precursors are added at the same time to UV-irradiated cells they label different sites in the nucleus. In contrast, even very short periods of simultaneous IdU+CldU labeling of S-phase cells produced mostly overlapped IdU and CldU replication foci. The differential labeling of repair sites might be due to compartmentalization of I-dUTP and Cl-dUTP pools, or to differential utilization of these thymidine analogs by DNA polymerases delta and epsilon (Poldelta and Polepsilon). To explore the latter possibility we used purified mammalian polymerases to find that I-dUTP is efficiently utilized by both Poldelta and Polepsilon. However, we found that the UV-induced incorporation of IdU was more strongly stimulated by treatment of cells with hydroxyurea than was incorporation of CldU. This indicates that there may be distinct IdU and CldU-derived nucleotide pools differentially affected by inhibition of the ribonucleotide reductase pathway of dNTP synthesis and that is consistent with the view that differential incorporation of IdU and CldU during NER may be caused by compartmentalization of IdU- and CldU-derived nucleotide pools. PMID- 15661660 TI - The multifunctional DNA repair/redox enzyme Ape1/Ref-1 promotes survival of neurons after oxidative stress. AB - Although correlative studies demonstrate a reduction in the expression of apurinic/apyrimidinic endonuclease/redox effector factor (Ape1/Ref-1 or Ape1) in neural tissues after neuronal insult, the role of Ape1 in regulating neurotoxicity remains to be elucidated. To address this issue, we examined the effects of reducing Ape1 expression in primary cultures of hippocampal and sensory neurons on several endpoints of neurotoxicity induced by H2O2. Ape1 is highly expressed in hippocampal and sensory neurons grown in culture as indicated by immunohistochemistry, immunoblotting and activity. Exposing hippocampal or sensory neuronal cultures to 25 or 50 nM small interfering RNA to Ape1 (Ape1siRNA), respectively, for 48 h, causes a reduction in immunoreactive Ape1 by approximately 65 and 54%, and an equivalent loss in endonuclease activity. The reduced expression of Ape1 is maintained for up to 5 days after the siRNA in the medium is removed, whereas exposing cultures to scrambled sequence siRNA (SCsiRNA) has no effect of Ape1 protein levels. The reduction in Ape1 significantly reduces cell viability in cultures 24 h after a 1-h exposure to 25 300 microM H2O2, compared to SCsiRNA treated controls. In cells treated with SCsiRNA, exposure to 300 microM H2O2 reduced cell viability by 40 and 30% in hippocampal and sensory neuronal cultures, respectively, whereas cultures treated with Ape1siRNA lost 93 and 80% of cells after the peroxide. Reduced Ape1 levels also increase caspase-3 activity in the cells, 2-3-fold, 60min after a 1-h exposure to 100 microM H2O2 in the cultures. Exposing neuronal cultures with reduced expression of Ape1 to 65 microM H2O2 (hippocampal) or 300 microM H2O2 (sensory) for 1h results in a 3-fold and 1.5-fold increase in the phosphorylation of histone H2A.X compared to cells exposed to SCsiRNA. Overexpressing wild-type Ape1 in hippocampal and sensory cells using adenoviral expression constructs results in significant increase in cell viability after exposure to various concentrations of H2O2. The C65A repair competent/redox incompetent Ape1 when expressed in the hippocampal and sensory cells conferred only partial protection on the cells. These data support the notion that both of functions of Ape1, redox and repair are necessary for optimal levels of neuronal cell survival. PMID- 15661661 TI - Product inhibition and magnesium modulate the dual reaction mode of hOgg1. AB - 8-Oxoguanine (8-oxoG) is a major mutagenic DNA base damage corrected by the base excision repair (BER) pathway, which is initiated by lesion specific DNA glycosylases. The human DNA glycosylase hOgg1 catalyses excision of 8-oxoG followed by strand incision 3' to the abasic site if cytosine is positioned in the complementary strand. Unlike most bifunctional glycosylases, hOgg1 uncouples base removal and strand cleavage. This paper addresses the significance of product inhibition and magnesium for the non-concerted action of hOgg1 activities. The enzymatic activities of hOgg1 were analysed on duplex DNA containing a single 8-oxoG or abasic site opposite cytosine. AP-lyase cleavage of abasic sites was inhibited in the presence of free 8-oxoG, indicating that the product of base excision inhibits the subsequent strand incision step. Assays with DNA containing 8-oxoG showed that free 8-oxoG also inhibited the glycosylase activity. This result suggests that the free 8-oxoG base may retain in the recognition site following N-glycosylic cleavage, implying that product inhibition contribute to uncoupling the activities of hOgg1. Magnesium reduced the efficiency of base excision and strand incision on DNA containing 8-oxoG under single turnover conditions; however, the reduction was more pronounced for the AP-lyase activity. Furthermore, Shiff-base formation between hOgg1 and 8-oxoG containing DNA was abrogated in the presence of magnesium. These results suggest that hOgg1 mainly operates as a monofunctional glycosylase under physiological concentrations of magnesium. PMID- 15661663 TI - Multiple PolK (POLK) transcripts in mammalian testis. AB - The Y-family of DNA polymerases are capable of translesion synthesis both in vitro and in vivo. The mouse/human gene (Polkappa/POLK) that encodes the Y-family member Polk is highly expressed in mouse/human testis. Using RT-PCR to amplify the coding sequence of the PolK gene, 11 forms of mouse PolK and 5 forms of human POLK transcripts were identified from testis. This phenomenon is apparently specific to PolK since several other specialized DNA polymerases capable of TLS do not express functional alternative transcripts in the testis. The multiple mouse/human (PolK/POLK) transcripts may encode multiple Polkappa isoforms in testis. PMID- 15661662 TI - Nucleosomal structure of undamaged DNA regions suppresses the non-specific DNA binding of the XPC complex. AB - The XPC protein complex is a DNA damage detector of human nucleotide excision repair (NER). Although the XPC complex specifically binds to certain damaged sites, it also binds to undamaged DNA in a non-specific manner. The addition of a large excess of undamaged naked DNA competitively inhibited the specific binding of the XPC complex to (6-4) photoproducts and the NER dual incision step in cell free extracts. In contrast, the addition of undamaged nucleosomal DNA as a competitor suppressed both of these inhibitory effects. Although nucleosomes positioned on the damaged site inhibited the binding of the XPC complex, the presence of nucleosomes in undamaged DNA regions may help specific binding of the XPC complex to damaged sites by excluding its non-specific binding to undamaged DNA regions. PMID- 15661664 TI - The evolution of truly social cognition: the core configurations model. AB - This article introduces a vocabulary suitable for evolutionary analyses in the human cognitive, social, and behavioral sciences. The vocabulary carves a middle way between advocates and critics of evolutionary perspectives by substituting the concept of repeated assembly for nature-nurture dualism. A model of core configurations-based on human morphology and ecology in human evolutionary history-is presented, and I argue that these configurations of face-to-face groups are the selective context for uniquely human mental systems. Hence, human cognition is "truly social,"" specialized for group living. The relevance of the core configuration model is illustrated with respect to two areas of interest to social psychologists: the self and social identity, and distributed cognition and shared reality. A final section illustrates the integrative power of the core configuration model with a brief comparison of the social and cognitive tasks faced by scientists and foragers. PMID- 15661665 TI - Estimates of social consensus by majorities and minorities: the case for social projection. AB - Meta-analyses of research on consensus estimation have identified an asymmetry in the error patterns between majorities and minorities (Gross & Miller, 1997; Mullen & Hu, 1988). Members of the majority slightly underestimate the size of their own group, whereas members of the minority strongly overestimate the size of theirs. The present analysis shows that a single psychological assumption about projection is sufficient to explain this asymmetry. Most people, regardless of whether they are actually members of the majority or members of the minority, believe themselves to be in the majority. It is not necessary to attribute different psychological mechanisms, such as ego protection or cognitive availability, to majority and minority members. A simple quantitative model and empirical data illustrate this point. PMID- 15661666 TI - Reciprocal strategies for large groups. AB - Reciprocal strategies have been shown to be effective in inducing long-term cooperation in social dilemmas involving small, but not large, groups. In this article, we argue that group-based reciprocal strategies (GBRS)--under which one reciprocates cooperative factions, not individuals, within the group--can induce cooperation in large groups. Results of two computer simulations of 100-member groups show that a GBRS accumulates a significantly larger payoff than do other commonly advocated mixed-motive strategies. A third simulation showed that the size of the cooperative faction is influenced by the nature of the payoff matrix; when there is considerable temptation to defect, one should cooperate only after a large number of others have done so, but if there is little temptation to defect, one should reciprocate the cooperative actions of a smaller faction. We also found that the use of a "win-cooperate, lose-defect" heuristic led to substantial total outcomes. Our results suggest several psychological factors are important in facilitating cooperation, yet need empirical study. PMID- 15661667 TI - The psychology of legitimacy: a relational perspective on voluntary deference to authorities. AB - People within organized groups often internalize their feelings of obligation to obey group rules and the decisions of group authorities. They believe that group authorities and rules are legitimate and, hence, entitled to be obeyed. Because of this belief, group members voluntarily accept and obey rules and decisions from group authorities. This review draws on evidence from studies of authorities in political, legal, managerial, educational, and family settings to explore why people view as legitimate and voluntarily defer to group authorities. Two theories about legitimacy are contrasted: resource-based theories, represented by instrumental models, and identification-based theories, represented by the relational model. The findings provide strong support for the existence of a relational component of legitimacy, suggesting that authorities draw an important part of their legitimacy from their social relationship with group members. The findings also show that there is an instrumental component to legitimacy. Hence, the psychology of legitimacy involves both instrumental and relational elements. PMID- 15661668 TI - The accuracy-confidence correlation in the detection of deception. AB - A meta-analysis was conducted of research on the relation between judges' accuracy at detecting deception and their confidence in their judgments. A total of 18 independent samples revealed an average weighted accuracy-confidence correlation of .04, a relation not significantly different from zero. However, confidence was positively correlated with judges' tendency to perceive messages as truthful, regardless of the actual truthfulness of the messages. Judges were also more confident when they really were rating truths compared to when they were rating lies. Also, men were more confident than women, and judges who had a closer relationship to the message sender felt more confident in their judgments of truths and lies. Methodological and theoretical explanations for these findings are discussed. PMID- 15661669 TI - Harming other people: perspectives on evil and violence. PMID- 15661670 TI - The roots of evil: social conditions, culture, personality, and basic human needs. AB - Evil actions are defined as repeated or persistent, not commensurate with provocation and causing extreme harm, at times due to repetition. Evil develops or evolves. As individuals and groups harm others, they tend to develop characteristics that make further and more intense harmdoing probable. In this article, I explore instigating conditions (difficult life conditions in a society, group conflict); cultural characteristics; the nature of evolution, with its psychological and social processes in individuals and groups; and the passivity and complicity of bystanders that lead to genocide and other collective violence. I consider the question of whether bystanders can be regarded as evil, focusing on the genocide in Rwanda as an example. I examine the socialization and experience of children and youth that lead to aggression and the subsequent evolution of aggression toward greater violence and evil. I explore the way personal characteristics and a system of relationships can lead to sexual abuse by fathers. One organizing concept in understanding the generation of violence that causes extreme harm is the frustration of basic human needs and their subsequent destructive fulfillment. PMID- 15661671 TI - Moral disengagement in the perpetration of inhumanities. AB - Moral agency is manifested in both the power to refrain from behaving inhumanely and the proactive power to behave humanely. Moral agency is embedded in a broader sociocognitive self theory encompassing self-organizing, proactive, self reflective, and self-regulatory mechanisms rooted in personal standards linked to self-sanctions. The self-regulatory mechanisms governing moral conduct do not come into play unless they are activated, and there are many psychosocial maneuvers by which moral self-sanctions are selectively disengaged from inhumane conduct. The moral disengagement may center on the cognitive restructuring of inhumane conduct into a benign or worthy one by moral justification, sanitizing language, and advantageous comparison; disavowal of a sense of personal agency by diffusion or displacement of responsibility; disregarding or minimizing the injurious effects of one's actions; and attribution of blame to, and dehumanization of, those who are victimized. Many inhumanities operate through a supportive network of legitimate enterprises run by otherwise considerate people who contribute to destructive activities by disconnected subdivision of functions and diffusion of responsibility. Given the many mechanisms for disengaging moral control, civilized life requires, in addition to humane personal standards, safeguards built into social systems that uphold compassionate behavior and renounce cruelty. PMID- 15661672 TI - The intrinsic appeal of evil: sadism, sensational thrills, and threatened egotism. AB - Three main sources of intrinsic appeal and satisfaction from performing violent acts are described. First, sadism involves deriving pleasure directly from the suffering of the victim. An opponent-process model is suggested. Second, the quest for thrilling sensations to escape from boredom can produce violent acts, including many in which the harmful consequences were not intended. Third, threatened egotism entails that one's favorable view of self (orpub1ic image) has been attacked, and violent responses are directed toward the source of this attack. Relevant individual differences (respectively, low guilt, high sensation seeking, and narcissism) moderate these patterns. Analyzing the intrinsic appeal of evil acts is a useful complement to analyzing situational determinants of violence. PMID- 15661673 TI - The second face of evil: wrongdoing in and by the corporation. AB - The second face of evil is hurtful, rather than malevolent. We argue that this is likely to be the form of wrongdoing characteristic of corporate actors. This article views the corporation at 3 levels: the individual self-interested actors within it, the individual actors who respond to their hierarchical position, and the hierarchy (corporation) itself. Research shows that action inside complex organizations such as corporations is not necessarily more rational and informed by greater foresight than individual actions, but several studies indicate that individuals hold organizations to a higher level of responsibility than individuals committing similar acts, and they do so in part because they perceive organizations to enjoy greater foresight. We address this paradox by suggesting that future research address how the corporation's obligations to foresee consequences and the sheer, primitive consequences themselves may shape perceptions of corporate foreseeability. PMID- 15661674 TI - The social construction of violence: the case of sexual and domestic violence. AB - What counts as "violence" is socially constructed, has varied over time, and reflects power relationships. Informed by social constructionism, we illustrate these points using as examples sexual violence and domestic violence. We review changes in how society and social scientists have defined and understood these topics during the last 30 years. We then discuss 3 areas of continuing controversy: who should decide if sexual or domestic violence has occurred, what to count as sexual and domestic violence, and the role of gender in defining sexual and domestic violence. PMID- 15661675 TI - Evil is more than banal: situationism and the concept of evil. AB - Social psychology as a discipline has given relatively little attention to the problem of evil in society, and those discussions in this field that do exist typically regard evil actions as only varieties of aggression without any characteristics that distinguish them from other forms of intentional mistreatment of others. Because of the field's situationistic perspective emphasizing the individual's susceptibility to the power of the immediate situation, social psychoiogists generally view the fairly high levels of obedience to authority displayed in Milgram's (1963, 1974) classic experiment as the paradigmatic example of evil behavior. For them, much evil is, in Arendt's (1963) well-known phrase, only "banal," and Milgram's findings are often viewed as illustrating the "central dynamic" involved in the slaughter of millions of Jews and other "undesirables" in the Holocaust. This article holds that Milgram's (1974) obedience research does not represent significant features of the Holocaust, especially the sadism that occurred not infrequently, and disregards the vital difference between those who initiated the murderous policy and the others who followed their orders. Building on Darley's (1992) earlier conjectures about the features that ordinary people might consider in judging whether any given action is evil, I suggest that many persons have a prototypic conception of evil and speculate about the dimensions that could be involved in this prototype. PMID- 15661676 TI - Accounting for evil and cruelty: is to explain to condone? AB - Analysts of evil and violence express the concern that to explain harmdoing may result in a condoning attitude toward perpetrators. An examination of research relevant to this hypothesis suggests that there are a variety of cognitive and affective processes that may produce a relatively condoning attitude toward perpetrators as a result of explaining their actions. Evidence from 3 exploratory studies supported the exonerating effects of explanations. Participants generating explicit explanations of harmdoing displayed a more condoning attitude toward pelpetrators than did those forming impressions of perpetrators without first explaining the acts. Participants reading social-psychological explanations of harmdoing also judged the researcher to be more condoning of perpetrators than those reading dispositional explanations of the same behavior. Implications of these findings are discussed. PMID- 15661677 TI - Methods for the study of evil-doing actions. AB - For obvious ethical reasons, experimental studies of severe harm-doing actions are precluded. What methods are available to experimental social psychologists for the study of harm- and evil-doing activities? Three are suggested: experiments that may have a component of role-playing but still can illuminate nodes in the socialization into harm-doing process, probes into the conceptual world of individuals who are enlisted into real-world harm-doing socialization processes, and secondary analyses of case studies written by those who have been caught up in harm doing. The methodological limits of each activity are examined, and it is argued that an approach in which combinations of methods are employed to arrive at theoretical constructions can both support generalizations that provide insights into the socialization process and be sufficiently rigorous to support prudent social action recommendations. PMID- 15661678 TI - Socially shared cognition, affect, and behavior: a review and integration. AB - In this article, we review 4 classes of models of socially shared cognition and behavior: supraindividual models, information-processing models, communication models, and social interaction models. Our review draws on research and theory in social psychology, sociology, and organization behavior. We conclude that these innovative perspectives on socially shared behavior represent a new approach to the study of groups and are distinct from traditional models of the group mind and crowd behavior. The key processes implicated in these models focus on the potency of immediate interaction, reciprocal influence processes between individuals and groups, goal-directed behavior, negotiated processing of information and ideas, and the maintenance and enhancement of social identity. This approach to socially shared understanding is not antagonistic toward the analysis of individual-level processes but rather maintains that individual-level processes are necessary but not sufficient to build a social psychology of shared understanding. PMID- 15661679 TI - The foot-in-the-door compliance procedure: a multiple-process analysis and review. AB - Research on the social compliance procedure known as the foot-in-the-door (FITD) technique is reviewed. Several psychological processes that may be set in motion with a FITD manipulation are identified: self-perception, psychological reactance, conformity, consistency, attributions, and commitment. A review of relevant investigations and several meta-analyses support the notion that each of these processes can influence compliance behavior in the FITD situation. I argue that the combined effects of these processes can account for successful FITD demonstrations as well as studies in which the technique was ineffective or led to a decrease in compliance. The experimental conditions most likely to produce an FITD effect are identified. PMID- 15661680 TI - Achievement goals and intrinsic motivation: a meta-analytic review. AB - This article presents a meta-analysis of the experimental literature that has examined the effect of performance and mastery achievement goals on intrinsic motivation. Summary analyses provided support for the hypothesis that the pursuit of performance goals has an undermining effect on intrinsic motivation relative to the pursuit of mastery goals. Moderator analyses were conducted in an attempt to explain significant variation in the magnitude and direction of this effect across studies. Results indicated that the undermining effect of performance goals relative to mastery goals was contingent on whether participants received confirming or nonconfirming competence feedback, and on whether the experimental procedures induced a performance-approach or performance-avoidance orientation. These findings provide conceptual clarity to the literature on achievement goals and intrinsic motivation and suggest numerous avenues for subsequent empirical work. PMID- 15661681 TI - Asch's social psychology: not as social as you may think. AB - This article discusses 2 commonly held ideas about Solomon Asch's work in social psychology: (a) Asch was primarily interested in social phenomena in general and in group processes in particular, and (b) Asch was a forerunner of social cognition. Asch's studies on social influence were translations of strictly perceptual experiments. For him, social stimuli had no specificity relative to physical ones provided that the perceptual context presented similar structural properties. Moreover, and contrary to Kurt Lewin (e.g., 1948) Asch focused his attention at the individual level and may have slowed down interest in social interactions or group processes. Asch's studies on impression formation presaged the social cognition approach. In his work, he foresaw the importance of online processing of information, the existence of implicit theories of personality, as well as perception based on exemplars and prototypes. However, Asch's reliance on immediate perceptual experience, on isomorphism between the properties of the external object and the phenomenal experience of this object, and his holistic and dynamic perspective clash with the main stream of social cognition research. PMID- 15661682 TI - Solomon Asch's legacy for group research. AB - Asch's work has had a profound impact on how psychologists think about and study social influence in groups. To appreciate this impact, we must go beyond his classic conformity experiments and consider his broader theoretical framework. This article examines 4 of Asch's ideas that have proven to be particularly influential in later efforts to understand social influence in groups: (a) Social interaction depends on individuals' ability to represent others' positions, define themselves as members of the same group, and regulate their behavior in terms of the norms and values of the group; (b) independence is critical to effective group functioning; (c) independence and conformity are not simply mirror images that can be explained in terms of a unitary psychological process; and (d) change of meaning is an important mechanism of social influence. Finally, Asch's role as a theorist and researcher in the wider area of group dynamics is considered. PMID- 15661683 TI - Disease without borders. PMID- 15661684 TI - Cancer statistics, 2005. AB - Each year, the American Cancer Society estimates the number of new cancer cases and deaths expected in the United States in the current year and compiles the most recent data on cancer incidence, mortality, and survival based on incidence data from the National Cancer Institute and mortality data from the National Center for Health Statistics. Incidence and death rates are age-standardized to the 2000 US standard million population. A total of 1,372,910 new cancer cases and 570,280 deaths are expected in the United States in 2005. When deaths are aggregated by age, cancer has surpassed heart disease as the leading cause of death for persons younger than 85 since 1999. When adjusted to delayed reporting, cancer incidence rates stabilized in men from 1995 through 2001 but continued to increase by 0.3% per year from 1987 through 2001 in women. The death rate from all cancers combined has decreased by 1.5% per year since 1993 among men and by 0.8% per year since 1992 among women. The mortality rate has also continued to decrease from the three most common cancer sites in men (lung and bronchus, colon and rectum, and prostate) and from breast and colorectal cancers in women. Lung cancer mortality among women has leveled off after increasing for many decades. In analyses by race and ethnicity, African American men and women have 40% and 20% higher death rates from all cancers combined than White men and women, respectively. Cancer incidence and death rates are lower in other racial and ethnic groups than in Whites and African Americans for all sites combined and for the four major cancer sites. However, these groups generally have higher rates for stomach, liver, and cervical cancers than Whites. Furthermore, minority populations are more likely to be diagnosed with advanced stage disease than are Whites. Progress in reducing the burden of suffering and death from cancer can be accelerated by applying existing cancer control knowledge across all segments of the population. PMID- 15661685 TI - American Cancer Society Guidelines for the Early Detection of Cancer, 2005. AB - Each January, the American Cancer Society (ACS) publishes a summary of its recommendations for early cancer detection, including guideline updates, emerging issues that are relevant to screening for cancer, and a summary of the most current data on cancer screening rates for US adults. In 2004, there were no updates to ACS guidelines. In this article, we summarize the current guidelines, discuss recent evidence and policy changes that have implications for cancer screening, and provide an update of the most recent data pertaining to participation rates in cancer screening by age, gender, and insurance status from the Centers for Disease Control and Prevention's Behavioral Risk Factor Surveillance System. PMID- 15661686 TI - Molecular epidemiology of cancer. AB - Epidemiology is very successful in identifying environmental and lifestyle factors that increase or reduce risk of specific cancers, leading to cancer prevention strategies. However, the etiology of many types of cancer is still poorly understood, despite extensive use of questionnaires and interview-based approaches in conventional epidemiologic studies. The integration of molecular techniques into epidemiology studies may provide new insights and has been referred to as molecular epidemiology. For instance, our ability to make connections between lifestyle and cancer risk is limited by difficulty in accurately measuring exposure to many carcinogens-newer molecular markers of exposure may provide better information. The completion of the Human Genome Project gives us knowledge of the genetic variations that presumably underlie the fact that a family history of cancer is a risk factor for most cancer types. Some of this excess risk has been explained over the last decade by identification of mutations in genes that give rise to a very high familial risk. Molecular epidemiologists are searching for genes that may give rise to much smaller increases in individual risk, but account for much of the residual risk associated with family history. These genes may also interact with environment and lifestyle factors such that cancer risk is not equally elevated in all persons exposed to an environmental factor (but not genetically susceptible), or all gene carriers (but not exposed to the environmental factor). Molecular markers may help to differentiate tumors with the same histologic appearance into different etiologic subtypes. Finally, response to treatment may be determined by molecular subtypes of the tumor, or inherited variation in drug metabolism. Examples will be given of how use of molecular techniques is informative in epidemiological studies of cancer and is predicted to lead to improvements in cancer incidence, early detection, and mortality. PMID- 15661687 TI - The silver anniversary of the American Journal of Neuroradiology: twenty-five years of documenting advances in neuroradiology. PMID- 15661688 TI - Functional neuroradiology: a call to action. PMID- 15661689 TI - What is right about MRI permeability studies. PMID- 15661690 TI - Development of the C1-C2 puncture in neuroradiology: a historical note. PMID- 15661692 TI - Experimental autoimmune encephalomyelitis in the rat spinal cord: lesion detection with high-resolution MR microscopy at 17.6 T. AB - BACKGROUND AND PURPOSE: Experimental autoimmune encephalomyelitis (EAE) is an inflammatory demyelinating disorder of the CNS and an animal model of multiple sclerosis. We used high-field MR microscopy at 17.6 T to image spinal cord inflammatory lesions in the acute stage of chronic relapsing rat EAE. We sought to compare lesions detected on MR imaging with histopathologic findings and to quantify the inflammatory lesion load. METHODS: Imaging of fixed spinal cord specimens was performed by using a 3D gradient-echo sequence with a spatial resolution of 35 x 35 x 58 microm3 and a total imaging time of 5.5 hours. Histopathologic analysis was performed by staining axial sections with hematoxylin-eosin or Luxol fast blue to identify cellular infiltration and demyelination. RESULTS: Clinical signs of EAE occurred on days 10-14 after immunization. On day 22, healthy white matter and gray matter were differentiated by high contrast on T2*-weighted images, with white matter lesions appearing as hyperintense areas in the normal-appearing white matter. Inflammatory lesions identified on histopathologic evaluation were readily detected with MR imaging and vice versa. MR imaging and histopathologic analysis had excellent correlation regarding the extent of white matter lesions. Inflammatory infiltrates of gray matter were not detectable with MR imaging. Using a semiautomatic segmentation of the acquired MR data, we could quantify white matter lesion load. CONCLUSION: Ex vivo high-resolution MR microscopy of the spinal cord at 17.6 T allows rapid and highly accurate determination of CNS inflammation by demonstrating virtually all histologically detectable white matter inflammatory lesions. PMID- 15661691 TI - Apparent diffusion coefficients in spinal cord transplants and surrounding white matter correlate with degree of axonal dieback after injury in rats. AB - BACKGROUND AND PURPOSE: Abnormal apparent diffusion coefficient (ADC) values in injured spinal cord white matter and fibroblast transplants have been shown to correspond with qualitative histologic findings of axonal loss or regeneration. We proposed that ADC values would correlate with quantitative axonal tracing in the transected rubrospinal tract (RST). METHODS: Eleven rats received right-sided lateral funiculus lesions at C3-4 (disrupting the RST) and transplantation of fibroblasts that were unmodified or modified to secrete brain-derived neurotrophic factor (BDNF). Behavioral tests measured hindlimb function at 1, 2, 4, 6, 8, 10, and 12 weeks after injury. At 12 weeks after injury, the antegrade axon tracer biotinylated dextran amine was stereotactically injected into the red nucleus to label the injured RST axons. Animals were sacrificed 2 weeks later. Diffusion-weighted MR imaging of the excised, fixed spinal cord specimens was then performed at 9.4 T. RESULTS: In white matter surrounding transplants, ADC values transverse to axons were elevated and ADC values longitudinal to axons were decreased. These ADC values were more abnormal closer to the transplant, and this correlated with decreases in numbers of labeled RST axons. ADC values in BDNF-expressing fibroblast transplants were significantly lower than those in unmodified fibroblast transplants, and these lower values correlated with decreased axonal dieback. Behaviorally, all animals showed partial recovery, but animals with BDNF-expressing fibroblast transplants had slightly improved hindlimb function compared to those with unmodified fibroblast transplants. CONCLUSION: ADC values may be able to evaluate graft function after spinal cord injury by demonstrating the degree of axonal dieback and preservation. PMID- 15661693 TI - West Nile Virus: a case report with flaccid paralysis and cervical spinal cord: MR imaging findings. AB - We present a case of serologically proved West Nile virus (WNV) flaccid paralysis of the right upper extremity. Radiologic correlation revealed striking T2 hyperintensities in the anterior horns of the cervical spinal cord, similar to those seen in cases of poliomyelitis. Recognition of the MR imaging findings in cases of WNV flaccid paralysis can provide early evidence of infection. PMID- 15661694 TI - Percutaneous treatment of pediatric aneurysmal bone cyst at C1: a minimally invasive alternative: a case report. AB - An 11-year-old girl presenting with neck pain was diagnosed as having an aneurysmal bone cyst involving the posterior arch of C1. Minimally invasive treatment was performed with percutaneous injections of calcitonin and methylprednisolone. Two injections separated in time by 2 months were performed with complete sclerosis and ossification of the cyst over a period of 6 months. Such percutaneous sclerosis offers an important primary therapeutic option or an adjunct to surgery of these otherwise vascular cystic masses and should be contemplated as the initial step in the treatment of these benign tumors, especially those in the spine where surgical treatment can involve complex resection and fixation procedures. PMID- 15661695 TI - Intradural spinal vein enlargement in craniospinal hypotension. AB - We present a case of craniospinal hypotension in a 45-year-old woman with an associated epidural pseudomeningocele extending the entire length of the spine. The epidural pseudomeningocele was caused by a CSF leak at the T8 level. In addition to typical low-pressure symptoms, the epidural pseudomeningocele caused atypical symptoms characterized by positional thoracic radiculopathy. Craniospinal hypotension was associated with massive cervical epidural venous engorgement, as well as enlargement of the posterior spinal cord vein, which was reminiscent of a dural arteriovenous fistula at CT myelography. Enlargement of the posterior spinal vein is explained by the Monro-Kellie hypothesis, and the spinal analog to enlarged cerebral veins known to be associated with intracranial hypotension. PMID- 15661696 TI - MR imaging findings in cauda equina gnathostomiasis. AB - We report a case of cauda equina syndrome caused by Gnathostoma spinigerum, which was confirmed by an immunoblotting test. MR imaging of the lumbosacral spine showed long, segmented hyperintensity along the cauda equina with irregular enhancement on the postcontrast study. The conus medullaris was slightly enlarged with abnormal enhancement. The patient was treated with corticosteroids, and her clinical condition improved. MR imaging, 9 months after treatment, showed the condition to be completely resolved. PMID- 15661697 TI - CT fluoroscopic-guided cervical nerve root blocks. AB - While both fluoroscopic and CT-guidance during cervical nerve root blocks have been well documented in the literature, the use of CT fluoroscopy (CTF) has not. CTF is well suited to provide imaging guidance during these procedures due to its combination of excellent anatomic detail, relatively low radiation dose and the ability to perform an initial dynamic contrast injection, and is a viable alternative to fluoroscopic guidance. Details of the technique along with the initial experience at one institution are presented. PMID- 15661698 TI - MR quantitation of volume and diffusion changes in the developing brain. AB - BACKGROUND AND PURPOSE: Brain volume and diffusion change during maturation. Quantitation of these changes may be helpful in understanding normal brain development. We used diffusion-weighted imaging to characterize the volumetric and diffusion changes in vivo. METHODS: We recruited 30 pediatric volunteers (aged 1 month-17 years; 14 male, 16 female). Diffusion was measured in three orthogonal directions with a b value of 1000 s/mm2. The diffusion parameters from the entire brain were calculated and fitted to a triple gaussian model. In addition, region-of-interest measurements were made in caudate, thalamus, genu and splenium of the corpus callosum, and periventricular white matter (PVWM). The brain volume was measured by counting pixels and by using the model. RESULTS: Water diffusion of the whole brain, caudate, thalamus, genu and splenium of the corpus callosum, and PVWM decreased during maturation, with the most significant change within the first 2 years. Robust negative correlations were found between age and the measured average diffusion constant (Dav) values in each of the measured locations (P <.005). Volumes of different cerebral compartments and the total intracranial volume (ICV) increased rapidly during the first 2 years of life and then had a slower growth process through adolescence. Age was correlated with the ICV and the volume of each brain compartment (P <.005). CONCLUSION: Brain diffusion decreases and brain volume increases during maturation, with the most significant changes occurring within the first 2 years of life. The brain model used in this study provides a good estimate of the increasing brain volume. PMID- 15661699 TI - MR venography in the pediatric patient. AB - BACKGROUND AND PURPOSE: Little is known about age-related changes in posterior fossa venous anatomy on 2D time-of-flight MR venography (MRV) or about artifacts that limit its accuracy in diagnosing venous occlusive disease. We evaluated pediatric appearances of posterior fossa venous drainage. METHODS: One hundred and eight children with normal MR imaging or minimal congenital anomalies underwent 2D MRV. Transverse sinus dominance and absence and the presence of an occipital sinus were correlated with age. Venous structure conspicuity was compared on source and maximum intensity projection images. RESULTS: Right, left, and codominance of the transverse sinus, respectively, was as follows: at < 25 months, 37%, 21%, and 42%; 25 months to 5 years, 35%, 30%, 35%; and > or =6 years, 50%, 16%, 34%. Transverse sinus dominance was not related to age between the three groups (P=.58, chi-square contingency), but some relationship was observed when patients <6 years were compared to those > or =6 years (P=.032). Chi-square trends showed a mildly positive correlation between age and an absent transverse sinus (P=.026) and a decreasing trend in the presence of an occipital sinus with age (P=.038). Saturation effects due to in-plane/slow flow were worse in patients <25 months; effects in the transverse sinuses or internal jugular veins were miminized with coronal or axial imaging, respectively. CONCLUSION: 2D TOF MRV shows age-related changes in venous anatomy. Caution should be used before posterior fossa venous occlusive disease is diagnosed on the basis of signal intensity loss, especially in neonates and young infants. PMID- 15661700 TI - Spontaneous partial thrombosis of a basilar artery giant aneurysm in a child. AB - We report the complete, spontaneous obliteration of a partially thrombosed dissecting giant aneurysm in the basilar artery by occlusion of both the lumen of the aneurysm and the parent artery in a 15-year-old girl. PMID- 15661701 TI - Spontaneous thrombosis of a traumatic posterior cerebral artery aneurysm in a child. AB - Traumatic posterior circulation aneurysms are rare, especially in children. They are typically associated with severe trauma and are thought to result from either direct osseous injury or stretching or compression of an artery against the tentorium. They may grow, rupture, or both. Although spontaneous thrombosis may occur, it rarely results in complete occlusion. We report the spontaneous and complete thrombosis of a large posterior cerebral artery aneurysm in a child who presented after minor head trauma. Five-year follow-up documents the complete occlusion of the aneurysm. PMID- 15661702 TI - Malformations of cortical development: high-resolution MR and diffusion tensor imaging of fiber tracts at 3T. AB - Patients with malformations of cortical development and epilepsy may have a variety of abnormal brain findings, including abnormal gyral patterns, cortical thickening, decreased volume of white matter, and increased diffusion of white matter. The status of individual white matter fiber tracts, however, is unknown. We present a case of bilateral frontal schizencephaly and subcortical heterotopia and illustrate alterations of white matter fascicles by combined structural and functional diffusion tensor imaging at 3 T. PMID- 15661703 TI - Hypothalamic hamartoma associated with a craniopharyngeal canal. AB - Hypothalamic hamartoma is a rare congenital lesion. We present the case of a 7 year-old girl who suffered from precocious puberty, the cause of which was diagnosed by using MR imaging and CT as pedunculated hypothalamic hamartoma associated with a large craniopharyngeal canal and sellar spine mimicking pituitary duplication. PMID- 15661704 TI - Sonographic NASCET index: a new doppler parameter for assessment of internal carotid artery stenosis. AB - BACKGROUND AND PURPOSE: Established Doppler parameters for carotid stenosis assessment do not reflect North American Symptomatic Carotid Endarterectomy Trial (NASCET)-style methodology. We derived a Doppler parameter, termed sonographic NASCET index (SNI), and hypothesized that the SNI would provide greater angiographic correlation and better accuracy in predicting stenosis of 70% or greater than that of currently used peak systolic velocity (PSV) measurements. METHODS: Inclusion criteria of angiographically proved carotid stenoses of 40-95% and measured proximal and distal internal carotid artery Doppler PSV values were established. Occlusions and near occlusions were specifically excluded. Doppler and angiographic data meeting the inclusion criteria from 32 carotid bifurcations were identified; actual angiographic stenoses ranged 40-89%. SNI values were calculated for each vessel. PSV and SNI were correlated with angiography by using linear regression analysis. Accuracies of SNI and PSV in predicting stenosis of 70% or greater were compared at two thresholds. RESULTS: Correlation between SNI and angiography was superior to that between PSV and angiography (r2=0.64 vs 0.38). PSV and SNI values that corresponded to 70% angiographic stenosis were 345 cm/s and 45.5, respectively. Accuracy of PSV of 345 cm/s or greater in predicting stenosis of 70% or greater was 78%, compared with 88% for SNI of 45.5 or greater. The SNI value that corresponded to a PSV threshold of 250 cm/s was 33. Accuracy of PSV of 250 cm/s or greater in predicting stenosis of 70% or greater was 81%, compared with 88% for SNI of 33 or greater. CONCLUSION: Correlation between SNI and angiography was greater than that between PSV and angiography. Accuracy of SNI in predicting stenosis of 70% or greater was also superior to that of PSV at two thresholds. These results suggest that SNI may be a better predictor of high grade carotid stenosis than is PSV. PMID- 15661706 TI - Early postoperative CT scanning for juvenile nasopharyngeal angiofibroma: detection of residual disease. AB - BACKGROUND AND PURPOSE: High recurrence rates and early recurrence have been reported for juvenile nasopharyngeal angiofibroma (JNA) involving the skull base. The purpose of this study was to evaluate the diagnostic accuracy of contrast enhanced CT scanning for the detection of residual disease (RD) in the early postoperative course after surgical excision of JNA. METHODS: We retrospectively reviewed data in 20 male patients (mean age +/- SD, 15.4 +/- 5 years; range, 10 32 years) who underwent enhanced helical CT in the days after apparent complete surgical excision of JNA with initial expansion in the skull base. Four independent, blinded readers evaluated the occurrence of RD. Final diagnoses were rendered on the basis of histologic examination of excised specimens of RD or clinical and radiologic follow-up. The Cohen kappa test was performed to examine interreader agreement. RESULTS: Postoperative contrast-enhanced CT had a sensitivity of 75%, a specificity of 83%, a positive predictive value of 75% and a negative predictive value of 83% for the detection of RD. The prevalence of RD was 40%. The base of pterygoids was the most frequent location of RD. Interreader agreement was high for the detection of putative RD (kappa=0.83). Variabilities in readers' interpretations were encountered for false-positive results and for disease in the foramen lacerum. False-negative results involved the base of pterygoids. Early postoperative CT scanning was well tolerated by all patients. CONCLUSION: Contrast-enhanced helical CT is an accurate tool to evaluate excision of JNA in the days after surgery. PMID- 15661705 TI - Poststenotic flow and intracranial hemodynamics in patients with carotid stenosis: transoral carotid ultrasonography study. AB - BACKGROUND AND PURPOSE: In severe carotid stenosis, blood supply via the poststenotic internal carotid artery (ICA) and collateral pathways determine cerebral perfusion. We investigated whether poststenotic flow on transoral carotid ultrasonography (TOCU) is predictive of cerebral hemodynamics. METHODS: Eighty-eight patients with unilateral carotid stenosis underwent TOCU to analyze blood flow velocity and poststenotic diameter of the extracranial ICA. Intracranial collaterals and cerebral hemodynamics were assessed with selective angiography and single photon emission CT. RESULTS: Poststenotic diameter (P <.0001) and velocities (peak systolic velocity [PSV], time-averaged mean velocity [TMV], end-diastolic velocity [EDV]; all P < or = .009) decreased with stenotic severity. Poststenotic diameter was correlated with PSV (r=0.36, P=.0005), EDV (r=0.32, P=.002), and TMV (r=0.39, P=.0001). Poststenotic flow was significantly lower with collateral pathways than without pathways (P < or = .02) and significantly decreased as the number of the collaterals increased (P <.0001). Flow <5 mL/s indicated collaterals (81% sensitivity, 80% specificity). When flow was <5 mL/s, the asymmetry index in the middle cerebral artery (MCA) territory was significantly low during rest (P=.003) and after acetazolamide challenge (P=.006). Poststenotic flow velocity was associated with baseline (P=.007) and postacetazolamide (P=.0009) MCA asymmetry indexes. CONCLUSION: Poststenotic ICA flow measured with TOCU reflects collateral flow and cerebral hemodynamics in patients with severe carotid stenosis. This technique may provide new parameters for screening patients with hemodynamically significant carotid stenosis. PMID- 15661707 TI - Evaluation of platybasia with MR imaging. AB - BACKGROUND AND PURPOSE: Platybasia, or abnormal obtuseness of the basal angle, was first measured on plain skull images. At present, evaluation of the brain and skull more commonly involves CT and MR imaging. We evaluated a new MR imaging method of evaluating platybasia. METHODS: We retrospectively evaluated midline sagittal MR images in 200 adults and 50 children. The basal angle of the skull base was measured by using two methods: The standard MR imaging technique measured the angle formed by two lines-one joining the nasion and the center of the pituitary fossa connected by a line joining the anterior border of the foramen magnum and center of the pituitary fossa. The modified technique measured the angle formed by a line across the anterior cranial fossa and dorsum sellae connecting a line along the clivus. RESULTS: With the standard MR imaging technique, we obtained mean angles of 129 degrees +/- 6 degrees for adults and 127 degrees +/- 5 degrees for children, compared with 135.3 degrees (composite mean) in previous series. The modified technique produced values of 117 degrees +/- 6 degrees for adults and 114 degrees +/- 5 degrees for children, which were significantly lower that those of standard MR imaging and traditional radiography (P <.05). CONCLUSION: Both the standard and modified MR imaging techniques produced basal angles lower than those previously reported with standard radiography. The modified technique uses clearly featured landmarks that can be reproduced consistently on midline sagittal T1 images. This technique and its corresponding values can be used as the new standard for evaluating the basal angle. PMID- 15661708 TI - Aortic arch variation: a unique case with anomalous origin of both vertebral arteries as additional branches of the aortic arch distal to left subclavian artery. AB - We present the case of bilateral anomalous origin of both vertebral arteries (VAs) in a 20-year-old male patient who presented for routine contrast-enhanced CT follow-up examination of the chest. Contrast-enhanced CT revealed abnormal origins of both the VAs from the aortic arch distal to the origin of the left subclavian artery. Following this, CT angiography was performed, which confirmed the findings. To our knowledge, this is the first report of anomalous origins of both VAs beyond the origin of the left subclavian artery. The possible embryonic mechanism and the clinical importance of this variant is also reviewed. PMID- 15661709 TI - Parapharyngeal neuroglial heterotopia presenting as a growing single locular cyst: MR imaging findings. AB - Heterotopic brain presenting as a giant, growing, single locular cyst at the parapharyngeal space has not been reported before, to our knowledge. We present such a case, with MR imaging findings, in a 13-month-old girl. A well-demarcated giant cystic mass was noted in the left parapharyngeal space from the skull base to the submandibular region. Airway compression and deformity of the left mandible with subluxation of the temporomandibular joint were noted. The cyst contained a clear fluid that was isointense to CSF with all pulse sequences. Wall enhancement was noted on contrast-enhanced T1-weighted images. No connection to intracranial structures was noted. Histologic findings were compatible with neuroglial heterotopia. PMID- 15661710 TI - MR imaging findings of uveal leiomyoma: three cases. AB - Leiomyoma is a rare tumor arising from the uveal tract. Fundoscopically, it appears as a yellowish-white, elevated mass and cannot be readily distinguished from melanoma or other uveal tumors. Cross-sectional imaging may have an important role, particularly when the opaque ocular media or a vitreous hemorrhage precludes a clear ophthalmoscopic view. In this respect, radiologists should be aware of suggestive findings of uveal leiomyoma to avoid an incorrect diagnosis and unnecessary radical surgery. We report MR imaging findings of three cases of uveal leiomyoma. PMID- 15661711 TI - Accuracy of dynamic perfusion CT with deconvolution in detecting acute hemispheric stroke. AB - BACKGROUND AND PURPOSE: Dynamic perfusion CT (PCT) with deconvolution produces maps of time-to-peak (TTP), mean transit time (MTT), regional cerebral blood flow (rCBF), and regional cerebral blood volume (rCBV), with a computerized automated map of the infarct and penumbra. We determined the accuracy of these maps in patients with suspected acute hemispheric stroke. METHODS: Forty-six patients underwent nonenhanced CT and dynamic PCT, with follow-up CT or MR imaging. Two observers reviewed the nonenhanced studies for signs of stroke and read the PCT maps for TTP, MTT, rCBF, and rCBV abnormalities. Sensitivity, specificity, accuracy, and interobserver agreement were compared (Wilcoxon tests). Nonenhanced CT and PCT data were reviewed for stroke extent according to previously reported methods. Sensitivity, specificity, and accuracy of the computerized maps in detecting ischemia and its extent were determined. RESULTS: Compared with nonenhanced CT, PCT maps were significantly more accurate in detecting stroke (75.7-86.0% vs. 66.2%; P <.01), MTT maps were significantly more sensitive (77.6% vs. 69.2%; P <.01), and rCBF and rCBV maps were significantly more specific (90.9% and 92.7%, respectively, vs. 65.0%; P <.01). Regarding stroke extent, PCT maps were significantly more sensitive than nonenhanced CT (up to 94.4% vs. 42.9%; P <.01) and had higher interobserver agreement (up to 0.763). For the computerized map, sensitivity, specificity, and accuracy, respectively, were 68.2%, 92.3%, and 88.1% in detecting ischemia and 72.2%, 91.8%, and 87.9% in showing the extent. CONCLUSION: Dynamic PCT maps are more accurate than nonenhanced CT in detecting hemispheric strokes. Despite limited spatial coverage, PCT is highly reliable to assess the stroke extent. PMID- 15661712 TI - Relationship between transcranial Doppler and CT data in acute intracerebral hemorrhage. AB - BACKGROUND AND PURPOSE: It would be useful to have a noninvasive test for correlation with CT findings in patients with intracerebral hemorrhage (ICH). We determined which transcranial Doppler (TCD) variables are related to which CT data in patients with ICH. METHODS: We prospectively included 51 patients (age +/ SD, 66.2 +/- 12.4 years; 30 men, 21 women) with first-ever supratentorial, nontraumatic ICH. CT and TCD examination were performed in the acute stage (less than 12 hours from symptom onset). TCD recordings were obtained from the middle cerebral arteries, and the following variables were analyzed: systolic (Vs), diastolic (Vd), mean (Vm) velocities, and pulsatiliy index from the affected (a) and unaffected (u) hemispheres. RESULTS: PIs obtained for both hemispheres were positively correlated with hematoma volume (aPI, r=0.43, P=.001; uPI, r=0.44, P=.001), volume of hypoattenuation (aPI r=0.64, P <.0001; uPI, r=0.39, P=.005), total volume (aPI, r=0.59, P <.0001; uPI, r=0.48, P <.0001), and midline shift (aPI, r=0.28, P=.04; uPI, r=0.29, P=.03). Both PIs were increased in patients with intraventricular hemorrhage (aPI, P=.01; uPI P=.004). No TCD parameter was correlated with ventricular size. CONCLUSION: Most TCD parameters were correlated with CT data in the acute stage of ICH. An increase in PI probably reflects intracranial hypertension and mass effect. Further studies are needed to determine the clinical application of our findings. PMID- 15661713 TI - Local tissue damage assessed with statistical mapping analysis of brain magnetization transfer ratio: relationship with functional status of patients in the earliest stage of multiple sclerosis. AB - BACKGROUND AND PURPOSE: In the early stage of Multiple Sclerosis (MS), conventional MR imaging parameters such as T2 lesion load fail to explain the clinical status of patients. In the present work, we aimed to determine the ability of magnification transfer imaging to better reflect the relationship between local tissue damage and functional status of MS patients. METHODS: We performed a comparative statistical mapping analysis on brain tissue magnetization transfer ratio (MTR) data measured in 18 patients with clinically isolated syndrome suggestive of MS (CISSMS) and 18 matched control subjects. RESULTS: In the patients with CISSMS, a pattern of significant low MTR values was observed in the white matter, corpus callosum, bilateral occipitofrontal fascicles, right fornix, right parietal white matter, external capsule, right superior longitudinal fasciculus (SLF), right inferior longitudinal fasciculus, optica radiata, parietal white matter, right cingulum, gray matter, bilateral thalamus, bilateral caudate, right insula, and left Brodmann area (BA) 8. No correlation was found between local MTR decrease and Expanded Disability Status Scale score. Significant correlations between MTR and MS Functional Composite scores (Spearman rank test, P <.05) were observed in the left BA40, right SLF, right frontal white matter, splenium, and anterior corpus callosum. Local MTR values correlated with Paced Auditory Serial Addition Test scores in the left BA40, right BA4, right SLF, and splenium. CONCLUSION: Statistical mapping analysis of brain MTR data provides valuable information on the relationship between the location of brain tissue damage and its functional impact in patients with MS, even in the earliest stage of the disease. PMID- 15661714 TI - Quantitative proton MR spectroscopic findings of cortical reorganization in the auditory cortex of musicians. AB - BACKGROUND AND PURPOSE: Brain has a capacity for reorganization that enables use dependent adaptations to acquire skills. Previous studies demonstrated morphometric and functional use-dependent changes in the brains of musicians. The purpose of this study was to investigate the differences in metabolite concentrations in the planum temporale, an area strongly associated with the processing of music perception, between trained musicians and non-musicians. We hypothesized that the microscopic changes leading to use-dependent adaptations in brain might cause neurometabolite changes that could be detected with quantitative proton MR spectroscopy. METHODS: We performed quantitative proton MR spectroscopy in the left planum temporale of 10 musicians (six men and four women; age range, 20-37 years) and in those of 10 age- and sex-matched control subjects who had no musical training. We calculated the major metabolite concentrations in the left planum temporale. RESULTS: The difference in N acetylaspartate (NAA) concentrations between the musicians and the non-musician control subjects was statistically significant (P <.01). No significant difference was noted in the choline and creatine concentrations between the musicians and the non-musician control subjects (P >.05). The NAA concentration of the musicians correlated with the total duration of musical training and activity (r=0.733, P <.05). CONCLUSION: Long-term, professional musical activity caused significant changes in the neurometabolite concentrations that might reflect the physiologic mechanism(s) of use-dependent adaptation in the brains of musicians. PMID- 15661715 TI - Automatic method to assess local CT-MR imaging registration accuracy on images of the head. AB - BACKGROUND AND PURPOSE: Precise registration of CT and MR images is crucial in many clinical cases for proper diagnosis, decision making or navigation in surgical interventions. Various algorithms can be used to register CT and MR datasets, but prior to clinical use the result must be validated. To evaluate the registration result by visual inspection is tiring and time-consuming. We propose a new automatic registration assessment method, which provides the user a color coded fused representation of the CT and MR images, and indicates the location and extent of poor registration accuracy. METHODS: The method for local assessment of CT-MR registration is based on segmentation of bone structures in the CT and MR images, followed by a voxel correspondence analysis. The result is represented as a color-coded overlay. The algorithm was tested on simulated and real datasets with different levels of noise and intensity non-uniformity. RESULTS: Based on tests on simulated MR imaging data, it was found that the algorithm was robust for noise levels up to 7% and intensity non-uniformities up to 20% of the full intensity scale. Due to the inability to distinguish clearly between bone and cerebro-spinal fluids in the MR image (T1-weighted), the algorithm was found to be optimistic in the sense that a number of voxels are classified as well-registered although they should not. However, nearly all voxels classified as misregistered are correctly classified. CONCLUSION: The proposed algorithm offers a new way to automatically assess the CT-MR image registration accuracy locally in all the areas of the volume that contain bone and to represent the result with a user-friendly, intuitive color-coded overlay on the fused dataset. PMID- 15661716 TI - evaluation of communication between intracranial arachnoid cysts and cisterns with phase-contrast cine MR imaging. AB - BACKGROUND AND PURPOSE: The demonstration of communication between arachnoid cysts (ACs) and the adjacent subarachnoid space is a prerequisite for their proper management. CT cisternography (CTC) is the conventional method for functional evaluation of ACs. The sensitivity of MR imaging to CSF flow has been demonstrated, but reports of the clinical usefulness of MR CSF flow techniques in this application are limited. The purpose of our study was to prospectively evaluate the accuracy of MR CSF flow study as an alternative to CTC in this setting. METHODS: MR CSF flow study with retrospective ECG-gated 2D, fast low angle shot, phase-contrast (PC), cine gradient-echo sequence was performed in 39 patients with an intracranial AC. Results were compared with intraoperative and CTC findings. RESULTS: PC cine MR imaging results were compatible with operative or CTC findings in 36 (92.3%) of 39 patients. Twenty-four cysts were noncommunicating, and 15 were communicating. Three cysts were evaluated as being noncommunicating on PC cine MR imaging (false-negative) but demonstrated contrast enhancement on CTC. No false-positive diagnoses occurred. All cysts regarded as being communicating on PC cine MR imaging were also found to be communicating on both confirmation methods. CONCLUSION: MR CSF flow imaging with a PC cine sequence can be incorporated in the imaging work-up of ACs. This is a reliable alternative to invasive CTC for the functional evaluation of ACs. PMID- 15661718 TI - De novo development of a lesion with the appearance of a cavernous malformation adjacent to an existing developmental venous anomaly. AB - A case is presented in which a complex multicystic hemosiderin-containing lesion developed adjacent to a previously documented developmental venous anomaly (venous angioma). This lesion had the characteristic MR imaging appearance of a cavernous malformation. Follow-up MR imaging demonstrated a decrease in both the size and complexity of this lesion, which suggests at least a portion of the lesion was due to sequelae of hemorrhage. This case further supports the association of a de novo, hemosiderin-containing lesion in association with developmental venous anomaly. Implications of these findings are that the commonly seen "cavernous malformations" in association with developmental venous anomaly are acquired lesions, and not congenital in origin. A review of the literature discussing the etiology of cavernous malformations and their reported association with the developmental venous anomaly is provided. PMID- 15661717 TI - Water excitation: a possible pitfall in cerebral time-of-flight angiography. AB - BACKGROUND AND PURPOSE: Time-of-flight MR angiography (TOF MRA), with its advantage of high spatial resolution, is widely used for visualization of intracranial arteries. Fat signal intensity from bone marrow can interfere with vessel signal intensity, especially in maximum intensity projection reconstructions. Use of a technique such as fat saturation or water excitation can reduce this signal intensity. METHODS: Ten volunteers were included in this study. TOF MRA was performed by using spoiled gradient echo sequences on a 1.5-T MR unit either with or without water excitation. Water excitation was performed by using a binomial excitation pulse pair, with a null in the excitation profile at the fat frequency to obtain fat suppression. Two blinded neuroradiologists then judged the images. Additional studies by using a phantom with a flow of about 2 mL/s were performed under the same conditions. Image quality obtained with and that without water excitation was graded as similar by both neuroradiologists. RESULTS: As the main finding, the sequences by using water excitation revealed an important pitfall: apparent carotid artery stenosis was detected in 4/10 and occlusions in 1/10 cases. Use of a flow phantom could reveal the same pitfall. Intracranial vessel disease was excluded in all volunteers by using Doppler sonography. CONCLUSION: The type of water excitation tested here can induce significant artifacts in cerebral TOF MRA. These artifacts can be misinterpreted as carotid artery stenosis or even occlusion, whereas the benefit brought by water excitation with respect to fat suppression is less significant. PMID- 15661719 TI - Secondary supratentorial primitive neuroectodermal tumor following irradiation in a patient with low-grade astrocytoma. AB - We report a case of a supratentorial primitive neuroectodermal tumor (PNET) that occurred 12 years after cranial irradiation for a grade II astrocytoma. Neuroimaging was unable to distinguish between a recurrence of the original neoplasm and the development of a new, distinct entity. Pathologic review assisted by immunohistochemical staining, however, revealed a high-grade PNET. Although rare, PNET needs to be included in the differential diagnoses for previously irradiated patients, who develop recurrent brain tumors in the presence of uncharacteristic imaging features. PMID- 15661720 TI - 3D rotational angiography: use of propeller rotation for the evaluation of intracranial aneurysms. AB - We compared two methods of 3D digital subtraction angiography (DSA)--propeller and standard rotation--for the assessment of aneurysmal morphology and its relation to neighboring vessels. Aneurysms were correctly visualized and localized with both techniques. 3D DSA with propeller rotation technique seems to be effective and allows us to reduce the amount of contrast material related to a shortened acquisition time. Technical progress including propeller rotation allows a larger range of rotation and faster rotational speeds. PMID- 15661721 TI - Preliminary results of endovascular stent-assisted angioplasty for symptomatic middle cerebral artery stenosis. AB - BACKGROUND AND PURPOSE: Stent placement for intracranial atherosclerosis has become an alternative treatment technique; however, stent placement for middle cerebral artery (MCA) stenosis remains a technical and clinical challenge. Our purpose was to assess the feasibility, safety, and effectiveness of stent placement for MCA stenosis. METHODS: Between May 1998 and August 2003, we performed stent placement for MCA stenosis (more than 50%) in 17 patients and retrospectively analyzed the technical success rate, complications, and outcomes over 10 months. RESULTS: Stent placement was technically successful in 16 (94.1%) patients and failed in one patient (5.9%), because the stent did not reach the lesion due to carotid artery tortuousity. In 16 patients, postprocedural angiography showed restoration of the normal luminal diameter. Acute in-stent thromboses occurred in nine patients (56.3%) and was lysed with abciximab. The parent artery ruptured in two patients (12.5%): One died from massive subarachnoid hemorrhage, and the other patient received a second stent over the tear site. Stent jumping (distal migration at the time of balloon inflation) occurred in one patient (6.3%) and was solved by implanting a second stent. Periprocedural complications included subacute in-stent thrombosis (n = 1, 6.3%) and minor stroke (n=1, 6.3%); these were relieved with heparin therapy or local thrombolysis. Fifteen patients experienced symptomatic relief or were stable at follow-up. Angiographic follow-up (n=6) revealed no in-stent restenosis. CONCLUSION: Stent placement for symptomatic MCA stenosis was technically feasible and effective in alleviating symptoms and improving cerebral blood flow. PMID- 15661722 TI - Predictive value of angiographic testing for tolerance to therapeutic occlusion of the carotid artery. AB - BACKGROUND AND PURPOSE: Controversy exists on how to assess a patient's tolerance before permanent carotid artery occlusion. We sought to determine the positive predictive value of synchronous opacification of hemispheric cortical veins at angiography of the contralateral carotid or vertebral artery as a predictor of tolerance to permanent carotid artery occlusion without development of ischemic injury. METHODS: Seventy-six angiographic test occlusions were performed in 74 consecutive patients considered for therapeutic occlusion. Angiography of collateral cerebral vessels was performed during test occlusion. Synchronous filling (a < 0.5-second delay of opacification between the cortical veins of the occluded and collateral vascular territories) was considered a predictor for tolerance to permanent occlusion. To detect clinically silent ischemic defects, MR imaging was performed before and 6-12 weeks after permanent occlusion. Positive predictive value (95% confidence interval [CI]) of synchronous venous filling for absence of ischemic deficits after permanent occlusion was calculated. RESULTS: No procedural complications of the test occlusion occurred. In 51 of 54 patients who passed the test, permanent occlusion was performed. Two patients, both in poor clinical condition after subarachnoid hemorrhage, died of diffuse vasospasm after permanent occlusion. Of the 49 surviving patients, one developed a transient discrete hemiparesis with small new hypoperfusion infarctions on MR images. All other patients remained neurologically unchanged with no new ischemic lesions on follow-up MR images. Positive predictive value of tolerance to carotid artery occlusion after passing the angiographic test was 98% (95% CI: 89-100%). CONCLUSION: The angiographic test occlusion protocol reliably predicts tolerance to therapeutic carotid artery occlusion. It is safe and easy to perform. PMID- 15661723 TI - Hemodynamic evaluation during balloon test occlusion of the sigmoid sinus: clinical and technical considerations. AB - Test occlusion of a patent dural sinus requires careful hemodynamic evaluation. We performed balloon test occlusion of the sigmoid sinus in a patient with an advanced-stage temporal bone malignancy that affected the dominant sigmoid sinus. Before and during the test occlusion, intrasinus pressure was monitored upstream from the balloon via the inner lumen of a double-lumen angioplasty balloon microcatheter. On the basis of the hemodynamic findings, permanent balloon occlusion was carried out without complications. PMID- 15661724 TI - Fatal cerebral hemorrhage early after subclavian artery endovascular therapy. AB - We report a case of cerebral hyperperfusion syndrome complicated by a fatal intracranial hemorrhage as a consequence of endovascular recanalization of an occluded left subclavian artery. This entity is thought to be secondary to the failure of the normal autoregulatory mechanism controlling cerebral blood flow after recanalization. Patients at risk for this relatively rare complication are not easily identified. Further investigation is warranted to identify patients at risk for this complication and to determine optimal periprocedural management to decrease risk. PMID- 15661726 TI - MR imaging findings of cortical blindness following cerebral angiography: is this entity related to posterior reversible leukoencephalopathy? PMID- 15661727 TI - An explanation for putaminal CT, MR, and diffusion abnormalities secondary to nonketotic hyperglycemia. PMID- 15661728 TI - Absence of memory dysfunction after bilateral mammillary body and mammillothalamic tract electrode implantation: preliminary experience in three patients. PMID- 15661729 TI - Desperate appliance. PMID- 15661731 TI - Ultrasonography of carotidynia. PMID- 15661730 TI - Rupture of a cerebral aneurysm during MR Imaging: leakage of gadolinium into the subarachnoid space. PMID- 15661732 TI - Lipid mixing between lipoplexes and plasma lipoproteins is a major barrier for intravenous transfection mediated by cationic lipids. AB - It has been previously shown that transfection activity of cationic liposome/DNA lipoplexes delivered systemically is drastically inhibited by lipoproteins (Tandia, B. M., Vandenbranden, M., Wattiez, R., Lakhdar, Z., Ruysschaert, J. M., and Elouahabi, A. (2003) Mol Ther. 8, 264-273). In this work, we have compared the binding/uptake and transfection activities of DOTAP (N-[1-(2,3 dioleoyloxy)propyl]-N,N,N-trimethylammonium chloride) and diC14-amidine (3 tetradecylamino-N-tert-butyl-N'-tetra-decylpropionamidine)-containing lipoplexes in the presence or absence of purified low density lipoproteins and high density lipoprotein. Binding/uptake of both lipoplexes by the mouse lung endothelial cell line was inhibited to a similar extent in the presence of lipoproteins. In contrast, transfection activity of diC14-amidine-containing lipoplexes was almost completely inhibited (approximately by 95%), whereas approximately 40% transfection activity of DOTAP-containing lipoplexes was preserved in the presence of lipoproteins. Interestingly, the ability of lipoproteins to inhibit the transfection efficiency of lipoplexes was well correlated with their ability to undergo lipid mixing with the cationic lipid bilayer as revealed by fluorescence resonance energy transfer assay. Incubation of lipoplexes with increased doses of lipoproteins resulted in enhanced lipid mixing and reduced transfection activity of the lipoplexes in mouse lung endothelial cells. The role of lipid mixing in transfection was further demonstrated using lipid-mixing inhibitor, lyso-phosphatidylcholine, or activator (dioleoylphosphatidylethanolamine). Incorporation of Lyso-PC into diC14-amidine containing lipoplexes completely abolished their capacity to undergo lipid mixing with lipoproteins and allowed them to reach a high transfection efficiency in the presence of lipoproteins. On the other hand, the incorporation of dioleoylphosphatidylethanolamine into DOTAP/DNA lipoplex activated lipid mixing with the lipoproteins and was shown to be detrimental toward the transfection activity of these lipoplexes. Taken together, these results indicate that fusion of lipoplexes with lipoproteins is a limiting factor for in vivo transfection. PMID- 15661733 TI - Lysophospholipid flipping across the Escherichia coli inner membrane catalyzed by a transporter (LplT) belonging to the major facilitator superfamily. AB - The transfer of phospholipids across membrane bilayers is protein-mediated, and most of the established transporters catalyze the energy-dependent efflux of phospholipids from cells. This work identifies and characterizes a lysophospholipid transporter gene (lplT, formally ygeD) in Escherichia coli that is an integral component in the 2-acylglycerophosphoethanolamine (2-acyl-GPE) metabolic cycle for membrane protein acylation. The lplT gene is adjacent to and in the same operon as the aas gene, which encodes the bifunctional enzyme 2-acyl GPE acyltransferase/acyl-acyl carrier protein synthetase. In some bacteria, acyltransferase/acyl-ACP synthetase (Aas) and LplT homologues are fused in a single polypeptide chain. 2-Acyl-GPE transport to the inside of the cell was assessed by measuring the Aas-dependent formation of phosphatidylethanolamine. The Aas-dependent incorporation of [3H]palmitate into phosphatidylethanolamine was significantly diminished in deltalplT mutants, and the LplT-Aas transport/acylation activity was independent of the proton motive force. The deltalplT mutants accumulated acyl-GPE in vivo and had a diminished capacity to transport exogenous 2-acylglycerophosphocholine into the cell. Spheroplasts prepared from wild-type E. coli transported and acylated fluorescent 2-acyl-GPE with an apparent K(d) of 7.5 microM, whereas this high-affinity process was absent in deltalplT mutants. Thus, LplT catalyzes the transbilayer movement of lysophospholipids and is the first example of a phospholipid flippase that belongs to the major facilitator superfamily. PMID- 15661734 TI - Resistance to a protein farnesyltransferase inhibitor in Plasmodium falciparum. AB - The post-translational farnesylation of proteins serves to anchor a subset of intracellular proteins to membranes in eukaryotic organisms and also promotes protein-protein interactions. Inhibition of protein farnesyltransferase (PFT) is lethal to the pathogenic protozoa Plasmodium falciparum. Parasites were isolated that were resistant to BMS-388891, a tetrahydroquinoline (THQ) PFT inhibitor. Resistance was associated with a 12-fold decrease in drug susceptibility. Genotypic analysis revealed a single point mutation in the beta subunit in resistant parasites. The resultant tyrosine 837 to cysteine alteration in the beta subunit corresponded to the binding site for the THQ and peptide substrate. Biochemical analysis of Y837C-PFT demonstrated a 13-fold increase in BMS-388891 concentration necessary for inhibiting 50% of the enzyme activity. These data are consistent with PFT as the target of BMS-388891 in P. falciparum and suggest that PFT inhibitors should be combined with other antimalarial agents for effective therapy. PMID- 15661735 TI - Cytoprotective peptide humanin binds and inhibits proapoptotic Bcl-2/Bax family protein BimEL. AB - Humanin (HN) is a recently identified endogenous peptide that protects cells against cytotoxicity induced by various stimuli. Recently, we showed that HN binds to and inhibits Bax, a proapoptotic Bcl-2 family protein, suggesting a mechanism for HN action. In this study, we identified Bim, a Bcl-2 homology 3 only member of the Bcl-2/Bax family, as an additional HN target protein. Using in vitro protein binding, immunoprecipitation, and coimmunolocalization assays, we demonstrated that HN binds directly to the extra long isoform of Bim (BimEL) but not the long (BimL) or short (BimS) isoforms. HN also protects cells against apoptosis induced by BimEL but not BimL and BimS in gene transfection studies. In contrast, mutants of HN which failed to bind BimEL failed to protect from BimEL induced cell death. Moreover, HN inhibited BimEL-induced release of SMAC and cytochrome c from mitochondria isolated from bax-/-cells, indicating that HN can suppress BimEL independently of its effect on Bax. Finally, we demonstrate that HN prevents BimEL-induced oligomerization of Bak using isolated mitochondria. Taken together, our results indicate that the inhibition of BimEL may contribute to the antiapoptotic properties of the HN peptide. PMID- 15661736 TI - Overexpression of proteasome beta5 assembled subunit increases the amount of proteasome and confers ameliorated response to oxidative stress and higher survival rates. AB - The proteasome is the major cellular proteolytic machinery responsible for the degradation of both normal and damaged proteins. Proteasomes play a fundamental role in retaining cellular homeostasis. Alterations of proteasome function have been recorded in various biological phenomena including aging. We have recently shown that the decrease in proteasome activity in senescent human fibroblasts relates to the down-regulation of beta-type subunits. In this study we have followed our preliminary observation by developing and further characterizing a number of different human cell lines overexpressing the beta5 subunit. Stable overexpression of the beta5 subunit in WI38/T and HL60 cells resulted in elevated levels of other beta-type subunits and increased levels of all three proteasome activities. Immunoprecipitation experiments have shown increased levels of assembled proteasomes in stable clones. Analysis by gel filtration has revealed that the recorded higher level of proteasome assembly is directly linked to the efficient integration of "free" (not integrated) alpha-type subunits identified to accumulate in vector-transfected cells. In support we have also found low proteasome maturation protein levels in beta5 transfectants, thus revealing an increased rate/level of proteasome assembly in these cells as opposed to vector transfected cells. Functional studies have shown that beta5-overexpressing cell lines confer enhanced survival following treatment with various oxidants. Moreover, we demonstrate that this increased rate of survival is due to higher degradation rates following oxidative stress. Finally, because oxidation is considered to be a major factor that contributes to aging and senescence, we have overexpressed the beta5 subunit in primary IMR90 human fibroblasts and observed a delay of senescence by 4-5 population doublings. In summary, these data demonstrate the phenotypic effects following genetic up-regulation of the proteasome and provide insights toward a better understanding of proteasome regulation. PMID- 15661737 TI - Humanin binds and nullifies Bid activity by blocking its activation of Bax and Bak. AB - Recently, we discovered that Humanin (HN), a small endogenous peptide of 24 amino acids, binds to and inhibits the proapoptotic protein Bax. We show here that HN also interacts with the BH3-only Bcl-2/Bax family protein, Bid, as well as a truncated form of Bid (tBid) associated with protease-mediated activation of this proapoptotic protein. Synthetic HN peptide binds purified Bid and tBid in vitro and blocks tBid-induced release of cytochrome c and SMAC from isolated mitochondria, whereas mutant peptides that fail to bind Bid or tBid lack this activity. Moreover, HN peptide also retained protective activity on bax-/ mitochondria, indicating that HN can block tBid-induced release of apoptogenic proteins from these organelles in a Bax-independent manner. HN peptide inhibits tBid-induced oligomerization of Bax and Bak in mitochondrial membranes, as shown by experiments with chemical cross-linkers or gel filtration. Gene transfection experiments showed that HN (but not an inactive mutant of HN) also protects intact cells from apoptosis induced by overexpression of tBid. We conclude that Bid represents an additional cellular target of HN, and we propose that HN mediated suppression of Bid contributes to the antiapoptotic activity of this endogenous peptide. PMID- 15661738 TI - The human TREX2 3' -> 5'-exonuclease structure suggests a mechanism for efficient nonprocessive DNA catalysis. AB - The 3' --> 5'-exonucleases process DNA ends in many DNA repair pathways of human cells. Determination of the human TREX2 structure is the first of a dimeric 3' deoxyribonuclease and indicates how this highly efficient nonprocessive enzyme removes nucleotides at DNA 3' termini. Symmetry in the TREX2 dimer positions the active sites at opposite outer edges providing open access for the DNA. Adjacent to each active site is a flexible region containing three arginines positioned appropriately to bind DNA and to control its entry into the active site. Mutation of these three arginines to alanines reduces the DNA binding capacity by approximately 100-fold with no effect on catalysis. The human TREX2 catalytic residues overlay with the bacterial DnaQ family of 3'-exonucleases confirming the structural conservation of the catalytic sites despite limited sequence identity, and mutations of these residues decrease the still measurable activity by approximately 10(5)-fold, confirming their catalytic role. PMID- 15661739 TI - Repression and coactivation of CCAAT/enhancer-binding protein epsilon by sumoylation and protein inhibitor of activated STATx proteins. AB - CCAAT/enhancer-binding protein epsilon (C/EBPepsilon) is a neutrophil-specific transcription factor whose activity is controlled by juxtaposed activating and regulatory domains. We previously determined that the function of the major regulatory domain (RD1) in C/EBPepsilon was dependent on the integrity of a five amino acid motif that was identical to the recognition site for members of the small ubiquitin-like modifier (SUMO) family of ubiquitin-related proteins. We show here that the SUMO attachment site (the regulatory domain motif) is necessary and sufficient both for the intrinsic inhibitory function of RD1 and for coactivation by PIASxalpha and PIASxbeta, two members of the protein inhibitor of activated STAT (PIAS) family of SUMO E3 ligases. PIASxbeta was a more potent coactivator than PIASxalpha of both full-length C/EBPepsilon and fusion proteins containing the N-terminal portion of C/EBPepsilon, whereas PIASxalpha was more active on fusion proteins containing a heterologous activation domain. Two modes of coactivation were observed, one that was dependent on the integrity of the RING finger (RF) domain and was shared by both PIASxalpha and PIASxbeta and a second mode that was independent of the RF and was only observed with PIASxbeta. Sumoylation of C/EBPepsilon was enhanced by coexpression of PIASxalpha, suggesting that this modification is associated with the enhanced activity of the target protein. These results suggest that a complex interplay of accessory factors, including SUMO and PIAS proteins, modulates the activity of C/EBPepsilon. PMID- 15661740 TI - Interleukin-6 regulation of transforming growth factor (TGF)-beta receptor compartmentalization and turnover enhances TGF-beta1 signaling. AB - Transforming growth factor (TGF)-beta1 is a key cytokine involved in the pathogenesis of fibrosis in many organs, whereas interleukin (IL)-6 plays an important role in the regulation of inflammation. Recent reports demonstrate interaction between the two cytokines in disease states. We have assessed the effect of IL-6 on TGF-beta1 signaling and defined the mechanism by which this occurred. Stimulation of Smad-responsive promoter (SBE)4-Lux activity by TGF beta1 was significantly greater in the presence of IL-6 than that induced by TGF beta1 alone. Augmented TGF-beta1 signaling following the addition of IL-6 appeared to be mediated through binding to the cognate IL-6 receptor, the presence of which was confirmed by fluorescence-activated cell sorting and Stat specific signaling. TGF-beta1 receptors internalize by both caveolin-1 (Cav-1) lipid raft and early endosome antigen 1 (EEA-1) non-lipid raft pathways, with non lipid raft-associated internalization increasing TGF-beta1 signaling. Affinity labeling of TGF-beta1 receptors demonstrated that IL-6 stimulation resulted in increased partitioning of TGF-beta receptors to the non-lipid raft fraction. There was no change in expression of Cav-1; however, following IL-6 stimulation, co-immunoprecipitation demonstrated decreased association of IL-6 receptor with Cav-1. Increased TGF-beta1-dependent Smad signaling by IL-6 was significantly attenuated by inhibition of clathrin-mediated endocytosis and augmented by depletion of membrane cholesterol. These results indicate that IL-6 increased trafficking of TGF-beta1 receptors to non-lipid raft-associated pools results in augmented TGF-beta1 Smad signaling. PMID- 15661741 TI - Rap1 GTPase inhibits leukocyte transmigration by promoting endothelial barrier function. AB - The passage of leukocytes out of the blood circulation and into tissues is necessary for the normal inflammatory response, but it also occurs inappropriately in many pathological situations. This process is limited by the barrier presented by the junctions between adjacent endothelial cells that line blood vessels. Here we show that activation of the Rap1 GTPase in endothelial cells accelerated de novo assembly of endothelial cell-cell junctions and increased the barrier function of endothelial monolayers. In contrast, depressing Rap1 activity by expressing Rap1GAP led to disassembly of these junctions and increased their permeability. We also demonstrate that endogenous Rap1 was rapidly activated at early stages of junctional assembly, confirming the involvement of Rap1 during junctional assembly. Intriguingly, elevating Rap1 activity selectively within endothelial cells decreased leukocyte transendothelial migration, whereas inhibiting Rap1 activity by expression of Rap1GAP increased leukocyte transendothelial migration, providing physiological relevance to our hypothesis that Rap1 augments barrier function of inter endothelial cell junctions. Furthermore, these results suggest that Rap1 may be a novel therapeutic target for clinical conditions in which an inappropriate inflammatory response leads to disease. PMID- 15661742 TI - Polo-like kinase 1 phosphorylates heat shock transcription factor 1 and mediates its nuclear translocation during heat stress. AB - Heat shock transcription factor 1 (HSF1) is activated by pathophysiologic stresses and activation leads to an increased cellular level of heat shock proteins (Hsp(s)). Although the activation of HSF1 occurs via multiple stress induced processes such as hyperphosphorylation, the exact cellular mechanism of HSF1 activation is still unclear. Here we show polo-like kinase 1 (PLK1) and HSF1 interact in vivo using the tandem affinity purification system. Although the interaction between HSF1 and PLK1 is increased by thermal stress, overexpression of PLK1 did not affect HSF1 trimerization or DNA binding activity. This interaction results in the phosphorylation of HSF1 on serine 419 by PLK1. Interestingly, mutation of serine 419 to alanine inhibited heat-stress induced HSF1 nuclear translocation. Our results suggest that the phosphorylation of HSF1 by PLK1 is an essential step for HSF1 nuclear translocation by heat stress. PMID- 15661743 TI - Role of protein phosphatase 2A in mGluR5-regulated MEK/ERK phosphorylation in neurons. AB - The regulation of protein phosphorylation requires coordinated interaction between protein kinases and protein phosphatases (PPs). Recent evidence has shown that the Galphaq-protein-coupled metabotropic glutamate receptor (mGluR) 5 up regulates phosphorylation of MAPK/ERK1/2. However, signaling mechanisms linking mGluR5 to ERK are poorly understood. In this study, roles of a major serine/threonine PP, PP2A, in this event were evaluated in cultured neurons. We found that the PP1/2A inhibitors okadaic acid and calyculin A mimicked the effect of the mGluR5 agonists (RS)-3,5-dihydroxyphenylglycine and (RS)-2-chloro-5 hydroxyphenylglycine in facilitating phosphorylation of ERK1/2 and its upstream kinase, MEK1/2, in a PP2A-dependent but not PP1-dependent manner. Co administration of either inhibitor with an mGluR5 agonist produced additive phosphorylation of ERK1/2. Enzymatic assays showed a basal level of phosphatase activity of PP2A under normal conditions, and activation of mGluR5 selectively inhibited PP2A, but not PP1, activity. In addition, a physical association of the cytoplasmic C terminus of mGluR5 with PP2A was observed, and ligand activation of mGluR5 reduced mGluR5-PP2A binding. Additional mechanistic studies revealed that mGluR5 activation increased tyrosine (Tyr307) phosphorylation of PP2A, which was dependent on activation of a p60c-Src family tyrosine kinase, but not the epidermal growth factor receptor tyrosine kinase and resulted in dissociation of PP2A from mGluR5 and reduced PP2A activity. Together, we have identified a novel, mGluR5-triggered signaling mechanism involving use- and Src-dependent inactivation of PP2A, which contributes to mGluR5 activation of MEK1/2 and ERK1/2. PMID- 15661744 TI - Brain contains a functional glucose-6-phosphatase complex capable of endogenous glucose production. AB - Glucose is absolutely essential for the survival and function of the brain. In our current understanding, there is no endogenous glucose production in the brain, and it is totally dependent upon blood glucose. This glucose is generated between meals by the hydrolysis of glucose-6-phosphate (Glc-6-P) in the liver and the kidney. Recently, we reported a ubiquitously expressed Glc-6-P hydrolase, glucose-6-phosphatase-beta (Glc-6-Pase-beta), that can couple with the Glc-6-P transporter to hydrolyze Glc-6-P to glucose in the terminal stages of glycogenolysis and gluconeogenesis. Here we show that astrocytes, the main reservoir of brain glycogen, express both the Glc-6-Pase-beta and Glc-6-P transporter activities and that these activities can couple to form an active Glc 6-Pase complex, suggesting that astrocytes may provide an endogenous source of brain glucose. PMID- 15661745 TI - Comparative agonist/antagonist responses in mutant human C5a receptors define the ligand binding site. AB - The C terminus is responsible for all of the agonist activity of C5a at human C5a receptors (C5aRs). In this report we have mapped the ligand binding site on the C5aR using a series of agonist and antagonist peptide mimics of the C terminus of C5a as well as receptors mutated at putative interaction sites (Ile(116), Arg(175,) Arg(206), Glu(199), Asp(282), and Val(286)). Agonist peptide 1 (Phe-Lys Pro-d-cyclohexylalanine-cyclohexylalanine-d-Arg) can be converted to an antagonist by substituting the bulkier Trp for cyclohexylalanine at position 5 (peptide 2). Conversely, mutation of C5aR transmembrane residue Ile(116) to the smaller Ala (I116A) makes the receptor respond to peptide 2 as an agonist (Gerber, B. O., Meng, E. C., Dotsch, V., Baranski, T. J., and Bourne, H. R. (2001) J. Biol. Chem. 276, 3394-3400). However, a potent cyclic hexapeptide antagonist, Phe-cyclo-[Orn-Pro-d-cyclohexylalanine-Trp-Arg] (peptide 3), derived from peptide 2 and which binds to the same receptor site, remains a full antagonist at I116AC5aR. This suggests that although the residue at position 5 might bind near to Ile(116), the latter is not essential for either activation or antagonism. Arg(206) and Arg(175) both appear to interact with the C-terminal carboxylate of C5a agonist peptides, suggesting a dynamic binding mechanism that may be a part of a receptor activation switch. Asp(282) has been previously shown to interact with the side chain of the C-terminal Arg residue, and Glu(199) may also interact with this side chain in both C5a and peptide mimics. Using these interactions to orient NMR-derived ligand structures in the binding site of C5aR, a new model of the interaction between peptide antagonists and the C5aR is presented. PMID- 15661746 TI - Insights into selective activation of p53 DNA binding by c-Abl. AB - As a transcription factor, p53 recognizes a specific consensus DNA sequence and activates the expression of the target genes involved in either growth arrest or apoptosis. Despite our wealth of knowledge on the genes that are targeted by p53 in growth arrest and apoptosis, relatively little is known about the promoter specificity triggered by p53 in these processes. Here we show that interaction with c-Abl stabilized p53 tetrameric conformation, and as a consequence c-Abl stimulated p53 DNA binding only when all quarter binding sites (a perfect binding sequence) on p53-responsive promoters were present. This result suggests that in response to DNA damage, c-Abl binding may specifically stimulate p53 DNA binding on the promoters with perfect binding sequences. A sequence comparison of several known p53-responsive elements illustrates the presence of the perfect binding sequences on the p21 but not the Bax promoter. Significantly, we show that c-Abl indeed enhanced p53 DNA binding and transcription from p21 but not Bax. These results suggest that the promoter specificity plays an important role in selective activation of p53 DNA binding by c-Abl. The implications of this with relation to selective activation of p53 target genes involved in either growth arrest or apoptosis are discussed. PMID- 15661748 TI - Evolutionary transition from resprouter to seeder life history in two Erica (Ericaceae) species: insights from seedling axillary buds. AB - BACKGROUND AND AIMS: The genus Erica represents the epitome of plant biodiversity in the South African Cape region. It includes seeder and resprouter species, but both species diversity and narrow endemism are tightly associated with the seeder habit. It also includes 'mixed' species, in which both seeder and resprouter life histories are found. This intraspecific variation in life history is genetically based. METHODS: The cotyledonary region and basal stem of seeder and resprouter seedlings of two 'mixed' species, Erica calycina and E. coccinea, were examined to detect morphological and anatomical differences in axillary bud development between regeneration forms. KEY RESULTS: While at least some bud activity was observed for resprouter seedlings, none was detected for seeder seedlings. A closer examination allowed the detection of some axillary buds in seeder seedlings of the two species, but they appeared in an unequivocally atrophied state. CONCLUSIONS: The seeder and resprouter life histories are two character states and the seeder one (i.e. loss of resprouting) is derived within these two Erica species. Results allow the hypothesis that the loss of resprouting in a fire-prone scenario such as the Cape fynbos has promoted high diversification rates in seeder Erica lineages. PMID- 15661747 TI - Low resolution structural study of two human HSP40 chaperones in solution. DJA1 from subfamily A and DJB4 from subfamily B have different quaternary structures. AB - Proteins that belong to the heat shock protein (Hsp) 40 family assist Hsp70 in many cellular functions and are important for maintaining cell viability. A knowledge of the structural and functional characteristics of the Hsp40 family is therefore essential for understanding the role of the Hsp70 chaperone system in cells. In this work, we used small angle x-ray scattering and analytical ultracentrifugation to study two representatives of human Hsp40, namely, DjA1 (Hdj2/dj2/HSDJ/Rdj1) from subfamily A and DjB4 (Hlj1/DnaJW) from subfamily B, and to determine their quaternary structure. We also constructed low resolution models for the structure of DjA1-(1-332), a C-terminal-deleted mutant of DjA1 in which dimer formation is prevented. Our results, together with the current structural information of the Hsp40 C-terminal and J-domains, were used to generate models of the internal structural organization of DjA1 and DjB4. The characteristics of these models indicated that DjA1 and DjB4 were both dimers, but with substantial differences in their quaternary structures: whereas DjA1 consisted of a compact dimer in which the N and C termini of the two monomers faced each other, DjB4 formed a dimer in which only the C termini of the two monomers were in contact. The two proteins also differed in their ability to bind unfolded luciferase. Overall, our results indicate that these representatives of subfamilies A and B of human Hsp40 have different quaternary structures and chaperone functions. PMID- 15661749 TI - Perfect syncarpy in apple (Malus x domestica 'Summerland McIntosh') and its implications for pollination, seed distribution and fruit production (Rosaceae: Maloideae). AB - BACKGROUND AND AIMS: The gynoecium of the domestic apple, Malus x domestica, has been assumed to be imperfectly syncarpic, whereby pollination of each stigmatic surface can result in fertilization within only one of the five carpels. Despite its implied effect on fruit quantity and quality, the resulting influence of flower form on seed set and distribution within the apple fruit has seldom been investigated. Instead, poor fruit quality is usually attributed to problems with pollination, such as low bee numbers and/or ineffective pollinators within apple agro-ecosystems. The objective of this study was to determine the true nature of gynoecial structure and its influence on fruit production in the apple cultivar 'Summerland McIntosh'. METHODS: A stigma-excision method was used to determine the effects of uneven pollination among the five stigmas on fruit quantity (as measured by fruit set), and quality (seed number and distribution). In addition, flowers were examined microscopically to determine pollen tube pathways. KEY RESULTS: Fruit set, seed number, seed distribution, and the microscopic examination of flower gynoecial structure reported in this study indicated that the gynoecium of the cultivar Summerland McIntosh is perfectly syncarpic and not imperfectly syncarpic as previously thought. CONCLUSIONS: Pollination levels among the five stigmas need not be uniform to obtain full seed development within Summerland McIntosh fruit; even if one stigmatic surface is adequately pollinated, a full complement of seeds is likely. The importance of perfect syncarpy in recognizing true causes of poor fruit quality in apple is discussed. PMID- 15661750 TI - Structure and function of plant canopies. AB - This section comprises a set of papers taken from those presented at a symposium held to commemorate the 50th anniversary of the Monsi-Saeki theory (1953), together with invited papers. The papers describe recent advances in the study of structure and function of plant canopies and are written by former students (and their collaborators) of Professors Monsi and Saeki. The topics cover construction and maintenance of efficient photosynthetic systems at leaf, individual plant and stand level. Canopy structure and function are analysed with respect to optimization and an evolutionarily stable strategy. A new translation of the original paper by Monsi and Saeki (1953) into English has been commissioned and is included in this section. PMID- 15661751 TI - On the factor light in plant communities and its importance for matter production. 1953. PMID- 15661752 TI - A letter from Professor Boysen Jensen to Professor Monsi, September 1955. PMID- 15661753 TI - Predisposition to atypical hemolytic uremic syndrome involves the concurrence of different susceptibility alleles in the regulators of complement activation gene cluster in 1q32. AB - The efficiency of the complement system as an innate immune defense mechanism depends on a fine control that restricts its action to pathogens and prevents non specific damage to host tissues. Genetic and functional analyses have shown that this critical control of complement activation may be impaired in atypical hemolytic uremic syndrome (aHUS) patients. Mutations in HF1, MCP or FI have been found in aHUS patients, but incomplete penetrance of the disease in individuals carrying these mutations is relatively frequent and no genetic defect has yet been found in a majority of aHUS patients. We report here the identification of a specific SNP haplotype block, spanning the MCP gene in the regulators of complement activation gene cluster, which is over-represented in aHUS patients and strongly associates with the severity of the disease. Linkage disequilibrium analyses suggest that this SNP haplotype also includes the CR1, DAF and C4BP genes. Initial studies identified two SNPs in the haplotype that influence the transcription activity of the MCP promoter in transient transfection experiments. Notably, the SNP haplotype block was found to be particularly frequent among patients who carry mutations in HF1, MCP or FI. These findings and the identification of aHUS patients carrying mutations in two complement regulatory genes provide an important insight into the etiology of aHUS. Together, they suggest that complement regulatory molecules act as a protein network and that multiple hits, involving plasma- and membrane-associated complement regulatory proteins, are necessary to impair protection to host tissues and to confer significant predisposition to aHUS. PMID- 15661754 TI - The Fanconi anemia pathway is required for the DNA replication stress response and for the regulation of common fragile site stability. AB - Fanconi anemia (FA) is a rare multi-genic, autosomal and X-linked recessive disorder characterized by hematological abnormalities, developmental defects and increased cancer susceptibility. Patient-derived FA cells display heightened sensitivity to DNA cross-linking agents such as mitomycin C (MMC). In response to DNA damaging agents, and during S-phase of the cell cycle, the FA pathway is activated via the mono-ubiquitination of FANCD2 (FANCD2-Ub), signaling its translocation to discrete nuclear foci, where it co-localizes with the central DNA repair proteins BRCA1 and RAD51. However, the exact function of activated FANCD2-Ub remains unclear. Here, we have characterized the role of the FA pathway in response to DNA replicative stress by aphidicolin (APH) and hydroxyurea (HU). The FA pathway is strongly activated in response to both agents. In addition, using patient-derived FA cell lines and siRNA targeting FANCD2, we demonstrate a functional requirement for the FA pathway in response to low doses of APH: a replicative stress treatment known to result in chromosome breakage at common fragile sites. Both the total number of chromosome gaps and breaks and breaks at the specific common fragile sites FRA3B and FRA16D were significantly elevated in the absence of an intact FA pathway. Furthermore, we demonstrate that APH activates the mono-ubiquitination of both FANCD2 and PCNA and the phosphorylation of RPA2, signaling processive DNA replication arrest. Following APH treatment, FANCD2-Ub co-localizes with PCNA (early) and RPA2 (late) in discrete nuclear foci. Our results demonstrate an integral role for the FA pathway in the DNA replication stress response. PMID- 15661755 TI - Neuronal dysfunction in a polyglutamine disease model occurs in the absence of ubiquitin-proteasome system impairment and inversely correlates with the degree of nuclear inclusion formation. AB - The accumulation of protein deposits in neurons, in vitro proteasome assays and over-expression studies suggest that impairment of the ubiquitin-proteasome system (UPS) may be a common mechanism of pathogenesis in polyglutamine diseases such as Huntington disease and spinocerebellar ataxias (SCAs). Using a knock-in mouse model that recapitulates the clinical features of human SCA7, including selective neuronal dysfunction, we assessed the UPS at cellular resolution using transgenic mice that express a green fluorescent protein (GFP)-based reporter substrate (Ub(G76V)-GFP) of the UPS. The levels of the reporter remained low during the initial phase of disease, suggesting that neuronal dysfunction occurs in the presence of a functional UPS. Late in disease, we observed a significant increase in reporter levels specific to the most vulnerable neurons. Surprisingly, the basis for the increase in Ub(G76V)-GFP protein can be explained by a corresponding increase in Ub(G76V)-GFP mRNA in the vulnerable neurons. An in vitro assay also showed normal proteasome proteolytic activity in the vulnerable neurons. Thus, no evidence for general UPS impairment or reduction of proteasome activity was seen. The differential increase of Ub(G76V)-GFP among individual neurons directly correlated with the down-regulation of a marker of selective pathology and neuronal dysfunction in SCA7. Furthermore, we observed a striking inverse correlation between the neuropathology revealed by this reporter and ataxin-7 nuclear inclusions in the vulnerable neurons. Altogether, these data show a protective role against neuronal dysfunction for polyglutamine nuclear inclusions and exclude significant impairment of the UPS as a necessary step for polyglutamine neuropathology. PMID- 15661756 TI - A mutation in NFkB interacting protein 1 results in cardiomyopathy and abnormal skin development in wa3 mice. AB - We have identified waved 3 (wa3), a novel recessive mutation that causes abnormalities of the heart and skin. The cardiac defect results in a severe and rapidly progressive dilated cardiomyopathy. We identified the gene mutated in these mice, which we call NFkB interacting protein1 (Nkip1), using positional cloning. Nkip1 is expressed in skin, heart and vascular endothelium and shares homology with a small family of proteins that play a role in the regulation of transcription factors. A C-terminal fragment of this protein was previously identified as the RelA associated inhibitor (RAI). We show that the full-length protein is larger than previously described, and we confirm that it interacts with NFkB in vivo. Expression analysis of genes known to be regulated by NFkB revealed that Intercellular adhesion molecule 1 (Icam1) expression is consistently elevated in mutant mice. This result suggests that wa3 mutant mice represent a potentially important model for the analysis of the role of inflammatory processes in heart disease. PMID- 15661757 TI - Localization and functional analysis of the LARGE family of glycosyltransferases: significance for muscular dystrophy. AB - The dystroglycanopathies are a novel group of human muscular dystrophies due to mutations in known or putative glycosyltransferase enzymes. They share the common pathological feature of a hypoglycosylated form of alpha-dystroglycan, diminishing its ability to bind extracellular matrix ligands. The LARGE glycosyltransferase is mutated in both the myodystrophy mouse and congenital muscular dystrophy type 1D (MDC1D). We have transfected various cell lines with a variety of LARGE expression constructs in order to characterize their subcellular localization and effect on alpha-dystroglycan glycosylation. Wild-type LARGE co localized with the Golgi marker GM130 and stimulated the production of highly glycosylated alpha-dystroglycan (hyperglycosylation). MDC1D mutants had no effect on alpha-dystroglycan glycosylation and failed to localize correctly, confirming their pathogenicity. The two predicted catalytic domains of LARGE contain three conserved DxD motifs. Systematically mutating each of these motifs to NNN resulted in the mislocalization of one construct, while all failed to have any effect on alpha-dystroglycan glycosylation. A construct lacking the transmembrane domain also failed to localize at the Golgi apparatus. These results indicate that LARGE needs to both physically interact with alpha-dystroglycan and function as a glycosyltransferase in order to stimulate alpha-dystroglycan hyperglycosylation. We have also cloned and overexpressed a homologue of LARGE, glycosyltransferase-like 1B (GYLTL1B). Like LARGE it localized to the Golgi apparatus and stimulated alpha-dystroglycan hyperglycosylation. These results suggest that GYLTL1B may be a candidate gene for muscular dystrophy and that its overexpression could compensate for the deficiency of both LARGE and other glycosyltransferases. PMID- 15661758 TI - Characterization of the nephrocystin/nephrocystin-4 complex and subcellular localization of nephrocystin-4 to primary cilia and centrosomes. AB - Nephrocystin and nephrocystin-4 are newly identified proteins involved in familial juvenile nephronophthisis, an autosomal recessive nephropathy characterized by cyst formation and renal fibrosis. Nephrocystin is an adaptor protein that is able to associate with signaling molecules involved in cell adhesion and actin cytoskeleton organization, such as p130Cas, Pyk2, tensin and filamins. Nephrocystin was recently shown to interact and to co-localize with the microtubule component beta-tubulin to the primary cilia in renal epithelial cells, an organelle known to play a key role in the pathogenesis of cystic kidney diseases. In this study, we demonstrated that nephrocystin-4 also localizes to the primary cilia in polarized epithelial tubular cells, particularly at the basal bodies, and associates with microtubule component alpha-tubulin, suggesting a common role for the nephrocystin proteins in ciliary function. However, the co localization of nephrocystin-4 with the microtubules is not restricted to the primary cilia, as nephrocystin-4 was also detected at the centrosomes of dividing cells and close to the cortical actin cytoskeleton in polarized cells. We also detected p130Cas and Pyk2 in the nephrocystin-4-containing complex, confirming the role of the nephrocystin proteins in cell-cell and cell-matrix adhesion signaling events. Finally, we refined the structural and functional regions involved in the interaction between nephrocystin and nephrocystin-4. These data suggest that nephrocystin and nephrocystin-4 belong to a multifunctional complex localized in actin- and microtubule-based structures involved in cell-cell and cell-matrix adhesion signaling as well as in cell division. PMID- 15661759 TI - George W Bush's second term. PMID- 15661760 TI - Opioids for persistent non-cancer pain. PMID- 15661761 TI - Reducing the transmission of genital herpes. PMID- 15661762 TI - Schizophrenia: a genetic disorder of the synapse? PMID- 15661763 TI - Would regional government have been good for your health? PMID- 15661764 TI - Eli Lilly defends itself against assertions in BMJ. PMID- 15661765 TI - Rich nations should pay more to developing countries. PMID- 15661766 TI - Pfizer found guilty of breaching code. PMID- 15661767 TI - Royalty payments to staff researchers cause new NIH troubles. PMID- 15661768 TI - Children face same social problems as they did 100 years ago. PMID- 15661769 TI - Drug company chiefs accept the need for more openness. PMID- 15661770 TI - FDA rejects sale of over the counter statins. PMID- 15661771 TI - MPs disappointed by response to report on allergy services. PMID- 15661772 TI - Bush pushes for limit to medical malpractice awards. PMID- 15661773 TI - New NHS IT system will preserve patient confidentiality. PMID- 15661774 TI - Only a few more deaths from vCJD likely in UK. PMID- 15661775 TI - Generosity after tsunami could threaten neglected crises. PMID- 15661777 TI - US drug industry's claims of other countries "freeloading" are a myth. PMID- 15661776 TI - US health agency scales down plans for free, online research results. PMID- 15661779 TI - Former Czech health minister could be jailed. PMID- 15661780 TI - Delays in implementing e-booking threaten patient choice agenda. PMID- 15661781 TI - Complementary medicine must prove its worth. PMID- 15661783 TI - Do we need to treat vulvovaginitis in prepubertal girls? PMID- 15661784 TI - Activity in later life. PMID- 15661785 TI - The marketing of a disease: female sexual dysfunction. PMID- 15661786 TI - Monitoring global health: bottom up approach is more likely to be successful. PMID- 15661787 TI - Monitoring global health: WHO has mandate and expertise. PMID- 15661788 TI - Monitoring global health: problems are primarily national. PMID- 15661789 TI - Specialised care for early psychosis: like was not compared with like. PMID- 15661790 TI - Limitations and potential of country of birth as proxy for ethnic group. PMID- 15661791 TI - Monitoring global health: Cochrane Collaboration in developing world could be an answer. PMID- 15661792 TI - Preventing and treating hepatitis B infection: immunisation is most important strategy to control hepatitis B. PMID- 15661793 TI - Computer assisted knee arthroplasty is here today. PMID- 15661794 TI - Specialised care for early psychosis: more detail is needed. PMID- 15661795 TI - A prisoner of conscience in Vietnam. PMID- 15661796 TI - High risk groups are still not being vaccinated. PMID- 15661797 TI - Small, fuzzy and interpretable gene expression based classifiers. AB - MOTIVATION: Interpretation of classification models derived from gene-expression data is usually not simple, yet it is an important aspect in the analytical process. We investigate the performance of small rule-based classifiers based on fuzzy logic in five datasets that are different in size, laboratory origin and biomedical domain. RESULTS: The classifiers resulted in rules that can be readily examined by biomedical researchers. The fuzzy-logic-based classifiers compare favorably with logistic regression in all datasets. AVAILABILITY: Prototype available upon request. PMID- 15661799 TI - Concept-based annotation of enzyme classes. AB - MOTIVATION: Given the explosive growth of biomedical data as well as the literature describing results and findings, it is getting increasingly difficult to keep up to date with new information. Keeping databases synchronized with current knowledge is a time-consuming and expensive task-one which can be alleviated by automatically gathering findings from the literature using linguistic approaches. We describe a method to automatically annotate enzyme classes with disease-related information extracted from the biomedical literature for inclusion in such a database. RESULTS: Enzyme names for the 3901 enzyme classes in the BRENDA database, a repository for quantitative and qualitative enzyme information, were identified in more than 100,000 abstracts retrieved from the PubMed literature database. Phrases in the abstracts were assigned to concepts from the Unified Medical Language System (UMLS) utilizing the MetaMap program, allowing for the identification of disease-related concepts by their semantic fields in the UMLS ontology. Assignments between enzyme classes and diseases were created based on their co-occurrence within a single sentence. False positives could be removed by a variety of filters including minimum number of co-occurrences, removal of sentences containing a negation and the classification of sentences based on their semantic fields by a Support Vector Machine. Verification of the assignments with a manually annotated set of 1500 sentences yielded favorable results of 92% precision at 50% recall, sufficient for inclusion in a high-quality database. AVAILABILITY: Source code is available from the author upon request. SUPPLEMENTARY INFORMATION: ftp.uni koeln.de/institute/biochemie/pub/brenda/info/diseaseSupp.pdf. PMID- 15661800 TI - Gene finding for the helical cytokines. AB - MOTIVATION: Gene finding remains an open problem well after the sequencing of the human genome. The low gene sensitivity of current methods is a problem for divergent protein families, because fairly accurate exon assemblies are required before sensitive fold recognition algorithms can be applied. This paper presents a new genomic threading algorithm which integrates the gene finding and fold recognition steps into a single process. The method is applicable to evolutionarily divergent protein families that have retained some trace of their common ancestry, number and phase of introns, sizes of exons and placement of structural elements on specific exons. Such conserved structural signals may be visible despite dramatic evolution of protein sequence. RESULTS: The method is evaluated on the family of helical cytokines by cross-validation sensitivity analysis. The method has also been applied to all intergenic regions of the human genome, and an expression and cloning approach has been coupled with the predictions of the method. Two genes discovered by this method are discussed. SUPPLEMENTARY INFORMATION: All data used and the results obtained in the cross validation analysis are available at http://www.soi.city.ac.uk/~conklin/papers/GT/ PMID- 15661801 TI - Tea and circulating estrogen levels in postmenopausal Chinese women in Singapore. AB - The role of tea in the etiology of breast cancer is controversial. We recently provided the first set of human evidence that breast cancer risk is significantly inversely associated with tea intake, largely confined to intake of green tea. Since black tea and green tea possess comparable levels of the total tea polyphenols that possess antioxidative activities, reasons for the paradoxical effects of green tea and black tea on breast cancer protection are not apparent. Some limited evidence suggests that green tea may have downregulatory effects on circulating sex-steroid hormones, whereas black tea may have upregulatory effects. We therefore, investigated the relationship between tea intake, and plasma estrogen and androstenedione levels in a cross-sectional study of healthy postmenopausal Chinese women in Singapore. In this group of 130 women, 84 were non or irregular (less than once a week) tea drinkers, 27 were regular (weekly/daily) green tea drinkers and 19 were regular (weekly/daily) black tea drinkers. Relative to plasma estrone levels in non- or irregular tea drinkers (29.5 pg/ml) the levels were 13% lower in regular green tea drinkers (25.8 pg/ml) and 19% higher in regular black tea drinkers (35.0 pg/ml). These differences in estrone levels were statistically significant (P = 0.03) inspite of adjusting for age, body mass index, intake of soy, and other covariates. A similar pattern of differences between tea intake, and plasma levels of estradiol (P = 0.08) and androstenedione (P = 0.14) were found. In addition, the tea-estrogen associations were observed irrespective of the genotype of catechol-O-methyltransferase (COMT), a major enzyme that aids in the excretion of tea polyphenols in humans. Larger studies are needed to confirm results from this cross-sectional study and to better understand the potentially differing effect of black and green tea on circulating estrogen levels and ultimately on the risk of breast cancer. PMID- 15661802 TI - Osteopontin silencing by small interfering RNA suppresses in vitro and in vivo CT26 murine colon adenocarcinoma metastasis. AB - Hepatic metastasis is a primary cause for failure of locoregional therapy in colorectal cancer. Increased expression of osteopontin (OPN), a ligand for alpha(v)beta3 integrin and CD44 receptors, is associated with metastasis in several types of cancer. However, the mechanism by which OPN mediates metastasis in colorectal cancer remains unknown. We hypothesized that OPN mediates invasion of colon cancer cells through basement membrane and migration through extracellular matrix (ECM). In this study, we used CT26 murine colon adenocarcinoma cells syngeneic to BALB/c mice to generate cell lines (pS-OPN) in which OPN expression was suppressed through small interfering RNA (siRNA) plasmids. CT26 wild-type cells (WT) and CT26 cells stably expressing murine mismatch siRNA (pS-MM) served as controls. Western blotting quantified OPN protein levels and our most downregulated clone, pS-OPN-A4, demonstrated a mean 3.0-fold decrease in OPN protein expression versus WT. In vitro cell motility and invasiveness were decreased in pS-OPN-A4 by 3.6-fold (P = 0.004 versus WT) and 4.1-fold (P = 0.01 versus WT), but proliferation was similar amongst cell lines. We demonstrated that OPN suppression significantly correlates with MMP-2 downregulation. In vivo hepatic metastasis was assessed by quantifying liver weights and surface tumor nodules in 33 BALB/c mice (11/group) subjected to intrasplenic injection of tumor cells. pS-OPN-A4 resulted in a 50.4% decrease in mean liver weight compared with WT (3.79 +/- 1.49 g versus 1.88 +/- 1.34 g, P = 0.009). Only 18% of pS-OPN-A4 livers had >20 metastatic surface nodules compared with 89% for WT and 75% for pS-MM-V6. This study demonstrates that RNA interference stably reduces CT26 tumor expression of OPN and significantly attenuates CT26 colon cancer metastasis by diminishing tumor cell motility and invasiveness. PMID- 15661804 TI - Human colon cancer cells lacking Bax resist curcumin-induced apoptosis and Bax requirement is dispensable with ectopic expression of Smac or downregulation of Bcl-XL. AB - Multiple apoptotic stimuli induce conformational changes in Bax, a proapoptotic protein from the Bcl-2 family and its deficiency is a frequent cause of chemoresistance in colon adenocarcinomas. Curcumin, a dietary compound from turmeric, is known to induce apoptosis in a variety of cancer cells. To understand the role of Bax in curcumin-induced apoptosis we used HCT116 human colon cancer cells with one allele of Bax gene (Bax+/-) and Bax knockout HCT116 (Bax-/-) cells in which Bax gene is inactivated by homologous recombination. Cell viability decreased in a concentration-dependent manner in Bax+/- cells treated with curcumin (0-50 microM) whereas only minimal changes in viability were observed in Bax-/- cells upon curcumin treatment. In Bax-/- cells curcumin induced activation of caspases 9 and 3 was blocked and that of caspase 8 remained unaltered. Curcumin-induced release of cytochrome c, Second mitochondria derived activator of caspase (Smac) and apoptosis inducing factor (AIF) was also blocked in Bax-/- cells and reintroduction of Bax, downregulation of the antiapoptotic protein Bcl-XL by antisense DNA as well as the overexpression of Smac, highly sensitized the Bax-/- cells toward curcumin-induced apoptosis. There was no considerable difference in the percentage of apoptotic cells in Bak RNAi transfected Bax+/- or Bax-/- cells treated with curcumin when compared with their corresponding vector transfected cells treated with curcumin. The present study demonstrates the role of Bax but not Bak as a critical regulator of curcumin induced apoptosis and implies the potential of targeting antiapoptotic proteins like Bcl-XL or overexpression of proapoptotic proteins like Smac as interventional approaches to deal with Bax-deficient chemo-resistant cancers for curcumin-based therapy. PMID- 15661803 TI - Increased 5-lipoxygenase expression and induction of apoptosis by its inhibitors in esophageal cancer: a potential target for prevention. AB - Arachidonic acid (AA) is the major precursor of several classes of signal molecules and the alteration of its metabolism is involved in human carcinogenesis. For instance, 5-lipoxygenase (5-LOX) converts AA to hydroxyeicosatetraenoic acids or leukotrienes (LTs), which are able to enhance proliferation, increase survival and suppress the apoptosis of human cells. To determine the potential use of 5-LOX inhibitors in the prevention of esophageal cancer, we first analyzed the 5-LOX expression in esophageal tissue samples using immunohistochemistry and then examined the effect of the 5-LOX inhibitors AA861 and REV5901 on cell viability and apoptosis in esophageal cancer cell lines. 5 LOX expression was present in 79% (127/161) of esophageal cancer but in only 13% (4/32) of normal esophageal mucosa. 5-LOX was also expressed in all the eight esophageal cancer cell lines. Moreover, 5-LOX inhibitors caused a dose- and time dependent reduction of cell viability, which was due to the induction of apoptosis and associated with LTB4 suppression. Our data also showed that both LTB4, a product of 5-LOX and LTB4 receptor antagonist U-75302 were able to prevent AA861 and REV5901 on induction of apoptosis. The present study demonstrated that 5-LOX protein expression is increased in esophageal cancer and that 5-LOX inhibitors can induce esophageal cancer cells to undergo apoptosis, suggesting that 5-LOX may be an effective target in the prevention of esophageal cancer. PMID- 15661805 TI - Combined inhibition of PDGF and VEGF receptors by ellagic acid, a dietary-derived phenolic compound. AB - The vascular endothelial growth factor (VEGF) and platelet-derived growth factor (PDGF) receptors play essential and complementary roles in angiogenesis and combined inhibition of these receptors has been shown to result in potent antitumor activity in vivo. In this study, we report that ellagic acid (EA), a natural polyphenol found in fruits and nuts, inhibits VEGF-induced phosphorylation of VEGFR-2 in endothelial cell (EC) as well as PDGF-induced phosphorylation of PDGFR in smooth muscle cells, leading to the inhibition of downstream signaling triggered by these receptors. EA also specifically inhibited VEGF-induced migration of ECs as well as their differentiation into capillary like tubular structures and abolished PDGF-dependent smooth muscle cell migration. Interestingly, EA presents a greater selectivity for normal cells than for tumor cells since the migration of the U87 and HT1080 cell lines were much less affected by this molecule. The identification of EA as a naturally occurring dual inhibitor of VEGF and PDGF receptors suggests that this molecule possesses important antiangiogenic properties that may be helpful for the prevention and treatment of cancer. PMID- 15661806 TI - Combinations of glutathione S-transferase genotypes and risk of early-onset lung cancer in Caucasians and African Americans: a population-based study. AB - Polymorphisms in GSTM1, GSTT1 and GSTP1 genes in humans are associated with the reduction of enzymatic activity toward several substrates, including those in tobacco smoke. To investigate the potential role these polymorphisms have, as modulators of early-onset lung cancer risk, a population-based case-control study involving early-onset lung cancer cases was performed. Biological samples were available for 350 individuals diagnosed <50 years of age identified from the metropolitan Detroit Surveillance, Epidemiology and End Results (SEER) program and 410 cases of age, race and sex-matched controls ascertained through random digit dialing. African Americans carrying at least one G allele at the GSTP1 locus were 2.9-fold more likely to have lung cancer compared with African Americans without a G allele after adjustment for age, sex, pack years of smoking and history of lung cancer in a first-degree relative (95% CI 1.29-6.20). African Americans with either one or two risk genotypes at the GSTM1 and GSTP1 loci were at increased risk of having lung cancer compared with those having fully functional GSTM1 and GSTP1 genes (OR = 2.8, 95% CI 1.1-7.2 and OR = 4.0, 95% CI 1.3-12.2, respectively). No significant single gene associations between GSTM1, GSTT1 or GSTP1 and early-onset lung cancer were identified in Caucasians, after adjusting for age, sex, pack years and family history of lung cancer. However, our results suggest that specific combinations of glutathione S-transferase polymorphisms increase the risk of early-onset of lung cancer. Joint analysis of these genotypes may identify individuals who are at a higher risk of developing early-onset lung cancer with a greater certainty than single gene studies. PMID- 15661807 TI - Prostaglandin E2 promotes migration and adhesion in hepatocellular carcinoma cells. AB - The effect of the expression of cyclooxygenase-2 (COX-2) and prostaglandin E2 (PGE2) synthesis on cell migration, the secretion of matrix metalloproteinases (MMPs) and the adhesion of human hepatoma cell lines has been investigated. A close correlation was observed between the expression of COX-2 under basal conditions and the secretion of MMP-2 and MMP-9. Cell migration in HuH-7 cells, which express high constitutive levels of COX-2 was significantly inhibited by selective inhibitors of COX-2 and enhanced by exogenous addition of PGE2. Hepatocellular carcinoma (HCC) cells expressed beta1 and alphaV beta3 integrins, exhibiting an increase in cell adhesion onto fibronectin and vitronectin. Moreover, addition of PGE2 increased the beta1 integrin levels and adhesion on vitronectin in HuH-7 cells. Inhibitors of MEK/ERK, p38 MAPK, protein kinases A and C impaired the migration of HuH-7 cells induced by PGE2, indicating the involvement of multiple pathways in the process. Taken together, these results support the existence of a relationship between COX-2-derived PGE2 synthesis, and migration and adhesion through an integrin-dependent pathway in HCC cells. PMID- 15661808 TI - Involvement of transcription factor Sp1 in quercetin-mediated inhibitory effect on the androgen receptor in human prostate cancer cells. AB - The transactivation function of the human androgen receptor (AR) can be regulated by several coregulators that may be either positive or negative. Ubiquitous transcription factor Sp1 not only regulates the basal expression of the AR but also acts as its coregulator. Our previous study has shown that quercetin, one of the main polyphenols, can effectively inhibit the expression and function of the AR. The present study is to address if quercetin may affect Sp1's action on AR transactivation activity in human prostate adenocarcinoma cell lines, LNCaP and PC-3. First, we showed that indeed in transient transfections Sp1 could enhance transcriptional activity of the AR promoter and of androgen upregulated gene promoters, i.e. the prostate-specific antigen and the hK2 genes. Interestingly, the enhancing activity of Sp1 could be repressed by quercetin. The gel shift and western blot analyses indicated that the specific DNA motif binding activity of Sp1 and its protein levels were not altered by quercetin. However, the state of interaction of Sp1 with the AR treated by quercetin plus androgen was different from that by androgen treatment or none as demonstrated by coimmunoprecipitation experiments and glutathione S-transferase (GST) pull-down assays. Moreover, we showed that quercetin caused changes in post-translational modification of AR protein. The above findings strongly suggest that changes induced by quercetin in post-translational modification of the AR and in states of physical interaction of Sp1 with the AR may be critical for the attenuation of AR's function. PMID- 15661809 TI - Silencing of genes by promoter hypermethylation: key event in rodent and human lung cancer. AB - Transcriptional silencing by CpG island hypermethylation has become a critical component in the initiation and progression of lung cancer. The ability of pharmacologic agents to reverse promoter hypermethylation also makes it an attractive target to pursue for prevention of lung cancer. Animal models, together with studies in humans have fostered significant advances in elucidating the role of gene-specific methylation in cancer initiation and progression, the modulation of promoter methylation by carcinogen exposure and the ability to block tumor development by preventing epigenetically mediated gene silencing. These advances represent the beginning of efforts to develop a progression model for lung cancer that should aid efforts for early detection and gene targeting for therapy, and the development of preventive interventions that reverse epigenetic-mediated gene silencing. PMID- 15661811 TI - 3,3'-Diindolylmethane inhibits angiogenesis and the growth of transplantable human breast carcinoma in athymic mice. AB - Studies have linked the consumption of broccoli and other cruciferous vegetables to a reduced risk of breast cancer. The phytochemical indole-3-carbinol (I3C), present in cruciferous vegetables, and its major acid-catalyzed reaction product 3,3'-diindolylmethane (DIM) have bioactivities relevant to the inhibition of carcinogenesis. In this study, the effect of DIM on angiogenesis and tumorigenesis in a rodent model was investigated. We found that DIM produced a concentration-dependent decrease in proliferation, migration, invasion and capillary tube formation of cultured human umbilical vein endothelial cells (HUVECs). Consistent with its antiproliferative effect, which was significant at only 5 microM DIM, this indole caused a G1 cell cycle arrest in actively proliferating HUVECs. Furthermore, DIM downregulated the expression of cyclin dependent kinases 2 and 6 (CDK2, CDK6), and upregulated the expression of CDK inhibitor, p27(Kip1), in HUVECs. We observed further in a complementary in vivo Matrigel plug angiogenesis assay that, compared with vehicle control, neovascularization was inhibited up to 76% following the administration of 5 mg/kg DIM to female C57BL/6 mice. Finally, this dose of DIM also inhibited the growth of human MCF-7 cell tumor xenografts by up to 64% in female athymic (nu/nu) mice, compared with the vehicle control. This is the first study to show that DIM can strongly inhibit the development of human breast tumor in a xenograft model and to provide evidence for the antiangiogenic properties of this dietary indole. PMID- 15661810 TI - Decreased n-6/n-3 fatty acid ratio reduces the invasive potential of human lung cancer cells by downregulation of cell adhesion/invasion-related genes. AB - Recent studies have shown opposing effects of n-6 and n-3 fatty acids on the development of cancer and suggest a role for the ratio of n-6 to n-3 fatty acids in the control of cancer. However, whether an alteration in the n-6/n-3 fatty acid ratio of cancer cells affects their invasive potential has not been well investigated. We recently developed a genetic approach to modify the n-6/n-3 ratio by expression of the Caenorhabditis elegans fat-1 gene encoding an n-3 desaturase that converts n-6 to n-3 fatty acids in mammalian cells. The objective of this study was to examine the effect of alteration in the n-6/n-3 fatty acid ratio on the invasive potential of human lung cancer A549 cells. Adenovirus mediated gene transfer of the n-3 desaturase resulted in a marked reduction of the n-6/n-3 fatty acid ratio, particularly the ratio of arachidonic acid to eicosapentaenic acid. Cell adhesion assay showed that the cells expressing fat-1 gene had a delayed adhesion and retarded colonization. Matrigel assay for invasion potential indicated a 2-fold reduction of cell migration in the fat-1 transgenic cells when compared with the control cells. An increased apoptosis was also observed in the fat-1 transgenic cells. Microarray and quantitative polymerase chain reaction revealed a downregulation of several adhesion/invasion related genes (MMP-1, integrin-alpha2 and nm23-H4) in the fat-1 transgenic cells. These results demonstrate that a decreased n-6/n-3 fatty acid ratio reduces the invasion potential of human lung cancer cells by probably downregulating the cell adhesion/invasion-related molecules, suggesting a role for the ratio of n-6 to n 3 fatty acids in the prevention and treatment of cancer. PMID- 15661812 TI - Antineoplastic cyclic astin analogues kill tumour cells via caspase-mediated induction of apoptosis. AB - Astins, a family of cyclopentapeptides, isolated from the roots of a medicinal plant Aster tataricus (Compositae), show antitumour activity. Their chemical structures consist of a 16-membered ring system containing a unique beta,gamma dichlorinated proline [Pro(Cl2)], other non-coded amino acid residues, and a cis conformation in one of the peptide bonds. The beta,gamma-dichlorinated proline residue is considered to play an important role in their antineoplastic activities in vitro on nasopharynx carcinoma (KB) cells and in vivo on sarcoma 180 ascites and P388 lymphocytic leukaemia in mice. The acyclic astins without Pro(Cl2) do not show antitumour activity against S-180 ascites in vivo, suggesting that the cyclic nature of astins plays an important role in their antitumour activities. We synthesized new astin-related cyclopeptides differing from the natural product for the presence of some non-proteinogenic amino acid residues: Aib, Abu, -S(beta3)-hPhe and a peptide bond surrogate (-SO2-NH-) and we tested for their antitumour effect. We observed cytotoxic effects of the newly synthesized cyclic astins, but not with the acyclic analogue astins. We also observed that the cyclic astin induced apoptosis in a human papillary thyroid carcinoma cell line (NPA cell line) and that apoptotis was associated with activation of caspases. The caspase family inhibitor, Z-Val-Asp-(OMe)-FMK, protected NPA cells from cyclic analogue astin-induced apoptosis. To determine which caspase was specifically activated, we assayed caspase activity in astin treated cells in the presence of specific caspase and 8, 9 or 3 inhibitors, i.e. Z-IETD-FMK, Z-LEHD-FMK Z-DEVD-FMK, which inhibit caspases 8, 9 and 3, respectively. The data presented here show selective antineoplastic properties of the newly synthesized cyclic astins, and suggest, for the first time, a mechanism for their antineoplastic action through the activation of apoptotic pathway. PMID- 15661813 TI - 5,7-Dimethoxyflavone downregulates CYP1A1 expression and benzo[a]pyrene-induced DNA binding in Hep G2 cells. AB - The objective of this study was to examine the ability of dietary polyphenols to inhibit cytochrome P450 (CYP) 1A1 expression and activity and benzo[a]pyrene (BaP) DNA binding, with the main emphasis on prevention of chemical-induced hepatic carcinogenesis. For this purpose we used Hep G2 cells, a good model of the normal human hepatocyte for CYP1A1 cell signaling. First, when these cells were exposed to a low concentration (1 microM) of BaP, DNA binding occurred, which dramatically increased after 6 h of treatment. BaP also dramatically induced CYP1A1 activity, protein expression and mRNA levels, the likely reason for the marked increase in DNA binding. Second, we screened 25 polyphenols with highly varying chemical structures for maximum ability to inhibit CYP1A1 activity in the Hep G2 cells. Highly varying responses were obtained, ranging from a 10 fold induction by some polyphenols to almost complete inhibition, in particular by 5,7-dimethoxyflavone (DMF), a flavonoid found in some tropical plants. Third, we examined the ability of DMF to inhibit DNA binding of BaP and the mechanisms involved. DMF (2-20 microM) inhibited BaP-induced DNA binding. DMF also inhibited BaP-induced CYP1A1 activity, CYP1A1 protein expression and mRNA levels. Moreover, DMF directly inhibited the catalytic activity of recombinant CYP1A1 with an IC50 of 0.8 microM. In conclusion, DMF was a highly potent inhibitor of BaP-induced DNA binding and CYP1A1 protein expression and activity in the Hep G2 cells. These properties may make DMF an effective chemoprotectant in chemical-induced liver cancer. PMID- 15661814 TI - The physiological regulation of toll-like receptor expression and function in humans. AB - Eleven mammalian toll-like receptors (TLRs 1-11) have been identified to date and are known to play a crucial role in the regulation of immune responses; however, the factors that regulate TLR expression and function in vivo are poorly understood. Therefore, in the present study, we investigated the physiological regulation of TLR expression and function in humans. To examine the influence of diurnal rhythmicity on TLR expression and function, peripheral venous blood samples were collected from healthy volunteers (n = 8) at time points coinciding with the peak and nadir in the endogenous circulating cortisol concentration. While no diurnal rhythmicity in the expression of TLRs 1, 2, 4 or 9 was observed, the upregulation of costimulatory (CD80 and CD86) and antigen-presenting (MHC class II) molecules on CD14(+) monocytes following activation with specific TLR ligands was greater (P < 0.05) in samples obtained in the evening compared with the morning. To examine the influence of physical stress on TLR expression and function, peripheral venous blood samples were collected from healthy volunteers (n = 11) at rest and following 1.5 h of strenuous exercise in the heat (34 degrees C). Strenuous exercise resulted in a decrease (P < 0.005) in the expression of TLRs 1, 2 and 4 on CD14(+) monocytes. Furthermore, the upregulation of CD80, CD86, MHC class II and interleukin-6 by CD14(+) monocytes following activation with specific TLR ligands was decreased (P < 0.05) in samples obtained following exercise compared with at rest. These results demonstrate that TLR function is subject to modulation under physiological conditions in vivo and provide evidence for the role of immunomodulatory hormones in the regulation of TLR function. PMID- 15661815 TI - Time-matched pre- and postsynaptic changes of GABAergic synaptic transmission in the developing mouse superior colliculus. AB - Developmental changes in the kinetics of GABAergic postsynaptic currents have been reported for various brain structures. However, it has remained unclear whether these modifications are matched by presynaptic changes. We addressed this question by analysing evoked IPSCs (eIPSCs) in mouse superior colliculus slices between postnatal day (P) 1 and 22. eIPSCs were elicited by electrical stimulation and measured in the whole-cell patch-clamp configuration. IPSCs were analysed using the binomial model of synaptic transmission. The readily releasable pool (RRP, N) was estimated from the cumulative eIPSC amplitude histograms during 50-Hz stimulation. Median delayed IPSC (dIPSC) amplitude was used as a quantal amplitude (q) estimate. The mean release probability (p) was determined as the mean eIPSC amplitude divided by the product of RRP and q. The experiments revealed that GABAergic synapses pass through two distinct periods of functional adjustment: (i) P1-3 (coincidental with the onset of glutamatergic spontaneous activity and a switch from depolarizing to hyperpolarizing GABA action) displayed a significant decrease of p, associated with an increase in the paired-pulse ratio (eIPSC(2)/eIPSC(1)); and (ii) P6-15 (the period before and shortly after eye opening) is characterized by a drastic reduction of IPSC duration. On the presynaptic side, it was accompanied by a down-regulation of asynchronous release in favour of stimulus-locked synchronous release. We conclude that postsynaptic modifications of GABAergic synaptic transmission in the superior colliculus (SC) are indeed accompanied by presynaptic changes, and this may guarantee the necessary efficacy of inhibition during the developmental reconstruction of the synaptic network in the SC. PMID- 15661816 TI - Mechanisms of acetylcholine-mediated vasodilatation in young and aged human skin. AB - Thermoregulatory cutaneous vasodilatation (VD) is attenuated in aged skin. While acetylcholine (ACh) plays a role in thermally mediated VD, the precise mechanisms through which ACh-mediated VD acts and whether those downstream mechanisms change with ageing are unclear. We tested the hypotheses that both nitric oxide (NO)- and prostanoid-mediated pathways contribute to exogenous ACh-mediated VD, and that both are attenuated with advanced age. Twelve young (Y: 23 +/- 1 years) and 10 older (O: 69 +/- 1 years) subjects underwent infusions of 137.5 mum ACh at four intradermal microdialysis sites: control (C, Ringer solution), NO synthase inhibited (NOS-I, 10 mm l-NAME), cyclooxygenase inhibited (COX-I, 10 mm ketorolac) and NOS-I + COX-I. Red blood cell flux was monitored using laser Doppler flowmetry, and cutaneous vascular conductance (CVC) was calculated (laser Doppler flux/mean arterial pressure) and normalized to maximal CVC (%CVC(max)) (28 mm sodium nitroprusside + local heating to 43 degrees C). Baseline %CVC(max) was increased in the O at COX-I sites (COX-I 16 +/- 1, NOS-I + COX-I 16 +/- 2 versus C 10 +/- 1%CVC(max); P < 0.001) but not in the young, suggesting an age related shift toward COX vasoconstrictors contributing to basal cutaneous vasomotor tone. There was no difference in peak %CVC(max) during ACh infusion between age groups, and the response was unchanged by NOS-I (O: NOS-I 35 +/- 5 versus C 38 +/- 5%CVC(max); P = 0.84) (Y: NOS-I 41 +/- 4 versus C 39 +/- 4%CVC(max); P = 0.67). COX-I and NOS-I + COX-I attenuated the peak CVC response to ACh in both groups (COX-I O: 29 +/- 3, Y: 22 +/- 2%CVC(max) versus C; P < 0.001 both groups; NOS-I + COX-I O: 32 +/- 3 versus Y: 29 +/- 2%CVC(max); versus C; P < 0.001 both groups). ACh mediates cutaneous VD through prostanoid and non NO-, non-prostanoid-dependent pathways. Further, older subjects have a diminished prostanoid contribution to ACh-mediated VD. PMID- 15661817 TI - Reduced glycine transporter type 1 expression leads to major changes in glutamatergic neurotransmission of CA1 hippocampal neurones in mice. AB - To investigate the effects of persistent elevation of synaptic glycine at Schaffer collateral-CA1 synapses of the hippocampus, we studied the glutamatergic synaptic transmission in acute brain slices from mice with reduced expression of glycine transporter type 1 (GlyT1+/-) as compared to wild type (WT) littermates using whole-cell patch-clamp recordings of CA1 pyramidal cells. We observed faster decay kinetics, reduced ifenprodil sensitivity and increased zinc-induced antagonism in N-methyl-d-aspartate receptor (NMDAR) currents of GlyT1+/- mice. Moreover, the ratio alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor (AMPAR)/NMDAR was decreased in mutants compared to WT. Surprisingly, this change was associated with a reduction in the number of AMPARs expressed at the CA1 synapses in the mutants compared to WT. Overall, these findings highlight the importance of GlyT1 in regulating glutamatergic neurotransmission. PMID- 15661818 TI - G protein {beta}{gamma} subunits mediate presynaptic inhibition of transmitter release from rat superior cervical ganglion neurones in culture. AB - The activation of presynaptic G protein-coupled receptors (GPCRs) is widely reported to inhibit transmitter release; however, the lack of accessibility of many presynaptic terminals has limited direct analysis of signalling mediators. We studied GPCR-mediated inhibition of fast cholinergic transmission between superior cervical ganglion neurones (SCGNs) in culture. The adrenoceptor agonist noradrenaline (NA) caused a dose-related reduction in evoked excitatory postsynaptic potentials (EPSPs). NA-induced EPSP decrease was accompanied by effects on the presynaptic action potential (AP), reducing AP duration and amplitude of the after-hyperpolarization (AHP), without affecting the pre- and postsynaptic membrane potential. All effects of NA were blocked by yohimbine and synaptic transmission was reduced by clonidine, consistent with an action at presynaptic alpha2-adrenoceptors. NA-induced inhibition of transmission was sensitive to pre-incubation of SCGNs with pertussis toxin (PTX), implicating the involvement of Galpha(i/o)betagamma subunits. Expression of Galpha transducin, an agent which sequesters G protein betagamma (Gbetagamma) subunits, in the presynaptic neurone caused a time-dependent attenuation of NA-induced inhibition. Injection of purified Gbetagamma subunits into the presynaptic neurone inhibited transmission, and also reduced the AHP amplitude. Furthermore, NA-induced inhibition was occluded by pre-injection of Gbetagamma subunits. The Ca(2+) channel blocker Cd(2+) mimicked NA effects on transmitter release. Cd(2+), NA and Gbetagamma subunits also inhibited somatic Ca(2+) current. In contrast to effects on AP-evoked transmitter release, NA had no clear action on AP-independent EPSPs induced by hypertonic solutions. These results demonstrate that Gbetagamma subunits functionally mediate inhibition of transmitter release by alpha2 adrenoceptors and represent important regulators of synaptic transmission at mammalian presynaptic terminals. PMID- 15661819 TI - Carbon dioxide sensitivity during hypoglycaemia-induced, elevated metabolism in the anaesthetized rat. AB - We have utilized an anaesthetized rat model of insulin-induced hypoglycaemia to test the hypothesis that peripheral chemoreceptor gain is augmented during hypermetabolism. Insulin infusion at 0.4 U kg (-1)min(-1) decreased blood glucose concentration significantly to 3.37 +/- 0.12 mmol l(-1). Whole-body metabolism and basal ventilation were elevated without increase in P(a,CO(2)) (altered non significantly from the control level, to 37.3 +/- 2.6 mmHg). Chemoreceptor gain, measured either as spontaneous ventilatory airflow sensitivity to P(a,CO(2)) during rebreathing, or by phrenic minute activity responses to altered P(a,CO(2)) induced by varying the level of artificial ventilation, was doubled during the period of hypermetabolism. This stimulatory effect was primarily upon the mean inspiratory flow rate, or phrenic ramp component of breathing and was reduced by 75% following bilateral carotid sinus nerve section. In vitro recordings of single carotid body chemoafferents showed that reducing superfusate glucose concentration from 10 mM to 2 mM reduced CO(2) chemosensitivity significantly from 0.007 +/- 0.002 Hz mmHg(-1) to 0.001 +/- 0.002 Hz mmHg(-1). Taken together, these data suggest that the hyperpnoea observed during hypermetabolism might be mediated by an increase in the CO(2) sensitivity of the carotid body, and this effect is not due to the insulin-induced fall in blood glucose concentration. PMID- 15661820 TI - Induced sharp wave-ripple complexes in the absence of synaptic inhibition in mouse hippocampal slices. AB - The characteristic, behaviour-related network oscillations of the mammalian hippocampus (, gamma and ripples) are accompanied by strongly phase-coupled action potentials in specific subsets of GABAergic interneurones. It has been suggested that the resulting phasic, repetitive inhibition shapes rhythmic coherent activity of the neuronal network. Here, we examined whether synaptic inhibition entrains approximately 200 Hz network ripples by applying the GABA(A) receptor antagonist gabazine to CA1 minislices of mouse hippocampus. Gabazine blocked spontaneously occurring sharp wave-ripple (SPW-R) activity. However, local application of KCl to the dendritic layer elicited excitatory sharp waves on which approximately 200 Hz ripple oscillations were superimposed with equal temporal properties of native SPW-R. The activity also persisted in the additional presence of blockers of glutamatergic synaptic transmission. In contrast, synchrony was largely abolished after addition of gap junction blockers. Thus, GABAergic transmission appears to be involved in the generation of sharp waves but phasic inhibition is no prerequisite for the precise synchronization of hippocampal neurones during high-frequency oscillations at approximately 200 Hz. Gap junctions on the other hand seem to be necessary to orchestrate coordinated activity within the ripple frequency domain. PMID- 15661821 TI - Impact of glucose infusion on the structural and functional characteristics of adipose tissue and on hypothalamic gene expression for appetite regulatory neuropeptides in the sheep fetus during late gestation. AB - In the present study, our aim was to determine whether intrafetal glucose infusion increases fetal adiposity, synthesis and secretion of leptin and regulates gene expression of the 'appetite regulatory' neuropeptides neuropepetide Y (NPY), agouti-related peptide (AGRP), pro-opiomelanocortin (POMC) and cocaine- and amphetamine-regulated transcript (CART) and receptors (leptin receptor (OB-Rb) and melancortin 3 receptor (MC3R)) within the fetal hypothalamus. Glucose (50% dextrose in saline) or saline was infused (7.5 ml h( 1)) into fetal sheep between 130 and 140 days gestation (term = 150 +/- 3 days gestation). Glucose infusion increased circulating glucose and insulin concentrations, mean lipid locule size (532.8 +/- 3.3 microm2 versus 456.7 +/- 14.8 microm2) and total unilocular fat mass (11.7 +/- 0.6 g versus 8.9 +/- 0.6 g) of the perirenal fat depot. The expression of OB-Rb mRNA was higher in the ventromedial nucleus compared to the arcuate nucleus of the hypothalamus in both glucose and saline infused fetuses (F= 8.04; P < 0.01) and there was a positive correlation between expression of OB-Rb and MC3R mRNA in the arcuate nucleus (r= 0.81; P < 0.005). Glucose infusion increased mRNA expression for POMC, but not for the anorectic neuropeptide CART, or the orexigenic neuropeptides NPY and AGRP, in the arcuate nucleus of the fetal hypothalamus. These findings demonstrate that increased circulating glucose and insulin regulate gene expression of the neuropeptides within the fetal hypothalamus that are part of the neural network regulating energy balance in adult life. PMID- 15661822 TI - Reduced ventricular flow propagation velocity in elite athletes is augmented with the resumption of exercise training. AB - Chronic exercise induces physiological enlargement of the left ventricle ('athlete's heart'), but the effects of current and long-term exercise training on diastolic function have not been investigated. Echocardiography and Doppler imaging were used to assess left ventricular (LV) dimensions and indices of diastolic filling in 22 elite athletes at the end of their 'off-season' (baseline) and, subsequently, following 3 and 6 months of training. Twelve matched controls were also studied at baseline, 3 and 6 months. Compared to controls at baseline, athletes exhibited significantly higher LV mass (235.7 +/- 7.1 g versus 178.1 +/- 14.5 g, P < 0.01) and reduced flow propagation velocity (V(P): 50.21 +/- 1.7 versus 72.2 +/- 3.6 cm s(-1), P < 0.01), a measure of diastolic function. Three months of training further increased LV mass in athletes (253.2 +/- 7.1 g; P < 0.01 versus baseline), and significantly increased their V(P) (66.7 +/- 2.5 cm s(-1); P < 0.05 versus baseline). These trends for increased mass and diastolic filling persisted following 6 months of training (LV mass 249.0 +/- 8.7 g P < 0.05 versus baseline; V(P) 75.7 +/- 3.0 cm s(-1); P < 0.01 versus baseline, and P = 0.01 versus 3 months). This study suggests that following a period of relative inactivity the rate of ventricular relaxation during early diastole may be slowed in athletes who exhibit ventricular hypertrophy, whilst resumption of training increases the speed of ventricular relaxation in the presence of further hypertrophy of the left ventricle. PMID- 15661823 TI - Simultaneous changes of cell volume and cytosolic calcium concentration in macula densa cells caused by alterations of luminal NaCl concentration. AB - Cell volume and cytosolic Ca(2+) concentration ([Ca(2+)](i)) were measured in rabbit macula densa (MD) cells loaded with calcein and Fura Red using confocal microscopy. [Ca(2+)](i) was also analysed with Indo-1 and fura-2. We used isolated microperfused thick ascending limbs with attached glomerulus. The results showed that when the luminal NaCl concentration ('NaCl') was decreased from 35 to 10 mM, the cell volume decreased by 10.4%, and [Ca(2+)](i) increased by 9.5%. This increase was inhibited in Ca(2+)-free solution. When luminal [NaCl] was changed from 35 to 135 mM, the cell volume increased by 15.1%, and [Ca(2+)](i) did not change. The cell volume alterations were not different in Ca(2+)-free solutions. Using Indo-1, basal [Ca(2+)](i) in MD cells was 107.8 nM. When luminal [NaCl] was changed from 135 to 10 mm, [Ca(2)](i) increased by 23.5 nM. Using fura-2, the basal [Ca(2+)](i) in MD cells was 115.3 nM, and when luminal [NaCl] was changed from 135 or 35 to 10 mM, [Ca(2+)](i) change was 30.1 or 10.6 nM, respectively. An increase in [NaCl] caused no change in [Ca(2+)](i). In Ca(2+)-free solution, no change in [Ca(2+)](i) occurred. A stepwise decrease in luminal [NaCl] resulted in a sigmoid increase in [Ca(2+)](i) in MD cells. The steepest part of the curve was between 70 and 10 mM. In conclusion, we found that MD cells have cell volume regulation, and that [Ca(2+)](i) elevation caused by decreased luminal [NaCl] is independent of the cell volume. PMID- 15661824 TI - Human postural sway results from frequent, ballistic bias impulses by soleus and gastrocnemius. AB - It has been widely assumed for nearly a century, that postural muscles operate in a spring-like manner and that muscle length signals joint angle (the mechano reflex mechanism). Here we employ automated analysis of ultrasound images to resolve calf muscle (soleus and gastrocnemius) length changes as small as 10 mum in standing subjects. Previously, we have used balancing of a real inverted pendulum to make predictions about human standing. Here we test and confirm these predictions on 10 subjects standing quietly. We show that on average the calf muscles are actively adjusted 2.6 times per second and 2.8 times per unidirectional sway of the body centre of mass (CoM). These alternating, small (30-300 microm) movements provide impulsive, ballistic regulation of CoM movement. The timing and pattern of these adjustments are consistent with multisensory integration of all information regarding motion of the CoM, pattern recognition, prediction and planning using internal models and are not consistent with control solely by local reflexes. Because the system is unstable, errors in stabilization provide a perturbation which grows into a sway which has to be reacted to and corrected. Sagittal sway results from this impulsive control of calf muscle activity rather than internal sources (e.g. the heart, breathing). This process is quite unlike the mechano-reflex paradigm. We suggest that standing is a skilled, trial and error activity that improves with experience and is automated (possibly by the cerebellum). These results complement and extend our recent demonstration that paradoxical muscle movements are the norm in human standing. PMID- 15661825 TI - Active, non-spring-like muscle movements in human postural sway: how might paradoxical changes in muscle length be produced? AB - In humans, during standing the calf muscles soleus and gastrocnemius actively prevent forward toppling about the ankles. It has been generally assumed that these postural muscles behave like springs with dynamic stiffness reflecting their mechanical properties, reflex gain including higher derivatives, and central control. Here, for the first time, we have used an ultrasound scanner and automated image analysis to record the tiny muscular movements occurring in normal standing. This new, non-invasive technique resolves changes in muscle length as small as 10 mum without disturbing the standing process. This technical achievement has allowed us to test the long-established mechano-reflex, muscle spring hypothesis that muscle length changes in a spring-like way during sway of the body. Our results contradict that hypothesis. Muscle length changes in a non spring-like manner: on average, shortening during forward sway and lengthening during backwards sway (paradoxical movements). This counter-intuitive result is a consequence of the fact that calf muscles generate tension through a series elastic component (SEC, Achilles tendon and foot) which limits maximal ankle stiffness to 92 +/- 20% of that required to balance the body. Paradoxical movements cannot be generated by stretch reflexes with constant intrafusal drive but might be produced by reflex coupling of extrafusal (alpha) and intrafusal (beta, gamma) drive or by positive force feedback. Standing requires the predictive ability to produce the observed muscle movements preceded (110 +/- 50 ms) by corresponding changes in integrated EMG signal. We suggest higher level anticipatory control is more plausible. PMID- 15661826 TI - The role of preimplantation genetic diagnosis in the management of severe rhesus alloimmunization: first unaffected pregnancy: case report. AB - Rhesus (Rh) D alloimmunization may cause haemolytic disease of the fetus and newborn if the fetal Rh blood type is positive. Although the incidence of severe RhD alloimmunization has decreased with prophylactic anti-D immunoglobulin administration during and after pregnancy, sensitization still occurs in a small group of women. In such women, Rh disease will continue to be significant problem and for their babies who may be affected. Preimplantation genetic diagnosis (PGD) may be utilized to avoid materno-fetal blood group incompatibility in an RhD sensitized woman. Biopsy of a single cell from early cleavage-stage embryos screening for RhD-negative embryos allows the transfer of only RhD-negative embryo(s) into the uterus. This avoids any complications related to haemolytic disease of the fetus and newborn. This article describes the first reported case of an unaffected pregnancy using PGD for Rh disease. IVF and embryo transfer resulted in a clinical pregnancy and the birth of a healthy girl confirmed to be blood type RhD negative. PGD in couples with a heterozygous RhD-positive male partner provides an option for avoiding haemolytic disease of the newborn in RhD alloimmunized mothers. PMID- 15661827 TI - Thymocyte stimulation by anti-TCR-beta, but not by anti-TCR-alpha, leads to induction of developmental transcription program. AB - Anti-T cell receptor (aTCR) antibody (Ab) stimulation of T cells results in TCR down-modulation and T cell activation. Differences in the effect of anti-alpha chain and beta-chain Ab have been reported on thymocytes. Anti-beta-chain Ab but not anti-alpha-chain reagents cause long-term TCR down-modulation. However, both types of Ab result in TCR cross-linking and activate early steps in signal transduction. In this study, we show that TCR internalization and calcium flux, hallmarks of T cell activation, are similar with aValpha and aVbeta treatment. Therefore, we have compared the gene expression profiles of preselection thymocytes stimulated with these reagents. We find that aValpha treatment does not cause any significant change in gene expression compared with control culture conditions. In contrast, aVbeta stimulation results in numerous changes in gene expression. The alterations of expression of genes known to be expressed in thymocytes are similar to changes caused by positive thymic selection, suggesting that the expression of some of the genes without known roles in thymocyte development and of novel genes whose expression is found to be altered may also be involved in this process. PMID- 15661829 TI - Fc gammaRIIIB stimulation promotes beta1 integrin activation in human neutrophils. AB - The molecular stimuli involved in receptor-induced integrin activation are still poorly defined. We have investigated the role of receptors for the Fc portion of immunoglobulin G molecules (Fc gammaR) on activation of integrins in human neutrophils. Cross-linking of Fc gammaRIIA induced an increase in surface expression of beta2 integrins but had no effect on beta1 integrins. In contrast, cross-linking of Fc gammaRIIIB not only increased beta2 integrins on the cell surface but also induced beta1 integrin activation, as indicated by an increase in binding to fibronectin and the appearance of an activation epitope detected by the monoclonal antibody 15/7. The Fc gammaRIIIB-induced increase of beta2 integrins required Src-family tyrosine kinases, Syk kinase, and phosphatidylinositol-3 kinase (PI-3K), as the corresponding, specific inhibitors, PP2, Piceatannol, and LY294002, completely blocked it. Contrary to this, Fc gammaRIIIB-induced beta1 integrin activation was not blocked by PP2 or LY294002. It was, however, enhanced by Piceatannol. After Fc gammaRIIIB cross-linking, colocalization of Fc gammaRIIIB and active beta1 integrins was detected on the neutrophil membrane. These data show, for the first time, that cross-linking of Fc gammaRIIIB induces an inside-out signaling pathway that leads to beta1 integrin activation. This activation is independent of Src-family kinases, and PI 3K and may be induced in part by the interaction of Fc gammaRIIIB with beta1 integrins. PMID- 15661828 TI - Analysis of the role of the PAC1 receptor in neutrophil recruitment, acute-phase response, and nitric oxide production in septic shock. AB - Infections caused by Gram-negative bacteria constitute one of the major causes of septic shock, which results from the inability of the immune system to limit bacterial spread during the ongoing infection. In the last decade, it has been demonstrated that vasoactive intestinal peptide (VIP) and pituitary adenylate cyclase-activating polypeptide (PACAP) are two endogenous immunopeptides, which together with three G protein-coupled receptors (VPAC1, VPAC2, and PAC1) exert a significant, therapeutic effect attenuating the deleterious consequences of septic shock by balancing pro- and anti-inflammatory factors. We have recently shown PAC1 receptor involvement in vivo as an anti-inflammatory receptor, at least in part, by attenuating lipopolysaccharide-induced production of proinflammatory interleukin-6. The present study deepens in the protective role of PAC1 receptor in septic shock, elucidating its involvement in the modulation of neutrophil recruitment and in the expression of different molecular sensors such as intercellular adhesion molecule-1, vascular cell adhesion molecule-1, fibrinogen, serum amyloid A, and nitric oxide as important, systemic players of the development of septic shock. Our results, using a mice deficient in PAC1 and a PAC1 antagonist, show that VIP and PACAP as well as the PAC1 receptor are involved in neutrophil recruitment in different target organs, in adhesion molecules expression, and in coagulation-related molecule fibrinogen synthesis. Thus, this study provides some important insights with respect to the involvement of PAC1 into the complexities of sepsis and represents an advantage for the design of more specific drugs complementing standard intensive care therapy in severe sepsis, confirming VIP and PACAP as candidates for multitarget therapy of septic shock. PMID- 15661830 TI - Coactivator proteins as determinants of estrogen receptor structure and function: spectroscopic evidence for a novel coactivator-stabilized receptor conformation. AB - The direct regulation of gene transcription by nuclear receptors, such as the estrogen receptor (ER), involves not just ligand and DNA binding but the recruitment of coregulators. Typically, recruitment of p160 coactivator proteins to agonist-liganded ER is considered to be unidirectional, with ligand binding stabilizing an ER ligand binding domain (LBD) conformation that favors coactivator interaction. Using fluorophore-labeled ERalpha-LBDs, we present evidence for a pronounced stabilization of ER conformation that results from coactivator binding, manifest by decreased ER sensitivity to proteases and reduced conformational dynamics, as well as for the formation of a novel coactivator-stabilized (costabilized) receptor conformation, that can be conveniently monitored by the generation of an excimer emission from pyrene labeled ERalpha-LBDs. This costabilized conformation may embody features required to support ER transcriptional activity. Different classes of coactivator proteins combine with estrogen agonists of different structure to elicit varying degrees of this receptor stabilization, and antagonists and coactivator binding inhibitors disfavor the costabilized conformation. Remarkably, high concentrations of coactivators engender this conformation even in apo- and antagonist-bound ERs (more so with selective ER modulators than with pure antagonists), providing an in vitro model for the development of resistance to hormone therapy in breast cancer. PMID- 15661831 TI - Peroxisome proliferator-activated receptor alpha physically interacts with CCAAT/enhancer binding protein (C/EBPbeta) to inhibit C/EBPbeta-responsive alpha1 acid glycoprotein gene expression. AB - Recently, the role of the peroxisome proliferator-activated receptor alpha (PPARalpha) in the hepatic inflammatory response has been associated to the decrease of acute phase protein transcription, although the molecular mechanisms are still to be elucidated. Here, we were interested in the regulation by Wy 14643 (PPARalpha agonist) of alpha1-acid glycoprotein (AGP), a positive acute phase protein, after stimulation by Dexamethasone (Dex), a major modulator of the inflammatory response. In cultured rat hepatocytes, we demonstrate that PPARalpha inhibits at the transcriptional level the Dex-induced AGP gene expression. PPARalpha exerts this inhibitory effect by antagonizing the CCAAT/enhancer binding protein (C/EBPbeta) transcription factor that is involved in Dex dependent up-regulation of AGP gene expression. Overexpression of C/EBPbeta alleviates the repressive effect of PPARalpha, thus restoring the Dex-stimulated AGP promoter activity. Furthermore, glutathione-S-transferase GST pull-down and coimmunoprecipitation experiments evidenced, for the first time, a physical interaction between PPARalpha and the C-terminal DNA binding region of C/EBPbeta, thus preventing it from binding to specific sequence elements of the AGP promoter. Altogether, these results provide an additional molecular mechanism of negative regulation of acute phase protein gene expression by sequestration of the C/EBPbeta transcription factor by PPARalpha and reveal the high potency of the latter in controlling inflammation. PMID- 15661832 TI - Thyrocytes express a functional toll-like receptor 3: overexpression can be induced by viral infection and reversed by phenylmethimazole and is associated with Hashimoto's autoimmune thyroiditis. AB - Toll-like receptors (TLRs) initiate an innate immune response. TLR3 on dendritic cells recognize double-stranded (ds) RNA and then signal increases in cytokines and recognition molecules important for immune cell interactions. In this report, we demonstrate TLR3 mRNA and protein are expressed on Fisher rat thyroid cell line-5 (FRTL-5) thyroid cells and are functional because incubating cells with polyinosine-polycytidylic acid causes 1) transcriptional activation of both the nuclear factor kappaB (NF-kappaB)/Elk1 and interferon (IFN) regulatory factor 3/IFN-beta signal paths, 2) posttranscriptional activation of NF-kappaB and ERK1/2, and 3) increased IFN-beta mRNA. TLR3 can be overexpressed, along with dsRNA-dependent protein kinase, major histocompatibility complex-I or II, and IFN regulatory factor-1, by transfecting dsRNA into the cells, infection with Influenza A virus, or incubation with IFN-beta, but not by incubation with dsRNA or IFNgamma, or by dsDNA transfection. A methimazole (MMI) derivative, phenylmethimazole, to a significantly greater degree than MMI, prevents overexpression by inhibiting increased transcriptional activation of IRF-3 and of IFN-stimulated response elements, phosphorylation of signal transducers and activation of transcription (STAT-1), but not NF-kappaB activation. TLR3 can be functionally overexpressed in cultured human thyrocytes by dsRNA transfection or IFN-beta treatment. Immunohistochemical studies show that TLR3 protein is overexpressed in human thyrocytes surrounded by immune cells in 100% of patients with Hashimoto's thyroiditis examined, but not in normal or Graves' thyrocytes. We conclude that functional TLR3 are present on thyrocytes; TLR3 downstream signals can be overexpressed by pathogen-related stimuli; overexpression can be reversed by phenylmethimazole to a significantly greater extent than MMI by inhibiting only the IFN regulatory factor-3/IFN-beta/signal transducers and activation of transcription arm of the TLR3 signal system; and TLR3 overexpression can induce an innate immune response in thyrocytes, which may be important in the pathogenesis of Hashimoto's thyroiditis and in the immune cell infiltrates. PMID- 15661833 TI - Hypothalamic growth hormone-releasing hormone (GHRH) deficiency: targeted ablation of GHRH neurons in mice using a viral ion channel transgene. AB - Animal and clinical models of GHRH excess suggest that GHRH provides an important trophic drive to pituitary somatotrophs. We have adopted a novel approach to silence or ablate GHRH neurons, using a modified H37A variant of the influenza virus M2 protein ((H37A)M2). In mammalian cells, (H37A)M2 forms a high conductance monovalent cation channel that can be blocked by the antiviral drug rimantadine. Transgenic mice with (H37A)M2 expression targeted to GHRH neurons developed postweaning dwarfism with hypothalamic GHRH transcripts detectable by RT-PCR but not by in situ hybridization and immunocytochemistry, suggesting that expression of (H37A)M2 had silenced or ablated virtually all the GHRH cells. GHRH M2 mice showed marked anterior pituitary hypoplasia with GH deficiency, although GH cells were still present. GHRH-M2 mice were also deficient in prolactin but not TSH. Acute iv injections of GHRH in GHRH-M2 mice elicited a significant GH response, whereas injections of GHRP-6 did not. Twice daily injections of GHRH (100 microg/d) for 7 d in GHRH-M2 mice doubled their pituitary GH but not PRL contents. Rimantadine treatment failed to restore growth or pituitary GH contents. Our results show the importance of GHRH neurons for GH and prolactin production and normal growth. PMID- 15661835 TI - VEGF-A splice variants and related receptor expression in human skeletal muscle following submaximal exercise. AB - VEGF-A contributes to muscle tissue angiogenesis following aerobic exercise training. The temporal response of the VEGF-A isoforms and their target receptors has not been comprehensively profiled in human skeletal muscle. We combined submaximal exercise with and without reduced leg blood flow to establish whether ischemia-induced metabolic stress was an important physiological stimuli responsible for regulating the VEGF-A system in humans. Nine healthy men performed two 45-min bouts of one-leg knee-extension exercise, with and without blood flow restriction. Muscle biopsies were obtained at rest and 2 and 6 h after exercise. Expression (mRNA) of the VEGF-A splice variants and related receptors [VEGF receptor (VEGFR)-1, VEGFR-2, and neuropilin-1] was determined by using qPCR. VEGF-A(total) expression increased more robustly after exercise with reduced blood flow, and initially this principally reflected an increase in VEGF A(165). Six hours after exercise, there was a relatively greater increase in VEGF A(189), and this response was not influenced by blood flow conditions. VEGFR-1 mRNA expression increased 2 h after exercise, and neuropilin-1 expression was transiently reduced, while all three receptors increased by 6 h. There was no evidence for the expression of the inhibitory VEGF-A(165B) variant in human skeletal muscle. Our study, reflecting both VEGF-A ligand and receptors, implicates metabolic perturbation as a regulator of human muscle angiogenesis and demonstrates that VEGF-A splice variants are distinctly regulated. Our findings also indicate that all three receptor genes exhibit different pretranslational regulation, in response to exercise in humans. PMID- 15661834 TI - Novel gender-related regulation of CYP2C12 gene expression in rats. AB - The expression of CYP2C12 by GH occurs in female but not in male rat livers. Direct injection of the CYP2C12 promoter-luciferase gene into male rat livers showed that the CYP2C12 promoter was active in both male and female rats. Thus, to further examine one or more factors that regulate the gender-related expression of CYP2C12, male rats were treated with trichostatin A, a specific inhibitor of histone deacetylase capable of condensing the chromatin structure. Interestingly, the expression of CYP2C12 by GH was seen even in the livers of male rats, indicating that histone deacetylase contributes to the suppression of CYP2C12 expression in male rats. Deoxyribonuclease I hypersensitive assay using nuclei from the livers of male or female rats revealed that the chromatin structure of the CYP2C12 gene was gender specific: a hypersensitive site at a position -4.2 kb containing GH-responsive element that bound to signal transducer and activator of transcription 5 (STAT5), termed as HS (hypersensitive site) 1, was specific for female rat livers, whereas a hypersensitive site at a position 3 kb, designated as HSm (male-specific hypersensitive site), was characteristic of male rat livers. A -3425/-3275 region within HSm functioned as a negative regulatory region, when the region was inserted in front of simian virus 40 promoter. Gel shift assay demonstrated that both CCAAT/enhancer-binding protein alpha and beta bound to the -3425/-3275 region. Based on these results, we conclude that the gender-related expression of the CYP2C12 gene results from the inaccessibility of to STAT5 to the GH-responsive element by chromatin condensation seen in male rat livers, and from the presence of the male-specific HSm that acts as a silencer. PMID- 15661836 TI - Effect of body tilt on calf muscle performance and blood flow in humans. AB - To explore the effect of posture on muscle performance, we tested the effects of body tilt angle on the strength, endurance, and fatigue of, and blood flow into, the plantar flexors. Human subjects were fixed to a tilt table that could tilt them from the horizontal (0 degrees ) to upright (90 degrees ) position and enabled force to be applied to a footplate through isometric action of the right calf muscle. In experiment 1, six subjects performed a strength test and graded test (intermittent contractions) to the point of failure at three tilt angles (0, 47, and 90 degrees ). In Experiment 2, seven subjects performed a strength test and constant-force test [70% maximum force (F(max)); intermittent contractions] to the point of failure in the horizontal and three inclined positions (32, 47, and 67 degrees ). In experiment 3, leg blood flow was assessed during constant force exercise at two intensities (30 and 70% F(max)) and two tilt angles (0 and 67 degrees ) in six subjects. Strength was not affected (P > 0.05) by tilt angle. Time to failure during the graded test was significantly higher at 47 degrees (25.9 +/- 2.0 min) and 90 degrees (25.1 +/- 3.0 min) than 0 degrees (22.2 +/- 2.6 min). Time to failure during the constant-force test was also significantly higher at 32 degrees (7.1 +/- 3.6 min), 47 degrees (8.0 +/- 5.2 min), and 67 degrees (8.6 +/- 5.6 min) compared with 0 degrees (4.0 +/- 2.6 min). When graded or constant-force exercise was performed with arterial flow to the leg eliminated, there were no differences in exercise time between the horizontal and an inclined position. During nonischemic exercise, leg blood flow was significantly higher during exercise in the inclined position. These results demonstrate that head-up tilt improves endurance of the plantar flexors, that this effect occurs in the absence of an effect on strength, and that it depends on an intact peripheral circulation. Moreover, the postural effect on muscle endurance appears to be due to a greater blood flow into the leg, an effect that is established during the initial contractions. PMID- 15661837 TI - Influence of phosphatidylglycerol on the uptake of liposomes by alveolar cells and on lung function. AB - The effect of phosphatidylglycerol on the uptake of surfactant-like liposomes by alveolar type II cells and alveolar macrophages as well as the effect on endogenous surfactant function was studied in vivo. Healthy ventilated rats were intratracheally instilled with fluorescent labeled liposomes with different concentrations of phosphatidylglycerol. Lung function was determined by monitoring arterial oxygenation and, at the end of the experiment, by recording static pressure-volume curves. In addition, alveolar cells were isolated, and cell-associated fluorescence was determined using flow cytometry. The results show that, in the presence of cofactors (Ca(2+), Mg(2+)), phosphatidylglycerol stimulates the uptake by alveolar macrophages but hardly affects the uptake by alveolar type II cells. High concentrations of phosphatidylglycerol reduce the number of alveolar macrophages in the alveolar space and deteriorate lung function. On the other hand, the presence of cofactors protects the lung against the negative effects of phosphatidylglycerol on endogenous surfactant and alveolar macrophages. This study indicates that the phosphatidylglycerol concentration may play a fundamental role in the surfactant function and metabolism depending on the presence of so-called cofactors like calcium and magnesium; further study is needed to clarify the mechanisms involved. PMID- 15661838 TI - Genetic determinants on rat chromosome 6 modulate variation in the hypercapnic ventilatory response using consomic strains. AB - To understand the genetic basis of pathways involved in the control of breathing, a large scale, high-throughput study using chromosomal substitution strains of rats is underway. Eight new consomic rat stains (SS-2(BN), SS-4(BN), SS-6(BN), SS 7(BN), SS-8(BN), SS-11(BN), SS-12(BN), SS-14(BN), SS-Y(BN)), containing one homozygous BN/NHsdMcwi (BN) chromosome on a background of SS/JrHsdMcwi (SS), were created by PhysGen (http://pga.mcw.edu) Program for Genomic Applications. Male and female rats were studied using standard plethysmography under control conditions and during acute hypoxia (inspired oxygen fraction = 0.12) and hypercapnia (inspired CO(2) fraction = 0.07). The rats were also studied during treadmill exercise. Both male and female BN rats had a significantly lower ventilatory response during 7% CO(2) compared with SS rats of the same gender. SS 6(BN) female rats had a significantly reduced ventilatory response, similar to BN rats due primarily to a reduced tidal volume. Male SS-6(BN) rats had a significantly reduced tidal volume response to hypercapnia but a slightly increased frequency response during hypercapnia. Gene(s) on the Y chromosome may play a role in this increased frequency response in the male rats because the SS Y(BN) hypercapnic ventilatory response involves a significantly increased frequency response. Several chromosomal substitutions slightly altered the ventilatory responses to hypoxia and exercise. However, genes on chromosomes 6 and Y of those studied are of primary importance in aspects of ventilatory control currently studied. PMID- 15661839 TI - Characterization of calves exhibiting a novel inheritable TNF-alpha hyperresponsiveness to endotoxin: associations with increased pathophysiological complications. AB - A subpopulation of calves, herein termed "hyperresponders" (HPR), was identified and defined by the patterns of plasma TNF-alpha concentrations that developed following two challenges with endotoxin (LPS, 0.8 mug Escherichia coli 055:B5 LPS/kg(0.75) live body wt) separated by 5 days. The principle characteristic of HPR calves was a failure to develop tolerance to repeated LPS challenge that was evident in the magnitude of the TNF-alpha concentrations and prolonged severity of pathological sequellae. Whereas calves failing to develop LPS tolerance were identified on the basis of their excessive in vivo plasma TNF-alpha concentration responses, in vitro TNF-alpha responses of peripheral blood mononuclear cells isolated from each calf and challenged with LPS or PMA did not correlate or predict the magnitude of in vivo plasma TNF response of the calf. Intentional breeding to obtain calves from bulls and/or cows documented as HPR resulted in offspring displaying the HPR character when similar progeny calves were tested with LPS in vivo, with extensive controls in place to account for sources of variability in the general TNF-alpha response to LPS that might compromise interpretation of the data. Feed intake, clinical serology and hematology profiles, and acute-phase protein responses of HPR calves following LPS were significantly different from those of calves displaying tolerance. These results suggest that the pattern of plasma TNF-alpha changes that evolve from a low-level double LPS challenge effectively reveal the presence of a genetic potential for animals to display excessive or prolonged pathological response to LPS-related stress and compromised prognosis for recovery. PMID- 15661840 TI - Cutaneous vascular responses to isometric handgrip exercise during local heating and hyperthermia. AB - The dramatic increase in skin blood flow and sweating observed during heat stress is mediated by poorly understood sympathetic cholinergic mechanisms. One theory suggests that a single sympathetic cholinergic nerve mediates cutaneous active vasodilation (AVD) and sweating via cotransmission of separate neurotransmitters, because AVD and sweating track temporally and directionally when activated during passive whole body heat stress. It has also been suggested that these responses are regulated independently, because cutaneous vascular conductance (CVC) has been shown to decrease, whereas sweat rate increases, during combined hyperthermia and isometric handgrip exercise. We tested the hypothesis that CVC decreases during isometric handgrip exercise if skin blood flow is elevated using local heating to levels similar to that induced by pronounced hyperthermia but that this does not occur at lower levels of skin blood flow. Subjects performed isometric handgrip exercise as CVC was elevated at selected sites to varying levels by local heating (which is independent of AVD) in thermoneutral and hyperthermic conditions. During thermoneutral isometric handgrip exercise, CVC decreased at sites in which blood flow was significantly elevated before exercise (-6.5 +/- 1.8% of maximal CVC at 41 degrees C and -10.5 +/- 2.0% of maximal CVC at 43 degrees C; P < 0.05 vs. preexercise). During isometric handgrip exercise in the hyperthermic condition, an observed decrease in CVC was associated with the level of CVC before exercise. Taken together, these findings argue against withdrawal of AVD to explain the decrease in CVC observed during isometric handgrip exercise in hyperthermic conditions. PMID- 15661841 TI - Effect of glutamine on water and sodium absorption in human jejunum at baseline and during PGE1-induced secretion. AB - Glutamine, a major fuel for enterocytes, stimulates water and sodium absorption in animal models of secretory diarrhea, but data in humans are still limited. The aim of this study was to investigate the effect of glutamine on jejunal absorption during hypersecretion in humans. In six healthy adults, the effects of glutamine on jejunal absorption were assessed with a triple-lumen tube on two occasions, at baseline and during PGE(1)-induced hypersecretion (0.1 microg.kg( 1).min(-1)) in a random order. Isoosmolar solutions containing polyethylene glycol 4000 as nonabsorbable marker were infused in the jejunum at 10 ml/min over 1-h periods: saline (sodium chloride 308 mmol/l), glucose-mannitol 45:45 mM, glucose 90 mM, alanine-glucose 45:45 mM, glutamine-glucose 45:45 mM, and glutamine 90 mM. Net absorptive and secretory fluxes were measured at steady state. At baseline, glutamine- and alanine-containing solutions induced a threefold increase of water and sodium absorption (P < 0.05); 90 mM glutamine stimulated water absorption more than 90 mM glucose (3.6 +/- 0.6 vs. 1.9 +/- 0.3 ml.min(-1).30 cm(-1), P < 0.05). PGE(1)-induced hypersecretion was reduced (P < 0.05) by solutions of alanine-glucose, glutamine-glucose, and glutamine 90 mM (P < 0.05) and reversed to absorption by alanine-glucose and glutamine-glucose. Glutamine and alanine absorption was nearly complete and was not influenced by PGE(1). In conclusion, glutamine stimulates water and electrolyte absorption in human jejunum, even during experimental hypersecretion. In addition to the metabolic effects of glutamine, these results support the evaluation of glutamine containing solutions for the rehydration and the nutritional support of patients with secretory diarrhea. PMID- 15661842 TI - An evaluation of detection methods for large lariat RNAs. AB - Ty1 elements are long terminal repeat (LTR) retrotransposons that reside within the genome of Saccharomyces cerevisiae. It has been known for many years that the 2'-5' phosphodiesterase Dbr1p, which debranches intron lariats, is required for efficient Ty1 transposition. A recent report suggested the intriguing possibility that Ty1 RNA forms a lariat as a transposition intermediate. We set out to further investigate the nature of the proposed Ty1 lariat branchpoint. However, using a wide range of techniques we were unable to find any evidence for the proposed lariat structure. Furthermore, we demonstrate that some of the techniques used in the initial study describing the lariat are capable of incorrectly reporting a lariat structure. Thus, the role of the Dbr1 protein in Ty1 retrotransposition remains elusive. PMID- 15661843 TI - Mutational analysis of the DEAD-box RNA helicase eIF4AII characterizes its interaction with transformation suppressor Pdcd4 and eIF4GI. AB - Eukaryotic initiation factor (eIF) 4A unwinds secondary and tertiary structures in the 5'-untranslated region of mRNA, permitting translation initiation. Programmed cell death 4 (Pdcd4) is a novel transformation suppressor and eIF4A binding partner that inhibits eIF4A helicase activity and translation. To elucidate the regions of eIF4A that are functionally significant in binding to Pdcd4, we generated point mutations of eIF4A. Two-hybrid analysis revealed that five eIF4A mutants completely lost binding to Pdcd4 while four eIF4A mutants retained wild-type levels of binding. The residues that, when mutated, inactivated Pdcd4 binding specified ATP binding, ATP hydrolysis, or RNA binding. With the exception of the Q-motif mutant eIF4AP56L, the eIF4A mutants inactivated for Pdcd4 binding were inactivated for binding to eIF4G (GM, GC, or both) and for enhancing translation. Several eIF4A mutants showing wild-type level binding to Pdcd4 were also inactivated for binding to eIF4G and for enhancing translation. Thus, significant dissociation of eIF4A's Pdcd4- and eIF4G-binding regions appears to occur. Because three of the four eIF4A mutants that retained Pdcd4 binding also suppressed translation activity in a dominant-negative manner, the structure that defines the Pdcd4-binding domain of eIF4A may be necessary but is insufficient for translation. A structural homology model of eIF4A shows regions important for binding to Pdcd4 and/or eIF4G lying on the perimeters of the hinge area of eIF4A. A competition experiment revealed that Pdcd4 competes with C terminal eIF4G for binding to eIF4A. In summary, the Pdcd4-binding domains on eIF4A impact both binding to eIF4G and translation initiation in cells. PMID- 15661844 TI - Interaction of RRF and EF-G from E. coli and T. thermophilus with ribosomes from both origins--insight into the mechanism of the ribosome recycling step. AB - Ribosome recycling factor (RRF), elongation factor-G (EF-G), and ribosomes from Thermus thermophilus (tt-) and Escherichia coli (ec-) were used to study the disassembly mechanism of post-termination ribosomal complexes by these factors. With tt-RRF, ec-EF-G can release bound-tRNA from ec-model post-termination complexes. However, tt-RRF is not released by ec-EF-G from ec-ribosomes. This complex with tt-RRF and ec-ribosomes after the tRNA release by ec-EF-G is regarded as an intermediate of the disassembly reaction. Not only tt-RRF, but also mRNA, cannot be released from ec-ribosomes by tt-RRF and ec-EF-G. These data suggest that the release of RRF from ribosomes is coupled or closely related to the release of mRNA during disassembly of post-termination complexes. With tt ribosomes, ec-EF-G cannot release ribosome-bound ec-RRF even though they are from the same species, showing that proper interaction of ec-RRF and ec-EF-G does not occur on tt-ribosomes. On the other hand, in contrast to a published report, tt EF-G functions with ec-RRF to disassemble ec-post-termination complexes. In support of this finding, tt-EF-G translocates peptidyl tRNA on ec-ribosomes and catalyzes ec-ribosome-dependent GTPase, showing that tt-EF-G has in vitro translocation activity with ec-ribosomes. Since tt-EF-G with ec-RRF can release tRNA from ec-post-termination complexes, the data are consistent with the hypothesis that the release of tRNA by RRF and EF-G from post-termination complexes is a result of a translocation-like activity of EF-G on RRF. PMID- 15661845 TI - U2AF binding selects for the high conservation of the C. elegans 3' splice site. AB - Caenorhabditis elegans is unusual among animals in having a highly conserved octamer sequence at the 3' splice site: UUUU CAG/R. This sequence can bind to the essential heterodimeric splicing factor U2AF, with U2AF65 contacting the U tract and U2AF35 contacting the splice site itself (AG/R). Here we demonstrate a strong correspondence between binding to U2AF of RNA oligonucleotides with variant octamer sequences and the frequency with which such variations occur in splice sites. C. elegans U2AF has a strong preference for the octamer sequence and exerts much of the pressure for 3' splice sites to have the precise UUUUCAG/R sequence. At two positions the splice site has a very strong preference for U even though alternative bases can also bind tightly to U2AF, suggesting that evolution can select against sequences that may have a relatively modest reduction in binding. Although pyrimidines are frequently present at the first base in the exon, U2AF has a very strong bias against them, arguing there is a mechanism to compensate for weakened U2AF binding at this position. Finally, the C in the consensus sequence must remain adjacent to the AG/R rather than to the stretch of U's, suggesting this C is recognized by U2AF35. PMID- 15661846 TI - Conserved spacing between the box C/D and C'/D' RNPs of the archaeal box C/D sRNP complex is required for efficient 2'-O-methylation of target RNAs. AB - RNA-guided nucleotide modification complexes direct the post-transcriptional nucleotide modification of both archaeal and eukaryotic RNAs. We have previously demonstrated that efficient 2'-O-methylation activity guided by an in vitro reconstituted archaeal box C/D sRNP requires juxtaposed box C/D and C'/D' RNP complexes. In these experiments, we investigate the importance of spatially positioning the box C/D and C'/D' RNPs within the sRNP complex for nucleotide modification. Initial sequence analysis of 245 archaeal box C/D sRNAs from both Eukyarchaeota and Crenarchaeota kingdoms revealed highly conserved spacing between the box C/D and C'/D' RNA motifs. Distances between boxes C to D' and C' to D (D' and D spacers, respectively) exhibit highly constrained lengths of 12 nucleotides (nt). Methanocaldococcus jannaschii sR8 sRNA, a model box C/D sRNA with D and D' spacers of 12 nt, was mutated to alter the distance between the two RNA motifs. sRNAs with longer or shorter spacer regions could still form sRNPs by associating with box C/D core proteins, L7, Nop56/58, and fibrillarin, comparable to wild-type sR8. However, these reconstituted box C/D sRNP complexes were severely deficient in methylation activity. Alteration of the D and D' spacer lengths disrupted the guided methylation activity of both the box C/D and C'/D' RNP complexes. When only one spacer region was altered, methylation activity of the corresponding RNP was lost. Collectively, these results demonstrate the importance of box C/D and C'/D' RNP positioning for preservation of critical inter-RNP interactions required for efficient box C/D sRNP-guided nucleotide methylation. PMID- 15661847 TI - Small molecule control of pre-mRNA splicing. AB - SR proteins regulate alternative splicing by binding to exonic sequences where, via an arginine/serine-rich splicing activation domain, they enhance the binding of the spliceosome to the adjacent splice sites. Here, a system is described in which a nontoxic derivative of the small molecule rapamycin is used to control pre-mRNA splicing in vitro. This involves the rapamycin-dependent recruitment of a splicing activation domain located on one protein to a second protein bound to the pre-mRNA. These results provide a new approach to explore for regulating gene expression in vivo with small molecules by controlling pre-mRNA splicing. PMID- 15661848 TI - Identification and characterization of critical cis-acting sequences within the yeast Ty1 retrotransposon. AB - The yeast long terminal repeat (LTR) retrotransposon Ty1, like retroviruses, encodes a terminally redundant RNA, which is packaged into virus-like particles (VLPs) and is converted to a DNA copy by the process of reverse transcription. Mutations predicted to interfere with the priming events during reverse transcription and hence inhibit replication are known to dramatically decrease transposition of Ty1. However, additional cis-acting sequences responsible for Ty1 replication and RNA dimerization and packaging have remained elusive. Here we describe a modular mini-Ty1 element encoding the minimal sequence that can be retrotransposed by the Ty1 proteins, supplied in trans by a helper construct. Using a mutagenic screening strategy, we recovered transposition-deficient modular mini-Ty1-HIS3 elements with mutations in sequences required in cis for Ty1 replication and integration. Two distinct clusters of mutations mapped near the 5'-end of the Ty1 RNA. The clusters define a GAGGAGA sequence at the extreme 5'-end of the Ty1 transcript and a complementary downstream UCUCCUC sequence, 264 nt into the RNA. Disruption of the reverse complementarity of these two sequences decreased transposition and restoration of complementarity rescued transposition to wild-type levels. Ty1 cDNA was reduced in cells expressing RNAs with mutations in either of these short sequences, despite nearly normal levels of Ty1 RNA and VLPs. Our results suggest that the intramolecular interaction between the 5' GAGGAGA and UCUCCUC sequences stabilizes an RNA structure required for efficient initiation of reverse transcription. PMID- 15661849 TI - Silencing of polo-like kinase (Plk) 1 via siRNA causes induction of apoptosis and impairment of mitosis machinery in human prostate cancer cells: implications for the treatment of prostate cancer. AB - Prostate cancer (PCa) is one of the most common cancers in men. Each year approximately 543,000 new cases are reported worldwide, and the disease kills 200,000 (mostly older men) in developed countries. The existing treatment approaches and surgical intervention have not been able to effectively manage this dreaded cancer and, therefore, continuing efforts are ongoing to explore novel targets and strategies for the management of PCa. The activity of polo-like kinase 1 (Plk1) is elevated in tissues and cells with a high mitotic index, including cancer cells. An increasing body of evidence suggests that the level of Plk1 expression has prognostic value for predicting outcomes in patients with some cancers. A close correlation between Plk1 expression and carcinogenesis has been documented. However, the role of Plk1 in PCa is not known. We propagated a hypothesis that Plk1 inhibition will result in elimination of human PCa cells via a mitotic arrest followed by apoptosis (1). To define the role of Plk1 in PCa, we used the technique of RNA silencing via small interfering RNA (siRNA). First, using a series of human prostate carcinoma cells and normal human prostate epithelial (PrEC) cells, we assessed Plk1 levels in PCa. Immunoblot analyses clearly showed a significant expression of Plk1 in LNCaP, DU145, and PC3 human PCa cells. Interestingly, Plk1 was not detectable in normal PrEC cells. Next, we transfected the PCa cells with Plk 1 siRNA, which resulted in a significant inhibition in Plk1 protein in all PCa cells. Plk1 depletion resulted in a decrease in cell viability and induction of apoptosis in PCa cells but had no appreciable effect in normal PrEC cells. Our data also demonstrated that Plk1 siRNA transfection of PCa cells resulted in 1) a mitotic cell cycle arrest, 2) failure of cytokinesis, and 3) defects in centrosome integrity and maturation. Thus, our study suggested that 1) Plk1 plays a critical role in the process of PCa development and 2) gene therapeutic approaches aimed at Plk1 or the pharmacological inhibitors of Plk1 may be developed for the management of PCa. PMID- 15661852 TI - Mandatory submission of microarray data to public repositories: how is it working? PMID- 15661851 TI - MAFFT version 5: improvement in accuracy of multiple sequence alignment. AB - The accuracy of multiple sequence alignment program MAFFT has been improved. The new version (5.3) of MAFFT offers new iterative refinement options, H-INS-i, F INS-i and G-INS-i, in which pairwise alignment information are incorporated into objective function. These new options of MAFFT showed higher accuracy than currently available methods including TCoffee version 2 and CLUSTAL W in benchmark tests consisting of alignments of >50 sequences. Like the previously available options, the new options of MAFFT can handle hundreds of sequences on a standard desktop computer. We also examined the effect of the number of homologues included in an alignment. For a multiple alignment consisting of approximately 8 sequences with low similarity, the accuracy was improved (2-10 percentage points) when the sequences were aligned together with dozens of their close homologues (E-value < 10(-5)-10(-20)) collected from a database. Such improvement was generally observed for most methods, but remarkably large for the new options of MAFFT proposed here. Thus, we made a Ruby script, mafftE.rb, which aligns the input sequences together with their close homologues collected from SwissProt using NCBI-BLAST. PMID- 15661850 TI - DNA analysis with multiplex microarray-enhanced PCR. AB - We have developed a highly sensitive method for DNA analysis on 3D gel element microarrays, a technique we call multiplex microarray-enhanced PCR (MME-PCR). Two amplification strategies are carried out simultaneously in the reaction chamber: on or within gel elements, and in bulk solution over the gel element array. MME PCR is initiated by multiple complex primers containing gene-specific, forward and reverse, sequences appended to the 3' end of a universal amplification primer. The complex primer pair is covalently tethered through its 5' end to the polyacryl- amide backbone. In the bulk solution above the gel element array, a single pair of unattached universal primers simultaneously directs pseudo monoplex PCR of all targets according to normal solution-phase PCR. The presence of a single universal PCR primer pair in solution accelerates amplification within gel elements and eliminates the problem of primer interference that is common to conventional multiplex PCR. We show 10(6)-fold amplification of targeted DNA after 50 cycles with average amplification efficiency 1.34 per cycle, and demonstrate specific on-chip amplification of six genes in Bacillus subtilis. All six genes were detected at 4.5 pg of bacterial genomic DNA (equivalent to 10(3) genomes) in 60 independent amplification reactions performed simultaneously in single reaction chamber. PMID- 15661853 TI - Cyclin G1 and cyclin G2 are expressed in the periimplantation mouse uterus in a cell-specific and progesterone-dependent manner: evidence for aberrant regulation with Hoxa-10 deficiency. AB - Because uterine cell-specific proliferation, differentiation, and apoptosis are differentially regulated during the periimplantation period, we speculated that negative cell cycle regulators are also operative in the uterus during this period. This prompted us to examine the roles of two negative growth-regulatory genes, cyclin G1 and cyclin G2, in the periimplantation mouse uterus. We show that cyclin G1 and cyclin G2 genes are differentially regulated in the uterus during this period (d 1-8 of pregnancy) in a spatiotemporal manner. The results suggest that cyclin G1 is primarily associated with epithelial cell differentiation before implantation and stromal cell proliferation and differentiation during decidualization, whereas cyclin G2 is associated with terminal differentiation and apoptosis of the luminal epithelial and stromal cells at the site of blastocyst after implantation. Pharmacological and genetic studies provide evidence that the expression of cyclin G1, not cyclin G2, is regulated by progesterone via its nuclear receptor. Furthermore, the expression of these genes is aberrantly up-regulated in homeo box A-10 mutant uteri, suggesting that cyclin G1 and cyclin G2 genes act as downstream targets of homeobox A-10 and negatively impact uterine cell proliferation. Collectively, our present and previous studies suggest that negative cell cycle regulators collaborate with growth-promoting regulators in regulating uterine cell-specific proliferation, differentiation, and apoptosis relevant to implantation and decidualization. PMID- 15661854 TI - Insulin-like growth factor-I gene transfer by electroporation prevents skeletal muscle atrophy in glucocorticoid-treated rats. AB - Catabolic states caused by injury are characterized by a loss of skeletal muscle. The anabolic action of IGF-I on muscle and the reduction of its muscle content in response to injury suggest that restoration of muscle IGF-I content might prevent skeletal muscle loss caused by injury. We investigated whether local overexpression of IGF-I protein by gene transfer could prevent skeletal muscle atrophy induced by glucocorticoids, a crucial mediator of muscle atrophy in catabolic states. Localized overexpression of IGF-I in tibialis anterior (TA) muscle was performed by injection of IGF-I cDNA followed by electroporation 3 d before starting dexamethasone injections (0.1 mg/kg.d sc). A control plasmid was electroporated in the contralateral TA muscle. Dexamethasone induced atrophy of the TA muscle as illustrated by reduction in muscle mass (403 +/- 11 vs. 461 +/- 19 mg, P < 0.05) and fiber cross-sectional area (1759 +/- 131 vs. 2517 +/- 93 mum(2), P < 0.05). This muscle atrophy was paralleled by a decrease in the IGF-I muscle content (7.2 +/- 0.9 vs. 15.7 +/- 1.4 ng/g of muscle, P < 0.001). As the result of IGF-I gene transfer, the IGF-I muscle content increased 2-fold (15.8 +/ 1.2 vs. 7.2 +/- 0.9 ng/g of muscle, P < 0.001). In addition, the muscle mass (437 +/- 8 vs. 403 +/- 11 mg, P < 0.01) and the fiber cross-sectional area (2269 +/- 129 vs. 1759 +/- 131 mum(2), P < 0.05) were increased in the TA muscle electroporated with IGF-I DNA, compared with the contralateral muscle electroporated with a control plasmid. Our results show therefore that IGF-I gene transfer by electroporation prevents muscle atrophy in glucocorticoid-treated rats. Our observation supports the important role of decreased muscle IGF-I in the muscle atrophy caused by glucocorticoids. PMID- 15661855 TI - Endogenous estrogens inhibit mouse fetal Leydig cell development via estrogen receptor alpha. AB - It is now accepted that estrogens play a role in male fertility and that exposure to exogenous estrogens during fetal/neonatal life can lead to reproductive disorders in the male. However, the estrogen receptor (ER)-mediated processes involved in the regulation of male reproduction during fetal and neonatal development are still largely unclear. We previously reported that ER beta deficiency affects gametogenesis in mice but changes neither the number nor the differentiated functions of fetal Leydig cells. We show here that ER alpha deficient mice (ER alpha-/-) display higher levels of testicular testosterone secretion than wild-type mice from fetal d 13.5 onwards. This results from higher levels of steroidogenic activity per fetal Leydig cell, as indicated by the hypertrophy of these cells and the higher levels of mRNA for StAR, P450c17 and P450scc in the testis, for a similar number of Leydig cells. Because LH is not produced on fetal d 13.5 and because no change in plasma LH concentration was observed in 2-d-old ER alpha-deficient mice, LH is probably not involved in the effects of estrogens on testicular steroidogenesis in fetal and early neonatal Leydig cells. Furthermore, inactivation of ER beta did not change the effect of ER alpha inactivation on steroidogenesis. Lastly, in an organ culture system, 1 mum diethylstilbestrol decreased the testosterone secretion of wild-type fetal and neonatal testes but not of ER alpha-/- testes. Thus, this study shows that endogenous estrogens physiologically inhibit steroidogenesis via ER alpha by acting directly on the testis early in fetal and neonatal development. PMID- 15661856 TI - A molecular cascade showing nitric oxide-heme oxygenase-1-vascular endothelial growth factor-interleukin-8 sequence in human endothelial cells. AB - Heme oxygenase (HO)-1 has been shown to be an important biological target of nitric oxide (NO). NO can induce HO-1 expression and IL-8 production, particularly, in endothelial cells. Interestingly, HO-1 tends to induce the production of vascular endothelial growth factor (VEGF) that is involved in endothelial IL-8 syntheses. Whether HO-1 expression by NO may provide a link with IL-8 or VEGF synthesis was investigated in human umbilical vein endothelial cells (HUVECs). The NO donor S-nitroso-N-acetyl-penicillamine (SNAP) dose-dependently increased IL-8 and VEGF productions and HO-1 expression in HUVECs. Transfection with either HO-1 small interfering RNA or HO-1 antisense oligodeoxynucleotide abrogated the ability of SNAP to induce HO-1 expression and IL-8 and VEGF productions. Both pharmacological induction and gene transfer of HO-1 directly induced IL-8 and VEGF productions. Anti-VEGF neutralizing antibody blocked SNAP mediated IL-8 production and VEGF itself induced IL-8 production, whereas anti-IL 8 neutralizing antibody had no effect on VEGF production in SNAP-treated HUVECs. Neither anti-VEGF nor anti-IL-8 antibodies influenced SNAP-induced HO-1 expression. Moreover, neither VEGF nor IL-8 showed an additive effect on SNAP induced HO-1 expression. HO-1 transfection had no significant effect on productions of other CXC chemokines, such as growth-related oncogen-alpha and epithelial neutrophil activation peptide-78. Taken together, these results provide a molecular cascade showing NO-HO-1-VEGF-IL-8 sequence in human endothelial cells. PMID- 15661858 TI - A dominant negative human peroxisome proliferator-activated receptor (PPAR){alpha} is a constitutive transcriptional corepressor and inhibits signaling through all PPAR isoforms. AB - Several missense mutations in the ligand-binding domain of human peroxisome proliferator-activated receptor (PPAR)gamma have been described in subjects with dominantly inherited severe insulin resistance associated with partial lipodystrophy, hypertension, and dyslipidemia. These mutant receptors behave as dominant-negative inhibitors of PPARgamma signaling when studied in transfected cells. The extent to which such dominant-negative effects extend to signaling through other coexpressed PPAR isoforms has not been evaluated. To examine these issues further, we have created a PPARalpha mutant harboring twin substitutions, Leu459Ala and Glu462Ala, within the ligand binding domain (PPARalpha(mut)), examined its signaling properties, and compared the effects of dominant-negative PPARalpha and PPARgamma mutants on basal and ligand-induced gene transcription in adipocytes and hepatocytes. PPARalpha(mut) was transcriptionally inactive, repressed basal activity from a PPAR response element-containing promoter, inhibited the coactivator function of cotransfected PPAR-gamma coactivator 1alpha, and strongly inhibited the transcriptional response to cotransfected wild type receptor. In contrast to PPARgamma, wild-type PPARalpha failed to recruit the transcriptional corepressors NCoR and SMRT. However, PPARalpha(mut) avidly recruited these corepressors in a ligand-dissociable manner. In hepatocytes and adipocytes, both PPARalpha(mut) and the corresponding PPARgamma mutant were capable of inhibiting the expression of genes primarily regulated by PPARalpha, gamma, or -delta ligands, albeit with some differences in potency. Thus, dominant negative forms of PPARalpha and PPARgamma are capable of interfering with PPAR signaling in a manner that is not wholly restricted to their cognate target genes. These findings may have implications for the pathogenesis of human syndromes resulting from mutations in this family of transcription factors. PMID- 15661857 TI - Oxytocin induces dephosphorylation of eukaryotic elongation factor 2 in human myometrial cells. AB - The oxytocin (OT) receptor (OTR) mediates a wide spectrum of biological actions and is expressed in a large number of different tissues, including uterine, breast, and lung tumors. To define more completely the intracellular signaling mechanisms linked to OTR activation, we have used a phosphoproteomics approach and have characterized changes in the phosphorylation states of intracellular proteins in response to OTR activation in OTR-expressing cell lines. Using a specific antiphosphothreonine antibody, we observed several distinct changes in the threonine phosphorylation patterns. The most prominent change involved dephosphorylation of a 95-kDa moiety. Purification by ion exchange chromatography combined with one- and two-dimensional polyacrylamide gel electrophoresis followed by N-terminal micro-sequence analysis revealed that the 95-kDa moiety corresponded to eukaryotic elongation factor 2. This protein is a key regulator of cellular protein synthesis and mediates, upon dephosphorylation, the translocation step of peptide chain elongation. Dose-response curves in myometrial cells expressing the endogenous OTR indicated a significant effect of OT on eukaryotic elongation factor 2 dephosphorylation at 1 nM, a concentration close to the dissociation constant (K(d)) of OT. Time course analysis indicates that the effect is rapid with a significant effect occurring at 5 min. To determine directly the effect of OT on protein synthesis, the incorporation of [35S]Met into total protein was assessed. In myometrial cells, OTR activation led to significant 29% increase in total protein synthesis over a 2-h period. These findings establish a novel link between OTR activation and cellular protein synthesis and thus define a mechanism by which OT assumes a so far unrecognized, physiologically relevant trophic function. PMID- 15661859 TI - Endofacial competitive inhibition of glucose transporter-4 intrinsic activity by the mitogen-activated protein kinase inhibitor SB203580. AB - The translocation of glucose transporter-4 (GLUT4) to the cell surface is a complex multistep process that involves movement of GLUT4 vesicles from a reservoir compartment, and docking and fusion of the vesicles with the plasma membrane. It has recently been proposed that a p38 mitogen-activated protein kinase (MAPK)-dependent step may lead to intrinsic activation of the transporters exposed at the cell surface. In contrast to data obtained in muscle and adipocyte cell lines, we found that no insulin activation of p38 MAPK occurred in rat adipose cells. However, the p38 MAPK inhibitor SB203580 consistently inhibited transport activity after preincubation with the adipose cells. These apparently contradictory findings led us to hypothesize that the inhibitor may have a direct effect on the transport catalytic activity of GLUT4 that was independent of inhibition of the kinase. Kinetic analysis of 3-O-methyl-d-glucose transport activity revealed that SB203580 was a noncompetitive inhibitor of zero-trans (substrate outside but not inside) transport, but was a competitive inhibitor of equilibrium-exchange (substrate inside and outside) transport. This pattern of inhibition of GLUT4 was also observed with cytochalasin B. The pattern of inhibition is consistent with interaction at the endofacial surface, but not the exofacial surface of the transporter. Occupation of the endofacial substrate site reduces maximum velocity under zero-trans conditions, because return of the substrate site to the outside is blocked, and no substrate is present inside to displace the inhibitor. Under equilibrium-exchange conditions, internal substrate competitively displaces the inhibitor, and the transport K(m) is increased. PMID- 15661860 TI - Kainate/estrogen receptor involvement in rapid estradiol effects in vitro and intracellular signaling pathways. AB - Although the interactions between sex steroids and GnRH have been extensively studied, little is known about the mechanism of estradiol (E2) effects on GnRH secretion. In the present study, we used retrochiasmatic hypothalamic explants of 50-d-old male rats, and we observed that E2 significantly increased the glutamate evoked GnRH secretion in vitro within 15 min in a dose-dependent manner. E2 also significantly increased the L-arginine-evoked GnRH secretion. E2 effects were time dependent because the initially ineffective 10(-9) M concentration became effective after 5 h of incubation. The E2 effects involved the estrogen receptor (ER) alpha because they were similarly obtained with the specific ER alpha agonist 1,3,5-tris(4-hydroxyphenyl)-4-propyl-1H-pyrazole. The use of glutamate receptor agonists and antagonists indicated that E2 effects on GnRH secretion evoked by both glutamate and L-arginine involved the 2-amino-3-hydroxy-5-methyl-4 isoxazol propionic acid/kainate receptors. Similar E2 effects on the kainate evoked secretion were observed throughout development in both sexes. The observation of similar E2 effects using explants containing the median eminence alone indicated that the median eminence was a direct target for E2 rapid effects on the glutamate-evoked GnRH secretion. The signaling pathways involved in E2 effects included an increase in intracellular calcium and the activation of protein kinase A, protein kinase C, and MAPK. It is concluded that E2 can stimulate the glutamate- and nitric oxide-evoked GnRH secretion in vitro through a rapid pathway involving the ER and kainate receptor as well as through a slower mechanism responding to lower E2 concentrations. PMID- 15661862 TI - The monocarboxylate transporter 8 linked to human psychomotor retardation is highly expressed in thyroid hormone-sensitive neuron populations. AB - Recent genetic analysis in several patients presenting a severe form of X-linked psychomotor retardation combined with abnormal thyroid hormone (TH) levels have revealed mutations or deletions in the gene of the monocarboxylate transporter 8 (MCT8). Because in vitro MCT8 functions as a TH transporter, the complex clinical picture of these patients indicated an important role for MCT8 in TH-dependent processes of brain development. To provide a clue to the cellular function of MCT8 in brain, we studied the expression of MCT8 mRNA in the murine central nervous system by in situ hybridization histochemistry. In addition to the choroid plexus structures, the highest transcript levels were found in neo- and allocortical regions (e.g. olfactory bulb, cerebral cortex, hippocampus, and amygdala), moderate signal intensities in striatum and cerebellum, and low levels in a few neuroendocrine nuclei. Colocalization studies revealed that MCT8 is predominantly expressed in neurons. Together with the spatiotemporal expression pattern of MCT8 during the perinatal period, these results strongly indicate that MCT8 plays an important role for proper central nervous system development by transporting TH into neurons as its main target cells. PMID- 15661861 TI - Androgens stimulate human vascular smooth muscle cell proteoglycan biosynthesis and increase lipoprotein binding. AB - Vascular smooth muscle cell (VSMC) proliferation and proteoglycan biosynthesis are two critical contributors to the development of atherosclerosis. We investigated the effects of specific androgens, androstenedione, dihydrotestosterone, and testosterone, on proteoglycan biosynthesis in human VSMC derived from internal mammary arteries. Vascular SMCs were metabolically labeled with [(35)S]sulfate or [(35)S]methionine/cysteine to assess glycosaminoglycans (GAGs) or proteoglycan core protein, respectively. The electrophoretic migration of radiolabeled proteoglycans was assessed by SDS-PAGE. Proteoglycan-low density lipoprotein (LDL) interactions were assessed using LDL affinity columns. Treatment of VSMCs with androstenedione (100 nm), dihydrotestosterone (10 nm), or testosterone (100 nm) increased [(35)S]sulfate incorporation into GAGs by 24.8% (P < 0.05), 22% (P < 0.05), and 32.5% (P < 0.05), respectively. Treatment of VSMCs with testosterone did not alter [(35)S]methionine/cysteine incorporation into proteoglycan core protein, suggesting that the effect of testosterone was associated with an increase in GAG length. Dihydrotestosterone (10 nm) and testosterone (100 nm) treatment of VSMCs resulted in the synthesis of biglycan and decorin that showed reduced electrophoretic mobility by SDS-PAGE, indicating an increase in GAG length. The effect of testosterone treatment on [(35)S]sulfate incorporation and GAG length was reversed by pretreatment of VSMCs with flutamide (1 mum), an androgen receptor antagonist. Proteoglycans from VSMCs treated with testosterone showed 11% (P < 0.01) higher binding capacity to LDL compared with proteoglycans from untreated cells. These results suggest a possible proatherogenic action of androgens through an elongation of GAG chains on proteoglycans in an androgen receptor-dependent manner. PMID- 15661863 TI - Differential estrogen receptor expression in autoimmune myasthenia gravis. AB - Myasthenia gravis (MG) is an autoimmune disease associated with thymic hyperplasia and is much more prevalent in women than men. In this study we investigated potential changes in estrogen receptor (ER) expression in thymic hyperplasia. We first quantified by real-time PCR the relative expression of ER alpha and ER beta in normal thymus and found that the ER beta to ER alpha ratio was inverted in thymocytes (8.6 +/- 1.2), compared with thymic epithelial cells (0.18 +/- 0.05). The ER transcript number gradually decreased in thymic epithelial cells during culture, indicating that the thymic environment influences ER expression. CD4+ helper T cells expressed higher level of ERs, compared with CD8+ cells, as assessed by flow cytometry in thymocytes and peripheral blood mononuclear cells. In MG patients, we found an increased expression of ER alpha on thymocytes and both ERs on T cells from peripheral blood mononuclear cells, indicating that the signals provided by thymic and peripheral microenvironments are distinct. Finally, activation of normal thymocytes by proinflammatory cytokines induced increased expression of ERs especially in the CD4+ subset, suggesting that an excess of proinflammatory cytokines could explain the increase of ERs expression on MG lymphocytes. The dysregulation of ER expression in MG lymphocytes could affect the maintenance of the homeostatic conditions and might influence the progression of the autoimmune response. PMID- 15661864 TI - Dihydrotestosterone increases hippocampal N-methyl-D-aspartate binding but does not affect choline acetyltransferase cell number in the forebrain or choline transporter levels in the CA1 region of adult male rats. AB - Testosterone, acting through its androgenic metabolite 5alpha-dihydrotestosterone (DHT), can increase dendritic spine density in the CA1 region of the male rat hippocampus. The mechanisms mediating this increase in spines are presently unknown. In female rats, estrogen (E) has been shown to increase spine density, which is in part mediated by increases in N-methyl-d-aspartate (NMDA) receptors in the CA1 region and cholinergic forebrain inputs to the hippocampus. Whether similar mechanisms are responsible for the DHT-induced increase in spines in the male remains to be determined. In the first experiment, we used [(3)H]glutamate NMDA receptor binding autoradiography to assess whether DHT-treated males had higher NMDA receptor levels in the CA1 region of the hippocampus, compared with oil-treated males. In the second set of experiments, we used choline acetyltransferase (ChAT) in situ hybridization and immunohistochemistry to assess whether DHT could affect ChAT cell number in the forebrain. We also investigated the effect of DHT on hemicholinium-3-sensitive choline transporter levels in the CA1 region of the male hippocampus. We found that DHT significantly increased NMDA receptor binding in the CA1 region of males but had no effect on ChAT cell number in the forebrain or hemicholinium-3-sensitive choline transporter protein levels in the CA1 region. These data indicate that, similar to E-induced spinogenesis in females, DHT-induced increases in spine formation in males may require increases in NMDA receptors. However, unlike E-treated females, these data suggest that DHT does not influence cholinergic inputs to the hippocampus. PMID- 15661865 TI - First sighting of the elusive T cell antigen receptor. PMID- 15661866 TI - Tumor-specific antigen of murine T-lymphoma defined with monoclonal antibody. 1982. PMID- 15661867 TI - Sphingolipids and the balancing of immune cell function: lessons from the mast cell. AB - Recent studies reveal that metabolites of sphingomyelin are critically important for initiation and maintenance of diverse aspects of immune cell activation and function. The conversion of sphingomyelin to ceramide, sphingosine, or sphingosine-1-phosphate (S1P) provides interconvertible metabolites with distinct biological activities. Whereas ceramide and sphingosine function to induce apoptosis and to dampen mast cell responsiveness, S1P functions as a chemoattractant and can up-regulate some effector responses. Many of the S1P effects are mediated through S1P receptor family members (S1P(1-5)). S1P(1), which is required for thymocyte emigration and lymphocyte recirculation, is also essential for Ag-induced mast cell chemotaxis, whereas S1P(2) is important for mast cell degranulation. S1P is released to the extracellular milieu by Ag stimulated mast cells, enhancing inflammatory cell functions. Modulation of S1P receptor expression profiles, and of enzymes involved in sphingolipid metabolism, particularly sphingosine kinases, are key in balancing mast cell and immune cell responses. Current efforts are unraveling the complex underlying mechanisms regulating the sphingolipid pathway. Pharmacological intervention of these key processes may hold promise for controlling unwanted immune responses. PMID- 15661868 TI - Cutting edge: TLR ligands are not sufficient to break cross-tolerance to self antigens. AB - Cross-presentation of peripheral self-Ags by dendritic cells (DC) can induce deletion of autoreactive CTL by a mechanism termed cross-tolerance. Activation of DC by microbial TLR ligands is thought to result in adaptive immunity. However, activation of tolerogenic DC may cause autoimmunity by stimulating instead of deleting autoreactive CTL. To investigate this scenario, we have monitored the response of autoreactive CTL in specific for the transgenic self Ag, OVA, expressed in pancreatic islets of RIP-mOVA mice injected with ligands of TLR2, 3, 4, and 9. This somewhat enhanced proliferation and cytokine production, and moderately reduced the CTL number able to induce autoimmunity. Nevertheless, physiological CTL numbers were deleted before disease ensued, unless specific CD4 T cell help was provided. In conclusion, DC activation by TLR ligands was insufficient to break peripheral cross-tolerance in the absence of specific CD4 T cell help, and triggered autoimmunity by stimulating the early effector phase of autoreactive CTL only when their precursor frequency was extremely high. PMID- 15661869 TI - Impaired recall of CD8 memory T cells in immunologically privileged tissue. AB - Foreign Ags that enter immunologically privileged sites such as the eye, brain, and testis persist for an extended period of time, whereas the same Ags are rapidly eliminated at conventional sites. Immune privilege, therefore, provides unwanted refuge for pathogens and tumor cells but is beneficial for the survival of allogeneic grafts. In this study, we asked whether memory T cells can eliminate foreign Ags deposited at an immunologically privileged site by studying CD8 memory T cell-mediated rejection of pancreatic islet allografts placed either in the testis (a privileged organ) or under the kidney capsule (a nonprivileged site) of diabetic mice. We found that CD8 memory T cells reject intratesticular grafts at a significantly slower rate than the rejection of intrarenal grafts. Delayed graft rejection in the testis was not due to reduced homing or proliferation of memory T cells but due to their increased apoptosis at that site. Apoptosis was mediated by the combined actions of two TNFR family members that are up-regulated on activated memory T cells, Fas, and CD30. Therefore, memory T cells survey immunologically privileged tissues but are subject to the immunosuppressive mechanisms present at these sites. PMID- 15661870 TI - RNase L and double-stranded RNA-dependent protein kinase exert complementary roles in islet cell defense during coxsackievirus infection. AB - Coxsackievirus (CV) is an important human pathogen that has been linked to the development of autoimmunity. An intact pancreatic beta cell IFN response is critical for islet cell survival and protection from type 1 diabetes following CV infection. In this study, we show that IFNs trigger an antiviral state in beta cells by inducing the expression of proteins involved in intracellular antiviral defense. Specifically, we demonstrate that 2',5'-oligoadenylate synthetases (2 5AS), RNase L, and dsRNA-dependent protein kinase (PKR) are expressed by pancreatic islet cells and that IFNs (IFN-alpha and IFN-gamma) increase the expression of 2-5AS and PKR, but not RNase L. Moreover, our in vitro studies uncovered that these pathways play important roles in providing unique and complementary antiviral activities that critically regulate the outcome of CV infection. The 2-5AS/RNase L pathway was critical for IFN-alpha-mediated islet cell resistance from CV serotype B4 (CVB4) infection and replication, whereas an intact PKR pathway was required for efficient IFN-gamma-mediated repression of CVB4 infection and replication. Finally, we show that the 2-5AS/RNase L and the PKR pathways play important roles for host survival during a challenge with CVB4. In conclusion, this study has dissected the pathways used by distinct antiviral signals and linked their expression to defense against CVB4. PMID- 15661871 TI - Different competitive capacities of Stat4- and Stat6-deficient CD4+ T cells during lymphophenia-driven proliferation. AB - The outcome of an immune response relies on the competitive capacities acquired through differentiation of CD4(+) T cells into Th1 or Th2 effector cells. Because Stat4 and Stat6 proteins are implicated in the Th1 vs Th2 generation and maintenance, respectively, we compare in this study the kinetics of Stat4(-/-) and Stat6(-/-) CD4(+) T cells during competitive bone marrow reconstitution and lymphopenia-driven proliferation. After bone marrow transplantation, both populations reconstitute the peripheral T cell pools equally well. After transfer into lymphopenic hosts, wild-type and Stat6(-/-) CD4(+) T cells show a proliferation advantage, which is early associated with the expression of an active phospho-Stat4 and the down-regulation of Stat6. Despite these differences, Stat4- and Stat6-deficient T cells reach similar steady state numbers. However, when both Stat4(-/-) and Stat6(-/-) CD4(+) T cells are coinjected into the same hosts, the Stat6(-/-) cells become dominant and out-compete Stat4(-/-) cells. These findings suggest that cell activation, through the Stat4 pathway and the down-regulation of Stat6, confers to pro-Th1 T cells a slight proliferation advantage that in a competitive situation has major late repercussions, because it modifies the final homeostatic equilibrium of the populations and favors the establishment of Th1 CD4(+) T cell dominance. PMID- 15661872 TI - Polarized development of memory cell-like IFN-gamma-producing cells in the absence of TCR zeta-chain. AB - TCR/CD3 complex-mediated signals play critical roles in regulating CD4(+) Th cell differentiation. In this report, we have examined the in vivo role of a key TCR/CD3 complex molecule zeta-chain in regulating the differentiation of Th cells. We have studied T cells from zeta-chain-deficient mice (zetaKO mice), zeta chain-bearing mice (zeta(+) mice), and from zetaKO mice expressing a FcRgamma chain transgene (FcRgammaTG, zetaKO mice). Our results demonstrated that, compared with those of control mice, CD4(+) T cells and not CD8(+) T cells from zetaKO mice were polarized into IFN-gamma-producing cells. Some of these IFN gamma-producing cells could also secrete IL-10. Interestingly, zetaKO mouse T cells produced IFN-gamma even after they were cultured in a Th2 condition. Our studies to determine the molecular mechanisms underlying the polarized IFN-gamma production revealed that the expression level of STAT4 and T-bet were up regulated in freshly isolated T cells from zetaKO mice. Further studies showed that noncultured zetaKO mice CD4(+) T cells and thymocytes bore a unique memory cell-like CD44(high), CD62L(low/neg) phenotype. Altogether, these results suggest that, in the absence of the zeta-chain, CD4(+) T cells develop as polarized IFN gamma-producing cells that bear a memory cell-like phenotype. The zeta-chain bearing T cells may produce a large amount of IFN-gamma only after they are cultured in a condition favoring Th1 cell differentiation. This study may provide important implications for the down-regulation of zeta-chain in T cells of patients bearing a variety of tumors, chronic inflammatory and infectious diseases. PMID- 15661873 TI - Activated NKT cells inhibit autoimmune diabetes through tolerogenic recruitment of dendritic cells to pancreatic lymph nodes. AB - NKT cell activation by alpha-galactosylceramide (alpha-GalCer) inhibits autoimmune diabetes in NOD mice, in part by inducing recruitment to pancreatic lymph nodes (PLNs) of mature dendritic cells (DCs) with disease-protective effects. However, how activated NKT cells promote DC maturation, and what downstream effect this has on diabetogenic T cells was unknown. Activated NKT cells were found to produce a soluble factor(s) inducing DC maturation. Initially, there was a preferential accumulation of mature DCs in the PLNs of alpha-GalCer-treated NOD mice, followed by a substantial increase in T cells. Adoptive transfer of a diabetogenic CD8 T cell population (AI4) induced a high rate of disease (75%) in PBS-treated NOD recipients, but not in those pretreated with alpha-GalCer (8%). Significantly, more AI4 T cells accumulated in PLNs of alpha-GalCer than PBS-treated recipients, while no differences were found in mesenteric lymph nodes from each group. Compared with those in mesenteric lymph nodes, AI4 T cells entering PLNs underwent greater levels of apoptosis, and the survivors became functionally anergic. NKT cell activation enhanced this process. Hence, activated NKT cells elicit diabetes protection in NOD mice by producing a soluble factor(s) that induces DC maturation and accumulation in PLNs, where they subsequently recruit and tolerize pathogenic T cells. PMID- 15661874 TI - Cathepsin S controls MHC class II-mediated antigen presentation by epithelial cells in vivo. AB - Epithelial cells at environmental interfaces provide protection from potentially harmful agents, including pathogens. In addition to serving as a physical barrier and producing soluble mediators of immunity, such as cytokines or antimicrobial peptides, these cells are thought to function as nonprofessional APCs. In this regard, intestinal epithelial cells are particularly prominent because they express MHC class II molecules at the site of massive antigenic exposure. However, unlike bone marrow-derived professional APC, such as dendritic cells or B cells, little is known about the mechanisms of MHC class II presentation by the nonprofessional APC in vivo. The former use the lysosomal cysteine protease cathepsin S (Cat S), whereas thymic cortical epithelial cells use cathepsin L (Cat L) for invariant chain degradation and MHC class II maturation. Unexpectedly, we found that murine Cat S plays a critical role in invariant chain degradation in intestinal epithelial cells. Furthermore, we report that nonprofessional APC present a class II-bound endogenous peptide to naive CD4 T cells in vivo in a Cat S-dependent fashion. These results suggest that in vivo, both professional and nonprofessional MHC class II-expressing APC use Cat S, but not Cat L, for MHC class II-mediated Ag presentation. PMID- 15661875 TI - Roles for common cytokine receptor gamma-chain-dependent cytokines in the generation, differentiation, and maturation of NK cell precursors and peripheral NK cells in vivo. AB - NK cells differentiate in adult mice from bone marrow hemopoietic progenitors. Cytokines, including those that signal via receptors using the common cytokine receptor gamma-chain (gamma(c)), have been implicated at various stages of NK cell development. We have previously described committed NK cell precursors (NKPs), which have the capacity to generate NK cells, but not B, T, erythroid, or myeloid cells, after in vitro culture or transfer to a fetal thymic microenvironment. NKPs express the CD122 Ag (beta chain of the receptors for IL 2/IL-15), but lack other mature NK markers, including NK1.1, CD49b (DX5), or members of the Ly49 gene family. In this report, we have analyzed the roles for gamma(c)-dependent cytokines in the generation of bone marrow NKP and in their subsequent differentiation to mature NK cells in vivo. Normal numbers of NKPs are found in gamma(c)-deficient mice, suggesting that NK cell commitment is not dependent on IL-2, IL-4, IL-7, IL-9, IL-15, or IL-21. Although IL-2, IL-4, and IL 7 have been reported to influence NK cell differentiation, we find that mice deficient in any or all of these cytokines have normal NK cell numbers, phenotype, and effector functions. In contrast, IL-15 plays a dominant role in early NK cell differentiation by maintaining normal numbers of immature and mature NK cells in the bone marrow and spleen. Surprisingly, the few residual NK cells generated in absence of IL-15 appear relatively mature, expressing a variety of Ly49 receptors and demonstrating lytic and cytokine production capacity. PMID- 15661876 TI - Enhanced production of IL-10 by dendritic cells deficient in CIITA. AB - Dendritic cells (DC) are professional APCs that play a critical role in regulating immunity. In DC, maturation-induced changes in MHC class II expression and Ag presentation require transcriptional regulation by CIITA. To study the role of CIITA in DC, we evaluated key cell functions in DC from CIITA-deficient (CIITA(-/-)) mice. The ability to take up Ag, measured by fluid phase endocytosis, was comparable between CIITA(-/-) and control DC. Although CIITA(-/ ) DC lack MHC class II, they maintained normal expression of costimulatory molecules CD80, CD86, and CD40. In contrast, CIITA(-/-) DC activated with LPS or CpG expressed increased IL-10 levels, but normal levels of TNF-alpha and IL-12 relative to control. Enhanced IL-10 was due to greater IL-10 mRNA in CIITA(-/-) DC. Abeta(-/-) DC, which lack MHC class II but express CIITA normally, had exhibited no difference in IL-10 compared with control. When CIITA was cotransfected with an IL-10 promoter-reporter into a mouse monocyte cell line, RAW 264.7, IL-10 promoter activity was decreased. In addition, reintroducing CIITA into CIITA(-/-) DC reduced production of IL-10. In all, these data suggest that CIITA negatively regulates expression of IL-10, and that CIITA may direct DC function in ways that extend beyond control of MHC class II. PMID- 15661877 TI - Phosphatidylinositol 3-kinase regulates thymic exit. AB - To understand the role of PI3K during T cell development, we generated transgenic mice expressing the N terminus of the PI3K catalytic subunit (p110(ABD); ABD, adaptor binding domain) in thymocytes. Expression of p110(ABD) activates endogenous p110 and results in the accumulation of mature single-positive CD3(high)heat-stable Ag(low) thymocytes. This is mostly due to a defect in emigration of those cells, as shown by the delayed appearance of peripheral T cells in neonatal transgenic mice and by competitive adoptive transfer experiments. Although the mechanisms underlying these effects of PI3K are not yet clear, our results show an important role for PI3K activity in the regulation of mature thymocyte exit to the periphery. PMID- 15661878 TI - MAPK p38 alpha is dispensable for lymphocyte development and proliferation. AB - Signals mediated by the p38alpha MAPK have been implicated in many processes required for the development and effector functions of innate and adaptive immune responses. As mice deficient in p38alpha exhibit embryonic lethality, most analyses of p38alpha function in lymphocytes have relied on the use of pharmacologic inhibitors and dominant-negative or constitutively active transgenes. In this study, we have generated a panel of low passage p38alpha(+/+), p38alpha(+/-), and p38alpha(-/-) embryonic stem (ES) cells through the intercrossing of p38alpha(+/-) mice. These ES cells were used to generate chimeric mice by RAG-deficient blastocyst complementation, with the lymphocytes in these mice being derived entirely from the ES cells. Surprisingly, B and T cell development were indistinguishable when comparing chimeric mice generated with p38alpha(+/+), p38alpha(+/-), and p38alpha(-/-) ES cell lines. Moreover, proliferation of p38alpha(-/-) B and T cells in response to Ag receptor and non Ag receptor stimuli was intact. Thus, p38alpha is not an essential component of signaling pathways required for robust B and T lymphocyte developmental, nor is p38alpha essential for the proliferation of mature B and T cells. PMID- 15661879 TI - Ig alpha/Ig beta complexes generate signals for B cell development independent of selective plasma membrane compartmentalization. AB - Ligand-induced BCR association with detergent-resistant plasma membrane compartments (lipid rafts) has been argued to be essential for initiating and/or sustaining Igalpha/Igbeta-dependent BCR signaling. Because a fraction of the BCR and an even larger fraction of the preBCR associates with lipid rafts in the apparent absence of ligand stimulation, it has been proposed that raft-associated receptor complexes mediate the ligand-independent basal signaling events observed in resting B lineage cells. However, there is no direct evidence that localization of Igalpha/Igbeta-containing complexes to detergent-resistant membrane compartments is absolutely required for the signaling events that drive B cell development. To address these issues we have designed surrogate preBCR/Igalpha/Igbeta complexes that are incapable of ligand-induced aggregation and that are preferentially targeted to either raft or nonraft compartments. An analysis of their ability to promote the preBCR-dependent proB-->preB cell transition of murine B cell progenitors revealed that expression of these surrogate receptor complexes at levels that approximate that of the conventional preBCR can drive B cell development in a manner independent of both aggregation and lipid raft localization. PMID- 15661880 TI - The low affinity Fc receptor for IgG functions as an effective cytolytic receptor for self-specific CD8 T cells. AB - We have recently described a population of self-Ag-specific murine CD8(+) T cells with a memory phenotype that use receptors of both the adaptive and innate immune systems in the detection of transformed and infected cells. In this study we show that upon activation with IL-2 with or without Ag, between 10 and 20% of the activated self-specific CD8(+) T cells express the low affinity FcR for IgG. By contrast, all IL-2-activated NK cells express high levels of this FcR. The FcR comprises the FcgammaRIIIalpha and FcRgamma subunits. However, the FcRgamma subunit also associates with the CD3 complex, and this association probably contributes to the low expression of FcR in activated cells. Although the FcR is expressed at a low level on activated self-specific CD8(+) T cells, it functions very efficiently as a cytolytic receptor in ADCC. FcR-dependent killing occurred in the absence of TCR stimulation, but could be augmented by concurrent stimulation of the TCR. In addition to mediating ADCC, engagement of the FcR on self-specific CD8(+) T cells results in the production of both IFN-gamma and TNF alpha. This is the first report of an activating FcR on self-specific murine CD8(+)alphabeta TCR(+) T cells and establishes the importance of innate immune system receptors in the function of these self-specific CD8(+) T cells. PMID- 15661881 TI - Synthetic TLR agonists reveal functional differences between human TLR7 and TLR8. AB - Although TLR7 and TLR8 are phylogenetically and structurally related, their relative functions are largely unknown. The role of TLR7 has been established using TLR7-deficient mice and small molecule TLR7 agonists. The absence of TLR8 selective agonists has hampered our understanding of the role of TLR8. In this study TLR agonists selective for TLR7 or TLR8 were used to determine the repertoire of human innate immune cells that are activated through these TLRs. We found that TLR7 agonists directly activated purified plasmacytoid dendritic cells and, to a lesser extent, monocytes. Conversely, TLR8 agonists directly activated purified myeloid dendritic cells, monocytes, and monocyte-derived dendritic cells (GM-CSF/IL-4/TGF-beta). Accordingly, TLR7-selective agonists were more effective than TLR8-selective agonists at inducing IFN-alpha- and IFN-regulated chemokines such as IFN-inducible protein and IFN-inducible T cell alpha chemoattractant from human PBMC. In contrast, TLR8 agonists were more effective than TLR7 agonists at inducing proinflammatory cytokines and chemokines, such as TNF-alpha, IL-12, and MIP-1alpha. Thus, this study demonstrated that TLR7 and TLR8 agonists differ in their target cell selectivity and cytokine induction profile. PMID- 15661882 TI - Bone marrow is a preferred site for homeostatic proliferation of memory CD8 T cells. AB - Proliferative renewal of memory CD8 T cells is essential for maintaining long term immunity. In this study, we examined the contributions that various tissue microenvironments make toward the homeostatic proliferation of Ag-specific memory CD8 T cells. We found that dividing memory T cells were present in both lymphoid and nonlymphoid tissues. However, the bone marrow was the preferred site for proliferation and contained a major pool of the most actively dividing memory CD8 T cells. Adoptive transfer studies indicated that memory cells migrated through the bone marrow and divided there preferentially. These results show that the bone marrow is not only the source of stem cells for generating naive T cells but also provides the necessary signals for the self-renewal of memory T cells. PMID- 15661883 TI - Induction of impaired antitumor immunity by fusion of MHC class II-deficient dendritic cells with tumor cells. AB - To dissect the role of Ag presentation through MHC class I and/or II pathways by dendritic cell (DC)-tumor fusion cells, we have created various types of DC-tumor fusion cells by alternating fusion cell partners. Fusions of MC38/MUC1 carcinoma cells with DC from wild-type (WT-DC), MHC class I knockout (IKO-DC), class II knockout (IIKO-DC), or class I and II knockout (I/IIKO-DC) mice created WTDC fusion cells (FC), IKO-FC, IIKO-FC, and I/IIKO-FC, respectively. MHC class II- and MUC1-positive fusion cells were constructed by fusion of B16/MUC1 melanoma cells with IKO-DC (IKO/B16-FC). Immunization of MUC1 transgenic mice with 5 x 10(5) WTDC-FC, IKO-FC, IIKO-FC, or I/IIKO-FC provided 100, 91.7, 61.5, and 15.4% protection, respectively, against tumor challenge with MC38/MUC1 cells. In contrast, all mice immunized with irradiated MC38/MUC1 tumor cells or WT-DC developed tumors. One group of mice was immunized with 5 x 10(5) IKO/B16-FC and then challenged with B16/Ia(+)/MUC1 on one flank and MC38/MUC1 on the other flank. Immunization of these mice with IKO/B16-FC resulted in 100 and 78.6% protection against B16/Ia(+)/MUC1 and MC38/MUC1 tumor challenge, respectively. The antitumor immunity induced by immunization with IKO/B16-FC was able to inhibit the growth of MHC class II-negative tumor. In addition, in vivo results correlated with the induction of Ag-specific CTL. Collectively, the data indicate that MHC class II Ag presentation targeting activation of CD4 T cells is indispensable for antitumor immunity. PMID- 15661884 TI - CD226 expression deficiency causes high sensitivity to apoptosis in NK T cells from patients with systemic lupus erythematosus. AB - Humans and mice with systemic lupus erythematosus (SLE) and related autoimmune diseases have reduced numbers of NK T cells. An association between NK T cell deficiency and autoimmune disease has been identified. However, the mechanisms for reduction of NK T cell number in patients with SLE are unknown. In the present study we report that NK T cells from active SLE patients are highly sensitive to anti-CD95-induced apoptosis compared with those from normal subjects and inactive SLE patients. CD226 expression is deficient on NK T cells from active SLE patients. The expression of one antiapoptotic member protein, survivin, is found to be selectively deficient in freshly isolated NK T cells from active SLE patients. CD226 preactivation significantly up-regulates survivin expression and activation, which can rescue active SLE NK T cells from anti-CD95 induced apoptosis. In transfected COS7 cells, we confirm that anti-CD95-mediated death signals are inhibited by activation of the CD226 pathway through stabilization of caspase-8 and caspase-3 and through activation of survivin. We therefore conclude that deficient expression of CD226 and survivin in NK T cells from active SLE is a molecular base of high sensitivity of the cells to anti-CD95 induced apoptosis. These observations offer a potential explanation for high apoptotic sensitivity of NK T cells from active SLE, and provide a new insight into the mechanism of reduction of NK T cell number in SLE and understanding the association between NK T cell deficiency and autoimmune diseases. PMID- 15661885 TI - The role of p53 and Fas in a model of acute murine graft-versus-host disease. AB - Graft-vs-host disease (GVHD) is a devastating, frequently fatal, pathological condition associated with lesions in specific target organs, including the intestine, liver, lung, and skin, as well as pancytopenia and alopecia. Bone marrow (BM) atrophy is observed in acutely diseased animals, but the underlying mechanisms of hemopoietic stem cell depletion remained to be established. We used an experimental mouse model of acute GVHD in which parental cells were injected into F(1) hosts preconditioned by sublethal irradiation. The resulting graft-vs host response was kinetically consistent, resulting in lethality within 3 wk. We observed disease pathology in the liver and small intestine, and consistent with previous observations, we found BM atrophy to be a factor in the onset of acute disease. The product of the protooncogene, p53, is known to be a key player in many physiological examples of apoptosis. We investigated the role of p53 in the apoptosis of BM cells (BMC) during the development of acute disease and found that at least one copy of the p53 gene is necessary for depletion of BM and subsequent lethality in host animals. BM depletion was preceded by induction of the death receptor, Fas, on the surface of host stem cells, and induction of Fas was coincidental with the sensitization of BMC to Fas-mediated apoptosis. Our data indicate that BM depletion in acute GVHD is mediated by p53-dependent up regulation of Fas on BMC, which leads to Fas-dependent depletion and subsequent disease. PMID- 15661886 TI - Heat shock protein 60: specific binding of lipopolysaccharide. AB - Human heat shock protein 60 (HSP60) has been shown to bind to the surface of innate immune cells and to elicit a proinflammatory response. In this study we demonstrate that the macrophage stimulatory property of recombinant human HSP60 is tightly linked to the HSP60 molecule and is lost after protease treatment. However, inhibition of macrophage stimulation was reached by the LPS-binding peptide magainin II amide. Indeed, HSP60 specifically bound [(3)H]LPS. [(3)H]LPS binding to HSP60 was saturable and competable by the unlabeled ligand. To identify the epitope region of the HSP60 molecule responsible for specific LPS binding, we analyzed the effect of several anti-HSP60 mAbs on HSP60-induced production of inflammatory mediators from macrophages. We identified only one mAb, clone 4B9/89, which blocked the macrophage stimulatory activity of the chaperone. The epitope specificity of this mAb points to the region aa 335-366 of HSP60. Clone 4B9/89 also strongly inhibited [(3)H]LPS binding to HSP60. A more detailed analysis was performed by screening with selected overlapping 20-mer peptides of the HSP60 sequence, covering the region aa 331-380. Only one peptide blocked LPS binding to HSP60, thereby restricting the potential LPS-binding region to aa 351-370 of HSP60. Finally, analysis of selected 15-mer peptides and a 13-mer peptide of the HSP60 sequence revealed that most of the LPS-binding region was accounted for by aa 354-365 of HSP60, with the motif LKGK being critical for binding. Our studies identified a defined region of HSP60 involved in LPS binding, thereby implicating a physiological role of human HSP60 as LPS binding protein. PMID- 15661887 TI - The pathway of antigen uptake and processing dictates MHC class II-mediated B cell survival and activation. AB - The influence of the pathway of Ag uptake and processing on MHC class II (CII) mediated B cell function is unknown. In this study, we investigate in resting and activated (via the BCR or CD40) B cells the biological properties of CII-peptide complexes (CII-peptide) generated by either the BCR-mediated Ag processing (type I complex) or fluid phase Ag processing (type II complex). Compared with type I complex, ligation of type II complex by either specific Ab or the TCR in Ag presenting assay results in significant decreases in B cell survival rate (50 100%) and expression levels of CII, CD86, and CD54. Loss of B cells following ligation of type II complex occurs in the presence of a comparatively good level of specific CD4(+) T cell division, indicating that B cell loss is a late event following T cell stimulation. Comparative analysis of T and B cell conjugates after Ab ligation of type I or II complex reveals decreased efficiency of the latter in forming conjugates. Neither initial differential levels of CII and other studied surface markers, B cell type inherent differences, BCR signaling, T cell proliferation, nor initial density of CII-peptide complexes could explain the T cell-induced B cell loss. We propose that the context in which CII-peptide complexes are present in the membrane following BCR uptake and processing leads to B cell survival. Thus, appropriate targeting of Ag ensures generation of relevant immune responses. PMID- 15661888 TI - Characterization of a B220+ lymphoid cell subpopulation with immune modulatory functions in nasal-associated lymphoid tissues. AB - Complex mechanisms operate on mucosal tissues to regulate immune responsiveness and tolerance. When the lymphocyte subpopulations from murine nasal-associated lymphoid tissues (NALT) were characterized, we observed an accumulation of B220(low)CD3(low)CD4(-)CD8(-)CD19(-)c-Kit(+) cells. TCR transgenic mice and athymic mice were used for monitoring T cell lineage and the presence of extrathymic T cell precursors. The majority of cells from NALT exhibited a T cell precursor phenotype (CD4(-)CD8(-)CD19(-)c-Kit(+)). Fas-independent apoptosis was their main mechanism of cell death. We also demonstrated that B220(low)CD4(-)CD8( )CD19(-) cells from NALT exhibited the potential to down-regulate the activation of mature T cells. However, the innate immunity receptor TLR2 was also highly expressed by this cell subpopulation. Moreover, nasal stimulation with a TLR2/6 agonist resulted in a partial activation of the double-negative cells. These results suggest that the immune responses in NALT may be in part modulated by a cell subpopulation that maintains a tolerogenic milieu by its proapoptotic status and suppressive activity, which can be reverted through stimulation of a TLR signaling cascade. PMID- 15661889 TI - Limited ability of antigen-specific Th1 responses to inhibit Th2 cell development in vivo. AB - Th1 and Th2 cells mutually antagonize each other's differentiation. Consequently, allergen-specific Th1 cells are believed to be able to suppress the development of Th2 cells and to prevent the development of atopic disorders. To determine whether a pre-existing Ag-specific Th1 response can affect the development of Th2 cells in vivo, we used an immunization model of Ag-pulsed murine dendritic cell (DC) transfer to induce distinct Th responses. When transferred into naive mice, Ag-pulsed CD8alpha(+) DCs induced a Th1 response and the production of IgG2a, whereas CD8alpha(-) DCs primed a Th2 response and the production of IgE. In the presence of a pre-existing Ag-specific Th2 environment due to Ag-pulsed CD8alpha( ) DC transfer, CD8alpha(+) DCs failed to prime Th1 cells. In contrast, CD8alpha( ) DCs could prime a Th2 response in the presence of a pre-existing Ag-specific Th1 environment. Moreover, exogenous IL-4 abolished the Th1-inducing potential of CD8alpha(+) DCs in vitro, but the addition of IFN-gamma did not effectively inhibit the potential of CD8alpha(-) DCs to prime IL-4-producing cells. Thus, Th1 and Th2 cells differ in their potential to inhibit the development of the other. This suggests that the early induction of allergen-specific Th1 cells before allergy sensitization will not prevent the development of atopic disorders. PMID- 15661890 TI - IFN-gamma suppresses STAT6 phosphorylation by inhibiting its recruitment to the IL-4 receptor. AB - Polarized Th1 cells show a stable phenotype: they become insensitive to IL-4 stimulation and lose the potential to produce IL-4. Previously, we reported that IFN-gamma played a critical role in stabilizing Th1 phenotype. However, the mechanism by which IFN-gamma stabilizes Th1 phenotype is not clear. In this study, we compared STAT6 phosphorylation in wild-type (WT) and IFN-gamma receptor knockout (IFNGR(-/-)) Th1 cells. We found a striking diminution of STAT6 phosphorylation in differentiated WT Th1 cells, but not in differentiated IFNGR( /-) Th1 cells. The impairment of STAT6 phosphorylation in differentiated WT Th1 cells was not due to a lack of IL-4R expression or phosphorylation. Jak1 and Jak3 expression and phosphorylation were comparable in both cell types. No differential expression of suppressor of cytokine signaling 1 (SOCS1), SOCS3, or SOCS5 was observed in the two cell types. In addition, Src homology 2-containing phosphatase mutation did not affect IL-4-induced STAT6 phosphorylation in differentiated Th1 cells derived from viable motheaten (me(v)/me(v)) mice. These results led us to focus on a novel mechanism. By using a pulldown assay, we observed that STAT6 in WT Th1 cells bound less effectively to the phosphorylated IL-4R/GST fusion protein than that in IFNGR(-/-) Th1 cells. Our results suggest that IFN-gamma may suppress phosphorylation of STAT6 by inhibiting its recruitment to the IL-4R. PMID- 15661891 TI - Phosphostim-activated gamma delta T cells kill autologous metastatic renal cell carcinoma. AB - Metastatic renal cell carcinoma, inherently resistant to conventional treatments, is considered immunogenic. Indeed, partial responses are obtained after treatment with cytokines such as IL-2 or IFN-alpha, suggesting that the immune system may control the tumor growth. In this study, we have investigated the ability of the main subset of peripheral gammadelta lymphocytes, the Vgamma9Vdelta2-TCR T lymphocytes, to induce an effective cytotoxic response against autologous primary renal cell carcinoma lines. These gammadelta T cells were expanded ex vivo using a Vgamma9Vdelta2 agonist, a synthetic phosphoantigen called Phosphostim. From 11 of 15 patients, the peripheral Vgamma9Vdelta2 T cells were amplified in vitro by stimulating PBMCs with IL-2 and Phosphostim molecule. These expanded Vgamma9Vdelta2 T cells express activation markers and exhibit an effector/memory phenotype. They display a selective lytic potential toward autologous primary renal tumor cells and not against renal NC. The lytic activity involves the perforin-granzyme pathway and is mainly TCR and NKG2D receptor dependent. Furthermore, an increased expression of MHC class I-related molecule A or B proteins, known ligands of NKG2D, are detected on primary renal tumor cells. Interestingly, from 2 of the 11 positive cultures in response to Phosphostim, expanded-Vgamma9Vdelta2 T cells present an expression of killer cell Ig-like receptors, suggesting their prior recruitment in vivo. Unexpectedly, on serial frozen sections from three tumors, we observe a gammadelta lymphocyte infiltrate that was mainly composed of Vgamma9Vdelta2 T cells. These results outline that Vgamma9Vdelta2-TCR effectors may represent a promising approach for the treatment of metastatic renal cell carcinoma. PMID- 15661892 TI - The CD200 receptor is a novel and potent regulator of murine and human mast cell function. AB - CD200R is a member of the Ig supergene family that is primarily expressed on myeloid cells. Recent in vivo studies have suggested that CD200R is an inhibitory receptor capable of regulating the activation threshold of inflammatory immune responses. Here we provide definitive evidence that CD200R is expressed on mouse and human mast cells and that engagement of CD200R by agonist Abs or ligand results in a potent inhibition of mast cell degranulation and cytokine secretion responses. CD200R-mediated inhibition of FcepsilonRI activation was observed both in vitro and in vivo and did not require the coligation of CD200R to FcepsilonRI. Unlike the majority of myeloid inhibitory receptors, CD200R does not contain a phosphatase recruiting inhibitory motif (ITIM); therefore, we conclude that CD200R represents a novel and potent inhibitory receptor that can be targeted in vivo to regulate mast cell-dependent pathologies. PMID- 15661893 TI - CD70 signaling is critical for CD28-independent CD8+ T cell-mediated alloimmune responses in vivo. AB - The inability to reproducibly induce robust and durable transplant tolerance using CD28-B7 pathway blockade is in part related to the persistence of alloreactive effector/memory CD8(+) T cells that are less dependent on this pathway for their cellular activation. We studied the role of the novel T cell costimulatory pathway, CD27-CD70, in alloimmunity in the presence and absence of CD28-B7 signaling. CD70 blockade prolonged survival of fully mismatched vascularized cardiac allografts in wild-type murine recipients, and in CD28 deficient mice induced long-term survival while significantly preventing the development of chronic allograft vasculopathy. CD70 blockade had little effect on CD4(+) T cell function but prevented CD8(+) T cell-mediated rejection, inhibited the proliferation and activation of effector CD8(+) T cells, and diminished the expansion of effector and memory CD8(+) T cells in vivo. Thus, the CD27-CD70 pathway is critical for CD28-independent effector/memory CD8(+) alloreactive T cell activation in vivo. These novel findings have important implications for the development of transplantation tolerance-inducing strategies in primates and humans, in which CD8(+) T cell depletion is currently mandatory. PMID- 15661895 TI - Intestinal dendritic cell subsets: differential effects of systemic TLR4 stimulation on migratory fate and activation in vivo. AB - Dendritic cells (DC) present peripheral Ags to T cells in lymph nodes, but also influence their differentiation (tolerance/immunity, Th1/Th2). To investigate how peripheral conditions affect DC properties and might subsequently regulate T cell differentiation, we examined the effects of a potent DC-activating, TLR-4 mediated stimulus, LPS, on rat intestinal and hepatic DC in vivo. Steady-state rat intestinal and hepatic lymph DC are alpha(E2) integrin(high) (CD103) and include two subsets, signal regulatory protein alpha (SIRPalpha)(hi/low), probably representing murine CD8alphaalpha(-/+) DC. Steady-state lamina propria DC are immature; surface MHC class II(low), but steady-state lymph DC are semimature, MHC class II(high), but CD80/86(low). Intravenous LPS induced rapid lamina propria DC emigration and increased lymph DC traffic without altering SIRPalpha(high)/SIRPalpha(low) proportions. CD80/86 expression on lymph or mesenteric node DC was not up-regulated after i.v. LPS. In contrast, i.v. LPS stimulated marked CD80/86 up-regulation on splenic DC. CD80/86 expression on intestinal lymph DC, however, was increased after in vitro culture with TNF-alpha or GM-CSF, but not with up to 5 mug/ml LPS. Steady-state SIRPalpha(low) DC localized to T cell areas of mesenteric nodes, spleen, and Peyer's patch, whereas SIRPalpha(high) DC were excluded from these areas. Intravenous LPS stimulated rapid and abundant SIRPalpha(high) DC accumulation in T cell areas of mesenteric nodes and spleen. In striking contrast, i.v. LPS had no effect on DC numbers or distribution in Peyer's patches. Our results suggest that any explanation of switching between tolerance and immunity as well as involving changes in DC activation status must also take into account differential migration of DC subsets. PMID- 15661894 TI - Dynamic interactions of Fc gamma receptor IIB with filamin-bound SHIP1 amplify filamentous actin-dependent negative regulation of Fc epsilon receptor I signaling. AB - The engagement of high affinity receptors for IgE (FcepsilonRI) generates both positive and negative signals whose integration determines the intensity of mast cell responses. FcepsilonRI-positive signals are also negatively regulated by low affinity receptors for IgG (FcgammaRIIB). Although the constitutive negative regulation of FcepsilonRI signaling was shown to depend on the submembranous F actin skeleton, the role of this compartment in FcgammaRIIB-dependent inhibition is unknown. We show in this study that the F-actin skeleton is essential for FcgammaRIIB-dependent negative regulation. It contains SHIP1, the phosphatase responsible for inhibition, which is constitutively associated with the actin binding protein, filamin-1. After coaggregation, FcgammaRIIB and FcepsilonRI rapidly interact with the F-actin skeleton and engage SHIP1 and filamin-1. Later, filamin-1 and F-actin dissociate from FcR complexes, whereas SHIP1 remains associated with FcgammaRIIB. Based on these results, we propose a dynamic model in which the submembranous F-actin skeleton forms an inhibitory compartment where filamin-1 functions as a donor of SHIP1 for FcgammaRIIB, which concentrate this phosphatase in the vicinity of FcepsilonRI and thereby extinguish activation signals. PMID- 15661896 TI - Kinase-independent functions for Itk in TCR-induced regulation of Vav and the actin cytoskeleton. AB - The Tec family kinase Itk is an important regulator of Ca(2+) mobilization and is required for in vivo responses to Th2-inducing agents. Recent data also implicate Itk in TCR-induced regulation of the actin cytoskeleton. We have evaluated the requirements for Itk function in TCR-induced actin polarization. Reduction of Itk expression via small interfering RNA treatment of the Jurkat human T lymphoma cell line or human peripheral blood T cells disrupted TCR-induced actin polarization, a defect that correlated with decreased recruitment of the Vav guanine nucleotide exchange factor to the site of Ag contact. Vav localization and actin polarization could be rescued by re-expression of either wild-type or kinase-inactive murine Itk but not by Itk containing mutations affecting the pleckstrin homology or Src homology 2 domains. Additionally, we find that Itk is constitutively associated with Vav. Loss of Itk expression did not alter gross patterns of Vav tyrosine phosphorylation but appeared to disrupt the interactions of Vav with SLP-76. Expression of membrane-targeted Vav, Vav-CAAX, can rescue the small interfering RNA to Itk-induced phenotype, implicating the alteration in Vav localization as directly contributing to the actin polarization defect. These data suggest a kinase-independent scaffolding function for Itk in the regulation of Vav localization and TCR-induced actin polarization. PMID- 15661897 TI - Interaction between phosphatidylserine and the phosphatidylserine receptor inhibits immune responses in vivo. AB - Phosphatidylserine (PS) on apoptotic cells promotes their uptake and induces anti inflammatory responses in phagocytes, including TGF-beta release. Little is known regarding the effects of PS on adaptive immune responses. We therefore investigated the effects of PS-containing liposomes on immune responses in mice in vivo. PS liposomes specifically inhibited responses to Ags as determined by decreased draining lymph node tissue mass, with reduced numbers of total leukocytes and Ag-specific CD4(+) T cells. There was also a decrease in formation and size of germinal centers in spleen and lymph nodes, accompanied by decreased levels of Ag-specific IgG in blood. Many of these effects were mimicked by an agonistic Ab-specific for the PS receptor. TGF-beta appears to play a critical role in this inhibition, as the inhibitory effects of PS were reversed by in vivo administration of anti-TGF-beta Ab. PS-containing liposomes did not appear to directly inhibit dendritic cell maturation in vitro in response to a variety of stimuli, nor did it prevent their migration to regional lymph nodes in vivo, suggesting that the inhibitory effects may have resulted from complicated interactions between tissue cells and dendritic cells, subsequently inhibiting their ability to productively activate T lymphocytes. PMID- 15661898 TI - Tim-3+ T-bet+ tumor-specific Th1 cells colocalize with and inhibit development and growth of murine neoplasms. AB - Although T cells infiltrate many types of murine and human neoplasms, in many instances tumor-specific cytotoxicity is not observed. Strategies to stimulate CTL-mediated antitumor immunity have included in vitro stimulation and/or genetic engineering of T cells, followed by adoptive transfer into tumor-bearing hosts. In this model of B cell lymphoma in SJL/J mice, we used Tim-3(+) T-bet(+) Th1 cells to facilitate the development of tumor-specific CTL. Tumor-specific Th1 cell lines were polarized with IL-12 during in vitro stimulation and long term maintenance. As few as 5 million Tim-3(+) T-bet(+) Th1 cells enabled recipients to resist growth of malignant transplantable cells. In addition, similar numbers of Th1 cells injected into 2- to 3-mo-old mice inhibited development of the spontaneous primary lymphomas, which normally arise in 90% of aging mice. CFSE(+) Th1 cells colocalized with injected tumor cells in vivo and formed conjugates with the tumor cells within follicles, whereas in nontumor-challenged recipients the CFSE(+) Th1 cells localized only within the T cell zones of the spleen. These results provide evidence that adoptive immunotherapy with Tim-3(+) T-bet(+) tumor specific Th1 cells can be used to induce host cytotoxic responses that inhibit the development and growth of neoplastic cells. PMID- 15661899 TI - IFN-gamma determines distinct clinical outcomes in autoimmune encephalomyelitis. AB - Experimental autoimmune encephalomyelitis (EAE) is an inflammatory disease of the CNS initiated by autoreactive CD4(+) T cells. EAE classically presents with a progressive ascending paralysis and is a model of multiple sclerosis that recapitulates some aspects of the disease. In this report we describe a mouse strain that spontaneously develops a severe, nonclassical form of EAE with 100% incidence. The distinct clinical phenotype is marked initially by a slight head tilt, progressing to a severe head tilt, spinning, or a rotatory motion. Classical EAE spontaneously occurs in myelin basic protein (MBP)-specific TCR transgenic RAG-1(-/-) mice (referred to as T/R(-)), whereas nonclassical EAE spontaneously occurs in T/R(-) IFN-gamma(-/-) mice (T/R(-)gamma(-)). Thus, the TCR recognizes the same Ag (MBP) and uses identical TCR in both cases. The cellular infiltrate in nonclassical EAE is predominantly found in the brainstem and cerebellum, with very little inflammation in the spinal cord, which is primarily affected in classical disease. Importantly, depending on the genetic makeup and priming conditions of the MBP-specific T cells, nonclassical disease can occur in the presence of an inflammatory infiltrate with eosinophilic, neutrophilic, or monocytic characteristics. Finally, we believe that nonclassical spontaneous EAE could be a useful model for the study of some characteristics of multiple sclerosis not observed in classical EAE, such as the inflammatory responses in the brainstem and cerebellum that can cause vertigo. PMID- 15661900 TI - Rolling of Th1 cells via P-selectin glycoprotein ligand-1 stimulates LFA-1 mediated cell binding to ICAM-1. AB - Activated T cells migrate from the blood into nonlymphoid tissues through a multistep process that involves cell rolling, arrest, and transmigration. P Selectin glycoprotein ligand-1 (PSGL-1) is a major ligand for P-selectin expressed on subsets of activated T cells such as Th1 cells and mediates cell rolling on vascular endothelium. Rolling cells are arrested through a firm adhesion step mediated by integrins. Although chemokines presented on the endothelium trigger integrin activation, a second mechanism has been proposed where signaling via rolling receptors directly activates integrins. In this study, we show that Ab-mediated cross-linking of the PSGL-1 on Th1 cells enhances LFA-1-dependent cell binding to ICAM-1. PSGL-1 cross-linking did not enhance soluble ICAM-1 binding but induced clustering of LFA-1 on the cell surface, suggesting that an increase in LFA-1 avidity may account for the enhanced binding to ICAM-1. Combined stimulation by PSGL-1 cross-linking and the Th1-stimulating chemokine CXCL10 or CCL5 showed a more than additive effect on LFA-1-mediated Th1 cell adhesion as well as on LFA-1 redistribution on the cell surface. Moreover, PSGL-1-mediated rolling on P-selectin enhanced the Th1 cell accumulation on ICAM 1 under flow conditions. PSGL-1 cross-linking induced activation of protein kinase C isoforms, and the increased Th1 cell adhesion observed under flow and also static conditions was strongly inhibited by calphostin C, implicating protein kinase C in the intracellular signaling in PSGL-1-mediated LFA-1 activation. These results support the idea that PSGL-1-mediated rolling interactions induce intracellular signals leading to integrin activation, facilitating Th1 cell arrest and subsequent migration into target tissues. PMID- 15661901 TI - OX40 signals during priming on dendritic cells inhibit CD4 T cell proliferation: IL-4 switches off OX40 signals enabling rapid proliferation of Th2 effectors. AB - In this study we examined the role and regulation of OX40 signals during CD4 T cell priming on dendritic cells (DCs). Contrary to expectation, OX40-deficient cells proliferated more rapidly than their normal counterparts, particularly when stimulated with peptide in the absence of added cytokines. This proliferative advantage was not apparent for Th2-differentiated cells. When the reasons for this were investigated, we found that the cytokine IL-4 specifically down regulated expression of OX40 ligand on T, B, and DCs, but not on the CD4(+)CD3(-) cells linked with selection of Th2 cells into the memory compartment. OX40 ligand expression was also down-regulated on rapidly proliferating Th1 effectors. These data are compatible with OX40 signals acting during priming as a check on naive T cell proliferation while T cells integrate additional DC signals. This would serve to limit inappropriate T cell responses. In contrast, OX40 signals from CD4(+)CD3(-) cells located in the outer T zone select proliferating Th2 effectors into the memory T cell pool. PMID- 15661903 TI - Differentiation and expansion of T cells with regulatory function from human peripheral lymphocytes by stimulation in the presence of TGF-{beta}. AB - T cells with immunoregulatory function have been described in human and mouse systems. In both systems these cells can be differentiated either in the thymus or from peripheral T cells. To date, more progress has been made in the study of murine regulatory T cells, because it has been very difficult to isolate human regulatory T cells of sufficient purity and in sufficient numbers to permit detailed examinations of their biochemistry. We report in this study that human T cells with regulatory function can be differentiated in vitro from naive (CD4(+)CD45RA(+)) cord blood or peripheral T cells by stimulation with anti-CD3 and anti-CD28 in the presence of TGF-beta. Cells derived in this manner express a surface phenotype (CD25(+), CD122(+), HLA-DR(+), glucocorticoid-induced TNF receptor-related gene(+), CD103(+), CTLA-4(+)) described for human and mouse regulatory T cells and express protein and message for the transcription factor forkhead/winged helix transcription factor (FOXP3). They produce primarily TGF beta and IL-10, with lesser amounts of IFN-gamma and IL-13, when stimulated through their TCRs and are capable of inhibiting cytokine production and proliferation by stimulated naive T cells. Unlike Th1 and Th2 cells, these TGF beta-derived regulatory T cells do not appear to be dependent on the protein kinase Ctheta; pathway of NF-kappaB activation for Ag-induced responses. PMID- 15661902 TI - Analysis of marginal zone B cell development in the mouse with limited B cell diversity: role of the antigen receptor signals in the recruitment of B cells to the marginal zone. AB - The quasimonoclonal (QM) mouse provides an intelligible model to analyze the B cell selection as the competition between two major 4-hydroxy-3-nitrophenylacetyl specific B cell populations whose BCR are comprised of the knockin V(H)17.2.25 (V(H)T)-encoded H chain and the lambda1 or lambda2 L chain. In this study, we show the QM system is useful to examine how BCR signals guide a subset of B cells to the marginal zone (MZ). Compared with the control C57BL/6 mice, the QM mice had approximately 2.7-fold increased number of B cells exhibiting the MZ B cell phenotype and a larger MZ area in the spleen. Interestingly, V(H)T/lambda2 B cells significantly predominated over V(H)T/lambda1 B cells in MZ (V(H)T/lambda1:V(H)T/lambda2 approximately 3:7) and transitional 2-B cell subsets, while these two populations were comparable in immature, transitional 1, and mature counterparts. Thus, the biased use of lambda2 in the MZ B cells may be the result of selection in the periphery. The enlargement of MZ B cell compartment and the preferred recruitment of the V(H)T/lambda2 B cells were further augmented by doubling the V(H)T gene, but dampened by the dysfunction of Bruton's tyrosine kinase, suggesting a positive role of BCR signaling in this selection. Comparison of Ag specificity between V(H)T/lambda1 and V(H)T/lambda2 IgM mAbs revealed a polyreactive nature of the V(H)T/lambda2 BCR, including the reactivity with ssDNA. Taken together, it is suggested that polyreactivity (including self-reactivity) of BCR is crucial in driving B cells to differentiate into the MZ phenotype. PMID- 15661904 TI - Sonic hedgehog is produced by follicular dendritic cells and protects germinal center B cells from apoptosis. AB - The Hedgehog (Hh) signaling pathway is involved in the development of many tissues during embryogenesis, but has also been described to function in adult self-renewing tissues. In the immune system, Sonic Hedgehog (Shh) regulates intrathymic T cell development and modulates the effector functions of peripheral CD4(+) T cells. In this study we investigate whether Shh signaling is involved in peripheral B cell differentiation in mice. Shh is produced by follicular dendritic cells, mainly in germinal centers (GCs), and GC B cells express both components of the Hh receptor, Patched and Smoothened. Blockade of the Hh signaling pathway reduces the survival, and consequently the proliferation and Ab secretion, of GC B cells. Furthermore, Shh rescues GC B cells from apoptosis induced by Fas ligation. Taken together, our data suggest that Shh is one of the survival signals provided by follicular dendritic cells to prevent apoptosis in GC B cells. PMID- 15661906 TI - Differential control of CD28-regulated in vivo immunity by the E3 ligase Cbl-b. AB - The E3 ubiquitin ligase Casitas B cell lymphoma-b (Cbl-b) plays a critical role in the development of autoimmunity and sets the threshold for T cell activation. In the absence of Cbl-b, T cells stimulated via the TCR respond similarly to those that have received a CD28-mediated costimulatory signal, suggesting that the absence of Cbl-b substitutes for CD28-mediated costimulation. In this study, we show that loss of Cbl-b restores Ig class switching and germinal center formation in Vav1 mutant mice in response to an in vivo viral challenge. Genetic inactivation of Cbl-b also rescues impaired antiviral IgG production in CD28 mutant mice. Moreover, loss of CD28 results in disorganization of follicular dendritic cell clusters, which is also rescued by the Cbl-b mutation. Intriguingly, despite restored antiviral in vivo immunity and follicular dendritic cell clusters, loss of Cbl-b did not rescue germinal center formation in CD28-deficient mice. Mechanistically, in vivo vesicular stomatitis virus induced IL-4 and IFN-gamma production and up-regulation of the inducible costimulatory molecule ICOS were dependent on CD28, and could not be rescued by the loss of Cbl-b. These data provide genetic evidence that CD28-dependent in vivo immune responses and Ig class switching can be genetically uncoupled from germinal center formation and ICOS induction by Cbl-b-Vav1-regulated signaling pathways. PMID- 15661905 TI - Immune selection of hot-spot beta 2-microglobulin gene mutations, HLA-A2 allospecificity loss, and antigen-processing machinery component down-regulation in melanoma cells derived from recurrent metastases following immunotherapy. AB - Scanty information is available about the mechanisms underlying HLA class I Ag abnormalities in malignant cells exposed to strong T cell-mediated selective pressure. In this study, we have characterized the molecular defects underlying HLA class I Ag loss in five melanoma cell lines derived from recurrent metastases following initial clinical responses to T cell-based immunotherapy. Point mutations in the translation initiation codon (ATG-->ATA) and in codon 31 (TCA- >TGA) of the beta(2)-microglobulin (beta(2)m) gene were identified in the melanoma cell lines 1074MEL and 1174MEL, respectively. A hot-spot CT dinucleotide deletion within codon 13-15 was found in the melanoma cell lines 1106MEL, 1180MEL, and 1259MEL. Reconstitution of beta(2)m expression restored HLA class I Ag expression in the five melanoma cell lines; however, the HLA-A and HLA-B,-C gene products were differentially expressed by 1074MEL, 1106MEL, and 1259MEL cells. In addition, in 1259MEL cells, the Ag-processing machinery components calnexin, calreticulin, and low m.w. polypeptide 10 are down-regulated, and HLA A2 Ags are selectively lost because of a single cytosine deletion in the HLA-A2 gene exon 4. Our results in conjunction with those in the literature suggest the emergence of a preferential beta(2)m gene mutation in melanoma cells following strong T cell-mediated immune selection. Furthermore, the presence of multiple HLA class I Ag defects within a tumor cell population may reflect the accumulation of multiple escape mechanisms developed by melanoma cells to avoid distinct sequential T cell-mediated selective events. PMID- 15661907 TI - CD45 signals outside of lipid rafts to promote ERK activation, synaptic raft clustering, and IL-2 production. AB - CD45 is dynamically repositioned within lipid rafts and the immune synapse during T cell activation, although the molecular consequences of CD45 repositioning remain unclear. In this study we examine the role of CD45 membrane compartmentalization in regulating murine T cell activation. We find that raft localized CD45 antagonizes IL-2 production by opposing processive TCR signals, whereas raft-excluded CD45 promotes ERK-dependent polarized synaptic lipid raft clustering and IL-2 production. We propose that these dual CD45 activities ensure that only robust TCR signals proceed, whereas signals meeting threshold requirements are potentiated. Our findings highlight membrane compartmentalization as a key regulator of CD45 function and elucidate a novel signal transduction pathway by which raft-excluded CD45 positively regulates T cell activation. PMID- 15661908 TI - Elemental analysis of Mycobacterium avium-, Mycobacterium tuberculosis-, and Mycobacterium smegmatis-containing phagosomes indicates pathogen-induced microenvironments within the host cell's endosomal system. AB - Mycobacterium avium and Mycobacterium tuberculosis are human pathogens that infect and replicate within macrophages. Both organisms live in phagosomes that fail to fuse with lysosomes and have adapted their lifestyle to accommodate the changing environment within the endosomal system. Among the many environmental factors that could influence expression of bacterial genes are the concentrations of single elements within the phagosomes. We used a novel hard x-ray microprobe with suboptical spatial resolution to analyze characteristic x-ray fluorescence of 10 single elements inside phagosomes of macrophages infected with M. tuberculosis and M. avium or with avirulent M. smegmatis. The iron concentration decreased over time in phagosomes of macrophages infected with Mycobacterium smegmatis but increased in those infected with pathogenic mycobacteria. Autoradiography of infected macrophages incubated with (59)Fe-loaded transferrin demonstrated that the bacteria could acquire iron delivered via the endocytic route, confirming the results obtained in the x-ray microscopy. In addition, the concentrations of chlorine, calcium, potassium, manganese, copper, and zinc were shown to differ between the vacuole of pathogenic mycobacteria and M. smegmatis. Differences in the concentration of several elements between M. avium and M. tuberculosis vacuoles were also observed. Activation of macrophages with recombinant IFN-gamma or TNF-alpha before infection altered the concentrations of elements in the phagosome, which was not observed in cells activated following infection. Siderophore knockout M. tuberculosis vacuoles exhibited retarded acquisition of iron compared with phagosomes with wild-type M. tuberculosis. This is a unique approach to define the environmental conditions within the pathogen containing compartment. PMID- 15661909 TI - Ig-independent Ig beta expression on the surface of B lymphocytes after B cell receptor aggregation. AB - In order for humoral immune responses to develop, B cells must be able to recognize, bind, and internalize Ags. These functions are performed by the BCR, which is also responsible for initiating and transducing activation signals necessary for B cell proliferation and differentiation. We have examined surface expression patterns of individual components of the BCR following anti-Ig- and Ag induced aggregation. Specifically, the localization and expression levels of the Ag-binding component, surface Ig (sIg), and the Igbeta component of the Igalpha/Igbeta signaling unit were investigated to determine their individual participation in the internalization and signal transduction. Using primary murine B cells, we found that while >95% of the sIg is internalized following anti-Ig-induced aggregation, 20-30% of Igbeta remains on the surface. These results suggest that sIg and Igbeta may function independently following the initial stages of signal transduction. PMID- 15661910 TI - A20 is a negative regulator of IFN regulatory factor 3 signaling. AB - IFN regulatory factor 3 (IRF-3) is a critical transcription factor that regulates an establishment of innate immune status following detection of viral pathogens. Recent studies have revealed that two IkappaB kinase (IKK)-like kinases, NF kappaB-activating kinase/Traf family member-associated NF-kappaB activator binding kinase 1 and IKK-i/IKKepsilon, are responsible for activation of IRF-3, but the regulatory mechanism of the IRF-3 signaling pathway has not been fully understood. In this study, we report that IRF-3 activation is suppressed by A20, which was initially identified as an inhibitor of apoptosis and inducibly expressed by dsRNA. A20 physically interacts with NF-kappaB-activating kinase/Traf family member-associated NF-kappaB activator-binding kinase 1 and IKK i/IKKepsilon, and inhibits dimerization of IRF-3 following engagement of TLR3 by dsRNA or Newcastle disease virus infection, leading to suppression of the IFN stimulation response element- and IFN-beta promoter-dependent transcription. Importantly, knocking down of A20 expression by RNA interference results in enhanced IRF-3-dependent transcription triggered by the stimulation of TLR3 or virus infection. Our study thus demonstrates that A20 is a candidate negative regulator of the signaling cascade to IRF-3 activation in the innate antiviral response. PMID- 15661911 TI - Functional and molecular analysis of the double-positive stage-specific CD8 enhancer E8III during thymocyte development. AB - Several developmental stage-, subset-, and lineage-specific Cd8 cis-regulatory regions have been identified. These include the E8(III) enhancer, which directs expression in double-positive (DP) thymocytes, and E8(II), which is active in DP cells and CD8(+) T cells. Using a transgenic reporter expression assay, we identified a 285-bp core fragment of the E8(III) enhancer that retains activity in DP thymocytes. In vitro characterization of the core enhancer revealed five regulatory elements that are required for full enhancer activity, suggesting that multiple factors contribute to the developmental stage-specific activity. Furthermore, deletion of E8(III) in the mouse germline showed that this enhancer is required for nonvariegated expression of CD8 in DP thymocytes when E8(II) is also deleted. These results indicate that E8(III) is one of the cis-elements that contribute to the activation of the Cd8a and Cd8b gene complex during T cell development. PMID- 15661913 TI - A common polymorphism in the SFTPD gene influences assembly, function, and concentration of surfactant protein D. AB - Surfactant protein D (SP-D) plays important roles in the host defense against infectious microorganisms and in regulating the innate immune response to a variety of pathogen-associated molecular pattern. SP-D is mainly expressed by type II cells of the lung, but SP-D is generally found on epithelial surfaces and in serum. Genotyping for three single-nucleotide variations altering amino acids in the mature protein in codon 11 (Met(11)Thr), 160 (Ala(160)Thr), and 270 (Ser(270)Thr) of the SP-D gene was performed and related to the SP-D levels in serum. Individuals with the Thr/Thr(11)-encoding genotype had significantly lower SP-D serum levels than individuals with the Met/Met(11) genotype. Gel filtration chromatography revealed two distinct m.w. peaks with SP-D immunoreactivity in serum from Met/Met(11)-encoding genotypes. In contrast, Thr/Thr(11) genotypes lacked the highest m.w. form. A similar SP-D size distribution was found for recombinant Met(11) and Thr(11) expressed in human embryonic kidney cells. Atomic force microscopy of purified SP-D showed that components eluting in the position of the high m.w. peak consist of multimers, dodecamers, and monomers of subunits, whereas the second peak exclusively contains monomers. SP-D from both peaks bound to mannan-coated ELISA plates. SP-D from the high m.w. peak bound preferentially to intact influenza A virus and Gram-positive and Gram-negative bacteria, whereas the monomeric species preferentially bound to isolated LPS. Our data strongly suggest that polymorphic variation in the N-terminal domain of the SP-D molecule influences oligomerization, function, and the concentration of the molecule in serum. PMID- 15661912 TI - A single nucleotide polymorphism on the promoter of eotaxin1 associates with its mRNA expression and asthma phenotypes. AB - Eotaxin1 plays a pivotal role in eosinophil-associated inflammation. Previously, we demonstrated 14 single-nucleotide polymorphisms (SNPs) in the human eotaxin1 gene and the association between the EOT+67G>A allele and the level of IgE. In this study, we investigated the association between the SNPs and plasma eotaxin1 levels, peripheral blood eosinophil counts, and PC20 methacholine values in normal and asthmatic subjects, and the effects of SNPs on the process of eotaxin1 production. The EOT-576C>T and EOT-384A>G polymorphisms and haplotypes (ht1 and ht4) were significantly associated with plasma eotaxin1 levels in the asthmatics (p < 0.001-0.040). The log [plasma eotaxin1] values correlated with the log [serum total IgE] values in the asthmatics and the normal controls (p = 0.012 and p = 0.004, respectively), and with the log [PC20 methacholine] values in the asthmatics (p = 0.014). A DNA-protein complex was formed with EOT-384A>G, but not with the other SNPs of the promoter. The interaction was stronger with the minor allele than with the common allele, and was reduced upon TNF-alpha exposure. TNF alpha-stimulated PBMCs from the asthmatics with the minor allele homozygote expressed significantly lower levels of eotaxin1 mRNA than those from individuals with the common allele. The EOT+67G>A polymorphism, which substitutes alanine with threonine, did not affect eotaxin1 production or activity. Our data suggest that the EOT-384A>G SNP participates in the regulation of eotaxin1 expression by providing a potential binding site for a repressor, and that the ANOVA of EOT 384A>G may predict asthma phenotypes. PMID- 15661914 TI - Signaling through CD14 attenuates the inflammatory response to Borrelia burgdorferi, the agent of Lyme disease. AB - Lyme disease is a chronic inflammatory disorder caused by the spirochetal bacterium, Borrelia burgdorferi. In vitro evidence suggests that binding of spirochetal lipoproteins to CD14, a pattern recognition receptor expressed on monocytes/macrophages and polymorphonuclear cells, is a critical requirement for cellular activation and the subsequent release of proinflammatory cytokines that most likely contribute to symptomatology and clinical manifestations. To test the validity of this notion, we assessed the impact of CD14 deficiency on Lyme disease in C3H/HeN mice. Contrary to an anticipated diminution in pathology, CD14(-/-) mice exhibited more severe and persistent inflammation than did CD14(+/+) mice. This disparity reflects altered gene regulation within immune cells that may engender the higher bacterial burden and serum cytokine levels observed in CD14(-/-) mice. Comparing their in vitro stimulatory activity, live spirochetes, but not lysed organisms, were a potent CD14-independent stimulus of cytokine production, triggering an exaggerated response by CD14(-/-) macrophages. Collectively, our in vivo and in vitro findings support the provocative notion that: 1) pattern recognition by CD14 is entirely dispensable for elaboration of an inflammatory response to B. burgdorferi, and 2) CD14-independent signaling pathways are inherently more destructive than CD14-dependent pathways. Continued study of CD14-independent signaling pathways may provide mechanistic insight into the inflammatory processes that underlie development of chronic inflammation. PMID- 15661915 TI - Monocyte chemoattractant protein-1 and CCR2 interactions are required for IFN alpha/beta-induced inflammatory responses and antiviral defense in liver. AB - IFN-alpha/beta-mediated functions promote production of MIP-1alpha (or CCL3) by mediating the recruitment of MIP-1alpha-producing macrophages to the liver during early infection with murine CMV. These responses are essential for induction of NK cell inflammation and IFN-gamma delivery to support effective control of local infection. Nevertheless, it remains to be established if additional chemokine functions are regulated by IFN-alpha/beta and/or play intermediary roles in supporting macrophage trafficking. The chemokine MCP-1 (or CCL2) plays a distinctive role in the recruitment of macrophages by predominantly stimulating the CCR2 chemokine receptor. Here, we examine the roles of MCP-1 and CCR2 during murine CMV infection in liver. MCP-1 production preceded that of MIP-1alpha during infection and was dependent on IFN-alpha/beta effects for induction. Resident F4/80(+) liver leukocytes were identified as primary IFN-alpha/beta responders and major producers of MCP-1. Moreover, MCP-1 deficiency was associated with a dramatic reduction in the accumulation of macrophages and NK cells, as well as decreased production of MIP-1alpha and IFN-gamma in liver. These responses were also markedly impaired in mice with a targeted disruption of CCR2. Furthermore, MCP-1- and CCR2-deficient mice exhibited increased viral titers and elevated expression of the liver enzyme alanine aminotransferase in serum. These mice also had widespread virus-induced liver pathology and succumbed to infection. Collectively, these results establish MCP-1 and CCR2 interactions as factors promoting early liver inflammatory responses and define a mechanism for innate cytokines in regulation of chemokine functions critical for effective localized antiviral defenses. PMID- 15661916 TI - Myeloperoxidase plays critical roles in killing Klebsiella pneumoniae and inactivating neutrophil elastase: effects on host defense. AB - Activated neutrophils use myeloperoxidase (MPO) to generate an array of potent toxic oxidants. In the current studies we used genetically altered mice deficient in MPO to investigate the role of the enzyme in host defense against the Gram negative bacterium Klebsiella pneumoniae, an important human pathogen. For comparison, we used mice deficient in the antimicrobial molecule, neutrophil elastase (NE). When challenged i.p., mice deficient in either MPO or NE were markedly more susceptible to bacterial infection and death. In vitro studies suggested that MPO impairs the morphology of bacteria in a distinctive way. Of importance, our in vitro studies found that MPO mediated oxidative inactivation of NE, an enzyme that has been widely implicated in the pathogenesis of various tissue-destructive diseases. This pathway of oxidative inactivation may be physiologically relevant, because activated neutrophils isolated from MPO deficient mice exhibited increased elastase activity. Our observations provide strong evidence that MPO, like NE, is a key player in the killing of K. pneumoniae bacteria. They also suggest that MPO may modulate NE to protect the host from the tissue-degrading activity of this proteinase. PMID- 15661917 TI - Surface-expressed TLR6 participates in the recognition of diacylated lipopeptide and peptidoglycan in human cells. AB - Recognition of microbial components by TLR2 requires cooperation with other TLRs. TLR6 has been shown to be required for the recognition of diacylated lipoproteins and lipopeptides derived from mycoplasma and to activate the NF-kappaB signaling cascade in conjunction with TLR2. Human TLR2 is expressed on the cell surface in a variety of cells, including monocytes, neutrophils, and monocyte-derived, immature dendritic cells (iDCs), whereas the expression profile of TLR6 in human cells remains obscure. In this study we produced a function-blocking mAb against human TLR6 and analyzed TLR6 expression in human blood cells and cell lines and its participation in ligand recognition. TLR6 was expressed, although at a lower level than TLR2, on the cell surface in monocytes, monocyte-derived iDCs, and neutrophils, but not on B, T, or NK cells. Confocal microscopic analysis revealed that TLR6 was colocalized with TLR2 at the plasma membrane of monocytes. Importantly, TLR2/6 signaling did not require endosomal maturation, and anti-TLR6 mAb inhibited cytokine production in monocytes and iDCs stimulated with synthetic macrophage-activating lipopeptide-2 or peptidoglycan, indicating that TLR6 recognized diacylated lipopeptide and peptidoglycan at the cell surface. In addition, TLR2 mutants C30S and C36S (Cys(30) and Cys(36) in TLR2 were substituted with Ser), which were expressed intracellularly in HEK293 cells, failed to induce NF-kappaB activation upon macrophage-activating lipopeptide-2 stimulation even in the presence of TLR6. Thus, coexpression of TLR2 and TLR6 at the cell surface is crucial for recognition of diacylated lipopeptide and peptidoglycan and subsequent cellular activation in human cells. PMID- 15661918 TI - VCAM-1 expression on CD8+ cells correlates with enhanced anti-HIV suppressing activity. AB - CD8(+) cells from HIV-infected individuals showing the CD8(+) cell noncytotoxic antiviral response unexpectedly revealed mRNA for VCAM-1, a cell surface molecule found on endothelial cells. Uninfected subjects had undetectable levels of VCAM-1 mRNA in their CD8(+) cells. Flow cytometry analysis showed that up to 12% of the CD8(+) cells from HIV-positive individuals expressed VCAM-1 compared with 0.8% of the CD8(+) cells of HIV-negative individuals. Enrichment of the CD8(+)VCAM-1(+) cell population and subsequent coculture with CD4(+) cells acutely infected with HIV-1 showed that the VCAM-1(+)CD8(+) cells were able to suppress viral replication with 50% less input cells than the unseparated CD8(+) cell population. This study demonstrates, for the first time, the expression of VCAM-1 on CD8(+) cells. Moreover, the CD8(+)VCAM-1(+) cells show enhanced CD8(+) cell noncytotoxic antiviral response activity that could have clinical importance in HIV infection. PMID- 15661920 TI - Leukotriene B4 protects latently infected mice against murine cytomegalovirus reactivation following allogeneic transplantation. AB - Human CMV is often associated with transplant rejection and opportunistic infections such as pneumonia in immunosuppressed patients. Current anti-CMV therapies, although effective, show relatively high toxicity, which seriously limits their long-term use. In this study, we provide evidence that leukotriene B(4) (LTB(4)) plays an important role in the fight against murine CMV (MCMV) infection in vivo. Intravenous administration of 50 and 500 ng/kg/day of LTB(4) to mice infected with a lethal dose of MCMV significantly increases their survival (50 and 70%, respectively), compared with the placebo-treated group (10% of survival). In mice infected with a sublethal dose of MCMV and treated daily with 50 ng/kg/day of LTB(4), the salivary gland viral loads were found to be reduced by 66% compared with the control group. Furthermore, using an allogeneic bone marrow transplantation mouse model, the frequency of MCMV reactivation from latently infected mice was much lower (38%) in LTB(4) (500 ng/kg)-treated mice than in the placebo-treated group (78%). Finally, in experiments using 5 lipoxygenase-deficient mice, MCMV viral loads in salivary glands were found to be higher in animals unable to produce leukotrienes than in the control groups, supporting a role of endogenous 5-lipoxygenase products, possibly LTB(4), in host defense against CMV infection. PMID- 15661919 TI - Prototype Alzheimer's disease vaccine using the immunodominant B cell epitope from beta-amyloid and promiscuous T cell epitope pan HLA DR-binding peptide. AB - Immunization of amyloid precursor protein transgenic mice with fibrillar beta amyloid (Abeta) prevents Alzheimer's disease (AD)-like neuropathology. The first immunotherapy clinical trial used fibrillar Abeta, containing the B and T cell self epitopes of Abeta, as the immunogen formulated with QS21 as the adjuvant in the vaccine. Unfortunately, the clinical trial was halted during the phase II stage when 6% of the participants developed meningoencephalitis. The cause of the meningoencephalitis in the patients that received the vaccine has not been definitively determined; however, analysis of two case reports from the AN-1792 vaccine trial suggest that the meningoencephalitis may have been caused by a T cell-mediated autoimmune response, whereas production of anti-Abeta Abs may have been therapeutic to the AD patients. Therefore, to reduce the risk of an adverse T cell-mediated immune response to Abeta immunotherapy we have designed a prototype epitope vaccine that contains the immunodominant B cell epitope of Abeta in tandem with the synthetic universal Th cell pan HLA DR epitope, pan HLA DR-binding peptide (PADRE). Importantly, the PADRE-Abeta(1-15) sequence lacks the T cell epitope of Abeta. Immunization of BALB/c mice with the PADRE-Abeta(1-15) epitope vaccine produced high titers of anti-Abeta Abs. Splenocytes from immunized mice showed robust T cell stimulation in response to peptides containing PADRE. However, splenocytes from immunized mice were not reactivated by the Abeta peptide. New preclinical trials in amyloid precursor protein transgenic mouse models may help to develop novel immunogen-adjuvant configurations with the potential to avoid the adverse events that occurred in the first clinical trial. PMID- 15661921 TI - Antigen-specific T cells maintain an effector memory phenotype during persistent Trypanosoma cruzi infection. AB - Infection with the protozoan parasite Trypanosoma cruzi is a major cause of morbidity and mortality in Central and South America. Control of acute experimental infection with T. cruzi is dependent on a robust T cell and type 1 cytokine response. However, little evidence exists demonstrating the development and persistence of a potent antiparasite T cell memory response, and there has been much speculation that the majority of the immune response to T. cruzi infection is not directed against the parasite. In this study, we used an experimental mouse model of T. cruzi infection to test both the Ag specificity and the functional and phenotypic characteristics of the responding T cell population. We observed a vigorous antiparasite response from both CD4(+) and CD8(+) T cells that was maintained in the face of persistent infection. T cells from infected mice also proliferated in response to re-exposure to Ag, and CD8(+) T cells underwent spontaneous proliferation when transferred to naive congenic mice, both characteristic of central memory T cells. Interestingly, T cells from infected mice showed significant down-regulation of CD62L, a characteristic associated with an effector memory phenotype. These results suggest that T cells maintained in mice with chronic T. cruzi infection are fully functional memory cells that cannot be easily categorized in the current central/effector memory paradigm. PMID- 15661922 TI - Immune activation of type I IFNs by Listeria monocytogenes occurs independently of TLR4, TLR2, and receptor interacting protein 2 but involves TNFR-associated NF kappa B kinase-binding kinase 1. AB - Type I IFNs are well established antiviral cytokines that have also been shown to be induced by bacteria. However, the signaling mechanisms regulating the activation of these cytokines during bacterial infections remain poorly defined. We show that although Gram-negative bacteria can activate the type I IFN pathway through TLR4, the intracellular Gram-positive bacterium Listeria monocytogenes (LM) can do so independently of TLR4 and TLR2. Furthermore, experiments using genetic mutants and chemical inhibitors suggest that LM-induced type I IFN activation occurs by an intracellular pathway involving the serine-threonine kinase TNFR-associated NF-kappaB kinase (TANK)-binding kinase 1 (TBK1). Interestingly, receptor-interacting protein 2, a component of the recently discovered nucleotide-binding oligomerization domain-dependent intracellular detection pathway, was not involved. Taken together, our data describe a novel signal transduction pathway involving TBK1 that is used by LM to activate type I IFNs. Additionally, we provide evidence that both the LM- and TLR-dependent pathways converge at TBK1 to activate type I IFNs, highlighting the central role of this molecule in modulating type I IFNs in host defense and disease. PMID- 15661924 TI - Opposing and hierarchical roles of leukotrienes in local innate immune versus vascular responses in a model of sepsis. AB - The 5-lipoxygenase (5-LO)-derived leukotrienes (LTs) influence both local innate immunity and vascular responses, but the relative importance of effects on these two processes in sepsis is unknown. In a cecal ligation and puncture model of peritonitis with severe sepsis, 5-LO(-/-) mice showed a reduction in peritoneal neutrophil accumulation and an increase in the number of bacteria in the peritoneal cavity. Despite this impairment of local innate immunity, the null mice exhibited a marked improvement in survival, and this protection was also seen in wild-type animals treated with the LT synthesis inhibitor MK 886. A survival advantage in severe sepsis was also observed in mice treated with the cysteinyl-LT receptor antagonist MK 571, but not with the LTB(4) receptor antagonist CP 105, 696. Protection in the 5-LO(-/-) mice was associated with reduced vascular leak and serum lactate levels. Moreover, wild-type mice treated with MK 571 exhibited less sepsis-induced hypotension. These data demonstrate opposing effects of cysteinyl-LTs on innate immune vs hemodynamic responses, demonstrating protective effects on local immunity and deleterious effects on the vasculature. They also suggest the possible therapeutic utility of targeting vascular events in sepsis with cysteinyl-LT blockade. PMID- 15661923 TI - Expression and activity of beta-defensins and LL-37 in the developing human lung. AB - Immaturity of innate immunity contributes to the increased susceptibility of human neonates to infection. The lung is a major portal of entry for potential pathogens in the neonate, and human beta-defensins (HBDs) and LL-37 participate in pulmonary innate immunity. We hypothesized that these antimicrobial factors would be developmentally regulated, expressed by neonatal pulmonary tissues, and participate in neonatal innate immunity. We found HBD-2 to be the predominant beta-defensin in human neonatal lung. HBD-2 mRNA expression was developmentally regulated, induced by the proinflammatory factor IL-1beta, and decreased by dexamethasone. Additionally, HBD-2 abundance in neonatal tracheal aspirates increased as a function of gestational age. HBD-1 had a lower level of expression compared with HBD-2 and was induced by dexamethasone. HBD-3 and LL-37 messages were not detected in airway epithelial cultures. Additionally, each antimicrobial peptide exhibited a unique spectrum of antimicrobial activity and salt sensitivity against bacteria commonly causing sepsis in the neonate. Lower levels of HBD-2 may be one factor contributing to the increased susceptibility of premature infants to pulmonary infections. PMID- 15661925 TI - A chemotactic peptide from laminin alpha 5 functions as a regulator of inflammatory immune responses via TNF alpha-mediated signaling. AB - Tissue injury triggers inflammatory responses that may result in release of degradation products or exposure of cryptic domains of extracellular matrix components. Previously, we have shown that a cryptic peptide (AQARSAASKVKVSMKF) in the alpha-chain of laminin-10 (alpha5beta1gamma1), a prominent basement membrane component, is chemotactic for both neutrophils (PMNs) and macrophages (Mphis) and induces matrix metalloproteinase-9 (MMP-9) production. To determine whether AQARSAASKVKVSMKF has additional effects on inflammatory cells, we performed microarray analysis of RNA from RAW264.7 Mphis stimulated with AQARSAASKVKVSMKF. Several cytokines and cytokine receptors were increased >3-fold in response to the laminin alpha5 peptide. Among these were TNF-alpha and one of its receptors, the p75 TNFR (TNFR-II), increasing 3.5- and 5.7-fold, respectively. However, the peptide had no effect on p55 TNFR (TNFR-I) expression. Corroborating the microarray data, the protein levels of TNF-alpha and TNFR-II were increased following stimulation of RAW264.7 cells with AQARSAASKVKVSMKF. In addition, we determined that the production of TNF-alpha and TNFR-II in response to AQARSAASKVKVSMKF preceded the production of MMP-9. Furthermore, using primary Mphis from mice deficient in TNFR-I, TNFR-II, or both TNF-alpha receptors (TNFRs), we determined that AQARSAASKVKVSMKF induces MMP-9 expression by Mphis through a pathway triggered by TNFR-II. However, TNF-alpha signaling is not required for AQARSAASKVKVSMKF-induced PMN release of MMP-9 or PMN emigration. These data suggest that interactions of inflammatory cells with basement membrane components may orchestrate immune responses by inducing expression of cytokines, recruitment of inflammatory cells, and release of proteinases. PMID- 15661926 TI - Sle1ab mediates the aberrant activation of STAT3 and Ras-ERK signaling pathways in B lymphocytes. AB - The Sle1ab genomic interval on murine chromosome 1 mediates the loss of immune tolerance to chromatin resulting in antinuclear Abs (ANA) production in the lupus prone NZM2410 mouse. Global gene expression analysis was used to identify the molecular pathways that are dysregulated at the initiation of B lymphocyte autoimmunity in B6.Sle1ab mice. This analysis identified that STAT3 and ras-ERK signaling pathways are aberrantly activated in Sle1ab B lymphocytes, consistent with increased production of IL-6 by splenic B lymphocytes and monocytes in B6.Sle1ab mice. In vitro treatment of splenic mononuclear cells isolated from ANA positive Sle1ab mice with anti-IL-6 Ab or AG490, an inhibitor of STAT3 signaling pathway, suppressed ANA production in short-term culture, indicating that this pathway was essential to the production of autoantibodies. In vivo treatment of ANA-positive B6.Sle1ab mice with the ras pathway inhibitor, perillyl alcohol, suppressed the increase of ANA. These findings identify IL-6 as a early key cytokine in Sle1ab-mediated disease development and indicate that the STAT3 and ras-ERK signaling pathways are potential therapeutic targets for treating systemic lupus erythematosus. PMID- 15661927 TI - TLR-induced inflammation in cystic fibrosis and non-cystic fibrosis airway epithelial cells. AB - Cystic fibrosis (CF) is a genetic disease characterized by severe neutrophil dominated airway inflammation. An important cause of inflammation in CF is Pseudomonas aeruginosa infection. We have evaluated the importance of a number of P. aeruginosa components, namely lipopeptides, LPS, and unmethylated CpG DNA, as proinflammatory stimuli in CF by characterizing the expression and functional activity of their cognate receptors, TLR2/6 or TLR2/1, TLR4, and TLR9, respectively, in a human tracheal epithelial line, CFTE29o(-), which is homozygous for the DeltaF508 CF transmembrane conductance regulator mutation. We also characterized TLR expression and function in a non-CF airway epithelial cell line 16HBE14o(-). Using RT-PCR, we demonstrated TLR mRNA expression. TLR cell surface expression was assessed by fluorescence microscopy. Lipopeptides, LPS, and unmethylated CpG DNA induced IL-8 and IL-6 protein production in a time- and dose-dependent manner. The CF and non-CF cell lines were largely similar in their TLR expression and relative TLR responses. ICAM-1 expression was also up regulated in CFTE29o(-) cells following stimulation with each agonist. CF bronchoalveolar lavage fluid, which contains LPS, bacterial DNA, and neutrophil elastase (a neutrophil-derived protease that can activate TLR4), up-regulated an NF-kappaB-linked reporter gene and increased IL-8 protein production in CFTE29o( ) cells. This effect was abrogated by expression of dominant-negative versions of MyD88 or Mal, key signal transducers for TLRs, thereby implicating them as potential anti-inflammatory agents for CF. PMID- 15661928 TI - Decoy receptor 3 increases monocyte adhesion to endothelial cells via NF-kappa B dependent up-regulation of intercellular adhesion molecule-1, VCAM-1, and IL-8 expression. AB - Decoy receptor 3 (DcR3), a soluble receptor for FasL, LIGHT and TL1A, is highly expressed in cancer cells. We show that pretreatment of HUVECs with DcR3 enhances the adhesion of THP-1 and U937 cells and primary monocytes. A similar stimulatory effect of DcR3 on THP-1 adhesion was also observed in human microvascular endothelial cells (HMVECs). Flow cytometry and ELISA showed that DcR3-treated HUVECs exhibited significant increases in ICAM-1 and VCAM-1 expression. We also demonstrate the ability of DcR3 to stimulate the secretion of IL-8 by HUVECs. RT PCR and reporter assays revealed that the expression of adhesion molecules and IL 8 are regulated at the level of gene transcription. Experiments with pyrrolidine dithiocarbamate indicated the involvement of an NF-kappaB signaling pathway. DcR3 was found to induce IkappaB kinase activation, IkappaB degradation, p65 nuclear translocation, and NF-kappaB DNA-binding activity. The enhancement by DcR3 of cell adhesion to HUVECs was not mimicked by the TL1A-Ab, which has been shown in our previous work to be a neutralizing Ab against TL1A, thereby inducing HUVECs angiogenesis. Moreover, DcR3-induced cell adhesion could be detected in human aortic endothelial cells (ECs) in which TL1A expression is lacking. Together, our data demonstrate that DcR3 increases monocyte adhesion to ECs via NF-kappaB activation, leading to the transcriptional up-regulation of adhesion molecules and IL-8 in ECs. This novel action appears not to be due to TL1A neutralization, but occurs through an as yet undefined target(s). This study implicates DcR3 in the relationship between inflammation and cancer development. PMID- 15661929 TI - Tissue transglutaminase-mediated formation and cleavage of histamine-gliadin complexes: biological effects and implications for celiac disease. AB - Celiac disease is an HLA-DQ2-associated disorder characterized by an intestinal T cell response. The disease-relevant T cells secrete IFN-gamma upon recognition of gluten peptides that have been deamidated in vivo by the enzyme tissue transglutaminase (transglutaminase 2 (TG2)). The celiac intestinal mucosa contains elevated numbers of mast cells, and increased histamine secretion has been reported in celiac patients. This appears paradoxical because histamine typically biases T cell responses in the direction of Th2 instead of the Th1 pattern seen in the celiac lesions. We report that histamine is an excellent substrate for TG2, and it can be efficiently conjugated to gluten peptides through TG2-mediated transamidation. Histamine-peptide conjugates do not exert agonistic effects on histamine receptors, and scavenging of biologically active histamine by gluten peptide conjugation can have physiological implications and may contribute to the mucosal IFN-gamma response in active disease. Interestingly, TG2 is able to hydrolyze the peptide-histamine conjugates when the concentrations of substrates are lowered, thereby releasing deamidated gluten peptides that are stimulatory to T cells. PMID- 15661930 TI - Cutaneous antigen priming via gene gun leads to skin-selective Th2 immune inflammatory responses. AB - It is becoming increasingly evident that the compartmentalization of immune responses is governed, in part, by tissue-selective homing instructions imprinted during T cell differentiation. In the context of allergic diseases, the fact that "disease" primarily manifests in particular tissue sites, despite pervasive allergen exposure, supports this notion. However, whether the original site of Ag exposure distinctly privileges memory Th2 immune-inflammatory responses to the same site, while sparing remote tissue compartments, remains to be fully investigated. We examined whether skin-targeted delivery of plasmid DNA encoding OVA via gene-gun technology in mice could generate allergic sensitization and give rise to Th2 effector responses in the skin as well as in the lung upon subsequent Ag encounter. Our data show that cutaneous Ag priming induced OVA specific serum IgE and IgG1, robust Th2-cytokine production, and late-phase cutaneous responses and systemic anaphylactic shock upon skin and systemic Ag recall, respectively. However, repeated respiratory exposure to aerosolized OVA failed to instigate airway inflammatory responses in cutaneous Ag-primed mice, but not in mice initially sensitized to OVA via the respiratory mucosa. Importantly, these contrasting airway memory responses correlated with the occurrence of Th2 differentiation events at anatomically separate sites: indeed cutaneous Ag priming resulted in Ag-specific proliferative responses and Th2 differentiation in skin-, but not thoracic-, draining lymph nodes. These data indicate that Ag exposure to the skin leads to Th2 differentiation within skin draining lymph nodes and subsequent Th2 immunity that is selectively manifested in the skin. PMID- 15661931 TI - The Salmonella pathogenicity island (SPI)-2 and SPI-1 type III secretion systems allow Salmonella serovar typhimurium to trigger colitis via MyD88-dependent and MyD88-independent mechanisms. AB - Salmonella typhimurium can colonize the gut, invade intestinal tissues, and cause enterocolitis. In vitro studies suggest different mechanisms leading to mucosal inflammation, including 1) direct modulation of proinflammatory signaling by bacterial type III effector proteins and 2) disruption or penetration of the intestinal epithelium so that penetrating bacteria or bacterial products can trigger innate immunity (i.e., TLR signaling). We studied these mechanisms in vivo using streptomycin-pretreated wild-type and knockout mice including MyD88(-/ ) animals lacking an adaptor molecule required for signaling via most TLRs. The Salmonella SPI-1 and the SPI-2 type III secretion systems (TTSS) contributed to inflammation. Mutants that retain only a functional SPI-1 (M556; sseD::aphT) or a SPI-2 TTSS (SB161; DeltainvG) caused attenuated colitis, which reflected distinct aspects of the colitis caused by wild-type S. typhimurium: M556 caused diffuse cecal inflammation that did not require MyD88 signaling. In contrast, SB161 induced focal mucosal inflammation requiring MyD88. M556 but not SB161 was found in intestinal epithelial cells. In the lamina propria, M556 and SB161 appeared to reside in different leukocyte cell populations as indicated by differential CD11c staining. Only the SPI-2-dependent inflammatory pathway required aroA-dependent intracellular growth. Thus, S. typhimurium can use two independent mechanisms to elicit colitis in vivo: SPI-1-dependent and MyD88-independent signaling to epithelial cells and SPI-2-dependent intracellular proliferation in the lamina propria triggering MyD88-dependent innate immune responses. PMID- 15661932 TI - Protein kinase C alpha-mediated chemotaxis of neutrophils requires NF-kappa B activity but is independent of TNF alpha signaling in mouse skin in vivo. AB - Protein kinase C (PKC) isoforms are major regulators of cutaneous homeostasis and mediate inflammation in response to 12-O-tetradecanoylphorbol-13-acetate (TPA). We have previously reported that transgenic mice overexpressing PKCalpha in the skin exhibit severe intraepidermal neutrophilic inflammation and keratinocyte apoptosis when treated topically with TPA. Activation of PKCalpha increases the production of TNFalpha and the transcription of chemotactic factors (MIP-2, KC, S100A8/A9), vascular endothelial growth factor, and GM-CSF in K5-PKCalpha keratinocytes. In response to PKCalpha activation, NF-kappaB translocates to the nucleus and this is associated with IkappaB phosphorylation and degradation. Preventing IkappaB degradation reduces both the expression of inflammation associated genes and chemoattractant release. To determine whether TNFalpha mediated NF-kappaB translocation and subsequent expression of proinflammatory factors, K5-PKCalpha mice were treated systemically with a dimeric soluble form of p75 TNFR (etanercept) or crossed with mice deficient for both TNFR isoforms, and keratinocytes were cultured in the presence of TNFalpha-neutralizing Abs. The in vivo treatment and TNFR deficiency did not prevent inflammation, and the in vitro treatment did not prevent NF-kappaB nuclear translocation after TPA. Together these results implicate PKCalpha as a regulator of a subset of cutaneous cytokines and chemokines responsible for intraepidermal inflammation independent of TNFalpha. PKCalpha inhibition may have therapeutic benefit in some human inflammatory skin disorders. PMID- 15661934 TI - NF-kappa B activation is not required for Chlamydia trachomatis inhibition of host epithelial cell apoptosis. AB - Chlamydia trachomatis, an obligate intracellular bacterial species, is known to inhibit host cell apoptosis. However, the chlamydial antiapoptotic mechanism is still not clear. Because NF-kappaB activation is antiapoptotic, we tested the potential role of NF-kappaB activation in chlamydial antiapoptotic activity in the current study. First, no obvious NF-kappaB activation was detected in the chlamydia-infected cells when these cells were resistant to apoptosis induced via either the intrinsic or extrinsic apoptosis pathways. Second, inhibition of NF kappaB activation with pharmacologic reagents failed to block the chlamydial antiapoptotic activity. Finally, NF-kappaB p65 gene deletion did not prevent chlamydia from inhibiting host cell apoptosis. These observations together have demonstrated that NF-kappaB activation is not required for the chlamydial antiapoptotic activity. PMID- 15661933 TI - A chemokine-dependent stromal induction mechanism for aberrant lymphocyte accumulation and compromised lymphatic return in rheumatoid arthritis. AB - According to the current model for tissue-specific homing, specificity is conferred by the selective recruitment of lymphocyte populations from peripheral blood, based on their expression of chemokine and adhesion receptors (endothelial selection). In this study, we provide evidence for an alternative stromal induction mechanism that operates in chronic inflammation. We show that the human rheumatoid synovial microenvironment directly induces functional inflammatory (CCR5 and CXCR3) and constitutive (CCR7 and CXCR4) chemokine receptors on infiltrating CD4(+) T cells. Expression of the corresponding inflammatory chemokine ligands (CCL5 and CXCL11) was confined to stromal areas in the synovium. However, expression of the constitutive ligands (CCL19 and CXCL12) was inappropriately high on both vascular and lymphatic endothelium, suggesting that the vascular to lymphatic chemokine gradient involved in lymphatic recirculation becomes subverted in the rheumatoid synovium. These results challenge the view that leukocyte trafficking is regulated solely by selective recruitment of pre existing chemokine receptor-positive cells from peripheral blood, by providing an alternative explanation based on aberrant lymphocyte retention and compromised lymphatic return. PMID- 15661936 TI - Profiling blood lymphocyte interactions with cancer cells uncovers the innate reactivity of human gamma delta T cells to anaplastic large cell lymphoma. AB - Quantifying the contacts that circulating lymphocytes have with cancer cells is useful, because their deficit favors malignancy progression. All normal lymphocytes contact, scan, and acquire membrane fragments (trogocytosis) from foreign cells for their immunosurveillance. So in this study, we used the in vitro trogocytosis of PKH67-stained cancer cell lines as a measure of their interactions with bulks of PBMC freshly isolated from healthy donors. Allogeneic PBMC mixed and coincubated in vitro for 1 h did not trogocytosis, whereas in the same conditions CD20(+), CD4(+), CD8(+), gammadelta T, and CD16(+) PBMC interacted strongly with the cancer cells. Although most unprimed lymphoid effectors of innate (NK) and adaptive (B and T) immunity from healthy donors spontaneously trogocytosed different tumoral cell lines, some carcinoma cell lines could escape them in the coculture. This also uncovered the strong interactions of circulating Vgamma9/Vdelta2(+) central memory gammadelta T cells with anaplastic large cell lymphoma. These interaction profiles were stable upon time for healthy blood donors but were different with other tumors and blood donors. This profiling provides interaction signatures for the immunomonitoring of cancer. PMID- 15661935 TI - Naturally acquired MAGE-A10- and SSX-2-specific CD8+ T cell responses in patients with hepatocellular carcinoma. AB - Immunotherapy is being proposed to treat patients with hepatocellular carcinoma (HCC). However, more detailed knowledge on tumor Ag expression and specific immune cells is required for the preparation of highly targeted vaccines. HCC express a variety of tumor-specific Ags, raising the question whether CTL specific for such Ags exist in HCC patients. Indeed, a recent study revealed CTLs specific for two cancer-testis (CT) Ags (MAGE-A1 and MAGE-A3) in tumor infiltrating lymphocytes of HCC patients. Here we assessed the presence of T cells specific for additional CT Ags: MAGE-A10, SSX-2, NY-ESO-1, and LAGE-1, which are naturally immunogenic as demonstrated in HLA-A2(+) melanoma patients. In two of six HLA-A2(+) HCC patients, we found that MAGE-A10- and/or SSX-2 specific CD8(+) T cells naturally responded to the disease, because they were enriched in tumor lesions but not in nontumoral liver. Isolated T cells specifically and strongly killed tumor cells in vitro, providing evidence that these CTL were selected in vivo for high avidity Ag recognition. Therefore, besides melanoma, HCC is the second solid human tumor with clear evidence for in vivo tumor recognition by T cells, providing the rational for specific immunotherapy, based on immunization with CT Ags such as MAGE-A10 and SSX-2. PMID- 15661937 TI - CCL18 is expressed in atopic dermatitis and mediates skin homing of human memory T cells. AB - CCL18 is a human chemokine secreted by monocytes and dendritic cells. The receptor for CCL18 is not yet known and the functions of this chemokine on immune cells are not fully elucidated. In this study, we describe that CCL18 is present in skin biopsies of atopic dermatitis (AD) patients but not in normal or psoriatic skin. CCL18 was specifically expressed by APCs in the dermis and by Langerhans and inflammatory dendritic epidermal cells in the epidermis. In addition, the serum levels of CCL18 and the percentages of CCL18-producing monocyte/macrophages and dendritic cells were significantly increased in AD patients compared with healthy controls. Furthermore, we demonstrate that CCL18 binds to CLA(+) T cells in peripheral blood of AD patients and healthy individuals and induces migration of AD-derived memory T cells in vitro and in human skin-transplanted SCID mice. These findings highlight a unique role of CCL18 in AD and reveal a novel function of this chemokine mediating skin homing of a subpopulation of human memory T cells. PMID- 15661938 TI - Cathepsin S is required for murine autoimmune myasthenia gravis pathogenesis. AB - Because presentation of acetylcholine receptor (AChR) peptides to T cells is critical to the development of myasthenia gravis, we examined the role of cathepsin S (Cat S) in experimental autoimmune myasthenia gravis (EAMG) induced by AChR immunization. Compared with wild type, Cat S null mice were markedly resistant to the development of EAMG, and showed reduced T and B cell responses to AChR. Cat S null mice immunized with immunodominant AChR peptides showed weak responses, indicating failed peptide presentation accounted for autoimmune resistance. A Cat S inhibitor suppressed in vitro IFN-gamma production by lymph node cells from AChR-immunized, DR3-bearing transgenic mice. Because Cat S null mice are not severely immunocompromised, Cat S inhibitors could be tested for their therapeutic potential in EAMG. PMID- 15661939 TI - IFN-gamma and IL-10 mediate parasite-specific immune responses of cord blood cells induced by pregnancy-associated Plasmodium falciparum malaria. AB - Available evidence suggests that immune cells from neonates born to mothers with placental Plasmodium falciparum (Pf) infection are sensitized to parasite Ag in utero but have reduced ability to generate protective Th1 responses. In this study, we detected Pf Ag-specific IFN-gamma(+) T cells in cord blood from human neonates whose mothers had received treatment for malaria or who had active placental Pf infection at delivery, with responses being significantly reduced in the latter group. Active placental malaria at delivery was also associated with reduced expression of monocyte MHC class I and II in vivo and following short term in vitro coculture with Pf Ag compared with levels seen in neonates whose mothers had received treatment during pregnancy. Given that APC activation and Th1 responses are driven in part by IFN-gamma and down-regulated by IL-10, we examined the role of these cytokines in modulating the Pf Ag-specific immune responses in cord blood samples. Exogenous recombinant human IFN-gamma and neutralizing anti-human IL-10 enhanced T cell IFN-gamma production, whereas recombinant human IFN-gamma also restored MHC class I and II expression on monocytes from cord blood mononuclear cells cocultured with Pf Ag. Accordingly, active placental malaria at delivery was associated with increased frequencies of Pf Ag-specific IL-10(+)CD4(+) T cells in cord blood mononuclear cell cultures from these neonates. Generation and maintenance of IL-10(+) T cells in utero may thus contribute to suppression of APC function and Pf Ag-induced Th1 responses in newborns born to mothers with placental malaria at delivery, which may increase susceptibility to infection later in life. PMID- 15661940 TI - IFN-gamma at the site of infection determines rate of clearance of infection in cryptococcal meningitis. AB - In animal models, immunity to cryptococcal infection, as in many chronic fungal and bacterial infections, is associated with a granulomatous inflammatory response, intact cell-mediated immunity, and a Th1 pattern of cytokine release. To examine the correlates of human immunity to cryptococcal infection in vivo, we analyzed immune parameters at the site of infection over time and assessed the rate of clearance of infection by serial quantitative cerebrospinal fluid (CSF) fungal cultures in 62 patients in a trial of antifungal therapy for HIV associated cryptococcal meningitis. CSF IL-6, IFN-gamma, TNF-alpha, and IL-8 were significantly higher in survivors compared with nonsurvivors. There were negative correlations between log TNF-alpha, IFN-gamma, and IL-6 levels and baseline cryptococcal CFU. Log IFN-gamma, G-CSF, TNF-alpha, and IL-6 were correlated positively with the rate of fall in log CFU/ml CSF/day. In a linear regression model including antifungal treatment group, baseline CFU, and these cytokines, only treatment group and log IFN-gamma remained independently associated with rate of clearance of infection. The results provide direct in vivo evidence for the importance of quantitative differences in IFN-gamma secretion in human immune control of granulomatous infections, and increase the rationale for adjunctive IFN-gamma in the treatment of refractory HIV-associated cryptococcosis. PMID- 15661942 TI - History of the American Institute of Ultrasound in Medicine. PMID- 15661941 TI - One NY-ESO-1-derived epitope that promiscuously binds to multiple HLA-DR and HLA DP4 molecules and stimulates autologous CD4+ T cells from patients with NY-ESO-1 expressing melanoma. AB - NY-ESO-1 is expressed by a broad range of human tumors and is often recognized by Abs in the sera of cancer patients with NY-ESO-1-expressing tumors. The NY-ESO-1 gene also encodes several MHC class I- and class II-restricted tumor epitopes recognized by T lymphocytes. In this study we report one novel pan-MHC class II restricted peptide sequence, NY-ESO-1 87-111, that is capable of binding to multiple HLA-DR and HLA-DP4 molecules, including HLA-DRB1*0101, 0401, 0701, and 1101 and HLA-DPB1*0401 and 0402 molecules. We also demonstrate that peptide NY ESO-1 87-111 stimulates Th1-type and Th-2/Th0-type CD4(+) T cells and clones when presented in the context of these HLA-DR and HLA-DP4 molecules. Both bulk CD4(+) T cells and CD4(+) T cell clones were capable of recognizing not only peptide pulsed APCs, but also autologous dendritic cells, either loaded with the NY-ESO-1 protein or transfected with NY-ESO-1 cDNAs. Using IFN-gamma and IL-5 ELISPOT assays and PBL from patients with NY-ESO-1-expressing tumors, we observed the existence of Th1-type circulating CD4(+) T cells recognizing peptide NY-ESO-1 87 111 in the context of HLA-DP4 molecules. Taken together, these data represent the first report of an HLA-DR- and HLA-DP-restricted epitope from a tumor Ag. They also support the relevance of cancer vaccine trials with peptides NY-ESO-1 87-111 in the large number of cancer patients with NY-ESO-1-expressing tumors. PMID- 15661943 TI - Sonographic evaluation of optic nerve diameter in children with raised intracranial pressure. AB - OBJECTIVE: The optic nerve is part of the central nervous system, and a rise in intracranial pressure (ICP) can directly affect the perioptic nerve space, leading to an increase in nerve diameter. Transorbital sonography is a safe and easy-to-perform method to measure optic nerve diameter for rapid diagnosis of increased ICP. METHODS: The optic nerve diameter was measured in 3 individual occasions by transorbital sonography in the transverse view, 3 mm posterior to the papilla in both eyes, and the mean was calculated. Two groups were examined: a control group of children with normal ICP and normal ophthalmologic and optic nerve examination results and a case group with increased ICP as determined by an alternative method. RESULTS: The sample consisted of 156 children, of which 78 (50%) were in the case group and 78 (50%) were in the control group. Eighty-four subjects (53.7%) were male, and 72 (46.1%) were female. The mean +/- SD ages were 6.9 +/- 5.6 years in the case group and 6.8 +/- 5.5 years in the control group. The mean optic nerve diameters were 5.6 +/- 0.6 mm (range, 4.55 +/- 0.7 to 7.6 +/ 0.6 mm) in the case group and 3.3 +/- 0.6 mm (range, 2 +/- 0.6 to 4.35 +/- 0.6 mm) in the control group. The difference in the means was significant at P < .001. CONCLUSIONS: Optic nerve diameter, measured by transorbital sonography, was significantly greater in pediatric patients with increased ICP compared with a control group. Transorbital sonography can be used to identify pediatric patients with raised ICP. PMID- 15661944 TI - The ratio of flow velocities in the middle cerebral and internal carotid arteries for the prediction of cerebral palsy in term neonates. AB - OBJECTIVE: This study evaluated whether the ratio of the mean flow velocities in the middle cerebral artery (MCA) and the internal carotid artery (ICA) of neonates in the first days of life can be used to identify future neurodevelopmental disabilities. METHODS: We observed 127 term neonates without congenital malformations, chromosomal aberrations, intracranial hemorrhage, or early onset sepsis. The mean cerebral blood flow velocities were measured in the right and left ICAs and in the right and left MCAs with a Doppler flowmeter once from day 1 to day 3. The Vm ratio was defined as the mean velocity in the right and left MCAs/mean velocity in the right and left ICAs. Neurologic examinations were performed at 12 months of age in the outpatient follow-up clinic to detect cerebral palsy (CP), and the subjects were divided into 4 groups according to the diagnosis of hypoxic-ischemic encephalopathy (HIE) and neurologic prognosis: HIE- and normal neurologic function, HIE- and CP, HIE+ and normal neurologic function, and HIE+ and CP. RESULTS: The Vm ratio in infants with the HIE- diagnosis and CP was significantly lower than that in infants with the HIE- diagnosis without CP (P < .05). There was no significant difference between the Vm ratios in infants with the HIE+ diagnosis without CP and infants with the HIE+ diagnosis and CP. CONCLUSIONS: The Vm ratio might be a useful index in estimating neurologic outcome at birth, especially in neonates without the diagnosis of HIE. PMID- 15661945 TI - Right-to-left shunt evaluated at the aortic arch by contrast-enhanced transesophageal echocardiography. AB - OBJECTIVE: The right-to-left shunt (RLS) is diagnosed by contrast-enhanced transesophageal echocardiographic monitoring of the bilateral atria (cTEE-BA). However, the procedure is often disturbed by nonsmoke spontaneous individual contrast (NSSIC) with fast motion, which appears in the left atrium after respiratory maneuvers without administration of a contrast medium and moves past in several seconds. We attempted to perform cTEE monitoring of the aortic arch (cTEE-AA) for evaluation of the RLS and compared the findings with those of cTEE BA. METHODS: Both cTEE-BA and cTEE-AA were performed in 168 patients with ischemic stroke (133 men and 35 women; mean age +/- SD, 62.0 +/- 14.4 years). The frequency of NSSIC in the left atrium was compared with that in the aortic arch during the respiratory maneuver. When contrast much brighter than the NSSIC was visualized in the left atrium and the aortic arch during the respiratory maneuver with administration of the contrast medium, we considered the RLS to be positive in the cTEE-BA and cTEE-AA, respectively. Findings were then compared between the 2 examinations. RESULTS: Nonsmoke spontaneous individual contrast was more frequently observed in the left atrium than the aortic arch (61.3% versus 14.9%; chi2 test, P < .0001). The RLS was positive in 34 patients in the cTEE-BA and in 39 patients in the cTEE-AA. The sensitivity and specificity of the cTEE-AA for the cTEE-BA were 100% and 96.3%, respectively. CONCLUSIONS: The cTEE-AA may be an alternative method for detection of an RLS, especially in patients with a large amount of NSSIC in the left atrium. PMID- 15661946 TI - Characterization of breast masses with sonography: can biopsy of some solid masses be deferred? AB - OBJECTIVE: To determine whether sonography can be used to categorize some solid breast masses as probably benign so that biopsy can be deferred. METHODS: We prospectively characterized 844 sonographically visible solid breast masses referred for biopsy. Mammographic and sonographic features of the masses were recorded, and all masses were categorized by American College of Radiology Breast Imaging Reporting and Data System classification before biopsy. Of the 844 masses, 148 were categorized as probably benign (Breast Imaging Reporting and Data System category 3). Sonographically guided biopsy (n = 804) or fine-needle aspiration (n = 40) was performed for pathologic correlation. RESULTS: Of the 148 masses that met the sonographic criteria for probably benign masses, there was 1 malignancy, for a negative predictive value of 99.3%. CONCLUSIONS: Follow-up can be an acceptable alternative to biopsy for sonographically probably benign solid masses. PMID- 15661947 TI - Effect of total abdominal hysterectomy on ovarian blood supply in women of reproductive age. AB - OBJECTIVE: The purpose of this study was to evaluate the effect of total abdominal hysterectomy on ovarian blood supply using transvaginal color Doppler ultrasonography in women of reproductive age. METHODS: This prospective study included 61 women aged 40 years or younger who were divided into 2 groups: group 1, comprising 31 patients who underwent total abdominal hysterectomy (TAH); and group 2, comprising 30 women with no abnormalities. Inclusion criteria included normal ovarian function at baseline, with basal follicle-stimulating hormone levels of less than 15 mUI/mL, normal body weight, no tobacco use, and no history of laparotomy or ovarian disease. Ovarian arterial blood supply by determination of the pulsatility index (PI) on Doppler analysis and ovarian volume on transvaginal ultrasonography were assessed at baseline and at 6 and 12 postoperative months. The Student t test, profile analysis, and Friedman and Mann Whitney tests were used in the statistical analysis of data. RESULTS: Statistical analysis of baseline data revealed that both groups were homogeneous. At months 6 and 12, greater ovarian volumes and lower PI values were observed in patients who underwent TAH (P < .05). By the end of the study, in 8 of the 31 patients who underwent TAH (25.5%), benign ovarian cysts were observed. In the control group, all the parameters studied remained unchanged. CONCLUSIONS: The reduced PI values observed on Doppler ultrasonography suggested a decrease in the resistance flow in the ovarian arteries in women of reproductive age who underwent TAH. PMID- 15661948 TI - Screening sonography in pregnant patients with blunt abdominal trauma. AB - OBJECTIVE: The purpose of this study was to evaluate the accuracy of screening sonography for the detection of clinically significant abdominal injury in pregnant patients with blunt trauma. METHODS: We retrospectively reviewed the records of 5173 patients with blunt abdominal trauma who underwent screening sonography. Pregnant patients were identified, and the prospective sonographic interpretations were compared with surgical findings, computed tomography (CT), subsequent sonography, cystography, and the clinical course. RESULTS: Of 1567 female patients with trauma, 947 were of reproductive age and, 102 (11%) of these 947 were pregnant. One patient was excluded because a truth standard was not available. Five (5%) of these 101 patients were found to have injuries at surgery. These injuries involved the placenta (2 injuries), spleen (2 injuries), liver (1 injury), and kidney (1 injury); all required surgery. Initial sonographic findings were positive in 4 of 5 patients with injuries. The missed injury was a placental injury detected 15 hours after screening sonography because of fetal bradycardia. After screening sonography, 6 patients underwent additional abdominal imaging: CT (3 patients), cystography (1 patient), and additional sonography (2 patients). Of 101 patients, 95 (94%) required no additional tests, and 97 (96%) required no test involving ionizing radiation. No pregnant patient underwent diagnostic peritoneal lavage. Sensitivity was 80% (95% confidence interval, 28%-100%), and specificity was 100% (96 of 96; 95% confidence interval, 96%-100%) for detecting major abdominal injury. CONCLUSIONS: Sonography is an effective screening examination that can obviate more hazardous tests such as CT, cystography, and peritoneal lavage in most pregnant patients with trauma requiring objective evaluation of the abdomen. PMID- 15661949 TI - Assessment of umbilical cord coiling during the routine fetal sonographic anatomic survey in the second trimester. AB - OBJECTIVE: The purpose of this study was to evaluate the sonographic accuracy to determine the umbilical coiling index (UCI) during the routine fetal anatomic survey in the second trimester. METHODS: In 300 consecutive women with singleton pregnancies and absence of gross fetal anomalies who had a routine second trimester fetal anatomic survey, a distance between 2 pairs of coils was measured from the longitudinal images of the umbilical cord, and the antenatal UCI (aUCI) was calculated. The aUCI was compared with true UCI results obtained after birth. RESULTS: Two hundred thirty-six patients had adequate sonographic umbilical cord images, and all required demographic, antenatal, and labor data collection to meet the inclusion criteria. A statistically significant correlation between aUCI and true UCI was found (P < .0001; r = 0.643). The mean aUCI was 0.402 (80% confidence interval, 0.382), and the true UCI at birth was 0.203 (80% confidence interval, 0.176). The sonographic evaluation showed 12.3% and 8.9% of hypocoiled and hypercoiled cords, whereas evaluation at birth found 10.6% and 8.1% hypocoiled and hypercoiled umbilical cords, respectively. The sensitivity values of sonography to predict hypocoiling and hypercoiling at birth were 78.9% and 25.4%, respectively. CONCLUSIONS: A sonographic evaluation of umbilical cord coiling in the second trimester correlates with the true UCI at birth, although the sensitivity in predicting coiling patterns as hypocoiled and hypercoiled cords is less accurate. A difference between the aUCI and matched UCI at birth could be explained by a sonographic error in the sampling of different umbilical cord segments with discordant coiling patterns or the possibility of a dynamically evolving UCI with advancing gestational age. PMID- 15661950 TI - Clinical implications of the prenatal sonographic finding of fetal myocardial echogenic foci. AB - OBJECTIVE: The purpose of this study was to investigate the clinical implications of fetal echogenic foci limited to the myocardium of the ventricular wall (EMF) or intraventricular septum detected during prenatal sonography. METHODS: A retrospective review of fetuses found to have EMF during prenatal sonography from April 1990 through December 2003 was undertaken. Videotapes and reports were reviewed to determine the indication for sonography, gestational age, location, sonographic characteristics of the masses, and additional fetal findings. Sources of outcome data included neonatal and pediatric records and echocardiographic reports. RESULTS: Five fetuses were identified. Indications for sonographic evaluation included fetal growth (n = 4) and follow-up twin-twin transfusion syndrome (n = 1). Mean gestational age +/- SD at diagnosis was 23.7 +/- 8.0 weeks (range, 17.1-36.4 weeks). The EMF identified were within the right ventricular myocardium (n = 2), intraventricular septum (n = 1), left ventricular myocardium (n = 1), and myocardium of the cardiac apex (n = 1). All foci appeared sonographically "bright as bone." The foci ranged in size from 0.1 x 0.1 to 1.2 x 0.9 cm. Neonatal echocardiographic findings were reported as normal for each of the cases. Follow-up examinations available for up to 2.5 years of age showed normal cardiac function. CONCLUSIONS: The prenatal identification of EMF is unlikely to lead to notable short-term pathologic conditions because the fetuses in this investigation had unremarkable neonatal and pediatric echocardiograms and normal cardiac function. PMID- 15661951 TI - Inversion mode: a new volume analysis tool for 3-dimensional ultrasonography. AB - OBJECTIVES: The main goal was to introduce the inversion mode as a new image analysis tool for the examination of fluid-filled structures using 3-dimensional ultrasonography during pregnancy. METHODS: Three-dimensional ultrasonography was performed on fetuses having fluid collections of noncardiac origin. Threshold adjustment was used to visually assign full transparency to voxels that were associated with fluid. A new postprocessing tool, called the inversion mode, was activated to transform this region of interest into opaque voxels. The morphologic appearance of fluid collections and their anatomic relationship to other organs were shown in this manner. RESULTS: Diagnostic features were shown by this technique in several fetuses with problems that included pleural effusion, duodenal atresia, urinary tract abnormalities, and hydrocephaly. Furthermore, the inversion mode also permitted surface reconstruction of an irregular pleural effusion that had close resemblance to results with the Virtual Organ Computer-Aided Analysis rotational slice technique. Acoustic shadowing was also documented as a potential technical limitation of this method. CONCLUSIONS: The inversion mode can display scattered or contiguous fluid-filled structures in ways that can be very difficult or impossible to accurately characterize with conventional ultrasonography. It may be particularly helpful for the evaluation of multiple fluid-filled cysts or irregular fluid collections in the fetus. PMID- 15661952 TI - Middle cerebral artery peak systolic and ductus venosus velocity waveforms in the hydropic fetus. AB - OBJECTIVE: The purpose of this study was to assess whether Doppler assessment of the middle cerebral artery (MCA) peak systolic velocity (PSV) and ductus venosus (DV) velocity waveforms during sonography of hydropic fetuses may specify the cause of fetal hydrops. METHODS: A level II sonographic examination was performed in 16 hydropic fetuses, and the MCA PSV and DV velocity waveforms were assessed. The MCA PSV values divided hydropic fetuses into anemic (group 1) and nonanemic (group 2) fetuses. In group 2 fetuses, the DV was defined as normal or abnormal. Sonographic examination and Doppler assessment of these vessels specified the cause of hydrops and indicated the use of specific investigations for diagnosing the etiology of fetal hydrops. RESULTS: Seven of 16 fetuses had MCA PSV values greater than 1.50 multiples of the median (group 1). Nine of 16 fetuses had normal MCA PSV values (group 2); among them, 7 of 9 had either absent or reversed flow in the DV, and 2 had a normal DV. In group 1, the cause of fetal anemia was investigated by maternal serum tests, and 5 cordocentesis procedures were performed. In group 2, 7 of 9 fetuses had reversed flow in the DV, which suggested a cardiac abnormality confirmed by echocardiography. Five cordocentesis procedures were performed for fetal karyotype, and in 2 fetuses, the cause of hydrops was idiopathic. CONCLUSIONS: Our data suggest that assessment of the MCA PSV and DV velocity waveforms in the hydropic fetus may further our knowledge of the etiology of hydrops and may indicate which investigations among the many available should be used for diagnosing the cause of fetal hydrops. PMID- 15661953 TI - Advantages of color B-mode imaging with contrast optimization in sonography of low-contrast musculoskeletal lesions and structures in the foot and ankle. AB - OBJECTIVE: The purpose of this study was to evaluate the utility of color B-mode imaging with contrast optimization in evaluating low-contrast lesions of the foot and ankle (Morton neuromas and plantar fasciitis). METHODS: The sonographic examinations of 49 consecutive patients with a diagnosis of plantar fasciitis or Morton neuroma imaged with both conventional gray scale imaging and color B-mode imaging with contrast optimization (Photopic; Siemens Medical Solutions, Mountain View, CA) were reviewed. In every patient, matched pairs of images obtained with conventional gray and color maps (Photopic) were acquired and stored as DICOM (Digital Imaging and Communications in Medicine) images on a sonographic workstation. Each image was assessed independently by 2 musculoskeletal radiologists trained in musculoskeletal sonography for overall image contrast (lesion to background), conspicuity of regional tissue boundaries, visualization of deep tissue boundaries, and how well the internal characteristics of the structure were visualized. RESULTS: Three-way analysis of variance showed that Photopic imaging resulted in statistically significantly improved overall image contrast, definition of regional soft tissue boundaries, including deep soft tissue boundaries, and depiction of the internal characteristics of the structure being examined. CONCLUSIONS: Color imaging with contrast optimization improves overall image contrast and better defines deep soft tissue boundaries and the internal morphologic characteristics of Morton neuromas and the plantar fascia compared with conventional gray scale imaging. PMID- 15661954 TI - Rudimentary horn pregnancy: first-trimester prerupture sonographic diagnosis and confirmation by magnetic resonance imaging. AB - OBJECTIVE: Two cases of rudimentary horn pregnancy diagnosed in the first trimester by sonography and confirmed by magnetic resonance imaging (MRI) are reported. We suggest criteria for early, prerupture sonographic diagnosis of this rare condition. METHODS: We report a case in which pregnancy in a rudimentary horn was suspected on routine sonographic examination. In the second case, sonographic examination at 11 weeks' gestation revealed a right unicornuate uterus and a noncommunicating left rudimentary horn containing a gestational sac. In both cases, MRI clearly confirmed the sonographic diagnosis, showing an empty cavity of the uterine body and a pregnant uterine horn without an endometrial communication to the uterine body. RESULTS: Both patients underwent surgery, and the pregnant rudimentary horns were resected with no complications. CONCLUSIONS: We suggest the following criteria for sonographic diagnosis of rudimentary horn pregnancy: (1) a pseudopattern of a asymmetrical bicornuate uterus, (2) absent visual continuity tissue surrounding the gestational sac and the uterine cervix, and (3) the presence of myometrial tissue surrounding the gestational sac. Typical hypervascularization of placenta accreta may support the diagnosis. Additionally, MRI can be used to confirm the diagnosis before an invasive procedure is undertaken. PMID- 15661955 TI - Congenital dacryocystocele: prenatal 2- and 3-dimensional sonographic findings. AB - OBJECTIVE: The purpose of this series is to present our experience with cases of dacryocystocele diagnosed prenatally. The role of prenatal 3-dimensional sonography, as an adjunct to 2-dimensional sonography, in the prenatal assessment of these cases is emphasized. METHODS: A retrospective review of cases was conducted. Information was obtained by reviewing the sonographic reports and medical records. Outcomes were obtained from the referring obstetricians or directly from the parents. RESULTS: Ten fetuses had the diagnosis of a congenital dacryocystocele at a median gestational age of 30.1 weeks (range, 27-33 weeks). In 6 cases, the cystic lesion was unilateral, and in 4 it was bilateral, with a mean largest diameter at the time of diagnosis of 7.5 mm (range, 4-11 mm). There were no other associated findings. Three-dimensional sonography, carried out in 3 cases, clearly depicted the anomaly, the degree of intranasal extension, and swelling below the medial canthal area. Spontaneous resolution was documented prenatally in 5 fetuses, and 1 additional case resolved between the last prenatal scan and the delivery. There were no reported long-term complications associated with this finding, although 1 infant required probing at 2 months of age to resolve the dacryocystocele. CONCLUSIONS: Prenatal diagnosis of dacryocystocele is straightforward. A considerable number of lesions are bilateral, and many resolve in utero spontaneously or neonatally after minimal intervention. For those not resolving by the time of the delivery, ophthalmologic or rhinologic consultation is warranted because of potential complications. Three-dimensional sonography may provide a noninvasive method for evaluating these cystic masses and may contribute to the avoidance of additional diagnostic techniques in the neonatal period. PMID- 15661956 TI - Usefulness of real-time 3-dimensional echocardiography in the assessment of infective endocarditis: initial experience. AB - OBJECTIVE: Our aim was to provide an initial description of the usefulness of real-time 3-dimensional (3D) echocardiography when evaluating patients with infective endocarditis. METHODS: For this purpose, we describe 3 cases in which the usefulness of real-time 3D echocardiography is shown. RESULTS: Better definition of the morphologic characteristics of the vegetation and valvular apparatus was achieved with real-time 3D echocardiography. Furthermore, the information provided by the real-time 3D echocardiogram to the surgical team was accurate and direct because it was not based on detailed verbal information for performing a 3D mental reconstruction but was a simple image similar to what would be found in the operating room. In all cases, the information provided by the real-time 3D echocardiography was confirmed during surgery. CONCLUSIONS: Real time 3D echocardiography is an important tool for aiding diagnosis or for fine tuning a suspected diagnosis when traditional echocardiography is not completely clear for both medical and surgical decision making. PMID- 15661957 TI - Diagnosis of a displacement of Guglielmi detachable coils by transcranial color coded sonography. PMID- 15661958 TI - Prenatal diagnosis of an inguinoscrotal hernia: sonographic and magnetic resonance imaging findings. PMID- 15661959 TI - Increased systolic peak velocity in fetal middle cerebral artery blood flow shown by Doppler sonography in acute fetomaternal hemorrhage. PMID- 15661960 TI - Medulloepithelioma of the ciliary body: ultrasonographic biomicroscopic findings. PMID- 15661961 TI - Entertainment ultrasound. PMID- 15661962 TI - Three-dimensional sonography of a fetus and an intrauterine device in a bicornuate uterus. PMID- 15661963 TI - Cutaneous vasoconstrictor responses to norepinephrine are attenuated in older humans. AB - Cutaneous vasoconstriction (VC) in response to cooling is impaired with human aging. On the basis of previous findings that older humans rely predominantly on norepinephrine (NE) for reflex VC of skin blood vessels, and that the VC effects of NE are blunted with age in many vascular beds, we tested the hypothesis that cutaneous VC responses to exogenous NE are attenuated in aged skin compared with young skin. In 11 young (18-30 yr) and 11 older (62-76 yr) men and women, skin blood flow was monitored at two forearm sites with laser Doppler (LD) flowmetry, while local skin temperature was clamped at 34 degrees C. At one site, five doses of NE (10(-10) to 10(-2) M) were sequentially infused via intradermal microdialysis while the other site served as control (C; Ringer). Cutaneous vascular conductance (CVC; LD flux/mean arterial pressure) was expressed as percent change from baseline (%DeltaCVCbase). At 10(-10), 10(-8), and 10(-6) M NE, older VC responses were attenuated compared with young [10(-10):-35 (95% confidence interval:-16, -52) vs.-49 (-40, -58) %DeltaCVCbase, P=0.02; 10(-8):-38 (-20, -56) vs.-50 (-40, -61) %DeltaCVCbase, P=0.03; 10(-6):-52 (-35, -70) vs.-67 (-60, -74) %DeltaCVCbase, P=0.01]. Older maximal VC responses were also blunted compared with young [-80 (confidence interval:-73,-87) vs.-88 (confidence interval:-87, -90) %DeltaCVCbase, P=0.03]. NE-mediated cutaneous VC is blunted at both physiological and superphysiological doses in older subjects compared with young subjects. Considering that NE is the only functional neurotransmitter mediating reflex VC in aged skin, attenuated NE-mediated VC may further predispose older humans to excess heat loss in the cold. PMID- 15661964 TI - Uteroplacental restriction in the rat impairs fetal growth in association with alterations in placental growth factors including PTHrP. AB - During pregnancy, parathyroid hormone-related protein (PTHrP) is one of many growth factors that play important roles to promote fetal growth and development, including stimulation of placental calcium transport. Angiotensin II, acting through the AT(1a) receptor, is also known to promote placental growth. We examined the effects of bilateral uterine artery and vein ligation (restriction), which mimics placental insufficiency in humans, on growth, intrauterine PTHrP, placental AT(1a), and pup calcium. Growth restriction was surgically induced on day 18 of pregnancy in Wistar-Kyoto female rats by uterine vessel ligation. Uteroplacental insufficiency reduced fetal body weight by 15% and litter size (P < 0.001) compared with the control rats with no effect on placental weight or amniotic fluid volume. Uteroplacental insufficiency reduced placental PTHrP content by 46%, with increases in PTHrP (by 2.6-fold), parathyroid hormone (PTH)/PTHrP receptor (by 11.6-fold), and AT(1a) (by 1.7-fold) relative mRNA in placenta following restriction compared with results in control (P < 0.05). There were no alterations in uterine PTHrP and PTH/PTHrP receptor mRNA expression. Maternal and fetal plasma PTHrP and calcium concentrations were unchanged. Although fetal total body calcium was not altered, placental restriction altered perinatal calcium homeostasis, as evidenced by lower pup total body calcium after birth (P < 0.05). The increased uterine and amniotic fluid PTHrP (P < 0.05) may be an attempt to compensate for the induced impaired placental function. The present study demonstrates that uteroplacental insufficiency alters intrauterine PTHrP, placental AT(1a) expression, and perinatal calcium in association with a reduction in fetal growth. Uteroplacental insufficiency may provide an important model for exploring the early origins of adult diseases. PMID- 15661965 TI - Mechanism of cardiovascular effects of nociceptin microinjected into the nucleus tractus solitarius of the rat. AB - Microinjections (100 nl) of 0.15, 0.31, 0.62, and 1.25 mmol/l of nociceptin into the medial nucleus tractus solitarius (mNTS) elicited decreases in mean arterial pressure (11 +/- 1.8, 20 +/- 2.1, 21.5 +/- 3.1, and 15.5 +/- 1.9 mmHg, respectively) and heart rate (14 +/- 2.7, 29 +/- 5.5, 39 +/- 5.2, and 17.5 +/- 3.1 beats/min, respectively). Because maximal responses were elicited by microinjections of 0.62 mmol/l nociceptin, this concentration was used for other experiments. Repeated microinjections of nociceptin (0.62 mmol/l) into the mNTS, at 20-min intervals, did not elicit tachyphylaxis. Bradycardia induced by microinjections of nociceptin into the mNTS was abolished by bilateral vagotomy. The decreases in mean arterial pressure and heart rate elicited by nociceptin into the mNTS were blocked by prior microinjections of the specific ORL1-receptor antagonist [N-Phe(1)]-nociceptin-(1-13)-NH(2) (9 mmol/l). Microinjections of the ORL1-receptor antagonist alone did not elicit a response. Prior combined microinjections of GABA(A) and GABA(B) receptor antagonists (2 mmol/l gabazine and 100 mmol/l 2-hydroxysaclofen, respectively) into the mNTS blocked the responses to microinjections of nociceptin at the same site. Prior microinjections of ionotropic glutamate receptor antagonists (2 mmol/l NBQX and 5 mmol/l d-AP7) also blocked responses to nociceptin microinjections into the mNTS. These results were confirmed by direct neuronal recordings. It was concluded that 1) nociceptin inhibits GABAergic neurons in the mNTS, 2) GABAergic neurons may normally inhibit the release of glutamate from the terminals of peripheral afferents in the mNTS, and 3) inhibition of GABAergic neurons by nociceptin results in an increase in the release of glutamate in the mNTS, which in turn elicits depressor and bradycardic responses via activation of ionotropic glutamate receptors on secondary mNTS neurons. PMID- 15661966 TI - Cytokine profiles in the testes of rats treated with lipopolysaccharide reveal localized suppression of inflammatory responses. AB - Evidence indicates that the testis possesses a reduced capacity to mount inflammatory and rejection responses, which undoubtedly contributes to the ongoing survival of the highly immunogenic germ cells. The contribution of local cytokine expression to this condition was investigated in adult male rats treated with lipopolysaccharide to induce inflammation. Cytokine mRNA and protein expression were determined in tissue extracts and fluids by Northern blot analysis, quantitative PCR, or RNAse protection assay and specific ELISAs. Testicular expression of the proinflammatory cytokines, interleukin (IL)-1beta and tumor necrosis factor-alpha was considerably attenuated compared with the liver (control tissue); in contrast, the testicular IL-6 response was enhanced. Expression of IL-10, a type 2 immunoregulatory cytokine, was similar in both testis and liver, whereas the immunoregulatory/anti-inflammatory cytokines transforming growth factor-beta(1) and activin A were constitutively elevated in both normal and inflamed testes. The IL-1beta and transforming growth factor beta(1) proteins were present principally in their latent (inactive) forms, indicating that enzymic processing is an important control mechanism for these two cytokines within the testis. These data indicate that inflammatory and regulatory cytokine activity is regulated at both transcriptional and posttranslational levels in a testis-specific manner. It is concluded that a novel pattern of suppression of proinflammatory cytokine responses and normal or elevated expression of immunoregulatory cytokines may be responsible for reduced inflammatory responses and enhanced graft survival in the testis. These data have important implications for the understanding and treatment of male autoimmune infertility, testicular inflammation. and carcinogenesis. PMID- 15661967 TI - Increased osmolality of conscious water-deprived rats supports arterial pressure and sympathetic activity via a brain action. AB - To test the hypothesis that high osmolality acts in the brain to chronically support mean arterial pressure (MAP) and lumbar sympathetic nerve activity (LSNA), the osmolality of blood perfusing the brain was reduced in conscious water-deprived and water-replete rats by infusion of hypotonic fluid via bilateral nonoccluding intracarotid catheters. In water-deprived rats, the intracarotid hypotonic infusion, estimated to lower osmolality by approximately 2%, decreased MAP by 9+/-1 mmHg and LSNA to 86+/-7% of control; heart increased by 25+/-8 beats per minute (bpm) (all P<0.05). MAP, LSNA, and heart rate did not change when the hypotonic fluid was infused intravenously. The intracarotid hypotonic fluid infusion was also ineffective in water-replete rats. Prior treatment with a V1 vasopressin antagonist did not alter the subsequent hypotensive and tachycardic effects of intracarotid hypotonic fluid infusion in water-deprived rats. In summary, acute decreases in osmolality of the carotid blood of water-deprived, but not water-replete, rats decreases MAP and LSNA and increases heart rate. These data support the hypothesis that the elevated osmolality induced by water deprivation acts via a region perfused by the carotid arteries, presumably the brain, to tonically increase MAP and LSNA and suppress heart rate. PMID- 15661968 TI - Frequency encoding in renal blood flow regulation. AB - With a model of renal blood flow regulation, we examined consequences of tubuloglomerular feedback (TGF) coupling to the myogenic mechanism via voltage gated Ca channels. The model reproduces the characteristic oscillations of the two mechanisms and predicts frequency and amplitude modulation of the myogenic oscillation by TGF. Analysis by wavelet transforms of single-nephron blood flow confirms that both amplitude and frequency of the myogenic oscillation are modulated by TGF. We developed a double-wavelet transform technique to estimate modulation frequency. Median value of the ratio of modulation frequency to TGF frequency in measurements from 10 rats was 0.95 for amplitude modulation and 0.97 for frequency modulation, a result consistent with TGF as the modulating signal. The simulation predicted that the modulation was regular, while the experimental data showed much greater variability from one TGF cycle to the next. We used a blood pressure signal recorded by telemetry from a conscious rat as the input to the model. Blood pressure fluctuations induced variability in the modulation records similar to those found in the nephron blood flow results. Frequency and amplitude modulation can provide robust communication between TGF and the myogenic mechanism. PMID- 15661969 TI - Trimethylthiazoline supports conditioned flavor avoidance and activates viscerosensory, hypothalamic, and limbic circuits in rats. AB - Interoceptive stimuli modulate stress responses and emotional state, in part, via ascending viscerosensory inputs to the hypothalamus and limbic forebrain. It is unclear whether similar viscerosensory pathways are recruited by emotionally salient exteroceptive stimuli, such as odors. To address this question, we investigated conditioned avoidance and central c-Fos activation patterns in rats exposed to synthetic trimethylthiazoline (TMT), an odiferous natural component of fox feces. Experiment 1 demonstrated that rats avoid consuming novel flavors that previously were paired with TMT exposure, evidence that TMT supports conditioned flavor avoidance. Experiment 2 examined central neural systems activated by TMT. Odor-naive rats were acutely exposed to low or high levels of TMT or a novel nonaversive control odor and were perfused with fixative 60-90 min later. A subset of rats received retrograde neural tracer injections into the central nucleus of the amygdala (CeA) 7-10 days before odor exposure and perfusion. Brain sections were processed for dual-immunocytochemical detection of c-Fos and other markers to identify noradrenergic (NA) neurons, corticotropin-releasing hormone (CRH) neurons, and retrogradely labeled neurons projecting to the CeA. Significantly greater proportions of medullary and pontine NA neurons, hypothalamic CRH neurons, and CeA-projecting neurons were activated in rats exposed to TMT compared with activation in rats exposed to the nonaversive control odor. Thus the ability of TMT to support conditioned avoidance behavior is correlated with significant odor-induced recruitment of hypothalamic CRH neurons and brain stem viscerosensory inputs to the CeA. PMID- 15661970 TI - Glucocorticoids increase osmotic water permeability (Pf) of neonatal rabbit renal brush border membrane vesicles. AB - During postnatal maturation, there is an increase in renal brush border membrane vesicle (BBMV) osmotic water permeability and a parallel increase in aquaporin-1 (AQP1) protein abundance. The mechanisms responsible for these changes remain unknown. Because serum glucocorticoid levels rise postnatally and have previously been linked to other maturational changes in renal function, we examined the effects of glucocorticoids on osmotic (Pf) and diffusional (P(DW)) water permeability and AQP1 protein abundance of renal BBMV. Neonatal rabbits were treated with dexamethasone (10 microg/100 g) for three days and compared with control neonates and adults. Pf and P(DW) were measured at 20 degrees C with a stopped-flow apparatus using light-scattering and aminonaphthalene trisulfonic acid (ANTS) fluorescence, respectively. Pf was significantly higher in BBMV from dexamethasone-treated neonates compared with vehicle-treated neonates, but remained lower than in BBMV from adults (P<0.05). P(DW) in dexamethasone and vehicle-treated neonatal BBMV was lower than in adult BBMV. Pf/P(DW) ratio increased from neonate (5.1+/-0.3) to dexamethasone (7.0+/-0.1) and adult BBMV (6.3+/-0.1). AQP1 expression was increased by dexamethasone treatment to adult levels. Membrane fluidity, which is inversely related to generalized polarization (GP) of steady-state laurdan fluorescence, was significantly higher in neonatal BBMV than both dexamethasone and adult BBMV (GP: neonate 0.285+/-0.002, dexamethasone treatment 0.302+/-0.006, and adult 0.300+/-0.005; P<0.05). These combined results show that dexamethasone-treatment during days 4-7 of life increases BBMV water permeability despite a decrease in membrane fluidity. This occurs by increasing channel-mediated water transport, as reflected in an increase in AQP1 protein abundance and a higher Pf/P(DW) ratio. This mimics the maturational changes and suggests a physiological role for glucocorticoids in maturation of proximal tubule water transport. PMID- 15661971 TI - Increased blood pressure in the offspring of diabetic mothers. AB - Studies were conducted in rats to determine the effect of maternal diabetes and the consequent hyperglycemia on cardiovascular function in the offspring. Diabetes was induced in pregnant Wistar rats through streptozotocin injection (50 mg/kg). Cardiovascular parameters were measured in 2-mo-old offspring animals of diabetic (OD, n=12) and control rats (OC, n=8). Arterial pressure (AP), heart rate (HR), baroreflex sensitivity, and vascular responsiveness to phenylephrine (PH) and sodium nitroprusside (SN) were measured. Angiotensin-converting enzyme (ACE) activity in heart, kidney, and lung was determined. OD rats exhibited increases in systolic AP (138+/-8 vs. 119+/-6 mmHg, OD vs. OC), with no change in HR (342+/-21 vs. 364+/-39 beats per minute (bpm), OD vs. OC). The reflex tachycardia elicited by SN was reduced in OD rats, as indicated by the slope of the linear regression (-2.2+/-0.4 vs. -3.6+/-0.8 bpm/mmHg, OD vs. OC). Vascular responsiveness to PH was increased 63% in OD rats compared with OC. OD rats showed increases in ACE activity in heart, kidney, and lung (1.13+/-0.24, 3.04+/ 0.86, 40.8+/-8.9 vs. 0.73+/-0.19, 1.7+/-0.45, 28.1+/-6 nmol His-Leu.min-1 mg protein-1, OD vs. OC). Results suggest that diabetes during pregnancy affects cardiovascular function in offspring, seen as hypertension, baroreflex dysfunction, and activation of tissue renin-angiotensin system. PMID- 15661972 TI - Autonomic nervous system and blood pressure regulation in RGS2-deficient mice. AB - Regulator of G protein signaling (RGS2) deletion in mice prolongs signaling by G protein-coupled vasoconstrictor receptors and increases blood pressure. However, the exact mechanism of the increase in blood pressure is unknown. To address this question we tested autonomic nervous system function and blood pressure regulation in RGS2-deficient mice (RGS2-/-). We measured arterial blood pressure and heart rate (HR) with telemetry, computed time and frequency-domain measures for blood pressure and HR variability (HRV) as well as baroreflex sensitivity [BRS-low frequency (LF)], and assessed environmental stress sensitivity. Mean arterial blood pressure (MAP) was approximately 10 mmHg higher in RGS2-/-compared with RGS2+/+mice, while HR was not different between the groups, indicating a resetting of the baroreceptor reflex. Atropine increased MAP more in RGS2-/-than in RGS2+/+mice while HR responses were not different. Urinary norepinephrine excretion was higher in RGS2-/-than in RGS2+/+mice. The blood pressure decrease following prazosin was more pronounced in RGS2-/-mice than in RGS2+/+mice. The LF and high-frequency (HF) power of HRV were reduced in RGS2-/-compared with controls while BRS-LF and SBP-LF were not different. Atropine and atropine+metoprolol markedly reduced the HRV parameters in the time (RMSSD) and frequency domain (LF, HF, LF/HF) in both strains. Environmental stress sensitivity was increased in RGS2-/-mice compared with controls. We conclude that the increase in blood pressure in RGS2-/-mice is not solely explained by peripheral vascular mechanisms. A central nervous system mechanism might be implicated by an increased sympathetic tone. This state of affairs could lead to a baroreceptor-HR reflex resetting, while BRS remains unimpaired. PMID- 15661973 TI - Abrupt and gradual extinction among Late Permian land vertebrates in the Karoo basin, South Africa. AB - The Karoo basin of South Africa exposes a succession of Upper Permian to Lower Triassic terrestrial strata containing abundant terrestrial vertebrate fossils. Paleomagnetic/magnetostratigraphic and carbon-isotope data allow sections to be correlated across the basin. With this stratigraphy, the vertebrate fossil data show a gradual extinction in the Upper Permian punctuated by an enhanced extinction pulse at the Permian-Triassic boundary interval, particularly among the dicynodont therapsids, coinciding with negative carbon-isotope anomalies. PMID- 15661974 TI - Chronic lymphocytic inflammation specifies the organ tropism of prions. AB - Prions typically accumulate in nervous and lymphoid tissues. Because proinflammatory cytokines and immune cells are required for lymphoid prion replication, we tested whether inflammatory conditions affect prion pathogenesis. We administered prions to mice with five inflammatory diseases of the kidney, pancreas, or liver. In all cases, chronic lymphocytic inflammation enabled prion accumulation in otherwise prion-free organs. Inflammatory foci consistently correlated with lymphotoxin up-regulation and ectopic induction of FDC-M1+ cells expressing the normal cellular prion protein PrPC. By contrast, inflamed organs of mice lacking lymphotoxin-alpha or its receptor did not accumulate the abnormal isoform PrPSc, nor did they display infectivity upon prion inoculation. By expanding the tissue distribution of prions, chronic inflammatory conditions may act as modifiers of natural and iatrogenic prion transmission. PMID- 15661975 TI - Photic zone euxinia during the Permian-triassic superanoxic event. AB - Carbon and sulfur isotopic data, together with biomarker and iron speciation analyses of the Hovea-3 core that was drilled in the Perth Basin, Western Australia, indicate that euxinic conditions prevailed in the paleowater column during the Permian-Triassic superanoxic event. Biomarkers diagnostic for anoxygenic photosynthesis by Chlorobiaceae are particularly abundant at the boundary and into the Early Triassic. Similar conditions prevailed in the contemporaneous seas off South China. Our evidence for widespread photiczone euxinic conditions suggests that sulfide toxicity was a driver of the extinction and a factor in the protracted recovery. PMID- 15661976 TI - Global chronic diseases. PMID- 15661977 TI - Planetary science. Titan, once a world apart, becomes eerily familiar. PMID- 15661978 TI - Disaster preparedness. Global tsunami warning system takes shape. PMID- 15661979 TI - Research policy. Facing a revolt, Pasteur board members offer to resign. PMID- 15661981 TI - Teaching evolution. Judge orders stickers removed from Georgia textbooks. PMID- 15661980 TI - Medicine. Low-power mitochondria may raise risk of cardiovascular problems. PMID- 15661982 TI - Paleontology. Fossil count suggests biggest die-off wasn't due to a smashup. PMID- 15661983 TI - Patent law. Inventor knocks Japan's system after settlement. PMID- 15661985 TI - Oceanography. Grim forecast for a fading fleet. PMID- 15661984 TI - Patent law. Shuji Nakamura speaks out. PMID- 15661987 TI - Fred Kavli profile. A physics home away from home. PMID- 15661986 TI - Fred Kavli profile. A new benefactor takes aim at basic scientific questions. PMID- 15661988 TI - Parasitology. Twisted parasites from "outer space" perplex biologists. PMID- 15661989 TI - Indian Ocean tsunami. Using scientific assessments to stave off epidemics. PMID- 15661990 TI - Society for Integrative and Comparative Biology meeting. Scurrying roaches outwit without their brains. PMID- 15661991 TI - Society for Integrative and Comparative Biology meeting. With flippers, two can equal four. PMID- 15661992 TI - Society for Integrative and Comparative Biology meeting. More than one way to dig a tunnel. PMID- 15661993 TI - Revisiting the taxonomic impediment. PMID- 15661994 TI - Public health. Cutting world hunger in half. PMID- 15661996 TI - Geoscience. The boon and bane of radiocarbon dating. PMID- 15661995 TI - Medicine. Treating neurodegenerative diseases with antibiotics. PMID- 15661997 TI - Chemistry. Short and sharp-spectroscopy with frequency combs. PMID- 15661998 TI - Ecology. A leap for lion populations. PMID- 15662000 TI - Genetic factors in type 2 diabetes: the end of the beginning? AB - The intensive search for genetic variants that predispose to type 2 diabetes was launched with optimism, but progress has been slower than was hoped. Even so, major advances have been made in the understanding of monogenic forms of the disease which together represent a substantial health burden, and a few common gene variants that influence susceptibility have now been unequivocally identified. Armed with a better understanding of the tools needed to detect such genes, it seems inevitable that the rate of progress will increase and the relevance of genetic information to the diagnosis, treatment, and prevention of diabetes will become increasingly tangible. PMID- 15662001 TI - How obesity causes diabetes: not a tall tale. AB - The epidemic of obesity-associated diabetes is a major crisis in modern societies, in which food is plentiful and exercise is optional. The biological basis of this problem has been explored from evolutionary and mechanistic perspectives. Evolutionary theories, focusing on the potential survival advantages of "thrifty" genes that are now maladaptive, are of great interest but are inherently speculative and difficult to prove. Mechanistic studies have revealed numerous fat-derived molecules and a link to inflammation that, together, are hypothesized to underlie the obesity-diabetes connection and thereby represent prospective targets for therapeutic intervention. PMID- 15662002 TI - Diabetes, obesity, and the brain. AB - Recent evidence suggests a key role for the brain in the control of both body fat content and glucose metabolism. Neuronal systems that regulate energy intake, energy expenditure, and endogenous glucose production sense and respond to input from hormonal and nutrient-related signals that convey information regarding both body energy stores and current energy availability. In response to this input, adaptive changes occur that promote energy homeostasis and the maintenance of blood glucose levels in the normal range. Defects in this control system are implicated in the link between obesity and type 2 diabetes. PMID- 15662003 TI - Type 2 diabetes-a matter of beta-cell life and death? AB - In type 2 diabetes, the beta cells of the pancreas fail to produce enough insulin to meet the body's demand, in part because of an acquired decrease in beta-cell mass. In adults, pancreatic beta-cell mass is controlled by several mechanisms, including beta-cell replication, neogenesis, hypertrophy, and survival. Here, I discuss evidence supporting the notion that increased beta-cell apoptosis is an important factor contributing to beta-cell loss and the onset of type 2 diabetes. Interestingly, a key signaling molecule that promotes beta-cell growth and survival, insulin receptor substrate 2 (IRS-2), is a member of a family of proteins whose inhibition contributes to the development of insulin resistance in the liver and other insulin-responsive tissues. Thus, the IRS-2 pathway appears to be a crucial participant in the tenuous balance between effective pancreatic beta-cell mass and insulin resistance. PMID- 15662004 TI - Mitochondrial dysfunction and type 2 diabetes. AB - Maintenance of normal blood glucose levels depends on a complex interplay between the insulin responsiveness of skeletal muscle and liver and glucose-stimulated insulin secretion by pancreatic beta cells. Defects in the former are responsible for insulin resistance, and defects in the latter are responsible for progression to hyperglycemia. Emerging evidence supports the potentially unifying hypothesis that both of these prominent features of type 2 diabetes are caused by mitochondrial dysfunction. PMID- 15662006 TI - Porous semiconductor chalcogenide aerogels. AB - Chalcogenide aerogels based entirely on semiconducting II-VI or IV-VI frameworks have been prepared from a general strategy that involves oxidative aggregation of metal chalcogenide nanoparticle building blocks followed by supercritical solvent removal. The resultant materials are mesoporous, exhibit high surface areas, can be prepared as monoliths, and demonstrate the characteristic quantum-confined optical properties of their nanoparticle components. These materials can be synthesized from a variety of building blocks by chemical or photochemical oxidation, and the properties can be further tuned by heat treatment. Aerogel formation represents a powerful yet facile method for metal chalcogenide nanoparticle assembly and the creation of mesoporous semiconductors. PMID- 15662005 TI - Ecological change, group territoriality, and population dynamics in Serengeti lions. AB - Territorial behavior is expected to buffer populations against short-term environmental perturbations, but we have found that group living in African lions causes a complex response to long-term ecological change. Despite numerous gradual changes in prey availability and vegetative cover, regional populations of Serengeti lions remained stable for 10- to 20-year periods and only shifted to new equilibria in sudden leaps. Although gradually improving environmental conditions provided sufficient resources to permit the subdivision of preexisting territories, regional lion populations did not expand until short-term conditions supplied enough prey to generate large cohorts of surviving young. The results of a simulation model show that the observed pattern of "saltatory equilibria" results from the lions' grouping behavior. PMID- 15662007 TI - Deep-ultraviolet quantum interference metrology with ultrashort laser pulses. AB - Precision spectroscopy at ultraviolet and shorter wavelengths has been hindered by the poor access of narrow-band lasers to that spectral region. We demonstrate high-accuracy quantum interference metrology on atomic transitions with the use of an amplified train of phase-controlled pulses from a femtosecond frequency comb laser. The peak power of these pulses allows for efficient harmonic upconversion, paving the way for extension of frequency comb metrology in atoms and ions to the extreme ultraviolet and soft x-ray spectral regions. A proof-of principle experiment was performed on a deep-ultraviolet (2 x 212.55 nanometers) two-photon transition in krypton; relative to measurement with single nanosecond laser pulses, the accuracy of the absolute transition frequency and isotope shifts was improved by more than an order of magnitude. PMID- 15662008 TI - Charging effects on bonding and catalyzed oxidation of CO on Au8 clusters on MgO. AB - Gold octamers (Au8) bound to oxygen-vacancy F-center defects on Mg(001) are the smallest clusters to catalyze the low-temperature oxidation of CO to CO2, whereas clusters deposited on close-to-perfect magnesia surfaces remain chemically inert. Charging of the supported clusters plays a key role in promoting their chemical activity. Infrared measurements of the stretch vibration of CO adsorbed on mass selected gold octamers soft-landed on MgO(001) with coadsorbed O2 show a red shift on an F-center-rich surface with respect to the perfect surface. The experiments agree with quantum ab initio calculations that predict that a red shift of the C-O vibration should arise via electron back-donation to the CO antibonding orbital. PMID- 15662009 TI - Creating order from random fluctuations in small spin ensembles. AB - We demonstrate the ability to create spin order by using a magnetic resonance force microscope to harness the naturally occurring statistical fluctuations in small ensembles of electron spins. In one method, we hyperpolarized the spin system by selectively capturing the transient spin order created by the statistical fluctuations. In a second method, we took a more active approach and rectified the spin fluctuations by applying real-time feedback to the entire spin ensemble. The created spin order can be stored in the laboratory frame for a period on the order of the longitudinal relaxation time of 30 seconds and then read out. PMID- 15662010 TI - Slip-rate measurements on the Karakorum Fault may imply secular variations in fault motion. AB - Beryllium-10 surface exposure dating of offset moraines on one branch of the Karakorum Fault west of the Gar basin yields a long-term (140- to 20-thousand year) right-lateral slip rate of approximately 10.7 +/- 0.7 millimeters per year. This rate is 10 times larger than that inferred from recent InSAR analyses ( approximately 1 +/- 3 millimeters per year) that span approximately 8 years and sample all branches of the fault. The difference in slip-rate determinations suggests that large rate fluctuations may exist over centennial or millennial time scales. Such fluctuations would be consistent with mechanical coupling between the seismogenic, brittle-creep, and ductile shear sections of faults that reach deep into the crust. PMID- 15662011 TI - Speciation by distance in a ring species. AB - Ring species, which consist of two reproductively isolated forms connected by a chain of intergrading populations, have often been described as examples of speciation despite gene flow between populations, but this has never been demonstrated. We used amplified fragment length polymorphism (AFLP) markers to study gene flow in greenish warblers (Phylloscopus trochiloides). These genetic markers show distinct differences between two reproductively isolated forms but gradual change through the ring connecting these forms. These findings provide the strongest evidence yet for "speciation by force of distance" in the face of ongoing gene flow. PMID- 15662012 TI - Large sulfur bacteria and the formation of phosphorite. AB - Phosphorite deposits in marine sediments are a long-term sink for an essential nutrient, phosphorus. Here we show that apatite abundance in sediments on the Namibian shelf correlates with the abundance and activity of the giant sulfur bacterium Thiomargarita namibiensis, which suggests that sulfur bacteria drive phosphogenesis. Sediments populated by Thiomargarita showed sharp peaks of pore water phosphate (/=50 grams of phosphorus per kilogram). Laboratory experiments revealed that under anoxic conditions, Thiomargarita released enough phosphate to account for the precipitation of hydroxyapatite observed in the environment. PMID- 15662013 TI - Cardiovascular risk factors emerge after artificial selection for low aerobic capacity. AB - In humans, the strong statistical association between fitness and survival suggests a link between impaired oxygen metabolism and disease. We hypothesized that artificial selection of rats based on low and high intrinsic exercise capacity would yield models that also contrast for disease risk. After 11 generations, rats with low aerobic capacity scored high on cardiovascular risk factors that constitute the metabolic syndrome. The decrease in aerobic capacity was associated with decreases in the amounts of transcription factors required for mitochondrial biogenesis and in the amounts of oxidative enzymes in skeletal muscle. Impairment of mitochondrial function may link reduced fitness to cardiovascular and metabolic disease. PMID- 15662014 TI - Mechanism of hsp70i gene bookmarking. AB - In contrast to most genomic DNA in mitotic cells, the promoter regions of some genes, such as the stress-inducible hsp70i gene that codes for a heat shock protein, remain uncompacted, a phenomenon called bookmarking. Here we show that hsp70i bookmarking is mediated by a transcription factor called HSF2, which binds this promoter in mitotic cells, recruits protein phosphatase 2A, and interacts with the CAP-G subunit of the condensin enzyme to promote efficient dephosphorylation and inactivation of condensin complexes in the vicinity, thereby preventing compaction at this site. Blocking HSF2-mediated bookmarking by HSF2 RNA interference decreases hsp70i induction and survival of stressed cells in the G1 phase, which demonstrates the biological importance of gene bookmarking. PMID- 15662015 TI - Mathematical modeling of planar cell polarity to understand domineering nonautonomy. AB - Planar cell polarity (PCP) signaling generates subcellular asymmetry along an axis orthogonal to the epithelial apical-basal axis. Through a poorly understood mechanism, cell clones that have mutations in some PCP signaling components, including some, but not all, alleles of the receptor frizzled, cause polarity disruptions of neighboring wild-type cells, a phenomenon referred to as domineering nonautonomy. Here, a contact-dependent signaling hypothesis, derived from experimental results, is shown by reaction-diffusion, partial differential equation modeling and simulation to fully reproduce PCP phenotypes, including domineering nonautonomy, in the Drosophila wing. The sufficiency of this model and the experimental validation of model predictions reveal how specific protein protein interactions produce autonomy or domineering nonautonomy. PMID- 15662016 TI - T helper cell fate specified by kinase-mediated interaction of T-bet with GATA-3. AB - Cell lineage specification depends on both gene activation and gene silencing, and in the differentiation of T helper progenitors to Th1 or Th2 effector cells, this requires the action of two opposing transcription factors, T-bet and GATA-3. T-bet is essential for the development of Th1 cells, and GATA-3 performs an equivalent role in Th2 development. We report that T-bet represses Th2 lineage commitment through tyrosine kinase-mediated interaction between the two transcription factors that interferes with the binding of GATA-3 to its target DNA. These results provide a novel function for tyrosine phosphorylation of a transcription factor in specifying alternate fates of a common progenitor cell. PMID- 15662017 TI - Carotenoid cation formation and the regulation of photosynthetic light harvesting. AB - Photosynthetic light harvesting in excess light is regulated by a process known as feedback deexcitation. Femtosecond transient absorption measurements on thylakoid membranes show selective formation of a carotenoid radical cation upon excitation of chlorophyll under conditions of maximum, steady-state feedback deexcitation. Studies on transgenic Arabidopsis thaliana plants confirmed that this carotenoid radical cation formation is correlated with feedback deexcitation and requires the presence of zeaxanthin, the specific carotenoid synthesized during high light exposure. These results indicate that energy transfer from chlorophyll molecules to a chlorophyllzeaxanthin heterodimer, which then undergoes charge separation, is the mechanism for excess energy dissipation during feedback deexcitation. PMID- 15662018 TI - Cryo-electron tomography reveals the cytoskeletal structure of Spiroplasma melliferum. AB - Evidence has accumulated recently that not only eukaryotes but also bacteria can have a cytoskeleton. We used cryo-electron tomography to study the three dimensional structure of Spiroplasma melliferum cells in a close-to-native state at approximately 4-nanometer resolution. We showed that these cells possess two types of filaments arranged in three parallel ribbons underneath the cell membrane. These two filamentous structures are built of the fibril protein and possibly the actin-like protein MreB. On the basis of our structural data, we could model the motility modes of these cells and explain how helical Mollicutes can propel themselves by means of coordinated length changes of their cytoskeletal ribbons. PMID- 15662019 TI - Modulation of estrogen signaling by the novel interaction of heat shock protein 27, a biomarker for atherosclerosis, and estrogen receptor beta: mechanistic insight into the vascular effects of estrogens. AB - OBJECTIVE: We sought to discover proteins that associate with estrogen receptor beta (ERbeta) and modulate estrogen signaling. METHODS AND RESULT: Using a yeast 2-hybrid screen, we identified heat shock protein 27 (HSP27) as an ERbeta associated protein. HSP27 is a recently identified biomarker of atherosclerosis that is secreted at reduced levels from atherosclerotic compared with normal arteries. In vitro protein-binding assays confirmed the specific interaction of HSP27 with ERbeta and not ERalpha. HSP27 expression was absent in coronary arteries with complex atherosclerotic lesions. Interestingly, HSP27 expression was also absent in 60% of coronary arteries from young males and females (27+/ 6.5 years) with normal histology or nonobstructive fatty streaks/atheromas. Moreover, the absence of HSP27 in these normal or minimally diseased arteries coincided with the loss of ERbeta expression. Only 35% of arteries showed coexpression of HSP27 and ERbeta. Relative to controls, estradiol-mediated transcription was reduced 20% with overexpression of HSP27 and increased 44% when HSP27 protein levels were reduced with HSP27 siRNA. CONCLUSIONS: HSP27, an ERbeta associated protein, shows attenuated expression with coronary atherosclerosis and modulates estrogen signaling. PMID- 15662020 TI - 15-deoxy-Delta12,14-prostaglandin J2 (15d-PGJ2) signals through retinoic acid receptor-related orphan receptor-alpha but not peroxisome proliferator-activated receptor-gamma in human vascular endothelial cells: the effect of 15d-PGJ2 on tumor necrosis factor-alpha-induced gene expression. AB - OBJECTIVE: 15-deoxy-Delta12,14-prostaglandin J2 (15d-PGJ2), a natural ligand of the peroxisome proliferator-activated receptor-gamma (PPARgamma), has been shown to inhibit proinflammatory gene expression, but the signaling mechanisms involved remain unclear. Because retinoic acid receptor-related orphan receptor-alpha (RORalpha) has been reported to suppress tumor necrosis factor-alpha (TNF-alpha) induced expression of proinflammatory genes, we hypothesized that 15d-PGJ2 may induce RORalpha expression resulting in inhibition of proinflammatory gene expression. METHODS AND RESULTS: We demonstrate that 15d-PGJ2 induced RORalpha1 and RORalpha4 expression and inhibited TNF-alpha-induced vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) expression in human umbilical vein endothelial cells (HUVECs). In contrast, the synthetic PPARgamma ligand pioglitazone weakly induced RORalpha4 expression but did not affect RORalpha1 expression or TNF-alpha-induced gene expression. Biphenol A diglycidyl ether, a PPARgamma antagonist, did not block the effect of 15d-PGJ2 on RORalpha expression. Adenovirus-mediated overexpression of RORalpha1 inhibited TNF-alpha-induced VCAM-1 and ICAM-1 expression, and overexpression of a mutant form of RORalpha1 (RORalpha1Delta), which inhibited transcriptional activity of RORalpha1 and RORalpha4, attenuated its inhibition. Furthermore, we found that RORalpha1Delta attenuated the inhibitory actions of 15d-PGJ2 on TNF-alpha-induced VCAM-1 and ICAM-1 expression. CONCLUSIONS: These results suggest that 15d-PGJ2 inhibits TNF-alpha-induced expression of proinflammatory genes mediated in part via induction of RORalpha in HUVECs. This mechanism provides a novel insight into PPARgamma-independent actions of 15d-PGJ2. PMID- 15662021 TI - Mesoangioblasts, vessel-associated multipotent stem cells, repair the infarcted heart by multiple cellular mechanisms: a comparison with bone marrow progenitors, fibroblasts, and endothelial cells. AB - OBJECTIVE: To test the potential of mesoangioblasts (Mabs) in reducing postischemic injury in comparison with bone marrow progenitor cells (BMPCs), fibroblasts (Fbs), and embryonic stem cell-derived endothelial cells (ECs), and to identify putative cellular protective mechanisms. METHODS AND RESULTS: Cells were injected percutaneously in the left ventricular (LV) chamber of C57BL/6 mice, 3 to 6 hours after coronary ligation, and detected in the hearts 2 days and 6 weeks later. Echocardiographic examinations were performed at 6 weeks. LV dilation was reduced and LV shortening fraction was improved with Mabs and BMPCs but not with ECs and Fbs. Donor cell colonization of the host myocardium was modest and predominantly in the smooth muscle layer of vessels. Capillary density was higher in the peripheral infarct area and apoptotic cardiomyocytes were fewer with Mabs and BMPCs. Mabs and BMPCs, but not Fbs or ECs, promoted survival of cultured cardiocytes under low-oxygen in culture. This activity was present in Mab-conditioned medium and could be replaced by a combination of basic fibroblast growth factor (bFGF), insulin-like growth factor (IGF)-1, and hepatocyte growth factor (HGF), all of which are produced by these cells. Conditioned medium from Mabs, but not from Fbs, stimulated proliferation of smooth muscle cells in vitro. CONCLUSIONS: Mabs appear as effective as BMPCs in reducing postinfarction LV dysfunction, likely through production of antiapoptotic and angiogenic factors. PMID- 15662022 TI - Hyperglycemia reduces survival and impairs function of circulating blood-derived progenitor cells. AB - OBJECTIVE: Function and availability of circulating progenitor cells (CPC) have been shown to be impaired in patients with diabetes mellitus (DM). Therefore, the aim of the present study was to analyze possible mechanisms leading to the reduction of CPC amount and function. METHODS AND RESULTS: Peripheral blood mononuclear cells (MNCs) of healthy donors (n=15) were cultivated under hyperglycemia (HG) conditions (12 mmol/L D-Glucose) or in osmotic control medium (Con) (5 mmol/L D-Glucose plus 7 mmol/L L-Glucose) for 7 days. CPC amount was determined by uptake of acetylated low-density lipoprotein and lectin binding. On the functional level, cell cycle status, nitric oxide (NO) production, and migrational and integrative capacity of CPCs were assessed. HG conditions caused a significant decrease in CPC amount derived from healthy MNCs. Furthermore, HG conditions led to a functional impairment, reflected in a decreased NO production and matrix metalloproteinase (MMP)-9 activity, as well as an impairment of the migrational and integrative capacities. CONCLUSIONS: HG, a main feature of DM, affects important functional characteristics of CPCs. These results may provide further insight into the pathomechanism of endothelial dysfunction in HG. PMID- 15662023 TI - Selective blockade of the intermediate-conductance Ca2+-activated K+ channel suppresses proliferation of microvascular and macrovascular endothelial cells and angiogenesis in vivo. AB - OBJECTIVE: Ca2+-activated K+ (K(Ca)) channels have been proposed to promote mitogenesis in several cell types. Here, we tested whether the intermediate conductance K(Ca) channel (IKCa1) and the large-conductance K(Ca) channel (BK(Ca)) contribute to endothelial cell (EC) proliferation and angiogenesis. MATERIAL AND RESULTS: Function and expression of IKCa1 and BK(Ca)/Slo were investigated by patch-clamp analysis and real-time RT-PCR in human umbilical vein ECs (HUVECs) and in dermal human microvascular ECs 1 (HMEC-1). HMEC-1 expressed IKCa1 and BK(Ca)/Slo, whereas HUVECs expressed IKCa1. A 48-hour exposure to basic fibroblast growth factor (bFGF) augmented IKCa1 current amplitudes and induced a 3-fold increase in IKCa1 mRNA expression in HUVECs and HMEC-1. Vascular endothelial growth factor (VEGF) was also effective in upregulating IKCa1. BK(Ca)/Slo expression and current amplitudes in HMEC-1 were not altered by bFGF. bFGF- and VEGF-induced EC proliferation was suppressed by charybdotoxin, clotrimazole, or the selective IKCa1 blocker 1-[(2-chlorophenyl)diphenylmethyl] 1H-pyrazole (TRAM-34), whereas inhibition of BK(Ca)/Slo by iberiotoxin was ineffective. In the Matrigel plug assay in mice, administration of TRAM-34 for 2 weeks significantly suppressed angiogenesis by approximately 85%. CONCLUSIONS: bFGF and VEGF upregulate expression of IKCa1 in human ECs. This upregulation of IKCa1 seems to be required for mitogen-induced EC proliferation and angiogenesis in vivo. Selective IKCa1 blocker might be of therapeutic value to prevent tumor angiogenesis. PMID- 15662024 TI - Activated forkhead transcription factor inhibits neointimal hyperplasia after angioplasty through induction of p27. AB - OBJECTIVE: We examined the effects of FKHRL1 (forkhead transcription factor in rhabdomyosarcoma like-1) overexpression on vascular smooth muscle cell (VSMC) proliferation, apoptosis, and cell cycle, in vitro, and the role of FKHRL1 and p27 in the pathophysiology of neointimal growth after balloon angioplasty, in vivo. Furthermore, we tested whether FKHRL1 overexpression can inhibit neointimal hyperplasia in a rat carotid artery model. METHODS AND RESULTS: Adenovirus expressing the constitutively active FKHRL1 (FKHRL1-TM; triple mutant) with 3 Akt phosphorylation sites mutated was transfected to subconfluent VSMCs. FKHRL1 overexpression in cultured VSMCs increased p27 expression, leading to G1 phase cell-cycle arrest and increased apoptosis. In vivo, the phosphorylation of FKHRL1 increased significantly 3 hours after balloon injury and decreased thereafter, with the subsequent downregulation of p27. Although the phosphorylation of FKHRL1 was greatest at 3 hours, the downregulation of p27 showed a temporal delay, only slightly starting to decrease after 3 hours and reaching a nadir at 72 hours after balloon injury. Gene transfer of FKHRL1-TM increased p27, decreased proliferation, and increased apoptosis of VSMCs, which resulted in a marked reduction in neointima formation (intima-to-media ratio: 0.31+/-0.13 versus 1.17+/-0.28, for FKHRL1-TM versus Adv-GFP; P<0.001). CONCLUSIONS: Balloon angioplasty leads to the phosphorylation of FKHRL1 and decreased expression of p27, thereby promoting a proliferative phenotype in VSMCs in vitro and in vivo. This study reveals the importance of FKHRL1 in proliferation and viability of VSMCs and suggests that it may serve as a molecular target for interventions to reduce neointima formation after angioplasty. PMID- 15662025 TI - Human atherosclerotic plaque contains viable invasive Actinobacillus actinomycetemcomitans and Porphyromonas gingivalis. PMID- 15662026 TI - Leukocytosis and ischemic vascular disease morbidity and mortality: is it time to intervene? AB - The association between leukocytosis and increased morbidity and mortality of ischemic vascular disease has been observed for more than half a century, and recent studies in >350,000 patients confirm the robustness of the association and the dramatically higher relative and absolute acute and chronic mortality rates in patients with high versus low leukocyte counts. Although there is reason to believe that the association is not causal (that is, that leukocytosis is simply a marker of inflammation), there is also reason to believe that the leukocytosis directly enhances acute thrombosis and chronic atherosclerosis. Leukocytosis also is associated with poor prognosis and vaso-occlusive events in patients with sickle cell disease, and experimental data suggest a direct role for leukocytes in microvascular obstruction. The only way to test whether leukocytes contribute directly to poor outcome in ischemic cardiovascular disease is to assess the effect of modifying leukocyte function or number. Because selective blockade of leukocyte integrin alphaMbeta2 and P-selectin have thus far been disappointing as therapeutic strategies in human cardiovascular and cerebrovascular disease, I discuss the potential risks and benefits of short-term treatment with hydroxyurea to decrease the leukocyte count in select populations of patients at the highest risk of short-term death. PMID- 15662027 TI - Lesion development and response to immunization reveal a complex role for CD4 in atherosclerosis. AB - Atherosclerosis is a complex disease, bearing many of the characteristics of a chronic inflammatory process. Both cellular and humoral immune responses may be involved in the disease development. Oxidized low-density lipoprotein (oxLDL) is suggested to be an autoantigen in atherosclerosis. A protective effect against atherosclerosis has been demonstrated in animals immunized with oxLDL. Such a protection is associated with elevation of T cell-dependent IgG antibodies against oxLDL. In addition, it has been shown that immunization with Freund adjuvant alone also confers protection against development of atherosclerosis. We therefore hypothesized that CD4+ T cells are critical in the development of atherosclerosis and that they are involved in protective immune reactions after immunization. The development of atherosclerosis was studied in apolipoprotein E knockout (apoE KO) mice and CD4/apoE double knockout (dKO) mice that were immunized with either oxLDL in Freund adjuvant or adjuvant alone, or left untreated. Our results show that (1) the absence of CD4+ cells in apoE KO mice leads to reduced atherosclerosis, indicating that CD4+ cells constitute a major proatherogenic cell population, and (2) the atheroprotective effect of LDL immunization does not depend on CD4+ cells, whereas (3) the atheroprotective effect of adjuvant injection is CD4-dependent. These findings demonstrate complex roles of immune cell-cell interactions in the regulation of the atherosclerotic process and point to several possible targets in the treatment and prevention of atherosclerosis. PMID- 15662028 TI - Haptoglobin genotype- and diabetes-dependent differences in iron-mediated oxidative stress in vitro and in vivo. AB - We have recently demonstrated in multiple independent population-based longitudinal and cross sectional analyses that the haptoglobin 2-2 genotype is associated with an increased risk for diabetic cardiovascular disease. The chief function of haptoglobin (Hp) is to bind to hemoglobin and thereby prevent hemoglobin-induced oxidative tissue damage. This antioxidant function of haptoglobin is mediated in part by the ability of haptoglobin to prevent the release of iron from hemoglobin on its binding. We hypothesized that there may be diabetes- and haptoglobin genotype-dependent differences in the amount of catalytically active redox active iron derived from hemoglobin. We tested this hypothesis using several complementary approaches both in vitro and in vivo. First, measuring redox active iron associated with haptoglobin-hemoglobin complexes in vitro, we demonstrate a marked increase in redox active iron associated with Hp 2-2-glycohemoglobin complexes. Second, we demonstrate increased oxidative stress in tissue culture cells exposed to haptoglobin 2-2 hemoglobin complexes as opposed to haptoglobin 1-1-hemoglobin complexes, which is inhibitable by desferrioxamine by either a chelation or reduction mechanism. Third, we demonstrate marked diabetes-dependent differences in the amount of redox active iron present in the plasma of mice genetically modified expressing the Hp 2 allele as compared with the Hp 1 allele. Taken together these data implicate redox active iron in the increased susceptibility of individuals with the Hp 2 allele to diabetic vascular disease. PMID- 15662029 TI - Vascular endothelial-cadherin tyrosine phosphorylation in angiogenic and quiescent adult tissues. AB - Vascular endothelial-cadherin (VE-cadherin) plays a key role in angiogenesis and in vascular permeability. The regulation of its biological activity may be a central mechanism in normal or pathological angiogenesis. VE-cadherin has been shown to be phosphorylated on tyrosine in vitro under various conditions, including stimulation by VEGF. In the present study, we addressed the question of the existence of a tyrosine phosphorylated form of VE-cadherin in vivo, in correlation with the quiescent versus angiogenic state of adult tissues. Phosphorylated VE-cadherin was detected in mouse lung, uterus, and ovary but not in other tissues unless mice were injected with peroxovanadate to block protein phosphatases. Remarkably, VE-cadherin tyrosine phosphorylation was dramatically increased in uterus and ovary, and not in other organs, during PMSG/hCG-induced angiogenesis. In parallel, we observed an increased association of VE-cadherin with Flk1 (VEGF receptor 2) during hormonal angiogenesis. Additionally, Src kinase was constitutively associated with VE-cadherin in both quiescent and angiogenic tissues and increased phosphorylation of VE-cadherin-associated Src was detected in uterus and ovary after hormonal treatment. Src-VE-cadherin association was detected in cultured endothelial cells, independent of VE cadherin phosphorylation state and Src activation level. In this model, Src inhibition impaired VEGF-induced VE-cadherin phosphorylation, indicating that VE cadherin phosphorylation was dependent on Src activation. We conclude that VE cadherin is a substrate for tyrosine kinases in vivo and that its phosphorylation, together with that of associated Src, is increased by angiogenic stimulation. Physical association between Flk1, Src, and VE-cadherin may thus provide an efficient mechanism for amplification and perpetuation of VEGF stimulated angiogenic processes. PMID- 15662030 TI - Increased PI3-kinase in presympathetic brain areas of the spontaneously hypertensive rat. AB - Existing evidence led us to hypothesize that increases in p85alpha, a regulatory subunit of PI3-kinase, in presympathetic brain areas contribute to hypertension. PI3-kinase p85alpha, p110alpha, and p110delta mRNA was 1.5- to 2-fold higher in the paraventricular nucleus (PVN) of spontaneously hypertensive rats (SHR) compared with their controls, Wistar Kyoto rats (WKY). The increase in p85alpha/p110delta was attenuated in SHR treated with captopril, an angiotensin (Ang)-converting enzyme inhibitor, from in utero to 6 months of age. In the rostral ventrolateral medulla (RVLM), p110delta mRNA was approximately 2-fold higher in SHR than in WKY. Moreover, the increases in mRNA were associated with higher PI3-kinase activity in both nuclei. The functional relevance was studied in neuronal cultures because SHR neurons reflect the augmented p85alpha mRNA and PI3-kinase activity. Expression of a p85 dominant-negative mutant decreased norepinephrine (NE) transporter mRNA and [3H]NE uptake by approximately 60% selectively in SHR neurons. In summary, increased p85alpha/p110delta expression in the PVN and RVLM is associated with increased PI3-kinase activity in the SHR. Furthermore, normalized PI3-kinase p85alpha/p110delta expression within the PVN might contribute to the overall effect of captopril, perhaps attributable to a consequent decrease in NE availability. PMID- 15662031 TI - Induced deletion of the N-cadherin gene in the heart leads to dissolution of the intercalated disc structure. AB - The structural integrity of the heart is maintained by the end-to-end connection between the myocytes called the intercalated disc. The intercalated disc contains different junctional complexes that enable the myocardium to function as a syncytium. One of the junctional complexes, the zonula adherens or adherens junction, consists of the cell adhesion molecule, N-cadherin, which mediates strong homophilic cell-cell adhesion via linkage to the actin cytoskeleton. To determine the function of N-cadherin in the working myocardium, we generated a conditional knockout containing loxP sites flanking exon 1 of the N-cadherin (Cdh2) gene. Using a cardiac-specific tamoxifen-inducible Cre transgene, N cadherin was deleted in the adult myocardium. Loss of N-cadherin resulted in disassembly of the intercalated disc structure, including adherens junctions and desmosomes. The mutant mice exhibited modest dilated cardiomyopathy and impaired cardiac function, with most animals dying within two months after tamoxifen administration. Decreased sarcomere length and increased Z-line thickness were observed in the mutant hearts consistent with loss of muscle tension because N cadherin was no longer available to anchor myofibrils at the plasma membrane. Ambulatory electrocardiogram monitoring captured the abrupt onset of spontaneous ventricular tachycardia, confirming that the deaths were arrhythmic in nature. A significant decrease in the gap junction protein, connexin 43, was observed in the N-cadherin-depleted hearts. This animal model provides the first demonstration of the hierarchical relationship of the structural components of the intercalated disc in the working myocardium, thus establishing N-cadherin's paramount importance in maintaining the structural integrity of the heart. PMID- 15662032 TI - Glycogen-Synthase Kinase3beta/beta-catenin axis promotes angiogenesis through activation of vascular endothelial growth factor signaling in endothelial cells. AB - Glycogen-Synthase Kinase 3beta (GSK3beta) has been shown to function as a nodal point of converging signaling pathways in endothelial cells to regulate vessel growth, but the signaling mechanisms downstream from GSK3beta have not been identified. Here, we show that beta-catenin is an important downstream target for GSK3beta action in angiogenesis and dissect the signal transduction pathways involved in the angiogenic phenotype. Transduction of human umbilical vein endothelial cells (HUVECs) with a kinase-mutant form of the enzyme (KM-GSK3beta) increased cytosolic beta-catenin levels, whereas constitutively active GSK3beta (S9A-GSK3beta) reduced beta-catenin levels. Lymphoid enhancer factor/T-cell factor promoter activity was upregulated by KM-GSK3beta and diminished by S9A GSK3beta, whereas manipulation of Akt signaling had no effect on this parameter. beta-Catenin transduction induced capillary formation in a Matrigel-plug assay in vivo and promoted endothelial cell differentiation into network structures on Matrigel-coated plates in vitro. beta-Catenin activated the expression of vascular endothelial growth factor (VEGF)-A and VEGF-C in endothelial cells, and these effects were mediated at the levels of protein, mRNA, and promoter activity. Consistent with these data, beta-catenin increased the phosphorylation of the VEGF receptor 2 (VEGF-R2) and promoted its association with PI3-kinase, leading to a dose-dependent activation of the serine-threonine kinase Akt. Inhibition of PI3-kinase or Akt signaling led to a significant reduction in the pro-angiogenic activity of beta-catenin. Collectively, these data show that the growth factor-PI3-kinase-Akt axis functions downstream of GSK3beta/beta-catenin signaling in endothelial cells to promote angiogenesis. PMID- 15662033 TI - Central role of PKCbeta in neointimal expansion triggered by acute arterial injury. AB - We tested the hypothesis that PKCbeta contributes to vascular smooth muscle cell (SMC) migration and proliferation; processes central to the pathogenesis of restenosis consequent to vascular injury. Homozygous PKCbeta null (-/-) mice or wild-type mice fed the PKCbeta inhibitor, ruboxistaurin, displayed significantly decreased neointimal expansion in response to acute femoral artery endothelial denudation injury compared with controls. In vivo and in vitro analyses demonstrated that PKCbetaII is critically linked to SMC activation, at least in part via regulation of ERK1/2 MAP kinase and early growth response-1. These data highlight novel roles for PKCbeta in the SMC response to acute arterial injury and suggest that blockade of PKCbeta may represent a therapeutic strategy to limit restenosis. PMID- 15662035 TI - Accessory Kvbeta1 subunits differentially modulate the functional expression of voltage-gated K+ channels in mouse ventricular myocytes. AB - Voltage-gated K+ (Kv) channel accessory (beta) subunits associate with pore forming Kv alpha subunits and modify the properties and/or cell surface expression of Kv channels in heterologous expression systems. There is very little presently known, however, about the functional role(s) of Kv beta subunits in the generation of native cardiac Kv channels. Exploiting mice with a targeted disruption of the Kvbeta1 gene (Kvbeta1-/-), the studies here were undertaken to explore directly the role of Kvbeta1 in the generation of ventricular Kv currents. Action potential waveforms and peak Kv current densities are indistinguishable in myocytes isolated from the left ventricular apex (LVA) of Kvbeta1-/- and wild-type (WT) animals. Analysis of Kv current waveforms, however, revealed that mean+/-SEM I(to,f) density is significantly (P< or =0.01) lower in Kvbeta1-/- (21.0+/-0.9 pA/pF; n=68), than in WT (25.3+/-1.4 pA/pF; n=42), LVA myocytes, and that mean+/-SEM I(K,slow) density is significantly (P< or =0.01) higher in Kvbeta1-/- (19.1+/-0.9 pA/pF; n=68), compared with WT (15.9+/-0.7 pA/pF; n=42), LVA cells. Pharmacological studies demonstrated that the TEA sensitive component of I(K,slow), I(K,slow2,) is selectively increased in Kvbeta1 /- LVA myocytes. In parallel with the alterations in I(to,f) and I(K,slow2) densities, Kv4.3 expression is decreased and Kv2.1 expression is increased in Kvbeta1-/- ventricles. Taken together, these results demonstrate that Kvbeta1 differentially regulates the functional cell surface expression of myocardial I(to,f) and I(K,slow2) channels. PMID- 15662034 TI - Action potential duration restitution and alternans in rabbit ventricular myocytes: the key role of intracellular calcium cycling. AB - Action potential duration (APD) restitution properties and repolarization alternans are thought to be important arrhythmogenic factors. We investigated the role of intracellular calcium (Ca2+i) cycling in regulating APD restitution slope and repolarization (APD) alternans in patch-clamped rabbit ventricular myocytes at 34 to 36 degrees C, using the perforated or ruptured patch clamp techniques with Fura-2-AM to record Ca2+i. When APD restitution was measured by either the standard extrastimulus (S1S2) method or the dynamic rapid pacing method, the maximum APD restitution slope exceeded 1 by both methods, but was more shallow with the dynamic method. These differences were associated with greater Ca2+i accumulation during dynamic pacing. The onset of APD alternans occurred at diastolic intervals at which the APD restitution slope was significantly <1 and was abolished by suppressing sarcoplasmic reticulum (SR) Ca2+i cycling with thapsigargin and ryanodine, or buffering the global Ca2+i transient with BAPTA-AM or BAPTA. Thapsigargin and ryanodine flattened APD restitution slope to <1 when measured by the dynamic method, but not by the S1S2 method. BAPTA-AM or BAPTA failed to flatten APD restitution slope to <1 by either method. In conclusion, APD alternans requires intact Ca2+i cycling and is not reliably predicted by APD restitution slope when Ca2+i cycling is suppressed. Ca2+i cycling may contribute to differences between APD restitution curves measured by S1S2 versus dynamic pacing protocols by inducing short-term memory effects related to pacing dependent Ca2+i accumulation. PMID- 15662038 TI - Improving trial power through use of prognosis-adjusted end points. AB - BACKGROUND AND PURPOSE: The stroke patient population is heterogeneous, leading to wide variation in outcome caused by differences in age, initial severity, and presence of concomitant disease. Setting an identical recovery target for all patients in intervention trials may conceal individually important therapeutic treatment effects. Instead, a variable end point that takes severity or likely prognosis into account may be more informative. METHODS: We used data from the Glycine Antagonist in Neuroprotection (GAIN) International trial to assess statistical power of various primary end points for intervention trials. We selected prognosis-adjusted cut points based on Barthel Index (BI) or Rankin Scale (RS) using a prognostic model, or assigned a fixed end point within subgroups of patients defined by their Oxford category or National Institutes of Health Stroke Scale (NIHSS) score. We simulated a treatment effect and estimated statistical power with standard formulae. RESULTS: Assignment of end points using a prognostic model for individual patients increased statistical power, when compared with assigning end points using only the Oxford classification. For the BI, power was increased from 60% to 88% (equivalent to a 49% reduction in sample size if power remains unchanged). With the RS end points, power was increased from 84% to 92% (or a 24% reduction in sample size). Versus a fixed end point for all patients, model-based methods increased power by 22 percentage points for BI> or =95 and 14 percentage points for RS< or =1 (effective sample size reductions 43% and 34%). CONCLUSIONS: Prognosis-adjusted end points can increase statistical power compared with fixed end points. Assessment is based on realistic goals for individual patients and yet trial results remain generalizable. PMID- 15662039 TI - Phenytoin exposure is associated with functional and cognitive disability after subarachnoid hemorrhage. AB - BACKGROUND AND PURPOSE: Phenytoin (PHT) is routinely used for seizure prophylaxis after subarachnoid hemorrhage (SAH), but may adversely affect neurologic and cognitive recovery. METHODS: We studied 527 SAH patients and calculated a "PHT burden" for each by multiplying the average serum level of PHT by the time in days between the first and last measurements, up to a maximum of 14 days from ictus. Functional outcome at 14 days and 3 months was measured with the modified Rankin scale, with poor functional outcome defined as dependence or worse (modified Rankin Scale > or =4). We assessed cognitive outcomes at 14 days and 3 months with the telephone interview for cognitive status. RESULTS: PHT burden was associated with poor functional outcome at 14 days (OR, 1.5 per quartile; 95% CI, 1.3 to 1.8; P<0.001), although not at 3 months (P=0.09); the effect remained (OR, 1.6 per quartile; 95% CI, 1.2 to 2.1; P<0.001) after correction for admission Glasgow Coma Scale, fever, stroke, age, National Institutes of Health Stroke Scale > or =10, hydrocephalus, clinical vasospasm, and aneurysm rebleeding. Seizure in hospital (OR, 4.1; 95% CI, 1.5 to 11.1; P=0.002) was associated with functional disability in a univariate model only. Higher quartiles of PHT burden were associated with worse telephone interview for cognitive status scores at hospital discharge (P<0.001) and at 3 months (P=0.003). CONCLUSIONS: Among patients treated with PHT, burden of exposure to PHT predicts poor neurologic and cognitive outcome after SAH. PMID- 15662040 TI - Left ventricular mass indexed to height and prevalent MRI cerebrovascular disease in an African American cohort: the Atherosclerotic Risk in Communities study. AB - BACKGROUND AND PURPOSE: Previous studies have demonstrated that echocardiographic left ventricular mass (LVM) is an independent risk factor for stroke in whites. Despite the greater burden of stroke, the echocardiographic predictors of stroke in African Americans remain poorly understood. METHODS: This investigation is a retrospective analysis of prospectively collected data from the Jackson, Miss (all African American), cohort of the Atherosclerotic Risk in the Communities study. Between 1993 and 1995, 2445 participants received an echocardiogram, and a random subset (n=778) received cerebral MRI evaluating presence of infarcts or white matter disease (WMD; >3 on a scale of 0 to 9). Compared with the entire Jackson cohort, the random subset was older, had a lower body mass index (BMI), and a higher systolic blood pressure (SBP). Logistic regression models examined the relations of LVM indexed by height (LVM/height) to MRI findings adjusted for age, gender, BMI, SBP, hypertensive medications, diabetes, total/high-density lipoprotein cholesterol, smoking status, and history of myocardial infarction. RESULTS: The 667 participants (63% women; 62+/-4 years of age) had a high prevalence of hypertension (68%), obesity (46%), echocardiographic left ventricular hypertrophy (49%), MRI stroke (n=133), and WMD (n=92). Adjusted LVM/height was significantly associated with prevalent MRI stroke (odds ratio [OR], 1.3; 95% CI, 1.1 to 1.7; P=0.02) and WMD (OR, 1.5; 95% CI, 1.1 to 1.9; P=0.006; OR expressed per 1 SD LVM/height, 45 g/m). CONCLUSIONS: In this randomized subset of a population-based cohort of African American adults, LVM/height was related to MRI evidence of prevalent cerebrovascular disease. The current study supports the hypothesis that LVM/height is an important risk factor for stroke in multiple ethnicities. PMID- 15662041 TI - Public awareness of warning symptoms, risk factors, and treatment of stroke in northwest India. AB - BACKGROUND AND PURPOSE: This study assessed public awareness of warning symptoms, risk factors, and treatment of stroke in Ludhiana, Punjab, North West India. METHODS: A hospital-based survey was conducted between February 2002 and September 2002 by the Stroke section of Christian Medical College. The study subjects were relatives of patients without history of stroke, attending the outpatient department of the hospital. Trained medical students, interns, and a nurse interviewed subjects using a structured, pretested, open-ended questionnaire. RESULTS: Nine hundred forty-two individuals were interviewed during the study period (56.4% men, mean age 40.1 years, age range 15 to 80 years). Forty-five percent of the subjects did not recognize the brain as the affected organ in stroke. In the multivariate analysis, higher education (P<0.001; odds ratio 2.6; 95%, CI 1.8 to 3.8) and upper socioeconomic status (P<0.005; odds ratio 1.6; CI, 1.1 to 2.2) correlated with a better knowledge of which organ was affected in stroke. Twenty-three percent of the participants did not know a single warning symptom of stroke. Twenty-one percent of the subjects could not identify even a single risk factor for stroke. Seven percent of the study population believed that oil massage would improve stroke victims. A small proportion of subjects believed in witchcraft, faith healing, homeopathic, and ayurvedic treatment (3%). CONCLUSIONS: This hospital-based survey reveals a better awareness of stroke warning signs and risk factors. However, knowledge regarding the organ involved, etiology, and treatment of stroke is lacking. Considerable education is needed to increase public awareness in modern concepts of stroke treatment. PMID- 15662042 TI - The new anticonvulsant retigabine favors voltage-dependent opening of the Kv7.2 (KCNQ2) channel by binding to its activation gate. AB - Retigabine (RTG) is an anticonvulsant drug with a novel mechanism of action. It activates neuronal KCNQ-type K(+) channels by inducing a large hyperpolarizing shift of steady-state activation. To identify the structural determinants of KCNQ channel activation by RTG, we constructed a set of chimeras using the neuronal K(v)7.2 (KCNQ2) channel, which is activated by RTG, and the cardiac K(v)7.1 (KCNQ1) channel, which is not affected by this drug. Substitution of either the S5 or the S6 segment in K(v)7.2 by the respective parts of K(v)7.1 led to a complete loss of activation by RTG. Trp236 in the cytoplasmic part of S5 and the conserved Gly301 in S6 (K(v)7.2), considered as the gating hinge (Ala336 in K(v)7.1), were found to be crucial for the RTG effect: mutation of these residues could either knockout the effect in K(v)7.2 or restore it partially in K(v)7.1/K(v)7.2 chimeras. We propose that RTG binds to a hydrophobic pocket formed upon channel opening between the cytoplasmic parts of S5 and S6 involving Trp236 and the channel's gate, which could well explain the strong shift in voltage-dependent activation. PMID- 15662043 TI - EDGE: a centralized resource for the comparison, analysis, and distribution of toxicogenomic information. AB - Transcriptional profiling via microarrays holds great promise for toxicant classification and hazard prediction. Unfortunately, the use of different microarray platforms, protocols, and informatics often hinders the meaningful comparison of transcriptional profiling data across laboratories. One solution to this problem is to provide a low-cost and centralized resource that enables researchers to share toxicogenomic data that has been generated on a common platform. In an effort to create such a resource, we developed a standardized set of microarray reagents and reproducible protocols to simplify the analysis of liver gene expression in the mouse model. This resource, referred to as EDGE, was then used to generate a training set of 117 publicly accessible transcriptional profiles that can be accessed at http://edge.oncology.wisc.edu/. The Web accessible database was also linked to an informatics suite that allows on-line clustering and K-means analyses as well as Boolean and sequence-based searches of the data. We propose that EDGE can serve as a prototype resource for the sharing of toxicogenomics information and be used to develop algorithms for efficient chemical classification and hazard prediction. PMID- 15662044 TI - Amino acids critical for substrate affinity of rat organic cation transporter 1 line the substrate binding region in a model derived from the tertiary structure of lactose permease. AB - To identify functionally relevant amino acids in the rat organic cation transporter 1 (rOCT1), 18 consecutive amino acids in the presumed fourth transmembrane alpha helix (TMH) were mutated and functionally characterized after expression in Xenopus laevis oocytes. After mutation of three amino acids on successive turns of the alpha helix, K(m) values for tetraethylammonium (TEA) and/or 1-methyl-4-phenylpyridinium (MPP) were decreased. After replacement of Trp218 by tyrosine (W218Y) and Tyr222 by leucine (Y222L), the K(m) values for both TEA and MPP were decreased. In mutants Y222F and T226A, only the K(m) values for TEA and MPP were decreased, respectively. The data suggest that amino acids Trp218 and Tyr222 participate in the binding of both TEA and MPP, whereas Thr226 is only involved in the binding of MPP. Using the crystal structure of the lactose permease LacY from Escherichia coli that belongs to the same major facilitator superfamily as rOCT1, we modeled the tertiary structure of the presumed 12 transmembrane alpha helices. The validity of the model was suggested because seven amino acids that have been shown to participate in the binding of cations by mutagenesis experiments [fourth TMH Trp218, Tyr222, and Thr226 (this work); 10th TMH Ala443, Leu447, and Gln448 (companion work in this issue of Molecular Pharmacology); 11th TMH Asp475 (previous report)] are located in one region surrounding a large cleft that opens to the intracellular side. The dimensions of TEA in comparison with the interacting amino acids in the modeled cleft suggest that more than one TEA molecule can bind in parallel to the modeled conformation of the transporter. PMID- 15662045 TI - Subtype-specific affinity for corticosterone of rat organic cation transporters rOCT1 and rOCT2 depends on three amino acids within the substrate binding region. AB - The affinity of corticosterone to organic cation transporters (OCTs) is subtype- and species-dependent. For example, the IC50 values for corticosterone inhibition of cation uptake by transporters rOCT1 and rOCT2 are approximately 150 and approximately 4 microM, respectively. By introducing domains and amino acids from rOCT2 into rOCT1, we found that the exchange of three amino acids in the presumed 10th transmembrane alpha helix is sufficient to increase the affinity of rOCT1 for corticosterone to that of rOCT2. Replacement of these amino acids in rOCT2 decreased the affinity for corticosterone. These amino acids (Ala443, Leu447, and Gln448 in rOCT1 and Ile443, Tyr447, and Glu448 in rOCT2) are probably located within the substrate binding region because in rOCT1 mutants, the K(m) values for uptake of tetraethylammonium (TEA) and 1-methyl-4-phenylpyridinium (MPP) were decreased in parallel with a decrease of the IC50 values for the inhibition of cation uptake by corticosterone. In mutant rOCT1(L447Y/Q448E), the IC50 value for the inhibition of [3H]MPP (0.1 microM) uptake by corticosterone (24 +/- 4 microM) was significantly higher compared with the IC50 value for inhibition of [14C]TEA (10 microM) uptake (5.3 +/- 1.7 microM). This finding suggests an allosteric interaction between transported cation and corticosterone. Because this substrate specific effect cannot be explained by differential replacement of corticosterone by MPP versus TEA and was observed after point mutations within the presumed substrate region, the data suggest that MPP or TEA bind to the substrate binding region simultaneously with corticosterone and cause a short-range allosteric effect on the corticosterone binding site. PMID- 15662046 TI - Role of glycosylation in trafficking of Mrp2 in sandwich-cultured rat hepatocytes. AB - The multidrug resistance-associated protein (MRP) family plays a major role in the hepatic excretion of organic anions. The expression, localization, and function of Mrp2 (Abcc2), a canalicular multispecific organic anion transport protein, were studied in sandwich-cultured rat hepatocytes. The amount of Mrp2 protein remained constant in sandwich-cultured rat hepatocytes over 4 days in culture, but the molecular mass increased approximately 10 kDa from 190 to 200 kDa. Mrp2 was internalized initially after hepatocyte isolation and was gradually sorted to the canalicular membrane. Disposition of 5-(6)-carboxy-2',7' dichlorofluorescein (CDF), an Mrp2 substrate, confirmed the changes in Mrp2 localization. CDF was localized predominantly inside hepatocytes at day 0 and gradually localized to the canalicular domain over time in culture. By day 4 in culture, CDF was localized exclusively in the canalicular networks. Tunicamycin, an inhibitor of glycosylation, decreased the molecular mass and simultaneously impaired the trafficking of Mrp2 to the canalicular membrane. Treatment of lysates from both day 0 (Mrp2, 190 kDa) and day 4 (Mrp2, 200 kDa) sandwich cultured rat hepatocytes with peptide N-glycosidase F, a deglycosylation agent, resulted in a band of 180 kDa, suggesting that Mrp2 from both day 0 and day 4 was glycosylated, but Mrp2 on day 4 was more glycosylated than on day 0. In conclusion, these data support the hypothesis that glycosylation of Mrp2 is responsible for the increase in molecular mass and may be involved in directing the canalicular localization of Mrp2 in sandwich-cultured rat hepatocytes over days in culture. PMID- 15662047 TI - H(2)O(2): a mediator of esophagitis-induced damage to calcium-release mechanisms in cat lower esophageal sphincter. AB - We previously reported that induction of acute experimental esophagitis by repeated perfusion of HCl may affect release of intracellular Ca(2+) stores. We therefore measured cytosolic Ca(2+) in response to a maximally effective dose of ACh in fura 2-AM-loaded lower esophageal sphincter (LES) circular muscle cells and examined the contribution of H(2)O(2) to the reduction in Ca(2+) signal. In normal cells, the ACh-induced Ca(2+) increase was the same in normal-Ca(2+) and Ca(2+)-free medium and was abolished by the phosphatidylinositol 4,5-bisphosphate specific phospholipase C inhibitor U-73122, confirming that the initial ACh induced contraction depends on Ca(2+) release from intracellular stores through production of inositol trisphosphate. In LES cells, the ACh-induced Ca(2+) increase in normal-Ca(2+) medium was significantly lower in esophagitis than in normal cells and was further reduced ( approximately 70%) when the cells were incubated in Ca(2+)-free medium. This reduction was partially reversed by the H(2)O(2) scavenger catalase. H(2)O(2) measurements in LES circular muscle showed significantly higher levels in esophagitis than in normal cells. When normal LES cells were incubated with H(2)O(2), the ACh-induced Ca(2+) increase was significantly reduced in normal-Ca(2+) and Ca(2+)-free medium and was similar to that observed in animals with esophagitis. The initial ACh-induced contraction was also reduced in normal cells incubated with H(2)O(2). H(2)O(2), when applied to cells at sufficiently high concentration, produced a visible and prolonged Ca(2+) signal in normal cells. H(2)O(2)-induced cell contraction was also sensitive to depletion of stores by thapsigargin (TG); conversely, H(2)O(2) reduced TG-induced contraction, suggesting that TG and H(2)O(2) may operate through similar mechanisms. Ca(2+)-ATPase activity measurement indicates that H(2)O(2) and TG reduced Ca(2+)-ATPase activity, confirming similarity of mechanism of action. We conclude that H(2)O(2) may be at least partly responsible for impairment of Ca(2+) release in acute experimental esophagitis by inhibiting Ca(2+) uptake and refilling Ca(2+) stores. PMID- 15662048 TI - Essential fatty acid deficiency in mice is associated with hepatic steatosis and secretion of large VLDL particles. AB - Essential fatty acid (EFA) deficiency in mice decreases plasma triglyceride (TG) concentrations and increases hepatic TG content. We evaluated in vivo and in vitro whether decreased hepatic secretion of TG-rich very low-density lipoprotein (VLDL) contributes to this consequence of EFA deficiency. EFA deficiency was induced in mice by feeding an EFA-deficient (EFAD) diet for 8 wk. Hepatic VLDL secretion was quantified in fasted EFAD and EFA-sufficient (EFAS) mice using the Triton WR-1339 method. In cultured hepatocytes from EFAD and EFAS mice, VLDL secretion into medium was measured by quantifying [(3)H]-labeled glycerol incorporation into TG and phospholipids. Hepatic expression of genes involved in VLDL synthesis and clearance was measured, as were plasma activities of lipolytic enzymes. TG secretion rates were quantitatively similar in EFAD and EFAS mice in vivo and in primary hepatocytes from EFAD and EFAS mice in vitro. However, EFA deficiency increased the size of secreted VLDL particles, as determined by calculation of particle diameter, particle sizing by light scattering, and evaluation of the TG-to-apoB ratio. EFA deficiency did not inhibit hepatic lipase and lipoprotein lipase activities in plasma, but increased hepatic mRNA levels of apoAV and apoCII, both involved in control of lipolytic degradation of TG-rich lipoproteins. EFA deficiency does not affect hepatic TG secretion rate in mice, but increases the size of secreted VLDL particles. Present data suggest that hypotriglyceridemia during EFA deficiency is related to enhanced clearance of altered VLDL particles. PMID- 15662049 TI - P2X and P2Y purinergic receptors on human intestinal epithelial carcinoma cells: effects of extracellular nucleotides on apoptosis and cell proliferation. AB - Extracellular nucleotides interact with purinergic receptors, which regulate ion transport in a variety of epithelia. With the use of two different human epithelial carcinoma cell lines (HCT8 and Caco-2), we have shown by RT-PCR that the cells express mRNA for P2X1, P2X3, P2X4, P2X5, P2X6, P2X7, P2Y1, P2Y2, P2Y4, P2Y6, P2Y11, and P2Y12 receptors. Protein expression for P2Y1 and P2Y2 receptors was also demonstrated immunohistochemically, and P2X receptor subtype protein was present in the following decreasing order: P2X4 > P2X7 > P2X1 > P2X3 > P2X6 > P2X5 >> P2X2. The functional presence of P2X7, P2Y1, P2Y2, and P2Y4 receptors was shown based on the effect of extracellular nucleotides on apoptosis or cell proliferation, and measurement of nucleotide-dependent calcium fluxes using a fluorometric imaging plate reader in the presence of different selective agonists and antagonists. ATP, at high concentrations, induced apoptosis through ligation of P2X7 and P2Y1 receptors; conversely, ATP, at lower concentrations, and UTP stimulated proliferation, probably acting via P2Y2 receptors. We therefore propose that stimulation or dysfunction of purinergic receptors may contribute at least partially to modulation of epithelial carcinoma cell proliferation and apoptosis. PMID- 15662050 TI - Ischemia of rat stomach mobilizes ECL cell histamine. AB - Microdialysis was used to study how ischemia-evoked gastric mucosal injury affects rat stomach histamine, which resides in enterochromaffin-like (ECL) cells and mast cells. A microdialysis probe was inserted into the gastric submucosa, and the celiac artery was clamped (30 min), followed by removal of the clamp. Microdialysate histamine was determined by enzyme-linked immunosorbent assay. In addition, we studied the long-term effects of ischemia on the oxyntic mucosal histidine decarboxylase activity in omeprazole-treated rats. Gastric mucosal lesions induced by the ischemia were enlarged on removal of the clamp. The microdialysate histamine concentration increased immediately on clamping (50-fold rise within 30 min) and declined promptly after the clamp was removed. In contrast, histidine decarboxylase activity of the ECL cells was lowered by the ischemia and returned to preischemic values 9 days later. Mast cell-deficient rats responded to ischemia-reperfusion much like wild-type rats with respect to histamine mobilization. Pretreatment with the irreversible inhibitor of histidine decarboxylase, alpha-fluoromethylhistidine, which is known to eliminate histamine from ECL cells, prevented the rise in microdialysate histamine. Pharmacological blockade of acid secretion (cimetidine or omeprazole) prevented the lesions induced by ischemia-reperfusion insult but not the mobilization of histamine. In conclusion, ischemia of the celiac artery mobilizes large amounts of histamine from ECL cells, which occurs independently of the gross mucosal lesions. The prompt reduction of the mucosal histidine decarboxylase activity in response to ischemia probably reflects ECL cell damage. The lesions develop not because of mobilization of histamine per se but because of ischemia plus reperfusion plus gastric acid. PMID- 15662052 TI - Role of networks in supporting emergency medicine research: findings from the Wessex emergency care research network (WECReN). PMID- 15662053 TI - Reporting of gunshot wounds by doctors in emergency departments: a duty or a right? Some legal and ethical issues surrounding breaking patient confidentiality. AB - Recent guidelines have been produced advising doctors working in emergency departments that they should report all gunshot injuries to the police (albeit with consent in all but very limited circumstances). This article will discuss some of the legal and ethical issues that surround breaking patient confidentiality in relation to gunshot wounds and other potentially dangerous patients; and looks at some cases from the UK and the USA where such issues have been ruled on. Finally, the issue of whether physicians do, or should, have a duty to warn when they feel that their patient may be dangerous will be discussed. PMID- 15662054 TI - Randomised controlled trial and cost consequences study comparing initial physiotherapy assessment and management with routine practice for selected patients in an accident and emergency department of an acute hospital. AB - OBJECTIVE: The Department of Health is reviewing the effectiveness of accident and emergency (A&E) departments. This study aimed to compare health and economic effects of physiotherapy initial assessment and management with routine practice in an A&E department. METHODS: Randomised controlled trial and cost and consequences study. Patients presenting at A&E were eligible if suspected at triage to have soft tissue injury without fracture. The efficacy end point was "days to return to usual activities". Secondary end points included patient satisfaction with their care and further health outcomes and cost data. RESULTS: 766 of 844 (915) patients were randomised. The median days before return to usual activities (available for 73% of those randomised) was greater in the physiotherapist group (41 days compared with 28.5 days; hazard ratio 0.85 p = 0.071). The physiotherapy group expressed greater satisfaction with their A&E care (on a scale of 1 to 5, median was 4.2 compared with 4.0, p<0.001), were more likely to be given advice and reassurance, and more likely to be provided with aids and appliances. Costs were the same between the two arms. CONCLUSION: There is evidence that physiotherapy leads to a prolonged time before patients return to usual activities. This study shows no clear danger from physiotherapy intervention and long term outcomes may be different but given these findings, a best estimate is that introducing physiotherapist assessment will increase costs to the health service and society. Routine care should continue be provided unless there is some reason why it is not feasible to do so and an alternative must be found. PMID- 15662055 TI - Who waits longest in the emergency department and who leaves without being seen? AB - OBJECTIVES: To determine which patient characteristics are associated with prolonged waiting times in the emergency department and which characteristics are associated with an increased risk of leaving without being seen. METHODS: Multivariate analysis of routine data collected at the Northern General Hospital, Sheffield between 1 January and 31 December 2001. Patient age, sex, triage priority, postcode, initiator of attendance, mode of arrival, time, day, and month of presentation were examined as potential predictors of waiting time and risk of leaving without being seen. RESULTS: Waiting time data for 71,331 patients were analysed, along with a further 5512 patients who left without being seen. Older patients and those with lower triage priority had longer waiting times, while ambulance borne patients had slightly shorter waiting times. Sex, source of referral, and postcode did not predict waiting times. The most powerful predictors of waiting time related to time of presentation, with longer waits being associated with presentation at night, on Mondays or Sundays, and during autumn months. Patients who left without being seen were more likely to be younger, male, lower triage priority, non-ambulance borne, self referred, and presenting at the times when waiting times were longest. CONCLUSION: Time of presentation, rather than individual patient characteristics, seem to be the most powerful predictors of waiting time. This suggests that concerns about inequity of waiting times should be addressed by reorganisation of staff duty rosters. PMID- 15662056 TI - Emergency department patients' opinions of screening for intimate partner violence among women. AB - BACKGROUND: Universal screening for intimate partner violence (IPV) in the emergency department (ED) has been advocated by many medical institutions. Policies implemented for IPV screening have met with numerous obstacles. One such obstacle is the perception by emergency personnel that patients might be offended by such screening if they presented to the ED for problems unrelated to trauma. OBJECTIVES: To assess opinions of adult ED patients regarding a policy of universal IPV screening for women presenting to the ED. METHODS: This study was conducted in EDs in Halifax, Nova Scotia, and St John's, Newfoundland. Patients were questioned as to whether it was appropriate for all women to be asked if they had experienced violent or threatening behaviour from someone close to them. Patients in significant pain or in extremis were not approached. RESULTS: The data consist of a convenience sample of 514 adult ED patients, aged 16-95 years. Two (0.4%) were excluded from the analysis. Of 512 analysed, 442 (86.0%) answered "yes" to the question, 53 (10.3%) answered "no", 17 (3.3%) had no opinion. There were no significant differences between the proportion of "yes" and "no" answers in the male and female groups. CONCLUSION: Universal screening for IPV of adult female patients presenting to the ED was supported by most patients. Patient objections should not be seen as a reason to withhold questioning on this issue. PMID- 15662057 TI - Can an airway assessment score predict difficulty at intubation in the emergency department? AB - AIM: To assess whether an airway assessment score based on the LEMON method is able to predict difficulty at intubation in the emergency department. METHODS: Patients requiring endotracheal intubation in the resuscitation room of a UK teaching hospital between June 2002 and September 2003 were assessed on criteria based on the LEMON method. At laryngoscopy, the Cormack and Lehane grade was recorded. An airway assessment score was devised and assessed. RESULTS: 156 patients were intubated during the study period. There were 114 Cormack and Lehane grade 1 intubations, 29 grade 2 intubations, 11 grade 3 intubations, and 2 grade 4 intubations. Patients with large incisors (p<0.001), a reduced inter incisor distance (p<0.05), or a reduced thyroid to floor of mouth distance (p<0.05) were all more likely to have a poor laryngoscopic view (grade 2, 3, or 4). Patients with a high airway assessment score were more likely to have a poor laryngoscopic view compared with those patients with a low airway assessment score (p<0.05). CONCLUSIONS: An airway assessment score based on criteria of the LEMON method is able to successfully stratify the risk of intubation difficulty in the emergency department. Patients with a poor laryngoscopic view (grades 2, 3, or 4) were more likely to have large incisors, a reduced inter-incisor distance, and a reduced thyroid to floor of mouth distance. They were also more likely to have a higher airway assessment score than those patients with a good laryngoscopic view. PMID- 15662058 TI - Mild head injury: reliability of early computed tomographic findings in triage for admission. AB - OBJECTIVE: To search the literature for case reports on adverse outcomes in patients with mild head injury where acute computed tomography (CT) findings had been normal. METHODS: Mild head injury was defined as head trauma involving amnesia or loss of consciousness, but where neurological findings are normal on arrival at hospital (GCS 15). The scientific literature was systematically searched for case reports where an early CT was normal and the patient deteriorated within two days. In these cases, early discharge despite a normal CT head scan would have been hazardous. RESULTS: Two prospective studies were found that investigated the safety of early CT in 3300 patients with mild head injury, as were 39 reports on adverse outcomes describing 821 patients. In addition, 52 studies containing over 62 000 patients with mild head injury were reviewed. In total, only three cases were deemed to have experienced an early adverse outcome despite a normal CT and GCS 15 on initial presentation. In another eight cases with incomplete descriptions, the interpretation was doubtful. Many reports of complications were not relevant to our question and excluded. These reports included cases with more severe head injury/not GCS 15 at presentation, complications that occurred after more than two days, or initial CT findings that were not fully normal. CONCLUSION: Very few cases were found where an early adverse event occurred after normal acute CT in patients with mild head injury. The strongest scientific evidence available at this time shows that a CT strategy is a safe way to triage patients for admission. PMID- 15662060 TI - Plastic baton round injuries. AB - OBJECTIVES: To review the injuries resulting from a new plastic baton round. METHODS: Review of case notes of patients presenting with injuries caused by plastic baton rounds over a four month period in Northern Ireland. RESULTS: Twenty nine patients were identified, 28 with 30 injuries were included in the study. Eighty nine per cent were male; the average age was 24.3 years. Seven patients required admission. There were no fatalities. Five injuries were to the upper limbs and 16 to the lower limbs. Three patients sustained pulmonary contusions. There were no head injuries. CONCLUSIONS: Although the numbers in this study are small it should be noted that no patient suffered a face, neck, or head injury. This is in contrast with previous studies in which up to 41.4% of attendances were for face, neck, or head injuries. In this study there were seven injuries to the trunk. Of the 14 deaths attributable to plastic baton rounds in Northern Ireland, all have been the result of head or chest trauma. The use of plastic baton rounds has decreased and, while a reduction in head injuries is noted, potentially serious chest injuries are still occurring. It is vital that guidelines on firing are adhered to. A large proportion of people who have been struck by plastic baton rounds do not attend an accident and emergency department and therefore doctors must be aware of patients with potentially serious injuries presenting late. PMID- 15662059 TI - Non-collision injuries in public buses: a national survey of a neglected problem. AB - OBJECTIVES: This prospective, nationally representative, multi-centre study was undertaken to assess non-collision injuries sustained by public bus passengers in Israel. METHODS: The emergency departments (EDs) of six medical centres, which participated in this eight month study, were chosen to represent both urban and rural catchment areas. All patients diagnosed with injuries sustained on a public bus not involved in a road traffic accident were promptly evaluated for mechanism and nature of injury and demographic parameters. RESULTS: The study cohort consisted of 120 patients (86 were female, 34 were male, age range 3-89 years). Over half were older than 55 years. The most common injuries were to the limbs, vertebral column, and head. The major mechanism of injury was acceleration/deceleration. Most patients were standing when they sustained the injuries. There were no fatalities, and 17 patients were admitted to hospital (9 of 17, 52% older than 55 years). Extrapolation to yearly national statistics suggests a probable total of 729 such injuries. CONCLUSION: The significant injuries inflicted on passengers of public buses not involved in road traffic accidents warrant decisive preventative measures by transportation authorities. PMID- 15662061 TI - Evaluation of blunt abdominal trauma: current practice in Taiwan. AB - OBJECTIVE: To gain an overview of the current practice of different major institutions in Taiwan in the evaluation of abdominal injuries. A further comparison was made between general surgeons and emergency physicians in this aspect. METHOD: A telephone survey was conducted of all emergency departments of 58 major institutions (14 medical centres, 44 district hospitals) that are capable of providing definitive care for trauma victims in Taiwan in June 2002. Respondents were asked to select the diagnostic modality of choice in the evaluation of a haemodynamically abnormal blunt trauma victim with suspected intra-abdominal injuries. In the same study period, this particular telephone scenario was also used to survey 109 individual doctors (45 emergency physicians, 64 general surgeons). RESULTS: Most respondents preferred ultrasound (also known as focused assessment with sonography for trauma or "FAST") instead of diagnostic peritoneal lavage (DPL) because DPL is invasive and most doctors in Taiwan have limited experience in performing DPL or interpreting the results. CONCLUSIONS: It seems reasonable to devote greater resources for emergency departments to incorporate a FAST based algorithm into their initial management of trauma victims, and to improve training in its use. It is also suggested that future ATLS teaching in Taiwan should include didactic material on FAST. PMID- 15662062 TI - Pilot study of a paediatric emergency department oral rehydration protocol. AB - OBJECTIVE: To ascertain if the use of a paediatric oral rehydration protocol in the emergency department changed rates of admission, total time spent in hospital, total time spent in the emergency department, or number of unscheduled returns to the emergency department. METHODS: A two month time period was analysed before and then after the adoption of an oral rehydration protocol for children presenting to the emergency department with symptoms suggestive of gastroenteritis. The rates of admission, total time spent in hospital, total time spent in the emergency department, and the rate of unscheduled returns were analysed using Fisher's exact and Wilcoxon testing for non-parametric data. RESULTS: The adoption of a paediatric rehydration protocol was associated with a significant reduction in admission rates (before = 22.5%, after 5.1%; p = 0.048) and mean total hospital time (before = 7 hours 54 minutes, after = 2 hours 17 minutes; p = 0.017). There was no significant difference in time spent in the emergency department (before = 1 hour 25 minutes, after 1 hour 35 minutes, p = 0.3). The number of unscheduled returns did not change significantly (p = 0.3). CONCLUSION: Adoption of a paediatric rehydration protocol significantly reduces admission rates and total time spent in hospital for children presenting to the emergency department with symptoms of gastroenteritis. The mean time spent in the actual emergency department does not significantly increase. The rate of unscheduled returns does not change. PMID- 15662063 TI - Snake envenomation in a north Indian hospital. AB - OBJECTIVES: To study the clinical profile of snake envenomation in a tertiary referral north Indian hospital. METHODS: Retrospective case note analysis of all cases of snakebite admitted to the medical emergency from January 1997 to December 2001. RESULTS: Of a total of 142 cases of snakebite there were 86 elapid bites presenting with neuroparalytic symptoms and 52 viper bites having haemostatic abnormalities. Some 60.6% of the cases of snakebite occurred when the patient was asleep. Urban to rural ratio was 1:4.7 and male to female ratio was 4.25:1. Median time to arrival at our hospital after the bite was nine hours and mean duration of hospital stay was eight days. Twenty seven cases had acute renal failure and 75% of all elapid bites required assisted ventilation. Seventeen of 119 patients who received antivenom had an adverse event. The average dose of antivenom was 51.2 vials for elapid bites and 31 vials for viper bites. Overall mortality rate was 3.5%. CONCLUSION: Snakebites are common in the rural population of developing countries. There is a need to educate the public about the hazards of snakebite, early hospital referral, and treatment. PMID- 15662064 TI - Best evidence topic report. Timing of lumbar puncture in suspected subarachnoid haemorrhage. AB - A short cut review was carried out to establish how long after onset of headache a lumbar puncture should be carried out to rule out subarachnoid haemorrhage. Altogether 142 papers were found using the reported search, of which one presented the best evidence to answer the clinical question. The author, date and country of publication, patient group studied, study type, relevant outcomes, results and study weaknesses of this best paper are tabulated. A clinical bottom line is stated. PMID- 15662066 TI - Best evidence topic report. Proteolytic enzymes for oesophageal meat impaction. AB - A short cut review was carried out to establish whether proteolytic enzymes are effective at resolving oesophageal meat impaction. Altogether 98 papers were found using the reported search, of which three presented the best evidence to answer the clinical question. The author, date and country of publication, patient group studied, study type, relevant outcomes, results and study weaknesses of these best papers are tabulated. A clinical bottom line is stated. PMID- 15662067 TI - Best evidence topic report. Effervescent agents for oesophageal food bolus impaction. AB - A short cut review was carried out to establish whether fizzy drinks alone are effective at resolving food bolus impaction. Altogether 46 papers were found using the reported search, of which six presented the best evidence to answer the clinical question. The author, date and country of publication, patient group studied, study type, relevant outcomes, results and study weaknesses of these best papers are tabulated. A clinical bottom line is stated. PMID- 15662068 TI - Best evidence topic report. Humeral fractures and non-accidental injury in children. AB - A short cut review was carried out to establish whether proximal humeral fractures in children are indicative of non-accidental injury. Altogether 44 papers were found using the reported search, of which two presented the best evidence to answer the clinical question. The author, date and country of publication, patient group studied, study type, relevant outcomes, results and study weaknesses of these best papers are tabulated. A clinical bottom line is stated. PMID- 15662069 TI - Best evidence topic report. Torn frenulum and non-accidental injury in children. AB - A short cut review was carried out to establish whether a torn frenulum in a child is indicative of non-accidental injury. Altogether 104 papers were found using the reported search, of which none presented any evidence to answer the clinical question. It is concluded that there is no evidence available to answer this question. Further research is needed. PMID- 15662070 TI - SOCRATES 4 (synopsis of Cochrane reviews applicable to emergency services). PMID- 15662071 TI - 10 assessment and care of ENT problems. PMID- 15662072 TI - Effect of automatic external defibrillator audio prompts on cardiopulmonary resuscitation performance. AB - OBJECTIVES: To determine the effectiveness of the cardiopulmonary resuscitation (CPR) audio prompts in an automatic external defibrillator in 24 lay subjects, before and after CPR training. METHODS: Untrained subjects were asked to perform CPR on a manikin with and without the assistance of audio prompts. All subjects were then trained in CPR, and retested them eight weeks later. RESULTS: Untrained subjects who performed CPR first without audio prompts performed poorly, with only (mean (SD)) 24.5% (32%) of compressions at the correct site and depth, a mean compression rate of 52 (31) per minute, and with 15% (32%) of ventilatory attempts adequate. Repeat performance by this group with audio prompts resulted in significant improvements in compression rate (91(12), p = 0.0002, paired t test), and percentage of correct ventilations (47% (40%), p = 0.01 paired t test), but not in the percentage correct compressions (23% (29%)). Those who performed CPR first with audio prompts performed significantly better in compression rate (87 (19), p = 003, unpaired t test), and the percentage of correct ventilations (51 (34), p = 0.003 unpaired t test), but not in the percentage of correct compressions (18 (27)) than those without audio prompts. After training, CPR performance was significantly better than before training, but there was no difference in performance with or without audio prompts, although 73% of subjects commented that they felt more comfortable performing CPR with audio prompts. CONCLUSIONS: For untrained subjects, the quality of CPR may be improved by using this device, while for trained subjects the willingness to perform CPR may be increased. PMID- 15662073 TI - Prehospital care in Indonesia. AB - BACKGROUND: Indonesia is a huge, diverse, and developing country that until recently had no public ambulance service let alone a system of prehospital care. It commonly experiences many natural disasters, manmade conflicts, and violence as well as the daily emergencies seen worldwide. CURRENT SYSTEM: Hospitals of varying standards are widespread but have no system of emergency ambulance or patient retrieval. Indonesia's only public emergency ambulance service, 118, is based in five of the biggest cities and is leading the way in paramedic training and prehospital care. CHALLENGES AND DEVELOPMENTS: There are many challenges faced including the culture of acceptance, vast geographical areas, traffic, inadequate numbers of ambulances, and access to quality training resources. Recently there have been a number of encouraging developments including setting up of a disaster response brigade, better provision of ambulances, and development of paramedic training. CONCLUSIONS: An integrated national regionalised hospital and prehospital system may seem fantastic but with the enthusiasm of those involved and perhaps some help from countries with access to training resources it may not be an unrealistic goal. PMID- 15662074 TI - Penetrating injury to the soft palate causing retropharyngeal air collection. AB - Pharyngeal injuries caused by trauma are common and have been reported previously in the medical literature. In some cases of a penetrating injury there is a collection of air in the retropharyngeal space that can be shown on lateral soft tissue radiography of the neck. If this condition is not diagnosed or adequately treated the patient may develop severe complications such as mediastinitis. A case is reported of a patient with penetrating injury caused by a curtain rail and the subsequent treatment is described. PMID- 15662075 TI - Accessory ossicle or intraepiphyseal fracture of lateral malleolus: are we familiar with these? AB - A case of intraepiphyseal injury (type 7) to the lateral malleolus in a 11 year old child is described. This rare injury cannot be classified by commonly used Salter Harris classification for epiphyseal injury. Although less common, accessory ossicle of the malleoli is an important differential diagnosis for such injury. Details of type 7 intraepiphyseal injuries and accessory ossicle are described. PMID- 15662076 TI - Severe interscapular pain and increased creatine kinase activity: the answer was in the ankles. AB - Severe thoracic back pain with increased creatine kinase activity is a clinical presentation that suggests a variety of life threatening conditions. If initial examination is unrevealing, multiple diagnostic tests are usually performed attempting to identify the origin of the problem, sometimes neglecting apparently unrelated subtle physical findings. A patient is described in whom this was the initial presentation of a sensory demyelinating neuropathy, resulting in a diagnostic challenge. This case expands the differential diagnosis of severe thoracic back pain and increased creatine kinase activity, and illustrates the importance of physical examination in reaching a final diagnosis. PMID- 15662077 TI - Life threatening hyperkalaemia with diarrhoea during ACE inhibition. AB - A 67 year old woman developed acute renal failure with serum potassium 9.4 mmol/l requiring emergency dialysis after seven days of diarrhoea while taking an ACE inhibitor for vascular disease. Review of the literature, the British National Formulary, and the patient information leaflets for each of the 11 ACE inhibitors currently marketed in the UK suggests that this potentially life threatening complication of ACE inhibition is not yet widely recognised. PMID- 15662078 TI - Appropriate analysis and reporting of cluster randomised trials. PMID- 15662079 TI - Brain dopamine receptors--research perspectives and potential sites of regulation. AB - Dopamine mediates its biological actions via at least five distinct G protein coupled receptors. Recently, significant progress has been made in the understanding of the molecular and cellular consequences of activation of the different dopamine receptors. Not only their anatomical localization has been revisited, what implicates new insights into well known D1/D2 receptor synergism, but also evidence has been provided that dopamine receptors can physically interact with each other as well as with other neurotransmitter receptors. Dopamine receptors are subject to phosphorylation by G protein-coupled receptor kinases, the specificity of which has started to be revealed. Some progress has been made in elucidating the functional significance of polymorphisms observed in genes encoding dopamine receptor. All these new findings need to be appreciated by the psychopharmacologists in order to better understand the behavioral consequences of activation of dopamine receptors. PMID- 15662080 TI - Neurogenesis in the adult brain. AB - Neurogenesis is a process that involves cell proliferation, migration and differentiation. Adult neurogenesis has been discovered by Altman in the mid 1960s. It is known now that neurogenesis occurs in two main neurogenic areas of the adult mammalian brain: the olfactory bulb and the hippocampal dentate gyrus, although other brain regions, such as cortex or substantia nigra cannot be excluded. The rate of neurogenesis can be regulated in a positive and negative manner by several factors like, age, growth factors, hormones, environmental or pharmacological stimuli. Functional significance of adult neurogenesis is still under investigation, however, several evidences suggest involvement of newly generated neurons in cognitive processes. There are also several findings indicating that the impairment of adult neurogenesis may be involved in the pathophysiology of some brain diseases, like depression, epilepsy, ischemia or neurodegenerative disorders. It appears that alterations in the rate of neurogenesis may have important functional and therapeutic implications. PMID- 15662081 TI - Apoptosis in the central nervous system: Mechanisms and protective strategies. AB - Naturally occurring neuronal cell death, which is essential for normal development, tissue homeostasis and as a defence against pathogens, is generally considered to be apoptotic. Apoptosis, called also a programmed cell death, though the meanings of these terms are not identical, is a common phenomenon induced either by specific insults mediated through so-called "death receptors" (external pathway) or by non-specific insults leading to reduction of mitochondrial potential (internal pathway). At the molecular level, apoptosis is regulated by the activation of the caspase cascade, which depends either upon the participation of mitochondria and the activation of procaspase-9 or on the interaction of a death receptor with its ligand and subsequent activation of procaspase-8. Considering various mechanisms of programmed cell death, either during neural development or neurodegeneration, finding the most suitable and effective protective strategy needs evaluation of risk-to-benefit ratio. Unlike malignant tissues, neuronal cells have to be protected, but only if the protection does not disturb natural cell elimination. Defining molecular mechanisms is, therefore, the necessary step preceding the adaptation of new protective and then, possibly, therapeutic strategies. At present, the most promising are discoveries related to caspases, JNK, and GSK-3beta and their crucial role in stress-dependent and spontaneous apoptosis. Inhibitors of caspases and JNKs succeed in defending cells against kainate-, beta-amyloid- or MPTP-induced neuronal cell death. Among protective agents those, which easily cross the blood-brain barrier and exert profound neuroprotective action without side effects, seem to be the most important. PMID- 15662082 TI - Potential role of the brain dopaminergic system in the regulation of cytochrome P 450 expression. AB - Although there are no literature data, which would clearly describe the role of catecholaminergic systems (including dopaminergic system) in the expression of the liver cytochrome P-450, there are well documented reports on physiological regulation of cytochrome P-450 by endogenous hormones (glucocorticoids, sex hormones, thyroid hormones, growth hormone, insulin) and by the immune system (cytokines), and all these factors remain under central nervous system control. Therefore, one can expect that dysfunction of catecholaminergic systems may lead to significant changes in activities of particular cytochrome P-450 isoenzymes by altering levels of endogenous hormones. The mechanism underlying dopamine-induced regulation of the liver cytochrome P-450 expression seems not to be direct, but rather mediated by pituitary hormones and cytokines. PMID- 15662083 TI - MPEP, mGlu5 receptor antagonist, regulates NPYmRNA expression in hippocampal and amygdalar neurons. AB - Our earlier studies showed that the metabotropic glutamate receptor 5 (mGluR5) antagonist, MPEP, could regulate neuropeptide Y (NPY) neurons in the amygdala, influencing both peptide expression and its antianxiety effects. Two brain structures are particularly engaged in a regulation of anxiety, namely the amygdala and also the hippocampus. They both belong to the limbic system and contain NPY neurons and mGlu5 receptors. Therefore, in the present study, we examined the effect of MPEP on NPY and NPYmRNA expression in the amygdala and the hippocampus of the rat brain. NPY expression was studied by immunohistochemical method, and radioimmunoassay, and the NPY synthesis was examined using NPYmRNA in situ hybridization. Immunohistochemical localization of mGluR5 was also carried out. It was found that MPEP given 3 times every 8 h potently decreased NPYmRNA expression 30 min after the last dose in both those structures (to 8-20% of the control level). After single MPEP treatment, we did not observe any changes in NPYmRNA level in the hippocampus, and its decrease in the amygdala 6 h after MPEP administration. The obtained results suggest a positive regulatory control of NPY synthesis by mGlu5 receptors in hippocampal and amygdalar neurons. PMID- 15662084 TI - Involvement of nitricoxidergic system in the hypnotic effects of benzodiazepines in mice. AB - The influence of nitric oxide (NO) on hypnotic activity of diazepam, chlordiazepoxide and clonazepam was studied in mice. Administration of both non selective NO synthase inhibitors: N(G)-nitro-L-arginine methyl ester (L-NAME), N(G)-nitro-L-arginine (L-NOARG) and selective NO synthase inhibitor 7 nitroindazole (7-NI) resulted in significant increase in the duration of diazepam , chlordiazepoxide- and clonazepam-induced sleep. The effects of co administration of the examined inhibitors with benzodiazepines were not changed by L-arginine, a substrate for NO formation. Administration of L-arginine alone had no effect on the duration of sleep induced by benzodiazepines. Methylene blue, the guanyl cyclase inhibitor, was able to increase the duration of benzodiazepine-induced sleep. These findings suggest that the cGMP/NO system may participate in hypnotic effects of benzodiazepines. PMID- 15662085 TI - Inhibition of rodent brain monoamine oxidase and tyrosine hydroxylase by endogenous compounds - 1,2,3,4-tetrahydro-isoquinoline alkaloids. AB - Four different noncatecholic and one catecholic tetrahydroisoquinolines (TIQs), cyclic condensation derivatives of beta-phenylethylamine and dopamine with aldehydes or keto acids, were examined for the inhibition of rat and mouse brain monoamine oxidase (MAO) and rat striatum tyrosine hydroxylase (TH) activity. Simple noncatecholic TIQs were found to act as moderate (TIQ, N-methyl-TIQ, 1 methyl-TIQ) or weak (1-benzyl-TIQ), MAO B and MAO A inhibitors. 1-Methyl-TIQ inhibited more potently MAO-A than MAO-B; the similar but more modest effect was exerted by salsolinol. Only salsolinol markedly inhibited TH activity, being competitive with the enzyme biopterin cofactor. The inhibition of MAO and TH by TIQs is discussed in relation to their ability to regulate monoamine metabolism. PMID- 15662086 TI - Effects of joint administration of imipramine and amantadine in patients with drug-resistant unipolar depression. AB - The paper describes the effect of amantadine (AMA) supplementation on imipramine (IMI) therapy in patients (with treatment-resistant unipolar depression) who fulfilled DSM IV criteria for major depression. Twelve patients were enrolled to the study on the basis of history of their illness and therapy. Following 2 weeks of washout period, the patients were treated with IMI twice daily (100-150 mg/day) for 6 weeks, and then AMA was introduced (twice daily, 100-150 mg/day) and administered jointly with IMI for further 6 weeks. Thereafter, AMA was withdrawn, and the patients were treated with IMI alone for 2 weeks. Hamilton Depression Rating Scale (HDRS) and Beck Depression Inventory (BDI) were used to assess efficacy of antidepressant therapy. IMI changed neither HDRS nor BDI score after 3 or 6 weeks of treatment when compared with washout (before treatment). AMA supplementation significantly reduced both HDRS and BDI scores after 3- or 6 week supplementation. AMA augmentation of IMI treatment was beneficial and lasted even after AMA withdrawal. Moreover, pharmacokinetic data indicate that AMA did not influence significantly the plasma concentration of the IMI and its metabolite, desipramine, in the patients during joint treatment with AMA and IMI, what suggests the lack of pharmacokinetic interaction. These results suggest that joint therapy with IMI and AMA may be successful in the treatment-resistant unipolar depression. PMID- 15662087 TI - New 4-[omega-(diarylmethylamino)alkyl]- and 4-[omega-(diarylmethoxy)alkyl]-1 arylpiperazines as selective 5-HT1A/5-HT2A receptor ligands with differentiated in vivo activity. AB - Two series of novel 4-ethyl- or 4-propyl-1-arylpiperazines (5-12) with the 4,4' disubstituted diphenylmethylamino (series a) or the diphenylmethoxy (series b) terminal fragment were synthesized and evaluated for their binding affinity at 5 HT1A and 5-HT2A receptors. The influence of the introduction of 4-methyl, 4 chloro or 4-fluoro substituents at both phenyl rings of that terminal moiety on in vitro and in vivo 5-HT1A receptor activity of those modified compounds was discussed. Compounds 5a, 6a, 9a-12a, 5b, 6b, 9b, 11b and 12b displayed high to fairly high affinity for 5-HT1A receptors (Ki = 2.4-72 nM). Compounds of both series showed low or very low 5-HT2A receptor affinity (Ki = 155-5400 nM). Amines 5a, 6a, 11a, and their ether analogs 5b, 6b and 11b, also possessed high or moderate alpha(1)-adrenoceptor affinity (K(i) = 6-104 nM). The functional activity of compounds 5a, 6a, 9a-12a, 5b, 8b, 9b, 11b and 12b was tested in vivo in the commonly used animal models. The majority of those ligands behaved like 5 HT1A receptor antagonists, their influence on the pre- and/or postsynaptic sites being diverse, though. They exhibited characteristics of partial agonists of postsynaptic 5-HT1A receptors (11a), of weak antagonists of pre- and postsynaptic sites (12a, 9b), of antagonists of presynaptic (5a) or of antagonists of postsynaptic 5-HT(1A) receptors (9a, 10a, 5b, 8b, 11b and 12b) while, 6a was devoid of functional activity at those receptors. The above findings indicate that introduction of 4-methyl, 4-chloro or 4-fluoro substituents to the diphenylmethyl part of the 1-(2-methoxyphenyl)piperazines tested in vivo may modify their 5-HT1A receptor functional activity. PMID- 15662088 TI - Synthesis and cardiovascular activity of new 8-alkylamino-1,3-dimethyl-7-(2 hydroxy-3- piperazinopropyl)-3,7-dihydro-1H-purine-2,6-diones. AB - 7-{2-Hydroxy-3-[4-(2-phenoxyethyl)-piperazinyl-1-yl]-propyl}-1,3-di-methyl-3,7 dihydro-1H-purine-2,6-dione dihydrochloride (2), and several of its 8-alkylamino substituted derivatives (11-17) were synthesized and tested for electrocardiographic, antiarrhythmic and hypotensive activity. Also their alpha(1)- and alpha(2)-adrenoreceptor affinities were determined. It was found that compound 2, and its analogue 15 with 8-(2-morpholin-4-yl-ethylamino) substituent displayed a strong prophylactic antiarrhythmic activity in experimentally induced arrhythmia (LD50/ED50 = 54.9 and 55.0, respectively). The hypotensive activity was observed for 8-benzylamino (11) or 8-(pyridin-2-yl methylamino) (12) analogues. All the new derivatives (11-17) and 2 showed a weak affinity for alpha1-(Ki = 0.225-1.400 microM) and alpha2-(Ki = 0.152-4.299 microM) receptors. PMID- 15662089 TI - Antiarrhythmic profile and endothelial action of novel decahydroquinoline derivatives. AB - We tested antiarrhythmic and endothelial action of novel decahydroquinoline derivatives. Antiarrhythmic activity was analyzed using models of aconitine-, calcium chloride-, and adrenaline-induced arrhythmias in rats. Potency to induce nitric oxide (NO)-dependent coronary vasodilation was assessed in isolated guinea pig heart perfused according to Langendorff technique. Among 15 novel decahydroquinoline derivatives (D1-15), four of them displayed antiarrhythmic activity (D12-D15). D12-D15 compounds were more active in the model of aconitine induced arrhythmias than in calcium chloride-induced arrhythmias and were inactive in the model of adrenaline-induced arrhythmias. Profile of antiarrhythmic activity of D12-D15 compounds was similar to that of quinidine and procainamide. Interestingly, in the isolated guinea pig heart D14 and D15 (10(-5) M) induced coronary vasodilation, that was mediated by endothelium-derived NO. In conclusion, novel decahydroquinoline derivatives described here (D12-D15) show antiarrhythmic activity typical of antiarrhythmic drugs of class I. Importantly, some of these compounds (D14, D15) release NO from coronary endothelium, which may provide an additional therapeutic benefit. PMID- 15662090 TI - Anti-inflammatory effects of selective COX-2 inhibitors. AB - In this study, effects of rofecoxib, celecoxib, nimesulide on the acute phase of inflammation were studied in the carrageenan-induced paw edema model and their influence on the chronic phase of inflammation was evaluated in the cotton pellet granuloma tests. Additionally, effects of these drugs on capillary vascular permeability were examined in the hyaluronidase test and were compared with that of indomethacin (nonselective COX inhibitor). The results of the study demonstrated that rofecoxib, celecoxib, nimesulide, indomethacin at a dose of 10 mg kg(-1) reduced the volume of paw edema by 40.6% (p < 0.05), 21.6% (p < 0.05), 20.3% (p < 0.05), 64.0% (p < 0.05), respectively. Anti-proliferative effect of rofecoxib was of 29%, while those of celecoxib and nimesulide were of 13.5 and 21.2%, respectively. Indomethacin had an anti-proliferative effect of 44.2%. When the drugs were given at a dose of 25 mg kg(-1) rofecoxib, celecoxib, nimesulide reduced carrageenan-induced paw edema by 50.6% (p < 0.004), 27.9% (p < 0.004) and 33.0% (p < 0.004), respectively. Positive control, indomethacin, reduced the paw edema by 86.1% (p < 0.004). As a result, indomethacin, rofecoxib, celecoxib, nimesulide significantly inhibited both acute and chronic inflammation. While indomethacin, celecoxib, nimesulide significantly reduced capillary vascular permeability, the effect of rofecoxib was insignificant. We could not clarify this observation. Further studies are required to enlighten this effect of rofecoxib. PMID- 15662091 TI - Effect of simvastatin and fluvastatin on plasma fibrinogen levels in patients with primary hypercholesterolemia. AB - The aim of the study was to compare the effect of treatment with two different statins on plasma fibrinogen levels in patients with primary isolated hypercholesterolemia. Sixty three patients enrolled into the study were randomly divided into two groups, treated with simvastatin (20 mg/d) or fluvastatin (40 mg/d), respectively. Plasma lipid profile and fibrinogen levels were measured after 4 and 12 weeks of the therapy. Both drugs decreased total and LDL cholesterol and apoprotein B levels. Simvastatin additionally reduced triglyceride levels. After 4 weeks of treatment both drugs tended to increase plasma fibrinogen levels, while after 12 weeks fibrinogen level was significantly increased in the simvastatin-treated patients. The effect on fibrinogen did not correlate with their lipid-lowering potential, gender and was similar in patients positive and negative for anti-Helicobacter pylori or anti-Chlamydia pneumoniae antibodies. Our results support the findings about a relatively weak effect of statin therapy on plasma fibrinogen level and partially explain contradictory results of previous studies. PMID- 15662092 TI - Hypoxic and pharmacological preconditioning preserves vasomotor response of porcine coronary artery. AB - Vasomotor response of the coronary artery depends on both endothelial and smooth muscle cells. Response is altered by hypoxia-reoxygenation-induced damages. Hypoxic preconditioning and pharmacological preconditioning as well can prevent these alterations. We compared the effectiveness of both types of preconditioning against hypoxia-reoxygenation-induced changes in vasomotor response of the isolated artery. Porcine arterial rings (3-4 mm wide) were cut from the left anterior descending porcine coronary artery and placed in Krebs-Henseleit solution. In order to obtain control response of the arteries, we contracted arterial rings with 20 mM KCl before ("standard contraction") and after 60-min hypoxia and 30-min reoxygenation. In other groups, nitric oxide-synthase and cyclooxygenase were inhibited. Then, the rings were pre-contracted with U46619 and relaxed by cumulative addition of the substance P. Contractions and relaxations of non-preconditioned and hypoxically or pharmacologically preconditioned rings were compared. Hypoxic preconditioning was performed by two periods of 5-min hypoxia and 10-min reoxygenation. For pharmacological preconditioning, we used application of adenosine, adrenaline, acetylcholine and angiotensin II. Analysis was performed with one-way ANOVA, followed by Dunnett's Multiple Comparison Test. After hypoxia-reoxygenation, in non-preconditioned rings KCl-induced contractions were significantly increased compared to standard contraction. Relaxations of hypoxically and pharmacologically preconditioned rings (expressed as percentages of U46619-induced pre-contraction) were significantly decreased (p < 0.01) compared to hypoxic but not to normoxic rings. Hypoxic and pharmacological preconditioning may preserve contraction and endothelium-dependent relaxation of porcine coronary artery after long-lasting hypoxia-reoxygenation. PMID- 15662094 TI - Changes and their regression in the osseous system in rats after administering a cytostatic drug inhibiting tumor cell division in the phase of DNA synthesis. AB - Chemotherapeutic drugs may disturb the bone tissue metabolism and cause osteopenia, however, the pathomechanism of the damaging effect of cytostatics on this tissue has not been well recognized so far. The detrimental effect may result from a direct cytotoxic action of these drugs on cells remodeling the bone, or on osteogenic cells present in the bone and in the bone marrow, or may be the result of hormonal disorder caused by impaired function of gonads. The aim of this study was to investigate the in vivo effect of 5-fluorouracil (5-FU), a cytostatic agent which inhibits tumor cell division in the phase of DNA synthesis, on the bone remodeling in rats and to examine whether the period of 4 weeks was sufficient for regression of changes elicited by administering 5-FU. Changes in the bone tissue following administration of 5-FU and their regression were evaluated by assessing macrometric and histomorphometric parameters as well as of mechanical properties of the femur. The tests were carried out on male Wistar rats. 5-FU was administered at the doses: 30 mg/kg per os (po) daily for 5 days every 2 weeks; 15 mg/kg im daily for 5 days every 2 weeks; 65 mg/kg im once weekly. Changes in the osseous tissue were examined 4 weeks after the first dose of 5-FU administration. Regression of the changes was examined 8 weeks after the first dose of 5-FU administration (the 5-FU was not administered between 30th and 57th day after the first dose of 5-FU administration). As a result of our research, it was established that 5-FU disturbed the bone remodeling processes in rats, mostly by impairing the process of new bone matrix synthesis, which leads to impaired mineralization process and decreased mechanical endurance of the femur. It was also established that the period of 4 weeks was not sufficient for regression of the changes in the osseous tissue caused by 5-FU administration. PMID- 15662093 TI - Role of adrenal gland hormones in the mechanism of antiulcer action of nimesulide and ranitidine. AB - In the present study, we investigated whether the antiulcer effects of nimesulide (100 mg kg(-1) and ranitidine (150 mg kg(-1) were dependent on the adrenal cortex hormones. The antiulcer effects of nimesulide and ranitidine were examined in the indomethacin-induced gastric ulcer model in rats (first experiment). The mean ulcer areas in the control and ranitidine-treated groups were 11.1 +/- 3.18, 1.4 +/- 1.11 mm2, respectively. There was not any gastric damage in nimesulide treated group. The mean ulcer area of control group (second experiment) administered metyrapone and indomethacin was 11.8 +/- 9.9, and it measured 2.0 +/ 1.41 mm2 in ranitidine-given group, while gastric damage was not observed in nimesulide-administered group. In adrenalectomized and indomethacin-treated rats (third experiment), the mean ulcer area was 17.9 +/- 11.5 mm2 in the nimesulide group, gastric ulcer was not seen in ranitidine group. In adrenalectomized rats (fourth experiment), the mean ulcer areas were 29 +/- 14.3, 23 +/- 11.2 and 1.3 +/- 2.4 mm2 in control group given indomethacin, only nimesulide or indomethacin + ranitidine, respectively. The obtained results indicated that adrenal cortex hormones played a role in antiulcer effect of nimesulide, but not ranitidine. PMID- 15662095 TI - Effect of administration of alendronate sodium and retinol on the mechanical properties of the femur in ovariectomized rats. AB - Alendronate sodium, an aminobisphosphonate with potent antiresorptive activity, is used in the treatment of postmenopausal osteoporosis. Retinol, as a component of multivitamin preparations, is frequently used especially by elderly people. There are no reports on the interaction of alendronate sodium and retinol. The aim of the present study was to investigate the effect of administration of alendronate sodium and retinol on mechanical properties of the femoral bone in bilaterally ovariectomized rats. The experiments were carried out on 3-month-old Wistar rats, divided into 7 groups: I - sham-operated control rats, II - ovariectomized control rats, III - ovariectomy + alendronate sodium 3 mg/kg po, IV - ovariectomy + retinol 700 IU/kg po, V - ovariectomy + retinol 3500 IU/kg po, VI - ovariectomy + alendronate sodium 3 mg/kg po + retinol 700 IU/kg po, VII - ovariectomy + alendronate sodium 3 mg/kg po + retinol 3500 IU/kg po. The drugs were administered to the rats daily by oral gavage for 28 days. Body mass gain, bone mass, bone mineral content and calcium content in the femur and L-4 vertebra and mechanical properties of the whole femur (extrinsic stiffness, ultimate load, breaking load, deformation caused by the ultimate load) and the neck of the femur (load at fracture), were examined. Bilateral ovariectomy induced osteopenic changes in the rat skeletal system. Alendronate sodium (3 mg/kg po) counteracted the development of osteopenia induced by ovariectomy. Retinol at both used doses unfavorably affected the examined bone parameters of ovariectomized rats. Retinol administered with alendronate sodium lessened the preventive action of alendronate on the development of osteopenic changes in the skeletal system of ovariectomized rats. PMID- 15662096 TI - Effect of imipramine treatment on plasma dopamine beta-hydroxylase activity in chronic mild stress in rats. AB - Dopamine beta-hydroxylase (DBH) which catalyzes conversion of dopamine into noradrenaline, may be a good blood marker of unipolar depression. Therefore, we studied the effect of classic antidepressant drug imipramine (10 mg/kg ip) on activity of this enzyme in plasma of rats subjected to chronic mild stress (CMS), the model of anhedonia. CMS induced reductions in DBH activity by the second day and 5th week of stress duration. Imipramine treatment minimized these CMS-induced reductions. The data indicate that, similarly to human depression, CMS also affects DBH activity, and, moreover, the CMS-induced alterations are normalized by imipramine treatment. PMID- 15662097 TI - Chronic treatment with citalopram does not affect the expression of alpha1 adrenergic receptor (alpha1-AR) subtypes. AB - We previously reported that chronic treatment with imipramine and electroconvulsive shock up-regulate the density and alpha1A-adrenergic receptor (alpha1A-AR) mRNA level in the rat prefrontal cortex, while the expression of the alpha1B subtype was unchanged. The present study examined whether repeatedly given citalopram, a selective serotonin reuptake inhibitor, induces any changes in the expression of alpha1A and alpha1B subtypes of alpha1-AR. The receptors density was assessed in the rat cerebral cortex by [3H]prazosin binding while the expression of alpha1A and alpha1B receptors' mRNA was measured in the rat prefrontal cortex by Northern blot analysis or competitive reverse transcription and polymerase chain reaction (RT-PCR), respectively. We did not find any changes in alpha1A- and alpha1B-AR density or mRNA expression in the investigated rat brain structures of citalopram-treated rats. Thus, it seems that up-regulation of alpha1A-AR subtype is characteristic only of those antidepressant agents in which a noradrenergic component is involved in their pharmacological mechanism of action. PMID- 15662098 TI - Influence of antidepressant drugs on macrophage cytotoxic activity in rats. AB - The aim of the study was to evaluate the in vivo and in vitro effects of antidepressant drugs on cytotoxic activity of rat spleen macrophages. In the in vivo experiment, rats were injected subcutaneously with two different doses (2 or 10 mg/kg) of desipramine, fluvoxamine and fluoxetine. The drugs were given once, for 2, 4 or 8 weeks. In the in vitro experiment, spleen macrophages were cultured with three different concentrations of desipramine (3.75, 0.75, or 0.075 mM), fluvoxamine (3.14, 0.62, or 0.062 mM), and fluoxetine (3.23, 0.64, or 0.064 mM) for 72 h. The cytotoxic activity of macrophages was evaluated by measuring the lysis of ((51)Cr) chromate-labelled P-815 target cells. In the in vivo experiment, a single dose of fluvoxamine (2 and 10 mg/kg) and fluoxetine (10 mg/kg) significantly decreased macrophage cytotoxic activity. Fluvoxamine (2 and 10 mg/kg), fluoxetine (10 mg/kg) and desipramine (10 mg/kg) administrated for 14 days also decreased macrophage cytotoxic activity. Twenty-eight day treatment with desipramine (2 and 10 mg/kg) decreased macrophage cytotoxic activity. Desipramine, fluvoxamine and fluoxetine given for 56 days did not affect macrophage cytotoxic activity. In the in vitro experiment, antidepressant drugs did not affect the cytotoxic activity of macrophages. The results of the study indicate that the effects of antidepressant drugs on macrophage cytotoxic activity depend on the drug type, dose and duration of the treatment. PMID- 15662099 TI - Introduction of a new complex imide system into the structure of LCAPs. The synthesis and a 5-HT1A, 5-HT2A and D2 receptor binding study. AB - A series of 17 long-chain arylpiperazines containing bulky, complex imide systems (5,8-dimethyl-3b,9-epoxy-(3a,4,5,6,7,8,9,9a)-octahydro-1H-benzo[e]isoindole 1,3(2H)-dione or 4,9-diphenyl-4,9-epoxy-3a,4,9,9a-tetra-hydro-1H benzo[f]isoindole-1,3(2H)-dione) was synthesized and evaluated for their affinity for serotonin 5-HT1A, 5-HT2A and dopamine D2 receptors. Most of the new compounds showed moderate activity at 5-HT1A binding sites (Ki = 100-492 nM), and two derivatives were found to have marked affinity for the 5-HT2A receptor subtype. None of the tested compounds displayed appreciable binding to dopamine D2 receptors Structure-activity relationships were discussed in respect to an arylpiperazine fragment, whereas the comparison of different imide terminals enabled determination of the size of a hydrophobic pocket (approximately 300 A3) within the 5-HT1A receptor. PMID- 15662100 TI - Effect of metyrapone supplementation on imipramine therapy in patients with treatment-resistant unipolar depression. AB - The paper describes the effect of metyrapone supplementation on imipramine therapy in patients (with treatment-resistant unipolar depression) who fulfilled DSM IV criteria for major depression. Nine patients were enrolled to the study on the basis of history of their illness and therapy. Following 2 weeks of washout period, the patients were treated with imipramine twice daily (100 mg/day) for 6 weeks, and then metyrapone was introduced (twice daily, 500 mg/day), and administered jointly with imipramine for further 6 weeks. Hamilton Depression Rating Scale (HDRS) and Beck Depression Inventory (BDI) were used to assess efficacy of antidepressant therapy. Imipramine changed neither HDRS nor BDI score after 6 weeks of treatment when compared with baseline (before treatment). Metyrapone supplementation significantly reduced both HDRS and BDI scores after 6 week supplementation. Moreover, pharmacokinetic data indicate that metyrapone did not influence significantly the plasma concentration of imipramine and its metabolite, desipramine in the patients during joint treatment with metyrapone and imipramine, what suggests the lack of pharmacokinetic interaction. This preliminary study is the first demonstration of the benefit of metyrapone supplementation in imipramine therapy of treatment-resistant unipolar depression and suggests that a change in the level of neurotransmitters, hormones and immunological parameters, which are disturbed in depression, may contribute to the mechanism of the action of this drug. PMID- 15662101 TI - Effects of chronic treatment with classic and newer antidepressants and neuroleptics on the activity and level of CYP2D in the rat brain. AB - The aim of the present work was to study the effect of chronic treatment with pharmacological doses of selected antidepressants (imipramine, mirtazapine) and neuroleptics (thioridazine, risperidone) on the activity and level of CYP2D in the rat brain. Our previous studies carried out on the liver showed that after chronic treatment with psychotropics, the activity of CYP2D was significantly decreased by imipramine, thioridazine and risperidone, but increased by mirtazapine. Our preliminary results suggest that the same may happen in the brain, where similar tendencies in changes in CYP2D activity were observed. Imipramine, thioridazine and risperidone diminished, while mirtazapine tended to accelerate the rate of ethylmorphine O-deethylation, a specific reaction for measurement of CYP2D activity. In the case of thioridazine, the observed decrease in the enzyme activity was the most pronounced and statistically significant. The level of brain CYP2D4 was not substantially changed by the prolonged administration of the investigated drugs (imipramine 136.3 +/- 14.9%, thioridazine 121.9 +/- 3.5%, risperidone 113.5 +/- 7.8%, mirtazapine 80.3 +/- 1.5% of the control), and did not correspond positively with the measured CYP2D activity. This may imply that the observed changes in the CYP2D activity were not caused by the involvement of those psychotropics in the regulation of CYP2D4. In conclusion, our preliminary results suggest that the effects of prolonged treatment with antidepressants and neuroleptics on the activity of CYP2D found in our previous study in the liver also occur in the brain, which may have an impact on the pharmacological and clinical profile of those drugs. PMID- 15662102 TI - Selective mGlu5 receptor antagonist MTEP attenuates naloxone-induced morphine withdrawal symptoms. AB - Several lines of evidence suggest a crucial involvement of glutamate in the mechanism of drug addiction. The involvement of group I mGlu receptors in the mechanism of addiction has also been proposed. Given the recent discovery of selective and brain penetrable mGlu5 receptor antagonists, the effects of 3-[(2 methyl-1,3-thiazol-4-yl)ethynyl]-pyridine (MTEP) were evaluated in the naloxone precipitated morphine withdrawal model. Experiments were performed on male C57BL/6J (20-25 g) mice. Mice were rendered morphine-dependent and withdrawal was precipitated with naloxone. Two hours and 15 min after the last dose of morphine, mice were injected with a mGlu5 receptor antagonist. MTEP (1-10 mg/kg) in a dose dependent manner inhibited the naloxone-induced symptoms of morphine withdrawal in morphine-dependent mice, remaining without any effect on the locomotor activity of mice. The data suggest that selective mGlu5 receptor antagonists may play a role in the therapy of drug-dependence states. PMID- 15662103 TI - Intraamygdaloid administration of BIBO 3304 increases water intake and extends anxiolytic effects. AB - The present study was designed to evaluate the effects of BIBO 3304 in the Vogel's conflict drinking test and in the water intake test in non-deprived rats after injection of the drug into the basolateral nucleus of the amygdaloid complex. BIBO 3304 was given at the doses of 25, 100 and 200 pmol/0.5 microl/site. We investigated also the effect of 5-hydroxytryptophan (5-HTP), given intraperitoneally at a dose of 20 mg/kg, which was used as a positive control in the water intake test. Water consumption was measured 1, 2, 4, 6 and 24 h after drug administration. We found that water intake was increased both after 5-HTP and BIBO 3304 administration. PMID- 15662104 TI - Influence of beta-adrenergic antagonists on tear secretion in children. AB - The aim of the study was to compare the results of tear film volume, conjunctival and corneal state of children eyes both treated with beta-blocker and healthy ones. We have examined 40 eyes of 20 children at the age from 7 to 17 years. Group I - 20 glaucomatous eyes treated with 0.5% timolol twice daily during at least 12 months. Group II - 20 eyes of control age-matched group. Schirmer I test without anesthesia and lissamine green staining was performed to evaluate conjunctival and corneal surface. The values of Schirmer I test were: group I from 12 to 24 mm, the mean 17.06 +/- 1.78 mm and group II from 16 to 35 mm, the mean 29.3 +/- 2.67, p = 0.000004. Keratoepitheliopathy was observed in 4 eyes (20%) of group I and was rated as 3rd and 6th degree of Franck classification. In the group II, there were only single staining points classified as 1st degree noticed in 2 eyes (10%), p = 0.37. The long-term treatment with timolol causes a decrease in tear secretion in children, which can further generate keratoepiteliopathy. Therefore, we suggest application of the artificial tears in these patients. PMID- 15662105 TI - Release bursts in English word-final voiceless stops produced by native English and Korean adults and children. AB - The aim of this study was to evaluate the acquisition of statistical properties of a second language (L2). Stop consonants are permitted in word-final position in both English and Korean, but they are variably released in English and invariably unreleased in Korean. Native Korean (K) adults and children living in North America and age-matched native English (E) speakers repeated English words ending in released tokens of /t/ and /k/ at two times separated by 1.2 years. The judgments of E-speaking listeners were used to determine if the stimuli were repeated with audible release bursts. Experiments 1 and 2 revealed fewer final releases for K than E adults, and fewer releases for /t/ (but not /k/) for K than E children. Nearly all /t/ and /k/ tokens were heard as intended in experiment 3, which evaluated intelligibility. However, the K adults' /k/ tokens were identified with less certainty than the E adults'. Taken together, the results suggested that noncontrastive (i.e., statistical) properties of an L2 can be learned by children, and to a somewhat lesser extent by adults. PMID- 15662106 TI - [The perception of the second language production]. AB - Non-native speakers of a second language often report that the speech rate in that language is faster than the rate in their own language. So as to compare speech rate perception by native (French) and non-native (Swiss German) speakers, and to determine if rate estimation by non-native speakers is correlated with their level of comprehension, we asked two groups of 96 participants, native and non-native speakers of French, to listen to short stories read at slow, medium and fast rates. They were asked to answer a few comprehension questions and to give an estimate of the speech rate. The results obtained show that there is indeed a difference between the two groups: the faster the physical speech rate, the greater the impression of speed in the non-native speakers as compared with the native speakers. In addition, when speech rate is slow and normal, there is a significant negative correlation between oral comprehension and perceived rate: the lower the comprehension, the higher the estimated rate. PMID- 15662107 TI - Underlying voicing in Majorcan Catalan word-final stop-liquid clusters. AB - This study investigates the extent to which phonetic voicing is maintained in word-final clusters composed of an underlying voiced stop followed by nonsyllabic /l/ or /r/ in Majorcan Catalan. Electropalatographic and acoustic data for five speakers of this Catalan dialect reveal that, in agreement with the non-syllabic status of the liquid, voicing for /l/ is only available if occurring during the preceding stop. The rhotic is always phonetically voiceless. Speakers differ regarding the extent to which they keep the underlying stop voicing distinction and the production strategies they use for that purpose. This distinction is highly robust and distributed over the entire syllable in nasal-stop-/l/ clusters for some speakers, but much less clear or absent for those speakers who devoice /l/ as a general rule. Underlying stop voicing is cued primarily by stop closure duration and vocal fold vibration, or else by closure duration rather than by voicing. It may be concluded that the word-final devoicing process operating in Catalan does not apply to Majorcan Catalan tautosyllabic stop clusters with a liquid, and that phonetic voicing may affect just the stop or both the stop and the liquid. The implications of these findings for sound change are discussed. PMID- 15662108 TI - Final consonants and glottalization: new perspectives from Hanoi Vietnamese. AB - The evolution from final obstruents to final glottal stop and then to rhyme glottalization (i.e. from /at/ to /a?/, then to /a'/) is a well-established general trend in the history of the Sino-Tibetan language family and beyond. It has further been shown by laryngoscopy that in three languages which retain the nonreleased syllable-final obstruents /p/, /t/ and /k/ (Standard Thai, and two Chinese dialects), these obstruents are often accompanied by a glottal stop. The present research raises the issue whether there is another typological possibility: can nonreleased final obstruents be accompanied consistently by modal phonation, without glottal stop? Analysis of electroglottographic recordings of 126 syllables in two carrier sentences spoken by 4 speakers shows that, in Hanoi Vietnamese, the final obstruents /p/, /t/ and /k/ are not accompanied by glottalization, and that the open quotient increases in the course of the syllable rhyme. Obstruent-final rhymes (which may carry either of two tones: D1 or D2) are compared with nasal-final rhymes which, under one of the tones (tone B2), are confirmed to be glottalized. Our finding is that tones D1 and D2 (i.e. obstruent-final rhymes) are both produced in modal voice, which shows that the typological paradigm of observed realizations of syllable-final obstruents must be enlarged. The discussion puts forward the hypothesis that the unusual association of segments and voice quality found in Hanoi Vietnamese is a strategy to maintain the opposition between B2-tone and D2-tone rhymes. PMID- 15662109 TI - Voice register in Suai (Kuai): an analysis of perceptual and acoustic data. AB - Analyses of the perceptual and acoustic characteristics of the Register 1 ('clear') versus Register 2 ('breathy') distinction have been carried out on the Kuai dialect of Suai, a Mon-Khmer language. The perception results were obtained from five-parameter synthesized stimuli. They showed that the primary parameter underlying the distinction is the frequency of onset of laryngeal excitation (F0). One other parameter making a significant contribution was the open quotient. The F0 result was confirmed by an acoustic analysis of eight pairs of natural utterances produced by native speakers. We conclude that the Suai language is in a state of flux with respect to the voice registers, although the distinction has not disappeared. The perceptual data reveal mixed levels of sensitivity, and the production data indicate that some speakers maintain a fairly good distinction, while others do not. The language seems to be replacing the register distinction with a prosodic one of pitch accent, possibly as a stage leading to tonogenesis. PMID- 15662110 TI - Effective treatment of indolent non-hodgkin's lymphomas with mitoxantrone, chlorambucil and prednisone. AB - Since indolent non-Hodgkin's lymphomas (NHL) represent about 35% of all malignant lymphomas and mainly affect elderly patients, availability of a conventional chemotherapy regimen with high efficacy and low toxicity is of clinical importance. PATIENTS AND METHODS: We retrospectively analysed 13 patients with advanced indolent NHL who were treated with 6-9 cycles of MCP: mitoxantrone 8 mg/m2 (days 1 and 2), chlorambucil 3 x 3 mg/m2 (days 1-5) and prednisone 25 mg (days 1-5) every 4 weeks. RESULTS: The overall response was 84% (61% complete response, 23% partial response), 1 patient had stable disease and 1 patient experienced progressive disease. Median time to progression was 37 months (95% CI: 20-53) and the median survival has not yet been reached. The main toxicity (66%) was neutropenia (WHO grade III). There was no hair loss and no cardial or neurologic adverse event. CONCLUSION: In summary, MCP is an effective and well tolerated chemotherapy regimen and is probably an alternative to the more toxic CHOP regimen, especially in older patients. PMID- 15662111 TI - Neoadjuvant chemotherapy with cyclophosphamide, mitoxantrone, and 5-fluorouracil in locally advanced breast cancer. AB - OBJECTIVES: Our primary objective was to determine the response rate; secondary objectives were to assess the toxicity rate, and disease-free and overall survival rates in patients with locally advanced breast cancer (LABC) receiving a cyclophosphamide (500 mg/m2), mitoxantrone (12 mg/m2) and 5-fluorouracil (500 mg/m2) (CMF) chemotherapy regimen. PATIENTS AND METHODS: The data from 74 patients with LABC with neoadjuvant CMF chemotherapy were analyzed retrospectively. Preoperatively, all patients received 3 cycles of CMF on day 1, repeated every 21 days. In 3 (4.1%) patients, breast-conserving surgery was given and in 71 (95.9%) modified radical mastectomy. All patients received radiotherapy and 3 additional cycles of CMF chemotherapy after surgery. RESULTS: Median age of the patients was 47 years (range: 17-74). 43 patients were premenopausal, whereas 31 were postmenopausal. 54 patients were in stage IIIA, and 20 were in stage IIIB. The overall clinical response rate was 88%; 11 (14.9%) had a complete response, 54 (73%) had a partial response, and 2 (2.8%) had progression. 14 (18.9%) had a pathological complete response. The median follow-up was 62 months. The median disease-free survival was 64.9 months, and the median overall survival was 97.5 months. The 5-year disease-free and overall survival rates were 52% and 79.9%, respectively. Most frequent side-effects were nausea/vomiting, mucositis, alopecia and leukopenia. CONCLUSION: The CMF regimen has a high overall response rate and an acceptable side effect profile in the treatment of locally advanced breast cancer. Further studies are needed to evaluate its effectiveness in breast conserving strategies. PMID- 15662113 TI - No one-way street: cross-talk between e-cadherin and receptor tyrosine kinase (RTK) signaling: a mechanism to regulate RTK activity. AB - E-cadherin was originally viewed exclusively as a structural protein mediating cell-cell adhesion. More recently, its signaling functions have been recognized. Loss or downregulation of E-cadherin releases proteins, such as b-catenin and p120 catenin, from a membrane-bound state into the cytoplasm, which are known to regulate transcriptional activity. E-cadherin is known to interact with receptor tyrosine kinases, such as epidermal growth factor receptor (EGFR). However, previously, only the regulation of E-cadherin mediated adhesion through EGFR has been described and activation of EGFR was implicated in loss of cell adhesion, and increased cell migration and invasion. Now, Qian et al. (EMBO J 2004, 23:1739 48) describe that E-cadherin mediated adhesion inhibits receptor tyrosine kinase (RTK) activity. E-cadherin was found to interact through its extracellular domain with EGFR and other receptor tyrosine kinases, thereby decreasing receptor mobility and ligand-affinity. This is a novel mechanism by which E-cadherin inhibits RTKs, and suggests that downregulation of E-cadherin may contribute to the frequently observed activation of RTKs in tumors. PMID- 15662112 TI - Antiemetic efficacy of an oral suspension of granisetron plus dexamethasone and influence of quality of life on risk for nausea and vomiting. AB - OBJECTIVES: To assess the antiemetic efficacy of an oral suspension of granisetron/dexamethasone in patients receiving chemotherapy and to determine whether quality-of-life parameters influence the risk for postchemotherapy nausea and vomiting (PCNV). PATIENTS AND METHODS: In an open monocentric study, an oral suspension containing 2 mg granisetron and 16 mg (4 mg for moderately emetogenic chemotherapy) dexamethasone was administered to 43 chemotherapy-naive patients before highly (n = 16) or moderately (n = 27) emetogenic chemotherapy and on the 3 subsequent days (2 for moderately emetogenic chemotherapy). Emetic episodes were recorded and quality of life was assessed prior to each cycle with a questionnaire based on EORTC QLQ-30. RESULTS: In the group undergoing highly (moderately) emetogenic chemotherapy, complete control of acute vomiting was achieved in 60-72.7% (92.6-95.0%), and complete control of delayed vomiting in 37.5-40.0% (75.0-92.2%), of patients within the first 3 (5) cycles. The following quality-of-life parameters were significantly associated with PCNV: tiredness (RR = 1.3, p < 0.05), pain (RR = 1.5), impairment of daily life by pain (RR = 1.7), sensation of abdominal pressure and fullness (RR = 2.5), impairment of social activities (RR = 2.9). CONCLUSIONS: Once-daily oral administration of a suspension of granisetron/dexamethasone is an active prophylaxis of nausea and vomiting and compares favorably with data reported on intravenous administration. Quality-of-life parameters assessed pre-treatment could help to identify patients at high risk for nausea and vomiting so that antiemetic therapy can be tailored to individual patient risk. PMID- 15662114 TI - Antitumor effect of valorphin in vitro and in vivo: combined action with cytostatic drugs. AB - The action of the cytostatic drugs (epirubicin and vincristine) in combination with the endogenous antiproliferative beta-hemoglobin fragment (33-39), valorphin, was studied in tumor (L929 and A549) cell cultures, primary culture of murine bone marrow cells and in murine model of breast carcinoma in vivo. Simultaneous application of 1 microM valorphin and 1 microM epirubicin, in vitro, did not result in an additive suppressive effect on cell culture growth. Additive effects were achieved with alternating applications of the peptide and the drugs, namely, 0.5 microM (but not 1 microM) epirubicin added 24 h prior to 1 microM valorphin; 1 microM valorphin added 48 h prior to 0.1 microM epirubicin, or 0.1 microM vincristine, or 0.05 microM vincristine, which resulted in 100% cell death in the both series with vincristine and up to 78% cell biomass reduction in the experiments with epirubicin. In the in vivo model (female BLRB mice with subcutaneously inoculated syngeneic mammary carcinoma), simultaneous treatment with 25 mg/m(2) epirubicin and 1 mg/kg valorphin resulted in 42% of tumor growth inhibition, as compared with the negative control group and 22% inhibition as compared with the epirubcin-treated group (at 20th day of treatment). Survival was significantly improved (69% compared to 39% in the group treated with epirubicin only) at day 26 after the treatment beginning. PMID- 15662115 TI - Effects of HIV-1 Nef, a cytotoxic viral protein, on the growth of primary colorectal cancer. AB - Colorectal cancer is the third leading cause of cancer deaths in American men and women. We describe the cytotoxic use of HIV-1 Nef protein and a cytotoxic peptide identified within the HIV-1 Nef structure in targeting human cancer cells in vitro and in vivo in a human xenograft model. A human colorectal tumor was implanted and propagated in the subcutaneous tissue of SCID mice. The mice were injected biweekly with the Nef apoptotic peptide. The tumor treated with Nef peptide underwent significant growth inhibition by as much as 300 percent when compared to the control (untreated) tumors. The Nef peptides were found to have an apoptotic effect on the human colon tumor similar to the effect seen on CD4 cells when the viral protein is secreted by the HIV-1 virus infected cells. The evidence from the xenograft mouse model suggests that the Nef peptides can be used to inhibit human colorectal cancer growth. PMID- 15662116 TI - Opposing roles of E2Fs in cell proliferation and death. AB - Progression through the cell cycle is dependent upon the temporal and spatial regulation of the various members of the E2F family of transcription factors. Two of these members, E2F1 and E2F4 have opposing roles in cell cycle progression, which were defined over a decade ago. While E2F1 is an activator of cell cycle progression, E2F4 functions as a transcriptional repressor. Recent data indicate that these transcription factors also play a role in the cellular response to DNA damage. In the case of E2F1, its overexpression leads to apoptosis. In contrast, the decreased expression of E2F4, in response to siRNA-mediated knockdown or to certain therapeutic agents, induces apoptosis. Conversely, increased levels of E2F4 may confer resistance to apoptosis-inducing therapies used in the clinic. The balance between the activities of these two proteins in tumor cells is of great interest. Directed control of E2F1 and E2F4 action may lead to better diagnosis of disease and improved therapeutic modalities. PMID- 15662117 TI - Defining the extent of cables loss in endometrial cancer subtypes and its effectiveness as an inhibitor of cell proliferation in malignant endometrial cells in vitro and in vivo. AB - Loss of Cables expression is associated with a high incidence of endometrial hyperplasia and endometrial adenocarcinoma in humans. The Cables mutant mouse develops endometrial hyperplasia and following exposure to chronic estrogen develops early endometrial adenocarcinoma. The objectives of the current study were to determine if: (1) loss of Cables expression occurred in high grade endometrioid adenocarcinoma, uterine serous and clear cell carcinoma as observed in endometrial hyperplasia and low grade endometrial adenocarcinoma; (2) overexpression of Cables inhibited cell proliferation in endometrial cancer (EC) cells in vitro and in vivo; and (3) progesterone could regulate the expression of Cables mRNA. Hyperplastic endometrium and low and high grade endometrioid adenocarcinoma showed loss of Cables expression when compared to benign control secretory endometrium. Loss of Cables expression in serous and clear cell tumors was similar to that observed in endometrioid adenocarcinomas with greater than 80% showing loss of protein expression. Treatment of EC lines with progesterone increased cables expression in low-grade EC whereas it had no effect on cables expression in cells derived from high-grade EC. The progesterone-induced increase in cables was abrogated in the presence of a progesterone receptor (PR) antagonist, suggesting the PR mediates the increase. Cables overexpression inhibited cell proliferation of well differentiated EC cells and had no effect on the poorly differentiated EC cells. The capacity to form tumors was dramatically reduced in the Cables overexpressing cell lines compared to those cells containing the control vector. Collectively these results suggest that Cables is an important regulator of cell proliferation and loss of Cables expression contributes to the development of all types of EC. PMID- 15662118 TI - The SERCA pump as a therapeutic target: making a "smart bomb" for prostate cancer. AB - Prostate cancer is uniformly fatal once it has spread outside of the prostate gland. Prostate cancers have a remarkably low proliferative rate, which may in part explain their relative unresponsiveness to conventional antiproliferative chemotherapy. New therapies for prostate cancer that activate proliferation independent cell death are therefore needed. The endoplasmic reticulum (ER) has emerged as an organelle that plays a major role in cell signaling pathways, cellular response to stress and cellular activation of apoptosis. In this review, the SERCA pump is identified as an ER protein whose normal function is required by all cells and represents a potential therapeutic target for cancer therapy. Sustained SERCA inhibition by agents such as thapsigargin results in activation of ER-stress response and simultaneous activation of apoptotic pathways within the ER and the mitochondria. Due to the SERCA pump's critical role in normal cellular metabolism, agents like thapsigargin directed toward inhibiting SERCA function would likely produce significant toxicity to normal cells and, therefore, must be selectively targeted to cancer sites. The cytotoxicity of thapsigargin can be attenuated, however by coupling to a targeting peptide to produce an inactive prodrug that is only activated by prostate cancer specific proteases such as the serine protease prostate-specific antigen (PSA). PSA activated thapsigargin prodrugs have been characterized that are selectively toxic to PSA-producing prostate cancer cells in vitro and in vivo. These prodrugs are currently undergoing preclinical evaluation as potential targeted therapy for prostate cancer. PMID- 15662119 TI - Unpulsed dendritic cells induce broadly applicable anti-tumor immunity in mice. AB - The discovery of dendritic cells (DC) as professional antigen presenting cells has opened up new possibilities for their use in the development of cancer vaccines. Here we show that unpulsed BM-derived CD11c+CD8alpha-DC administered s.c. to mice enhanced their resistance to subsequent lethal tumor challenges. The tumor resistance-inducing activity of unpulsed DC was dependent on their maturation status, involved CD4+ and CD8+ T cell activities, and was abrogated by pulsing with an irrelevant class I MHC-restricted peptide. Although the BALB/c Meth A tumor system was employed extensively to demonstrate the tumor resistance inducing activity of unpulsed DC, the immunogenicity of this vaccine was evident in other inbred strains of mice against a variety of syngeneic tumors. The broad anti-tumor responses induced by unpulsed DC appears to be due to their capacity to present "self" tumor-associated antigens, such as the H2-Kd-restricted, wild type sequence p53(232-240) peptide. These findings highlight the potential broad applicability of unpulsed DC as a tumor vaccine. PMID- 15662120 TI - DNA ploidy and S phase fraction of breast and ovarian tumor cells treated with a natural anthracycline analog (aloin). AB - DNA ploidy and S phase fraction analysis by flow cytometry on breast and ovarian tumor cells continuously exposed to aloin, a natural anthraquinone, at two concentrations (20-60 microg/ml) was done. Untreated breast and ovarian tumor cells (control) showed an aneuploid pattern, with a mean DNA index of 2.10+/-0.10 and S phase fraction of 28.46+/-1.5 and 17.40+/-0.75%, respectively. Treatment of breast and ovarian tumor cells with aloin showed a persistent aneuploid pattern and a significantly dose-dependent increase in the percentage of S phase fraction and in the proportion of cells cycling at a higher ploidy level (>G2M). The polyploidization indicates that aloin does not inhibit initiation of DNA synthesis and that cells replicated a full complement of DNA but had difficulty in M phase. PMID- 15662121 TI - Cell cycle regulation and hepatocarcinogenesis. AB - Hepatocellular carcinoma (HCC) develops on a background of chronic hepatitis or cirrhosis. The slow progression of this disease has facilitated the identification of discrete pathologic stages. Increased rates of hepatocyte proliferation in preneoplastic nodules is an early event in the progression of HCC. Increased cell turnover results in the selection of monoclonal hepatocyte populations that subsequently undergo genomic alterations that lead to the development of HCC. The heterogeneous nature of genomic alterations identified in tumors from patients with HCC has impeded the identification of regulatory pathways whose disruption are critical for tumor initiation. However, several regulatory networks important for liver cell proliferation have been characterized using the partial hepatectomy model, an in vivo model of liver cell cycle progression, and are likely relevant to the pathogenesis of hepatocellular carcinoma. PMID- 15662122 TI - ZNRD1 gene suppresses cell proliferation through cell cycle arrest in G1 phase. AB - ZNRD1, a transcription-associated gene, was recently found in our laboratory significantly suppress the cell proliferation of stomach cancer cells in vitro and in vivo. In this study, we firstly characterized ZNRD1 expression in a wide spectrum of gastric diseases by immunohistochemistry and RT-PCR. We also investigated its antiproliferative effects and associated molecular alterations in human gastric cancer cell line AGS and mouse fibroblast cell line NIH3T3. Anti ZNRD1 monoclonal antibody H6 was found to react with 38 (63%) of normal gastric tissues, and 51 (81%) of gastritis. In contrast, no positive expression was found in gastric adenocarcinomas. Thus, the expression of ZNRD1 in normal gastric tissues was significantly higher than that in gastric adenocarcinomas. Compared with the control clones, ZNRD1-transfected cells exhibited significant inhibition of cell growth with G1 cell cycle arrest mediated by the suppression of cyclin D1 expression. These results showed that ZNRD1 may play an important role in the regulation of gastric carcinogenesis and could be used as a new target in treatment of stomach cancer. PMID- 15662123 TI - The relationships between p53 protein expression and the clinicopathological features in the uveal melanomas. AB - BACKGROUND: p53 alterations are common events in cancer and may have some prognostic ramifications. OBJECTIVES: To investigate the uveal melanoma for: (1) the p53 expression patterns, (2) the relationships among p53 expression and clinicopathological features of these lesions. MATERIALS AND METHODS: Using Immunoperoxidase-staining methods and mouse monoclonal antibodies, the expression of p53 protein in 60 uveal melanomas was examined. RESULTS: Based on p53 immunoreactivity, the lesions were divided into two groups: the first group included 9 tumors (15%) and had p53 protein overexpression. The second group included 51 (85%) tumors and was lacking p53 expression. p53 protein overexpression was significantly associated with extraocular extension and vascular invasion (p values, 0.01 and 0.04). No significant correlation was found between p53 protein expression and the clinicopathologic features of these lesions. CONCLUSIONS: p53 protein overexpression occurs in the uveal melanoma and may be associated with some unfavorable histological features (invasion). PMID- 15662124 TI - Homozygous deletion of the MTAP gene in invasive adenocarcinoma of the pancreas and in periampullary cancer: a potential new target for therapy. AB - Methylthioadenosine phosphorylase (MTAP) plays an important role in the salvage pathway for the synthesis of adenosine. Novel chemotherapeutic strategies exploiting the selective loss of MTAP function in cancers have been proposed. The MTAP gene, on chromosome 9p21, is frequently included within homozygous deletions of the p16INK4A/ CDKN2A gene. Biallelic deletions of the p16INK4A/CDKN2A gene are found in 40% of pancreatic cancers, suggesting that the MTAP gene may be frequently inactivated in pancreatic cancer and that selected patients with pancreatic cancer may benefit from therapies targeting this loss. We immunolabeled six xenografted pancreatic cancers with known MTAP and p16INK4A/CDKN2A gene status and found that immunolabeling mirrored gene status. Loss of expression of both MTAP and p16 was observed only in those pancreatic cancers with homozygous deletions that encompassed both the MTAP and p16INK4A/CDKN2A genes. We then immunolabeled a series of 320 microarrayed infiltrating pancreatic adenocarcinomas, 35 biliary adenocarcinomas, 54 ampullary cancers, and 35 noninvasive intraductal papillary mucinous neoplasms. Immunolabeling for MTAP was lost in 91 of the 300 (30%) evaluable pancreatic cancers, 9 of 54 (17%) ampullary cancers, 4 of 33 (12%) biliary cancers, and in 1 of 35 (3%) IPMNs. All neoplasms with loss of MTAP labeling also demonstrated loss of p16 labeling. These results suggest that MTAP expression is lost in approximately 30% of infiltrating pancreatic cancers and in a lower percentage of other periampullary neoplasms, that this loss is the result of homozygous deletions encompassing both the MTAP and p16INK4A/CDKN2A genes. Thus, pancreatic cancer is a promising cancer type in which to explore novel chemotherapeutic strategies to exploit the selective loss of MTAP function. PMID- 15662125 TI - Mitochondrial mutations and nasopharyngeal carcinoma. PMID- 15662126 TI - Epigenetic regulation of protein phosphatase 2A (PP2A), lymphotactin (XCL1) and estrogen receptor alpha (ER) expression in human breast cancer cells. AB - Absence of the estrogen receptor alpha (ER) in human breast cancer cells is an indicator of poor prognosis, and predictive of lack of response to hormonal therapy. Previous studies in our laboratory and others have shown that epigenetic regulation, including DNA methylation and histone deacetylation, are common mechanisms leading to ER gene silencing. Through the use of pharmacologic inhibitors, 5-aza 2'deoxycytidine (AZA) and Trichostatin A (TSA), we have shown that alterations in both of these mechanisms results in synergistic reexpression of ER mRNA and functional protein. These alterations may play a larger role in stimulation of cell signaling pathways leading to ER expression. We have utilized newly developed genome wide screening microarray techniques to identify gene(s) contributing to the hormone independent phenotype and AZA/TSA mediated ER expression. From this screen, we identified and confirmed expression of 4 candidate genes (PP2A, XCL1, THY1 and NBC4) as potential regulators of the hormone independent phenotype. Expression of two genes, XCL1 and PP2A, appeared to be correlated with ER expression. PP2A expression was not changed with ER degradation using ICI 182,780 whereas XCL1 expression decreased in the presence of AZA/TSA and ICI 182,780. This suggests that PP2A may be a determinant of ER expression while XCL1 appears to be ER responsive and downstream of ER expression. These gene products may be novel targets to be further explored in the development of new therapeutics for ER negative breast cancer. PMID- 15662127 TI - Genotyping and functional analysis of the D104N variant of human endostatin. AB - Endostatin is an endogenous inhibitor of angiogenesis derived from the extracellular matrix protein collagen XVIII. It has been reported that a variation at the 104 position (D104N) of human endostatin is associated with an increased risk of prostate cancer, potentially indicating that this protein variant is less active as an anti-angiogenic agent. Herein we reported the results of genotyping 389 patients with androgen independent prostate cancer (AIPC) and 352 normal control individuals for D104N endostatin. There was no significant association between the frequency of 104N endostatin and the incidence of AIPC in either Caucasian or African American patients compared to controls (15% Caucasian AIPC versus 13.7% in Caucasian controls, p=0.79; 7.4% African American AIPC versus 5.6% in African American controls, p=0.64). Actuarial analysis revealed no statistically significant association between incidence of the DN heterozygous genotype and survival (p=0.62 by logrank test). To study the functional significance of the D104N conversion, we have expressed and purified insoluble recombinant human 104D and 104N endostatin and compared their respective activities in human umbilical vein endothelial cell (HUVEC) tube formation assays. The 104N variant of human endostatin inhibited HUVEC tube formation at least as well as the wild-type form. We concluded that the D104N variation in human endostatin is neither clinically relevant nor suitable as a pharmacogenomic endpoint to assess the risk for developing AIPC. PMID- 15662128 TI - The proteasome inhibitor bortezomib (PS-341) inhibits growth and induces apoptosis in primary effusion lymphoma cells. AB - The ubiquitin-proteasome pathway is responsible for degrading many critical regulatory proteins involved in immune and inflammatory responses, control of cell growth and apoptosis. Recently, proteasome inhibitors have emerged as promising new therapeutic agents in hematological malignancies. Here we show that Bortezomib (PS-341), a proteasome-inhibitor, inhibits cellular proliferation and induces apoptosis in cell lines derived from Primary Effusion Lymphoma (PEL), a subtype of non-Hodgkin's lymphoma associated with infection by human herpes virus 8 (HHV-8). Bortezomib demonstrated more cytotoxicity against PEL cells than against cell lines derived from multiple myeloma, a disease for which is in current clinical use. Apoptosis induced by Bortezomib was associated with inhibition of the classical and alternative NF-kappaB pathways, upregulation of p53, p21 and p27 and activation of caspase cascade. Finally, treatment of PEL cells with Bortezomib exerted a synergistic or additive cytotoxic effect in combination with chemotherapeutic drugs or TRAIL. Taken together, these findings suggest that Bortezomib represents a promising agent for the treatment of PEL. PMID- 15662129 TI - Inhibition of gastric cancer angiogenesis by vector-based RNA interference for Raf-1. AB - Tumor angiogenesis plays an important role in the malignancy of solid tumors. A number of recent studies including our own have suggested that Raf-1 is involved in this process, and may be critical in regulating gene activation of several angiogenesis factors. To further define Raf-1 function in angiogenesis and to explore novel approaches to modulate angiogenesis, we employed the small interference RNA technique to knock-down gene expression of Raf-1 in gastric cancer cell line SGC7901 that expresses a high level of Raf-1. The protein level of Raf-1 in the SGC7901cells was decreased dramatically after transfection with a Raf-1 specific siRNA vector. Further study proved that, VEGF and HIF-1alpha, two angiogenesis promoting factors, were found to be downregulated. And we also find that Vector-based RNA interference for Raf-1 increases transfected gastric cell apoptosis and inhibits cellular proliferation. Our results suggest that Raf-1 may be involved in angiogenesis by controlling the expression of angiogenesis-related factors and provide a possible strategy for the treatment of tumor angiogenesis by targeting Raf-1. PMID- 15662130 TI - Upregulation of endogenous angiogenesis inhibitors: a mechanism of action of metronomic chemotherapy. AB - Antiangiogenic or metronomic chemotherapy, the frequent administration of conventional cytotoxic agents at low doses, is believed to target activated tumor endothelial cells. The mechanisms of action of such regimen remain poorly understood. In the March 2004 issue of Cancer Research, Hamano et al. demonstrated that low-dose cyclophosphamide inhibits tumor growth by upregulating the endogenous angiogenesis inhibitor thrombospondin-1 in tumor and perivascular cells. Thrombospondin-1, in turn, promotes endothelial cell apoptosis. It was also proposed that thrombospondin-1 levels might be used as a surrogate marker to monitor response to low-dose cyclophosphamide therapy in the clinic. PMID- 15662131 TI - Efficacy of beta-lapachone in pancreatic cancer treatment: exploiting the novel, therapeutic target NQO1. AB - NAD(P)H:quinone oxidoreductase (NQO1) is elevated in human pancreatic cancers. We hypothesized that beta-lapachone, a novel 1,2-naphthoquinone with potential antitumor activity in cancer cells expressing elevated levels of NQO1, would induce cytotoxicity in pancreatic cancer cells, wherein this two-electron reductase was recently found elevated. beta-lapachone decreased clonogenic cell survival, metabolic cell viability, and anchorage- independent growth in soft agar. The cytotoxic in vitro effects of beta-lapachone were inhibited with coadministration of dicumarol, a specific inhibitor of NQO1. In preestablished human pancreatic tumor xenografts in nude mice, beta-lapachone demonstrated greater tumor growth inhibition when given intratumorally compared to when complexed with cyclodextrin to increase its bioavailability. Due to the poor prognosis of patients with pancreatic cancer and the limited effectiveness of surgery, chemotherapy, and radiation therapy, treatment regimens based on sound, tumor-specific rationales are desperately need for this disease. PMID- 15662132 TI - Dystroglycan: to adhere or not adhere during cancer progression. PMID- 15662133 TI - The chemokine receptor CXCR4 is regulated by DNA methylation in pancreatic cancer. AB - Despite the biological and clinical importance of the interaction between the chemokine receptor CXCR4 and its ligand CXCL12 (SDF-1alpha) in human cancers, little is known about transcriptional regulation of the CXCR4 gene. Although aberrant hypermethylation in cancer has been described typically in genes with tumor-suppressor properties, this epigenetic alteration has also been observed to affect potential cancer-promoting genes. We now demonstrate that DNA methylation influences CXCR4 expression in human pancreatic cancer. Gene expression profiling and reverse transcription-PCR identified a significant proportion of pancreatic cancer cell lines that displayed little or no CXCR4 mRNA expression. Using methylation-specific PCR, combined bisulfite restriction analysis, and bisulfite sequencing, we found the 5' CpG islands of the CXCR4 gene to be unmethylated in normal pancreas, whereas promoter hypermethylation was detected in 45% (9 of 20) of pancreatic cancer cell lines and in 46% (46 of 100) of primary pancreatic adenocarcinomas. There was a significant inverse correlation between methylation and mRNA expression level of CXCR4 (P=0.008) in a large panel of pancreatic cancer cell lines. Constitutive as well as inducible expression of CXCR4 could be restored in methylated cell lines pharmacologically using epigenetic modifying drugs. These findings demonstrate the first evidence for epigenetic regulation of CXCR4 in human cancers, providing new insights into the role of CXCR4/CXCL12 interactions in tumor progression. PMID- 15662134 TI - Paraneoplastic leukemoid reaction and rapid progression in a patient with malignant melanoma: establishment of KT293, a novel G-CSF-secreting melanoma cell line. AB - Paraneoplastic leukemoid reaction (PLR) is a rare condition of leucocytosis in cancer patients. Here we report the rapid progression of a patient suffering from a metastasized malignant melanoma and PLR. The patient's white blood cell count exceeded 200,000 cells per mul and the serum level of Granulocyte Colony Stimulating Factor (G-CSF) was elevated up to 780 pg/mul. A Tc-m99-labeled anti NCA90/95 based granulocyte scan demonstrated reactive bone marrow expansion, splenomegaly and granulocyte infiltration into the tumor. KT293, a S100, gp100 and CD68 positive melanoma cell line derived from an axillary metastasis, produced large amounts of G-CSF in vitro and induced rapidly growing tumors and PLR after subcutaneous inoculation in SCID mice. In contrast to G-CSF-secreting cancer cells of other tissue origin, G-CSF-neutralizing antibodies failed to inhibit the growth of KT293 cells. In addition, KT293 cells did not express G-CSF receptor. These observations suggest that paracrine effects of G-CSF-secretion and PLR might promote an aggressive melanoma phenotype, as seen in this patient. PMID- 15662136 TI - Dual role of p53 in DNA binding. PMID- 15662135 TI - Altered gene expression in phenotypically normal renal cells from carriers of tumor suppressor gene mutations. AB - BACKGROUND: The inherently complex signaling networks of tumors result from genetic and epigenetic alterations that occur during cancer initiation and progression. METHODS: In an attempt to identify early molecular changes associated with dominantly inherited predisposition to "two-hit" renal tumors, the expression profiles of primary cultures of phenotypically normal renal epithelial cells from individuals bearing a germline mutation in either the von Hippel-Lindau (VHL) or the tuberous sclerosis complex (TSC) gene were compared to that of renal epithelial cells from control nonmutation carriers by microarray analysis. RESULTS: Reliability of the microarray data from pooled samples was confirmed by real-time RT-PCR. Principal Component Analysis revealed substantial differences in the gene expression profiles of the renal epithelial cells from VHL and TSC mutation carriers. In several instances, the microarray data confirm our present knowledge of the cellular pathways affected by biallelic VHL and TSC mutations. CONCLUSIONS: These findings demonstrate that heterozygosity for a mutant tumor suppressor gene may alter the expression profiles of phenotypically normal epithelial cells in a gene-specific manner. Detectable effects of "one hit" represent early molecular changes in tumorigenesis that may serve as targets for chemopreventive intervention. PMID- 15662137 TI - RNA interference: a potential strategy for isoform-specific phosphatidylinositol 3-kinase targeted therapy in ovarian cancer. AB - Phosphatidylinositol 3-kinase (PI3-kinase) is a novel intracellular transducer involved in a wide range of cancer-associated signaling pathways, which comprises various isoforms and splice variants with distinct biologic activities and clinical implications. Especially, the class Ia PI3-kinase 110 kD catalytic subunit alpha (PIK3CA) is the most important isoform in tumorigenesis and possibly, tumor angiogenesis. Several strategies have been developed to block PI3 kinase for cancer therapy; however, the approach to target specific PI3-kinase isoform has not been explored to date. In the present study, we show that RNA interference (RNAi) through small interfering (siRNA) sequences targeting PIK3CA has potential applications in isoform-specific "knock-down" of PI3-kinase. This strategy provides a novel tool to study the function of various PI3-kinase isoforms and may contribute to isoform-specific targeting of PI3-kinase in human cancer. PMID- 15662138 TI - The tumor suppressor p33ING1b enhances taxol-induced apoptosis by p53-dependent pathway in human osteosarcoma U2OS cells. AB - p33ING1b can stimulate cell cycle arrest, DNA repair, apoptosis and chemosensitivity. The actions of p33ING1b involve p53-dependent and p53 independent mechanisms. To investigate if the p33ING1b isoform is involved in the chemosensitivity of osteosarcoma cells, p33ING1b was overexpressed in p53+/+ U2OS cells or p53-mutant MG63 cells, and then cell growth arrest and apoptosis were assessed after treatment with taxol. The results showed that p33ING1b markedly increased taxol-induced growth inhibition and apoptosis in p53+/+ U2OS cells, but not in p53-mutant MG63 cells. Moreover, ectopic expression of p33ING1b could obviously upregulate p53, p21WAF1 and bax protein levels and activate caspase-3 in taxol-treated U2OS cells. Taken together, our data demonstrate that p33ING1b enhances taxol-induced apoptosis through p53-dependent pathway in human osteosarcoma cells. p33ING1b may be an important marker and/or therapeutic target in the prevention and treatment of osteosarcoma. PMID- 15662140 TI - The use of two neuroleptics to treat bipolar illness in two children. PMID- 15662141 TI - Aripiprazole in the treatment of bipolar disorder in children and adolescents. PMID- 15662144 TI - Psychosurgery for self-injurious behavior in Tourette's disorder. AB - One of the most serious and difficult-to-treat conditions in child and adolescent psychiatry is self-injurious behavior (SIB). SIB can be associated with a number of psychiatric disorders, including mental retardation, schizophrenia, borderline personality disorder, pervasive developmental disorders, stereotypic movement disorder, and Tourette's Disorder. A variety of neurosurgical procedures have been used to treat both intractable SIB and severe Tourette's Disorder. Understandably, there are few reports concerning psychosurgery in children and adolescents for any condition or disorder. This report describes the use of cingulotomy and subsequent limbic leucotomy in an adolescent boy with Tourette's Disorder for SIB. His repetitive and medically serious SIB and failure of all other treatments prompted this intervention after careful, comprehensive review and discussion. Following the second surgery, the severity and frequency of his SIB were reduced. PMID- 15662145 TI - Quality is in the eye of the beholder. PMID- 15662146 TI - A double-blind, placebo-controlled withdrawal trial of dexmethylphenidate hydrochloride in children with attention deficit hyperactivity disorder. AB - OBJECTIVES: d,l-threo-methylphenidate HCl (D,L-MPH) is the most common treatment of attention deficit hyperactivity disorder (ADHD). A previous report showed placebo-controlled efficacy for the purified d-isomer (dexmethylphenidate hydrochloride, d-MPH, Focalin) with a 2:1 potency compared to dl, and suggested a 6-hour duration of action. This study complements that report by studying the effect of placebo-controlled discontinuation and retesting the duration of action. METHODS: A 6-week, open-label titration of d-MPH (2.5-10 mg twice-a-day) was followed by a double-blind, placebo-controlled, 2-week withdrawal study of responders. RESULTS: In the open titration, 82% of the 89 enrolled patients achieved a Clinical Global Impression-Improvement (CGI-I) rating of much or very much improved. Only 5 patients discontinued for adverse events. Seventy-five patients continued into the placebo-controlled discontinuation. For the randomly assigned d-MPH (n=35) and placebo (n=40) groups, mean ages, respectively, were 10.1 +/- 2.9 and 9.9 +/- 2.7 years, 86% and 78% were male, and 70.6% and 80.0% took the ceiling dose of 10 mg twice-daily, respectively. Each group had 80% combined type ADHD and 20% inattentive type. By the end of the 2-week, placebo masked withdrawal, significantly more placebo patients (24 of 39) than d-MPH continuers (6 of 35) relapsed (61.5% versus 17.1%, p=0.001). Compared to d-MPH continuers, placebo patients deteriorated significantly more in the 2-week period on teacher ratings of the 18 ADHD symptoms rated 0-3 (p=0.028), the 3 p.m. and 6 p.m. parent ADHD symptom ratings (p=0.0026 and p=0.0381, respectively), and clinic (2-3 p.m.) and home (6 p.m.) Math Tests (p=0.024 and p<0.0001, respectively). The 6 p.m. scores replicated the significant effect at 6 hours reported in the previous study. CONCLUSIONS: d-MPH is safe, tolerable, and effective, with a 6-hour duration of effect suggested by the significant difference from placebo at 6 hours on a double-blind discontinuation. PMID- 15662147 TI - Open-label study of dexmethylphenidate hydrochloride in children and adolescents with attention deficit hyperactivity disorder. AB - BACKGROUND: This pilot study examined the efficacy and duration of the effect of dexmethylphenidate (d-MPH) given once-daily in subjects with attention deficit hyperactivity disorder (ADHD). METHOD: Subjects aged 6-18 years (inclusive) with ADHD were enrolled in this 8-week, openlabel study. Outcome measures included the Conners'Teacher and Parent Rating Scales, the Attention Deficit Disorder Rating Scale (ADDRS), the Clinical Global Impression (CGI) Scale, and teacher and parent visual analog scales to estimate the duration of efficacy. d-MPH was initiated at a dose of 2.5 mg/day. The dose was flexible, based on response and tolerability, and could be increased in increments of 2.5 mg/day to a maximum daily dose of 30 mg/day. RESULTS: Twenty-two subjects (mean age, 8.7 +/- 0.4 years) were treated. Significant improvements (p <0.0001) from baseline occurred in the Conners' Teacher and Parent Rating Scales after 8 weeks. Of the evaluated subjects, 85.7% (18 of 21) showed at least a 30% improvement from baseline on the Conners' Teacher Rating Scale, and 86.4% (19 of 22) of subjects showed at least a 30% improvement from baseline on the Conners' Parent Rating Scale. Most subjects demonstrated an improvement on the ADDRS and the CGI-Improvement (CGI-I) scale. Median duration of effect was estimated at 6.2 hours (teachers) and at 7.5 hours (parents). On average, patients gained 2.4 pounds over the course of the study. CONCLUSIONS: A single daily dose of d-MPH was effective in controlling ADHD in children and was well tolerated. Future studies are needed to confirm these findings and to evaluate chronic dosing with d-MPH. PMID- 15662148 TI - Dopamine receptor 4 (DRD4) 7-repeat allele predicts methylphenidate dose response in children with attention deficit hyperactivity disorder: a pharmacogenetic study. AB - OBJECTIVE: Genetic polymorphisms of the dopamine neurotransmitter system have been identified in attention deficit hyperactivity disorder (ADHD). Since stimulant medications act through this system, we sought to determine if the 48 base pair VNTR polymorphism (7- repeat allele) of dopamine receptor gene DRD4 predicts methylphenidate responsiveness. METHODS: Forty-five children, aged 7-15 years, with ADHD, confirmed by NIMH-DISC-IV, participated in this prospective pharmacogenetic study. Subjects received increasing methylphenidate doses based on serial Conners' Global Index-Parent assessments. Doses to obtain a 10-point improvement and normalization (T-score, 60) were determined. Blood and buccal screening for DRD4 7R was correlated with outcomes. RESULTS: Mean dose for a 10 point CGI-P improvement with DRD4 7R (n=20) was 30 mg (1.00 mg/kg) versus 20 mg (0.49 mg/kg) without 7R (n=25) (log rank=13.69; df=1; p=0.0002). Mean dose for CGI-P normalization for children with 7R was 47 mg (1.70 mg/kg) of methylphenidate versus 31 mg (0.79 mg/kg) of methylphenidate without 7R (log rank=14.17; df=1; p=0.0002). ADHD symptom normalization at < or =50 mg methylphenidate was achieved in 58% with 7R versus 95% without (log rank=9.45; df=1; p=0.002). CONCLUSIONS: Children with ADHD possessing the DRD4 7R allele require higher doses of methylphenidate for symptom improvement and symptom normalization. This pharmacogenetic study demonstrates that the 7-repeat allele of the DRD4 gene VNTR polymorphism correlates with treatment outcomes. PMID- 15662149 TI - Effect of methylphenidate formulation for attention deficit hyperactivity disorder on patterns and outcomes of treatment. AB - OBJECTIVES: The aim of this study was to assess whether once-daily Concerta extended-release (XR) methylphenidate (MPH) is associated with: (1) better compliance; (2) decreased likelihood of accidents/injuries; and (3) lower health care resource use over a 1-year period than 3-times-daily immediate-release (IR) MPH in children with attention deficit hyperactivity disorder (ADHD). METHODS: Multivariate regression analyses were performed using an administrative database. Inclusion criteria were: (1) an age of 6-12 years at the date of first prescription for XR/IR MPH (index date); (2) patient-level data available for least 6 months before and 12 months after the index date; (3) no ADHD medications in the 6-month prior period; and (4) no XR MPH use by the IR MPH group. RESULTS: IR MPH: n=344, mean age 9.55 years, 76% male; XR MPH: n=1431, mean age 9.78 years, 75% male. XR MPH patients were less likely to discontinue, less likely to switch, and more likely to persist (i.e., no gaps >14 days) with therapy (p <0.0001). Children receiving XR MPH were less likely to experience an accident or injury (OR=0.58, 95% CI 0.353-0.945). Use of XR MPH was associated with significantly fewer emergency room visits and general practitioner visits per patient, on average, over 1 year. CONCLUSIONS: These data support the use of XR MPH with its simplified dosing regimen for children with ADHD. PMID- 15662150 TI - Nutritional supplements and complementary/alternative medicine in Tourette syndrome. AB - OBJECTIVE: Tourette syndrome (TS) is a neuropsychiatric disorder associated with motor and vocal tics. Some people with TS have reported using alternative or complementary medicine (CAM), including nutritional supplements to control their tics. In a recent national survey, approximately 40% of people reported having used CAM in the prior year. We attempted to explore the use of supplements and other CAM among TS patients. METHOD: We developed a survey instrument based on anecdotal accounts of CAM use and distributed it to the mailing list of the New York Chapter of the Tourette Syndrome Association (n=500) and the subscription list of Latitudes, a newsletter exploring CAM treatments for neurological conditions (n=750). Responses were entered in a database and analyzed using SPSS version 10. RESULTS: Of 115 respondents, 87.8% reported using 1 or more of 29 nutritional supplements to control symptoms. Many also reported using other CAM. Most supplement users reported an improvement in tics. CONCLUSIONS: Although these results are not generalizable, they provide no evidence that use of supplements and other CAM is rarer among TS patients than in the general population. Given that most of our respondents were also using conventional medication, further study of the use of supplements and other CAM therapies by people with TS, the safety and efficacy of such therapies in this population, and possible interactions of such therapies with conventional treatment seems warranted. PMID- 15662151 TI - Intramuscular ziprasidone in youth: a retrospective chart review. AB - OBJECTIVE: The aim of this study was to assess the tolerability and effectiveness of intramuscular ziprasidone in inpatient children and adolescents. METHOD: A retrospective chart review was conducted of child and adolescent inpatients at a private psychiatric hospital in central New York between January 1, 2003 and August 30, 2003. Subjects who had received intramuscular ziprasidone were identified, and their age, gender, and dosage were recorded. Nursing-shift notes were reviewed to ascertain the need for repeat injection and for adverse reactions. RESULTS: Forty-nine youths received intramuscular ziprasidone; 43 injections were 20 mg and 6 injections were 10 mg. Readministration within 4 hours was required in only 1 subject. No adverse events were recorded. CONCLUSION: Intramuscular ziprasidone was well tolerated and effective for acutely agitated and aggressive inpatient youth. PMID- 15662152 TI - The effectiveness and tolerability of aripiprazole for pediatric bipolar disorders: a retrospective chart review. AB - OBJECTIVE: The aim of this retrospective chart review was to evaluate the effectiveness and tolerability of aripiprazole for the treatment of children and adolescents with bipolar disorders. METHODS: The medical charts of all children and adolescents with a DSM-IV diagnosis of bipolar disorder, type I, type II, not otherwise specified (NOS), or schizoaffective disorder, bipolar type, and who were treated with aripiprazole were reviewed by two child and adolescent psychiatrists who independently confirmed their DSM-IV diagnoses, severity, and the improvement of illness using the Clinical Global Impression (CGI) Severity and Improvement scores for bipolar disorder (CGI-BP) and the Clinical Global Assessment Scale (CGAS). RESULTS: Thirty patients who were treated with aripiprazole were identified (mean starting dose=9 +/- 4 mg/day, mean final dose=10 +/- 3 mg/day). The overall response rate, defined by a CGI-Improvement score of < or = 2 at endpoint, was 67%. There was a statistically significant improvement in CGAS scores (48 +/- 11 to 65 +/- 11, signed rank = 191, p <0.0001) and CGI-S scores (4.2 +/- 0.8 to 2.8 +/- 1.0, signed rank=-172, p <0.0001, effect size=1.90) from baseline to endpoint. No serious adverse events were identified. Common side effects were sedation (n=10, 33%), akathisia (n=7, 23%), and gastrointestinal disturbances (n=2, 7%). Baseline and endpoint weights were available for 14 (47%) of the patients. Change in weight ranged from +5 to -21 kg and 12 (86%) of 14 patients lost weight (mean weight loss was 3 +/- 6 kg). CONCLUSIONS: This retrospective chart review suggests that aripiprazole may be effective and well tolerated for children and adolescents with bipolar disorders. Controlled studies of aripiprazole for the treatment of pediatric bipolar disorder are necessary. PMID- 15662153 TI - Behavioral training for pill-swallowing difficulties in young children with autistic disorder. AB - INTRODUCTION: Difficulty with swallowing pills is a common problem, leading to noncompliance with treatment recommendations. Many young children with autistic disorder (AD) who also show comorbid symptoms associated with attention deficit hyperactivity disorder (ADHD) have difficulty swallowing pills. This pilot study describes our experience in teaching pill-swallowing skills to 4 children with AD who also had comorbid symptoms associated with ADHD. METHODS: Four children, aged 5-;6.5 years, were enrolled for pill-swallowing training, 3 of the children were Caucasian boys and 1 child was a Hispanic girl. All children met the Diagnostic and Statistical Manual of Mental Disorders, 4th edition (DSM-IV) diagnostic criteria for AD and ADHD-like symptoms. The children's verbal IQ ranged from 54 96, their nonverbal IQ ranged from 85-107, and the Preschool Language Scale-3 total language score ranged from 50-98. RESULTS: At the end of the pilot study, 2 children (50%) successfully learned to swallow the study capsules, 1 child (25%) was able to swallow the study capsules with the behavior therapist but had difficulty with the caregiver, and 1 child (25%) made slow progress and was withdrawn by the caregiver in favor of proceeding with a crushable medication for clinical care. CONCLUSION: Caregivers were appreciative of the opportunity for this short intervention. Behavioral training for pill swallowing may be indicated in some circumstances in young children with AD and/or other developmental disorders. PMID- 15662154 TI - Olanzapine-induced diabetes in a seven-year-old boy. AB - The atypical antipsychotics have been recognized to induce diabetes mellitus and ketoacidosis in the adult psychiatric population. This report notes the onset of weight gain, diabetes, and apparent ketosis in a prepubertal boy diagnosed with bipolar disorder and treated with olanzapine. The hyperglycemia rapidly normalized after discontinuation of the olanzapine. Within 2 years, the diabetes recurred. In spite of the normalization of blood-glucose levels, urine ketone tests remained positive and were explained by the fact that patients taking valproic acid may have a false-positive urine test for ketones. Regular monitoring of glucose should be considered in children and adolescents who gain weight while treated with atypical antipsychotics. PMID- 15662155 TI - Adolescent weight loss during treatment with olanzapine. AB - Antipsychotic medications are increasingly used in adolescents for a variety of psychiatric difficulties, including psychosis, bipolar disorder, and aggression. Weight gain is a significant side effect of neuroleptics, which may limit adolescents' compliance with these medications. This report presents a case of significant weight loss while on olanzapine, with a body mass index (BMI) falling from 25 to 19.5 during 8 months of treatment. Possible mechanisms for this effect are discussed. PMID- 15662156 TI - The effectiveness of risperidone in the treatment of three children with feeding disorders. AB - Children with histories of extensive medical interventions in early childhood, especially those involving the gastrointestinal tract, are at-risk for residual feeding problems long after the medical issues have resolved. This case series describes the inpatient multidisciplinary treatment of 3 consecutive preadolescent children (ages 9, 8, and 7 years) admitted to our rehabilitation facility. Each child was admitted with a severe feeding disorder and histories of failure-to-thrive (FTT) that had required enteral nutritional interventions. Each child also had comorbid anxiety and mood symptoms. The addition of risperidone to behavioral and psychopharmacologic treatments was observed to significantly increase oral intake and accelerate weight gain. In 2 of 3 patients, assistive feeding interventions were successfully discontinued; and in a 3rd patient, enteral nutritional support was reduced by 74%. These cases suggest that risperidone may be a safe and effective adjunctive treatment, when behavioral feeding therapy is not sufficiently successful for children who have chronic and complicated medical and psychiatric presentations. PMID- 15662164 TI - Respecting each other. PMID- 15662165 TI - Tell it to the Judge. PMID- 15662167 TI - Palliative sedation in the control of refractory symptoms. PMID- 15662168 TI - Palliative care outcomes research: the next steps. PMID- 15662169 TI - None left behind: urban poverty, social experience, and rethinking palliative care. PMID- 15662171 TI - Impact of palliative care case management on resource use by patients dying of cancer at a Veterans Affairs medical center. AB - BACKGROUND: The VA Puget Sound Health Care System (VAPSHCS) started a palliative care service (PCS) in October 2001 to provide case management for patients with advanced cancer. OBJECTIVE: To examine resource use during the last 60 days of life for patients dying of cancer who received PCS compared to patients receiving usual care (non-PCS) during the same time period. DESIGN: Retrospective nonrandomized comparison of resource use using administrative data. SETTING: Tertiary care Veterans Affairs medical center. PARTICIPANTS: All patients who died of cancer between October 1, 2001 and October 31, 2002 at VAPSHCS. RESULTS: Two hundred sixty-five patients died of cancer during the specified time period, including 82 PCS and 183 non-PCS patients. PCS patients received case management for a mean of 79 days, and were younger, had more comorbid conditions, and were more likely to have had chemotherapy in the last 60 days of life than non-PCS patients. Variables associated with more acute care bed days in the last 60 days of life included: chemotherapy in the last 60 days of life, and a length of stay on PCS less than 60 days. Variables associated with fewer acute care bed days within the last 60 days of life included: being married, and a length of stay on PCS 60 days or more. Compared to non-PCS patients, the place of death for PCS patients was less likely to be acute care. CONCLUSION: PCS for 60 or more days prior to death was associated with decreased use of acute care hospital resources for patients dying of cancer. PMID- 15662170 TI - Sedation for terminally ill patients with cancer with uncontrollable physical distress. AB - BACKGROUND: Relief of distressful symptoms in terminally ill patients with cancer is of prime importance. Use of sedation to accomplish this has been the focus of recent medical studies in countries other than Japan. We investigated the influence on consciousness of sedative drugs in a Japanese hospice. DESIGN AND SUBJECTS: We defined sedation as medical procedure to decrease level of consciousness in order to relieve severe physical distress refractory to standard interventions. We excluded increases in doses of morphine or other analgesic drugs resulting in secondary somnolence from the present study. We reviewed medical records of patients receiving sedation among 124 consecutive patients admitted to our palliative care unit between January and December in 1999. RESULTS: The 63 patients who received sedation (50.3%) died an average of 3.4 days after its initiation. Major symptoms requiring sedation were dyspnea, general malaise/restlessness, pain, agitation, and nausea/vomiting. The Palliative Performance Status (PPS) just before sedation was 20 or less in 83% of patients. Drugs administered for sedation were midazolam, haloperidol, scopolamine hydrobromide, and chlorpromazine. During the few days before death, sedated patients were significantly more drowsy and less responsive than that in those receiving non-sedative treatment. CONCLUSIONS: Our data suggest the effectiveness of sedation in relieving severe, refractory physical symptoms in terminally ill Japanese patients with cancer. Further investigation to confirm safety and effectiveness of sedation in this context is warranted. PMID- 15662172 TI - Attitudes, experiences, and beliefs affecting end-of-life decision-making among homeless individuals. AB - BACKGROUND: Individuals who are homeless may encounter various barriers to obtaining quality end-of-life (EOL) care, including access barriers, multiple sources of discrimination, and lack of knowledge among health care providers (HCPs) of their preferences and decision-making practices. Planning for death with individuals who have spent so much energy surviving requires an understanding of their experiences and preferences. OBJECTIVE: This study sought to increase HCPs' awareness and understanding of homeless or similarly marginalized individuals' EOL experiences and treatment preferences. DESIGN: Focus groups were conducted with homeless individuals using a semi-structured interview guide to elicit participants' EOL experiences, decision-making practices, and personal treatment preferences. SETTING/SUBJECTS: Five focus groups were conducted with 20 inner-city homeless individuals (4 per group) at a free urban health care clinic for homeless individuals in the United States. Sixteen of the 20 participants were African American; 4 were Caucasian. None were actively psychotic. All had experienced multiple losses and drug addiction. FINDINGS: Five main themes emerged: valuing an individual's wishes; acknowledging emotions; the primacy of religious beliefs and spiritual experience; seeking relationship-centered care; and reframing advance care planning. CONCLUSIONS: The narrative process of this qualitative study uncovered an approach to EOL decision making in which participants' reasoning was influenced by emotions, religious beliefs, and spiritual experience. Relationship-centered care, characterized by compassion and respectful, two-way communication, was obvious by its described absence--reasons for this are discussed. Recommendations for reframing advance care planning include ways for HCPs to transform advance care planning from that of a legal document to a process of goal-setting that is grounded in human connection, respect, and understanding. PMID- 15662173 TI - Parenteral ketamine as an analgesic adjuvant for severe pain: development and retrospective audit of a protocol for a palliative care unit. AB - BACKGROUND: Ketamine is an effective analgesic agent for treating a variety of neuropathic and cancer pain syndromes. Recent studies indicate that ketamine may have a particular role in the management of patients with neuropathic and/or pain syndromes that are poorly responsive to opioids. OBJECTIVE: To develop, implement, and subsequently assess a protocol designed to maximize the analgesic effect of ketamine while minimizing its side effects. DESIGN: A retrospective chart audit of 16 patients who had used the ketamine protocol over a 12-month period. Criteria for assessing the effectiveness of ketamine were defined. RESULTS: Ketamine was an effective, well-tolerated analgesic adjuvant for 11 of 16 patients with previously uncontrolled pain. Pain scores were reduced by at least 4 of 10 in 15 of the 16 patients. Median opioid dose reduction on starting ketamine was 25%. CONCLUSION: The audit confirmed the safety and effectiveness of ketamine as an analgesic adjuvant for patients with severe pain. Baseline opioid dose reduction and prophylactic use of haloperidol or benzodiazepine were effective in minimizing psychotomimetic side effects. PMID- 15662174 TI - End-of-life hospitalization for African American and non-Latino white nursing home residents: variation by race and a facility's racial composition. AB - BACKGROUND: Hospitalization of nursing home residents at the end of life is common, more so among African Americans. Whether a nursing home's racial mix is associated with hospitalization is unknown. OBJECTIVE: This study examined the association between race, a nursing home's racial mix, and end-of-life hospitalization. DESIGN: This was a retrospective cohort study. SETTING/SUBJECTS: Studied were nursing home residents in New York (n = 14,159) and Mississippi (n = 1481) who died in 1995-1996 and had a minimum data set (MDS) assessment within 120 days of death. MEASUREMENTS: The outcome measure was the odds of hospitalization in the last 90 days of life. A variable reflecting a nursing home's proportion of African American residents (in 1995-1996) represented a nursing home's racial mix. RESULTS: Forty-six percent of African Americans and 32% of whites were hospitalized in the last 90 days of life. After controlling for demographics, diagnoses, function, patient preferences (do-not-resuscitate [DNR]), and facility resources, nursing home residents in facilities having higher proportions of African American residents had greater odds of hospitalization (adjusted odds ratio [AOR] 1.14; 95% confidence interval [CI] 1.10, 1.18 in New York and AOR 1.35; 95% CI 1.24, 1.46 in Mississippi). Age and frailty interacted with race; older African Americans had a 16% greater likelihood (95% CI 1.08, 1.24) of hospitalization, and African Americans with more functional limitations had a 37% (95% CI 1.24, 1.51) greater likelihood of hospitalization than did comparable whites. CONCLUSIONS: It appears higher end-of life hospitalization rates for African American residents are attributable to the facilities where most reside, and to differential hospitalization of older or more functional limited residents. PMID- 15662175 TI - Service preferences among family caregivers of the terminally ill. AB - OBJECTIVE: The ability of families to assume caregiving responsibilities is contingent on material, social, and professional support. Inadequate or inappropriate support to the terminally ill and their family caregivers can result in the misuse of resources and add burden to the family. In this report, we describe service preferences among informal caregivers of the terminally ill. DESIGN: Three hundred seventy-three caregivers participated in telephone interviews at two points in time: when the terminally ill person was designated as palliative and 5 months subsequent to the first interview. In the case that the care recipient died during the study period, the caregiver participated in the interview three months after the death. MEASURES: After reviewing possible services received by the care recipients and caregivers, caregivers were asked to identify the five services they found most valuable and which services they would have liked to have had or received more of when caregiving. RESULTS: The five services caregivers reported as most valuable included: in-home nursing care, (90.7%); family physicians, (45.6%); medical specialists, (46.4%); housekeeping, (23.6%); and, religious support, (11.3%). The five most frequently reported services that family caregivers would have liked to have received or had more available included: housekeeping, (13.1%); caregiver respite, (10.2%); in-home nursing care, (8.0%); personal support workers, (4.6%); and, self-help/support groups, (3.8%). Analyses revealed that most (64.8%) perceived service needs were of a supportive nature for caregivers. Caregiver perceptions of the value and perceived need of services were consistent over time and into bereavement. Logistic regression analyses suggested that younger caregivers who were not employed, reported higher levels of burden and cared for someone with a diagnosis of cancer had greater perceived service needs. CONCLUSIONS: The findings reported in this paper provide important insights into caregiver perceptions of valued services when caring for a terminally ill family member. These finding also highlight the stability of caregiver service perceptions over time and into bereavement. PMID- 15662176 TI - The long-term impact of dialysis discontinuation on families. AB - BACKGROUND: Little is known about the long-term psychological impact of stopping life support treatments on surviving loved ones. OBJECTIVE: The authors sought to determine if there was an increase in pathologic grief in family members left behind after deaths that followed dialysis discontinuation. DESIGN: Phone interviews were used to collect data on demographics, attitudes, and families' comfort levels with the decision to withdraw dialysis. The Impact of Event Scale was administered to assess adaptation and stress levels. Avoidance and Intrusiveness subscales were calculated and associations with other survey data were examined using chi2 tests and analysis of variance (ANOVA). SETTING/SUBJECTS: The authors contacted families in New England who had previously participated in the Baystate Dialysis Discontinuation Study. MEASUREMENTS/RESULTS: Twenty-six family members (66% of the original study sample) were interviewed approximately 55 months after patient deaths. There was a low overall level of distress and the Avoidance subscale had insufficient variability for analysis. Intrusiveness was highest for spouses and primary caregivers. Only one respondent remembered the death as having been "bad," although 62% of patients were recalled as having suffered distressing symptoms in their last days. In ascending order of importance, respondents characterized good deaths as involving mental alertness, occurring at home, taking place while asleep, being peaceful, happening in the company of loved ones, and being painless or largely painfree. Almost all of the families reported becoming more comfortable with the decision to hasten death than originally. CONCLUSIONS: After nearly 5 years after dialysis discontinuation, families report low levels of distress. A higher frequency of intrusive thoughts was more likely if respondents were spouses or primary caregivers as compared to adult children, siblings, or other relatives. The findings suggest that families successfully adapt to the impact of dialysis withdrawal deaths. PMID- 15662177 TI - Cancer-related deaths in children and adolescents. AB - BACKGROUND: To add to the data regarding the quality of care given to dying children and their families. OBJECTIVE: To develop baseline of end-of-life care at a single pediatric facility to evaluate institution-wide palliative care initiative. DESIGN: Retrospective chart review of all known deaths during an 18 month time period. SETTING/SUBJECTS: One hundred forty-five charts of patients from a single pediatric cancer facility who died during designated time period. MEASUREMENTS: Variables included: cause and place of death; CPR/DNR status prior to death; length of end of life care prior to death; sibling counseling and bereavement counseling offered to family after death; and wishes or preferences of patient/family regarding the death experience. RESULTS: Results included: solid tumor patients more likely to die of progressive disease than leukemia or bone marrow transplant patients; bone marrow transplant patients 2-3 times more likely to die of cardiopulmonary or cardiovascular complications; solid tumor patients were more likely to die at home than leukemia patients; solid tumor/brain tumor patients had a median time in end of life or palliative care of 29 days compared to leukemia patients' median of 11 days; 48% of DNRs completed 11 days prior to death. CONCLUSIONS: Relationship exists between diagnosis, cause and place of death in this population; findings replicate findings of 4 similar studies; accurate and consistent quality standards of care need to be established for this population as well as methods of documentation before reviewing/accrediting agencies impose standards that are not evidence based. PMID- 15662178 TI - Health behaviors associated with better quality of life for older bereaved persons. AB - BACKGROUND: Bereavement is a risk factor for declines in health, adverse health behaviors, increased physician visits, and mortality, and occurs with greatest frequency in later life. Little is known about health behaviors that are associated with better quality of life among recently bereaved older persons. OBJECTIVE: The objective of this community-based longitudinal, observational study was to examine the influence of health behaviors on the quality of life of 200 elderly bereaved persons. DESIGN AND MEASUREMENTS: Health behaviors (i.e., exercise, monitoring caloric intake, sleep, daily vitamin intake, annual health check-ups) were examined at approximately 6 months postloss (baseline) and 11 months postloss (Wave 2). Quality of life was assessed at approximately 11 months postloss and 19 months postloss (Wave 3), using the RAND 36-Item Health Survey, which measures 8 domains of health and functioning, plus a single item assessing change in health. RESULTS: Consistently exercising 1 or more days per week at Waves 1 and 2 significantly (p < .05) predicted better self-rated health, physical functioning, fewer physical role limitations, and greater energy at Wave 3 in models that adjusted for age, gender, prior psychiatric disorder, baseline reports of functional disabilities and chronic conditions. Consistently monitoring caloric intake at Waves 1 and 2 predicted better self-rated health (p < .05), greater energy (p < .01), and positive change in health (p < .05) at Wave 3 in models that adjusted for the above set of control variables. Sleeping 6.5-9 hours per night at baseline alone predicted better social functioning (p < .001), fewer emotional role limitations (p < .01), better emotional health (p < .001), and greater energy (p < .01). CONCLUSIONS: Should future research confirm these results, clinicians would be advised to recommend the identified preventive and protective health behaviors to recently bereaved older patients. PMID- 15662179 TI - Evaluation of the End-of-Life Nursing Education Consortium undergraduate faculty training program. AB - BACKGROUND: Previous research has demonstrated that nursing education has not prepared nurses to provide optimum end-of-life (EOL) care; and yet, care of patients at the EOL is contingent on adequate preparation of nurses. To date, there has not been a unified or organized effort to broadly address the preparation of nurses in EOL care. OBJECTIVE: The purpose of the End-of-Life Nursing Education Consortium (ELNEC), a Robert Wood Johnson Foundation funded project (2000-2004), was to develop and implement a comprehensive national effort to improve EOL care by nurses through a joint collaboration between the American Association of Colleges of Nursing (AACN) and the City of Hope Cancer Center. DESIGN AND SETTING: Based on the AACN Peaceful Death document, the ELNEC curriculum focuses on nine EOL core areas. This project is a synthesis of research and knowledge in EOL care and is intended to assist clinical nurses with implementing scientifically based care in practice. Eight national training courses followed the development of the core training curriculum to enhance EOL expertise in faculty in undergraduate nursing programs (five courses) as well as in continuing education programs (three courses). Development of the ELNEC program included detailed teaching materials to integrate EOL content into existing nursing curricula and clinical teaching and extensive follow up evaluation. RESULTS: The data revealed significant outcomes in the report of implementation in the nursing curriculum including an increase in the amount of content, perceived effectiveness of new graduates, and of faculty expertise in EOL care, and a broad dissemination of all modules geographically. CONCLUSION: This national organized effort is a major step toward preparing nurses in EOL care to improve care of the dying. PMID- 15662180 TI - Who enrolls in observational end of life research? Report from the cultural variations in approaches to end of life study. AB - BACKGROUND: Response bias in end of life research is important though not well understood. OBJECTIVES: To compare consenting advanced cancer patient participants and refusers in observational end of life research. We hypothesized that, compared with refusers, consenters would: 1) have a stronger belief in personal gain from research participation, 2) be more satisfied with their medical care, 3) be more satisfied with support from family and friends, and 4) be in less physical discomfort compared with refusers. METHODS: Sixty eight patients consented to enroll in the 'parent' observational study. Thirty six patients refused to participate. Reasons for refusal were recorded verbatim and coded using qualitative techniques. Both patient consenters and patient refusers were asked the same questions regarding personal gain, satisfaction with medical care, family and social support, and physical discomfort. RESULTS: Consenters believed that they had more to gain from participation in research compared with refusers (p = 0.04). Consenters felt that aches or pain were more of a problem for them compared with refusers (p < 0.001). Both satisfaction with medical care and with support from family and friends were similar between consenters and refusers. CONCLUSIONS: Consenting study participants in observational research at end of life believe they have more to gain from study participation than do refusers. Contrary to our hypothesis, consenting participants were those who were experiencing greater physical discomfort compared to refusers. PMID- 15662181 TI - Evaluation of the Missoula-VITAS Quality of Life Index--revised: research tool or clinical tool? AB - BACKGROUND: Quality of life (QOL) is a central outcome measure in caring for seriously ill patients. The Missoula-VITAS Quality of Life Index (MVQOLI) is a 25 item patient-centered index that weights each of five QOL dimensions (symptoms, function, interpersonal, wellbeing, transcendence) by its importance to the respondent. The measure has been used to assess QOL for hospice patients, and has been found to be somewhat complex to use and analyze. OBJECTIVE: This study aimed to simplify the measure, and evaluate the reliability and validity of a revised version as either a research or clinical tool (i.e., "psychometric" versus "clinimetric"). DESIGN: Two data collection efforts are described. The psychometric study collected QOL data from 175 patients at baseline, 3-5 days, and 21 days later. The implementation study evaluated the feasibility and utility of the MVQOLI-R during over six weeks of use. SETTING/SUBJECTS: End-stage renal patients on dialysis, hospice, or long-term care patients participated in the psychometric study. The implementation study was done in hospice, home health, and palliative care settings. MEASUREMENTS: The MVQOLI-R and the Memorial Symptom Assessment Scale. RESULTS: The psychometric and implementation studies suggest that the MVQOLI-R performs well as a clinical tool but is not powerful as an outcome research instrument. The MVQOLI-R has the heterogeneous structure of clinimetric tools, and demonstrated both relevance and responsiveness. Additionally, in a clinical setting the MVQOLI-R was useful therapeutically for stimulating communication about the psychosocial and spiritual issues important to the tasks of life completion and life closure. CONCLUSIONS: The MVQOLI-R has clinical utility as a patient QOL assessment tool and may have therapeutic utility as a tool for fostering discussion among patients and their clinicians, as well as for helping patients identify sources of suffering and opportunities during this time in their lives. PMID- 15662182 TI - Controlled sedation for refractory suffering: part I. PMID- 15662183 TI - Saying goodbye. PMID- 15662184 TI - Waiting time. PMID- 15662185 TI - Controlled sedation for physical and existential suffering? PMID- 15662189 TI - The family conference. PMID- 15662195 TI - LD-SPatt: large deviations statistics for patterns on Markov chains. AB - Statistics on Markov chains are widely used for the study of patterns in biological sequences. Statistics on these models can be done through several approaches. Central limit theorem (CLT) producing Gaussian approximations are one of the most popular ones. Unfortunately, in order to find a pattern of interest, these methods have to deal with tail distribution events where CLT is especially bad. In this paper, we propose a new approach based on the large deviations theory to assess pattern statistics. We first recall theoretical results for empiric mean (level 1) as well as empiric distribution (level 2) large deviations on Markov chains. Then, we present the applications of these results focusing on numerical issues. LD-SPatt is the name of GPL software implementing these algorithms. We compare this approach to several existing ones in terms of complexity and reliability and show that the large deviations are more reliable than the Gaussian approximations in absolute values as well as in terms of ranking and are at least as reliable as compound Poisson approximations. We then finally discuss some further possible improvements and applications of this new method. PMID- 15662196 TI - Modeling and analysis of heterogeneous regulation in biological networks. AB - In this study, we propose a novel model for the representation of biological networks and provide algorithms for learning model parameters from experimental data. Our approach is to build an initial model based on extant biological knowledge and refine it to increase the consistency between model predictions and experimental data. Our model encompasses networks which contain heterogeneous biological entities (mRNA, proteins, metabolites) and aims to capture diverse regulatory circuitry on several levels (metabolism, transcription, translation, post-translation and feedback loops, among them). Algorithmically, the study raises two basic questions: how to use the model for predictions and inference of hidden variables states, and how to extend and rectify model components. We show that these problems are hard in the biologically relevant case where the network contains cycles. We provide a prediction methodology in the presence of cycles and a polynomial time, constant factor approximation for learning the regulation of a single entity. A key feature of our approach is the ability to utilize both high-throughput experimental data, which measure many model entities in a single experiment, as well as specific experimental measurements of few entities or even a single one. In particular, we use together gene expression, growth phenotypes, and proteomics data. We tested our strategy on the lysine biosynthesis pathway in yeast. We constructed a model of more than 150 variables based on an extensive literature survey and evaluated it with diverse experimental data. We used our learning algorithms to propose novel regulatory hypotheses in several cases where the literature-based model was inconsistent with the experiments. We showed that our approach has better accuracy than extant methods of learning regulation. PMID- 15662197 TI - Permutation pattern discovery in biosequences. AB - Functionally related genes often appear in each other's neighborhood on the genome; however, the order of the genes may not be the same. These groups or clusters of genes may have an ancient evolutionary origin or may signify some other critical phenomenon and may also aid in function prediction of genes. Such gene clusters also aid toward solving the problem of local alignment of genes. Similarly, clusters of protein domains, albeit appearing in different orders in the protein sequence, suggest common functionality in spite of being nonhomologous. In the paper, we address the problem of automatically discovering clusters of entities, be they genes or domains: we formalize the abstract problem as a discovery problem called the (pi)pattern problem and give an algorithm that automatically discovers the clusters of patterns in multiple data sequences. We take a model-less approach and introduce a notation for maximal patterns that drastically reduces the number of valid cluster patterns, without any loss of information, We demonstrate the automatic pattern discovery tool on motifs on E. Coli protein sequences. PMID- 15662198 TI - PROuST: a comparison method of three-dimensional structures of proteins using indexing techniques. AB - We present a new method for protein structure comparison that combines indexing and dynamic programming (DP). The method is based on simple geometric features of triplets of secondary structures of proteins. These features provide indexes to a hash table that allows fast retrieval of similarity information for a query protein. After the query protein is matched with all proteins in the hash table producing a list of putative similarities, the dynamic programming algorithm is used to align the query protein with each protein of this list. Since the pairwise comparison with DP is applied only to a small subset of proteins and, furthermore, DP re-uses information that is already computed and stored in the hash table, the approach is very fast even when searching the entire PDB. We have done extensive experimentation showing that our approach achieves results of quality comparable to that of other existing approaches but is generally faster. PMID- 15662199 TI - Robust sparse hyperplane classifiers: application to uncertain molecular profiling data. AB - Molecular profiling studies can generate abundance measurements for thousands of transcripts, proteins, metabolites, or other species in, for example, normal and tumor tissue samples. Treating such measurements as features and the samples as labeled data points, sparse hyperplanes provide a statistical methodology for classifying data points into one of two categories (classification and prediction) and defining a small subset of discriminatory features (relevant feature identification). However, this and other extant classification methods address only implicitly the issue of observed data being a combination of underlying signals and noise. Recently, robust optimization has emerged as a powerful framework for handling uncertain data explicitly. Here, ideas from this field are exploited to develop robust sparse hyperplanes, i.e., classification and relevant feature identification algorithms that are resilient to variation in the data. Specifically, each data point is associated with an explicit data uncertainty model in the form of an ellipsoid parameterized by a center and covariance matrix. The task of learning a robust sparse hyperplane from such data is formulated as a second order cone program (SOCP). Gaussian and distribution free data uncertainty models are shown to yield SOCPs that are equivalent to the SCOP based on ellipsoidal uncertainty. The real-world utility of robust sparse hyperplanes is demonstrated via retrospective analysis of breast cancer related transcript profiles. Data-dependent heuristics are used to compute the parameters of each ellipsoidal data uncertainty model. The generalization performance of a specific implementation, designated "robust LIKNON," is better than its nominal counterpart. Finally, the strengths and limitations of robust sparse hyperplanes are discussed. PMID- 15662200 TI - Blind source separation and the analysis of microarray data. AB - We develop an approach for the exploratory analysis of gene expression data, based upon blind source separation techniques. This approach exploits higher order statistics to identify a linear model for (logarithms of) expression profiles, described as linear combinations of "independent sources." As a result, it yields "elementary expression patterns" (the "sources"), which may be interpreted as potential regulation pathways. Further analysis of the so-obtained sources show that they are generally characterized by a small number of specific coexpressed or antiexpressed genes. In addition, the projections of the expression profiles onto the estimated sources often provides significant clustering of conditions. The algorithm relies on a large number of runs of "independent component analysis" with random initializations, followed by a search of "consensus sources." It then provides estimates for independent sources, together with an assessment of their robustness. The results obtained on two datasets (namely, breast cancer data and Bacillus subtilis sulfur metabolism data) show that some of the obtained gene families correspond to well known families of coregulated genes, which validates the proposed approach. PMID- 15662201 TI - Sequencing from compomers: using mass spectrometry for DNA de novo sequencing of 200+ nt. AB - One of the main endeavors in today's life science remains the efficient sequencing of long DNA molecules. Today, most de novo sequencing of DNA is still performed using the electrophoresis-based Sanger concept of 1977, in spite of certain restrictions of this method. Methods using mass spectrometry to acquire the Sanger sequencing data are limited by short sequencing lengths of 15-25 nt. We propose a new method for DNA sequencing using base-specific cleavage and mass spectrometry that appears to be a promising alternative to classical DNA sequencing approaches. A single stranded DNA or RNA molecule is cleaved by a base specific (bio-)chemical reaction using, for example, RNAses. The cleavage reaction is modified such that not all, but only a certain percentage of bases are cleaved. The resulting mixture of fragments is then analyzed using MALDI-TOF mass spectrometry, whereby we acquire the molecular masses of fragments. For every peak in the mass spectrum, we calculate those base compositions that will potentially create a peak of the observed mass and, repeating the cleavage reaction for all four bases, finally try to uniquely reconstruct the underlying sequence from these observed spectra. This leads us to the combinatorial problem of sequencing from compomers and, finally, to the graph-theoretical problem of finding a walk in a subgraph of the de Bruijn graph. Application of this method to simulated data indicates that it might be capable of sequencing DNA molecules with 200+ nt. PMID- 15662202 TI - Identifying interventional and pathogenic mechanisms by generative inverse modeling of gene expression profiles. AB - MOTIVATION: The characterization of genetic mechanisms underlying normal cellular function, cancer development, pathogenesis, and the effect of drug treatment is one of the most challenging topics for cancer research and molecular biology. Existing methods for inferring genetic regulatory networks from genome-wide expression profiles provide important information about gene interactions and regulatory relationships. However, these methods do not provide information about the impact of possible interventions or changes on such regulatory networks to study cause-effect relationships at a systems-biology level. RESULTS: We present a data-driven method called generative inverse modeling, which simulates the effect of local genetic changes on the global cellular state, as reflected by an altered genome-wide expression profile. For each genetic change we define a pathogenic score by calculating to what extent it transforms the simulated expression patterns into patterns measured for pathologically altered tissues. The method can be used to estimate the relevance of genes for disease-specific genetic mechanisms, e.g., as presented here for pathogenesis. Generative inverse modeling is based on a Bayesian probability density estimation from a set of measured gene-expression patterns. PMID- 15662203 TI - Folding dynamics of proteins from denatured to native state: principal component analysis. AB - Several trajectories starting from random configurations and ending in the native state for chymotrypsin inhibitor 2, CI2, are generated using a Go-type model where the backbone torsional angles execute random jumps on which a drift towards their native values is superposed. Bond lengths and bond angles are kept fixed, and the size of the backbone atoms and side groups are recognized. The large datasets obtained are analyzed using a particular type of principal component analysis known as Karhunen-Loeve expansion (KLE). Trajectories are decomposed separately into modes in residue space and time space. General features of different folding trajectories are compared in the modal space and relationships between the structure of CI2 and its folding dynamics are obtained. Dynamic scaling and order reduction of the folding trajectories are discussed. A continuous wavelet transform is used to decompose the nonstationary folding trajectories into windows exhibiting different features of folding dynamics. Analysis of correlations confirms the known two-state nature of folding of CI2. All of the conserved residues of the protein are shown to be stationary in the small modes of the residue space. The sequential nature of folding is shown by examining the slow modes of the trajectories. The present model of protein folding dynamics is compared with the simple Rouse model of polymer dynamics. Principal component analysis is shown to be a very effective tool for the characterization of the general folding features of proteins. PMID- 15662204 TI - Spectral decomposition for the search and analysis of RNA secondary structure. AB - Scales in RNA, based on geometrical considerations, can be exploited for the analysis and prediction of RNA structures. By using spectral decomposition, geometric information that relates to a given RNA fold can be reduced to a single positive scalar number, the second eigenvalue of the Laplacian matrix corresponding to the tree-graph representation of the RNA secondary structure. Along with the free energy of the structure, being the most important scalar number in the prediction of RNA folding by energy minimization methods, the second eigenvalue of the Laplacian matrix can be used as an effective signature for locating a target folded structure given a set of RNA folds. Furthermore, the second eigenvector of the Laplacian matrix can be used to partition large RNA structures into smaller fragments. An illustrative example is given for the use of the second eigenvalue to predict mutations that may cause structural rearrangements, thereby disrupting stable motifs. PMID- 15662205 TI - An empirical Bayesian significance test of cDNA library data. AB - Automated high-throughput sequencing of cDNA clones from numerous libraries has generated a wealth of information about both genome sequence and relative transcript abundances. A common statistical challenge in the analysis of library sequences is to infer whether there is differential expression for the same transcript under two different conditions, such as normal and diseased tissue. In contrast to the continuously variable intensity measurements from microarray experiments, data from cDNA library sequencing presents itself as a discrete count of the incidence of some clone or transcript in a finite sample. In this paper, we first propose a statistical model for data generated from cDNA library sequencing efforts. The model is based on the Poisson mixed with generalized inverse Gaussian (PGIG), introduced by Sichel (1971, 1975). PGIG has been used in modeling population abundance, ecological studies, word frequencies in publications, etc. Using data from the literature, we show that the proposed model provides a good fit to the observed data. Using this new model for cDNA library data, we developed an empirical Bayesian significance test (EBST) for inferring the statistical significance of differential gene expression from discrete data. PMID- 15662206 TI - Recognition of the helical cytokine fold. AB - Cytokines are an important class of proteins responsible for intercellular communication. The helical cytokines have a four-helix bundle fold, and they have remained largely intractable for sequence alignment methods due to their high evolutionary divergence. This paper presents a method that is specifically designed to recognize the helical cytokine fold in preprotein sequences such as full-length cDNA translations or transcripts predicted by gene finding methods. A protein fold is modeled by multiple sequence profiles, each representing a structurally conserved region. Nonstructural profiles are used to represent additional signals found in preprotein sequences. Profiles are connected by loop regions, each of a specified minimum and maximum length. A model for the helical cytokines is created by progressively improving a placement of four amphipathic helices onto training sequences. The sensitivity and specificity of the method are evaluated by a cross-validation procedure, demonstrating that cytokines with no intrafamily sequence similarity can be recognized. The method has been successfully used for the discovery of several new helical cytokines in the human genome. PMID- 15662207 TI - Endothelial function and dysfunction. Part II: Association with cardiovascular risk factors and diseases. A statement by the Working Group on Endothelins and Endothelial Factors of the European Society of Hypertension. AB - Dysfunction of the vascular endothelium is a hallmark of most conditions that are associated with atherosclerosis and is therefore held to be an early feature in atherogenesis. However, the mechanisms by which endothelial dysfunction occurs in smoking, dyslipidaemia, hyperhomocysteinaemia, diabetes mellitus, arterial hypertension, cerebrovascular diseases, coronary artery disease and heart failure are complex and heterogeneous. Recent data indicate that endothelial dysfunction is often associated with erectile dysfunction, which can precede and predict cardiovascular disease in men. This paper will provide a concise overview of the mechanisms causing endothelial dysfunction in the different cardiovascular risk factors and disease conditions, and of the impact of the intervention measures and treatments. PMID- 15662208 TI - Small artery structure and hypertension: adaptive changes and target organ damage. AB - Hypertension is known to be associated with an increase in the wall/lumen ratio (W/L) of the resistance arteries. Growth, eutrophic remodelling and changes in arterial distensibility can all contribute to an increase W/L. Wall stress may stimulate growth, whereas remodelling and/or reduced distensibility may be the result of prolonged contraction. Impaired or overwhelmed pressure-induced myogenic constriction, causing increased wall stress, may explain the small artery growth seen in diabetes and secondary hypertension, respectively. We find that small artery growth is a greater predictor of cardiovascular events than an increased W/L. PMID- 15662209 TI - Effect of resistance training on resting blood pressure: a meta-analysis of randomized controlled trials. AB - OBJECTIVE: To assess the influence of resistance training on resting blood pressure in healthy sedentary adults. METHODS: A comprehensive literature search with the MEDLINE computerized database was conducted and reference lists of published articles and reviews on the topic were checked. Inclusion criteria were as follows: the study involved a randomized, controlled trial; resistance training was the sole intervention; participants were sedentary normotensive and/or hypertensive adults with no other concomitant disease; the article was published in a peer-reviewed journal up to December 2003. We identified nine randomized controlled trials, involving 12 study groups and 341 participants. A standard protocol was used to extract information on sample size, participant characteristics, study design, training method and duration, and study outcomes. Pooled blood pressure estimates were obtained, weighted by either the number of participants in the training group or the inverse of the variance for blood pressure change. RESULTS: The weighted net changes of blood pressure, after adjustment for control observations, averaged -3.2 [95% confidence limits (CL) 7.1 to +0.7]/-3.5 (95% CL -6.1 to -0.9) mmHg when weighted for the number of trained participants, and -6.0 (95% CL -10.4 to -1.6)/ -4.7 (95% CL -8.1 to -1.4) mmHg, when weighted by the reciprocal of the variance for the blood pressure change. CONCLUSIONS: Our results suggest that moderate intensity resistance training is not contraindicated and could become part of the non-pharmacological intervention strategy to prevent and combat high blood pressure. However, additional studies are needed, especially in the hypertensive population. PMID- 15662210 TI - Acute stress and long-lasting blood pressure elevation: a possible cause of established hypertension? PMID- 15662211 TI - Physical activity, physical fitness and the incidence of hypertension. PMID- 15662212 TI - Multiple gene approaches to delineate the role of the renin-angiotensin aldosterone system in nephropathy. PMID- 15662213 TI - The potential advantage of omapatrilat: dual anti-fibrotic and anti-inflammatory effects in one single molecule. PMID- 15662214 TI - Blood pressure in acute stroke: in search of evidence. PMID- 15662215 TI - Sustained blood pressure increase after an acute stressor: the effects of the 11 September 2001 attack on the New York City World Trade Center. AB - BACKGROUND: Effects of catastrophic stress on blood pressure are well documented, but usually few measurements were taken before the event occurred, and the people studied were directly involved or geographically close to the disaster. The impact of the 11 September 2001 (9/11) attacks in New York City had far greater reach, and has been sustained by subsequent events. OBJECTIVE: To test the hypothesis that, after the 9/11 attacks, blood pressures in the population affected would be increased compared with that in both the preceding months and the same period during the previous year. METHODS: We used data available from a current study of blood pressure in four sites in the USA that enabled us to examine them from two perspectives: a mixed (within and between groups) analysis that tested the overall differences in blood pressure before and after 9/11, and a within-subjects model to examine the more focused issue of individual change in blood pressure after 9/11. The blood pressures of 427 hypertensive individuals were telemonitored at four sites. An additional 101 patients had been monitored at two sites during the same period in the previous year. RESULTS: Mean systolic blood pressure was significantly greater during the 2 months after 9/11, across the four sites, compared with that assessed during the previous 2 months (range of observed differences 1.7-3.8 mmHg). At the two sites for which data were available for the same period in the year 2000, there was also a significant effect for the same period during the preceding year for systolic blood pressure. However, at both these sites the effect at 2000 was significantly smaller than the effect at 2001. Blood pressure also generally increased among those individuals in whom monitoring overlapped the 9/11 event. CONCLUSION: The World Trade Center attacks produced a substantial and sustained increase in blood pressure that appears to be independent of seasonal effects, and which has important implications for morbidity and financial burden. The ubiquitous continuing reference to the events in the news reports may contribute to the sustained effects. PMID- 15662216 TI - Hemodynamic after-effects of acute dynamic exercise in sedentary normotensive postmenopausal women. AB - OBJECTIVES: To determine, in sedentary normotensive postmenopausal women, the after-effects of exercise on systemic and regional hemodynamics, and whether changes in total peripheral conductance after exercise relate to changes in brachial artery flow-mediated vasodilation (FMD). METHODS: In 13 sedentary postmenopausal women, the blood pressure (BP), cardiac output, total peripheral resistance and total peripheral conductance, calf vascular resistance and FMD were measured during baseline rest, and again commencing 45 min after treadmill exercise. Fourteen premenopausal women completed the identical protocol to obtain reference values for the after-effects of exercise in healthy females. RESULTS: In postmenopausal women, exercise was followed by falls in systolic BP (P < 0.01) and diastolic BP (P < 0.001). BP did not fall after exercise in premenopausal women. In both groups the cardiac output (P < 0.01) increased and the calf vascular resistance (P < 0.01) and total peripheral resistance (P < 0.05) decreased after exercise, but resistance fell more (P < 0.05) in postmenopausal women. Baseline FMD was greater in premenopausal women (12.1 +/- 1.5 versus 5.3 +/- 1.3%, P < 0.01), and similar before and after exercise, whereas prior exercise nearly doubled the FMD of postmenopausal women (to 9.9 +/- 1.4%, P < 0.01). These increases in FMD correlated with baseline values (r = -0.75, P < 0.01) and with relative changes in total peripheral conductance (r = 0.72, P < 0.02). The latter relationship was absent in premenopausal women (r = -0.29). CONCLUSIONS: In postmenopausal women, acute dynamic exercise elicits sustained increases in FMD that could facilitate post-exercise hypotension in this population. These observations reinforce the concept of exercise as an important non-pharmacological intervention to modify cardiovascular risk in postmenopausal women. PMID- 15662217 TI - Low physical activity as a predictor for antihypertensive drug treatment in 25-64 year-old populations in eastern and south-western Finland. AB - OBJECTIVE: To determine whether low leisure-time physical activity, occupational physical activity and commuting activity independently increase the risk of hypertension when adjusted for most risk factors for hypertension and for different forms of physical activity. DESIGN: Population-based prospective cohort study. SETTING: Eastern and south-western Finland. PARTICIPANTS: Men (n = 5935) and women (n = 6227) aged 25-64 years. MAIN OUTCOME MEASURE: Initiation of free of-charge medication for hypertension during a mean follow-up time of 11.3 years. RESULTS: Men with high leisure-time physical activity had a reduced risk of hypertension when adjustment had been made for age, area and year of survey, education, smoking, alcohol intake, baseline systolic blood pressure (SBP), body mass index (BMI), commuting activity and occupational physical activity [hazard ratio (HR) 0.79; 95% confidence interval (CI) 0.63 to 0.99]. Women with high leisure-time physical activity had a reduced risk of hypertension when adjusted for age, area and time of survey (HR 0.65; 95% CI 0.46 to 0.91). This association was no longer significant when further adjustments were made for other covariates (HR 0.73; 95% CI 0.52 to 1.03). High occupational physical activity reduced the risk of hypertension only among men and women combined when adjustment was made for age, area and time of survey, education, smoking and alcohol intake, in addition to baseline SBP, BMI, commuting activity and leisure-time physical activity (HR 0.83; 95% CI 0.72 to 0.96). Commuting activity was not associated with risk of hypertension in multivariate models. CONCLUSION: High levels of leisure-time physical activity are associated with a reduced risk of hypertension, independently of most common risk factors for hypertension, occupational physical activity and commuting activity. Promoting leisure-time physical activity is essential to prevent hypertension. PMID- 15662218 TI - Peroxisome proliferator-activated receptor-gamma2 Pro12Ala and endothelial nitric oxide synthase-4a/b gene polymorphisms are not associated with hypertension in diabetes mellitus type 2. AB - OBJECTIVE AND DESIGN: The Pro12Ala and the 4a/b polymorphisms of the peroxisome proliferator-activated receptor gamma (PPARgamma) and the endothelial nitric oxide-synthase (eNOS) genes, respectively, have been associated with hypertension in some but not all studies. The purpose of this study was to investigate the association between these polymorphisms and hypertension in patients with diabetes mellitus type 2 (DM2). METHODS: We determined, by polymerase chain reaction (PCR), the Pro12Ala PPARgamma2 and the eNOS 4a/b gene polymorphisms in a total of 395 patients with diabetes mellitus 2 (DM2) (225 men and 170 women) from the LIANCO (Lipid-Analytic-Cologne) study. Hypertension was defined as known or newly diagnosed hypertension according to current national guidelines. Associations were determined using chi-square statistics. The influence of genotype and other parameters on blood pressure was determined by analysis of variance (ANOVA) and multivariate analyses. RESULTS: The genotype frequencies of the Pro12Ala polymorphism were 3% AlaAla, 23% ProAla and 74% ProPro and of the eNOS 4a/b polymorphism 3% a/a, 25% b/a and 72% b/b. There were 65% patients with, and 35% without hypertension. A total of 77% of the patients with hypertension were under pharmacological treatment. The mean systolic and diastolic blood pressure (SBP, DBP) was 148 +/- 22 and 84 +/- 11 mmHg in patients with, and 131 +/- 12 and 79 +/- 8 mmHg in patients without, hypertension. There was no difference in the occurrence of hypertension among ProAla and AlaAla subjects compared with ProPro subjects (P = 0.98). There was also no difference between a allele carriers and non-carriers of the eNOS polymorphism (P = 0.42). There were no differences between men and women in the associations. Analysis of variance did neither identify an influence on systolic or diastolic blood pressure by the presence of the Ala or the a-allele of the respective genotypes nor a significant interaction of the two. CONCLUSIONS: In DM2 the Pro12Ala and 4a/b gene polymorphisms of the PPARgamma2 and eNOS genes, respectively, are not associated with systolic or diastolic blood pressure, either in men or in women. Our results in a large cohort fail to confirm reports of recent studies suggesting an association of lower blood pressure in patients with DM2 and carriers of Pro12Ala polymorphism. PMID- 15662219 TI - Genetic polymorphisms of the renin-angiotensin-aldosterone system and renal insufficiency in essential hypertension. AB - OBJECTIVE: The renin-angiotensin-aldosterone system (RAAS) plays an important role in the control of renal function both in physiological and pathological conditions. The aim of the present study was to evaluate the relation between four genetic polymorphisms of the RAAS and renal insufficiency in a population of patients with essential hypertension living in north-east Italy. DESIGN AND METHODS: Eighty-six hypertensive patients with renal insufficiency and 172 hypertensive patients without renal damage matched for age and hypertension duration to within 2 years were evaluated. Genotyping for insertion/deletion of angiotensin-converting enzyme (ACE I/D), angiotensinogen (AGT) M235T, angiotensin II type 1 receptor (AT1R) A1166C and aldosterone synthase (CYP11B2) -344C/T polymorphisms were performed using polymerase chain reaction, with further restriction analysis when required. RESULTS: Each of the genetic polymorphisms of the RAAS genes was associated with renal failure; the adjusted odds ratios were 1.47 and 1.89 for ACE D allele, assuming a co dominant and a recessive mode of inheritance, respectively; 1.51 for AGT T235 allele assuming a co dominant, and 1.98 assuming a recessive, pattern of inheritance; 1.79 for AT1R C1166 allele considering a dominant pattern; and 3.89 for CYP11B2 -344C allele as a recessive effect. However, CYP11B2 genotypes were not in Hardy-Weinberg equilibrium among controls. The associations AGT TT-AT1R AC and CYP11B2 CC-ACE DD showed a possible positive interaction in the development of renal insufficiency among hypertensive subjects. The association AGT MM-AT1R AA and AGT MM-AT1R AA-CYP11B2 TT or TC combinations were associated with a reduced risk for renal failure. CONCLUSIONS: Our findings suggest that in patients with essential hypertension an unfavorable genetic pattern of RAAS may contribute to the increased risk for the development of renal failure. PMID- 15662220 TI - Angiotensin-converting enzyme inhibition restores endothelial but not medial connexin expression in hypertensive rats. AB - OBJECTIVE AND DESIGN: Remodelling in the media and decreases in connexin (Cx) expression and size of endothelial cells occur in the caudal artery of spontaneously hypertensive rats (SHR). The objective of this study was to determine whether similar changes are found in the aorta and whether effects in both aorta and caudal artery are present in the pre-hypertensive period or can be reversed by antihypertensive treatment. METHODS AND RESULTS: In the aorta of SHR, there was no difference in endothelial cell size although Cxs 37 and 40 were decreased, compared with normotensive Wistar-Kyoto rats. Cxs 37 and 43 were also reduced in the media. These differences were not apparent in pre-hypertensive SHR. Inhibition of angiotensin-converting enzyme (ACE) in SHR decreased blood pressure and restored Cx expression in the endothelium of both aorta and caudal artery. The decreased endothelial cell size in the caudal artery or the reduced Cxs in the media of the aorta of SHR were unaffected by ACE inhibition. CONCLUSION: We conclude that cellular coupling is reduced in the endothelium of arteries of SHR, but this can be restored by inhibition of the renin-angiotensin system. Decreased cellular coupling in the media or decreased endothelial size in SHR were not reversed by this antihypertensive treatment. PMID- 15662221 TI - Position of Src tyrosine kinases in the interaction between angiotensin II and endothelin in in vivo vascular protein synthesis. AB - OBJECTIVES: Endothelin is a necessary intermediate in the trophic action of angiotensin II during hypertension-induced resistance artery remodeling in vivo. Since Src tyrosine kinases can be activated by both agonists, we studied their role in the trophic action of angiotensin II, endothelin and their interaction in rat small mesenteric arteries. METHODS AND RESULTS: Twenty-six hour infusion of high-dose angiotensin II (400 ng/kg per min) or endothelin (5 pmol/kg per min) via osmotic pumps significantly enhanced vascular protein synthesis in vivo. When angiotensin II was used as the trophic stimulus, treatment with a Src tyrosine kinase inhibitor (PP2, 0.5 mg/kg, starting at 21 h of the 26-h stimulation) produced a significant attenuation of extracellular regulated kinase 1 (ERK 1) phosphorylation and of protein synthesis. However, PP2 administered at 21 h or throughout the 26-h infusion did not abrogate the elevation of protein synthesis induced by endothelin. Moreover, endothelin did not enhance the phosphorylation of ERK 1/2 in small mesenteric arteries. We confirmed that angiotensin II stimulated the expression of prepro-endothelin mRNA in small mesenteric arteries in a Src-dependent manner, as the response was inhibited by PP2. To support the specific inhibitory activity of PP2 on Src tyrosine kinases in vivo, angiotensin II-induced phosphorylation of cortactin, a Src-specific substrate, was inhibited by PP2. CONCLUSION: Src tyrosine kinases represent an important signaling element in angiotensin II-induced endothelin production in small arteries in vivo. However, Src tyrosine kinases did not appear to contribute to the trophic signaling of endothelin, suggesting that they lie upstream of endothelin in the angiotensin II-endothelin-protein synthesis cascade. PMID- 15662222 TI - Role of L-type calcium channel blocking in epidermal growth factor receptor independent activation of extracellular signal regulated kinase 1/2. AB - OBJECTIVE: Vascular smooth muscle cell (VSMC) differentiation, growth and survival, key events in the development of cardiovascular diseases, are under the control of signaling enzymes including extracellular signal regulated kinase 1/2 (ERK 1/2), Akt and epidermal growth factor receptor (EGFR) activation. EGFR trans activation is known to mediate thrombin- or angiotensin II (AII)-stimulated ERK 1/2 activation. However, our laboratory has demonstrated, in thrombin-stimulated VSMC, that the prevention of intracellular Ca2+ elevation ([Ca2+]i) by BAPTA-AM pretreatment unveiled EGFR-independent ERK 1/2 activation. Since calcium channel blockers (CCBs) also impair agonist-induced [Ca2+]i elevation, we investigated whether EGFR-independent ERK 1/2 activation could occur in VSMCs treated by CCBs such as amlodipine, isradipine and verapamil, and examined the possible role of Akt. METHODS: Cultured VSMCs were pretreated or not with CCBs and with various inhibitors of the signaling pathways under study, prior to stimulation by thrombin or AII, and the phosphorylation/activation status of EGFR, Akt and ERK 1/2 was determined by Western blotting using phospho-specific antibodies. RESULTS AND CONCLUSION: Unlike BAPTA, CCBs did not impair stimulus-induced EGFR trans activation, hence ERK1/2 phosphorylation. However, when EGFR kinase was inhibited, CCBs and BAPTA dose-dependently protected stimulus-induced ERK1/2 phosphorylation. The effect of amlodipine could not be mimicked by its R+ enantiomer, which is devoid of CCB activity, suggesting that the effects of CCBs were accounted for by their L-type Ca2+ channel-blocking property. Altogether, our results indicated that in G-protein-coupled receptor (GPCR)-stimulated VSMCs, the prevention of [Ca2+]i elevation by CCBs unmasked an EGFR kinase-independent phosphorylation of ERK 1/2. Since EGFR kinase inhibitors are supposed to be efficient in the treatment of some cancers, such a mechanism might be clinically relevant in hypertensive patients with cancer. PMID- 15662224 TI - Induction of tissue kallikrein in human carotid atheroma does not lead to kallikrein-kinins pathway activation. AB - OBJECTIVE: The impairment of the tissue kallikrein-kinin system (KKS) may result in atheroma development. To determine the involvement of KKS in pathophysiology of human atherosclerosis, we examined the expression of all components of this system as well as angiotensinogen (another tissue kallikrein (TK) substrate), at messenger ribonucleic acid (mRNA) and protein levels in the human carotid artery with and without atheroma. METHODS: mRNA levels were compared with semi quantitative reverse transcriptase-polymerase chain reaction (RT-PCR) between atheroma plaque and intact tissue obtained during carotid endarterectomy in 15 patients. The cellular localization of the transcripts and proteins was analyzed with in situ hybridization and immunohistochemistry. TK activity was measured using chromogenic substrate. RESULTS: The kininogen mRNA was not detected in carotid wall. The TK mRNA was increased four-fold and TK activity 23-fold in atheroma plaque compared with intact tissue. No difference was observed for B1, B2 receptors, kallistatin, angiotensinogen and protein-kinase G type 1alpha (PK G) mRNAs. The TK and angiotensinogen transcripts as well as kininogen and angiotensinogen proteins were present in both intimal and medial cells. The kininogen immunoreactivity was weaker in atheroma. CONCLUSIONS: All KKS components were synthesized in arterial wall except kininogen probably coming from plasma. The absence of PK-G mRNA down-regulation in atheroma suggests that the kallikrein induction does not lead to KKS activation. PMID- 15662223 TI - Vascular responses to IGF-I and insulin are impaired in aortae of hypertensive rats. AB - OBJECTIVE: Insulin-like growth factor-I (IGF-I) and insulin are important vasoactive peptides but little is known about their effects in hypertension. DESIGN: We compared the responses of stroke-prone spontaneously hypertensive rats (SHRSP) and Wistar-Kyoto (WKY) rat aortae to IGF-I and insulin. METHODS: Aortae were removed from WKY and SHRSP, cut into 2-3 mm rings, and contractile responses to phenylephrine and endothelin-1 studied in organ chambers in the presence of vehicle, IGF-I (0.1 micromol/l) or insulin (0.1 micromol/l). In addition, the effects of nitric oxide synthase (NOS) inhibition, phosphatidylinositol 3-kinase (PI3-kinase) inhibition and superoxide scavenging on these responses were investigated. RESULTS: Incubation with IGF-I and insulin caused attenuation of phenylephrine-induced and endothelin-1-induced vasoconstriction in arteries from normotensive but not hypertensive animals. In the arteries from WKY rats, co incubation with either wortmannin or LY294002, inhibitors of PI3-kinase, attenuated the effect of IGF-I. The vasorelaxant effect of IGF-I was also abolished by removal of the endothelium or addition of the NOS inhibitor, N-nitro L-arginine methyl ester (L-NAME). Co-incubation with tiron, a superoxide scavenger, suggested that the attenuation of IGF-I vasodilation in SHRSP arteries was not due to excess superoxide production. CONCLUSION: In WKY, IGF-I/insulin attenuate phenylephrine-mediated constrictions via PI3-kinase/nitric oxide pathways. In contrast, in SHRSP these pathways are dysfunctional and IGF-I has little effect on vascular responses. PMID- 15662225 TI - Enhanced angiotensin II-mediated effects in fibroblasts of patients with familial hypercholesterolemia. AB - OBJECTIVE: Familial hypercholesterolemia (FH) is characterized by a high incidence of coronary heart disease. Evidence suggests an important role for angiotensin II (AngII) in the fibrotic response to tissue injury, and in promoting myocardial hypertrophy via paracrine mechanisms mediated by fibroblasts. We sought to determine whether AngII promotes proliferative and pro atherogenic responses in FH patients. METHODS: We used primary fibroblasts -- from five patients with heterozygous FH and five control subjects (C) -- to study AngII-induced cell growth, intracellular calcium fluxes, and expression/release of matrix components and pro-inflammatory peptides [transforming growth factor beta1 (TGFbeta1) and endothelin-1 (ET-1)] and metalloproteinases involved in plaque remodeling and vulnerability. RESULTS: AngII stimulated cell replication (5.1 +/- 0.03 versus 3.2 +/- 0.04 cells/50 cells per well, P < 0.001), and induced a larger increase in intracellular calcium content in FH cells than in C cells, in a dose-dependent fashion (mean difference = 76 nmol/l, P < 0.001). Similarly, TGFbeta1 and ET-1 expression and release were potentiated (after 24-h incubation with 1 micromol/l AngII: TGFbeta1 was 190 +/- 12 in C and 376 +/- 9 pg/ml per 10(6) cells in FH, and ET-1 was 93 +/- 5 in C and 192 +/- 7 pmol/ml per 10(6) cells in FH; P < 0.001 for both). AngII-induced release of the metalloproteinases MMP-1 and MMP-2 was also increased in FH versus C cells (0.52 +/- 0.04 versus 0.36 +/- 0.05 and 24 +/- 4 versus 13 +/- 3 ng/mg protein with 1 micromol/l AngII). These enhanced responses were likely due to an increased angiotensin receptor 1 (AT1) expression in cells from FH patients induced by AngII, and were prevented by pretreating cells with the selective AT1 antagonist irbesartan. CONCLUSIONS: These findings show that some AngII-mediated pathways are enhanced in FH subjects irrespective of the presence of low-density lipoprotein (LDL), thus contributing to the development and progression of atherosclerosis in these patients. PMID- 15662227 TI - Effect of insulin and angiotensin II receptor subtype-1 antagonist on myocardial remodelling in rats with insulin-dependent diabetes mellitus. AB - OBJECTIVES: To assess the role of insulin or an angiotensin II receptor antagonist (losartan), or both, in preventing cardiomyocyte damage in rats suffering from insulin-dependent diabetes mellitus (IDDM), and to correlate it with insulin receptor modulation at the cardiomyocyte, coronary endothelium and skeletal muscle cell level. DESIGN: Animals were divided into groups of normal rats, diabetic rats, and diabetic rats given insulin, each subdivided into a control group and an experimental group treated with losartan. METHODS: The animals were killed 1 month after enrollment to the study. Perfusion of the heart with iodine-125-labelled insulin was carried out for all the groups and the binding kinetics of insulin to its receptors on the coronary endothelial cells and the cardiomyocytes were determined using a physical/mathematical model. In addition, tissue samples from the heart and intercostal skeletal muscle were snap frozen and used for histological, indirect immunofluorescence and western blot analysis. RESULTS: Cardiac muscle from diabetic animals exhibited diffuse cardiomyopathic changes consisting of widespread vacuolation, loss of striation and cellular hypertrophy, which were reduced and even prevented by treatment with insulin and losartan. In addition, losartan seemed to mediate the upregulation of insulin receptor density on cardiomyocytes and skeletal muscle, and increase insulin receptor affinity at the coronary endothelial site. Finally, treatment with losartan induced a significant decrease in glucose concentrations in the diabetic group compared with the appropriate controls. CONCLUSIONS: Addition of losartan to the standard insulin treatment in non-hypertensive animals with IDDM offers new benefits concerning cardiac protection and prevention of damage. This may be attributed, in part, to insulin receptor density and sensitization. PMID- 15662226 TI - Magnesium supplementation prevents angiotensin II-induced myocardial damage and CTGF overexpression. AB - OBJECTIVES AND DESIGN: Magnesium deficiency promotes vasoconstriction and myocardial damage. Recent studies provide evidence that Ang II mobilizes intracellular Mg through AT1 receptor-mediated pathways. We tested the hypothesis of whether magnesium supplementation prevents Ang II-induced myocardial damage and induction of the profibrotic connective tissue growth factor (CTGF). METHODS: Four-week-old double transgenic rats harboring human renin and angiotensinogen genes (dTGR) were given dietary magnesium supplementation (0.6%) for 3 weeks. Control dTGR and normotensive Sprague-Dawley (SD) rats received normal diet (Mg 0.2%). Histopathological, immunohistochemical and mRNA analysis were used to detect the treatment-related effects of dietary magnesium in dTGR. RESULTS: Magnesium (Mg) supplementation decreased blood pressure, ameliorated cardiac hypertrophy, protected against the development of Ang II-induced myocardial damage and increased serum ionized Mg2+ concentration (all variables P < 0.05). There was no difference in serum ionized Mg2+ concentration between dTGR and SD rats. Myocardial connective tissue growth factor (CTGF) mRNA and protein expressions were increased by 300% in dTGR (P < 0.05), especially in areas with myocardial infarctions and vascular inflammation. Magnesium supplementation prevented Ang II-induced myocardial CTGF overexpression (P < 0.05). Magnesium supplementation also improved the therapeutic effects of the calcineurin inhibitor tacrolimus, which produced marked hypomagnesemia when given as monotherapy. CONCLUSION: Our findings suggest a salutary effect for magnesium supplementation in the treatment of Ang II-induced myocardial complications. PMID- 15662228 TI - ACE inhibitor and angiotensin II type 1 receptor blocker differently regulate ventricular fibrosis in hypertensive diastolic heart failure. AB - BACKGROUND: Promoted myocardial stiffening has a crucial role in the transition to overt diastolic heart failure (DHF) in hypertensive hearts and is attributed to progressive ventricular fibrosis. Previous studies revealed the effects of an angiotensin II type 1 receptor blocker (ARB) and an angiotensin-converting enzyme inhibitor (ACEI) on the synthesis and degradation of collagens in the other phenotype of heart failure, systolic heart failure, which has a different pathophysiology; however, little is known about their effects in DHF. OBJECTIVE: To investigate effects of an ACEI and an ARB on the regulatory system of ventricular fibrosis in hypertensive DHF. DESIGN AND METHODS: Dahl salt-sensitive rats fed a diet containing 8% NaCl from age 7 weeks (DHF model) were divided into three groups: six untreated rats, six rats treated with a subdepressor dose of an ARB, candesartan cilexetil (1 mg/kg per day), from age 8 weeks, and six rats treated with a subdepress or dose of an ACEI, temocapril hydrochloride (0.2 mg/kg per day), from age 8 weeks. Six Dahl salt-sensitive rats fed on normal chow served as controls. Data were collected when animals were aged 20 weeks. RESULTS: The administration of an ARB or an ACEI inhibited ventricular fibrosis to the same degree. The ACEI decreased the level of type I collagen mRNA, but the decrease was less than that induced by the ARB. The difference in collagen synthesis was probably cancelled out by that in degradation: both in-vitro and in situ zymography showed that gelatinase activity was greater in the rats treated with the ACEI than in those treated with the ARB. CONCLUSIONS: An ARB and an ACEI inhibited ventricular fibrosis through different mechanisms in hypertensive DHF. PMID- 15662229 TI - Dual angiotensin-converting enzyme/neutral endopeptidase inhibition on cardiac and renal fibrosis and inflammation in DOCA-salt hypertensive rats. AB - OBJECTIVES: The relative roles of angiotensin-converting enzyme (ACE) and neutral endopeptidase (NEP) inhibition on cardiac and renal fibrosis in deoxycorticosterone acetate (DOCA)-salt hypertensive rats were studied. METHODS: The ACE/NEP inhibitor omapatrilat (40 mg/kg per day), the ACE inhibitor enalapril (10 mg/kg per day) and the NEP inhibitor CGS 25462(100 mg/kg per day) were administrated for 3 weeks to DOCA rats. Collagen was stained with Sirius red, and mediators of inflammation were identified by immunolabeling (vascular cell adhesion molecule, nuclear factor-kappaB, infiltrating ED-1-positive macrophages and monocyte chemotactic protein-1) or by western blot (platelet-endothelial cell adhesion molecule-1). RESULTS: Elevated systolic blood pressure of DOCA rats was significantly reduced (P < 0.05) by omapatrilat and CGS 25462. Omapatrilat and CGS 25462 significantly (P < 0.05) decreased interstitial collagen density in the left ventricle of DOCA rats compared with untreated DOCA rats. Enalapril only decreased the subepicardial collagen of DOCA rats. Omapatrilat significantly (P < 0.05) decreased renal mesangial collagen deposition in DOCA rats. Cardiac and renal expression of surface adhesion molecules, nuclear factor-kappaB, monocyte chemotactic protein and ED-1-positive cells were decreased in omapatrilat-treated DOCA rats compared with untreated DOCA rats. Enalapril and CGS 25462 did not alter mesangial collagen of DOCA rats. CONCLUSIONS: Dual ACE/NEP inhibition was more effective than ACE or NEP inhibition in decreasing inflammatory mediators, and improving cardiac and renal fibrosis. This suggests a role for NEP inhibition added to blockade of the renin-angiotensin system that may explain the greater efficacy of omapatrilat. PMID- 15662230 TI - Adrenocorticotropic hormone is produced in the ventricle of patients with essential hypertension. AB - OBJECTIVE: Aldosterone is produced in the ventricle of patients with hypertension. The present study was designed to examine whether adrenocorticotropic hormone (ACTH) and cortisol are also produced from the heart in patients with essential hypertension. METHODS: The study population consisted of 57 patients with essential hypertension and 28 control subjects. Plasma levels of ACTH, aldosterone, and cortisol were measured in the aortic root, the anterior interventricular vein and the coronary sinus during cardiac catheterization. RESULTS: The plasma levels of ACTH were significantly higher at the anterior interventricular vein and coronary sinus than at the aortic root (12.7 +/- 1.0 versus 10.7 +/- 0.9 pmol/l, P < 0.001; and 12.3 +/- 1.0 versus 10.7 +/- 0.9 pmol/l, P < 0.001, respectively) in the hypertension group, whereas there were no significant differences in the levels among these sites in the control group. The plasma levels of aldosterone were significantly higher at the anterior interventricular vein and the coronary sinus than at the aortic root (261.7 +/- 16.4 versus 239.1 +/- 15.1 pmol/l, P < 0.001; and 258.8 +/- 17.0 versus 239.1 +/- 15.1 pmol/l, P < 0.01, respectively) in the hypertension group, whereas there were no significant differences in the levels among these sites in the control group. CONCLUSIONS: ACTH as well as aldosterone is produced, but cortisol is not produced, from the ventricle of patients with essential hypertension. PMID- 15662232 TI - The combination of vitamin C and grape-seed polyphenols increases blood pressure: a randomized, double-blind, placebo-controlled trial. AB - BACKGROUND: There is growing evidence that oxidative stress contributes to the pathogenesis of hypertension and endothelial dysfunction. Thus, dietary antioxidants may beneficially influence blood pressure (BP) and endothelial function by reducing oxidative stress. OBJECTIVE: To determine if vitamin C and polyphenols, alone or in combination, can lower BP, improve endothelial function and reduce oxidative stress in hypertensive individuals. DESIGN: A total of 69 treated hypertensive individuals with a mean 24-h ambulatory systolic blood pressure > or = 125 mmHg participated in a randomized, double-blind, placebo controlled, factorial trial. Following a 3-week washout, participants received 500 mg/day vitamin C, 1000 mg/day grape-seed polyphenols, both vitamin C and polyphenols, or neither for 6 weeks. At baseline and post-intervention, 24-h ambulatory BP, ultrasound-assessed endothelium-dependent and -independent vasodilation of the brachial artery, and markers of oxidative damage, (plasma and urinary F2-isoprostanes, oxidized low-density lipoproteins and plasma tocopherols), were measured. RESULTS: A significant interaction between grape seed and vitamin C treatments for effects on BP was observed. Vitamin C alone reduced systolic BP versus placebo (-1.8 +/- 0.8 mmHg, P = 0.03), while polyphenols did not (-1.3 +/- 0.8 mmHg, P = 0.12). However, treatment with the combination of vitamin C and polyphenols increased systolic BP (4.8 +/- 0.9 mmHg versus placebo; 6.6 +/- 0.8 mmHg versus vitamin C; 6.1 +/- 0.9 mmHg versus polyphenols mmHg, each P < 0.0001) and diastolic BP (2.7 +/- 0.6 mmHg, P < 0.0001 versus placebo; 1.5 +/- 0.6 mmHg, P = 0.016 versus vitamin C; 3.2 +/- 0.7 mmHg, P < 0.0001 versus polyphenols). Endothelium-dependent and -independent vasodilation, and markers of oxidative damage were not significantly altered. CONCLUSION: Although the mechanism remains to be elucidated, these results suggest caution for hypertensive subjects taking supplements containing combinations of vitamin C and polyphenols. PMID- 15662231 TI - Aliskiren, a novel, orally effective renin inhibitor, lowers blood pressure in marmosets and spontaneously hypertensive rats. AB - OBJECTIVES: Aliskiren is a new renin inhibitor of a novel structural class that has recently been shown to be efficacious in hypertensive patients after once daily oral dosing. We report the results of animal experiments performed in marmosets and rats in order to characterize aliskiren before its recent investigation in humans. METHODS: The effects of aliskiren were investigated in sodium-depleted marmosets (oral dosing) and in spontaneously hypertensive rats (dosing via subcutaneous osmotic minipumps). Blood pressure (BP) and heart rate were measured by radiotelemetry. RESULTS: In sodium-depleted marmosets, single oral doses of aliskiren (1-30 mg/kg) dose-dependently lowered BP. At a dose of 3 mg/kg, peak effects were observed 1 h after dosing (-30 +/- 4 mmHg, n = 6) and the response persisted for more than 12 h. A single oral dose of 3 mg/kg aliskiren was more effective than the same dose of either remikiren or zankiren, two orally active renin inhibitors previously tested in humans. Aliskiren (10 mg/kg) was at least as effective as equal doses of the AT1-receptor blocker valsartan or the angiotensin-converting enzyme inhibitor benazepril. In spontaneously hypertensive rats, aliskiren dose-dependently (10-100 mg/kg per day) decreased BP. Aliskiren also potentiated the antihypertensive effects of low doses of valsartan or benazeprilat (1 or 3 mg/kg per day). CONCLUSIONS: Aliskiren is an orally effective, long-lasting renin inhibitor that shows antihypertensive efficacy in animals superior to previous renin inhibitors and at least equivalent to angiotensin-converting enzyme inhibitors and AT1-receptor blockers. Aliskiren may therefore represent an effective, novel approach to the treatment of hypertension and related disorders, alone or in combination with other antihypertensive agents. PMID- 15662233 TI - Effects of candesartan cilexetil and enalapril on inflammatory markers of atherosclerosis in hypertensive patients with non-insulin-dependent diabetes mellitus. AB - OBJECTIVE: Circulating adhesion molecules may have a prognostic significance as markers of endothelial damage. Drugs which inhibit the renin-angiotensin system may be effective in reducing circulating or tissue adhesion molecules, albeit data available are scarce. The aim of the study was to investigate the effects of an angiotensin-converting enzyme (ACE) inhibitor, enalapril and a highly selective angiotensin receptor blocker, candesartan cilexetil, on circulating adhesion molecules in a large sample of patients with non-insulin-dependent diabetes mellitus (NIDDM). The study was comparative, multicenter, randomized and double blind, with two parallel groups. PATIENTS AND METHODS: NIDDM patients with a diagnosis of mild (grade 1) essential hypertension were included in the study, at the end of a 2-week placebo run-in period. The primary end-point of the study was to evaluate changes of intercellular adhesion molecule-1 (ICAM-1) plasma levels during treatment. The secondary end-points were: changes in vascular cells adhesion molecule-1 (VCAM-1), von Willebrand factor (vWF), fibrinogen and plasminogen activator inhibitor-1 (PAI-1) circulating levels and of urinary albumin excretion rate (AER) as well; 129 patients were randomized: 66 in the candesartan group and 63 in the enalapril group, 118 of them completed the scheduled 24-week treatment period. RESULTS: Candesartan and enalapril equally reduced circulating level of ICAM-1 and exerted comparable effects on changes of other adhesion molecules and coagulation factors. A similar blood pressure lowering effect was observed with the two drugs (candesartan: from 148/90 +/- 11/8 to 132/82 +/- 12/7 mmHg, P < 0.01, enalapril: from 148/91 +/- 12/8 to 131/85 +/- 14/6 mmHg, P < 0.01). Candesartan was more effective than enalapril in the reduction of albuminuria (P < 0.05 between treatments), although urinary protein excretion can be considered normal in the majority of patients. The two drugs were comparable in terms of adverse events reported. CONCLUSION: Candesartan and enalapril showed similar effects on blood pressure and on circulating adhesion molecules. In this study urinary protein excretion was reduced more by candesartan. PMID- 15662234 TI - Controlled-release nifedipine and candesartan low-dose combination therapy in patients with essential hypertension: the NICE Combi (Nifedipine and Candesartan Combination) Study. AB - OBJECTIVE: To compare the clinical efficacy of low-dose controlled-release (CR) nifedipine (20 mg/day) plus candesartan (8 mg/day) combination therapy with that of up-titrated candesartan (12 mg/day) monotherapy. DESIGN: Randomized, double blind study. SETTING: Outpatient study. PATIENTS AND PARTICIPANTS: Patients with essential hypertension, who did not achieve their target blood pressure with baseline treatment of candesartan 8 mg/day for 8 weeks. MAIN OUTCOME MEASURES: Blood pressure, pulse pressure, urinary microalbumin excretion. RESULTS: Blood pressure was significantly reduced in both groups (P < 0.05), but the reduction was significantly greater in the combination therapy group (12.1 +/- 1.4/8.7 +/- 0.9 mmHg) than in the up-titrated monotherapy group (4.1 +/- 1.4/4.6 +/- 0.9 mmHg) (P < 0.0001). The reduction in pulse pressure was significantly greater in the combination therapy group (3.3 +/- 1.2 mmHg) than in the up-titrated monotherapy group (0.7 +/- 1.2 mmHg) (P = 0.0031). Urinary microalbumin excretion decreased significantly in the combination therapy group (from 61.9 to 40.5 mg/g creatinine; P < 0.05), but not in the up-titrated monotherapy group. CONCLUSIONS: These findings suggest that the low-dose combination therapy of nifedipine CR and candesartan is superior to the up-titrated monotherapy of candesartan in terms of blood pressure control and renal protection in patients with essential hypertension. PMID- 15662235 TI - COSSACS (Continue or Stop post-Stroke Antihypertensives Collaborative Study): rationale and design. AB - RATIONALE: Up to 40% of acute stroke patients are already taking antihypertensive therapy on hospital admission, and most will develop elevated blood pressure levels as an acute complication of the stroke. However, no clear data exist as to whether antihypertensive therapy should be continued or discontinued in the acute situation. Surveys of clinical practice reveal significant physician variability and no clear guidelines exist. OBJECTIVES: The primary aim of the Continue or Stop post-Stroke Antihypertensives Collaborative Study (COSSACS) is to assess whether existing antihypertensive therapy should be continued or discontinued within the first 24 h for the first 2 weeks following acute ischaemic and haemorrhagic stroke onset. DESIGN: COSSACS is a multi-centre, prospective, randomized, open, blinded-endpoint study, in which patients on pre-existing antihypertensive therapy, admitted to hospital within 24 h of onset of suspected stroke, and within 36 h of their last dose of antihypertensive medication, are randomized to continue or stop current antihypertensive therapy. SETTING: Acute Stroke Units/ Medical Units of at least 25 UK Teaching and District General Hospitals. PATIENTS: The study will involve 2900 patients with suspected stroke without specific indication to continue or stop their antihypertensive medication in the opinion of their treating clinician. STUDY OUTCOMES: The primary outcome for COSSACS is the proportion of patients who are dead or dependent (defined by a modified Rankin score > 2) at 14 days post-stroke. Secondary outcomes include blood pressure changes, and neurological and functional status at 2 weeks and 6 months post-ictus. PMID- 15662236 TI - Prorenin cryoactivation as a possible cause of normal renin levels in patients with primary aldosteronism. PMID- 15662237 TI - Serial echocardiographic assessment of left ventricular mass. PMID- 15662240 TI - Future directions in imaging of neck and brain vessels. PMID- 15662241 TI - Utility of source images of three-dimensional time-of-flight magnetic resonance angiography in the diagnosis of indirect carotid-cavernous sinus fistulas. AB - BACKGROUND: We sought to assess the relative contribution of magnetic resonance imaging (MRI), maximum intensity projection (MIP), and source images of three dimensional (3D) time-of-flight (TOF) magnetic resonance angiography (MRA) to the diagnosis of indirect (dural) carotid-cavernous sinus fistulas (CCFs). METHODS: MRI and 3D TOF MRA were obtained in eight consecutive patients with indirect CCFs confirmed by conventional catheter angiography. Two radiologists masked to the angiographic results reviewed images retrospectively to evaluate the efficacy of MRI and 3D TOF MRA source and MIP images in the diagnosis of CCF. RESULTS: MRI disclosed CCF in five of eight cases; MIP images of TOF MRA disclosed CCF in four cases; source images of TOF MRA disclosed all eight CCF cases. CONCLUSIONS: The MRA source images are indispensable for a confirmatory diagnosis of indirect (dural) CCF. Underdiagnosis may occur by relying on MRI or 3D TOF MIP images alone. PMID- 15662243 TI - Optic nerve calcification after trauma. AB - Two patients with remote histories of severe optic nerve trauma displayed profound intraorbital optic nerve calcification on imaging studies. The presumed mechanism is optic nerve hemorrhage. Although calcification is known to occur long after brain hemorrhage, no comparable cases have been previously reported. PMID- 15662242 TI - Chronic myokymia limited to the eyelid is a benign condition. AB - BACKGROUND: Eyelid myokymia, unlike myokymia of the other facial muscles, is assumed to be a benign, self-limited disorder. However, no systematic follow-up study has been performed on patients with chronic, isolated eyelid myokymia to verify its benign nature. METHODS: Retrospective single-institution chart review of 15 patients examined between 1983 and 2002 with a diagnosis of isolated eyelid myokymia who have had at least 12 months of follow-up. RESULTS: In all patients, symptoms began as unilateral, weekly or biweekly, intermittent eyelid spasms, and progressed to daily spasms over several months. The mean duration of symptoms at first examination was 91 months (range 2.5 months to 20 years). In no patient was the myokymia the first manifestation of a neurologic disease, although one patient progressed to ipsilateral hemifacial spasm. Thirteen patients (86.7%) underwent neuroimaging that gave negative results. The myokymia resolved spontaneously in four patients. Of the remaining 11 patients, eight were treated with botulinum toxin injection at regular intervals, with most reporting an improvement in symptoms. CONCLUSION: Chronic isolated eyelid myokymia is a benign condition. It tends not to progress to other facial movement disorders or to be associated with other neurologic disease. It responds well to treatment with botulinum toxin. PMID- 15662244 TI - Magnetic resonance imaging of choroidal inflammation in Vogt-Koyanagi-Harada disease. AB - Acute binocular visual loss, photophobia, headache, and pulsatile tinnitus developed in a 51-year-old woman. Ophthalmologic examination showed bilateral optic disc edema with peripapillary nerve fiber layer hemorrhages. Lumbar puncture disclosed a monocytic pleocytosis. A diagnosis of Vogt-Koyanagi-Harada disease was made. Magnetic resonance imaging showed striking enhancement and thickening of the posterior ocular wall. A macular star figure appeared several days after prednisone treatment was begun. Laboratory evaluation was entirely negative. Within weeks, the clinical manifestations had resolved except for retinal striae. This is the third report of the magnetic resonance imaging visualization of choroidal inflammation in Vogt-Koyanagi-Harada disease and shows the imaging abnormalities in finer detail than earlier reports. PMID- 15662245 TI - Progression from anomalous optic discs to visible optic disc drusen. AB - At age 5, a patient underwent fundus photography that disclosed elevated optic discs without drusen. A head computed tomography did not show optic nerve calcification. At age 9, no disc drusen were evident by ophthalmoscopy, but a CT now showed optic nerve calcification. At age 12, optic disc drusen were faintly evident on photographs; visual fields showed blind spot enlargement OD and an arcuate defect OS. At age 21, he had numerous discrete disc drusen in both eyes, disc pallor, and slight progression of the visual field defects. This case documents the progression from anomalous optic discs to ophthalmoscopically visible optic nerve drusen over a 16-year period. PMID- 15662246 TI - Cervicocranial arterial dissections. AB - Cervicocranial dissections are an increasingly recognized cause of stroke in the young. When the dissections narrow the vascular lumen, they often alter blood flow enough to cause transient ischemic attacks in the brain. Alterations in the endothelium activate the coagulation cascade, leading to the formation of intramural clot that may embolize distally to cause brain infarction. Pain and neuro-ophthalmic symptoms and signs are common manifestations. PMID- 15662247 TI - Computed tomography and magnetic resonance angiography in cervicocranial vascular disease. AB - Although catheter angiography, or digital subtraction angiography (DSA), is still regarded as the gold standard for imaging of cervicocranial vascular disease, its morbidity, cost, and time-consuming features have prompted the development of noninvasive techniques based on computed tomography (CT) and magnetic resonance imaging. With the advent of powerful software, CT and magnetic resonance angiography are complementing and, in some cases, even replacing DSA in the diagnostic evaluation of carotid atherostenosis, unruptured aneurysms, dissections, stroke, penetrating trauma to the neck, and dural venous sinus occlusive disease. They offer advantages over DSA not only in reduced morbidity and time-saving but also in assessment of brain parenchyma, quantitative perfusion, and abnormalities of vessel walls. In the evaluation of blunt neck injuries and intracranial vascular malformations, fistulas, and vasculitis, CT and magnetic resonance angiography still do not provide as much information as DSA. PMID- 15662248 TI - Magnetic resonance imaging of suspected cervicocranial arterial dissections. AB - The authors propose that the optimal screening protocol for evaluation of suspected cervicocranial arterial dissections is magnetic resonance imaging (MRI) that includes three components: 1) contrast-enhanced three-dimensional time-of flight magnetic resonance angiography (MRA) through the superior mediastinum, neck, and skull base; 2) three-dimensional multiple overlapping thin-section acquisition MRA of the skull base and Circle of Willis region; and 3) axial non contrast, non-fat-suppressed T1-weighted, fat-suppressed T1-weighted, and T2 weighted spin-echo MRI from the level of the aortic arch through the level of the circle of Willis. MRA permits visualization of vascular luminal narrowing or obliteration, which can suggest vascular dissection but can also be caused by congenital variation, dysplasia, intraluminal thrombus, vasospasm, or extramural compression by tumor. By directly visualizing the blood vessel wall, axial T1 weighted and T2-weighted spin-echo MRI can identify the intramural hemorrhage of vascular dissection. This protocol is designed to maximize the sensitivity of a noninvasive technique and may eliminate the need for conventional endovascular angiography. PMID- 15662250 TI - Annual meeting of the Association for Research in Vision and Ophthalmology, Fort Lauderdale, Florida, April 25-29, 2004. PMID- 15662251 TI - The 56th annual meeting of the American Academy of Neurology, San Francisco, California, April 24-30, 2004. PMID- 15662252 TI - Neuro-ophthalmology at the Mayo Clinic. PMID- 15662253 TI - Intracranial hypertension associated with transverse myelitis. PMID- 15662254 TI - Persistent visual loss from neurotrophic corneal ulceration after dorsolateral medullary infarction (Wallenberg syndrome). PMID- 15662255 TI - Isolated granulomatous uveitis presenting twenty-two years before multiple sclerosis. PMID- 15662285 TI - The provisional approval step. PMID- 15662286 TI - Regression of left ventricular hypertrophy with moexipril, an angiotensin converting enzyme inhibitor, in hypertensive patients. AB - Left ventricular hypertrophy (LVH) is a common complication of essential hypertension and an independent risk factor for the development of cardiovascular disease. Therefore, antihypertensive treatment should decrease blood pressure (BP) and reverse LVH. However, antihypertensive drugs have been shown to have different effects on LVH despite similar effects on BP reduction. Although lowering BP produces a beneficial effect on LVH per se, meta-analyses of clinical trials have indicated that angiotensin-converting enzyme (ACE) inhibitors decrease left ventricular mass (LVM) to a greater extent than do some other antihypertensives. The aim of this study was to evaluate the effect of a 24-week treatment with the ACE inhibitor moexipril (15 mg once daily) on the regression of LVH in hypertensive patients. This was a multicenter, international, single blind, single-group, nonrandomized study. After a wash-out placebo period of 2 weeks, 15 mg moexipril once daily was administered for 24 weeks followed by a 2 week follow-up placebo period. Subjects with mild to moderate essential hypertension were screened; those with LVH [defined as an LVM indexed for body surface area (LVMIs) >111 g/m in men and LVMIs >106 g/m in women] were eligible to participate in this study. Echocardiograms were recorded on videotape and sent to a centralized laboratory for reading by 2 independent experts blinded for treatment, center, and visit; the mean values of these readings were calculated and used for analysis. Valid echocardiographic data were obtained from 72 patients (50 males, 22 females) with a mean age of 49 +/- 11 years. Analysis showed significant decrease of LVMIs (121 +/- 20 versus 103 +/- 17 g/m; P < 0.001) and BP (152 +/- 12/96 +/- 9 versus 140 +/- 13/86 +/- 9 mm Hg; P < 0.001) with moexipril. For patients who met LVMI inclusion criteria after centralized, blinded readings, the decrease from baseline in LVMIs was 23.4 g/m. The decrease in LVMIs was independent from the regression to the mean phenomenon as observed from the follow-up placebo period. Moexipril 15 mg once daily administered for 24 weeks resulted in a significant reversal of LVH in patients with essential hypertension. The result compares favorably with results previously obtained in trials of similar duration with other ACE inhibitors. PMID- 15662287 TI - Pharmacology and toxicology of a new aqueous formulation of intravenous amiodarone (Amio-Aqueous) compared with Cordarone IV. AB - Hypotension is the most frequent adverse event reported with intravenous amiodarone (Cordarone IV). The hypotension has been attributed to the vasoactive solvents of the formulation, polysorbate 80 and benzyl alcohol, both known to exhibit negative inotropy and hypotensive effect. A new aqueous formulation of intravenous amiodarone (Amio-Aqueous) does not contain vasoactive excipients and may be less toxic and cause less hypotension than Cordarone IV. This hypothesis was tested in a series of animal studies with direct comparison of Amio-Aqueous and Cordarone IV. All studies were performed on Sprague-Dawley rats. The acute toxicology study showed that both LD50 and LD100 were 30% greater for Amio Aqueous than for Cordarone. At the dose at which all animals expired on Cordarone, 50% of animals were still alive on Amio-Aqueous. The study on myocardial contractility showed that Amio-Aqueous was a far less negative inotropic than Cordarone IV (P < 0.001). Amio Aqueous did not have an effect on contractility at 5- and 10-mg/kg dose levels while Cordarone resulted in a 25% (P < 0.01) and 29% (P < 0.002) decrease, respectively. The study on arterial blood pressure showed that Cordarone caused a significant decrease in blood pressure at each of the 3, 5, 10, and 20 mg/kg doses (P < 0.05 to P < 0.001) while Amio Aqueous did not. The study on the antiarrhythmic effects showed comparable efficacy for both formulations. In conclusion, Cordarone IV was more toxic and caused significant hypotension and negative inotropy while Amio-Aqueous lacked the hypotensive and cardiotoxic properties of Cordarone. Therefore, Amio-Aqueous is a safer alternative than the standard formulation. PMID- 15662288 TI - Cutaneous resorption of lead after external use of lead-containing ointments in volunteers with healthy skin. AB - Lead-containing ointments are frequently used in anthroposophic medicine. In a prospective, open-label phase 1 study, 33 volunteers at the Ambulatory Clinic for Complementary and Alternative Medicine at the Free University of Berlin, aged 18 65 years, were exposed to 100 g Plumbum metallicum 0.4% ointment (Weleda, Germany) over a 4-week period. The lead-containing ointment was regularly applied to the cubital side of the forearm. Regular measurements of lead concentrations in whole blood, urine, and scalp hair were determined. None of the 33 volunteers showed an increase in lead concentrations in the 3 investigated compartments after 4 and 8 weeks. Blood lead levels (average value) decreased significantly from baseline to the first week (P < 0.05). Average values in the following investigations (weeks 3, 4, and 8) were significantly lower than at baseline (P < 0.05). There was no increase in lead levels in the scalp hair after 8 weeks (P < 0.05). The results show that the commonly prescribed lead-containing ointment Plumbum metallicum 0.4% in humans with an intact skin does not present a toxic risk. PMID- 15662289 TI - Pharmacokinetics and pharmacodynamics of febuxostat, a new non-purine selective inhibitor of xanthine oxidase in subjects with renal impairment. AB - To assess the safety, pharmacokinetics, and pharmacodynamics of febuxostat in subjects with normal renal function or renal impairment, febuxostat (80 mg/d) was orally administered for 7 days to subjects with normal renal function (n = 11, CLcr >80 mL/min/1.73 m) or to subjects with mild (n = 6, CLcr 50-80 mL/min/1.73 m), moderate (n = 7, CLcr 30-49 mL/min/1.73 m), or severe renal impairment (n = 7, CLcr 10-29 mL/min/1.73 m). The pharmacokinetics of febuxostat and its active quantifiable metabolites 67M-1, 67M-2, and 67M-4 as well as the pharmacodynamics of uric acid, xanthine, and hypoxanthine were determined in plasma (or serum) and urine. Febuxostat was safe and well tolerated. Regression analyses indicated that febuxostat tmax and Cmax,u values were not affected by CLcr. However, for AUC24,u, CLu/F, and t1/2z, regression analyses indicated a statistically significant relationship with CLcr. With the exception of 67M-1 Cmax, regression analyses for 67M-2 and 67M-4 Cmax, and for AUC24 for all 3 metabolites indicated a statistically significant linear relationship with CLcr. Irrespective of renal function group, the mean serum uric acid concentrations decreased by 55% to 64% by day 7. Although plasma exposure to febuxostat and its metabolites was generally higher in subjects with increasing degrees of renal impairment, the percentages of decrease in serum uric acid were comparable regardless of the renal function group. A once-daily 80-mg dose of febuxostat appears to be safe and well tolerated in different renal function groups and does not appear to require any dose adjustment based on differences in renal function. PMID- 15662290 TI - Trimetazidine in Angina Combination Therapy--the TACT study: trimetazidine versus conventional treatment in patients with stable angina pectoris in a randomized, placebo-controlled, multicenter study. AB - The aim of this study was to assess the efficacy and acceptability of trimetazidine (TMZ) in combination with hemodynamic agents (beta-blockers or long acting nitrates) in 177 stable angina patients. In this randomized, placebo controlled study (TACT: Trimetazidine in Angina Combination Therapy), stable angina patients resistant to nitrates or beta-blockers were selected. After a 1 week selection period (W0), patients who had a difference of <10% in duration between 2 positive exercise tests, defined as 1-mm ST-segment depression (STD) 80 milliseconds after J point with angina pain or 1.5 mm without pain were randomly treated with TMZ (20 mg t.i.d., n = 90) or placebo (Pbo t.i.d., n = 87) orally. A final exercise test was performed after 12 weeks of treatment (W12). The efficacy was assessed by exercise test duration, time to 1-mm STD, time to angina onset, mean number of angina attacks, mean short-acting nitrate consumption, and rate pressure product. Differences (W12 - W0) in these parameters were analyzed using the Student t test. All statistical tests were conducted at the 5% significance level. At inclusion and during the study, 52% of patients received long-acting nitrates, and 48% were treated with a beta-blocker as monotherapy. At the beginning of the study, the TMZ and Pbo groups were statistically homogeneous with respect to all analyzed characteristics (demographic characteristics, characteristics of anamnesis, characteristics used for evaluation of antianginal therapy efficacy). For various reasons, 11 patients (7 from the Pbo group and 4 from the TMZ group) were excluded from the trial. A total of 166 patients (80 from the Pbo group and 86 from the TMZ group) completed the study in full compliance with the protocol. After 12 weeks of therapy, exercise test duration increased from 417.7 +/- 14.2 (W0) to 506.8 +/- 17.7 seconds (W12) in the TMZ group versus 435.3 +/- 14.8 (W0) to 458.9 +/- 16.2 seconds (W12) in the Pbo group (P < 0.05). Time to 1-mm STD increased from 389.0 +/- 15.3 (W0) to 479.6 +/- 18.6 seconds (W12) in the TMZ group versus 411.8 +/- 15.2 (W0) to 428.5 +/- 17.3 seconds (W12) in the Pbo group (P < 0.05). Time to onset of anginal pain increased from 417.0 +/- 16.9 (W0) to 517.3 +/- 21.0 seconds (W12) in the TMZ group versus 415.1 +/- 16.5 (W0) to 436.4 +/- 18.5 seconds (W12) in the Pbo group (P < 0.005). The mean number of anginal attacks per week decreased from 5.6 +/- 0.6 to 2.7 +/- 0.5 in the TMZ group versus 6.8 +/- 0.7 to 5.1 +/- 0.7 in the Pbo group (P < 0.05), mean consumption short-acting nitrates per week decreased from 5.2 +/- 0.9 to 2.8 +/- 0.8 in the TMZ group versus 5.5 +/- 0.8 to 4.1 +/- 0.9 in the Pbo group (NS). No change in the rate-pressure product was seen in both. The combination of trimetazidine with beta-blockers or long-acting nitrates significantly improves exercise stress test parameters and angina symptoms compared with placebo. Due to its metabolic effect, free of any hemodynamic action, trimetazidine has proven to be beneficial for combination in patients with stable angina. PMID- 15662292 TI - Mechanism of action of paracetamol. AB - Paracetamol (acetaminophen) is generally considered to be a weak inhibitor of the synthesis of prostaglandins (PGs). However, the in vivo effects of paracetamol are similar to those of the selective cyclooxygenase-2 (COX-2) inhibitors. Paracetamol also decreases PG concentrations in vivo, but, unlike the selective COX-2 inhibitors, paracetamol does not suppress the inflammation of rheumatoid arthritis. It does, however, decrease swelling after oral surgery in humans and suppresses inflammation in rats and mice. Paracetamol is a weak inhibitor of PG synthesis of COX-1 and COX-2 in broken cell systems, but, by contrast, therapeutic concentrations of paracetamol inhibit PG synthesis in intact cells in vitro when the levels of the substrate arachidonic acid are low (less than about 5 mumol/L). When the levels of arachidonic acid are low, PGs are synthesized largely by COX-2 in cells that contain both COX-1 and COX-2. Thus, the apparent selectivity of paracetamol may be due to inhibition of COX-2-dependent pathways that are proceeding at low rates. This hypothesis is consistent with the similar pharmacological effects of paracetamol and the selective COX-2 inhibitors. COX-3, a splice variant of COX-1, has been suggested to be the site of action of paracetamol, but genomic and kinetic analysis indicates that this selective interaction is unlikely to be clinically relevant. There is considerable evidence that the analgesic effect of paracetamol is central and is due to activation of descending serotonergic pathways, but its primary site of action may still be inhibition of PG synthesis. The action of paracetamol at a molecular level is unclear but could be related to the production of reactive metabolites by the peroxidase function of COX-2, which could deplete glutathione, a cofactor of enzymes such as PGE synthase. PMID- 15662293 TI - Drug interactions with paracetamol. AB - Paracetamol (acetaminophen) is one of the most commonly used analgesic antipyretic drugs worldwide, and it is widely available by prescription and over the counter (OTC). Fortunately, few clinically significant drug interactions have been documented. There is probable potentiation of hepatotoxicity following an overdose from the paracetamol metabolite NAPQI by enzyme-inducing drugs. There is considerable controversy regarding the possible interaction with warfarin in its potential to increase its anticoagulant effects because of discrepancies between observational studies and those in healthy volunteers. Otherwise, no serious adverse drug interactions with therapeutic doses of paracetamol have been confirmed in humans. Because the absorption of paracetamol is so dependent on gastric emptying, other drugs that alter gastric emptying can change its pharmacokinetics; but this would not cause serious adverse effects. Although animal experiments have demonstrated that many compounds can modify paracetamol hepatotoxicity, these are unlikely to be important at therapeutic doses. PMID- 15662294 TI - Selecting nonprescription analgesics. AB - Worldwide, there has been an emerging patient demand for access to efficient drugs without consulting a doctor and obtaining a prescription. As a result, there has been an expanding movement of prescription-only drugs to over-the counter (OTC) status. An increasing number of drugs are becoming available OTC, empowering patients to treat themselves. Where the principle of empowering individuals to treat themselves can fail is when consumers lack the knowledge to do so safely. This potentially applies to the self-selection of analgesic drugs by consumers. When used inappropriately, these drugs pose significant risks. The nonsteroidal antiinflammatory drugs (NSAIDs) are associated with many adverse reactions, interactions, and contraindications in a number of patient groups, even at OTC doses. In particular, in the elderly, the high incidence of cardiovascular and gastrointestinal disease, coupled with age-related decline in renal function and multiple medication use, all warrant extra caution with the use of NSAIDs and make paracetamol the simple analgesic drug of first choice. Despite the possibility of hepatotoxicity in overdose, paracetamol represents a better all-round option for most patients requiring OTC analgesic therapy. PMID- 15662295 TI - The role of paracetamol in chronic pain: an evidence-based approach. AB - Chronic pain is a significant public health burden. Several international guidelines and influential reviews recommend the use of paracetamol (acetaminophen) as the first-line analgesic of choice for the management of chronic pain. These recommendations are based largely on the balance of evidence, which favorably demonstrates the efficacy, safety, and low cost of paracetamol relative to other analgesics.A decade ago, March et al suggested that because of the dangers associated with conventional nonsteroidal antiinflammatory (NSAID) use, particularly in the elderly, they should ideally not be used without an individual n-of-1 trial to show that they are more effective than paracetamol. Today, the results of our investigations into the individualization of pain management options continue to support this suggestion. Based on the data available to date, it still seems prudent to use NSAIDs only in those patients in whom there is good evidence of improved efficacy over paracetamol. In patients with chronic pain, paracetamol can play an important role as an NSAID sparer, with resultant benefits in terms of reduced adverse effects and cost savings. PMID- 15662297 TI - Analgesics and osteoarthritis: are treatment guidelines reflected in clinical practice? AB - Osteoarthritis (OA) treatment is complex and multifactorial, with pharmacological regimens requiring sufficient flexibility to be adapted to individual disease progression, flare ups, and response to treatment. Coexisting conditions are common and can lead to problems regarding polypharmacy. Several guidelines have been published for the management of OA pain. While differences exist, most recommend paracetamol as the initial oral drug for OA, based on its efficacy, tolerability, and cost; in patients who respond inadequately to paracetamol, supplementary or replacement analgesics should be recommended. This article considers the reality of analgesic use for OA in clinical practice and the extent to which guidelines are followed both in primary and secondary care. An international survey of rheumatologists (n = 610) found that paracetamol was recommended as first-choice analgesic for OA by 82% of those surveyed. Similarly, in a survey of French GPs, 90% of those surveyed recommended paracetamol first line; NSAIDs were recommended more frequently for stronger pain relief but were also recommended alongside paracetamol as a first-line treatment of mild to moderate pain by 43% of GPs. Finally, a UK patient survey, conducted at a London hospital (n = 200), found that 64% of patients were taking more than 1 drug for treatment of painful OA of the knee or hip; 76% were taking paracetamol and 40% were taking an NSAID. A further 39% had used an NSAID in the past but switched treatment, primarily due to side effects. These findings reinforce the case for the simple analgesic paracetamol to be seen as the cornerstone of pharmacological OA treatment, both as a first-line analgesic and as a foundation to which additional treatment modalities, including NSAIDs, can be added if and when necessary. PMID- 15662296 TI - Do individualized medication effectiveness tests (n-of-1 trials) change clinical decisions about which drugs to use for osteoarthritis and chronic pain? AB - To assess the impact of individualized medication effectiveness tests (IMETs, or n-of-1 trials), on patients' short-term decision making about medications for chronic pain. Survey evaluation of patients undergoing a double-blind, crossover comparison of drug versus placebo, drug versus drug, or drug versus drug combination using paracetamol and ibuprofen in 3 pairs of treatment periods, randomized within pairs. General practice patients (supplemented by a few from 2 tertiary pain clinics) with either chronic pain (> or =3 months), or osteoarthritis (with pain for > or =1 month) severe enough to warrant consideration of long-term nonsteroidal antiinflammatory drug (NSAID) use but for whom there was doubt about the efficacy of NSAID or alternative. Pain and stiffness in sites nominated by the patient, global pain, use of escape analgesia, and side effects. Of 116 IMETs started, 71 were completed. Drug management changed for 46 of 71 (65%). The most common change was to add paracetamol or to substitute the NSAID or COX-2 inhibitor with paracetamol (25 of 71 patients and 54% of changes). Of the 37 who were using NSAIDs or COX-2 inhibitors before the IMET, 12 (32%) ceased these afterward. Paracetamol was as effective or more effective than ibuprofen in 37 (68%) of the 54 IMETs directly comparing these drugs. IMETs provide useful information for clinical decisions. Paracetamol continues to be useful for patients with chronic pain whose optimal drug choice is in doubt. Our results provide a new (individual) perspective on the well-known recommendation for paracetamol as first-line treatment for chronic pain and demonstrate that it is feasible to provide IMETs nationally by mail and telephone. PMID- 15662298 TI - Total hip replacement: a successful interaction of biology, mechanics, and materials science. PMID- 15662299 TI - The long-term results of low-friction arthroplasty of the hip performed as a primary intervention. 1970. PMID- 15662300 TI - Strain adaptive bone remodelling in total joint replacement. AB - Histomorphologic analyses of artificial joint components implanted into bone need special technology for processing and for documentation; published histological work systematically done therefore is rare. The histopathology, three dimensionally analyzed in a complete sequence of sections is, however, the only precise answer in terms of biocompatibility and bone response. A complete analysis allows a type-related predictable prognosis of an implantation that is at least comparable to a finite element analysis with respect to load transfer to host bone. The histopathologic collection of the ZOW Munich is comprised of more than 5000 nondemineralized bone and joint specimens and more than 500 artificial joint components implanted in the human skeleton for up to 25 years. Fifty-nine implant-bone specimens without signs of loosening already have been processed and analyzed systematically. According to the strain-adapted bone remodelling, different types of anchorage clearly were differentiated and their morphologic substrate could be worked out. Based on that, the cemented standard anchorage could be distinguished histologically from the cemented press-fit procedure, and the noncemented press-fit from the porous ingrowths pattern. In terms of the topography of the bony integration, the proximal and distal press-fit and ingrowth pattern were analyzed; beside that, the cemented and noncemented epiphyseal resurfacings could be defined histologically. In all histologic specimens the remodelling appeared as a result of stress-related strain, reflecting stiffness of the implant and the resistance of bone to deformation. It clearly was worked out that all success of cemented components is based on preserved cancellous bone honeycombs stiffened by bone cement, representing an adaptation of bone in terms of stiffness to the stiff implants. PMID- 15662301 TI - The microenvironment around total hip replacement prostheses. AB - The metal stem of the totally replaced hip carries load and resists fatigue, but it is electrochemically corroded. Metallic atoms act as haptens, induce type 1 T helper cells/Th1-type immune responses and enhance periprosthetic osteolysis. Stiff metal implants, which do not have the same elasticity as the surrounding bone, cause stress shielding. Cyclic loading and lack of ligamentous support lead to mechanical and ischemia reperfusion injury and particle formation from bone, polymethylmethacrylate, and porous implant surfaces, which accelerate third-body polyethylene wear. Surgical injury and micromotion induce the formation of a fibrous capsule interface. Type-B lining cells produce lubricin and surface active phospholipids to promote solid-to-solid lubrication but may loosen the implant from bone. The pumping action of the cyclically loaded joint and synovial fluid pressure waves dissect the implant-host interface and transports polyethylene particles and pro-inflammatory mediators to the interface. Hyaluronan induces formation of a synovial lining like layer. Because of its localization close to bone, foreign body inflammation at the interface stimulates osteoclastogenesis and peri-implant bone loss. Metal-on-metal and ceramic-on ceramic pairs might minimize third body wear, but can lead to high-impact load of the acetabulum. Diamond coating of a metal-on-polyethylene couple might solve both of these problems. The basic biomaterial solutions allow good mechanical performance and relatively long life in-service, but surface modifications (porous coating, hydroxyapatite, diamond, bioglass, and others) may facilitate performance of the implant and improve the biomaterial and body interfaces. PMID- 15662302 TI - Pharmacologic modulation of periprosthetic osteolysis. AB - Wear and periprosthetic osteolysis of total joint replacements continue to be the most important problems in arthroplasty surgery. Despite the introduction of improved technologies including alternative bearing surfaces for TJRs, wear is inevitable because of relative movement at different interfaces and processes such as electrolysis and material degradation. Worn, clinically failing implants need to be followed closely and revised when appropriate. However, early wear and minor osteolysis do not result necessarily in progressive failure of the prosthesis. Indeed such cases may be followed up clinically and radiographically to establish the functional and biologic sequelae of wear and the timeline of these events. This scenario provides an opportunity to modulate the adverse biologic reaction associated with wear particles that includes chronic inflammation, the foreign body response, and periprosthetic bone destruction. Currently, immunological events associated with wear particles are becoming understood more clearly. Strategies to mitigate adverse processes associated with wear debris include local or systemic administration of immune modulators, signaling molecules, anti-inflammatory agents and growth factors, and altering osteoclast function. Ultimately, prevention of accelerated wear and periprosthetic osteolysis will be achieved with improved bearing surfaces and prosthetic designs. PMID- 15662303 TI - A review of current cross-linked polyethylenes used in total joint arthroplasty. AB - Major improvements have been made in new polyethylenes in regards to wear resistance and oxidation resistance. The background for and explanation of these improvements are presented in this study. The various manufacturing processes are described along with the possible features that the differences in manufacturing processes may have on wear, rate of particle generation, oxidation and mechanical properties. The role of some of the new polyethylenes in permitting the use of larger-diameter heads and the advantages of large-head diameters are discussed. Some of the advantages of metal-on-polyethylene versus hard-on-hard bearings also are described. PMID- 15662304 TI - Metal-on-metal total hip replacement. AB - Metal-on-metal bearings have wear rates that are 20 to 100 times lower than metal on-conventional polyethylene. The amount of wear generally is the same order of magnitude for the head and the cup. There is an initial run-in period of higher wear followed by lower, steady-state wear. Wear rate is a function of the interplay of material(s), macrogeometry, microgeometry, and the resultant type and amount of lubrication. The wear resistance and clinical performance of a metal-on-metal bearing are more sensitive to macrogeometry and lubrication than a metal-on-polyethylene bearing. Metal wear particles are nanometers in linear dimension. They are much smaller and more numerous than the submicron polyethylene wear particles, but the volume of periprosthetic inflammatory tissue is less. Osteolysis seems to be relatively rare. Little is known about the systemic distribution of metal particles and ions. The significance of systemic distribution also is not known. The levels of serum and urine Co and Cr ions are elevated in patients with metal-on-metal bearings, but the long-term, steady state levels are not much higher than those from corrosion of modular femoral components. Because of the elevated levels of Co and Cr ions, there is a greater risk of delayed type hypersensitivity. There also is concern about the potential for malignant degeneration secondary to prolonged exposure to these elements. The available data are insufficient to address this concern. Rigorous long-term studies are needed. It will take decades of close clinical observation to determine if the benefits of metal-on-metal bearings outweigh the associated risks. PMID- 15662305 TI - Ceramics in total hip replacement. AB - Alumina-on-alumina total hip arthroplasty has been used for 32 years in Europe. The theoretical advantages of this combination are represented by its remarkable sliding characteristics, its very low wear debris generation, and its improved fracture toughness. These advantages are achieved if the material is processed properly with high density, high purity and small grains. We summarize the results obtained with the alumina-on-alumina combination concerning in vitro and in vivo wear behavior with special emphasis on wear debris characterization and quantification and histologic tissue examinations. Alumina-on-alumina seems to be one of the best choices in young and active patients provided that sound socket fixation is maintained in the long term. PMID- 15662306 TI - Potential of coatings in total hip replacement. AB - In total hip replacements, the bulk properties of materials, such as proper elasticity and hardness, are important. However, the material interacts with the body mainly at the surfaces. Wear and corrosion are initiated at the surfaces also. Therefore, the control of surface properties using different kinds of treatments or coatings may improve total hip replacements considerably. The most studied surface treatments include ion implantation and methods to control surface topography, such as grit or sand blasting or plasma treatments. Among the large variety of coatings, hydroxyapatite, titanium oxide and nitride, zirconium oxide, pyrolytic carbon, and diamondlike carbon coatings have shown the most promising results. These coatings mainly are used to enhance bone growth; to minimize friction, wear, and corrosion; and to improve biocompatibility of total joint prostheses. The potential of novel coatings to solve some present problems in joint prostheses is discussed based on the structure and properties of different kind of coatings. It can be concluded that currently, coating methods exist to improve the tribologic performance and longevity of the total hip replacements. However, coatings must fulfill two essential requirements: no delamination in biochemical and biomechanical environments and sufficient protection of substrate from corrosion. PMID- 15662308 TI - Kinetically critical sites of femoral head roughening for wear rate acceleration in total hip arthroplasty. AB - Polyethylene wear acceleration from (scratching) damage to the femoral head is a recognized hazard from constructs prone to generate third-body debris, but the phenomenon is nebulous and therefore often is subordinated to more direct and immediate considerations. To help delineate tangible quantitative relationships between counterface roughening and accelerated polyethylene wear, an experimentally validated sliding-distance-coupled finite element model of total hip replacement wear was adapted to incorporate regions of localized femoral head roughening. This computational formulation was used systematically to identify the sites on the femoral head for which a given severity of local roughening (parameterized in terms of roughening patch size and tribologic wear coefficient) was most consequential in terms of elevated polyethylene wear. Two such sites, of nominally comparable kinetic importance, were consistently evident throughout a wide range of roughening severities. These critical sites were located quasi superiorly near the sagittal midline of the head, one slightly anterior and one slightly posterior of the coronal midline. PMID- 15662307 TI - Surface-gradient cross-linked polyethylene acetabular cups: oxidation resistance and wear against smooth and rough femoral balls. AB - Two methods were developed and evaluated for cross-linking the bearing surface of a polyethylene acetabular cup to a limited depth, in order to improve its resistance to wear without degrading the mechanical properties of the bulk of the component. In the first method, low-energy electron beams were used to cross-link only the bearing surface of the cups to a maximum depth of about 2 mm. The cups then were annealed at 100 degrees C in vacuum for 3 or 6 days to reduce the residual free radicals, and the resultant resistance to oxidation was compared by artificially aging the cups at 80 degrees C in air. Chemically cross-linked surface layers were produced by coating the bearing surfaces of the cups with a thin layer of polyethylene powder mixed with 1% weight peroxide, and compressing them at 6.9 MPa (1000 psi) and 170 degrees C. This resulted in a cross-linked surface layer that extended about 3 mm deep, with a gradual transition to conventional (noncross-linked) polyethylene in the bulk of the implant. In hip simulator wear tests with highly polished (implant quality) femoral balls, both types of surface cross-linking were found to improve markedly the wear resistance of the acetabular cups. In tests with roughened femoral balls, the wear rates were much higher and were comparable to those obtained with similarly roughened balls against noncross-linked polyethylene cups in a previous study, indicating that the full benefit of cross-linking may not be realized under conditions of severe third-body abrasion. Nevertheless, these results show a promising approach for optimizing the wear resistance and the bulk mechanical properties of polyethylene components in total joint arthroplasty. PMID- 15662309 TI - Prevention of shoulder stiffness after rotator cuff repair. AB - Predisposing factors for shoulder stiffness after rotator cuff repair have yet to be determined. The potential for recovery of range of motion and amelioration of pain in patients with this complication also remains unclear. Accordingly, data collected prospectively for 209 patients with a primary rotator cuff repair were retrospectively reviewed. Two groups, Group A (early motion recovery) and Group B (shoulder stiffness), were selected according to passive shoulder range of motion outcomes 6 weeks postoperatively. Both groups were compared for 10 descriptive and clinical characteristics, and for passive range of motion, muscle force, and functional outcomes obtained 0, 6, 12, 24, and 76 weeks postoperatively. Of the potential prognostic factors examined, restriction of range of motion for the preoperative hand behind the back best predicted shoulder stiffness at 6 weeks postoperatively. For patients with postoperative shoulder stiffness, pain had subsided by 24 weeks postoperatively, whereas range of motion steadily improved between 6 weeks and 76 weeks postoperatively. Results of the current study support a predictive role for restriction of range of motion for the preoperative hand behind the back, and affirms the potential for nearly complete recovery of range of motion and amelioration of pain in patients who have shoulder stiffness after rotator cuff repair. LEVEL OF EVIDENCE: Prognostic study, Level I-1 (prospective study). PMID- 15662310 TI - Reconstruction of the deltoid and acromion after failed acromionectomy. AB - Ten patients who had reconstruction of the acromion and deltoid in a painful dysfunctional shoulder after an acromionectomy were reviewed with respect to pain, range of motion, strength, patients' satisfaction, and complications. There were four total acromionectomies, five subtotal acromionectomies, and one partial acromionectomy. The retracted deltoid and defect of the bony acromion were reconstructed with a composite graft consisting of tricortical iliac bone with fascia lata. At an average followup of 58 months, the mean active forward flexion improved from 53 degrees to 122 degrees. All patients reported considerable relief of pain and three had no pain; however, the patients who had a supplementary cuff repair had poor results. The average UCLA score of the patients improved from 6.6 to 26.3. The score of the patients with intact rotator cuffs improved from 7.0 to 32.0, whereas patients who had rotator cuff tears had scores that improved from 6.2 to 20.4. Our clinical results suggest that reconstruction of the acromion and deltoid using an iliac bone graft and fascia in failed acromionectomy can lead to relief of pain and the improvement of shoulder function. LEVEL OF EVIDENCE: Therapeutic study, Level IV (case series no, or historical control group). PMID- 15662311 TI - Maintenance of interbody space in one- and two-level anterior cervical interbody fusion: comparison of the effectiveness of autograft, allograft, and cage. AB - The use of allografts, autologous iliac crest grafts, and cages for anterior cervical fusion is well documented, however there is no comparison regarding the effectiveness of maintaining the interbody space with the three approaches. We retrospectively measured the rate and amount of interspace collapse, segmental sagittal angulations, clinical results, and radiographic fusion success rates to determine which is the best fusion material. We assessed 73 patients who had one- and two-level cervical discectomies and interbody fusions without instrumentation. The three groups had similar clinical results and fusion rates. However, in the autograft group union occurred in 4 months. In the allograft group, union did not occur until 5.54 months. Moreover, the loss of cervical lordosis (2.75 degrees) was less in the cage group than in the allograft group (9.23 degrees). Additionally, the anterior interspace collapse (1.73 mm) in the cage group was less than the collapse recorded in the autograft group (2.82 mm) and in the allograft group (4 mm). An interspace collapse of 3 mm or greater was observed in 56.1% of the patients in the allograft group, compared with only 19% of the patients in the cage group. We showed that the cage is superior to the allograft and autograft in maintaining cervical interspace height and cervical lordosis after one-level and two-level anterior cervical decompression procedures. LEVEL OF EVIDENCE: Therapeutic study, Level III-2 (retrospective cohort study). PMID- 15662312 TI - Implant survivorship and complication rates after total knee arthroplasty with a third-generation cemented system: 5 to 8 years followup. AB - We evaluated implant survivorship, reoperation rates, and complication rates of a group of patients who had total knee arthroplasty with a third-generation cemented prosthetic device using cruciate-retaining and posterior-stabilized designs at 5 to 8 years followup. Three hundred thirty-four consecutive primary total knee arthroplasties (186 cruciate retaining and 148 posterior stabilized) were done in 287 patients at our institution during a 2-year period. Kaplan Meier survivorship using revision for any reason and revision for aseptic loosening as endpoints were 95.9% and 99.5% respectively at 8 years. Nine patients (four with cruciate-retaining total knee arthroplasties, five with posterior-stabilized total knee arthroplasties; 3.1%) had reoperations for any reason. No patients had reoperation for problems related to the patellofemoral joint. Thirty-two patients (11.1%) had intraoperative or postoperative complications. There were no differences in any of the outcomes analyzed between patients who had cruciate retaining or posterior-stabilized total knee replacements. Our results show that with appropriate patient selection and meticulous attention to surgical technique, excellent clinical and radiographic results can be achieved with a third-generation total knee arthroplasty system at intermediate followup. LEVEL OF EVIDENCE: Therapeutic study, Level III-2 (retrospective cohort study). PMID- 15662313 TI - Treatment of infected total knee arthroplasty using an articulating spacer: 2- to 12-year experience. AB - Fifty consecutive patients with late infected total knee arthroplasties were treated by debridement and removal of all components and cement, preserving the collateral ligaments. At the time of debridement, an articulating spacer was made to allow partial weightbearing and range of motion of the knee during rehabilitation. This spacer was implanted using antibiotic-impregnated bone cement. For this purpose, 4.8 g powdered tobramycin was mixed with 40 g Simplex cement. Cement was applied early to the components, but applied late to the femur, tibia, and patella to allow molding to the defects and bone without adherence to bone. Patients had tailored intravenous antibiotic therapy for 6 weeks for treatment of various gram-positive and gram-negative organisms. All patients had cemented revision total knee arthroplasty using antibiotic impregnated cement with standard cementing techniques. Range of motion before reimplantation was 6 degrees -91 degrees . Followup averaged 73 months (range, 24 150 months). The average modified Hospital for Special Surgery knee score after revision was 89 points (range, 70-100 points) with 90% good to excellent results, excluding the results of patients with reinfection. Range of motion after reimplantation was 4 degrees -104 degrees. Six patients had recurrences of infection, and one patient with a poor postoperative range of motion had a fusion. Use of an articulating spacer achieved soft tissue compliance, allowed for ease of operation, reduced postoperative pain, improved function, and eradicated infection equal to standards reported in the literature. LEVEL OF EVIDENCE: Therapeutic study, Level IV (case series-no, or historical controls). PMID- 15662314 TI - Total knee arthroplasty: range of motion across five systems. AB - Range of motion after total knee arthroplasty is an important variable in determining clinical outcome. The goal of this study was to assess range of motion across five types of posterior-stabilized knee prostheses used sequentially in the same institution during 17 years. The hypothesis was that absolute flexion would improve in newer models of this basic prosthesis design. Only primary knee arthroplasties in patients with osteoarthritis were evaluated. A retrospective analysis was done. Three hundred fifty-eight knees with osteoarthritis were reviewed. The average arc of motion was 103 degrees before surgery and 111 degrees after surgery. Absolute flexion was clinically similar but improved from before surgery (110 degrees ) to after surgery (113 degrees ). No difference was found when comparing improvements in range of motion among the different types of prostheses used. This study did not show that any knee system made a difference in determining the final range of motion postoperatively. Height emerged as a predictive factor of absolute flexion. Preoperative range of motion is the most important variable in determining improvements in range of motion, with height playing a secondary role. LEVEL OF EVIDENCE: Prognostic study, Level II-1. PMID- 15662315 TI - Surgery for equinovarus deformity in Friedreich's ataxia improves mobility and independence. AB - Friedreich's ataxia results in morbidity because of many factors; progressive equinovarus deformity is one of these. We studied the risk factors and incidence of this deformity. We sought to assess whether surgical management of fixed equinovarus deformity leads to functional improvement. Thirty-six patients with Friedrich's ataxia were assessed for this deformity. These patients were treated by splinting, botulinum toxin Type A injection, and surgery, as indicated by the severity, followed by an ongoing rehabilitation program. The effect of surgery was assessed using subscales of the Barthel index and functional independence measure. Severe foot deformities in which either surgery or botulinum toxin injection was recommended correlated with current age, years since disease onset, and years that the patient required a wheelchair for mobility, but not with the GAA repeat size or age at disease onset. Function and mobility were improved after surgery compared with a similar period before surgery. Three of seven patients who had surgery had significant complications. Aggressive management of foot deformities should be considered, and active measures to prevent permanent foot deformities should be pursued to maximize quality of life and independence of patients with Friedreich's ataxia. LEVEL OF EVIDENCE: Therapeutic study, Level IV (case series-no, or historical control group). PMID- 15662316 TI - TP-3 immunotoxins improve antitumor activity in mice with osteosarcoma. AB - We measured the antitumor activity of two types of TP-3 immunotoxins that target an antigen expressed in tumors associated with osteosarcoma. Development of novel agents for treatment of patients with osteosarcoma is important. We previously described a monovalent-disulfide-stabilized recombinant immunotoxin made from the TP-3 antibody. This agent is called TP-3(dsFv)-PE38 and is cytotoxic to human osteosarcoma cells in vitro. To improve antigen binding, we designed and produced a bivalent immunotoxin, TP-3(dsFv)2-PE38. We evaluated the activity of both molecules in vitro and in vivo using tumor-bearing mice. Compared with the monovalent TP-3 immunotoxin, the bivalent TP-3 immunotoxin showed an approximately sevenfold increase in cytotoxic activity against three osteosarcoma cell lines which react with the TP-3 monoclonal antibody. The apparent affinity of the bivalent TP-3 immunotoxin was 12-fold greater than that of the monovalent TP-3 immunotoxin. The antitumor activities of both TP-3 immunotoxins were measured using severe combined immunodeficient mice bearing osteosarcoma cell line OHS-M1 tumors. The dose at which the bivalent TP-3 immunotoxin produces complete regressions of tumors is (1/2) that of the monovalent TP-3 immunotoxin. Increasing the avidity of TP-3(dsFv)-PE38 significantly improves its cytotoxic activity in vitro and results in a twofold increase in antitumor activity in vivo. Because TP-3-based immunotoxins have good antitumor activity in mice, these molecules merit additional development for possible treatment of osteosarcoma in humans. PMID- 15662317 TI - Childhood cancer survivors: an at-risk cohort for ankle osteonecrosis. AB - We aimed to determine if the ankle is at risk for corticosteroid-induced osteonecrosis as a cause of pain in a cohort of childhood cancer survivors. We retrospectively reviewed magnetic resonance imaging scans of the ankle in patients treated at our institution between 1993 and 2003. Among the 15 patients who met study criteria, 20 of 30 (67%) ankles were involved. Older children had the highest incidence of the disease. The tibial metaphysis, epiphysis, and talus were the most frequent sites of osteonecrotic lesions. Ten patients were symptomatic with extended weightbearing activity. Surgery was required in four with continued amelioration of pain at last followup (17-44 months). As early detection and intervention lead to prevention of ankle morbidity, childhood cancer survivors would benefit from a prospective multi-institutional study that would provide understanding of ankle osteonecrosis and development of effective interventions. LEVEL OF EVIDENCE: Prognostic study, Level II-1 (retrospective study). PMID- 15662318 TI - Functional recovery after a reverse prosthesis for reconstruction of the proximal humerus in tumor surgery. AB - An alternative treatment for primary bone tumors of the proximal humerus was assessed. Four patients, who made full functional recovery after complete resection of the proximal humerus inclusive of the rotator cuff and subsequent reconstruction with a reverse shoulder prosthesis, were examined clinically and radiographically. Distinct medialization of the center of rotation of the glenohumeral joint (28 mm) and elongation of the remaining deltoid muscle (116%) were measured. Increased scapular rotation (118%) was observed. The radiologic results and thoracoscapular rhythm analyses were implemented in a three dimensional computerized model of the glenohumeral joint. This allowed us to calculate a doubling of the moment of the deltoid abductor muscle in the true scapular plane. After tumor surgery, in which the proximal humerus is resected without reinserting the rotator cuff, full functional recovery of the shoulder can be obtained with a total shoulder prosthesis, medializing the glenohumeral center of rotation and elongating the remaining deltoid muscle. LEVEL OF EVIDENCE: Therapeutic study, Level IV (case series-no, or historical control group). PMID- 15662319 TI - Which is a better method for Perthes' disease: femoral varus or Salter osteotomy? AB - The optimum surgical treatment of patients with Perthes' disease has not been decided. We compared the clinical and radiographic results at skeletal maturity of the femoral varus osteotomy and the Salter innominate osteotomy for treatment of patients with Perthes' disease. Treatment in 46 patients was by femoral varus osteotomy and in 30 patients by Salter innominate osteotomy. There were no significant differences between the two groups. The clinical results were similar in the two groups except for scarring after surgery, which was more prominent in the femoral varus osteotomy group. There was no significant difference in the sphericity of the femoral head and congruity of the hip between the two groups. However, coverage of the femoral head by the acetabulum, the neck-shaft angle, and the articular-trochanteric distance were closer to normal in the Salter innominate osteotomy group. We think that the Salter innominate osteotomy is the better treatment for patients with Perthes' disease to decrease residual problems such as coxa vara, trochanteric prominence, poor acetabular coverage, and the surgical scar. LEVEL OF EVIDENCE: Therapeutic study, Level IV (case series-no, or historical control group). PMID- 15662320 TI - Clarithromycin destroys biofilms and enhances bactericidal agents in the treatment of Pseudomonas aeruginosa osteomyelitis. AB - In this study, the effect of clarithromycin on the destruction of bacterial biofilm in Pseudomonas aeruginosa osteomyelitis was investigated. Foreign body related osteomyelitis caused by ceftazidime-sensitive Pseudomonas aeruginosa was produced in the tibias of 26 rats. After osteomyelitis was verified on Day 14, 10 rats had ceftazidime (1500 mg/kg/day) given subcutaneously, and 10 rats had ceftazidime given subcutaneously and clarithromycin (100 mg/kg/day, two 50-mg/kg doses every 12 hours) given orally; three rats formed the control group. After a treatment period of 20 days, the tibias and the foreign bodies were removed, cultured, and examined by electron microscopy. The number of microorganisms growing on the bone tissue in the group receiving combined treatment was significantly lower than in the other groups. The number of microorganisms growing on the foreign body in the group receiving only ceftazidime was significantly higher than that of the group receiving combined treatment. Electron microscope examination revealed that the biofilm layer was eradicated in the group that had combined therapy; however, biofilm formation was evident on the foreign body in the group receiving only ceftazidime. Clarithromycin enhanced the activity of concomitantly used bactericidal agents by destroying biofilm on the surface of the materials. PMID- 15662322 TI - Image-guided reconstruction of femoral fractures: is the staff progeny safe? AB - The potential of adverse effects to progeny caused by preconceptual and fetal exposure to ionizing radiation is an issue of increasing concern to orthopaedic surgeons and assisting staff. Are these fears justified? How effectively is the embryo or fetus protected, and should pregnant staff alter their duties? In this study, an anthropomorphic phantom was exposed fluoroscopically at two geometries common in surgical reconstruction of proximal femur fractures. Scatter radiation was converted to gonadal dose and embryo or fetal dose with and without use of a protective apron. The genetic risk for the orthopaedic surgeon after 10 years of occupational exposure was estimated to be 16,000 times lower than the natural frequency of heritable disease. The excess risk of childhood cancer associated with the dose accumulated during gestation was at least 600 times lower than the corresponding natural frequency. A properly shielded pregnant orthopaedic surgeon is allowed to do 14 hours of hip fluoroscopy during gestation, whereas 2100 hours of fluoroscopy are required for the induction of gross malformation or mental retardation to the growing embryo or fetus. The supplementary dose constraints for pregnant staff provide adequate protection to the unborn child without affecting regular personnel duties. PMID- 15662321 TI - A new locking plate for unstable fractures of the proximal humerus. AB - In a prospective study, 28 patients with 29 proximal humeral fractures were treated with the Locking Proximal Humerus Plate from 2001-2002. Most of these fractures were complex, Type B (n = 15) and Type C (n = 9) fractures, according to the AO classification. Followups included radiographs, clinical examinations, and recording Constant scores. After 1 year, the average Constant score for all fractures was 74.6 (range, 37-96). For Type A fractures, it was 82.6 (range, 60 96), for Type B it was 78.3 (range, 37-95), and for Type C it was 64.6 (range, 40 91). Complications related to the implant included breakage of the plate in one patient and redislocation of the fracture in four patients (one fracture was associated with deep infection), which required reoperation in two patients. Partial osteonecrosis was seen in two patients, once after deep infection. There were no nonunions. Our data show that using the Locking Proximal Humerus Plate for treatment of proximal humeral fractures of all types is a reliable procedure, with good results being obtained with careful planning and familiarity with the special features of the operative technique. LEVEL OF EVIDENCE: Therapeutic study, Level II-1 (prospective cohort study). PMID- 15662323 TI - Low-intensity ultrasound enhances maturation of callus after segmental transport. AB - The purpose of this study was to determine whether low-intensity ultrasound can be used to enhance callus maturation. Fifteen-millimeter bone defects at the metatarsal bones of sheep were treated with a segmental bone transport for 16 days. The callus formations in the bone defects were allowed to mature for 63 days before the animals were sacrificed. Eighteen sheep were operated on and divided into two groups. One group was treated with low-intensity ultrasound for 20 minutes per day, whereas the other group served as an untreated control group. Biomechanical tests after removal of the metatarsals showed significantly higher axial compression stiffness and significantly higher indentation stiffness of callus tissue in the healing zone in the group treated with ultrasound. Also, histologic analysis of the cortical defect zone showed significantly more callus formation and more active zones of endochondral ossification in the group treated with ultrasound. Stimulation of callus maturation by ultrasound is possible, similar to stimulation of fresh fracture healing, and may be used to shorten clinical treatment times. PMID- 15662325 TI - Articular cartilage adjacent to experimental defects is subject to atypical strains. AB - We tested the hypothesis that articular cartilage adjacent to experimental osteochondral defects is not subject to unusual strains under load. A 2.5-mm drill hole was made in the medial femoral condyle of 15 knees from 10 adult rabbits. Experimental joints were loaded with simulated quadriceps force, then frozen under load and preserved by freeze-substitution fixation. Deformation in the region of the defect was evaluated by scanning electron and light microscopy and compared with nondrilled and nonloaded control knees. To simulate blood clot, alginate was placed into some defects before loading. In loaded knees, articular cartilage at the edge of the drill hole was abnormally flattened and folded into the defect. Opposing tibial cartilage or meniscus intruded into the femoral defect beyond the cement line. Alginate did not prevent incursion of opposing cartilage. In this standard drill-hole model, the articular cartilage defect is occupied by the opposing surface when a joint is loaded. Any tissue growing or surgically implanted in the defect is subject to loading and displacement, therefore complicating attempts to characterize the healing or regenerative potential in similar drill-hole models. Deformation of cartilage at the defect edge suggests load concentration or increased compliance. Either phenomenon would contribute to subsequent degeneration of the cartilage adjacent to defects. PMID- 15662324 TI - Alendronate inhibits bone resorption at the bone-screw interface. AB - In the current study, we investigated whether the systemic administration of alendronate, a third-generation bisphosphonate, suppressed the loosening of screws at the bone-screw interface. We systemically administered alendronate to rats fitted with external fixators. External fixators with two half pins were applied to the right femurs of rats, and alendronate was administrated once a week during a 5-week postoperative period. Radiographic, histologic, and immunohistochemical findings subsequently were analyzed. Treatment with alendronate reduced the width of the fibrous loosening membrane and the number of osteoclasts at the bone-screw interface. These findings indicate that systemic treatment with alendronate exerts an inhibitory effect on local bone resorption at the bone-screw interface. PMID- 15662326 TI - Repair of steps and gaps in articular fracture models. AB - To examine the healing of surface defects in articular fractures, gaps measuring 0.5 mm wide and 2 mm deep were created in 0.5-mm coronal step-offs on the medial femoral condyles of 16 rabbits, and identical gaps without step-offs were created in another set of 13 rabbits. Evaluation of repair was done 6, 12, and 24 weeks postoperatively. Histologically, subchondral bone restoration of gaps in step offs was complete by 24 weeks, whereas restoration was incomplete in most gap only lesions. Bone density measured by peripheral quantitative computed tomography was normal in gaps in step-offs by 24 weeks, but values were less than in controls in the gap-only group. A moderate degree of degeneration was detected only at the high sides of step-off + gap lesions. The joint surface was restored by fibrous cartilage that showed gradually improving maturity in all defects, however, real integration with adjacent original cartilage did not occur. Immunohistologic examination showed decreasing collagen Type I and increasing Type II staining intensity in repair tissue of both types of lesions. These observations suggest that minor surface gaps and steps of articular fractures may regenerate without inducing severe early joint degeneration. However, certain repair features make the long-term outcome of these defects uncertain. PMID- 15662328 TI - James Platt White, MD (1811-1881): his interesting and remarkable accident. AB - James Platt White, MD (1811-1881), one of the founders and leading figures of the Buffalo Medical College and a pioneer in American obstetrics and gynecology, suffered an interesting and remarkable accident to his neck at the age of 26 while traveling in a stagecoach in Western New York. He was confined to bed until after 45 days, a piece of bone was discharged into his pharynx and then expectorated. The segment of bone proved to be the anterior arch of his atlas (C1) vertebra. He recovered completely from this injury except for permanent loss of rotation of his head and neck. However, he was without functional disability until his death, 44 years later, at the age of 70. This case documents the clinical result during a 44-year period after traumatic loss of the anterior arch of C1. Such cases have been reported only rarely in the literature. Only limited information is available regarding the long-term clinical significance of a Jefferson fracture with exfoliation of the anterior arch of C1. My analysis suggests that White suffered an open Jefferson's fracture that became infected. The anterior arch of C1 became a sequestrum and was discharged spontaneously into his pharynx and then expectorated. This case report with decades of followup should be of interest to all who care for patients with cervical spine injuries and those who are interested in the history of medicine in Western New York. PMID- 15662327 TI - Bone mesenchymal stem cells transplanted into rabbit intervertebral discs can increase proteoglycans. AB - We sought to determine whether transplanted allogeneic bone mesenchymal stem cells can survive and increase the amount of proteoglycans in intervertebral discs. We used the rabbit intervertebral disc as a model, creating three groups: an uninjected control group, a group injected with saline, and a group injected with 1 x 10(5) of bone mesenchymal stem cells containing trace marker gene LacZ from young rabbits. At 1, 3, and 6 months, X-gal staining and DNA-polymerase chain reaction of the neomycin-resistant gene were used to ascertain cell location. Reverse transcription-polymerase chain reaction and enzyme-linked immunosorbant assay analysis were done to evaluate the effect on the disc matrix. Transplanted mesenchymal stem cells were located and identified in the group injected with mesenchymal stem cells, whereas we detected none in the saline and control groups. The amount of mRNA and protein of proteoglycan and collagen Type II in the mesenchymal stem cells group was increased, whereas the amount of collagen Type I did not change. We found no changes in the saline group. Our data suggest transplanted allogenic bone mesenchymal stem cells can survive and increase proteoglycan amount, supporting its potential use as a treatment of intervertebral disc degeneration. PMID- 15662329 TI - Compressive intrapelvic synovial cysts: an early complication of an HA-coated cup. AB - Synovial cysts arising from the hip are not common but can develop in response to intraarticular derangement and increased intraarticular pressures. We present a case of a compressive intrapelvic synovial cyst 3 years after a total hip replacement with a hydroxyapatite-coated acetabular component. At surgery, cracking of the coating of the acetabular component was observed in the areas where the cup was not covered by host bone. The cyst was excised and the bearing surface was replaced. Three years after surgery there has been no recurrence. Migration of hydroxyapatite granules from uncovered portions of hydroxyapatite coated cups may be a source of third-body wear and could accelerate synovial cyst formation. PMID- 15662330 TI - Resection and reconstruction of a massive femoral hemophilic pseudotumor. AB - Hemophilic pseudotumor is a rare, but well-known, complication of hemophilia. We describe a 50-year-old man with mild hemophilia A, but with no previous need for Factor VIII supplementation, who presented with a pathologic fracture of the right femoral neck and shaft caused by a large hemophilic pseudotumor. Initial nonoperative therapy with factor replacement and skeletal traction resulted in radiographic evidence of fracture healing, but the patient's pain persisted. Therefore, he had a radical resection of his hemophilic pseudotumor (soft tissue component and entire femur), and reconstruction with a custom total femoral replacement. Six months after resection, the patient returned to full-time employment. Although pseudotumor formation is a well-recognized complication of hemophilia, the pseudotumor in our study is one of the largest yet described. More importantly, to our knowledge this is the first report of a pseudotumor treated by radical resection and reconstruction with a custom femoral prosthesis. We think that radical resection and reconstruction with a custom total femoral prosthesis is a valuable alternative to amputation in massive pseudotumors of the femur and soft tissues of the thigh. PMID- 15662331 TI - Salmonella spondylodiscitis in patients without sickle cell disease. AB - The optimal treatment of salmonella spondylodiscitis is controversial. The cases of eight patients who had salmonella spondylitis without sickle cell disease were reviewed. Back pain (100%), fever (75%), and elevated C-reactive protein levels (100%) were common, but gastrointestinal symptoms were not (0%). Six patients had positive blood cultures, and the other two had positive tissue cultures. Group C1 salmonella was the most common serotype. Two patients with coexisting aortic mycotic aneurysms had immediate aneurysm resection. Three others responded favorably to appropriate antibiotics, and three required subsequent surgical reconstruction because of neurologic impairment or osseous instability. Clinical outcomes were significantly better than those of 46 previously reported patients. Salmonella spondylodiscitis usually responds favorably to appropriate antibiotics; consequently, a tissue diagnosis is important. Operative interventions are necessary only for patients with coexisting aneurysms or ongoing osseous instability. A ruptured aortic aneurysm with pseudoaneurysm may mimic a paravertebral abscess, and surgery at the site of an unsuspected aneurysm may precipitate life-threatening hemorrhage. Satisfactory results may be depend on early surgical intervention for a mycotic aneurysm and also are related to host immunity. PMID- 15662332 TI - Lower back pain and mass in a 13-year-old girl. PMID- 15662333 TI - A 23-year-old woman with complete paraplegia and anesthesia below the T8 level. PMID- 15662334 TI - Thigh mass in a 73-year-old woman. PMID- 15662335 TI - Unicompartmental knee replacement: introduction: where have we been? Where are we now? Where are we going? PMID- 15662337 TI - Words that serve as pillars. PMID- 15662338 TI - The truth about capillary refill. PMID- 15662339 TI - Jim Page on leadership. PMID- 15662340 TI - Shivers: preheating your scoop stretcher. PMID- 15662342 TI - What stands out in the ECG crowd? PMID- 15662344 TI - 2005 JEMS Platinum Resource Guide. PMID- 15662346 TI - Bioterrorism: EMS response to deadly infections. PMID- 15662347 TI - DNP 2,4-dinitrophenol: a deadly way to lose weight. PMID- 15662351 TI - Electronic & data management issues: avoid scheduling hiccups. PMID- 15662354 TI - Who's minding the kids? PMID- 15662355 TI - Enrolling children in the State Children's Health Insurance Program (S-CHIP): can we do more? PMID- 15662356 TI - Physical abuse: Recognition and reporting. AB - This article provides primary care providers, including pediatric nurse practitioners, with a framework for understanding the dynamics of child abuse, recognizing physical abuse injuries, and reporting concerns of suspected physical abuse to child protective services. Three children die in America every day as a result of child abuse or neglect. Many children who have severe injuries at the time that physical abuse is diagnosed have previously presented with less severe injuries, and physical abuse was overlooked. Physical assessment for children presenting with bruises, bite marks, burns, skeletal injuries, abdominal trauma, and head injuries will be discussed. Prompt recognition and reporting of physical abuse injuries by primary care providers is imperative for the protection of children. PMID- 15662357 TI - The lactation consult: problem solving, teaching, and support for the breastfeeding family. AB - Requests from parents for lactation support have increased significantly in primary care settings. Pediatric nurse practitioners with the ability to assess the newborn as well as the breastfeeding couple are a valuable asset. This article seeks to identify early predictors of breastfeeding problems and how to perform a lactation consultation to address these concerns. Good infant weight gain and maternal comfort while nursing are described as key evaluation criteria when assessing the nursing couple. The importance of achieving a good latch at the breast is stressed. A step-by-step guide to both the lactation consultation and common latch-on techniques are reviewed. PMID- 15662358 TI - Primary care provider interventions for the delayed disclosure of adolescent sexual assault. AB - Acute sexual assault is a serious and underreported crime with the potential for causing grave physical and emotional harm to its victim. As a result of developmental and psychological factors, the adolescent victim may delay the disclosure of such an assault and therefore experience detrimental, acute, and long-term effects. By understanding the reasons for delayed disclosure and integrating this with currently established guidelines for acutely assaulted patients, primary care providers can better tailor the care they provide when faced with the delayed disclosure of adolescent sexual assault. Furthermore, based on this review, it becomes clear that standardized protocols are necessary to more efficiently care for these patients. Recommendations are provided to allow tailoring of primary care provider's interventions based on established protocols and new understandings when caring for adolescents who delay the disclosure of their sexual assault. PMID- 15662359 TI - Profile of a primary care practice asthma program: improved patient outcomes in a high-risk population. AB - Asthma touches all of our lives. Asthma is a disease entity of huge proportions nationally and locally. It is the most common cause of school absences from a chronic disease and a common reason for parents and caregivers to miss work. The purpose of this article is to provide pediatric nurse practitioners with an understanding of the impact of asthma on children, their families, and on clinical practice. It will examine an asthma program created in a pediatric primary care setting to treat a high-risk population that proved very successful. The discussion of these issues incorporates results from a 2-year clinical project that focused on the reduction of asthma-related emergency department visits, asthma-related hospitalizations, and asthma-related missed school days. PMID- 15662360 TI - Weight and weight concerns: are they associated with reported depressive symptoms in adolescents? AB - INTRODUCTION: The primary purpose of this study was to examine the incidence of obesity, perception of weight, and weight management behaviors in a nationwide sample of adolescents. Further, the relationship between these variables and the adolescents' self-reported depressive symptoms was also explored. METHOD: Cross sectional data from 16- to 18-year-olds who participated in the 1999 (n=9795) and 2001 (n=8190) Youth Risk Behavioral Surveillance System were used for this secondary analysis. RESULTS: No relationship was found between depressive symptoms and body mass index. However, perception of weight as either underweight or overweight was related to higher depressive symptoms. The risk profile for reporting depressive symptoms includes use of any weight control strategies, female, and perception of weight. DISCUSSION: These findings underscore the importance of addressing adolescents' perceptions of weight in relation to their actual body mass index and psychological well-being. PMID- 15662361 TI - EMLA for painful procedures in infants. AB - OBJECTIVES: Based on the content of the article, you will be able to: Identify the two drugs in EMLA and the youngest pediatric age for its approved use. List the procedures for which EMLA has been used to control pain in infants. Describe the efficacy of EMLA vs. other agents in treating pain. Discuss the potential adverse effects of EMLA in infants. See page 48 for instructions. PMID- 15662363 TI - A new approach to treating acute otitis media. PMID- 15662364 TI - Boy with an unwanted travel companion. PMID- 15662365 TI - Educating parents about portion sizes for preschoolers. PMID- 15662366 TI - Secondary analysis of data. PMID- 15662367 TI - Perceptions of battered women. PMID- 15662368 TI - The role of models in understanding CD8+ T-cell memory. AB - Immunological memory - the ability to 'remember' previously encountered pathogens and respond faster on re-exposure - is a central feature of the immune response of vertebrates. We outline how mathematical models have contributed to our understanding of CD8(+) T-cell memory. Together with experimental data, models have helped to quantitatively describe and to further our understanding of both the generation of memory after infection with a pathogen and the maintenance of this memory throughout the life of an individual. PMID- 15662369 TI - Host persistence: exploitation of anti-inflammatory pathways by Toxoplasma gondii. AB - Hosts that are infected with Toxoplasma gondii must mount a powerful immune response to contain dissemination of the parasite and to prevent mortality. After parasite proliferation has been contained by interferon-gamma-dependent responses, the onset of the chronic phase of infection is characterized by continuous cell-mediated immunity. Such potent responses are kept under tight control by a class of anti-inflammatory eicosanoid, the lipoxins. Here, we review such immune-containment strategies from the perspective of the host, which attempts to keep pro-inflammatory responses under control during chronic disease, as well as from the perspective of the pathogen, which hijacks the lipoxygenase machinery of the host for its own advantage, probably as an immune-escape mechanism. PMID- 15662370 TI - Evolutionary genomics: codon bias and selection on single genomes. AB - The idea that natural selection on genes might be detected using only a single genome has been put forward by Plotkin and colleagues, who present a method that they claim can detect selection without the need for comparative data and which, if correct, would confer greater power of analysis with less information. Here we argue that their method depends on assumptions that confound their conclusions and that, even if these assumptions were valid, the authors' inferences about adaptive natural selection are unjustified. PMID- 15662371 TI - Evolutionary genomics: codon volatility does not detect selection. AB - Plotkin et al. introduce a method to detect selection that is based on an index called codon volatility and that uses only the sequence of a single genome, claiming that this method is applicable to a large range of sequenced organisms. Volatility for a given codon is the ratio of non-synonymous codons to all sense codons accessible by one point mutation. The significance of each gene's volatility is assessed by comparison with a simulated distribution of 10(6) synonymous versions of each gene, with synonymous codons drawn randomly from average genome frequencies. Here we re-examine their method and data and find that codon volatility does not detect selection, and that, even if it did, the genomes of Mycobacterium tuberculosis and Plasmodium falciparum, as well as those of most sequenced organisms, do not meet the assumptions necessary for application of their method. PMID- 15662374 TI - Tales of the unexpected. PMID- 15662372 TI - Evolutionary genomics: detecting selection needs comparative data. AB - Positive selection at the molecular level is usually indicated by an increase in the ratio of non-synonymous to synonymous substitutions (dN/dS) in comparative data. However, Plotkin et al. describe a new method for detecting positive selection based on a single nucleotide sequence. We show here that this method is particularly sensitive to assumptions regarding the underlying mutational processes and does not provide a reliable way to identify positive selection. PMID- 15662375 TI - Einstein is dead. PMID- 15662376 TI - Titan team claims just deserts as probe hits moon of creme brulee. PMID- 15662377 TI - All parties on edge as NIH delays open-access briefing. PMID- 15662378 TI - Georgia court bans biology textbook stickers. PMID- 15662379 TI - Pasteur board quits in bid to resolve crisis at troubled institute. PMID- 15662381 TI - Antinuclear groups push to keep treaty review in the air. PMID- 15662380 TI - Indian Ocean fault line poses threat of further earthquakes. PMID- 15662382 TI - Science lobby urges UK to divert funds from military fields. PMID- 15662383 TI - Brain-scan ethics come under the spotlight. PMID- 15662385 TI - All pain, no gain? PMID- 15662386 TI - The premier division. PMID- 15662387 TI - Alternative views of amphibian toe-clipping. PMID- 15662388 TI - Insect collection ready to spread its wings. PMID- 15662394 TI - Schrodinger's mousetrap. Part 1. PMID- 15662389 TI - Croatian minister rejects 'meddling' claim. PMID- 15662395 TI - Endocrinology: fertility hormone in repose. PMID- 15662396 TI - Climatology: will soil amplify climate change? PMID- 15662397 TI - Evolution: a taste for mimicry. PMID- 15662398 TI - Astronomy: weighing the baby. PMID- 15662399 TI - Signal transduction: a new canon. PMID- 15662402 TI - Animal mimicry: choosing when to be a cleaner-fish mimic. AB - Mimicry in vertebrates is usually a permanent state--mimics resemble and normally accompany their model throughout the life stages during which they act as mimics. Here we show that the bluestriped fangblenny fish (Plagiotremus rhinorhynchos), which aggressively attacks other coral-reef fish, can turn off the mimetic colours that disguise it as the benign bluestreak cleaner wrasse, Labroides dimidiatus, and assume a radically different appearance. This opportunistic facultative mimicry extends the fangblenny's scope by allowing it to blend into shoals of small reef fish as well as to remain inconspicuous at cleaning stations. PMID- 15662403 TI - Behavioural ecology: transient sexual mimicry leads to fertilization. AB - Sexual mimicry among animals is widespread, but does it impart a fertilization advantage in the widely accepted 'sneak-guard' model of sperm competition? Here we describe field results in which a dramatic facultative switch in sexual phenotype by sneaker-male cuttlefish leads to immediate fertilization success, even in the presence of the consort male. These results are surprising, given the high rate at which females reject copulation attempts by males, the strong mate guarding behaviour of consort males, and the high level of sperm competition in this complex mating system. PMID- 15662405 TI - 1905 and all that. PMID- 15662406 TI - Einstein as icon. PMID- 15662407 TI - Brownian motion. PMID- 15662408 TI - In and out of equilibrium. AB - Albert Einstein's work on brownian motion showed how thermal equilibrium could be brought about by work exchanged through thermal fluctuations and viscous dissipation. Glasses are out-of-equilibrium systems in which this exchange happens at widely different timescales simultaneously. Theory then suggests the fascinating possibility that such behaviour may lead to a more general form of thermalization, in which the effective temperature shared by all components differs at each timescale. PMID- 15662409 TI - Quantum criticality. AB - As we mark the centenary of Albert Einstein's seminal contribution to both quantum mechanics and special relativity, we approach another anniversary--that of Einstein's foundation of the quantum theory of solids. But 100 years on, the same experimental measurement that puzzled Einstein and his contemporaries is forcing us to question our understanding of how quantum matter transforms at ultra-low temperatures. PMID- 15662410 TI - Happy centenary, photon. AB - One hundred years ago Albert Einstein introduced the concept of the photon. Although in the early years after 1905 the evidence for the quantum nature of light was not compelling, modern experiments--especially those using photon pairs -have beautifully confirmed its corpuscular character. Research on the quantum properties of light (quantum optics) triggered the evolution of the whole field of quantum information processing, which now promises new technology, such as quantum cryptography and even quantum computers. PMID- 15662411 TI - In search of symmetry lost. AB - Powerful symmetry principles have guided physicists in their quest for nature's fundamental laws. The successful gauge theory of electroweak interactions postulates a more extensive symmetry for its equations than are manifest in the world. The discrepancy is ascribed to a pervasive symmetry-breaking field, which fills all space uniformly, rendering the Universe a sort of exotic superconductor. So far, the evidence for these bold ideas is indirect. But soon the theory will undergo a critical test depending on whether the quanta of this symmetry-breaking field, the so-called Higgs particles, are produced at the Large Hadron Collider (due to begin operation in 2007). PMID- 15662412 TI - The state of the Universe. AB - The past 20 years have seen dramatic advances in cosmology, mostly driven by observations from new telescopes and detectors. These instruments have allowed astronomers to map out the large-scale structure of the Universe and probe the very early stages of its evolution. We seem to have established the basic parameters describing the behaviour of our expanding Universe, thereby putting cosmology on a firm empirical footing. But the emerging 'standard' model leaves many details of galaxy formation still to be worked out, and new ideas are emerging that challenge the theoretical framework on which the structure of the Big Bang is based. There is still a great deal left to explore in cosmology. PMID- 15662413 TI - A theory of everything? PMID- 15662414 TI - A theoretical look at the direct detection of giant planets outside the Solar System. AB - Astronomy is at times a science of unexpected discovery. When it is, and if we are lucky, new intellectual territories emerge to challenge our views of the cosmos. The recent indirect detections using high-precision Doppler spectroscopy of more than 100 giant planets orbiting more than 100 nearby stars is an example of such rare serendipity. What has been learned has shaken out preconceptions, for none of the planetary systems discovered so far is like our own. The key to unlocking a planet's chemical, structural, and evolutionary secrets, however, is the direct detection of the planet's light. Because there have been as yet no confirmed detections, a theoretical analysis of such a planet's atmosphere is necessary for guiding our search. PMID- 15662415 TI - Structure of human follicle-stimulating hormone in complex with its receptor. AB - Follicle-stimulating hormone (FSH) is central to reproduction in mammals. It acts through a G-protein-coupled receptor on the surface of target cells to stimulate testicular and ovarian functions. We present here the 2.9-A-resolution structure of a partially deglycosylated complex of human FSH bound to the extracellular hormone-binding domain of its receptor (FSHR(HB)). The hormone is bound in a hand clasp fashion to an elongated, curved receptor. The buried interface of the complex is large (2,600 A2) and has a high charge density. Our analysis suggests that all glycoprotein hormones bind to their receptors in this mode and that binding specificity is mediated by key interaction sites involving both the common alpha- and hormone-specific beta-subunits. On binding, FSH undergoes a concerted conformational change that affects protruding loops implicated in receptor activation. The FSH-FSHR(HB) complexes form dimers in the crystal and at high concentrations in solution. Such dimers may participate in transmembrane signal transduction. PMID- 15662416 TI - Role of the proto-oncogene Pokemon in cellular transformation and ARF repression. AB - Aberrant transcriptional repression through chromatin remodelling and histone deacetylation has been postulated to represent a driving force underlying tumorigenesis because histone deacetylase inhibitors have been found to be effective in cancer treatment. However, the molecular mechanisms by which transcriptional derepression would be linked to tumour suppression are poorly understood. Here we identify the transcriptional repressor Pokemon (encoded by the Zbtb7 gene) as a critical factor in oncogenesis. Mouse embryonic fibroblasts lacking Zbtb7 are completely refractory to oncogene-mediated cellular transformation. Conversely, Pokemon overexpression leads to overt oncogenic transformation both in vitro and in vivo in transgenic mice. Pokemon can specifically repress the transcription of the tumour suppressor gene ARF through direct binding. We find that Pokemon is aberrantly overexpressed in human cancers and that its expression levels predict biological behaviour and clinical outcome. Pokemon's critical role in cellular transformation makes it an attractive target for therapeutic intervention. PMID- 15662417 TI - A dynamical calibration of the mass-luminosity relation at very low stellar masses and young ages. AB - Mass is the most fundamental parameter of a star, yet it is also one of the most difficult to measure directly. In general, astronomers estimate stellar masses by determining the luminosity and using the 'mass-luminosity' relationship, but this relationship has never been accurately calibrated for young, low-mass stars and brown dwarfs. Masses for these low-mass objects are therefore constrained only by theoretical models. A new high-contrast adaptive optics camera enabled the discovery of a young (50 million years) companion only 0.156 arcseconds (2.3 au) from the more luminous (> 120 times brighter) star AB Doradus A. Here we report a dynamical determination of the mass of the newly resolved low-mass companion AB Dor C, whose mass is 0.090 +/- 0.005 solar masses. Given its measured 1-2 micrometre luminosity, we have found that the standard mass-luminosity relations overestimate the near-infrared luminosity of such objects by about a factor of approximately 2.5 at young ages. The young, cool objects hitherto thought to be substellar in mass are therefore about twice as massive, which means that the frequency of brown dwarfs and planetary mass objects in young stellar clusters has been overestimated. PMID- 15662418 TI - High-velocity streams of dust originating from Saturn. AB - High-velocity submicrometre-sized dust particles expelled from the jovian system have been identified by dust detectors on board several spacecraft. On the basis of periodicities in the dust impact rate, Jupiter's moon Io was found to be the dominant source of the streams. The grains become positively charged within the plasma environment of Jupiter's magnetosphere, and gain energy from its co rotational electric field. Outside the magnetosphere, the dynamics of the grains are governed by the interaction with the interplanetary magnetic field that eventually forms the streams. A similar process was suggested for Saturn. Here we report the discovery by the Cassini spacecraft of bursts of high-velocity dust particles (> or = 100 km s(-1)) within approximately 70 million kilometres of Saturn. Most of the particles detected at large distances appear to originate from the outskirts of Saturn's outermost main ring. All bursts of dust impacts detected within 150 Saturn radii are characterized by impact directions markedly different from those measured between the bursts, and they clearly coincide with the spacecraft's traversals through streams of compressed solar wind. PMID- 15662419 TI - Stable sea surface temperatures in the western Pacific warm pool over the past 1.75 million years. AB - About 850,000 years ago, the period of the glacial cycles changed from 41,000 to 100,000 years. This mid-Pleistocene climate transition has been attributed to global cooling, possibly caused by a decrease in atmospheric carbon dioxide concentrations. However, evidence for such cooling is currently restricted to the cool upwelling regions in the eastern equatorial oceans, although the tropical warm pools on the western side of the ocean basins are particularly sensitive to changes in radiative forcing. Here we present high-resolution records of sea surface temperatures spanning the past 1.75 million years, obtained from oxygen isotopes and Mg/Ca ratios in planktonic foraminifera from the western Pacific warm pool. In contrast with the eastern equatorial regions, sea surface temperatures in the western Pacific warm pool are relatively stable throughout the Pleistocene epoch, implying little long-term change in the tropical net radiation budget. Our results challenge the hypothesis of a gradual decrease in atmospheric carbon dioxide concentrations as a dominant trigger of the longer glacial cycles since 850,000 years ago. Instead, we infer that the temperature contrast across the equatorial Pacific Ocean increased, which might have had a significant influence on the mid-Pleistocene climate transition. PMID- 15662420 TI - Long-term sensitivity of soil carbon turnover to warming. AB - The sensitivity of soil carbon to warming is a major uncertainty in projections of carbon dioxide concentration and climate. Experimental studies overwhelmingly indicate increased soil organic carbon (SOC) decomposition at higher temperatures, resulting in increased carbon dioxide emissions from soils. However, recent findings have been cited as evidence against increased soil carbon emissions in a warmer world. In soil warming experiments, the initially increased carbon dioxide efflux returns to pre-warming rates within one to three years, and apparent carbon pool turnover times are insensitive to temperature. It has already been suggested that the apparent lack of temperature dependence could be an artefact due to neglecting the extreme heterogeneity of soil carbon, but no explicit model has yet been presented that can reconcile all the above findings. Here we present a simple three-pool model that partitions SOC into components with different intrinsic turnover rates. Using this model, we show that the results of all the soil-warming experiments are compatible with long-term temperature sensitivity of SOC turnover: they can be explained by rapid depletion of labile SOC combined with the negligible response of non-labile SOC on experimental timescales. Furthermore, we present evidence that non-labile SOC is more sensitive to temperature than labile SOC, implying that the long-term positive feedback of soil decomposition in a warming world may be even stronger than predicted by global models. PMID- 15662421 TI - Early Pliocene hominids from Gona, Ethiopia. AB - Comparative biomolecular studies suggest that the last common ancestor of humans and chimpanzees, our closest living relatives, lived during the Late Miocene Early Pliocene. Fossil evidence of Late Miocene-Early Pliocene hominid evolution is rare and limited to a few sites in Ethiopia, Kenya and Chad. Here we report new Early Pliocene hominid discoveries and their palaeoenvironmental context from the fossiliferous deposits of As Duma, Gona Western Margin (GWM), Afar, Ethiopia. The hominid dental anatomy (occlusal enamel thickness, absolute and relative size of the first and second lower molar crowns, and premolar crown and radicular anatomy) indicates attribution to Ardipithecus ramidus. The combined radioisotopic and palaeomagnetic data suggest an age of between 4.51 and 4.32 million years for the hominid finds at As Duma. Diverse sources of data (sedimentology, faunal composition, ecomorphological variables and stable carbon isotopic evidence from the palaeosols and fossil tooth enamel) indicate that the Early Pliocene As Duma sediments sample a moderate rainfall woodland and woodland/grassland. PMID- 15662422 TI - Definitive fossil evidence for the extant avian radiation in the Cretaceous. AB - Long-standing controversy surrounds the question of whether living bird lineages emerged after non-avian dinosaur extinction at the Cretaceous/Tertiary (K/T) boundary or whether these lineages coexisted with other dinosaurs and passed through this mass extinction event. Inferences from biogeography and molecular sequence data (but see ref. 10) project major avian lineages deep into the Cretaceous period, implying their 'mass survival' at the K/T boundary. By contrast, it has been argued that the fossil record refutes this hypothesis, placing a 'big bang' of avian radiation only after the end of the Cretaceous. However, other fossil data--fragmentary bones referred to extant bird lineages- have been considered inconclusive. These data have never been subjected to phylogenetic analysis. Here we identify a rare, partial skeleton from the Maastrichtian of Antarctica as the first Cretaceous fossil definitively placed within the extant bird radiation. Several phylogenetic analyses supported by independent histological data indicate that a new species, Vegavis iaai, is a part of Anseriformes (waterfowl) and is most closely related to Anatidae, which includes true ducks. A minimum of five divergences within Aves before the K/T boundary are inferred from the placement of Vegavis; at least duck, chicken and ratite bird relatives were coextant with non-avian dinosaurs. PMID- 15662423 TI - Field parameterization and experimental test of the neutral theory of biodiversity. AB - Ecologists would like to explain general patterns observed across multi-species communities, such as species-area and abundance-frequency relationships, in terms of the fundamental processes of birth, death and migration underlying the dynamics of all constituent species. The unified neutral theory of biodiversity and related theories based on these fundamental population processes have successfully recreated general species-abundance patterns without accounting for either the variation among species and individuals or resource-releasing processes such as predation and disturbance, long emphasized in ecological theory. If ecological communities can be described adequately without estimating variation in species and their interactions, our understanding of ecological community organization and the predicted consequences of reduced biodiversity and environmental change would shift markedly. Here, I introduce a strong method to test the neutral theory that combines field parameterization of the underlying population dynamics with a field experiment, and apply it to a rocky intertidal community. Although the observed abundance-frequency distribution of the system follows that predicted by the neutral theory, the neutral theory predicts poorly the field experimental results, indicating an essential role for variation in species interactions. PMID- 15662424 TI - Evolutionary dynamics on graphs. AB - Evolutionary dynamics have been traditionally studied in the context of homogeneous or spatially extended populations. Here we generalize population structure by arranging individuals on a graph. Each vertex represents an individual. The weighted edges denote reproductive rates which govern how often individuals place offspring into adjacent vertices. The homogeneous population, described by the Moran process, is the special case of a fully connected graph with evenly weighted edges. Spatial structures are described by graphs where vertices are connected with their nearest neighbours. We also explore evolution on random and scale-free networks. We determine the fixation probability of mutants, and characterize those graphs for which fixation behaviour is identical to that of a homogeneous population. Furthermore, some graphs act as suppressors and others as amplifiers of selection. It is even possible to find graphs that guarantee the fixation of any advantageous mutant. We also study frequency dependent selection and show that the outcome of evolutionary games can depend entirely on the structure of the underlying graph. Evolutionary graph theory has many fascinating applications ranging from ecology to multi-cellular organization and economics. PMID- 15662425 TI - An autoregulatory circuit for long-range self-organization in Dictyostelium cell populations. AB - Nutrient-deprived Dictyostelium amoebae aggregate to form a multicellular structure by chemotaxis, moving towards propagating waves of cyclic AMP that are relayed from cell to cell. Organizing centres are not formed by founder cells, but are dynamic entities consisting of cores of outwardly rotating spiral waves that self-organize in a homogeneous cell population. Spiral waves are ubiquitously observed in chemical reactions as well as in biological systems. Although feedback control of spiral waves in spatially extended chemical reactions has been demonstrated in recent years, the mechanism by which control is achieved in living systems is unknown. Here we show that mutants of the cyclic AMP/protein kinase A pathway show periodic signalling, but fail to organize coherent long-range wave territories, owing to the appearance of numerous spiral cores. A theoretical model suggests that autoregulation of cell excitability mediated by protein kinase A acts to optimize the number of signalling centres. PMID- 15662426 TI - A pentatricopeptide repeat protein is essential for RNA editing in chloroplasts. AB - RNA editing is a process of RNA maturation involved in the insertion, deletion or modification of nucleotides. In organellar transcripts of higher plants, specific cytidine residues are converted into uridine residues. In many cases, editing results in the restoration of conserved amino acid residues, a process that is essential for protein function in plastids. Despite the technical breakthrough in establishing systems in vivo and in vitro for analysing RNA editing, its machinery still remains to be identified in higher plants. Here we introduce a genetic approach and report the discovery of a gene responsible for the specific RNA editing event in the chloroplast. PMID- 15662428 TI - Closing the gap. PMID- 15662427 TI - Molecular dynamics of cyclically contracting insect flight muscle in vivo. AB - Flight in insects--which constitute the largest group of species in the animal kingdom--is powered by specialized muscles located within the thorax. In most insects each contraction is triggered not by a motor neuron spike but by mechanical stretch imposed by antagonistic muscles. Whereas 'stretch activation' and its reciprocal phenomenon 'shortening deactivation' are observed to varying extents in all striated muscles, both are particularly prominent in the indirect flight muscles of insects. Here we show changes in thick-filament structure and actin-myosin interactions in living, flying Drosophila with the use of synchrotron small-angle X-ray diffraction. To elicit stable flight behaviour and permit the capture of images at specific phases within the 5-ms wingbeat cycle, we tethered flies within a visual flight simulator. We recorded images of 340 micros duration every 625 micros to create an eight-frame diffraction movie, with each frame reflecting the instantaneous structure of the contractile apparatus. These time-resolved measurements of molecular-level structure provide new insight into the unique ability of insect flight muscle to generate elevated power at high frequency. PMID- 15662429 TI - Save now, don't pay later. PMID- 15662431 TI - Scientists and societies. International Institute for Applied Systems Analysis, Austria. PMID- 15662433 TI - Hard times at the FDA. PMID- 15662434 TI - Amplifying Igh translocations. PMID- 15662436 TI - Lymphoid development: it's not 'all Greek to us' any more. PMID- 15662437 TI - No driving without a license. PMID- 15662438 TI - STOP! In the name of positive selection. PMID- 15662439 TI - A move to exclude. PMID- 15662440 TI - T cell receptor CDRs: starring versus supporting roles. PMID- 15662442 TI - Mast cells in the development of adaptive immune responses. AB - Mast cells are so widely recognized as critical effector cells in allergic disorders and other immunoglobulin E-associated acquired immune responses that it can be difficult to think of them in any other context. However, mast cells also can be important as initiators and effectors of innate immunity. In addition, mast cells that are activated during innate immune responses to pathogens, or in other contexts, can secrete products and have cellular functions with the potential to facilitate the development, amplify the magnitude or regulate the kinetics of adaptive immune responses. Thus, mast cells may influence the development, intensity and duration of adaptive immune responses that contribute to host defense, allergy and autoimmunity, rather than simply functioning as effector cells in these settings. PMID- 15662447 TI - Opioid receptors and heart resistance to arrhythmogenic factors. AB - We present detailed data on the role of central and peripheral opioid receptors in the regulation of heart resistance to arrhythmogenic factors. Stimulation of peripheral delta2- and kappa1-receptors increases heart resistance to the arrhythmogenic effect of acute ischemia and reperfusion. Activation of peripheral mu- and delta1-opioid receptors improves electrical stability of the heart in animals with postinfarction cardiosclerosis. Possible mechanisms for opioidergic regulation of heart resistance to arrhythmogenic factors are discussed. PMID- 15662448 TI - Vagal stimulation modifies parameters of heterochromatin in the nuclei of vagosolitary complex neurons of medulla oblongata in rats. AB - New data were obtained on modification of heterochromatin parameters in the nuclei of medulla oblongata neurons in Wistar rats after stimulation of the vagus nerve: decrease in the area of heterochromatin regions and redistribution of chromocenters within the neuronal nuclear system. It was concluded that realization of the viscero-visceral reflex is associated with rearrangement of chromatin in neurons involved in transmission of the corresponding information. PMID- 15662449 TI - Induction of apoptosis in bone marrow cells is mediated via purinergic receptors. AB - ATP activity in mouse bone marrow cells was in vitro estimated by expression of phosphatidylserine on the outer membrane surface using FITC-labeled annexin. ATP induced apoptosis in bone marrow cells. Purinergic receptor antagonists PPADS and suramin modulated the apoptotic effect of ATP on hemopoietic cells. Acute and subacute administration of doxorubicin, an inductor of oxidative burst, decreased cell sensitivity to ATP and abolished its apoptotic effect. PMID- 15662450 TI - Spectral parameters of phrenic nerve activity in mature rats during electrical stimulation of retrotrapezoid nucleus. AB - In mature rats, electrical stimulation of the retrotrapezoid nucleus increased the amplitude and frequency of the high-frequency peak in the firing spectrum of the phrenic nerve, while the amplitude of medium-frequency peak and the amplitude ratio of medium- to high-frequency peaks decreased. These changes in spectral parameters were associated with accelerated increase in central inspiratory activity, decreased amplitude of phrenic nerve firing, and increased frequency of respiratory rhythm. It is hypothesized that being a relay structure of central chemosensitive mechanism, the retrotrapezoid nucleus regulates parameters of medium- and high-frequency spectral peaks of efferent electrical activity in the respiratory center together with the dorsal and ventral respiratory groups. PMID- 15662451 TI - Histidine increases beta-adrenoreactivity of myocardium in frogs. AB - Histidine (3x10(-5) g/ml) had no effect on contractility and chronoinotropic relationships in frog myocardium, but rapidly and reversibly increased myocardial beta-adrenoreactivity (increased myocardial response to 7x10(-8), 3x10(-7), and 4x10(-6) g/ml epinephrine) and potentiated the positive effect of epinephrine (7x10(-8), 3x10(-7) g/ml) on chronoinotropic relationships in the myocardium. Histidine is considered to be a component of endogenous sensitizer of beta adrenoceptors in human blood modulating the function of cardiomyocytes. PMID- 15662452 TI - Serotoninergic nervous system in intact heart and abdominal organs. AB - The participation of serotoninergic nervous system in the synergistic inhibitory effect of different subdivisions of the autonomic nervous system on the heart and their stimulatory effect on smooth muscle organs of the biliary excretion system, stomach, and small intestine were studied on rabbits under normal conditions. PMID- 15662453 TI - Maintenance of equilibrium during tracing eye movements. AB - Horizontal tracing movements of eyes modify the type of vertical posture maintenance decreasing the role of the lower segment in the regulation of the position of the pressure center. The relationship between fluctuations of the pressure center in the frontal and sagittal planes increases. Periodicity of eye movements corresponds to fluctuations of the pressure center and these signals were phase shifted relatively to each other. PMID- 15662454 TI - Ratio between the contents of 11-dehydrocorticosterone and corticosterone after acute and repeated stress: effect of dehydroepiandrosterone sulfate. AB - Acute stress was accompanied by reduction of 11-dehydrocorticosterone to corticosterone in male rats. The reverse reaction predominated during repeated stress and increased after administration of dehydroepiandrosterone sulfate. Treatment with mu-opioid receptor antagonist naltrexone in a dose of 0.1 mg/kg 20 min before administration of dehydroepiandrosterone sulfate abolished this effect. PMID- 15662455 TI - Stress response and apoptosis in pro- and antiinflammatory macrophages. AB - We showed that stress response and apoptosis in macrophages depend on the phenotype of their secretory activity and specific biological and physical characteristics of the factor inducing stress-response or apoptosis. PMID- 15662456 TI - Energostim in therapy of chronic ischemia of the lower limbs. AB - Positive changes were more pronounced in patients with chronic ischemia of the lower limbs treated with energostim alone and in combination with trental in comparison with patients receiving trental monotherapy. The best effect was attained in patients treated with energostim in combination with trental. PMID- 15662457 TI - Reactions and functional relationships between nonrespiratory neurons in the medulla oblongata after central application of penicillin. AB - Changes in impulse activity of nonrespiratory neurons in the medulla oblongata produced by central administration of penicillin were studied in acute experiments on narcotized immobilized rats. The mean firing frequency increased in most neurons. The peaks on histograms for the distribution of interspike intervals were shifted toward shorter intervals and their amplitude increased; the type of distribution was also changed. Tonic activity of neurons was transformed into burst activity. Study of auto- and cross-correlation histograms for neuronal pairs showed that hyperactivation of structures was accompanied by an increase in the degree of synchronization. These changes reflect the appearance of new functional relationships between neurons in the respiratory center. We found that nonrespiratory reticular neurons are involved in the mechanisms of normal and pathological respiratory rhythm generation and serve as a functionally labile component of the neuronal respiratory network. PMID- 15662458 TI - Pyroglutamyl-asparagine amide normalizes long-term potentiation in rat hippocampal slices. AB - Preapplication of peptide piracetam analogue pyroglutamyl-asparagine amide to rat hippocampal slices facilitates long-term potentiation of focal responses in the CA1 field after weak tetanization of the synaptic input (30 pulses, 100 Hz). This treatment normalized the development of long-term potentiation after standard tetanization (100 pulses, 100 Hz) impaired by ethanol. PMID- 15662459 TI - Comparative analysis of accumulation of chlorine e6 and hematoporphyrin derivatives in subpopulations of peripheral blood lymphocytes. AB - We studied accumulation of porphyrin photosensitizers chlorine e6, hematoporphyrin, and their derivatives by different lymphocyte subpopulations. The intensity of staining of B lymphocytes and natural killer cells with photosensitizers was higher compared to T lymphocytes. T cell subpopulation differed by their ability to bind photosensitizers. Relative accumulation of dimethyl esters of chlorine e6 and hematoporphyrin in cells surpassed that of nonesterified porphyrins. PMID- 15662460 TI - A new property of endogenous immunostimulator Taftsin. AB - Taftsin, an endogenous peptide immunostimulator, exhibits also high antiulcer activity. In a dose of 0.6 micromol/kg Taftsin decreases the area of ulcerative lesions in albino rats on various models of ulceration. PMID- 15662461 TI - Study of mutagenic activity of dioxidine by the polyorgan micronuclear method. AB - Antibacterial preparation dioxidine administered four times in doses of 10, 100, and 300 mg/kg increased the incidence of micronucleated cells in the bone marrow, lungs, and large intestine of mice. Bone marrow cells were most sensitive, while cells of the lungs and large intestine exhibited lower sensitivity to the cytogenetic effect of dioxidine. PMID- 15662462 TI - Ultrastructural changes in small intestinal lipofibroblasts of suckling rabbits with experimental cholera. AB - Ultrastructural analysis of the jejunum in suckling rabbits showed that lipofibroblasts localized in the submucosa and adjacent to crypts contain lipid inclusions (granules) with typical "melting" surface. Lipofibroblasts contained moderately widened cisternae of the granular endoplasmic reticulum and few mitochondria with dense matrix and poorly developed cristae. Experimental cholera was usually accompanied by a decrease in the number of lipid inclusions, and only in some cases by accumulation of lipid material. Our results suggest that the material accumulated in granules plays a role in the pathogenesis of cholera. PMID- 15662463 TI - Effect of trekrezan on immunogenesis under experimental conditions. AB - We studied the dynamics of immune reactions to trekrezan in mouse pups after vaccination of adult mice during pregnancy and mouse pups at different terms after birth with Salmonella typhimurium 34-96. The preparation modulated activity of the immune system and stimulated antibody production in the pre- and postnatal period. PMID- 15662464 TI - In vitro genotoxic and cytotoxic effects of protein somatic products from helminths on donor blood lymphocytes. AB - Protein somatic products of adult helminths Hymenolepis nana and Toxocara canis and secretory-excretory somatic product of Trichinella spiralis larvae in vitro produced genotoxic and cytotoxic effects on donor blood lymphocytes, which manifested in accumulation of single-chain breaks, alkaline-labile sites in nuclear DNA, and apoptotic cells. This effect depended on the concentration of parasitic protein products during coculturing. PMID- 15662465 TI - Conformation changes in albumin molecule as a marker of dissemination of the tumor process. AB - In 20 patients with stomach cancer the total and effective concentrations of blood albumin were measured using a fluorescent probe and albumin binding reserve was calculated. These parameters in two groups of patients (with local and disseminated process) differed significantly. Evaluation of the effective concentration and albumin binding reserve in cancer patients gives additional valuable information about patient's status. The effective concentration of albumin below 30 g/liter and albumin binding reserve below 70% can be regarded as a prognostically unfavorable sign indicating more extensive dissemination of the tumor process. PMID- 15662466 TI - Disorders in restructuring of T-cell receptor gamma-chain in malignant skin lymphomas. AB - Disorders in restructuring of T-cell receptor gamma-chain in DNA from skin biopsy specimens and peripheral blood lymphocyte of patients with malignant skin lymphomas at different stages of the disease were detected by PCR. Nucleotide sequences of monoclonal T-cell receptors were determined. The study showed that DNA sequences from the skin and lymphocytes did not coincide in some cases. PMID- 15662467 TI - Morphofunctional characteristic of mast cells in BALB/c and C57Bl/6 mice during cold exposure. AB - Mast cells of the mesentery and subcutaneous tissue in BALB/c and C57Bl/6 mice were studied after single and repeated cold exposure (-20 degrees C, 3 min). Immediate adaptive reactions of mast cells in BALB/c and C57Bl/6 mice did not differ after single cold exposure and were manifested in increased degranulation. Repeated cold exposure of BALB/c mice was followed by an adaptive reaction, which included an increase in the count of mast cells in subcutaneous tissue and normalization of the degranulation index. In C57Bl/6 mice the count of mast cells in subcutaneous tissue decreased, while the degranulation index remained high. These changes reflect the disadaptive response of mast cells to repeated cold exposure. PMID- 15662468 TI - Effect of antithrombin III on local hemostasis in the kidneys during experimental nephritis. AB - Administration of antithrombin III-enriched plasma to rabbits with acute Masugi nephritis inhibited prothrombinase formation and increased the release of component C3 from the kidneys. This treatment had a cytoprotective effect and was probably followed by dissociation of antigen--antibody complexes. PMID- 15662469 TI - Effect of the age of pregnant females on brain development in the offspring. AB - We examined offspring of 9-10 and 3.5-4 month-old female rats. Female offspring (14, 21, 28, 35, and 40 days) of old rats had higher body weight than offspring of young animals. No intergroup differences were revealed in the body weight of male offspring. At the age of 40 days the offspring of old females differed from the offspring of young rats by higher absolute weight of the brain (females), lower size of ganglionic neurons in the parietal lobe (males and females), and lower blood testosterone concentration (males). Thirty-day-old offspring of old rats exhibited higher locomotor activity and lower degree of anxiety compared to the offspring of young animals. PMID- 15662470 TI - In vitro effect of acetylcholine on function of sinoatrial node in rat heart. AB - Electrophysiological parameters of true pacemakers in the sinoatrial node of rat heart were recorded intracellularly using glass microelectrodes. In 11 of 13 experiments acetylcholine in increasing doses did not induce migration of the dominant pacemaker region, while in two cases its minor migration upstream the sinus node artery was observed. PMID- 15662471 TI - New data on the involvement of ectodermal epithelium in extrathymic differentiation of human T-lymphocytes. AB - Thymalin accumulates only in young cells of the skin ectodermal epithelium and mucosa of the anterior portions of the digestive and respiratory systems. After appearance of keratin the cells no longer accumulate thymalin. Similarly as in the thymus, thymalin content in the ectodermal epithelium of the organs is subjected to age-associated involution. PMID- 15662472 TI - Differences in modification of stress mechanisms in rat pups exposed to continuous and intermittent maternal deprivation. AB - During antenatal short-term intermittent maternal deprivation the content of epinephrine increased by 123% in the cortex and by 135% in the hypothalamus of 15 day-old rat pups, which is higher than during physiological development, while dopamine content, instead of increasing, decreased by 24% in the cortex and by 35% in the hypothalamus on day 15 of life. Shifts in norepinephrine and dopamine concentrations in the cortex were less pronounced in rat pups exposed to permanent maternal deprivation compared to intermittent deprivation, while the shifts in the hypothalamus were even more pronounced. Presumably, discoordination of the dopaminergic and sympathoadrenal systems in the course of development of the nervous system underlies behavioral disorders in rat pups under conditions of maternal deprivation. PMID- 15662473 TI - Histological method for evaluation of the efficiency of Enerlit-Clima. AB - We propose a method of evaluation of anticlimacteric efficiency of a drug by its effect on the estrous cycle. The study was carried out on 9-month-old mice with retained, but notably reduced reproductive function. Analysis of the cell components of the estrous cycle was carried out on histological preparations of vaginal smears. PMID- 15662474 TI - Active control and its stress effects. AB - The effects of 5-fold injections of normal saline were studied in Wistar rats in two conditioned reflex situations: defense and alimentary. Intact animals developed conditioned pain avoidance reflex in repeated tail-flick test after 60 presentations over 3 days. Two injections of normal saline were sufficient for the formation of a stable negative stress status in experimental rats, which manifested by a drastic increase in tactile sensitivity persisting for 5 days after discontinuation of injections. Only 15% experimental rats vs. 40% intact animals developed the food-procuring response in a multialternative maze. Presumably, repeated injections of normal saline should be regarded as a potent negative stress factor modulating animals behavior, and these signs should be taken into consideration, when interpreting the effects observed in pharmacological investigations. PMID- 15662475 TI - Simulation of electrolyte nephropathy in rats. AB - Stages of modeling of polyetiological electrolyte nephropathy in rats are described. Morphological study confirmed the presence of interstitial and tubular disorders in rat kidneys. The model is intended for studies of the mechanisms underlying the formation, development, and correction of nephrological diseases. PMID- 15662476 TI - Scoring CT/HRCT findings among asbestos-exposed workers: effects of patient's age, body mass index and common laboratory test results. AB - We studied the effects of age, body mass index (BMI) and some common laboratory test results on several pulmonary CT/HRCT signs. Five hundred twenty-eight construction workers (age 38-80, mean 63 years) were imaged with spiral and high resolution CT. Images were scored by three radiologists for solitary pulmonary nodules, signs indicative of fibrosis and emphysema, ground glass opacities, bronchial wall thickness and bronchiectasis. Multivariate statistical analyses were adjusted for smoking and asbestos exposure. Increasing age, blood haemoglobin value and erythrocyte sedimentation rate correlated positively with several HRCT signs. Increasing BMI was associated with a decrease in several signs, especially parenchymal bands, honeycombing, all kinds of emphysema and bronchiectasis. The latter finding might be due to the suboptimal image quality in obese individuals, which may cause suspicious findings to be overlooked. Background data, including patient's age and body constitution, should be considered when CT/HRCT images are interpreted. PMID- 15662477 TI - Well-differentiated adenocarcinoma of the gallbladder with intratumoral cystic components due to abundant mucin production: a mimicker of adenomyomatosis. AB - The prevalence and etiology of the cystic components within gallbladder carcinomas as seen on MR images were evaluated. A retrospective review of MR images was performed for 35 proven gallbladder carcinomas in search of radiologically detectable intratumoral cystic components. The pathologic specimens were meticulously reviewed to determine the etiology. MR images of 31 adenomyomatoses were also reviewed for comparison to clarify the difference in MR features between these two entities. Three cases out of 35 proven gallbladder carcinomas were found to have intratumoral cystic components. They were all well differentiated adenocarcinomas, and the cystic components consisted of dilated neoplastic glands filled with abundant mucin pool. Adenomyomatosis tended to have more and rounded cystic components (Rokitansky-Aschoff sinuses) lined in a linear fashion and were flat-elevated in shape, smaller in size and had a regular surface, as compared to the three carcinomas. Although rare, radiologists need to be aware that well-differentiated gallbladder carcinoma with mucin production can have cystic components, which may mimic adenomyomatosis. Careful interpretation of MR images may provide useful information in the differentiation of these two entities. PMID- 15662478 TI - CT and MR findings of a retrorectal cystic hamartoma confused with an adnexal mass on ultrasound. AB - We describe the imaging features of a tailgut cyst mistaken for an adnexal mass. A pelvic ultrasound in a 28-year-old woman showed a 10-cm hypoechoic left pelvic mass. Having not seen the left ovary, the radiologist concluded that the mass was an endometrioma. CT disclosed a retrorectal cystic lesion with wall calcifications and internal septa. MR confirmed the extra-ovarian location of the tumor, which was hyperintense on T2-weighted images and had an intermediate signal on T1-weighted images. Surgery revealed a retrorectal cystic hamartoma. Radiological diagnosis of a tailgut cyst requires first correct localization of the tumor and then differentiation from other retrorectal masses. PMID- 15662479 TI - Flat panel computed tomography of human ex vivo heart and bone specimens: initial experience. AB - The aim of this technical investigation was the detailed description of a prototype flat panel detector computed tomography system (FPCT) and its initial evaluation in an ex vivo setting. The prototype FPCT scanner consists of a conventional radiographic flat panel detector, mounted on a multi-slice CT scanner gantry. Explanted human ex vivo heart and foot specimens were examined. Images were reformatted with various reconstruction algorithms and were evaluated for high-resolution anatomic information. For comparison purposes, the ex vivo specimens were also scanned with a conventional 16-detector-row CT scanner (Sensation 16, Siemens Medical Solutions, Forchheim, Germany). With the FPCT prototype used, a 1,024x768 resolution matrix can be obtained, resulting in an isotropic voxel size of 0.25x0.25x0.25 mm at the iso-center. Due to the high spatial resolution, very small structures such as trabecular bone or third degree, distal branches of coronary arteries could be visualized. This first evaluation showed that flat panel detector systems can be used in a cone-beam computed tomography scanner and that very high spatial resolutions can be achieved. However, there are limitations for in vivo use due to constraints in low contrast resolution and slow scan speed. PMID- 15662480 TI - Percutaneous vertebroplasty immediately relieves pain of osteoporotic vertebral compression fractures and prevents prolonged immobilization of patients. AB - To assess the immediate efficacy of percutaneous vertebroplasty (PVP) in relief of pain and improving mobility of patients with vertebral compression fractures (VCF) secondary to osteoporosis, 205 cases (175 patients) underwent 250 percutaneous injections of polymethylmethacrylate (PMMA; unilateral, 247 levels; bilateral, 3 levels) into vertebrae under CT and fluoroscopic guidance for 34 months. Patients were prospectively asked to quantify their pain on a visual analog scale (VAS) before and a day after PVP. The interval to mobilization was recorded in those who were immobilized because of pain and/or bed-rest therapy (115 cases). PVP was technically successful in all patients, with three cases of minimal complications. The mean VAS score available for 196 cases was improved from 7.22+/-1.89 (range, 3-10) to 2.07+/-1.19 (range, 0-10) by PVP. Ninety-four of 115 immobilized cases (81.7%) were mobile by 24 h after PVP, and the mean value was 1.9+/-2.8 days. The incidence of recurrent and new fractures was 15.6% in 4-25 months (mean, 15.3 months). PVP is a safe and effective treatment for relieving the pain associated with osteoporotic VCF and strengthening the vertebrae, avoiding refractures. This therapy leads to early mobilization and avoidance of the dangers of conservative therapy of bed-rest. PMID- 15662481 TI - Characterization of hypervariable region in hepatitis C virus envelope protein during acute and chronic infection. AB - Hepatitis C virus (HCV) causes persistent infection in most patients. To clarify the mechanisms underlying establishment of this persistent infection, nucleotide sequences of the E1/E2 region were characterized in 5 patients with acute and chronic HCV infection. We used direct DNA sequencing methods to identify the major sequence of HCV in each patient. Each HCV genome displayed a high frequency of nucleotide sequence variation in the hypervariable region (HVR) of E2. However, patient-specific conserved nucleotide sequences were identified in the E1/E2 region during the course of infection and conserved the higher-order protein structure. In the acute phase HCV infection, amino acid substitution in HVR-1 as the monthly rate of amino acids substitution per site (%) between each point exceeded 10.2%. In the chronic phase HCV infection, a significantly lower rate of amino acid substitution was observed in patients. The host immune responses to HVR-1 of each HCV isolates from all clinical courses were characterized using synthetic peptides and ELISA. One chronic patient serum (genotype 1b) did not react at all to its own HVR-1 peptides, however another patient (genotype 2b) reacted to all clinical course. These results indicated that HVR-1 might not always exhibit neutralizing epitopes of HCV infection. The sequence variation in HVR-1 may instead indicate the existence of various clones in acute phase infection and the adaption of these clones is thought to have caused persistent and chronic infection in each patient. PMID- 15662482 TI - Phylogeny, genome evolution, and antigenic variability among endemic foot-and mouth disease virus type A isolates from India. AB - The capsid-coding (P1) and 3A regions of foot-and-mouth disease virus (FMDV) type A field isolates including two vaccine strains collected during 1977-2000 were analyzed. In the phylogenies, the isolates were distributed into two previously identified genotypes VI and VII, with multiple sub-genotypes that are temporally clustered. Comparison of the antigenic relationships of field isolates with the two vaccine strains (IND 17/77 and IND 490/97) and the reference strains of the genotypes VI (IND 233/99) and VII (IND 40/00) indicated two broad antigenic groups that correlate with the phylogenetic groupings (genotypes VI and VII), and are highly divergent from the vaccine strains. The maximum likelihood method of selection analysis identified a number of amino acid sites within the P1 region to be under weak positive selection. Some of the positively selected sites were mapped at/near the antigenically critical amino acid sites of the P1 region, indicating host immune pressure as one of the important driving force behind the observed genetic and antigenic diversity in FMDV. A small number of selected sites are located in the heparan sulphate-binding pocket of the virus, suggesting a fitness advantage for cell entry of the virus. No positive selection was detected in the 3A dataset. PMID- 15662483 TI - A loop mediated isothermal amplification (LAMP) method for detection of infectious hematopoietic necrosis virus (IHNV) in rainbow trout (Oncorhynchus mykiss). AB - A reverse transcription loop-mediated isothermal amplification (RT-LAMP) protocol was developed for detection of infectious hematopoietic necrosis virus (IHNV) RNA in rainbow trout (Oncorhynchus mykiss). A set of four primers, two outer and two inner primers for the RT-LAMP and the LAMP assay, were designed based on the sequence of G-protein of IHNV. Time and temperature conditions were optimized for 60 min at 63 degrees C for both RT-LAMP and LAMP protocols. The detection limit was found to be similar for both RT-LAMP and LAMP. When the sensitivity of RT LAMP and LAMP were compared with conventional nested PCR, a10-fold higher sensitivity was seen for the LAMP protocols. PMID- 15662484 TI - West Nile virus in the vertebrate world. AB - West Nile virus (WNV), an arthropod-borne virus belonging to the family Flaviviridae, had been recognized in Africa, Asia and the south of Europe for many decades. Only recently, it has been associated with an increasing number of outbreaks of encephalitis in humans and equines as well as an increasing number of infections in vertebrates of a wide variety of species. In this article, the data available on the incidence of WNV in vertebrates are reviewed. Moreover, the role of vertebrates in the transmission of WNV, the control of WNV infections in veterinary medicine as well as future perspectives are discussed. A wide variety of vertebrates, including more than 150 bird species and at least 30 other vertebrate species, are susceptible to WNV infection. The outcome of infection depends on the species, the age of the animal, its immune status and the pathogenicity of the WNV isolate. WNV infection of various birds, especially passeriforms, but also of young chickens and domestic geese, results in high titred viremia that allows arthropod-borne transmission. For other vertebrate species, only lemurs, lake frogs and hamsters develop suitable viremia levels to support arthropod-borne transmission. The role of vertebrates in direct, non arthropod-borne transmission, such as via virus-contaminated organs, tissues or excretions is less well characterized. Even though direct transmission can occur among vertebrates of several species, data are lacking on the exact amounts of infectious virus needed. Finally, the increased importance of WNV infections has led to the development of killed, live-attenuated, DNA-recombinant and chimeric veterinary vaccines. PMID- 15662485 TI - Protective immune response against foot-and-mouth disease virus challenge in guinea pigs vaccinated with recombinant P1 polyprotein expressed in Pichia pastoris. AB - Vaccination of the susceptible livestock with potent, safe and cost effective vaccine is the primary requirement to control foot-and-mouth disease (FMD) in an endemic country. In this study, an alternative approach was used in which structural protein genes of all the four serotypes of FMDV (O, Asia 1, A22 and C) were expressed separately in methylotrophic yeast Pichia pastoris. The recombinant polyproteins (P1) were characterized by SDS-PAGE and in Western Blot analysis. Partially purified protein was used for immunization in guinea pigs with different adjuvant formulations and immune response studied. Ninety micrograms of the recombinant protein per monovalent dose was used for immunization. A single injection of a monovalent or polyvalent vaccine was given to guinea pigs with various adjuvant combinations viz., Monovalent recombinant protein either adjuvanted with Montanide-ISA50V or Indigenous oil, Monovalent recombinant protein mixed with 1/10th dose of inactivated oil-adjuvanted virus vaccine and Polyvalent recombinant protein with Montanide ISA50V. FMDV specific humoral immune response was observed at about 28th day post vaccination. The immune response as assessed by indirect ELISA and Serum neutralization test titres was found to be 320-640 and 16-32, respectively. When challenged with virulent homologous type 'O' virus, the guinea pigs showed protective C index of 2.01,1.81, 2.56 and 2.48, respectively, with above said adjuvant combinations. The study has shown that yeast-expressed FMDV P1 polyprotein in a single dose could elicit a protective immune response in guinea pigs, and this could be a possible future vaccine candidate in homologous host. PMID- 15662486 TI - A monoclonal antibody that recognizes Epstein-Barr virus nuclear antigen 2 (EBNA 2) amino acids 1-58 does not react with EBNA 2 in native form, consistent with the self-association of EBNA 2 through the amino-terminus. AB - We have generated a mouse IgG1 monoclonal antibody (mAb) that recognizes amino acids 1-58 of Epstein-Barr virus (EBV) nuclear antigen 2 (EBNA 2) of type 1 EBV strain B95-8. mAb Y101 also reacted with EBNA 2 of EBV type 2 strains MISP and Jijoye in immunoblots, whereas Jijoye EBNA 2 was not detected by the widely used mAb PE2. mAb Y101, in contrast to mAb PE2, reacted with faster migrated, hypophosphorylated proteins of type 1 EBNA 2 as intensely as slower migrated, hyperphosphorylated ones. mAb Y101 did not react in fixed-cell immunostaining or cell extract immunoprecipitation. The results implicate that the amino-terminal epitope is not exposed in a native form, consistent with the previously reported idea of self-association of EBNA 2 through the amino-terminus. mAb Y101 is the first mAb to the EBNA 2 amino-terminus and will be useful for further analyses of the structure and function of EBNA 2. PMID- 15662487 TI - Use of CD34+ autologous stem cell transplantation in the treatment of children with refractory systemic lupus erythematosus. AB - We report on the unique effects and benefits of autologous stem cell transplantation in childhood systemic lupus erythematosus (SLE) and describe this procedure in two young girls with severe and refractory disease. The patients' stem cells were mobilized with granulocyte colony-stimulating factor (G-CSF) and collected by CS-3000 Blood Cell Separator (Baxter Healthcare, Round Lake, Ill., USA), and the CliniMACS CD34+ cell selection device (Miltenyi Biotech, Bergisch Gladbach, Germany) was used to obtain CD34+ cells. A total of 1.7x10(6) and 1.0x10(6)/kg CD34+ cells were obtained, with 2.0x10(5) and 1.0x10(4)/kg of CD3+ cells remaining, respectively. The conditioning regimen consisted of cyclophosphamide (50 mg/kg per day for 4 days) plus antithymocyte globulin (ATG Fresenius, 5 mg/kg per day for 3 days). Neutrophil counts recovered within 9 days in both cases. Within 15 days, the platelet counts recovered and were sustained over 100x10(9)/l. Cushingoid features disappeared completely 3 months after transplantation because of the removal of corticosteroid medication. One 13-year old child increased her height by 5 cm in 6 months after stopping steroids. She had not increased her height in her previous 7 years of disease. As of the time of this report, the first patient remains in clinical and laboratory remission for nearly 4 years, while the second suffered a relapse of thrombocytopenia 9 months post-transplantation. One residual effect of their treatment is that their CD4+ cell counts remained in the lower range after one year of transplant. The effect of this conditioning regimen plus CD34+ autologous stem cell transplantation on these two children with refractory SLE was beneficial, but long-term follow-up data and additional experience with this procedure are required. Autologous stem cell transplantation may limit the long-term toxicity of therapy in childhood SLE. PMID- 15662488 TI - Treatment of acute neuropsychiatric lupus with intravenous immunoglobulin (IVIG): a case report and review of the literature. AB - Neuropsychiatric lupus can be difficult to diagnose, and little prospective data exists to help direct management. In this case report we describe the acute onset of symptoms of depression, mania, and psychosis and their complete resolution 48 h following a 5-day treatment course of intravenous immunoglobulin (IVIG) in a 20 year-old woman with systemic lupus erythematosus (SLE). We review the literature on IVIG for the management of neuropsychiatric lupus. We propose that when more toxic therapies are refused or symptoms do not remit with other treatments, IVIG should be considered in patients with neuropsychiatric lupus. PMID- 15662489 TI - A new vaginal speculum for pelvic organ prolapse quantification (POPQ). AB - The purposes of this study were to introduce a new vaginal speculum, describe the technique of using the new speculum in identifying and measuring the severity of pelvic organ prolapse (POP), and present results of a pilot study comparing the new speculum to the conventional instruments used in performing POP quantification (POPQ). The new speculum has retractable upper and lower blades marked in centimeters. POPQ was performed with one instrument using the new speculum and multiple instruments performing the conventional technique. Twenty two patients underwent POPQ-11 using the new speculum and 11 using conventional instruments. The duration of the procedure and the level of discomfort were assessed. The POPQ method using the new speculum is described. Preliminary experience with the new speculum showed that the length of examination is significantly shorter (p<0.001) and the comfort level is better than with the conventional technique (p=0.088). A new vaginal speculum with adjustable blades simplifies POPQ. Preliminary testing suggests potential savings in procedure time and reduction in patient discomfort. PMID- 15662490 TI - Impact of duloxetine on quality of life for women with symptoms of urinary incontinence. AB - The objective of this study was to evaluate the effectiveness of duloxetine in improving quality of life among women with stress and mixed urinary incontinence. The study included 451 women with self-reported stress incontinence episodes (>or=1/week) who were randomized to duloxetine (40 mg BID) or placebo in a double blind, usual care design. Patients and physicians were allowed to titrate, augment, and/or discontinue treatment. Concomitant treatments were permitted. The primary outcome was the Incontinence Quality of Life Questionnaire (I-QOL) score, with assessments at 3, 6, and 9 months. Other measures included the Patient Global Impression of Improvement (PGI-I) and adverse events. The adjusted mean change in I-QOL total score was greater in the duloxetine group than in the placebo group and at a level comparable to that found in previous clinical trials, but the difference between placebo and duloxetine was not statistically significant in the intent-to-treat, last observation carried forward (LOCF) analysis. The difference approached statistical significance in favor of duloxetine at 3 months (p=0.07). PGI-I ratings did not demonstrate significant superiority for duloxetine in LOCF analysis; however, study completers taking duloxetine were significantly more likely to rate themselves as "better" (70.2%) than completers taking placebo (50.8%, p<0.05). Women utilized a variety of treatment methods including pelvic floor muscle training, estrogen, anticholinergic medication, weight reduction, and smoking cessation. In this study, while mean I-QOL change scores were numerically higher for the duloxetine group than mean change scores for the placebo group, this difference was not statistically significant. Among women who completed the study on study drug, a significantly greater proportion of duloxetine women versus placebo women rated their condition to be better. PMID- 15662491 TI - CT imaging in acute pulmonary embolism: diagnostic strategies. AB - Computed tomography pulmonary angiography (CTA) has increasingly become accepted as a widely available, safe, cost-effective, and accurate method for a quick and comprehensive diagnosis of acute pulmonary embolism (PE). Pulmonary catheter angiography is still considered the gold standard and final imaging method in many diagnostic algorithms. However, spiral CTA has become established as the first imaging test in clinical routine due to its high negative predictive value for clinically relevant PE. Despite the direct visualization of clot material, depiction of cardiac and pulmonary function in combination with the quantification of pulmonary obstruction helps to grade the severity of PE for further risk stratification and to monitor the effect of thrombolytic therapy. Because PE and deep venous thrombosis are two different aspects of the same disease, additional indirect CT venography may be a valuable addition to the initial diagnostic algorithm-if this was positive for PE-and demonstration of the extent and localization of deep venous thrombosis has an impact on clinical management. Additional and alternate diagnoses add to the usefulness of this method. Using advanced multislice spiral CT technology, some practitioners have advocated CTA as the sole imaging tool for routine clinical assessment in suspected acute PE. This will simplify standards of practice in the near future. PMID- 15662493 TI - Imaging in acute stroke. AB - Stroke is a syndrome characterized by a sudden neurological deficit caused by intracranial hemorrhage or ischemia. Computed tomography (CT) maintains a primary role in the evaluation of patients with acute stroke. The optimal magnetic resonance imaging (MRI) protocol in acute stroke includes diffusion-weighted imaging (DWI) to show acute ischemic lesion and MR perfusion study to estimate brain perfusion. Careful selection of patients for a thrombolytic therapy is crucial to improve safety and efficacy. PMID- 15662492 TI - Radiologic assessment of maxillofacial, mandibular, and skull base trauma. AB - Cranio-maxillofacial injuries affect a significant proportion of trauma patients either in isolation or concurring with other serious injuries. Contrary to maxillofacial injuries that result from a direct impact, central skull base and lateral skull base (petrous bone) fractures usually are caused by a lateral or sagittal directed force to the skull and therefore are indirect fractures. The traditional strong role of conventional images in patients with isolated trauma to the viscerocranium is decreasing. Spiral multislice CT is progressively replacing the panoramic radiograph, Waters view, and axial films for maxillofacial trauma, and is increasingly being performed in addition to conventional films to detail and classify trauma to the mandible as well. Imaging thus contributes to accurately categorizing mandibular fractures based on location, into alveolar, mandibular proper, and condylar fractures-the last are subdivided into intracapsular and extracapsular fractures. In the midface, CT facilitates attribution of trauma to the categories central, lateral, or combined centrolateral fractures. The last frequently encompass orbital trauma as well. CT is the imaging technique of choice to display the multiplicity of fragments, the degree of dislocation and rotation, or skull base involvement. Transsphenoid skull base fractures are classified into transverse and oblique types; lateral base (temporal bone) trauma is subdivided into longitudinal and transverse fractures. Supplementary MR examinations are required when a cranial nerve palsy occurs in order to recognize neural compression. Early and late complications of trauma related to the orbit, anterior cranial fossa, or lateral skull base due to infection, brain concussion, or herniation require CT to visualize the osseous prerequisites of complications, and MR to define the adjacent brain and soft tissue involvement. PMID- 15662494 TI - Percutaneous treatment in patients presenting with malignant cardiac tamponade. AB - The percutaneous treatment of pericardial effusion resulting in cardiac tamponade has undergone an evolution in recent years with the use of less invasive drainage techniques in selected cases. To determine optimal therapy modalities for oncology patients with malignant pericardial tamponade (MPT), the authors review their institutional experience with percutaneous needle puncture routes, means of imaging-guided drainage and percutaneous management of the pericardial fluid effusion (pericardial sclerosis and balloon pericardiotomy). Advantages and limits of the percutaneous techniques will be compared to the surgical treatment. PMID- 15662495 TI - Safety of ultrasound contrast agents. AB - The use of ultrasound contrast agents has increased over recent years. The Contrast Media Safety Committee (CMSC) of the European Society of Urogenital Radiology (ESUR) decided to review the safety of ultrasound contrast agents in humans and to draw up guidelines. A comprehensive literature search and review was carried out. The resulting report was discussed by the CMSC of ESUR and at the 11th European Symposium on Urogenital Radiology in Santiago de Compostela, Spain, in 2004. Ultrasound contrast agents approved for clinical use are well tolerated, and serious adverse reactions are rarely observed. Adverse events are usually minor (e.g. headache, nausea, altered taste, sensation of heat) and self resolving. These symptoms may not be related to the ultrasound contrast materials as they have also been observed in placebo-control groups. Intolerance to some components may occur. Generalized allergy-like reactions occur rarely. Ultrasound contrast agents are generally safe. The ultrasound scanning time and the acoustic output should be kept to the lowest level consistent with obtaining diagnostic information. Adverse reactions should be treated symptomatically. PMID- 15662496 TI - Evaluation of spatial and temporal resolution for ECG-gated 16-row multidetector CT using a dynamic cardiac phantom. AB - Measurements of spatial and temporal resolution for ECG-gated scanning of a stationary and moving heart phantom with a 16-row MDCT were performed. A resolution phantom with cylindrical holes from 0.4 to 3.0 mm diameter was mounted to a cardiac phantom, which simulates the motion of a beating heart. Data acquisition was performed with 16x0.75 mm at various heart rates (HR, 60-120 bpm), pitches (0.15-0.30) and scanner rotation times (RT, 0.42 and 0.50 s). Raw data were reconstructed using a multi-cycle real cone-beam reconstruction algorithm at multiple phases of the RR interval. Multi-planar reformations (MPR) were generated and analyzed. Temporal resolution and cardiac cycles used for image reconstruction were calculated. In 97.2% (243/250) of data obtained with the stationary phantom, the complete row of holes with 0.6 mm was visible. These results were independent of heart rate, pitch, scanner rotation time and phase point of reconstruction. For the dynamic phantom, spatial resolution was determined during phases of minimal motion (116/250). In 40.5% (47/116), the resolution was 0.6 mm and in 37.1% (43/116) 0.7 mm. Temporal resolution varied between 63 and 205 ms, using 1.5-4.37 cardiac cycles for image reconstruction. PMID- 15662497 TI - The establishment of endovascular aneurysm coiling at a neurovascular unit: report of experience during early years. AB - The treatment of cerebral aneurysms is changing from surgical clipping to endovascular coiling (EVC) in many neurovascular centres. The aim of this study was to evaluate the technical results and clinical outcome at 6 months in a consecutive series of subarachnoid hemorrhage (SAH) patients treated with EVC, in a situation when the EVC had been established very rapidly as the first line of treatment at a neurovascular centre. The patient material comprised 239 SAH patients (155 women and 84 men, mean age 55 years, age range 16-81) allocated to EVC as the first line of treatment in the acute stage (within 3 weeks of rupture) between September 1996 and December 2000. Clinical grade on admission was Hunt & Hess (H&H) I and II in 42%, H&H III in 25% and H&H grade IV and V in 33% of the patients. The aneurysm was located in the anterior circulation in 82% of the cases. EVC was performed on days 0-3 in 77% of the cases. EVC of the target aneurysm was able to be completed in 222 patients (93%). Complete occlusion was achieved in 126 patients (53%). Procedural complications occurred in 39 patients (16%). Favourable clinical outcome was observed in 57%, severe disability in 28% and poor outcome in 14% of the patients. Favourable outcome was achieved in 77% of H&H I and II patients and in 43% of H&H III-V patients. The multivariate logistic regression analysis revealed that younger age, good neurological grade on admission, absence of intracerebral hematoma and intraventricular hematoma respectively, ICA-PcomA aneurysm location, later treatment and absence of complications were significant predictors of favourable outcome. After interventional training and installation of the X-ray system, the introduction and establishment of EVC at a neurovascular unit can be done in a short period of time and with favourable results. Future studies must concentrate on identifying factors of importance for the choice of interventional or surgical therapy. The results of this study indicate that endovascular therapy may be particularly beneficial in poor-grade patients and in patients with aneurysms in the ICA-PcomA territory. PMID- 15662498 TI - Intracranial angiolipoma as cause of subarachnoid haemorrhage. Case report and review of the literature. AB - A 33-year-old female with a longstanding history of seizures was admitted to our hospital with subarachnoid haemorrhage (SAH). Computed tomography (CT), magnetic resonance imaging (MRI) and digital subtraction angiography (DSA) depicted a vascular fat-containing lesion overlying a right frontal cortical polymicrogyria. The diagnosis of angiolipoma was established. Conservatory management was undertaken with full recovery. She has been followed for 5 years since, with neither re-bleeding nor morphologic change of the lesion. This is a rare intracranial lesion, with only 11 intracranial angiolipomas published in the literature, and is the first case reported which is associated with SAH caused by this lesion. PMID- 15662499 TI - [Simultaneous analysis of t(X;18) by FISH- und SYT/SSX-RT-PCR in synovial sarcoma]. AB - Synovial sarcoma diagnosis and differential diagnostic distinction from other spindle cell sarcomas may be difficult. In these cases the detection of the t(X;18) translocation by FISH and RT-PCR is diagnostically extremely helpful. This study was aimed at the question whether or not simultaneous use of both methods is required for evidence of t(X;18) translocation.Paraffin-embedded tumour specimens from 53 patients were included in the study which were considered to be possible synovial sarcomas on the basis of histological aspect and immunohistochemical profile. Detection of t(X;18) was performed using FISH and RT-PCR simultaneously. Nuclei and amplifiable RNA could be isolated from 39 of the 53 included cases (75%). In 72% of these 39 cases FISH and RT-PCR showed identical negative or positive results. The remainder of the cases (28%) showed either a typical PCR product or a positive FISH signal.In conclusions FISH could be confirmed by typical PCR products and is therefore qualified as an internal quality control. Nevertheless tumour biological and methodical reasons have an important influence on both methods. Consequently in difficult cases simultaneous FISH and RT-PCR analysis is necessary for a clear evidence of t(X;18) translocation. PMID- 15662500 TI - Gene silencing studies in the gymnosperm species Pinus radiata. AB - A biolistic transformation procedure was used to transform embryogenic Pinus radiata tissue with constructs containing the Zea mays UBI1 (ubiquitin)-promoter followed by the P. radiata CAD (cinnamyl alcohol dehydrogenase) cDNA in sense or anti-sense orientation or in the form of an inverted-repeat. The effect of the different constructs on silencing the endogenous CAD gene was monitored in embryogenic tissue and somatic seedlings of 28 P. radiata transclones. Quantitative CAD measurements demonstrated that the construct containing an inverted-repeat of the CAD cDNA was most efficient in triggering gene silencing in P. radiata. Northern hybridization experiments with silenced transclones revealed that reduced CAD activities were the result of reduced steady state levels of the targeted CAD mRNA. Monitoring of the activity of the UBI1-promoter in the P. radiata transclones and heat-shock experiments with transgenic somatic P. radiata seedlings indicated that gene silencing is positively correlated with the expression level of the transgene. The obtained data are also consistent with a role for the expression level of the endogenous CAD gene in gene silencing. PMID- 15662501 TI - Excision of a selectable marker in transgenic rice (Oryza sativa L.) using a chemically regulated Cre/loxP system. AB - Removal of a selectable marker gene from genetically modified (GM) crops alleviates the risk of its release into the environment and hastens the public acceptance of GM crops. Here we report the production of marker-free transgenic rice by using a chemically regulated, Cre/loxP-mediated site-specific DNA recombination in a single transformation. Among 86 independent transgenic lines, ten were found to be marker-free in the T0 generation and an additional 17 lines segregated marker-free transgenic plants in the T1 generation. Molecular and genetic analyses indicated that the DNA recombination and excision in transgenic rice were precise and the marker-free recombinant T-DNA was stable and heritable. PMID- 15662502 TI - [Influence of cutoff filters on reading behavior in age-related macular degeneration]. AB - BACKGROUND: Cutoff filters may improve contrast sensitivity in different retinal dystrophies and therefore lead to improved visual function. We investigated whether cutoff filters result in improved reading ability in age-related macular degeneration (AMD). METHODS: Reading performance was examined in 22 patients with AMD aged 63-91 years (visual acuity 0.1-0.5) using the Radner reading charts. These charts were presented in random order either with or without cutoff filters (Zeiss CF 540 and CF 580) monocularly to the better eye using a defined luminance of the test charts of 105 cd/m(2). RESULTS: Using the CF 540 cutoff filter, there was no significant change in visual acuity while there was a tendency to decrease. Calculating the LogRAD score, which takes reading errors into account, three patients improved by one line, while nine deteriorated, six of them by one line or more. There was no significant improvement using the CF 580 compared to either the CF 540 or to normal reading glasses. CONCLUSIONS: Overall, we found no measurable improvement using the CF 540 cutoff filter. The cutoff filter CF 580, tested for additional comparison, did not show any advantage. PMID- 15662503 TI - [Photodynamic therapy with verteporfin for recurrent choroidal neovascularisation (CNV) following prior argon laser coagulation]. AB - BACKGROUND: Photodynamic therapy (PDT) with verteporfin has been successfully introduced to treat choroidal neovascularisations (CNV) that have more than 50% classic CNV components due to exudative age-dependent macular degeneration (AMD) and other diagnoses. However, what is still unclear is if patients with recurrent subfoveal CNV who have had prior laser coagulation can also profit from PDT for CNV. METHODS: The course of visual acuity (ETDRS) and the fluorescein angiographic findings were retrospectively analysed after standard PDT of recurrent subfoveal CNV after prior argon laser coagulation of an extrafoveal or juxtafoveal CNV in AMD or other diagnoses. RESULTS: A total of 20 consecutive eyes from 19 AMD patients were evaluated. After 12 months in 14 of the 20 eyes (70%) deterioration in visual acuity of 3 or more ETDRS lines could be avoided and likewise in 7 of 16 eyes (44%) after 24 months. After the 1st year, 5 of the 7 eyes (71%) with recurrent subfoveal CNV after laser coagulation of a myopic, postinflammatory or idiopathic CNV had deterioration of visual acuity of less than three lines or an improvement in visual acuity. CONCLUSIONS: Despite the low number of patients examined, we could see an indication for PDT in recurrent subfoveal CNV after prior argon laser coagulation of extrafoveal or juxtafoveal CNV. When the patient presented with a CNV cause other than AMD, a better prognosis for visual acuity was achieved. PMID- 15662504 TI - Are age references for waist circumference, hip circumference and waist-hip ratio in Dutch children useful in clinical practice? AB - The aim of this study was to present age references for waist circumference (WC), hip circumference (HC), and waist/hip ratio (WHR) in Dutch children. Cross sectional data were obtained from 14,500 children of Dutch origin in the age range 0-21 years. National references were constructed with the LMS method. This method summarises the distribution by three smooth curves representing skewness (L curve), the median (M curve), and coefficient of variation (S curve). The correlations between body mass index-standard deviation score (BMI-SDS), the circumferences and their ratio, and demographic variables were assessed by (multiple) regression analysis for three age groups: 0-<5 years (1), 5-<12.5 years (2), and 12.5-<21 years (3). A cut-off for clinical use was suggested based on the International Obesity Task Force criteria for BMI. Mean WC and HC values increased with age. Mean WC was slightly higher in boys than in girls, and this difference was statistically significant from 11 years of age onwards. In contrast, HC was significantly higher in girls than in boys from 9 years onwards. The correlation between WC-SDS and BMI-SDS ( r =0.73, P <0.01) and between HC and BMI-SDS ( r =0.67, P <0.01) increased with age. With regard to WHR-SDS, a low correlation was found for 12.5-20 years of age ( r =0.2, P <0.01). WC-SDS correlated positively with height SDS ( r =0.35, P <0.01). CONCLUSION: Waist circumferences can be used to screen for increased abdominal fat mass in children, whereby a cut-off point of 1.3 standard deviation score seems most suitable. PMID- 15662505 TI - Procalcitonin in hypoxic brain damage. PMID- 15662506 TI - Pharmacokinetics of intravenous paracetamol in children and adolescents under major surgery. AB - BACKGROUND: The aim of this study was to describe the pharmacokinetics of paracetamol (acetaminophen) and to get primary information on its metabolism after first indicated intravenous administration of paracetamol in children and adolescents undergoing major surgery. METHODS: About 4 weeks after the last chemotherapy, seven children and adolescents (five osteosarcoma, two Ewing tumors) received paracetamol infusion (median: 15.0 mg/kg) for analgesia. Sparse serum (37 samples; 4-7 per patient) and urine samples (27 samples; 0-15 per patient) were analyzed for paracetamol, paracetamol-glucuronide, paracetamol sulfate, paracetamol-mercapturate and paracetamol-cysteine using capillary electrophoresis. Nonlinear mixed-effect models were used to describe the pharmacokinetics of paracetamol in plasma. RESULTS: Pharmacokinetics of paracetamol after intravenous administration was best described by a two compartment model with clearance of 13.2 l/h per 70 kg (between-subject variability: 30%), intercompartmental clearance of 45.7 l/h per 70 kg (both parameters standardized to a 70-kg person using allometric "1/4 power models"), central volume of distribution of 13.2 l per 70 kg (between-subject variability: 71%) and peripheral volume of distribution of 33.0 l per 70 kg. Paracetamol, the glucuronide- and sulfate conjugates as well as cysteine and mercapturic acid conjugates, both products of oxidative pathways of paracetamol, were excreted in urine. CONCLUSIONS: Surgery, with all its potential influencing factors, together with chemotherapy given about 4 weeks previously do not seem to have a major impact on the pharmacokinetic behavior and the between-subject variability of paracetamol after intravenous administration. PMID- 15662507 TI - Tramadol concentrations in blood and in cerebrospinal fluid in a neonate. AB - Based on blood and cerebrospinal fluid samples collected in a full-term neonate, the penetration of tramadol in the central nervous system is described. Following intravenous administration of tramadol, a lag time of about 4 h was observed until full blood-brain equilibration was achieved. This pharmacokinetic observation is in line with a recent pharmacodynamic evaluation of the central opioid effects of tramadol in adults. PMID- 15662509 TI - Measuring the solubilities of ionic liquids in water using ion-selective electrodes. AB - Polyvinyl chloride-plasticized membrane ion-selective electrodes (ISE) based on conventional ion-exchangers have been proposed as a cheap universal tool to measure the solubilities of ionic liquids (ILs) in water. They are applicable for ILs with a wide range of solubilities in water, since the linear range of a potentiometric response spans several orders of magnitude. As an example, we have fabricated and tested ISEs for widely used alkylimidazolium ionic liquids. The aqueous solubilities of four typical ILs have been determined at 21 degrees C: 0.075 +/- 0.001 mol l(-1) (1-butyl-3-methylimidazolium, BMIm, hexafluorophosphate); 0.018 +/- 0.001 mol l(-1) (BMIm bis(triflylimide)); 0.054 +/- 0.007 mol l(-1) (1-butyl-2,3-dimethylimidazolium, BDMIm, hexafluorophosphate); 0.014 +/- 0.001 mol l(-1) (BDMIm bis(triflylimide)). PMID- 15662508 TI - The effect of the CYP2C19 genotype on the hydroxylation index of omeprazole in South Indians. AB - To investigate the relationship between CYP2C19 genotypes and the hydroxylation index (HI) of omeprazole in the South Indian population. Healthy unrelated South Indian subjects (n=300) were separated into three groups based on their CYP2C19 genotypes. They were administered a single oral dose of 20 mg omeprazole, and venous blood was collected 3 h later. Plasma was assayed using reversed-phase high-performance liquid chromatography, and the omeprazole HI was calculated. The means of HIs in individuals with CYP2C19*1/*1 (n=124), *1/*2 (n=129) and *2/*2,*2/*3 (n=47) were 2.4, 5.3 and 22.5, respectively, and were found to be significantly different between any two groups (P<0.0001). A good correlation was established between CYP2C19 genotype and omeprazole HI (r=0.54, 95% CI 0.45-0.62; P<0.0001). Of the 300 subjects, 42 (14.0%; 95% CI 10.1-17.9) were phenotypic poor metabolizers (PMs), but only 33 of them had two mutant alleles and the remaining 9 PMs had at least one wild-type allele. Among the 258 extensive metabolizers, 14 had two mutant alleles. The prevalence of PMs in the South Indian population was 14.0%, which is similar to that in North Indians and Orientals but significantly higher than in Caucasians and Africans. A genotype-phenotype relationship was established between the CYP2C19 genotype and HI of omeprazole, but 7.7% of subjects deviated from expected genotype-phenotype associations. This could be due to an additional mutation, either in the exons/introns or in the 5' regulatory region of the CYP2C19 gene. PMID- 15662510 TI - Investigation of the recovery of selenomethionine from selenized yeast by two dimensional LC-ICP MS. AB - Determination of selenomethionine in selenized yeast by HPLC-ICP MS has been revisited with the focus on recovery of this amino acid during the proteolytic digestion and chromatography steps. Recovery of the extracted selenium from an anion-exchange column was 100% but selenomethionine quantified by the method of standard additions accounted only for 67% of the selenium injected. Analysis (by size-exclusion LC-ICP MS) of the eluate collected before and after the selenomethionine peak showed the presence of oxidized selenomethionine (ca. 3%) and selenomethionine likely to be unspecifically associated with the biological matrix continuum (ca. 11%). This finding was validated by two-dimensional LC-ICP MS using a different elution order, i.e. size-exclusion anion-exchange. The approach developed enabled demonstration that more than 80% of selenium in the selenized yeast is actually present in the form of selenomethionine and suggests that many results reported elsewhere for the concentration of this vital amino acid in selenized yeast may be negatively biased. The research also provided insight into speciation of selenium in the solid residue after proteolytic extraction but the additional amount of selenomethionine recovered was negligible (<1.5%). PMID- 15662512 TI - Pretreatment procedures for characterization of arsenic and selenium species in complex samples utilizing coupled techniques with mass spectrometric detection. AB - Research interest in analyzing arsenic and selenium is dictated by their species dependent behavior in the environment and in living organisms. Different analytical methodologies for known species in relatively simple chemical systems are well established, yet the analysis of complex samples is still a challenge. Owing to the complex matrix and low concentrations of target species that may be chemically labile, suitable pretreatment of the sample becomes a critical step in any speciation procedure. In this paper, the pretreatment procedures used for arsenic and selenium speciation are reviewed with the emphasis on the link between the analytical protocol applied and the biologically-significant information provided by the results obtained. In the first approach, the aim of pretreatment is to convert the original sample into a form that can be analyzed by a coupled (hyphenated) technique, preventing possible losses and/or species interconversion. Common techniques include different leaching and extraction modes, enzymatic hydrolysis, species volatilization, and so on, with or without species preconcentration. On the other hand, if the speciation analysis is performed for elucidation of elemental pathways and specific functions in a living system, more conscious pretreatment and/or fractionation is needed. The macroscopic separation of organs and tissues, isolation of certain types of cells, cell disruption and separation of sub-cellular fractions, as well as isolation of a specific biomolecules become important. Furthermore, to understand molecular mechanisms, the identification of intermediate-often highly instable- metabolites is necessary. Real life applications are reviewed in this work for aquatic samples, soils and sediments, plants, yeast, and urine. PMID- 15662513 TI - Surface plasmon resonance-based immunoassay for 17beta-estradiol and its application to the measurement of estrogen receptor-binding activity. AB - A rapid and simple surface plasmon resonance (SPR)-based immunoassay for detection of 17beta-estradiol was developed. The assay was designed as an inhibitive format, in which 17beta-estradiol-BSA conjugates are immobilized on an SPR sensor chip and the binding of antibody to the chip is measured. The binding was inhibited by 17beta-estradiol in the concentration range 0.468 to 21.4 nmol L 1 with an IC50 value of 2.29+/-0.10 nmol L-1. Although not as sensitive as traditional radioimmunoassay (RIA) and enzyme-linked immunoassay (ELISA), this method requires no separation and washing after addition of the antibody, steps which are relatively time-consuming. Estrogen receptor (ER)-binding was then investigated using this SPR immunoassay for the determination of the amount of unbound 17beta-estradiol after competition with test compounds for the ER binding. Inhibition of the binding of 17beta-estradiol to ER by diethylstilbestrol (DES) was successfully measured by injecting the reaction mixture into the SPR sensor after addition of the antibody. This binding assay requires no separation of unbound 17beta-estradiol from the mixture and no radioisotope- or fluorescence-labeling of 17beta-estradiol. These results show the potential usefulness of the SPR sensor both detecting 17beta-estradiol and evaluating the ER-binding activity of xenoestrogens such as DES in a single assay system. PMID- 15662514 TI - Extraction of methylmercury from tissue and plant samples by acid leaching. AB - A simple and efficient extraction method based on acidic leaching has been developed for measurement of methylmercury (MeHg) in benthic organisms and plant material. Methylmercury was measured by speciated isotope-dilution mass spectrometry (SIDMS), using gas chromatography interfaced with inductively coupled plasma mass spectrometry (GC-ICP-MS). Reagent concentration and digestion temperature were optimized for several alkaline and acidic extractants. Recovery was evaluated by addition of MeHg enriched with CH3 201Hg+. Certified reference materials (CRM) were used to evaluate the efficiency of the procedure. The final digestion method used 5 mL of 4 mol L(-1) HNO3 at 55 degrees C to leach MeHg from tissue and plant material. The digest was further processed by aqueous phase ethylation, without interference with the ethylation step, resulting in 96 +/- 7% recovery of CH3 201Hg+ from oyster tissue and 93+/-7% from pine needles. Methylmercury was stable in this solution for at least 1 week and measured concentrations of MeHg in CRM were statistically not different from certified values. The method was applied to real samples of benthic invertebrates and inter laboratory comparisons were conducted using lyophilized zooplankton, chironomidae, and notonectidae samples. PMID- 15662515 TI - [Treatment of vitally endangered anorexia nervosa patients based on guardianship laws]. AB - Anorexia nervosa is a psychiatric disorder occurring primarily in young women. Especially when chronic or the body mass index is less than 12 kg/m(2), it has a mortality of up to 20%. Often these patients are admitted to emergency units and treated internistically. If they accept psychiatric treatment, they are normally transferred to specialized psychosomatic units for further therapy. If patients are not able to accept necessary further therapy to overcome the danger to health and life, the question arises in terms of guardianship law of whether they are able to handle their personal concerns alone. After several vitally endangered anorexia nervosa patients had been admitted to our closed psychiatric ward, we developed a therapeutic concept for this subgroup of patients, taking the possibilities of guardianship law into account. This concept aims at restoring the body weight continuously and finally enable the patients to be transferred to less restrictive psychosomatic units. The chosen treatment is shown with the therapy courses of 25 patients treated according to this concept. PMID- 15662516 TI - Role of anal dilatation in treatment of idiopathic constipation in children: long term follow-up of a double-blind randomized controlled study. AB - Constipation in childhood is a common symptom, with an estimated incidence between 0.3% and 8%. Most of the evidence for the current management of constipation and fecal soiling in children is based on reports of nonrandomized retrospective trials. Anal dilatation has had an established role in the management of idiopathic constipation but has never been evaluated by a randomized study. A double-blind randomized controlled trial was done of children who failed to respond to medical treatment and were admitted for investigation and treatment of idiopathic constipation to Guy's Hospital, London, between April 2001 and April 2003. All children had intestinal transit study on admission. They were randomized, using a computer-generated allocation in sealed envelopes, to receive no anal dilatation (control group) or anal dilatation (anal dilatation group). Anorectal manometry and endosonography were done under ketamine anesthesia followed by anal dilatation if necessary under the same anesthesia. Disimpaction of feces from the rectum was done at the end of the procedure under general anesthesia using propofol muscle relaxant to minimize stretching of anal sphincter muscles in the control group. All children had intensification of medical treatment, toilet training, and monitoring of their response to treatment during their hospital stay, which ranged from 3 to 5 days. Outcome was measured using a parent's questionnaire of symptom severity at 3 and 12 months of follow up by one of the authors, who was blinded to randomization. The symptom severity score ranged between 0 and 65 and consisted of scores for the following: delay in defecation (score range 0-10), difficulty and pain with passing stool (0-5), soiling problem (0-10), intensity of laxative treatment (0-10), child's general health (0-5), behavior related to the bowel problem (0-5), overall improvement of symptoms (0-12,) and assessment of megarectum on abdominal examination (0-8). Of 60 neurologically normal children, 31 (19 males) were randomized in the control group and 29 (18 males) in the anal dilatation group. All children had findings consistent with idiopathic constipation and positive anorectal reflex on manometry, no anal sphincter damage on endosonography, and no anal fissure on examination under anesthesia. The median age for control and anal dilatation groups was 7.97 (range 4.1-14.25) years and 7.78 (4-13.25) years, respectively. Both groups were also comparable with regard to median of duration of laxative treatment (32 months vs. 31.5 months), internal anal sphincter thickness on endosonography (0.90 mm vs. 0.80 mm), resting anal sphincter pressure on manometry (51 mmHg vs. 51 mmHg), total rectal capacity on manometry (260 mmHg vs. 260 mmHg), and total symptom severity score before admission (33 vs. 29), respectively. At 12-month follow-up, the median pre-admission symptom severity score had improved significantly, from 33 (range 12-49) in the control group and 29 (16-51) in the dilatation group to 15 (0-51, p < 0.0001) and 19 (1-46, p < 0.0001), respectively. There was no significant difference between the two groups with regard to symptom severity score improvement at 12-month follow-up (p < 0.92). We found a significant correlation between total rectal capacity measured on manometry and symptom severity score before admission and at 12-month follow up (r = 0.30, p < 0.01 and r = 0.25, p < 0.05, respectively). Our results indicate that anal dilatation does not contribute to the management of school aged children with idiopathic constipation. Admission to hospital for clarification of diagnosis and intensification of medical treatment with disimpaction of stool from the rectum is beneficial. PMID- 15662517 TI - [Working memory in healthy subjects and schizophrenics: studies using BOLD fMRT]. AB - Functional magnetic resonance imaging uses the blood oxygen level-dependent effect (BOLD MRI) for noninvasive display of cerebral correlatives of cognitive function. The importance for the understanding of physiological and pathological processes is demonstrated by investigations of working memory in schizophrenics and healthy controls. Working memory is involved in processing rather than storage of information and therefore is linked to complex processes such as learning and problem solving. In schizophrenic psychosis, these functions are clearly restricted. Training effects in the working memory task follow an inverse U-shape function, suggesting that cerebral activation reaches a peak before economics of the brain find a more efficient method and activation decreases. PMID- 15662518 TI - [Schizophrenia as a disconnection syndrome. Studies with functional magnetic resonance imaging and structural equation modeling]. AB - Aside from characteristic psychopathological symptoms, cognitive deficits are a core feature of schizophrenia. These deficits can only be addressed within the context of widespread functional interactions among different brain areas. To examine these interactions, structural equation modeling (SEM) was used for the analysis of fMRI datasets. In a series of studies, both in antipsychotic-treated and drug-free schizophrenic patients, a pattern of enhanced thalamocortical functional connectivity could be observed as an indicator for possible disruptions of frontostriatal thalamocortical circuitry. Moreover, drug-free patients and those receiving typical antipsychotic drugs were characterized by reduced interhemispheric corticocortical connectivity. This difference relative to normal controls was less in patients under atypical antipsychotic drugs. The results could be interpreted as a beneficial effect of atypical antipsychotic drugs on information processing in schizophrenic patients. The present findings are consistent with the model of schizophrenia as a disconnection syndrome and earlier concepts of "cognitive dysmetria" in schizophrenia. PMID- 15662519 TI - [Positron emission tomography in neuroscience. An integrative part of clinical diagnostic methods and experimental research]. AB - The role of molecular neuroimaging techniques is increasing in the understanding of pathophysiological mechanism of diseases. To date, positron emission tomography is the most powerful tool for the non-invasive study of biochemical and molecular processes in humans and animals in vivo. With the development in radiochemistry and tracer technology, a variety of endogenously expressed and exogenously introduced genes can be analyzed by PET. This opens up the exciting and rapidly field of molecular imaging, aiming at the non-invasive localisation of a biological process of interest in normal and diseased cells in animal models and humans in vivo. Besides its usefulness for basic research positron emission tomography has been proven to be superior to conventional diagnostic methods in several clinical indications. This is illustrated by detection of biological or anatomic changes that cannot be demonstrated by computed tomography or magnetic resonance imaging, as well as even before symptoms are expressed. The present review summarizes the clinical use of positron emission tomography in neuroscience that has helped elucidate the pathophysiology of a number of diseases and has suggested strategies in the treatment of these patients. Special reference is given to the neurovascular, neurodegenerative and neurooncological disease. PMID- 15662521 TI - [Imaging of sacral chordoma: comparison between MRI and CT]. AB - AIM: Sacral chordoma is a primarily benign tumor with a high tendency to recur. A correct definition of tumor volume is a significant prognostic factor. We compared the value of CT and MRI in diagnosing a sacral chordoma. METHODS: 31 patients with a histologically proven sacral chordoma were included in this study. Following parameters were analyzed by two independent radiologists: septation, signs of blood and ossification in the tumor tissue, contrast enhancement, maximal tumor diameter, infiltration of the soft tissue, the dural salc and the cauda equina and multifocality. RESULTS: In CT all chordomas showed a hypodensity to the normal tissue and in MRI a hyperintensity on T2w images with a low level of contrast enhancement. On the basis of the more precise soft tissue contrast of MRI compared with CT, MRI was significantly more accurate in all tested parameters (p <0.05) besides in detecting tumor ossification. In CT tumor volume was frequently underestimated. CONCLUSION: MRI in sacral chordoma is an essential tool in the pretherapeutic diagnostic regimen and in a state of relapse. PMID- 15662523 TI - Cavernous malformations after cerebral irradiation during childhood: report of nine cases. AB - INTRODUCTION: Cavernous hemangioma is increasingly recognized as a late complication of cerebral irradiation (CI); however, the significance of the problem, especially the risk of hemorrhage, is not documented in the literature. In order to discover this, we reviewed our experience of radiation-induced cavernous hemangiomas (RICH) in patients who had received CI during childhood. METHODS: We reviewed retrospectively our pediatric database of 965 brain tumors, 419 of which were irradiated between 1964 and 2003. We collected nine cases of RICH diagnosed in patients who had received CI during childhood. RESULTS: The CI dosage ranged between 25 and 55 Gy. The interval between CI and the diagnosis of RICH ranged from 4 to 22 years. The RICH was the cause of brain hemorrhage in five cases, three of which required surgical evacuation. DISCUSSION: Radiation induced cavernous hemangiomas are an underestimated problem, and systematic screening of irradiated patients with gradient-echo MRI will probably yield more asymptomatic cases. We think that not all RICH require surgery, but only those responsible for intracerebral hemorrhage or that show radiological progression. CONCLUSION: With respect for the long interval between CI and diagnosis of RICH, we advise control with MRI, including gradient-echo sequence, as late as 15 years after CI, and closer monitoring of asymptomatic RICH. PMID- 15662524 TI - The prognostic factor for outcome following second resection for intrahepatic recurrence of hepatocellular carcinoma with a hepatitis B virus infection background. AB - PURPOSE: Second resection has been proved to be a safe and effective treatment for patients with intrahepatic recurrent HCC after primary resection; however, preoperative prognostic factors for outcome following second resection in patients with a hepatitis B virus (HBV) infection background remains to be clarified. METHODS: Fifty-seven patients with intrahepatic recurrent an HCC and HBV infection background received second resection from 1997 to 2003 in our institute. All of them were negative for anti-hepatitis C virus (HCV) and positive regarding HBV profile. Patient and tumor factors were analyzed. RESULTS: At the time of preparing this paper, 31 had re-recurrence and 21 patients had died. No postoperative mortality was noted. The 1-, 3-, and 5-year overall survival after second resection were 69.9%, 61.2%, and 30.6%, respectively. Univariate and multivariate analysis showed that vascular invasion and time to recurrence were the independent prognostic factors for overall survival following second resection. The 3- and 4-year overall survival after second resection were 57.7% and 46.6% in patients with the presence of any of two risk factors (n = 46), and 100% and 100% in those with absence of both risk factors (n = 11, P = 0.008). CONCLUSIONS: Vascular invasion and time to recurrence were the prognostic factors for overall survival following second resection of intrahepatic recurrent HCC. PMID- 15662522 TI - Immunoreactivity for calcium-binding proteins defines subregions of the vestibular nuclear complex of the cat. AB - The vestibular nuclear complex (VNC) is classically divided into four nuclei on the basis of cytoarchitectonics. However, anatomical data on the distribution of afferents to the VNC and the distribution of cells of origin of different efferent pathways suggest a more complex internal organization. Immunoreactivity for calcium-binding proteins has proven useful in many areas of the brain for revealing structure not visible with cell, fiber or Golgi stains. We have looked at the VNC of the cat using immunoreactivity for the calcium-binding proteins calbindin, calretinin and parvalbumin. Immunoreactivity for calretinin revealed a small, intensely stained region of cell bodies and processes just beneath the fourth ventricle in the medial vestibular nucleus. A presumably homologous region has been described in rodents. The calretinin-immunoreactive cells in this region were also immunoreactive for choline acetyltransferase. Evidence from other studies suggests that the calretinin region contributes to pathways involved in eye movement modulation but not generation. There were focal dense regions of fibers immunoreactive to calbindin in the medial and inferior nuclei, with an especially dense region of label at the border of the medial nucleus and the nucleus prepositus hypoglossi. There is anatomical evidence that suggests that the likely source of these calbindin-immunoreactive fibers is the flocculus of the cerebellum. The distribution of calbindin-immunoreactive fibers in the lateral and superior nuclei was much more uniform. Immunoreactivity to parvalbumin was widespread in fibers distributed throughout the VNC. The results suggest that neurochemical techniques may help to reveal the internal complexity in VNC organization. PMID- 15662527 TI - Lysophosphatidic acid stimulates fas ligand microvesicle release from ovarian cancer cells. AB - Previous reports support that lysophosphatidic acid (LPA) upregulates Fas ligand (FasL) cell surface presentation on the ovarian cancer cells. In this study, we aim to investigate soluble FasL (sFasL) secretion associated with the small membrane microvesicles upon LPA stimulation, and to analyze the roles of cytoskeletal reorganization in FasL transport induced by LPA. Ovarian cancer cells were stimulated with LPA and spent media were harvested, concentrated, and ultracentrifugated to collect the supernatant and pellet. Western blot suggested that sFasL released from ovarian cancer cells were the mature form, and these sFasL are released with the small membrane microvesicles. Flow cytometry showed that the majority of microvesicles secreted contained FasL on their membrane, and these small membrane microvesicles are bioactive against activated human T lymphocytes. The microtubule-disrupting reagent nocodazole, not the actin filament-disrupting reagent cytochalasin D pretreatment blocked FasL-expressing small membrane microvesicle release stimulated by LPA, suggesting that microtubules play an essential role in FasL microvesicle transport and exocytosis. LPA may promote ovarian cancer metastasis by counterattacking peritoneal cavity anti-tumor immunity. PMID- 15662525 TI - Interferon alpha 2a down-regulates VEGF expression through PI3 kinase and MAP kinase signaling pathways. AB - An earlier report demonstrated that interferon alpha (IFN-alpha) inhibited tumor growth and recurrence in an MHCC97 xenograft model in nude mice by suppressing tumor angiogenesis rather than by inhibiting tumor cell proliferation. However, the underlying molecular mechanism was not fully elucidated. In this study, we demonstrated that IFN-alpha 2a could down-regulate VEGF expression both in mRNA and in protein levels, as well as down-regulating HIF-1 alpha mRNA expression in MHCC97 cells in vitro. A cDNA micro array analysis followed by Northern and Western blot analysis revealed that PI3 kinase and MAP kinase signaling pathways might be inhibited by IFN-alpha 2a. Blocking the function of IFN-alpha receptor with a specific peptide could eliminate the inhibitory effects of IFN-alpha 2a on VEGF expression. In addition, wortmannin and PD098059, respective inhibitors of the PI3 kinase and the MAP kinase signaling pathways, when used independently or in combination, could also down-regulate the VEGF synthesis and secretion in a similar pattern of IFN-alpha 2a. These observations may lead to the conclusion that IFN-alpha 2a could suppress VEGF synthesis and secretion by down-regulating HIF-1 alpha expression, via inhibition of the PI3 kinase and/or the MAP kinase signaling pathways. PMID- 15662526 TI - Relationship between intracellular localization of p34cdc2 protein and differentiation of esophageal squamous cell carcinoma. AB - PURPOSE: G2/M cyclins including cyclins A and B can exert their biologic functions of mitosis and proliferation of the tumor cells by being combined by protein kinase p34cdc2. The aim of the current study was to elucidate the clinicopathologic significance of immunohistochemical expression of p34(cdc2) in esophageal squamous cell carcinoma (ESCC), which has not been resolved. METHODS: Immunohistochemical expression of p34(cdc2) was examined for 91 cases of ESCC, and the relationship between the type of p34(cdc2) expression and the clinicopathologic features of the patients and tumors was analyzed. RESULTS: Forty-one ESCCs demonstrated cytoplasm dominant expression of p34(cdc2) and the other 50 ESCCs showed nuclei dominant p34(cdc2) expression. This differential expression pattern of p34(cdc2) did not reflect a prognostic aspect; however, the proportion of keratinizing tumors ESCCs with cytoplasm dominant expression of p34(cdc2) was significantly higher than that among ESCCs presenting nuclei dominant p34(cdc2) expression (P=0.006). CONCLUSION: Cellular differentiation in squamous cell carcinoma of the esophagus may be mediated by an intracellular localization of p34(cdc2). PMID- 15662528 TI - Dramatic development of severe SLE in a patient with an incomplete disease. AB - This case report describes the previously-unreported clinical course of a patient with a so-called incomplete systemic lupus erythematosus (SLE), i.e. symptoms related to one organ system only, together with the presence of ANA. He had an indolent course initially and developed, 6 months after the first symptoms, a severe disease with rapid appearance of major and unusual manifestations. The possibility of fast progression and a grave course of an incomplete SLE should be kept in mind. This report is meant to heighten awareness of such an atypical presentation so that prompt and aggressive immunosuppressive therapy may be instituted. PMID- 15662531 TI - A rare type of incarcerated lumbar hernia: CT and barium enema demonstration. PMID- 15662530 TI - A comparative study of the structure of human and murine greater omentum. AB - In humans, the greater omentum is a fatty peritoneal fold that extends from the greater curvature of the stomach to cover most abdominal organs. It performs many functions, which include acting as a reservoir of resident peritoneal inflammatory cells, a storage site for lipid, and a regulator of fluid exchange in and out of the peritoneal cavity. Most importantly, the omentum readily adheres to areas of inflammation and peritoneal damage, often leading to adhesion formation. Despite its clinical importance, the omentum remains an understudied organ, and discrepancies exist as to its exact morphology. This study uses a combination of phase contrast microscopy, scanning electron microscopy (SEM), and transmission electron microscopy (TEM) to elucidate the structure of the greater omentum of both human and mouse and determine whether it possesses a typical surface mesothelial cell lining similar to other serosa. Results indicated that both human and murine omenta were of similar structure and composed of two distinct types of tissue, one adipose-rich and the other translucent and membranous. The adipose-rich regions were well-vascularised and covered by a continuous mesothelial cell layer except at the sites of milky spots. In contrast, translucent areas were poorly vascularised and contained numerous fenestrations of varying size. The possible function and developmental origin of these gaps is unclear; however, their role in promoting omental adhesion formation and in the successful use of omental graft material is discussed. PMID- 15662532 TI - Permeability of marginal hybrid layers in composite restorations. AB - The goal of adhesive dentistry is to restore the peripheral seal of dentin lost from removal of enamel. Unfortunately, the hybrid layer (HL) that is used to create that seal is permeable to small ions or molecules, even in the absence of detectable, interfacial gap formation via nanoleakage. This nanoleakage results from several mechanisms including incomplete infiltration of adhesive monomers into demineralized collagen matrix, presence of hydrophilic monomers, and insufficient removal of solvent or water that remains trapped inside the HL. These mechanisms lead to a porous interface with nanometer-sized channels that increase the permeability of the HL. The null hypothesis tested in this study was that water and acidic solution storage are able to alter in vitro the resin dentin interface, further increasing the marginal hybrid layer (MHL) permeability. Class II cavities were made in vitro. The specimens were stored in water for 1 week and in lactic acid solution for 3 days. Polyvinyl siloxane impressions of restoration margins were taken before and after storage in water and lactic acid solution. Polyether replicas were obtained using the silicon impressions as molds. Replicas and original samples were observed under scanning electron microscopy. Lines of water droplets were detected on MHLs and overlying adhesive only after storage. Replicas obtained after acidic solution storage showed great numbers of irregularities such as gaps, voids, and degradation of the dentin-restoration surface margin, but also a great number of droplets. Dentin-restoration resin interfaces absorb water and are damaged by storage in dilute lactic acid. The presence of water droplets probably indicates water that flows out of the interface during the setting time of the impression and thus represents an index of marginal HL water permeability. PMID- 15662529 TI - The transformer2 gene in Musca domestica is required for selecting and maintaining the female pathway of development. AB - We present the isolation and functional analysis of a transformer2 homologue Mdtra2 in the housefly Musca domestica. Compromising the activity of this gene by injecting dsRNA into embryos causes complete sex reversal of genotypically female individuals into fertile males, revealing an essential function of Mdtra2 in female development of the housefly. Mdtra2 is required for female-specific splicing of Musca doublesex (Mddsx) which structurally and functionally corresponds to Drosophila dsx, the bottom-most regulator in the sex-determining pathway. Since Mdtra2 is expressed in males and females, we propose that Mdtra2 serves as an essential co-factor of F, the key sex-determining switch upstream of Mddsx. We also provide evidence that Mdtra2 acts upstream as a positive regulator of F supporting genetic data which suggest that F relies on an autocatalytic activity to select and maintain the female path of development. We further show that repression of male courtship behavior by F requires Mdtra2. This function of F and Mdtra2 appears not to be mediated by Mddsx, suggesting that bifurcation of the pathway at this level is a conserved feature in the genetic architecture of Musca and Drosophila. PMID- 15662533 TI - [Syphilis on the march. New reporting requirements]. AB - New public health laws in Germany require the reporting of some infectious agents directly but anonymously to the Robert Koch Institute (RKI). Included in this group are Treponema pallidum und HIV. Reporting responsibility lies with the director of the establishment where the causative agent was identified, usually a laboratory. The doctor who submits the clinical material for diagnosis is required to support the laboratory director in obtaining relevant information. The reporting form is filled out with the available information by the laboratory, sent to the RKI and a copy is sent to the submitting physician, who provides additional information and then sends the second form to the RKI, meeting his legal responsibilities. This new reporting system has provided reliable data about syphilis infections in Germany. In order to accumulate accurate data and plan effective prevention strategies, it is essential that all physicians comply with the new regulations. PMID- 15662534 TI - [Refractory cutaneous necrotizing vasculitis. Successful sulfone therapy]. AB - The clinical picture in vasculitis varies from few self-limiting symptoms to life threatening illnesses affecting multiple organs. We describe the course of a woman patient who had had a severe cutaneous necrotizing vasculitis for more than 10 years. Various immunsuppressants, including cyclophosphamide, had failed to improve the clinical course, although adequate doses were given for a sufficient period. Treatment with DADPS (diaminodiphenylsulfone, dapsone), however, finally led to complete remission of the severe and chronic vasculitic ulcerations. Indications for DADPS treatment of inflammatory cutaneous diseases are discussed, and an overview of cutaneous vasculitis is provided. PMID- 15662535 TI - [Long-term locally-recurrent melanoma]. AB - Malignant melanoma is a highly malignant skin tumor with a widely variable individual course. Currently, approximately 75% of German patients can be cured of malignant melanoma by early diagnosis and excision, while more than 20% die of distant metastases of the melanoma. In contrast, there are single cases of long term locally recurrent melanomas with neither complete cure nor development of distant metastases. We report on a female patient with a nevus-associated melanoma who was treated in 1968 with X-ray radiation (Dermopan, 100 Gy) with curative intent. In the following 31 years, the patient developed four phases of in-transit and satellite metastases in a zosteriform pattern as well as ilioinguinal lymph node metastasis. Since 1999, the patient has been free of recurrence. Besides the unusual biological features of this case, this report reflects the changes of paradigms in the therapy of melanoma over the last forty years. PMID- 15662536 TI - [Toxic phytodermatitis caused by Euphorbia helioscopia L. (sun spurge)]. AB - A 6-year-old patient presented with linear erythema and bullae on the face. The lesions developed after playing with plants the day before. The plant was identified as Euphorbia helioscopia L. (sun spurge). The sun spurge belongs to the Euphorbiaceae plant family. These plants produce a typical milky juice that causes toxic reactions following contact with skin and mucous membranes. In the literature several cases of toxic dermatitis caused by plants of the Euphorbiaeae family have been described. The most important differential diagnosis of these skin lesions is the bullous phototoxic dermatitis caused by psoralens. Plant induced toxic dermatitis is of increasing importance in dermatology. The exact determination of the causative plants is a prerequisite for the diagnosis of phytodermatitis. PMID- 15662537 TI - Orexigenic and anorexigenic mechanisms in the control of nutrition in chronic kidney disease. AB - Malnutrition is defined as abnormalities caused by an inadequate diet, but this term is often used inappropriately to describe the syndrome of loss of body weight with muscle mass being replaced by fatty tissue and declining serum proteins present in adults and children with chronic kidney disease (CKD). This syndrome is more accurately described as cachexia, and manifests as growth failure in children with CKD. Cachexia is common and is an important risk factor for poor quality of life and increased mortality and morbidity in both adults and children with CKD. Anorexia, acidosis and inflammation are important causes of cachexia, but the underlying molecular mechanism is not well understood. Dietary intake is often poor and resting metabolic rate is increased in CKD. The energy cost of growth is increased in experimental CKD. Circulating concentrations of cytokines, such as leptin, tumor necrosis factor-alpha and interleukins 1 and 6 are increased in patients with CKD and correlate with the degree of cachexia in these individuals. We hypothesize that cytokines signal through orexigenic neuropetides such as agouti-related peptide and neuropeptide Y (NPY), and anorexigenic neuropetides such as proopiomelanocortin and alpha-melanocyte stimulating hormone in the arcuate nucleus in the hypothalamus. This signaling system also involves the NPY receptor and the melanocortin receptors and controls appetite and metabolic rate in health and disease. Furthermore, the first order neurons of this system are located outside the blood-brain barrier and can therefore sense the circulating levels of cytokines, as well as long-term satiety hormones such as leptin and insulin and short-term satiety hormones such as ghrelin and peptide (P) YY. There is experimental evidence that this hypothalamic neuropeptide signaling system may have an important role in the pathogenesis of cachexia in CKD. Understanding the molecular mechanism of cachexia in CKD may lead to novel therapeutic strategies. PMID- 15662538 TI - Treatment of secondary hyperparathyroidism of dialysis patients with calcimimetics as a valuable addition to established therapeutic means. AB - Currently available options for the treatment of hyperparathyroidism secondary to chronic renal failure do not allow the achievement of target values for plasma calcium, phosphorus, and parathyroid hormone in the majority of patients with chronic kidney disease (CKD) stage 5. This is particularly true for CKD patients who have been referred to nephrologists late in the course of their disease and in whom prevention has not been possible. The advent of a new class of therapeutic agents, the calcimimetics, will allow an easier control of already established parathyroid overfunction, as has been demonstrated in several phase II studies and one phase III study with cinacalcet. Future studies will show whether an earlier start of treatment in patients with CKD stage 2, 3, and 4 allows the prevention of secondary hyperparathyroidism. Since all available experience has been gathered in adult patients it is also necessary to test the efficacy and safety of the calcimimetics in children with CKD. PMID- 15662539 TI - Gene expression profiling: cell cycle deregulation and aneuploidy do not cause breast cancer formation in WAP-SVT/t transgenic animals. AB - Microarray studies revealed that as a first hit the SV40 T/t antigen causes deregulation of 462 genes in mammary gland cells (ME cells) of WAP-SVT/t transgenic animals. The majority of deregulated genes are cell proliferation specific and Rb-E2F dependent, causing ME cell proliferation and gland hyperplasia but not breast cancer formation. In the breast tumor cells a further 207 genes are differentially expressed, most of them belonging to the cell communication category. In tissue culture breast tumor cells frequently switch off WAP-SVT/t transgene expression and regain the morphology and growth characteristics of normal ME cells, although the tumor-revertant cells are aneuploid and only 114 genes regain the expression level of normal ME cells. The profile of retransformants shows that only 38 deregulated genes are tumor specific, and that none of them is considered to be a typical breast cancer gene. PMID- 15662540 TI - Modulation of Toll-interleukin 1 receptor mediated signaling. AB - Toll-like receptors (TLRs) belong to the Toll-interleukin 1 receptor superfamily, which is defined by a common intracellular Toll-IL-1 receptor (TIR) domain. A group of TIR domain containing adaptors (MyD88, TIRAP, TRIF and TRAM), are differentially recruited to the Toll-IL-1 receptors, contributing to the specificity of signaling. The IL-1 mediated signaling pathway serves as a "prototype" for other family members. Genetic and biochemical studies reveal that IL-1R uses adaptor molecule MyD88 to mediate a very complex pathway, involving a cascade of kinases organized by multiple adapter molecules into signaling complexes, leading to activation of the transcription factor NFkappaB. Several Toll-like receptors utilize variations of the "prototype" pathway by employing different adaptor molecules. Double-stranded RNA triggered, TLR3-mediated signaling is independent of MyD88, IRAK4, and IRAK. The adapter molecule TRIF is utilized by TLR3 to mediate the activation of NFkappaB and IRF3. LPS-induced, TLR4-mediated signaling employs multiple TIR-domain containing adaptors, MyD88/TIRAP to mediate NFkappaB activation, TRIF/TRAM for IRF3 activation. Recent studies have also begun to unravel how these pathways are negatively regulated. SIGIRR (also known as TIR8), a member of TIR superfamily that does not activate the transcription factors NFkappaB and IRF3, instead negatively modulates responses. Cells from SIGIRR-null mice show enhanced activation in response to either IL-1 or certain Toll ligands. In addition to SIGIRR, several other negative regulators have been shown to inhibit the TIR signaling, including ST2, IRAKM, MyD88s, SOCS1, and Triad3A. The coordinated positive and negative regulation of the TIR signaling ensures the appropriate modulation of the innate and inflammatory responses. PMID- 15662541 TI - Low back pain prevention's effects in schoolchildren. What is the evidence? AB - Given the high prevalence rates of back pain, as early as in childhood, there has been a call for early preventive interventions. To determine which interventions are used to prevent back problems in schoolchildren, as well as what the evidence is for their utility, the literature was searched to locate all investigations that used subjects under the age of 18 and not seeking treatment. Included investigations were specifically designed as an intervention for low back pain (LBP) prevention. Additionally, a literature search was performed for modifiable risk factors for LBP in schoolchildren. The literature-update search was performed within the scope of the "COST Action B13" of the European Commission, approved for the development of European guidelines for the management of LBP. It was concluded that intervention studies in schoolchildren focusing on back-pain prevention are promising but too limited to formulate evidence-based guidelines. On the other hand, since the literature shows that back-pain reports about schoolchildren are mainly associated with psychosocial factors, the scope for LBP prevention in schoolchildren may be limited. However, schoolchildren are receptive to back-care-related knowledge and postural habits, which may play a preventive role for back pain in adulthood. Further studies with a follow-up into adulthood are needed to evaluate the long-term effect of early interventions and the possible detrimental effect of spinal loading at young age. PMID- 15662542 TI - Radiolucent cage for cervical vertebral reconstruction: a prospective study of 17 cases with 2-year minimum follow-up. AB - In cervical spondylotic myelopathy, extended anterior spinal cord decompression necessitates subsequent stable vertebral reconstruction. Reconstruction with an iliac crest graft and screw-plate fixation gives satisfactory clinical and radiological results, but they are often compromised by morbidity involving the bone harvest. The purpose of this study was to evaluate the contribution to cervical reconstruction of a biocompatible, radiolucent cage combined with screw plate fixation, making use of bone harvested in situ. This prospective study was performed between July 2000 and March 2001 in eight women and nine men (mean age, 55 years) operated for cervical spondylotic myelopathy. Situated between levels C3 and C6, the cage was inserted after one corporectomy in ten patients, two corporectomies in five patients, and three corporectomies in two patients. The cage consisted of a polyester mesh impregnated with poly-L-lactic acid (PLLA) conferring temporary rigidity to the cage during bony fusion. Clinical and radiological follow-up (plain films, computed tomographic reconstruction in three cases) was performed at 2 months, 6 months, 12 months, 24 months and 36 months, postoperatively, with a mean follow-up of 30 months. Functional results were evaluated according to the Japanese Orthopaedic Association's scoring system. An independent surgeon assessed the radiological evidence of anterior cervical fusion using the grades proposed by Bridwell [6]. Every patient experienced neurological recovery. At last follow-up, radiological findings were consistent with grade I (complete fusion) in five cases, grade II (probable fusion) in ten cases, grade III (radiolucent halo in favor of non fusion) in one case, and grade IV (graft lysis) in one case with persistent neck pain. In three cases there was screw breakage (two grade II, one grade IV). None of these cases required surgical revision at latest follow-up. In extensive spinal cord decompression through an anterior approach, cervical reconstruction using the present type of cage can achieve clinical results comparable to conventional techniques. The rigidity of the cage meets biomechanical imperatives. Its radiolucency permits one to monitor the course of consolidation, contrary to metal cages. The cases of probable non-fusion and screw breakage were not accompanied by signs of instability on the flexion extension films. This cage meets the biologic and biomechanical imperatives of cervical reconstruction. It obviates complications involving bone harvest. PMID- 15662544 TI - Scale-up of biopesticide production processes using wastewater sludge as a raw material. AB - Studies were conducted on the production of Bacillus thuringiensis (Bt)-based biopesticides to ascertain the performance of the process in shake flasks, and in two geometrically similar fermentors (15 and 150 l) utilizing wastewater sludge as a raw material. The results showed that it was possible to achieve better oxygen transfer in the larger capacity fermentor. Viable cell counts increased by 38-55% in the bioreactor compared to shake flasks. As for spore counts, an increase of 25% was observed when changing from shake flask to fermentor experiments. Spore counts were unchanged in bench (15 l) and pilot scale (5.3-5.5 e(+08) cfu/ml; 150 l). An improvement of 30% in the entomotoxicity potential was obtained at pilot scale. Protease activity increased by two to four times at bench and pilot scale, respectively, compared to the maximum activity obtained in shake flasks. The maximum protease activity (4.1 IU/ml) was obtained in pilot scale due to better oxygen transfer. The Bt fermentation process using sludge as raw material was successfully scaled up and resulted in high productivity for toxin protein yield and a high protease activity. PMID- 15662545 TI - The effect of nitrogen source on yield and glycosylation of a human cystatin C mutant expressed in Pichia pastoris. AB - Human cystatin C is a cysteine proteinase inhibitor with potential applications as an anti-viral agent, cancer tumor growth inhibitor, and in prevention of proteolysis during food processing. A glycosylated cystatin C mutant with increased temperature stability was developed for the latter application [Nakamura et al. (1998) FEBS Lett 427:252-254]. A recombinant variant of cystatin C [Nakamura et al. (2000) International patent no. PLTCA99/00717] with two potential sites for N-linked glycosylation was expressed in Pichia pastoris Mut(s). Little of the cystatin C produced was in the glycosylated form under fermentation conditions of pH 6, temperature 28 degrees C, methanol only feed, and ammonium hydroxide as a nitrogen source. Thus, the effect of addition of complex nitrogen sources, peptone and amino acid supplements, on the yield and glycosylation of this mutant cystatin C were investigated. A full factorial design experiment using 2-l fermenters was performed with three factors: ammonium hydroxide, peptone, and an amino acid mix, at two levels, absent or present. Peptone addition was found to have a positive, and the most significant, effect on cell specific cystatin C yield. A maximum mutant cystatin C yield of 0.82 mumol (g-dry cell weight)(-1) min(-1) was obtained when all three nitrogen sources were used together. However, under these conditions only 16% of protein was in the glycosylated form since ammonia was found to have a significant negative effect on glycosylation extent. The maximum extent of glycosylation was 30% when peptone and amino acid mix were the only nitrogen sources added. PMID- 15662546 TI - Fermentation of a milk-soymilk and Lycium chinense Miller mixture using a new isolate of Lactobacillus paracasei subsp. paracasei NTU101 and Bifidobacterium longum. AB - A milk-soymilk mixture was fermented using Lactobacillus paracasei subsp. paracasei NTU101 and Bifidobacterium longum BCRC11847 at different inoculum ratios (1:1, 1:2, 1:5, 2:1, and 5:1). When the inoculum ratio was 1:2, the cell numbers of both strains were balanced after 12 h of cultivation. The pH and titratable acidity were very similar at the various inoculum ratios of cultivation. The milk-soymilk mixture was supplemented with 5, 10, 15, and 20% Lycium chinense Miller juice and fermented with Lactobacillus paracasei subsp. paracasei NTU101 and B. longum BCRC11847. Sensory evaluation results showed that supplementation with 5% Lycium chinense Miller juice improved the acceptability of the fermented milk-soymilk. The fermented beverage was stored at 4 degrees C for 14 days; variations in pH and titratable acidity were slight. The cell numbers of L. paracasei subsp. paracasei NTU101 and B. longum BCRC11847 in the fermented beverage were maintained at 1.2x10(9) CFU/ml and 6.3x10(8) CFU/ml, respectively, after 14 days of storage. PMID- 15662547 TI - Amputation as a marker of the quality of foot care in diabetes. AB - Strategic targets for the management of foot ulcers focus on reducing the incidence of amputation. While data on the incidence of amputation can be obtained relatively easily, the figures require very careful interpretation. Variation in the definition of amputation, population selection and the choice of numerator and denominator make comparisons difficult. Major and minor amputation have to be distinguished as they are undertaken for different reasons and are associated with different costs and functional implications. Many factors influence the decision of whether or not to remove a limb. In addition to disease severity, co-morbidities, and social and individual patient factors, many aspects of the structure of care services affect this decision, including access to primary care, quality of primary care, delays in referral, availability and quality of specialist resources, and prevailing medical opinion. It follows that a high incidence of amputation can reflect a higher disease prevalence, late referral, limited resources, or a particularly interventionist approach by a specialist team. Conversely, a low incidence of amputation can indicate a lower disease prevalence or severity, good management of diabetes in primary and secondary care, or a particularly conservative approach by an expert team. An inappropriately conservative approach could conceivably enhance suffering by condemning a person to months of incapacity before they die with an unhealed ulcer. The reported annual incidence of major amputation in industrialised countries ranges from 0.06 to 3.83 per 10(3) people at risk. Some centres have documented that the incidence is falling, but this is often from a baseline value that was unusually high. Other centres have reported that the incidence has not changed. The ultimate target is to achieve not only a decrease in incidence, but also a low overall incidence. This must be accompanied by improvements in morbidity, mortality, and patient function and mood. PMID- 15662548 TI - Acute effects of glucose and insulin on vascular endothelium. AB - AIMS/HYPOTHESIS: Chronic exposure to high concentrations of glucose has consistently been demonstrated to impair endothelium-dependent, nitric oxide (NO) mediated vasodilation. In contrast, several clinical investigations have reported that acute exposure to high glucose, alone or in combination with insulin, triggers vasodilation. The aim of this study was to examine whether elevated glucose itself stimulates endothelial NO formation or enhances insulin-mediated endothelial NO release. METHODS: We measured NO release and vessel tone ex vivo in porcine coronary conduit arteries (PCAs). Intracellular Ca(2+) was monitored in porcine aortic endothelial cells (PAECs) by fura-2 fluorescence. Expression of the Na(+)/glucose cotransporter-1 (SGLT-1) was assayed in PAECs and PCA endothelium by RT-PCR. RESULTS: Stimulation of PCAs with D: -glucose, but not the osmotic control L: -glucose, induced a transient increase in NO release (EC(50) approximately 10 mmol/l), mediated by a rise in intracellular Ca(2+) levels due to an influx from the extracellular space. This effect was abolished by inhibitors of the plasmalemmal Na(+)/Ca(2+) exchanger (dichlorobenzamil) and the SGLT-1 (phlorizin), which was found to be expressed in aortic and coronary endothelium. Alone, D: -glucose did not relax PCA, but did augment the effect of insulin on NO release and vasodilation. CONCLUSIONS/INTERPRETATION: An increased supply of extracellular D: -glucose appears to enhance the activity of the endothelial isoform of nitric oxide synthase by increasing intracellular Na(+) concentrations via SGLT-1, which in turn stimulates an extracellular Ca(2+) influx through the Na(+)/Ca(2+) exchanger. This mechanism may be responsible for glucose-enhanced, insulin-dependent increases in tissue perfusion (including coronary blood-flow), thus accelerating glucose extraction from the blood circulation to limit the adverse vascular effects of prolonged hyperglycaemia. PMID- 15662549 TI - Antioxidative treatment prevents activation of death-receptor- and mitochondrion dependent apoptosis in the hearts of diabetic rats. AB - AIMS/HYPOTHESIS: The mechanisms by which glucose injures cells of the cardiovascular system include generation of reactive oxygen species and induction of cellular apoptosis. To date, little is known about the molecular events of hyperglycaemia-induced apoptosis in the heart in vivo. METHODS: Male Sprague Dawley rats were rendered diabetic by a single intraperitoneal injection of 60 mg/kg body weight streptozotocin. Caspase activities in cardiac ventricular tissue were determined using fluorometric and immunoassay caspase-activity assay kits respectively. Expression levels of proteins of the apoptotic cascade were determined with western blot analyses using specific antibodies. RESULTS: Four weeks of hyperglycaemia induced significant apoptosis in cardiac tissue. Determining the initiators of death-receptor-dependent apoptosis revealed induction of CD95/Fas and caspase-8. Examination of the activities of effector caspases revealed increased activity of caspase-6, but not caspase-3 and -7. On evaluating inhibitors of apoptosis, we found up-regulation of caspase-3 and -7 inhibiting X-linked inhibitors of apoptosis in diabetic rats. Hyperglycaemia also induced significant mitochondrion-dependent apoptosis. Our evaluation of expression levels of Bcl-2 family members showed increased expression of pro apoptotic Bak and Bax in diabetic rats. Antioxidative treatment with lipoic acid significantly suppressed apoptosis and down-regulated caspase-6, -8 and -9 activity, as well as expression levels of pro-apoptotic Bcl-2 proteins without changing blood glucose levels. CONCLUSIONS/INTERPRETATION: The present study indicates that reactive oxygen species induced by high glucose are involved in both death-receptor- and mitochondrion-dependent apoptosis in the heart in vivo. It also suggests that antioxidants may be a therapeutic option for preventing cardiovascular damage in diabetes mellitus in humans. PMID- 15662550 TI - Characterisation of glucose transporters in the intact coronary artery endothelium in rats: GLUT-2 upregulated by long-term hyperglycaemia. AB - AIMS/HYPOTHESIS: We have examined the effects of streptozotocin-induced type 1 diabetes on the expression and subcellular distribution of the classic sugar transporters (GLUT-1 to 5 and sodium-dependent glucose transporter-1 [SGLT-1]) in the endothelial cells of an en face preparation of septal coronary artery from Wistar rats. METHODS: The presence of the GLUT isoforms and SGLT-1 in the endothelial cell layer was determined by immunohistochemistry using wide-field fluorescence microscopy coupled to deconvolution, and was quantified by digital image analysis. RESULTS: We found that all of the transporters were expressed within these cells and that all except SGLT-1 were preferentially located on the abluminal side. The heaviest labelling was adjacent to the cell-to-cell junctions where the luminal and abluminal membranes are in close proximity, which may reflect a spatial organisation specialised for vectorial glucose transport across the thinnest part of the cytoplasm. Long-term hyperglycaemia, induced by streptozotocin, significantly downregulated GLUT-1, 3, 4 and 5 and dramatically upregulated GLUT-2, leaving SGLT-1 unchanged. CONCLUSIONS/INTERPRETATION: We conclude that the high susceptibility of endothelial cells to glucose toxicity may be the result of the subcellular organisation of their GLUTs and the increased expression of GLUT-2. PMID- 15662551 TI - Chronic exposure to high glucose impairs bradykinin-stimulated nitric oxide production by interfering with the phospholipase-C-implicated signalling pathway in endothelial cells: evidence for the involvement of protein kinase C. AB - AIMS/HYPOTHESIS: Overwhelming evidence indicates that endothelial cell dysfunction in diabetes is characterised by diminished endothelium-dependent relaxation, but the matter of the underlying molecular mechanism remains unclear. As nitric oxide (NO) production from the endothelium is the major player in endothelium-mediated vascular relaxation, we investigated the effects of high glucose on NO production, and the possible alterations of signalling pathways implicated in this scenario. METHODS: NO production and intracellular Ca(2+) levels ([Ca(2+)](i)) were assessed using the fluorescent probes 4,5 diaminofluorescein diacetate and fura-2 respectively. RESULTS: Exposure of cultured bovine aortic endothelial cells to high glucose for 5 or 10 days significantly reduced NO production induced by bradykinin (but not by Ca(2+) ionophore) in a time- and dose-dependent manner. This was probably due to an attenuation in bradykinin-induced elevations of [Ca(2+)](i) under these conditions, since a close correlation between [Ca(2+)](i) increases and NO generation was observed in intact bovine aortic endothelial cells. Both bradykinin-promoted intracellular Ca(2+) mobilisation and extracellular Ca(2+) entry were affected. Moreover, bradykinin-induced formation of Ins(1,4,5)P(3), a phospholipase C product leading to increases in [Ca(2+)](i), was also inhibited following high glucose culture. This abnormality was not attributable to a decrease in inositol phospholipids, but possibly to a reduction in the number of bradykinin receptors. The alterations in NO production, the increases in [Ca(2+)](i), and the bradykinin receptor number due to high glucose could be largely reversed by protein kinase C inhibitors and D: -alpha-tocopherol (antioxidant). CONCLUSIONS/INTERPRETATION: Chronic exposure to high glucose reduces NO generation in endothelial cells, probably by impairing phospholipase-C mediated Ca(2+) signalling due to excess protein kinase C activation. This defect in NO release may contribute to the diminished endothelium-dependent relaxation and thus to the development of cardiovascular diseases in diabetes. PMID- 15662552 TI - Prediction of the risk of cardiovascular mortality using a score that includes glucose as a risk factor. The DECODE Study. AB - AIMS/HYPOTHESIS: Risk scores have been developed to predict cardiovascular or coronary risk, and while most have included diabetes as a risk factor, none have included lower glucose concentrations, either at fasting or following a 2-h oral glucose tolerance test. This article develops 5- and 10-year risk scores for cardiovascular mortality that include glucose concentrations as well as known diabetes status. METHODS: Data is from the DECODE cohort: 16,506 men and 8,907 women from 14 European studies. The risk factors studied were as follows: age, fasting and 2-h glucose (including cases of known diabetes), fasting glucose alone (including cases of known diabetes), cholesterol, smoking status, systolic blood pressure and BMI. For an absolute risk score the 1995 country- and sex specific cardiovascular death rates were used. RESULTS: In men, for both 5- and 10-year cardiovascular mortality, after adjusting for age and study centre, all studied risk factors, except BMI, were significantly associated with cardiovascular mortality (p<0.05). These results were unchanged in multivariate models with all factors included. In women, after adjusting for age and centre, glucose categories, systolic blood pressure and BMI were predictive of 5-year cardiovascular mortality. With all factors in the model, only age and glucose categories were predictive. In terms of 10-year cardiovascular mortality, smoking status and blood pressures were also predictive in the women. For men and women, the same scores were used for the risk factors, except for age and glucose categories where the hazard ratios differed significantly. CONCLUSIONS/INTERPRETATION: Including glucose concentrations as well as diabetic status provides quantitative information on cardiovascular risk prediction. PMID- 15662553 TI - Joint role of non-HDL cholesterol and glycated haemoglobin in predicting future coronary heart disease events among women with type 2 diabetes. AB - AIMS/HYPOTHESIS: Non-HDL cholesterol (the sum of LDL, VLDL and IDL cholesterol) is considered to be particularly valuable in the management of dyslipidaemia in type 2 diabetes. However, it remains uncertain whether the association between non-HDL cholesterol and cardiovascular risk in type 2 diabetes depends on the status of hyperglycaemia. We aimed to determine whether non-HDL cholesterol predicts CHD events among diabetic women independently of currently established risk factors and the status of glycaemic control. METHODS: We prospectively followed 921 diabetic women in the Nurses' Health Study, who were free of cardiovascular disease at the time that blood was drawn in 1989/90. During 10 years of follow-up, we identified 122 incident CHD cases. RESULTS: After adjustment for age, BMI, smoking, alcohol consumption, and other lifestyle risk factors, the multivariate relative risks (RRs) of CHD for extreme quartiles were 1.97 (95% CI: 1.14-3.43) for non-HDL cholesterol, 1.78 (1.02-3.11) for apolipoprotein B-100, and 1.93 (1.15-3.22) for LDL cholesterol. However, the association between non-HDL cholesterol and CHD risk was only apparent among women with elevated fasting triglycerides (RR for extreme quartiles: 3.80; p=0.045). HbA(1)c was strongly associated with increased CHD risk (RR for increase by 1 unit: 1.24; 95% CI: 1.13-1.35), and both non-HDL cholesterol and HbA(1)c additively predicted CHD risk (RR for the combination of high non-HDL cholesterol and high HbA(1)c [tertiles]: 4.59). CONCLUSIONS/INTERPRETATION: Our study suggests that non-HDL cholesterol and HbA(1)c are potent predictors of CHD risk in diabetic women. Therapies to lower CHD risk in diabetic patients should emphasise both glycaemic control and lipid lowering. PMID- 15662554 TI - Type 2 diabetes and risk of non-embolic ischaemic stroke in Japanese men and women. AB - AIMS/HYPOTHESIS: The aim of this study was to examine the relationship between type 2 diabetes and risk of ischaemic stroke in Asian populations. METHODS: We conducted a 17-year prospective cohort study in 10,582 Japanese individuals (4287 men and 6295 women) aged 40-69 years living in five communities in Japan. All subjects were free of stroke and CHD at baseline. Diabetes was defined as a fasting glucose level of >/=7.0 mmol/l, a non-fasting glucose of >/=11.1 mmol/l, or receiving medication for diabetes. RESULTS: The risk of non-embolic ischaemic stroke was approximately two-fold higher in diabetic subjects than in subjects with normal glucose levels. The multivariate relative risk after adjustment for age, community, hypertensive status, BMI, triceps and subscapular skinfold thickness (TSF and SSF), and other known cardiovascular risk factors was 1.8 (95% CI 1.0-3.2) for men and 2.2 (1.2-4.0) for women. This excess risk was primarily observed among non-hypertensive subjects and individuals with higher values for measures of adiposity (BMI, TSF and SSF values above the median), particularly those with higher values for SSF. The association between non-embolic ischaemic stroke and glucose abnormality was particularly strong among non-hypertensive subjects with higher SSF values: the multivariate relative risk was 1.9 (1.0-3.7) for borderline diabetes and 4.9 (2.5-9.5) for diabetes. CONCLUSIONS/INTERPRETATION: In this cohort, type 2 diabetes was a significant risk factor for non-embolic ischaemic stroke, particularly in non-hypertensive and non-lean individuals. Due to the nationwide decrease in blood pressure and increase in mean BMI among the Japanese population, with current levels approaching those observed in Western countries, the impact of glucose abnormalities on risk of ischaemic stroke represents a forthcoming public health issue in Japan. PMID- 15662555 TI - Coffee consumption and glucose tolerance status in middle-aged Japanese men. AB - AIMS/HYPOTHESIS: Several studies have reported that coffee has a protective effect against the development of type 2 diabetes. However, few of these studies used the standard glucose tolerance test to diagnose type 2 diabetes. The aim of this study was to investigate the relationship between coffee and green tea consumption and glucose tolerance status as determined using a 75-g OGTT. METHODS: We performed a cross-sectional study of 3224 male officials of the self defence forces. Glucose tolerance status was determined in accordance with the 1998 World Health Organization criteria, and average intakes of coffee and green tea over the previous year were assessed by a self-administered questionnaire. The figures obtained were adjusted for BMI, physical activity and other factors. RESULTS: A total of 1130 men were identified as having glucose intolerance (IFG, IGT or type 2 diabetes). Compared with those who did not consume coffee on a daily basis, fasting and 2-h post-load plasma glucose levels were 1.5% and 4.3% lower in those who drank 5 cups of coffee or more per day respectively. The adjusted odds ratios of glucose intolerance for categories of <1, 1-2, 3-4 and >/=5 cups of coffee per day were 1.0 (referent), 0.8 (95% CI 0.6-1.0), 0.7 (95% CI 0.6-0.9) and 0.7 (95% CI 0.5-0.9) respectively (p=0.0001 for trend). No clear association was observed between green tea drinking and glucose tolerance status. CONCLUSIONS/INTERPRETATION: Coffee consumption may inhibit postprandial hyperglycaemia and thereby protect against the development of type 2 diabetes mellitus. PMID- 15662556 TI - Coffee consumption and incidence of impaired fasting glucose, impaired glucose tolerance, and type 2 diabetes: the Hoorn Study. AB - AIMS/HYPOTHESIS: Coffee contains several substances that may affect glucose metabolism. The aim of this study was to evaluate the relationship between habitual coffee consumption and the incidence of IFG, IGT and type 2 diabetes. METHODS: We used cross-sectional and prospective data from the population-based Hoorn Study, which included Dutch men and women aged 50-74 years. An OGTT was performed at baseline and after a mean follow-up period of 6.4 years. Associations were adjusted for potential confounders including BMI, cigarette smoking, physical activity, alcohol consumption and dietary factors. RESULTS: At baseline, a 5 cup per day higher coffee consumption was significantly associated with lower fasting insulin concentrations (-5.6%, 95% CI -9.3 to -1.6%) and 2-h glucose concentrations (-8.8%, 95% CI -11.8 to -5.6%), but was not associated with lower fasting glucose concentrations (-0.8%, 95% CI -2.1 to 0.6%). In the prospective analyses, the odds ratio (OR) for IGT was 0.59 (95% CI 0.36-0.97) for 3-4 cups per day, 0.46 (95% CI 0.26-0.81) for 5-6 cups per day, and 0.37 (95% CI 0.16-0.84) for 7 or more cups per day, as compared with the corresponding values for the consumption of 2 or fewer cups of coffee per day (p=0.001 for trend). Higher coffee consumption also tended to be associated with a lower incidence of type 2 diabetes (OR 0.69, CI 0.31-1.51 for >/=7 vs /=7 vs C SNP in the 3' untranslated region showed borderline association (p~0.06) when evidence for linkage was taken into account. However, replication studies (350 young-onset diabetic patients; 747 controls) failed to confirm any relationship with diabetes for this variant. No significant associations were seen for diabetes-related traits including BMI and waist-to-hip ratio. CONCLUSIONS/INTERPRETATION: The present study, the first reported analysis of this gene, indicates that the SCD variants typed do not explain chromosome-10-encoded susceptibility to type 2 diabetes. Although this study provided no evidence that SCD sequence variation influences diabetes susceptibility or related traits, SCD remains a major target for pharmaceutical and/or environmental manipulation. PMID- 15662558 TI - Effects of lipase inhibition on gastric emptying of, and on the glycaemic, insulin and cardiovascular responses to, a high-fat/carbohydrate meal in type 2 diabetes. AB - AIMS/HYPOTHESES: We examined the effects of lipase inhibition with orlistat on (i) gastric emptying of, and (ii) the glycaemic, glucagon-like peptide-1 (GLP-1) and cardiovascular responses to, a high-fat/carbohydrate meal in type 2 diabetic patients. METHODS: Eight type 2 diabetic patients, who were aged 62 years (median range: 49-68 years) and managed by diet alone, consumed a meal containing 65 g powdered potato, 20 g glucose reconstituted with 200 ml water (labelled with 20 MBq (99m)Tc-sulphur-colloid) and 45 g margarine. They did this on two separate occasions, with and without 120 mg orlistat, and while in the seated position with their back against a gamma camera. Venous blood samples for measurement of blood glucose, plasma insulin and GLP-1 were obtained immediately before the meal and at regular intervals afterwards. Blood pressure (systolic and diastolic) and heart rate were measured using an automated device. RESULTS: Gastric emptying of the meal was faster after orlistat than without orlistat (50% emptying time [mean +/- SEM], 61+/-8 min vs 98+/-5 min; p=0.0001). In the first 60 min after the meal blood glucose (p=0.001) and plasma insulin (p=0.01) concentrations were higher in patients who had taken orlistat; between 60 and 180 min plasma GLP-1 (p=0.02) concentrations were lower after orlistat than without orlistat. Between 0 and 30 min systolic blood pressure (p=0.003) was lower, and heart rate (p=0.03) greater in subjects who had taken orlistat than in those who had not. CONCLUSIONS/INTERPRETATION: Inhibition of fat digestion by orlistat may-as a result of more rapid gastric emptying-exacerbate postprandial glycaemia and the postprandial fall in blood pressure in patients with type 2 diabetes after ingestion of meals containing fat and carbohydrate. PMID- 15662559 TI - Thiazolidinediones improve insulin sensitivity in adipose tissue and reduce the hyperlipidaemia without affecting the hyperglycaemia in a transgenic model of type 2 diabetes. AB - AIM/HYPOTHESIS: The aim of this study was to examine the effects of thiazolidinediones on the MKR mouse model of type 2 diabetes. METHODS: Six-week old wild-type (WT) and MKR mice were fed with or without rosiglitazone or pioglitazone for 3 weeks. Blood was collected from the tail vein for serum biochemistry analysis. Hyperinsulinaemic-euglycaemic clamp analysis was performed to study effects of thiazolidinediones on insulin sensitivity of tissues in MKR mice. Northern blot analysis was performed to measure levels of target genes of PPAR gamma agonists in white adipose tissue and hepatic gluconeogenic genes. RESULTS: Thiazolidinedione treatment of MKR mice significantly lowered serum lipid levels and increased serum adiponectin levels but did not affect levels of blood glucose and serum insulin. Hyperinsulinaemic-euglycaemic clamp showed that whole-body insulin sensitivity and glucose homeostasis failed to improve in MKR mice after rosiglitazone treatment. Insulin suppression of hepatic endogenous glucose production failed to improve in MKR mice following rosiglitazone treatment. This lack of change in hepatic insulin insensitivity was associated with no change in the ratio of HMW : total adiponectin, hepatic triglyceride content, and sustained hepatic expression of PPAR gamma and stearoyl-CoA desaturase 1 mRNA. Interestingly, rosiglitazone markedly enhanced glucose uptake by white adipose tissue with a parallel increase in CD36, aP2 and GLUT4 gene expression. CONCLUSIONS/INTERPRETATION: These data suggest that potentiation of insulin action on tissues other than adipose tissue is required to mediate the antidiabetic effects of thiazolidinediones in our MKR diabetic mice. PMID- 15662560 TI - High-dose thiamine therapy counters dyslipidaemia in streptozotocin-induced diabetic rats. AB - AIMS/HYPOTHESIS: Cardiovascular disease in diabetes is linked to increased risk of atherosclerosis, increased levels of triglyceride-rich lipoproteins and enhanced hepatic lipogenesis. The hepatic hexosamine pathway has been implicated in signalling for de novo lipogenesis by the liver. In this study, we assessed if decrease of flux through the hexosamine pathway induced by high-dose thiamine therapy counters diabetic dyslipidaemia. METHODS: The model of diabetes used was the streptozotocin-induced diabetic rat with maintenance insulin therapy. Normal control and diabetic rats were studied for 24 weeks with and without oral high dose therapy (7 and 70 mg/kg) with thiamine and benfotiamine. Plasma total cholesterol, HDL cholesterol and triglycerides were determined at 6-week intervals and hepatic metabolites and transketolase activity after death of the rats at 24 weeks. RESULTS: We found that thiamine therapy (70 mg/kg) prevented diabetes-induced increases in plasma cholesterol and triglycerides in diabetic rats but did not reverse the diabetes-induced decrease of HDL. This was achieved by prevention of thiamine depletion and decreased transketolase activity in the liver of diabetic rats. There was a concomitant decrease in hepatic UDP-N acetylglucosamine and fatty acid synthase activity. Thiamine also normalised food intake of diabetic rats. A lower dose of thiamine (7 mg/kg) and the thiamine monophosphate prodrug benfotiamine (7 and 70 mg/kg) were ineffective. CONCLUSIONS/INTERPRETATION: High-dose thiamine therapy prevented diabetic dyslipidaemia in experimental diabetes probably by suppression of food intake and hexosamine pathway signalling but other factors may also be involved. Benfotiamine was ineffective. PMID- 15662561 TI - Allosteric mechanisms in ACT domain containing enzymes involved in amino acid metabolism. AB - An important sequence motif identified by sequence analysis is shared by the ACT domain family, which has been found in a number of diverse proteins. Most of the proteins containing the ACT domain seem to be involved in amino acid and purine synthesis and are in many cases allosteric enzymes with complex regulation enforced by the binding of ligands. Here we explore the current understanding of the ACT domain function including its role as an allosteric module in a selected group of enzymes. We will further describe in more detail three of the proteins where some understanding is available on function and structure: i) the archetypical ACT domain protein E. coli 3PGDH, which catalyzes the first step in the biosynthesis of L-Ser, ii) the bifunctional chorismate mutase/prephenate dehydratase (P-protein) from E. coli, which catalyzes the first two steps in the biosynthesis of L-Phe, and iii) the mammalian aromatic amino acid hydroxylases, with special emphasis on phenylalanine hydroxylase, which catalyzes the first step in the catabolic degradation of L-phenylalanine (L-Phe). The ACT domain is commonly involved in the binding of a small regulatory molecule, such as the amino acids L-Ser and L-Phe in the case of 3PGDH and P-protein, respectively. On the other hand, for PAH, and probably for other enzymes, this domain appears to have been incorporated as a handy, flexible small module with the potential to provide allosteric regulation via transmission of finely tuned conformational changes, not necessarily initiated by regulatory ligand binding at the domain itself. PMID- 15662562 TI - Pseudo-peptides derived from isomannide as potential inhibitors of serine proteases. AB - Hepatitis C, Dengue and West Nile virus are among of the most important flaviviruses that share one important serine protease enzyme. Serine proteases belong to the most studied class of proteolytic enzymes, and are a primary target in the drug development field. In this paper, we describe the synthesis and preliminary molecular modeling studies of a novel class of N-t-Boc amino acid amides derived of isomannide as potential serine proteases inhibitors. PMID- 15662563 TI - Biochemical properties of new synthetic carnosine analogues containing the residue of 2,3-diaminopropionic acid: the effect of N-acetylation. AB - Three novel carnosine analogues 7-9 containing the residue of L(+)2,3 diaminopropionic acid with different degree of N-acetylation instead of beta alanine have been synthesized and characterized. Comparative analysis of hydrolysis by carnosinase revealed that the mono- and bis-acetylated compounds 8 and 9 are resistant to enzymatic hydrolysis and act as competitive inhibitors of this enzyme. The hydroxyl radical scavenging potential of the three analogues was evaluated by their ability to inhibit iron/H(2)O(2)-induced degradation of deoxyribose. The second-order rate constants of the reaction of compounds 7-9 with hydroxyl radical were almost identical to that of carnosine. These compounds were also found to act as protective agents against peroxynitrite-dependent damage as assessed by their ability to prevent nitration of free tyrosine induced by this species. PMID- 15662564 TI - Improved brain protection at decompressive craniectomy--a new method using Palacos R-40 (methylmethacrylate). AB - A new method is described for protecting the brain after decompressive craniectomy in which a temporary methylmethacrylate flap is formed, somewhat larger than the original bone flap, thus gaining "extra" volume for the oedematous brain in which to expand. The present procedure was developed as a part of ordinary clinical practice particularly in response to demands from the NICU staff and our colleagues at other clinics who were responsible for the care of the patient in the post NICU period. They made us keenly aware that these patients frequently lack optimal co-ordination and balance and therefore run an increased risk of trauma to the unprotected brain when falling. This prompted us to develop a method for brain protection after decompressive craniectomy aiding in the care and rehabilitation until the final installation of the patient's own bone flap can be performed. PMID- 15662565 TI - Predicting the risk of rupture of intracranial aneurysms based on anatomical location. AB - The location of intracranial aneurysms may be a significant independent risk factor for predicting aneurysm rupture. A recent report found high bleed rates from posterior communicating artery aneurysms which had not previously been noted. On this distinction hangs the decision whether to treat a large number of unruptured aneurysms. In the recent publication by the International Study of Unruptured Intracranial Aneurysms (ISUIA), two bleeds from small incidental posterior communicating artery aneurysms were noted and these aneurysms were reported to have a similar risk to aneurysms of the posterior circulation and as a result were grouped with them. This was a post hoc analysis so the justification for this assertion is tenuous. The hypothesis that posterior communicating aneurysms are of similar risk to posterior circulation aneurysms requires further testing on other data before it can be confidently accepted. A review of the literature was undertaken to define relative risks of rupture for different anatomical locations and to test the above hypothesis. Eleven papers were found to contain sufficient data to calculate rupture rates for anatomical sub groups. Studies contained a total of 30,204 patient years of follow up. Results showed the internal carotid artery to be the commonest site for unruptured aneurysms (38%). Aneurysms located in the posterior circulation had an overall annual bleed rate of 1.8%. This compares with 0.49% for the anterior circulation. The bleed rate from aneurysms of the posterior communicating artery (0.46% per year) was similar to that of the rest of the anterior circulation. The ISUIA post hoc hypothesis fails when tested on these data and the ISUIA data should be re-analysed with posterior communicating artery aneurysms grouped with the anterior circulation where they more traditionally belong. PMID- 15662566 TI - Dwarfism, occult spinal dysraphism, and presacral myxopapillary ependymoma with an epidermoid cyst in a child. AB - The authors present a case of a child with dwarfism that was noted to be developmentally delayed. Imaging revealed atlantoaxial instability, occult spinal dysraphism, and a presacral mass. Histopathology of the presacral lesion was that of a myxopapillary ependymoma with epidermoid cyst. We believe this to be the first report in the extant medical literature of this constellation of findings in the same patient. However, there are rare reports indicating a possible association of occult spinal dysraphism and the simultaneous occurrence of spinal ependymomas. Further case reports are necessary to discern whether these pathological entities are true low rate associations that the clinician should consider in their evaluation of these patients. PMID- 15662568 TI - Multiple spinal dural arteriovenous fistulas. AB - Multiple spinal dural arteriovenous fistulas (DAVFs) have been rarely reported and only two such cases are found in the literature. A 71-year-old man complained of muscle weakness and hypesthesia in both legs and angiographically diagnosed as thoracic DAVF. The fistula was surgically treated, however, the symptoms recurred 14 months after the first treatment. Angiography revealed a new fistula in the lumbar region and this was also treated surgically. In the previously reported cases of multiple spinal DAVFs, the second fistulas were also diagnosed after the initial treatment. Symptomatic patients after the initial treatment of DAVF should be re-examined angiographically. PMID- 15662567 TI - Simultaneous multi-section perfusion CT and CT angiography for the assessment of acute ischemic stroke. AB - BACKGROUND: Introduction of helical computed tomography (CT) scanning has enabled rapid imaging of the vascular status by means of CT angiography and perfusion CT. By virtue of recent multi-detector technology, helical CT has the ability to perform both CT angiography and multi-section perfusion CT simultaneously. This study investigated the clinical feasibility of simultaneous assessment of perfusion CT and CT angiography in patients with acute ischemic stroke. METHOD: Perfusion CT and CT angiography were performed simultaneously in a series of consecutive 31 acute ischemic stroke patients. The time required for the entire processing was about 15 minutes. Contrast agent was used in a total dose of 100 ml (35 ml for perfusion CT and 65 ml for CT angiography). FINDINGS: Simultaneous perfusion CT scans and CT angiographies were of diagnostic quality for 29 patients (94%). In large territorial infarct patients, perfusion CT could predict all perfusion deficits of the final lesions (10 out of 10 lesions) and CT angiography could detect 9 of 10 occlusions of major cerebral arteries (90%). In patients with small lacunar or subcortical infarcts, perfusion CT could predict 9 out of 19 lesions (47.4%), and false-negative were encountered in small lesions (three patients) or in inadequate coverage of data acquisition (seven patients). Acute stage thrombolytic intervention could be carried out based on the findings, and the success of thrombolytic therapy could be demonstrated by follow-up study. CONCLUSIONS: Simultaneous perfusion CT and CT angiography is the very useful tool for the rapid and adequate diagnosis of almost all of the large territorial infarcts and some of non-territorial lacunar infarcts. It is an easy-to-perform and safe imaging technique to assess acute ischemic stroke. PMID- 15662569 TI - Cranio-cervical stabilization of traumatic atlanto-occipital dislocation with minimal resultant neurological deficit. AB - Our purpose is to describe a case of atlanto-occipital dislocation and discuss treatment approaches to minimize subsequent neurological deficits. Traumatic atlanto-occipital dislocation, has traditionally been considered rare and lethal, due to resulting high levels of spinal cord injury. Outcomes are generally expected to be poor. However, recent case reports indicate that survival is increasing. Of patients who survive cranio-cervical dislocation, many endure resulting neurological deficits. We present a rare case of a 23-year-old male, who sustained an atlanto-occipital dislocation in a motor vehicle accident. The patient presented with a Glasgow Coma Scale (GCS) of 11T. Lateral C-spine x-ray and thin-section slices CT delineated a C1 ring fracture on the left side with approximately 1 cm anterior and superior subluxation of the occipital condyles of the cranium in reference to C1. The patient was completely awake, alert, and was following commands. The patient underwent a cranio-cervical stabilization from occiput to C3, using lateral mas screws (C1-C3) and transarticular screws (C2 C3). The Vertex (Medtronics) system used included longitudinal bars connected to the lateral mas plating system, which was subsequently used to place screws within the keel of the occipital bone. Motor strength and sensation remained intact following surgery. One-week post-operation, the patient was ambulating 140 feet, conversationally appropriate, and had a GCS of 15. This case illustrates the possibility for neurosurgical intervention of cranio-cervical dislocations to achieve optimal outcome and demonstrates that survival from this injury is not only conceivable, but recovery of function is also possible. PMID- 15662570 TI - Superficial siderosis of the central nervous system following cervical nerve root avulsion: the importance of early diagnosis and surgery. AB - Superficial siderosis (SS) of the central nervous system is an insidious, progressive, irreversible and debilitating neurological disorder caused by recurrent haemorrhage within the subarachnoid space. The subsequent deposition of haemorrhagic breakdown products in the spinal cord and nervous tissues leads to the loss of neurones and myelin, and to the development of a neurological deficit. In a small number of patients, the source of haemorrhage is related to traumatic cervical nerve root avulsion occurring several years prior to the onset of symptoms. Surgical ablation of the source has been shown to halt the progression of the disease, at least in the short term. We review the literature on SS secondary to cervical nerve root avulsion and report a further case in which surgical management was successful in halting disease progression. We emphasize that early detection and recognition of the initial non-progressive symptoms related to this poorly known disease, coupled with timely surgical management, minimizes the degree of neurological disability. PMID- 15662571 TI - Thrombosed basilar apex aneurysm presenting as a third ventricular mass and hydrocephalus. AB - Aneurysms presenting as third ventricular masses are uncommon; most are giant aneurysms arising from the basilar apex. We present a case of a thrombosed basilar apex aneurysm presenting as a third ventricular mass and hydrocephalus in a 55-year-old man. The case is unique in the literature as the aneurysm was completely thrombosed and angiographically occult. The lesion was explored to verify the diagnosis and a third ventriculostomy resolved the patient's symptoms. Completely thrombosed aneurysms should be considered in the differential diagnosis of symptomatic third ventricular masses, even when angiographic studies are negative. The literature on aneurysms presenting as third ventricular masses is reviewed. PMID- 15662572 TI - Intramedullary spinal cord germinoma expresses the protooncogene c-kit. AB - A 33-year-old woman first noticed numbness in the both lower extremities and the numbness gradually extended up to the thorax. Magnetic resonance (MR) imaging demonstrated a mass with moderate and heterogeneous enhancement and peritumoural intramedullary cysts in the spinal cord at the T-1 to T-3 levels. The tumour was explored partially and intra-operative pathological examination demonstrated the so-called "two-cell pattern" typical of germinoma. The margin of the tumour was not clear. Histological examination showed germinoma with strong c-kit protein expression on the tumour cell surface. Chemotherapy and radiation therapy were given following surgery. Her neurological deficits were improved but not resolved. PMID- 15662573 TI - External radiotherapy and anaemia treatment: state of the art. AB - Anaemia is considered a common problem in many cancers secondary to the disease itself or related to chemo- and/or radiotherapy. Several clinical trials have advocated the prognostic value of anaemia and hypoxia in the outcome of many cancers. Erythropoietin is recognised as an effective treatment for anaemia, which also improves the quality of life in patients with malignant disease. External radiotherapy plays an important role in the treatment of loco-regional cancer but its efficacy can be compromised by many factors. Tumor hypoxia is considered by many authors as an important factor contributing to radioresistance. We report in this article the radiobiological rationale in favour of combining radiotherapy and erythropoietin, and review relevant clinical papers published in this field. PMID- 15662575 TI - Self-reported health needs and use of primary health care services by adolescents enrolled in post-mandatory schools or vocational training programmes in Switzerland. AB - BACKGROUND: The second Swiss Multicenter Adolescent Survey on Health (SMASH02) was conducted among a representative sample (n = 7428) of students and apprentices aged 16 to 20 from the three language areas of Switzerland during the year 2002. This paper reports on health needs expressed by adolescents and their use of health care services over the 12 months preceding the survey. METHODS: Nineteen cantons representing 80% of the resident population agreed to participate. A complex iterative random cluster sample of 600 classes was drawn with classes as primary sampling unit. The participation rate was 97.7% for the classes and 99.8% for the youths in attendance. The self-administered questionnaire included 565 items. The median rate of item non-response was 1.8%. Ethical and legal requirements applying to surveys of adolescent populations were respected. RESULTS: Overall more than 90% of adolescents felt in good to excellent health. Suffering often or very often from different physical complaints or pain was also reported such as headache (boys: 15.9%, girls: 37.4%), stomach-ache (boys: 9.7%, girls: 30.0%), joint pain (boys: 24.7%, girls: 29.5%) or back pain (boys: 24.3%, girls: 34.7%). Many adolescents reported a need for help on psychosocial and lifestyle issues, such as stress (boys: 28.5%, girls: 47.7%) or depression (boys: 18.9%, girls: 34.4%). Although about 75% of adolescents reported having consulted a general practitioner and about one-third having seen another specialist, reported reasons for visits do not correspond to the expressed needs. Less than 10% of adolescents had visited a psychiatrist, a family planning centre or a social worker. CONCLUSIONS: The reported rates of health services utilisation by adolescents does not match the substantial reported needs for help in various areas. This may indicate that the corresponding problems are not adequately detected and/or addressed by professionals from the health and social sectors. PMID- 15662576 TI - Junior physicians' workplace experiences in clinical fields in German-speaking Switzerland. AB - BACKGROUND AND OBJECTIVES: To date, there have been several prospective cohort studies investigating the workplace experiences of junior physicians, but with limited focus on gender issues. The objective of the present study is to explore the workplace experiences of first-year residents according to gender, type of training hospital, and clinical field. METHODS: Data reported are from the second assessment of the longitudinal Swiss physicians' career development study, begun in 2001. In 2003, 497 residents (54.7% females, 45.3% males) assessed their workplace conditions, social support at work, and effort-reward imbalance. RESULTS: There are few, but relevant, gender related differences in workplace experiences, with female physicians experiencing less mentoring and higher over commitment, yet more positive social relationships at work. In a multivariate model, significant differences in some workplace variables with regard to type of training hospital and/or clinical field are found: workplace conditions are rated worse in type "A" hospitals (university and cantonal hospitals) than in type "B"/"C"/"D" hospitals (regional hospitals and highly specialised units), and in surgical fields than in internal medicine. In "A" hospitals mentoring is assessed as better, but positive social relationships as worse. Both scales are rated worse in surgical fields than in internal medicine. The effort-reward imbalance (ERI) is rated significantly higher (unfavourable) in "A" hospitals than in "B"/"C"/"D" hospitals, regardless of gender and clinical field. Significantly more subjects with an ERI quotient above 1 (which is unfavourable) work in "A" hospitals, and in surgical fields regardless of hospital type. Of the total sample, 81 subjects (16.3%), 41 males and 40 females, show an ERI quotient above 1. The greater the workload, the worse the rating of workplace conditions, effort reward imbalance, and over-commitment. CONCLUSION: Institutional determinants are crucial factors for the workplace experiences and first career steps of junior physicians. Medical educators, especially those in "A" hospitals, should become more involved in structured residency programs and be aware of potential gender inequalities in the career support of female physicians. PMID- 15662577 TI - Body mass index and the risk of male cancer mortality of various sites: 17-year follow-up of the Basel cohort study. AB - QUESTIONS UNDER STUDY: There is growing evidence for a link between body weight and cancer risk, but there is not a clear consensus yet. METHODS: We studied the association between body mass index (BMI) and overall, lung, prostate and colon cancer mortality. In 1971/73, weight and height were measured in 2974 men working in Basel, Switzerland. In 1990, the vital status of all participants was assessed. RESULTS: 290 men had died from cancer, 87 from lung, 30 from prostate, and 22 from colon cancer. In the predefined Cox Proportional Hazards Regression Models for survival analysis, a baseline hazard was modified multiplicatively by covariates, i.e. the untransformed continuous variable "BMI" was chosen as covariate. In addition it was assumed that the baseline hazard may be different for smokers, non-smokers and different age groups (age at entry into study). Thus, multiple strata, i.e. combinations of smoking status and age groups were allowed. With increasing BMI overall cancer mortality did not change. Accordingly, the relative risk (RR) per 1-unit increase of BMI (unit = 1 kg/m2) was 1.03 (95% CI: 0.99-1.07). In relation to lung cancer, mortality did neither increase nor decrease with increasing BMI (RR = 1.0; 95% CI 0.93- 1.07). The results for prostate cancer mortality were similar, i.e. no correlation with BMI was observed (RR = 0.95; 95% CI: 0.93-1.18). The same was true for colon cancer mortality (RR = 1.09; 95% CI: 0.92-1.24). CONCLUSIONS: This investigation provides little evidence of an association between BMI and mortality of all cancers combined, cancer of the lung, the prostate and the colon. PMID- 15662578 TI - Docetaxel-induced radiation recall dermatitis. PMID- 15662579 TI - [Prevention of diabetes mellitus type 1]. AB - Primary prevention of type 1 diabetes is as yet not feasible. In contrast, prevention of acute and chronic complications of type 1 diabetes is highly effective. With a near normal blood glucose control the appearance and progression of microvascular complications (i.e. retinopathy, nephropathy and neuropathy) can be significantly reduced. Intensive insulin therapy is the gold standard for treatment of type 1 diabetes. PMID- 15662580 TI - [Malaria in the emergency room. Results of the emergency treatment of 137 patients with symptomatic malaria]. AB - OBJECTIVE: To assess characteristics and outcome of emergency patients with acute malaria. PATIENTS AND METHODS: We retrospectively assessed the clinical and laboratory parameters of 137 consecutive patients (87 males, 50 females; median age 37 years, range 17 - 67 years) presenting with acute malaria to our tertiary care center between 1992 and 2002. RESULTS: Falciparum malaria was diagnosed in 116/137 and tertian malaria in 19/137 patients; a single patient was infected with both parasites while in another case the type of parasite remained unclear. Infections were acquired in Africa (121), Asia , and in the Americas . One traveler visited multiple continents. Only 36 % (50/137) of patients had used malaria chemoprophylaxis. 128/137 patients were treated as in-patients; 22 of these had to be treated on an intensive care unit. According to the criteria of the German Society of Tropical Medicine, 44/137 (32 %; 95 % confidence interval (CI): 25 - 40 %) patients suffered from complicated malaria. The overall mortality rate was 2/137 (1.5 %; 95 % CI: 0,4 - 5.2 %); the mortality rate of complicated malaria tropica was 2/44 (4,5 %; 95 % CI 1,3 - 15 %). Patients with complicated malaria were significantly older than those with uncomplicated malaria. Median length of hospital stay was 4 days in uncomplicated and 9 days in complicated cases. Based on costs of EUR 2500 per case, an attack rate of > 3 % in East African travelers and a cost of EUR 55 for a chemoprophylaxis with mefloquine, chemoprophylaxis is cost-effective. CONCLUSION: In our retrospective analysis, complicated malaria tropica was associated with older age. Although malaria causes considerable morbidity, the overall mortality from severe malaria is low. Reinforcement of chemoprophylaxis especially in travelers to Africa could reduce malaria cases and is cost-effective. PMID- 15662581 TI - [Angioedema in hereditary deficiency of complement factor 1 esterase inhibitor and alpha 1-antitrypsin]. AB - HISTORY AND CLINICAL FINDINGS: A 15-year-old girl had suffered from episodic, sometimes threatening angioedema of the face, nasopharyngeal space and distal extremities beginning at age 13. EXAMINATIONS: A C1-esterase inhibitor (C1-INH) deficiency was revealed protein-chemically and functionally. There was also an alpha (1)-antitrypsin (AAT) deficiency with heterocygotic phenotype PiMZ. The combination of C1-INH and AAT deficiency was also found in the patient's mother and brother. THERAPY AND COURSE: Under 8-month therapy with 200 mg/d danazol per os (reduction of the dosis in the last month to 100 mg/d), there was no further edema, the C1-INH concentration normalized and there was also an increase in C1 INH function. During the observation period, use of the emergency set with C1-INH concentrate was not required. CONCLUSIONS: This is the first reported case of angioedema in combination of two hereditary enzyme defects C1-INH deficiency (autosomal-dominant genetics) and AAT deficiency (autosomal-recessive). In addition to a survey of current literature, the current state of diagnostics and therapy of the hereditary angioedema is presented. PMID- 15662582 TI - [Ambulatory management of hepatitis B before and after liver transplantation]. AB - HISTORY AND CLINICAL FINDINGS: A 48-year-old patient from Pakistan was referred with scleral icterus, ascites and lower limb edema. DIAGNOSIS: The patient was suffering from post-hepatitis liver cirrhosis, CHILD stage B, with underlying chronic hepatitis B and C coinfection. TREATMENT AND COURSE: One year after diagnosis, orthotopic liver transplantation (OLT) was performed. To avoid recurrence of the hepatitis B in the transplant, the patient was given a treatment with lamivudine preoperatively. A prophylaxis with a combination of lamivudine and hepatitis B immunoglobulin peri- and postoperatively has been given for more than 5 years. CONCLUSION: The use of lamivudine and hepatitis B immunoglobulin improves transplant survival in patients who have undergone OLT on the basis of post-hepatitis liver cirrhosis secondary to chronic hepatitis B. Costs are reduced by administering the hepatitis B immunoglobulin intramuscularly and by managing the preoperative and aftercare of the patient on an outpatient basis. PMID- 15662583 TI - [Tuberculosis--current therapeutic concepts]. PMID- 15662584 TI - [Secondary osteoporosis in endocrine diseases]. PMID- 15662585 TI - [Dehydration. Ethical standards finding in the social controversy]. AB - Three case histories are presented, in which terminal hydration was omitted. Two patients were in the terminal phase of a menigiosis carcinomatosa and Creutzfeldt Jacob disease, the third was completely aphasic and hemiplegic after ischemic stroke. All three died peacefully without signs of suffering or thirst. The psychological aspects of therapeutic-palliative change and the archetypical horror of death are considered. The moral justification is discussed in relation to the "Principles for Physicians Accompanying the Dying" of the German chamber of physicians. The respect of the patient's will is the guiding principle. These ethical controversies should be discussed sincerely and frankly so that the society is enabled to follow. PMID- 15662586 TI - [Hypothenar-hammer syndrome as the cause of a severe digital ischemia. Re the article in DMW 44/2004]. PMID- 15662587 TI - [Complicated course of Churg-Strauss syndrome with eosinophilic perimyocarditis and pericardial tamponade. From DMW Nr. 41/2004]. PMID- 15662588 TI - Frequent promoter methylation of tumor-related genes in sporadic and men2 associated pheochromocytomas. AB - Hypermethylation of CpG island promoters is associated with transcriptional inactivation of tumor suppressor genes in neoplasia. Inactivation of p16 and Pten was related to the development of pheochromocytomas. In this report, we investigated the methylation status of the p16INK4a cell cycle inhibitor gene and other prominent tumor-related genes ( PTEN, RASSF1 A, CDH1, MSH2, MLH1, VHL, and TIMP3) in sporadic and multiple endocrine neoplasia type 2 (MEN2) pheochromocytomas by methylation-specific PCR. Hypermethylation was detected in 48 % of pheochromocytomas for RASSF1 A, 24 % for p16, 36 % for MSH2, 16 % for CDH1, and 8 % for PTEN. No VHL, MLH1, and TIMP3 methylation was observed. Interestingly, the frequency of p16 inactivation in familial tumors was higher (5 out of 12, 42 %) than in sporadic tumors (1 out of 13, 8 %; p = 0.047) and RASSF1 A inactivation was more common in the hereditary tumors (58 %) compared to the sporadic tumors (38 %). Combined methylation of RASSF1 A and p16 was found only in MEN2-related pheochromocytomas. Thus, a subset of hereditary pheochromocytomas displays preferential methylation of p16 and RASSF1 A. PMID- 15662589 TI - Pilot study: tendency of increasing iodine content in human milk and cow's milk. AB - The iodine supply in Germany has improved throughout the last decade, albeit with enormous differences between individuals and regions. In the Thuringian city of Jena, analyses of the iodine content of human milk have been undertaken regularly since 1982. Significantly increasing iodine concentrations in human and cow's milk have been found. Therefore, the current situation and the effectiveness of measures to prevent iodine deficiency demands re-evaluation. The iodine content of human milk from 32 lactating mothers was analysed on the 5th day (mean) postpartum and mothers' dietary iodine intake during the last two months of pregnancy was assessed by means of a food frequency questionnaire. To corroborate the assumption that the increasing iodine levels of cow's milk are one of the main reasons for the improved iodine supply, the iodine concentration of 34 cow's milk bulk-samples was also determined. Both human and cow's milk samples were analysed by the ICP-MS method. Twenty women took iodine supplements (mean daily intake = 175 microg). The average daily iodine intake of the 20 supplemented and 12 non-supplemented women was 258 microg and 116 microg, respectively. Daily iodine intake from food and beverages was significantly lower in supplemented women (83 microg/day). The average iodine content of human milk was 169 +/- 88 microg/l with a range of 33 - 348 microg/l. This content is two times higher than levels from 1994 in the same area. There was no difference in the human milk iodine content between mothers taking supplements and those who did not. Cow's milk samples showed a mean iodine concentration of 178 +/- 131 microg/l (range 48 - 661 microg/l). PMID- 15662590 TI - Functional thyrotropin receptor expression in the pituitary folliculo-stellate cell line TtT/GF. AB - Thyrotropin secretion from the anterior pituitary is regulated mainly through TRH and thyroid hormones. Recent findings of a TSH receptor (TSHR) on folliculo stellate (FS) cells in the human anterior pituitary indicate that TSH secretion might, in addition, be regulated in a paracrine manner via FS cells. In order to elucidate the physiological relevance of TSHR expression in FS cells we evaluated the effects of TSH on a murine FS cell line, TtT/GF. First, Western blot analysis confirmed the expression of TSHR protein in these cells. Second, three potential second messenger pathways were studied. Last, cDNA array hybridization was used to evaluate the effect of TSH on gene expression levels. TSH failed to induce either the adenylate cyclase/cAMP pathway, the phosphatidylinositol/calcium pathway, or the Janus kinase (JAK)/signal transducer and activator of transcription (STAT) 3 pathway. Most of the genes regulated by TSH were related to cell proliferation, cell differentiation, and apoptosis. Moreover, TSH induced STAT5a and TGFbeta2 expression. We report that TtT/GF cells express a functional TSHR that is not coupled to cAMP nor IP (3) but probably signals through the JAK/STAT5a pathway. Functional TSHR expression in this cell line offers an in vitro model to study the role of TSHR in FS cells. PMID- 15662591 TI - Signal transduction mediating gene expression of SP1, LHbeta-subunit and GH in response to GnRH or GHRH in the postnatal and fetal porcine anterior pituitary in vitro. AB - To clarify signal transduction pathways mediating putative gene expression of transcription factor SP1 (selective promoter factor 1 or specificity protein 1) by GnRH or GHRH porcine anterior pituitary monolayer cultures were exposed for various time periods to GnRH, GHRH, activators of adenylate cyclase (AC) or proteinkinase C (PKC), and mRNA levels of SP1, LHbeta-subunit, and GH were determined by multiplex RT-PCR. In many experiments LH and GH release were measured as well for comparison. Another approach was to inactivate AC, PKC, or proteinkinase A (PKA) by specific inhibitors, MDL, GFX, and H89, respectively. Postnatally (4 weeks) SP1 mRNA level was maximally increased by GnRH, GHRH and both by activation of AC or PKC after 2 h of exposure. Two-hour stimulation of SP1 mRNA levels by dbcAMP was totally blocked by H89, while this inhibitor not clearly blocked GHRH stimulated SP1 mRNA levels. Stimulation of LHbeta mRNA by GnRH was suppressed by inactivation of AC or of PKC but not by inactivation of PKA. Inactivation of AC or PKA but not of PKC inhibited GHRH induced GH mRNA. Already at day 50 of fetal life (and likewise day 80) SP1 mRNA levels were stimulated by GHRH or activation of AC, but not by GnRH or activation of PKC. The results are consistent with the notion that SP1 plays an important role 1) in conferring GnRH responsiveness to the LHbeta-subunit gene by mediating the actions of both AC and PKC and 2) in conferring GHRH responsiveness to the GH gene through activation of the AC probably PKA pathway. Furthermore, the data are in line with the view that the GHRH/AC/SP1/GH pathway develops earlier during fetal life than the GnRH/PKC/SP1/LHbeta pathway. PMID- 15662592 TI - Identification of a novel calcium-sensing receptor gene mutation causing familial hypocalciuric hypercalcemia by single-strand conformation polymorphism analysis. AB - Calcium-sensing receptor gene (CASR) mutations that alter the function of the G protein coupled Ca (2+)-sensing receptor are reported in patients with familial hypocalciuric hypercalcemia (FHH), autosomal dominant hypocalcemia (ADH), and neonatal severe hyperparathyroidism (NSHPT). In search for novel disease causing mutations in the CASR gene, we screened exons 2 - 7 of the CASR gene of a family with FHH using single-strand conformation polymorphism analysis. We identified a novel CASR mutation (c.518 T > C; L173 P) in exon 4 encoding for the extracellular domain of the Ca (2+)-sensing receptor. This region seems to represent a hot spot within the CASR gene with at least 13 reported disease causing mutations thus far. PMID- 15662593 TI - Levels of plasma fibrinogen and d-dimer in patients with impaired fasting glucose. AB - BACKGROUND: Cardiovascular risk associated with impaired fasting glucose has been examined in various studies with conflicting results. During the last 10 years, several risk markers for atherosclerosis such as fibrinogen and D-dimer have been identified. The present study was designed to evaluate plasma fibrinogen and D dimer levels in patients with impaired fasting glucose compared with normal subjects and those with type 2 diabetes mellitus. METHODS: Age-, sex-, and body mass index-matched 30 normal subjects, 30 patients with impaired fasting glucose (fasting glucose 110 to 125 mg/dl), and 30 patients with type 2 diabetes mellitus (fasting glucose >/= 126 mg/dl) were included in the study. RESULTS: The levels of plasma fibrinogen in patients with type 2 diabetes mellitus, impaired fasting glucose, and normal subjects were 449 (306 - 605) mg/dl, 348 (264 - 468) mg/dl, and 216 (179 - 260) mg/dl, respectively. Patients with impaired fasting glucose had significantly lower plasma fibrinogen levels than patients with type 2 diabetes mellitus (p < 0.05). There were significantly higher plasma fibrinogen levels in patients with impaired fasting glucose than in normal subjects (p < 0.05). The levels of plasma D-dimer in patients with type 2 diabetes mellitus, impaired fasting glucose, and normal subjects were 615 (505 - 768) mg/l, 518 (412 - 664) mg/l, and 424 (356 - 557) mg/l, respectively. Patients with impaired fasting glucose had significantly lower plasma D-dimer levels than patients with type 2 diabetes mellitus (p < 0.05). There were significantly higher plasma D dimer levels in patients with impaired fasting glucose than in normal subjects (p < 0.05). The levels of plasma fibrinogen and D-dimer were related to fasting glucose in type 2 diabetes mellitus and impaired fasting glucose groups (p < 0.05). We also detected positive correlation between plasma fibrinogen levels and age in study groups (p < 0.05). CONCLUSION: Our data suggest that patients with impaired fasting glucose pose a hypofibrinolytic status and cardiovascular risk, although this was lower than in patients with type 2 diabetes mellitus. PMID- 15662594 TI - Gastric electrical stimulation results in improved metabolic control in diabetic patients suffering from gastroparesis. AB - AIMS/HYPOTHESIS: Symptoms of gastroparesis possess a heavy impact on the quality of life; delayed gastric emptying may result in poor metabolic control in diabetics. Gastric electrical stimulation (GES) has recently been introduced as a treatment option in patients with drug refractory gastroparesis to increase the quality of life by alleviating nausea and vomiting frequencies. However, the effect of GES on metabolic control has not been assessed yet. METHODS: We performed a prospective single center study on the long-term effect (12 months) of continuous high-frequency/low-energy GES on symptoms, gastric emptying (measured scintigraphically), and metabolic control (HbA1c) in insulin-dependent diabetic subjects suffering from drug-refractory gastroparesis for more than one year. RESULTS: Seventeen (12 female, 5 male) patients entered the study; all were available for analysis at all time points. No therapy-associated adverse events occurred. Weekly vomiting and nausea frequencies decreased significantly at 6 and 12 months. Gastric retention rates improved significantly from 83 % (2 h) and 38 % (4 h) to 35 % (2 h)/14 % (4 h) and 25 % (2 h)/17 % (4 h) at 6 and 12 months, respectively. HbA1c values were lowered in all 17 subjects; initially, all HbA1c values were above 7.5 %; at 6 and 12 months, mean values had significantly decreased from 8.6 % to 6.2 % and 6.5 %, respectively. CONCLUSIONS/INTERPRETATION: Gastric electrical stimulation offers symptom control in diabetics with drug-refractory gastroparesis and decreases gastric retention. This study, for the first time, documents a positive effect of this therapy on metabolic control as indicated by HbA1c, a surrogate marker of the risk of diabetic complications. PMID- 15662595 TI - Differential regulation of human blood glucose level by interleukin-2 and -6. AB - While the acute phase reaction to infection is associated with hyperglycemia, during progressing infection hypoglycemia can develop. The cytokines regulating the dynamics of host defense may concurrently contribute to blood glucose regulation. To examine this hypothesis, changes in blood glucose concentrations in healthy men were compared following administration of interleukin-2 (IL-2) and IL-6 representing, respectively, major mediators of the adaptive and the early innate immune response to bacterial infection. Doses of 10 000 IU/kg IL-2 and 0.5 microg/kg IL-6 (vs. placebo) were administered subcutaneously in two groups of men (n = 18 and 16) at 1900 h before a period of nocturnal rest allowing an assessment of changes under basal conditions. Serum concentrations of glucose and of various hormones were assessed every 60 min. Despite generally lowered glucose concentration at night, IL-2 induced a transient but distinct decrease in blood glucose concentration most consistent 8 - 9 hours following injection (p < 0.01). The hypoglycemic response to IL-2 was not accompanied by changes in serum insulin, C-peptide or cortisol. In contrast to IL-2, IL-6 led to an increase in cortisol, followed by a pronounced increase in blood glucose again peaking about 8 hours after injection (p < 0.001). Results indicate a differential regulation of blood glucose concentration by cytokines. Contrasting with the hyperglycemic effects of the acute phase regulator IL-6, the T-cell cytokine IL-2 seems to support glucose uptake and utilization by immune cells. PMID- 15662596 TI - Effect of pioglitazone on lipids in well controlled patients with diabetes mellitus type 2 -- results of a pilot study. AB - INTRODUCTION: Patients with diabetes mellitus type 2 are characterized by a typical dyslipoproteinemia. Improvement in glucose control usually also ameliorates this dyslipoproteinemia. It is unclear whether different antidiabetic strategies differ in their effects on the lipid profile. Particularly, it is unknown whether glitazones improve lipid values independently of their effects on glucose metabolism. METHODS: Ten patients well controlled on sulfonylureas (HbA1c 6.9 +/- 0.5 %) with diabetic dyslipoproteinemia were treated with additional pioglitazone (30 mg/d) for 3 months. Every 4 weeks the sulfonylurea dose was adjusted to keep HbA1c and fasting glucose constant. Before and after 3 months of pioglitazone therapy lipid metabolism was determined in detail (cholesterol, triglyceride, LDL-cholesterol, HDL-cholesterol, VLDL-cholesterol, VLDL triglycerides, lipoprotein(a), LDL-subtype distribution by isopycnic density gradient ultracentrifugation). RESULTS: Although glucose control remained unchanged (HbA1c 6.9 +/- 0.5 % vs. 6.8 +/- 0.6 %; fasting glucose concentration 7.7 +/- 1.1 vs. 7.3 +/- 1.3 mmol/l) we observed a significant reduction in triglyceride concentration (1.9 +/- 0.6 vs. 1.4 +/- 0.5 mmol/l, - 26 %, p < 0.01), a significant increase in HDL-cholesterol concentration (1.2 +/- 0.2 vs. 1.4 +/- 0.2 mmol/l, + 14 %, p < 0.05), a significant decrease in LDL/HDL-ratio (3.03 +/- 0.77 vs. 2.51 +/- 0.61, - 24 %, p < 0.05) and non-significant improvements in total cholesterol, LDL-cholesterol, VLDL-triglycerides, and VLDL cholesterol concentrations. The LDL-subtype profile improved (significant reduction [- 20 %] in small dense LDL). CONCLUSIONS: This pilot study indicates that at comparable fasting glucose concentration and at comparable HbA1c value pioglitazone is superior to sulfonylureas concerning the improvement of diabetic dyslipoproteinemia. Whether this relates to indirect effects (improvement in insulin sensitivity) or direct effects (stimulation of PPARalpha) remains to be determined. PMID- 15662597 TI - Moderate exercise with a dietary vitamin C and e combination protects against streptozotocin-induced oxidative damage to the kidney and lens in pregnant rats. AB - Moderate exercise and vitamin C and E (VCE) supplementation can be beneficial to diabetes due to reducing free radical production in lens and kidney of diabetic pregnant rats. We investigated the effect of VCE supplementation and moderate exercise on lipid peroxidation (MDA) and scavenging enzyme activity in the kidneys and lens of STZ-induced diabetic pregnant rats. Fifty female Wistar rats were used and were randomly divided into five groups. First and second were used as the control and pregnant control group. Third group was the pregnant diabetic group. The fourth group was the diabetic-pregnant-exercise group. VCE supplemented feed was given to pregnant-diabetic-exercise rats constituting the fifth group. Animals in the exercised groups were moderately exercised daily on a treadmill (16.1 m/min, 45 min/d) for three weeks (five days a week). Diabetes was induced on day zero of the study. Plasma, lens, and kidney samples were taken from all animals on day 20. Exercise and administration of VCE to pregnant diabetic rats resulted in significant decrease in the albumin and total protein values and the elevated MDA, plasma creatinine, and urea levels as an indicator of oxidative stress and renal functional parameters. Exercise and VCE supplementation also increased glutathione peroxidase (GSH-Px), reduced glutathione (GSH), vitamin E, and beta-carotene levels in the kidney, GSH-Px and GSH in the lens, the albumin and total protein values in plasma. In the diabetic pregnant animals, the decreased vitamins A and E concentration and GSH levels in kidney, creatinine, and urea values in plasma did not improve through exercise only although their concentrations were increased by VCE supplementation. Kidney weight did not also affect either by exercise or VCE supplementation. In conclusion, these results suggest that exercise plus VCE affects antioxidant metabolism and reduces lipid peroxidation, thereby improving the damage caused by oxidative stress involved in the pathogenesis of lens and kidney in diabetic pregnant rats. Moderate exercise with dietary VCE may play a role in preventing nephropathy and cataract formation in diabetic pregnant rat. PMID- 15662598 TI - Schmidt syndrome presenting as acute liver failure. AB - The Schmidt Syndrome (Type II Autoimmune-Syndrome) is characterised by an autoimmune adrenalitis in combination with a chronic lymphocellular thyreoiditis resulting in insufficiency of these organs in adulthood. Combination with diabetes is possible. The diagnosis is usually established by clinical examination and analysis of serum hormone levels (adrenocorticotropin hormone [ACTH], cortisol, thyroid stimulating hormone [TSH], triiodothyronine [fT3], thyroxine [fT4]). In the present case, initial diagnosis was rapid progressive liver failure of unknown origin with consecutive multiple organ dysfunction syndrome including dysfunction of heart, lungs, and kidneys. Frequent and less frequent causes of liver failure were ruled out, e.g. viral or autoimmune hepatitis, Budd-Chiari-syndrome, toxic, or drug induced liver failure. In retrospect, the multiple organ dysfunction syndrome was caused by hypoperfusion due to severe hypovolemia and hypoperfusion was induced by adrenocortical insufficiency proven by endocrinological testing. The clinical course of this case stresses the importance of the hormone balance in the critical ill patient. The guideline for treatment of patients with assumed hormonal dysregulation should include a full hormone status prior to substitution. The present case report also illustrates the importance of clinical signs and careful consideration of the medical history in detecting an autoimmune endocrine disease. PMID- 15662601 TI - Molecular markers in bladder cancer. AB - Bladder cancer is one of the malignancies for which extensive information regarding molecular pathogenesis and genetic predictors of natural history as well as response to various modalities of treatment based on molecular profile is available. As more prognostic markers are being investigated in clinical trial settings, in the not very distant future we will be able to use these predictive markers in clinical decision-making. Bladder cancer is the second most common genitourinary tumor and is a significant cause of morbidity and mortality. A need for tumor markers that can be incorporated into clinical practice to add prognostic information and to refine the conventional TNM and grading systems in terms of treatment response and prognosis is crucial. Intravesical and systemic chemotherapy in bladder cancer are limited in their efficacy in the treatment of bladder cancer patients primarily when they are unable to induce apoptosis in bladder tumor cells. Understanding the apoptotic signals and the cascade of reactions that give pro-survival signals will go a long way in refining the treatments and will help in the future to individualize cancer therapies. It is imperative to study the role of these mechanisms in prospective clinical trials in a quest to find predictive markers that can help to tailor treatments, keeping in view the molecular heterogeneity. PMID- 15662599 TI - Functional consequences of PRODH missense mutations. AB - PRODH maps to 22q11 in the region deleted in the velocardiofacial syndrome/DiGeorge syndrome (VCFS/DGS) and encodes proline oxidase (POX), a mitochondrial inner-membrane enzyme that catalyzes the first step in the proline degradation pathway. At least 16 PRODH missense mutations have been identified in studies of type I hyperprolinemia (HPI) and schizophrenia, 10 of which are present at polymorphic frequencies. The functional consequences of these missense mutations have been inferred by evolutionary conservation, but none have been tested directly. Here, we report the effects of these mutations on POX activity. We find that four alleles (R185Q, L289M, A455S, and A472T) result in mild (<30%), six (Q19P, A167V, R185W, D426N, V427M, and R431H) in moderate (30%-70%), and five (P406L, L441P, R453C, T466M, and Q521E) in severe (>70%) reduction in POX activity, whereas one (Q521R) increases POX activity. The POX encoded by one severe allele (T466M) shows in vitro responsiveness to high cofactor (flavin adenine dinucleotide) concentrations. Although there is limited information on plasma proline levels in individuals of known PRODH genotype, extant data suggest that severe hyperprolinemia (>800 microM) occurs in individuals with large deletions and/or PRODH missense mutations with the most-severe effect on function (L441P and R453C), whereas modest hyperprolinemia (300-500 microM) is associated with PRODH alleles with a moderate reduction in activity. Interestingly, three of the four alleles associated with or found in schizophrenia (V427M, L441P, and R453C) resulted in severe reduction of POX activity and hyperprolinemia. These observations plus the high degree of polymorphism at the PRODH locus are consistent with the hypothesis that reduction in POX function is a risk factor for schizophrenia. PMID- 15662602 TI - The surgical management of muscle invasive bladder cancer: a contemporary review. AB - Muscle-invasive bladder cancer is a potentially lethal disease with a high rate of cure if timely and effective therapy is applied while the disease is confined to the bladder or regional lymph nodes. Radical cystectomy is the gold standard to which all other local therapies including multimodality bladder-preserving strategies should be compared. Contemporary cystectomy combined with regional lymphadectomy may be performed with an acceptably low morbidity, provides unparalleled local control, and may result in durable disease-free survival even among patients with locoregional lymph node metastases. Refinements in surgical technique coupled with the expanded application of continent urinary diversion have resulted in excellent functional outcomes without compromising cancer control in properly selected patients. An increasing awareness of the importance of quality-of-life issues combined with an enhanced understanding of tumor biology have resulted in the surgical modifications which include an expanded role for lymphadectomy and preservation of uninvolved adjacent organs. PMID- 15662603 TI - Management of muscle invasive bladder cancer--British approaches to organ conservation. AB - British groups have long favored a more conservative approach to the management of invasive bladder cancer. They have been involved in several randomized trials looking at the efficacy of these approaches and have also been leaders in the study of neoadjuvant chemotherapy. This article documents the concerns that linger over the efficacy and morbidity of cisplatin-based chemotherapy and describes a national trial investigating a potentially less morbid 5FU and mitomycin alternative. PMID- 15662604 TI - Organ preservation by combined modality treatment in bladder cancer: the European perspective. AB - Combined modality therapy, including transurethral resection, radiation therapy, and systemic chemotherapy, has been shown to produce survival rates comparable to those of radical cystectomy. With these programs, cystectomy has been reserved for patients with incomplete response or local relapse after trimodality treatment. This review summarizes European series of combined modality treatment of muscle-invasive bladder cancer and reflects our emphasis on the importance of combining radiation with a transurethral resection and cisplatin-based chemotherapy. It also documents our belief that high-grade T1 disease may be effectively treated by the same approach. This represents a true distinction between the European and US strategies. We also review the role of predictive and prognostic factors in selecting patients for the respective treatment alternatives. PMID- 15662605 TI - Selective bladder preservation by trimodality therapy for patients with muscularis propria-invasive bladder cancer and who are cystectomy candidates--the Massachusetts General Hospital and Radiation Therapy Oncology Group experiences. AB - The Massachusetts General Hospital and the Radiation Therapy Oncology Group have been leading the charge for organ conservation in bladder cancer in North America for over two decades. In a series of six successive studies the group has refined the techniques and is now moving toward a translational future in which novel biologic agents will be combined with the best current strategies. The North American approach is characterized by its selective nature, in that it preselects patients likely to do well with a trimodality approach and then further selects according to the response to an induction course of chemotherapy and radiation. Only those who are complete responders move onto full dose. This "check point" allows salvage cystectomy to be performed on incomplete responders before they have had full-dose radiation. This preserves the urinary diversion options open to the surgeon as well as brings forward the time to a salvage procedure should it be needed. PMID- 15662606 TI - Radiation therapy for muscle-invasive bladder cancer: treatment planning and delivery in the 21st century. AB - Advances in radiation treatment planning, verification, and delivery provide a means to optimize radiation treatment for bladder cancer and overcome difficulties which have previously limited the success of this treatment. They offer the opportunity to enhance the therapeutic ratio by reducing the volume of normal tissue irradiated and by increasing radiation dose or using more intensive fractionation and synchronous chemotherapy regimes. In this review, we aim to present a practical overview of radiation protocols for bladder cancer, covering issues relating to patient selection, choice of target volumes, verification, dose, and fractionation. Alternative methods of improving treatment accuracy such as image-guided radiotherapy and intensity modulated radiotherapy are also discussed. PMID- 15662607 TI - Brachytherapy in the treatment of invasive bladder cancer. PMID- 15662608 TI - Quality of life after radical treatment for invasive bladder cancer. AB - Considerable effort has been put into improving the quality of life after radical therapy for bladder cancer, though it has proved difficult to demonstrate conclusively that these aims have been achieved. There can be little doubt that the impact of a cystectomy is substantial but that it can be lessened by the use of continent and orthotopic diversion. Quality-of-life studies have, however, shown the remarkable ability of patients to adapt well to the more commonly used incontinent ileal conduit. Chemoradiation appears to have little impact on bladder function for about three-quarters of all patients and this effect will likely be lessened in the future by the use of partial bladder irradiation or better targeting using fiducial markers. Both radical surgery and chemoradiation can perturb bowel function but this may be reduced by choosing less functionally critical portions of bowel for diversion or by better targeted pelvic nodal radiation. Male sexual function is profoundly affected by both treatment approaches although conformal radiation or nerve-sparing cystectomy may help in the future. Female sexual function has never been fully examined but the impact of both approaches is likely very high. PMID- 15662609 TI - Chemotherapy for local treatment of bladder cancer. AB - This review documents the evidence that combination chemotherapy, known to be useful in metastatic disease, offers an advantage in the context of apparently localized disease. The use of neoadjuvant chemotherapy before cystectomy shows that pathologic complete responses can be obtained one-third of the time and that this, in combination with an aggressive TURBT, can be one strategy to preserve the bladder. The response to chemotherapy strongly predicts ultimate survival whatever the local therapy. Two recently updated randomized trials raise the possibility that neoadjuvant chemotherapy has a small but detectable survival benefit yet its use is controversial in everyday practice because of the desire of surgeons to perform an immediate cystectomy. The role of adjuvant chemotherapy remains undefined. PMID- 15662610 TI - von Willebrand factor: a prima ballerina on two different stages. PMID- 15662611 TI - New perspectives on von Willebrand factor functions in hemostasis and thrombosis. AB - The adhesive protein, von Willebrand factor (VWF), mediates the initiation and progression of thrombus formation at sites of vascular injury by means of specific interactions with extracellular matrix components and platelet receptors. The essential biologic properties of VWF have been elucidated, with progress particularly in the areas of genetic regulation, biosynthesis, and specific bimolecular interactions. The three-dimensional structure of selected domains has been solved, but our understanding of detailed structure-function relationships is still fragmented, partly because of the complexity and size of the VWF molecule. The biomechanical properties of the interaction between the VWF A1 domain and the platelet receptor glycoprotein (GP) Ibalpha also are better known, but we can still only hypothesize how this adhesive bond can oppose the fluid dynamic effects of rapidly flowing blood to initiate thrombus formation and contribute to platelet activation. Elucidating the details of VWF and GPIbalpha function will lead to a more satisfactory definition of the role of platelets in atherothrombosis, since hemodynamic forces greatly influence responses to vascular injury in stenosed and partially occluded arteries. Progress in this direction is also aided by rapidly expanding novel information on the mechanisms that regulate VWF multimer size in the circulation, a topic of relevance to explain microvascular thrombosis and, perhaps, arterial thrombosis in general. Developments in these areas of research will refine our understanding of the role played by VWF in vascular biology and pathology. PMID- 15662612 TI - Phenotypic and genotypic diagnosis of von Willebrand disease: a 2004 update. AB - In the last two decades, progress in the diagnosis of von Willebrand disease (VWD) came from the rapidly developing field of molecular techniques that allowed the first phenotype-genotype correlations. In particular, structural and functional defects of von Willebrand factor (VWF) that underlie VWD type 2 and their molecular basis not only helped to understand the pathophysiology of VWD but also the complex post-translation processing of VWF and the multiple VWF functions. In contrast to the dramatic development of molecular techniques, improvement of methods for phenotypic description, a prerequisite for phenotype genotype comparisons, has been neglected. The gold standard to differentiate VWD type 2 from type 1 and between diverse type 2 subtypes is the electrophoretic analysis of VWF multimers, a demanding technique that itself is not easily standardized but of crucial relevance for correct classification. This article summarizes the current knowledge on phenotype-genotype correlations as well as up to-date phenotypic and genotypic methods in the diagnosis of VWD. PMID- 15662613 TI - Treatment of von Willebrand disease. AB - The bleeding tendency in von Willebrand disease (VWD) is heterogeneous and some patients with the mildest form of the disease have no significant bleeding symptoms throughout their lives. In some cases, the most difficult task for a clinician is to decide whether any treatment is actually required. However, cases with moderate to severe factor VIII (FVIII) and von Willebrand factor (VWF) deficiency usually require treatment to stop or prevent bleeding. Increasing autologous FVIII/VWF by desmopressin administration or providing normal allogeneic VWF through the infusion of plasma-derived concentrates can correct FVIII and VWF deficiencies and normalize or shorten bleeding time (BT). FVIII levels are the best predictors of soft tissue or surgical bleeding, while BT normalization, reflecting the correction of platelet-dependent functions of VWF, is considered a reliable indicator of an effective treatment of mucosal bleeding. Recombinant concentrates of FVIII are not indicated (apart from cases with alloantibodies against exogenous VWF), since they are devoid of VWF and lack its stabilizing effect on circulating FVIII. A very-high-purity concentrate of VWF has recently been made available, but its advantages over conventional concentrates containing both FVIII and VWF moieties are not obvious. The best way to select the appropriate treatment is to perform a test infusion with desmopressin in any patient with clinically significant VWD, provided that he/she has no contraindication to the compound or belongs to subtype with an anticipated lack of response (for example, type 3 VWD with FVIII/VWF lower than 5%). PMID- 15662614 TI - von Willebrand disease in the developing world. AB - von Willebrand disease (VWD) is considered to be the most common inherited bleeding disorder. Data on its epidemiology and impact in developing countries are limited. The biologic heterogeneity and variable presentation of VWD make diagnosis difficult. Although there is no accurate estimate of the prevalence of VWD in developing countries, available data suggest that the proportion of diagnosed cases is lower than the expected number, often accounting for only 6% to 13% of patients with hereditary bleeding disorders. Although accurate subtyping is often not possible, the number with severe disease tends to be much higher, particularly in those parts of the world where consanguinity is common. Agents used to treat patients with VWD range from plasma to purified factor concentrates. Desmopressin (DDAVP) is commonly used. Preliminary data on molecular genetics suggests that there are significant population differences. There is inadequate awareness of this condition and lack of support for these patients from the health care system in many developing countries. Concerted efforts are needed at the scientific and social levels to improve this situation. PMID- 15662615 TI - von Willebrand disease and women's health. AB - In 1926 von Willebrand described a bleeder family in Aland; this condition became known as von Willebrand disease (VWD). von Willebrand noted that "the trait seemed especially to be seen among the women." Today, the use of a pictorial bleeding assessment chart (PBAC) has enabled the prevalence of VWD to be established among women presenting with menorrhagia, as well as the documentation of this symptom in women with known VWD and the assessment of treatment response in menorrhagia. Treatments for menorrhagia include tranexamic acid, desmopressin (DDAVP) administered either intranasally or subcutaneously, the oral contraceptive pill, the "Mirena" coil (Schering Oy, Turku, Finland), and endometrial ablation. Von Willebrand factor (VWF) shows strong cyclical variation, with peak values occurring in the luteal phase. Although increased in pregnancy, levels of VWF decline postnatally and the incidence of both primary and secondary postpartum hemorrhage is high (20% to 25%). Baseline VWF levels less than 15 IU/dL are unlikely to reach greater than 50 IU/dL in the third trimester, and therefore prophylaxis with DDAVP or VWF-containing concentrate to cover delivery should be considered. PMID- 15662616 TI - von Willebrand factor and factor VIII as risk factors for arterial and venous thrombosis. AB - Since the early 1990s attempts have been made to elucidate whether high concentrations of von Willebrand factor (VWF) and factor VIII (FVIII) in plasma are associated with an increased risk of thrombosis. Several prospective studies on the role of VWF in arterial thrombosis, mainly coronary heart disease, were performed in healthy individuals and patients with previous cardiovascular disease. Although the majority showed an association between high VWF levels and arterial thrombosis, others failed to confirm such findings. A smaller number of studies have evaluated FVIII, mainly for its association with venous thrombosis. Two prospective observations, together with several case-control studies, provided solid evidence of an association between high FVIII levels and a first or recurrent episode of venous thrombosis. On the whole, high levels of VWF and FVIII in plasma confer a moderately high risk of arterial and venous thrombosis, respectively. These findings have no therapeutic implication, but they should be taken into account in the assessment of the individual risk profile. PMID- 15662617 TI - Interplay between ADAMTS13 and von Willebrand factor in inherited and acquired thrombotic microangiopathies. AB - The presence of unusually large multimers of von Willebrand factor (VWF) is thought to be a major pathogenic factor for thrombotic thrombocytopenic purpura (TTP). ADAMTS13 is a protease that regulates the multimeric size and function of VWF by cleaving VWF. Hence, congenital or acquired deficiency of ADAMTS13 causes life-threatening illness of TTP. Mutations in the ADAMTS13 gene cause inherited TTP, and the development of autoantibodies that inhibit ADAMTS13 activity frequently are associated with acquired TTP. ADAMTS13 consists of 1,427 amino acid residues and is composed of multiple structural and functional domains, containing a signal peptide, a propeptide, a reprolysin-like metalloprotease domain, a disintegrin-like domain, a thrombospondin type-1 (Tsp1) motif, a cysteine-rich domain, a spacer domain, seven additional Tsp1 repeats, and two CUB domains. In particular, the cysteine-rich/spacer domains are essential for VWF cleavage and are the principal epitopes recognized by autoantibodies in patients with acquired TTP. Therefore, it is likely that these domains are involved in the recognition and binding of ADAMTS13 to VWF. ADAMTS13 circulates in the blood in an active state, and efficiently cleaves unfold form of VWF induced under shear stress caused by blood flow, preventing the accumulation of pathogenic unusually large VWF multimers (ULVWF). Thus, ADAMTS13 helps maintain vascular homeostasis by preventing the excess thrombus formation. PMID- 15662618 TI - New approach to quantifying developmental variation in the dentition using serial microtomographic imaging. AB - Dental morphogenesis and cellular differentiation are expressed in the fully formed tooth by the topography of the dentin-enamel junction and outer enamel surface. These boundaries can be differentiated using a variety of imaging systems. In this study, we used serial microCT imaging to provide accurate 3D reconstructions of developing lower human second deciduous molars. These were used to quantify the volume of enamel and dentin of individual cusps in relation to basal crown height. As growth and differentiation proceed apically, the spatial orientation of cusp tips and their bases were used to estimate their order of initiation and coalescence. We found that the order of coalescence differed from the order of initiation. We also found that dentin cusp height and volume as well as rate and quantity of enamel apposition varied along mesio distal and bucco-lingual axes, and were independent of order of initiation and duration of growth. These results demonstrate that the potential for variation in crown size and form is maintained throughout development. We propose that the microCT model developed in this study constitutes a new approach for the investigation of developmental variation and its contribution to phylogenetic variation expressed in crown form and size. PMID- 15662619 TI - Determination of the distribution of consolidants and interpretation of mercury porosimetry data in a sandstone porous network using LSCM. AB - Knowing the 3D distribution of a consolidant within the porous network of a rock is essential for understanding the porosity quantitative data obtained by mercury porosimetry and for observing the effect of consolidants on pore interconnection. In this work, we show for the first time that the distribution of consolidant in the porous network can be determined using laser scanner confocal microscopy (LSCM). Results indicate that consolidants are concentrated in pore throats of less than 40 microm in diameter, affecting both the porous interconnection and the circulation of fluids. LSCM allowed demonstration of the fact that the increase in microporosity detected by mercury porosimetry is due to the development of fissures within the consolidants. No consolidant that produces this kind of fissure can be used in the consolidation of building stones, since it would increase microporosity and, in consequence, vulnerability to weathering agents. PMID- 15662620 TI - GFAP immunoreactivity and transcription in trigeminal and dental tissues of rats and transgenic GFP/GFAP mice. AB - Sensory mechanisms in teeth are not well understood and may involve pulpal-neural interactions. Tooth cells that proliferate in vitro have polyclonal immunoreactivity (IR) for glial fibrillary acidic protein (GFAP), growth associated protein (GAP-43), and vimentin, plus glial-like ion channels. Here, we analyzed GFAP-IR patterns in dental and trigeminal tissues of rats, for comparison with green fluorescent protein (GFP) associated with GFAP transcription in transgenic mice, in order to better characterize glial-like cells in dental tissues. Astrocytes, ganglion satellite cells, and epineurial Schwann cells were demonstrated by anti-GFAP antibodies and GFP-GFAP, as expected. Odontoblasts did not stain by any of these methods and cannot be the glial-like cells. Fibroblasts and undifferentiated mesenchymal cells in pulp had polyclonal GFAP-IR and vimentin-IR, while nerve fibers reacted only with polyclonal antibody. Some Schwann cell subtypes in trigeminal nerve and oral mucosa were positive for GFP and for polyclonal anti-GFAP, but not for monoclonal antibody. In pulp almost all Schwann cells were unstained, but many Schwann cells in periodontal ligament had polyclonal GFAP-IR. These results show greater heterogeneity for Schwann cells than expected, and suggest that the glial-like pulp cells are fibroblasts and/or undifferentiated mesenchymal cells or stem cells. We also found that polyclonal GFAP revealed intermediate filaments in preterminal sensory nerve fibers, thereby providing a useful marker for that neural subregion. GFP transcription by some Schwann cell subtypes in oral mucosae and trigeminal nerve, but not trigeminal root was a novel finding that reveals more complexity in peripheral glia than previously recognized. PMID- 15662621 TI - Effects of objective numerical apertures on achievable imaging depths in multiphoton microscopy. AB - Multiphoton microscopy is a powerful technique for achieving three-dimensional submicron imaging in biological specimens. However, specimen optical parameters such as refractive indices and scattering coefficients can result in the loss of image resolution and decreased signal in depth. These factors are coupled to the focusing objective's numerical aperture (NA) in limiting the achievable imaging depths. In this work, we performed multiphoton imaging on aqueous fluorescent solution, human skin, and rat tail tendon to show that, under the same immersion condition, lower NA objectives can examine more deeply into biological specimens and should be used when optimal imaging depths is desired. PMID- 15662622 TI - Technique to quantify local clustering of synaptic vesicles using single section data. AB - Synaptic vesicles are organelles that specialize in the storage of a neurotransmitter that continuously undergo an exo-endocytotic cycle. During this cycle vesicles change their positions within a presynaptic terminal and their numbers as well as spatial arrangement can provide insight into a neurotransmitter turnover. This article introduces a technique based on the nearest-neighbor formalism to quantify the proximity of vesicles to active zones and vesicle clustering in different regions of a terminal. The technique, implemented in a software package, uses the two-dimensional coordinates of features identified in digitized electron micrographs as an input. It has been validated in the analysis of asymmetric synapses of the rat hippocampal CA1 stratum radiatum affected by transient cerebral ischemia. It was shown that a 15 minute-long ischemic episode influenced the spatial arrangement of vesicles that were more distant from active zones and had larger intervesicle spacings with respect to the control. The latter effect was apparently stronger within 200 nm distance of active zones. PMID- 15662623 TI - Method for localization of sialic acid on cell surface and cell interior in peripheral blood: a pilot study. AB - This article presents a method for identification and localization of cell surface and intracellular sialoglycoconjugates of peripheral blood cells. To reveal cell surface conjugates, a sample of peripheral blood was incubated with lectin after centrifugation and rinsing. For intracellular localization in leukocytes, RBCs were lysed and the membranes were permeabilized prior to cytochemical reaction. Fluorescein isothiocyanate conjugated lectins were used for visualization in fluorescence microscope. All lectins bound specifically to the surface of erythrocytes. Confocal microscopy showed surface and intracellular labeling of permeabilized leukocytes. A part of the signal in eosinophils originated from binding of anionic fluorophore to cationic granular proteins. PMID- 15662624 TI - High-power laser vaporization of the canine prostate using a 110 W Thulium fiber laser at 1.91 microm. AB - INTRODUCTION: The Thulium fiber laser may have several advantages over current urology lasers, including smaller size, more efficient operation, improved spatial beam quality, more precise tissue incision, and operation in pulsed or continuous-wave modes. Previous laser-tissue interaction studies utilizing the Thulium fiber laser have been limited to laser powers of less than 5 W, restricting potential medical applications. This study describes the preliminary testing of a high-power Thulium fiber laser for vaporization of the canine prostate, ex vivo. STUDY DESIGN/MATERIALS AND METHODS: A continuous-wave, 110-W Thulium fiber laser operating at a wavelength of 1.91 microm, delivered 88.5+/ 2.3 W of power through a 600-microm-core silica fiber for non-contact vaporization of canine prostates (n = 6). RESULTS: The Thulium fiber laser vaporized prostate tissue at a rate of 0.83+/-0.11 g/minute. The thermal coagulation zone measured 500-2,000 microm, demonstrating the potential for hemostasis. CONCLUSIONS: The high-power Thulium fiber laser is capable of rapid vaporization and coagulation of the prostate. In vivo animal studies are currently being planned for evaluation of the Thulium fiber laser for potential treatment of benign prostate hyperplasia (BPH). PMID- 15662625 TI - Immunochemical detection of Lonomia obliqua caterpillar venom in rats. AB - Severe cases of human envenoming by caterpillars of the saturniid moth Lonomia obliqua in Brazil can result in renal damage, leading to renal failure, and intracerebral hemorrhaging. In this work, we used immunohistochemical staining with rabbit antiserum raised against L. obliqua venom to examine venom distribution in selected tissues of the brain (cerebellum and hippocampus), kidneys, and liver of male Wistar rats injected with a single dose of venom (200 microg/kg, i.v.) and sacrificed 6, 18, 24, and 72 hours later. The immunolabeling of GFAP was also examined to assess the venom effects on perivascular astrocytic end-feet in the microvasculature of the hippocampus and cerebellum. Venom was detected in the kidneys (6 and 18 hours) and in the liver (6 hours) but not in the brain at any of the time intervals examined. In contrast, immunolabeling for GFAP revealed astrogliosis in the cerebellum and enhanced expression of this protein in the glial processes of the cerebellum and hippocampus, with a maximum response from 24 hours onwards. The high immunoreactivity seen in the kidneys agreed with the renal damage and dysfunction reported for some patients. The lack of venom detection in the brain, despite the altered expression of GFAP in astrocytes, suggested either that the venom does not enter this organ or that its entrance is transient and fast. Alternatively, the circulating venom may induce the release of mediators that could serve as second messengers to provoke the late astrocytic reactivity and astrogliosis. It is possible that both of these mechanisms may contribute to the effects observed. PMID- 15662626 TI - Analysis of hyperpigmentation and hypopigmentation after Er:YAG laser skin resurfacing. AB - BACKGROUND AND OBJECTIVES: Pigmentary disorders--such as hyperpigmentation and hypopigmentation, are devastating complications of erbium:yttrium-aluminum-garnet (Er:YAG) laser resurfacing. This study was undertaken to assess the clinical and histopathologic features of hyperpigmentation and hypopigmentation following Er:YAG laser resurfacing, especially in darker skin. STUDY DESIGN/MATERIALS AND METHODS: One hundred and ninety patients (skin phototypes III and above), treated with Er:YAG lasers--short-pulsed and modulated (variable-pulsed and dual-mode) Er:YAG lasers--for skin resurfacing were recruited. The clinical features of hyperpigmentation and hypopigmentation were evaluated retrospectively using medical charts and serial photographs. For histopathologic examinations, skin biopsies were performed in three patients at hyperpigmentation sites and in four patients at hypopigmentation sites. RESULTS: Hyperpigmentation was observed in 38.4% of the patients. Mean onset and duration were 3.5 and 7.2 weeks, and then it has faded away within 16 weeks in 93.2% of cases. Hypopigmentation was observed in 13.7% of the patients; its mean onset was 2 months after treatment, and it faded within 1 year in 85% of cases. The incidences and mean durations of these side effects were more intense and longer in patients treated with short pulsed, variable-pulsed, and dual-mode Er:YAG lasers, in increasing order. In terms of histopathologic examinations, melanin amounts in the epidermal basal layer were observed to vary. CONCLUSIONS: Hyperpigmentation and hypopigmentation are frequent complications of Er:YAG laser resurfacing. Long pulse duration induced thermal damage seems to be the most important factor in terms of the induction of pigmentary disorders. PMID- 15662627 TI - Suppressed TGF-beta1 expression is correlated with up-regulation of matrix metalloproteinase-13 in keloid regression after flashlamp pulsed-dye laser treatment. AB - BACKGROUND AND OBJECTIVES: Flashlamp pulsed-dye lasers (PDLs) has shown effectiveness in the treatment of keloids. In this study, we investigated whether PDL treatments decreased transforming growth factor-beta1 (TGF-beta1)-induction and up-regulation of matrix metalloproteinase (MMP) expression in keloid regression. STUDY DESIGN/MATERIALS AND METHODS: Keloid tissues obtained from 10 patients with intra-lesional or punch biopsies before and 7 days after PDL treatments [fluence per pulse was 10-18 J/cm2 (mean 14.0 J/cm2)]. Immunohistochemical (IHC) staining of TGF-beta1 and MMP-1 and MMP-13 expressions in keloid tissue was performed. Western blot analysis of MMP-1 and MMP-13 expressions in extracellular matrix was evaluated. RESULTS: IHC staining indicated that expression of TGF-beta1 was significantly reduced in keloid tissues after PDL irradiation. MMP-13 but not MMP-1 expression on IHC staining significantly increased in extracellular matrix of keloid tissues after PDL treatment. Western blot analysis also showed MMP-13 but not MMP-1 significant increased in keloid tissues after PDL treatment. CONCLUSIONS: Regression of keloids regressed after PDL treatments are associated with down-regulation of TGF beta1 expression and up-regulation of MMP-13 activity. PMID- 15662628 TI - Influence of physiological conditions on laser damage thresholds for blood, heart, and liver cells. AB - BACKGROUND AND OBJECTIVES: Damage to blood and other tissues during laser interventions depends mainly upon absorption of laser radiation by cells. The objective of this work was to evaluate the influence tissue-specific physiological factors on photo-damage thresholds of individual cells: Red blood cells (blood), hepatocytes (liver), and miocytes (heart). STUDY DESIGN/MATERIALS AND METHODS: Laser-induced damage to individual cells was detected and studied with Laser Load Test (LLT). Probability and thresholds of RBC damage after one laser pulse (532 nm, 10 nanoseconds) were obtained experimentally as functions of physiological conditions. Using in vitro models, we have studied influence of the oxygen level, pH, temperature, and cell heterogeneity on RBC, the inhibition of metabolic activity on miocytes and drug toxicity on hepatocytes. RESULTS: Single laser pulse induced cell lyses through a vapor bubble. The decrease of the O2 level and temperature caused increase of damage thresholds at 532 nm. Deviation of the pH level from neutral to any side caused also the increase of the damage threshold. Inhibition of metabolism of miocytes and toxic damage to hepatocytes also resulted in the increase of the damage threshold. CONCLUSIONS: Resistance of various tissues at cell level against photo-damage significantly depends on physiological properties of cells. A general rule for such dependence is that the better the cell state the lower its threshold for laser-damage. PMID- 15662629 TI - Spectral evaluation of laser-induced cell damage with photothermal microscopy. AB - BACKGROUND AND OBJECTIVES: Determining cell photo-damage is important for laser medicine and laser safety standards. This work evaluated the potential of photothermal (PT) technique for studying invasive laser-cell interaction, with a focus on PT evaluation of spectral dependence of laser-induced damage in visible region at single intact cell level. STUDY DESIGN/MATERIALS AND METHODS: PT is based on irradiation of a single intact cells with a tunable pump laser pulse (420-570 nm, 8 nanoseconds, 0.1-300 microJ) and monitoring of temperature dependent variations of the refractive index with a second, collinear probe beam in pulse (imaging) mode (639 nm, 13 nanoseconds, 10 nJ), and continuous (integrated PT response) mode (633 nm, 2 mW). The local and the integrated PT responses from the individual living red blood cells, lymphocytes, and cancer cells (K562) in vitro were obtained at different pump laser fluence and wavelength and compared with data obtained by conventional viability tests (Annexin V--propidium iodide). RESULTS: The cell damage with pump pulse lead to specific change in PT response's temporal shape and PT image's structure. The photodamage thresholds varied in the range of 0.5-5 J/cm2 for red blood cells, 4.4-42 J/cm2 for lymphocytes, and 36-90 J/cm2 for blast cells in the pump wavelength range of 417-555 nm. CONCLUSION: Damage threshold at different wavelength depends on absorption spectra of cells. Spectral evaluation of laser damage thresholds can be done in two supplements for each PT mode--PT imaging and integrated PT response. The correlation between specific change of PT parameters and cell damage permits using PT technique to rapidly estimate the invasive conditions of the laser-cell interactions. PMID- 15662630 TI - Evaluation of a KTP (potassium-titanyl-phosphate) 532 nm laser for endovaporization of choroidal melanomas. AB - BACKGROUND AND OBJECTIVES: The purpose of this in vitro study is to evaluate the use of the KTP (potassium-titanyl-phosphate) 532 nm laser for future use in endovaporization of large choroidal melanomas. STUDY DESIGN/MATERIALS AND METHODS: A KTP 532 nm green laser coupled to a 0.3 mm fiberoptic probe was used to perform in vitro studies on human cadaver eyes and on one enucleated melanoma eye. The specimens were examined by histological staining (Masson-Trichrome) and scanning electron microscopy (SEM). RESULTS: Histological analysis of sclera, choroid, retina, and melanoma revealed different zones of collateral thermal tissue damage. These zones were analyzed quantitatively. We determined the best suitable laser parameters and the time for sclera penetration. CONCLUSIONS: The KTP-laser showed good potential in intraocular vaporization of choroidal melanomas. However, great differences in laser absorption between sclera and melanoma can cause significant complications such as perforation. PMID- 15662631 TI - Effect of wavelength on low-intensity laser irradiation-stimulated cell proliferation in vitro. AB - BACKGROUND AND OBJECTIVES: There exist contradictory reports about low-intensity laser light-stimulated cell proliferation. The purpose of this study was to determine the effect of wavelength on proliferation of cultured murine cells. STUDY DESIGN/MATERIALS AND METHODS: Proliferation of primary cell cultures was measured after irradiation with varying laser wavelengths. RESULTS: Fibroblasts proliferated faster than endothelial cells in response to laser irradiation. Maximum cell proliferation occurred with 665 and 675 nm light, whereas 810 nm light was inhibitory to fibroblasts. CONCLUSIONS: These observations suggest that both wavelength and cell type influence the cell proliferation response to low intensity laser irradiation. PMID- 15662633 TI - Histologic evaluation of the pulsed Nd:YAG laser for laser lipolysis. AB - BACKGROUND AND OBJECTIVES: Laser lipoplasty with pulsed Nd:YAG laser, widely used in Europe and Latin America, has recently been introduced in Japan and the USA. We report histologic analyses of the effects of the laser on human fat tissue. STUDY DESIGN/MATERIALS AND METHODS: Freshly excised human skin and subcutaneous fat were irradiated with the pulsed Nd:YAG laser (SmartLipo, DEKA, Italy). A 1,064 nm laser at 40 Hz and 150 mJ and 100 microseconds-long pulses were used. Methods of exposure were the same as in the clinical application. In the control group, the specimens were cannulated by the handpiece without irradiation. The tissue was studied by scanning electron microscopy and hematoxylin eosin staining. RESULTS: Scanning electron microscopy after irradiation showed greater destruction of human adipocytes than in the control. Degenerated cell membrane, vaporization, liquefaction, carbonization, and heat-coagulated collagen fibers were observed. CONCLUSIONS: Our study showed that the SmartLipo appeared to be histologically effective for destruction of human fat tissue. PMID- 15662632 TI - Activation of ERK and p38 kinase mediated keloid fibroblast apoptosis after flashlamp pulsed-dye laser treatment. AB - BACKGROUND AND OBJECTIVES: Flashlamp pulsed-dye lasers (PDLs) revealed effective regression or arrest in patients with keloids in our clinical studies [Kuo YR et al., Laser Surg Med 2004;34:104-108]. In this study, we further investigated whether the induction of keloid regression seen with PDL treatment through activation in mitogen-activated protein (MAP) kinase and caspase promotes cell apoptosis and reduces fibroblast proliferation. STUDY DESIGN/MATERIALS AND METHODS: Keloid tissues were obtained from 10 patients with intralesional or punch biopsies prior to and 7 days after PDL treatments [fluence per pulse was 10 18 J/cm2 (mean 14 J/cm2)]. Prior to and after PDL treatments, the proliferating fibroblasts in keloid tissue were immunohistochemically detected by proliferating cell nuclear antigen (PCNA) expression. The apoptotic cell was detected by terminal deoxynucleotidyl transferase dUTP-nick end labeling (TUNEL) staining and fragmented caspase-3 expression. MAP kinase activation as represented by extracellular signal-regulated kinase (ERK), p38 kinase (p38), and c-Jun N terminal kinase (JNK) expression of keloid tissues was investigated by immunohistochemical (IHC) staining, respectively. RESULTS: IHC staining indicated that PCNA expression of fibroblasts was significantly reduced in keloid tissue after PDL irradiation. TUNEL assay revealed lower apoptotic cells expression in the keloid tissue prior to laser treatment. Following laser treatment, apoptotic cells with relatively strong DNA damage and fragmentation were seen in all keloid biopsy samples, especially in the keloid fibroblast population. The activation of ERK and p38 MAP kinase increased significantly in keloid tissue after PDL treatment. JNK was shown to be unchanged. CONCLUSIONS: The PDL treatment is shown to induce keloid regression through suppression of keloid fibroblast proliferation, induction of apoptosis, and upregulation of ERK and p38 MAP kinase activity. PMID- 15662634 TI - Deflection of an eye shield by the cryogen cooling spray causing a cryogen induced conjunctivitis in a patient receiving treatment with the pulsed dye laser. PMID- 15662635 TI - Measuring dynamics of caspase-3 activity in living cells using FRET technique during apoptosis induced by high fluence low-power laser irradiation. AB - BACKGROUND AND OBJECTIVES: Low-power laser irradiation (LPLI) has been used for therapies such as curing spinal cord injury, healing wound etc. Yet, the mechanism of LPLI remains unclear. In order to determine the effects of high fluence LPLI on cell growth and caspase-3 activity, we have measured the dynamics of caspase-3 activity during cell apoptosis induced by high fluence LPLI treatment. STUDY DESIGN/MATERIALS AND METHODS: He-Ne laser was used to irradiate human lung adenocarcinoma cells (ASTC-a-1). Cell Counting Kit-8 was used for cytotoxicity assay. A fluorescent microscope was used to perform fluorescence resonance energy transfer (FRET) imaging. A luminescence spectrometer was used to acquire the fluorescent emission spectrum. Statistical analysis was performed with Student's paired t-test. RESULTS: Cytotoxicity assay showed that when light irradiation fluence exceeded 60 J/cm2, LPLI treatment induced ASTC-a-1 cell apoptosis in a fluence-dependent manner. FRET imaging and spectrofluorometric analysis demonstrated that caspase-3 was activated during high fluence LPLI induced cell apoptosis. CONCLUSIONS: Using FRET technique, we have reported that high fluence LPLI can induce human lung adenocarcinoma cells (ASTC-a-1) apoptosis. The activation of caspase-3 plays an important role in the apoptotic process. PMID- 15662636 TI - QTL mapping with discordant and concordant sibling pairs: new statistics and new design strategies. AB - The term "extreme discordant and concordant" (EDAC) sampling has been used to describe a variety of strategies for quantitative trait locus mapping using sibling pairs sampled from the corners of the bivariate trait distribution. The principle of the design is to gain efficiency by genotyping only the most informative of the available sibling pairs. EDAC-type designs have been studied in a number of papers, and have been applied in a few others. This literature is somewhat out of date, however, because there are many new statistics that are appropriate for EDAC data. With newer statistics, the power of EDAC designs can be improved. Moreover, the relative power of different designs must be re evaluated, because the newer statistics improve the power of some designs more than others. That is, there is a circular relationship between design and statistic choices. In this report, we review a number of available design and statistic choices for EDAC studies, and use simulation to show what statistics are most powerful for each design. We then use those more powerful statistics to suggest strategies for making design choices among various EDAC and non-EDAC designs that use sibling pairs. We find that when genotyping must be minimized, an EDAC design with predominantly discordant pairs is the best choice, and when a balance of genotyping and phenotyping effort must be achieved, single proband ascertainment can do better. We also show that moderately selected samples (as opposed to very extreme samples) can be an efficient choice for many studies. PMID- 15662637 TI - Stress fracture of the distal tibia and fibula through heavy lifting. AB - BACKGROUND: Occupational stress fractures are reported among athletes, soldiers, ballet dancers, and in connection with shoveling. METHODS: A literature search on occupational stress fractures was performed. The biomechanical stress in association with special lifting tasks was estimated. RESULTS: A 59-year-old male welder presented with a stress fracture of the left distal tibia and fibula in connection with heavy lifting. The object was in front and lifted while he partly stood on one leg, which was rotated during the lifting. There was no considerable previous exposure to heavy lifting. CONCLUSION: It is likely, that this stress fracture was the result of strain on tibia and fibula through this composite lifting operation. PMID- 15662639 TI - Occupational exposure to metalworking fluids and risk of breast cancer among female autoworkers. AB - BACKGROUND: Metalworking fluids (MWF) are used for lubrication during metal manufacturing. Previous studies have observed increased risks of several cancers among MWF-exposed workers. We hypothesized that MWF may be associated with risk of breast cancer because they can contain carcinogenic or endocrine-disrupting chemicals. METHODS: We conducted a case-control study nested in a cohort of 4,680 female automobile workers employed for at least 3 years between 1/1/41 and 1/1/85, with follow-up through 1994. Cases were identified using the National Death Index (NDI), Michigan cancer registries, and company records. Detailed quantitative MWF exposure data were available for each subject, although data on known breast cancer risk factors were not. RESULTS: Ninety-nine cases of breast cancer and 626 matched controls were identified. There was a weak positive association between lifetime cumulative exposure to soluble MWF and breast cancer risk, but no evidence of association with either straight or synthetic fluids. When exposure was divided into time-windows, the association was strongest for soluble MWF in the decade preceding diagnosis. Controlling for earlier exposures, there was an odds ratio of 1.18 (95% CI=1.02-1.35) per mg/m3-year of cumulative exposure to soluble MWF in this 10-year period. CONCLUSION: This hypothesis generating study provides some preliminary evidence for an association between exposure to soluble MWF and increased risk of breast cancer. Additional studies of MWF and breast cancer, with data on known breast cancer risk factors, are warranted. PMID- 15662640 TI - Severe lead poisoning in the plastics industry: a report of three cases. AB - BACKGROUND: Lead stabilizers (e.g., lead sulfate, lead stearate) are common additives in plastics used in electrical devices. In 1997, three plastics compounders at one California company were severely lead-poisoned. METHODS: The poisonings were investigated by interviewing the workers, employer, and treating physician and reviewing medical records and environmental monitoring results. In addition to measuring blood lead levels (BLLs), noninvasive K X-ray fluorescence was used to measure bone lead concentration of the index case. RESULTS: Blood lead concentrations of the three workers at time of diagnosis were 159, 114, and 108 microg/dl. The worker with highest exposure presented with clinical findings of crampy abdominal pain, constipation, normocytic anemia, fatigue, and reversible azotemia. Bone lead concentration in his tibia, calcaneous, and patella were 102, 219, and 182 ppm, respectively. The poisonings resulted from uncontrolled use of powdered lead sulfate stabilizer. CONCLUSION: Clinicians should be aware of potential serious overexposure to lead in compounding of plastics. PMID- 15662641 TI - Determinants of work absence following surgery for carpal tunnel syndrome. AB - BACKGROUND: The objective was to identify factors across multiple domains associated with return to work in a community-based cohort of workers with carpal tunnel syndrome. METHODS: Workers scheduled for carpal tunnel release were recruited into this prospective study. Subjects completed questionnaires preoperatively and at 2, 6, and 12 months postoperatively. The questionnaires contained demographic, clinical, and psychosocial factors and physical and psychosocial workplace stressors. Predictors of work absence at 6 and 12 months were examined in bivariate and multivariate logistic regression analyses. RESULTS: Six months following surgery of 181 subjects, 29 (19%) were out of work. Twelve months postoperatively 33 subjects (22%) were out of work. In bivariate analyses, the factors associated with work absence at 6 months, at P < or = 0.01, included preoperative physical functional status, change in self-efficacy between preoperative assessment and 2 months, lower income, workers' compensation, representation by an attorney, work exposure to force and repetition, higher psychological job demands and lower control, lower social support by coworkers, lower job security and more supportive organizational policies and practices. The factors associated with work absence at 12 months in bivariate analyses included preoperative physical functional status, lower self-efficacy at 2 months, workers' compensation, and less supportive organizational policies and practices. Multivariate analyses documented a multidimensional model, with predictors from multiple domains. CONCLUSIONS: Clinical, demographic, economic, and workplace factors were associated with work absence. Strategies to reduce work absence following carpal tunnel release should address multiple dimensions of the worker and workplace. PMID- 15662642 TI - Change in permeation parameters and the decontamination efficacy of three chemical protective gloves after repeated exposures to solvents and thermal decontaminations. AB - BACKGROUND: Chemical protective clothing (CPC) and gloves, which provide adequate protection, are usually too expensive to be considered disposable. Repeated use of CPC without effective decontamination may result in secondary exposure and injury. However, decontamination may change the physical and/or chemical properties of the barrier material, causing variations in breakthrough time (BT) and steady-state permeation rate (SSPR). METHODS: Glove materials including neoprene, Guardian butyl rubber, and nitrile synthetic rubber were selected for this study. Toluene and acetone were chosen as the challenge chemicals. Permeation was measured in a closed loop system using a 2.5 cm permeation cell and a MIRAN infrared analyzer (Foxboro, MA). Following the permeation test, the samples were thermally decontaminated. After each exposure/decontamination cycle, BT and SSPR were measured. A total of 260 permeation tests were conducted. Permeation test results were collected on each material/chemical combination for up to 10 exposure/decontamination cycles. RESULTS: On average, changes in BT and SSPR in comparison with respect to new swatches were 11.5% and 13.7% after seven exposure/decontamination cycles. The percentages increased to 26.6% and 15.9% after 10 exposure/decontamination cycles, respectively. For at least seven cycles, the BT mean for four out of five material/chemical combinations tested (neoprene/acetone, neoprene/toluene, nitrile/acetone, and nitrile/toluene) was not significantly different from the original value of the BT for each corresponding swatch. Similarly, the SSPR mean for each of the five material/chemical combinations after at least five cycles was not significantly different from those for new swatches. The BT mean for the butyl/toluene combination, however, was significantly different from the new swatches even after the first exposure/decontamination. The SSPR mean was significantly different after five cycles compared to the new swatches. CONCLUSIONS: Except for the butyl/toluene combination, thermal decontamination was an effective method in removing the solvents from the matrix of selected glove materials. Multiple reuses of some chemical protective gloves could be safe if effective decontamination methods are used and the glove materials do not have significant degradation. PMID- 15662643 TI - Non-fatal occupational injuries related to slips, trips and falls in seafaring. AB - BACKGROUND: Merchant seafaring often involves hazardous occupational operations and several studies have shown increased overall injury mortality. The aim of this study was to describe the risks involved in slip, trip and fall (STF) injuries in merchant seafaring, in order to point out areas for prevention. METHODS: A questionnaire study was carried out in 11 countries with 6,461 participants. The seafarers gave information on whether they were injured during their latest tour of duty, and whether STF preceded the injury. RESULTS: Of the total reported injuries 43% (n=467) were STF related. Fractures and sprains accounted for 42% of the STF injuries compared with 17% for non-STF injuries. The proportions of STF injuries for different types of ships and areas of the ships varied from < 30% to >60%, particularly work on passenger ship decks, where STF injuries were >60%. The proportions of STF injuries increased by age and by severity of the injury. CONCLUSIONS: Injuries related to STF on merchant ships were more frequent than previous estimates. There is a need for improvements in defining, reporting, and preventing STF injuries in merchant seafaring. PMID- 15662644 TI - Certification of occupational diseases as common diseases in a primary health care setting. AB - BACKGROUND: It is often difficult to discern whether a disease is an occupational or common disease, especially in a primary care setting. METHODS: From a randomly selected sample of 322 workers attending a Primary Health Care Center, 207 workers (response rate of 64.3%) agreed to participate. An occupational questionnaire was administered. General practitioners provided medical records for each worker. Medical records and occupational questionnaires were independently reviewed by three professionals. They assessed whether a relationship between disease and working conditions was probable or improbable. RESULTS: Thirty-three of the 207 cases (15.9%) were considered probably related to working conditions according to the expert's opinion. The most frequent were musculoskeletal diseases (20 cases). Of the 207 workers, 74 (35.7%) judged that their diseases could be related to their working conditions. CONCLUSIONS: A significant proportion of diseases attended in primary care setting was not recognized as occupational, and they were hence not reflected in official statistics. PMID- 15662645 TI - Flock workers' exposures and respiratory symptoms in five plants. AB - BACKGROUND: Sentinel cases of lymphocytic bronchiolitis in flock production and coating operations triggered a five-plant study of airborne respirable dust and fiber exposures and health symptoms. METHODS: Job histories from 219 current workers were linked to a job-exposure matrix derived from personal exposure measurements of respirable dust and fibers. Univariate group comparisons and multivariate modeling tested for relations between indices of cumulative and current exposure, and respiratory and systemic symptom outcomes. RESULTS: Respiratory symptoms and repeated flu-like illnesses were associated with use of compressed air to clear equipment (blow-downs) and with respirable dust exposure (current and cumulative) after controlling for smoking. Blow-downs had an equal or greater effect than smoking status on most symptoms. CONCLUSIONS: Eliminating compressed air cleaning, engineering control of dust exposure, and respirators are needed to limit exposures to particulates. Longitudinal follow up may provide guidance for a dust or fiber level without adverse respiratory health effects. PMID- 15662646 TI - Outcomes in work-related injuries: a comparison of older and younger workers. AB - BACKGROUND: The "graying of the workforce" has generated concerns about the physical capacity of older workers to maintain their health and productivity on the job, especially after an injury occurs. There is little detailed research on age-related differences in work outcomes after an occupational injury. METHODS: A self-report survey about occupational, health, and financial outcomes, and related factors was administered 2-8 weeks post-injury to workers aged < 55 and > or = 55 who had lost time due to a work injury. RESULTS: Despite more severe injuries in older workers, most outcomes were similar in both age groups. In multivariate models, age was unrelated or inversely related to poor outcomes. Injury severity, physical functioning, and problems upon return to work were associated with adverse work injury outcomes. CONCLUSIONS: Older workers appear to fare better than younger workers after a work injury; their relative advantage may be primarily due to longer workplace attachment and the healthy worker effect. PMID- 15662647 TI - Occupational and environmental neurology: a pioneer. In memorium: Robert G. Feldman, MD, 1933-2003. PMID- 15662648 TI - Truck drivers and heart disease in the United States, 1979-1990. AB - BACKGROUND: Studies of truck drivers and cardiovascular disease (CVD), myocardial infarction, or ischemic heart disease (IHD) are limited, although studies of other professional drivers reported increased risk. METHODS: US mortality data from 1979 to 1990 for ages 15-90 were used to calculate proportional mortality ratios (PMRs) for heart disease and lung cancer for short and long haul truck drivers. Analysis was performed for Black (998 short haul and 13,241 long haul) truck drivers and White (4,929 short and 74,315 long haul) truck drivers separately. RESULTS: The highest significantly elevated proportionate heart disease (IHD, acute myocardial infarction (AMI), and other forms of heart disease) and lung cancer mortality was found for White and Black male long haul truck drivers age 15-54. Mortality was not significantly elevated for short haul truck drivers of either race or gender, nor for truck drivers who died after age 65, except for lung cancer among White males. An indirect adjustment suggested that smoking could explain the excess IHD mortality, but no direct data for smoking or the other known risk factors for heart disease were available and occupational exposures were not measured. CONCLUSIONS: The highest significant excess proportionate mortality for lung cancer, IHD and AMI was found for long haul truck drivers who were under age 55 at death. A cohort or longitudinal study of heart disease among long haul truck drivers, that obtains data for occupational exposures as well as lifestyle risk factors, could help explain inconsistencies between the findings of this and previous studies. PMID- 15662650 TI - Repellency and toxicity of aromatic plant extracts against the mosquito Culex pipiens molestus (Diptera: Culicidae). AB - The insecticidal activities of essential oil extracts from leaves, flowers and roots of aromatic plants against fourth-instar larvae of the mosquito Culex pipiens molestus Forskal were determined. Extracts of Foeniculum vulgare Mill were the most toxic, followed by those of Ferula hermonis Boiss, Citrus sinensis Osbeck, Pinus pinea L, Laurus nobilis L and Eucalyptus spp with LC50 values of 24.5, 44.0, 60.0, 75.0, 117.0 and 120.0 mg litre(-1), respectively. Combination tests between the LC50 and the maximum sub-lethal concentration (MSLC) were determined. Over 20 major components were identified in extracts from each plant species tested. Five essential oils and nine pure components were studied for their repellency against mosquito bites. Terpineol and 1,8-cineole were the most effective against Culex pipiens molestus bites offering complete protection for 1.6 and 2 h, respectively. PMID- 15662652 TI - Investigation of nickel suppression and cytocompatibility of surface-treated nickel-titanium shape memory alloys by using plasma immersion ion implantation. AB - Nickel-titanium (NiTi) shape memory alloys are increasingly being used in orthopedic applications. However, there is a concern that Ni is harmful to the human body. We have recently investigated the use of nitrogen, or oxygen plasma immersion ion implantation to mitigate this deleterious effect. Our results reveal that the near-surface Ni concentration in all the treated samples is significantly suppressed. In addition, our in vitro tests show that the plasma treated surfaces are cytologically compatible allowing the attachment and proliferation of osteoblasts. Among the two types of samples, the best biological effects are found on the samples with nitrogen implantation. PMID- 15662653 TI - All science is interdisciplinary--from magnetic moments to molecules to men (Nobel lecture). PMID- 15662654 TI - A boryl bridged complex: an unusual coordination mode of the BR2 ligand. PMID- 15662655 TI - Theoretical and synthetic studies on [Zn2(eta5-C5Me5)2]: analysis of the Zn-Zn bonding interaction. PMID- 15662656 TI - Nanophotonic light sources for fluorescence spectroscopy and cellular imaging. PMID- 15662657 TI - Detection of DNA strand breaks and oxidized DNA bases at the single-cell level resulting from exposure to estradiol and hydroxylated metabolites. AB - Long-term exposure to steroidal estrogens is a key factor contributing to increases in the risk of developing breast cancer. Proposed mechanisms include receptor-activated increases in the rate of cell proliferation leading to the accumulation of genetic damage resulting from reading errors, and the production of DNA damage by species arising from metabolism of 17beta-estradiol (E2) resulting in mutations. In support of the second mechanism, catechol metabolites of E2 induce DNA damage in vitro. In the present study, utilizing the single-cell gel electrophoresis (Comet) assay, we observed increases in the number of single strand breaks in estrogen receptor alpha-positive (MCF-7) and -negative (MDA-MB 231) breast cancer cells exposed to E2 (for 24 hr) or 4-hydroxy-17beta-estradiol (4-OH-E2; for 2 hr). The concentrations of 4-OH-E2 sufficient to induce these effects were approximately 100 nM, substantially lower than reported previously. The catechol 2-hydroxy-17beta-estradiol (2-OH-E2) also induced strand breaks. 2 OH-E2, often referred to as an improbable carcinogen in humans, is not a major metabolite of E2 in the breast; however, our findings show that it is as DNA damaging as 4-OH-E2. Formamidopyrimidine glycosylase posttreatment of E2-, 4-OH E2-, and 2-OH-E2-exposed MCF-7 cells led to an up to sixfold increase in mean tail moment, suggesting that oxidative DNA damage was formed. Comet formation could be partially attenuated by coincubation with dimethylsulfoxide, attributing a small DNA-damaging role to oxyradicals emanating from catechol redox cycling. Similar findings were obtained with MDA-MB-231 cells, indicating that estrogen receptor status is not relevant to these effects. Our observations show that exposure to E2 adds to the oxidative load of cells, and this may contribute to genomic instability. PMID- 15662658 TI - Genotoxicity of size-fractionated samples of urban particulate matter. AB - Urban particulate matter (UPM) includes particles of size smaller than 10 microm (PM10), which may impact on human respiratory and cardiovascular health. It has been reported previously that PM10 can induce DNA damage. We have collected size fractionated PM10 at the roadside and measured the induction of DNA damage by different-sized UPM using the alkaline Comet assay and the plasmid strand-break assay. We found that foil disks were more suitable for collecting UPM than quartz fiber filters, as the UPM could be easily extracted from the foil disks and accurately weighed. Using the Comet assay, all size fractions induced DNA damage in A549 lung epithelial cells, with the finer fractions (D50% = 0.65 microm and lower) inducing the most damage. In the plasmid strand-break assay, in which DNA damage is induced by free-radical species generated in solution, the most damage was also induced by the finer fractions, although the finest fraction (D50% < 0.43 microm) did not induce as much damage as D50% = 0.65 and 0.43 microm. When an organic extract of a standard UPM sample was compared to the whole particles and the washed particles in the Comet assay, it was found that around 75% of the damage induced by the whole UPM could be induced by the organic extract. These results show that finer particulates have the greatest ability to induce DNA damage in lung epithelial cells and naked DNA, and that both organic and inorganic components of the UPM contribute to its genotoxic effects. PMID- 15662659 TI - Bioimaging TOF-SIMS of tissues by gold ion bombardment of a silver-coated thin section. AB - The imaging time-of-flight secondary-ion-mass-spectrometry (TOF-SIMS) method was utilized to address the problem of cholesterol localization in rat tissues. Rat kidneys were fixed, cryoprotected by sucrose, frozen, sectioned by cryoultramicrotomy, and dried at room temperature. The samples were either covered with a thin silver layer or analyzed uncovered in an imaging TOF-SIMS instrument equipped with an Au1-3(+)-source. The yield of desorbed secondary ions for some species was up to 600-fold higher after silver coating of the samples. Reference samples of cholesterol were silver-coated and analyzed by TOF-SIMS to define significant peaks, specific for cholesterol. Such peaks were found at m/z = 386 (C27H46O+), m/z = 493 (C27H46O107Ag+), m/z = 495 (C27H46O109Ag+), m/z = 879 (C54H92O2 107Ag+), and m/z = 881 (C54H92O2 109Ag+). The silver-cationized cholesterol (493 < or = m/z < or = 495) signal was localized by imaging TOF-SIMS in the kidney sections and showed a high cholesterol content in the kidney glomeruli. A more diffuse distribution of cholesterol was also found over areas representing the cytoplasm or plasma membrane of the epithelial cells in the proximal tubules of rat kidney. PMID- 15662660 TI - Cortisol receptor blockade and seawater adaptation in the euryhaline teleost Fundulus heteroclitus. AB - To examine the role of cortisol in seawater osmoregulation in a euryhaline teleost, adult killifish were acclimated to brackish water (10 per thousand) and RU486 or vehicle was administered orally in peanut oil daily for five days at low (40 mg.kg(-1)) or high dose (200 mg.kg(-1)). Fish were transferred to 1.5 x seawater (45 per thousand) or to brackish water (control) and sampled at 24 h and 48 h after transfer, when Cl- secretion is upregulated. At 24 h, opercular membrane Cl- secretion rate, as Isc, was increased only in the high dose RU486 group. Stimulation of membranes by 3-isobutyl-1-methylxanthine and cAMP increased Isc in vehicle treated controls but those from RU486-treated animals were unchanged and membranes from brackish water animals showed a decrease in Isc. At 48 h, Isc increased and transepithelial resistance decreased in vehicle and RU486 groups, compared to brackish water controls. Plasma cortisol increased in all groups transferred to high salinity, compared to brackish water controls. RU486 treated animals had higher cortisol levels compared to vehicle controls. Vehicle treated controls had lower cortisol levels than untreated or RU486 treated animals, higher stimulation of Isc, and lower hematocrit at 24 h, beneficial effects attributed to increased caloric intake from the peanut oil vehicle. Chloride cell density was significantly increased in the high dose RU486 group at 48 hours, yet Isc was unchanged, suggesting a decrease in Cl- secretion per cell. Thus cortisol enhances NaCl secretion capacity in chloride cells, likely via glucocorticoid type receptors. PMID- 15662661 TI - Nitric oxide (NO) in normal and hypoxic vascular regulation of the spiny dogfish, Squalus acanthias. AB - Nitric oxide (NO) is a potent vasodilator in terrestrial vertebrates, but whether vascular endothelial-derived NO plays a role in vascular regulation in fish remains controversial. To explore this issue, a study was made of spiny dogfish sharks (Squalus acanthias) in normoxia and acute hypoxia (60 min exposure to seawater equilibrated with 3% oxygen) with various agents known to alter NO metabolism or availability. In normoxia, nitroprusside (a NO donor) reduced blood pressure by 20%, establishing that vascular smooth muscle responds to NO. L arginine, the substrate for NO synthase, had no hemodynamic effect. Acetylcholine, which stimulates endothelial NO and prostaglandin production in mammals, reduced blood pressure, but also caused marked bradycardia. L-NAME, an inhibitor of all NO synthases, caused a small 10% rise in blood pressure, but cell-free hemoglobin (a potent NO scavenger and hypertensive agent in mammals) had no effect. Acute hypoxia caused a 15% fall in blood pressure, which was blocked by L-NAME and cell-free hemoglobin. Serum nitrite, a marker of NO production, rose with hypoxia, but not with L-NAME. Results suggest that NO is not an endothelial-derived vasodilator in the normoxic elasmobranch. The hypertensive effect of L-NAME may represent inhibition of NO production in the CNS and nerves regulating blood pressure. In acute hypoxia, there is a rapid up regulation of vascular NO production that appears to be responsible for hypoxic vasodilation. PMID- 15662662 TI - Mixed reproductive strategy in Tubifex tubifex (Oligochaeta, Tubificidae)? AB - Despite the wide use of the tubificid oligochaete Tubifex tubifex in eco toxicological studies, the reproductive strategy adopted by the species, that can reasonably be assumed to be the key to its ecological success, is still not well investigated. This study reports on breeding experiments analysed by allozyme markers (Pgi, Pgm, and Idh), accompanied by a study of the sperm production in the species, with the aim to: (1) clarify the type of uniparental reproduction adopted by the species, (2) test the hypothesis that uniparental reproduction is reversible, (3) investigate the occurrence of biparental reproduction in laboratory cultures. Studies of parent-offspring comparison at polymorphic allozyme loci showed parthenogenetic reproduction and maintenance of asexuality in the parthenogenetic individuals. A cross-breeding test performed with couples made up of randomly assorted individuals, whose genotype combinations were suitable for discriminating between sexual and asexual reproduction, failed to show biparental reproduction in laboratory cultures: T. tubifex always reproduced parthenogenetically. Unexpectedly, spermiogenetic analysis indicated that both kinds of sperm produced by the species (eusperm and parasperm) were differentiated in individuals raised either in cohort cultures or in isolation (first, second, and third parthenogenetic generations), with a similar pattern of sperm production correlated to the sexual stage. Interestingly, there was no avoidance of mating in any of the collective cultures analysed. Concomitance between parthenogenetic reproduction and a "normal" male functionality, which is typical of a sexually reproducing species, could be justified by a mixed reproductive strategy or a pseudogamy process occurring in T. tubifex. However, several aspects of the reproductive behaviour of the species deserve further investigation. PMID- 15662663 TI - Glucocorticoid metabolism and Na+ transport in chicken intestine. AB - The role of aldosterone in regulation of electrogenic Na+ transport is well established, though mineralocorticoid receptors bind glucocorticoids with similar binding affinity as aldosterone and plasma concentration of aldosterone is much lower than glucocorticoids. In mammals, the aldosterone specificity is conferred on the low-selective mineralocorticoid receptors by glucocorticoid inactivating enzyme 11beta-hydroxysteroid dehydrogenase (11HSD) that converts cortisol or corticosterone into metabolites (cortisone, 11-dehydrocorticosterone) with lower affinity for these receptors. The present study examined the chicken intestine, whether changes in 11HSD activity are able to modulate the effect of corticosterone on Na+ transport, and how the metabolism of this hormone is distributed within the intestinal wall. This study shows that not only aldosterone, but also corticosterone (B), was able to increase the electrogenic Na+ transport in chicken caecum in vitro. The effect of corticosterone was higher in the presence of carbenoxolone, an inhibitor of steroid dehydrogenases, and was comparable to the effect of aldosterone. The metabolism of B in the intestine was studied; results showed oxidation of this steroid to 11-dehydrocorticosterone (A) and reduction to 11-dehydro-20beta-dihydrocorticosterone (20diA) as the main metabolic products at low nanomolar concentration of the substrate. In contrast, 20beta-dihydrocorticosterone and 20diA were the major products at micromolar concentration of B. Progesterone was converted to 20beta-dihydroprogesterone. The metabolism of corticosterone was localized predominantly in the intestinal mucosa (enterocytes). In conclusion, the oxidation at position C11 and reduction at position C20 suggest that both 11HSD and 20beta-hydroxysteroid dehydrogenase (20HSD) operate in the chicken intestine and that the mucosa of avian intestine possesses a partly different system of modulation of corticosteroid signals than mammals. This system seems to protect the aldosterone target tissue against excessive concentration of corticosterone and progesterone. PMID- 15662664 TI - Glucose transport by English sparrow (Passer domesticus) skeletal muscle: have we been chirping up the wrong tree? AB - Glucose uptake by mammalian skeletal muscle has been extensively covered in the literature, whereas the uptake of glucose by avian skeletal muscle has yet to be examined. As skeletal muscle provides the majority of postprandial glucose uptake in mammals, this study was designed to characterize the glucose transport mechanisms and glycogen content of avian skeletal muscle. In addition, plasma glucose levels were measured. English sparrow extensor digitorum communis (EDC) skeletal muscles were used for this study to quantify in vitro radiolabeled glucose uptake. Uptake of labeled glucose was shown to decrease in the presence of increasing unlabeled glucose and was maximal by 60 minutes of incubation. Various agents known to increase glucose transport in mammalian tissues, via the insulin and contraction-responsive pathways, were used to manipulate and characterize in vitro transport in birds. The typical effectors of the mammalian insulin pathway, insulin (2 ng/ml) and insulin-like growth factor-1 (48 ng/ml), did not increase skeletal muscle glucose transport. Likewise, inducers of the mammalian contraction-responsive pathway had no effect on glucose transport by in vitro avian skeletal muscle (5 mM caffeine, 2 mM AICAR (5'-aminoimidazole-4 carboxamide-1-b-D-ribofuranoside). Interestingly, 200 microM phloretin, an agent used to block glucose transport proteins, significantly inhibited its uptake (P<0.001). These results suggest that a glucose transporter is responsible for glucose uptake by avian skeletal muscle, albeit at unexpectedly low levels, considering the high plasma glucose concentrations (265.9+/-53.5 mg/dl) and low skeletal muscle glycogen content (9.1+/-4.11 nM glucose/mg) of English sparrows. PMID- 15662665 TI - Estradiol-17beta content in developing eggs and induced sex reversal of the medaka (Oryzias latipes). AB - To clarify the effect of exogenous estradiol-17beta (E2) on sex differentiation, the E2 content of developing eggs of Oryzias latipes was measured by radioimmunoassay. Endogenous E2 was present in lower concentrations in ovulated, mature eggs in the ovarian cavity than in intrafollicular oocytes. The E2 content of eggs precipitously declined to a minimum level by 2 days post-fertilization. The E2 content of eggs was affected by 24 hr of incubation in medium containing exogenous E2 at concentrations above 10 ng/ml. Short (24 hr) exposure of fertilized eggs in the early developmental stage to exogenous E2 at concentrations of 10 ng/ml induced sex reversal of some genotypic males to functional females. However, endogenous E2 levels in fertilized eggs might not influence sexual differentiation in embryogenesis. The present results suggest the possibility that concentrations of exogenous E2 higher than that of endogenous E2 triggers a priming step in the cascade of sex differentiation toward the female, and this effect is maintained. PMID- 15662666 TI - Regulation of capacitative Ca2+ entry by prothoracicotropic hormone in the prothoracic glands of the silkworm, Bombyx mori. AB - Measurements of Ca2+ influx in Fura-2/AM loaded steroidogenic cells (prothoracic glands; PGs) of the silkworm, Bombyx mori showed that application of the neuropeptide prothoracicotropic hormone (PTTH) can increase the intracellular [Ca2+]i. This PTTH-mediated Ca2+ influx in PG cells had kinetic patterns and pharmacological characteristics similar to those induced by thapsigargin. Namely, it produced increases in intracellular Ca2+ levels only in the presence of extracellular Ca2+, it was blocked by Gd3+ and 2-Aminoethoxydiphenylborate (2 APB), and it was unaffected by several toxins or compounds that block voltage activated Ca2+ channels. Moreover, the PTTH-stimulated increase of Ca2+ levels was eliminated in the presence of heparin (an IP3 receptor blocker), and by TMB-8 which also blocked any PTTH-dependent increase of ecdysteroid secretion. The PTTH mediated increase of Ca2+ levels was not affected by the non-hydrolysable GDP analogue, GDPbetaS, an indication that a G protein is not downstream of the PTTH receptor. These results argue strongly in favor of gating by the PTTH receptor of capacitative Ca2+ entry (CCE) channels (or store-operated Ca2+ channels (SOCs)) by a mechanism that does not involve any G proteins but requires the presence of functional IP3 receptors. Because the ability of PTTH to stimulate the [Ca2+]i levels of PG cells was completely mimicked by thapsigargin and exhibited a pharmacological profile similar to CCE mechanisms, we believe that PTTH directly regulates a CCE pathway in PG cells thereby activating a plethora of downstream regulators responsible for ecdysteroid secretion by the PGs of Bombyx mori. PMID- 15662667 TI - Function of the hepatic melanogenesis in the newt, Triturus carnifex. AB - Like the majority of lower vertebrates, the newt Triturus carnifex holds varying quantities of melanin and hemosiderin in the Kupffer cells of the liver. Following hypoxic treatment, the amount of these two pigments can increase to such an extent that they can occupy nearly a quarter of the surface of histological sections. A group of six specimens, anesthetised with chlorbutol, were subjected to hypoxic treatment by keeping them in a respiratory chamber containing degassed water under vacuum, with only 1.1 ppm of residual oxygen, until they had consumed the oxygen completely (4 hours, at a temperature of 18 degrees C). Using hematological and histochemical techniques and computerised image analysis, it has been shown that hypoxic animals not only increase the extent of the melanic areas of the liver from about 5-7% to almost 24% compared to control groups kept under two different respiratory conditions (6 anesthetised specimens exposed to the air and 6 submerged in normoxic water), they also went through a remarkable hemolytic process to justify a parallel increase in hemosiderin deposits. Melanin was extracted from the liver by keeping fragments of the organ for one hour at 37 degrees C in an oxidising solution (20 mL of benzyl alcohol, 10 mL of acetone, 5 mL of 10% hydrogen peroxide, and 4 drops of concentrated ammonia solution), then quickly rinsing them in 50% acetone and subsequently letting them stand for 6 hours in 10 mL of distilled water alkalised to pH 12 with a drop of ammonia solution. The extract was then left to sediment at pH 2.5 and the black precipitate washed and dried under vacuum. Elemental and spectrophotometric analyses revealed a significant presence of purines in the melanic pigment. This phenomenon can be explained by the animals' need under hypoxic crisis to rapidly neutralise purines resulting from lysis of the nucleated red blood cells by introducing them into an inert molecular complex. A partial model of structure is proposed here. Synthesis of the mixed polymer is possible through the well-known capacity of ferrous iron to activate tyrosinase (the enzyme responsible for melanogenesis) even in the absence of DOPA. PMID- 15662668 TI - The level of ADAM12-S in maternal serum is an early first-trimester marker of fetal trisomy 18. AB - BACKGROUND: ADAM12-S is a pregnancy-associated insulin-like growth factor binding protein-3 (IGFBP-3) and IGFBP-5 protease present in human gestational serum. Recently, maternal serum levels of ADAM12-S were found to be markedly reduced during the first trimester of pregnancies with a Down syndrome (DS) fetus. On the basis of this finding, it was suggested that ADAM12-S might be a useful maternal serum marker of fetal chromosomal disease. OBJECTIVE: Retrospective examination of the use of ADAM12-S as a marker for fetal trisomy 18. METHOD: Serum samples were obtained from ten women during the first semester of their pregnancies with fetuses that had trisomy 18. An ELISA was used to determine the levels of ADAM12 in maternal serum. Results were compared to ADAM12-S levels, previously measured in the serum of 170 women carrying normal pregnancies during the first trimester. RESULTS: In all cases, the ADAM12-S concentration in maternal serum samples was lower in trisomy 18 pregnancies than in normal pregnancies, with a median multiple of the median (MoM) of 0.28 (p < 0.001) CONCLUSION: A reduced concentration of ADAM12-S in maternal serum is a promising marker for foetal trisomy 18, as well as for DS. PMID- 15662669 TI - Sonographic atypical vascular coiling of the umbilical cord. AB - OBJECTIVE: To investigate whether an atypical umbilical coiling pattern at prenatal sonography is associated with adverse pregnancy outcome. METHODS: A targeted sonographic evaluation of the umbilical cord (UC) was performed in 758 women with singleton gestation, and gestational age above 20 weeks. Atypical coiling was defined as the presence of a spring-shape UC (supercoiling) or an unusual, aperiodic coiling pattern (uncoordinated coiling). Umbilical artery Doppler assessment was conducted in cases with atypical coiling. Pregnancy and neonatal outcomes were investigated. RESULTS: Of the study population, 7 and 16 fetuses had an umbilical cord with uncoordinated coiling and supercoiling respectively. Three umbilical cords had a single umbilical artery. Eight patients delivered before 34 weeks of gestation. Eight fetuses were growth restricted. In seven cases, abnormal sonographic findings were detected (three meconium peritonitis, two severe hydronephrosis and two cardiac anomalies). One fetus affected by trisomy 18 presented multiple anomalies. Perinatal death occurred in three cases. Of the surviving newborns, eight were admitted to NICU. Umbilical artery Doppler waveforms presented a systolic notch in seven (30.4%) cases. CONCLUSIONS: The presence of an atypical umbilical cord vascular coiling is associated with an increased risk of unfavourable pregnancy outcome. The identification of an umbilical artery notch at Doppler investigation is frequently associated with an atypical UC coiling pattern. PMID- 15662670 TI - Atom-transfer radical reactions under mild conditions with [{RuCl2(1,3,5 C6H3iPr3)}2] and PCy3 as the catalyst precursors. PMID- 15662671 TI - Template-assisted cross olefin metathesis. PMID- 15662672 TI - One-pot conversion of phospholide ions into beta-functional phosphinines. PMID- 15662673 TI - Photoswitchable microlens arrays. PMID- 15662674 TI - Methane and carbon dioxide storage in a porous van der Waals crystal. PMID- 15662675 TI - Formation of [60]fullerene nanoclusters with controlled size and morphology through the aid of supramolecular rod-coil diblock copolymers. PMID- 15662676 TI - Ba(4)Mo(12)S(18): A superconductor containing the dimeric unit (Mo(6))(2)S(24), the missing link between the Mo(6)S(14) and Mo(9)S(17) units. PMID- 15662677 TI - Physicochemical characterization of porous materials: spatially resolved accessibility of zeolite crystals. PMID- 15662678 TI - Lamellar bridged silsesquioxanes: self-assembly through a combination of hydrogen bonding and hydrophobic interactions. AB - The synthesis of four bis(trialkoxysilylated) organic molecules capable of self assembly--(EtO)3Si(CH2)3NHCONH-(CH2)n-NHCONH(CH2)3Si(OEt)3 (n = 9-12)- associating urea functional groups and alkylidene chains of variable length is described. These compounds behave as organogelators, forming supramolecular assemblies thanks to the intermolecular hydrogen bonding of urea groups. Whereas fluoride ion-catalysed hydrolysis in ethanol in the presence of a stoichiometric amount of water produced amorphous hybrids, acid-catalysed hydrolysis in an excess of water gave rise to the formation of crystalline lamellar hybrid materials through a self-organisation process. The structural features of these nanostructured organic/inorganic hybrids were analysed by several techniques: attenuated Fourier transformed infrared (ATR-FTIR), solid-state NMR spectroscopy (13C and 29Si), scanning and transmission electron microscopy (SEM and TEM) and powder X-ray diffraction (PXRD). The reaction conditions, the hydrophobic properties of the long alkylidene chains and the hydrogen-bonding properties of the urea groups are determining factors in the formation of these self-assembled nanostructured hybrid silicas. PMID- 15662679 TI - Size-selective formation of C78 fullerene from a three-dimensional polyyne precursor. AB - Multicyclic cagelike cyclophanes 2 a and 2 b containing cyclobutene rings have been prepared as precursors of three-dimensional polyynes C78H18 (1a) and C78H12Cl6 (1b), respectively. Laser irradiation of 2a and 2b induced expulsion of the aromatic fragment, indane, to give the three-dimensional polyyne anions C78H18- and C78H12Cl6-, respectively. Whereas the former anion lost only four hydrogen atoms to form C78H14-, complete loss of all hydrogen and chlorine atoms was observed from the latter anion, to yield a C78- ion that has a fullerene structure which was proven by its characteristic fragmentation pattern. PMID- 15662680 TI - 3-Fluoropiperidines and N-methyl-3-fluoropiperidinium salts: the persistence of axial fluorine. AB - It has previously been shown that the fluorine atom in N-protonated 3 fluoropiperidine salts in water strongly prefers the axial orientation in the six membered ring chairs. In the present work we examine the proposition that the N methyl salts are equally disposed to present axial fluorine. Initially, we explored this point by comparing the structures of the corresponding NH2+, NHMe+, and NMe2+ salts by means of density functional theory (DFT), ab initio, and MMFF force field calculations with and without aqueous solvation models. The predictions unambiguously pointed to axial fluorine for all salts investigated, including those with simultaneous axial F and (N)Me. The calculations were followed by synthesis of the corresponding series of 4,4-diphenylpiperidinium salts. These were evaluated by one- and two-dimensional NMR spectroscopy in [D6]DMSO to fully corroborate the axial disposition of the fluorine in each of the compounds. X-ray crystal structure determinations were likewise performed for the diphenyl-3-fluoro NH2+ and NMe2+ systems to substantiate axial-F. Comparison of the X-ray structures of the fluorinated and unfluorinated NMe2+ salts reveals that the fluorine resides axial in spite of substantial steric compression. While the charge-dipole phenomenon responsible for the axial-F conformation in the parent protonated fluoropiperidinium compounds carries over to doubly alkylated salts, we show that it extends to molecular orientation in the packing of the unit cells in the solid state as well. Finally, using the computational methods that successfully motivated our synthesis and structural work, we have made predictions for a number of new structures and re-examined some parallel results reported by the Eliel group in the early 1970s. Although C-F...H-N hydrogen bonds are reported to be weak and few in number, the CF...HN charge-dipole orienting effect is a powerful directing force that matches the hydrogen-bond in both its energetic contribution and conformational consequences. PMID- 15662681 TI - [MnIII2 (5-Rsaltmen)2NiII(pao)2(L)]2+: an S(T)=3 building block for a single chain magnet that behaves as a single-molecule magnet. AB - Mn(III)-Ni(II)-Mn(III) linear-type trinuclear complexes bridged by oximate groups were selectively synthesized by the assembly reaction of [Mn2(5 Rsaltmen)2(H2O)2](ClO4)2 (5-Rsaltmen2-=N,N'-(1,1,2,2-tetramethylethylene) bis(5-R salicylideneiminate); R=Cl, Br) with [Ni(pao)2(phen)] (pao-=pyridine-2 aldoximate; phen=1,10-phenanthroline) in methanol/water: [Mn2(5 Rsaltmen)2Ni(pao)2(phen)](ClO4)2 (R=Cl, 1; R=Br, 2). Structural analysis revealed that the [Mn(III)-ON-Ni(II)-NO-Mn(III)] skeleton of these trimers is in every respect similar to the repeating unit found in the previously reported series of 1D materials [Mn2(saltmen)2Ni(pao)2(L1)2](A)(2) (L(1)=pyridine, 4-picoline, 4 tert-butylpyridine, N-methylimidazole; A=ClO4-, BF4-, PF6-, ReO4-). Recently, these 1D compounds have attracted a great deal of attention for their magnetic properties, since they exhibit slow relaxation of the magnetization (also called single-chain magnet (SCM) behavior). This unique magnetic behavior was explained in the framework of Glauber's theory, generalized for chains of ferromagnetically coupled anisotropic spins. Thus, in these 1D compounds, the [Mn(III)-ON-Ni(II)-NO Mn(III)] unit was considered as an S(T)=3 anisotropic spin. Direct-current magnetic measurements on 1 and 2 confirm their S(T)=3 ground state and strong uniaxial anisotropy (D/k(B) approximately -2.4 K), in excellent agreement with the magnetic characteristic deduced in the study on the SCM series. The ac magnetic susceptibility of these trimers is strongly frequency-dependent and characteristic of single-molecule magnet (SMM) behavior. The relaxation time tau shows a thermally activated (Arrhenius) behavior with tau0 approximately 1x10(-7) s and Delta(eff)/k(B) approximately 18 K. The effective energy barrier for reversal of the magnetization Delta(eff) is consistent with the theoretical value (21 K) estimated from |D| S2T. The present results reinforce consistently the interpretation of the SCM behavior observed in the [Mn2(saltmen)2Ni(pao)2(L1)2](A)2 series and opens new perspectives to design single-chain magnets. PMID- 15662682 TI - Aryl palladium carbene complexes and carbene-aryl coupling reactions. AB - Transmetalation of an aminocarbene moiety from [W(CO)5{C(NEt2)R}] to palladium leads to isolable monoaminocarbene palladium aryl complexes [{Pd(mu Br)Pf[C(NEt2)R]}2] (R = Me, Ph; Pf = C6F5). When [W(CO)5{C(OMe)R}] is used, the corresponding palladium carbenes cannot be isolated since these putative, more electrophilic carbenes undergo a fast migratory insertion process to give alkyl palladium complexes. These complexes could be stabilized in the eta3-allylic form for R = 2-phenylethenyl or in the less stable eta3-benzylic fashion for R = Ph. Hydrolysis products and a pentafluorophenylvinylic methyl ether (when R = Me) were also observed. The monoaminocarbenes slowly decompose through carbene-aryl coupling to produce the corresponding iminium salts and, depending on the reaction conditions, the corresponding hydrolysis products. The electrophilicity of the carbene carbon, which is mainly determined by the nature of the heteroatom group, controls the ease of evolution by carbene-aryl coupling. Accordingly, no carbene-aryl coupling was observed for a diaminocarbene palladium aryl complex. PMID- 15662683 TI - Reaction mechanism of porphyrin metallation studied by theoretical methods. AB - We have studied the reaction mechanism for the insertion of Mg2+ and Fe2+ into a porphyrin ring with density functional calculations with large basis set and including solvation, zero-point and thermal effects. We have followed the reaction from the outer-sphere complex, in which the metal is coordinated with six water molecules and the porphyrin is doubly protonated, until the metal ion is inserted into the deprotonated porphyrin ring with only one water ligand remaining. This reaction involves the stepwise displacement of five water molecules and the removal of two protons from the porphyrin ring. In addition, a step seems to be necessary in which a porphyrin pyrrolenine nitrogen atom changes its interaction from a hydrogen bond to a metal-bound solvent molecule to a direct coordination to the metal ion. If the protons are taken up by a neutral imidazole molecule, the deprotonation reactions are exothermic with minimal barriers. However, with a water molecule as an acceptor, they are endothermic. The ligand exchange reactions were approximately thermoneutral (+/-20 kJ mol(-1), with one exception) with barriers of up to 72 kJ mol(-1) for Mg and 51 kJ mol(-1) for Fe. For Mg, the highest barrier was found for the formation of the first bond to the porphyrin ring. For Fe, a higher barrier was found for the formation of the second bond to the porphyrin ring, but this barrier is probably lower in solution. No evidence was found for an initial pre-equilibrium between a planar and a distorted porphyrin ring. Instead, the porphyrin becomes more and more distorted as the number of metal-porphyrin bonds increase (by up to 191 kJ mol( 1)). This strain is released when the porphyrin becomes deprotonated and the metal moves into the ring plane. Implications of these findings for the chelatase enzymes are discussed. PMID- 15662684 TI - Organic-inorganic hybrids based on novel bimolecular [Si2W22Cu2O78(H2O)]12- polyoxometalates and the polynuclear complex cations [Cu(ac)(phen)(H2O)]n n+ (n=2, 3). AB - The reaction of a monosubstituted Keggin polyoxometalate (POM) generated in situ with copper-phenanthroline complexes in excess ammonium or rubidium acetate led to the formation of the hybrid metal organic-inorganic compounds A7[Cu2(ac)2(phen)2(H2O)2][Cu3(ac)3(phen)3(H2O)3][Si2W22Cu2O78(H2O)].approximately 18 H2O (A=NH4+ (1), Rb+ (2); ac=acetate; phen=1,10-phenanthroline). These compounds are constructed from inorganic and metalorganic interpenetrated sublattices containing the novel bimolecular Keggin POM, [Si2W22Cu2O78(H2O)]12-, and Cu-ac-phen complexes, [Cu(ac)(phen)(H2O)]n n+ (n=2, 3). The packing of compound 1 can be viewed as a stacking of open-framework layers parallel to the xy plane built of hydrogen-bonded POMs, and zigzag columns of pi-stacked Cu-ac phen complex cations running along the [111] direction. Magnetic and EPR results are discussed with respect to the crystal structure of the compounds. DFT calculations on [Cu(ac)(phen)(H2O)]n n+ cationic complexes have been performed, to check the influence of packing in the complex geometry and determine the magnetic exchange pathways. PMID- 15662685 TI - A novel L1CAM mutation with L1 spectrum disorders. AB - X-linked hydrocephalus, HSAS (hydrocephalus due to stenosis of aqueduct of Sylvius), MASA (mental retardation, aphasia, shuffling gait, and adducted thumbs), and CRASH (corpus callosum hypoplasia, retardation, adducted thumbs, spastic paraplegia, and hydrocephalus) syndromes are allelic disorders. X-linked hydrocephalus and associated phenotypes are due to mutations in the L1CAM gene, which has been identified as a coding neural cell adhesion molecule. We report two cases of L1 spectrum disorders within the same family. The first case was diagnosed by ultrasonographic examination prenatally and the second case was diagnosed postnatally. Both patients and their mothers carry a novel mutation of the L1CAM gene. In this family, nine X-linked hydrocephalus and five female carriers were found in three generations, and molecular genetic analysis was performed to detect the asymptomatic carriers. PMID- 15662686 TI - DNA-based prenatal diagnosis for severe and variant forms of multiple acyl-CoA dehydrogenation deficiency. AB - OBJECTIVES: Multiple acyl-CoA dehydrogenation deficiency (MADD) is a clinically heterogeneous disorder of mitochondrial fatty acid, amino acid, and choline oxidation due to mutations in the genes encoding electron transfer flavoprotein (ETF) or ETF ubiquinone oxidoreductase (ETFQO). So far, prenatal diagnosis of MADD has relied mostly on second-trimester biochemical analyses of amniotic fluid or cultured amniocytes. We report here on an alternative DNA-based approach for prenatal diagnosis in pregnancies at risk of MADD. METHODS: We used our knowledge of the mutational status in three unrelated families with a history of MADD to perform direct sequencing for the familial mutations using genomic DNA isolated from chorionic villus samples (CVS) at gestational week 10 to 11. RESULTS: Within two days, we were able to carry out accurate DNA-based prenatal testing in one pregnancy at risk of severe MADD, and in two pregnancies at risk of variant forms of MADD. CONCLUSION: This is the first report of DNA-based prenatal diagnosis of MADD. Our molecular approach is suitable for fast and reliable first-trimester prenatal diagnosis in pregnancies at risk of severe and variant forms of MADD. PMID- 15662687 TI - Prenatal diagnosis of Aicardi-Goutieres syndrome. AB - OBJECTIVES: Aicardi-Goutieres syndrome is an autosomal recessive neurodegenerative disorder inducing cerebral atrophy, intracerebral calcification and developmental arrest. Diagnosis requires the presence of progressive encephalopathy with clinical onset shortly after birth, typical neuroimaging features associated with a raised blood and cerebrospinal fluid interferon-alpha level.A case of prenatal diagnosis of Aicardi-Goutieres syndrome is reported. METHODS: An MRI performed at 26 gestational weeks showed bilateral calcifications and white matter abnormalities, cerebral anomalies typically described in this disease. The fetal blood analysis revealed an increase in interferon-alpha. RESULTS: Therefore, the prenatal diagnosis of Aicardi-Goutieres syndrome in this fetus was based on the following facts: the familial background with the affected first child and consanguineous parents, a normal pregnancy and normal fetal growth, cerebral anomalies diagnosed on prenatal ultrasound and cerebral MRI, raised interferon-alpha in the fetal serum and no evidence of any infectious etiology. The autopsy performed postdelivery at 28 1/2 weeks' gestation confirmed the diagnosis of Aicardi-Goutieres syndrome. CONCLUSION: To the best of our knowledge, this is the first prenatal diagnosis of this syndrome. Such a diagnosis may prove useful for families at risk as long as genetic screening is not available. PMID- 15662688 TI - Is fetal gender a risk factor for severe congenital cytomegalovirus infection? AB - Cytomegalovirus is the main cause of congenital viral infection and amniotic fluid viral load appears to be the single nonclinical prognostic factor. However, as in other infectious diseases, host genetics may influence the severity of the disease. To test this hypothesis, we looked retrospectively at the fetal gender in cases of severe congenital cytomegalovirus infection in our database. We also analyzed the international English literature covering this subject between 1985 and 2003. The proportion of females with brain abnormalities was statistically different from that of males (62/258: 24% vs 30/251: 12%, p = 0.004). The risk of abnormal brain development in infected fetuses was twice as high in females than in males (Chi(2) = 8.7; OR = 2, IC [1.26-3.21]). In our cases, amniotic fluid CMV DNA load was not significantly higher in males than in females (p = 0.06) and was also similar in severely and non-severely infected fetuses (p = 0.09). PMID- 15662689 TI - Maternal cell contamination of prenatal samples assessed by QF-PCR genotyping. AB - OBJECTIVES: To establish the genotype of cultured cells from a cohort of amniotic fluid and chorionic villus samples, and compare this genotype with that obtained from uncultured material from the same sample, in order to assess the frequency and significance of maternal cell contamination of prenatal samples. METHODS: Quantitative fluorescence-polymerase chain reaction (QF-PCR) was carried out by amplification of microsatellite markers using fluorescence-labelled primers, followed by quantitative analysis of the allele peaks on a genetic analyser. A multiplex of 12 primer pairs for four loci on each of chromosomes 13, 18 and 21 was used. RESULTS: A total of 307 prenatal samples were tested. Of the 254 amniotic fluid samples, 39.8% had some degree of bloodstaining, ranging from 5% bloodstaining in the cell pellet to heavily bloodstained fluid. Uncultured samples were tested by QF-PCR analysis and the cultured cells were tested by both QF-PCR and karyotype analysis. Of the samples, 90.2% had the same single genotype on direct and cultured material. Two samples (0.65%) were mosaic for an aneuploidy cell line. A second genotype, interpreted as maternal cell contamination, was identified in direct and/or cultured preparations in 9.1% of samples, 17.8% of which were not bloodstained. Seven amniotic fluid samples (2.8%) showed maternal cell contamination in cultured material. CONCLUSIONS: For heavily bloodstained amniotic fluid samples, a maternal blood specimen may help interpret the results of rapid trisomy testing, followed by confirmation of the fetal origin of cultured cells. QF-PCR analysis has established a higher incidence of maternal cell contamination of cultured amniocytes than previous reports; the presence of MCC (maternal cell contamination) in cultured cells from samples with no bloodstaining underlines the need for karyotype analysis of more than one XX culture. PMID- 15662690 TI - Accepting or declining the offer of prenatal screening for congenital defects: test uptake and women's reasons. AB - OBJECTIVES: Prenatal screening for Down syndrome has become standard practice in many western countries. In the Netherlands, however, prenatal screening tests for congenital defects are not offered routinely. The present study aims to assess test uptake in a large, unselected population of pregnant women, and to give more insight into the decision for or against prenatal screening through nuchal translucency measurement or maternal serum screening. PATIENTS AND METHODS: The study is part of a randomized controlled trial with two groups, each being offered a different prenatal screening test, and a control group. Pregnant women received postal questionnaires at three stages of their pregnancy. RESULTS: Of the women being offered the nuchal translucency measurement or the second trimester maternal serum test, 53 and 38% respectively accepted the test offer. The main reasons for accepting were 'gaining knowledge about the health of the foetus/curiosity' (50%), 'favourable characteristics of the screening test' (18%), and 'increased risk of having a child with DS' (15%). The main reasons for declining were 'unfavourable characteristics of the screening test' (42%), 'not applicable/not necessary' (35%), 'anxiety/uncertainty' (36%), 'adverse characteristics of the invasive tests' (32%), and 'being against abortion' (15%). DISCUSSION: The uptake of prenatal screening was relatively low, and different distributions of reasons were reported, compared to other studies. These differences may be due to the specific Dutch situation in which prenatal screening is not part of standard prenatal care. The question arises as to whether informed decision-making would be reduced if prenatal screening became routinised. PMID- 15662691 TI - Detection of mosaicism for primary trisomies in prenatal samples by QF-PCR and karyotype analysis. AB - OBJECTIVES: QF-PCR can be used to rapidly diagnose primary trisomy in prenatal samples. Our objectives were to estimate the prevalence of primary trisomy mosaicism for chromosomes 13, 18 or 21 in a cohort of prenatal samples, and to compare and contrast the detection of this mosaicism using both QF-PCR and karyotype analysis. METHODS: Data was collated from all prenatal samples displaying mosaicism for a primary trisomy between June 2000 and March 2004. Levels of mosaicism were estimated and samples were categorised according to the cell population in which the mosaicism was detected. RESULTS: In a total of 8983 samples, 18 samples (0.20%) displaying mosaicism were detected, including trisomy 13 (three samples), trisomy 18 (seven samples), trisomy 21 (seven samples) and mosaic triploidy (one sample). This included 7 amniotic fluid and 11 chorionic villus samples. Mosaicism was detected by QF-PCR in 12 samples and by karyotype analysis in 8 samples. CONCLUSIONS: QF-PCR can detect mosaicism when the abnormal cell line contributes at least 15% of the whole sample. Use of both karyotype and QF-PCR analysis leads to the detection of more cases of mosaicism than either test alone. PMID- 15662692 TI - Characterization of a small supernumerary marker chromosome as r(8) at prenatal diagnosis by MFISH. AB - OBJECTIVE: To identify the mosaic marker chromosome detected in amniotic fluid cells of a 26-year-old woman, with raised triple test values and an ultrasound scan, which showed a fetus with echogenic bowels. METHODS: Routine karyotyping with G- and C-banding was carried out for both, amniotic fluid at 18 weeks of gestation as well as fetal blood at 22(+6) weeks. Peripheral blood of both parents was karytoyped. MFISH and the all centromeric human probe were used on fetal lymphocytes to identify the marker chromosome. RESULTS: Both parents had a normal karyotype. Amniotic fluid culture showed a de novo supernumerary marker chromosome (SMC) in 14 of the 30 colonies from four different cover slip cultures. The marker was confirmed in 50% of the fetal lymphocytes. G- and C banding provided little information except that the marker had some heterochromatic material. The all centromeric human probe also showed the presence of a centromere along with a rim of euchromatic material. MFISH identified this ring marker to be belonging to chromosome 8. CONCLUSIONS: SMCs with chromosome 8 have been shown to be variable phenotypes. Presence of only heterochromatic material seems to have no discernable phenotypic effects, but, with the presence of euchromatic material, mental and physical developmental delay has been reported. The parents opted to go ahead with the pregnancy and an apparently normal female baby was born at 40 weeks with no complications. PMID- 15662693 TI - Down syndrome screening marker levels in women with a previous aneuploidy pregnancy. AB - BACKGROUND: In Down syndrome screening programmes, women with a previous affected pregnancy are assumed to have the same marker distribution as those without a family history. This assumption needs to be tested. METHODS: Information on previous aneuploidy pregnancies was routinely sought on the test request forms in three centres, Leeds, Romford and the Fetal Medicine Centre, London. For each woman with a previous aneuploidy (case), five unaffected pregnancies to women without a history were selected as controls. The markers tested included maternal serum free beta-human chorionic gonadotrophin (hCG), pregnancy-associated plasma protein A (PAPP-A), alpha-fetoprotein, unconjugated estriol and ultrasound nuchal translucency thickness. RESULTS: There were 375 cases: 303 with previous Down syndrome, 63 with Edwards syndrome and 9 with Patau's syndrome. There was a statistically significant difference between cases and controls, in the distribution of free beta-hCG and PAPP-A levels, adjusted for gestation. On average, free beta-hCG was increased by 10% in a subsequent pregnancy after aneuploidy (p < 0.005, Wilcoxon rank sum test) and for PAPP-A the increase was 15% (p < 0.0001). No other marker was significantly different. CONCLUSION: Risk calculation algorithms need to be modified to take account of the increased marker levels. Until data from sufficient affected pregnancies are available for study, it would be prudent to assume that the same increase as in unaffected pregnancies applies. PMID- 15662694 TI - Correlation between biomagnetic and Doppler findings of the uterine artery in normal and preeclamptic pregnancies. AB - OBJECTIVES: To elucidate the hemodynamics of the uterine artery in normal and abnormal pregnancies by use of Doppler ultrasound and SQUID biomagnetometry. METHOD: Two hundred and three women (gestational age 28-42 weeks) were included in the study. Forty-three of them had preeclampsia and/or intrauterine growth restriction and 160 were normal. Uterine artery waveform measurements were evaluated by use of pulsatility index (PI) (normal value PI<1.45). Biomagnetic signals of uterine arteries were recorded and analyzed with Fourier analysis. The biomagnetic signals were distributed according to spectral amplitudes as high (140-300 fT/ radicalHz), low (50-110 fT/ radicalHz) and borderline (111-139 fT/ radicalHz). RESULTS: There was a statistically significant difference between normal and abnormal pregnancies concerning the waveform amplitudes (p < 0.001) and the PI index (p < 0.001). Specifically, we noticed high biomagnetic amplitudes in most normal pregnancies (92.5%) and low biomagnetic amplitudes in most preeclamptic cases (90.7%). CONCLUSIONS: Our study results indicated that biomagnetic measurements of uterine artery might prove to be useful in the evaluation of the fetal well being, especially in cases of preeclampsia and intrauterine growth restriction. PMID- 15662695 TI - Sonographic markers of exencephaly below 10 weeks' gestation. AB - We report two cases of exencephaly diagnosed by transvaginal ultrasonography at 8 weeks 4 days and at 9 weeks 3 days of gestation. Both cases presented an irregular cephalic pole, and, in the case seen at 8 weeks 4 days, brain vesicles were also absent, whereas in the case seen at 9 weeks 3 days, the midline echo was indistinguishable with disorganized choroid plexuses. In both cases, anencephaly was evident at 11 and 12 weeks' gestation and the postmortem confirmed the diagnosis. PMID- 15662696 TI - Increased nuchal translucency and split-hand/foot malformation in a fetus with an interstitial deletion of chromosome 2q that removes the SHFM5 locus. AB - OBJECTIVES: To describe and discuss the clinical, cytogenetic and molecular findings in a fetus with the first prenatally detected interstitial deletion of chromosome 2q. CASE REPORT: A fetus with increased nuchal translucency on routine ultrasound examination at 13 weeks' gestation was found to have severe upper-limb abnormalities on follow-up ultrasound examination at 16 weeks. The pregnancy was terminated, and the autopsy revealed monodactyly of the right upper limb, oligodactyly of the left upper limb and bilateral split foot, as well as atrial and ventricular septal defects and mild facial dysmorphism. RESULTS: Cytogenetic studies and haplotype analysis of the fetus and both parents showed that the fetus carried a de novo deletion encompassing a region of about 30 Mb on the paternal chromosome 2q (karyotype 46,XX,del(2)(q24.2-q32.2)). CONCLUSION: This is the first instance of increased nuchal translucency associated with a chromosome 2q deletion. Moreover, the striking malformations affecting all four of the fetus' limbs support previous suggestions that a novel locus for split-hand/foot malformation (SHFM5) lies on chromosome 2q31. PMID- 15662697 TI - Comparison of the use of amniocentesis in two countries with different policies for prenatal testing: the case of France and the United States. AB - OBJECTIVE: To compare maternal age- and education-specific use of amniocentesis in France and the United States. METHODS: We used two nationally representative datasets, National Perinatal Survey of 1998 in France (n = 12 816) and National Center for Health Statistics birth data for 1997 in the United States (n = 3 799 975). Analyses included binomial regression with test of interactions between country, maternal age and education. RESULTS: Amniocentesis use was more than threefold greater in France than in the United States (Risk Ratio (RR) 3.2, 95% CI, 3.1-3.4). This was true across maternal age and education groups. Differences in use of amniocentesis were greatest, however, for women with lower levels of education and older (>/=38 years) women. CONCLUSION: Our results suggest greater use and lesser disparities in maternal age- and education-specific use of amniocentesis in France as compared to that in the United States. These differences may be due to several factors, including differences in women's cultural values and preferences. They may also represent barriers to effective access to prenatal testing, particularly for women in lower socioeconomic groups, in the United States. PMID- 15662698 TI - Split notochord syndrome variant: prenatal findings and neonatal management. AB - Spilt notochord syndrome is an extremely rare form of spinal dysraphism characterized by a complete cleft of the spine and a persistent communication between endoderm and ectoderm. A variant of split notochord syndrome was diagnosed in a 25-week-old fetus showing a prolapsed congenital colostomy and a spinal cystic lesion. The final diagnosis included protruding colon segment, imperforate anus with a rectourethral fistula and lipomyelomeningocele. The ultrasound features of the condition and the post-natal management are discussed. The neonate was successfully treated with a posterior sagittal anorectoplasty, while the lipomyelomeningocele was resected at a later stage. PMID- 15662699 TI - Blinded review of Papanicolaou smears. PMID- 15662700 TI - Aberrant, highly hyperdiploid cells in human papillomavirus-positive, abnormal cytologic samples are associated with progressive lesions of the uterine cervix. AB - BACKGROUND: Infection with oncogenic-type human papillomavirus (HPV) and consecutive cytologic abnormalities of the uterine cervix precede the evolution of carcinoma. However, the specificity of both changes is too low to predict the true malignant potential of the change in a given time point, because the majority of the HPV infections revert to normal with time. In preliminary studies, the authors demonstrated that, among many dysregulatory phenomena at the cytologic level, the occurrence of significant DNA content aberrations were in good correlation with progressive cervical changes; and, as a marker for this, the significance of cells with nuclear DNA content > 9c (9c cells) was investigated using slide-based cytometry. The objective of the current study was to determine whether 9c cells in cytologic samples that presented with dysplasia and with high-risk HPV types were associated with the development of higher grade cervical intraepithelial neoplasia (CIN II+). METHODS: Samples with positive cytologic diagnoses (atypical squamous cells of undetermined significance, low grade squamous intraepithelial lesion [LSIL], and high-grade squamous intraepithelial lesion [HSIL]) were evaluated for both parameters and were related to outcomes and, if available, to histology results over a follow-up of up to 4 years. RESULTS: Although the presence of high-risk HPV was demonstrable in almost all samples with CIN II+ (96.7%), the virus genome alone proved to have a poor positive predictive value (56.1%) for higher grade CIN. Cytology and the demonstration of 9c cells resulted in less sensitivity compared with the occurrence of high-risk HPV (70.4% and 73.7%, respectively) but in significantly greater specificity (91.3% and 89.2%, respectively). Moreover, a positive predictive value of 81.8% for CIN II+ could be calculated for 9c cells. HSIL morphology, unfavorable HPV type, and DNA cytometry together excluded the highest rate of CIN > or = II with 100% sensitivity and 91.3% specificity. CONCLUSIONS: Biologic parameters, in addition to cytology, help to define the nature of cervical dysplasias more accurately. Oncogenic HPV types and hyperdiploid cells with DNA content > 9c disclosed cytologic changes with the greatest malignant potential. Cancer PMID- 15662701 TI - DNA damage in lymphocytes of rural Indian women exposed to biomass fuel smoke as assessed by the Comet assay. AB - The Comet assay has found wide acceptance in monitoring human genotoxicity caused by lifestyle and occupational and environmental factors. In the present study, we have used the Comet assay to measure the DNA damage in a population of rural Indian women cooking with biomass fuels (BMFs; fire wood and cow dung cakes). Out of 144 volunteers, 70 used BMFs for domestic cooking, while the remaining 74 used liquefied petroleum gas (LPG) and served as a reference population. All the individuals had comparable socioeconomic backgrounds and were between 20 and 55 years of age. Significantly higher levels of DNA damage were observed for BMF users than for LPG users. For BMF users in comparison with the reference population, Olive tail moment was 3.83 +/- 0.15 (arbitrary units) vs. 2.77 +/- 0.07 (P < 0.001); % tail DNA was 11.19 +/- 0.35 vs. 8.29 +/- 0.20 (P < 0.001); and comet tail length (microm) was 51.15 +/- 1.37 vs. 40.26 +/- 0.88 (P < 0.001). Similar significant differences were found when the groups were stratified by age and length of exposure. This study suggests that exposure to BMF smoke leads to greater levels of DNA damage than exposure to LPG combustion products. PMID- 15662702 TI - Cytotoxicant-induced trophoblast dysfunction and abnormal pregnancy outcomes: role of zinc and metallothionein. AB - Normal trophoblast function, including implantation, hormone production, and formation of the selectively permeable maternofetal barrier, is essential for the establishment and maintenance of the fetoplacental unit and proper fetal development. Maternal cytotoxicant exposure causes the destruction of these cells, especially the terminally differentiated syncytiotrophoblasts, and results in a myriad of poor pregnancy outcomes. These outcomes range from intrauterine growth retardation and malformation to spontaneous abortion or stillbirth. There is recent evidence that the metal-binding protein, metallothionein, is involved in the protection of human trophoblastic cells from heavy metal-induced and severe oxidative stress-induced apoptosis. Metallothionein, with its unique biochemical structure, can both bind essential metal ions, such as the transcription modulator zinc, and yet allow their ready displacement by toxic nonessential metal ions or damaging free radicals. These properties suggest that metallothionein may be responsible not only for sequestering the cytotoxic agents, but also for altering signal transduction in the affected cells. Here, we review several identified causes of adverse pregnancy outcomes (specifically, prenatal exposure to cigarette smoke and alcohol, gestational infection, and exposure to environmental contaminants), discuss the role of zinc in modulating the cellular response to these toxic insults, and then propose how metallothionein may function to mediate this protective response. PMID- 15662703 TI - "Atypical" cells in fine-needle aspiration biopsy specimens of benign thyroid cysts. AB - BACKGROUND: Cystic lesions of the thyroid are common. Most are benign nodules with degenerative changes in a multinodular goiter. Fine-needle aspiration biopsy (FNAB) specimens from these cystic nodules usually are easily interpreted as benign. However, occasionally, cells with atypical features are encountered, increasing the possibility of a cystic malignant neoplasm. To the authors' knowledge, the microscopic features of these benign cells, presumed to be of cyst lining origin, have not been well described to date. To refine the description of their morphologic features, with the belief that better recognition will avoid unnecessary surgery, the authors examined the cytologic and corresponding histologic features of thyroid cysts with "atypical" cells. METHODS: A total of 149 FNAB specimens from thyroid cysts containing atypical cells were identified. Seventy-five specimens with subsequent histologic correlation showing a benign cystic thyroid nodule were selected for study. The cytologic features of the atypical cyst-lining cells were reviewed and correlations were made with histologic, immunohistochemical, and clinical features. In addition, 12 FNAB specimens of histologically proven cystic papillary carcinoma diagnosed as atypical were reviewed for comparison. RESULTS: The majority of specimens (94%) were diagnosed cytologically as atypical thyroid cysts. However, in 29% of these specimens, a papillary or Hurthle cell neoplasm could not be excluded. The cytologic features of the atypical cells most often resembled classic reparative epithelial cells consistent with a cyst-lining origin. The most common features were cohesive flat sheets (84%), distinct cell borders (96%), nuclear enlargement (92%), nuclear grooves (79%), dense granular cytoplasm (79%), small distinct nucleoli (85%), fine chromatin (87%), and elongate to spindled cytomorphology (57%). In contrast to the atypical cells from benign cysts, cystic papillary carcinomas lacked the repair-like spindled cytomorphology, and showed nuclear crowding (100%), as well as papillary microarchitecture (50%), and rare intranuclear pseudoinclusions (42%). Histologically, the atypical cells in aspirate specimens corresponded to cyst-lining cells, which exhibited a spectrum ranging from flattened elongate cells with eosinophilic cytoplasm to more polygonal cells with abundant eosinophilic cytoplasm, enlarged irregular nuclei with pale chromatin, prominent nuclear grooves, and distinct nucleoli. Immunohistochemical staining of a subset of the resected thyroid cysts showed that the cyst-lining cells were positive for keratin and thyroglobulin, consistent with thyroid follicular cells. CONCLUSIONS: Atypical cyst-lining cells were found to have characteristic features (e.g., distinct cell borders, elongated shape, eosinophilic cytoplasm, and distinct nucleoli) and lacked nuclear crowding, intranuclear pseudoinclusions, and papillary architecture that, in many specimens, allowed them to be recognized as benign. The authors recommended that the subset of cells with the characteristic features described in the current study be reported as "consistent with benign cyst lining cells". PMID- 15662704 TI - Mechanisms for the environmental regulation of gene expression. AB - The environment can play a significant role in the production of phenotypes. However, the developmental mechanisms by which the environment can affect normal development are only now being elucidated. At least three paths have been found through which the environment can modify gene expression. The first is the neuroendocrine route, wherein the nervous system transmits signals from the environment to the endocrine system and the hormones alter gene expression. The second pathway involves environmental agents that change the methylation pattern of genes, thereby altering their transcriptional capacities. The third route involves the direct induction of gene expression in the host by its microbial symbionts. The normal environmental regulation of phenotype production should be considered a normal component of development and developmental biology. PMID- 15662705 TI - Response characteristics of the mitochondrial DNA genome in developmental health and disease. AB - This review focuses on mitochondrial biology in mammalian development; specifically, the dynamics of information transfer from nucleus to mitochondrion in the regulation of mitochondrial DNA genomic expression, and the reverse signaling of mitochondrion to nucleus as an adaptive response to the environment. Data from recent studies suggest that the capacity of embryonic cells to react to oxygenation involves a tradeoff between factors that influence prenatal growth/development and postnatal growth/function. For example, mitochondrial DNA replication and metabolic set points in nematodes may be determined by mitochondrial activity early in life. The mitochondrial drug PK11195, a ligand of the peripheral benzodiazepine receptor, has antiteratogenic and antidisease action in several developmental contexts in mice. Protein malnutrition during early life in rats can program mitochondrial DNA levels in adult tissues and, in humans, epidemiological data suggest an association between impaired fetal growth and insulin resistance. Taken together, these findings raise the provocative hypothesis that environmental programming of mitochondrial status during early life may be linked with diseases that manifest during adulthood. Genetic defects that affect mitochondrial function may involve the mitochondrial DNA genome directly (maternal inheritance) or indirectly (Mendelian inheritance) through nuclear-coded mitochondrial proteins. In a growing number of cases, the depletion of, or deletion in, mitochondrial DNA is seen to be secondary to mutation of key nuclear-coded mitochondrial proteins that affect mitochondrial DNA replication, expression, or stability. These defects of intergenomic regulation may disrupt the normal cross-talk or structural compartmentation of signals that ultimately regulate mitochondrial DNA integrity and copy number, leading to depletion of mitochondrial DNA. PMID- 15662706 TI - Investigations into the etiology of neural tube defects. AB - Neural tube defects (NTDs) are serious malformations affecting approximately 1 per 1000 births, yet the mechanisms by which they arise are unknown. There have been consistent efforts in many fields of research to elucidate the etiology of this multifactorial condition. While no single gene has been identified as a major independent risk factor for NTDs, candidate genes have been proposed that may modify the effects of maternal and/or embryonic exposures. Folate supplementation effectively reduces the occurrence of NTDs and, consequently, has focused much research on metabolism of folate-related pathways during pregnancy and development. Further understanding of normal development and how teratogens can perturb these orchestrated processes also remains at the fore of modern scientific endeavors. The composite of these factors remains fragmented; the aim of this review is to provide the reader with a summary of sentinel and current works in the body of literature addressing NTD disease etiology. PMID- 15662707 TI - Mechanisms regulating toxicant disposition to the embryo during early pregnancy: an interspecies comparison. AB - The dose of toxicant reaching the embryo is a critical determinant of developmental toxicity, and is likely to be a key factor responsible for interspecies variability in response to many test agents. This review compares the mechanisms regulating disposition of toxicants from the maternal circulation to the embryo during organogenesis in humans and the two species used predominantly in regulatory developmental toxicity testing, rats and rabbits. These three species utilize fundamentally different strategies for maternal embryonic exchange during early pregnancy. Early postimplantation rat embryos rely on the inverted visceral yolk sac placenta, which is in intimate contact with the uterine epithelium and is equipped with an extensive repertoire of transport mechanisms, such as pinocytosis, endocytosis, and specific transporter proteins. Also, the rat yolk sac completely surrounds the embryo, such that the fluid-filled exocoelom survives through most of the period of organogenesis, and can concentrate compounds such as certain weak acids due to pH differences between maternal blood and exocelomic fluid. The early postimplantation rabbit conceptus differs from the rat in that the yolk sac is not closely apposed to the uterus during early organogenesis and does not completely enclose the embryo until relatively later in development (approximately GD13). This suggests that the early rabbit yolk sac might be a relatively inefficient transporter, a conclusion supported by limited data with ethylene glycol and one of its predominant metabolites, glycolic acid, given to GD9 rabbits. In humans, maternal embryo exchange is thought to occur via the chorioallantoic placenta, although it has recently been conjectured that a supplemental route of transfer could occur via absorption into the yolk sac. Knowledge of the mechanisms underlying species specific embryonic disposition, factored together with other pharmacokinetic characteristics of the test compound and knowledge of critical periods of susceptibility, can be used on a case-by-case basis to make more accurate extrapolations of test animal data to the human. PMID- 15662708 TI - Pineal gland lesions: a cytopathologic study of 20 specimens. AB - BACKGROUND: Pineal gland lesions are rare, with only a few cytologic descriptions occurring in the literature, according to the authors' knowledge. The current article describes the cytopathologic characteristics of 20 such lesions with discussion of differential diagnoses. METHODS: Cytologic material was obtained either by fine-needle aspiration biopsy (FNAB) under stearotactic radiologic guidance or by touch imprinting (TI) at the time of frozen sectioning. The 20 specimens include pineoblastoma (five specimens), pineocytoma (four specimens), astrocytoma (three specimens), germ cell tumor (three specimens), meningioma (one specimen), epidermoid cyst (three specimens), and pineal cyst (one specimen). Smears were stained with Diff-Quik and with Papanicolaou and hematoxylin and eosin stains. In selected specimens, immunoperoxidase (IPOX) stains were performed on cell block sections using synaptophysin, neuron-specific enolase, placental alkaline phosphatase, glial fibrillary acidic protein, leukocyte common antigen, cytokeratins, and human chorionic gonadotropin antibodies. RESULTS: Several cytomorphologic characteristics unique to each lesional category with occasional overlapping features were observed. The unique features included the following: small, hyperchromatic, round to oval cells with frequent rosetting (pineocytoma), with a few specimens in addition showing hypercellularity, crowding, mitoses, and necrosis (pineoblastoma); pleomorphic round cells in a fibrillary background (astrocytoma); large polygonal cells with prominent nucleoli and clear cytoplasm (germ cell tumor); spindled fibroblastic cells (meningioma); anucleate squames and mature squamous cells (epidermoid cyst); and small uniform polygonal cells (pineal cyst). When necessary, IPOX studies supported the morphologic diagnoses. CONCLUSIONS: FNAB and TI cytology were found to provide a rapid and reliable diagnosis of pineal lesions. This is particularly important when dealing with minute amounts of tissue material. Both techniques appeared to provide equally good cytomorphology on smears. IPOX studies played an important complementary role in difficult cases when performed on cell blocks. PMID- 15662709 TI - Embryonic and fetal programming of physiological disorders in adulthood. AB - In the past decade, data from numerous epidemiological studies have indicated strong inverse associations between birth weight and risk of coronary heart disease, hypertension, type 2-diabetes, and other diseases in adulthood. The "Barker hypothesis" thus postulates that a number of organ structures and functions undergo programming during embryonic and fetal life. This developmental programming determines the set points of physiological and metabolic responses in adult life. Alterations of nutrient availability during gestation may lead to developmental adaptations, via hormonal maneuvers by the embryo and fetus that readjust these set points. These adaptive measures have short-term benefits to the embryo and fetus, so that the newborn will be better prepared for the adverse environment (e.g., undernutrition). However, adequate nutritional support during postnatal life that enables catch-up growth may create metabolic conflicts that predispose the adult to aberrant physiological functions and, ultimately, increased risk of disease. It is plausible that other adverse in utero conditions, including exposure to developmental toxicants, may similarly alter adult disease susceptibility. This article provides an overview of the Barker hypothesis, its supporting evidence, the current advances in understanding the biological mechanisms underlying this phenomenon, and its implications for developmental toxicology. PMID- 15662710 TI - Prenatal diagnosis of Fanconi anemia (Group C) subsequent to abnormal sonographic findings. AB - Manifestations of Fanconi Anemia Complementation Group C (FA-C) include multiple major congenital malformations, hypoplastic radius, absent thumb, growth retardation, elfin-like facial features, microphthalmia, microcephaly, cafe-au lait spots, early onset of hematologic disease and poor survival (Auerbach, 1997). We describe two cases in which second-trimester sonographic findings led to parental carrier testing for FA-C and subsequent prenatal diagnosis of affected fetuses. PMID- 15662711 TI - Diagnosis and outcome of fetal lower urinary tract obstruction in the northern region of England. AB - OBJECTIVE: We reviewed the prenatal and postnatal management of fetal lower urinary tract obstruction (LUTO) in a large geographically defined population. METHODS: The records of 113 cases of LUTO seen over a 14-year period were examined. The predictive accuracy of prenatal findings for chronic renal failure (CRF) and a comparison of prenatal-suspected and non-suspected cases were made. RESULTS: The incidence of LUTO was 2.2 in 10 000 births. During the study period, prenatal detection improved from 33 to 62%. Sensitivity of prenatal ultrasound detection of renal dysplasia and fetal urinary sodium, calcium, and beta2 microglobulin for CRF or renal dysplasia on autopsy were 59, 33, 66, and 63% respectively. Compared to undetected cases, those detected prenatally had higher mortality and a higher rate of CRF at 24 months (17% vs 57%, p < 0.01). CONCLUSION: Our observations confirm the poor prognosis associated with fetal LUTO. The value of serial fetal urine biochemistry, other prenatal predictors of postnatal renal function, and the benefits of vesicoamniotic shunting require larger series and longer follow-up. PMID- 15662712 TI - Timing of mid-trimester sacral ossification. PMID- 15662713 TI - Timing of mid-trimester sacral ossification: reply. PMID- 15662714 TI - Identification of Muir-Torre syndrome among patients with sebaceous tumors and keratoacanthomas: role of clinical features, microsatellite instability, and immunohistochemistry. AB - BACKGROUND: The Muir-Torre syndrome (MTS) is an autosomal-dominant genodermatosis characterized by the presence of sebaceous gland tumors, with or without keratoacanthomas, associated with visceral malignancies. A subset of patients with MTS is considered a variant of the hereditary nonpolyposis colorectal carcinoma, which is caused by mutations in mismatch-repair genes. The objective of the current study was to evaluate whether a combined clinical, immunohistochemical, and biomolecular approach could be useful for the identification of Muir-Torre syndrome among patients with a diagnosis of sebaceous tumors and keratoacanthomas. METHODS: The authors collected sebaceous skin lesions and keratoacanthomas recorded in the files of the Pathology Department of the University of Modena during the period 1986-2000. Through interviews and examination of clinical charts, family trees were drawn for 120 patients who were affected by these skin lesions. RESULTS: Seven patients also were affected by gastrointestinal tumors, thus meeting the clinical criteria for the diagnosis of MTS. In the MTS families, a wide phenotypic variability was evident, both in the spectrum of visceral tumors and in the type of skin lesions. Microsatellite instability was found in five MTS patients: These patients showed concordance with immunohistochemical analysis; moreover, a constitutional mutation in the MSH2 gene was found in 1 patient. Lack of expression of MSH2/MSH6 or MLH1 proteins was evident in the skin lesions and in the associated internal malignancies of 3 patients and 2 patients with MTS, respectively. CONCLUSIONS: The clinical, biomolecular, and immunohistochemical characterization of sebaceous skin lesions and keratoacanthomas may be used as screening for the identification of families at risk of MTS, a disease that is difficult to recognize and diagnose. PMID- 15662715 TI - Tracking vaccine lot lifecycles using reports to the vaccine adverse event reporting system (VAERS). AB - PURPOSE: There is currently no systematically available information available on how rapidly a specific lot of vaccine is used once distributed. We used data from reports to the Vaccine Adverse Event Reporting System (VAERS) to develop a proxy means of surveillance for the lifecycle of selected vaccine lots. METHODS: A convenience sample, consisting of selected lots of: diphtheria, tetanus, and acellular pertussis (DTaP), Haemophilus influenzae type b (Hib), Hepatitis B, and varicella vaccines, was selected for lifecycle analysis. Assuming that circulation of a vaccine lot is proportional to vaccine-specific adverse event (AE) reporting for that vaccine type, we constructed Gamma distributed usage models and compared them with lot-specific VAERS reports to estimate the actual lifecycle of lots in the system. RESULTS: Evidence of lot circulation was detected within 1-2 months, and a peak was observed 3-4 months after the vaccine release date for most of the study vaccines. Ninety percent of the vaccine doses in each lot were estimated to be used within 5-9 months of distribution. The length of time a vaccine lot was in use ranged from 5 to 17 months from earliest vaccination date. CONCLUSIONS: Our modeled and inferred administration of the selected lots of different vaccines were concordant. This method may be useful for spatial and temporal tracking of vaccine lot utilization. PMID- 15662716 TI - Quantifying levels of p53 mutation in mouse skin tumors. AB - Allele-specific competitive blocker PCR (ACB-PCR) amplification and quantification was developed for mouse p53 codon 270 CGT-->TGT base substitution and codon 244/245 AAC/CGC-->AAT/TGC tandem mutation. PCR products corresponding to p53 mutant and wild-type DNA sequences were generated. These DNAs were mixed in known proportions to construct samples with defined mutant fractions and the allele-specific detection of each mutation was systematically optimized. Each assay was used to analyze eight simulated solar light (SSL)-induced tumors. By analyzing mutant fraction (MF) standards in parallel with PCR products generated from tumor samples, p53 mutants could be quantified as subpopulations within the tumors. All eight tumors contained detectable levels of p53 codon 270 CGT-->TGT mutation. Three tumors had p53 MFs between 10(-4) and 10(-3). Five tumors had p53 MFs between 10(-3) and 10(-2). None of the eight mouse skin tumors had measurable levels of p53 codon 244/245 tandem mutation. Frequent detection of p53 codon 270 CGT-->TGT mutation provides additional evidence that a pyrimidine dinucleotide overlapping a methylated CpG site (Pyr(me)CG) is a susceptible target for SSL induced mutagenesis. The absence of p53 codon 244/245 mutation in tumors may be explained by its mutant p53 phenotype and/or indicate that this site is not methylated. These initial results indicate that p53 codon 270 CGT-->TGT mutation may be a sensitive biomarker for SSL- or UV-induced mutagenesis. This mutational endpoint may be useful for evaluating the co-carcinogenicity of compounds administered in combination with UV or SSL. PMID- 15662718 TI - Comparative actions of clomazone on beta-carotene levels and growth in rice (Oryza sativa) and watergrasses (Echinochloa spp). AB - Seedlings of rice, early watergrass (thiobencarb-resistant and thiobencarb susceptible biotypes, R and S, respectively), and late watergrass (thiobencarb resistant and thiobencarb-susceptible biotypes, R and S, respectively) were hydroponically exposed to clomazone at concentrations ranging from 0.08 to 7.9 microM. Whole-plant growth (mg fresh wt) and beta-carotene concentrations (microg g(-1) fresh wt) were measured after a 7-day exposure period. For growth, the no observed effect concentrations (NOECs) were 7.9, 0.21, 0.21, 0.46 and 0.46 microM clomazone for rice, early watergrass (R), early watergrass (S), late watergrass (R) and late watergrass (S), respectively, while the concentrations causing 25% inhibition in response (IC25) were 5.6 (+/-1.6), 0.46 (+/-0.06), 0.42 (+/-0.08), 0.92 (+/-0.45) and 0.79 (+/-0.08) microM clomazone, respectively. Clomazone inhibits beta-carotene synthesis via inhibition of the non-mevalonate isoprenoid synthetic pathway. For assessment of clomazone effects, beta-carotene levels proved to be a more sensitive toxicological endpoint than growth. For rice, early watergrass (R), early watergrass (S), late watergrass (R) and late watergrass (S), the beta-carotene NOECs were 0.21, <0.08, <0.08, 0.08 and 0.46 microM clomazone respectively, while IC25 values were 0.42 (+/-0.26), 0.08 (+/-0.02), 0.08 (+/-0.02), 0.33 (+/-0.09) and 0.54 (+/-0.15) microM, respectively. No evidence was found that the thiobencarb-resistance mechanisms present in early and late watergrasses impart resistance to clomazone. Due to similar sensitivity between rice and late watergrass, use of clomazone in rice culture will require the use of a safening technique. PMID- 15662717 TI - Nonrandom aneuploidy of chromosomes 1, 5, 6, 7, 8, 9, 11, 12, and 21 induced by the benzene metabolites hydroquinone and benzenetriol. AB - The loss and gain of whole chromosomes (aneuploidy) is common in the development of leukemia and other cancers. In acute myeloid leukemia, the loss (monosomy) of chromosomes 5 and 7 and the gain (trisomy) of chromosome 8 are common clonal chromosomal abnormalities. Here, we have tested the hypothesis that metabolites of the human leukemogen benzene cause a higher rate of gain and loss among the chromosomes involved in leukemogenesis and, as such, are nonrandom and selective in their effects. Human peripheral blood was exposed to two metabolites of benzene, namely, hydroquinone (HQ) and benzenetriol (BT), and the ploidy status of nine different chromosomes (1, 5, 6, 7, 8, 9, 11, 12, and 21) was examined using fluorescence in situ hybridization of metaphase spreads. Poisson regression was used to provide interpretable incidence rate ratios and corresponding P values for all nine chromosomes. Statistically significant differences were found between the sensitivity of the nine chromosomes to gain or loss. Chromosomes 5 and 7 were highly sensitive to loss following HQ and BT exposure, whereas chromosomes 7, 8, and 21 were highly sensitive to gain in comparison to other chromosomes. Significant support for the a priori hypothesis that chromosomes 5 and 7 are more sensitive to loss induced by HQ and BT than the other seven chromosomes was also obtained. These data support the notion that benzene metabolites affect the ploidy status of specific chromosomes more than others and may initiate or promote leukemia induction through these specific effects. PMID- 15662719 TI - Endogenous estrogen status, but not genistein supplementation, modulates 7,12 dimethylbenz[a]anthracene-induced mutation in the liver cII gene of transgenic big blue rats. AB - A growing number of studies suggest that isoflavones found in soybeans have estrogenic activity and may safely alleviate the symptoms of menopause. One of these isoflavones, genistein, is commonly used by postmenopausal women as an alternative to hormone replacement therapy. Although sex hormones have been implicated as an important risk factor for the development of hepatocellular carcinoma, there are limited data on the potential effects of the estrogens, including phytoestrogens, on chemical mutagenesis in liver. Because of the association between mutation induction and the carcinogenesis process, we investigated whether endogenous estrogen and supplemental genistein affect 7,12 dimethylbenz[a]anthracene (DMBA)-induced mutagenesis in rat liver. Intact and ovariectomized female Big Blue rats were treated with 80 mg DMBA/kg body weight. Some of the rats also received a supplement of 1,000 ppm genistein. Sixteen weeks after the carcinogen treatment, the rats were sacrificed, their livers were removed, and mutant frequencies (MFs) and types of mutations were determined in the liver cII gene. DMBA significantly increased the MFs in liver for both the intact and ovariectomized rats. While there was no significant difference in MF between the ovariectomized and intact control animals, the mutation induction by DMBA in the ovariectomized groups was significantly higher than that in the intact groups. Dietary genistein did not alter these responses. Molecular analysis of the mutants showed that DMBA induced chemical-specific types of mutations in the liver cII gene. These results suggest that endogenous ovarian hormones have an inhibitory effect on liver mutagenesis by DMBA, whereas dietary genistein does not modulate spontaneous or DMBA-induced mutagenesis in either intact or ovariectomized rats. PMID- 15662720 TI - Glyphosate-resistant crops: history, status and future. AB - The commercial launch of glyphosate-resistant soybeans in 1996 signaled the beginning of a new era in weed management in row crops. Today, over 80% of the soybeans grown in the USA are glyphosate resistant. Since that time, many crops have been transformed that have allowed crop applications of many classes of herbicide chemistries. Crops currently under production include maize, soybean, cotton and canola. Transformation technology and selection methods have improved and the rate of development as well as the breadth of crops being considered as commercial targets has increased. On the basis of recent adoption rates by growers around the world, it appears that glyphosate-resistant crops will continue to grow in number and in hectares planted. However, global public acceptance of biotechnology-derived products will continue to impact the rate of adoption of this and other new innovations derived from transformation technology. PMID- 15662721 TI - Control of blue mold (Penicillium expansum) by fludioxonil in apples (cv Empire) under controlled atmosphere and cold storage conditions. AB - A reduced risk fungicide, fludioxonil, was tested for its efficacy against blue mold caused by thiabendazole-resistant and -sensitive Penicillium expansum (Link) Thom in apples under three storage conditions. In a co-treatment, fludioxonil and inoculum were applied together to test the protective activity of the fungicide on wounds that had been aged for 1 or 2 days. The fungicide was also tested for its curative activity in post-inoculation treatment on apples that had been inoculated for 1 or 2 days. Fludioxonil was very effective as co-treatment and as post-inoculation treatment. At a concentration of 300 mg litre(-1), fludioxonil gave complete control of post-harvest blue mold caused by the thiabendazole resistant and -sensitive P expansum for 105 days in controlled atmosphere (CA) storage at 2 (+/-1) degrees C, for 42 days in common cold storage at 4 (+/-1) degrees C and also in a shelf-life study for 6 days at 20 (+/-1) degrees C. Comparison on the effect of fludioxonil in CA storage and common cold storage showed that higher concentrations of fungicide were needed in cold storage than in CA storage. Fludioxonil at a concentration of 450 mg litre(-1), gave 98 and 92% control of blue mold of apples in the simulated shelf-life studies after CA and common cold storages, respectively. Fludioxonil has a potential to be incorporated in the fungicide resistance management strategies for control of blue mold in apples stored for 105 days. PMID- 15662722 TI - What it takes to get a herbicide's mode of action. Physionomics, a classical approach in a new complexion. AB - Discovering new herbicides with novel modes of action is a priority assignment in plant protection research. However, for active compounds identified in greenhouse screens, the crucial point is to tread the most efficient path in determining a herbicide's target site, regarding chance of success, time and research costs. Today, in the literature, molecular (functional genomics, transcriptomics), biochemical (proteomics) and analytical (metabolomics) approaches are particularly discussed. So far, less attention has been focused on the comprehensive physiological profiling of the complex plant system as a procedure which enables new herbicides, with an unknown target site for their mode of action, to be screened rapidly. Here, the concept of an array of 'functional' bioassays is presented which has ultimately been developed from the classical tool of mode of action diagnosis by symptoms. These bioassays are designed to differentiate between the distinct responses of the multiple organization units (plant, tissue, meristematic cell, organelle), developmental stages, types of metabolism (phototrophic, heterotrophic) and physiological processes in the plant organism. The response pattern to a herbicide can be viewed as the end result of changes induced in the molecular and biochemical process chain and should be diagnostic of its physiological mode of action. The results can be interpreted directly or a fingerprint database for all known modes of action to be screened for analogy. The term 'physionomics' is proposed for this comprehensive physiological profiling of the plant system, following the parallel terminology of the molecular and biochemical 'omics' technologies. Physionomics procedures provide a first clue to the mode of action of a new herbicide that can direct more time-consuming and costly molecular, biochemical, histochemical or analytical studies to identify a target site more efficiently. PMID- 15662723 TI - Technical performance of some commercial glyphosate-resistant crops. AB - Glyphosate-resistant (GR) crops have been sold commercially in the USA since 1996. The use of glyphosate alone or with conventional pre- and post-emergence herbicides with different modes of action gives growers many options for affordable, safe, easy, effective wide-spectrum weed control. Despite the overwhelming popularity of this technology, technical issues have surfaced from time to time as US growers adopt these crops for use on their farms. The types of concern raised by growers vary from year to year depending on the crop and the environment, but include perceptions of increased sensitivity to diseases, increased fruit abortion, reduced pollination efficiency, increased sensitivity to environmental stress, and differences in yield and agronomic characteristics between transgenic and sister conventional varieties. Although several glyphosate resistant crops are commercially available, maize, soybean and cotton constitute the largest cultivated acreage and have likewise been associated with the highest number of technical concerns. Because glyphosate is rapidly translocated to and accumulates in metabolic sink tissues, reproductive tissues and roots are particularly vulnerable. Increased sensitivity to glyphosate in reproductive tissues has been documented in both glyphosate-resistant cotton and maize, and results in reduced pollen production and viability, or increased fruit abortion. Glyphosate treatments have the potential to affect relationships between the GR crop, plant pathogens, plant pests and symbiotic micro-organisms, although management practices can also have a large impact. Despite these potential technical concerns, this technology remains popular, and is a highly useful tool for weed control in modern crop production. PMID- 15662724 TI - Haplotype sharing transmission/disequilibrium tests that allow for genotyping errors. AB - The present study introduces new Haplotype Sharing Transmission/Disequilibrium Tests (HS-TDTs) that allow for random genotyping errors. We evaluate the type I error rate and power of the new proposed tests under a variety of scenarios and perform a power comparison among the proposed tests, the HS-TDT and the single marker TDT. The results indicate that the HS-TDT shows a significant increase in type I error when applied to data in which either Mendelian inconsistent trios are removed or Mendelian inconsistent markers are treated as missing genotypes, and the magnitude of the type I error increases both with an increase in sample size and with an increase in genotyping error rate. The results also show that a simple strategy, that is, merging each rare haplotype to a most similar common haplotype, can control the type I error inflation for a wide range of genotyping error rates, and after merging rare haplotypes, the power of the test is very similar to that without merging the rare haplotypes. Therefore, we conclude that a simple strategy may make the HS-TDT robust to genotyping errors. Our simulation results also show that this strategy may also be applicable to other haplotype based TDTs. PMID- 15662725 TI - The motility of glioblastoma tumour cells is modulated by intracellular cofilin expression in a concentration-dependent manner. AB - The invasive behaviour of tumour cells has been attributed in part to dysregulated cell motility. Members of the ADF/Cofilin family of actin-binding proteins are known to increase microfilament dynamics by increasing the rate at which actin monomers leave the pointed end of the filament and by a filament severing activity. As depolymerisation is a rate-limiting step in actin dynamics, ADF/Cofilins are suspected to facilitate the motility of cells. To test this, we investigated the influence of cofilin on tumour motility by transient and stably overexpressing cofilin in the human glioblastoma cell line, U373 MG. Several different methods were used to ascertain the level of cofilin in overexpressing clones and this was correlated with their rate of random locomotion. A biphasic relationship between cofilin level and locomotory rate was found. Clones that displayed a moderate amount of overproduction of cofilin were found to have increased rates of locomotion approximately linear to the overproduction of cofilin up to an optimal cofilin level of about 4.5 times that of wild type cells at which the cells were almost twice as fast. However, clones producing more than this optimal amount were found to locomote at progressively reduced speeds. Cells that overexpress cofilin have reduced stress fibres compared to control cells showing that the excess cofilin affects the actin cytoskeleton. We conclude that overexpression of cofilin enhances the motility of glioblastoma tumour cells in a concentration-dependent fashion, which is likely to contribute to their invasiveness. PMID- 15662726 TI - Calcium transients regulate titin organization during myofibrillogenesis. AB - Titin has a Ca2+-dependent kinase domain and may act as a molecular template for myofibrillogenesis. Therefore, we examined the relationship between endogenous Ca2+ transients and titin organization in embryonic myocytes. When transients were blocked during sarcomere assembly, titin organization was disrupted. Titin was distributed in punctate aggregates on an otherwise diffuse background, resulting in a 66% decrease in organization. Myosin, as reported previously, was also disrupted in a similar manner (75% decrease). In titin-actin-myosin triple labeling experiments, myosin and titin were highly colocalized, although titin aggregates without significant myosin accumulation were also observed. This suggests that myosin-titin association is not dependent on Ca2+ transients, although terminal aspects of titin-myosin organization require transients. We also examined whether titin organization is dependent on actin filament dynamics. The data indicate that (1) the normal sarcomeric arrangement of titin depends on Ca2+ transients, (2) titin-myosin association does not require Ca2+ transients, and (3) titin filament organization does not depend on barbed-end actin dynamics. PMID- 15662727 TI - RNAi knockdown of the focal adhesion protein TES reveals its role in actin stress fibre organisation. AB - TES was originally identified as a candidate tumour suppressor gene and has subsequently been found to encode a novel focal adhesion protein. As well as localising to cell-matrix adhesions, TES localises to cell-cell contacts and to actin stress fibres. TES interacts with a variety of cytoskeletal proteins including zyxin, mena, VASP, talin and actin. There is evidence that TES may function in actin-dependent processes as overexpression of TES results in increased cell spreading and decreased cell motility. Together with TES's interacting partners, these data suggest that TES might be involved in regulation of the actin cytoskeleton. Here, for the first time, we have used RNAi to successfully knockdown TES in HeLa cells and we demonstrate that loss of TES from focal adhesions results in loss of actin stress fibres. Similarly, and as previously reported, RNAi-mediated knockdown of zyxin results in loss of actin stress fibres. TES siRNA treated cells show reduced RhoA activity, suggesting that the Rho GTPase pathway may be involved in the TES RNAi-induced loss of stress fibres. We have also used RNAi to examine the requirement of TES and zyxin for each other's localisation at focal adhesions, and we propose a hierarchy of recruitment, with zyxin being first, followed by VASP and then TES. Cell Motil. PMID- 15662730 TI - Nomenclature for factors of the HLA system, update May 2004. PMID- 15662728 TI - Inflammation and life-span. PMID- 15662731 TI - Challenges and changes. PMID- 15662732 TI - [Reactivation in immunosuppression. Complicated hepatitis comeback]. PMID- 15662733 TI - 'Knock, knock'... Fashion Robin to the rescue. PMID- 15662734 TI - [Studies of the relationship of HLA polymorphisms and the infection of H. pylori in the population of Linqu in Shandong Province]. AB - In order to analyze the relationship of HLA polymorphisms and the infection of H. pylori in the population of Linqu County in Shandong Province, polymerase chain reaction with sequence-specific primers(PCR-SSP) was used to determine the alleles of HLA type I and II in 90 Hp-positive persons and 49 Hp-negative controls. The results showed that among the 68 alleles of HLA type I ,4 alleles were found significantly different between Hp-positive and Hp-negative population,while no significant difference was found among the 22 alleles of HLA type UI. Hp-positive persons had a lower allele frequency of A* 02 (OR=0.56, 95% CI=0.33 approximately 0. 94; P=0.029), B* 48 (OR=0.15, 95% CI=0.03 approximately 0.72; P=0. 007), CW* 08 (OR=0. 32, 95% CI=0.15 approximately 0.69; P=0. 003) and a higher allele frequency of CW* 15 (OR=5.11, 95% CI=0.63 approximately 40.90; P=0. 024) compared with Hp-negative controls. Our results indicated that the polymorphisms of HLA type I is involved in the genetic susceptibility of Hp infection in Linqu County,while the polymorphisms of HLA type I may have no relationship with the genetic susceptibility of Hp infection. It was shown that among the alleles of HLA type I , CW* 15 might be a susceptible gene of Hp infection while A * 02 ,B * 48 and CW* 08 might be protective genes. PMID- 15662735 TI - Anesthesia during intralesional bare fiber laser treatment of a giant hemangioma of head and neck region of an infant using the laryngeal mask airway. PMID- 15662736 TI - [The genetic variation of two microsatellite markers of Escherichia coli F4(K88) receptor in different swine breeds]. AB - The genetic variation of ETEC F4 receptor in Shaziling and Yorkshire breeds were studied using two micro-satellite markers(S0223 and S0068). The results showed that there were polymorphisms in the two markers, and there were great variations of the gene heterozygosity and Shannon information index in the two breeds. It was also reported that there were differences in K88ab and K88ac receptors in Chinese native breeds and foreign breeds, so the two markers might be the genetic markers of F4 receptor gene. PMID- 15662737 TI - [Association between genetic factor and physical performance]. AB - Correlation study between physical performance and gene has become a hot topic. Evidences have indicated that human capacity is partially influenced by genetic factors. Since 2000, the genetic studies of locomotion have progressed rapidly. More than 70 genetics markers associated with physical performance have been studied, such as VO(2max) (maximal oxygen uptake), muscle strength and lactate threshold. This research could be used to detect the special individual in spaceflight project and national defense project. Obviously, genetic detection of the special individual at early stage will save time and fund as compared with traditional training system. The present paper is to summarize some major international studies on aerobic exercise related genes and loci. We expect that the present article may provide helpful information to promote the project in China. PMID- 15662738 TI - Footnote added to National Patient Safety Goal 7, Requirement 7A, for behavioral health care. PMID- 15662739 TI - [Two approaches of quantitative-trait linkage analysis]. AB - ln this article, we discussed two model-free methods for detecting genetic linkage for quantitative traits, Haseman-Elston regression approach and variance components approach. The former is a regression approach for detecting linkage based on the squared difference or squared sums in quantitative trait values of sib-pairs and their estimated marker IBD scores. The latter can jointly model covariate effects along with variance components, including genetic component and non-genetic sources of variability. We have outlined the model assumption, the algorithm and the extensions for the both methods. PMID- 15662740 TI - [Strengthening study on influenza and face to the new challenge of influenza]. PMID- 15662741 TI - Has the DoH lost its clout? PMID- 15662742 TI - Working lives. Primary care: crossing the great divide. Interview by Nick Edwards. PMID- 15662743 TI - HSJ management challenge. The Stepford vibes. PMID- 15662745 TI - Working lives. Making drama out of a crisis. Interview by Marie-Anne Hamilton. PMID- 15662746 TI - Challenges to deliver therapies. AB - Medical science and pharmaceutical technology have progressed in enormous leaps. The professions have access to knowledge of improved therapies that cover a large selection of diagnoses. This state of affairs is a successful result of the long and purposeful co-operation between science and technology, theory and practice. The therapies have changed but their delivery to the patient remains the same. During the recent international recession and the continuous cost containment programmes the question of availability has risen to the fore. Availability is a question of means. That is usually interpreted as equal to saving and cost cutting. At times of affluence it could mean allowance but that seems far in the past. Availability also concerns the physical distribution of the therapies such as pharmaceuticals. So far, advances in the science have not affected either the distribution or availability of pharmaceutical products. With the rise in patient awareness, pressure is rising to make full use of the research results. This is a natural, albeit unmet, demand and represents an opportunity for the pharmacy institutions with their well trained personnel, modern outlets and the unique as well as traditional position in direct contact with the patients. This role could gain added weight through the progress of the technologies that allow ambulatory therapies instead of cost consuming institutionalisation. It would appear quite natural that the providers of such services would seek the pharmacy outlet networks as their proper points of delivery. There is no doubt that this choice would be taken as a convenient solution by the consumers of the services. This would offer considerable savings to society and efficiency to the patients. PMID- 15662744 TI - Behavioral graded activity following first-time lumbar disc surgery: 1-year results of a randomized clinical trial. AB - STUDY DESIGN AND OBJECTIVES: In a randomized clinical trial, the effectiveness of behavioral graded activity was assessed as compared to usual care provided by physiotherapists for patients after first-time lumbar disc surgery (n = 105). SUMMARY OF BACKGROUND DATA: Little is known about the effectiveness of rehabilitation programs following lumbar disc surgery. Most programs focus on biomechanical aspects, whereas psychosocial factors are hardly addressed. The aim of the behavioral graded activity program, which is an operant treatment, is to alter psycho-social factors such as fear of movement and pain catastrophizing, which might subsequently lead to improved functional status and higher rates of recovery. Behavioral treatments for patients following lumbar disc surgery have not yet been assessed in a randomized clinical trial. METHODS: INCLUSION CRITERIA: age between 18 and 65 years; first-time lumbar disc surgery; restrictions in normal activities of daily living. EXCLUSION CRITERIA: surgical complications and confirmed and relevant underlying diseases. Outcome assessment took place at 6 and 12 months after randomization. RESULTS: Six months after randomization, 62% of the patients had recovered following usual care versus 65% of the patients following behavioral graded activity. After 12 months, 73% and 75%, respectively, had recovered. Differences between intervention groups, 3% and 2% respectively, after 6 and 12 months are not statistically significant. Furthermore, there were no differences between the two groups regarding functional status, pain, pain catastrophizing, fear of movement, range of motion, general health, social functioning or return to work. After 1 year, 4 of the behavioral graded activity cases had undergone another operation versus 2 of usual care cases. CONCLUSION: Both fear of movement and pain catastrophizing seem to be unaffected by either treatment in these patients. It is concluded that treatment principles derived from theories within the field of chronic low back pain might not apply to these patients. After 1 year of follow-up, there were no statistically significant or clinically relevant differences between the behavioral graded activity program and usual care as provided by physiotherapists for patients following first-time lumbar disc surgery. PMID- 15662747 TI - In memory of Dr Hirotsugu Shiraki (1917-2004). PMID- 15662748 TI - [118th Internal Medicine Seminar and the 27th Vanysek Seminar organized by the Czech and Slovak Internal Medicine Society, the Association of Czech Physicians in Brno and the Czech Oncological Society on the anniversary of the birth of Dr. Rudolf Vanysek, founder of the Brno School of Internal Medicine. Friday, 2 March 2001 in Brno. Oncology of Solid Tumors in Internal Medicine]. PMID- 15662749 TI - [Comment on: Myocardial infarct and malignant diseases]. PMID- 15662750 TI - [Mucormycosis]. AB - Over recent years the clinical importance of invasive fungal infections has significantly increased. Mucormycosis refers to a rare, life threatening, opportunistic disease caused by filamentous fungi of the order Mucorales. Most frequently mucormycosis occurs in patients with ketoacidotic diabetes and in neutropenic patients with acute leukaemia. This study of fifteen patients was conducted to evaluate the clinical characteristics of invasive mucormycosis, and goal was also to propose for an early diagnosis and management as survival can only be improved by aggressive surgery and antifungal therapy. In a retrospective analysis of the biopsy and autopsy records of the Department of Pathology at Szent Laszlo Hospital between 1992-2003 were evaluated fifteen histologically documented cases of mucormycosis. The underlying diseases were hematological malignancy with neutropenia in ten cases, chronic leukaemia with neutropenia, chronic leukaemia with latent diabetes mellitus and diabetes mellitus one case each. Three cases did not belong to any typical risk group. An in vivo diagnosis was made only in four patients with a rhinocerebral manifestation. Radical necrectomy and long-term antifungal therapy cured two of them. Two patients died of a generalised infection despite therapy. None of the systemic infections could be cured. The diagnosis was made post mortem in eleven patients. Selected clinical forms of mucormycosis are illustrated with short case reports. PMID- 15662751 TI - [Risk and prevention in cardiovascular diseases]. AB - The RCTs are the most important sources of drug therapy related scientific evidence (RCT--Randomised Controlled clinical Trials). In most cases RCTs studying identical or similar clinical issues are slightly or considerably varied. However, for such cases we have the appropriate methodology to compare the more or less different RCT results. This publication was written to provide a general overview of this methodology such as relative and absolute risk reduction (RRR, ARR), odd ratio (OR), number needed to treat and number needed to harm (NNT, NNH). PMID- 15662752 TI - [Cytokines, prostaglandins, nutritive and non-nuitritive factors in inflammatory bowel diseases]. AB - Therapeutic interventions in the case of gastrointestinal disease are based on the understanding of the role of different inflammatory mediators. Reactive O2 and N2 metabolites are involved in IBD. Pro-inflammatory cytokines, apoptosis signalling and redox-response transcription factors are depended on free radicals. NO activates COX enzymes. PGE2 negatively modulates induction of NO synthase by interleukins and therefore regulation of gastric mucosal integrity by endogenous NO depends on arachidonic acid cascade. PG-s have pro-inflammatory and anti-inflammatory effects on the immune system. Dietary PUFA-s and eicosanoids have potential effects on the modulation of inflammatory processes and immune cells. The cholesterol level lowering activity of several cytokines and colony stimulating factor can be observed. Therapeutic efficacy of N-3 PUFA is described in cases of patients with chronic gastrointestinal disorders, but N-3 PUFA-s only delay early relapse of ulcerative colitis in remission. TNF is known as a pleiotropic cytokine. Strategies for TNF in IBD is very important part of therapeutical approaches. Therapy with infliximab and related ones are encouraging in critical cases. It is also believed recently, that NF-kappaB also may be a target of IBD treatment. It became known, that oxidized LDL can inhibit LPS-induced binding of the NF-kappaB to DNA and the subsequent expression of TNF alpha and interleukin-1beta in macrophages as well as oxidized LDL modulates activation of NF-kappaB in mononuclear phagocytes by altering the degradation of I-kappaBs. 15-d-PGJ2 inhibits multiple steps in the NF-kappaB signaling pathway. 15-d-PGJ2 metabolite binds PPAR-gamma promotes adipocyte differentiation. PPAR gamma ligand inhibits growth of cells through induction of apoptosis. Several nutritional polyphenols (the secondary metabolites of plants) are COX2 and/or LOX inhibitors and iNOS activators. The moderate nutritional customs with natural antioxidants can help restore to normal function of gastrointestinal tract, but the immoderate consumption of vitamins and polyphenol type antioxidant molecules is contraindicated. PMID- 15662754 TI - [An interesting and short-lived periodical in the country-side]. PMID- 15662753 TI - [Treatment of primary mediastinal large B-cell lymphomas]. AB - INTRODUCTION: Primary mediastinal large B-cell non-Hodgkin's lymphoma is a relatively rare disease with specific clinical symptoms. This tumour originates from a subset of B-cells of the thymus and at the time of the diagnosis the disease is predominantly localised in the mediastinum. The tumor grows rapidly and frequently involves other thoracic structures. The majority of the patients are young females. There are no histologic features that reliably distinguish these tumors from other diffuse large B-cell lymphomas. This is the only lymphoma subtype which can only be defined by the combination of clinical and pathologic features. Analysis with DNA microarrays verified that primary mediastinal and diffuse large B-cell lymphomas are different diseases. AIMS: Comparing the effectiveness of two types of anthracycline-based standard chemotherapy regimens and the evaluation of the prognostic markers which are applied in large B-cell lymphomas. METHODS: 27 patients with primary mediastinal lymphoma were treated by the authors with anthracycline-based polychemotherapy with complementary radiotherapy from January 1995 to December 2002. RESULTS: Complete remission was obtained in 15 patients (56%) and no relapse was observed in this group. 9 additional patients (33%) achieved partial remission, while in 3 cases (11%) the treatment was ineffective. The patients who failed to achieve complete remission were subsequently treated with more intensive chemotherapy. Afterwards, those patients who were chemosensitive, underwent high-dose chemotherapy with autologous peripheral blood stem-cell transplantation. The chemoresistant patients received palliative chemotherapy. The 5-year overall survival rate of the 27 patients was 62.11%. CONCLUSION: The authors found that the procarbazine, prednisolone, adriamycin, cyclophosphamide, etoposide, cytosine-arabinoside, bleomycin, vincristine, methotrexate treatment was more effective than the cyclophosphamide, adriamycin, vincristine, prednisolone combination. The expected 5-year overall survival rates were 83.57% vs. 33.33%, respectively. This difference was significant (p = 0.017). No prognostic value of age adjusted international prognostic index, LDH- and b2-microglobulin levels were found. The results with the new standard of combined immuno-chemotherapy (rituximab- cyclophosphamide, adriamycin, vincristine, prednisolone) seem to be hopeful and more effective than earlier treatments. PMID- 15662756 TI - Legal corner. Nurse Practice Act. PMID- 15662755 TI - [Under the torrid zone. To the memory of Dr. Laszlora Scheitz]. PMID- 15662757 TI - What to say...when managing difficult individuals. PMID- 15662760 TI - Nurses--defenders of environmental health: Mercury. PMID- 15662758 TI - Healthy way to lose weight. PMID- 15662761 TI - Ethical issues--clinical research. PMID- 15662762 TI - Lactose intolerance. AB - Although lactose intolerance is very common it is not a serious health condition. The diagnosis is relatively simple and minimally invasive. Treatment is geared towards a life-long plan of management. Persons who have difficulty digesting lactose will learn by trial and error what food items cause distress and learn to avoid offending milk sugars. In addition many products are available over-the counter to aid in digestion of lactose. Often these additives enable the person to consume lactose. Adequate amounts of calcium may be consumed by eating a carefully chosen diet containing lactose free sources of calcium in order to maintain healthy bone, nerve, and muscle development. PMID- 15662763 TI - Colorectal cancer. PMID- 15662764 TI - [Alarming symptoms in vertebrobasilar circulatory disorders. Part I]. AB - Dizziness and vertigo--like headache--are the most common complaints which leads patients to visit the doctor. In spite of the headache--which may be primary (e.g. migraine) or symptomatic--dizziness and vertigo do not appear to be a separate nosologic entity but rather the symptoms of several neurological disorders. For differential diagnosis, interdisciplinary thinking and activity is needed because the vestibular, neurological and psychiatric disorders might have a common role in the development of symptoms and further overlapping can also occur. The vascular disorders of the vertebrobasilar system are discussed in detail in this review. The importance, occurrence and causes of vertigo as a warning symptom is in the focus. The author draws attention to life-threatening conditions with acute onset in cases of the posterior scale ischemia and emphasizes the importance of the correct and early diagnosis. The author tries to clear up the nihilistic aspect in treating of stroke and stresses the necessity of thrombolysis and interventional radiological procedures which may be the only chance for the recovery of the patients. The pharmacological prevention of recurrent vascular events is also important and obligatory for the clinicians. PMID- 15662765 TI - [New chapter in the study of visual evoked potentials--clinical application of the multifocal VEP method]. AB - The multifocal visual evoked potentials are the evoked responses over the visual cortex in response to the stimulation of circumscribed small areas in the central 30 degree region of the retina. The recording of multifocal visual evoked potentials was made possible by the computer algorhythm elaborated by Sutter in 1991. Multifocal electroretinography, developed upon the same theoretical principles, is already an routine clinical examination method for the topographic analysis of functional damages in the central part of the retina and for the differential diagnostics in neuro-ophthalmology. The multifocal visual evoked potential, however, has not been introduced into the clinical practice, although it displays the function of ganglion cells in a given region of the retina in a more detailed way than the sensitivity threshold in the perimetry. This examination makes the objective verification of defects possible in the visual pathway, too. In our department the recording of multifocal visual evoked potentials was started in 2002. In this paper we present the basics of this method and also deal with the problems concerning its application and its status in the history of visual evoked potentials. PMID- 15662766 TI - The impact of neuroleptic medication on seizure threshold and duration in electroconvulsive therapy. AB - INTRODUCTION: In most patients diagnosed with psychotic depression or schizophrenia and treated with electroconvulsive therapy, parallelly administered antipsychotic medication cannot be stopped. Antipsychotic drugs can influence both seizure threshold and seizure activity in different ways. PATIENTS AND METHOD: The present study processes the data of 77 patients treated parallelly with electroconvulsive therapy and antipsychotic drugs. Oral doses of the antipsychotic medication administered the day before the electroconvulsive therapy, stimulus intensity, seizure durations, and impedance were analysed from session to session. RESULTS: One group of antipsychotics (haloperidol, fluphenazine, risperidone, sulpirid) was not found to influence seizure activity: there was no significant difference in EEG and EMG registered seizure duration or in stimulus intensity between the treated and non-treated group. However, significant difference was found between the next treated and non-treated groups in 40% of the sessions in case of olanzapine, in 50% of the sessions in case of clozapine and in 57% of the sessions in case of zuclopenthixol in EEG or EMG registered seizure duration as well as in stimulus intensity. In the third group (quetiapine) there was a significant difference in each session (2nd session: EMG, p=0.02; 5th session: EEG, p=0.05, EMG, p=0.04). Most of the antipsychotics (olanzapine, clozapine, zuclopenthixol) have been shown to possess epileptogenic properties; only quetiapine reduces seizure activity. CONCLUSION: In the clinical use of olanzapine, clozopine and zuclopenthixol seems epileptogenic, whereas in the case of quetiapine seizure reducing properties must be taken into account. Together with the consideration of the accompanying somatic and neurologic disturbances and with the concomitant medications this can influence the treatment of choice. PMID- 15662767 TI - Temozolomide chemotherapy of patients with recurrent anaplastic astrocytomas and glioblastomas. AB - INTRODUCTION: Anaplastic astrocytomas and glioblastomas are the most frequent and most malignant hemispherial tumours. Unfortunately, astrocytic tumours are of infiltrative character and radical removal is not possible. Recurrent malignant gliomas are rarely suitable for reoperation. In most of the cases of recurrent gliomas chemotherapy is the last choice. PATIENTS AND METHOD: Seventy-five consecutive patients with recurrent malignant astrocytomas and glioblastomas had been treated at our institute with per os temozolomide for five days every month. The patients received two to 16 courses of chemotherapy. The toxicity, quality of life, response to chemotherapy and survival data were analysed. RESULTS: Out of 75 patients four were excluded following the first treatment due to myelotoxicity, and allergic reactions. Among the patients treated with temozolomide in seven cases complete response, 17 partial response, 14 progressive disease were observed. In 33 cases the disease stabilized and out of them in 27% a significant neurological improvement was detected. The time to progression was 6.8 months and the median survival time 8.75 months for patients with glioblastoma and with malignant astrocytoma or malignant mixed oligoastrocytoma 9.45 and 11.15 months, respectively. The overall survival for patients with originally lower grade glioma was 70.32 and for patients with glioblastoma multiforme 17.43 months. CONCLUSIONS: Temozolomide chemotherapy in patients with recurrent malignant astrocytoma and glioblastoma proved to be efficacious and similar good results were achieved as with a nitrosourea based combined chemotherapy. Even in those patients who received previous chemotherapy temozolomide is well tolerated and a relatively long time to progression was achieved in cases of recurrent malignant gliomas. In a few number of patients where BCNU had been previously failed with temozolomide stable disease was achieved. Temozolomide seems to be a promising drug in the chemotherapy of malignant gliomas and can be applied as a second line chemotherapy, as well. PMID- 15662768 TI - [Immunomodulatory therapy in multiple sclerosis]. AB - During the past decade, several disease-modifying agents have been established and have become available for the treatment of multiple sclerosis. The disease modifying agents could be grouped into immunomodulatory and immunosuppressive therapies altering the long-term course of multiple sclerosis. Therapy is now available for relapsing-remitting, secondary progressive and progressive relapsing multiple sclerosis. Different disease-modifying agents became also available for the treatment of relapsing-remitting multiple sclerosis in Hungary which makes the therapeutic decision difficult. This overview might help to give an answer for different questions in the management of multiple sclerosis: Which agent to choose? When to initiate the therapy? Which dose to apply? Are the drugs safe? How long to treat the patients with immunomodulatory drugs? We give a review from the literature to assess the efficacy of disease-modifying therapies and to compare the data from phase three trials of interferon beta1b, two preparations of interferon beta1a or glatiramer acetate for the treatment of multiple sclerosis. We analyzed the efficacy and safety of these agents on physical, inflammatory and cognitive measures of disease activity. Comparison of study results indicated similar effects of immunomodulatory agents on relapse related and inflammatory measures in relapsing multiple sclerosis. Interferon beta1a slowed the progression of disability in relapsing multiple sclerosis. One interferon beta1a preparation (intramuscularly injected) demonstrated efficacy in slowing progression of cognitive dysfunction. The interferons reduced relapses at early phase of secondary progressive multiple sclerosis, but their efficacy have not yet been proven in the later phase of secondary progressive multiple sclerosis without relapses. Mitoxantrone demonstrated efficacy in slowing the progression of disability in secondary progressive multiple sclerosis. All of the disease modifying agents are safe and tolerable, if the indication is correct and the patients are strictly controlled. PMID- 15662769 TI - [Early onset dementias: three cases]. AB - INTRODUCTION: Dementia is a decline of intellectual abilities. The etiology of dementia syndrome is diverse. The authors describe three patients with early onset dementia. CASE REPORTS: The first patient was a 44 years old male with mild gait, body ataxia, memory loss, slowness and apathy Investigations proved AIDS dementia syndrome. In the second case of a 37 years old female patient, herpes simplex encephalitis was suspected due to sudden onset of speech arrest and to brain MRI and CSF findings. Her symptoms improved during antiviral treatment but later progressive dementia developed. CSF serological tests proved the presence of neurolues-dementia paralytica. The third patient was a 38-years-old female. Neurological examination was performed because of progressive memory loss, changed behaviour and impaired attention. Neuropsychological test showed severe dementia. Metachromatic leukodystrophy was proven by decreased arylsulfatase activity. CONCLUSIONS: It is not easy to recognize the early symptoms of dementia. In these cases, besides detailed history, neurological examination and neuropsychological tests, brain MRI and cerebral spinal fluid serological tests were indispensable for a correct diagnosis, especially in the young patients. PMID- 15662770 TI - Mechanisms of edema after gamma knife surgery for meningiomas. Report of two cases. AB - The authors describe two patients in whom tumor swelling and brain swelling (and possible tumor swelling), respectively, developed after undergoing gamma knife surgery. One had a skull defect with a palpable parasagittal tumor. One had neurofibromatosis Type 2 with multiple tumors, one of which was parasagittal. PMID- 15662771 TI - A simple method for predicting imaging-based complications following gamma knife surgery for cerebral arteriovenous malformations. AB - OBJECT: The authors studied the relationship between dose planning parameters and complications in the treatment of cerebral arteriovenous malformations (AVMs). METHODS: There were 41 continuous unselected patients. The mean follow-up period was 19 months; the mean age was 28 years; the male/female ratio was 2.2:1.0; the median prescription dose was 25 Gy (range 14-25 Gy); the median prescription isodose was 50%. The median lesion volume was 4.4 cm3. The median lesion coverage was 93%; and the mean conformity index was 1.22. The authors found no relationship between lesion volume or integral dose and the development of the clinical effects based on the adverse radiation effects (AREs); however, there was a significant relationship between both target volume and integral dose with the development of AREs as well as the severity of the AREs. CONCLUSIONS: The integral dose could be used as a guideline for the prescription dose. Arguments are made for maximizing the prescription dose for the long-term safety of the patient. PMID- 15662772 TI - Does new magnetic resonance imaging technology provide better geometrical accuracy during stereotactic imaging? AB - OBJECT: The authors sought to compare the accuracy of stereotactic target imaging using the Siemens 1T EXPERT and 1.5T SYMPHONY magnetic resonance (MR) units. METHODS: A water-filled cylindrical Perspex phantom with axial and coronal inserts containing grids of glass rods was fixed in the Leksell stereotactic frame and subjected to MR imaging in Siemens 1T EXPERT and Siemens 1.5T SYMPHONY units. Identical sequences were used for each unit. The images were transferred to the GammaPlan treatment planning system. Deviations between stereotactic coordinates based on MR images and estimated real geometrical positions given by the construction of the phantom insert were evaluated for each study. The deviations were further investigated as a function of the MR unit used, MR sequence, the image orientation, and the spatial position of measured points in the investigated volume. CONCLUSIONS: Larger distortions were observed when using the SYMPHONY 1.5T unit than those with the EXPERT 1T unit. Typical average distortion in EXPERT 1T was not more than 0.6 mm and 0.9 mm for axial and coronal images, respectively. Typical mean distortion for SYMPHONY 1.5T was not more than 1 mm and 1.3 mm for axial and coronal images, respectively. The image sequence affected the distortions in both units. Coronal T2-weighted spin-echo images performed in subthalamic imaging produced the largest distortions of 2.6 mm and 3 mm in the EXPERT 1T and SYMPHONY 1.5T, respectively. Larger distortions were observed in coronal slices than in axial slices in both units, and this effect was more pronounced in SYMPHONY 1.5T. Noncentrally located slice positions in the investigated volume of the phantom were associated with larger distortions. PMID- 15662773 TI - Computational model for the estimation of the extracranial doses received during Leksell gamma knife model C treatment. AB - OBJECT: Extracranial doses received by patients undergoing Leksell gamma knife surgery (GKS) can be of clinical concern. Therefore, the ability to preestimate peripheral doses received outside the treatment field during the GKS would be beneficial and could be used for the optimization of treatment planning by providing a reference for practitioners to calculate the extracranial dose burden to the body before the start of treatment. METHODS: A dose of 40 Gy was delivered to the Rando phantom in a single fraction to a midline hypothetical target close to the center of the skull. Treatment planning was performed for multiple isocenters with a prescription to the 50% isodose of each collimator. Treatment plans were produced for 1, 5, 10, 15, and 20 shots, keeping the same dose and dose distribution at different target volumes. An automatic positioning system was used for positioning the phantom during the treatment. The doses to different organs were measured during GKS using a thermoluminescent dosimeter. In vivo measurements were also made in 200 patients who underwent GKS with the model C unit for different diagnoses at Na Homolce Hospital. CONCLUSIONS: The peripheral dose depended on the collimator size with a logarithmic dependence on collimator size and a linear dependence on the number of shots. This model can be used for the estimation of peripheral doses with a total error less than 20%. This information can help clinicians with treatment planning optimization, especially in patients with long survival expectancy. PMID- 15662774 TI - Low-grade gliomas treated by fractionated gamma knife surgery. AB - OBJECT: The authors sought to evaluate local tumor control, complications, and progression-free survival in patients harboring low-grade gliomas who were treated with Leksell gamma knife surgery (GKS). METHODS: During a 6-year period 70 patients were treated for verified low-grade gliomas (Grade I or II) by GKS. Statistical analysis was based on 68 patients; two patients were lost to follow up. The median patient age was 17 years. The median target volume was 4200 mm.3 The median prescription dose was 25 Gy. The median number of fractions was five. Ninety-five percent of patients were treated in five daily fractions. Partial or complete tumor regression was achieved in 83% of patients with a median time to response of 18 months. There was moderate acute or late toxicity in not more than 5% of patients. In this series the progression-free survival was 92% at 3 years and 88% at 5 years. CONCLUSIONS: Relatively high local tumor control with minimal complications was achieved. PMID- 15662775 TI - A comparison of the gamma knife model C and the automatic positioning system with Leksell model B. AB - OBJECT: The authors sought to compare the quality of treatment planning, radiation protection, and the time taken for treatment in the Leksell gamma knife model B with that using the model C Automatic Positioning System (APS). METHODS: Data were obtained in 463 patients treated with the B model and 518 patients treated with the C model. Data were analyzed in patients in whom the following diagnoses had been made: vestibular schwannoma, pituitary adenoma, meningioma, solitary metastasis, and other benign and malignant solitary tumors. Patients with arteriovenous malformations, ocular lesions, and functional diagnoses were excluded from this study. CONCLUSIONS: With the C model there was a better conformity for most treated targets, such as vestibular schwannomas (p = 0.005) and meningiomas (p = 0.015). The level of radiation exposures to personnel was significantly decreased when using the model C (p < 0.001). There was no significant difference in radiation exposure of extracranial structures for the same number of shots in patients treated by both models. The mean time saved using the C model with the APS was 41 minutes per treatment. It would seem that the gamma knife model C permits better dose conformity, shorter treatment times, and less radiation exposure to personnel. PMID- 15662776 TI - Treatment of essential trigeminal neuralgia with gamma knife surgery. AB - OBJECT: The authors present the long-term follow-up results (minimum 5 years) of patients with essential trigeminal neuralgia (TN) who were treated with gamma knife surgery (GKS). METHODS: One hundred seven patients (61 females and 46 males) underwent GKS. The median follow up was time was 60 months (range 12-96 months). The target was the trigeminal root, and the maximum dose was 70 to 80 Gy. Repeated GKS was performed in 19 patients for recurrent pain, and the same dose was used. Initial successful results were achieved in 96% of patients, with complete pain relief in 80.4%. Relief was achieved after a median latency of 3 months (range 1 day-13 months). Gamma knife surgery failed in 4% of patients. Pain recurred in 25% of patients after a median latent interval of 36 months (6 94 months). The initial success rate after a second GKS was 89% and 58% of patients were pain free. Pain relapse occurred in only one patient in this group. Hypesthesia was observed in 20% of patients after the first GKS and in 32% after the second GKS. The median interval to hypaesthesia was 35 months (range 3-94 months) after one treatment and 21 months (range 1-72 months) after a second treatment. CONCLUSIONS: The initial success rate of pain relief was high and comparable to that reported in other studies. A higher than usual incidence of sensory impairment after GKS could be the long duration of follow-up study and due to the detailed neurological examination. PMID- 15662777 TI - Long-term complications after gamma knife surgery for arteriovenous malformations. AB - OBJECT: The authors analyzed of the long-term complications that occur 2 or more years after gamma knife surgery (GKS) for intracranial arteriovenous malformations (AVMs). METHODS: Patients with previously untreated intracranial AVMs that were managed by GKS and followed for at least 2 years after treatment were selected for analysis (237 cases). Complete AVM obliteration was attained in 130 cases (54.9%), and incomplete obliteration in 107 cases (45.1%). Long-term complications were observed in 22 patients (9.3%). These complications included hemorrhage (eight cases), delayed cyst formation (eight cases), increase of seizure frequency (four cases), and middle cerebral artery stenosis and increased white matter signal intensity on T2-weighted magnetic resonance imaging (one case of each). The long-term complications were associated with larger nidus volume (p < 0.001) and a lobar location of the AVM (p < 0.01). Delayed hemorrhage was associated only with incomplete obliteration of the nidus (p < 0.05). Partial obliteration conveyed no benefit. Delayed cyst formation was associated with a higher maximal GKS dose (p < 0.001), larger nidus volume (p < 0.001), complete nidus obliteration (p < 0.01), and a lobar location of the AVM (p < 0.05). CONCLUSIONS: Incomplete obliteration of the nidus is the most important factor associated with delayed hemorrhagic complications. Partial obliteration does not seem to reduce the risk of hemorrhage. Complete obliteration can be complicated by delayed cyst formation, especially if high maximal treatment doses have been administered. PMID- 15662778 TI - Gamma knife surgery of the pituitary: new treatment for thalamic pain syndrome. AB - OBJECT: Although reports in the literature indicate that thalamic pain syndrome can be controlled with chemical hypophysectomy, this procedure is associated with transient diabetes insipidus. It was considered reasonable to attempt gamma knife surgery (GKS) to the pituitary gland to control thalamic pain. METHODS: Inclusion criteria in this study were poststroke thalamic pain, failure of all other treatments, intolerance to general anesthetic, and the main complaint of pain and not numbness. Seventeen patients met these criteria and were treated with GKS to the pituitary. The target was the pituitary gland together with the border between the pituitary stalk and the gland. The maximum dose was 140 to 180 Gy. All patients were followed for more than 3 months. CONCLUSIONS: An initial significant pain reduction was observed in 13 (76.5%) of 17 patients. Some patients experienced pain reduction within 48 hours of treatment. Persistent pain relief for more than 1 year was observed in five (38.5%) of 13 patients. Rapid recurrence of pain in fewer than 3 months was observed in four (30.8%) of 13 patients. The only complication was transient diabetes insipidus in one patient. It would seem that GKS of the pituitary might have a role to play in thalamic pain arising after a stroke. PMID- 15662779 TI - The effect of gamma knife irradiation on functions of striatum in rats. AB - OBJECT: An animal model has been developed to study the effect of gamma knife surgery(GKS) on cerebral function. METHODS: A rat was fixed in a newly developed Regis-Valliccioni frame that enables the target region to be planned directly on the magnetic resonance images. The left striatum was irradiated with 150 Gy via a 4-mm collimator of the Leksell gamma knife. Apomorphine (dopamine agonist) was administered to elicit a circling behavior (apomorphine test) after the GKS so as to examine the time course of the changes in dopaminergic functions of irradiated striatum. After a series of behavioral analyses, irradiated brains were subjected to histological examination. Necrosis was observed in the irradiated area surrounded by hemorrhage and gliosis. The distance between the histologically estimated and planned centers of the irradiation areas was 1.0 +/- 0.5 mm. The extent of the distance was due to errors along dorsoventral axis. The distribution of the irradiation areas influenced the activity and the circling behaviors in apomorphine test, which was suggestive of involvement of the nigrostriatal pathway. CONCLUSIONS: Targeting by using the Regis-Valliccioni frame was very accurate compared with targeting with coordinates based on brain maps used hitherto. Although targeting improved the accuracy, further effort will still be necessary to reduce errors along dorsoventral axis. The apomorphine test indicated a reduced dopaminergic function of the irradiated area including striatum, which accompanied histological changes after a high dose of irradiation (150 Gy). PMID- 15662780 TI - Gamma knife surgery for arteriovenous malformations involving the corpus callosum. AB - OBJECT: The purpose of this study was to evaluate the safety and efficacy of gamma knife surgery (GKS) for the treatment of arteriovenous malformations (AVMs) involving the corpus callosum. METHODS: Thirty-two patients aged from 7 to 65 years (median 25 years) with AVMs of the corpus callosum underwent GKS between 1990 and 2002. The maximum AVM diameter was more than 3 cm in 11 patients (34%). The AVM volume ranged from 0.1 to 19.1 cm3 (median 1.6 cm3). The median dose to the AVM margin was 20 Gy (range 17-28 Gy). Patients were followed for 1 to 12 years (median 9 years). The angiographically confirmed actuarial obliteration rate was 64% and 74% at 4 and 6 years, respectively. Younger patient age (p < 0.05) and lower radiosurgery-based grading score (calculated from the patient age and AVM volume; p < 0.01) were the significant factors affecting successful AVM obliteration. No patient suffered a hemorrhage after GKS, although 28 patients (88%) had a history of hemorrhage from their AVMs. Radiation-induced neurological deficit was observed only in one patient (3%) who had undergone previous radiotherapy (50 Gy). No patient experienced complications of occlusion or stenosis of the normal vascular structures adjacent to the AVM. CONCLUSIONS: Gamma knife surgery is a safe and effective treatment for selected patients with AVMs involving the corpus callosum, and it carries a low risk of damaging adjacent critical vascular structures. Even ruptured AVMs with relatively large diameter can be successfully treated, especially in younger patients, with minimal morbidity and a low risk of repeated hemorrhage. PMID- 15662782 TI - Histopathological findings in a surgically resected thalamic cavernous hemangioma 1 year after 40-Gy irradiation. AB - Stereotactic radiosurgery is a controversial treatment modality in the management of cerebral cavernous hemangiomas (CHs), and results vary from center to center. Even the interpretation of treatment failure is controversial. It is suggested that the systematic pathological investigation of irradiated specimens could help to resolve the controversy. A hemorrhagic lesion in the posterior part of the thalamus had been diagnosed as a tumor and was treated with 40-Gy fractionated radiotherapy. One year after this treatment the case was reconsidered based on new imaging evidence, and the lesion was removed by conventional craniotomy. Histopathological examination revealed a CH with postirradiation changes. Compared with nonirradiated control CH tissue samples, there was endothelial cell destruction and marked fibrosis with scar tissue formation in the stroma of the treated lesion. The histopathological findings in this specimen were similar to those described in arteriovenous malformations after gamma knife surgery. The results of light microscopic investigations suggest that the ionizing effect of radiation energy evokes vascular and connective tissue stroma changes in CHs as well. PMID- 15662781 TI - Gamma knife surgery for hypothalamic hamartomas accompanied by medically intractable epilepsy and precocious puberty: experience in Mexico. AB - OBJECT: Hypothalamic hamartoma is a nonneoplastic malformative mass of neurons and glia in the region of the hypothalamus. Because of its location, open surgery is associated with high morbidity and mortality rates. Gamma knife surgery (GKS) may be an efficient and safe treatment approach, which produces little morbidity. The authors describe the results of GKS in three patients with hypothalamic hamartomas. METHODS: All patients were male, aged 3, 12, and 15 years. The lesions were classified according to the Valdueza scale: one was Type IIb and two were Type IIa. The patients presented with gelastic seizures (15-20 per day), generalized epilepsy, behavioral abnormalities, and alterations of the sleep cycle. Precocious puberty was present in one patient. The Type IIb tumor had a volume of 1.8 cm3, and the Type IIa tumors were 597 mm3 and 530.1 mm3. The lesions received 12.5 Gy, 14 Gy, and 15 Gy, respectively, to the 50% isodose line. The patients were followed for 30 to 50 months. After 3 months, all patients showed improvement of their sleep, behavior, and epilepsy. At the present time, these patients are receiving low-dose antiepileptic agents and have achieved adequate social development and school integration. CONCLUSIONS: Gamma knife surgery appears to be a good, safe, and effective option for the treatment of selected hypothalamic hamartomas. No morbidity or mortality was associated with these three cases. PMID- 15662783 TI - Relative roles of microsurgery and stereotactic radiosurgery for the treatment of patients with cranial meningiomas: a single-surgeon 4-year integrated experience with both modalities. AB - OBJECT: The authors sought to assess the respective roles of microsurgery and gamma knife surgery (GKS) in the treatment of patients with meningiomas. METHODS: The authors culled from a 4-year prospective database data on 74 cases of meningiomas. Thirty-eight were treated with GKS and 35 with microsurgery. Simpson Grade 1 or 2 resection was achieved in 86.1 % of patients who underwent microsurgery. Patients who underwent GKS received a mean margin dose of 16.4 Gy (range 14-20 Gy). The mean tumor coverage was 94.7%, and the mean conformity index was 1.76. Significant differences between the two treatment groups (GKS compared with microsurgery) included age (mean 60 compared with 50.7 years), volume (mean 7.85 cm3 compared with 44.4 cm3), treatment history (55.3% compared with 14.3%), and tumor location (cavernous sinus/petroclival, 14 compared with three). The median follow up was 21.5 months (range 1.5-50 months). In patients with benign meningiomas GKS tumor control was 96.8% with one recurrence at the margin. The recurrence rate was zero of 27 for Simpson Grade 1 or 2 resection and three of four for higher grades in those patients who underwent microsurgery. There was no procedure-related mortality or permanent major neurological morbidity. The mean Karnofsky Performance Scale score was maintained for both forms of treatment. Symptoms improved in 48.4% of patients undergoing microsurgery and 16.7% of those who underwent GKS. Transient and permanent cranial nerve morbidity was 7.9 compared with 2.9%, and 5.3 compared with 8.5% for GKS and microsurgery, respectively. In a patient satisfaction survey 93.1% of microsurgery patients and 91.2% of GKS patients were highly satisfied. CONCLUSIONS: Both GKS and microsurgery serve important roles in the overall management of patients with meningiomas. Both are safe and effective and provide high degrees of satisfaction when used for differentially selected patients. PMID- 15662784 TI - Radiosurgery and the prevention of regrowth of incompletely removed nonfunctioning pituitary adenomas. AB - OBJECT: The authors studied the efficacy of gamma knife radiosurgery (GKS) in the prevention of regrowth of nonfunctioning pituitary adenomas (NPA). METHODS: One hundred nineteen patients were included in this study and were divided into two groups. All patients had undergone surgery in our department and recurrent or residual adenoma was demonstrated on postoperative MR imaging. Group A consisted of 68 patients who were followed without additional treatment. Group B was composed of 51 patients who received GKS within 1 year after microsurgery. There was no significant demographic difference between the two groups. In Group B the mean margin dose was 16.5 +/- 0.3 Gy (range 13-21 Gy). Fifty one and one tenth percent of patients in Group A were recurrence free at 5 years and 89.8% in Group B (p < 0.001). In Group B patients, tumor volume decreased from a baseline value of 2.4 +/- 0.2 cm3 to 1.6 +/- 0.2 cm3 at last follow up (p < 0.001). CONCLUSIONS: The results of this study suggest that GKS is effective in controlling growth of residual NPA for at least 5 years following initial maximal surgical debulking compared with no radiation therapy. Thus, GKS is recommended after microsurgery when visible tumor can be detected on imaging studies. PMID- 15662785 TI - Analysis of long-term outcomes and prognostic factors in patients with non-small cell lung cancer brain metastases treated by gamma knife radiosurgery. AB - OBJECT: The authors conducted a study to evaluate the long-term outcomes and prognostic factors for survival in a large series of patients treated by gamma knife surgery (GKS) for non-small cell lung cancer (NSCLC) brain metastases. METHODS: The study is based on the retrospective analysis of clinical and radiological records obtained during a 10-year period (1993-2003), concerning 836 lesions in 504 patients. The lesions were primary in 86% and recurrent 14% of the cases; they were solitary in 31%, single in 29%, and multiple in 40%. The mean follow-up period was 16 months (range 4-113 months). The most common histological types were adenocarcinoma (51%) and squamous cell carcinoma (27%). Dose planning parameters were as follows: mean target volume 6.2 cm3 (range 0.06-22.5 cm3); mean prescription dose 21.4 Gy (range 15.5-28 Gy); and mean number of isocenters 6.7 (range one-18). Progression-free and actuarial survival curves were calculated using the Kaplan-Meier method. The main factors affecting survival were determined by unimultivariate analysis (log-rank test and Cox proportional hazard models). Analysis of long-term outcomes seemed to confirm that GKS is a primary therapeutic option in these patients. The 1-year local tumor control rate was 94%. The overall median survival was 14.5 months, with extremely rewarding quality of life indices. The recursive partitioning analysis classification was the dominant prognostic factor. CONCLUSIONS: Gamma knife surgery is a useful treatment for brain metastases from NSCLC. PMID- 15662786 TI - Gamma knife surgery for cavernous hemangiomas: an analysis of 125 patients. AB - OBJECT: The authors sought to determine the value of gamma knife surgery (GKS) in the treatment of cavernous hemangiomas (CHs). METHODS: Between 1993 and 2002, a total of 125 patients with symptomatic CHs were treated with GKS. Ninety-seven patients presented with bleeding and 45 of these had at least two bleeding episodes. Thirteen patients presented with seizures combined with hemorrhage, and 15 patients presented with seizures alone. The mean margin dose of radiation was 12.1 Gy and the mean follow-up time was 5.4 years. In the 112 patients who had bled the number of rebleeds after GKS was 32. These rebleeds were defined both clinically and based on magnetic resonance imaging for an annual rebleeding rate of 32 episodes/492 patient-years or 6.5%. Twenty-three of the 32 rebleeding episodes occurred within 2 years after GKS. Nine episodes occurred after 2 years; thus, the annual rebleeding rate after GKS was 10.3% for the first 2 years and 3.3% thereafter (p = 0.0038). In the 45 patients with at least two bleeding episodes before GKS, the rebleeding rate dropped from 29.2% (55 episodes/188 patient-years) before treatment to 5% (10 episodes/197 patient-years) after treatment (p < 0.0001). Among the 28 patients who presented with seizures, 15 (53%) had good outcomes (Engel Grades I and II). In this study of 125 patients, symptomatic radiation-induced complications developed in only three patients. CONCLUSIONS: Gamma knife surgery can effectively reduce the rebleeding rate after the first symptomatic hemorrhage in patients with CH. In addition, GKS may be useful in reducing the severity of seizures in patients with CH. PMID- 15662787 TI - Gamma knife surgery for vestibular schwannoma: 10-year experience of 195 cases. AB - OBJECT: The authors conducted a study to determine the optimal radiation dose for vestibular schwannoma (VS) and to examine the histopathology in cases of treatment failure for better understanding of the effects of irradiation. METHODS: A retrospective study was performed of 195 patients with VS; there were 113 female and 82 male patients whose mean age was 51 years (range 11-82 years). Seventy-two patients (37%) had undergone partial or total excision of their tumor prior to gamma knife surgery (GKS). The mean tumor volume was 4.1 cm3 (range 0.04 23.1 cm3). Multiisocenter dose planning placed a prescription dose of 11 to 18.2 Gy on the 50 to 94% isodose located at the tumor margin. Clinical and magnetic resonance (MR) imaging follow-up evaluations were performed every 6 months. A loss of central enhancement was demonstrated on MR imaging in 69.5% of the patients. At the latest MR imaging assessment decreased or stable tumor volume was demonstrated in 93.6% of the patients. During a median follow-up period of 31 months resection was avoided in 96.8% of cases. Uncontrolled tumor swelling was noted in five patients at 3.5, 17, 24, 33, and 62 months after GKS, respectively. Twelve of 20 patients retained serviceable hearing. Two patients experienced a temporary facial palsy. Two patients developed a new trigeminal neuralgia. There was no treatment-related death. Histopathological examination of specimens in three cases (one at 62 months after GKS) revealed a long-lasting radiation effect on vessels inside the tumor. CONCLUSIONS: Radiosurgery had a long-term radiation effect on VSs for up to 5 years. A margin 12-Gy dose with homogeneous distribution is effective in preventing tumor progression, while posing no serious threat to normal cranial nerve function. PMID- 15662788 TI - Gamma knife surgery for multiple hemangioblastomas. AB - OBJECT: The authors describe their experience in treating patients with hemangioblastoma, especially multiple lesions, with gamma knife surgery (GKS). METHODS: Nine patients with 84 hemangioblastomas underwent GKS between July 1992 and May 2003. Three patients harbored a single lesion and six patients had multiple lesions. Of the six patients with multifocal tumors, a diagnosis of von Hippel-Lindau disease had been established in five. In the patients with multiple lesions, the mean radiation dose delivered to the tumor margin was 16.6 Gy (range 12.8-29.75 Gy). The mean margin isodose was 60% (range 40-95%). Three of the 84 lesions failed to be controlled after a mean follow-up period of 4.3 years (range 8.6-141 months). One patient who had undergone two GKS treatments suffered delayed radiation-induced complications, and posterior fossa decompression and ventriculoperitoneal shunt insertion were required. CONCLUSIONS: To achieve tumor control and avoid morbidity, the surgeon should keep in mind minimizing field overlapping by using a small-diameter collimator or applying a steep dose gradient, and by accurate dose prescription. PMID- 15662789 TI - Gamma knife surgery for intracranial cavernous hemangioma. AB - OBJECT: The purpose of this study was to assess the benefits of radiosurgery for cavernous hemangioma. METHODS: Sixty-five cavernous hemangiomas were treated with gamma knife surgery (GKS) between October 1994 and December 2002. Forty-two patients attended follow up. The mean patient age was 37.6 years (range 7-60 years). The lesions were located in the frontal lobe in 12 cases, deep in the parietal lobe in five, in the basal ganglia in five, in the temporal in three, in the cerebellum in three, in the pons/midbrain in six, and in multiple locations in eight cases. The presenting symptoms were seizure in 12, hemorrhage in 11, and other in 19. The maximum dose was 26.78 Gy, and the mean margin dose was 14.55 Gy. The mean follow-up period after radiosurgery was 29.6 months (range 5-93 months). The tumor decreased in size in 29 cases, was unchanged in 12, and increased in size in one. In the seizure group, seizures were controlled without anticonvulsant medication in nine cases (81.8%) after 31.3 months (range 12-80 months). After 93 months, one patient developed a cyst, which was resected. Rebleeding occurred in one case (2.3%). On T2-weighted imaging changes were seen in 11 cases (26.2%), in three (7.1 %) of which neurological deterioration was correlated with imaging changes. In other cases these deficits were temporary. CONCLUSIONS: The authors found that GKS was an effective treatment modality for cavernous hemangiomas, especially for those located within the brainstem, basal ganglia, or deep portions of the brain. It can reduce seizure frequency significantly although this takes time. In the group receiving a marginal dose below 15 Gy the patients fared better than when the dose exceeded 15 Gy. PMID- 15662791 TI - Low-dose radiosurgery for large vestibular schwannomas: long-term results of functional preservation. AB - OBJECT: The author conducted a study to assess the long-term results obtained in patients who underwent GKS for large vestibular schwannomas (> 3 cm in diameter). Facial and cochlear nerve functions were evaluated. METHODS: Twenty consecutive large tumors in 18 patients (including two cases of neurofibromatosus Type 2 [NF2]) were followed for more than 6 years. There were eight tumors that were more than 4 cm in maximum diameter. Microsurgery had already been performed prior to GKS in 11 patients (nine recurrent and two residual tumors). Four patients (including one with NF2) died during the follow-up period of other diseases or by accident. Fourteen of 15 tumors were stable or decreased in size. Microsurgery was performed in one patient 2 years after radiosurgery. Facial nerve function was preserved in all patients and hearing preserved in four of five patients with cochlear nerve function prior to radiosurgery. No adverse effects of radiosurgery have been observed to date. CONCLUSIONS: Gamma knife surgery seems to have a place in the low-dose treatment of selected large vestibular schwannoma in patients with a reasonable chance of retaining facial function and pretreatment hearing level. Patients with severe brainstem compression should first be undergo microsurgery. PMID- 15662790 TI - Targeting the cranial nerve: microradiosurgery for trigeminal neuralgia with CISS and 3D-flash MR imaging sequences. AB - OBJECT: The authors undertook a study to identify magnetic resonance (MR) imaging techniques that can be used reliably during gamma knife surgery (GKS) to identify the trigeminal nerve, surrounding vasculature, and areas of compression. METHODS: Preoperative visualization of the trigeminal nerve and surrounding vasculature as well as targeting the area of vascular compression may increase the effectiveness of GKS for trigeminal neuralgia. During the past years our gamma knife centers have researched different MR imaging sequences with regard to their ability to visualize cranial nerves and vascular structures. Constructive interference in steady-state (CISS) fusion imaging with three-dimensional gradient echo sequences (3D-Flash) was found to be of greatest value in the authors' 25 most recent patients. In 24 (96%) out of the 25 patients, the fifth cranial nerve, surrounding vessels, and areas of compression could be reliably identified using CISS/3D-Flash. The MR images were acceptable despite patients' history of microvascular decompression, radiofrequency (RF) ablation, or concomitant disease. In one of 25 patients with a history of multiple RF lesions, the visualization was inadequate due to severe trigeminal nerve atrophy. CONCLUSIONS: The CISS/3D-Flash fusion imaging has become the preferred imaging method at the authors' institutions during GKS for trigeminal neuralgia. It affords the best visualization of the trigeminal nerve, surrounding vasculature, and the precise location of vascular compression. PMID- 15662792 TI - Outpatient gamma knife surgery for vestibular schwannoma: definition of the therapeutic profile based on a 10-year experience. AB - OBJECT: The purpose of the study was to define the therapeutic profile of outpatient gamma knife surgery (GKS) for vestibular schwannoma (VS) by using sequential tumor volumetry to quantify changes following treatment. METHODS: A total of 111 patients met the inclusion criteria. The median follow-up duration was 7 years (range 5-9.6 years). Thirty-seven patients (33%) had undergone surgery before GKS and 10 (9%) had neurofibromatosis Type 2 (NF2). The median VS volume was 1.6 cm3 (range 0.08-8.7 cm3). The actuarial 6-year tumor control rate after a single GKS treatment was 95%. Tumor swelling was observed in 43 patients (38.7%). Recurrence was significantly associated with NF2 (p < 0.003) and the reduced dose (p < 0.03) delivered to these tumors. The incidence of facial nerve neuropathy was mainly determined by surgery prior to GKS (p < 0.0001). Facial nerve radiation toxicity was mild and transient. No permanent facial nerve toxicity was observed. Trigeminal neuropathy occurred in 13 patients, and this was correlated with the VS volume (p < 0.02). The median hearing loss was -10 dB (range + 20 dB to -70 dB). The risk of hearing loss was correlated with age and transient tumor swelling (p < 0.05) but not with dose parameters or NF2. CONCLUSIONS: Outpatient GKS is feasible, effective, and safe. Its therapeutic profile compares favorably with that of microsurgery. PMID- 15662793 TI - Long-term results of gamma knife surgery for growth hormone-producing pituitary adenoma: is the disease difficult to cure? AB - OBJECT: The authors conducted a study to determine the long-term results of gamma knife surgery for residual or recurrent growth hormine (GH)-producing pituitary adenomas and to compare the results with those after treatment of other pituitary adenomas. METHODS: The series consisted of 67 patients. The mean tumor diameter was 19.2 mm and volume was 5.4 cm3. The mean maximum dose was 35.3 Gy and the mean margin dose was 18.9 Gy. The mean follow-up duration was 63.3 months (range 13-142 months). The tumor resolution rate was 2%, the response rate 68.3%, and the control rate 100%. Growth hormone normalization (GH < 1.0 ng/ml) was found in 4.8%, nearly normal (< 2.0 ng/ml) in 11.9%, significantly decreased (< 5.0 ng/ml) in 23.8%, decreased in 21.4%, unchanged in 21.4%, and increased in 16.7%. Serum insulin-like growth factor (IGF)-1 was significantly decreased (IGF-1 < 400 ng/ml) in 40.7%, decreased in 29.6%, unchanged in 18.5%, and increased in 11.1%, which was almost parallel to the GH changes. CONCLUSIONS: Gamma knife surgery was effective and safe for the control of tumors; however, normalization of GH and IGF-1 secretion was difficult to achieve in cases with large tumors and low-dose radiation. Gamma knife radiosurgery is thus indicated for small tumors after surgery or medication therapy when a relatively high-dose radiation is required. PMID- 15662794 TI - Late cyst formation following gamma knife surgery of arteriovenous malformations. AB - OBJECT: The authors present data concerning the development of cysts following gamma knife surgery (GKS) in 1203 consecutive patients with arteriovenous malformations (AVMs) treated by the senior author (L.S.). The cyst was defined as a fluid-filled cavity at the site of a treated AVM. Cases involving regions corresponding to previous hematoma cavities were excluded. The incidence of cyst formation was assessed using magnetic resonance imaging studies performed in 196 cases with more than 10 years of follow up, in 332 cases with 5 to 10 years of follow up, and in 675 cases with less than 5 years of follow up. One hundred five cases were lost to follow-up study. The Cox regression method was used to analyze the factors related to cyst formation. METHODS: The incidence of cyst formation in the entire patient population was 1.6 and 3.6% in those undergoing follow-up examination for more than 5 years. Ten of 20 cysts developed between 10 to 23 years, nine between 5 to 10 years, and one in less than 5 years following the treatment. Cyst fluid aspiration, cystoperitoneal shunt placement, or craniotomy were used in three symptomatic cases. Analysis of age, sex, and treatment parameters yielded no significant relationship with cyst formation; however, radiation-induced tissue change following GKS (p = 0.027) and prior embolization (p = 0.011) were related to cyst formation. CONCLUSIONS: Overall, the incidence of cyst formation in patients who underwent GKS for AVM was 1.6%. The development of the cyst was related to the duration of the follow-up period. When cysts are symptomatic, surgical intervention should be performed. PMID- 15662795 TI - Gamma knife surgery for brain metastases from lung cancer. AB - OBJECT: The authors conducted a study to evaluate the safety and efficacy of gamma knife surgery (GKS) for the treatment of brain metastases from lung cancer. METHODS: Between February 1993 and May 2003 191 patients underwent treatment for 424 brain metastases from non-small (171 cases) and small cell lung carcinoma (20 cases). Imaging and clinical status were monitored every 3 months following the treatment. Kaplan-Meier survival curves, Cox proportional hazards regression for risk factor analysis, and nonparametric methods for evaluating tumor response were used. There was no difference in median survival following combined whole brain radiation therapy (WBRT) and gamma knife surgery (14 months) and GKS alone (15 months). There was also no difference between the median survival rates for either tumor type. In the multivariate analysis, age less than 65 years, Karnofsky Performance Scale score greater than 70, normal neurological status, multiple GKS treatments, and pre-GKS craniotomy were related to longer survival. Tumor control rates varied according to the volume of the metastases and were as follows: 84.4% (< 0.5 cm3), 94% (0.5-2 cm3), 89.1% (2-4 cm3), 93.4% (4-8 cm3), 85.7% (8-14 cm3), and 87.5% (> 14 cm3). Four lesions required post-GKS craniotomy due to swelling or rapid tumor progression. The rate of tumor shrinkage was higher when a volume was 2 cm3, lower in cystic lesions, lower in tumors with previous WBRT, and lower for margin doses less than 14 Gy. CONCLUSIONS: The risk benefit ratio of GKS in this series was satisfactory. There was no difference in response rates of the two tumor types, and WBRT did not improve the duration of survival. PMID- 15662796 TI - Cyst formation following gamma knife surgery for intracranial meningioma. AB - OBJECT: The authors conducted a study to evaluate the clinical significance of cyst formation or enlargement after gamma knife surgery (GKS) for intracranial benign meningiomas. METHODS: The medical records of 160 patients with 184 tumors were examined for those with follow-up data of more than 2 years among 270 patients who underwent GKS for intracranial meningiomas between February 1992 and November 2001. Cyst formation or enlargement following GKS was observed in five patients, one man and four women (mean age 61.2 years). The tumor location was the sphenoid ridge in one case, petroclival in two, tentorium in one, and parasagittal region in one. All patients underwent surgery before GKS. The mean tumor volume was 10.5 cm3, the mean margin dose was 13.4 Gy (median 14 Gy), and the mean maximum dose was 27.5 Gy (median 24.1 Gy). At the time of GKS three tumors were associated with cyst, of which two enlarged after radiosurgery. Three cysts developed de novo after GKS. Three of the five patients needed surgery to treat the cyst formation or enlargement. Histological examination demonstrated various findings such as tumor necrosis, proliferation of small vessels, vascular obliteration, and hemosiderin deposits. CONCLUSIONS: New cyst formation following GKS for benign intracranial meningioma is relatively rare; however, both preexisting and newly developed cysts tend to enlarge after GKS and often require surgery. PMID- 15662797 TI - Stereotactic noninvasive volume measurement compared with geometric measurement for indications and evaluation of gamma knife treatment. AB - OBJECT: Volume estimation is one of the most important criteria in the evaluation and follow up of radiosurgical treatments and outcomes; however, several limitations are involved in the calculation estimation of target volumes. METHODS: Retrospective and prospective studies were conducted to evaluate the efficacy of a new noninvasive stereotactic method when it is compared with geometric volume calculation of intracranial tumors for planning stereotactic radiosurgery treatment as well as for follow up and outcome evaluation. Two equations were created that permit comparison of the calculated and measured volumes. These equations took linear and quadratic forms, respectively. Volume estimation using the stereotactic approach compared with traditional volume calculation gave more accurate results regardless of the shape and size of the lesion. CONCLUSIONS: The use of stereotactic volume calculation is highly recommended in planning, follow up, and determination of the outcome in patients participating in radiosurgical treatment and should lead to more uniform reports of the response to treatment. PMID- 15662798 TI - Visual changes after gamma knife surgery for optic nerve tumors. Report of three cases. AB - Tumors involving the optic nerve (optic glioma, optic nerve sheath meningioma) are benign but difficult to treat. Gamma knife surgery (GKS) may be a useful treatment. The authors present data obtained in three such cases and record the effects of GKS. PMID- 15662799 TI - Gamma knife surgery for brain metastases in patients harboring four or more lesions: survival and prognostic factors. AB - OBJECT: This study was performed to evaluate the role of gamma knife surgery (GKS) in patients with a large number (four or more) of metastatic brain lesions. METHODS: The authors retrospectively reviewed the outcome in 130 patients who underwent GKS for metastatic lesions. Eighty-four patients presented with one to three lesions (Group A) and 46 presented with four or more lesions (Group B). The overall median survival time after GKS was 35 weeks. The median survival time in Group A (48 weeks) was significantly longer (p = 0.005) than the survival time in Group B (26 weeks). The recursive partitioning analysis (RPA) class was the only significant prognostic factor identified in multivariate analysis. The median survival for patients in RPA Classes I, II, and III was 72, 48, and 19 weeks, respectively, in Group A and 36 and 13 weeks for Classes II and III in Group B. The number of lesions, tumor volume, whole brain radiotherapy, primary tumor site, age, and sex did not affect survival significantly. CONCLUSIONS: It is suggested that GKS provides an increase in survival time even in patients with a large number (four or more) of metastatic lesions. Concerning the selection of patients for GKS, RPA class should be considered as the most important factor and multiplicity of the lesions alone should not be a reason for withholding GKS. PMID- 15662800 TI - Development of neurosurgery in Austria. Historical vignette. PMID- 15662801 TI - Glossopharyngeal neuralgia treated with gamma knife surgery: treatment outcome and failure analysis. Case report. AB - Glossopharyngeal neuralgia (GPN) is a rare condition in which patients present with intractable deep throat pain. Similar to trigeminal neuralgia (TN), treatment with microvascular decompression (MVD) has been successful in both. Because gamma knife surgery (GKS) has also been shown to be effective in treating TN, it seemed reasonable to apply it to GPN. The authors present the first report of GKS-treated GPN in a patient who presented with severe, poorly controlled GPN and who refused MVD. PMID- 15662802 TI - Analyzing 3-tesla magnetic resonance imaging units for implementation in radiosurgery. AB - OBJECT: The limiting factor affecting accuracy during gamma knife surgery is image quality. The new generation of magnetic resonance (MR) imaging units with field strength up to 3 teslas promise superior image quality for anatomical resolution and contrast. There are, however, questions about chemical shifts or susceptibility effects, which are the subject of this paper. METHODS: The 3-tesla MR imaging unit (Siemens Trio) was analyzed and compared with a 1-tesla unit (Siemens Magnetom Expert) and to a 1.5-tesla unit (Philips Gyroscan). Evaluation of the magnitude of error was performed within transverse slices in two orientations (axial/coronal) by using a cylindrical phantom with an embedded grid. Deviations were determined for 21 targets in a slab phantom with known geometrical positions within the stereotactic frame. Distortions caused by chemical shift and/or susceptibility effects were analyzed in a head phantom. Inhouse software was used for data analyses. The mean deviation was less than 0.3 mm in axial and less than 0.4 mm in coronal orientations. For the known targets the maximum deviation was 1.16 mm. By optimizing these parameters in the protocol these inaccuracies could be reduced to less than 1.1 mm. Due to inhomogeneities a shift in the z direction of up to 1.5 mm was observed for a dataset, which was shown to be compressed by 1.2 mm. CONCLUSIONS: The 3-tesla imaging unit showed superior anatomical contrast and resolution in comparison with the established 1 tesla and 1.5-tesla units; however, due to the high field strength the field within the head coil is very sensitive to inhomogeneities and therefore 3-tesla imaging data will have be handled with care. PMID- 15662803 TI - Evaluation of a new concept for the management of skull base chordomas and chondrosarcomas. AB - OBJECT: Chordomas and chondrosarcomas of the skull base are rare locally invasive tumors associated with high recurrence rates. The aim of this study was to evaluate the concept of microsurgical tumor volume reduction followed by early gamma knife surgery (GKS). METHODS: Thirteen patients with 15 tumors were treated between October 2000 and June 2003. There were three patients (23.1%) with chordomas and 10 (76.9%) with chondrosarcomas. There were nine men and four women who ranged in age between 19 and 69 years. All patients first underwent maximal tumor resection. Within 2 to 10 months after surgery they were treated with GKS. The mean postoperative tumor volume treated with GKS was 9.7 cm3 (range 1.4-20.3 cm3). Follow-up computerized tomography and magnetic resonance imaging examinations with volumetric tumor analysis were performed every 6 months after GKS. The mean treatment dose was 17 Gy and the mean isodose was 52%. The mean follow-up duration was 17 months during which there was only one tumor recurrence at the margin of the radiation field. The mean volume reduction was 35.4%. CONCLUSIONS: Results of this treatment strategy are encouraging but the efficacy of this multimodal treatment combining surgery and early GKS requires a longer follow up. PMID- 15662804 TI - Gamma knife surgery for hemangioblastomas. AB - OBJECT: The authors reviewed their 14-year experience using stereotactic radiosurgery for the treatment of hemangioblastomas and define the role and the proper strategy for radiosurgery of this condition. METHODS: This is a retrospective study of 38 hemangioblastomas in 13 patients. Seven patients had von Hippel-Lindau disease. All patients have undergone at least one follow-up visit. The median and mean tumor volumes were 0.23 cm3 and 0.72 cm3 respectively (range 0.004::4.84 cm3). Twenty-eight tumors received 20 Gy to the margin, and the remainder received 18 Gy. The median clinical follow-up period was 36 months (range 3-159 months). No patient died. The survival rate was 84.6% (11 of 13 patients). The actuarial 5- and 10-year survival rates were both 80.8%. The median radiological follow-up period was 35 months (range 7-147 months). Only one tumor increased in volume 24 months after treatment in association with an intratumoral hemorrhage. The tumor control rate was 97.4% (37 of 38 tumors). Actuarial 5- and 10-year control rates were both 96.2%. New lesions and/or those increasing in size outside the irradiated area were discovered in five patients (38.5%). Nine tumors revealed peritumoral contrast enhancement which was seen more frequently in larger tumors with a volume greater than 0.5 cm3 (p = 0.0034). CONCLUSIONS: Gamma knife surgery is a safe and effective method to control hemangioblastomas for as many as 10 years. Higher doses and smaller tumors probably contribute to good outcomes. Recurrence outside the original irradiated area is common. Peritumoral contrast enhancement may be seen in larger tumors. The authors recommend regular imaging follow up and early repeated treatment in the face of new or growing tumors. PMID- 15662805 TI - Pineal tumors: analysis of treatment results in 20 patients. AB - OBJECT: The authors evaluate their results when using gamma knife surgery (GKS) in the management of patients with tumors in the pineal region. METHODS: This is a retrospective clinical evaluation of 20 patients with primary tumors of the pineal region treated with GKS from November 1994 through August 2003. There were 13 germ cell tumors, two pineoblastomas, two low-grade gliomas, one primitive neuroectodermal tumor, one teratoma, and one pineocytoma. There were 10 male and 10 female patients. Their median age was 15.5 years (range 5-71 years). The median margin dose was 11 Gy (range 8-20 Gy). The median target volume was 3.1 cm3 (range 0.1-49.9 cm3). Five patients received sequential systemic chemotherapy and four underwent adjuvant conventional radiation therapy. Seventeen (85%) of 20 patients are alive with a median survival of 30.4 months (range 0-85.7 months). Two patients required retreatment. Three patients died: one of unrelated causes, one who presented with extensive local disease, and the other of meningeal carcinomatosis with local control of the primary tumor. No complications from GKS were noted. CONCLUSIONS: This initial experience suggests that GKS is a valuable treatment modality for the management of pineal region tumors. This technique offers excellent local tumor control and minimal patient morbidity, allowing for immediate use of systemic chemotherapy and/or conventional radiation if indicated. PMID- 15662806 TI - Does gamma knife surgery stimulate cellular immune response to metastatic brain tumors? A histopathological and immunohistochemical study. AB - OBJECT: The aim of this study was to analyze the cellular immune response and histopathological changes in secondary brain tumors after gamma knife surgery (GKS). METHODS: Two hundred ten patients with cerebral metastases underwent GKS. Seven patients underwent subsequent craniotomy for tumor removal between 1 and 33 months after GKS. Four of these patients had one tumor, two patients had two tumors, and one patient had three. Histological and immunohistochemical investigations were performed. In addition to routine H & E and Mallory trichrome staining, immunohistochemical reactions were conducted to characterize the phenotypic nature of the cell population contributing to the tissue immune response to neoplastic deposits after radiosurgery. Light microscopy revealed an intensive lymphocytic infiltration in the parenchyma and stroma of tumor samples obtained in patients in whom surgery was performed over 6 months after GKS. Contrary to this, extensive areas of tissue necrosis with either an absent or scanty lymphoid population were observed in the poorly controlled neoplastic specimens obtained in cases in which surgery was undertaken in patients less than 6 months after GKS. Immunohistochemical characterization demonstrated the predominance of CD3-positive T cells in the lymphoid infiltration. CONCLUSIONS: Histopathological findings of the present study are consistent with a cellular immune response of natural killer cells against metastatic brain tumors, presumably stimulated by the ionizing energy of focused radiation. PMID- 15662807 TI - Gamma knife surgery for trigeminal neuralgia: improved initial response with two isocenters and increasing dose. AB - OBJECT: The authors sought to evaluate the initial response of trigeminal neuralgia (TN) to gamma knife surgery (GKS) based on the number of shots delivered and radiation dose. METHODS: Between September 1998 and September 2003, some 63 patients with TN refractory to medical or surgical management underwent GKS at Upstate Medical University. Ten patients had multiple sclerosis and 25 patients had undergone prior invasive treatment. Gamma knife surgery was delivered to the trigeminal nerve root entry zone in one shot in 27 patients or two shots in 36 patients. The radiation dose was escalated to less than or equal to 80 Gy in 20 patients, 85 Gy in 21 patients, and greater than or equal to 90 Gy in 22 patients. Pain before and after GKS was assessed using the Barrow Neurological Institute Pain Scale and the improvement score was analyzed as a function of dose grouping and number of shots. Sixty patients were available for evaluation, with an initial overall and complete response rate of 90% and 27%, respectively. There was a greater improvement score for patients who were treated with two shots compared with one shot, mean 2.83 compared with 1.72 (p < 0.001). There was an increased improvement in score at each dose escalation level: less than or equal to 80 Gy (p = 0.017), 85 Gy (p < 0.001), and greater than or equal to 90 Gy (p < 0.001). Linear regression analysis also indicated that there was a greater response with an increased dose (p = 0.021). Patients treated with two shots were more likely to receive a higher dose (p < 0.001). There were no severe complications. Five patients developed mild facial numbness. CONCLUSIONS: Gamma knife surgery is an effective therapy for TN. Initial response rates appear to correlate with the number of shots and dose. PMID- 15662808 TI - Volume reduction in meningiomas after gamma knife surgery. AB - OBJECT: The purpose of this study was to evaluate the volume-reducing effects of gamma knife surgery (GKS) of meningiomas with and without previous surgical treatment. METHODS: A group of 127 patients with a mean age of 57.1 years (range 9-81 years) with 142 meningiomas (128 World Health Organization Grade I and 14 Grade II) were included in this study. The management strategy reduces tumor volume with surgery when necessary (81 patients). Stereotactic GKS with a Gamma Knife model C was performed in all tumors of suitable size. Magnetic resonance imaging follow-up examinations with volumetric tumor analysis was performed 6 months after treatment and annually thereafter. The mean tumor volume was 5.9 cm3 (range < 5 to > 40 cm3). The mean follow-up time after GKS was 29.3 months (range 11-61 months). The mean prescription dose was 13.8 Gy (range 10-18 Gy). A reduction in volume occurred in 117 (82.4%) of all tumors, and in 20 tumors (14.1%) growth ceased. The overall tumor control rate of 96.4%. The mean volume reduction achieved with GKS was more than 46.1%. Only five tumors (3.5%) stowed a volume increase. CONCLUSIONS: Gamma knife surgery was effective in reducing meningioma volume at short-term follow up. Further studies are needed to examine the development of these findings over a longer period. PMID- 15662809 TI - Radiosurgery of vestibular schwannomas: summary of experience in 829 cases. AB - OBJECT: Management options for vestibular schwannomas (VSs) have greatly expanded since the introduction of stereotactic radiosurgery. Optimal outcomes reflect long-term tumor control, preservation of cranial nerve function, and retention of quality of life. The authors review their 15-year experience. METHODS: Between 1987 and 2002, some 829 patients with VSs underwent gamma knife surgery (GKS). Dose selection, imaging, and dose planning techniques evolved between 1987 and 1992 but thereafter remained stable for 10 years. The average tumor volume was 2.5 cm3. The median margin dose to the tumor was 13 Gy (range 10-20 Gy). No patient sustained significant perioperative morbidity. The average duration of hospital stay was less than 1 day. Unchanged hearing preservation was possible in 50 to 77% of patients (up to 90% in those with intracanalicular tumors). Facial neuropathy risks were reduced to less than 1%. Trigeminal symptoms were detected in less than 3% of patients whose tumors reached the level of the trigeminal nerve. Tumor control rates at 10 years were 97% (no additional treatment needed). CONCLUSIONS: Superior imaging, multiple isocenter volumetric conformal dose planning, and optimal precision and dose delivery contributed to the long-term success of GKS, including in those patients in whom initial microsurgery had failed. Gamma knife surgery provides a low risk, minimally invasive treatment option for patients with newly diagnosed or residual VS. Cranial nerve preservation and quality of life maintenance are possible in long-term follow up. PMID- 15662810 TI - Gamma knife surgery for choroidal neovascularization in age-related macular degeneration. Technical note. AB - The authors conducted a study to determine a way of overcoming the poor-quality demonstration of choroidal neovascularization (CNV) associated with age-related macular degeneration (AMD) on conventional magnetic resonance (MR) imaging studies. The poor MR imaging demonstration of CNV in patients with AMD makes the use of gamma knife surgery more difficult. This difficulty, however, can be overcome by use of a modified time-of-flight MR imaging,sequence with Gd enhancement and coronal reconstruction. PMID- 15662811 TI - Increased preservation of functional hearing after gamma knife surgery for vestibular schwannoma. AB - OBJECT: Gamma knife surgery (GKS) for vestibular schwannoma is still associated with an additional hearing loss of approximately 30%. The purpose of this study was to record the effect on hearing preservation of maintaining a margin dose of 13 Gy while reducing the maximum dose to 20 Gy. METHODS: Seventy-eight of 95 patients who entered a prospective protocol with a follow up of at least 12 months (mean 22 months) were evaluated. The mean tumor volume was 2.28 cm3. After a mean follow-up duration of 22 months, the magnetic resonance imaging-based tumor control rate was 87%. In two cases a second procedure (surgery) was necessary. Thus, the clinical control rate was 97.5%. In two cases there was an increase in trigeminal dysesthesia. One patient suffered transient facial nerve impairment. Functional hearing was preserved in 83.4% of the patients with functional hearing preoperatively. CONCLUSIONS: Reducing the maximum dose to 20 Gy seems to be an effective treatment, which probably increases preservation of functional hearing without sacrificing the high tumor control rates achieved in radiosurgery. Postradiosurgery tumor swelling occurred in 25% of the cases and was not correlated with hearing deterioration. PMID- 15662812 TI - Gamma knife surgery of brain cavernous hemangiomas. AB - OBJECT: The authors conducted a study to record more detailed information about the natural course and factors predictive of outcome following gamma knife surgery (GKS) for cavernous hemangiomas. METHODS: One hundred twelve patients with brain cavernous hemangiomas underwent GKS between 1993 and 2000. The median prescription dose was 16 Gy. One hundred seven patients were followed for a median of 48 months (range 6-114 months). The rebleeding rate was 1.6%, which is not significantly different with that prior to radiosurgery (2%). An increase in volume was observed in 1.8% of cases and a decrease in 45%. Perilesional edema was detected in 27% of patients, which, together with the rebleeding, caused a transient morbidity rate of 20.5% and permanent morbidity rate of 4.5%. Before radiosurgery 39% of patients suffered from epilepsy and this improved in 45% of them. Two patients with brainstem cavernous hemangiomas died due to rebleeding. Rebleeding was more frequent in female middle-aged patients with a history of bleeding, a larger lesion volume, and a prescription dose below 13 Gy. Edema after GKS occurred more frequently in patients who had surgery, a larger lesion volume, and in those in whom the prescription dose was more than 13 Gy. CONCLUSIONS: Gamma knife surgery of cavernous hemangiomas can produce an acceptable rate of morbidity, which can be reduced by using a lower margin dose. Lesion regression was observed in many patients. Radiosurgery seems to remain a suitable treatment modality in carefully selected patients. PMID- 15662813 TI - Progress in glaucoma treatment research: a nonrandomized prospective study of 102 patients with advanced refractory glaucoma treated by Leksell gamma knife irradiation. AB - OBJECT: The authors performed a study to characterize the effects of ciliary body gamma knife surgery (GKS) for advanced glaucoma. METHODS: In addition to a pilot project involving 14 patients, 88 other patients were treated. Four 8-mm isocenters were used. Blind eyes received 20 Gy to the 50% isodose. If there was preserved vision the dose was 15 Gy. The median follow up was 20 months (range 2 54 months). Severe pain was present in 93% of patients with secondary glaucoma; after GKS it was substantially improved in 89% of cases. In primary open angle glaucoma, the pain was less frequent and severe. In more than half the patients it disappeared during a median latency of 6 weeks. The median intraocular pressure value was lowered in patients with secondary glaucoma (from 51.3-27 mm Hg). The same values for primary glaucoma were 25.3 and 16.1 mm Hg, respectively, after a median latency of 12 weeks. Twenty-seven of 40 patients with neovascular glaucoma showed a marked reduction of neovascularization after a median latency of 18 weeks. Treatment complications were slight. There was postoperative lacrimation in 61%. Two patients underwent postirradiation cataract extraction. Noninfectious keratitis was observed in two patients. Pharmacotherapy could be reduced in all patients and ceased in approximately one third. There has been no recurrence of the initial symptoms and no worsening of visual acuity. CONCLUSIONS: Gamma knife surgery ameliorates the main symptoms in advanced glaucomas and precludes the need for eventual ocular enucleation. PMID- 15662814 TI - Long-term results of Leksell gamma knife surgery for trigeminal schwannomas. AB - OBJECT: The authors evaluated the long-term efficacy of gamma knife surgery (GKS) in patients with trigeminal schwannomas. METHODS: Fifty-six patients, 31 women and 25 men (mean age 42 years), underwent GKS for trigeminal schwannomas. Fourteen had previously undergone surgery, and GKS was the primary treatment in the remaining 42 patients. The mean target volume was 8.7 cm3 (range 0.8-33 cm3); the mean maximum dose was 27 Gy (range 20-40 Gy); the mean tumor margin dose was 13.3 Gy (range 10-15 Gy); and the mean follow-up period was 68 months (range 27 114 months). Disappearance of the tumor occurred in seven patients. An obvious decrease in tumor volume was observed in 41 patients, four tumors remained unchanged, and four tumors progressed at 5, 26, 30, and 60 months, respectively. One patient with disease progression died of tumor progression at 36 months after GKS. The tumor growth control rate in this group was 93% (52 of 56 cases). Mild numbness or diplopia was relieved completely in 14 patients. Improvement of other neurological deficits was demonstrated in 25 patients. Trigeminal nerve dysfunction was either unchanged or slightly worse in 13 patients after GKS. Four patients experienced mild symptom deterioration related to tumor progression. CONCLUSIONS: Radiosurgery proved to be an effective treatment for small- and medium-sized trigeminal schwannomas. Some larger tumors are also suitable for radiosurgery if there is no significant brainstem compression. PMID- 15662815 TI - The long-term results of gamma knife radiosurgery for hemangioblastomas of the brain. AB - OBJECT: The authors assessed the long-term result of gamma knife surgery (GKS) for hemangioblastomas of the brain (HABs) and show histopathological findings after GKS. METHODS: Thirty-five patients, 28 men and seven women, with a mean age of 36 years underwent GKS. Eighteen patients presented with multiple tumors and 17 with a solitary tumor. Twenty-one patients had von Hippel-Lindau (VHL) disease. The mean tumor diameter was 13 mm (range 5-55 mm). The mean follow up after GKS was 66 months (range 24-114 months). The mean prescription dose was 17.2 Gy (range 12-24 Gy) at the tumor margin. For tumors close to or within the brainstem a prescription dose of 12 to 13 Gy was used. At the most recent follow up, 29 patients were alive, six were dead, and satisfactory tumor control had been achieved in 29. A stable or improved neurological status was obtained in 21 patients. Eight patients underwent open surgery because of tumor-associated cyst enlargement or the development of new tumors after GKS. Seven patients developed new tumors and five of them required a second GKS. The 1-year tumor control rate was 94%; 2 years, 85%; 3 years, 82%; 4 years, 79%; and 5 years, 71%. Histopathology showed that no tumor cells were found and there was degeneration and necrosis in a tumor nodule 48 months after GKS with a prescription dose of 18 Gy. CONCLUSIONS: Gamma knife surgery was a useful choice for small- or medium sized, solid HAB in the long term, especially when the tumor margin dose was 18 Gy. Although GKS can treat multiple tumors in a single session, for HABs associated with VHL disease, GKS faces the dual problems of tumor recurrence or development of a new tumor. PMID- 15662816 TI - Tolerance dose in gamma knife surgery of lesions extending to the anterior visual pathway. AB - OBJECT: The authors performed a retrospective analysis of the radiation dose to the anterior visual pathway (AVP) to assess its tolerance to gamma knife surgery. METHODS: They examined five cases followed for more than 3 years. The AVP was treated with 10-Gy doses or higher. The mean maximum delivered dose to the AVP was 14 Gy. Ten gray or more was delivered to 25.5% of the ipsilateral AVP, 12 Gy or more to 12.5% of the ipsilateral AVP, and 14 Gy or more to 5.7% of the ipsilateral AVP. Although the mean follow-up period was 40.8 months (36-51 months), no cases of visual function deterioration developed. CONCLUSIONS: The tolerance dose of the AVP is considered to be less than 8 to 10 Gy; however, although the delivered dose to the AVP definitely exceeded the tolerance dose in all five cases, no visual disturbance has been identified. Longer follow up is required before any final conclusions may be drawn. Nonetheless, it is suggested that a visual disturbance may be avoided by using careful accurate dose planning even if the dose delivered to the AVP is higher than currently believed to be acceptable. PMID- 15662817 TI - Gamma knife thalamotomy for movement disorders: evaluation of the thalamic lesion and clinical results. AB - OBJECT: The authors studied the effects of gamma knife thalamotomy (GKT) on Parkinson disease-related tremor and essential tremor before and after reloading of radioactive cobalt. METHODS: Based on experience in stereotactic thalamotomy aided by depth microrecording, the target was located at the lateral border of the thalamic ventralis intermedius nucleus (VIM). For more precise targeting, the percentage representation of the thalamic VIM in relation to the entire thalamic length is useful. The location of the target was determined on magnetic resonance (MR) imaging and computerized tomography scanning. A maximum dose of 130 Gy was delivered to the target by using a single isocenter with the 4-mm collimator. In more recent cases, a systematic follow-up examination was performed at 3, 6, 12, 18, and 24 months after GKT. Since 1993, the authors have treated 70 patients with PD. Throughout the series the same dosimetric technique has been used. The course after GKT was compared between the 25 cases with PD treated before reloading and the 35 cases treated after reloading. In the majority (80-85%) treated after reloading, tremor and rigidity were reduced around 6 months after GKT. In the cases treated before reloading this effect took approximately 1 year. The thalamic reaction on MR imaging showed the same two lesion types in both series: a restricted and a diffuse. After reloading the restricted lesion was more frequent and the lesion volume was smaller. CONCLUSIONS: The shorter delay in clinical improvement and smaller lesion size may be related to an increased radiation dose. PMID- 15662818 TI - Gamma knife surgery for glomus jugulare tumors: an intermediate report on efficacy and safety. AB - OBJECT: Glomus jugulare tumors are rare tumors that commonly involve the middle ear, temporal bone, and lower cranial nerves. Resection, embolization, and radiation therapy have been the mainstays of treatment. Despite these therapies, tumor control can be difficult to achieve particularly without undo risk of patient morbidity or mortality. The authors examine the safety and efficacy of gamma knife surgery (GKS) for glomus jugulare tumors. METHODS: A retrospective review was undertaken of the results obtained in eight patients who underwent GKS for recurrent, residual, or unresectable glomus jugulare tumors. The median radiosurgical dose to the tumor margin was 15 Gy (range 12-18 Gy). The median clinical follow-up period was 28 months, and the median period for radiological follow up was 32 months. All eight patients demonstrated neurological stability or improvement. No cranial nerve palsies arose or deteriorated after GKS. In the seven patients in whom radiographic follow up was obtained, the tumor size decreased in four and remained stable in three. CONCLUSIONS: Gamma knife surgery would seem to afford effective local tumor control and preserves neurological function in patients with glomus jugulare tumors. If long-term results with GKS are equally efficacious, the role of stereotactic radiosurgery will expand. PMID- 15662819 TI - Radiosurgery for patients with recurrent small cell lung carcinoma metastatic to the brain: outcomes and prognostic factors. AB - OBJECT: Lung carcinoma is the leading cause of death from cancer. More than 50% of those with small cell lung cancer develop a brain metastasis. Corticosteroid agents, radiotherapy, and resection have been the mainstays of treatment. Nonetheless, median survival for patients with small cell lung carcinoma metastasis is approximately 4 to 5 months after cranial irradiation. In this study the authors examine the efficacy of gamma knife surgery for treating recurrent small cell lung carcinoma metastases to the brain following tumor growth in patients who have previously undergone radiation therapy, and they evaluate factors affecting survival. METHODS: A retrospective review of 27 patients (47 recurrent small cell lung cancer brain metastases) undergoing radiosurgery was performed. Clinical and radiographic data obtained during a 14 year treatment period were collected. Multivariate analysis was utilized to determine significant prognostic factors influencing survival. The overall median survival was 18 months after the diagnosis of brain metastases. In multivariate analysis, factors significantly affecting survival included: 1) tumor volume (p = 0.0042); 2) preoperative Karnofsky Performance Scale score (p = 0.0035); and 3) time between initial lung cancer diagnosis and development of brain metastasis (p = 0.0127). Postradiosurgical imaging of the brain metastases revealed that 62% decreased, 19% remained stable, and 19% eventually increased in size. One patient later underwent a craniotomy and tumor resection for a tumor refractory to radiosurgery and radiation therapy. In three patients new brain metastases were demonstrating on follow-up imaging. CONCLUSIONS: Stereotactic radiosurgery for recurrent small cell lung carcinoma metastases provided effective local tumor control in the majority of patients. Early detection of brain metastases, aggressive treatment of systemic disease, and a therapeutic strategy including radiosurgery can extend survival. PMID- 15662820 TI - Gamma knife surgery of superficially located meningioma. AB - OBJECT: The authors analyzed tumor control rates and complications in patients with superficially located meningiomas after gamma knife surgery (GKS). METHODS: Between 1998 and 2003, GKS was performed in 23 patients with 26 lesions in whom follow-up imaging for 1 year or more was available. The male/female ratio was 1:22. The mean age was 59 years. The median tumor volume was 4.7 cm3, and the mean margin dose was 16 Gy at the 50% isodose line. Peritumoral edema was revealed on magnetic resonance (MR) imaging in four patients before GKS. Magnetic resonance imaging and clinical examinations were performed every 6 months after GKS. The mean follow-up duration was 32 months. The tumor shrank in eight cases, was stable in 17, and enlarged in one; thus 25 (95%) of 26 tumors were controlled. A peritumoral high signal on T2-weighted MR images was found in eight lesions and preexisting edema was aggravated in three lesions after GKS. Ten of these 11 patients complained of severe headache, and three patients experienced neurological deficits at the same time after a mean latency of 3 months; however, high signal was not demonstrated on imaging before 6 months on average. Steroid agents, when required, gave relief to all patients. The complication rate was 43% (10 of 23 cases). High signal disappeared in nine patients and decreased in the remaining two. High signal was associated with a high integral dose and a large tumor volume. Tumor shrinkage at the last follow-up examination was more prominent in the patients with symptomatic high signal (p = 0.03). CONCLUSIONS: There was a good tumor control rate with a high complication rate. Longer follow up of more patients is needed. Adjusting the dose-volume relationship should be considered to reduce complications. PMID- 15662821 TI - Microvascular decompression after gamma knife surgery for trigeminal neuralgia: intraoperative findings and treatment outcomes. AB - OBJECT: The authors sought to determine whether the results of trigeminal microvascular decompression (MVD) are influenced by prior gamma knife surgery (GKS). METHODS: Gamma knife surgery is an established procedure for treating medically intractable trigeminal neuralgia but failures do occur. The authors assessed six patients (two men and four women; mean age 52 years) who experienced pain recurrence after GKS and elected to undergo trigeminal MVD via retrosigmoid craniotomy. Three patients underwent a single GKS to a maximal dose of 80 Gy, whereas three others underwent a second GKS to total of 120 to 135 Gy. At surgery, none of the six patients demonstrated excess arachnoid thickening, grossly apparent changes in the nerve itself, or any other tissue alterations that made successful mobilization of a blood vessel from the trigeminal root entry zone technically more difficult. A single individual had a small atherosclerotic plaque in the superior cerebellar artery near its contact point with the trigeminal nerve. Follow up at a mean of 25.4 months (range 7.5-42 months) indicated that five patients were pain free. One patient had improved but still relied on medications for pain control. CONCLUSIONS: In the authors' experience, trigeminal MVD can be performed without added difficulty in patients who have previously undergone GKS. The success rates seem similar to those normally associated with MVD. Patients who elect the less invasive option of GKS can be assured that trigeminal MVD remains a viable alternative at a later date if further surgery is required. PMID- 15662822 TI - Prolonged survival in a subgroup of patients with brain metastases treated by gamma knife surgery. AB - OBJECT: The authors analyzed the factors involved in determining prolonged survival (> or = 24 months) in patients with brain metastases treated by gamma knife surgery (GKS). METHODS: Between 1995 and 2003, a total of 116 patients underwent 167 GKS procedures for brain metastases. There was no special case selection. Smaller and larger lesions were treated with different protocols. The mean patient age was 56.9 years, the mean number of initial lesions was 3.15, and the mean lesion volume was 10.45 cm.3 The mean follow-up time was 9.2 months. The median patient survival was 8.68 months. One-, 2-, 3-, 4-, and 5-year actuarial survival rates were 31.8%, 19.8%, 14.6%, 7.7%, and 6.9%, respectively. Patient age, number of lesions at presentation, and lesion volume had no influence on patient survival. Twenty-three (19.8%) patients survived for 24 months or more. Certain factors were associated with increased survival time. These were stable primary disease (21 of 23 patients), a long latency between diagnosis of the primary tumor and the occurrence of brain metastases (mean 28.4 months, median 16 months), absence of third-organ involvement, and repeated local procedures. Ten patients underwent repeated GKS (mean 3.4 per patient). Seven patients required open surgery for local treatment failures (recurrence or radiation necrosis). Two patients had both. Fifteen patients underwent repeated procedures. CONCLUSIONS: Aggressive local therapy with GKS, repeated GKS, and GKS plus surgery can achieve increased survival in a subgroup of patients with stable primary disease, no third-organ involvement, and long primary-brain secondary intervals. PMID- 15662823 TI - Diagnostic value of thallium-201 chloride single-photon emission computerized tomography in differentiating tumor recurrence from radiation injury after gamma knife surgery for metastatic brain tumors. AB - OBJECT: The authors assessed the diagnostic value of 201Tl Cl single-photon emission computerized tomography (SPECT), performed after gamma knife surgery (GKS) for metastatic brain tumors in differentiating tumor recurrence from radiation injury. METHODS: Of 6503 metastatic brain tumors treated with GKS, 201Tl SPECT was required in 72 to differentiate between tumor recurrence and radiation injury. When the Tl index was greater than 5, the lesion was diagnosed as a tumor recurrence. When the index was < 3.0 it was called radiation injury. In cases with a Tl index between 3 and 5, 201Tl SPECT was repeated once per month until the Tl index was greater than 5 or less than 3. If the Tl index fluctuated between 3 and 5 for 2 months, the lesion was diagnosed as radiation injury. The final diagnosis was based on histological examination or clinical course. The sensitivity of the method was 91%; thus 201Tl SPECT is effective for differentiating between tumor recurrence and radiation injury in metastatic brain tumors treated with GKS. Caution is necessary, however, for the following reasons: 1) simple interinstitutional comparisons of Tl indices are not possible because measurement methods are institute specific; 2) steroid administration decreases the Tl index to a variable degree; and 3) a severe radiation injury lesion, as is often seen after repeated GKS or very high dose GKS, may have a Tl index greater than 5. CONCLUSIONS: Used with critical insight 201Tl Cl SPECT can be useful in distinguishing between tumor regrowth and radiation necrosis in patients with cerebral metastases. PMID- 15662824 TI - Initial experience with gamma knife surgery for endocrine ophthalmopathy. AB - OBJECT: The aim of this study was to evaluate both the effectiveness and safety of the treatment of endocrine ophthalmopathy with gamma knife surgery (GKS). METHODS: Five patients were included in a prospective study designed to assess the results of GKS of endocrine ophthalmopathy secondary to Graves disease. All the patients completed a 2-year follow-up period. During this period, the patients were evaluated both clinically and by means of additional methods, including computerized tomography and magnetic resonance imaging studies. The minimum dose delivered to the 50% isodose line was 6.5 Gy in all the patients. In all cases, a clinical improvement was observed. The best effect was seen in symptom regression related to soft-tissue involvement. No treatment-related side effects were detected. CONCLUSIONS: In light of the results obtained the authors consider that GKS may be a safe and effective way to treat endocrine ophthalmopathy. PMID- 15662825 TI - Gamma knife surgery for refractory postherpetic trigeminal neuralgia: targeting in one session both the retrogasserian trigeminal nerve and the centromedian nucleus of the thalamus. AB - OBJECT: The authors tested the hypothesis that two targets are needed to treat postherpetic trigeminal neuralgia (TN): one in the trigeminal nerve for the direct sharp pain and one in the thalamus for the diffuse burning pain. METHODS: Three patients with refractory postherpetic TN were treated with gamma knife surgery (GKS) through a novel two-target approach. In a single treatment session, both the trigeminal nerve and centromedian nucleus were targeted. First, the trigeminal nerve, ipsilateral to the facial pain, was treated with 60 to 80 Gy. Second, the centromedian nucleus was localized using standard coordinates and by comparing magnetic resonance images with a stereotactic atlas. A single dose of 120 to 140 Gy was delivered to the target point with a single 4-mm isocenter. Patients were followed clinically and with neuroimaging studies. Pain relief was scored as excellent (75-100%), good (50-75%), poor (25-50%); or none (0-25%). Follow up ranged from 6 to 53 months. There were no GKS-related complications. Two patients died of unrelated medical illnesses but had good or excellent pain relief until death. One patient continues to survive with 44 months follow up and no decrease in pain intensity, but with a decreased area of pain. CONCLUSIONS: Combined GKS of the centromedian nucleus and trigeminal nerve in a single treatment session is feasible and safe, and the effect was promising. A larger study is required to confirm and expand these results. PMID- 15662826 TI - Gamma knife surgery for atypical meningiomas. AB - OBJECT: Complete resection is the optimal treatment for atypical meningiomas (AMs) but its feasibility depends on the tumor site. The object of this study was to assess the effect of gamma knife surgery (GKS) on AM. METHODS: In 15 patients 21 AMs were treated by GKS. Four patients had residual lesions and 10 patients had recurrent tumors after one or more microsurgical interventions. Three patients were treated twice with GKS because of tumor tissue outside the treatment volume, either at the margin or at a distant location. The median clinical and neuroimaging follow-up period was 35 months (range 21-67 months). Ten tumors shrank 6 to 12 months after GKS, 10 remained stable, and one grew. Between 18 and 36 months after GKS, four patients had a distant recurrence, and two had a margin recurrence. In one of these cases, an additional local recurrence was demonstrated 1 year later, and the patient underwent standard radiotherapy. No patient suffered persistent adverse effects after radiosurgery. CONCLUSIONS: After early tumor shrinkage, high recurrence rates were demonstrated both at the treatment margin and at distant locations in cases treated for AM. There was only one recurrence within the GKS radiation field. For small- and medium-sized AMs GKS may be a safe adjunct to other treatment modalities. PMID- 15662827 TI - Tumor recurrence and survival following gamma knife surgery for brain metastases. AB - OBJECT: The authors evaluated prognostic factors for tumor recurrence and patient survival following gamma knife surgery (GKS) for brain metastasis. METHODS: A retrospective review of 101 patient charts was undertaken for those patients treated with GKS for brain metastases from 1994 to 2001. Recurrence rates of brain metastasis following GKS depended on the duration of patient survival. Long term survival was associated with a higher risk of tumor recurrence and shorter term survival was associated with a lower risk. The duration of survival following GKS for brain metastases seems to be characteristic of the primary disease rather than the cerebral disease. CONCLUSIONS: Recurrence rates of brain metastasis following GKS are related to duration of survival, which is in turn mostly dependent on the nature and course of the primary tumor. PMID- 15662828 TI - Ultrastructural changes in arteriovenous malformations after gamma knife surgery: an electron microscopic study. AB - OBJECT: The authors analyzed morphological alterations at the subcellular level by undertaking transmission electron microscopy in arteriovenous malformations (AVMs) after gamma knife surgery (GKS). METHODS: Histological, immunohistochemical, and electron microscopic investigations were performed in a series of pathological specimens obtained in seven patients. The patients harbored cerebral AVMs that had been previously treated with GKS and had suffered subsequent bleeding 10 to 52 months after treatment. Histological studies revealed spindle cell proliferation in the connective tissue stroma and in the subendothelial region of the irradiated AVM vessels. Electron microscopy demonstrated different ultrastructural characteristics of this spindle cell population. There were cells with a smooth-edged oval nuclei surrounded by massive bundles of collagen fibers in the extracellular matrix. Other cells with the same nuclear morphology contained abundant intracytoplasmic filaments. Nuclear deformation was connected to a fibrillary system developed within the cytoplasm, and peripheral attachment sites were related to an extracellular layer of basement membrane-like material arranged parallel to the cell border. Also present were cells containing well-developed cisterns of rough endoplasmic reticulum and dense bodies at the periphery of the cytoplasm with folded, irregular nuclei. CONCLUSIONS: The ultrastructural and histological characteristics of the spindle cell population in the GKS-treated AVMs are similar to those designated as myofibroblasts in wound healing processes and pathological fibromatoses. Because similar cell modifications have not been demonstrated in control nonirradiated AVM specimens, these myofibroblasts may contribute to the shrinking process and final occlusion of AVMs after radiosurgery. PMID- 15662829 TI - Temporary symptomatic swelling of meningiomas following gamma knife surgery. Report of two cases. AB - In two patients in whom gamma knife surgery was performed for meningiomas clinically significant volume increases were observed in the first 3 months after treatment. Clinical examination and various imaging studies form the basis of the report in these patients. In each case, the volume increase was temporary. PMID- 15662830 TI - Mitochondria and neuronal death/survival signaling pathways in cerebral ischemia. AB - Apoptotic cell death pathways have been implicated in acute brain injuries, including cerebral ischemia, brain trauma, and spinal cord injury, and in chronic neurodegenerative diseases. Experimental ischemia and reperfusion models, such as transient focal/global ischemia in rodents, have been thoroughly studied and suggest the involvement of mitochondria and the cell survival/death signaling pathways in cell death/survival cascades. Recent studies have implicated mitochondria-dependent apoptosis involving pro- and antiapoptotic protein binding, the release of cytochrome c and second mitochondria-derived activator of caspase, the activation of downstream caspases-9 and -3, and DNA fragmentation. Reactive oxygen species are known to be significantly generated in the mitochondrial electron transport chain in the dysfunctional mitochondria during reperfusion after ischemia, and are also implicated in the survival signaling pathway that involves phosphatidylinositol-3-kinase (PI3-K), Akt, and downstream signaling molecules, like Bad, 14-3-3, and the proline-rich Akt substrate (PRAS), and their bindings. Further studies of these survival pathways may provide novel therapeutic strategies for clinical stroke. PMID- 15662831 TI - Impairment of brain mitochondrial oxidative phosphorylation accompanying vitamin E oxidation induced by iron or reactive nitrogen species: a selective review. AB - Mitochondria are exposed to large fluxes of iron, and reactive oxygen and nitrogen species. Hence they are susceptible to oxidative stress, a process inhibited by vitamin E. Our investigations show that iron uncouples oxidative phosphorylation whereas peroxynitrite and nitrite are inhibitors of oxidative phosphorylation. Oxidation of mitochondrial vitamin E is accompanied by generation of lipid peroxidation products, altered enzyme activity and electrical conductance etc., and result in inefficient oxidative phosphorylation. Vitamin E is important for mitochondrial function because: (1) Prior investigations have shown that vitamin E is essential for maintaining mitochondrial respiration. (2) Vitamin E is the most potent, lipid-soluble antioxidant localized ideally in mitochondrial membranes. (3) The decline in respiratory control ratios (RCR) of rat brain mitochondria exposed to peroxynitrite closely paralleled the oxidative elimination of vitamin E. (4) Finally, iron is a strong uncoupler of oxidative phosphorylation in brain mitochondria from vitamin E deficient animals and not from controls. PMID- 15662832 TI - Biochemical aspects of neurodegeneration in human brain: involvement of neural membrane phospholipids and phospholipases A2. AB - Neural membrane phospholipids are hydrolyzed by a group of enzymes known as phospholipases. This process results in the generation of second messengers such as arachidonic acid, eicosanoids, platelet activating factor, and diacylglycerols. High levels of these metabolites are neurotoxic and are associated with neurodegeneration. The collective evidence from many studies suggests that neural membrane phospholipid metabolism is disturbed in neural trauma and neurodegenerative diseases. This disturbance is caused by the stimulation of phospholipases A2. Stimulation of these enzymes produces changes in membrane permeability, fluidity, and alteration in ion homeostasis. Low calcium influx produces mild oxidative stress and results in neurodegeneration promoted by apoptosis, whereas a calcium overload generates high oxidative stress and causes neurodegeneration associated with necrosis. Alterations in phospholipid metabolism along with the accumulation of lipid peroxides and compromised energy metabolism may be responsible for neurodegeneration in ischemia, spinal cord trauma, head injury, and Alzheimer disease. The synthesis of phospholipases A2 inhibitors that cross the blood-brain barrier without harm may be useful for the treatment of acute neural trauma and neurodegenerative diseases. PMID- 15662833 TI - Discovering the mechanisms of neurodegeneration in prion diseases. AB - The purpose of my chapter in this issue of Neuroscience Reviews dedicated to Dr. Lawrence Eng is to summarize my contributions to understanding the mechanisms of neurodegeneration in prion diseases. I explain that I was able to advance the field of prion disease neuropathology largely because of the foundation of neurochemistry and immunohistochemistry that I learned while working 5 years in Dr. Eng's laboratory. In my review, I relate how my Neuropathology Research Laboratory began as a collaboration with Dr. Stanley Prusiner 20 years ago that led from immunohistochemical staining of amyloid plaques in rodent and human brains using prion protein-specific antibodies to molecular evidence that the abnormal prion protein, PrP(Sc), is the cause of the clinically relevant neuropathological changes in animal and human prion diseases. PMID- 15662834 TI - Cryptic axonal antigens and axonal loss in multiple sclerosis. AB - Axonal loss is well correlated with functional deficits in Multiple Sclerosis (MS); however, the molecular mechanisms that underlie this axonal loss are not understood. In this review we summarize evidence that antibodies to axolemma enriched fractions (AEF) isolated from CNS myelinated axons may play a role in axonal destruction. AEF contains potent antigens that elicit high-titer antisera, which destroy neurites in vitro, prevent neurite outgrowth, cause reactive changes in the neuronal cell bodies of origin and prevent myelination. We propose that these AEF antigens are cryptic because they are shielded from immune surveillance in vivo via the tightly sealed paranodal loops of myelin. Antibodies to AEF are found in cerebrospinal fluid (CSF) and sera of MS patients at higher levels compared with CSF or sera derived from patients with other neurological diseases. The potential identity of these cryptic antigens and their role in the axonal destruction characteristic of MS is discussed. PMID- 15662835 TI - Estrogen as a multi-active neuroprotective agent in traumatic injuries. AB - In the past decade, research has demonstrated that estrogen's role in physiology and development is far more complicated than previously assumed. Among these discoveries, there has been an increased recognition of the impact estrogen has in neurodevelopment, central nervous system physiology, and neuropathophysiology. These observations have led many researchers to consider using estrogen pharmacotherapeutically, at physiologic or supraphysiologic doses, for a variety of injury and toxicity models. In this short review, the effects of estrogen as an anti-apoptotic agent, as an anti-oxidant, and as an anti-inflammatory agent are discussed. Finally, the possibility of using estrogen as a neuroprotectant in neurotrauma is addressed. PMID- 15662836 TI - Zippers make signals: NCAM-mediated molecular interactions and signal transduction. AB - The neural cell adhesion molecule, NCAM, is involved in multiple cis- and trans homophilic interactions (NCAM binding to NCAM) thereby facilitating cell-cell adhesion through the formation of zipper-like NCAM-complexes. NCAM is also involved in heterophilic interactions with a number of proteins and extracellular matrix molecules. Some of these heterophilic interactions are mutually exclusive, and some interfere with or are dependent on homophilic NCAM interactions. Furthermore, both homo- and heterophilic interactions are modulated by posttranslational modifications of NCAM. Heterophilic NCAM-interactions initiate several intracellular signal transduction pathways ultimately leading to biological responses involving cellular differentiation, proliferation, migration and survival. Both homo- and heterophilic NCAM-interactions can be mimicked by synthetic peptides, which can induce NCAM-like signalling, and in vitro and in vivo studies suggest that such NCAM mimetics may be used for the treatment of neurodegenerative disorders. PMID- 15662837 TI - Signaling from nucleotide receptors to protein kinase cascades in astrocytes. AB - In the CNS, extracellular ATP can function as an excitatory neurotransmitter as well as a trophic factor. These short-term and long-term actions are mediated by nucleotide receptors. Extracellular ATP can also act as a co-mitogen in conjunction with polypeptide growth factors such as basic fibroblast growth factor (FGF2). Cellular proliferation, differentiation and survival are regulated by signaling cascades composed of protein kinases, including extracellular signal regulated protein kinase (ERK) and protein kinase B (also called Akt). Here we summarize recent studies on nucleotide receptor signaling to ERK and Akt in astrocytes and the role of protein kinase cascades in mediating the trophic actions of extracellular ATP, alone or together with FGF2. Because extracellular ATP and FGF2 contribute to the hyperplastic and hypertrophic response of astrocytes to CNS injuries, an understanding of their protein kinase signaling mechanisms may lead to novel therapeutic approaches for neurological conditions that involve gliosis and the generation of reactive astrocytes, such as trauma, stroke, seizure and neurodegenerative and demyelinating disorders. PMID- 15662838 TI - The adhesive role of acetylcholinesterase (AChE): detection of AChE binding proteins in developing rat spinal cord. AB - Acetylcholinesterase (AChE) is expressed by dorsal root ganglion (DRG) neurons during developmental periods when their central axons are growing into and through the spinal cord. Importantly, our previous studies have shown that AChE induces DRG axonal outgrowth by an adhesive mechanism and thus, have now employed a blot overlay technique to screen for potential AChE binding proteins in the developing spinal cord. Our results show that: (1) AChE binds to proteins with apparent molecular weights of 200, 110, 35, and 33 kDa; (2) these proteins are developmentally expressed during periods of axonal outgrowth from DRG neurons; (3) all four proteins are synthesized by astrocytes; and (4) AChE binding to these proteins is highly dependent on ionic strength supporting an electrostatic mechanism of adhesion. Taken together, these data provide further documentation for the participation of AChE in adhesive interactions during morphogenesis of the central nervous system and suggest a role for astrocytes in regulating AChE mediated axonal growth. PMID- 15662839 TI - Combined effects of ammonia and manganese on astrocytes in culture. AB - Ammonia has been strongly implicated in the pathogenesis of hepatic encephalopathy (HE), and astrocytes appear to be the primary target of ammonia neurotoxicity. Recent work has shown that manganese also plays a role in the pathogenesis of HE and causes astrocyte morphologic and functional changes similar to ammonia. We therefore investigated whether a combination of these compounds could produce additive/synergistic effects. Cultured astrocytes treated with 5 mM ammonia (NH4Cl) along with 100 microM manganese acetate (MnAc) for 3 h showed a 55-65% increase in free radical production over ammonia or manganese alone (P < 0.05). There was also a 50% decrease in the mitochondrial membrane potential (beta psi(m)) at 24 h following treatment with NH4Cl (5 mM) plus MnAc (50 microM) (P < 0.05), as compared to ammonia or manganese alone. Astrocytes treated with ammonia or manganese alone for 24 h showed no cell death, as determined by LDH release and light microscopic examination. However, cultures treated with ammonia plus manganese showed 80-90% necrotic cell death as estimated by light microscopy and 59% cell death as determined by LDH release. LDH release by ammonia plus manganese was blocked by the antioxidant superoxide dismutase (25 units/ml) as well as by the nitric oxide synthase inhibitor N(omega)-nitro-L-argininemethyl ester (500 microM). In conclusion, ammonia plus manganese exert additive/synergetic effects on the induction free radicals, mitochondrial inner membrane depolarization and cellular integrity, which may contribute to the tissue injury associated with chronic forms of HE. PMID- 15662840 TI - Metabolism of [U-(13)C]leucine in cultured astroglial cells. AB - Leucine is rapidly metabolized in astroglial primary cultures. Therefore, it is considered as valuable fuel in brain energy metabolism. Only few of the leucine metabolites generated and released by astroglial cells have been identified. Therefore, a more detailed study was performed analyzing by NMR techniques the 13C-labeled metabolites, which were released by astroglial primary cultures during the degradation of [U-(13)C]leucine. Confirming a former radioactive study this analysis revealed 13C-labeled 2-oxoisocaproate and ketone bodies. Additionally, 13C-labeled alanine, citrate, glutamine, lactate and succinate were identified. Their detailed isotopomer analysis proves that 13C-labeled acetyl-CoA enters the tricarboxylic acid cycle, that intermediates with a characteristic 13C labeling pattern can be withdrawn at several positions of the cycle, and that in the case of lactate and alanine there appears to be a participation of an active phosphoenolpyruvate carboxykinase and/or malic enzyme pathway. Thus, astroglial cells generate and release into the extracellular fluid not only the leucine catabolites 2-oxoisocaproate and ketone bodies, but also several tricarboxylic acid cycle dependent metabolites. PMID- 15662841 TI - Increased cell proliferation and neurogenesis in the hippocampal dentate gyrus of old GFAP(-/-)Vim(-/-) mice. AB - In response to central nervous system (CNS) injury, and more discretely so also during aging, astrocytes become reactive and increase their expression of the intermediate filament proteins glial fibrillary acidic protein (GFAP) and vimentin. Studies of mice deficient in astrocytic intermediate filaments have provided insights into the function of reactive gliosis. Recently we demonstrated robust integration of retinal transplants (1) and increased posttraumatic synaptic regeneration (2) in GFAP(-/-)Vim(-/-) mice, suggesting that modulation of astrocyte activity affects the permissiveness of the CNS environment for regeneration. Neurogenesis in the adult mammalian CNS is restricted to essentially two regions, the hippocampus and the subventricular zone. Here, we assessed neurogenesis in the hippocampus of 18-month-old GFAP(-/-)Vim(-/-) mice. In the granular layer of the dentate gyrus, cell proliferation/survival was 34% higher and neurogenesis 36% higher in GFAP(-/-)Vim(-/-) mice than in wildtype controls. These findings suggest that the adult hippocampal neurogenesis in healthy old mice can be increased by modulating astrocyte reactivity. PMID- 15662843 TI - Analyses of proteolipid protein mutants show levels of proteolipid protein regulate oligodendrocyte number and cell death in vitro and in vivo. AB - Previous tissue culture studies indicate that the level of native proteolipid protein (PLP) or mutated PLP regulates the number of oligodendrocytes (Olgs). The regulation of Olg number is most likely due to toxicity of over-expression of native PLP or mis-sense mutations of PLP. We tested, in vivo and in vitro, the hypothesis that the absence of native PLP or reduced amounts of mutated PLP leads to an increase in numbers of Olgs and a corresponding decrease in the number of apoptotic Olgs. In cultures derived from PLP deficient mice, the number of Olgs is twofold greater than in wild-type mice. In primary glial cultures or in enriched OLG cultures, in which the synthesis of native PLP is blocked using antisense technology, the number of apoptotic cells is several-fold reduced. Injection of PLP antisense oligodeoxynucleotides into jimpy (jp) mice reduces the number of dying glia in spinal cord 3x compared to controls, and increased the number of myelinated fibers. These studies demonstrate that inhibition of native or mutant PLP synthesis directly reduces apoptosis. The regulation of apoptosis by PLP gene expression occurs independently of myelination, indicating that the PLP gene has multiple primary functions. PMID- 15662842 TI - Expression specificity of GFAP transgenes. AB - Glial fibrillary acidic protein (GFAP) is an intermediate filament protein found predominantly in astrocytes. This specificity has recommended the GFAP gene promoter for targeting transgene expression to astrocytes. Although both we [Brenner et al. J. Neurosci. 14:1030-1037, (1994)] and others [Mucke et al. New Biol. 3:465-474, (1991)] have reported astrocyte specificity for GFAP promoters, we demonstrate here that these DNA sequences can also direct activity in neurons. The pattern of neuronal activity varied with both the nature of the expressed sequence and the transgene insertion site. Specifically, neuronal expression was very high for a protective protein/cathepsin A minigene, moderate for lacZ and undetectable for GFP. These findings, coupled with a survey of the literature, recommend that investigators using GFAP-driven transgenes verify specificity for each line studied, using a detection system whose sensitivity is sufficient to detect a compromising level of misexpression. PMID- 15662844 TI - Resveratrol protects against neurotoxicity induced by kainic acid. AB - Increased oxidative stress has been implicated in the mechanisms of excitotoxicity in hippocampus induced by kainic acid (KA), an excitatory glutamate receptor agonist. Resveratrol, a polyphenolic antioxidant compound enriched in grape, is regarded as an important ingredient in red wine to offer cardiovascular and neural protective effects. This study was designed to investigate whether resveratrol treatment may ameliorate neuronal death after KA administration. Adult Sprague Dawley male rats were treated with KA (8 mg/kg) daily for 5 days and another group was treated similarly with KA plus resveratrol (30 mg/kg/day). Three hr after the last treatment protocol, animals were sacrificed, and brain sections were obtained for histochemical and immunohistochemical identification of neurons, astrocytes and microglial cells. After KA administration, significant neuronal death and activation of astrocytes and microglial cells were observed in the hippocampal CA1, CA3 and polymorphic layer (hilar) of the dentate gyrus (DG) (P < 0.001). The KA-induced hippocampal neuronal damage was significantly attenuated by treatment with resveratrol (P < 0.001). Resveratrol also suppressed KA-induced activation of astrocytes and microglial cells. Since increased oxidative stress is a key factor for KA-induced neurotoxicity, this study demonstrated the ability of resveratrol to act as free radical scavenger to protect against neuronal damage caused by excitotoxic insults. PMID- 15662845 TI - NMDA receptor in nucleus accumbens is implicated in morphine withdrawal in rats. AB - The purpose of the present study is to elucidate whether ketamine, a non competitive antagonist of the NMDA receptor, can suppress the morphine withdrawal syndrome in rats at a dose without affecting motor functions and to identify its site of action in the central nervous system. Rats were made dependent on morphine by multiple injections of morphine hydrochloride for 5 days. They were then given ketamine at the following doses and routes of administration: (a) intraperitoneal (i.p.) injections (2-16 mg/kg), (b) intracerebroventricular (i.c.v.) injections (4-100 microg), and (c) intra-nucleus accumbens (NAc) or intra-amygdalar microinjections (0.4-10 microg). Naloxone HCl (1 mg/kg, i.p.) was administered 3 h after the last ketamine injection to precipitate withdrawal syndrome, which was scored within a period of 30 min. Results showed that some of the precipitated withdrawal signs were dose-dependently suppressed by repeated injections of ketamine at 8 and 16 mg/kg, i.p. or 100 microg, i.c.v. Dose dependent suppression was observed by repeated microinjections (0.4-10 microg) of ketamine to NAc, but not to amygdala. These results indicate that the NMDA receptor antagonist ketamine has the ability to suppress morphine withdrawal syndrome in experimental settings without motor interference, and NAc could be the critical CNS site mediating such effect. PMID- 15662846 TI - Analysis of glutamine accumulation in rat brain mitochondria in the presence of a glutamine uptake inhibitor, histidine, reveals glutamine pools with a distinct access to deamidation. AB - Rat cerebral nonsynaptic mitochondria were incubated in medium containing 2 mM glutamine (Gln) or 2 mM glutamate (Glu), in the presence of a Gln uptake inhibitor histidine (His) as well as other basic amino acids, lysine and arginine (Lys, Arg) not inhibiting Gln uptake. Subsequently, the mitochondrial contents of Glu and Gln were determined by HPLC. Incubation in the presence of Glu alone increased the Glu content from approximately 3.5 to 15 nmol/mg protein, without affecting the Gln content. On the other hand, incubation with Gln increased the content of Gln from approximately 1.5 to approximately 12 nmol/mg, and that of Glu to 10 nmol/mg. As expected, addition of His did not alter the Glu and Gln content resulting from incubation with Glu. However, His significantly decreased to almost the preincubation level the content of Glu in mitochondria incubated with Gln, without affecting the content of Gln. No other amino acid had any effect on these parameters. The results point to the existence of distinct Gln pools, one of which is accessible to external Gln via a His-sensitive transporter and is accessible for deamidation in the mitochondria. PMID- 15662847 TI - Sodium butyrate induces apoptosis in MSN neuroblastoma cells in a calcium independent pathway. AB - Sodium butyrate (NaBt), a histone deacetylase inhibitor, can cause apoptosis in a number of cancer cells. However, the mechanism of this action is poorly understood. Increased intracellular [Ca(2+)] level has been suggested as a likely mechanism, but there is little corroborating data. In this report we provide evidence that NaBt-treated MSN neuroblastoma cells undergo massive apoptosis in the presence of serum and regardless of external or internal [Ca(2+)] levels. Presented data suggest that apoptotic effect of NaBt is both time- and dose dependent (LD50 1 mM); and that, presence of serum or cAMP, a second messenger molecule that modulates the apoptotic program in a wide variety of cells could not circumvent the apoptotic effect of NaBt. Our findings suggest that NaBt induced apoptosis in MSN neuroblastoma cells occurs via a pathway that is independent of Ca(2+) flux, intracellular [Ca(2+)] or cAMP levels. Further, we also present data that exclude a role for PKC or histones acetylation. PMID- 15662848 TI - Presence of sodium-calcium exchanger/GM1 complex in the nuclear envelope of non neural cells: nature of exchanger-GM1 interaction. AB - Previous studies have revealed the presence of Na+ / Ca2+ exchanger (NCX) activity associated with GM1 ganglioside in the nuclear envelope (NE) of neurons and glia as well as various neural cell lines. The nuclear NCX1 exchanger, unlike that in the plasma membrane, was shown to be tightly associated with GM1 and potentiated by the latter. One non-neural cell line, Jurkat, was found to contain no Na+ / Ca2+ exchanger of the NCX1, NCX2, or NCX3 types in either nuclear or plasma membrane. To determine whether such absence in the NE is generally characteristic of non-neural cells we have examined two more such cell lines in addition to human lymphocytes. RT-PCR showed NCX1 expression in both HeLa and NCTC cell lines and also NCX2 in the latter; NCX3, a subtype previously observed in NG108-15 cells, was not expressed in either. Immunocytochemical and immunoblot studies indicated NCX1 on the cell surface and nuclear envelope of both cell types. Some alternatively spliced isoforms of NCX1 in the nuclear envelope of both cell types were tightly associated with ganglioside GM1. Human lymphocytes, a mixed population of T and B cells, showed similar evidence for plasma membrane and nuclear expression in some but not all cells. The high affinity association between NCX1 and GM1, explored by reaction with base, acid, and proteases, was found to involve charge-charge interaction with a requirement for a positively charged moiety in NCX. PMID- 15662849 TI - Lack of apparent neurological abnormalities in rabbits sensitized by gangliosides. AB - Two very high titer polyclonal antibodies against two ganglioside antigens, GM1 and GD1a, have been raised in New Zealand white rabbits using a homogeneous suspension of the highly purified antigens in Keyhole Limpet Hemocyanin and Freund's adjuvant. The antisera were prepared over a period of 6 months with repeated injections of the ganglioside suspension, followed by an intravenous injection of the purified ganglioside solution, and collecting the serum (approximately 50 ml) at defined time intervals. The GM1-antibody, thus prepared, showed a cross reactivity toward GDlb and asialo-GM1 (GA1), while the GDla antibody reacted with GD1a, GM1 and GA1 and GD1b as determined by immuno-overlay and ELISA methods. The titer for GM1 antiserum, determined by'ELISA, was greater than 1/10,000 dilution while the titer for GD1a antibody was greater than 1/5000 dilution. No neurological or behavioral abnormality was observed during the period of antiserum production. To evaluate any likely pathological damage caused by such a high titer ganglioside-antibody, autopsy of CNS as well PNS tissues from the rabbits were carried out after the final bleeding. No obvious pathological changes, including demyelination, were noted in any of the four rabbits. These observations cast doubt as to the direct effect of anti ganglioside antibody induced neurological and pathological disorders. PMID- 15662850 TI - Cytochrome c release and caspase-3 activation in retinal ganglion cells following different distance of axotomy of the optic nerve in adult hamsters. AB - This study examined the relationship between the distance of axotomy and the death of injured retinal ganglion cells (RGCs) in adult hamsters and the relationship of cytochrome c and caspase-3 on the death pathway of RGCs. The left optic nerve (ON) of adult hamsters was transected either at 1 or 3 mm away from the optic disc, and retrogradely labeled with Flurogold on the ON stump. After a predetermined period of postoperative time, the surviving RGCs were counted by retina flat-mount, and the activation of cytochrome c and caspase-3 were investigated by immunohistochemistry. Cell loss was found to be much faster (P < 0.01), more cells with cytochrome c were observed (P < 0.05) and the activation of caspase-3 was earlier when ON was transected 1 mm away from the optic disc than when was transected 3 mm away from the optic disc. Distance of axotomy affects the axotomized cell death rate where more RGCs died when the ON transection was applied closer to the eye. The timing of activation of caspase-3 in the RGCs may be linked to the distance of axotomy. PMID- 15662851 TI - Anoxia effects on CNS function and survival: regional differences. AB - The effects of anoxia on function and survival of different central nervous system (CNS) areas were tested. As expected, synaptic function in a typical gray matter area of the brain, hippocampus, failed rapidly during 30 min of anoxia and did not recover. Mouse optic nerve and corpus callosum, two white matter (WM) areas of the brain, showed persistent function during total anoxia for periods as long as two hours. Moreover, even after two hours of anoxia followed by a recovery period, nearly half of the axons that were excitable at the outset remained functional. The corpus callosum contains a high percentage of unmyelinated axons while optic nerve axons are completely myelinated. These studies indicate that CNS structures vary greatly in their ability to function and survive anoxia. Mammalian WM, independent of myelination, is remarkably tolerant of anoxia implying that CNS axons generate enough ATP by anaerobic energy metabolism to sustain function. PMID- 15662852 TI - A model to induce low temperature trauma for in vitro astrogliosis study. AB - Astrogliosis is an inevitable and rapid response of astrocytes to physical, chemical and pathological injuries. To study astrogliosis, we developed a reproducible in vitro model in which low temperature injury to cultured astrocytes could be induced by placing the culture dish onto a copper pipe pre cooled by liquid nitrogen. Using this model, the relationship between the temperature decline and the severity of cellular damage was analyzed. An increase in the expression of some known injury-related proteins, such as glial fibrillary acidic protein (GFAP), immediate early response genes (IEGs), and heat shock proteins 70 (HSP70), was demonstrated in astrocytes after low temperature trauma. With the use of this low temperature trauma model, the flexibility in the temperature control and injury area may allow researchers to evaluate cryotherapy and cryosurgery, which could be applicable to future development of quality health care. PMID- 15662853 TI - An enzyme-linked immunosorbent assay for the rapid quantification of intracellular and extracellular guanosine 3',5'-cyclic monophosphate in cultured glial cells. AB - A sensitive enzyme-linked, indirect immunosorbent assay (ELISA) for the determination of guanosine 3',5'-cyclic monophosphate (cGMP) in glial cells is described. The assay uses an antiserum produced against succinylated cGMP and is based on the competition between endogenous cGMP and a fixed amount of immobilized antigen. The antibody exhibited a high degree of specificity with negligible cross reactivity with other nucleotides or related compounds. The standard curve, linearized using a logit-log transformation, had an operating range of 1 fmol/50 microl to 5 pmol/50 microl. The sensitivity of the assay was significantly increased by acetylation of standards and samples. Recoveries of cGMP from samples of cultured cells ranged from 85% to 105% with a mean recovery of 97% +/- 7%. Levels of cGMP measured with the ELISA were in agreement with the corresponding values obtained using radioimmunoassay. The present method provides for a cheap, sensitive and simple alternative to the commercially available cGMP assays. PMID- 15662854 TI - Mobil refinery proudly waves flag as an OSHA 'Star' VPP participant. Labor management commitment and cooperation resolved hazards, changed attitudes. PMID- 15662856 TI - Is your chemical protective clothing quickly becoming landfill material? Users need to know how and when to dispose of or decontaminate their CPC. PMID- 15662855 TI - 'What's in a name?'--OSHA policy may find database a tough obstacle. Can the agency demonstrate repeat violations based on insufficient information? PMID- 15662857 TI - Can you take the heat? Any strenuous job in hot, humid environments runs the risk of heat stress and a general-duty clause citation. PMID- 15662858 TI - Combined chemical, physical hazards make exposures harder to calculate. Studies show multiple exposures must be evaluated wholly rather than 'adding up' individual risks. AB - Clearly, current research adds to the mounting scientific evidence that combined exposures can produce greater toxicity than would be predicted using an additive model. Although there is not yet a quantitative method to determine the risks associated with multiple exposures, safety and health specialists must begin to consider the health effects from the entire work process, rather than individual chemical or physical agents. Moreover, organizations involved in risk assessment and standard-setting must evaluate the interactive effects of combined chemical and physical agent exposures. This will make personnel exposure standards comprehensive in addressing realistic occupational health risks. PMID- 15662859 TI - Software scrutiny keeps paper trail from becoming 'Dos error' avalanche. How compatible is the cost and the system with your long-term record processing needs? PMID- 15662860 TI - Readily-avaliable methods estimate air flow for HVAC. PMID- 15662862 TI - Tuberculosis revival forces health care to implement strict preventive policies. New guidelines will affect every health-care setting, whether private or municipal, paid or volunteer. PMID- 15662861 TI - Back injury prevention in health care requires training techniques, exercise. Lifting teams, job analysis, ergonomics help trim the risks. PMID- 15662863 TI - Spirometers' technological evolution spurs increased confidence in results. Four generations of advances enable more useful diagnoses for asthma, emphysema or tuberculosis. AB - A century and a half of spirometry testing has seen the most common lung disease change from tuberculosis which reduces lung volumes to emphysema and asthma which reduce flow rates. Spirometers have evolved to highly accurate, relatively inexpensive hand-held instruments. These new spirometers use disposable flow sensors which eliminate the chance of cross-contamination and have microprocessors with programs which monitor and report the quality of the maneuvers as they are performed. Such improvements enable this highly useful diagnostic tool to be used more widely and with greater confidence. PMID- 15662864 TI - Operationally defined training helps employees achieve adaptive readiness. A routine inventory of "critical" skills establishes the foundation for safe behaviors. PMID- 15662865 TI - Measures of security. PMID- 15662866 TI - OSHA releases ergonomics draft. PMID- 15662867 TI - Stop spread of tuberculosis in hospitals. PMID- 15662868 TI - ACGIH's threshold limit values useful, but formulas are still controversial. Toxicologic methods in determining TLVs, scientific adequacy and political factors questioned. PMID- 15662869 TI - A safe haven from violence emerges as another duty. AB - Education is the key to avoiding violence in any situation. Safety personnel must become more knowledgeable of security procedures and how to handle situations which could lead to violence. Managers and employees must be trained to respond properly to a variety of stress, personnel and security problems. The safety manager's role in providing a safe haven for workers in a violent society is critical to the employees' morale and well-being. Employees who are not consumed with their own personal safety will perform better and add to the company's bottom line. Safety must be allowed to evolve to include the effects of ever changing societal issues which will continue to challenge loss prevention in the workplace. PMID- 15662870 TI - Employee focus groups cultivate openness about company policies. Informal, confidential discussions can prove helpful in workplace health planning. PMID- 15662871 TI - Simple procedures estimate airflow through HVAC units, occupied spaces. Approximations of fresh outdoor air can be achieved using temperature and O2 monitors. PMID- 15662872 TI - Behavior-based involvement recognized in OSHA voluntary protection programs. Meaningful employee involvement satisfies OSHA and helps the corporate bottom line. PMID- 15662873 TI - OSHA revises, rewords hazcom rule to foster knowledge about hazards. Hazard communication violations are common, but changes could make it easier to comply. PMID- 15662874 TI - New rule mandates hazard assessment to specify protective equipment needs. 1994 revisions clarify requirements for PPE in contemporary work environments. PMID- 15662876 TI - Falling into place. Panel hopes negotiated rulemaking will acheive consensus in steel erection industry. PMID- 15662875 TI - Accommodation between a rock and a hard place. Case by case, the ADA addresses back impairment and fights workers' comp malingerers by keeping the disabled at work and productive. PMID- 15662877 TI - Skin care plan involves more than washing hands. Hygiene strategy counters spread of dermatitis among employees. PMID- 15662878 TI - Listening in. Voice communication plays an important part in the safety, well being and productivity of those who work in permit-required confined spaces. PMID- 15662879 TI - Closed-loop power and focus control of laser welding for full-penetration monitoring. AB - We describe a closed-loop control system ensuring full penetration in welding by controlling the focus position and power of a 4-kW Nd:YAG laser. A focus position monitoring system was developed based on the chromatic aberration of the focusing optics. With the laser power control system we can determine the degree of penetration by analyzing the keyhole image intensity profile. We demonstrate performance in bead-on-plate welding of Inconel 718 and titanium. The focus control system maintained a focal position on tilted and nonflat workpieces, and the penetration monitoring technique successfully controlled the laser power to maintain the full-penetration regime in the presence of linear and step changes of thickness. Finally we discuss the performances and the limits of the systems when applied to a realistic complex aerospace component. PMID- 15662880 TI - Versatile in-fiber sensing by use of core-replaced D-fiber. AB - We present a method of replacing a section of the core of a D-shaped optical fiber with sensing materials as a platform for various extrinsic fiber sensors. In this configuration light guides within the sensing material allow for strong interaction between the sensing material and the optical field. Initial experimental results indicate that replacing the fiber core with polymer enhances its temperature sensitivity by at least a factor of 5. The new technique is promising as a means for incorporating various sensing materials into the path of a beam traveling in an optical fiber. PMID- 15662881 TI - Detection of surface strain by three-dimensional digital holography. AB - Three-dimensional digital holography with three object-illuminating beams has been successfully used for the detection of surface strain in metallic objects. The optical setup that uses illuminating beams to irradiate the object from three directions means that all three object surface displacement components, x, y, and z, can be independently calculated and used to find the strain gradients on the surface. The results show the conversion of the complete surface displacement field into a surface strain field. The method is capable of measuring microstrains for out-of-plane surface displacements of less than 10 microm. PMID- 15662883 TI - Spatial self-filtering with polarizer sheets. AB - A method for phase visualization and edge enhancement by spatial self-filtering by use of a polarizer sheet in the Fourier plane of an optical processor is described. Light absorbed by the polarizer sheet induces a thermal lens, which, in turn, produces selective action on certain spatial frequencies of the image to be processed. Some experiments that demonstrate the self-filtering action of the proposed system are presented. PMID- 15662882 TI - Geometric superresolution by code division multiplexing. AB - In many highly resolved optical systems the resolution is limited not by the optics but by the CCD's nonzero pixel size. As a result, overall resolution is decreased. Here we propose a novel approach to enhancing resolution beyond the limit set by the CCD's pixels. This method does not involve additional mechanical elements, such as those used for microscans. In this scheme neither the CCD nor additional elements are moved. The geometric superresolving procedure is based on code-division multiplexing, with all its inherent benefits, such as relative noise immunity to single-tone interference. A setup is proposed for coherent and incoherent illumination, with slight modifications for the latter. A theoretical analysis of the setup is presented and compared with empirical results. This scheme is shown to enhance one-dimensional image resolution with the use of only a simple mask that doubles image resolution. This method can easily be expanded to two-dimensional images and to resolution-enhancement factors greater than 2. PMID- 15662884 TI - Application of supergeneralized matched filters to target classification. AB - The matched filter (MF) is the optimum linear operator for distinguishing between a fixed signal and noise, given the noise statistics. A generalized matched filter (GMF) is a linear filter that can handle the more difficult problem of a multiple-example signal set, and it reduces to a MF when the signal set has only one member. A supergeneralized matched filter (SGMF) is a set of GMFs and a procedure to combine their results nonlinearly to handle the multisignal problem even better. Obviously the SGMF contains the GMF as a special case. An algorithm for training SGMFs is presented, and it is shown that the algorithm performs quite well even for extremely difficult classification problems. PMID- 15662885 TI - Comment on "Redundancy of phase-space distribution functions in complex field recovery problems". AB - In a recent publication [Appl. Opt. 42, 1932 (2003)], redundancies of phase-space representations were studied. In particular, signal recovery from a single section of the ambiguity function was explored. It is shown that this signal recovery method can be associated with interferometry. PMID- 15662886 TI - Modulation effect of the atmosphere in a pyramid wave-front sensor. AB - The pyramid wave-front sensor in its original form works with a mechanical modulation that adapts the linear range of the sensor to seeing and sensing conditions. For adaptive optics systems working in an astronomical context, the way in which the aberrations produced by the atmospheric turbulence, which are not seen by the sensor owing to its limited temporal bandwidth, act as modulators is shown. These aberrations have the same effect of increasing the linear range and localizing the measurement as does mechanical modulation. The effect of residual wave-front aberrations is estimated for some example conditions of telescope diameter, system bandwidth, wind velocity, and Fried parameter. PMID- 15662887 TI - Alternative to the effective transmittance approach for the calculation of polychromatic transmittances in rapid transmittance models. AB - Many current rapid transmittance algorithms, specifically the Optical Path Transmittance (OPTRAN), are based on use of effective transmittances to account for the effects of polychromatic radiation on the transmittance calculations. We document how OPTRAN was modified by replacing the effective transmittance concept with a correction term. Use of the correction term solves some numerical problems that were associated with use of effective transmittances, greatly reduces the line-by-line computational burden, and allows for the efficient inclusion of more gases. This correction method can easily be applied to any other fast models that use the effective transmittance approach. PMID- 15662888 TI - Polarized erbium-doped superfluorescent fiber source utilizing double-pass backward configuration. AB - An erbium-doped superfluorescent fiber source utilizing a double-pass backward configuration is analyzed when a polarizer is inserted into an erbium-doped fiber to obtain polarized output light. Such a polarized configuration is simulated and experimentally confirmed to have the following characteristics: high polarization power conversion efficiency, pump-power-independent mean-wavelength operation, and low sensitivity to polarizer insertion loss. PMID- 15662889 TI - Performance of two-stage discrete fiber Raman amplifiers with and without all optical gain clamping. AB - Using numerical simulations, we analyze the properties of two-stage discrete fiber Raman amplifiers without and with all-optical gain clamping. In both cases a two-stage amplifier can be designed to have the same gain characteristics as a single-stage amplifier with improved noise performance by use of either the same total length of gain fiber (but with increased pump power) or total pump power (but with increased total length of gain fiber). PMID- 15662890 TI - Lightweight diode laser spectrometer CHILD (Compact High-altitude iN-situ Laser Diode) for balloonborne measurements of water vapor and methane. AB - A new lightweight near-infrared tunable diode laser spectrometer CHILD (Compact High-altitude In-situ Laser Diode spectrometer) was developed for flights to the stratosphere as an additional in situ sensor on existing balloonborne payloads. Free-air absorption measurements in the near infrared are made with an open-path Herriott cell with new design features. It offers two individual absorption path lengths optimized for CH4 with 74 m (136 pass) and H2O with 36 m (66 pass). New electronic features include a real-time gain control loop that provides an autocalibration function. In flight-ready configuration the instrument mass is approximately 20 kg, including batteries. It successfully measured stratospheric CH4 and H2O profiles on high-altitude balloons on four balloon campaigns (Environmental Satellite validation) between October 2001 and June 2003. On these first flights, in situ spectra were recorded from ground level to 32,000-m altitude with a sensitivity of 0.1 ppm [(parts per million), ground] to 0.4 ppm (32,000 m) for methane and 0.15-0.5 ppm for water. PMID- 15662891 TI - Experimental demonstration of multiple pulse nonlinear optoacoustic signal generation and control. AB - Generating underwater acoustic signals from a remote, aerial location by use of a high-energy pulsed infrared laser has been demonstrated. The laser beam is directed from the air and focused onto the water surface, where the optical energy was converted into a propagating acoustic wave. Sound pressure levels of 185 dB re microPa (decibel re microPa) were consistently recorded under freshwater laboratory conditions at laser-pulse repetition rates of up to 1000 pulses/s. The nonlinear optoacoustic transmission concept is outlined, and the experimental results from investigation of the time-domain and frequency-domain characteristics of the generated underwater sound are provided. A high repetition rate, high-energy per pulse laser was used in this test under freshwater laboratory conditions. A means of deterministically controlling the spectrum of the underwater acoustic signal was investigated and demonstrated by varying the laser-pulse repetition rate. PMID- 15662892 TI - Optical frequency measurements with the global positioning system: tests with an iodine-stabilized He-Ne laser. AB - Global positioning system- (GPS-) referenced optical frequency combs based on mode-locked lasers offer calibrations for length metrology traceable to international length standards through the SI second and the speed of light. The absolute frequency of an iodine-stabilized He-Ne laser [127I2 R(127) 11-5 f component] was measured with a femtosecond comb referenced to a multichannel GPS timing receiver. The expected performance and limitations of GPS-referenced comb measurements are discussed. PMID- 15662893 TI - Imaging at 3.4 Thz with a quantum-cascade laser. AB - We have assembled a single-frequency imaging system at 3.4 THz with a quantum cascade laser. Images of electronic and biological applications are demonstrated. We operate the laser with a peak output power of 2.5 mW at a 7% duty cycle and a 22 K operating temperature. The minimum spot size is 340 microm. The signal is detected with a single-element deuterated triglycine sulfate detector, and images are captured by scanning of the sample. PMID- 15662894 TI - Evidence for wavelength dependence of the scattering phase function and its implication for modeling radiance transfer in shelf seas. AB - More than 90% of stations from the Irish and Celtic Seas are found to have significantly higher back-scattering ratios in the blue (470 nm) than in the red (676 nm) wave band. Attempts to obtain optical closure by use of radiance transfer modeling were least successful for stations at which backscattering ratios are most strongly wavelength dependent. Significantly improved radiance transfer simulation results were obtained with a modified scattering correction algorithm for AC-9 absorption measurements that took wavelength dependency in the scattering phase function into account. PMID- 15662895 TI - Comparative analysis of selected radiative transfer approaches for aquatic environments. AB - A comparative analysis is presented of simple approaches to radiative transfer in plane-parallel layers, such as the self-consistent Haltrin approach, the Chandrasekhar-Klier exact solution for isotropic scatters, an extended version of two-flux radiative Kubelka-Munk theory, the neutron-diffuse Gate-Brinkworth theory, and different versions of the delta-Eddington theory. It is shown that the Haltrin approach is preferable to others and can be used for the solution of an inverse optical problem of the estimation of absorption and backscattering coefficients of aquatic environments from measured apparent optical properties. Two different methods of transformation from measured irradiance reflectance at combined illumination to irradiance reflectance induced by diffuse illumination only are developed. An analysis of the use of the different models for estimation of the effect of the bottom albedo is also presented. PMID- 15662896 TI - Quantitative comparison of terahertz emission from (100) InAs surfaces and a GaAs large-aperture photoconductive switch at high fluences. AB - InAs has previously been reported to be an efficient emitter of terahertz radiation at low excitation fluences by use of femtosecond laser pulses. The scaling and saturation of terahertz emission from a (100) InAs surface as a function of excitation fluence is measured and quantitatively compared with the emission from a GaAs large-aperture photoconductive switch. We find that, although the instantaneous peak radiated terahertz field from (100) InAs exceeds the peak radiated signals from a GaAs large-aperture photoconductive switch biased at 1.6 kV/cm, the pulse duration is shorter. For the InAs source the total energy radiated is less than can be obtained from a GaAs large-aperture photoconductive switch. PMID- 15662898 TI - [Pain reduced and quality of life improved by sustained-release hydromorphone. Results of an observational study]. AB - METHOD: A total of 487 patients with severe pain of varying pathogenesis received sustained-release hydromorphone (Palladon) in an observational study. After an initial examination, two follow-ups took place on the 3rd and 7th day and a final examination after about 21 days. RESULTS: The intensity of the pain, measured using a numerical rating scale from 0 to 10 (none--most severe pain), decreased by 65.3% from 7.2 at the start to 2.5 at the final examination. The quality of life, characterized by the patients themselves assessing the variables of sleep, vitality, mood, social contacts, activity, resilience and walking ability, increased by 53.7% in comparison with the initial examination. Symptoms typical of opioids (tiredness, nausea, constipation and vomiting) were present in 65.4% of the patients at the start of the treatment with hydromorphone. On sustained release hydromorphone, with appropriate treatment a markedly smaller proportion of patients suffered from these accompanying symptoms. Both the efficacy and tolerability as well as patient compliance were assessed as "very good" or "good" by the doctors in more than 85% of cases. CONCLUSION: This observational study confirms the good pain-relieving efficacy of sustained-release hydromorphone together with very good tolerability and very good compliance. Patients assessed their own quality of life as markedly improved. PMID- 15662897 TI - [Six months treatment of restless legs syndrome with levodopa/benserazide in domiciliary practice]. AB - Restless legs syndrome is a common neurological condition with 2-10% prevalence in the general population. Patients suffer from uncomfortable or unpleasant sensations in legs or arms and motor restlessness with a compelling desire to move the extremities. The symptoms worsen at rest with at least temporary relief by activity but also later in the day and in the evening. Dopaminergic drugs are considered to be the first choice in therapy, however, only levodopa/benserazide is approved until now. In this six months observational study, eighty patients with diagnosed RLS were treated with levodopa/ benserazide. The RLS symptoms improved in 90%, 77.5% of the patients continued with the treatment after the end of the study. Levodopa/Benserazide proved to be efficacious independently of severity of RLS and was well tolerated in long-term therapy. PMID- 15662899 TI - [Improvement in quality of life in the elderly. Results of a placebo-controlled study on the efficacy and tolerability of lecithin fluid in patients with impaired cognitive functions]. AB - Lecithin, a precursor of the neurotransmitter acetylcholine, has a positive effect on brain and memory functions. In a prospective, randomized, double-blind study, the effect of buerlecithin fluid (BLF) was investigated in comparison with placebo in patients with mild cognitive disorders. A total of 96 ambulatory patients (> 55 years) were admitted to the study. Treatment duration was 84 days. In both treatment groups, a clear improvement in all the cognitive parameters tested was seen. The main target measure, the overall Sandoz Clinical Assessment Geriatric (SCAG) score improved by 18.7 (test substance) and 16.4 (placebo) points (p = 0.1620). A statistically relevant improvement of the secondary target parameter, response in the SCAG score, was achieved with BLF (85.4%) in comparison with placebo (62.5%) (p = 0.018). Furthermore, BLF demonstrated significant superiority in a number of the other target measures. The study also confirmed the very good tolerability of BLF. PMID- 15662900 TI - [Adult mesenchymal stem cells for the regeneration of musculoskeletal tissue]. AB - Mesenchymal stem cells can be isolated from human bone marrow aspirates, and, thanks to their differentiation potential and good in vitro culture properties, represent an attractive cell line for the replacement of mesenchymal tissue, i. e. tissue engineering. Both in vitro and in vivo, they can differentiate into cartilage, bone, tendons and fat cells, and--in contrast to embryonic stem cells- are not underethical scrutiny. Cultured in three-dimensions on a variety of carrier substances, they have already been successfully employed for the reconstruction of mesenchymal tissues in numerous studies involving both small and large animal models. While a clinical application in humans has also been successful in pilot studies, numerous points still remain to be clarified, underscoring the need for further intensive research before wide-scale clinical application can be contemplated. Only then can it be shown whether the associated high expectations are justified. PMID- 15662901 TI - [Safety and antihypertensive efficacy of the dihydropyridine calcium antagonist Amlodipine besylate--results from an observational study in 9,672 patients]. AB - OBJECTIVES: This observational study investigated prescription behavior and the efficacy and safety of Amlodipine besylate in hypertensive patients. METHODS: 9,672 hypertensive patients were treated with Amlodipine besylate 5-10 mg/day for 12 weeks. At the end of the observation period, the participating physicians were asked to give their subjective evaluation of the efficacy and safety of treatment with Amlodipine besylate as well as their therapeutic considerations regarding the use of Amlodipine besylate. RESULTS: 55.2% of the patients received Amlodipine besylate in combination with other antihypertensive agents. The mean baseline blood pressure was 168.0/96.7 mmHg and the mean blood pressure reduction achieved at the end of the observation period was -28.5/-14.5 mmHg. In 80.3% of the total population, a reduction to diastolic blood pressure values of < 90 mmHg was achieved. A reduction to systolic blood pressure values of < 140 mmHg was obtained in 41.6% of the patients. Adverse events were reported by only 1.2% of the patients. For the vast majority of the patients, efficacy (96.6%) and safety (98.9%) were rated "very good" or "good" by their physicians. 24-hour efficacy with once daily dosing was given as the most important argument for using Amlodipine besylate. CONCLUSION: The observation study confirmsthat Amlodipine besylate is an effective and safe antihypertensive drug both in mono and combination therapy. PMID- 15662902 TI - [S2 guideline for infectious endocarditis]. AB - Microbe-induced (infectious) endocarditis is an endovascular infection, caused mainly by bacteria, of cardiovascular structures. The major predilection site are the native heart valves, but involvement of implanted intracardiac foreign material is increasingly being seen. The mortality rate of infectious endocarditis depends on clinical factors and the causal agent, but also on the time of the establishment of the diagnosis and the initiation of appropriate treatment. In Germany, the current mortality rate ranges up to 18%. Between January 2003 and July 2004, with the aim of improving patient care and thus the outcome of this condition, a guideline commission worked out recommendations for the diagnosis, treatment and management of the disease for the use of general practitioners and hospital physicians, in particular microbiologists, infectiologists, cardiologists and cardiac surgeons. The basis for this guideline was the systematic search through the literature of the European guideline. On the 16th and 28th of June 2004, the entire guideline was formerly approved in a nominal group process. PMID- 15662903 TI - [Prenosological diagnosis in the environment-human health area]. PMID- 15662904 TI - [Ecological and hygienic markers in evidence-based medicine]. PMID- 15662905 TI - [A problem of health monitoring in preventive medicine]. PMID- 15662906 TI - [Health indicators in the problem of the human environment and environmental hygiene]. PMID- 15662907 TI - [Health indicators of children and adolescents in the sociohygienic monitoring system]. PMID- 15662908 TI - [Assessment of schoolchildren's adaptation under the conditions of iodine deficiency and ambient air pollution]. PMID- 15662909 TI - [Genetic methods for monitoring multifactorial diseases]. PMID- 15662910 TI - [Biochemical methods in the evaluation of the influence of environmental factors on organism]. PMID- 15662911 TI - [Biomonitoring of toxic substances in the human body]. PMID- 15662912 TI - [Monitoring of lifestyle]. PMID- 15662913 TI - [Monitoring of reproductive and genetic health]. PMID- 15662914 TI - [Assessment of children's health in the Aral Sea region]. PMID- 15662915 TI - [Monitoring of congenital anomalies and malformations in the industrial centers of the Sverdlovsk region]. PMID- 15662916 TI - [Monitoring of congenital malformations in the Altai Territory]. PMID- 15662918 TI - [Monitoring of bronchial asthma in the town of Vladimir]. PMID- 15662917 TI - [Ultrasound studies in human health monitoring]. PMID- 15662919 TI - [Specific features of children's health monitoring in the town of Kemerovo]. PMID- 15662920 TI - [Health indices in adolescent girls in the sociohygienic monitoring of the population of a large city on the lower Volga]. PMID- 15662921 TI - [The scientific bases of diagnosis of prenosological homeostatic disorders in case of chronic chemical loads]. PMID- 15662922 TI - [Hygienic diagnosis of the endoecological status]. PMID- 15662923 TI - [Prenosological changes in children from petrochemical regions]. PMID- 15662924 TI - [Evaluation of local immunity of respiratory tract mucosae in the diagnosis of prenosological entities]. PMID- 15662925 TI - [Noninvasive method of sociohygienic monitoring of children]. PMID- 15662926 TI - [Evaluation of premorbid changes in children]. PMID- 15662927 TI - [Study of genetic polymorphism and cytogenetic disorders upon chemical exposures]. PMID- 15662928 TI - [Evaluation of sensibilizers of the dwelling environment]. PMID- 15662929 TI - [Polymorphism of NAT2 gene in petrochemical workers]. PMID- 15662931 TI - [Resolution of the scientific-and-practical conference on problems of diagnosis or prenosological entities in preventive and clinical medicine]. PMID- 15662930 TI - [A new approach to solving the problem of prenosological diagnosis]. PMID- 15662932 TI - [Main results of researches by the state institution A. N. Sysin Research Institute of Human Ecology and Environmental Hygiene, Russian Academy of Medical Sciences, in 2003]. PMID- 15662933 TI - [Malaria: from genetic and molecular biology to disease control]. AB - The knowledge of the genomic structure of Plasmodium falciparum and of its main vector, Anopheles gambiae, may offer new perspectives for malaria therapy, vaccines or control of mosquito-borne transmission. New targets for future antimalarial drugs were identified, mainly apicoplast (a vestige of a vegetal structure incorporated by the parasite) and several enzymes, particularly proteases. The practical difficulty is now to select a few number of these "promising molecules", probably no more than 3 or 4, for a preclinical and clinical pharmaceutical development. Indeed, several other antimalarial drugs are already under development, and the industrial possibilities for developing new drugs are evidently limited. Many new vaccination targets and antigenic proteins were also identified. According to scientific and industrial limitations, a complete evaluation of these antigens is absolutely necessary to select a few of them for clinical development. For anti-malarial vaccinations, DNA vaccines may offer the most interesting perspectives, with the possibility of simultaneous immunisation against different Plasmodium stages and of an adjuvant effect by adding a gene encoding certain cytokines. In Anopheles gambiae genome, several genes encoding key-proteins (particularly odorant receptors necessary for blood feeding) were identified, as other genes encoding for proteins limiting the sexual development of Plasmodium inside its vector. From a theoretical viewpoint, genetically modified non biting or non transmitting mosquitoes offer new perspectives for the control of malaria transmission, but until now, the preliminary practical attempts gave rather poor results. On the whole, the genomic and proteomic of Plasmodium falciparum and Anopheles gambiae yielded exciting scientific results, but it is still too early and very speculative to imagine their practical applications for the control of malaria. PMID- 15662934 TI - [Cerebral malaria, a key role for endothelial cells?]. AB - Five to six hundred millions of people, throughout the world, suffer from malaria and more than one million die each year as a consequence, in about 20% of the cases, of cerebral malaria, an important complication of Plasmodium falciparum infection (Holding & Snow, 2001). Despite many studies, the physiopathology of these cerebral occurrences is not understood, especially concerning the intricacy and respective roles of the various mechanisms identified: sequestration of parasitized red cells in microvessels, cytokine secretion, changes in the T lymphocyte repertoire, host genetic factors driving sensitivity pathogenic factors from Plasmodium (Mazier et al., 2000). PMID- 15662935 TI - [Malaria research in the post-genomic era]. AB - Genomic sequence determination of Plasmodium falciparum and other species of the genus, as well as that of Anopheles gambiae, and human, rat and mouse genome sequencing have completely changed the landscape of fundamental research about malaria. These data should urgently be exploited, in order to develop new tools to combat the disease: new drugs, fine dissection of the cascade of events following infection of the various vector species and vertebrate host, analysis of the complex interaction leading to the pathology or, inversely, contributing to sustained protection. Powerful population biology tools are now available, allowing to investigate genetic exchanges within natural population and to identify factors structuring parasitic and vector populations. Nevertheless, important impediments persist, including the complexity of experimental systems and the unclear relevance of animals models. Numerous challenges are to be faced; they call upon a more efficient organisation of research efforts in the systematic explorations using the powerful novel post-genomic technologies, as well as the development of new tools and experimental models required by functional genomics and integrative biology. PMID- 15662936 TI - [Drug susceptibility of Plasmodium falciparum from polyclonal isolates]. AB - Msp-1 and Msp-2 genes, each present as a unique copy in the genome of Plasmodium, contain polymorphic repeats in bloc 2. We studied allelic polymorphism of Msp-1 and Msp-2 by amplifying bloc 2 with a fluorescent primer, and analysing the fragment generated. We validated this method by mixing two cloned strains: chloroquine-susceptible HB3-Honduras and chloroquine-resistant FCM29-Cameroon. This method was then used to quantify the clones in natural isolates of 19 infected persons during quinine treatment. The fragment analysis method detects efficiently clone numbers and the proportions of each in isolates. PMID- 15662937 TI - [The PAL+ program, an incentive concerted action about malaria and associated infectious diseases, for developing countries]. AB - The French Ministry in charge of Research has launched a multi-institutional incentive concerted action to assist Southern countries on malaria: the PAL+ program. PAL+ aims at bringing out: 1) conditions to promote novel preventive and therapeutic tools adapted to existing situations in the countries concerned; 2) a contribution to help research teams in Southern countries become competitive. PAL+ plans to strengthen cooperative relationships with developing countries (subsaharian Africa, South East Asia and South American countries). Research programs were oriented towards public health needs in malaria-endemic countries and thus mainly focused on: i) development of new antimalarial drugs and new therapeutical strategies: new targets and new leads for drugs, clinical assays for recognition of malaria and optimization of effective treatment or prophylactic drug dosage; ii) pathophysiology of severe malaria: mechanisms of immunity, biology and genome of host and parasite and research leading to vaccine trials; iii) basic and field research on mosquito genetics and biology which may lead to new prevention and control opportunities; iv) social studies on behaviours and habits around prevention and medication of malaria. The objective is to help Southern countries increase their capacity in clinical research, epidemiology, therapeutics, public health and social science (e.g. behaviours and habits accompanying medicine-taking). This means a true partnership and training adapted to specific needs and based on sound science. Research was therefore largely pursued in the laboratories of Southern countries and PAL+ supported the initiative in different ways by: i) providing easier opportunities for scientists from the North to collaborate with scientists from the South; ii) supporting networks of scientist collaborations. This was achieved by setting up a new type of relationships between scientists, based on a continuous dialogue and on bringing them together in small meetings on thematic discussions, the so-called Ateliers de PAL+. The Ateliers should play a major role in increasing the scientific capacity in developing countries. PAL+ program is a commitment to speed up better understanding of the disease by helping endemic countries contribute to research for their own benefit. PMID- 15662938 TI - [Neural control of erection]. AB - Activation of sacral parasympathetic pathways elicits penile erection through the release of vasorelaxant neurotransmitters that increase blood flow to the penis and relax the penile erectile tissue. Sympathetic pathways are antierectile. The pudendal pathway, responsible for the contraction of the perineal striated muscles, enhances an already present erection. All pathways originate in the spinal cord, but at various levels and areas. The convergence of information from peripheral and supra-spinal origins onto spinal neurones is very likely activating more specifically the spinal pro-erectile network. Peripheral information is the afferent limb of reflexive erections, impinges onto spinal interneurones and is able to activate or regulate the activity of sympathetic, parasympathetic and somatic nuclei. Supra-spinal information impinges onto either the same or a different spinal network. Premotor neurones located in supra-spinal structures, that project directly onto spinal sympathetic, parasympathetic or pudendal motoneurones, are present in the medulla, pons and diencephalon. Several of these premotor neurones may in turn be activated by sensory information from the genitals. Descending pathways release a variety of aminergic and peptidergic neurotransmitters in the vicinity of spinal neurones, thereby exerting complex effects on the spinal pro-erectile network. Brainstem and hypothalamic nuclei (among the latter, the paraventricular nucleus and the medial preoptic area) may not reach directly the spinal pro-erectile network. They are prone to regulate penile erection in more integrated and coordinated responses of the body, as those occurring during sexual behaviour. The pro-erectile central and spinal effects of neuropeptides such as oxytocin, melanocortins and endorphins have only recently been analyzed. Such compounds may represent therapeutic strategies to treat erectile dysfunction through a central site of action. PMID- 15662939 TI - [Physiology and pharmacology of ejaculation]. AB - Ejaculation requires an interplay of peripheral actors comprising, among others, smooth and skeletal fibers, glandular and endothelial cells. These actors are driven by vegetative and somatic innervations, deriving essentially from the spinal cord, in turn controlled by cerebral structures and endocrine factors, mostly steroids; These controls require sensitive afferences and command two steps, emission under autonomic control, and ejaculation per se which further involves somatic motoneurons. This review first describes the peripheral innervation of the part of the genital tract concerned in ejaculation, in which the sympathetic component is predominant and releases noradrenalin and neuropeptides; however parasympathetic and somatic components also play a role. At the spinal level, control circuits are organized into networks influenced by spinal structures, which have been discovered through selective lesions or stimulations, as well as by retrograde trans-synaptic tracing with neurotropic viruses. Among these structures, the median preoptic area and the hypothalamic paraventricular nucleus are major regulation sites. On the other hand, serotoninergic and also dopaminergic and adrenergic systems are implicated as well in the command of ejaculation; the latter constitute priviledged targets for a pharmacological treatment of dysfunctions. PMID- 15662940 TI - [Cardiovascular risk factors, erection disorders and endothelium dysfunction]. AB - Upon sexual stimulation, penile erection, occurring in response to the activation of pro-erectile autonomic pathways, is greatly dependent on adequate inflow of blood to the erectile tissue and requires coordinated arterial endothelium dependent vasodilatation and sinusoidal endothelium-dependent corporal smooth muscle relaxation. Nitric oxide (NO) is the principal peripheral pro-erectile neurotransmitter which is released by both non-adrenergic, non-cholinergic neurons and the sinusoidal endothelium to relax corporal smooth muscle through the cGMP pathway. Any factors modifying the basal corporal tone, the arterial inflow of blood to the corpora, the synthesis/release of neurogenic or endothelial NO are prime suspects for being involved in the pathophysiology of erectile dysfunction (ED). In fact, conditions associated with altered endothelial function, such as ageing, hypertension, hypercholesterolemia and diabetes, may, by changing the balance between contractant and relaxant factors, cause circulatory and structural changes in penile tissues, resulting in arterial insufficiency and defect in smooth muscle relaxation and thus, ED. There is increasing evidence to suggest that ED is predominantly a vascular disease and may even be a marker for occult cardiovascular disease. Recent results illustrating the importance of endothelial dysfunction in the pathophysiology of different forms of experimental ED are discussed. These pathways may represent new potential treatment targets. PMID- 15662941 TI - [Genito-sexual dysfunction in patients with a medullary lesion]. AB - Multiple dramatic consequences follow medullary lesions. Not only are voluntary motor control and sensitivity of the body segment below the lesion lost, but it also becomes impossible to control erection and ejaculation as well as urinary and faecal continency. The first investigations into genito-sexual function in paraplegics have brought about the idea, commonly admitted in the medical world, that this kind of patient is impotent and sterile. Fortunately this idea is disappearing gradually and many data have demonstrated that appropriate treatment is required and some therapies efficient. This is particularly important in the case of the population concerned, namely young men in 70% of the cases, since the usual age bracket at trauma is between 25 and 35 years old. At this time of life, sexual activity is often at its peak, so that the fertility potential becomes erased. PMID- 15662942 TI - [Identifying distinct components in the cerebral treatment of visual sexual information through functional neuroimaging]. AB - For now several years, the growing developement of neuroimaging techniques such as Positron Emission Tomography (PET) or functional Magnetic Resonance Imaging (fMRI) allowed a better understanding of neural processes involved in human emotions and goal-directed behaviors. In particular, several studies are now available on the neural correlates of male sexual arousal. A neurobehavioral model of neural processes involved in sexual arousal has been proposed (Redoute et al., 2000) comprising: i) a cognitive component; ii) an emotional component; iii) a motivational component and iv) an autonomic component. Among other regions, several cerebral areas have been found to be linked to: 1) the cognitive component which comprises: i) the orbitofrontal cortex involved in attentional processes directed toward the target and the superior parietal lobules; ii) the inferior parietal lobules involved in motor imagery processes; 2) the motivational component which involves the caudal part of the anterior cingulate cortex, related to motor preparation processes; 3) the autonomic component: concurrent measures of cerebral activations by functional neuroimaging and of erectile response by penile plethysmography allow the demonstration of the involvement of the hypothalamus, the insula, and the rostral part of the anterior cingulate cortex in this component. PMID- 15662943 TI - [Epigenetic post-transcriptional mechanisms for regulating physiological functions, and their decline during aging]. AB - Sequencing of genomes did not reveal the "secret of life" and left biologists and philosophers unsatisfied. It became clear that post-genetic, epigenetic (taken in a wide sense, post-translational) events play an important role even during development and adult homeostasis. Such mechanisms become predominant during aging, and explain the differential decline of functions (aging in "spare parts"). Some of these post-translational mechanisms, leading to auto-amplifying vicious circles, will be described. PMID- 15662944 TI - [Proteasome and proteolysis]. AB - The maintenance of cellular homeostasis and the ability of cells to respond to their environment depend on the degradation of bulk proteins and orderly degradation of key regulatory proteins and their inhibitors. The 26S proteasome plays an essential role in these degradations. It is involved in the activation and inactivation of many cellular processes such as cell cycle progression, apoptosis and regulation of gene expression. It presents unique structural and functional properties. It degrades proteins by an unusual mechanism. Several series of proteasome inhibitors have been developed, useful to elucidate the biological roles of this multicatalytic enzyme. Velcade (bortezomid) was the first proteasome inhibitor to undergo, in may 2003, clinical trials in cancer patients. PMID- 15662945 TI - [Effect of glucose concentration on vascular function in aging. Action on calcium fluxes and vasomotricity induced by elastin peptides]. AB - Glycemia is a physiological parameter tightly regulated for an optimal energetic supply to the organism, in spite of variable tissular glucose needs. Physiopathological alteration of glycemic regulation leads to dysfunctions of many cell types. For example, diabetes considerably increases morbidity and mortality linked to cardiovascular pathologies and constitute nowadays a serious public health problem. Many in vivo and in vitro studies have investigated the impact of extracellular glucose concentration on smooth muscle and endothelial cells. Glycemia regulates expression and activity of proteins implicated in various processes, such as vasodilation (eNOS), cellular adherence (ICAM-1, VCAM 1), glucose transport (GLUT-1) or free radical generation. Nuclear receptors of the PPAR (peroxisome proliferator-activated receptors) family which are implicated in glucose and lipid metabolism control, seem to have direct vascular actions, in the regulation of cellular functions by extracellular glucose, reinforcing their status of pharmacological targets for preservation and improvement of vascular function. More general processes, such as cellular proliferation and cell death, are also influenced by glucose concentration. Concerning the contractile function, hypoglycemia and hyperglycemia modulate vascular reactivity while acting on the vasoactive substances level and the cellular response to these molecules. In particular they act on variation of ionic channels (K+, Ca2+) activity or by interfering with some signaling pathways (NO). For example, the age-dependant vasodilation and endothelial calcium influx induced by elastin peptide are modulated by extracellular glucose levels. In conclusion, abnormal chronic variations of circulating glucose levels seem to be directly responsible for endothelial and smooth muscle cell dysfunction in the pathogenesis of cardiovascular abnormalities of patients presenting glycemia dysregulations. PMID- 15662946 TI - [Thirty years after fibronectin discovery: role in malignant transformation and in ageing]. AB - 1973 was the year of a major discovery: two proteins were shown to disappear from cell surface when cells were transformed by tumor viruses. Later on a large number of glycoproteins were recognized as identical and coined as fibronectin. In spite of 30 years of studies, interest in fibronectin remains vivid. Thanks to its study, understanding of extracellular matrix organisation, cell-matrix interactions and signalling through integrins, formation of fibronectin based fibrillar matrix made great progress. Furthermore importance of matricryptic sites emerged as well as matricryptins. Such peptides show properties that original proteins do not possess. Some of these peptides are used as therapeutic agents. On the contrary others appear to be involved in vicious circles underlying age-dependent decline of tissue function. This fascinating molecule and the parent molecules will certainly remain of interest for some time. PMID- 15662947 TI - [Epistemological views about vicious epigenetic circles due to aging]. AB - Aging is not a disease. Age changes occur in every adult animal. They take place in all species. Yet, malignant tumours and neurodegenerative diseases grow with age. Nobody dies from aging. Death results from age-associated diseases. Muscular and digestive atrophies rise with age. But should it be said that digestive atrophy leads to malnutrition, or is it the contrary? Where is the chicken and where is the egg? We are enclosed in a vicious circle. A vicious circle is not a simple cycle. It is an irreversible phenomenon that happens when a system of objects is acting on itself. Vicious circles now reach the molecular level. This paper tries to show that the destruction of elastin and of fibronectin in connective tissues produces a self destruction effect. Von Neumann showed that a model of self reproduction with the help of reproductive automata could be imagined. We attempt now to show that nature has imagined a model to produce self destruction, trough degradation products of fibronectin and of elastin. This self destruction process is neither purely stochastic, nor genetically programmed. It is an epigenetical process. It is not present in the beginning, as an a priori instruction, it emerges. We try to give here a unusual specific philosophical definition of the word "emergence". PMID- 15662948 TI - Synthesis and antileishmanial activity of 3-imidazolylalkylindoles. Part I. AB - The present study was designed to investigate conazoles as new antileishmanial agents. Several 3-imidazolylalkyl-indoles were prepared under mild reaction conditions and pharmacomodulation at N1 and C5 of the indole ring and at the level of the alkyl chain (R) was carried out starting from the corresponding 3 formylindoles 7-10. All target imidazolyl compounds 38-52 were evaluated in vitro against Leishmania mexicana promastigotes; ketoconazole, amphotericin B and meglumine antimoniate were used as references. Eight out of fifteen compounds (40,43,44,47,48, 50, 51 and 52) exerted similar activity to ketoconazole, with IC50 values in the range of 2.10-3.30 microg/mL. However the most potent compound, 1-(2-bromobenzyl)-3-(1H-imidazol-1-ylmethyl)-1H-indole (38), exhibited IC50 value (0.011+/-0.003 microg/mL) 270-fold lower than that of ketoconazole. Four compounds (38, 43, 50 and 52) were also tested against intracellular amastigotes of L. mexicana; compound 38 exhibited the highest activity with an IC50 value of 0.018+/-0.004 microg/mL. PMID- 15662949 TI - Synthesis and evaluation of disubstituted N1- and N3-imidazolidin-2-ones acting as potential immunosuppressive agents. AB - New N1-mono and N1, N2-disubstituted imidazolidin-2-one with a significant immunosuppressive activity have been discovered. Among the 17 synthesized and tested compounds, five of them showed maximal inhibition of proliferation of concanavallin A (Con A)- stimulated splenocytes at 90 microM, identical to that obtained with cyclosporin A (CsA) at 5 microM, an optimal concentration. PMID- 15662950 TI - Microwave-assisted synthesis of novel thiazolocarbazoles and evaluation as potential anticancer agents. Part III. AB - Novel 6-substituted thiazolocarbazole derivatives have been synthesized under microwave irradiation via the corresponding imino-1,2,3-dithiazoles. In vitro antitumor potential of these polyheterocyclic compounds was evaluated. Among all the tested thiazolocarbazoles, compound 10 is the most effective in inhibiting cell growth. PMID- 15662951 TI - Synthesis and potential anti-inflammatory activity of some tetrahydrophthalazinones. AB - A solid-phase route for the preparation of 4a,5,8,8a-tetrahydrophthalazinon-1 ones employing the Diels-Alder reaction has been developed. Some of the new compounds have been tested for inhibition of LPS-stimulated TNF-alpha production in human whole blood from patients with chronic obstructive pulmonary disease (COPD). This evaluation revealed two compounds 17 and 18 of interest, incorporating an arylpiperazine moiety, which were found to inhibit LPS-induced TNF-alpha release like the well known anti-inflammatory PDE4 inhibitors, rolipram and roflumilast. PMID- 15662952 TI - Synthesis and antiviral effect against herpes simplex type 1 of 12-substituted benzo[c]phenanthridinium salts. AB - The synthesis of benzo[c]phenanthridine alkaloid derivatives is described. In vitro antiviral activity against herpes simplex type 1 (HSV1) has been investigated. Contrary to the natural product fagaronine, which did not have any activity in the HSV1 antiviral tests, four 12-alkoxy derivatives showed good activity demonstrating the importance of the 12-substitution in the structure activity relationships. PMID- 15662953 TI - Synthesis and preliminary in vitro evaluation of antimycobacterial activity of new pyrrolo[1,2-a] quinoxaline-carboxylic acid hydrazide derivatives. AB - New pyrrolo[1,2-a]quinoxaline-2- or -4-carboxylic acid hydrazide derivatives were synthesized from nitroaniline or 1,2-phenylenediamine, and evaluated in vitro for their antimycobacterial activity as part of a TAACF TB screening program. Two compounds 7c and 13 showed an interesting activity at 6.25 microg/mL against Mycobacterium tuberculosis H37Rv, with a 94 and 100 percentage inhibition, respectively. PMID- 15662954 TI - Synthesis and anti-HIV activity of some [Nucleoside Reverse Transcriptase Inhibitor]-C5'-linker-[Integrase Inhibitor] heterodimers as inhibitors of HIV replication. AB - Selected for their expected ability to inhibit HIV replication, a series of eight heterodimers containing a Nucleoside Reverse Transcriptase Inhibitor (NRTI) and an Integrase Inhibitor (INI), bound by a linker, were designed and synthesized. For the NRTIs, d4U, d2U and d4T were chosen. For the INIs, 4-[1-(4-fluorobenzyl) 1H-pyrrol-2-yl]-2,4-dioxobutyric acid (6) and 4-(3,5-dibenzyloxyphenyl)-2,4 dioxobutyric acid (9) (belonging to the beta-diketo acids class) were chosen. The conjugation of the two different inhibitors (NRTI and INI) was performed using an amino acid (glycine or beta-alanine) as a cleavable linker. PMID- 15662955 TI - Synthesis of 5-alkynylated d4T analogues as potential HIV-1 reverse transcriptase inhibitors. AB - A series of 2',3'-didehydro-2',3'-dideoxynucleosides substituted with an alkynylhydroxy- (6, 7, 12 and 13) and alkynylamino- (20) groups at the C-5 position were synthesized. All these five target modified nucleosides were tested for anti-human immunodeficiency virus type 1 activity in CEM-SS and MT-4 cells and unfortunately displayed no improvement in antiviral activity. PMID- 15662956 TI - Naphthalene combretastatin analogues: synthesis, cytotoxicity and antitubulin activity. AB - Synthesis and evaluation of new combretastatin analogues with varied modifications on the bridge and the aromatic rings, have shown that the 2 naphthyl moiety is a good surrogate for the 3-hydroxy-4-methoxyphenyl (B-ring) of combretastatin A-4. Other bicyclic systems, such as 6(7)-quinolyl and 5-indolyl, can replace the B-ring, but they produce less potent analogues in the cytotoxicity and tubulin polymerization inhibition assays. Other modifications are detrimental for the potency of the studied analogues. The 2-naphthyl combretastatin 53 and the related 6-quinolyl combretastatin 106 analogues are the most potent among the derivatives of this type, whereas 92 and 95 are the most potent among the naphthalene derivatives with a heterocycle in the bridge. Previous and new results in this family of combretastatin analogues are discussed. PMID- 15662957 TI - Docking study of ligands into the colchicine binding site of tubulin. AB - Cancer is a major cause of mortality in developed countries, following only cardiovascular diseases. Death of cancerous cells can be achieved by stopping mitosis and the antimitotic class of drugs formed by the spindle poisons can be used for this purpose. Their role is to disorganize the mitotic spindle by targeting its main constituent, the microtubules, themselves made of heterodimers of alpha and beta-tubulin. They disrupt the dynamics of the microtubules either by stabilizing them, as do paclitaxel or epothilones, or destabilizing them, as do colchicine. The binding site of colchicine seems to lie between the two units of the tubulin dimer. Here, we report on the characterization of this site by the docking of a series of reference compounds, and the subsequent docking of ligands prepared in our laboratory. PMID- 15662958 TI - 2- and 3-[(aryl)(azolyl)methyl]indoles as potential non-steroidal aromatase inhibitors. AB - The present study was designed to follow our pharmacomodulation work in the field of non-steroidal aromatase inhibitors. All target compounds 12a-h and 28a-h were tested in vitro for human placental aromatase inhibition, using testosterone or androstenedione as the substrate for the aromatase enzyme and the IC50 and relative potency to aminoglutethimide data are included. A SAR study indicated that 3-[(4-fluorophenyl)(1H-imidazol-1-yl)methyl]-1-ethyl-2-methyl-1H-indole (28 g) was a highly potent and selective aromatase inhibitor with IC50 value of 0.025 microM. 28 g was also a weak inhibitor of androstenedione synthesis. PMID- 15662959 TI - Synthesis and biological evaluation with plant cells of new fosmidomycin analogues containing a benzoxazolone or oxazolopyridinone ring. AB - Fosmidomycin, 3-(N-formyl-N-hydroxyamido) propylphosphonic acid sodium salt, is an efficient inhibitor of 1-deoxy-D-xylulose-5-phosphate (DOXP) reductoisomerase, the second enzyme of the 2C-methyl-D-erythritol-4-phosphate (MEP) pathway notably present in Plasmodium species. We have synthesized a new series of analogues of fosmidomycin, containing a benzoxazolone, benzoxazolethione or oxazolopyridinone ring. As the MEP pathway is involved in the biosynthesis of all isoprenoids, accumulation of ajmalicine in Catharanthus roseus cells was chosen as a marker of monoterpenoid indole alkaloid (MIA) production. None of the twelve studied phosphonic esters 3 and phosphonic acids 4 affected periwinkle cell growth, but some of them (3c, 3e, 3g and 3h) showed a significant inhibition of ajmalicine accumulation: 45-85% at 125 microM. Surprisingly, this effect disappeared by conversion of 3c and 3g into the corresponding acids 4c and 4g, respectively. PMID- 15662960 TI - Synthesis and cytotoxic activity of thiazolofluorenone derivatives. AB - The synthesis and biological evaluation of some novel thiazolofluorenones, thiazolofluorenes and thiazoloanthraquinones, substituted with amino side-chains are described. These polyheterocyclic compounds have been synthesized via the corresponding imino-1,2,3-dithiazoles. Their cytotoxic activity and their eventual selective effect on a phase of the cell cycle were evaluated in vitro, using the murine lymphocytic L1210 leukaemia cell line. PMID- 15662961 TI - Pharmacomodulation of a sulfamide 5-HT6 receptor ligand. AB - A series of N-omega-aminoalkyl- or N-omega-amidinoalkyl-2,4,6-triisopropyl benzenesulfonamides has been synthesized and their respective affinity indices on 5-HT6 receptor determined. This evaluation clearly showed that the compounds possessing an arylpiperazine moiety or an amidine function exhibited good affinity for the model. PMID- 15662962 TI - Synthesis and biological evaluation of thienopyrrolizines, a new family of CDK/GSK-3 inhibitors. AB - Fifteen new thieno[2,3-b ]- and thieno[3,4-b]pyrrolizines were synthesized and tested against two protein kinases, CDK1/cyclin B and GSK-3. Among these compounds, 3-(3-hydroxy-4-methoxyphenyl)-8H-thieno[2,3-b]pyrrolizin-8-one 4g was identified as a moderate inhibitor of these kinases. Its molecular modeling study brought to the fore the pivotal role of the 2-methoxyphenol grouping and the interest in replacing it by bioisosteric moieties in future pharmacomodulations. PMID- 15662963 TI - Nasal eosinophils correlate with FEV1 in patients with perennial allergic rhinitis associated to asthma. AB - BACKGROUND: Eosinophil inflammation is present in nasal and bronchial mucosa and a clear link between upper and lower airways exists. Our aim was to evaluate whether nasal inflammation correlates with lung function in patients with perennial allergic rhinitis associated with asthma. METHODS: Eighty perennial allergic rhinitics with mild asthma, aged from 18 to 46 years, were enrolled. Nasal scraping and spirometry were performed in all subjects. RESULTS: A close relationship between the nasal eosinophil number and the percentage of predicted FEV1 was demonstrated (r = -0.76; p<0.0001). CONCLUSIONS Nasal cytology may be considered a non-invasive tool to assess airway inflammation in mild asthmatics with associated allergic rhinitis. PMID- 15662964 TI - Latex and chickpea (Cicer arietinum) allergy: first description of a new association. AB - In this paper we describe the existence of cross-reactivity between allergens from latex and chickpea, a food from the Leguminosae family, which is common in the Mediterranean diet. We present the case report of a spina bifida boy with a clinical relevant food allergy to chickpea (oral syndrome + dysphonia), developing after the appearance of latex allergy symptoms (lip angioedema + intraoperative anaphylaxis). Specific IgE to latex and chickpea was demonstrated by skin prick tests, measurement of patient's serum specific IgE and IgE immunoblotting. Cross-reactivity was studied by means of EAST-inhibition and western blotting-inhibition. A strong inhibition was observed in several IgE binding bands when latex extract was used in solid phase and patient serum was preincubated with chickpea extract (chickpea extract as inhibitor phase). As far as we know, this is the first report of cross-reactivity between latex and chickpea, a food which should therefore be added to the extensive list of latex cross-reactive foods. PMID- 15662965 TI - Prevalence of allergic reactions to Hymenoptera stings in northern Italy. AB - The epidemiology of Hymenoptera venom allergy in Europe showed prevalence of systemic reactions of about 3%, but is scantly investigated in Italy. We studied two samples of the general and an at risk population, i.e. the foresters, in northern Italy. A standardized questionnaire dealing with allergic reactions to Hymenoptera stings was submitted to the subjects included in the study, who were factory workers in Milan and Verona and foresters in Lombardy. An overall number of 574 subjects participated to the study, 462 (299 males and 163 females, mean age 40.1 yrs, range 20-63 yrs) were factory workers and 112 (all males, mean age 38.8 yrs, range 24-59 yrs) were foresters. Among factory workers, 302 (65.4%) were stung by Hymenoptera, 49 (10.6%) had had large local reactions, and 13 (2.8%) systemic reactions. Among foresters, 76 (67.8%) were stung, 11 (9.8%) had had large local reactions, and 5 (4.5%) systemic reactions. The systemic reactions were life-threatening in 3 (0.6%) industrial workers and 2 (1.8%) foresters. The prevalence of systemic reaction of 2.8% in the general population agrees with the data reported from a number of countries with temperate climate. The prevalence of 4.5% in foresters attributed to this population a risk only slightly increased. PMID- 15662966 TI - Kinetics and dynamic evaluation of specific immunotherapy. AB - Specific immunotherapy (SIT) is frequently used in the treatment of allergic diseases. However, the mechanisms by which SIT achieves clinical improvement remained unclear. We decided to study the in vivo kinetics of this therapy, using a nuclear medicine approach (leukocytes labelled with 99mTc-HMPAO) in patients on maintenance doses of specific immunotherapy with confirmed clinical efficacy. MATERIAL AND METHODS: We studied 13 allergic patients grouped according to different treatment schedules: subcutaneous aqueous allergenic extract (3 latex and 2 hymenoptera venom), subcutaneous depot extract (2 house dust mite and 2 pollens), subcutaneous modified allergens (2 pollens), sublingual extract (2 house dust mites). The control group included two allergic patients submitted to subcutaneous injections of bacterial extract (1 patient--positive control), and aqueous solution (1 patient). At the same time that the therapeutic allergen was administered subcutaneously, the autologous labelled white cells were injected intravenously in a peripheral vein in the contralateral arm. A thoracic dynamic acquisition of 60 mins, 64x64 matrix, 2 frame/min, in anterior view was performed. Static acquisition for 256x256 matrix, during 5 mins each at 60, 90, 120, 180, 240, 300 and 360 mins after the administration of the radiolabelled leukocytes, in thoracic (anterior and posterior), and abdominal view were performed. During the examination, the local erythema was monitored. A similar procedure was undertaken for Sublingual administration of immunotherapy. RESULTS: The inflammatory activity at the site of SIT injection (aqueous depot extract) started in the first hour and the increase was time related. For modified allergen extract and sublingual SIT the activity was present since the beginning of the administration. The ascendant lymphatic drainage, which was directed to the homolateral axillary region, to the lymphoid tissue of the upper mediastinum and to the anterior region of the neck began earlier. Thoracic focalisations were present for all the patients, whereas bowel focalisations were only observed for the subcutaneous route of administration. Sublingual SIT did not induce axillary or intestinal inflammatory focalisations, even though the patients had swallowed the allergenic extract. The uptake coefficient in individualized areas corrected to the uptake coefficient background was also studied. CONCLUSIONS: For the subcutaneous route of administration, except for glutaraldehyde-modified allergen, the local inflammatory activity at the allergenic injection site was significantly higher in depth and was time dependent, maintaining activity even after complete disappearance of the erythema and/or wheal. These results express a prompt inflammatory involvement of the immune system with this allergenic therapy, which was unexpected until now. We also observed differences concerning allergic diseases, the type of allergenic extracts and routes of administration. PMID- 15662967 TI - Zygomycosis in Qatar: a retrospective review of six cases. AB - Six cases of zygomycosis that were diagnosed at Hamad Medical Corporation over an eleven-year period are retrospectively reviewed. All patients had at least one predisposing condition. Renal transplant and diabetes mellitus were the most common predisposing conditions. Sites of involvement were pulmonary in two patients, rhinocerebral two, rhino-orbital, and cutaneous in one each. The clinical features in patients with rhinocerebral and rhino-orbital involvement were very suggestive of the diagnosis, while patients with pulmonary and cutaneous involvement, the disease was not suspected and was only made after the histopathology or culture results were available. Diagnosis was made premortum in all patients. Diagnosis was confirmed by histopathology in three patients and by culture in the other three. Five of the six patients died. Poor outcome correlated with rhinocerebral and pulmonary involvement, while cutaneous disease was associated with favorable outcome. Therapy with amphotericin B, surgical debridement, and correction of the underlying predisposing condition offers the best chance for survival. PMID- 15662968 TI - Response of low-grade oligodendroglial tumors to temozolomide. PMID- 15662969 TI - Transduction of human glial and neuronal tumor cells with different lentivirus vector pseudotypes. AB - Lentiviral vectors have proven to be valuable tools for in vitro and in vivo gene delivery because they can transduce dividing and non-dividing cells efficiently, and mediate long-term gene expression. Pseudotyping of lentiviral vectors with envelope proteins other than VSV-G has resulted in enhanced transduction of certain cell types and tissues. In order to improve lentiviral vector-based gene therapy for peripheral neuroectodermal and brain tumors, we compared the efficiency of eight different lentivirus pseudotypes in transducing neuronal and glial tumor cell lines. Here, lentiviral vectors pseudotyped with the envelopes from human foamy virus, rabies, Mokola or amphotropic murine leukemia virus displayed the highest transduction efficiency in neuroblastomas, whereas pseudotyping with the lymphocytic choriomeningitis virus glycoprotein from strain Armstrong 53b resulted in the highest transduction efficiency in gliomas. PMID- 15662970 TI - Enhanced cytotoxic effects of 5-aminolevulinic acid-mediated photodynamic therapy by concurrent hyperthermia in glioma spheroids. AB - During photodynamic therapy (PDT) both normal and pathological brain tissue, in close proximity to the light source, can experience significant temperature increases. The purpose of this study was to investigate the anti-tumor effects of concurrent 5-aminolevulinic acid (ALA)-mediated PDT and hyperthermia (HT) in human and rat glioma spheroids. Human or rat glioma spheroids were subjected to PDT, HT, or a combination of the two treatments. Therapies were given concurrently to simulate the conditions that will occur during patient PDT. Predictions of diffusion theory suggest that brain tissue immediately adjacent to a spherical light applicator may experience temperature increases approaching 8 degrees C for laser input powers of 2 W. In the in vitro model employed here, HT had no effect on spheroid survival at temperatures below 49 degrees C, while sub threshold fluence PDT results in only modest decrease in survival. HT (40-46 degrees C) and PDT interact in a synergistic manner if the two treatments are given concurrently. The degree of synergism increases with increasing temperature and light fluence. Apoptosis is the primary mode of cell death following both low fluence rate PDT and combined HT + PDT. PMID- 15662971 TI - Up-regulation of CC chemokine, CCL3L1, and receptors, CCR3, CCR5 in human glioblastoma that promotes cell growth. AB - Human CC ligand 3-like protein 1 (CCL3L1), a member of the CC chemokine family, that induces MCP1 and RANTES, exhibits a variety of proinflammatory activities including chemotaxis, and functional and proliferative activation of leukocytes, lymphocytes and macrophages. Its signal is transmitted through transmembrane receptors, CC chemokine receptors, CCR1, CCR3 and CCR5. To examine gene expression of chemokine, CCL3L1, and its receptors, CCR1, CCR3 and CCR5, we analyzed tumor tissues from 21 patients with several types of primary gliomas. CCL3L1, CCR3 and CCR5 gene exhibited over-expression in 70% (7/10), 60% (6/10), and 60% (6/10) of glioblastoma, in comparison with lower frequencies seen in lower-grade gliomas. Transfection of CCL3L1-expression vector to glioblastoma cell line enhanced proliferation of the tumor cells. These data suggest that increased expression of the CCL3L1, CCR3 and CCR5 chemokine-receptors system is involved in brain tumorigenesis, especially in the progression of glioblastoma. PMID- 15662972 TI - Quantification of thrombospondin-1 secretion and expression of alphavbeta3 and alpha3beta1 integrins and syndecan-1 as cell-surface receptors for thrombospondin 1 in malignant glioma cells. AB - Malignant glioma cells secrete thrombospondin-1 (TSP-1) which participates in the motility of glioma cells, and binds to cell surface alphavbeta3 and alpha3beta1 integrins, and syndecan-1. This study evaluated the amount of TSP-1 secretion from malignant glioma cells, and the expression of alphavbeta3 and alpha3beta1 integrins, and syndecan-1. The amounts of TSP-1 in the supernatants from 10 malignant glioma cell lines and eight non-glioma malignant tumor cell lines were measured by enzyme-linked immunosorbent assay. Expression of alphavbeta3 and alpha3beta1 integrins, and syndecan-1 were examined by flow cytometry. The amounts of TSP-1 secreted by malignant glioma cells were 43 to 2431 ng/l x 10(6) cells/24 h (mean +/- SD = 626 +/- 792). Seven of 10 glioma cell lines secreted more than 100 ng of TSP-1 and three of these cell lines secreted more than 1 microg. Seven of eight non-glioma cell lines secreted less than 100 ng of TSP-1. All glioma cell lines expressed alpha3beta1 integrin and syndecan-1, and seven of 10 glioma cell lines expressed alphavbeta3 integrin. Treatment of the glioma cell lines with TGF-beta2 did not change the expression of alphavbeta3 integrin. These results suggest that malignant glioma cells secrete high levels of TSP-1, which may be important in the migration of glioma cells via interactions with alphavbeta3 and alpha3beta1 integrins, and syndecan-1. PMID- 15662973 TI - Hyperthermia induces translocation of apoptosis-inducing factor (AIF) and apoptosis in human glioma cell lines. AB - In the hyperthermal treatment, the wild type (wt) p53 plays an important role in apoptosis induction in the tumor cells. In human gliomas, p53 frequently has some form of mutation. The mutant type (mt) p53 does not work properly as a tumor suppressor and this may result in poor responses during treatment. We investigated the relationship between apoptosis-inducing factor (AIF) and apoptosis under various thermal conditions (43, 45, and 47 degrees C for 1 h) using four p53-wild or -mutant human glioma cell lines (A172, T98G, U251MG, and YKG-1). AIF translocation from the mitochondria to the nucleus under hyperthermal conditions was demonstrated by confocal laser microscopy. The percentage of AIF positive nuclei increased significantly in comparison with the control in all cell lines and in all temperature groups except for YKG-1 at 47 degrees C. Immunoblot analyses of the nuclear fraction of each cell line revealed temperature-dependent increases in AIF. A simultaneous release of cytochrome c from the mitochondria to the cytosol was noted. A flow cytometric analysis showed that apoptosis induction occurred more often in a temperature-dependent manner in the 45 and 47 degrees C groups than in the control group. These findings indicate that the hyperthermal conditions can lead to AIF translocation and apoptotic cell death in the p53-mutant human glioma cells. The present report is the first description of AIF-induced apoptosis in hyperthermia. PMID- 15662975 TI - An unusual presentation of glioblastoma multiforme. PMID- 15662974 TI - Immunohistochemical study for O6-methylguanine-DNA methyltransferase in the non neoplastic and neoplastic components of gliomas. AB - Although the expression O6-methylguanine-DNA methyltransferase (MGMT) is an important hallmark for decision of nitrosourea chemotherapy for glioma patients, no immunohistochemical method for analysis of MGMT has been standardized yet. Gliomas usually contain non-neoplastic cells even deep in the tumor. It is not known which of these components expresses MGMT. To clarify this point, we investigated MGMT expression in the non-neoplastic cells in autopsy and surgical specimens by immunohistochemistry. High grade gliomas were also studied to find a cut-off point for treatment decision. MGMT immunohistochemistry in the normal brain or brain with non-neoplastic disease revealed nuclear staining in some endothelial cells, inflammatory cells, ependymal cells, astrocytes and oligodendroglias. Some cells were double stained with CD68 (macrophages or microglias). The neurons were consistently MGMT-negative. High grade gliomas always contained an MGMT-positive non-neoplastic component. Although, the endothelial cells were easily distinguished from the neoplastic cells, other cells were often mistaken for tumor cells. The population of MGMT-positive non neoplastic cells was usually less than 10%. We set a cut off-point at 10% between the positive and negative groups because the statistical difference in the overall survival was most distinct at this value. In 51 high grade glioma patients, who received both radiotherapy and chemotherapy with nimustine (ACNU), the median overall survival of the MGMT-negative group (23 months) was significantly longer than that of the MGMT-positive group (14 months) (P < 0.009). Multivariate analysis revealed that the negative MGMT expression was a significant prognostic variable next to the degree of surgical removal for the overall survival. In the MGMT-positive group, addition of platinum-based chemotherapy did not improve the survival. PMID- 15662976 TI - Use of fractional anisotropy value by diffusion tensor MRI for preoperative diagnosis of astrocytic tumors: case report. AB - The fractional anisotropy (FA) value calculated by diffusion tensor MRI can indicate the degree of directionality of water diffusion in astrocytic tumors. Here, we report a case of anaplastic astrocytoma in which FA proved invaluable for the preoperative differential diagnosis. A 60-year-old man complained of headache, and underwent routine neuroimaging and DTI. The routine images suggested a low-grade glioma in the left temporal lobe, based on lack of enhancement on MRI with contrast medium and lack of tumor staining on angiograms, whereas FA value was very high. Based on these findings, a preoperative diagnosis of high-grade glioma was suspected. The surgical specimen exhibited the histological features of anaplastic astrocytoma with a high density of spindle shaped cells and low vascularity. In this report, we discuss the relationship between FA and other characteristics of the present tumor, and discuss the utility of FA measurement in astrocytic tumors. PMID- 15662977 TI - The relationship between peritumoral brain edema and the expression of vascular endothelial growth factor and its receptors in intracranial meningiomas. AB - We examined the radiological and histological features of, and the influences of the expression of VEGF and its two major receptors, Flt-1 and Flk-1, on the development of peritumoral brain edema (PTBE) in patients with intracranial meningiomas. The expressions of VEGF and VEGF receptors in the immunohistochemical study were analyzed in relation to several factors, including tumor size, location, vascularity, and blood supply, as seen on digital subtraction angiographic studies. The edema volume (P = 0.0003) and edema index (P < 0.0001) had a significantly positive relation to VEGF expression. The positivity of Flt-1 and Flk-1 was mainly observed in tumor vessels; 44 cases (37.2%) were positive for the Flt-1 antibody and 37 cases (31.4%) for the Flk-1 antibody. The mean value of the edema index of the positive-Flt-1 group (5.220 +/ 11.586) was significantly higher than that of the negative-Flt-1 group (1.782 +/ 2.559) (P < 0.0001). The mean value of the edema index of the positive-Flk-1 group (3.925 +/- 5.870) was slightly higher than that of the negative-Flk-1 group (2.671 +/- 8.136) (P < 0.0001). Our data suggest that the expressions of VEGF and VEGF receptors positively relate to each other and to the formation of PTBE in patients with meningiomas. PMID- 15662979 TI - Clinical guidelines--are they of any use? PMID- 15662978 TI - Phase II study of temozolomide and cisplatin as primary treatment prior to radiotherapy in newly diagnosed glioblastoma multiforme patients with measurable disease. A study of the Spanish Medical Neuro-Oncology Group (GENOM). AB - This phase II study evaluates the activity of temozolomide and cisplatin administered before radiation therapy in newly diagnosed glioblastoma multiforme patients, in terms of response, time to progression and survival. PATIENTS AND METHODS: Forty patients with measurable disease after surgery, a Karnofsky status > 60, and Barthel Index > 10 were included. They were treated with three cycles of temozolomide 200 mg/m2/day for 5 days and cisplatin 100 mg/m2 on day 1. Conventional focal radiation therapy to 60 Gy was administered after response evaluation. RESULTS: Three patients were not evaluable for central reviewed response but all 40 were evaluable for toxicity, time to progression and survival. Objective responses by Macdonald criteria on an intent to treat basis were 45% including complete response in three patients (7.5%), and partial response in 15 patients (37.5%). Responses were seen in biopsy-only patients (33.4%) as well as in partial surgery patients (52%). Median survival for all patients was 12.5 months. Biopsy-only patients had a median survival of 12.8 months. Grade 3 to 4 neutropenia was the most important toxicity, and occurred in 37.5% of patients. A delay in 18.2% and a dose reduction in 9.6% of cycles were necessary due to myelosuppression on day 28. Two patients had neutropenic fever resulting in one treatment-related death. Eighty-two percent of patients received radiotherapy. CONCLUSION: This regimen has significant activity, as it induces objective responses even in biopsy-only patients, appearing to improve their median survival. A better combination schedule is needed to improve the toxicity profile. PMID- 15662980 TI - PMTCT programme--partial assessments can build the picture. PMID- 15662983 TI - Impact of chronic diseases on the health-related quality of life of South Africans. PMID- 15662981 TI - Sedation of children undergoing MRI--a risky business! PMID- 15662984 TI - Tactical shift to improve SAMA's prospects? PMID- 15662985 TI - Miracles in the land of non-accountability. PMID- 15662986 TI - Bisho's uncoded maladies. PMID- 15662987 TI - Door-handle symptoms. PMID- 15662988 TI - Naropin: safety and efficacy not established in children under 1 year of age. PMID- 15662989 TI - Life and medicine in the Islamic Republic of Pakistan. PMID- 15662990 TI - Imaging of carotidynia. PMID- 15662991 TI - Drug policy for methamphetamine use urgently needed. PMID- 15662992 TI - From research presentation to publication. PMID- 15662993 TI - Instant vertebral assessment/lateral vertebral assessment: an integral and essential part of osteoporosis assessment. PMID- 15662994 TI - Gastric luminal epidermal growth factor is affected by diet. AB - OBJECTIVE: Diet is an area of major interest to those investigating the causes of cancer of the oesophagus in the Transkei. This study looked at the associations between intragastric epidermal growth factor level, diet and intragastric pH. SETTING AND SUBJECTS: A dietary survey was co-ordinated with studies of gastric luminal epidermal growth factor and gastric fluid pH in 120 rural Transkeians. RESULTS: Gastric fluid epidermal growth factor was associated with low dietary intake of animal products (p = 0.002) and vegetables (p = 0.026). There was no association with pH. CONCLUSION: A dietary subgroup has been identified in the Transkei population with high levels of epidermal growth factor in the upper gastrointestinal lumen. This adds to previously demonstrated diet-related changes in the upper gastrointestinal tract in Transkei. These changes may affect the disease pattern of the population. PMID- 15662995 TI - Efficacy of albendazole against the whipworm trichuris trichiura--a randomised, controlled trial. AB - OBJECTIVES AND DESIGN: To test the efficacy of albendazole against the whipworm Trichuris trichiura for school-based deworming in the south-western Cape, South Africa. Children infected with Trichuris were randomised to 3 doses of albendazole (400, 800 or 1200 mg), each repeated 4 times. The boy/girl ratio was 1. A group not infected with worms was treated with placebo, creating a negative control. SUBJECTS AND SETTING: Pupils at a primary school serving a wine producing area approximately 90 km east of Cape Town. OUTCOME MEASURES: Trichuris cure rates and reduction in the number of eggs/g in faeces, as well as the infection dynamics of Trichuris and Ascaris during treatment with placebo. RESULTS: Albendazole treatment was associated with Trichuris cure rates of 23% (400 mg), 56% (800 mg) and 67% (1200 mg) after the final treatment. The corresponding reductions in the number of eggs/g of faeces were 96.8%, 99.3% and 99.7%. Environmental pollution by human faeces was confirmed because worm egg negative children in the placebo group became egg-positive while the study was in progress. CONCLUSION: The 400 mg stat dose had a low Trichuris cure rate. To repeat the dose on 2 or 3 days would increase cost, reduce compliance and complicate management. Albendazole cannot be used in deworming programmes in South Africa because it is a Schedule 4 prescription medicine. De-scheduling is needed urgently, particularly because of high efficacy against hookworm in KwaZulu-Natal and neighbouring countries. PMID- 15662996 TI - The impact of endovascular treatment on cerebral aneurysm outcome at Groote Schuur Hospital. AB - OBJECTIVES: Neuro-endovascular treatment was introduced at Groote Schuur Hospital in 2001. We sought to assess whether this has resulted in a change in overall outcomes for patients treated for cerebral aneurysms. DESIGN: Retrospective cohort study. The first cohort included cerebral aneurysm patients seen between 1994 and 1998 when only surgical treatment was available. The second cohort consisted of a group treated using both surgical and endovascular methods between 2002 and 2004. SUBJECTS: Patients with ruptured and unruptured aneurysms were included in the study. OUTCOME MEASURES: The Glasgow Outcome Score (GOS) wasused to assess outcome. The primary comparator between groups was major disability (GOS 3 and 2) and death (GOS 1). RESULTS: Cohorts were comparable regarding age, sex and presenting clinical condition as assessed using the World Federation of Neurological Surgeons grading for subarachnoid haemorrhage. There was an absolute reduction in major disability and death of 16% in the later cohort where 55% of patients had endovascular treatment. CONCLUSIONS: The option of endovascular treatment for cerebral aneurysms at Groote Schuur Hospital has allowed for more rapid treatment of patients, which has reduced morbidity and mortality from re bleeding. We are also able to select the best treatment option for each patient and believe this has contributed to our improved results. PMID- 15662997 TI - Endovascular treatment of cerebral aneurysms--a cost analysis. AB - OBJECTIVES: To determine if endovascular treatment of cerebral aneurysms is cost effective when compared with conventional surgical treatment. DESIGN: A retrospective study. SUBJECTS: Seventeen patients treated between August 2002 and August 2003 had posterior communicating artery aneurysms. This group was selected for detailed analysis. OUTCOME MEASURES: Total treatment costs, in South African rands (R), were calculated for each patient. Average treatment costs for each group were then compared. RESULTS: Surgically clipped and endovascularly coiled groups were comparable for age, sex and clinical condition. The average cost for endovascular treatment per patient was R37 041. Surgical treatment was more expensive at R44 104, a difference of 16%. CONCLUSIONS: Despite the high cost of endovascular devices, appropriate use of this technology ultimately offers less expensive treatment than microsurgical clipping of aneurysms. PMID- 15662998 TI - Effect of Nigerian citrus (Citrus sinensis Osbeck) honey on ethanol metabolism. AB - The effect of Nigerian citrus (Citrus sinensis Osbeck) honey on ethanol metabolism was tested using 45 consenting individuals in apparent good health and between the ages of 25 and 35 years. The subjects were moderate social drinkers matched in terms of body weight and build. The results obtained showed that on average, honey significantly (p < 0.05) increased the blood ethanol clearance rate by 68% and decreased the intoxication period by 43%, but insignificantly (p > 0.05) reduced the degree of intoxication by 9%. Honey could be a promising anti intoxicating agent, but its long-term biochemical evaluation, possibly as a complement in the management of alcohol intoxication, deserves further study. PMID- 15662999 TI - Homology modeling, force field design, and free energy simulation studies to optimize the activities of histone deacetylase inhibitors. AB - As an effort to develop therapeutics for cancer treatments, a number of effective histone deacetylase inhibitors with structural diversity have been discovered. To gain insight into optimizing the activity of an identified lead compound, a computational protocol sequentially involving homology modeling, docking experiments, molecular dynamics simulation, and free energy perturbation calculations was applied for rationalizing the relative activities of known histone deacetylase inhibitors. With the newly developed force field parameters for the coordination environment of the catalytic zinc ion in hand, the computational strategy proved to be successful in predicting the rank orders for 12 derivatives of three hydroxamate-based inhibitor scaffolds with indole amide, pyrrole, and sulfonamide moieties. The results showed that the free energy of an inhibitor in aqueous solution should be an important factor in determining the binding free energy. Hence, in order to enhance the inhibitory activity by adding or substituting a chemical group, the increased stabilization in solution due to the structural changes must be overcome by a stronger enzyme-inhibitor interaction. It was also found that to optimize inhibitor potency, the hydrophobic head of an inhibitor should be elongated or enlarged so that it can interact with Pro29 and His28 that are components of the flexible loop at the top of the active site. PMID- 15663000 TI - QSAR and classification models of a novel series of COX-2 selective inhibitors: 1,5-diarylimidazoles based on support vector machines. AB - The support vector machine, which is a novel algorithm from the machine learning community, was used to develop quantitation and classification models which can be used as a potential screening mechanism for a novel series of COX-2 selective inhibitors. Each compound was represented by calculated structural descriptors that encode constitutional, topological, geometrical, electrostatic, and quantum chemical features. The heuristic method was then used to search the descriptor space and select the descriptors responsible for activity. Quantitative modelling results in a nonlinear, seven-descriptor model based on SVMs with root mean square errors of 0.107 and 0.136 for training and prediction sets, respectively. The best classification results are found using SVMs: the accuracy for training and test sets is 91.2% and 88.2%, respectively. This paper proposes a new and effective method for drug design and screening. PMID- 15663001 TI - BODIL: a molecular modeling environment for structure-function analysis and drug design. AB - BODIL is a molecular modeling environment geared to help the user to quickly identify key features of proteins critical to molecular recognition, especially (1) in drug discovery applications, and (2) to understand the structural basis for function. The program incorporates state-of-the-art graphics, sequence and structural alignment methods, among other capabilities needed in modern structure function-drug target research. BODIL has a flexible design that allows on-the-fly incorporation of new modules, has intelligent memory management, and fast multi view graphics. A beta version of BODIL and an accompanying tutorial are available at http://www.abo.fi/fak/mnf/bkf/research/johnson/bodil.html. PMID- 15663002 TI - Tuning of hydrogen bond strength using substituents on phenol and aniline: a possible ligand design strategy. AB - Using Density Functional Theory, the hydrogen bonding energy is calculated for the interaction of phenol and aniline with four model compounds representing the protein backbone and various amino acid side chain residues. The models are methanol, protonated methylamine, formaldehyde and acetate anion. The H-bond energies for the uncharged species are approximately 2.5 kcal mol(-1), whereas the charged model compounds bind with much higher energies of approximately 20 kcal mol(-1). The effect of para-substitution on the hydrogen bond energies is determined. Substitution has little effect on the H-bond energy of the neutral complexes (<2 kcal mol(-1)), but for the positively and negatively charged systems substitution drastically alters the binding energies, e.g., 14.3 kcal mol(-1) for para-NO2. In the context of protein-ligand binding, relatively small changes in binding energy can cause large changes in affinity due to their exponential relationship. This means that for -NO2 an enormous change of 10 orders of magnitude for the affinity constant is predicted. These calculations allow prediction of H-bonds, using different substituents, in order to fine-tune and optimize ligand-protein interactions in the search for drug candidates. PMID- 15663003 TI - Interactive essential dynamics. AB - Essential dynamics (ED) is a useful method for analyzing trajectories generated by molecular dynamics (MD), but current tools are awkward to use, limiting the usefulness of the technique. This paper describes a new interactive graphical interface for visualization of ED results, including filtering a trajectory on an arbitrary set of eigenvectors and manipulation of a structure's projection along any eigenvector. PMID- 15663004 TI - The interaction of PTP-BL PDZ domains with RIL: an enigmatic role for the RIL LIM domain. AB - PDZ domains are protein-protein interaction modules that are crucial for the assembly of structural and signaling complexes. PDZ domains specifically bind short carboxyl-terminal peptides and occasionally internal sequences that structurally resemble peptide termini. Previously, using yeast two-hybrid methodology, we studied the interaction of two PDZ domains present in the large submembranous protein tyrosine phosphatase PTP-BL with' the C-terminal half of the LIM domain-containing protein RIL. Deletion of the extreme RIL C-terminus did not eliminate binding, suggesting the presence of a PDZ binding site within the RIL LIM moiety. We have now performed experiments in mammalian cell lysates and found that the RIL C-terminus proper, but not the RIL LIM domain, can interact with PTP-BL, albeit very weakly. However, this interaction with PTP-BL PDZ domains is greatly enhanced when the combined RIL LIM domain and C-terminus is used, pointing to synergistic effects. NMR titration experiments and site directed mutagenesis indicate that this result is not dependent on specific interactions that require surface exposed residues on the RIL LIM domain, suggesting a stabilizing role in the association with PTP-BL. PMID- 15663005 TI - Abscisic acid promoted changes in the protein profiles of rice seedling by proteome analysis. AB - This study investigates the influence of exogenously applied abscisic acid (ABA) on the leaves and leaf sheaths of two-week-old rice seedling at the level of the proteome. Significant differences were observed in the two-dimensional polyacrylamide gel electrophoresis protein profiles between control and ABA treated samples. Amino-acid sequence analysis of affected proteins revealed that ABA caused drastic changes in the major photosynthetic protein, ribulose 1,5 bisphosphate carboxylase/oxygenase and accumulation of certain defense/stress related proteins. Moreover, cutting or treating leaf sheaths with jasmonic acid (JA) rapidly increased the endogenous level of ABA, suggesting a role for ABA during the defense/stress-response. Comparative study indicated a potential overlap between ABA and JA as detected at the level of the proteome. Furthermore, in vitro protein phosphorylation experiments and in-gel kinase assays also revealed considerable changes in the phosphorylation status of some proteins, and differential effects on myelin basic protein and calcium-dependent protein kinase activities by ABA treatment, which suggests involvement of kinase in the downstream signaling cascade. These results provide evidence at proteome level for the involvement of ABA in stress-response in rice seedling. PMID- 15663007 TI - Protein and mRNA characterization in high and low metastasis adenoid cystic carcinoma cell lines. AB - Metastasis and invasion, the important characteristics of malignant tumors, are closely associated with a series of changes in the expression of genes and proteins. In this study, we compare mRNA and protein expression in high and low metastasis adenoid cystic carcinoma cell lines by mRNA suppression subtractive hybridization and two-dimensional electrophoresis combined with peptide mass fingerprint analysis. 34 differentially expressed genes were obtained using suppression subtractive hybridization experiments including 6 highly expressed gene sequences in the high metastasis cell line, and 28 in the low metastasis cell line. RNA dot blot hybridization further confirmed the results after excluding false positives. For protein analysis, ten significantly different protein spots were detected using two-dimensional gel electrophoresis technique combined with matrix-assisted laser desorption/ionization time-of-flight mass spectrometer (MALDI- TOF-MS). The results then compare with the SWISS PROT database. These results suggest that high tumor metastasis of adenoid cystic carcinoma is associated with multiple genes whose function include angiogenesis, protein synthesis, signal transduction, modulation of cell cycle, molecular chaperones, and immune co-stimulating molecule. Moreover, the results of the phenotypic function-related expression mapping analysis at the mRNA and protein level revealed obvious complementarities, providing important clues for further study of the molecular mechanism of metastasis, metastasis control and possible targets for cancer gene therapy. PMID- 15663006 TI - Nucleotide sequence and molecular characterization of the structural glycoprotein gp41 gene homologue of Spodoptera litura nucleopolyhedrosis virus (spltNPV-I). AB - The structural glycoprotein gene gp41 homologue of Spodoptera litura nucleopolyhedrosis virus (SpltNPV-I *) was identified in the 4.0 kb EcoRI-L fragment of the viral genome. The nucleotide sequence of 2063 bp of this fragment revealed an open reading frame of 1014 nucleotides to encode a polypeptide of 337 amino acids. Analysis of nucleotide and deduced amino acid sequences of the putative ORF indicated its identity with gp41 protein of other baculoviruses sharing maximum homology with that of Spodoptera frugiperda nucleopolyhedrosis virus (SfNPV). The coding sequence was preceded by an AT-rich region containing the consensus baculoviral late promoter motif RTAAG. The putative SpltNPV gp41 ORF was abundantly expressed as a 37 kDa apoprotein in E. coli and as a 50 kDa glycoprotein in Sf9 cells. The recombinant protein expressed in insect cells was glycosylated (20%) and has GlcNAc as the terminal sugar. The gene is conserved among baculoviruses and places SpltNPV-I close to Spodoptera frugiperda and Spodoptera exigua NPVs in phylogenetic tree. PMID- 15663008 TI - Molecular cloning and expression analysis of rice phosphoribulokinase gene that is regulated by environmental stresses. AB - Phosphoribulokinase (PRKase, EC 2.7.1.19) plays an important role in regulating the flow of sugar through the Calvin cycle. To investigate its regulatory character and expression pattern, the gene encoding PRKase in rice was cloned by RACE. A full-length cDNA with an open reading frame of 1212 bp encoding 403 amino acids residues was obtained from an indica rice variety, 9311. The OsPrk is a single locus gene in the rice genome. It is localized in the region of 28.32 approximately 28.33 Mb on the Chromosome 2, flanked by the genetic markers RM450 and MRG0168. RT-PCR analysis revealed that OsPrk gene was expressed in all the tissues we tested and OsPrk transcript level could be dramatically boosted by light illumination. Its expression was down-regulated by externally applied NaCl, ABA, MeJA and glucose over 24 hr, whereas it was up-regulated by GA after 24 hr treatments. These results also indicated that OsPrk gene expression is modulated by these factors at multiple levels. PMID- 15663009 TI - Apolipoprotein E polymorphism in Saudis. AB - Apolipoprotein E (apoE) genotypes were determined in 165 Saudis. The prevalence of genotype, E3/E3, E3/E4 and E4/E4 was found to be 71, 27 and 2% respectively. The E3/E3 was the most prevalent genotype among the Saudis followed by E3/E4. However, other genotypes E2/E2, E2/E3 and E2/E4 were absent showing the absence of E2 allele in the test population. The high frequencies of the E3 allele (0.845) and E3/E3 genotype (0.71) and absence of E2 allele in Saudis under study are similar to those reported earlier for Native Americans, Mexican-Americans, Mayans, Cayapa, Mazatecan Indians and Mexican Mestizos populations. PMID- 15663011 TI - Intestinal transplantation. PMID- 15663010 TI - Identification and expression of a conserved ubiquitin gene homologue of Spodoptera litura nucleopolyhedrovirus (spltNPV-I). AB - An ORF having a potential to code for a polypeptide of 79 amino acids has been identified within 993 nt sequence of 2 kb EcoRI-W fragment of Spodoptera litura nucleopolyhedrovirus (SpltNPV-I). Nucleotide and deduced amino acid sequence analyses showed its identity with the ubiquitin homologue of eukaryotes (79-80%), Melanoplus sanguinipes entomopoxvirus (76%) and other baculoviruses (72-89%). The ORF is under baculovirus late promoter motif RTAAG but unlike other baculoviruses, three such motifs at -6, -10 and -27 position are present in SpltNPV. The ORF expresses as a 10 kDa proteinin E. coli and the purified recombinant protein showed crossreactivity with the rabbit anti-ubiquitin antibodies. PMID- 15663012 TI - An analysis of the long-term complications of intestine transplant recipients. AB - BACKGROUND: One-year survival rates for intestine transplant recipients have increased to 70%, but it is believed that the complications following the procedure do not diminish after the first year as is seen in other forms of solid organ transplantation. OBJECTIVE: To review the ongoing medical requirements of an increasing number of long-term survivors of intestinal transplantation. METHOD: A retrospective medical chart review was completed on all patients who received intestinal transplantation at the University of Nebraska Medical Center from September 1990 through March 2003. One hundred forty-six transplantations were performed on 128 patients-53 intestinal, 70 liver/intestinal, and 23 intestinal with liver and pancreas or kidney. RESULTS: Seventy-six patients survived longer than 365 days and form the study group. Major reasons for readmissions were infections, gastrointestinal complications, dehydration, and rejection. The average number of rehospitalizations per patient per year remained constant. Death in patients with more than 365 days survival (N=23) was the result of sepsis (56%) and multiple other complications (44%). CONCLUSION: The number of readmissions each year per patient remains constant and medical problems remain complex and life threatening. Patients with intestinal transplantation will continue to require regular expert follow-up care and careful attention to even the smallest medical problem even years following transplantation. PMID- 15663013 TI - Quality of life after intestinal transplantation. AB - As technology introduces more advanced therapies, individuals have realized that quantity or length of life may not always be the ultimate goal in treating disease. The quality of life at times is as important as length of life. In this article, we review the literature regarding quality of life after intestinal transplantation. Published data on patient survival are briefly reviewed and the available studies investigating quality of life are examined. Few studies have been performed to date, but the limitations and implications of the available studies are summarized. PMID- 15663014 TI - Intestinal rehabilitation: a management program for short-bowel syndrome. AB - Over the last 3 decades, there has been significant improvement in the survival and quality of life of patients who require home parenteral nutrition; however, parenteral nutrition remains costly, is associated with multiple complications, and does not promote the function of the remaining bowel. Intestinal rehabilitation refers to the process of restoring enteral autonomy and decreasing dependence on parenteral nutrition by utilizing dietary, pharmacological, and, occasionally, surgical interventions. A major focus of research has been to identify a trophic factor that will enhance adaptation of the remaining gastrointestinal tract following massive gut resection and allow enteral autonomy. Whether intestinal rehabilitation occurs as the result of increased intestinal adaptation or as the result of a comprehensive approach to care has yet to be determined. This article reviews intestinal failure as the result of short-bowel syndrome and the management strategy of an intestinal rehabilitation program in the care of these patients. PMID- 15663015 TI - Home care management of intestinal transplant recipients. AB - During the past 14 years, significant advances have been made in the field of intestinal transplantation. Patients' survival rates have increased, influencing referral patterns, and there has been a growing trend toward shortened hospitalizations with earlier discharge. Home care has also evolved during this same period, with development of the resources, training, and processes required for in-home management of patients with multiple complex needs. Together these trends have led to the ability to provide care and services to higher acuity patients in the outpatient and home setting after intestinal transplantation. In this article, we review the infusion management of intestinal transplant recipients before and after they go home and the rationale for the need for specialized home care services. PMID- 15663016 TI - Financial considerations insurance and coverage issues in intestinal transplantation. AB - OBJECTIVE: To increase healthcare workers' knowledge of reimbursement concerns. METHODS: Chronological survey of transplants reimbursed at the University of Nebraska Medical Center from December 1997 to October 2003, which include accounts of 30 patients who received intestine transplants. CONCLUSIONS: Gross billed hospital charges for the past 30 transplantations ranged from dollars 112094 to dollars 667597. Length of stay ranged from 18 to 119 days. Charges include organ procurement fees. All 30 intestine transplants were reimbursed by third-party healthcare coverage; combination of coverage; and/or patient and family payments, which resulted in adherence to financial guidelines prearranged by the hospital. Financial guidelines are usually cost plus a percentage. Thirteen transplantations occurred after April 2001, when Medicare made a national coverage decision to reimburse this form of transplantation. Since then, obtaining surgical authorization and reimbursement is easier. Most insurance companies and state public health agencies accept intestinal transplantations as a form of treatment. Researching transplant coverage before evaluation is essential to be compensated adequately. Financial guidelines will secure the fiscal success of the program. Educating patients to insurance and entitlements may reduce the out-of-pocket cost to patients. Transplant financial coordinators coordinate these efforts for the facility. The best coverage option for the patient and transplant programs is a combination of commercial healthcare coverage, secondary entitlement program, and fund-raising. With length of stay ranging up to 119 days and a lifetime of posttransplant outpatient follow-up care, it is beneficial for the facility to also have a fundraising program to assist patients. PMID- 15663017 TI - Is there a role for living donor intestine transplants? AB - The use of living donors with intestinal transplantation is controversial because it may not significantly improve candidate access to organs when intestine-only grafts are needed, and may involve excessive donor risk when combined liver intestine grafts are required. Although limited data are available for comparison at this time, graft and patient survival rates for intestinal transplantations using living donors are no different than for deceased donor transplantations. Potential benefits that may be provided to the intestine transplant recipient through the use of living donors include better HLA matching, shorter ischemia times, better bowel preparation, and better opportunities for introducing immunomodulatory strategies. Conversely, living intestine donors are at risk for mortality, significant morbidity, financial loss, and psychologic trauma. The long-term outcomes of living intestine donors have not yet been reported. Ultimately, these data are essential before the wider use of living donors can be advocated for intestinal transplantation. PMID- 15663018 TI - Quality of life 1 year after lung transplantation. AB - Lung transplantation is a viable therapeutic option for patients with end-stage lung disease. The focus of interest has shifted from advances in surgical techniques to improved quality of life for the transplant recipient. A prospective longitudinal repeated measures design was used to determine the impact of lung transplantation on quality of life and life satisfaction. Using 4 measurement points (before transplantation, after 8 weeks, and after 6 and 12 months), 61 patients were followed from before to 1 year after successful lung transplantation. Quality of life was measured using 2 generic (SF-36, Quality of Life Profile for Chronic Diseases) and 1 lung-specific (Saint George's Respiratory Questionnaire) questionnaires. All dimensions of the health-related scores improved significantly after transplantation, except bodily pain. Patients reported the most significant improvements until 6 months after transplantation. Lung transplantation had no influence on social functioning or role emotional. The Saint George's Respiratory Questionnaire ratings showed similar significantly better scores for transplant recipients at 6 months, and stagnations for most dimensions afterward. PMID- 15663019 TI - Validation and refinement of the Questionnaire for Lung Transplant Patients. AB - CONTEXT: The Questionnaire for Lung Transplant Patients was designed to assess symptoms and activity tolerance in lung transplant recipients during their post transplant evaluations. The initial psychometric evaluation determined that the questionnaire was clinically useful, reliable, and valid. OBJECTIVE: To report the results of further psychometric analyses in a new, expanded sample of lung transplant recipients and to demonstrate the iterative manner by which instruments are refined and tested. METHODS: Internal consistency, test-retest stability, convergent validity, factorial validity, and group differences attributable to age, gender, and transplant type were determined in a pooled sample of 177 lung transplant recipients. Sensitivity to change over time was measured in a subsample (n = 51) who provided repeated measures data. RESULTS: The Questionnaire for Lung Transplant Patients and its subscales were internally consistent (Kuder Richardson reliability of 0.73-0.95). Test-retest stability was high (intraclass correlations >0.70). Symptoms showed a significant curvilinear pattern with a tendency to decrease over time before rising again at the 12-month measurement for the total questionnaire (F=6.8, P=.012) and 2 subscales- Respiratory (F=5.6, P=.022) and Activities of Daily Living (F=19.7, P<.001). Convergent construct validity correlations ranged from 0.29 to 0.53 and were consistent with theoretical expectations. Factorial analysis confirmed 3 domains that coincided with the Respiratory, General, and Activities of Daily Living subscales. CONCLUSIONS: The Questionnaire for Lung Transplant Patients is a reliable and valid measure for assessing physical symptoms and activity intolerance after lung transplantation in individual recipients, recipients in aggregate, and comparison groups, on one occasion and serially over time. These results will guide future refinement and testing of the Questionnaire for Lung Transplant Patients. PMID- 15663020 TI - Influenza vaccine antibody responses in lung transplant recipients. AB - CONTEXT: Lung transplant recipients are at high risk of morbidity and mortality from influenza infection because of altered lung physiology and immunosuppression. Annual influenza immunization is recommended, but the ability to mount an antibody response may be limited by immunosuppressant medications. OBJECTIVE: To compare the antibody response rate to influenza vaccine in lung transplant recipients to healthy controls. DESIGN: Open label study. SETTING: Lung transplant clinic and General Clinical Research Center at a university hospital. SUBJECTS: Sixty-eight single and bilateral lung transplant recipients and 35 healthy controls were enrolled in October and November 2002. METHODS: Each individual underwent blood sampling before receiving the 2002-2003 influenza vaccine and 4 weeks later. Influenza antibody concentrations were measured by hemagglutination inhibition assay. Vaccine response rates (antibody concentration >40 hemagglutination units and at least 4-fold increase in antibody concentration) were compared using chi2. The influence of specific immunosuppressants on vaccine response was compared. RESULTS: The influenza vaccine response rate for lung transplant recipients was 29/68 (43%) and 22/35 (63%) for the healthy individuals (P < .05; chi2). Among the recipients, mycophenolate mofetil was associated with poorer influenza vaccine antibody response (> 40 hemagglutination units) (62% vs 91%; P = .01), whereas sirolimus (91% vs 63%; P = .02) was associated with better influenza antibody response compared to those not taking mycophenolate mofetil or sirolimus, respectively. CONCLUSION: Lung transplant recipients had lower influenza vaccine response rates than healthy individuals. Influenza vaccine antibody response is influenced by concomitant administration of mycophenolate mofetil or sirolimus. Future studies should measure protection from influenza infection conferred by immunization and alternative vaccination strategies. PMID- 15663021 TI - The future of family medicine project: embracing the future. PMID- 15663022 TI - Personalizing prevention: the U.S. Surgeon General's Family History Initiative. PMID- 15663023 TI - Off-label uses for selective serotonin reuptake inhibitors. PMID- 15663024 TI - Vitamin D insufficiency as a cause of hyperparathyroidism. PMID- 15663025 TI - Use of the term conjunctival icterus instead of scleral icterus. PMID- 15663026 TI - Glucocorticoids for treatment of croup. PMID- 15663027 TI - Antidepressants and smoking cessation. PMID- 15663028 TI - Hormonal contraceptives and weight gain. PMID- 15663029 TI - Imaging for suspected appendicitis. AB - Acute appendicitis is the most common reason for emergency abdominal surgery and must be distinguished from other causes of abdominal pain. Family physicians play a valuable role in the early diagnosis and management of this condition. However, the overall diagnostic accuracy achieved by traditional history, physical examination, and laboratory tests has been approximately 80 percent. The ease and accuracy of diagnosis varies by the patient's sex and age, and is more difficult in women of childbearing age, children, and elderly persons. If th diagnosis of acute appendicitis is clear from the history and physical examination, prompt surgical referral is warranted. In atypical cases, ultrasonography and computed tomography (CT) may help lower the rate of false-negative appendicitis diagnoses, reduce morbidity from perforation, and lower hospital expenses. Ultrasonography is safe and readily available, with accuracy rates between 71 and 97 percent, although it is highly operator dependent and difficult in patients with a large body habitus. While there is controversy regarding the use of contrast media and which CT technique is best, the accuracy rate of CT scanning is between 93 and 98 percent. Disadvantages of CT include radiation exposure, cost, and possible complications from contrast media. PMID- 15663030 TI - Procedural sedation in the acute care setting. AB - Many patients require sedation during diagnostic or therapeutic procedures. Ideally, procedural sedation minimizes the patient's awareness and discomfort while maintaining the patient's safety. Appropriate monitoring by trained personnel is the key to successful procedural sedation. These techniques should be used only by health care professionals skilled in managing complications, including cardiorespiratory compromise. It is important to take a complete history and perform a thorough physical examination, paying special attention to the selection of pharmacologic agents. Common sedative agents include etomidate, ketamine, fentanyl, and midazolam. These have become the agents of choice for procedural sedation because of their ease of use, predictable action, and excellent safety profiles. All patients requiring procedural sedation should be monitored by qualified staff at the bedside until they have recovered to an age appropriate baseline mental status and function. PMID- 15663031 TI - Insertion and removal of intrauterine devices. AB - The intrauterine device (IUD) is an effective contraceptive for many women. The copper-releasing IUD can be used for 10 years before replacement and is a good choice for women who cannot, or choose not to, use hormone-releasing contraceptives. However, some women experience an increase in menstrual blood loss and dysmenorrhea. The progestin-releasing IUD can be used for five years. It may reduce menorrhagia and dysmenorrhea, although some women have increased spotting and bleeding during the first months after insertion. The ideal candidates for IUD use are parous women in stable, monogamous relationships. Pregnancy, unexplained vaginal bleeding, and a lifestyle placing the woman at risk for sexually transmitted diseases are contraindications to IUD use. Insertion of the IUD can take place at any time during the menstrual cycle provided the woman is not pregnant. Before insertion, a bimanual examination and a sounding of the uterus are necessary to determine the uterus position and the depth of the uterine cavity. The IUD is inserted into the uterus according to individual protocols, with the threads cut at a length to allow the patient to check the device's position. Expulsion may occur with both types of IUDs. PMID- 15663032 TI - Understanding and interpreting serum protein electrophoresis. AB - Serum protein electrophoresis is used to identify patients with multiple myeloma and other serum protein disorders. Electrophoresis separates proteins based on their physical properties, and the subsets of these proteins are used in interpreting the results. Plasma protein levels display reasonably predictable changes in response to acute inflammation, malignancy, trauma, necrosis, infarction, burns, and chemical injury. A homogeneous spike-like peak in a focal region of the gamma-globulin zone indicates a monoclonal gammopathy. Monoclonal gammopathies are associated with a clonal process that is malignant or potentially malignant, including multiple myeloma, Waldenstrom's macroglobulinemia, solitary plasmacytoma, smoldering multiple myeloma, monoclonal gammopathy of undetermined significance, plasma cell leukemia, heavy chain disease, and amyloidosis. The quantity of M protein, the results of bone marrow biopsy, and other characteristics can help differentiate multiple myeloma from the other causes of monoclonal gammopathy. In contrast, polyclonal gammopathies may be caused by any reactive or inflammatory process. PMID- 15663033 TI - Intravenous fluids for children with gastroenteritis. PMID- 15663035 TI - Information from your family doctor. Kidney cysts. PMID- 15663034 TI - Glowing teeth. PMID- 15663036 TI - Information from your family doctor. Urinary tract infections. PMID- 15663037 TI - Information from your family doctor. Microscopic hematuria. PMID- 15663038 TI - Information from your family doctor. Polycystic kidney disease (PKD). PMID- 15663039 TI - Is this patient really incompetent? PMID- 15663040 TI - Radio-frequency ablation of large, nonresectable hepatic tumors. AB - Patients with nonresectable hepatic metastases who are not treated survive an average of 6 months. We report our experience with radio-frequency ablation (RFA) of nonresectable hepatic tumors 4 cm or greater in size. A retrospective chart review of all patients undergoing RFA of hepatic tumors 4 cm or greater from October 1, 1999, through August 31, 2002, was performed. Thirty-six patients were identified who underwent RFA of tumors 4 cm or greater. There were a total of 81 tumors ablated in the 36 patients. Twenty patients underwent RFA only; seven patients received RFA plus a wedge resection. Five patients were treated with RFA followed by chemoembolization. Two patients underwent RFA plus placement of a hepatic artery infusion pump. The median tumor size was 5 cm (range, 4-14 cm). Median patient follow-up was 26 months (range, 1-54 months). Patients with metastatic colon cancer had the longest median survival of 28 months (range, 1 and 48 months). The survival of primary hepatocellular carcinoma was worse with a median survival of 20 months (range, 1-36 months). At last follow-up, 11 (30%) of the patients remain alive and disease free. There were no perioperative deaths and one intraoperative complication. In our experience, RFA of larger tumors is effective and safe. Tumor size should not be an absolute contraindication to RFA of nonresectable hepatic tumors. PMID- 15663041 TI - Stent-graft repair for acute traumatic thoracic aortic rupture. AB - Traumatic rupture of the thoracic aorta is potentially life-threatening and leads to death in 75 to 90 per cent of cases at the time of injury. In high-risk patients, as traumatic injuries of the aorta combine with multiple associated injuries, endoluminal repair is now reported as a promising therapeutic strategy with encouraging results. This study determined the outcome of patients with traumatic thoracic aortic injury treated endovascularly during the past 7 years at our institution. Thirteen patients, 11 males and 2 females (mean age, 39 years; range, 19-82), with traumatic rupture of the otherwise unremarkable descending aorta (10 acute, 3 chronic), out of a series of 64 endovascular thoracic stent-graft procedures, were treated by implantation of Talent (n = 8), Vanguard (n = 5), and Excluder (n = 2) self-expanding devices between January 1996 and August 2003. The immediate technical success rate was 92 per cent (12/13). One patient showed a proximal endoleak type I, which was treated successfully by an additional stent-graft procedure. Secondary success rate was 100 per cent. The mortality rate was 0 per cent. Two additional stent-graft procedures were performed due to type I endoleaks after 18 and 28 months. There was no other intervention-related morbidity or mortality during the mean follow up time of 26.4 months' (range, 6-86). Endovascular stent-graft repair of traumatic thoracic aortic injuries is a safe, effective, and low-morbidity alternative to open thoracic surgery and has promising midterm results. PMID- 15663042 TI - Impact of neoadjuvant therapy on postoperative complications in patients undergoing resection for rectal adenocarcinoma. AB - Surgical resection continues to be the mainstay of treatment for rectal cancer. Neoadjuvant therapy (chemotherapy and radiation) has also been shown to be efficacious. The impact of preoperative chemotherapy and radiation on postoperative complications is unclear. The purpose of this study is to evaluate the relationship of neoadjuvant therapy on postoperative complications in patients undergoing a resection of rectal cancer. A total of 325 patients who underwent curative resection for rectal cancer from 1984 to 2001 were retrospectively reviewed. Only cases with complete data sets who had undergone surgery at this institution were evaluable (257). The patients were divided into groups based on the operative procedure performed; abdominoperineal resection (APR) versus sphincter-sparing (SS) procedures (LAR/Transanal) and whether or not preoperative chemotherapy or radiation was administered. There was no significant difference between complication rates for APR and SS with 19 per cent and 14 per cent, respectively. The preoperative therapy had no effect on complications after APR. However, the SS group showed 21 per cent of the patients who received radiation had complications compared to 11 per cent in those who did not (P = 0.087). Complications in the SS group included leaks, wound infections, abscess, embolism, cardiac dysrhythmias, and myocardial infarctions. The 30-day mortality was 1.9 per cent for the entire cohort with no clear difference between groups. There was no significant difference in complication rate between APR and SS. In the APR group, neoadjuvant therapy had no impact on the incidence of complications. However, the SS group did show a trend between preoperative chemotherapy and radiation and complication rate. However, this may not outweigh the advantages of preoperative therapy in this setting. PMID- 15663043 TI - Preoperative lymphoscintigraphy and internal mammary sentinel lymph node biopsy do not enhance the accuracy of lymphatic mapping for breast cancer. AB - Lymphoscintigraphy (LS) may identify sentinel lymph nodes (SLNs) outside the axilla. Biopsy of these nodes could improve the accuracy of lymphatic mapping (LM) for breast cancer (BC) if a significant number of tumor-positive extra axillary sentinel nodes are identified. To address this, we evaluated the impact of the use of preoperative LS and biopsy of axillary and internal mammary SLNs in women with BC. From October 1997 to July 2003, 175 women with breast cancer received technetium sulfur colloid, and images were obtained. Isosulfan blue dye was injected intraoperatively, and LM of the axillary and internal mammary lymph node basins was performed with a hand-held gamma probe. The anatomic location and histologic status of all SLNs identified with LS and LM was recorded, and the impact of the findings on LS and internal mammary LM were evaluated. LS showed SLN in 127/175 (73%) women and "hot spots" were found with the gamma probe in 142/175 (81%). At least one SLN was identified by LM in 168/175 (96%) patients, and 48/168 (29%) had metastases. One hundred sixty-two of 168 (96%) patients had SLN exclusively in the axilla. Only 10 of 175 (6%) women had internal mammary (IM) SLNs seen on LS. LM identified IM sentinel nodes in 6 of these 10 patients, but none were involved with tumor. Preoperative lymphoscintigraphy and biopsy of internal mammary sentinel nodes do not enhance the accuracy of lymphatic mapping for breast cancer. Omitting lymphoscintigraphy reduces the complexity and cost of lymphatic mapping without compromising the identification of tumor-positive sentinel nodes. PMID- 15663044 TI - Retrospective analysis of traumatic bladder injury: does suprapubic catheterization alter outcome of healing? AB - The role of suprapubic catheters in traumatic bladder injuries is not well defined. Current literature suggests that suprapubic catheters are only necessary with large intraperitoneal bladder ruptures. The purpose of this study is to show that all bladder injuries can be managed with transurethral catheterization alone with a similar leak rate, morbidity, and healing time. Retrospective analysis was done of all patients with traumatic bladder injuries at a level I urban trauma center from June 1992 through June 2003. Medical records were reviewed and data analyzed according to type of bladder catheterization (i.e., transurethral or suprapubic). All patients with urethral injuries were excluded. Fifty-six patients met inclusion criteria. Twenty-seven patients suffered penetrating bladder injuries, and 29 bladder injuries were secondary to blunt trauma. Forty seven patients were treated with transurethral catheter drainage, two were treated with suprapubic catheters, and seven were treated with both transurethral and suprapubic catheters. Forty patients had follow-up cystograms prior to catheter removal. Of the 47 patients treated with transurethral drainage alone, 3 (6%) developed urinary leaks. Of the 9 patients with suprapubic catheters, 2 (22%) developed urinary leaks. The mean time to removal of transurethral catheters was 15 days. The mean time to removal of suprapubic tubes was also 15 days. All study patients successfully healed their bladder injuries regardless of catheterization method. Suprapubic catheter drainage may increase morbidity without improving healing time. These results effectively support the decision to use transurethral catheter drainage alone in all patients with traumatic bladder injuries. PMID- 15663046 TI - Management of blunt pediatric hepatic and splenic injury: similar process, different outcome. AB - Though nonoperative management of stable children with blunt solid organ injury has been shown to be effective, we hypothesize that hepatic injuries represent a higher mortality risk than splenic injuries and that combination hepatosplenic injury is a marker of even greater mortality potential. A multi-institutional pediatric trauma registry was queried for all children with blunt injuries to the liver (H) or spleen (S), excluding those with severe brain injury. Incidence and mortality of H, S, and all combinations of H/S were compared. The mortality rate for patients with H was significantly higher (2.5%) than in patients with S (0.7%), and the overall mortality of H/S (8.6%) was significantly higher than both. Furthermore, the mortality of H/S injuries was associated with increasing severity of either the hepatic or splenic injury. In childhood injury, H and S occur with almost equal frequency but with different mortality, and H/S is less common but associated with increased mortality. PMID- 15663045 TI - Reoperative fundoplications are effective treatment for dysphagia and recurrent gastroesophageal reflux. AB - With wide application of antireflux surgery, reoperations for failed fundoplications are increasingly seen. This study was undertaken to document outcomes after reoperative fundoplications. Sixty-four patients, 26 men and 38 women, of average age 55 years+/-15.6 (SD), underwent reoperative antireflux surgery between 1992 and 2003. Fundoplication prior to reoperation had been undertaken via celiotomy in 27 and laparoscopically in 37. Both before and after reoperative antireflux surgery, patients scored their reflux and dysphagia on a Likert Scale (0 = none, 10 = continuous). Reoperation was undertaken because of dysphagia in 16 per cent, recurrent reflux in 52 per cent (median DeMeester Score 52), or both in 27 per cent. Failure leading to reoperation was due to hiatal failure in 28 per cent, wrap failure in 19 per cent, both in 33 per cent, and slipped Nissen fundoplication in 20 per cent. Laparoscopic reoperations were completed in 49 of 54 patients (91%); 15 had reoperations undertaken via celiotomy. Eighty-eight per cent of reoperations were Nissen fundoplications. With reoperation, Dysphagia Scores improved from 9.5+/-0.7 to 2.6+/-2.8, and Reflux Scores improved from 9.1+/-1.4 to 1.8+/-2.7. Seventy-nine per cent of patients with reflux prior to reoperation, 100 per cent with dysphagia, and 74 per cent with both noted excellent or good outcomes after reoperation. We conclude that failure after fundoplication occurs. Reoperations reduce the severity of dysphagia and reflux, thus salvaging excellent and good outcomes in most. Laparoscopic reoperations are generally possible. Reoperative fundoplications are effective treatment for dysphagia and recurrent gastroesophageal reflux, and their application is encouraged. PMID- 15663047 TI - Intramuscular myxomas: a clinicopathologic study with emphasis on surgical management. AB - Intramuscular myxomas (IMs) are rare myxoid tumors named for their abundance of noncollagenous mucinous stroma. IMs are benign tumors characterized by a paucity of cells, diminished vascularity, and minimal mitotic figures. The objectives of this study were to examine clinicopathologic features of IM and to discuss clinical management of these unusual soft tissue tumors. A 10-year retrospective study at Orlando Regional Medical Center was conducted from May 1993 to May 2003. A case report of a 48-year-old male with a right gluteal mass is presented. Four cases with histologically confirmed diagnosis of intramuscular myxoma were reported. Three patients presented with lesions of the lower extremity and one with an extrafascial IM of the eighth rib. One of the lower extremity cases was shown to be cellular myxoma, a recently characterized subtype of IM. All cases were treated with at least marginal excision of the mass. There have been no known recurrences. Our findings are consistent with described characteristics for IM with respect to anatomic location and gross and microscopic appearance. As it may be unreliable to differentiate IM from malignant myxoid tumors preoperatively, we favor excision with wide margins in the surgical management of these tumors. PMID- 15663048 TI - Overcrowded motor vehicle trauma from the smuggling of illegal immigrants in the desert of the Southwest. AB - Overcrowded motor vehicle crashes caused by the very active criminal enterprise of smuggling illegal immigrants in the desert of the Southwest is a recent and under-recognized trauma etiology. A computerized database search from 1990 through 2003 of local newspaper reports of overcrowded motor vehicle crashes along the 281 miles of Arizona's border with Mexico was conducted. This area was covered by two level I trauma centers, but since July 2003 is now served only by the University Medical Center. Each of these crashes involved a single motor vehicle in poor mechanical shape packed with illegal immigrants. Speeding out of control on bad tires, high-speed rollovers result in ejection of most passengers. Since 1999, there have been 38 crashes involving 663 passengers (an average of 17 per vehicle) with an injury rate of 49 per cent and a mortality rate of 9 per cent. This relatively recent phenomenon (no reports from before 1998) of trauma resulting from human smuggling is lethal and demonstrates the smugglers' wanton disregard for human life, particularly when facing apprehension. Even a few innocent bystanders have been killed. These crashes overwhelm a region's trauma resources and must be recognized when planning the distribution of trauma resources to border states. PMID- 15663049 TI - Chylous ascites after nephrectomy for trauma. AB - Chylous ascites has many etiologies including malignancy, infection, and inflammation. Management of this condition varies from conservative treatment to surgery. We present a case of chylous ascites after laparotomy for trauma and review the pathophysiology, etiology, diagnosis, and treatment options for this condition. PMID- 15663050 TI - Nonoperative management of neonatal splenic rupture. AB - Neonatal injury of the spleen is an uncommon but serious condition. Although the standard management of children with splenic injury is nonoperative, there is scant evidence in the literature to support handling neonates in the same way. We report a case of neonatal splenic rupture that was managed nonoperatively. A 3.6 kg full-term female born vaginally became tachycardic and pale on the second day of life. She had a distended abdomen and a hemoglobin of 5.8 g/dL. Her blood pressure remained within normal limits. She was transfused 20 cc/kg packed red blood cells. CT scan showed a grade V splenic rupture. Coagulopathy workup was negative. The assumption was that she had a ruptured spleen secondary to a traumatic delivery. She remained stable after the transfusion. It took 32 weeks for a CT scan to show complete healing. Traditionally, neonatal splenic rupture has been treated with splenectomy or splenorrhaphy. The first case of a neonate to be treated nonoperatively was reported in 2000. Our patient is only the second reported case. We chose to follow her with imaging to document healing and to rule out a tumor, as epidermoid cysts and hemangioendotheliomas can cause neonatal splenic rupture. We also review the literature to try to gain some insight into the management of this rare problem. PMID- 15663051 TI - A review of upper extremity fasciotomies in a level I trauma center. AB - The purpose of this study was to review recent experience with upper extremity fasciotomy. This study is a retrospective review of injured patients undergoing fasciotomy in the upper extremity at an urban trauma center. Mechanisms of injury, indications for and timing of fasciotomy, role of compartment pressures, techniques of closure, amputation rate, and patient outcomes were collected. Over a 3-year period, 201 fasciotomies were performed in the extremities of 157 injured patients, including 37 in the upper extremities of 27 patients. The mechanisms of injury were penetrating trauma in 13 patients (10 GSW, three SW), blunt or crush in 9, and burns (4 electric, 1 flame) in 5. Vascular injuries and fractures were present in 15 (56%) and 9 (33%) patients, respectively. The decision to perform a fasciotomy was a clinical one in 21 patients (75%), and only 6 patients had compartment pressures measured (range, 40-87 mm Hg; mean, 52). Upper extremity fasciotomy was performed at a first operation in 24 patients, whereas only 3 patients had a delayed fasciotomy from 6 to 48 hours after injury. Two patients died on the first hospital day, and 5 others had an amputation of an upper extremity at a mean of 8 days (range 2 to 26) after injury; however, no amputation was due to the failure to perform a timely fasciotomy. In the remaining 20 patients, closure of the fasciotomy site was performed at a mean of 9 days (range, 2 to 22) after injury, most commonly by split thickness skin grafting. Hospital stay was a mean of 20 days (range, 7-35). We conclude that 1) upper extremity fasciotomy accounts for less than 20 per cent of all fasciotomies performed; 2) a clinical decision is the most common reason for performing upper extremity fasciotomy, and only 11 per cent of patients underwent a delayed fasciotomy in this review; 3) the need for upper extremity fasciotomy is associated with a length of stay longer than expected for overall injury severity. PMID- 15663052 TI - Apoptosis and necrosis in the development of acute lung injury after hemorrhagic shock. AB - Acute lung injury can be a complication of hemorrhagic shock. Mechanisms of injury include neutrophil-derived inflammatory products that induce necrosis within the lung. Recent data has shown apoptosis, in addition to necrosis, as a pathway leading toward acute lung injury in shock models. This study quantitates apoptotic and necrotic cells in the lung after hemorrhagic shock. Mongrel pigs (20-30 kg) under general anesthesia (with pancuronium and pentobarbital) underwent instrumentation with placement of carotid and external jugular catheters. The animals were randomized to sham hemorrhage (n = 6) and to hemorrhagic shock (n = 7). The hemorrhagic shock group then underwent hemorrhage (40-45% blood volume) to a systolic blood pressure of 40-50 mm Hg for 1 hour. The animals were then resuscitated with shed blood plus crystalloid to normalization of heart rate and blood pressure. The animals were observed under general anesthesia for 6 hours after resuscitation, then sacrificed, and lungs were harvested. Lung injury parameters including histology (H&E stain), apoptosis [terminal deoxynucleotidyl transferase-mediated dUTP biotin nick end labeling (TUNEL)], and myeloperioxidase activity (spectrophotometric assay) were assessed. Hemorrhagic shock induced marked loss of lung architecture, neutrophil infiltration, alveolar septal thickening, hemorrhage, and edema in H&E staining. Furthermore, MPO activity, a marker for neutrophil infiltration and activation, was more than doubled as compared to controls (44.0 vs 20.0 Grisham units activity/g). Apoptosis (cell shrinkage, membrane blebbing, apoptotic bodies) and necrosis (cellular swelling, membrane lysis) in neutrophils, macrophages, as well as in alveolar cells was demonstrated and quantified by H&E staining use. Apoptosis was confirmed and further quantified by positive TUNEL signaling via digital semiquantitative analysis, which revealed a significant increase in apoptotic cells (16.0 vs 2.5 cells/hpf, shock vs control, respectively) and necrotic cells (16.0 vs 2.0 cells/hpf, shock vs control, respectively). Acute lung injury is a complex pathophysiologic process. Apoptosis in cells (neutrophils, macrophages, alveolar cells) is induced within the lung after hemorrhagic shock. The role of apoptosis in pulmonary dysfunction after hemorrhagic shock has yet to be determined. PMID- 15663053 TI - Admission serum albumin is predicitve of outcome in critically ill trauma patients. AB - There is a paucity of data evaluating serum albumin on admission as a predictor of outcome in adult trauma patients. Our objectives were to evaluate whether or not hypoalbuminemia on admission is a predictor of adverse outcome in trauma patients. Prospective data was collected daily on 1023 patients over a 2-year period. Patients were stratified by serum albumin level on admission, age, gender, injury severity, and comorbid conditions. Outcome was measured by ICU and hospital length of stay, ventilator days, incidence of infection, and mortality. Student t test, chi2, and multilinear regression analysis were used to determine level of significance. Blunt injuries accounted for the majority (78%) of the admissions. The mean age of the study population was 43+/-21 years with a mean Injury Severity Score (ISS) of 21.4+/-12. The majority of patients were male (74.5%). The mean albumin level on admission was 2.9+/-1.8. Five hundred ninety three (58%) patients were admitted with a serum albumin level of > or =2.6 as compared to 430 patients (42%) with an admission albumin level of <2.6. Patients with a lower serum albumin level were found to have a significantly greater ICU (17.1 vs 14.2 days) and hospital length of stay (17.3 vs 20.1 days, P'< 0.05), ventilator days (11.1 vs 13.5 days, P < 0.05), and mortality (P = 0.008) when matched for age and injury severity. The relative risk of infection and mortality increased greater than 2.5-fold in patients with increased age and low serum albumin when analyzed by multilinear regression analysis, P < 0.001. An admission serum albumin level of <2.6 g/dL is a significant independent predictor of morbidity and mortality in trauma patients. The combination of increased age and low albumin level was most predictive of infection and mortality. Early nutrition should be considered in these high-risk patients. PMID- 15663054 TI - Neoadjuvant chemotherapy for locally advanced breast cancer results in alterations in preoperative tumor marker status. AB - Neoadjuvant therapy followed by breast-conserving surgery has become an acceptable option for patients with locally advanced breast cancer. Although a distinct survival benefit has not been demonstrated using this approach, several questions have been raised following such therapy including its effects on receptor status and tumor markers. The current study retrospectively reviews estrogen receptor (ER), progesterone receptor (PR), and HER2-neu status in 55 consecutive patients treated by neoadjuvant chemotherapy. Preoperative and postoperative tumor markers were available for 43 of the 55 patients (78%). The pathologic complete tumor response rate (pCR) for this group was 19 per cent (8/43). Of those patients who did not achieve a pCR (n = 35), a change in tumor markers was seen in 25.7 per cent (9/35) of patients. When compared to a control group not undergoing neoadjuvant therapy, a significantly higher percent change in marker expression was noted in the neoadjuvant group (25.7% vs 5.9%, P = 0.046). ER, PR, and HER2-neu status remain important prognostic indicators for breast cancer. Tumor markers are useful in planning adjuvant therapy regimens. In this review, nearly 19 per cent of patients achieved a pCR. In patients not achieving a pCR, one in four patients had at least one change in tumor marker status. This study demonstrates the importance of establishing receptor and marker status prior to neoadjuvant therapy, as many patients will achieve a pCR and make tumor analysis impossible. Postoperative marker studies should be performed given the possibility of a change in status. The clinical relevance of this data will require further long-term follow-up. Until such data becomes available, caution should be considered when basing adjuvant therapy regimens on preoperative tumor marker studies alone. PMID- 15663055 TI - A nutrition support team led by general surgeons decreases inappropriate use of total parenteral nutrition on a surgical service. AB - The purpose of this study was to decrease the number of inappropriate orders for total parenteral nutrition (TPN) in surgical patients. From February 1999 through November 2000 and between July 2001 and June 2002, the surgeon-guided adult nutrition support team (NST) at a university hospital monitored new TPN orders for appropriateness and specific indication. In April 1999, the NST was given authority to discontinue inappropriate TPN orders. Indications, based on the American Society for Parenteral and Enteral Nutrition (ASPEN) standards, included short gut, severe pancreatitis, severe malnutrition/catabolism with inability to enterally feed > or =5 days, inability to enterally feed >50 per cent of nutritional needs > or =9 days, enterocutaneous fistula, intra-abdominal leak, bowel obstruction, chylothorax, ischemic bowel, hemodynamic instability, massive gastrointestinal bleed, and lack of abdominal wall integrity. The number of inappropriate TPN orders declined from 62/194 (32.0%) in the first 11 months of the study to 22/168 (13.1%) in the second 11 months (P < 0.0001). This number further declined to 17/215 (7.9%) in the final 12 months of data collection, but compared to the second 11 months, this decrease was not statistically significant (P = 0.1347). The involvement of a surgical NST was associated with a reduction in inappropriate TPN orders without a change in overall use. PMID- 15663056 TI - Three-year financial analysis of minimally invasive radio-guided parathyroidectomy. AB - Minimally invasive radio-guided parathyroidectomy (MIRP) has had a high success rate in correcting hypercalcemia, along with a low morbidity rate and high patient satisfaction. Our study was conducted in an attempt to analyze the cost effectiveness of MIRP in patients treated for primary hyperparathyroidism. We conducted a retrospective study of the total charges of three groups of patients undergoing surgery for previously untreated hyperparathyroidism in a single health care system. The three study groups included patients undergoing traditional bilateral neck exploration, MIRP, and neck exploration guided by intraoperative parathormone (PTH) assay. Charges were stratified into preoperative, intraoperative, and postoperative categories. The average total charge was $8,512 for MIRP, $12,723 for traditional neck exploration, and $13,011 for bilateral neck exploration with PTH assay. The decreased charge for MIRP was due to reduced operating room time, anesthesia costs, length of hospitalization, and an avoidance of the use of intraoperative tissue analysis and PTH assay. There was a greater than $4,000 savings with MIRP as compared with the more extensive neck exploration. These savings more than compensate for the cost of technology (preoperative sestamibi scan and intraoperative gamma probe) necessary to perform radio-guided parathyroidectomy. PMID- 15663057 TI - The coexistence of anaplastic and papillary carcinomas of the thyroid: a case presentation and literature review. AB - Papillary carcinomas of the thyroid are the most common malignant growth affecting the thyroid, currently representing 60-65 per cent of malignant thyroid neoplasm. Although the etiology of this neoplasm is unknown, they are thought to be related to neck irradiation, adenoma transformation, and Hashimoto thyroiditis. Papillary carcinomas are usually purely papillary but occasionally have areas of histologically different neoplasm, most commonly follicular. Overall, these carcinomas represent an indolent group of neoplasm and have an excellent prognosis. The occurrence of an anaplastic area in a papillary carcinoma represents the dedifferentiation of the primary neoplasm. This is an extremely rare occurrence and is considered to have negative prognostic significance. The purpose of this presentation is to discuss an unusual clinical case of a coexisting anaplastic and papillary carcinoma of the thyroid, diagnosed by fine needle aspiration (FNA) analysis presenting in a 67-year-old African American woman. Evaluation and treatment will be discussed. PMID- 15663058 TI - Applying rapid microbiology techniques in the war against bioterrorism. PMID- 15663059 TI - Towards an improved sterility test. PMID- 15663060 TI - Pharmaceutical development of an intravenous dosage form of diacetylmorphine hydrochloride. AB - A solid dosage form for multiple use was developed for parenteral administration of diacetylmorphine in a clinical trial on co-prescription of heroin to heroin addicts. A 300-mg/mL diacetylmorphine hydrochloride solution was lyophilised as 10-mL aliquots in 30-mL glass vials, to be reconstituted to 150 mg/mL with water for injection before use. Addition of bulking agents for improvement of the cake structure of the lyophilised product appeared unnecessary. Stability studies indicated good stability of the lyophilised product under prescribed storage conditions (25 degrees C, 60% relative humidity) and under more extreme conditions (40 degrees C, 75% relative humidity). The reconstituted product was found to be stable for six days at room temperature. Suitability of the product for multiple use was supported by the fact that the reconstituted product was found to be antimicrobially active. PMID- 15663061 TI - Photostability of symphatomimetic agents in commonly used infusion media in the absence and presence of bisulfite. AB - Previous studies have indicated that epinephrine is photodegraded in sulfite containing solutions by adrenochrome sulfonate. The present study was undertaken to investigate if the same degradation pattern is found for the symphatomimetic compounds isoprenaline, dopamine, and metaraminol. It was found that isoprenaline and dopamine followed the same degradation pattern as epinephrine--they are photodegraded more rapidly in the presence of bisulfite than in its absence, probably by formation of an aminochrome sulfonate. Metraminol was photostable under the given conditions. The photostability of isoprenaline, dopamine, and epinephrine was further investigated in the infusion media sodium chloride (9 mg/ml), glucose (50 mg/ml), Macrodex (dextran 70, 60 mg/ml, sodium chloride 9 mg/ml) and Ringer acetate. The influence of pH, conductivity, viscosity and media concentration were analyzed. In the presence of bisulfite it was found that Ringer acetate had a photodestabilizing effect, while Macrodex had a slight photoprotective effect, on the catecholamines. The pH of the media was shown to be the most important factor, a low pH being protective. Glucose had a destabilizing effect on epinephrine, dopamine, and isoprenaline in the absence of bisulfite. PMID- 15663062 TI - A novel approach to the statistical evaluation of media fill tests by the difference from no contamination data. AB - It is obvious that, in the media fill test and process simulation test, positive numbers in total fills should not have any significant difference from zero or asepsis. There are many reports concerning the definition of "sterility" or "asepsis." However, any scientific and practical methods to demonstrate "no significant difference from zero" have not been reported up to now. The existing criteria, such as "less than 0.1%," "less than 0.05%," and "less than two positives" are not appropriate to assure the integrity of processes, and sometimes lead to erroneous results. The purpose of this report is to demonstrate novel, reasonable and practical methods and criteria based on scientific and statistical consideration. According to the ISO 13408-1 Aseptic Processing of Health Care Products, Part 1 (1998), General Requirement for Aseptic Processing, the action level for the number of positive units in media fill tests is specified as 0.1%, and the alert level is 0.05%. In this paper it is shown that the existing ISO standard and other official methods are inappropriate in that zero contaminated units (sterile product) is outside the confidence range of probable distribution of contaminated units, even though the contaminated units are less than 0.1% in larger numbers of fills, and even less than 0.05%. This indicates that the limit of 0.1% or 0.05% is inappropriate in cases of larger numbers of fills. For sterile products, the number of contaminated units other than "zero" at the statistical confidence range must be judged to be contaminated units in process and as non-sterile. In order to harmonize this criteria-"no significant difference from zero"-with the existing criteria, the new criteria may be combined with only the existing criteria of 0.05% in smaller number of fills. PMID- 15663063 TI - Alcohol policies on college campuses. AB - State and local alcohol policies can minimize opportunities for people to use alcohol, thereby reducing consumption and alcohol-related problems. Little is known, however, about the prevalence of campus policies aimed at reducing college students' alcohol use and related problems. The authors surveyed school administrators in Minnesota and Wisconsin to assess the frequency of alcohol policies and whether institutional characteristics were likely to predict campus policies. They also compared administrators' responses to policies posted on college Web sites. Most schools prohibited beer kegs and provided alcohol-free housing for students. A minority of schools prohibited all alcohol use on campus or at Greek organizations or banned advertisements in school newspapers for alcohol or off-campus bars. The prevalence of policies varied with school characteristics, and agreement was poor between Web-site policy information and that provided by administrators. Further research on the prevalence of college alcohol policies might be useful for assessing trends and future prevention needs on campuses. PMID- 15663064 TI - Using social norms theory to explain perceptions and sexual health behaviors of undergraduate college students: an exploratory study. AB - The authors and associates conducted a social norms-based intervention targeting high-risk sexual behaviors among undergraduate students at 4 college campuses. Social norms theory predicts that widely held misperceptions may encourage risky behavior in a misguided attempt to conform to perceived norms and that information correcting these misperceptions will lead to a decrease in such behaviors. Students overestimated their peers' levels of sexual activity, numbers of partners, incidence of sexually transmitted infections, and rates of unintended pregnancies, but underestimated rates of condom use. Rates of HIV test taking, however, were accurately estimated. Although some components of sexual risk behaviors lend themselves well to social norms-based interventions, others, specifically inconsistent condom use and avoiding HIV tests, do not. Although no changes in reported beliefs or practices were apparent at the end of a 9-month intervention period, longer or modified interventions may be needed to make a fair assessment of the efficacy of this approach. PMID- 15663065 TI - Illicit use of prescribed stimulant medication among college students. AB - The authors investigated illicit use of stimulant medications at a midwestern university. They used a questionnaire to (a) examine the extent to which university students illicitly used stimulant medications prescribed for attention deficit hyperactivity disorder; (b) determine why college students abused such drugs; and (c) identify the factors that predicted illicit use of prescribed stimulant medication. Findings revealed that 17% of 179 surveyed men and 11% of 202 women reported illicit use of prescribed stimulant medication. Forty-four percent of surveyed students stated that they knew students who used stimulant medication illicitly for both academic and recreational reasons. Students reported they experienced time pressures associated with college life and that stimulants were said to increase alertness and energy. Regression analysis revealed that the factor that predicted men's use was knowing where to get easily acquired stimulant medication, whereas the main predictor for women was whether another student had offered the prescribed stimulants. PMID- 15663066 TI - The impact of a health campaign on hand hygiene and upper respiratory illness among college students living in residence halls. AB - Hand hygiene is a key element in preventing the transmission of cold and flu viruses. The authors conducted an experimental-control design study in 4 campus residence halls to determine whether a message campaign about hand hygiene and the availability of gel hand sanitizer could decrease cold and flu illness and school and work absenteeism. Their findings indicate that students who were exposed to the message campaign and provided with gel hand sanitizer increased their knowledge about the potential health benefits of hand washing and sanitizer use; they reported higher rates of hand washing and using sanitizer than did the control group. These students also experienced fewer cold and flu illnesses during the study than those in the control group and missed fewer class or work engagements because of colds or flu. Conducting a health promotion campaign in residence halls may therefore help prevent colds and flu and decrease absenteeism on university campuses. PMID- 15663067 TI - Internet use for health information among college students. AB - Use of the Internet to retrieve health information is increasingly common. The authors surveyed 743 undergraduate students at 2 academic institutions to examine their Internet use, health-seeking behaviors, and attitudes related to the use of the Internet to obtain health information. Fifty-three percent of the respondents indicated that they would like to get health information online, and 28% reported that they would like to attend a health program online. Overall, 74% of the students reported having ever received health information online, and more than 40% reported that they frequently searched the Internet for information. They used various search engines and multiple Web sites to find health information. Issues related to the credibility of the information on health Web sites were crucial considerations for students. The study found differences in Internet use for health information by gender and by level of Internet experience. PMID- 15663068 TI - Compassion for first-time phlebotomy patients. PMID- 15663069 TI - Systemic manifestations and treatment of brown recluse spider bites. PMID- 15663070 TI - Coombs-positive hemolytic anemia secondary to brown recluse spider bite: a review of the literature and discussion of treatment. AB - The bite of the brown recluse spider (Loxosceles reclusa) typically results in local, dermonecrotic skin lesions. Rarely, these bites may precipitate systemic disturbances of varying severity collectively known as systemic loxoscelism. The more severe systemic alterations attributed to the venom of this arachnid include hemolytic anemia, multiorgan failure, disseminated intravascular coagulation, or even death. Coombs-positive hemolysis associated with brown recluse spider bites has rarely been documented in the literature. We report 2 cases of systemic loxoscelism in young women associated with severe Coombs-positive hemolytic anemia and systemic symptoms requiring hospitalization. Both patients were treated with aggressive wound care, hematologic monitoring with blood transfusion, and intravenous fluid replacement. Recovery was excellent in both cases. We review the literature and discuss the controversies surrounding the treatment of more severe brown recluse bite reactions. PMID- 15663071 TI - Merkel cell carcinoma: a case report with treatment summary and updates. AB - Merkel cell carcinoma (MCC) is a rare primary cutaneous neoplasm known for its propensity to develop early regional and distant metastasis. Fewer than 400 cases occur annually in the United States. MCC ranks as the most deadly of cutaneous malignancies, with a fatality rate of approximately 25%. Because of its aggressive nature, MCC is often resistant to surgery, radiation, and chemotherapy regimens. Standardized treatment patterns have not been established, and difficulty arises finding appropriate treatment for the elderly, who comprise the majority of patients with MCC. PMID- 15663072 TI - A microsponge formulation of hydroquinone 4% and retinol 0.15% in the treatment of melasma and postinflammatory hyperpigmentation. AB - Disorders of hyperpigmentation such as melasma and postinflammatory hyperpigmentation (PIH) are common, particularly among people with darker skin types. Hydroquinone (HQ) bleaching creams are considered the gold standard for treating hyperpigmentation. Recently, a new formulation of HQ 4% with retinol 0.15% entrapped in microsponge reservoirs was developed for the treatment of melasma and PIH. Microsponges were used to release HQ gradually to prolong exposure to treatment and to minimize skin irritation. The safety and efficacy of this product were evaluated in a 12-week open-label study. A total of 28 patients were enrolled, and 25 completed the study. Study end points included disease severity, pigmentation intensity, lesion area, and colorimetry assessments. Adverse events also were recorded. Patients applied the microentrapped HQ 4% formulation to the full face twice daily (morning and evening). A broad-spectrum sunscreen was applied once in the morning, 15 minutes after application of the test product. Patients were evaluated at baseline and at 4, 8, and 12 weeks. The microentrapped HQ 4%/retinol 0.15% formulation produced improvement at all study end points. Improvement in disease severity and pigmentation intensity was statistically significant at weeks 4, 8, and 12 compared with baseline (P<.001). Lesion area and colorimetry measurements also were significantly improved at each visit (P<.001). Microentrapped HQ 4% was well tolerated, with only one patient discontinuing because of an allergic reaction, which was not considered serious. In this open-label study, microentrapped HQ 4% with retinol 0.15% was safe and effective. PMID- 15663073 TI - Evaluation of the inheritance of the complex vertebral malformation syndrome by breeding studies. AB - To investigate the congenital complex vertebral malformation syndrome (CVM) in Holstein calves, two breeding studies were performed including 262 and 363 cows, respectively. Cows were selected from the Danish Cattle Database based on pedigree and insemination records. Selected cows were progeny of sires with an established heterozygous CVM genotype and pregnant after insemination with semen from another sire with heterozygous CVM genotype. Following calving the breeders should state, if the calf was normal and was requested to submit dead calves for necropsy. In both studies, significantly fewer CVM affected calves than expected were obtained; a finding probably reflecting extensive intrauterine mortality in CVM affected foetuses. The findings illustrate increased intrauterine mortality as a major potential bias in observational studies of inherited disorders. PMID- 15663074 TI - Effect of anionic salt and highly fermentable carbohydrate supplementations on urine pH and on experimentally induced hypocalcaemia in cows. AB - The objective of this experiment was to determine the effect of dietary grain on calcium homeostasis. Six rumen-fistulated dairy cows with 3 or more previous lactations and no history of parturient paresis were randomly assigned to a sequence of diets in a crossover study with 4 periods of 10 days each. Dietary treatments were: A control ration consisting of wrap grass silage alone (1), the control ration supplemented with ammonium chloride and ammonium sulphate salt solution (2), control ration following a period with supplementation (3) and control ration supplemented with increasing amounts of barley from 4 to 10 kg/cow per day, expected to produce subclinical rumen acidosis (4). Daily intake of the diets was adjusted to 14 kg DM/cow per day. On day 11, the calcium-regulating mechanisms in cows were challenged until recumbency by a standardized intravenous EDTA infusion and cows were left to recover spontaneously. Anion supplementation and the feeding of highly fermentable carbohydrate lowered urine pH below 7.0 due to subclinical acidosis. During spontaneous recovery from EDTA induced hypocalcaemia, the cows more quickly regained a whole blood free calcium concentration of 1.00 mmol/L if they had most recently been supplemented with either anionic salts or with increasing amounts of barley, as compared to the basic ration. It is concluded that so-called slug-feeding or 'steaming up' with highly fermentable carbohydrates before parturition in milk fever susceptible cows enhanced calcium homeostasis similar to the effect seen in cows on anionic diets. PMID- 15663075 TI - Evaluation of copper supplementation to control Haemonchus contortus infections of sheep in Sweden. AB - A pen study was conducted to assess the effect of providing daily copper mineral supplement, or copper wire particle (COWP) capsules, on established or incoming mixed nematode infections in young sheep. For lambs with established (6 week old) infections, COWP resulted in 97% and 56% reduction of the adult and early L4 stages of H. contortus, respectively, compared with controls (p<0.001). Additionally there was a 74% reduction in Teladorsagia circumcincta infections in the COWP lambs compared with controls (p<0.01). However, no effect was observed when COWP were given at the commencement of a larval dosing period of 6 weeks. There was no significant effect of copper mineral supplement (given at the recommended rate to prevent Cu deficiency) on either established, or developing parasite infections. In addition, a field trial was conducted on a commercial farm to assess the effects of COWP in the management of recurrent H. contortus infections, but lack of parasites during the grazing season prevented an adequate assessment from being made. These results indicate that there is little, if any, benefit from a parasite control standpoint in recommending copper therapy, specifically to control parasites in Swedish sheep flocks. PMID- 15663076 TI - Effects of complex vertebral malformation on fertility in Swedish holstein cattle. AB - Complex vertebral malformation (CVM) is an autosomal recessive inherited defect in the Holstein breed. It causes intra-uterine mortality through the entire gestation period leading to repeat breeding and involuntary culling of cows and thereby economic losses. The defect was first reported in Denmark in 1999 and a direct DNA test for the defect has been available since February 2001. The aim of this study was to investigate if Holstein bulls heterozygous for the CVM gene had reduced reproductive performance, measured as non-return rate (NRR) and in a daughter fertility index. All genotyped Swedish Holstein bulls born between 1995 and 1999 were included. Altogether 228 bulls were analysed, of which 53 bulls, i.e. 23%, were confirmed CVM carriers. A statistically significant difference between carriers and non-carriers in the relative breeding value for NRR was observed for 168 days NRR (101.1+/-0.9 vs. 103.1+/-0.6, p<0.05). There was no difference for 28 days NRR whereas the difference approached significance for 56 days NRR. No significant effect of the paternal CVM genotype on the daughter fertility index was shown probably due to the complexity of traits this index is composed of. In conclusion, the study showed that carriers of the CVM defect have an inferior NRR compared with non-carriers. PMID- 15663077 TI - Experimental contagious caprine pleuropneumonia: a long term study on the course of infection and pathology in a flock of goats infected with Mycoplasma capricolum subsp. capripneumoniae. AB - Contagious caprine pleuropneumonia (CCPP) is a major threat to goat farming in parts of Africa and Asia. It classically causes acute high morbidity and mortality early in infection, but little is known of its long term epizootiology and course. In this study, 10 goats were inoculated with Mycoplasma capricolum subsp. capripneumoniae (M. capripneumoniae) and then mixed with 15 goats for contact transmission. The disease course was monitored in each goat for 56-105 days, whereafter the goats were killed and necropsied. Varying features signifying infection occurred in altogether 17 goats (7 inoculated, 10 in contact). Clinical signs were severe in 8 goats but no fatalities occurred. Only 6 goats had serum antibody titres against M. capripneumoniae in ELISA. Fourteen goats (5 inoculated, 9 in-contact) had chronic pleuropulmonary lesions compatible with CCPP at necropsy and 7 of those showed M. capripneumoniae antigen in the lung by immunohistochemistry. Neither cultivation nor PCR tests were positive for the agent in any goat. The results indicate that the clinical course of CCPP in a flock may be comparatively mild, M. capripneumoniae-associated lung lesions may be present at a late stage of infection, and chronic infection may occur without a significant serological response. PMID- 15663078 TI - BVDV and BHV-1 infections in dairy herds in northern and northeastern Thailand. AB - Bulk milk samples from 220 dairy herds were collected at 9 public milk collection centres in the northeastern and northern Thailand, and a subset of 11 herds was selected for individual testing. The samples were tested for presence of antibodies to BVDV and BHV-1 using an indirect ELISA. The results from the bulk milk testing demonstrated a moderate level of exposure to BVDV and BHV-1 (73% and 67%, respectively). However, the low proportion of herds with high BVDV antibody levels (13%) and the low within-herd seroprevalence of BVDV and BHV-1 in the 11 herds (24% and 5%, respectively), particularly among the young stock (15% and 0%, respectively), demonstrated a low prevalence of active BVDV infection and a low rate of reactivation of latent BHV-1. The presence of a self-clearance process was also indicated by the results from the individual testing. Moreover, a surprisingly low prevalence of BVDV and BHV-1 antibody-positive herds at one of the milk centres was found. This centre was established 5-10 years before the others. Our impression is that this reflects the self-clearance process, where consecutive replacement of imported infected animals without further spread has resulted in a nearly total elimination of the infections. Based on our experiences and on these results we are convinced that this process can continue if there is awareness of herd biosecurity. This is especially important in the context of a future intensification of the dairy production. PMID- 15663079 TI - Viral and bacterial pathogens in bovine respiratory disease in Finland. AB - Pathogens causing bovine respiratory tract disease in Finland were investigated. Eighteen cattle herds with bovine respiratory disease were included. Five diseased calves from each farm were chosen for closer examination and tracheobronchial lavage. Blood samples were taken from the calves at the time of the investigation and from 86 calves 3-4 weeks later. In addition, 6-10 blood samples from animals of different ages were collected from each herd, resulting in 169 samples. Serum samples were tested for antibodies to bovine parainfluenza virus-3 (PIV-3), bovine respiratory syncytial virus (BRSV), bovine coronavirus (BCV), bovine adenovirus-3 (BAV-3) and bovine adenovirus-7 (BAV-7). About one third of the samples were also tested for antibodies to bovine virus diarrhoea virus (BVDV) with negative results. Bacteria were cultured from lavage fluid and in vitro susceptibility to selected antimicrobials was tested. According to serological findings, PIV-3, BAV-7, BAV-3, BCV and BRSV are common pathogens in Finnish cattle with respiratory problems. A titre rise especially for BAV-7 and BAV-3, the dual growth of Mycoplasma dispar and Pasteurella multocida, were typical findings in diseased calves. Pasteurella sp. strains showed no resistance to tested antimicrobials. Mycoplasma bovis and Mannheimia haemolytica were not found. PMID- 15663080 TI - Cumulative risk of bovine mastitis treatments in Denmark, Finland, Norway and Sweden. AB - Data from the national dairy cow recording systems during 1997 were used to calculate lactation-specific cumulative risk of mastitis treatments and cumulative risk of removal from the herds in Denmark, Finland Norway and Sweden. Sweden had the lowest risk of recorded mastitis treatments during 305 days of lactation and Norway had the highest risk. The incidence risk of recorded mastitis treatments during 305 days of lactation in Denmark, Finland, Norway and Sweden was 0.177, 0.139, 0.215 and 0.127 for first parity cows and 0.228, 0.215, 0.358 and 0.204 for parities higher than three, respectively. The risk of a first parity cow being treated for mastitis was almost 3 times higher at calving in Norway than in Sweden. The period with the highest risk for mastitis treatments was from 2 days before calving until 14 days after calving and the highest risk for removal was from calving to 10 days after calving in all countries. The study clearly demonstrated differences in bovine mastitis treatment patterns among the Nordic countries. The most important findings were the differences in treatment risks during different lactations within each country, as well as differences in strategies with respect to the time during lactation mastitis was treated. PMID- 15663081 TI - A serological survey of ruminant livestock in Kazakhstan during post-Soviet transitions in farming and disease control. AB - The results of a serological survey of livestock in Kazakhstan, carried out in 1997--1998, are reported. Serum samples from 958 animals (cattle, sheep and goats) were tested for antibodies to foot and mouth disease (FMD), bluetongue (BT), epizootic haemorrhagic disease (EHD), rinderpest (RP) and peste des petits ruminants (PPR) viruses, and to Brucella spp. We also investigated the vaccination status of livestock and related this to changes in veterinary provision since independence in 1991. For the 2 diseases under official surveillance (FMD and brucellosis) our results were similar to official data, although we found significantly higher brucellosis levels in 2 districts and widespread ignorance about FMD vaccination status. The seroprevalence for BT virus was 23%, and seropositive animals were widespread suggesting endemicity, despite the disease not having being previously reported. We found a few seropositives for EHDV and PPRV, which may suggest that these diseases are also present in Kazakhstan. An hierarchical model showed that seroprevalence to FMD and BT viruses were clustered at the farm/village level, rather than at a larger spatial scale. This was unexpected for FMD, which is subject to vaccination policies which vary at the raion (county) level. PMID- 15663082 TI - Preliminary diagnostic survey on causes of prenatal mortality in Icelandic cattle. PMID- 15663083 TI - [Inmunohistochimic expresion of p53 protein and TTS prognostic value in the larynx cancer]. AB - We carried out an immunohistochemical study of p53 (DO7) expression in a series of 195 patients with laryngeal squamous cell carcinoma, treated and followed at the Department of Otolaryngology Virgen de la Salud Hospital (Toledo, Spain). In the cases with lymph node metastasis we also studied p53 expression at this site. We have investigated the value of p53 expression as a prognostic factor (tumor recurrence, deads due tu cancer and survival) and we have evaluated the relationship between p53 expression and other clinicopathologic characteristics. PMID- 15663084 TI - [Cochlear-vestibular ototoxicity by gentamicin. Report of a case and literature review]. AB - Gentamicin is an antibiotic member of the aminoglycosides family with well known potential to cause permanent ototoxicity by damaging more likely the cochlea vestibular system. We are reporting the clinical case of a 48 year-old female who was treated with such IV drug because of abdominal surgery. She had no otologic past history, and was complaining of tinnitus, periferical vertigo and progressive bilateral neurosensorial hearing loss, it est, cochlear-vestibular symptoms. After the withdrawol of gentamicin, the vestibular pathologic symptoms but not the hearing loss had improved, docummented by the persistance of the audiometric decay in the acute frequencies. Prevention is the best way to avoid iatrogenic lesions with thesse climps. PMID- 15663085 TI - [Rhinocerebral mucormycosis: differential diagnosis of invasive fungal sinusitis]. AB - Mucormycosis is an opportunistic infection caused by molds Mucoraceae of the family Phycomycetes. This invasive FS is found in debilitated hosts, most frequently in patients with hematologic malignancies, burn patients and diabetes mellitus. There are several differents forms: rhino-cerebral, pulmonary or disseminated. We describe a case of invasive fungal sinusitis (FS): Rhino-orbital cerebral mucormycosis, treated at our hospital with radical surgery and medical treatmen We reviewed the clinical presentation of the different types. PMID- 15663086 TI - [Hyperparathyroidism due to chronic renal failure. Analysis of 32 operated patients]. AB - We report our study of 32 patients diagnosed as hyperparathyroidism due to chronic renal failure (27 secondary forms and 5 tertiary ones), between June 1990 and 2003, that were sent from Nefrology Department for to surgical intervention. The following parameters have been analysed: Age, sex, clinical symptoms, kind of surgery performed, postoperative complications and anatomopathologic result (AP). On 64 nerves exposed, there were no recurrent nerve palsies. 12 patients suffered a symptomatic hypocalcemia (37,5%) although only one was permanent. The control of calcium, phosporus and calcitriol is essential to try to avoid a hyperplasia of the parathyroid glands. PMID- 15663087 TI - [Lingual tonsillitis. Report of 3 cases and literature review]. AB - Lingual tonsillitits is a relatively uncommon pathology that affects specially patients with previous palatine tonsillectomy and its clinical behaviour is even more notorious than an acute tonsillitis. We report 3 cases hospitalized by our ENT Department analysing the initial symptoms, complementary tests, treatment and clinical evolution. In one of the patients that diagnosis probably was secondary to a complication due to an infectious mononucleosis and finally he was operated by surgical cauterization the lingual tonsils. PMID- 15663088 TI - [Facial palsy caused by otitis media with effusion in a 9-year old child]. AB - We report the case of a 9 years old child diagnosed as right otitis media with effusion associated to homolateral and peripherical facial palsy, fact very uncommon in the reviewed literature. The patient was treated by miringotomy and insertion of transtympanic drainage recovering his facial function totally after 9 weeks. PMID- 15663089 TI - [Pronostic value of the MDM-2 protein expression in the larynx cancer]. AB - In this paper we carried out an immunohistochemical study of Mdm-2 (IF2) expression in a series of 195 patients with laryngeal carcinoma that were diagnosticated, treated and followed at the Department of Otolaryngology at Virgen de la Salud Hospital (Toledo, Spain). In the cases with lymph node metastasis we also studied Mdm-2 expression at this level. We also wanted to investigate the value of Mdm-2 expression as a prognostic factor (tumor recurrence, deads due to cancer and survival) and we have evaluated the relationship between Mdm-2 expression and other clinic and pathologic characteristics. PMID- 15663090 TI - [Non invasive fungal sinusitis: mycetoma and alergical fungal sinusitis]. AB - We have made a study about different cases of non-invasive fungal sinusitis (FS) treated at our hospital with surgery (endoscopic sinus approach) and medical treatment. We review two cases: Mycetoma and allergic fungal sinusitis, following the recent classification based on physiopathology, treatment and prognosis. We review the clinical presentation of the different types. PMID- 15663091 TI - [Cogan sindrome. Audio-vestibular findings in a patient and literature review]. AB - Cogan Sindrome is described as the association of non-syphilic interstitial keratitis with vestibulo-auditory symptoms in its typical form. We are reporting a case corresponding to a 32 years old male who consulted at first place for ophthalmic clinic and, two months later, affected of bilateral neurosensorial hearing loss with tinnitus and vestibular dysfunction. In spite of the corticoid treatment our patient has not improved. His hearing disorder was even worse after one year-follow up. A bibliographic review has been performed related to this pathology whose etiology is still unknown. PMID- 15663092 TI - [PTHi value in the surgical treatment of hyperparathyroidism]. AB - The prevalence of hyperparathyroidism as increased in the last 20 years due to a longer number of biochemical determinations, in which we observe in many cases hipercalcemia by casualty. The simptomatology of the primary hyperparathyroidism is very subtile and sometimes none in an 80% of the patients. Between november 1996 and march 2003 we have operated 55 parathyroidectomies. 36 patients (65,4%) had primary hyperparathyroidism, 16 (29,1%) secondary hyperparathyroidism and the rest (5,4%) tertiary hyperparathyroidism. Our hospital has a device of rapid detection of the parathyroid hormone level since february 2000, allowing us its satisfactory intraoperative determination in few minutes, evaluating the decrease to values inferior to the half of its basal level. PMID- 15663093 TI - [Acute resection in unusual cases of symptomatic aneurysms of the abdominal aorta]. AB - On three unusual cases of symptomatic abdominal aortic aneurysms, the authors demonstrate the need for satisfactory erudition of a vascular surgeon, who must manage the above serious conditions. In the first case, the authors deal with a symptomatic aneurysm of the abdominal aorta and a homolateral dystopy of the right kidney co-existence. In the second case, the authors describe a rare, nevertheless a very serious communication between a vein and an adjacent aneurysm. In the third case, the authors prefer the left-sided retroperitoneal approach in the management of the juxtarenal AAA in combination with a "hostile abdomen". PMID- 15663094 TI - [What to expect from the neoadjuvant therapy of the oesophageal carcinoma?]. AB - AIM: The aim of this study was to assess the neoadjuvant therapy of the oesophageal carcinoma firstly from the perspective of the immediate effect on the resection procedure itself and, secondly, from the perspective of the long-term results. METHODOLOGY: Prior to the resection of the oesophagus, the patients were administered the neoadjuvant therapy. The patients were allocated to three groups with the following branches of the cytostatic treatment: CDDP+FU, TAX+FU a CDDP. A smaller group with a less advanced disorder was treated only surgically, with the oesophageal resection. Furthermore, the study aimed at assessing chemoresistance of the tumors according to the MTT test and at correcting the individual branches appropriately. RESULTS: The total of 70 patients were operated from 2001 to V/2004. Out of this number, 15 oesophgeal resections without the neoadjuvant therapy were conducted. None of the patients exited, and fistules were the commonest complications. CONCLUSION: No differences in postoperative complications were reported between the groups with or without the neodjuvant therapy. Therefore, this therapy has no positive effect on the resection results themselves. The MTT test proved to be of low significance in the neodjuvant therapy assessment. Chemoresistance can be assessed only retrospectively and, furthermore, the results are likely to be affected by a relatively high sensitivity to the neodjuvant therapy itself. The pCR rate reaches 20%. Long-term results had not been assessed due to a short follow-up period. PMID- 15663095 TI - [Diagnosis of spontaneous arteriovenous fistula complicating aortoiliac aneurysm using spiral CT. Report of 3 cases]. AB - Acute spontaneous arterio-venous fistula complicating atherosclerotic abdominal aortic aneurysm (AAA) is rare. This life-threatening setting is observed in 1-2% of all AAAs and 2-4% of ruptured of AAAs. The triad of abdominal or lower back pain, pulsatile abdominal mass, and continual abdominal machinery-like bruit is seen only in half of cases. Currently, CT angiography is a noninvasive technique which enables a rapid and exact preoperative diagnosis. The authors describe three cases of aortoiliac aneurysm complicated by an acute arteriovenous fistula which were diagnosed using spiral CT. PMID- 15663096 TI - [Mini-invasive radiation-navigated parathyreoidectomy--MIRP]. AB - AIM: The aim was to introduce a new surgical method and to verify its validity. METHODOLOGY: In 20 patients whose ultrasound findings correlated with the MIBI scintigraphic results, a radiation-navigated parathyreoidectomy using a C-Track was conducted, following a radionucleotide application. RESULTS: From 2001 to 2003, 20 patients with hyperparathyreosis were operated. In all cases, an altered parathyroid gland with increased activity was detected using radionavigation. After the surgery, the calcium and the parathormone blood levels decreased. CONCLUSION: The MIRP is a miniinvasive surgical method indicated for the use in cases when hyperparathyreosis is confirmed and the ultrasound findings correlate with the MIBI-scintigraphic findings. PMID- 15663097 TI - [Reoperations following laparoscopic fundoplication]. AB - During an eight-year-period, 623 laparoscopic fundoplications were conducted in the Surgical Clinic in Olomouc. Out of the total number, 14 of them were reoperations and 4 reoperations were conducted on patients, primarily operated in other clinics. 9 patients were reoperated for dislocations of the rim transhiatally into the mediastinum, respectively aborally to the gastric fundus. 4 patients were operated for oesophageal constricions in the hiatus region and 2 patients for haemoperitoneum caused by haemorrhaging from a wound after the port removal. The remaining 3 patients had the reoperation indicated for a biliary reflux in the oesophageus, the rim loosening or for a non-standard primary procedure, conducted to treat the oesophageal reflux disorder in another clinic. This study discusses causes of individual complications and their treatment alternatives. The authors point-out prevention of complications based on following certain principles during the laparoscopic fundoplication procedures. PMID- 15663098 TI - [An aorto-enteric fistula]. AB - The most experienced surgeons should be called to conduct reoperations in the aorto-femoral region. It is always advisable to be aware of possible injuries to the urether and to prevent such cases by bandaging. A long-term follow-up of the patient in the same clinic, or at least consultation of his/hers condition helps to minimize any diagnostic hesitations as well as iatrogening harmings. Haemorrhaging aorto-enteric fistules must be surgically managed in clinics specialized in vascular surgery. PMID- 15663099 TI - [Non-surgical aproach in blunt injuries to the liver and spleen]. AB - INTRODUCTION: Non-surgical approach in blunt injuries of the liver and spleen in patients haemodynamically stable, is widely accepted as their therapeutical method of choice. Based on a retrospective analysis, the aim of this study is to assess the injured patients with blunt injuries of the liver and spleen who were treated non-surgically. SUBJECTS AND METHODOLOGY: 75 injured patients with blunt intraabdominal injuries were hospitalized from 2001 to 2003. The spleen injuries were diagnosed in 40 patients, out of which 12 of them suffered from concomittent injuries of other intraabdominal organs, the liver injuries were diagnosed in 37 cases, out of which 24 injuries were isolated. The non-surgical approach was indicated in haemodynamically stable patients with isolated injuries of the spleen and the liver of the ISt-IVth grade with haemoperitoneum up to 300 ml, diagnosed on a spiral CT examination. RESULTS: 47 (63%) patients were urgently operated for intraabdominal injuries, 7 of them (15%) exited within 30 days after the surgery. The non-surgical approach was indicated in 28 (37%) cases. In this group, 19 patients suffered from liver injuries, 7 from spleen injuries and 2 from pancreatic injuries. The severity of the injury was classified as the It degree in 7 patients, as the IInd degree in 7 patients, as the IIIrd degree in 7 patients and as the IVth degree in 5 patients. The non-surgical procedure failed in 5 (19%) cases of patients with IIIrd or IVth degree liver or spleen injuries. The failure was caused by a delayed bleeding from the injured spleen in 3 patients and by a continuous bleeding in the IVth degree liver injury and the bile collection from the biliary bladder rupture in the patient with the IIIrd degree liver injury. The spleen injury was treated using splenectomy. In the IVth degree liver injury a debridement of the dilacerated tissue of the 7th segment and a ligature of the injured branch of the right hepatic vein, were conducted. The patient with the IIIrd degree injury was treated using cholecystectomy with a suture of the liver rupture. CONCLUSION: The non-surgical approach to the treatment of the isolated blunt liver and spleen injuries is indicated for the injured patients who are haemodynamicaly stable, with Ist-IVth degree injuries, without or with minor (up to 300 ml) haemoperitoneum and with continuous intensive care. The non- surgical approach to the higher degree blunt liver and spleen injuries has higher treatment failure risk rates. PMID- 15663100 TI - [Jejunal diverticulosis as a rare cause of massive bleeding into digestive tract]. AB - The authors present a case of an elderly man, who was admitted to the hospital due to unusual cause of massive bleeding into digestive tube. During diagnostic work-up the source of gastrointestinal bleeding was localized into jejunal divertuculum. The patient was operated on and resection of involved jejunal segment with primary anastomosis was performed. The possible complications ofjejunal diverticulosis are discussed. PMID- 15663101 TI - [Radiofrequence ablation in contemporary surgery--our experience]. AB - Radio-frequency ablation (RFA) is one of the methods used in multimodale treatment of liver tumors. The opportunities for the use of RFA are more wide recently. Authors present their own experience with RFA in the treatment of liver tumors and also its use in the treatment of parenchymatous abdominal organs injury. They compare the different operative approaches for RFA with concentration to their advantages. PMID- 15663102 TI - [A retrorectal cystic tumor--case reports]. AB - Cystic retrorectal tumors are rare tumors with a risk potential for a malignant growth. Their diagnostics, location and decision on the surgical approach- abdominal, perineal or combined--are based on ultrasound, CT and MRI examination findings. A complete removal of the neoplasm without opening the wall of the cystic tumor is a pre-requisite for a good therapeutical result. PMID- 15663103 TI - [The onset of physiological activity in the stomach in the postoperative period. A comparative study with a prokinetic preparation, Ganaton]. AB - Postoperative gastroparesis decreases patient's postoperative comfort. The aim of this prospective study was to assess efficacy of the peroperatively administered prokinetic preparation Ganaton (Itopridi hydrochloridum, Abbott) on the postoperative gastroparesis. METHODOLOGY: This prospective study was conducted in the Ist Surgical Clinic of the 1st Medical Faculty in Prague in 2001-2001. The total of 64 patients took part in this study. The patients underwent either a non adjustable bandaging of the stomach via laparoscopy for a severe obesity, or a laparoscopic cholecystectomy. In the postoperative period, a subjective status of each patient, as well as objective examinations (auscultaion, gas excretion, stool excretion) and a percutaneous electrographic record were recorded. RESULTS: The patients after both the gastric bandaging and the laparoscopic cholecystectomy, demonstrated faster restoration of the physiological stomach contractions frequency in the group with itopride, compared with the placebo group. CONCLUSION: The prokinetic preparation was well tolerated and the authors did not record any undesirable side-effects. The preparation significantly speeded up restoration of the physiological stomach contractions frequency compared with the placebo group. Based on our results, its administration is a suitable part of the prophylaxis and treatment of the postoperative gastroparesis. PMID- 15663104 TI - [Surgical treatment of the colorectal carcinoma synchronous with liver metastases]. AB - The authors present their results of the surgical treatment of the synchrone liver metastases in the second stage and following three courses of chemotherapy. Although the trial group is small, counting 25 patients operated between April 2002 and October 2003 (i.e. 18 months), the procedure's clinical benefits may be deducted, based on the immediate postoperative results. PMID- 15663105 TI - Giant basal cell carcinomas: a result of neglect? AB - Although the occurrence of giant basal cell carcinoma in clinical practice is quite rare, there are some references to these malignancies in the literature. Our study includes 12 patients without previous treatment who had giant basal cell carcinoma with no evidence of metastatic dissemination. Depending on the anatomic region, we used a primary suture, local flap and split-thickness skin grafts for the reconstruction of the defect after the wide excision of the carcinoma. All patients have been followed up and no recurrence has been detected to date. Nine lesions diagnosed on the face (69%) were obvious, and therefore they should have been diagnosed at their early stage. In our series most of the patients had had tumors for more than 3 years (except 2 cases) with no history of radiation exposure or recurrence after previous treatment. Therefore our findings suggest to us that the primary reason for a tumor to achieve this giant size was neglect. PMID- 15663106 TI - GLADI: Gluteal Adipose Implant. A new technique for the reshaping of the gluteal trochanteric region. AB - The author describes a new technique of fat grafting of the buttocks, GLADI (Gluteal Adipose Implant) based on Coleman procedure, associated with the lipoplasty of the gluteal-trochanteric region in order to improve harmony of the lower limbs. 132 patients were treated by this technique with very good results, no major complication, and a durable increase of the gluteal region in 125 (94.7%) of them. GLADI technique is a valid alternative to silicone implantations and offers durable improvements in gluteal shape without the need of additional surgery or the risk of implant rupture. PMID- 15663107 TI - Recurrent branchial sinus tract with aberrant extension. AB - Second branchial cysts are the commonest lesions among congenital lateral neck anomalies. Good knowledge of anatomy and embryology are necessary for proper treatment. Surgical treatment involves resection of all branchial remnants, which extend laterally in the neck, medial to the sternocleidomastoid muscle with cranial extension to the pharynx and ipsilateral tonsillar fosa. However, infections and previous surgery can distort anatomy, making the approach to branchial anomalies more difficult. We present a case of a 17-year-old patient who presented with a second branchial tract anomaly with an aberrant extension to the midline and part of the contralateral neck. Previous surgical interventions and chronic infections may have been the primary cause for this aberrant tract. All head and neck surgeons should bear in mind that aberrant presentations may exist when reoperating on chronic branchial cysts fistulas. PMID- 15663108 TI - Mandibular reconstruction with local live vascularised bone transfer. AB - There are different methods to provide a skeletal support for mandibular reconstruction following the trauma or ablation for tumor in head and neck region. These methods vary from complicated techniques like vascularised free bone grafts to methods like free bone grafts, local vascularised bone grafts, cadaveric irradiated bone grafts and alloplastic material spacers including silastic rubber, polyvinyl plastics, ceramics, hydroxylapatite or metal. The technique employed solely depends upon many factors like expertise available, existing infrastructure, the available time factor and complexity of the defects including the type and extent of tissue loss in the particular patient. The sliding mandibulectomy is a simple method of mandibular reconstruction. This surgical technique is indicated for repairing short mandibular defects up to 9 cm in overall length including the mandibular arch for which extra vertical osteotomies are needed to bend the segment. More so it has its value in patients with poor prognosis and poor general conditions in whom longer and complicated mandibular reconstruction procedures with free vascularised bone flaps are not advisable. This article describes different types of sliding osteotomies used to harvest live bone flaps for rebuilding the mandibular defects including the arch of the mandible for patients who have undergone malignant tumor ablation. Primary reconstruction for the mandibular skeletal support was obtained in 18 cases with vascularised live-bone graft through various sliding osteotomies in different combinations. Complications, failures and success rate of sliding mandibulectomies and osteotomies have been analyzed and discussed in detail. PMID- 15663109 TI - The influence of age and exercise on the mobility of hand joints: 1: Metacarpophalangeal joints of the three-phalangeal fingers. AB - Mobility of metacarpophalangeal joints (MP) of the three-phalangeal fingers was measured in university students (52 males and 49 females), senior citizens (30 males and 30 females), and pianists (21 males and 31 females). We consider the student data file to be a control group with hand mobility unchanged by external influence. Extension, flexion, and total abduction in this group are greater in the left than the right hand. Only extensions were greater in females compared to males. In seniors, all types of studied movements are, with the exception of total abduction in females, lesser that in the control group. The difference is more apparent in males than in females. Intersexual difference showing better MP joint mobility in females than males is thus greater in seniors than in students; however, greater mobility of MP joints in left compared to right hand is less noticeable. Compared to students, pianists show greater finger abduction, and- less markedly--also passive and active extensions, while we did not notice difference in finger flexion. Intersexual difference in MP joint mobility in pianists were not recorded, and better mobility on the left hand compared to the right hand was evident only in dorso-palmar movements in males (the exception was total finger abduction, which is greater for the left hand in males as well as in females). In the three studied series we did not register differences in interfinger abduction between the left and right hand or between sexes. Active dorso-palmar MP joint range of motion is greatest in the little finger and smallest in the index finger, smallest in seniors and greatest in pianists. In all three groups, the range is greater in the left than the right hand and in females compared to males. PMID- 15663110 TI - The development of intracranial relations in patients with complete unilateral cleft lip and palate in relation to surgery method and gender aspect. AB - This study focuses on cranium development during puberty growth spurt in patients with complete unilateral cleft of the lip and palate (UCLPc) after use of various surgery methods. Next, this study focuses on cranium development differences between the genders in patients undergoing operations by the same method, as well as comparison of intracranial relations between impaired and healthy individuals. The work is based on longitudinal cephalometric measurement of X-ray films and it is interpreted by cluster analysis. It focuses mainly on mutual interrelations of linear dimensions (meaning longitudinal and vertical) with angular dimensions (characteristic of shape and position). Mutual relationships of linear characteristics were closer than its relationship with the angular characteristics and characteristics of shape and position were mutually closely related than with characteristics of the size. The development of the cranial shape in regards to the linear dimensions is influenced particularly by the depth of maxilla, the length of the ramus of the mandible, and the height of the upper face. The study confirms the least positive development of cranium in boys with a bone graft. Girls undergoing operations with the same technique have the advantage of earlier growth termination of most parts of the cranium, which makes it possible to maintain the results of the therapeutical compensation of the defect. PMID- 15663111 TI - Twin-to-twin transfusion syndrome: treatment with endoscopic laser surgery or serial amnio-reduction? PMID- 15663112 TI - The ethics of research on pregnant women: is maternal consent sufficient? PMID- 15663113 TI - Tubal evaluation in the investigation of subfertility: a structured comparison of tests. PMID- 15663114 TI - Failed sterilisation: evidence-based review and medico-legal ramifications. PMID- 15663115 TI - Delayed versus early pushing in women with epidural analgesia: a systematic review and meta-analysis. PMID- 15663116 TI - What do they know?: a content analysis of women's perceptions of trial information. AB - OBJECTIVE: To examine interpretations of study information by women participating in ORACLE, a trial of antibiotics in preterm labour. DESIGN: Questionnaire survey sent to women recruited to the ORACLE trial. SETTING: United Kingdom. POPULATION: A questionnaire was sent to 3074 ORACLE participants in a purposively selected sample of 55 collaborating maternity units, chosen to reflect a range of regions and of district general and teaching hospitals. METHODS: Content analysis was applied to verbatim text provided in response to an open question. Responses were also compared with a framework based on key points about the purpose of ORACLE. Closed questions were analysed using descriptive statistics. MAIN OUTCOME MEASURES: Participants' interpretations of the purpose of the study. RESULTS: A response rate of 61% was achieved, and 1462 participants provided written answers to a specific question on why the study was being carried out. Content analysis suggested that the information leaflet was highly valued as a source of information about the trial. There was evidence that women's interpretations of the purpose of the trial were not identical to those that the investigators intended. Of the five key points about the trial described in the information leaflet, 400 (27%) participants reported one key point, 550 (38%) two key points, 229 (16%) three key points and 23 (1.5%) four key points. None reported five key points and it was not possible to classify 46 responses (3%). Vague, confused understanding or poor recall were evident in 204 (14%) of responses. CONCLUSION: Although the ORACLE trial was run as a model of good practice at the time, this study suggests that it may not be possible to demonstrate full understanding of trial purpose and design by all participants. Emphasis should be on the provision of full information that involves consumers in its design and evaluation. PMID- 15663117 TI - Hypoxia induced activin secretion by the fetoplacental unit: differential responses related to gestation. AB - OBJECTIVE: To determine whether activin A levels reflect oxygen availability in basal and hypoxic conditions in the late pregnant fetus and newborn lamb. DESIGN: In vivo animal experimental study. SETTING: Department of Physiology, Monash University. POPULATION: Chronically catheterised fetal sheep in late gestation. METHODS: Fetal hypoxia was induced at 125 (n = 4), 135 (n = 4) or 145 days ('term'; n = 3) gestational age by maternal nitrogen exposure, for 4 hours, during which maternal and fetal arterial, and amniotic fluid samples were collected. Lambs (age one, five and eight days; n = 3) were exposed to 1 hour of hypoxia via nitrogen exposure. MAIN OUTCOME MEASURES: Activin A, prostaglandin E2 (PGE2) and cortisol were analysed in plasma and amniotic fluid, and whole blood was used to determine Pao2, Paco2, %O2, lactate and pH. RESULTS: Basal activin A concentrations in the fetal arterial circulation remained unchanged between 125 days (0.230 [0.10] ng/mL) and term (0.28 [0.10] ng/mL), as did fetal oxygen saturation (59.11% [4.74%] to 52.25% [4.84%]) and pH (7.35 [0.02] to 7.37 [0.02]). Moderate fetal hypoxia (50% fall in fetal arterial %O2) produced a significant increase in circulating activin A (2.05 [0.67] ng/mL) and a significant decrease in pH (7.27 [0.03]) at 125 days of gestation, however, at 135 and 145 days, activin A and pH remained unchanged. Fetal activin A concentration was significantly correlated with pH (P = 0.036) but not %O2 (P = 0.072). Hypoxia in the lambs did not alter circulating activin A. CONCLUSIONS: In response to hypoxia, activin A is increased in the circulation of 125-day-old fetuses, but not in older fetuses. Fetal arterial activin A levels sensitively reflect pH but not oxygen saturation, with increasing activin A in conditions of metabolic acidosis. PMID- 15663118 TI - Prevalence, nature, severity and correlates of postpartum depressive symptoms in Vietnam. AB - OBJECTIVE: To examine depressive symptomatology in women after childbirth in Ho Chi Minh City, Vietnam. DESIGN: A cross sectional survey. SETTING: Hung Vuong Obstetrics and Gynaecology Hospital and the Maternal, Child Health and Family Planning Center of Ho Chi Minh City, Vietnam. POPULATION: Mothers of infants aged +/- six weeks attending well-baby clinics. METHOD: Participants were recruited consecutively in the postnatal wards and invited to take part in the study at the first clinic visit. Individual structured interviews about health and social circumstances, including the Edinburgh Postnatal Depression Scale (EPDS) were administered during clinic visits. The interview schedule was translated into Vietnamese, back translated for verification and piloted. Interviewers were specifically trained members of staff of the two centres. MAIN OUTCOME MEASURES: EPDS scores and responses to structured questions about specific and non-specific symptoms. RESULTS: Of 506 women who participated, 166 (33%) had EPDS scores in the clinical range of > 12 and 99 (19%) acknowledged suicidal ideation. In a forward stepwise logistic regression analysis, 77% of cases with EPDS scores > 12 were correctly classified in a model which included unwelcome pregnancy, lack of a permanent job, < 30 days complete rest after childbirth, an unsettled baby, not being given special foods, avoiding proscribed foods and being unable to confide in their husbands. CONCLUSION: Depressive symptomatology is more prevalent among parturient women in Ho Chi Minh City, Vietnam than reported rates in developed countries and is at present unrecognised. PMID- 15663119 TI - Rupture of the sphincter ani: the recurrence rate in second delivery. AB - BACKGROUND: Injury to the genital tract sustained during childbirth can lead to transient or protracted morbidity. Attention should be paid to avoidable risk factors that can cause this complication. AIM: To analyse the recurrence, at a later delivery, of trauma to the genital tract, subsequent to perineal laceration of the sphincter ani (third or fourth degree), sustained at an earlier delivery. DESIGN: A population-based study. SETTING: In Sweden, 1973-1997 inclusive. POPULATION: All women with a vaginal, singleton delivery in Sweden. METHODS: The Medical Birth Registry, the National Board of Health and Welfare, was used to identify cases of ruptured sphincter ani. MAIN OUTCOME MEASURES: OR was calculated with 95% confidence interval. A stratified analysis was performed using the Mantel-Haenszel technique. Major end point Rupture of the sphincter ani (third or fourth degree) at second delivery. RESULTS: The incidence of anal sphincter rupture increased sixfold during the study period, from 0.5% in 1973 to 3.0% in 1997. Women who had sustained a laceration of this type ran a significantly increased risk of a recurrence at a later delivery. This effect persisted even after stratification for birthweight, year of birth, parity and maternal age (OR 4.74, 95% confidence interval 4.34-5.17). When only fourth degree rupture was considered (rupture of both anal sphincter and rectum), the corresponding figures were 6.52 (95% CI 5.29-8.04). This effect also persisted after stratification for birthweight, year of birth, parity and maternal age. The OR for giving birth a second time, subsequent to a third or fourth degree perineal laceration at first delivery, was 0.68 (95% CI 0.67-0.70). CONCLUSION: Our findings suggest that the risk of an anal sphincter rupture at delivery increases five to sevenfold when there has been a similar rupture at a previous delivery. Further study is needed before safe recommendations can be made concerning the subsequent mode of delivery to be adopted, following rupture in the sphincter ani at a previous birth. PMID- 15663120 TI - The relationship between delivery mode and mortality in very low birthweight singleton vertex-presenting infants. AB - OBJECTIVE: To investigate the factors associated with caesarean delivery and the relationship between mode of delivery and mortality in singleton vertex presenting very low birthweight (< or = 1500 g) live born infants. DESIGN: Observational population-based study. SETTING: Data collected from all 28 neonatal departments comprise the Israel National Very Low Birth Weight Infant Database. POPULATION: 2955 singleton vertex-presenting very low birthweight infants registered in the database from 1995 to 2000, and born at 24-34 weeks of gestation. METHODS: The demographic, obstetric and perinatal factors associated with caesarean delivery and subsequent mortality were studied. The independent effect of the mode of delivery on mortality was tested by multiple logistic regression. MAIN OUTCOME MEASURE: Mortality was defined as death prior to discharge. RESULTS: Caesarean delivery rate was 51.7%. Caesarean delivery was directly associated with increasing maternal age and gestational age, small for gestational age infants, maternal hypertensive disorders and antepartum haemorrhage, and was inversely related to premature labour and prolonged rupture of membranes. Factors associated with increased survival were increasing gestational age, antenatal corticosteroid therapy, maternal hypertensive disorders and no amnionitis. Mortality rate prior to discharge was lower after caesarean delivery (13.2% vs 21.8%), but in the multivariate analysis, adjusting for the other risk factors associated with mortality, delivery mode had no effect on infant survival (OR 1.00, 95% CI 0.74-1.33). In a subgroup with amnionitis, a protective effect of caesarean delivery was found. CONCLUSIONS: Caesarean delivery did not enhance survival of vertex-presenting singleton very low birthweight babies. Caesarean delivery cannot be routinely recommended, unless there are other obstetric indications. PMID- 15663121 TI - A qualitative study of conceptions and attitudes regarding maternal mortality among traditional birth attendants in rural Guatemala. AB - OBJECTIVE: To explore conceptions of obstetric emergency care among traditional birth attendants in rural Guatemala, elucidating social and cultural factors. STUDY: design Qualitative in-depth interview study. SETTING: Rural Guatemala. SAMPLE: Thirteen traditional birth attendants from 11 villages around San Miguel Ixtahuacan, Guatemala. METHOD: Interviews with semi-structured, thematic, open ended questions. Interview topics were: traditional birth attendants' experiences and conceptions as to the causes of complications, attitudes towards hospital care and referral of obstetric complications. MAIN OUTCOME MEASURES: Conceptions of obstetric complications, hospital referrals and maternal mortality among traditional birth attendants. RESULTS: Pregnant women rather than traditional birth attendants appear to make the decision on how to handle a complication, based on moralistically and fatalistically influenced thoughts about the nature of complications, in combination with a fear of caesarean section, maltreatment and discrimination at a hospital level. There is a discrepancy between what traditional birth attendants consider appropriate in cases of complications, and the actions they implement to handle them. CONCLUSION: Parameters in the referral system, such as logistics and socio-economic factors, are sometimes subordinated to cultural values by the target group. To have an impact on maternal mortality, bilateral culture-sensitive education should be included in maternal health programs. PMID- 15663122 TI - Incidence of breast abscess in lactating women: report from an Australian cohort. AB - OBJECTIVES: To report the incidence of breast abscess in lactating women. DESIGN: Data were combined from two studies (a randomised controlled trial and a survey) to provide a cohort of women for this report. SETTING: Women were recruited from two hospitals on one site in Melbourne, Victoria, the Royal Women's Hospital (public) (1999-2001) and Frances Perry House (private) (2000-2001). PARTICIPANTS: A total of 1193 of 1311 (91%) primiparous, English-speaking women from a diverse range of backgrounds, including those receiving public clinic care, private care and birth centre care. METHODS: A structured telephone interview was conducted on breastfeeding at six months postpartum. MAIN OUTCOME MEASURES: Lactational mastitis and breast abscess. RESULTS: Two hundred and seven women experienced mastitis. Five women developed a breast abscess: 0.4% of women who commenced breastfeeding (95% CI 0.14-0.98); 2.9% of women who took antibiotics for mastitis (95% CI 1.0-6.7). CONCLUSION: Although many authors estimate that 11% of women with mastitis develop a breast abscess, the incidence of lactating breast abscesses in Australia appears to be lower than reported in the past. Our estimate is that 3% of women with mastitis will develop a breast abscess. PMID- 15663123 TI - Carbohydrate solution intake during labour just before the start of the second stage: a double-blind study on metabolic effects and clinical outcome. AB - OBJECTIVE: To study the effects of oral carbohydrate ingestion on clinical outcome and on maternal and fetal metabolism. DESIGN: Prospective, double-blind, randomised study. SETTING: Leyenburg Hospital, The Hague, The Netherlands. POPULATION: Two hundred and two nulliparous women. METHODS: In labour, at 8 to 10 cm of cervical dilatation, the women were asked to drink a solution containing either 25 g carbohydrates or placebo. In a subgroup of 28 women, metabolic parameters were measured. MAIN OUTCOME MEASURES: Number of instrumental deliveries, fetal and maternal glucose, free fatty acids, lactate, pH, Pco2, base excess/deficit and beta-hydroxybutyrate. RESULTS: Drinking a carbohydrate enriched solution just before starting the second stage of labour did not reduce instrumental delivery rate (RR 1.1, 95% CI 0.9-1.3). Caesarean section rate was lower in the carbohydrate group, but the difference did not reach statistical significance (1% vs 7%, RR 0.2, 95% CI 0.02-1.2). In the carbohydrate group, maternal free fatty acids decreased and the lactate increased. In the umbilical cord there was a positive venous-arterial lactate difference in the carbohydrate group and a negative one in the placebo group, but the differences in pH and base deficit were comparable. CONCLUSION: Intake of carbohydrates just before the second stage does not reduce instrumental delivery rate. The venous-arterial difference in the umbilical cord suggested lactate transport to the fetal circulation but did not result in fetal acidaemia. PMID- 15663124 TI - Do maternity care provider groups have different attitudes towards birth? AB - OBJECTIVE: To compare family physicians', obstetricians' and midwives' self reported practices, attitudes and beliefs about central issues in childbirth. DESIGN: Mail-out questionnaire. SETTING/POPULATION; All registered midwives in the province, and a sample of family physicians and obstetricians in a maternity care teaching hospital. Response rates: 91% (n = 50), 69% (n = 97) and 89% (n = 34), respectively. METHODS: A postal survey. MAIN OUTCOME MEASURES: Twenty-three five-point Likert scale items (strongly agree to strongly disagree) addressing attitudes toward routine electronic fetal monitoring, induction of labour, epidural analgesia, episiotomy, doulas, vaginal birth after caesarean section (VBACs), birth centres, provision educational material, birth plans and caesarean section. RESULTS: Cluster analysis identified three distinct clusters based on similar response to the questions. The 'MW' cluster consisted of 100% of midwives and 26% of the family physicians. The 'OB' cluster was composed of 79% of the obstetricians and 16% of the family physicians. The 'FP' cluster was composed of 58% the family physicians and 21% the obstetricians. Members of the 'OB' cluster more strongly believed that women had the right to request a caesarean section without maternal/fetal indications (P < 0.001), that epidurals early in labour were not associated with development of fetal malpositions (P < 0.001) and that increasing caesarean rates were a sign of improvement in obstetrics (P < 0.001). The 'OB' cluster members were more likely to say they would induce women as soon as possible after 41 3/7 weeks of gestation (P < 0.001) and were least likely to encourage the use of birth plans (P < 0.001). The 'MW' cluster's views were the opposite of the 'OBs' while the 'FP' cluster's views fell between the 'MW' and 'OB' clusters. CONCLUSIONS: In our environment, obstetricians were the most attached to technology and interventions including caesarean section and inductions, midwives the least, while family physicians fell in the middle. While generalisations can be problematic, obstetricians and midwives generally follow a defined and different approach to maternity care. Family physicians are heterogeneous, sometimes practising more like midwives and sometimes more like obstetricians. PMID- 15663125 TI - Risks of induction of labour in women with a uterine scar from previous low transverse caesarean section. AB - OBJECTIVE: Recent studies have shown that among women with uterine scars from previous caesarean section of any type, induction of labour is associated with increased risk of uterine rupture compared with spontaneous labour. We have assessed the risk of uterine rupture in a cohort of women with a previous low transverse caesarean section in whom induction and management of labour were performed according to a strict protocol. DESIGN: Cohort study. SETTING: University Hospital. POPULATION: All women with a singleton pregnancy and a previous low transverse caesarean section requiring induction of labour from 1/1/1992 to 12/30/2001 (n = 310) were compared with a control cohort during the same study period constituted of women with a previous low transverse caesarean section in spontaneous labour (n = 1011). METHODS: Clinical characteristics and rate of uterine rupture of women with previous caesarean section undergoing induction of labour were compared with those of women with previous caesarean section in spontaneous labour. MAIN OUTCOME MEASURE: Incidence of uterine rupture. RESULTS: Uterine rupture occurred in 0.3% in the previous caesarean section--induction group versus 0.3% in the previous caesarean section- spontaneous labour group (P = 0.9). Logistic regression analysis showed no significant difference in the rate of uterine rupture between the induction and spontaneous labour group (P = 0.67) after controlling for maternal age, parity, duration of labour, gestational age at delivery and birthweight. CONCLUSION: Among women with a previous low transverse caesarean section, induction of labour is not associated with significantly higher rates of uterine rupture compared with spontaneous labour, provided a consistent protocol with strict criteria for intervention is adopted. PMID- 15663126 TI - Trends in the cause of late fetal death, 1982-2000. AB - BACKGROUND: Progress in reducing late fetal deaths has slowed in recent years, despite changes in intrapartum and antepartum care. OBJECTIVES: To describe recent trends in cause-specific fetal death rates. DESIGN: Retrospective cohort study. SETTING: North of England. POPULATION/SAMPLE: 3,386 late fetal deaths (> or = 28 weeks of gestation and at least 500 g), occuring between 1982 and 2000. METHODS: Data on deaths were obtained from the Northern Perinatal Mortality Survey. Data on live births were obtained from national birth registration statistics. Rate ratios (RR) and 95% confidence intervals (CI) for fetal deaths in 1991-2000 compared with 1982-1990 were calculated. MAIN OUTCOME MEASURES: Cause-specific late fetal death rates per 10,000 total births. RESULTS: Mortality in singletons declined from 51.5 per 10,000 births in 1982-1990 to 42.0 in 1991 2000 (RR 0.82, 95% CI 0.76-0.87). There was a greater decline in multiples, from 197.9 to 128.0 per 10,000 (RR 0.65, 95% CI 0.51-0.83). In singletons, the largest reductions occurred in intrapartum-related deaths, and deaths due to congenital anomalies, antepartum haemorrhage and pre-eclampsia. There was little change in the rate of unexplained antepartum death occurring at term (RR 0.97, 95% CI 0.84 1.11) or preterm (RR 0.94, 95% CI 0.82-1.07), these accounting for about half of all late fetal deaths. Unexplained antepartum deaths declined in multiple births and in singletons of birthweight < 1500 g. CONCLUSIONS: While late fetal mortality due to many specific causes has declined, unexplained antepartum death rates have remained largely unchanged. Improved identification of deaths due to growth restriction and infection, which may otherwise be classified as unexplained, is important. Further investigation of the underlying aetiologies of genuinely unexplained deaths is needed. PMID- 15663127 TI - Management of genital chlamydial infections at termination of pregnancy services in England and Wales: where are we now? AB - OBJECTIVE: To determine the range of policies and practices related to the management of genital chlamydial infection employed at termination of pregnancy services in England and Wales. DESIGN: Cross-sectional descriptive study. SETTING: England and Wales. POPULATION: Termination of pregnancy providers. METHODS: Survey questionnaire administered to termination of pregnancy providers. MAIN OUTCOME MEASURES: Policies and practices for the management of genital chlamydial infection in women seeking termination of pregnancy with comparison to the national guidelines of the chief medical officer (CMO) and the Royal College of Obstetricians and Gynaecologists (RCOG). RESULTS: One hundred and thirty-eight (48%) practices responded to the survey, with representation across England and Wales. Policies for screening and/or treatment of chlamydial infection existed for 70% of providers. We found three practice patterns for the management of genital chlamydial infection among termination of pregnancy attenders: 70% of providers tested their own attenders prior to termination and treated if necessary; about 25% of providers administered prophylaxis without testing; and a small number of providers (< 5%) neither tested nor treated attenders. CONCLUSION: These patterns may be the result of differences in the CMO and RCOG guidelines. Given the impact of untreated genital chlamydial infection in women attending for termination, consistent recommendations from the CMO and RCOG may encourage uniform practice for the management of chlamydial infection in this vulnerable population. PMID- 15663128 TI - Ignorance of electrosurgery among obstetricians and gynaecologists. AB - OBJECTIVE: The purpose of this study was to assess the level of skill of laparoscopic surgeons in electrosurgery. DESIGN: Subjects were asked to complete a practical diathermy station and a written test of electrosurgical knowledge. SETTING: Tests were held in teaching and non-teaching hospitals. SAMPLE: Twenty specialists in obstetrics and gynaecology were randomly selected and tested on the Monash University gynaecological laparoscopic pelvi-trainer. Twelve candidates were consultants with 9-28 years of practice in operative laparoscopy, and 8 were registrars with up to six years of practice in operative laparoscopy. Seven consultants and one registrar were from rural Australia, and three consultants were from New Zealand. METHODS: Candidates were marked with checklist criteria resulting in a pass/fail score, as well as a weighted scoring system. We retested 11 candidates one year later with the same stations. MAIN OUTCOME MEASURES: No improvement in electrosurgery skill in one year of obstetric and gynaecological practice. RESULTS: No candidate successfully completed the written electrosurgery station in the initial test. A slight improvement in the pass rate to 18% was observed in the second test. The pass rate of the diathermy station dropped from 50% to 36% in the second test. CONCLUSION: The study found ignorance of electrosurgery/diathermy among gynaecological surgeons. One year later, skills were no better. PMID- 15663129 TI - The validity of transvaginal ultrasound measurement of endometrial thickness: a comparison of ultrasound measurement with direct anatomical measurement. AB - OBJECTIVES: To compare transvaginal ultrasound measurements of endometrial thickness with direct anatomical measurements and consider the implications of these findings on clinical practice. DESIGN: Prospective observational study using two modalities blinded to each other's findings. SETTING: Singleton Hospital, Swansea, a medium-sized District General Hospital. SAMPLE: Forty-seven women admitted for hysterectomy. METHODS: All women underwent transvaginal ultrasound scan to measure the endometrial thickness within 16 hours of surgery. Anatomical measurement of the fresh specimen was carried out immediately after surgery. MAIN OUTCOME MEASURES: Agreement between ultrasound and anatomical measurements of the endometrial thickness. RESULTS: No ultrasound measurement was possible in 15% of patients. When both values were obtained, transvaginal ultrasound measurements were > 2 mm different from the ruler measurement in 13/40 (33%) with an obvious tendency of the ultrasound scan to over-estimate the endometrial thickness. The mean difference between the two measurements was -0.8 mm (limits of agreement -7.1 to +5.5 mm). The discrepancy was greater in women with endometrial thickness < or = 5 mm (-1.6 mm, limits of agreement -5.7 to +2.6 mm) compared with that in women with endometrial thickness > 5 mm (-0.2 mm, limits of agreement -7.6 to +7.2 mm). Kappa statistics showed good agreement between the two measurements in discriminating between thin and thick endometrium in 77% (kappa = 0.55). Transvaginal ultrasound misdiagnosed a thick endometrium as thin in 3/40 (8%) and misdiagnosed a thin endometrium as thick in 6/40 (15%). CONCLUSIONS: Transvaginal ultrasonography is of limited value as a screening test for abnormal endometrium in patients with postmenopausal bleeding if the only parameter of normality is an endometrial thickness of 5 mm or less. PMID- 15663130 TI - The use of levonorgestrel-releasing intrauterine system for treatment of menorrhagia in women with inherited bleeding disorders. AB - BACKGROUND: The levonorgestrel-releasing intrauterine system (LNG-IUS) is used commonly by gynaecologists as a contraceptive and to treat menorrhagia. However, its efficacy has not been examined in women with inherited bleeding disorders. DESIGN: A prospective pilot study. SETTING: A teaching hospital in north London with a designated haemophilia centre. POPULATION: Female patients with a known inherited bleeding disorder. METHODS: Sixteen women with subjective and objective menorrhagia caused by inherited bleeding disorders (13 von Willebrand's Disease, two factor XI deficiency and one Hermansky-Pudlak syndrome), who had previously undergone unsuccessful medical treatment were followed up for nine months after LNG-IUS insertion. Bleeding was measured by pictorial chart and haemoglobin concentration. RESULTS: All women reported that their periods were improved, pictorial chart scores were lower and 56% became amenorrhoeic. None reported side effects. CONCLUSION: The LNG-IUS is well tolerated and effective and improves quality of life. PMID- 15663131 TI - Women's perceived experiences of abuse in the health care system: their relationship to childhood abuse. AB - OBJECTIVES: The aim of this study was to determine whether there was an association between any lifetime experiences of emotional, physical and/or sexual abuse and perceived abuse in the health care system. Furthermore, we wanted to ascertain if adult victims of perceived abuse in the health care system reported exposure to childhood emotional, physical and/or sexual abuse more often than non victims did. DESIGN: A cross sectional questionnaire study. The first hypothesis was tested in the total sample, and the second hypothesis in a case-control analysis. The cases were those women who reported perceived experiences of abuse in the health care system as adults. Exposure was defined as experience of emotional, physical and/or sexual abuse in childhood. SETTINGS: Three Swedish gynaecological clinics. SAMPLE: A total of 2439 gynaecology patients (response rate 81%). METHODS: Postal questionnaire. MAIN OUTCOME MEASURE: Associations between experiences of emotional, physical and/or sexual abuse, and perceived abuse in the health care system; all operationalised in The NorVold Abuse Questionnaire (NorAQ). RESULTS: A general association was found between lifetime experiences of emotional, physical and/or sexual abuse and perceived abuse in the health care system. Adult victims of abuse in the health care system reported experiences of emotional, physical and/or sexual abuse in childhood more often than non-victims did. These findings also held after adjustment for age and educational level. CONCLUSIONS: We found associations between experiences of any lifetime abuse and perceived abuse in the health care system. Adult victimisation in the health care system was associated with childhood exposure to emotional, physical and/or sexual abuse. These associations call for attention and need to be further investigated. PMID- 15663132 TI - Testing positive for human papillomavirus in routine cervical screening: examination of psychosocial impact. AB - OBJECTIVE: To examine the psychosocial impact of testing positive for high risk human papillomavirus (HPV) among women attending primary cervical screening. DESIGN: Cross sectional survey. Measures were taken at baseline and one week after the receipt of HPV and cytology screening results. SETTING: Well women's clinic in London, UK. Population or Sample Four hundred and twenty-eight women aged 20-64 years. METHODS: Postal questionnaire survey. MAIN OUTCOME MEASURES: Psychosocial and psychosexual outcomes were anxiety, distress and feelings about current, past and future sexual relationships. RESULTS: Women with normal cytology who tested positive for HPV (HPV+) were significantly more anxious and distressed than women who were negative (HPV-) using both a state anxiety measure [F(1,267) = 29, P < 0.0001] and a screening specific measure of psychological distress [F(1,267) = 69, P < 0.0001]. Women with an abnormal or unsatisfactory smear result, who tested HPV+, were significantly more distressed than HPV- women with the same smear result [F(1,267) = 8.8, P = 0.002], but there was no significant difference in state anxiety. Irrespective of cytology result, HPV+ women reported feeling significantly worse about their sexual relationships. Approximately one-third of women who tested positive reported feeling worse about past and future sexual relationships compared with less than 2% of HPV- women. CONCLUSION: The findings suggest that testing positive for HPV may have an adverse psychosocial impact, with increased anxiety, distress and concern about sexual relationships. Psychosocial outcomes of HPV testing need further investigation and must be considered alongside clinical and economic decisions to include HPV testing in routine cervical screening. PMID- 15663133 TI - The role of clinical follow up in early stage cervical cancer in South Wales. AB - OBJECTIVE: To assess the effectiveness of clinical follow up after primary surgery for early stage cervical cancer. DESIGN: Retrospective analysis of clinical follow up after radical hysterectomy and node dissection for early stage cervical cancer. SETTING: Gynaecological Oncology Cancer Centre. SAMPLE: Two hundred and ninety-one patients who underwent surgery for cervical cancer. METHODS: Follow up data were collected retrospectively from hand-searched patients notes, as well as a computerised database (Information System for Clinical Organisation [ISCO]). The data were analysed using the SPSS for windows (SPSS, Chicago, Illinois) statistics package, using chi2, Kaplan-Meier life tables and Cox Linear regression analysis. MAIN OUTCOME MEASURES: To determine whether routine follow up was useful for detecting early recurrent disease. RESULTS: Two hundred and ninety-one patients treated by radical hysterectomy and node dissections were followed up. The cumulative five-year survival for all cases in our series was 80% and 53/291 patients (18.2%) were found to have recurrent disease. The median period from surgery to recurrence was 17.6 months (3.0-60.0). Seven patients with recurrence were detected at a routine follow up examination, and two out of seven of the patients were asymptomatic. Detection of the recurrence on routine follow up was not an independent prognostic factor for survival when compared with age, stage and whether the patient received post operative adjuvant therapy. CONCLUSIONS: Routine follow up in patients following radical hysterectomy and node dissection for early stage cervical cancer is not a sensitive way of detecting recurrent disease, as a high proportion of patients were symptomatic at the time of detection. As there are other reasons for follow up, we propose alternative methods of structuring the programme. PMID- 15663134 TI - Gestational trophoblastic disease: is intensive follow up essential in all women? AB - OBJECTIVE: To determine the timescale of the registration process for gestational trophoblastic disease and its impact on hCG level at registration and subsequent need for chemotherapy. DESIGN: A prospective observational study using a standardised protocol for registration, assessment and treatment for molar pregnancy. SETTING: A supra-regional tertiary referral centre for gestational trophoblastic disease. PARTICIPANTS: A total of 2046 consecutive women registered between January 1994 and December 1998 with a diagnosis of molar pregnancy. METHODS: Data at and after registration, collected prospectively on a computerised database, were statistically analysed (by multiple logistic regression and ANOVA). MAIN OUTCOME MEASURES: Relationship between length of time to and hCG value at registration; also the subsequent need for chemotherapy. RESULTS: A total of 2046 women with a diagnosis of molar pregnancy were registered in the study period. The mean time interval between first evacuation and registration at the referral centre was 47 days (median 37, range 0-594). One hundred and five out of 2046 (5.1%) women needed chemotherapy. Sixty-three precent of the women (1296 out of 2046) had a normal level of urinary hCG (less than 40 IU/24 hours) at the time of registration and only one (0.08%) needed chemotherapy. Binary logistic regression analysis showed a statistically significant relationship between time to registration, hCG value, histology, pretreatment risk score and decision to administer chemotherapy. CONCLUSION: Women with gestational trophoblastic disease who were registered late were significantly more likely to have normal levels of hCG and were less likely to need chemotherapy. A less intensive follow up may be justified in women with gestational trophoblastic disease who are registered with a normal hCG level. PMID- 15663135 TI - Laparoscopic surgery following abdominal wall reconstruction: description of a novel method for safe entry. AB - We report a technique for safe laparoscopic access for gynaecological surgery in women who have had their abdominal wall reconstructed by plastic surgery. The abdominal wall incisions for placement of the trocars are made in the abdominoplasty scars and are cosmetically acceptable. In a limited series there appears to be no excess morbidity. PMID- 15663136 TI - Research jobs: how good is the training? AB - OBJECTIVES: To determine if a period in a research job improves confidence in research skills and to assess the quality of research training. DESIGN AND SETTING: A questionnaire was designed and piloted to assess the content, structure and process of research training. POPULATION/SAMPLE: All individuals who had passed MRCOG in the five years prior to 1999 and were residents of England or Wales. METHODS: Confidence scores were compared between those with and those without research experience. The availability and value of differing strategies for research training were compared. MAIN OUTCOME MEASURES: Confidence in research skills and attitudes to training. RESULTS: Of the 532 usable questionnaires returned, 226 respondents had done or were doing research and these individuals had higher confidence on a variety of research skills than those with no experience of a formal research job. Confidence was patchy with less than 50% feeling confident at assessing bias in a case-controlled study, understanding the statistics used in a paper or assessing the power of a study. Of those who had done research, 50% or less felt their training had been good or excellent in any area. Self-directed learning and discussion with peers were felt to be the most useful strategies for research training. Short intensive courses were not available for many respondents, but were felt to be useful. CONCLUSION: The high levels of dissatisfaction with the training in key skills required for research suggests that there is a need for a system for recognition of research posts. Reform of training in the research job should build on the current strength of encouraging self-directed learning. PMID- 15663137 TI - Unadjusted and customised weight centiles in the identification of growth restriction among stillborn infants. AB - The study aims to compare the utility of unadjusted with customised weight standards in the identification of intrauterine growth restriction (IUGR) among unexplained stillborn infants undergoing postmortem examination. Unadjusted and customised birthweight centiles were determined for 51 unexplained stillborn infants undergoing perinatal autopsy. Unadjusted centiles were calculated from an ultrasonically derived fetal weight standard. Customised centiles were calculated from an online calculator which adjusts the standard to account for important physiological variables. IUGR was defined as moderate or severe according to brain/liver ratios of > 3 and > 5, respectively. The relationship between the weight centiles and abnormal brain/liver weight ratios was explored. Neither unadjusted nor customised standards identify stillborn infants with brain/liver ratios > 3:1. Both unadjusted and customised weight standards identify stillborn infants with brain/liver ratios > 5 equally well with high sensitivity (95%) but low specificities (63% and 66%, respectively). Customising weight standards to account for physiological variables does not identify growth restricted stillborn infants more usefully than an unadjusted fetal weight standard. PMID- 15663138 TI - Prevalence of Mycoplasma genitalium in early pregnancy and relationship between its presence and pregnancy outcome. AB - Mycoplasma genitalium is associated with cervicitis and pelvic inflammatory disease but little is known about its role in pregnancy. We investigated the prevalence of M. genitalium by polymerase chain reaction assay on urine specimens from 1216 pregnant women (mean age 31years) presenting before 10 weeks of gestation in 32 general practices. The prevalence of M. genitalium was 0.7% (6/915, 95% CI 0.1-1.2). It was more common in women aged < 20 years, women of Afro-Caribbean or black African ethnic origin, women in social classes 3-5 and single women. Only one woman with M. genitalium infection miscarried, and none of those followed up to term had a preterm birth, although the numbers were small. The low prevalence of M. genitalium infection suggests it is unlikely to be an important risk factor in adverse pregnancy outcome in healthy women in the community. PMID- 15663139 TI - Ex vivo analysis of HIV-1 co-receptors at the endocervical mucosa of women using oral contraceptives. AB - Combined oral contraceptives may alter the microenvironment of the female genital tract and, thus, influence susceptibility of endocervical cells to HIV-1 transmission. The mechanism for this effect is unknown but might involve combined oral contraceptive up-regulation of chemokine receptors on CD4+ endocervical cells. We measured chemokine co-receptor (CCR5 and CXCR4) expression on cervical intraepithelial CD4+ T lymphocytes, macrophages and dendritic cells using flow cytometry in 32 healthy women, 16 of whom were combined oral contraceptive users and 16 non-users. All women tested negative for sexually transmitted infections. Combined oral contraceptive users showed a higher proportion of CCR5+ CD4+ T lymphocytes compared with combined oral contraceptive non-users (P < 0.05). However, expression of both co-receptors on cervical intraepithelial macrophages and dendritic cells was no different between the two groups. Up-regulation of CCR5 on cervical intraepithelial CD4+ T lymphocytes offers a potential explanation by which women receiving combined oral contraceptives may be at increased risk of HIV transmission. PMID- 15663140 TI - Infant cortisol response after prolonged antenatal prednisolone treatment. AB - Prednisolone is widely used to treat medical conditions in pregnancy, despite the lack of long-term safety studies on infants. Animal studies have shown that antenatal glucocorticoid treatment can cause in utero growth restriction and up regulation of the offsprings' hypathalamic-pituitary-adrenal axis. We recruited women treated antenatally with prednisolone, and followed 12 of the infants up to four months, using routine infant vaccinations as a stressor. Birthweights were similar to controls (n = 289, uncomplicated, singleton term pregnancies), as were infants' baseline and stress-induced cortisol levels. Mothers rated their infants as less difficult and more adaptable than controls. This study provides initial reassurance about the safety of prednisolone in pregnancy. PMID- 15663141 TI - Treatment of external genital warts with 5% imiquimod cream during pregnancy: a case report. PMID- 15663142 TI - Fetal survival after conservative management of cord prolapse for three weeks. PMID- 15663143 TI - Ventricular outflow obstruction, valve aplasia, bradyarrhythmia, pulmonary hypoplasia and non-immune fetal hydrops because of a large rhabdomyoma in a case of unknown tuberous sclerosis: a prenatal diagnosed cardiac rhabdomyoma with multiple symptoms. PMID- 15663144 TI - Pregnancy following monofollicular ovulation induction with recombinant FSH, recombinant LH and timed coitus in an amenorrheic woman with long-standing hypogonadotrophic hypogonadism. PMID- 15663145 TI - Transection of a colonic diverticulum during enterocele repair. PMID- 15663146 TI - Cornual pregnancy. PMID- 15663147 TI - Routine ultrasound for pregnancy termination requests increases women's choice and reduces inappropriate treatments. PMID- 15663148 TI - Epidemiology of incontinence and prolapse. PMID- 15663149 TI - Pathophysiology of urinary incontinence and pelvic organ prolapse. PMID- 15663150 TI - Oestrogens and the lower urinary tract. PMID- 15663151 TI - Urinary incontinence--older people: where are we now? PMID- 15663152 TI - Dispelling the myth--does hysterectomy cause pelvic organ dysfunction? PMID- 15663153 TI - Role of imaging in urogynaecology. PMID- 15663154 TI - Measuring quality of life in urogynaecology. PMID- 15663155 TI - The role of pelvic floor muscle training in urinary incontinence. PMID- 15663156 TI - Mid-urethral tapes: which? Review of available commercial mid-urethral tapes for the correction of stress incontinence. PMID- 15663157 TI - Is there a role for colposuspension? PMID- 15663158 TI - The treatment of stress incontinence--is there a role for laparoscopy? AB - The laparoscopic colposuspension procedure has been performed by a relatively few urogynaecologists over the last 12 or 13 years. It is a procedure that takes time and skill to learn and is recognized as a level 3 minimal access procedure. Comparative studies between it and other surgical procedures are relatively few but would appear to suggest that it is less successful than open colposuspension or TVT in terms of curing USI. Adding to this the fact that TVT would appear to have, like the laparoscopic colposuspension, the advantages attached to a minimal access procedure, it is hard to see the procedure having a significant place in the treatment of USI in the longer term. It may well be that a few skilled practitioners will continue with the procedure, especially if performing other laparoscopic surgery at the same time. PMID- 15663159 TI - Detrusor overactivity--an update. PMID- 15663160 TI - Interstitial cells of the bladder: the missing link? PMID- 15663161 TI - Devices for continence and prolapse. PMID- 15663162 TI - Management of prolapse of the anterior compartment. PMID- 15663163 TI - Management of the posterior compartment. PMID- 15663164 TI - The vaginal vault. PMID- 15663165 TI - Pathophysiology and nonsurgical treatment of anal incontinence. PMID- 15663166 TI - How will the scaling up of zinc for treatment of childhood diarrhoea affect caretakers' practices? PMID- 15663167 TI - Serum lipid profiles and homocysteine levels in adults with stroke or myocardial infarction in the town of Gombe in northern Nigeria. AB - While the incidence of infectious diseases has been on the decline in developing countries, the toll of cardiovascular diseases, including stroke and myocardial infarction, has been increasing. The impression of physicians in certain regions of the western Sahel, including the state of Gombe in northeastern Nigeria, is that macrovascular disease in the indigenous population is on the rise. This study was, therefore, undertaken to compare well-established risk factors for cardiovascular disease in a group of 53 men (n=34) and women (n=19) in the town of Gombe who had suffered a stroke or heart attack with the corresponding parameters in 48 age- and gender-matched healthy adults living in the same environment. The parameters of cardiovascular diseases considered were: overweight and obesity, blood pressure, lipid profiles, and homocysteine. While the male and female patients who had suffered stroke (n=48) or heart attack (n=5) were borderline hypertensive, their mean body mass index values were not different from the corresponding values of their control groups. Although the serum homocysteine levels of the patients and controls were not significantly different, 85% of the stroke patients had serum homocysteine levels greater than 10 microM. These high homocysteine levels could not be accounted for by sub optimal folate or vitamin B 12 status. The serum levels of HDL-cholesterol and triglyceride were not significantly different between the male and female patients and their respective controls. However, the males, but not the females, with macrovascular disease had significantly higher levels of total cholesterol (161 vs 137 mg/dL, p=0.04) and LDL-cholesterol (91 vs 70 mg/dL, p=0.02). In addition, both female and male stroke/myocardial infarction patients exhibited an elevated LDL-cholesterol/HDL-cholesterol ratio. These results indicate that blood pressure and the LDL-cholesterol/HDL-cholesterol ratio are associated with stroke and myocardial infarction in adults in northern Nigeria, thereby creating potential opportunities for possible public-health interventions. PMID- 15663168 TI - Gestational nightblindness among women attending a public maternity hospital in Rio de Janeiro, Brazil. AB - This study evaluated the prevalence of gestational nightblindness among postpartum women seen at the University Maternal Hospital of the Federal University in Rio de Janeiro, Brazil and the association of this symptom with a biochemical indicator (serum retinol levels) and sociodemographic, anthropometric and antenatal care variables. In total, 262 postpartum women, who did not receive vitamin A supplementation during pregnancy, were interviewed. Gestational nightblindness was diagnosed through the standardized interview as proposed by WHO. Serum retinol levels were evaluated by spectrophotometry. Gestational nightblindness relating to low levels of serum retinol (<1.05 micromol/L, p = 0.000) was diagnosed in 17.9% of subjects interviewed. Less than five antenatal care appointments (odds ratio [OR] = 2.179; confidence interval [CI] 95% = 1.078 4.402) and a history of one or more miscarriage(s) (OR = 2.306; CI 95% = 1.185 - 4.491) were predictors for gestational nightblindness. These findings justify the need for nutritional counselling, aimed at improving the vitamin A nutritional status, especially among pregnant women with a history of previous miscarriages and poor antenatal care. PMID- 15663169 TI - Risk factors for cardiovascular disease and diet of urban and rural dwellers in northern Nigeria. AB - Over the last 30 years, cardiovascular diseases (CVDs), including stroke and myocardial infarction, have increased in developing countries. Serum lipids and diet of the Fulani, a rural Nigerian population, were previously studied. Despite their consumption of a diet rich in saturated fat, the overall blood lipid profiles of Fulani men and women are generally favourable. However, Fulani males in the same study had mean serum levels of homocysteine, an emerging risk factor for CVD, that exceeded the upper limit of the homocysteine reference range. The authors were interested in knowing if these findings in the Fulani nomads were representative of the biochemical parameters of CVD risk in other ethnic groups in the same region of Nigeria. To address this question, the nutrient content of diets of 55 men, aged 20-75 years, and 77 women, aged 20-70 years, who were inhabitants of a large urban centre in northern Nigeria, was assessed, and their serum levels of total cholesterol, LDL-cholesterol, HDL-cholesterol, triglycerides, and homocysteine were determined. These data were compared with those of the same rural Fulani population studied previously. Urban subjects consumed more calories than rural subjects (men: 2061 vs 1691 kcal; women: 1833 vs 1505 kcal) and had a significantly higher mean body mass index (BMI) and percentage of body fat than rural subjects. Both urban males and females had carbohydrate intakes that were greater than those of Fulani pastoralists (men: 56% vs 33% total calories; women: 51% vs 38% total calories), but had a significantly lower dietary intake of total fat and saturated fat (men: 36% vs 51% of total calories; women: 40% vs 51% of total calories). With the exception of HDL-cholesterol levels, which were significantly lower in the rural population, the blood lipid profiles of rural subjects were more favourable compared to those of urban subjects. Both urban and rural males had homocysteine levels above the upper limit of the reference range for healthy adults (urban males--12.7 micromol/L; rural males-15.2 micromol/L). The dietary intakes of folate and vitamin B12 were lower for rural Fulani subjects, and this was reflected in their significantly lower serum concentrations of these two vitamins. Results of this study suggest that, although the lipid profiles of urban and rural men and women in northern Nigeria indicate a relatively low risk for CVD, their elevated serum homocysteine levels are a cause for concern. The high homocysteine levels among rural men and women could be explained in part at least by their marginal status with respect to folate and vitamin B12. PMID- 15663170 TI - Prevalence of enterotoxigenic Escherichia coli-associated diarrhoea and carrier state in the developing world. AB - This study assesses the importance of enterotoxigenic Escherichia coli (ETEC) as a diarrhoeal agent in developing countries. Odds ratios were calculated for incurring ETEC-associated diarrhoea based on data reported between 1970 and 1999. Carriage of ETEC was associated with diarrhoea in children aged less than five years, except for hospitalized infants aged 0-11 month(s) and children aged 1-4 year(s) at outpatient clinics. Two hundred and eighty million episodes of diarrhoea due to ETEC were seen yearly among those aged less than five years, and close to 50 million children of this age group were asymptomatic carriers of ETEC. Every 7th diarrhoeal episode in children aged less than one year and close to 25% of diarrhoeal cases in children aged 1-4 year(s) were due to ETEC. A child born in a developing country is likely to experience 0.5 diarrhoeal episodes per year caused by ETEC until the age of five years, after which the yearly incidence drops to 0.1. To conclude, ETEC remains an important diarrhoeal pathogen among children in the developing world. PMID- 15663171 TI - Triaging patients with post-abortion complications: a prospective study in Nepal. AB - The first manual vacuum aspiration (MVA) services unit in Nepal was established in 1995 at the country's largest national maternity hospital in Kathmandu. This research sought to assess and evaluate the safety, acceptability, and effectiveness of MVA services. This prospective study was conducted during 12 months in 1998, and follow-up was made at six weeks. Two groups of patients were compared: 529 patients treated in the MVA unit and 236 patients who were clinically eligible for treatment in the MVA unit but were treated instead in the main operation theatre (OT) owing to the unavailability of services in the MVA unit during the hours of their admission. The two groups differed with respect to some of their background characteristics but were similar in their clinical characteristics. The MVA group received contraceptive counselling and services and had significantly shorter stays in hospital. However, the direct cost incurred by the patients, regardless of the type of facility they used, was about the same. Follow-up at six weeks revealed that the MVA patients had significantly fewer complaints and were generally more satisfied with the services they had received than their counterparts. Slightly more than half of the women in the MVA group were using contraception at the time of follow-up compared to no women in the OT group. It is concluded that the MVA unit provided safe, effective, and efficient services to about 50% of all the patients admitted to the hospital with post-abortion complications. An additional 25% of the post-abortion patients could be served if the unit were kept open 24 hours a day, saving resources and time for patients and hospital staff. As a parallel development, both MVA and main OT services would need to be more effectively integrated with outside antenatal and family-planning clinics to address the reproductive health needs of women, thereby reducing the number of patients requiring post-abortion care. PMID- 15663172 TI - Environmental temperature, cholera, and acute diarrhoea in adults in Lima, Peru. AB - The effects of environmental temperature, presence and severity of El Nino, presence of cholera in the community, and interactions among these variables on the number of adult diarrhoeal patients attending the Hospital Nacional Cayetano Heredia in Lima, Peru, during 1991-1998, were evaluated. During 1991-1996, an increased number of visits to the hospital due to acute diarrhoea in the warmer months was observed. This periodic pattern was altered in 1997, when rising of the environmental temperature was observed in Lima secondarily associated with a strong El Nino event. A multivariate model was built in which environmental temperature and interaction between environmental temperature and presence of cholera predicted the number of adult patients with acute diarrhoea attending the Hospital Nacional Cayetano Heredia. Monitoring of environmental temperature and presence of cholera may be used as a warning system to predict epidemics of diarrhoea in adults, which may have a tremendous impact on healthcare strategies and management of health services in general. PMID- 15663173 TI - Cost of delivering child immunization services in urban Bangladesh: a study based on facility-level surveys. AB - This facility-based study estimated the costs of providing child immunization services in Dhaka, Bangladesh, from the perspective of healthcare providers. About a quarter of all immunization (EPI) delivery sites in Dhaka city were surveyed during 1999. The EPI services in urban Dhaka are delivered through a partnership of the Government of Bangladesh (GoB) and non-governmental organizations (NGOs). About 77% of the EPI delivery sites in Dhaka were under the management of NGOs, and 62% of all vaccinations were provided through these sites. The outreach facilities (both GoB and NGO) provided immunization services at a much lower cost than the permanent static facilities. The average cost per measles-vaccinated child (MVC), an indirect measure of number of children fully immunized (FIC-the number of children immunized by first year of life), was 11.61 U.S. dollars. If all the immunization doses delivered by the facilities were administered to children who were supposed to be immunized (FVC), the cost per child would have been 6.91 U.S. dollars. The wide gap between the cost per MVC and the cost per FVC implies that the cost of immunizing children can be reduced significantly through better targeting of children. The incremental cost of adding new services or interventions with current EPI was quite low, not significantly higher than the actual cost of new vaccines or drugs to be added. NGOs in Dhaka mobilized about 15,000 U.S. dollars from the local community to support the immunization activities. Involving local community with EPI activities not only will improve the sustainability of the programme but will also increase the immunization coverage. PMID- 15663174 TI - Knowledge of men and women about reproductive tract infections and AIDS in a rural area of north India: impact of a community-based intervention. AB - This study of a community-level health-education intervention on reproductive tract infections/sexually transmitted diseases (RTIs/STDs) was conducted in three villages of Haryana in north India. The study was aimed at increasing awareness among men and women of reproductive age about the prevention and treatment of RTIs, modes of HIV/AIDS transmission, and methods of prevention. Health education was imparted through one-to-one interactions with men and women during home visits, at village-based clinics and health camps, and through health-education talks with men and women. Cumulative effects of the intervention were examined at the end of the survey by comparing the change in knowledge from the baseline. Records of clinic attendance were examined to assess the probable impact of the intervention on clinic attendance. Baseline and follow-up evaluations revealed that there was an improvement in the median total knowledge score of women from 0 to 6, whereas it remained at 5 for men both at baseline and follow-up. Knowledge about RTI/STIs increased among both men and women from the median score of 0 to 2 and from 0 to 3 respectively. The median knowledge score on HIV/AIDS declined among men from 4 to 2 but increased from 0 to 3 among women. Clinic attendance for RTI/STI cases, referred to the Naraingarh hospital, showed an eight-fold rise from an average of four cases per month in 1998-1999 to an average of 35 cases per month in 1999-2000. The findings of the study suggest that health-education strategy through home visits, RTI case management and counselling, and organizing a weekly clinic and occasional camps and health-education talks can increase the level of awareness about RTIs/STIs among both men and women and improve clinic attendance. PMID- 15663175 TI - Maternal deaths and their causes in Ankara, Turkey, 1982-2001. AB - This study was carried out to determine the incidence and causes of maternal deaths about a 20-year period at the Zekai Tahir Burak Women's Health Education and Research Hospital (ZTBWHERH), Ankara, Turkey. All maternal deaths from January 1982 to July 2001 were reviewed and classified retrospectively. Using a computer-generated list, 348 patients admitted to the Labour Department of ZTBWHERH during 1982-2001 were selected as controls. Medical records were reviewed for demographic data, history of antenatal care, route of delivery, referral history, and perinatal mortality. Cases and controls were compared, and standard tests were used for calculating odds ratio (OR) and 95% confidence interval (CI) for the association of demographic and delivery characteristics. During this period, there were 174 maternal deaths and 430,559 livebirths, giving a maternal mortality ratio of 40.4/100,000 livebirths. The mortality rate declined from 85.1/100,000 in 1982 to 11.6/100,000 in 2001. One hundred thirty (74.7%) deaths were due to direct obstetric causes and 24 (13.7%) were abortion related, while 20 (11.4%) were due to indirect obstetric causes. The most common cause of direct obstetric deaths was pre-eclampsia/eclampsia, followed by obstetric haemorrhage and embolism. Abortion-related sepsis and haemorrhage, anesthesia-related deaths, obstetric sepsis, acute fatty liver of pregnancy, and ectopic pregnancy accounted for other causes of deaths. Cardiovascular disease was the leading indirect cause of death. Referral, lack of antenatal care, and foetal death at admittance were associated with 8-, 3-, and 6-fold increased risk of maternal mortality respectively (OR 8.89, 95% CI 5.7-13.8; OR 3.74, 95% CI 2.5 5.5; OR 6.38, 95% CI 3.1-13.1). Although maternal mortality ratios have declined at the hospital, especially in the past five years, the rate is still high, and further improvements are needed. The problem of maternal mortality remains multifactorial. Short-term objectives should be focused on improving both medical and administrative practices. Improving the status of women will necessarily remain a long-term objective. PMID- 15663176 TI - An evaluation of post-campaign knowledge and practices of exclusive breastfeeding in Uganda. AB - Despite the benefits of exclusive breastfeeding for the health of mothers and children, its practice has markedly declined throughout the developing world. Mass media-communication programmes could play an important role in reversing this trend. This study evaluated the extent to which exposure to behaviour change communication (BCC) messages in the media determined recent improvements in exclusive breastfeeding knowledge and practices in areas targeted by the Delivery of Improved Services for Health (DISH) Project of Uganda. Data were drawn from the 1999 DISH Evaluation Survey. The survey collected information from representative samples of women and men of reproductive age. Multiple logistic regressions were used for assessing the independent influences of BCC exposure on breastfeeding knowledge and practices, controlling for several confounding factors. The results indicated that the exposure to BCC messages was strongly associated with women's knowledge of six months as the ideal duration for exclusive breastfeeding. Positive influences on knowledge of men were also found. Media effects on women's current practice of exclusively breastfeeding their infants up to six months were less conclusive, possibly because of the short interval between the launch of the BCC campaign and survey implementation. While there was some evidence of bias of self-reported exposure, results of exploratory analysis of the indirect effects of communication campaigns suggest that impacts may be compounded as overall awareness is increased at the community level eventually leading to improved knowledge among individuals. PMID- 15663177 TI - Zinc therapy for diarrhoea increased the use of oral rehydration therapy and reduced the use of antibiotics in Bangladeshi children. AB - Excessive use of antibiotics for diarrhoea is a major contributing factor towards increasing rates of antimicrobial resistance in developing countries. Zinc therapy for diarrhoea has been shown to be beneficial in controlled efficacy trials, and it is of interest to determine if availability of zinc syrup for treatment of diarrhoea would satisfy the demand for a 'medicine' for diarrhoea, thus reducing the use of antibiotics, without competing with the use of oral rehydration therapy (ORT). This community-based controlled trial was conducted from November 1998 to October 2000, and all children aged 3-59 months in the study area were included. In this trial, the availability of zinc supplements, along with ORT and appropriate education programmes, was associated with significantly higher use of ORT and lower use of antibiotics. PMID- 15663178 TI - Influences of different therapy protocols for chronic hepatitis B infection in Turkish children on nutritional anthropometric data. PMID- 15663179 TI - Shigellosis: report of a workshop. PMID- 15663180 TI - The mechanism of biogenesis and potential function of the two alternatively spliced mRNAs encoded by the murine Msx3 gene. AB - The homeodomain-containing Msx3 gene, the newest member of the Msx family, encodes two mRNAs, with an unknown relationship to each other. To elucidate how Msx3 gene generates the two transcripts, we cloned their corresponding cDNAs from an E10.5 mouse embryo cDNA library. The alignment of sequences of the two Msx3 specific cDNAs with the corresponding regions of the genomic DNA revealed that read-through of the sequences preceding a cryptic splice donor site in the first intron of Max3 gene generated the longer transcript. The longer Msx3 mRNA (Msx3 l) contains 66 nucleotides spliced in frame that would encode a protein with 22 additional amino acids. These extra 22 amino acids are inserted between the residues 72 and 73, exactly 14 amino acids upstream of the homeodomain of the smaller Msx3 protein. In situ hybridization and competitive RT-PCR experiments revealed that both Msx3-s and Msx3-l mRNAs elicited similar spatio-temporal patterns of expression in the developing embryo, with maximal expression of both mRNAs occurring in the embryos between 8.5 and 12.5 days post-coitus. We found that while Msx3-s down-regulated Msx1 promoter in transfected C2C12 cells, co expression of Msx3-l alone did not affect the activity of the Msx1 promoter. In contrast, a concomitant expression of Msx3-s and Msx3-l in the transfected C2C12 cells neutralized the repressive effect of Msx3-s on the Msx1 promoter. In transient expression assays, the repressive action of Msx3-s on Msx1 promoter could also be reversed by co-expression of exogenous Sp1. Our data indicate that the potential interactions among the protein products encoded by the alternately spliced Msx3 mRNAs and the putative constituents of transcriptional co-activators and co-repressors may have functional consequences in vivo. PMID- 15663181 TI - Isolation and characterization of cytosolic alanine aminotransferase isoforms from starved rat liver. AB - Alanine is the most effective precursor for gluconeogenesis among amino acids and the initial reaction is catalyzed by alanine aminotransferases (AlaATs). It is a less extensively studied enzyme under starvation and known to that the enzyme activity increases in liver under starvation. The present study describes the purification and characterization of two isoforms of alanine aminotransferases from starved male rat liver under starvation. The molecular mass of isoforms was found to be 17.7 and 112.2 kDa with isoelectric points of 4.2 and 5.3 respectively for AlaAT I and AlaAT II. Both the enzymes showed narrow substrate specificity for L-alanine with different Km for alanine and 2-oxoglutarate. Both the enzymes were glycoprotein in nature. Inhibition, modification and spectroscopic studies showed that both PLP and free-SH groups are directly involved in the enzymatic catalysis. PLP activated both the enzymes with a Km 0.057 mM and 0.2 mM for AlaAT I and II respectively. PMID- 15663182 TI - The age-related increase in N-methyl-2-pyridone-5-carboxamide (NAD catabolite) in human plasma. AB - N-Methyl-2-pyridone-5-carboxamide (2PY) is a major catabolite of NAD, which is excreted in the urine. Assay of plasma concentration and urine exrection of 2PY is of interest for monitoring niacin nutritional state and NAD catabolism. Aging has major impact on number of metabolic processes, but its effect on plasma and tissue 2PY concentration has not been studied so far. The present study was undertaken to investigate the effect of aging on human plasma 2PY concentration. Blood samples were collected from 78 (56 females and 22 males) healthy subjects at different age ranging from 5 to 90 years. Plasma ultrafiltrates were analyzed with liquid chromatography/mass spectrometry (LC/MS) for 2PY concentration. 2PY concentration was also evaluated in plasma and liver of young (2 month) and old (24 month) rats. The mean plasma 2PY concentration in young (5-16 years old) healthy subjects was 0.39 +/- 0.22 micromol/l while in old healthy subjects (50 90 years old) it was approximately 2.6-fold higher. No gender differences was found in plasma 2PY concentration. Univariate regression analysis displayed positive correlation between age and human plasma 2PY concentration (r = 0.57, p < 0.01) and between age and 2PY/creatinine ratio (r = 0.4, p < 0.01). Multivariate regression analysis indicates that both age and plasma creatinine concentrations were independently correlated with the increase in plasma 2PY concentration. Daily 2PY excretion with urine corrected for weight was approximately 30% lower in old subjects than in young humans. 2PY concentrations in liver and plasma of 2-month-old rats were lower than in 24-month-old rats. In conclusion, our results demonstrated that human plasma 2PY concentration increases with age. This could be a consequence of both decreased renal excretion and increased production in the the liver or other organs. Our results indicate that age should be taken into consideration in the interpretation of plasma 2PY concentration. PMID- 15663183 TI - Hyperglycemia does not alter state 3 respiration in cardiac mitochondria from type-I diabetic rats. AB - Cardiovascular complications are the primary cause of death for diabetic patients. Clinically, the development of dysfunctional cardiomyopathy is one of the main complications of diabetes. Experimental evidence indicates that the mitochondrion is one of the main sites implicated in the development of cardiac dysfunction. Yet, the precise cause and mechanisms involved in the process are largely debated. We report here that heart mitochondria from streptozotocin induced diabetic Sprague-Dawley rats present a gradual reduction in state 3 oxygen consumption that reaches 35% by the fourth week following diabetes onset. Rats presenting a level of hyperglycemia similar to diabetic animals, but not showing the marked weight loss or appearance of urinary ketones typical of the later group present no decline in state 3 mitochondrial oxygen consumption, the values being indistinguishable from those of mitochondria from control animals. Mitochondria from hyperglycemic non-ketotic rats, however, show a 15-20% increase in state 4 respiration, but only when glutamate is used as energetic substrate. Mitochondria from diabetic rats, instead, show a 40-50% increase in state 4 respiration with glutamate and 20-25% with succinate as energetic substrate. Interestingly, hyperglycemic non-ketotic animals present a level of serum insulin intermediate between those of controls and diabetic animals. These functional modifications are unrelated to the time elapsed since the onset of diabetes, as they are observed at 2, 4, 6 as well as 8 and 12 weeks after diabetes onset. Taken together, these data argue against hyperglycemia per se being a direct cause of the decline in state 3 oxygen consumption observed in cardiac mitochondria of type-I diabetic rats. Rather, they point to insulin level and subsequent metabolic alterations as a possible cause for the insurgence of mitochondrial dysfunction. PMID- 15663184 TI - Effect of orally administered L-tryptophan on serotonin, melatonin, and the innate immune response in the rat. AB - To assess the effects of external administration of L-tryptophan on the synthesis of serotonin and melatonin as well as on the immune function of Wistar rats, 300 mg of the amino acid were administered through an oral cannula either during daylight (08:00) or at night (20:00) for 5 days. Brain, plasma, and peritoneal macrophage samples were collected 4 h after the administration. The accumulation of 5-hydroxytryptophan (5-HTP) after decarboxylase inhibition was used to measure the rate of tryptophan hydroxylation in vivo. Circulating melatonin levels were determined by radioimmunoassay, and the phagocytic activity of macrophages was measured by counting, under oil-immersion phase-contrast microscopy, the number of particles ingested. The results showed a diurnal increase (p < 0.05) in the brain 5-HTP, serotonin (5-hydroxytryptamine, 5-HT), and 5-hydroxyindolacetic acid (5-HIAA) of the animals which had received tryptophan at 08:00 and were killed 4 h later. In the animals which received tryptophan during the dark period, the 5 HT declined but the 5-HT/5-HIAA ratio remained unchanged. There was also a significant increase (p < 0.05) in nocturnal circulating melatonin levels and in the innate immune response of the peritoneal macrophages in the animals which had received tryptophan at 20:00. The results indicated that the synthesis of serotonin and melatonin, as well as the innate immune response, can be modulated by oral ingestion of tryptophan. PMID- 15663185 TI - Time course studies on the functional evaluation of experimental chronic myocardial infarction in rats. AB - In vivo models of myocardial infarction induced by coronary artery ligation (CAL) in rats usually suffer from high early mortality and a low rate of induction. This study investigated the time course initiation of chronic myocardial infarction (CMI) in albino rats and the possibility of reducing early mortality rate due to myocardial infarction by modification of the surgical technique. CAL was carried out by passing the suture through the epicardial layer around the midway of the left anterior descending coronary artery including a small area of the myocardium to avoid mechanical damage to the heart geometry. In addition, the role of endothelin-1 (ET-1) in rat heart with congestive heart failure was critically assessed. Time course initiation experiments were designed by sacrificing the animals at different time intervals and by carrying out physiological, biochemical, histopathological, electron microscopical and immunohistochemical studies. Specific markers of myocardial injury, viz. cardiac troponin-T (cTnT), high sensitivity C-reactive protein, lactate dehydrogenase and fibrinogen were measured at different time points. Serum marker enzymes and activities of lysosomal hydrolases were found to be elevated on the eighth day post-ligation. Histopathological studies demonstrated focal areas showing fibrovascular tissue containing fibroblasts, collagenous ground substance and numerous small capillaries replacing cardiac muscle fibers. Transmission electron micrographs exhibited mitochondrial changes of well-developed irreversible cardiac injury, viz. swelling, disorganization of cristae, appearance of mitochondrial amorphous matrix densities, significant distortion of muscle fibers and distinct disruption of the intercalated discs. Immunoblotting studies confirmed the presence of alpha 2-macroglobulin which supported the inflammatory response. The severity of the CMI was inferred by the measurement of the level of ET-1 in plasma and left ventricle which was significantly higher in the CMI rats than in the sham-operated rats. Immunohistochemical studies at different time intervals showed that there was a significant immunoexpression of ET-1 on the eighth day post-ligation. This study conclusively showed that ligation of left anterior descending artery minimized mortality and ET-1 was expressed during CMI. PMID- 15663186 TI - Effect of some volatile oils on the affinity of intact and oxidized low-density lipoproteins for adrenal cell surface receptors. AB - Extensive research has shown that a high plasma concentration and oxidation of low-density lipoproteins (LDL) has an important role in atherogenesis. The affinity of LDL to its classic receptor is reduced due to oxidation. Instead, it is taken up by scavenger molecules in macrophages, as a result of which foam cells are formed that have a major role in increasing the subendothelial fat layers of the blood vessels. In the present study the antioxidant effect of eight volatile compounds in plant essences, namely, anethol, eugenol, limonen, linalool, p-cymol, pulegon, thymol, and geraniol, and their effect on the affinities of intact and oxidized (with Cu(+2)) LDL for LDL receptor in sheep adrenal tissue cells in the presence of labeled LDL with fluorescein isothiocynate (FITC) were investigated. The results obtained show that eugenol and thymol have the highest antioxidant effect, on the uptake of LDL (intact and oxidized) by the adrenal cells. The order of the compounds studied with regard to their antioxidant effect on intact and oxidized LDL is as follows: On intact LDL: eugenol > or = thymol > linalool > p-cymol > limonen > geraniol > anethol; on oxidized LDL: thymol > or = eugenol > geraniol > p-cymol > linalool > pulegon. Our findings also show that the compounds, particularly thymol and eugenol, have an antioxidant property and can change the affinity of the LDL particles for the LDL receptor probably due to their lipophylic property. Further research may prove that these compounds can be used clinically, especially in atherosclerotic and hypercholesterolemic cases. PMID- 15663187 TI - Effects of polyphenols from grape seeds on oxidative damage to cellular DNA. AB - Grape seed polyphenols have been reported to exhibit a broad spectrum of biological properties. In this study, eleven phenolic phytochemicals from grape seeds were purified by gel chromatography and high performance liquid chromatography (HPLC). The antioxidant activities of five representative compounds with different structure type were assessed by the free radical scavenging tests and the effects of the more potent phytochemicals on oxidative damage to DNA in mice spleen cells were investigated. Procyanidin B4, catechin, epicatechin and gallic acid reduced ferricyanide ion and scavenged the stable free radical, alpha, alpha-diphenyl-beta-picrylhydrazyl (DPPH) much more effectively than the known antioxidant vitamin ascorbic acid, while epicatechin lactone A, an oxidative derivative of epicatechin, did not reduce ferricyanide ion appreciably at concentrations used and was only about half as effective on free radical-scavenging as epicatechin. Mice spleen cells, when pre-incubated with relatively low concentration of procyanidin B4, catechin or gallic acid, were less susceptible to DNA damage induced by hydrogen peroxide (H2O2), as evaluated by the comet assay. In contrast, noticeable DNA damage was induced in mice spleen cells by incubating with higher concentration (150 microM) of catechin. Collectively, these data suggest that procyanidin B4, catechin, gallic acid were good antioxidants, at low concentration they could prevent oxidative damage to cellular DNA. But at higher concentration, these compounds may induce cellular DNA damage, taking catechin for example, which explained the irregularity of dose-effect relationship. PMID- 15663188 TI - Reactivity of typhoid patients sera with stress induced 55 kDa phenotype in Salmonella enterica serovar Typhi. AB - Salmonella faces a variety of stresses including acid and heat, in the natural environment whether in the gastrointestinal tract of mammalian host or in the external environment during transmission where survival and multiplication is a priority for the pathogen. In the present study, Salmonella enterica serovar Typhi was grown under acid (inorganic and organic) as well as heat stress and the outermembrane protein (OMP) profiles were compared. A 55 kDa OMP was found to be expressed with high intensity under the selected stress conditions in comparison to normal conditions. The protein expressed under acidic stress reacted with antibodies raised against heat shock protein indicating the similarity of atleast some of the epitopes. In vivo immunogenicity (reactivity with typhoid patient sera) revealed that the 55kDa protein under each stress condition was reactive with 83% of the typhoid sera. In the light of role of the stress induced proteins in pathogenesis of microbial infections and their immunogenic potential, these findings may be relevant for a better understanding of the host-microbe interactions and for future development of diagnostic and preventive strategies. PMID- 15663189 TI - The effects of insulin-like growth factor binding protein-3 (IGFBP-3) on T47D breast cancer cells enriched for IGFBP-3 binding sites. AB - To investigate insulin-like growth factor (IGF)-independent effects of IGF binding protein-3 (IGFBP-3), T47D cells were enriched for a population of cells that expressed binding sites for biotinylated-IGFBP-3 by panning on streptavidin coated plate. Proliferation of cell enriched for IGFBP-3 binding sites was significantly inhibited by IGFBP-3, whereas IGFBP-3 had no significant effect on the non-enriched cell population. Enriched and non-enriched cells were equally responsive to IGF-I, TGF-beta and EGF. Conditioned medium from enriched cells had less IGFBP-3 than that from non-enriched cells. Cross-linking of biotinylated IGFBP-3 to T47D cell membranes identified complexes with Mr of 32, 80 and 100 kDa. All of these complexes were more abundant in enriched cells compared with the non-enriched cell population. These data demonstrate that despite the anti proliferative effects of IGFBP-3 it is possible to selectively enriched for cell populations with more abundant IGFBP-3 binding sites. These enriched cells are more responsive to IGFBP-3 and secrete less of this binding protein than non enriched cells, supporting the concept that IGFBP-3 secretion by human breast cancer cells may function as an autocrine or paracrine modulator of cell proliferation. PMID- 15663190 TI - Identification of an atypical form of 30 kDa SOD--a possible virulence factor in clinical isolates of Shigella spp. AB - Reviews on the pathogenic mechanisms of Shigella species show a lacunae in the understanding of the bacterial antioxidant defense system and its regulations. This study was done to investigate the regulation of expression of antioxidant enzymes in clinical isolates of Shigella species, under various growth conditions. The in vitro expression of superoxide dismutase in the clinical isolates of Shigella spp., is modulated by both endogenous and exogenous factors. During aerobic and iron repleted growth conditions, the expression of the MnSOD and FeSOD enzymes were higher, and an atypical SOD was also expressed. However, under anaerobic growth conditions and in plasmid-cured strains, the antioxidant enzyme activities were decreased and the atypical SOD was not expressed. Absence of the atypical form of SOD may be due to the low oxygen environment. Plasmid encoded factors may also play a role in the expression of this SOD, which had a molecular weight of approximately 30 kDa. In the rat ileal loop ligation assay, mild lesions were observed only in the intestinal microvilli of rats injected with plasmid-cured strains of Shigella spp., suggesting that plasmid-encoded factors, including those that regulate the expression of the atypical SOD, are essential for the virulence of Shigella spp. PMID- 15663191 TI - Streptozotocin-induced diabetes influences the activity of ecto-nucleoside triphosphate diphosphohydrolase 1 of rat osseous plate membranes. AB - We report the kinetic characterization of an ecto-nucleosidetriphosphate diphosphohydrolase 1 from rat osseous plate membranes in streptozotocin-induced diabetic rats, which arises during ectopic mineralization twenty days after a subcutaneous implantation of demineralized bone matrix, Insulin deficiency decreased the ecto-nucleoside triphosphate diphosphohydrolase activity from 1293.1 +/- 39.8 (control rats) to 556.0 +/- 8.2 nmol Pi/(min mg). Two families of ATP hydrolyzing sites showed cooperative effects with specific activities of 256.2 +/- 7.7 nmol Pi/(min mg) and 299.8 +/- 8.9 nmol Pi/(min mg), and studies on the stimulation of the enzyme by magnesium and calcium ions showed that the decrease in enzyme activity results from changes in the affinity of the enzyme for these ions. To our knowledge this is the first study associating the effects of type I diabetes with an ecto-nucleoside triphosphate diphosphohydrolase activity from rat osseous plate membranes. PMID- 15663192 TI - Calcium-modulated ciliary membrane guanylate cyclase transduction machinery: constitution and operational principles. AB - Odorant transduction is a biochemical process by which the odorant signal generates the electric signal. The cilia of the olfactory neuroepithelium are the sites of this process. This study documents the detailed biochemical, structural and functional description of an odorant-responsive Ca2+ -modulated membrane guanylate cyclase transduction machinery in the cilia. Myristoylated (myr) neurocalcin delta is the Ca2+ -sensor component and the cyclase, ONE-GC, the transduction component of the machinery. Myr-neurocalcin delta senses increments in free Ca2+, binds to a defined domain of ONE-GC and stimulates the cyclase. The findings enable the formulation of an odorant transduction model in which three pivotal signaling components--Ca2+, myr-neurocalcin delta and ONE-GC--of the transduction machinery are locked. A glaring feature of the model is that its Ca2+ -dependent operational principle is opposite to the phototransduction model. PMID- 15663193 TI - Adenoviral SERCA1 overexpression triggers an apoptotic response in cultured neonatal but not in adult rat cardiomyocytes. AB - Although adenoviral SERCA gene transfer improves cardiac function in the failing heart, there is controversy regarding the cytotoxic effect of such overexpression. We sought to examine the effect of SERCA1 overexpression on neonatal (NRCMs) and adult rat cardiomyocytes (ARCMs). Cultured NRCMs and ARCMs were infected with adenoviral vector expressing EGFP (Ad.EGFP) or SERCA1 (Ad.SERCA1). Gel electrophoresis and microarray analysis were performed to examine DNA fragmentation and apoptosis-related genes, respectively. Northern and western blot were used to examine the expression level of SERCA. The optimal viral titer for Ad.EGFP was found to be 2-5 pfu/cell in NRCMs and 100 pfu/cell in ARCMs. Infection of NRCMs with 4 pfu/cell of Ad.SERCA1 resulted in loss of cells and DNA fragmentation, while no apoptosis was detected in Ad.EGFP-infected cells. Gene array analysis also revealed that Ad.SERCA1 induced expression of apoptosis related genes in NRCMs. However, neither loss of cell viability nor DNA fragmentation and induction of apoptosis-related genes was observed in ARCMs infected with 100 pfu/cell of Ad.SERCA1. Following the optimal infection, the SERCA1 expression level in NRCMs was 4-fold higher than that in ARCMs. Interestingly, the endogenous SERCA2 protein in NRCMs was completely replaced by exogenous SERCA1 protein. Furthermore, the activity of Ca2+ ATPase was increased by 4-fold in NRCMs but only by 1.5-fold in ARCMs. Our results indicate that adenoviral SERCA1 gene transfer has appreciably different effects on NRCMs and ARCMs. SERCA1 overexpression triggers an apoptotic response in NRCMs but not in ARCMs. PMID- 15663194 TI - Differential expression of Hyaluronic Acid Binding Protein 1 (HABP1)/P32/C1QBP during progression of epidermal carcinoma. AB - Our laboratory has characterized a novel cell surface glycoprotein, Hyaluronic Acid Binding Protein 1 (HABP1), interacting specifically with hyaluronan (HA) and regulating HA-mediated cellular event. The involvement of HA in different stages of carcinoma is well documented. In the present communication, the expression profile of HABP1 was investigated from initiation to progression of epidermal carcinoma in mice, induced by benzo[a]pyrene (B[a]P) exposure. During tumor initiation, HABP1 accumulated in inflammatory subsquamous tissue and with progression, the protein, was also seen to overexpress in papillomatic and acanthotic tissue. With the onset of metastasis, HABP1 overexpression was confined to metastatic islands, while it disappeared gradually from the surrounding mass. Such expression profiles in metastasized tissue were supported by decreased levels of HABP1, both at protein and transcript levels. These observations taken together suggest that the changes in HABP1 level coincide with specific stages of tumor progression, that lead to disruption of its interaction with HA, implying a role in the regulation of tumor metastasis. PMID- 15663195 TI - Peroxynitrite and fibrinolytic system: the effect of peroxynitrite on plasmin activity. AB - We have shown that peroxynitrite (ONOO-) inhibits streptokinase-induced conversion of plasminogen to plasmin in a concentration-dependent manner and reduces both amidolytic (IC5o approximately 280 microM at 10 microM concentration of enzyme) and proteolytic activity of plasmin. Spectrophotometric and immunoblot analysis of peroxynitrite-treated plasminogen demonstrates a concentration dependent increase in its nitrotyrosine residues that correlates with a decreased generation of active plasmin. Peroxynitrite (1 mM) causes the nitration of 2.9 tyrosines per plasminogen molecule. Glutathione, like deferoxamine, partially protects plasminogen from peroxynitrite-induced inactivation and reduces the extent of tyrosine nitration. These data suggest that nitration of plasminogen tyrosine residues by peroxynitrite might play an important role in the inhibition of plasmin catalytic activity. PMID- 15663196 TI - Inhibition of amyloid fibrillogenesis and toxicity by a peptide chaperone. AB - Aggregation of proteins in tissues is associated with several diseases, including Alzheimer's disease. It is characterized by the accumulation of amyloid beta peptide (Abeta) in the extracellular spaces of the brain cells, resulting in neuronal death and other pathological changes. alpha-Crystallin, a small heat shock protein in lens, and a peptide chaperone having the functional site sequence DFVIFLDVKHFSPEDLTVK of alphaA-crystallin may inhibit Abeta fibrillogenesis and toxicity. The peptide chaperone (mini-alphaA-crystallin), having an Abeta interacting domain and a complex solubilizing domain, was shown in previous studies to prevent aggregation of several proteins under denaturing conditions. In this in vitro study, using transmission electron microscopy and thioflavin T binding assay, we show that mini-alphaA-crystallin arrests the fibril formation of Abeta peptides. Mini-alphaA-crystallin also suppresses the toxic action of Abeta on rat pheochromocytoma (PC 12) cells. The wide chaperoning capability of the peptide and its ability to inhibit amyloid fibril formation and suppress toxicity suggest that mini-alphaA-crystallin may serve as a universal chaperone in controlling diseases of protein aggregation, including Alzheimer's disease. PMID- 15663197 TI - Regional differences in oxidative capacity of rat white adipose tissue are linked to the mitochondrial content of mature adipocytes. AB - Two metabolic pathways of the white adipocytes (i.e. de novo lipogenesis and lipolysis) require mitochondria functionality. In this report, the oxidative capacity of two white adipose tissues of rat and their respective isolated adipocytes were evaluated. Two major white fat pads, namely inguinal and epididymal tissues, were chosen as subcutaneous and visceral adipose tissues, respectively. The mitochondrial content of these tissues was estimated using cytological and biochemical analysis. Electron microscopy analysis showed higher mitochondrial density in epididymal than in inguinal adipocytes. The mitochondrial DNA content and mitochondrial enzymatic equipment were also higher in the former than in the latter tissue. A positive correlation between two mitochondrial enzymatic activities, namely cytochrome c oxidase and citrate synthase, and the mtDNA content of adipose tissue was reported. Moreover, NRF1 protein, which belongs to the transcriptional activator family and is thought to be involved in mitochondrial biogenesis regulation, was present in higher proportions in nuclei isolated from epididymal cells than in those from inguinal cells. Finally, greater abundance of mitochondria in epididymal tissue is in agreement with higher cytochrome c oxidase activity as well as increased respiration (i.e. basal and noradrenaline-stimulated) of adipocytes isolated from epididymal tissue as compared to adipocytes isolated from inguinal tissue. Therefore, white adipose tissue appears as a heterogeneous organ with marked variation in mitochondrial content depending on its anatomical location. PMID- 15663198 TI - Radioprotective mechanism of Podophyllum hexandrum during spermatogenesis. AB - RP-1, a herbal preparation of Podophyllum hexandrum has already been reported to provide protection against whole body lethal gamma irradiation (10 Gy). It has also been reported to render radioprotection to germ cells during spermatogenesis. Present study was undertaken to unravel the cellular and molecular mechanism of action of RP-1 on testicular system in strain 'A' mice. Various antioxidant parameters such as thiol content, glutathione peroxidase (GPx), glutathione reductase (GR), glutathione-S-transferase (GST) enzyme activity, lipid peroxidation (LPO) and total protein levels were investigated. Thiol content was seen to increase significantly (p < 0.05) in both RP-1 alone and RP-1 pretreated irradiated groups over the irradiated groups at 8, 16 and 24 h. Irradiation (10 Gy) significantly decreased GPx, GST and GR activity in comparison to untreated control but RP-1 treatment before irradiation significantly (p < 0.05) countered radiation-induced decrease in the activity of these enzymes. Radiation-induced LPO was also found to be reduced at all time intervals by RP-1 treatment before irradiation. As compared to irradiated group the protein content in testicular tissue was increased in RP-1 pretreated irradiated group at 4 and 16 h significantly (p < 0.05). Comets revealed by single-cell gel electrophoresis were significantly longer (p < 0.001) in irradiated mice than in unirradiated control. RP-1 treatment before irradiation, however, rendered significant increase (p < 0.05) in comet length over the corresponding control and irradiated group initially at 4 h but at later time points, this was reduced significantly (p < 0.01) as compared to the irradiated group. RP-1 treatment alone rendered shorter comets at 8, 16 and 24 h than irradiated groups (p < 0.001). This study implies that RP-1 offers radioprotection at biochemical and cytogenetic level by protecting antioxidant enzymes, reducing LPO and increasing thiol content. PMID- 15663200 TI - Effect of temperature and duration of hyperthermia on HSP72 induction in rat tissues. AB - The aim of the present study was to determine whether heat shock protein 72 (HSP72) is induced in a heated rat model at rectal temperatures below 42 degrees C. Rats were divided into a control group and six groups (n = 6) heated to different rectal temperatures: 39 degrees C for 1 h (39), 40.0 degrees C for either 15 min (40S) or 1 h (40L), 41.0 degrees C for either 15 min (41S) or 1 h (41L) and 42.0 degrees C for 15 min (42). Tissues were sampled 4 h after heating. Following 1 h at 40.0 degrees C, HSP72 was significantly elevated in heart (p < 0.005), but not in gut or liver tissue. In all three tissues, HSP72 was significantly elevated under the conditions 41L and 42 compared to control tissue (p < 0.005). Marked differences were found in the amount of HSP72 induced in different tissues in response to the same heat stress. Duration of heating was important in modulating HSP72 induction, with a significantly greater induction of HSP72 following 1 h compared to 15 min at 41 degrees C in all three tissues (p < 0.02). A correlation was found between thermal load and HSP72 content in liver, heart (both p < 0.01) and gut (p < 0.001) for the rats heated to 41 and 42 degrees C. These data show that HSP72 is induced at temperatures below 42 degrees C, with striking differences between tissues. PMID- 15663199 TI - Regulated expression of galectin-1 during T-cell activation involves Lck and Fyn kinases and signaling through MEK1/ERK, p38 MAP kinase and p70S6 kinase. AB - Recent evidence has implicated galectins and their carbohydrate ligands as novel regulators of T-cell homeostasis. Galectin-1 (Gal-1), a member of this family, inhibits clonal expansion, induces apoptosis of antigen-primed T lymphocytes and suppresses the development of T-cell-mediated autoimmune diseases in vivo. Because the beta-galactoside-binding protein is expressed in activated but not resting T cells, it has been hypothesized that Gal-1-induced apoptosis may constitute an autocrine suicide mechanism to eliminate activated T cells contributing to the termination of an effector immune response. We undertook this study to investigate the signals and intracellular pathways leading to Gal-1 expression during T-cell activation. When T cells were stimulated either with anti-CD3 or anti-CD28 monoclonal antibody plus PMA in the presence of accessory cells, a sustained up-regulation of Gal-1 was observed, reaching a plateau between days 3 and 5 following CD3 engagement or costimulation through CD28. Investigation of the signal transduction events involved in this process revealed a role for Lck and Fyn kinases, since the Src kinase inhibitor PP1 inhibited the up-regulated expression of Gal-1 following T-cell activation. Downstream signaling routes involve mitogen-activated protein kinase (MAPK) kinase (MEK)1/extracellular signal-regulated kinase (ERK) and p38 MAPK, as Gal-1 expression was prevented by U0126 and SB202190. In addition, expression of Gal-1 involves interleukin (IL)-2-dependent signaling routes triggered by p70S6 kinase, as it could be inhibited by rapamycin. This is the first demonstration of the intracellular pathways that control activation-induced expression of Gal-1, which may reveal potential targets for immune intervention to modulate expression of this beta-galactoside-binding protein in pathological disorders. PMID- 15663201 TI - Influence of plasma total antioxidant ability on lipid and protein oxidation products in plasma and erythrocyte ghost obtained from developing and adult rats pretreated with two vitamin K formulations. AB - Ferric reducing ability of plasma (FRAP), as an index of total antioxidant capacity of plasma was found to be enhanced significantly (p < 0.05) in suckling rats pretreated either with vitamin K1 (28, 56 or 84 mg/kg/3 days) or menadione (vitamin K3) at a dose of 15 mg/kg b.w./3 days. The effect of vitamin K1 on FRAP was dose-dependent and it was inversely related to the formation of lipid peroxidation products in plasma as judged by thiobarbituric acid reacting substances (TBARS). Lack of influence of the drugs on FRAP in adults was corroborated with elevation in the levels of plasma TBARS. Possible role of FRAP on the rate of lipid peroxidation and protein oxidation (protein carbonyls) on erythrocyte membrane was also investigated following isolation of erythrocyte ghost from control and treated rats. Vitamin K1 as well as menadione failed to change the levels of protein carbonyls in erythrocyte ghost obtained from both the age groups. Analysis of major erythrocyte membrane proteins, using SDS polyacrylamide gel electrophoresis (SDS-PAGE) substantiated these results. In spite of higher antioxidant capacity of plasma and erythrocytes obtained from young rats, the rate of lipid peroxidation in erythrocyte ghost preparation was also high in this age group (p < 0.05). These results suggest that the drug related induction in FRAP occurs only in immature animals as a part of protective mechanism against lipid peroxidation products generated in plasma. PMID- 15663202 TI - Natural vanadium-containing Mt. Fuji ground water improves hypo-activity of liver insulin receptor in Goto-Kakisaki rats. AB - After daily treatments with Mt. Fuji ground water containing natural vanadium (approximately 65 microg/l) at doses of 0.53 microg/kg/day for 12 weeks, blood glucose (BG), serum hemoglobin A1C (HbA1C) levels and insulin secretion from the pancreas of Goto-Kakisaki (GK) rats, a genetic model of Type 2 diabetes, were improved. In GK rat liver insulin receptors, the binding properties of [125I] insulin, and the activities of insulin receptor beta subunit and primary insulin like growth factor-1beta all recovered to normal levels of those found in Wistar rats. These results suggest that daily treatment with small concentrations of natural vanadium improves hyperglycemia by ameliorating liver insulin receptor activity. PMID- 15663203 TI - Remedial effect of DL-alpha-lipoic acid against adriamycin induced testicular lipid peroxidation. AB - Adriamycin (ADR), a cytotoxic antineoplastic drug is used in the treatment of various solid tumors. However, its efficacy continues to be challenged by significant toxicities including testicular toxicity. In the present study, the effect of lipoic acid, a "universal antioxidant" was investigated on ADR induced peroxidative damages in rat testis. Adult male albino rats of Wistar strain were administered ADR (1 mg/kg body weight, i.v.) once a week for 10 weeks. ADR injected rats showed a significant decline in the activities of enzymic antioxidants (catalase, superoxide dismutase, glutathione peroxidase, glutathione reductase and glutathione-S-transferase) and non-enzymic antioxidants (reduced glutathione, Vitamin A, Vitamin C and Vitamin E) with high malondialdehyde levels. The extent of testicular toxicity was evident from the decreased activities of testicular marker enzymes (sorbitol dehydrogenase and glucose-6 phosphate dehydrogenase). Treatment with lipoic acid (35 mg/kg body weight, i.p.) one day prior to ADR administration, maintained near normal activities of the enzymes and significantly reduced lipid peroxidation, thereby proving it to be an effective cytoprotectant. PMID- 15663204 TI - AJCP as a vehicle for improving community life: an historic-analytic review of the journal's contents. AB - The current article is an analysis of the research represented in American Journal of Community Psychology (AJCP) from 1993 to 1998, including a comparison to two previously published analytic reviews by Lounsbury et al. (J. W. Lounsbury, D. S. Leader, E. P. Meares, & M. P. Cook, 1980) and Speer et al. (P. Speer, A. Dey, P. Griggs, C. Gibson, B. Lubin, & J. Hughey, 1992), respectively. Observed trends are examined with references to major epistemological frameworks and methods used to define community psychology. Four guiding principles were examined to determine the representation of the epistemological frameworks of the articles published in the journal. The frameworks include social action, human diversity and cultural relativity, person-environmental fit, and methodological procedures. The results document the transition of community psychology from its early beginnings to an independent field conducting research consistent with the values articulated at the Swampscott Conference. This paper also comments on how well, after three decades of publication, the journal has served as a vehicle for improving community life. PMID- 15663205 TI - Social support mediates the effects of dual-focus mutual aid groups on abstinence from substance use. AB - BACKGROUND: Specialized 12-step based groups have emerged to address the needs of persons recovering from both substance abuse and psychiatric illness. OBJECTIVE: This study investigates the role of social support in mediating the association between mutual aid participation and subsequent substance use for dually diagnosed persons. METHOD: A cohort of Double Trouble in Recovery (DTR) members in New York City were studied prospectively over a two-year period. FINDINGS: Longer DTR participation during the first year of the study was associated with lower substance use in the second year; that effect was partially explained by the maintenance of high level of social support. CONCLUSION: These findings speak of the enduring influence of 12-step attendance on reducing substance use, and underline the importance of both 12-step attendance and supportive networks for dually diagnosed persons. PMID- 15663206 TI - The contributions of community, family, and school variables to student victimization. AB - An ecological perspective was used to predict school violence from a number of variables describing the students and their families based on nationally representative sample of 10,400 students in grades 7-11 in 162 schools across Israel. Self-administered anonymous questionnaires that included a scale for reporting victimization by serious and moderate physical violence, threats, and verbal-social victimization were filled out during class. Hierarchical linear modeling (HLM) examined the relationships between students' reports of victimization and student level variables (gender, age) and the school-level variables--cultural affiliation (Jewish vs. Arab), the socio-economic status (SES) of the school's neighborhood and students' families, school and class size, school level (junior high vs. high), and school climate. Variance between schools accounted for 9-15% of the variance in student victimization (major factors being school climate characteristics and percent of boys at the school). Boys reported higher victimization than girls for all forms of school violence. Students in junior high schools reported more victimization than high school students. Higher levels of victimization were reported in overcrowded classes, while school size was not associated with students' reports of victimization. Although there were almost no differences between Jewish and Arab schools in students' reports of victimization, the SES of the school's neighborhood and students did have a moderate effect. The discussion highlights the importance of improving school climate and the need to allocate more resources to schools in low SES contexts in order to protect students from school violence. PMID- 15663207 TI - The relationship between perceptions of neighborhood characteristics and substance use among urban African American adolescents. AB - Although the neighborhood microsystem is recognized as an important domain for adolescent development, relative to the family and peer contexts, neighborhood factors have been understudied in relation to adolescent substance abuse. In addition, recent research suggests that risk factors for adolescent substance use may differ for African Americans when compared to Caucasian youth. This study investigated the association between perceived neighborhood disorganization and later substance use, as well as possible mediators of that association, among a community sample of urban African American adolescents. Perceptions of neighborhood disorganization (i.e., violence/safety and drug activity) in grade 7 were associated with increased tobacco, alcohol, and marijuana use in grade 9. For females, this association was mediated by attitudes about drug use and perceptions of drug harmfulness. Findings highlight the importance of neighborhood contextual variables for African American substance use. Implications and directions for future research are presented. PMID- 15663208 TI - Crime, social capital, and community participation. AB - Social disorganization at the neighborhood and community levels has been consistently linked to various forms of criminal activity. However,a very much smaller body of literature addresses the effects of crime on community organizations. In some studies, crime appears to energize communities while in others, crime leads to withdrawal from community life. Using department of health crime victimization data and interviews with 2,985 low-income inner city residents living in 487 multi-family dwellings, a multi-level model examined the relationships among crime victimization, social organization, and participation in neighborhood organizations. Social organization at the individual and building levels was measured using recent formulations of social capital theory. Findings regarding crime suggested more signs of a chilling effect on participation than of an energizing effect, especially at the building level. Social capital at the building level was more strongly and consistently related to participation in community organizations than was crime. PMID- 15663209 TI - Delinquency and substance use among gang-involved youth: the moderating role of parenting practices. AB - This study uses longitudinal data from an ethnically diverse sample of 300 ninth grade students to examine the moderating effect of parenting practices upon the relationship between gang involvement and adolescent problem behavior. Results of hierarchical linear modeling indicate that gang involvement is a highly significant positive predictor of each of three categories of problem behavior (minor delinquency, major delinquency, and substance use). Three of the four parenting variables (behavioral control, psychological control, parent-adolescent conflict, and warmth) are found to moderate the relationship between gang involvement and problem behavior, with the most consistent effects found for behavioral control and warmth. These findings indicate that intervention efforts aimed at reducing the impact of gang involvement on adolescent development should consider factors that may decrease the deleterious behavioral outcomes associated with youth gangs. PMID- 15663210 TI - An empowerment evaluation model for sexual assault programs: empirical evidence of effectiveness. AB - Using an empowerment evaluation approach, the Sexual Assault and Rape Prevention (SARP) Evaluation Project brought together university researchers, public health evaluators, state funders, and program staff to provide evaluation training and consultation to all state-funded rape prevention and victim services programs in Michigan. In this paper, the specific activities of the SARP project are described, and process and outcome data are presented that address the effectiveness of this empowerment evaluation approach. Based on subjective reports from program staff and objective data obtained from state funders, results suggest that this evaluation model was successful in helping 90% of the prevention programs and 75% of the victim services programs successfully develop and launch program evaluations. One-year follow-up data indicate that 90% of the programs had sustained their evaluation efforts after the formal work of the SARP project had ended. PMID- 15663211 TI - Transmission of classical swine fever. A review. AB - Classical swine fever (CSF) is one of the most important viral pig diseases. Basic measures to control epidemics of CSF comprise eradication of infected herds and preventive emptying of herds at risk. Identification of these herds at risk is based on knowledge of the different transmission routes of the virus. Direct transmission of CSF is undoubtedly the most efficient way of CSF virus transmission. Data on indirect transmission are variable and often equivocal. Various indirect transmission routes like swill feeding, wild boar and artificial insemination are well described, and the importance is beyond discussion. Mechanical transmission via vehicles and persons are categorized as very important based upon epidemiological research, whereas in experiments they can only be reproduced under worst case conditions. The role of arthropods, birds, rodents, and other animals in the spread of CSF virus remains doubtful. Active transmission by these has never been demonstrated and also very sparse indications for mechanical transmission are available. Also the role of airborne transmission remains debated. However epidemiological as well as experimental data indicate that airborne spread over short distances is probable. PMID- 15663212 TI - Mast cells and canine mast cell tumours. A review. AB - This article reviews the literature on mast cells and tumours derived from mast cells in the dog. Mast cells play a central role in inflammatory and immune reactions. Mast cells, normal and neoplastic, contain and release important biologically active substances: heparin, histamine, eosinophilic chemotactic factor and proteolytic enzymes. Mast cell tumours occur in the dog, particularly in the boxer and related breeds, in the skin and less frequently in the intestines. Cytology usually provides an accurate diagnosis, but histological examination adds further information concerning the histologic grade and the completeness of surgical therapy. Cutaneous mast cell tumours should be regarded as potentially malignant and therefore be removed widely (3 cm. margin). Local recurrence, regional and distant metastases together with paraneoplastic disorders may cause the death of the pet. Histologic grading (2 or 3 grades) and clinical staging together with kinetic parameters and breed (boxers have relatively benign tumours) are important prognostic parameters. Based on prognostic criteria, surgical treatment should be completed with adjuvant radiotherapy, corticosteroids and eventually with combined chemotherapy. A novel, promising therapy is the application of the receptor kinase inhibitor. The study of the pathogenesis of mast cell tumours received new impetus by the finding of mutations, deletions and duplications, in exons 11 and 12 of the C-kit oncogene. Further study of physiological and oncological aspects of mast cells are favoured by the availability of mast cells isolated from spontaneous mast cell tumours and of cultured cell lines. PMID- 15663213 TI - Postweaning multisystemic wasting syndrome--PMWS. the first year with the disease in Sweden. AB - Postweaning multisystemic wasting syndrome (PMWS) is causally associated with porcine circovirus type 2 (PCV2) infection of pigs. PCV2 has been present in the Swedish pig population for at least ten years. Despite this, no signs of PMWS were observed in pigs of Sweden until 2003. Since then the disease has spread slowly in Sweden. The pig population of Sweden is geographically isolated, the density of pigs and the pathogen load in the country is low and the trade of animals is organised in a restricted way. To date, there has been little information placed in the peer-reviewed literature on the study of the spread of PMWS in a country during the early phase of the disease outbreak, mainly because diagnosis of PMWS has only occurred after the disease has been recognised on a number of farms. This manuscript documents the studies to date on the PMWS outbreak in Sweden which has been closely monitored since the initial outbreak of diseases on a single farm in 2003 was diagnosed. In Sweden PMWS was officially reported for the first time at a progeny test station during December 2003. One year later sixteen herds have been diagnosed and reported to the authorities: one progeny test station, one nucleus herd, thirteen piglet producers or integrated herds and one specialised fattening herd. Fifteen suspected cases have been deemed negative for PMWS with disease scenarios associated with Lawsonia intracellularis, Brachyspira pilosicoli, Mycoplasma hyopneumoniae, shortcomings in feed composition etc. To date, one year after the index case was reported ten other herds suspected for PMWS are under observation. The PMWS cases are concentrated into the southern part of Sweden. However, the disease is spreading slowly, and in general there are no obvious physical links between affected herds. The data generated to date in Sweden suggest that introduction of a new contagious microbe into the country that is responsible for the PMWS outbreaks appears unlikely, as does the spread of PMWS via semen. PMID- 15663214 TI - [Application and prospect on the researches of ecological cultivation of Coptis chinensis]. AB - Traditional cultivation of Coptis chinensis was carried out under shield by disafforestation, which has been used for over 300 years and lead to the severe destruction of natural environment. Several ecological modes for cultivation of Coptis chinensis have been developed, which increase the yields of Coptis chinensis, protect the resources of forest, and obtain economic and ecologic benefit. PMID- 15663215 TI - [Ecological technique of cultivating Coptis chinensis in woods and co-cultivating it with maize]. AB - OBJECTIVE: To establish ecologically new modes of cultivating Coptis chinensis in woods and co-cultivating it with maize. METHODS: Based on the experience obtained from plot comparative test and production test, we used application-oriented research methods, and established new Coptis chinensis cultivation techniques that protect the natural environment. RESULTS: Coptis chinensis was harvested 6 years after cultivation. Yield of Coptis chinensis cultivated in forest (69.5 kg per 0.067 hm2) was higher with 3.7% than that in controls which cultivated under shed (67 kg). Yield of Coptis chinensis co-cultivated together with maize (168.4 kg per 0.067 hm2) was lower with 15.8% than that in controls which cultivated under shed (200 kg). CONCLUSION: The new cultivation modes of Coptis chinensis is an ecologically new technique that assures the good growth of medicinal plants, cereals, forest and animal husbandry. PMID- 15663216 TI - [Quality assessment for Coptis chinensis planted with ecological techniques]. AB - OBJECTIVE: To evaluate the quality difference between Coptis chinensis planted with ecological techniques and shelf planted Coptis chinensis. METHODS: Ultraviolet-visible spectrophotometry, alcohol extract mensuration, moisture mensuration, and ash mensuration were used to determine the contents of total alkaloids, alcohol extract, water, and total ash of Coptis chinensis, which were planted in shelf, Rhus chinensis wood, M mulbery wood, corn wood, Magnolia officinalis wood, fruiter wood, shading net, and firry wood, respectively. RESULTS: The contents of total alkaloids and alcohol extract of Coptis chinensis Table planted with ecological techniques were higher than those of Coptis chinensis planted in shelf. The contents of water and total ash were less than 12% and 5%, respectively, which met the provisions of the pharmacopoeia. CONCLUSION: The quality of Coptis chinensis planted with ecological techniques is similar to that of Coptis chinensis planted in shelf. These ecological techniques for Coptis chinensis have become mature and practical. PMID- 15663217 TI - [Technique of storing Coptis chinensis seeds in damp sand under shed and fine cultivation of seedling]. AB - OBJECTIVE: To develop technique of storing Coptis chinensis seeds in damp sand under shed and fine cultivation of seedling and to enhance the cultivation seedling rate from seeds of Coptis chinensis. METHODS: With the old technique of seedling in woods as control, we screened the new method for storing seeds and cultivating seedlings. RESULTS: Seeds were picked up at May and stored in damp sand under shed until November, and then planted with technique of fine cultivation of seedling. The germination rate of seeds was up to 98%. One hundred and forty thousands seedlings could be getting from per kilogram seeds. CONCLUSION: Compared to the old method of cultivating seedling in woods, the technique of fine cultivation of seedling significantly increased the rate of cultivation seedlings by 8 times. This technique has been widely applied. PMID- 15663218 TI - [Analysis and assessment of Coptis chinensis for different parts, ages, and heights using Fourier transform infrared spectroscopy]. AB - OBJECTIVE: To study the infrared (IR) fingerprint spectra of Coptis chinensis for different parts, ages, and heights, and to analyze the integrate rules about the content of berberine component in Coptis chinensis for different parts, ages, and heights. METHOD: The Fourier transform infrared (FTIR) spectroscopy was applied to detect the infrared spectra of Coptis chinensis samples rapidly and non separately. RESULTS: The IR spectra of the root, leafstalk, and fibre of Coptis chinesis have their unique features. The content of berberine component in Coptis chinensis was different for different parts and the sequence: root > leafstalk > fibre. When Coptis chinensis grows, the content of berberine component in leafstalk also increases. The content of berberine component in leafstalk which planting in 1 200 m was less than that in 1 300, 1 400, and 1 500 m. The ages and heights provide no obvious influences on the content of berberine in the root of Coptis chinensis. CONCLUSION: Using FTIR spectroscopy, the quality of Coptis chinensis can be controlled, which provides a useful method for the standardized planting of Coptis chinensis. PMID- 15663219 TI - [Analysis of contents of berberine in Coptis chinensis of Lichuan]. AB - OBJECTIVE: To determine the contents of berberine in Coptis chinensis of Lichuan and establish its best cultivation scheme. METHOD: We used high performance liquid chromatography (HPLC) to measure the contents of berberine at different altitude, growth age, and leaves. RESULTS: Analytic data showed that the growth age and firry woods shading did not affect the contents of berberine in rhizome of Coptis chinensis. Low altitude was more suitable for Coptis chinensis to synthesize berberine. The contents of berberine in rhizome of Coptis chinensis with floral leaf were higher than those with lusterless and lustrous leaves, but no significant difference. CONCLUSION: The synthesis of berberine is closely correlated with shading conditions. It is recommended that the shading ratio should be reduced or the sheds removed in the middle growth age (2-3 years), and then the herb should be reshaped at the last year to enhance the synthesis of berberine, so as to obtain high-quality Coptis chinensis in the harvest. PMID- 15663220 TI - [Establishment of the control substance of plant drug and fingerprints of Coptis chinensis]. AB - OBJECTIVE: To establish the control substance of plant drug (CSPD) of Coptis chinensis Franch. and its proton nuclear magnetic resonance (1H NMR) and high performance liquid chromatography (HPLC) fingerprints for the purpose of original identification. METHODS: The CSPD and their 1H NMR and HPLC fingerprints of Coptis chinensis were obtained by standardized procedure. Chemical components were isolated from the CSPD by silica gel column chromatography. By elucidation of their structures, the assignments of the characteristic signals in fingerprints could be achieved. RESULTS; The 1H NMR and HPLC fingerprints of the samples from various sources had wonderful reproducibility and characteristic features. Furthermore, five main compounds were isolated from CSPD and their structures were authenticated by spectral analysis as palmatine chloride, berberine chloride, epiberberine chloride, coptisine chloride, and jatrorrhizine chloride, respectively. The 1H NMR and HPLC fingerprints of the CSPD of Coptis chinensis showed mainly the characteristic signals of the berberine-type compounds isolated in this work. CONCLUSION: The 1H NMR and HPLC fingerprints of the CSPD of Coptis chinensis exhibit the structures and total composition of the main active constituents in it, and can be used for its original identification and quality evaluation. PMID- 15663221 TI - [Effects of two test-meals on transient lower esophageal sphincter relaxation in patients with gastroesophageal reflux disease and mechanism of gastroesophageal reflux]. AB - OBJECTIVE: To investigate the effects of standard meal and fat meal distending the fundus on transient lower esophageal sphincter relaxation (TLESR) and esophageal motility and to explore the mechanism of gastroesophageal reflux (GER) in patients with gastroesophageal reflux disease (GERD). METHODS: Eight patients with GERD (3 male, 5 female; median age: 43.5 ys) were enrolled in the study. All received 2 times of esophageal manometry and pH monitoring simultaneously for 30 min during fasting and 2 h after two different test-meals, including standard meal (SM) and fat meal (FM) on separate day at least 1 week apart. RESULTS: The frequency of TLESR significantly increased after 2 test-meals (P < 0.05). There were no significant difference in the frequency and duration of TLESR between SM group and FM group 1 h after meal (P > 0.05). However, the frequency of TLESR in FM group 2 h after meal was more than that in SM group and during fasting (P < 0.05). Lower esophageal sphincter pressure (LESP) significantly decreased in FM group than in SM group (P < 0.05). The contractive amplitude of post lower esophageal sphincter relaxation and the contractive amplitude of the distal esophagus had no difference after FM and SM. Acid reflux episodes and duration of pH < 4 were larger after FM than after SM (P < 0.05). A total of 50.2% of GER occurred during decreased LESP and 37.8% during TLESR after FM, while 61.7% of GER occurred during TLESR after SM. CONCLUSIONS: Both the SM and FM can increase the frequency of TLESR in patients with GERD. Decreased LESP and increased frequency of TLESR after FM are the major mechanism of GER, while reflux after SM may attribute to the increased frequency of TLESR. PMID- 15663222 TI - [Comparative activity of antihistamines on area under dose-response curve from histamine-induced wheal and flare responses in human skin]. AB - OBJECTIVE: To compare the activity of antihistamines by the index of area under dose-response curve (AUDRC) obtained from histamine-induced wheal and flare reactions. METHODS: Mizolastine 10 mg, loratadine 10 mg, and placebo were given to 90 healthy volunteers and 60 allergic patients in a double-blind and randomized manner. Histamine titration tests (histamine concentrations 54.3, 20.0, 7.3, and 2.7 mmol/L) were performed for each one before dosing and 2, 4, and 24 hours after dosing. The reactivity was evaluated by histamine-induced wheal and flare areas. The AUDRC values of the wheal and flare areas as a function of the natural logarithm transformed histamine concentration were calculated for each subject, and compared. RESULTS: There was no significant difference of the wheal and flare areas between health volunteers and allergic patients. The AUDRC(27-54.3 mmol/l.) for wheal and flare of mizolastine was 115.7, 23.4, 7.7, 49.8 and 902.1, 40.9, 2.6, 46.9 ln (mmol/L) x mm2 at each time (before dosing and 2, 4, 24 hours after dosing) respectively. Compared with loratadine [116.2, 80.2, 49.7, 71.9 and 957.6, 495.3, 153.5, 205.9 ln (mmol/L) x mm2], mizolastine decreased AUDRC(2.7 - 54.3 mmol/L significantly (P < 0.01). CONCLUSION: Histamine-induced wheal and flare inhibition test is a reliable pharmacodynamic model for antihistamines, and AUDRC may be an useful index to predict antihistamines pharmacodynamic activity. PMID- 15663223 TI - [Effect of insulin, cAMP, and dexamethasone on phosphoenolpyruvate carboxykinase gene promoter in vitro]. AB - OBJECTIVE: To study the effect of insulin, cyclic adenosine monophosphate (cAMP), and dexamethasone (DEX) on 550 bp (-600 -/+ 69) fragment of phosphoenolpyruvate carboxykinase (PEPCK) gene promoter by reporter gene. METHODS: The recombinant pGL2-PEPCK-Luc and the control plasmid pSV-beta-Galactosidase were co-transfected to rat hepatoma cell line (CBRH7919) by lipofectin. By measuring luciferase activity, we evaluated in vitro regulation of PEPCK gene promoter on reporter gene transcription. RESULTS: cAMP and DEX stimulated PEPCK promoter obviously; meanwhile, they also had accumulative effects. At different physiological concentrations, insulin had a suppressive effect on PEPCK promoter, which was dose-independent. CONCLUSION: There is a perfect feedback mechanism for PEPCK promoter in hepatoma cell. 550 bp (-600 -/+ 69) fragment of PEPCK may be a candidate gene in the gene therapy of diabetes. PMID- 15663224 TI - [Treatment of vertebral hemangioma with percutaneous vertebroplasty]. AB - OBJECTIVE: To investigate the efficacy of percutaneous vertebroplasty in the treatment of vertebral hemangioma. METHODS: Seven patients with vertebral hemangiomas were treated by percutaneous vertebroplasty, including one case of cervical, three cases of thoracic, and three cases of lumbar hemangiomas. The average score of the 6-point behavioral pain rating scale was 2.67 +/- 0.41, and the average score of ambulation was 2.83 +/- 0.33. Guilty vertebral bodies were orientated with fluoroscopy. The procedures were performed under local anesthesia. The image features were also analyzed. Unipedicular or bipedicular approaches were used in 6 cases of thoracic and lumbar hemangiomas. The cervical anterior-lateral approach was adopted in one case of cervical hemangioma. 4-7 ml of 15%-20% bone cement was mixed and injected into the vertebral body to form a cast in the lesions. Re-examination of clinical symptoms, plain film, and CT were made for 1, 3, and 9 months of post-procedure follow-up. RESULTS: Good results were achieved in all the seven cases. Pain was completely relieved in 5 cases and partially relieved in 2 cases. Symptom was also recovered in 2 patients with radiculopathy. No recurrence was found after 1-9 months of postoperative follow up. CONCLUSION: Treatment of vertebral hemangioma with percutaneous vertebroplasty is safe and effective with minimal invasion. PMID- 15663225 TI - [Application of Neuroform stent in the treatment of intracranial aneurysm]. AB - OBJECTIVE: To introduce the application of Neuroform stent in the treatment of intracranianl wide neck aneurysm. METHODS: Eight patients (9 aneurysms), including 3 males and 5 females, aged from 41 to 74, were treated. Among the 9 aneurysms, there were 3 wide-neck aneurysms in internal carotid artery (ICA), 4 vertebral aneurysms (in 3 cases), and 2 basilar tip giant aneurysms. Heparinization were given for all procedures after femoral artery Seldinger's puncture. Stents were released through 3 m Transcend 0.014 guide wires, which were posited in the aneurysm-carry arteries first, or, through a 205 cm Transcend 0.014 guiding wire. Further coiling was selected for some cases. After the treatment 24 h heparinization were maintained. For coiling cases, aspirin (300 (mg/d) and ticlopiding (250 mg/d) were given at first 6 weeks, and aspirin (300 mg/d) was given following 6 months. For stent alone cases, only 1 month aspirin (300 mg/d) was given. RESULTS: In one ICA aneurysm, the stent moved to the bifurcation of ICA while a coil was pushed into the aneurysm lumen, and the ICA spasmed. Partial occlusion achieved in the aneurysm. The patient died due to bleeding of the aneurysm 20 h after anticoagulation treatment with heparin. One dissection aneurysm and 2 fusiform aneurysms (in bilateral vertebral arterys of one patient) were treated with stents only. The dissection aneurysm was completely occluded after 3 months. In 1 basilar artery (BA) tip giant aneurysm, the distal part of BA spasmed immediately after stenting. Complete occlusion was achieved with coils 4 months later. Another BA tip aneurysm was partially occluded after the stent was deployed. The other 3 aneurysms were completely embolized with the protection of the Neuroform stents. There is no further DSA follow-up of the cases. CONCLUSIONS: The Neuroform stent is easy to pass through the tortuous vessels. Combine with coils, it may be used in the treatment of wide neck aneurysms. However, it has the risk of migration because of the softness. PMID- 15663226 TI - [Tracheal intubation under general anesthesia in patients with difficult laryngoscopy]. AB - OBJECTIVE: To evaluate the safety and efficacy of oral endotracheal intubation in the patients with difficult laryngoscopy undergoing general anesthesia. METHODS: A total of 1 683 patients with difficult laryngoscopy, aged 1.5-67 yr, and scheduled for the elective plastic surgery were observed in this study from 1989 1997. All these patients were at American Society of Anesthesiologist physical status I. According to the preoperative predictive results for difficult laryngoscopy, we classified these patients into two groups: Group I included 1 375 patients, whose epiglottis could be viewed (laryngoscopic view grades II and III); and Group II, included 308 patients, whose epiglottis could not be viewed (laryngoscopic view grade IV). For group I, anesthesia was induced with thiopentone 4-5 mg/kg and succinylcholine 1 mg/kg; Laryngoscopy was carried out using modified Macintosh method. For Group II, anesthesia was induced with a total intravenous anesthesia or inhaled anesthesia; anesthetic depth was required to effectively inhibit laryngeal reflexes with reservation of spontaneous breathing. Tracheal intubation was performed by fiberoptic stylet laryngoscope (FOSL). During anesthesia induction and tracheal intubation procedures, electrocardiogram, arterial pressure, heart rate and pulse oxygen saturation (SpO2) were continuously monitored. Complications of intubation (arrhythmia, and so on) were observed and recorded. Immediately after laryngoscopy and successful intubation, patients were examined for any traumatic injuries at teeth, lips, tongue, and oropharyngeal tissues. RESULTS: In group I, tracheal intubation was accomplished by the first attempt in 1 279 cases (93.0%) and the intubation time was less than 3 min in 1 304 cases (94.8%). In group II, tracheal intubation was accomplished by the first attempt in 114 patients (37.0%), and 123 patients (39.9%) had the intubation time of less than 3 min. Tracheal intubation was successful by the second or third attempt in 96 patients of group I and 156 patients of group II, respectively. Thirty-eight patients required four or more attempts, which only occurred in group II. Of all the complications of tracheal intubation, the traumatic complications were most common. The incidences of traumatic complications in the patients with laryngoscopic view grade II, III (group I ) and IV (group II) were 0.7%, 3.9% and 14.3%, respectively. Other complications such as respiratory depression were only seen in group II. A pooled incidence of the intubation complications was 6.7% (113/1 683). CONCLUSION: An anesthesiologist who is skillful in difficult airway management may safely manage the airway in the patients with difficult laryngoscopy under general anesthesia. PMID- 15663227 TI - [Comparison of endoscopic versus open surgical treatment of carpal tunnel syndrome]. AB - OBJECTIVE: To compare the results of endoscopic surgical decompression of carpal tunnel release (CTR) with open CTR in patients with idiopathic carpal tunnel syndrome (CTS). METHODS: Forty patients with CTS were randomly chosen for a prospective study from April 2000 to August 2002. Endoscopic CTR was performed in 23 CTS patients (26 sides) and open CTR was performed in 17 CTS patients (21 sides). Five parameters were evaluated, including each patient's symptom amelioration, complication, operation time, hospitalization time, and the time needed to resume normal lifestyle. The mean follow-up time was 2 years. RESULTS: No significant difference was observed between the endoscopic CTR group and open CTR group in regard to symptom amelioration, electromyographic testing, and complications. In comparison to open CTR, endoscopic CTR significantly decreased operation time, hospitalization time, and the time needed to resume normal lifestyle and activity (P < 0.01). CONCLUSION: Compared with open CTR, endoscopic CTR has the advantage of reduced pain of scar and shortened time of hospitalization and recovery. It is a safe and effective method for treating idiopathic CTS. PMID- 15663228 TI - [A strategy for targeting gene therapy against cancer mediated by epidermal growth factor receptor]. AB - OBJECTIVE: To establish a protocol for the targeting gene therapy against cancer with rich epidermal growth factor receptor (EGFR). METHODS: A recombinant pcDNA3.1-PE III mut was constructed and combined with a non-viral vector, a fusion protein histone H1, epidermal growth factor C-loop previously expressed by us, to be a protein-DNA complex in vitro. Using the complex to treat BT-325 and Hela cancer cells with EGFR and JK cells without EGFR. The killing rates of the cells was calculated after 48 h of incubation at 37 degrees C. RESULTS: To BT-325 and Hela cells, the killing rates were 46.03% and 48.12% respectively. To JK cells, the complex had no killing function. CONCLUSION: The protocol for targeting gene therapy against cancer with EGFR has been established successfully. PMID- 15663229 TI - [Comparison of preemptive analgesia efficacy between etoricoxib and rofecoxib in ambulatory gynecological surgery]. AB - OBJECTIVE: To compare the preemptive analgesia efficacy between two cycloxygenase 2 inhibitors, rofecoxib and etoricoxib in the ambulatory uterine evacuation patients. METHODS: In this randomized, double-blinded, placebo-controlled trial 60 patients were randomly divided into three groups and received a single dose of placebo, rofecoxib 50 mg, or etoricoxib 120 mg, respectively, before operation. Patient's visual analogue score (VAS) was rated postoperatively at 15 min, 30 min, 60 min, time-to-discharge, 6 h and 24 h. Fentanyl (in post-anesthesia care unit) and paracetamol (at home) were supplementary analgesics and the dosage was also recorded. Patient's satisfaction score was rated at 24 h postoperatively. RESULTS: Etoricoxib 120 mg and rofecoxib 50 mg were significantly superior to placebo at 6 h postoperatively (P < 0.05) while there was no significant differences of VAS at other time points. The amounts of Fentanyl used in post anesthesia care unit were similar in three groups, but paracetamol taken at home was much less in rofecoxib group and etoricoxib group than in placebo group (P < 0.01). Compared to rofecoxib, etoricoxib provided better pain relief after discharge (P < 0.05). The overall pain management satisfaction score was significantly higher in etoricoxib group (96 +/- 7) than in other groups (P < 0.01). CONCLUSION: Preemptive rofecoxib 50 mg and etoricoxib 120 mg may significantly decrease VAS at 6 h postoperatively, and reduce the usage of analgesics in ambulatory uterine evacuation patients. Etoricoxib 120 mg offeres better pain relief at home compared with rofecoxib 50 mg. PMID- 15663230 TI - [Palmitic acid induces apoptosis in human hepatoma cell line, HepG2 cells]. AB - OBJECTIVE: To investigate the effects of palmitic acid (PA) on human hepatocytes and its mechanism. METHODS: We administered a mimic hyperlipidemia condition of 0.2-0.4 mmol/L PA to human hepatoma cell line, HepG2 cells. Cell viability was determined by Trypan blue staining. Cell cycle and early apoptosis were determined by propidium iodide and/or Annexin V staining, and the levels of Bcl-2 and Bax were analyzed by flow cytometry. RESULTS: An inhibition of cell growth was observed at a dose- and time-dependent manner in HepG2 cells after the treatment of PA. An apoptosis with appearance of sub-G1 fraction determined by cell cycle analysis significantly increased after the treatment of PA for 4 days. Bcl-2 level slightly decreased; in contrast, Bax level elevated markedly, which resulted in a significant decrease of Bcl-2/Bax ratio. CONCLUSION: PA may induce cell death on hepatocytes via mitochondria-mediated apoptosis by reducing the level of Bcl-2/Bax. PMID- 15663231 TI - [Clinical validity of anti-thyroperoxidase antibody and anti-thyroglobulin antibody]. AB - OBJECTIVE: To evaluate the clinical validity of anti-thyroperoxidase antibody (anti-TPOAb) and anti-thyroglobulin antibody (anti-TgAb). METHOD: Serum levels of anti-TPOAb and anti-TgAb were assayed using chemiluminescence immunoassay in 434 subjects, including 51 patients with Hashimoto's thyroiditis, 58 with Graves' disease, 68 with nodular goiter, 56 with thyroid adenoma and carcinoma, 56 with subacute thyroiditis, 65 with euthyroid non-thyroid endocrine disease, 35 with euthyroid non-thyroid autoimmune diseases, and 45 euthyroid controls. RESULTS: The highest level and most positive results of serum anti-TgAb and anti-TPOAb were observed in patients with Hashimoto's thyroiditis (median 373 and 6 974 U/ml, positive rate 84.3% and 86.3%), followed by patients with Graves' disease (median 84 and 1 369 U/ml, positive rate 44.8% and 72.4%). Serum anti-TgAb and anti-TPOAb were also more common in patients with subacute thyroiditis and other autoimmune diseases than in the controls. CONCLUSION: The assay of serum anti TPOAb and anti-TgAb by chemiluminescence immunoassy are useful in the differential diagnosis of autoimmune thyroid disease. PMID- 15663232 TI - [Tear film function of patients with type 2 diabetes]. AB - OBJECTIVE: To study the tear film function of patients with type 2 diabetes and to investigate the risk factors of dry eye in these patients. METHODS: Totally 111 patients with type 2 diabetes and 100 age- and sex-matched control subjects were studied. Tear film function was evaluated by dry eye syndrome, tear breaking up time (BUT), corneal fluorescein staining, Schirmer I test (SIt), and tear film lipid layer observation with tear scope. Dye eye score was calculated with the results of these tests. RESULTS: When compared with the controls, patients with type 2 diabetes showed higher dry eye score (diabetics 3.28 +/- 1.56, control 2.31 +/- 1.50, P < 0.01) and faster BUT [diabetics (6.50 +/- 4.84) s, control (12.26 +/- 7.16) s, P < 0.01], but similar SIt [diabetics (10.61 +/- 6.86) s, control (10.92 +/- 7.05) s, P > 0.05]. More diabetic patients were diagnosed as dry eye(diabetics 19.8%, control 8.0%, P < 0.05). According to their retinopathy, the diabetic patients were divided into three groups: without diabetic retinopathy (DR), with background DR, and with proliferative DR. For these three groups, the dry eye scores were 2.95 +/- 1.50, 3.38 +/- 1.48 and 4.11 +/- 1.60, respectively (P < 0.01); the SIt were (10.95 +/- 6.89) mm, (11.71 +/- 7.30) mm and (7.63 +/- 5.20) mm, respectively (P > 0.05); the BUT were (7.53 +/- 5.23) s, (5.88 +/- 4.10) s and (4.47 +/- 4.17) s (P < 0.05). Patients with DR were then devided into two groups: with photocoagulation and without photocoagulation. For these two groups, the dry eye scores were 4.71 +/- 1.14 and 3.26 +/- 1.15, respectively (P < 0.01); the BUT were (2.93 +/- 2.06) s and (6.26 +/- 4.36) s, respectively (P < 0.01); the SIt were (7.21 +/- 6.51) mm and (11.33 +/- 6.73) mm, respectively (P < 0.05); the rates of corneal fluorescein staining were 50.0% and 17.9%, respectively (P < 0.05). Dry eye score had a good correlation with diabetic retinopathy and photocoagulation (P < 0.01), but was poorly correlaed with age, gender, insulin, duration of diabetes mellitus, and metabolic control (P > 0.05). CONCLUSIONS: Patients with type 2 diabetes tend to develop tear film dysfunction. The disorders of tear film quantity and quality seem relevant to the stage of diabetic retinopathy and photocoagulation. PMID- 15663233 TI - [Relationship of msp AI polymorphism in cytochrome P450c 17alpha gene with bone mass and bone size in Shanghai men of Han nationality]. AB - OBJECTIVE: To investigate the relationship of Msp AI polymorphism in the promoter region of cytochrome P450c 17alpha (CYP17) gene with bone mass and bone size in Shanghai men of Han nationality. METHODS: The CYP17 Msp AI genotype was determined by polymerase chain reaction-restriction fragment length polymorphism in 397 unrelated men (324 healthy men, 73 osteoporosis patients) aged 46-80 years of Han nationality in Shanghai. Bone mineral density (BMD), bone mineral content (BMC), and bone cross-section area (CSA) at lumber spine 1-4 and at any sites of proximal femur, including femoral neck, trochanter and Ward's triangle were measured by duel-energy X-ray absorptiometry. RESULTS: Frequency distributions of CYP17 genotype were TC (51.1%), CC (33.8%), and TT (15.1%). The allele frequencies T and C were 40.7% and 59.3%, respectively. Allele frequencies did not deviate from Hardy-Weinberg equilibrium. The frequencies of CYP17 Msp AI genotype did not show difference between osteoporosis cases and healthy controls. In group of all population, or in subgroups of osteoporosis patients and healthy men, CYP17 Msp AI genotype was not significantly associated with BMD, BMC, and CSA at lumber spine 1-4 and at any sites of proximal femur after having been adjusted for age, weight, and height with analysis of covariance. CONCLUSION: Msp AI polymorphism of CYP17 gene is not a genetic factor that influence the variation of bone mass and bone size in Shanghai men of Han nationality. PMID- 15663234 TI - [Clinical aspects of mature ovarian teratoma: result of 695 cases]. AB - OBJECTIVE: To evaluate the clinical aspects of mature ovarian teratoma. METHOD: We retrospectively studied 695 patients operated in PUMC Hospital between January 1990 and December 2000. RESULTS: Laparoscopy surgery had shorter hospitalization day, less blood loss during surgery, shorter operation time compared with laparotomy (P < 0.001). Routine contralateral wedge resection was done in 601 cases, among which 459 had normal inspection and palpatation. Only one (0.22%) was pathologically confirmed bilateral tumor. Torsion was the major complication (7.5%). Average tumor size of torsion was (10.2 +/- 4.5) cm. Probability of torsion increased as the tumor enlarged, which was less than 4.4% when the tumor size was less than 6 cm and greater than 10% if size greater than 8 cm. Rupture, infection, and malignant transformation were rare. CONCLUSIONS: Laparoscopy is superior to laparotomy in the treatment of mature ovarian teratoma. If the contralateral ovarian is within normal configuration, contralateral wedge resection is unnecessary. When the tumor size is greater than 8 cm in diameter, it will have more chance to twist. PMID- 15663235 TI - [Gene expression of type X collagen in the intervertebral disc of idiopathic scoliosis patients]. AB - OBJECTIVE: To observe the characteristics of gene expression of type X collagen in the cartilage of end-plate and the fibrous annulus in the intervertebral disc of idiopathic scoliosis (IS) patients. METHOD: Investigating the expression of type X collagen in the peak disc and the lower end disc of 21 IS patients, the peak disc of 16 congenital scoliosis (CS) and the lumbar disc of 3 normal people (according with the principle of medical ethnics) by reverse transcript polymerase chain reaction. RESULTS: The expression of type X collagen in the concave side of IS peak disc was higher than the convex side (P < 0.05). There was no significant difference of gene expression of type X collagen between the convex side and the concave side of the lower end disc (P > 0.05). The gene expression of type X collagen in the IS peak disc was higher than those of lower end disc (P < 0.05). For the CS peak discs, the expression of type X collagen of the concave side was higher than the convex side (P < 0.05). CONCLUSION: The expression of type X collagen of the IS peak disc increases, and the expression of type X collagen of the concave side is higher than the convex side. These changes may be secondary. PMID- 15663236 TI - [Clinical effectiveness of ropivacaine mesylate in epidural anesthesia and postoperative analgesia]. AB - OBJECTIVE: To compare the clinical effectiveness of ropivacaine mesylate and ropivacaine hydrochloride in epidural anesthesia and postoperative analgesia. METHODS: Forty-four patients undergoing epidural anesthesia for intraperitoneal hysterectomy or hysteromyomectomy were randomly assigned to two equal groups. Group ropivacaine hydrochloride received 15 ml ropivacaine hydrochloride of 7.5 mg/ml in anesthetic and 2 mg/ml in postoperative analgesic subsequently while group ropivacaine mesylate correspondingly received ropivacaine mesylate of both 15 ml of 8.94 mg/ml and 2.374 mg/ml. RESULTS: No significant differences were found in the onset, extent, and duration of sensory and motor blockage, and also in the hemodynamic stability, when ropivacaine mesylate was comparable with ropivacaine hydrochloride. No severe adverse events were observed in this study. CONCLUSION: Ropivacaine mesylate is an effective and safe alternative to ropivacaine hydrochloride in epidural anesthesia and postoperative analgesia. PMID- 15663237 TI - [Research development of Coptis chinensis]. AB - To summarize the research and application of ecological planting technique for Coptis chinensis, and describe the recent development of its chemical components, pharmacological effects and clinical applications. PMID- 15663238 TI - [Mesopothamic origin of angels:How did an angel acquire wings?]. PMID- 15663239 TI - [Angels in the medieval sky:. Philosophical questions on the angelologia]. PMID- 15663240 TI - [Angels and other glossy pictures]. PMID- 15663241 TI - [Story behind a dozen]. PMID- 15663242 TI - [Hippocrates is alive]. PMID- 15663243 TI - [Memoires of Matti Ayarapaa]. PMID- 15663244 TI - [Georg Car von Dobeln and his doctors]. PMID- 15663245 TI - [Asklepios and his temple tell the pictorial story of antique medicine]. PMID- 15663246 TI - [Practice of medicine and a piece of art]. PMID- 15663247 TI - [Neurophysiology of pleasure]. PMID- 15663248 TI - [Traditional drinks of Christmas]. PMID- 15663249 TI - [Santa with a cool game in his sack]. PMID- 15663250 TI - [Adventures of a clinician in the jungle of bureaucracy. What every researcher should know before starting a clinical study?]. PMID- 15663251 TI - [Is there enough water to improve hygiene especially in developing countries?]. PMID- 15663252 TI - [Sources of medicine for developing countries]. PMID- 15663254 TI - Bovine genomics update: making a cow jump over the moon. AB - Recent research in bovine genetics has focused on characterization of the biological differences underlying phenotypic variation for qualitative and quantitative traits of economic value in existing cattle populations. The much anticipated benefits of DNA-based tools to routinely guide selection decisions for more efficient genetic gain and widened profit margins have not been fully met since the origin of this premise over two decades ago. However, the impending release of a high-quality draft genome sequence in 2005 should mark a turning point in these efforts. The following discussion summarizes how the bovine genetics research community has positioned itself to fully utilize a genome sequence resource and frames how genome sequence information can not only be applied to better implement marker-assisted selection, but also address rising consumer concerns relative to animal well-being and food safety. PMID- 15663255 TI - Genetic studies of two sister species in the Drosophila melanogaster subgroup, D. yakuba and D. santomea. AB - We performed genetic analysis of hybrid sterility and of one morphological difference (sex-comb tooth number) on D. yakuba and D. santomea, the former species widespread in Africa and the latter endemic to the oceanic island of Sao Tome, on which there is a hybrid zone. The sterility of hybrid males is due to at least three genes on the X chromosome and at least one on the Y, with the cytoplasm and large sections of the autosomes having no effect. F1 hybrid females carrying two X chromosomes from either species are perfectly fertile despite their genetic similarity to completely sterile F1 hybrid males. This implies that the appearance of Haldane's rule in this cross is at least partially due to the faster accumulation of genes causing male than female sterility. The larger effects of the X and Y chromosomes than of the autosomes, however, also suggest that the genes causing male sterility are recessive in hybrids. Some female sterility is also seen in interspecific crosses, but this does not occur between all strains. This is seen in pure-species females inseminated by heterospecific males (probably reflecting incompatibility between the sperm of one species and the female reproductive tract of the other) as well as in inseminated F1 and backcross females, probably reflecting genetically based incompatibilities in hybrids that affect the reproductive system. The latter 'innate' sterility appears to involve deleterious interactions between D. santomea chromosomes and D. yakuba cytoplasm. The difference in male sex-comb tooth number appears to involve fairly large effects of the X chromosome. We discuss the striking evolutionary parallels in the genetic basis of sterility, in the nature of sexual isolation, and in morphological differences between the D. santomea/D. yakuba divergence and two other speciation events in the D. melanogaster subgroup involving island colonization. PMID- 15663256 TI - Microsatellite variation among divergent populations of stalk-eyed flies, genus Cyrtodiopsis. AB - Microsatellite primers are often developed in one species and used to assess neutral variability in related species. Such analyses may be confounded by ascertainment bias (i.e. a decline in amplification success and allelic variability with increasing genetic distance from the source of the microsatellites). In addition, other factors, such as the size of the microsatellite, whether it consists of perfect or interrupted tandem repeats, and whether it is autosomal or X-linked, can affect variation. To test the relative importance of these factors on microsatellite variation, we examine patterns of amplification and allelic diversity in 52 microsatellite loci amplified from five individuals in each of six populations of Cyrtodiopsis stalk-eyed flies that range from 2.2 % to 11.2% mitochondrial DNA sequence divergence from the population used for microsatellite development. We find that amplification success and most measures of allelic diversity declined with genetic distance from the source population, in some cases an order of magnitude faster than in birds or mammals. The median and range of the repeat array length did not decline with genetic distance. In addition, for loci on the X chromosome, we find evidence of lower observed heterozygosity compared with loci on autosomes. The differences in variability between X-linked and autosomal loci are not adequately explained by differences in effective population sizes of the chromosomes. We suggest, instead, that periodic selection events associated with X-chromosome meiotic drive, which is present in many of these populations, reduces X-linked variation. PMID- 15663257 TI - Predicting the annual effective size of livestock populations. AB - Effective population size (Ne) is an important parameter determining the genetic structure of small populations. In natural populations, the number of adults (N) is usually known and Ne can be estimated on the basis of an assumed ratio Ne/N, usually found to be close to 0.5. In farm animal populations, apart from using pedigrees or genetic marker information, Ne can be estimated from the number N of breeding animals, and a value of 1 is commonly assumed for the ratio Ne/N. The purpose of this paper is to show the relation between effective population size and breeding herd size in livestock species. With overlapping generations, Ne can be predicted knowing the number of individuals entering the population per generation and the variance of family size, the latter being directly related to the survival pattern (or replacement policy) in the breeding herd. Assuming an ideal survivorship leading to a geometric age distribution, it can be shown that the number of breeding animals tends to overestimate effective size, particularly in early-maturing species. The ratio of annual effective size to the number of breeding animals is shown to be equal to [1 + (a- 1)(1 - s)]2/(1 - s2), where a is the age at first offspring and s is the survival rate (including culling) of the parents between successive births. This expression shows to what extent inbreeding may be determined by demography or culling policy independently of the actual herd size. In many situations a fast replacement or an early culling will increase annual effective size. Consequences for the management of small populations are discussed. PMID- 15663258 TI - The effect of an imprecise map on interval mapping QTLs. AB - The statistical analysis of quantitative trait locus (QTL) experiments relies on the use of a linkage map of the markers genotyped. Such a map is, at best, a good estimate of the true map. Resources might be diverted into developing better marker maps or improved maps become available after the analysis, raising concerns over the original analysis. It is therefore important to understand the sensitivity of QTL analysis to map inaccuracy. We have used simulation methods to investigate the consequences of an incorrect map on the results of a QTL analysis using interval mapping. Backcross data sets were generated with a particular map and then analysed with both the correct map and incorrect maps. If the incorrect maps maintained the true linkage groups (i.e. no markers were incorrectly assigned to another linkage group), the accuracy of the map had little or no impact on the ability to detect QTLs, the true significance levels of the tests or the relative placement of QTLs. When a marker was incorrectly placed on another linkage group, there was a small increase in the level of the test. After adjusting for this increase, there was a decrease in power to detect a QTL near the misplaced marker. This decrease was of a similar magnitude to that found when using a single-marker analysis compared with interval mapping. These results mean that QTL analyses can proceed without the need for very accurate marker maps, and that estimated QTL positions can be translated onto updated maps without the need for reanalysis. PMID- 15663259 TI - Quantitative trait loci with parent-of-origin effects in chicken. AB - We investigated potential effects of parent-of-origin specific quantitative trait loci (QTL) in chicken. Two divergent egg-layer lines differing in egg quality were reciprocally crossed to produce 305 F2 hens. Searching the genome using models with uni-parental expression, we identified four genome-wide significant QTL with parent-of-origin effects and three highly suggestive QTL affecting age at first egg, egg weight, number of eggs, body weight, feed intake, and egg white quality. None of these QTL had been detected previously using Mendelian models. Two genome-wide significant and one highly suggestive QTL show exclusive paternal expression while the others show exclusive maternal expression. Each of the parent-of-origin specific QTL explained 3-5 % of the total phenotypic variance, with the effects ranging from 0.18 to 0.4 phenotypic SD in the F2. Using simulations and further detailed analyses, it was shown that departure from fixation in the founder lines, grand-maternal effects (i.e. mitochondrial or W linked) and Z-linked QTL were unlikely to give rise to any spurious parent-of origin effects. The present results suggest that QTL with parent-of-origin specific expression are a plausible explanation for some reciprocal effects in poultry and deserve more attention. An intriguing hypothesis is whether these effects could be the result of genomic imprinting, which is often assumed to be unique to eutherian mammals. PMID- 15663260 TI - Influence of the intraluminal thrombus in abdominal aortic aneurysms. PMID- 15663261 TI - What have we learnt after two years working with the da Vinci robot system in digestive surgery? AB - Robotic- assisted surgery has been introduced recently in order to overcome some of the difficulties surgeons encounter during advanced laparoscopic surgery. Due to the 3D vision equipment, higher number of degrees of freedom in manipulating instruments and better ergonomics it is hoped that by using robot techniques the indications of minimally invasive surgery in the field of digestive surgery can be broadened or that difficult procedures will be easier to perform. Since the introduction of the system in our hospital now almost two years ago 70 procedures have been performed with the aid of the da Vinci system covering the whole spectrum of GI surgery. Conversion took place in 2.5% and peroperative morbidity related to the use of robotic techniques was 10%. Although we had the subjective feeling that the procedures were indeed easier to perform and more relaxing for the surgeon, some major problems still exist as the complete lack of tactile feedback and the cost effectiveness of these procedures. Before robotics can be introduced in the every day clinical practice of the surgeon, its true benefit still needs to be established. This can only be done by well randomised prospective studies comparing one technique with the other. PMID- 15663262 TI - Diagnostic and therapeutic strategies in endocrine duodenopancreatic tumours. PMID- 15663263 TI - Surgical management of cystic pancreatic tumours. PMID- 15663264 TI - Combined carotid and coronary artery disease. AB - Stroke is a devastating complication of coronary artery bypass surgery. It increases the mortality and morbidity of the operation, and prolongs the length of hospital stay. Functional disability from the stroke may dramatically reduce the quality of life post-operatively, and some patients will require permanent institutional care. Although there are many causes of stroke extra-cerebral carotid stenosis is the most important predictor of stroke for patients undergoing coronary artery bypass grafting (CABG). If patients are identified before coronary intervention, they may benefit from treatment of their carotid disease. Carotid endarterectomy can be performed before or simultaneously with coronary artery bypass. However, carotid angioplasty with stenting and cerebral protection may offer a superior method to conventional surgery in this high-risk group of patients. PMID- 15663265 TI - How robust is the evidence supporting prophylactic carotid endarterectomy in patients undergoing coronary bypass? PMID- 15663266 TI - Occlusion of internal mammary grafts: a review of the potential causative factors. AB - The outcome of patient undergoing CABG is largely dependant on the long-term patency of the conduit used. Internal mammary artery (IMA) is considered whenever possible due to its improved long-term functionality over saphenous vein graft. However, a 10% rate of late arterial closure is described without well-known predictors. Chronic competition induced by a moderate coronary lesion on the bypassed native vessel is thought to be a major factor of arterial graft shrinkage even if conflicting data are reported in the available literature. Therefore, the decision to use an IMA to bypass a moderate native coronary lesion should be carefully weighted. When angiography is doubtful, more accurate functional investigations should be considered. Among them, pressure-derived fractional flow reserve could give an immediate answer of whether an intermediate lesion should be bypassed. PMID- 15663267 TI - Endofibrosis of the iliac arteries: an underestimated problem. AB - Arterial endofibrosis is a recently discovered artery disease that is specific to endurance athletes. Cycling is the sport that has shown the greatest number of cases. The endofibrosis is located most frequently in the external iliac artery, but other locations must also be sought. Thigh pain during supramaximal exercise and patient questioning provide strong indications. The diagnosis is confirmed when there is concordance of exercise-induced symptoms, a positive exercise test result and discrete arteriographic characteristics. Treatment is based on folate supplements and standardised surgery carried out by an experienced surgical team. PMID- 15663268 TI - Aortoiliac steno-occlusion in young women: a single center experience and review of the literature. AB - Peripheral obliterating arterial disease characterized by aortoiliac steno occlusion occurring in relatively young women of small stature, is frequently reported in the literature under the name small aorta syndrome. Although it remains unclear whether small aorta syndrome represents a separate entity, the small size of the distal aorta increases risk for aortoiliac occlusive disease. Patients usually present with lower extremity claudication and typical risk factors. This paper shows an analysis of the literature focusing on the pathogenesis, clinical features, risk factors and treatment, as well as a single center experience with this disorder. PMID- 15663269 TI - The multidisciplinary approach to limb salvage. AB - There is considerable morbidity and mortality associated with ulcerations of the lower limbs in patients with diabetes as well as non-diabetic patients. The role of the primary care physician and podiatrist in the evaluation, diagnosis, and management of lower extremity wounds is critical. Careful assessment and management of vascular disease by vascular surgeons and interventionalists, biomechanical foot abnormalities by the podiatrist, wound coverage by the plastic surgeons and aggressive treatment of infections and metabolic derangements by the physicians are required. The multidisciplinary approach provides a comprehensive treatment protocol and significantly increases the chances of successfully healing the ulcer and prevents recurrence. It also provides for increased academic output through publications, participation in clinical trials and educational venues. PMID- 15663270 TI - Pedal artery bypass. AB - Pedal artery bypass has proven to be a safe, simple and durable procedure. Failure of a pedal artery reconstruction seldom results in an amputation that is more proximal than it would have been without previous attempted bypass. Thus an aggressive strategy in the revascularisation of the forefoot in case if critical ischemia based on tibial occlusive disease is worthwhile. PMID- 15663271 TI - The value of modern ultrasonographic techniques and computed tomography in detecting and staging of pancreatic carcinoma. AB - OBJECTIVE: to assess the clinical value of ultrasonographic methods and computed tomography in diagnosing and staging pancreatic carcinoma. METHODS: prospective clinical trial of 140 patients (64 women and 77 men; mean age 59,6) operated on for pancreatic carcinoma between 2000 and 2004. In each case helical CT, routine , color- and power Doppler and 3-D USG were performed to detect and stage cancer. Analyses of accuracy, sensitivity, specificity, PPV and NPV of ultrasonographic methods and CT were made. RESULTS: 3-D USG showed the best accuracy of local staging (T): 95.6%. CT was the most accurate in lymph node assessment: 91.3%. The accuracy of CT, 3-D USG and power-Doppler at detecting vascular infiltration was 93.1%. CONCLUSIONS: diagnostic accuracy of modern ultrasound techniques is comparable to helical CT in detecting and staging pancreatic carcinoma. USG is recommended due to the relatively low cost, non-invasiveness and availability of the procedure. PMID- 15663272 TI - Pylorus-preserving pancreaticoduodenectomy with external pancreatic remnant drainage. AB - BACKGROUND: Pancreaticoduodenectomy is the mainstay of surgical treatment for various benign and malignant diseases of the head of the pancreas and the periampullar region. The incidence of postoperative morbidity remains significantly high. Various modifications have been advocated for the restoration of the digestive continuity and drainage of bile and pancreatic ducts in order to diminish this high morbidity rate. METHODS: A new modification with the combination of an end-to-end duodenojejunal anastomosis and creation of an external pancreatic fistula following pylorus-preserving pancreaticoduodenectomy is described. Our initial experience with this modification in seven patients is reported and its advantages are discussed. RESULTS: There was no mortality, while morbidity was 29%, including postoperative pneumonia and psychosis in one patient, and intra-abdominal abscess in another patient. No significantly delayed gastric emptying was observed and no patient developed diabetes, anastomotic ulcer or reflux cholangitis postoperatively. CONCLUSIONS: The reported technique seems to be a safe modification of restoration after pancreaticoduodenectomy, with maintenance of anatomical gastrointestinal continuity, avoidance of complications from anastomosis with the pancreatic remnant, as well as maintenance of endocrine pancreatic function. However, long-term follow-up and larger comparative studies are needed to evaluate the impact of this modification on morbidity and mortality, including long-term nutritional complications. PMID- 15663273 TI - Gastric cancer in young patients--is there any hope for them? AB - Gastric cancer usually affects people older than sixty years. This type of cancer is very rare in adults under thirty years of age. In addition, the prognosis in this part of population is grave due to the high incidence of undifferentiated tumours and advanced stage at time of diagnosis. Radical surgery affords the only chance for long term survival, but even this option is often limited upon finding locally advanced disease or peritoneal seeding. The following are case studies of three young adults from a group of 45 patients, who were treated between January 1st, 2000 and December 31st, 2003. PMID- 15663274 TI - Clinicopathological characteristics of patients with proximal third gastric carcinoma. AB - OBJECTIVE: Epidemiological studies show a continuing rise in the prevalence of proximal third gastric carcinoma (PGC), and the prognosis of patients with this carcinoma is poorer than that of patients with more distally located gastric carcinomas. We compared the clinicopathological features and prognosis of PGC patients with those of patients with middle/distal gastric carcinoma (MGC/DGC). MATERIAL AND METHODS: Of the 2696 patients diagnosed with gastric carcinoma who underwent surgery in our hospital in a 15-year period, 271 patients (10.1%) were diagnosed with PGC and retrospectively reviewed. RESULTS: T1-category tumours were less common in patients with PGC than in patients with MGC/DGC (p < 0.001). Lymph node invasion was more common in patients with PGC than in patients with MGC/DGC (p < 0.05). Tumour stage (stage I) and T category (T1) significantly influenced the 5-year survival rates of patients whose tumours were resected with curative intent. The 5-year survival rate of patients whose PGC were resected with curative intent was higher than that of patients whose PGC were resected with palliative intent (57.4 vs. 12.6%, p < 0.001). The 5-year survival rate was 49.3% for patients with PGC and 57.3% for patients with MGC/DGC (p = 0.0273). Multivariate analysis showed that tumour size, lymph node status, and resection with curative intent were significant prognostic factors for survival in patients with PGC. CONCLUSION: The poor prognosis of PGC is mainly due to its more advanced stage at diagnosis compared with that of more distally located gastric carcinoma. Early detection is important for improving the prognosis of patients with proximal third gastric carcinoma. PMID- 15663275 TI - Surgical interventions during STI 571 treatment of metastatic GIST: experience in six patients. AB - Gastrointestinal stromal tumours (GIST), previously classified as smooth muscle tumours, are the most common mesenchymal tumours of the digestive tract. Since the discovery of KIT (CD 117) expression, these tumours can be diagnosed confidently by pathology. Until recently, surgery was the only treatment available because these tumours were not sensitive to chemotherapy nor radiation therapy. In the long run most of these tumours recurred in the abdominal cavity or in the liver. Recently a new drug STI 573 (Glivec) showed very promising results in metastatic disease with response rates of about 65%. In six patients treated at our center, surgery was indicated during STI therapy because of subobstruction, skin necrosis, abdominal distention, bleeding. Surgery proved to be safe and efficient, allowing continuation of STI therapy in much better circumstances. PMID- 15663276 TI - Carotid artery surgery--critical analysis of indications in a regional Finnish vascular center. AB - BACKGROUND: The purpose of this study was to analyze retrospectively the treatment of patients referred for carotid artery stenosis to a vascular surgical unit in the 1990's. Main attention was paid to the appropriateness of the indications for CEA. MATERIAL AND METHODS: In the Pirkanmaa region (population of 440 000), all carotid surgery is performed in the regional University Hospital. All new referrals for vascular surgery because of carotid stenosis or bruit in 1990, 1992, 1994, 1996 and 1998 were included and case records reviewed. RESULTS: Four hundred patients were referred. Indication for referral was a neurologic event in 46.2%, indefinite symptom in 27.9% and asymptomatic stenosis or carotid bruit in 25.9%. Most patients underwent carotid ultrasound as first imaging (93.7%). Almost half of the patients were operated (n=176). The 30-day combined stroke and death rate was 6.5%. Appropriate indication for CEA was found in 31.6%. Over half (57.0%) of the indications were considered uncertain and 11.4% inappropriate, most of these patients having asymptomatic stenosis. CONCLUSION: Patients with asymptomatic stenosis underwent CEA relatively often and the proportion of inappropriate indications was too high. Evaluation of indications and perioperative complications is highly important in carotid surgery. PMID- 15663277 TI - The surgical treatment of abdominal aortic aneurysms in patients with light to moderate renal dysfunction. AB - OBJECTIVE: The incremental surgical risk caused by mild or moderate renal dysfunction in patients undergoing surgery for an abdominal aortic aneurysm is not well defined. The aim of this study is to analyze the prognostic significance of mild or moderate renal dysfunction in 79 patients who have undergone repair of an infrarenal abdominal aortic aneurysm at our institution. METHODS: The postoperative results of 35 non-dialysis-dependent patients (Group I) with renal dysfunction were included and compared with a group of 44 patients (Group II) with similar characteristics of age, gender and pre-operative risk factors. All operations were performed through a standard transperitoneal median laparotomy. RESULTS: The mean operation and aortic cross-clamp times did not differ significantly between the two groups (234+/-15 min vs. 189+/-17min ; p = 0.25 and 57+/-12 min. vs. 52+/-9 min. ; p = 0.52). Early operative mortality was 5.71% in patients with renal dysfunction and did not differ significantly when compared with the other group of patients 4.5%; p = 0.083). Blood requirement was higher (p < 0.01) and the hospital stay was longer (p < 0.001) in group I. There were 18 patients in group I (51.4%) and three patients in group II (6.8%) in whom the pre operative serum creatinine level increased above 2.5 mg/dl during the postoperative period. The postoperative increase of creatinine level differed significantly in patients with pre-operative renal dysfunction (p < 0.001). There were nine patients (25.7%) in group I and four patients (9.1%) in group II who required mechanical ventilation for more than 24 hours (p = 0.025). There was no significant difference between the two groups in terms of cardiac morbidity p = 0.234). CONCLUSION: Patients with even a mild or moderate degree of renal dysfunction require close monitoring and prolonged hospital care during the postoperative period. Although satisfactory early surgical outcomes may be expected in this particular group of patients, pulmonary and renal morbidity rates are higher than in patients with normal renal function. PMID- 15663278 TI - Intravaginal slingplasty: short term results. AB - BACKGROUND: Numerous surgical techniques have been described for the treatment of vaginal vault prolapses. In 1997, a new minimally invasive procedure was introduced by Petros: the infracoccygeal sacropexy, also known as Intravaginal Slingplasty (IVS). This technique is used to place a mesh in the recto-vaginal fascia and to reinforce the uterosacral ligament by placing a polypropylene tape between the perineum and the vaginal vault. Since July 2002, we have changed our approach to the treatment of vaginal vault prolapses and now perform IVS. The aim of this study was to report our early experience and short-term results with IVS. METHODS: Prospective single-institution non-randomized trial of patients who underwent IVS. Indications, intra- and post-operative complications were recorded as well as early post-operative results. RESULTS: 34 patients with a mean age of 60+/-13 years, were operated during a 12-month period. Surgical indications included rectoceles (n = 27), enteroceles (n = 26), cystoceles (n = 15) and hysteroceles (n = 9). 85% of the patients (n = 29) had more than one prolapse. Mean operative time was 63+/-19 minutes, with a 0% intra-operative complication rate. Post-operative complication rate was 2.9%: bleeding from an internal haemorrhoid required surgical haemostasis. Median post-operative stay was 3 days (range: 2-7 days). There was also one post operative complication (2.9%, a mesh erosion). Recurrence rate was 8.8% (two cystoceles and one rectocele recurred after surgery). CONCLUSION: Posterior IVS provides a safe and efficacious treatment for posterior vaginal vault prolapses. Long-term results are required to assess the functional results and recurrence rate of the technique. PMID- 15663279 TI - Laparoscopic intraperitoneal repair of incisional and ventral hernias. AB - BACKGROUND: After reports in the literature on the safety and feasibility of laparoscopic repair of ventral hernia, and with the potential advantages of the minimal invasive approach, we started to perform this technique in 2001. This study was done to evaluate the results of our initial experience. METHODS: From March 2001 to October 2003, all patients with a ventral hernia greater than 4 cm were planned to have a laparoscopic repair. Patients were studied retrospectively, collecting data on preoperative and intraoperative variables, complications and recurrences. RESULTS: In 49 patients, out of 52 patients planned, laparoscopic repair was performed. The indication was incisional hernia in 88% and recurrence after open hernia repair in 43%. The mean hernia surface area was 86,6 cm2 and 43% had a width greater than 10 cm. There were no intraoperative complications and the mean operating time was 103 min. Postoperative complications were seen in 9 patients (18,4%). Mean hospital stay was 5,9 days. Mean follow-up was 14,3 months. Late complications were seen in three patients (6,1%). Recurrence was present in one patient (2,0%). CONCLUSIONS: Laparoscopic repair of incisional and ventral hernia is a safe alternative for open mesh repair. Further definition of indications is needed, based on the dimension and the localization of the hernia. If the omission of transabdominal wall sutures improves the postoperative course with no adverse effect on recurrence rate, will be the subject of a randomized trial we have started this year. PMID- 15663280 TI - Treatment of deep cartilage defects of the knee using autologous chondrograft transplantation and by abrasive techniques--a randomized controlled study. AB - AIM OF THE STUDY: To compare outcomes of surgical treatment of deep cartilage defects of the knee in a group of patients treated by autologous chondrograft transplantation versus patients treated by abrasive techniques. MATERIALS AND METHODS: An original method of chondrograft preparation based on cultivated autologous chondrocytes in a three-dimensional carrier - fibrin glue (Tissucol, Baxter, Austria) has been described. Preclinical tests in human cadavres and porcine models have established the possibility of chondrograft use in humans. Of the 50 patients included in the study, 25 patients (50%) underwent autologous chondrograft transplantation (group I) and 25 patients (50%) were treated using abrasive techniques according to Johnson (group II). These two groups were similar with respect to age, size of defect, depth and localization, and presence of concomitant knee injuries. The Lysholm knee and IKDC (International Knee Documentation Committee) subjective scores were used to evaluate the results. RESULTS: The preoperative value of the Lysholm knee score for patients in group I was 47.60 points; 5 months after surgery 77.20 points; and 12 months after surgery 86.48 points. The values for the Lysholm knee score for patients in group II preoperatively, 5 months postop, and 12 months postop were 52.60, 69.20, and 74.48 respectively. Results 12 months after surgery were significantly better in group I as compared to group II (p < 0.001). The preoperative value of the IKDC subjective score in group I was 41.28 points; 5 months after surgery 67.00 points; and 12 months after surgery 76.48 points. The values for the IKDC subjective score in group II preoperatively, 5 months postop, and 12 months postop were 45.00, 62.28, and 68.08 respectively. Results 12 months after surgery were significantly better in group I when compared to group II (p < 0.05). CONCLUSIONS: The results obtained in this study have confirmed the better outcome in patients treated with autologous chondrograft transplantation. This original method was found to be just as effective as abrasive techniques. We recommend its use in clinical practice. PMID- 15663281 TI - The effect of diosmin hesperidin on intestinal ischaemia--reperfusion injury. AB - AIM: The effect of Diosmin Hesperidin on intestinal ischaemia reperfusion injury was evaluated in an experimental model in rats. MATERIAL AND METHODS: Forty Spraque-Dawley rats were divided into 4 groups of (n = 10) (sham, sham + Diosmin Hesperidin, Reperfusion, Reperfusion + Diosmin Hesperidin). Diosmin Hesperidin oral gavage was administrated at a dose of 50 mg/kg to rats 14 and 2 hours before the operation and 30 minutes of ischaemia and 30 minutes of reperfusion was performed in the groups when appropriate. Ileum samples were resected for histopathological evaluation and tissue malondialdehyde (MDA) and myeloperoxidase (MPA) level determination. RESULTS: Mean mucosal injury score of IR group (4,50+/ 0,23) was significantly higher than the other groups (p < 0.05). Although mean mucosal injury score of IR + DH group was higher than sham and sham + DH groups, difference was not statistically significant (p > 0.05). Tissue MDA and MPO activities of IR group were 45,55+/-2.61 nmol/g/wet tissue and 1.68+/-0.25 U/g/wet tissue respectively and were significantly higher than the other groups (p < 0.008). Although tissue MDA and MPO activities of IR + DH group was higher than sham and sham + DH groups, differences were not statistically significant (p > 0.008). CONCLUSION: Diosmin Hesperidin seems to be effective in the prevention of intestinal reperfusion injury. PMID- 15663282 TI - Update on Spigelian hernia: diagnosis and treatment by means of two cases. AB - The rarity of Spigelian hernias and the frequent subtle clinical findings can cause an important delay in diagnosis, especially in obese patients. Furthermore it has a high risk of incarceration. When this occurs, a fast recognition and adequate treatment are necessary. Treatment can be primary suture or mesh repair. More recently, the laparoscopic approach has become more popular. We present two cases of incarcerated Spigelian hernia and we give a review of the literature, with specific attention for the anatomical features and pathogenesis. PMID- 15663283 TI - Subtotal thyroidectomy with transposition of the remnant gland to the subcutaneous space. A new alternative technique in benign thyroid disease. AB - BACKGROUND: Thyroidectomy is the preferred surgical option for the treatment of benign disease of the thyroid in a wide rage of indications. Controversy exists concerning the extent of the primary excision for optimal results. A subtotal thyroidectomy with transposition of the thyroid remnant to the subcutaneous space prevents thyroid hormone supplementation for life and laryngeal nerve damage during a re-operation. METHODS: We present the case of a 42-year-old female with nontoxic multinodular goitre who underwent subtotal thyroidectomy in which a thyroid remnant of the left upper pole was placed subcutaneously through a buttonhole incision at the junction of the left sternocleidomastoid and the pre thyroidal muscles. The remnant of thyroid nourished by the vascular pedicle of the superior pole vessels was sutured to the pre-thyroidal muscles. RESULTS: One month after operation, the remnant was palpated as a small well-limited mass without signs of inflammation and thyroid function tests were normal. A contrast enhanced computed tomography (CT) scan confirmed the subcutaneous position of the thyroid tissue without pathological signs. The scintigraphy showed uptake radioactivity by the transported thyroid remnant. One year after operation the patient was clinically euthyroid with TSH and T4 levels within normal limits. CONCLUSIONS: This report documents the feasibility and efficacy of subtotal thyroidectomy with transposition of the thyroid remnant to the subcutaneous space in multinodular goitre. PMID- 15663284 TI - Peliosis hepatis revealed by biliary colic in a patient with oral contraceptive use. AB - Peliosis hepatis is a rare condition associated with a variety of diseases and drugs. We report a patient who developed peliosis hepatis while taking oral contraceptives. Three months after withdrawal of this oral contraceptive, reduction in size of these lesions was observed. PMID- 15663285 TI - Intrathoracic aneurysm of the right subclavian artery. AB - Arterial aneurysms may occur in all arteries of the human body. Arteriosclerosis takes the first place in the aetiology of the aneurysms of the great arteries and lower extremities. We would like to present a rare atherosclerotic aneurysm in the intrathoracic region of the right subclavian artery, in which vascular continuity was re-established by resection and direct end-to-end anastomosis instead of the generally used resection + revascularization technique with vascular grafts. PMID- 15663286 TI - Antiphospholipid antibody syndrome (APS) presenting as splenic thrombosis and acute acalculous cholecystitis. AB - Antiphospholipid syndrome is an uncommon auto-immune disease presenting with various clinical manifestations that may lead to surgical intervention and sometimes even life-threatening emergencies. This syndrome presents with venous and arterial thrombosis of many organs such as liver, kidney and of the skin etc. Clinical manifestations may mimic hematological disorders and be misdiagnosed in some cases due to the complexity of the symptoms. In the present study, a 65-year old man with APS syndrome presenting with severe abdominal organ pathologies that required surgical intervention, is reported. PMID- 15663287 TI - The case of acute appendicitis and appendiceal duplication. AB - BACKGROUND: Appendiceal anomalies are extremely rare malformations that are usually found in the adult population as an incidental finding during laparotomy performed for other reasons. Abnormal development of the appendix usually takes the form of a double appendix. Accompanying intestinal, genito-urinary or vertebral malformations may be present when appendiceal duplications are detected in childhood. CASE REPORT: Presented herein is a case of perforated double appendix, which causes acute abdomen in a child, without any co-existing pathology. CONCLUSION: Appendiceal anomalies are of great practical importance and a surgeon must bear them in mind during an operation. If he overlooks them, the patient undergoing surgery may experience grave consequences. They also may be a forensic issue in cases when a second explorative laparotomy reveals 'previously removed' vermiform appendix. PMID- 15663288 TI - Spontaneous pneumoperitoneum associated with colonic pseudo-obstruction. AB - Pneumoperitoneum is almost always pathognomonic of a perforated abdominal viscus requiring urgent surgical intervention. Spontaneous or non-surgical pneumoperitoneum is a rare clinical condition arising secondary to abdominal, thoracic, gynaecologic or idiopathic causes. In addition to good clinical judgement, an important component in the management process is to rule out other causes of pneumoperitoneum by performing appropriate investigations. We describe a 60-year-old man who presented with clinical features of pseudo-obstruction, following an injury to his back which was compounded by hypokalaemia. Roentgenography revealed massive pneumoperitoneum and colonic distension. As there were no overt clinical features of peritonitis, the patient was managed conservatively with parenteral nutrition and close observation. A water-soluble contrast enema and computed tomography of the abdomen were of no help in identifying the cause of his pneumoperitoneum but were helpful in eliminating the presence of hollow viscus perforation or an obvious inflammatory focus. The aetiology of pneumoperitoneum in our patient was most likely due to dissection of air through the distended colonic wall, secondary to large bowel pseudo obstruction. The diagnosis of spontaneous or non-surgical pneumoperitoneum is one of exclusion and we stress the importance of relying on clinical parameters when managing such patients conservatively. PMID- 15663289 TI - Pneumatosis cystoides intestinalis : report of two cases. AB - We present two cases of Pneumatosis Cystoides Intestinalis (PCI) involving the small intestine: emergency operations was mandatory. At surgery, the affected ileal segments were resected. After surgery, clinical courses were uneventful. PCI is a sign, not a disease, and therefore its significance is directly related to the patient's overall clinical status. Treatment is not indicated in asymptomatic patients, whereas immediate surgical intervention is necessary in patients with abdominal catastrophes. PMID- 15663290 TI - Malignant fibrous histiocytoma and squamous carcinoma derived from a burn scar. AB - Malignant fibrous histiocytoma is the most common type of soft tissue sarcoma in late adult life. The tumour may be seen rarely in burn scars and accompanying other kinds of malignancies. We herein report a case of a tumour that derived from an old burn scar on the scalp. The histological examination revealed that the tumour contained both malignant fibrous histiocytoma and squamous carcinoma areas. Malignant transformation in a burn scar is a well-known development; therefore, a new lesion occurring within the scar should not be neglected. PMID- 15663291 TI - Food-drug interactions via human cytochrome P450 3A (CYP3A). AB - Food-drug interactions have been reported to occur in various systems in the body. The causes of these interactions are mainly divided into pharmacodynamic and pharmacokinetic processes. Among these processes, drug metabolism plays a crucial role in drug interactions. Metabolic food-drug interactions occur when a certain food alters the activity of a drug-metabolizing enzyme, leading to a modulation of the pharmacokinetics of drugs metabolized by the enzyme. A variety of interactions have been documented so far. Foods consisting of complex chemical mixtures, such as fruits, alcoholic beverages, teas, and herbs, possess the ability to inhibit or induce the activity of drug-metabolizing enzymes. According to results obtained thus far, cytochrome P450 3A4 (CYP3A4) appears to be a key enzyme in food-drug interactions. For example, interactions of grapefruit juice with felodipine and cyclosporine, red wine with cyclosporine, and St John's wort with various medicines including cyclosporine, have been demonstrated. The results indicate the requirement of dosage adjustment to maintain drug concentrations within their therapeutic windows. The CYP3A4-related interaction by food components may be related to the high level of expression of CYP3A4 in the small intestine, as well as its broad substrate specificity, as CYP3A4 is responsible for the metabolism of more than 50% of clinical pharmaceuticals. This review article summarizes the findings obtained to date concerning food-drug interactions and their clinical implications. It seems likely that more information regarding such interactions will accumulate in the future, and awareness is necessary for achieving optimal drug therapy. PMID- 15663292 TI - Enhanced oral bioavailability of paclitaxel by recombinant interleukin-2 in mice with murine Lewis lung carcinoma. AB - The effect of recombinant interleukin-2 (rIL-2) pretreatment on the pharmacokinetics of paclitaxel was investigated in the murine Lewis lung carcinoma model in C57B1/6 mice. Paclitaxel 15 mg/kg was administrated orally to mice, either alone or after 3 days pretreatment with twice daily dose of 16.5 microg rIL-2. Plasma concentrations of paclitaxel were estimated by reversed phase HPLC. Pharmacokinetic parameters were determined using MicroPharm software. Using Bailer's method, a significant difference was observed in the AUCs of paclitaxel administrated alone and with rIL-2 pretreatment (928.2 +/- 136.8 vs 2549.6 +/- 131.3 ng.h.ml(-1), p <0.0001). Pretreatment with rIL-2 resulted in a 3 fold increase in the oral bioavailability of paclitaxel without altering its elimination half-life (0.798 vs 0.747 h). This could be due to the inhibition of P-glycoprotein (P-gp) mediated transport, thus enhancing paclitaxel intestinal absorption. The combination of these two drugs could be of interest in clinical practice due to their activity in pulmonary cancer. PMID- 15663293 TI - Inhibition of trans-permethrin hydrolysis in human liver fractions by chloropyrifos oxon and carbaryl. AB - Permethrin, a pyrethroid insecticide, is one of several deployment-related chemicals that have been suggested as causative agents for Gulf War related illnesses. Hydrolysis of trans-permethrin (tPMT) is a major route of detoxication and a potential locus for interactions with chemicals with similar use patterns. This study examined the potential inhibitory effects of chlorpyrifos, carbaryl, pyridostigmine bromide and the insect repellent N,N-diethyl-m-toluamide (DEET) on tPMT hydrolysis in human liver fractions. Although chlorpyrifos was not inhibitory, its toxic metabolite, chlorpyrifos oxon, strongly and irreversibly inhibited tPMT hydrolysis at low concentrations (cytosolic and microsomal Ki values of 3 and 16 nM, respectively). Carbaryl, a known anticholinesterase agent, showed non-competitive inhibition kinetics, with Ki values two orders of magnitude higher than those for chlorpyrifos oxon. Although DEET was much less effective than either chlorpyrifos oxon or carbaryl, equimolar concentrations inhibited up to 45% of tPMT hydrolysis. Pyridostigmine bromide showed no inhibitory effects. This study suggests that interaction potential between organophosphorus and pyrethroid insecticides should be considered in safety assessments when both insecticides are deployed simultaneously. PMID- 15663294 TI - CYP1A2 in a smoking and a non-smoking population; correlation of urinary and salivary phenotypic ratios. AB - The use of caffeine as a probe for CYP1A2 phenotyping has been extensively investigated over the last 25 years. Numerous metabolic ratios have been employed and various biological fluids analysed for caffeine and its metabolites. These investigations have used non-smoking, smoking and numerous disease populations to investigate the role of CYP1A2 in possible disease aetiology and for induction and inhibition studies in vivo using dietary, environmental and pharmaceutical compounds. This investigation found that the 17X/137X CYP1A2 metabolic ratio in a 5 h saliva sample and 0-5 h urine collection was not normally distributed in both a non-smoking and a smoking population. The urinary and salivary CYP1A2 metabolic ratio was log normally distributed in the non-smoking population but the smoking population showed a bi- (or tri-)modal distribution on log transformation of both the urinary and salivary CYP1A2 metabolic ratios. The CYP1A2 metabolic ratios were significantly higher in the smoking population compared to the non-smoking population when both the urinary and salivary CYP1A2 metabolic ratios were analysed. These results indicate that urinary flow rate was not a factor in the variation in CYP1A2 phenotype in the non-smoking and smoking populations studied here. The increased CYP1A2 activity in the smoking population was probably due to induction of the CYP1A2 gene via the Ah receptor causing an increase in the concentration of CYP1A2 protein. PMID- 15663295 TI - Effects of Cassia auriculata and Cardospermum halicacabum teas on the steady state blood levels of theophylline in rats. AB - The effects of concurrent administration of herbal tea prepared from dried flowers of Cassia auriculata or aerial parts of Cardospermum halicacabum and steady state serum levels of theophylline was investigated in Wistar rats. Results obtained demonstrate that a significant increase in the steady state levels of theophylline occur when this drug is administered concurrently with herbal tea prepared from either of the above plants. C. auriculata and C. halicacabum enhanced the steady state levels of theophylline by 32.5% (p < 0.02) and 48.2% (p < 0.02), respectively, when compared with the levels in animals receiving theophylline alone for the same time period. Herbal teas prepared from C. auriculata or C. halicacabum should therefore be avoided by patients treated with theophylline as these herbal teas have the potential to influence the bioavailability of the prescription drug. PMID- 15663296 TI - A pharmacokinetic interaction study between omeprazole and the H2-receptor antagonist ranitidine. AB - The effect of ranitidine pretreatment on the pharmacokinetics of omeprazole was investigated in 14 male human volunteers. Omeprazole (40 mg, gastroresistant pellets) was administered to the volunteers in a two-treatment study design, either alone or after 5 days pretreatment with b.i.d. doses of 150 mg ranitidine. Plasma concentrations of omeprazole were determined over a 24-hour period following drug administration, by a validated RP-HPLC method. Pharmacokinetic parameters were calculated with compartmental and non-compartmental analysis, using the computer program Kinetica (Inna Phase). In the two periods of treatments, the mean peak plasma concentrations Cmax were 730.8 ng/ml for omeprazole alone and 802.1 ng/ml for omeprazole co-administered with ranitidine (not significant). The time taken to reach the peak, Tmax, was 1.29 h and 1.42 h, respectively (not significant). The areas under the curve (AUC0-10) were 1,453.3 ng.h/ml and 1,736.8 ng.h/ml for the two periods of treatment; thus a greater AUC was obtained after pretreatment with multiple doses of ranitidine. Our data show that the pharmacokinetics of omeprazole might be inhibited by pretreatment with ranitidine; however, the clinical relevance of this interaction still has to be confirmed. PMID- 15663297 TI - The characterization and distribution of inorganic chemicals in tributary waters of the lower Athabasca river, Oilsands region, Canada. AB - At present, there are two large industrial plants recovering oil from the lower Athabasca oil sands area and there are plans for several more mines in the area. There are environmental concerns for aquatic life in areas downstream of current and future oil sands activities. To assess and predict potential impacts of industrial activities, it is important to separate impacts from those produced by naturally occurring oil sands deposit. Studies were therefore conducted to determine whether the water quality of tributaries to the Athabasca River, which have not been impacted by anthropogenic activities, is affected by inorganic constituents resulting from flowing through reaches with natural oilsands deposit. Three tributaries, Steepbank River, Mackay River, and Ells River at upstream and downstream locations on each stream were investigated during four surveys from 1998 to 2000. In addition to some physical parameters such as pH, conductance and hardness and the major ions (calcium, magnesium, sodium, potassium, bicarbonate, chloride, sulfate, and silicates), seventeen trace metals were investigated. Some of these metals, especially iron and manganese, were of high concentrations and in some instances, particularly in a survey conducted during the spring freshets in April 1999, exceeded guidelines for the protection of aquatic life. The observed concentrations of metals seem to be of natural origin and can be used as base-line data for future assessment of anthropogenic activities in the oil sand region. PMID- 15663298 TI - Polycyclic aromatic hydrocarbons in surface soils of Novi Sad and bank sediment of the Danube River. AB - Concentrations of 16 EPA polycyclic aromatic hydrocarbons (PAHs) in the surface soil samples of Novi Sad city urban area and in the Danube bank sediment sample were measured in 2001. The sampling sites were chosen in order to cover the whole city area and the results correspond to a network of six representative sampling sites. The total content of PAHs ranged from 307 to 1452 microg/kg with arithmetic mean value of 667 microg/kg and median of 382 microg/kg that represented typical PAH level of soils throughout Novi Sad urban area. Value of total PAHs content obtained in bank sediment sample along the Danube River was 975 microg/kg. Benzo(a)pyrene known to be carcinogenic formed 6-14% of the sum concentration of the identified PAHs for all investigated samples. Data were compared with the ones found for soils and river sediments throughout the world, and with target values set by Dutch authorities for unpolluted soil. The total carcinogenic potency for each sampling site was calculated and compared with the ones obtained on the base of the target concentrations. It was revealed only one sampling site with carcinogenic potency below the value calculated for unpolluted soil. PMID- 15663299 TI - Clophen A60 composition and content of CBs, CNs, CDFs, and CDDs after 2D-HPLC, HRGC/LRMS, and HRGC/HRMS separation and quantification. AB - Chlorobiphenyl (CB) and by-side chlorodibenzofuran (CDF), chlorodibenzo-p-dioxin (CDD), and chloronaphthalene (CN) homologue group and congener composition, and concentrations have been examined in technical CBs mixture Clophen A60. 101 peaks representing 116 CB congeners were quantified in Clophen A60, and most contributing were CBs nos. 138, 153,134/144/149, and 180 with 15.4, 12.3,8.2, and 6.5%, respectively. Di-, tri-, tetra-, penta-, hexa-, hepta-, and octa-CBs constituted, respectively, 0.03, 0.03, 0.32, 9.9, 52.7, and 4.9% of chlorobiphenyls content of the Clophen A60, while mono-, nona-, and decaCB were not quantified. Tetra- to octaCDDs were absent in Clophen A60 at concentration above the method limit of quantification of <0.01 microg/g, while the total CDFs and CNs were found at 12 and 42 microg/g, respectively. Tetra-, penta-, hexa-, and hepta-CDF with 27, 46, 19, and 7% contribution, respectively, dominated in homologue group profile of CDFs, while octa-CDF was absent in Clophen A60. In compositional profile of CDF congeners the most abundant were 1,2,4,7,8-PeCDF, 2,3,4,7,8-PeCDF, and 1,2,3,4,7,8-HxCDF and each had >5% contribution. Amongst the CN homologue groups the profile in descending order followed by hepta-, hexa-, octa-, and penta-CNs with 56, 26, 16, and 2%, respectively. In compositional profile of CNs the most abundant were the congeners such as 1,2,3,4,5,6,7-HpCN (no. 73), 1,2,3,4,6,7-/1,2,3,5,6,7-HxCN (nos. 66/67), and 1,2,3,4,5,6,7,8-OcCN (no. 75) with 55, 21, and 16%, respectively. In terms of dioxin-like toxicity of Clophen A60 the contribution from planar non-ortho and mono-ortho CBs, CDFs, and CNs was 5280, 594.5, and 33.1 ng TCDD TEQ/g, respectively, and the total TEQ of planar analogues was 5908 ng/g. PMID- 15663300 TI - By-side impurities in chloronaphthalene mixtures of the Halowax series: all 135 chlorodibenzofurans. AB - One hundred twenty five congeners of CDF off 135 theoretically possible were quantified in seven of various type Halowax formulations but still 45 co-eluted under the gas chromatographic conditions applied. The total CDFs concentration of Halowax formulations was between 250 and 16,000 ng/g. The compositional profile of CDF homologue groups of the Halowaxes frequently followed a degree of chlorination of the parent chloronaphthalene mixture and an exception was Halowax 1031. The compositional profile of many isomers of trichlorodibenzofuran, found and often highly abundant, was characteristic to majority of the formulations examined, and each of them demonstrated to have its own pattern but mysterious to explain remained Halowax 1031. In term of dioxin-like toxicity the most potent due to CDFs content was Halowax 1014 with 210 ng TCDD TEQ/g and next was Halowax 1013 with 36 ng TEQ/g, while between 1.3 and 5.0 ng TEQ/g were for other formulations. A tentative estimation made implies that the net total CDFs production due to manufacture of the technical CNs in the XX century could reach an amount between 420 kg (median) and 825 kg (mean), and for most toxic dioxin like congeners between 705 g (median) and 5700 g (mean) TCDD TEQ, while for the worst case of Halowax 1014 alone scenario it could be 750 kg of the total CDFs and 16 kg of TCDD TEQ for most toxic congeners. PMID- 15663301 TI - By-side impurities in chloronaphthalene mixtures of the Halowax series: all 75 chlorodibenzo-p-dioxins. AB - A by-side chlorodibenzo-p-dioxins (CDDs) has been identified as impurity in concentration between 1.5 and 370 ng/g in the Halowax formulations of all type. Halowax 1014 was relatively richer in number of CDD congeners detected when compared to six other CN formulations examined. Amongst the mono- to tri-CDDs, the most prevalent in the Halowaxes were 1- and 2-MoCDD, and especially they were abundant in the formulations of a lower than a higher degree of chlorination. Amongst the tetra- to octaCDD only 1,2,3,4-/1,2,4,6-/1,2,4,9-/ 1,2,3,8-TeCDD, 1,2,3,4,6,7,8-HpCDD and OcCDD were found in all the Halowaxes, and 1,2,3,4,6,7,9 HpCDD remained undetected only in Halowax 1099 and 1013, while most of TeCDDs, PeCDD, and HxCDDs were absent in a majority of the formulations examined. The compositional profile of 1,2,3,4-/1,2,4,6-/1,2,4,9,-/1,2,3,8-TeCDD and OcCDD congeners found in the Halowaxes seem to indicate, that after an initial in situ formation of mono- and di-CDDs during CNs synthesis, a further increase of reaction time, temperature, and pressure can lead to successive chlorination of the already established chlorodibenzo-p-dioxin molecule, and so to enrichment in 1,2,3,4-/ 1,2,4,6-/1,2,4,9,-/1,2,3,8-TeCDD but also OcCDD content for most of the final products obtained. Nevertheless, also due to the co-synthesis of chlorophenols in the Halowaxes, their condensation reactions could also contribute to the formation of CDDs. In term of dioxin-like toxicity the most potent due to CDDs content was Halowax 1014 with 0.95 ng TCDD TEQ/g, and between 0.00068 and 0.058 ng/g were for other formulations. A rough estimation made implies that a net CDDs production due to manufacture of the technical CNs in the XX century could reach an amount between 3.0 and 12.6 kg, while for most toxic dioxin-like constituents between 5.25 and 24 g TCDD TEQ, For the worst case scenario and involvement of Halowax 1014 only the net total CDDs production was estimated to be 1.5 kg, and for highly-toxic congeners 71 g TCDD TEQ. All these figures are much lower when compared to co-production of CDFs. PMID- 15663302 TI - Origins and mobility of phosphorus forms in the sediments of Lakes Taihu and Chaohu, China. AB - Phosphorus release from the sediment is generally an important factor for lake eutrophication. We have investigated phosphorus forms in surface sediments from Lake Taihu and Lake Chaohu by a chemical extraction method. The results showed that the concentrations of Fe/Al-bound phosphorus (Fe/Al-P) and organic phosphorus (Org-P) had significant correlations with those of amorphous Fe/Al oxides and organic matter in the sediments. Furthermore, Ca-bound phosphorus (Ca P) concentrations were well related to total organic carbon (TOC) concentrations. The results also indicated that higher pH weakened the association of phosphates with the Fe/Al hydroxides and then resulted in Fe/Al-P release. The increase of pH was companied with autogenetic progresses of calcium carbonate and then elevated Ca-P concentrations in the sediments. In conclusion, external P loadings determine spatial distribution of P fractions, especially Fe/Al-P and Org-P, and partially contribute to additional Ca-P fractions in these highly human-disturbed lakes. The pH-induced phosphorus mobility in the sediments is very important for these lakes. PMID- 15663303 TI - Sorption and biodegradation of phthalic acid esters in freshwater sediments. AB - Pathalic acid esters (PAEs) have been used as plasticizers in many products so that they could enter the aquatic environment. Three freshwater sediments in Taiwan were selected to explore the sorption and biodegradation processes of PAEs. Results indicated that di-n-butyl phthalate (DBP) and di-2-ethylhexyl phthalate (DEHP) are the only detectable PAEs which ranged in 24.9-68.3 mg/kg and 39.1-71.9 mg/kg in this study. The adsorption capacities of DBP and DEHP were identified by Kf values of the nonlinear Freundlich model associated with R2 values more than 0.90. This is expected that partition is the main mechanism controlling the transfer of PAEs between water and sediment phases. After 30 days contact, much of DBP and DEHP were removed immediately within 1 day in the desorption process. Because microorganism is as the major routes of breakdown of PAEs in the environment, much lower degraded amounts of DBP and DEHP occurred in the sterilized sediment than those in the unsterilized sediments. PMID- 15663304 TI - Phytoaccumulation of lead by sunflower (Helianthus annuus), tobacco (Nicotiana tabacum), and vetiver (Vetiveria zizanioides). AB - The ability of three plant species: Helianthus annuus, Nicotiana tabacum, and Vetiveria zizanioides for phytoaccumulation of Pb was studied. Plants were grown in hydroponic solution containing Pb(NO3)2 at concentration of 0.25 and 2.5 mM Pb in the presence or absence of chelating agents (EDTA or DTPA). Lead (Pb) transport and localization within the tissues of the plant species was determined using scanning electron microscope equipped with energy dispersive X-ray spectrometers (SEM-EDS). The addition of chelators increased Pb uptake as compared to plants grown in solution containing Pb alone. Lead taken up by the plant species were concentrated in both leaf and stem at the region of vascular bundles with greater amounts in the leaf portion. Lead granules were also found in the H. annuus root tissue from the epidermis layer to the central axis. After four weeks of growth a 23-fold increase in shoot Pb content for H. annuus and N. tabacum and 17-fold increase in shoot Pb for V. zizanioides resulted from plants grown in the 2.5 mM Pb-EDTA treatment. The higher Pb treatment (2.5 mM Pb containing EDTA) resulted in higher concentrations of Pb in plant tissue at the fourth week of exposure as compared to Pb treatment containing DTPA. Overall, Pb accumulation potential of H. annuus was greater than that of N. tabacum and V. zizanioides as indicated by the bioconcentration factor (171, 70, and 88, respectively). The highest measured Pb concentrations were found in H. annuus roots, stems, and leaves (2668, 843, and 3611 microg/g DW, respectively) grown in the 2.5 mM Pb-EDTA treatment. The addition of chelators caused some reduction in plant growth and biomass. Results showed that the three plant species tested have potential for use in phytoaccumulation of Pb since the Pb was concentrated in leaf and stem as compared to control plants. H. annuus however best meet the prerequisites for a hyperaccumulator plant and would have the potential for use in the restoration of abandoned mines and factories sites contaminated with elevated Pb levels in the soil. PMID- 15663305 TI - Characterization of odors from limed biosolids treated with nitrate and anthraquinone. AB - Complaints from the public due to odor emissions are one of the biggest problems associated with any biosolids land application program. Chemical additives to reduce or mask odors are one option for producers; however, many chemicals are too expensive or are too unstable to use safely. This project provides a preliminary evaluation of nitrate or nitrate + anthraquinone as additives in controlling odors from limed biosolids. Over a twenty-four day period, odors were measured in the headspace over several treatment levels using two different chemical analysis tools along with olfactometric evaluation of odor intensity and hedonic tone. On six days during the sample period, hydrogen sulfide was measured using a Jerome 631X, a sensor that also responds to other reduce sulfur gases. Other specific sulfides, amines, and mercaptans were also determined using solid phase microextraction with gas chromatography-mass spectrometry. A simple sniff test approach was used with six panelists on five days during the project. The chemical analysis results revealed that the addition of nitrate and especially nitrate + anthraquinone was effective in reducing concentrations of hydrogen sulfide and methylmercaptan when compared to untreated limed biosolids. However, the olfactometric results did not reveal any significant differences between treatments. The panelists also found that all treatments exhibited a fishy or ammonical character, indicative of amines, or ammonia. More advance olfactometric analysis utilizing dilution techniques might have been able to distinguish between treatments, but it is likely that amines were the dominant odorant released from all treatments. This preliminary project suggests that chemical addition of nitrate or nitrate + anthraquinone would be most effective in controlling odors from unlimed biosolids such as anaerobically digested materials. PMID- 15663306 TI - Effect of Nannochloris sp. on the toxicity of four algae. AB - The effect of Nannochloris sp. on the toxicity of cultures of four algae was evaluated using a Microtox 500 that measures the effect on a light-producing bacterium, Vibrio fischeri. Cultures of four algae produced a toxic effect, but in the presence of Nannochloris sp., the effect was reversed, and a stimulatory effect was observed. The effect was tested for Microcystis aeroginosa, Cyclotella menengheniana, Scenedesmus dimorphis, and Lyngbya sp. using cultures obtained from the University of Texas Culture Collection of Algae. PMID- 15663307 TI - Levels of biochemical constituents of fish associated with water dispersed fractions of used automobile lubricants. AB - The effects of sublethal concentrations of water-dispersed fractions of used automobile lubricant (Mobil oil SAE 50) on some biochemical constituents in fish were investigated under laboratory conditions. The fish exposed was Tilapia zilli (mean weight: 7.15+/-0.01 g), the sublethal concentrations were 125.00, 62.50, 31.25, 15.63, 7.81, and 0.00 ppm and the exposure period was 10 weeks. There was no mortality in the exposed fish throughout the exposure period; however, the biochemical constituents investigated revealed that the various concentrations led to physiological impairment in the fish, which was directly proportional to concentration and exposure period. Severely reduced feed intake was observed in fish exposed to concentrations of 62.50 and 125.00 ppm. Depletion in protein and lipid reserves, anemia and hyperglycemia were observed in all exposed fish. The implications of water dispersed fractions of used automobile lubricants in the aquatic environment and their effects on fish biodiversity are discussed. PMID- 15663308 TI - Adsorption of anionic surfactant on alumina and reuse of the surfactant-modified alumina for the removal of crystal violet from aquatic environment. AB - The adsorption characteristics of sodium dodecyl sulfate (SDS), an anionic surfactant on neutral alumina were studied in detail. Alumina was found to be an efficient adsorbent for SDS and could be used for the removal of SDS from its highly concentrated (several thousand ppm) solution. The equilibrium time found was 2 h. Though the removal efficiency was low (approximately 65%) at neutral pH, but in slightly acidic condition and in the presence of NaCl the efficiency could be increased dramatically (up to >98%). The adsorption isotherm study showed distinct four regions. The effects of various other parameters such as adsorbent dose, time, and the presence of different ions (Cl-, NO3-, SO4(2-), and Fe3+), and nonionic surfactant on the SDS adsorption were also studied. It was observed that the adsorption capacity was increased due to the presence of these ions in general. After the adsorption of SDS on alumina, the surfactant-modified alumina (SMA) was used for the removal of crystal violet (CV), a well-known cationic dye from aquatic environment. The kinetic studies showed that 1 h shaking time was sufficient to achieve the equilibrium. The removal of CV followed the second order kinetics. Studies were conducted to see the effects of adsorbent dose and initial CV concentration on the removal of CV using SMA. The pH was maintained at 6.7+/-0.1. SMA was found to be very efficient, and approximately 99% efficiency could be achieved under optimised conditions for the removal of CV when present even at a high concentration (200 ppm). To test whether the removal of CV was possible from real water using SMA, the adsorption study was conducted using CV spiked samples using distilled water, tap water, and synthetically prepared wastewater. It was interesting to note that the removal efficiency was even better for tap water and much better for wastewater when compared to that using distilled water. Desorption of both SDS and CV from the SMA surface was possible using 1 M sodium hydroxide solutions. PMID- 15663309 TI - Path concentration distribution of toluene using remote sensing FTIR and one dimensional reconstruction method. AB - A one-dimensional mapping technique coupled with Open-Path remote sensing FTIR was presented in this article. This technique was applied to one of the air toxic volatile organic compounds (VOCs)-toluene. The Path Integrated Concentrations (PICs) of toluene in different path lengths along one beam path were fitted by polynomial fitting method with degrees from 3 to 7. The 6th degree polynomial fitting showed the best fitting result. Moreover, the methods of reconstructing path concentration distribution along the beam path by applying a Gaussian model and the derivative of 6th degree polynomial fitting function were established in this article. The reconstructed concentrations of toluene along the beam path by the two methods were experimentally close. Results showed that the concentration peaks were at 1.05 and 2.40 m from the instrument by applying the Gaussian model with the maximum concentration of 0.85 and 3.19 ppm, and at 0.99 and 2.49 m from the instrument with the maximum concentration of 0.78 and 2.80 ppm by applying the 6th polynomial fitting function. It was obvious that the reconstruction results by these two methods were very close. This approach could be appreciated for its fast calculation, exact peak location orientation and concentration flow tendency mapping. In can be concluded that this method can provide the path concentration distributions of much more releasing gases in a briefly and intuitionistic way. The remote sensing FTIR coupled with these mathematical reconstruction techniques can be applied to the real world environmental and industrial hygiene monitoring, thus works as an alert system for the VOCs pollution. PMID- 15663310 TI - Recovering industrial sludge-derived slag as fine aggregate. AB - This study presents the result of using melting to recover both industrial sludge slag (the main constituent of which is calcium fluoride) and water works sludge slag as fine aggregate in cement. The main characteristics of both slag and cement mortars were measured to evaluate the feasibility of using slag as aggregate. In this study, the slag replacement ratios were 0, 10, 20, 30, 40, and 50% (w/w), and the curing periods were 7, 28, and 90 days. Slag quality was determined according to the standards of fine aggregates in the ASTM specifications, and cement mortars with various slag replacement ratios were evaluated based on their compressive strength, and Toxicity Characteristic Leaching Procedure (TCLP). The crushed slag produced in this study met the ASTM standards for fine aggregate, including gravity, unit weight, absorption, and grading, and the TCLP leached concentrations are far below existing limits, establishing the safety and suitability of slag as fine aggregate. The TCLP leached concentrations of slag and cement mortar were not significantly related to the replacement ratio, and declined with increasing curing period, revealing that the hydration strongly influenced metal leaching. The compressive strength test results of the cement mortars demonstrated that the optimal replacement ratio for maximizing compressive strength was 40%. This study also discussed the effects of replacement ratio and curing periods on cement mortars. PMID- 15663311 TI - A study for the thermal treatment of dehydrated sewage sludge with gas-agitated double screw type dryer. AB - A specially designed dryer has been developed for the thermal treatment of dehydrated, highly viscous sewage sludge with moisture content more than 80 wt.% by an gas-agitated, double-screw type dryer system. The treatment capacity of sludge dryer was about 100 kg/h. It consists of burner, feeding hopper, hot gas supplying ducts with double screw conveyor, damper and outlets of drying gas and dried material. The sewage sludge was transported by the revolution of the cylinder conveyor together with the tumbling and mixing action of the screw and lifters. The heating of the sludge was made efficient by the combination action of conduction and convection modes together with the gas-agitation process. The conduction occurred across the surface of the combustor cylinder, the convection was made by the flue gas interaction over the sludge and the agitation action through holes of conveyor cylinder into the bottom of the sludge. The number and location of gas-agitation holes and thereby the fraction of drying gas into the agitation holes were evaluated by numerical calculation of turbulent reacting flow. To evaluate the performance of the dryer developed in this study, a series of parametric experiments were performed in terms of important variables. In general drying process occurred successfully even for the highly agglomerating municipal sewage sludge through the first lumped plastic phase to a fine granulate state when the heat and mass balance was matched by adjusting rpm of the conveyor, burner capacity, and the fraction of gas-agitation. Further the avoidance of initial sludge agglomeration was considered to be important by the combination action of screw and lifter. The weight of the sludge was reduced approximately by 60% and therefore the volume of sludge was decreased to approximately 75% of initial volume with the water content of 10-20%. Energy efficiency of dryer was evaluated in the range of 70-75% at the sludge feed rate, 100 kg/h. The results obtained show the development of a robust, drying action applicable to highly viscous and high moisture content sewage sludge by the novel design of conveyor system and gas-agitation process developed in this study. PMID- 15663312 TI - Determination of the optimal and economical biofilter depth in an anaerobic hybrid reactor for treating livestock industrial wastewater. AB - A laboratory scale anaerobic hybrid reactor (AHR) being comprised of UASB and anaerobic biofilter was used to study the different biofilter depth with corresponding different critical HRT (1) and HRT (2) in treating livestock industrial wastewater, where the critical HRT (1) was defined as the HRT which resulted in the total COD in the effluent of the AHR exceeding in the influent at unsteady state, and the HRT (2) was defined as the HRT which resulted in the total COD in the effluent of the AHR exceeding the quality standard for discharging to sewer (in Hong Kong is Total COD < 1000 mg L1) at steady state. Two formulas expressing the relationship of different biofilter depth in the AHR and it's critical HRT (1) and HRT (2) in treating livestock industrial wastewater were obtained and used to design optimal and economical biofilter depth in a full scale AHR for treating livestock industrial wastewater on site. The performance of the full scale AHR indicated that using the experience formulas to design the optimal and economical biofilter depth in the full scale AHR for treating livestock industrial wastewater was successful. Experimental results also showed that the biofilter in the full scale AHR had not only physical filtration effect, but also had powerful biodegradation capability, furthermore, at unsteady state, the effect of biofilter on pollutant degrading in the full scale AHR was more great. PMID- 15663313 TI - Evaluation of a novel integrated bioreactor--AOS system for treating oil containing restaurant wastewater on site in Hong Kong. AB - AOS system is an anaerobic digestion reactor (A-section, UASB or UBF), oxidation degradation reactor (O-section, aerobic biofilm reactor), and physical sedimentation tank (S-section) 3-in-1 integrated reactor. The compact bioreactor was applied to treat oil-containing restaurant's wastewater on site in Hong Kong. The treatment efficiency was observed at different HRT. Experimental results indicated that the AOS system for treating oil-containing restaurant wastewater on site was feasible when HRT at 0.6 days above. During the running period, the net sludge generation rate of the AOS system was only about 0.08-0.09 g g( 1)removed COD, and the utmost COD specific volumetric removal rate (SVRR) of the system reached 2.77 gL(-1) d(-1). The utmost specific biomass substrate utilization rate (SBSUR) of COD in O-section and A-section reached 1.22 d(-1) and 0.128 d(-1), respectively. In addition, the experiment results showed that the AOS system with a filter in UASB (that is UBF's AOS system) could bear much more great HLR comparing with UASB's AOS and was more efficient for treating oil containing restaurant's wastewater on site. PMID- 15663314 TI - Kerry's story: the challenges of facing a recurrent craniopharyngioma. AB - Craniopharyngiomas account for approximately nine per cent of pediatric brain tumours and one to two per cent of adult brain tumours. Though considered benign, the compressive nature of these relentless tumours can produce catastrophic sequelae. This case study pays tribute to a 33-year-old woman who faced numerous surgeries, adjuvant therapy, and many of the complications associated with a craniopharyngioma. The interdisciplinary neuroscience team, along with colleagues from neuroradiology, ophthalmology, and endocrinology, joined together to care for Kerry and her family to meet their complex health and psychosocial needs. However, it was Kerry who taught us about perseverance, strength, and the power of an unconquerable human spirit. PMID- 15663315 TI - Neurodevelopmental treatment stroke rehabilitation: a critique and extension for neuroscience nursing practice. AB - The aim of this article is to review neurodevelopmental treatment (NDT) literature and existing stroke NDT nursing research, as well as explore issues related to professional collaboration in stroke rehabilitation and implications for neuroscience nursing practice. NDT or the Bobath approach is used to encourage stroke patients to use the affected side of their body in order to promote and relearn normal movement and to reduce muscle spasticity. Neuroscience nurses have an important role in facilitating stroke patients to practise transferring out of bed and performing activities of daily living outside of physiotherapy and occupational therapy sessions. Neuroscience nurses also care for stroke patients over a 24-hour perio. Therefore, it is important that nurses understand physiotherapy and occupational therapy strategies in stroke rehabilitation. PMID- 15663316 TI - Advances in MR imaging of vertebral collapse. AB - Vertebral collapse is a frequently encountered entity in clinical practice. While diagnosing acute, benign traumatic types is usually not complicated, difficulties arise in determining the etiology of vertebral collapse when there is no history of significant trauma, especially in older populations. Considering up to one third of vertebral collapse in patients with a known primary malignancy are benign, the diagnosis becomes even more difficult when there is a history of malignancy. Additionally, because primary cancer patients may be immunocompromised, the possibility of infectious vertebral collapse should also be entertained. The purpose of this article is to review all recent magnetic resonance imaging aspects of differentiating osteoporotic, malignant, and infectious vertebral collapse. PMID- 15663317 TI - Infectious spondylitis. AB - Infection of the spine is a rare but serious cause of back pain. Conventional radiographs remain the initial screening procedure. Typically two adjacent vertebral bodies and the intervening disk space are affected. Early in the course of the disease, radiographs may be normal or nondiagnostic. Magnetic resonance imaging or radionuclide bone scan will establish pathology centered in the vertebral body. Because of the ability to image soft tissues, magnetic resonance imaging is particularly helpful in detecting paravertebral and extradural abscesses. Four other conditions may mimic infectious spondylitis: degenerative disk disease associated with Modic type 1 changes, pseudoarthrosis in ankylosing spondylitis, dialysis spondyloarthropathy, and neuropathic spondyloarthropathy. Advanced imaging studies in combination with radiographs and clinical information are essential in determining the correct diagnosis. PMID- 15663318 TI - Imaging of the lumbar spine neoplasms. AB - The most common lumbar spine tumors are metastatic disease, myeloma, and hemangioma. Primary osseous lesions of the lumbar spine are unusual. When encountered, they often exhibit characteristic imaging properties, aiding the radiologist to provide a short list of differential diagnoses. We provide a discussion of imaging appearance of lumbar spine neoplasms. Emphasis of this review is on osseous lesions. Few common neurogenic intradural, extraaxial lesions are also discussed. PMID- 15663319 TI - Imaging of low back pain in children and adolescents. AB - In children with low back pain (LBP), a specific cause is often identified. LBP has a relatively high prevalence during school years. However, only a minority of the children suffering from LBP seek medical attention. Protracted back pain in childhood is a serious condition that should be thoroughly investigated. This article is a systematic review of the intrinsic causes of LBP. Imaging modalities are discussed, with emphasis on magnetic resonance imaging. We have divided the intrinsic causes of LBP into four main groups: mechanical, developmental, infectious/inflammatory, and neoplastic. Disk protrusion is prevalent in young athletes. Spondylolysis and spondylolisthesis are the most common causes of chronic LBP in children. Thoracic or thoracolumbar Scheuermann disease causes kyphosis while a lumbar localization is more painful. Childhood diskitis is associated with fever and leukocytosis. Spinal inflammatory arthritides in children include juvenile rheumatoid arthritis, the juvenile spondyloarthropathies, and SAPHO syndrome, where spine as well as sacroiliac joint changes may be seen. Cysts, tumors, tumor-like lesions, and metastases are infrequent causes of back pain in children. Several of these conditions are described and illustrated in this review of LBP in children and adolescents. PMID- 15663320 TI - Imaging of disk disease and degenerative spondylosis of the lumbar spine. AB - Low back pain is a common but poorly understood entity. Features of degeneration depend on which component of the motion segment is predominantly affected, and include disk space narrowing, vacuum phenomenon, disk desiccation, vertebral osteophyte formation, disk herniation, and facet arthrosis, but these features do not necessarily have any relationship to symptoms. Since most episodes of back pain resolve on their own, and most disk herniations spontaneously regress, imaging of low back pain, although widely performed, is probably not necessary in most cases. PMID- 15663321 TI - Imaging of the postoperative spine. AB - With the widespread prevalence of back pain, and approximately 200,000 lumbar spine operations annually in the United States, imaging of the postoperative lumbar spine has become a major diagnostic issue. The many causes of the failed back surgery syndrome make the diagnosis of the etiology of the patient's symptom complex very important to the treating physician. With the varied types of surgery, the increasing use of fusion hardware, and the myriad of complicating factors, the imaging strategy must be flexible enough to be comprehensive in the evaluation of these difficult patients. This article will address the normal and abnormal findings after lumbar spine surgery. PMID- 15663322 TI - The promise of metabolomics in cancer molecular therapeutics. AB - A systematic elucidation of cancer cell dysfunction and therapeutic mechanisms seems within reach with modern functional genomics and proteomics tools. However, for this potential to be realized, the metabolic consequences of gene expression and protein activity must be understood. 'Metabolomics' is currently a major missing component. It differs from classical metabolic studies by its greater breadth, depth and speed, enabled by the huge advances in analytical instrumentation (especially nuclear magnetic resonance and mass spectrometry) over the past two decades. Multiple metabolic pathways and networks can now be traced by the flow of atoms through metabolites, known as isotopomer analyses. Thus, metabolomics demands both high-throughput and high-information content analyses, and interpretation: the latter is currently a bottleneck. There are currently very few metabolomic studies in cancer therapeutics, despite this great need and potential. PMID- 15663323 TI - Recent advances in transcription profiling of human cancer. AB - During the past 20 years, significant success in the therapy of certain cancer types has given rise to the hope that cancer will soon be curable. However, survival rates of most patients diagnosed with cancer have not substantially increased. Instead, it is becoming clear that many tumor types, which were previously regarded as homogeneous disease entities, are composed of different subtypes which lead to varying patient prognosis and survival rates. These findings complicate diagnosis and treatment of cancer patients and call for new anti-oncogenic therapies. DNA microarrays are powerful tools to investigate global gene expression in tumors and their corresponding normal tissues, to classify tumors based on their molecular properties, and to identify novel targets for future tumor therapy. Recent results of global gene expression analyses in various tumor types are reviewed, and their implications for diagnosis and therapy of human cancer are discussed. PMID- 15663324 TI - Genomic and proteomic profiling for biomarkers and signature profiles of toxicity. AB - Toxicity profiling measures and compares all gene expression changes among biological samples after toxicant exposure. Toxicity profiling with DNA microarrays to measure all mRNA transcripts (transcriptomics), or by global separation and identification of proteins (proteomics), has led to the discovery of better descriptors of toxicity, toxicant classification and exposure monitoring than current indicators. A shared goal in transcript and proteomic profiling is the development of biomarkers and signatures of chemical toxicity. In this review, biomarkers and signature profiles are described for specific chemical toxicants that affect target organs such as liver, kidney, neural tissues, gastrointestinal tract and skeletal muscle, for specific disease models such as cancer and inflammation, and for unique chemical-protein adducts underlying cell injury. The recent introduction of toxicogenomics databases support researchers in sharing, analyzing, visualizing and mining expression data, assist the integration of transcriptomics, proteomics and toxicology datasets, and eventually will permit in silico biomarker and signature pattern discovery. PMID- 15663325 TI - Gels and more gels: probing toxicity. AB - Two-dimensional electrophoresis (2-DE) remains an important technology in the study of protein expression. In many applications, 2-DE has been supplanted by liquid chromatographic and mass spectrometric approaches that overcome some of its limitations and labor intensiveness. Nevertheless, 2-DE has exceptional relevance in toxicology and, despite the challenges, its implementation continues to support toxicologists in understanding the biological effects of chemical exposures in living systems. This review will address the use of 2-DE-based proteomics in toxicology and recent developments in this strategy, considering the unique nature of toxicity testing and the issues of dynamic range and reproducibility that have previously limited the overall utility of 2-DE in this field. PMID- 15663326 TI - Artificial neural network technologies to identify biomarkers for therapeutic intervention. AB - High-throughput technologies such as DNA/RNA microarrays, mass spectrometry and protein chips are creating unprecedented opportunities to accelerate towards the understanding of living systems and the identification of target genes and pathways for drug development and therapeutic intervention. However, the increasing volumes of data generated by molecular profiling experiments pose formidable challenges to investigate an overwhelming mass of information and turn it into predictive, deployable markers. Advanced biostatistics and machine learning methods from computer science have been applied to analyze and correlate numerical values of profiling intensities to physiological states. This article reviews the application of artificial neural networks, an information-processing tool, to the identification of sets of diagnostic/prognostic biomarkers from high throughput profiling data. PMID- 15663327 TI - p21WAF-1/Cip-1 gene therapy as an adjunct to glaucoma filtration surgery. AB - Glaucoma is a blinding eye disease characterized by elevated intraocular pressure (IOP). Glaucoma filtration surgery (GFS) is designed to reduce IOP, but wound healing responses to the procedure can result in surgical failure. Anti metabolites used in conjunction with GFS are commonly employed to control the wound healing response, but have unwanted side effects. This review describes the therapeutic potential of ocular gene therapy using an adenovirus vector containing the human p21WAF-1/Cip-1 gene (rAd-p21) to control unwanted wound healing post-GFS. Here, we summarize encouraging preclinical data in relevant models, and propose rAd-p21 gene therapy as an alternative to the currently used methods of wound healing modulation. PMID- 15663328 TI - Bacteria in the treatment of cancer. AB - Incidental observations of cancer regression following spontaneous bacterial infection have led to the preclinical development of bacteria as potential therapeutics in the treatment of cancer. A variety of natural and gene-modified bacterial species have now been explored as potential cancer treatments. The selectivity of bacterial species towards cancer will be discussed, direct bacterial oncolytic activity will be summarized and the use of bacteria as gene and/or protein delivery vehicles will be described. Preclinical and early clinical results presented indicate a high safety profile and demonstrate clear anticancer activity, thereby justifying further investigation of bacteria as a therapeutic approach in the treatment of cancer. PMID- 15663329 TI - Orphan GPCR target validation. AB - Nearly half of the identified non-olfactory G protein-coupled receptors (GPCRs) have no identified cognate ligand. These 'orphan' receptors are likely to have important physiological roles that could potentially be exploited therapeutically. However, by definition, such receptors are not immediately open for pharmacological investigation of their function. Here we summarize several strategic approaches to facilitate the discovery of orphan GPCR biology. PMID- 15663330 TI - Technology evaluation: Kahalalide F, PharmaMar. AB - Kahalalide F is a depsipeptide under development by PharmaMar as a potential treatment for solid tumors. It is currently undergoing phase II clinical trials. PMID- 15663331 TI - Technology evaluation: cantuzumab mertansine, ImmunoGen. AB - ImmunoGen is developing cantuzumab mertansine, in which the CanAg antigen targeted humanized antibody C242 is conjugated to the company's proprietary cytotoxic agent, DM1, using ImmunoGen's tumor-activated prodrug technology. Cantuzumab mertansine is undergoing phase II trials for the potential treatment of CanAg-expressing cancers, including pancreatic and colorectal cancers and non small-cell lung cancer. PMID- 15663332 TI - Technology evaluation: lumiliximab, Biogen Idec. AB - Biogen Idec (formerly IDEC Pharmaceuticals) is developing lumiliximab, a primatized anti-CD23 macaque/human chimeric antibody that inhibits the production of the IgE antibody, for the potential treatment of allergic conditions. PMID- 15663334 TI - Topical therapy for dermatophytoses: should corticosteroids be included? AB - Dermatophytoses, commonly known as ringworm or tinea, represent superficial fungal infections caused by dermatophytes, which are among the most common infections encountered in medicine. The use of corticosteroid-containing combinations in dermatophyte infections that are usually treated with topical medications is still a much-debated issue. The addition of a corticosteroid to local antifungal therapy may be of value in reducing local inflammatory reaction and thus carries the theoretical advantage of rapid symptom relief in acute dermatophyte infections associated with heavy inflammation. However, the use of such combinations requires caution as they have some potential risks, especially with long-term use under occlusive conditions. Corticosteroid-induced cutaneous adverse effects have been reported primarily in pediatric patients due to inappropriate application of these preparations on diaper areas. Additionally, the corticosteroid component may interfere with the therapeutic actions of the antifungal agent, or fungal growth may accelerate because of decreased local immunologic host reaction, such that underlying infection may persist, and dermatophytes may even acquire the ability to invade deeper tissues. Analysis of the literature documenting clinical study data and adverse reactions related to combination therapy, drew the following conclusions: (i) combination products containing a low potency nonfluorinated corticosteroid may initially be used for symptomatic inflamed lesions of tinea pedis, tinea corporis, and tinea cruris, in otherwise healthy adults with good compliance; (ii) therapy should be substituted by a pure antifungal agent once symptoms are relieved, and should never exceed 2 weeks for tinea cruris and 4 weeks for tinea pedis/corporis; and (iii) contraindications for the use of these combinations include application on diaper or other occluded areas and facial lesions, as well as in children <12 years of age and in immunosuppressed patients for any reason. PMID- 15663335 TI - Newer therapies for cutaneous sarcoidosis: the role of thalidomide and other agents. AB - Skin involvement occurs in a third of patients with sarcoidosis. The type of lesions can range from the transient erythema nodosum to the chronic facial lesion lupus pernio. For some patients with sarcoidosis, lesions on the face or elsewhere on the body may be the major or only indication for therapy. These lesions are often chronic and the use of corticosteroids may lead to more long term complications. Conventional alternatives to corticosteroids include antimalarial agents, methotrexate, and azathioprine. Recently, several drugs have been studied for chronic cutaneous sarcoidosis; thalidomide has been the most widely used. Thalidomide has been demonstrated to suppress tumor necrosis factor (TNF) release, which may be important at both the initial and chronic phases of the inflammation of sarcoidosis and appears to be crucial as part of the initial granulomatous response. Thalidomide has a different toxicity profile than corticosteroids or immunosuppressives. The usual dosage has recently been investigated in a dose-escalation trial, with the majority of patients responding to 100 mg/day. Drug toxicity has been reported in the sarcoidosis trials. The most serious adverse effect has been peripheral neuropathy, which often resolves by reducing the dose or discontinuing the medication. Other drugs that have been studied for sarcoidosis include infliximab and tetracyclines. Infliximab is a chimeric monoclonal antibody against TNF, and several published reports have shown it to be effective for the treatment of cutaneous sarcoidosis. The efficacy of tetracyclines for cutaneous sarcoidosis could be on the basis of their immunologic properties. In addition, these drugs have potent antimicrobial activity against Propionibacterium acnes; there is increasing evidence to suggest this may be one of the causes of sarcoidosis. However, most of the newer agents for cutaneous sarcoidosis have only been studied in small series. Over the next few years, it is hoped that there will be clinical trials to determine the role of each new therapy in the treatment of cutaneous sarcoidosis. PMID- 15663336 TI - Optimum treatment strategies for actinic keratosis (intraepidermal squamous cell carcinoma). AB - Actinic keratoses are superficial squamous cell carcinomas. Treatment of these lesions is indicated to prevent the cells from invading the dermis and possibly metastasizing. If a lesion exhibits evidence of possible dermal invasion, such as marked erythema, ulceration, tenderness, bleeding, and especially induration, the physician should always consider performing a biopsy. Cryosurgical destruction, the most common treatment employed, has been shown to be 98.8% effective in eliminating the lesions. Adverse reactions such as scarring, textural changes, infection, and pigmentation alteration rarely occur. Physical destruction using electrodesiccation and curettage is particularly effective when the patient has hyperkeratotic lesions. When a patient has a multitude of actinic keratoses, the use of other treatments including fluorouracil, nonsteroidal anti-inflammatory preparations, immune response modifiers, and photodynamic therapy should be considered. However, none of these treatments has proven to be as effective overall as cryosurgical destruction. If a lesion does not respond to treatment, obtaining a biopsy of the lesion should be considered to be certain that the lesion is not an invasive squamous cell carcinoma. PMID- 15663337 TI - Dermatologic treatment of cutaneous graft versus host disease. AB - Cutaneous involvement in graft versus host disease (GVHD) after allogeneic hematopoietic cell transplant can be separated into acute GVHD (aGVHD), lichenoid chronic GVHD (cGVHD) and sclerodermatous cGVHD. It seems clear that these syndromes result from different mechanisms and entail different treatment approaches. Standard treatment of cutaneous aGVHD involves the intensification of immunosuppressive therapy with adequate topical supportive management. In skin limited disease, phototherapy has shown promising results. In cutaneous cGVHD, the combination of corticosteroids and cyclosporine (ciclosporin) is the recommended therapy, and other immunosuppressants may be added depending on whether lichenoid or sclerodermatous lesions are present. High response rates to phototherapy have been found in lichenoid disease, while sclerodermatous disease responds better to etretinate or extracorporeal photochemotherapy. Localized cutaneous cGVHD may be treated with topical corticosteroids alone. Few reports on the effect of treatments in GVHD clearly describe the cutaneous involvement and the influence of the treatment on the skin. Therefore, dermatologists should be deeply involved in the diagnosis and treatment of GVHD, and good dermatologic grading systems should be developed. Theses changes will increase our knowledge of cutaneous GVHD, and relevant data in the evaluation of the effect of therapy in the disease will be obtained. PMID- 15663338 TI - Role of antifungal agents in the treatment of seborrheic dermatitis. AB - Seborrheic dermatitis is a superficial fungal disease of the skin, occurring in areas rich in sebaceous glands. It is thought that an association exists between Malassezia yeasts and seborrheic dermatitis. This may, in part, be due to an abnormal or inflammatory immune response to these yeasts. The azoles represent the largest class of antifungals used in the treatment of this disease to date. In addition to their antifungal properties, some azoles, including bifonazole, itraconazole, and ketoconazole, have demonstrated anti-inflammatory activity, which may be beneficial in alleviating symptoms. Other topical antifungal agents, such as the allylamines (terbinafine), benzylamines (butenafine), hydroxypyridones (ciclopirox), and immunomodulators (pimecrolimus and tacrolimus), have also been effective. In addition, recent studies have revealed that tea tree oil (Melaleuca oil), honey, and cinnamic acid have antifungal activity against Malassezia species, which may be of benefit in the treatment of seborrheic dermatitis. In cases where seborrheic dermatitis is widespread, the use of an oral therapy, such as ketoconazole, itraconazole, and terbinafine, may be preferred. Essentially, antifungal therapy reduces the number of yeasts on the skin, leading to an improvement in seborrheic dermatitis. With a wide availability of preparations, including creams, shampoos, and oral formulations, antifungal agents are safe and effective in the treatment of seborrheic dermatitis. PMID- 15663339 TI - Optimal management of facial telangiectasia. AB - Telangiectasia can be treated with many different modalities. Electrosurgery is effective but dependent on technique. Sclerotherapy can be used on larger blue telangiectasia but may be complicated from inadvertent injection into arterioles. Surgical phlebectomy can be used on reticular veins but is technically difficult. Many different laser wavelengths can be successfully used to minimize and/or eliminate vascular lesions on the face. Each type of laser has advantages unique to its profile of wavelength, pulse duration, spot size and cutaneous cooling. The benefits of using 532-595 nm lasers are the multiple applications of these lasers on treating not only telangiectasia, but also pigmentation and even fine wrinkles. Newer methods of cutaneous cooling and lengthening of pulse duration make these lasers less painful with fewer adverse effects (purpura) than previous lasers. The benefit in using a 1064 nm laser is that its longer wavelength can penetrate more deeply, allowing effective thermosclerosis of vessels up to 3-4 mm in diameter. In addition, the 1064 nm wavelength permits treatment of patients of skin types I-VI with or without a tan, since melanin absorption is minimal. PMID- 15663340 TI - Selecting an oral contraceptive agent for the treatment of acne in women. AB - In women, acne can be successfully treated with oral contraceptives. This article focuses on the results of clinical studies on the treatment of acne with oral contraceptives. From the literature, 12 such studies could be traced over the last 10 years. Most of the studies were multicenter, randomized, and double blind. Five studies were placebo controlled. All of these studies were analyzed and compared with each other. Statistically, all placebo-controlled studies showed a better result with the active treatment, than with the placebo. However, the placebo groups also showed an improvement in the acne lesions. In seven studies, two oral contraceptives were compared for their effect on acne vulgaris. All oral contraceptives studied showed a beneficial effect on the severity of acne. It can be concluded, based on these studies, that all oral contraceptives have a more or less beneficial effect on mild to moderate acne in women. Comparing the various clinical trials, ethinylestradiol/drospirenone is as good as ethinylestradiol/cyproterone, which is slightly better than ethinylestradiol/desogestrel and ethinylestradiol/gestodene. Ethinylestradiol/chlormadinone is slightly better than ethinylestradiol/levonorgestrel, which is as effective as ethinylestradiol/norethindrone, which is far better than the placebo. Ethinylestradiol/norgestimate is better than placebo. PMID- 15663341 TI - Topical therapy for fungal infections. AB - Fungi often infect the skin surface and subsequently invade the stratum corneum to avoid being shed from the skin surface by desquamation. Pharmacologic agents applied to the surface of the skin in the form of creams, lotions, or sprays, readily penetrate into the stratum corneum to kill the fungi (fungicidal agents), or at least render them unable to grow or divide (fungistatic agents). Thus, topical therapies work well to rid the skin of topical fungi and yeasts. Azole drugs such as miconazole, clotrimazole, and ketoconazole are fungistatic, limiting fungal growth but depending on epidermal turnover to shed the still living fungus from the skin surface. Allylamines and benzylamines such as terbinafine, naftifine, and butenafine are fungicidal, actually killing the fungal organisms. Fungicidal drugs are often preferred over fungistatic drugs for treatment of dermatophytic fungal infections, since treatment times as short as one application daily for 1 week are associated with high cure rates. Furthermore, patients often stop treatments when the skin appears healed, usually after about a week of treatment. If this short-term treatment is stopped, fungi recur more often when fungistatic, rather than fungicidal, drugs have been used. Yeast infections such as those caused by Candida albicans respond less well to allylamine drugs. The azole drugs are often preferred for these types of infections. Nail infections are difficult to cure with topical therapies because the infections usually occur under the nail instead of on top of it and products penetrate poorly, if at all, through the nail plate. Infections of hair follicles, nails, and widespread infections often require systemic treatments. Antifungal agents are compounded into many different types of vehicles. Patients often prefer to treat weeping infections with spray formulations. Most physicians prescribe branded products in cream or lotion bases. Cost is a factor dictating prescription choice, especially since most products work well regardless of mechanism of action. Cost becomes especially important when infections involve large areas of the body surface. This article reviews various treatments of cutaneous fungal infections, with special emphasis on cure rates and rationales for choosing particular products. PMID- 15663342 TI - Role of imiquimod in skin cancer treatment. AB - Cancer of the skin is by far the most common form of all cancers. Given the increasing incidence of skin cancer worldwide, it seems feasible to reconsider the treatment options available for dealing with this growing problem. Despite the lower costs associated with classical methods such as surgery and radiotherapy, immune response modifiers such as imiquimod appear to be good candidates for the future given their good cosmetic effects, the possibility of treating large areas, and the simpleness of patient-applied treatment with a cream formulation. This article reviews current literature on the use of imiquimod in the treatment of nonmelanoma and melanoma skin cancer. PMID- 15663343 TI - Acne and its management beyond the age of 35 years. AB - Although acne is not usually considered to be a disorder that affects the elderly, the disorder occurs sufficiently often in mature individuals to be noteworthy. The variety known as 'persisting acne' is, as its name suggests, ordinary acne that continues into adult life. 'Chin acne' is a curious type that occurs premenstrually in mature women, while 'sporadic acne' describes the sudden development of significant acne in later life for no apparent reason. When acne develops in an individual outside the usual susceptible age group precipitating causes such as exposure to comedogenic substances or drugs must be excluded. Similarly, endocrine causes such as androgen-secreting tumors and the administration of anabolic steroids need to be considered. All inflammatory processes are decreased in the elderly and this may be one reason for the persistence and intransigence of acne lesions in older age groups. The principles of treatment of acne in the elderly do not differ from those in other age groups, although the emphasis during counseling needs to focus on explanation and reassurance rather than prognosis. Topical retinoids and benzoyl peroxide easily irritate elderly skin, so azelaic acid and even sulfur preparations are preferable. Low-dose systemic isotretinoin is reported to be helpful to patients in this age group and is certainly worth a trial. PMID- 15663344 TI - Calcipotriol/betamethasone dipropionate: a review of its use in the treatment of psoriasis vulgaris. AB - The two-compound product containing calcipotriol 50 microg/g plus betamethasone dipropionate 0.5 mg/g (Dovobet, Daivobet) [referred to here as calcipotriol/betamethasone dipropionate], is a topical treatment for psoriasis vulgaris, combining a vitamin D analog and a corticosteroid. For most adult patients with psoriasis vulgaris on the trunk and limbs, up to 4 weeks of therapy with calcipotriol/betamethasone dipropionate provides an effective and well tolerated treatment. In clinical trials, patients with a mean baseline psoriasis area and severity index (PASI) of 9.5-10.9 experienced a mean 65.0-74.4% PASI improvement within 4 weeks, significantly better than improvements with calcipotriol 50 microg/g monotherapy, betamethasone dipropionate 0.5 mg/g monotherapy, or placebo. In addition, in 6.4%-20.1% of patients, lesions cleared. In patients who were subsequently treated with calcipotriol maintenance therapy, benefits were retained for at least 4 weeks. The safety of calcipotriol/betamethasone dipropionate in patients treated for up to 1 year was generally good; fewer than 5% of patients experienced adverse events possibly associated with long-term corticosteroid use. PMID- 15663345 TI - Pimecrolimus: a review of its use in atopic dermatitis. AB - Pimecrolimus (Elidel) is a topically active, nonsteroid, calcineurin inhibitor that has shown efficacy in controlling symptoms of atopic dermatitis in adult and pediatric patients. Topical pimecrolimus 1% cream is approved in the US for the short-term and intermittent long-term treatment of mild-to-moderate atopic dermatitis in non-immunocompromised patients aged >/=2 years who do not respond well to, or may have adverse effects with, conventional treatments. Pimecrolimus 1% cream is an effective and well tolerated treatment for atopic dermatitis in infants, children, adolescents, and adults. Pimecrolimus is effective at reducing the incidence of disease flares and, thus, the need for rescue treatment with topical corticosteroids. The drug also improves the health-related quality of life (HR-QOL) of children and adolescents, and improves the QOL of parents of children with atopic dermatitis. Furthermore, pimecrolimus does not cause skin atrophy, a problem commonly associated with topical corticosteroids, and is not associated with clinically relevant systemic adverse events. Thus, topical pimecrolimus 1% cream is an effective treatment option for the management of mild to-moderate atopic dermatitis. PMID- 15663346 TI - Management of glaucoma: focus on pharmacological therapy. AB - Glaucoma represents a major cause of vision loss throughout the world. Primary open-angle glaucoma, the most common form of glaucoma, is a chronic, progressive disease often, though not always, accompanied by elevated intraocular pressure (IOP). In this disorder, retinal ganglion cell loss and excavation of the optic nerve head produce characteristic peripheral visual field deficits. Patients with normal-tension glaucoma present with typical visual field and optic nerve head changes, without a documented history of elevated IOP. A variety of secondary causes, such as pigment dispersion syndrome and ocular trauma, can result in glaucoma as well. Treatment of all forms of glaucoma consists of reducing IOP. With proper treatment, progression of this disease can often be delayed or prevented. Treatment options for glaucoma include medications, laser therapy and incisional surgery. Laser techniques for the reduction of IOP include argon laser trabeculoplasty and selective laser trabeculoplasty. Both techniques work by increasing outflow of aqueous humour through the trabecular meshwork. Surgical options for glaucoma treatment include trabeculectomy, glaucoma drainage tube implantation and ciliary body cyclodestruction. While each of these types of procedures is effective at lowering IOP, therapy usually begins with medications. Medications lower IOP either by reducing the production or by increasing the rate of outflow of aqueous humour within the eye. Currently, there are five major classes of drugs used for the treatment of glaucoma: (i) cholinergics (acetylcholine receptor agonists); (ii) adrenoceptor agonists; (iii) carbonic anhydrase inhibitors (CAIs); (iv) beta-adrenoceptor antagonists; and (v) prostaglandin analogues (PGAs). Treatment typically begins with the selection of an agent for IOP reduction. Although beta-adrenoceptor antagonists are still commonly used by many clinicians, the PGAs are playing an increasingly important role in the first-line therapy of glaucoma. Adjunctive agents, such as alpha adrenoceptor agonists and CAIs are often effective at providing additional reduction in IOP for patients not controlled on monotherapy. As with any chronic disease, effective treatment depends on minimising the adverse effects of therapy and maximising patient compliance. The introduction of a variety of well tolerated and potent medications over the past few years now allows the clinician to choose a treatment regimen on an individual patient basis and thereby treat this disorder more effectively. PMID- 15663347 TI - Hypothyroidism in the elderly: pathophysiology, diagnosis and treatment. AB - Some degree of hypothyroidism is common in the elderly. It affects 5-20% of women and 3-8% of men. The occurrence varies with genetics with a high prevalence in Caucasians, and the disease is more common in populations with a high iodine intake. The common causes of hypothyroidism are autoimmune destruction of the thyroid gland and previous thyroid surgery or radioiodine therapy. Various types of medication, including amiodarone, cytokines and lithium, often induce hypothyroidism. Symptoms may be atypical and measurement of serum thyroid stimulating hormone (TSH) levels should be part of biochemical testing for undiagnosed medical conditions in elderly subjects. The finding of an elevated serum TSH level should be confirmed by repeated testing and supplemented with measurements of serum levels of thyroxine (T(4)) and thyroid peroxidase antibodies to verify, quantify and subclassify the abnormality. The recommended and appropriate replacement therapy for hypothyroidism is levothyroxine sodium. The initial replacement dose should be low if heart disease is suspected. Because of the long half-life of levothyroxine sodium small dosage adjustments may be performed by adding or withdrawing a tablet once or twice weekly. Levothyroxine sodium is only partly absorbed after oral ingestion, and food, minerals, drugs and tablet composition influence absorption. Studies performed a few years ago suggested that a combination of levothyroxine sodium and liothyronine may improve clinical results, but recent more comprehensive studies have not supported this hypothesis. Accordingly, liothyronine replacement is not documented to be of benefit. If liothyronine is added to replacement, the liothyronine dose should be kept low, within the physiological range and, preferably be administered twice daily. Thyroid hormone therapy has no beneficial effect above placebo in elderly individuals with normal serum TSH levels and T(4) levels. The major risk of levothyroxine sodium therapy is over-replacement, with anxiety, muscle wasting, osteoporosis and atrial fibrillation as adverse effects. Subclinical hypothyroidism with elevated serum TSH levels but T(4) levels within the laboratory reference range is a mild variant of overt hypothyroidism. Patients with subclinical hypothyroidism should be informed about the disease and offered the possibility of replacement. Only some patients treated for subclinical hypothyroidism will feel better after therapy. In elderly patients on replacement therapy, care should include estimation of serum TSH level once or twice a year, with small dosage adjustments of levothyroxine sodium to keep serum TSH level within the normal range. PMID- 15663348 TI - Managing bipolar disorder in the elderly: defining the role of the newer agents. AB - Clinical research in geriatric psychopharmacology has been a relatively neglected focus compared with the wealth of information on younger populations, and there is a dearth of published, controlled trials. Similarly, these are limited data in the area of geriatric bipolar disorder. Although there is an absence of rigorous, evidence-based information, preliminary data on older adults with bipolar disorder suggest some promising treatment options and important differences in older versus younger patients with bipolar illness. Lithium, while widely utilised in younger populations, is often poorly tolerated in the elderly. Clinical evidence regarding use of antiepileptic compounds in late-life bipolar disorder is generally compiled from bipolar disorder studies in mixed populations, studies in older adults with seizure disorders, and studies on dementia and psychotic conditions other than bipolar disorder. Valproate semisodium and carbamazepine are widely prescribed compounds in older adults with bipolar disorder. However, the popularity of these compounds has occurred in context of an absence of evidence-based data. The atypical antipsychotics have expanded the treatment armamentarium for bipolar disorder in mixed populations and may offer particular promise in management of bipolar illness in older populations as well. Olanzapine, risperidone, quetiapine, ziprasidone and aripiprazole are atypical antipsychotics that have been approved by the US FDA for the treatment of bipolar disorder; however, there are no published, controlled trials with atypical antipsychotics specific to mania in geriatric patients. Preliminary reports on the use of clozapine, risperidone, olanzapine and quetiapine suggest a role for the use of these agents in late-life bipolar disorder. Information with ziprasidone and aripiprazole specific to geriatric bipolar disorder is still lacking. PMID- 15663349 TI - Postural blood pressure changes and orthostatic hypotension in the elderly patient: impact of antihypertensive medications. AB - With age our ability to maintain haemodynamic homeostasis during position changes becomes less effective. This predisposes elderly patients to significant changes in blood pressure upon standing and orthostatic hypotension (OH). The prevalence of OH varies according to the population being studied. A range of between 5% and 60% has been reported with the lower rate in elderly individuals living in the community and higher rates in those living in an institution or in the acute-care setting. Multiple factors have been linked to OH including age, bed rest, low body mass index and medications. Although antihypertensive medications can theoretically, as a group, worsen OH, the majority of cross-sectional studies have found no association. In addition, prospective randomised trials have demonstrated an improvement in postural blood pressure (PBP) changes with antihypertensive medications. When considering the individual classes, peripheral vasodilators, specifically alpha-adrenoceptor antagonists and nondihydropyridine calcium channel antagonists, can exacerbate PBP changes and lead to OH. ACE inhibitors, angiotensin-receptor antagonists and beta-adrenoceptor antagonists with intrinsic sympathomimetic activity are less likely to worsen OH. Careful management of electrolyte disturbance can decrease the risk of developing OH with diuretic use. With the aging population, this problem will be encountered by the clinicians at a much higher rate. A detailed patient history, an accurate orthostatic blood pressure measurement and careful evaluation of the autonomic nervous system can provide clinical guidance for management of OH. In hypertensive individuals with no pre-treatment OH, the use of antihypertensive medication can be safe and lead to a low risk of developing OH. In individuals with pre-treatment OH or who develop OH while on antihypertensive medications avoidance of the classes that may exacerbate OH and a judicious use of antihypertensive classes that may improve PBP changes may be safe and adequate treatment. PMID- 15663351 TI - Rasagiline. AB - Rasagiline is a second-generation potent, irreversible and selective inhibitor of monoamine-oxidase type B (MAO-B), which has been evaluated for the treatment of Parkinson's disease. Rasagiline also possesses neuroprotective properties that are independent of its MAO inhibitory activity. Unlike selegiline, rasagiline has no amphetamine-like metabolites and its major metabolite, 1-(R)-aminoindan, has demonstrated favourable pharmacological activity in experimental studies. Rasagiline has shown significant beneficial effects as monotherapy in the treatment of early Parkinson's disease. Monotherapy with rasagiline 1 or 2 mg once daily significantly attenuated the worsening of symptoms, compared with placebo, in patients with early Parkinson's disease in a randomised, double-blind trial (n = 404). Furthermore, patients treated with rasagiline for 12 months had less functional decline than patients whose treatment was delayed for 6 months (n = 371). In patients with moderate-to-advanced disease receiving background therapy with levodopa and additional anti-parkinsonian medications (n = 1159), rasagiline 0.5 or 1 mg once daily reduced the daily 'off' time by 0.49-0.94 hours relative to that in placebo recipients in two randomised, double-blind trials. The efficacy of rasagiline 1 mg once daily was similar to entacapone 200 mg administered with each levodopa dose. Rasagiline was generally well tolerated in clinical trials as both monotherapy and when administered with other antiparkinsonian drugs. Adverse events with rasagiline were generally similar in frequency to those seen in placebo or entacapone recipients. PMID- 15663350 TI - The association of inappropriate drug use with hospitalisation and mortality: a population-based study of the very old. AB - BACKGROUND AND OBJECTIVE: Inappropriate prescribing is an important and possibly preventable risk factor for adverse drug reactions (ADRs) in the elderly, and hospital-based studies have shown that a large proportion of admissions is a result of ADRs. However, little is known about how inappropriate drug use (IDU) affects the elderly at the population level. The aim of this study was to explore possible associations of IDU with acute hospitalisation and mortality in an elderly population during 3 years of follow-up. PATIENTS AND METHODS: Data from a rural, population-based, longitudinal cohort study within the Kungsholmen Project, Sweden, were used. 785 participants, > or=75 years of age, had complete data on drug use and selected covariates collected during baseline investigation from 1995 to 1998, and were included in the study. Hospitalisation and mortality data during 3 years after inclusion were collected. IDU was assessed at baseline using consensus-based criteria applicable to available data (derived from Beers' criteria, Canadian criteria and clinical indicators of drug-related morbidity in older adults) with the addition of potentially dangerous drug duplication and additional potentially hazardous drug-drug interactions. IDU was defined as presence of at least one inappropriate drug regimen according to the study criteria. Logistic regression and proportional hazard models were used, respectively, to study the association of IDU with hospitalisation and mortality. RESULTS: Drugs were used on a regular or 'as needed' basis by 91.6% of the study population, with a mean of 4.4 drugs per person. IDU was common, with a prevalence of 18.6% and was associated with increased risk of at least one acute hospitalisation in community-living elderly, after adjustment for age, sex, education, comorbidity, dependency in activities of daily living (ADL) and smoking. The odds ratio was 2.72 (95% CI 1.64, 4.51). No association with mortality was found, after adjustment for age, sex, housing, education, comorbidity, ADL-dependency, smoking and body mass index. CONCLUSION: Polypharmacy and IDU are common among the elderly and IDU is associated with acute hospitalisation in community-living elderly. Although causality cannot be established with this study design, the results are consistent with the high prevalence of drug-related hospital admissions found in hospital-based studies. Our results indicate that it is desirable with current knowledge, to reduce IDU through information to physicians and careful prescribing. PMID- 15663354 TI - Wound-healing technologies: low-level laser and vacuum-assisted closure. PMID- 15663355 TI - Mechanisms of transformation by MLL. AB - Rearrangements of the mixed-lineage leukemia gene MLL1 (MLL, HRX, ALL1), the human homologue of the Drosophila gene trithorax, are associated with aggressive acute leukemias in both children and adults. Transformation by rearranged forms of MLL1, including in-frame fusion proteins, partial tandem duplications, and amplification of MLL1 through upregulation of Hox gene and cofactor expression apparently results in a block in hematopoietic differentiation. MLL1 regulates Hox gene expression via direct promoter binding and histone H3 Lys 4 methylation mediated by the intrinsic methyltransferase activity of the SET domain. Mll1 knockout leads to loss of Hox gene expression, defects in hematopoiesis, and embryonic lethality. A close homologue, MLL2 is amplified in some solid tumors. MLL2 also has histone H3 Lys 4 methyltransferase activity that is dependent on menin, a protein mutated in multiple neoplasia type I (MEN1) and which is required for normal Hox expression. These findings underscore the importance of the MLL histone methyltransferases in development and disease. PMID- 15663356 TI - Regulatory mechanisms operative in osteoclasts. AB - The osteoclast is hematopoietic in origin and is the primary bone-resorbing cell derived from monocyte/macrophage lineage. Tumor necrosis factor (TNF) family member, RANK ligand (RANKL) expressed on marrow stromal/osteoblast cells in response to several osteotropic factors, is critical for osteoclast precursor differentiation to form multinucleated osteoclasts, which resorb bone. M-CSF is required for proliferation, survival, and expression of receptor activator of nuclear factor kappa B (RANK) in osteoclast precursors. The interaction of RANKL RANK results in activation of various signaling cascades during osteoclast development and activation. The osteoclast is an autocrine/paracrine, intracrine regulatory cell that produces factors such as IL-6, Annexin II, TGF-beta, OIP 1/hSca, which influence its own formation and activity. In addition, integrin expression in osteoclasts mediate cell-matrix and cell-cell interactions in the bone microenvironment through specific signaling pathways resulting in cytoskeletal organization, polarization, and activation of osteoclasts to resorb bone. Recent molecular genetic studies have identified several transcription factors, such as NF-kappaB, c-Fos, MITF, and NFATc1, which are essential for osteoclast differentiation. Although a wide variety of molecules, including the reactive oxygen species (ROS) that are differentially regulated during osteoclastogenesis, the precise signal transduction pathways, and molecular mechanisms underlying the gene expression in osteoclasts, are just beginning to be defined. In this review, we discuss the molecular regulatory mechanisms operative during osteoclast differentiation, bone resorption, and survival. PMID- 15663357 TI - The regulation and regulatory activities of alternative splicing of the SMN gene. AB - Alternative splicing is an essential process that produces protein diversity in humans. It is also the cause of many complex diseases. Spinal muscular atrophy (SMA), the second most common autosomal recessive disorder, is caused by the absence of or mutations in the Survival Motor Neuron 1 (SMN1) gene, which encodes an essential protein. A nearly identical copy of the gene, SMN2, fails to compensate for the loss of SMN1 because exon 7 is alternatively spliced, producing a truncated protein, which is unstable. SMN1 and SMN2 differ by a critical C-to-T substitution at position 6 of exon 7 in SMN2 (C6U transition in mRNA). This substitution alone is enough to cause an exon 7 exclusion in SMN2. Various cis- and trans-acting factors have been shown to neutralize the inhibitory effects of C6U transition. Published reports propose models in which either abrogation of an enhancer element associated with SF2/ASF or gain of a silencer element associated with hnRNP A1 is the major cause of exon 7 exclusion in SMN2. Most recent model suggests the presence of an EXtended INhibitory ContexT (Exinct) that is formed as a consequence of C6U transition in exon 7 of SMN2. In Exinct model, several factors may affect exon 7 splicing through cooperative interactions. Such regulation may be common to many alternatively spliced exons in humans. Recent advances in our understanding of SMN gene splicing reveals multiple challenges that are specific to in vivo regulation, which we now know is intimately connected with other biological pathways. PMID- 15663358 TI - Neuroendocrine cells in prostate cancer. AB - Neuroendocrine (NE) cells are found in prostate tumors, and their incidence is considered a promising prognostic indicator for the development of androgen independent disease. NE cells are derived from non-NE prostate cancer cells and secrete factors that can act in a paracrine manner to stimulate the survival, growth, motility, and metastatic potential of prostatic carcinoma cells. Factors such as IL-6, epinephrine, and forskolin induce NE differentiation in prostate cancer cells; the mechanisms involve increases in intracellular cAMP, protein kinase A (PKA) activation and reduced intracellular calcium levels. Transcription factors implicated in the acquisition of NE characteristics by prostate cancer cells include STAT3, CREB, EGR1, c-fos, and NF-kappaB. Expression of Chromogranin A, neuron-specific enolase, bcl-2, and the androgen receptor are modulated during NE differentiation and serve as molecular markers for NE cells. Most importantly, NE cells secrete neuropeptides, such as bombesin, neurotensin, PTHrP, serotonin, and calcitonin, which trigger growth and survival responses in androgen independent prostate cancer cells. Prostate cancer cell receptors that play a role in these processes include the gastrin-releasing peptide (GRP) receptor, neurotensin receptors, and the epidermal growth-factor receptor (EGFR). Signal transduction molecules activated by these neuropeptides include Src, focal adhesion kinase (FAK), ERK, and PI3K/Akt, with subsequent activation of Elk-1, NF kappaB, and c-myc transcription factors. A multitude of genes are then expressed by prostate cancer cells, which are involved in proliferation, anti-apoptosis, migration, metastasis, and angiogenesis. Targeting of these pathways at multiple levels can be exploited to inhibit the process by which NE cells contribute to the progression of androgen-independent, treatment-refractory prostate cancer. PMID- 15663359 TI - A hierarchical approach to finite element modeling of the human spine. AB - Finite element modeling has become an important part of biomechanics research in the last 30 years. In particular, virtual experimentation on the spine bypasses several limitations on mechanical testing: any load and constraint may be applied to a finite element model, and experiments may be repeated with slight alterations on the same specimen. Because computer power is still limited, each virtual model is designed with a particular scale in mind. To analyze spinal motion, a motion segment consisting of two or more vertebrae and the connective tissue is modeled. For vertebral strength, a single vertebra, with or without posterior elements, is analyzed. To understand the biomechanics of the cancellous bone and to examine bone remodeling, the individual trabeculae comprising the cancellous bone are examined. These three areas represent the bulk of current research. Scientists also explore whole-spine mechanics and ultrastructural dynamics, but these suffer greatly from a lack of physiological data on the observed phenomena. To a large extent, the spinal physiological loading scenario and the load distribution at the different structural levels are unknown. As in vivo measurement techniques and computational power grow, so will the usefulness of finite element modeling. PMID- 15663360 TI - Regulation, regulatory activities, and function of biglycan. AB - Biglycan is a member of the small leucine repeat proteoglycan family (SLRP). The biglycan gene is located on the X chromosome. Based on the amino acid sequence, the protein core of biglycan can be divided into six distinct domains: (1) a signal sequence, (2) a propeptide region, (3) a N-terminal glycosaminoglycan attachment region, (4) a cysteine loop, followed by (5) a leucine- rich repeat region domain (that makes up over 66% of the core protein), and (6) a final cysteine loop. Biglycan has been found in almost every organ within our body, but it is not uniformly distributed within an organ. Biglycan has been shown to be expressed on the cell surface, pericellularly, and sometimes within the extracellular matrices of a range of specialized cell types within the organ. Its expression pattern has been shown to be altered by growth factors and certain pathologic conditions. The regulation of biglycan expression occurs by both transcriptional and nontranscriptional mechanisms. The currently proposed biglycan functions appear to be dependent on the particular microenvironment and on the organ in question. In this review, we will focus on gene and protein structure, localization, expression, regulation, and function. PMID- 15663361 TI - Effect of age on the immunoglobulin class switch. AB - Aging represents a complex remodelling in which both specific and innate immunity deteriorate. Age-related changes in humoral immunity involve reduced vaccine responses and increased production of auto-antibodies. Although T-cell alterations play a significant role in age-related humoral immune changes, alterations in B cells also occur. In this review, we provide an overview of age related changes in B-cell functions and markers, including transcription factors, and also discuss controversies in the field of B-cell aging. We summarize our recent results, showing that splenic B cells from senescent mice are deficient in production of secondary isotypes (IgG1, IgG2a, IgG3, IgE), class switch recombination (CSR), and expression of the transcription factor E47. We also demonstrate that there is more Id2 (a negative regulator of E47) in old activated B cells. E47 is required for CSR, at least in part, via expression of activation induced cytidine deaminase (AID). Our studies show that impaired induction of E47, and, subsequently, AID, contribute to poor CSR and production of secondary isotypes in senescence. We also present new data indicating the absence of DNA switch region excision circles for CSR in old activated B cells, confirming the location of the defect at the DNA endonucleolytic step. And, finally, we show that there is no change in NF-kappaB or Blimp-1 in old vs young stimulated B cells. PMID- 15663362 TI - Ly-49 receptors and their functions. AB - Ly-49 receptors are lectin-like type II transmembrane disulfide-bonded homodimers expressed on natural killer (NK) cells and some T-cell subsets. Cell-mediated cytotoxicity and release of cytokines/chemokines are functions regulated by Ly-49 recognition of class I major histocompatibility complex proteins (MHC-I) or virus encoded MHC-like product(s). Here we examine diversity and conservation found within the Ly-49 gene family and explore the importance of polymorphism in Ly-49 receptor expression, specificity, and function. Several parallels are evident between Ly-49 receptors in rodents and killer Ig-related (KIR) receptors in humans, including receptor gene amplification and diversification, expression patterns, MHC-I specificity, shared signaling, and ultimate effects on NK-cell functions. These similarities suggest that insights gained in defining Ly-49 receptor functions in small animal models could have direct relevance to existing clinical challenges where there may be opportunities to manipulate human NK cells and KIR receptors for therapeutic benefit. PMID- 15663363 TI - Role of the D prostanoid receptor 1 in the modulation of immune and inflammatory responses. AB - Prostaglandins (PGs) are potent eicosanoid lipid mediators derived from phospholipase-released arachidonic acid, which are involved in numerous homeostatic biological functions and inflammation. They are generated by the sequential action of cyclooxygenase isozymes and cell-specific PG synthases. Along with their role in inflammatory responses, recent accumulating evidence strongly suggests that PGs, including PGD2, are part of a complex regulatory network that modulates the immune system. PGD2 is the major prostanoid secreted by activated mast cells and has long been implicated in allergic diseases. The aim of this review is to discuss our current understanding of the mode of action of PGD2 during Th2-mediated inflammation. We also discuss recent findings, which suggest that PGD2 exerts important effects on both immune and inflammatory responses by targeting the D prostanoid receptor 1 on dendritic cells, the most potent antigen-presenting cells. PMID- 15663371 TI - Recent advances in urogynaecology. Proceedings of the joint RCOG/BSUG meeting on incontinence and prolapse, 16-19 June 2003. PMID- 15663364 TI - The impact of extracellular acidosis on dendritic cell function. AB - Dendritic cells (DCs) are the most efficient antigen-presenting cells. They are activated in the periphery by conserved pathogen molecules and by inflammatory mediators produced by a variety of cell types in response to danger signals. It is widely appreciated that inflammatory responses in peripheral tissues are usually associated with the development of acidic microenvironments. Surprisingly, there are relatively few studies directed to analyze the effect of extracellular acidosis on the immune response. We focus on the influence of extracellular acidosis on the function of immature DCs. The results presented here show that acidosis activates DCs. It increases the acquisition of extracellular antigens for MHC class I-restricted presentation and the ability of antigen-pulsed DCs to induce both specific CD8+ CTL and B-cell responses. These findings may have important implications to our understanding of the mechanisms through which DCs sense the presence of infection or inflammation in nonlymphoid tissues. PMID- 15663383 TI - Interstitial cells of the bladder: the missing link? PMID- 15663390 TI - Shifting the balance of care--or imaginative healthcare accounting? PMID- 15663391 TI - Inherited metabolic diseases and pregnancy. PMID- 15663392 TI - Microwave and thermal balloon ablation for heavy menstrual bleeding: a systematic review. AB - OBJECTIVE: To compare the effectiveness of two second generation endometrial ablation techniques (microwave and thermal balloon endometrial ablation) with first generation techniques of endometrial ablation to treat heavy menstrual bleeding in women. SEARCH STRATEGY: We searched the Cochrane Library (issue 3, 2002), the National Research Register, MEDLINE (1966 to August 2002), Embase (1980 to August 2002) and Web of Science Proceedings (all years). We also searched reference lists and contacted experts and manufacturers in the field. SELECTION CRITERIA: Randomised controlled trials and controlled trials of microwave endometrial ablation and thermal balloon endometrial ablation versus transcervical resection and rollerball ablation, alone or in combination, to treat heavy menstrual bleeding were included. DATA COLLECTION AND ANALYSIS: Two reviewers independently selected studies for inclusion and extracted data. As there was considerable clinical and methodological heterogeneity among the studies, meta-analysis was not undertaken and results are presented descriptively. RESULTS: Two randomised controlled trials of microwave endometrial ablation and eight trials (six randomised controlled trials) of thermal balloon endometrial ablation were included in the review. No significant differences were found between first and second generation techniques in terms of amenorrhoea, bleeding patterns, pre-menstrual symptoms, patient satisfaction or quality of life. Microwave endometrial ablation and thermal balloon endometrial ablation had significantly shorter operating and theatre times than first generation techniques. Adverse effects were few with all techniques, but there were fewer peri-operative adverse effects with second generation techniques. CONCLUSION: Microwave endometrial ablation and thermal balloon endometrial ablation are alternatives to first generation techniques for treating heavy menstrual bleeding. No head-to-head trials of microwave endometrial ablation and thermal balloon endometrial ablation have been undertaken and there is not yet enough evidence of differences in clinical effectiveness between these two techniques. PMID- 15663393 TI - Accuracy of fetal echocardiography in the routine detection of congenital heart disease among unselected and low risk populations: a systematic review. AB - OBJECTIVE: To determine among unselected and low risk populations the accuracy with which fetal echocardiography during the second trimester detects congenital heart disease. DESIGN: A systematic review of studies that assess the accuracy of fetal echocardiography. SETTING: District General Hospital and Tertiary referral centres. POPULATION: Women during the second trimester attending for ultrasound assessment. METHODS: General bibliographic databases (e.g. MEDLINE, EMBASE) and specialist computerised databases (e.g. Cochrane Library, National Research Register), grey literature, manual searching of reference lists of primary and review articles and personal contact with experts were used to identify studies. Studies were included if fetal echocardiography among unselected or low risk pregnant women was compared against a postnatal reference standard. Data were extracted on quality, study design and characteristics, and accuracy data to construct 2 x 2 tables. Data were synthesised qualitatively, and sensitivity and specificity with 95% confidence intervals were calculated. MAIN OUTCOME MEASURE: Sensitivity and specificity. RESULTS: Five primary studies met the inclusion criteria and comprised 60,901 subjects. One study assessed the accuracy of fetal echocardiography among an unselected population and four studies among low risk populations. All eligible studies found that fetal echocardiography helped to correctly diagnose normal fetus among unselected and low risk populations. Correct diagnosis of babies for congenital heart defects was higher among the unselected population (85%) than among the low risk populations (range from 35% to 86%); however, the potential for ascertainment bias and the choice of reference standard limits the validity of this finding. The variation in sensitivity across studies was not explainable by clinical factors such as scanning regime, operator skill and equipment. CONCLUSIONS: The evidence from this review about the accuracy of fetal echocardiography does not lend support to its routine use among unselected and low risk populations during the second trimester to detect congenital heart disease. PMID- 15663394 TI - Association between congenital toxoplasmosis and preterm birth, low birthweight and small for gestational age birth. AB - OBJECTIVE: To determine the association between congenital toxoplasmosis and preterm birth, low birthweight and small for gestational age birth. DESIGN: Multicentre prospective cohort study. SETTING: Ten European centres offering prenatal screening for toxoplasmosis. POPULATION: Deliveries after 23 weeks of gestation in 386 women with singleton pregnancies who seroconverted to toxoplasma infection before 20 weeks of gestation. Deliveries after 36 weeks in 234 women who seroconverted at 20 weeks or later, and tested positive before 37 weeks. METHODS: Comparison of infected and uninfected births, adjusted for parity and country of birth. MAIN OUTCOME MEASURES: Differences in gestational age at birth, birthweight and birthweight centile. RESULTS: Infected babies were born or delivered earlier than uninfected babies: the mean difference for seroconverters before 20 weeks was -5.4 days (95% CI: -1.4, -9.4), and at 20 weeks or more, -2.6 days (95% CI: -0.5, -4.7). Congenital infection was associated with an increased risk of preterm delivery when seroconversion occurred before 20 weeks (OR 4.71; 95% CI: 2.03, 10.9). No significant differences were detected for birthweight or birthweight centile. CONCLUSION: Babies with congenital toxoplasmosis were born earlier than uninfected babies but the mechanism leading to shorter length of gestation is unknown. Congenital infection could precipitate early delivery or prompt caesarean section or induction of delivery. We found no evidence for a significant association between congenital toxoplasmosis and reduced birthweight or small for gestational age birth. PMID- 15663395 TI - Rupture of the uterine scar during term labour: contractility or biochemistry? AB - OBJECTIVE: Vaginal birth after a prior low transverse caesarean section (VBAC) is advocated as a safe and effective method to reduce the total caesarean section rate. However, the risk of uterine rupture has dampened the enthusiasm of practising clinicians for VBAC. Uterine rupture occurs more frequently in women receiving prostaglandins in preparation for the induction of labour. We hypothesised that similar to the cervix, prostaglandins induces biochemical changes in the uterine scar favouring dissolution, predisposing the uterus to rupture at the scar of the lower segment as opposed to elsewhere. DESIGN: We tested aspects of this hypothesis by investigating the location of uterine rupture associated with prostaglandins and compared it with the sites of rupture in the absence of prostaglandins. SETTINGS: Two North American University Hospitals. POPULATION: Twenty-six women with a prior caesarean section, experiencing uterine rupture in active labour. METHODS: Retrospective review of all pregnancies complicated by uterine rupture at two North American teaching hospitals from 1991 to 2000. MAIN OUTCOME MEASURE: Site of the uterine rupture. RESULTS: Thirty-four women experienced rupture after a previous caesarean section with low transverse uterine incision. Ten of the women who ruptured (29%) received prostaglandins for cervical ripening (dinoprostone: n= 8 or misoprostol: n= 2) followed by either spontaneous contractions (n= 3) or oxytocin augmentation during labour (n= 7). In 16 women (47%), oxytocin alone was sufficient for the induction/augmentation of labour. Eight (23%) women ruptured at term before reaching the active phase of labour in the absence of pro-contractile agents or attempted VBAC. There were no differences among the groups in terms of age, body mass index, parity, gestational age, fetal weight or umbilical cord pH measurements. Women treated with prostaglandins experienced rupture at the site of their old scar more frequently than women in the oxytocin-alone group whose rupture tended to occur remote from their old scar (prostaglandins 90%vs oxytocin 44%; OR: 11.6, 95% CI: 1.2-114.3). CONCLUSION: Women in active labour treated with prostaglandins for cervical ripening appear more likely to rupture at the site of their old scar than women augmented without prostaglandins. We propose that prostaglandins induce local, biochemical modifications that weaken the scar, predisposing it to rupture. PMID- 15663396 TI - A comparison of fetal organ measurements by echo-planar magnetic resonance imaging and ultrasound. AB - OBJECTIVES: To compare fetal organ size measured using echo-planar magnetic resonance imaging and 2D ultrasound. To determine the relative accuracy with which each technique can predict fetal growth restriction. DESIGN: A cross sectional, observational study comparing two different measurement techniques against a gold standard, in a normal clinical population and an abnormal population. SETTING AND POPULATION: Seventy-four pregnant women (33 who were ultimately found to be normal and 37 with fetal growth restricted fetuses) were recruited from the City Hospital Nottingham UK to be scanned once (at various gestations). METHODS: Each fetus had a standard ultrasound biometry assessment followed by magnetic resonance imaging measurement of organ volumes. MAIN OUTCOME MEASURES: For each measurement for both techniques, the normal population was plotted with 90% confidence intervals. Fetal growth restricted subjects were compared with the normal population using this plot; 2 x 2 tables were created for each measurement. This was used to calculate the relative sensitivities and positive predictive value of the different measurements. A Bland-Altman plot was used to compare the ultrasound and magnetic resonance imaging measurements of fetal weight. RESULTS: Brain sparing was seen in ultrasonic head circumference measurements, but an overall reduction in fetal growth restriction brain volume was apparent using magnetic resonance imaging at late gestations. Across the whole range of gestational ages, ultrasound assessment of fetal weight was the best predictor of fetal growth restriction. CONCLUSION: Ultrasound fetal weight assessment appears to identify more fetuses with fetal growth restriction than abdominal circumference. The brain sparing apparent in ultrasonic head circumference measurements of fetuses with fetal growth restriction masks a reduction in brain volume observed with magnetic resonance imaging. PMID- 15663397 TI - Seroprevalence, incidence of prenatal infections and reliability of maternal history of varicella zoster virus, cytomegalovirus, herpes simplex virus and parvovirus B19 infection in South-Western Finland. AB - OBJECTIVE: To study seroprevalence and incidence and fetal transmission of varicella zoster virus (VZV), cytomegalovirus (CMV), herpes simplex virus (HSV) types 1 and 2 and parvovirus B19 infections during pregnancy and to evaluate the reliability of maternal past history of VZV, HSV and parvovirus infections. DESIGN: Prospective study of parturient women. SETTING: South-Western Finland. PARTICIPANTS: Five hundred and fifty-eight parturient women. METHODS: IgG and IgM antibodies against VZV, CMV, HSV-1 and -2, and parvovirus B19 were measured from maternal serum in the first trimester and at delivery and from cord serum, mother's own information of her past infections was compared with her serological status. MAIN OUTCOME MEASURES: Seroprevalence, seroconversions and fetal transmission of VZV, CMV, HSV and parvovirus B19, reliability of maternal history of VZV, HSV and parvovirus B19. RESULTS: Seroprevalences were 96.2% for VZV, 56.3% for CMV, 54.3% for HSV, 46.8% for HSV-1, 9.3% for HSV-2 and 58.6% for parvovirus B19. Parity was associated with CMV seropositivity, maternal age differed only between HSV-2 seropositive and seronegative women, while area of residence (urban or rural) had no effect. Six seroconversions were observed: two VZV, one CMV and three parvovirus infections. No cases of primary HSV infections occurred. Fetal transmission was observed in two cases of parvovirus infection. No infants with anti-CMV IgM antibodies were born to CMV immunised women. False positive history of chickenpox was given only by 1.5% of the women, history of herpes infections was less reliable, and history of parvovirus infection was unreliable. CONCLUSIONS: Seroprevalence and the risk of viral infections during pregnancy cannot be extrapolated from one pregnant population to another. PMID- 15663398 TI - Smell perception during early pregnancy: no evidence of an adaptive mechanism. AB - OBJECTIVE: It has been suggested that nausea and vomiting in pregnancy is an evolutionary adaptive mechanism to avoid the ingestion of potentially harmful foods. It has also been suggested that the mechanism that triggers nausea and vomiting in pregnancy may be olfaction and that olfactory senses are invoked to provide this protection. This study aimed to test this theory in a systematic design. DESIGN: Cross sectional study. SETTING: The antenatal department of a maternity hospital in the north of England. SAMPLE: Three groups of participants: pregnant women (n= 55), non-pregnant women (n= 42) and men (n= 48). METHODS: Sensitivity was tested towards the odours of six standard stimuli (half safe and half associated with potentially harmful compounds). MAIN OUTCOME MEASURES: Odour rating of likeness, strength and pleasantness. RESULTS: Pregnant women rated safe and odours with potentially harmful compounds differently but not more so than men or non-pregnant women. There was no evidence that pregnancy changed the olfactory processes from the non-pregnant state and only slight differences between pregnant women and men were recorded. CONCLUSIONS: There was no evidence that olfactory processes had undergone any adaptation during pregnancy. The ability to differentiate safe from potentially harmful compounds was common to all three groups studied. PMID- 15663399 TI - The long term neurologic outcome of children from pregnancies complicated by twin to-twin transfusion syndrome. AB - OBJECTIVE: To assess long term outcomes of children from pregnancies complicated by twin-to-twin transfusion syndrome. DESIGN: Comparison of children from pregnancies with twin-to-twin transfusion syndrome in Western Australia with a contemporaneous regional comparison cohort of preterm and term infants studied using an identical assessment procedure. POPULATION AND SETTING: All infants aged > or =18 months were identified from a geographically based longitudinal cohort of monochorionic twin pregnancies with an antenatal diagnosis of twin-to-twin transfusion syndrome conducted prospectively since 1992. METHODS: Children were evaluated using age-specific developmental and behavioural assessments. Cerebral palsy was diagnosed clinically and ascertainment confirmed through the Western Australian Cerebral Palsy Register. MAIN OUTCOME MEASURES: Intellectual disability, cerebral palsy, behavioural and cognitive function. RESULTS: Fifty two children were identified as eligible for study and assessments were performed on 49 (94%). Three surviving children had a diagnosis of cerebral palsy (5.8%). The mean IQ score was 8 points lower in twin-to-twin transfusion syndrome children compared with the comparison cohort although this was mainly due to a decrement of 13 points in those born before 33 weeks of gestation. There was no difference between the donor and the recipient twin in terms of IQ scores (median difference -3, 95% CI -9 to 6). There was no relationship of IQ score to the worst stage of the twin-to-twin transfusion syndrome. Child Behavior Check List and Vineland Adaptive Behavior Scale scores did not differ between twin-to-twin transfusion syndrome children and the comparison group. CONCLUSIONS: Twin-to-twin transfusion syndrome is associated with a significant reduction in IQ score in very preterm survivors. There seems to be no increase in the prevalence of cerebral palsy. Overall behaviour and adaptive behaviour scale scores are similar to a comparison group. PMID- 15663400 TI - Effects of long term administration of KUR-1246, a selective beta(2)-adrenoceptor agonist, on pregnant sheep and their fetuses. AB - OBJECTIVE: To evaluate the safety of KUR-1246 as a tocolytic agent, we examined the effects of its long term infusion on respiratory and cardiovascular systems and general metabolism in pregnant sheep and their fetuses. DESIGN: Animal experiment with chronically instrumented ewes and their fetuses. SETTING: Center for animal experiments, Hokkaido University School of Medicine, Japan. SAMPLE: Eight Suffolk ewes at 117 to 120 days of gestation. METHODS: At 120-124 days of gestation, ewes (n= 4) were infused intravenously for 24 hours with KUR-1246 at 0.03 microg/kg/minute, a dose that completely inhibits oxytocin-induced uterine contractions in pregnant sheep. The controls received saline instead (n= 4). Statistical comparisons were carried out by repeated-measures ANOVA followed by Dunnett's test. MAIN OUTCOME MEASURES: Maternal and fetal values of heart rate, blood pressure, plasma electrolytes, glucose, insulin and non-esterified fatty acid levels, and blood gases and lactate level. RESULTS: The maternal plasma levels of KUR-1246 increased and reached a plateau at 15 hours or later from the start of the infusion, whereas the fetal levels of it were below the lower limit of quantification (0.1 ng/mL) throughout the experiment. Significant differences over time between the ewes that had received with KUR-1246 and the controls were found for the following parameters: maternal heart rate, blood lactate, plasma glucose, and plasma insulin levels, and fetal plasma glucose and plasma insulin levels (P < 0.05). In the KUR-1246 treated ewes, significant changes from the pre infusion value were detected in maternal blood lactate and fetal plasma glucose levels within 6 hours from the start of the infusion (P < 0.05). No significant differences were observed in other parameters in either ewes or fetuses. CONCLUSION: The physiologic changes induced by a 24-hour infusion of KUR-1246 were transient and considered to be within the compensatory capacity in both pregnant ewes and their fetuses, suggesting that KUR-1246 is a potentially safe tocolytic agent for use by long term infusion. PMID- 15663401 TI - Concentration of cord serum placenta growth factor in normal and diabetic pregnancies. AB - OBJECTIVE: To investigate whether maternal diabetes or diabetes-related complications, such as macrosomia and chronic fetal hypoxia, are associated with altered placenta growth factor (PlGF) levels in cord serum. DESIGN: Case-control study. SETTING: Helsinki University Central Hospital, Helsinki, Finland. POPULATION: Sixty-two normal pregnancies, 67 pregnancies complicated by type 1 diabetes and 28 pregnancies complicated by insulin-treated gestational diabetes. METHODS: Cord serum PlGF concentration was measured by an enzyme-linked immunosorbent assay. Amniotic fluid erythropoietin concentration was measured by a chemiluminescent immunologic method. Umbilical artery gas variables were analysed with standard blood gas and pH electrodes. MAIN OUTCOME MEASURE: PlGF concentration in cord serum at birth. RESULTS: Cord serum PlGF concentration was similar in normal pregnancies [13.4 (1.0) ng/L], in pregnancies complicated by type 1 diabetes [15.1 (1.8) ng/L, P= 0.583 vs controls] and in pregnancies complicated by insulin-treated gestational diabetes [13.6 (0.9) ng/L, P= 0.991 vs controls]. Cord serum PlGF did not correlate with relative birthweight. In diabetic pregnancies, cord serum PlGF correlated negatively with amniotic fluid erythropoietin (r=-0.449, P < 0.0001) and positively with umbilical artery Po(2) (r= 0.333, P= 0.001). There was a trend toward lower cord serum PlGF levels in diabetic pregnancies with pre-eclampsia compared with those without any hypertensive disorders. CONCLUSIONS: Maternal diabetes per se is not associated with altered PlGF levels in cord serum. The correlation between PlGF and indices of fetal hypoxia in diabetic pregnancies may be related to the role of PlGF in potentiating the angiogenic response to vascular endothelial growth factor in ischaemia. PMID- 15663402 TI - Repeated measures screening for Down's Syndrome. AB - OBJECTIVE: To demonstrate the potential value of screening for Down's Syndrome using highly correlated repeated measures of serum markers taken in the first and second trimesters of pregnancy. DESIGN: A Monte Carlo simulation study. POPULATION: Detection rates and false positive rates relating to the maternal age distribution of England and Wales for the period 1996 to 1998 were obtained using marker distributions from the SURUSS study. RESULTS: Screening using first trimester nuchal translucency and repeated measures of uE3 and PAPP-A in the first and second trimester has an estimated false positive rate of 0.3% for an 85% detection rate. This should be compared with the integrated test with an estimated false positive rate of 1.2% for the same detection rate. CONCLUSIONS: The performance of repeated measures screening tests, and their acceptability to women, should be assessed in further prospective studies. PMID- 15663403 TI - Early onset severe pre-eclampsia: expectant management at a secondary hospital in close association with a tertiary institution. AB - OBJECTIVES: Early onset severe pre-eclampsia is ideally managed in a tertiary setting. We investigated the possibility of safe management at secondary level, in close co-operation with the tertiary centre. DESIGN: Prospective case series over 39 months. SETTING: Secondary referral centre. POPULATION: All women (n= 131) between 24 and 34 weeks of gestation with severe pre-eclampsia, where both mother and fetus were otherwise stable. METHODS: After admission, frequent intensive but non-invasive monitoring of mother and fetus was performed. Women were delivered on achieving 34 weeks, or if fetal distress or major maternal complications developed. Transfer to the tertiary centre was individualised. MAIN OUTCOME MEASURES: Prolongation of gestation, maternal complications, perinatal outcome and number of tertiary referrals. RESULTS: Most women [n= 116 (88.5%)] were managed entirely at the secondary hospital. Major maternal complications occurred in 44 (33.6%) cases with placental abruption (22.9%) the most common. One maternal death occurred and two women required intensive care admission. A mean of 11.6 days was gained before delivery with the mean delivery gestation being 31.8 weeks. The most frequent reason for delivery was fetal distress (55.2%). There were four intrauterine deaths. The perinatal mortality rate (> or =1000 g) was 44.4/1000, and the early neonatal mortality rate (> or =500 g) was 30.5/1000. CONCLUSIONS: The maternal and perinatal outcomes are comparable to those achieved by other tertiary units. This model of expectant management of early onset, severe pre-eclampsia is encouraging but requires close co-operation between secondary and tertiary institutions. Referrals to the tertiary centre were optimised, reducing their workload and costs, and patients were managed closer to their communities. PMID- 15663404 TI - Incidence of severe pre-eclampsia, postpartum haemorrhage and sepsis as a surrogate marker for severe maternal morbidity in a European population-based study: the MOMS-B survey. AB - OBJECTIVE: To describe the incidence of three conditions of acute severe maternal morbidity in selected regions in nine European countries. DESIGN: A population based questionnaire survey. SETTING: Eleven regions in nine countries of Europe. POPULATION: All the pregnant women in each region who had delivered during the period covered by the study. METHODS: Standard definitions of three severe obstetric conditions, pre-eclampsia, postpartum haemorrhage and sepsis were established by a steering committee. A common questionnaire was used in each participating country. The incidence of the three obstetric conditions and characteristics of the study women were compared. MAIN OUTCOME MEASURES: Incidence of three severe obstetric conditions: pre-eclampsia, postpartum haemorrhage and sepsis. RESULTS: The study identified 1734 women with at least one of the three conditions, with 847 experiencing severe haemorrhage, 793 experiencing severe pre-eclampsia and 142 experiencing severe sepsis. There were wide variations in incidence of three conditions combined, ranging from 14.7 per thousand deliveries in Brussels, Belgium to 6.0 per thousand deliveries in Upper Austria. CONCLUSIONS: This study sets a simple and straightforward approach to the definition of three severe obstetric conditions and allows population-based comparisons between developed countries in Europe, even though difficulties may have been present with applying the definition across countries. The reported incidence of these severe obstetric conditions in general and severe haemorrhage varied significantly between countries. Overall, severe haemorrhage in particular was the most common of the three conditions, followed closely by severe pre eclampsia. PMID- 15663405 TI - Misoprostol for second trimester pregnancy termination in women with prior caesarean section. AB - OBJECTIVE: To examine whether a previous caesarean section increases the risk for complications in women undergoing a mid-trimester pregnancy termination by labour induction. DESIGN: Retrospective analysis of case records between 1997 and 2002. SETTING: Fetal Medicine Unit of a large teaching hospital. POPULATION: One hundred and eight women with a previous caesarean section (study group) and 216 women without such a history (controls), who underwent a second trimester termination of pregnancy. METHODS: All the terminations were performed between 17 and 24 weeks of gestation by using 400 mug of oral administration of misoprostol in combination with 400 mug of intravaginal misoprostol. The same dose of intravaginal misoprostol was repeated every 6 hours for a maximum of five doses. MAIN OUTCOME MEASURES: Severe haemorrhage requiring blood transfusion, post abortal infection, retained placenta and uterine rupture. RESULT: Complications occurred in 16 out of 108 women of the study group (15%) and in 26 out of 216 of the controls (12%), with only one ruptured uterus in the control group. CONCLUSION: We found no evidence that a previous caesarean delivery affects the incidence of complications when women with such a history undergo a mid-trimester pregnancy termination with misoprostol. PMID- 15663406 TI - Previous abdominal surgery and closed entry for gynaecological laparoscopy: a prospective study. AB - OBJECTIVES: To assess the morbidity from closed laparoscopic access and define the role of previous surgery on the occurrence of these complications. DESIGN AND METHODS: We prospectively recorded data on all laparoscopic procedures between January 2000 and January 2001. We compared results from patients without previous abdominal surgery (group I) with patients with prior abdominal surgery (group II). The insertion site for the Verres needle was trans-umbilical for patients in group I and in the left upper quadrant for patients in group II. SETTING: Gynaecology department of a University Hospital. POPULATION: All laparoscopic procedures between January 2000 and January 2001. MAIN OUTCOME MEASURES: We compared results from patients without previous abdominal surgery (group I) with patients with prior abdominal surgery (group II). The insertion site for the Verres needle was trans-umbilical for patients in group I and the left upper quadrant for patients in group II. RESULTS: Four hundred and seventy-seven laparoscopies were carried out during the study period, 368 women without previous surgery were included in group I, and 109 women were included in group II. We recorded 1 complication (overall complication rate of 0.2% and 31 incidents (6.4%). One complication (small bowel injury) was related to the insertion of the Verres needle (0.2%). The incidents and complications occurred only in group II (P < 0.05). All the complications were treated by laparoscopy. CONCLUSION: The complication rate of the entry step is low in gynaecological laparoscopy. A previous history of laparotomy increases the risk of these complications and incidents. Safety rules and other access method should be investigated for these patients. PMID- 15663407 TI - Evaluation of endocervical, first-void urine and self-administered vulval swabs for the detection of Chlamydia trachomatis in a miscarriage population. AB - OBJECTIVE: To compare, in parallel, different approaches of opportunistically screening women with bleeding in early pregnancy for Chlamydia trachomatis. DESIGN: Prospective observational [corrected] study. SETTING: Early pregnancy assessment unit, University hospital, Scotland, UK. POPULATION: Over 200 consecutive women admitted to an early pregnancy assessment unit were recruited. All had a positive pregnancy test, a history of vaginal bleeding and were less than 24 weeks of gestation. Women with recent antibiotic use, heavy vaginal bleeding and cervical shock excluded. METHODS: Each women provided two or more of the following specimens: a self-administered vulval swab, first-void urine and/or endocervical swab. Following screening, each completed a semi-structured questionnaire assessing the acceptability of each method undertaken. MAIN OUTCOME MEASURE: Subjective rating of the screening methods; prevalence; method performance, including proportion requiring repeat testing. RESULTS: The majority accepted screening, with moderate prevalence rates (95% CI) 3.9% (2.0-7.4%) identified. All positive women were less than 30 years of age. Parallel screening exposed the potential of reduced test performance with urine. Non-invasive sampling was more acceptable, but more likely to require repeat testing. CONCLUSION: Both acceptability and the effect of bleeding on test performance need further assessment before a particular specimen can be recommended for screening this population of women for C. trachomatis. PMID- 15663408 TI - Functional and anatomical outcome of anterior and posterior vaginal prolapse repair with prolene mesh. AB - OBJECTIVE: To evaluate the effects of prolene mesh on urinary, bowel and sexual function in prolapse surgery. DESIGN: Prospective observational study on consecutive women. SETTING: Two referral uorgynaecological units in Italy. POPULATION: Women requiring prolapse repair for anterior or posterior vaginal prolapse. METHODS: All women were assessed for urinary, bowel, prolapse symptoms and dyspareunia pre- and post-operatively. Urodynamics was performed in selected cases. Surgery consisted of an anterior or posterior repair plus a prolene mesh. Follow up was after 1, 6 and 12 months. The ANOVA test was used for statistical analysis. MAIN OUTCOME MEASURES: Vaginal anatomical restoration, urinary, bowel and sexual function. RESULTS: We recruited 63 women (mean age 63 years) with a mean follow up of 17 months. Anatomically, the success rate was 94%. Thirty-two women had an anterior repair. Among this group, the sexual activity rate did not alter but dyspareunia increased by 20%. Urge and stress incontinence did not change post-operatively but urgency improved in 10% and 13% had vaginal erosion of the mesh. Thirty-one women had a posterior repair. Among this group, sexual activity decreased by 12% and dyspareunia increased in 63%. Constipation improved in 15% and anal incontinence in 4%, and 6.5% of women had vaginal erosion of the mesh and one required mesh removal for pelvic abscess. CONCLUSIONS: Although this study shows good anatomical results with the use of prolene mesh for prolapse repair, there was a high rate of morbidity. We believe that the use of prolene mesh should be abandoned. PMID- 15663409 TI - Bladder mucosal autografts for repair of vesicovaginal fistula. AB - Twenty-two women with primary and secondary (five patients) vesicovaginal fistula attending a tertiary level urological unit in India were treated by repair of the fistula using bladder mucosal autografts. The fistula was approached transabdominally or via a combined abdominal and vaginal approach (for those involving the trigone). After closure of the vaginal layer, bladder mucosa was harvested from the dome of the bladder and laid over the fistula with sutures at each corner to fix it in place. Patients were catheterised for 12-14 days. At follow up after 3 to 12 months, 20 out of 22 patients were continent, with no other symptoms. The two failures had undergone two previous repairs each. This series is the first from India, and demonstrates the efficacy of bladder mucosal autografts for managing large fistulae, those where a previous repair has failed and fistulae adjacent to the ureteric orifice without the need for uretero neocystostomy. PMID- 15663410 TI - Total laparoscopic hysterectomy as a primary surgical treatment for endometrial cancer in morbidly obese women. AB - To evaluate the feasibility of total laparoscopic hysterectomy as the primary treatment for endometrial cancer in morbidly obese women, an audit was carried out during an 18-month period in a tertiary referral centre for gynaecological oncology. Four women who had laparoscopic surgery were compared with a similar cohort who had open surgery. The mean operating time was equivalent, without evidence of excess morbidity with the laparoscopic approach. However, inpatient stay was longer with open versus laparoscopic surgery (11.5 vs 4 days). Laparoscopic surgery is safe to use in morbidly obese women with endometrial cancer. PMID- 15663411 TI - Pelvic abscess following microwave endometrial ablation. PMID- 15663412 TI - Vertex haemangioma: an unusual cause of intrapartum haemorrhage. PMID- 15663413 TI - Uterus and bladder necrosis after uterine artery embolisation for postpartum haemorrhage. PMID- 15663414 TI - Pregnancy outcome in a renal transplant recipient with residual mild tertiary hyperparathyroidism. PMID- 15663415 TI - Partial ischemic necrosis of the uterus following a uterine brace compression suture. PMID- 15663416 TI - Partial ischemic necrosis of the uterus following a uterine brace compression suture. PMID- 15663417 TI - Antenatal screening and intrapartum management of Group B streptococcus in the UK. PMID- 15663419 TI - The future of antenatal screening for Down's Syndrome. PMID- 15663421 TI - Visions in monochrome: families, marriage and the individualization thesis. AB - This paper takes issue with the way in which the individualization thesis--in which it is assumed that close relationships have become tenuous and fragile--has become so dominant in 'new' sociological theorizing about family life. Although others have criticized this thesis, in this paper the main criticism derives from empirical research findings carried out with members of transnational families living in Britain whose values and practices do not fit easily with ideas of individualization. It is argued that we need a much more complex and less linear notion of how families change across generations and in time. PMID- 15663422 TI - Symbolic violence and the neighbourhood: the educational aspirations of 7-8 year old working-class girls. AB - This article focuses on the experiences of 7-8 year old working-class girls in Belfast, Northern Ireland and their attitudes towards education. It shows how their emerging identities tend to emphasize relationships, marriage and motherhood at the expense of a concern with education and future careers. The article suggests that one important factor that can help explain this is the influence of the local neighbourhood. In drawing upon Bourdieu's concepts of symbolic violence and habitus and Elias' notion of figuration, the article shows how the local neighbourhood represents the parameters of the girls' social worlds. It provides the context within which the girls tend to focus on social relations within their community and particularly on family relationships, marriage and children. It also provides the context within which the girls tend to develop strong interdependent relationships with their mothers that also tend to encourage and reinforce the girls' particular gendered identities. The article concludes by arguing that there is a need for more research on working-class girls and education to look beyond the school to incorporate, more fully, an understanding of the influence of the family and local neighbourhood on their attitudes towards education and their future career aspirations. PMID- 15663424 TI - The globalization of football: a study in the glocalization of the 'serious life'. AB - Sport, in particular football, constitutes one of the most dynamic, sociologically illuminating domains of globalization. This paper examines the globalization of football with particular reference to Robertson's theorizations of global processes. We examine football's cultural globalization through the concept of 'glocalization', which highlights the interdependence of local and global processes within the game's identities and institutions. We address economic globalization in football by considering the world's leading clubs as 'glocal' transnational corporations. We assess the political globalization of football with reference to the possible enhancement of democracy within the game's international governance. We conclude by affirming the utility of sport in advancing our empirical and theoretical understanding of globalization processes. PMID- 15663439 TI - Teaching oral hygiene to children with autism. AB - BACKGROUND: The need for improved oral hygiene routines in individuals with disabilities has been documented in many reports. The aim of this study was to evaluate whether visual pedagogy is a suitable way to teach children with autism how to brush their teeth. METHODS: The investigation took the form of a prospective study including clinical examinations and structured interviews. Based on visual pedagogy, a series of pictures were produced that showed a structured method and technique of tooth brushing. The pictures were placed in the bathroom or wherever tooth brushing was performed. Fourteen children with autism aged between 5 and 13 years (mean age = 9.3 years), and their parents participated. RESULTS: Before the study, all parents found it difficult/very difficult to maintain good oral hygiene in their child. All children had visible plaque on their maxillary incisors and canines. After 12 months, the amount of visible plaque was reduced. After 18 months, most parents found maintaining good oral hygiene easier than before the study. All but one child/parent adopted the programme. CONCLUSIONS: Visual pedagogy is a useful tool in helping people with autism to improve their oral hygiene. PMID- 15663440 TI - A study into dentists' knowledge of the treatment of traumatic injuries to young permanent incisors. AB - OBJECTIVE: The aims of this study were to evaluate dentists' knowledge of the emergency treatment of traumatic injuries to young permanent incisors, and to investigate barriers to treatment. DESIGN: A closed-ended questionnaire was sent to 1023 general dental practitioners (GDPs) and community dental officers (CDOs) in West/North Yorkshire and Humberside, UK. METHODS: The questionnaire comprised 17 questions. Six questions asked for general information about the participants (i.e. profession, age, gender, year of graduation, training or education on dental trauma, and willingness to provide emergency care), 10 were relevant to the emergency treatment of crown fractures, root fractures, luxation and avulsion injuries, and the last question queried any perceived barriers to treatment. Results. Seven hundred and twenty-four questionnaires were returned, a response rate of 71%, and these indicated that dentists' knowledge of the emergency treatment of dentoalveolar trauma in children was inadequate. The CDOs were significantly more knowledgeable than the GDPs, as were younger and more recently graduated dentists compared with older ones. The GDPs regarded the difficulty of treating children and the inadequate fees of the UK National Health Service as important barriers to treatment. Dentists who attended continuing dental education courses on dental traumatology had a more thorough knowledge than those who did not. CONCLUSION: Overall, the dentists' knowledge of the emergency treatment of dentoalveolar trauma in children was inadequate. Greater emphasis on undergraduate and postgraduate education in this area is indicated. PMID- 15663441 TI - Hypomineralized molars and incisors of unknown origin: treatment outcome at age 18 years. AB - OBJECTIVE: To assess the outcome of treatment of hypomineralized molars and incisors of unknown aetiology (MIH) in 18-year-olds. DESIGN: A follow-up study including clinical examination, panoramic radiography and intraoral photos. SAMPLE AND METHOD: Seventy-six individuals treated at the Eastman Dental Institute in Stockholm during 1978-2001 with the diagnosis MIH. Severity of enamel defects in molars and incisors, prevalence and distribution of extracted molars, type, quality and median duration of restorations, periradicular condition of affected molars, dental occlusion and space closure in cases of extraction, as well as the individual's satisfaction with the treatment, were assessed. RESULTS: Severe defects with enamel surface breakdown in all four molars occurred in 42% of the individuals and 29% had at least one incisor with yellow/brown opacity in the enamel. At follow up, 42% of the individuals had at least one molar extracted; 18% had all four molars extracted. The median duration of the molar restorations (n = 153) was 5 years. Of the individuals with restored molars, 48% had at least one unacceptable restoration. Periradicular pathology was observed in three molars. The sagittal relations did not differ between individuals with and without extraction of molars. Space closure was acceptable in 87% of the individuals with extracted molars. Eighty percent were satisfied with the treatment. CONCLUSIONS: Extraction of molars with severe enamel defects gave good or acceptable results in a majority of the patients while conservative restorative treatment resulted in a need for additional treatment in approximately half of the patients. PMID- 15663442 TI - Relationship between oral parafunctional/nutritive sucking habits and temporomandibular joint dysfunction in primary dentition. AB - The aim of this study was to evaluate the relationship between nutritive and parafunctional habits and the presence of temporomandibular dysfunction (TMD) in children with primary dentition. METHODS: Ninety nine children, aged 3-5 years, were examined to check for the presence or absence of signs and symptoms of TMD (headache, preauricular pain, earache, masticatory muscle tenderness, deviation on opening, occlusal interference and asymmetric movement of the mandible), oral parafunctions (bruxism, nail biting, finger/thumb sucking, speech alteration, mouth breathing, pacifier and atypical swallowing) and nutritive habits (breast- or bottle-feeding) through interview and clinical examination. The results were submitted to descriptive statistical analysis and Fisher's exact test. RESULTS: The results showed that only atypical swallowing was positively related to TMD (P < 0.0001); other oral parafunctional and nutritive habits were not related to TMD. CONCLUSIONS: It was concluded that parafunctional habits, with the exception of atypical swallowing, and feeding methods were not determinants for the presence of signs and/or symptoms of TMD in the sample of children included in the study. PMID- 15663443 TI - Clinical observations of odontomas in Japanese children: 39 cases including one recurrent case. AB - Retrospective investigations of odontomas in Japanese children and one recurrent case were carried out. Thirty-nine cases of odontoma in 38 children were treated in the Paediatric Dentistry Clinic of Niigata University Dental Hospital between September 1979 and December 2002. The patients consisted of 23 males and 15 females and their ages ranged from 1 year 2 months to 14 years old. The chief complaints were delayed tooth eruption in 19 cases (five: primary teeth, 14: permanent teeth), retention of primary teeth in 11, incidentally found on the radiographic examination in eight cases, and swelling of the jaw in one case. Thirty-four cases (87%) were associated with tooth eruption disturbances. The most frequently affected region was the maxillary anterior region. Treatment consisted of surgical removal of odontomas in all cases, after which if the impacted teeth did not erupt, exposure of the crown and/or orthodontic traction was performed. Pathological diagnoses were compound odontoma in 30 cases, complex odontoma (n = 7), and compound and complex odontoma (n = 2). A retrospective study of the radiographs revealed the developing process of odontomas in four cases and odontoma disturbed tooth eruption since the early uncalcified developing stage. A recurrent case was a boy aged 6 years 5 months in whom the first surgical removal of odontoma was performed at the age of 1 year 8 months. Recurrence of an odontoma is very rare, but in very young children odontomas are in the early developing stages, containing uncalcified portions, so it is important to perform periodical observations until the succedaneous teeth erupt. PMID- 15663444 TI - Randomized, controlled trial comparing the retention of a flowable restorative system with a conventional resin sealant: one-year follow up. AB - This clinical trial evaluated the retention rate of a flowable restorative system (Bond 1 + Flow-It!) used as a pit-and-fissure sealant compared with a conventional filled resin sealant (Fluroshield) over a 1-year period. METHOD: Using a half-mouth design, 160 sealants (80 in primary and 80 in permanent teeth) were placed on sound first/second primary molars and first permanent molars of 40 children aged between 4 and 7 years. For both primary and permanent dentitions, half the teeth (n = 40) were sealed with Fluroshield and half (n = 40) with Bond 1 + Flow-It! Teeth were evaluated at baseline, 6- and 12-month intervals. RESULTS: For both materials, there was no total loss of sealants placed on either the primary or permanent molars over 1 year. From Fluroshield sealants placed on primary teeth, 33 were completely intact after 6 months and 31 after 1 year. From those placed on permanent molars, no loss of material was observed after 6 months, while partial loss was noticed on 5% of teeth at 1-year recall. For Flow It! resin applied on primary molars, partial loss of material was observed in only 1 sealant after 6 months and in 2 sealants after 1 year. On permanent teeth, 100% retention rate was observed over a 1-year follow up. There was a statistically significant difference (P < 0.01) between the sealing materials on primary but not permanent teeth, and, overall, Flow-It! sealants presented a higher retention rate at both 6-month and 1-year evaluations. Significant differences (P < 0.01) between baseline and the other evaluation periods were also observed. CONCLUSION: It may be concluded that the flowable restorative system yielded optimal retention on both primary and permanent molars. Its retention rate was significantly higher than that of the conventional pit-and fissure sealant on primary teeth. PMID- 15663445 TI - In-vitro microleakage of repaired fissure sealants: a randomized, controlled trial. AB - The aim of this study was to investigate whether differences in surface treatment prior to repair influenced the seal of a resin fissure sealant placed on the occlusal surfaces of permanent molar teeth. SETTING: In-vitro study. SAMPLE AND METHODS: One hundred and twenty-eight extracted human first and second molars were randomly allocated to one of four groups of 32 teeth each. A light cured, unfilled, opaque resin fissure sealant (Delton) was placed on their occlusal surface following cleaning by prophylaxis and acid etching. Following storage in artificial saliva (Saliva Orthana) for a week, duplication of sealant failure was carried out. The teeth were then subjected to one of four different surface treatments: Group 1: with a slow-speed prophylaxis brush followed by acid etching (control method); Group 2: a slow-speed bur and acid etching; Group 3: air abrasion and acid etching; and Group 4: acid etching and application of a bonding agent. Following a further week of storage in artificial saliva two layers of impermeable varnish were applied to the nonocclusal surfaces of the teeth; their apices were then sealed with wax and the teeth were immersed in 1% methylene blue for 48 h. The teeth were then sectioned (ISOMET 1000) to achieve three cuts resulting in a maximum of four blocks, i.e. six surfaces per tooth. A total of 715 sections from 126 teeth were scored for microleakage on the intact and repaired side of the fissure sealant. RESULTS: Statistical analysis did not demonstrate any one single method of repair to be superior to the control method for reapplication of the sealant. CONCLUSION: All four techniques compared in this study seem to be acceptable for replacing or repairing lost or fractured fissure sealants. As prophylaxis with a brush rotating at slow speed followed by acid etching, which probably represents current practice, is also the simplest technique that can be practised on children, it is therefore recommended. PMID- 15663446 TI - Clinical performance of a non-rinse conditioning sealant in three paediatric dental practices: a retrospective study. AB - The present clinical retrospective study describes the retention rates of a compomer sealant (Dyract Seal, Dentsply-De Trey, Germany) with non-rinse conditioning (NRC) placed in three paediatric dentistry practices. Three hundred and seventeen sealants were applied in 220 primary and 97 permanent molars of 176 children aged 2.5-13 years. The tooth surface was freshened with a #1/2 round bur mounted on a slow speed engine, and isolated with cotton rolls. Application of the NRC and Dyract Seal followed the manufacturer's instructions. Sealant retention was classified as A (fully retained), B (partially lost) or C (completely missing). From a total of 220 sealants placed in primary molars, 38 were in the mouth between 12 and 18 months, 29 functioned between 19 and 24 months and 46 were followed up between 25 and 36 months. One hundred and thirteen (51%) were fully retained (A), 73 (33%) scored B (27 for 12-18 months; 20 for 19 24 months, 26 for 25-36 months) and 34 (16%) were lost and scored C (12 for 12-18 months; 12 for 19-24 months; 10 for 25-36 months). From a total of 97 sealed permanent molars, 45 (46%) were fully retained (score A). Of these, 25 were followed up for 12-18 months, 10 for 19-24 months and 10 for 25-36 months. Thirty eight sealed permanent molars (19%) scored B [17 for 12-18 months, 10 for 19-24 months and seven for 25-36 months and 14 (15%) were completely lost (score C: 6 for 12-18 months, seven for 19-24 months and two for 25-36 months)]. Dyract Seal has a lower retention rate than conventional sealants. It may be appropriate for sealing primary molars of very young children for a limited period of time and for permanent molars of children with pronounced gag reflex, where rinsing can become a problem and lead to disruptive behaviour. PMID- 15663447 TI - Talon cusp in a primary lateral incisor from a medieval child. AB - This paper describes talon cusp in an ancient skeleton and is a rare report of its occurrence in a primary lateral incisor. As well as talon cusp, the affected incisor also shows abnormal widening, probably representing a double tooth. There is also a supernumerary permanent incisor. The report shows that talon cusp existed in British populations more than 600 years before the first written description. A brief review of the literature on the occurrence of talon cusp in the primary dentition is presented. PMID- 15663449 TI - An unusual intranasal foreign body in an unsuspecting 9-year old. AB - This report describes the incidental finding and removal of an unusual intranasal foreign body in a 9-year-old boy. The spectrum of items lost and found in the nasal cavity are reviewed before discussing the management of this case. Dental Practitioners should remain vigilant as radiopaque foreign bodies can be identified on commonly taken dental radiographic views. Patients should be referred promptly for foreign body removal in order to minimize potential complications. PMID- 15663448 TI - A point mutation of the ED1 gene in a Japanese family with X-linked hypohidrotic ectodermal dysplasia. AB - X-linked hypohidrotic ectodermal dysplasia (EDA) is characterized by the hypoplasia or absence of hair, teeth and sweat glands. In this study, the authors investigated the ED1 gene in a Japanese family with X-linked hypohidrotic ectodermal dysplasia. The only affected male fulfils the diagnostic criteria for this disorder. His parents were not consanguineous and both of them were healthy. After informed consent, genomic DNA was isolated from the peripheral blood lymphocytes or oral buccal epithelial cells of all members of the family. A polymerase chain reaction fragment containing exon 9 of the ED1 gene was amplified using primers. The patient's amplified fragment, as well as those from his father, mother and sister, were directly sequenced. The sequence from the patient revealed a point mutation (G1149A) in exon 8 of the ED1 gene, which changes codon 291 from glycine to arginine. Heterozygosity was demonstrated in his mother and sister. This mutation has not been reported previously. The amino acid substitution is predicted to disrupt the transmembrane domain, which strongly implies that this is the disease-causing mutation in the family. PMID- 15663456 TI - The performance and subjective responses of call-center operators with new and used supply air filters at two outdoor air supply rates. AB - A 2 x 2 replicated field intervention experiment was conducted in a call-center providing a telephone directory service: outdoor air supply rate was adjusted to be 8% or 80% of the total airflow of 430 l/s (3.5 /h) and the supply air filters were either new or had been in place for 6 months. One of these independent variables was changed each week for 8 weeks. The interventions did not affect room temperature, relative humidity or noise level. The 26 operators were blind to conditions and each week returned questionnaires recording their environmental perceptions and Sick Building Syndrome (SBS) symptoms. Their performance was continuously monitored by recording the average talk-time every 30 min. Replacing a used filter with a clean filter reduced talk-time by about 10% at the high ventilation rate but had no significant effect at the low rate. Increasing the outdoor air supply rate reduced talk-time by 6% with a new filter in place but increased talk-time by 8% with a used filter in place. The interventions also had significant effects on some SBS symptoms and environmental perceptions. The present results indicate that increasing outdoor air supply rate and replacing filters can have positive effects on health, comfort and performance. PRACTICAL IMPLICATIONS: Supply air filters should be changed frequently not just because their airflow resistance increases progressively but because they degrade air quality with negative consequences for health, comfort and the performance of office work, all of which are factors that affect office productivity (profitability). Increasing outdoor air supply rates may only be beneficial when new filters are installed. Unwanted negative effects may be produced when used filters are in place. Filter condition (used or new) should always be recorded to make it possible to draw sound conclusions in studies of the effects of outdoor air supply rates on building occupants. PMID- 15663457 TI - Human response to personalized ventilation and mixing ventilation. AB - The response of 60 human subjects to a personalized ventilation system (PVS), providing control of positioning of the air terminal device and the airflow rate, was compared with the response to mixing ventilation (MV). Perceived air quality, thermal comfort, intensity of Sick Building Syndrome symptoms and performance of subjects were studied during 3 h 45 min exposures. In case of MV alone the room air temperature was 23 degrees C and 26 degrees C. The PVS supplied outdoor air at 23 degrees C or 20 degrees C or recirculated room air at 23 degrees C when the room temperature was 23 degrees C, and outdoor air at 20 degrees C when the room temperature was 26 degrees C. The PVS providing outdoor air improved perceived air quality and decreased SBS symptoms compared to MV alone and when the room air was re-circulated through the PVS. The percentage dissatisfied with air quality, 3 min after initial occupancy, decreased from 22% with MV to 7% with PVS; and from 49% to 20%, at room temperatures 23 degrees C and 26 degrees C, respectively. Over time, these differences in percentage dissatisfied decreased markedly. Headache and decreased ability to think clearly were reported as least intense when the PVS supplied outdoor air at 20 degrees C, while the most intense symptoms occurred with MV. PVS increased self-estimated performance. PRACTICAL IMPLICATIONS: Personalized ventilation can improve occupants' thermal comfort, perceived air quality and decrease the intensity of SBS symptoms compared to mixing ventilation. Occupants will use the provided individual control of airflow rate and positioning of the air terminal device to obtain preferred microenvironment in rooms where the air temperature is within the range recommended by indoor climate standards. Development of more efficient air terminal devices is recommended. PMID- 15663458 TI - The effects of moderate heat stress and open-plan office noise distraction on SBS symptoms and on the performance of office work. AB - Three air temperatures (22/26/30 degrees C) and two acoustic conditions-quiet (35 dBA) or open-plan office noise (55 dBA)-were established in an office. Thirty subjects aged 18-29 years (16 male), clothed for thermal neutrality at 22 degrees C, performed simulated office work for 3 h under all six conditions. Many more (68% vs. 4%) were dissatisfied with noise in the noise condition (P < 0.01). Warmth decreased thermal acceptability (P < 0.001) and perceived air quality (P < 0.01) and increased odour intensity (P < 0.05) and stuffiness (P < 0.01). After 2 h, some forehead sweating was observed on 4, 36 and 76% of subjects (P < 0.001) at 22, 26 and 30 degrees C, while 0, 21 and 65% felt "warm" (P < 0.001). Raised temperature increased eye, nose and throat irritation (P < 0.05), headache intensity (P < 0.05), difficulty in thinking clearly (P < 0.01) and concentrating (P < 0.01), and decreased self-estimated performance (P < 0.001). Noise increased fatigue (P < 0.05) and difficulty in concentrating (P < 0.05) but did not interact with thermal effects on subjective perception. In an addition task, noise decreased workrate by 3% (P < 0.05), subjects who felt warm made 56% more errors (P < 0.05) and there was a noise-temperature interaction (P < 0.01): the effect of warmth on errors was less in the noise condition. Typing speed (P < 0.05) and reading speed (P < 0.05) were higher in noise. PRACTICAL IMPLICATIONS: This paper demonstrates that open office noise distraction, even at the realistic level of 55 dBA, increases fatigue and has many negative effects on the performance of office work, as does a moderately warm air temperature. These findings may be used to provide economic justification for the provision of private offices and air temperature control in hot weather. The additional finding that noise distraction and heat stress can sometimes counteract each other in the short term is of academic interest only, as they both increase subjective distress and fatigue. In practice, neither should be deliberately introduced to counteract the other. PMID- 15663459 TI - Worker performance and ventilation in a call center: analyses of work performance data for registered nurses. AB - We investigated the relationship between ventilation rates and individual work performance in a call center, and controlled for other factors of the indoor environment. We randomized the position of the outdoor air control dampers, and measured ventilation rate, differential (indoor minus outdoor) carbon dioxide (DeltaCO(2)) concentration, supply air velocity, temperature, humidity, occupant density, degree of under-staffing, shift length, time of day, and time required to complete two different work performance tasks (talking with clients and post talk wrap-up to process information). DeltaCO(2) concentrations ranged from 13 to 611 p.p.m. We used multivariable regression to model the association between the predictors and the responses. We found that agents performed talk tasks fastest when the ventilation rate was highest, but that the relationship between talk performance and ventilation was not strong or monotonic. We did not find a statistically significant association between wrap-up performance and ventilation rate. Agents were slower at the wrap-up task when the temperature was high (> 25.4 degrees C). Agents were slower at wrap-up during long shifts and when the call center was under-staffed. PRACTICAL IMPLICATIONS: The productivity benefits of ventilation rates that exceed common standards such as ASHRAE Standard 62 may be small (0-2%), and other factors may have a larger impact on productivity. Understaffing and long shifts should be avoided because both showed a negative impact on performance. In this study, high temperature had the largest statistically significant impact on productivity and was caused by occupants fighting over the thermostat setpoint. Care should be taken to avoid high temperatures in call centers. If occupants are allowed to adjust temperature setpoints, then the size and/or duration of the setpoint change should be restricted. PMID- 15663460 TI - Study of effect of adsorptive building material on formaldehyde concentrations: development of measuring methods and modeling of adsorption phenomena. AB - In this paper, a method is developed to assess the performance of adsorptive building materials that are used for reducing indoor pollutant concentrations. Mass transfer has a great influence on the materials' performance. To control the mass transfer rate precisely in the performance test, the authors have developed the Boundary-Layer-Type Small Test Chamber in which airflow along the test materials can be controlled precisely. A new index of adsorption performance, the equivalent ventilation rate (Q(ads)), is defined that corresponds to the mass transfer coefficient when the surface pollutant concentration is zero. Modeling and experimental verification of adsorption were done, demonstrating the pollutant concentration decrease caused by adsorptive building materials. The pollutant reduction phenomena were modeled, including pollutant degradation by chemical reaction and adsorption in building materials. Adsorption tests of gypsum board containing a substance that decomposes HCHO within the board are reported. The adsorption rate of the gypsum board predicted by numerical analysis (CFD, Computational Fluid Dynamics) corresponds well with experimental results. PRACTICAL IMPLICATIONS: Development and verification of a method to measure the decrease in indoor pollutant concentration caused by an adsorptive building material are reported. Mass transfer has a great influence on the material's performance. The equivalent ventilation rate (Q(ads)) of the adsorption performance is defined as a new index that corresponds to the mass transfer coefficient. The equivalent ventilation rate (Q(ads)) can be used directly to compare the effect of pollutant concentration decrease via adsorption with the effect of ventilation. PMID- 15663461 TI - Listening to the occupants: a Web-based indoor environmental quality survey. AB - Building occupants are a rich source of information about indoor environmental quality and its effect on comfort and productivity. The Center for the Built Environment has developed a Web-based survey and accompanying online reporting tools to quickly and inexpensively gather, process and present this information. The core questions assess occupant satisfaction with the following IEQ areas: office layout, office furnishings, thermal comfort, indoor air quality, lighting, acoustics, and building cleanliness and maintenance. The survey can be used to assess the performance of a building, identify areas needing improvement, and provide useful feedback to designers and operators about specific aspects of building design features and operating strategies. The survey has been extensively tested and refined and has been conducted in more than 70 buildings, creating a rapidly growing database of standardized survey data that is used for benchmarking. We present three case studies that demonstrate different applications of the survey: a pre/post analysis of occupants moving to a new building, a survey used in conjunction with physical measurements to determine how environmental factors affect occupants' perceived comfort and productivity levels, and a benchmarking example of using the survey to establish how new buildings are meeting a client's design objectives. PRACTICAL IMPLICATIONS: In addition to its use in benchmarking a building's performance against other buildings, the CBE survey can be used as a diagnostic tool to identify specific problems and their sources. Whenever a respondent indicates dissatisfaction with an aspect of building performance, a branching page follows with more detailed questions about the nature of the problem. This systematically collected information provides a good resource for solving indoor environmental problems in the building. By repeating the survey after a problem has been corrected it is also possible to assess the effectiveness of the solution. PMID- 15663462 TI - An experimental comparison of a kinetic model for the reaction of alpha-pinene and Delta(3)-carene with ozone and nitrogen oxides. AB - A kinetic model was compiled to simulate reactions of the monoterpenes, alpha pinene and Delta(3)-carene, with O(3), NO(2) and NO. The influence of different initial settings of O(3), NO(2) and NO on the monoterpene reaction was evaluated. At initial levels of 75 p.p.b. of O(3), NO(2) and NO each, 1.5% of alpha-pinene and 1.1% of Delta(3)-carene were calculated to react after 215 s. The corresponding experimental results showed that 9.3-12.2% of alpha-pinene and 9.9 11.7% of Delta(3)-carene reacted. The calculated levels of O(3), NO(2) and NO were compared to experimental measurements and were shown to correspond well. However, comparison of the amount of monoterpene reacted between calculated and experimental results, demonstrated that the calculations underestimated the amount of monoterpene reacted in the experimental chamber. The difference between experimental and calculated results could, e.g., be the effect of surfaces and the presence of water, which are parameters not included in the kinetic model known to have influence on these reactions. PRACTICAL IMPLICATIONS: A compiled kinetic model could be used to predict concentrations of O(3), NO(2), NO and to predict the effects on the monoterpene reaction in a Teflon reaction chamber when changing the initial concentrations of O(3), NO(2). However, the results in the present work show that surface and moisture effects have to be settled and the effects included in future models. PMID- 15663463 TI - Room chamber assessment of the pollutant emission properties of (nominally) low emission unflued gas heaters. AB - Pollutant emissions from unflued gas heaters were assessed in CSIRO's Room Dynamic Environmental Chamber. This paper describes the chamber assessment procedure and presents findings for major commercial heaters that are nominally "low-emission". The chamber was operated at controlled conditions of temperature, humidity, ventilation and air mixing, representative of those encountered in typical indoor environments. A fixed rate of heat removal from the chamber air ensured that the heaters operated at constant heating rates, typically approximately 6 MJ/h which simulated operation of a heater after warm-up in an insulated dwelling in south-east Australia. The pollutants assessed were nitrogen dioxide, carbon monoxide, formaldehyde, VOCs and respirable suspended particulates. One type of heater was lower emitting for nitrogen dioxide, but emitted greater amounts of carbon monoxide and formaldehyde (the latter becoming significant to indoor air quality). When operated with low line pressure or slight misalignment of the gas burner, this heater became a hazardous source of these pollutants. Emissions from the heaters changed little after continuous operation for up to 2 months. PRACTICAL IMPLICATIONS: Unflued gas heaters have been popular as primary heating sources in Australian homes for many years due to their ease of installation and energy efficiency, with approximately 600,000 now installed in housing and schools. However, with concerns over potential health impacts to occupants, manufacturers have reduced the nitrogen dioxide emissions from unflued gas heaters in Australia over recent years. They have done so with a target level for nitrogen dioxide in indoor air of 300 p.p.b. This is somewhat higher than the ambient air (and WHO) guideline of 110 p.p.b. Several studies of child respiratory health show an impact of unflued gas combustion products. A full characterization of the combustion products is needed under conditions that simulate heater operation in practice-this study was undertaken to provide such characterization. Key findings are that the focus needs to be on total gas emissions (not just nitrogen dioxide), and that heater installation can be very sensitive to small faults which lead to very high levels of toxic pollutants. These findings have influenced current government proposals for emission limits for these heaters. PMID- 15663464 TI - Ventilation efficiencies and thermal comfort results of a desk-edge-mounted task ventilation system. AB - In chamber experiments, we investigated the ventilation effectiveness and thermal comfort of a task ventilation system with an air supply nozzle located underneath the front edge of a desk and directing air towards a heated mannequin or a human volunteer seated at the desk. The task ventilation system provided outside air, while another ventilation system provided additional space cooling but no outside air. Test variables included the vertical angle of air supply (-15 degrees to 45 degrees from horizontal), and the supply flow rate of (3.5-6.5 l/s). Using the tracer gas step-up and step-down procedures, the measured air change effectiveness (i.e., exhaust air age divided by age of air in the breathing zone) in experiments with the mannequin ranged from 1.4 to 2.7 (median, 1.8), whereas with human subjects the air change effectiveness ranged from 1.3 to 2.3 (median, 1.6). The majority of the air change effectiveness values with the human subjects were less than values with the mannequin using comparable tests. Similarly, the tests run with supply air temperature equal to the room air temperature had lower air change effectiveness values than comparable tests with the supply air temperature lower ( approximately 5 degrees C) than the room air temperature. The air change effectiveness values are higher than typically reported for commercially-available task ventilation or displacement ventilation systems. Based on surveys completed by the subjects, operation of the task ventilation system did not cause thermal discomfort. PRACTICAL IMPLICATIONS: With a desk-edge mounted task ventilation system it is possible to obtain an increase in the effective ventilation rate of 50%. This could lead to reduced energy use. Also, this improvement can be gained while maintaining thermal comfort for occupants. Thus occupants can be thermally comfortable and save energy at the same time. PMID- 15663465 TI - VOC- and SVOC-emissions from adhesives, floor coverings and complete floor structures. AB - Emissions of volatile organic compounds (VOC) and semivolatile organic compounds (SVOC) from materials for flooring installation (primer, screed, adhesive, floor covering) were measured by means of emission test chambers and cells over a time period of at least 28 days at 23 degrees C, 50% relative humidity and an area specific air flow rate of q = 1.25 m(3)/m(2) h. Single components were tested in comparison to three complete structures (same concrete, primer, screed, adhesive) with different types of floor covering (PVC, carpet, linoleum). Sorption into concrete/screed and different permeability of flooring materials affected the emissions from the complete structures. The complete structures with linoleum and PVC showed the same types of emission and emission rates as the individual floor coverings themselves. Emissions from the carpet-covered structure resulted also from the lower layers. In all cases emissions from the complete structures were lower than the sum of emissions from the single components. For two adhesives the formation of secondary emissions (aldehydes and organic acids) was observed starting after the standard testing time of 28 days. PRACTICAL IMPLICATIONS: This paper gives a survey of possible emissions of VOCs and SVOCs from flooring materials and adhesives. On the example of these materials it is shown that the determination of SVOC-emissions from materials is important because after a few weeks the emission rates for SVOCs might be higher than for VOCs. In the real indoor environment SVOCs will be probably adsorbed to dust but by means of emission test chambers or cells the determination of emission rates from materials is possible. With the knowledge of this "emission potential" it is possible to estimate also the release of SVOCs into the (indoor) environment. PMID- 15663466 TI - Detection of single and mixed VOCs by smell and by sensory irritation. AB - We have measured complete concentration-detection (i.e., psychometric or detectability) functions to study the olfactory and ocular/nasal chemesthetic (a term that includes sensory irritation) impact of VOCs presented singly and in various binary mixtures. Such functions provide considerably more information than that provided by measuring only a "threshold". The outcome for single VOCs confirmed the much higher absolute sensitivity of olfaction compared to chemesthesis, but also demonstrated that the detection of ocular and nasal sensory irritation increases as a function of vapor concentration at a much higher rate than that for the detection of odor. The outcome for the binary mixtures revealed that, for both olfaction and chemesthesis, complete additivity of detection of individual components held at relatively low levels of detectability but broke down at higher levels. The breakdown for odor detection, compared to that for sensory irritation detection, was, first, more extensive, and, second, dependent to a larger extent on the degree of structural and chemical similarity/dissimilarity between the mixed VOCs. PRACTICAL IMPLICATIONS: Concentration-detection functions for the chemesthetic and olfactory detectability of VOCs have shown that, even when nasal pungency and eye irritation begin to be evoked at concentrations orders of magnitude larger than those evoking odor, they sharply increase in detectability to become clearly noticeable. In contrast, odor detectability increases with concentration at a much lower rate. As a result, any fixed reduction (e.g., 10-times) in the concentration of a VOC will reduce detectability of sensory irritation much more dramatically than detectability of odor, within their respective ranges. Concentration-detection functions are particularly informative when employed to probe into the rules of dose- and response-additivity in mixtures. Our results for olfaction, and to a lesser extent for chemesthesis, indicate that additivity of detection of individual VOCs in mixtures is level-dependent: as detectability increases, the degree of additivity decreases. This suggests that a substantial improvement of perceived air quality could follow from control of just the few dominating chemosensory sources. PMID- 15663467 TI - Reducing the ingress of urban noise through natural ventilation openings. AB - For buildings in busy urban areas affected by high levels of road traffic noise the potential to use natural ventilation can be limited by excessive noise entering through ventilation openings. This paper is concerned with techniques to reduce noise ingress into naturally ventilated buildings while minimizing airflow path resistance. A combined experimental and theoretical approach to the interaction of airflow and sound transmission through ventilators for natural ventilation applications is described. A key element of the investigation has been the development of testing facilities capable of measuring the airflow and sound transmission losses for a range of ventilation noise control strategies. It is demonstrated that a combination of sound reduction mechanisms -- one covering low frequency sound and another covering high frequency sound -- is required to attenuate effectively noise from typical urban sources. A method is proposed for quantifying the acoustic performance of different strategies to enable comparisons and informed decisions to be made leading to the possibility of a design methodology for optimizing the ventilation and acoustic performance of different strategies. PRACTICAL IMPLICATIONS: The need for employing techniques for combating low frequency sound in tandem with techniques for reducing high frequency sound in reducing the ingress of noise from urban sources such as road traffic to acceptable levels is demonstrated. A technique is proposed for enabling the acoustic and airflow performance of apertures for natural ventilation systems to be designed simultaneously. PMID- 15663468 TI - Mucous membrane and lower respiratory building related symptoms in relation to indoor carbon dioxide concentrations in the 100-building BASE dataset. AB - Indoor air pollutants are a potential cause of building related symptoms and can be reduced by increasing ventilation rates. Indoor carbon dioxide (CO(2)) concentration is an approximate surrogate for concentrations of occupant generated pollutants and for ventilation rate per occupant. Using the US EPA 100 office-building BASE Study dataset, we conducted multivariate logistic regression analyses to quantify the relationship between indoor CO(2) concentrations (dCO(2)) and mucous membrane (MM) and lower respiratory system (LResp) building related symptoms, adjusting for age, sex, smoking status, presence of carpet in workspace, thermal exposure, relative humidity, and a marker for entrained automobile exhaust. In addition, we tested the hypothesis that certain environmentally mediated health conditions (e.g., allergies and asthma) confer increased susceptibility to building related symptoms. Adjusted odds ratios (ORs) for statistically significant, dose-dependent associations (P < 0.05) for combined mucous membrane, dry eyes, sore throat, nose/sinus congestion, sneeze, and wheeze symptoms with 100 p.p.m. increases in dCO(2) ranged from 1.1 to 1.2. Building occupants with certain environmentally mediated health conditions were more likely to report that they experience building related symptoms than those without these conditions (statistically significant ORs ranged from 1.5 to 11.1, P < 0.05). PRACTICAL IMPLICATIONS: These results suggest that provision of sufficient per-person outdoor ventilation air, could significantly decrease prevalence of selected building related symptoms. The observed relationship between indoor minus outdoor CO(2) concentrations and mucous membrane and lower respiratory symptoms suggests that air contaminants are implicated in the etiology of building related symptoms. Levels of indoor air pollutants that are suspected to cause building related symptoms could be reduced by increasing ventilation rates, improving ventilation effectiveness, or reducing sources of indoor air pollutants, if known. PMID- 15663469 TI - Comparison of predicted and derived measures of volatile organic compounds inside four new relocatable classrooms. AB - Our objective was to develop a process for selecting interior finish materials having low impacts with respect to emissions of toxic and odorous volatile organic compounds (VOCs) for school relocatable classrooms (RCs). A laboratory study identified alternate materials with low VOC emissions. Two pairs of RCs then were constructed. One RC per pair contained standard interior materials; the other incorporated alternate materials. The pairs were sited side-by-side at two California elementary schools in fall 2001. Fifteen target VOCs, including the toxicants formaldehyde, acetaldehyde, vinyl acetate, phenol, toluene, and naphthalene, were measured during school hours over 8 weeks in the succeeding fall cooling season. Indoor minus outdoor VOC concentrations with an advanced HVAC operated were low; only formaldehyde concentrations exceeded 5 p.p.b. Classroom VOC concentrations were predicted based on emission factors (microg/m(2)/h), material quantities and design ventilation rates. These were compared to average adjusted concentrations measured when the occupied classrooms were operating at near the code-minimum ventilation rate. For 16 of the possible 42 comparisons, measured concentrations agreed within a factor of two of predicted the predicted values. Concentrations of six of 10 VOCs were significantly lower in modified RCs though average differences were mostly less than 1 p.p.b. PRACTICAL IMPLICATIONS: Laboratory-based material testing combined with modeling and field validation to select low VOC-impact interior finish materials helped achieve the aim of providing generally acceptable air quality in new school relocatable classrooms (RCs). The accuracy of the combined process was evidenced by the correct prediction of air quality impacts, though small, due to material VOC emissions when the study RCs were ventilated at code-minimum requirements. The process could be generalized to other manufacturers and classroom types. Material selection also is important to accommodate reduced ventilation rate conditions, which likely occur in many classrooms. PMID- 15663470 TI - Charge selectivity of the Cys-loop family of ligand-gated ion channels. AB - The determinants of charge selectivity of the Cys-loop family of ligand-gated ion channels have been studied for more than a decade. The investigations have mainly covered homomeric receptors e.g. the nicotinic acetylcholine receptor alpha7, the glycine receptor alpha1 and the serotonin receptor 5-HT(3A). Only recently, the determinants of charge selectivity of heteromeric receptors have been addressed for the GABA(A) receptor alpha2beta3gamma2. For all receptor subtypes, the selectivity determinants have been located to an intracellular linker between transmembrane domains M1 and M2. Two features of the M1-M2 linker appear to control ion selectivity. A central role for charged amino acid residues in selectivity has been almost universally observed. Furthermore, recent studies point to an important role of the size of the narrowest constriction in the pore. In the present review, these determinants of charge selectivity of the Cys-loop family of ligand-gated ion channels will be discussed in detail. PMID- 15663471 TI - Alzheimer's disease beta-secretase BACE1 is not a neuron-specific enzyme. AB - The brains of Alzheimer's disease (AD) patients are morphologically characterized by neurofibrillar abnormalities and by parenchymal and cerebrovascular deposits of beta-amyloid peptides. The generation of beta-amyloid peptides by proteolytical processing of the amyloid precursor protein (APP) requires the enzymatic activity of the beta-site APP cleaving enzyme 1 (BACE1). The expression of this enzyme has been localized to the brain, in particular to neurons, indicating that neurons are the major source of beta-amyloid peptides in brain. Astrocytes, on the contrary, are known to be important for beta-amyloid clearance and degradation, for providing trophic support to neurons, and for forming a protective barrier between beta-amyloid deposits and neurons. However, under certain conditions related to chronic stress, the role of astrocytes may not be beneficial. Here we present evidence demonstrating that astrocytes are an alternative source of BACE1 and therefore may contribute to beta-amyloid plaque formation. While resting astroyctes in brain do not express BACE1 at detectable levels, cultured astrocytes display BACE1 promoter activity and express BACE1 mRNA and enzymatically active BACE1 protein. Additionally, in animal models of chronic gliosis and in brains of AD patients, there is BACE1 expression in reactive astrocytes. This would suggest that the mechanism for astrocyte activation plays a role in the development of AD and that therapeutic strategies that target astrocyte activation in brain may be beneficial for the treatment of AD. Also, there are differences in responses to chronic versus acute stress, suggesting that one consequence of chronic stress is an incremental shift to different phenotypic cellular states. PMID- 15663472 TI - Nurr1 co-localizes with EphB1 receptors in the developing ventral midbrain, and its expression is enhanced by the EphB1 ligand, ephrinB2. AB - Both ephrins and the transcription factor, Nurr1, are critically involved in CNS development and, particularly, in the ontogenesis of the nigro-striatal system. Here we examined whether the ephrin receptor, EphB1, and Nurr1 share a similar expression pattern in the embryonic brain and whether expression of Nurr1 is under the control of EphB1 activation. The transcripts of EphB1 receptor and Nurr1 showed a similar pattern of expression in the ventral midbrain of mice at early stages of embryonic development (E11.5 and E12.5). At later stages (E15.5), only Nurr1 mRNA could still be detected in significant amounts in the A9-A10 regions of the ventral midbrain, whereas the two transcripts still showed a similar pattern of expression in discrete regions of the hindbrain. To examine whether activation of EphB1 receptor could induce the expression of Nurr1 in the ventral midbrain, we applied the EphB1 ligand, ephrinB2, to explants of embryonic mouse ventral midbrain. Low concentrations of clustered ephrinB2 (0.25 microg/mL) enhanced Nurr1 mRNA and protein levels, whereas higher concentrations were inactive. We conclude that activation of EphB1 receptors by appropriate concentrations of its ligand ephrinB2 might contribute to the acquisition of a dopaminergic fate in developing midbrain ventral neurones. PMID- 15663473 TI - Involvement of 3',5'-cyclic adenosine monophosphate-dependent protein kinase in regulation of Fos expression and tyrosine hydroxylase levels during morphine withdrawal in the hypothalamic paraventricular nucleus and medulla oblongata catecholaminergic cell groups. AB - Morphine withdrawal stimulates the hypothalamic-pituitary-adrenocortical axis activity by activation of nucleus tractus solitarius (NTS)/ventrolateral medulla (VLM) noradrenergic pathways innervating the hypothalamic paraventricular nucleus (PVN). We investigated whether cAMP-dependent protein kinase (PKA) plays a role in this process by estimating changes in PKA immunoreactivity and the influence of inhibition of PKA on Fos protein expression and tyrosine hydroxylase (TH) immunoreactivity levels in the PVN and NTS/VLM during morphine withdrawal. Dependence on morphine was induced by a 7-day s.c. implantation of morphine pellets. Morphine withdrawal was precipitated on day 8 by an injection of naloxone (5 mg/kg s.c.). When opioid withdrawal was precipitated, an increase in PKA immunoreactivity levels was observed 90 min after naloxone administration in the PVN and NTS/VLM areas. Morphine withdrawal induced expression of Fos in the PVN and NTS/VLM, indicating an activation of neurones in those nuclei. TH immunoreactivity in NTS/VLM was increased 90 min after induction of morphine withdrawal, whereas there was a decrease in TH levels in the PVN at the same time point. When the selective PKA inhibitor HA-1004 was infused it greatly diminished the Fos expression observed in morphine-withdrawn rats. Furthermore, the changes in TH immunoreactivity were significantly modified by infusion of HA-1004. The present findings suggest that an up-regulated PKA-dependent transduction pathway might contribute to the activation of the hypothalamic-pituitary-adrenocortical axis in response to morphine withdrawal. PMID- 15663474 TI - Protein glycation, oxidation and nitration adduct residues and free adducts of cerebrospinal fluid in Alzheimer's disease and link to cognitive impairment. AB - Increased damage to proteins by glycation, oxidation and nitration has been implicated in neuronal cell death leading to Alzheimer's disease (AD). Protein glycation, oxidation and nitration adducts are consequently formed. Quantitative screening of these adducts in CSF may provide a biochemical indicator for the diagnosis of AD. To assess this, we measured 11 glycation adducts, three oxidation adducts and a nitration adduct, determining both protein adduct residues and free adducts, in CSF samples of age-matched normal healthy subjects (n = 18) and subjects with Alzheimer's disease (n = 32). In CSF protein, the concentrations of 3-nitrotyrosine, N(epsilon)-carboxymethyl-lysine, 3 deoxyglucosone-derived hydroimidazolone and N-formylkynurenine residues were increased in subjects with Alzheimer's disease. In CSF ultrafiltrate, the concentrations of 3-nitrotyrosine, methylglyoxal-derived hydroimidazolone and glyoxal-derived hydroimidazolone free adducts were also increased. The Mini Mental State Examination (MMSE) score correlated negatively with 3-nitrotyrosine residue concentration (p < 0.05), and the negative correlation with fructosyl lysine residues just failed to reach significance (p = 0.052). Multiple linear regression gave a regression model of the MMSE score on 3-nitrotyrosine, fructosyl-lysine and N(epsilon)-carboxyethyl-lysine residues with p-values of 0.021, 0.031 and 0.052, respectively. These findings indicate that protein glycation, oxidation and nitration adduct residues and free adducts were increased in the CSF of subjects with Alzheimer's disease. A combination of nitration and glycation adduct estimates of CSF may provide an indicator for the diagnosis of Alzheimer's disease. PMID- 15663475 TI - Neuronal precursor-specific activity of a human doublecortin regulatory sequence. AB - The doublecortin (DCX) gene encodes a 40-kDa microtubule-associated protein specifically expressed in neuronal precursors of the developing and adult CNS. Due to its specific expression pattern, attention was drawn to DCX as a marker for neuronal precursors and neurogenesis, thereby underscoring the importance of its promoter identification and promoter analysis. Here, we analysed the human DCX regulatory sequence and confined it to a 3.5-kb fragment upstream of the ATG start codon. We demonstrate by transient transfection experiments that this fragment is sufficient and specific to drive expression of reporter genes in embryonic and adult neuronal precursors. The activity of this regulatory fragment overlapped with the expression of endogenous DCX and with the young neuronal markers class III beta-tubulin isotype and microtubule-associated protein Map2ab but not with glial or oligodendroglial markers. Electrophysiological data further confirmed the immature neuronal nature of these cells. Deletions within the 3.5 kb region demonstrated the relevance of specific regions containing transcription factor-binding sites. Moreover, application of neurogenesis-related growth factors in the neuronal precursor cultures suggested the lack of direct signalling of these factors on the DCX promoter construct. PMID- 15663476 TI - Diminished Akt phosphorylation in neurons lacking glutathione peroxidase-1 (Gpx1) leads to increased susceptibility to oxidative stress-induced cell death. AB - We have previously identified an increased susceptibility of glutathione peroxidase-1 (Gpx1)-/- mice to neuronal apoptosis following mid-cerebral artery (MCA) occlusion. This study was designed to elucidate the mechanisms involved in elevated neuronal cell death arising from an altered endogenous oxidant state. This was addressed in both an in vitro and in vivo model of oxidative stress in the form of exogenous H2O2 and cerebral ischaemia, respectively. Increased levels of cell death were detected in primary neurons lacking Gpx1 following the addition of exogenous H2O2. This increased apoptosis correlated with a down regulation in the activation of the phospho-inositide 3-kinase [PI3K]-Akt survival pathway. The importance of this pathway in protecting against H2O2 induced cell death was highlighted by the increased susceptibility of wildtype neurons to apoptosis when treated with the PI3K inhibitor, LY294002. The Gpx1-/- mice also demonstrated elevated neuronal cell death following MCA occlusion. Although Akt phosphorylation was detected in the Gpx1-/- brains, activation was not seen in later reperfusion events, as demonstrated in wildtype brains. Previous studies have highlighted the importance of Akt phosphorylation in protecting against neuronal cell death following cerebral ischaemia-reperfusion. Our results suggest that the increased susceptibility of Gpx1-/- neurons to H2O2 induced apoptosis and neuronal cell death in vivo following cerebral ischaemia reperfusion injury can be attributed in part to diminished activation of Akt. Perturbations in key anti-apoptotic mechanisms as a result of an altered redox state may have implications in the study of oxidative stress-mediated neuropathologies. PMID- 15663477 TI - Presenilin 2 familial Alzheimer's disease mutations result in partial loss of function and dramatic changes in Abeta 42/40 ratios. AB - Gene knockout studies in mice suggest that presenilin 1 (PS1) is the major gamma secretase and that it contributes disproportionately to amyloid beta (Abeta) peptide generation from beta-amyloid precursor protein (APP), whereas PS2 plays a more minor role. Based on this and other observations we hypothesized that familial Alzheimer's disease (FAD) mutations in PS2 would have a dramatic effect on function in order to have an observable effect on Abeta levels in the presence of normal PS1 alleles. Only four of the eight reported FAD mutations in PS2 have altered function in vitro suggesting that the other variants represent rare polymorphisms rather than disease-causing mutations. In support of our hypothesis, the four verified PS2 FAD mutations cause substantial changes in the Abeta 42/40 ratio, comparable with PS1 mutations that cause very-early-onset FAD. Most of the PS2 mutations also cause a significant decrease in Abeta 40, APP C terminal fragment (CTF)gamma and Notch intracellular domain (NICD) production suggesting that they are partial loss of function mutations. PS2 M239V, its PS1 homolog M233V, and other FAD mutations within transmembrane (TM) 5 of PS1 differentially affect CTFgamma and NICD production suggesting that TM5 of PS are important for gamma-secretase cleavage of APP but not Notch. PMID- 15663478 TI - Inhibition of the alpha-ketoglutarate dehydrogenase complex by the myeloperoxidase products, hypochlorous acid and mono-N-chloramine. AB - Abstract alpha-Ketoglutarate dehydrogenase (KGDHC) complex activity is diminished in a number of neurodegenerative disorders and its diminution in Alzheimer Disease (AD) is thought to contribute to the major loss of cerebral energy metabolism that accompanies this disease. The loss of KGDHC activity appears to be predominantly due to post-translation modifications. Thiamine deficiency also results in decreased KGDHC activity and a selective neuronal loss. Recently, myeloperoxidase has been identified in the activated microglia of brains from AD patients and thiamine-deficient animals. Myeloperoxidase produces a powerful oxidant, hypochlorous acid that reacts with amines to form chloramines. The aim of this study was to investigate the ability of hypochlorous acid and chloramines to inhibit the activity of KGDHC activity as a first step towards investigating the role of myeloperoxidase in AD. Hypochlorous acid and mono-N-chloramine both inhibited purified and cellular KGDHC and the order of inhibition of the purified complex was hypochlorous acid (1x) > mono-N-chloramine (approximately 50x) > hydrogen peroxide (approximately 1,500). The inhibition of cellular KGDHC occurred with no significant loss of cellular viability at all exposure times that were examined. Thus, hypochlorous acid and chloramines have the potential to inactivate a major target in neurodegeneration. PMID- 15663479 TI - Different properties of the central and peripheral forms of human tryptophan hydroxylase. AB - Tryptophan hydroxylase (TPH) catalyses the rate-limiting reaction in the biosynthesis of serotonin. In humans, two different TPH genes exist, located on chromosomes 11 and 12, respectively, and encoding two enzymes (TPH1 and TPH2) with an overall sequence identity of 71%. We have expressed both enzymes as various fusion proteins in Escherichia coli and using an in vitro transcription/translation system, and compared their solubility and kinetic properties. TPH2 is more soluble than TPH1, has a higher molecular weight and different kinetic properties, including a lower catalytic efficiency towards phenylalanine than TPH1. Both enzymes are phosphorylated by cAMP-dependent protein kinase A. TPH2 was phosphorylated at Ser19, a phosphorylation site not present in TPH1. The differences between TPH1 and TPH2 have important implications for the regulation of serotonin production in the brain and the periphery and may provide an explanation for some of the diverging results reported for TPH from different sources in the past. PMID- 15663480 TI - Role of basic region leucine zipper transcription factors cyclic AMP response element binding protein (CREB), CREB2, activating transcription factor 2 and CAAT/enhancer binding protein alpha in cyclic AMP response element-mediated transcription. AB - The transcription factor cAMP response element binding protein (CREB), a member of the basic region leucine zipper (bZIP) family of proteins, is the major cAMP response element (CRE) binding. Other bZIP proteins, including CREB2, activating transcription factor 2 (ATF2), or CAAT/enhancer binding protein (C/EBP) have been reported to transactivate CRE-containing genes or to interfere with transactivation by CREB. We have designed a simple transactivation assay using expression of either a constitutively active CREB mutant or a nuclear targeted mutant of the catalytic subunit of cAMP-dependent protein kinase. In both cases, a striking stimulation of transcription of CRE-containing reporter genes was observed in noradrenergic locus coeruleus-like CATH.a cells. In addition, a constitutively active mutant of ATF2 specifically transactivated a secretogranin II promoter/luciferase reporter gene, but had no effect on the tyrosine hydroxylase promoter. In contrast, CREB2 and C/EBPalpha did not transactivate CRE containing reporter genes, indicating that these bZIP proteins target distinct genetic elements. Experiments involving dominant-negative bZIP mutants revealed that CREB does not heterodimerize with CREB2, ATF2, c-Jun or C/EBP. Rather, CREB and ATF2 compete for binding to the CRE, and are independently able to up regulate transcription of genes containing CRE motifs in their regulatory regions. PMID- 15663481 TI - Molecular mechanisms involved in the adenosine A and A receptor-induced neuronal differentiation in neuroblastoma cells and striatal primary cultures. AB - Adenosine A1 receptors (A1Rs) and adenosine A(2A) receptors (A(2A)Rs) are the major mediators of the neuromodulatory actions of adenosine in the brain. In the striatum A1Rs and A(2A)Rs are mainly co-localized in the GABAergic striatopallidal neurons. In this paper we show that agonist-induced stimulation of A1Rs and A(2A)Rs induces neurite outgrowth processes in the human neuroblastoma cell line SH-SY5Y and also in primary cultures of striatal neuronal precursor cells. The kinetics of adenosine-mediated neuritogenesis was faster than that triggered by retinoic acid. The triggering of the expression of TrkB neurotrophin receptor and the increase of cell number in the G1 phase by the activation of adenosine receptors suggest that adenosine may participate in early steps of neuronal differentiation. Furthermore, protein kinase C (PKC) and extracellular regulated kinase-1/2 (ERK-1/2) are involved in the A1R- and A(2A)R mediated effects. Inhibition of protein kinase A (PKA) activity results in a total inhibition of neurite outgrowth induced by A(2A)R agonists but not by A1R agonists. PKA activation is therefore necessary for A(2A)R-mediated neuritogenesis. Co-stimulation does not lead to synergistic effects thus indicating that the neuritogenic effects of adenosine are mediated by either A1 or A(2A) receptors depending upon the concentration of the nucleoside. These results are relevant to understand the mechanisms by which adenosine receptors modulate neuronal differentiation and open new perspectives for considering the use of adenosine agonists as therapeutic agents in diseases requiring neuronal repair. PMID- 15663482 TI - Direct interaction of myosin regulatory light chain with the NMDA receptor. AB - NMDA receptors interact with a variety of intracellular proteins at excitatory synapses. In this paper we show that myosin regulatory light chain (RLC) isolated from mouse brain is a NMDA receptor-interacting protein. Myosin RLC bound directly to the C-termini of both NMDA receptor 1 (NR1) and NMDA receptor 2 (NR2) subunits, rendering the interaction of myosin RLC with NMDA receptors distinct from that of calmodulin which is considered a NR1-interacting protein. Myosin RLC co-localized with NR1 in the dendritic spines of isolated hippocampal neurons, and was co-immunoprecipitated from brain extracts in a complex with NR1, NR2A, NR2B, PSD-95, Adaptor protein-2 and myosin II heavy chain. The C0 region of NR1 was necessary and sufficient for binding myosin RLC. Ca2+/calmodulin, but not calmodulin alone, displaced recombinant myosin RLC from the carboxy tail of NR1 indicating that myosin RLC and Ca2+/calmodulin can compete for a common binding site on NR1 in vitro. Myosin RLC is the only known substrate for myosin regulatory light chain kinase, which has recently been shown to modulate NMDA receptor function in isolated hippocampal neurons. Our results suggest that an additional level of NMDA receptor regulation may be mediated via a direct interaction with a light chain of myosin II. Thus, myosin RLC-NMDA receptor interactions may contribute to the contractile and motile forces that are placed upon NMDA receptor subunits during changes associated with synaptic plasticity and neural morphogenesis. PMID- 15663483 TI - An RNAi strategy for treatment of amyotrophic lateral sclerosis caused by mutant Cu,Zn superoxide dismutase. AB - Amyotrophic lateral sclerosis (ALS or Lou Gehrig's disease) is a neurodegenerative disease characterized by motor neuron degeneration, paralysis and death. One cause of this disease is mutations in the Cu,Zn superoxide dismutase (SOD1) gene. As mutant SOD1 acquires a toxic property that kills motor neurons, by reducing the mutant protein the disease progression may be slowed or prevented. While mutant SOD1 is toxic, the wild-type SOD1 is indispensable for motor neuron health. Therefore, the ideal therapeutic strategy would be to inhibit selectively the mutant protein expression. Previously we have demonstrated that RNA interference (RNAi) can selectively inhibit some mutant SOD1 expression. However, more than 100 SOD1 mutants can cause ALS and all mutants cannot be inhibited selectively by RNAi. To overcome this obstacle, we have designed a replacement RNAi strategy. Using this strategy, all mutants and wild-type genes are inhibited by RNAi. The wild-type SOD1 function is then replaced by designed wild-type SOD1 genes that are resistant to the RNAi. Here we demonstrate the concept of this strategy. PMID- 15663484 TI - Stimulation of the locus coeruleus elicits noradrenaline and dopamine release in the medial prefrontal and parietal cortex. AB - Our previous studies have suggested that dopamine and noradrenaline may be coreleased from noradrenergic nerve terminals in the cerebral cortex. To further clarify this issue, the effect of electrical stimulation of the locus coeruleus on extracellular noradrenaline, dopamine and DOPAC in the medial prefrontal cortex, parietal cortex and caudate nucleus was analysed by microdialysis in freely moving rats. Stimulation of the locus coeruleus for 20 min with evenly spaced pulses at 1 Hz failed to modify cortical catecholamines and DOPAC levels. Stimulation with bursts of pulses at 12 and 24 Hz increased, in a frequency related manner, not only noradrenaline but also dopamine and DOPAC in the two cortices. In both cortices noradrenaline returned to baseline within 20 min of stimulation, irrespective of the stimulation frequency, whereas dopamine returned to normal within 20 and 60 min in the medial prefrontal cortex and within 60 and 80 min in the parietal cortex after 12 and 24 Hz stimulation, respectively. DOPAC remained elevated throughout the experimental period. Phasic stimulation of the locus coeruleus at 12 Hz increased noradrenaline in the caudate nucleus as in the cerebral cortices but was totally ineffective on dopamine and DOPAC. Tetrodotoxin perfusion into the medial prefrontal cortex dramatically reduced noradrenaline and dopamine levels and suppressed the effect of electrical stimulation. These results indicate that electrical stimulation-induced increase of dopamine is a nerve impulse exocytotic process and suggest that cortical dopamine and noradrenaline may be coreleased from noradrenergic terminals. PMID- 15663485 TI - Mutagenesis analysis of the serotonin 5-HT2C receptor and a Caenorhabditis elegans 5-HT2 homologue: conserved residues of helix 4 and helix 7 contribute to agonist-dependent activation of 5-HT2 receptors. AB - An alignment of serotonin [5-hydroxytryptamine (5-HT)] G protein-coupled receptors identified a lysine at position 4.45 (helix 4) and a small polar residue (serine or cysteine) at 7.45 (helix 7) that occur exclusively in the 5 HT2 receptor family. Other serotonin receptors have a hydrophobic amino acid, typically a methionine, at 4.45 and an invariant asparagine at 7.45. The functional significance of these class-specific substitutions was tested by site directed mutagenesis of two distantly related 5-HT2 receptors, Caenorhabditis elegans 5-HT2ce and rat 5-HT2C. Residues 4.45 and 7.45 were each mutated to a methionine and asparagine, respectively, or an alanine and the resulting constructs were tested for activity. A K4.45M mutation decreased serotonin dependent activity (Emax) of the rat 5-HT2C receptor by 60% and that of the C. elegans homologue by 40%, as determined by a fluorometric plate-based calcium assay. The rat mutant also exhibited nearly sixfold higher agonist binding affinity and significantly lower constitutive activity compared with wildtype. Mutagenesis of S7.45 in the C. elegans receptor increased serotonin binding affinity by up to 25-fold and decreased Emax by up to 65%. The same mutations of the cognate C7.45 in rat 5-HT2C produced a smaller fourfold change in the affinity for serotonin and decreased agonist efficacy by up to 50%. Substitutions of S/C7.45 did not produce a significant change in the basal activity of either receptor. All mutants tested exhibited levels of receptor expression similar to the corresponding wildtype based on measurements of specific [3H]-mesulergine binding or flow cytometry analyses. Taken together, these results suggest that K4.45 and S/C7.45 play an important role in the conformational rearrangements leading to agonist-induced activation of 5-HT2 receptors. PMID- 15663486 TI - Calcium-dependent dephosphorylation of brain mitochondrial calcium/cAMP response element binding protein (CREB). AB - Calcium-mediated signaling regulates nuclear gene transcription by calcium/cAMP response element binding protein (CREB) via calcium-dependent kinases and phosphatases. This study tested the hypothesis that CREB is also present in mitochondria and subject to dynamic calcium-dependent modulation of its phosphorylation state. Antibodies to CREB and phosphorylated CREB (pCREB) were used to demonstrate the presence of both forms in isolated mitochondria and mitoplasts from rat brain. When energized mitochondria were exposed to increasing concentrations of Ca2+ in the physiological range, pCREB was lost while total CREB remained constant. In the presence of Ru360, an inhibitor of the mitochondrial Ca2+ uptake uniporter, calcium-dependent loss of pCREB levels was attenuated, suggesting that intramitochondrial calcium plays an important role in pCREB dephosphorylation. pCREB dephosphorylation was not, however, inhibited by the phosphatase inhibitors okadaic acid and Tacrolimus. In the absence of Ca2+, CREB phosphorylation was elevated by the addition of ATP to the mitochondrial suspension. Exposure of mitochondria to the pore-forming molecule alamethicin that causes osmotic swelling and release of intermembrane proteins enriched mitochondrial pCREB immunoreactivity. These results further suggest that mitochondrial CREB is located in the matrix or inner membrane and that a kinase and a calcium-dependent phosphatase regulate its phosphorylation state. PMID- 15663487 TI - Thyroid hormone regulates Galphai1 gene expression in the rat cerebellar cortex during post-natal development. AB - Thyroid hormone regulates the expression of G protein in tissues such as fat and heart. In the brain, very little information is available relative to the regulation by thyroid hormone of G proteins. Here, we show that the expression of the Galphai1 gene is induced by thyroid hormones in the rat cerebellum during development. Hence, the levels of Galphai1 transcripts and protein were decreased in the cerebellum of hypothyroid neonates. In situ hybridization studies showed that the neurons of the cerebellar cortex, particularly Purkinje cells, were affected. Surprisingly, and in contrast with the in vivo stimulatory effect described above, thyroid hormone repressed the activity of the rat Galphai1 promoter in vitro, suggesting that the effect of this hormone in the cerebellum is indirect. In this regard, we present data suggesting that the transcription factor C/EBPbeta could be implicated. First, there are active CEBP binding sites in the Galphai1 promoter. Second, we have found a diminished DNA binding activity of hypothyroid nuclear proteins to a Galphai1 promoter sequence containing a C/EBP binding site. Third, this complex is likely to contain C/EBPbeta protein as it is displaced by specific anti-C/EBPbeta antibodies. Finally, there is a significant decrease in the C/EBPbeta protein content in the hypothyroid cerebellar cortex. PMID- 15663488 TI - Group I and II metabotropic glutamate receptors alter brain cortical metabolic and glutamate/glutamine cycle activity: a 13C NMR spectroscopy and metabolomic study. AB - Metabotropic glutamate receptors (mGluR) modulate neuronal function. Here, we tested the effect on metabolism of a range of Group I and II mGluR ligands in Guinea pig brain cortical tissue slices, applying 13C NMR spectroscopy and metabolomic analysis using multivariate statistics. The effects of Group I agonists (S)-3,5-dihydroxyphenylglycine (DHPG) and (RS)-2-chloro-5 hydroxyphenylglycine (CHPG) depended upon concentration and were mostly stimulatory, increasing both net metabolic flux through the Krebs cycle and glutamate/glutamine cycle activity. Only the higher (50 microm) concentrations of CHPG had the opposite effect. The Group I antagonist (RS)-1-aminoindan-1,5 dicarboxylic acid (AIDA), consistent with its neuroprotective role, caused significant decreases in metabolism. With principal components analysis of the metabolic profiles generated by these ligands, the effects could be separated by two principal components. Agonists at Group II mGluR [(2S,2'R,3'R)-2-(2',3' dicarboxycyclopropyl)glycine (DCG IV) and 2R,4R-4-aminopyrrolidine-2,4 dicarboxylate (APDC)] generally stimulated metabolism, including glutamate/glutamine cycling, although this varied with concentration. The antagonist (2S)-alpha-ethylglutamic acid (EGLU) stimulated astrocyte metabolism with minimal impact on glutamate/glutamine cycling. (RS)-1-Aminophosphoindan-1 carboxylic acid (APICA) decreased metabolism at 5 microm but had a stimulatory effect at 50 microm. All ligand effects were separated from control and from each other using two principal components. The ramifications of these findings are discussed. PMID- 15663489 TI - Brain response to traumatic brain injury in wild-type and interleukin-6 knockout mice: a microarray analysis. AB - Traumatic injury to the brain is one of the leading causes of injury-related death or disability. Brain response to injury is orchestrated by cytokines, such as interleukin (IL)-6, but the full repertoire of responses involved is not well known. We here report the results obtained with microarrays in wild-type and IL-6 knockout mice subjected to a cryolesion of the somatosensorial cortex and killed at 0, 1, 4, 8 and 16 days post-lesion. Overall gene expression was analyzed by using Affymetrix genechips/oligonucleotide arrays with approximately 12,400 probe sets corresponding to approximately 10,000 different murine genes (MG_U74Av2). A robust, conventional statistical method (two-way anova) was employed to select the genes significantly affected. An orderly pattern of gene responses was clearly detected, with genes being up- or down-regulated at specific timings consistent with the processes involved in the initial tissue injury and later regeneration of the parenchyma. IL-6 deficiency showed a dramatic effect in the expression of many genes, especially in the 1 day post-lesion timing, which presumably underlies the poor capacity of IL-6 knockout mice to cope with brain damage. The results highlight the importance of IL-6 controlling the response of the brain to injury as well as the suitability of microarrays for identifying specific targets worthy of further study. PMID- 15663490 TI - Changing pattern of imported cutaneous leishmaniasis in the Netherlands. AB - Cutaneous leishmaniasis (CL) in western countries seems to be appearing more frequently. Our aim was to determine if there has been a shift in countries where CL is acquired and whether the incidence has changed, and to assess current diagnostic procedures and treatment modalities. In a retrospective study medical records of patients with the diagnosis of CL at the Departments of Tropical Dermatology and Tropical Medicine, Academic Medical Center, Amsterdam, the Netherlands, from 1990 to 2000 were analysed. CL was diagnosed in 78 patients. The majority was acquired in Belize, Surinam, French Guyana and Bolivia. Giemsa stains were positive for the parasite in impression smears from 43% and in biopsies from 71%. Seventy-eight per cent of cases were culture-positive and 89% were PCR-positive. Sixty-two patients were treated systemically: pentavalent antimony (32), pentamidine isetionate (11), itraconazole (19), and 13 locally, the majority with a combination of cryosurgery and intralesional pentavalent antimony. Imported CL is becoming more frequent, with South and Middle American countries being important sources of infection. Multiple tests, of which PCR is the most sensitive, are required to confirm the diagnosis. Systemic treatment was given to the majority of the patients. PMID- 15663491 TI - Quality of life issues for South Africans with acne vulgaris. AB - The adverse effects of acne on the psyche have been established in patients from 'first world' countries. There has been no in depth study in predominantly black patients from Africa addressing this issue. This was a prospective cross sectional study of acne patients attending a dermatology unit in KwaZulu-Natal, South Africa. A questionnaire was completed and acne graded by the Global Acne Grading scale. Psychological morbidity and quality of life (QOL) were assessed by the General Health Questionnaire and Dermatology Specific Quality of Life Questionnaires, respectively. We found that clinical severity was not associated with patient perception or psychological distress. The QOL measures such as feelings, social activities, performance at work or school, activities of daily living and overall mental health were found to be associated with distress with associated P-values of 0.0002, 0.0168, 0.0032, 0.033 and < 0.0001, respectively. The severity of acne was not associated with psychological distress. Painful and bleeding lesions were associated with distress levels; P = 0.042 and P = 0.019, respectively. In conclusion, South African patients with acne vulgaris suffer significant psychological distress, which affects the quality of their lives. PMID- 15663492 TI - Staphylococcus aureus re-colonization in atopic dermatitis: beyond the skin. AB - Patients with atopic dermatitis (AD) are often heavily colonized by Staphylococcus aureus, which adversely affects eczema severity. Strategies to control S. aureus in AD include antibiotic and or antiseptics. However long-term efficacy is unclear. In this study we consider extra-cutaneous factors that may cause S. aureus re-colonization in adult AD. Twenty-one patients with AD were recruited and were assessed for: duration of AD, use of topical or oral antibiotic within the preceding 3 months, the number of hospital admissions during the preceding year and current treatment. The types of topical treatments used, vehicle, container and the expiry dates were also recorded. The severity of AD was assessed by SCORAD index. Microbiological assessment for S. aureus carriage from affected skin, anterior nares, emollient and topical steroid was undertaken using culture, Staphaurex test and antibiotic resistance. Of the patients 86% had S. aureus colonization. The median SCORAD score were greater in those colonized with S. aureus (P = 0.02) and those with contaminated treatments (P = 0.05). Prior antibiotic treatment, prior hospital admission and nasal carriage did not influence the median SCORAD. Three extra-cutaneous mechanisms by which S. aureus can re-colonize the skin were identified: antibiotic resistance, nasal carriage and treatment contamination. PMID- 15663493 TI - Randomized, double-blind, placebo-controlled pilot study to assess the value of free radical scavengers in reducing inflammation induced by cryotherapy. AB - The inflammation which follows cryotherapy is a significant disadvantage of this therapeutic modality. To date, the only treatment shown to reduce this inflammation is application of topical corticoids. We have therefore conducted a pilot study to investigate whether pretreatment with the free radical scavengers, vitamins C and E might alleviate the signs and symptoms of inflammation following liquid nitrogen cryotherapy of common warts. We undertook a randomized, double blind, placebo-controlled, parallel group study. We recruited 40 adult patients, of whom 38 returned for evaluation. Treatments comprised vitamin C (2000 mg) and vitamin E (800 IU) daily or matching placebo for 7 days prior to cryotherapy to a hand wart. Oedema volume, erythema level, pain intensity and the presence or absence of blistering were assessed 24 h after cryotherapy. There were no significant differences between the two treatment groups in any of the parameters assessed. This study yielded no suggestion of benefit from the use of pretreatment with free radical scavengers in conjunction with liquid nitrogen cryotherapy. PMID- 15663494 TI - Pigmented seborrheic keratoses of the vulva clinically mimicking a malignant melanoma: a clinical, dermoscopic-pathologic case study. AB - The diagnosis of seborrheic keratosis is, in general, a clinical one, but in some cases, the differential diagnosis between pigmented seborrheic keratosis and malignant melanoma is difficult. Dermoscopy may improve the early diagnosis of vulvar melanoma and thus play a role in the preoperative classification of pigmented lesions at this particular site. We report the first case of a pigmented seborrheic keratosis of the vulva clinically mimicking a malignant melanoma, whose dermoscopic features have been investigated together with their pathologic correlates. Dermoscopically our case shows the absence of comedo-like openings and the presence of the pseudo-network. Dermoscopy is therefore a useful method for the differential diagnosis of pigmented lesions even in the vulva. PMID- 15663495 TI - Spontaneous regression of subungual keratoacanthoma with reossification of underlying distal lytic phalynx. AB - A 45-year-old man presented with a rapidly enlarging keratotic lesion of the distal subungual right middle finger. An X-ray of the digit revealed a well defined cup-shaped lytic lesion of the phalynx underlying the subungual nodule. The lesion resolved spontaneously with reossification of the underlying bony defect. The clinical history and X-ray is consistent with a diagnosis of spontaneously resolving subungual keratoacanthoma. PMID- 15663496 TI - The use of a novel biodegradable preparation capable of the sustained release of bacteriophages and ciprofloxacin, in the complex treatment of multidrug-resistant Staphylococcus aureus-infected local radiation injuries caused by exposure to Sr90. AB - In December 2001, three Georgian lumberjacks from the village of Lia were exposed to a strontium-90 source from two Soviet-era radiothermal generators they found near their village. In addition to systemic effects, two of them developed severe local radiation injuries which subsequently became infected with Staphylococcus aureus. After hospitalization in Tbilisi, Georgia, the patients were treated with various medications, including antibiotics and topical ointments; however, wound healing was only moderately successful, and their S. aureus infection could not be eliminated. Approximately 1 month after hospitalization, treatment with PhagoBioDerm (a wound-healing preparation consisting of a biodegradable polymer impregnated with ciprofloxacin and bacteriophages) was initiated. Purulent drainage stopped within 2-7 days. Clinical improvement was associated with rapid (7 days) elimination of the aetiologic agent, a strain of S. aureus resistant to many antibiotics (including ciprofloxacin), but susceptible to the bacteriophages contained in the PhagoBioDerm preparation. PMID- 15663497 TI - PNP with Waldenstrom's macroglobulinaemia. AB - We describe a 65-year-old Caucasian man with Waldenstrom's macroglobulinaemia who developed paraneoplastic pemphigus (PNP) 3 years after his haematological diagnosis. This is a very rare malignancy that is associated with PNP. The evolution of PNP in this patient appears to exhibit the postulated immunological phenomenon of epitope spreading. PMID- 15663498 TI - Linear basaloid follicular hamartoma on the Blaschko's line of the face. AB - We report a 5-year-old girl with localized basaloid follicular hamartoma. The lesion consisted of a linear, hypopigmented, atrophic patch on the right lateral side of nose. Of great interest is that the linear distribution of lesion may follow the Blaschko's line of the face. PMID- 15663499 TI - Three cases of focal dermal hypoplasia (Goltz syndrome). AB - Focal dermal hypoplasia or Goltz syndrome is a rare genodermatosis, characterized by multiple abnormalities of ectodermal and mesodermal origin. It is found predominantly in females and is characterized by hypoplasia of skin and papillomas. Three cases of focal dermal hypoplasia in infancy with unusual inheritance patterns are reported. Cutaneous features were atrophic reticulated scars involving the trunk and extremities following the lines of Blaschko. Papillomas were present on the genitalia and in a periorificial distribution. Skeletal abnormalities included syndactyly, polydactyly and lobster claw deformities. Ophthalmological examination revealed strabismus and retinal colobomas. PMID- 15663500 TI - Verrucous trichilemmal cyst containing human papillomavirus. AB - Verrucous cyst is an uncommon nonplantar epidermoid (infundibular) cyst with verrucous changes in its wall. We present the first case of trichilemmal cyst (isthmus-catagen cyst) with verrucous changes of its squamous lining. PMID- 15663501 TI - A rare case of the cutaneous form of adult T-cell leukaemia/lymphoma: assessment of remission by PCR for clonal T-cell receptor gamma gene rearrangements in an electron beam-irradiated cutaneous lesion. AB - Adult T-cell leukaemia/lymphoma is a lymphoproliferative disorder aetiologically associated with human T-cell lymphotropic virus type I infection. A cutaneous lesion often develops in the disease, and in rare cases, is even the only manifestation. Here we report a rare case of 'cutaneous' adult T-cell leukaemia/lymphoma with neither atypical cells in the peripheral blood nor lymph node involvement. All nodular lesions were completely eliminated after local electron beam irradiation (20 Gy/nodule in total). To evaluate whether or not there were residual lymphoma cells in the skin, we performed PCR to detect clonal T cell receptor gamma gene rearrangements. The sample from the nodule before irradiation showed evidence of a rearranged band, which was not detected at the same site after treatment nor in any peripheral blood. The findings suggest that this procedure is useful for the evaluation of therapeutic effects and the early detection of lymphoma recurrence. PMID- 15663502 TI - Decline of anti-desmoglein 1 IgG ELISA scores by withdrawal of D-penicillamine in drug-induced pemphigus foliaceus. AB - A case of pemphigus foliaceus arising during the administration of D penicillamine was reported. A 72-year-old woman with systemic sclerosis developed flaccid vesicles and crusted erythema of the skin after administration of D penicillamine for 1-year. She was positive for anti-desmoglein 1 IgG by ELISA. The withdrawal of D-penicillamine resulted in improvement of the skin lesions and ELISA scores for anti-demoglein 1 antibodies revealed a surprisingly rapid decline. This case is a classical incidence of drug-induced pemphigus foliaceus, in which D-penicillamine was constantly required not only to induce, but also to maintain the production of the autoantibodies. PMID- 15663503 TI - Treatment of radiation-relapsing primary cutaneous B-cell lymphoma with an anti CD20 monoclonal antibody. AB - Primary cutaneous B cell lymphomas have a high recurrence rate after treatment with surgery and/or local radiation therapy. Two men are described in whom radiotherapy-relapsing cutaneous B-cell lymphomas were successfully treated with the monoclonal anti-CD20 antibody rituximab. Both patients had a complete response with no recurrence at follow-up at 17 and 24 months for the large B-cell lymphoma of the leg and the follicle centre cell lymphoma, respectively. These are two of the few cases in the literature showing that rituximab is an effective and well-tolerated treatment for radiotherapy-relapsing primary cutaneous B cell lymphoma. PMID- 15663504 TI - Basal cell carcinoma: from host response and polymorphic variants to tumour suppressor genes. AB - The molecular factors and events that characterize susceptibility and outcome in cutaneous basal cell carcinoma (BCC) have been the focus of much research interest. As a result, we are beginning to understand the complex relationships between exposure to ultraviolet radiation (UVR), host response and the resulting damage to key genes that characterize these tumours. In this review, we will focus on genetic factors that influence susceptibility and outcome. While the search for susceptibility genes has generally resulted in the identification of low penetrance allelic variants, studies on modifier genes influencing outcome variables such as tumour number, age of onset and tumour subtype have identified factors with higher potential impact. Here we will briefly describe some recent work on the genetic basis of the immune response to UVR, the effect of UVR on the generation of reactive oxygen species and their detoxification, and the role of onco- and tumour suppressor genes. Areas for further research are highlighted, together with a consideration of possible applications in clinical practice. PMID- 15663505 TI - N-acetyltransferase 2 polymorphisms in patients with Behcet's disease. AB - It is possible that dietary, environmental factors and/or genetic polymorphisms in xenobiotic-metabolizing enzymes may contribute to the development of Behcet's disease. As N-acetyltransferase (NAT) 2 is an important xenobiotic-metabolizing enzyme and theoretically the nonacetylated xenobiotics may induce an autoimmune mechanism, the aim of the present study was to investigate whether the genetic polymorphism of NAT2 plays a role in susceptibility to Behcet's disease. Forty Behcet's disease patients and 82 control subjects were enrolled in the study. NAT2*5A, NAT2*6A, NAT27*A/B and NAT2*14A polymorphisms were detected by using real time PCR with LightCycler (Roche Diagnostics GmbH, Mannheim, Germany). The NAT2*5A and NAT2*6A mutant genotypes carried an increased risk of developing Behcet's disease [odds ratio (OR) = 66.29, 95% confidence interval (CI) = 8.21 535.33; and OR = 24; 95% CI = 2.04-304.98, respectively]. The NAT2*7A/B and NAT2*14A gene polymorphisms were not an increased risk for developing Behcet's disease. As a result of this study we conclude the NAT2 slow acetylator status may be a determinant in susceptibility to Behcet's disease. This finding may have implications for the theories of the pathogenesis of the disease as well as for therapeutic aspects. PMID- 15663506 TI - Serum interleukin 18 and tumour necrosis factor-alpha levels are increased in Behcet's disease. AB - Inflammation in Behcet's disease is thought to be mediated by cytokines derived from T-helper type 1 (Th1) lymphocytes. In this study, we tried to determine serum interleukin (IL)-18 and tumour necrosis factor (TNF)-alpha levels of patients with Behcet's disease. Twenty-seven patients with active Behcet's disease, and 20 healthy control subjects were included in this study. Differences between mean serum IL-18 and TNF-alpha level of patients with Behcet's disease were significantly increased when compared with the control group. A significant correlation was found between serum IL-18 and TNF-alpha levels of Behcet patients (rs = 0.627, P < 0.0001). IL-18 and TNF-alpha levels may be related to disease pathogenesis. Increased levels of IL-18 also support Th1 predominance in Behcet's disease. PMID- 15663507 TI - Epidermolytic hyperkeratosis type PS-1 caused by aberrant splicing of KRT1. AB - Mutations in the keratin 1 (KRT1) gene underlie epidermolytic hyperkeratosis (EHK). This autosomal dominant disorder is characterized by phenotypic heterogeneity. In the present study, we assessed a 33-year-old individual presenting with severe palmoplantar keratoderma and histopathological findings suggestive of EHK. We analysed genomic DNA extracted from the patient's blood lymphocytes for pathogenic mutations in KRT1. A heterozygous 4-bp deletion was identified in intron 1 of the gene (591+3_+6delGAGT), suggesting the possibility that it may interfere with the normal splicing of intron 1. We detected a 66-bp deletion in KRT1 mRNA extracted from the patient's skin, predicted to result in the translation of a mutant KRT1 lacking 22 amino acids, including the conserved helix initiation motif. The identification of this unusual and novel mutation underscores the diagnostic importance of sequence analysis of keratin gene noncoding regions. PMID- 15663508 TI - Evaluation of germline CDKN2A, ARF, CDK4, PTEN, and BRAF alterations in atypical mole syndrome. AB - Atypical mole syndrome is a sporadic or an inherited condition with an increased risk of melanoma. Germline mutations in the CDKN2A, ARF, CDK4 and somatic mutations in the PTEN and BRAF genes have been associated with melanoma. In this study, we evaluated genes associated with familial and sporadic melanoma for mutations in 28 probands with the atypical mole syndrome. No sequence alterations in the coding regions or in the splice junctions of CDKN2A, ARF, CDK4, PTEN or BRAF were identified. These data suggest that genes evaluated in this study are unlikely to be candidate genes for atypical mole syndrome and support the notion that unknown susceptibility gene/s for this disease exist. PMID- 15663509 TI - Complete paternal uniparental isodisomy of chromosome 1 resulting in Herlitz junctional epidermolysis bullosa. AB - Herlitz junctional epidermolysis bullosa (JEB) is an autosomal recessive mechanobullous disorder that results from loss-of-function mutations in the genes encoding the basement membrane component, laminin 5. Typically, there are frameshift, splice site or nonsense mutations on both alleles of either the LAMA3, LAMB3 or LAMC2 genes, with affected individuals inheriting one mutated allele from each parent. In this report, we describe a patient with Herlitz JEB in whom DNA analysis revealed homozygosity for the recurrent nonsense mutation R635X in LAMB3, located on chromosome 1q32.2. However, screening of parental DNA showed that although the patient's father was a heterozygous carrier of this mutation, the mother's DNA showed only wild-type sequence. Subsequent genotype analysis using 13 microsatellite markers spanning chromosome 1 revealed that the affected child was homozygous for the entire series of markers tested and that all of the alleles originated from the father. These results indicate that the Herlitz JEB phenotype in this patient is due to complete paternal isodisomy of chromosome 1 and reduction to homozygosity of the mutant LAMB3 gene locus. This is the fourth case of uniparental disomy to be described in Herlitz JEB, but it represents the first example of complete paternal isodisomy for chromosome 1 with a pathogenic mutation in the LAMB3 gene. These findings have important implications for mutation screening in JEB and for genetic counselling. PMID- 15663510 TI - Familial multiple cutaneous and uterine leiomyomas associated with papillary renal cell cancer. AB - Multiple cutaneous and uterine leiomyomas is an autosomal dominant condition that results in benign smooth muscle tumours of the skin and, in females, uterine fibroids. This syndrome overlaps with hereditary leiomyomatosis and renal cell cancer syndrome in which affected individuals may develop the rare type II papillary renal cell cancer, in addition to skin leiomyomas. Recently, heterozygous mutations in the gene encoding fumarate hydratase have been found to underlie both conditions. Fumarate hydratase is an enzyme that catalyses the conversion of fumarate to malate in the Kreb's cycle and may also function as a tumour suppressor gene. We report a family with multiple leiomyomas, uterine fibroids and papillary renal cell cancer. The proband is a 77-year-old Polish woman who developed multiple cutaneous leiomyomas on her right upper arm in her thirties and subsequently underwent a hysterectomy for uterine fibroids in her forties. She has four offspring: her eldest daughter also has skin and uterine leiomyomas with a similar onset; her son has multiple skin leiomyomas and in addition was diagnosed with metastatic papillary renal cell cancer at the age of 50 years; the two youngest daughters are unaffected. DNA sequencing in all the affected individuals disclosed a heterozygous G-->C substitution at nucleotide 173 of the fumarate hydratase gene, that converts an arginine residue (CGA) to proline (CCA). This missense mutation has not been reported previously and is designated R58P. Interestingly, the clinically asymptomatic 20-year-old son of the individual with renal cancer was also found to be heterozygous for R58P. It is likely that he will develop skin leiomyomas in the future but the risk of renal cancer is difficult to predict. Nevertheless, detection of this mutation has important implications for screening and genetic counselling in this and other family members. PMID- 15663511 TI - UVA1 and UVB irradiated skin investigated by optical coherence tomography in vivo: a preliminary study. AB - In histological studies, it has frequently been demonstrated that ultraviolet (UV) exposure, in particular UVB, can induce significant thickening of the viable epidermis and/or stratum corneum. Since skin biopsy alters the original skin morphology and always requires an iatrogenic trauma, we aimed to introduce optical coherence tomography (OCT) in vivo for the investigation of changes of epidermal thickness (ET) following UVA1 and UVB irradiation. Twelve healthy subjects received daily 60 J/cm2 of UVA1 and 1.5 minimal erythema doses UVB on their upper back over 3 consecutive days. Twenty-four hours after the last irradiation, OCT assessments were performed on UV exposed and adjacent nonirradiated control sites. Data of ET as expressed by comparison of the averaged A-scans differed significantly between nonirradiated (94.2 +/- 15.7 microm), UVA1 (105.4 +/- 12.8 microm) and UVB (125.7 +/- 22.1 microm) exposed sites. In comparison to the nonirradiated sites, UVA1 exposed skin showed significant (P = 0.022) increase of ET of 11% and UVB exposed sites a significant (P < 0.001) increase of 25%. ET of UVA1 and UVB exposed skin sites differed significantly (P =0.005). Our results obtained from OCT in vivo measurements confirm data of previous histological studies indicating that not only erythemogenic doses of UVB, but also suberythemogenic doses of UVA1 may have a significant impact on ET. OCT appears to be a promising bioengineering technique for photobiological studies. However, further studies are needed to establish its measurement precision and validity, and to investigate in vivo spectral dependence on UV induced skin changes such as skin thickening. PMID- 15663512 TI - Subcutaneous administration of collagen-polyvinylpyrrolidone down regulates IL 1beta, TNF-alpha, TGF-beta1, ELAM-1 and VCAM-1 expression in scleroderma skin lesions. AB - In this study the effect of collagen-polyvinylpyrrolidone (collagen-PVP) vs. triamcinolone acetonide (Triam) in scleroderma (SSc) skin lesions was evaluated. Ten SSc patients were treated weekly with subcutaneous injections of 0.2 mL Triam (8 mg/mL) or 0.2 mL collagen-PVP (1.66 mg collagen). Skin biopsies were obtained from lesions before and after treatment. Tissue sections were evaluated by histology and immunohistochemistry (ELAM-1, VCAM-1, IL-1beta, TNF-alpha, TGF beta1 and PDGF). The corticoid-treated group showed abnormal tissue architecture while the biodrug-treatment restored cutaneous appendages and type I/III collagen proportion. Cytokine and adhesion molecule expression was almost inhibited with Triam, while collagen-PVP down-regulated it. Collagen-PVP improved the tissue architecture of SSc lesions and down-regulated some proinflammatory parameters, without the side effects induced by corticoids. PMID- 15663513 TI - Extensive cutaneous fibrosis and ulceration caused by methadone injection. PMID- 15663514 TI - Gingival hyperplasia due to metal allergy. PMID- 15663515 TI - Cutaneous lymphadenoma. PMID- 15663516 TI - 'Slot machine' finger: an occupational dermatosis? PMID- 15663517 TI - Vulval eczema associated with propolis sensitization from topical therapies treated successfully with pimecrolimus cream. PMID- 15663518 TI - Dermatitis artefacta in a teenager after awareness of his HIV-positive diagnosis. PMID- 15663519 TI - Dysphagia in dermatomyositis secondary to bladder cancer: rapid response to combined immunoglobulin and methylprednisolone. PMID- 15663520 TI - Keloid development within adolescent striae distensae. PMID- 15663521 TI - Herpes simplex of the vulva evoked by topical tacrolimus treatment. PMID- 15663522 TI - Serum lipid fractions, nitric oxide and plasma endothelin-1 levels in actinic keratosis. PMID- 15663523 TI - Dermatological aspects of medicine: recent advances in nephrology. PMID- 15663524 TI - An unusual tender eruption. PMID- 15663525 TI - An erythematous nodule on the sole of the foot. PMID- 15663526 TI - A rapidly growing nodule on the face. PMID- 15663527 TI - Evaluating peer support for prostate cancer: the Prostate Cancer Peer Support Inventory. AB - OBJECTIVE: To develop and test a measure for assessing peer support for men attending prostate cancer support groups, and to describe socio-demographic, medical and adjustment characteristics of Australian men who attend these support groups. PATIENTS AND METHODS: In all, 1224 men (51% response) from 44 prostate cancer support groups across Australia were recruited by mail. Men completed self report measures that included the Prostate Cancer Peer Support Inventory (PCSI), the UCLA Prostate Cancer Index bother scales, psychological distress, quality of life (QoL), bother from pain and tiredness, perception of the clinician's support for group participation. Group-level variables were also included in the analyses. RESULTS: Peer support was rated positively by most men; a high satisfaction with support groups was related to better QoL, lower pain, younger age, higher perceived clinician support for group participation, use of alternative therapies, lower education, and regular attendance; dissatisfaction with support groups was related to higher psychological distress, lower QoL, and lower perceived clinician support for group participation. Group variables did not predict positive or negative support. Overall QoL was similar to community norms and psychological distress was low, with only 8% of men reporting high distress. The most common physical symptom was sexual bother, with 74% of men reporting moderate or high bother. CONCLUSIONS: The PCSI was a useful measure of peer support. Perception of the benefits of peer support was related to individual but not group differences. The clinicians' attitudes to participation in support groups influenced the men's experience of these groups, and this finding has implications for developing support services for these men. PMID- 15663528 TI - Baseline predictors of end-stage renal disease risk in patients with type 2 diabetes and nephropathy: new lessons from the RENAAL Study. PMID- 15663531 TI - Care resources for the aging population. PMID- 15663532 TI - An overview of regular dialysis treatment in Japan (as of 31 December 2002). AB - The Japanese Society for Dialysis Therapy (JSDT) has annually conducted a nationwide statistical survey of all dialysis facilities in Japan. The Society conducted this survey of 3625 dialysis facilities at the end of 2002, and responses were received from 3612 facilities (99.61%). Based on the survey investigation results tabulated at the end of 2002, the population of dialysis patients in Japan was 229,538. The gross mortality rate was 9.2% for the year extending from the end of 2001 to the end of 2002. The mean age of patients beginning dialysis was 64.7 years. The mean age of the overall dialysis population in the study year was 62.2 years. In the patients who began dialysis in 2002, the number of patients with diabetic nephropathy as the primary disease increased to 39.1% of patients. 6.5% of 40-64-year-old dialysis patients had taken out long-term-care insurance, but 31% of 65-year-old or older dialysis patients had taken out this insurance. 65.7% of the three-times-weekly facility hemodialysis patients underwent daytime dialysis, while 11.9% of patients underwent evening dialysis. PMID- 15663533 TI - Evaluation of improved coronary flow velocity reserve using transthoracic Doppler echocardiography after single LDL apheresis. AB - The purpose of this study was to clarify whether coronary flow velocity reserve (CFVR), evaluated by adenosine 5'-triphosphate-induced hyperemia, is improved by single low-density lipoprotein (LDL) apheresis. Lipid lowering therapy is known to improve endothelium-dependent vasodilatation in forearm or coronary resistant vessels. However, few reports have studied the effect of acute LDL reduction on CFVR. METHODS: Seven patients with familial hypercholesterolemia and significant coronary stenosis except in the left anterior descending artery (LAD) were enrolled in this study. Coronary flow velocity reserve was estimated before and after LDL apheresis using transthoracic Doppler echocardiography (TTDE), which detects the flow velocity at the distal site of the LAD. Although the averaged diastolic peak velocity (ADPV) during ATP-induced hyperemia was similar before and after LDL apheresis, the ADPV at baseline decreased from 30.69 to 25.56 cm/s, resulting in an increased CFVR from 1.78 to 2.10 (P < 0.001). Plasma bradykinin and 6 keto PGF1alpha increased while fibrinogen and plasma viscosity decreased after apheresis. Single LDL apheresis improves CFVR according to TTDE analysis because of the decreasing ADPV at baseline, which is thought to be induced by epicardial coronary artery dilatation and improved microvessel function. This is the result of various factors, such as changes in plasma LDL cholesterol, bradykinin and PGI2 levels with LDL apheresis. PMID- 15663534 TI - Continuous hemodiafiltration in pediatric critical care patients. AB - Continuous hemodiafiltration (CHDF) is an essential procedure in critical care. However, application of this therapy to pediatric patients is associated with several problems derived from their smaller body size and weight compared with adults. We have successfully conducted CHDF in pediatric patients including newborns by taking such problems into consideration and carefully coping with them. The present study consisted of 60 pediatric patients treated with CHDF. Clinical efficacy and safety of CHDF in pediatric patients were assessed in these patients by reviewing patient clinical records. The 60 patients treated with CHDF included 27 males and 33 females. Their body weight ranged from 700 g to 53.0 kg. The mean CHDF duration was 6.80 +/- 6.94 days. Blood access was provided in a veno-venous mode in 42 patients, and an arterio-venous mode in 18 patients. Of the 60 pediatric patients receiving CHDF, 31 patients survived without serious complications, achieving a survival rate of 51.7%. Successful CHDF in pediatric patients was achieved by careful and exact execution of the following countermeasures to overcome problems specific to application of this therapy to pediatric patients: minimization of the priming volume; use of colloid solutions or whole blood as priming solution; maintaining secure blood access; selection of an appropriate anticoagulant; temperature control of both the patient's body and components of the hemofiltration circuit. In pediatric critical care, CHDF is safely applicable to the critically ill and expected to produce a wide spectrum of clinical efficacy just as in adults. PMID- 15663535 TI - A last resort modality using cryofiltration apheresis for the treatment of cold hemagglutinin disease in a Veterans Administration hospital. AB - Cold hemagglutinin disease is a cold autoimmune hemolytic anemia (cAIHA) caused by an autoantibody, such as IgM, directed against the I-antigen present on the surface of erythrocytes. Cold exposure can activate this system causing hemolysis, hemagglutination, microvascular thrombosis, or acrocyanosis. Thus, surgical procedures requiring hypothermia, such as coronary artery bypass surgery, present a significant problem in patients with cAIHA. The purpose of this study was to evaluate the safety and effectiveness of cryofiltration apheresis (CFA), used as a last resort, for the treatment of cAIHA. Effectiveness was evaluated by clinical assessment and laboratory evaluations of cold agglutinin titer, immunoglobulins, and other plasma proteins. Safety was evaluated by vital signs, monitoring, and laboratory measurements of complements, hematology and blood chemistry. Five patients with cAIHA were treated by CFA using the cryoglobulin (CG) filter (Pall Medical, Ann Arbor, MI, USA). Four patients received only one CFA procedure, while one patient received four CFA treatments. The cold agglutinin titers were fairly low, ranging from 1 : 1 to 1 : 2048. However, a wide thermal amplitude(4-37 degrees C) was observed in most patients. Two out of five patients responded favorably with reduction in titer. The two responders had acute forms of cAIHA with serum positive for cryoglobulins. The three non-responders had chronic forms of cAIHA with negative cryoglobulins. CFA effectively removed cryoprotein precipitates while conserving other plasma components. The CG filter was biocompatible with no complement activation or observed complications due to CFA or CG filter. While the mechanism of action in treating this type of patient population with CFA is unknown, the plausible theories are discussed. PMID- 15663536 TI - Double filtration plasmapheresis for the treatment of a rheumatoid arthritis patient with extremely high level of c-reactive protein. AB - A 68-year-old-male who was diagnosed as having rheumatoid arthritis (RA) 7 years previously was admitted the Chiba Social Insurance Hospital due to general fatigue, spiking fever, and appetite loss. Blood tests showed extremely high levels of C-reactive protein (CRP, 318.5 mg/dL), and hypergammapathy (IgG 3228 mg/dL, IgA 905 mg/dL, IgM 2537 mg/dL) and high titers of rheumatoid factor (RAPA 40960X). He was diagnosed as having RA with vasculitis, according to interstitial pneumonitis, cutaneous nodules and polyneuropathy. Prednisolone (30 mg/day) was prescribed, however, myeloperoxidase-antineutrophil cytoplasmic antibody proved to be positive (86EU) and cyclophosphamide (50 mg/day) was added one week later. Additionally, IgM K-chain M-protein was revealed and the differentiation between auto-immune and hematologic diseases was required for further drug prescriptions. Therefore, double filtration plasmapheresis (DFPP) was initiated weekly. Hematologic diseases were negated and the hypergammapathy was improved. C reactive protein and MPO-ANCA decreased to the normal level after three sessions (IgG 1064 mg/dL, IgA 331 mg/dL, IgM 94 mg/dL, CRP 0.04 mg/dL) and the patients symptoms improved. Prednisolone was tapered and he was discharged. It was suggested that the case presented here was quite rare, having an extremely high level of CRP which was successfully managed by DFPP. PMID- 15663537 TI - Plasmapheresis treatment in Guillain-Barre syndrome: potential benefit over IVIg in patients with axonal involvement. AB - Response to therapeutic plasma exchange (TPE) was evaluated in patients diagnosed with Guillain-Barre Syndrome (GBS). Our aim was to assess response to TPE in patients who had failed treatment with intravenous immune globulin (IVIg). We conducted a retrospective chart review of 10 patients with the diagnosis of Guillain-Barre Syndrome who required TPE. Patients were identified by reviewing data from log books for TPE at The University of Connecticut Health Center, Farmington CT, USA. Patients who had failed IVIg treatment prior to being referred for TPE were also identified. Eight out of 10 patients treated with TPE showed improvement in their neurological exam. Four patients had axonal involvement on electromyelogram (EMG). Three patients were referred for TPE after failing IVIg treatment. All three of these had axonal involvement on EMG. Three of the four patients with axonal involvement demonstrated improvement with TPE. TPE may be a superior treatment option as compared to IVIg in patients with GBS and EMG findings of axonal involvement. PMID- 15663538 TI - Peripheral digit ischemic syndrome can be a manifestation of postoperative thrombotic thrombocytopenic purpura. AB - In addition to common dysfunction of the brain and kidney, thrombotic thrombocytopenic purpura (TTP) may present with atypical clinical features due to the involvement of other organs such as the lung, pancreas, heart, eye, and skin. We have also observed the unusual presentation of peripheral digit ischemic syndrome (PDIS) in some patients with postoperative TTP. To clarify this relationship between TTP and PDIS, the hematologic data from the medical records of patients with known diagnoses of thrombotic microangiopathy (TM) were examined in a single institution. A total of 94 patients were diagnosed with TM. Among these patients, PDIS developed in six patients and in all these patients PDIS occurred with postoperative TTP. Four patients also had acute respiratory distress syndrome (ARDS). Because of delayed diagnosis of TTP, only two patients survived and four died. One patient responded to plasma exchange and survived, and another patient recovered from postoperative TTP without plasma exchange. However, both patients required the amputation of multiple digits. In conclusion, PDIS is another atypical manifestation of TTP and has occurred exclusively in patients with postoperative TTP in this series. Once PDIS developed, the prognosis was poor and amputation of digits was needed in surviving patients. Early recognition of this atypical manifestation of TTP is essential for a favorable outcome. PMID- 15663539 TI - Effect of skin care with an emollient containing a high water content on mild uremic pruritus. AB - Skin care is very important for preventing uremic pruritus. However, mild uremic pruritus has usually been treated with antihistamine and urea-containing ointments. We therefore examined the effects of an aqueous gel with higher water content. Twenty hemodialysis patients with mild pruritus who were not being treated with any emollient were divided into two groups of 10 each. Patients in one group were treated with an aqueous gel containing 80% water. This emollient was applied twice daily for 2 weeks. No emollient was applied for the next 2 weeks. The other group of patients were not treated with any emollient for the 4 weeks. Visual analog scale scores for itching in the experimental group at week 2 were significantly decreased compared with that at week 0 (3.5 +/- 0.3 vs 0.6 +/- 0.2, P < 0.01). Skin dryness in the experimental group was significantly improved at week 2 compared with that at week 0. The visual analog scale score for itching increased to 1.2 +/- 0.5 and skin dryness reappeared in 40% of patients by week 4, i.e. after the emollient was stopped. There were no significant changes in the control group during the study. It is concluded that the aqueous gel with high water content reduced itching and improved xerosis in patients with mild uremic pruritus. It is reasonable that skin care with an emollient containing a high water content is first started for hemodialysis patients with xerosis, even if they do not feel itching. PMID- 15663544 TI - 2004 Japanese Society for Dialysis Therapy guidelines for renal anemia in chronic hemodialysis patients. AB - The guideline committee of Japanese Society for Dialysis Therapy (JSDT), chaired by Professor F. Gejyo of Niigata University, now publishes an original Japanese guideline entitled 'Guidelines for Renal Anemia in Chronic Hemodialysis Patients'. It includes the re-evaluation of the usage of recombinant human erythropoietin (rHuEPO) with the medical and economical arguments regarding the prognosis and the quality of life of Japanese hemodialysis patients. This guideline consists of 7 sections. The first section comprises the general definition and the differential diagnosis of anemia. The hemoglobin (Hb) level of the Japanese population seemed to be low when compared with that of the European and American populations. The second section describes the target Hb level in hemodialysis patients. Multivariate analysis of the data that were collected from dialysis institutions throughout the country showed that an Hb level of 10-11 g/dL (Ht level 30-33%) at the first dialysis session in a week is the ideal range for chronic hemodialysis patients in terms of the 3-5 year survival rate. The supine position at blood sampling and the sampling timing at the first dialysis session in a week might affect the lower setting of target Hb hematocrit (Ht), compared to that of European and American guidelines. However, we particularly recommended that an Hb level of 11-12 g/dL (Ht level from 33 to 36%) at the first dialysis session in a week is desirable in relatively young patients. In the third section, the markers of iron deficiency are discussed. The Transferin saturation test (TSAT) and serum ferritin were emphasized as the standard markers. The routes of administration of rHuEPO and its dosages are written in the fourth section. The subcutaneous route was associated with the occurrence of secondary red cell aplasia due to anti-rHuEPO antibodies; however, secondary red cell aplasia was seldom observed in the venous injection. From this fact we recommend venous injection for chronic hemodialysis patients. We advocate an initial dosage of 1500 U three times per week. The fifth section deals with the factors refractory to treatment with rHuEPO. If the patient shows an inadequate response to the usage of 9000 U per week, this condition defines the inadequate response to rHuEPO in Japan. Blood transfusion must be avoided where possible. The reasons for this and the adverse effects are interpreted in section six. In the final section, the adverse effects of rHuEPO are listed. Among them, hypertension, thrombotic events and secondary red cell aplasia were emphasized as the major complications. PMID- 15663545 TI - A comparison of bicarbonate hemodialysis, hemodiafiltration, and acetate-free biofiltration on cytokine production. AB - Acetate-free biofiltration (AFB) is a special hemodiafiltration (HDF) modality performed with a base-free dialysate and simultaneous injection of non-pyrogenic sodium bicarbonate solution. The purpose of this study was to investigate the difference of cytokine production by conventional bicarbonate hemodialysis (BCD), standard HDF and AFB in the same patients. Eight stable hemodialysis patients were treated in random order with BCD, HDF and AFB every 4 weeks. The production of interleukin-1 beta (IL-1 beta) and interleukin-1 receptor antagonist (IL-1Ra) by peripheral blood mononuclear cells (PBMC) was investigated without stimulation and with stimulation by a small amount of endotoxin (ET)-contaminated beta 2 microglobulin (beta 2M) and lipopolysaccharide (LPS) before and after dialysis treatment in the last sessions during all periods. To serve as controls, 14 healthy volunteers participated in this study. In spontaneous IL-1Ra production, the values of before and after AFB were not significantly different from that of the controls, and the values of before and after BCD and before HDF were significantly higher than that of the controls. In LPS-stimulated PBMC, IL-1 beta production before and after AFB was not significantly different from that of the controls, and before and after BCD and HDF was significantly higher than that of the controls. In ET-contaminated beta 2M-stimulated PBMC, IL-1 beta production before and after AFB was not significantly different compared to the controls, and the production was significantly lower than that before and after BCD and HDF. In addition, IL-1Ra production after AFB was not significantly different from the controls, and the production was significantly lower than that before and after BCD and HDF. It was concluded that a lower cytokine production by AFB may have the effect of preventing dialysis-related complications. PMID- 15663546 TI - Renal cell carcinoma detected by screening shows better patient survival than that detected following symptoms in dialysis patients. AB - There is an apparent advantage to early or incidental diagnosis of renal cell carcinoma in the general population. We investigated whether the prognosis of dialysis patients with renal cell carcinoma is better if renal cell carcinoma is detected by screening of patients without symptoms rather than by examination after the appearance of the symptoms. Two groups in which renal cell carcinomas were detected by screening (N = 721) and by symptoms (n = 76) were compared. The model tested consisted of patient age and duration of dialysis in addition to the detection by screening or symptoms. Cox proportional hazard model was used to compare survival rates between the groups. The hazard ratio of death from all causes in patients with renal cell carcinoma detection by screening was 0.653 and the risk of death was reduced by 35%, compared with that in the group with detection due to symptoms. Moreover, the 50% survival rate in the group detected by symptoms was 80 months and the ratio of those detected by screening was 119 months. Therefore, screening provided a survival benefit of 39 months (3.3 years) in death from all causes after adjustment for age and duration of dialysis. The survival rate was best in young patients with a short duration of dialysis and renal cell carcinoma detected by screening. In dialysis patients with renal cell carcinoma, the survival rate in the group detected by screening was better than that in the group detected by symptoms. Especially, there was a large effect of screening for renal cell carcinoma in the young dialysis patients. PMID- 15663547 TI - Bio-intact parathyroid hormone and intact parathyroid hormone in hemodialysis patients with secondary hyperparathyroidism receiving intravenous calcitriol therapy. AB - Intact parathyroid hormone (iPTH) assay has been the most widely used for the diagnosis of secondary hyperparathyroidism and evaluation of vitamin D therapy. However, 1-84 PTH assay might be a better diagnostic tool since iPTH detects not only 1-84 PTH but also large C-terminal fragments, which would antagonize PTH action. Therefore, we conducted a multicenter study to evaluate the clinical usefulness of a newly developed immunochemiluminometric assay for 1-84 PTH, Bio Intact PTH (BiPTH). Thirty-five uremic patients with secondary hyperparathyroidism participated in the study. Intravenous calcitriol therapy was continued for 12 months. iPTH and bone-specific alkaline phosphatase (BAP) were monitored at each dialysis center to control the dose of calcitriol. Serum and plasma samples were collected from each center and both iPTH and BiPTH were measured using Allegro-Lite assay reagents from Nichols Institute Diagnostics (San Clemente, CA, USA). Intravenous calcitriol suppressed iPTH after 1 month as well as BiPTH. Bone-specific alkaline phosphatase decreased after 3 months. A high degree of correlation between Nichols iPTH and BiPTH (y = 0.3913 x + 19.517, r = 0.9561) was demonstrated with a BiPTH/iPTH ratio of approximately 0.44. Significant correlation between BAP and iPTH, or between BAP and BiPTH was not observed. Our limited data failed to demonstrate the superiority of BiPTH to iPTH. Therefore, further investigations would be necessary to examine the relationship between BiPTH and bone histomorphometry. PMID- 15663548 TI - Dose-response study of 22-oxacalcitriol in patients with secondary hyperparathyroidism. AB - The dose-response relationships and the safety of administering 22-oxacalcitriol (OCT) to patients with secondary hyperparathyroidism (2HPT) under regular three times-weekly hemodialysis (HD) were evaluated by double-blind parallel group design. A total of 203 patients with 2HPT were randomly allocated into four groups, and 5 microg (Group L), 10 microg (Group M), or 15 microg (Group H) OCT, or placebo (Group P) was administrated at the end of every HD for 12 weeks. Reductions of intact-parathyroid hormone (iPTH) concentration greater than 30% from baseline were observed in 7.7% of Group P as compared to 77.3% of the pooled OCT groups after 12 weeks of treatment (Mantel test: P < 0.001). Time-trends (slopes) of log-iPTH concentration calculated by least-squares line fitting to each patient's data during treatment differed between Group P and the pooled OCT groups (t-test: P < 0.001) and these iPTH slopes decreased dose-dependently (linear trend by t-test: P < 0.001). Slopes of serum calcium corrected for albumin (corrected-sCa) concentrations also differed between Group P and the pooled OCT groups (t-test: P < 0.001), and increased dose-dependently (linear trend by t-test: P < 0.0001). Serum phosphorus and Ca x P product increased significantly only in high dose groups. Slopes of log(iPTH) and corrected-sCa concentrations were reciprocally related. Most adverse events were hypercalcemia and dose-related, but occasionally comprised pruritus or increased serum creatinine phosphokinase. These results indicate that OCT produced a strong and dose-dependent suppression of PTH and an increase of corrected-sCa concentration in patients with 2HPT. The recommended initial dosages of OCT would appear to be 5 microg when pretreatment iPTH concentrations are less than 500 pg/mL, and 10 microg when greater than 500 pg/mL for safe and effective treatment. As in the case of PTH, calcium and phosphorus showed dose-dependent increases. It is therefore essential to take precautions as to possible increases in calcium and phosphorus. PMID- 15663549 TI - A preliminary report on multiple family discussion groups for patients with chronic medical illness and its repercussions in the management of the hemodialysis process. AB - The hemodialysis (HD) process involves an important degree of stress, not only for the patient but also for the family. The available data suggest that the quality of the family's performance predicts the degree of commitment that the patient acquires with the dialysis center. The establishment of a program of multiple family discussion groups allows exploration of the effect of the treatment on the patient in their context, not only regarding the illness, but also regarding the quality of life related to health, satisfaction and functional state. After the startup in the Hospital Virxe da Xunqueira of the multiple family discussion group (MFDG) for patients with chronic medical illness, the objective of this work was to evaluate the repercussions of the MFDG over the therapeutic fulfillment, the quality of life and the expectations of the patients in the chronic HD program, through the assessment of these indicators before and after their participation in this group. The MFDG was performed for a total of eight people, the families of four patients in the chronic HD program. Six weekly 1.5 hour sessions were performed and structured according to the following general contents: chronic illness impact component (2 sessions), family development component (3 sessions) and family illness integration component (1 session). Although no objectives were made for changes in relation to the therapeutic fulfillment, the average auto-effectiveness, locus of control, success and family general expectations went up slightly after the participation in the MFDG. The average specific self-effectiveness and family expectations in the presence of the illness reflected a modest increase, while the specific expectations of control locus and success in the presence of the illness decreased slightly. The scores obtained regarding the general state of health reflected a small decrease, while the evaluation of the quality of life of patients and family members showed a slight increase. We can conclude in the first place highlighting the viability of the MFDG, since no impediments were found either in recruitment of the families of the participants, or in preventing their excellent participation in the beginning and through the course of the group. Although no objectives were made for changes in relation to the therapeutic fulfillment, the high indices of satisfaction which the group obtained indicate that the discussion group is useful for the patients to find more support from their families, to change their view of the illness, to learn from other families new ways to resolve the difficulties and to increase their perception of capacity in the presence of the illness. The obtained data are preliminary and derived from only four families, but are encouraging as far as the improvement in the quality of life and the adjustment of the participants to the illness. Studies with the inclusion of more families are still pending in order to be able to arrive at conclusions based on a greater empiric basis. The records of MFDG for the families of HD patients are scarce. With this work it is attempted to reveal that these types of groups can be applied with these patients and their families, and they seem to prove beneficial for all those involved: patients, family and health professionals. PMID- 15663550 TI - Therapeutic plateletpheresis in a case of symptomatic thrombocytosis in chronic myeloid leukemia. AB - Extreme thrombocytosis is a frequent feature in myeloproliferative disorders which can predispose a person to thrombotic complications. As opposed to other myeloproliferative disorders, symptomatic thrombocytosis is rare in chronic myeloid leukemia. We describe a second case report of chronic myeloid leukemia (Ph chromosome positive) in a patient in chronic phase on hydroxyurea who presented with sudden onset digital cyanosis of the left hand, giddiness, headache and malaise due to extreme thrombocytosis. A 67% global reduction in the platelet count from 1553 x 10(9)/L to 513 x 10(9)/L after two therapeutic plateletpheresis procedures was seen. There was simultaneous improvement in all symptoms except cyanosis on the tip of the middle finger that progressed to dry gangrene. Dramatic reduction in the platelet count and ablation of symptoms by therapeutic plateletpheresis is an effective therapy and should begin as soon as possible. PMID- 15663551 TI - A case report of plasma exchange therapy in non-paraneoplastic cerebellar ataxia associated with anti-Yo antibody. AB - A 71-year-old-woman was admitted to the S. Eugenio Hospital for a history of progressively impaired standing and gait. Anamnesis revealed systemic hypertension, gastric polyposis and juvenile pulmonary tuberculosis. Neurological examination showed a severe truncal and gait ataxia, without any sensory-motor impairment. Motor and somato-sensory evoked potentials were normal. Brain Magnetic Resonance Imaging (MRI) showed minimal signs of chronic ischemia only at a supratentorial level. Cerebral Single Photon Emission Computed Tomography, spinal MRI, total body computed tomography, Esophagogastroduodenoscopy, and finally total body Positron Emission Tomography resulted negative for neoplasms. Oncological serum markers were negative. Serum antibody against Purkinje's cells (Anti-Yo) was detected and titer was 1:80, while normally it should be undetectable. Other autoantibodies (Anti-Hu, Anti-Ri) were undetectable. Two sessions of plasma exchange (PE) were thus performed, leading to a rapid, marked and durable improvement of standing and gait and to a reduction of the autoantibody, which became undetectable. No serious adverse effect was noted. Although no definite therapy for autoimmune cerebellar ataxia has been established, PE should be considered as one of the main therapeutic choices. PMID- 15663552 TI - A case report of the effect of plasma exchange on reactive hemophagocytic syndrome associated with toxic shock syndrome. AB - We report here the case of a patient suffering from hemophagocytic syndrome (HPS) associated with toxic shock syndrome (TSS). A 50-year-old man was admitted because of fever, watery diarrhea and shortness of breath. Clinical analysis revealed systemic cyanosis, sunburn-like erythema and septic shock. Staphylococcus aureus was identified from both blood and sputum culture and the serum enterotoxin A antibody test was positive, suggesting that this was a case of TSS. Though the respiratory and hemodynamic status improved by the mechanical ventilation, fluid resuscitation with catecholamine and antibiotic therapy, the platelet count decreased rapidly. Bone marrow aspiration revealed a large quantity of hemophagocytosis by macrophages. This reactive HPS was treated not with immunosuppressive drugs but with therapeutic plasma exchange in order to prevent worsening of S. aureus infection. After plasma exchange, the circulating macrophage colony-stimulating factor (M-CSF) level was reduced and the platelet count increased rapidly. Bacteria associated HPS remains a difficult diagnosis with high mortality and there is a crucial question of whether this should be treated with immunosuppressive drugs. The patient's clinical course would suggest that the therapeutic plasma exchange should be considered as a therapeutic tool for the bacteria associated HPS instead of immunosuppressive drugs. PMID- 15663553 TI - Surfactant protein D and asthma. PMID- 15663554 TI - Surfactant protein D deficiency influences allergic immune responses. AB - BACKGROUND: The collectin surfactant protein D (SP-D) confers protection against pulmonary infection and inflammation. Recent data suggest a role for SP-D in the modulation of allergic inflammation. OBJECTIVE: The aim of this study is to characterize the immune responses of SP-D-deficient (SP-D(-/-)) mice in a kinetic model of allergic inflammation. We determined whether allergic parameters were enhanced in SP-D(-/-) mice in vivo. Further, we examined whether functional immune responses in vitro such as lymphocyte proliferation (LP) and cytokine production were modulated in the absence of SP-D. METHODS: In vivo, wild-type (WT) and SP-D(-/-) mice were sensitized and challenged with the allergen ovalbumin (OVA) and assessed for allergic parameters (bronchoalveolar lavage (BAL) eosinophils, IL-13 production, pulmonary IFN-gamma, IL-10 expression) at early time points (1 and 3 days of challenge) in comparison with late time points (7 days of challenge). In vitro, spleen cells from WT and SP-D(-/-) mice were stimulated with the mitogen concanavalin A (ConA) and lipid A (LpA) and analysed for LP, IL-13 and IFN-gamma production. Toll-like receptor 4 (TLR4), ligand for LpA, was assessed by mRNA expression and immunohistochemistry in vivo. RESULTS: Following allergen exposure in vivo, SP-D(-/-) mice expressed higher BAL eosinophils and IL-13 concentrations and lower IFN-gamma expression at early time points compared with WT mice. IL-10 expression was increased at early time points in SP-D(-/-) compared with WT mice. Allergen-induced TLR4 expression was increased in WT, but not in SP-D(-/-) mice. After stimulation with LpA and ConA in vitro LP was increased and IFN-gamma concentration was decreased in SP-D(-/-) mice. CONCLUSION: SP-D may be critical for the modulation of early stages of allergic inflammation in vivo. PMID- 15663555 TI - Serum surfactant protein D is elevated in allergic patients. AB - BACKGROUND: There is evidence that surfactant protein (SP)-D is important in the innate, as well as in the adaptive pulmonary immune response. Serum concentrations of SP-D have been proposed as parameter of the integrity of the blood-airspace barrier in interstitial lung diseases. We hypothesized that serum SP-D concentrations are affected in allergic patients and correlate with changes in allergic airway inflammation. OBJECTIVE: To determine levels of serum SP-D in allergic patients compared with non-allergic controls. Furthermore, to investigate associations between serum SP-D concentrations on the one hand and changes in commonly used markers of bronchial inflammation in allergic airways disease on the other hand. MATERIALS AND METHODS: Fifty allergic patients were studied and bronchial allergen challenge was used as a model to increase bronchial allergic inflammation in these patients. Serum SP-D concentrations, inflammatory parameters in induced sputum and bronchial hyper-responsiveness (BHR) were determined before and after allergen challenge. Twenty-five non allergic volunteers served as controls. RESULTS: Baseline serum SP-D was significantly higher in allergic patients as compared with controls (mean serum SP-D concentration (95% confidence interval): 62.7 (55.5, 70.0) in allergic patients vs. 49.5 (36.7, 62.3) ng/mL in non-allergic controls, P=0.006). In addition, baseline serum SP-D appeared to be an independent predictor for the magnitude of the late asthmatic response after allergen challenge. Furthermore, serum SP-D was predictive for the sputum eosinophil cationic protein concentration after allergen challenge. CONCLUSION: We propose that serum SP-D concentrations are associated with allergic bronchial inflammation and may give additional information, beside BHR and sputum eosinophils, about the degree of bronchial inflammation in allergic patients. PMID- 15663556 TI - Differential capacity of CD8+ alpha or CD8- alpha dendritic cell subsets to prime for eosinophilic airway inflammation in the T-helper type 2-prone milieu of the lung. AB - BACKGROUND: Different subsets of dendritic cells (DCs), identified in mouse spleen by their differential expression of CD8 alpha, can induce different T helper (Th) responses after systemic administration. CD8 alpha(-) DCs have been shown to preferentially induce Th type 2 (Th2) responses whereas CD8 alpha(+) DCs induce Th1 responses. OBJECTIVE: To study if these DC subsets can still induce different Th responses in the Th2-prone milieu of the lung and differentially prime for eosinophilic airway inflammation, typical of asthma. METHODS: Donor mice first received daily Flt3L injections to expand DC numbers. Purified CD8 alpha(+) or CD8 alpha(-) splenic DCs were pulsed with ovalbumin (OVA) or phosphate-buffered saline and injected intratracheally into recipient mice in which carboxyfluorescein diacetate succinimidyl ester-labelled OVA-specific T cell receptor transgenic T cells had been injected intravenously 2 days earlier. T cell proliferation and cytokine production of Ag-specific T cells were evaluated in the mediastinal lymph nodes (MLNs) 4 days later. The capacity of both subsets of DCs, to prime for eosinophilic airway inflammation was determined by challenging the mice with OVA aerosol 10 days later. RESULTS: CD8 alpha(-) DCs migrated to the MLN and induced a vigorous proliferative T cell response accompanied by high-level production of IL-4, IL-5, IL-10 and also IFN-gamma during the primary response and during challenge with aerosol, leading to eosinophilic airway inflammation. In the absence of migration to the MLN, CD8 alpha(+) DCs still induced a proliferative response with identical levels of IFN gamma but reduced Th2 cytokines compared with CD8 alpha(-) DCs, which led to weak eosinophilic airway inflammation upon OVA aerosol challenge. Unpulsed DCs did not induce proliferation or cytokine production in Ag-specific T cells. CONCLUSION: CD8 alpha(-) DCs are superior compared with CD8 alpha(+) DCs in inducing Th2 responses and eosinophilic airway inflammation in the Th2-prone environment of the lung. PMID- 15663557 TI - Single nucleotide polymorphism screening and association analysis--exclusion of integrin beta 7 and vitamin D receptor (chromosome 12q) as candidate genes for asthma. AB - BACKGROUND: The human genes coding for integrin beta 7 (ITGB7) and vitamin D receptor (VDR) are two of the several candidate genes for asthma and related phenotypes found in a promising candidate region on chromosome 12q that has been identified in multiple genomewide screens and candidate gene approaches. METHODS: All exons, including parts of the neighbouring introns, and the predicted promoter region of the ITGB7 gene were screened for common polymorphisms in 32 independent asthmatic and healthy probands, resulting in the detection of two single nucleotide polymorphisms (SNPs) unknown so far. In addition to these SNPs, five already described SNPs of the ITGB7 and one in the human VDR gene were analysed in a Caucasian sib pair study of 176 families with at least two affected children, using matrix assisted laser desorption/ionization time of flight mass spectrometry. All confirmed SNPs were tested for linkage/association with asthma and related traits (total serum IgE level, eosinophil cell count and slope of the dose-response curve after bronchial challenge). RESULTS: Two new variations in the ITGB7 gene were identified. The coding SNP in exon 4 causes a substitution of the amino acid GLU by VAL, whereas the other variation is non-coding (intron 3). None of the eight analysed SNPs, of either the ITGB7 or the VDR genes, showed significant linkage/association with asthma or related phenotypes in the family study. CONCLUSIONS: These findings indicate that neither the human ITGB7 nor the VDR gene seem to be associated with the pathogenesis of asthma or the expression of related allergic phenotypes such as eosinophilia and changes in total IgE level. PMID- 15663558 TI - Mothers of very low birth weight infants have less atopy than mothers of full term infants. AB - BACKGROUND: Studies on the pregnancy outcome of asthmatic mothers have suggested an increased rate of preterm deliveries. In contrast, our earlier study suggests that mothers of very low birth weight (VLBW) (<1500 g) infants less frequently had atopy than did mothers of full-term infants. METHODS: We inquired about symptoms of atopy and doctor-diagnosed atopy in parents of 370 infants of VLBW (<1500 g) and 544 parents of full-term infants. Odds ratios for atopic symptoms and diagnosed atopy were calculated, and groups were compared with a trend test. RESULTS: Mothers of preterm infants of birth weight (BW) <1000 g significantly less often had physician-diagnosed allergic rhinitis (AR) (P=0.02). Among all the mothers, a trend test showed that maternal AR was significantly (P=0.03) higher in parallel with a higher infant BW. Fathers of infants with different BWs showed no differences in prevalence of atopic symptoms. CONCLUSION: We thus infer that maternal balance between T-helper type 1 (Th1) and Th2 cells, shifted towards Th2 in those with AR, may have a favourable effect on maintenance of pregnancy before gestational week 30. PMID- 15663559 TI - Serum ovalbumin-specific immunoglobulin G responses during pregnancy reflect maternal intake of dietary egg and relate to the development of allergy in early infancy. AB - BACKGROUND: The value of allergen elimination diets during pregnancy for primary prevention of infant allergy has been questioned. However, dietary compliance may influence effectiveness. OBJECTIVES: To monitor egg intake during a randomized controlled trial of egg avoidance throughout pregnancy and lactation by serial measurements of serum ovalbumin (OVA) IgG concentration in conjunction with dietary diary record and also, to analyse specific IgG concentrations at birth in relation to infant allergic outcome. METHODS: Pregnant women, with personal or partner atopy, were randomized to complete dietary egg exclusion or an unmodified healthy diet before 20 weeks gestation. The infants were evaluated for atopy at 6 months of age. Serum food-specific IgG concentrations were determined by ELISA in maternal samples collected at study recruitment and during labour, and in infant samples at birth (umbilical cord). RESULTS: Serum-specific IgG to OVA, but not the unrelated allergen, cow's milk beta-lactoglobulin, decreased over pregnancy in egg-avoiding women only (P<0.001). Cord OVA IgG concentration correlated with maternal IgG at delivery (r=0.944; P<0.001), and for infants born to atopic women, cord concentration was higher than that of their mother's (P<0.001). Infants with the lowest and highest cord IgG concentrations were the least likely, and those with mid-range concentrations were the most likely, to be atopic by 6 months of age (P=0.008). CONCLUSION: Serum OVA IgG concentration reflects egg consumption, thereby indicating dietary allergen doses to which the developing immune system might be exposed. Trans-placental maternal IgG must be considered among early life factors that regulate infant atopic programming. PMID- 15663560 TI - The effect of specific immunotherapy on the expression of costimulatory molecules in late phase reaction of the skin in allergic patients. AB - BACKGROUND: Specific immunotherapy (SIT) modulates immune responses to allergens resulting in improvement of allergic symptoms. However, the mechanisms behind the clinical changes are not clear. Participation of costimulatory molecules on antigen-presenting cells and T cells in the process of antigen recognition is suggested to be of essential importance. The SIT effect on expression of costimulatory molecules has not been earlier examined. METHODS: Forty-one birch allergic patients were treated with SIT or placebo. After 1 year of treatment skin biopsies were obtained 24 h following allergen challenge. Sections were stained with antibodies against: EG2 (eosinophils), CD4 (T cells), CD68 (macrophages), CD1a (Langerhans cells), CD28 (on T cells) and costimulatory molecules (CD80, CD86). RESULTS: Following allergen challenge number of the CD4(+) and CD68(+) cells increased significantly (P=0.002, 0.0001, respectively) in the placebo, but not in the SIT-treated patients. The difference between groups was significant (P=0.003, 0.01, respectively). The numbers of EG2(+) cells increased significantly in both groups. CD80(+) cell numbers increased in the placebo (P=0.01) but not in the SIT group. The number of CD86(+) cells increased in both groups (placebo, P=0.001; SIT, P=0.01) but significantly less in the SIT group (P=0.05). The numbers of CD28(+) cells increased in the placebo (P=0.001) but remained unchanged in the SIT group. The difference between the groups was significant (P=0.05). CONCLUSION: There were lower numbers of cells expressing costimulatory molecules in SIT-treated than in placebo-treated patients. Decreased costimulation may lead to diminished immune response following allergen exposure. This could be an important factor contributing to the clinical improvement after SIT. PMID- 15663561 TI - Identification of anti-inflammatory drugs according to their capacity to suppress type-1 and type-2 T cell profiles. AB - BACKGROUND: Down-regulation or modulation of T cell activity by immunosuppressive drugs is an effective treatment in diseases where exaggerated T cell responses play a role. A primary effect of the anti-inflammatory drugs (AIDs) is inhibition of the synthesis of growth factors, such as IL-2, thereby down-regulating T cell proliferation. However, it is still largely unknown to what extent these AIDs are able to down-regulate specifically type-1 or type-2 T cell cytokine production, and whether they can down-modulate chemokine receptor expression, thereby preventing migration of T cells to the site of inflammation. OBJECTIVE: We investigated the suppressive effect of dermatologically used AID (cyclosporin A (CsA), lactoferrin (LF), 1 alpha, 25-dihydroxyvitamin D(3) (VD(3)), hydrocortisone (HC), di-methyl-fumarate (DMF), diclofenac (DF)) on both type-1 and type-2 T cells. Since allergic contact dermatitis is a skin disorder in which an exaggerated T cell response of both types of T cell subsets can be observed, we used this disorder as a model to study the capacity of AID to suppress type-1 or type-2 T cell responses. METHODS: Peripheral blood mononuclear cells of nickel allergic patients were cultured in the presence of allergen and increasing concentrations of AID. Proliferation was determined by measuring (3)H thymidine incorporation; chemokine receptor (CCR10, CCR4, CXCR3) expression was studied by flow cytometric analysis and IFN-gamma or IL-5 cytokine production was measured by ELISA. RESULTS: Three major patterns can be distinguished regarding the effect of AID on T cell responses. The first group, including CsA and LF, inhibited non selectively T cell proliferation, chemokine receptor expression and cytokine production, with CsA as the most potent drug tested. A second group of AID, which included VD(3), HC and DMF, suppressed mainly type-1 T cell responses, as revealed by strong interference with IFN-gamma production and CXCR3 expression, and limited effects on either or both IL-5 and CCR4 expression. The third pattern was displayed by DF, which down-regulated IL-5 production and CCR4 expression, whereas IFN-gamma and CXCR3 were unaltered. CONCLUSIONS: Using a contact allergy model, we have demonstrated that various AIDs show distinct pharmacological profiles in that either type-1 or type-2 or both T cell responses are suppressed. These results should contribute to a more rational selection of AID in treating inflammatory skin diseases mediated by either or both of these T cell subsets. PMID- 15663562 TI - Biological effects of montelukast, a cysteinyl-leukotriene receptor-antagonist, on T lymphocytes. AB - BACKGROUND: Montelukast (MNT), a cysteinyl-leukotriene receptor (Cys-LTR) antagonist, has anti-inflammatory activity in the treatment of allergic diseases. If this effect is due only to blocking leukotrienes or also owing to inhibiting proliferation and survival of inflammatory cells, is actually unknown. OBJECTIVE: Testing the hypothesis that MNT could influence T lymphocyte functional behaviour in vitro. METHODS: Normal T lymphocytes were analysed for surface expression of Cys-LTR(1) and Cys-LTR(2) by means of monoclonal antibodies (mAbs), in the resting state and after activation with T helper type 2 cytokine or T cell receptor (TcR) stimulation. Proliferative activity, as well as IL-4 andIFN-gamma production, were simultaneously determined in samples exposed to molar concentrations of MNT from 10(-8) to 10(-5). Programmed cell death in cultured samples was evaluated by means of propidium iodide and fluorescein isothiocyanate conjugated anti-Annexin V mAb staining. The complementary DNA microarray technique was adopted to identify gene products involved in apoptosis induction. RESULTS: Resting T cells expressed low levels of Cys-LTR. Upon anti-CD3 mAb activation, a progressive increase in Cys-LTR(1) and -LTR(2) expression was observed. Exposure to MNT reduced proliferative response to TcR engagement, increased IFN-gamma production and led to apoptosis at minimal concentrations of 10(-6) M. A progressive loss in BAD and B cell lymphoma/leukaemia-2 activities, and an increase in the expression of CD27, TRAF3, TRAIL, p53 and Fas genes were also observed. CONCLUSIONS: Biological effects of MNT delineate a complex picture of gene activation and repression, probably induced by Cys-LTR blockade. The induction of apoptosis in allergen-specific T cell population, as a final result, appears fundamental in the treatment of asthma. PMID- 15663563 TI - The effectiveness of intranasal corticosteroids in combined allergic rhinitis and asthma syndrome. AB - BACKGROUND: Allergic rhinitis (AR) and asthma often coexist and may represent two manifestations of the same disease recently named combined AR and asthma syndrome (CARAS). AIM: To review the common pathophysiology of combined AR and asthma and to investigate the efficacy of intranasal corticosteroids (INCS). METHODS: Medline was used to identify articles relevant to mechanisms. A Cochrane systematic review was performed to assess the efficacy of INCS in CARAS. RESULTS: There is cross-talk, evidence of a common inflammatory response in both sites, linked by a systemic component. The efficacy of anti-inflammatory INCS on asthma outcomes was assessed in a systematic review of 12 randomized controlled trials involving 425 subjects. After INCS there were non-significant trends for improvement in asthma symptom score (standardized mean difference (SMD) of 0.61; P=0.07), forced expiratory volume in 1 s (SMD of 0.31; P=0.08), and morning peak expiratory flow (weighted mean difference of 36.51; P=0.06). There was no impact on methacholine airways responsiveness (SMD of -0.20; P=0.4). The review identified two promising new treatment options in united airway disease such as INCS as monotherapy in rhinitis and mild asthma, and a combined intranasal and intrabronchial corticosteroid (IBCS) deposition technique. CONCLUSION: Common mucosal inflammatory responses occur in CARAS. This systematic review shows trends for a benefit of INCS in CARAS, but recognizes that more research is needed. At this stage, the current best practice is to treat asthma conventionally with IBCS with or without beta(2)-agonist and to add INCS to improve specific rhinitis symptoms. PMID- 15663564 TI - Suppressive activity of fexofenadine hydrochloride on metalloproteinase production from nasal fibroblasts in vitro. AB - BACKGROUND: Allergic rhinitis (AR) is an inflammatory disease characterized by nasal wall remodelling with intense infiltration of eosinophils and mast cells/basophils. Matrix metalloproteinases (MMPs), MMP-2 and MMP-9, are the major proteolytic enzymes that induce airway remodelling. These enzymes are also important in the migration of inflammatory cells through basement membrane components. OBJECTIVE: We evaluated whether fexofenadine hydrochloride (FEX), the carboxylic acid metabolite of terfenadine with selective H(1)-receptor antagonist activity, could inhibit MMP production from nasal fibroblasts (NFs) in response to TNF-alpha stimulation in vitro. METHODS: NFs were established from nasal polyp derived fibroblasts (PFs) taken from patients with AR. Nasal mucosal fibroblasts (MFs) were also induced from nasal mucosal tissues from septal deformity patients without allergy. PF and MF (2 x 10(5) cells/mL, each) were stimulated with TNF alpha in the presence of various concentrations of FEX. After 24 h, culture supernatants were obtained and assayed for MMP-2, MMP-9, tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2 levels by ELISA. The influence of FEX on mRNA expression of MMPs and TIMPs in 4 h-cultured cells was also evaluated by real-time RT-PCR. Furthermore, nuclear factor-kappa B (NF-kappa B) activation in fibroblasts treated with FEX for 4 h was examined by ELISA. RESULTS: FEX at more than 350 ng/mL inhibited the production of MMP-2 and MMP-9 from both PF and MF in response to TNF-alpha stimulation, whereas TIMP-1 and TIMP-2 production was scarcely affected by FEX. FEX also inhibited MMP mRNA expression and NF-kappa B activation in PF and MF after TNF-alpha stimulation. CONCLUSION: The present data suggest that the attenuating effect of FEX on MMP-2 and -9 production from NFs induced by inflammatory stimulation may underlie the therapeutic mode of action of the agent on allergic diseases, including AR. PMID- 15663565 TI - Exhaled eicosanoids following oral aspirin challenge in asthmatic patients. AB - BACKGROUND: Biochemical analysis of expiratory breath condensate is an emerging non-invasive technique for assessment of airway inflammation. OBJECTIVE: We wondered whether application of expiratory breath condensate could facilitate diagnosis of aspirin-intolerant asthma and reproduce eicosanoids mediators' abnormalities described in this disease. METHODS: We measured prostaglandins (PGs) E(2), F(2 alpha), 9 alpha 11 beta F(2) and iso-F(2) by gas chromatography/mass-spectrometry and cysteinyl leukotrienes (cys-LTs) by radioimmunoassay in breath condensates of asthmatic patients undergoing oral aspirin challenge. Fourteen patients with aspirin-induced asthma and 20 aspirin tolerating asthmatics, most of them on chronic inhaled corticotherapy, were studied and compared with 10 healthy subjects. Additionally, plasma 9 alpha 11 beta PGF(2), the metabolite of PGD(2) and urinary leukotriene (LT) E(4) were measured before and following the challenge. RESULTS: At baseline, PG did not differ between the groups, except for lower 9 alpha 11 beta PGF(2) in aspirin intolerant asthma. Their concentrations were not changed by the challenge. Breath condensate cys-LTs were similar in the groups studied at base, and after aspirin challenge increased only in aspirin-intolerant patients. Elevated baseline urinary LTE(4) and its further increase following aspirin challenge was highly diagnostic for aspirin-intolerant asthma. The discriminatory value of cys-LTs increase in breath condensates was lower (72.8%) than either basal (99%) or post challenge increase (94%) of urinary LTE(4). CONCLUSIONS: In asthmatic patients on chronic corticotherapy measurement of urinary LTE(4) excretion rather than cys LTs in breath condensate is of greater value for diagnosis of aspirin hypersensitivity. PMID- 15663566 TI - Reticular basement membrane thickening in airways of lung transplant recipients is not affected by inhaled corticosteroids. AB - INTRODUCTION: Chronic rejection is a major problem for all lung transplant programmes, which is functionally manifested by fixed airflow limitation, Bronchiolitis Obliterans Syndrome (BOS). The inclusion of a Pre-BOS category, BOS(0 approximately p), in newly revised guidelines, recognizes the potential importance of early changes. We have previously demonstrated reticular basement membrane (Rbm) thickening in clinically stable lung transplant recipients free from BOS. The present study extends this, testing the hypothesis that inhaled corticosteroid (ICS) therapy will lead to a decrease in Rbm thickness in lung transplant recipients. METHODS: A parallel group, bronchoscopic intervention study of clinically stable lung allograft recipients, free from BOS, but with evidence of airway inflammation. Following baseline assessment of Rbm thickening, subjects were randomized to 3 months of either chlorofluorocarbon-driven beclomethasone diproprionate (BDP) 400 microg b.i.d., or a formulation designed to yield at least an equivalent dose, hydrofluoroalkane-driven BDP, 200 microg b.i.d. RESULTS: Three months treatment with a moderate dose of ICS, including a formulation designed for preferential small airway deposition, had no effect on Rbm thickening (13+/-3 vs. 14+/-5 microm post-ICS). CONCLUSION: Our data would suggest that airway remodelling can occur early in lung allografts and is not affected by moderate dose ICS therapy. Longitudinal studies are required to describe the pathophysiological processes involved in BOS, and specifically to elucidate potential relationships between airway remodelling, airflow obstruction and allograft failure. PMID- 15663567 TI - Latex allergy in infants younger than 1 year. AB - BACKGROUND: The prevalence of latex allergy in children is increasing worldwide. Previous multiple operations or atopic predisposition are known risk factors. In contrast, only sporadic cases of latex allergy have been reported in infants younger than 1 year, and the causative latex-containing products or symptoms in young infants have not been studied in detail. OBJECTIVE: The purpose of this study is to analyse the symptoms and risk factors of latex allergy in young infants. METHODS: Cases of latex allergy in infants younger than 1 year were studied in detail. Clinical course, causative latex-containing products were spotted and detailed analysis for latex allergy in patients and patients' parents was performed. CONCLUSION: We report nine cases of latex allergy in infants younger than 1 year. None of them have any abnormality or previous operations. Six patients had atopic eczema/dermatitis syndrome, one patient had bronchial asthma, whereas two patients had no overt allergic diseases. Symptoms of latex allergy were wheezing, swelling of face or lips, facial rash, or anaphylaxis, and causative latex-containing products were teat, pacifier, nose cleaner, teether, balloon, or enema tube. All of the nine patients had positive skin prick test to latex and extract from causative latex-containing products, whereas eight patients had positive serum latex-specific IgE. Study for family history revealed that latex allergy was noted in either father or mother in six patients, in both father and mother in one patient, whereas no latex allergy was noted in parents in two patients. It should be noted that all of these patients had latex-induced symptoms at home. Latex allergy in young infants may not be unusual. Physicians should be aware of latex allergy, and care should be taken to avoid contact with latex in young infants, especially when there is family history for latex allergy. PMID- 15663568 TI - Heparin-induced recurrent anaphylaxis. AB - BACKGROUND: Heparin-related immediate-type hypersensitivity reactions like urticaria, angio-oedema or bronchospasm are very rare, and only a few cases of anaphylaxis-like responses because of heparin have been described. However, the mechanisms underlying these reactions and the role of mast cells in their pathogenesis have not been elucidated. OBJECTIVES: We report a patient with end stage renal disease who presented with recurrent anaphylaxis after receiving heparin during haemodialysis. The underlying aetiology was obscured by the initiation of haemodialysis with its known anaphylactic-like side-effects. The diagnosis of hypersensitivity to heparin was confirmed by the clinical picture, positive skin tests and elevated serum tryptase levels. MATERIALS AND METHODS: We performed prick and intradermal skin tests with heparin, enoxaparin and danaparoid heparinoid. Total and mature tryptase levels were measured in serum by ELISAs at 1, 24 and 36 h following the reaction. RESULTS: An elevated mature tryptase level was found at 1 h, which returned to normal levels at 24 and 36 h. A high total tryptase level was detected at 1 h, but remained somewhat elevated at 24 h. Prick tests were negative with the three compounds. Intradermal skin tests with heparin and enoxaparin were both positive, while with danaparoid negative. Following negative skin test results, danaparoid was used as an anticoagulant during dialysis for the next 3 years without any adverse effects. CONCLUSIONS: In conclusion, we report the first case of heparin-induced anaphylaxis confirmed by an elevated level of mature tryptase in serum. Following skin tests, the patient was treated with danaparoid during haemodialysis sessions three times a week without any adverse effects. Because of increasing use of heparin in daily medical practice, physicians should be aware of possible immediate hypersensitivity reactions to this medication and know how to diagnose and treat them. PMID- 15663569 TI - Determination of the T cell epitopes of the lipocalin allergen, Rat n 1. AB - BACKGROUND: Laboratory animal allergy (LAA) is an important cause of occupational sensitization and asthma. Rats are a frequent cause of LAA and the major rat allergen, Rat n 1, is a member of the lipocalin protein family, which includes several other animal allergens such as the cow allergen, Bos d 2. To date, Bos d 2 is the only mammalian lipocalin allergen to have been studied in detail. OBJECTIVE: We undertook a cross-sectional study of a large population of individuals exposed to laboratory rats to determine the proliferative responses of peripheral blood mononuclear cells (PBMCs) to the major rat allergen, Rat n 1. METHODS: Eighty-three cases (defined by a positive skin prick test (SPT) > or =3 mm and/or a positive RAST > or =2% binding) and 274 referents without specific IgE to rats were tested for their proliferative responses of PBMCs to rat allergen. Cytokine release to rat urinary protein was examined in 28 sensitized and 42 non-sensitized exposed individuals. RESULTS: Proliferation to rat urinary protein was weak in all individuals. Four regions within Rat n 1 were identified as containing potential immunodominant T cell epitopes and three of these co localized within the conserved regions of the lipocalin molecule. All four regions within Rat n 1 overlapped considerably with the characterized epitopes of the lipocalin allergen, Bos d 2. IL-5 and ratios of IL-5/IFN-gamma were significantly increased in cases. CONCLUSION: The response to Rat n 1 is remarkably similar to the cow lipocalin allergen Bos d 2. T cell epitopes within lipocalins appear to co-localize with the conserved regions of the molecule. LAA is characterized by an increased production of IL-5. Investigation of other lipocalin allergens will provide further information about the allergenicity of this group of proteins. PMID- 15663570 TI - Molecular and structural analysis of immunoglobulin E-binding epitopes of Pen ch 13, an alkaline serine protease major allergen from Penicillium chrysogenum. AB - BACKGROUND: Through proteomic and genomic approaches we have previously identified and characterized an alkaline serine protease that is a major allergen (88% frequency of IgE binding) of Penicillium chrysogenum (Pen ch 13). OBJECTIVE: The aim of the present study is to identify the linear IgE-binding epitopes of Pen ch 13. METHODS: IgE-binding regions were identified by dot-blot immunoassay using 11 phage-displayed peptide fragments spanning the whole molecule of Pen ch 13. The minimal epitope requirements for IgE binding were further defined with overlapping peptides synthesized on derivatized cellulose membranes using SPOTs technology. The critical residues on the immunodominant epitopes were mapped through site-directed mutagenesis. The locations of the IgE epitopes identified were correlated with a three-dimensional structure of Pen ch 13. RESULTS: IgE antibodies in 35 serum samples reacted with at least one of the 11 peptide fragments of Pen ch 13. Peptide f-2n (residues 31-61) showed a high-intensity and the highest frequency (77%) of IgE binding. The frequencies of IgE binding to peptide f-4 (residues 93-133), f-1 (residues 1-37) and f-7 (residues 168-206) were 51%, 34% and 31%, respectively. SPOTs assay narrowed down the region of IgE binding of f-2n to residues 48-55 (GHADFGGR). Three, two and one epitope(s) that are four to nine amino acids in length, within f-4, f-1 and f-7, respectively, were found. Site-directed mutagenesis of Pen ch 13 revealed that substitution of His49 and/or Phe52 on Pen ch 13 with methionine resulted in proteins with drastic loss of IgE binding in seven sera tested. Proteins with amino acid replacements at residues 15-18 (RISS), or at residues 112 (I) and 116 (D) have lower IgE binding reactivity in one of the two patient's sera tested. Substituting residues 117 (W), 119 (V) and 120 (K) also block most of the IgE binding in one of the two patient's sera tested. In addition, replacing residues 203 (V) and 204 (D) along with a deletion at residue 206 (Y) diminished the IgE binding in two serum samples tested. A model was constructed based on the structure of P. cyclopium subtilisin protease that has >90% (256 out of 283 amino acids) sequence identity with Pen ch 13. The major epitope (GHADFGGR) on Pen ch 13 formed a loop-like structure and was located at the surface of the allergen. CONCLUSIONS: Several linear IgE-reactive epitopes and their critical core amino acid residues were identified for the Pen ch 13 allergen. The major linear IgE-binding epitope, 48GHADFGGR55, formed a loop-like structure at the surface of the allergen. Substitution of His49 and/or Phe52 with methionine significantly reduced IgE binding to Pen ch 13. Mapping of these results on a 3D model of the allergen provides valuable information about the molecular basis of allergenicity for Pen ch 13 and for designing specific immunotherapeutics. PMID- 15663571 TI - Expression of tyrosine hydroxylase and neuropeptide tyrosine in mouse sympathetic airway-specific neurons under normal situation and allergic airway inflammation. AB - BACKGROUND: The traditional neurotransmitter catecholamine and the neuropeptide tyrosine in sympathetic airway nerves have been proposed to be involved in the pathogenesis of airway diseases. OBJECTIVE: The aim of the present study was to investigate the effect of allergic airway inflammation on the expression of catecholamine enzyme tyrosine hydroxylase (TH), neuropeptide tyrosine (NPY) and tachykinins in mouse sympathetic airway ganglia. METHODS: Using neuronal tracing in combination with immunohistochemistry, the present study was designed to characterize TH, NPY and tachykinin profiles of superior cervical (SCG) and stellate ganglia after allergen challenge. RESULTS: The vast majority of fast blue-labelled SCG neurons (allergen: 97.5+/-1.22% (mean+/-SEM) vs. controls: 94.5+/-1.48%, P=0.18) and stellate neurons (allergen: 95.3+/-1.01% vs. controls: 93.6+/-1.33%, P=0.34) were immunoreactive for TH. Of the TH immunoreactive and fast blue-labelled SCG neurons, 52.0+/-1.01% allergen vs. 51.2+/-3.58% controls (P=0.83) and stellate neurons, 57.3%+/-0.97 allergen vs. 56.4+/-1.65% controls (P=0.64) were positive for TH only but not NPY, whereas 45.3+/-1.05% allergen vs. 43.3+/-1.18% controls (P=0.47) of fast blue-labelled SCG neurons and 37.9+/-0.86% allergen vs. 37.1+/-1.24% controls (P=0.62) of fast blue-labelled stellate neurons were immunoreactive for both TH and NPY immunoreactivities. There was a trend of an increase, but not significant one, in the percentage of TH-/NPY immunoreactive and fast blue-labelled neurons in allergen-treated animals in comparison with the controls. Tachykinins, however, were not expressed by sympathetic neurons and were also not induced in sympathetic neurons after allergen challenge. CONCLUSION: The present study indicates that allergic airway inflammation does not alter the expression of noradrenalin and NPY in sympathetic ganglia and also shows that sympathetic neurons do not respond to allergic airway inflammation with tachykinins induction. However, a participation of catecholamine and NPY in the pathogenesis of allergic airway inflammation cannot be excluded in the present study as a higher neurotransmitter output per neuron following allergen challenge could be possible. PMID- 15663573 TI - The research assessment exercise and academic obstetrics and gynaecology. PMID- 15663574 TI - Cell salvage in obstetrics: the time has come. PMID- 15663575 TI - The future of hysterectomy. PMID- 15663576 TI - Gender differences of placental dysfunction in severe prematurity. AB - OBJECTIVE: Rates of several obstetric complications have been reported to vary with fetal gender. We investigated whether a sex difference exists in findings at placental histology of extreme prematurity. DESIGN: Case-control study. SETTING: University Hospital. POPULATION: Four hundred and thirty-seven consecutive deliveries before 32 weeks of gestation of singleton, liveborn, non-anomalous infants. METHODS: Obstetric, neonatal and placental histologic findings were compared between male (n= 232) and female (n= 205) neonates. MAIN OUTCOME MEASURES: Histologic evidence of acute placental inflammation, uteroplacental vascular pathology, intraplacental vascular pathology and chronic placental inflammation. RESULTS: Male fetuses had similar racial distributions, rate of nulliparity, maternal age, gestational age at delivery, placental weight and fetoplacental weight ratio as female fetuses, but higher birthweight centiles (41 [27] vs 33 [26]). Placental histology showed no association between fetal gender and lesions of acute inflammation (P= 0.08), intraplacental vascular pathology (P= 0.16) or uteroplacental vascular pathology (P= 0.83). However, lesions of chronic inflammation had a significantly higher score in male than in female fetuses (P= 0.001). When we examined the distribution of chronic placental inflammation, significantly more severe lesions were noted in male than in female fetuses at the implantation site (i.e. the area of interstitial trophoblast invasion of the maternal decidua and maternal endovascular trophoblast remodelling), than within the placental villi (chronic villitis) or in the amniochorionic membranes (where interstitial trophoblast invasion is minimal). CONCLUSION: In premature deliveries at <32 weeks, male fetal gender is associated with placental lesions suggestive of a maternal immune response against the invading interstitial trophoblast. The immunologic basis of these findings deserves further studies. PMID- 15663577 TI - Determinants and consequences of discrepancies in menstrual and ultrasonographic gestational age estimates. AB - OBJECTIVES: To assess the association between maternal and fetal characteristics and discrepancy between last normal menstrual period and early (<20 weeks) ultrasound-based gestational age and the association between discrepancies and pregnancy outcomes. DESIGN: Hospital-based cohort study. SETTING: Montreal, Canada. SAMPLE: A total of 46,514 women with both menstrual- and early ultrasound based gestational age estimates. MAIN OUTCOME MEASURES: Positive (last normal menstrual period > early ultrasound, i.e. menstrual-based gestational age is higher than early ultrasound-based gestational age, so that the expected date of delivery is earlier with the menstrual-based gestational age) discrepancies > or =+7 days, mean birthweight, low birthweight, stillbirth and in-hospital neonatal death. RESULTS: Multiparous mothers and those with diabetes, small stature or high pre-pregnancy body mass index were more likely to have positive discrepancies. The proportion of women with discrepancies > or =+7 days was significantly higher among chromosomally malformed and female fetuses. The mean birthweight declined with increasingly positive differences. The risk of low birthweight was significantly higher for positive differences. Associations with fetal growth measures were more plausible with early ultrasound estimates. CONCLUSIONS: Although most discrepancies between last normal menstrual period- and early ultrasound-based gestational age are attributable to errors in menstrual dating, our results suggest that some positive differences reflect early growth restriction. PMID- 15663578 TI - Significant paternal contribution to the risk of small for gestational age. AB - OBJECTIVE: The aim of this study is to investigate both maternal and paternal contributions in the familial aggregation of small for gestational age. DESIGN: Nested case-control study. SETTING: Metropolitan area of Haguenau, France. POPULATION: Data were drawn from a French population-based maternity registry. After selection, 256 cases born either small for gestational age or average for gestational age were included. METHODS: Controlling for known pregnancy-related risk factors, logistic regression models were used to determine the risk of the child being small for gestational age, given that the mother, father or both were small for gestational age, and to examine interactions between maternal small for gestational age and pregnancy risk factors. MAIN OUTCOME MEASURES: Specifically, we investigate to what extent having either or both parents born small for gestational age increases the risk of small for gestational age in their offspring, after controlling for the established risk factors of small for gestational age and maternal and paternal characteristics. We also explore the extent to which the intergenerational predictors of small for gestational age may modify the effect of current pregnancy-related risk factors. RESULTS: The risk of a small for gestational age offspring was 4.7 times greater for mothers and 3.5 times greater for fathers who were small for gestational age, compared with average for gestational age counterparts. Furthermore, the risk of a small for gestational age offspring was 16.3 times greater when both parents were small for gestational age. No significant interactions between maternal small for gestational age and maternal smoking, hypertension or parity were observed. CONCLUSION: These results indicate that small for gestational age in both mother and father significantly influences the risk of their offspring being small for gestational age. While previous research has indicated that the birth outcome of the mother is an important determinant of the birth outcome of her offspring, these data indicate that the birth outcome of the father plays an equally critical role in determining fetal growth, strongly suggesting a genetic component in the familial aggregation of small for gestational age. PMID- 15663579 TI - Fetal heart rate patterns and ECG ST segment changes preceding metabolic acidaemia at birth. AB - OBJECTIVES: To compare the rates of abnormal ST segment patterns of the ECG and cardiotocographic (CTG) abnormalities in fetuses with metabolic acidaemia at birth and controls. To evaluate the inter-observer agreement in interpretation of ST analysis and CTG. DESIGN: Case-control study. SETTING: Three University hospitals in southern Sweden. POPULATION: Cases and controls were selected from the Swedish randomised controlled trial on intrapartum monitoring, including 4966 fetuses monitored with a scalp electrode. METHODS: Two obstetricians independently assessed the CTG and ST traces of 41 fetuses with metabolic acidaemia at birth and 101 controls, blinded to group, outcome and all clinical data. They classified each CTG trace and ST analysis as abnormal or not abnormal, and whether there was indication to intervene according to the CTG or to the CTG + ST guidelines. If their classification differed, assessment by a third obstetrician determined the final classification. MAIN OUTCOME MEASURES: Rates of CTG and ST abnormalities and decisions to intervene. Rates of inter-observer agreement. RESULTS: CTG was classified as abnormal in 50% and ST in 63% of cases with acidaemia, and in 20% and 34% of controls, respectively. CTG abnormalities were judged to be indication for intervention in 45% and CTG + ST abnormalities in 56% of cases with acidaemia, and in 15% and 8% of controls, respectively. The proportion of agreement between the two initial observers was significantly higher for ST abnormalities (94%) than for CTG abnormalities (73%), and for indication to intervene according to CTG + ST (89%) than according to CTG alone (76%). CONCLUSIONS: The inter-observer agreement rate was higher for a decision to intervene based on CTG + ST than on CTG alone. PMID- 15663580 TI - Pre- and periconceptional primary cytomegalovirus infection: risk of vertical transmission and congenital disease. AB - OBJECTIVE: To estimate the risk of congenital cytomegalovirus infection and disease following primary maternal infection around the time of conception compared with the risk during later stages of pregnancy. DESIGN: Cohort study between 1990 and 2003. SETTING: Germany. PARTICIPANTS: One hundred and sixty-six pregnant women with serologically confirmed primary cytomegalovirus infection and known outcome. METHODS: Timing of primary cytomegalovirus infection by analysing the kinetics of cytomegalovirus-specific IgG and IgM antibodies, the IgG avidity index and neutralising antibodies. MAIN OUTCOME MEASURE: Onset of maternal primary infection in relation to congenital infection and disease. RESULTS: Preconceptional (between eight and two weeks before onset of the last menstrual period) was determined in three women and did not lead to congenital infection. Periconceptional infection (between one week before and five weeks after last menstrual period) occurred in 20 women with congenital infection in nine cases (45%). Timing was less precise (between eight weeks before and five weeks after last menstrual period) in an additional 10 women, three cases of which resulted in congenital infection. Of the 12 pregnancies in which congenital infection occurred, seven were terminated, six before the 12th week of gestation (WG 12) and one at WG 19 due to fetal hyperechogenic bowel. One of the five infected live born infants delivered to a mother with periconceptional infection showed dystrophy and mild microcephaly at birth, but had a rather normal development at two years of age. Primary infections occurring between WG 6-20 and WG 20-38 resulted in transmission rates of 30% (27/89) and 58% (18/31), respectively. CONCLUSIONS: Counselling of women with periconceptional primary cytomegalovirus infection should be adjusted to offer prenatal diagnosis and high-level ultrasound controls due to the considerable risk for fetal infection and uncertainty of clinical outcome and disease. PMID- 15663581 TI - Proteomic biomarker analysis of amniotic fluid for identification of intra amniotic inflammation. AB - BACKGROUND: Intra-amniotic inflammation is associated with poor neonatal outcome independent of prematurity. We applied proteomic technology (SELDI: surface enhanced laser desorption ionisation) to identify the proteomic profile of intra amniotic inflammation. DESIGN: One hundred and four samples of amniotic fluid were analysed. In stage 1, samples from patients with symptoms of preterm labour and known outcomes were tested to identify the characteristic profile for inflammation. We extracted the profile using a novel, stepwise logical approach comparing SELDI tracings from patients who delivered preterm and had intra amniotic inflammation in response to infection to the tracings of patients who had symptoms of preterm labour but delivered at term. In stage 2, we applied the algorithm to samples from pregnancies whose outcomes were unknown to the investigators. SETTING: North-American university in collaboration with Ciphergen field demonstration laboratory. SAMPLE: One hundred and four samples of human amniotic fluid from transabdominal amniocentesis. METHODS: SELDI (surface enhanced laser desorption ionisation) and Mass Restricted analysis, a novel algorithm for extraction of clinical and biological relevant biomarkers from proteomic SELDI tracings. MAIN OUTCOME MEASURE: Presence of intra-amniotic inflammation and/or infection leading to preterm birth. RESULTS: Patients with intra-amniotic inflammation that deliver preterm have a distinctive amniotic fluid proteomic profile of three or four of the following proteins: neutrophil defensins-1 and -2, and calgranulins A and C. Based on the presence or absence of these biomarkers, we devised the mass restricted (MR) score ranging from 0 (all biomarker peaks absent) to 4 (all biomarker peaks present). In stage 1, MR score > 2 had 92.9% sensitivity (95% CI 76.5-98.9) and 91.8% specificity (95% CI 80.4 97.7) for detection of intra-amniotic inflammation. In blind testing (stage 2), MR score > 2 provided 100% specificity and sensitivity (95% CI 100-100). A MR score > 2 was associated with imminent preterm delivery. CONCLUSION: Proteomic analysis of amniotic fluid reveals the presence of biomarkers characteristic of intrauterine inflammation. This methodology may identify the subgroup of patients that might benefit most from interventions to prevent fetal damage in utero. PMID- 15663582 TI - Cardiovascular and endocrine effects of a single course of maternal dexamethasone treatment in preterm fetal sheep. AB - OBJECTIVE: To determine the effects of a single course of maternally administered dexamethasone on preterm fetal sheep in utero. DESIGN: Prospective randomised controlled trial. SETTING: University laboratory. SAMPLE: Pregnant sheep at 0.7 of gestation. METHODS: Pregnant ewes at 103 days of pregnancy (term = 147 days) were given two intramuscular injections of vehicle (n= 7) or 12 mg of dexamethasone (DEX; n= 8) 24 hours apart. Fetuses were continuously monitored for five days. MAIN OUTCOME MEASURES: Fetal mean arterial blood pressure, carotid and femoral arterial blood flow and vascular resistance, heart rate, heart rate variability, fetal plasma cortisol and ACTH and fetal body movements. RESULTS: DEX injections led to an acute increase in mean arterial blood pressure with a rise in carotid and femoral vascular resistance, a fall in femoral arterial blood flow, and a brief fall in fetal heart rate followed by significant tachycardia. From 24 hours after the injections, mean arterial blood pressure and vascular resistance returned to control values, however, a mild tachycardia [200 (3) vs 184 (4) bpm, P < 0.05] and loss of the circadian pattern of fetal heart rate variability persisted until the end of recording. Plasma ACTH and cortisol were markedly suppressed by DEX (P < 0.05), with values returning to control levels 32 and 72 hours after the first injection, respectively. There was no effect on basal fetal heart rate variability, body movements, carotid arterial blood flow, or the circadian pattern of fetal heart rate. CONCLUSION: In contrast to previous experiments utilising direct fetal infusion of steroids, maternal administration of DEX was associated with only transient hypertension. PMID- 15663583 TI - Comparison of digital and ultrasonographic examination of the cervix in predicting time interval from induction to delivery in women with a low Bishop score. AB - OBJECTIVE: To compare pre-induction ultrasonographic cervical length and Bishop score in predicting time to delivery after labour induction with prostaglandins. DESIGN: Prognostic cohort study. SETTING: Tertiary referral maternity unit in a teaching hospital. POPULATION: Two hundred and sixty-six women with singleton pregnancies at between 34(+0) and 41(+3) weeks of gestation requiring induction of labour with prostaglandins for medical indications. METHODS: A secondary analysis of a trial comparing two prostaglandins. Assessment of the Bishop score and measurement of the cervical length by transvaginal sonography were performed by two operators, blinded to each other's results. We estimated the predictive effects on the outcomes of ultrasonographic cervical length and Bishop score. MAIN OUTCOME MEASURE: Time intervals from induction to delivery and to vaginal delivery. RESULTS: Cervical length and Bishop score were associated with the time interval from induction to delivery, based on univariable analyses. When considered jointly in a multivariable model, only the Bishop score was significantly related to the outcome: The higher the Bishop score, the higher the hazard to delivery [hazard ratio (HR): 1.2, 95% confidence interval (CI): 1.1 1.3], illustrating that once the Bishop score is taken into account, further knowledge of cervical ultrasound length (HR: 0.99, 95% CI: 0.98-1.0) did not add any predictive information. Also, Bishop score was predictive of time interval between induction and vaginal delivery (HR: 1.2, 95% CI: 1.1-1.4) while cervical length had no additional predictive value (HR: 0.99, 95% CI: 0.98-1.0) when both cervical length and Bishop score were introduced in the model. CONCLUSIONS: The Bishop score appears to be a better predictor of the time interval from induction to delivery and to vaginal delivery than cervical length after induction of labour for medical reasons. PMID- 15663584 TI - Domestic violence, lifetime trauma and psychological health of childbearing women. AB - OBJECTIVE: Although histories of abuse are associated with psychiatric illness in women, health professionals rarely enquire directly about such experiences. This study examined the association between physical and sexual violence and lifetime trauma and depressive and posttraumatic stress symptoms in women receiving maternity care. DESIGN: Cross sectional study. SETTING: South London Hospital maternity services. POPULATION: Two hundred women receiving postnatal or antenatal care. METHODS: Two hundred women receiving postnatal or antenatal care at a South London maternity service were screened for lifetime experiences of trauma and domestic violence. Information was obtained about self-harming behaviour, suicidal thoughts and attempts and psychiatric history. Women completed the Edinburgh Postnatal Depression Scale (EPDS) and the Posttraumatic Diagnostic Scale (PTDS). MAIN OUTCOME MEASURES: RESULTS: One hundred and twenty one (60.5%) women reported at least one traumatic event and two-thirds of these had experienced multiple traumatic events. The most frequent (34%) was witnessing or experiencing physical assault by a family member. Forty-seven (23.5%) women had experienced domestic violence. Physical and sexual abuse commonly co occurred. Thirteen (10.7%) women with a trauma history had current posttraumatic stress disorder. Severe posttraumatic symptoms were associated with physical and sexual abuse histories and repeat victimisation. Adult and childhood physical and sexual abuse histories were also associated with more severe depressive symptomatology. Significant social factors associated with depression were being single, separated or in a non-cohabiting relationship. CONCLUSION: Traumatic events are under-recognised risk factors in the development of depressive and posttraumatic stress symptoms in childbearing women. Childhood abuse creates a vulnerability to re-traumatisation in adulthood. Awareness of the impact of trauma and abuse on psychological health may enable more appropriate targeting of clinical services and support for women receiving maternity care. PMID- 15663585 TI - The effect of the Term Breech Trial on medical intervention behaviour and neonatal outcome in The Netherlands: an analysis of 35,453 term breech infants. AB - OBJECTIVE: To examine the effects of the Term Breech Trial on the medical behaviour of Dutch obstetricians and on neonatal outcomes. DESIGN: Retrospective observational study. SETTING: The Netherlands. POPULATION: Infants born at term in breech presentation in the Netherlands between 1998 and 2002, with birthweights < or =4000 g (n= 33,024) and >4000 g (n= 2429), respectively. Multiple pregnancies, antenatal death and major congenital malformations were excluded. METHODS: Data derived from the Dutch Perinatal Database were used to compare modes of delivery and neonatal outcome of infants born in breech position in the 33 months preceding publication of the Term Breech Trial and in the 25 months thereafter. MAIN OUTCOME MEASURES: Incidence of planned and emergency caesarean section, vaginal breech delivery, perinatal death, 5-minute Apgar score and birth trauma. RESULTS: Within two months after publication of the Term Breech Trial, the overall caesarean rate increased from 50% to 80% and has remained stable thereafter. In the group of infants < or =4000 g, this was associated with a significant decrease of perinatal mortality from 0.35% to 0.18%, a decrease of the incidence of a 5-minute Apgar score <7 from 2.4% to 1.1% and a decrease of birth trauma from 0.29% to 0.08%. In the (small) group of infants >4000 g, a similar trend was observed. CONCLUSIONS: The Term Breech Trial has resulted in an exceptionally rapid change in medical behaviour by Dutch obstetricians. This change was followed by improved neonatal outcome. PMID- 15663586 TI - Physical symptoms after childbirth: prevalence and associations with self-rated health. AB - OBJECTIVE: The aims of the present study were to describe the prevalence of a number of physical symptoms, as described by women themselves, two months and one year after childbirth in a national Swedish sample and to investigate the association between specific symptoms and women's self-rated health. DESIGN: Cohort study. SETTING: Swedish antenatal clinics. POPULATION: A total of 2413 women recruited from 593 antenatal clinics in Sweden during three one-week periods evenly spread over one year (1999-2000), representing 54% of women eligible for the study and 76% of those who consented to participate. The representativity of the sample was assessed by comparison with the total Swedish birth cohort of 1999. METHODS: Data were collected by means of questionnaires in early pregnancy, two months and one year after the birth and from the Swedish Medical Birth Register. MAIN OUTCOME MEASURES: Self-reported symptoms and self rated health. RESULTS: Tiredness, headache, neck, shoulder and low back pain were common problems at two months as well as one year after childbirth. At two months, pain from caesarean section, dyspareunia and haemorrhoids were frequent problems, whereas stress incontinence was often reported at one year. Ninety-one percent of the women said self-rated health was 'very good' or 'good' at two months after birth, and 86% at one year. Low self-rated health was associated with symptoms that affected general physical functioning and wellbeing, such as tiredness, headache, musculoskeletal problems, mastitis, perineal pain, dysuria, stomachache and nausea. Complaints related to more specific situations, such as dyspareunia, constipation and stress incontinence were not associated with self rated health. CONCLUSION: Despite the fact that physical symptoms were common two months and one year after the birth, the vast majority of women rated their health as 'very good' or 'good'. PMID- 15663587 TI - Outcome of term breech births: 10-year experience at a district general hospital. AB - OBJECTIVE: To review the short and long term outcomes among singleton infants with breech presentation at term delivered in a geographically defined population over a 10-year period. DESIGN: Retrospective, cohort study. SETTING: District General Hospital. POPULATION: 1433 term breech infants alive at the onset of labour and born between January 1991 and December 2000. METHODS: Data abstracted from birth registers, neonatal discharge summaries and the child health database system were used to compare the short and long term outcomes of singleton term breech infants born by two different modes of delivery (prelabour caesarean section and vaginal or caesarean section in labour). Fisher's exact test was used to compare the categorical variables. MAIN OUTCOME MEASURES: Short term outcomes: perinatal mortality, Apgar scores, admission to the neonatal unit, birth trauma and neonatal convulsions. Long term outcomes: deaths during infancy, cerebral palsy, long term morbidity (development of special needs and special educational needs). RESULTS: Of 1433 singleton term infants in breech presentation at onset of labour, 881 (61.5%) were delivered vaginally or by caesarean section in labour and 552 (38.5%) were born by prelabour caesarean section. There were three (0.3%) non-malformed perinatal deaths among infants born by vaginal delivery or caesarean section in labour compared with none in the prelabour caesarean section cohort. Compared with infants born by prelabour caesarean section, those delivered vaginally or by caesarean section in labour were significantly more likely to have low 5-minute Apgar scores (0.9% vs 5.9%, P < 0.0001) and require admission to the neonatal unit (1.6% vs 4%, P= 0.0119). However, there was no significant difference in the long term morbidity between the two groups (5.3% in the vaginal/caesarean section in labour group vs 3.8% in the prelabour caesarean group, P= 0.26); no difference in rates of cerebral palsy; and none of the eight infant deaths were related to the mode of delivery. CONCLUSIONS: Vaginal breech delivery or caesarean section in labour was associated with a small but unequivocal increase in the short term mortality and morbidity. However, the long term outcome was not influenced by the mode of delivery. PMID- 15663588 TI - Vaginal hysterectomy for benign disorders in obese women: a prospective study. AB - OBJECTIVE: To compare the morbidity of vaginal hysterectomy in obese and non obese women in a single institution. DESIGN: Obese and non-obese women with benign uterine disorders matched for age, parity and race underwent vaginal hysterectomy without laparoscopic assistance. Peri-operative outcome complications were compared. Thirty-eight women had a BMI over 30 kg/m(2) (study group) and 178 women had a BMI below 30 kg/m(2) (control group). SETTING: Gynaecologic department of a university hospital. POPULATION: Women who were referred to our department with an indication of vaginal hysterectomy for benign disorders. METHODS: Case control study. MAIN OUTCOME MEASURES: Peri-operative complications, the fall in the haemoglobin concentration, the duration of the procedure, the length of the hospital stay and uterine weight were analysed. RESULTS: Mean BMI was 33.2 and 23.7 kg/m(2) in the study and control groups, respectively. None of the obese women had severe co-morbidity contraindicating surgery. There were no significant differences in surgical or anaesthetic risk factors, including parity, hormonal status, pre-existing disease and estimated mean uterine weight. The overall complication rates were 14% and 16% in the obese and non-obese groups, respectively (P= 0.7). Obesity did not increase the duration of the procedure (48 [7] minutes vs 50 [10] minutes, P= 0.1) or the length of hospitalisation (5.8 [1.4] days vs 5.5 [1.2] days, P= 0.2). CONCLUSION: Vaginal hysterectomy can be successfully performed, with acceptable morbidity, in obese women. PMID- 15663589 TI - The effect of tibolone in postmenopausal women receiving tamoxifen after surgery for breast cancer: a randomised, double-blind, placebo-controlled trial. AB - OBJECTIVE: To assess the effects of tibolone on climacteric symptoms, endometrium and serum lipid/lipoproteins in postmenopausal women receiving tamoxifen after surgery for breast cancer. DESIGN: Double-blind, randomised, placebo-controlled, multicentre pilot study. SETTING: Hospital outpatient clinic. SAMPLE: Seventy postmenopausal women receiving tamoxifen following surgery for early breast cancer. METHODS: Women received 20 mg/day oral tamoxifen plus either 2.5 mg/day oral tibolone or placebo for 12 months. MAIN OUTCOME MEASURES: Frequency and severity of hot flushes (diary cards); intensity of hot flushes and sweats (Landgren scale); interference of hot flushes and sweats with normal life; frequency and intensity of other climacteric symptoms; endometrial thickness and histology; vaginal bleeding; breast cancer recurrence and serum lipid/lipoproteins. RESULTS: Daily card data showed no change in the daily number of hot flushes with either tibolone or placebo (P= 0.219) after three months. There was a significant reduction in the severity of flushes with tibolone compared with placebo (-0.4 vs 0.2, P= 0.031). The Landgren scale showed a mean change in the number of hot flushes of -0.6 with tibolone and +1.1 with placebo after 12 months (P= 0.022). Endometrial biopsies were normal and vaginal bleeding was similar in both groups. A significant decrease in triglycerides (-23% vs 1.4%) and HDL (-12% vs 19%) was seen with tibolone compared with placebo after 12 months. CONCLUSIONS: Tibolone prevented an increase in hot flushes in postmenopausal women given tamoxifen following surgery for breast cancer without untoward effects on the endometrium. Beneficial effects on serum lipid profile were noted. PMID- 15663590 TI - The effectiveness of live lactobacilli in combination with low dose oestriol (Gynoflor) to restore the vaginal flora after treatment of vaginal infections. AB - OBJECTIVE: To evaluate the effectiveness of live lactobacilli in combination with low dose oestriol for restoration of the vaginal flora after anti-infective treatment. DESIGN: The study was designed as a single centre, randomised, placebo controlled, double-blind clinical trial. SETTING: University Hospital. SAMPLE: Three hundred and sixty women out of 1750 were randomised. METHODS: Three hundred and sixty women with the complaints of vaginal infections (bacterial vaginosis, candidiasis, trichomoniasis or fluor vaginalis) were randomly assigned two to seven days after the end of the anti-infective therapy, to therapy with live lactobacilli in combination with low dose oestriol (study group, n= 240) or placebo (n= 120). The follow up visits occurred three to seven days and four to six weeks after the end of the restoration therapy. MAIN OUTCOME MEASURES: The Normal Flora Index (NFI), which consists of numbers of lactobacilli, pathogenic microorganisms, leucocytes and vaginal pH, was used as the primary outcome of the study. Secondary outcomes included the total symptoms score, the degree of purity of the vaginal flora and the global assessment of the treatment by the investigator and the women. RESULTS: During restoration therapy, the NFI increased significantly more in the study group than in the control group in both first and second control visits (P= 0.002 and P= 0.006, respectively). The degree of purity of the vaginal flora also increased significantly more in the study group compared with the control group (P < 0.0001 and P= 0.001, respectively). No serious adverse event was reported during restoration therapy. CONCLUSION: Restoration of the vaginal flora can be significantly enhanced by the administration of live lactobacilli in combination with low dose oestriol. PMID- 15663591 TI - Reactivation of ocular toxoplasmosis during pregnancy. AB - We wished to assess whether ocular toxoplasmosis can be reactivated during pregnancy in immunocompetent females, and whether such reactivation is confined to the eye or whether the fetus is exposed to the risk of vertical transmission. For this purpose, we retrospectively examined 18 females with ocular toxoplasmosis during the course of 35 pregnancies. Of these 18 patients, seven developed recurrences during seven pregnancies. Due to the potential risk of functional damage to the mother and the possibility of vertical disease transmission to the fetus, we suggest following such cases carefully during pregnancy until the dimensions of the problem are more fully appreciated. PMID- 15663592 TI - Measurement of beliefs about effectiveness of laparoscopic uterosacral nerve ablation. AB - To explore gynaecologists' 'prior' beliefs on effectiveness of laparoscopic uterosacral nerve ablation (LUNA), a structured survey was used to gather information on the distribution of their prior beliefs regarding the effects of LUNA on pelvic pain, both numerically [on a 10-point visual analogue scale] and by responses to a questionnaire. None of the 25 gynaecologists responding to the questionnaire stated that LUNA would increase pain, while two of the 25 gave numerical answers suggesting they believed that the intervention would worsen the pain. The most widely held 'prior belief', reflected in both questionnaire and numerical responses, was that LUNA would have a small beneficial effect on pain. PMID- 15663593 TI - The use of botulinum toxin type A (BOTOX) as treatment for intractable chronic pelvic pain associated with spasm of the levator ani muscles. PMID- 15663594 TI - Proteomics technology for the accurate diagnosis of inflammation in twin pregnancies. PMID- 15663595 TI - Aplasia cutis congenita and low molecular weight heparin. PMID- 15663596 TI - Review of the accuracy of various diagnostic tests for bacterial vaginosis to predict preterm birth (Honest et al., BJOG, May 2004). PMID- 15663597 TI - Quantifying severe maternal morbidity: a Scottish population study. PMID- 15663599 TI - An insight into the dendritic cells at the maternal-fetal interface. AB - The conditions that permit the genetically distinct fetus to survive and develop within the mother are among the most fascinating immunologic puzzles. The presence of dendritic cells in the maternal decidua pointed to a biologic role of antigen-presenting cells in maternal-fetal interaction. The method of study included recent findings on the lineage, maturity, phenotype and function of dendritic cells at the maternal-fetal interface. The increment of uterine dendritic cells occurs simultaneously with the decisive phase of gestation, when implantation takes place. Decidual dendritic cells of the first trimester pregnancy, with a phenotype characteristic of the mature myeloid lineage, express MHC class II, co-stimulatory and adhesion molecules, control Th1/Th2 balance and activate the proliferative response of autologous NK cells. Dendritic cells are specifically equipped to control immunity, to trigger immune response and also to maintain tolerance, avoiding the rejection of the conceptus by the maternal immune system. PMID- 15663600 TI - Protective effect of vitamin E in an animal model of LPS-induced inflammation. AB - PROBLEM: Many sterility outcomes may be associated to the presence of an inflammatory response that would lead to an inability of the endometrium to support implantation and maintain viable embryos. We have established an animal model of inflammation in which the systemic administration of lipopolysaccharyde (LPS) results in a low embryo implantation rate. The purpose of this work was to investigate the effect of the inflammatory agent LPS on embryo viability and to verify the ability of vitamin E to modulate the inflammatory effect of LPS on embryo viability. METHOD OF STUDY: For pre-implantation studies B6CBAF1 mice, which were intraperitoneally inoculated with LPS (4-10 mg/kg), were used. Mice were also treated with vitamin E (4-10 mg/kg) before or after LPS injection. Embryos were obtained from the oviduct after each treatment. RESULTS: The LPS produces a decrease in the number of pre-implantational embryos in a concentration dependent manner. The LPS effect can be partially reversed or prevented by vitamin E. Preliminary results show that inflammatory cytokines are secreted by intraperitoneal macrophages in LPS treated mice. CONCLUSION: Our results demonstrate the ability of vitamin E to avoid an inflammatory environment and to allow viability of embryos. PMID- 15663601 TI - Cleavage of integrin by mu-calpain during hypoxia in human endometrial cells. AB - PROBLEM: The distribution and activation of mu-calpain and possible cleavage of integrin in human endometrial cells under hypoxic condition were investigated. METHOD OF STUDY: Human endometrial epithelial and stromal cells were subjected to hypoxia, and subsequently used for immunostaining and western blot analysis. RESULTS: The proform of mu-calpain was detected in the cytoplasm of normal cells, and displayed a substantial decrease after hypoxia. Conversely, the active form of mu-calpain was not detected in normal cells, but was abundant after hypoxia. The cytoplasmic domain of integrin beta3 was also detected in the cytoplasm of endometrial cells. Western blot analysis confirmed that both the proform of mu calpain and the integrin beta3 cytoplasmic domain decreased during hypoxia. CONCLUSIONS: Mu-calpain is activated in human endometrial cells during hypoxia and that subsequent cleavage of the integrin beta3 cytoplasmic domain may give some adverse effects to the function of human endometrium. PMID- 15663603 TI - B- and T-cells in the follicular fluid and peripheral blood of patients undergoing IVF/ET procedures. AB - PROBLEM: To analyse percentage of total and memory CD27(+) B-cells and other lymphocyte subpopulations in the peripheral blood (PB) and follicular fluid (FF) of infertile married couples. METHOD OF STUDY: Forty-eight couples from in vitro fertilization/embryo transfer (IVF/ET) programme were divided into four groups: patients with previous unsuccessful fertilization (n = 13), ectopic pregnancy (n = 8), multiple (at least three) failed IVF/ET (n = 18) and missed abortions (n = 9). Control group consisted of 15 married couples with healthy children. RESULTS: PB memory CD27(+) B-cells were significantly decreased in all groups of infertile patients compared with controls. First group had increased memory B-cells percentages compared with the second group. The differences in the percentages of PB memory B-cells in third and fourth group compared with the first group were not statistically significant. FF memory B-cells in the first and third group were significantly increased compared with second and fourth group. The percentage of total FF B-cells in all groups were significantly decreased compared with their percentage in PB. Male partners of women from the first group had had significantly increased percentages of memory B-cells compared with the partners of women from the second group. Percentage of total T- and B-cells, CD4+ and CD8+ T-cells, NK cells and activated HLA-DR(+) T-cells in all groups were not significantly different from controls. We found no statistically significant difference between immunoglobulin E levels in all groups of patients. We found lower levels of IgA and IgM in FF compared with serum in all groups. CONCLUSION: Infertile patients have significantly decreased percentage of CD27(+) B-cells in the PB. Abnormalities in the memory B-cell compartment may contribute to the pathogenesis of infertility. In the T-cell compartment abnormalities were not detected. It appears that hormonal stimulation did not influence cellular immunity parameters. PMID- 15663602 TI - Ecology of danger-dependent cytokine-boosted spontaneous abortion in the CBA x DBA/2 mouse model. I. Synergistic effect of LPS and (TNF-alpha + IFN-gamma) on pregnancy loss. AB - PROBLEM: Previous data have shown "danger" signals, such as bacterial lipopolysaccharide (LPS) acting via toll-like (tlr) receptors are required for early pregnancy failure in several murine abortion models. Indeed, the abortion rate increased in the CBA x DBA/2 model after a gestation day (gd) 7.5 injection of tumour necrosis factor (TNF)-alpha + interferon (IFN)-gamma only if the LPS tlr signalling pathway was intact. High rates of cytokine-boosted abortion >80% loss can be achieved in certain animal colonies, that have a high endogenous (spontaneous) rate of resorption (30-50%). A specific role for LPS has been postulated to determine both the endogenous and cytokine-boosted losses. METHODS: To test the role of LPS in spontaneous and cytokine-boosted abortions, recombinant TNF-alpha + IFN-gamma, and LPS were injected in different doses and sequences intraperitoneally (i.p.) into CBA x DBA/2 mated mice in the Toronto General Research Institute animal facility where the endogenous abortion rate is <30%. The effects of poly IC, a tlr3 agonist that induces IFN-gamma that can reverse LPS-induced tolerance, and effects of anti-MD-1 on TNF-alpha induction by LPS, poly IC, CPG, or HSP in vitro were also examined. RESULTS: A high endogenous rate of loss similar to that seen in Clamart could be achieved by increasing exposure to LPS on the morning after mating (gd 0.5). The magnitude by which the abortion rate could be increased by an i.p. injection of 2000 u TNF-alpha + 1000 u IFN-gamma on gd 7.5 was independent of the endogenous rate of loss, and could not be increased by doubling the dose. One microgram of LPS given on day 7.5 achieved a similar rate of loss, and if given with the cytokines, synergistically boosted the rate of loss to near Clamart rates. LPS given 1 day prior to the cytokines abrogated the cytokine effect, whereas LPS given day 0.5 had no significant effect on the response to day 7.5 cytokine injection. Blocking MD-1 inhibited TNF-alpha stimulation by poly IC, LPS, CPG, or HSP in vitro, and reduced abortion rates. Poly IC did not avert LPS-type tolerance effects in vivo. CONCLUSIONS: High endogenous rates of abortion in the CBA x DBA/2 model may be explained by exposure to LPS at the time of mating. Increased rates of loss triggered by cytokines later in pregnancy may depend on increased absorption of LPS from intestinal flora. PMID- 15663606 TI - Correlation between lipophilicity and triptan outcomes. AB - BACKGROUND: It has been suggested that triptans achieving higher central nervous system (CNS) levels should have an advantage in efficacy, if central actions are important. Objective.-Our aim was to correlate the efficacy and tolerability results of triptans with their lipophilicity. METHODS: Data for response and pain free at 2 hours, recurrence, adverse events (AE), CNS AE, and chest symptoms taken from Ferrari et al's meta-analysis publications for the recommended doses of oral triptans were correlated with their lipophilicity coefficients (logD(pH)7.4 = -2.1 almotriptan < -1.5 sumatriptan < -1.0 zolmitriptan < -0.7 rizatriptan < -0.2 naratriptan < 0.5 eletriptan). RESULTS: We found no significant correlation between lipophilicity coefficients and any of the analyzed parameters. There was, however, some correlation between lipophilicity and CNS AE (P = .09, r = 0.74) and, to a lesser degree, with a reduction in recurrence rate (r = -0.36). The r values for response and pain free with placebo correction ranged from 0.04 to 0.34, suggesting almost no correlation between lipophilicity and efficacy variables. CONCLUSIONS: According to this analysis, a higher lipophilicity does not seem crucial to improve triptan efficacy. This physico-chemical property, however, correlates with higher CNS AE and, possibly, lower recurrence rates. PMID- 15663607 TI - Zolmitriptan 5 mg nasal spray: efficacy and onset of action in the acute treatment of migraine--results from phase 1 of the REALIZE Study. AB - OBJECTIVES: The objective of phase 1 (reported here) of this two-phase study was to assess the efficacy of zolmitriptan 5 mg nasal spray, in terms of ability to provide relief from all migraine symptoms, in a controlled setting, designed to replicate clinical practice. BACKGROUND: Zolmitriptan nasal spray has been shown to be fast acting and highly effective in the treatment of migraine, as assessed using standard endpoints, such as headache response and pain-free rates. METHODS: In the double-blind first phase of the study, patients with migraine were randomized to receive zolmitriptan 5 mg nasal spray or placebo to treat a single migraine attack. Attacks were treated according to patients' normal patterns of use, in order to closely reflect clinical practice; that is, no specific regimen was dictated in terms of time to treatment or at what level of pain intensity the headache should be treated. Patients could take a second dose of study medication or an agreed escape medication if adequate pain relief had not been achieved 2 hours after the first dose. The primary efficacy endpoint was total symptom relief (freedom from pain, nausea, photophobia, and phonophobia) 1 hour after the first dose. Secondary efficacy endpoints included headache response, pain-free status and sustained pain-free status, and ability to perform normal activities. RESULTS: The intention-to-treat population comprised 461 zolmitriptan nasal spray recipients and 451 placebo recipients. The total symptom relief rate 1 hour post dose was significantly higher in the zolmitriptan 5 mg nasal spray group than in the placebo group (14.5% vs. 5.1%; P < .0001); the difference between the groups was significant from 30 minutes post-dose. Treatment with zolmitriptan nasal spray, compared with placebo, also produced a higher headache response rate from 10 minutes post-dose (15.1% vs. 9.1%; P = .0079) and a higher pain-free rate from 30 minutes post-dose (7.7% vs. 3.2%; P = .0039). Zolmitriptan nasal spray was also significantly superior to placebo in terms of sustained pain-free status and patients' ability to perform normal activities. Zolmitriptan nasal spray was well tolerated. CONCLUSIONS: These findings confirm the efficacy demonstrated by zolmitriptan nasal spray in previous clinical trials. PMID- 15663608 TI - Zolmitriptan nasal spray exhibits good long-term safety and tolerability in migraine: results of the INDEX trial. AB - BACKGROUND: Previous studies have shown that zolmitriptan 5 mg nasal spray has a fast onset of action, high efficacy, and good tolerability in the acute treatment of migraine. Objective.-This open-label, noncomparative, multicenter, multinational phase III study was designed to further evaluate the long-term safety and tolerability of zolmitriptan 5 mg nasal spray in a population of migraineurs largely naive to triptan nasal sprays who treated multiple migraine attacks over a 1-year period. METHODS: Patients were required to have an established diagnosis of migraine with or without aura (based on International Headache Society criteria), a high frequency of migraine attacks, and were allowed to treat migraine with any baseline headache intensity. A secondary objective of the study was to assess the long-term efficacy of zolmitriptan nasal spray. A subgroup analysis aimed to determine whether rhinitis had any influence on outcomes of treatment. RESULTS: The safety population consisted of 538 patients who treated 20,717 migraine attacks with zolmitriptan 5 mg nasal spray. Overall, adverse events occurred in 32.8% of attacks, and led to treatment withdrawal in 4.5% of patients. The most common adverse events were unusual taste (19.0%) and paresthesia (6.8%). Adverse events were generally of mild intensity, transient, and well tolerated, showing a decline in incidence over time. Serious adverse events were rare. The presence of rhinitis and use of a second dose of trial medication had no effect on the incidence of adverse events. At 2 hours, 53.8% of attacks treated with zolmitriptan nasal spray 5 mg were rendered pain free. The highest 2-hour pain free rates were seen for headaches of mild baseline intensity (83.3%), followed by headaches of moderate (56.5%), and severe (32.2%) baseline intensity. The 2-hour pain-free rate remained consistent throughout the study period. The presence of rhinitis had no effect on efficacy. CONCLUSIONS: Zolmitriptan 5 mg nasal spray demonstrated a well-tolerated and efficacious profile in the acute treatment of multiple migraine attacks over a 1-year period. PMID- 15663609 TI - Low income and education levels may cause medication overuse and chronicity in migraine patients. AB - BACKGROUND: Frequent analgesic drug intake, especially in migraine patients, may induce the risk of medication overuse headache (MOH). The various conditions that may affect the development and the features of MOH have not been determined yet. AIM: To compare MOH patients with migraine as pre-existing headache and episodic migraine patients according to socioeconomic and educational variables. METHODS: Forty-six MOH patients with migraine as pre-existing headache and 61 migraine patients were included into study. The headache characteristics, socioeconomic and educational variables of MOH and migraine groups and subgroups divided according to the education (low/high education subgroups), and income (low/high income subgroups) levels were evaluated. RESULTS: We found that mean duration of education was shorter in MOH patients than migraine patients. There was a negative correlation between duration of education and duration of MOH. The mean duration of MOH was longer and rate of low-income level was higher in patients with low-education level. The duration of education was lower in MOH patients with low income. The frequency of migraine attacks and low-income rate was also higher in low-educated migraine patients. The duration of education was shorter in migraine patients with low income. CONCLUSION: We report that migraineurs with low socioeconomic status may have risk of developing MOH. A better identification of patients at risk of drug-associated headache may contribute to improved health in a group of patients with MOH. PMID- 15663610 TI - Migraine prevalence and some related factors in Turkey. AB - OBJECTIVES: The aim of our study is to determine the prevalence of migraine in Provincial Center of Edirne. METHODS: A systematic random sample of 386 inhabitants over 14 years old were interviewed face to face. Diagnosis of migraine was based on the International Headache Society (IHS) classification. RESULTS: Migraine was identified in 77 subjects (60 female, 17 male). The lifetime prevalence of migraine was found to be 19.9% (95% CI: 18.3-21.5) (9.34% (95% CI: 8.6-10) in males, 29.3% (95% CI: 27-31.6) in females). CONCLUSIONS: These results show that while migraine prevalence in Turkish females is higher than that reported in previous studies from Europe and United States, lifetime prevalence is similar in all populations. PMID- 15663611 TI - Development and positioning reliability of a TMS coil holder for headache research. AB - OBJECTIVE: Accurate and reproducible coil positioning is important for headache research using transcranial magnetic stimulation protocols. We aimed to design a transcranial magnetic stimulation coil holder and demonstrate reliability of test retest coil positioning. METHODS: A coil holder was developed and manufactured according to three principles of stability, durability, and three-dimensional positional accuracy. Reliability of coil positioning was assessed by stimulating over the motor cortex of four neurologically normal subjects and recording finger muscle responses, both at a test phase and a retest phase several hours later. RESULTS: In all four subjects, repositioning of the transcranial magnetic stimulation coil solely on the basis of coil holder coordinates was accurate to within 2 mm. CONCLUSIONS: The coil holder demonstrated good test-retest reliability of coil positioning, and is thus a promising tool for transcranial magnetic stimulation-based headache research, particularly studies of prophylactic drug effect where several laboratory visits with identical coil positioning are necessary. PMID- 15663612 TI - Comparison of intravenous valproate with intravenous lysine-acetylsalicylic acid in acute migraine attacks. AB - OBJECTIVE: The study compared efficacy and tolerability of intravenous valproate (iVPA) with intravenous lysine-acetylsalicylic acid (iLAS) in acute migraine attacks. Background.-iLAS has been proven to be a highly effective treatment in acute migraine attacks, but it is not available in many countries and contraindicated in patients with asthma or peptic ulcers. Current data suggest that iVPA may be effective in the treatment of acute migraine attacks. DESIGN/METHODS: In this randomized, double-blind, parallel-group phase-II study, 40 patients with acute migraine attacks (onset <5 hours, severe or moderate headache on a four-point IHS scale) alternately received iVPA 800 mg or iLAS 1000 mg. Primary outcome criteria were the percentage of patients reporting pain relief after 1 hour and patients who remained sustained pain free for 24 hours following drug administration. Secondary outcome criteria were relief of pain and associated migrainous symptoms (nausea, photophobia, and phonophobia) at 1, 2, 24, and 48 hours following drug administration. RESULTS: There were no significant differences in demographic and clinical features between both treatment groups. Percentage of pain relief after 1 hour in the iVPA and iLAS groups were 25% and 30%, respectively, and of sustained pain free for 24 hours were 20% and 30%, respectively, without significant differences (P = 1 and P= .72, respectively). Both drugs improved associated migrainous symptoms without significant differences at the different time points, but again with a trend in favor of iLAS. No adverse events were observed. CONCLUSION: Both drugs were effective in acute migraine attacks with a trend in favor of iLAS. As both drugs were well tolerated, further studies with higher doses of iVPA for the treatment of acute migraine attacks are recommended. PMID- 15663613 TI - Migraine therapy: a survey of pharmacists' knowledge, attitudes, and practice patterns. AB - OBJECTIVE: Study pharmacists' knowledge, attitudes, and practice patterns with regard to migraine therapy. BACKGROUND: Pharmacists interact with headache sufferers at least 53,000 times daily, thus are well positioned to improve the less than optimal medication management of these disorders. Methods.-Two hundred self-administered surveys, distributed at a migraine symposium, assessing pharmacists' demographic characteristics and level of agreement or disagreement with treatment approaches were conducted. RESULTS: A total of 171 useable surveys (86%) were received. Of the sample, 35% were community pharmacists, 29% were hospital pharmacists, and the remainder were from other work environments. Exclusively among community pharmacists, 80% feel that headache is an important part of their practice, 85% make between one and five over-the-counter (OTC) headache product suggestions per day, and 12% make six or more daily OTC recommendations. Among all the sample's pharmacists, more than half feel migraine patients should try OTC drugs prior to prescription medications, only half ask patients about headache-related morbidity, and one-third feel migraine-specific medications should be reserved only for patients who initially fail nonspecific drugs. Few pharmacists utilize published migraine treatment guidelines. Approximately two-thirds of pharmacists do not feel migraine is a neurobiological illness. The majority is comfortable with their ability to identify people needing a physician referral. CONCLUSIONS: Our results show pharmacists, particularly those in community pharmacies, interact with headache sufferers multiple times daily. Most pharmacists were neither familiar with nor practice migraine therapies endorsed by evidence-based guidelines. Further training of pharmacists is warranted. PMID- 15663614 TI - Chronic post-traumatic neck and head pain. AB - OBJECTIVE: This is a review of current concepts of chronic post-traumatic neck and head pain. In this article, I will emphasize the physiological and sociological aspects of these disorders. BACKGROUND: The pathophysiology of chronic post-traumatic neck and head pain has not been well understood. Some have emphasized the organic factors and others the psychogenic aspects of these conditions. Only in recent years have this dichotomy been integrated with sociocultural concepts. METHODS: The history of chronic post-traumatic head and neck pain is reviewed. Paradoxes are discussed, ie, the great differences in prevalence around the world, the inconsistent relationship of symptoms to degree of trauma, the curious phenomena of structural disease without symptoms, and symptoms without structural disease. The organic and pathophysiologic factors are reviewed, then those factors that modulate pain in these conditions are discussed. CONCLUSION: Chronic post-traumatic neck and head pain is rarely either organic or psychogenic. Rather physiological, social, and cultural factors play major roles in modulating pain and either perpetuate or ameliorate these chronic pain conditions. PMID- 15663615 TI - MRI of a brain under pressure. PMID- 15663616 TI - The young woman with postpartum "thunderclap" headache. PMID- 15663617 TI - Intracranial hypertension induced by rofecoxib. AB - Several secondary causes are implicated in the etiology of idiopathic intracranial hypertension. Rofecoxib is a selective cyclo-oxygenase (COX)-2 inhibitor, now being increasingly used in place of nonselective nonsteroidal anti inflammatory drugs (NSAIDs). We report a case of intracranial hypertension in a 69-year-old man 3 weeks after the commencement of rofecoxib therapy with reversal of clinical findings on drug withdrawal. PMID- 15663618 TI - Spontaneous intracranial hypotension presenting as mental deterioration. AB - A 55-year-old woman had new onset of postural headache followed by change of mental status 3 weeks later. Magnetic resonance imaging (MRI) of the brain and whole spine showed typical spontaneous intracranial hypotension (SIH) findings, bilateral subdural hematoma, and cerebrospinal fluid leakage over the T7-T9. Her headache and mentality improved after epidural blood patches. Early recognition and correct diagnosis are crucial for successful treatment in patients with SIH presenting with mental confusion. PMID- 15663619 TI - Cluster-like headache aura status. AB - We describe a patient with successive attacks (40 to 90 minutes) of cluster-like headache associated with aphasia, and contralateral hemihypesthesia and hemiplegia. The condition can best be described as cluster-like headache aura status. PMID- 15663620 TI - Cluster headache with obstructive sleep apnea and periodic limb movements during sleep: a case report. AB - We report the case of a man with episodic cluster headache who suffered from severe obstructive sleep apnea (OSA) as well as periodic limb movements during sleep (PLMS). His attacks of headache occurred primarily during sleep being timely to REM sleep as 90 to 120 minutes interval. OSAs were more frequent and prolonged during REM sleep and oxygen saturation decreased to 81% during this sleep period. Periodic limb movements were also observed in our patient that were more frequent during the first half of the polysomnographic recordings. This case is one of the few reporting cases with CH who had both OSA and PLMS. PMID- 15663621 TI - Peripheral postganglionic sympathicoplegia mimicking cluster headache attacks. AB - After dissection with complete occlusion of the internal carotid artery, a 58 year-old man started to suffer from intense cluster headache-like attacks. Magnetic imaging showed signs of nonsymptomatic cerebral emboli, which could be dated to have occurred in temporal relation to the start of the attacks, all on the right side. This case and two similar ones indicate that peripheral postganglionic sympathicoplegia can cause attacks with similar pain characteristics, accompanying symptoms, duration, and regularity as in cluster headache. PMID- 15663622 TI - Severe headaches following smallpox vaccination. AB - Headaches are common following smallpox vaccination; the re-introduction of civilian vaccination necessitates better understanding of the clinical features and outcome of postvaccination headache. We identified patients reporting headache following vaccination from among those reported to the U.S. Vaccine Adverse Events Reporting System to characterize demographic and clinical features. One-hundred and eight reports were obtained from among 627 smallpox vaccine-related reports, including 15 hospitalized persons. None had neurologic dysfunction or acute laboratory abnormalities; headache resolved in all except 2 hospitalized patients within 3 months. Severe headache following smallpox vaccination is generally transient, but debilitating headache may occur and further characterization is needed. PMID- 15663624 TI - Migraine, epilepsy, and brain neuronal hyperexcitation. PMID- 15663633 TI - QTc interval and QTc dispersion during haemodiafiltration. AB - BACKGROUND AND AIM: Our aim was to evaluate QTc interval and QTc dispersion in 27 end-stage renal disease (ESRD) patients undergoing Acetate Free Biofiltration (AFB) in order to ascertain any correlations between the electrrocardiographic (ECG) parameters, serum Na+, K+, Ca++, Mg++ and intraerythrocytic Mg++ (Mg++e) concentrations. All measures were made at t0 (session beginning), t1 (first hour), t2 (second hour), t3 (third hour), and t4 (session end). RESULTS: Blood pressure, heart rate, bodyweight and total ultrafiltration in the three dialysis sessions were constant. A significant progressive increase occurred in serum Ca++ during the sessions, while there was a significant diminution in serum K+. The pattern for Mg++ concentrations in serum and erythrocytes differed: in serum it decreased, whereas Mg++e increased. At t4, the QTc interval was reduced to a significant extent with respect to the baseline value. QTc dispersion significantly increased at t1 without there being significant variations at other times with respect to t0. At t2, t3 and t4, values promptly returned to baseline levels. QTc had a negative correlation with serum Ca++ levels at t4. In contrast, an inverse correlation was found between QTc dispersion and serum K+ at t1. No other correlations could be found between any other electrolytes, QTc interval or QTc dispersion. CONCLUSION: In conclusion, the decrease observed in the QTc interval at the end of an AFB session was inversely related to serum Ca++ concentrations. Moreover, an increase in QTc dispersion occurred during the first hour of the session, and was negatively correlated with serum K+. PMID- 15663634 TI - Increases in peritoneal small solute transport in the first month of peritoneal dialysis predict technique survival. AB - BACKGROUND: Peritoneal transport of small solutes generally increases during the first month of peritoneal dialysis (PD). The aim of this study was to prospectively evaluate the ability of the peritoneal equilibration test (PET), carried out 1 and 4 weeks after the commencement of PD, to predict subsequent technique survival. METHODS: Fifty consecutive patients commencing PD at the Princess Alexandra Hospital between 1 February 2001 and 31 May 2003 participated in the study. Paired 1 week and 1 month PET data were collated and correlated with subsequent technique survival. RESULTS: A significant increase was observed in the dialysate : plasma creatinine ratio at 4 h (D/P Cr) between 1 and 4 weeks after the onset of PD (0.55 +/- 0.12 vs 0.66 +/- 0.11, P <0.001). Mean death censored technique survival was superior in patients who experienced > or =20% rise in D/P Cr during the first month of PD compared with those who did not (2.3 +/- 0.2 vs 1.6 +/- 0.2 years, P <0.05). Using a multivariate Cox proportional hazards model analysis, the significant independent predictors of death-censored technique survival were an increase in D/P Cr of greater than 20% during the first month (adjusted hazard ratio [HR] 0.20, 95% CI 0.05-0.75), the absence of diabetes mellitus, the absence of ischaemic heart disease, body mass index and baseline peritoneal creatinine clearance. CONCLUSIONS: A 20% or greater rise in D/P Cr during the first month of commencing PD is independently predictive of PD technique survival. Further investigations of the mechanisms underlying this phenomenon are warranted. PMID- 15663635 TI - Ultrasonography in the management of exit site infections in peritoneal dialysis patients. AB - AIM: To assess the efficacy of using ultrasonography (USG) in monitoring the progress of exit site infection (ESI) in patients undergoing continuous ambulatory peritoneal dialysis (CAPD). METHODS: Twenty-two cases of newly diagnosed ESI and 20 cases with normal exit sites as controls were assessed by using USG. The exit sites were reassessed by using USG after finishing a course of antibiotic therapy, and the sonographic findings were correlated with the clinical outcome. RESULTS: Out of the 22 cases of ESI, 21 cases had definite sonolucent zones around the external cuffs, while one case had normal sonographic findings. Of the 20 control cases of normal exit sites, 16 had normal sonographic findings, and four had sonolucent zones around the external cuffs. Exit site infections correlated with positive sonographic findings as compared to normal exits (P <0.0001). The 21 cases of ultrasonic-positive ESI were re-examined after antibiotic therapy, and 10 of these had a post-treatment sonolucent rim around the distal cuff < or =1 mm thick, while 11 cases were persistently > mm thick. The former group was shown to have a more favourable outcome (P=0.013). And despite variable USG findings, all eight patients with Pseudomonas aeruginosa related ESI had an unfavourable clinical outcome. CONCLUSION: Ultrasonography of the exit sites in CAPD patients is a useful adjunctive tool in the management of ESI. A sonolucent zone around the external cuff >1 mm thick following a course of antibiotic treatment and the involvement of the proximal cuff are associated with poor clinical outcome. In ESI caused by Pseudomonas aeruginosa, the clinical outcome was uniformly poor irrespective of the sonographic findings. PMID- 15663636 TI - Periodic limb movement disorder and restless legs syndrome in dialysis patients. AB - BACKGROUND: Sleep disturbances, in particular restless legs or limb movements, during the night are often reported by uremic patients. However, polysomnography (PSG) studies have never been carried out to confirm the actual occurrence of these disorders and the association with other objective and self-reported sleep wake data. METHODS: Forty-eight participants were subjected to a 2-day PSG. These data on sleep including periodic limb movements, which are associated with restless legs, were correlated with clinical observations, quality of sleep-wake and life questionnaires, and with biochemical and neurographical measures. RESULTS: Restless legs syndrome (RLS) was observed in 58.3% of the patients and periodic limb movement disorder (PLMD) occurred in 70.8% of the patients. PLMD was revealed polysomnographically in almost 90% of the RLS patients. Patients with both PLMD and RLS had significantly poorer sleep quality than those with neither disorder or with PLMD alone, both in terms of self-reported data and the PSG. Quality of life was significantly worse in patients with RLS and PLMD compared to those patients with neither disorder. PLMD patients also tended to have a lower quality of life. All other metabolic measures and the results of a nerve conduction test were not correlated with RLS and/or PLMD. CONCLUSION: There was a high prevalence of severe RLS and PLMD in the present sample of uraemia patients. Nearly all RLS patients had severe PLMD. RLS (in combination with PLMD) in dialysis is associated with poor sleep quality, insomnia complaints, depression and emotional distress. Our results suggest that PLMD per se is also clinically relevant. PMID- 15663637 TI - Non-convulsive status epilepticus and consciousness disturbance after star fruit (Averrhoa carambola) ingestion in a dialysis patient. AB - Star fruit ingestion may induce severe neurological complications in chronic renal failure patients. We present a case on maintenance dialysis therapy who developed a consciousness disturbance without convulsion after eating star fruit. The symptoms became aggravated after haemodialysis. The brain computed tomography scan showed no abnormal findings, but the electroencephalogram found active focal sharp waves in the left central regions and diffusion-weighted magnetic resonance imaging also showed hyperintense lesions in the left central regions that were compatible with non-convulsive status epilepticus. His condition improved dramatically after anticonvulsant therapy and regular haemodialysis. The patient was discharged 20 days later without neurological sequela. PMID- 15663638 TI - Anti-oxidant status in relation to lipoproteins, leptin and pro-inflammatory cytokines in children with steroid-sensitive nephrotic syndrome. AB - BACKGROUND: Reactive oxygen species and cytokines are reported to play a role in the proteinuria of nephrotic syndrome. The aim of this study was to investigate indirect evidence of oxidant activity together with leptin, lipoproteins and pro inflammatory cytokines in children with steroid-sensitive nephrotic syndrome. METHODS: A total of 40 children with steroid-sensitive nephrotic syndrome (20 with newly onset or relapse comprised group I and 20 in remission while receiving steroids comprised group II) and 20 sex and age matched healthy control children were included. The following indirect parameters of oxidant activity were determined: serum malondialdehyde, erythrocyte superoxide dismutase, catalase and whole-blood-reduced glutathione. Serum leptin, lipids and lipoproteins were also determined. RESULTS: Similar glutathione, increased malondialdehyde levels and decreased superoxide dismutase and catalase activity were observed in group I patients compared with controls. There was no significant difference in these variables between group I and group II (P >0.05). Tumour necrosis factor-alpha and interleukin-6 concentrations were similar in patients and controls. Concentrations of interleukin-1beta and interleukin-8 were higher in the active phase of nephrotics compared with controls (P <0.05). Significant positive correlations were found between malondialdehyde and interleukin-1beta, interleukin-6, leptin and lipoprotein (a) (P <0.05). There were significant negative correlations between anti-oxidants and leptin, lipoprotein (a) and several cytokines (P <0.05). CONCLUSIONS: Changes in the concentrations of malondialdehyde, superoxide dismutase, catalase and glutathione are compatible with increased amounts of oxidation in steroid-sensitive nephrotic syndrome. Leptin and pro-inflammatory cytokines may be related to excessive protein permeability in nephrotic syndrome. PMID- 15663639 TI - Caveolins in the repair phase of acute renal failure after oxidative stress. AB - Ischaemia-reperfusion and toxic injury are leading causes of acute renal failure (ARF). Both of these injury initiators use secondary mediators of damage in oxygen-derived free radicals. Several recent publications about ischaemia reperfusion and toxin-induced ARF have indicated that plasma membrane structures called caveolae, and their proteins, the caveolins, are potential participants in protecting or repairing renal tissues. Caveolae and caveolins have previously been ascribed many functions, a number of which may mediate cell death or survival of injured renal cells. This review proposes possible pathophysiological mechanisms by which altered caveolin-1 expression and localization may affect renal cell survival following oxidative stress. PMID- 15663640 TI - Managing Henoch-Schonlein purpura in children with fish oil and ACE inhibitor therapy. AB - BACKGROUND: Henoch-Schonlein purpura (HSP) is a vasculitic syndrome with palpable purpura and renal involvement. The treatment for HSP with persistent renal disease remains controversial. The kidney biopsy in HSP shows IgA deposits and fish-oil therapy has proven to be promising in halting the progression of IgA nephropathy. METHODS: Five children with biopsy-proven HSP with repeated episodes of haematuria and proteinuria were treated with fish oil (1 g orally twice daily). In three of the five patients an angiotensin-converting enzyme inhibitor (ACEI) was added for hypertension. RESULTS: The mean duration of follow up after starting fish-oil therapy was 49.2 weeks. The protein excretion rate prior to starting fish oil was 1041 mg/day and on the last follow-up visit the rate had decreased to 104 mg/day (P <0.05). The average blood pressure (BP) prior to therapy was 135/82. On the last follow-up visit the average BP off ACEI had decreased to 100/54 (P <0.05). After a year of follow up serum creatinine and glomerular filtration rates have remained stable at 51.2 micromol/L and 128 mL/min/1.73 m2, respectively. CONCLUSION: This is the first report of abatement of HSP with fish oil and ACEI in children. There is a need for randomized prospective trials to confirm this observation. PMID- 15663641 TI - Glomerulosclerosis: a paraneoplastic phenomenon? AB - Glomerulosclerosis is not classically considered a paraneoplastic glomerular lesion. Focal and segmental glomerulosclerosis (FSGS) has rarely been reported in association with solid tumours. We report three cases of FSGS and an additional case of collapsing glomerulosclerosis in patients presenting with nephrotic syndrome and malignancy. PMID- 15663642 TI - Multicentric Castleman's disease complicated by secondary renal amyloidosis. PMID- 15663643 TI - Does asymptomatic hyperuricaemia contribute to the development of renal and cardiovascular disease? An old controversy renewed. AB - Recent studies in both humans and experimental animals have led to renewed interest in uric acid and its association with hypertension, cardiovascular events and renal disease progression. This has also refuelled a longstanding debate regarding the precise role of this ubiquitous breakdown product of purine metabolism in these disease processes. Various lines of evidence suggest that uric acid may have a direct role in the pathogenesis of hypertension and vascular disease. Regardless of this possibility, it is apparent that serum uric acid levels serve as a powerful 'biomarker' or independent predictor of prognosis and outcome in certain renal, cardiovascular and cerebrovascular diseases. Whether these outcomes can be improved by specifically treating asymptomatic hyperuricaemia remains inadequately resolved at this stage. Data from various animal studies suggests that lowering uric acid levels may be of benefit, but the crucial human studies are still lacking. This review will examine some of the recent evidence supporting a causal and contributory role for uric acid in cardiovascular and renal disease. How clarification of the role of uric acid may guide future treatment strategies will also be discussed. PMID- 15663644 TI - Medicinal herbal extracts--renal friend or foe? Part two: herbal extracts with potential renal benefits. AB - In this second of two articles regarding the renal toxicities or benefits of medicinal herbs, herbs are reported as being 'potentially beneficial' to the kidneys if there is strong in vivo evidence of renal protection from toxic substances or drugs; potent, specific renal anti-oxidant effects; in vivo cancer antiproliferative effects specific to the kidneys; or in vivo evidence of being beneficial in renal disease or failure. Among the herbs, polyherbal formulae and fungi with potential renal benefits are Cordyceps sinensis, Sairei-to, Rheum spp., Salvia miltiorrhiza and its component, magnesium lithospermate B and others. PMID- 15663645 TI - Efficacy and side-effect profile of sevelamer hydrochloride used in combination with conventional phosphate binders. AB - BACKGROUND: Poor phosphate control is common among patients with end-stage renal disease. Sevelamer hydrochloride has been demonstrated to be a safe and effective phosphate binder when used as a monotherapy. However, cost limits its usefulness in many countries. Data assessing its effectiveness and safety in combination with conventional phosphate binders are lacking. METHODS: Dialysis patients meeting the following inclusion criteria participated in this study: (i) hyperphosphataemia >1.8 mmol/L (5.6 mg/dL); and (ii) an inability to tolerate currently available binders. The trial was conducted in three phases each lasting 3 months: (i) an observation phase (patients continued on their regular phosphate binders); (ii) a titration phase (sevelamer was added at a dose of 403 mg three times daily with meals, titrated to a maximum of 1209 mg three times daily); and (iii) a maintenance phase. RESULTS: Twenty-five patients were recruited into the study. Eighteen patients completed all three trial phases. Mean serum phosphate dropped from 2.11 +/- 0.06 mmol/L (6.6 +/- 0.2 mg/dL) during the observation period to 1.91 +/- 0.01 mmol/L (5.9 +/- 0.003 mg/dL) during the maintenance phase (P=0.02). Calcium x phosphate product fell from 5.49 +/- 0.17 mmol2/L2 (68.64 +/- 2.11 mg2 dL2) to 4.89 +/- 0.27 mmol2/L2 (61.36 +/- 3.35 mg2 dL2) (P=0.02). There was no significant change in serum calcium or parathyroid hormone. Total serum cholesterol fell from 3.8 mmol/L (3.4-4.37) 147 mg/dL (131-169) to 3.55 mmol/L (2.97-4.2) 137 mg/dL (115-162) (P=0.02). Serum low-density lipoprotein cholesterol also fell significantly from 1.67 +/- 0.10 mmol/L (65 +/- 4 mg/dL) to 1.52 +/- 0.11 mmol/L (59 +/- 4 mg/dL) (P=0.04). The average prescribed dose of sevelamer was 2.4 g/day. Elemental calcium dropped from 3.4 g/day (1.4 to 4.6) to 1.2 g/day (0.6-2.4) (P=0.04). Seventy-two per cent of patients reported mild flatulence, nausea and indigestion. Three patients discontinued treatment because of adverse effects. CONCLUSIONS: Sevelamer in combination with conventional phosphate binders is effective in lowering serum phosphate and calcium-phosphate product in patients with refractory hyperphosphataemia. Beneficial effects on lipid profile were also observed. Mild gastrointestinal upset is common. PMID- 15663646 TI - Effectiveness of high trough levels of cyclosporine for 5 months in a case of steroid-dependent nephrotic syndrome with severe steroid toxicity. AB - Glucocorticoid treatment for steroid-dependent nephrotic syndrome (NS) is associated with severe adverse effects, such as bone fractures and epidural lipomatosis. Furthermore, a high trough level of cyclosporine (CsA) over an extended period of time is known to induce CsA nephropathy. We present a girl with steroid-dependent NS and steroid-induced vertebral compression fractures and epidural lipomatosis who was treated with a high-dose of prednisolone after experiencing several relapses. A high CsA trough level (between 147 and 225 ng/mL) over a period of only 5 months was effective in improving the vertebral compression fractures, alleviating the epidural lipomatosis by enabling the discontinuation of prednisolone treatment. Thus, high trough levels of CsA over a short period of time may enable prednisolone to be discontinued in cases of steroid-dependent NS without causing any clinical, histological, serum and/or urinary CsA-related adverse effects. PMID- 15663647 TI - Switchover to generic cyclosporine in stable renal transplant recipients: a single unit experience. AB - BACKGROUND: The introduction of the immunosuppressant cyclosporine has significantly improved renal transplant survival. It is an expensive drug and generic alternatives may offer cost advantages. However, generic alternatives must be shown to provide equivalent therapeutic efficacy and safety. This study reports our experience of a switch from the microemulsion formulation of cyclosporine, Neoral (Novartis), to the generic equivalent, Cysporin (Mayne Pharma). METHOD: A two-period, single-sequence, cross-over study was done to compare cyclosporine blood levels and the area under the curve (AUC) of Neoral with Cysporin 2 weeks after a 1:1 dose switch. cyclosporine blood levels were measured at time points 0, 2, 4 and 8 h (C0, C2, C4, C8) after the switch. The cyclosporine AUC at 0-4 h and 0-12 h were calculated using the trapezoidal method. The two formulations were considered to result in equivalent blood levels if the 95% confidence interval (CI) of the ratio of the two levels was within 0.8 1.25. RESULTS AND CONCLUSION: A total of 38 stable renal transplant patients aged 49.79 +/- 11.38 years (mean +/- SD), who were 7.84 +/- 3.97 years postrenal transplantation, were studied. The Neoral dose at the time of the switch was 2.38 +/- 1.21 mg per kg bodyweight. At all measured time points the 95% CI for the cyclosporine drug level ratio was between 0.9 and 1.15. There were no significant adverse events during the period of study. We conclude that the generic formulation of cylosporin, Cysporin, after a 1:1 switch from Neoral results in equivalent blood levels in stable renal transplant recipients. After switchover cyclosporine levels at C0 or C2 can continue to be monitored as per the institution's current monitoring practice. PMID- 15663648 TI - Reversal of pancytopenia following kidney transplantation in a patient of primary hyperoxaluria with bone marrow involvement. AB - Combined liver and kidney transplantation is the ideal treatment for patients with end-stage renal failure secondary to primary hyperoxaluria and systemic oxalosis, with a functioning liver providing replacement of the deficient enzyme and a functioning kidney providing the route of excretion for the oxalate crystals. Pancytopenia from bone marrow infiltration of oxalate crystals is a rare complication of primary hyperoxaluria, and its reversal following transplant has not been described. We report the first case of pancytopenia from marrow infiltration by oxalate crystals reversing following a successful kidney transplant alone. Although kidney alone transplants do not provide the best chance of survival or quality of life as compared to a combined kidney and liver transplant, a well functioning kidney transplant is able to take care of the systemic oxalate load and ameliorate, at least for a period of time, the systemic complications of oxalosis. PMID- 15663649 TI - Epidermolytic hyperkeratosis: a keratin 1 or 10 mutational event. AB - Epidermolytic hyperkeratosis is an unusual type of ichthyosis. This inherited keratinization disorder is characterized clinically by erythema, blistering, and peeling shortly after birth. It may resolve and be replaced with thick scaling. It can lead to life-threatening complications, such as sepsis. Histologically, there is a hyperkeratosis and vacuolar degeneration. Genetically, this is an autosomal dominant disease with complete penetrance; however, 50% are spontaneous mutations. The clinical phenotype is a result of alterations in the gene(s) for keratin 1 and/or 10. We review this disorder and its therapy, which is mainly symptomatic with emollients and retinoids. PMID- 15663650 TI - Prevalence of different skin conditions in an outpatients' setting in north western Nigeria. AB - BACKGROUND: Information on skin diseases in north-western Nigeria is scanty. We therefore conducted a prospective survey of 2611 new patients seen between August 1999 and July 2001 at the consultant medical/dermatology clinics of Usmanu Danfodiyo University Teaching Hospital, Sokoto, Nigeria. METHODS: The personal bio-data of all consecutive patients with skin diseases were documented. They were examined and the diagnoses, which were based on the history of presentation and physical examination, were recorded. Bacteriologic, mycologic, and histologic confirmations were obtained as appropriate. RESULTS: Seven hundred and forty-six patients (28.6%) had skin diseases. There were 407 male and 339 female patients aged between 0.17 and 90 years; the median age was 27 years. The median ages for male and female patients were 28 and 25 years, respectively. There were 900 skin diseases seen in the 746 patients. The majority of skin diseases were found amongst the younger age group, which represented over 85% of the patients. Infectious and parasitic skin diseases accounted for 44.4% of cases; eczema, acne, papulosquamous, and pigmentary skin disorders were observed in 14.1%, 7.0%, 6.4%, and 6.0% of cases, respectively. CONCLUSIONS: Preventable skin diseases are common in north-western Nigeria and predominantly affect individuals in the highly productive age group. Health education is therefore necessary to curb their spread, reduce the associated morbidity, and improve the health status of the population. A concerted effort should be made to train health workers in the diagnosis and treatment of the more common dermatologic conditions. PMID- 15663651 TI - Lesional T-cell subset in post-kala-azar dermal leishmaniasis. AB - BACKGROUND: The pathogenesis of post-kala-azar dermal leishmaniasis (PKDL) is ill understood. This study was carried out to find the percentage of T-helper and T suppressor cells in lesional tissue and their probable role in the pathogenesis of PKDL. METHODS: An immunoperoxidase monoclonal antibody technique was used to characterize and quantify the subsets of T lymphocytes in the infiltrate in 25 patients with PKDL. RESULTS: The ratio of T-helper to T-suppressor cells was 0.87 in hypopigmented macules and 0.85 in nodule and/or plaque lesions of PKDL. CONCLUSIONS: In this study, there was a definite preponderance of T-suppressor cells over T-helper cells in both types of skin lesions of PKDL. Further studies should be undertaken on larger numbers of patients to compare T-cell subsets both in skin lesions and the circulation, in order to determine the pathogenesis of PKDL. PMID- 15663653 TI - Becker's melanosis: a report of 12 cases with atypical presentation. AB - Becker's melanosis is a distinct clinical entity. It usually presents in adolescence as a unilateral, hyperpigmented, hairy cutaneous hamartoma. We report herein 12 cases of Becker's melanosis that differ in their presentation from classical Becker's melanosis, and we believe that such presentations are not uncommon. PMID- 15663652 TI - Imiquimod is a strong inhibitor of tumor cell-induced angiogenesis. AB - BACKGROUND: Imiquimod, a potent immunomodulator, not having a direct antiproliferative activity, was found to be effective in genital and cutaneous premalignancies and malignancies. As tumor development depends on blood vessel supply, the inhibition of angiogenesis could be responsible for the antitumor activity. OBJECTIVE: To find in a murine model whether imiquimod has antiangiogenic activity and whether this activity is mediated by locally induced cytokines. METHODS: The study was performed in two cell lines: Skv human keratinocytes containing multiple integrated copies of HPV16 derived from bowenoid papulosis, and murine L1 lung sarcoma cells of Balb/c mice. The murine model of cutaneous angiogenesis was used to assess and count the new blood vessel formation. The mice were immunosuppressed by a total body X-ray irradiation and treated with 5% or 2.5% imiquimod cream before or after induction of angiogenesis with intradermally injected tumor cell suspension. In some experiments the mice were, in addition, treated intraperitoneally with monoclonal antibodies against murine IFNalpha, TNFgamma or IL-18. RESULTS: Topical application of imiquimod on the murine skin resulted in reduction of angiogenesis (P < 0.001) induced by intradermal injection of both human and mouse tumor cells, more pronounced when 5% cream was applied on three consecutive days. Antibodies against murine IFNgamma, TNFalpha and IL-18 completely abolished the inhibitory effect of imiquimod on angiogenesis induced by murine L1 sacroma cells. When human Skv cells were used in angiogenesis assay, the effect of imiquimod was abolished by antibodies against IL-18 but not against TNFalpha, which may be due to overproduction of TNFalpha by Skv cells. CONCLUSIONS: Antiangiogenic effect of imiquimod was found to be mediated by IL-18, probably through promoting production of INFgamma, the most important inhibitor of angiogenesis. PMID- 15663654 TI - Oral bexarotene in a therapy-resistant Sezary syndrome patient: observations on Sezary cell compartmentalization. AB - BACKGROUND: A 63-year-old man with therapy-resistant Sezary syndrome was enrolled in a multicenter trial of oral bexarotene for advanced-stage cutaneous T-cell lymphoma (CTCL). METHODS: Monthly evaluations for efficacy and side-effects were conducted and documented. RESULTS: Gradual improvement in erythema, pruritus, and scale was noted during the initial 16-week trial period and treatment was extended to 40 weeks. From week 20 to week 40, the erythroderma continued to improve and the lymph node burden decreased, but the absolute Sezary cell count inversely increased. By week 40, intractable pruritus and erythroderma abruptly recurred, and bexarotene was discontinued. CONCLUSIONS: Bexarotene is well tolerated and can be efficacious in patients with Sezary syndrome. Shifting of Sezary cells between different compartments was noted. Further studies on the interaction between the skin, lymph nodes, and peripheral blood compartments during bexarotene treatment in this subset of patients with CTCL are needed. PMID- 15663655 TI - Skin diseases in south-east Nigeria: a current perspective. AB - BACKGROUND AND OBJECTIVE: The incidence of skin diseases in south-east Nigeria during the present decade was analyzed and compared with results from other parts of Nigeria, particularly those in the same zone, obtained more than 30 years ago. This study was carried out to update the recent clinical picture of skin diseases in our environment in view of the rapid development, urbanization and advances in the region. PATIENTS AND METHODS: Consecutive new patients seen at the dermatology clinic of the University of Nigeria, Teaching Hospital, Enugu, Nigeria, from February 1999 to March 2001 were included in this prospective study. Only cases seen and examined by the author were included in this study to ensure uniformity of diagnosis. RESULTS: A total of 2871 patients were observed within the study period. Adults accounted for 69.7% and were aged between 18 and 73 years, while the male:female ratio was 1.3:1. Allergic skin diseases (24.9%) were the commonest skin disorders identified, as opposed to infestations which accounted for an extremely high result of 33.7% (for the same region between 1968 and 1971). In second place was infections/infestations (19.1%). A reversal of picture was thus observed. Within the allergic disorders; eczemas/dermatitis were found to be the most prevalent followed by follicular (13.7%) and pigmentary disorders (11.1%). Sexually transmitted diseases and HIV/AIDs have increased significantly and accounted for 5.4%. Blistering diseases (1.1%) and malignancies (0.5%) occurred less frequently, similar to results found in recent decades for the same region. CONCLUSION: The current picture of skin diseases in south-east Nigeria has changed significantly from mere infections to allergic skin, follicular and pigmentary disorders. Cutaneous lesions secondary to STDs and HIV/AIDs have also increased. Skin lesions related to malnutrition, kwashiorkor and starvation were not observed nor were cutaneous tuberculosis, yaws or pediculosis, while blistering disorders and malignancies remained almost the same. The current picture is similar to that in other developing and Afro Caribbean countries. Primary-care physicians and health-care providers in Nigeria/Africa need to be aware of the globally changing pattern of skin diseases in the region to enable the allocation of necessary resources (financial, material and human) to manage these skin diseases. PMID- 15663656 TI - The value of Doppler ultrasound in diagnosis in 25 cases of furunculoid myiasis. AB - BACKGROUND: The larvae of the botfly Dermatobia hominis cause furunculoid myiasis in endemic areas. Lack of knowledge of this condition outside these areas leads to confusion in diagnosis and delays appropriate treatment. OBJECTIVE: To describe the clinical findings of furunculoid myiasis encountered in Quintana Roo, in the south-east of Mexico. MATERIALS AND METHODS: We performed an observational study on 25 cases diagnosed over a period of 4 years. RESULTS: Diagnosis of furunculoid myiasis was made in 14 males and 11 females with an average age of 24.5 years. Most of our patients were students, farmers, or housewives. The number of lesions varied from one to four. In 20% of cases, more than one parasite was present in each lesion. The scalp was the most commonly affected region (40%), followed by the trunk and the extremities. Doppler ultrasound study (DUSG) of the furuncle-like lesions confirmed the clinical diagnosis in all cases. CONCLUSIONS: Furunculoid myiasis is frequent in the state of Quintana Roo, Mexico. We found no association with occupation, gender, social background or age. DUSG can be used to evaluate the number of parasites per furunculoid lesion avoiding misdiagnoses and treatment delays. PMID- 15663657 TI - Cutaneous leishmaniasis: an atypical case. PMID- 15663658 TI - Pierre Marie-Bamberger syndrome (secondary hypertrophic osteoarthropathy). PMID- 15663659 TI - Persistent Mongolian spots in Chinese adults. AB - Four Chinese females aged 21-35 years with Mongolian spots are reported. Two patients had the spots on their arms while the other two had the spots on their shoulders. The persistence of Mongolian spots in Chinese adults has not been previously reported. PMID- 15663660 TI - Acanthosis nigricans in a plaque of scleredema on the back of a diabetic patient: a case report. AB - Acanthosis nigricans (AN) and scleredema are two skin conditions that have been reported in association with diabetes mellitus. Few cases associating scleredema and AN have been reported. A literature search did not reveal any reports of diabetic patients developing AN on top of a scleredema plaque. Here we report a patient with diabetes mellitus who developed AN within the same indurated scleredema plaque. PMID- 15663661 TI - Metastasis of lung cancer to the finger: a report of two cases. PMID- 15663663 TI - Basal cell carcinoma of the scrotum with lymph node metastasis: report of a case and review of the literature. PMID- 15663662 TI - Hair removal using an 800-nm diode laser: comparison at different treatment intervals of 45, 60, and 90 days. AB - BACKGROUND: Some laser irradiation parameters such as wavelength, fluence, pulse duration, and spot size have been shown to influence the damage of any target inside the skin, however, the role of some patients' factors such as hair growth cycle is still under debate. OBJECTIVE: To determine the association of treatment interval and laser treatment outcome. METHODS: In a retrospective chart review of 176 patients undergoing laser-assisted hair removal with a diode laser 24 patients were selected. All accepted to cease the therapy, and be followed-up for 5 months. At the end of the study the patients were questioned about the efficacy of the treatment as well as the adverse effects. Hair counting was also performed. The adverse effects (pain, blister or erosion, hyperpigmentation, hypopigmentation, and folliculitis) were questioned during the follow-up period. RESULTS: The mean hair reduction was 78.1%, 45.8%, and 28.7% in 45, 60, and 90 day interval groups, respectively (P < 0.0001). CONCLUSION: The treatment interval was related to the treatment outcome in our study. PMID- 15663664 TI - Narrow-band UVB for the treatment of vitiligo: an emerging effective and well tolerated therapy. AB - BACKGROUND: Vitiligo is an acquired depigmentation disorder of great cosmetic importance, affecting 1% of the general population. Photochemotherapy is the most commonly used treatment modality in extensive vitiligo, but is associated with many short- and long-term side-effects. Recently, narrow-band ultraviolet B (NBUVB) therapy has been reported to be an effective and safe therapeutic option in patients with vitiligo. We studied the efficacy and safety of NBUVB (311 nm) therapy in Indian patients with generalized vitiligo. METHODS: Fourteen patients (six males and eight females), aged 12-56 years, with generalized vitiligo, were treated thrice weekly with NBUVB radiation therapy for a maximum period of 1 year. RESULTS: At the end of 1 year, 10 patients (71.4%) had marked to complete repigmentation and two each (14.3%) had moderate or mild repigmentation. Repigmentation sites showed an excellent color match. The response to therapy was correlated with the sites of involvement, duration of disease, and compliance to therapy. Adverse events were limited and transient. CONCLUSION: NBUVB therapy is effective and safe in Indian patients with vitiligo. Long-term follow up is required, however, to establish the stability of repigmentation. PMID- 15663665 TI - Thalidomide for dermatology: a review of clinical uses and adverse effects. PMID- 15663666 TI - Scrotal ulceration induced by all-trans retinoic acid in a patient with acute promyelocytic leukemia. AB - All-trans retinoic acid (ATRA) has been shown to improve the outcome in patients with acute promyelocytic leukemia compared with chemotherapy alone, but it is associated with adverse effects. We report the development of scrotal ulcer in a patient with acute promyleocytic leukemia (APL) within 10 days of treatment with ATRA at a dose of 40 mg orally twice daily. The ulcer did not respond to antibiotic treatment and healed shortly after withholding ATRA. The biopsy showed inflammation only, and other microbiological workup was negative. PMID- 15663668 TI - Nipped in the bud: effects of politics and socioeconomics in the development of Venezuelan and Latin American dermatology. PMID- 15663667 TI - Use of pimecrolimus cream 1% (Elidel) in the treatment of atopic dermatitis in infants and children: the effects of ethnic origin and baseline disease severity on treatment outcome. AB - BACKGROUND: Pimecrolimus cream 1%, a cell-selective inhibitor of inflammatory cytokines, has been shown to be effective in treating atopic dermatitis (AD). This report examines the effect of ethnic origin and baseline disease severity on treatment outcomes in pediatric patients with AD treated with pimecrolimus cream 1%. METHODS: The analysis included 589 patients aged 3 months to 17 years from three 6-week, randomized, multicenter studies of similar design. Patients were treated with pimecrolimus cream 1% or vehicle twice daily. Efficacy, safety and tolerability in Caucasian and non-Caucasian groups were compared. In addition, the effect of baseline disease severity on treatment outcome was investigated. RESULTS: A total of 321 Caucasian and 268 non-Caucasian patients [Blacks, Asians and others (including Hispanics)] with mild, moderate or severe disease at baseline were included. Baseline characteristics were comparable between the pimecrolimus and vehicle control groups and between Caucasian and non-Caucasian groups. Significantly higher efficacy [measured by Investigators' Global Assessment and Eczema Area and Severity Index (EASI) scores] was achieved in the pimecrolimus-treated group, compared with the vehicle group, irrespective of ethnic origin. Baseline disease severity had no effect on treatment outcome: patients with both mild and moderate AD responded well to pimecrolimus (absolute change from baseline in EASI score -2.60 and -5.48, respectively; both P < 0.001). Pimecrolimus cream 1% was safe and well tolerated in all ethnic groups and at all levels of disease severity. CONCLUSIONS: Ethnic origin and baseline disease severity had no effect on treatment outcome with pimecrolimus cream 1% in patients with AD. PMID- 15663669 TI - Familial mucosal lichen planus in three successive generations. PMID- 15663670 TI - Serum levels of proinflammatory cytokines in psoriasis patients from Saudi Arabia. PMID- 15663671 TI - Mondor's phlebitis of penis following recurrent candidal balanoposthitis. PMID- 15663674 TI - Radio-frequency ablation of renal cell carcinoma in patients who were at significant risk. AB - OBJECTIVE: Although radio-frequency ablation (RFA) has been recently applied as a minimally invasive treatment option for renal cell carcinoma (RCC), indication of this modality remains a critical issue due to the lack of complete tumor destruction as well as the uncertainty of its long-term efficacy. We report the efficacy of RFA for nine carefully selected patients with RCC who had significant reason to avoid invasive surgical treatment under general anesthesia. METHODS: Radio-frequency ablation was performed under epidural or local anesthesia by ultrasound or computed tomography (CT) guidance in nine patients with biopsy proven RCC (mean diameter, 38 mm; range, 20-53 mm), who were at significant operative or anesthetic risk for invasive surgery. Follow-up enhanced CT scans or magnetic resonance images were evaluated every 3-6 months and an evaluation of metastasis was performed every 6 months. RESULTS: At a mean follow-up of 17 months, seven (78%) of the nine patients with renal tumor showed no tumor enhancement. The renal function of all patients was well preserved. All patients were able to continue undergoing their respective treatments for active diseases in other organs in parallel to the RFA treatment. No distant metastasis, urine leakage were reported and one case of temporary hematuria and one case of peri renal hemorrhage not requiring blood transfusion were encountered. Intra operative ultrasonography was useful in the real-time monitoring of the minimally excessive extension of ablation into the normal parenchyma. CONCLUSION: Radio frequency ablation appears to be an effective and safe minimally invasive therapeutic option for selected patients with RCC who have reason to avoid invasive surgery under general anesthesia. PMID- 15663675 TI - Primary ureteroscopic treatment for obstructive ureteral stone-causing fornix rupture. AB - BACKGROUND: Management of fornix rupture (FR) by obstructive stone is comprised of extravasation control and the elimination of the obstruction. For all patients, management initially remains conservative under close follow up. Endoscopic management of FR involved with an obstructive stone of the ureter or the pelvi-ureteric junction (UPJ) consists mainly of stenting the ureter. Our endoscopic approach to this pathological entity comprises of the sole stenting of the ureter, as well as primary ureteroscopic lithotripsy followed by ureter stenting. PATIENTS AND METHODS: In the Department of Urology at the General Hospital of Rhodos Island, Rhodos, Greece, over the last 15 years, 51 of 86 patients with FR due to an obstructive stone, were treated endoscopically. Twenty two patients underwent sole stenting of the ureter (option A) and 29 patients underwent primary ureteroscopic lithotripsy and stenting (option B). RESULTS: The overall primary 'successful outcome' was achieved in nine of the 22 patients (40.9%) in the group treated with sole stenting, while the remaining 59.1% required secondary interventions. However, 27 of the 29 patients (93.1%) treated with primary ureteroscopic lithotripsy and stenting required no auxiliary treatment. The primary successful outcome results for obstructive middle and lower ureteral stones with FR were eight out of 12 (66.6%) and 26 out of 27 (96.3%) for therapeutic options A and B, respectively. Upper obstructive ureteral stones with FR required secondary intervention in most cases, regardless of the therapeutic option chosen. (In nine out of 10 and one out of two cases for options A and B, respectively). The mean duration of hospitalization for options A and B were 7.6 and 5.3 days, respectively. The mean duration that the ureter stent remained in situ for A and B treatment options was 30.9 and 10.2 days, respectively. CONCLUSIONS: Sole stenting of the ureter is reserved for infected FR or for stones of the upper ureter or the UPJ. Ureteroscopic lithotripsy followed by double-J stenting of the ureter may offer a quick and safe therapeutic alternative for distal and middle obstructive ureteral stones with FR. PMID- 15663676 TI - Effect of ureteral reimplantation on prevention of urinary tract infection and renal growth in infants with primary vesicoureteral reflux. AB - AIM: We retrospectively reviewed the results of ureteral reimplantation in infants with primary vesicoureteral reflux (VUR) to evaluate the effect on prevention of urinary tract infection (UTI) and renal growth. MATERIALS AND METHODS: From July 1991 to December 2001, a total of 205 infants (180 boys and 25 girls) with primary VUR underwent ureteral reimplantation at the Department of Urology, Osaka Medical Center and Research Institute for Maternal and Child Health, Osaka, Japan. Indications for surgery were high-grade reflux (grade IV V), breakthrough UTI and non-compliance of medical treatment. Age at surgery raged from 1 to 11 months (mean, 6.4 months). Ureteral reimplantation was performed according to Cohen's method. Only two of 336 refluxing ureters required ureteral tailoring. Follow-up ranged from 12 to 110 months (mean, 64 months). Surgical outcome, frequency of UTI and individual renal growth measured by (99m)Tc-dimercaptosuccinic acid (DMSA) scintigraphy was evaluated. RESULTS: Postoperative ultrasound and voiding cystourethrography showed neither residual reflux nor ureterovesical obstruction. Contralateral low grade reflux occurred in six of 74 patients (8.1%) who had unilateral reflux preoperatively. After reimplantation, 10 patients documented 13 febrile UTI. Eleven of the 13 episodes occurred early in the postoperative period (<6 months). Frequency of febrile UTI reduced from 0.23538 before surgery to 0.00894 and 0.00081 per patient per month at 6 and 12 months after surgery, respectively. No development of renal scarring was seen in postoperative DMSA scan. Changes of differential renal function was <0.05 in all patients. CONCLUSION: The present results show ureteral reimplantation in infants is safe and very effective for the prevention of UTI. After surgical treatment in infancy, individual renal growth of children with primary VUR is stable. PMID- 15663677 TI - Association of positive serum anti-p53 antibodies with poor prognosis in bladder cancer patients. AB - AIMS: To assess the association of serum anti-p53 antibodies and overexpression of tumor p53 protein with survival and prognostic factors in patients with urinary bladder tumors. METHODS: Seventy-six patients with transitional cell carcinoma of the urinary bladder were assessed prospectively (Ta, 18; T(1), 30; > or =T(2), 28). Serum anti-p53 antibodies were detected by enzyme-linked immunosorbent assay. Tumor p53 gene overexpression was assessed by immunohistochemical staining. The mean follow-up time was 34 months. RESULTS: Serum anti-p53 antibodies were positive in 25 patients (33%). Overexpression of tumor p53 protein was positive in 41 patients (54%). There was an association between the presence of serum anti-p53 antibodies and tumor p53 gene overexpression (P = 0.001). The total survival of the patients with positive serum anti-p53 antibodies was shorter than the patients with positive tumor p53 gene overexpression (P < 0.001, P = 0.344, respectively). In the multivariate survival analysis, both tumor stage and serum-p53 antibodies were found to be independent survival predictors (P = 0.004, P = 0.006, respectively). CONCLUSION: Serum anti-p53 antibody positive tumors had a worse prognosis than those with negative serum levels, regardless of the p53 status of the tumor. PMID- 15663678 TI - Unreliable residual volume measurement after increased water load diuresis. AB - BACKGROUND: The aim of the present study was to examine whether increased water load intake prior to ultrasound examination of the lower urinary tract, affects the measurement of postvoid residual urine volume. PATIENTS AND METHODS: Ninety four male patients with postvoid residual volumes after increased water load diuresis of more than 100 mL (PVR1) were included in the present study. All patients underwent a second PVR measurement by an urologist with a portable transabdominal bladder ultrasound scan without having received an increased water load (PVR2). A comparison of the measurements was performed and the PVR values were also correlated with other parameters, such as age, International Prostate Symptom Score, prostate volume and serum prostate-specific antigen (PSA) values. RESULTS: Postvoid residual urine volume after increased water load diuresis (PVR1) differed from that recorded after normal bladder filling and voiding at first desire (PVR2), with the former being larger than the later in every patient (P < 0.001). The PVR values, PVR1 and PVR2, were independent to patient age, symptom score prostate volume and PSA value. CONCLUSIONS: Measurement of postvoid residual volume in a relaxed patient, who voids at first desire, represents everyday life and should be the correct method of testing. PMID- 15663679 TI - Telomerase activity is correlated with lower grade and lower stage bladder carcinomas. AB - BACKGROUND: Telomerase is a ribonucleoprotein enzyme that compensates for the progressive erosion of telomeres. The increasing interest in telomerase is motivated by the demonstration that most human carcinomas are telomerase positive. The potential use of telomerase activity in bladder carcinomas using a urine sample has been reported in several studies. However, little is known about the detection of telomerase activity in bladder carcinoma tissues. Herein, we investigate telomerase activity in bladder carcinoma tissues according to grade (G) and stage. MATERIAL AND METHODS: Telomerase activity was assayed by polymerase chain reaction enzyme-linked immunosorbent assay methods. Malignant lesions were assessed in 37 patients with bladder carcinoma and no malignant lesions were assessed in five patients with dysplasia or inflammatory bladder lesions. RESULTS: Twenty-three out of 37 carcinoma samples were telomerase positive and one out of five control samples without carcinoma was telomerase positive. The positive rates according to stage and grade were 83.3% for superficial and 42.1% for invasive stages and 83.3% for G1, 66.7% for G2 and 40.0% for G3. Telomerase activity was correlated with lower grade and lower stage bladder carcinomas. CONCLUSION: These results strongly suggest that reactivation of telomerase may differ between superficial and invasive bladder carcinomas and also between low grade and high grade bladder carcinomas. PMID- 15663680 TI - Three-dimensional ultrasonography in evaluation of benign prostatic hyperplasia. AB - BACKGROUND: The clinical application of volume estimation by 3-D ultrasound has recently gained much attention. However, there have been no reports evaluating the prostate volume by 3-D transrectal ultrasound (TRUS) before transurethral resection of the prostate (TURP). The purpose of the present study was to evaluate the value of 3-D TRUS for prediction of prostate morphology and resected weight before TURP and to investigate whether 3-D TRUS is a more useful examination than 2D TRUS in patients with benign prostatic hyperplasia (BPH). METHODS: Transurethral resection of the prostate was performed in 23 patients with BPH. We evaluated the prostate morphology and measured both the volumes of the whole prostate and the transition zone using 2-D and 3-D TRUS, respectively. The actual resected weight was recorded and compared with the volume of the whole prostate and that of the transition zone measured by 2D and 3-D TRUS. RESULTS: The volume of the transition zone measured by 3-D TRUS correlated most strongly with the resected weight (r = 0.84). A large median lobe was seen in three patients in whom the transition zone volume measured by 2-D TRUS was considerably larger than the resected weight. However, overestimation in the three patients decreased by the use of 3-D TRUS. CONCLUSIONS: It was concluded that 3-D TRUS was equal or superior to 2-D TRUS in prediction of the resected weight and 3-D TRUS offers better information as a diagnostic tool before TURP. PMID- 15663681 TI - Combined androgen blockade in the management of advanced prostate cancer: a sensible or ostensible approach. AB - BACKGROUND: To compare the efficacy of orchiectomy alone and orchiectomy plus flutamide in treating patients with advanced carcinoma prostate. MATERIALS AND METHODS: The study was initiated on 1 July 1997 and closed after enrolling 100 patients on 30 June 2000. Patients were prospectively randomized to orchiectomy alone (O) and orchiectomy plus flutamide (OF). A complete response (CR) was defined as the normalization of bone scans and serum prostate-specific antigen (PSA) levels returning to normal (< 4 ng/mL). A partial response (PR) was defined as a 50% reduction in metastasis mass compared to the initial study or a decrease in the PSA level of 50% of the initial value. Progressive disease (PD) was defined as the development of any new hot spot on bone scan or any increase in previously existing PSA level by 25%. RESULTS: A total of 100 patients were entered in the study. The maximum percentage change in PSA levels in both groups was found in the first 3 months after orchiectomy, that is, 95% and 97% for the O and OF groups, respectively. In more than 80% of the patients this decrease in PSA was maintained for 3 years. The mean percentage change at 3 years in the O and OF groups was 70% and 75% (P = 0.95), respectively, and the overall response rate (CR + PR) was 88.50% and 86.53% in the two groups, respectively (P = 0.85). The follow-up period ranged between 3 and 5 years (mean, 3.5 years). The mean time to progression was 27 and 29 months in the O and OF groups, respectively. The overall survival rate at 3 and 5 years in two treatment groups was 45.83% and 48.07%, 20.83% and 23.07% in the O and OF groups, respectively (P = 0.75). CONCLUSIONS: Maximum percentage decrease in PSA is seen within the first 3 months of therapy. Orchiectomy alone is as effective as combination therapy in decreasing serum PSA. Overall survival at 3 and 5 years in the orchiectomy only group was as good as that of combination therapy. These data suggest that the routine addition of flutamide to orchiectomy is not advisable. PMID- 15663682 TI - Transurethral resection in women with lower urinary tract symptoms. AB - BACKGROUND: To evaluate the preliminary safety, efficacy and complications of transurethral resection (TUR) in the management of women with lower urinary tract symptoms. METHODS: One hundred and eight women (mean age, 57.6 years) with moderate to severe lower urinary tract symptoms underwent TUR between September 1998 and December 2002. At preoperative baseline, 6 months and 24 months postoperatively, all patients underwent clinical evaluations including the standardized American Urological Association symptom score, peak urine flow, postvoid residual urine volume and quality-of-life assessments. Operative time, catheter time, hospital stay and incidence of side-effects were also recorded. RESULTS: The total mean operative time was 19.25 +/- 6.70 min. The mean catheter time was 26.2 +/- 12.8 h and the mean hospital stay was 2.08 +/- 0.84 days. No patient needed a blood transfusion or experienced hyponatremia. The total storage and voiding symptom scores and quality of life score had improved significantly 6 months after the surgical procedure (P < 0.01). There were little changes in peak flow rate and residual urine volume. Complications of the procedure included urinary tract infections (n = 4) and stress incontinence (n = 3). Two patients later underwent bladder neck suspension procedure. CONCLUSIONS: In this preliminary study, there was significant clinical improvement maintained at all follow-up intervals and the results were not comparable to TUR of the prostate in men with clinical benign prostatic hyperplasia. The risks associated with TUR in women, especially of stress incontinence, are significant. A multicenter clinical trial is currently underway to determine the long-term efficacy and safety of TUR in women. PMID- 15663684 TI - Congenital absence of the testis in human fetuses and in cryptorchid patients. AB - BACKGROUND: The aim of the present study is to make a comparative study in human fetuses and in patients with cryptorchidism, analyzing the incidence of a number anomalies of the testes for both populations. METHODS: We studied 326 testes from 163 human fetuses ranging in age from 10 to 35 weeks postconception (WPC) and 133 testes from 101 cryptorchid patients aged from 1 to 15 years old (mean, 6.4 years). The Fisher's exact test was used for comparison. RESULTS: Among 326 fetal testes, 224 (68.7%) were abdominal, 45 (13.8%) were inguinal and 55 (16.8%) were scrotal. In one fetus at 23 WPC, both testes (0.6%) were absent. Of the 133 cryptorchid testes, 17 (12.78%) were abdominal, 92 (69.1%) were inguinal and 24 (18%) were high scrotal. Of the 17 abdominal testes, three (17.6%) were atrophic and two were vanished (11.7%). Of the 92 inguinal testes, one (1.08%) was vanished. Twenty-eight (21%) of the cryptorchid testes were impalpable and among these, 17 were located in the abdomen (60.7%) and 11 (38.2%) in the inguinal region (internal ring). CONCLUSIONS: Testicular agenesis is a very rare anomaly, both in fetuses and patients with cryptorchidism. PMID- 15663683 TI - Clinical application of prognostic factors for patients with organic causes of erectile dysfunction on 100 mg of sildenafil citrate. AB - OBJECTIVES: There are still no clinical predictive factors to determine the response rate of erectile dysfunction (ED) patients to sildenefil citrate. The aim of the present study is to evaluate and stratify the risk factors and attempted to determine the prognostic factors in clinical practice to predict the response rate. This is important in improving cost effectiveness and avoiding side-effects. MATERIAL AND METHODS: This is an open label prospective study including patients attending the andrology clinic in Tan Tock Seng Hospital, Singapore, over 2 years. The patients were evaluated and investigated for possible underlying causes and ED severity was assessed by five-items of the International Index of Erectile Function questionnaire (IIEF-5), together with the duration, degree and rigidity of erection. Psychogenic causes were excluded with a minimal follow up of 6 months. All patients were placed on 100 mg of sildenafil citrate and were reassessed at the end of 6 months. Logistic regression with univariate and multivariate was used as the method of statistical analysis. RESULTS: A total of 232 patients were in the cohort. The overall response rate was 43%, with the best response rate in veno-occlusive cases and the worst responses from neurogenic causes. Age, smoking, diabetes mellitus, hypertension, hyperlipidemia, pretreatment IIEF-5 score, interval to achieve erection and duration of erection were significant in univariate analysis, but only age, smoking and IIEF-5 score were significant in multivariate analysis. With a combination of these factors, a table was formed to determine the possible response rate in clinical practice. This will assist physicians in selecting patients with potentially favorable responses and avoid side-effects and an unnecessary wastage of time and cost. CONCLUSION: Possible factors could be determined and used clinically to predict the response rate to sildenafil citrate. PMID- 15663685 TI - Comparison of two in vivo models for prostate cancer: orthotopic and intratesticular inoculation of LNCaP or PC-3 cells. AB - BACKGROUND: The critical events in the clinical course of prostate cancer are the occurrence of metastasis and the induction of the hormone-refractory status of the disease. In order to investigate the factors responsible for these events, we need appropriate in vivo models. MATERIALS AND METHODS: Orthotopic and intratesticular models were created by the injection of LNCaP cells or PC-3 cells into the prostate or testis of severe combined immunodeficient mice. RESULTS: LNCaP cells in the intratesticular model showed a higher incidence of tumor formation and lymph node metastasis when compared with those in the orthotopic model, while PC-3 cells were highly tumorigenic and metastastic in both models. A high concentration of androgens might play a role in tumor aggressiveness of LNCaP cells, given that enhanced mRNA expressions of integrin alphaV and vascular endothelial growth factor was induced by dehydrotestosterone administration in vitro. The high expression of metastasis-related genes, including the urokinase plasminogen activator system, metalloproteinases and vascular endothelial growth factor-C, might be attributed to the high metastatic potential in both models. Interestingly, testicular xenografts of LNCaP cells were able to survive on the subcutis back of castrated male mice as well female mice. CONCLUSIONS: Intratesticular models of prostate cancer appear to be suitable for studying the mechanisms of metastasis and for evaluating various treatment strategies. PMID- 15663686 TI - Effect of intraurethral Mitomycin-C on healing and fibrosis in rats with experimentally induced urethral stricture. AB - BACKGROUND: In the pathogenesis of urethral stricture, fibrosis is associated with an excessive collagen increase. After the recognition that topical application of Mitomycin-C (MMC) inhibits fibroblast proliferation and is effective in preventing scar formation, many studies have been carried out on this subject outside the scope of urology. The aim of the present study is to observe the intraurethral impact of the employment of low doses of MMC on scar formation and fibrosis in experimental rat model. METHODS: Urethral injuries were made by internal urethrotomy knife. The study was carried out with 35 adult male Wistar albino rats. Five rats were allocated to the control group (group 1), 10 to a group that was administered 2 mg/L MMC (group 2) and 20 to a group that was administered 10 mg/L MMC (group 3). Mitomycin-C was administered to the injured urethra in the form of irrigation for 5 min. The rats were sacrificed 14 days later in order to evaluate chronic inflammation and fibrosis and their penises were histopathologically examined under light microscopy with hematoxilen eosin and trichrom stains. RESULTS: When group 2 was compared with control group, the differences in hemosiderin-laden macrophages (HLM), mononuclear cell infiltration (MCI) and fibrosis were found to be statistically significant (P < 0.01, P < 0.05, P < 0.005, respectively). When group 3 was compared with control group, the differences in HLM, MCI and fibrosis were also found to be statistically significant (P < 0.05, P < 0.05, P < 0.005, respectively). In the comparison of group 2 with group 3, no statistically significant differences were found in terms of the these parameters. CONCLUSIONS: Although MMC is toxic at high doses, the antifibrotic effect of the intraurethral low dose MMC may be useful in combination therapy for internal urethrotomy. PMID- 15663687 TI - Long-term survival after bilateral adrenalectomy for metachronous adrenocortical cancer. AB - We report the case of a female patient with bilateral metachronous adrenocortical cancer who survived long-term after adrenalectomy. In 1991, the patient underwent left adrenalectomy to remove a huge adrenal mass (10 x 9 cm) displaying no hormonal abnormality. Histological diagnosis was adrenocortical cancer. A right adrenal mass (7 x 6 cm) was found 4 years after left adrenalectomy. Right adrenalectomy was performed, and histological diagnosis was again adrenocortical cancer. The patient remains alive with no evidence of disease 8 years after last surgery. PMID- 15663688 TI - Enhancement of T cell proliferative response against autologous cancer cells of a metasatic renal cell carcinoma patient after unexplained regression. AB - The unexplained regression of a metastasis of renal cell carcinoma (RCC) is rare phenomenon. While an immune mechanism has been proposed for spontaneous or unexplained regression of RCC, only limited data are available regarding immunological response. We report a case of a 62-year-old man with RCC whose paravertebral pleural tumor regressed after the withdrawal of interferon-alpha. In the present case, we demonstrate the enhancement of T cell proliferative response against autologous RCC cells secondary to the unexplained regression using the mixed-lymphocyte tumor culture. To our knowledge, the enhancement of an antitumor immunity secondary to an unexplained regression of RCC has not previously been reported. PMID- 15663689 TI - Multicentric liposarcoma. AB - Abstract A 72-year-old man complaining of upper abdominal discomfort was diagnosed as having retroperitoneal liposarcoma by means of diagnostic imaging. He then underwent an operation. One mass existed on the curvatura ventriculi major, extending to the hilum splenicum and pressing back the pancreal head and body. There was another mass to the left of the first, situated on the ventral side of the left kidney. Also, another mass was intramurally found adjacent to the curvatura ventriculi major. Histologically, the mass on the curvatura ventriculi major ranged from the peritoneal cavity to the retroperitoneum Its intraperitoneal portion was classified as a differentiated lipoma-like type and the retroperitoneal mass was of mucous type. The mass on the left kidney was of a differentiated fibrosing type. The intramural mass in the gastric curvature was found to be a differentiated lipoma-like type. The patient has been under observation for 12 months and has shown no recurrence. PMID- 15663690 TI - Carcinosarcoma of the bladder diverticulum and a review of the literature. AB - We report a carcinosarcoma within a bladder diverticulum together with a review of 18 previously reported cases of intradiverticular sarcomas and carcinosarcomas with respect to their clinical features, treatments and outcomes. Frequent deaths with intra-abdominal recurrences suggest the need for total cystectomy, rather than diverticulectomy. PMID- 15663691 TI - Synchronous anal and penile cancers. AB - Malignancies of the anogenital region have similar etiology, including viral infections, sexual habits, social and educational factors. Condylomata acuminata represent precancerous lesions. In the available literature, we found no cases of synchronous cancers of anus and penis reported. We report a case of synchronous cancer of the penile foreskin and cancer of the anus in a 36 year-old man. The microscopic diagnosis of the anal lesion was squamous cell cancer, basaloid type. The penile lesion was squamous cell cancer, keratinizing type. Both cancers presented with the in situ component. Immunohistochemical staining (MIB-1) and histological features (koilocytosis, acanthosis and dyskeratosis) indirectly confirmed the past viral infection. Numerous etiological similarities in anogenital cancers entail careful clinical inspection in patients with a malignant lesion in this region and suggest searching for possible other concomitant cancers or precancerous lesions. PMID- 15663692 TI - Testis-sparing surgery in an adult with bilateral synchronous seminomatous tumor. AB - Bilateral synchronous testicular cancer is a rare occurrence and is usually associated with similar histological findings in each testicle. The standard therapy of bilateral testis cancer is generally considered to be orchiectomy. We present a case of synchronous bilateral testicular germ cell tumor treated with testis-sparing surgery. The patient was disease free for 30 months after surgery without local recurrence or distant metastases. Testis-sparing surgery provides a better quality of life and may be considered a safe, feasible alternative in the treatment of carefully selected patients with bilateral testicular germ cell tumor. PMID- 15663693 TI - Spontaneous rupture of varicocele testis associated with advanced pancreatic cancer. AB - We report a rare case of strain-induced spontaneous rupture of varicocele associated with renal vein involvement by advanced pancreatic cancer. Computed tomography and color Doppler sonography yielded the correct diagnosis and the patient could maintain quality of life without surgery for acute scrotum. PMID- 15663694 TI - Paratesticular mucinous cystadenocarcinoma: metastasis from pancreatic cancer. AB - We experienced a case of a paratesticular mucinous adenocarcinoma from primary pancreatic cancer. A 67-year old man presented with a scrotal mass. Scrotal ultrasound showed a cystic mass on the testis. Radical orchiectomy was performed and the tumor was revealed as a mucinous cystadenocarcinoma separated from epididymis and testis. Metastatic cancer was suspected and abdominal computed tomography showed pancreatic cancer. We report this rare case of metastatic paratesticular cystoadenocarcinoma. PMID- 15663695 TI - Median raphe cyst in the scrotum, mimicking a serous borderline tumor, associated with cryptorchidism after orchiopecxy. AB - Median raphe cyst (MRC) is a benign lesion occurring predominantly in the ventral surface of the penises of young men and is an embryological developmental anomaly of the male genitalia. Serous borderline tumors (SBT) are found most frequently in the female ovary and only several cases with SBT of the male genitalia have been reported. We describe a case of MRC with features of SBT, which appeared in the scrotum of a 9-year-old boy after orchiopexy and was associated with surgery for cryptorchidism. The cyst arose on the right testicular tunica and consisted of cystic components with intracystic papillae lined by stratified epithelial cells, some of which showed mild cytological atypia and sporadic mitosis. These epithelial cells expressed CA 125, CA 19-9, carcinoembryonic antigen, estrogen receptor and progesterone receptor. Although no cases of MRC with characteristics of SBT in association with the rete testis has been described, the current report gives additional information for follow-up of cryptorchidism. PMID- 15663696 TI - Low-grade renal epithelial tumor originating from the distal nephron. PMID- 15663698 TI - How does race matter, anyway? PMID- 15663699 TI - Reverse translation in health policy and management: from bedside to bench and beyond. PMID- 15663700 TI - The impact of Medicaid managed care on hospitalizations for ambulatory care sensitive conditions. AB - OBJECTIVE: To determine whether Medicaid managed care is associated with lower hospitalization rates for ambulatory care sensitive conditions than Medicaid fee for-service. We also explored whether there was a differential effect of Medicaid managed care by patient's race or ethnicity on the hospitalization rates for ambulatory care sensitive conditions. DATA SOURCES/STUDY SETTING: Electronic hospital discharge abstracts for all California temporary assistance to needy families (TANF)-eligible Medicaid beneficiaries less than age 65 who were admitted to acute care hospitals in California between 1994 and 1999. STUDY DESIGN: We performed a cross-sectional comparison of average monthly rates of admission for ambulatory care-sensitive conditions among TANF-eligible Medicaid beneficiaries in fee-for-service, voluntary managed care, and mandatory managed care. DATA COLLECTION/EXTRACTION METHODS: We calculated monthly rates of ambulatory care-sensitive condition admission rates by counting admissions for specified conditions in hospital discharge files and dividing the monthly count of admissions by the size of the at-risk population derived from a separate monthly Medicaid eligibility file. We used multivariate Poisson regression to model monthly hospital admission rates for ambulatory care-sensitive conditions as a function of the Medicaid delivery model controlling for admission month, admission year, patient age, sex, race/ethnicity, and county of residence. PRINCIPAL FINDINGS: The adjusted average monthly hospitalization rate for ambulatory care-sensitive conditions per 10,000 was 9.36 in fee-for-service, 6.40 in mandatory managed care, and 5.25 in voluntary managed care (p<.0001 for all pairwise comparisons). The difference in hospitalization rates for ambulatory care sensitive conditions in Medicaid fee-for-service versus managed care was significantly larger for patients from minority groups than for whites. CONCLUSIONS: Selection bias in voluntary Medicaid managed care programs exaggerates the differences between managed care and fee-for-service, but the 33 percent lower rate of hospitalizations for ambulatory care sensitive conditions found in mandatory managed care compared with fee-for-service suggests that Medicaid managed care is associated with a large reduction in hospital utilization, which likely reflects health benefits. The greater effect of Medicaid managed care for minority compared with white beneficiaries is consistent with other findings that suggest that managed care is associated with improvements in access to ambulatory care for those patients who have traditionally faced the greatest barriers to health care. PMID- 15663701 TI - How well does Medicaid work in improving access to care? AB - OBJECTIVE: To provide an assessment of how well the Medicaid program is working at improving access to and use of health care for low-income mothers. DATA SOURCE/STUDY SETTING: The 1997 and 1999 National Survey of America's Families, with state and county information drawn from the Area Resource File and other sources. STUDY DESIGN: Estimate the effects of Medicaid on access and use relative to private coverage and being uninsured, using instrumental variables methods to control for selection into insurance status. DATA COLLECTION/EXTRACTION METHOD: This study combines data from 1997 and 1999 for mothers in families with incomes below 200 percent of the federal poverty level. PRINCIPAL FINDINGS: We find that Medicaid beneficiaries' access and use are significantly better than those obtained by the uninsured. Analysis that controls for insurance selection shows that the benefits of having Medicaid coverage versus being uninsured are substantially larger than what is estimated when selection is not accounted for. Our results also indicate that Medicaid beneficiaries' access and use are comparable to that of the low-income privately insured. Once insurance selection is controlled for, access and use under Medicaid is not significantly different from access and use under private insurance. Without controls for insurance selection, access and use for Medicaid beneficiaries is found to be significantly worse than for the low-income privately insured. CONCLUSIONS: Our results show that the Medicaid program improved access to care relative to uninsurance for low-income mothers, achieving access and use levels comparable to those of the privately insured. Our results also indicate that prior research, which generally has not controlled for selection into insurance coverage, has likely understated the gains of Medicaid relative to uninsurance and overstated the gains of private coverage relative to Medicaid. PMID- 15663702 TI - From Medicaid to uninsured: drop-out among children in public insurance programs. AB - OBJECTIVES: To estimate a national disenrollment rate among children in Medicaid and the Children's Health Insurance Program (CHIP); to determine what share of disenrollment is due to acquiring other insurance or losing eligibility; and to examine what demographic and policy factors make disenrollment more likely. DATA SOURCES: Insurance status, income, and demographics from the Current Population Survey (CPS) March Supplement (1998-2001); eligibility data from the National Governors Association; and policy data from the former Health Care Financing Administration (HCFA), state welfare offices, and previous research. STUDY DESIGN: The study used a nationally representative sample of 5,551 children in Medicaid or CHIP. The key outcomes were the percentage of children still enrolled 1 year later, and the share of disenrollees who became uninsured despite remaining eligible. Multivariate logistic regression was used to explore demographics and policies predictive of disenrollment. DATA COLLECTION AND ANALYSIS: CPS data were extracted using the Census Bureau's Federal Electronic Research and Review Extraction Tool 1.0. Data analysis was performed using Stata 7 (Stata Corporation 2001). PRINCIPAL FINDINGS: Of the children enrolled in Medicaid or CHIP, 27.7 percent were no longer enrolled 12 months later. Of those, 45.4 percent dropped out despite apparently remaining eligible and having no other insurance--corresponding to 3.0 million children annually. Drop-out varied significantly across states. Children without siblings in public insurance were at a higher risk for drop-out. Children with more educated parents were more likely to leave Medicaid for private insurance or to lose Medicaid eligibility, while black children and infants were less likely to lose their eligibility. Decreased Medicaid provider reimbursement rates were strongly associated with drop-out, while Medicaid managed care increased the exodus to private insurance. CONCLUSIONS: Drop-out from Medicaid and CHIP is a significant policy concern and helps explain the persistence of uninsurance among millions of eligible children. Clinical encounters with providers appear to play a key role in preventing drop out. PMID- 15663703 TI - Incentives in a Medicaid carve-out: impact on children with special health care needs. AB - OBJECTIVE: To evaluate whether a specialty care payment "carve-out" from Medicaid managed care affects caseloads and expenditures for children with chronic conditions. DATA SOURCE: Paid Medicaid claims in California with service dates between 1994 and 1997 that were authorized by the Title V Children with Special Health Needs program for children under age 21. STUDY DESIGN: A natural experiment design evaluated the impact of California's Medicaid managed care expansion during the 1990s, which preserved fee-for-service payment for certain complex medical diagnoses. Outcomes in time series regression include Title V program participation and expenditures. Multiple comparison groups include children in managed care counties who were not mandated to enroll, and children in nonmanaged care counties. DATA COLLECTION/EXTRACTION METHODS: Data on the study population were obtained from the state health department claims files and from administrative files on enrollment and managed care participation. PRINCIPAL FINDINGS: The carve-out policy increased the number of children receiving Title V authorized services. Recipients and expenditures for some ambulatory services increased, although overall expenditures (driven by inpatient services) did not increase significantly. Cost intensity per Title V recipient generally declined. CONCLUSIONS: The carve-out policy increased identification of children with special health care needs. The policy may have improved children's access to prevailing standards of care by motivating health plans and providers to identify and refer children to an important national program. PMID- 15663704 TI - The "minimizing antibiotic resistance in Colorado" project: impact of patient education in improving antibiotic use in private office practices. AB - OBJECTIVE: To assess the marginal impact of patient education on antibiotic prescribing to children with pharyngitis and adults with acute bronchitis in private office practices. DATA SOURCES/STUDY SETTING: Antibiotic prescription rates based on claims data from four managed care organizations in Colorado during baseline (winter 2000) and study (winter 2001) periods. STUDY DESIGN: A nonrandomized controlled trial of a household and office-based patient educational intervention was performed. During both periods, Colorado physicians were mailed antibiotic prescribing profiles and practices guidelines as part of an ongoing quality improvement program. Intervention practices (n=7) were compared with local and distant control practices. DATA COLLECTION/EXTRACTION METHODS: Office visits were extracted by managed care organizations using International Classification of Diseases-9-Clinical Modification codes for acute respiratory tract infections, and merged with pharmacy claims data based on visit and dispensing dates coinciding within 2 days. PRINCIPAL FINDINGS: Adjusted antibiotic prescription rates during baseline and study periods increased from 38 to 39 percent for pediatric pharyngitis at the distant control practices, and decreased from 39 to 37 percent at the local control practices, and from 34 to 30 percent at the intervention practices (p=.18 compared with distant control practices). Adjusted antibiotic prescription rates decreased from 50 to 44 percent for adult bronchitis at the distant control practices, from 55 to 45 percent at the local control practices, and from 60 to 36 percent at the intervention practices (p<.002 and p=.006 compared with distant and local control practices, respectively). CONCLUSIONS: In office practices, there appears to be little room for improvement in antibiotic prescription rates for children with pharyngitis. In contrast, patient education helps reduce antibiotic use for adults with acute bronchitis beyond that achieved by physician-directed efforts. PMID- 15663705 TI - Self-medication of mental health problems: new evidence from a national survey. AB - OBJECTIVE: To evaluate the association between past 30-day use of alcohol, marijuana, and other illicit drugs and past year unmet need for and use of mental health care. DATA SOURCE: A subsample of 18,849 respondents from the 2001 National Household Survey on Drug Abuse and the 2002 National Survey on Drug Use and Health. Subjects were between the ages of 18 and 65 years and had least one past year mental disorder symptom and no past year substance dependency. STUDY DESIGN: Logistic regressions of past 30-day substance use on past 12-month unmet need for mental health care and past 12-month use of mental health services controlling for clinical and sociodemographic characteristics. Predicted probabilities and corresponding standard errors are reported. PRINCIPAL FINDINGS: Use of illicit drugs other than marijuana increased with unmet need for mental health care (4.4 versus 3.2 percent, p=.046) but was not reduced with mental health-care use. Heavy alcohol use was not associated with increased unmet need for mental health care, but was higher among individuals with no mental health care use (4.4 percent versus 2.7 percent, p<.001). By contrast, marijuana use did not appear associated with either unmet need or mental health care use. CONCLUSIONS: Substance use varies with past year unmet need for mental health care and mental health care use in ways consistent with the self-medication hypothesis. Results suggest that timely screening and treatment of mental health problems may prevent the development of substance-use disorders among those with mental disorders. Further research should identify subgroups of individuals for whom timely and appropriate mental health treatment would prevent the development of substance-use disorders. PMID- 15663706 TI - The effects of geography and spatial behavior on health care utilization among the residents of a rural region. AB - OBJECTIVE: This analysis determines the importance of geography and spatial behavior as predisposing and enabling factors in rural health care utilization, controlling for demographic, social, cultural, and health status factors. DATA SOURCES: A survey of 1,059 adults in 12 rural Appalachian North Carolina counties. STUDY DESIGN: This cross-sectional study used a three-stage sampling design stratified by county and ethnicity. Preliminary analysis of health services utilization compared weighted proportions of number of health care visits in the previous 12 months for regular check-up care, chronic care, and acute care across geographic, sociodemographic, cultural, and health variables. Multivariable logistic models identified independent correlates of health services utilization. DATA COLLECTION METHODS: Respondents answered standard survey questions. They located places in which they engaged health related and normal day-to-day activities; these data were entered into a geographic information system for analysis. PRINCIPAL FINDINGS: Several geographic and spatial behavior factors, including having a driver's license, use of provided rides, and distance for regular care, were significantly related to health care utilization for regular check-up and chronic care in the bivariate analysis. In the multivariate model, having a driver's license and distance for regular care remained significant, as did several predisposing (age, gender, ethnicity), enabling (household income), and need (physical and mental health measures, number of conditions). Geographic measures, as predisposing and enabling factors, were related to regular check-up and chronic care, but not to acute care visits. CONCLUSIONS: These results show the importance of geographic and spatial behavior factors in rural health care utilization. They also indicate continuing inequity in rural health care utilization that must be addressed in public policy. PMID- 15663708 TI - Initial home health outcomes under prospective payment. AB - OBJECTIVE: To assess initial changes in home health patient outcomes under Medicare's home health Prospective Payment System (PPS), implemented by the Centers for Medicare and Medicaid Services (CMS) in October 2000. DATA SOURCES/STUDY SETTING: Pre-PPS and early PPS data were obtained from CMS Outcome and Assessment Information Set (OASIS) and Medicare claims files. STUDY DESIGN: Regression analysis was applied to national random samples (n=164,810) to estimate pre-PPS/PPS outcome and visit-per-episode changes. DATA COLLECTION/EXTRACTION METHODS: Outcome episodes were constructed from OASIS data and linked with Medicare claims data on visits. PRINCIPAL FINDINGS: Outcome changes (risk adjusted) were mixed and generally modest. Favorable changes included higher improvement rates under PPS for functioning and dyspnea, higher community discharge rates, and lower hospitalization and emergent care rates. Most stabilization (nonworsening) outcome rates also increased. However, improvement rates were lower under PPS for wounds, incontinence, and cognitive and emotional/behavioral outcomes. Total visits per episode (case-mix adjusted) declined 16.6 percent although therapy visits increased by 8.4 percent. CONCLUSIONS: The outcome and visit results suggest improved system efficiency under PPS (fewer visits, similar outcomes). However, declines in several improvement rates merit ongoing monitoring, as do subsequent (posthome health) hospitalization and emergent care use. Since only the early PPS period was examined, longer-term analyses are needed. PMID- 15663707 TI - The administrative control system of substance abuse managed care. AB - OBJECTIVE: This article searches for the dimensions of the administrative structures in outpatient substance abuse managed care that control the behavior of agency providers. It also ascertains how these dimensions, and several financial mechanisms, affect key aspects of the providers services: the average number of sessions of care that are delivered, the rate of completion of care, and the (estimated) rate at which clients control their substance use. DATA SOURCES: The data were collected in 1999 for this investigation. STUDY DESIGN: These data come from a nationally representative, cross-sectional sample of individual contracts between outpatient drug treatment providers and the Behavioral Health Managed Care Organizations (BHMCOs) that are empowered to regulate the delivery of services. Provider responses are analyzed here. DATA COLLECTION METHODS: Factor analyses at a contract level examine the structural dimensions of the control system. Multivariate analyses at the same level rely on generalized linear models to predict the dependent variables by the structural dimensions and financial mechanisms. FINDINGS: The factor analyses suggest that there are six multiple variable structural dimensions. The multivariate analyses suggest that the dimension that mandates follow-up of discharged clients tends to relate to more sessions of care and perhaps a higher rate of service completion. Most other dimensions are found to relate to fewer sessions of care, lower rates of service completion, or lower rates of control of substance abuse. No structural dimension relates to all dependent variables. Financial mechanisms evince varying relations to the sessions of care. They rarely relate to the other dependent variables. CONCLUSION: The results generally suggest that providers, payers, or policymakers might affect service provision by selecting BHMCOs that stress particular structural dimensions and financial mechanisms. However, managed care contracts most heavily rely on structural dimensions that restrict treatment sessions and fail to predict superior client outcomes. PMID- 15663709 TI - The determinants of private medical insurance prevalence in England, 1997-2000. AB - OBJECTIVE: The aim of this paper is to identify the factors that determine the prevalence of private medical insurance (PMI) in England. DATA SOURCES/STUDY SETTING: Secondary data sources are the British Household Panel Survey (BHPS) 1997-2000, Laing's Healthcare Market Review 1999-2000, the United Kingdom (U.K.) Department of Health's National Health Service Waiting Times Team, and the Work Force Statistics Branch of the Department of Health. STUDY DESIGN: Logistic regression models for panel data were used to compare non-PMI subscribers with individual subscribers and those with employer-provided PMI. DATA COLLECTION/EXTRACTION METHODS: The BHPS data are collected by the Institute for Social and Economic Research at the University of Essex. Other data used were collected by Laing and Buisson and the U.K. Department of Health. PRINCIPAL FINDINGS: Individual PMI is more prevalent among the well-educated and healthy. Income, age, sex, and political preference are key determinants of PMI prevalence for both individual and employer paid PMI. Individuals are also likely to reflect on information with regard to waiting times in deciding whether or not to purchase PMI cover. The withdrawal of the tax subsidy in 1997 to PMI subscribers over 60 years of age did not impact on their rate of withdrawal from PMI coverage relative to the rate among all PMI subscribers, but may have discouraged potential new subscribers. CONCLUSIONS: Current trends in the PMI market suggest that, over time, individually purchased PMI is likely to be partially displaced by PMI purchased as part of a company-based plan. However, having PMI is linked to economic factors in both groups, suggesting a similar segment of the population valuing the responsiveness that PMI provides. Geographic factors relating to waiting times and supply-side factors are associated with both individual and company-based PMI. The withdrawal of the tax subsidy to individual subscribers older than age of 60 resulted in a significant decline in the demand for PMI. In particular, the number of new subscribers in this group declined substantially. PMID- 15663710 TI - Response rates and response bias for 50 surveys of pediatricians. AB - RESEARCH OBJECTIVE: To track response rates across time for surveys of pediatricians, to explore whether response bias is present for these surveys, and to examine whether response bias increases with lower response rates. DATA SOURCE/STUDY SETTING: A total of 63,473 cases were gathered from 50 different surveys of pediatricians conducted by the American Academy of Pediatrics (AAP) since 1994. Thirty-one surveys targeted active U.S. members of the AAP, six targeted pediatric residents, and the remaining 13 targeted AAP-member and nonmember pediatric subspecialists. Information for the full target samples, including nonrespondents, was collected using administrative databases of the AAP and the American Board of Pediatrics. STUDY DESIGN: To assess bias for each survey, age, gender, location, and AAP membership type were compared for respondents and the full target sample. Correlational analyses were conducted to examine whether surveys with lower response rates had increasing levels of response bias. PRINCIPAL FINDINGS: Response rates to the 50 surveys examined declined significantly across survey years (1994-2002). Response rates ranged from 52 to 81 percent with an average of 68 percent. Comparisons between respondents and the full target samples showed the respondent group to be younger, to have more females, and to have less specialty-fellow members. Response bias was not apparent for pediatricians' geographical location. The average response bias, however, was fairly small for all factors: age (0.45 years younger), gender (1.4 percentage points more females), and membership type (1.1 percentage points fewer specialty-fellow members). Gender response bias was found to be inversely associated with survey response rates (r=-0.38). Even for the surveys with the lowest response rates, amount of response bias never exceeded 5 percentage points for gender, 3 years for age, or 3 percent for membership type. CONCLUSIONS: While response biases favoring women, young physicians, and nonspecialty-fellow members were found across the 52-81 percent response rates examined in this study, the amount of bias was minimal for these factors that could be tested. At least for surveys of pediatricians, more attention should be devoted by investigators to assessments of response bias rather than relying on response rates as a proxy of response bias. PMID- 15663711 TI - Testing for statistical discrimination in health care. AB - OBJECTIVE: To examine the extent to which doctors' rational reactions to clinical uncertainty ("statistical discrimination") can explain racial differences in the diagnosis of depression, hypertension, and diabetes. DATA SOURCES: Main data are from the Medical Outcomes Study (MOS), a 1986 study conducted by RAND Corporation in three U.S. cities. The study compares the processes and outcomes of care for patients in different health care systems. Complementary data from National Health And Examination Survey III (NHANES III) and National Comorbidity Survey (NCS) are also used. STUDY DESIGN: Across three systems of care (staff health maintenance organizations, multispecialty groups, and solo practices), the MOS selected 523 health care clinicians. A representative cross-section (21,480) of patients was then chosen from a pool of adults who visited any of these providers during a 9-day period. DATA COLLECTION: We analyzed a subsample of the MOS data consisting of patients of white family physicians or internists (11,664 patients). We obtain variables reflecting patients' health conditions and severity, demographics, socioeconomic status, and insurance from the patients' screener interview (administered by MOS staff prior to the patient's encounter with the clinician). We used the reports made by the clinician after the visit to construct indicators of doctors' diagnoses. We obtained prevalence rates from NHANES III and NCS. FINDINGS: We find evidence consistent with statistical discrimination for diagnoses of hypertension, diabetes, and depression. In particular, we find that if clinicians act like Bayesians, plausible priors held by the physician about the prevalence of the disease across racial groups could account for racial differences in the diagnosis of hypertension and diabetes. In the case of depression, we find evidence that race affects decisions through differences in communication patterns between doctors and white and minority patients. CONCLUSIONS: To contend effectively with inequities in health care, it is necessary to understand the mechanisms behind the problem. Discrimination stemming from prejudice is of a very different character than discrimination stemming from the application of rules of conditional probability as a response to clinical uncertainty. While in the former case, doctors are not acting in the best interests of their patients, in the latter, they are doing the best they can, given the information available. If miscommunication is the culprit, then efforts should be aimed at reducing disparities in the ways in which doctors communicate with patients. PMID- 15663712 TI - Application of a propensity score approach for risk adjustment in profiling multiple physician groups on asthma care. AB - OBJECTIVES: To develop a propensity score-based risk adjustment method to estimate the performance of 20 physician groups and to compare performance rankings using our method to a standard hierarchical regression-based risk adjustment method. DATA SOURCES/STUDY SETTING: Mailed survey of patients from 20 California physician groups between July 1998 and February 1999. STUDY DESIGN: A cross-sectional analysis of physician group performance using patient satisfaction with asthma care. We compared the performance of the 20 physician groups using a novel propensity score-based risk adjustment method. More specifically, by using a multinomial logistic regression model we estimated for each patient the propensity scores, or probabilities, of having been treated by each of the 20 physician groups. To adjust for different distributions of characteristics across groups, patients cared for by a given group were first stratified into five strata based on their propensity of being in that group. Then, strata-specific performance was combined across the five strata. We compared our propensity score method to hierarchical model-based risk adjustment without using propensity scores. The impact of different risk-adjustment methods on performance was measured in terms of percentage changes in absolute and quintile ranking (AR, QR), and weighted kappa of agreement on QR. RESULTS: The propensity score-based risk adjustment method balanced the distributions of all covariates among the 20 physician groups, providing evidence for validity. The propensity score-based method and the hierarchical model-based method without propensity scores provided substantially different rankings (75 percent of groups differed in AR, 50 percent differed in QR, weighted kappa=0.69). CONCLUSIONS: We developed and tested a propensity score method for profiling multiple physician groups. We found that our method could balance the distributions of covariates across groups and yielded substantially different profiles compared with conventional methods. Propensity score-based risk adjustment should be considered in studies examining quality comparisons. PMID- 15663713 TI - Variations among Institutional Review Board reviews in a multisite health services research study. AB - OBJECTIVE: To document the Institutional Review Board (IRB) review process and to explore the impact of different patient notification procedures. DATA SOURCES/STUDY SETTING: Review of IRB application and correspondence records prospectively collected during a multisite study of health care quality involving telephone interviews of 3,000 participants across 15 primary care sites. STUDY DESIGN: Records were reviewed to ascertain: (1) the type of IRB review conducted, (2) the number of days from submission to approval of the IRB application, (3) whether the IRB required patient notification and/or consent prior to the release of names, and (4) patient participation rates. DATA COLLECTION/EXTRACTION METHODS: The study coordinating center prepared a common study protocol for IRB submission and assisted sites with submission. The application, correspondence with the IRB, consent script, and patient letters were collected, reviewed, coded, and analyzed. PRINCIPAL FINDINGS: IRBs at the 15 sites and survey center varied in the type of IRB required and the number of days from submission to approval (range of 5-172 days). Four sites required patient notification in advance of the study; 2-11 percent of patients refused in opt-out sites and 37 percent in the single opt-in site. Participation among contacted patients did not appear to be related to patient notification procedures. CONCLUSIONS: Variations in IRB requirements can affect response rates and sample generalizability. PMID- 15663714 TI - Institutional review boards and multisite studies in health services research: is there a better way? AB - OBJECTIVE: The following paper examines the issue of whether the current system for ethics review of multisite health services research protocols is adequate, or whether there exist alternative methods that should be considered. PRINCIPAL FINDINGS: (1) Investigators at different sites in a multisite project often have very different experiences with respect to the requirements and requests of the review board. Other problems include the waste of time and resources spent on document preparation for review boards, and delays in the commencement of research activities. (2) There are several possible reasons why there is variability in ethics review. These include the absence of standardized forms, differences in the background and experiences of board members, the influence of institutional or professional culture, and regional thinking. (3) Given the limited benefits derived from the variability in recommendations of multiple boards and the numerous problems encountered in seeking ethics approval from multiple boards suggest that some sort of reform is in order. CONCLUSIONS: The increasing number of multisite, health services research studies calls for a centralized system of ethics review. The local review model is simply not conducive to multisite studies, and jeopardizes the integrity of the research process. Centralized multisite review boards, together with standardized documents and procedure, electronic access to documentation, and training for board members are all possible solutions. Changes to the current system are necessary not only to facilitate the conduct of multisite research, but also to preserve the integrity of the ethics approval process in general. PMID- 15663716 TI - Circulating endothelial (progenitor) cells reflect the state of the endothelium: vascular injury, repair and neovascularization. AB - An increase in the number of circulating endothelial cells (CEC) and of bone marrow-derived endothelial progenitor cells (EPC) in the peripheral blood is associated with vascular injury, repair and neovascularization. The phenotype and number of CEC may serve as diagnostic or prognostic parameters of vascular injury and tumour growth. An increase in the number of EPC may reflect repair of ischaemic vascular injury, a finding which has resulted in the initiation of clinical cardiovascular pilot trials using cell therapy. However, there is no consensus on the exact phenotype of the EPC and haematopoietic stem cells (HSC) and therefore the best candidate cell for transplant has not been established. Although the use of peripheral blood stem cells following mobilization, or of ex vivo-expanded cells, may improve EPC-mediated vascular graft endothelialization or tissue vascularization, sustained EPC-induced neovascularization still needs to be proven. Flow cytometric characterization, in combination with functional assays, will further elucidate the phenotype of the CEC and EPC, thereby providing reliable detection to appreciate their role in vascular diseases and cancer and to evaluate and, if possible, improve their therapeutic potential. PMID- 15663717 TI - Epidemiology of blood donors in Japan, positive for hepatitis B virus and hepatitis C virus by nucleic acid amplification testing. AB - BACKGROUND AND OBJECTIVES: The Japanese Red Cross screens seronegative blood donors by nucleic acid amplification testing (NAT) for hepatitis B, hepatitis C and human immunodeficiency virus-1 markers. NAT-positive donors thus identified seemed to have a different infectious background from serologically positive donors. The purpose of our study was to characterize this background in the hepatitis B virus (HBV) and hepatitis C virus (HCV) NAT-positive donors. MATERIALS AND METHODS: Some 328 HBV DNA-positive and 44 HCV RNA-positive donors were detected by NAT testing of seronegative blood donors. These were characterized regarding age, gender and genotype of HBV and HCV. RESULTS: Those who were HBV NAT-positive were mainly young, in particular teenage girls. In Japan, genotypes C and B have previously been dominant, but recently genotype A has increased, and genotype H was recently detected. In HBV NAT-positive donors, the rate of genotype A was high (12.2%) compared with patients in hospital (1.7 2%). Donors who were HCV NAT-positive were also young, but mostly men in their twenties. The ratio of genotype 1b to 2a or 1b to 2b in HCV NAT-positive donors differed from that of hospitalized patients in Japan. We did not find genotype 1a, which is dominant in the USA. CONCLUSIONS: The high-risk donors detected by NAT were mainly young, with a different distribution of genotypes from that of hospitalized patients, regarding both HBV and HCV. The rare HBV genotype H has been found for the first time in Japan. The findings reflect the present spread of hepatitis viruses B and C. PMID- 15663718 TI - The impact of two whole blood inline filters on markers of coagulation, complement and cell activation. AB - BACKGROUND AND OBJECTIVES: There exists a current lack of information about the impact of different inline filters, used for the leucoreduction of whole blood (WB), on the levels of clotting factors and markers of coagulation, complement and cell activation in plasma. Only a few small comparisons of different types of WB inline filters have been published to date. MATERIALS AND METHODS: This study compared two plasma types of 200 units each. Both study groups were derived from WB, inline-filtered and held for 2 h at 20 degrees between donation and filtration. Then, 200 units (Group A) were filtered using a positively charged polyester filter (Baxter RZ2000) and the other 200 units (Group B) were filtered using an uncharged polyester filter (Fresenius). After filtration, both groups were analysed for fibrinogen, factors V and VIII:C (FV and FVIII:C, respectively), immunoglobulin G (IgG), residual leucocytes and platelets, and markers of coagulation, complement and cell activation. Predonation plasma samples from CPDA1-anticoagulated blood were obtained from 100 different individuals and served as controls. RESULTS: WB inline filtration did not influence fibrinogen, FV, FVIII:C or IgG levels. Neither filter induced thrombin or fibrin formation. The charged filter caused substantial complement activation and neutrophil elastase and platelet factor 4 release. In contrast, the plasma filtered through the uncharged filter showed markedly lower levels of C3a-desArg, C5a, neutrophil elastase and platelet factor 4, and moderately reduced levels of prothrombin fragments 1+2 and D-dimer, compared with controls. CONCLUSIONS: Filter type has a significant impact on the quality of plasma derived from WB filtered through inline filtration systems. Some filters produce substantial coagulation and complement activation and cell release, while others appear to reduce the plasma levels of activation markers. The clinical significance of these findings remains to be determined. PMID- 15663719 TI - Haemovigilance in a general university hospital: need for a more comprehensive classification and a codification of transfusion-related events. AB - BACKGROUND AND OBJECTIVES: The purpose of this study was to analyse the transfusion-related events recorded in a general university hospital. MATERIALS AND METHODS: The method we used was retrospective analysis of the data collected between 1999 and 2003. RESULTS: The incidence of transfusion reactions (n = 394) was 4.19 per 1000 blood products distributed: 59% (n = 231) were febrile non haemolytic transfusion reactions; 22% (n = 88) were caused by allergy; 5% (n = 21) were caused by bacterial infection; and 14% (n = 54) were classified as other reactions. Platelet concentrates gave rise to a significantly greater number of reactions than erythrocyte concentrates and fresh-frozen plasma. Transfusion errors and near-miss events were also observed and were analysed separately. A series of transfusion-related events, such as haemosiderosis, metabolic disturbances or volume overload, were not reported. CONCLUSIONS: Our experience prompts us to propose a more comprehensive classification and codification of transfusion-related events. PMID- 15663720 TI - Association of ABO gene mutations resulting in a rare B subgroup. AB - BACKGROUND AND OBJECTIVES: B subgroups are rare and the genetic analysis reported to date has been limited. MATERIALS AND METHODS: Serological and molecular investigations were performed in blood from a B-subgroup donor. RESULTS: Red cells did not react with anti-B and anti-AB reagents. However, cells absorbed anti-B. Red cells presented positive reactions with anti-H, and saliva secreted H substance. The molecular study demonstrated a B allele with the substitutions 467C>T, 646T>A, 681G>A, 771C>T, 796C>A, 803G>C, 829G>A and an O allele with the sequence of O02. CONCLUSIONS: It is probable that the presence in exon 7 of some of the O02 substitutions could have weakened the enzymatic activity of the encoded B transferase. PMID- 15663721 TI - Systemic analysis and zygosity determination of the RHD gene in a D-negative Chinese Han population reveals a novel D-negative RHD gene. AB - BACKGROUND AND OBJECTIVES: The aim of this study was to systemically analyse the genetic background of D negativity in a Chinese Han population. MATERIALS AND METHODS: DNA of 74 D-negative samples was analysed by using an RHD multiplex polymerase chain reaction (MPX PCR) for the presence of RHD and by PCR restriction fragment length polymorphism (PCR-RFLP) for RHD zygosity determination. Sixty-five samples were additionally analysed by using real-time quantitative PCR on RHD exon 7. RHD exon-specific sequencing was performed on discrepant samples. RESULTS: Forty-six samples (62%) showed the absence of RHD specific exons by RHD MPX PCR and homozygous RHD negativity by PCR-RFLP. Twenty two samples (30%) showed a 1227G>A mutation, characteristic for the Del phenotype. Five (7%) samples showed all characteristics of the RHD(1-2)-CE(3-9) D(10) hybrid gene. One sample (1.4%) showed a novel 933C>A nonsense mutation in RHD exon 6, which resulted in a premature stop codon. CONCLUSIONS: The RHD gene deletion, RHD-CE-D hybrid genes and one novel 93C>A mutation were found to be the three mechanisms that cause D negativity in our samples. The 1227G>A Del mutation was found to be the major cause of discrepant results between genotyping and phenotyping strategies, favouring genotyping of D-negative samples. PMID- 15663722 TI - Heterogeneity of platelet responsiveness to anti-CD36 in plasma associated with adverse transfusion reactions. AB - BACKGROUND AND OBJECTIVES: Antibodies to CD36 (anti-CD36) are clinically important. As some platelet immunoglobulins produced by transfusion or pregnancy have been shown to induce platelet activation and to play roles in non-haemolytic transfusion reactions (NHTRs), we investigated the in vitro response of platelets to plasma containing anti-CD36. MATERIALS AND METHODS: Plasma containing anti CD36, implicated in the development of NHTRs and subsequent thrombocytopenia, was incubated with CD36-positive platelets. Plasma-induced platelet activation was examined by evaluating platelet aggregation and RANTES (regulated on activation, normal, T-cell expressed, and presumably secreted) release. RESULTS: Platelet activation was induced by plasma alone in four out of 20 CD36-positive subjects. In seven subjects, platelet activation was synergistically induced by the combination of epinephrine priming and the plasma. The platelets of the nine remaining subjects failed to respond to the plasma. Platelet activation induced by either the plasma alone or by synergy with epinephrine required the involvement of Fc gamma RIIa. The different responsiveness of the platelets was partially associated with the surface levels of CD36 and Fc gamma RIIa, but not with Fc gamma RIIa polymorphisms. CONCLUSIONS: Plasma containing anti-CD36, implicated in the development of NHTRs, exhibited a platelet-activating capability. Additionally, platelets from healthy human subjects exhibited a considerable degree of heterogeneity in their responsiveness to this plasma. The heterogeneity of these responses may determine the occurrence of anti-CD36 related NHTRs. PMID- 15663723 TI - Primary autoimmune neutropenia in children: a study of neutrophil antibodies and clinical course. AB - BACKGROUND AND OBJECTIVES: Primary autoimmune neutropenia (AIN) in children is characterized by severe neutropenia, but mild bacterial infections and a spontaneous resolution. Neutrophil autoantibodies are involved in the disease. The precise relationship between the specificity and level of reactivity of the antibodies with the absolute neutrophil count and frequency of infections is not known. To obtain a better insight into this relationship, we performed a follow up study in 15 patients with primary AIN. In addition, we performed two different neutrophil antibody tests to evaluate their sensitivity and specificity. MATERIALS AND METHODS: Blood samples from 15 children were tested for neutrophil antibodies, at different time-points during the disease, by using the indirect granulocyte immunofluorescence test (GIFT) and the monoclonal antibody-specific immobilization of granulocyte antigens (MAIGA) assay. Clinical data related to the occurrence of bacterial infections and treatment, and neutrophil counts were collected. RESULTS: Early in the disease, antibodies with pan-FcRIIIb specificity were detected, and HNA-1a or HNA-1b specificity of the antibodies developed over time. The sensitivity and specificity of neutrophil antibody detection tests were higher in the GIFT than in the MAIGA assay. Variables predicting time of recovery from neutropenia were not found. Prophylactic antibiotics led to the almost complete disappearance of infections. CONCLUSIONS: In patients with primary neutropenia, neutrophil antibody specificity changes over time. Prophylactic antibiotics do benefit the patients. PMID- 15663724 TI - Transfusion-transmitted bacterial infection: risks, sources and interventions. PMID- 15663725 TI - The ability of HPA-5b typing to predict vascular occlusion events. PMID- 15663726 TI - Prophylactic treatment of patients with haemophilia A or haemophilia B. PMID- 15663733 TI - Castration of piglets: the analgesic effects of intratesticular and intrafunicular lidocaine injection. AB - OBJECTIVE: The aim of this study was to evaluate the analgesic effect of intratesticular and intrafunicular lidocaine for the surgical castration of piglets and to investigate the degree of nociception induced by lidocaine injection. STUDY DESIGN: Prospective controlled experimental study. ANIMALS: Forty-seven male Norwegian landrace piglets with normal testicular anatomy, aged 22 (+/-2.6 SD) days and weighing 7.4 +/- 1.4 kg. MATERIALS AND METHODS: Anaesthesia was induced and maintained using halothane delivered in oxygen. End tidal halothane was stabilized at 1.3% for 20 minutes before mean arterial blood pressure (MAP) pulse rate and electroencephalography (EEG) monitoring began. After 5 minutes of data collection, scrotal skin was desensitized with lidocaine before either an intrafunicular (IF) (n = 15) or an intratesticular (IT) (n = 16) lidocaine injection was made. Pigs in the control group (n = 16) did not receive lidocaine. Ten minutes later, a scalpel and an emasculator were used to cut the funiculus spermaticus. The MAP, pulse rate and EEG were monitored continuously for 5 minutes after castration. RESULTS: During castration, MAP increased significantly, while pulse rate and EEG theta power fell significantly more in control, compared with the IT or IF groups. EEG alpha power fell more in the control group than in the IF group. No significant differences were found between the IF and IT groups. EEG, MAP and pulse rate responses to castration in the control group were significantly larger than the response to lidocaine injection. CONCLUSION/CLINICAL RELEVANCE: Injecting lidocaine into the funiculus spermaticus or into the testes is effective in reducing signs of nociception caused by castration. Lidocaine injection is less noxious than castration without local anaesthetic. PMID- 15663734 TI - Effects of peri-operative morphine administration during halothane anaesthesia in horses. AB - OBJECTIVE: To study the effects of morphine on haemodynamic variables, blood gas values and the requirement for additional anaesthetic drugs in horses undergoing surgery. STUDY DESIGN: Prospective randomized study. METHODS: Thirty-eight client owned horses, ASA(American Society of Anesthesiologists) category I or II, undergoing elective surgical procedures, were studied. Horses were divided between two groups, and were paired according to operation, anaesthetist, body position during surgery, mass and breed. Group M+ received morphine by intravenous (IV) injection (0.15 mg kg(-1)) before induction of anaesthesia and then by infusion (0.1 mg kg(-1) hour(-1)) throughout anaesthesia. Group M- received the same anaesthetic technique (pre-anaesthetic medication with romifidine (100 microg kg(-1)) IV; induction with ketamine (2.2 mg kg(-1)) and diazepam (50 microg kg(-1)) IV; maintenance with halothane), except that morphine was excluded. Both groups received flunixin IV (1.1 mg kg(-1)) before surgery. Both groups also received 50% nitrous oxide for the first 10 minutes of anaesthesia. During anaesthesia, end-tidal halothane was maintained at 0.9% (+/ 0.1%) in both groups. Heart rate (HR) and respiratory rate (fr), systolic, mean and diastolic arterial pressures were recorded every 5 minutes. Arterial blood samples were analysed every 20 minutes. Additional anaesthetics (ketamine and midazolam) were administered whenever the horse moved. Dobutamine was infused to maintain mean arterial pressure (MAP) > 58 mm Hg, but was discontinued when MAP reached 68 mm Hg. Mechanical ventilation was imposed when PaCO(2) exceeded 9.3 kPa (70 mm Hg). RESULTS: Haemodynamic data (HR and MAP) and blood gas measurements were analysed using repeated measure analysis using a mixed covariance pattern model (SAS version 8.2). A Student's t-test was used to investigate differences between groups in the doses of additional anaesthetics required. There were no significant differences between M+ or M- groups in MAP (p = 0.65), HR (p = 0.74), PaO2 (p = 0.40) or PaCO2 (p = 0.20). Fewer horses in the M+ group received additional anaesthetics (15.8% compared to 21.1% in M- group), and the mean dose of ketamine required was higher in the M- group (mean +/- SD: M , 0.93 +/- 0.70; M+, 0.45 +/- 0.17). These differences were not statistically significant (p = 0.28). CONCLUSIONS: Pre-anaesthetic and peri-operative morphine administration is not associated with significant haemodynamic or ventilatory changes. Horses receiving morphine tended to receive fewer and lower doses of additional anaesthetic drugs, although this was not statistically significant. PMID- 15663735 TI - Comparison of morphine and butorphanol as pre-anaesthetic agents in combination with romifidine for field castration in ponies. AB - OBJECTIVE: The aim of this study was to compare two different alpha2 agonist opioid combinations in ponies undergoing field castration. STUDY DESIGN: Prospective double-blind randomized clinical trial. ANIMAL POPULATION: Fifty-four ponies undergoing field castration. MATERIALS AND METHODS: The ponies were randomly allocated to receive one of three different pre-anaesthetic medications [intravenous (IV) romifidine 100 microg kg(-1) and butorphanol 50 micro kg(-1); romifidine 100 microg kg(-1) and morphine 0.1 mg kg(-1) IV, or romifidine 100 microg kg(-1) and saline IV] before induction of anaesthesia with ketamine 2.2 mg kg(-1) IV. Further doses of romifidine (25 microg kg(-1)) and ketamine (0.5 mg kg(-1)) were given when required to maintain anaesthesia. Quality of sedation, induction of anaesthesia, maintenance of anaesthesia, recovery, and surgical condition were assessed using a visual analogue scale scoring system and compared. The effects of the different drug combinations on heart and respiratory rate were evaluated and the recovery time was recorded. RESULTS: Anaesthesia was considered adequate for surgery in all ponies. No anaesthetic complications were observed. Quality of sedation was significantly better in the butorphanol group compared with the control group (p = 0.0428). Overall quality of anaesthesia was better in the butorphanol group compared with morphine (p = 0.0157) and control (p < 0.05) groups. Quality of induction of anaesthesia and recovery were not significantly different between groups, nor were the surgical conditions, recovery time and the number of repeated anaesthetic doses required during the procedure. Muscle twitches were observed in both the control and morphine groups. Maintenance of anaesthesia was judged to be smoother in the butorphanol group compared with the morphine and control groups (p = 0.006). Heart rate decreased significantly (p < 0.01) in all groups after administration of sedatives but did not differ significantly between groups at any time point. CONCLUSION: The combination of butorphanol and romifidine was found to provide better sedation compared with the other drug combinations. CLINICAL RELEVANCE: The combination of butorphanol and romifidine provided better sedation, but morphine was found to be a suitable alternative to butorphanol. Use of morphine and butorphanol in combination with alpha2 agonists should be further investigated to assess their analgesic effects. PMID- 15663736 TI - The effects of two endotracheal tube cuff inflation pressures on liquid aspiration and tracheal wall damage in horses. AB - OBJECTIVE: To investigate the effect of endotracheal tube cuff inflation pressure on the occurrence of liquid aspiration and tracheal wall damage. STUDY DESIGN: Prospective, randomized experimental study. ANIMALS: Ten healthy horses, weighing 535 +/- 55 kg. METHODS: Horses were anesthetized, orotracheally intubated, placed in dorsal recumbency, and maintained on isoflurane in oxygen with controlled ventilation for 175 +/- 15 minutes. The horses were randomly assigned to an endotracheal cuff pressure of 80-100 or 120 cm H2O. The cuff pressure was continuously monitored and maintained at a constant pressure. Methylene blue in saline was instilled proximal to the cuff. After euthanasia, the trachea was opened distal to the endotracheal tube tip to check for evidence of dye leaking past the cuff. The cervical trachea was then resected and opened longitudinally for gross and histologic examinations. RESULTS: No blue staining was found distal to the cuff in any horse. Visual examination of the tracheal mucosa revealed hyperemic and hemorrhagic lesions at the site of the cuff contact. Histologic changes included epithelium attenuation or erosion, submucosal neutrophilic infiltration, and submucosal hemorrhages. Lesions were absent or less extensive in the lower cuff pressure group as compared to the high cuff pressure group. CONCLUSIONS: The endotracheal tube cuff produced a seal sufficient to prevent leakage in both groups. Tracheal wall damage was more severe and occurred more frequently in the higher cuff pressure group. CLINICAL RELEVANCE: Tracheal mucosal damage induced by cuff inflation is pressure-dependent. Cuff pressure monitoring is recommended. PMID- 15663737 TI - Evaluation of analgesia provided by the administration of epidural ketamine in dogs with a chemically induced synovitis. AB - OBJECTIVE: To determine if epidural ketamine provides analgesia in dogs with a chemically induced synovitis. STUDY DESIGN: Prospective randomized experimental trial. ANIMALS: Thirty-two healthy, adult mongrel dogs (13-30 kg). METHODS: (Part I) Synovitis was induced in the right stifle of 16 dogs and allowed to develop for 12 hours. Epidural injection at the lumbosacral space of either ketamine (2 mg kg(-1); n = 8) or placebo (n = 8) was performed. Limb use and pain were measured using a force platform and numerical rating scale (NRS). Assessments were performed before and at 12, 14, 16, 18, 20, and 24 hours after the induction of synovitis. (Part II) Epidural injection of either ketamine (n = 8) or placebo (n = 8) was performed immediately before the induction of synovitis. Analgesia was assessed as in Part I. Assessments occurred before and at 2, 4, 6, 8, and 12 hours after the induction of synovitis. RESULTS: (Part I) Vertical ground reaction forces (VGRF) significantly decreased and NRS scores of total pain significantly increased after the induction of synovitis in all dogs (p < 0.05). No significant differences in VGRF or NRS scores were measured between treatment groups at any assessment period. (Part II) Dogs that received ketamine had significantly lower NRS scores 2 hours after treatment (p < 0.05). NRS scores did not differ between groups at any other evaluation. VGRF did not differ significantly between treatment groups at any assessment period. CONCLUSION: Epidural ketamine at a dose of 2 mg kg(-1) administered after the development of synovitis does not provide significant levels of analgesia. Administration of ketamine before the induction of synovitis significantly decreased the NRS score 2 hours post-induction. CLINICAL RELEVANCE: Administration of epidural ketamine before tissue injury may provide analgesia of short duration in dogs. PMID- 15663738 TI - Preliminary comparison of four anaesthetic techniques in badgers (Meles meles). AB - OBJECTIVE: To investigate the use of four ketamine-based anaesthetic combinations in wild badgers. STUDY DESIGN: Prospective, randomized, clinical trial. ANIMALS: Twenty-four adult badgers. MATERIALS AND METHODS: Animals were divided into four groups of six and were anaesthetized using either intramuscular (IM) ketamine alone (20 mg kg(-1)), ketamine (15 mg kg(-1) IM) and midazolam (0.4 mg kg(-1) IM), ketamine (10 mg kg(-1) IM) and midazolam (1 mg kg(-1) IM) or ketamine (5 mg kg(-1) IM) and medetomidine (80 microg kg(-1) IM) antagonized with atipamezole (0.8 mg kg(-1); IM). Features of each technique, i.e. quality of induction, maintenance and recovery, and the need for additional doses, were assessed using a simple descriptive scale. Physiological variables, i.e. rectal temperature, respiratory rate, heart rate and blood pressure, were also recorded. RESULTS: Combinations of ketamine and midazolam did not produce adequate anaesthesia. The combination of medetomidine and ketamine had few advantages over ketamine alone. CONCLUSIONS AND CLINICAL RELEVANCE: These data will contribute to a wider study attempting to refine anaesthetic techniques in badgers. PMID- 15663739 TI - High rectal temperature indicates an increased risk of unexpected recovery in anaesthetized badgers. AB - OBJECTIVE: To identify factors associated with sudden early recovery (SER) from anaesthesia in badgers (Meles meles). STUDY DESIGN: Experimental trial. ANIMALS: Ninety-three adult wild badgers. METHODS: Animals were randomly assigned to receive one of four anaesthetics based on medetomidine (M) ketamine (K) and butorphanol (B) combined in different ratios: (i) MKB 20:40:80 microg kg(-1); (ii) MKB 20:40:60 microg kg(-1); (iii) MKB 20:60:40 microg kg(-1); and (iv) ketamine alone 0.2 mg kg(-1). For each animal, induction time was measured and physiological variables (heart rate, respiratory rate and rectal temperature) were recorded at 5-minute intervals during anaesthesia. Cases of SER were recorded and binary logistic regression applied to identify predictive factors. RESULTS: Fourteen animals (15%) exhibited SER. Rectal temperature was the only variable that was a significant predictor of SER. Animals showing SER had significantly higher rectal temperatures which, in contrast to other cases, did not fall during the first 10 minutes of anaesthesia, which was when most SERs occurred. CONCLUSION AND CLINICAL RELEVANCE: We recommend that (i) rectal temperature is closely monitored during wild badger anaesthesia and (ii) that animals with higher than expected temperatures are treated with additional caution. PMID- 15663740 TI - Overdose during chemical restraint in a black rhinoceros (Diceros bicornis). AB - A juvenile female black rhinoceros (Diceros bicornis) was successfully treated after overdose of drugs used for chemical restraint. Subsequent general anaesthesia for surgical reduction of a recurrent rectal prolapse was uneventful. Over a 25-minute period before transportation to the veterinary hospital, the animal received a total dose of 1.225 mg etorphine, 30 mg acepromazine and 30 mg detomidine. Based on an estimated mass of 200 kg, these corresponded to doses of 6.1 microg kg(-1) etorphine, 150 microg kg(-1) acepromazine, and 150 microg kg( 1) detomidine which constitutes considerable overdose for each drug given separately, notwithstanding the synergy that probably resulted when the three drugs were present concurrently. The estimated body mass may have substantially overestimated the actual body mass and exacerbated overdosage. The animal was recumbent and apnoeic on arrival at the hospital. Heart sounds were auscultated and a weak peripheral pulse was palpated; no pulse deficits were detected, although the heart rate was low. The trachea was intubated, inspired breath was enriched with oxygen and the lungs ventilated manually. Diprenorphine (1.5 mg) was given intravenously and spontaneous breathing resumed 11 minutes later. After induction of general anaesthesia using isoflurane, emergency surgery for correction of rectal prolapse was performed, from which the animal recovered uneventfully. The case highlights some of the practical problems that may be encountered in dealing with dangerous and unfamiliar species. PMID- 15663742 TI - Allelic polymorphism in introns 1 and 2 of the HLA-DQA1 gene. AB - Human leukocyte antigen (HLA) class II antigens are highly polymorphic membrane glycoproteins, encoded by the A and B genes of DR, DQ, and DP. The polymorphism is mainly located in exon 2, with the exception of DQA1. Of the 27 DQA1 alleles presently known, 18 cannot be identified on the basis of exon 2 alone, but need additional information from the other exons. DQA1 has been reported to be the most ancient class II gene. For evolutionary comparison and to assess the degree of polymorphism outside the exons, the sequences of introns 1 and 2 were determined from 30 different cell lines, encompassing 15 different DQA1 alleles. The sequences revealed major nucleotide differences between the different lineages, whereas within each lineage few differences were present. Phylogenetic analysis of intron and exon sequences confirmed this lineage specificity. Altogether, the present data indicate that the HLA-DQA1 lineages represent ancient entities. The observed variation of the introns in alleles with identical exon sequences implicates conservative selection of the exons within a given lineage. Intron sequences may provide the means to set up an accurate typing system. PMID- 15663741 TI - The HLA Dictionary 2004: a summary of HLA-A, -B, -C, -DRB1/3/4/5 and -DQB1 alleles and their association with serologically defined HLA-A, -B, -C, -DR and DQ antigens. AB - This report presents serologic equivalents of human leucocyte antigen (HLA)-A, B, -C, -DRB1, -DRB3, -DRB4, -DRB5 and -DQB1 alleles. The dictionary is an update of the one published in 2001. The data summarize equivalents obtained by the World Health Organization Nomenclature Committee for factors of the HLA System, the International Cell Exchange, the National Marrow Donor Program, recent publications and individual laboratories. This latest update of the dictionary is enhanced by the inclusion of results from studies performed during the 13th International Histocompatibility Workshop and from neural network analyses. A summary of the data as recommended serologic equivalents is presented as expert assigned types. The tables include remarks for alleles, which are or may be expressed as antigens with serologic reaction patterns that differ from the well established HLA specificities. The equivalents provided will be useful in guiding searches for unrelated hematopoietic stem cell donors in which patients and/or potential donors are typed by either serology or DNA-based methods. The serological DNA equivalent dictionary will also aid in typing and matching procedures for organ transplant programs whose waiting lists of potential donors and recipients comprise of mixtures of serologic and DNA-based typings. The tables with HLA equivalents and a questionnaire for submission of serologic reaction patterns for poorly identified allelic products will be made available through the WMDA web page: www.worldmarrow.org. and in the near future also in a searchable form on the IMGT/HLA database. PMID- 15663743 TI - New strategies for efficient typing of HLA class-II loci DQB1 and DRB1 by using Pyrosequencing. AB - The characterization of genetic risk factors for complex diseases located on chromosome-6 frequently requires human leucocyte antigen (HLA) genotyping of large patient cohorts. Currently available methods do not support high-throughput HLA typing beyond the major allele group level. We, thus, developed a high throughput approach for the HLA-DQB1 and HLA-DRB1 loci that is based on Pyrosequencing. Pyrosequencing offers a higher degree of automation than direct sequencing or oligotyping. Using a dispensation order optimized for the particular HLA locus, rapid group typing and fine resolution can be achieved. We implemented the method for two important HLA loci--DQB1 and DRB1. The HLA-DQB1 typing method comprises the following steps: splitting the potential alleles after a generic polymerase chain reaction (PCR) amplification into groups with a first Pyrosequencing reaction and resolving the split allele groups by means of five further Pyrosequencing reactions. The HLA-DR gene family is known to be the most polymorphic one in the HLA class-II region because of a large number of DRB1 alleles. Because of this complex nature, HLA-DRB1 typing was performed by means of a combination of sequence-specific PCR typing and Pyrosequencing. HLA-DQB1 typing and HLA-DRB1 typing were performed successfully by using standard DNA samples with the help of known HLA genotypes and in a blind study by using the samples from the Deutscher Zell Austausch 2002 and 2003. The approach was optimized and was practically tested for genotyping in disease association studies. Our well-elaborated Pyrosequencing-based protocols offer a new alternative to the existing HLA class-II typing methods and represent a convenient and economic solution, a unique combination of high accuracy with high sample throughput. PMID- 15663744 TI - A novel blocking monoclonal antibody recognizing a distinct epitope of human CD40 molecule. AB - CD40, a member of the tumor necrosis factor receptor superfamily, is an important costimulatory molecule during the immune response. Here, we report a blocking mouse antihuman CD40 monoclonal antibody, mAb 3G3, of which the specificity was verified by flow cytometry and Western blot. It was shown by competition test that 3G3 bound to a different site (epitope) of CD40 from the reported CD40 mAbs, including clone mAb89, 3B2, and 5C11. It was also found that mAb 3G3 could inhibit homotypic aggregation of Daudi cells induced by the agonistic anti-CD40 mAb 5C11. Furthermore, mAb 3G3 effectively inhibited the proliferation of peripheral blood mononuclear cells in mixed lymphocyte reaction assay. Finally, a sensitive and specific soluble CD40 (sCD40) ELISA kit was established by matching mAb 3G3 with 5C11, and it was found that the levels of sCD40 in sera from patients with immune disorders such as hyperthyroidism, chronic nephritis, and rheumatoid arthritis were obviously higher than those from normal individuals. Thus, this blocking anti-CD40 mAb provides a novel tool for the study of CD40. PMID- 15663745 TI - Association of interleukin-18 gene single-nucleotide polymorphisms with susceptibility to inflammatory bowel disease. AB - Interleukin-18 (IL-18) is believed to be one of the most important cytokines in the pathogenesis of inflammatory bowel disease (IBD). The aim of the study was to clarify the significance of single-nucleotide polymorphisms (SNPs) at the 5'-end of the IL-18 gene in the development of IBD. DNA was obtained from peripheral blood of 99 patients with ulcerative colitis (UC), 79 patients with Crohn's disease (CD), and 102 healthy controls. All participants were Japanese. SNPs at 656G/T, -607C/A, -137G/C, +113T/G, and +127C/T were determined by means of direct sequencing, and a genetic association with IBD was examined. The frequencies of the G allele at +113 and the T allele at +127 were significantly higher in patients with CD and UC compared with controls. The differences in allelic frequencies were more striking in patients with CD than in patients with UC, and at position +127 than at position +113. The haplotype estimation, according to the E-M algorithm, suggested that TACGT is closely associated with IBD, especially with CD. It was concluded that SNPs at the 5'-end of IL-18 gene might be closely related to the etiology of IBD. PMID- 15663746 TI - TNF-alpha-, TNF-beta-, IL-6-, and IL-10-promoter polymorphisms in patients with chronic obstructive pulmonary disease. AB - Chronic obstructive pulmonary disease (COPD) is a major health problem. The disease is driven by abnormal inflammatory reactions in response to inhaled particles and fumes. Therefore, inflammatory mediators are postulated to be of distinct importance. In the present case-control study, we investigated interleukin (IL)-promoter polymorphisms known to correlate with altered transcription levels of their gene products in patients with COPD. We analyzed tumor necrosis factor-alpha (TNF-alpha)-308, TNF-beta-intron1-252, IL-6-174, IL 10-819, and IL-10-1082 polymorphisms in 469 individuals using restriction fragment length polymorphism-based converted polymerase chain reaction. The study population consisted of 113 patients with COPD based on chronic bronchitis, divided into subgroups by severity (I degrees -III degrees ), 113 matched hospitalized individuals suffering from severe coronary heart disease without pulmonary disease (age-, sex-, and smoking-matched control group), and 243 healthy individuals (population control group). The matched analysis showed no significant differences in genotype distribution of all tested polymorphisms between the matched controls and the COPD patients. However, comparison with the population controls revealed significant differences in IL-10-1082 A/G genotype frequencies (P = 0.0247 for the whole COPD group, P = 0.009 for smokers only), with the genotypes carrying the G allele more common in the COPD cases [odds ratio (OR) = 1.66, 95% confidence interval (CI) 1.01-2.75; P = 0.046]. Interestingly, this shift toward more G alleles was even more pronounced in the matched control group (OR = 2.55, 95% CI 1.47-4.41; P = 0.0007), suggesting both presented groups share corresponding underlying mechanisms. The IL-10-1082_G allele is known to correlate with altered IL-10 levels. Therefore, it might be associated with altered or abnormal inflammatory response, a mechanism that could be postulated to be important in both chronic bronchitis and coronary heart disease. PMID- 15663747 TI - Emerging new alleles suggest high diversity of HLA-C locus. AB - Human leukocyte antigen (HLA)-C has only recently emerged as an important transplantation antigen and as a receptor for natural killer cells. Over the last few years, sequence-based typing (SBT) revealed the true diversity of HLA-C locus; however, the frequency at which new alleles are detected still remains high. During routine SBT of 3500 samples for the National Marrow Donor Program, we have identified 20 new HLA-C alleles reported in this article in 26 individuals. New variants have been characterized by direct sequencing of polymerase chain reaction product obtained by allele-specific amplification of potential new alleles. Most of the new alleles carry coding substitutions of residues located within the antigen-binding groove. The substitutions are predominantly located in the alpha2-helix which is consistent with the unique to HLA-C conservation of alpha1-helix. Seven new alleles, or 35%, have been identified in African Americans, two of them in three and four individuals each, suggesting that these alleles may not be rare. This observation reflects the fact that the minority groups, previously under-represented in the HLA research pools subjected to SBT, now begin to emerge as a main source of new HLA-C alleles. This study further confirms that HLA-C locus is at least as polymorphic as HLA-A and HLA-B. PMID- 15663748 TI - Sequence of a novel HLA-A*0301 intronic variant (A*03010103). AB - We report here the full-length sequence of a novel HLA-A*0301 allele, A*03010103, which differs from A*03010101 by a single nucleotide substitution (G>T) at position 492 within intron 2. The variant was originally identified by Reference Strand-mediated Conformational Analysis (RSCA) and was confirmed by cloning and sequencing. The difference in RSCA mobility between A*03010101 and A*03010103 demonstrates the sensitivity of RSCA to detect single nucleotide polymorphisms. PMID- 15663749 TI - Interleukin-6 (IL-6)--373 A9T11 allele is associated with reduced susceptibility to chronic periodontitis in Japanese subjects and decreased serum IL-6 level. AB - We investigated whether interleukin-6 (IL-6) gene polymorphisms could be associated with chronic periodontitis (CP) and serum IL-6 level. One hundred and twelve CP and 77 non-CP Japanese subjects were analyzed for IL-6 -597 (G/A), -572 (C/G), -373 (A(n)T(m)), -190 (C/T), and -174 (G/C) polymorphisms. We could only detect -572 and -373 polymorphisms and found that the frequency and carriage rate of the -373 A9T11 allele were significantly higher in non-CP subjects. Enzyme linked immunosorbent assay confirmed that the -572 and -373 G[A9T11] haplotypes were associated with lower serum IL-6 level. These findings suggest that IL-6 373 A9T11 allele could be associated with reduced susceptibility to CP among Japanese subjects and decreased serum IL-6 level. PMID- 15663750 TI - Extended DR3-D6S273-HLA-B haplotypes are associated with increased susceptibility to type 1 diabetes in US Caucasians. AB - Human leukocyte antigen (HLA)-DR3 haplotypes are associated with susceptibility to type 1 diabetes (T1D). Reports from Northern European populations show that an allele (D6S273*2) at a microsatellite mapping to HLA class III marks an extended DR3-B18 haplotype associated with increased susceptibility to T1D. Consistent with previous reports, D6S273*2 marked a highly predisposing DR3 haplotype in European origin, multiplex families from the USA. Furthermore, we observed on DR3 haplotypes that other D6S273 alleles were also significantly associated with both increased transmission (D6S273*5; P < 0.02) and decreased transmission (D6S273*7; P < 0.05) to affected individuals. The differential transmission was most evident among DR3-B8 haplotypes. Neither HLA-B*1801 nor any alleles of D6S273 were associated with increased T1D predisposition on DR4 haplotypes. These data indicate that multiple alleles of D6S273 mark a susceptibility locus whose effect we were able to detect only among DR3 haplotypes but not limited to DR3-B18 haplotypes. PMID- 15663751 TI - High-resolution sequence-based DPA1 typing identified two novel DPA1 alleles, DPA1*010303 and DPA1*0303, from a Kenyan population. AB - We report here two novel DPA1 alleles, DPA1*010303 and DPA1*0303, identified from a Kenyan population during sequence-based HLA-DPA1 typing. Molecular cloning and sequencing of multiple clones confirmed that one of the new DPA1 alleles is identical to DPA1*010301 at exon 2, except for a single nucleotide substitution (ACG ACC) at codon 15. The new allele has been named by the WHO Nomenclature Committee as DPA1*010303. The second novel DPA1 allele is identical to DPA1*0301, except for a single nucleotide difference (GAA GAC) at codon 28 that changed the amino acid from Glu to Asp. The new allele has been named by the WHO Nomenclature Committee as DPA1*0303. Identification of the two novel DPA1 alleles reflects the genetic diversity of this East African population. PMID- 15663752 TI - L-carnitine supplementation in dialysis patients: does the evidence justify its use? AB - Beginning January 1, 2003, the Centers for Medicare and Medicaid Services (CMS) initiated a national coverage determination for the use of levo-carnitine (L carnitine) in end-stage renal disease (ESRD) patients as a supplement for individuals defined as having a carnitine deficiency, accompanied by either erythropoietin (EPO)-resistant anemia and/or symptomatic intradialytic hypotension. The CMS criteria established for use of this supplement is minimal because EPO-resistant anemia and symptomatic intradialytic hypotension are never defined with any degree of scientific rigor. Even with the minimal criteria, CMS rejected 97% of the claims submitted over the first 6 months of the national coverage. Most importantly, scientific studies to justify use of the supplement do not exist. Based on limited scientific investigations, there appears to be no justification for the government paying for this supplement. A randomized, prospective, controlled study is needed to determine if there is any benefit from L-carnitine supplementation. Pending appropriate scientific proof, I do not believe that we should continue to waste our limited ESRD resources on a treatment of unproven benefit. PMID- 15663753 TI - The new arteriovenous fistula: the need for earlier evaluation and intervention. AB - The National Kidney Foundation's Dialysis Outcomes Quality Initiative recommends an ideal 3- to 4-month arteriovenous fistula (AVF) maturation period prior to cannulation. This recommendation may be misinterpreted by practicing nephrologists to mean that they should wait 3 months before performing a clinical assessment of the AVF. Such a delay in evaluating AVF maturity results in unnecessarily prolonged catheter use in dialysis patients whose ultimately unsatisfactory fistula could already have been diagnosed and modified at a much earlier point. Clinical assessment at 1 month can detect the common etiologies for nonmaturity, including stenosis in the vein adjacent to the anastomosis and accessory vein diversion of flow from the main fistula vein. Duplex ultrasound provides additional information in selected cases. Early evaluation allows for early fistulography for problems and potentially early cannulation for adequately maturing fistulas. Prospective trials of early AVF clinical assessment at 1 month should be performed. PMID- 15663754 TI - Phosphorus management in end-stage renal disease. AB - Chronic kidney disease is an important public health problem, with an increasing number of patients worldwide. One important outcome of renal failure is disordered mineral metabolism, most notably involving calcium and phosphorus balance. Of importance is that increased serum phosphorus levels are associated with increased mortality rates. Despite dietary restrictions, patients receiving dialysis invariably experience hyperphosphatemia and require treatment with phosphate binders. Existing phosphate binders are effective in reducing serum phosphorus levels, but are associated with a number of important disadvantages. Lanthanum carbonate, a new noncalcium, nonaluminum phosphate binder, represents a promising treatment for hyperphosphatemia. PMID- 15663755 TI - What clinical insights from the early days of dialysis are being overlooked today? PMID- 15663760 TI - Should the target hemoglobin for patients with chronic kidney disease treated with erythropoietic replacement therapy be changed? AB - Recombinant human erythropoietin (rHuEPO, epoetin) revolutionized the treatment of anemia in patients with chronic kidney disease (CKD) when it was approved for use in the United States in 1989. Among dialysis patients, the mean hemoglobin (Hb) in patients undergoing dialysis rose from 7-8 g/dl prior to 1989 to 11-12 g/dl today. Among patients with CKD not on dialysis, epoetin use has not been as broadly applied as among dialysis patients, and although the mean Hb level in this patient population has increased, the impact has been less than in patients on dialysis. The optimal treatment target for epoetin remains controversial. Consistent with clinical practice guidelines, current practice in dialysis patients in the United States aims to maintain a target Hb of 11-12 g/dl, a level that is still well below the normal range. Debate centers on whether the current Hb target is too low and whether the target range is too narrow. Quality of life clearly improves in many individuals as Hb rises into the normal range from lower levels. In retrospective studies, higher Hb levels have been associated with lower risks of hospitalization and mortality. However, one large, prospective clinical trial has raised concern about normalizing Hb in hemodialysis patients with cardiac disease, and other prospective studies have not yet provided convincing evidence of significant benefits from normalizing Hb in dialysis dependent and non-dialysis-dependent patients with CKD. A relative lack of information on non-dialysis-dependent patients with CKD and changes in fiscal policies regulating reimbursement for epoetin have contributed to uncertainty as to the best practices for anemia management in patients with CKD. There is increasing interest in the potential benefits of broadening the current target Hb range or eliminating an upper limit altogether and instead establishing a minimum Hb goal. While some extension of the upper limit of the currently recommended target Hb range might appear to be reasonable, the extent to which this should be extended, the benefits, risks, and costs of maintaining higher Hb levels in patients with CKD, and whether target Hb levels should be different in different CKD patient groups remains to be determined. Future efforts are likely to focus on selecting patient populations most likely to benefit from normalizing Hb, while adjusting the range of a subnormal Hb target for others. PMID- 15663761 TI - Determinants of small solute clearance in hemodialysis. AB - Recent outcome trials in chronic dialysis patients raise concerns about the relationship between delivered urea Kt/V and survival. Nevertheless, measurement of delivered small solute clearance remains the most common approach to quantify therapy. The purpose of this review is to provide an overview of the numerous factors influencing small solute clearance during hemodialysis. Although the focus of the review is on the manner in which dialyzer characteristics influence small solute clearances, factors related to other aspects of the extracorporeal circuit and to the patient will also be discussed. PMID- 15663763 TI - Genomic medicine, gene polymorphisms, and human biological diversity. AB - There is considerable variation in the prevalence of comorbid conditions such as malnutrition, inflammation, cardiovascular disease, and bone disease among end stage renal disease (ESRD) patients on maintenance hemodialysis. Besides traditional risk factors, these differences also arise from variability in genetic factors and gene-environment interactions. Recent publication of the human genome sequence through the Human Genome and Celera projects has provided a major impetus to the field of molecular genetics and genomic medicine. This review provides a broad overview of recent developments in this field, basic information and terminology in common use, and explores the impact of genetic variations or polymorphisms on human biological diversity. PMID- 15663764 TI - Model for gene-environment interaction: the case for dialysis. AB - Organ complications in end-stage renal disease (ESRD) such as cardiovascular disease, anemia, bone disease, malnutrition, inflammation, and infections occur in many organ systems and are caused by a multitude of underlying disease-, uremia-, and therapy-related factors, and with a wide range of manifestations and severity. Interindividual variability in the pathophysiologic response of the uremic host to environmental factors, including renal replacement therapy, may be governed to a significant degree by genetic susceptibility factors. Specific genes regulate the pathophysiologic responses of organ systems affected by ESRD and can serve as candidate genes for the host-environment interaction. This review summarizes emerging clinical and translational research work in the field of genetic polymorphism of candidate genes and their effects on the development of organ complications in ESRD. Methodologic limitations of the existing published data, the need for large prospective cohort studies, and potential future risks for the use of genomic information in resource allocation are also highlighted. PMID- 15663765 TI - Resolving controversies regarding hemodiafiltration versus hemodialysis: the Dutch Convective Transport Study. AB - Hemodialysis patients suffer from a high incidence of cardiovascular disease. Among the many predisposing factors, such as high blood pressure, dyslipidemia, and fluid overload, the accumulation of high molecular weight uremic toxins, the so-called middle molecules, may play an important role. Since convective therapies such as online hemodiafiltration have a better clearance profile for these compounds than standard hemodialysis, it has been suggested that these dialysis strategies may reduce cardiovascular morbidity and mortality. As reliable data on these issues are not available, the Dutch Convective Transport Study (CONTRAST) was recently initiated. This prospective randomized trial was designed to compare online hemodiafiltration with low-flux hemodialysis with respect to cardiovascular morbidity and mortality. PMID- 15663766 TI - National surveillance of dialysis-associated diseases in the United States, 2002. AB - In December 2002, all U.S. chronic hemodialysis centers were surveyed regarding selected patient care practices and dialysis-associated diseases. The results were compared with similar surveys conducted in previous years. In 2002, 85% of hemodialysis centers were free-standing and 81% operated for profit; the proportion of centers operating for profit has increased each year since 1985. During 1995-2002, the percentage of patients who received dialysis through central catheters increased from 13% to 26%; this trend is worrisome, as infections and antimicrobial use are higher among patients receiving dialysis through catheters. However, during the same period, the percentage of patients receiving dialysis through fistulas increased from 22% to 33%. The percentage of centers reporting one or more patients infected or colonized with vancomycin resistant enterococci (VRE) increased from 12% in 1995 to 30% in 2002. During 1997-2002, the percentage of patients vaccinated against hepatitis B virus (HBV) infection increased from 47% to 56% and the percentage of staff vaccinated increased from 87% to 90%. In 2002, routine testing for antibody to hepatitis C virus (anti-HCV) was performed on patients at 64% of centers; anti-HCV was found in 7.8% of patients. In 2001, the Centers for Disease Control (CDC) published Recommendations for Preventing Transmission of Infections among Chronic Hemodialysis Patients. Centers were surveyed regarding their awareness of the recommendations and about a variety of infection control practices. In general, the incidence of HBV and HCV was not substantially different for the infection control practices evaluated, including where staff obtain clean supplies for patient treatment, reuse of unused and unopened supplies, and practices for changing external transducer filters/protectors. However, in 2002, the incidence of HBV infection was higher among patients in centers where injectable medications were prepared on a medication cart or medication area located in the treatment area compared to a dedicated medication room. Also, those centers that used a disposable container versus a nondisposable container for priming the dialyzer had a significantly lower incidence of HCV. PMID- 15663767 TI - Extracorporeal management of valproic acid toxicity: a case report and review of the literature. AB - The incidence of intentional or accidental valproic acid (VPA) overdose is increasing. Severe VPA toxicity may lead to coma and death. Traditionally the treatment of patients with VPA toxicity has been limited to supportive measures. VPA is highly protein bound and therefore it is considered not to be removable by extracorporeal means. However, studies of VPA toxicokinetics indicate that at blood levels that exceed therapeutic concentrations, VPA protein binding sites become saturated, leading to increased concentration of the free unbound drug. The free unbound drug has a small molecular weight and therefore it is theoretically amenable to removal by extracorporeal means. We present a patient with VPA toxicity who was successfully treated with "in-series" hemodialysis and hemoperfusion followed by continuous venovenous hemodiafiltration (CVVHDF) and review the literature on the management of VPA toxicity using extracorporeal therapies. PMID- 15663770 TI - Insect glutathione transferases and insecticide resistance. AB - Glutathione transferases (GSTs) are a diverse family of enzymes found ubiquitously in aerobic organisms. They play a central role in the detoxification of both endogenous and xenobiotic compounds and are also involved in intracellular transport, biosynthesis of hormones and protection against oxidative stress. Interest in insect GSTs has primarily focused on their role in insecticide resistance. GSTs can metabolize insecticides by facilitating their reductive dehydrochlorination or by conjugation reactions with reduced glutathione, to produce water-soluble metabolites that are more readily excreted. In addition, they contribute to the removal of toxic oxygen free radical species produced through the action of pesticides. Annotation of the Anopheles gambiae and Drosophila melanogaster genomes has revealed the full extent of this enzyme family in insects. This mini review describes the insect GST enzyme family, focusing specifically on their role in conferring insecticide resistance. PMID- 15663771 TI - Gene expression of ecdysteroid-regulated gene E74 of the honeybee in ovary and brain. AB - To facilitate studies of hormonal control in the honeybee (Apis mellifera L.), a cDNA for a honeybee homologue of the ecdysteroid-regulated gene E74 (AmE74) was isolated and its expression was analysed. Northern blot analysis indicated strong expression in the adult queen abdomen, and no significant expression in the adult drone and worker abdomens. In situ hybridization demonstrated that this gene was expressed selectively in the ovary and gut in the queen abdomen. Furthermore, this gene was also expressed selectively in subsets of mushroom body interneurones in the brain of the adult worker bees. These findings suggest that AmE74 is involved in neural function as well as in reproduction in adult honeybees. PMID- 15663772 TI - piggyBac internal sequences are necessary for efficient transformation of target genomes. AB - A previously reported piggyBac minimal sequence cartridge, which is capable of efficient transposition in embryo interplasmid transposition assays, failed to produce transformants at a significant frequency in Drosophila melanogaster compared with full-length or less extensive internal deletion constructs. We have re-examined the importance of these internal domain (ID) sequences for germline transformation using a PCR strategy that effectively adds increasing lengths of ID sequences to each terminus. A series of these piggyBac ID synthetic deletion plasmids containing the 3xP3-ECFP marker gene are compared for germline transformation of D. melanogaster. Our analyses identify a minimal sequence configuration that is sufficient for movement of piggyBac vectored sequences from plasmids into the insect genome. Southern hybridizations confirm the presence of the piggyBac transposon sequences, and insertion site analyses confirm these integrations target TTAA sites. The results verify that ID sequences adjacent to the 5' and 3' terminal repeat domains are crucial for effective germline transformation with piggyBac even though they are not required for excision or interplasmid transposition. Using this information we reconstructed an inverted repeat cartridge, ITR1.1k, and a minimal piggyBac transposon vector, pXL-BacII ECFP, each of which contains these identified ID sequences in addition to the terminal repeat configuration previously described as essential for mobility. We confirm in independent experiments that these new minimal constructs yield transformation frequencies similar to the control piggyBac vector. Sequencing analyses of our constructs verify the position and the source of a point mutation within the 3' internal repeat sequence of our vectors that has no apparent effect on transformation efficiency. PMID- 15663773 TI - Gene expression profiles among immature and adult reproductive castes of the termite Reticulitermes flavipes. AB - Array-based genomic studies were conducted with the goal of identifying immature (i.e. nymph) and adult reproductive caste-biased gene expression in the termite Reticulitermes flavipes. Using cDNA macro-arrays, we identified thirty-four nymph biased genes falling into eight ontogenic categories. Based on gene expression profiles among diverse castes and developmental stages (determined by quantitative PCR), several important trends emerged. These findings highlight the importance of several developmental and survival-based factors among immature and adult termite reproductives, including: vitellogenesis, nutrient storage, juvenile hormone sequestration, ribosomal translational and filtering mechanisms, fatty acid biosynthesis, apoptosis inhibition, and both endogenous and symbiont cellulase-assisted nutrition. These findings are highly significant as they are the first to elucidate the molecular biology underlying termite reproductive caste differentiation and reproductive caste-specific biology. Other gene expression results are in agreement with previous findings that suggest roles for vitellogenin-like haemolymph proteins in soldier caste differentiation. PMID- 15663774 TI - Tissue localization and regulation by juvenile hormone of human allergen Bla g 4 from the German cockroach, Blattella germanica (L.). AB - The German cockroach, Blattella germanica (L.), produces several potent protein aeroallergens, including Bla g 4, a approximately 20 kDa lipocalin. RT-PCR, Northern analyses and in situ hybridization showed that Bla g 4 is expressed only in the adult male reproductive system. Western blotting and ELISA with rBla g 4 antiserum detected immunoreactivity in the utricles and the conglobate gland, but not in other tissues of the male reproductive system. The Bla g 4 protein content of males increased from adult emergence to day 14, but during copulation Bla g 4 was depleted in the male and transferred to the female within the spermatophore. Topical application of juvenile hormone III stimulated Bla g 4 production by both conglobate gland and utricles. PMID- 15663775 TI - The mosquito Aedes aegypti (L.) leucokinin receptor is a multiligand receptor for the three Aedes kinins. AB - A cDNA cloned from Aedes aegypti (L.) (Aedae) female Malpighian tubule (AY596453) encodes a 584 amino acid residue protein (65.2 kDa) predicted as a G protein coupled receptor and orthologue of the drosokinin receptor from Drosophila melanogaster and highly similar to the tick Boophilus microplus myokinin receptor (AF228521). Based on the similarity to this Aedes sequence, we also propose a correction for the Anopheles gambiae protein sequence EAA05450. When expressed in CHO-K1 cells, the Aedes receptor behaved as a multiligand receptor and functionally responded to concentrations > or = 1 nM of Aedae kinins 1-3, respectively, as determined by a calcium bioluminescence plate assay and single cell intracellular calcium measurements by confocal fluorescence cytometry. Estimates of EC50 values by the plate assay were 16.04 nM for Aedae-K-3, 26.6 nM for Aedae-K-2 and 48.8 nM for Aedae-K-1 and were statistically significantly different. These results suggest that the observed differences in physiological responses to the three Aedes kinins in the Aedes isolated Malpighian tubule reported elsewhere could now be explained by differences in intracellular signalling events triggered by the different peptides on the same receptor and not necessarily due to the existence of various receptors for the three Aedes kinins. PMID- 15663776 TI - Expression of Cyp6g1 and Cyp12d1 in DDT resistant and susceptible strains of Drosophila melanogaster. AB - The Rst(2)DDT locus (loci) in Drosophila is associated with the over-expression of two cytochrome P450 genes, Cyp6g1 and Cyp12d1. Using northern and western blot analysis we observed the expression pattern of these two genes in two DDT susceptible (Canton-S and 91-C) and three DDT resistant strains (Wisconsin, 91-R and Hikone-R). In Canton-S and 91-R, the CYP6G1 protein was constitutively expressed throughout development. In the Wisconsin strain, CYP6G1 was not expressed in third instar larvae unless the larvae are exposed to DDT. CYP12D1 protein was only expressed in adults. Cyp12d1 mRNA is induced in DDT resistant strains post-exposure to DDT and the expression patterns of Cyp12d1 mRNA varied across DDT resistant strains. Our data support the hypothesis that there is evolutionary plasticity in the expression patterns of P450s associated with metabolic pesticide resistance. PMID- 15663777 TI - Aedes aegypti transferrin. Gene structure, expression pattern, and regulation. AB - Mosquitoes and all other insects so far examined have an abundant haemolymph transferrin (Tsf). The exact function of these proteins has not been determined, but they may be involved in iron transport, in oogenesis and in innate immune defence against parasites and pathogens. The Tsf gene of Aedes aegypti has been cloned and sequenced. It contains a single small intron, which contrasts it to vertebrate Tsf genes that contain up to sixteen introns. The promoter region of the gene is rich in putative NF-kappaB binding sites, which is consistent with the postulated role of Tsf in insect innate immunity. Tsf message levels are very low in embryos and early larvae, but high in late larvae, pupae and adults. Western blotting experiments revealed high levels of Tsf protein in pupae and adults. Late larvae and ovaries of blood-fed mosquitoes have little intact protein, but two prominent proteolytic degradation products. These may represent biologically active peptides, as has been shown for other organisms. Tsf message is down-regulated by inorganic iron in the diet or environment, but up-regulated by a blood meal in the adult female. The up-regulation following a blood meal may, in part, be due to the decrease in juvenile hormone (JH) that is known to follow blood feeding. Treatment of blood-fed females with methoprene, an analogue of JH, resulted in decrease of the Tsf message. PMID- 15663778 TI - Molecular cloning and transcriptional activation of lysozyme-encoding cDNAs in the mosquito Aedes aegypti. AB - Lysozymes are enzymes characterized by their ability to break down bacterial cell walls. In insects certain lysozymes are only found in the midgut, whereas others are only found in the haemolymph and fat body after immune challenge. We identified two lysozyme-encoding cDNAs from Aedes aegypti. Both deduced protein sequences are basic in nature, contain 148 amino acids including eight highly conserved cysteine residues, and their genomic sequences contain a single intron. Transcriptional profiles indicated that the predominant form is constitutively expressed and up-regulated upon immune challenge and blood feeding in adult mosquitoes. The second form is expressed during early developmental stages, larvae and pupae, and at low levels in adults after immune challenge. Lysozymes in Aedes aegypti play both roles, defined by the spatial and temporal regulation of their expression. PMID- 15663780 TI - Preservation of a transgenic strain of the sawfly, Athalia rosae (Hymenoptera) by artificial fertilization using cryopreserved sperm. AB - Germline transformation using a piggyBac-derived vector is feasible in the sawfly, Athalia rosae. A previously generated transgenic line carrying green fluorescence protein (GFP) genes as reporters was successfully maintained and preserved without consecutive rearing. Sperm taken from males that were frozen directly in liquid nitrogen and stored at -80 degrees C for a year were microinjected into mature unfertilized eggs dissected from female ovaries. A fraction of the sperm-injected eggs was fertilized and developed into diploid females, and all of them expressed GFP. Haploid male progeny from these females segregated into GFP-positive and GFP-negative individuals in a ratio of 1:1 indicating heterozygosity of the parental females. The GFP genes were stably inherited staying at the location where they were originally integrated. PMID- 15663779 TI - The accumulation of specific mRNAs following multiple blood meals in Anopheles gambiae. AB - One approach to genetic control of transmission of the parasites that cause human malaria is based on expressing effector genes in mosquitoes that disable the pathogens. Endogenous mosquito promoter and other cis-acting DNA sequences are needed to direct the optimal tissue-, stage- and sex-specific expression of the effector molecules. The mRNA accumulation profiles of eight different genes expressed specifically in the midgut, salivary glands or fat body tissues of the malaria vector, Anopheles gambiae, were characterized as a measure of their suitability to direct the expression of effector molecules designed to disable specific stages of the parasites. RT-PCR techniques were used to determine the abundance of the gene products and their duration following multiple blood meals. Transcription from the midgut-expressed carboxypeptidase-encoding gene, AgCP, follows a cyclical, blood-inducible expression pattern with maximum accumulation every 3 h post blood meal. Other midgut-expressed genes encoding a trypsin and chymotrypsin, Antryp2 and Anchym1, respectively, and the fat body-expressed genes, Vg1 and Cathepsin, also show a blood-inducible pattern of expression with maximum accumulation 24 h after every blood meal. Expression of the Lipophorin gene in the fat body and apyrase and D7-related genes (AgApy and D7r2) in the salivary glands is constitutive and not significantly affected by blood meals. Promoters of the midgut- and fat body-expressed genes may lead to maximum accumulation of antiparasite effector molecule transcripts after multiple blood meals. The multiple feeding behaviour of An. gambiae thus can be an advantage to express high levels of antiparasite effector molecules to counteract the parasites throughout most of adult development. PMID- 15663781 TI - Histopathological changes in the human larynx following expanded polytetrafluroethylene (Gore-Tex(R)) implantation. AB - BACKGROUND: Expanded polytetrafluroethelyne (e PTFE, Gore-Tex(R)) has been advocated as an implant material for medialization of the vocal fold. Animal studies involving rabbits and a porcine model have demonstrated host tolerance of the implant. There have been no reports describing the histological changes in a human laryngectomy specimen with a Gore-Tex implant. CASE PRESENTATION: The histological findings in a laryngectomy specimen of a patient previously implanted with e PTFE for medialization of a paralyzed vocal fold following excision of a vagal neurofibroma were studied.Histopathology revealed a mild foreign-body giant cell granulomatous reaction with some associated fibrosis. The granulomatous response was limited to the periphery of the Gore-Tex and although it closely followed the profile of the material it did not encroach into or significantly break up the material. There was no significant neutrophilic or lymphocytic inflammation. CONCLUSIONS: Our findings are consistent with the animal models confirming that Gore-Tex implantation does not result in a significant granulomatous reaction in the human larynx over a 13-month period. Moreover, there is no evidence of resorption or infection. Further, the lack of lymphocytes in association with the granulomas indicates that there is no significant immunological hypersensitivity. Histologically, the slight permeation by connective tissue is similar to that seen in Gore-Tex vascular and cardiac implants. The degree of the slight giant cell response appears to be dependent on the profile of the material; a sharp edge incited more of a response than a flat surface. PMID- 15663782 TI - Inference of demographic history from genealogical trees using reversible jump Markov chain Monte Carlo. AB - BACKGROUND: Coalescent theory is a general framework to model genetic variation in a population. Specifically, it allows inference about population parameters from sampled DNA sequences. However, most currently employed variants of coalescent theory only consider very simple demographic scenarios of population size changes, such as exponential growth. RESULTS: Here we develop a coalescent approach that allows Bayesian non-parametric estimation of the demographic history using genealogies reconstructed from sampled DNA sequences. In this framework inference and model selection is done using reversible jump Markov chain Monte Carlo (MCMC). This method is computationally efficient and overcomes the limitations of related non-parametric approaches such as the skyline plot. We validate the approach using simulated data. Subsequently, we reanalyze HIV-1 sequence data from Central Africa and Hepatitis C virus (HCV) data from Egypt. CONCLUSIONS: The new method provides a Bayesian procedure for non-parametric estimation of the demographic history. By construction it additionally provides confidence limits and may be used jointly with other MCMC-based coalescent approaches. PMID- 15663783 TI - Chlamydia pneumoniae induces aponecrosis in human aortic smooth muscle cells. AB - BACKGROUND: The intracellular bacterium Chlamydia pneumoniae is suspected to play a role in formation and progression of atherosclerosis. Many studies investigated cell death initiation versus inhibition by Chlamydia pneumoniae in established cell lines but nothing is known in primary human aortic smooth muscle cells, a cell type among others known to be involved in the formation of the atherosclerotic plaque. Type of cell death was analyzed by various methods in primary aortic smooth muscle cells after infection with Chlamydia pneumoniae to investigate a possible pathogenic link in atherosclerosis. RESULTS: Chlamydiae were found to be localized up to 72 h post infection in aortic smooth muscle cells either as single bacteria or inside of large inclusions. Quantification of host cell death by lactate dehydrogenase release assay revealed strictly dose and time dependent lysis for all tested isolates of Chlamydia pneumoniae. Phosphatidylserine exposure was detected by flow cytometry in Chlamydia pneumoniae infected cells. Ultrastructure of Chlamydia pneumoniae infected human aortic smooth muscle cells showed extensive membrane- and organelle damage, chromatin condensation but no nuclear fragmentation. DNA fragmentation as well as cell membrane permeability was analyzed by TUNEL and NHS-biotin staining and occurred exclusively in cells carrying Chlamydia pneumoniae spots but not in smooth muscle cells with inclusions. These morphological features of cell death were not accompanied by an activation of caspase-3 as revealed by analysis of enzyme activity but involved mitochondrial membrane depolarization as shown by TMRE uptake and release of cytochrome c from mitochondria. CONCLUSION: This study provides evidence that Chlamydia pneumoniae induce a spot like infection in human aortic smooth muscle cells, which results in a chimeric cell death with both apoptotic and necrotic characteristics. This aponecrotic cell death may assist chronic inflammation in atherosclerotic blood vessels. PMID- 15663784 TI - Towards an understanding of barriers to condom use in rural Benin using the Health Belief Model: a cross sectional survey. AB - BACKGROUND: HIV/AIDS is the most dramatic epidemic of the century that has claimed over two decade more than 3 million deaths. Sub Saharan Africa is heavily affected and accounts for nearly 70% of all cases. Despite awareness campaigns, prevention measures and more recently promotion of anti viral regimens, the prevalence of cases and deaths is still rising and the prevalence of systematic condom use remains low, especially in rural areas. This study identifies barriers to condom use based on the Health Belief Model (HBM) in Benin, West Africa. METHODS: The study was a cross-sectional survey conducted from June to July 2002. Two hundred fifty one (251) individuals were interviewed using a structured questionnaire adapted from a standardized WHO/GAP questionnaire. A logistic regression was used to identify factors associated with condom use. RESULTS: In spite of satisfactory knowledge on HIV/AIDS transmission, participants are still at high risk of contracting the infection. Sixty three (63) percents of the interviewees reported being able to recognize infected people, and condom use during the last occasional intercourse was declared by only 36.8% of males and 47.5% of females. Based on the HBM, failure to use condom was related to its perceived lack of efficacy [OR = 9.76 (3.71-30.0)] and perceived quality [OR = 3.61 (1.31-9.91)]. CONCLUSIONS: This study identifies perceived efficacy (incomplete protective effect) and perceived utilization-related problem (any reported problem using condoms) as the main barriers to condom use. Hence, preventions strategies based on increasing perceived risk, perceived severity or adequate knowledge about HIV/AIDS may not be sufficient to induce condom use. These data will be useful in designing and improving HIV/AIDS prevention outreach programs in Sub Saharan Africa. PMID- 15663785 TI - Asthma is a risk factor for acute chest syndrome and cerebral vascular accidents in children with sickle cell disease. AB - BACKGROUND: Asthma and sickle cell disease are common conditions that both may result in pulmonary complications. We hypothesized that children with sickle cell disease with concomitant asthma have an increased incidence of vaso-occlusive crises that are complicated by episodes of acute chest syndrome. METHODS: A 5 year retrospective chart analysis was performed investigating 48 children ages 3 18 years with asthma and sickle cell disease and 48 children with sickle cell disease alone. Children were matched for age, gender, and type of sickle cell defect. Hospital admissions were recorded for acute chest syndrome, cerebral vascular accident, vaso-occlusive pain crises, and blood transfusions (total, exchange and chronic). Mann-Whitney test and Chi square analysis were used to assess differences between the groups. RESULTS: Children with sickle cell disease and asthma had significantly more episodes of acute chest syndrome (p = 0.03) and cerebral vascular accidents (p = 0.05) compared to children with sickle cell disease without asthma. As expected, these children received more total blood transfusions (p = 0.01) and chronic transfusions (p = 0.04). Admissions for vasoocclusive pain crises and exchange transfusions were not statistically different between cases and controls. SS disease is more severe than SC disease. CONCLUSIONS: Children with concomitant asthma and sickle cell disease have increased episodes of acute chest syndrome, cerebral vascular accidents and the need for blood transfusions. Whether aggressive asthma therapy can reduce these complications in this subset of children is unknown and requires further studies. PMID- 15663786 TI - Harnessing genomics to improve health in the Eastern Mediterranean Region - an executive course in genomics policy. AB - BACKGROUND: While innovations in medicine, science and technology have resulted in improved health and quality of life for many people, the benefits of modern medicine continue to elude millions of people in many parts of the world. To assess the potential of genomics to address health needs in EMR, the World Health Organization's Eastern Mediterranean Regional Office and the University of Toronto Joint Centre for Bioethics jointly organized a Genomics and Public Health Policy Executive Course, held September 20th-23rd, 2003, in Muscat, Oman. The 4 day course was sponsored by WHO-EMRO with additional support from the Canadian Program in Genomics and Global Health. The overall objective of the course was to collectively explore how to best harness genomics to improve health in the region. This article presents the course findings and recommendations for genomics policy in EMR. METHODS: The course brought together senior representatives from academia, biotechnology companies, regulatory bodies, media, voluntary, and legal organizations to engage in discussion. Topics covered included scientific advances in genomics, followed by innovations in business models, public sector perspectives, ethics, legal issues and national innovation systems. RESULTS: A set of recommendations, summarized below, was formulated for the Regional Office, the Member States and for individuals.* Advocacy for genomics and biotechnology for political leadership;* Networking between member states to share information, expertise, training, and regional cooperation in biotechnology; coordination of national surveys for assessment of health biotechnology innovation systems, science capacity, government policies, legislation and regulations, intellectual property policies, private sector activity;* Creation in each member country of an effective National Body on genomics, biotechnology and health to:- formulate national biotechnology strategies- raise biotechnology awareness- encourage teaching and training of biotechnology- devise integration of biotechnology within national health systems. CONCLUSION: The recommendations provide the basis for a road map for EMR to take steps to harness biotechnology for better and more equitable health. As a result of these recommendations, health ministers from the region, at the 50th Regional Committee Meeting held in October 2003, have urged Member States to establish national bodies of biotechnology to formulate a strategic vision for developing biotechnology in the service of the region's health. These efforts promise to raise the profile of genomics in EMR and increase regional cooperation in this exciting new field. PMID- 15663787 TI - Molecular models of NS3 protease variants of the Hepatitis C virus. AB - BACKGROUND: Hepatitis C virus (HCV) currently infects approximately three percent of the world population. In view of the lack of vaccines against HCV, there is an urgent need for an efficient treatment of the disease by an effective antiviral drug. Rational drug design has not been the primary way for discovering major therapeutics. Nevertheless, there are reports of success in the development of inhibitor using a structure-based approach. One of the possible targets for drug development against HCV is the NS3 protease variants. Based on the three dimensional structure of these variants we expect to identify new NS3 protease inhibitors. In order to speed up the modeling process all NS3 protease variant models were generated in a Beowulf cluster. The potential of the structural bioinformatics for development of new antiviral drugs is discussed. RESULTS: The atomic coordinates of crystallographic structure 1CU1 and 1DY9 were used as starting model for modeling of the NS3 protease variant structures. The NS3 protease variant structures are composed of six subdomains, which occur in sequence along the polypeptide chain. The protease domain exhibits the dual beta barrel fold that is common among members of the chymotrypsin serine protease family. The helicase domain contains two structurally related beta-alpha-beta subdomains and a third subdomain of seven helices and three short beta strands. The latter domain is usually referred to as the helicase alpha-helical subdomain. The rmsd value of bond lengths and bond angles, the average G-factor and Verify 3D values are presented for NS3 protease variant structures. CONCLUSIONS: This project increases the certainty that homology modeling is an useful tool in structural biology and that it can be very valuable in annotating genome sequence information and contributing to structural and functional genomics from virus. The structural models will be used to guide future efforts in the structure-based drug design of a new generation of NS3 protease variants inhibitors. All models in the database are publicly accessible via our interactive website, providing us with large amount of structural models for use in protein-ligand docking analysis. PMID- 15663788 TI - Brainstem levels of transcription factor AP-2 in rat are changed after treatment with phenelzine, but not with citalopram. AB - BACKGROUND: Before therapeutic effect is obtained after treatment with antidepressant drugs, like serotonin selective reuptake inhibitors (SSRIs), tricyclic antidepressants (TCAs) and monoamine oxidase inhibitors (MAO-Is) there is an initial lag-period of a few weeks. Neuronal adaptations on a molecular level are supposed to be involved in the initiation of the antidepressant effect. Transcription factor AP-2 is essential for neuronal development and many genes involved in the brainstem monoaminergic systems have binding sites for AP-2 in their regulatory regions. The genotype of the AP-2beta isoform has been associated with e.g. anxiety-related personality traits and with platelet MAO activity. In addition, previous studies have shown that the levels of AP-2alpha and AP-2beta in rat whole brain were decreased after 10 days of treatment with citalopram (SSRI) and imipramine (TCA), and were increased with phenelzine (MAO I). RESULTS: In the present study, we report that treatment with citalopram for 1, 7 or 21 days did not have effect on the AP-2 levels in rat brainstem. However, after treatment with phenelzine for 1, 7 or 21 days the levels of AP-2alpha and AP-2beta had increased after 7 days, but had returned to control levels at day 21. CONCLUSION: The decrease in AP-2 levels in rat whole brain previously seen after treatment with citalopram does not seem to be localised to the brainstem, it may rather occur in the monoaminergic terminal projection areas. The present data suggest that the increase in AP-2 levels previously seen in rat whole brain after subchronic treatment with phenelzine is located in the brainstem. It cannot, however, be excluded that other brain regions are involved. PMID- 15663789 TI - A robust two-way semi-linear model for normalization of cDNA microarray data. AB - BACKGROUND: Normalization is a basic step in microarray data analysis. A proper normalization procedure ensures that the intensity ratios provide meaningful measures of relative expression values. METHODS: We propose a robust semiparametric method in a two-way semi-linear model (TW-SLM) for normalization of cDNA microarray data. This method does not make the usual assumptions underlying some of the existing methods. For example, it does not assume that: (i) the percentage of differentially expressed genes is small; or (ii) the numbers of up- and down-regulated genes are about the same, as required in the LOWESS normalization method. We conduct simulation studies to evaluate the proposed method and use a real data set from a specially designed microarray experiment to compare the performance of the proposed method with that of the LOWESS normalization approach. RESULTS: The simulation results show that the proposed method performs better than the LOWESS normalization method in terms of mean square errors for estimated gene effects. The results of analysis of the real data set also show that the proposed method yields more consistent results between the direct and the indirect comparisons and also can detect more differentially expressed genes than the LOWESS method. CONCLUSIONS: Our simulation studies and the real data example indicate that the proposed robust TW SLM method works at least as well as the LOWESS method and works better when the underlying assumptions for the LOWESS method are not satisfied. Therefore, it is a powerful alternative to the existing normalization methods. PMID- 15663790 TI - Inhibition of TNFalpha in vivo prevents hyperoxia-mediated activation of caspase 3 in type II cells. AB - BACKGROUND: The mechanisms during the initial phase of oxygen toxicity leading to pulmonary tissue damage are incompletely known. Increase of tumour necrosis factor alpha (TNFalpha) represents one of the first pulmonary responses to hyperoxia. We hypothesised that, in the initial phase of hyperoxia, TNFalpha activates the caspase cascade in type II pneumocytes (TIIcells). METHODS: Lung sections or freshly isolated TIIcells of control and hyperoxic treated rats (48 hrs) were used for the determination of TNFalpha (ELISA), TNF-receptor 1 (Western blot) and activity of caspases 8, 3, and 9 (colorimetrically). NF-kappaB activation was determined by EMSA, by increase of the p65 subunit in the nuclear fraction, and by immunocytochemistry using a monoclonal anti-NF-kappaB-antibody which selectively stained the activated, nuclear form of NF-kappa B. Apoptotic markers in lung tissue sections (TUNEL) and in TIIcells (cell death detection ELISA, Bax, Bcl-2, mitochondrial membrane potential, and late and early apoptotic cells) were measured using commercially available kits. RESULTS: In vivo, hyperoxia activated NF-kappaB and increased the expression of TNFalpha, TNF receptor 1 and the activity of caspase 8 and 3 in freshly isolated TIIcells. Intratracheal application of anti-TNFalpha antibodies prevented the increase of TNFRI and of caspase 3 activity. Under hyperoxia, there was neither a significant change of cytosolic cytochrome C or of caspase 9 activity, nor an increase in apoptosis of TIIcells. Hyperoxia-induced activation of caspase 3 gradually decreased over two days of normoxia without increasing apoptosis. Therefore, activation of caspase 3 is a temporary effect in sublethal hyperoxia and did not mark the "point of no return" in TIIcells. CONCLUSION: In the initiation phase of pulmonary oxygen toxicity, an increase of TNFalpha and its receptor TNFR1 leads to the activation of caspase 8 and 3 in TIIcells. Together with the hyperoxic induced increase of Bax and the decrease of the mitochondrial membrane potential, activation of caspase 3 can be seen as sensitisation for apoptosis. Eliminating the TNFalpha effect in vivo by anti-TNFalpha antibodies prevents the pro apoptotic sensitisation of TIIcells. PMID- 15663791 TI - Immunoreactivity of the Mycobacterium avium subsp. paratuberculosis 19-kDa lipoprotein. AB - BACKGROUND: The Mycobacterium tuberculosis 19-kDa lipoprotein has been reported to stimulate both T and B cell responses as well as induce a number of Th1 cytokines. In order to evaluate the Mycobacterium avium subsp. paratuberculosis (M. avium subsp. paratuberculosis) 19-kDa lipoprotein as an immunomodulator in cattle with Johne's disease, the gene encoding the 19-kDa protein (MAP0261c) was analyzed. RESULTS: MAP0261c is conserved in mycobacteria, showing a 95% amino acid identity in M. avium subspecies avium, 84% in M. intracellulare and 76% in M. bovis and M. tuberculosis. MAP0261c was cloned, expressed, and purified as a fusion protein with the maltose-binding protein (MBP-19 kDa) in Escherichia coli. IFN-gamma production was measured from 21 naturally infected and 9 control cattle after peripheral blood mononuclear cells (PBMCs) were stimulated with a whole cell lysate (WCL) of M. avium subsp. paratuberculosis or the recombinant MBP-19 kDa. Overall, the mean response to MBP-19 kDa was not as strong as the mean response to the WCL. By comparison, cells from control, non-infected cattle did not produce IFN-gamma after stimulation with either WCL or MBP-19 kDa. To assess the humoral immune response to the 19-kDa protein, sera from cattle with clinical Johne's disease were used in immunoblot analysis. Reactivity to MBP-19 kDa protein, but not MBP alone, was observed in 9 of 14 infected cattle. Antibodies to the 19-kDa protein were not observed in 8 of 9 control cows. CONCLUSIONS: Collectively, these results demonstrate that while the 19-kDa protein from M. avium subsp. paratuberculosis stimulates a humoral immune response and weak IFN gamma production in infected cattle, the elicited responses are not strong enough to be used in a sensitive diagnostic assay. PMID- 15663792 TI - What do hospital decision-makers in Ontario, Canada, have to say about the fairness of priority setting in their institutions? AB - BACKGROUND: Priority setting, also known as rationing or resource allocation, occurs at all levels of every health care system. Daniels and Sabin have proposed a framework for priority setting in health care institutions called 'accountability for reasonableness', which links priority setting to theories of democratic deliberation. Fairness is a key goal of priority setting. According to 'accountability for reasonableness', health care institutions engaged in priority setting have a claim to fairness if they satisfy four conditions of relevance, publicity, appeals/revision, and enforcement. This is the first study which has surveyed the views of hospital decision makers throughout an entire health system about the fairness of priority setting in their institutions. The purpose of this study is to elicit hospital decision-makers' self-report of the fairness of priority setting in their hospitals using an explicit conceptual framework, 'accountability for reasonableness'. METHODS: 160 Ontario hospital Chief Executive Officers, or their designates, were asked to complete a survey questionnaire concerning priority setting in their publicly funded institutions. Eight-six Ontario hospitals completed this survey, for a response rate of 54%. Six close-ended rating scale questions (e.g. Overall, how fair is priority setting at your hospital?), and 3 open-ended questions (e.g. What do you see as the goal(s) of priority setting in your hospital?) were used. RESULTS: Overall, 60.7% of respondents indicated their hospitals' priority setting was fair. With respect to the 'accountability for reasonableness' conditions, respondents indicated their hospitals performed best for the relevance (75.0%) condition, followed by appeals/revision (56.6%), publicity (56.0%), and enforcement (39.5%). CONCLUSIONS: For the first time hospital Chief Executive Officers within an entire health system were surveyed about the fairness of priority setting practices in their institutions using the conceptual framework 'accountability for reasonableness'. Although many hospital CEOs felt that their priority setting was fair, ample room for improvement was noted, especially for the enforcement condition. PMID- 15663793 TI - Factors influencing emergency medical readmission risk in a UK district general hospital: A prospective study. AB - BACKGROUND: Over recent years increased emphasis has been given to performance monitoring of NHS hospitals, including overall number of hospital readmissions, which however are often sub-optimally adjusted for case-mix. We therefore conducted a study to examine the effect of various patient and disease factors on the risk of emergency medical readmission. METHODS: The study setting was a District General Hospital in Greater Manchester and the study period was 4.5 years. All index emergency medical admission during the study period leading to a live discharge were included in the study (n = 20,209). A multivariable proportional hazards modelling was used, based on Hospital Episodes Statistics data, to examine the influence of various baseline factors on readmission risk. Deprivation status was measured with the Townsend deprivation index score. Hazard ratios (HR) and associated 95% confidence intervals (CI) of unplanned emergency medical admission by sex, age group, admission method, diagnostic group, number of coded co-morbidities, length of stay and patient's deprivation status quartile, were calculated. RESULTS: Significant independent predictors of readmission risk at 12 months were male sex (HR 1.13, CI: 1.07-1.2), age (age >75 (HR 1.57, CI 1.45-1.7), number of coded co-morbidities (HR for >4 coded co morbidities: 1.49 CI: 1.26-1.76), admission via GP referral (HR 0.93, CI 0.88 0.99) and primary diagnosis of heart failure (HR 1.33, CI: 1.16-1.53) and chronic obstructive pulmonary disease/asthma (HR 1.34, CI: 1.21-1.48). Higher level of deprivation was also significantly and independently associated and with increased emergency medical readmission risk at three (HR for the most deprived quartile 1.21, CI: 1.08-1.35), six (HR 1.21, CI: 1.1-1.33) and twelve months (HR 1.25, CI: 1.16-1.36). CONCLUSIONS: There is a potential for improving health and reducing demand for emergency medical admissions with more effective management of patients with heart failure and chronic obstructive airways disease/asthma. There is also a potential for improving health and reducing demand if reasons for increased readmission risk in more deprived patients are understood. The potential influence of deprivation status on readmission risk should be acknowledged, and NHS performance indicators adjustment for deprivation case-mix would be prudent. PMID- 15663794 TI - The absence of reactive oxygen species production protects mice against bleomycin induced pulmonary fibrosis. AB - BACKGROUND: Reactive oxygen species and tissue remodeling regulators, such as metalloproteinases (MMPs) and their inhibitors (TIMPs), are thought to be involved in the development of pulmonary fibrosis. We investigated these factors in the fibrotic response to bleomycin of p47phox -/- (KO) mice, deficient for ROS production through the NADPH-oxidase pathway. METHODS: Mice are administered by intranasal instillation of 0.1 mg bleomycin. Either 24 h or 14 days after, mice were anesthetized and underwent either bronchoalveolar lavage (BAL) or lung removal. RESULTS: BAL cells from bleomycin treated WT mice showed enhanced ROS production after PMA stimulation, whereas no change was observed with BAL cells from p47phox -/- mice. At day 1, the bleomycin-induced acute inflammatory response (increased neutrophil count and MMP-9 activity in the BAL fluid) was strikingly greater in KO than wild-type (WT) mice, while IL-6 levels increased significantly more in the latter. Hydroxyproline assays in the lung tissue 14 days after bleomycin administration revealed the absence of collagen deposition in the lungs of the KO mice, which had significantly lower hydroxyproline levels than the WT mice. The MMP-9/TIMP-1 ratio did not change at day 1 after bleomycin administration in WT mice, but increased significantly in the KO mice. By day 14, the ratio fell significantly from baseline in both strains, but more in the WT than KO strains. CONCLUSIONS: These results suggest that NADPH-oxidase-derived ROS are essential to the development of pulmonary fibrosis. The absence of collagen deposition in KO mice seems to be associated with an elevated MMP-9/TIMP 1 ratio in the lungs. This finding highlights the importance of metalloproteinases and protease/anti-protease imbalances in pulmonary fibrosis. PMID- 15663795 TI - An online operational rainfall-monitoring resource for epidemic malaria early warning systems in Africa. AB - Periodic epidemics of malaria are a major public health problem for many sub Saharan African countries. Populations in epidemic prone areas have a poorly developed immunity to malaria and the disease remains life threatening to all age groups. The impact of epidemics could be minimized by prediction and improved prevention through timely vector control and deployment of appropriate drugs. Malaria Early Warning Systems are advocated as a means of improving the opportunity for preparedness and timely response. Rainfall is one of the major factors triggering epidemics in warm semi-arid and desert-fringe areas. Explosive epidemics often occur in these regions after excessive rains and, where these follow periods of drought and poor food security, can be especially severe. Consequently, rainfall monitoring forms one of the essential elements for the development of integrated Malaria Early Warning Systems for sub-Saharan Africa, as outlined by the World Health Organization. The Roll Back Malaria Technical Resource Network on Prevention and Control of Epidemics recommended that a simple indicator of changes in epidemic risk in regions of marginal transmission, consisting primarily of rainfall anomaly maps, could provide immediate benefit to early warning efforts. In response to these recommendations, the Famine Early Warning Systems Network produced maps that combine information about dekadal rainfall anomalies, and epidemic malaria risk, available via their Africa Data Dissemination Service. These maps were later made available in a format that is directly compatible with HealthMapper, the mapping and surveillance software developed by the WHO's Communicable Disease Surveillance and Response Department. A new monitoring interface has recently been developed at the International Research Institute for Climate Prediction (IRI) that enables the user to gain a more contextual perspective of the current rainfall estimates by comparing them to previous seasons and climatological averages. These resources are available at no cost to the user and are updated on a routine basis. PMID- 15663796 TI - Large scale hierarchical clustering of protein sequences. AB - BACKGROUND: Searching a biological sequence database with a query sequence looking for homologues has become a routine operation in computational biology. In spite of the high degree of sophistication of currently available search routines it is still virtually impossible to identify quickly and clearly a group of sequences that a given query sequence belongs to. RESULTS: We report on our developments in grouping all known protein sequences hierarchically into superfamily and family clusters. Our graph-based algorithms take into account the topology of the sequence space induced by the data itself to construct a biologically meaningful partitioning. We have applied our clustering procedures to a non-redundant set of about 1,000,000 sequences resulting in a hierarchical clustering which is being made available for querying and browsing at http://systers.molgen.mpg.de/. CONCLUSIONS: Comparisons with other widely used clustering methods on various data sets show the abilities and strengths of our clustering methods in producing a biologically meaningful grouping of protein sequences. PMID- 15663797 TI - Genetics and geometry of canalization and developmental stability in Drosophila subobscura. AB - BACKGROUND: Many properties of organisms show great robustness against genetic and environmental perturbations. The terms canalization and developmental stability were originally proposed to describe the ability of an organism to resist perturbations and to produce a predictable target phenotype regardless of random developmental noise. However, the extent to which canalization and developmental stability are controlled by the same set of genes and share underlying regulatory mechanisms is largely unresolved. RESULTS: We have analyzed the effects of clinical genetic variation (inversion polymorphism) on wing asymmetry by applying the methods of geometric morphometrics in the context of quantitative genetics using isochromosomal lines of Drosophila subobscura. For the analysis of overall size, developmental stability was positively correlated with levels of heterozygosity and development at the optimal temperature. For analyses of shape, the overall comparisons by matrix correlations indicate that inter- and intraindividual variation levels were poorly correlated, a result also supported when comparing the vectors describing patterns of variation of landmark position. The lack of similarity was basically due to the discrepancy between the genetic and environmental components of the interindividual variation. Finally, the analyses have also underscored the presence of genetic variation for directional asymmetry. CONCLUSIONS: The results strongly support the hypothesis that environmental canalization and developmental stability share underlying regulatory mechanisms, but environmental and genetic canalization are not functionally the same. A likely explanation for this lack of association is that natural wing shape variation in Drosophila populations is loosely related to individual fitness. PMID- 15663798 TI - Genetical and functional investigation of fliC genes encoding flagellar serotype H4 in wildtype strains of Escherichia coli and in a laboratory E. coli K-12 strain expressing flagellar antigen type H48. AB - BACKGROUND: Serotyping of O-(lipopolysaccharide) and H-(flagellar) antigens is a wideley used method for identification of pathogenic strains and clones of Escherichia coli. At present, 176 O- and 53 H-antigens are described for E. coli which occur in different combinations in the strains. The flagellar antigen H4 is widely present in E. coli strains of different O-serotypes and pathotypes and we have investigated the genetic relationship between H4 encoding fliC genes by PCR, nucleotide sequencing and expression studies. RESULTS: The complete nucleotide sequence of fliC genes present in E. coli reference strains U9-41 (O2:K1:H4) and P12b (O15:H17) was determined and both were found 99.3% (1043 of 1050 nucleotides) identical in their coding sequence. A PCR/RFLP protocol was developed for typing of fliC-H4 strains and 88 E. coli strains reacting with H4 antiserum were investigated. Nucleotide sequencing of complete fliC genes of six E. coli strains which were selected based on serum agglutination titers, fliC-PCR genotyping and reference data revealed 96.6 to 100% identity on the amino acid level. The functional expression of flagellin encoded by fliC-H4 from strain U9 41 and from our strain P12b which is an H4 expressing variant type was investigated in the E. coli K-12 strain JM109 which encodes flagellar type H48. The fliC recombinant plasmid carrying JM109 strains reacted with both H4 and H48 specific antisera whereas JM109 reacted only with the H48 antiserum. By immunoelectron microscopy, we could show that the flagella made by the fliC-H4 recombinant plasmid carrying strain are constituted of H48 and H4 flagellins which are co-assembled into functional flagella. CONCLUSION: The flagellar serotype H4 is encoded by closely related fliC genes present in serologically different types of E. coli strains which were isolated at different time periods and geographical locations. Our expression studies show for the first time, that flagellins of different molecular weigh are functionally expressed and coassembled in the same flagellar filament in E. coli. PMID- 15663802 TI - Which patients with prostate cancer are actually candidates for hormone therapy? AB - In this article, we will try to address the following aspects: which factors are responsible of the introduction of new candidates for hormone therapy in prostate cancer, who are actually candidates for hormone therapy, classifying them on the basis of the stage of the disease, and which treatment modalities can be proposed for each candidate. Since the introduction of hormone therapy for the treatment of prostate cancer, there has been a debate about the optimal timing of hormone therapy. A modification in the timing of hormone therapy produced new candidates for hormone manipulation. In particular, the use of hormone treatment for younger patients, longer periods and early prostate cancer, absolutely requires a whole re-evaluation of which therapy is indicated and it may produce new problems such as higher risk of over-treatment, need of a better evaluation of quality of life in younger patients and the research for better tolerated therapies. Therapies that resist for longer periods without the production of a hormone-refractory disease are also required. PMID- 15663803 TI - Interobserver variation of prostatic volume estimation with digital rectal examination by urological staffs with different experiences. AB - PURPOSE: To compare the accuracy of estimating prostatic volume with digital rectal examination (DRE) by urological staffs with different experiences. Measurement of prostatic volume with transrectal ultrasonography (TRUS) serves as the reference standard. MATERIALS AND METHODS: Thirty-nine consecutive male patients admitted with acute urinary retention had their prostatic volume estimated with DRE by a urology junior trainee, a urology higher trainee and a trained urologist. All patients had TRUS to measure their prostatic volumes. Pearson correlation coefficients (r) were used to assess the relationships between the prostatic volume measured with TRUS and that estimated with DRE by the 3 urological staffs. Wilcoxon signed ranks tests were used to compare the discrepancies between the prostatic volume measured with TRUS and that estimated with DRE for the 3 Urological staffs, and to assess the inter-observer differences of these discrepancies. RESULTS: The correlation coefficients for the 3 urological staffs were r = 0.573 for the urology junior trainee, r = 0.541 for the urology higher trainee, and r = 0.640 for the trained urologist. The median discrepancies between the prostatic volume measured with TRUS and that estimated with DRE were -9.1 mL for the urology junior trainee, -1.3 mL for the urology higher trainee and 0.9 mL for the trained urologist. These discrepancies were statistically significant only in the case of urology junior trainee (p = 0.015, Wilcoxon signed ranks test). The difference in these discrepancies was statistically significant only between the urology junior trainee and the trained urologist (p = 0.003, Wilcoxon signed ranks test). CONCLUSIONS: The trained urologist was more accurate in estimating prostatic volume with DRE than the urology junior trainee. PMID- 15663804 TI - Analysis of risk factors of involvement of seminal vesicles in patients with prostate cancer undergoing radical prostatectomy. AB - OBJECTIVE: To determine through preoperative serum PSA level, Gleason score on biopsy and percentage of fragments affected by tumor on biopsy, the probability of involvement of the seminal vesicles. MATERIALS AND METHODS: During the period between March 1991 to December 2002, we selected 899 patients undergoing radical prostatectomy for treatment of localized prostate adenocarcinoma. The analyzed preoperative variables were PSA, percentage of positive fragments and Gleason score on the biopsy. Pre-operative PSA was divided in scales from 0 to 4.0 ng/mL, 4.1 to 10 ng/mL, 10.1 to 20 ng/mL and > 20 ng/mL, Gleason score was categorized in scales from 2 to 6. 7 and 8 to 10, and the percentage of affected fragments was divided in 0 to 25%, 25.1% to 50%, 50.1% to 75%, and 75.1% to 100%. All these variables were correlated with the involvement of seminal vesicles in the surgical specimen. RESULTS: Of the 899 patients under study, approximately 11% (95% CI, [9% - 13%]) had involvement of seminal vesicles. On the multivariate analysis, when PSA was < or = 4, the Gleason score was 2 to 6, and less than 25% of fragments were involved on the biopsy, only 3.6%, 7.6% and 6.2% of patients respectively, had involvement of seminal vesicles. On the multivariate analysis, we observed that PSA, Gleason score and the percentage of involved fragments were independent prognostic factors for invasion of seminal vesicles. CONCLUSION: The preoperative variables used in the present study allow the identification of men with minimal risk (lower than 5%) if involvement of seminal vesicles. PMID- 15663805 TI - Concurrent validity, internal consistency and responsiveness of the Portuguese version of the King's Health Questionnaire (KHQ) in women after stress urinary incontinence surgery. AB - OBJECTIVE: To evaluate the concurrent validity, internal consistency and responsiveness of King's Health Questionnaire (KHQ) in patients who underwent sling procedures for the treatment of stress urinary incontinence. MATERIALS AND METHODS: We performed a prospective open label multicenter study in 4 tertiary referral centers. Sixty-eight female patients were enrolled with urodynamically diagnosed urinary stress incontinence. Patients were treated using surgical procedures, mostly (73%) with the synthetic sling procedure, which has been considered one of the gold standard methods for the treatment of urinary incontinence. The patients were assessed before and after one month of postoperative follow up, using the KHQ in its validated Portuguese version. Patients also underwent preoperative urodynamic test, Stamey incontinence grading, pad usage and the assessment of number of pads used per day. After surgery, patients underwent stress test, Stamey incontinence grading pad usage and the assessment of number of pads used per day. RESULTS: The concurrent validity showed good correlations in some domains of KHQ to clinical parameters. The internal consistency was higher after treatment compared to preoperative values. Objective parameters, such as pad usage and the assessment of number of pads used per day, had significant correlation with changes in post-treatment scores on KHQ. The responsiveness expressed in terms of standardized effect size (SES) and standardized response mean (SRM) was large. CONCLUSION: The results showed moderate concurrent validity, strong internal consistency and high responsiveness for KHQ, indicating that it is suitable for measuring outcomes in clinical trials among female patients with stress urinary incontinence. PMID- 15663806 TI - Feasibility of refreezing human spermatozoa through the technique of liquid nitrogen vapor. AB - OBJECTIVE: To assess the feasibility of refreezing human semen using the technique of liquid nitrogen vapor with static phases. MATERIALS AND METHODS: Twenty samples from 16 subjects who required disposal of their cryopreserved semen were thawed, corresponding to 6 cancer patients and 10 participants in the assisted reproduction (AR) program. Samples were refrozen using the technique of liquid nitrogen vapor with static phases, identical to the one used for the initial freezing, and thawed again after 72 hours. We assessed the concentration of motile spermatozoa, total and progressive percent motility and spermatic vitality, according to criteria of the World Health Organization (WHO), as well as spermatic morphology according to the strict Kruger criterion, after the first and after the second thawing. RESULTS: We observed a significant decrease in all the parameters evaluated between the first and the second thawing. Median values for the concentration of motile spermatozoa decreased from 2.0x10(6)/mL to 0.1x10(6)/mL (p < 0.01); total percent motility from 42% to 22.5% (p < 0.01); progressive percent motility from 34% to 9.5% (p < 0.01); vitality from 45% to 20% (p < 0.01); and morphology from 5% to 5% (p = 0.03). There was no significant difference in the spermatic parameters between the cancer and assisted reproduction groups, both after the first and after the second thawing. We observed that in 100% of cases there was retrieval of motile spermatozoa after the second thawing. CONCLUSION: Refreezing of human semen by the technique of liquid nitrogen vapor allows the retrieval of viable spermatozoa after thawing. PMID- 15663807 TI - Cholesteatoma of the upper urinary tract. AB - We report the case of a 57-year old patient with complex cystic image in right kidney. Following radical nephrectomy, the pathological study established the diagnosis of renal cholesteatoma. We discuss the frequency, pathogenesis, clinical presentation, propedeutics, histological findings and proposes for intervention observed in the literature. PMID- 15663809 TI - Non-Hodgkin lymphoma of the bladder. AB - Lymphomas of the bladder are rare lesions, representing approximately 0.2% of the primary neoplastic lesions and approximately 1.8% of the secondary lesions in this organ. The authors report the case of a 41-year old patient with secondary lymphoma of the bladder occurring 2 years after treatment for non-Hodgkin lymphoma, diagnosed by biopsy of cervical lymph node, and analyze the clinical and prognostic aspects of bladder lymphomas. PMID- 15663808 TI - Ureteral endometriosis and coexistent urethral leiomyoma in a postmenopausal woman. AB - We report the case of a postmenopausal woman with a synchronous obstructing intrinsic endometrioma of the left ureter and a coexistent periurethral leiomyoma. Endometriosis in postmenopausal women is a rare clinical entity usually associated with exogenous estrogen use. Urethral leiomyomas are also rare, with only 40 cases reported in the literature. Ovarian hormones are believed to influence the growth of leiomyomas. We report the genitourinary presentation of 2 separate disease entities with known hormonal influence in a postmenopausal woman receiving estrogen replacement therapy. We believe the patient's hormonal milieu affected the development of her concurrent pathology. PMID- 15663810 TI - Testicular schistosomiasis mimicking tumour. AB - Schistosomiasis or bilharziasis is a disease caused by Schistosoma. When infecting men the most common parasites are Schistosoma mansoni, Schistosoma japonicum and Schistosoma haematobium. The Schistosoma mansoni is the only endemic parasite in Brazil. We present a case of testicular schistosomiasis simulating malignancy. The case was treated successfully by excisional biopsy and praziquantel therapy. A review of the literature is discussed. PMID- 15663811 TI - Findings in cystourethrography that suggest lower urinary tract dysfunction in children with vesicoureteral reflux. AB - PURPOSE: Children with lower urinary tract dysfunction and vesicoureteral reflux, at cystography assessment, frequently present alterations in the lower urinary tract anatomy such as dilated posterior urethra, irregularity of the bladder wall and diverticula. However, the significance of these findings is unknown. The objective of this study is to evaluate the incidence of these findings, their time of disappearance and their correlation with the severity of the reflux. MATERIALS AND METHODS: 193 children with vesicoureteral reflux, considered simple, in the age group above 5 years at the moment of diagnosis, were analyzed. The recommendation for follow-up of these patients was one voiding cystoureterography (VCUG) each year. Only patients with a minimum of 2 VCUGs performed in a period of at least 6 months were considered. The VCUGs were classified as positive and negative in relation to findings that were characteristic of lower urinary tract dysfunction (LUTD). RESULTS: From the 193 children analyzed, 50 (26%) presented positive VCUG and 143 negative VCUG. From the patients without symptoms of lower urinary tract dysfunction (n = 135), 12 (9%) presented positive VCUG and 123 (91%) a negative VCUG. From the patients with negative VCUG, 68 (48%) presented unilateral reflux and 75 (52%) presented bilateral reflux. From those with positive VCUG, 26 (52%) had unilateral reflux and 24 bilateral reflux (48%). This difference was not statistically significant. A higher incidence of grade II reflux was more evident in patients with negative VCUG and degree III in patients with positive VCUG (p < 0.05). CONCLUSIONS: Our study demonstrated that 64% of the patients with LUTD and reflux presented findings in the VCUG that suggest dysfunction. PMID- 15663812 TI - Adriamycin-induced fetal hydronephrosis. AB - INTRODUCTION: At the end of pregnancy, the amniotic fluid (AF) depends basically on renal function, corresponding to fetal urine. Changes in AF, especially oligohydramnios, are reported in association with fetal hydronephrosis (FH). The experimental model using adriamycin in pregnant female rats has a teratogenic effect and has been classically employed to study esophageal atresia. Nevertheless, adriamycin promotes FH with high frequency as well. In the present study, using this animal model, we tried to identify the incidence and microscopic changes of FH, as well as its correlation with AF weight. MATERIALS AND METHODS: Eight Spreague-Dawley pregnant female rats received adriamycin 2.2 mg/kg on the 8th and 9th gestational days (considering term gestation = 22 days). Those fetuses that received adriamycin (Adriamycin Group) were compared with fetuses from 2 female rats (Control Group), which received 0.9% saline solution. On the 21.5 gestational day, the fetuses were collected by cesarean incision, sacrificed, and examined for macro and microscopic changes in kidneys and ureters. Fetuses with bilateral hydronephrosis formed the Hydronephrosis Group. AF weight was determined as well. RESULTS: Hydronephrosis occurred in 70 (95%) of the 74 fetuses in the adriamycin group against none of the 21 fetuses from the control group. The amniotic fluid weight was increased in the adriamycin group in relation to the control group (p < 0.001). The histomorphometric study revealed dilation of the renal pelvis and reduction of renal parenchyma in the hydronephrosis group in relation to the control group. Severe cortical atrophy, cortical tubular atrophy and medullar atrophy were observed in the hydronephrosis group. CONCLUSIONS: Slight renal lesions were in agreement with changes in AF weight, since they suggest that there was production of urine with the maintenance of AF. PMID- 15663830 TI - The development of the FOLFOX regimens as a treatment standard of advanced colorectal cancer. PMID- 15663831 TI - Evolution of FOLFOX regimens in the treatment of advanced colorectal cancer. PMID- 15663833 TI - Staging of colorectal cancer: past, present, and future. AB - Since the inception of staging for colorectal cancer (CRC), anatomic extent of disease has provided a solid foundation for predicting prognosis and guiding treatment in patients with CRC. However, the rapid advances in biologic and genetic technology have provided many new possibilities for enhancing the predictive power of the staging system. In this article, we will briefly discuss the history of CRC staging, identify the current parameters for American Joint Committee on Cancer staging including recent changes, and offer guidelines for future staging modifications. Although anatomic parameters have served us well for many years, their continued use as sole contributors to CRC staging is questionable. Staging in the future is likely to be a complicated combination of anatomic and biologic markers used together in mathematic modeling to group patients into prognostic risk groups, which can be used to guide tailored therapy. PMID- 15663832 TI - The addition of bevacizumab to FOLFOX4 prolongs survival in relapsed colorectal cancer: interim data from the ECOG 3200 trial. PMID- 15663834 TI - Colorectal cancer in patients with inflammatory bowel disease. AB - Patients with inflammatory bowel disease (IBD) are subject to increased risks for the development of colorectal cancer (CRC), risks that are attributed to the duration and anatomic extent of disease in patients with ulcerative colitis and Crohn's disease. Although IBD contributes only 1%-2% to all cases of CRC, the mortality rate in patients with a diagnosis of CRC in the setting of IBD is higher than for those afflicted with sporadic cases of CRC. Given the length of time from IBD onset to the development of CRC, surveillance continues to be widely practiced. Although still under development, novel techniques for the earlier detection of dysplastic lesions have moved to the forefront in an attempt to optimize surveillance strategies and decrease the risk of CRC development. PMID- 15663835 TI - Lymphatic mapping and sentinel node analysis in colon cancer. AB - Accurate staging of colon cancer is prognostically and therapeutically important. By identifying those patients who would most benefit from adjuvant chemotherapy, accurate staging should decrease recurrence rates and improve overall survival. Lymphatic mapping and sentinel node analysis allow for a focused review of the lymph nodes, which are most likely to harbor a metastasis and may make ultrastaging techniques, such as immunohistochemistry and reverse-transcriptase polymerase chain reaction, more practical. The prognostic significance of micrometastatic disease detected via ultrastaging techniques remains controversial. PMID- 15663836 TI - 5-Fluorouracil-based chemotherapy for advanced colorectal cancer in elderly patients: a north central cancer treatment group study. AB - Although 5-fluorouracil (5-FU)-based chemotherapy is commonly used in patients with advanced colorectal cancer (CRC), little data exist on the tolerability and benefit of therapy in elderly patients. To compare toxicity, dose intensity, response rate, time to tumor progression, and overall survival for older and younger patients, we conducted a pooled analysis of 1748 patients, divided into 4 quartile-based age groups, from 4 North Central Cancer Treatment Group trials testing 5-FU with or without leucovorin for advanced CRC. Patients aged > 65 years had modestly higher rates of severe toxicity (grade >/= 3) overall (53% vs. 46%) and higher rates of diarrhea (21% vs. 16%), stomatitis (17% vs. 13%), and infection (4% vs. 2%). Toxicity rates were similar between patients aged 66-70 years and patients aged > 70 years. The response rate did not differ by age group (2-sided; P = 0.90); it was significantly lower for patients with higher performance status scores (30% for score of 0/1; 17% for 2/3; 2-sided; P = 0.001). Performance status, not age, was predictive of time to tumor progression and overall survival. The older patients with CRC treated with 5-FU have modestly higher rates of severe toxicity, mainly diarrhea and stomatitis. Supportive measures to control diarrhea and stomatitis may be particularly important in elderly patients. Age alone should not be used to determine whether older patients are treated, because performance status is predictive of dose intensity, response rate, time to tumor progression, and overall survival. PMID- 15663837 TI - Long-term results of local excision with and without chemoradiation for adenocarcinoma of the rectum. AB - The role of local excision for rectal carcinoma remains controversial. We reviewed 285 patients undergoing curative resection for rectal cancer between 1984 and 2001. Surgical procedures were local excision (LE; n = 49), abdominoperineal resection (APR; n = 124), and low anterior resection (LAR; n = 112). Median follow-up for all patients was 6.2 years. For patients undergoing local excision, postoperative tumor stages were Tis (22%), T1 (41%), T2 (18%), and T3 (18%). Twelve patients received postoperative radiation >/= 45 Gy, and 4 patients received adjuvant chemotherapy. Of the 49 patients who underwent LE, the 5- and 10-year overall survival rates were 76% and 42%, respectively. The 5- and 10-year disease-free survival rates were 69% and 58%, respectively. The incidence of local recurrence was 16% and the incidence of distant recurrence was 6%. For the 11 patients who experienced disease recurrence, the median time to recurrence was 13 months (range, 1-59 months). Of the 8 patients who developed local recurrence, 4 refused salvage treatment, 2 underwent salvage APR, and 2 underwent repeat excision. Of the 4 who underwent salvage surgery, one is alive with no evidence of disease, one developed distant disease, and 2 died with unknown disease status. Adjuvant therapy did not affect survival or recurrence rates in patients undergoing LE compared with other surgeries. The rate of local failure (16%) is comparable to that observed in the Cancer and Leukemia Group B (CALGB) 8984 prospective study and suggests that highly selected patients undergoing local excision can expect good local control of rectal cancer. PMID- 15663838 TI - Synergistic antitumor activity of capecitabine in combination with irinotecan. AB - 5-Fluorouracil (5-FU) and capecitabine alone and in combination with irinotecan/oxaliplatin are clinically active in the treatment of colorectal and other solid tumors. Studies of the antitumor activity and toxicity of capecitabine or irinotecan alone and in combination with each other, were compared with 5-FU and raltitrexed in human tumor xenografts of colorectal and squamous cell carcinoma of the head and neck using clinically relevant schedules. Antitumor activity and toxicity were evaluated in nude mice bearing human colon carcinomas of HCT-8 and HT-29 and in head and neck squamous cell carcinomas of A253 and FaDu xenografts using the maximum tolerable dose of single-agent capecitabine, 5-FU, or raltitrexed, or each of the drugs in combination with irinotecan. Mice were treated with capecitabine and irinotecan alone or in combination using 2 different schedules: (1) capecitabine orally once a day for 7 days and a single dose of irinotecan (50 mg/kg intravenously [I.V.]), with each drug alone or in combination, and (2) capecitabine orally 5 days a week for 3 weeks and irinotecan 50 mg/kg (I.V. injection) once a week for 3 weeks, with each drug alone or in combination. For comparative purposes, the antitumor activity of single-agent capecitabine, 5-FU, or raltitrexed, or each drug in combination with irinotecan was carried out at its maximum tolerated dose (MTD) using a 3-week schedule. Results indicated that HT-29 and A253 xenografts were de novo resistant (no cure) to capecitabine and irinotecan alone at the MTD, whereas HCT-8 and FaDu xenografts were relatively more sensitive, yielding 10%-20% cures. The combination of irinotecan/capecitabine was much more active than either drug alone against all 4 tumor models. The cure rates were increased from 0 to 20% in A253 and HT-29 xenografts and from 10%-20% to 80%-100% in HCT-8 and FaDu tumor xenografts, respectively. Irinotecan/capecitabine had clear advantage over irinotecan/5-FU and irinotecan/raltitrexed in efficacy and selectivity in that they were more active and less toxic. The extent of synergy with irinotecan/capecitabine appears to be tumor-dependent and independent of the status of p53 expression. The potential impact of the preclinical results on clinical practice for the use of these drugs in combination needs clinical validation. PMID- 15663839 TI - Importance of preclinical investigations of the integration of capecitabine into polychemotherapy regimens. PMID- 15663840 TI - Wildlife as source of zoonotic infections. AB - Zoonoses with a wildlife reservoir represent a major public health problem, affecting all continents. Hundreds of pathogens and many different transmission modes are involved, and many factors influence the epidemiology of the various zoonoses. The importance and recognition of wildlife as a reservoir of zoonoses are increasing. Cost-effective prevention and control of these zoonoses necessitate an interdisciplinary and holistic approach and international cooperation. Surveillance, laboratory capability, research, training and education, and communication are key elements. PMID- 15663841 TI - Potential mammalian filovirus reservoirs. AB - Ebola and Marburg viruses are maintained in unknown reservoir species; spillover into human populations results in occasional human cases or epidemics. We attempted to narrow the list of possibilities regarding the identity of those reservoir species. We made a series of explicit assumptions about the reservoir: it is a mammal; it supports persistent, largely asymptomatic filovirus infections; its range subsumes that of its associated filovirus; it has coevolved with the virus; it is of small body size; and it is not a species that is commensal with humans. Under these assumptions, we developed priority lists of mammal clades that coincide distributionally with filovirus outbreak distributions and compared these lists with those mammal taxa that have been tested for filovirus infection in previous epidemiologic studies. Studying the remainder of these taxa may be a fruitful avenue for pursuing the identity of natural reservoirs of filoviruses. PMID- 15663842 TI - Nipah virus encephalitis reemergence, Bangladesh. AB - We retrospectively investigated two outbreaks of encephalitis in Meherpur and Naogaon, Bangladesh, which occurred in 2001 and 2003. We collected serum samples from persons who were ill, their household contacts, randomly selected residents, hospital workers, and various animals. Cases were classified as laboratory confirmed or probable. We identified 13 cases (4 confirmed, 9 probable) in Meherpur; 7 were in persons in two households. Patients were more likely than nonpatients to have close contact with other patients or have contact with a sick cow. In Naogaon, we identified 12 cases (4 confirmed, 8 probable); 7 were in persons clustered in 2 households. Two Pteropus bats had antibodies for Nipah virus. Samples from hospital workers were negative for Nipah virus antibodies. These outbreaks, the first since 1999, suggest that transmission may occur through close contact with other patients or from exposure to a common source. Surveillance and enhancement of diagnostic capacity to detect Nipah virus infection are recommended. PMID- 15663843 TI - Risk factors for alveolar echinococcosis in humans. AB - We conducted a case-control study to investigate risk factors for acquiring autochthonous alveolar echinococcosis in Germany. Forty cases and 120 controls matched by age and residence were interviewed. Patients were more likely than controls to have owned dogs that killed game (odds ratio [OR] = 18.0), lived in a farmhouse (OR = 6.4), owned dogs that roamed outdoors unattended (OR = 6.1), collected wood (OR = 4.7), been farmers (OR = 4.7), chewed grass (OR = 4.4), lived in a dwelling close to fields (OR = 3.0), gone into forests for vocational reasons (OR = 2.8), grown leaf or root vegetables (OR = 2.5), owned cats that roamed outdoors unattended (OR = 2.3), and eaten unwashed strawberries (OR = 2.2). Sixty-five percent of cases were attributable to farming. Measures that prevent accidental swallowing of possibly contaminated material during farming or adequate deworming of pet animals might reduce the risk for alveolar echinococcosis. PMID- 15663844 TI - Exposure to nonhuman primates in rural Cameroon. AB - Exposure to nonhuman primates has led to the emergence of important diseases, including Ebola hemorrhagic fever, AIDS, and adult T-cell leukemia. To determine the extent of exposure to nonhuman primates, persons were examined in 17 remote villages in Cameroon that represented three habitats (savanna, gallery forest, and lowland forest). Questionnaire data were collected to assess whether persons kept wild animal pets; hunted and butchered wild game; had experienced bites, scratches, or injuries from live animals; or had been injured during hunting or butchering. While all villages had substantial exposure to nonhuman primates, higher rates of exposure were seen in lowland forest sites. The study demonstrates that exposure is not limited to small groups of hunters. A high percentage of rural villagers report exposure to nonhuman primate blood and body fluids and risk acquiring infectious diseases. PMID- 15663845 TI - Origin of the amphibian chytrid fungus. AB - The sudden appearance of chytridiomycosis, the cause of amphibian deaths and population declines in several continents, suggests that its etiologic agent, the amphibian chytrid Batrachochytrium dendrobatidis, was introduced into the affected regions. However, the origin of this virulent pathogen is unknown. A survey was conducted of 697 archived specimens of 3 species of Xenopus collected from 1879 to 1999 in southern Africa in which the histologic features of the interdigital webbing were analyzed. The earliest case of chytridiomycosis found was in a Xenopus laevis frog in 1938, and overall prevalence was 2.7%. The prevalence showed no significant differences between species, regions, season, or time period. Chytridiomycosis was a stable endemic infection in southern Africa for 23 years before any positive specimen was found outside Africa. We propose that Africa is the origin of the amphibian chytrid and that the international trade in X. laevis that began in the mid-1930s was the means of dissemination. PMID- 15663846 TI - Nonsusceptibility of primate cells to Taura syndrome virus. AB - Taura syndrome virus (TSV), a pathogen of penaeid shrimp and member of the family Dicistroviridae, was recently reported to have the ability to infect primate cells. We independently retested this hypothesis. Three lines of primate cells FRhK-4, MA-104, and BGMK, which are highly susceptible to infection by human picornaviruses, were challenged with TSV. Viral replication was assayed by real time reverse transcription-polymerase chain reaction using cell media samples collected on days 0, 4, and 7 postchallenge. By day 7, genome copy numbers had decreased 25%-99%. No cytopathic effect was observed after 7 days. An in situ hybridization assay, with gene probes specific for detection of TSV, was negative for TSV in challenged cells. The infectivity of residual virus in the cell culture media at day 7 was confirmed by bioassay using TSV-free indicator shrimp (Litopenaeus vannamei). TSV did not infect the primate cells tested, and no evidence of zoonotic potential was found. PMID- 15663847 TI - Venezuelan equine encephalitis virus, southern Mexico. AB - Equine epizootics of Venezuelan equine encephalitis (VEE) occurred in the southern Mexican states of Chiapas in 1993 and Oaxaca in 1996. To assess the impact of continuing circulation of VEE virus (VEEV) on human and animal populations, serologic and viral isolation studies were conducted in 2000 to 2001 in Chiapas State. Human serosurveys and risk analyses indicated that long-term endemic transmission of VEEV occurred among villages with seroprevalence levels of 18% to 75% and that medical personnel had a high risk for VEEV exposure. Seroprevalence in wild animals suggested cotton rats as possible reservoir hosts in the region. Virus isolations from sentinel animals and genetic characterizations of these strains indicated continuing circulation of a subtype IE genotype, which was isolated from equines during the recent VEE outbreaks. These data indicate long-term enzootic and endemic VEEV circulation in the region and continued risk for disease in equines and humans. PMID- 15663848 TI - Opisthorchiasis from imported raw fish. AB - Liver fluke infection caused by Opisthorchiidae is a major public health problem in many parts of the Far East, Southeast Asia, and eastern Europe. However, with the growing volume of international travel and population migration, the infection is increasingly diagnosed in countries where the disease is not endemic, particularly in North America. We report an outbreak of acute opisthorchiasis in a family that was infected in a non-disease-endemic area after eating raw carp illegally imported from a highly disease-endemic area in Siberia. With the growing numbers of former Soviet Union citizens immigrating to other countries, western physicians need to be alert regarding Opisthorchis-associated pathology in this population. The epidemiology and biology of Opisthorchiidae in the former Soviet Union are reviewed. PMID- 15663850 TI - West Nile virus outbreak in North American owls, Ontario, 2002. AB - From July to September 2002, an outbreak of West Nile virus (WNV) caused a high number of deaths in captive owls at the Owl Foundation, Vineland, Ontario, Canada. Peak death rates occurred in mid-August, and the epidemiologic curve resembled that of corvids in the surrounding Niagara region. The outbreak occurred in the midst of a louse fly (Icosta americana, family Hippoboscidae) infestation. Of the flies tested, 16 (88.9 %) of 18 contained WNV RNA. Species with northern native breeding range and birds >1 year of age were at significantly higher risk for WNV-related deaths. Species with northern native breeding range and of medium-to-large body size were at significantly higher risk for exposure to WNV. Taxonomic relations (at the subfamily level) did not significantly affect exposure to WNV or WNV-related deaths. Northern native breeding range and medium-to-large body size were associated with earlier death within the outbreak period. Of the survivors, 69 (75.8 %) of 91 were seropositive for WNV. PMID- 15663849 TI - Identifying rodent hantavirus reservoirs, Brazil. AB - We describe the genetic analysis of samples from hantavirus pulmonary syndrome (HPS) patients from southern and southeastern states of Brazil and rodents captured at the presumed site of infection of these patients. A total of 65 samples that were antibody-positive for Sin Nombre or Laguna Negra virus by enzyme-linked immunosorbent assay were processed by nested reverse transcription polymerase chain reaction (RT-PCR) by using several primer combinations in the M and S genome segments. PCR products were amplified and sequenced from samples from 11 HPS patient and 7 rodent samples. Phylogenetic analysis of nucleotide sequence differences showed the cocirculation of Araraquara and Juquitiba-like viruses, previously characterized from humans. Our genetic data indicate that Araraquara virus is associated with Bolomys lasiurus (hairy-tailed Bolo mouse) and the Juquitiba-like virus is associated with Oligoryzomys nigripes (black footed pigmy rice rat). PMID- 15663851 TI - Crimean-Congo hemorrhagic fever, Mauritania. AB - From February to August 2003, 38 persons were infected with Crimean-Congo hemorrhagic fever (CCHF) virus in Mauritania; 35 of these persons were residents of Nouakchott. The first patient was a young woman who became ill shortly after butchering a goat. She transmitted the infection to 15 persons in the hospital where she was admitted and four members of her family. In Nouakchott, two disease clusters and 11 isolated cases were identified. The case-fatality ratio was 28.6%. Of the patients not infected by the first case-patient, almost half were butchers, which suggests that the primary mode of animal-to-human transmission was direct contact with blood of infected animals. The hospital outbreak alerted health authorities to sporadic cases that occurred in the following weeks, which would have probably gone otherwise unnoticed. Studies must be conducted to determine the potential risk for continued sporadic outbreaks of CCHF in humans and to propose prevention measures. PMID- 15663852 TI - Alligators as West Nile virus amplifiers. AB - Recent evidence suggests that American alligators (Alligator mississippiensis) may be capable of transmitting West Nile virus (WNV) to other alligators. We experimentally exposed 24 juvenile alligators to WNV parenterally or orally. All became infected, and all but three sustained viremia titers >5.0 log10 PFU/mL (a threshold considered infectious for Culex quinquefasciatus mosquitoes) for 1 to 8 days. Noninoculated tankmates also became infected. The viremia profiles and multiple routes of infection suggest alligators may play an important role in WNV transmission in areas with high population densities of juvenile alligators. PMID- 15663854 TI - Differential virulence of West Nile strains for American crows. AB - Crow deaths were observed after West Nile virus (WNV) was introduced into North America, and this phenomenon has subsequently been used to monitor the spread of the virus. To investigate potential differences in the crow virulence of different WNV strains, American Crows were inoculated with Old World strains of WNV from Kenya and Australia (Kunjin) and a North American (NY99) WNV genotype. Infection of crows with NY99 genotype resulted in high serum viremia levels and death; the Kenyan and Kunjin genotypes elicited low viremia levels and minimal deaths but resulted in the generation of neutralizing antibodies capable of providing 100% protection from infection with the NY99 strain. These results suggest that genetic alterations in NY99 WNV are responsible for the crow virulent phenotype and that increased replication of this strain in crows could spread WNV in North America. PMID- 15663853 TI - H3N2 influenza virus transmission from swine to turkeys, United States. AB - In 1998, a novel H3N2 reassortant virus emerged in the United States swine population. We report the interspecies transmission of this virus to turkeys in two geographically distant farms in the United States in 2003. This event is of concern, considering the reassortment capacity of this virus and the susceptibility of turkey to infection by avian influenza viruses. Two H3N2 isolates, A/turkey/NC/16108/03 and A/turkey/MN/764/03, had 98.0% to 99.9% nucleotide sequence identity to each other in all eight gene segments. All protein components of the turkey isolates had 97% to 98% sequence identity to swine H3N2 viruses, thus demonstrating interspecies transmission from pigs to turkeys. The turkey isolates were better adapted to avian hosts than were their closest swine counterparts, which suggests that the viruses had already begun to evolve in the new host. The isolation of swine-like H3N2 influenza viruses from turkeys raises new concerns for the generation of novel viruses that could affect humans. PMID- 15663855 TI - Cats as a risk for transmission of antimicrobial drug-resistant Salmonella. AB - To determine whether cats were a risk for transmission of Salmonella to humans, we evaluated the excretion of Salmonella by pet cats. Rectal-swab specimens were taken from 278 healthy house cats, from 58 cats that died of disease, and from 35 group-housed cats. Group-housed cats were kept in one room with three cat trays and a common water and feed tray. Eighteen (51.4%) of 35 group-housed cats, 5 (8.6%) of 58 diseased cats (5/58), and 1 (0.36%) of 278 healthy house cats excreted Salmonella. Salmonella isolates were of serotypes Typhimurium, Enteritidis, Bovismorbificans and 4:i:-. Acquired antimicrobial resistance was found in serotype Typhimurium (resistance to ampicillin, chloramphenicol, and tetracycline; to ampicillin; and to chloramphenicol) and 4:i:- strains (resistance to ampicillin, chloramphenicol, sulfonamides, trimethoprim, and sulfamethoxazole/trimethoprim). Cats that excrete Salmonella can pose a public health hazard to people who are highly susceptible to Salmonella, such as children, the elderly, and immunocompromised persons. PMID- 15663856 TI - VecTest as diagnostic and surveillance tool for West Nile virus in dead birds. AB - The VecTest antigen-capture assay for West Nile virus was performed on oral and tissue swabs from dead birds in New York State from April 2003 through July 2004. Results were compared with those from real-time reverse transcriptase-polymerase chain reaction of kidney or brain. Oral VecTest sensitivity is adequate for surveillance in American Crows (Corvus brachyrhynchos) (87%), Blue Jays (Cyanocitta cristata) (80%), and House Sparrows (Passer domesticus) (76%). Oral VecTest performed well for small samples of American Kestrels (Falco sparverius), Northern Cardinals (Cardinalis cardinalis), Common Grackles (Quiscalus quiscula), and House Finches (Carpodacus mexicanus). Poor sensitivity occurred in most raptors, Mourning Doves (Zenaida macroura), Fish Crows (Corvus ossifragus), and American Robins (Turdus migratorius). Specificity was excellent (98%), except for false-positive results that occurred mostly in Gray Catbirds (Dumatella carolinensis), Green Herons (Butorides virescens), and tests of blood and tissues. Feather pulp and kidney may be useful for VecTest assays in corvids. PMID- 15663858 TI - Avian influenza H5N1 in tigers and leopards. AB - Influenza virus is not known to affect wild felids. We demonstrate that avian influenza A (H5N1) virus caused severe pneumonia in tigers and leopards that fed on infected poultry carcasses. This finding extends the host range of influenza virus and has implications for influenza virus epidemiology and wildlife conservation. PMID- 15663857 TI - Experimental Everglades virus infection of cotton rats (Sigmodon hispidus). AB - Everglades virus (EVEV), an alphavirus in the Venezuelan equine encephalitis (VEE) serocomplex, circulates among rodents and vector mosquitoes and infects humans, causing a febrile disease sometimes accompanied by neurologic manifestations. EVEV circulates near metropolitan Miami, which indicates the potential for substantial human disease, should outbreaks arise. We characterized EVEV infection of cotton rats in South Florida, USA, to validate their role in enzootic transmission. To evaluate whether the viremia induced in cotton rat populations regulates EVEV distribution, we also infected rats from a non-EVEV endemic area. Viremia levels developed in rats from both localities that exceeded the threshold for infection of the vector. Most animals survived infection with no signs of illness, despite virus invasion of the brain and the development of mild encephalitis. Understanding the mechanisms by which EVEV-infected cotton rats resist clinical disease may be useful in developing VEE therapeutics for equines and humans. PMID- 15663859 TI - Novel avian influenza H7N3 strain outbreak, British Columbia. AB - Genome sequences of chicken (low pathogenic avian influenza [LPAI] and highly pathogenic avian influenza [HPAI]) and human isolates from a 2004 outbreak of H7N3 avian influenza in Canada showed a novel insertion in the HA0 cleavage site of the human and HPAI isolate. This insertion likely occurred by recombination between the hemagglutination and matrix genes in the LPAI virus. PMID- 15663860 TI - Human illness from avian influenza H7N3, British Columbia. AB - Avian influenza that infects poultry in close proximity to humans is a concern because of its pandemic potential. In 2004, an outbreak of highly pathogenic avian influenza H7N3 occurred in poultry in British Columbia, Canada. Surveillance identified two persons with confirmed avian influenza infection. Symptoms included conjunctivitis and mild influenzalike illness. PMID- 15663861 TI - SARS molecular detection external quality assurance. AB - Inactivated severe acute respiratory syndrome-associated coronavirus samples were used for an external quality assurance study within the World Health Organization SARS Reference and Verification Network and other reference institutions. Of 58 participants, 51 correctly detected virus in all samples > or =9,400 RNA copies per milliliter and none in negative samples. Commercial test kits significantly improved the outcome. PMID- 15663862 TI - Detecting West Nile virus in owls and raptors by an antigen-capture assay. AB - We evaluated a rapid antigen-capture assay (VecTest) for detection of West Nile virus in oropharyngeal and cloacal swabs, collected at necropsy from owls (N = 93) and raptors (N = 27). Sensitivity was 93.5%-95.2% for northern owl species but <42.9% for all other species. Specificity was 100% for owls and 85.7% for raptors. PMID- 15663863 TI - Parastrongylus cantonensis in a nonhuman primate, Florida. AB - Parastrongylus (= Angiostrongylus) cantonensis is a parasitic nematode of Norway rats throughout tropical regions. This parasite is neurotropic and causes disease and death in humans and other mammals. We report the first identification of P. cantonensis as the cause of a debilitating neurologic disease in a captive primate in Florida. PMID- 15663864 TI - Naturally acquired Plasmodium knowlesi malaria in human, Thailand. AB - We describe a case of naturally acquired infection with Plasmodium knowlesi in Thailand. Diagnosis was confirmed by the small subunit ribosomal RNA and the mitochondrial cytochrome b sequences. The occurrence of simian malaria in human has signified the roles of wild primate populations in disease transmission in some malaria-endemic areas. PMID- 15663866 TI - Protective effectiveness of hantavirus vaccine. AB - A case-control study in the Republic of Korea evaluated the protective effectiveness of the hantavirus vaccine. Point estimates showed increasing effectiveness with increasing numbers of doses received: 25% for one dose, 46% for two doses, and 75% for three doses. All 95% confidence intervals overlapped zero; therefore, the findings could be due to chance. PMID- 15663865 TI - Rabies in endangered Ethiopian wolves. AB - With rabies emerging as a particular threat to wild canids, we report on a rabies outbreak in a subpopulation of endangered Ethiopian wolves in the Bale Mountains, Ethiopia, in 2003 and 2004. Parenteral vaccination of wolves was used to manage the outbreak. PMID- 15663867 TI - Genome sequence and attenuating mutations in West Nile virus isolate from Mexico. AB - The complete genome sequence of a Mexican West Nile virus isolate, TM171-03, included 46 nucleotide (0.42%) and 4 amino acid (0.11%) differences from the NY99 prototype. Mouse virulence differences between plaque-purified variants of TM171 03 with mutations at the E protein glycosylation motif suggest the emergence of an attenuating mutation. PMID- 15663868 TI - Animal-to-human transmission of Salmonella Typhimurium DT104A variant. AB - Salmonella enterica serovar Typhimurium was isolated from a pig, a calf, and a child on a farm in the Netherlands. The isolates were indistinguishable by phenotyping and genotyping methods, which suggests nonfoodborne animal-to-animal and animal-to-human transmission. Persons in close contact with farm animals should be aware of this risk. PMID- 15663869 TI - Isolation and molecular identification of Nipah virus from pigs. AB - Nipah viruses from pigs from a Malaysian 1998 outbreak were isolated and sequenced. At least two different Nipah virus strains, including a previously unreported strain, were identified. The findings highlight the possibility that the Malaysia outbreaks had two origins of Nipah virus infections. PMID- 15663870 TI - Serologic evidence of lyssavirus infection in bats, Cambodia. AB - In Cambodia, 1,303 bats of 16 species were tested for lyssavirus. No lyssavirus nucleocapsid was detected in 1,283 brains tested by immunofluorescence assay. Antibodies against lyssaviruses were detected by enzyme-linked immunosorbent assay in 144 (14.7%) of 981 serum samples. Thirty of 187 serum samples contained neutralizing antibodies against different lyssaviruses. PMID- 15663871 TI - Human-to-dog transmission of methicillin-resistant Staphylococcus aureus. AB - Methicillin-resistant Staphylococcus aureus (MRSA) was cultured from the nose of a healthy dog whose owner was colonized with MRSA while she worked in a Dutch nursing home. Pulsed-field gel electrophoresis and typing of the staphylococcal chromosome cassette mec (SCCmec) region showed that both MRSA strains were identical. PMID- 15663873 TI - First human cases of tickborne encephalitis, Norway. AB - The first reported case of tickborne encephalitis (TBE) in Norway occurred in 1997. From 1997 to 2003, from zero to two cases of human TBE have been diagnosed per year in Norway, for a total of eight cases. Clinical TBE cases in dogs are not reported in Norway. PMID- 15663872 TI - Francisella tularensis peritonitis in stomach cancer patient. AB - Tularemia with peritonitis developed in a 50-year-old man soon after diagnosis of stomach cancer with metastasis. The ascites grew Francisella tularensis subsp. holarctica, which was identified by sequencing analysis of the 16S rDNA. The infection resolved with antimicrobial treatment. Antibodies detected 4 weeks after onset disappeared after chemotherapy-associated lymphopenia. PMID- 15663874 TI - Antibodies to SARS coronavirus in civets. AB - Using three different assays, we examined 103 serum samples collected from different civet farms and a market in China in June 2003 and January 2004. While civets on farms were largely free from SARS-CoV infection, approximately 80% of the animals from one animal market in Guangzhou contained significant levels of antibody to SARS-CoV, which suggests no widespread infection among civets resident on farms, and the infection of civets in the market might be associated with trading activities under the conditions of overcrowding and mixing of various animal species. PMID- 15663875 TI - Salmonella Typhimurium outbreak associated with veterinary clinic. AB - A Salmonella enterica serovar Typhimurium outbreak was associated with a veterinary clinic. Confirmed cases were in one cat, two veterinary technicians, four persons associated with clinic patients, and a nurse not linked to the clinic. This outbreak emphasizes the importance of strong public health ties to the animal health community. PMID- 15663876 TI - West Nile virus viremia in wild rock pigeons. AB - Feral rock pigeons were screened for neutralizing antibodies to West Nile virus (WNV) during late winter/spring and summer of 2002 and 2003. Additionally, virus isolation from serum was attempted from 269 birds collected during peak transmission periods. The observed viremia levels and seroprevalence indicate that this species could be involved in amplifying WNV in urban settings. PMID- 15663877 TI - Health monitoring and life on the Mississippi. PMID- 15663878 TI - Using survey data for diabetes surveillance among minority populations: a report of the Centers for Disease Control and Prevention's expert panel meeting. AB - INTRODUCTION: Data on diabetes morbidity and mortality and the quality of care among U.S. minority populations are necessary to assess progress toward eliminating racial/ethnic disparities and to design and implement effective interventions. This paper summarizes the discussions and recommendations of an expert panel to address the use of survey data for diabetes surveillance among minority populations. METHODS: The Centers for Disease Control and Prevention's Division of Diabetes Translation convened an expert panel of persons with survey experience and awareness of the problems in conducting health-related surveys among minority populations. Panel members were asked to 1) determine ways to enhance the ability of existing survey systems to address diabetes surveillance among minority populations; 2) identify survey systems that could be used to address surveillance needs; and 3) determine whether new minority-specific survey systems need to be developed. RESULTS: Panel members concluded that, although no existing survey system is completely adequate for diabetes surveillance among minority populations, new systems should not be developed. They recommended 1) investigating the use of community-based surveys; 2) exploring the ability of national surveys to increase sample sizes and produce state-level estimates; and 3) encouraging government agencies and public health programs to coordinate and integrate diabetes-related survey data and share analytic methodology. CONCLUSION: No existing survey is suitable for conducting minority-specific diabetes surveillance. Modifying and expanding existing surveys to establish a diabetes surveillance system of sentinel minority populations would be more feasible than developing a new one. Interagency coordination and collaboration will be critical in this effort. PMID- 15663879 TI - State-based diabetes surveillance among minority populations. PMID- 15663880 TI - The burden of chronic disease: the future is prevention. Introduction to Dr. James Marks' presentation, "The Burden of Chronic Disease and the Future of Public Health". PMID- 15663881 TI - Policy and environmental indicators for heart disease and stroke prevention: data sources in two states. AB - INTRODUCTION: Investigators in South Carolina and Alabama assessed the availability of data for measuring 31 policy and environmental indicators for heart disease and stroke prevention. The indicators were intended to determine policy and environmental support for adopting heart disease and stroke prevention guidelines and selected risk factors in 4 settings: community, school, work site, and health care. METHODS: Research teams used literature searches and key informant interviews to explore the availability of data sources for each indicator. Investigators documented the following 5 qualities for each data source identified: 1) the degree to which the data fit the indicator; 2) the frequency and regularity with which data were collected; 3) the consistency of data collected across time; 4) the costs (time, money, personnel) associated with data collection or access; and 5) the accessibility of data. RESULTS: Among the 31 indicators, 11 (35%) have readily available data sources and 4 (13%) have sources that could provide partial measurement. Data sources are available for most indicators in the school setting and for tobacco control policies in all settings. CONCLUSION: Data sources for measuring policy and environmental indicators for heart disease and stroke prevention are limited in availability. Effort and resources are required to develop and implement mechanisms for collecting state and local data on policy and environmental indicators in different settings. The level of work needed to expand data sources is comparable to the extensive work already completed in the school setting and for tobacco control. PMID- 15663884 TI - Featured abstracts from the 18th National Conference on Chronic Disease Prevention and Control. PMID- 15663882 TI - Social and cultural barriers to diabetes prevention in Oklahoma American Indian women. AB - INTRODUCTION: The prevalence of diabetes is disproportionately higher among minority populations, especially American Indians. Prevention or delay of diabetes in this population would improve quality of life and reduce health care costs. Identifying cultural definitions of health and diabetes is critically important to developing effective diabetes prevention programs. METHODS: In-home qualitative interviews were conducted with 79 American Indian women from 3 tribal clinics in northeast Oklahoma to identify a cultural definition of health and diabetes. Grounded theory was used to analyze verbatim transcripts. RESULTS: The women interviewed defined health in terms of physical functionality and absence of disease, with family members and friends serving as treatment promoters. Conversely, the women considered their overall health to be a personal issue addressed individually without burdening others. The women presented a fatalistic view of diabetes, regarding the disease as an inevitable event that destroys health and ultimately results in death. CONCLUSION: Further understanding of the perceptions of health in at-risk populations will aid in developing diabetes prevention programs. PMID- 15663883 TI - Pediatric hospitalizations for asthma: use of a linked file to separate person level risk and readmission. AB - INTRODUCTION: Disparities in asthma hospitalization by gender, age, and race/ethnicity are thought to be driven by a combination of 2 factors: disease severity and inadequate health care. Hospitalization data that fail to differentiate between numbers of admissions and numbers of individuals limit the ability to derive accurate conclusions about disparities and risks. METHODS: Hospitalization records for pediatric asthma patients (aged one to 14 years) were extracted from New Jersey Hospital Discharge Files (for the years 1994 through 2000) and then linked by patient identifiers using a probabilistic matching algorithm. The analysis file contained 30,400 hospital admissions for 21,016 children. Hospitalization statistics were decomposed into persons hospitalized and number of hospitalizations. Analysis of readmission within 180 days of discharge used additional records from 2001 to avoid bias due to truncated observation. RESULTS: Overall, 22.9% of children in our analysis had repeat asthma admissions within the same age interval, accounting for 30.9% of all hospitalizations. Also among all children, 11.7% had at least one readmission within 180 days of a prior discharge. The risk of hospitalization was higher for boys, decreased by age for both genders, was lowest for white children and highest for black children. Readmission rates were higher for black and Hispanic girls than boys in older age groups, but were otherwise relatively uniform by gender and age. CONCLUSION: Decomposition of ratios of total hospitalizations to population illuminates components of risk and suggests specific causes of disparity. PMID- 15663885 TI - A model community skin cancer prevention project in Maine. PMID- 15663887 TI - Response to S. Leonard Syme's essay. PMID- 15663886 TI - Law as a tool for preventing chronic diseases: expanding the spectrum of effective public health strategies. PMID- 15663888 TI - Development of novel therapies for Huntington's disease: hope and challenge. AB - Huntington's disease (HD) is an autosomal dominant neurological disease. It is a fatal neurological disorder affecting 5-10 out of 10,000 people. While there are intensive research efforts focusing on uncovering molecular mechanisms of the pathogenesis of HD, a number of studies have begun to look for effective therapies for HD. There is a large body of encouraging news on novel therapeutic developments. The present paper reviews drugs used for symptomatic treatment of HD and experimental therapies targeting HD molecular pathology. PMID- 15663889 TI - Anti-amnestic and anti-aging effects of ginsenoside Rg1 and Rb1 and its mechanism of action. AB - In the present paper, we overview the discovery of new biological activities induced by ginsenoside Rg1 and Rb1 and discuss possible mechanisms of action. Both compounds could increase neural plasticity in efficacy and structure; especially Rg1, as one small molecular drug, can increase proliferation and differentiation of neural progenitor cells in dentate gyrus of hippocampus of normal adult mice and global ischemia model in gerbils. This finding has great value for treatment of Alzheimer's disease and other neurodegenerative disorders which is characterized by neurons loss. Increase of expression of brain derived neurotrophic factor, Bcl-2 and antioxidant enzyme, enhanced new synapse formation, inhibition of apoptosis and calcium overload are also important neuron protective factors. Rg1 and Rb1 have common effects, but there are some differences in pharmacology and mechanism. These differences may attribute to their different chemical structure. Rg1 is panaxtriol with two sugars, while Rb1 is panaxtriol with four sugars. PMID- 15663890 TI - Caspase-1 inhibitor Ac-YVAD-CHO attenuates quinolinic acid-induced increases in p53 and apoptosis in rat striatum. AB - AIM: To study the effects of the caspase-1 inhibitor Ac-YVAD-CHO on quinolinic acid (QA)-induced apoptosis. METHODS: Rats were pre-treated with intrastriatal infusion of Ac-YVAD-CHO (2-8 microg) before intrastriatal injection of QA (60 nmol). Striatal total proteins, genomic DNA, and nuclear proteins were isolated. The effects of Ac-YVAD-CHO on QA-induced caspase-1 activity, internucleosomal DNA fragmentation, IkappaB-alpha degradation, NF-kappaB, and AP-1 activation, and increases in p53 protein levels were measured with enzyme assays, agarose gel electrophoresis, electrophoresis mobility shift assays, and Western blot analysis. RESULTS: Pre-treatment with Ac-YVAD-CHO inhibited QA-induced internucleosomal DNA fragmentation. Ac-YVAD-CHO inhibited QA-induced increases in caspase-1 activity and p53 protein levels, but had no effect on QA-induced IkappaB-alpha degradation, NF-kappaB or AP-1 activation. CONCLUSION: Caspase-1 is involved in QA-induced p53 upregulation but not IkappaB-alpha degradation. Inhibition of caspase-1 attenuates QA-induced apoptosis in rat striatum. PMID- 15663891 TI - Orexin A promotes histamine, but not norepinephrine or serotonin, release in frontal cortex of mice. AB - AIM: To investigate the effects of orexin A on release of histamine, norepinephrine, and serotonin in the frontal cortex of mice. METHODS: Samples for measuring histamine, norepinephrine, and serotonin contents were collected by in vivo microdialysis of the frontal cortex of anesthetized mice. The histamine, noradrenaline, and serotonin content in dialysates were measured by HPLC techniques. RESULTS: Intracrebroventricular injection of orexin A at doses of 12.5, 50, and 200 pmol per mouse promoted histamine release from the frontal cortex in a dose-dependent manner. At the highest dose given, 200 pmol, orexin A significantly induced histamine release, with the maximal magnitude being 230% over the mean basal release. The enhanced histamine release was sustained for 140 min, and then gradually returned to the basal level. However, no change in norepinephrine or serotonin release was observed under application of the same dose of orexin A. CONCLUSION: These results suggest that the arousal effect of orexin A is mainly mediated by histamine, not by norepinephrine or serotonin. PMID- 15663892 TI - Modulating effect of adenosine deaminase on function of adenosine A1 receptors. AB - AIM: To study the modulating effect of adenosine deaminase (ADA) on the adenosine A1 receptor (A1R) in HEK293 cells stably expressing the human A1R. METHODS: cDNA was amplified by RT-PCR using total RNA from human embryo brain tissue as the template. The PCR products were subcloned into the plasmid pcDNA3 and cloned into the plasmid pcDNA3.1. The cloned A1R cDNA was sequenced and stably expressed in HEK293 cells. The modulating effect of adenosine deaminase on A1R was studied by using [3H]DPCPX binding assay and an intracellular calcium assay. RESULTS: HEK293 cells stably expressing human A1R were obtained. Saturation studies showed that the K(D) value and B(max) value of [3H]DPCPX were 1.6+/-0.2 nmol/L and 1.819+/ 0.215 nmol/g of protein respectively, in the absence of ecto-ADA respectively, and 1.3+/-0.2 nmol/L and 1.992+/-0.130 nmol/g of protein in the presence of ecto ADA respectively, suggesting that the K(D) value and B(max) value of [3H]DPCPX were unaffected by ecto-ADA. In the case of [3H]DPCPX competition curves obtained from intact cells or membranes, A1R agonist CCPA/[3H]DPCPX competition curve could be fitted well to a one-site model in the absence of ecto-ADA and a two site model in the presence of ecto-ADA with a K(H) value of 0.74 (0.11+/-4.8) nmol/L (intact cells) or 1.8 (0.25+/-10) nmol/L (membrane) and a K(L) value of 0.94 (0.62+/-1.41) micromol/L (intact cells) or 0.77 (0.29+/-0.99) micromol/L (membrane). The K(L) value is not significantly different from the IC50 value of 0.84(0.57+/-1.23) micromol/L (intact cells) or 0.84 (0.63+/-1.12) micromol/L (membrane) obtained in the absence of ecto-ADA. Similar results were obtained from the CPA/[3H]DPCPX competition curve in the absence or presence of ecto-ADA on intact cells or membranes. Intracellular calcium assay demonstrated that the EC50 value of CPA were 10 (5+/-29) nmol/L and 94 (38+/-229) nmol/L in the presence or absence of ecto-ADA, respectively. CONCLUSION: A1R stably expressed in the HEK293 cells display a low affinity for agonists in the absence of ADA and high and low affinities for agonists in the presence of ADA. The presence of ADA may promote the signaling through the adenosine A1 receptor in HEK293 cells. PMID- 15663893 TI - Modulation of P-glycoprotein function by amlodipine derivatives in brain microvessel endothelial cells of rats. AB - AIM: To investigate whether the amlodipine derivatives, CJX1 and CJX2, have a modulative effect on P-glycoprotein (P-gp) function in rat brain microvessel endothelial cells (RBMEC). METHODS: Isolated RBMEC were cultured in DMEM/F12 (1:1) medium. The amount of intracellular rhodamine (Rh123) was determined, using a fluorescence spectrophotometer, to evaluate the function of P-gp. RESULTS: The accumulation of Rh123 in RBMEC was potentiated in a concentration-dependent manner after incubation with CJX1 and CJX2 at 1, 2.5, 5, and 10 micromol/L (P<0.01), but no accumulation of Rh123 was observed in human umbilical vein endothelial cells after incubation with CJX1 and CJX2 10 micromol/L (P>0.05). Accumulation of intracellular Rh123 was increased and efflux of intracellular Rh123 was decreased in a time-dependent manner from 0-100 min after CJX1 and CXJ2 at 10 micromol/L treatment. The inhibitory effect of CJX1 and CJX2 on P-gp function was reversible and remained even at 120 min after removal of CJX1 and CJX2 at 2.5 micromol/L from the medium. CONCLUSION: CJX1 and CJX2 exhibited a potent effect in the inhibition of P-gp function in vitro. PMID- 15663894 TI - Estrogen stimulates release of secreted amyloid precursor protein from primary rat cortical neurons via protein kinase C pathway. AB - AIM: To investigate the mechanism of the action of estrogen, which stimulates the release of secreted amyloid precursor protein alpha (sAPP(alpha)) and decreases the generation of amyloid-beta protein (A(beta)), a dominant component in senile plaques in the brains of Alzheimer's disease patients. METHODS: Experiments were carried out in primary rat cortical neurons, and Western blot was used to detect sAPP(alpha) in a culture medium and the total amount of cellular amyloid precursor protein (APP) in neurons. RESULTS: 17beta-Estradiol (but not 17alpha estradiol) and beta-estradiol 6-(O-carboxymethyl) oxime: BSA increased the secretion of sAPP(alpha) and this effect was blocked by protein kinase C (PKC) inhibitor calphostin C, but not by the classical estrogen receptor antagonist ICI 182,780. Meanwhile, 17beta-estradiol did not alter the synthesis of cellular APP. CONCLUSION: The effect of 17beta-estradiol on sAPP(alpha) secretion is likely mediated through the membrane binding sites, and needs molecular configuration specificity of the ligand. Furthermore, the action of the PKC-dependent pathway might be involved in estrogen-induced sAPP(alpha) secretion. PMID- 15663895 TI - Visual recognition memory is related to basic expression level of NMDA receptor NR1/NR2B subtype in hippocampus and striatum of rats. AB - AIM: To examine the basic expression levels of N-methyl-D-aspartate (NMDA) receptor NR1 and NR2B subunits in six brain regions of Sprague-Dawley (SD) rats with different visual recognition memory. METHODS: Rats were tested by a novel object-recognition model and grouped into the high and the low visual recognition memory groups. The expression levels of NR1 and NR2B subunits in the cortex, hippocampus, striatum, amygdala, diencephalon, and olfactory bulb were measured by semiquantitative immunoblotting. RESULTS: The NR1 and NR2B subunit protein levels in the hippocampus of the high visual recognition memory group were 35.9% (P<0.01) and 53.4% (P<0.05) higher respectively than those in the low group. In addition, the NR2B level in the striatum in the high visual recognition memory group was 25.0% (P<0.05) higher than that in the low one. However, no significant difference was found in the levels of the subunits between the two groups in other brain regions. CONCLUSION: The visual recognition memory in rats is related to the basic expression level of NMDA receptor NR1/NR2B subtype in the hippocampus and striatum. PMID- 15663896 TI - Effects of adenosine agonist R-phenylisopropyl-adenosine on halothane anesthesia and antinociception in rats. AB - AIM: To investigate the antinociceptive effect of adenosine agonist R phenylisopropyl-adenosine (R-PIA) given to conscious rats by intracerebroventricular (ICV) and intrathecal (IT), and identify the effect of R PIA on minimum alveolar concentration (MAC) of halothane with pretreatment of A1 receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) or K+ channel blocker 4-aminopyridine (4-AP). METHODS: Sprague-Dawley rats were implanted with 24-gauge stainless steel guide cannula using stereotaxic apparatus and ICV method, and an IT catheter (PE-10, 8.5 cm) was inserted into the lumbar subarachnoid space, while the rats were under pentobarbital anesthesia. After one week of recovery from surgery, rats were randomly assigned to one of the following protocols: MAC of halothane, or tail-flick latency. All measurements were performed after R-PIA (0.8-2.0 microg) microinjection into ICV and IT with or without pretreatment of DPCPX or 4-AP. RESULTS: Microinjection of adenosine agonist R-PIA in doses of 0.8-2.0 microg into ICV and IT produced a significant dose- and time-dependent antinociceptive action as reflected by increasing latency times and ICV administration of adenosine agonist R-PIA (0.8 microg) reducing halothane anesthetic requirements (by 29%). The antinociception and reducing halothane requirements effected by adenosine agonist R-PIA was abolished by DPCPX and 4-AP. CONCLUSION: ICV and IT administration of adenosine agonist R PIA produced an antinociceptive effect in a dose-dependent manner and decreased halothane MAC with painful stimulation through activation of A1 receptor subtype, and the underlying mechanism involves K+ channel activation. PMID- 15663897 TI - Effects of intrathecal 6-hydroxydopamine, alpha1 and alpha2 adrenergic receptor antagonists on antinociception of propofol in mice. AB - AIM: To investigate the relationship between spinal cord norepinephrine, alpha1 and alpha2 adrenergic receptors and antinociception of propofol in mice. METHODS: Kunming mice were used. Antinociceptive tests were investigated with the tail immersion test and the acetic acid-induced writhing test. The effects of subcutaneous (sc), intrathecal (ith) and intracerebroventricular (icv) injection propofol on pain threshold were observed. The influences of pretreatment with ith 6-hydroxydopamine, alpha1R antagonist prazosin, or alpha2R antagonist yohimbine on the antinociception of propofol were studied. RESULTS: Significant antinociception was produced by propofol (25, 50 mg/kg, sc) and propofol (20, 40 microg, ith) in tail-immersion test and acetic the acid-induced writhing test (P<0.05 or P<0.01). Icv propofol (10, 20, and 40 microg) did not produce any effect on pain threshold in mice (P>0.05). The 6-hydroxydopamine (5 and 10 microg), prazosin (5 and 10 microg), or yohimbine (5 and 10 microg) ith alone did not affect basal tail-flick latency (TFL) in conscious mice, but significantly reduced the TFL as measured by tail-immersion test in propofol (50 mg/kg, sc) treated mice, compared with basal TFL and vehicle groups (P<0.05 or P<0.01). CONCLUSION: The spinal cord is a target of propofol antinociception. In mice propofol antinociception is partly mediated by spinal norepinephrine, alpha1R and alpha2R. PMID- 15663898 TI - Effects of complete Freund's adjuvant on immunohistochemical distribution of IL 1beta and IL-1R I in neurons and glia cells of dorsal root ganglion. AB - AIM: To investigate the effects of complete Freund adjuvant (CFA) on inflammatory hyperalgesia and morphological change of the coexistence of interleukin-1 beta (IL-1beta) and type I IL-1 receptor (IL-1RI) in neurons and glia cells of rat dorsal root ganglion (DRG). METHODS: The pain-related parameters and the expression of IL-1RI and IL-1beta positive neurons and glia cells of DRG in normal saline (NS) and adjuvant-induced arthritic (AA) group were examined with pain behavior assessment methods and immunohistochemical assay, respectively. RESULTS: Five hours, 1 d, and 2 d after intra-articular injection of 50 microL CFA, tactile hyperalgesia induced by CFA was observed in the foot flexion and extension scores of the ipsilateral hindpaw of AA group. Three days after injection, the distribution of IL-1RI/IL-1beta double-stained coexisted neurons and glia cells were observed in ipsilateral DRG of both groups. The number of IL 1beta positive neurons, IL-1RI positive neurons, IL-1beta/IL-1RI double-stained neurons, and IL-1RI positive glia cells in ipsilateral DRG of the AA group were higher than that of NS group (P<0.05 or P<0.01). CONCLUSION: The coexistence of IL-1beta and IL-1RI in neurons and nonneuronal cells suggests an as yet unknown autocrine and/or paracrine function of IL-1beta in the DRG. The function was enhanced in articular arthritis induced by CFA and could play an important role in hyperalgesia under inflammatory conditions. PMID- 15663899 TI - Inhibition of ATP-induced calcium influx in HT4 cells by glucocorticoids: involvement of protein kinase A. AB - AIM: In our previous observations, adenosine triphosphate (ATP) was found to evoke immediate elevations in intracellular free calcium concentration ([Ca2+]i) in HT4 neuroblastoma cells of mice. We tried to see if a brief pretreatment of glucocorticoids could inhibit the Ca2+ response and reveal the underlying signaling mechanism. METHODS: Measurement of [Ca2+]i was carried out using the dual-wavelength fluorescence method with Fura-2 as the indicator. RESULTS: Pre incubation of HT4 cells for 5 min with corticosterone (B) or bovine serum albumin conjugated corticosterone (B-BSA) inhibited the peak [Ca2+]i increments in a concentration-dependent manner. Cortisol and dexamethasone had a similar action, while deoxycorticosterone and cholesterol were ineffective. Both extracellular Ca2+ influx and internal Ca2+ release contributed to ATP-induced [Ca2+]i elevation. The brief treatment with only B attenuated Ca2+ influx. Furthermore, the [Ca2+]i elevation induced by the P2X receptor agonist adenosine 5'-(beta, gamma-methylene) triphosphate (beta, gamma-meATP) was also suppressed. The rapid inhibitory effect of B can be reproduced by forskolin 1 mmol/L and blocked by H89 20 mmol/L. Neither nuclear glucocorticoid receptor antagonist mifepristone nor protein kinase C inhibitors influenced the rapid action of B. CONCLUSION: Our results suggest that glucocorticoids modulate P2X receptor-medicated Ca2+ influx through a membrane-initiated, non-genomic and PKA-dependent pathway in HT4 cells. PMID- 15663900 TI - Onychin inhibits proliferation of vascular smooth muscle cells by regulating cell cycle. AB - AIM: To investigate the effects of onychin on the proliferation of cultured rat artery vascular smooth muscle cells (VSMCs) in the presence of 10% new-born calf serum (NCS). METHODS: Rat VSMCs were incubated with onychin 150 micromol/L or genistein 10 micromol/L in the presence of 10% NCS for 24 h. The proliferation of VSMCs was measured by cell counting and MTS/PMS colorimetric assays. Cell cycle progression was evaluated by flow cytometry. Retinoblastoma (Rb) phosphorylation, and expression of cyclin D1 and cyclin E were measured by Western blot assays. The tyrosine phosphorylation of ERK1/2 was examined by immunoprecipitation techniques using anti-phospho-tyrosine antibodies. RESULTS: The proliferation of VSMCs was accelerated significantly in the presence of 10% NCS. Onychin reduced the metabolic rate of MTS and the cell number of VSMCs in the presence of 10% NCS in a dose-dependent manner. Flow cytometry analysis revealed that the G1-phase fraction ratio in the onychin group was higher than that in the 10% NCS group (85.2% vs 70.0%, P<0.01), while the S-phase fraction ratio in the onychin group was lower than that in 10% NCS group (4.3% vs 16.4%, P<0.01). Western blot analysis showed that onychin inhibited Rb phosphorylation and reduced the expression of cyclin D1 and cyclin E. The effects of onychin on proliferation, the cell cycle and the expression of cyclins in VSMCs were similar to those of genistein, an inhibitor of tyrosine kinase. Furthermore immunoprecipitation studies showed that both onychin and genistein markedly inhibited the tyrosine phosphorylation of ERK1/2 induced by 10% NCS. CONCLUSION: Onychin inhibits the proliferation of VSMCs through G1 phase cell cycle arrest by decreasing the tyrosine phosphorylation of ERK1/2, and the expression of cyclin D1 and cyclin E, and sequentially inhibiting Rb phosphorylation. PMID- 15663901 TI - Effect of a single dose of mifepristone on expression of pinopodes in endometrial surface of mice. AB - AIM: To investigate the effect of mifepristone (RU486) as a single dose on pinopodes expression in the endometrial surface of mice at the time of implantation. METHODS: Pregnant mice in the treated four groups received mifepristone subcutaneously (0.1 mg) between 07:00 and 08:00 AM on Pd (day of pregnancy)1, Pd2, Pd3, and Pd4. Pregnant mice in the non-treated group were used as controls. The uterine horns were collected randomly from two mice in each group between 21:30 and 22:00 PM on Pd4, and from another two mice of the same group between 09:30 and 10:00 AM on Pd5. The specimens were examined by scanning electron microscopy for the detection of pinopodes. RESULTS: When mifepristone was given on Pd1, developing and fully developed pinopodes were observed, but the expression was markedly reduced compared to the control group. When mifepristone was administered on Pd2, only a few developing pinopodes were present. When mifepristone was administered on Pd3, developing pinopodes were observed. When mifepristone was administered on Pd4, different development stage pinopodes were present in specimens collected between 21:30 and 22:00 PM on Pd4, but no pinopodes was observed in specimens taken between 09:30 and 10:00 AM on Pd5. CONCLUSION: These findings suggest that administration of a single dose of RU486 subcutaneously on Pd1, Pd2, Pd3, and Pd4 might play a role in inhibiting development and maturation of endometrium, hence affecting embryo implantation in mice. PMID- 15663902 TI - Swietenia mahagony extract shows agonistic activity to PPAR(gamma) and gives ameliorative effects on diabetic db/db mice. AB - AIM: To search the peroxisome proliferator-activated receptor g (PPAR(gamma)) agonists from Swietenia mahagony extract (SmE) and observe the possible ameliorative effects of SmE on diabetic db/db mice. METHODS: The PPAR(gamma) agonistic activity of SmE was screened by yeast-two hybrid system. The blood glucose levels of diabetic db/db mice were measured using a blood glucose level monitor and the data were statistically analyzed by NDST8.8W software. RESULTS: By using the clinical drug rosiglitazone as a positive control, it was found that the PPARg agonistic activity of SmE at a concentration of 50 microg/L was approximately half that of 35.7 microg/L (0.1 micromol/L) of rosiglitazone. At the dose of 1000 mg/kg, SmE remarkably decreased the blood glucose concentration of db/db mice from (15.26+/-2.98) to (7.58+/-2.20) mmol/L, and reduced the blood glucose levels by 55.49% compared with the control group (P<0.01). CONCLUSION: SmE shows agonistic activity to PPARg and can ameliorate the blood glucose levels of diabetic db/db mice. SmE may be thus used as a potential agent for diabetes therapy. PMID- 15663903 TI - Triptolide suppresses CD80 and CD86 expressions and IL-12 production in THP-1 cells. AB - AIM: To investigate the effects of triptolide, a diterpenoid triepoxide from Tripterygium wilfordii Hook F (TWHF), on the co-stimulatory molecule expression and interleukin-12 (IL-12) production from THP-1 cells. METHODS: THP-1 cells were differentiated into macrophage-like cells by Me2SO, and then cultured with IFN gamma (500 kU/L) and lipopolysaccharide (LPS) (1 mg/L) with or without triptolide. The surface molecule expressions were analyzed on a FACScan flow cytometer. IL-12p40, IL-12p70 were assayed by ELISA. RESULTS: Triptolide suppressed CD80 and CD86 expressions on IFN-gamma (500 kU/L) and LPS (1 mg/L) activated THP-1 cells at nontoxic dosages of 2.5-0.625 microg/L. Furthermore, the production of IL-12p40 and IL-12p70 were also significantly reduced in THP-1 cells exposed to triptolide. CONCLUSION: Triptolide impairs the antigen presenting function by inhibiting CD80 and CD86 expressions and decreased IL 12p40 and IL-12p70 (bioactive form) productions from the activated THP-1 cells. PMID- 15663904 TI - RNA interference by expression of short hairpin RNAs suppresses bcl-xL gene expression in nasopharyngeal carcinoma cells. AB - AIM: To evaluate a new plasmid mediated RNA interference (RNAi) system and investigate whether knock-down of bcl-xL by short hairpin RNA (shRNA) can induce apoptosis of human nasopharyngeal carcinoma (NPC) cell line CNE-2Z in vitro. METHODS: The plasmid containing mU6 promoter was subcloned to yield the pmU6 plasmid, recombinant plasmid expressing shRNA targeting bcl-xL gene was designed and constructed, and were co-transfected cells with green fluorescence protein expressing plasmid. Flow cytometry was used to evaluate transfection efficiency, and RT-PCR and Western blot were applied to analyze bcl-xL mRNA and protein levels, respectively. RESULTS: The shRNA expressed by the recombinant plasmid efficiently suppressed bcl-xL gene expression and induced apoptosis of NPC cells in vitro. CONCLUSION: The recombinant plasmid can sufficiently mediate RNAi in CNE-2Z cells, and knock-down of the bcl-xL expression by shRNA significantly induced apoptosis in CNE-2Z cells. The results suggest this new system, mediated RNAi can be used as a tool for the study of gene function and gene therapy. PMID- 15663905 TI - Synthesis and anti-tumor activity of alkenyl camptothecin esters. AB - AIM: To study the degrees of influence of changing side ester chains at position C20 of camptothecin on the anti-tumor activity of the molecules. METHODS: The esterification reaction of camptothecin 1 and 9-nitrocamptothecin 2 with crotonic anhydride in pyridine gave the corresponding esters 3 and 4, respectively. The acylation of 1 and 2 with cinnamoyl chloride gave products 7 and 8. Epoxidation reaction of 3 and 4 with m-chloroperoxybenzoic acid in benzene solvent gave the products 5 and 6. Esters 3, 4, and 5 were tested for anti-tumor activity against 14 human cancer cell lines. RESULTS: Both in vitro and in vivo anti-tumor activity studies for these esters were conducted and the data demonstrated positive results, that is, these esters were active against the tested tumor lines. CONCLUSION: Alkenyl esters 3 and 4 showed strong anti-tumor activity in vitro against 14 different cancer cell lines. Ester 3 was active against human breast carcinoma in mice and the toxicity of the agent was not observed in mice during the treatment, implying that this agent is effective for treatment with low toxicity. PMID- 15663906 TI - Effect of MePEG molecular weight and particle size on in vitro release of tumor necrosis factor-alpha-loaded nanoparticles. AB - AIM: To study the in vitro release of recombinant human tumor necrosis factor alpha (rHuTNF-alpha) encapsulated in poly (methoxypolyethyleneglycol cyanoacrylate-co-n-hexadecyl cyanoacrylate) (PEG-PHDCA) nanoparticles, and investigate the influence of methoxypolyethyleneglycol (MePEG) molecular weight and particle size. METHODS: Three sizes (approximately 80, 170, and 240 nm) of PEG-PHDCA nanoparticles loading rHuTNF-alpha were prepared at different MePEG molecular weights (M(r) =2000, 5000, and 10,000) using the double emulsion method. The in vitro rHuTNF-alpha release was studied in PBS and rat plasma. RESULTS: A higher burst-release and cumulative-release rate were observed for nanoparticles with higher MePEG molecular weight or smaller particle size. A decreased cumulative release of rHuTNF-alpha following the initial burst effect was found in PBS, while the particle sizes remained constant and MePEG liberated. In contrast, in rat plasma, slowly increased cumulative-release profiles were obtained after the burst effect. During a 5-h incubation in rat plasma, more than 50% of the PEG-PHDCA nanoparticles degraded. CONCLUSION: The MePEG molecular weight and particle size had an obvious influence on rHuTNF-alpha release. rHuTNF alpha released from PEG-PHDCA nanoparticles in a diffusion-based pattern in PBS, but in a diffusion and erosion-controlled manner in rat plasma. PMID- 15663907 TI - Pharmacokinetics and tissue distribution of 5-fluorouracil encapsulated by galactosylceramide liposomes in mice. AB - AIM: To study the pharmacokinetics and tissue distribution of 5-fluorouracil encapsulated by galactosylceramide liposomes (5-Fu-GCL) in mice. METHODS: The concentration of 5-fluorouracil (5-Fu) in serum was detected by high performance liquid chromatography after 5-Fu-GCL (80, 40, 20 mg/kg) and free 5-Fu (40 mg/kg) were injected intravenously into mice. The tissue distribution of 5-Fu-GCL (40 mg/kg) and free 5-Fu (40 mg/kg) was investigated, and concentration-time profile of the two preparations in the liver were studied. Data were analyzed by 3p97 program. RESULTS: Serum concentration-time curves of 5-Fu-GCL and free 5-Fu conformed to one compartment model of first order absorption. 5-Fu-GCL at 80, 40, and 20 mg/kg had T(1/2Ke) of 25.8+/-4.2, 27.3+/-4.4, and 28.2+/-5.6 min; C0 of 24.9+/-4.9, 17.7+/-3.6, and 11.5+/-2.7 mg/L; and AUC of 990.0+/-45.2,622.5+/ 38.3, and 340.4+/-25.6 mg x min x L(-1), respectively. In contrast free 5-Fu at 40 kg/mg had T(1/2Ke) of 15.8+/-2.2 min, C0 of 35.8+/-6.2 mg/L, AUC of 639.0+/ 35.9 mg x min x L(-1). The tissue distribution of 5-Fu-GCL in the liver and immune organs was significantly increased, while in heart and kidney it was remarkably decreased. The AUC of 5-Fu-GCL in the liver was 3 times higher than that of free 5-Fu. CONCLUSION: The pharmacokinetics and tissue distribution of 5 Fu-GCL appears to be linear-related and dose-dependent, and exhibits sustained release and hepatic target characteristics. PMID- 15663908 TI - Statins and the prevention of stroke in diabetes. PMID- 15663911 TI - Atherosclerosis and restenosis: is there a role for RAGE? AB - Diabetic vascular complications are a major cause of morbidity and mortality. Furthermore, such vascular disease is only incompletely explained by "traditional" risk factors in the nondiabetic complications. This situation has prompted the search for factors contributing to the pathogenesis of accelerated and more severe vascular disease in patients with diabetes. We review evidence that receptor for advanced glycation end products (RAGE), via its interaction with ligands, serves as a cofactor exacerbating diabetic vascular disease. RAGE is a member of the immunoglobulin superfamily of cell surface molecules with a diverse repertoire of ligands reminiscent of pattern recognition receptors. In the diabetic milieu, two classes of RAGE ligands, products of nonenzymatic glycoxidation and S100 proteins, appear to drive receptor-mediated cellular activation and, potentially, acceleration of vascular disease. PMID- 15663912 TI - Novel features of nitric oxide, endothelial nitric oxide synthase, and atherosclerosis. AB - There is a complex pathophysiologic scenario involving nitric oxide (NO), endothelial nitric oxide synthase (eNOS), and the development of atherosclerosis and unstable atheroma. Endothelial damage induced by atherosclerosis leads to the reduction in bioactivity of eNOS with subsequent impaired release of NO. An important mechanism is local enhanced degradation of NO by increased generation of reactive oxygen species and other free radicals, with subsequent cascade of oxidation-sensitive mechanisms in the arterial wall. Novel molecular approaches have resulted in the development of new strains of mice lacking eNOS. These experimental models will help to understand how to implement NO-based therapies against atherosclerosis. L-arginine, the precursor of NO, has demonstrated beneficial effects in atherosclerosis and disturbed shear stress. The target or goal for new drugs should be the complete restoration of NO-mediated signaling pathways in atherosclerotic arteries. PMID- 15663913 TI - Peripheral arterial disease: clinical assessment and indications for revascularization in the patient with diabetes. AB - Peripheral arterial disease (PAD) is an under-recognized complication of diabetes. Recently, prevalence estimates in patients with diabetes over 50 years of age have been placed at 25% to 30%. The main reason for under-reporting is the largely asymptomatic nature of PAD in diabetes. Nonetheless, it is important to diagnose PAD because it is a marker of systemic atherosclerosis with excess cardiovascular risk, and it may identify a patient who may develop progressive disability and risk of limb loss. The most sensitive and specific diagnostic tool is an ankle-brachial index. Imaging studies are performed in patients who are candidates for revascularization. The most durable and effective revascularization procedure for PAD in diabetes is surgical bypass with saphenous vein as the conduit. Endovascular interventions are best used in patients with proximal disease with short-segment stenoses. The indications for revascularization have been immutable for decades, namely rest pain, ischemic ulceration, or gangrene. Presently, clinicians would include "selected" patients with intermittent claudication who have disabling symptoms and proximal disease above the inguinal ligament. PMID- 15663914 TI - FFAs: do they play a role in vascular disease in the insulin resistance syndrome? AB - The insulin resistance syndrome, otherwise known as the metabolic syndrome, describes a cluster of cardiovascular and metabolic abnormalities, which are strongly associated with overweight and obesity. The importance of the syndrome is due to its increased rates of cardiovascular morbidity and mortality. Insulin resistance is also characterized by elevated free fatty acid (FFA) levels. In otherwise healthy human subjects, elevation of FFA impairs endothelial function. This appears to be largely the result of blunting of nitric oxide-dependent tone, most likely at the level of the endothelial isoform of nitric oxide synthase (eNOS). Some of the potential mediatory mechanisms include oxidative stress, proinflammatory cytokines, C-reactive protein, or endogenous inhibitors of eNOS. Regardless of the mechanism(s) that mediates the effects of increased FFA on the vasculature, impaired vascular function is likely to account, at least in part, for the increase in cardiovascular mortality in subjects with the insulin resistance syndrome. PMID- 15663915 TI - ACE inhibitors and mineralocorticoid receptor blockade in patients with congestive heart failure. AB - The two major outcome trials on the combination of angiotensin-converting enzyme (ACE) inhibitors and mineralocorticoid receptor (MR) antagonists in heart failure are RALES (Randomized Aldactone Evaluation Study) and EPHESUS (Eplerenone Post Acute Myocardial Infarction Heart Failure Efficacy and Survival Study). There have also been studies in essential hypertension, and in diabetic hypertensive patients, on the cardiac and renal effects of ACE inhibitors and MR antagonists, individually and in combination. In the clinical studies on heart failure, in outcome trials and the smaller studies using surrogate end points, a combination of ACE inhibition and MR blockade is superior to ACE inhibition alone, and in the hypertension studies to either agent alone. Some insight into their distinct sites of protective action may be gained from studies on experimental animal preparations. The principal caveat in the use of combination therapy is the possibility of hyperkalemia, which should be minimal in patients with creatine clearance greater than 30 mL/min and with the low doses of MR antagonist shown to be effective in outcome trials. PMID- 15663918 TI - Interactions between obstructive sleep apnea and the metabolic syndrome. AB - The metabolic syndrome, an emerging public health problem, represents a constellation of cardiovascular risk factors. It has been suggested that the presence of obstructive sleep apnea (OSA) may increase the risk of developing some of the features of the metabolic syndrome, including hypertension, insulin resistance, and type 2 diabetes. In this article, we discuss the parallels between the metabolic syndrome and obstructive sleep apnea and describe possible OSA-related factors that may contribute to the metabolic syndrome, specifically the roles of obesity, hypertension, dyslipidemia, sex hormones, inflammation, vascular dysfunction, leptin, insulin resistance, and sleep deprivation. PMID- 15663917 TI - PPAR ligands: potential therapies for metabolic syndrome. AB - Metabolic syndrome (MS), a condition characterized by multiple related clinical disorders including insulin resistance, central obesity, hyperlipidemia, hypertension, and heart disease, is an increasingly prevalent disease in industrialized societies. The intense research interest in the peroxisome proliferator-activated receptors (PPARs), by both the pharmaceutical industry and academia, stems largely from the well-documented therapeutic actions of their synthetic agonists in alleviating several of the maladies associated with MS. This report focuses on the current understanding of the mechanisms of action of PPAR agents and their clinical use in the context of MS. PMID- 15663919 TI - Diabetes and brain aging: epidemiologic evidence. AB - Increasingly, data from epidemiologic studies suggest diabetes is a risk factor in old age for brain aging, including cognitive impairment and dementia. These associations may reflect a direct effect on the brain of hyperglycemia, or the effects of the diabetes-associated comorbidities of hypertension, dyslipidemia, or hyperinsulinemia. Epidemiologic data on diabetes and brain aging are reviewed. A brief overview is also given of the physiologic mechanisms supporting the epidemiologic data. PMID- 15663920 TI - Erectile dysfunction: interrelationship with the metabolic syndrome. AB - Erectile dysfunction (ED) is more commonly seen in men with various components of the metabolic syndrome (a constellation of various cardiovascular and diabetes risk factors). ED can be considered as a risk marker of the metabolic syndrome and its associated conditions. The patient with ED should be thoroughly evaluated for coexisting vascular disease. Any cardiovascular risk factors should be modified or treated (ie, smoking, diabetes, hypertension, and hyperlipidemia). Endothelial dysfunction is a major unifying etiology for many of the aspects of the metabolic syndrome, especially diabetes and cardiovascular disease. It also plays a major role in ED. The multifactorial etiology of ED, especially in patients with the metabolic syndrome, increases the complexity of managing this problem so clinicians need to be aware of the underlying pathophysiology to ensure the best possible outcomes in management. PMID- 15663921 TI - The evolving role of inflammation in obesity and the metabolic syndrome. AB - Advances in adipose tissue biology over the past 10 years have led to an improved understanding of the mechanisms linking obesity with the metabolic syndrome and other complications. Obesity is characterized by a chronic, systemic low-grade state of inflammation. Biomarkers of inflammation, such as the leukocyte count, tumor necrosis factor-alpha (TNF-alpha), interleukin 6 (IL-6), and C-reactive protein, are increased in obesity, associated with insulin resistance, and predict the development of type 2 diabetes and cardiovascular disease. It is now clear that the adipocyte is an active participant in the generation of the inflammatory state in obesity. Adipocytes secrete a variety of cytokines, including IL-6 and TNF-alpha, that promote inflammation. Moreover, recent studies suggest that obesity is associated with an increase in adipose tissue macrophages, which also participate in the inflammatory process through the elaboration of cytokines. An improved understanding of the role of adipose tissue in the activation of inflammatory pathways may suggest novel treatment and prevention strategies aimed at reducing obesity-associated morbidities and mortality. PMID- 15663922 TI - The high-resolution Structure of LeuB (Rv2995c) from Mycobacterium tuberculosis. AB - The crystal structure of the enzyme 3-isopropylmalate dehydrogenase (IPMDH) from Mycobacterium tuberculosis (LeuB, Mtb-IPMDH, Rv2995c) without substrate or co factor was determined at 1.65 A resolution, which is the highest resolution reported for an IPMDH to date. The crystals contain two functional dimers in the asymmetric unit in an arrangement close to a tetramer of D2 symmetry. Despite the absence of a substrate or inhibitor bound to the protein, the structure of the monomer resembles the previously observed closed form of the enzyme more closely than the open form. A comparison with the substrate complex of IPMDH from Thiobacillus ferrooxidans and the co-factor complex of the Thermus thermophilus enzyme revealed a close relationship of the active-site architecture between the various bacterial enzymes. The inhibitor O-isobutenyl oxalylhydroxamate was found to bind to the active site of IPMDH in a mode similar to the substrate isopropylmalate. PMID- 15663923 TI - Localization of a critical interface for helical rod formation of bacterial adhesion P-pili. AB - Pyelonephritic Escherichia coli cause urinary tract infections that involve the kidneys. Initiation of infection is dependent on P-pili expressed on the bacterial surface. In this work, an essential interface for assembly of the helical rod structure of P-pili has been located on the major pilin subunit, PapA. Based on primary sequence alignment, secondary structure analysis, and quaternary structure modeling of the PapA subunit, we predicted the location of a site that is critical for in vivo assembly of the native macromolecular structure of P-pili. A rigid helical rod of PapA subunits comprising most of the pilus length is stabilized by n to n+3 subunit-subunit interactions, and is important for normal function of these pili. Using site-directed mutagenesis, ultrastructural analysis by electron cryomicroscopy, immunocytochemistry, and molecular modeling we show that residues 106-109 (Asn, Gly, Ala, Gly) are essential for assembly of native P-pilus filaments. Mutation of these residues disrupts assembly of the native P-pilus helix. Extended fibrillar structures do still assemble, verifying that n to n+1 subunit-subunit interactions are maintained in the mutant fiber morphology. Observation of this fibrillar morphology in the mutant fiber was predicted by our modeling studies. These mutant P-pili data validate the predictive value of our model for understanding subunit-subunit interactions between PapA monomers. Alteration of the pilus structure from a 7-8 nm helical rod to a 2 nm fibrillar structure may compromise the ability of these bacteria to adhere and remain bound to the host cell, thus providing a possible therapeutic target for antimicrobial drugs. PMID- 15663924 TI - Conformational changes observed in enzyme crystal structures upon substrate binding. AB - The theory of induced fit predicts that enzymes undergo conformational changes as they bind their substrate. We have analysed the structures of 60 different enzymes to see if conformational changes are observed between the apo form, and the substrate (or substrate analog) bound form. In each enzyme the residues responsible for catalysis and substrate binding are known and are examined to see how the active site area is affected by conformational changes. Surprisingly, we find that induced fit motions in most enzymes is very small (usually 1 A RMSD between the apo and substrate-bound forms across the whole protein). We also find that there is a significant difference between the motions undergone by the binding residues and those undergone by the catalytic residues. The binding residues tend to exhibit larger backbone motions, but both binding and catalytic residues show the same, considerable, amount of side-chain flexibility. Knowing the extent of induced fit in enzymes is important for our understanding of the principles of enzyme catalysis and also for improving ligand docking and structural template searching. PMID- 15663925 TI - PfMyb1, a Plasmodium falciparum transcription factor, is required for intra erythrocytic growth and controls key genes for cell cycle regulation. AB - During the complex life cycle of Plasmodium falciparum, divided between mosquito and human hosts, the regulation of morphologic changes implies a fine control of transcriptional regulation. Transcriptional control, however, and in particular its molecular actors, transcription factors and regulatory motifs, are as yet poorly described in Plasmodium. In order to decipher the molecular mechanisms implicated in transcriptional regulation, a transcription factor belonging to the tryptophan cluster family was studied. In a previous work, the PfMyb1 protein, contained in nuclear extracts, was shown to have DNA binding activity and to interact specifically with myb regulatory elements. We used long pfmyb1 double stranded RNA (dsRNA) to interfere with the cognate messenger expression. Parasite cultures treated with pfmyb1 dsRNA exhibited a 40% growth inhibition when compared with either untreated cultures or cultures treated with unrelated dsRNA, and parasite mortality occurred during trophozoite to schizont transition. In addition, the pfmyb1 transcript and protein decreased by as much as 80% in treated trophozoite cultures at the time of their maximum expression. The global effect of this partial loss of transcript and protein was investigated using a thematic DNA microarray encompassing genes involved in signal transduction, cell cycle and transcriptional regulation. SAM software enabled us to identify several genes that were differentially expressed and probably directly or indirectly under the control of PfMyb1. Using chromatin immuno-precipitation, we demonstrated that PfMyb1 binds, within the parasite nuclei, to several promoters and therefore participates directly in the transcriptional regulation of the corresponding genes. This study provides the first evidence of a regulation network involving a Plasmodium transcription factor. PMID- 15663926 TI - Asymmetric allosteric activation of the symmetric ArgR hexamer. AB - Hexameric arginine repressor, ArgR, bound to L-arginine serves both as the master transcriptional repressor/activator at diverse regulons in a wide range of bacteria and as a required cofactor for resolution of ColE1 plasmid multimers. Multifunctional ArgR is thus unusual in possessing features of specific gene regulators, global regulators, and non-specific gene organizers; its closest functional analog is probably CAP, the cyclic AMP receptor/activator protein. Isothermal titration calorimetry, surface plasmon resonance, and proteolysis indicate that binding of a single L-argine [corrected] per ArgR hexamer triggers a global conformation [corrected] change and resets the affinities of the remaining five sites, making them 100-fold weaker. The analysis suggests a novel thermodynamic signature for this mechanism of activation. PMID- 15663927 TI - Crystal structure of leucyl-tRNA synthetase from the archaeon Pyrococcus horikoshii reveals a novel editing domain orientation. AB - The editing domains of the closely homologous leucyl, isoleucyl, and valyl-tRNA synthetases (LeuRS, IleRS, and ValRS, respectively) contribute to accurate aminoacylation, by hydrolyzing misformed non-cognate aminoacyl-tRNAs. The editing domain is inserted at the same point of the sequence in IleRS, ValRS, and the archaeal/eukaryal LeuRS, but at a distinct point in the bacterial LeuRS. Here, we showed that LeuRS from the archaeon Pyrococcus horikoshii has editing activity against the nearly cognate isoleucine. The conserved Asp332 in the editing domain is crucial for this activity. A deletion mutant lacking the C-terminal region has only negligible aminoacylation activity, but retains the full activity of adenylate synthesis and editing. We determined the crystal structure of this editing-active, truncated form of P.horikoshii LeuRS at 2.1 A resolution. The structure revealed that it has a novel editing domain orientation. The editing domain of P.horikoshii LeuRS is rotated by approximately 180 degrees (rotational state II), with the two-beta-stranded linker untwisted by a half-turn, as compared to those in IleRS and ValRS (rotational state I). This editing domain rotational state in the archaeal LeuRS is similar to that in the bacterial LeuRS. However, because of the insertion point difference, the orientation of the editing domain relative to the enzyme core in the archaeal LeuRS differs completely from that in the bacterial LeuRS. An insertion region specific to the archaeal/eukaryal LeuRS editing domains interacts with the enzyme core and stabilizes the unique orientation. Thus, we established that there are three types of editing domain orientations relative to the enzyme core, depending on the combination of the editing domain insertion point (i or ii) and the rotational state (I or II): [i, I] for IleRS and ValRS, [ii, II] for the bacterial LeuRS, and now [i, II] for the archaeal/eukaryal LeuRS. PMID- 15663928 TI - Monitoring uncharged tRNA during transcription of the Bacillus subtilis glyQS gene. AB - Expression of the Bacillus subtilis glyQS gene, encoding glycyl-tRNA synthetase, depends on stabilization of an antiterminator element during transcription of the 5' region of the mRNA by binding of uncharged tRNA(Gly). The glyQS gene is a member of the T box family of genes, all of which are involved in generation of charged tRNA. Each gene in this family exhibits an increase in readthrough of a termination signal located upstream of the start of the coding sequence in response to a decrease in the ratio of charged to uncharged tRNA. Many structural features of T box RNAs that are necessary for tRNA-dependent antitermination have been defined, but little is known about the timing or sequence of events that lead to a productive interaction with uncharged tRNA and discrimination against charged tRNA. To investigate these issues, transcription complexes were blocked artificially at specific positions along the leader sequence and tested for the ability to recognize tRNA. Although the sequence element that binds the tRNA anticodon is located more than 100 nt before the termination signal, complexes with nascent transcripts extending to just upstream of the termination site were still competent for antitermination. This result indicates that the transcript can fold into a receptive structure in the absence of the tRNA, and that tRNA is not necessary prior to this point. A mimic of charged tRNA(Gly) inhibited antitermination by uncharged tRNA unless the leader RNA-tRNA(Gly) complexes contained the complete antiterminator. These results suggest that the transcription complex can interact with either uncharged or charged tRNA until it approaches the termination point, allowing maximal flexibility in monitoring the ratio of charged to uncharged tRNA. PMID- 15663929 TI - DBC2 significantly influences cell-cycle, apoptosis, cytoskeleton and membrane trafficking pathways. AB - The tumor suppressor DBC2 belongs to a previously uncharacterized gene family, RHOBTB (Bric-a-brac, Tramtrack, Broad-complex). The biological roles of RHOBTB proteins, including DBC2, remain unclear. To understand the physiological functions of DBC2, a global approach was applied. Expression of DBC2 was manipulated in HeLa cells and RNA profiling of the cells was performed by microarray analyses. DBC2 was introduced into HeLa cells by a mammalian expression vector with a constitutive promoter. DBC2 knockdown was achieved by RNA interference with small interfering RNA. RNA profiles of these samples were performed by microarray analysis using Affymetrix GeneChip HG-U133A 2.0. The microarray data were analyzed by Microarray Suite 5.0 (MAS 5.0) and Robust Multichip Average (RMA). A list of genes whose expression was significantly altered (p<0.001) was generated and overlaid onto a cellular pathway map in the Ingenuity Systems' Pathway Knowledge Base (Winter'04 Release). Two networks were found to react substantially to DBC2 expression; namely, more than half of participating genes are affected. One of the networks regulates cell growth through cell-cycle control and apoptosis. The other network is related to cytoskeleton and membrane trafficking. Our findings suggest that the biological roles of DBC2 are related directly and/or indirectly to these cellular machineries. PMID- 15663930 TI - Shape-specific nucleotide binding of single-stranded RNA by the GLD-1 STAR domain. AB - Proteins containing the STAR RNA-binding domain fulfill vital roles in RNA biogenesis, yet a detailed understanding of STAR domain RNA binding specificity is lacking. In Caenorhabditis elegans, the STAR protein GLD-1 directly binds the 28 nucleotide recognition element TGE within the 3' untranslated region of tra-2 mRNA. The GLD-1:TGE interaction promotes translational silencing of tra-2 mRNA, marking a pivotal event in the spermatogenesis to oogenesis switch in C.elegans hermaphrodites. By measuring the binding affinities of both GLD-1 and TGE mutants, we have explored the molecular determinants of STAR domain specificity. Site-directed GLD-1 mutants were guided by sequence homology with human splicing factor 1 (SF1), for which an RNA:protein complex structure is available in the work done by Liu et al. The RNA binding affinity of 11 mutant GLD-1 proteins was measured, and their binding specificity was assessed with a series of TGE RNAs containing natural or modified nucleotides. This combinatorial analysis of both RNA and protein mutants revealed a diverse array of specificities of individual nucleotide-binding pockets along the interface. At nucleotide position 18, adenosine appears to be specified by the overall shape of a pocket lined with aliphatic side-chains. At position 19, the high preference for cytidine is dependent on both the length of an amino acid side-chain and the identity of terminal functional groups. The nucleotide 21 binding pocket exhibits low discrimination for cytidine, and accommodates most nucleobases. The highly hydrophobic binding interface and apparent small number of hydrogen bonding read out interactions at these positions is consistent with our finding that few amino acids seem to function individually in establishing binding specificity. Rather, specificity is conferred by the shape of the nucleotide-binding pocket. Our data provide the first detailed, quantitative analysis of the STAR domain, and highlight features of STAR:RNA recognition that are distinct among single stranded RNA-binding proteins. PMID- 15663931 TI - Structural snapshots of the KMSKS loop rearrangement for amino acid activation by bacterial tyrosyl-tRNA synthetase. AB - Tyrosyl-tRNA synthetase (TyrRS) has been studied extensively by mutational and structural analyses to elucidate its catalytic mechanism. TyrRS has the HIGH and KMSKS motifs that catalyze the amino acid activation with ATP. In the present study, the crystal structures of the Escherichia coli TyrRS catalytic domain, in complexes with l-tyrosine and a l-tyrosyladenylate analogue, Tyr-AMS, were solved at 2.0A and 2.7A resolution, respectively. In the Tyr-AMS-bound structure, the 2' OH group and adenine ring of the Tyr-AMS are strictly recognized by hydrogen bonds. This manner of hydrogen-bond recognition is conserved among the class I synthetases. Moreover, a comparison between the two structures revealed that the KMSKS loop is rearranged in response to adenine moiety binding and hydrogen-bond formation, and the KMSKS loop adopts the more compact ("semi-open") form, rather than the flexible, open form. The HIGH motif initially recognizes the gamma phosphate, and then the alpha and gamma-phosphates of ATP, with a slight rearrangement of the residues. The other residues around the substrate also accommodate the Tyr-AMS. This induced-fit form presents a novel "snapshot" of the amino acid activation step in the aminoacylation reaction by TyrRS. The present structures and the T.thermophilus TyrRS ATP-free and bound structures revealed that the extensive induced-fit conformational changes of the KMSKS loop and the local conformational changes within the substrate binding site form the basis for driving the amino acid activation step: the KMSKS loop adopts the open form, transiently shifts to the semi-open conformation according to the adenosyl moiety binding, and finally assumes the rigid ATP-bound, closed form. After the amino acid activation, the KMSKS loop adopts the semi-open form again to accept the CCA end of tRNA for the aminoacyl transfer reaction. PMID- 15663932 TI - Defining the basis for the specificity of aminoglycoside-rRNA recognition: a comparative study of drug binding to the A sites of Escherichia coli and human rRNA. AB - 2-Deoxystreptamine (2-DOS) aminoglycoside antibiotics exert their antimicrobial activities by targeting the decoding region A site of the rRNA and inhibiting protein synthesis. A prokaryotic specificity of action is critical to therapeutic utility of 2-DOS aminoglycosides as antibiotics. Here, isothermal titration calorimetry (ITC) and fluorescence studies are presented that provide insight into the molecular basis for this prokaryotic specificity of action. Specifically, the rRNA binding properties of the 2-DOS aminoglycosides paromomycin and G418 (geneticin) are compared, using both human and Escherichia coli rRNA A site model oligonucleotides as drug targets. Paromomycin and G418 differ with respect to their specificities of action, with only paromomycin exhibiting a specificity for prokaryotic versus human ribosomes. G418 binds to both the human and E. coli rRNA A sites with a markedly lower affinity than paromomycin, with the affinities of both drugs for the human rRNA A site being lower than those they exhibit for the E. coli rRNA A site. Paromomycin induces the destacking of the base at position 1492 (by E. coli numbering) upon binding to the E. coli rRNA A site, but not the human rRNA A site. By contrast, the binding of G418 induces the destacking of base 1492 when either rRNA A site serves as the drug target. In the aggregate, these results suggest that binding induced base destacking at the rRNA A site is a critical factor in determining the prokaryotic specificity of aminoglycoside action, with binding affinity for the A site being of secondary importance. PMID- 15663933 TI - Specific contributions of histone tails and their acetylation to the mechanical stability of nucleosomes. AB - The distinct contributions of histone tails and their acetylation to nucleosomal stability were examined by mechanical disruption of individual nucleosomes in a single chromatin fiber using an optical trap. Enzymatic removal of H2A/H2B tails primarily decreased the strength of histone-DNA interactions located approximately +/-36bp from the dyad axis of symmetry (off-dyad strong interactions), whereas removal of the H3/H4 tails played a greater role in regulating the total amount of DNA bound. Similarly, nucleosomes composed of histones acetylated to different degrees by the histone acetyltransferase p300 exhibited significant decreases in the off-dyad strong interactions and the total amount of DNA bound. Acetylation of H2A/H2B appears to play a particularly critical role in weakening the off-dyad strong interactions. Collectively, our results suggest that the destabilizing effects of tail acetylation may be due to elimination of specific key interactions in the nucleosome. PMID- 15663934 TI - Crystal structure of a free kappaB DNA: insights into DNA recognition by transcription factor NF-kappaB. AB - The dimeric NF-kappaB transcription factors regulate gene expression by recognizing specific DNA sequences located within the promoters of target genes. The DNA sequences, referred to as kappaB DNA, are divided into two broad classes. Class I kappaB DNA binds optimally to p50 and p52 NF-kappaB subunits, while class II kappaB DNAs are recognized specifically by the NF-kappaB subunits c-Rel and p65. We determined the X-ray crystal structure of a class II kappaB DNA sequence at 1.60 A resolution. This structure provides a detailed picture of kappaB DNA hydration, counter ion binding, and conformation in the absence of NF-kappaB binding partner. X-ray structures of both class I and class II kappaB DNA bound to NF-kappaB dimers were determined previously. Additionally, the NMR solution structure of a class I kappaB DNA is known. Comparison of the protein-bound and unbound kappaB DNA structures reveals that the free form of both classes approximates ideal B-form DNA more closely. Local geometries about specific DNA bases differ significantly upon binding to NF-kappaB. This is particularly evident at the 5'-GG/CC base-pairs; a signature of NF-kappaB specific DNA binding sequences. Differential phosphate group conformations, minor groove widths, buckle, twist, and tilt angles are observed between bound and unbound kappaB DNA. We observe that the presence of an extra G:C base-pair, 5'- to the GGA sequence in class I kappaB DNA, alters the geometry of the two internal G:C base-pairs within the GGGA tetranucleotide, which explains, at least in part, the structural basis for distinct NF-kappaB dimer recruitment by the two different classes of kappaB DNA. Together, these observations suggest that NF-kappaB dimers recognize specific structural features of kappaB DNA in order to make sequence-specific complexes. PMID- 15663935 TI - Target-site preferences of Sleeping Beauty transposons. AB - The Sleeping Beauty (SB) transposon is a Tc1/mariner family transposon that has applications in vertebrate animals for gene transfer, gene-tagging, and human gene therapy. In this study, we analyzed the target-site preferences of the SB transposon. At the genomic level, integration of SB transposons with respect to genes (exons and introns) and intergenic regions appears fairly random but not on a micro-scale. Although there appears to be a consensus sequence around the vicinity of the target sites, the primary sequence is not the determining factor for target selection. When integrations were examined over a limited topography, the sites used most often for integration did not match the consensus sequence. Rather, a unique deformation inherent in the sequence may be a recognition signal for target selection. The deformation is characterized by an angling of the target site such that the axis around the insertion site is off-center, the rotation of the helix (twisting) is non-uniform and there is an increase in the distance between the central base-pairs. Our observations offer several hypothetical insights into the transposition process. Our observations suggest that particular deformations of the double helix predicted by the V(step) algorithm can distinguish TA sites that vary by about 16-fold in their preferences for accommodating insertions of SB transposons. PMID- 15663936 TI - N and C-terminal sub-regions in the c-Myc transactivation region and their joint role in creating versatility in folding and binding. AB - The proto-oncogene c-myc governs the expression of a number of genes targeting cell growth and apoptosis, and its expression levels are distorted in many cancer forms. The current investigation presents an analysis by proteolysis, circular dichroism, fluorescence and Biacore of the folding and ligand-binding properties of the N-terminal transactivation domain (TAD) in the c-Myc protein. A c-Myc sub region comprising residues 1-167 (Myc1-167) has been investigated that includes the unstructured c-Myc transactivation domain (TAD, residues 1-143) together with a C-terminal segment, which appears to promote increased folding. Myc1-167 is partly helical, binds both to the target proteins Myc modulator-1 (MM-1) and TATA box-binding protein (TBP), and displays the characteristics of a molten globule. Limited proteolysis divides Myc1-167 in two halves, by cleaving in a predicted linker region between two hotspot mutation regions: Myc box I (MBI) and Myc box II (MBII). The N-terminal half (Myc1-88) is unfolded and does not alone bind to target proteins, whereas the C-terminal half (Myc92-167) has a partly helical fold and specifically binds both MM-1 and TBP. Although this might suggest a bipartite organization in the c-Myc TAD, none of the N and C-terminal fragments bind target protein with as high affinity as the entire Myc1-167, or display molten globule properties. Furthermore, merely linking the MBI with the C terminal region, in Myc38-167, is not sufficient to achieve binding and folding properties as in Myc1-167. Thus, the entire N and C-terminal regions of c-Myc TAD act in concert to achieve high specificity and affinity to two structurally and functionally orthogonal target proteins, TBP and MM-1, possibly through a mechanism involving molten globule formation. This hints towards understanding how binding of a range of targets can be accomplished to a single transactivation domain. PMID- 15663937 TI - Allosteric network of cAMP-dependent protein kinase revealed by mutation of Tyr204 in the P+1 loop. AB - Previous studies on the catalytic subunit of cAMP-dependent protein kinase (PKA) identified a conserved interaction pair comprised of Tyr204 from the P+1 loop and Glu230 at the end of the alphaF-helix. Single-point mutations of Tyr204 to Ala (Y204A) and Glu230 to Gln (E230Q) both resulted in alterations in enzymatic kinetics. To understand further the molecular basis for the altered kinetics and the structural role of each residue, we analyzed the Y204A and the E230Q mutants using hydrogen/deuterium (H/D) exchange coupled with mass spectrometry and other biophysical techniques. The fact that the mutants exhibit distinct molecular properties, supports previous hypotheses that these two residues, although in the same interaction node, contribute to the same enzymatic functions through different molecular pathways. The Tyr204 mutation appears to affect the dynamic properties, while the Glu230 mutation affects the surface electrostatic profile of the enzyme. Furthermore, H/D exchange analysis defines the dynamic allosteric range of Tyr204 to include the catalytic loop and three additional distant surface regions, which exhibit increased deuterium exchange in the Y204A but not the E230Q mutant. Interestingly, these are the exact regions that previously showed decreased deuterium exchange upon binding of the RIalpha regulatory subunit of PKA. We propose that these sites, coupled with the P+1 loop through Tyr204, represent one of the major allosteric networks in the kinase. This coupling provides a coordinated response for substrate binding and enzyme catalysis. H/D exchange analysis also further defines the stable core of the catalytic subunit to include the alphaE, alphaF and alphaH-helix. All these observations lead to an interesting new way to view the structural architecture and allosteric conformational regulation of the protein kinase molecule. PMID- 15663938 TI - An integrative approach to gain insights into the cellular function of human ataxin-2. AB - Spinocerebellar ataxia type 2 (SCA2) is a hereditary neurodegenerative disorder caused by a trinucleotide expansion in the SCA2 gene, encoding a polyglutamine stretch in the gene product ataxin-2 (ATX2), whose cellular function is unknown. However, ATX2 interacts with A2BP1, a protein containing an RNA-recognition motif, and the existence of an interaction motif for the C-terminal domain of the poly(A)-binding protein (PABC) as well as an Lsm (Like Sm) domain in ATX2 suggest that ATX2 like its yeast homolog Pbp1 might be involved in RNA metabolism. Here, we show that, similar to Pbp1, ATX2 suppresses the petite (pet-) phenotype of Deltamrs2 yeast strains lacking mitochondrial group II introns. This finding points to a close functional relationship between the two homologs. To gain insight into potential functions of ATX2, we also generated a comprehensive protein interaction network for Pbp1 from publicly available databases, which implicates Pbp1 in diverse RNA-processing pathways. The functional relationship of ATX2 and Pbp1 is further corroborated by the experimental confirmation of the predicted interaction of ATX2 with the cytoplasmic poly(A)-binding protein 1 (PABP) using yeast-2-hybrid analysis as well as co-immunoprecipitation experiments. Immunofluorescence studies revealed that ATX2 and PABP co-localize in mammalian cells, remarkably, even under conditions in which PABP accumulates in distinct cytoplasmic foci representing sites of mRNA triage. PMID- 15663939 TI - Imaging calpain protease activity by multiphoton FRET in living mice. AB - Constant efforts are ongoing for the development of new imaging methods that allow the investigation of molecular processes in vivo. Protein-protein interactions, enzymatic activities and intracellular Ca2+ fluxes, have been resolved in cultured cells using a variety of fluorescence resonance energy transfer (FRET) detection methods. However, FRET has not been used so far in conjunction with 3D intravital imaging. We evaluated here a combination of multiphoton microscopy (MPM), method of choice for non-destructive living tissue investigation, and FRET imaging to monitor calpain proteolytic activity in living mice muscle. We show that kinetics of ubiquitous calpains activation can be efficiently and quantitatively monitored in living mouse tissues at cellular level with a FRET-based indicator upon calcium influx. The ability to visualize calpain activity in living tissue offers a unique opportunity to challenge remaining questions on the biological functions of calpains and to evaluate the therapeutic potential of calpain inhibitors in many degenerative conditions. PMID- 15663941 TI - Catalytic mechanism of C-C hydrolase MhpC from Escherichia coli: kinetic analysis of His263 and Ser110 site-directed mutants. AB - C-C hydrolase MhpC (2-hydroxy-6-keto-nona-1,9-dioic acid 5,6-hydrolase) from Escherichia coli catalyses the hydrolytic C-C cleavage of the meta-ring fission product on the phenylpropionic acid catabolic pathway. The crystal structure of E. coli MhpC has revealed a number of active-site amino acid residues that may participate in catalysis. Site-directed mutants of His263, Ser110, His114, and Ser40 have been analysed using steady-state and stopped-flow kinetics. Mutants H263A, S110A and S110G show 10(4)-fold reduced catalytic efficiency, but still retain catalytic activity for C-C cleavage. Two distinct steps are observed by stopped-flow UV/Vis spectrophotometry, corresponding to ketonisation and C-C cleavage: H263A exhibits very slow ketonisation and C-C cleavage, whereas S110A and S110G exhibit fast ketonisation, an intermediate phase, and slow C-C cleavage. H114A shows only twofold-reduced catalytic efficiency, ruling out a catalytic role, but shows a fivefold-reduced K(M) for the natural substrate, and an ability to process an aryl-containing substrate, implying a role for His114 in positioning of the substrate. S40A shows only twofold-reduced catalytic efficiency, but shows a very fast (500 s(-1)) interconversion of dienol (317 nm) to dienolate (394 nm) forms of the substrate, indicating that the enzyme accepts the dienol form of the substrate. These data imply that His263 is responsible for both ketonisation of the substrate and for deprotonation of water for C-C cleavage, a novel catalytic role in a serine hydrolase. Ser110 has an important but non-essential role in catalysis, which appears not to be to act as a nucleophile. A catalytic mechanism is proposed involving stabilisation of reactive intermediates and activation of a nucleophilic water molecule by Ser110. PMID- 15663940 TI - High-affinity, peptide-specific T cell receptors can be generated by mutations in CDR1, CDR2 or CDR3. AB - The third complementarity-determining regions (CDR3s) of antibodies and T cell receptors (TCRs) have been shown to play a major role in antigen binding and specificity. Consistent with this notion, we demonstrated previously that high affinity, peptide-specific TCRs could be generated in vitro by mutations in the CDR3alpha region of the 2C TCR. In contrast, it has been argued that CDR1 and CDR2 are involved to a greater extent than CDR3s in the process of MHC restriction, due to their engagement of MHC helices. Based on this premise, we initiated the present study to explore whether higher affinity TCRs generated through mutations in these CDRs or other regions would lead to significant reductions in peptide specificity (i.e. the result of greater binding energy gained through interactions with major histocompatibility complex (MHC) helices). Yeast-display technology and flow sorting were used to select high-affinity TCRs from libraries of CDR mutants or random mutants. High-affinity TCRs with mutations in the first residue of the Valpha, CDR1, CDR2, or CDR3 were isolated. Unexpectedly, every TCR mutant, including those in CDR1 and CDR2, retained remarkable peptide specificity. Molecular modeling of various mutants suggested that such exquisite specificity may be due to: (1) enhanced electrostatic interactions with key peptide or MHC residues; or (2) stabilization of CDRs in specific conformations. The results indicate that the TCR is positioned so that virtually every CDR can contribute to the antigen-specificity of a T cell. The conserved diagonal docking of TCRs could thus orient each CDR loop to sense the peptide directly or indirectly through peptide-induced effects on the MHC. PMID- 15663942 TI - The structure of the C-C bond hydrolase MhpC provides insights into its catalytic mechanism. AB - 2-Hydroxy-6-ketonona-2,4-diene-1,9-dioic acid 5,6-hydrolase (MhpC) is a 62 kDa homodimeric enzyme of the phenylpropionate degradation pathway of Escherichia coli. The 2.1 A resolution X-ray structure of the native enzyme determined from orthorhombic crystals confirms that it is a member of the alpha/beta hydrolase fold family, comprising eight beta-strands interconnected by loops and helices. The 2.8 A resolution structure of the enzyme co-crystallised with the non hydrolysable substrate analogue 2,6-diketo-nona-1,9-dioic acid (DKNDA) confirms the location of the active site in a buried channel including Ser110, His263 and Asp235, postulated contributors to a serine protease-like catalytic triad in homologous enzymes. It appears that the ligand binds in two separate orientations. In the first, the C6 keto group of the inhibitor forms a hemi-ketal adduct with the Ser110 side-chain, the C9 carboxylate group interacts, via the intermediacy of a water molecule, with Arg188 at one end of the active site, while the C1 carboxylate group of the inhibitor comes close to His114 at the other end. In the second orientation, the C1 carboxylate group binds at the Arg188 end of the active site and the C9 carboxylate group at the His114 end. These arrangements implicated His114 or His263 as plausible contributors to catalysis of the initial enol/keto tautomerisation of the substrate but lack of conservation of His114 amongst related enzymes and mutagenesis results suggest that His263 is the residue involved. Variability in the quality of the electron density for the inhibitor amongst the eight molecules of the crystal asymmetric unit appears to correlate with alternative positions for the side-chain of His114. This might arise from half-site occupation of the dimeric enzyme and reflect the apparent dissociation of approximately 50% of the keto intermediate from the enzyme during the catalytic cycle. PMID- 15663943 TI - Structure of the mature P3-virus particle complex of cauliflower mosaic virus revealed by cryo-electron microscopy. AB - The cauliflower mosaic virus (CaMV) has an icosahedral capsid composed of the viral protein P4. The viral product P3 is a multifunctional protein closely associated with the virus particle within host cells. The best-characterized function of P3 is its implication in CaMV plant-to-plant transmission by aphid vectors, involving a P3-virion complex. In this transmission process, the viral protein P2 attaches to virion-bound P3, and creates a molecular bridge between the virus and a putative receptor in the aphid's stylets. Recently, the virion bound P3 has been suggested to participate in cell-to-cell or long-distance movement of CaMV within the host plant. Thus, as new data accumulate, the importance of the P3-virion complex during the virus life-cycle is becoming more and more evident. To provide a first insight into the knowledge of the transmission process of the virus, we determined the 3D structures of native and P3-decorated virions by cryo-electron microscopy and computer image processing. By difference mapping and biochemical analysis, we show that P3 forms a network around the capsomers and we propose a structural model for the binding of P3 to CaMV capsid in which its C terminus is anchored deeply in the inner shell of the virion, while the N-terminal extremity is facing out of the CaMV capsid, forming dimers by coiled-coil interactions. Our results combined with existing data reinforce the hypothesis that this coiled-coil N-terminal region of P3 could coordinate several functions during the virus life-cycle, such as cell-to-cell movement and aphid-transmission. PMID- 15663944 TI - Dynamics in the unfolded state of beta2-microglobulin studied by NMR. AB - Many proteins form amyloid-like fibrils in vitro under conditions that favour the population of partially folded conformations or denatured state ensembles. Characterising the structural and dynamic properties of these states is crucial towards understanding the mechanisms of self-assembly in amyloidosis. The aggregation of beta2-microglobulin (beta2m) into amyloid fibrils in vivo occurs in the condition known as dialysis-related amyloidosis (DRA) and the protein has been shown to form amyloid-like fibrils under acidic conditions in vitro. We have used a number of 1H-15N nuclear magnetic resonance (NMR) experiments in conjunction with site-directed mutagenesis to study the acid-unfolded state of beta2m. 15N NMR transverse relaxation experiments reveal that the acid-denatured ensemble, although predominantly unfolded at the N and C termini, contains substantial non-native structure in the central region of the polypeptide chain, stabilised by long-range interactions between aromatic residues and by the single disulphide bond. Relaxation dispersion studies indicate that the acid-unfolded ensemble involves two or more distinct species in conformational equilibrium on the micro- to millisecond time-scale. One of these species appears to be hydrophobically collapsed, as mutations in an aromatic-rich region of the protein, including residues that are solvent-exposed in the native protein, disrupt this structure and cause a consequent decrease in the population of this conformer. Thus, acid-unfolded beta2m consists of a heterogeneous ensemble of rapidly fluctuating species, some of which contain stable, non-native hydrophobic clusters. Given that amyloid assembly of beta2m proceeds with lag kinetics under the conditions of this study, a rarely populated species such as a conformer with non-native aromatic clustering could be key to the initiation of amyloidosis. PMID- 15663945 TI - The Escherichia coli twin-arginine translocation apparatus incorporates a distinct form of TatABC complex, spectrum of modular TatA complexes and minor TatAB complex. AB - The Tat system transports folded proteins across bacterial plasma and plant thylakoid membranes. To date, three key Tat subunits have been identified and mechanistic studies indicate the presence of two types of complex: a TatBC containing substrate-binding unit and a separate TatA complex. Here, we used blue native gel electrophoresis and affinity purification to study the nature of these complexes in Escherichia coli. Analysis of solubilized membrane shows that the bulk of TatB and essentially all of the TatC is found in a single 370kDa TatABC complex. TatABC was purified to homogeneity using an affinity tag on TatC and this complex runs apparently as an identical band. We conclude that this is the primary core complex, predicted to contain six or seven copies of TatBC together with a similar number of TatA subunits. However, the data indicate the presence of an additional form of Tat complex containing TatA and TatB, but not TatC; we speculate that this may be an assembly or disassembly intermediate of the translocator. The vast majority of TatA is found in separate complexes that migrate in blue-native gels as a striking ladder of bands with sizes ranging from under 100 kDa to over 500 kDa. Further analysis shows that the bands differ by an average of 34 kDa, indicating that TatA complexes are built largely, but possibly not exclusively, from modules of three or four TatA molecules. The range and nature of these complexes are similar in a TatC mutant that is totally inactive, indicating that the ladder of bands does not stem from ongoing translocation activity, and we show that purified TatA can self-assemble in vitro to form similar complexes. This spectrum of TatA complexes may provide the flexibility required to generate a translocon capable of transporting substrates of varying sizes across the plasma membrane in a folded state. PMID- 15663946 TI - Structural and energetic consequences of mutations in a solvated hydrophobic cavity. AB - The structural and energetic consequences of modifications to the hydrophobic cavity of interleukin 1-beta (IL-1beta) are described. Previous reports demonstrated that the entirely hydrophobic cavity of IL-1beta contains positionally disordered water. To gain a better understanding of the nature of this cavity and the water therein, a number of mutant proteins were constructed by site-directed mutagenesis, designed to result in altered hydrophobicity of the cavity. These mutations involve the replacement of specific phenylalanine residues, which circumscribe the cavity, with tyrosine, tryptophan, leucine and isoleucine. Using differential scanning calorimetry to determine the relative stabilities of the wild-type and mutant proteins, we found all of the mutants to be destabilizing. X-ray crystallography was used to identify the structural consequences of the mutations. No clear correlation between the hydrophobicities of the specific side-chains introduced and the resulting stabilities was found. PMID- 15663947 TI - Folding, stability and polymerization properties of FtsZ chimeras with inserted tubulin loops involved in the interaction with the cytosolic chaperonin CCT and in microtubule formation. AB - To attain its native conformation, the cytoskeletal protein tubulin needs the concourse of several molecular chaperones, among others the cytosolic chaperonin CCT. It has been previously described that denatured tubulin interacts with CCT in a quasi-folded conformation using several loops located throughout its sequence. These loops are also involved in microtubule formation and are absent in its prokaryote homologue FtsZ, which in vitro folds by itself and does not interact with CCT. Several FtsZ/tubulin chimeric proteins were generated by inserting consecutively one, two or three of the CCT-binding domains of tubulin into the corresponding sequence of FtsZ from Methanococccus jannaschii. The insertion of any of the CCT-binding loops generates in the FtsZ/tubulin chimeras the ability to interact with CCT. The accumulation of CCT-binding loops induces in the FtsZ/tubulin chimeras unfolding and refolding properties that are more similar to tubulin than to its prokaryote counterpart. Finally, the insertion of some of these loops generates in the FtsZ/tubulin chimeras more complex polymeric structures than those found for FtsZ. These results reinforce the notion that CCT has coevolved with tubulin to deal with the folding problems encountered by the eukaryotic protein with the appearance of the new sequences involved in microtubule formation. PMID- 15663948 TI - Effects of heme on the structure of the denatured state and folding kinetics of cytochrome b562. AB - Heme-linked proteins, such as cytochromes, are popular subjects for protein folding studies. There is the underlying question of whether the heme affects the structure of the denatured state by cross-linking it and forming other interactions, which would perturb the folding pathway. We have studied wild-type and mutant cytochrome b562 from Escherichia coli, a 106 residue four-alpha helical bundle. The holo protein apparently refolds with a half-life of 4 micros in its ferrous state. We have analysed the folding of the apo protein using continuous-flow fluorescence as well as stopped-flow fluorescence and CD. The apo protein folded much more slowly with a half-life of 270 micros that was unaffected by the presence of exogenous heme. We examined the nature of the denatured states of both holo and apo proteins by NMR methods over a range of concentrations of guanidine hydrochloride. The starting point for folding of the holo protein in concentrations of denaturant around the denaturation transition was a highly ordered native-like species with heme bound. Fully denatured holo protein at higher concentrations of denaturant consisted of denatured apo protein and free heme. Our results suggest that the very fast folding species of denatured holo protein is in a compact state, whereas the normal folding pathway from fully denatured holo protein consists of the slower folding of the apo protein followed by the binding of heme. These data should be considered in the analysis of folding of heme proteins. PMID- 15663950 TI - The relationship between domain duplication and recombination. AB - Protein domains represent the basic evolutionary units that form proteins. Domain duplication and shuffling by recombination are probably the most important forces driving protein evolution and hence the complexity of the proteome. While the duplication of whole genes as well as domain-encoding exons increases the abundance of domains in the proteome, domain shuffling increases versatility, i.e. the number of distinct contexts in which a domain can occur. Here, we describe a comprehensive, genome-wide analysis of the relationship between these two processes. We observe a strong and robust correlation between domain versatility and abundance: domains that occur more often also have many different combination partners. This supports the view that domain recombination occurs in a random way. However, we do not observe all the different combinations that are expected from a simple random recombination scenario, and this is due to frequent duplication of specific domain combinations. When we simulate the evolution of the protein repertoire considering stochastic recombination of domains followed by extensive duplication of the combinations, we approximate the observed data well. Our analyses are consistent with a stochastic process that governs domain recombination and thus protein divergence with respect to domains within a polypeptide chain. At the same time, they support a scenario in which domain combinations are formed only once during the evolution of the protein repertoire, and are then duplicated to various extents. The extent of duplication of different combinations varies widely and, in nature, will depend on selection for the domain combination based on its function. Some of the pair-wise domain combinations that are highly duplicated also recur frequently with other partner domains, and thus represent evolutionary units larger than single protein domains, which we term "supra-domains". PMID- 15663949 TI - Detection of a hidden folding intermediate of the third domain of PDZ. AB - The folding pathway of the third domain of PDZ from the synaptic protein PSD-95 was characterized using kinetic and equilibrium methods by monitoring the fluorescence signal from a Trp residue that is incorporated at a near-surface position. Kinetic folding of this domain showed multiple exponential phases, whereas unfolding showed a single exponential phase. The slow kinetic phases were attributed to isomerization of proline residues, since there are five proline residues in this domain. We found that the logarithms of the rate constants for the fast phase of folding and unfolding are linearly dependent on the concentrations of denaturant. The unfolding free energy derived from these rate constants at zero denaturant was close to the value measured using the equilibrium method, suggesting the absence of detectable sub-millisecond folding intermediates. However, native-state hydrogen exchange experiments detected a partially unfolded intermediate under native conditions. It was further confirmed by a protein engineering study. These data suggest that a hidden intermediate exists after the rate-limiting step in the folding of the third domain of PDZ. PMID- 15663951 TI - Structure of the armadillo repeat domain of plakophilin 1. AB - The p120ctn subfamily of armadillo domain proteins has roles in modulating intercellular adhesion by cadherin-containing junctions. We have determined the crystal structure of the arm repeat domain from plakophilin-1 (PKP1), a member of the p120ctn subfamily that is found in desmosomes. The structure reveals that the domain has nine instead of the expected ten arm repeats. A sequence predicted to be an arm repeat is instead a large insert which serves as a wedge that produces a significant bend in the overall domain structure. Structure-based sequence alignments indicate that the nine repeats and large insert are common to this subfamily of armadillo proteins. A prominent basic patch on the surface of the protein may serve as a binding site for partners of these proteins. PMID- 15663952 TI - A study of the membrane-water interface region of membrane proteins. AB - The most conspicuous structural characteristic of the alpha-helical membrane proteins is their long transmembrane alpha-helices. However, other structural elements, as yet largely ignored in statistical studies of membrane protein structure, are found in those parts of the protein that are located in the membrane-water interface region. Here, we show that this region is enriched in irregular structure and in interfacial helices running roughly parallel with the membrane surface, while beta-strands are extremely rare. The average amino acid composition is different between the interfacial helices, the parts of the transmembrane helices located in the interface region, and the irregular structures. In this region, hydrophobic and aromatic residues tend to point toward the membrane and charged/polar residues tend to point away from the membrane. The interface region thus imposes different constraints on protein structure than do the central hydrocarbon core of the membrane and the surrounding aqueous phase. PMID- 15663953 TI - Epidermal growth factor gene polymorphism is different between schizophrenia and lung cancer patients in Korean population. AB - Low incidence of cancer in schizophrenia is one of the interesting puzzles in psychiatric field over decades. Analysis of genetic difference between schizophrenia and lung cancer might provide us with possible clues to understand molecular mechanisms of pathophysiology of schizophrenia. Epidermal growth factor (EGF), one of the potent growth promoting factors, has been studied for its roles in cancer development. EGF is also known to be involved in cognitive function. In order to analyze the genetic difference between schizophrenia and lung cancer, polymorphism of EGF gene was studied from 174 schizophrenia patients, 122 lung cancer patients and 132 controls in Korean population. Genotype frequency analysis of EGF gene (AluI restriction site, 5'-UTR, rs4444903) in the EGF gene was studied. The genotype and allele frequencies of the AluI polymorphism showed significant differences between schizophrenia and lung cancer patients [p<0.0001; p<0.0001, odds ratio (95% CI), 0.3690 (0.2600-0.5236)]. When compared with controls, schizophrenia patients showed no significant differences from controls in genotype and allele frequencies [p=0.5151; p=0.3516, odds ratio (95% CI), 0.8589 (0.6235-1.1830)]. However, lung cancer patients showed significant differences from controls in genotype and allele frequencies [p<0.0001; p<0.0001, odds ratio (95% CI), 2.3275 (1.6082-3.3687)]. These results indicate that schizophrenia is not associated with AluI polymorphism of EGF gene and EGF gene polymorphism is different between schizophrenia and lung cancer patients. PMID- 15663954 TI - Penetrating brain injury leads to activation of ciliary neurotrophic factor receptors. AB - Endogenous injury response mechanisms likely reduce secondary neuronal loss following CNS trauma by activating growth factor receptors. Therefore, it is important to determine which growth factor receptors are activated in vivo by CNS trauma and which signal transduction pathways are affected in which cell types. We present a model of penetrating brain injury utilizing stereotaxic insertion of a fine needle. This procedure can be used to anatomically characterize injury response mechanisms through immediate, local application of pharmacological agents. We find, through immunohistochemistry, that injury of the rat facial motor nucleus leads to activation of STAT3, a neuronal survival factor, in the dendrites, nuclei and cytoplasm of the motor neurons. A similar response was observed with the trigeminal motor nucleus. Use of the ciliary neurotrophic factor (CNTF) receptor antagonist, AADH-CNTF, indicated that the STAT3 activation resulted largely, and perhaps entirely, from injury-induced activation of CNTF receptors. PMID- 15663955 TI - Different physiological properties in a pool of mandibular closer motor neurons in a caterpillar, Bombyx mori. AB - To determine whether a large pool of mandibular closer motor neurons in an insect consists of different functional groups, the physiological properties of the motor neurons were investigated by intracellular recording techniques. The mandibular closer muscle of a caterpillar, Bombyx mori, was innervated by at least 11 motor neurons with large somata and 2-3 neurons with small somata. The large 11 motor neurons could be divided into slow and fast types on the basis of thresholds of neurons and frequency of soma spikes by current injections. The fast motor neurons usually evoked relatively larger muscle potentials, whereas the slow neurons evoked smaller potentials. The slow neurons spiked while the mandible was held in a fully closed position, whereas the fast neurons did not. The slow neurons did not spike while the mandible was free to move. These results suggest different physiological properties among closer motor neurons, supporting the hypothesis that the closer motor neurons in a large neuron pool can be divided into different functional groups. PMID- 15663956 TI - Lack of expression of the liver-type glutaminase (LGA) mRNA in human malignant gliomas. AB - In the central nervous system (CNS), liver-type glutaminase (LGA) shows a unique nuclear localization suggesting its role in the regulation of transcription rather than in the cellular glutamine metabolism. RT-PCR analysis of RNA derived from postoperative tissue samples revealed the absence or only traces of LGA mRNA in all (9) cases of malignant gliomas (astrocytoma anaplasticum, AA, WHO grade III; glioblastoma multiforme, WHO grade IV) examined. The RNA was strongly expressed in the non-neoplastic tissue derived from the same patients (6 cases), and in most of the brain metastases from different organs (5 out of 7 cases). By contrast, the mRNAs coding for the kidney-type glutaminase (KGA) and its less ubiquitous isoform GAC, which catalyze degradation of the cytoplasmic pool of Gln, were expressed in all the tissues examined. The lack of LGA may be thus considered as a useful negative diagnostic marker of highly malignant gliomas in situ. PMID- 15663957 TI - Applications of random field theory to electrophysiology. AB - The analysis of electrophysiological data often produces results that are continuous in one or more dimensions, e.g., time-frequency maps, peri-stimulus time histograms, and cross-correlation functions. Classical inferences made on the ensuing statistical maps must control family wise error (FWE) when searching across the map's dimensions. In this paper, we borrow multiple comparisons procedures, established in neuroimaging, and apply them to electrophysiological data. These procedures use random field theory (RFT) to adjust p-values from statistics that are functions of time and/or frequency. This RFT adjustment for continuous statistical processes plays the same role as a Bonnferonni adjustment in the context of discrete statistical tests. Here, by analysing the time frequency decompositions of single channel EEG data we show that RFT adjustments can be used in the analysis of electrophysiological data and illustrate the advantages of this method over existing approaches. PMID- 15663959 TI - Neuroprotective effect of geranylgeranylacetone, a noninvasive heat shock protein inducer, on cerebral infarction in rats. AB - The present study evaluated the neuroprotective effect of geranylgeranylacetone (GGA), which is known as an antiulcer agent and more recently as a heat shock protein (HSP) inducer, against cerebral infarction induced by permanent left middle cerebral artery (MCA) occlusion. GGA was given orally in various regimens prior to MCA occlusion in rats. Pretreatment with a single oral GGA dose (800 mg/kg) 48 h before ischemia significantly attenuated cerebral infarction volume (81.7+/-18.4 mm3 versus 369.1+/-70.2 mm3; P<.01). A significant increase in HSP70 immunoreactivity was found in the neocortex in GGA-treated animals with or without ischemia. Pretreatment with a single oral dose of GGA provides an important tool for exploring the mechanisms of neuroprotection against cerebral ischemic neuronal damage. PMID- 15663958 TI - Transient expression of hypoxia-inducible factor-1 alpha and target genes in peripheral nerves from diabetic rats. AB - Decreased blood flow is one of the earliest physiological changes observed after the onset of either clinical or experimental diabetes. The reduction in blood flow is believed to lead to nerve hypoxia, which in conjunction with other metabolic alterations and degenerative processes in different nerve compartments, results in the dysfunction known as diabetic neuropathy. The transcriptional regulator hypoxia-inducible factor-1 alpha (HIF-1alpha) accumulates rapidly under hypoxic conditions and modulates the expression of several target genes that protect tissues against ischemia and infarction. At present it is unclear whether diabetic nerve injury results from an abnormal response of HIF-1alpha and its protective target genes. In the present study we have analyzed the expression and activity of HIF-1alpha and its target genes in diabetic nerves as a first step to determine their possible contribution to the development or maintenance of diabetic neuropathy. We observed a transient increase in the expression of HIF 1alpha that peaked between 4 and 6 weeks and declined 8 weeks after induction of experimental diabetes in rats. The increase in HIF-1alpha in diabetic nerves coincided with a similarly transient increase in the expression of several HIF 1alpha target genes including vascular endothelial growth factor, lactate dehydrogenase and erythropoietin, which subsided 8-10 weeks after induction of diabetes. These results suggest that the transient activation of neurotrophic and angiogenic genes, as opposed to a more sustained effect in response to the chronic injury, may be responsible for the alterations in nerve function and regeneration that characterize the diabetic neuropathy. PMID- 15663960 TI - Unaltered alpha-synuclein blood levels in juvenile Parkinsonism with a parkin exon 4 deletion. AB - We recently reported here that SNCA triplication results in a doubling in the amount of alpha-synuclein protein in blood from cases with hereditary Lewy body disease. This observation shows that alpha-synuclein levels in blood accurately reflect gene dosage, which we assume drives pathogenesis in these individuals. A previous report has suggested that parkin can affect alpha-synuclein metabolism in human brain. Here we have tested whether there is also an increase of alpha synuclein in autosomal recessive juvenile Parkinsonism (ARJP). We find there is not and discuss this result in terms of the putative relationships between alpha synuclein and parkin. PMID- 15663961 TI - Interleukin-6 protects cultured cerebellar granule neurons against glutamate induced neurotoxicity. AB - Cytokine interleukin-6 (IL-6) has been showed to be an important mediator of neuroimmune responses. However, effects of IL-6 in the central nervous system (CNS) are quite complex and diverse, and mechanisms through which IL-6 influences neuronal functions are primarily unknown. In the present study, we explored protective effect of IL-6 that was chronically applied to cerebellar granule neurons (CGNs) in culture against neurodamage induced by glutamate and mechanisms involved in the neuroprotective effect of IL-6. The chronic IL-6 exposure significantly prevented the CGNs from the glutamate-induced attenuation of neuronal vitality. This neuroprotective effect of IL-6 depended on its concentrations. IL-6 at 2.5 ng/ml did not markedly improve the neuronal vitality, but IL-6 at 5 and 10 ng/ml notably improved the neuronal vitality. The glutamate evoked neuronal apoptosis also was strikingly inhibited by the chronic IL-6 pretreatment. Intracellular Ca2+ in the CGNs lacking IL-6 pretreatment acutely rose as soon as these neurons were stimulated by glutamate and were maintained at higher levels during the whole 18-min period of glutamate attack. Although intracellular Ca2+ in the IL-6-pretreated CGNs also produced an acute and transient elevation in response to the glutamate insult, they quickly dropped and recovered to basal levels before the glutamate application. Anti-gp130 monoclonal antibody (mAb) blocked the suppressive effect of IL-6 on the glutamate-induced intracellular Ca2+ overload. These results reveal that IL-6 can protect neurons against glutamate-induced neurotoxicity, and suggest that the neuroprotective effect of IL-6 may be via gp130 signal transducing pathway to suppress the glutamate-evoked intracellular Ca2+ overload. PMID- 15663962 TI - Acute and long-term alteration of chemokine mRNA expression after anti-viral and anti-inflammatory treatment in herpes simplex virus encephalitis. AB - Mortality and morbidity rates remain high among patients with herpes simplex virus encephalitis (HSVE). Chemokine-mediated recruitment and activation of leukocytes to focal areas of viral CNS infection are crucial steps in antiviral response and clearance. However, the inflammatory reaction and cellular antiviral response may enhance collateral damage to neurons and account for chronic progressive brain damage. We identified a specific mRNA expression of the interferon-gamma-inducible chemokines (CXCL9, CXCL10 and CXCL11), and RANTES (CCL5) in the acute course and long-term of experimental HSVE. This pattern was substantially altered by anti-viral and anti-inflammatory treatment. Our findings indicate a pivotal role of these chemokines in the immunopathogenesis of HSVE. PMID- 15663963 TI - Identification of the epitope of a monoclonal antibody to DJ-1. AB - Mutations in DJ-1 can cause early onset parkinsonism. Various antibodies have been generated to detect this protein, one of which is a commonly used monoclonal antibody (clone 3E8). Since results of in situ examinations of DJ-1 expression with this antibody have differed from analyses with species-specific antibodies (e.g. rat), it would be useful to know the epitope for this antibody. Using GFP tagged deletion constructs of human DJ-1, we have localized the epitope region for this antibody to within residues 56-78 of human DJ-1. Mapping this region to the published three-dimensional structure of DJ-1 indicates that this is a solvent-accessible surface epitope. Immunonegativity of E64D mutant DJ-1 with the monoclonal antibody suggests that glutamate 64 of human DJ-1 contributes to the epitope recognized by this antibody. Moreover, the loss of immunoreactivity due to such a small substitution demonstrates the remarkable sensitivity of the monoclonal antibody 3E8 to DJ-1. PMID- 15663964 TI - Divergent distribution of cytoglobin and neuroglobin in the murine eye. AB - Neuroglobin (Ngb) and cytoglobin (Cygb) are two vertebrate globins with yet poorly defined functions. Previous studies had demonstrated a high expression level of neuroglobin in the mammalian retina, being in line with a respiratory function. Here we show that in the mouse eye, cytoglobin is localised in fibroblasts of the ciliary processes and the choroidea. In the neuronal retina, cytoglobin is expressed in a subset of neurons of the ganglion cell and inner nuclear layers. Cytoglobin is also present in the inner plexiform layer, but absent from the pigment cells. Neuroglobin is localised in photoreceptor inner segments, the plexiform layers and the ganglion cell layer. The divergent distribution of neuroglobin and cytoglobin in the mammalian retina suggests distinct functions of these proteins in the vertebrate nervous system. While neuroglobin seems to be associated with oxygen consumption, a respiratory function of cytoglobin is unlikely. PMID- 15663965 TI - Electrophysiological evidence of enhanced distractibility in ADHD children. AB - Abnormal involuntary attention leading to enhanced distractibility may account for different behavioral and cognitive problems in children with attention deficit hyperactivity disorder (ADHD). This was investigated in the present experiment by recording event-related brain potentials (ERPs) to distracting novel sounds during performance of a visual discrimination task. The overall performance in the visual task was less accurate in the ADHD children than in the control children, and the ADHD children had a higher number of omitted responses following novel sounds. In both groups, the distracting novel sounds elicited a biphasic P3a ERP component and a subsequent frontal Late Negativity (LN). The early phase of P3a (180-240 ms) had significantly smaller amplitudes over the fronto-central left-hemisphere recording sites in the ADHD children than in the control group presumably due to an overlapping enhanced left-hemisphere dominant negative ERP component elicited in the ADHD group. Moreover, the late phase of P3a (300-350 ms) was significantly larger over the left parietal scalp areas in the ADHD children than in the controls. The LN had a smaller amplitude and shorter latency over the frontal scalp in the ADHD group than in the controls. In conclusion, the ERP and behavioral effects caused by the novel sounds reveal deficient control of involuntary attention in ADHD children that may underlie their abnormal distractibility. PMID- 15663966 TI - Influence of a tumor necrosis factor gene polymorphism in Japanese patients with multiple system atrophy. AB - We studied promoter region polymorphisms in the tumor necrosis factor (TNF), interleukin (IL)-6, IL-10, and transforming growth factor (TGF)-beta1 genes in Japanese patients with multiple system atrophy (MSA) (n=122) and normal controls (n=277). The frequency of the TNF-1031C, a high producer allele of TNF, was increased significantly in MSA patients compared with controls (chi2=12.36, P=0.0021, Pc=0.0084). In contrast, there was no difference in the genotype or allele frequency in the other cytokine gene polymorphisms. We also failed to detect any difference in the disease onset between each genotype of the polymorphisms examined. Our findings suggest that TNF might have a toxic effect in MSA. PMID- 15663970 TI - History of the development of international emergency medicine. AB - A future challenge for international EM development is expanding the specialty to Africa, where it is non-existent. Another goal for international EM development is to assist in the peace process in the Middle East. There is increasing need for good EM development throughout the world. Development of international EM has been rapid, with most of the spread of development occurring in the last 10 years. There is great opportunity and reward for students, residents, and EM physicians to further promote and develop EM internationally. PMID- 15663967 TI - Effect of acidic oligosaccharide sugar chain on scopolamine-induced memory impairment in rats and its related mechanisms. AB - In this study we evaluated the effect of a novel, marine-derived, acidic oligosaccharide on scopolamine-induced amnesia in rats using the Morris water maze test. The results show that 30-day administration of this oligosaccharide, referred to as acidic oligosaccharide sugar chain (AOSC), to rats attenuates memory impairment by scopolamine, as evaluated by shortened escape latency, swimming distance, and increased swimming time of rats with memory impairment induced by scopolamine in the quadrant where the platform is placed. The data additionally suggest that an appropriate dose of scopolamine, a traditional muscarinic receptor antagonist, elevates oxidative damage in brain, characterized by inactivation of superoxide dismutase (SOD) and glutathione peroxidase (GSH Px), and consequently, inhibition of ATPase in the hippocampus and cerebral cortex. AOSC ameliorates oxidative injuries caused by scopolamine by increasing the activities of SOD, GSH-Px, and ATPase. Further investigation by flow cytometry revealed that AOSC significantly reduces the overloading of intracellular free calcium ion ([Ca2+]i), thus suppressing apoptosis induced by H2O2 in human neuroblastoma SH-SY5Y cells. These findings suggest that AOSC can induce cognitive improvement via its antioxidant activity. PMID- 15663971 TI - Priorities in global emergency medicine development. AB - EM initiatives are gaining global acceptance as a result of emergency physicians; local advocates; national, transnational, and international EM organizations; and governmental leadership, organizations, and agencies involved in international health and an evolving global health agenda. Spanning the spectrum from basic initiatives to improve acute care services to mature EM specialty development, all countries acknowledge the need for emergency care. The level of EM development in a country is fluid and depends on many variables, including status of health development, burden of disease,resources, advocacy, available expertise, and public demand. Emergency physicians should support the promotion of EM in the context of essential public health and primary care initiatives in these developing countries. Additionally, emergency physicians should work closely with stakeholders, health policy experts, health economics, and international organizations involved in health care to promote the advancement of EM worldwide. PMID- 15663972 TI - Regional highlights in global emergency medicine development. AB - There are a number of notable trends in the development of EM that have been highlighted at the regional level globally. From the establishment of formal EM training as the standard of care to practice in an ED to the influence of transnational regulatory bodies and financing mechanisms on specialty development and the broad-based issues of health security that affect EM, the specialty continues to grow around the world. As practitioners in each of these regions struggle to respond effectively to the development issues and challenges presented here, they continue to advance EM as one of the more dynamic young specialties in medicine. The regions presented here are as distinctive as the people who practice our specialty; however, in many respects the highlights and challenges are universal. This recognition is perhaps what motivates the appeal for collaboration in international EM development. PMID- 15663973 TI - Funding emergency medicine development in low- and middle-income countries. AB - The specialty of EM is developing rapidly throughout the world. This growth is relatively lacking in the LICs, however. The lack of resources and financing capabilities in these regions may hinder specialty development. Further growth of the specialty in these countries requires an understanding of their health priorities and the global health and development agencies that often assist these countries in supporting the health sector. Identifying health priorities in these regions that intersect with EM is crucial and may form the basis for further expansion of EM. Many potential funding opportunities exist within the governmental and private sector, but all require some familiarity with application mechanisms and project cycles. Building relationships with personnel within these agencies and countries of interest is often fundamental to successful programmatic funding. PMID- 15663974 TI - Hot topics in international emergency medicine. AB - The Global Burden of Disease report published in 1996 predicted that by 2020 the majority of illness burden globally will be from in-juries, mental illness, and chronic diseases. Patients with many of the disorders leading the list frequently require emergent care. This article provides a brief literature-based introduction to these and other important topics relevant to international development of emergency medicine. PMID- 15663975 TI - Injury prevention and other international public health initiatives. AB - Injuries, whether caused by unintentional or intentional events, area significant public health problem. The burden of injury is greatest in low-and middle-income countries and among individuals of low socioeconomic status living in high-income countries. Most of these injuries are prevent-able. Emergency physicians can play an important role in reducing the global burden of injuries by providing expert care and by identifying, implementing, and evaluating population-based countermeasures to prevent and control injuries. The strategy used in a particular country depends in large part on the nature of the local problem, the concerns of the population, the availability of resources, and competing demands. Even simple countermeasures may have a big impact in reducing the global burden of death and disability due to injury. PMID- 15663976 TI - Understanding diversity among prehospital care delivery systems around the world. AB - Understanding the diversity in EMS systems around the world isa daunting challenge. It is difficult to describe or categorize these complex systems, but examination of whether a country has an informal or formal system is a starting point. Further understanding of access within the system allows additional insight into the care available. Personnel, training,equipment, communication systems, and service provider levels offer clear information regarding the sophistication of the system. The focus of an EMS system can help gauge its development and role in the community. Clear descriptions of each of these dimensions allows for better communication and understanding of a country's prehospital care system. This dimensional analysis enhances communication regarding the existing system and provides an opportunity to understand where improvements and interventions might be effective. PMID- 15663977 TI - International humanitarian assistance: where do emergency physicians belong? AB - As human civilization faces new and challenging humanitarian crises, the entrance of EPs into the realm of HA is needed and timely. As noted by Jennifer Leaning [35], an EP at Harvard University, "Medical and public health personnel who respond to these crises enter a domain of perilous complexity. Road maps do not exist, but the possibility of good favors the prepared mind." Thus, with proper training in the principles of public health and experience, EPs have a tremendous opportunity to affect the realm of international HA, particularly via the analysis and development of international emergency health systems and building the capacity for effective relief. EPs with training in HA can be valuable assets to relief programs in the field and administratively, especially after the initial disaster and during the transition to health system reconstruction and development. PMID- 15663978 TI - Lessons learned from international emergency medicine development. AB - The lessons learned from development of EM around the world span several key areas including general development, systems comparisons, models of EM practice, and education and training. Neither definitive nor exhaustive, these lessons learned are intended to be viewed as sign posts along the road traveled at this point in international EM development. It is hoped that future participants in international EM development can assimilate these lessons learned, adopt the most relevant ones, and add their own insight and wisdom to this growing list. Most importantly, it is hoped that by whichever path future development takes, we all reach the same destination of providing the best possible emergency medical care for the people of the world. PMID- 15663979 TI - Preparing for international medical service. AB - In many ways, preparation for medical service outside the United States is not greatly different from preparation for a self-guided vacation tour of another country. The major differences are in the rigor that one should apply to being sure all details are set, particularly if one is planning to go to a site away from a capital city. Additionally, the mental preparation is required in terms of the understanding of global health issues in a holistic sense and the personal mental preparation for working under what may be austere conditions. With solid preparation, anyone interested in IH experiences should be able to have a rewarding, trouble-free experience. PMID- 15663980 TI - Developing and implementing emergency medicine programs globally. AB - International EM development requires a comprehensive assessment of the current system and careful planning to ensure that the most important needs are addressed. Modeling a country's EM system after an existing foreign system underestimates the complex needs for instituting appropriate system interventions. Planning must include all stakeholders. With appropriate planning, international interventions can contribute to health system advancement. PMID- 15663982 TI - Using distance-based technologies for emergency medicine training and education. AB - Cost-effective and sustainable ways of continuing to improve emergency medical services and education worldwide must be pursued if the field is to continue to expand globally. Distance-based learning and the use of telecommunications advances present us with an ideal opportunity to improve international medical education. Such technologies can overcome the financial and logistic constraints of travel and can complement existing exchanges. PMID- 15663981 TI - International emergency medicine fellowships. AB - The active interchange of intellectual ideas in the quest to improve healthcare globally will likely be best served by active interchange among physicians around the world. Subspecialty fellowship training programs for United States and foreign graduates will provide a focused path to development of a global network of physicians dedicated to the delivery of high-quality emergency health services. PMID- 15663983 TI - Research agendas in global emergency medicine. AB - As the global culture moves forward into the 21st century with increasing interaction of populations through direct contact and electronic interchange, those citizens of our planet who have not benefited from the material gains that have been realized in the "mature economy" countries will increasingly seek equity on all levels, beginning with the most fundamental aspect of health care. There is a need to develop a capacity for treatment of urgent and emergent health conditions globally, a need that will only increase with advancing global economic development. PMID- 15663984 TI - Reflections on becoming a specialty and its impact on global emergency medical care: our challenge for the future. AB - International emergency medicine development includes many activities. Among them are efforts to establish and support the development of the specialty of emergency medicine. In carrying out such activities, it is important for emergency physicians to be aware of the story of the establishment and development of the specialty of emergency medicine in the United States and to seek ways to support similar efforts in other countries. The benefits of specialty development toward improving emergency care globally are likely to be immense. PMID- 15663985 TI - Steroids and prostate cancer. PMID- 15663986 TI - Hormonal, cellular, and molecular regulation of normal and neoplastic prostatic development. AB - This review on normal and neoplastic growth of the prostate emphasizes the importance of epithelial-mesenchymal/stromal interactions. Accordingly, during prostatic development urogenital sinus mesenchyme (a) specifies prostatic epithelial identity, (b) induces epithelial bud formation, (c) elicits prostatic bud growth and regulates ductal branching, (d) promotes differentiation of a secretory epithelium, and (e) specifies the types of secretory proteins expressed. In reciprocal fashion, prostatic epithelium induces smooth muscle differentiation in the mesenchyme. Epithelial-mesenchymal interactions during development continue postnatally into adulthood as stromal-epithelial interactions which play a homeostatic role and in so doing reciprocally maintain epithelial and stromal differentiation and growth-quiescence. Prostatic carcinogenesis involves perturbation of these reciprocal homeostatic cell-cell interactions. The central role of mesenchyme in prostatic epithelial development has been firmly established through analysis of tissue recombinants composed of androgen-receptor-positive wild-type mesenchyme and androgen-receptor-negative epithelium. These studies revealed that at the very least ductal morphogenesis, epithelial cytodifferentiation, epithelial apoptosis and epithelial proliferation are regulated by stromal and not epithelial androgen receptors. Likewise, progression from non-tumorigenesis to tumorigenesis elicited by testosterone plus estradiol proceeds via paracrine mechanisms. Thus, stromal-epithelial interactions play critical roles in the hormonal, cellular, and molecular regulation of normal and neoplastic prostatic development. PMID- 15663987 TI - The epidemiology of sex steroid hormones and their signaling and metabolic pathways in the etiology of prostate cancer. AB - The purpose of this review is to discuss the epidemiologic literature on the association of sex steroid hormones and components of their signaling and metabolic pathways with prostate cancer and to describe data evaluating racial variation in sex steroid hormone pathways as a possible explanation for the notably higher risk of prostate cancer in African-American men compared to white or Asian men. Although sex steroid hormones likely contribute to the growth and progression of prostate cancer, associations between hormones and prostate cancer risk across the range of normal levels have been difficult to reliably demonstrate epidemiologically. Methodologic issues no doubt have made the detection of these associations difficult. Of particular importance are (1) the inadequacy of measuring circulating hormones in middle age as a surrogate for the exposure in the target cells in the prostate at the relevant time in life and (2) the current inability to integrate across components of the sex steroid hormone signaling pathway to fully capture target cell androgenic and estrogenic stimulation. Although the approach of evaluating polymorphisms in genes involved in sex steroid hormone signaling or metabolism as a way to minimize some of the issues in the direct measurement of hormones is logical, the findings among these studies are somewhat difficult to reconcile as well. The problems of the changing case mix due to screening for elevated PSA, small sample sizes increasing the likelihood of false negative and false positive results, the controls and their allele frequencies not being representative of the population at risk, and lack of knowledge of the functional consequence of a polymorphism in relation to other polymorphisms in that gene or without consideration of other genes involved in the same pathway may be contributory. The primary result of the Prostate Cancer Prevention Trial confirms that intraprostatic dihydrotestosterone levels in the normal range indeed do contribute to the growth of prostate adenocarcinoma. However, the secondary result of higher-grade disease in cases in the finasteride arm coupled with clinical studies showing higher grade disease in non-metastatic cases with lower serum androgens, if not a pathological artifact or detection bias in the finasteride arm, possibly suggests a complex relationship between androgens and the growth versus differentiation of a prostate tumor. Finally, racial variation in components of the sex steroid hormone pathway do appear to exist, but whether the extent of the variation is adequately great such that it accounts for some of the substantial differences in prostate cancer incidence among blacks, whites, and Asians is unclear. PMID- 15663988 TI - Alterations of androgen receptor in prostate cancer. AB - The significance of androgens in the development of prostate cancer has been known for more than half century. During the last decade, a lot of effort has been put to study the significance of the specific nuclear receptor of the hormone, androgen receptor (AR). It has been suggested that polymorphisms, especially the length of CAG repeat in exon 1 of the gene, are associated with the risk of prostate cancer. However, not all studies have confirmed the association. Most surprisingly, it has now become clear that prostate carcinomas emerging during the androgen withdrawal therapy (i.e. hormone-refractory tumors) are capable of reactivating the AR-mediated signalling despite of the low levels of androgens. In addition, it has been shown that AR gene itself is genetically targeted. One-third of the hormone-refractory prostate carcinomas contains amplification of the gene. In addition, 10-30% of prostate carcinomas treated by antiandrogens acquire point mutation in the AR gene. The genetic alterations in AR indicate that receptor should be considered as putative treatment target. Evidently, the currently available antiandrogens are not capable to abolish the AR-mediated signalling efficiently enough. PMID- 15663989 TI - Expression and function of androgen receptor coactivators in prostate cancer. AB - Human androgen receptor (AR) associates with coactivator or corepressor proteins that modulate its activation in the presence of ligand. Early studies on AR coactivators in carcinoma of the prostate were hampered because of lack of respective antibodies. Investigations at mRNA level revealed that most benign and malignant prostate cells express common coactivators. AR coactivators SRC-1 and TIF-2 are up-regulated in tissue specimens obtained from patients who failed prostate cancer endocrine therapy. Increased expression of these coactivators is associated with enhanced activation of the AR by the adrenal androgen dehydroepiandrosterone. Similar association between AR coactivator expression and high prostate cancer grade and stage was reported for RAC-3 (SRC-3). The transcriptional integrator CBP was detected in clinical specimens representing organ-confined prostate cancer, lymph node metastases and tumour cell lines. Agonistic effect of the nonsteroidal antiandrogen hydroxyflutamide was strongly potentiated in prostate cells transfected with CBP cDNA. A functional homologue of CBP, p300, is implicated in ligand-independent AR activation by interleukin-6. The AR coactivator Tip60, which is up-regulated by androgen ablation, is recruited to the promoter of the prostate-specific antigen gene in the absence of androgen in androgen-independent prostate cancer sublines. It was proposed that the cofactor ARA70 is a specific enhancer of AR action. However, research from other laboratories has demonstrated interaction between ARA70 and other steroid receptors. Although in some cases dominant-negative coactivator mutants inhibited proliferation of prostate cancer cells in vitro, confirmation from in vivo tumour models is missing. In summary, several abnormalities in AR coactivator expression and function are associated with prostate cancer progression. PMID- 15663990 TI - Androgens, lipogenesis and prostate cancer. AB - Both experimental and epidemiological data indicate that androgens are among the main factors controlling the development, maintenance and progression of prostate cancer. Identifying the genes that are regulated by androgens represents a major step towards the elucidation of the mechanisms underlying the impact of androgens on prostate cancer cell biology and is an attractive approach to find novel targets for prostate cancer therapy. Among the genes that have been identified thus far, several genes encode lipogenic enzymes. Studies aimed at the elucidation of the mechanisms underlying androgen regulation of lipogenic genes revealed that androgens coordinately stimulate the expression of these genes through interference with the molecular mechanism controlling activation of sterol regulatory element-binding proteins (SREBPs), lipogenic transcription factors governing cellular lipid homeostasis. The resulting increase in lipogenesis serves the synthesis of key membrane components (phospholipids, cholesterol) and is a major hallmark of cancer cells. Pharmacologic inhibition of lipogenesis or RNA-interference-mediated down-regulation of key lipogenic genes induces apoptosis in cancer cell lines and reduces tumor growth in xenograft models. While increased lipogenesis is already found in the earliest stages of cancer development (PIN) and initially is androgen-responsive it persists or re emerges with the development of androgen-independent cancer, indicating that lipogenesis is a fundamental aspect of prostate cancer cell biology and is a potential target for chemoprevention and for antineoplastic therapy in advanced prostate cancer. PMID- 15663991 TI - Sex steroid hormone metabolism and prostate cancer. AB - The growth and function of the prostate is dependent on androgens. The two predominant androgens are testosterone, which is formed in the testis from androstenedione and 5alpha-dihydrotestosterone, which is formed from testosterone by 5alpha-reductases and is the most active androgen in the prostate. Prostate cancer is one of the most common cancers among men and androgens are involved in controlling the growth of androgen-sensitive malignant prostatic cells. The endocrine therapy used to treat prostate cancer aims to eliminate androgenic activity from the prostatic tissue. Most prostate cancers are initially responsive to androgen withdrawal but become later refractory to the therapy and begin to grow androgen-independently. Using LNCaP prostate cancer cell line we have developed a cell model to study the progression of prostate cancer. In the model androgen-sensitive LNCaP cells are transformed in culture conditions into more aggressive, androgen-independent cells. The model was used to study androgen and estrogen metabolism during the transformation process. Our results indicate that substantial changes in androgen and estrogen metabolism occur in the cells during the process. A remarkable decrease in the oxidative 17beta-hydroxysteroid dehydrogenase activity was seen whereas the reductive activity seemed to increase. The changes suggest that during transformation estrogen influence is increasing in the cells. This is supported by the cDNA microarray screening results which showed over-expression of several genes up-regulated by estrogens in the LNCaP cells line representing progressive prostate cancer. Since local steroid metabolism controls the bioavailability of active steroid hormones in the prostate, the variations in steroid-metabolizing enzymes during cancer progression may be crucial in the regulation of the growth and function of the organ. PMID- 15663992 TI - Endocrine treatment of prostate cancer. AB - Although androgen deprivation as a treatment for patients with prostate cancer was described more than 60 years ago its optimal use remains controversial. The widespread use of prostate-specific (PSA) assay has lead to earlier diagnosis and earlier detection of recurrent disease. This means that the systemic side effects of androgen deprivation and quality of life have become more important. Debates continue regarding the proper use and timing of endocrine therapy with orchiectomy, oestrogen agonists, gonadotropin hormone-releasing hormone (GnRH) agonists, GnRH antagonists, and androgen antagonists. A critical review of the literature was performed. Data support that androgen deprivation is an effective treatment for patients with advanced prostate cancer. However, although it improves survival, it is not curative, and creates a spectrum of unwanted effects that influence quality of life. Castration remains the frontline treatment for metastatic prostate cancer, where orchiectomy, oestrogen agonists and GnRH agonists produce equivalent clinical responses. Maximum androgen blockade (MAB) is not significantly more effective than single agent GnRH agonist or orchiectomy. Nonsteroidal antiandrogen monotherapy is as effective as castration in treatment of locally advanced prostate cancer offering quality of life benefits. Adjuvant endocrine treatment is able to delay disease progression at any stage. There is, however, controversy of the possible survival benefit of such treatment, including patients having PSA relapse after definitive local treatment for prostate cancer. Neoadjuvant endocrine treatment has its place mainly in the external beam radiotherapy setting. Intermittent androgen blockade is still considered experimental. The decision regarding the type of androgen deprivation should be made individually after informing the patient of all available treatment options, including watchful waiting, and on the basis of potential benefits and adverse effects. Several large studies are under way to investigate the role of adjuvant endocrine treatment in the field of early prostate cancer, intermittent androgen deprivation and endocrine therapy alone compared with endocrine therapy with radiotherapy. The real challenge, however, is to develop better means to avert hormone-refractory prostate cancer and better treatments for patients with hormone-refractory disease when it occurs. PMID- 15663993 TI - Role of estrogens in development of prostate cancer. AB - Estrogens have previously been extensively used in prostate cancer treatment. Serious side effects, primarily in cardiovascular system have, however, limited their use. The therapeutic effect of estrogen in preventing prostate cancer growth was mainly obtained indirectly by feedback inhibition of the hypothalamic release of LRH leading to lowered serum androgen levels and castration like effects. Prostate tissue is also most probably a target for direct regulation by estrogens. Prostate contains estrogen receptor alpha (ERalpha) and beta (ERbeta), which are localized characteristically in stroma and epithelium, respectively. The physiological function of these receptors is not known but there is evidence of the role of estrogens in prostatic carcinogenesis. Developing prostate seems particularly sensitive to increased level of endogenous and/or exogenous estrogens. Perinatal or neonatal exposure of rats and mice to estrogens leads to "imprinting" of prostate associated with increased proliferation, inflammation and dysplastic epithelial changes later in life. Prolonged treatment of adult rodents with estrogens along with androgens also leads to epithelial metaplasia, PIN-like lesions and even adenocarcinoma of prostate speaking for the role of estrogen in prostate cancer development. Recent results concerning antiestrogen inhibition of prostate cancer development beyond PIN-type lesions in transgenic mouse models further suggests a role for estrogens in prostate cancer progression. These results also suggest that direct inhibition of estrogen action at the level of prostate tissue may provide an important novel principle of development of prostate cancer therapies. PMID- 15663994 TI - Interaction of nuclear receptor ligands with the Vitamin D signaling pathway in prostate cancer. AB - A number of hormonal ligands and/or the nuclear receptors that mediate their actions have been targeted for prostate cancer therapy. Androgens, the ligands for the androgen receptor (AR), are critical for the growth of prostate cancer. Inhibition of androgen production has been the mainstay of treatment for advanced prostate cancer for decades. Other more recently tested targets include retinoid receptors (RAR and RXR), glucocorticoid receptors (GR), estrogen receptors (ER) and peroxisome proliferator-activated receptors (PPAR). Calcitriol, acting through the Vitamin D receptor (VDR), has many tumor suppressive activities in the prostate, including inhibition of proliferation, induction of apoptosis and/or differentiation, and reduction of cellular invasion. Because of these properties, calcitriol and its less hypercalcemic analogs are being evaluated as agents to prevent or treat prostate cancer. Androgens, retinoids, glucocorticoids, estrogens and agonists of PPAR directly or indirectly impact Vitamin D signaling pathways, and vice versa. In order to design the most effective strategies to use calcitriol to prevent or treat prostate cancer, the interactions of other nuclear receptors and their ligands with the Vitamin D signaling pathway need to be considered. PMID- 15663995 TI - The role of Vitamin D3 metabolism in prostate cancer. AB - Vitamin D deficiency increases risk of prostate cancer. According to our recent results, the key Vitamin D hormone involved in the regulation of cell proliferation in prostate is 25(OH) Vitamin D3. It is mainly acting directly through the Vitamin D receptor (VDR), but partially also through its 1alpha hydroxylation in the prostate. A deficiency of 25(OH) Vitamin D is common especially during the winter season in the Northern and Southern latitudes due to an insufficient sun exposure, but Vitamin D deficient diet may partially contribute to it. A lack of Vitamin D action may also be due to an altered metabolism or Vitamin D resistance. Vitamin D resistance might be brought up by several mechanisms: Firstly, an increased 24-hydroxylation may increase the inactivation of hormonal Vitamin D metabolites resulting in a Vitamin D resistance. This is obvious in the cancers in which an oncogenic amplification of 24-hydroxykase gene takes place, although an amplification of this gene in prostate cancer has not yet been described. During the aging, the activity of 24 hydroxylase increases, whereas 1alpha-hydroxylation decreases. Furthermore, it is possible that a high serum concentration of 25(OH)D3 could induce 24-hydroxylase expression in prostate. Secondly, Vitamin D receptor gene polymorphism or defects may result in a partial or complete Vitamin D resistance. Thirdly, an overexpression or hyperphosphorylation of retinoblastoma protein may result in an inefficient mitotic control by Vitamin D. Fourthly, endogenous steroids (reviewed by [D.M. Peehl, D. Feldman, Interaction of nuclear receptor ligands with the Vitamin D signaling pathway in prostate cancer, J. Steroid Biochem. Mol. Biol. (2004)]) and phytoestrogens may modulate the expression of Vitamin D metabolizing enzymes. In summary, the local metabolism of hormonal Vitamin D seems to play an important role in the development and progression of prostate cancer. PMID- 15663996 TI - Severe hypophosphatemic osteomalacia in hormone-refractory prostate cancer metastatic to the skeleton: natural history and pitfalls in management. AB - We report the case of a severe symptomatic hypophosphatemic osteomalacia in a 66 year-old patient with hormone-refractory prostate cancer metastatic to the skeleton. A follow-up of 2 years from diagnosis to development of hormone refractoriness and death allowed us to study the natural history of this uncommon disturbance of mineral homeostasis in this common malignancy. Relevant to the difficult management of the late stages of prostate cancer is the failure of hypophosphatemia to respond to conventional therapeutic approaches and the favorable outcome of antitumor therapy suggesting that this group of patients, although having a poor prognosis, could still benefit from aggressive second line therapy. In this malignancy in which metastases have a predilection for bone, failure to recognize osteomalacia can only result in significantly increasing the burden of skeletal complications. PMID- 15663997 TI - Infrared imaging of calcified tissue in bone biopsies from adults with osteomalacia. AB - Osteomalacia is a pathological bone condition in which there is deficient primary mineralization of the matrix, leading to an accumulation of osteoid tissue and reduced bone mechanical strength. The hypothesis that there are no qualitative or quantitative differences in osteomalacic bone mineral or matrix compared to disease-free bones was tested by examining unstained sections of polymethyl methacrylate (PMMA) embedded iliac crest biopsies using Fourier transform infrared imaging (FTIRI) at approximately 6-microm spatial resolution. Controls were seven female subjects, aged 36-57, without apparent bone disease. The experimental group consisted of 11 patients aged 22-72, diagnosed with osteomalacia. The spectroscopic parameters analyzed in each data set were previously established as sensitive to bone quality: phosphate/amide I band area ratio (mineral content), 1660/1690 cm(-1) peak ratio (collagen cross-links), and the 1030/1020 cm(-1) peak ratio (mineral crystallinity). The correspondence between spectroscopic mineral content (phosphate/amide I ratio) and ash weight was validated for apatite crystals of different composition and crystallite size. The FTIRI results from the biopsies expressed as color-coded images and pixel population means were compared with the nonparametric Mann-Whitney U test. There were no significant differences in the cortical parameters. Significant difference was found in the mineral content of the trabecular regions with a lower mean value in osteomalacia (P = 0.01) than in controls. Mineral crystallinity tended to be decreased in the trabecular bone (P = 0.09). This study supports the hypothesis that, in osteomalacia, the quality of the organic matrix and of mineral in the center of bone does not change, while less-than optimal mineralization occurs at the bone surface. This study provides the first spectroscopic evaluation of whole bone mineral and matrix properties in osteomalacia, demonstrating that there are few differences in collagen cross links between biopsies from patients with osteomalacia and from individuals without histological evidence of bone disease. PMID- 15663998 TI - Biochemical markers of bone formation reflect endosteal bone loss in elderly men- MINOS study. AB - In the skeleton of elderly men, two opposite activities occur: bone loss at the endosteal envelope, which increases bone fragility, and periosteal apposition, which improves bending strength of bone. Both may contribute to serum bone formation markers although they have an opposite effect on bone fragility. The aim of this study was to determine if circulating bone formation markers reflect periosteal bone formation and endosteal bone remodelling in 640 men aged 55-85 years belonging to the MINOS cohort. We measured biochemical markers of bone formation (osteocalcin, bone alkaline phosphatase, N-terminal extension propeptide of type I collagen) and bone resorption (urinary and serum beta isomerised C-terminal telopeptide of collagen type I, total and free deoxypyridinoline). Parameters of bone size (cross-sectional surface of third lumbar vertebral body measured by X-ray, projected areas of total hip, femoral neck, radius and ulna measured by dual-energy X-ray absorptiometry) increased with age (r = 0.20-0.32, P < 0.0001). In contrast, parameters related to bone loss (areal bone mineral density [aBMD], volumetric bone mineral density [vBMD] and cortical thickness) and determined mainly by bone resorption, decreased with ageing (r = -0.14 to -0.23, P < 0.005-0.0001). Men in the highest quartile of bone resorption markers had lower aBMD (3.8-10.2%, P < 0.05-0.0001), lower vBMD (3.9-13.0%, P < 0.05-0.0001), and lower cortical thickness (1.5-9.6%, P < 0.05 0.0001) than men in the lowest quartile. Markers of bone resorption were not significantly associated with estimates of bone size at any skeletal site. Markers of bone formation were not associated with estimates of periosteal formation after adjustment for covariates. In contrast, men in the highest quartile of the bone formation markers had significantly lower aBMD (4.0-11.7%, P < 0.05-0.0001), lower vBMD (4.2-16.3, P < 0.05-0.0001) and lower cortical thickness (4.0-7.4%, P < 0.05-0.0001) than men in the lowest quartile. In summary, serum levels of bone formation markers are negatively correlated with the estimates of endosteal bone loss. In contrast, they disclose no association with parameters reflecting periosteal apposition. Thus, in elderly men, bone formation markers reflect endosteal bone remodelling, probably because of the coupling between resorption and formation activities. In contrast, they do not reflect the periosteal bone formation, probably because the periosteal surface is smaller and has a slower remodelling rate than the endosteal surface. PMID- 15663999 TI - Intervention thresholds for osteoporosis in the UK. AB - The aim of this study was to determine the threshold of fracture probability at which interventions became cost-effective in women based on data from the UK. We modelled the effects of an intervention costing pound 350 per year given for 5 years that decreased the risk of all osteoporotic fractures by 35% followed by a waning of effect (offset time) for a further 5 years. Sensitivity analyses included a range of treatment duration (3-10 years), intervention costs (pound 300-400/year) and offset times (0-15 years). Data on costs and risks were from the UK. Costs included direct costs, but excluded indirect costs due to morbidity. A threshold for cost-effectiveness of pound 30,000/QALY gained was used. With the base case ( pound 350 per year; 35% efficacy) treatment in women was cost-effective with a 10-year hip fracture probability that ranged from 1.1% at the age of 50 years to 9.0% at the age of 85 years. Intervention thresholds were sensitive to the assumed costs and offset time. The exclusion of osteoporotic fractures other than hip fracture significantly increased the cost effectiveness ratio because of the substantial morbidity from such other fractures, particularly at younger ages. Cost-effective scenarios were found for women at the threshold for osteoporosis from the age of 60 years. Treatment of established osteoporosis was cost-effective irrespective of age. We conclude that the inclusion of all osteoporotic fractures has a marked effect on intervention thresholds, that these vary with age and that available treatments can be targeted cost-effectively to individuals from the UK at moderately increased fracture risk. PMID- 15664001 TI - The effect of intranasal salmon calcitonin therapy on bone mineral density in idiopathic male osteoporosis without vertebral fractures--an open label study. AB - The aim of this study was to examine the effect of intranasal salmon calcitonin therapy on bone mineral density (BMD) in idiopathic male osteoporosis without vertebral fractures. We conducted a randomized, open label, controlled trial in 71 male patients (mean age 59 +/- 6 years) suffering from idiopathic osteoporosis (femoral neck T-score < -2.5) without vertebral deformity. Patients in the control group (n = 31) received 400 IU Vitamin D + 1000 mg elemental calcium daily while the treatment group (n = 40) received 400 IU Vitamin D, 1000 mg elemental calcium plus 200 IU calcitonin nasal spray daily during alternate months. The study period was 18 months. Compared to controls, nasal calcitonin was associated with significant increases in bone mineral density at the lumbar spine (+3.5 +/- (-4.3%) vs. +0.83 +/- 6.4%, P = 0.04) and the femoral neck (+3.2 +/- 3.9% vs. +0.68 +/- 5.7%, P = 0.004). No significant difference was observed at the radius between the treatment groups (+1.4 +/- 8.8% vs. +1.4 +/- 10.9%, P = 0.98). Treatment was well tolerated with no premature discontinuations or significant side effects compared to the control group. We conclude that 200 IU salmon calcitonin nasal spray used daily, intermittently proved to be an effective and safe therapy in male idiopathic osteoporosis. PMID- 15664000 TI - Surface plasmon resonance (SPR) confirms that MEPE binds to PHEX via the MEPE ASARM motif: a model for impaired mineralization in X-linked rickets (HYP). AB - Matrix Extracellular Phospho-glycoprotEin (MEPE) and proteases are elevated and PHEX is defective in HYP. PHEX prevents proteolysis of MEPE and release of a protease-resistant MEPE-ASARM peptide, an inhibitor of mineralization (minhibin). Thus, in HYP, mutated PHEX may contribute to increased ASARM peptide release. Moreover, binding of MEPE by PHEX may regulate this process in normal subjects. The nature of the PHEX-MEPE nonproteolytic interaction(s) (direct or indirect) is/are unknown. Our aims were to determine (1) whether PHEX binds specifically to MEPE, (2) whether the binding involves the ASARM motif region, and (3) whether free ASARM peptide affects mineralization in vivo in mice. Protein interactions between MEPE and recombinant soluble PHEX (secPHEX) were measured using surface plasmon resonance (SPR). Briefly, secPHEX, MEPE, and control protein (IgG) were immobilized on a Biacore CM5 sensor chip, and SPR experiments were performed on a Biacore 3000 high-performance research system. Pure secPHEX was then injected at different concentrations, and interactions with immobilized proteins were measured. To determine MEPE sequences interacting with secPHEX, the inhibitory effects of MEPE-ASARM peptides (phosphorylated and nonphosphorylated), control peptides, and MEPE midregion RGD peptides on secPHEX binding to chip-immobilized MEPE were measured. ASARM peptide and etidronate-mediated mineralization inhibition in vivo and in vitro were determined by quenched calcein fluorescence in hind limbs and calvariae in mice and by histological Sanderson stain. A specific, dose-dependent and Zn-dependent protein interaction between secPHEX and immobilized MEPE occurs (EC50 of 553 nM). Synthetic MEPE PO4-ASARM peptide inhibits the PHEX-MEPE interaction (K(D(app)) = 15 uM and B(max/inhib) = 68%). In contrast, control and MEPE-RGD peptides had no effect. Subcutaneous administration of ASARM peptide resulted in marked quenching of fluorescence in calvariae and hind limbs relative to vehicle controls indicating impaired mineralization. Similar results were obtained with etidronate. Sanderson-stained calvariae also indicated a marked increase in unmineralized osteoid with ASARM peptide and etidronate groups. We conclude that PHEX and MEPE form a nonproteolytic protein interaction via the MEPE carboxy-terminal ASARM motif, and the ASARM peptide inhibits mineralization in vivo. The binding of MEPE and ASARM peptide by PHEX may explain why loss of functional osteoblast-expressed PHEX results in defective mineralization in HYP. PMID- 15664002 TI - Calcium phosphate drug delivery system: influence of local zoledronate release on bone implant osteointegration. AB - Despite total hip replacement (THR) gives generally satisfactory results, the quality of outcome in young patients is markedly decreased compared to the average THR outcome. For this population, pharmacological treatment with bisphosphonate would be beneficial to decrease the peri-implant osteolysis. However, as this population does not necessarily suffer from osteoporosis, a nonsystemic treatment would be preferable. Zoledronate was then grafted to hydroxyapatite (HA) coating of titanium implants. The implants were inserted in rat condyles with various zoledronate concentrations. A positive concentration dependent effect was observed on the peri-implant bone density and on different histomorphometric parameters. Importantly for the outcome of the implants, the mechanical fixation was increased by the local presence of zoledronate. The obtained results open the way of an easy transformation of currently existing HA coated implants by grafting bisphosphonate onto the coating in order to increase their service life in the patients. PMID- 15664003 TI - RETRACTED: Menatetrenone and vitamin D2 with calcium supplements prevent nonvertebral fracture in elderly women with Alzheimer's disease. AB - This article has been retracted: please see Elsevier Policy on Article Withdrawal (https://www.elsevier.com/about/our-business/policies/article-withdrawal).This article has been retracted at the request of the Corresponding Author, Yoshihiro Sato, and the co-authors have been informed.Dr. Sato wishes to retract this article on the grounds that it contains fabricated clinical trial data, which he was responsible for producing. In addition, Dr. Sato claims he listed all of the named co-authors without their consent. The co-authors were therefore unaware of the presence of fabricated data in this publication and their participation in the publication. This retraction was initiated by Dr. Sato, and the Editor-in Chief of Bone was informed by the author directly. PMID- 15664005 TI - Human reaming debris: a source of multipotent stem cells. AB - The biological characteristics of human reaming debris (HRD) generated in the course of surgical treatment of long bone diaphyseal fractures and nonunions are still a matter of dispute. Therefore, the objective of the present investigation has been to characterize the intrinsic properties of human reaming debris in vitro. Samples of reaming debris harvested from 12 patients with closed diaphyseal fractures were examined ultrastucturally and were cultured under standard conditions. After a lag phase of 4-7 days, cells started to grow out from small bone fragments and established a confluent monolayer within 20-22 days. The cells were characterized according to morphology, proliferation capacity, cell surface antigen profile, and differentiation repertoire. The results reveal that human reaming debris is a source of multipotent stem cells which are able to grow and proliferate in vitro. The cells differentiate along the osteogenic pathway after induction and can be directed toward a neuronal phenotype, as has been shown morphologically and by the expression of neuronal markers after DMSO induction. These findings have prompted interest in the use of reaming debris-derived stem cells in cell and bone replacement therapies. PMID- 15664004 TI - Bladder exstrophy: effects on bone age, bone mineral density, growth, and metabolism. AB - Bladder exstrophy patients with or without augmentation have not been investigated according to metabolic bone problems, bone ages and growth, and development in details yet. We studied alterations in growth, bone ages, biochemistry of bone, bone mineral densities (BMD) of the forearm, neck of femur and lumbar vertebrae, blood gases, glomerular filtration rates (GFR), and electrolytes of 15 bladder exstrophy patients with augmentation and in those who had no augmentation. In six patients, a sigmoid colon was used for bladder augmentation and one patient underwent a ureterosigmoidostomy. Growth charts of all children were analyzed for determination of the percentiles. The parameters were compared with normal children and a comparison between augmented and nonaugmented patients were made. Growth retardation and decreased bone age were detected in all of the children. Ten patients with bladder exstrophy were below the 10th percentile for height. The mean age/bone age ratio of the patients was 1.59. The mean lumbar and femoral Z scores of the patients were -1.00 and -0.49, respectively. Mean BMD for distal radius was 0.239 g/cm2. Seven patients had a marked BMD decrease, their femoral and/or lumbar Z scores were below -1. Four cases had a pH lower than 7.35. In five patients, a HCO3 level less than 19 mmol/l was detected, four of them had an augmentation. Chloride measurements were slightly increased in six patients and alkaline phosphatase levels in five cases. Reduced GFR values were detected in two patients. There were no significant difference in laboratory values, in percentile height, and weights, in BMDs of femur, vertebra, forearm nor were any differences noted in age/bone age ratios in patients with augmentation when compared with those who had no augmentation. We found varying alteration in bone mineral density and HCO3 levels in patients with bladder exstrophy. Patients with bladder exstrophy, with or without augmentation, may develop serious growth retardation. As much as 45% of them, regardless of presence of augmentation, have an osteopenia or osteoporosis. We found a considerable difference in percentiles of heights as well as bone ages in bladder exstrophy patients when compared with normal population. We recommend close follow up of children with bladder exstrophy for linear growth, development of osteopenia, and bone ages. PMID- 15664007 TI - Prostaglandin-dependent activation of ERK mediates cell proliferation induced by transforming growth factor beta in mouse osteoblastic cells. AB - Transforming growth factor beta (TGF(beta)) is a major coupling factor for bone turnover and is known to stimulate osteoblastic proliferation. Recent information indicates that, in addition to the Smad pathway, TGF(beta) also activates MAP kinases in osteoblastic cells. The role of these signaling cascades in cell proliferation induced by TGF(beta) as well as the cellular and molecular mechanisms of their activation by TGF(beta) has been investigated in this study. In MC3T3-E1 cells, TGF(beta) enhanced cell proliferation by about 2-fold and induced activation of the three MAP kinases, extracellular regulated kinase (ERK), p38, and c-Jun N-terminal kinase (JNK). Surprisingly, however, whereas activation of Smad2 was rapid and maximal after 15-min incubation, activation of MAP kinases was delayed with p38 stimulation detected after 1-h exposure and activation of ERK and JNK after 3 h, suggesting indirect activation of MAP kinases by TGF(beta). Among factors known to be released in response to TGF(beta) in osteoblastic cells and influence their growth, prostaglandins (PGs) were good candidates that were further investigated for mediating TGF(beta)-induced activation of MAP kinases and cell proliferation. Indomethacin, a selective inhibitor of PG synthesis, completely blunted cell proliferation induced by TGF(beta) and markedly reduced activation of MAP kinases without influencing Smad2 phosphorylation. EP4A, a specific PGE2 receptor antagonist, also blunted TGF(beta)-induced osteoblastic proliferation. In addition to these effects, PGE2 rapidly activated MAP kinases in MC3T3-E1 cells and increased cell proliferation by about 2-fold. The role of each MAP kinases in mediating TGF(beta)- and PGE2 induced cell proliferation was investigated using selective inhibitors. U0126, a specific inhibitor of the ERK pathway, completely blocked both TGF(beta)- and PGE2-induced cell proliferation whereas SB203580 and SP600125, which are selective inhibitors of, respectively, p38 and JNK pathways, had no effect. Finally, the effect of PGE2 on activation of ERK was mimicked by phorbol esters and not by forskolin, and was associated with activation of protein kinase C. This latter effect and the stimulation of ERK induced by PGE2 were completely blocked by a specific inhibitor of PKC. In conclusion, data presented in this study strongly suggest that the local release of PGE2 is involved in cell proliferation induced by TGF(beta) in osteoblastic cells. This effect is mediated by the ERK pathway activated by a PKC-dependent mechanism. PMID- 15664006 TI - Quantitation of osteoblast-like cell mineralization on tissue culture polystyrene and Ti-6Al-4V alloy disks by Tc-99m-MDP labeling and imaging in vitro. AB - Technetium-99m-methylene-diphosphonate (Tc-99m-MDP) labeling was used to quantify mineralization of cultures of MC3T3 osteoblast-like cells in vitro. MC3T3 cells were cultured on tissue culture polystyrene (TCPS) in mineralizing and non mineralizing media, and then labeled by Tc-99m-MDP. The gamma signal from labeled samples was imaged with a gamma camera and compared with the calcium content in the same samples determined by inductively coupled plasma. The high correlation (coefficient of determination, 0.88) between these two values validated the radiotracer uptake method as a quantitative analytical tool for certain mineralization studies in vitro. There was an association between mineralization and radionuclide uptake in the MC3T3 cultures on titanium alloy, but the attenuation of the gamma photons by the metal resulted in a less robust correlation. The results warrant implementation of this scintigraphic method for quantitative studies of osteoblast-mediated mineralization in vitro. PMID- 15664008 TI - Positive, site-specific associations between bone mineral status, fitness, and time spent at high-impact activities in 16- to 18-year-old boys. AB - The incidence of hip and forearm fracture in elderly men in the United Kingdom is a public health issue, but there is limited knowledge on lifestyle factors affecting male bone health. The aim of this cross-sectional study was to evaluate the relationships between whole body and regional bone mineral status and self reported participation time in no-, low-, moderate-, and high-impact activities and fitness measurements in 16- to 18-year-old boys. One hundred twenty-eight boys underwent absorptiometry (DXA) measurements (Hologic QDR 1000W) of bone mineral content (BMC), bone area (BA), and bone mineral density (BMD) at the whole body, hip, spine, and forearm. They also completed the EPIC (European Prospective Investigation of Cancer) physical activity questionnaire, which allowed categorization of activities according to impact and aerobic intensity. Fitness and strength were assessed in each subject using estimated VO2 max, grip strength, and back strength. Significant positive relationships were found between BMC, BA, and BMD and the fitness and strength measurements and participation time in high-impact sports at most skeletal sites. The relationships were further examined after adjustment of BMC for height, weight, and bone area, thereby minimizing the influence of body and bone size on these relationships. VO2 max was a significant positive determinant of size-adjusted BMC at the whole body, the ultradistal and one-third radius, and all the hip sites, except the trochanter. Size-adjusted BMC at the forearm sites and trochanter was significantly positively associated with grip strength. Size adjusted BMC at the whole body and all the hip sites was significantly positively associated with time spent at high-impact activities. Differences in size adjusted BMC across thirds of time spent at high-impact activities were also examined. Boys in the highest third of high-impact activity had significantly greater size-adjusted whole body BMC and total hip BMC compared to subjects in the lowest third [+3.4 (1.2)% for whole body and +8.5 (2.2)% for hip, both P = 0.001]. Boys in the highest third of high-impact activity spent most activity time jogging, playing tennis, football and rugby, basketball, and exercising with weights. Back strength and lean mass were significantly greater in subjects in the highest third compared to those in the middle (P = 0.02) and lowest third (P = 0.03). No significant differences were found between subjects in each third of high-impact activity for potential confounders including TV viewing, calcium intake, body fat, and height. These findings indicate that participation of male adolescents in a range of high-impact activities for 1 h or more a day is associated with greater bone size and mineral content, especially at the hip. PMID- 15664009 TI - Genetic variation in femur extrinsic strength in 29 different inbred strains of mice is dependent on variations in femur cross-sectional geometry and bone density. AB - The femurs from groups of mice from 29 different inbred strains were characterized to study the genetic variations in bone parameters. For these analyses, we used peripheral quantitative computed tomography to assess bone size and density in addition to three-point bend testing to assess bone mechanical properties. Highly significant differences between inbred strains were found for all size, density, and mechanical parameters measured (P < 0.0001). Correcting femoral cross-sectional geometry values or bone mechanical properties values for body weight or femur length reduced but did not eliminate the variations in bone geometry or bone mechanical properties. Mice of similar body size had as much as a 40% difference in the midshaft total area of the femur. Regression analysis suggested that 50.9% of the variation in maximum load among strains was related to variations in section modulus, i.e., cross-sectional geometry, 21.5% was related to variations in material bone density, and 27.7% to variations in quality. These components were further analyzed to show that 3.9-27.8% of the variation in maximum load was related to adaptation to mechanical stress. These findings indicate that there is a significant genetic variation in the femur cross-sectional area, density, and mechanical properties between inbred mouse strains. These studies identify inbred mouse strains suitable for future studies identifying genes regulating bone geometry and mechanical properties. PMID- 15664010 TI - Growth hormone can reverse glucocorticoid-induced low bone turnover on cortical but not on cancellous bone surfaces in adult Wistar rats. AB - We evaluated the effect of glucocorticoids (GC) and growth hormone (GH) on cortical and cancellous bone turnover in adult rats using random vertical sections giving valid measurements of bone surfaces and bone formation parameters. GH administration could reverse GC-induced osteopenia and low bone turnover of cortical bone. However, GH could not reverse the GC-induced low bone turnover of cancellous bone. METHODS: Seventy female Wistar rats, 7 months of age, were divided into five groups: (1) start control, (2) saline, (3) GC 9 mg/kg/day (Solu Medrol), (4) GH 5 mg/kg/day, and (5) GC 9 mg/kg/day + GH 5 mg/kg/day, and injected for 3 months. The vertebral body was examined using dynamic histomorphometry and biomechanical tests. Nonparametric methods were used. RESULTS: Glucocorticoid administration induced a low bone turnover state of both the cortical and cancellous bone of the vertebral body, without altering the absolute amount of bone or the biomechanical competence of the vertebral body. GH administration induced a small increase in longitudinal bone growth and ventral modeling drift. This growth increased the total amount of cortical, endocortical, and cancellous bone in the vertebra. The biomechanical competence of a 3.5-mm high cylinder of the central vertebral body was also increased due to an increase in the amount of cortical bone, whereas the total amount of cancellous bone in the cylinder was unaltered. The cancellous bone density (CBV) was, however, increased due to thicker trabeculae probably induced by an accelerated mineral appositional rate (MAR) induced by GH. GH also increased longitudinal and ventral modeling drifts in the GC-injected animals. GH increased the amount of cortical bone and also the amount of cancellous bone close to the epiphyseal growth plate, whereas the cancellous bone volume of the central vertebral cylinder was unaffected by GH administration in GC-injected animals. GH could also increase parameters of bone formation (bone mineralizing surface (MS) and MAR) on cortical bone surfaces in GC-injected animals, whereas parameters of bone formation [MS and bone formation rates (BFR)] on cancellous bone surfaces were even lower than those of animals injected with GC alone. CONCLUSION: GH can reverse GC-induced low bone turnover on cortical but not on cancellous bone surfaces. PMID- 15664011 TI - Intrinsic bone tissue properties in adult rat vertebrae: modulation by dietary protein. AB - Bone strength depends on bone mass, geometry, microarchitecture, and intrinsic tissue quality. Whether intrinsic bone tissue properties could be influenced by changes in dietary protein is not known. To address this issue, nanoindentation tests were performed on the lateral, anterior, and posterior site of L5 vertebral bodies in adult female rats fed a normal protein containing diet and in ovariectomized (OVX) rats receiving an isocaloric low protein diet with or without isocaloric essential amino acids supplements. The tissue properties varied significantly between the different sites (anterior, posterior, lateral), suggesting possible effects of heterogeneous stress distribution on the vertebrae in vivo. Isocaloric low protein intake associated with ovariectomy led to significant decreases of indentation modulus, hardness, and dissipated energy on the posterior vertex. Axial compression tests of adjacent vertebral bodies were correlated with the indentation results. Correlations between macroscopic mechanical data obtained by axial compression of vertebral body, and intrinsic tissue properties measured by nanoindentation test suggest that postelastic behavior strongly varied with material fragility detected on the tissue level. Macroscopic stiffness however may be dominated by bone geometry changes and less by variations of intrinsic bone tissue properties. Combining parameters of tissue properties and bone mineral density was highly predictive of vertebral body ultimate strength. Besides geometry and microarchitecture, intrinsic bone tissue properties are important determinants of the mechanical competence of rat vertebrae. Changes in intrinsic tissue properties could thus contribute to the increased bone fragility found in protein undernutrition. PMID- 15664012 TI - Bone defect repair in immobilization-induced osteopenia: a pQCT, biomechanical, and molecular biologic study in the mouse femur. AB - The present study was carried out to determine whether immobilization-induced (Im) osteopenic bone possesses the same reparative capacity as normal healthy bone. Furthermore, the effects of mechanical loading versus immobilization on bone defect healing were studied. Three-week cast-immobilization was used to induce local osteopenia in mice. A standardized metaphyseal bone defect of the distal femur was created unilaterally both in immobilization-induced (Im) osteopenic mice and in nonimmobilized (Mo) age-matched control animals. After creation of the bone defect, the animals in both groups were further divided into two groups: 3-week cast-immobilization (Im-Im and Mo-Im) groups, and unrestricted weight-bearing (Im-Mo and Mo-Mo) groups. The healing process was followed up to 3 weeks using RNA analysis, histomorphometry, biomechanical testing, and pQCT measurements. At 3 weeks of healing without immobilization, bone mineral density (BMD), as well as bone bending stiffness and strength were higher in normal (Mo Mo) than in osteopenic (Im-Mo) bone. Although the levels of mRNAs characteristic to chondrocytes (Sox9 and type II collagen), hypertrophic chondrocytes (Type X collagen), osteoblasts (type I collagen and osteocalcin), and osteoclasts (cathepsin K) during the bone defect healing exhibited similarities in their expression profiles, mechanical loading conditions also caused characteristic differences. Mechanical loading during healing (Mo-Mo group) induced stronger expression of cartilage- and bone-specific genes and resulted in higher BMD than that seen in the cast-immobilized group (Mo-Im). In biomechanical analysis, increased bending stiffness and strength were also observed in animals that were allowed weight-bearing during healing. Thus, our study shows that bone healing follows the same molecular pathway both in osteopenic and normal bones and presents evidence for reduced or delayed regeneration of noncritical size defects in immobilization-induced osteopenic bone. PMID- 15664013 TI - Differential effects of alendronate treatment on bone from growing osteogenesis imperfecta and wild-type mouse. AB - Bisphosphonates have been reported to decrease the number of fractures in children with osteogenesis imperfecta (OI). The current study sought to further explore bisphosphonate-associated bone changes in OI by investigating the effects of alendronate (ALN) treatment on bone mechanical and material properties in osteogenesis imperfecta (oim/oim) and wild-type (+/+) mice treated with 26-73 microg kg(-1) day(-1) of ALN for 8 weeks via subcutaneously implanted pumps. Femoral three-point bend tests to evaluate cortical bone were combined with geometric and material density analysis. Cortical and trabecular architecture of metaphyseal bone were histomorphometrically evaluated and material density assessed by quantitative backscattered electron imaging (qBEI). For the cortical oim/oim bone, which revealed principally inferior biomechanical properties compared to +/+ bone, ALN neither improved cortical strength or any other mechanical property, nor affected cortical width (Ct.Wi.) or material density. In contrast, for the +/+ mice, bone strength was enhanced (+22%, P < 0.05) though coupled with increased brittleness (+28%, P < 0.05). This mechanical improvement was associated with an increase in Ct.Wi. (+17.3%, P = 0.02) and a reduction in heterogeneity of cortical mineralization (Ca(Width), -4%, P = 0.04). In the metaphysis, ALN raised cancellous bone volume (BV/TV) significantly in oim/oim as well as in +/+ mice (+97%, P = 0.008 and +200%, P < 0.0001, respectively). This occurred without any change in either material density or trabecular thickness (Tb.Th.) in the oim/oim mice, while in the +/+ mice, material density increased slightly but significantly (+3%, P = 0.004), and Tb.Th. increased by 77% (P < 0.0001). Taken together, these results illustrate the differential effects of ALN on oim/oim vs. +/+ bone, as well as on specific skeletal sites, i.e., cortical vs. trabecular bone. ALN augmented the mechanical, geometrical, and material properties of +/+ cortical and trabecular bone, while the only observable improvement to the oim/oim bone was increased cancellous bone volume. This suggests that in this mouse model of OI, the previously demonstrated bisphosphonate-associated reduction in fractures is primarily attributable to increased metaphyseal bone mass. PMID- 15664015 TI - Effect of simulated weightlessness on osteoprogenitor cell number and proliferation in young and adult rats. AB - Experiments with rats flown in space or hind limb unloaded (HU) indicate that bone loss in both conditions is associated with a decrease in bone volume and osteoblast surface in cancellous and cortical bone. We hypothesize that the decrease in osteoblastic bone formation and osteoblast surface is related to a decrease in the number of osteoprogenitors and/or decreased proliferation of their progeny. We tested this hypothesis by evaluating the effect of 14 days of HU on the number of osteoprogenitors (osteoblast colony forming units; CFU-O), fibroblastic colony forming units (CFU-F), and alkaline phosphatase-positive CFU (CFU-AP) in cell populations derived from the proximal femur (unloaded) and the proximal humerus (normally loaded) in 6-week-old and 6-month-old rats. To confirm the effect of unloading on bone volume and structure, static histomorphometric parameters were measured in the proximal tibial metaphysis. Effects of HU on proliferation of osteoprogenitors were evaluated by measuring the size of CFU-O. HU did not affect the total number of progenitors (CFU-F) in young or adult rats in any of the cell populations. In femoral populations of young rats, HU decreased CFU-O by 71.0% and mean colony size was reduced by 20%. HU decreased CFU-AP by 31.3%. As expected, no changes in CFU-O or CFU-AP were seen in cell populations from the humerus. In femoral cell populations of adult rats, HU decreased CFU-O and CFU-AP by 16.6% and 36.6%, respectively. Again, no effects were seen in cell populations from the humerus. In 6-week-old rats, there was a greater decrease in bone volume, osteoblast number, and osteoblast surface in the proximal tibial metaphysis than that observed in adult rats. Both trabecular thickness and trabecular number were decreased in young rats but remained unaffected in adults. Neither osteoclast number nor surface was affected by unloading. Our results show that the HU-induced decrease in the number of osteoprogenitors observed in vitro parallels the effects of HU on bone volume and osteoblast number in young and old rats in vivo, suggesting that the two may be interdependent. HU also reduced CFU-O colony size in femoral populations indicating a diminished proliferative capacity of osteoblastic colonies. PMID- 15664014 TI - Impaired bone resorption in cathepsin K-deficient mice is partially compensated for by enhanced osteoclastogenesis and increased expression of other proteases via an increased RANKL/OPG ratio. AB - Previous reports indicate that mice deficient for cathepsin K (Ctsk), a key protease in osteoclastic bone resorption, develop osteopetrosis due to their inability to properly degrade organic bone matrix. Some features of the phenotype of Ctsk knockout mice, however, suggest the presence of mechanisms by which Ctsk deficient mice compensate for the lack of cathepsin K. To study these mechanisms in detail, we generated Ctsk-deficient (Ctsk-/-) mice and analyzed them at the age of 2, 7, and 12 months using peripheral quantitative computed tomography, histomorphometry, resorption marker measurements, osteoclast and osteoblast differentiation cultures, and gene expression analyses. The present study verified the previously published osteopetrotic features of Ctsk-deficient mice. However, these changes did not exacerbate during aging indicating the absence of Ctsk to have its most severe effects during the rapid growth period. Resorption markers ICTP and CTX were decreased in the media of Ctsk-/- osteoclasts cultured on bone slices indicating impaired bone resorption. Ctsk-/- mice exhibited several mechanisms attempting to compensate for Ctsk deficiency. The number of osteoclasts in trabecular bone was significantly increased in Ctsk-/- mice compared to controls, as was the number of osteoclast precursors in bone marrow. The mRNA levels for receptor activator of nuclear factor (kappa)B ligand (RANKL) in Ctsk-/- bones were increased resulting in increased RANKL/OPG ratio favoring osteoclastogenesis. In addition, expression of mRNAs of osteoclastic enzymes (MMP 9, TRACP) and for osteoblastic proteases (MMP-13, MMP-14) were increased in Ctsk /- mice compared to controls. Impaired osteoclastic bone resorption in Ctsk-/- mice results in activation of osteoblastic cells to produce increased amounts of other proteolytic enzymes and RANKL in vivo. We suggest that increased RANKL expression mediates enhanced osteoclastogenesis and increased protease expression by osteoclasts. These observations underline the important role of osteoblastic cells in regulation of osteoclast activity and bone turnover. PMID- 15664016 TI - Projection from the ventral bed nucleus of the stria terminalis to the retrorubral field in rats and the effects of cells in these areas on mating in male rats versus gerbils. AB - This research identified the rat counterpart of the lateral cell group of the sexually dimorphic area (SDA) found in medial preoptic area (MPOA) gerbil of gerbils. The lateral SDA (lSDA) is critical for mating in male gerbils and contains most of the SDA cells projecting to the retrorubral field (RRF), a projection that is also important for mating. Therefore, to locate the counterpart of the lateral SDA, we traced the inputs to the rat RRF, which were dense in the ventral part of the bed nucleus of the stria terminalis (BST). To determine if the ventral BST or its projection to the RRF affects mating in male rats, we disrupted them bilaterally by placing cell-body lesions bilaterally in the ventral BST or unilaterally there and in the contralateral RRF. We also studied the effects of RRF lesions in both rats and gerbils. Bilateral ventral BST lesions, which left the medial preoptic nucleus intact, produced persistent and severe mating deficits. Disconnecting the ventral BST from the RRF also had long-lasting, but less severe, consequences. RRF lesions produced only temporary mating deficits in rats, but virtually eliminated mating in gerbils. The recovery of mating in rats after RRF, but not ventral BST, lesions, and the intermediate effects of disconnecting these areas from each other suggest that the ventral BST may contain mating-related projection neurons other than those projecting to the RRF or that its RRF-projecting cells send collaterals to another site. In either case, the pedunculopontine tegmental nucleus or raphe nuclei may be involved. PMID- 15664017 TI - Age-specific threats induce CRF expression in the paraventricular nucleus of the hypothalamus and hippocampus of young rats. AB - Young animals respond to threatening stimuli in an age-specific way. Their endocrine and behavioral responses reflect the potential threat of the situation at a given age. The aim of the present study was to determine whether corticotropin-releasing factor (CRF) is involved in the endocrine and behavioral responses to threat and their developmental changes in young rats. Preweaning 14 day-old and postweaning 26-day-old rats were exposed to two age-specific threats, cat odor and an adult male rat. The acute behavioral response was determined during exposure. After exposure, the time courses of the corticosterone response and of CRF expression in the paraventricular nucleus of the hypothalamus (PVN) and in extrahypothalamic areas were assessed. Preweaning rats became immobile when exposed to cat odor or the male rat, whereas postweaning rats became immobile to cat odor only. Male exposure increased serum corticosterone levels in 14-day-old rats, but cat odor failed to increase levels at either age. Exposure induced elevation of CRF mRNA levels in the PVN that paralleled changes in corticosterone levels. CRF may thus play a role in endocrine regulation and its developmental changes during early life. Neither cat odor nor the adult male altered CRF mRNA levels in the bed nucleus of the stria terminalis (BNST) or the amygdala, but both stimuli increased levels in the hippocampus. Hippocampal CRF mRNA expression levels did not parallel cat odor or male-induced immobility, indicating that CRF is not involved in this response in young rats but may be involved in aspects of learning and memory. PMID- 15664018 TI - Diminished maternal responsiveness during pregnancy in multiparous female common marmosets. AB - Common marmoset monkeys (Callithrix jacchus) live in small groups in which, usually, only a single female breeds and all group members provide infant care. When two females breed concurrently, however, they may commonly kill one another's infants, especially during the peripartum period. To investigate the mechanisms underlying infanticide by breeding females, we characterized responses of multiparous females to infants and determined circulating hormone levels in adult females during early pregnancy, late pregnancy, and the early postpartum period. Additionally, we compared the responses of postpartum females to their own infants and infants of other females (unfamiliar infants). Postpartum females were highly maternal toward both their own and unfamiliar infants, and showed no differences in their behavioral or hormonal responses to the two. During both early and late pregnancy, however, these females exhibited longer latencies to initially approach unfamiliar infants and spent less time carrying unfamiliar infants. Moreover, females spent less time carrying unfamiliar infants during late pregnancy than early pregnancy. Most late pregnant females never carried infants, and those that did rejected them quickly. Prolactin concentrations were higher and progesterone concentrations lower postpartum than in early or late pregnancy, while estradiol concentrations, the estradiol-to-progesterone ratio, and cortisol levels were higher during late pregnancy. Within reproductive conditions, however, maternal behaviors were not correlated with hormone levels. These results suggest that maternal responsiveness in marmosets may be attenuated during pregnancy, especially late pregnancy, and this may contribute to infanticide by breeding females. PMID- 15664019 TI - Menstrual cycle-related changes in plasma oxytocin are relevant to normal sexual function in healthy women. AB - Circulating levels of the neuro-hypophysial nonapeptide oxytocin increase during sexual arousal and orgasm in both men and women. A few studies have evaluated the effect of the menstrual cycle on plasma oxytocin in normally cycling, sexually active, healthy fertile women using or not using contraceptive pills. In 20 ovulating women and 10 women taking an oral contraceptive (group 1 and group 2, respectively), sexual function, hormonal profile, and plasma oxytocin (OT) were evaluated throughout the menstrual cycle. In group 1, plasma OT was significantly lower during the luteal phase in comparison with both the follicular and ovulatory phases. Plasma oxytocin was significantly correlated with the lubrication domain of the Female Sexual Function Index (FSFI) during the luteal phase and showed a trend towards statistical significance during the follicular phase. In group 2, plasma OT did not show any significant fluctuation throughout the menstrual cycle, even though a significant correlation was evident with both the arousal and the lubrication domain of the FSFI during the assumption of the contraceptive pill. These findings suggest that plasma OT fluctuates throughout the menstrual cycle in normally cycling healthy fertile women with adequate sexual activity but not taking any oral contraceptive pill. Moreover, plasma OT levels significantly relates to the genital lubrication in both women taking and not taking oral contraceptive pill apparently confirming its role in peripheral activation of sexual function. PMID- 15664020 TI - A phylogenetically controlled test of hypotheses for behavioral insensitivity to testosterone in birds. AB - In most male birds that exhibit paternal care, extending the spring testosterone (T) peak throughout the breeding season reduces nestling provisioning. However, in some species, this trade-off between high T and expression of paternal care is absent. For example, during some or all of the nestling period, T did not affect paternal behavior in Male Lapland longspurs (Calcarius lapponicus), chestnut collared longspurs (Calcarius ornatus), and great tits (Parus major). Two ecological constraints have been hypothesized to drive insensitivity to T after eggs hatch: (1) a short breeding season that limits breeding opportunities, and (2) a need for paternal care to ensure reproductive success. However, because two of the three species that exhibit T insensitivity are closely related, potential phylogenetic confounds limit determination of which, if either, factor constrains some males to T insensitivity. We examined the effects of supplementary T on paternal behavior in the Snow Bunting (Plectrophenax nivalis), a member of the monophyletic Calcarius/Plectrophenax clade. Male Snow Buntings are constrained to a short breeding season, but paternal care is not essential for survival of nestlings. We administered exogenous T during the parental phase to mimic the early spring T peak. T treatment increased song rates and interfered with paternal behavior such that nestlings of T-implanted males grew more slowly than controls. Our data suggest that T insensitivity in this clade is related to relatively recent constraints of the breeding environment (i.e., not simply common ancestry) and that the necessity of paternal care in some species may be a strong selective factor driving behavioral insensitivity to T during the parental phase. PMID- 15664021 TI - Estrogenic effects of zearalenone on the expression of progestin receptors and sexual behavior in female rats. AB - Zearalenone is a resorcylic acid lactone compound that is produced by fungal infection of edible grains and is believed to influence reproduction by binding to estrogen receptors. In order to study the potential estrogenic effects of this compound in the brain, we examined the effects of zearalenone on the expression of neuronal progestin receptors and feminine sexual behavior in female rats. Ovariectomized rats were treated with zearalenone (0.2, 1.0, or 2.0 mg), estradiol benzoate, or vehicle daily for 3 days. They were then either perfused, and progestin receptors visualized by immunocytochemistry, or injected with progesterone and tested for sexual receptivity with male rats. Progestin receptor containing cells were counted in the medial preoptic area and ventromedial hypothalamus. The two highest doses of zearalenone increased the concentration of neuronal progestin receptors, as did 10 microg of estradiol. The highest dose of zearalenone (2 mg) also induced progestin receptor staining density comparable to that of 10 microg of estradiol benzoate. In behavioral tests, ovariectomized animals treated with 2 mg of zearalenone followed by progesterone showed levels of sexual receptivity comparable to females treated daily with estradiol benzoate (2 microg) followed by progesterone. These studies suggest that, although structurally distinct and less potent than estradiol, zearalenone can act as an estrogen agonist in the rat brain. PMID- 15664022 TI - Enhanced yolk testosterone influences behavioral phenotype independent of sex in Japanese quail chicks Coturnix japonica. AB - Studies have demonstrated an effect of yolk testosterone levels on the physiology and behavior of nestling birds. In order to investigate this phenomenon experimentally in a precocial bird, we enhanced yolk testosterone, but within the physiological range, by injecting 50 ng testosterone in ethanol into Japanese quail Coturnix japonica eggs prior to incubation. The chicks hatching from these as well as from control eggs that had received the carrier-only or were left unmanipulated were subject to a number of behavioral tests from hatching to the age of 3 weeks. In addition, fecal samples were taken during a 90-min isolation period to determine a physiological response to a stressor. Experimental chicks performed a detour task faster and approached novel objects sooner than did the controls. Chicks from treated eggs took a longer time to start distress vocalizing and also produced less distress calls during open-field trials, took on average a larger number of trials for them to show tonic immobility and also excreted lower levels of corticosterone metabolites (BM) than did the controls. In response to a stressor, excreted BM was initially higher in the control chicks, as compared to the experimental birds. Induced behavioral effects were independent of sex with no sex treatment interactions found. In sum, experimentally enhanced testosterone levels in the eggs shifted individual behavioral phenotype towards "bold" or "proactive", irrespective of sex. We conclude that testosterone in the yolk influences the coping style of hatchlings and may be a potential means of maternal influence on offspring phenotype. PMID- 15664023 TI - Female sexual behavior, but not sex skin swelling, reliably indicates the timing of the fertile phase in wild long-tailed macaques (Macaca fascicularis). AB - The extent to which catharrine primate males are able to discern the fertile phase during the female ovarian cycle under natural conditions is still debated. In a recent study, we showed that wild male long-tailed macaques are able to detect the fertile phase, but the cues males used to assess female reproductive status remained unclear. In the present study, we tested female sex skin swelling and specific female behaviors for their reliability in signaling the fertile phase, as determined by measurement of fecal estrogens (E) and progestogens (P) during nine ovulatory cycles in seven free-ranging females. We found that changes in sex skin swellings showed a significant positive correlation to the E/P ratio, but swelling size did not significantly differ between cycle phases. In contrast, the frequency of two of the tested female behaviors, namely initiation of sexual interactions and reaching back for the male during copulation, was not only correlated with female reproductive hormones, but was significantly elevated during the fertile phase compared to nonfertile phases of the cycle. We thus conclude that female sex skin swelling does not reliably indicate the timing of the fertile phase in long-tailed macaques, whereas certain female behaviors do. Since cycles differed considerably in the number of males with which females had sexual interactions as well as in the number of sexual interactions with dominant males, the signaling character of these specific female behaviors appears to be robust against inconsistencies in these social variables. Female behavior might therefore play an important role in the recognition of the fertile phase by male macaques under natural conditions. PMID- 15664024 TI - Testosterone, cognition, and social status. AB - What leads some people to perform better than others on certain cognitive tasks? One explanation involves individual differences in testosterone. Testosterone is associated with higher performance on spatial tests, and lower performance on verbal tests. However, a large literature suggests that testosterone only predicts behavior when status is in jeopardy. In the present study, we manipulated status before administering a spatial and verbal test. In a high status position, high-testosterone individuals performed well on both tests, and blood pressure dropped. In a low-status position, high-testosterone individuals performed relatively poorly on both tests, and blood pressure did not change. These data suggest that differences in cognitive performance stem from an interaction between testosterone and the social situation. PMID- 15664025 TI - The effects of mating stimulation on c-Fos immunoreactivity in the female hamster medial amygdala are region and context dependent. AB - During mating in hamsters, both tactile and nontactile sensory stimulation experienced by the female affect sexual behavior and progestational neuroendocrine reflexes. To test the interactions of these types of mating stimulation, c-Fos immunohistochemistry measured brain cellular activity during sexual behavior under conditions that included combinations of tactile and nontactile mating stimulation. Test groups received: (1) mating stimulation from a male, females being either fully mated or mated while wearing a vaginal mask, or (2) experimenter applied manual vaginocervical stimulation (VCS)-with or without males present, or (3) handling similar to VCS but without insertions-with or without males present. Numbers of c-Fos immunoreactive cells were counted in specific subdivisions of the posterior medial amygdala (MeP) and ventromedial hypothalamus (VMH). The medial amygdala dorsal and ventral subdivisions responded differentially to components of mating stimulation. The posterodorsal Me (MePD) cellular activation was greatest during mating conditions that included VCS and/or males present. However, the posteroventral Me (MePV) was sensitive to male exposure and not to VCS. Also, MePV and VMH shell responses mirrored each other, both being primarily sensitive to male exposure. In separate tests, manual VCS induced pseudopregnancy, though the procedure was most effective with additional nontactile stimulation from males present. In summary, contextual cues provided by nontactile male stimulation enhance the effect of vaginocervical and other tactile stimulation on reproductive processes. Furthermore, c-Fos expression in the female hamster medial amygdala is region and context dependent. PMID- 15664026 TI - Arginine vasotocin activates advertisement calling and movement in the territorial Puerto Rican frog, Eleutherodactylus coqui. AB - Arginine vasotocin (AVT) is a neuropeptide that modulates social behavior in amphibians and activates calling in frogs. The Puerto Rican coqui frog, Eleutherodactylus coqui, is a terrestrial anuran that exhibits complex social behaviors, including territoriality and paternal care. Males have a distinctive social hierarchy and can be any of the following: satellite (non-calling), territorial (calling), or paternal (guards and broods embryos). Field experiments were conducted to determine the effects of AVT on satellite behavior and the male social hierarchy of E. coqui. Satellite males were captured within the territory held by a resident male, given injections (i.p.) of AVT or saline (control) and placed back in their original location. To determine if AVT affects all males, not merely satellite males, territorial (calling) males were injected with AVT. Significantly more satellite males commenced advertisement calling following AVT injections than did control males injected with saline. AVT-activated satellites did not challenge the resident territorial male for possession of the territory but instead moved into a new area before commencing to call. In fact, AVT activated satellite males were significantly more likely to move into a new territory following AVT injections than AVT-injected territorial males. The effect of AVT was short lived, lasting only one night in all but two cases. It is concluded that AVT stimulates advertisement calling and AVT-activated males displayed territorial characteristics of E. coqui. PMID- 15664027 TI - Is there an effect of prenatal testosterone on aggression and other behavioral traits? A study comparing same-sex and opposite-sex twin girls. AB - Men and women differ in temperament and personality traits, such as aggression and sensation seeking. The sex hormone testosterone could play a role in the origin of these differences, but it remains unclear how and when testosterone could have these effects. One way to investigate the prenatal exposure effect of testosterone is to compare opposite-sex (OS) and same-sex (SS) female twins. It has been suggested that OS twin girls are exposed prenatally to elevated testosterone levels and that this may result in some masculinization of their personality and behavior. We measured sexually dimorphic traits and circulating testosterone levels in 13-year-old OS (n = 74) and SS (n = 55) twins. Testosterone levels showed a clear circadian rhythm, with higher levels in the morning than in the afternoon. Testosterone was higher in boys than girls, but similar in OS and SS twin girls. Testosterone was not in any way systematically related to the different personality traits. However, a sex difference in aggression proneness was observed, and OS girls showed a more masculine pattern of aggression proneness than the SS girls. It is argued that it is unlikely that this difference is due to social factors, such as a gender-specific upbringing. PMID- 15664028 TI - A complete rezoning of the Great Barrier Reef Marine Park. PMID- 15664029 TI - A UK perspective on the development of marine ecosystem indicators. AB - This paper reviews the suite of marine ecosystem indicators currently in use or under development in the UK to support the major national and international biodiversity and ecosystem policies. Indicators apply to a range of different ecosystem components, and range from those that can only be used for high level environmental health monitoring, to those which actively support management. Assessment of indicators against a management framework of driving force, pressure, state, impact and response, has shown that there are many indicators of state for ecosystem components, but relatively few for pressure of human activities on the environment, or of the socio-economic response to those pressures. This outcome, a result of unplanned sectorally driven indicator development, is not a co-ordinated contribution to marine environmental management and must be addressed if we are to avoid high monitoring costs and duplication of effort. PMID- 15664030 TI - Byssal detachment underestimates tolerance of mussels to toxic compounds. AB - Mussels are sedentary organisms attached to solid substrata by means of byssus threads. Mussels detached from their substratum tend to reattach by producing new byssus threads. Therefore, in bioassays using mussels, if the test animals are in an unattached status, increased byssogenic activity would expose their soft body parts to the toxic compound used. We test the tolerance of two mussel species (Mytilopsis leucophaeata and Mytilus edulis) to an oxidizing biocide (chlorine) and show that detached mussels are 24-28% less tolerant than byssally attached ones. Detached mussels also showed higher oxygen consumption, filtration rate, foot activity, byssus thread production and byssogenesis index, parameters which are associated with valve opening. A review of literature showed that most of the published data on toxicity against heavy metals and biocides are generated using unattached mussels. The data, therefore, represent an underestimation of the toxicity of the tested compound. Correction data are presented for chlorine tolerance of Dreissena polymorpha, Mytilopsis leucophaeata and Mytilus edulis. The present study suggests the importance of maintaining mussels in their attached status, while undertaking toxicity bioassays. PMID- 15664031 TI - The role of organochlorines in cancer-associated mortality in California sea lions (Zalophus californianus). AB - Wild California sea lions (Zalophus californianus) have an unusually high prevalence of neoplasms (18% of stranded dead adults) and high levels of contaminants. The contribution of organochlorine (OC) tissue burdens to the probability of sea lions dying from carcinoma was explored using a logistic regression model. Levels of PCBs and DDTs were determined in blubber of sea lions diagnosed with metastatic carcinoma and animals that had died from non-carcinoma related incidents (e.g., gunshot, domoic acid poisoning). Animals with carcinoma had higher mean concentrations (based on wet weight) of PCBs and DDTs (more than 85% and 30% higher, respectively) in blubber than did sea lions without carcinoma; the highest concentrations of OCs in the sea lions affected with carcinoma were measured in the males. Blubber thickness was significantly different between the two groups of sea lions, but after controlling for this difference, there was still a significant effect of PCBs, but not DDTs, on the probability of sea lions dying with carcinoma. Age, sex, mass and length did not affect the probability of dying from carcinoma. PMID- 15664032 TI - Development of macrofaunal communities on dredged material used for mudflat enhancement: a comparison of three beneficial use schemes after one year. AB - In recent years, dredged material has become regarded as a potential resource and used to create and/or improve intertidal habitats ('beneficial use' schemes). This paper presents the results of a sampling programme to investigate the short term macrofaunal recovery of three beneficial use schemes in south-east England in terms of species and functional diversity. Environmental parameters (sediment redox potential, and water, organic carbon and silt/clay contents) and univariate community attributes (total individuals and species, diversity, evenness and biomass) at the recharge sites had attained reference levels at two schemes while assemblages differed significantly in terms of species composition at all three schemes. While trophic group proportionality had re-established at one scheme, an increased grazer dominance was apparent at another while the proportion of sub surface deposit feeders decreased at the third. Total individuals and species number of the developing communities were negatively correlated with sediment redox potential at 4 cm and % silt/clay, respectively. The implications of these results for monitoring the recovery of future fine-grained beneficial use schemes are discussed. PMID- 15664033 TI - Nitrogen and phosphorus budget in coastal and marine cage aquaculture and impacts of effluent loading on ecosystem: review and analysis towards model development. AB - Being an essentially open system, cages are usually characterized by a high degree of interaction with environment and cage systems are highly likely to produce large bulk of wastes that are released directly into the environment. Therefore, large-scale cage aquaculture development has been put into question and concerns have been raised that cage aquaculture produces large bulk of wastes that are rich in organic matter and nutrients and are released into coastal and nearshore environment. Recent information on cage aquaculture nutrient budget is scarce and most published reports are dated. This paper reviews cage aquaculture nutrient budget and nutrient loadings and propose a model for nutrient (nitrogen, N and phosphorus, P) budget in a hypothetical cage aquaculture farm with values of feed loss, FCR (feed conversion ratio) and nutrient contents in feed and fish taken from published literature in order to calculate the amount (kg) of N and P produced and released to the environment for each ton of fish produced. The paper proposes, in addition, a critically analyzed nutrient budget based on the dry matter conversion rate instead of the usual feed conversion rate. The conceptual model shows that 132.5 kg N and 25.0 kg P are released to the environment for each ton of fish produced; these values are as high as 462.5 kg N and 80.0 kg P when calculated on the basis of dry matter conversion rate instead of usual feed conversion rate. Thus, the annual global N and P loadings from cage aquaculture (10,000 tons fish and 3000 tons dry matter) are 1325 tons N and 250 tons P and 1387.5 tons N and 240.0 tons P based on usual feed conversion rate and dry matter conversion rate respectively. The paper also proposes, by analyzing the existing data, an FCR-based regression model for predicting nutrient loadings for a given diet. Finally, attempt was made to calculate the annual global loading and release of N and P from cage aquaculture to the coastal and marine environment, the potential impacts of nutrient loading on the ecosystem were discussed and critical points to be considered for minimizing nutrient output in cage aquaculture were suggested. PMID- 15664034 TI - Metal speciation and environmental impact on sandy beaches due to El Salvador copper mine, Chile. AB - Several coastal rocky shores in northern Chile have been affected by the discharges of copper mine tailings. The present study aims to analyze the chemical speciation of heavy metals in relation to the diversity of sessile species in the rocky intertidal benthic community on the northern Chilean coast, which is influenced by the presence of copper mine tailings. In particular, the chemical forms of Cd, Cu, Fe, Mn, Ni, Pb and Zn in beach sediment samples collected in the area influenced by El Salvador mine tailings were studied using a sequential chemical extraction method. In general, all the elements present a maximum concentration in the area near the actual discharge point (Caleta Palito). With regard to Cu and Mn, the concentrations range between 7.2-985 and 746-22,739 microg/g respectively, being lower than background levels only in the control site of Caleta Zenteno. Moreover, the correlation coefficients highlight that Fe, Mn and Ni correlate significantly and positively in the studied area, showing a possible common, natural origin, whilst Cu shows a negative correlation with Fe, Mn and Ni. It could be possible that Cu has an anthropogenic origin, coming from mining activity in the area. Cd, Fe, Mn, Ni, Pb and Zn are mostly associated with the residual phase, whilst Cu presents a different speciation pattern, as resulted from selective extractions. In fact, Cu is highly associated with organic and exchangeable phases in contaminated localities, whilst it is mainly bound to the residual phase in control sites. Moreover, our results, compared to local biological diversity, showed that those sites characterized by the highest metal concentrations in bioavailable phase had the lowest biodiversity. PMID- 15664035 TI - Dissolved oxygen--a target indicator in determining use of the Gulf of Kachchh waters. AB - Spatial and temporal variations of Dissolved Oxygen (DO) and Biochemical Oxygen Demand (BOD) in the Gulf of Kachchh (GoK), India are assessed based on data collected since 1976. DO concentrations in the interior GoK vary within a narrow range (6.05-6.86 mg l(-1)), whereas near-shore waters show wider variations (3.5 7.8 mg l(-1)). The DO concentration levels in the Gulf waters are close to saturation, varying from 75.4% to 108.6%, which is much higher than the 60% saturation level set for the SW-1 waters. BOD-DO simulation shows that when effluents containing a BOD concentration of 50.0 mg l(-1) are introduced at three select locations, DO reduces from 5.1 mg l(-1) to 3.1, 5.0 and 3.8 mg l(-1), respectively, indicating a strong sensitivity to effluent load. Based on the DO analysis, BOD assimilation capacity of coastal waters around the Marine National Park (MNP) and Marine Sanctuary (MS) in the Gulf is determined. DO available for utilization for various categories of water use is illustrated, keeping in mind the ecology of MNP and MS areas. The significance of DO as a target indicator to zone the Gulf for different water use, is highlighted in this study. PMID- 15664036 TI - Occurrence and distribution antibiotic resistance of heterotrophic bacteria isolated from a marine beach. AB - Antibiotic resistance of heterotrophic bacteria isolated from a sandy beach in Sopot, at the Southern Baltic Sea coast was determined. The levels of resistance of bacteria to various antibiotics differed considerably. Bacteria inhabiting the middle part of the beach and the dune were most resistant; the least resistant were bacteria isolated from the sea-beach contact zone. Generally, there were no significant differences in antibiotic resistance between pigmented and non pigmented bacteria. Bacteria isolated from the surface layer of the sand were more resistant to the tested antibiotics than bacteria from the subsurface layers. The majority of bacterial strains were resistant to 3-8 antibiotics. Bacterial resistance to antibiotics was dependent on their chemical structure. PMID- 15664037 TI - Tendencies in the numbers of beverage containers on the Polish coast in the decade from 1992 to 2001. PMID- 15664038 TI - Phytoplankton succession during a red tide of Skeletonema costatum in Jiaozhou Bay of China. PMID- 15664039 TI - Persistent organic pollutants in marine birds, arctic hare and ringed seals near Qikiqtarjuaq, Nunavut, Canada. PMID- 15664040 TI - Polycyclic aromatic hydrocarbons in capelin (Mallotus villosus) in the Barents Sea by use of fixed wavelength fluorescence measurements of bile samples. PMID- 15664041 TI - Expertise in coastal zone environmental impact assessments. PMID- 15664042 TI - Nutrient enrichment and coral reproduction: between truth and repose (a critique of Loya et al.). PMID- 15664044 TI - Adaptation of an Invader assay for the detection of African swine fever virus DNA. AB - A closed tube isothermal Invader assay (Third Wave Technologies Inc., Madison, Wisconsin, USA) was adapted for the detection of African swine fever virus (ASFV) DNA. Several ASFV Invader assays were designed successfully and tested on a real time PCR instrument (iCycler, BioRad). The assay exhibiting the lowest signal/noise ratio (VP73 ASFV Invader Assay) was analysed further using serial 10 fold dilutions of Lisbon 60 ASFV viral genome. The assay sensitivity was determined to be in the order of 2500 copies of ASFV DNA and showed a dynamic range of 4 logs, from 2.5x10(6) to 2500 copies. The high specificity of the test was demonstrated by the lack of cross-reactivity to the clinically similar but heterologous virus, classical swine fever virus. The sensitivity of the Invader assay is sufficient for the testing of acutely infected viremic animals in which the viral load will be high. The robustness and ease of use of the ASFV Invader assay, combined with the possibility to run and read the assay using simple and relatively inexpensive equipment, makes it suitable for laboratories lacking containment facilities and/or real-time PCR instrumentation or on a regional basis for on-site diagnosis close to putative sites of ASFV outbreaks. PMID- 15664045 TI - Real-time PCR for quantitation of porcine reproductive and respiratory syndrome virus and porcine circovirus type 2 in naturally-infected and challenged pigs. AB - Real-time PCR assays were developed for quantitative detection of porcine reproductive and respiratory syndrome virus (PRRSV) and porcine circovirus type 2 (PCV2). The established real-time PCR for the quantitation of PRRSV cDNA and PCV2 DNA were found to be in the 9-log(10) linear dynamic range with excellent linearity and reliable reproducibility. Using these techniques, the distribution and quantitation of PRRSV and PCV2 in naturally infected and challenged pigs were investigated. The viral concentrations were expressed as the mean log(10) viral DNA or cDNA copy numbers per mg or ml of tested samples. For pigs infected naturally with both viruses, the lung, spleen, tonsil and lymphoid organs had the highest viral burdens with ranges from 5.73 to 8.38 and 5.65 to 6.91 for PRRSV and PCV2, respectively. The injection of formalin-inactivated Salmonella choleraesuis emulsified in complete Freund's adjuvant 1 week before and after the inoculation of both viruses resulted in PRRSV replication enhancement 2 weeks post-challenge. However, this facilitated the clearance of PRRSV 4 weeks post challenge. Results from this study show that the established quantitative PCR could be a useful tool when applied to vaccine development and pathogenesis studies in the future. PMID- 15664046 TI - Identification of Dobrava, Hantaan, Seoul, and Puumala viruses by one-step real time RT-PCR. AB - We developed four assays for specifically identifying Dobrava (DOB), Hantaan (HTN), Puumala (PUU), and Seoul (SEO) viruses. The assays are based on the real time one-step reverse transcriptase polymerase chain reaction (RT-PCR) with the small segment used as the target sequence. The detection limits of DOB, HTN, PUU, and SEO assays were 25, 25, 25, and 12.5 plaque-forming units, respectively. The assays were evaluated in blinded experiments, each with 100 samples that contained Andes, Black Creek Canal, Crimean-Congo hemorrhagic fever, Rift Valley fever and Sin Nombre viruses in addition to DOB, HTN, PUU and SEO viruses. The sensitivity levels of the DOB, HTN, PUU, and SEO assays were 98%, 96%, 92% and 94%, respectively. The specificity of DOB, HTN and SEO assays was 100% and the specificity of the PUU assay was 98%. Because of the high levels of sensitivity, specificity, and reproducibility, we believe that these assays can be useful for diagnosing and differentiating these four Old-World hantaviruses. PMID- 15664047 TI - Construction and properties of a gene-silencing vector based on Poplar mosaic virus (genus Carlavirus). AB - A gene-silencing vector based on a full-length genomic clone of Poplar mosaic virus (PopMV) was constructed, with coat protein and movement protein genes removed, and containing instead, the coding sequence for green fluorescent protein (GFP). This paper demonstrates that the PopMV-derived gene-silencing vector was able to silence GFP expression in GFP transgenic Nicotiana benthamiana plants. The full-length genome of an Oxford isolate of PopMV (PV275) was cloned and sequenced. A full-length PopMV clone, under transcriptional control of the 35SCaMV promoter was then constructed, and the clone was able to replicate locally in Nicotiana species. Several autonomous plant RNA and DNA viruses have been converted into vectors and implemented for virus-induced gene-silencing (VIGS) of transgenes and endogenous genes [Burton, R., Gibeaut, D., Bacic, A., Findlay, K., Roberts, K., Hamilton, A., Baulcombe, D., Fincher, G., 2000. Virus induced silencing of a plant cellulose synthase gene. Plant Cell 12, 691-706; Dalmay, T., Horsefield, R., Braunstein, T.H., Baulcombe, D.C., 2001. SDE3 encodes an RNA helicase required for post-transcriptional gene silencing in Arabidopsis. EMBO J. 20, 2069-2077; Gossele, V., Fache, I., Meulewaeter, F., Cornelissen, M., Metzlaff, M., 2002. SVISS--a novel transient gene silencing system for gene function discovery and validation in tobacco plants. Plant J. 32, 859-866; Holzberg, S., Brosio, P., Gross, C., Pogue, G.P., 2002. Barley stripe mosaic virus-induced gene silencing in a monocot plant. Plant J. 30, 315-327; Ratcliff, F., Martin-Hernandez, A., Baulcombe, D., 2000. Tobacco rattle virus as a vector for analysis of gene function by silencing. Plant J. 25, 237-245; Ruiz, M., Vionnet, O., Baulcombe, D., 1998. Initiation and maintenance of virus-induced gene silencing. Plant Cell 10, 937-946]. The use of a virus that naturally infects trees as a gene-silencing vector has not been demonstrated before. The ability to systemically silence a plant transgene following the production of a gene-silencing signal from a locally replicating viral-construct derived from a carlavirus has not to our knowledge been shown before. PMID- 15664048 TI - Investigation of the morphology of cell clones, derived from the mammalian EBTr cell line and their susceptibility to vaccine avian poxvirus strains FK and Dessau. AB - The ability for replication of vaccine avian pox viral strains FK and Dessau in cell clones, derived from the EBTr cell line, derived from embryonic bovine trachea, was studied. The derived seven cell clones showed different morphological characteristics and diverse sensitivity to both vaccine avian pox viral strains. Hence, the EBTr-derived cell clones could be used for cultivation, as well as for differentiation of vaccine avian pox viral strains. In addition, studies have been undertaken to elucidate the possible use of cultivated strains in these heterologous cell culture system's vaccine avian pox viral strains for biotechnology, as well as for solving problems, related to infection of people with avian viruses. PMID- 15664049 TI - A multiplex RT-PCR assay capable of distinguishing beet necrotic yellow vein virus types A and B. AB - A multiplex reverse-transcription polymerase chain assay (mRT-PCR) was developed, based on primers designed to distinguish the A and B types of beet necrotic yellow vein virus (BNYVV). RNA was extracted from 72 BNYVV isolates from Asia, Europe and North America, and the type of each isolate determined using an established detection method based on single strand conformation polymorphisms (SSCPs). An area of the 'triple gene block' region on RNA 2 was amplified and sequenced from 16 isolates of the A and B types. These sequences were aligned and two sets of PCR primers were designed to amplify unique areas common to each type. The A type assay produced a single 324 base-pair RT-PCR fragment when positive samples were amplified. The B type assay produced a 178 base-pair product from positive samples. No amplification was observed from healthy Chenopodium quinoa or sugar beet plants and from plants infected by others sugar beet soil-borne viruses. Fragment length differed sufficiently to allow both assays to be run in a single PCR tube. The results obtained using the new multiplex RT-PCR assay were consistent with those from the established SSCP method for all 72 reference samples. PMID- 15664050 TI - Simultaneous detection of six stone fruit viruses by non-isotopic molecular hybridization using a unique riboprobe or 'polyprobe'. AB - A new strategy for the simultaneous detection of plant viruses by molecular hybridization has been developed. Two, four or six viral sequences were fused in tandem and transcribed to render unique riboprobes and designated as 'polyprobes'. The 'polyprobe four' (poly 4) covered the four ilarviruses affecting stone fruit trees including apple mosaic virus (ApMV), prunus necrotic ringspot virus (PNRSV), prune dwarf virus (PDV), and American plum line pattern virus (APLPV) whereas the 'polyprobe two' (poly 2) was designed to detect simultaneously, plum pox virus (PPV) and apple chlorotic leaf spot virus (ACLSV), the two more important viruses affecting these trees. Finally, a 'polyprobe six' (poly 6) was generated to detect any of the six viruses. The three polyprobes were comparable to the individual riboprobes in terms of end-point dilution limit and specificity. The validation of the new simultaneous detection strategy was confirmed by the analysis of 46 field samples from up to seven different hosts collected from 10 different geographical areas. PMID- 15664052 TI - Development of a TaqMan RT-PCR assay without RNA extraction step for the detection and quantification of African Chikungunya viruses. AB - Chikungunya virus (CHIKV), a member of the alphavirus genus, is of considerable public health concern in Southeast Asian and African countries. However, despite serological evidence, the diagnosis of this arthropod-borne human disease is confirmed infrequently and needs to be improved. In fact, illness caused by CHIKV can be confused with diseases such as dengue or yellow fever, based on the similarity of the symptoms, and laboratory confirmation of suspected cases is required to launch control measures during an epidemic. Moreover, no quantitative molecular tool is described to study CHIKV replication or detection in clinical samples and cell culture supernatants. In this study, a specific and sensitive CHIKV one-step TaqMan RT-PCR assay was developed as a tool for the diagnosis of African CHIKV as well as a rapid indicator of active infection by quantifying viral load. This study also showed that a simple heat viral RNA release during the reverse transcription step constituted an alternative to the conventional RNA extraction method. PMID- 15664051 TI - Quantitation of defective and ecotropic viruses during LP-BM5 infection by real time PCR and RT-PCR. AB - Murine AIDS (MAIDS) is a pathology induced by the LP-BM5 murine leukaemia virus mixture in susceptible strains of mice such as C57BL/6J resulting in lymphoproliferation and progressive immunodeficiency. The etiologic agent of this pathology is BM5d, a replication defective virus. BM5e is a replication competent virus in the viral mixture that functions as a helper virus. This paper describes real time PCR and RT-PCR assays for quantitation of the proviral DNA and viral RNA of BM5d and BM5e. Data is presented describing the change in BM5d and BM5e proviral DNA levels and viral RNA levels in both blood and spleen in the first 8 weeks of infection. Infected mice have increasing levels of BM5d and BM5e viral DNA and RNA detectable from as early as 2 weeks post infection. Similar levels of proviral DNA was found for BM5d and BM5e in PBMC and spleen, however higher levels of BM5e viral RNA were observed in both tissues throughout infection. The assays described can be used as both a diagnostic tool and to investigate the direct effect of treatments on the BM5d and BM5e viruses and MAIDS development. PMID- 15664053 TI - The use of reverse transcriptase for efficient first- and second-strand cDNA synthesis from single- and double-stranded RNA templates. AB - Molecular characterization of eight distinct, difficult-to-clone RNA plant viruses was accomplished after the development of a reverse transcriptase-based first- and second-strand cDNA synthesis method. Double-stranded (ds) RNA templates isolated from strawberry and blackberry and several herbaceous hosts (mint, pea and tobacco) were cloned using this method. Templates, combined with random primers, were denatured with methyl mercuric hydroxide. Reverse transcriptase was added followed by the addition of RNase H. The resulting dsDNA was then digested with restriction endonucleases to produce shorter fragments that could be cloned efficiently into a T-tailed vector after adding an A overhang using Taq polymerase. This procedure resulted in a high number of cloned fragments and allowed insert sizes up to three kilobase-pairs. Unlike traditional cDNA construction methods, there is no need for additional enzymes/steps for second-strand synthesis, PCR amplification or prior sequence information. Synthesis and cloning of cDNA derived from dsRNA templates is much more efficient than with previously described methods. This procedure also worked well for cloning gel-purified dsRNA and with single-stranded RNA templates. PMID- 15664054 TI - The heminested RT-PCR for the study of rabies virus pathogenesis. AB - The aim of the present trial was to evaluate the heminested RT-PCR for the study of rabies virus distribution in mice inoculated experimentally. Inoculation was by the intramuscular route in 150 mice, using the dog street rabies virus. Groups of five animals were killed at different times. Fragments of different organs were collected and the material was tested by Fluorescent Antibody Test (FAT) and heminested RT-PCR (hn RT-PCR). Positive results were obtained beginning on the 10th day after inoculation in the brain, spinal cord, salivary gland, limbs, lungs, liver, spleen, urinary bladder, tongue and right kidney. Hn RT-PCR was shown to be more efficient for the study of rabies virus distribution in different tissues and organs. PMID- 15664056 TI - A new cell-based assay for measuring the forward mutation rate of HIV-1. AB - Over 20 years into the ever-worsening AIDS pandemic, genetic variation remains the greatest obstacle for treating and preventing HIV-1 infection. Mutation rate assays for HIV-1 have been reported; however, none measure directly the forward mutation rate during replication of the virus in cell culture while still retaining the ability to propagate and further study mutant proviruses. Therefore, the objective of the current study was to develop such a phenotypic cell-based assay for measuring the forward mutation rate of HIV-1. Conventional recombinant DNA techniques and polymerase chain reaction were used to create a replication defective HIV-1 vector, pNL4-3Delta+cass, which is based on the NL4-3 strain and contains the thymidine kinase gene from human herpes virus type 1 as the mutational target. A series of transfection and infection steps were used to introduce the vector into 143B cells, which are negative for thymidine kinase function, and produce vector virus for a single cycle of replication. Viral titers were measured by counting the number of drug resistant colonies on the assay plates, and forward mutation rates were calculated from the viral titers. Mutant proviruses were sequenced to determine the types of genetic alterations that occurred. The average forward mutation rate for HIV-1 was 2.2 x 10( 5)mutations/base/cycle. The majority of mutations were base substitutions, including high frequencies of C-->U and G-->A transitions. Single adenosine insertions were also observed frequently. The new assay is economical and provides a direct measurement of the mutation rate during a single cycle of viral replication. Target cells containing mutant proviruses survive the drug selection process and may be propagated for further analysis. The new assay is novel and has many advantages over previous mutation rate assays and thus will be very useful in future studies on genetic variation of HIV-1. PMID- 15664055 TI - Development of a polymerase chain reaction for the detection of Anguillid herpesvirus DNA in eels based on the herpesvirus DNA polymerase gene. AB - Anguillid herpesvirus (AnHV, also known as Herpesvirus anguillae or HVA) is found in both Japanese and European eels. Based on restriction enzyme analysis a small number of differences were found between AnHV isolated from Japanese eels and from European eels. The total genome size of both is about 245 kb, which is confirmed by alternating-field electrophoresis. Using a set of degenerate primers based on conserved regions within DNA-directed DNA polymerase coding regions, a 463 base pair fragment was isolated from both Japanese and European AnHV. Nucleotide sequence analysis showed that the cloned regions of both viruses have identical sequences. Based on this part of the DNA-polymerase sequence, primers were selected and used to develop a sensitive PCR to detect AnHV DNA in eel tissue samples. To avoid false negative results and to estimate the number of AnHV genome copies found in tissues, 100 copies of an internal control plasmid were added to the tissue samples. This semi-quantitative AnHV PCR can be used for both the European and Japanese isolates of AnHV, detects as few as 10 genome copies and is 100 times more sensitive than standard virus isolation. PMID- 15664057 TI - Evaluation of the NucliSens EasyQ assay in HIV-1-infected individuals in South Africa. AB - We compared the performance of the NucliSens EasyQ assay (bioMerieux) combined with the manual NucliSens miniMag extraction methodology to the Roche Cobas Ampliprep/Standard Amplicor Monitor methodology (Roche Diagnostics) for HIV-1 RNA quantitation in HIV-1-infected individuals in South Africa. Plasma samples (284) from HIV sero-positive patients at different stages of infection were analyzed. The distribution of results was typical of the clinical samples received at the laboratory where 20% have viral load results <400 copies/ml (2.6 log) and 18% have viral load results >750000 copies/ml (5.8 log) using the Roche Amplicor Monitor standard assay. All statistical analyses were performed using log10 transformed values for all the variables in the analyses, i.e. log10EasyQIU/ml, and log10RNA (log10 copies/ml, Amplicor). Roche values were converted from RNA copies per ml to IU/ml by multiplying the Roche value by 0.51. HIV RNA levels quantitated by the NucliSens EasyQ assay correlated significantly with those of the Roche Cobas Amplicor Monitor assay (r=0.874, p<0.0001). Reproducibility of the NucliSens EasyQ assay in the log6IU range yielded CV variance of 1.3-2.84% for two well-trained technologists. In addition, a retrospective evaluation of the performance of the NucliSens EasyQ assay in 102 runs (2448) samples was conducted in the laboratory over a 4-month interval. Factors considered during this evaluation included time taken to perform the assay, volume requirements, number of required repeats, potential for contamination. PMID- 15664058 TI - A mRNA PCR for the diagnosis of feline infectious peritonitis. AB - A reverse transcriptase polymerase chain reaction (RT-PCR) for the detection of feline coronavirus (FCoV) messenger RNA in peripheral blood mononuclear cells (PBMCs) is described. The assay is evaluated as a diagnostic test for feline infectious peritonitis (FIP). It is based on a well-documented key event in the development of FIP: the replication of virulent FCoV mutants in monocytes/macrophages. To detect most feline coronavirus field strains, the test was designed to amplify subgenomic mRNA of the highly conserved M gene. The test was applied to 1075 feline blood samples (424 from healthy, 651 from sick cats suspected of FIP) and returned 46% of the diseased cats as positive for feline coronavirus mRNA in their peripheral blood cells; of the healthy cats, 5% tested positive. Of a group of 81 animals in which FIP had been confirmed by post-mortem examination, 75 (93%) tested positive, whereas 17 cats with different pathologies (non-FIP cases) all tested negative. In view of the low rate of false-positive results (high specificity) the mRNA RT-PCR may be a valuable addition to the diagnostic arsenal for FIP. PMID- 15664060 TI - Evaluation of a modified version of Heteroduplex Mobility Assay for rapid screening of HIV-1 isolates in epidemics characterized by mono/dual clade predominance. AB - The geographical distribution of human immunodeficiency virus type 1 (HIV-1) subtypes show, with the exception of some African Countries, a specific pattern. However, the significant phenomenon of migration to Western Countries, coupled to inter-ethnic blending, may result in a constant introduction and spread of novel subtypes and/or recombinant forms in previously homogeneous HIV-1 epidemics. The need to identify and trace these events prompted the development of a rapid and specific bio-molecular tool for the HIV-1 screening, based on the well established Heteroduplex Mobility Assay (HMA). This modified version of HMA (rHMA) has been designed to detect, by a short electrophoretic analysis, HIV-1 isolates remarkably divergent form the local predominant clade, for subsequent more accurate genetic and phylogenetic analyses. The method has been validated for both C2-V5 region of env gene and the p24-p7 region of the gag gene, by proof of-concept experiments performed on a panel of reference standards representing the globally most prevalent HIV-1 subtypes, and applied to screen Italian and Ugandan field isolates. The rHMA experimental conditions identified in this study have been shown to be specific and reliable for both sub-genomic regions of each subtype used. In the context of nationwide monitoring programs, the rHMA may represent a powerful tool for the HIV-1 molecular surveillance in both developed and developing countries, particularly those characterized by mono/dual-clade HIV 1 epidemic, which is relevant for epidemiological studies and for the development of preventive and therapeutic strategies. PMID- 15664059 TI - An efficient virus concentration method and RT-nested PCR for detection of rotaviruses in environmental water samples. AB - Water samples were concentrated by the modified adsorption-elution technique followed by speedVac reconcentration of the filter eluates. Reverse transcriptase nested polymerase chain reaction (RT-nested PCR) was used to detect rotavirus RNA in concentrates of the water. The detection limit of the rotavirus determined by RT-nested PCR alone was about 1.67 plaque forming units (PFU) per RT-PCR assay and that by RT-nested PCR combined with concentration from 1l seeded tap water sample was 1.46 plaque forming units per assay. Water samples were collected from various sources, concentrated, and determined rotavirus RNA. Of 120 water samples, rotavirus RNA was detected in 20 samples (16.7%); 2/10 (20%) of the river samples, 8/30 (26.7%) of the canal samples, and 10/40 (25%) of the sewage samples but was not found in any tap water samples (0/40). Only three water samples were positive for rotavirus antigen determined using an enzyme-linked immunosorbent assay (ELISA). Alignment analysis of the sequenced PCR product (346 bp fragment) was performed in eight rotavirus-positive samples using the rotavirus sequence deposited in the GenBank. All samples gave the correct VP7 sequence. Results of analysis showed two samples similar to human rotavirus (97 98%), five similar to rotavirus G9 sequence (94-99%), and one sample similar to animal rotavirus (97%). PCR inhibitors were not observed in any concentrated water samples. In all 20 (of 120) samples where rotaviruses were found, fecal coliforms including Escherichia coli were also found, but of the samples testing negative for rotaviruses, 76 were fecal coliforms positive and 69 were E. coli positive. The combination of the virus concentration method and RT-nested PCR described below made it possible to effectively detect rotaviruses in environmental water samples. PMID- 15664061 TI - Recombinant Jembrana disease virus gag proteins identify several different antigenic domains but do not facilitate serological differentiation of JDV and nonpathogenic bovine lentiviruses. AB - In Indonesia, it is suspected that there are two bovine lentiviruses circulating in the cattle population: a pathogenic Jembrana disease virus (JDV), and a nonpathogenic bovine immunodeficiency-like virus (BIV). Both viruses cross-react antigenically and cannot be differentiated by current serological tests using JDV antigens. To identify possible type-specific epitopes, a series of recombinant protein constructs including the matrix, capsid and nucleocapsid proteins were produced from JDV gag and the expressed proteins were tested by Western blot using JDV and BIV hyperimmune sera. JDV matrix and truncated capsid proteins were recognised by both JDV and BIV hyperimmune sera indicating that there were multiple cross-reactive epitopes present in JDV gag. At least three epitopic regions were identified in these constructs, including the major homology region, by monoclonal antibody binding studies. JDV nucleocapsid recombinant protein was not recognised by either JDV or BIV hyperimmune sera and none of the recombinant gag proteins were able to differentiate between JDV positive sera from Jembrana disease endemic and Jembrana disease-free areas. Additionally, a 40 amino acid recombinant subunit protein encompassing the region recently found to contain an epitope unique to BIV [Zheng, L., Zhang, S., Wood, C., Kapil, S., Wilcox, G.E., Loughin, T.A., Minocha, H.C., 2001. Differentiation of two bovine lentiviruses by a monoclonal antibody on the basis of epitope specificity. Clin. Diagn. Lab. Immunol. 8, 283-287] was tested but was not recognised by either JDV positive sera from Jembrana disease-endemic or Jembrana disease-free areas. PMID- 15664062 TI - Detection of white spot syndrome virus (WSSV) from hemolymph of Penaeid shrimps Penaeus japonicus by reverse passive latex agglutination assay using high-density latex particles. AB - A simple reverse passive latex agglutination (RPLA) method for detecting white spot syndrome virus (WSSV) in the hemolymph of infected Kuruma shrimp (Penaeus japonicus) was developed. It was confirmed that WSSV could be detected from the shrimp hemolymph when the latex particles blocked with a casein protein were used as detection reagent. It became clear from the result of the infection trial that viruses are detectable by RPLA before the appearance of overt symptoms of this disease. In addition, an amplification product of 982 bp (s) derived from WSSV by PCR was detected in all the samples in which WSSV was detected by RPLA. This newly developed RPLA assay can examine many samples in a simple manner since hemolymph can be extracted more easily than any other organs. This assay can be used conveniently for virus detection in the culture pond of shrimps or in the field. PMID- 15664063 TI - Development and evaluation of a method to detect and quantify enteroviruses using NASBA and internal control RNA (IC-NASBA). AB - We have developed a rapid, sensitive, and specific assay for the detection and quantification of enteroviruses using nucleic acid sequence-based amplification (NASBA). The inclusion of an internal control (IC) increased the precision and accuracy of the method over a standard NASBA assay and provided a way to detect assay inhibition. The assay was sensitive to 10 viral particles with amplification and detection occurring in as little as 18 min. The assay detected a variety of different enteroviruses to the exclusion of non-target viruses. The standard NASBA method resulted in predictions of viral load to within an order of magnitude of the expected number, as compared with prediction to within less than a half order of magnitude using the IC-NASBA method. Rapid and sensitive detection of enteroviruses is important in both clinical samples to diagnose illness and in environmental samples to assess risk of wastewater contamination and potential health hazards. PMID- 15664064 TI - New estimation method for highly sensitive quantitation of human immunodeficiency virus type 1 DNA and its application. AB - A new estimation method for quantitation of HIV-1 DNA was established by introducing a pre-quantitation polymerase chain reaction (PCR) before conventional real-time PCR. Two alternative methods for estimating the copy number can be used: the first method utilizes the rate of beta2-microglobulin (beta2M) gene amplification during the pre-quantitation PCR, and the second utilizes a calibration curve of the crossing point of real-time PCR versus the standard HIV-1-plasmid concentration. These methods could be used to reproducibly and accurately detect a provirus density down to five copies/10(6) cells (for methods 1 and 2, inter-assay CV=17 and 16% and accuracy=81 and 92%, respectively). The levels of HIV-1 DNA could be measurable using as little as 100 microl of whole blood or buffy coat cells. Using a combination of a conventional and highly sensitive methods, we found that the amount of HIV-1 DNA ranged from 2 to 5960 copies/10(6) cells (median of 830 copies/10(6) cells) in CD4-positive T lymphocytes isolated from 30 patients responding well to highly active antiretroviral therapy (HAART). Thus, the highly sensitive method developed in this study allows estimation of the HIV-1 reservoirs in peripheral CD4-positive T lymphocytes of patients responding well to HAART. PMID- 15664065 TI - Assays for glucosidase inhibitors with potential antiviral activities: secreted alkaline phosphatase as a surrogate marker. AB - As secretion of the middle (MHBs) glycoprotein of hepatitis B virus is highly dependent upon the action of the host oligosaccharide processing enzymes glucosidase I and II, drugs that inhibit this enzyme have been proposed as potential antiviral agents. To facilitate the identification of new, more effective inhibitors of MHBs secretion, an assay has been developed based on the expression of this glycoprotein alone by transfection of Huh7 hepatoma cells. The data clearly demonstrate that both mono- and di-glycosylated forms of MHBs are produced in this system and both forms are equally dependent upon glucosidase processing for secretion. In addition, inclusion of a co-transfected reporter construct that encodes secreted alkaline phosphatase (SEAP) to permit normalization of transfection revealed that the SEAP gene product was itself sensitive to glucosidase inhibition. This sensitivity also was observed in HepG2 human hepatoma cells. Thus, measuring SEAP secretion may be another method for evaluating glucosidase inhibition. In addition, this finding has important implications for the use of a SEAP reporter in screens of potential antiviral agents. PMID- 15664066 TI - Validation of IgG-sandwich and IgM-capture ELISA for the detection of antibody to Rift Valley fever virus in humans. AB - Rift Valley fever (RVF) virus is an important zoonotic and a potential biothreat agent. This paper describes validation of sandwich and capture enzyme-linked immunoassays (ELISA) based on gamma-irradiated antigens for the detection of RVFV specific IgG and IgM antibody in humans. Validation data sets derived from testing field-collected sera from Africa (n=2400) were dichotomised according to the results of a virus neutralisation test. In addition, sera from laboratory workers immunized with inactivated RVF vaccine (n=93) and serial sera (n=3) from a single RVF case were used. ELISA data were expressed as percentage of high positive control serum (PP). Cut-off values at 95% accuracy level were optimised using the misclassification cost term option of the two-graph receiver operating characteristics analysis. During the routine use of assays there was no evidence for excessive intra- and inter-plate variations within and between runs of assays. At a cut-off of 13.2 PP the sensitivity of the IgG-sandwich ELISA was 100% and specificity 99.95%, while for the IgM-capture ELISA the values were 96.47 and 99.44%, respectively, at a cut-off of 7.1 PP. Compared to the virus neutralisation test, the IgG-sandwich ELISA was more sensitive in detection of immunological responses in vaccines. Following natural infection class-specific antibodies were detected in serum taken 6 days after onset of symptoms. The results demonstrate that both assays will be useful for early diagnosis of infection, epidemiological surveillance and for monitoring of immune response after vaccination. As highly accurate, robust and safe tests, they have the potential to replace traditional diagnostic methods which are unable to distinguish between different classes of immunoglobulins, and pose health risks necessitating their use being restricted to high containment facilities outside RVF endemic areas. PMID- 15664067 TI - Rapid differentiation of avian infectious bronchitis virus isolates by sample to residual ratio quantitation using real-time reverse transcriptase-polymerase chain reaction. AB - A rapid diagnostic assay for differentiating avian infectious bronchitis virus (IBV) isolates was developed. The basis of the assay is the cleavage of target RNA by RNase H mediated by sequence-specific chimeric oligonucleotides followed by sample to residual ratio quantitation (SRRQ) using RRT-PCR. Four serotype specific chimeric oligonucleotides were designed, one each for the Massachusetts, Connecticut, Arkansas, and Delaware/Georgia 98 serotypes, and tested for their ability to mediate specific cleavage of target RNA from known homologous and heterologous strains of IBV. Specific cleavage of target RNAs by each chimeric oligonucleotide was verified using agarose gel analysis and RRT-PCR. There were no non-specific cleavage products. Eight different IBV strains representing seven serotypes were tested and each chimeric oligonucleotide mediated cleavage of target RNA only from strains within the serotype that the chimeric was designed against. The SRRQ assay was evaluated on 15 samples without prior knowledge of their grouping and correctly identified the serotype of each sample. The assay is rapid; six samples can be tested in approximately 4 h. In addition, the primer set amplifies all IBV RNAs tested to date and provides a built in control for detecting IBV whether it is typeable or not. PMID- 15664068 TI - Isolation of recombinant antibodies (scFvs) to grapevine virus B. AB - A panel of 15 recombinant single chain antibodies (scFv) specific to grapevine virus B (GVB) were recovered from a human combinatorial scFv antibody library using the phage display technique against purified virus particles. Two selected scFv-encoding genes were expressed in recombinant Escherichia coli cells as dimeric antibodies. Successful detection of GVB in tissues of herbaceous hosts and grapevine was obtained in a direct binding assay using dimeric scFvs. This reagent was also shown to substitute efficiently for a GVB polyclonal serum in standard DAS-ELISA test used routinely for diagnosis. PMID- 15664069 TI - Automatic quantitation of vacuolar lesions in the brain of mice infected with transmissible spongiform encephalopathies. AB - The spongiform change induced in the brain tissue is one of the major features investigated in the neuropathology in natural and experimental transmissible spongiform encephalopathies (TSEs). In this context, a reproducible quantitation of the magnitude of these vacuolar lesions is described using image analysis techniques. To be exploited successfully, this image analysis must be able to distinguish the vacuolar lesions from vascular elements. The present study describes the different calibration stages of image analysis using hematoxylin eosin stained slices of brain from mice infected with TSEs. In order to select automatically a maximum of vacuoles and to eliminate a maximum of vascular elements, relevant criteria based on the threshold values of the optical density, shape factor and surface of detected objects were determined. Compared to visual scoring, this method has the advantages of enhanced precision of the measure, reproducibility and moreover, the collection of numerical data for more detailed statistical analysis. In addition, an original scale change function is proposed allowing a comparative analysis with values from the visual scoring method. The method of automatic recognition and quantitation of vacuolar lesions described in this paper represent a useful tool for large-scale analysis of spongiform lesions induced by different TSE isolates transmitted to mice. PMID- 15664070 TI - Simple and rapid strategy for genetic characterization of influenza B virus reassortants. AB - Genetic reassortment of influenza viruses is widely used for creating viruses with specific phenotypes. Reassortment of two influenza viruses, each with eight RNA segments potentially yields as many as 256 gene segment combinations. Therefore, confirmation that progeny viruses possess genomes corresponding to the specified phenotypes can be laborious and time-consuming. To establish a convenient method for genotyping influenza virus reassortants, we adapted single strand conformation polymorphism analysis (SSCP) using standard laboratory equipment. By varying the concentration of polyacrylamide between 4-6% and the concentration of glycerol between 5-8% in the gel, together with adding PCR primers to the DNA sample during the denaturing step, optimal conditions can be found for SSCP with little effort. The described method has high accuracy and reliability, and provides a tool for rapid, cost-effective genetic screening and assessment of the purity and genetic stability of the reassortant viruses. This method should be useful in basic research applications and in preparing reassortant viruses for vaccine use. PMID- 15664071 TI - Assessment of human papillomavirus mRNA detection over time in cervical specimens collected in liquid based cytology medium. AB - Little is known about the stability of human papillomavirus (HPV) RNA within cervical samples collected in liquid based cytology (LBC) preservation media. We addressed this by analysing patient LBC specimens for the presence of HPV RNA over a prospective time course. LBC samples in PreservCyt were obtained from seven women referred to colposcopy due to a cytological diagnosis of moderate or severe dyskaryosis. Aliquots were removed and subject to RNA extraction at, 6h (base-line), 4, 7 and 14 days, post-collection. HPV mRNA was detected using the PreTect HPV Proofer, which detects HPV 16, 18, 31, 33 and 45 E6/E7 transcripts and human small ribonucleoprotein U1A mRNA as a sample control. HPV DNA genotyping was also performed at base-line to assess the range of types in our group. In addition to assessment of viral RNA, overall integrity of the cellular RNA extract was analysed by the RNA 6000 pico assay. Control human RNA was amplified successfully in all seven samples at each time point. Five of the seven women were HPV positive for E6/E7 viral transcripts at base-line and positivity was maintained in all five up to 14 days. Although the pattern of cellular RNA profiles generated from the samples was variable, results indicated that this extract could be amenable to gene expression profiling and that degradation did not increase as a result of storage time. It is concluded that HPV RNA in routinely collected LBC specimens in PreservCyt can be detected for at least 14 days from sample collection. PMID- 15664072 TI - Human papillomavirus DNA detection and typing in male urine samples from a high risk population from Argentina. AB - The aim of the present study was to evaluate the first void urine (FVU) as a non invasive sampling method for HPV detection and genotyping in a high-risk population. Men presenting with HPV associated penile lesions or HPV positive partners attending a urological department in La Plata, Argentina were enrolled for HPV detection and genotyping. DNA from 185 first-void urine samples was evaluated for the presence of HPV by nested polymerase chain reaction using MY09/11 and GP05/06 primers. The viral genotype was analyzed by means of the single-stranded conformation polymorphisms (SSCP) method. Seventy-three percent (135/185) of the FVU specimens were positive for HPV-DNA. The viral prevalence in patients with HPV-DNA positive partners was 68.8% (77/112), and 79.5% (58/73) of patients with penile lesions were found to be HPV positive. The most frequent viral type was HPV-11 (26.7%), followed by HPV-6 (23%), HPV-16 (21.5%), HPV-18 (6%), and HPV-31 (4.4%). In this study, 11.1% (15/135) of the HPV positive specimens were double infections. These results indicate that high-risk HPVs can be found in clinical lesions in a high percentage (43.8%), as simple or double infections. In this sense, the male population represents an important reservoir for the virus and may play a role in the transmission and perpetuation of the infection in the general population. The method described below provides a tool for detection and typing of HPV-DNA using samples obtained by non-invasive techniques and thus easy to obtain. PMID- 15664073 TI - High-level expression of recombinant 3AB1 non-structural protein from FMDV in insect larvae. AB - For its potential usefulness in diagnosis, the non-structural protein 3AB1 from foot-and-mouth disease virus was expressed as a soluble protein by using Autographa californica nuclear polyhedrosis virus as a vector. The 3AB1 coding sequence was introduced into AcNPV genome via pBAcPAK3AB1 transfer vector to originate Ac3AB1 recombinant baculovirus of phenotype occ-. Rachiplusia nu larvae were injected with supernatants of Sf9 cells infected with Ac3AB1 and 5 days post infection total protein extracts were obtained. An intense band of approximately 21.5 kDa was observed when total larvae extracts were SDS-PAGE resolved and the recombinant protein detected by an FMDV-infected guinea pig serum. ELISA tests and Western blot experiments were carried out using sera both from FMDV-infected cattle and from vaccinated animals. The recombinant protein was only recognized by sera from infected animals, suggesting that this method of production in insect larvae could be applied to an efficient mass production of proteins of diagnostic interest. PMID- 15664074 TI - Fabrication of amino silane-coated microchip for DNA extraction from whole blood. AB - A simple microchip device for DNA extraction was constructed based on electrostatic interactions between surface amine groups and DNA. Microchannel was fabricated on silicon wafer by photolithography and coated with 3 aminopropyltriethoxysilane (APTES) or 3-[2-(2-aminoethylamino)-ethylamino] propyltrimethoxysilane (AEEA) to introduce amine groups on the surface. Determination of the number of surface amine groups and optimization of DNA capture condition were demonstrated to characterize the microchip. Capacities of capturing DNA were approximately 97 ng/cm2 in APTES and 194 ng/cm2 in AEEA modified microchips, respectively. The amount of DNA captured in the microchip increased depending on surface amine density. Furthermore, DNA extraction using amine-coated microchip from whole blood was examined. Quantification of DNA and proteins in washing or eluting fraction indicates that proteins were removed at washing steps and only DNA was effectively eluted by changing alkalinity of buffer from pH 7.5 to 10.6. The amount of DNA extracted from whole blood was approximately 10 ng and its recovery ratio was 27-40%. Performance of PCR for the eluted fraction indicates that DNA extracted from whole blood was well purified using amine-coated microchip. PMID- 15664076 TI - Expression profiling by oligonucleotide microarrays spotted on coated polymer slides. AB - We have developed a ready-to-spot polymer microarray slide, which is coated with a uniform layer of reactive electrophilic groups using anthraquinone-mediated photo-coupling chemistry. The slide coating reduces the hydrophobicity of the native polymer significantly, thereby enabling robust and efficient one-step coupling of spotted 5' amino-linked oligonucleotides onto the polymer slide. The utility of the coated polymer slide in gene expression profiling was assessed by fabrication of spotted oligonucleotide microarrays using a collection of 5' amino linked 70-mer oligonucleotide probes representing 96 yeast genes from Operon. Two colour hybridizations with labelled cDNA target pools derived from standard grown and heat-shocked wild type yeast cells could reproducibly measure heat shock induced expression of seven different heat shock protein (HSP) genes. Moreover, the observed fold changes were comparable to those reported previously using spotted cDNA arrays and high-density 25-mer oligonucleotide arrays from Affymetrix. The low hybridization signals obtained from the DeltaSSA4 mutant cDNA target, together with the high signal detected in two-colour hybridizations with heat-shocked wild type yeast relative to the DeltaSSA4 mutant strain implies that unspecific binding of cDNA target to the SSA4-specific 70-mer oligonucleotide probes is negligible. Combined, our results indicate that the coated polymer microarray slide represents a robust and cost-effective array platform for pre spotted oligonucleotide arrays. PMID- 15664075 TI - Stable long-term indigo production by overexpression of dioxygenase genes using a chromosomal integrated cascade expression circuit. AB - In our laboratory we have analyzed different factors to maximize the yield in heterologous protein expression for long-term cultivation, by combination of an efficient cascade expression system and stable integration in the bacterial chromosome. In this work, we have explored this system for the production of indigo dye as a model for biotechnological production, by expressing in Escherichia coli the thnA1A2A3A4 genes from Sphingomonas macrogolitabida strain TFA, which encode the components of a tetralin dioxygenase activity. We compared Ptac, and the Pm-based cascade expression circuit in a multicopy plasmid and stably integrated into the bacterial chromosome. Plasmid-based expression systems resulted in instability of indigo production when serially diluted batch experiments were performed without a selective pressure. This problem was solved by integrating the expression module in the chromosome. Despite the gene dosage reduction, the synergic effect of the cascade expression system produced comparable expression to the dioxygenase activity in the plasmid configuration but could be stably maintained for at least 5 days. Here, we show that the cascade amplification circuit integrated in the chromosome could be an excellent system for tight control and stable production of recombinant products. PMID- 15664077 TI - The deletion of the pif gene improves the biosafety of the baculovirus-based technologies. AB - Our goal was to improve the biosafety of baculovirus-based technologies by deleting the pif (per os infectivity factor) gene from baculovirus expression vectors. Such a deletion would block transmission in nature without disturbing protein production. A pif deletion mutant of Autographa californica multiplecapsid nucleopolyhedrovirus (AcMNPV) was constructed and its infectivity to two host species was tested by oral or intrahemocoelic inoculation. Virus replication after oral inoculation was monitored using PCR. Oral inoculations with a mixture of the wild type and the pif deletion viruses were carried out. The pif deletion blocked oral infection but it did not hamper infectivity in cell culture. The blockage took place early after inoculation and could not be overcome by mixed inoculations with the wild type. The cat gene was inserted under the control of the polyhedrin promoter in the deletion mutant and the wild type CAT yield was measured in Spodoptera frugiperda insect cells (Sf9) infected with either recombinant. The pif deletion did not hamper CAT production. This deletion significantly improved CAT yields early in the infection. Hence, expression vectors lacking pif may produce higher quality protein. The pif deletion is a simple measure that dramatically reduces the chances of virus spread or gene transfer in nature. PMID- 15664078 TI - Development of a targeted transgenesis strategy in highly differentiated cells: a powerful tool for functional genomic analysis. AB - Functional genomic analysis is a challenging step in the so-called post-genomic field. Identification of potential targets using large-scale gene expression analysis requires functional validation to identify those that are physiologically relevant. Genetically modified cell models are often used for this purpose allowing up- or down-expression of selected targets in a well defined and if possible highly differentiated cell type. However, the generation of such models remains time-consuming and expensive. In order to alleviate this step, we developed a strategy aimed at the rapid and efficient generation of genetically modified cell lines with conditional, inducible expression of various target genes. Efficient knock-in of various constructs, called targeted transgenesis, in a locus selected for its permissibility to the tet inducible system, was obtained through the stimulation of site-specific homologous recombination by the meganuclease I-SceI. Our results demonstrate that targeted transgenesis in a reference inducible locus greatly facilitated the functional analysis of the selected recombinant cells. The efficient screening strategy we have designed makes possible automation of the transfection and selection steps. Furthermore, this strategy could be applied to a variety of highly differentiated cells. PMID- 15664079 TI - Single nucleotide polymorphism for animal fibre identification. AB - Animal fibres are highly valuable industrial products often adulterated during marketing. Currently, there is no precise method available to identify and differentiate the fibres. In this study, a PCR-RFLP technique was exploited to differentiate cashmere and wool fibres derived from goat and sheep, respectively. The presence of DNA in animal hair shafts has enabled the isolation of DNA from scoured cashmere and wool fibres. The mitochondrial cytochrome b sequences of both species were amplified by PCR using primers designed from conserved regions. The polymorphism observed between the two species was detected by restricting the amplified product by endonucleases viz., BamH1 and Ssp1. The RFLP profile clearly distinguishes the cashmere and wool fibres and this technique can also be exploited to test adulteration in animal fibres qualitatively. PMID- 15664081 TI - Bioleaching of spent refinery processing catalyst using Aspergillus niger with high-yield oxalic acid. AB - A spent refinery processing catalyst was physically and chemically characterized, and subjected to one-step and two-step bioleaching processes using Aspergillus niger. During bioleaching of the spent catalysts of various particle sizes ("as received", 100-150 microm, <37 microm, and x =2.97 (average) microm) and pulp densities, the biomass dry weight and pH were determined. The corresponding leach liquor was analysed for excreted organic acids along with heavy metal values extracted from the catalyst. Chemical characterization of the spent catalyst confirmed the presence of heavy metal including Al (33.3%), Ni (6.09%) and Mo (13.72%). In general, the presence of the spent catalyst caused a decrease in the biomass yield and an increase in oxalic acid secretion by A. niger. The increase in oxalic acid secretion with a decrease in the catalyst particle size (up to <37 microm) led to corresponding increase in the extraction of metal values. The highest extraction of metal values from the spent catalyst (at 1% w/v pulp density and particle size <37 microm) were found to be 54.5% Al, 58.2% Ni and 82.3% Mo in 60 days of bioleaching. Oxalic acid secretion by A. niger in the presence of the spent catalyst was stimulated using 2-[N Morpholino]ethanesulfonic acid (MES) buffer (pH 6), which resulted in comparable metal extraction (58% Al, 62.8% Ni and 78.9% Mo) in half the time required by the fungus in the absence of the buffer. Spent medium of A. niger grown in the absence and in the presence of MES buffer were found to leach almost similar amounts of Al and Ni, except Mo for which the spent medium of buffered culture was significantly more effective than the non-buffered culture. Overall, this study shows the possible use of bioleaching for the extraction of metal resources from spent catalysts. It also demonstrated the advantages of buffer-stimulated excretion of organic acids by A. niger in bioleaching of the spent catalyst. PMID- 15664080 TI - Bioleaching of spent fluid catalytic cracking catalyst using Aspergillus niger. AB - The use of the fungus Aspergillus niger for the bioleaching of heavy metals from spent catalyst was investigated, with fluid catalytic cracking (FCC) catalyst as a model. Bioleaching was examined in batch cultures with the spent catalysts at various pulp densities (1-12%). Chemical leaching was also performed using mineral acids (sulphuric and nitric acids) and organic acids (citric, oxalic and gluconic acids), as well as a mixture of organic acids at the same concentrations as that biogenically produced. It was shown that bioleaching realised higher metal extraction than chemical leaching, with A. niger mobilizing Ni (9%), Fe (23%), Al (30%), V (36%) and Sb (64%) at 1% pulp density. Extraction efficiency generally decreased with increased pulp density. Compared with abiotic controls, bioleaching gave rise to higher metal extractions than leaching using fresh medium and cell-free spent medium. pH decreased during bioleaching, but remained relatively constant in both leaching using fresh medium and cell-free spent medium, thus indicating that the fungus played a role in effecting metal extraction from the spent catalyst. PMID- 15664082 TI - High throughput easy microinjection with a single-cell manipulation supporting robot. AB - A single-cell manipulation supporting robot (SMSR) has been developed for the high throughput and easy microinjection. Its concept is to let an experimenter concentrate his/her attention only on the microinjection by facilitating other associated works. SMSR was applied to the microinjection into rice protoplasts and mouse embryonic stem (ES) cells. The microinjection into these cells is exceptionally difficult than usual animal cells such as fibroblasts. In the case of rice protoplast, for example, non-stop microinjection into 100 cells could be done within 1h that was 17-times faster than that of the robot-less work. The success rate was 7-8% that was same level obtained by the robot-less work. The present results indicate that SMSR is a useful machine for the microinjection of specific genes and proteins in living cells to analyze their respective functions, which is an urgent and important subject in the post-genome era. PMID- 15664083 TI - Adaptive multiscale principal component analysis for on-line monitoring of a sequencing batch reactor. AB - In recent years, multiscale monitoring approaches, which combine principal component analysis (PCA) and multi-resolution analysis (MRA), have received considerable attention. These approaches are potentially very efficient for detecting and analyzing diverse ranges of faults and disturbances in chemical and biochemical processes. In this work, multiscale PCA is proposed for fault detection and diagnosis of batch processes. Using MRA, measurement data are decomposed into approximation and details at different scales. Adaptive multiway PCA (MPCA) models are developed to update the covariance structure at each scale to deal with changing process conditions. Process monitoring by a unifying adaptive multiscale MPCA involves combining only those scales where significant disturbances are detected. This multiscale approach facilitates diagnosis of the detected fault as it hints to the time-scale under which the fault affects the process. The proposed adaptive multiscale method is successfully applied to a pilot-scale sequencing batch reactor for biological wastewater treatment. PMID- 15664085 TI - Burn treatment's evolution in the 20th century. PMID- 15664086 TI - Reconstruction of the common bile duct. PMID- 15664087 TI - The alphabet versus the goddess: The conflict between word and image. PMID- 15664088 TI - Cervical spine evaluation in urban trauma centers: lowering institutional costs and complications through helical CT scan. AB - BACKGROUND: In the evaluation of the cervical spine (c-spine), helical CT scan has higher sensitivity and specificity than plain radiographs in the moderate- and high-risk trauma population, but is more costly. We hypothesize that institutional costs associated with missed injuries make helical CT scan the least costly approach. STUDY DESIGN: A cost-minimization study was performed using decision analysis examining helical CT scan versus radiographic evaluation of the c-spine. Parameter estimates were obtained from the literature for probability of c-spine injury, probability of paralysis after missed injury, plain film sensitivity and specificity, CT scan sensitivity and specificity, and settlement cost of missed injuries resulting in paralysis. Institutional costs of CT scan and plain radiography were used. Sensitivity analyses tested robustness of strategy preference, accounted for parameter variability, and determined threshold values for individual parameters on strategy preference. RESULTS: C spine evaluation with helical CT scan has an expected cost of US 554 dollars per patient compared with US 2,142 dollars for plain films. CT scan is the least costly alternative if threshold values exceed US 58,180 dollars for institutional settlement costs, 0.9% for probability of c-spine fracture, and 1.7% for probability of paralysis. Plain films are least costly if CT scan costs surpass US 1,918 dollars or plain film sensitivity exceeds 90%. CONCLUSIONS: Helical CT scan is the preferred initial screening test for detection of cervical spine fractures among moderate- to high-risk patients seen in urban trauma centers, reducing the incidence of paralysis resulting from false-negative imaging studies and institutional costs, when settlement costs are taken into account. PMID- 15664089 TI - Monitoring performance: longterm impact of trauma verification and review. AB - BACKGROUND: This study documents how the verification process at a Level I pediatric trauma center affected patient care through changes in care indicators (CIs) from predesignation through four postverification time frames. An important component of any verification program is its effectiveness, not only at the time of verification but during the time between "examinations." To date, few data exist describing the interval periods and the progression and maturation of a trauma program after initial verification. STUDY DESIGN: Forty-seven distinct CIs were monitored monthly through data generated from the trauma registry. Six distinct time periods were identified. PRE (January, June, October 1997), trauma care without monitoring; VER (November 1999 to September 2000), preparation for verification; and four postverification periods: P1 (January to June 2001), P2 (July to December 2001), P3 (January to June 2002), and P4 (July to September 2002). RESULTS: Between 1997 and 2002, trauma admissions increased from 200 per year to 313 per year. Mortality rate and Injury Severity Score distributions remained unaltered. Statistically significant (p < 0.05) quantitative and qualitative changes were observed in numbers (percent) of patients reaching clinical criteria. These included prehospital, emergency department, and hospital based trauma competencies. Trauma patient evaluation (including radiology) and disposition out of the emergency department (<120 minutes) improved in each study section and remained high during the postverification time period. There was a strong pair-wise correlation (p < 0.005, Cronbach alpha 0.8) between CNS charting and acquisition of head CAT scans. Pediatric ICU duration of stay increased in both the (summer) P2 and P4 time periods. Prehospital and emergency department fluid monitoring remained unsatisfactory. CONCLUSIONS: Statistically significant changes in patient care indicators were noted to improve during the trauma center designation process, and other key deficiencies were identified and addressed. Maintaining these improvements requires constant monitoring or performance may revert below accepted levels. PMID- 15664090 TI - Postinjury serum secretory phospholipase A2 correlates with hypoxemia and clinical status at 72 hours. AB - BACKGROUND: Although trauma patients often suffer direct lung damage, an equally destructive mechanism of lung injury involves postinjury systemic inflammation. We postulate that secretory phospholipase A(2) (sPLA(2)) release induced by trauma relates to systemic inflammation that compromises both lung function and clinical status after injury. The objectives of this study were: to relate Injury Severity Score to postinjury sPLA(2); to determine whether circulating sPLA(2) relates to pulmonary oxygenation and compliance; and to determine whether early or persistent increases in sPLA(2) are associated with abnormal chest x-ray at 72 hours after injury. STUDY DESIGN: The prospective cohort study comprised 54 consecutive intensive care admissions in patients with traumatic injury admitted over a 6-month period from November 1, 1996, to May 1, 1997. RESULTS: Postinjury peak sPLA(2) values were associated with increased ISS (r = 0.49, r(2) = 0.24, p < 0.001). Patients with elevated sPLA(2) had poor oxygenation compared with those with normal sPLA(2) levels (Pa0(2)/Fi0(2) ratio 164 +/- 16 versus 260 +/- 26 mmHg [mean +/- SEM], p < 0.01) and also required additional PEEP (5.5 +/- 0.9 versus 2.5 +/- 0.4 cm H(2)O, p = 0.01). Secretory PLA(2) levels in patients with abnormal chest x-ray 72 hours after injury were higher (1.08 +/- 0.2 versus 0.34 +/- 0.1 activity units, p < 0.001) than levels seen in patients with normal x rays. CONCLUSIONS: Increasing injury magnitude is associated with elevated sPLA(2) levels, and increased sPLA(2) is related to postinjury hypoxemia and clinical status. PMID- 15664091 TI - The stages of change questionnaire as a predictor of trauma patients most likely to decrease alcohol use. AB - BACKGROUND: Research has shown that negative trauma-related consequences of drinking can predict readiness to change drinking behavior. These findings are confined to patients with positive blood alcohol levels at admission. The current study extends such findings by examining whether stage of readiness to change among all adult trauma activations admitted to the emergency department for 24 hours or more can be used to predict a change in behavior at followup. STUDY DESIGN: Patients 18 years of age or older admitted to a Level II trauma center between December 1, 2001, and January 31, 2003, with a trauma activation were eligible to participate. Enrolled patients were screened for alcohol use and readiness to change with the Alcohol Use Disorders Identification Test (AUDIT) and Short Form Stages of Change (SFSC), respectively, within 2 days of discharge. Blood alcohol level (BAL) was obtained for all study patients. Six to 18 months after discharge, patients were followed up with the AUDIT. The ability of the SFSC to predict change in alcohol behavior at followup was analyzed by multiple regression. RESULTS: Of 253 eligible patients, 146 patients were enrolled and had BAL taken. Most were men (64%) and 36% were women. Mean positive BAL (n = 57) was 186 mg/dL (range 10 to 537 mg/dL). Of these, 23% (13 of 57) met AUDIT criteria for harmful drinking and 47% (27 of 57) met criteria for dependent drinking. Even among those with undetectable BAL (n = 89), harmful or dependent drinking was identified by the AUDIT for 11% (10 of 89). A readiness to change was common among those reporting harmful or dependent drinking (26%) at baseline. The SFSC independently and significantly predicted change in drinking behavior among the 72 patients with followup (p = 0.05). Patients indicating greater readiness to change at baseline were those more likely to decrease their consumption pattern at followup. CONCLUSIONS: Findings suggest that administering the brief SFSC questionnaire to all trauma patients and providing assistance to those demonstrating a willingness to change may prove to be an effective strategy for reducing problem alcohol use. PMID- 15664092 TI - An analysis of the Research Fellowship Scheme of the Royal College of Surgeons of England. AB - BACKGROUND: The Research Fellowship Scheme of the Royal College of Surgeons of England commenced in 1993 with the aim of exposing selected surgical trainees to research techniques and methodology, with the hope of having an impact on surgical research and increasing the cadre of young surgeons who might decide to pursue an academic career in surgery. Over 11 million pounds sterling (approximately US 20 million dollars) has been invested in 264 fellowships. The College wished to evaluate the impact of the Scheme on the careers of research fellows, surgical research, and patient care. As the 10th anniversary of the Scheme approached. STUDY DESIGN: Two-hundred and sixty research fellows whose current addresses were available were sent a questionnaire. Two-hundred and thirty-eight (91.5%) responded. RESULTS: Three-quarters of the research fellows conducted laboratory-based research, with most of the remainder conducting patient-based clinical research. One-third of the fellows who have reached consultant status have an academic component to their post. The total number of publications based on fellowship projects was 531, with a median impact factor of 3.5. Almost all fellows had been awarded a higher degree or were working toward this. Half of the fellows received subsequent funding for research, mostly awarded by national or international funding bodies. CONCLUSIONS: The Research Fellowship Scheme of the Royal College of Surgeons of England has successfully supported many trainee surgeons in the initial phase of their research career. It has helped surgical research by increasing the pool of surgeons willing to embark on an academic career. Indirectly, patient care has benefited by promoting an evidence-based culture among young surgeons. Such schemes are relevant to surgical training programs elsewhere if more young surgeons are to be attracted into academic surgery. PMID- 15664093 TI - Completely laparoscopic total and partial gastrectomy for benign and malignant diseases: a single institute's prospective analysis. AB - BACKGROUND: Laparoscopic surgery for gastrointestinal benign disease has gained worldwide acceptance; totally laparoscopic surgery for malignant diseases remains controversial. The purposes of this study were to examine prospectively our experience with laparoscopic gastric resections, to evaluate the surgical outcomes, and to discuss the role of these procedures in the treatment of benign and malignant diseases of the stomach. To the best of our knowledge, this is the largest prospective study of totally laparoscopic total and partial gastrectomies in Western countries. STUDY DESIGN: Thirty-three patients who underwent totally laparoscopic gastric resection between April 1995 and January 2004 were studied prospectively. Eight patients underwent laparoscopic total gastrectomy and 25 patients had laparoscopic partial gastrectomy. There were 21 women and 12 men with a mean age of 71 +/- 10 years. RESULTS: Twenty-one patients (63.6%) were operated on for malignant diseases and 12 patients (36.4%) had benign lesions. Conversion to laparotomy was not required in any case. Mean operative time was 138 +/- 40 minutes and mean blood loss was 58 +/- 85 mL. There were no major intraoperative complications except for one splenectomy, and there were no perioperative deaths. Two postoperative complications occurred; one patient developed an intraperitoneal abscess with a small duodenal fistula after total gastrectomy and was treated by peritoneal lavage and drain placement. The other patient developed delayed gastric emptying after subtotal gastrectomy and was managed conservatively. Mean ambulation time and mean hospital stay were 2.3 +/- 0.7 days and 14.6 +/- 5 days, respectively. All resected margins were tumor free. The mean number of retrieved lymph nodes for the malignant lesions was 22 +/- 12 (range 10 to 53). CONCLUSIONS: This prospective trial demonstrated that totally laparoscopic total and partial gastric resections had good results and were feasible and safe procedures. In addition, we concluded that the totally laparoscopic approach for early and advanced gastric cancer can obtain adequate margins and follow oncologic principles. PMID- 15664094 TI - Evaluation of continuous infusion of 0.5% bupivacaine by elastomeric pump for postoperative pain management after open inguinal hernia repair. AB - BACKGROUND: Postoperative pain management and narcotic usage after inguinal hernia repair is an important concern for anesthesiologists and surgeons. Regional anesthesia incisional infiltration techniques may reduce the need for systemic medications. STUDY DESIGN: Double-blind study of 52 patients undergoing open hernia repair with the Prolene Hernia System prospectively randomized to receive either 0.5% bupivacaine or saline continuously for 48 hours at 2 mL/hour through use of an elastomeric continuous infusion pump (ONQ Pain Relief System). Outcomes measured over the 120 hours postoperatively were: narcotic use, Visual Analogue Scale pain scores, site inflammation (visual inspection), and side effects. Comparisons between the groups were made by applying the Mann-Whitney rank sum test and analysis of variance with treatment as a fixed effect in the model. RESULTS: In the bupivacaine group, 24% (versus 4% in the placebo group) required no narcotics (p < 0.05). Daily and total narcotic usages for all 5 study days were significantly less (p < 0.05) in the bupivacaine group. There were no reports of complications at the catheter-insertion site or surgical-wound site. CONCLUSIONS: Demonstrable benefits include reduction of narcotic usage and pain with no apparent increase in the risk of infection or complication. Continuous infusion of 0.5% bupivacaine at 2 mL/h through the ONQ pump is a safe and effective adjunct in postoperative pain management for open inguinal hernia repair. This represents a viable and possibly superior option for prolonged pain management, minimizing use of narcotics in patients undergoing this procedure. PMID- 15664095 TI - Management and prognosis of metastases to the thyroid gland. AB - BACKGROUND: Intrathyroid metastases (ITM) of extrathyroid cancer are rare and have a poor prognosis. The aim of this work was to identify the sites of primary cancers and the treatment and prognosis of these lesions. STUDY DESIGN: This retrospective study was carried out on patients treated between 1982 and 2002 in the surgical departments of the University hospitals of Poitiers, Limoges, Tours, and Nantes, France. All diagnoses were confirmed by cytology or histologic examination. RESULTS: Twenty-nine patients (41 to 78 years) had ITM. Primary cancers were renal cell in 16 patients, lung in 4, digestive in 4, sarcoma in 1, melanoma in 1, neuroendocrine in 1, and of unknown origin in 1 patient. For 10 patients, diagnoses of primary cancer and ITM were synchronous. For 19 patients, delay between diagnosis of the primary cancer and ITM was 6.8 years (2 months to 16 years). Diagnosis was confirmed with fine-needle aspiration 3 times and with histologic examination of the thyroid 26 times. Twenty-seven patients had thyroidectomy; two were not operated on. After treatment of ITM, 13 patients had new metastatic sites. Mean followup for all patients was 2.3 years. Seven patients (24%) (6 with renal cancer) were disease free (followup 4.5 years). Four patients were alive with disease (followup 1.4 years). Eighteen patients (62%) died of their disease at a mean delay of 1.4 years. CONCLUSIONS: ITM are rare but the diagnosis should be borne in mind when patients have a history of cancer (mainly renal cancer). Preoperative diagnosis and complete evaluation could avoid unnecessary thyroidectomy because numerous patients had diffuse metastases. PMID- 15664097 TI - Behavioral associations between prostate and colon cancer screening. AB - BACKGROUND: Despite the controversy over the utility of routine prostate-specific antigen (PSA) screening in reducing prostate cancer-specific mortality, it has gained widespread use throughout the United States. Although colorectal cancer (CRC) screening reduces CRC mortality and appears to be cost effective, CRC screening adherence is suboptimal. To better understand screening behaviors among men, the Behavioral Risk Factors Surveillance Survey was used to identify potential relationships that would allow interventions to enhance CRC screening. STUDY DESIGN: We included in our analysis 22,304 men, 50 years and older, who participated in the 2002 Behavioral Risk Factors Surveillance Survey. Chi-square and multivariate analyses were performed to determine the independent correlates of adherence to CRC screening. Independent variables evaluated were age, race, educational level, employment status, income, health insurance, the presence of a personal physician, self-reported general health, current smoking status, and receiving a PSA test. RESULTS: Men were more adherent with PSA screening than CRC screening (50.4% versus 47.6%;p < 0.002). In multivariate analysis, adherence to PSA screening (adjusted odds ratio [OR] 3.24, p < 0.001) exerted the largest independent effect on CRC screening adherence. Other positive correlates of adherence to CRC screening were having health insurance (adjusted OR 1.39, p < 0.01) and a personal physician (adjusted OR 2.01, p < 0.01). Age predicted CRC screening with an inverse-U correlation. Failure to adhere to CRC screening was associated with self-reported good health (adjusted OR 0.87, p < 0.01) and being a current smoker (adjusted OR 0.65, p < 0.01). Even in men who were compliant with PSA testing, CRC screening remained suboptimal (65%). CONCLUSIONS: More men received PSA testing than CRC screening. Men who received PSA testing were more likely to adhere to CRC screening. Taken together, PSA testing may represent a "teachable moment" for a behavior-related intervention aimed at reducing the risk of colon cancer. Targeting men who already accept one form of cancer screening can potentially increase CRC screening adherence. PMID- 15664096 TI - Antioxidants inhibit fatty acid and glucose-mediated induction of neutral endopeptidase gene expression in human microvascular endothelial cells. AB - BACKGROUND: Neutral endopeptidase (NEP) is a membrane-bound metallopeptidase that degrades tachykinins and may regulate their role in wound repair. NEP enzyme activity is increased in diabetic wounds and skin compared with normal controls. We have shown that unsaturated fatty acids and glucose upregulate NEP activity in human microvascular endothelial cells (HMECs) and that vitamins E and C reduce this effect. STUDY DESIGN: To determine whether these changes involve NEP gene expression regulation, we analyzed NEP mRNA levels in HMECs cultured with elevated glucose (40 mM) and fatty acids oleate (40 microM) and linoleate (40 microM) for 48 hours or 1 month. Cells were exposed for an additional 48 hours to antioxidants vitamins E or C or N-acetylcysteine. Total RNA was extracted and analyzed for NEP mRNA using real-time reverse transcriptase polymerase chain reaction. NEP gene expression was standardized to beta-actin mRNA and results were analyzed using ANOVA. RESULTS: Elevated glucose, oleate, and linoleate upregulated NEP mRNA in short and longterm HMEC cultures, but did not alter rate of NEP mRNA degradation. Vitamins E and C and N-acetylcysteine blocked glucose- and fatty acid-induced NEP mRNA (p < or = 0.05). The potential role of oxidative stress in NEP activation was confirmed by demonstrating that elevated glucose and fatty acids increase H(2)O(2) levels in HMECs. CONCLUSIONS: Regulation of NEP enzyme activity by glucose and fatty acids appears to include gene expression transcription as well as modulation of enzyme activity. Our results also suggest that oxidative stress may be involved in upregulation of NEP by fatty acids and glucose. PMID- 15664098 TI - Bisegmentectomy 7-8 as alternative to more extensive liver resections. AB - BACKGROUND: Preservation of hepatic parenchyma should be attempted whenever possible in order to reduce the risk of liver failure and increase the chance to re-resect the recurrence. STUDY DESIGN: The presence of a lesion in segments 7-8 infiltrating the right hepatic vein is usually an indication for right hepatectomy. If a thick inferior right hepatic vein is seen, a bisegmentectomy 7 8 can be performed. We review our experience with this uncommon liver resection. RESULTS: In 11 of 332 patients with colorectal liver metastases, a lesion was localized in segments 7-8 infiltrating the right hepatic vein. Six underwent resection of segments 7-8. The mean estimated rate of remnant liver volume (segments 2-4 plus caudate lobe) was 23.7%; 4 patients had neoadjuvant chemotherapy. Intraoperative mean blood loss was 200 mL without transfusions; no patients developed postoperative liver failure, and there was no in-hospital mortality. Surgical margin was negative in all patients. Median survival was 25 months, with 3 patients alive and disease-free. One patient with an intrahepatic recurrence underwent re-resection. CONCLUSIONS: Bisegmentectomy 7-8 is an uncommon but safe procedure that allows curative resections without unnecessary sacrifice of functional parenchyma. PMID- 15664099 TI - Making surgery safer. PMID- 15664100 TI - Update on bone morphogenetic proteins and their application in spine surgery. PMID- 15664101 TI - Transfusions in surgical patients. PMID- 15664102 TI - Male breast disease. PMID- 15664103 TI - Are bariatric surgical outcomes worth their cost? A systematic review. PMID- 15664106 TI - Part 2: Surgical palliation of advanced illness: what's new, what's helpful. PMID- 15664107 TI - The surgeon and acts of civilian terrorism: biologic agents. PMID- 15664108 TI - Complete colorectal duplication. PMID- 15664109 TI - Posttraumatic pneumatocele. PMID- 15664110 TI - Liver resection using a saline-linked radiofrequency dissecting sealer for transection of the liver. PMID- 15664111 TI - Value of extended lymphadenectomy in laparoscopic subtotal gastrectomy for advanced gastric cancer. PMID- 15664112 TI - Association between apolipoprotein E polymorphism and Alzheimer disease in Tehran, Iran. AB - Epsilon 4 allele of apolipoprotein E (APOE-epsilon4) is a major risk factor for Alzheimer's disease (AD). The association of APOE allele frequencies with AD remains unknown in developing countries. We examined the frequency of APOE alleles in 105 patients with AD and 129 cognitively normal subjects of similar age and sex (control group), in Tehran, Iran. The APOE-epsilon4 allele frequency was significantly higher in the AD subjects than in the control group (21% versus 6.2%, p < 0.001). In addition, the OR for APOE-epsilon4 heterozygous and homozygous subjects were 3.2 (p = 0.001) and 12.75 (p = 0.01), respectively. The OR was not uniform across age groups. The AD subjects carrying one or two APOE epsilon4 allele showed earlier age-at-onset (p < 0.001). These data suggest that the APOE-epsilon4 allele increase the risk for AD in Tehran population in a dose and age-dependent manner. Although the APOE-epsilon2 allele frequency was lower in the AD subjects than in the control group (0.95% versus 2.7%, p = 0.15), APOE epsilon2 was not associated with the onset of AD in Tehran's population. The OR for epsilon2 allele in AD subjects was 0.34 (p = 0.21). The genotype frequencies for epsilon3, epsilon4, and epsilon2 alleles in control subjects were 91.2, 6.1, and 2.7%, respectively. These values were similar to that reported for Turkish, Greece, Japanese, Spanish, and Moroccan populations, but they were significantly different from the reported values for the other ethnic populations. This observation emphasizes the importance of geographical location and ethnical background of the subjects in the study of APOE genotypes and their association with AD. PMID- 15664113 TI - Activity deprivation-dependent induction of the proapoptotic BH3-only protein Bim is independent of JNK/c-Jun activation during apoptosis in cerebellar granule neurons. AB - Bcl-2-interacting mediator of cell death (Bim), a proapoptotic BH3-only protein, plays a critical role in neuronal apoptosis. Cerebellar granule neurons (CGNs) depend on activity for their survival and undergo apoptosis when deprived of depolarizing concentration of KCl. While it has been proposed that the activation of c-Jun NH2-terminal protein kinase (JNK)/c-Jun pathway contributes to the upregulation of bim gene in neurons subjected to survival signaling withdrawal, here we show that neither inhibition of JNK activity nor expression of dominant negative c-Jun suppresses the expression of bim gene induced by activity deprivation in CGNs. We conclude that induction of bim gene is independent of the activation of JNK/c-Jun signaling pathway by activity deprivation during apoptosis of CGNs. PMID- 15664114 TI - Coexpression of the cannabinoid receptor type 1 with the corticotropin-releasing hormone receptor type 1 in distinct regions of the adult mouse forebrain. AB - The high abundance of the cannabinoid receptor type 1 (CB1) in the brain and the discovery of its endogenous ligands possessing neuromodulatory activities suggest an important potential of the endocannabinoid system to influence the functions of other receptor systems in the brain, including the corticotropin releasing hormone (CRH) system. Several studies evidenced a cross-talk between these two receptor systems. In trying to detail functional interactions between CB1 and the CRH receptor type 1 (CRHR1), we performed double-label-in situ hybridisation on mouse forebrain sections to localise the transcripts encoding the two receptors at a cellular level. Colocalisation of both receptor mRNAs was only detected in low CB1-expressing cells, which are mainly principal projecting neurons, whereas high CB1-expressing cells, which are considered to be mostly GABAergic did not contain mRNA encoding CRHR1. CB1 is differentially coexpressed with CRHR1 in olfactory regions, in several cortical and limbic structures, and in some hypothalamic and thalamic nuclei. These observations suggest a complex mechanism underlying the mutual interrelation and modulation of the two receptor systems. In particular, high levels of coexpressing cells in cortical and limbic areas may relate to cognitive functions, such as working memory, emotional and declarative learning. Colocalisation of CB1 and CRHR1 in hypothalamic regions strongly suggests functional interactions regarding the neuroendocrine homeostasis, including feeding behaviour. PMID- 15664115 TI - Facilitation of respiratory rhythm by a mu-opioid agonist in newborn rat pons medulla-spinal cord preparations. AB - We investigated the effect of a mu-opioid agonist, DAGO, on the respiratory frequency of pons-medulla-spinal cord preparations from newborn rats. Bath application of a low concentration of DAGO (0.2 microM) facilitated respiratory rhythm in pons-medulla-spinal cord preparations, whereas it induced respiratory depression in medulla-spinal cord preparations (without pons). At a higher concentration (1.0 microM), at which the inspiratory burst generation in the medulla was strongly depressed, the respiratory rhythm in half of the pons medulla-spinal cord preparations increased and then decreased, thus showing a biphasic response. In the other half of these preparations, only the facilitatory effect was observed. The burst rate of pre-inspiratory neurons in the rostral ventrolateral medulla was also facilitated by DAGO application. Such facilitation of the respiratory rhythm is probably due to disinhibition of a pontine inhibitory system. Our findings also suggest the existence of a pontine excitatory system, which is depressed by the pontine inhibitory system under control conditions. PMID- 15664116 TI - Illusory percepts of moving patterns due to discrete temporal sampling. AB - Continuously, moving objects under continuous illumination can be seen to move in a direction opposite to their actual motion. This illusory reversed motion can be explained as due to discrete temporal sampling of the moving stimulus by the visual system. If temporal sampling lies behind the illusory motion, then the probability of illusory motion should depend on the temporal frequency of the motion stimulus. By presenting contracting bull's-eye gratings of various spatial frequencies we were able to tease apart the drift speed and temporal frequency. The prevalence of illusory percepts depended on the temporal frequency, not the speed. The data suggest that the human visual system samples the incoming stimulation at a rate near 16 Hz. PMID- 15664117 TI - Fibroblast growth factor 2 mediated disruption of myelin-forming oligodendrocytes in vivo is associated with increased tau immunoreactivity. AB - We have previously shown that fibroblast growth factor 2 (FGF2) disrupts myelin formation by oligodendrocytes in vivo. Here, we have investigated the possibility that this is associated with changes in the expression of tau, a major microtubule-associated protein (MAP) involved in the production of myelin membranes by oligodendrocytes. FGF2, or saline vehicle in controls, was delivered into the brain ventricles of deeply anaesthetised young rats, and their actions were examined on the anterior medullary velum (AMV), a thin sheet of tissue that roofs part of the ventricular system underlying the cerebellum. The results show that the FGF2-induced loss of myelin is associated with increased immunostaining for tau within oligodendrocyte somata. Immunohistochemical and Western blot analyses demonstrate a 50% decrease in myelin-forming oligodendrocytes, axonal myelin sheaths, and levels of myelin-related proteins, with a correlative 100% increase in the level of tau. The results identify a potential mechanism by which FGF2-mediated accumulation of tau disrupts the transport of myelin-related gene products, resulting in disruption and eventual loss of oligodendrocytes and myelin, which are features of ischemia and a variety of demyelinating and neurodegenerative diseases. PMID- 15664118 TI - Feature uncertainty: a novel test to probe prefrontal dysfunction in unaffected siblings of schizophrenia patients. AB - Previous studies indicated that neuropsychological impairments are potential endophenotypes of schizophrenia. However, the sensitivity of these procedures is not sufficient and their brain substrates are poorly defined. The aim of this study was to measure the behavioral performance of siblings of schizophrenia patients and controls on a novel feature uncertainty (FU) task that selectively activates dorsal anterior cingulate cortex relative to orientation (OR) and spatial frequency (SF) discrimination. During the FU task, two subsequent sinusoidal gratings are presented and participants are asked to remember both the OR and SF of the gratings. After the disappearance of the gratings, a color cue signifies the perceptual dimension (OR or SF) to discriminate. Results revealed that the siblings of schizophrenia patients (n = 25) showed a selective deficit on the FU task as compared with controls (n = 20). The FU deficit was more severe than that found on neuropsychological tests of executive functions, psychomotor speed, and verbal memory. These results suggest that anterior cingulate dysfunction is a potential endophenotype of schizophrenia. PMID- 15664119 TI - Evaluation of cholinergic markers in Alzheimer's disease and in a model of cholinergic deficit. AB - Cognitive deficits in neuropsychiatric disorders, such as Alzheimer's disease (AD), have been closely related to cholinergic deficits. We have compared different markers of cholinergic function to assess the best biomarker of cognitive deficits associated to cholinergic hypoactivity. In post-mortem frontal cortex from AD patients, acetylcholine (ACh) levels, cholinacetyltransferase (ChAT) and acetylcholinesterase (AChE) activity were all reduced compared to controls. Both ChAT and AChE activity showed a significant correlation with cognitive deficits. In the frontal cortex of rats with a selective cholinergic lesion, all cholinergic parameters measured (ACh levels, ChAT and AChE activities, "in vitro" and "in vivo" basal ACh release) were significantly reduced. AChE activity was associated to ChAT activity, and even more, to "in vivo" and "in vitro" basal ACh release. Quantification of AChE activity is performed by an easy and cheap method and therefore, these results suggest that determination of AChE activity may be used as an effective first step method to evaluate cholinergic deficits. PMID- 15664120 TI - Changes of 5-HT receptor subtype mRNAs in rat dorsal root ganglion by bee venom induced inflammatory pain. AB - The reverse transcriptase polymerase chain reaction (RT-PCR) technique was used to examine the changes of the expression of 5-hydroxytryptamine (5-HT) receptors in the rat lumbar dorsal root ganglion (DRG) following subcutaneous bee venom (BV) injection into the plantar surface of the unilateral hindpaw. In the DRG ipsilateral to the BV injection, significant increase of mRNA levels of 5-HT(1A), 5-HT(1B), 5-HT(2A) and 5-HT(3) receptor subtypes were observed at 1 and 4h after the BV injection, while increase of 5-HT(2C), 5-HT(4), 5-HT(6) and 5-HT(7) receptor subtype mRNAs was detected at 4h only. No such changes were observed in the expressions of 5-HT(1D), 5-HT(1F) and 5-HT(5A) receptor subtype mRNAs. Upregulation of 5-HT(1A), 5-HT(1B) and 5-HT(2A) receptor subtype mRNAs was also observed in the contralateral DRG at 4 h. The presence of 5-HT(1E), 5-HT(2B) and 5-HT(5B) receptor subtype mRNAs was not detected in the rat DRG. The present results suggest that different sets of 5-HT receptor subtypes work at different stages of the inflammatory pain induced by subcutaneous BV injection. PMID- 15664121 TI - Cell culture isolation of a transmissible cytotoxicity from a human sample of cerebrospinal fluid. AB - We investigated a transmissible cytotoxicity isolated in VERO cell cultures from a sample of cerebrospinal fluid (CSF) drawn from a woman with ischemic brain injury. Amorphous aggregates formed by subunities of approximately 11 nm of diameter were detected in ultracentrifugates from partially purified cytotoxic cell preparations in the absence of virion-like particles which might justify the trasmissibility of this cytotoxic activity. Results of chemico-physical studies provided indications on the presence in the CSF of two protease-resistant acidic glycoproteins of about 39 and 27 kDa, respectively. The conformational change of a proteinic molecule may associate with particular properties such as tendency to aggregation, resistance to proteolysis, cytotoxicity. Considering that these same properties are shared by proteins present in the CSF sample under study, a hypothesis to pursue is that the CSF inoculum we isolated contained misfolded proteins formed in vivo following the ischemic injury of brain tissue. As far as the in vitro transmissibility of the cytotoxic activity, this could take place following the reproduction of the alterations of those proteins, independently of the original cause(s) which have fostered their formation in vivo. PMID- 15664122 TI - Multichannel NIRS analysis of brain activity during semantic differential rating of drawing stimuli containing different affective polarities. AB - We used 24-channel near-infrared spectroscopy (NIRS) to measure activity in the temporal, parietal, and frontal regions of the brain in eight Japanese women while the participants rated line drawings using semantic differential scales. Participants rated the seven line drawings on 15 bipolar semantic scales, each of which belonged to one of three semantic classes: Evaluation, Activity, or Potency. Suzuki et al. [M. Suzuki, J. Gyoba, Y. Sakuta, Multichannel near infrared spectroscopy analysis of brain activities during semantic differential rating of drawings, Tohoku Psychologica Folia 62 (2003) 86-98.] had reported previously that the right superior temporal gyrus and the right inferior parietal lobule are associated with Activity rating, while the brain regions around the central fissure were related to Potency rating. Based on these suggestions, we investigated the brain activity in these regions during rating of stimuli containing different affective polarities. When drawings were reported as 'static' or 'calm', oxyhemoglobin concentration was higher around the right superior temporal gyrus as compared to when they were considered 'noisy' or 'excitable'. Oxyhemoglobin concentrations around the central fissure were also higher when drawings were rated as 'soft', 'smooth', or 'blunt' compared to 'hard', 'rough', or 'sharp'. Any characteristic oxyhemoglobin changes were not found during the ratings on the evaluation scales. Our results suggest that activation patterns of the temporal and parietal regions are significantly modified by semantic polarities of Activity and Potency. PMID- 15664123 TI - Inhibitors of iNOS protects PC12 cells against the apoptosis induced by oxygen and glucose deprivation. AB - It has been shown that deletion of the gene encoding the inducible form of nitric oxide synthase (iNOS) results in a reduction of ischemia-induced apoptotic cell death, suggesting the detrimental role of iNOS. The signaling pathways by which iNOS mediates apoptotic cell death under ischemic conditions remain unclear. Understanding the molecular mechanisms of iNOS-mediated apoptotic cell death in ischemia may offer opportunities for potential therapeutic intervention. In the current study, undifferentiated rat pheochromocytoma PC12 cells, exposed to oxygen and glucose deprivation (OGD) followed by reperfusion (adding back oxygen and glucose, OGD-R), were used as an in vitro model of ischemia. The iNOS expression and activity were increased during OGD-R. OGD-R-induced apoptosis was demonstrated by the increase of LDH release, cytosolic release of cytochrome C and caspase-3 activity. Inhibition of iNOS activity by selective iNOS inhibitors, aminoguanidine and 1400W, reduces OGD-R-induced apoptotic cell death, as demonstrated by the decrease of LDH release, cytochrome C release, and caspase-3 activity. These results suggest the critical role of iNOS in mediating apoptosis under ischemic conditions, likely through the induction of caspase-3 activity. PMID- 15664124 TI - Visual discrimination performance is related to decreased alpha amplitude but increased phase locking. AB - This study investigated the question whether good and bad performance in a visual discrimination task is related to resting alpha power in a different way as it is known from memory tasks. The results show that good perceptual but not memory performance is related to low alpha amplitudes. In addition, we found that large phase resetting in the alpha band, and enhanced early components in the ERP are related to good performance in the discrimination task. The conclusion of this study is that in contrast to memory performance which is related to large resting alpha activity low alpha amplitudes are an indicator for good perceptual performance. PMID- 15664125 TI - Stability of metronidazole, tetracycline HCl and famotidine alone and in combination. AB - Metronidazole, tetracycline HCl and famotidine are commonly used for the treatment of Helicobacter pylori-associated peptic ulcer. In this paper, stabilities of these drugs and their combinations in solid and liquid states were studied as part of preformulation in the development of a combination drug delivery system. Solubility studies of metronidazole and tetracycline HCl were investigated, which indicated that both metronidazole and tetracycline HCl have high solubilities at and around pH 2.0. Metronidazole is relatively stable with little degradation in liquid phase. Tetracycline HCl in the dry state is stable when stored at room temperature regardless of exposure to light or humidity in the range of 20-65%. Enhanced temperature associated humidity effect was responsible for the instabilities of tetracycline HCl and famotidine to different extents. Elevated temperature accelerated the degradation of all the drugs in liquid phase but light exposure was not a factor for the degradation. The degradation processes of tetracycline HCl and famotidine were highly dependent on the pH of the solution, and relatively stable profiles were achieved at pH 4.0. No potential incompatibility between the drugs under storage conditions was observed in the development of a new multi-drug delivery tablet. PMID- 15664126 TI - Physicochemical and release properties of pellets coated with Kollicoat SR 30 D, a new aqueous polyvinyl acetate dispersion for extended release. AB - Kollicoat SR 30 D is a new aqueous colloidal polyvinyl acetate dispersion used for extended release coatings. Kollicoat SR 30 D is stable against sedimentation, has a low viscosity (54 mPas) and a negative zeta potential of -23.2 mV because of the presence of the anionic surfactant, sodium dodecyl sulfate. Because of its low minimum film formation temperature (MFT = 18 degrees C), plasticizer addition and a thermal after-treatment (curing) of coated pellets was not required. Coated pellets showed no aging or curing effect. The rate of release could be easily adjusted by varying the coating level. A subcoating layer of the hydrophilic polymer, polyvinyl alcohol, between an ibuprofen-containing core and the Kollicoat SR coating prevented the diffusion of the lipophilic, low melting ibuprofen into the polymer coating during storage. The drug release from Kollicoat SR 30 D coated pellets was almost independent of the pH and ionic strength of release medium. PMID- 15664127 TI - Skin permeation of physostigmine from fatty acids-based formulations: evaluating the choice of solvent. AB - This study was conducted to gain an understanding of the enhancement mechanism of fatty acids in skin permeation of physostigmine (PHY) by using a series of fatty acids and two solvents of opposing lipophilicity (propylene glycol (PG) and mineral oil (MO)). Interaction between fatty acid and drug was proven using NMR and conductivity measurements that showed a dependence on type of solvent used. Permeation flux of physostigmine from mineral oil-based formulations to skin was increased as solubility of physostigmine in mineral oil was enhanced in the presence of fatty acids having a longer chain. Thus, the dominant role of fatty acids in mineral oil was to increase solubility of physostigmine in the formulations that increased the driving force for physostigmine permeation through skin. As for propylene glycol, enhancement caused by fatty acids was attributed to their ability to increase the lipophilicity of formulation and to disrupt the lipid bilayers within the stratum corneum (SC). In conclusion, fatty acids enhancement for drug permeation across the skin was found to be dependent on the solvent used. Among various formulations here, oleic acid in mineral oil yielded fast permeation of PHY with a short lag time, which may be a good vehicle for transdermal delivery of PHY. PMID- 15664128 TI - Functional characterization of adenosine transport across the BBB in mice. AB - We investigated transport characteristics of adenosine across the blood-brain barrier (BBB) in mice. Uptake clearance across the BBB was measured by using an in situ mouse brain perfusion technique and cultured mouse brain capillary endothelial cell line (MBEC4 cells). Nucleoside transporter was cloned by RT-PCR and expressed on Xenopus laevis oocyte. Both in situ and in vitro studies revealed that the adenosine uptake is concentration-dependent, Na(+)-independent and S-(p-nitrobenzyl)-6-thioinosine (NBMPR)-sensitive. The K(t) values of in situ and in vitro studies were 31.7 +/- 13.8 microM and 11.9 +/- 2.84 microM, respectively. A good correlation was found for the inhibitory effects of nucleoside analogs to adenosine uptake between in situ and in vitro studies. RT PCR revealed the expression of RNA of mouse equilibrative nucleoside transporter (mENT1) in mouse brain capillary and MBEC4 cells. In mENT1 expressed on X. laevis oocyte, K(t) value of adenosine transport was 6.9 +/- 2.7 microM (and comparable to those in situ and in vitro studies). In conclusion, we characterized the adenosine transport across the BBB in mice by using in situ brain perfusion technique and MBEC4 cells and found that these transports share common characteristics with mENT1-mediated transport. Transport of adenosine across the BBB in mice may be attributable to mENT1. PMID- 15664129 TI - Chitosan-alginate multilayer beads for controlled release of ampicillin. AB - The aim of this study is to develop multilayer beads with improved properties for controlled delivery of the antibiotic ampicillin. Ionotropic gelation was applied to prepare single and multilayer beads using various combinations of chitosan and Ca(2+) as cationic components and alginate and polyphosphate as anions. Beads prepared with higher concentrations of chitosan entrapped more ampicillin. During incubation in simulated gastric fluid, the beads swelled and started to float but did not show any sign of erosion. Single layer chitosan-alginate beads released 70% of the drug within 4 h. Multilayer beads released only 20-30% in the same period of time. During subsequent incubation in simulated intestinal fluid, both single and multilayer beads continued to release drug. At least part of this release is due to disintegration of the beads. The rate of release both in gastric and intestinal fluid and the kinetics of disintegration in intestinal fluid can be controlled by changing the chitosan concentration in the coagulation fluid. The release of the drug can also be controlled by the degree of cross linking using polyphosphate. Cross-linked multilayer beads were prepared that released only 40% of the entrapped drug during 24 h. It is concluded that chitosan-alginate multilayer beads, cross-linked with polyphosphate offer an opportunity for controlled gastrointestinal passage of compounds with low molecular weight like ampicillin. PMID- 15664130 TI - Biodisposition of PEG-coated lipid microspheres of indomethacin in arthritic rats. AB - Conventional lipid microspheres (LM) were prepared using soybean oil and lipid at a 5.5:1 weight ratio with lipid phase consisting of PC (phosphatidyl choline):CH (cholesterol) (1:0.5) by molar ratio. The average diameter of the particles was 150 nm. Long-circulating microspheres (S-LM) were also prepared similarly but the lipid phase consisted of PC:CH:DSPE-PEG (phosphatidyl choline:cholesterol:distearoyl phosphatidyl ethanolamine-polyethylene glycol) 1:0.5:0.16 by molar ratio. A comparative biodistribution study was conducted between free indomethacin and lipo-indomethacin (LM and S-LM) in the arthritic rats by administering the formulations at a dose equivalent to 12 mg of indomethacin/kg. It was observed that the free drug as well as the encapsulated drug followed biphasic clearance from the blood. Pharmacokinetic parameters, such as AUC(0-t), terminal half-life, MRT increased significantly when the drug was used in encapsulated form (p < 0.05). Clearance of the drug was reduced 1.4 times with the conventional lipid microspheres and was reduced three-fold when encapsulated in polyethylene glycol-coated lipid microspheres. The overall drug targeting efficiency (T(e)) with the PEG-coated lipid microspheres was 7.5-fold higher than the conventional lipid microspheres. The high accumulation of the drug in arthritic paw with S-LM system may be accounted for by the reduced uptake by RES cells, and thereby, availability for extravascularization in the inflammatory tissues. PMID- 15664131 TI - Erythritol-based dry powder of glucagon for pulmonary administration. AB - Glucagon, a key regulatory element of glycogen metabolism, is known to be effective in the clinical treatment of hypoglycemia and the maintenance of normal circulating glucose levels in patients with total pancreatectomy, however the clinical use of this gut hormone has been restricted to parenteral administration. In this investigation, we prepared dry powder dosage forms of glucagon, which were formulated by mixing micronized glucagon particles and excipients with larger carrier particles. To achieve alveolar deposition for subsequent systemic absorption, a dry powder inhalant (DPI) of glucagon was size reduced to a mass median diameter between 1 and 6 microm, as measured by laser diffraction analysis. The use of erythritol as both excipient and carrier in DPI of glucagon resulted in high and reproducible flowability and dispersibility of the powder mixtures, and therefore it provided a low dosing of the active substances. Distinct transpulmonary absorption of glucagon was confirmed after intratracheal administration of the glucagon dry powder to anesthetized rats, as evidenced by the increase in the blood glucagon and blood sugar levels. These results suggested the usefulness of an erythritol-based powder form of glucagon for systemic administration. PMID- 15664132 TI - Stability of chitosan-pDNA complex powder prepared by supercritical carbon dioxide process. AB - The present study examined the stability of a gene in powders prepared with supercritical carbon dioxide (CO(2)) from the viewpoints of the ternary structure of DNA and in vivo transfection potential. An aqueous chitosan-pCMV-Luc complex solution containing mannitol was injected into the stream of a supercritical CO(2)/ethanol admixture to precipitate a gene powder. The obtained gene powders and gene solutions were placed in stability chambers at 25 or 40 degrees C for 4 weeks. The integrity and transfection potency of the gene were examined by electrophoresis and in vivo pulmonary transfection study in mice. The supercritical CO(2) process decreased the supercoiled DNA during the manufacturing process; however, the decrease in the remaining supercoiled and open circular DNA in the powders during storage was much slower than that in solutions. In addition, the powders had higher transfection potency than the solutions containing the same amount of DNA. The effect of chitosan on the stability of DNA in solutions was not obvious in the solutions but it improved the stability of DNA in powders during manufacturing and storage. Thus, a gene powder with a cationic vector is a promising ready-to-use formulation for inhalation therapy of pulmonary diseases. PMID- 15664134 TI - Effects of sugar ester and hydroxypropyl methylcellulose on the physicochemical stability of amorphous cefditoren pivoxil in aqueous suspension. AB - The improvement in physicochemical stability of amorphous cefditoren pivoxil (CDTR-PI) in aqueous suspensions by addition of sugar ester (SE) and hydroxypropyl methylcellulose (HPMC) was explained by prolonging the induction period prior to crystallization and the reduction in crystal peak intensity. Furthermore, the stabilizing effect of these additives in a multiple additive system was greater than in a single additive system. To determine the mechanism, by which these additives stabilized the amorphous CDTR-PI, we evaluated the surface states of CDTR-PI in suspension by measuring Raman spectra and zeta potential. The change in Raman spectra demonstrated that SE and HPMC interacted with CDTR-PI at the same interaction sites on CDTR-PI. The zeta potential reflected the adsorption phenomena of the additives and indicated that both SE and HPMC adsorbed onto particles of CDTR-PI with no apparent competitive interaction and the response was complementary. It was considered, based on this study, that HPMC and SE would stabilize amorphous CDTR-PI by different mechanisms; HPMC would mainly inhibit crystal growth by small amount of adsorption and SE would inhibit both crystal growth and nucleation by large amount of adsorption. This was considered to result in the hybrid effect in the multiple additive system. PMID- 15664133 TI - Synergistic effect of EDTA and boric acid on corneal penetration of CS-088. AB - In order to investigate the effects of EDTA and boric acid (EDTA/boric acid) on the corneal penetration of CS-088, an ophthalmic agent, the apparent permeability coefficient of CS-088 in the presence of EDTA/boric acid across the isolated corneal membranes of rabbits was measured using an in vitro penetration chamber system. FITC-dextran (M.W. 4400) and an electrical method based on membrane resistance were used to provide a quantitative assessment of the enhancing effect of EDTA/boric acid. The corneal penetration of CS-088 was significantly enhanced in the presence of EDTA/boric acid by approximately 1.6-fold. The permeability enhancing effect of EDTA/boric acid was apparently synergistic and concentration dependent on both EDTA and boric acid. The penetration of FITC-dextran, a paracellular marker, and electrical resistance of corneal membranes were not affected in the presence of EDTA/boric acid. Furthermore, no enhancing effect of EDTA/boric acid was observed in de-epithelialized corneas, although de epithelialized corneas exhibited a markedly higher permeability of CS-088 that was 24-fold greater than that for intact corneas. In conclusion, EDTA/boric acid synergistically enhances the transcellular permeability of CS-088 in the outer layer but not in the inner layers of the corneal membrane. PMID- 15664135 TI - Preparation of prostaglandin E1-hydroxypropyl-beta-cyclodextrin complex and its nasal delivery in rats. AB - The potential use of hydroxypropyl-beta-cyclodextrin (HP-betaCD) in the solubilization and stabilization of prostaglandin E(1) (PGE(1)) was investigated. The solubility and chemical stability of PGE(1) were significantly improved upon complexation with HP-betaCD. The nasal delivery of PGE(1) from the complex formulation was also studied in Wistar rats and compared with intravenous administration. PGE(1) complex after nasal administration caused a rapid decrease of blood pressure and exhibited an obvious dose-efficacy relationship, showing results nearly similar to those obtained for intravenous route. The time to reach the peak effect (T(max)) was approximately 3-4 min. Except T(max), other pharmacodynamic parameter values such as the maximal percent of blood pressure decrease (E(max), %), the lasting time of effect (T(d)), and the area under the curve (AUC, blood pressure decrease % min) were increased with increasing the administered doses. The E(max), T(d), and in particular AUC values between doses were significantly different (P < 0.01), but T(max) between doses were not significantly different (P < 0.05). The AUC values per unit dose of PGE(1) for nasal administration, however, were smaller than those for intravenous route, probably due to the incomplete absorption of nasally administered PGE(1). Besides, the in vitro effect of the PGE(1) complex on nasal mucociliary movement was also investigated with a toad palate model. The PGE(1) complex formulation exerted only minor effect on nasal mucociliary movement. These results indicate that the PGE(1)-HP-betaCD complex formulation for nasal delivery is a very promising preparation with advantages such as rapid and effective absorption, good chemical stability, ease of administration, and minor nasal ciliotoxicity. PMID- 15664136 TI - XEDS-mapping for explaining release patterns from single pellets. AB - A common way to formulate controlled-release (CR) pharmaceuticals is to coat pellets of active substance with a polymer film, decrease the size of the pellets and distribute them as multiple-unit dosages in capsules. To increase the understanding of the release mechanism, the pellet shape and surface structure of pellets, before and after release in microtitre plates, have been studied by scanning electron microscope and X-ray energy-dispersive spectrometry. By performing these studies we associate release profiles during the first few hours to the microscopic structure. Pellets were divided into three classes (spherical pellets, dumbbell shaped pellets and twin-pellets) according to pellet form. Cases of burst release occurred for all three shape classes due to "open-window defects" at the surface. Areas of thinner polymer film in the neck-region of dumbbell shaped pellets broaden the range of intermediate release rates for this pellet shape. The surface of twin pellets and dumbbell shaped pellets showed more defects, which increases the release rates in comparison to spherical pellets. All pellets with high release rates revealed ruptures in the polymer film, whereas only small cracks could be traced for pellets with slow release rates. The information gained is necessary for the development of future formulations and mathematical modelling of release patterns. The pharmaceutical used as model was remoxipride coated with a polymer film of ethyl cellulose and 10 wt.% triethyl citrate. PMID- 15664137 TI - An investigation of the disintegration of tablets in biorelevant media. AB - The purpose of the study was to examine the disintegration of tablets in media designed to simulate conditions pertaining in the stomach. Although many studies have been performed to determine dissolution rates in these media, little work has been undertaken on the preliminary step in dissolution, namely disintegration. Two tablet formulations were prepared. One disintegrated rapidly (under 25 s in water) and the other more slowly (8 min in water). The disintegration times were measured by the BP 2000 test using discs. For the rapidly disintegrating tablets, disintegration times were similar in all media except for whole milk. This media is used to simulate the fed stomach and disintegration times were over five times longer than in the other media (P < 0.05). A similar effect was seen with the poorly disintegrating tablets in milk, and prolonged times were also observed in some of the other media. For these latter media, there was a good correlation between the penetration rate of the fluid into the tablet and the disintegration time. Penetration rates for milk were also slow which may be a reflection of its relatively high viscosity and low surface tension. PMID- 15664138 TI - The effect of the amount of binder liquid on the granulation mechanisms and structure of microcrystalline cellulose granules prepared by high shear granulation. AB - The structure of granules changes during the high shear granulation process. The purpose of this research was to investigate the effect of the amount of binder liquid on the structure of the granules and the structural changes which occur during the granulation process, using microcrystalline cellulose (MCC) and water as the model system. The structure is the result of the granulation mechanism; therefore, conclusions can be drawn about the latter by studying the former. X ray microtomography and scanning electron microscopy (SEM) were applied in order to visualise the densification process of granules, which were first freeze dried in order to preserve their structure. Variations in their porosity were quantified by applying image analysis to the tomography results. In order to link the granule mechanical properties to their structural differences, a micromanipulation technique was used to measure granule resistance to deformation. MCC granules granulated with 100% (w/w) water showed increased densification with time, as expected; detailed examination showed that densification is more pronounced in the core of the granule; whereas the outer part remained more porous. Increased densification reduces deformability, so that granules become more resistant to breakage. The lower deformability of the densified granules in the final stages of granulation might result in establishment of equilibrium between attrition and growth, without substantial gross breakage. On the other hand, when more water was used (125%, w/w), densification was hardly observed; the porosity of the granule core was still high even after prolonged granulation times. This may be explained by the fact that higher water content increases the ease of deformation of granules. This increased deformability led to significant granule breakage even during the final phases of the granulation process. Therefore, for these granules a final equilibrium between breakage and coalescence might be established. This also explains why more granules produced with 125% granulation liquid were composed of fragments of irregular shape. Our results establish the link between the granulation behaviour of MCC in the latter stages and the material structure of these granules, which is determined by their liquid content. The process conditions (amount of liquid) to be chosen depend largely on the final purpose for which the granular material is produced. PMID- 15664139 TI - PLGA nanoparticles containing praziquantel: effect of formulation variables on size distribution. AB - Praziquantel has been shown to be highly effective against all known species of Schistosoma infecting humans. Spherical nanoparticulate drug carriers made of poly(D,L-lactide-co-glycolide) acid with controlled size were designed. Praziquantel, a hydrophobic molecule, was entrapped into the nanoparticles with theoretical loading varying from 10 to 30% (w/w). This study investigates the effects of some process variables on the size distribution of nanoparticles prepared by emulsion-solvent evaporation method. The results show that sonication time, PLGA and drug amounts, PVA concentration, ratio between aqueous and organic phases, and the method of solvent evaporation have a significant influence on size distribution of the nanoparticles. PMID- 15664140 TI - The influence of excipients on the diffusion of ibuprofen and paracetamol in gastric mucus. AB - The aim of this study was to examine the diffusion of commonly administered analgesics, ibuprofen and paracetamol, through gastric mucus. As ibuprofen and paracetamol are often formulated with alkalising excipients, or are commonly co administered with antacids that have been demonstrated to alter their absorption, diffusion was also studied in the presence of a range of soluble and insoluble antacids or buffering agents. The effect of pH, which has been demonstrated to modify the properties of mucus, was also studied. Mucus was a significant barrier to diffusion for both drugs, compared to an unstirred aqueous layer with diffusion rates significantly lower in the presence of a mucus barrier for both drugs; ibuprofen diffusion also demonstrated a significant increase in the lag time. Paracetamol diffusion was not significantly affected by addition of any antacid, whereas ibuprofen rates were affected and the diffusion lag time for ibuprofen was significantly reduced in all cases. Isolated increases in pH increased the rate and reduced the lag time for ibuprofen diffusion. It was shown that mucus acts as a passive barrier in the case of paracetamol diffusion, and an interactive barrier to ibuprofen diffusion. Changes in mucus viscosity at different pH values may be responsible for the observed changes in ibuprofen diffusion rate. PMID- 15664141 TI - Improved pharmacodynamics of timolol maleate from a mucoadhesive niosomal ophthalmic drug delivery system. AB - In the present study chitosan (REVTMbio1) or Carbopol (REVTMbio2 and 3) coated niosomal timolol maleate (0.25%) formulations were prepared by reverse phase evaporation (REV) and compared to timolol solution (TMS; 0.25%) in terms of in vitro release and IOP lowering pharmacodynamic effect. The in vitro release phase of timolol (91% release in 2 h) was extended significantly by its incorporation into niosomes and further by the polymer coating (40-43% release upto 10 h). The developed formulations were evaluated for their pharmacodynamics in albino rabbits, by measuring intraocular pressure (IOP) using a non-contact pneumatonometer, and were compared to a marketed in situ gel forming solution of timolol (Timolet GFS, 0.5%; Sun Pharma). REVTMbio1 formulation showed a more sustained effect of upto 8h (vis a vis 6 h for carbopol-coated niosomes). TMS in comparison showed effect for only 2 h though the peak effect was slightly more (14%). Lowering of IOP in the contralateral eye (20-40% as compared to 100% in case of TMS), considerably reduces with REV and REVbio formulations indicating lesser systemic side effects. Moreover, the results of REVTMbio1formulation containing 0.25% of timolol maleate compared well with the 0.5% marketed gel formulation, indicating our formulation to be significantly better considering that similar effect is obtained at half the concentration. The later becomes especially important in context to the cardiovascular side effects associated with ocular timolol maleate therapy. PMID- 15664142 TI - Loading efficiency of stavudine on polybutylcyanoacrylate and methylmethacrylate sulfopropylmethacrylate copolymer nanoparticles. AB - Loading efficiency (LE) of stavudine (D4T), a human immunodeficiency antiretroviral agent, on the external surfaces of polybutylcyanoacrylate (PBCA) and methylmethacrylate-sulfopropylmethacrylate (MMA-SPM) was investigated. The experimental results indicate that the larger the polymeric nanoparticles (NPs), the smaller LE of D4T on the two kinds of biomaterials. Freeze drying of the two NPs, however, yields an increase in particle size and an increase in LE of D4T, in general. Preservation of the two D4T-loaded NPs through cold storage at 4 degrees C over 6 weeks leads to an increase in particle size and a decrease in LE of D4T. LE of D4T on both of the two NPs decreases with a variation in pH value from pH 7.2 of loading medium. LE of D4T on MMA-SPM NPs is larger than that on PBCA NPs at pH 7.4; and for the case of variation in pH value of loading medium from pH 7.2, the extent of decrease in LE of D4T for MMA-SPM NPs is higher than that on PBCA NPs. These outcomes imply that for oral administration, D4T-loaded MMA-SPM NPs may be more advantageous than D4T-loaded PBCA NPs, and D4T-loaded PBCA NPs may be more favorable than D4T-loaded MMA-SPM NPs for intravenous injection. PMID- 15664143 TI - Inhomogeneities in muscle activation reveal motor unit recruitment. AB - This paper presents a novel method to quantify spatial changes in muscle activation pattern by multi-channel surface electromyography (MCSEMG) in order to investigate motor unit recruitment variation. The method is based on non-uniform distributions of motor units that cause spatial inhomogeneous muscle activation. To evaluate the method, 15 subjects performed three isometric elbow flexion contractions consisting of slow sinusoidal changes in force ranging from 0% to 80% of the maximal voluntary contraction. MCSEMG electrodes were placed in a 10 x 13 grid over the biceps brachii muscle. From all channels, root mean square (RMS) values of bipolar leadings were computed over 0.5 s epochs over the whole recording. Thereafter, correlation coefficients were calculated between the RMS values at one epoch, with the RMS values at another epoch. Results showed consistent spatial changes in the distribution of RMS at different contraction levels up to 80% of maximal voluntary contraction and when comparing increasing and decreasing contractions at the same force level. These findings are reproducible within and between subjects, and in agreement with physiological phenomena and therefore indicate that the spatial inhomogeneities of motor unit properties in the biceps brachii muscle can be used to study changes in motor unit recruitment. PMID- 15664144 TI - Voluntary low-force contraction elicits prolonged low-frequency fatigue and changes in surface electromyography and mechanomyography. AB - Controversies exist regarding objective documentation of fatigue development with low-force contractions. We hypothesized that non-exhaustive, low-force muscle contraction may induce prolonged low-frequency fatigue (LFF) that in the subsequent recovery period is detectable by electromyography (EMG) and in particular mechanomyography (MMG) during low-force rather than high-force test contractions. Seven subjects performed static wrist extension at 10% maximal voluntary contraction (MVC) for 10 min (10%MVC10 min). Wrist force response to electrical stimulation of extensor carpi radialis muscle (ECR) quantified LFF. EMG and MMG were recorded from ECR during static test contractions at 5% and 80% MVC. Electrical stimulation, MVC, and test contractions were performed before 10%MVC10 min and at 10, 30, 90 and 150 min recovery. In spite of no changes in MVC, LFF persisted up to 150 min recovery but did not develop in a control experiment omitting 10%MVC10 min. In 5% MVC tests significant increase was found in time domain of EMG from 0.067+/-0.028 mV before 10%MVC10 min to 0.107+/-0.049 and 0.087+/-0.05 mV at 10 and 30 min recovery, respectively, and of the MMG from 0.054+/-0.039 ms(-2) to 0.133+/-0.104 and 0.127+/-0.099 ms(-2), respectively. No consistent changes were found in 80% MVC tests. In conclusion, non-exhaustive low force muscle contraction resulted in prolonged LFF that in part was identified by the EMG and MMG signals. PMID- 15664145 TI - Simulation analysis of the performance of a novel high sensitive spectral index for quantifying M-wave changes during fatigue. AB - A high sensitive fatigue index is desired to improve stimulation strategy and to prevent muscle damage in functional electrical simulations. The great number of indexes used shows that there is no index that satisfies all investigators. A way to develop a high sensitive index for quantifying M-wave changes during fatigue and to estimate its performance was analyzed. The changes in M-wave and its frequency distribution due to variations of intracellular action potential (IAP) and muscle fibre propagation velocity (MFPV) with fatigue were simulated. It was found that the ratio between the spectral moments of order -1 and 2 was considerably more sensitive to peripheral muscle fatigue than the mean (the ratio between the spectral moments of order 1 and 0) and median frequency traditionally used. The sensitivity of the new index depended on the electrode arrangement and position in respect to the active fibres. The belly-tendon detection promised the highest index sensitivity. The length of the active fibres also affected the index sensitivity. The shorter the fibres the lower was the index sensitivity. The sensitivity of the new index could be relatively high even in the case of traditionally used high-pass cut-off frequencies that could distort the M-wave shape. PMID- 15664146 TI - Intracellular action potential generation and extinction strongly affect the sensitivity of M-wave characteristic frequencies to changes in the peripheral parameters with muscle fatigue. AB - Changes in muscle fibre propagation velocity (MFPV) and shape of intracellular action potentials (IAPs) accompany peripheral muscle fatigue. We have shown through mathematical simulations that the effects of IAP generation and extinction reduced the sensitivity of the mean (fmean) and median (fmed) frequency of M-wave power spectra to individual changes in MFPV. Due to the differences in weighting of the spectral components used for calculation of the characteristic frequencies, the highest spectral components of the M-wave affected the fmean more than the fmed. These components are related to the M-wave leading edge that reflects the IAP depolarization phase. They reduced the sensitivity of the spectral moment of order 1 to individual changes in MFPV and increased its sensitivity to IAP changes. Since the changes of the IAP depolarization phase during the final stages of peripheral muscle fatigue affected the fmean more, the range of the relative reductions of the fmean and fmed were approximately the same under combined changes in IAP and MFPV. The sensitivities of M-wave characteristic frequencies depended also on the electrode arrangement and position as well as on the length of active muscle fibres. PMID- 15664147 TI - The effects of gender on quadriceps muscle activation strategies during a maneuver that mimics a high ACL injury risk position. AB - While the increased incidence of serious knee injuries in female athletes is well established, the underlying neuromuscular mechanisms related to the elevated ACL injury rate has yet to be delineated. Video analysis of ACL injury during competitive sports play indicates a common body position associated with non contact ACL injury; the tibia is externally rotated, the knee is close to full extension, the foot is planted and as the limb is decelerated it collapses into valgus. The purpose of the current prospective study was to evaluate gender differences in quadriceps muscle activation strategies when performing a physically challenging, but reproducible maneuver that mimics the high ACL injury risk position (in the absence of high velocity and high loads). Twenty physically active college-aged subjects (10 male and 10 female) performed multiple sets of the prescribed exercise. EMG recordings were employed to measure the ratio of activation between the medial and lateral quadriceps during the 4, 8, 12, 16, and 20th sets of exercise. Females demonstrated decreased RMS medial-to-lateral quadriceps ratios compared to males (F(1,18)=5.88, p=0.026). There was no main effect of set number on RMS quadriceps ratio (p>0.05). The results of this study suggest that females utilize neuromuscular activation strategies which may contribute to "dynamic valgus" and ACL rupture when performing high-risk maneuvers. PMID- 15664148 TI - Comparison of Fourier and wavelet transform procedures for examining the mechanomyographic and electromyographic frequency domain responses during fatiguing isokinetic muscle actions of the biceps brachii. AB - The primary purpose of the present study was to compare the fast Fourier transform (FFT) with the discrete wavelet transform (DWT) for determining the mechanomyographic (MMG) and electromyographic (EMG) center frequency [mean power frequency (mpf), median frequency (mdf), or wavelet center frequency (cf)] patterns during fatiguing isokinetic muscle actions of the biceps brachii. Seven men (mean+/-SD age=23+/-3 years) volunteered to perform 50 consecutive maximal, concentric isokinetic muscle actions of the dominant forearm flexors at a velocity of 180 degrees s(-1). Non-parametric "run" tests indicated significant (p<0.05) trends in the MMG and EMG signals for the 5th, 25th, and 45th muscle actions for all subjects, thereby confirming non-stationarity of the MMG and EMG signals. There were significant (p<0.05) correlations among the average normalized mpf, mdf, and cf values for contractions 1-50 for both MMG (r=0.671 0.935) and EMG (r=0.956-0.987). Polynomial regression analyses demonstrated quadratic decreases in normalized MMG mpf (R2=0.439), MMG mdf (R2=0.258), MMG cf (R2=0.359), EMG mpf (R2=0.952), EMG mdf (R2=0.914) and EMG cf (R2=0.888) across repetitions. The primary finding of this study was the similarity in the mpf, mdf, and cf patterns for both MMG and EMG, which suggested that, despite the concerns over non-stationarity, Fourier based methods are acceptable for determining the patterns for normalized MMG and EMG center frequency during fatiguing dynamic muscle actions at moderate velocities. PMID- 15664149 TI - The effect of sampling frequency on EMG measures of occupational mechanical exposure. AB - The purpose of the present study was to determine how much it is possible to downsample EMG without loosing the accuracy of some EMG measures of occupational mechanical exposure frequently used in the literature. The EMG signals of four muscles (two trapezius, right deltoid and right extensor digitorum) were collected (sampling frequency: 1024 Hz) from 20 subjects while performing a 20 min computer work task. The EMG RMS amplitude was computed from 0.125 s successive time-windows for the original (1024 Hz) and four additional digitally resampled (at 512, 256, 128 and 64 Hz) EMG signals. Three of the most frequently used data reduction methods (1. gaps analysis, 2. amplitude probability distribution function and 3. exposure variation analysis) were used to summarise the 20 min EMG activation profiles. The results from the exposure variation analysis were summarized into three variables using a new method detailed here. In general, the ANOVA for repeated measures demonstrated that a decrease of the sampling frequency significantly changed the EMG measures (relative to the 1024 Hz sampling condition) most of the time at 64 and 128 Hz, occasionally at 256 Hz, but practically never at 512 Hz. An analysis of the maximal errors (relative to the 1024 Hz condition) across all subjects supported these findings. Consequently, it was concluded that 512 Hz is quite conservative and should practically never lead to invalid EMG parameters estimations. Conversely, 256 Hz represents the lowest limit tolerable for some EMG parameters (gaps analysis, amplitude probability distribution function) while it is unacceptable for others (modified exposure variation analysis). PMID- 15664150 TI - An EMG fractal indicator having different sensitivities to changes in force and muscle fatigue during voluntary static muscle contractions. AB - During a sustained contraction, electromyographic signals (EMGs) undergo a spectral compression. This fatigue behaviour induces a shift of the mean and the median frequencies to lower frequencies. On the other hand, several studies conclude that the mean/median frequency can increase, decrease or remain constant with an increasing force level. Such inconsistency is embarrassing since the fatigue state may be influenced by the force level. In this paper, we propose a frequency indicator which is sensitive to the force level independently of the fatigue state evaluated at 70% of the maximal voluntary contraction. Ten healthy volunteers participated in the study and both surface EMGs (from the short head of the biceps brachii) and force signals were measured. This study compared force and fatigue effects on the EMGs during short (3-s) isometric contractions at different strength intensities and during a sustained isometric contraction until exhaustion. The EMGs partly show 1/falpha spectral behaviours since their power spectral densities may experimentally fit with two linear segments in a log-log representation. The measured "right" slope produces variations of force as 20 times the variations of fatigue. 1/falpha Behaviour may be related to stochastic fractals. This fractal indicator is a new frequency indicator that is thus complementary to other known classical frequency indicators when studying force during unknown fatigue states. PMID- 15664151 TI - Quantification the relationship between FITA scores and EMG skill indexes in archery. AB - Forearm electromyographic (EMG) data are assumed to be an effective method in estimating performance level in archery. The aim of the current study was to establish archery skill indexes based on EMG data. Elite (n=7, FITA score=1303.4+/-26.2), beginner (n=6, FITA score=1152+/-9.0) and non-archers (n=10, assumed FITA score=250+/-0), were involved in the study. EMG activity of Muscle flexor digitorum superficialis and Muscle extensor digitorum were quantified. Two-second periods--1 s before and 1 s after the fall of the clicker- were used to obtain averaged and rectified EMG data. The averaged and rectified EMG data were filtered by averaging finite impulse response filter with 40 ms time window and then normalized with respect to maximum voluntary contraction. To estimate FITA scores from EMG data, the following skill indexes that based on mean area under some parts of processed EMG waveforms was offered for archery. These were the pre-clicker archery skill index (PreCASI), post-clicker archery skill index (PostCASI), archery skill index (ASI) and post-clicker archery skill index 2 (PostCASI2). The correlations between rank of FITA scores and natural logarithms of archery skill indexes were significant for log(PreCASI): r=-0.66, p<0.0008; for log(PostCASI): r=-0.70, p<0.0003; for log(ASI): r=-0.74, p<0.0001; log(PostCASI2): r=-0.63, p<0.002. It is concluded that EMG skill indexes may be useful for: (a) assessing shooting techniques, (b) evaluation of archers' progress and (c) selection of talented archers. PMID- 15664153 TI - Conduit for privileged communications in the lymph node. AB - A study by Sixt et al. in this issue of Immunity identifies conduit-associated dendritic cells whose privileged access to antigen arriving by the conduit enables uptake and processing of antigen within 90 min of antigen inoculation, long before the arrival of dendritic cells from skin. PMID- 15664154 TI - Oxidative modification of leukocyte adhesion. AB - In this issue of Immunity, Marko Salmi and colleagues describe mice lacking AOC3, an endothelial cell monoaminooxidase that is involved in modulating leukocyte rolling, adhesion, and migration (Stolen et al., 2005). Their data demonstrate the importance of oxidative modification of (unknown) adhesion molecules in regulating inflammation and lymphocyte homing. PMID- 15664155 TI - Lyn tyrosine kinase: accentuating the positive and the negative. AB - Lyn, one of several Src-family tyrosine kinases in immune cells, is noted for its ability to negatively regulate signaling pathways through phosphorylation of inhibitory receptors, enzymes, and adaptors. Somewhat paradoxically, it is also a key mediator in several pathways of B cell activation, such as CD19 and CD180. Whether Lyn functions to promote or inhibit immune cell activation depends on the stimulus and the developmental state, meaning that the consequences of Lyn activity are context dependent. The importance of regulating Lyn activity is exemplified by the pathological conditions that develop in both lyn-/- and lyn gain-of-function mice (lynup/up), including lethal antibody-mediated autoimmune diseases and myeloid neoplasia. Here, we review the outcomes of altered Lyn activity within the framework of B cell development and differentiation and the circumstances that appear to dictate the outcome. PMID- 15664156 TI - The conduit system transports soluble antigens from the afferent lymph to resident dendritic cells in the T cell area of the lymph node. AB - Resident dendritic cells (DC) within the T cell area of the lymph node take up soluble antigens that enter via the afferent lymphatics before antigen carrying DC arrive from the periphery. The reticular network within the lymph node is a conduit system forming the infrastructure for the fast delivery of soluble substances from the afferent lymph to the lumen of high endothelial venules (HEVs). Using high-resolution light microscopy and 3D reconstruction, we show here that these conduits are unique basement membrane-like structures ensheathed by fibroblastic reticular cells with occasional resident DC embedded within this cell layer. Conduit-associated DC are capable of taking up and processing soluble antigens transported within the conduits, whereas immigrated mature DC occur remote from the reticular fibers. The conduit system is, therefore, not a closed compartment that shuttles substances through the lymph node but represents the morphological equivalent to the filtering function of the lymph node. PMID- 15664157 TI - Identification of FcalphaRI as an inhibitory receptor that controls inflammation: dual role of FcRgamma ITAM. AB - Serum IgA is considered a discrete housekeeper of the immune system with multiple anti-inflammatory functions, whereas IgA-immune complexes mediate inflammatory responses. Here, we identify FcalphaRI as a molecular device that determines the nature of IgA responses. In the absence of sustained aggregation, receptor targeting by serum IgA or anti-FcalphaRI Fab inhibits activating responses of heterologous FcgammaR or FcepsilonRI. The inhibitory mechanism involves recruitment of tyrosine phosphatase SHP-1 to FcalphaRI and impairment of Syk, LAT, and ERK phosphorylation induced by FcepsilonRI engagement. SHP-1 recruitment is dependent on ERK. Conversely, sustained aggregation of FcalphaRI by multimeric ligands stimulates cell activation by recruiting high amounts of Syk and aborting SHP-1 binding. Both types of signals require the FcRgamma-ITAM motif. Anti FcalphaRI Fab treatment suppresses manifestations of allergic asthma in FcalphaRI transgenic mice. These findings redefine FcalphaRI as a bifunctional inhibitory/activating receptor of the immune system that mediates both anti- and proinflammatory functions of IgA. PMID- 15664158 TI - CD34 and CD43 inhibit mast cell adhesion and are required for optimal mast cell reconstitution. AB - CD34 is a cell-surface sialomucin expressed by hematopoietic stem cells (HSC), mast cells, and vascular endothelia. Despite its popularity as an HSC marker, the function of CD34 on hematopoietic cells remains enigmatic. Here, we have addressed this issue by examining the behavior of mutant mast cells lacking CD34, the related sialomucin, CD43, or both molecules. Loss of these molecules leads to a gene-dose-dependent increase in mast cell homotypic aggregation with CD34/CD43KOs > CD43KO > CD34KO > wild-type. Importantly, reexpression of CD34 or CD43 in these cells caused reversal of this phenotype. Furthermore, we find that loss of these sialomucins prevents mast cell repopulation and hematopoietic precursor reconstitution in vivo. Our data provide clear-cut evidence for a hematopoietic function for CD34 and suggest that it acts as a negative regulator of cell adhesion. PMID- 15664159 TI - The KSHV immediate-early transcription factor RTA encodes ubiquitin E3 ligase activity that targets IRF7 for proteosome-mediated degradation. AB - Many viruses encode proteins that counteract the development of the interferon (IFN)-mediated antiviral state. Here, we report that interferon regulatory factor 7 (IRF7), a key mediator of type I IFN induction, is targeted for degradation by binding to the RTA immediate-early nuclear transcription factor encoded by Kaposi's sarcoma-associated herpesvirus (KSHV or HHV8). Cotransfection with RTA blocked IRF7-mediated IFNalpha and IFNbeta mRNA production and promoted the ubiquitination and degradation of IRF7 protein in a proteasome-dependent fashion. Addition of RTA also promoted polyubiquitination of IRF7 in an in vitro cell free assay, demonstrating that RTA itself acts as a ubiquitin E3 ligase. RTA also autoregulated its own polyubiquitination and stability, and both activities were abolished by point mutations in a Cys plus His-rich N-terminal domain. Therefore, manipulation of the stability and function of IRF7 by the KSHV RTA transcription factor provides an unexpected regulatory strategy for circumventing the innate immune defence system. PMID- 15664160 TI - Tumor recognition following Vgamma9Vdelta2 T cell receptor interactions with a surface F1-ATPase-related structure and apolipoprotein A-I. AB - Vgamma9Vdelta2 T lymphocytes, a major gammadelta T lymphocyte subset in humans, display cytolytic activity against various tumor cells upon recognition of yet uncharacterized structures. Here, we show that an entity related to the mitochondrial F1-ATPase is expressed on tumor cell surface and promotes tumor recognition by Vgamma9Vdelta2 T cells. When immobilized, purified F1-ATPase induces selective activation of this lymphocyte subset. The Vgamma9Vdelta2 T cell receptors (TCR) and the F1-ATPase also bind a delipidated form of apolipoprotein A-I (apo A-I), as demonstrated by surface plasmon resonance. Moreover, the presence of apo A-I in the culture medium is required for optimal activation of Vgamma9Vdelta2 T cells by tumors expressing F1-ATPase. This study thus describes an unanticipated tumor recognition mechanism by Vgamma9Vdelta2 lymphocytes and a possible link between gammadelta T cell immunity and lipid metabolism. PMID- 15664161 TI - Structure of an autoimmune T cell receptor complexed with class II peptide-MHC: insights into MHC bias and antigen specificity. AB - T cell receptor crossreactivity with different peptide ligands and biased recognition of MHC are coupled features of antigen recognition that are necessary for the T cell's diverse functional repertoire. In the crystal structure between an autoreactive, EAE T cell clone 172.10 and myelin basic protein (1-11) presented by class II MHC I-Au, recognition of the MHC is dominated by the Vbeta domain of the TCR, which interacts with the MHC alpha chain in a manner suggestive of a germline-encoded TCR/MHC "anchor point." Strikingly, there are few specific contacts between the TCR CDR3 loops and the MBP peptide. We also find that over 1,000,000 different peptides derived from combinatorial libraries can activate 172.10, yet the TCR strongly prefers the native MBP contact residues. We suggest that while TCR scanning of pMHC may be degenerate due to the TCR germline bias for MHC, recognition of structurally distinct agonist peptides is not indicative of TCR promiscuity, but rather highly specific alternative solutions to TCR engagement. PMID- 15664162 TI - Distinct and nonredundant in vivo functions of TNF produced by t cells and macrophages/neutrophils: protective and deleterious effects. AB - Tumor necrosis factor (TNF, TNFalpha) is implicated in various pathophysiological processes and can be either protective, as in host defense, or deleterious, as in autoimmunity or toxic shock. To uncover the in vivo functions of TNF produced by different cell types, we generated mice with TNF ablation targeted to various leukocyte subsets. Systemic TNF in response to lipopolysaccharide was produced mainly by macrophages and neutrophils. This source of TNF was indispensable for resistance to an intracellular pathogen, Listeria, whereas T-cell-derived TNF was important for protection against high bacterial load. Additionally, both T-cell derived TNF and macrophage-derived TNF had critical and nonredundant functions in the promotion of autoimmune hepatitis. Our data suggest that T-cell-specific TNF ablation may provide a therapeutic advantage over systemic blockade. PMID- 15664163 TI - Absence of the endothelial oxidase AOC3 leads to abnormal leukocyte traffic in vivo. AB - Leukocyte migration from the blood to tissues is a prerequisite for normal immune responses. We produced mice deficient in an endothelial cell-surface oxidase (amine oxidase, copper containing-3 [AOC3], also known as vascular adhesion protein-1 [VAP-1]) and found that this enzyme is needed for leukocyte extravasation in vivo. Real-time imaging shows that AOC3 mediates slow rolling, firm adhesion, and transmigration of leukocytes in vessels at inflammatory sites and lymphoid tissues. Absence of AOC3 results in reduced lymphocyte homing into lymphoid organs and in attenuated inflammatory response in peritonitis. These data alter the paradigm of leukocyte extravasation cascade by providing the first physiological proof for the concept that endothelial cell surface enzymes regulate the development of inflammatory reactions in vivo and suggest that this enzyme should be useful as an anti-inflammatory target. PMID- 15664164 TI - Cooperation of human tumor-reactive CD4+ and CD8+ T cells after redirection of their specificity by a high-affinity p53A2.1-specific TCR. AB - Efficient immune attack of malignant disease requires the concerted action of both CD8+ CTL and CD4+ Th cells. We used human leukocyte antigen (HLA)-A*0201 (A2.1) transgenic mice, in which the mouse CD8 molecule cannot efficiently interact with the alpha3 domain of A2.1, to generate a high-affinity, CD8 independent T cell receptor (TCR) specific for a commonly expressed, tumor associated cytotoxic T lymphocyte (CTL) epitope derived from the human p53 tumor suppressor protein. Retroviral expression of this CD8-independent, p53-specific TCR into human T cells imparted the CD8+ T lymphocytes with broad tumor-specific CTL activity and turned CD4+ T cells into potent tumor-reactive, p53A2.1-specific Th cells. Both T cell subsets were cooperative and interacted synergistically with dendritic cell intermediates and tumor targets. The intentional redirection of both CD4+ Th cells and CD8+ CTL by the same high-affinity, CD8-independent, tumor-specific TCR could provide the basis for novel broad-spectrum cancer immunotherapeutics. PMID- 15664165 TI - Interleukin-32: a cytokine and inducer of TNFalpha. AB - We describe the gene structure, regulation, signal transduction. and functions of a cytokine, interleukin (IL)-32. An IL-18 unresponsive cell was converted to a responsive cell by transfection of the IL-18 receptor beta chain, and IL-18 induced microarray revealed high expression of a cytokine-like gene. Although IL 32 does not share sequence homology with known cytokine families, IL-32 induces various cytokines, human TNFalpha, and IL-8 in THP-1 monocytic cells as well as mouse TNFalpha and MIP-2 in Raw macrophage cells. IL-32 activates typical cytokine signal pathways of nuclear factor-kappa B (NF-kappaB) and p38 mitogen activated protein kinase. IL-32 mRNA is highly expressed in immune tissue rather than other tissues. Human IL-32 exists as four splice variants, and IL-32 from other species were found as expressed sequence tag clones in the databank. Induced in human peripheral lymphocyte cells after mitogen stimulation, in human epithelial cells by IFNgamma, and in NK cells after exposure to the combination of IL-12 plus IL-18, IL-32 may play a role in inflammatory/autoimmune diseases. PMID- 15664166 TI - Wake up and smell the pheromones. AB - Odorant binding proteins (OBPs) are abundant proteins of unknown function expressed at high levels in insect and vertebrate chemosensory organs. In this issue of Neuron, Xu et al. show that Drosophila OBP76a is necessary for fruit flies to respond to the aggregation pheromone 11-cis vaccenyl acetate. The results suggest a mechanism by which this OBP is intimately involved in pheromone signal transduction. PMID- 15664167 TI - Toward establishing a therapeutic window for rTMS by theta burst stimulation. AB - In this issue of Neuron, Huang et al. show that a version of the classic theta burst stimulation protocol used to induce LTP/LTD in brain slices can be adapted to a transcranial magnetic stimulation (TMS) protocol to rapidly produce long lasting (up to an hour), reversible effects on motor cortex physiology and behavior. These results may have important implications for the development of clinical applications of rTMS in the treatment of depression, epilepsy, Parkinson's, and other diseases. PMID- 15664168 TI - Progress in realizing the promise of microarrays in systems neurobiology. AB - The power of microarrays in neuroscience has been challenged by the cellular heterogeneity and complexity of the central nervous system. In this issue of Neuron, Arlotta, Molyneaux, and colleagues have developed a technique combining retrograde labeling, flow cytometry, and microarrays to purify and molecularly characterize a specific population of neurons from the brain, focusing here on cortical projection neurons. We discuss these findings and the implications of this development for both systems and molecular neuroscience. PMID- 15664169 TI - The molecular machinery of resurgent sodium current revealed. AB - Some TTX-sensitive sodium channels open transiently during recovery from inactivation, generating a "resurgent" sodium current that flows immediately following action potentials. In this issue of Neuron, Grieco and colleagues present evidence that resurgent sodium current results from a novel form of inactivation in which the cytoplasmic tail of the beta4 subunit acts as a classic open-channel blocker. PMID- 15664170 TI - Patterning molecules; multitasking in the nervous system. AB - Classical patterning molecules that were previously implicated in controlling cell fate choices in the nervous system have recently been shown to play additional roles in axon guidance and synaptogenesis. Bone morphogenetic proteins (BMPs), Sonic hedgehog (Shh), Wnts, and fibroblast growth factors (FGFs) all participate in multiple acts of controlling neuronal circuit assembly. Depending on the cellular context, they can provide instructive signals at the growth cone or synapse or alternatively can elicit responses in the nucleus initiating transcriptional changes. Differences in signal transduction pathways may contribute to the diversity of the functional repertoire of these versatile molecules. PMID- 15664171 TI - Drosophila OBP LUSH is required for activity of pheromone-sensitive neurons. AB - Odorant binding proteins (OBPs) are extracellular proteins localized to the chemosensory systems of most terrestrial species. OBPs are expressed by nonneuronal cells and secreted into the fluid bathing olfactory neuron dendrites. Several members have been shown to interact directly with odorants, but the significance of this is not clear. We show that the Drosophila OBP lush is completely devoid of evoked activity to the pheromone 11-cis vaccenyl acetate (VA), revealing that this binding protein is absolutely required for activation of pheromone-sensitive chemosensory neurons. lush mutants are also defective for pheromone-evoked behavior. Importantly, we identify a genetic interaction between lush and spontaneous activity in VA-sensitive neurons in the absence of pheromone. The defects in spontaneous activity and VA sensitivity are reversed by germline transformation with a lush transgene or by introducing recombinant LUSH protein into mutant sensilla. These studies directly link pheromone-induced behavior with OBP-dependent activation of a subset of olfactory neurons. PMID- 15664172 TI - Theta burst stimulation of the human motor cortex. AB - It has been 30 years since the discovery that repeated electrical stimulation of neural pathways can lead to long-term potentiation in hippocampal slices. With its relevance to processes such as learning and memory, the technique has produced a vast literature on mechanisms of synaptic plasticity in animal models. To date, the most promising method for transferring these methods to humans is repetitive transcranial magnetic stimulation (rTMS), a noninvasive method of stimulating neural pathways in the brain of conscious subjects through the intact scalp. However, effects on synaptic plasticity reported are often weak, highly variable between individuals, and rarely last longer than 30 min. Here we describe a very rapid method of conditioning the human motor cortex using rTMS that produces a controllable, consistent, long-lasting, and powerful effect on motor cortex physiology and behavior after an application period of only 20-190 s. PMID- 15664174 TI - Coupled tertiary folding and oligomerization of the T1 domain of Kv channels. AB - Acquisition of secondary, tertiary, and quaternary structure is critical to the fabrication, assembly, and function of ion channels, yet the relationship between these biogenic events remains unclear. We now address this issue in voltage-gated K(+) channels (Kv) for the T1 domain, an N-terminal Kv recognition domain that is responsible for subfamily-specific, efficient assembly of Kv subunits. This domain forms a 4-fold symmetric tetramer. We have identified residues along the axial T1-T1 interface that are critical for tertiary and quaternary structure, shown that mutations at one end of the axial T1 interface can perturb the crosslinking of an intersubunit cysteine pair at the other end, and demonstrated that tertiary folding and tetramerization of this Kv domain are coupled. A threshold level of tertiary folding is required for monomers to oligomerize. Coupling between tertiary and quaternary structure formation may be a common feature in the biogenesis of multimeric proteins. PMID- 15664173 TI - Neuronal subtype-specific genes that control corticospinal motor neuron development in vivo. AB - Within the vertebrate nervous system, the presence of many different lineages of neurons and glia complicates the molecular characterization of single neuronal populations. In order to elucidate molecular mechanisms underlying the specification and development of corticospinal motor neurons (CSMN), we purified CSMN at distinct stages of development in vivo and compared their gene expression to two other pure populations of cortical projection neurons: callosal projection neurons and corticotectal projection neurons. We found genes that are potentially instructive for CSMN development, as well as genes that are excluded from CSMN and are restricted to other populations of neurons, even within the same cortical layer. Loss-of-function experiments in null mutant mice for Ctip2 (also known as Bcl11b), one of the newly characterized genes, demonstrate that it plays a critical role in the development of CSMN axonal projections to the spinal cord in vivo, confirming that we identified central genetic determinants of the CSMN population. PMID- 15664175 TI - Open-channel block by the cytoplasmic tail of sodium channel beta4 as a mechanism for resurgent sodium current. AB - Voltage-gated sodium channels with "resurgent" kinetics are specialized for high frequency firing. The alpha subunits interact with a blocking protein that binds open channels upon depolarization and unbinds upon repolarization, producing resurgent sodium current. By limiting classical inactivation, the cycle of block and unblock shortens refractory periods. To characterize the blocker in Purkinje neurons, we briefly exposed inside-out patches to substrate-specific proteases. Trypsin and chymotrypsin each removed resurgent current, consistent with established roles for positively charged and hydrophobic/aromatic groups in blocking sodium channels. In Purkinje cells, the only known sodium channel associated subunit that has a cytoplasmic sequence with several positive charges and clustered hydrophobic/aromatic residues is beta4 (KKLITFILKKTREK; beta4(154 167)). After enzymatic removal of block, beta4(154-167) fully reconstituted resurgent current, whereas scrambled or point-mutated peptides were ineffective. In CA3 pyramidal neurons, which lack beta4 and endogenous block, beta4(154-167) generated resurgent current. Thus, beta4 may be the endogenous open-channel blocker responsible for resurgent kinetics. PMID- 15664176 TI - Sorting and activity-dependent secretion of BDNF require interaction of a specific motif with the sorting receptor carboxypeptidase e. AB - Activity-dependent secretion of BDNF is important in mediating synaptic plasticity, but how it is achieved is unclear. Here we uncover a sorting motif receptor-mediated mechanism for regulated secretion of BDNF. X-ray crystal structure analysis revealed a putative sorting motif, I(16)E(18)I(105)D(106), in BDNF, which when mutated at the acidic residues resulted in missorting of proBDNF to the constitutive pathway in AtT-20 cells. A V20E mutation to complete a similar motif in NGF redirected a significant proportion of it from the constitutive to the regulated pathway. Modeling and binding studies showed interaction of the acidic residues in the BDNF motif with two basic residues in the sorting receptor, carboxypeptidase E (CPE). (35)S labeling experiments demonstrated that activity-dependent secretion of BDNF from cortical neurons was obliterated in CPE knockout mice. Thus, we have identified a mechanism whereby a specific motif I(16)E(18)I(105)D(106) interacts with CPE to sort proBDNF into regulated pathway vesicles for activity-dependent secretion. PMID- 15664177 TI - Phospholipase Cbeta serves as a coincidence detector through its Ca2+ dependency for triggering retrograde endocannabinoid signal. AB - Endocannabinoids mediate retrograde signal and modulate transmission efficacy at various central synapses. Although endocannabinoid release is induced by either depolarization or activation of G(q/11)-coupled receptors, it is markedly enhanced by the coincidence of depolarization and receptor activation. Here we report that this coincidence is detected by phospholipase Cbeta1 (PLCbeta1) in hippocampal neurons. By measuring cannabinoid-sensitive synaptic currents, we found that the receptor-driven endocannabinoid release was dependent on physiological levels of intracellular Ca(2+) concentration ([Ca(2+)](i)), and markedly enhanced by depolarization-induced [Ca(2+)](i) elevation. Furthermore, we measured PLC activity in intact neurons by using exogenous TRPC6 channel as a biosensor for the PLC product diacylglycerol and found that the receptor-driven PLC activation exhibited similar [Ca(2+)](i) dependence to that of endocannabinoid release. Neither endocannabinoid release nor PLC activation was induced by receptor activation in PLCbeta1 knockout mice. We therefore conclude that PLCbeta1 serves as a coincidence detector through its Ca(2+) dependency for endocannabinoid release in hippocampal neurons. PMID- 15664178 TI - Bidirectional synaptic plasticity regulated by phosphorylation of stargazin-like TARPs. AB - Synaptic plasticity involves protein phosphorylation cascades that alter the density of AMPA-type glutamate receptors at excitatory synapses; however, the crucial phosphorylated substrates remain uncertain. Here, we show that the AMPA receptor-associated protein stargazin is quantitatively phosphorylated and that stargazin phosphorylation promotes synaptic trafficking of AMPA receptors. Synaptic NMDA receptor activity can induce both stargazin phosphorylation, via activation of CaMKII and PKC, and stargazin dephosphorylation, by activation of PP1 downstream of PP2B. At hippocampal synapses, long-term potentiation and long term depression require stargazin phosphorylation and dephosphorylation, respectively. These results establish stargazin as a critical substrate in the bidirectional control of synaptic strength, which is thought to underlie aspects of learning and memory. PMID- 15664179 TI - Transient and persistent dendritic spines in the neocortex in vivo. AB - Dendritic spines were imaged over days to months in the apical tufts of neocortical pyramidal neurons (layers 5 and 2/3) in vivo. A fraction of thin spines appeared and disappeared over a few days, while most thick spines persisted for months. In the somatosensory cortex, from postnatal day (PND) 16 to PND 25 spine retractions exceeded additions, resulting in a net loss of spines. The fraction of persistent spines (lifetime > or = 8 days) grew gradually during development and into adulthood (PND 16-25, 35%; PND 35-80, 54%; PND 80-120, 66%; PND 175-225, 73%), providing evidence that synaptic circuits continue to stabilize even in the adult brain, long after the closure of known critical periods. In 6-month-old mice, spines turn over more slowly in visual compared to somatosensory cortex, possibly reflecting differences in the capacity for experience-dependent plasticity in these brain regions. PMID- 15664180 TI - Circadian pacemaker neurons transmit and modulate visual information to control a rapid behavioral response. AB - Circadian pacemaker neurons contain a molecular clock that oscillates with a period of approximately 24 hr, controlling circadian rhythms of behavior. Pacemaker neurons respond to visual system inputs for clock resetting, but, unlike other neurons, have not been reported to transmit rapid signals to their targets. Here we show that pacemaker neurons are required to mediate a rapid behavior. The Drosophila larval visual system, Bolwig's organ (BO), projects to larval pacemaker neurons to entrain their clock. BO also mediates larval photophobic behavior. We found that ablation or electrical silencing of larval pacemaker neurons abolished light avoidance. Thus, circadian pacemaker neurons receive input from BO not only to reset the clock but also to transmit rapid photophobic signals. Furthermore, as clock gene mutations also affect photophobicity, the pacemaker neurons modulate the sensitivity of larvae to light, generating a circadian rhythm in visual sensitivity. PMID- 15664181 TI - Spatial memory in the rat requires the dorsolateral band of the entorhinal cortex. AB - The extensive connections of the entorhinal cortex with the hippocampus and the neocortex point to this region as a major interface in the hippocampal neocortical interactions underlying memory. We asked whether hippocampal dependent recall of spatial memory depends on the entorhinal cortex, and, if so, which parts are critical. After training in a Morris water maze, rats received fiber-sparing lesions in the dorsolateral band of the entorhinal cortex, which mediates much of the visuospatial input to the dorsal hippocampus. These lesions entirely disrupted retention and retarded new learning. Spatial memory was spared by lesions in the ventromedial band, which connects primarily with ventral hippocampus, but these lesions reduced defensive behavior on an elevated plus maze, mirroring the effects of damage to ventral hippocampus. The results suggest that the functional differences between dorsal and ventral hippocampus reflect their connectivity with modules of the entorhinal cortex that are differently linked to the rest of the cortex. PMID- 15664182 TI - Expansion of direction space around the cardinal axes revealed by smooth pursuit eye movements. AB - It is well established that perceptual direction discrimination shows an oblique effect; thresholds are higher for motion along diagonal directions than for motion along cardinal directions. Here, we compare simultaneous direction judgments and pursuit responses for the same motion stimuli and find that both pursuit and perceptual thresholds show similar anisotropies. The pursuit oblique effect is robust under a wide range of experimental manipulations, being largely resistant to changes in trajectory (radial versus tangential motion), speed (10 versus 25 deg/s), directional uncertainty (blocked versus randomly interleaved), and cognitive state (tracking alone versus concurrent tracking and perceptual tasks). Our data show that the pursuit oblique effect is caused by an effective expansion of direction space surrounding the cardinal directions and the requisite compression of space for other directions. This expansion suggests that the directions around the cardinal directions are in some way overrepresented in the visual cortical pathways that drive both smooth pursuit and perception. PMID- 15664183 TI - MAPKAP kinase-2: three's company at the G(2) checkpoint. AB - Elegant studies in fission yeast by and in mammalian cells by offer new insights into the mechanism through which stress-induced p38 activation inhibits mitotic entry in eukaryotic cells. PMID- 15664184 TI - Raf phosphorylation: one step forward and two steps back. AB - We understand Raf-1 activation relatively well but know less about how it is inactivated. An exciting study in this issue of Molecular Cell now describes the molecular basis underlying the transient nature of Raf-1 signaling. PMID- 15664185 TI - Domain swapping and retroviral assembly. AB - The structure of a mammalian SCAN domain has been recently reported. The molecule is a structural homolog of the HIV-1 capsid protein and forms a domain-swapped dimer in solution. The authors propose a similar domain-swapping event facilitates HIV-1 assembly, providing a new model for protein-protein interactions underlying viral particle formation. PMID- 15664186 TI - Heterochromatin focuses on senescence. AB - A key step in cellular senescence is the packaging of proliferation-promoting genes into repressive chromatin or heterochromatin. Recent work has described a novel histone component and mode of assembly of this senescence-associated heterochromatin. PMID- 15664187 TI - Osmotin is a homolog of mammalian adiponectin and controls apoptosis in yeast through a homolog of mammalian adiponectin receptor. AB - The antifungal activity of the PR-5 family of plant defense proteins has been suspected to involve specific plasma membrane component(s) of the fungal target. Osmotin is a tobacco PR-5 family protein that induces apoptosis in the yeast Saccharomyces cerevisiae. We show here that the protein encoded by ORE20/PHO36 (YOL002c), a seven transmembrane domain receptor-like polypeptide that regulates lipid and phosphate metabolism, is an osmotin binding plasma membrane protein that is required for full sensitivity to osmotin. PHO36 functions upstream of RAS2 in the osmotin-induced apoptotic pathway. The mammalian homolog of PHO36 is a receptor for the hormone adiponectin and regulates cellular lipid and sugar metabolism. Osmotin and adiponectin, the corresponding "receptor" binding proteins, do not share sequence similarity. However, the beta barrel domain of both proteins can be overlapped, and osmotin, like adiponectin, activates AMP kinase in C2C12 myocytes via adiponectin receptors. PMID- 15664188 TI - A polybasic motif allows N-WASP to act as a sensor of PIP(2) density. AB - Phosphatidylinositol 4,5-bisphosphate (PIP(2)) activates the actin regulatory protein N-WASP by binding to a short polybasic region involved in N-WASP autoinhibition. Here, we show that unlike canonical lipid binding modules, such as PH domains, this polybasic motif binds PIP(2) in a multivalent, cooperative manner. As a result, PIP(2) activation of N-WASP-mediated actin polymerization in vitro and in extracts is ultrasensitive: above a certain threshold, N-WASP responds in a switch-like manner to a small increase in the density of PIP(2) (Hill coefficient n(H) = approximately 20). We show that the sharpness of the PIP(2) activation threshold can be tuned by varying the length of the polybasic motif. This sharp activation threshold may help suppress N-WASP activation by quiescent PIP(2) levels yet leave it poised for activation upon subtle, signaling induced perturbations in PIP(2) distribution. PMID- 15664189 TI - Crystal structure of the ligand binding suppressor domain of type 1 inositol 1,4,5-trisphosphate receptor. AB - Binding of inositol 1,4,5-trisphosphate (IP(3)) to the amino-terminal region of IP(3) receptor promotes Ca(2+) release from the endoplasmic reticulum. Within the amino terminus, the first 220 residues directly preceding the IP(3) binding core domain play a key role in IP(3) binding suppression and regulatory protein interaction. Here we present a crystal structure of the suppressor domain of the mouse type 1 IP(3) receptor at 1.8 A. Displaying a shape akin to a hammer, the suppressor region contains a Head subdomain forming the beta-trefoil fold and an Arm subdomain possessing a helix-turn-helix structure. The conserved region on the Head subdomain appeared to interact with the IP(3) binding core domain and is in close proximity to the previously proposed binding sites of Homer, RACK1, calmodulin, and CaBP1. The present study sheds light onto the mechanism underlying the receptor's sensitivity to the ligand and its communication with cellular signaling proteins. PMID- 15664190 TI - RhoA GTPase regulates B cell receptor signaling. AB - The RhoA GTPase controls many cellular functions, including gene transcription and actin polymerization. Several lines of evidence suggest that Rho GTPases are required for B cell receptor (BCR) signaling, but whether RhoA is necessary has not been investigated. Here, we show that RhoA is activated, downstream of PI3K, in response to BCR stimulation and is important for BCR-dependent calcium flux and cell proliferation. A RhoA dominant-negative mutant strongly inhibited BCR dependent calcium mobilization. The RhoA-specific inhibitor, C3 toxin, inhibited both BCR-dependent calcium flux and cell proliferation. RhoA is important for BCR dependent synthesis of IP(3) by PLCgamma2, but is not required for tyrosine phosphorylation of PLCgamma2. BCR-dependent synthesis of phosphatidylinositol-4,5 bisphosphate (PtdIns-4,5-P(2)) is inhibited in the absence of RhoA function. Providing exogenous PtdIns-4,5-P(2) restores BCR-dependent calcium flux in cells lacking functional RhoA. Our findings support a function for RhoA in BCR dependent PtdIns-4,5-P(2) synthesis, PLCgamma2 activation, calcium mobilization, and cell proliferation. PMID- 15664191 TI - Regulation of Raf-1 by direct feedback phosphorylation. AB - The Raf-1 kinase is an important signaling molecule, functioning in the Ras pathway to transmit mitogenic, differentiative, and oncogenic signals to the downstream kinases MEK and ERK. Because of its integral role in cell signaling, Raf-1 activity must be precisely controlled. Previous studies have shown that phosphorylation is required for Raf-1 activation, and here, we identify six phosphorylation sites that contribute to the downregulation of Raf-1 after mitogen stimulation. Five of the identified sites are proline-directed targets of activated ERK, and phosphorylation of all six sites requires MEK signaling, indicating a negative feedback mechanism. Hyperphosphorylation of these six sites inhibits the Ras/Raf-1 interaction and desensitizes Raf-1 to additional stimuli. The hyperphosphorylated/desensitized Raf-1 is subsequently dephosphorylated and returned to a signaling-competent state through interactions with the protein phosphatase PP2A and the prolyl isomerase Pin1. These findings elucidate a critical Raf-1 regulatory mechanism that contributes to the sensitive, temporal modulation of Ras signaling. PMID- 15664192 TI - Macrophage migration inhibitory factor MIF interferes with the Rb-E2F pathway. AB - Macrophage migration inhibitory factor (MIF) is implicated in the regulation of inflammation and cell growth. We previously showed that MIF is a potent modulator of p53- and E2F-dependent pathways that are activated in response to oncogenic signaling. Here, we characterize the functional link between MIF and E2F transcription factors. Our results demonstrate that MIF-deficient cells exhibit E2F-dependent growth alterations and reduced susceptibility to oncogenic transformation. The basis for this transformation resistance is a perturbed function of the C-terminal Rb binding region of E2F4. However, inactivation of Rb or substitution of the E2F4 C-terminal domain by the E2F1 C-terminal region rescues the transformation defect. Importantly, the involvement of E2F factors in DNA replication rather than in regulation of transcription determines their oncogenic properties in the context of MIF deficiency. A proinflammatory molecule interfering with tumor suppression and DNA replication provides a compelling molecular link for the association of chronic inflammation and tumorigenesis. PMID- 15664193 TI - Regulation of p21(WAF1/CIP1) stability by WISp39, a Hsp90 binding TPR protein. AB - p21(WAF1/CIP1), a cyclin-dependent kinase inhibitor and a critical regulator of cell cycle, is controlled transcriptionally by p53-dependent and -independent mechanisms and posttranslationally by the proteasome. We have identified WISp39, a tetratricopeptide repeat (TPR) protein that binds p21. WISp39 stabilizes newly synthesized p21 protein by preventing its proteasomal degradation. WISp39, p21, and hsp90 form a trimeric complex in vivo. The interaction of WISp39 with Hsp90 is abolished by point mutations within the C-terminal TPR domain of WISp39. Although this WISp39 TPR mutant binds p21 in vivo, it fails to stabilize p21. Our results suggest that WISp39 recruits Hsp90 to regulate p21 protein stability. WISp39 downregulation by siRNA prevents the accumulation of p21 and cell cycle arrest after ionizing radiation. The results demonstrate the importance of posttranslational stabilization of p21 protein by WISp39 in regulating cellular p21 activity. PMID- 15664194 TI - E6 oncoprotein represses p53-dependent gene activation via inhibition of protein acetylation independently of inducing p53 degradation. AB - The mechanism employed by DNA tumor viruses to inhibit p53-dependent transcription from chromatin is poorly understood. Here, we use in vitro reconstituted chromatin and UV-irradiated cells to define the mechanism of human papillomavirus E6 oncoprotein in repressing p53-dependent transcription. We demonstrate that E6 does not prevent p53 or p300 recruitment to the chromatin but inhibits p300-mediated acetylation on p53 and nucleosomal core histones. This suppression of protein acetylation requires the E6-interacting regions of p300. Moreover, E6 mutants unable to interact with p53 or p300, but not deficient in inducing p53 degradation, fail to inhibit p53-mediated activation, indicating that a p53-E6-p300-containing protein complex is critical for repressing p53 targeted gene activation. That E6 acts as a molecular switch converting p53-p300 from an activating complex to a repressing entity on the chromatin, which occurs independently of E6AP-mediated protein degradation pathway, may represent a general mechanism for gene regulation. PMID- 15664195 TI - A conformational change in the eukaryotic translation preinitiation complex and release of eIF1 signal recognition of the start codon. AB - During eukaryotic translation initiation, ribosomal 43S complexes scan mRNAs for the correct AUG codon at which to begin translation. Start codon recognition triggers GTP hydrolysis, committing the complex to engagement at that point on the mRNA. While fidelity at this step is essential, the nature of the codon recognition event and the mechanism by which it activates GTP hydrolysis are poorly understood. Here we report the changes that occur within the 43S.mRNA complex in response to AUG codon recognition. eIF1 and eIF1A are key players in assembly of 43S.mRNA complexes capable of locating initiation codons. We observed FRET between these two factors when bound to the 40S subunit. Using steady-state FRET, anisotropy, and kinetic analyses, we demonstrate that start codon recognition results in a conformational change and release of eIF1 from the ribosome. These rearrangements probably play a role in triggering GTP hydrolysis and committing the complex to downstream events. PMID- 15664196 TI - Formation of a dynamic kinetochore- microtubule interface through assembly of the Dam1 ring complex. AB - How kinetochore proteins form a dynamic interface with microtubules is largely unknown. In budding yeast, the 10-protein Dam1 complex is an Aurora kinase target that plays essential roles maintaining the integrity of the mitotic spindle and regulating interactions with the kinetochore. Here, we investigated the biochemical properties of purified Dam1 complex. The complex oligomerized into rings around microtubules. Ring formation was facilitated by microtubules but could occur in their absence. Mutant alleles led to partially assembled complexes or reduced microtubule binding. The interaction between rings and microtubules is mediated by the C termini of both Dam1 and alphabeta-tubulin. Ring formation promotes microtubule assembly, stabilizes against disassembly, and promotes bundling. A GTP-tubulin lattice is the preferred binding partner for the complex, and Dam1 rings can exhibit lateral mobility on microtubules. These observations suggest a mechanism by which the kinetochore can recognize and stay attached to the plus ends of microtubules. PMID- 15664197 TI - LexA cleavage is required for CTX prophage induction. AB - The physiologic conditions and molecular interactions that control phage production have been studied in few temperate phages. We investigated the mechanisms that regulate production of CTXphi, a temperate filamentous phage that infects Vibrio cholerae and encodes cholera toxin. In CTXphi lysogens, the activity of P(rstA), the only CTXphi promoter required for CTX prophage development, is repressed by RstR, the CTXvphi repressor. We found that the V. cholerae SOS response regulates CTXvphi production. The molecular mechanism by which this cellular response to DNA damage controls CTXphi production differs from that by which the E. coli SOS response controls induction of many prophages. UV-stimulated CTXphi production required RecA-dependent autocleavage of LexA, a repressor that controls expression of numerous host DNA repair genes. LexA and RstR both bind to and repress P(rstA). Thus, CTXphi production is controlled by a cellular repressor whose activity is regulated by the cell's response to DNA damage. PMID- 15664198 TI - Human Asf1 regulates the flow of S phase histones during replicational stress. AB - Maintenance of chromosomal integrity requires tight coordination of histone biosynthesis with DNA replication. Here, we show that extracts from human cells exposed to replication stress display an increased capacity to support replication-coupled chromatin assembly. While in unperturbed S phase, hAsf1 existed in equilibrium between an active form and an inactive histone-free pool, replication stress mobilized the majority of hAsf1 into an active multichaperone complex together with histones. This active multichaperone complex was limiting for chromatin assembly in S phase extracts, and hAsf1 was required for the enhanced assembly activity in cells exposed to replication stress. Consistently, siRNA-mediated knockdown of hAsf1 impaired the kinetics of S phase progression. Together, these data suggest that hAsf1 provides the cells with a buffering system for histone excess generated in response to stalled replication and explains how mammalian cells maintain a critical "active" histone pool available for deposition during recovery from replication stresses. PMID- 15664200 TI - Health and poverty: a new Marshall plan? PMID- 15664199 TI - An important role for RNase R in mRNA decay. AB - mRNA decay is a major determinant of gene expression. In Escherichia coli, message degradation initiates with an endoribonucleolytic cleavage followed by exoribonuclease digestion to generate 5'-mononucleotides. Although the 3' to 5' processive exoribonucleases, PNPase and RNase II, have long been considered to be mediators of this digestion, we show here that another enzyme, RNase R, also participates in the process. RNase R is particularly important for removing mRNA fragments with extensive secondary structure, such as those derived from the many mRNAs that contain REP elements. In the absence of RNase R and PNPase, REP containing fragments accumulate to high levels. RNase R is unusual among exoribonucleases in that, by itself, it can digest through extensive secondary structure provided that a single-stranded binding region, such as a poly(A) tail, is present. These data demonstrate that RNase R, which is widespread in prokaryotes and eukaryotes, is an important participant in mRNA decay. PMID- 15664201 TI - thnx sir bill. c u in 2009? PMID- 15664202 TI - Translating compassion into psychosocial aid after the tsunami. PMID- 15664203 TI - Managing tsunami risk. PMID- 15664204 TI - Controlling meticillin-resistant Staphylococcus aureus, aka "Superbug". PMID- 15664205 TI - Unbiased studies are needed before virtual colonoscopy can be dismissed. PMID- 15664206 TI - Hepatitis C virus in blood donation. PMID- 15664207 TI - Treatment of AIDS in conflict-affected settings: a failure of imagination. PMID- 15664208 TI - Poly-ticks: Blue State versus Red State for Lyme disease. PMID- 15664212 TI - UNICEF: suggestions for change. PMID- 15664213 TI - UNICEF: suggestions for change. PMID- 15664214 TI - UNICEF: suggestions for change. PMID- 15664215 TI - UNICEF: suggestions for change. PMID- 15664216 TI - UNICEF: suggestions for change. PMID- 15664217 TI - UNICEF: suggestions for change. PMID- 15664218 TI - UNICEF: suggestions for change. PMID- 15664219 TI - UNICEF: suggestions for change. PMID- 15664220 TI - UNICEF: suggestions for change. PMID- 15664222 TI - UNICEF: suggestions for change. PMID- 15664221 TI - Corticosteroids and acute asthma. PMID- 15664223 TI - UNICEF: suggestions for change. PMID- 15664224 TI - Isolation of patients in single rooms or cohorts to reduce spread of MRSA in intensive-care units: prospective two-centre study. AB - BACKGROUND: Hospital-acquired infection due to meticillin-resistant Staphylococcus aureus (MRSA) is common within intensive-care units. Single room or cohort isolation of infected or colonised patients is used to reduce spread, but its benefit over and above other contact precautions is not known. We aimed to assess the effectiveness of moving versus not moving infected or colonised patients in intensive-care units to prevent transmission of MRSA. METHODS: We undertook a prospective 1-year study in the intensive-care units of two teaching hospitals. Admission and weekly screens were used to ascertain the incidence of MRSA colonisation. In the middle 6 months, MRSA-positive patients were not moved to a single room or cohort nursed unless they were carrying other multiresistant or notifiable pathogens. Standard precautions were practised throughout. Hand hygiene was encouraged and compliance audited. FINDINGS: Patients' characteristics and MRSA acquisition rates were similar in the periods when patients were moved and not moved. The crude (unadjusted) Cox proportional hazards model showed no evidence of increased transmission during the non-move phase (0.73 [95% CI 0.49-1.10], p=0.94 one-sided). There were no changes in transmission of any particular strain of MRSA nor in handwashing frequency between management phases. INTERPRETATION: Moving MRSA-positive patients into single rooms or cohorted bays does not reduce crossinfection. Because transfer and isolation of critically ill patients in single rooms carries potential risks, our findings suggest that re-evaluation of isolation policies is required in intensive-care units where MRSA is endemic, and that more effective means of preventing spread of MRSA in such settings need to be found. PMID- 15664225 TI - Analysis of air contrast barium enema, computed tomographic colonography, and colonoscopy: prospective comparison. AB - BACKGROUND: The usefulness of currently available colon imaging tests, including air contrast barium enema (ACBE), computed tomographic colonography (CTC), and colonoscopy, to detect colon polyps and cancers is uncertain. We aimed to assess the sensitivity of these three imaging tests. METHODS: Patients with faecal occult blood, haematochezia, iron-deficiency anaemia, or a family history of colon cancer underwent three separate colon-imaging studies--ACBE, followed 7-14 days later by CTC and colonoscopy on the same day. The primary outcome was detection of colonic polyps and cancers. Outcomes were assessed by building an aggregate view of the colon, taking into account results of all three tests. FINDINGS: 614 patients completed all three imaging tests. When analysed on a per patient basis, for lesions 10 mm or larger in size (n=63), the sensitivity of ACBE was 48% (95% CI 35-61), CTC 59% (46-71, p=0.1083 for CTC vs ACBE), and colonoscopy 98% (91-100, p<0.0001 for colonoscopy vs CTC). For lesions 6-9 mm in size (n=116), sensitivity was 35% for ACBE (27-45), 51% for CTC (41-60, p=0.0080 for CTC vs ACBE), and 99% for colonoscopy (95-100, p<0.0001 for colonoscopy vs CTC). For lesions of 10 mm or larger in size, the specificity was greater for colonoscopy (0.996) than for either ACBE (0.90) or CTC (0.96) and declined for ACBE and CTC when smaller lesions were considered. INTERPRETATION: Colonoscopy was more sensitive than other tests, as currently undertaken, for detection of colonic polyps and cancers. These data have important implications for diagnostic use of colon imaging tests. PMID- 15664226 TI - Ethnic variations in UK asthma frequency, morbidity, and health-service use: a systematic review and meta-analysis. AB - BACKGROUND: The frequency of asthma varies between countries, and may also vary between ethnic groups in more geographically confined areas. We sought evidence of such ethnic variations in the UK for asthma frequency, morbidity, and health services use, and to understand possible reasons for any differences. METHODS: We searched MEDLINE, EMBASE, CINAHL, PSYCHInfo, PREMEDLINE, HEALTHSTAR, Cambridge Register of Conference Abstracts, the Dissertation and Thesis Database, and the National Registry of Research. Additionally, we searched the bibliographies of reports identified and websites of health authorities, and contacted experts in this discipline. Our main outcomes were comparisons of asthma rate, morbidity, and health-services use. We did meta-analyses using random-effects models. FINDINGS: 13 studies contained relevant data. All prevalence studies were of children and showed that south Asian children had a lower frequency of symptoms suggestive of asthma compared with black and white children (pooled rate of history of wheeze in the previous 12 months: south Asians 9.6% [95%CI 8.0-11.2%], black people 16.2% [12.8-19.6%], white people 14.6% [11.5-17.8%]). The pooled frequency of clinician-diagnosed asthma in children followed a similar pattern (south Asians 7.6% [3.7-11.4%], black people 15.0% [3.5-26.5%], white people 10.6% [4.6-16.7%]. However, relative to white people, the risk of admission for asthma in children and adults was higher for south Asians (odds ratio 2.9 [2.4 3.4]) and black people (2.1 [1.8-2.5]). INTERPRETATION: The differences in admission are not explained by differences in asthma frequency between groups; they could relate to ethnic variations in asthma severity, differences in health seeking behaviour, or difficulties in accessing high-quality primary care services. PMID- 15664227 TI - Clinical outcome of individualised treatment of multidrug-resistant tuberculosis in Latvia: a retrospective cohort study. AB - BACKGROUND: Latvia has one of the highest rates of multidrug-resistant tuberculosis (MDRTB). Our aim was to assess treatment outcomes for the first full cohort of MDRTB patients treated under Latvia's DOTS-Plus strategy following WHO guidelines. METHODS: We retrospectively reviewed all civilian patients who began treatment with individualised treatment regimens for pulmonary MDRTB in Latvia between Jan 1, and Dec 31, 2000. We applied treatment outcome definitions for MDRTB, developed by an international expert consensus group, and assessed treatment effectiveness and risk factors associated with poor outcome. FINDINGS: Of the 204 patients assessed, 55 (27%) had been newly diagnosed with MDRTB, and 149 (73%) had earlier been treated with first-line or second-line drugs for this disease. Assessment of treatment outcomes showed that 135 (66%) patients were cured or completed therapy, 14 (7%) died, 26 (13%) defaulted, and treatment failed in 29 (14%). Of the 178 adherent patients, 135 (76%) achieved cure or treatment completion. In a multivariate Cox proportional-hazards model of these patients, independent predictors of poor outcome (death and treatment failure) included having previously received treatment for MDRTB (hazard ratio 5.7, 95% CI 1.9-16.6), the use of five or fewer drugs for 3 months or more (3.2, 1.1-9.6), resistance to ofloxacin (2.6, 1.2-5.4), and body-mass index less than 18.5 at start of treatment (2.3, 1.1-4.9). INTERPRETATION: The DOTS-Plus strategy of identifying and treating patients with MDRTB can be effectively implemented on a nationwide scale in a setting of limited resources. PMID- 15664228 TI - T-cell responses and previous exposure to hepatitis C virus in indeterminate blood donors. AB - Blood donors are routinely screened for hepatitis C virus infection. Some individuals have weak or restricted virus-specific antibody responses, and are classed as indeterminate. Such donors are almost always negative for viral RNA in blood. We postulated that previous transient virus exposure might account for some of these cases. With sensitive ex-vivo analyses of T-cell responses, we identified virus-specific responses in 15 of 30 indeterminate blood donors tested, compared with none in controls (p=0.0013). Additionally, these responses were typically focused on core-derived peptides. These findings suggest previous exposure to the virus in many indeterminate blood donors. PMID- 15664229 TI - Intraocular haemorrhage after thrombolysis. PMID- 15664230 TI - Chronic kidney disease: the global challenge. AB - The worldwide rise in the number of patients with chronic kidney disease (CKD) and consequent end-stage renal failure necessitating renal replacement therapy is threatening to reach epidemic proportions over the next decade, and only a small number of countries have robust economies able to meet the challenges posed. A change in global approach to CKD from treatment of end-stage renal disease (ESRD) to much more aggressive primary and secondary prevention is therefore imperative. In this Seminar, we examine the epidemiology of CKD worldwide, with emphasis on early detection and prevention, and the feasibility of methods for detection and primary prevention of CKD. We also review the risk factors and markers of progressive CKD. We explore current understanding of the mechanisms underlying renal scarring leading to ESRD to inform on current and future interventions as well as evidence relating to interventions to slow the progression of CKD. Finally, we make strategic recommendations based on future research to stem the worldwide growth of CKD. Consideration is given to health economics. A global and concerted approach to CKD must be adopted in both more and less developed countries to avoid a major catastrophe. PMID- 15664231 TI - Treating individuals 4: can meta-analysis help target interventions at individuals most likely to benefit? AB - Meta-analyses of randomised trials aim to summarise the effects of interventions across many patients, and can seem remote from the clinical issue of how individual patients should be treated and which patient groups will benefit the most from treatment. One method that attempts to address this point entails relating the overall effect in every trial to summaries of patient characteristics. This is called meta-regression. The interpretation of such analyses is not straightforward, however, because of a combination of confounding and other biases. Much more useful is to compare the outcomes for patient subgroups within trials and combine these results across trials. Unfortunately this method is rarely possible using published information, so analyses of individual patient data from trials are necessary. Also, although meta-analyses generally summarise an intervention's effect as a relative risk reduction, the groups of patients with the greatest absolute risk reduction have the most to gain. PMID- 15664232 TI - The Millennium Project: a plan for meeting the Millennium Development Goals. PMID- 15664233 TI - Hard choices: rationing antiretroviral therapy for HIV/AIDS in Africa. PMID- 15664234 TI - Unrecognised comorbidities. PMID- 15664235 TI - A fatal encephalitis. PMID- 15664237 TI - The value and limitations of long-term nucleoside antiviral therapy in chronic hepatitis B. PMID- 15664238 TI - Paralysed monocytes in acute on chronic liver disease. PMID- 15664239 TI - Mushrooms, amatoxins and the liver. PMID- 15664240 TI - Intestinal flora and bilirubin. PMID- 15664241 TI - Extended lamivudine treatment in patients with HBeAg-negative chronic hepatitis B. AB - BACKGROUND/AIMS: The histological and clinical outcome of lamivudine 100mg/day was assessed in 76 HBeAg-negative chronic hepatitis B patients previously randomised to a double-blind comparison study of lamivudine and placebo. METHODS: Paired liver biopsies were available before 1 year of randomised lamivudine treatment and after 2 years of further open-label treatment for 48 patients. Serum samples were analysed for hepatitis B markers and ALT levels (n=74). RESULTS: The histological activity index improved, remained unchanged and worsened in 64, 32 and 5%, respectively, for patients without YMDD-variant HBV compared to 15, 54 and 31% with the variant. None of the 42/48 patients without cirrhosis at baseline progressed to cirrhosis. Of 24/48 patients without bridging fibrosis at pre-treatment, 83% (20/24) did not progress to bridging fibrosis. Median HBV DNA remained below the lower limit of detection and ALT < or =1 times the ULN for patients without the variant whereas levels gradually increased to 11.3Meq/ml (bDNA assay) and 2 times the upper limit of normal by month 24 for patients with variant. CONCLUSIONS: The clinical benefit of lamivudine is greatest for patients without YMDD variants over 2 years of extended treatment. Additional therapies should be considered for patients with YMDD variants. PMID- 15664242 TI - Sequence-specific inhibition of duck hepatitis B virus reverse transcription by peptide nucleic acids (PNA). AB - BACKGROUND/AIMS: Peptide nucleic acids (PNAs) appear as promising new antisense agents, that have not yet been examined as hepatitis B virus (HBV) inhibitors. Our aim was to study the ability of PNAs targeting the duck HBV (DHBV) encapsidation signal epsilon to inhibit reverse transcription (RT) and to compare their efficacy with phosphorothioate oligodeoxynucleotides (S-ODNs). METHODS: The effect of two partly overlapping PNAs targeting epsilon and of analogous S-ODNs was tested in cell-free transcription and translation system for DHBV RT expression. In addition their antiviral effect was investigated in primary duck hepatocytes (PDH). RESULTS: Both PNAs reproducibly inhibited DHBV RT in a dose dependent manner with IC(50) of 10nM, whereas up to 600-fold higher concentration of S-ODNs was required for similar inhibition. The PNA targeting the bulge and upper stem of epsilon appeared as more efficient RT inhibitor than the PNA targeting only the bulge. Importantly, the inhibition was highly sequence specific since double-mismatched PNA had no effect on the RT reaction. Moreover, in PDH the PNA coupled to Arg(7) cationic delivery peptide decreased DHBV replication. CONCLUSIONS: We provide the first evidence that PNAs targeting the bulge and upper stem of epsilon can efficiently and in a sequence-specific manner inhibit DHBV RT. PMID- 15664243 TI - Long-term clinical and histological outcomes in patients with spontaneous hepatitis B surface antigen seroclearance. AB - BACKGROUND/AIMS: During the natural course of hepatitis B virus (HBV) infection, the long-term clinical and histological outcomes following spontaneous hepatitis B surface antigen (HBsAg) seroclearance remain unclear. METHODS: Between 1984 and 2003, 49 (9.5%) out of 432 inactive HBsAg carriers had no detectable level of circulating HBsAg. Fifteen of 49 patients had undergone paired peritoneoscopic liver biopsies. RESULTS: During a mean follow-up period of 19.6 months after HBsAg seroclearance, 5 of 49 (10.2%) patients were noted to have HCC. Liver cirrhosis (P=0.040), a history of perinatal infection (P=0.005) and long-standing duration (at least 30 years) of HBsAg positivity (P=0.002) were associated with a significantly higher risk of developing HCC. Despite HBsAg seroclearance, HBV DNA was detected in the liver tissues from all 15 patients who underwent paired liver biopsies. Necroinflammation was significantly ameliorated (P<0.0001). On the other hand, amelioration of the fibrosis score did not reach a statistically significant level (P=0.072). Interestingly, aggravation of liver fibrosis was evident in 2 patients (13.3%) including one who had rapidly progressed to overt cirrhosis. CONCLUSIONS: In patients with spontaneous HBsAg seroclearance, necroinflammation was markedly improved and liver fibrosis was unchanged or regressed despite occult HBV infection. However, HCC developed in a minority of cases. PMID- 15664244 TI - Patients with acute on chronic liver failure display "sepsis-like" immune paralysis. AB - BACKGROUND/AIMS: Cellular immune depression is linked to high mortality in sepsis, but has yet to be systematically analysed in liver cirrhosis. The aim of the present study was to directly compare functional immune parameters in patients with acute on chronic liver failure (ACLF), severe sepsis, and non decompensated cirrhosis. METHODS: Patients with ACLF (n=27) were investigated at admission to a medical ICU. Patients with stable liver cirrhosis (n=24) and severe sepsis (n=31) served as control groups. In all subjects, serum levels of IL-6 and IL-10, ex vivo production of TNF-alpha in a whole blood assay, and monocyte surface HLA-DR expression were determined. RESULTS: In patients with ACLF or sepsis, ex vivo TNF-alpha production and HLA-DR expression were severely decreased compared to subjects with stable cirrhosis (both P<0.001). Contrary, IL 6 levels were highest in septic patients, followed by subjects with ACLF and cirrhotic patients (both P<0.05). Immune dysfunction in ACLF was independent of aetiology of liver cirrhosis and associated with high mortality. CONCLUSIONS: Patients with ACLF and severe sepsis show a similar degree of cellular immune depression. The reduced cellular immune function in subjects with ACLF might contribute to the increased infectious morbidity of these patients and provide a rational basis for prevention strategies. PMID- 15664245 TI - Indication of liver transplantation following amatoxin intoxication. AB - BACKGROUND/AIMS: Indication of liver transplantation in acute liver failure following amatoxin intoxication is still uncertain. METHODS: One hundred and ninety-eight patients were studied retrospectively. The laboratory parameters alanine-aminotransferase, serum bilirubin, serum creatinine and prothrombin index were analyzed over time. Predictors of fatal outcome and survival were determined by receiver-operating-characteristic and sensitivity-specificity analysis. RESULTS: Twenty-three patients died in the median 6.1 days (range, 2.7-13.9 days) after ingestion. Using a single parameter as predictor of fatal outcome the area under the receiver-operating-characteristic curve of prothrombin index (0.96) and serum creatinine (0.93) were both significantly greater (P<0.05) compared with serum bilirubin (0.82) and alanine-aminotransferase (0.69). Prediction of fatal outcome had an optimum, if a prothrombin index less than 25% was combined with a serum creatinine greater than 106 micromol/l from day 3 after ingestion onwards (sensitivity 100%, 95% confidence interval 87-100; specificity 98%, 95% confidence interval 94-100). The median time period between the first occurrence of this predictor in non-survivors and death was 63h (range, 3-230h). CONCLUSIONS: A decision model of liver transplantation following amatoxin intoxication using prothrombin index in combination with serum creatinine from day 3 to 10 after ingestion enables an early and reliable assessment of outcome. PMID- 15664246 TI - Apoptosis on hepatoma cells but not on primary hepatocytes by histone deacetylase inhibitors valproate and ITF2357. AB - BACKGROUND/AIMS: Due to a particular resistance against conventional chemotherapeutics, palliative treatment of hepatocellular carcinomas (HCC) is highly ineffective. Recent demonstration of both proliferation-inhibition and apoptosis of hepatoma cells by a histone deacetylase inhibitor (HDAC-I) treatment opens up a promising new approach. However, little is known about tumor cell death mechanisms and HDAC-I influences on healthy hepatocytes. METHODS: HDAC-I substances with favourable in vivo profiles, valproate (VPA) and ITF2357, were investigated on HCC cell lines and primary human hepatocytes (PHH). Histone acetylation and apoptosis-modulating proteins were investigated by western blotting, proliferation by sulforhodamin B binding, toxicity by enzyme release, apoptosis by FACS analysis. RESULTS: VPA and ITF2357 inhibited proliferation in HCC cell lines. Both substances induced considerable cellular damage in HCC derived cells, but PHH tolerated these substances well. A downregulation of anti- and upregulation of proapoptotic factors was found. Moreover, Bcl-X(L) transfection into HCC cells abrogated apoptosis induced by both substances, indicating that modulation of intracellular pro- and anti-apoptotic proteins is a key event in VPA or ITF2357 induced tumor-cell death. CONCLUSIONS: Preferential induction of cell death in HCC-derived cell lines, without toxicity in PHH, demonstrates the potential of VPA and ITF2357 to become promising new tools in the fight against HCC. PMID- 15664247 TI - Alcohol, tobacco and obesity are synergistic risk factors for hepatocellular carcinoma. AB - BACKGROUND/AIMS: Alcohol has been shown to be an important risk factor for hepatocellular carcinoma (HCC). The role of tobacco as a risk factor for HCC is controversial. Recently, obesity has been reported to be a risk factor for HCC. We investigated whether these factors increase the risk of HCC in American patients. METHODS: Consecutive patients with HCC, cirrhosis without HCC and, control patients without liver disease were enrolled and exposure to risk factors was assessed. RESULTS: When HCC cases were compared to cirrhotic controls, the risk of HCC increased 6-fold for alcohol (OR 5.7; 95% CI: 2.4-13.7), 5-fold for tobacco (OR 4.9; 95% CI: 2.2-10.6), and 4-fold with obesity (OR 4.3; 95% CI: 2.1 8.4). Using spline regression, a dose-dependent relationship between alcohol and tobacco exposure with risk of HCC was noted. There was significant interaction between alcohol, tobacco and obesity, with synergistic indices greater than 1. CONCLUSIONS: Alcohol, tobacco and obesity are independent risk factors for HCC in our patient population, and they interact synergistically to increase the risk of HCC. Data from this study may allow us to stratify cirrhotics into low- and high risk groups for the development of HCC surveillance strategies. PMID- 15664248 TI - Multistep and multicentric development of hepatocellular carcinoma: histological analysis of 980 resected nodules. AB - BACKGROUND/AIMS: Histological observations support the concept of multistep and multicentric development of hepatocellular carcinoma (HCC) in cases of chronic liver disease. However, the relationship between the incidence of such a modality of development of HCC and the type of background liver disease has not been fully investigated. METHODS: A total of 980 HCC nodules resected from 664 patients were analyzed. Multistep HCC was defined as well differentiated HCC containing the portal tracts (early HCC), or the presence of early HCC-like areas in the periphery of the nodule. In cases with multiple nodules, if the smaller nodule showed the features of multistep HCC, or if each nodule showed a distinct histology, the case was defined to have multicentric HCC. RESULTS: Of the 980 nodules, 369 (37.7%) met the criteria of multistep HCC. Of the 664 patients, 177 (26.7%) had multiple nodules that met the criteria of multicentric HCC. Both the incidences of multistep and multicentric HCC were significantly higher in HCV-Ab positive cases than in HBs-Ag-positive cases (46.0 vs. 19.1%, P<0.001 and 34.1 vs. 16.5%, P=0.005, respectively). CONCLUSIONS: Multistep and multicentric HCC develops most frequently in patients with HCV infection. PMID- 15664249 TI - Potent induction of apoptosis in human hepatoma cell lines by targeted cytotoxic somatostatin analogue AN-238. AB - BACKGROUND/AIMS: The efficacy of a targeted cytotoxic hybrid somatostatin analogue AN-238 and of its superactive radical 2-pyrrolinodoxorubicin (AN-201) to induce apoptosis of HepG2 and Hep3B human hepatoma cell lines were studied. AN 238 was designed to selectively target tumor cells expressing somatostatin receptor subtypes (sst(s)). Its effects on HepG2 or Hep3B cells displaying or lacking tumor suppressor p53, respectively, were compared. Normal rat isolated hepatocytes were also tested. METHODS: sst(s) were characterized by binding assays and RT-PCR. Cytotoxicity was quantified by flow cytometry. DNA fragmentation was studied by gel electrophoresis, PARP cleavage by Western blot and ROS formation using fluorescent probes. RESULTS: Specific binding of iodinated RC-160 to HepG2 and Hep3B cells, and its displacement by AN-238 was characterized. mRNA for hsst(2A) was found in both cell lines. Flow cytometry showed a stronger effect of AN-238 than AN-201 to induce sub-G1 phase. DNA fragmentation, nuclear bodies, and PARP cleavage were observed. In addition, AN 238 increased formation of ROS more potently than AN-201. However, no inductions of DNA fragmentation by AN-201 or AN-238 were observed on rat hepatocytes. CONCLUSIONS: Our results indicate that, in liver cancer, the cytotoxic somatostatin analogue AN-238 is a powerful agent that can induce apoptosis, through sst(s) and independently of p53. PMID- 15664250 TI - The impact of intestinal microflora on serum bilirubin levels. AB - BACKGROUND/AIMS: Intestinal microflora plays an important role in the pathogenesis of neonatal jaundice by inhibiting enterosystemic circulation of bilirubin. The present study aimed to investigate the influence of intestinal microflora on serum bilirubin levels in hyperbilirubinemic Gunn rats. METHODS: After a baseline phase Gunn rats received oral antibiotics (either clindamycin/neomycine or co-trimethoxazole for four days, phase II). Intestinal colonization was carried out either with a bilirubin-reducing strain of C. perfringens or C. pasteurianum incapable of reducing bilirubin (phase III). Serum bilirubin and fecal bile pigments were determined at the end of each phase. RESULTS: Oral administration of clindamycin/neomycine resulted in the disappearance of fecal urobilinoids. Simultaneously, serum bilirubin increased dramatically (186+/-31 vs. 289+/-35 micromol/l, P=0.004). Intestinal colonization with C. perfringens led to reappearance of fecal urobilinoid production accompanied with a partial decrease of serum bilirubin (289+/-35 vs. 239+/-17 micromol/l, P=0.013), whereas the effect of C. pasteurianum on bile pigment metabolism was negligible. Co-trimethoxazole therapy had no effect on serum and intestinal metabolism of bilirubin. CONCLUSIONS: Intestinal microflora greatly affects intravascular metabolism of bilirubin. Prolonged use of certain antibiotics in man may lead to an increase in serum bilirubin levels, while the enhancement of intestinal catabolism may have an opposite effect. PMID- 15664252 TI - Increased prevalence of primary sclerosing cholangitis among first-degree relatives. AB - BACKGROUND/AIMS: The aim of the present study was to investigate the familial occurrence of autoimmune diseases in a large group of patients with primary sclerosing cholangitis (PSC). METHODS: All patients with PSC treated at Huddinge University Hospital between 1984 and 1999 were included (n=145). For every patient with PSC and inflammatory bowel disease (IBD) (n=126) we randomly selected a control patient with IBD (n=126), matched for age, sex and type of IBD. A questionnaire comprising information about autoimmune diseases among first degree relatives was answered by all patients and controls. RESULTS: We identified 22 index cases with PSC from 21 families with a first-degree relative with either chronic liver disease and/or IBD. Five patients with PSC had a first degree relative with PSC (3.4%). The prevalence of PSC among first-degree relatives was 0.7% (5/717). In siblings the prevalence was 1.5% (4/269). The prevalence of first-degree relatives with autoimmune diseases outside the liver was similar in PSC patients and controls. CONCLUSIONS: First-degree relatives of patients with PSC have a PSC prevalence of 0.7%. This represents a nearly 100 fold increased risk of developing PSC compared with the general population, supporting the hypothesis that genetic factors are of importance for development of PSC. PMID- 15664251 TI - Cholestatic bile acids inhibit gap junction permeability in rat hepatocyte couplets and normal rat cholangiocytes. AB - BACKGROUND/AIMS: The aim of this work was to study the effects of different bile acids on the permeability of gap junction channels (PGJC). We also looked at the effects of some bile acids on the coordination of intercellular calcium oscillations. METHODS: The permeability of gap junctions was assessed by fluorescent dye transfer and calcium signalling on fluorescent microscopy. RESULTS: Cholestatic bile acids such as taurolithocholate, taurolithocholate sulfate and taurochenodeoxycholate inhibit the permeability of gap junctions in a dose-dependent and reversible manner in hepatocytes. Experiments performed in other cell types suggest that this effect is specific for cells having bile salt transporters, independently of the type of connexin expressed in these cells. Thus, cholestatic bile acids inhibit PGJC in normal rat cholangiocytes which express Cx43, but not in HeLa cells transfected with Cx26 or 32, which are expressed in hepatocytes. Calcium oscillations induced by bile acids in rat hepatocyte couplets are not coordinated and, by inhibiting the PGJC, cholestatic bile acids prevent the coordination of calcium oscillations induced by noradrenaline in these cells. CONCLUSIONS: Cholestatic, but not choleretic bile acids inhibit the PGJC in cells able to accumulate bile acids. This inhibition might contribute to the cholestatic effect of these bile acids. PMID- 15664253 TI - Adaptative bile duct proliferative response in experimental bile duct ischemia. AB - BACKGROUND/AIMS: A rat model of bile duct ischemia was established and used to examine the potential of bile duct proliferation to provide an adaptative response in cholestatic disorders. METHODS: Rats underwent partial or complete arterial deprivation of the liver. Serum biochemical tests, histological analyses and bile secretion measurements were performed at different time points up to 6 weeks after surgery. RESULTS: Rats developed biochemical signs of cholestasis exclusively after complete arterial deprivation. Within 4h, cholangiocytes in these rats showed morphological signs of cell damage. After 48h, they displayed VEGF expression and became proliferative. The proportion of Ki67-labeled cholangiocytes ( approximately 30%) was similar in interlobular bile ducts and periportal ductules. A ductular reaction made of well-formed bile ducts confined to portal tracts developed within 1 week. Bile flow which was initially decreased, was restored at 3 weeks, while the biochemical signs of cholestasis completely resolved at 6 weeks. At this time, the number of bile duct sections was maximal. Fibrosis intensity was also maximal, although moderate (100 microM when bacteria are exposed to DCB and light at the same time (1.2 J/cm2 of UVA and 2.1 J/cm2 of visible light). Furthermore, DCB is both photocytotoxic and photogenotoxic to human Jurkat T-cells. Under a light irradiation dose of 2.3 J/cm2 of UVA and 4.2 J/cm2 of visible light, it causes the Jurkat T-cells to become nonviable in a DCB dose-dependent manner and the nonviable cells reaches 60% at DCB concentrations higher than 50 microM. At the same time, DNA fragmentation is observed for cells exposed to both DCB and light, determined by single cell gel electrophoresis (alkaline comet assay). As much as 5% (average) DNA fragmentation was observed when exposed to 200 microM DCB and light irradiation. This suggests that DCB can penetrate the cell membrane and enter the cell. Upon light activation, DCB in the cells can cause various cellular damages, leading to nonviable Jurkat T-cells. It appears, the nonviable cells are not caused solely by fragmentation of cellular DNA, but by other damages such as to proteins and cell membranes, or DNA alkylation. Therefore, persons exposed to DCB through environmental contamination or through tattoo piercing using DCB-containing inks must not only concern about its toxicity without exposing to light, but also its phototoxicity. PMID- 15664271 TI - Immuno-histochemical detection of MRPs in human lung cells in culture. AB - The transport of molecules across membranes is an essential function of all living organisms and a large number of specific transporters have evolved to carry out this function. The largest transporter gene family is the ATP-binding cassette (ABC) transporter superfamily. The multidrug resistance-associated protein (MRP) family is comprised of nine related ABC transporters. The intra cellular distribution of the different MRP isoforms in relation to their physiological and non physiological function is still a point of discussion. For this purpose we used normal human lung cells (bronchial epithelial cells, NHBEC, and peripheral lung cells, PLC) as well as tumor cell cultures as test tools to investigate the intracelluar localization of these proteins under classical culture conditions and under air-liquid interface by means of indirect fluorescence microscopy. Characterization of the cultured cells as lung epithelial cells was performed by means of immuno-histochemical analysis. MRP1 and MRP3 were localised to the cellular membrane in all tested lung cell types. In contrast to that MRP2, MRP4 and MRP5 could be described as intracellular proteins in NHBEC and PLC. All MRP1-MRP5 isoforms could be characterized in A549 tumor cell line as membrane proteins. In order to imitate the physiological in vivo circumstances in the lung, we have established a dry/wet method (air-liquid interface) for cell cultivation so that cultured cells have the option to polarize between air and basal membrane and this might influence the distribution pattern of MRP1 and MRP2 in NHBEC. Using confocal laser scanning techniques we could show that in cells kept under dry/wet conditions MRP1 was found to be localised to baso-lateral cell regions while MRP2 was localised to all cell regions. Under classical culture conditions MRP1 was not localized to particular membrane regions and MRP2 was found to be an intracellular protein. PMID- 15664272 TI - Functional assays in marine biotoxin detection. AB - The contamination of seafood by algal toxins regularly affects animals living in several areas of the world, and the number of toxic phycotoxins which are being characterized is steadily increasing. The extreme dynamics characterizing the field of algal toxins has stimulated the development of tools to be implemented in the monitoring of contamination of seafood by individual toxin classes. Under these circumstances, functional assays which can encompass the analytical potential of chemical methods and the predictive features of biological tests are sought. A variety of functional assays for the detection of phycotoxins has been developed in the last 20 years, and the analysis of their features reveals that their specificity is related to the hierarchical level of the biological response to the toxin that has been exploited for its detection. Ideally, analytical methods which could allow accurate estimates of the overall toxicity of multiple classes of toxins in a single procedure would provide the best means for the highest standards in consumer protection and the most rational and economical tools in the management of risks posed by phycotoxins in a wider scale. The achievement of a "systemic functional assay for marine biotoxins" does not appear to be at hand, but its inclusion among the foreseeable events is fully justified by the new research tools and approaches which have become available for the high throughput analysis of entire molecular domains at the cellular level. PMID- 15664273 TI - Effect of molsidomine and L-arginine in cyclosporine nephrotoxicity: role of nitric oxide. AB - Cyclosporine A (CsA) is a potent and effective immunosuppressive agent, but its action is frequently accompanied by severe renal toxicity. To determine if the renal alterations are mediated directly by cyclosporine or by secondary homodynamic alterations induced by cyclosporine, we evaluated if L-arginine and a nitric oxide donor, molsidomine could prevent these alterations. Eight groups of rats were employed in this study, group 1 served as control, group 2 rats were treated with CsA (20 mg/kg, s.c. for 21 days), group 3 received CsA along with L arginine (125 mg/kg in drinking water concurrently with CsA), groups 4 and 5 received CsA along with molsidomine (5 and 10 mg/kg, p.o. 24 h before and 21 days concurrently with CsA), group 6 received CsA along with L-arginine (125 mg/l in drinking water concurrently with CsA) and L-NAME (10 mg/kg), groups 7 and 8 received L-NAME (10 mg/kg) along with CsA and molsidomine (5 and 10 mg/kg), respectively. Renal function was assessed by measuring serum creatinine, blood urea nitrogen, creatinine and urea clearance. Tissue and urine nitrite and nitrate levels were measured to estimate the total nitric oxide levels. The renal oxidative stress was measured by renal malondialdehyde levels, reduced glutathione levels and by enzymatic activity of catalase and superoxide dismutase. Renal morphological alterations were assessed by histopathological examination. CsA administration for 21 days resulted in a marked renal oxidative stress, significantly deranged the renal functions as well as renal morphology. Treatment with L-arginine as well as with molsidomine significantly improved the renal dysfunction; tissue and urine total nitric oxide levels, renal oxidative stress and prevented the alterations in renal morphology. This protection against CsA nephrotoxicity was attenuated by treatment with L-NAME, clearly indicating that NO plays a pivotal role in renoprotective effect of L-arginine and molsidomine against cyclosporine nephrotoxicity. PMID- 15664274 TI - Effect of N'-nitrosonornicotine (NNN) on murine palatal fusion in vitro. AB - Maternal smoking has been linked to an increased risk for orofacial clefts. N' nitrosonornicotine (NNN) is one of the tobacco-specific nitrosamines that has been shown to be linked to the deleterious effects of tobacco and could be linked to the formation of cleft palate birth defects. The effect of NNN on palatal fusion was examined using an in vitro organ culture model of palatal development. The organ cultures were exposed to NNN (0.01, 0.1, 1, 10 and 100 mM) and the effects on palatal development characterized at defined points. Palatal fusion was evaluated at embryonic day 13 (E13)+72 h by characterizing the remaining medial edge epithelium (MEE) and determining the extent of fusion compared to controls. The NNN-treated group (1 mM) had more MEE remaining in the palatal midline than the untreated group at E13+72 h (P<0.05). Changes in cell proliferation in the MEE resulting from NNN exposure were examined by BrdU incorporation in replicating DNA. Changes in the pattern of MEE cell death were examined by TUNEL. BrdU incorporation and TUNEL staining showed that the NNN (1 mM)-treated palates had more MEE cell proliferation and less apoptosis than the untreated-palates at E13+24 h (P<0.05). The mechanism altered by NNN was further evaluated by characterizations of extracellular signal-regulated kinase (ERK) 1/2, p38 and c-jun amino-terminal kinase (JNK). NNN at 1 mM induced ERK1/2 phosphorylation, but reduced p38 phosphorylation (P<0.05, P<0.01, respectively) in the MEE. The results suggest that NNN inhibited palatal fusion by effects on cell proliferation and MEE cell death. PMID- 15664275 TI - Cytokine mRNA profiles for isocyanates with known and unknown potential to induce respiratory sensitization. AB - Isocyanates are low-molecular-weight chemicals implicated in allergic asthmatic type reactions. Identification of chemicals likely to cause asthma is difficult due to the lack of a validated test method. One hypothesis is that differential cytokine induction (Th1 versus Th2 profiles) in the draining lymph node following dermal application can be used to identify asthmagens and distinguish them from contact allergens. In this study, we compared the cytokine mRNA profiles of six chemicals: toluene diisocyanate (TDI), diphenylmethane-4,4'-diisocyanate (MDI), dicyclohexylmethane-4,4'-diisocyanate (HMDI), isophorone diisocyanate (IPDI), p tolyl(mono)isocyanate (TMI), and meta-tetramethylene xylene diisocyanate (TMXDI). Whereas TDI and MDI are well-known respiratory sensitizers, documentation for HMDI, IPDI, TMI, and TMXDI is limited, but suggests that HMDI and IPDI may have respiratory sensitization potential in humans and TMI and TMXDI do not. Following dermal exposure of BALB/c mice, all six isocyanates induced cytokines characteristic of a Th2 response. Although LLNAs suggested that the doses chosen for the RPA were immunologically equivalent, the isocyanates tested differentiated into two groups, high responders and low responders. However, two of the low responders (TMI and TMXDI) were further tested and induced higher levels of Th2 cytokine message than dinitrochlorobenzene (not an asthmagen). Further study of these chemicals is needed to determine whether the Th2 cytokine responses observed for these low responders is predictive of asthmagenic potential or represents an insufficient signal. PMID- 15664276 TI - Glutathione metabolism is impaired in vitro by thallium(III) hydroxide. AB - The possibility that Tl(OH)3, the main Tl3+ specie present in water solutions, could interfere with the normal functioning of the glutathione-dependent antioxidant defense system was investigated. For this purpose, we used both the purified components of this system and rat brain cytosolic fractions. Tl(OH)3 (1 25 microM) significantly decreased the content of reduced glutathione (GSH) in both experimental systems, caused by GSH oxidation. In the same range of concentrations Tl(OH)3 inhibited glutathione peroxidase (GPx) activity in both models, using cumene hydroperoxide as the substrate. No alterations in the capacity of GPx activity to metabolize H2O2 were observed. Both in purified GR as well as in the cytosolic fraction, Tl(OH)3 (1-5 microM) inhibited GR activity, with a partial recovery of the activity at higher concentrations. While Tl(OH)3 inhibited the GR diaphorase activity of purified GR, in a concentration (1-25 microM) dependent manner, this effect was only observed in the cytosolic fractions at the highest concentration assessed (25 microM). Results indicate that, similarly to previous findings for Tl+ and Tl3+, Tl(OH)3 also alters the glutathione-dependent antioxidant defense system. The observed alterations of this important antioxidant protective pathway by the major Tl3+ specie in water solutions could be one mechanism involved in the oxidative stress associated to Tl-intoxication. PMID- 15664277 TI - Dynamic MR imaging for diagnosis of lesions adjacent to pituitary gland. AB - In this study, we evaluated the usefulness of dynamic studies for the preoperative diagnosis of tumor lesions adjacent to the pituitary gland. We examined 13 tumors. We obtained pre- and post-contrast thin slice T1-weighted images of the sellar regions. FSE sequences were used for dynamic study. The capability to distinguish the tumor from the pituitary gland and additional information obtained from the dynamic study were evaluated. Two meningiomas were identified more clearly on the dynamic than on the delayed study. In two cavernous hemangiomas, the peripheral nodular stains gradually increased during the dynamic study. For the preoperative evaluation of lesions adjacent to the pituitary gland, the dynamic study can often identify the lesions more clearly and may provide additional information. PMID- 15664278 TI - Vertebrocarotid collateral in extracranial carotid artery occlusions: digital subtraction angiography findings. AB - The internal and external carotid arteries are usually considered occluded distal to a common carotid artery occlusion but some collateral vessels may provide blood keeping the internal and external carotid arteries patent distal to the occlusion. Most common communication in such a case is diversion of blood from muscular branches of the vertebral artery to occipital branch of the external carotid artery which in turn could maintain blood flow into the internal carotid artery, a condition called carotid steal. We encountered vertebrocarotid anastomoses maintaining the patency of carotid circulation in six patients. Patients were four females and two males, ages ranging from 40 to 67 (mean age: 56) years. Five of the patients had ischemic cerebral symptoms. The origin of the external carotid artery was occluded in two and the whole common carotid artery in the remaining four patients. Two patients had double steal, carotid and subclavian at the same time. There was also severe stenosis or occlusion of at least one other major extracranial cerebral artery in all the cases. This concomitant involvement of the second extracranial cerebral artery was thought to be the main reason for the development of vertebrocarotid collateral. In contrast to most of the previously published reports claiming the inadequacy of angiography when compared with colour Doppler ultrasonography, angiography finely depicted the distal patency of the carotid circulation and all the collaterals in detail in every case. Selective injection of the vertebral artery ipsilateral to the occlusion, is the key to demonstrate distal patency of the carotid circulation in cases of proximal carotid occlusion. Demonstration of patency of the distal circulation is very important because some of the patients might get benefit from a reconstructive surgery. PMID- 15664279 TI - MRI patterns in orbital malignant lymphoma and atypical lymphocytic infiltrates. AB - OBJECTIVE: To evaluate the potential of MRI in differentiating between malignant lymphomas and atypical lymphocytic infiltrates in the orbit. MATERIALS AND METHODS: MRI, clinical and histopathological findings in 30 patients presenting with orbital lymphoproliferative diseases (malignant lymphoma and atypical lymphocytic infiltrates) were evaluated. RESULTS: MRI detected 28 out of 30 (93%) orbital lymphoid tumors. Seven out of eight (87.5%) atypical lymphocytic infiltrates and 21/22 (95.4%) of malignant lymphomas were detected. One conjunctival malignant lymphoma and one conjunctival atypical lymphocytic infiltrate were missed. Only malignant lymphoma lesions were hyperintense compared to the extraocular muscles on precontrast and postcontrast T1-weighted images. The lacrimal duct was involved only with malignant lymphoma. Bilateral disease was more likely to be malignant lymphoma. Intraconal lesions were more likely to be associated with lesions elsewhere. No malignant lymphoma involved the extraconal fat. Only intraconal atypical lymphocytic infiltrates had indistinct margins. CONCLUSION: In the orbit, MRI features alone may not allow clear-cut differentiation of malignant lymphomas from atypical lymphocytic infiltrates. However, certain imaging features increase the likelihood of distinguishing them. MRI may miss conjunctival disease. Both orbits should be imaged when orbital lymphoid disease is a probability. The presence of intraconal disease should prompt a search for lesions elsewhere. The use of a head coil instead of a superficial coil may be advantageous by eliminating coil shine effect and allowing evaluation of both orbits. Comparison of lesion signal to extraocular muscle signal appears to be a better alternative than cerebral gray matter or periorbital fat in differentiating malignant lymphoma from atypical lymphocytic infiltrates. Since imaging findings overlap, histopathological diagnosis is necessary in all cases. PMID- 15664280 TI - How does nose blowing effect the computed tomography of paranasal sinuses in chronic sinusitis? AB - OBJECTIVE: Our aim was to determine whether inward or outward movement of the secretions in the paranasal sinuses due to nose blowing after nasal decongestion has any effect on the paranasal sinus computed tomography (CT) images in patients with sinusitis and to asses whether nose blowing may result in misdiagnosis or overdiagnosis in radiological evaluation of sinusitis. MATERIALS AND METHODS: Twenty-four patients with chronic sinusitis were evaluated in an academic tertiary care hospital and data were collected prospectively. After coronal sinus computed tomography scans were performed at 100 mA setting which was half the value of the standard radiation dose suggested by the manufacturer, topical decongestion was applied to each nostril followed by nose blowing 10 min later. Sinus CT scans were then repeated at the same setting. We evaluated the mucosal thickness of medial, lateral, superior and inferior maxillary and frontal sinus walls and the maximal thickness in anterior ethmoidal cells. The measurements prior to and following nose blowing were compared with Wilcoxon signed ranks test. The obtained images were also staged using Lund-McKay staging system separately and the scores were compared with Student's t-test. RESULTS: We observed a tendency towards reduction in mucosal thickness after nose blowing. There were statistically significant differences between maxillary sinus inferior wall and frontal sinus inferior wall mucosal thickness values prior to and after nose blowing. The difference however was very small, about 0.5 mm in magnitude and Lund-McKay score did not change in any of the patients after nose blowing. CONCLUSION: Nose blowing and topical nasal decongestion does not have any effect on the diagnostic accuracy of sinus CT in chronic sinusitis patients. PMID- 15664281 TI - Multiplanar reformat and volume rendering of a multidetector CT scan for path planning a fluoroscopic procedure on Gasserian ganglion block-a preliminary report. AB - In the treatment of trigeminal neuralgia, Gasserian block under fluoroscopical guidance may be difficult because of anatomic variability, and difficulty in identification of the foramen ovale. We introduce how to use three-dimensional CT in the preprocedural planning. We determine the skull-rotation angle in which the foramen ovale is best visualized, the relationship (distance, angle) between the virtual puncture point and anatomical landmarks, and the distance between the virtual puncture point and the foramen. PMID- 15664282 TI - Comparison of quantitative and visual analysis of Tc-99m MIBI scintimammography for detection of primary breast cancer. AB - Tc-99m MIBI scintimammography (SMM) is known to be a useful diagnostic tool for primary breast cancer. We conducted this study to compare the quantitative and visual analysis for detection of primary breast cancer and also investigated the incremental role of quantitative index of SMM. METHODS: 520 highly suspected breast cancer patients (malignant 370: palpable 232, nonpalpable 138; benign 150: palpable 67, nonpalpable 83) were included in this study. The SMM was performed 10 min after the injection of 750 MBq of Tc-99m MIBI. For visual analysis, three scoring based method was used. The lesions to non-lesion ratios (L/Ns) were calculated. Receiver operating characteristic curve (ROC) analyses were performed to determine the optimal visual grade, to calculate cut-off value of L/N and to investigate whether the L/N could provide incremental value additive to visual analysis. The patients were divided into four groups according to the tumor size (group A: size 5 cm). Also, the patients were divided into two groups according to age (under and over 45 years) to investigate incremental value of quantitative analysis. RESULTS: When visual grade 3 was used as the cut-off grade, the sensitivity and specificity were 82.7 and 78%, respectively. The L/N of malignant breast disease was significantly higher than that of benign (2.00 +/ 1.88 versus 0.60 +/- 0.7, P < 0.01). When L/N of 1.27 was used as the cut-off value, the sensitivity and specificity of SMM were 77.6 and 83.3%, respectively. When the L/N was added to visual grade, the area under curve (AUC) of visual + quantitative (V + Q) analyses was higher than that of visual (V) and quantitative (Q) analyses (AUC 0.874 versus 0.803, P < 0.01). In group A and B, the AUCs of V + Q (0.861, 0.895) were higher than those of V (0.808, 0.781) and Q (0.808, 0.813). In group C, the AUC of V + Q (0.847) was higher than that of Q (0.803, P = 0.041). However, the AUC of V (0.915) was higher than that of V + Q (P = 0.009). In group D, there were no statistical differences between V + Q (0.685) and V (0.570, P = 0.058) and Q (0.620, P = 0.145). The V + Q revealed incremental value in the detection of primary breast cancer in both age groups. Also, in older age patients group (over 45 years), the specificities of Q and V + Q were higher than younger group (under 45 years). For axillary lymph node involvement, the sensitivity, specificity, and accuracy of SMM were 66.9, 70.1, and 68%, respectively. CONCLUSION: From this study, the optimal visual grade for diagnosis of breast cancer was grade 3 and cut-off value of L/N was 1.27. Also, we found that the L/N provides incremental value additive to visual analysis. Especially, when the tumor is small (size 0.75). CONCLUSION: The venous segments distinguish liver areas divided by the left and middle hepatic vein in exactly the same pattern as Couinaud segments and portal vein segments do. However, the comparison of shape and position of venous subsegments showed no correlation with both liver segmental approaches. PMID- 15664289 TI - Accuracy of thin section magnetic resonance using phased-array pelvic coil in predicting the T-staging of rectal cancer. AB - Magnetic resonance (MR) imaging may contribute to staging rectal cancer and inform the decision regarding administration of pre-operative radiotherapy. The accuracy of MR has been debated. The aim of the present study was to determine the accuracy of thin section T2-weighted MR images in rectal cancer patients. MR results were compared with histological assessment of resection specimens. Over a 2-year period, 42 patients were studied. Histological staging was pT2 n = 13, pT3 n = 25 and pT4 n = 4. MR diagnostic accuracy was 74%. MR sensitivity and specificity was 62% and 79% for pT2 lesions, 84% and 59% for pT3 lesions and 50% and 76% for pT4 lesions. Estimation of tumour penetration by thin section MR imaging of rectal cancers using pelvic phased-array coil has moderate diagnostic accuracy. The limitations of MR should be acknowledged when selecting rectal cancer patients for pre-operative radiotherapy. PMID- 15664290 TI - Functional evaluation of the postoperative gastrointestinal tract using kinematic MR imaging: Quantitative assessment of peristaltic activity. AB - The purpose of this study is to demonstrate the feasibility of kinematic MR imaging in visualizing peristaltic activity in the reconstructed gastrointestinal tract with quantitative measurements. Sixteen patients with gastrointestinal reconstruction were studied with kinematic MR imaging using HASTE and/or true FISP. Peristaltic waves were noted in the retrosternal gastric segment after gastric pull through (frequency = 3.0 +/- 0.5 times/min, velocity = 2.6 +/- 0.6 mm/s) and in the residual antrum after cardiectomy (frequency = 3 times/min, velocity = 4.0 mm/s). Peristaltic waves were not apparent in the residual fundus after distal gastrectomy and in the reconstructed jejunum after pancreatoduodenectomy and gastrojejunostomy. PMID- 15664291 TI - Advantages and disadvantages of hysterosonosalpingography in the assessment of the reproductive status of uterine cavity and fallopian tubes. AB - BACKGROUND: Hysterosonosalpingography as a contrast ultrasound method is safer, cheaper and easier to perform than hysterosalpingography in the assessment of the uterine cavity and fallopian tubes. Is it feasible for all patients? Which is the main problem in the evaluation of target structures by ultrasound? METHODS: In a prospective study, 68 patients in the initial stage of the infertility treatment were examined by hysterosonosalpingography using saline NaCl infundibile and Echovist as contrast media. Subsequently, further status of the tubes and uterine cavity was assessed by the "gold standards", laparoscopy and hysteroscopy. RESULTS: Sensitivity and specificity of hysterosonosalpingography using NaCl infundibile for evaluation of the uterine cavity was 100 and 88.8%, respectively. Negative predictive value was 100% and positive predictive value 97%. Sensitivity and specificity of the method for the assessment of the tubal status was 100 and 66%, respectively, negative predictive value was 100% and positive predictive value was 61%. For the assessment of tubal patency using positive contrast Echovist the method has shown 100% sensibility and negative predictive value again but it reached a specificity of 77% and a positive predictive value of 70%. There were no evident complications during or after the procedure. CONCLUSION: Hysterosonosalpingography is useful in making decisions regarding further procedures for the diagnosis and treatment of infertility. Uterine cavity evaluation using saline is the method of choice. Tubal patency can be assessed only under ideal sonographic conditions. The method is feasible for early assessment of the reproductive status of uterine cavity and fallopian tubes as a simple, safe and cheap outpatient method prior to any following invasive procedure or even histerosalpingography. PMID- 15664292 TI - Is hysterosalpingography able to diagnose all uterine malformations correctly? A retrospective study. AB - BACKGROUND: Mullerian duct anomalies are an uncommon but often a treatable cause of infertility. MATERIAL AND METHODS: From January 2002 until November 2003, a total of 705 hysterosalpingographies were realised of which 658 were analysed retrospectively in order to determine the importance of uterine malformations in infertile and sterile patients. In ten cases, the results of hysterosalpingography were compared with hysteroscopy and laparoscopy. RESULTS: The frequency of uterine malformations in infertile and sterile women was 10%. In this population, arcuate uterus (57.6%) was the most common malformation, followed by subseptate uterus (18.2%), uterus bicornis unicollis (10.6%), uterus bicornis bicollis (3.0%), septate uterus (6.1%), unicornuate uterus (3.0%) and unicornuate uterus with double vagina (1.5%). DISCUSSION: Our bibliographic research confirmed that arcuate uterus is the most frequent congenital malformation, but in gestations it does not show a significantly reduced live birth rate. Nevertheless, the septate uterus presents an important decrease of live birth rate that can be improved significantly by doing hysteroscopic metroplasty. As this is a completely different surgical intervention compared to bicornuate uterus, a correct diagnosis is very important. Differentiation of these two malformations is quite difficult by hysterosalpingographic exploration, wherefore an additional exploration by means of endovaginal or three-dimensional ultrasound is recommended. PMID- 15664293 TI - Transabdominal pulse inversion harmonic imaging improves assessment of ovarian morphology in virgin patients with PCOS: comparison with conventional B-mode sonography. AB - OBJECTIVE: In virgin policystic ovary syndrome (PCOS) patients transabdominal sonography is the preferential method of the pelvic examination. The purpose of this study was to determine ovarian morphology by the transabdominal route by pulse inversion harmonic imaging (PIHI) in virgin PCOS patients and to compare the diagnostic image quality with conventional B-mode ultrasonography (CBU). METHODS: Fifty-two ovaries in 26 virgin patients were evaluated by the transabdominal approach. Each ovary was examined using both PIHI and CBU. The sharpness of the follicular cysts walls, degree of internal echo definitions of the follicle cysts and overall ovarian conspicuity was assessed subjectively, using 4 point scoring (0, being worst; 3, being best score). The number of countable follicles, the size of largest and smallest ovarian follicle and ovarian volumes were assessed quantitively by both techniques. The effect of body mass index (BMI) on qualitative and quantitative scoring was evaluated. RESULTS: The sharpness of the cyst wall and internal echo structure was significantly better with PIHI than with CBU (P < 0.001 P < 0.001 and P < 0.001, respectively). PIHI improved overall ovarian conspicuity in 41 (78.8%) of 52 examination. The number of countable follicles was significantly lower with CBU (P < 0.001). The maximum diameter of the largest follicle was larger with PIHI sonography to compared CBU (P < 0.001). Mean ovarian volume was significantly larger with CBU (P < 0.001). When data were analyzed separately according to BMI, number of non diagnostic overall ovarian conspicuity scores with CBU was markedly high in obese patients (88% with CBU versus 3.8% with PIHI). On the other hand, mean number of countable follicles with CBU became much more lower in the obese group (P < 0.001). CONCLUSION: In virgin PCOS patients, when compared to transabdominal CBU, PIHI significantly improved the detection of ovarian follicles, especially in high BMI obese subjects, through increased contrast sensitivity for cystic structures and decreased intrafollicular artifactual echoes. Transabdominal ultrasonographic examination by PIHI mode can contribute to the evaluation of ovaries. PMID- 15664294 TI - Balloon angioplasty of popliteal and crural arteries in elderly with critical chronic limb ischemia. AB - OBJECTIVE: Elderly patients with extensive infrainguinal peripheral vascular disease and critical chronic limb ischemia (CCLI) are poor surgical candidates. Our purpose was to evaluate angiographic and clinical results of popliteal, infrapopliteal, and multi-level disease percutaneous transluminal angioplasty (PTA) in such patients. DESIGN: Retrospective study of angiographic and clinical files in selected group. MATERIALS AND METHODS: Between 1996 and 2002, 38 elderly patients aged 80-94 years old (mean age 83.3) with critical leg ischemia were treated with PTA. All patients were at high surgical risk. 31/38 (81.5%) patients had chronic non-healing wounds, and 14/38 (37%) had multi-level disease of superficial femoral, popliteal and crural arteries. One hundred and two lesions were treated by angioplasty. Immediate angiographic and 1 year clinical results were retrospectively analyzed. RESULTS: The overall procedural success rate was 32/38 (84.2%). There were three major complications (7.9%), but no deaths, and three technical failures, all were of infrapopliteal lesions. After 1 year, 27 patients could be followed, five patients died during the first year of unrelated causes. Twenty-three patients (85.2%), were clinically re-occluded within 1 year, but complete and partial wound healing was achieved in 80% (16/20) and rest pain improvement in 57% (4/7), so that overall limb salvage was 74% (20/27). CONCLUSIONS: Elderly patients with multi-level CCLI have a short patency term following angioplasty of 14.8% after 1 year. Nevertheless, this temporary vascular patency enables wound healing or improvement in 74% of these patients, thus such endovascular interventions are recommended in this age group. PMID- 15664295 TI - Enhancing ultrasound images by morphology filter and eliminating ringing effect. AB - Various medical image compression techniques have been proposed for accelerating image propagation in many applications. JPEG2000 is a new generation technique that can encode near lossless ultrasound images at medium bit-rate with diagnostically acceptable quality. Because the coder of JPEG2000 is based on wavelet transform, the reconstructed image will contain some ringing artifacts. Some de-ringing algorithm must be applied to enhance image quality. This study presents quad-tree decomposition and a set of morphological filters for reducing the ringing artifacts of ultrasound images. Specifically, the presented morphological filters use eight predefined morphological operations, including four structuring elements (SE) that include both dilation and erosion. The proposed voting strategy can be used to select the morphological filter for each block to optimize decoded image quality. Image quality can be enhanced by applying the appropriate morphological filter to each block. Experimental results demonstrate that the proposed technique enhances reconstructed ultrasound image quality compared to JPEG2000 at the same bit-rate in terms of both PSNR and the perceptual results. PMID- 15664296 TI - Analysis of radiological examination request forms in conjunction with justification of X-ray exposures. AB - OBJECTIVES: To evaluate the adequacy of patient data and clinical information transmitted to the radiological departments by the referring clinicians in view of the justification demands concerning the radiological examinations, as those are described in the Radiation Protection Regulations of the European Union Nations. MATERIALS AND METHODS: Request forms for 1708 patients referred to radiology department and 410 referred to the CT department were evaluated concerning the proper completion of the form fields. Furthermore, for CT examinations a more detailed evaluation was carried out, to assess the rationalistic use of CT within the diagnostic algorithm. RESULTS: Not properly completed request forms to both departments were observed, however, the omissions were more prominent for the referrals to the radiology department, where the age and the probable clinical diagnosis was not given in 81.5 and 46% of the forms, respectively. Furthermore, the emergency indication was less cautiously used compared to the CT requests. For the CT department 33.3% of requests were justified and the final diagnosis was included in the referring doctor's probable clinical diagnosis. U/S was the examination of choice in 51% of the cases where an examination should have been performed before CT and in 70.6% of the cases where an alternative examination was proposed. CONCLUSIONS: The inadequate transmission of clinical information and the large number of not fully justified CT requests observed in this study are typical examples of the various problems that radiologists have to face in order to apply the justification directives. PMID- 15664297 TI - Abdominal virtual ultrasonographic images reconstructed by multi-detector row helical computed tomography. AB - BACKGROUND: Three-dimensional (3D) images can be generated using thin sections from multi-detector row computed tomography (CT) and computer software, simulating images obtained using conventional ultrasonography (US). This software allows easy diagnosis of abdominal lesions and subsequent treatment of focal liver lesions such as hepatocellular carcinoma (HCC). The present study used newly developed virtual US software for diagnose and treatment of hepatobiliary disease. METHODS: The software was used to create virtual US images in 10 subjects. Radiofrequency ablation (RFA) was performed by virtual US in seven patients with HCC. RESULTS: Slices were easily reconstructed from various angles, and each slice was continuously animated as with conventional US in all subjects. Moreover, when seven patients with HCC were examined using virtual US, HCC nodules were visualized and could be treated with RFA. CONCLUSIONS: Virtual US should prove useful for visualization of HCC nodules that cannot be seen under conventional US. Virtual US is a useful tool for US-guided treatment of HCC. PMID- 15664298 TI - Content and composition of fatty acids in normal and inflamed gingival tissues. AB - The balance of essential fatty acid is important for good health and normal development. Essential fatty acids (EFA) are the precursors of prostaglandins (PGs), thromboxanes and leukotrienes (LT). The aim of this clinical study was to determine the total fatty acid level of polyunsaturated fatty acids (PUFA), monounsaturated fatty acids (MUFA), saturated fatty acids (SFA) and each fatty acids level of inflamed and normal gingival tissues. Twenty-seven subjects were included the present study. Nineteen samples of inflamed human gingival tissue (nine of fibrous hyperplasia (FH), ten of peripheral giant cell granuloma (PGCG) and eight samples of normal human gingival tissue were analyzed. The characteristics of inflammation were assessed histologically. Variance analyses were performed to assess the differences among tissues. The total cellular fatty acid profiles of the tissues in inflamed human gingival tissue and in eight samples of normal human gingival tissue were determined by gas chromatography using Sherlock microbial identification system (MIS) software (Microbial ID, Newark, DE, USA) with a database of FAME profiles for eukary. PUFAs, MUFAs, and SFAs were quantified by Sherlock microbial identification system (MIS) or database gas chromatography (DGC). There were statistically significant differences between the concentrations in inflamed (FH, PGCG) and healthy gingival tissues for PUFA and MUFA (P<0.001, P<0,011, respectively). There were statistically significant differences among the concentrations in FH, PGCG, and healthy gingival tissues for SFA (P<0.0001). Arachidonic acid, docosahexaenoic acid, linoleic acid were increased in inflamed tissue. The results of this study showed that unsaturated fatty acids (PUFA and MUFA) significantly increased in inflamed gingival tissues. PMID- 15664299 TI - Dietary red palm oil improves reperfusion cardiac function in the isolated perfused rat heart of animals fed a high cholesterol diet. AB - It has been shown that dietary red palm oil (RPO) supplementation improved reperfusion function. However, no exact protective cellular mechanisms have been established. Our aim was to search for a possible cellular mechanism and a role for fatty acids. Rats were fed a standard rat chow, plus cholesterol and/or RPO supplementation for 6 weeks. Functional recovery, myocardial phospholipid and cAMP/cGMP levels were determined in isolated rat hearts subjected to 25 min of normothermic total global ischaemia. Dietary RPO in the presence of cholesterol improved aortic output (AO) recovery (63.2+/-3.06%, P<0.05) vs. cholesterol only (36.5+/-6.2%). The improved functional recovery in hearts supplemented with RPO vs. control was preceded by an elevation in the cGMP levels early in ischaemia (RPO 132.9+/-36.3% vs. control 42.7+/-24.4%, P<0.05). Concurrently, cAMP levels decreased (RPO -8.3+/-6.9% vs. control 19.9+/-7.7%, P<0.05). Our data suggest that dietary RPO-supplementation improved reperfusion AO through mechanisms that may include activation of the NO-cGMP and inhibition of the cAMP pathway. PMID- 15664300 TI - The effect of conjugated linoleic acid on transepithelial calcium transport and mediators of paracellular permeability in human intestinal-like Caco-2 cells. AB - Conjugated linoleic acid (CLA) increases paracellular permeability across human intestinal-like Caco-2 cell monolayers, which transport Ca predominantly by the transcellular route. In vivo, however, paracellular Ca transport is the predominant route of Ca transport. Therefore, the objective of this study was to investigate the effect of CLA on transepithelial Ca transport in Caco-2 cells transporting Ca predominantly by the paracellular route. Cells were seeded onto permeable transport membranes and allowed to differentiate, over 14 d, into intestinal-like cell monolayers. Monolayers (n=9/treatment) were exposed to 0 (control) or 80 microM- 18:2, -cis-9, trans-11 CLA or -trans-10, cis-12 CLA for 14 d prior to Ca transport studies. Overall transepithelial Ca transport as well as transcellular and parcellular Ca transport was significantly increased (P<0.001) by exposure of Caco-2 cells to both isomers of CLA, an effect which appeared to be related to altered localization of zona occludens 1 (a tight junction protein). PMID- 15664301 TI - Long-chain polyunsaturated fatty acids, endothelial lipase and atherosclerosis. AB - Endothelial lipase (EL), a new member of the lipase gene family, was recently cloned and has been shown to have a significant role in modulating the concentrations of plasma high-density lipoprotein levels (HDL). EL is closely related to lipoprotein and hepatic lipases both in structure and function. It is primarily synthesized by endothelial cells, functions at the cell surface, and shows phospholipase A1 activity. Overexpression of EL decreases HDL cholesterol levels whereas blocking its action increases concentrations of HDL cholesterol. Pro-inflammatory cytokines suppress plasma HDL cholesterol concentrations by enhancing the activity of EL. On the other hand, physical exercise and fish oil (a rich source of eicosapentaenoic acid and docosahexaenoic acid) suppress the activity of EL and this, in turn, enhances the plasma concentrations of HDL cholesterol. Thus, EL plays a critical role in the regulation of plasma HDL cholesterol concentrations and thus modulates the development and progression of atherosclerosis. The expression and actions of EL in specific endothelial cells determines the initiation and progression of atherosclerosis locally explaining the patchy nature of atheroma seen, especially, in coronary arteries. Both HDL cholesterol and EPA and DHA enhance endothelial nitric oxide (eNO) and prostacyclin (PGI2) synthesis, which are known to prevent atherosclerosis. On the other hand, pro-inflammatory cytokines augment free radical generation, which are known to inactivate eNO and PGI2. Thus, interactions between EL, pro- and anti inflammatory cytokines, polyunsaturated fatty acids, and the ability of endothelial cells to generate NO and PGI2 and neutralize the actions of free radicals may play a critical role in atherosclerosis. PMID- 15664303 TI - Effects of soy oil on murine salivary tumorigenesis. AB - Dietary fat influences dimethylbenzanthracene (DMBA)-induced tumorigenesis of several organs, including the salivary glands. There is not enough evidence to suggest that soy oil could also affect growth of salivary tumors. The main purpose of this work therefore was to study the effects of dietary soy oil on macroscopic parameters of chemically induced murine salivary gland tumors. Eighty BALB/c male mice were assigned to four groups: soy oil (SO), corn oil (CO, control), fish oil (FO) and olein (O). Two weeks later, tumors were induced by 9,10-dimethyl-1,2-benzanthracene (DMBA). At the 13th post-injection week, the animals were sacrificed. In vivo tumor diameter, gland volume (total resected mass), tumor volume (microscopically measured), tumor remission and tumor histopathology were analyzed. The initial in vivo tumor diameter, gland and tumor volume were significantly greater in soy oil than in fish oil group. 26.7% of animals on the soy oil diet showed tumor remission. Sarcomas were more often found in the SO group, carcinomas in FO and the mixed-type tumors both in SO and CO groups. This study shows that the soy oil treatment resulted in larger tumors, some of which later became undetectable. It is necessary to further investigate these divergent results. PMID- 15664302 TI - Pretreatment tumor prostaglandin E2 concentration and cyclooxygenase-2 expression are not associated with the response of canine naturally occurring invasive urinary bladder cancer to cyclooxygenase inhibitor therapy. AB - The purpose of this study was to determine the extent to which pretreatment prostaglandin E2 (PGE2) concentration and cyclooxygenase-2 (cox-2) expression could be used to predict the antitumor activity of cox inhibitor treatment in naturally occurring canine transitional cell carcinoma of the urinary bladder (TCC). Snap frozen tissues (to measure PGE2) and formalin-fixed TCC samples (for cox-2 immunohistochemistry) were obtained by cystoscopy or surgery. Complete tumor staging was performed before and after one month of treatment with the cox inhibitor, piroxicam (0.3 mg/kg q24 h po). The pretreatment PGE2 concentration ranged from 57 to 1624 ng/g of TCC tissue; n=18 dogs). Cox-2 immunoreactivity was observed in all TCC samples. There was no association between PGE2 concentration, cox-2 expression, and change in tumor volume with piroxicam treatment. In conclusion, cox-2 expression or PGE2 concentration alone, or the combination of the two was not useful in predicting response to piroxicam treatment in canine TCC. PMID- 15664304 TI - Effect of Cleome arabica leaf extract, rutin and quercetin on soybean lipoxygenase activity and on generation of inflammatory eicosanoids by human neutrophils. AB - The effects of Cleome arabica leaf extract, rutin and quercetin on soybean lipoxygenase (Lox) activity and on calcium ionophore (A23187)-stimulated generation of the leukotriene B4 and prostaglandin E2 by human neutrophils were examined. The extract (25 microg/ml), rutin (25 microM) and quercetin (25 microM) inhibited LTB4 synthesis at all concentrations of A23187 used. The extract at 1 100 microg/ml and rutin at 1-100 microM inhibited LTB4 generation by neutrophils stimulated with 1 microM A23187 by about 50%. PGE2 production in response to different concentrations of A23187 was affected in a biphasic manner by the extract and rutin. Quercetin at 1-100 microM caused concentration-dependent inhibition of LTB4 and PGE2 production. The extract, rutin and quercetin caused concentration-dependent inhibition of soybean Lox activity. These results indicate that rutin, quercetin and an extract of C. arabica containing these compounds inhibit Lox activity, consequently decreasing LTB4 production. Thus, these compounds or extracts containing them may be beneficial for the treatment of inflammatory conditions, particularly those characterised by excessive leukotriene generation. PMID- 15664305 TI - Lipoprotein-associated PAF-acetylhydrolase activity in subjects with the metabolic syndrome. AB - Plasma- and lipoprotein-associated activity of the platelet activating factor acetylhydrolase (PAF-acetylhydrolase, PAF-AH) plays an important role in inflammation and in atherosclerotic process, which are present in the metabolic syndrome (MS). Paraoxonase 1 (PON1) is an esterase associated with high-density lipoprotein (HDL) which contributes to the anti-atherogenic effects of this lipoprotein. We investigated the activities of both enzymes in 60 patients with MS and 110 age- and sex-matched subjects without it (non-MS group). Plasma PAF-AH activity was higher in the MS compared to the non-MS group, while HDL-PAF-AH and serum PON1 activities were lower in the MS compared to the non-MS group. Univariate regression analysis in the MS group showed that plasma PAF-AH activity was positively associated with systolic blood pressure, whereas HDL-PAF-AH activity was inversely associated with the homeostasis model assessments (HOMA) index. Both associations remained significant in the multivariate regression analysis, suggesting that insulin resistance and systolic hypertension are major determinants for the alterations in plasma and HDL-associated PAF-AH activity among those observed in MS patients. PMID- 15664306 TI - Randomised double-blind placebo-controlled trial of fish oil in the treatment of depression. AB - Converging evidence suggests that omega-3 polyunsaturated fatty acids have aetiological importance in depression. To determine the effect of adding fish oil to existing therapy in participants who were being treated for depression in a community setting, 77 participants were randomly assigned to receive 8 g of either fish or olive oil per day in addition to their existing therapy. Fifty nine (77%) participants completed 12 weeks of treatment. Dietary, biochemical and lifestyle factors were measured throughout the study. Mood was assessed using the Short Form Hamilton Depression Rating Scale (HDRS-SF) and the Beck Depression Inventory II. Sample size calculations were based on the HDRS-SF. Intention-to treat and per protocol analyses were carried out using residual maximum likelihood. There was no evidence that fish oil improved mood when compared to the placebo oil, despite an increase in circulating omega-3 polyunsaturated fatty acids. However, mood improved significantly in both groups within the first 2 weeks of the study (P<0.001) and this improvement was sustained throughout. In conclusion, fish oil was no more effective than the control as an add-on therapy for depression in this setting. PMID- 15664307 TI - Effect of resveratrol and beta-sitosterol in combination on reactive oxygen species and prostaglandin release by PC-3 cells. AB - The objective of this project was to identify some possible mechanisms by which two common phytochemicals, resveratrol and beta-sitosterol, inhibit the growth of human prostate cancer PC-3 cells. These mechanisms include the effect of the phytochemicals on apoptosis, cell cycle progression, prostaglandin synthesis and the production of reactive oxygen species (ROS). Prostaglandins have been known to play a role in regulating cell growth and apoptosis. PC-3 cells were supplemented with 50 microM resveratrol or 16 microM beta-sitosterol alone or in combination for up to 5 days. Phytochemical supplementation resulted in inhibition in cell growth. beta-Sitosterol was more potent than resveratrol and the combination of the two resulted in greater inhibition than supplementation with either alone. Long-term supplementation with resveratrol or beta-sitosterol elevated basal prostaglandin release but beta-sitosterol was much more potent than resveratrol in this regard. beta-Sitosterol was more effective than resveratrol in inducing apoptosis and the combination had an intermediate effect after 1 day of supplementation. Cells supplemented with resveratrol were arrested at the G1 phase and at the G2/M phase in the case of beta-sitosterol while the combination resulted in cell arrest at the two phases of the cell cycle. beta Sitosterol increased ROS production while resveratrol decreased ROS production. The combination of the two phytochemicals resulted in an intermediate level of ROS. The observed changes in prostaglandin levels and ROS production by these two phytochemicals may suggest their mediation in the growth inhibition. The reduction in ROS level and increase by resveratrol supplementation in PC-3 cells reflects the antioxidant properties of resveratrol. It was concluded that these phytochemicals may induce the inhibition of tumor growth by stimulating apoptosis and arresting cells at different locations in the cell cycle and the mechanism may involve alterations in ROS and prostaglandin production. PMID- 15664308 TI - Motility, ATP levels and metabolic enzyme activity of sperm from bluegill (Lepomis macrochirus). AB - Male bluegill displays one of two life history tactics. Some males (termed "parentals") delay reproduction until ca. 7 years of age, at which time they build nests and actively courts females. Others mature precociously (sneakers) and obtain fertilizations by cuckolding parental males. In the current study, we studied the relations among sperm motility, ATP levels, and metabolic enzyme activity in parental and sneaker bluegill. In both reproductive tactics, sperm swimming speed and ATP levels declined in parallel over the first 60 s of motility. Although sneaker sperm initially had higher ATP levels than parental sperm, by approximately 30 s postactivation, no differences existed between tactics. No differences were noted between tactics in swimming speed, percent motility, or the activities of key metabolic enzymes, although sperm from parentals had a higher ratio of creatine phosphokinase (CPK) to citrate synthase (CS). In both tactics, with increasing CPK and CS activity, sperm ATP levels increased at 20 s postactivation, suggesting that capacities for phosphocreatine hydrolysis and aerobic metabolism may influence interindividual variation in rates of ATP depletion. Nonetheless, there was no relation between sperm ATP levels and either swimming speed or percent of sperm that were motile. This suggests that interindividual variation in ATP levels may not be the primary determinant of variation in sperm swimming performance in bluegill. PMID- 15664309 TI - Physiological and morphological characteristics of the rhythmic contractions of the amnion in veiled chameleon (Chamaeleo calyptratus) embryogenesis. AB - A morpho-functional study of the veiled chameleon (Chamaeleo calyptratus) amnion was performed as part of a general comparative study of amnion rhythmic contractions (ARC) in reptile and bird embryogenesis. Eggs incubated at 27.5 degrees C were used at different developmental stages from day 80 (D80) to D184 for the recording of ARC using a force transducer. Slow ARC, about 1 min in duration, were revealed from D88 (stage 31) to the near-hatching stages (incubation time was 183-198 days). The frequency and amplitude of slow ARC increased significantly towards the end of incubation. This ARC enhancement correlated with the differentiation of smooth muscle elements in the amniotic membrane from single spindle-shaped cells to complex "star-like" structures and with the development of the actin fibers, revealed by phalloidin, in the amniotic muscle layer. Short-term changes in temperature influenced ARC and heart rate (HR). Cooling to 25 degrees C from the control (27.5 degrees C) resulted in a significant decrease in both ARC frequency and HR. Heating to 30 degrees C significantly increased the embryonic HR, but not ARC frequency. Within the temperature range from 25 to 30 degrees C the temperature effect on ARC frequency and HR was reversible. PMID- 15664310 TI - Factorial effects of salinity, dietary carbohydrate and moult cycle on digestive carbohydrases and hexokinases in Litopenaeus vannamei (Boone, 1931). AB - Litopenaeus vannamei were reared in close cycle over seven generations and tested for their capacity to digest starch and to metabolise glucose at different stages of the moulting cycle. After acclimation with 42.3% of carbohydrates (HCBH) or 2.3% carbohydrates (LCBH) diets and at high salinity (40 g kg(-1)) or low salinity (15 g kg(-1)), shrimp were sampled and hepatopancreas (HP) were stored. Total soluble protein in HP was affected by the interaction between salinity and moult stages (p<0.05). Specific activity of alpha-amylase ranged from 44 to 241 U mg protein(-1) and a significant interaction between salinity and moult stages was observed (p<0.05), resulting in highest values at stage C for low salinity (mean value 196.4 U mg protein(-1)), and at D0 in high salinity (mean value 175.7 U mg protein(-1)). Specific activity of alpha-glucosidase ranged between 0.09 and 0.63 U mg protein(-1), an interaction between dietary CBH and salinity was observed for the alpha-glucosidase (p<0.05) and highest mean value was found in low salinity-LCBH diet treatment (0.329 U mg protein(-1)). Hexokinase specific activity (range 9-113 mU mg protein(-1)) showed no significant differences when measured at 5 mM glucose (p>0.05). Total hexokinase specific activity (range 17 215 mU mg protein(-1)) showed a significant interaction between dietary CBH and salinity (p<0.05) with highest value (mean value 78.5 mU mg protein(-1)) found in HCBH-high salinity treatment, whereas in the other treatments the activity was not significantly different (mean value 35.93 mU mg protein(-1)). A synergistic effect of dietary CBH, salinity and moult stages over hexokinase IV-like specific activity was also observed (p<0.05). As result of this interaction, the highest value (135.5+/-81 mU mg protein(-1)) was observed in HCBH, high salinity at D0 moult stage. Digestive enzymes activity is enhanced in the presence of high starch diet (HCBH) and hexokinase can be induced at certain moulting stages under the influence of blood glucose level. Perspectives are opened to add more carbohydrates in a growing diet, exemplifying the potential approach for less polluting feed. PMID- 15664311 TI - Physiological measurements from native and transplanted mussel (Mytilus galloprovincialis) in the canals of Venice. Survival in air and condition index. AB - The Venice Lagoon has been the focus of many environmental studies due to its heavy contaminant load derived from a variety of sources, including industrial activity, oil tanker traffic, and waste runoff from the mainland. In recent years, there has been increasing concern about water quality in the urban areas of Venice related to the discharge of untreated sewage directly into canals, adding to the pollutant load already existing in these areas. One way of gauging the impact of these chemicals is monitoring the local fauna. In the search for good indicators of water quality in the Venice urban area, two physiological indices for mussels-survival in air and condition index-have been evaluated. In 2002, a seasonal study was undertaken; mussels (Mytilus galloprovincialis) were collected in three sites located in the canals of the city and a clean reference site. At the same time, two transplantation experiments were performed, deploying farmed mussels for 1,2, 4, and 12 weeks in the historic centre of Venice and also at a reference site. Data from survival in air test and the condition index of native and transplanted mussels are reported. The results suggest an impact of the complex mixture of pollutants on mussel health status. Both native mussels and those transplanted to the urban area showed reduced survivability in air and decreased condition index values, indicating a less healthy status in animals from the canals of the city. Data are discussed in relation to pollutant bioaccumulation. PMID- 15664312 TI - Flight muscle enzyme activities do not differ between pelagic and near-shore foraging seabird species. AB - Common terns (Sterna hirundo), sooty terns (S. fuscata) and brown noddies (Anous stolidus) are phylogenetically related seabirds that differ in field activity levels and daily energy expenditure. To test whether muscle metabolic capacities co-evolve with activity levels and energy expenditure, we collected pectoral muscle biopsies from members of each species, and measured the activities of key enzymes in oxidative metabolism (citrate synthase, CS), anaerobic metabolism (lactate dehydrogenase, LDH), glycolysis (pyruvate kinase, PK), fatty acid oxidation (3-hydroxyacyl CoA dehydrogenase) and phosphocreatine hydrolysis (creatine phosphokinase, CPK). We hypothesized that temperate-breeding common terns would have higher enzyme activities than the two tropical species (sooty terns and brown noddies); consistent with the higher activity level of common terns. There were no differences in enzyme activities among adults of the three species. Common tern chicks within 2-3 days of flight had two-fold higher pyruvate kinase activity than adults, suggesting an increased glycolytic capacity in the chicks. Given the lack of difference among species at the enzymatic level, our results support the notion that behavior and whole organism performance can evolve considerably before there are detectable changes in underlying lower-level physiological/biochemical traits. PMID- 15664313 TI - Transfer of methyl-branched hydrocarbons from the parasitoid, Eretmocerus mundus, to silverleaf whitefly nymphs during oviposition. AB - The parasitic wasp Eretmocerus mundus (Hymenoptera: Aphelinidae), a natural enemy of the silverleaf whitefly Bemisia argentifolii (Homoptera: Aleyrodidae), deposits eggs beneath nymphs and not within them. Experiments were designed to establish whether ovipositing E. mundus females leave marking chemicals on nymphs to enable searching females to discriminate parasitized from unparasitized hosts. Cuticular lipids from three experimental treatment groups were characterized: parasitoid-exposed nymphs that had a parasitoid egg between the nymph and leaf; control nymphs not exposed to E. mundus; and a third treatment condition of parasitized nymphs, held for 10 days after wasp exposure. Lipids were solvent extracted from the nymphal cuticles of the various treatment groups and the lipid components were characterized and quantified by gas chromatography and mass spectrometry. Results indicated the presence of quantities of C31 and C33 dimethylalkanes only from parasitoid-exposed groups of nymphs and not in the extracts from control nymphs or the parasitized nymphs after 10-day exposure. Furthermore, the C31 and C33 dimethylalkanes were shown to be major lipid components of the hexane extracts from E. mundus females. Since the lipids were removed from parasitoid-exposed nymphs before interaction with hatched parasitoid larvae, the findings indicated that the dimethylalkanes were transferred onto nymphal cuticles by ovipositing E. mundus females. PMID- 15664315 TI - Collecting baseline corticosterone samples in the field: is under 3 min good enough? AB - Evaluating corticosterone (CORT) responses to stress in free-living vertebrates requires knowing the unstressed titers prior to capture. Based upon laboratory data, the assumption has been that samples collected in less than 3 min of capture will reflect these unstressed concentrations. This assumption was tested for six species using samples collected from 945 individuals at 0-6 min after capture. Samples were from five avian species trapped at multiple times of year and one reptilian species, comprising a total of 14 different data sets for comparisons. For seven of 14 data sets, including five species, there was no significant increase in corticosterone titers within 3 min of capture. In six of the 14 data sets, corticosterone titers increased significantly after 2 min, and in one data set, the increase started at 1.5 min. In all seven of the cases showing an increase before 3 min, however, corticosterone titers from the time of increase to 3 min were significantly lower than titers after 30 min of restraint stress. These results indicate a high degree of confidence for these species that samples collected in less than 2 min reflect unstressed (baseline) concentrations, and that samples collected from 2-3 min also will likely reflect baseline concentrations but at worst are near baseline. PMID- 15664314 TI - Tannin-binding salivary proteins in three captive rhinoceros species. AB - Tannin-binding salivary proteins (TBSP) are considered to be counter-defences acquired in the course of evolution by animals whose natural forage contains such tannins. As tannins mostly occur in browse material but not in grasses, it is assumed that grazers do not have a need for TBSP. Whereas it has been shown in several non-ungulate species that TBSP can be induced by dietary tannins, their presence or absence in ungulates has, so far, been shown to be a species-specific characteristic independent of dietary manipulations. We investigated saliva from three rhinoceros species from zoological gardens fed comparable, conventional zoo diets. As expected, saliva from white rhinoceroses (Ceratotherum simum, grazer) had lower tannin-binding capacities than that from black rhinoceroses (Diceros bicornis, browser). Surprisingly, however, Indian rhinoceroses (Rhinoceros unicornis), commonly regarded as grazers as well, displayed the highest tannin binding capacities of the three species investigated. It is speculated that this discrepancy might be a result of an evolutionarily recent switch to a grass dominated diet in Indian rhinoceroses, and that the black rhinoceros, which is closer related to the white rhinoceros than the Indian species, has evolved an inducible mechanism of TBSP production. In separate trials during which the tannin content of the diets of black rhinoceroses was increased by the addition of either tannic acid or quebracho, the tannin-binding capacity of black rhinoceros saliva was increased to levels within the same range as that of Indian rhinoceroses on the conventional diets. While induction trials in white and Indian rhinoceroses remain to be performed for a full understanding of salivary anti-tannin defence in rhinoceroses, these results are the first report of an induced salivary response to increased dietary tannin levels in an ungulate species. PMID- 15664316 TI - Faecal corticosteroid levels as an indicator of well-being in the tammar wallaby, Macropus eugenii. AB - Faecal corticosteroid levels were measured in five female tammar wallabies, Macropus eugenii, at Macquarie University, NSW, Australia, to assess their reliability as indicators of well-being in this species. Animals were challenged with a change in conditions over the course of approximately 1 week, comprising movement from group yards to isolation in individual yards, in order to impose a disturbance to homeostasis ("stress"). Faecal samples were collected in 24-h intervals during the study period and analysed for corticosteroid concentration. The use of enzyme immunoassay for the measurement of corticosteroids in marsupial faecal pellets was validated. We observed a significant increase in faecal corticosteroids upon isolation and movement. Faecal corticosteroids remained above initial levels in all five animals throughout the study period, suggesting that faecal corticosteroid concentrations may be a useful indicator of a change in animal well-being. Faecal corticosteroid levels did not correlate with serum cortisol levels, implying that the use of noninvasive methods in a representative marsupial, the tammar wallaby, has the potential to provide information that is not readily apparent using blood-based protocols. Faecal corticosteroid analysis therefore has the potential for application in monitoring the well-being of captive and managed marsupial populations, as part of an integrated system of measures of animal health and well-being. PMID- 15664317 TI - The hearing abilities of the prawn Palaemon serratus. AB - The mechanism of sound reception and the hearing abilities of the prawn (Palaemon serratus) have been studied using a combination of anatomical, electron microscopic and electrophysiological approaches, revealing that P. serratus is responsive to sounds ranging in frequency from 100 to 3000 Hz. It is the first time that the Auditory Brainstem Response (ABR) recording technique has been used on invertebrates, and the acquisition of hearing ability data from the present study adds valuable information to the inclusion of an entire sub-phylum of animals when assessing the potential impact of anthropogenic underwater sounds on marine organisms. Auditory evoked potentials were acquired from P. serratus, using two subcutaneous electrodes positioned in the carapace close to the supraesophageal ganglion and the statocyst (a small gravistatic organ located below the eyestalk on the peduncle of the bilateral antennules). The morphology of the statocyst receptors and the otic nerve pathways to the brain have also been studied, and reveal that P. serratus possesses an array of sensory hairs projecting from the floor of the statocyst into a mass of sand granules embedded in a gelatinous substance. It is the purpose of this work to show that the statocyst is responsive to sounds propagated through water from an air mounted transducer. The fundamental measure of the hearing ability of any organism possessing the appropriate receptor mechanism is its audiogram, which presents the lowest level of sound that the species can hear as a function of frequency. The statocyst of P. serratus is shown here to be sensitive to the motion of water particles displaced by low-frequency sounds ranging from 100 Hz up to 3000 Hz, with a hearing acuity similar to that of a generalist fish. Also, recorded neural waveforms were found to be similar in both amplitude and shape to those acquired from fish and higher vertebrates, when stimulated with low-frequency sound, and complete ablation of the electrophysiological response was achieved by removal of the statocyst. PMID- 15664318 TI - Growth rate and thermoregulation in reared king quails (Coturnix chinensis). AB - Growth rate was investigated in king quails between 1st and 60th day of life. Gompertz growth constants were 0.075 in males and 0.056 in females. Colonic temperature (Tb) was measured in quails divided into four age groups (1-3, 7-10, 16-19, and 44-59 days old) in ambient temperatures set separately for each group. Metabolic rate was measured only in 44-59-day-old birds. The mean value of the thermoneutral body temperature (Tb at TNZ) in the active phase in the youngest quails was 39.0 degrees C. In 44-59-day-old quails, the resting metabolic rate in the thermoneutral zone (RMR at TNZ) was on average 9.44 mW g(-1) (1.66 cm3 O2 g( 1)h(-1)), without sex-specific differences. No such differences were found in this age group neither in Tb at TNZ, nor in minimal thermal conductance (Cmin). However, differences were found in the rate of metabolic heat production below the thermoneutral zone, even when mass-independent units were used. The maximum metabolic rate (Mmax) in 2-month-old males was 34.08 mW g(-1) (5.98 cm3 O2 g( 1)h(-1)), while in females 29.73 mW g(-1) (5.21 cm3 O2 g(-1)h(-1)). Heat-stressed 44-59-day-old quails elevated their Tb to as much as an average 44.1 degrees C in Ta of about 45 degrees C. The obtained growth model and a gradual development of the body temperature regulation mechanism in king quails followed the known strategy of development, typical for precocial birds. The sexual size dimorphism in the studied quails did not result in differences in thermoregulation parameters between the sexes, except for the rate of metabolic rate below thermoneutral zone. PMID- 15664319 TI - Hyperthyroidism changes nociceptive response and ecto-nucleotidase activities in synaptosomes from spinal cord of rats in different phases of development. AB - Changes in transport, receptors and production of extracellular adenosine have been observed after induction of hyperthyroidism. Adenosine is associated with inhibitory actions such as reduction in release of excitatory neurotransmitters and antinociception at spinal site. In contrast, ATP acts as an excitatory neurotransmitter and produces pronociceptive actions. ATP may be completely hydrolyzed to adenosine by an enzyme chain constituted by an ATP diphosphohydrolase and an ecto-5'-nucleotidase, as previously described in the spinal cord. Thus, we now investigated the effects of the hyperthyroidism on adenine nucleotide hydrolysis in the spinal cord and verified the nociceptive response in this pathology during different phases of development. Hyperthyroidism was induced in male Wistar rats, aged 5, 60 and 330 days by daily intraperitoneal injections of L-thyroxine (T4) for 14 days. Nociception was assessed with a tail-flick apparatus. Rats starting the treatment aged 5 days demonstrated a significant increase in ADP and AMP hydrolysis and increased tail flick latency (TFL). In contrast, in the spinal cord from hyperthyroid rats aged 60 and 330 days old, the hydrolysis of ATP, ADP and AMP were significantly decreased. Accordingly, the tail-flick latency was decreased, indicating a hyperalgesic response. These results suggest the involvement of ecto nucleotidases in the control of the hyperthyroidism-induced nociceptive response in rats at distinct developmental stages. PMID- 15664320 TI - Improving estimates of trophic shift in Nile tilapia, Oreochromis niloticus (L.), using measurements of lipogenic enzyme activities in the liver. AB - To test whether the measurement of selected enzyme activities could be used to estimate more precisely the trophic shift of C isotopes, Nile tilapia (Oreochromis niloticus) were fed semi-synthetic diets differing in their lipid contents (1.7%, 5.0%, 10.8% and 20.0%). The diets were formulated to contain the same amount of nitrogen and metabolizable energy and were made from casein, wheat starch, corn germ oil supplemented with vitamins, minerals and L-arginine. The influence of the different diets on the activity of two lipogenic enzymes, ATP citrate lyase and malic enzyme, on delta13C values in the whole fish, the liver and their correlation was investigated. There was a strong positive correlation between delta13C values in the lipids of whole fish and those of their livers. The activities of lipogenic enzymes increased significantly with increasing trophic shift of C isotopes (Deltadelta13Cdiet-fish values) in the lipids. If the relationship between trophic shift and enzyme activity can be confirmed in situations where feed quantity and quality are not known, the determination of enzyme activities would enable better estimates of the trophic shift to be made thus significantly improving back-calculation of diets from stable isotope data. PMID- 15664322 TI - Phenotypic flexibility in the intestinal enzymes of the African clawed frog Xenopus laevis. AB - The intestinal plasticity of digestive enzymes of amphibian species is poorly known. The goal of this study was to characterize digestive enzyme profiles along the small intestine of adult frogs, Xenopus laevis, in response to an experimental diet. We acclimated adult X. laevis for 30 days either to carbohydrate-rich or protein-rich diets, and determined the morphology and digestive enzymes of the small intestine. We found a significant difference of aminopeptidase-N activity between carbohydrate-rich and protein-rich acclimated animals. We also found a little variation in the expression of maltase activity, which contrast with the proposed hypothesis about the existence of digestive tradeoff in vertebrates. This finding supports the adaptive modulation hypothesis and suggests that caution is called for when analyzing physiological data regarding assumed discrete trophic category of species. PMID- 15664321 TI - Biological and enzymatic activities of Micrurus sp. (Coral) snake venoms. AB - The venoms of Micrurus lemniscatus carvalhoi, Micrurus frontalis frontalis, Micrurus surinamensis surinamensis and Micrurus nigrocinctus nigrocinctus were assayed for biological activities. Although showing similar liposome disrupting and myotoxic activities, M. frontalis frontalis and M. nigrocinctus nigrocinctus displayed higher anticoagulant and phospholipase A2 (PLA2) activities. The latter induced a higher edema response within 30 min. Both venoms were the most toxic as well. In the isolated chick biventer cervicis preparation, M. lemniscatus carvalhoi venom blocked the indirectly elicited twitch-tension response (85+/ 0.6% inhibition after a 15 min incubation at 5 microg of venom/mL) and the response to acetylcholine (ACh; 55 or 110 microM), without affecting the response to KCl (13.4 mM). In mouse phrenic nerve-diaphragm preparation, the venom (5 microg/mL) produced a complete inhibition of the indirectly elicited contractile response after 50 min incubation and did not affect the contractions elicited by direct stimulation. M. lemniscatus carvalhoi inhibited 3H-L-glutamate uptake in brain synaptosomes in a Ca2+-, but not time, dependent manner. The replacement of Ca2+ by Sr2+ and ethylene glycol-bis(beta-aminoethyl ether) (EGTA), or alkylation of the venom with p-bromophenacyl bromide (BPB), inhibited 3H-L-glutamate uptake. M. lemniscatus carvalhoi venom cross-reacted with postsynaptic alpha-neurotoxins short-chain (antineurotoxin-II) and long-chain (antibungarotoxin) antibodies. It also cross-reacted with antimyotoxic PLA2 antibodies from M. nigrocinctus nigrocinctus (antinigroxin). Our results point to the need of catalytic activity for these venoms to exert their neurotoxic activity efficiently and to their components as attractive tools for the study of molecular targets on cell membranes. PMID- 15664323 TI - Intraspecific differences in thermal tolerance of the diamondback watersnake (Nerodia rhombifer): effects of ontogeny, latitude, and sex. AB - Ontogenetic shifts in microhabitat use are widespread among taxa and can result in drastic shifts in thermal habitat among age classes. Likewise, geographic variation in climate along latitudinal gradients can cause differences in thermal environments among populations of a species. Using a common garden design, we examined four populations of a single species of semi-aquatic snake, Nerodia rhombifer, to determine whether ontogenetic shifts in habitat use (and/or body size) and latitudinal differences in ambient temperature have resulted in evolutionary changes in thermal tolerance. We found ontogenetic differences in thermal tolerance for all populations, with neonates tolerating temperatures 2 degrees C higher than adults, a pattern that is consistent with ontogenetic shifts in body size and microhabitat use in this species. There were differences in thermal tolerance among latitudes in neonates, suggesting genetic differences among populations, but adults showed no latitudinal differences. In combination, the increased thermal tolerance of neonates and the age-specific response to latitude suggest individuals may be most sensitive to selection on thermal tolerance as neonates. Although latitudinal differences exist in neonates, their tolerances were not ranked according to latitude, suggesting the effects of some other local factor (e.g., microclimate) may be important. Lastly, among neonates, females tolerate higher temperatures than males. PMID- 15664324 TI - Influence of food type on specific dynamic action of the Chinese skink Eumeces chinensis. AB - We used the Chinese skink (Eumeces chinensis) as an experimental model to study influence of food type on specific dynamic action (SDA) of feeding. Thirty-three adult males collected from a natural population were divided equally into three (one control and two experimental) groups. We starved all skinks at 30 degrees C for 3 days and then provided the experimental skinks with a single meal consisting of either mealworms or meat [the flesh of the bullfrog (Rana catesbeiana)]. Food ingested by skinks of the two experimental groups differed in lipid content and lean dry mass but not in total dry mass and energy. Defecation following feeding occurred slightly earlier in skinks ingesting mealworms (mean=41.7 h) than in those ingesting meat (mean=47.7 h), but the difference was not significant. Analyses of variance (ANOVAs) with repeated measures showed that temporal variation in oxygen consumption over 72 h after feeding was evident in the experimental skinks but not in the control ones. Oxygen consumption was higher in the experimental skinks than in the control ones during the time interval between 4.5 and 36 h after feeding. The peak metabolic rate was greater but occurred later in skinks ingesting meat than in those ingesting mealworms. The estimated amounts of oxygen consumed by mealworm-fed, meat-fed and unfed skinks at 30 degrees C over 72 h after feeding were 356.5, 393.8 and 295.2 mL, respectively. Our results provide a support for the previous prediction that SDA is affected by types of food ingested by animals as skinks ingesting mealworms and meat differed in the time to reach a peak metabolic rate, the level of the peak metabolic rate and the magnitude of the SDA effect. PMID- 15664325 TI - The effects of fasting and refeeding on biochemical parameters in the red-legged partridge (Alectoris rufa). AB - Every year, about three million farm-reared red-legged partridges are released in Spain for hunting purposes, most of them for restocking where the success is very low. Some factors such as the sudden shift from abundant commercial energy-rich diet to natural food and absence of experience at looking for natural food resources could cause a rapid physical deterioration, being more vulnerable to predation and disease. To determine fasting capacity in red-legged partridges, we fasted 32 birds during 4 days and then we refed them, collecting blood samples and body masses during the experiment. Red-legged partridges entered in the third critic phase of fasting, in which body proteins became the main energetic source, after 48 h of fasting. The fasting period caused an average loss of 17% of their initial body mass and only attained a 93.6% of their initial body mass after 24 days of refeeding. The limited capacity to resist fasting in the red-legged partridge could be one of the reasons for the low success in restocking, mainly in those where neither supplementary feeding nor acclimation period is allowed for birds before they are finally released. PMID- 15664326 TI - Optimisation of the coupled monoclonal antibody density for recombinant hepatitis B virus surface antigen immunopurification. AB - Using immunosorbents based upon cyanogen bromide-Sepharose CL-4B, we have examined different ligand densities in coupling of monoclonal antibody (MAb) to find the best performance, for recombinant hepatitis B surface antigen (rHBsAg) purification. Three replicates of 5 and 15 cycles of densities ranges: 2.17-2.19, 3.18-3.62, 4.06-4.17, and 5.13-5.40 mg/ml (control); or 1.81-2.47, 3.17-3.41, 4.16-4.28, and 5.16-5.19 mg/ml (control), respectively were evaluated in terms of binding capacity, antigen recovery, ligand leakage and purity of antigen, and compared to the control. Adsorption and antigen recovery of immunosorbents manufactured were not different statistically, eventhough increased 8.08 and 9.90% at a range of 3.17-3.41 mg/ml. At this range, efficiency expressed as productivity and MAb saving was optimal. Ligand leakage and purity of antigen showed similar behaviour among all densities. Aspects related to ligand density in antigen immunoaffinity purification are discussed. PMID- 15664327 TI - Development and application of a multi-target immunoaffinity column for the selective extraction of natural estrogens from pregnant women's urine samples by capillary electrophoresis. AB - In this paper, a methodology for the determination of three naturally occurring estrogens (estradiol, estrone and estriol) in pregnant women's urine has been described. The procedure included immunoaffinity column (IAC) extraction of 4 mL of urine sample and subsequent analysis of the extraction by micellar electrokinetic chromatography (MEKC). A multi-target polyclonal antibody that has high affinity to three estrogens was produced. Then the IAC was developed by coupling polyclonal antibody to CNBr-activated Sepharose 4B. The IAC showed high affinity for these estrogens. Recoveries of three estrogens from human serum matrix were greater than 92% with R.S.D. less than 4.5%. The final elute of urine sample was diluted with running buffer and then quantitated with MEKC. The experimental results demonstrated that IAC was a useful technique for extraction and concentration of estrogens from biological samples. Three estrogens levels in six pregnant women's urine were measured by both the present method and enzyme linked immunoadsorbent assay (ELISA). The results of this method have been found to correlate well with those of ELISA. PMID- 15664328 TI - Confirmation of carbadox and olaquindox metabolites in porcine liver using liquid chromatography-electrospray, tandem mass spectrometry. AB - A method is described for the quantitative determination of quinoxaline-2 carboxylic acid (QCA) and methyl-3-quinoxaline-2-carboxylic acid (MQCA), the metabolites that have been designated as the marker residues for the veterinary drugs, carbadox and olaquindox, respectively, in swine tissue. The method is suitable for use as a confirmatory method under EU National Surveillance Schemes. Porcine liver samples were subjected to protease digestion followed by liquid liquid extraction. Further clean-up was performed by automated solid phase extraction (SPE) and was followed by a final liquid-liquid extraction step. Analysis was performed using a narrow bore column HPLC coupled to electrospray MS/MS, operated in positive ion mode. MS/MS product ions were monitored at m/z 102 and 75 amu for QCA, m/z 145 and 102 amu for MQCA and at m/z 106 and 152 amu for the d(4)-QCA and d(7)-MQCA internal standards, respectively. The method has been validated at 3.0, 10, 50 and 150 microg kg(-1) for both metabolites. The method performance characteristics-the decision limit (CCalpha) and the detection capability (CCbeta) have been determined for QCA at 0.4 and 1.2 microg kg(-1), respectively, and for MQCA at 0.7 and 3.6 microg kg(-1), respectively. PMID- 15664329 TI - High-performance liquid chromatography measurement of hyperforin and its reduced derivatives in rodent plasma. AB - A reverse-phase high-performance liquid chromatography method was developed for the determination of hyperforin and its reduced derivatives octahydrohyperforin and tetrahydrohyperforin in rodent plasma. The procedure includes solid-phase extraction from plasma using the Baker 3cc C8 cartridge, resolution on the Symmetry Shield RP8 column (150 mm x 4.6 mm, i.d. 3.5 microm) and UV absorbance detection at 300 nm. The assay was linear over a wide range, with an overall coefficient of variation less than 10% for all compounds. The precision and accuracy were within acceptable limits and the limit of quantitation was sufficient for studies preliminarily assessing the disposition of tetrahydrohyperforin and octahydrohyperforin in the mouse and rat. PMID- 15664330 TI - Application of solid phase microextraction to the determination of strychnine in blood. AB - A simple and rapid method based on solid phase microextraction (SPME) via direct immersion followed by gas chromatography coupled with electron impact ionization/mass spectrometry (GC/EI-MS) was developed for the determination of strychnine in blood. Papaverine was used as internal standard (I.S.). Two types of fibre coating were tested, 100 microm polydimethylsiloxane and 65 microm Carbowax/Divinylbenzene, the latter giving higher recoveries of the compound. The main factors affecting the SPME process, such as sample dilution (1:10), adsorption and desorption times (20 and 10 min, respectively), carry-over effect (not observed), pH and salt addition (no modifications on pH or salt concentration) were optimized. The procedure was validated in terms of linearity (r(2)=0.9992 for concentrations ranging from 0.10 to 5.00 microg/mL), intra and interday precision (0.93 and 4.62%, respectively at 0.50 microg/mL; 3.33 and 8.06%, respectively at 2.50 microg/mL), sensitivity (6.83 and 8.91 ng/mL for LOD and LOQ, respectively) and extraction recovery (0.54 and 0.39% at 0.50 and 2.50 microg/mL, respectively). The developed procedure was found suitable for forensic investigations and was considered a good alternative to the liquid-liquid extraction methods normally used for the determination of this compound in biological media. PMID- 15664331 TI - Liquid chromatography-electrospray tandem mass spectrometry method for determination of indapamide in serum for single/multiple dose bioequivalence studies of sustained release formulations. AB - Indapamide and internal standard (5-chloro-2-methoxy-N-[2-(4 sulphamoylphenyl)ethyl]benzamide) were isolated from plasma by a single step liquid-liquid extraction in t-butyl methyl ether. The chromatographic separation was achieved on a reversed-phase C(18) monolithic column with a mobile phase consisting in a methanol/aqueous 0.1% formic acid mixture and a flow rate of 0.8 ml/min, in isocratic conditions, within 11 min. Target compounds were transferred in an ion trap analyzer via an atmospheric pressure electrospray interface (AP ESI). The mass analyzer was used in a selected reaction monitoring (SRM) mode, in order to enhance on detection selectivity. Whole method produces quantitation limit for indapamide of 1 ng/ml. Method was successfully applied to assess bioequivalence of two sustained release marketed pharmaceutical formulations of indapamide 1.5 mg coated tablets, carried-out in a single/multiple doses, randomized design. PMID- 15664332 TI - Determination of plasma Vitamin K by high-performance liquid chromatography with fluorescence detection using Vitamin K analogs as internal standards. AB - A HPLC fluorescence determination method for Vitamin K derivatives (Vitamin K(1), phylloquinone, PK and K(2), menaquinones, MK-4 and MK-7) using post-column reduction and internal standards was developed. Selectivity and reproducibility were increased by optimized chromatography conditions and satisfactory precision and accuracy were attained by using synthetic internal standards. After addition of internal standards to plasma samples, lipids were extracted with ethanol and hexane. Chromatography was performed by isocratic reverse phase separation on a C18 column. Vitamin K derivatives were detected at 430 nm with excitation at 320 nm for MK-4 and 240 nm for PK and MK-7. The detection limits for MK-4, PK and MK 7 were 4, 2 and 4 pg, respectively. The recoveries of MK-4, PK and MK-7 were greater than 92% and the inter- and intra-assay R.S.D. values were 5.7-9.2% for MK-4, 4.9-9.6% for PK and 6.3-19.3% for MK-7. The data showed good correlation between proposed method and LC-APCI/MS method for MK-4 (R(2)=0.988), PK (R(2)=0.979) and MK-7 (R(2)=0.986). The method allows the determination of Vitamin K for evaluating their clinical and nutritional status. PMID- 15664333 TI - Determination of carotenoids and all-trans-retinol in fish eggs by liquid chromatography-electrospray ionization-tandem mass spectrometry. AB - A novel method was developed for the combined determination of carotenoids and retinoids in fish eggs, which incorporates prior analyte isolation using liquid liquid partitioning to minimize analyte degradation, and fraction analysis using high-performance liquid chromatography-electrospray (positive)-quadrupole mass spectrometry (LC-ESI(+)-MS; SIM or MRM modes). Eggs from Chinook salmon (Oncorhynchus tshawytscha) were used as the model fish egg matrix. The methodology was assessed and validated for beta-carotene, lutein, zeaxanthin, and beta-cryptoxanthin (molecular ion radicals [M](+)), canthaxanthin and astaxanthin ([M+Na](+) adducts) and all-trans-retinol ([(M+H)-H(2)O](+)). Using replicate egg samples (n=5) spiked with beta-cryptoxanthin and beta-carotene before and after extraction, matrix-sourced ESI(+) enhancement was observed as evidenced by comparable %matrix effect and %process efficiency values for beta-cryptoxanthin and beta-carotene of 114-119%. In aquaculture-raised eggs from adult Chinook salmon astaxanthin, all-trans-retinol, lutein and canthaxanthin were identified and determined at concentrations of 4.12, 1.06, 0.12 and 0.45 microg/g (egg wet weight), respectively. To our knowledge, this is the first report on a method for LC-MS determination of carotenoids and retinoids in a fish egg matrix, and the first carotenoid-specific determination in any fish egg sample. PMID- 15664334 TI - Chromatographic analysis of carbamazepine binding to human serum albumin. AB - In this study, high-performance affinity chromatography was used to characterize the binding of carbamazepine to an immobilized human serum albumin (HSA) column. Frontal analysis was first used to determine the association equilibrium constant and binding capacity for carbamazepine on this column at various temperatures. The non-specific binding of carbamazepine within the column was also considered. The results indicated that carbamazepine had a single binding site on HSA with an association equilibrium constant of 5.3 x 10(3)M(-1) at pH 7.4 and 37 degrees C. This was confirmed through zonal elution self-competition studies. The value of DeltaG for this reaction was -5.35 kcal/mol at 37 degrees C, with an associated change in enthalpy (DeltaH) of -6.45 kcal/mol and a change in entropy (DeltaS) of -3.56 cal/molK. The location of this binding region was examined by competitive zonal elution experiments using probe compounds with known sites on HSA. It was found that carbamazepine had direct competition with l-tryptophan, a probe for the indole-benzodiazepine site of HSA, but allosteric interactions with probes for the warfarin, tamoxifen and digitoxin sites. Changes in the pH, ionic strength, and organic modifier content of the mobile phase were used to identify the predominant forces in the carbamazepine-HSA interaction. PMID- 15664336 TI - Quantitative determination of pravastatin and R-416, its main metabolite in human plasma, by liquid chromatography-tandem mass spectrometry. AB - A quantitative method was developed and validated for rapid and sensitive analysis of pravastatin and R-416, the main metabolite of pravastatin, in human plasma. The analytes were extracted from plasma samples by a solid phase extraction method using a Bond Elut C(8). The method involved the use of liquid chromatography coupled with atmospheric pressure chemical ionization (APCI) and selected reaction monitoring (SRM) mass spectrometry. A pravastatin analog, R 122798, was used as the internal standard (I.S.). Separation of pravastatin, R 416 and the I.S. was accomplished using a reverse-phase column (C(18)). The components eluted were ionized by the APCI source (negative ion) and subsequently detected by a highly selective triple quadrupole mass spectrometer in the SRM mode. Linear standard curves were obtained from 0.1 ng/mL (lower limit of quantification, LLOQ) to 100 ng/mL. The intra-assay precisions (coefficient of variation) for the samples at the LLOQ were 1.8% for pravastatin and 1.6% for R 416. The intra-assay accuracy values were 95.8-107.6% for pravastatin, and 92.6 109.0% for R-416, respectively. Precision and accuracy of quality control (QC) samples were determined at concentrations of 0.5, 10 and 80 ng/mL for all analytes. The intra- and inter-assay precision calculated from QC samples were within 10% for pravastatin and within 11% for R-416. The overall recoveries for pravastatin and R-416 were 75.7-82.1% and 68.6-74.3%, respectively. Pravastatin and R-416 were stable in human plasma for 3 weeks at -20 degrees C in a freezer, up to 6h at room temperature, and up to 48 h at 6 degrees C. This assay method was successfully used to evaluate the pravastatin and R-416 levels in healthy volunteers following oral administration of Mevalotin. PMID- 15664335 TI - Determination of thiamine and its phosphate esters in rat tissues analyzed as thiochromes on a RP-amide C16 column. AB - A new reversed-phase chromatographic method is described for the separation and quantification of thiamine (T), thiamine monophosphate (TMP) and diphosphate (TDP) in rat tissues. Sample extraction with perchloric acid (HClO(4)) was found more suitable than extraction with trichloroacetic acid (TCA), as regards convenience and background fluorescence. Derivatization of thiamine vitamers to thiochromes was optimized and complete separation of TDP and TMP thiochromes was obtained on a RP-amide C16 column in isocratic elution, with T thiochrome eluting in less than 10 min. The precision and the accuracy of the HPLC procedure were assessed: ranging from 0.5 to 7.7% for intra-day and from 2.0 to 9.4% for inter day precision, a recovery average of 101% was determined (range 90-111%). Mean values of recovery for TDP, TMP or T were 91, 96 and 90% for liver extracts, respectively. Analysis of vitamers in tissues of rat submitted to 8 days thiamin deficiency, followed by a 14 days repletion, showed a significant reduction of TPP after 8 days of depletion in liver (-67%), brains (-50%), kidneys (-60%), followed by a complete recovery upon repletion. PMID- 15664337 TI - Sensitive method for the determination of ibutilide in human plasma by liquid chromatography-tandem mass spectrometry. AB - A rapid, selective and sensitive liquid chromatography-tandem mass spectrometry (LC-MS-MS) method was developed and validated for determination of ibutilide in human plasma. The analyte and internal standard sotalol were extracted from plasma samples by liquid-liquid extraction, and separated on a C(18) column, using acetonitrile-water-10% butylamine-10% acetic acid (80:20:0.07:0.06, v/v/v/v) as the mobile phase. Detection was performed on a triple-quadrupole tandem mass spectrometer by multiple reaction monitoring (MRM) mode via TurboIonSpray ionization (ESI). Linear calibration curves were obtained in the concentration range of 20-10,000 pg/ml, with a lower limit of quantitation of 10 pg/ml. The intra- and inter-day precision values were below 8% and accuracy was within +/-3% at all three QC levels. The method was utilized to support clinical pharmacokinetic studies of ibutilide in healthy volunteers following intravenous administration. PMID- 15664338 TI - New lignan metabolites in rat urine. AB - Ten potential lignan metabolites were quantified in rat urine extracts using liquid chromatography-tandem mass spectrometry. The rats were orally administered with the plant lignans 7-hydroxymatairesinol, matairesinol, lariciresinol or secoisolariciresinol, or with the mammalian lignan enterolactone. The samples were enzymatically hydrolysed and solid-phase extracted before analysis. Of the analysed compounds, only trace amounts of 7-oxoenterolactone could be detected in the urine extracts before administration, but after administration of any of the lignans, the excretion of 7-oxoenterolactone increased and monodemethylated matairesinol and 4,4'-dihydroxyenterolactone could be detected. In addition, other novel lignan metabolites were detected, i.e., 7-oxomatairesinol, alpha conidendrin, and alpha- and beta-conidendric acid. PMID- 15664340 TI - Determination of urinary 5-hydroxytryptophol glucuronide by liquid chromatography mass spectrometry. AB - 5-Hydroxytryptophol glucuronide (GTOL) is the major excretion form of 5 hydroxytryptophol (5-HTOL), a minor serotonin metabolite under normal conditions. Because the concentration of 5-HTOL is markedly increased following consumption of alcohol, measurement of 5-HTOL is used as a sensitive biomarker for detection of recent alcohol intake. This study describes the development and evaluation of a liquid chromatography-electrospray ionization mass spectrometry (LC-MS) procedure for direct quantification of GTOL in human urine. Deuterium labelled GTOL (GTOL-(2)H(4)) was used as internal standard. GTOL was isolated from urine by solid-phase extraction on a C(18) cartridge prior to injection onto a gradient eluted Hypurity C(18) reversed-phase HPLC column. The detection limit of the method was 2.0 nmol/L and the measuring range 6-8500 nmol/L. The intra- and inter assay coefficients of variation were <3.5% (n=10) and <6.0% (n=9), respectively. The new LC-MS method was highly correlated with an established GC-MS method for urinary 5-HTOL (r(2)=0.99, n=70; mean 5-HTOL/GTOL ratio=1.10). This is the first direct assay for quantification of GTOL in urine. The method is suitable for routine application. PMID- 15664339 TI - High-performance liquid chromatography with ultraviolet detection for therapeutic drug monitoring of everolimus. AB - We developed and validated a high-performance liquid chromatography-ultraviolet (HPLC-UV) method for determining everolimus concentrations in human whole blood. Sample preparation involved a solid-phase extraction after protein precipitation. The separation of everolimus from internal standard (IS) and endogenous components was achieved using an isocratic elution on an octyl column. The method showed a linear relationship between peak height ratios and blood concentrations in the range of 1-200 ng/mL (r(2)=0.9997). The observed intra- and inter-day assay imprecision had a coefficient of variation (CV)=12.8%, and inaccuracy was 11.4%. The method was found to be precise, accurate, and sensible making it useful for routine therapeutic monitoring of everolimus. PMID- 15664341 TI - Methanol plug assisted sweeping-micellar electrokinetic chromatography for the determination of dopamine in urine by violet light emitting diode-induced fluorescence detection. AB - The use and limitations of a methanol plug assisted sweeping-micellar electrokinetic chromatography (sweeping-MEKC) method is described. Using naphthalene-2,3-dicarboxaldehyde (NDA)-labeled dopamine as a model compound, this new method was also used in the determination of dopamine in actual urine samples. An inexpensive violet light emitting diode (LED) was used for the light source, because this is suitable for fluorescence excitation. The number of theoretical plates of the analyte was determined to be approximately 1 x 10(5) and approximately 2 x 10(5) by means of MEKC and sweeping-MEKC and this was improved to approximately 1 x 10(6) when the methanol plug assisted mode was applied. In addition, the detection limit of NDA-labeled dopamine was determined to be 9.1 x 10(-7) and 1.2 x 10(-8)M by means of MEKC and sweeping-MEKC and this was improved to 4.7 x 10(-9)M when the methanol plug assisted sweeping-MEKC mode was applied. PMID- 15664342 TI - Simultaneous determination of the HIV nucleoside analogue reverse transcriptase inhibitors lamivudine, didanosine, stavudine, zidovudine and abacavir in human plasma by reversed phase high performance liquid chromatography. AB - A reversed phase high performance liquid chromatography method was developed for the simultaneous quantitative determination of the nucleoside reverse transcriptase inhibitors (NRTIs) lamivudine, didanosine, stavudine, zidovudine and abacavir in plasma. The method involved solid-phase extraction with Oasis MAX cartridges from plasma, followed by high performance liquid chromatography with a SymmetryShield RP 18 column and ultraviolet detection set at a wavelength of 260 nm. The assay was validated over the concentration range of 0.015-5 mg/l for all five NRTIs. The average accuracies for the assay were 92-102%, inter- and intra day coefficients of variation (CV) were <2.5% and extraction recoveries were higher than 97%. This method proved to be simple, accurate and precise, and is currently in use in our laboratory for the quantitative analysis of NRTIs in plasma. PMID- 15664343 TI - Simultaneous determination of amphetamine and methamphetamine enantiomers in urine by simultaneous liquid-liquid extraction and diastereomeric derivatization followed by gas chromatographic-isotope dilution mass spectrometry. AB - A simple, rapid, reliable, and economic analytical scheme starting with in situ liquid-liquid extraction and asymmetric (or diastereomeric) chemical derivatization (ChD) followed by gas chromatography (GC)-isotope dilution mass spectrometry (MS) is described for the simultaneous determination of D- and L amphetamine (AP) and methamphetamine (MA) in urine which could have resulted from the administration of various forms of questioned amphetamines or amphetamines generating drugs. By using L-N-trifluoroacetyl-1-prolyl chloride (L-TPC) as chiral derivatizing agent, resolutions of 2.2 and 2.0 were achieved for the separation of AP and MA enantiomeric pairs, respectively, on an ordinary HP-5MS capillary column. The GC-MS quantitation was carried out in the selected ion monitoring (SIM) mode using m/z 237 and 251 as the quantifier ions for the respective diastereomeric pairs of AP-L-TPC and MA-L-TPC. The calibration curves plotted for the two pairs of analytes stretch with good linearity down to 45 ng/mL, and the limits of detection and quantitation determined were as low as 40 and 45 ng/mL, respectively. Also, a comparative study using 10 real-case urine specimens previously screened as positive for MA administration showed mostly tolerable biases between the two sums (of concentration) of D- and L-MA obtained via an asymmetric L-TPC-ChD approach and via an ordinary pentafluoropropionylation (PFPA-ChD) approach, respectively, as well as between the two sums of D- and L-AP obtained thereupon, thus validating the proposed analytical scheme as a promising forensic protocol for the detailed analysis of enantiomeric amphetamines in urine. PMID- 15664344 TI - Highly sensitive determination of the methylated p16 gene in cancer patients by microchip electrophoresis. AB - The p16 tumor suppressor gene is inactivated by promoter region hypermethylation in many types of tumor. Recent studies showed that aberrant methylation of the p16 gene is an early event in many tumors, especially in lung cancer, and may constitute a new biomarker for early detection and monitoring of prevention trials. We detected tumor-associated aberrant hypermethylation of the p16 gene in plasma and tissue DNA from 153 specimens using a modified semi-nested methylation specific PCR (MSP) combining plastic microchip electrophoresis or slab gel electrophoresis, respectively. Specimens were from 79 lung cancer patients, 15 abdominal tumor patients, 30 positive controls and 30 negative controls. The results showed that the positive rate obtained by microchip electrophoresis was more than 26.6% higher and the same specificity was kept when compared with slab gel electrophoresis. The microchip electrophoresis can rapidly and accurately analyze the PCR products of methylated DNA and obviously improve the positive rate of diagnosis of cancer patients when compared with gel electrophoresis. This method with the high assay sensitivity might be used for detection of methylation of p16 gene and even to facilitate early diagnosis of cancer patients. PMID- 15664345 TI - High-performance liquid chromatographic-tandem mass spectrometric method for the determination of clemastine in human plasma. AB - A highly sensitive high-performance liquid chromatographic-tandem mass spectrometric method (HPLC-MS-MS) has been developed to quantitate clemastine in human plasma for the purpose of pharmacokinetic studies. Sample preparation was carried out by liquid-liquid extraction using deuterated clemastine as an internal standard. Chromatographic separation used a C18 reversed phase polymer column giving an extremely fast total run time of 2 min. The method was validated and used for the bioequivalence study of clemastine tablets in healthy male volunteers (n=28). The lower limit of detection proved to be 0.01 ng/ml for clemastine. PMID- 15664346 TI - Determination of flavonoids from Orthosiphon stamineus in plasma using a simple HPLC method with ultraviolet detection. AB - A simple liquid chromatographic method was developed for the simultaneous determination of flavonoids from Orthosiphon stamineus Benth, namely sinensitin, eupatorin and 3'-hydroxy-5,6,7,4'-tetramethoxyflavone, in plasma. Prior to analysis, the flavonoids and the internal standard (naproxen) were extracted from plasma samples using a 1:1 mixture of ethyl acetate and chloroform. The detection and quantification limits for the three flavonoids were similar being 3 and 5 ng/ml, respectively. The within-day and between-day accuracy values, expressed as percentage of true values, for the three flavonoids were between 95 and 107%, while the corresponding precision, expressed as coefficients of variation, for the three flavonoids were less than 14%. In addition, the mean recovery values of the extraction procedure for all the flavonoids were between 92 and 114%. The calibration curves were linear over a concentration range of 5-4000 ng/ml. The present method was applied to analyse plasma samples obtained from a pilot study using rats in which the mean absolute oral bioavailability values for sinensitin, eupatorin and 3'-hydroxy-5,6,7,4'-tetramethoxyflavone was 9.4, 1.0 and 1.5%, respectively. PMID- 15664347 TI - Removal of lipopolysaccharides from protein-lipopolysaccharide complexes by nonflammable solvents. AB - During the recovery of recombinant proteins from gram negative bacteria, many of the methods used to extract proteins from cells release lipopolysaccharides (LPS, endotoxin) along with the protein of interest. In many instances, LPS will co purify with the target protein due to specific or non-specific protein-LPS interactions. We have investigated the ability of alkanediols to effect the separation of LPS from protein-LPS complexes while the complexes are immobilized on ion exchange chromatographic resins. Proteins were complexed with fluorescently labeled LPS and bound to ion exchange resin. Alkanediol washes of the resins were preformed and the proteins eluted. Column eluates were monitored for LPS and protein by fluorescence and UV spectroscopy, respectively. Alkanediols were effective agents for dissociating LPS from protein-LPS complexes. The efficiency of LPS removal increased with increasing alkanediol chain length. The 1,2-alkanediol isomers were more effective than terminal alkanediol isomers in the separation of LPS from protein-LPS complexes, while the separation of LPS from protein-LPS complexes was more efficient on cation exchangers than on anion exchangers. In addition, it was noted during these investigations that the 1,2-alkanediols increased the retention time of the proteins on the ion exchange resins. Alkanediols provide a safer alternative to the use of other organics such as alcohols or acetonitrile for the separation of LPS from protein due to their lower toxicity and decreased inflammability. In addition, they are less costly than many of the detergents that have been used for similar purposes. PMID- 15664348 TI - A novel matrix for high performance affinity chromatography and its application in the purification of antithrombin III. AB - Viscose fiber, a regenerated cellulose, was evaluated for using as a novel matrix for high performance affinity chromatography. With a one-step activation with epichlorohydrin, heparin can be readily covalently attached to the matrix. This heparin-viscose fiber material was used for purifying antithrombin III (AT III) from human plasma. The purity of the AT III from this one-step purification is 93% as measured by SDS-PAGE and the protein recovery yield is about 90%. This column is highly specific as described by the dissociation constant of the complex of immobilized heparin and AT III, which was 2.83 x 10(-5)mol/L. And more important, this viscose fiber material demonstrated its excellent mechanical property that allows the flow rate to reach up to 900 cm/h or more. PMID- 15664349 TI - High-performance liquid chromatographic assay for the determination of total and free topotecan in the presence and absence of anti-topotecan antibodies in mouse plasma. AB - A rapid and sensitive high-performance liquid chromatographic (HPLC) assay has been developed to allow determination of total (i.e. bound and unbound) and free (i.e. unbound) topotecan (TPT) in mouse plasma in the presence and absence of anti-TPT antibodies. The chromatographic analysis was carried out using reversed phase isocratic elution with a Nova-Pak C18 column (3.9 mm x 150 mm, 4 microm) protected by a Nova-Pak C18 guard column (3.9 mm x 20 mm, 4 microm), where 10 mM KH(2)PO(4)-methanol-triethylamine (72:26:2 (v/v/v), pH 3.5) was used as the mobile phase. Topotecan was quantified with fluorescence detection using an excitation wavelength of 361 nm and an emission wavelength of 527 nm. The retention time for the internal standard, acridine, and TPT were 7.4 and 9.0 min, respectively. The lower limit of quantitation (LOQ) for TPT was determined as 0.02 ng in mouse plasma and mouse plasma ultrafiltrate, corresponding to a concentration of 1 ng/ml in 20 microl mouse plasma. The assay was shown to be linear over a concentration range of 1-500 ng/ml. The recoveries of free and total TPT from spiked mouse plasma were within 10% of theoretical values (assessed at 1, 20 and 500 ng/ml). The validated HPLC assay was applied to evaluate TPT pharmacokinetics following administration of TPT to Swiss Webster mice and to hyperimmunized and control BALB/c mice. The assay has been shown to be capable for measuring total and free TPT in mouse plasma with high sensitivity and will allow the testing of the effect of anti-TPT antibodies on the disposition of TPT. PMID- 15664350 TI - Use of ion trap gas chromatography-multiple mass spectrometry for the detection and confirmation of 3'hydroxystanozolol at trace levels in urine for doping control. AB - Stanozolol, a synthetic anabolic androgenic steroid, is often abused in sports to enhance performance. Consequently, the anti-doping laboratories daily screen for its metabolites (3'hydroxystanozolol and 4beta hydroxystanozolol) in all urines, mainly by GC-MS, after enzymatic hydrolysis and TMS derivatization. A sensitive and specific method by GC-MS(3) has been developed for the identification in urine of 3'hydroxystanozolol at trace levels. Full mass spectra and diagnostic ions are presented and a case report is commented. In this case, it was possible to attest the presence of a low concentration of stanozolol metabolite in a sample obtained from a competition test. This would have not been possible with other analytical techniques used in the laboratory. Through this case report, it was also possible to demonstrate the importance of sampling and the difficulties that has to face the laboratory when the pre-analytical step is not correctly performed. PMID- 15664351 TI - Investigation of the pharmacokinetics and determination of tramadol in rabbit plasma by a high-performance liquid chromatography-diode array detector method using liquid-liquid extraction. AB - An HPLC system using a new, simple and rapid liquid-liquid extraction and high performance liquid chromatography-diode array detector method (HPLC-DAD) detection was validated to determine tramadol concentration in rabbit plasma. The method described was applied to a pharmacokinetic study of intravenous tramadol injections in rabbits. The extraction with ethylacetate yielded good response. The recovery of tramadol from plasma averaged 90.40%. Serial plasma samples were obtained prior to, during and after completion of the infusion for determination of tramadol concentrations. Tramadol concentrations were measured using reverse phase high-performance liquid chromatography and pharmacokinetic application with intravenous tramadol in rabbits revealed that tramadol followed one-compartment open model. Maximum plasma concentration (C(max)) and area under the plasma concentration-time curve (AUC) for tramadol were 14.3 microg mL(-1) and 42.2 microg h mL(-1), respectively. The method developed was successfully applied to a simple, rapid, specific, sensitive and accurate HPLC method for investigation of the pharmacokinetics of tramadol in rabbit plasma. PMID- 15664352 TI - High-performance liquid chromatography method for the quantification of rabeprazole in human plasma using solid-phase extraction. AB - A simple, sensitive and selective HPLC method with UV detection (284 nm) was developed and validated for quantitation of rabeprazole in human plasma, the newest addition to the group of proton-pump inhibitors. Following solid-phase extraction using Waters Oasistrade mark SPE cartridges, the analyte and internal standard (Pantoprazole) were separated using an isocratic mobile phase of 5 mM ammonium acetate buffer (pH adjusted to 7.4 with sodium hydroxide solution)/acetonitrile/methanol (45/20/35, v/v) on reverse phase Waters symmetry C(18) column. The lower limit of quantitation was 20 ng/mL, with a relative standard deviation of less than 8%. A linear range of 20-1000 ng/mL was established. This HPLC method was validated with between- and within-batch precision of 2.4-7.2% and 2.2-7.3%, respectively. The between- and within-batch bias was -1.7 to 2.6% and -2.6 to 2.1%, respectively. Frequently coadministered drugs did not interfere with the described methodology. Stability of rabeprazole in plasma was excellent, with no evidence of degradation during sample processing (autosampler) and 3 months storage in a freezer. This validated method is sensitive, simple and repeatable enough to be used in pharmacokinetic studies. PMID- 15664353 TI - Validated liquid chromatographic ultraviolet method for the quantitation of Etoricoxib in human plasma using liquid-liquid extraction. AB - A simple, sensitive and specific HPLC method with UV detection (284 nm) was developed and validated for quantitation of Etoricoxib in human plasma, the newest addition to the group of nonsteroidal anti-inflammatory drugs-a highly selective cyclooxygenase-2 inhibitor. Following a single-step liquid-liquid extraction with diethyl ether/dichloromethane (70/30, v/v), the analyte and internal standard (Zaleplon) were separated using an isocratic mobile phase of water/acetonitrile (58/42, v/v) on reverse phase Waters symmetry C(18) column. The lower limit of quantitation was 5 ng/mL, with a relative standard deviation of less than 20%. A linear range of 5-2500 ng/mL was established. This HPLC method was validated with between- and within-batch precision of 4.1-5.1% and 1.1 2.4%, respectively. The between- and within-batch bias was -3.8-4.7% and -0.6 9.4%, respectively. Frequently coadministered drugs did not interfere with the described methodology. Stability of Etoricoxib in plasma was >90%, with no evidence of degradation during sample processing (autosampler) and 30 days storage in a freezer. This validated method is sensitive and simple with between batch precision of <6% and was used in pharmacokinetic studies. PMID- 15664354 TI - In vivo rat metabolism and pharmacokinetic studies of ginsenoside Rg3. AB - Metabolism of an anti-tumor active component of Panax ginseng, ginsenoside (20R) Rg(3), was studied for better understanding its pharmacokinetics in rat. LC-MS was used to determine Rg(3) and its metabolites in rat plasma, urine and feces samples. An average half-life of 18.5 min was obtained after the ginsenoside was intravenously dosed at 5 mg/kg. However, Rg(3) was not detected in rat plasma collected after oral administration at 100 mg/kg. Only 0.97-1.15% Rg(3) of the dosed amount was determined in feces. Hydrolysis and oxygenated metabolites were detected and identified in feces collected after oral administration by using LC MS and MS-MS. PMID- 15664355 TI - Development and validation of a high-performance liquid chromatography-mass spectroscopy assay for determination of artesunate and dihydroartemisinin in human plasma. AB - A sensitive method has been developed and validated for the determination of artesunate and its active metabolite dihydroartemisinin (DHA) in human plasma using artemisinin as an internal standard. Solid phase extraction (SPE) using Oasis HLB extraction cartridges was used for sample preparation and analysis was performed on a Shimadzu LCMS-2010 in single ion monitoring positive mode using atmospheric pressure chemical ionization (APCI) as an interface. Positive ions were measured using extracted ion chromatogram mode. The extracted ion for artesunate, alpha- and beta-DHA was m/z 221 and for artemisinin was m/z 283. Chromatography was carried out using a Synergi Max-RP, 4 mu, 75 mm x 4.6 mm column using glacial acetic acid 0.1%, acetonitrile and methanol mixture (38:46.5:15.5) as a mobile phase delivered at a flow rate of 0.5 mL/min. The retention times of artesunate, alpha- and beta-DHA and artemisinin were 17.4, 11.8, 18.7 and 13.4 min, respectively, with a total run time of 21 min. The assay was linear over the range 1-3000 ng/mL for artesunate and DHA. The analysis of quality control samples for artesunate 50, 300, 1300 and 2600 ng/mL demonstrated excellent precision with relative standard deviation of 14.3, 11.3, 7.5 and 12.1%, respectively (n=5). Recoveries at concentration of 50, 300, 1300 and 2600 ng/mL were 75, 94.5, 74.3 and 75.5%, respectively; similar results were obtained for precision and recovery of DHA. This liquid chromatography-mass spectroscopy (LC-MS) method for the determination of artesunate and DHA in human plasma has superior specification for sensitivity, sample throughput and robustness than previous methods and can reliably quantitate concentrations of both (artesunate and DHA) compounds as low as 1 ng/mL. PMID- 15664356 TI - pH-dependent functional activity of P-glycoprotein in limiting intestinal absorption of protic drugs 1. Simultaneous determination of quinidine and permeability markers in rat in situ perfusion samples. AB - A simple, specific and sensitive reversed-phase high performance liquid chromatographic (RP-HPLC) method with UV absorbance detection was developed and validated for simultaneous determination of quinidine, verapamil and passive permeability markers, in samples obtained from rat intestinal in situ single-pass perfusion studies. Chromatography was carried out on C18 column with mobile phase comprising of acetate buffer (pH 5.0) and methanol in the ratio of 40:60 (v/v) pumped at a flow rate of 0.6 ml/min and UV detection was employed at 230 and 275 nm. The average retention times for hydrochlorthiazide, frusemide, quinidine, propranolol, and verapamil were 4.9, 5.8, 6.9, 8.9 and 11.3 min, respectively. The calibration curves were linear (R(2)>0.9995) in the selected range for each analyte. The method is specific and sensitive with limit of quantification as 25 ng/ml for quinidine and verapamil. The intra- and inter-day accuracy and precision were found to be good for all the five analytes. The method was found to be reliable in permeability determination and to estimate pH-dependent P glycoprotein (P-gp)-mediated efflux transport of quinidine. Weak bases quinidine, propranolol and verapamil showed pH-dependent permeability, where quinidine permeability increased by 3.6-fold when the luminal pH was changed from pH 4.5 7.4. Inhibition of P-gp by verapamil (200 microM) indicated that about 68% and only 35% of passive transport of quinidine was attenuated by P-gp-mediated efflux at pH 4.5 and 7.4, respectively. In conclusion, low passive transport rates of weakly basic P-gp substrates at lower pH, may lead to more accessibility of these molecules to P-gp within enterocytes thus resulting in pH-dependent functional activity of P-gp as protic drugs moves along the gastrointestinal tract. PMID- 15664357 TI - The trace analysis of alkyl alkylphosphonic acids in urine using gas chromatography-ion trap negative ion tandem mass spectrometry. AB - A sensitive method has been developed for the trace analysis of alkyl alkylphosphonic acids, metabolites of nerve agents, in urine using a benchtop ion trap mass spectrometer. The acids were isolated from urine by simple solid phase extraction and converted to their pentafluorobenzyl esters. An ion trap mass spectrometer in selected reaction monitoring mode provided limits of detection of 0.1 ng/ml for isopropyl, isobutyl, pinacolyl and cyclohexyl methylphosphonic acids and for ethyl ethylphosphonic acid. The detection limit for ethyl methylphosphonic acid was higher (0.5 ng/ml) due to a lower recovery. PMID- 15664358 TI - Evaluation of immobilized metal membrane affinity chromatography for purification of an immunoglobulin G1 monoclonal antibody. AB - The large scale production of monoclonal antibodies (McAbs) has gaining increased relevance with the development of the hybridoma cell culture in bioreactors creating a need for specific efficient bioseparation techniques. Conventional fixed bead affinity adsorption commonly applied for McAbs purification has the drawback of low flow rates and colmatage. We developed and evaluated a immobilized metal affinity chromatographies (IMAC) affinity membrane for the purification of anti-TNP IgG(1) mouse McAbs. We immobilized metal ions on a poly(ethylene vinyl alcohol) hollow fiber membrane (Me(2+)-IDA-PEVA) and applied it for the purification of this McAbs from cell culture supernatant after precipitation with 50% saturation of ammonium sulphate. The purity of IgG(1) in the eluate fractions was high when eluted from Zn(2+) complex. The anti-TNP antibody could be eluted under conditions causing no loss of antigen binding capacity. The purification procedure can be considered as an alternative to the biospecific adsorbent commonly applied for mouse IgG(1) purification, the protein G-Sepharose. PMID- 15664359 TI - Biological monitoring of the five major metabolites of di-(2-ethylhexyl)phthalate (DEHP) in human urine using column-switching liquid chromatography-tandem mass spectrometry. AB - We present a fast and reliable on-line clean-up HPLC-method for the simultaneous determination of the five major urinary metabolites of di-(2-ethylhexyl)phthalate (DEHP) namely mono-(2-ethyl-5-carboxypentyl)phthalate (5carboxy-MEPP), mono-[2 (carboxymethyl)hexyl]phthalate (2carboxy-MMHP), mono-(2-ethyl-5 hydroxyhexyl)phthalate (5OH-MEHP), mono-(2-ethyl-5-oxohexyl)phthalate (5oxo-MEHP) and mono-(2-ethylhexyl)phthalate (MEHP). These metabolites represent about 70% of an oral DEHP dose. We for the first time succeeded to reliably quantify 5carboxy MEPP and to identify 2carboxy-MMHP as major metabolites in native urines of the general population. The analytical procedure consists of an enzymatic hydrolysis, on-line extraction of the analytes from urinary matrix by a restricted access material column (RAM), back-flush transfer onto the analytical column (betasil phenylhexyl), detection by ESI-tandem mass spectrometry and quantification by isotope dilution (limit of detection (LOD) 0.25 microg/l). Median concentrations of a small collective taken from the general population (n=19) were 85.5 microg/l (5carboxy-MEPP), 47.5 microg/l (5OH-MEHP), 39.7 microg/l (5oxo-MEHP), 9.8 microg/l (MEHP) and about 37 microg/l (2carboxy-MMHP). The presented method can provide insights into the actual internal burden of the general population and certain risk groups. It will help to further explore the human metabolism of DEHP an occupational and environmental toxicant of great concern. PMID- 15664360 TI - Pitfalls in trimethylsilylation of anabolic steroids. New derivatisation approach for residue at ultra-trace level. AB - Mixtures such as N-methyl-N-trimethylsilyl-trifluoroacetamide (MSTFA), ammonium iodide and dithioerythreitol (DTE) or MSTFA, trimethyliodosilane and DTE were used for derivatisation of anabolic steroids extracted from 2 g kidney fat and present at ng kg(-1) level. They are leading to unexpected products. Their identity and mechanism of formation have been discussed. A new silylation mixture was developed to overcome these pitfalls: N,O-bis-trimethylsilyl-acetamide was used in combination of 2.5% of MSTFA/I(2) (1000:10 (v/w)). A single product consisting in ether-TMS and/or enol-TMS derivative was observed for all tested steroids with a stability demonstrated for at least 48 h. Quantitative application was proved even at the low ng kg(-1) level in a complex biological matrices, i.e. kidney fat. PMID- 15664361 TI - Analysis of the interaction between hyaluronan and hyaluronan-binding proteins by capillary affinity electrophoresis: significance of hyaluronan molecular size on binding reaction. AB - We developed a method for the analysis of the interaction between hyaluronan (HA) oligosaccharides and hyaluronan-binding proteins (HABPs) using capillary affinity electrophoresis (CAE). The method is based on high-resolution separation of fluorescent-labeled HA molecules in the presence of hyaluronan-binding proteins at different concentrations by capillary electrophoresis (CE) with laser-induced fluorescent detection. Hyaluronan-binding protein from bovine nasal cartilage interacts strongly with HA decasaccharide or larger oligosaccharides. Effect of the molecular size of HA oligomers clearly showed that longer carbohydrate chains than decasaccharide were required for recognition by HA binding protein. Interestingly, the interaction did not cause retardation of HA oligomers as observed in many binding reactions such as the interaction between pharmaceuticals and serum albumin, but showed disappearance of the oligomer peak. Although we cannot explain the accurate mechanism on the interaction, disappearance is probably due to low equilibrium rate between free and conjugate states. The present technique will be useful to compare the relative binding affinity, and to understand the mechanism on the interaction between hyaluronan and hyaluronan-binding proteins. PMID- 15664362 TI - Determination of dextromethorphan and its metabolite dextrorphan in human urine using high performance liquid chromatography with atmospheric pressure chemical ionization tandem mass spectrometry: a study of selectivity of a tandem mass spectrometric assay. AB - Analytical method for the simultaneous determination of dextromethorphan (1) and dextrorphan (2) in urine, based on solid-phase extraction of drug from acidified hydrolyzed biological matrix, were developed. The analytes (1 and 2) and the internal standard (levallorphan, 3, IS) were detected by high-performance liquid chromatography-mass spectrometry (HPLC-MS/MS) in positive ionization mode using a heated nebulizer (HN) probe and monitoring their precursor-->product ion combinations of m/z 272-->215, 258-->201, and 284-->201 for 1, 2, and 3, respectively, in multiple reaction monitoring mode. The analytes and IS were chromatographed on a Keystone Prism reverse phase (50 mm x 2.0 mm) 5 microm column using a mobile phases consisting of a 35/65 or 27/73 mixtures of methanol/water containing 0.1% TFA adjusted to pH 3 with ammonium hydroxide pumped at 0.4 ml/min for 1 and 2, respectively. The limits of reliable quantification of 1 and 2 were 2 and 250 ng/ml, respectively, when 1 ml of urine was processed. The absence of matrix effect was demonstrated by analysis of neat standards and standards spiked into urine extracts originating from five different sources. The linear ranges of the assay were 2-200 and 250-20,000 ng/ml for 1 and 2, respectively. Assay selectivity was evaluated by monitoring the "cross-talk" effects from other metabolites into the MS/MS channels used for monitoring 1, 2, and 3. In addition, an interfering peak originating from an unknown metabolite of 1 into the quantification of dextromethorphan was detected, requiring an effective chromatographic separation of analytes from other metabolites of 1. The need for careful assessment of selectivity of the HPLC MS/MS assay in the presence of metabolites, and the assessment of matrix effect, are emphasized. PMID- 15664363 TI - Determination of lansoprazole and two of its metabolites by liquid-liquid extraction and automated column-switching high-performance liquid chromatography: application to measuring CYP2C19 activity. AB - A simple and sensitive column-switching high-performance liquid chromatographic (HPLC) method for the simultaneous determination of lansoprazole, a proton pump inhibitor and its major metabolites: 5-hydroxylansoprazole and lansoprazole sulfone in human plasma. The test compounds were extracted from 1 mL of plasma using diethyl ether-dichloromethane (7:3, v/v) mixture and the extract was injected into a column I (TSK-PW precolumn, 10 microm, 3.5 mm x 4.6 mm i.d.) for clean-up and column I (C(18) STR ODS-II analytical column, 5 microm, 150 mm x 4.6 mm i.d.) for separation. The peak was detected by a ultraviolet detector set at a wavelength of 285 nm, and the total time for a chromatographic separation was approximately 25 min. The method was validated for the concentration range from 3 to 5000 ng/mL. Mean recoveries were 74.0% for lansoprazole, 68.3% for 5 hydroxylansoprazole, and 79.4% for lansoprazole sulfone. Intra- and inter-day relative standard derivatives were less than 6.1 and 5.1% for lansoprazole, 5.8 and 5.8% for 5-hydroxylansoprazole, 4.4 and 5.9% for lansoprazole sulfone, respectively, at the different concentration ranges. This method is suitable for use in therapeutic drug monitoring and pharmacokinetic studies, and provides use tool for measuring CYP2C19 activity. PMID- 15664365 TI - Quantification of the bombesin/gastrin releasing peptide antagonist RC-3095 by liquid chromatography-tandem mass spectrometry. AB - Bombesin (BN) and its mammalian equivalent, gastrin-releasing peptide (GRP), stimulate cell proliferation and are involved in the pathogenesis of several types of human cancer. BN/GRP and their receptors were shown to be critical for the growth of various human malignancies, such as small-cell lung, prostate, ovary, stomach and breast cancers in the human tumor xenograft model. In the present study, a fast, sensitive, robust method was developed for the determination and quantification of a BN/GRP receptor antagonist RC-3095 (D-Tpi Gln-Trp-Ala-Val-Gly-His-Leupsi(CH2NH)Leu-NH2), in human plasma by liquid chromatography coupled with tandem mass spectrometry. RC-3095 was extracted from 0.2 ml human plasma by protein precipitation using cold acetonitrile (0.4 ml). The method has a chromatographic run of 10 min using a C(8) analytical column (150 mm x 4.6 mm i.d.) and the linear calibration curve over the range was linear from 20 to 10000 ng ml(-1) (r(2)>0.994). The between-run precision, based on the relative standard deviation replicate quality controls, was 5.7% (60 ng ml(-1)), 7.1% (600 ng ml(-1)) and 6.8% (8000 ng ml(-1)). The between-run accuracy was +/ 0.0, 2.1 and 3.1% for the above-mentioned concentrations, respectively. The developed procedure allows the quantitative determination of peptide RC-3095 for pharmacokinetics studies in human plasma. PMID- 15664364 TI - Determination of bulleyaconitine A in human plasma by liquid chromatography electrospray ionization tandem mass spectrometry. AB - A sensitive and specific high-performance liquid chromatography (HPLC) electrospray ionization tandem mass spectrometry (MS-MS) was developed for the determination of bulleyaconitine A (BLA) in human plasma. BLA and internal standard (I.S.) ketoconazole were extracted from the plasma by a liquid-liquid extraction. The supernatant was evaporated to complete dryness and reconstituted with acetonitrile containing 0.1% acetic acid before injecting into an ODS MS column. The gradient mobile phase was composed of a mixture of acetonitrile (containing 0.1% acetic acid, v/v) and 0.1% acetic acid aqueous solution eluted at 0.3 ml/min. BLA and I.S. were determined by multiple reaction monitoring using precursor-->product ion combinations at m/z 644.6-->584.3 and 531.2-->81.6, respectively. Linearity was established for the concentration range of 0.12-6 ng/ml. The recoveries of BLA ranged from 96.93 to 113.9% and the R.S.D. was within 20%. The method is rapid and applicable to the pharmacokinetic studies of BLA in human. PMID- 15664367 TI - Simultaneous determination of urinary creatinine and UV-absorbing amino acids using a novel low-capacity cation-exchange chromatography for the screening of inborn errors of metabolism. AB - A simple and versatile low-capacity cation-exchange chromatography system for the simultaneous determination of creatinine and UV-absorbing amino acids was developed. The separation column was packed with a newly developed low-capacity sulfoacylated macro-porous polystyrene-divinylbenzene resin selective for amino acid cations. Urinary creatinine, creatine, tyrosine, histidine, phenylalanine, and tryptophan were simultaneously separated and determined by an isocratic elution with phosphate/acetonitrile eluent in 25 min. Relative standard deviations (R.S.D.) of the retention times for the analytes were between 0.28 and 1.06%. R.S.D. of peak area responses for the analytes were between 0.75 and 3.51%. The r(2) values for the calibration lines were between 0.9994 and 0.9999. The method could provide the creatinine ratios for the analytes, and was applicable to the screening and/or chemical diagnosis of several inherited disorders of amino-acid metabolism such as phenylketonuria (PKU). PMID- 15664366 TI - Determination of acyclovir in human serum by high-performance liquid chromatography using liquid-liquid extraction and its application in pharmacokinetic studies. AB - A fast, simple and sensitive high performance liquid chromatographic (HPLC) method has been described for determination of acyclovir in human serum. Since acyclovir is a polar compound and soluble in aqueous medium and practically insoluble in most of organic solvents, its analysis in biological fluids in currently published HPLC methods, involve pre-treatment of acyclovir plasma sample including deproteinization or solid phase extraction. In present method liquid-liquid extraction of acyclovir and internal standard (vanillin) is achieved using dichloromethane-isopropyl alcohol (1:1, v/v) as an extracting solvent. Analysis was carried out on ODS column using methanol-phosphate buffer (0.05 M) containing sodium dodecyl sulfate (200 mg/L) and triethylamine (2 mL/L, v/v) as mobile phase (pH=2.3; 5:95, v/v) at flow rate of 2 ml/min. The method was shown to be selective and linear into the concentration range of 10-2560 ng/mL. Accuracy and precision of the method were also studied. The limit of quantitation was evaluated to be 10 ng/mL. This method was applied in bioequivalence study of two different acyclovir preparations after administration of 400mg in 12 healthy volunteers. PMID- 15664369 TI - [Rectum cancer: sphincter saving and quality of life]. PMID- 15664368 TI - Development and validation of an HPLC-UV method for determination of iohexol in human plasma. AB - An HPLC-UV analytical method for estimation of iohexol in human plasma was developed and validated. Protein precipitation and iohexol extraction from plasma (100 microl) was carried out by adding 800 microl perchloric acid (5%, v/v in water) containing iohexol related compound B as the internal standard followed by vortex mixing and centrifugation. The supernatant (90 microl) was then injected onto a microBondapak C(18) column (150 mm x 3.9 mm, 10 microm) maintained at 30 degrees C. The mobile phase comprised of various proportions of acetonitrile and water with a total run time of 12 min and the wavelength of the UV detector was set at 254 nm. The extraction recovery of iohexol from plasma was >95% and the calibration curve was linear (r(2)=0.99) over iohexol concentrations ranging from 10 to 750 microg/ml (n=8). The method had an accuracy of >92% and intra- and inter-day CV of <3.7% and <3.6%, respectively. The method reported is simple, reliable, precise, accurate and has the capability of being used for determination of iohexol in clinical settings. PMID- 15664370 TI - [Pancreas divisum and the dominant dorsal duct syndrome]. AB - Pancreas divisum, the most frequent congenital malformation of the pancreas, results from the absence of embryologic fusion of the dorsal and ventral pancreatic ducts which keep an autonomy of drainage. The dorsal pancreatic duct is dominant and drains the major part of the pancreatic fluid through a non adapted accessory papilla. The high prevalence of pancreas divisum in patients presenting recurrent acute pancreatitis, the presence of obstructive pancreatitis electively located on the dorsal pancreatic duct and the results of the treatments targeted on the accessory papilla are the arguments pleading for the pathogenic character of the pancreas divisum. Currently, the diagnosis of pancreas divisum is based on magnetic resonance imaging. For symptomatic patients (after exclusion of patients with intestinal functional disorders), results of endoscopic sphincterotomy or surgical sphincteroplasty are favourable in 75% of patients with recurrent acute pancreatitis. They are worse in patients with chronic pain. Surgical sphincteroplasty must be discussed in the same manner as the endoscopic treatment for sometimes avoiding multiplication of the procedures. PMID- 15664371 TI - [Pseudocontinent perineal colostomy]. AB - AIM: This prospective study was designed to evaluate functional results of a pseudocontinent perineal colostomy (PCPC) using Schmidt's technique. METHODS: Functional results in eight patients whose rectum were resected due to cancer or anal epidermoid carcinoma and reconstructed by PCPC between January 1995 and July 2002 in our institution were evaluated. Surgical technique and post-operative care were described. Morbidity, functional results and degree of patient satisfaction were reported. Median follow-up was 40 months (18-70 months) and was completed in 100% of patients. RESULTS: There were no operative deaths. Four patients had post-operative complications, whose two patients had partial disunion of sutures, and two patients had urinary infection that was treated by adequate antibiotherapy. No conversion to a definitive abdominal colostomy was performed. On a functional level, one patient has normal continence, six had gas incontinence, and one has occasional minimal soiling. Seventy-five percent of patients were either highly satisfied with their continence. DISCUSSION: PCPC is a reliable technique, which can be proposed as an alternative to a left iliac colostomy following amputation of the rectum due to cancer, provided that certain requirements are met: careful selection of patients, informed consent, flawless surgical technique and lifetime daily colic irrigation. PMID- 15664372 TI - [Diaphragmatic hernia in a rare complication of oesophagectomy for cancer]. AB - INTRODUCTION: Diaphragmatic hernia is a rare complication of oesophagectomy for cancer. We report a series of seven patients to determine characteristics of this entity. PATIENTS AND METHODS: Seven patients (six male and one female, 61 to 68 years old) were operated on for diaphragmatic hernia following oesophagectomy for carcinoma (adenocarcinoma N =4, squamous-cell carcinoma N =3). Oesophagectomy had been performed through abdominal transhiatal approach in four patients and transthoracically in three, with hiatal enlargement in all cases. RESULTS: Three patients, all symptomatic, underwent emergency surgery within two years following oesophagectomy. Of the four patients operated between two and seven years after oesophagectomy, two were symptomatic. Presence of symptoms were neither related with technique of oesophagectomy, nor to type of hiatal enlargement (anterior, or by crura division). All patients with hernia containing small bowel were symptomatic. All patients were operated through abdominal approach. Hernia contained colon three times, small bowel once, and both three times. Hernia reduction needed additional phrenotomy in six patients. Two patients underwent colectomy to treat peroperative colonic ischemia. Diaphragmatic hiatus was calibrated around the gastric tube by direct suture in six patients or with absorbable mesh in one. There was no death. No recurrences occurred with a follow up ranging from one to five years. CONCLUSION: The diaphragmatic hernia after oesophagectomy is due to excessive hiatal enlargement. Hernias occurring early after oesophagectomy are badly tolerated and need urgent reoperation. To prevent this complication of oesophagectomy, we advocate calibration of diaphragmatic hiatus fit to width of gastroplasty. PMID- 15664373 TI - [Results of infrainguinal revascularization in patients with end-stage renal disease]. AB - OBJECTIVES: The aim of this retrospective study was to evaluate the efficacy of bypass in patients with endstage renal disease (ESRD) and to determine predictive factors and precise bypass indications. METHOD: Forty one patients with ESRD underwent 50 bypass, 6 limbs were stage II and 44 stage III or IV according to Leriche and Fontaine classification. Revascularisations procedures were 47 infrainguinal bypass and 3 miscellaneous. Median follow up was 17,0+/-15,7 months. RESULTS: Perioperative mortality rate was 12% (n = 6). Morbidity was as follow : 1 perioperative major imputation and 8 secondary ones. There were 26 secondary death (12 from cardiac events), cumulative survival rate declined to 42,9+/-7,7% and limb salvage rate to 77,2+/-7,5% at 2 years. Primary and secondary potency rates were 53,5+/-10,4% and 70,6+/-10%. Quality of life was good in 28% of revascularised patients. Among risk factors, myocardial events showed a statistical significance in predicting survival, good runoff and bypass occlusion showed a statistical significance in predicting limb salvage. CONCLUSION: Revascularisation can be performed in ESRD patients. However to improve the results full evaluation of myocardial risks, skin lesions and infection of the feet, available autologous vein and nutritional status may be needed in those patients. PMID- 15664375 TI - [Is laparoscopy suitable for large adrenal tumors?]. PMID- 15664374 TI - [Value of surgical ampullectomy in the management of benign ampullary tumors]. AB - AIM OF THE STUDY: To report the results of transduodenal excision (TDE) for tumors of the ampulla of Vater. PATIENTS AND METHODS: From 1998 to 2003, 10 patients underwent a transduodenal excision for presumed benign tumors of the ampulla of Vater. After resection, frozen sections were performed to ensure negative margins. RESULTS: There was no operative mortality. A postoperative pancreatitis occurred in one patient. For nine patients the postoperative course was uneventful. The mean duration of hospital stay was 18 +/-11 days. The final pathology showed adenoma in 8 patients, an adenocarcinoma in one patient and inflammatory lesions in other one. With a mean follow-up of 20 months, endoscopy did not show any recurrence in patients with benign lesion. Patient with an invasive cancer developed recurrence. CONCLUSION: Transduodenal excision is safe and effective treatment for benign ampullary tumors. TDE should be the operation of choice for patients with histologically-proven benign ampulloma, staged as uT1 by endoscopic ultrasound. This approach could reduce the rate of pancreaticaduodenoctomy performed for benign ampullomas. PMID- 15664376 TI - [Desmoid tumor in a male breast in the context of Gardner's syndrome. Case report]. AB - Fibromatosis (Desmoid tumor) of the male breast is an exceptional location. We present a case of such a tumor, in the context of Gardner's syndrome. A palpable mass was discovered in the right breast of a 52 years old man, with a history of rectocolic adenomatous polyposis. Mammography ant thoracic CT scan showed a stellar tumor, mimicking a breast cancer. Treatment consisted of wide excision and histology revealed a desmoid tumor. Desmoid tumors of the breast are benign lesions; they should be widely excised because of a high risk of recurrence. Coloscopy is indicated in the presence of mammary fibromatosis, to look for associated multiple polyps, confirming the diagnosis of Gardner's syndrome. PMID- 15664377 TI - [Ehlers-Danlos syndrome revealed by sigmoid perforation]. AB - We report a case of colonic perforation revealing Ehlers -Danlos syndrome type IV in a male adult. This syndrome is a heritable disorder of collagen synthesis. Its prognosis is severe resulting in vascular rupture or bowel perforation. In his surgical strategy, the surgeon has to cope with the recurrent feature of the colonic perforation. In order to prevent other perforations, a therapeutic scheme has been set up according to literature. PMID- 15664378 TI - [Rectal prolapse resection by trans-anal approach and stapling technique]. AB - We are introducing a simplified cure, derived from Alteimeier's intervention, of rectal prolapse by perineal way resection using a linear cutting stapler. It's a reproducible, quick and easy technique, which permits the elderly patients to come back in their middle of life rapidly with few complications. PMID- 15664379 TI - [Improvement of surgical ampullectomy's postoperative course. To the memory of Henry Doubilet]. PMID- 15664380 TI - Modulation of growth factor action in the cardiovascular system. PMID- 15664381 TI - The biology of vascular endothelial growth factors. AB - The discovery of the vascular endothelial growth factor (VEGF) family members VEGF, VEGF-B, placental growth factor (PlGF), VEGF-C and VEGF-D and their receptors VEGFR-1, -2 and -3 has provided tools for studying the vascular system in development as well as in diseases ranging from ischemic heart disease to cancer. VEGF has been established as the prime angiogenic molecule during development, adult physiology and pathology. PlGF may primarily mediate arteriogenesis, the formation of collateral arteries from preexisting arterioles, with potential future therapeutic use in for example occlusive atherosclerotic disease. VEGF-C and VEGF-D are primarily lymphangiogenic factors, but they can also induce angiogenesis in some conditions. While many studies have addressed the role of angiogenesis and the blood vasculature in human physiology, the lymphatic vascular system has until recently attracted very little attention. In this review, we will discuss recent advances in angiogenesis research and provide an overview of the molecular players involved in lymphangiogenesis. PMID- 15664382 TI - Transcriptional regulation of the Vascular Endothelial Growth Factor gene--a concert of activating factors. AB - The vascular endothelial growth factor A (VEGF-A) is essential during embryonic development as inactivation of only one allele of its gene results in embryonic lethality. Up-regulation of VEGF under physiological situations allows for adaptation to hypoxic stress, to transient inflammatory processes, and to wounding. Its expression also increases all along the process of neovascularization of solid and hematological tumors. The object of this article is to focus on the transcriptional regulation of its gene. The major cis-acting sequences and trans-activating factors will be described as well as the physiological and pathological situations leading to the intervention of such sequences and factors. We will also focus on two transcription factors essential to VEGF gene transcription: the hypoxia-inducible factor-1, which is responsible for its increased by hypoxia, as well as Sp1, which is implicated in the response to various extracellular stimuli. PMID- 15664383 TI - Growth factor signal transduction defects in the cardiovascular system. AB - Growth factors are important molecules mediating both the development as well as adaptive and pathological changes within the cardiovascular system. Growth factors therefore mediate both beneficial and nonbeneficial effects. The beneficial actions include the improvement of endothelial function, stimulation of vascular repair, the formation of new capillaries (angiogenesis), and the growth of collateral arteries (arteriogenesis). These actions represent the conceptual basis for the therapeutic use of growth factors, based on the idea to therapeutically induce or accelerate a given beneficial process. The beneficial effects of growth factors have to be separated from effects where growth factors act as mediators of pathological processes such as atherogenesis and plaque destabilization. This review article focuses on the physiological and beneficial effects of growth factors in the vessel wall and describes and explains the dysfunction of these effects under certain conditions now referred to as "growth factor signal transduction defects". PMID- 15664384 TI - PDGF receptor kinase inhibitors for the treatment of restenosis. AB - The clogging of arteries by neointima is a hallmark of atherosclerosis and of restenosis following balloon angioplasty. The realization in the 1980s that PDGF and its receptor play a key role in the onset of neointimal formation led us to develop PDGFR kinase inhibitors as antirestenosis agents. In this review, we describe the development of these inhibitors and their implementation as antirestenosis agents by localized delivery to the site of injury. PMID- 15664385 TI - Tyrosine phosphatases in vessel wall signaling. AB - Protein tyrosine phosphatases (PTPs) are critical regulators of cellular processes like migration, proliferation and differentiation that are involved in physiological and pathological vessel wall function. In this review, we summarize the biochemistry of this enzyme family, discuss the present knowledge concerning the identity and involvement of PTPs in vascular cells and in pathways of relevance to cardiovascular diseases. We also briefly introduce ongoing efforts to develop inhibitors of PTPs, and finally point to some opportunities for use of such agents in novel treatment strategies. PMID- 15664386 TI - TGF-beta receptor function in the endothelium. AB - Genetic studies in mice and humans have revealed the pivotal role of transforming growth factor-beta (TGF-beta) signaling during angiogenesis. Mice deficient for various TGF-beta signaling components present an embryonic lethality due to vascular defects. In patients, mutations in the TGF-beta type I receptor ALK1 or in the accessory TGF-beta receptor endoglin are linked to an autosomal dominant disorder of vascular dysplasia termed Hereditary Haemorrhagic Telangiectasia (HHT). It has puzzled researchers for years to explain the effects of TGF-beta being a stimulator and an inhibitor of angiogenesis in vitro and in vivo. Recently, a model has been proposed in which TGF-beta by binding to the TGF-beta type II receptor can activate two distinct type I receptors in endothelial cells (ECs), i.e., the EC-restricted ALK1 and the broadly expressed ALK-5, which have opposite effects on ECs behavior. ALK1 via Smad1/5 transcription factors stimulates EC proliferation and migration, whereas ALK5 via Smad2/3 inhibits EC proliferation and migration. Here, the new findings are presented concerning the molecular mechanisms that take place in ECs to precisely regulate and even switch between TGF-beta-induced biological responses. In particular, the role of the accessory TGF-beta receptor endoglin in the regulation of EC behavior is addressed and new insights are discussed concerning the possible mechanisms that are implicated in the development of HHT. PMID- 15664387 TI - The NR4A subfamily of nuclear receptors: new early genes regulated by growth factors in vascular cells. AB - The molecular mechanisms regulating endothelial cell activation and vascular smooth muscle cell proliferation are critical in the pathological processes underlying atherosclerosis. Numerous growth factors and cytokines trigger the complex and redundant signaling pathways that regulate cell cycle entry; however, the genes controlling these processes are not fully known. Applying techniques for differential gene expression analysis, new transcription factors have been identified in these mechanisms, among them the three members of the NR4A subfamily of nuclear receptors (NRs). These transcription factors (NOR-1, Nur77 and Nurr1) are products of immediate-early genes whose expression and activity is regulated in a cell-specific manner by a variety of extracellular mitogenic, apoptotic and differentiation stimuli. Unlike most NRs whose transcriptional activity is regulated by direct modulatory ligands, NR4A genes do not appear to require ligand binding for activation, and in vascular cells they are highly responsive to growth factors, cytokines, lipoproteins and thrombin. In this review, we discuss our present knowledge on the role of this subfamily of NRs in vascular cell function. PMID- 15664388 TI - Control of arterial branching morphogenesis in embryogenesis: go with the flow. AB - Formation of a properly branched vascular system during embryogenesis is crucial for embryo survival. Here we review the regulation of the morphogenesis of the arterial and venous system during embryogenesis. We show that in addition to deterministic patterning mechanisms and plasticity of endothelial cells, arterial venous differentiation and branching morphogenesis involves a prominent role for blood flow. Based on in vivo observations of developing arteries, we identified a novel morphological event crucial for the morphogenesis of the arterial tree, disconnection of small side branches. This disconnection of side branches occurs exactly at the point of bifurcation. The rate of disconnection of side branches depends on flow velocity and branching angle. The balance between disconnection and maintenance of arterial side branches determines the number of side branches connected to a large artery. Based on these observations, we postulate that the number of pre-existing collaterals connected to a large artery is a function of the disconnection process and can be regulated by hemodynamics. We furthermore show that embryonic arteries already adapt their lumen diameter to the amount of flow carried. Taken together, we suggest that hemodynamics plays a pivotal role in shaping the arterial system. We suggest that flow-evoked remodeling processes determine the number of preexisting collaterals during critical periods of embryo fetal development. Insight into these basic principles of arterial growth and branching during embryogenesis may aid to understanding the observed variability in the capacity to establish a collateral circulation in patients with ischemic diseases and finding new strategies for therapeutic arteriogenesis. PMID- 15664389 TI - Role of neural guidance signals in blood vessel navigation. AB - Despite the tremendous progress achieved in both vasculogenesis and angiogenesis in the last decade, little is still known about the molecular mechanisms underlying the pathfinding of blood vessels during their formation. However, emerging evidence shows that different axonal guidance cues, including members of the Slit and semaphorin families, are also involved in the blood vessel guidance, suggesting that blood vessels and nerves share common mechanisms in choosing and following specific paths to reach their respective targets. These promising findings open novel avenues not only in vascular biology but also in therapeutic angiogenesis. Indeed, the identification of new molecules involved in the guidance of blood vessels may be helpful in designing angiogenic strategies, which would insure both the formation of new blood vessels and their guidance into an organized and coordinated network. PMID- 15664390 TI - Update on therapeutic neovascularization. AB - Therapeutic neovascularization for cardiovascular ischemia is a promising avenue in spite of disappointing early clinical trial results. The concept of three different mechanisms of neovascularization has served to define potential therapeutic targets such as vascular remodeling and stem cell recruitment, but it is anticipated that this will lose significance as the pleiotropic nature of angiogenic cytokines becomes fully understood. With the rapidly growing body of data on growth factors and pro-angiogenic strategies, approaches will emerge that are more effective than the ones that have been tested clinically thus far. Combinations of growth factors, for instance to stabilize vessels, or growth factors combined with cell transplants deserve more attention but will make the design of preclinical and clinical studies increasingly complex. Recent developments suggest that when using the appropriate dose and treatment regimens, even single growth factor therapy can result in stable and functional vessels. Whether gene therapy or protein therapy will be optimal for this purpose depends mainly on technical developments in vector design and production and on progress in the engineering of slow release matrix formulations for proteins. With the increasing complexity of therapeutic strategies, it remains imperative that these approaches are rationally based on fundamental and preclinical data. PMID- 15664391 TI - Growth factor-induced therapeutic angiogenesis in the heart: protein therapy. AB - Therapeutic angiogenesis, stimulated growth of new vasculature to compensate for tissue ischemia, remains an unfulfilled promise. After nearly a decade of trials, the development of drugs capable of stimulating revascularization of underperfused tissues remains an exciting but unrealized goal in cardiovascular therapeutics. This review will summarize experiences in therapeutic angiogenesis studies employing protein therapies and will draw conclusions based on our current understanding of biological processes of new vessel growth. PMID- 15664392 TI - Growth factor-induced therapeutic angiogenesis and arteriogenesis in the heart- gene therapy. AB - Myocardial ischemia is one of the most promising targets of gene therapy. Although several growth factors and delivery approaches have yielded positive results in preclinical studies, first clinical studies have shown little or no real clinical benefit to the patients. It is likely that less than optimal gene therapy approaches have been used so far, and more thorough preclinical studies are needed in order to establish safe, efficient pro-angiogenic therapy. Growth factor, gene transfer vector, delivery method and target microenvironment need to be chosen based on the therapeutic target. It has become apparent that induction of large collateral arteries in the myocardium may need a different approach than rapid growth of neovasculature around infarction scar. Large animal models are necessary in the determination of optimal therapeutic agent, dose and clinically relevant delivery strategy. PMID- 15664393 TI - Vitamins C and E prevent endothelial VEGF and VEGFR-2 overexpression induced by porcine hypercholesterolemic LDL. AB - OBJECTIVE: Vascular endothelial growth factor (VEGF) is believed to play a role in the development of atherosclerosis and has been found to be increased in hypercholesterolemia. We examined the hypothesis that endothelial VEGF and VEGF receptor-2 (VEGFR-2) expression is upregulated by hypercholesterolemic low density lipoprotein (LDL) and, because it could be driven by oxidative stress, we tested whether vitamin C and E supplementation could modulate it. METHODS: Native LDL were characterized after isolation from adult normal (C-LDL), hypercholesterolemic (HC-LDL) and hypercholesterolemic mini-pigs receiving vitamins C and E (HCV-LDL). VEGF, VEGFR-2, HIF-1 alpha and superoxide anion (O(2)(-)) productions were measured in porcine coronary endothelial cells (ECs) incubated for 48 h with native LDL. The effect of exogenous ascorbic acid and alpha- or beta-tocopherol was also studied. RESULTS: HC-LDL, with high cholesterol (P<0.05) and reduced tocopherol/cholesterol ratio (P<0.05), increased significantly VEGF and VEGFR-2 (p<0.001) in EC, associated with higher O(2)(-) and HIF-1 alpha expression, in comparison with C-LDL and HCV-LDL. The addition of vitamin C and alpha- or beta-tocopherol to the culture medium prevented the induction of VEGF and VEGFR-2 expression by HC-LDL, both at mRNA and protein levels. CONCLUSIONS: Our data suggest HC-LDL induce endothelial VEGF and VEGFR-2 overexpression at least by increasing oxidative stress, and HIF-1 alpha is one of the signaling mechanisms involved. Prevention of VEGF and VEGFR-2 upregulation could help explain the beneficial effects of vitamins C and E in hypercholesterolemia-induced experimental atherosclerosis. PMID- 15664394 TI - Activation of vascular smooth muscle cells by TNF and PDGF: overlapping and complementary signal transduction mechanisms. AB - OBJECTIVE: Because tumor necrosis factor-alpha (TNF) has been implicated in the pathogenesis of vein graft neointimal hyperplasia, we sought to determine mechanisms by which TNF could induce proliferative and migratory responses in smooth muscle cells (SMCs). METHODS AND RESULTS: In rabbit jugulocarotid interposition vein grafts, SMCs expressed TNF as early as four days postoperatively. In rabbit aortic SMCs, TNF and platelet-derived growth factor (PDGF) elicited comparable migration (1.7-fold/basal), and their effects were partially additive. In contrast, while TNF failed to promote SMC [(3)H]thymidine incorporation alone, it doubled the [(3)H]thymidine incorporation observed with PDGF alone. To gain mechanistic insight into these phenomena, we found that TNF and PDGF each activated p38(mapk) equivalently in SMCs, but that PDGF was two to three times more efficacious than TNF in activating SMC extracellular signal regulated kinases (ERK) 1 and 2 and phosphoinositide 3-kinase. However, only TNF activated NF kappa B. SMC [(3)H]thymidine incorporation that depended on TNF, but not PDGF, was abolished by overexpression of a dominant-negative I kappa B alpha mutant. Inhibition of ERK activation by U0126 reduced SMC migration stimulated only by PDGF (by 35%, P<0.05), but not by TNF. Inhibition of phosphoinositide 3 kinase by LY294002, however, significantly reduced both TNF- and PDGF-stimulated chemotaxis (by 38-54%, P<0.05). In contrast, both U0126 and LY294002 abolished SMC [(3)H]thymidine incorporation induced by either TNF, PDGF, or both agonists. CONCLUSIONS: In primary rabbit SMCs, TNF promotes migration and mitogenesis through signaling mechanisms that are both distinct from and overlapping with those employed by PDGF. TNF-induced SMC mitogenesis requires complementary co stimulation with other growth factors. PMID- 15664395 TI - Diversity and similarity in signaling events leading to rapid Cox-2 induction by tumor necrosis factor-alpha and phorbol ester in human endothelial cells. AB - OBJECTIVE: This study examines whether cyclooxygenase 2 (Cox-2) synthesis in human endothelial cells involves different signaling pathways when induced by the proinflammatory cytokine tumor necrosis factor-alpha (TNF alpha) or by the tumor and angiogenic promoter phorbol ester (PMA). Moreover, the hypothesis that reactive oxygen species (ROS) and an altered redox status within the cell are fundamental steps for Cox-2 synthesis is verified. METHODS: Human endothelial cells isolated from umbilical vein (HUVEC) were exposed to PMA and TNF alpha and Cox-2 protein and mRNA levels were evaluated by Western blot and Real-Time Quantitative Reverse Transcription-PCR analysis. Prostaglandin E(2) (PGE(2)) and 6-keto prostaglandin F(1 alpha) (6-keto-PGF(1 alpha)) levels were measured in cell medium as an index of Cox-2 activity. Intracellular ROS formation was detected by flow cytometry in HUVEC loaded with the oxidant-sensitive 2',7' dichlorofluorescein diacetate (DCFH-DA) and by nitroblue tetrazolium (NBT) reduction. Reduced and oxidized glutathione (GSH and GSSG) were measured by HPLC. RESULTS: Data show that TNF alpha and PMA signal for early Cox-2 induction through distinct pathways. PMA-induced Cox-2 expression involves a small GTPase dependent pathway acting via tyrosine kinase, activation of protein kinase C (PKC) and of the mitogen-activated protein kinase (MAPK) ERK1/2. Conversely, MAPK p38 is critical for Cox-2 induction by TNF alpha. Of interest, intracellular ROS generation and consequent GSH/GSSG ratio reduction represents a common step through which PMA and TNF alpha signal for early Cox-2 induction. In addition, we provide evidence that phosphatidylinositol 3 (PI3)-kinase activation plays a regulatory role for Cox-2 synthesis in HUVEC. CONCLUSION: Cox-2 represents a critical link among vascular homeostasis, inflammatory response, angiogenesis and tumor growth. The finding that two independent pathways and an overlapping upstream event signal for Cox-2 induction in HUVEC may be of relevance to develop strategies aimed at selectively interfering with Cox-2 regulating pathways. PMID- 15664396 TI - Tranilast attenuates cardiac matrix deposition in experimental diabetes: role of transforming growth factor-beta. AB - OBJECTIVE: The pathological accumulation of extracellular matrix is a characteristic feature of diabetic cardiomyopathy that is directly related to a loss of function. Tranilast (n-[3,4-anthranilic acid), used for the treatment of fibrotic skin diseases, has also been shown to inhibit transforming growth factor beta (TGF-beta)-induced matrix production in kidney epithelial cells. METHODS: To investigate the effects of tranilast in the diabetic heart, we examined its effects in cultured cardiac fibroblasts and then assessed its effects in (mRen 2)27 diabetic rats with established disease (8 weeks after streptozotocin). RESULTS: In vitro studies demonstrated a 58% reduction in TGF-beta1-induced 3[H] hydroxyproline incorporation with tranilast 30 microM (p<0.01). At 16 weeks, diabetes in the Ren-2 rat was associated with increased cardiac fibrosis and evidence of TGF-beta1 activation, as measured by the abundance of phosphorylated Smad2. Despite persistent hyperglycaemia and hypertension, tranilast attenuated cardiac fibrosis by 37% (p<0.05) in association with reduction in phospho-Smad2 (p<0.01). CONCLUSION: These findings indicate that tranilast has antifibrotic actions in the Ren-2 model of experimental diabetic cardiac disease by mechanisms that might attributable to reduced TGF-beta activity. PMID- 15664397 TI - In vitro differences between venous and arterial-derived smooth muscle cells: potential modulatory role of decorin. AB - OBJECTIVE: We analyzed the phenotypic and functional differences between venous and arterial smooth muscle cells (SMC) and the role of decorin in modulating these differences. METHODS AND RESULTS: SMC were isolated from the jugular veins and carotid arteries of male white New Zealand rabbits. Venous SMC demonstrated increased proliferation (2-fold, p<0.001), migration (1.7-fold, p<0.001), and collagen synthesis (4-fold, p<0.001), with decreased adhesion to collagen and fibronectin (1.2-fold, p<0.01) compared to arterial SMC. Higher levels of gelatinase activity (MMP-2 and MMP-9) and tissue inhibitor of metalloproteinase (TIMP) were also observed in venous SMC. Venous SMC demonstrated increased expression of SMemb and decreased expression of SM1--markers of a dedifferentiated and differentiated phenotype, respectively. Arterial SMC produced increased levels of the inhibitory proteoglycan, decorin, compared to venous SMC. Conditioned medium from arterial SMC (ASMC-CM) significantly decreased DNA synthesis, collagen synthesis, and gelatinase activity in venous SMC. Removal of decorin from ASMC-CM by immunoprecipitation significantly reversed the inhibitory effects of ASMC-CM on venous SMC proliferation and collagen synthesis but did not affect gelatinase activities. CONCLUSION: Venous SMC are more dedifferentiated and demonstrate increased proliferative and synthetic capacity than arterial SMC. Differential decorin expression between arterial and venous SMC contributes to these differences in biologic behavior. Venous SMC properties may contribute to accelerated atherosclerosis in venous bypass grafts. PMID- 15664398 TI - Roles of forkhead transcription factor Foxc2 (MFH-1) and endothelin receptor A in cardiovascular morphogenesis. AB - OBJECTIVE: Foxc2/MFH-1 is a member of the forkhead family of transcription factors and Foxc2-deficient mice exhibit aortic arch anomalies (type B interruption of the aortic arch). Endothelin receptor type-A (ETA) is one of the two known endothelin receptors that belong to the G-protein-coupled receptor family. ETA-deficient mice show defects in the great arteries, primarily type B interruption of the aortic arch. Based on similar phenotypes in the cardiovascular system of Foxc2- and ETA-deficient mice, we investigated whether Foxc2 and ETA have a close relationship in aortic arch patterning. METHODS: The Foxc2 and ETA homozygotes were obtained by crossing the Foxc2 and ETA heterozygotes, respectively. The double Foxc2/ETA homozygotes were obtained by crossing the double Foxc2/ETA heterozygotes. RESULTS: We investigated the expression of ETA in Foxc2-null mice and the expression of Foxc2 in ETA-null mice and found that the absence of either Foxc2 or ETA had no effect on the expression of the other. Next, we analyzed mice lacking both Foxc2 and ETA to examine the relationship between Foxc2 and ETA on aortic arch patterning in vivo. We found that the majority of Foxc2/ETA double-mutant embryos died around 11.5 dpc and that all surviving mice had persistent truncus arteriosus. CONCLUSIONS: The results suggest that Foxc2- and ETA-expressing cells additively form the aorticopulmonary septum. PMID- 15664400 TI - Liposomal Hsp90 cDNA induces neovascularization via nitric oxide in chronic ischemia. AB - OBJECTIVE: Induction of angiogenesis has been reported subsequent to eNOS overexpression or activation, the latter involving Hsp90 as a chaperone protein. Here, we investigated the potential of regional Hsp90 overexpression to induce therapeutic neovascularization in vivo in a chronic rabbit hindlimb ischemia model. METHODS: In rabbits (n=7 per group), the external femoral artery was excised at day 0 (d0). At d7, liposomes containing eGFP (control group) or Hsp90 were retroinfused into the anterior tibial vein. At day 7 and day 35, angiographies were obtained and analyzed for collateral formation and perfusion velocity (frame count score) (% of d7 values). Capillary/muscle fiber (C/MF) ratio was calculated from five muscle areas of the ischemic limb. L-NAME and Geldanamycin were co-applied, where indicated. RESULTS: Compared to mock-treated controls, Hsp90 transfected increased C/MF ratio at day 35 (1.78+/-0.15 vs. 1.19+/-0.13, p<0.05), an effect blunted by L-NAME (1.39+/-0.11). Hsp90 transfection increased collateral formation (157+/-11% vs. 110+/-13%) and frame count score (174+/-18% vs. 117+/-10%), both sensitive to inhibition by L-NAME coapplication (135+/-17% and 134+/-14%, respectively). Of note, C/MF ratio was found elevated 3 days after Hsp90 transfection (1.61+/-0.16 at d10), at a time point when collateral formation was unchanged (106+/-6%), and tended to remain elevated in the presence of L-NAME applied thereafter (1.64+/-0.35 at d35), though L-NAME blocked subsequent changes in collateral growth or increase in perfusion at d35. CONCLUSIONS: We conclude that Hsp90 is capable of inducing angiogenesis and arteriogenesis via nitric oxide (NO) in a rabbit model of chronic ischemia. Our findings describe the capillary level as an initial site of Hsp90-cDNA-induced neovascularization, followed by growth of larger conductance vessels, resulting in an improved hindlimb perfusion. PMID- 15664399 TI - Erythropoietin prevents the acute myocardial inflammatory response induced by ischemia/reperfusion via induction of AP-1. AB - OBJECTIVE: Erythropoietin (EPO) prevents the myocardial dysfunction induced by ischemia/reperfusion (I/R). Since I/R-induced myocardial dysfunction is associated with an acute inflammatory response, we assessed the anti-inflammatory properties of EPO using in vitro and in vivo models of I/R. METHODS: Isolated cardiac myocytes were exposed to anoxia/reoxygenation (A/R; the in vitro counterpart to I/R). Hearts were challenged with I/R in situ. RESULTS: In vitro, A/R increased myocyte oxidant stress and converted the myocytes to a proinflammatory phenotype (these myocytes induced PMN transendothelial migration). Pretreatment of the myocytes with EPO prevented the A/R-induced proinflammatory effects. EPO increased myocyte (1) nuclear translocation of AP-1 (c-fos/c-jun), (2) eNOS, but not iNOS, protein expression, and (3) NO production. An AP-1 "decoy" oligonucleotide prevented the induction of eNOS by EPO and reversed the beneficial effect of EPO. An inhibitor of phosphatidylinostol 3 (PI3)-kinase prevented the nuclear translocation of AP-1 induced by EPO. In vivo, in wild type mice, I/R induced an increase in myocardial MPO activity (indicative of PMN infiltration); an effect prevented by pretreatment of the mice with EPO. This anti-inflammatory effect of EPO was not observed in cardiac specific c-fos( /-) mice. CONCLUSIONS: Collectively, these findings indicate that EPO can ameliorate the myocardial inflammatory response in both in vitro and in vivo models of I/R. This beneficial effect of EPO is mediated by eNOS-derived NO via a PI3-kinase-dependent activation of AP-1. PMID- 15664401 TI - Tenascin-C is an essential factor for neointimal hyperplasia after aortotomy in mice. AB - OBJECTIVE: Neointimal hyperplasia at the arterial anastomotic site is a critical problem during cardiovascular surgery. It has been suggested that tenascin-C (TN C), an extracellular matrix (ECM) glycoprotein, might play an important role in neointimal hyperplasia. In this study, the direct contribution of tenascin-C to neointimal hyperplasia after aortotomy was examined using tenascin-C-deficient (TNKO) mice. METHODS AND RESULTS: A simple aortotomy model was constructed in mice. In wild-type (WT) mice, neointimal hyperplasia was observed at the suture sites at days 14 and 28. Immunohistochemical staining showed strong expression of tenascin-C in both neointima and media around the suture line at day 14. At day 28, tenascin-C staining was detected in neointima, but not in media. In tenascin C-deficient mice, much less neointimal hyperplasia was seen compared to that in wild-type mice, and the mean neointima/media area ratio decreased to 52.8% and 34.3% at days 14 and 28, respectively. The proliferating cell nuclear antigen indices in wild-type mice were twice those in tenascin-C-deficient mice at day 14. There were fewer Alcian blue-positive proteoglycans deposited in the neointima of tenascin-C-deficient mice than in wild-type mice. These results suggest that tenascin-C promotes neointimal cell migration and proliferation, and the deposition of proteoglycans. CONCLUSIONS: We have presented direct evidence that tenascin-C is a crucial molecule in neointimal hyperplasia at anastomotic sites. PMID- 15664402 TI - Effects of all-trans retinoic acid on orphan receptor APJ signaling in spontaneously hypertensive rats. AB - OBJECTIVE: Studies show general agreement that all-trans retinoic acid (atRA) has been linked to the regulation of G protein-coupled receptor (GPCRs) signaling. To further validate effects of atRA on the cardiovascular GPCRs, the present study was designed to assess whether atRA will modulate orphan receptor APJ, a homologue of angiotensin II type 1 (AT(1)) receptor. METHODS: Real-time polymerase chain reaction and Western blot methods were performed to examine the expression of APJ and its endogenous ligand apelin in spontaneously hypertensive rats (SHR) and Wistar-Kyoto (WKY) rats after chronic atRA treatment. RESULTS: APJ and apelin expression were markedly depressed in placebo-treated SHR, compared with WKY rats (p<0.01). However, in atRA-treated SHR, a significant upregulation of APJ and apelin expression was observed in both heart and aorta (p<0.05), accompanied by a reduction of AT(1) expression, an elevation of serum nitric oxide levels and a subsequent decrease of blood pressure. CONCLUSIONS: Chronic atRA treatment activates gene and protein expression of APJ and apelin and reduces blood pressure in SHR, suggesting that atRA may regulate the balance between apelin-APJ and angiotensin II-AT(1) signaling and have potential clinical value in the prevention and treatment of human hypertension. PMID- 15664403 TI - Continuous inhalation of carbon monoxide attenuates hypoxic pulmonary hypertension development presumably through activation of BKCa channels. AB - OBJECTIVE: We tested the hypothesis that inhalation of a low concentration of exogenous carbon monoxide (CO) attenuates the development of hypoxic pulmonary artery hypertension by activation of large-conductance voltage and Ca(2+) activated K(+) channels (BK(Ca)). METHODS: The BK(Ca) activity was measured using whole-cell and inside-out patch clamp recordings in Wistar rat pulmonary artery (PA) myocytes. Pulmonary artery pressures were measured in vivo and membrane potentials were recorded in vitro in pressurized resistance arteries. RESULTS: Chronic CO inhalation slightly increases single-channel conductance of BK(Ca) channels and induces a large increase in the sensitivity of BK(Ca) channels to Ca(2+) of PA myocytes from normoxic and chronic hypoxic rats. Consequently, BK(Ca) currents are increased and play a more prominent role in controlling resting membrane potential of PA myocytes. Chronic CO inhalation also reduces hemodynamic changes induced by chronic hypoxia and attenuates hypoxic pulmonary artery hypertension. CONCLUSION: Chronic inhalation of CO attenuates hypoxic pulmonary artery hypertension development presumably through activation of BK(Ca) channels. These results highlight the potential use of CO as a novel avenue for research on the treatment of pulmonary artery hypertension (PAHT) in a similar manner to another gasotransmitter, nitric oxide. PMID- 15664405 TI - Evidence of a genetic heterogeneity of familial hydatidiform moles. AB - We previously mapped a maternal recessive locus responsible for familial hydatidiform moles (HMs) to 19q13.4. The candidate region has recently been narrowed down to 1.1 megabases. Here, we report the segregation of alleles at 18 genetic markers, including nine new ones, from the HMs candidate region in a recently reported consanguineous family. In this family, five affected women had a total of seven HMs, three miscarriages, and three normal children. Linkage and haplotype analyses exclude linkage to 19q13.4 and indicate the presence of a second recessive locus responsible for familial molar pregnancies. The heterogeneity in the phenotype of the conceptuses of patients with familial HMs is in agreement with previous observations and seems to be a common feature of this condition. This indicates that the homozygous genetic defects leading to hydatidiform moles can be modulated by other genetic or environmental factors. The identification of these factors may unravel natural ways to treat these forms of reproductive wastage and reverse the infertility of women with recurrent moles. PMID- 15664406 TI - Fetal and placental chromosomal mosaicism revealed by QF-PCR in severe IUGR pregnancies. AB - A number of genetic and environmental factors are taken into account as responsible for intrauterine growth restriction (IUGR); nevertheless, the relevance of genetic alteration in IUGR aetiology remains to be determined. The aim of this study was to investigate using a combined cytogenetic-molecular approach, improved by a new application of QF-PCR method, the presence of mosaic chromosomal changes in fetal/placental samples from 12 pregnancies with unexplained severe IUGR. This multiple approach allowed us to reveal and quantify subtle chromosomal mosaicisms with less than 5% of trisomic cells even in cases in which cytogenetic and FISH analyses failed to reveal them. These are three pregnancies with a mosaic trisomy for chromosomes 7, 2 and 14; the former case presented matUPD7 and was previously described in this journal (Placenta 22 (2001) 813) in association with pre- and postnatal growth restriction. It is intriguing that chromosomes 7, 2 and 14 are known or suspected to harbour imprinted genes, so that an unbalanced gene dosage in a subset of cells during embryonic development could lead to an early impairment of placental function. Our findings indicate that extensive molecular and cytogenetic studies of IUGR fetal and placental tissues are necessary to reveal at least part of the heterogeneous genetic lesions implicated in IUGR phenotypes. PMID- 15664407 TI - Characterisation of deep arterio-venous anastomoses within monochorionic placentae by vascular casting. AB - OBJECTIVE: To characterise arterio-venous anastomoses (AVA) in monochorionic (MC) placentae and determine (i) whether shared cotyledons lie beneath the co termination of an artery from one twin and a vein to the contralateral twin and (ii) whether all AVA can be detected by visual inspection of the chorionic plate. METHODS: Vascular casts were made of 15 MC placentae. The number of typical AVAs suspected visually before digestion was compared with the number of AVAs identified after acid digestion. RESULTS: Thirty-three of 67 (49%) suspected typical AVAs were confirmed as typical after casting. There were five false positives and no false negatives. The remainder were classified as atypical AVAs, found in > or =90% of MC placentae. Type I (small vascular connections between two apparently normal cotyledons not seen before casting) and Type II (shared cotyledons arising within larger apparently normal cotyledons) atypical AVAs were found in 53% and 73% of placentae, respectively. CONCLUSIONS: Only half the shared cotyledons in MC placentae are characterised by co-termination of an artery and vein on the chorionic plate. We report the existence of deep anastomoses beneath the chorionic plate that cannot be visualised by chorionic plate inspection. These findings have implications for laser treatment of twin twin transfusion syndrome. PMID- 15664409 TI - Cloning, sequencing, structural and molecular biological characterization of placental protein 20 (PP20)/human thiamin pyrophosphokinase (hTPK). AB - Full-length cDNAs of placental protein 20 (PP20) were cloned by screening a human placental cDNA library, which encode a 243 amino acid protein, identical to human thiamin pyrophosphokinase (hTPK) as confirmed by protein sequence analysis. Genomic alignment showed that the PP20/hTPK gene contains 9 exons. It is abundantly expressed in placenta, as numerous EST clones were identified. As thiamine metabolism deficiencies have been seen in placental infarcts previously, these indicate that PP20/hTPK may have a role in placental diseases. Analysis of the 1kb promoter region showed numerous putative transcription factor binding sites, which might be responsible for the ubiquitous PP20/hTPK expression. This may also be in accordance with the presence of the protein in tissues responsible for the regulation of the exquisite balance between cell division, differentiation and survival. TPK activity of the purified and recombinant protein was proved by mass spectrometry with electrospray ionization. By Western blot, PP20/hTPK was found in all human normal and tumorous adult and fetal tissues in nearly equal amounts, but not in sera. By immunohistochemical and immunofluorescent confocal imaging methods, diffuse labelling in the cytoplasm of the syncytiotrophoblasts and weak staining of the trophoblasts were observed, and the amount of PP20/hTPK decreased from the first trimester to the end of gestation. A 3D model of PP20/hTPK was computed (PDB No.: 1OLY) by homology modelling. A high degree of structural homology showed that the thiamin binding site was highly similar to that of the mouse enzyme, but highly different from the bacterial ones. Comparison of the catalytic centre sequences revealed differences, raising the possibility of designing new drugs which specifically inhibit bacterial and fungal enzymes without affecting PP20/hTPK and offering the possibility for safe antimicrobial therapy during pregnancy. PMID- 15664408 TI - The effects of acute nutrient restriction in the mid-gestational ewe on maternal and fetal nutrient status, the expression of placental growth factors and fetal growth. AB - This study explores the hypothesis that acute under-nutrition in mid-gestation reduces maternal and fetal nutrient status and affects the expression of specific regulators of placental growth and function. Welsh Mountain ewes were fed a concentrate diet plus wheat straw to provide 100% of their maintenance requirements. The concentrate ration of nutrient restricted (NR) ewes was reduced from day (d) 83 of gestation and withdrawn from d85 to d90. At d90, half the ewes (NR m = 7, control n = 8) were euthanased. The remainder (NR n = 9, control n = 9) were fed their maintenance diet until slaughter at d135. Maternal plasma insulin and IGF-I concentrations decreased during nutrient restriction and NEFA concentrations increased. Fetal IGF-I and insulin concentrations were unaltered by maternal diet. Placental VEGF mRNA expression was reduced at d90 (P < 0.05). IGFBP-3 and IGFBP-2 mRNA expression was reduced at d90 (P < 0.05) and d135 (P < 0.05), respectively. Placental weight was significantly lower in NR ewes at d90 (P < 0.05) and the distribution of placentomes shifted towards the everted phenotype at d135 (P < 0.05). Reduced thoracic girth and uterine fluid volume at d90 (P < 0.05) and decreased fetal lung weight at d90 (P < 0.05) and d135 (P < 0.05) suggest spatial limitation of lung expansion. In summary, acute NR in mid gestation reduced anabolic drive and mobilised lipid stores in the maternal compartment, whilst fetal nutrient status was maintained. This was accompanied by changes in placental VEGF and IGFBP expression. The growth of the fetal lung appears to have been compromised and this may have adverse consequences for subsequent neonatal respiratory function. PMID- 15664410 TI - Leptin expression in the fetus and placenta during mouse pregnancy. AB - During pregnancy, leptin concentrations in the maternal circulation are elevated in both humans and rodents but decrease to pre-pregnancy levels at birth, suggesting a role for leptin in the maintenance of pregnancy. Synthesis of leptin by the human placenta is established but whether the murine placenta synthesizes leptin remains controversial. The aims of this study were to determine (a) if the mouse wild-type placenta expresses the ob gene using Reverse Transcription Polymerase Chain Reaction (RT-PCR) and (b) whether the mouse fetus and placenta contribute to the significant increase of leptin in the maternal circulation during pregnancy. The mouse placenta did not express the ob gene at a level that could be readily detected using RT-PCR. Moreover, both maternal gain in weight and undetectable concentrations of leptin in sera in leptin-deficient ob/ob mothers bearing heterozygote (ob/+) fetuses suggested that the mouse fetus and placenta do not make a significant contribution to the dramatic increase in maternal plasma concentrations of leptin during late gestation. It is therefore concluded that neither fetal- nor placental-derived leptin modulates maternal weight gain during pregnancy. PMID- 15664411 TI - Increased biological oxidation and reduced anti-oxidant enzyme activity in pre eclamptic placentae. AB - Oxidative stress occurs when cellular levels of reactive oxygen species exceed anti-oxidant capabilities and has been implicated in the pathogenesis of pre eclampsia. In this study we have examined the tissue levels of endogenous anti oxidant proteins (superoxide dismutase, glutathione peroxidase, thioredoxin reductase and thioredoxin) and the level of lipid and protein oxidation in placental samples from normal and pre-eclamptic pregnancies. Pre-eclamptic tissue homogenates demonstrated significantly increased levels of lipid peroxidation (20.68 +/- 7.811 microM protein versus 5.33 +/- 4.03 microM/mg protein, P < 0.001) and a trended increase in protein carbonyl concentration (248.1 +/- 97.71 units/mg protein versus 209.7 +/- 82.6 U/mg protein) when compared to controls. The levels and activities of the anti-oxidant proteins superoxide dismutase (2.48 +/- 0.6 U/mg protein versus 2.02 +/- 0.51 U/mg protein, P <0.02), thioredoxin reductase (19.25 +/- 9.81 U/mg protein versus 13.02 +/- 5.66 U/mg protein,P = 0.02), thioredoxin (107.00 +/- 18.11 ng/mg protein versus 91.12 +/- 21.18 ng/mg protein, P = 0.02) and glutathione peroxidase (17.33 +/- 6.63 mmol/min/mg protein versus 11.50 +/- 3.11 mmol/min/mg, P < 0.02) were all found to be significantly reduced when comparing pre-eclamptic placental tissue homogenates to gestational age-matched control placentae from non-pre-eclamptic pregnancies. The results of this study demonstrate a decreased enzymatic anti-oxidant capacity and increased oxidation in placental tissue from pre-eclamptic women, which may contribute to the pathogenesis of this complex disorder. PMID- 15664412 TI - A comparative study of the effect of three different syncytiotrophoblast micro particles preparations on endothelial cells. AB - Pre-eclampsia is a pregnancy-associated multi-system disorder of unknown etiology, characterized by damage to the maternal endothelium. The latter facet has been suggested to be mediated in part by elevated shedding of inflammatory placental syncytiotrophoblast micro-particles (STBM) into the maternal circulation. In this study, we have examined STBM prepared by three different methods: mechanical dissection, in vitro placental explant culture and perfusion of placental cotyledons. All three preparations yielded morphologically similar STBM, as confirmed by scanning electron microscopy, and all contained syncytiotrophoblast-specific proteins as determined by the presence of placental alkaline phosphatase. The functional properties of the three STBM preparations were examined on human umbilical vein endothelial cells (HUVEC), where the mechanically prepared particles were found to inhibit proliferation to the greatest extent. Furthermore, only mechanically prepared STBM lead to the detachment and apoptosis of HUVEC cells. Our study, therefore, suggests that STBM prepared from placental perfusion or in vitro explant culture are biologically different from mechanically prepared ones, and may provide a better approximation of physiologically produced placental micro-particles. PMID- 15664413 TI - Expression of GLUT12 in the fetal membranes of the human placenta. AB - The aim of this study was to characterize the expression of the novel glucose transporter GLUT12 in the fetal membranes of the human placenta. RT-PCR and Western blotting of extracts of amnion and choriodecidua from four normal term placentas identified GLUT12 mRNA and protein expression. In all four samples the signals for GLUT12 were markedly stronger in the choriodecidua than in the amnion, whereas the signals for GLUT1, a glucose transporter know to be expressed in fetal membranes, were similar for the two tissues. In further studies, paraffin sections of fetal membranes were analyzed by immunohistochemistry with GLUT12 and GLUT1-specific polyclonal antibodies. GLUT12 immunoreactivity was localized predominantly to the trophoblast cells in the chorion and to a lesser extent to decidual cells and to epithelial and fibroblast cells of the amnion. GLUT1 was localized to chorionic trophoblast cells and amniotic epithelial and fibroblast cells. GLUT12 expression was predominantly cytoplasmic, whereas GLUT1 was associated with the membrane of the cells. These results show that GLUT12 is expressed in cells of human fetal membranes and suggest that GLUT12 may play a role in the facilitation of glucose transport into these cells. PMID- 15664414 TI - The expression of Cdk inhibitors p27kip1 and p57kip2 in mouse placenta and human choriocarcinoma JEG-3 cells. AB - This study was to investigate the expression of Cdk inhibitors p27kip1 and p57kip2 during the development of mouse placenta and during the steroid-treated culture of human choriocarcinoma JEG-3 cells. The p27kip1 mRNA in mouse placenta was highly expressed in 18 days p.c. than that in other groups. But, p57kip2 mRNA expression was high in 12, 14, and 16 days p.c., then decreased in 18 days p.c. The p27kip1 protein expression pattern was similar to mRNA. But, p57kip2 expression was higher in 14 days p.c. than that in other groups. The p27kip1 protein in mouse placenta was gradually increased in labyrinth zone from 12 days to 18 days p.c. However, p57kip2 protein was slightly decreased in labyrinth zone from 12 days to 18 days p.c. These reverse patterns of p27kip1 and p57kip2 expression were also shown in decidua and spongiotrophoblast. The p27kip1 mRNA expression was very low in human choriocarcinoma JEG-3 cells with estradiol concentration-independent manner. In 5 and 50 ng DEX-treated groups, p27kip1 mRNA was dramatically increased in comparison with control groups. The p57kip2 mRNA was not detected in JEG-3 cells. This result shows that p27kip1 may play a role in late period of mouse placental development and p57kip2 may play a role in middle period of mouse placental development, and that p27kip1 may play a role in growth inhibition of human choriocarcinoma cells and could be up-regulated by DEX in human choriocarcinoma. PMID- 15664415 TI - Rho-associated protein kinase II (rock II) expression in normal and preeclamptic human placentas. AB - Rho-associated kinases are serine/threonine kinases that have several functions which might contribute to various physiological and pathological states in cells. There are two isoforms of these enzymes known as rho-associated kinase I (rock I, ROKbeta) and II (rock II, ROKalpha). It has been shown that rock II may be a potential mediator of apoptosis. In addition, rocks play a crucial role in the formation of microvilli-like structures. Increased placental apoptosis and microvilli shedding were shown in preeclampsia. We hypothesized that altered rock expression may lead to the pathologies seen in preeclampsia. Therefore in this study we compared placental rock II expression between normotensive and preeclamptic women using immunohistochemistry and Western blot. Immunohistochemistry and Western blot experiments revealed that rock II is mainly localized in syncytiotrophoblast cells of the placental villi and increased in preeclampsia. In addition to 160 kDa rock II molecule we also detected biologically relevant smaller active form of rock II in preeclamptic but not in normal placentas. We suggest that increased rock II expression in preeclamptic placentas may contribute to etiology or pathogenesis of this syndrome. However, it is considered that rock II expression may also increase to compensate for placental functions changing in preeclampsia. PMID- 15664416 TI - Placental alkaline phosphatase (PLAP) study in diabetic human placental villi infected with Trypanosoma cruzi. AB - Previous work has demonstrated that PLAP activity decreases in serum and placental villi from term chagasic and diabetic pregnant women. In vitro, T. cruzi induces changes in human syncytiotrophoblast's PLAP. Our aim was to determine if infection with T. cruzi induces changes in PLAP activity in diabetic and chagasic women's placenta, in order to elucidate if PLAP plays a role in the mechanisms of interaction between placenta and T. cruzi, and whether hyperglycemic conditions could worsen the placental infection. Using zymogrammes, Western blot, biochemical and immunohistological techniques, PLAP activity was determined in placental villi from diabetic and chagasic women, and in normal placentas cultured under hyperglycemic conditions with or without trypomastigotes. A significant reduction of PLAP expression was immunologically detected in infected diabetic and normal placental villi cultured under hyperglycemic conditions of 71 and 81%, respectively, compared with controls. A significant decrease of PLAP specific activity was registered in homogenates and in the culture media from both infected diabetic and normal placentas under hyperglycemic conditions (of about 50-70%), and in chagasic ones (of about 87%), when compared with controls. Thus, PLAP might be involved in parasite invasion and diabetic and hyperglycemic placentas could be more susceptible to T. cruzi infection. PMID- 15664417 TI - Gestational profile of Na+/H+ exchanger and Cl-/HCO3- anion exchanger mRNA expression in placenta using real-time QPCR. AB - The onset of maternal blood flow (10-12 weeks gestation) results in increased oxygenation of the placenta. We investigated whether the expressions of Na+/H+ exchanger (NHE) and Cl-/HCO3- anion exchanger (AE), thought to have an important role in maintaining intracellular pH of the syncytiotrophoblast and fetal pH homeostasis, are altered at the same time as this increase in blood flow. Real time quantitative PCR was used to examine steady state levels of NHE (NHE1, 2, 3) and AE (AE1, 2) mRNA expression in early (6-9 weeks) and late (10-13 weeks) first trimester and full-term (38-40 weeks) placentas. beta-Actin, IF2B and GAPDH mRNA was also measured. None of the genes showed a significant difference in expression between the early and late first trimester groups. However, NHE2 (p < 0.001) and GAPDH (p < 0.05) mRNA expression significantly increased 18- and 3.7 fold between early first trimester and term. In conclusion, this study provides additional evidence that GAPDH is an unsuitable housekeeping gene for normalization of transcript levels in placenta. The expression of NHE and AE in the villous placenta is not altered concomitant with the onset of maternal blood flow. However, NHE2 transcripts appear to be gestationally regulated, which may contribute to changes in NHE activity. PMID- 15664418 TI - Localization of ubiquitin carboxyl-terminal hydrolase-L1 in cynomolgus monkey placentas. AB - Ubiquitin carboxyl-terminal hydrolase-L1 (UCH-L1) is a restrictedly expressed enzyme in neural and reproductive tissues, and it is considered to have a significant role in reproduction. In the present study, we investigated the localization of UCH-L1 in placenta of cynomolgus monkeys (Macaca fascicularis). UCH-L1 protein was detected in cytotrophoblasts of chorionic plate and villi, and decidual cells of decidua basalis in cynomolgus monkey placenta, and the amount of UCH-L1 protein in whole placenta increased as pregnancy progressed. These results supported that UCH-L1 is necessary for placental and fetal development in primate placenta. This is the first report to demonstrate the presence of UCH-L1 in primate placenta, and the cynomolgus monkey may be a useful model for the study of the functions of the ubiquitin-proteasome system in human pregnancy. PMID- 15664422 TI - Mandated reporting: a policy without reason. PMID- 15664423 TI - Definition and measurement of neglectful behavior: some principles and guidelines. PMID- 15664425 TI - Suicidal phenomena and abuse in adolescents: a review of epidemiological studies. AB - OBJECTIVE: To conduct a systematic review of the international literature to determine whether adolescents who had experienced abuse were more likely to also experience suicidal thoughts and behaviors than other adolescents, and to investigate the nature of this association. METHOD: Reports of community- or school-based studies of adolescents on the association between the prevalence of suicidal phenomena and abuse were searched for via electronic databases, reviewing reference lists and contacting experts in the field. RESULTS: Ten relevant studies were identified. Adolescents who had been physically or sexually abused were significantly more likely to experience suicidal thoughts and behaviors than other adolescents. This association appeared to be direct in most studies in which multivariate analyses were conducted. There is evidence that other factors (e.g., low self-esteem) may play a mediating role in the relationship between abuse and suicidal phenomena. CONCLUSIONS: There is a clear link between abuse and suicidal phenomena occurring during childhood and adolescence. PMID- 15664424 TI - The long-term effects of war experiences on children's depression in the Republic of Croatia. AB - OBJECTIVE: The aim of the study was to investigate whether different levels of depressive symptoms in early adolescent boys and girls could be predicted on the basis of war experiences, perceived available social support (instrumental support, support to self-esteem, belonging and acceptance) and extraversion. METHODS: The sample consisted of 583 children ages 12 to 15 years; 283 children were displaced from different parts of Croatia for a period of approximately three and a half years. The following instruments were administered: Questionnaire on Children's Stressful and Traumatic War Experiences, Reynolds Adolescent Depression Scale, Junior Eysenck Personality Questionnaire, and Interpersonal Support Evaluation List. RESULTS: Regression analyses showed that more war experiences were related to more depressive symptoms for boys only. The greater extent of perceived available social support for boys (instrumental support, support to self-esteem, belonging and acceptance) related to fewer depressive symptoms. For girls, perceived instrumental support and self-esteem were related to fewer depressive symptoms. Predictors in the boys' sample accounted for 35% of the variance in the results on the depression scale, and 27% in the girls' sample. CONCLUSIONS: We conclude that boys suffer more from the long-term effects of war than girls. In situations where children cannot be shielded from stressful events, such as war, a greater level of perceived social support is related to fewer depressive symptoms both for boys and girls in early adolescence. PMID- 15664426 TI - Caregivers' readiness for change: predictive validity in a child welfare sample. AB - OBJECTIVE: To assess the predictive validity of continuous measures of problem recognition (PR), intentions to change (ITC), and overall readiness for change (RFC) among primary caregivers who received in-home services following substantiated reports of child abuse or neglect. METHOD: A modified version of the University of Rhode Island Change Assessment scale was included in interviews with a sample of 353 primary caregivers at 4 weeks, 16 weeks, and 1 year after referral for in-home services. Additional data were obtained from administrative records and caseworker surveys. Hierarchical linear and nonlinear models were used to assess relationships between PR, ITC, RFC and changes over time in measures of individual and family functioning (e.g., parenting behaviors, children's behaviors, housing and economic problems, social support, and life events). Bivariate probit regression analysis was used to examine relationships between PR, ITC, RFC and the likelihood of subsequent reports of child maltreatment and out-of-home placements within 1 year after referral. RESULTS: Initial problem recognition and intentions to change predict a few improvements in individual and family functioning, along with significant reductions in the likelihood of additional reports of child maltreatment within 1 year. Initial intentions to change also predict reductions in the substantiation of subsequent reports of maltreatment. An overall measure of readiness for change predicts reductions in the likelihood of out-of-home placement. CONCLUSIONS: Problem recognition and intentions to change predict somewhat different outcomes; hence, there are few advantages of a combined measure of readiness for change. Further inquiry is needed to determine whether and how these associations are mediated by intervention processes or other factors in child welfare services populations. PMID- 15664427 TI - Post-traumatic stress disorder among school age Palestinian children. AB - OBJECTIVES: This study was designed to assess the prevalence of PTSD among Palestinian school-age children. Variables that distinguish PTSD and non-PTSD children were examined, including child characteristics, socioeconomic status, family environment, and parental style of influence. METHOD: Participants were 1,000 children aged 12 to 16 years. They were selected from governmental, private, and United Nations Relief Work Agency (UNRWA) schools in East Jerusalem and various governorates in the West Bank. Questionnaires were administered in an interview format with children at school, and with the available parent at home. RESULTS: A substantial number of children experienced at least one lifetime trauma (54.7%). Post-traumatic stress disorder (PTSD) was diagnosed in 34.1% of the children, most of whom were refugees, males, and working. Although the expected association between family environment, parental style of influence and PTSD symptomatoplogy was found in this study, family ambiance (child's experience of anxiety in home environment) was the only predictor in the final model. CONCLUSIONS: The results stress the importance of assessing PTSD in schools settings. PMID- 15664428 TI - Effect of TCDD on mRNA expression of genes encoding bHLH/PAS proteins in rat hypothalamus. AB - 2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) brings about a wide variety of toxic and biochemical effects via an AH receptor (AHR)-mediated signalling pathway. Wasting syndrome and acute lethality are TCDD-induced endpoints showing a striking sensitivity difference between two rat strains, TCDD-sensitive Long Evans (Turku/AB) (L-E) and TCDD-resistant Han/Wistar (Kuopio) (H/W). These rat strains were used to study hypothalamic effects of TCDD on expression of genes encoding AHR-regulated bHLH/PAS proteins potentially involved in molecular pathogenesis of the wasting syndrome. In addition, two well-established target genes of TCDD, CYP1A1 and CYP1A2 were also examined. Quantitative RT-PCR was used to measure mRNA levels in hypothalamus, which is a major center of food intake and body weight regulation. At both 6 and 96 h after a single dose of 50 microg/kg TCDD, significant elevations were found in mRNA levels of AHR repressor (AHRR), CYP1A1 and CYP1A2, but not those of AHR, ARNT or ARNT2. Likewise, TCDD (100 microg/kg) did not alter the expression of SIM1, implicated in the suppressive impact of TCDD on food intake, nor that of PER2, involved in regulation of circadian rhythms. Differences between H/W and L-E rats appeared in constitutive levels of AHR and ARNT and in TCDD-induced levels of CYP1A2, AHRR, AHR and ARNT, which all were about two- to four-fold lower in H/W rats. Thus, although the changes found do not account for the wasting syndrome, expression of all principal genes of the AHR-signalling pathway in rat hypothalamus make it a candidate target for TCDD. PMID- 15664429 TI - Carbofuran-induced oxidative stress in slow and fast skeletal muscles: prevention by memantine and atropine. AB - Acute toxic effects of acetylcholinesterase (AChE) inhibitors on skeletal muscles are thought to involve oxidative stress with increased generation of free radicals such as reactive oxygen species (ROS) and reactive nitrogen species (RNS). Muscle hyperactivity with its increased oxygen and energy consumption appear to be the primary cause of oxidative stress. The present investigation was therefore undertaken to establish the normal levels of F(2)-isoprostanes (F(2) IsoPs, specific markers of ROS/oxidative stress), citrulline (determinant of NO/NOS and marker of RNS), and high-energy phosphates (HEP: adenosine triphosphate, ATP and phosphocreatine, PCr) in slow (soleus) and fast (extensor digitorum longus, EDL) muscles of rats. In addition, we aimed to determine if memantine HCl (MEM), in combination with atropine sulfate (ATS), prevents carbofuran-induced changes in markers of oxidative stress. Control values were not significantly different for F(2)-IsoPs (1.142 +/- 0.027 and 1.177 +/- 0.092 ng/g) and citrulline (469.7 +/- 31.8 and 417.8 +/- 18.5 nmol/g) in soleus and EDL muscles, while the values were different for HEP (ATP, 3.66 +/- 0.11 and 5.85 +/- 0.14 micromol/g; PCr, 7.91 +/- 0.26 and 13.14 +/- 0.31 micromol/g). Rats acutely intoxicated with carbofuran (1.5 mg/kg, s.c.) showed the signs of maximal toxicity including muscle hyperactivity within 60 min of exposure. At this time, F(2)-IsoPs (177 and 153%) and citrulline (267 and 304%) levels were significantly increased, while ATP (46 and 43%) and PCr (44 and 46%) levels were decreased in soleus and EDL, respectively. Rats pretreated with MEM (18 mg/kg, s.c.) and ATS (16 mg/kg, s.c.), 60 and 15 min prior to carbofuran, respectively, showed no signs of toxicity. MEM in combination with ATS protected muscles from carbofuran induced hyperactivity and attenuated increases in F(2)-IsoPs and citrulline, and depletion of HEP. Carbofuran-induced changes and protection by MEM and ATS were of similar magnitude in both muscles. These findings indicate that carbofuran induced muscle hyperactivity produces oxidative stress as measured by increased ROS and RNS generation, and HEP depletion. MEM and ATS prevent the carbofuran induced chain of events involved in oxidative stress. PMID- 15664430 TI - Prospective protective role of melatonin against arsenic-induced metabolic toxicity in Wistar rats. AB - Subchronic exposure to arsenic is associated with alteration of glucose homeostasis. Arsenic treatment (as sodium arsenite) of male Wistar rats (weighing 130-150 g) at a dose of 5.55 mg kg(-1) body weight (equivalent to 35% of LD(50)) (i.p.) per day for a period of 30 days produced hypoglycemia, with associated increased urinary excretion of glucose and depletion of liver glycogen and pyruvic acid contents. Mobilization of free amino acids from kidney to liver was facilitated by arsenic treatment. Arsenic exposure significantly decreased the glutamate-pyruvate transaminase activity in kidney. Glucose 6-phosphatase activity in liver tissue was also significantly decreased after arsenic treatment. In addition to these, liver lactate dehydrogenase activity was elevated due to arsenic treatment. Melatonin supplementation (i.p.) at a dose of 10 mg kg(-1) day(-1) for last five days prior to sacrifice reversed most of the above changes caused by arsenic. Melatonin, being a potent free radical scavenger may reduce arsenic-induced free radical production, and thereby, eliminating its toxic effects. So, arsenic-induced hypoglycemia, with associated glycogenolytic as well as glycolytic activities of liver can be partially counteracted by melatonin supplementation. Accordingly, it may be suggested that melatonin can serve as a prospective protective agent against arsenic-induced metabolic toxicity. PMID- 15664431 TI - Down-regulation of GAT-1 mRNA expression in the microdissected hypothalamic medial preoptic area of rat offspring exposed maternally to ethinylestradiol. AB - Steroid hormones are powerful regulators of gene transcription in the brain and have the potential to permanently alter the structure and function of the developing brain. Steroid-mediated altered gene expression may thus be responsible for the molecular cascade for sexual differentiation. In this study, to assess effects of maternal exposure to ethinylestradiol (EE) on brain sexual differentiation of offspring, region-specific mRNA expression of two estrogen responsive genes, gamma-aminobutyric acid transporter type 1 (GAT-1) and anti apoptotic bcl-xL was measured in the medial preoptic area (MPOA), including sexually dimorphic nucleus (SDN), at the late stage of brain sexual differentiation in rats. Pregnant Sprague-Dawley animals were fed diets containing EE at concentrations of 0, 0.02, 0.1, and 0.5 ppm from day 15 of pregnancy to day 9 after delivery. In another group, neonates were directly injected with estradiol benzoate (EB: 10 microg/pup, sc) on postnatal day (PND) 2. The MPOA on PND 9 was microdissected from methacarn-fixed paraffin-embedded brain sections to measure mRNA levels by competitive RT-PCR, followed by plate hybridization. EE-exposure decreased GAT-1 expression dose-dependently from 0.02 ppm in females and at 0.5 ppm in males, while EB-treatment caused reduction only in females. EE-exposure did not alter Bcl-xL levels. At week 11, EE-exposed females exhibited a similar spectrum of histopathological changes in endocrine linked organs as with EB, evident from 0.1 ppm, while in males EE-exposure did not cause histopathological alteration despite clear change with EB-treatment. Measurement of SDN-POA dimensions at week 11 revealed volume reduction in males exposed to 0.5 ppm EE or EB. The results suggest that GAT-1 expression in the developing MPOA is a sensitive measure for the level of disruption of brain sexual differentiation due to maternal dietary exposure to estrogens, despite definite reproductive abnormalities may not be detectable in males with this exposure protocol. PMID- 15664432 TI - Effects of butylated hydroxyanisole on the development and functions of reproductive system in rats. AB - Butylated hydroxyanisol (BHA) is a widely used antioxidant for long preservation of food products, cosmetics and pharmaceuticals. Although BHA is generally recognized as safe, it is classified as a suspected endocrine-disrupting compound. We investigated the effects of BHA on reproductive function and development by the treatment of mature male and female SD rats (F0) through pre gestation, gestation and lactation period and of their offspring (F1) until 13 weeks old via gavage with BHA 0 (corn oil, vehicle control), 10, 100 and 500 mg/kg bw/day. Organ weights of liver, adrenal gland and thyroid gland of F0 rats were increased by BHA 500 mg/kg but those of spleen and ventral prostate were decreased without significant difference in terminal body weight. Reduced serum testosterone and thyroxine (T4) were observed with dose-dependent manner in F0 male rats. Mating rate was decreased and cohabitation duration for conception was longer without differences in the number, motility and morphology of sperm by BHA 500 mg/kg. Body weight of F1 offspring was significantly decreased with change of relative weight of liver and brain by BHA 500 mg/kg at PND21. Sexual maturation indicated by vaginal opening and preputial separation was delayed by BHA 500 mg/kg. The weights of liver and adrenal gland were increased while those of spleen, vagina, testes and ventral prostate were decreased in F1 rats exposed to BHA 100 or 500 mg/kg for 13 weeks. Also, BHA 500 mg/kg reduced the velocity of sperm motion and number with smaller-sized sperm head in F1 male rats and slightly shortened estrous cycle length with higher frequency of estrus and lower frequency of diestrus stages in F1 female rats. Lower serum T4 and testosterone contents with higher serum cholesterol levels were also observed by BHA 500 mg/kg. Increased follicular cell height, and exfoliated and vacuolated follicular epithelial cells were observed in thyroids of F1 female and males rats exposed to BHA 500 mg/kg. This study elucidates that high dose of BHA induce weak dysfunction and underdevelopment of reproductive system of male and female rats with the change of T4 and testosterone levels, sex organ weights and sexual maturation and histological lesions of thyroid gland. PMID- 15664433 TI - Evaluation of the clinical, immunologic, and biochemical effects of nitroso sulfamethoxazole administration to dogs: a pilot study. AB - Sulfonamide antimicrobials such as sulfamethoxazole (SMX) have been associated in humans with hypersensitivity reactions, to include fever, skin eruptions, hepatotoxicity, and blood dyscrasias. These reactions also occur in dogs, the only non-human species known to develop a similar spectrum of sulfonamide hypersensitivity. Sulfonamide hypersensitivity is not well understood, but has been hypothesized to be due to the generation of the reactive oxidative metabolite, nitroso sulfamethoxazole (SMX-NO). SMX-NO, unlike the parent sulfonamide, is cytotoxic in vitro, haptenizes tissue proteins, and is immunogenic in rodents. The purpose of this pilot study was to determine whether SMX-NO, when administered to dogs, would lead to drug-tissue adducts, anti-drug antibodies, antioxidant depletion, or clinical evidence of drug hypersensitivity. Four dogs were randomized to one of four treatments: SMX-NO 1 mg/kg; SMX-NO 3 mg/kg; SMX-NO 10 mg/kg; or vehicle control. Dosing was by the intraperitoneal route, once daily for four consecutive days per week, for 2 weeks total, followed by a third week of observation. Following this, all dogs were challenged with trimethoprim-sulfamethoxazole, 25 mg/kg for 12 h for 2 weeks. No dog developed clinical or biochemical evidence of drug hypersensitivity. Plasma cysteine and leukocyte reduced glutathione were not depleted during dosing; however, ascorbate was significantly depleted by week 2 following SMX-NO at 10 mg/kg. Anti-SMX antibodies (IgG or IgM by ELISA) were not detected in any dogs at any time points. SMX-hemoglobin adducts were detected in the spleen in SMX-NO dosed dogs; however, these adducts were not accompanied by an immunologic or systemic response. The results of this pilot study indicate that SMX-NO dosing in dogs, using a dosing protocol shown to be immunogenic in other species, produces modest ascorbate depletion and hemoglobin adduct formation, but is insufficient to produce an immunologic response or a clinical syndrome of sulfonamide hypersensitivity in this susceptible species. PMID- 15664434 TI - Different cytoprotective effect of antioxidants and change in the iron regulatory system in rodent cells exposed to paraquat or formaldehyde. AB - To study the mechanism of toxicity of paraquat and formaldehyde, the response of oxidant-exposed cultured NIH3T3 cells to antioxidants or an iron chelator was investigated. Paraquat-induced cell death was reduced by treatment with 10 microM pyrrolidine dithiocarbamate (PDTC) and 10 microM desferrioxamine (DFO), but not with N-acetyl-L-cysteine (NAC). Cells were protected from formaldehyde-induced cytotoxicity by 1 mM NAC, but not by PDTC or DFO. Moreover, paraquat modulated the cellular iron regulatory system. Paraquat induced a time-dependent increase in the binding of iron regulatory protein 1 (IRP1) to iron-responsive element (IRE), and the enhanced IRP1 activity continued over 24 h. On the other hand, no induction of increased IRP1 binding to IRE was observed in rodent cells exposed to formaldehyde. Previously, we observed stimulation of EpRE-mediated ferritin mRNA expression in the cells exposed to hydrogen peroxide. However, paraquat did not induce any transcriptional activation of ferritin genes. These results suggest that intracellular iron may be involved in paraquat-mediated cytotoxicity and the influence of paraquat on iron metabolism differs from that of hydrogen peroxide. PMID- 15664435 TI - Chemoprotective and toxic potentials of synthetic and natural chalcones and dihydrochalcones in vitro. AB - Cytochrome P4501A activity, oxidative stress and inhibition of gap junctional intercellular communication (GJIC) are involved in metabolic activation of promutagens and tumor-promoting activity of various xenobiotics, and their prevention is considered to be an important characteristic of chemoprotective compounds. In this study, a series of 31 chalcones and their corresponding dihydroderivatives, substituted in 2,2'-, 3,3'-, 4- or 4'-position by hydroxyl or methoxy group, were tested for their ability to inhibit Fe(II)/NADPH-enhanced lipid peroxidation and cytochrome P4501A-dependent 7-cethoxyresorufin-O deethylase (EROD) activity in rat hepatic microsomes. Effects of the compounds on GJIC were determined in rat liver epithelial WB-F344 cells. Most of the chalcones and dihydrochalcones inhibited EROD activity in a dose-dependent manner at the range 0.25-25 microM, which was comparable to model flavonoid inhibitors alpha naphthoflavone and quercetin. The chalcones exhibited higher inhibition activity than the corresponding dihydroderivatives. Mono and dihydroxylated chalcones, and dihydrochalcones showed none or only a weak antioxidant activity; trihydroxyderivatives inhibited in vitro lipid peroxidation significantly only at 50 microM concentration. Potential adverse effects, namely inhibition of GJIC and/or cytotoxicity were detected after treatment of WB-F344 cells with a number of chalcone and dihydrochalcone derivatives, suggesting that they should be excluded from additional screening as chemoprotective compounds. Chalcones and dihydrochalcones substituted at 4- and/or 4'-position, which elicited no inhibition of GJIC, were further tested for the potential enhancing effects on GJIC. The present data seem to suggest that 4-hydroxy, 2',4'-dihydroxy-3-methoxy, 2,4,4'-trihydroxy, and 2',4,4'-trihydroxychalcone, 2',4-dihydroxy and 2'-hydroxy 3,4-dimethoxydihydrochalcone might be promising chemoprotective compounds against CYP1A activity, and partly also against oxidative damage without inducing adverse effects, such as GJIC inhibition. In general, determination of potencies of tested compounds to inhibit GJIC should be involved in any set of methods for the in vitro screening of chemoprotective characteristics of potential drugs, in order to reveal their potential adverse effects associated with tumor promotion. PMID- 15664436 TI - Inhibition of cyclooxygenase activity, platelet aggregation and thromboxane B2 production by two environmental toxicants: m- and o-cresol. AB - Cresol is a well-known environmental pollutant, toluene metabolite, uremic toxicant and accidental poisoning product. Formocresol, a preparation of formalin and cresol, is also used as a root canal medicament and for pulpotomy of primary teeth. However, little is known about its effect on cardiovascular system. In this study, m-cresol inhibited the AA-induced platelet aggregation by 43-97% at concentrations ranging from 0.25 to 1 mM. Collagen-induced platelet aggregation was also inhibited by 0.25-1 mM of m-cresol by 47-98%. Accordingly, o-cresol (0.1 0.5 mM) also inhibited the AA-induced platelet aggregation by 46-96% and the collagen-induced platelet aggregation by 35-88% at concentrations of 0.1-1 mM. AA and collagen-induced platelet thromboxane B(2) (TXB(2)) production was inhibited by even 0.1 mM of m-cresol with 88 and 54% of inhibition, respectively. The o cresol (0.1 mM) also inhibited the AA- and collagen-induced platelet TXB(2) production with 91 and 97% respectively. Although m- and o-cresol (<1 mM) showed little effect on thrombin-induced platelet aggregation, they effectively inhibited the thrombin-induced platelet TXB(2) production. The m-cresol (2 and 5 mM) inhibited the COX-1 activity by 55-99%, but showed little effect on COX-2 enzyme activity. Moreover, o-cresol (0.5 and 1 mM) inhibited the COX-1 activity by 40-95%. COX-2 enzyme activity was inhibited by 68% at a concentration of 5 mM o-cresol. These results indicate that acute cresol-poisoning, direct root canal medication with formocresol or long-term occupational exposure to cresol and toluene may potentially suppress blood clot formation and lead to tissue hemorrhage via inhibition of platelet aggregation, TXB(2) production and COX enzyme activity. PMID- 15664438 TI - Evaluation of anti-androgenic activity of di-(2-ethylhexyl)phthalate. AB - DEHP is a widely used platiciser in the manufacture of PVC-based materials. It is known to disrupt the reproductive tract development in male rats. We have performed the Hershberger assay with DEHP on an immature castrated rat model to check if DEHP antagonise the testosterone propionate androgenic effect on the accessory sex organs development. DEHP significantly decreased the BC/LA muscles, the prostate, and the seminal vesicles relative weights from 100, 200, and 400 mg/kg bw/day, respectively. DEHP increased the liver relative weight from 100 mg/kg bw/day. A study was also performed on MDA-MB453 cell line stably transfected with pMMTVneo-Luc with DEHP and its major metabolites (MEHP and metabolites VI and IX) to identify anti-androgenic activity. Neither DEHP nor MEHP antagonised DHT activity in the MDA-MB453 transfected cells. In contrast, metabolites VI and IX were anti-androgenic in vitro. DEHP appeared not to be a 5alpha-reductase inhibitor and acted in an independent mechanism from the testicular production in the young rat. PMID- 15664437 TI - Evaluation of particle translocation across the alveolo-capillary barrier in isolated perfused rabbit lung model. AB - Particulate air pollution is associated with respiratory and cardiovascular morbidity and mortality. It has been suggested that ultrafine particles are able to translocate from the airways into the bloodstream in vivo. We have investigated this in an isolated perfused and ventilated rabbit lung preparation lacking pulmonary lymphatic flow. Fluorescent polystyrene particles of different diameters (24, 110 or 190 nm) and surface chemistry (carboxylate or amine modified) were injected either intratracheally (i.t.) or intravascularly (i.v.) and, after a period of 2 h, their presence in the perfusion liquid or in the bronchoalveolar lavage (BAL) fluid, was assessed by spectrofluorimetry. Vascular pressures and lung weights were monitored. Following the i.t. administration, no particle translocation was observed from the alveoli into the vascular compartment. Similarly, no particle translocation was found after i.v. administration of particles. However, when microvascular permeability was pharmacologically increased by administering histamine (10(-4) M) in the vascular compartment, inducing a positive driving force provided by fluid filtration, a fluorescent signal in BAL was recorded (2.5 +/- 1% of the dose of particles administered), suggesting a translocation of particles through the alveolo capillary barrier. We conclude that ultrafine polystyrene particles cannot significantly diffuse from lung into the vascular compartment in our model, but they are able to translocate in the opposite direction when the microvascular permeability is increased by histamine. The relevance of these ex vivo findings for the in vivo translocation of inhaled ultrafine particles remains to be established. PMID- 15664439 TI - Effects of P-glycoprotein inhibitors on transepithelial transport of cadmium in cultured renal epithelial cells, LLC-PK1 and LLC-GA5-COL 150. AB - The purpose of this study using LLC-PK(1) cells and P-glycoprotein (P-gp) overexpressed LLC-PK(1) cells (LLC-GA5-COL 150 cells) was to investigate the secretory transport of cadmium (Cd) via endogenous and overexpressed P-gp, respectively. Cell monolayers cultured on permeable membranes were incubated at 37 degrees C for 60 min with 1 microM CdCl(2) from either the apical or the basolateral side. The basolateral-to-apical transport of Cd was 1.7 times higher than the apical-to-basolateral transport of Cd in LLC-GA5-COL 150 cells, while the transport from apical and basolateral sides was almost the same in LLC-PK(1) cells. Treatment with a P-gp monoclonal antibody, UIC2, significantly decreased the basolateral-to-apical transport of Cd in LLC-PK(1) and LLC-GA5-COL 150 cells, and significantly increased the apical-to-basolateral transport of Cd in both cells. The effects of UIC2 were more marked in LLC-GA5-COL 150 cells than in LLC PK(1) cells. Furthermore, typical P-gp inhibitors such as cyclosporin A, and doxorubicin decreased the basolateral-to-apical transport of Cd slightly in LLC PK(1) cells and significantly in LLC-GA5-COL 150 cells. These results suggest that Cd is extruded from the apical membrane of LLC-PK(1) and LLC-GA5-COL 150 cells, probably depending on the level of P-gp expression. PMID- 15664440 TI - Increased levels of metallothionein in placenta of smokers. AB - Experiments were designed to evaluate and compare metallothionein (MT), zinc and cadmium levels in human placentas of smoking and non-smoking women. Smoking was assessed by self-reported cigarette consumption and urine cotinine levels before delivery. Smoking pregnant women with urine cotinine levels higher than 130 ng/ml were included in the smoking group. Determination of placental MT was performed by western blot analysis after tissue homogenization and saturation with cadmium chloride (1000 ppm). Metallothionein was analyzed with a monoclonal antibody raised against MT-1 and MT-2 and with a second anti mouse antibody conjugated to alkaline phosphatase. Zinc and cadmium were determined by neutron activation analysis and atomic absorption spectrometry respectively. Smokers showed higher placental MT and cadmium levels, together with decreased newborn birth weights, as compared to non-smokers. The semi-quantitative analysis of western blots by band densitometry indicated that darker bands corresponded to MT present in smokers' samples. This study confirms that cigarette smoking increases cadmium accumulation in placental tissue and suggests that this element has a stimulatory effect on placental MT production. PMID- 15664441 TI - In vitro genotoxicity of the West African anti-malarial herbal Cryptolepis sanguinolenta and its major alkaloid cryptolepine. AB - Cryptolepine (CLP), the major alkaloid of the West African anti-malarial herbal Cryptolepis sanguinolenta (Periplocaceae) is a DNA intercalator that exhibits potent toxicity to a variety of mammalian cells in vitro. We have hypothesized that the DNA intercalating properties of cryptolepine could trigger genetic damage in mammalian cells. The objective of the present study was therefore to assess the ability of both cryptolepine (CLP) and the traditional anti-malarial formulation, the aqueous extract from the roots (CSE) to induce mutation at the hprt locus and micronuclei (MN) formation in V79, a Chinese hamster fibroblast cell line commonly used in genetic toxicity studies. CSE at a high concentration (50 microg/ml) induced an apparent significant ten fold increase in mutant frequency compared to vehicle control (mean of 38 versus 4 mutant clones/10(6) surviving cells) but, this concentration of CSE was very toxic (<15% cell survival). CLP did not appear to be mutagenic in the dosage range used (up to 2.5 microM, equivalent to 1.1 microg/ml). However, after 24h treatment of V79 cells both CSE and CLP induced a dose-dependent increase in micronuclei of 4.15% and 6.43% (25 microg/ml CSE and 2.5 microM, equivalent to 1.1 microg/ml CLP, respectively) compared to 0.36% in vehicle control. These results show that treatment of mammalian cells with CSE and CLP can lead to DNA damage and we suggest that the routine use of CSE and the potential use of CLP derivatives in malaria chemotherapy could carry a genotoxic risk. PMID- 15664443 TI - Trichloroethylene as human carcinogen. PMID- 15664442 TI - The immunosuppressive effect of methylmercury does not preclude development of autoimmunity in genetically susceptible mice. AB - Methylmercury (MeHg) is a common environmental pollutant due to both natural and anthropogenic sources. Although the central nervous system (CNS) is considered the critical organ for the toxic effect of MeHg, it has recently been suggested that the immune system might be at least as sensitive as the CNS. We have examined the effects of MeHg on the immune system in genetically metal susceptible mice. Subcutaneous (sc) injections of 2 mg MeHg/kg body weight (bw) every third day (internal dose ca. 540 microg Hg/kg bw/day) to A.SW mice of the H 2(s) haplotype, caused during the first week a 47 and 9% reduction of B- and T cells, respectively, which indicates immunosuppression. Subsequently, an autoimmune syndrome developed which shared certain features with the syndrome induced by inorganic mercury in H-2(s) mice, including antibodies targeting the 34 kDa nucleolar protein fibrillarin, increased expression of IL-4 mRNA, increase of Th2-type of immunoglobulins (IgE and IgG1), and increased MHC class II expression on B-cells. However, the response using MeHg was attenuated compared with even lower doses of Hg in the form of inorganic mercury, and specifically lacked the increased expression of IL-2 and IFN-gamma mRNA, the polyclonal B-cell activation (PBA), and the systemic immune-complex (IC) deposits which are induced by inorganic mercury. Increasing the dose of MeHg increased the titre of anti nucleolar antibodies and shortened the induction time, but did not lead to stronger immunostimulation or systemic IC-deposits. The kidney and liver selectively accumulated MeHg, while the blood, spleen and lymph nodes showed lower levels of MeHg. The accumulation of MeHg and Hg(2+) increased throughout the 30-day period. The fraction of Hg(2+) in the kidney varied between 4 and 22%, and the lymph nodes showed a maximum of 30% Hg(2+). We conclude first that MeHg has quantitatively different effect on the immune system compared with inorganic mercury, and secondly that an initial immunosuppression induced by a xenobiotic does not preclude subsequent immunostimulation and autoimmunity. PMID- 15664444 TI - Possible carcinogenicity of trichloroethylene in humans. PMID- 15664445 TI - Imaging protein behavior inside the living cell. AB - The genetically encoded fluorescent proteins (FPs) have transformed studies in cell biology by allowing the behavior of proteins to be tracked within the natural environment of the living cell. Progressively more complex imaging methods are being used to measure the mobility, co-localization and interactions of proteins labeled with the FPs. This review provides an overview of recent developments in live-cell imaging techniques to analyze the subcellular distribution and interactions of proteins in living cells. PMID- 15664446 TI - Pulsed estradiol exposure has a limited ability to induce uterine proliferation in ovariectomised female Wistar rats. AB - All post-menopausal hormone replacement therapies (HRT) aim to provide a steady mid-follicular serum concentration of estrogen, with the exception of pulsed estrogen therapy, which concentrates estradiol (E2) exposure in the few hours following administration. This study was carried out to identify and characterise cellular and molecular mechanisms specifically involved in the response of the uterus to pulsed E2. Ovariectomised Wistar rats were treated with E2 for 10 days via IV route to mimic pulsed therapy (1, 4, 10 and 250 microg/kg) or with a subcutaneous pump to mimic standard HRT (10 and 250 microg/kg). Pulsed estrogen therapy effects on uterus was revealed by general E2 sensitivity markers (C3 mRNA, progesterone receptor (PR)) from the lower dose with no over stimulation even at the highest dose conversely to what observed with continuous exposure. Uterotrophic effect of pulsed E2 (uterine weight and epithelium thickness) was observed at all dose administered but with a limited maximal effect comparable to the ranges measurable in sham animals. This data corroborates with proliferating cell nuclear antigen (PCNA) expression in the uterine epithelium used as a marker of proliferation. PCNA was significantly induced after continuous administration but only slightly after pulsed E2 (250 microg/kg). In summary, pulsed E2 leads to a more limited proliferative effect than with continuous E2 in the uterus. PMID- 15664447 TI - Sex specific expression of progesterone receptor in mouse lower urinary tract. AB - Progesterone receptor (PR) was investigated immunohistochemically in the lower urinary tract of the male and female mouse. Estrogen receptor (ER)-subtype deficient mice (ERKO, BERKO) were used to determine the possible regulation of PR expression in an ER-subtype-specific manner. PR was found to be co-expressed with ERalpha in cell nuclei of urothelium, lamina propria fibroblasts and smooth muscle cells in the female urethra. Only few PR positive cells were seen in female ERKO mice. Ovariectomy reduced and estrogen treatment restored the urethral PR expression in female wild type and BERKO mice. Thus, the expression of PR in the female urethra is estrogen-inducible via ERalpha. In male urethra, PR was co-expressed with ERbeta in the rhabdosphincter. In male, no evidence was obtained for the ER-linked control of the PR expression. No PR-positive cells were observed in the body of the bladder of either sex or any strain. PMID- 15664448 TI - Estrogen receptor independent rapid non-genomic effects of environmental estrogens on [Ca2+]i in human breast cancer cells. AB - The aim of this study was to identify and characterize an alternative pathway through which environmental estrogenic compounds may mediate their intracellular effects. Three human breast cancer cell lines were employed including MCF-7 cells, which express both ERalpha and ERbeta; MDA-MB-231 cells, which express ERbeta but not ERalpha; and SKBR-3 cells, which express neither ERalpha nor ERbeta. The effect of environmental estrogenic compounds on intracellular calcium ion concentration ([Ca(2+)](i)) was measured and compared to that of 17beta estradiol (E2). A rapid and maintained increase in [Ca(2+)](i) was observed following the application of nanomolar concentrations of environmental estrogens and E2 regardless of the expression of ERalpha and ERbeta. Removal of extracellular Ca(2+) completely abolished the steroid-induced [Ca(2+)](i) increase. Pre-treatment of cells with the estrogen receptor (ER) antagonist ICI 182,780 had no effect on either basal [Ca(2+)](i) or the steroid-triggered [Ca(2+)](i) response. In summary, we have demonstrated ER independent rapid non genomic effects of environmental estrogenic compounds, at nanomolar concentrations, on [Ca(2+)](i). The results of this study demonstrate an alternative pathway to explain potent intracellular effects of endocrine disrupting chemicals. PMID- 15664449 TI - Sex hormone-binding globulin antagonizes the anti-apoptotic effect of estradiol in breast cancer cells. AB - Sex hormone-binding globulin, the plasma carrier for sex steroids, inhibits the estradiol-induced proliferation of breast cancer cells. Estradiol induces cell proliferation triggering multiple mechanisms. Besides regulating growth factors, it activates Erk-1/-2, thus inhibiting apoptosis. In the present study, we investigated the effect of SHBG on estradiol-mediated anti-apoptotic effect in MCF-7 breast cancer cells. As expected, estradiol reduced the number of cells undergoing apoptosis. Although no modification of estradiol action was observed in cells treated contemporarily with estradiol and SHBG, pre-incubation with SHBG before estradiol treatment contrasted the anti-apoptotic effect completely. A mutant form of SHBG, lacking the O-linked oligosaccharide in Thr(7), displayed no such effect. Moreover, SHBG prevented the estradiol-induced phosphorylation of Erk-1/-2, whereas it had no effect on estradiol-induced transcription. Taken together, our observations suggest that the interaction of SHBG with MCF-7 cell membranes causes inhibition of the anti-apoptotic effect of estradiol which might account for SHBG's inhibitory effect on breast cancer cell growth. PMID- 15664451 TI - KATP-channels in beta-cells in tissue slices are directly modulated by millimolar ATP. AB - In pancreatic beta-cells, inhibition of K(ATP)-channels plays a pivotal role in signal transduction of glucose-induced insulin release. However, the extreme sensitivity of K(ATP)-channels to its ligand ATP as found in inside-out patches is not directly compatible with modulation of these channels at physiological [ATP](i). We studied K(ATP)-channel sensitivity to ATP in beta-cells in dispersed culture and in fresh pancreatic tissue slices. Physiological [ATP](i) blocks more than 99% of K(ATP)-channels in cultured beta-cells, while only 90% in beta-cells in slices, indicating reduced sensitivity to ATP in the fresh slices. Applying cytosolic factors like ADP, phosphatidylinositol-4,5-bisphosphate (PIP(2)) or oleoyl-CoA did not restore the K(ATP)-channel sensitivity in cultured beta-cells. Our data suggest that interaction between SUR1 and Kir6.2 subunit of the K(ATP) channel could be a factor in sensitivity modulation. Tissue slices are the first beta-cell preparation to study direct K(ATP)-channel modulation by physiological [ATP](i). PMID- 15664450 TI - Distinct Grb10 domain requirements for effects on glucose uptake and insulin signaling. AB - The adapter protein Grb10 binds to phosphotyrosine residues in insulin receptors via its C-terminal region and regulates insulin signaling. This study investigated Grb10 regulation of glucose uptake and the importance of the Grb10 N terminal region using 3T3-L1 adipocytes overexpressing full-length (FL-Grb10) or N-terminally truncated Grb10 (BPS-SH2). Overexpression of FL-Grb10 inhibited insulin-stimulated receptor autophosphorylation and glucose uptake. In contrast, the BPS-SH2 fragment of Grb10 had no effect on receptor phosphorylation or glucose uptake. In spite of these differences, both FL-Grb10 and the BPS-SH2 fragment inhibited insulin-stimulated phosphorylation of IRS1, IRS2, Akt/PKB, Shc, ERK1/2, APS, and c-Cbl to a similar extent. Co-precipitation studies demonstrated more sustained binding of the BPS-SH2 fragment than FL-Grb10 to insulin receptors. Although receptor binding domains of Grb10 are sufficient to inhibit insulin effects on proximal post-receptor signaling responses, N-terminal domains of Grb10 are essential for the effects of this adapter protein on receptor phosphorylation and glucose uptake. PMID- 15664452 TI - Regulation of the human vitamin D3 receptor promoter in breast cancer cells is mediated through Sp1 sites. AB - 1,25-Dihydroxyvitamin D(3) (1,25(OH)(2)D(3)), the active form of Vitamin D, mediates gene transcription through the Vitamin D receptor (VDR), a nuclear receptor expressed in multiple normal and transformed cell types. In mammary epithelial cells, including those derived from breast cancers, 1,25(OH)(2)D(3) induces growth arrest and/or apoptosis through VDR dependent mechanisms, and VDR agonists represent potential therapeutic agents for hyperproliferative diseases, including cancer. Since target cell sensitivity to 1,25(OH)(2)D(3) and its analogs reflects VDR expression, understanding the transcriptional regulation of the VDR gene is fundamental to development of VDR agonists as therapeutic agents. The studies reported here focused on molecular characterization of the promoter region upstream of exon 1c in the human VDR gene. In transient transfection assays, luciferase reporter constructs containing -800 to +31 of the VDR gene exhibit basal promoter activity in T47D breast cancer cells which is enhanced by 1,25(OH)(2)D(3), estrogen and the phytoestrogen resveratrol. Deletion constructs and site-directed mutagenesis were used to map three distinct GC-rich Sp1 consensus sites that independently mediate the effects of estrogen, resveratrol, and 1,25(OH)(2)D(3) on VDR promoter activity. Up-regulation of the VDR promoter by 1,25(OH)(2)D(3) was mapped to an Sp1 site 261bp upstream of exon 1c, estrogen responsiveness to a proximal Sp1 site beginning at -50, and resveratrol regulation to a distal Sp1 site beginning at -381. Studies with estrogen receptor (ER) subtype specific ligands suggest that the effect of estrogen on VDR promoter is dependent on both ERalpha and ERbeta, whereas the effect of resveratrol is dependent only on ERalpha. In summary, these studies demonstrate transcriptional regulation of the exon 1c VDR promoter in breast cancer cells, and identify three distinct GC-rich, Sp1 consensus sites that differentially confer responsiveness to estrogen, resveratrol and 1,25(OH)(2)D(3). PMID- 15664453 TI - Structure and expression of the chicken proglucagon gene. AB - The rat and human proglucagon gene immediate promoter regions have differing transcriptional activities in islet cell lines. It is unclear upon which lineage the expression changes have occurred, or which base changes and transcription factors explain the changes in promoter activity. We have studied the expression of the chicken proglucagon gene to better understand changes in mammalian proglucagon gene promoter activity. The chicken proglucagon gene is composed of seven exons, and contains an additional exon that contains much of intervening peptide 1 that has not been seen in any other proglucagon gene. Despite very limited sequence identity between chicken and rat proglucagon promoters, the chicken proglucagon promoter supports reporter gene expression in rodent glucagon expressing cell lines. These results suggest that the immediate proglucagon promoter region likely has retained ability to be expressed in islet cells, but a few changes to this sequence on the human lineage have extinguished expression of the immediate promoter region. Our work also demonstrate that in contrast to the conserved human proglucagon promoter region which has lost promoter activity that significant changes in proglucagon promoter sequence can occur yet retain expression potential. PMID- 15664454 TI - Tob proteins suppress steroid hormone receptor-mediated transcriptional activation. AB - Although sex steroid hormones have significant effects on bone metabolism, the molecular mechanisms of these actions have not been fully elucidated yet. We examined the functional relationship between steroid hormone receptors and Tob, a member of an anti-proliferative protein family and a negative regulator of osteoblast proliferation and differentiation. Luciferase assay using promoters carrying hormone-responsive elements revealed that both Tob1 and Tob2 proteins but not PC3 suppressed steroid hormone receptor-dependent transcriptional activation in MC3T3-E1 osteoblastic cells. Mutated Tob proteins carrying amino acid substitutions at an LXXLL motif also showed the same degree of inhibition of the transcriptional activation as the wild type. By observation of androgen receptor (AR)-tagged with green fluorescent protein under a confocal laser scanning microscope, we found that Tob1 inhibits the nuclear foci formation of dihydrotestosterone-bound AR. These results indicate that Tob family proteins may negatively regulate sex steroid hormone action in bone formation. PMID- 15664456 TI - Proteomic patterns: their potential for disease diagnosis. AB - Alterations in proteins abundance, structure, or function, act as useful indicators of pathological abnormalities prior to development of clinical symptoms and as such are often useful diagnostic and prognostic biomarkers. The underlying mechanism of diseases such as cancer are, however, quite complicated in that often multiple dysregulated proteins are involved. It is for this reason that recent hypotheses suggest that detection of panels of biomarkers may provide higher sensitivities and specificities for disease diagnosis than is afforded with single markers. Recently, a novel approach based on the analysis of protein patterns has emerged that may provide a more effective means to diagnose diseases, such as ovarian and prostate cancer. The method is based on the use of surface-enhanced laser desorption/ionization (SELDI) time-of-flight mass spectrometry (TOF-MS) to detect differentially captured proteins from clinical samples, such as serum and plasma. This analysis results in the detection of "proteomic" patterns that have been shown in recent investigations to distinguish diseased and unaffected subjects to varying degrees. This review will discuss the basics of SELDI protein chip technology and highlight its recent applications in disease biomarker discovery with emphasis on cancer diagnosis. PMID- 15664455 TI - The pathway to femaleness: current knowledge on embryonic development of the ovary. AB - Increasing evidence indicates that organogenesis of the ovary is not a passive process arising by default in the absence of the testis pathway. A coordinated interaction is actually in force between somatic cells and female germ cells in embryonic ovaries, thus creating a unique microenvironment that facilitates the formation of follicles. Identification of the functional roles of several novel regulatory elements such as Figalpha, Foxl2, follistatin, and Wnt4 reveals the complexity of early ovarian organization. Challenges await us to establish the molecular connections of these molecules as well as to discover new candidates in the pathway of early ovarian development. PMID- 15664457 TI - Traditional phytotherapy in Central Italy (Marche, Abruzzo, and Latium). AB - In this study, the more significant results of extensive ethnopharmacobotanical research carried out by the author in the years 1977-2000 in 175 localities of three regions of Central Italy (Marche, Abruzzo, and Latium) have been reported and compared. The usages of 80 species belonging to 36 families are described, of which 71 were used in human therapy and 29 in veterinary medicine. Uses are suited with the number of localities in which they have been mentioned. Among the wild plant mainly still used, Malva sylvestris, Urtica dioica, and Sambucus nigra are particularly highly considered, while major uses of plants concern these plants in addition to Allium sativum, Rubus ulmifolius, Parietaria diffusa, Cynodon dactylon, and Ficus carica. Unusual phytotherapic uses concern Brassica oleracea, Taraxacum officinale (warts), Ruscus aculeatus, S. nigra (chilblains), Allium cepa (chilblains; to remove thorns and splinters), Juglans regia, R. ulmifolius (burns), and Euphorbia paralias (bites of weevers). Among new uses with only one quotation, we remember Cirsium arvense (intestinal disturbances), Centaurea bracteata (cough), Lupinus albus (calluses), Melittis melissophyllum (eye inflammations, antispasmodic), and Artemisia absinthium (tendon inflammations), while among plants employed in various regions with interesting less-known properties, there are C. arvense (emergency haemostatic), P. diffusa (insect bites), and Scrophularia canina (antiseptic and cicatrizing agent for wounds in bovines and sheep). PMID- 15664458 TI - Trypanocidal activity of extracts and fractions of Bertholletia excelsa. AB - Crude extracts and fractions of Bertholletia excelsa stem barks were tested for trypanocidal activity. Acetone and methanol extracts showed significant in vitro trypanocidal activity against trypomastigote form of Trypanosoma cruzi since in the concentration of 500 microg/ml, the parasites were reduced in 100% and 90.3% respectively, whereas the triterpene betulinic acid pure isolated from hexane extract presented 75.4%. PMID- 15664460 TI - Estrogenic activity of a polyphenolic extract of the leaves of Epimedium brevicornum. AB - The estrogenic activity of a polyphenolic extract of the leaves of Epimedium brevicornum and five fractions obtained by solid-phase extraction were investigated using estrogen-responsive bioassays, a yeast cell assay and the Ishikawa Var-I assay. The extract was found to exhibit significant estrogenic activity in both assays. Furthermore, bioassay-guided fractionation led to localisation of the estrogenicity in the relatively non-polar fractions of the polyphenolic extract. PMID- 15664459 TI - Evaluation of oral therapy on Mansonial Schistosomiasis using single dose of Balanites aegyptiaca fruits and praziquantel. AB - The efficacy of Balanites aegyptiaca fruit mesocarp was compared with praziquantel in mice infected with Sudanese strain of Schistosoma mansoni. Infected mice were given a single dose of 200 mg/kg body weight of B. aegyptiaca fruit mesocarp and 200 mg/kg b.w. of praziquantel after 6 weeks from the onset of the infection. A significant reduction was observed in EPG (egg count per gram of faeces), eggs burden in tissues and recovery of adult worms (P<0.05) for both the plant and the drug-treated animals. PMID- 15664461 TI - Stem and leaf morphoanatomy of Maytenus ilicifolia. AB - Maytenus ilicifolia is a woody medicinal plant, employed mainly for its antiulcerogenic properties. The stem and leaf morphoanatomy has been studied, aiming to supply knowledge for the pharmacognostic and taxonomic species identification. The vegetative material was fixed, freehand sectioned and stained according to usual microtechniques. The stem organization, in secondary growth, shows periderm beneath the remaining epidermis, conspicuous sclerenchymatic ring in the cortex and cambium forming phloem outside and xylem inside. The leaf is simple, alternate and lanceolate and has sparsely spiny teeth along the margin. Epidermal cells containing calcium oxalate crystals, thick cuticle that forms cuticular flanges, dorsiventral mesophyll and amphicrival bundle in the midrib and petiole are observed. PMID- 15664462 TI - Cytotoxic activity of extracts from Linum cell cultures. AB - Callus and suspension cultures of Linum narbonense and Linum leonii were developed to study the production of lignans and their cytotoxic activity. Justicidin B was determined to be the main lignan. The maximal yield of justicidin B up to 2.22 mg/g of the cell dry weight was detected in the callus cultures of L. leonii, followed by the callus cultures of L. narbonense (1.57 mg/g dwt). The cytotoxicity of the obtained extracts was measured using the MTT dye assay. L. narbonense and L. leonii both showed cytotoxic activity. PMID- 15664464 TI - Quantitative determination of alpha-tocopherol in Pistacia lentiscus, Pistacia lentiscus var. chia, and Pistacia terebinthus by TLC-densitometry and colorimetry. AB - A quantitative determination of alpha-tocopherol in Pistacia lentiscus, Pistacia lentiscus var. chia, and Pistacia terebinthus, leaves was established by TLC densitometry and colorimetry. The highest amount of alpha-tocopherol was found in P. lentiscus var. chia. PMID- 15664463 TI - Antifilarial activity of Azadirachta indica on cattle filarial parasite Setaria cervi. AB - Alcohol and aqueous extracts of flowers of Azadirachta indica were tested in vitro for their potential antifilarial activity against whole worm, nerve muscle (n.m.) preparation and microfilariae of Setaria cervi. The effects of alcohol and aqueous extracts were similar in nature on the spontaneous movements of whole worm and nerve muscle preparation. On the whole worm, the response was characterized by initial increase in tone, rate and amplitude of contractions followed by reversible paralysis. The initial stimulant effect is likely to be due to irritant effect on the cuticle. Nerve muscle preparation responded to both extracts by inhibition of spontaneous movements followed by reversible paralysis; initial stimulation phase was absent. The inhibition was concentration related. Alcohol and aqueous extracts had almost similar lethal effect on the microfilariae of S. cervi, the LC50 being 15 and 18 ng/ml, respectively. PMID- 15664465 TI - Antibacterial activity of seed proteins of Robinia pseudoacacia. AB - A low molecular weight cationic peptide was isolated from Robinia pseudoacacia seed and tested in vitro against seven bacteria (Corynebacterium michiganense, Staphylococcus aureus, Bacillus subtilis, Erwinia carotovora subsp. carotovora, Pseudomonas syringae pv syringae, Xanthomonas campestris pv campestris, and Escherichia coli). The peptide inhibited the growth of the tested strains. The effective concentrations required for 50% inhibition of bacterial growth ranged between 20 and 120 microg ml(-1) protein. S. aureus was found to be the most sensitive strain, however, E. coli was not affected much when compared with others. Reduction of antibacterial activity of the peptide with CaCl2 addition into the growth medium was also observed. PMID- 15664467 TI - Methyljasmonate accelerates catabolism of monoterpenoid indole alkaloids in Catharanthus roseus during leaf processing. AB - Variations in alkaloid pattern during drying of leaves (leaf processing) showed that treatment with methyljasmonate can induce formation of bisindole alkaloids as a result of catabolism of the monomeric alkaloids catharanthine and vindoline. A two-fold increase in 3',4'-anhydrovinblastine was shown in treated leaves especially from day 8 until day 21. Serpentine also increased in the same period under the treatment as a catabolic product of ajmalicine. Basic peroxidases that are responsible for the formation of anhydrovinblastine and serpentine showed high activity at days 8 and 21 in treated leaves, causing the increase in anhydrovinblastine and serpentine. PMID- 15664466 TI - Chemical constituents of Lychnophora pohlii and trypanocidal activity of crude plant extracts and of isolated compounds. AB - Crude extracts of Lychnophora pohlii were tested in vitro against trypomastigote forms of Trypanosoma cruzi, and the dichloromethane and methanol crude extracts from leaves plus inflorescences were found to have trypanocidal activity. The bioassay-guided fractionation of the extracts yielded seven active compounds: the sesquiterpene lactones lychnopholide, centratherin, goyazensolide and 15 desoxygoyazensolide in the dichloromethane extract, and caffeic acid and the flavonoids luteolin and vicenin-2 in the methanol extract. One active caffeoyl quinic acid derivative was isolated from the inactive hydroalcoholic extract of leaves plus inflorescences. Chemically, the plant has sesquiterpene lactone type furanoheliangolides, flavonoids, caffeic acid, a caffeoyl quinic acid derivative, which are characteristic of the Vernonieae. PMID- 15664468 TI - Analgesic and antipyretic activities of the aqueous extract of Urtica macrorrhiza in experimental animals. AB - Oral administration of the aqueous extract of the stem of Urtica macrorrhiza (100 400 mg/kg) dose dependently reduced the number of writhings and stretchings induced by acetic acid and decreased licking activity of the late phase in formalin test at doses of 200 and 400 mg/kg. At doses of 200 and 400 mg/kg, p.o. it suppressed yeast-induced fever in rats. PMID- 15664469 TI - Inhibition of the Carpobrotus edulis methanol extract on the growth of phagocytosed multidrug-resistant Mycobacterium tuberculosis and methicillin resistant Staphylococcus aureus. AB - The Carpobrotus edulis methanol extract, inactive against the methicillin resistant Staphylococcus aureus or the multidrug-resistant Mycobacterium tuberculosis, does inhibit the growth of these two bacteria once they are phagocytosed by monocyte derived human macrophages. PMID- 15664470 TI - Antinociceptive activity of Eupatorium buniifolium aqueous extract. AB - The antinociceptive properties of the aqueous extract of Eupatorium buniifolium aerial parts were investigated in chemical and thermal models of nociception in mice. Oral doses of 250 and 500 mg/kg produced an inhibition of 42.3% and 73.9%, respectively, of the abdominal writhes induced by acetic acid in mice. This antinociceptive effect was not reversed by pretreatment with naloxone. In the formalin test, the administration of 500 and 1000 mg/kg p.o. had no effects in the first phase (0-5 min) but produced a dose-dependent analgesic effect on the second phase (15-30 min) with inhibitions of the licking time of 38.9% and 57.3%, respectively. In the hot plate test, no effect was seen at the dose of 500 mg/kg p.o. PMID- 15664471 TI - In vitro anti-Vibrio cholerae activity of essential oil from Lepechinia caulescens. AB - Lepechinia caulescens is a plant employed by the Purepecha (a Mexican ethnic group) to treat infectious gastrointestinal ailments. The essential oil of this species was active against some strains of Vibrio cholerae with 4 microl/ml MIC and 6 microl/ml MBC. The major components of the oil found by GC-MS were borneol, camphor and trans-caryophyllene. PMID- 15664472 TI - Antifungal activity of Cynara scolymus L. extracts. AB - Chloroform, ethanol and ethyl acetate extracts of Cynara scolymus L. leaves, heads and stems were tested for their antifungal activity using the agar-well diffusion assay technique. The leaves extracts and the ethanol fractions were found to be the most effective extract against all the tested organisms. PMID- 15664473 TI - Antimicrobial activity of Chenopodium botrys essential oil. AB - Essential oil, isolated by hydrodistillation from Chenopodium botrys aerial parts, expressed significant bactericidal and fungicidal activity against selected strains of microorganisms. PMID- 15664474 TI - Flavonol glycosides with free radical-scavenging activity of Saururus chinensis. AB - An activity-guided fractionation procedure was used to identify the antioxidative components of the aerial parts of Saururus chinensis. The antioxidant activity was investigated with the 1,1-diphenyl-2-picrylhydrazyl (DPPH) radical- and superoxide anion-scavenging assays. Three active compounds (flavonol glycosides) were identified. PMID- 15664476 TI - Ethnobotany of medicinal plants from Dibru-Saikhowa Biosphere Reserve of Northeast India. AB - A survey of medicinal plants used by the ethnic communities of Dibru-Saikhowa Biosphere Reserve in Northeast India has been reported. Fifty-five plant species belonging to 34 families and 52 different genera were included. For each plant species, botanical name, vernacular name, part(s) used, popular medicinal use, forms of preparation and applications of the herbal remedies are provided. PMID- 15664475 TI - Antimicrobial activity of Datura innoxia and Datura stramonium. AB - The antibacterial activity of the methanol extracts of the aerial parts of the Datura innoxia and Datura stramonium was investigated. The extracts showed activity against Gram (+) bacteria in a dose dependent manner. Little or no antibacterial activity was found against Escherichia coli and Pseudomonas aeruginosa. PMID- 15664477 TI - Flavonoids from Blumea balsamifera. AB - 3,4',5-Trihydroxy-3',7-dimethoxyflavanone was isolated from the ligroin extract of the leaves of Blumea balsamifera, while the acetone extract yielded 3',4',5 trihydroxy-7-methoxyflavanone and a new biflavonoid identifed as 3-O-7'' biluteolin (1). The isolation of 1 is significant since a biflavonoid with a C-O C linkage of the type [I-3-O-II-7] has not been previously reported from a plant. PMID- 15664478 TI - Oleanane triterpenes from Aegiceras corniculatum. AB - A new oleanane triterpene, 16alpha-hydroxy-l3,28-epoxyoleanan-3-one 1, was isolated from the stem bark of Aegiceras corniculatum, together with protoprimulagenin, aegicerin, embelinone, syringic acid, gallic acid, isorhamnetin and isorhamnetin 3-O-alpha-L-rhamnofuranosyl-(1-->6)-beta-D glucopyranoside. Their structures were determined by spectroscopic methods. PMID- 15664479 TI - Antimicrobial susceptibility of uncommonly isolated non-enteric Gram-negative bacilli. AB - The frequency of occurrence and antimicrobial susceptibility patterns of 3059 non enteric Gram-negative bacilli (NGB), other than Pseudomonas aeruginosa and Acinetobacter spp., consecutively collected as part of the SENTRY Antimicrobial Surveillance Program (1997-2003) were reviewed. During this period, a total of 221,084 bacterial isolates were collected from several clinical specimens worldwide, including 25,305 (11.5%) NGB. Acinetobacter spp. and P. aeruginosa accounted for 82.7% of the NGB isolates and have been excluded from this analysis. The antimicrobial susceptibility results of 3509 strains from 13 species/genera have been analysed in this review. The isolates were tested by reference broth microdilution methods in three central laboratories using common reagents and procedures. More than 30 antimicrobial agents were tested and the results for the 18 most active compounds are reported here. Stenotrophomonas maltophilia (2076 strains; 59.2%) was the most frequently isolated pathogen in this group, followed by Aeromonas spp. (385 strain; 11.0%), Burkholderia cepacia (269 strains; 7.7%), Pseudomonas fluorescens/putida (253 strains; 7.2%) and Alcaligenes spp. (236 strains; 6.7%). All other species/genera accounted for less than 3% of the isolates analysed. The antimicrobial agents with the most consistent activity against the NGB evaluated in the present study were the newer fluoroquinolones gatifloxacin and levofloxacin with 84.1 and 84.9% susceptibility overall. Trimethoprim/sulphamethoxazole was active against 85.3% of the isolates tested, but showed reduced activity against P. fluorescens/putida (22.1% susceptibility). Antimicrobial susceptibility varied significantly between species/genera and the geographical regions evaluated. Thus, proper identification and quantitative susceptibility testing will be required for the treatment of NGB infections. Extensive worldwide surveillance programmes remain extremely important to guide empirical antimicrobial therapy for rarely isolated pathogens and also for pathogens that are not routinely tested due to the lack of standardised susceptibility testing methods. PMID- 15664481 TI - Pharmacokinetic-pharmacodynamic modelling of antibacterial activity of cefpodoxime and cefixime in in vitro kinetic models. AB - The bacterial time-kill curves of cefpodoxime and cefixime against four bacterial strains (Haemophilus influenzae, Moraxella catarrhalis, Streptococcus pneumoniae/penicillin sensitive and S. pneumoniae/penicillin intermediate) were compared in in vitro infection models in which various human pharmacokinetic profiles of unbound antibiotic were simulated. This approach offers more detailed information than the minimum inhibitory concentration (MIC) does about the time course of antibacterial efficacy of an antibiotic. A pharmacokinetic pharmacodynamic (PK-PD) model based on unbound antibiotic concentrations at the site of infection, and a sigmoid Emax-relationship with EC50 as the antibiotic concentration necessary to produce 50% of the maximum effect, effectively described the antimicrobial efficacy of both cefpodoxime and cefixime. The EC50 values of cefpodoxime and cefixime were consistent with their respective MIC values. Both antibiotics had similar high potency against H. influenzae (EC50: 0.04 mg/L) and M. catarrhalis (EC50: 0.12 mg/L), while the potency of cefpodoxime against S. pneumoniae strains was about 10-fold higher than that of cefixime (EC50s/sensitive strain: 0.02 mg/L versus 0.27 mg/L; EC50s/intermediate strain: 0.09 mg/L versus 0.69 mg/L). Applications of this model and unbound tissue PK profiles obtained from a previous clinical study performed in our group, showed that cefpodoxime has higher bacteriological potency than cefixime against S. pneumoniae. Simulations based on this model allow the comparison of antibacterial efficacy of different antibiotics and dosing regimens. PMID- 15664480 TI - Efficacy of a new pharmacokinetically enhanced formulation of amoxicillin/clavulanate (2000/125 mg) in adults with community-acquired pneumonia caused by Streptococcus pneumoniae, including penicillin-resistant strains. AB - Community-acquired pneumonia (CAP) is a common respiratory illness, frequently caused by Streptococcus pneumoniae. The prevalence of S. pneumoniae resistance to common antimicrobials has increased over recent years. A new pharmacokinetically enhanced formulation of amoxicillin/clavulanate (2000/125 mg) has been developed, designed to combat infections caused by S. pneumoniae, including penicillin resistant (PRSP, penicillin minimum inhibitory concentrations (MICs) >or=2mg/l) isolates, and those with elevated amoxicillin/clavulanic acid MICs, while maintaining coverage of beta-lactamase-producing pathogens. A pooled efficacy analysis of four randomized (1:1) and one non-comparative clinical trials of amoxicillin/clavulanate, 2000/125 mg, given twice daily, was conducted in adult patients with CAP. Comparator agents were conventional amoxicillin/clavulanate formulations. At follow-up (days 16-39), efficacy (eradication of the initial pathogen or clinical cure in patients for whom no repeat culture was performed) in patients with S. pneumoniae infection was 92.3% (274/297) for amoxicillin/clavulanate, 2000/125 mg and 85.2% (46/54) for comparators (P=0.11). Twenty-four of 25 PRSP-infected patients receiving amoxicillin/clavulanate, 2000/125 mg were treated successfully. Both amoxicillin/clavulanate, 2000/125 mg and comparators were well tolerated, with few patients withdrawing from the studies. PMID- 15664482 TI - Antiproliferative synergism of azasterols and antifolates against Toxoplasma gondii. AB - The antiproliferative effects of two azasterols, 22,26-azasterol (20-piperidin-2 yl-5alpha-pregnan-3beta-20(R,S)-diol, AZA) and 24,25(R,S)-epiminolanosterol (EIL), in combination with sulphadiazine (SDZ) and pyrimethamine (PYR) were studied against Toxoplasma gondii tachyzoites growing in cultured mammalian cells. We had previously shown that AZA and EIL, two known inhibitors of Delta24(25)sterol methyl transferase in fungi and protozoa, have a potent and selective anti-T. gondii activity, although no 24-alkyl sterols have been detected in this parasite. We now report that when these sterol analogues were used in combination with the conventional SDZ/PYR treatment, potent synergistic effects were observed, ranging from 10- to 100-fold reductions of the IC50 values in the presence of sub-optimal doses of azasterols. When exposed to these drug combinations, intracellular T. gondii parasites displayed diverse subcellular alterations, including mitochondrial swelling, the arrest of the endodiogeny process with fragmented nuclei and subsequent cell lysis. These results suggest a potential new approach for the treatment of toxoplasmosis, which could significantly lower the required levels of antifolates and thus their adverse side effects. PMID- 15664483 TI - In vitro activity of C20-diterpenoid alkaloid derivatives in promastigotes and intracellular amastigotes of Leishmania infantum. AB - The in vitro anti-proliferative effects are described of several atisine-type diterpenoid alkaloids against the protozoan parasite Leishmania infantum, which causes human visceral leishmaniasis and canine leishmaniasis in the Mediterranean basin, as well as human cutaneous leishmaniasis throughout the Mediterranean region. From a total of 43 compounds tested, including C19- and C20-diterpene alkaloids from several chemical classes, only 15,22-O-diacetyl-19-oxo dihydroatisine, azitine and isoazitine were highly active against cultures of the parasite (promastigote form) with IC50 values within the range of the reference drug pentamidine-isothionate (7.39-12.80 mg/L for the test compounds, 11.32 mg/L for the positive control). These compounds were not toxic to the host cell. When treated with a dosage of 5 microg/mL of the active compounds (half of their IC50), the promastigote forms lost 80% of their infection capacity and the multiplication of extracellular forms of L. infantum was severely affected. The study showed that atisine-type C20-diterpenoid alkaloids exhibited promising anti leishmanial properties with strong molecular selectivity. These might have implications for other intracellular pathogens- or phylogenetically related parasites, such as Trypanosoma spp. PMID- 15664484 TI - Molecular mechanisms of high level tetracycline-resistance in group A streptococcal isolates, T serotypes 4 and 11. AB - The molecular mechanism of high level tetracycline resistance in T serotypes 4 and 11 group A streptococcal (GAS) isolates was examined in 61 tetracycline resistant isolates in Japan. PCR and sequencing analyses revealed that the T serotype/emm genotype, T4/4 isolates carried tet(O) genes, which were genetically homogenous. The T11/11 and T11/89 isolates carried different subtypes of tet(M) genes, which were present on transposons Tn916 and Tn1545, respectively. In addition, these T11 isolates may have obtained the tet(M) gene after the 1990s, because resistance to tetracycline in T11 isolates was rarely found before then. These results strongly suggested that the T4 and T11 GAS isolates acquired tetracycline-resistance via different molecular mechanisms. PMID- 15664485 TI - Antimicrobial resistance in Streptococcus pneumoniae, Haemophilus influenzae, Moraxella catarrhalis and group A beta-haemolytic streptococci in 2002-2003. Results of the multinational GRASP Surveillance Program. AB - A multinational surveillance study, GRASP, was conducted between November 2002 and April 2003 with the aim of assessing rates of antimicrobial resistance among 2656 isolates of Streptococcus pneumoniae, 2486 isolates of group A beta haemolytic streptococci, 1358 isolates of Haemophilus influenzae and 1047 of Moraxella catarrhalis from 20 countries in Europe, eastern Asia and southern Africa. Conspicuous differences between various countries were noted in the S. pneumoniae resistance rates observed for penicillin (0-79.2%) and erythromycin (4 66%), along with other antimicrobials. The percentage of MDR strains was above 25% in 8 of the 20 countries studied. Group A streptococcal macrolide resistance rates ranged from 0% to 35% by country, while rates of beta-lactamase production ranged from 0% to 39% for H. influenzae and 80-100% for M. catarrhalis. Antibiotic resistance in S. pneumoniae remains a significant problem world wide. PMID- 15664486 TI - Evolution of CTX-M-type beta-lactamases in isolates of Escherichia coli infecting hospital and community patients. AB - Escherichia coli isolates collected at our Institution from 1999 to 2003 (n=20,258) were studied to evaluate the production of CTX-M-type extended spectrum beta-lactamases (ESBL). Isolates suspected of producing CTX-M enzymes were analyzed by the double-disk synergy test, hybridization with specific probes, PCR and direct DNA sequencing. Overall, 53 ESBL-positive isolates were found to carry CTX-M-type genes (blaCTX-M-1, n=51; blaCTX-M-15, n=2). The isolation of CTX-M-positive strains increased from 1 per year (1999) to 26 per year (2003). The first isolate carrying the blaCTX-M-15 gene appeared in 2003 and was obtained from a patient previously treated with ceftazidime. CTX-M-positive isolates were characterized by multi-drug resistance and were obtained both from inpatients (n=29) and outpatients (n=24). Most patients were over 60-year-old (n=45), had underlying chronic diseases (n=32), and had been hospitalized more than once (n=33). Strains were frequently isolated from the urinary tract, often after recurrent infections. Our study demonstrates that CTX-M-producing isolates are increasing among E. coli strains. Adequate laboratory detection may help in choosing appropriate treatment and in limiting the spread of this resistance trait. PMID- 15664487 TI - In vivo activity of gemifloxacin, moxifloxacin and levofloxacin against pneumococci with gyrA and parC point mutations in a sepsis mouse model measured with the all or nothing mortality end-point. AB - A dose-decreasing immunocompetent sepsis mouse model was used to evaluate the in vivo effect of levofloxacin, moxifloxacin and gemifloxacin, using a ciprofloxacin/levofloxacin susceptible serotype 6B strain (ciprofloxacin MIC: 1 mg/l) and two resistant serotype 14 and 19F strains with gyrA and parC point mutations (ciprofloxacin MICs of 32 and 64 mg/l, respectively). Significant higher in vivo activity was found for moxifloxacin and gemifloxacin than for levofloxacin against strains 1 and 2, and for gemifloxacin versus moxifloxacin or levofloxacin against strain 3. Gemifloxacin treatment resulted in 100% survival against strains 1 and 2(AUC0-24 h/MIC of 30 and 62) but against strain 3, survival was 60-80% (AUC0-24 h/MIC of 93). Similar AUC0-24 h/MIC values produced different therapeutic results suggesting that in vitro parameters other than the MIC could influence efficacy predictions based on in vitro susceptibility tests (MICs) or pharmacodynamic parameters (AUC0-24 h/MIC). PMID- 15664489 TI - Influence of various criteria for elimination of duplicates when calculating the prevalence and antibiotic susceptibility of microorganisms associated with urinary infections. AB - The effect of applying various criteria for elimination of repeated isolates in the same patient on the prevalence and antibiotic susceptibility of microorganisms frequently associated with urinary infections were analyzed. When time criteria were applied, there was a decrease in the number of isolates of 13.4-27.5%, whereas when variations in antibiotic susceptibility were considered, between 4.3 and 20.3% of the isolates were eliminated. The absolute number of isolates varied considerably depending on which of these two criteria were used, but neither criterion was seen to modify significantly the antibiotic susceptibility of these microorganisms and this confirms the recommendations of the NCCLS for this group of pathogens. PMID- 15664490 TI - Heterogeneous vancomycin-resistant Staphylococcus aureus (hetero-VISA) in Singapore. PMID- 15664488 TI - Clarithromycin co-administered with amikacin attenuates systemic inflammation in experimental sepsis with Escherichia coli. AB - To assess the efficacy of clarithromycin as an immunomodulator in experimental sepsis with Escherichia coli, acute pyelonephritis was induced after ligation of the right ureter and injection of the test isolate into the renal pelvis in 40 rabbits. Four groups of treatment were applied with administration of therapy on advent of sepsis-associated pulmonary oedema, as follows: A: controls; B: clarithromycin; C: amikacin, D: both agents. Survival was recorded along with estimation of serum levels of endotoxins (LPS), of tumour necrosis factor-alpha (TNFalpha), malondialdehyde (MDA) and of bacterial counts. Mean survival of groups A, B, C and D was 2.51, 7.60, 10.25 and 11.40 days, respectively. Serum levels of TNFalpha and of MDA of group A increased over-time. Pulmonary oedema at 6 h after bacterial challenge was accompanied by increase of TNFalpha and MDA; administration of clarithromycin decreased their values. It is concluded that intravenous clarithromycin might constitute a promising immunomodulatory agent for the management of sepsis since its efficacy was proved after administration on presentation of sepsis-associated pulmonary oedema. The presented findings emphasise the need for further clinical research of the use of clarithromycin for the therapy of Gram-negative sepsis. PMID- 15664491 TI - In vitro activity of new quinoxalin 1,4-dioxide derivatives against strains of Mycobacterium tuberculosis and other mycobacteria. PMID- 15664493 TI - Prevalence of antibiotic use for acute respiratory infection in a primary care clinic in post-war Kabul province, Afghanistan. PMID- 15664492 TI - In vitro activity of meropenem in combination with ciprofloxacin against clinical isolates of Pseudomonas aeruginosa. PMID- 15664494 TI - Use of Etest to assess synergy of antibiotic combinations against clinical isolates of Pseudomonas spp. PMID- 15664495 TI - Ultrafast relaxation dynamics of a biologically relevant probe dansyl at the micellar surface. AB - We report picosecond-resolved measurement of the fluorescence of a well-known biologically relevant probe, dansyl chromophore at the surface of a cationic micelle (cetyltrimethylammonium bromide, CTAB). The dansyl chromophore has environmentally sensitive fluorescence quantum yields and emission maxima, along with large Stokes shift. In order to study the solvation dynamics of the micellar environment, we measured the fluorescence of dansyl chromophore attached to the micellar surface. The fluorescence transients were observed to decay (with time constant approximately 350 ps) in the blue end and rise with similar timescale in the red end, indicative of solvation dynamics of the environment. The solvation correlation function is measured to decay with time constant 338 ps, which is much slower than that of ordinary bulk water. Time-resolved anisotropy of the dansyl chromophore shows a bi-exponential decay with time constants 413 ps (23%) and 1.3 ns (77%), which is considerably slower than that in free solvents revealing the rigidity of the dansyl-micelle complex. Time-resolved area normalized emission spectroscopic (TRANES) analysis of the time dependent emission spectra of the dansyl chromophore in the micellar environment shows an isoemissive point at 21066 cm-1. This indicates the fluorescence of the chromophore contains emission from two kinds of excited states namely locally excited state (prior to charge transfer) and charge transfer state. The nature of the solvation dynamics in the micellar environments is therefore explored from the time-resolved anisotropy measurement coupled with the TRANES analysis of the fluorescence transients. The time scale of the solvation is important for the mechanism of molecular recognition. PMID- 15664496 TI - Flow-cytometric monitoring of mitochondrial depolarisation: from fluorescence intensities to millivolts. AB - Redistribution potentiometric dyes represent a powerful tool for monitoring membrane potential of mitochondria, especially when these dyes are used with flow cytometry. In particular, tetramethylrhodamine methyl ester proved to be suitable for the screening of mitochondrial membrane potential in cultured human skin fibroblasts from patients suffering from different defects of oxidative phosphorylation. We have developed a method that makes it possible to measure the changes in mitochondrial membrane potential, or to assess the differences between respective mitochondrial membrane potentials in investigated cells and controls in the absolute scale of millivolts. Our approach employs the fact that a logarithmic transformation of Nernst equation-controlled intensity of fluorescence from potentiometric dyes accumulated in mitochondria leads to a linear scale for mitochondrial membrane potentials. PMID- 15664497 TI - Temperature-induced isomerization of violaxanthin in organic solvents and in light-harvesting complex II. AB - Three main xanthophyll pigments are bound to the major photosynthetic pigment protein complex of Photosystem II (LHCII): lutein, neoxanthin and violaxanthin. Chromatographic analysis of the xanthophyll fraction of LHCII reveals that lutein appears mainly in the all-trans conformation, neoxanthin in the 9'-cis conformation and major fraction of violaxanthin in the all-trans conformation. Nevertheless, a small fraction of violaxanthin appears always in a cis conformation: 9-cis and 13-cis (approximately 4% and 2% in the darkness, respectively). Illumination of the isolated complex (5 min, 445 nm, 250 micromolm 2s-1) results in the substantial increase in the concentration of the cis steric conformers of violaxanthin: up to 6% of 9-cis and 4% of 13-cis. Similar effect can be obtained by dark incubation of the same preparation for 30 min at 60 degrees C. Heating-induced isomerization of the all-trans violaxanthin can also be obtained in the organic solvent system but the formation of the 9-cis stereoisomer has not been observed under such conditions. The fact that the appearance of the 9-cis form of violaxanthin is specific for the protein environment suggests that violaxanthin may replace neoxanthin in LHCII in the N1 xanthophyll binding pocket and that the protein stabilizes this particular conformation. The analysis of the electronic absorption spectra of LHCII and the FTIR spectra of the protein in the Amid I band spectral region indicates that violaxanthin isomerization is associated with the disaggregation of the complex. It is postulated that this reorganization of LHCII provides conditions for desorption of violaxanthin from the pigment protein complexes, its diffusion within the thylakoid membrane and therefore, availability to the enzymatic deepoxidation within the xanthophyll cycle. It is also possible that violaxanthin isomerization plays the role of a security valve, by consuming an energy of excessive excitations in the antenna pigment network (in particular, exchanged at the triplet state levels). PMID- 15664498 TI - Role of LHCII organization in the interaction of substituted 1,4-anthraquinones with thylakoid membranes. AB - The chlorophyll fluorescence, photochemical activity and surface electric properties of thylakoid membranes with different stoichiometry of pigment-protein complexes and organization of the light-harvesting chlorophyll a/b protein complex of photosystem II (LHCII) were studied in the presence of substituted 1,4 anthraquinones. Data show strong dependence of the quenching of the chlorophyll fluorescence on the structural organization of LHCII. The increase of the LHCII oligomerization, which is associated with significant reduction of the transmembrane electric charge asymmetry and electric polarizability of the membrane, correlates with enhanced quenching effect of substituted 1,4 athraquinones. Crucial for the large quinone-induced changes in the membrane electric dipole moments is the structure of the quinone molecule. The strongest reduction in the values of the dipole moments is observed after interaction of thylakoids with 3-chloro-9-hydroxy-1,4-anthraquinone (TF33) which has the highest quenching efficiency. The quinone induced changes in the photochemical activity of photosystem II (PSII) correlate with the total amount of the supramolecular LHCII-PSII complex and depend on the number of substituents in the 1,4 anthraquinone molecule. PMID- 15664499 TI - Photochemical activities of plant photosystem I particles reconstituted into phosphatidylglycerol liposomes. AB - Phosphatidylglycerol (PG) is the only anionic phospholipid in photosynthetic membrane and the important component of photosystem I (PSI). In this study, the interaction of PG with PSI particle from spinach was investigated by using reconstitution method. The results from the properties of electron transport, fluorescence emission, turbidity, and protein secondary structures in PSI complex incorporated into PG liposomes revealed the existence of PSI-PG interactions. A stimulation and an inhibition of oxygen uptake in PSI particle at a low and higher PG/chlorophyll mass ratio, respectively, were observed. Moreover, an additional enhancement and depression of electron flow in the PSI-PG complexes were occurred in the reaction medium containing CaCl2 at concentrations below and above 5 mM, the aggregation threshold of the reconstituted membranes, respectively. The results demonstrated that the maintenance of the structural optimization was needed for a stimulation of electron transport at a low PG/PSI mass ratio, while a decay of this PSI activity at high PG/PSI ratio was the result of inhibition of the energy transfer from LHCI to PSI reaction center induced by the dissociation of LHCI-680. PMID- 15664501 TI - 1,25-Dihydroxyvitamin D3 and analogues protect primary human keratinocytes against UVB-induced DNA damage. AB - Exposure to UVB irradiation is a major risk factor for the development of skin cancer. Therefore, it is important to identify agents that can offer protection against UVB-caused damage. Photocarcinogenesis is caused largely by mutations at sites of incorrectly repaired DNA photoproducts, of which the most common are the cyclobutane pyrimidine dimers (CPDs). In this study, we demonstrated that 1,25 dihydroxyvitamin D3 [1,25(OH)2D3] protects primary human keratinocytes against the induction of CPDs by UVB. This protection required pharmacologic doses 1,25(OH)2D3 and an incubation period of at least 8 h before irradiation. Furthermore, we provided arguments indicating that the anti-proliferative capacity of 1,25(OH)2D3 underlies its protective effect against UVB-induced DNA damage. Finally, we showed that 19-nor-14-epi-23-yne-1,25(OH)2D3 (TX 522) and 19 nor-14,20-bisepi-23-yne-1,25(OH)2D3 (TX 527), two low-calcemic analogues of 1,25(OH)2D3, were even 100 times more potent than the parent molecule in inhibiting UVB-caused DNA damage. These molecules are therefore promising candidates for the chemoprevention of UVB-induced skin cancer. PMID- 15664500 TI - Effects of photodynamic therapy using a fractionated dosing of mono-L-aspartyl chlorin e6 in a murine tumor. AB - One of the 'second generation' photosensitizing agents is N-acetyl chlorin e6 (NPe6). This product has a strong absorbance band at 665 nm, permitting treatment at a greater depth of tumor than earlier agents based on porphyrin structures. We examined the effects of fractionated drug administration on photodynamic efficacy. Prior studies had shown that it is the level of NPe6 in the circulation that predicts for photodynamic efficacy, indicating vascular shut-down to be the predominant mode of tumor control. Although pharmacokinetic studies revealed that >99% of NPe6 was lost from the circulation, it appears that a fractionated dosage protocol can promote photodamage to neoplastic tissue in vivo. This study also indicated the potential utility of an implantable micro array for tumor irradiation. PMID- 15664502 TI - Year-long biophoton measurements: normalized frequency count analysis and seasonal dependency. AB - Biophoton emissions from three healthy human subjects were measured for 52 weeks. The active nature of dorsal hands and personal discernable patterns in palmar hands were investigated through frequency count analysis of biophoton emission rates. Also, the seasonal dependency of biophoton emission rates from human hands was studied and we found that emission rates were lowest in autumn. There was a reversion of relative emission rates from the palms and the dorsa depending upon the season. The emission rates from the palms remain rather stable throughout the year, but those from the dorsa vary widely depending upon the season. These features of biophoton emission rates were considered in light of the diagnostic view of traditional Chinese medicine. PMID- 15664503 TI - Phototoxicity, distribution and kinetics of association of UVA-activated chlorpromazine, 8-methoxypsoralen, and 4,6,4'-trimethylangelicin in Jurkat cells. AB - Extracorporeal phototherapy (ECP) is a therapeutic approach based on photobiological effects of 8-methoxypsoralen (8-MOP) on white blood cells isolated from the blood, exposed to UVA and then reinfused into the patient. 8 MOP is presently the only drug approved for clinical application of ECP; therefore, identification of other photosensitizers with better photochemical and pharmacokinetic properties might enhance the efficacy of this treatment modality. Among such alternative drugs are 4,6,4'-trimethylangelicin (TMA) and chlorpromazine (CPZ), which have previously been studied in an animal model for ECP. In this current study, cellular bioavailability of 8-MOP, TMA and CPZ was investigated in vitro, using low doses of UVA relevant for the clinical setting of ECP. Our fluorescence microscopy study revealed that 8-MOP and CPZ penetrated readily into the cells, where they accumulated with similar kinetics. No distinct fluorescence was observed in cells incubated with TMA. We found that the phototoxic efficiency of 8-MOP was an order of magnitude greater than that of CPZ, i.e., to obtain a similar reduction in survival of cells subjected to photosensitization by the drugs, the concentration of CPZ needed to be 10 times higher than that of 8-MOP. The photoactivated TMA exhibited the highest pro apoptotic efficiency. A clear indication of photoinduced formation of reactive oxygen species and peroxidation of lipids was observed only in CPZ-sensitized cells, suggesting different mechanisms for phototoxicity mediated by CPZ and by the two furocoumarins. PMID- 15664505 TI - Effect of low-level laser therapy on the healing of second-degree burns in rats: a histological and microbiological study. AB - This paper presents the results of a study on the effects of two different doses of low-level laser therapy on healing of deep second-degree burns. Sixty rats were randomly allocated to one of four groups. A deep second-degree burn was inflicted in each rat. In the control group burns remained untreated; in two laser treated groups the burns were irradiated daily with low-level helium-neon laser with energy densities of 1.2 and 2.4 J/cm2, respectively. In the fourth group the burns were treated topically with 0.2% nitrofurazone cream every day. The response to treatments was assessed histologically at 7, 16 and 30 days after burning, and microbiologically at Day 15. The number of macrophages at day 16, and the depth of new epidermis at day 30, was significantly less in the laser treated groups in comparison with control and nitrofurazone treated groups (P=0.000). Staphylococcus epidermidis was found in the 70% of rat wounds in the laser treated groups in comparison with 100% of rats in the control group. S. aureus was found in the 40% rat wounds in the nitrofurazone treated group, but there was not found in the wounds of laser treated, and control groups. It is concluded that low-level laser therapy of deep second-degree burn caused significant decrease in the number of macrophage and depth of new epidermis. In addition, it decreased incidence of S. epidermidis and S. aureus. PMID- 15664504 TI - Kinetic nature of the thermal destabilization of LHCII macroaggregates. AB - The main light-harvesting chl a/b pigment-protein complex of photosystem II (LHCII) in isolated state forms macroaggregates with different ultrastructure and lipid content [I. Simidjiev, V. Barzda, L. Mustardy, G. Garab, Anal. Biochem. 250 (1997) 169-175]. The thermodynamic stability of highly delipidated tightly bound LHCII macroaggregates is studied by differential scanning calorimetry and fluorescence spectroscopy. The calorimetric profile of LHCII is asymmetric, the denaturation transition is taking place at around 72 degrees C. A shoulder, which overlaps with the main denaturation transition, appears around 58 degrees C. The denaturation temperature strongly depends on the scanning rate indicating the kinetic nature of the thermal destabilization of LHCII macroaggregates. The fluorescence data prove that the thermal denaturation of LHCII is an irreversible and kinetically controlled process. PMID- 15664506 TI - Nurturing biotechnology's future. PMID- 15664507 TI - Targeting cancer cells with dendrimers. PMID- 15664508 TI - Chitinases and peptide mimotopes. PMID- 15664509 TI - The end of single-molecule envy. PMID- 15664510 TI - Riboswitch structures: purine ligands replace tertiary contacts. PMID- 15664511 TI - Crosslinking photosensitized by a ruthenium chelate as a tool for labeling and topographical studies of G-protein-coupled receptors. AB - The purpose was to apply oxidative crosslinking reactions to the study of recognition and signaling mechanisms associated to G-protein-coupled receptors. Using a ruthenium chelate, Ru(bipy)(3)(2+), as photosensitizer and visible light irradiation, in the presence of ammonium persulfate, we performed fast and efficient covalent labeling of the B(2) bradykinin receptor by agonist or antagonist ligands possessing a radio-iodinated phenol moiety. The chemical and topographical specificities of these crosslinking experiments were investigated. The strategy could also be applied to the covalent labeling of the B(1) bradykinin receptor, the AT(1) angiotensin II receptor, the V(1a) vasopressin receptor and the oxytocin receptor. Interestingly, we demonstrated the possibility to covalently label the AT(1) and B(2) receptors with functionalized ligands. The potential applications of metal-chelate chemistry to receptor structural and signaling studies through intramolecular or intermolecular crosslinking are presented. PMID- 15664512 TI - Direct in vitro selection of a 2'-O-methyl aptamer to VEGF. AB - Aptamers (protein binding oligonucleotides) have potential as a new class of targeted therapeutics. For applications requiring chronic systemic administration, aptamers must achieve high-affinity target binding while simultaneously retaining high in vivo stability, tolerability, and ease of chemical synthesis. To this end, we describe a method for generating aptamers composed entirely of 2'-O-methyl nucleotides (mRmY). We present conditions under which 2'-O-methyl transcripts can be generated directly and use these conditions to select a fully 2'-O-methyl aptamer from a library of 3 x 10(15) unique 2'-O methyl transcripts. This aptamer, ARC245, is 23 nucleotides in length, binds to vascular endothelial growth factor (VEGF) with a Kd of 2 nM, and inhibits VEGF activity in cellular assays. Notably, ARC245 is so stable that degradation cannot be detected after 96 hr in plasma at 37 degrees C or after autoclaving at 125 degrees C. We believe ARC245 has considerable potential as an antiangiogenesis therapeutic. PMID- 15664513 TI - Synthesis and functional evaluation of DNA-assembled polyamidoamine dendrimer clusters for cancer cell-specific targeting. AB - We sought to produce dendrimers conjugated to different biofunctional moieties (fluorescein [FITC] and folic acid [FA]), and then link them together using complementary DNA oligonucleotides to produce clustered molecules that target cancer cells that overexpress the high-affinity folate receptor. Amine terminated, generation 5 polyamidoamine (G5 PAMAM) dendrimers are first partially acetylated and then conjugated with FITC or FA, followed by the covalent attachment of complementary, 5'-phosphate-modified 34-base-long oligonucleotides. Hybridization of these oligonucleotide conjugates led to the self-assembly of the FITC- and FA-conjugated dendrimers. In vitro studies of the DNA-linked dendrimer clusters indicated specific binding to KB cells expressing the folate receptor. Confocal microscopy also showed the internalization of the dendrimer cluster. These results demonstrate the ability to design and produce supramolecular arrays of dendrimers using oligonucleotide bridges. This will also allow for further development of DNA-linked dendrimer clusters as imaging agents and therapeutics. PMID- 15664514 TI - Focusing mutations into the P. fluorescens esterase binding site increases enantioselectivity more effectively than distant mutations. AB - Rational design of enzymes with improved properties, such as enantioselectivity, usually focuses mutations within the substrate binding site. On the other hand, directed evolution of enzymes usually targets the entire protein and discovers beneficial mutations far from the substrate binding site. In this paper, we propose an explanation for this discrepancy and show that a combined approach- random mutagenesis within the substrate binding site--is better. To increase the enantioselectivity (E) of a Pseudomonas fluorescens esterase (PFE) toward methyl 3-bromo-2-methylpropionate, we focused mutagenesis into the substrate binding site at Trp28, Val121, Phe198, and Val225. Five of the catalytically active mutants (13%) showed better enantioselectivity than wild-type PFE. The increases in enantioselectivity were higher (up to 5-fold, reaching E = 61) than with mutants identified by random mutagenesis of the entire enzyme. PMID- 15664515 TI - A genome-wide overexpression screen in yeast for small-molecule target identification. AB - We describe a multicopy gene suppression screen of drug sensitivity in Saccharomyces cerevisiae that facilitates the identification of cellular targets of small molecules. An array of yeast transformants harboring a multicopy yeast genomic library was screened for resistance to growth inhibitors. Comparison of array growth patterns for several such inhibitors allowed the differentiation of general and molecule-specific genetic suppressors. Specific resistance to phenylaminopyrimidine (1), an inhibitor identified from a kinase-directed library, was associated with the overexpression of Pkc1 and a subset of downstream kinases. Components of two other pathways (pheromone response/filamentous growth and Pho85 kinase) that genetically interact with the PKC1 MAPK signaling cascade were also identified. Consistent with the suppression screen, inhibitor 1 bound to Pkc1 in yeast cell lysate and inhibited its activity in vitro. These results demonstrate the utility of this approach for the rapid deconvolution of small-molecule targets. PMID- 15664516 TI - Specificity and affinity of natural product cyclopentapeptide inhibitors against A. fumigatus, human, and bacterial chitinases. AB - Family 18 chitinases play key roles in organisms ranging from bacteria to man. There is a need for specific, potent inhibitors to probe the function of these chitinases in different organisms. Such molecules could also provide leads for the development of chemotherapeuticals with fungicidal, insecticidal, or anti inflammatory potential. Recently, two natural product peptides, argifin and argadin, have been characterized, which structurally mimic chitinase chitooligosaccharide interactions and inhibit a bacterial chitinase in the nM-mM range. Here, we show that these inhibitors also act on human and Aspergillus fumigatus chitinases. The structures of these enzymes in complex with argifin and argadin, together with mutagenesis, fluorescence, and enzymology, reveal that subtle changes in the binding site dramatically affect affinity and selectivity. The data show that it may be possible to develop specific chitinase inhibitors based on the argifin/argadin scaffolds. PMID- 15664517 TI - Simple, helical peptoid analogs of lung surfactant protein B. AB - The helical, amphipathic surfactant protein, SP-B, is a critical element of pulmonary surfactant and hence is an important therapeutic molecule. However, it is difficult to isolate from natural sources in high purity. We have created and studied three different, nonnatural analogs of a bioactive SP-B fragment (SP-B(1 25)), using oligo-N-substituted glycines (peptoids) with simple, repetitive sequences designed to favor the formation of amphiphilic helices. For comparison, a peptide with a similar repetitive sequence previously shown to be a good SP mimic was also studied, along with SP-B(1-25) itself. Surface pressure-area isotherms, surfactant film phase morphology, and dynamic adsorption behavior all indicate that the peptoids are promising mimics of SP-B(1-25). The extent of biomimicry appears to correlate with peptoid helicity and lipophilicity. These biostable oligomers could serve in a synthetic surfactant replacement to treat respiratory distress syndrome. PMID- 15664518 TI - Enhanced ligand affinity for receptors in which components of the binding site are independently mobile. AB - Using calmodulin antagonism as a model, it is demonstrated that, under circumstances in which binding sites are motionally independent, it is possible to create bifunctional ligands that bind with significant affinity enhancement over their monofunctional counterparts. Suitable head groups were identified by using a semiquantitative screen of monofunctional tryptophan analogs. Two bifunctional ligands, which contained two copies of the highest-affinity head group tethered by rigid linkers, were synthesized. The bifunctional ligands bound to calmodulin with a stoichiometry of 1:1 and with an affinity enhancement over their monofunctional counterparts; the latter bound with a stoichiometry of 2:1 ligand:protein. A lower limit to the effective concentrations of the domains of calmodulin relative to each other (0.2-2 mM) was determined. A comparable effective concentration was achieved for bifunctional ligands based on higher affinity naphthalene sulphonamide derivatives. PMID- 15664519 TI - Wortmannin, a widely used phosphoinositide 3-kinase inhibitor, also potently inhibits mammalian polo-like kinase. AB - Polo-like kinases (PLKs) play critical roles throughout mitosis. Here, we report that wortmannin, which was previously thought to be a highly selective inhibitor of phosphoinositide (PI) 3-kinases, is a potent inhibitor of mammalian PLK1. Observation of the wortmannin-PLK1 interaction was enabled by a tetramethylrhodamine-wortmannin conjugate (AX7503) that permits rapid detection of PLK1 activity and expression in complex proteomes. Importantly, we show that wortmannin inhibits PLK1 activity in an in vitro kinase assay with an IC(50) of 24 nM and when incubated with intact cells. Taken together, our results indicate that, at the concentrations of wortmannin commonly used to inhibit PI 3-kinases, PLK1 is also significantly inhibited. PMID- 15664521 TI - Conformation of DNA modified by monofunctional Ru(II) arene complexes: recognition by DNA binding proteins and repair. Relationship to cytotoxicity. AB - We analyzed DNA duplexes modified at central guanine residues by monofunctional Ru(II) arene complexes [(eta(6)-arene)Ru(II)(en)(Cl)](+) (arene = tetrahydroanthracene or p-cymene, Ru-THA or Ru-CYM, respectively). These two complexes were chosen as representatives of two different classes of Ru(II) arene compounds for which initial studies revealed different binding modes: one that may involve DNA intercalation (tricyclic-ring Ru-THA) and the other (mono-ring Ru CYM) that may not. Ru-THA is approximately 20 times more toxic to cancer cells than Ru-CYM. The adducts of Ru-THA and Ru-CYM have contrasting effects on the conformation, thermodynamic stability, and polymerization of DNA in vitro. In addition, the adducts of Ru-CYM are removed from DNA more efficiently than those of Ru-THA. Interestingly, the mammalian nucleotide excision repair system has low efficiency for excision of ruthenium adducts compared to cisplatin intrastrand crosslinks. PMID- 15664520 TI - Single-molecule observation of the catalytic subunit of cAMP-dependent protein kinase binding to an inhibitor peptide. AB - An engineered version of the staphylococcal alpha-hemolysin protein pore, bearing a peptide inhibitor near the entrance to the beta barrel, interacts with the catalytic (C) subunit of cAMP-dependent protein kinase. By monitoring the ionic current through the pore, binding events are detected at the single-molecule level. The kinetic and thermodynamic constants governing the binding interaction and the synergistic effect of MgATP are comparable but not identical to the values in bulk solution. Further, the values are strongly dependent on the applied membrane potential. Additional exploration of these findings may lead to a better understanding of the properties of enzymes at the lipid/water interface. Despite the complications, we suggest that the engineered pore might be used as a sensor element to screen inhibitors that act at either the substrate or ATP binding sites of the C subunit. PMID- 15664522 TI - Tailoring of glycopeptide scaffolds by the acyltransferases from the teicoplanin and A-40,926 biosynthetic operons. AB - The teicoplanin acyltransferase (Atf) responsible for N-acylation of the glucosamine moiety to create the teicoplanin lipoglycopeptide scaffold has recently been identified. Here we use that enzyme (tAtf) and the cognate acyltransferase from the related A-40,926 biosynthetic cluster (aAtf) to evaluate specificity for glycopeptide scaffolds and for the acyl-CoA donor. In addition to acylation of 2-aminoglucosyl glycopeptide scaffolds with k(cat) values of 400 2000 min(-1), both Atfs transfer acyl groups to regioisomeric 6-aminoglucosyl scaffolds and to glucosyl scaffolds at rates of 0.2-0.5 min(-1) to create variant lipoglycopeptides. Using the teicoplanin glycosyltransferase tGtfA, tAtf, and GtfD, a glycosyltransferase from the vancomycin producer, it is possible to assemble a novel lipoglycopeptide with GlcNAc at beta-OH-Tyr(6) and an N(6)-acyl glucosaminyl-vancosamine at Phegly(4). This study illustrates the utility of chemo- and regioselective acyltransferases and glycosyltransferases to create novel lipoglycopeptides. PMID- 15664523 TI - Angiostrongylus vasorum: a real heartbreaker. AB - Recent reports suggest that the canine heartworm Angiostrongylus vasorum is expanding from traditional endemic foci in several parts of the world. We are ill placed to judge the causes and potential consequences of this expansion because of a lack of knowledge about fundamental aspects of the biology of the parasite. We call for a renewed focus on this important but neglected nematode. PMID- 15664524 TI - Climate variability and malaria epidemics in the highlands of East Africa. AB - Malaria epidemics in the highlands of East Africa garner significant research attention, due, in part, to their proposed sensitivity to climate change. In a recent article, Zhou et al. claim that increases in climate variance, rather than simple increases in climate mean values, have had an important role in the resurgence of malaria epidemics in the East African highlands since the early 1980s. If proven, this would be an interesting result but we believe that the methods used do not test the hypothesis suggested. PMID- 15664526 TI - Chemical ecology of insect vectors: temporal, environmental and physiological aspects. PMID- 15664525 TI - Climate variability and malaria epidemics in the highlands of East Africa. AB - The causes of the recent re-emergence of malaria in the East African highlands probably involve a complex interplay among multiple factors, including climate, land use, topography, inadequate use of antimalarial drugs and drug resistance, socioeconomic status, health policy and public health control measures. It is important to determine the relative contribution of these factors. In our study, we statistically attributed the effects of autocorrelation, seasonality and climate variability to the temporal variation in the number of malaria patients in several highland sites in East Africa. We found that in three out of seven sites, climate variability contributed more variance to malaria patient numbers than did autocorrelation and seasonality. In all seven study sites, we found highly significant nonlinear, synergistic effects of the interaction between rainfall and temperature on malaria patient time series. PMID- 15664527 TI - The impact of HIV-protease inhibitors on opportunistic parasites. AB - Opportunistic parasitic infections are an important cause of morbidity and mortality in people infected with HIV. Since the introduction of highly active antiretroviral therapy (HAART), there has been a marked reduction in the occurrence and clinical course of these parasitic infections. Although these changes have been attributed to the restoration of cell-mediated immunity induced by either non-nucleoside reverse transcriptase inhibitors or HIV protease inhibitors, in combination with at least two nucleoside reverse transcriptase inhibitors included in HAART, there is evidence that HIV protease inhibitors have a direct inhibitory effect on the proteases of parasites. The results of studies on opportunistic parasitic infections conducted both before and during the HAART era indicate the need to develop clinical trials on the efficacy of HIV protease inhibitors in controlling parasitic infections in individuals with HIV or other immunocompromised individuals and laboratory investigations on aspartyl proteases of parasites as an important target for the development of new drugs. PMID- 15664528 TI - Gene drive systems in mosquitoes: rules of the road. AB - Population replacement strategies for controlling transmission of mosquito-borne diseases call for the introgression of antipathogen effector genes into vector populations. It is anticipated that these genes, if present at high enough frequencies, will impede transmission of the target pathogens and result in reduced human morbidity and mortality. Recent laboratory successes in the development of virus- and protozoan-resistant mosquito strains make urgent research of gene drive systems capable of moving effector genes into wild populations. A systematic approach to developing safe and effective gene drive systems that includes defining the requirements of the system, identifying naturally occurring or synthetic genetic mechanisms for gene spread upon which drive systems can be based and the successful adaptation of a mechanism to a drive system, should mitigate concerns about using genetically engineered mosquitoes for disease control. PMID- 15664529 TI - The unusual mitochondrial compartment of Cryptosporidium parvum. AB - Recent studies, including the Cryptosporidium parvum Genome Project, have provided evidence for a mitochondrial-derived compartment in this parasite. This organelle appears to lack a genome, and thus must be entirely dependent on nuclear-encoded proteins. Here, we review the evidence for such an organelle in C. parvum and its probable function. There is no adequate treatment for infection by this parasite and so the elucidation of the role of this organelle and the effective targeting of its functions by antimicrobial agents might provide new treatments for infection by C. parvum. PMID- 15664530 TI - The importance of the spleen in malaria. AB - There are several malaria vaccine candidates at various stages of development. Many of these target blood stages of Plasmodium falciparum. The spleen is a key site for removal of parasitized red blood cells, generation of immunity and production of new red blood cells during malaria. This article describes how all of these processes are modified following infection, and suggests that until we fully understand how these processes function and are modulated by infection, appropriate malaria vaccine design and delivery will be extremely difficult to achieve. PMID- 15664532 TI - Determinants of the eradicability of filarial infections: a conceptual approach. AB - Lymphatic filariasis and onchocerciasis are subject to major intervention programs by the WHO. The Onchocerciasis Control Programme in West Africa was launched 30 years ago and has led to considerable insights into the control of this infection. The Global Alliance to Eliminate Lymphatic Filariasis is a relatively recent control program with ambitious targets concerning its efficacy and its schedule. These expectations, however, are based on certain assumptions about the density-dependent processes of limitation and facilitation which determine eradicability: the levels of transmission thresholds and breakpoints. Here, we review these processes operating in filarial infections and show their impact on the persistence of the parasite, as well as pointing out those issues where more information is required to develop sound predictions about the eradicability of these infections. PMID- 15664531 TI - The response of Cryptosporidium parvum to UV light. AB - Ultraviolet (UV) light is being considered as a disinfectant by the water industry because it appears to be very effective for controlling potential waterborne pathogens, including Cryptosporidium parvum. However, many organisms have mechanisms such as nucleotide excision repair and photolyase enzymes for repairing UV-induced DNA damage and regaining preirradiation levels of infectivity or population density. Genes encoding UV repair proteins exist in C. parvum, so the parasite should be able to regain infectivity following exposure to UV. Nevertheless, there is an increasing body of evidence that the organism is unable to reactivate following UV irradiation. This paper describes the effective inactivation of C. parvum by UV light, identifies nucleotide excision repair genes in the C. parvum and Cryptosporidium hominis genomes and discusses the inability of UV-exposed oocysts to regain infectivity. PMID- 15664533 TI - Clear guidance for tough choices. PMID- 15664534 TI - Treatment of emotional lability in ALS. PMID- 15664535 TI - Injuries in patients with epilepsy. PMID- 15664536 TI - Donepezil for memory impairment in multiple sclerosis. PMID- 15664537 TI - Prevention of in-hospital mortality after stroke. PMID- 15664538 TI - Evidence for use of glatiramer acetate in multiple sclerosis. PMID- 15664539 TI - Evidence for use of glatiramer acetate in multiple sclerosis. PMID- 15664541 TI - Reaching beyond the midline: why are human brains cross wired? AB - The crossing of nerve tracts from one hemisphere in the brain to the contralateral sense organ or limb is a common pattern throughout the CNS, which occurs at specialised bridging points called decussations or commissures. Evolutionary and teleological arguments suggest that midline crossing emerged in response to distinct physiological and anatomical constraints. Several genetic and developmental disorders involve crossing defects or mirror movements, including Kallmann's and Klippel-Feil syndrome, and further defects can also result from injury. Crossed pathways are also involved in recovery after CNS lesions and may allow for compensation for damaged areas. The development of decussation is under the control of a host of signalling molecules. Growing understanding of the molecular processes underlying the formation of these structures offers hope for new diagnostic and therapeutic interventions. PMID- 15664542 TI - Essential tremor. AB - Essential tremor (ET) is one of the most common neurological diseases; although it is a disease about which we are only beginning to develop an understanding. Effective treatment options for the disorder are severely limited. The traditional view of ET as a benign, familial, monosymptomatic disorder is being replaced by one of ET as a disease or family of diseases with aetiological, clinical, and pathological heterogeneity. Recent identification of putative environmental toxins linked to ET provide hope for disease prevention through a reduction in exposure to risk factors. Pathological and genetic studies will yield new insights into disease pathogenesis and mechanisms, which may result in the development of more effective symptomatic therapies developed with an understanding of the disease biology. These insights also have the potential to form the basis for neuroprotective therapies. PMID- 15664543 TI - The neuro-ophthalmology of multiple sclerosis. AB - Multiple sclerosis (MS) is the most common disabling neurological disease in young people. Most CNS lesions involve neuroanatomically non-eloquent zones that often do not result in symptomatic complaints. By contrast, tissue-injury mechanisms involving inflammatory demyelination can involve more eloquent sites, such as the optic nerve and brainstem, which can correspondingly produce the development of well recognised syndromes such as optic neuritis and internuclear ophthalmoplegia, respectively. In this review we discuss the broad landscape of abnormalities that affect the afferent visual system and the ocular motor apparatus, and emphasise relevant features, the recognition and treatment of which are of importance to general neurological practice. The commonness of visual sensory and eye movement abnormalities in MS highlights the importance of understanding the principles addressed in this review. PMID- 15664544 TI - Intracranial aneurysm screening: indications and advice for practice. AB - The advent of non-invasive methods of imaging intracranial blood vessels has facilitated screening for intracranial aneurysms in people who are at risk. A strong risk factor for intracranial aneurysm, which causes subarachnoid haemorrhage, is a positive family history, defined as two or more first-degree relatives with subarachnoid haemorrhages (relative risk [RR]=6.6). The greatest familial risk is associated with an affected sibling. Another strong risk factor is polycystic kidney disease (RR=4.4). People who have these risk factors are potential candidates for screening. Even if screening does not find abnormality, there is a high risk of new aneurysms 5 years later. Repeated screening might be done, although the optimum interval between screening assessments and the duration of repeated screening is unclear. Patients who have survived a subarachnoid haemorrhage are at increased risk of another from a newly developed aneurysm, but whether screening is beneficial in such patients is not clear. Most occurrences of subarachnoid haemorrhage in the general population are related to the more common risk factors of hypertension (RR=2.8) and smoking (RR=1.9). Therefore, screening of individuals who have a family history of subarachnoid haemorrhage or polycystic kidney disease will have little effect on the incidence of subarachnoid haemorrhage in the general population. When patients present for screening, the construction of a family tree and discussion of the history of relatives with stroke can be revealing. Before referring a patient for non invasive imaging of the circle of Willis, the physician should discuss the risks and benefits of screening, the implications for holding a licence to drive or fly and for life insurance, and whether or not the patient wants to be informed about other incidental findings. PMID- 15664545 TI - A fatal demyelinating illness in a young woman 10 weeks post partum. PMID- 15664546 TI - Echocardiographic diagnosis of pulmonary embolism: a rise and fall of McConnell sign? PMID- 15664547 TI - Echocardiographic volumetry of the right ventricle. PMID- 15664548 TI - Regional right ventricular dysfunction in acute pulmonary embolism and right ventricular infarction. AB - BACKGROUND: A normally contracting right ventricular apex associated to a severe hypokinesia of the mid-free wall ('McConnell sign') has been considered a distinct echocardiographic pattern of acute pulmonary embolism. OBJECTIVE: To evaluate the clinical utility of the 'McConnell sign' in the bedside diagnostic work-up of patients presenting to the Emergency Department with an acute right ventricular dysfunction due to pulmonary embolism or right ventricular infarction. DESIGN: Among 201 patients, consecutively selected from our clinical database and diagnosed as having massive or submassive pulmonary embolism or right ventricular infarction, 161 were suitable for an echocardiographic review of regional right ventricular contraction and were included in the study. There were 107 cases with pulmonary embolism (group 1) and 54 cases with right ventricular infarction (group 2). All echocardiographic studies were randomly examined by two experienced and independent echocardiographers, blinded to the patient diagnosis and without Doppler informations. RESULTS: The McConnell sign was detected in 75 of 107 patients in group 1 (70%) and in 36 of 54 patients in group 2 (67%); the finding was absent in 32 cases in group 1 and in 18 cases in group 2 (P=0.657). The sensitivity, specificity, positive and negative predictive values of the McConnell sign for the diagnosis of pulmonary embolism were respectively 70, 33, 67 and 36%. CONCLUSIONS: In a clinical setting of patients with acute right ventricular dysfunction the McConnell sign cannot be considered a specific marker of pulmonary embolism. PMID- 15664549 TI - Echocardiographic volumetry of the right ventricle. AB - AIMS: The aim of our study was to define the optimal geometric model for two dimensional volumetry of the right ventricle and its clinical validation. METHODS AND RESULTS: It has been shown in previous reports that an ellipsoidal shell model best reflects the complex right ventricular geometry. We investigated 82 patients without tricuspid regurgitation and intracardiac shunt. Measurements of right ventricular stroke volume were made in modified apical four chamber and parasternal short axis views both in end-diastole and end-systole. The following controls were used:stroke volume of right ventricle estimated by thermodilution in 22 patients, stroke volume of left ventricle calculated with Teichholz formula in 77 patients, stroke volume of left ventricle estimated by Doppler volumetry in 41 patients. The correlations between right ventricular stroke volume by echocardiography and thermodilution, left ventricular volumetry by Teichholz and Doppler were r=0.87, p<0.001; r=0.91, p<0.001; and r=0.83, p<0.001, respectively. CONCLUSION: The ellipsoidal shell model of the right ventricle is a good model for right ventricular volumetry. PMID- 15664550 TI - The feasibility and usefulness of contrast exercise echocardiography for the assessment of left ventricular function in master athletes. AB - BACKGROUND: The number of competitive master athletes (MA, over 40 years) has been rising. Since the incidence of coronary artery disease (CAD) is increasing in this population, cardiovascular pre-participation screening, including a maximum exercise test, is recommended. In this context the addition of contrast to echo could be useful because wall thickening and motion are better markers of myocardial function when the whole endocardial border (EB) is visible. OBJECTIVE: To evaluate the feasibility and usefulness of rest and exercise contrast echo for the assessment of LV wall motion in competitive master athletes with suboptimal acoustic windows. METHODS: Forty consecutive MA underwent echo and contrast echo both at rest and during exercise. Contrast-enhanced images were achieved at rest and at peak exercise after administration of SonoVue (BR1), using apical 4 and 2 chamber views divided into 6 myocardial segments (MS). The EB resolution for each segment was graded as: 0=not visible, 1=barely visible, 2=well-delineated. RESULTS: In the baseline at-rest echo 17/40 (45%) patients were graded as score 0, 22/40 (55%) were graded as score 1 and only 1 athlete has reached score 2. In at-rest echo contrast 28/40 (70%) athletes have reached score 1 and 12/40 (30%) score 2. Nobody was graded as score 0. At the same time in the baseline peak exercise echo 12/40 (30%) athletes were graded as score 0, 24/40 (60%) athletes reached score 1 and 4/40 (10%) score 2, while using contrast at peak-exercise echo 28/40 (70%) were grated as score 1 and 12/40 (30%) reached score 2. The differences about the grading of the score between the two groups with and without contrasts, at rest and at peak exercise, are statistically significant (p<0.001). Considering the whole of the MS analyzed in the majority of the athletic population studied, we can see that at-rest echo, 64/480 MS (13.3%) were graded as 0, 156/480 MS (32.5%) as 1 and 260/480 MS (54.2%) were graded 2, while in the peak-exercise 96/480 MS (20%) were graded as 0, 235/480 MS (48.9%) as 1 and 209/480 MS (43.5%) were graded 2. On the other hand, using contrast, in at rest echo, 480/480 MS (100%) were graded as 2 while in the peak-exercise echo 460/480 MS (95.9%) were graded as 2 and 20/480 MS (4.1%) as 1. The percentage of the MS graded as 2 reach almost the whole number of the analyzed MS with a percentage increment in the at-rest and peak-exercise echo of 85% and 120%, respectively. CONCLUSIONS: Our results show that the use of contrast echo improves the visibility of the EB in a way that the two groups of competitive athletes show at rest and after exercise a significant increment of the score 2 compatible with a better visibility of the EB. These results suggest that contrast echo, both at rest and during exercise, is a better method for EB analysis to understand the behaviour of the wall motion in subjects with suboptimal acoustic windows, and could be suitable for cardiovascular screening in master athletes. PMID- 15664551 TI - Real-time perfusion adenosine stress echocardiography in the coronary care unit: a feasible bedside tool for predicting coronary artery stenosis in patients with acute coronary syndrome. AB - Myocardial contrast echocardiography using power modulation real-time perfusion (RTP) is an appealing method for bedside risk stratification of patients with acute coronary syndrome. In this study, we aimed to evaluate the accuracy in predicting significant coronary stenosis of a bedside RTP adenosine stress protocol in patients with acute coronary syndrome. METHODS: Prior to coronary angiography, 36 consecutive in-patients with acute coronary syndrome underwent a bedside adenosine stress echocardiography with RTP in the coronary care unit. Visual assessment of both perfusion and wall motion was made, comparing rest and hyperaemia images. Each segment was attributed to one of the three main coronary vessel areas. RESULTS: The sensitivity of predicting significant stenosis was 87, 83 and 81% for left anterior descending, circumflex and right posterior descending areas, respectively. Specificity was 69, 67 and 60%, respectively. The positive predictive values were 83, 79 and 74%, respectively. CONCLUSIONS: RTP using adenosine is a feasible bedside tool in predicting the area of significant coronary stenosis and could be helpful as a bedside decision-making tool in the clinical setting. More studies are required to assess the clinical value of RTP adenosine stress echocardiography. PMID- 15664552 TI - Reference values from M-mode and Doppler echocardiography for normal Syrian hamsters. AB - AIMS: Echocardiography has recently been introduced to small animal research, allowing serial measurements of cardiac diseases. In addition, the hamster model has been increasingly used, as it mimics many human heart conditions. However, no reference range of echocardiographic values reflecting normal left ventricular (LV) function exists for hamsters. The purpose of this study was to provide one. METHODS AND RESULTS: The study group consisted of 118 10-week-old, female, Syrian golden hamsters, which underwent high-resolution echocardiography. LV mass was calculated using the corrected cube formula, and LV systolic and diastolic function were assessed by fractional shortening and mitral inflow pulsed-wave Doppler, respectively. The myocardial performance index (MPI) measured the time spent in isovolumic activity and reflected both systolic and diastolic function. The mean+/-SD LV mass, fractional shortening, and MPI were 0.19+/-0.04 g, 44.7+/ 6.6%, and 0.39+/-0.1, respectively. E and A waves were differentiated in 52% of all animals. Logistic regression adjusted with a cutoff of 378 bpm revealed that the risk of E/A wave fusion was 35 times greater (95% CI: 12.6; 98.4) in animals with a heart rate >378 bpm. CONCLUSION: This study documents echocardiographic characteristics in normal Syrian hamsters, which can be used as control values for future studies. PMID- 15664553 TI - Non-invasive assessment of coronary flow reserve in idiopathic dilated cardiomyopathy: hemodynamic correlations. AB - BACKGROUND: Impaired vasodilator myocardial blood flow response has been observed in dilated cardiomyopathy (DCMP). However, the mechanisms responsible for this blunted response are not clear. In the present study, we investigated whether the blunted vasodilator flow response is related to indices of left ventricular performance in patients with idiopathic dilated cardiomyopathy. METHODS AND RESULTS: Eighteen DCMP patients and 12 healthy subjects (C) underwent transoesophageal echocardiography within 48 h from cardiac catheterization. Coronary flow velocity reserve (CFR) was measured in the proximal LAD as the ratio of the peak diastolic coronary flow velocity (Vd-M) after intravenous administration of adenosine to peak baseline diastolic flow velocity (Vd-R). Left ventricular (LV) mass index was positively correlated with baseline coronary diastolic velocity (r=0.415; p=0.043) and inversely correlated with coronary flow reserve (r=-0.570; p=0.003). The baseline coronary diastolic velocity was higher in DCMP vs C (56+/-13 cm/s vs 35+/-12 cm/s; p=0.04). In DCMP pts Vd-R positively correlated with end-diastolic wall stress (r=0.654; p=0.01). Vd increased in both C (96+/-32 cm/s; p<0.05 vs baseline) and DCMP patients (108+/-20 cm/s; p<0.01 vs baseline). The CFR was lower in DCMP patients vs C (1.93+/-0.78 vs 2.99+/-1.01; p=0.009). In DCMP pts CFR was negatively correlated with right atrial pressure (r=-0.595; p=0.015), LVEDP (r=-0.576; p=0.015), pulmonary capillary wedge pressure (PCWP: r=-0.772; p<0.001) and positively with ejection fraction (EF: r=0.683; p=0.003). CONCLUSION: Pts with DCMP have lower CFR compared to controls. This blunted CFR is due to higher baseline coronary flow and reflects higher wall stress. The close relation between CFR and EF, PCWP and LVEDP suggests that not only a higher baseline Vd but also compressive forces due to left ventricular dysfunction might be responsible for the observed blunted adenosine-mediated coronary vasodilation. PMID- 15664554 TI - Functional diagnosis of coronary stenosis using tissue tracking provides best sensitivity and specificity for left circumflex disease: experience from the MYDISE (myocardial Doppler in stress echocardiography) study. AB - AIMS: To evaluate the diagnostic capacity of quantitative analysis of segmental longitudinal myocardial displacement images (tissue tracking, TT) during dobutamine stress echocardiography for the detection of patients with coronary artery disease (CAD). METHODS AND RESULTS: TT-generated colour-coded maps of systolic segmental longitudinal displacement were obtained by post-processing of echocardiographic data from 105 patients with CAD and 90 low risk individuals selected from MYDISE database. Quantitative analysis of the distribution pattern of segmental displacement during dobutamine stress was most successful when a ratio of basal (high amplitude) to apical (low amplitude) colour-coded displacement bands (B/A ratio) was employed. Applied in four different left ventricular sectors, the B/A ratio provided a significant discrimination of patients with CAD (p<0.05 in the anterior and p<0.001 in the inferior wall) as assessed by receiver operating characteristic analysis. The procedure was most sensitive when applied in inferior wall for the detection of left circumflex coronary artery disease, the B/A ratio of 0.8 giving the best combination of sensitivity (77+/-8%) and specificity (77+/-5%) values. CONCLUSION: Quantification of dobutamine stress echocardiography using TT is an efficient diagnostic approach and a valuable additional modality in functional cardiac imaging for the initial identification of patients suspected for CAD. PMID- 15664555 TI - Aorto-right ventricular fistula as an occasional finding. AB - Aorto-right ventricular fistulas are defects of the aortic wall in the area above the right coronary cusp, where it separates aorta and right ventricular outflowtract. Often, these injuries are due to trauma or infective endocarditis. We report an occasional finding of such a fistula, without these causes. There were no other abnormalities on the aortic valve, root or the ascending aorta. PMID- 15664556 TI - Long-term echocardiographic follow-up of a patient with primary pulmonary hypertension receiving iloprost inhalations. AB - Iloprost inhalation has recently emerged as an alternative therapy for severe primary pulmonary hypertension. In the studies documenting the effect of iloprost inhalation therapy, hemodynamic variables have been measured invasively. We have followed a patient with primary pulmonary hypertension receiving iloprost inhalation therapy for 5 years using echocardiography to monitor changes in pulmonary artery pressure and right ventricular function. Echocardiography was also used to evaluate the initial response to iloprost inhalation therapy. This case illustrates the feasibility and utility of echocardiography in the testing and follow-up of iloprost inhalation therapy in patients with pulmonary hypertension. PMID- 15664557 TI - Incomplete myocardial rupture after coronary embolism of an isolated single coronary artery. AB - An 82-year-old female was admitted to the coronary care unit with an anterior wall myocardial infarction and cardiogenic shock. She was in chronic atrial fibrillation without oral anticoagulation. Coronary angiography showed occlusion of the left main coronary artery which originated together with a normal right coronary artery from the right sinus of Valsalva. The advanced age, the presence of chronic atrial fibrillation not anticoagulated and the normal appearance of the remaining coronary arteries suggested a thromboembolic origin. Transthoracic echocardiography showed an abrupt interruption of the myocardial wall, in the apical portion of the interventricular septum, not communicating with the pericardial sac or right ventricular cavity suggesting the presence of an incomplete contained rupture of the myocardial wall at this location. She died in cardiogenic shock due to the extensive left ventricular damage. PMID- 15664558 TI - Traditional contrast echocardiography may fail to demonstrate a patent foramen ovale: negative contrast in the right atrium may be a clue. AB - We describe a patient who suffered a stroke of unknown origin and presented a patent foramen ovale (PFO) at contrast echocardiography. This PFO was clearly detectable after contrast delivery in the femoral vein, while repeated contrast delivery in an antecubital vein only showed a negative contrast effect, which suggests that the blood crossing the PFO originated from the vena cava inferior. However, enhanced detection of a PFO by femoral contrast delivery, compared to antecubital injection has been published many years ago, this mode is not widely implemented yet. With this case report we would like to illustrate that the negative contrast effect may be used as an indicator that a PFO cannot be excluded and a switch to femoral contrast injection is then mandatory. PMID- 15664560 TI - Angiotensin II inhibition of Ca2+ currents is independent of ATR1 angiotensin II receptor activation in rat adult vagal afferent neurons. AB - Angiotensin II (ANG II) has the ability to modulate the activity of neurons involved in the cardiovascular regulation. One effective way of doing that is by changing calcium currents. In the present study, we investigated the effects of ANG II on high-voltage-activated (HVA) Ca2+ currents measured in adult vagal afferent neurons using the whole-cell patch-clamp technique. In addition, we demonstrated the presence of ATR1 and ATR2 receptors mRNA at nodose neurons using conventional reverse transcriptase-polymerase chain reaction (RT-PCR). ANG II (100 nM) decreased the HVA Ca2+ current (peak current recorded at 0 mV: -60.9+/ 8.7 pA/pF in control conditions versus -31.9+/-5.7 pA/pF in the presence of ANG II) and shifted the Ca2+ current activation to a more negative membrane potential (control V0.5=-12.5+/-1.5 mV versus -18.4+/-2.8 mV during perfusion with ANG II). Losartan (500 nM) was not able to prevent the ANG II effect on the HVA Ca2+ current making unlikely the involvement of the ATR1 receptor. When ANG II was perfused in the continuous presence of saralasin, a non-selective ANG II receptor antagonist, we observed a faster but transient inhibition of HVA Ca2+ current. The inhibition was not sustained as observed when we applied ANG II alone and the HVA Ca2+ current recovered with time reaching levels close to the control. Unexpectedly, treatment with the ATR2 blocker PD 123,319 (500 nM) caused a significant inhibition on the HVA Ca2+ current making rather difficult any further conclusions. The above results allow us to conclude that ANG II induced inhibition on the HVA Ca2+ current is probably not via ATR1 receptor activation. PMID- 15664561 TI - Noradrenergic innervation of rabbit pancreatic ganglia. AB - Sympathetic nerve stimulation indirectly regulates pancreatic endocrine and exocrine secretion, in part, through actions on the cholinergic parasympathetic innervation of the secretory tissues. Earlier work identified noradrenergic nerves in pancreatic ganglia and demonstrated the effects of exogenous norepinephrine (NE) on synaptic transmission but no quantitative studies of ganglionic NE content and release exist. Therefore, the distribution and density of catecholamine (CA)-containing nerves in rabbit pancreatic ganglia were studied using paraformaldehyde/glutaraldehyde (FAGLU) staining and HPLC analysis of CA concentrations. Neural release of [3H]NE was measured in ganglia isolated from the head/neck or body regions of the pancreas. CA-containing nerves densely innervated most ganglia (86%) from both regions, while neural and non-neural CA containing cell bodies were rarely found. Ganglia from the head/neck region contained significantly higher concentrations of NE. Both 40 mM K+ and veratridine evoked Ca2+-dependent [3H]NE release and tetrodotoxin inhibited 80% of veratridine-stimulated release. omega-Conotoxin GVIA alone antagonized veratridine-stimulated release by 40% but the addition of nifedipine or omega agatoxin IVA caused no further inhibition. There were no apparent regional differences in the Ca2+-dependence or toxin-sensitivity of NE release. In conclusion, ganglia throughout the rabbit pancreas receive a dense, functional noradrenergic innervation and NE release is dependent upon N- but not P/Q- or L type voltage-dependent Ca2+ channels. These noradrenergic nerves may indirectly regulate pancreatic secretion through actions on ganglionic transmission. PMID- 15664562 TI - Cardiovascular and autonomic nervous functions during acclimatization to hypoxia in conscious rats. AB - The time courses of changes in cardiovascular and autonomic nervous functions during acclimatization to hypoxia were studied in conscious Sprague-Dawley rats. The animals were kept under a 12:12-h light-dark cycle and exposed to hypoxia (1 atm, 10% O2). Implanted telemetry transmitters were used to record blood pressure (BP). Changes in heart rate (HR) and BP were monitored over a 21-day period, and variations before and during hypoxia were analyzed using the wavelet transform method. The HR, high-frequency power of HR variability (HR-HF) and low-frequency power of BP variability (BP-LF) were all significantly increased after 1 h of hypoxia, whereas the LF/HF ratio of HR variability did not change. After this initial increase, both HR and the BP-LF were found to decrease. On the first day of hypoxia, HR and BP-LF values were significantly lower than those of the control rats, whereas the HR-HF was higher. Subsequently, these values altered so that they were similar to the control after 14 days of hypoxia. In addition, the amplitude of diurnal variation in HR was reduced during hypoxia. These results suggest that a sequence of dynamic interactions between sympathetic and parasympathetic nervous activities might have important roles in the regulation of cardiovascular function during acclimatization to hypoxia. PMID- 15664559 TI - Dynamics of fast synaptic excitation during trains of stimulation in myenteric neurons of guinea-pig ileum. AB - Fast excitatory postsynaptic potentials (fEPSPs) occur in bursts in the myenteric plexus during evoked motor reflexes in the guinea-pig ileum in vitro. This study used electrophysiological methods to study fEPSPs during stimulus trains to mimic bursts of synaptic activity in vitro. The amplitude of fEPSPs or fast excitatory postsynaptic currents (EPSCs) declined (rundown) during stimulus trains at frequencies of 0.5, 5, 10 and 20 Hz. At 0.5 Hz, fEPSP or fEPSC amplitude declined by 50% after the first stimulus but remained constant for the remainder of the train. At 5, 10 and 20 Hz, synaptic responses ran down completely with time constants of 0.35, 0.21 and 0.11 s, respectively. Recovery from rundown occurred with a time constant of 7 s. Mecamylamine, a nicotinic cholinergic receptor antagonist, or PPADS, a P2X receptor antagonist, reduced fEPSP amplitude, but they had no effect on rundown. Responses caused by trains of ionophoretically applied ATP or ACh (to mimic fEPSPs) did not rundown. Blockade of presynaptic inhibitory muscarinic, adenosine A1, opioid, alpha2-adrenergic and 5-HT1A receptors or pertussis toxin (PTX) treatment did not alter rundown. Antidromic action potentials followed a 10-Hz stimulus train. Iberiotoxin (100 nM), a blocker of large conductance calcium activated K+ (BK) channels, did not alter rundown. These data suggest that synaptic rundown is not due to: (a) action potential failure; (b) nicotinic or P2X receptor desensitization; (c) presynaptic inhibition mediated by pertussis-toxin sensitive G-proteins, or (d) BK channel activation. Synaptic rundown is likely due to depletion of a readily releasable pool (RRP) of neurotransmitter. PMID- 15664563 TI - Autonomic cardiovascular responses to hypercapnia in conscious rats: the roles of the chemo- and baroreceptors. AB - The role of the autonomic nervous system, the central and peripheral chemoreceptors, and the arterial baroreceptors was examined in the cardiovascular response to hypercapnia in conscious rats chronically instrumented for the measurement of arterial blood pressure (ABP), heart rate (HR), and renal sympathetic nerve activity (RSNA). Rats were exposed to hypercapnia (6% CO2), and the cardiovascular and autonomic nervous responses in intact and carotid chemo- and/or aortic denervated rats were compared. In intact and carotid chemo denervated rats, hypercapnia induced significant increases in mean ABP (MABP) and RSNA, and a significant decrease in HR. The HR decrease was reversed by atropine and eliminated by bilateral aortic denervation, which procedure, however, did not affect the MABP or RSNA response. Bilateral carotid chemo-denervation did not affect the baroreflex control of HR, although this control was attenuated by aortic denervation. Hypercapnia did not affect baroreflex sensitivity in intact rats. These results suggest that hypercapnia induces an increase in MABP due to an activation of sympathetic nervous system via central chemoreceptors and a decrease in HR due to a secondary reflex activation of the parasympathetic nervous system via arterial baroreceptors in response to the rise in ABP. In addition, carotid chemoreceptors do not play a major role in the overall cardiovascular response to hypercapnia in conscious rats. The mechanism responsible for the parasympatho-excitation may also involve CO2 induced aortic chemoreceptor simulation. PMID- 15664564 TI - A hypothesis: autonomic rhythms are reflected in growth lines of teeth in humans and extinct archosaurs. AB - A major determinant of tooth architecture is the arrangement of lines in dentin and in the enamel following the contour of the surface. Since the original description of these lines in the 19th century, they have been attributed to recurring events during tooth development. They have also attracted the attention of dental scientists and anthropologists; however, to date, studies of these structures have been largely theoretical and microscopic. We show here that the statistical properties of the spacing between the lines are similar in teeth from both ancient and modern humans and from extinct archosaurs, reptiles that lived tens or hundreds of millions of years ago-they also resemble heart rate variability of living humans. We propose that the deposition of these recurring structures is controlled by the autonomic nervous system. This control accounts for their regularity and recurrent nature and implies that the lines are an expression of a biologic rhythm which has been conserved throughout evolution. Details of the rhythms give clues to life styles in ancient civilizations and to the physiology of extinct archosaurs. PMID- 15664565 TI - Botulinum toxin A (Botox) and sweating-dose efficacy and comparison to other BoNT preparations. AB - BACKGROUND: Botulinum toxin type A (BoNT/A) is 20-50 times more effective than Botulinum toxin type B (BoNT/B) concerning the treatment of muscular hypercontractions [Sloop, R.R., Cole, B.A., Escutin, R.O., 1997. Human response to botulinum toxin injection: type B compared with type A. Neurology 49, 189 194]. Botulinum toxins block motor nerves as well as autonomic fibres [Rand, M.J., Whaler, B.C., 1965. Impairment of sympathetic transmission by botulinum toxin. Nature 206, 588-591]. OBJECTIVE: Purpose of this study was to analyse the dose dependent reduction of sweating using the BoNT/A preparation Botox and to compare the results with our earlier results analysing Dysport [Braune, C., Erbguth, F., Birklein, F., 2001. Dose thresholds and duration of the local anhidrotic effect of botulinum toxin injections: measured by sudometry. Br. J. Dermatol. 144, 111-117] and Neurobloc (BoNT/B) [Birklein, F., Eisenbarth, G., Erbguth, F., Winterholler, M., 2003. Botulinum toxin type B blocks sudomotor function effectively: a 6 month follow up. J. Invest. Dermatol. 121, 1312-1316]. METHODS: Different doses of Botox were injected subcutaneously (n=27 healthy subjects). Planimetrical analyses of the area of anhidrosis and quantitative sudomotor-axon-reflex testing (QSART) were done after 3 weeks, 3 and 6 months. RESULTS: A threshold dose of 1.25 MU Botox led to anhidrotic skin spots after 3 weeks. The duration of anhidrosis was prolonged for 3 months when 17.5 MU and for 6 months when 50 MU were injected. Anhidrotic area size decreased with time (p=0.001), indicating partial recovery at the edges. After 3 weeks, QSART had significantly decreased to 29% of baseline. With doses of 70 MU or more it decreased to zero. After 3 months, QSART had returned to 68% of baseline and after 6 months to 87%. CONCLUSIONS: Botox dose-dependently suppressed sweating. Comparison to Dysport and Neurobloc revealed a strikingly similar efficacy after 3 weeks and 3 months for all preparations. BoNT/A in general induced a more sustained anhidrosis than BoNT/B. PMID- 15664566 TI - Nonlinear analysis of RR interval in euthymic bipolar disorder. AB - BACKGROUND: The bipolar affective disorder is long been speculated to have an autonomic involvement. Nonlinear analysis of heart rate variability has recently been shown to be a reliable noninvasive test for quantitative assessment of the central sympathovagal interaction that modulates cardiovascular autonomic function. METHODS: We studied 32 euthymic bipolar patients and 24 controls. A high-resolution electrocardiogram was obtained during complete rest. Nonlinear analysis (Poincare plot, largest Lyaponuv exponent, minimal embedding dimension, symbolic dynamic) was used. RESULTS: There wasn't a statistically significant difference in the nonlinear analysis of the heart rate variability, between the euthymic bipolar patients and controls, in the rest situation. CONCLUSION: The nonlinear analysis of heart rate variability didn't support the notation that there is a disturbance in the autonomic nerves system of bipolar patients in the euthymic state. PMID- 15664567 TI - Spectral analysis of slow modulation of EEG amplitude and cardiovascular variables in subjects with postural tachycardia syndrome. AB - OBJECTIVE: Previous studies have reported slow (<0.5 Hz) modulation of electroencephalographic (EEG) background amplitude and suggested that this reflects periodic neuronal activity in the brainstem, such as may be recorded from cardiovascular and respiratory centers in animals. We searched for a relationship between EEG amplitude modulation and modulation of simultaneously recorded cardiovascular variables and attempted to determine whether this relationship was altered in subjects with postural tachycardia syndrome (POTS). METHODS: We recorded EEG, blood flow velocity in the middle cerebral artery (MCA), heart rate, respirations, and blood pressure from subjects with POTS and controls during head-up tilt. Time-frequency analysis of 0.512-s epochs of EEG was performed to determine peak alpha amplitude. Spectra were divided into 3 bands: ultraslow, middle, and respiratory. RESULTS: EEG alpha amplitude modulation in all frequency bands was reduced in POTS subjects while supine. EEG modulation decreased in controls with head-up tilt but not in POTS subjects. Heart rate modulation in the respiratory frequency band decreased with head-up tilt and was significantly less (P<0.02) in ultraslow and respiratory frequency bands in POTS subjects after head-up tilt. Blood pressure and MCA flow velocity modulation in middle and respiratory bands increased with head-up tilt to a greater degree in POTS subjects. Blood pressure and MCA flow velocity modulation frequencies were moderately correlated, but correlations between EEG and cardiovascular variable modulation frequencies were generally low, being highest in the respiratory band but not statistically significant. CONCLUSION: There are subtle differences in EEG amplitude modulation in subjects with POTS. Altered EEG amplitude modulation in POTS may reflect altered brainstem physiology in this disorder. PMID- 15664568 TI - Imaging for thyroglossal duct cyst: the bare essentials. AB - Thyroglossal duct cyst is the most common congenital cyst in the head and neck, and imaging features have been well documented in the literature. However, there are several practical important points to bear in mind during preoperative imaging, which are often overlooked. This review aims to summarize the imaging findings and emphasize important points for trainees and radiologists, particularly those who may encounter this lesion infrequently. PMID- 15664569 TI - Musculoskeletal infections: ultrasound appearances. AB - Musculoskeletal infections are commonly encountered in clinical practice. This review will discuss the ultrasound appearances of a variety of musculoskeletal infections such as cellulitis, infective tenosynovitis, pyomyositis, soft-tissue abscesses, septic arthritis, acute and chronic osteomyelitis, and post-operative infection. The peculiar sonographic features of less common musculoskeletal infections, such as necrotizing fasciitis, and rice body formation in atypical mycobacterial tenosynovitis, and bursitis will also be presented. PMID- 15664570 TI - MR imaging in the diagnosis and management of inheritable musculoskeletal disorders. AB - Although the initial identification of a congenital skeletal malformation is usually made by a combination of clinical examination, conventional radiology and genetic tests, this review illustrates the additional value of magnetic resonance imaging (MRI) in the diagnosis, determination of the extent of the disease, demonstration of associated abnormalities, therapy planning, disease monitoring and prognosis evaluation. PMID- 15664571 TI - The dural tail sign--beyond meningioma. AB - There have been somewhat conflicting reports published about the significance of linear meningeal thickening and enhancement adjacent to peripherally located cranial mass lesions on contrast-enhanced magnetic resonance (MR) images. Most of the authors consider this so-called "dural tail sign" or "flare sign" almost specific for meningioma. This review illustrates the MR imaging findings of a wide spectrum of disorders that show this dural sign. Causes include other extra axial lesions and also peripherally located intra-axial lesions such as neuromas, chloromas, metastases, lymphoma, gliomas, pituitary diseases, granulomatous disorders, and also cerebral Erdheim-Chester disease. The dural tail sign is not specific to a particular pathological process. Nevertheless, useful conclusions can be drawn from the morphology of the lesion, its enhancement pattern, and its solitary or multifocal presentation. The final diagnosis must be based on cerebrospinal fluid studies or histological studies after biopsy. PMID- 15664572 TI - Image guidance of foot joint injections. PMID- 15664573 TI - CT guided diagnostic foot injections. AB - AIM: To describe a CT technique for guiding diagnostic and therapeutic injections in the hind- and mid-foot. MATERIALS AND METHODS: Over a period of 50 months, 28 individuals were referred for diagnostic and therapeutic hind- and mid-foot injections before possible arthrodesis. A CT technique was developed that allowed entry into the various joints using a vertical approach. Numbers of joints injected were as follows: posterior subtalar, 21; talonavicular, 4; calcaneonavicular, calcaneocuboid, navicular-cuneiform and 5th metatarsocuboid joints, 1 each. RESULTS: All injections but one were technically successful. Significant relief of symptoms was noted by 16 participants, whereas for 9 there was no improvement and for 3 a partial response was achieved. CONCLUSION: CT is a simple and safe alternative to fluoroscopy for guiding diagnostic and therapeutic foot injections, and may be the technique of choice in cases of disordered anatomy. PMID- 15664574 TI - Computer-aided detection of pulmonary nodules: influence of nodule characteristics on detection performance. AB - AIM: To evaluate prospectively the influence of pulmonary nodule characteristics on detection performances of a computer-aided diagnosis (CAD) tool and experienced chest radiologists using multislice CT (MSCT). MATERIALS AND METHODS: MSCT scans of 20 consecutive patients were evaluated by a CAD system and two independent chest radiologists for presence of pulmonary nodules. Nodule size, position, margin, matrix characteristics, vascular and pleural attachments and reader confidence were recorded and data compared with an independent standard of reference. Statistical analysis for predictors influencing nodule detection or reader performance included chi-squared, retrograde stepwise conditional logistic regression with odds ratios and nodule detection proportion estimates (DPE), and ROC analysis. RESULTS: For 135 nodules, detection rates for CAD and readers were 76.3, 52.6 and 52.6%, respectively; false-positive rates were 0.55, 0.25 and 0.15 per examination, respectively. In consensus with CAD the reader detection rate increased to 93.3%, and the false-positive rate dropped to 0.1/scan. DPEs for nodules < or = 5 mm were significantly higher for ICAD than for the readers (p < 0.05). Absence of vascular attachment was the only significant predictor of nodule detection by CAD (p = 0.0006-0.008). There were no predictors of nodule detection for reader consensus with CAD. In contrast, vascular attachment predicted nodule detection by the readers (p = 0.0001-0.003). Reader sensitivity was higher for nodules with vascular attachment than for unattached nodules (sensitivities 0.768 and 0.369; 95% confidence intervals = 0.651-0.861 and 0.253 0.498, respectively). CONCLUSION: CAD increases nodule detection rates, decreases false-positive rates and compensates for deficient reader performance in detection of smallest lesions and of nodules without vascular attachment. PMID- 15664575 TI - Intra-individual comparison of patient acceptability of multidetector-row CT colonography and double-contrast barium enema. AB - AIMS: To compare the subjective acceptability of CT colonography in comparison with barium enema in older symptomatic patients, and to ascertain preferences for future colonic investigation. MATERIALS AND METHODS: The study population comprised 78 persons aged 60 years or over with symptoms suggestive of colorectal neoplasia, who underwent CT colonography followed the same day by barium enema. A 25-point questionnaire was administered after each procedure and an additional follow-up questionnaire a week later. Responses were compared using Wilcoxon matched pairs testing, Mann-Whitney test statistics and binomial exact testing. RESULTS: Participants suffered less physical discomfort during CT colonography (p = 0.03) and overall satisfaction was greater compared with barium enema (p = 0.03). On follow-up, respondents reported significantly better tolerance of CT colonography (p = 0.002), and were less prepared to undergo barium enema again (p < 0.001). Of 52 subjects expressing an opinion, all preferred CT to barium enema. CONCLUSION: Patient satisfaction was higher with CT colonography than barium enema. CT colonography caused significantly less physical discomfort and was overwhelmingly preferred by patients. PMID- 15664576 TI - Bladder transitional cell carcinoma: correlation of contrast enhancement on computed tomography with histological grade and tumour angiogenesis. AB - AIM: To investigate the correlation between the degree of contrast enhancement of bladder cancer in the early enhanced phase of helical computed tomography (CT) and microvessel density (MVD), vascular endothelial growth factor (VEGF) and histological grade. MATERIALS AND METHODS: Sixty-five patients with transitional cell carcinoma of the bladder were examined by incremental unenhanced CT and helical CT at 40-45 s after initiation of intravenous administration of contrast medium before surgery. The CT density in Hounsfield units of bladder carcinomas were measured in the middle of the maximum diameter section of the cancer lesions on unenhanced and enhanced CT. The degree of contrast enhancement of the tumour was determined as the absolute increase in Hounsfield units. Histological grade, VEGF and MVD were analysed for each cancer. The Pearson and Spearman correlation tests were used to determine the strength of the relationships between CT enhancement and histological grade, VEGF expression and MVD. RESULTS: Different degrees of enhancement were observed in 91 cancers during the early enhanced phase of helical CT. Mean MVDs and mean CT enhancing values of different histological grade groups were statistically different (p < 0.001). A positive correlation was found in the CT-enhancing value of bladder cancer and MVD (Pearson correlation test; r = 0.938, p < 0.001) and histological grade (Spearman rank correlation; r = 0.734, p < 0.001). VEGF of bladder cancer did not correlate with the change in CT attenuation (Spearman rank correlation; r = 0.087, p = 0.410) and MVD (Spearman rank correlation, r = 0.103, p = 0.330). CONCLUSION: In bladder cancer, the degree of contrast enhancement during the early enhanced helical CT is correlated with the MVD and histological grade of tumour. It is possible that MVD is the histopathological basis of early contrast enhancement of bladder cancer. PMID- 15664577 TI - Effect of reduction in tube current on reader confidence in paediatric computed tomography. AB - BACKGROUND: Computed tomography (CT) contributes significantly to the total radiation dose derived from medical imaging in children. As the number of CT examinations increases there is renewed interest in possible ways to minimize such radiation. AIM: To study the effect of decremental reduction in tube current settings on structural resolution and on reader confidence in being able to reach a final diagnosis. MATERIALS AND METHODS: The study involved spiral CT of children aged 0-13 years. CT performed at reduced mA settings (60-75, 76-90 and 91-130 mA for thoracic, abdominal and pelvic examinations, and 76-90 and 91-130 mA for cranial examinations) were compared with similar investigations at conventional settings (180-240 mA). Images were scored by two blinded readers for structural resolution and diagnostic confidence. Structural resolution was scored on a binary (adequate or not) scale for six structures each in cranial, thoracic and pelvic examinations and for eight structures in abdominal studies, and reader confidence was scored on a four-point scale as 25-100%. Scores from the two readers were averaged for comparison in subgroup analyses. RESULTS: Reduction of tube current settings to 75-90 mA for thoracic, abdominal and pelvic examinations and to 90-130 mA for cranial examinations was not associated with any significant deterioration in image quality. The number of scans considered satisfactory were comparable even at 60-75 mA, although the total relative scores for structural resolution at this setting were lower. CONCLUSION: Tube current settings can be significantly reduced in all paediatric age groups, in all regions. In younger children the impact of such reduction in tube current is likely to be greater. Cranial scans are more sensitive to such reductions as compared to thoracic or abdominopelvic examinations. PMID- 15664578 TI - Accuracy of radiographer plain radiograph reporting in clinical practice: a meta analysis. AB - AIM: To determine the accuracy of radiographer plain radiograph reporting in clinical practice. MATERIALS AND METHODS: Studies were identified from electronic sources and by hand searching journals, personal communication and checking reference lists. Eligible studies assessed radiographers' plain radiograph reporting in clinical practice compared with a reference standard, and provided accuracy data to construct 2 x 2 contingency tables. Data were extracted on study eligibility and characteristics, quality and accuracy. Summary estimates of sensitivity and specificity and receiver operating characteristic curves were used to pool the accuracy data. RESULTS: Radiographers compared with a reference standard, report plain radiographs in clinical practice at 92.6% (95% CI: 92.0 93.2) and 97.7% (95% CI: 97.5-97.9) sensitivity and specificity, respectively. Studies that compared selectively trained radiographers and radiologists of varying seniority against a reference standard showed no evidence of a difference between radiographer and radiologist reporting accuracy of accident and emergency plain radiographs. Selectively trained radiographers were also found to report such radiographs as accurately as those not solely from accident and emergency, although some variation in reporting accuracy was found for different body areas. Training radiographers improved their accuracy when reporting normal radiographs. CONCLUSION: This study systematically synthesizes the literature to provide an evidence-base showing that radiographers can accurately report plain radiographs in clinical practice. PMID- 15664579 TI - MRI findings of eosinophilic myelomeningoencephalitis due to Angiostrongylus cantonensis. AB - AIM: To study the imaging characteristics of eosinophilic myelomeningoencephalitis due to Angiostrongylus cantonensis using magnetic resonance imaging (MRI). MATERIALS AND METHODS: Seventeen MRI examinations of the brain and spinal cord in five patients with angiostrongyliasis cantonensis of the central nervous system were performed. The final diagnosis was based on typical clinical symptoms, results of blood and cerebrospinal fluid (CSF) tests, and the presence of nematode larvae in the CSF. The sequential MRI follow-up examinations were carried out at a different stage for every patient from 1 to 28 weeks after the onset of symptoms. The features of the lesions in the brain, spinal cord, meninges and nerve roots on MRI were studied, moreover, the development of the lesions was analysed on follow-up MRI. RESULTS: Abnormalities were demonstrated on MRI in all five cases. They included three cases of meningoencephalitis, one case of encephalitis and one myelomeningitis. The locations and appearances of the lesions were as follows: (1) brain involvement in four cases (including cerebrum in four, cerebellum in two and brain stem in three), and spinal cord involvement in one case. These lesions were diffuse or scattered and appeared as similar or slightly reduced signal intensity on T1-weighted images (T1WI), high signal intensity on T2-weighted images (T2WI) and turbo fluid attenuated inversion recovery pulse sequence (FLAIR) images. After administration of gadolinium chelate (Gd-DTPA), multiple round or oval enhancing nodules, with diameters ranging from 3 to 10 mm, were seen on T1WI, a few lesions appeared as stick-shaped enhancement whose longest measurement was 14 mm. Diffuse or local oedema around the lesion could be seen. (2) Meningeal involvement in four cases, a case of ependymal involvement and a case of nerve root involvement were among them. These lesions appeared as linear or nodular enhancement of the leptomeninges and ependyma, as well as nerve root enhancement. (3) There was a mild ventricular enlargement in two cases. On follow-up MRI lesions were most severe from the 5th week to the 8th week and it took at least 4-8 weeks (1-2 months) for a lesion to resolve completely, the resolution of larger lesion needed more than 22 weeks. CONCLUSION: Multiple enhancing nodules in the brain and linear enhancement in the leptomeninges were the main features; stick-shaped enhancement was the characteristic sign of the disease on Gd-DTPA enhanced-T1 weighted images. PMID- 15664580 TI - CT herniography in the diagnosis of occult groin hernias. AB - AIM: To evaluate the role of computed tomography (CT) after herniography in the diagnosis and management of primary and recurrent groin hernias not detectable on clinical examination. MATERIAL AND METHODS: Fifty-one patients underwent CT post herniography over a 6-year period for suspected primary or recurrent inguinal hernia. The herniography and post-herniography CT findings were retrospectively compared with clinical and surgical follow-up. Statistical analysis was performed to assess the role of herniography and CT post-herniography in the primary and recurrent groups. RESULTS: Of the 51 patients investigated for occult inguinal hernia, 19 had previous hernia repair with possible recurrence. The most common symptom at presentation was groin pain or discomfort (84%). Seventy-five percent in the primary group and 84% in the recurrent group had no findings on herniography or CT. Nine percent in the primary group and 16% in the recurrent group had hernias diagnosed by herniography. CT did not enhance the detection of hernia. Sensitivity for herniography and CT herniography in the primary groin hernia group was 75% as against specificity, which was 100 and 90%, respectively. For the recurrent groin hernias, sensitivity was 60% for herniography and 40% for CT herniography and specificity 100% for both. CONCLUSION: CT performed post herniography did not provide any benefit over performing herniography alone in the diagnosis of occult primary or recurrent inguinal hernias. PMID- 15664581 TI - Combined antegrade and retrograde ureteral stenting: the rendezvous technique. AB - Ureteral stenting is a routine procedure in endourology. To increase the success rate in difficult cases, it may be helpful to use the rendezvous technique, a combined antegrade and retrograde approach. We performed 16 urological rendezvous in 11 patients with ureteral strictures or urologic lesions. The combined approach was successful in all patients, without morbidity or mortality. In our experience the rendezvous technique increased the success rate of antegrade ureteral stenting from 78.6 to 88.09% (p > 0.05). This procedure is a valid option in case of failure of conventional ureteral stenting. PMID- 15664582 TI - A comparison of inflammatory myopathies at whole-body turbo STIR MRI. PMID- 15664583 TI - "Cystic" pulmonary schistosomiasis. PMID- 15664584 TI - Metastatic manifestations of invasive lobular breast carcinoma. PMID- 15664586 TI - Managing errors in radiology: a working model (1). PMID- 15664587 TI - Managing errors in radiology: a working model (2). PMID- 15664588 TI - Septic arthritis of the major joints of the lower limb after periarticular external fixation application: are conventional safe corridors enough to prevent it? AB - Septic arthritis as a result of pin track infection, following application of external fixators in periarticular fractures of the lower limb, is a rare, but serious complication. Several studies, combining cadaver dissection and MRI scans or conventional X-ray measurements, have tried to define the exact anatomy of the capsular reflection in the major joints of the lower limb (hip, knee and ankle), in order to provide specific safe corridors for extra-capsular wire and pin placement. These studies are reviewed, their methods and results are presented, and their conclusions are evaluated as suggested guidelines for safe extra capsular wire and pin insertion. PMID- 15664589 TI - Cervical spine clearance: a review. AB - Ethical concerns have hindered any randomised control blinded studies on the imaging required to assess the cervical spine in an unconscious trauma patient. The issue has been contentious for many years and has resulted in burgeoning but inconclusive guidance. MRI and multislice CT technology have made rapid advances, but the literature is slower to catch up. Never the less there appears to be an emerging consensus for the multiply injured patient. The rapid primary clinical survey should be followed by lateral cervical spine, chest and pelvic radiographs. If a patient is unconscious then CT of the brain and at least down to C3 (and in the USA down to D1) has now become routine. The cranio-cervical scans should be a maximum of 2 mm thickness, and probably less, as undisplaced type II peg fractures, can be invisible even on 1 mm slices with reconstructions. If the lateral cervical radiograph and the CT scan are negative, then MRI is the investigation of choice to exclude instability. Patients with focal neurological signs, evidence of cord or disc injury, and patients whose surgery require pre operative cord assessment should be imaged by MRI. It is also the investigation of choice for evaluating the complications and late sequela of trauma. If the patient is to have an MRI scan, the MR unit must be able to at least do a sagittal STIR sequence of the entire vertebral column to exclude non-contiguous injuries, which, since the advent of MRI, are now known to be relatively common. Any areas of oedema or collapse then require detailed CT evaluation. It is important that cases are handled by a suitably skilled multidisciplinary team, and avoid repeat imaging due to technical inadequacies. The aim of this review is to re-examine the role of cervical spine imaging in the context of new guidelines and technical advances in imaging techniques. PMID- 15664591 TI - Injuries from biologic material of suicide bombers. AB - BACKGROUND: The management of penetrating wounds caused by suicide bomber bone fragments, contaminated by infectious viruses such as hepatitis or human immunodeficiency virus, is a new medical challenge. The aim of this study was to review the literature and add our experience in the treatment of such wounds. PATIENTS AND METHODS: The files of 94 patients with multiple penetrating fragment injuries of the musculoskeletal system were reviewed. Following any necessary life-saving procedures, the patients underwent wound debridement with delayed wound closure. Broad-spectrum antibiotic treatment was started immediately on admission, and all patients were inoculated with antitetanus toxin and hepatitis B vaccine. RESULTS: The most common injuries were multiple lacerations caused by penetration of small metal fragments. Those that were not removed on initial debridement and continued to cause pain were removed during follow-up. Twenty three patients presented with 49 open fractures caused by the penetrating nails or impact against other objects. Three cases were complicated by chronic osteomyelitis. Three patients presented with injuries caused by bone fragments from the suicide bomber. Samples of bone from one suicide bomber tested positive for hepatitis B virus. None of the patients developed clinical signs of hepatitis B, human immunodeficiency virus or other severe infections during follow-up. CONCLUSIONS: The penetration of biologic material may transmit severe incurable infectious disease. PMID- 15664592 TI - Delayed reaction to shrapnel retained in soft tissue. AB - BACKGROUND: Treatment of penetrating injuries to soft tissues does not require surgical excision of shrapnel. Metals usually remain inert and do not cause damage and are therefore left in soft tissue. OBJECTIVE: Characterization of delayed reaction to shrapnel retained for many years in soft tissue. PATIENTS: Four patients sustained penetrating injuries to the limbs with embedded shrapnel. Many years later, they experienced delayed reaction to the metals that required surgery, with very unusual findings. CONCLUSIONS: Although nonsurgical treatment of shrapnel in soft tissues is the treatment of choice in most cases, we need to be aware of the possibility of late complications requiring surgical treatment. PMID- 15664593 TI - Ballistic variables and tissue devitalisation in penetrating injury--establishing relationship through meta-analysis of a number of pig tests. AB - BACKGROUND: Assessment of the injury potential is central to ascertaining that a law enforcement bullet does not cause unjustified and excessive injuries. There seems to prevail an understanding that tissue devitalisation correlates with kinetic energy dissipated into the tissue. Other views exist too. The purpose of the study was to find out whether such a correlation can be found and at what level of confidence. METHODS: A number of reported tests done with live pigs with sufficient primary data have first been brought to the same temporally comparable level and then analysed. The tests comprise of 140 shots. To maintain consistency tests with other animals were excluded. RESULTS: The best correlation was obtained between excised muscle tissue and dissipated kinetic energy per millimetre of wound channel. An equation describing the relationship between dissipated energy E(d) and devitalised tissue m(deb) is presented as a regression function m(deb)=44.575xE(d)+10.319 with R2=0.293. An experimental method for estimating the energy used for bullet deformation of controlled deformation bullets is also presented. CONCLUSIONS: A method for using the regression function for obtaining tissue destruction figures for any point of wound channel formed in tissue simulant is presented. The figures are intended for meaningful comparison of the injury potential of various bullets and not for forecasting actual tissue injuries. The documentation of the ballistic properties in animal tests also seems somewhat lacking. Some changes in documenting firearms injuries are proposed in order to validate the methods and further enhance the fidelity of simulant testing. PMID- 15664594 TI - Management of polytraumatized patients with associated blunt chest trauma: a comparison of two European countries. AB - BACKGROUND: Blunt chest trauma represents one of the most common injuries in polytrauma patients. Blunt chest injury complicating polytrauma is associated with significant prolongation of intensive care stay. Further, it has a great impact on the timing of fixation of skeletal injuries, possibly contributing to adverse outcome. The purpose of this study is to assess whether there are any differences in the management and outcome of polytrauma patients with blunt chest trauma between trauma units in two different countries. Detailed information about advantages and disadvantages of these two systems might allow optimising the management of blunt chest trauma. PATIENTS AND METHODS: This investigation was performed using the polytrauma database of the German Trauma Society and the British Trauma Audit Research Network. After the definition of the inclusion abbreviated injury scale (AIS(chest) > or = 3) and injury severity score (ISS > 16) and exclusion (AIS(head/neck) > or = 2, referral from outside institutions) criteria, patients were recruited solely from these databases. RESULTS: 188 patients from the German database and 181 patients from the British database were enrolled in this study. Demographic data and injury pattern of the two patient populations did not significantly differ. The volume of initial red blood cell transfusion and length of the intensive care stay were significantly higher in Germany (p < 0.05). Mortality in the UK was 9% higher than in Germany (p = 0.057). Time to death in non-survivors was also significantly longer in Germany (p < 0.05). CONCLUSIONS: The reasons for the differences regarding survival times and survival rates seem to be multiple. German patients received more red blood cells, had a longer hospital stay in intensive care and a better survival rate. The use of kinetic therapy in Germany, not standard in the UK, may contribute to a more favourable outcome. PMID- 15664595 TI - The hormonal and inflammatory responses to pelvic reconstructive surgery following major trauma. AB - Patients undergoing trauma sustain an initial injury followed by further physiological challenges during surgery. Plasma osteocalcin (OC), a marker of osteoblastic activity, declines after major surgery. Increased cortisol secretion, and other components of the perioperative stress response, may play a role in mediating this response. We have examined the osteocalcin, hormonal and cytokine responses in twenty patients undergoing post-traumatic pelvic reconstruction surgery. We measured plasma osteocalcin, serum cortisol, bone specific alkaline phosphatase (BSAP), IL-6, IL-8, IL-10, plasma epinephrine and norepinephrine concentrations for up to 3 days after surgery. We recorded an increase in IL-6, IL-10 and epinephrine concentrations perioperatively and a fall in OC and BSAP concentrations. There were no significant changes in cortisol or IL-8 concentrations. Patients undergoing pelvic reconstruction surgery following trauma have a preserved inflammatory and catecholamine response but the cortisol response may be obtunded. Osteocalcin concentrations are affected by factors other than glucocorticoids. PMID- 15664596 TI - Improving performance in the management of severely injured patients in critical care. AB - OBJECTIVE: To determine opportunities for improvement (OI) in the critical care management of severely injured patients in a general adult intensive care unit through a performance improvement (PI) process. METHODS: Retrospective review of patient records from intensive care patients who had sustained traumatic injuries, except isolated head injury, over a 1-year period. Three assessors independently audited the notes using performance improvement methodology to determine complications, errors in management and preventability. Complications were included when two or more assessors independently detected the complication. MEASUREMENTS AND RESULTS: Records from 90 patients with a diagnosis of 'trauma' were reviewed, 14 patients with isolated head injury were excluded. The mean injury severity score was 23 (range 4-43). No complications or errors of management were identified from 41 patients, including ten patients who died. Seventy-two complications were identified in 35 patients including 15 pneumonias, 6 cases of peri-operative hypothermia and 5 recurrent pneumothoraces. Fourteen preventable complications were identified. CONCLUSIONS: The PI OI process highlighted specific opportunities for the improvement of critical care management of trauma patients in our unit. These will be addressed through the introduction of formal tertiary surveys and clinical management guidelines addressing hypothermia and management of coagulopathy. PMID- 15664597 TI - Open reduction and internal fixation of distal radial fractures using the Pi plate. AB - The authors reviewed 22 patients with 23 fractures of the distal radius, who were treated by open reduction and internal fixation using the low profile contourable titanium dorsal distal radius AO Pi-plate. There were four men and 18 women with a mean age of 57 years. The mean follow up period was 26 months (range, 12-48 months). The modified Lidstrom's radiographic scoring system and the modified Gartland and Werley' functional scoring systems were used for evaluation of the outcome. An excellent clinical outcome was present in four patients, a good in 10, a fair in seven, and a poor in two. Radiographic results were excellent in 12 patients, good in 10, and fair in one patient. Seven patients required removal of the plate for median nerve compression and tendonitis. The Pi-plate can be used in a wide spectrum of distal radial fractures including complex comminuted intra articular (AO type C3) fractures with satisfactory functional outcome and low rate of complication. PMID- 15664598 TI - Distal radioulnar joint dislocation in association with elbow injuries. AB - Traumatic distal radioulnar joint (DRUJ) dislocation with or without an associated fracture is a rare injury. When coupled with a radial head fracture this is commonly known as the Essex-Lopresti injury. We report two cases of elbow dislocation with ipsilateral radial neck fractures and associated true DRUJ dislocations. This has not been previously described in the literature. In elbow injuries with wrist involvement, symptoms in the latter may be subtle. Due to inadequate examination of the affected joint, poor initial radiographic views, and general rarity of this injury, distal radioulnar joint dislocations are frequently missed. We hope our experience illustrates the need to examine thoroughly the joint above and below the injured site, and to be aware of the potential for DRUJ instability in all patients with elbow injuries. PMID- 15664599 TI - Superficial radial nerve damage due to Kirschner wiring of the radius. AB - For displaced fractures of the radius, the use of Kirschner wires (K Wires) is accepted practice either alone or to supplement external fixation. Complications related to K wires include infection, migration and damage to tendons and nerves. We set out to investigate to incidence of superficial radial nerve damage due to radial styloid K wires. Injury to the sensory branch of the superficial radial nerve was seen in eight (20%) out of the 40 patients. K wiring is a popular technique to help maintain anatomic reduction of distal radial fractures. It has the advantage of being a semi-closed procedure, which is simple to perform. However, the morbidity of nerve damage is often underestimated and can be avoided. PMID- 15664600 TI - Use of bio-resorbable implants for stabilisation of distal radius fractures: the United Kingdom patients' perspective. AB - Bio-resorbable implants have been, recently, introduced in the United Kingdom. To our knowledge there have been no randomised studies to assess perception of today's well-informed patients about this new method of fracture stabilisation. In order to assess the patients' perception a prospective study was performed on 100 consecutive adult patients with distal radius fractures. Following detailed verbal and written information about both resorbable and metal implants, the patients were asked to complete a specifically designed questionnaire. Ninety five percent of the patients appreciated the 'resorbable' feature and responded that they would prefer to have their fracture stabilised with a resorbable implant. Conversely, 91% of the participants considered removal as the most negative aspect of the metal implant (p<0.0001). While 56% of the patients felt that it was relatively a new and evolving technology, 29% of them had apprehension about the relative strength of the resorbable implant. Eighty percent of the patients stated that they would be happy to participate in clinical trials to compare the use of bio-resorbable implants versus metal ones (p=0.0001). This study sets the foundation for the implementation of prospective randomised trials to assess the efficacy of the new generation of bio-resorbable implants. PMID- 15664601 TI - External fixation for unstable intra-articular distal radial fractures in women older than 55 years. Acceptable functional end results in the majority of the patients despite significant secondary displacement. AB - Unstable intra-articular distal radial fractures in women older than 55 years were treated by closed reduction and external fixation to achieve the best functional outcome. Sixteen women had radiographic and functional assessment. Despite initial good alignment secondary displacement occurred in 11 patients, probably due to the comminution of the fracture and possibly influenced by osteoporosis. Malunion of the distal radius was seen in two patients and intra articular incongruity with an intra-articular step exceeding 1 mm was observed in two other patients. The functional outcome was excellent or good in 10 and fair in two patients. Four patients had a poor functional outcome. Two of these patients had a significant loss of reduction, one resulting in a malunion. The other two had an intra-articular incongruity of more than 1 mm. Three of the four patients with a poor functional outcome had clinical signs of reflex sympathetic dystrophy. CONCLUSION: Closed reduction and external fixation of "bad case" severely comminuted unstable distal radial fractures in the elderly may result in an acceptable functional outcome in the majority of the cases, although significant secondary displacement occurred in eleven of the sixteen patients. PMID- 15664602 TI - Isolated lateral compartment syndrome after Weber C fracture dislocation of the ankle: a case report and literature review. PMID- 15664603 TI - A rare case of HITTS following prophylactic LMWH therapy in a blunt trauma victim. PMID- 15664604 TI - A fail safe method of inserting end caps into intramedullary nails. PMID- 15664605 TI - An easy and accurate preoperative method for determining tibial nail lengths. PMID- 15664606 TI - Sagittal index in thoraco-lumbar burst fractures. PMID- 15664607 TI - Growth and trace metal accumulation of two Salix clones on sediment-derived soils with increasing contamination levels. AB - The growth and metal uptake of two willow clones (Salix fragilis 'Belgisch Rood' and Salix viminalis 'Aage') was evaluated in a greenhouse pot experiment with six sediment-derived soils with increasing field Cd levels (0.9-41.4 mg kg-1). Metal concentrations of eight elements were measured in roots, stems and leaves and correlated to total and soil water metal concentrations. Dry weight root biomass, number of leaves and shoot length were measured to identify eventual negative responses of the trees. No growth inhibition was observed for both clones for any of the treatments (max. 41.4 mg kg-1 Cd, 1914 mg kg-1 Cr, 2422 mg kg-1 Zn, 655 mg kg-1 Pb), allowing their use for phytoextraction on a broad range of contaminated sediments. However, dry weight root biomass and total shoot length were significantly lower for S. viminalis compared to S. fragilis for all treatments. Willow foliar Cd concentrations were strongly correlated with soil and soil water Cd concentrations. Both clones exhibited high accumulation levels of Cd and Zn in aboveground plant parts, making them suitable subjects for phytoextraction research. Cu, Cr, Pb, Fe, Mn and Ni were found mainly in the roots. Bioconcentration factors of Cd and Zn in the leaves were highest for the treatments with the lowest soil Cd and Zn concentration. PMID- 15664608 TI - Characterization of adsorbent composition in co-removal of hexavalent chromium with copper precipitation. AB - Mechanisms of hexavalent chromium co-removal with copper precipitation by dosing Na2CO3 were studied with a series of well-designed batch tests using solutions containing 150 mg l-1 Cu(II) and 60 mg l-1 Cr(VI). It was found that direct precipitation of chromium through formation of copper-chromium bearing precipitates (in the form of CuCrO4) was one of the main mechanisms contributing to chromium co-removal at pH close to 5.0, and adsorption of chromium at a higher pH by freshly formed copper-carbonate precipitates (adsorbent) contributed to further chromium co-removal. Since, according to solubility products, neither copper-carbonate nor copper-hydroxide precipitates can be produced at pH around 5.0 for a pure 150 mg l-1 copper precipitation, characterization of copper carbonate precipitates (adsorbent) was carried out through developing pC-pH curves of the systems by both equilibrium calculations and MINEQL+ 4.5 (a chemical equilibrium modeling software), and also through laboratory determination of the precipitate composition, such as gravimetric analyses, inorganic carbon percentage and EDAX spectrum analyses. CuCO3.Cu(OH)2, or a combination of CuCO3.Cu(OH)2 (in majority) and Cu(OH)2 (in minority) were suggested to be the major constituent of the precipitates obtained from the copper solution with Na2CO3 dosing. PMID- 15664609 TI - Comparison of EDTA and EDDS as potential soil amendments for enhanced phytoextraction of heavy metals. AB - Phytoextraction has been proposed as an alternative remediation technology for soils polluted with heavy metals or radionuclides, but is generally conceived as too slow working. Enhancing the accumulation of trace pollutants in harvestable plant tissues is a prerequisite for the technology to be practically applicable. The chelating aminopolycarboxylic acid, ethylene diamine tetraacetate (EDTA), has been found to enhance shoot accumulation of heavy metals. However, the use of EDTA in phytoextraction may not be suitable due to its high environmental persistence, which may lead to groundwater contamination. This paper aims to assess whether ethylene diamine disuccinate (EDDS), a biodegradable chelator, can be used for enhanced phytoextraction purposes. A laboratory experiment was conducted to examine mobilisation of Cd, Cu, Cr, Ni, Pb and Zn into the soil solution upon application of EDTA or EDDS. The longevity of the induced mobilisation was monitored for a period of 40 days after application. Estimated effect half lives ranged between 3.8 and 7.5 days for EDDS, depending on the applied dose. The minimum observed effect half life of EDTA was 36 days, while for the highest applied dose no decrease was observed throughout the 40 day period of the mobilisation experiment. Performance of EDTA and EDDS for phytoextraction was evaluated by application to Helianthus annuus. Two other potential chelators, known for their biodegradability in comparison to EDTA, were tested in the plant experiment: nitrilo acetic acid (NTA) and citric acid. Uptake of heavy metals was higher in EDDS-treated pots than in EDTA-treated pots. The effects were still considered insufficiently high to consider efficient remediation. This may be partly due to the choice of timing for application of the soil amendment. Fixing the time of application at an earlier point before harvest may yield better results. NTA and citric acid induced no significant effects on heavy metal uptake. PMID- 15664610 TI - Long-term operation of biofilters for biological removal of ammonia. AB - Biological removal of ammonia was investigated using two types of packing materials, compost and sludge in laboratory-scale biofilters (8l reactor volume). The aim of this study is to investigate the potential of unit systems packed with these supports in terms of ammonia emissions treatment. Experimental tests and measurements included analysis of removal efficiency, metabolic products, and results of long-term operation. The inlet concentration of ammonia applied was 20 200 mg m-3. The ammonia loading rates of 24.9-566 g NH3 m-3 d-1 to compost biofilter (BF3) and 24.9-472 g NH3 m-3 d-1 to sludge biofilter (BF4) were applied for 210 days, respectively. Removal efficiencies of the compost and sludge biofilters were in the range of 97-99% and 95-99%, respectively when the inlet concentration of ammonia was below 110 mg m-3, and the maximum elimination capacities were 288 and 243 g NH3m-3d-1, respectively. However, removal efficiency and elimination capacity of both biofilters significantly decreased as the inlet concentration increased to above 110 mg m-3. By using kinetic analysis, the maximum removal rate of ammonia, Vm, and the saturation constant, Ks, were determined for both packing materials and the value of Vm for compost was found to be larger. Periodic analysis of the biofilter packing materials showed the accumulation of the nitrification product NO3- in the operation. During the experiment, the pressure drops measured were very low. The use of both packing materials requires neither nutritive aqueous solution nor buffer solution. PMID- 15664611 TI - Process evaluation for optimization of EDTA use and recovery for heavy metal removal from a contaminated soil. AB - This study aimed to establish an optimized, closed loop application of ethylenediaminetetraacetic acid (EDTA) in heavy metal removals from a contaminated soil through integrating EDTA recovery/regeneration and metal precipitation processes in the treatment train. Three divalent heavy metals were investigated, namely, Pb, Cd, and Ni. The extractability of the metals by EDTA followed the decreasing order of CdPb>>Ni. The first part of this study was to search for the optimal use of the fresh EDTA in removing these heavy metals from the contaminated soil. The second part of this study was devoted to the recovery/regeneration of the spent EDTA which followed the sequential processes involving (1) complex destabilization by adding ferric ion (Fe(III)) to liberate Pb, Cd, and Ni, (2) precipitation of the liberated Pb, Cd, and Ni in phosphate (PO4(3-)) forms, and (3) precipitation of the excess Fe(III) which eventually produced free EDTA for reuse. The process variables were dosages of Fe(III) and PO4(3-), pH and reaction times. Laborious trial experiments would be needed in searching for the optimum conditions for the above processes. To expedite this exercise, a geochemical equilibrium model, MINTEQA2, was used to find the thermodynamically favorable conditions for recoveries of both EDTA and heavy metals. This was then followed by experimental examination of the process kinetics to observe for the optimal reaction time for each thermodynamically favorable process. This study revealed that 2 h of reaction time each for the complex destabilization reaction and the metal phosphate precipitation reaction was sufficient to achieve equilibrium. With the optimized process condition identified in this study, a total of 95%, 89% and 90% of the extracted Pb, Cd and Ni, respectively, could be precipitated from the spent EDTA solution, with 84% EDTA recovery. The reused EDTA maintained more than 90% of its preceding extraction power in each cycle of reuse. PMID- 15664612 TI - Microbial community and biochemistry process in autosulfurotrophic denitrifying biofilm. AB - The 16S rDNA-based molecular technique was applied to analyze the microbial community of autotrophic denitrification bacteria in a biofilm developed on the surface of sulfur particles and then the biochemistry process involved in this biofilm was discussed based on the microbial community analysis. Six key operational taxonomy units were identified, which were all unknown species belonging to a wide range of bacteria from four major subdivisions (alpha, beta, gamma and delta) of the kingdom Proteobacteria and from the kingdom Chlorobia (green sulfur bacteria). One species was chemoautotrophic and related to Thiobacillus denitrificans, two species were photoautotrophic, and three were chemoheterotrophic. Contrary to expectation, T. denitrificans-like bacteria constituted only 32% of the microbial community. As a result of the study, the entire microbiology of the autosulfurotrophic denitrification process as well as the interactions between the different microbial groups in the biofilm may need to be reconsidered. PMID- 15664613 TI - Adsorption of phenolic compounds by activated carbon--a critical review. AB - Adsorption of phenol and its derivatives on activated carbons is considered based on numerous papers related to this issue. Special attention is paid to the effects of carbon surface functionalities, pH of solution and heterogeneity effects that accompany adsorption of phenolic compounds. Moreover, in this paper the most important aspects are overviewed referring to irreversible adsorption of phenols and impact of different substituents of phenolic compounds on their uptake by activated carbons is considered. Finally, some remarks pertaining to applications of novel adsorbents for phenol adsorption are discussed and illustrated by means of a few examples. PMID- 15664614 TI - Photocatalytic oxidation of gaseous DMF using thin film TiO2 photocatalyst. AB - The heterogeneous photocatalytic oxidation of gaseous N,N'-dimethylformamide (DMF) widely used in the manufacture of synthetic leather and synthetic textile was investigated. The experiments were carried out in a plug flow annular photoreactor coated with Degussa P-25 TiO2. The oxidation rate was dependent on DMF concentration, reaction temperature, water vapor, and oxygen content. Photocatalytic deactivation was observed in these reactions. The Levenspiel deactivation kinetic model was used to describe the decay of catalyst activity. Fourier transform infrared (FTIR) was used to characterize the surface and the deactivation mechanism of the photocatalyst. Results revealed that carbonylic acids, aldehydes, amines, carbonate and nitrate were adsorbed on the TiO2 surface during the photocatalytic reaction. The ions, NH4+ and NO3-, causing the deactivation of catalysts were detected on the TiO2 surface. Several treatment processes were applied to find a suitable procedure for the regeneration of catalytic activity. Among these procedures, the best one was found to be the H2O2/UV process. PMID- 15664615 TI - Adsorption study for the removal of a basic dye: experimental and modeling. AB - An effective adsorbent is developed from saw dust and its various adsorption characteristics are studied for removing a basic dye (crystal violet) from its aqueous solution. Equilibrium data are fitted to various adsorption isotherms. It is seen that about 341 mg of crystal violet can be removed using 1g of the adsorbent at 298 K. Kinetic study is also carried out to observe the effects of various process parameters viz. particle size of the adsorbent, initial concentration of the dye, temperature and adsorbent amount. A generalized two resistance mass transfer model, which includes a film mass transfer coefficient (k(f)) and an internal effective diffusivity (Dp), is used to interpret the adsorption kinetic data. The model parameters (k(f) and Dp) are estimated by fitting the experimental data to the model. The evaluated parameters are used to predict the concentration profiles at various other operating conditions. The average deviation of the predicted values lies within 10% in all the cases. Sensitivity analysis is performed to observe the sensitivity of the model to the variations in the model parameters. PMID- 15664616 TI - The effect of EDTA on Helianthus annuus uptake, selectivity, and translocation of heavy metals when grown in Ohio, New Mexico and Colombia soils. AB - The use of two EDTA concentrations for enhancing the bioavailability of cadmium, chromium, and nickel in three natural soils (Ohio, New Mexico and Colombia) was investigated. The resulting uptake, translocation and selectivity with Helianthus annuus after mobilization were also examined. In general, plants grown in the sandy-loam Ohio soil had a higher uptake that resulted in a selectivity and total metal content of Cd>Cr>>Ni and 0.73 mg and Cr>Cd>>Ni and 0.32 mg for 0.1 and 0.3 g kg-1 EDTA, respectively. With the silty-loam New Mexico soil, although the total metal uptake was not statistically different the EDTA level did alter the selectivity; Cd>Cr>>Ni (0.1 g kg-1 EDTA) and Cd>>Cr>Ni (0.3 g kg-1 EDTA). Conversely, with the Colombian (sandy clay loam) soil increasing the EDTA level resulted in a higher total metal uptake (0.62 mg) than the 0.1 g kg-1 (0.59 mg) treatment. For all three soils, the translocation of Cd was limited. Evaluating the mobile metal fraction with and without EDTA determined that the chelator was capable of overcoming mass transfer limitations associated with the expandable clay fraction in the soils. Root wash results and root biomass concentrations indicated that Cd sorption was occurring. Therefore limited Cd translocation was attributed to insufficient phytochelatin levels. PMID- 15664617 TI - Acid azo dye degradation by free and immobilized horseradish peroxidase (HRP) catalyzed process. AB - Acid azo (Acid Black 10 BX) dye removal by plant based peroxidase catalyzed reaction was investigated. Horseradish peroxidase (HRP) was extracted from horseradish roots and its performance was evaluated in both free and immobilized form. HRP showed its ability to degrade the dye in aqueous phase. Studies are further carried out to understand the process parameters such as aqueous phase pH, H2O2 dose, dye and enzyme concentrations during enzyme-mediated dye degradation process. Experimental data revealed that dye (substrate) concentration, aqueous phase pH, enzyme and H2O2 dose play a significant role on the overall enzyme-mediated reaction. Acrylamide gel immobilized HRP showed effective performance compared to free HRP and alginate entrapped HRP. Alginate entrapped HRP showed inferior performance over the free enzyme due to the consequence of non-availability of the enzyme to the dye molecule due to polymeric immobilization. Standard plating studies performed with Pseudomonas putida showed enhanced degradation of HRP catalyzed dye compared to control. PMID- 15664618 TI - Multivariate analysis of photo-Fenton degradation of the herbicides tebuthiuron, diuron and 2,4-D. AB - The degradation of herbicides in aqueous solution by photo-Fenton process using ferrioxalate complex (FeOx) as source of Fe2+ was evaluated under blacklight irradiation. The commercial products of the herbicides tebuthiuron, diuron and 2,4-D were used. The multivariate analysis, more precisely, the response surface methodology was applied to evaluate the role of FeOx and hydrogen peroxide concentrations as variables in the degradation process, and in particular, to define the concentration ranges that result in the most efficient degradation of the herbicides. The degradation process was evaluated by the determination of the remaining total organic carbon content (TOC), by monitoring the decrease of the concentrations of the original compounds using HPLC and by the chloride ion release in the case of diuron and 2,4-D. Under optimized conditions, 20 min were sufficient to mineralize 93% of TOC from 2,4-D and 90% of diuron, including oxalate. Complete dechlorination of these compounds was achieved after 10 min reaction. It was found that the most recalcitrant herbicide is tebuthiuron, while diuron shows the highest degradability. However, under optimized conditions the initial concentration of tebuthiuron was reduced to less than 15%, while diuron and 2,4-D were reduced to around 2% after only 1 min reaction. Furthermore, it was observed that the ferrioxalate complex plays a more important role than H2O2 in the photodegradation of these herbicides in the ranges of concentrations investigated. PMID- 15664619 TI - Adsorption of Co(II) by a carboxylate-functionalized polyacrylamide grafted lignocellulosics. AB - A new adsorbent (PGBS-COOH) having carboxylate functional group at the chain end was synthesized by graft copolymerization of acrylamide onto banana stalk, BS (Musa Paradisiaca) using ferrous ammonium sulphate/H2O2 redox initiator system. The efficiency of the adsorbent in the removal of cobalt [Co(II)] from water was investigated using batch adsorption technique. The adsorbent exhibits very high adsorption potential for Co(II) and under optimum conditions more than 99% removal was achieved. The maximum adsorption capacity was observed at the pH range 6.5-9.0. The equilibrium isotherm data were analysed using three isotherm models, Langmuir, Freundlich and Scatchard, to determine the best fit equation for the sorption of Co(II) on the PGBS-COOH. A comparative study with a commercial cation exchanger, Ceralite IRC-50, having carboxylate functional group showed that PGBS-COOH is 2.8 times more effective compared to Ceralite IRC-50 at 30 degrees C. Synthetic nuclear power plant coolant water samples were also treated by the adsorbent to demonstrate its efficiency in removing Co(II) from water in the presence of other metal ions. Acid regeneration was tried for several cycles to recover the adsorbed metal ions and also to restore the sorbent to its original state. PMID- 15664620 TI - Degradation of some biorecalcitrant pesticides by homogeneous and heterogeneous photocatalytic ozonation. AB - Photo-Fenton/ozone (PhFO) and TiO2-photocatalysis/ozone (PhCO) coupled systems are used as advanced oxidation processes for the degradation of the following biorecalcitrant pesticides: alachlor, atrazine, chlorfenvinfos, diuron, isoproturon and pentachlorophenol. These organic compounds are considered Priority Hazardous Substances by the Water Framework Directive of the European Commission. The degradation process of the different pesticides, that occurs through oxidation of the organic molecules by means of their reaction with generated OH radical, follows a first and zero-order kinetics, when PhFO and PhCO are applied, respectively. These two Advanced Oxidation Processes, together with the traditional ozone+UV, have been used to investigate TOC reduction of the different pesticide aqueous solutions. The best results of pesticide mineralization are obtained when PhFO is applied; with the use of this advanced oxidation process the aqueous pesticide solutions become detoxyfied except in the case of atrazine and alachlor aqueous solutions for which no detoxification is achieved at the experimental conditions used in the work, at least after 2 and 3 h of treatment, respectively. PMID- 15664621 TI - Catalytic dechlorination kinetics of p-dichlorobenzene over Pd/Fe catalysts. AB - p-Dichlorobenzene (p-DCB) was dechlorinated using Pd/Fe bimetallic catalytic reductants synthesized by chemical deposition. Batch experiments demonstrated that the Pd/Fe bimetallic particles could effectively dechlorinate p-DCB, p-DCB and its intermediate chlorobenzene were removed completely at a Pd loading of 0.02% (weight ratio of Pd to Fe) and Pd/Fe power to solution ratio about 4g 75 ml 1 in 90 min. Dechlorination was affected by various factors such as the reaction temperature, pH, Pd loading percentage over Fe and the introduction of Pd/Fe catalysts et al. Chlorobenzene represents partially stable dechlorinated intermediates in the generation of benzene and part of p-DCB was dechlorinated to benzene indirectly on the surface of Pd/Fe. The dechlorination of p-DCB took place on the surface of the Pd/Fe bimetallic particles in a pseudo-first-order reaction, the activation energy of the dechlorination reaction was determined to be 80.0 kJ mol-1 at the temperature range of 287-313 K. PMID- 15664622 TI - Effects of polyelectrolytes on reduction of model compounds via coagulation. AB - The objective of this research work was to evaluate the performance of enhanced coagulation by alum and polymer. Synthetic source waters containing high molecular weight humic acids, medium molecular weight tannic acids and low molecular weight p-hydroxybenzoic acid were formulated by adjusting the concentration of turbidity and pH; and jar tests were used to study the effect of various types and dosages of polymer on reducing the above model compounds. At a specific pH condition, the applied alum dosage would efficiently decrease the turbidity to 2 NTU follows the order: humic>tannic>p-hydroxybenzoic acid. Adjustment of pH influenced the performance of alum obviously but not of p DADMAC. High p-DADMAC dosage overwhelming the effects of alum is less affected by pH adjustment. The results of this investigation reveal that enhanced coagulation with p-DADMAC was founded to be very effective for removing high-molecular-weight THM precursors, i.e., humic acid and tannic acid, and markedly reduced the alum dosages required for turbidity removal. The other two polymers, i.e., cationic PAM and non-ionic PAM, which had higher molecular weight but lower charge density than p-DADMAC, were not capable of removing organic precursors. It was thus concluded that enhanced coagulation with polymer, p-DADMAC, could be considered as a promising technique for removal of NOMs with hydrophobic and higher-molar mass (>1K) in water treatment plants. PMID- 15664623 TI - Superoxide dismutase evolution and life span regulation. AB - Superoxide is among the most abundant reactive oxygen species (ROS) produced by the mitochondria, and is involved in cellular signaling pathways. Superoxide and other ROS can damage cellular macromolecules and levels of oxidative damage products are positively correlated with aging. Superoxide dismutase (SOD) enzymes catalyze the breakdown of superoxide into hydrogen peroxide and water and are therefore central regulators of ROS levels. Genetic and transgenic manipulation of SOD activities in model systems such as S. cereviseae, mouse and Drosophila are consistent with a central role for SOD enzymes in regulating oxidative stress resistance. Over-expression of SOD in S. cereviseae and Drosophila can reduce oxidative damage and extend life span, but the mechanism(s) are not yet clear. A phylogenetic analysis of publicly available SOD protein sequences suggests several additional conserved gene families. For example, in addition to the well characterized soluble Cu/Zn enzyme (Sod) and mitochondrial manganese-containing form (Sod2), Drosophila melanogaster is found to contain a putative copper chaperone (CCS), an extracellular Cu/Zn enzyme (Sod3), and an extracellular protein distantly related to the Cu/Zn forms (Sodq). C. elegans and blue crab are unusual in having two Mn-containing SODs, and A. gambiae contains an unusual internally repeated SOD. The most parsimonius conclusion from the analysis of the extracellular SODs is that they evolved independently multiple times by addition of a signal peptide to cytoplasmic SOD. PMID- 15664624 TI - Broad spectrum detoxification: the major longevity assurance process regulated by insulin/IGF-1 signaling? AB - Our recent survey of genes regulated by insulin/IGF-1 signaling (IIS) in Caenorhabditis elegans suggests a role for a number of gene classes in longevity assurance. Based on these findings, we propose a model for the biochemistry of longevity assurance and ageing, which is as follows. Ageing results from molecular damage from highly diverse endobiotic toxins. These are stochastic by products of diverse metabolic processes, of which reactive oxygen species (ROS) are likely to be only one component. Our microarray analysis suggests a major role in longevity assurance of the phase 1, phase 2 detoxification system involving cytochrome P450 (CYP), short-chain dehydrogenase/reductase (SDR) and UDP-glucuronosyltransferase (UGT) enzymes. Unlike superoxide and hydrogen peroxide detoxification, this system is energetically costly, and requires the excretion from the cell of its products. Given such costs, its activity may be selected against, as predicted by the disposable soma theory. CYP and UGT enzymes target lipophilic molecular species; insufficient activity of this system is consistent with age-pigment (lipofuscin) accumulation during ageing. We suggest that IIS-regulated longevity assurance involves: (a) energetically costly detoxification and excretion of molecular rubbish, and (b) conservation of existing proteins via molecular chaperones. Given the emphasis in this theory on investment in cellular waste disposal, and on protein conservation, we have dubbed it the green theory. PMID- 15664625 TI - Growth hormone alters methionine and glutathione metabolism in Ames dwarf mice. AB - Reduced signaling of the growth hormone (GH)/insulin-like growth factor-1(IGF 1)/insulin pathway is associated with extended life span in several species. Ames dwarf mice are GH and IGF-1 deficient and live 50-64% longer than wild type littermates (males and females, respectively). Previously, we have shown that Ames mice exhibit elevated levels of antioxidative enzymes and lower oxidative damage. To further explore the relationship between GH and antioxidant expression, we administered GH or saline to dwarf mice and evaluated components of the methionine and glutathione (GSH) metabolic pathways. Treatment of dwarf mice with GH significantly suppressed methionine adenosyltransferase (40 and 38%) and glycine-N-methyltransferase (44 and 43%) activities (in 3- and 12-month-old mice, respectively). Growth hormone treatment elevated kidney gamma-glutamyl cysteine synthetase protein levels in 3- and 12-month-old dwarf mice. In contrast, the activity of the GSH degradation enzyme, gamma-glutamyl transpeptidase, was suppressed by GH administration in heart and liver. The activity of glutathione-S-transferase, an enzyme involved in detoxification, was also affected by GH treatment. Taken together, the current results along with data from previous studies support a role for growth hormone in the regulation of antioxidative defense and ultimately, life span in organisms with altered GH or IGF-1 signaling. PMID- 15664627 TI - The effects of larval density on adult life-history traits in three species of Drosophila. AB - There is evidence that longevity and starvation resistance are determined by a common genetic mechanism. Starvation resistance in Drosophila strongly correlates with both fat content and longevity, and is affected by density during rearing. In this study, we examine how three species, Drosophila melanogaster, Drosophila ananassae and Drosophila willistoni, respond to three larval density treatments. Starvation resistance after adult eclosion, and after 2 days of feeding, and longevity were examined in each sex. D. willistoni reacted differently to larval density than the other two species. This species showed an effect of density on longevity whilst D. ananassae and D. melanogaster showed no such effects. The results also indicate that starvation resistance is not solely determined by fat content. Resistance to starvation at two time points after eclosion differed among species. This may reflect differences in resource acquisition and allocation, and we discuss our findings in relation to how selection may operate in the different species. PMID- 15664626 TI - T-kininogen can either induce or inhibit proliferation in Balb/c 3T3 fibroblasts, depending on the route of administration. AB - T-kininogen (T-KG) is a precursor of T-kinin, the most abundant kinin in rat serum, and also acts as a strong and specific cysteine proteinase inhibitor. Its expression is strongly induced during aging in rats, and expression of T-KG in Balb/c 3T3 fibroblasts results in inhibition of cell proliferation. However, T-KG is a serum protein produced primarily in the liver, and thus, most cells are only exposed to the protein from the outside. To test the effect of T-KG on fibroblasts exposed to exogenous T-KG, we purified the protein from the serum of K-kininogen-deficient Katholiek rats. In contrast to the results obtained by transfection, exposure of Balb/c 3T3 fibroblasts to exogenously added T-KG leads to a dose-dependent increase in [3H]-thymidine incorporation. This response does not require kinin receptors, but it is clearly mediated by activation of the ERK pathway. As a control, we repeated the transfection experiments, using a different promoter. The results are consistent with our published data showing that, under these circumstances, T-KG inhibits cell proliferation. We conclude that T-KG exerts opposite effects on fibroblast proliferation, depending exclusively on the way that it is administered to the cells (transfection versus exogenous addition). PMID- 15664628 TI - Interleukin-6 -174G/C polymorphism and longevity: a follow-up study. AB - Increased rate of inflammation has been observed to be associated with aging. This is manifested, e.g. as increased blood levels of proinflammatory cytokines, such as interleukin-6 (IL-6). The production of IL-6 is, at least partially, genetically determined the single nucleotide polymorphism (SNP) at the promoter ( 174G/C) being decisive. Consequently, some studies have demonstrated that the 174G/C genotype frequencies are different in very old persons as compared to younger ones. However, the results published this far have been conflicting. One of the main confounding factors in these kind of case/control association studies is the undetected difference in the population structure. To avoid this, we now have collected the mortality data of our cohort of 285 nonagenarians (representing mortality between 90 and 95 years of age) and correlated these to the IL-6 genotype. The frequency of -174 allele G was clearly higher in the survivors (n = 114) than in the non-survivors (n = 171). PMID- 15664630 TI - What evidence is there for the existence of individual genes with antagonistic pleiotropic effects? AB - Classical evolutionary theory predicts the existence of genes with antagonistic effects on longevity and various components of early-life fitness. Quantitative genetic studies have provided convincing evidence that such genes exist. However, antagonistic pleiotropic effects have rarely been attributed to individual loci. We examine several classes of longevity-assurance genes: those involved in regulation of the gonad; the insulin-like growth factor pathway; free-radical scavenging; heat shock proteins and apoptosis. We find initial evidence that antagonistic pleiotropic effects are pervasive in each of these classes of genes and in various model systems--although most studies lack explicit studies of fitness components. This is particularly true of human studies. Very little is known about the early-life fitness effects of longevity loci. Given the possible medical importance of such effects we urge their future study. PMID- 15664632 TI - What accounts for the wide variation in life span of genetically identical organisms reared in a constant environment? AB - Individual organisms show marked variability in life span, even when they are of the same genotype and are raised in a common environment protected from extrinsic hazards. This intrinsic variability of life span is thought to arise from the stochastic nature of the cellular and molecular mechanisms controlling development and ageing. In this article we review what is currently understood about the factors underlying the variability of life span and consider the implications for research that aims to improve the predictability of health in old age. PMID- 15664631 TI - What are the effects of maternal and pre-adult environments on ageing in humans, and are there lessons from animal models? AB - An open issue in research on ageing is the extent to which responses to the environment during development can influence variability in life span in animals, and the health profile of the elderly in human populations. Both affluence and adversity in human societies have profound impacts on survivorship curves, and some of this effect may be traceable to effects in utero or in infancy. The Barker Hypothesis that links caloric restriction in very early life to disruptions of glucose-insulin metabolism in later life has attracted much attention, as well as some controversy, in medical circles. It is only rarely considered by evolutionary biologists working on phenotypic plasticity, or by biogerontologists studying model organisms such as C. elegans or Drosophila. One crucial mechanism by which animals can respond in an adaptive manner to adverse conditions, for example in nutrition or infection, during development is phenotypic plasticity. Here we begin with a discussion of adaptive plasticity in animals before asking what such phenomena may reveal of relevance to rates of ageing in animals, and in humans. We survey the evidence for effects on adult ageing of environmental conditions during development across mammalian and invertebrate model organisms, and ask whether evolutionary conserved mechanisms might be involved. We conclude that the Barker Hypothesis is poorly supported and argue that more work in human populations should be integrated with multi disciplinary studies of ageing-related phenomena in experimental populations of different model species that are subjected to nutritional challenges or infections during pre-adult development. PMID- 15664633 TI - Comparative study on the properties of acrylic bone cements prepared with either aliphatic or aromatic functionalized methacrylates. AB - Bone cements prepared with methacrylic acid (MAA) and diethyl amino ethyl methacrylate (DEAEM) were compared with formulations employing 4 methacryloyloxybenzoic acid (MBA) and 4-diethyaminobenzyl methacrylate (DEABM) as comonomer. The influence of these new aromatic monomers on various physicochemical, setting and mechanical properties was assessed. Surface characterization demonstrated that bone cements prepared with any of the functionalized monomers exhibited increasing hydrophilicity with monomer concentration and that the aromatic monomers provided more hydrophilic cements than their aliphatic counterparts for low concentrations of the functional monomer. It was also found that bone cements prepared with high amounts of the acidic aliphatic monomer provided the highest exotherm of reaction and their setting times were shorter than MBA based cements. On the other hand, DEABM containing bone cements exhibited shorter setting times than DEAEM formulations and slightly higher peak temperatures. In general, it was found that the glass transition temperature increased with the presence of acidic comonomer and decreased when alkaline comonomers were present, especially aliphatic ones. When aromatic methacrylates were used at 0.05 molar fraction, the highest tensile and compressive strength were achieved i.e. 46 and 118 MPa for MBA and 51 and 108 MPa for DEABM formulations. A further increase in the aromatic monomer concentration led to cements of low mechanical properties due to solubility problems as revealed by SEM. PMID- 15664634 TI - Development of a strontium-containing hydroxyapatite bone cement. AB - A new route was developed to synthesis a new type of strontium-containing hydroxyapatite (Sr-HAP) bone cement with precursors of tetracalcium phosphate (TTCP), strontium hydrogen phosphate (DSPA), dicalcium phosphate (DCPA), phosphate acid and water. The processing parameters and fundamental properties including pH value, setting time, compressive strength of final hardened body and the cytotoxicity for serial extracts of each cements were investigated. The result shows that the final product of the cement after setting for 24h is nonstoichiometic Sr-containing hydroxyapatite (Ca(10-m-x)Sr(x) square(m)(HPO4)y(PO4)6-y(OH)2-2m square2m, 0 mexiletine > indapamide. PMID- 15664746 TI - On-line membrane preconcentration for continuous monitoring of trace pharmaceuticals. AB - Membrane pervaporation is presented as a method for on-line concentration and monitoring of trace analytes in a simulated pharmaceutical process stream. Pervaporation involves the selective transport of volatile organics across a membrane and into a gas stream. Experiments were carried out using a polar solvent-permeable Nafion membrane and several model pharmaceutical compounds. Solvent reductions greater than 90% and enrichment factors in excess of 7.9 were observed. Residence time and temperature were found to be important operating parameters. Interaction with membrane bound sulfonic acid residues resulted in the loss of reactive analytes such as 1,2-diphenylhydrazine. The concentrated stream was monitored using HPLC and UV/vis detection. Method detection limits were 0.5-1.2 microg/mL and the relative standard deviation for six repeat injections was 3.9-6.2%. PMID- 15664747 TI - Simultaneous quantification of 12 bioactive components of Ligusticum chuanxiong Hort. by high-performance liquid chromatography. AB - A sensitive and specific HPLC-UV method has been developed, for the first time, to simultaneously quantify 12 bioactive ingredients in Ligusticum chuanxiong Hort. (Rhizoma Chuanxiong). This assay was fully validated in respect to precision, accuracy and sensitivity. This method was successfully applied to quantify twelve ingredients in six different Chuanxiong samples. The results demonstrated significant variations in the total content and quantity of each of the main bioactive compounds in different herbs, indicating that quality control of bioactive ingredients in Chuanxiong is critical to ensure its clinical benefits. This assay can be readily utilized as quality control method for Chuanxiong. PMID- 15664749 TI - Implementation of a simple semi-quantitative near-infrared method for the classification of clinical trial tablets. AB - Near infrared transmission spectroscopy combined with chemometrical methods can be applied for identity confirmation of double-blind clinical trial tablets. Samples of two clinical studies, investigating the dose and placebo effect of an experimental drug, were studied. The identity of the blistered tablets was checked using partial least squares beta classification (PLSBC) applied to their NIR transmission spectra. PLSBC is a new supervised classification approach based on partial least squares (PLS) regression combined with beta-error driven class boundaries. It has the ability to limit the probability for misclassification to a known number and therefore providing the method developer a tool for deciding whether the NIR spectra of the different strengths of tablets are specific enough to obtain a robust classification model. The presented approach has the advantage to be applicable on most commercial available near infrared spectroscopy (NIRS) instrumentation software and it can be applied in a GMP environment since validation according to the ICH Q2A and Q2B guidelines on analytical method validation is fast and relatively easy. PMID- 15664748 TI - Separation and characterization of synthetic impurities of triclabendazole by reversed-phase high-performance liquid chromatography/electrospray ionization mass spectrometry. AB - A simple high-performance liquid chromatography/electrospray ionization mass spectrometry (HPLC/ESI-MS) method for the separation and characterization of impurities in the synthesis of triclabendazole has been developed. The analytical separation was achieved on a reversed-phase C18 column using acetonitrile and water (60:40, v/v) as mobile solvent at a flow-rate of 1.0 ml/min at 25 degrees C, and an UV detection at 230 nm. The on-line HPLC/ESI-MSn examinations were performed using ion trap analyzer with extraction ion chromatography (EIC) technique in positive or negative ion modes. The semi-preparative separation was performed with a reversed-phase column using methanol and water (75:25, v/v) as mobile solvent at a flow-rate of 4 ml/min at 25 degrees C, and an UV detection at 230 nm. Thus, two impurities were detected and identified as 5-chloro-6-(2,3,4 trichlorophenoxy)-2-methylsulfanyl-1H-benzoimidazole and 5-chloro-6-(2,3 dichlorophenoxy)-1-methyl-2-methylsulfanyl-1H-benzoimidazole. Meanwhile, some intermediates of impurity-1 in multi-step synthetic reactions, were tracked. Structural elucidation by 1D and 2D NMR and ESI-MSn was discussed. PMID- 15664750 TI - Determination of cocaine and benzoylecgonine by direct injection of human urine into a column-switching liquid chromatography system with diode-array detection. AB - A method for the determination of cocaine (COC) and benzoylecgonine (BZE) in human urine using a column-switching liquid chromatography system is reported. A homemade precolumn (20 mm x 4.6 mm i.d.) dry-packed with Alltech ODS-C18 (35-750 microm) was employed as an extraction precolumn in order to extract and concentrate the COC and BZE from the human urine sample. The analytes were continuously transferred to the analytical column (Spherisorb-C8, 250 mm x 4.6 mm i.d.; dp = 5 microm) by means of the switching arrangement in the backflush mode. Detection was carried out at 235 nm in a UV-diode array detector. The validation of the method revealed analytes quantitative recoveries (96-102%) at three concentrations in the range from 0.25 to 4.00 and from 0.5 to 12.0 microg/mL for COC and BZE, respectively. These values demonstrate the excellent extraction efficiency of the precolumn. The detection limits for COC and BZE at a signal-to noise ratio of 3 were 0.08 and 0.15 microg/mL when a sample volume of 50 microL was injected. The overlap of sample preparation, analysis and recondition of the precolumn increases the sample throughput to four samples per hour. The proposed method has been applied to the determination of COC and BZE in human urine samples from 73 suspecting drug addicts. Urine concentrations of 1.0-118.10 microg of BZE/mL and 0.1-41.0 microg of COC/mL were found. PMID- 15664751 TI - Determination of fluvoxamine and its metabolite fluvoxamino acid by liquid-liquid extraction and column-switching high-performance liquid chromatography. AB - This study describes a new simultaneous determination of fluvoxamine and fluvoxamino acid by automated column-switching high-performance liquid chromatography. The test compounds were extracted from 1.5 ml of plasma using chloroform-toluene (15:85, v/v), and the extract was injected into a hydrophilic metaacrylate polymer column for clean-up and a C18 analytical column for separation. The mobile phase for separation consisted of phosphate buffer (0.02 M, pH 4.6), acetonitrile and perchloric acid (60%) (62.4:37.5:0.1, v/v/v) and was delivered at a flow rate of 0.6 ml/min. The peak was detected using a UV detector set at 254 nm. The method was validated for the concentration range 0.8-153.6 ng/ml for fluvoxamine and 0.6-115.2 ng/ml for fluvoxamino acid, and their good linearity (r > 0.998) were confirmed. Intra-day coefficient variations (CVs) for fluvoxamine and fluvoxamino acid were less than 6.6 and 6.0%, respectively. Inter day CVs for corresponding compounds were 6.3 and 6.5%, respectively. Relative errors ranged from -18 to 9% and mean recoveries were 96-100%. The limit of quantification was 1.2 and 0.9 ng/ml for fluvoxamine and fluvoxamino acid, respectively. This method shows successful application for pharmacokinetic studies and therapeutic drug monitoring. PMID- 15664753 TI - A simple and sensitive LC/MS/MS assay for 7-ethyl-10-hydroxycamptothecin (SN-38) in mouse plasma and tissues: application to pharmacokinetic study of liposome entrapped SN-38 (LE-SN38). AB - An LC/MS/MS method to quantify SN-38 in mouse plasma and tissue homogenates containing liposome entrapped SN-38 (LE-SN38) was developed. Camptothecin (CPT) was used as the internal standard (IS). Sample preparation consisted of simple protein precipitation by acetonitrile containing 0.5% acetic acid. SN-38 and IS were separated by a C18 HPLC column and detected using a mass spectrometer operating in the multiple reaction monitoring (MRM) mode. The peak area of the m/z 393.3-->349.1 transition of SN-38 and that of the m/z 349.1-->305.2 transition of the IS were measured and a standard curve was generated from their ratios. The method had a LLOQ of 0.5 ng/mL in mouse plasma, which corresponds to 2.5 pg for the 5 microL injection volume. The linear range was 0.5-1000 ng/mL of SN-38 in plasma sample spiked with LE-SN38. The LLOQ in tissue homogenates (5%, w/v) quantitation was 1 ng/mL (20 ng/g tissue) of SN-38 in kidney, liver, lung, and spleen homogenates, and 2 ng/mL (40 ng/g tissue) in heart homogenate containing LE-SN38. The assay was linear up to 400 ng/mL of SN-38 in tissue homogenates, and may be extended to 120 microg/mL by proper dilution of samples over the upper limit of quantitation. Acceptable precision and accuracy were obtained for concentrations over the entire standard curve range, both between run and within-run for plasma and tissue homogenates. The method was successfully used to quantify SN-38 in plasma and tissues samples for pharmacokinetic and tissue distribution studies of LE-SN38 in mice. PMID- 15664752 TI - A high throughput approach for simultaneous estimation of multiple synthetic trioxane derivatives using sample pooling for pharmacokinetic studies. AB - The present study describes the application of concept of sample pooling to increase the throughput of pharmacokinetic screening at drug discovery and development stage. An HPLC-UV method for the simultaneous estimation of three synthetic antimalarial compounds 99/357, 99/408 and 99/411 has been developed and validated in rat serum with internal standard for pharmacokinetic profiling. Drug compounds in serum were extracted by two-step liquid-liquid extraction with 2% isopropyl alcohol in n-hexane and quantitated using a validated gradient HPLC-UV method, which was made feasible for all compounds using gradient elution scheme. The method was validated in terms of HPLC reproducibility, linearity, specificity, recovery, accuracy and precision, freeze thaw stability and long term storage stability. Excellent linear relationships (r>0.99) were obtained for calibration as well as analytical standards over a concentration range of 25-1000 ng/ml for three analytes. Recoveries were fond to be >85% for 99/408 and 99/357 and >70% for 99/411. The method developed for three analytes was found to be accurate and precise as bias and percent relative standard deviation (% R.S.D.) values were within limits (<20%). By employing sample pooling approach, plasma level - time profile following single intravenous dose of all three compounds were obtained in a fraction of the time required by conventional single compound dosing and analysis. PMID- 15664754 TI - Rapid determination of tramadol in human plasma by headspace solid-phase microextraction and capillary gas chromatography-mass spectrometry. AB - A simple, rapid and sensitive method for determination of tramadol in plasma samples was developed using headspace solid-phase microextraction (HS-SPME) and gas chromatography with mass spectrometry (GC-MS). The optimum conditions for the SPME procedure were: headspace extraction on a 65-microm polydimethylsiloxane/divinylbenzene (PDMS/DVB) fiber; 0.5 mL of plasma modified with 0.5 mL of sodium hydroxide (0.1 M); extraction temperature of 100 degrees C, with stirring at 2000 rpm for 30 min. The calibration curve showed linearity in the range of 1-400 ng mL(-1) with regression coefficient corresponding to 0.9986 and coefficient of the variation of the points of the calibration curve lower than 10%. The detection limit for tramadol in plasma was 0.2 ng mL(-1). The proposed method was successfully applied to determination of tramadol in human plasma samples from 10 healthy volunteers after a single oral administration. PMID- 15664755 TI - Combined liquid and solid-surface room temperature fluorimetric determination of naproxen and salicylate in serum. AB - A rapid and sensitive method for the determination of naproxen and salicylate in serum is presented. The employed strategy combines solid-phase extraction on a reverse-phase membrane with spectrofluorimetry. Solid-phase extraction under optimum pH conditions makes NX to be retained over the solid surface (where it is directly determined by a fluorimetric technique). Salicylate passes through the disk and is also fluorimetrically determined, but in solution. The linear calibration ranges for NX in the membrane and salicylate in solution were 0.014 0.250 and 0.010-0.250 microg ml(-1), respectively. The lowest value, in each case, is the corresponding limit of quantitation. The performance of the method is demonstrated with the successful determination of both drugs in spiked and real human serum samples. PMID- 15664756 TI - Determination of plasma and brain levels of isotretinoin in mice following single oral dose by high-performance liquid chromatography. AB - An isocratic reversed-phase high-performance liquid chromatographic method was established and validated according to FDA's Guidance for Industry, "Bioanalytical Method Validation", for the determination of isotretinoin in plasma and brain tissue from mice following single and multiple oral doses of Accutane. Plasma sample preparation included deproteination with acetonitrile perchloric acid followed by centrifugation. Brain tissue was homogenized and extracted with acetonitrile-perchloric acid followed by centrifugation. The supernatants were analyzed by high-performance liquid chromatography (HPLC). Benz[alpha]anthrancene-7,12-dione was used as the internal standard. Chromatographic separation was achieved on a C18 column using an acetonitrile aqueous 0.5% acetic acid (85:15, v/v) elution. The average extraction efficiency was >95% for plasma and >82% for brain. The lower limit of quantification was 30 ng/mL for plasma and was 30 ng/0.1g for brain tissue, respectively. The linear range for plasma was 30-600 ng/mL, and 15-300 ng/0.1g for brain. Maximum concentrations of isotretinoin in both plasma and brain were observed at 1h after single oral dosing (25 mg/kg). The maximum concentrations in plasma and brain were 2.36 microg/mL and 0.34 microg/g, respectively. The mean area under curve (AUC) in plasma was 6.13 microg h/mL. The mean eliminate half-life in plasma was estimated as 46 min. PMID- 15664757 TI - Simultaneous determination of N-hydroxymethyl-N-methylformamide, N methylformamide and N-acetyl-S-(N-methylcarbamoyl)cystein in urine samples from workers exposed to N,N-dimethylformamide by liquid chromatography-tandem mass spectrometry. AB - N-Hydroxymethyl-N-methylformamide (HMMF) and N-methylformamide (NMF) in urine samples from workers exposed to N,N-dimethylformamide (DMF) cannot be distinguished by a gas chromatographic method because HMMF is converted to NMF at the injection port of gas chromatography (GC). Total NMF (HMMF+NMF) has been measured instead. Also, the determination of N-acetyl-S-(N methylcarbamoyl)cystein (AMCC), which is supposed to be related to the toxicity of DMF, needs multiple treatments to convert to a volatile compound before GC analysis. There is no previous report of a simultaneous determination of three major metabolites of DMF in urine. The aim of this study is to develop a simple and selective method for the determination of DMF metabolite in urine. By using a liquid chromatography-tandem mass spectrometry, we can directly distinguish these three major metabolites of DMF in a single run. The diluted urine samples were analyzed on Capcell Pak MF SG80 column with the mobile phase of methanol in 2mM formic acid (10:90, v/v). The analytes were detected by an electrospray ionization tandem mass spectrometry in the multiple-reaction-monitoring mode. The standard curves were linear (r>0.999) over the concentration ranges of 0.004-8 microg/mL. The precision and accuracy of quality control samples for inter-batch (n=6) analyses were in the range of 1.3-9.8% and 94.7-116.8, respectively. The sum of each HMMF and NMF concentration determined by LC-MS/MS method shows high correlation (r=0.9927 with the slope of 1.0415, p<0.0001) with NMF included HMMF concentration determined by GC method for 13 urine samples taken from workers exposed to DMF. The excretion ratio of HMMF:NMF:AMCC is approximately 4:1:1 in molar concentration. PMID- 15664759 TI - Evaluation of solid state form of troglitazone by solid state NMR spectroscopy. AB - The solid state forms of troglitazone drug substance and diastereomers were characterized using solid state nuclear magnetic resonance (SSNMR) spectroscopic method. The SSNMR spectroscopy could distinguish the hydrated and the non hydrated RR/SS forms more clearly than powder X-ray diffractometry (PXRD). The SSNMR result supported that troglitazone drug substance consists of diastereomers as a simple physical mixture. SSNMR spectroscopy was also able to characterize the solid state forms of troglitazone in tablets while PXRD was unable to because of interference from the pharmaceutical additives. Troglitazone was proved to exist in amorphous form in tablets, and keep its solid state form amorphous against heat and humidity. SSNMR spectroscopy thus provides very important information for the development of the pharmaceutical formulation of troglitazone. PMID- 15664758 TI - Diagnostic value of bone remodeling markers in the diagnosis of bone metastases in patients with breast cancer. AB - Metastatic spread to bone is common in patients with breast cancer and its early detection is required for the better management of these patients. Several biochemical markers of bone remodeling have been recently developed, in order to assess metastatic bone disease with non radiologic methods. The pyridinolin cross linked amino-terminal telopeptide of type I collagen (NTx) has been measured in serum and urine as a specific marker of bone collagen breakdown, while the bone isoform of alkaline phosphatase (BAP) has been used to determine bone formation activity. Thirty-three consecutive ambulatory patients with metastatic breast cancer and bone metastases and 31 with extraskeletal metastases only, matched for age and menopausal status, were studied. Serum levels of NTx and BAP were measured by enzyme-linked immunosorbent assays. The diagnostic accuracy of both markers was evaluated by receiver operating characteristic (ROC) analysis. Patients with bone metastases had significantly higher levels of NTx (37.0+/-36.9 nM BCE versus 23.5+/-21.0 nM BCE, P<0.05) and BAP (57.8+/-31.7 U/L versus 36.5+/ 28.5 U/L, P<0.01) compared to those without bone metastases. NTx was positively correlated with BAP (R=0.340, P<0.01). The area under the ROC curve was 0.671 for NTx and 0.755 for BAP. Using a cut-off value of 29.7 nM BCE for NTx, specificity and sensitivity were 87.1% and 45.5%, respectively; in the case of BAP, using a cut-off value of 50.6 U/L, the specificity and sensitivity were 90.3% and 54.5%, respectively. In patients not receiving concomitant hormonal treatment, the area under the ROC curve was 0.724 for NTx and 0.822 for BAP; in this subgroup of patients, using a cut-off value of 30.0 nM BCE for NTx, the specificity and sensitivity were 96.2% and 47.1%, respectively, while using a cut-off value of 50.0 U/L for BAP, the corresponding percentages were 92.3% and 70.6%. Although serum NTx and BAP are quite specific, they are not sensitive enough to diagnose bone metastases in patients with advanced breast cancer. Their diagnostic accuracy, however, is considerably enhanced in patients not receiving hormonal therapy. PMID- 15664760 TI - Simultaneous LC determination of tizanidine and rofecoxib in tablets. AB - A reverse phase high performance liquid chromatographic method to determine tizanidine (TZ) and rofecoxib (RF) in combination is proposed and applied to the pharmaceuticals. This method allows the determination of 0.1-0.5 microg/ml of TZ and 1.2-6.0 microg/ml of RF along with 10 microg/ml of nimesulide (internal standard), in a mobile phase consisting of 1% (v/v) triethylamine (pH adjusted to 2.5 using dilute orthophosphoric acid):acetonitrile in the ratio 55:45% (v/v). Detection wavelength of 303 nm and flow rate of 0.8 ml/min were fixed for the study. The limit of detection (LOD) for TZ and RF were found to be 10 and 1 ng/ml, respectively. The limit of quantification (LOQ) for TZ and RF were found to be 80 and 12 ng/ml, respectively. The amount of drug present in the tablet and the recovery studies were also carried out. The % R.S.D. of recovery studies for TZ and RF were found to be 0.0673 and 0.0146, respectively. The method is validated for accuracy, precision, ruggedness and robustness. PMID- 15664761 TI - High-performance liquid chromatography electrospray ionization mass spectrometry determination of tulobuterol in rabbit's plasma. AB - A sensitive and specific liquid chromatography electrospray ionization mass spectrometry (LC-ESI-MS) method has been developed and validated for the identification and quantification of tulobuterol in rabbits' plasma. After the addition of clenbuterol-HCl, the internal standard (IS) and 1.0 M sodium hydroxide solution, plasma samples were extracted using a solvent mixture comprised of 5% isopropanol in n-hexane. The compounds were separated on a prepacked Lichrospher CN (5 microm, 150 mm x 2.0 mm) column using a mixture of methanol-water (10 mM CH3COONH4, pH 4.0) as mobile phase. A Shimadzu LCMS-2010A mass spectrometer connected to a Shimadzu high performance liquid chromatograph (HPLC) was used to develop and validate the method. The method has shown to be sensitive and specific by testing six different blank plasma batches. Linearity was established for the range of concentrations 0.50-40.0 ng/mL with a coefficient of determination (r) of 0.9998. The intra-day precision was better than 15%. The lower limit of quantification (LLOQ) was identifiable and reproducible at 0.50 ng/mL. The proposed method enables the unambiguous identification and quantification of tulobuterol for pharmacokinetic, bioavailability or bioequivalence studies. PMID- 15664762 TI - Differential pulse voltammetric determination of the dopaminergic agonist bromocriptine at glassy carbon electrode. AB - The electrochemical oxidation of bromocriptine at glassy carbon electrode has been carried out in Britton-Robinson (B-R) buffer solutions in the pH range 2.0 11.0 employing cyclic, linear sweep and differential pulse voltammetry (DPV). Bromocriptine showed one well-defined oxidation peak accompanied by a smaller one. The oxidation process was found irreversible. For analytical purposes, the well-resolved diffusion controlled voltammetric peak at pH 5 was critically investigated. The linear relationship between peak current height and bromocriptine concentration allowed the differential pulse voltammetric determination of the drug over a wide concentration range, from 0.04 to 5.00 microg ml(-1) with a detection limit of 0.01 microg ml(-1). A relative standard deviation of 1.44% at 0.1 microg ml(-1) level was obtained. The proposed DPV method was successfully applied for the individual tablet assay to verify the uniformity content of bromocriptine in commercial tablets. PMID- 15664763 TI - Voltammetric study of danazol and its determination in capsules and spiked biological fluids. AB - The voltammetric behaviour of danazol DZ (antigonadotropin) was studied using cyclic voltammetry, direct current, differential pulse polarography (DPP) and alternating current polarography. Danazol exhibited irreversible cathodic waves over the pH range of 1-5 in Britton Robinson buffers. At pH 1 (the analytical pH), a well-defined wave with E1/2 of -1.04 V versus Ag/AgCl reference electrode was obtained. The diffusion current constant (Id) was 4.8+/-0.14 microA.L.m mole( 1) and the current-concentration plot was rectilinear over the range from 5 x 10( 6) to 1 x 10(-4) M with correlation coefficient (n = 11) of 0.995. The calculated detection limit was 1 x 10(-6) M using the DPP mode. The wave was characterized as being irreversible, diffusion-controlled although adsorption phenomenon played a limited role in the electrode process. The proposed method was applied to commercial capsules and the average percentage recovery was in agreement with that obtained by the official USP method. The method was extended to the in vitro determination of DZ in spiked human urine and plasma samples, the percentage recoveries were 96+/-4 and 97+/-5, respectively. A proposal of the electrode reaction was postulated. PMID- 15664764 TI - Zonisamide monotherapy for epilepsy in children and young adults. AB - Several of the newer antiepilepsy drugs have not been tested as monotherapy in controlled trials. Zonisamide is a broad-spectrum antiepilepsy drug indicated for the adjunctive treatment of partial seizures in adults. However, several small, open-label studies have indicated that it may be safe and effective as monotherapy. The present chart review study was conducted to evaluate the safety and effectiveness of zonisamide monotherapy in a pediatric and young adult patient group. Patient records at the Blue Bird Circle Clinic for Pediatric Neurology were reviewed to identify patients receiving zonisamide monotherapy. Efficacy was assessed from seizure diaries and patients' subjective evaluations. Safety and tolerability were evaluated by analysis of adverse events and change in body weight. The study included 131 patients aged 1 to 21.8 years with a broad spectrum of seizure types and epilepsy syndromes. A total of 101 patients (77.1%) achieved a 50% or greater decrease in seizure frequency, including 39 patients who achieved seizure freedom. Zonisamide monotherapy was well tolerated, with three patients (2.3%) discontinuing for adverse events. These results support open-label studies from Japan reporting that zonisamide monotherapy is safe and effective in pediatric and young adult patients. PMID- 15664766 TI - Popliteal angle in preterm infants with periventricular leukomalacia. AB - The aim of this study is to clarify the usefulness of popliteal angle in infants with periventricular leukomalacia. The popliteal angle was measured at 1, 4, 8, and 12 months of corrected age in 47 infants with periventricular leukomalacia and in 103 control infants with normal development. The popliteal angle was categorized into two groups; tight, when it was < or =120 degrees , and wide, when it was >120 degrees . The severity of diplegia was defined at 2 years as follows: mild, when an infant could walk; moderate, when an infant could sit but could not walk; severe, when an infant could not sit. Tight popliteal angle was more often observed in infants with periventricular leukomalacia than in normal infants at any corrected age. The rate of tight popliteal angle was significantly lower in infants with mild diplegia than in those with moderate or severe diplegia at 8 and 12 months of corrected age. Specificity and positive predictive value were high at 8 and 12 months of corrected age, whereas sensitivity was relatively low. The results of this study suggest that evaluation of popliteal angle is useful for detection of infants with periventricular leukomalacia, although false-negative rate is high during late infancy. PMID- 15664765 TI - Ketogenic diet in pediatric epilepsy patients with gastrostomy feeding. AB - Ketogenic diet is effective in the control of intractable seizures. Poor compliance is a major limiting factor. In one study, only 50% of children receiving the oral ketogenic diet remained on the diet after 1 year. Twelve children with static encephalopathy and intractable symptomatic epilepsy were given the ketogenic diet via gastrostomy tube. Mean age was 3 years (range, 7 months to 6.5 years). Mean seizure frequency at baseline was 199/month. Seizure frequency after 12 and 18 months of diet was compared with baseline. After 12 months on the diet, the number of antiepileptic drugs was compared with baseline. Median seizure reduction at 1 year and 18 months was 61% and 66%, respectively (P = 0.02). Individually, six patients had 90% seizure reduction, one had 75% reduction, three had 50% reduction, and two patients did not improve. Mean antiepileptic drugs at baseline was 2.8; at 12 months 1.6 (49% reduction). Three patients had weight loss. Two patients discontinued the diet at 13 months and 21 months, respectively, because of diarrhea and weight loss. Compliance with diet was 100% during treatment. This study suggests that the ketogenic diet via gastrostomy feeding tube is safe and effective in children with intractable seizures and ensures compliance. PMID- 15664767 TI - Improved outcome for very low birth weight multiple births. AB - This study describes time trends for very low birth weight multiple births in relation to very low birth weight singletons. Two cohorts of very low birth weight (less than 1250 gm) children recruited between 1983-85 (cohort 1, n = 115) and 1992-94 (cohort 2, n = 144) were compared. The Bayley Scales of Infant Development and a standardized neurologic examination were administered at 2 years corrected age. Neurodevelopmental outcome did not change between cohort 1 and 2 for singletons. For multiple births, mean Mental Developmental Index increased after adjustment for neonatal risk factors [adjusted mean (standard deviation) 81.8 (11.7) to 96.5 (18.6), analysis of covariance P = 0.007]. The prevalence of cerebral palsy decreased, however not significantly [adjusted odds ratio (95% confidence interval) 0.3 (0.1-1.5), P = 0.14]. The proportion of disease-free survival (no cerebral palsy and no developmental delay) increased for multiple births (7-37%, P = 0.002), but not for singletons. In cohort 2, neurodevelopmental outcome of multiple births was similar to that of singletons. The cognitive outcome of very low birth weight multiple births improved, possibly because of changes in perinatal practice. However, neurodevelopmental outcome was similar to that of very low birth weight singletons who were unaffected by changes in neonatal care with high proportions of motor delay and cerebral palsy. PMID- 15664768 TI - Spectrum of corpus callosum agenesis. AB - Magnetic resonance imaging and clinical features of 16 children with agenesis of the corpus callosum are reviewed. The patients were classified in three groups. Eight children had agenesis of the corpus callosum without substantial involvement of other brain areas. This group included patients with interhemispheric cyst, lipoma, or partial agenesis (hypogenesis) of the corpus callosum. Four patients in the second group manifested severe telencephalic dysgenesis in addition to agenesis of the corpus callosum. In the third group, agenesis of the corpus callosum was a component of a syndrome in four children. The outcome was not favorable for the majority of patients, even in the first group without significant involvement of other systems. Severe handicaps with developmental delay, intellectual disability, and neurologic deficit were observed in patients with additional telencephalic dysgenesis or syndromic form of corpus callosum agenesis. PMID- 15664769 TI - Accelerated head growth in early development of individuals with autism. AB - Macrocephaly is one of the most consistent physical findings reported in autistic individuals. Previous studies attempted to determine if macrocephaly is associated with risk for autism. This study hypothesizes that an abnormal acceleration in head growth during early development, rather than macrocephaly, is associated with autism risk. To investigate this hypothesis, head circumference data were examined in 251 individuals from 82 multiplex (at least two individuals with autism) and 113 sporadic (no family history) families with autism. This examination included longitudinal measurements for 79 individuals. Nineteen percent of the original 251 individuals were found to have macrocephaly (head circumference >97%). Abnormal acceleration in head growth was defined as an increase of 25 or more percentile points in head circumference between two consecutive measurements. Thirty-five percent of individuals with multiple head circumference records had an abnormal increase in head circumference. Furthermore, autistic individuals with accelerated head growth in early childhood displayed higher levels of adaptive functioning and less social impairment. This study confirms the presence of abnormal acceleration in head growth during the first and second months of life in a subgroup of autistic individuals. PMID- 15664770 TI - Frontal white matter reductions in healthy males with complex stereotypies. AB - The pathophysiologic mechanism for stereotypic, bilateral repetitive movements involving the arms and hands (complex motor stereotypies) is unknown. This study used volumetric magnetic resonance imaging to compare cerebral lobes and caudate nucleus in six males with complex stereotypies and average intelligence to age matched control subjects. Results indicated volumetric reductions in frontal white matter, disproportionate to total cerebral white volume, and in the left and right caudate nuclei. These preliminary data suggest a possible dysfunction of cortico-striatal-thalamo-cortical circuitry in children with nonautistic, physiologic motor stereotypies. PMID- 15664771 TI - Effect of antiepileptic drug polytherapy on crystalluria. AB - Urolithiasis is a rare side effect of antiepileptic drugs. To clarify the risk factors for urolithiasis induced by antiepileptic drugs, the effect of antiepileptic drug monotherapy on crystalluria was studied, and zonisamide or sulthiame therapy and alkaline urine were demonstrated to be risk factors. In the next investigation, the effect of antiepileptic drug polytherapy on crystalluria was retrospectively studied in epilepsy patients treated for more than 1 month during the last 7 years. A total of 278 urine specimens from epilepsy patients aged between 7 months and 36 years were enrolled in this study. The mean age was 12.3 years. There were 109 samples from females and 169 from males. Antiepileptic drugs administered in this study were valproate (174 urinary samples), zonisamide (139), carbamazepine (138), phenobarbital (65), phenytoin (52), acetazolamide (17), clonazepam (15), sulthiame (6), ethosuximide (6), nitrazepam (4), and clobazam (4). Epilepsy patients treated with antiepileptic drug polytherapy were frequently found to have crystalluria in patients demonstrating alkaline urine and taking acetazolamide, zonisamide (particularly with high serum levels), or many antiepileptic drugs in combination. Regular urinalysis seems to be necessary in these patients, and the evaluation for urolithiasis should be performed if persistent crystalluria is demonstrated. PMID- 15664772 TI - Maternal uniparental disomy chromosome 14: case report and literature review. AB - Uniparental disomy is a genetic cause of disease implicated in a wide variety of neurologic disorders. A recently identified condition is maternal uniparental disomy for chromosome 14 (mUPD14) syndrome. A child with hypotonia and developmental delay was found to have mUPD14 after identification of a balanced karyotypic rearrangement involving both chromosomes 14. We explore the genetic mechanisms by which uniparental disomy can cause clinical abnormalities and karyotypic findings that should raise suspicion for uniparental disomy, review the literature on the mUPD14, and discuss clinical indications on which to suspect this diagnosis. Although it is more difficult to establish a diagnosis in the absence of visible karyotypic abnormalities involving chromosome 14, a distinct phenotype exists in mUPD14 syndrome: in utero growth restriction, congenital hypotonia, gross motor delay, arrested hydrocephalus, mild to moderate mental retardation, joint hyperextensibility, short stature, and precocious puberty. Testing for mUPD14 should be considered in infants with generalized hypotonia who have a history of in utero growth restriction. PMID- 15664773 TI - Underdiagnosis of mild congenital disorders of glycosylation type Ia. AB - Congenital disorders of glycosylation-Ia are the most frequent type of congenital disorders of glycosylation. This condition affects the nervous system as well as other organs. The estimated incidence of congenital disorders of glycosylation-Ia is higher than the number of identified cases, therefore underdiagnosis of this heterogeneous disorder is probable. Neurologic and biologic signs are hallmarks for the identification of patients with congenital disorders of glycosylation-Ia. This report describes two children with congenital disorders of glycosylation-Ia syndrome confirmed by phosphomannomutase gene mutations with normal development and absence of biologic anomalies such as elevated transaminases and altered hemostasis. In conclusion, congenital disorders of glycosylation should be considered in cases of unexplained behavioral symptoms such as hyperactivity and concentration difficulties and mild neurologic signs. Intellectual retardation is often overestimated because of dysarthria and motor difficulties. Psychomotor reeducation might improve quality of life. PMID- 15664774 TI - Spinal cord infarction in meningitis: polygenic risk factors. AB - We report a male with spinal cord infarction and tetraplegia after Streptococcus pneumoniae meningitis. He was subsequently found to have both a Chiari I malformation and factor V Leiden mutation. A literature search was conducted to identify previously reported cases of pediatric spinal cord infarction associated with acute bacterial meningitis, anatomic brain anomalies, and hypercoagulability disorders. This article is the first report of spinal cord infarction in a child with hypercoagulability disorder and structural brain anomaly in the setting of acute bacterial meningitis. The confluence of infection, inflammation, localized pressure, and predisposition to hypercoagulability produced unique conditions resulting in infarction of the cervical spine. This report emphasizes the polygenic nature of the expression of spinal cord infarction. PMID- 15664775 TI - Visual disturbance associated with postoperative cerebellar mutism. AB - Cerebellar mutism is an uncommon complication of posterior fossa surgery. Manifestations include disturbances of articulation, prosody, and pitch, and, if severe, complete mutism. Symptoms are independent of recognizable cortical or brainstem injury, and recovery is variable, with permanent deficits frequently observed. Cerebellar dysfunction is commonly invoked as an etiology, although controversy remains concerning the mechanism. Visual impairment has been reported only once before in the setting of this disorder. We report a confirmatory case of sudden, severe visual loss in association with cerebellar mutism after resection of a midline medulloblastoma in a 7-year-old. PMID- 15664776 TI - Migraine-like attacks in child with Sturge-Weber syndrome without facial nevus. AB - The Sturge-Weber syndrome was recently subdivided into type I (facial and leptomeningeal angioma, possible glaucoma), type II (facial angioma, without evident endocranial involvement), and type III (exclusive leptomeningeal angioma). Thus far in the literature only 24 cases of Sturge-Weber syndrome type III have been reported. This study presents a case of a 2-year 9-month-old child with normal psychomotor development and skin free (no angiomas), who presented repeated episodes of severe headache, vertiginous symptoms, vomiting, and drowsiness, separated by complete recovery. The cranial computed tomography and magnetic resonance imaging with gadolinium revealed left occipital leptomeningeal angiomatosis with calcifications, suggesting a diagnosis of Sturge-Weber syndrome type III. Considering the normal psychomotor development, the improved electroencephalographic reports between the episodes, and the absence of hypoperfusion areas on single-photon emission computed tomography at 30 months of follow-up, the symptomatology appears an expression of migraine-like symptoms resulting from vasomotor disturbances within and around the angioma, more than an expression of partial seizures arising through an epileptic focus in the ischemic region around the vascular malformation. PMID- 15664777 TI - An inborn error of metabolism presenting as hypoxic-ischemic insult. AB - This case report profiles two children whose sole presentation is intractable seizures. The index case is a 1-year-old female. She presented to the emergency department with intractable seizures. Her initial metabolic evaluation was nonconclusive. Electroencephalogram was abnormal. Brain magnetic resonance imaging yielded a picture consistent with profound ischemic hypoxic injury. The second case was the 8-year-old brother of the index case. He suffered from intractable seizures since birth. On examination he was microcephalic with spastic quadriparesis and bilateral dislocation of lenses. Computed tomography of the brain revealed a low-density area in the white and cortical matter consistent with hypoxic-ischemic injury. His urinalysis for sulfocysteine produced findings consistent with isolated sulfite oxidase deficiency. PMID- 15664778 TI - Cerebellar hypoperfusion in infantile neuroaxonal dystrophy. AB - An identical abnormal pattern was detected by means of (99m)Tc-hexamethyl propyleneamine-oxime single-photon emission computed tomography in two siblings with infantile neuroaxonal dystrophy. The markedly decreased cerebellar perfusion, along with the early motor symptoms, characteristic magnetic resonance imaging and pathologic findings, points to a preferential cerebellar involvement in this disease. A relative increase in the perfusion to the basal ganglia correlated with the magnetic resonance imaging abnormalities, highly resembling that of Hallervorden-Spatz disease in one of the males, at this site. PMID- 15664779 TI - Subdural hematoma as clinical presentation of osteogenesis imperfecta. AB - Osteogenesis imperfecta is an inherited collagenous disease. The mildest form may present with less severe findings, for example blue sclera, but can also lead to significant handicap such as deafness or multiple bone fractures. We describe an 11-month-old female in whom bilateral chronic subdural hematoma was the leading clinical presentation of osteogenesis imperfecta type I. She was hospitalized due to epileptic seizures caused by these bilateral subdural hematomas without preceding trauma. Osteogenesis imperfecta type I was diagnosed on the basis of clinical and radiologic findings. This case demonstrates that nontraumatic chronic subdural hematoma in patients with osteogenesis imperfecta type I may be caused by impaired bone calcification, vascular fragility, and permanent friction between multiple bone fragments of the skull. Osteogenesis imperfecta type I should be considered as an underlying disease in cases of nontraumatic subdural hematoma. A thorough clinical examination is recommended to exclude subtle characteristics of the disease. PMID- 15664780 TI - Brain glutamine by MRS in a patient with urea cycle disorder and coma. AB - In patients who undergo metabolic decompensation from urea cycle disorders, cerebrospinal fluid glutamine level may be a better marker of cerebral dysfunction than blood ammonia or glutamine levels. However, obtaining cerebrospinal fluid by lumbar puncture carries risk in these acutely ill patients with cerebral edema. Using magnetic resonance single voxel spectroscopy as an alternative to cerebrospinal fluid analysis, elevated brain glutamine/glutamate complex levels were detected in a patient with carbamyl phosphate synthetase deficiency, who had been comatose for many days after normalization of blood ammonia and improvement in blood glutamine levels. Brain glutamine by single voxel spectroscopy decreased toward normal with neurologic recovery. We conclude that brain glutamine may be a better marker than serum ammonia for the management of urea cycle disorders, particularly in patients with prolonged mental status changes. PMID- 15664781 TI - Stress-associated changes in the steady-state expression of latent Epstein-Barr virus: implications for chronic fatigue syndrome and cancer. AB - Antibodies to several Epstein-Barr virus (EBV)-encoded enzymes are observed in patients with different EBV-associated diseases. The reason for these antibody patterns and the role these proteins might play in the pathophysiology of disease, separate from their role in virus replication, is unknown. In this series of studies, we found that purified EBV deoxyuridine triphosphate nucleotidohydrolase (dUTPase) can inhibit the replication of human peripheral blood mononuclear cells in vitro and upregulate the production of TNF-alpha, IL 1beta, IL-6, IL-8, and IL-10. It also enhanced the ability of natural killer cells to lyse target cells. The EBV dUTPase also significantly inhibited the replication of mitogen-stimulated lymphocytes and the synthesis of IFN-gamma by cells isolated from lymph nodes and spleens obtained from mice inoculated with the protein. It also produced sickness behaviors known to be induced by some of the cytokines that were studied in the in vitro experiments. These symptoms include an increase in body temperature, a decrease in body mass and in physical activity. The data provide a new perspective on how an early nonstructural EBV encoded protein can cause immune dysregulation and produce clinical symptoms observed in patients with chronic fatigue syndrome (CFS) separate from its role in virus replication and may serve as a new approach to help identify one of the etiological agents for CFS. The data also provide additional insight into the pathophysiology of EBV infection, inflammation, and cancer. PMID- 15664782 TI - Immune-to-brain communication dynamically modulates pain: physiological and pathological consequences. AB - This review examines recently recognized roles of immunological processes in pain modulation and explores the potential implications of these immunologically derived phenomena for human chronic pain control. The focus is an examination of how activation of immune-like glial cells within the spinal cord can amplify pain by modulating the excitability of spinal neurons. Such glially driven enhancement of pain can be physiological, as occurs in response to peripheral infection or inflammation. Here, immune-to-brain-to-spinal cord communication leads to pain enhancement (hyperalgesia) as one component of the well-characterized sickness response. This sickness-induced hyperalgesia, like many sickness responses, is mediated by the activation of glia and the consequent release of proinflammatory cytokines. However, glially driven pain can also occur under pathological conditions, such as occurs following peripheral nerve inflammation or trauma. Here, immune- and trauma-induced alterations in peripheral nerve function lead to the release of substances within the spinal cord that trigger the activation of glia. Evidence is reviewed that such pathologically driven glial activation is associated with enhanced pain states of diverse etiologies and that such pain facilitation is driven by glial release of proinflammatory cytokines and other neuroexcitatory substances. This recently recognized role of spinal cord glia and glially derived proinflammatory cytokines as powerful modulators of pain is exciting as it may provide novel approaches for controlling human chronic pain states that are poorly controlled by currently available therapies. PMID- 15664783 TI - How important is it to consider prophylaxis to counter non-specific immunosuppression following cancer surgery? PMID- 15664784 TI - Marginating pulmonary-NK activity and resistance to experimental tumor metastasis: suppression by surgery and the prophylactic use of a beta-adrenergic antagonist and a prostaglandin synthesis inhibitor. AB - Surgery is imperative for cancer treatment, but was suggested to suppress immunity and facilitate metastasis. Here we study the involvement of catecholamines and prostaglandins (PG) in such outcomes, and the role played by marginating-pulmonary (MP)-NK cells in controlling MADB106 metastasis. Non operated and laparotomized F344 rats were injected postoperatively with a PG synthesis inhibitor (indomethacin, 4 mg/kg i.p.), a beta-blocker (nadolol, 0.6 mg/kg s.c.), both drugs, or vehicle. Rats were then inoculated intravenously with non-immunogenic syngeneic MADB106 cells, and 24 h later lung tumor retention was assessed, or 3 weeks later lung metastases were counted. Additionally, 12 h after surgery we harvested MP-NK cells and circulating-NK cells and compared their numbers and cytotoxicity against MADB106 cells and standard YAC-1 target cells. Surgery significantly increased MADB106 metastasis. Nadolol and indomethacin reduced this effect by approximately 50% when used alone, and significantly more (75%) when used together. Only MP-leukocytes exhibited NK cytotoxicity against MADB106 cells. Surgery markedly suppressed it, and nadolol and indomethacin additively restored it. Similar effects were observed assessing MP-NK and circulating-NK cytotoxicity against YAC-1 target cells. Alterations in the numbers of NK cells were partly associated with alterations in total MP-NK activity, but not with circulating-NK activity. Last, administrating nai ve rats with physiologically relevant doses of a beta-adrenergic agonist (metaproterenol), and/or with PGE2, additively and independently of each other promoted MADB106 metastasis, simulating the effects of surgery. These findings point at potential prophylactic measures in cancer patients undergoing surgery, and suggest a role for MP-NK cells in resisting metastasis of apparently insensitive tumors. PMID- 15664786 TI - The role of central corticotropin-releasing hormone in the anorexic and endocrine effects of the bacterial T cell superantigen, Staphylococcal enterotoxin A. AB - Bacterial superantigens, such as the staphylococcal enterotoxins, exert a strong capacity for in vivo stimulation of T cell proliferation and cytokine production. Previously, staphylococcal enterotoxin A (SEA) was shown to induce an anorexic effect under novel contextual conditions of testing, and produced an increase in plasma ACTH and corticosterone levels in C57BL/6J mice. In the present study, the role of corticotropin releasing hormone (CRH) in promoting these effects of SEA was addressed via intracerebroventricular (icv) administration of alpha-helical CRH(9-41) ((alpha)hCRH), a non-selective CRH receptor antagonist, and astressin 2B, a selective CRH receptor 2 antagonist. The efficacy of (alpha)hCRH and astressin-2B in blocking anorexic responses to CRH and urocortin under the current conditions of testing was first confirmed. Subsequently, it was found that (alpha)hCRH (20 microg icv), but not astressin-2B (10 and 25 microg icv), significantly attenuated the anorexia induced by SEA. This suggested that central CRH is involved in mediating the anorexia induced by SEA, but potentially through CRH receptor 1. Additional results revealed that plasma ACTH stimulation in response to SEA was not significantly attenuated by either antagonist administered icv. However, the plasma corticosterone elevation showed a modest, but significant, attenuation in SEA challenged mice given (alpha)hCRH. These data suggest a possible influence of central CRH on adrenocorticoid activity subsequent to SEA challenge. More importantly, it appears that central activation of CRH receptors is a consequence of SEA challenge, and this likely contributes to its anorexic effects. PMID- 15664785 TI - IL-1 type I receptor plays a key role in mediating the recruitment of leukocytes into the central nervous system. AB - This study investigates the role of type I IL-1 receptor (IL-1R1) in mediating the recruitment of leukocytes into the brain parenchyma in mice. Intracerebroventricular (icv) injection of interleukin IL-1beta induced infiltration of leukocytes between 8 and 72 h after the injection. Leukocytes were rarely found in the brain tissue of saline-injected animals. At 8h after IL 1beta injection, leukocytes were seen lining the blood vessels of the brain and sparsely scattered infiltration of leukocytes was found in the cortex. Peak infiltration of leukocytes, which distributed evenly throughout the brain, was seen at 16 h post-injection. The number of leukocytes in the brain declined thereafter and no leukocytes were found 72 h post-injection. This phenomenon was replicated in mice deficient in lymphotoxin-alpha (LT(alpha)), IL-6, interferon (IFN)-gamma receptor, or the tumor necrosis factor (TNF)-alpha receptor, but abrogated in animals deficient in IL-1R1. ICV injection of IFN-gamma or TNF alpha, but not IL-6 or IL-12, also induced leukocyte infiltration into the brain. Injection of IL-1beta, IFN-gamma, TNF-alpha, IL-6, and IL-12 induced IL-1beta expression in the brain, with IL-6 and IL-12 being the least effective. Leukocyte infiltration induced by icv IFN-gamma and TNF-alpha was also abrogated in IL-1R1 knockout animals. The induced infiltrating leukocytes were identified as neutrophils. Chronic infection with Trypanosoma brucei resulted in the recruitment of T cells, but no other cell types, into the brain. This did not occur in IL-1R1-knockout mice. Thus, IL-1R1 appears to be important for the recruitment of leukocytes across the blood-brain barrier. PMID- 15664787 TI - Does genotype mask the relationship between psychological factors and immune function? AB - This paper examined the interaction between genetic influences of the polymorphic human leukocyte antigens (DRB1 and DQB1) and psychological distress on the development of cellular immunity to the novel antigen, keyhole limpet hemocyanin (KLH). Participants (n = 227) were immunized with KLH and the development of cutaneous delayed-type hypersensitivity (DTH) against KLH was examined 3 weeks later. Distress was assessed using the Profile of Mood States. DNA was typed for the serologically defined DRB1 and DQB1 antigens. There was a significant correlation between distress at immunization and the development of DTH skin test responses to KLH (n = 214, r = .24, p = .003). HLA DQ2 was weakly associated with a decreased likelihood of developing a cutaneous delayed-type hypersensitivity response against KLH (odds ratio [OR] = 1.6; confidence interval [CI] 0.9-2.7). HLA DQ5 was weakly associated with an increased likelihood of responding to the antigen (OR=0.6; CI=0.3-1.0). The correlation between distress and immune function in HLA DQ2 negative individuals was .34 (n = 136, p = .00) and in HLA DQ2 positive individuals it was .06 (n = 74, p =. 64). For HLA DQ5 negative individuals the correlation was .26 (n = 140, p = .00) and for HLA DQ5 positive individuals it was .22 (n = 70, p = .07). These results suggest that the distress/immune relationship in genetically susceptible or protected individuals may be underestimated in psychoneuroimmunology research. PMID- 15664788 TI - Stress-induced modulation of NK activity during influenza viral infection: role of glucocorticoids and opioids. AB - Activation of the hypothalamic-pituitary-adrenal axis (HPA) and sympathetic nervous system by stress has been shown to modulate both innate and adaptive immunity during an experimental influenza A/PR8 viral infection. HPA activation alters levels of glucocorticoids (GC) and opioids which are associated with suppression of lymphoid cellularity and NK activity. These experiments were designed to investigate the role that stress-induced GC and opioids have in modulating NK activity during an influenza viral infection. C57BL/6 mice were treated daily with mifepristone (RU486), a GC receptor antagonist or naltrexone (NTX), a non-specific opioid receptor antagonist. Mice were infected intranasally with A/PR8 virus and underwent daily restraint stress (RST). Phenotypic analysis of splenic cell populations and NK cytotoxicity were assessed at 3 days post infection. RST of infected mice significantly suppressed splenic CD3(-)DX5+ cellularity and NK cytolytic activity. RU486 administration fully restored splenic NK cellularity but not cytolytic activity. NTX administration restored NK cytolytic activity but not splenic NK cell number. A similar restoration in NK cytolytic activity was observed after administration of beta-funaltrexamine (FNA), a mu-specific opioid receptor antagonist, but not the delta- or kappa specific opioid receptor antagonists naltrindole or nor-binaltorphimine, respectively. Co-administration of RU486 and NTX restored both NK cellularity and cytolytic activity. The restoration of RST-induced suppression of NK activity by RU486 and NTX or FNA suggests that glucocorticoids modulate NK cellularity and opioids that bind to the mu-opioid receptor modulate NK cytotoxicity during periods of stress and viral infection. PMID- 15664789 TI - The association between interleukin-6, sleep, and demographic characteristics. AB - We examined the relationship between the pro-inflammatory cytokine IL-6 and sleep architecture in 70 healthy men and women. Blood was drawn in the early morning for assessment of IL-6 followed by nocturnal sleep monitoring with polysomnography. Sleep records were scored for sleep stages using standard criteria. Morning IL-6 levels were positively correlated with REM latency after sleep onset [rho = .31, p = .01], percent (%) stage 1 sleep [rho = .23, p = .053], % wake after sleep onset (WASO) [rho = .29, p<.05]. IL-6 levels were negatively correlated with sleep efficiency [rho = -.36, p<.01] and slow wave sleep (SWS) [rho = -.26, p<.05]. After controlling for demographic variables including race, gender, age, and BMI, multiple hierarchical regression analyses revealed that morning IL-6 levels accounted for a significant portion of the variance of REM latency (p<.01), sleep efficiency (p<.01), and % WASO (p = .01). IL-6 was no longer associated with % stage 1 sleep, SWS, and total sleep time after controlling for the demographic characteristics. These findings suggest that the inflammatory marker IL-6 is associated with sleep quality and that certain individual characteristics such as race, gender, and age modify that relationship. Higher IL-6 levels were associated with lower quality of sleep among healthy asymptomatic men and women. PMID- 15664790 TI - Brain-derived neurotrophic factor supports facial motoneuron survival after facial nerve transection in immunodeficient mice. AB - Numerous studies have shown that motoneuron survival can be facilitated by neurotrophic factors (NTF) after injury. However, the ability of specific NTF to rescue facial motoneurons (FMN) from axotomy-induced death in immunodeficient mice has not been tested. Therefore, one goal of this study was to determine if brain-derived neurotrophic factor (BDNF), an NTF with a known ability to rescue FMN from axotomy-induced death, supports FMN from axotomy-induced death in recombinase activating gene-2 knockout (RAG-2 KO) mice that lack functional T and B lymphocytes. Nerve growth factor, which has been shown not to play a role in motoneuron survival, was used as a negative control. Brain derived neurotrophic factor treatment restored FMN survival to wild-type (WT) control levels 4 weeks post-operative (wpo) (80% +/- 1.9, 83% +/- 2.4, respectively). The second goal of this study was to begin to elucidate if CD4+ T cells produce NTF after facial nerve axotomy. Cervical lymph nodes were collected from WT mice 9 days post operative, re-activated with anti-CD3 and supernatant collected 24 h later. Immediately after injury, the supernatant was administered to RAG-2 KO mice leading to an increase in FMN survival equivalent to WT controls (80% +/- 1.4, 84% +/- 2.1, respectively, 4 wpo). In addition, cervical lymph node supernatant treated with anti-BDNF attenuated FMN rescue in RAG-2 KO mice (62% +/- 3.3) 4 wpo. These data lend support to the hypothesis that CD4+ T cells produce NTF that support motoneuron survival before target reconnection occurs. PMID- 15664791 TI - Testosterone administration attenuates regional brain hypometabolism in women with anorexia nervosa. AB - Abnormalities in brain metabolism have not been consistently well localized in anorexia nervosa (AN), and effects of specific therapies on these functional abnormalities have not been studied. Androgen replacement therapy improves mood, well-being and cognitive function in men with androgen deficiency. We therefore hypothesized that women with AN and relative androgen deficiency would exhibit regional brain hypometabolism compared with healthy controls, and that low-dose physiologic androgen replacement would attenuate the hypometabolism in some of these brain loci. We used FDG PET and statistical parametric mapping methods to investigate regional brain glucose metabolism in (1) 14 women with AN and 20 healthy control subjects of similar mean age and (2) women with AN after randomization to low-dose replacement testosterone therapy or placebo. Cerebral metabolism was decreased in the posterior cingulate, pregenual anterior cingulate, left middle temporal, right superior temporal, and left dorsolateral prefrontal cortex in the AN group compared with controls. In AN patients receiving testosterone, cerebral metabolism increased in the posterior cingulate, subgenual anterior cingulate, premotor cortex, right caudate and right parietal lobes. In conclusion, our data demonstrate distinct loci of regional brain hypometabolism in women with AN compared with controls. Moreover, abnormalities in one of these regions-the posterior cingulate cortex-were attenuated towards normal with low-dose testosterone replacement therapy. Further study is warranted to replicate these findings, as well as to determine their physiological and clinical significance. PMID- 15664792 TI - Magnetic resonance detects brainstem changes in chronic, active heavy drinkers. AB - Neuropathological and neuroimaging studies show cortical and subcortical volume loss in alcohol-dependent individuals. Using quantitative magnetic resonance imaging (MRI) and proton magnetic resonance spectroscopic imaging ((1)H MRSI), we studied the size and potential cellular injury of the brainstem in untreated heavy alcohol drinkers. The brainstem is considered critical in the development and maintenance of drug and alcohol dependence. Two methods of brainstem size determination were compared: standard volumetry vs. midsagittal MR image area measurement. Heavy drinkers (n=12) and light drinkers (n=10) were compared with MRI; (1)H MRSI brainstem data were obtained from a subset of this cohort. Chronic heavy drinking was associated with significantly smaller midsagittal areas of the brainstem, midbrain, and pons, and with significantly smaller overall brainstem volume. Heavy drinking was also associated with significantly lower ratios of N acetyl-aspartate and choline-containing metabolites compared with creatine containing compounds in the brainstem, independent of brainstem atrophy. Additionally, brainstem volume and midsagittal brainstem area were correlated (r=0.78). These structural and metabolite findings are consistent with neuronal injury in the brainstem of untreated chronic heavy drinkers. The results also indicate that the midsagittal MRI brainstem area is an easily determined and reliable indicator of brainstem volume. PMID- 15664793 TI - Glucose utilization in the medial prefrontal cortex correlates with serotonin turnover rate and clinical depression in alcoholics. AB - We measured the serotonin metabolite 5-hydroxyindoleacetic acid (5-HIAA) in cerebrospinal fluid (CSF), regional cerebral glucose uptake (rCMRglc) as assessed with positron emission tomography in the medial prefrontal cortex (PFC) and severity of clinical depression (Beck's Depression Inventory, BDI) in detoxified male alcoholics and age-matched healthy men. In alcoholics, the severity of clinical depression was negatively correlated with rCMRglc in the medial PFC and positively with CSF 5-HIAA concentrations. A voxel-based analysis showed that the strongest correlation between CSF 5-HIAA levels and rCMRglc was found in alcoholics in the left orbitofrontal and medial PFC (BA10 and BA11); no significant correlations were observed among healthy control subjects. This pilot study indicates that a dysfunction of medial PFC may interact with central serotonin turnover and negative mood states during early abstinence. PMID- 15664794 TI - Amygdala activity in obsessive-compulsive disorder with contamination fear: a study with oxygen-15 water positron emission tomography. AB - Previous imaging studies of obsessive-compulsive symptom states have implicated frontal-striatal and limbic regions in the pathophysiology of obsessive compulsive disorder (OCD). Functional imaging studies, however, have yielded inconsistent results, presumably due to methodological differences (patient inclusion criteria, stimulus paradigm, imaging technique, and absence of control groups). In the present study, randomized presentation of contamination-related and neutral visual stimuli was used to investigate the neurophysiological correlates of contamination fear in a group of medication-free OCD patients with washing behaviors and healthy controls. A total of 21 subjects (11 OCD patients and 10 healthy controls) were scanned using H(2)(15)O positron emission tomography (PET). Subjects were presented with pictures of clean and dirty surroundings and were requested to make indoor/outdoor decisions to control for attention differences. State anxiety and obsessionality were rated after each scan using visual analogue scales. Main effects of stimulus type (contamination vs. neutral) were found in bilateral occipital cortex in both groups. A significant group interaction effect was observed in the left amygdala reflecting enhanced activity in response to contamination stimuli in OCD patients. Sensitization effects were observed in the right amygdala in the OCD group; these paralleled an increase in levels of distress and obsessionality as well as a decrease in dorsolateral prefrontal activity. The findings of the present study are consistent with the hypothesis of decreased frontal-striatal control of limbic structures, specifically the amygdala, resulting in an inadequate fear response in OCD patients with contamination fear. PMID- 15664796 TI - Gray matter abnormalities in paranoid schizophrenia and their clinical correlations. AB - Numerous imaging studies have shown structural brain abnormalities in schizophrenia. Recently, voxel-based morphometry enabled whole brain analysis beyond the regions of interest (ROI). Regional gray matter concentrations of magnetic resonance (MR) images from 35 patients with paranoid schizophrenia were compared with those from 35 age- and sex-matched controls, and their clinical correlations were explored using voxel-based morphometry. Gray matter concentrations in the patients were significantly reduced in the left insular and dorsolateral prefrontal regions, and bilaterally in the medial frontal, anterior cingulate, inferior frontal and superior temporal regions. On the other hand, gray matter concentrations in the bilateral cerebellum and right striatum were significantly increased in the patients compared with controls. A negative correlation between the score for the severity of 'lack of insight and judgment' and gray matter concentrations in the left posterior and right anterior cingulate and bilateral inferior temporal regions including the lateral fusiform gyri was found. These results suggest the important roles of the paralimbic structures in the pathophysiology of schizophrenia and the involvement of the perceptual and monitoring systems in the mechanism of insight. PMID- 15664795 TI - Functional MRI changes before and after onset of reported emotions. AB - The social nature of emotion is evident in the importance of facial and vocal displays in emotion-related behavior. This is the first brain-imaging study to use simulated face-to-face social interactions to evoke emotional responses and to compare valence-related activations before and after subjective onset of emotional response. Videotapes were prepared with actresses who described happy or unhappy experiences. Functional magnetic resonance imaging (fMRI) at 1.5 T was used to acquire BOLD images in 21 healthy young adults before, after, and during viewing of the happy and sad tapes. Subjects pushed buttons to indicate the onset of subjective emotional responses. Group data were analyzed by a bootstrap randomization method after anatomical normalization. Significant activation was detected in frontal and sensory regions prior to the reported onset of emotional response, and this activity showed a marked decrease after the report of conscious emotional experience. Significant differences between happy and sad conditions were evident in multiple brain regions both before and after the reported onset of emotional response, including the middle and superior temporal gyri, the middle frontal gyrus, the caudate, and the hippocampus. Socially relevant emotional stimulation is feasible and evokes robust responses. The neural correlates of the evoked emotion are multiple, widely distributed, and inclusive of areas important in many cognitive tasks. Positive and negative emotional responses include activation of common and distinctive brain regions. PMID- 15664797 TI - Shape changes in prefrontal, but not parieto-occipital regions: brains of schizophrenic patients come closer to a circle in coronal and sagittal view. AB - There is some evidence for prefrontal (PF) lobe changes in schizophrenia while the parieto-occipital (PO) region seems to be unaffected. This magnetic resonance imaging (MRI) study was performed to examine shape differences as part of the spectrum of structural abnormalities in schizophrenia. The measurements were done on families affected with schizophrenia to identify the influence of genetic and environmental factors on these changes. The sample under study consisted of 164 subjects including 45 family members (FM) suffering from schizophrenia, 27 FM with other psychiatric disorders and 51 FM without psychiatric disorders based on ICD-10 criteria. In addition, 41 nonpsychiatric control subjects were included in the study. On defined planes at the corpus callosum boundary of the PF and the PO, brain width, height and length were measured on coronal slices. Ratios of these linear measurements were also calculated based on the idea that a plane can be approximated by a circle if the concerning ratio comes close to 1. It was hypothesized that these relative brain shape parameters, especially the PF ratio width/height, would show differences between schizophrenic patients and control subjects. For all members from families with schizophrenia compared with control subjects, there were significant differences in the PF, but not in the PO region. PF height was increased. PF ratios of width/height and height/(2 x length) were closer to 1 in affected families than in control subjects. The results can be interpreted as an indication for PF brain shape changes in subjects with a disposition for schizophrenia. On coronal and sagittal planes situated at the corpus callosum, their PF could be approximated by a circle better than in control subjects. As the frontal lobe takes shape late in brain development, underlying genetic mechanisms may be dysregulated in schizophrenic patients and subjects at risk to develop the disorder. PMID- 15664798 TI - Altered functioning of the cingulate gyrus in two cases of chromosome 22q11 deletion syndrome. AB - The 22q11 microdeletion syndrome (22q11-DS) is strongly associated with schizophreniform disorders and, in turn, the 22q11 deletion region harbours several candidate genes for schizophrenia. Here, we present the case of an adolescent female patient with 22q11-DS associated with impaired cognitive abilities and behavioural abnormalities. The patient was studied with magnetic resonance imaging (MRI) and positron emission tomography (PET) as well as extensive neurophysiological investigations. Although no structural or functional abnormalities were found in MRI and PET, assessment of event-related potentials elicited during the Continuous Performance Test revealed a lack of NoGo anteriorisation. The latter was replicated in a second case of 22q11-DS with schizoaffective disorder but devoid of a severe somatic syndrome. This electrophysiological finding, which indicates modified functioning of the cingulate gyrus, has previously been demonstrated only in patients suffering from schizophrenia and attention deficit/hyperactivity disorder, two psychopathological conditions frequently associated with 22q11-DS. We conclude that more extensive study of NoGo anteriorisation as a potential endophenotype of 22q11-DS patients at risk for 22q11-associated psychiatric conditions is warranted. PMID- 15664799 TI - Acetazolamide-enhanced neuroSPECT scan reveals functional impairment after minimal traumatic brain injury not otherwise discernible. AB - A patient suffering from aggressive personality changes and cognitive impairment following head trauma, without neurological or anatomical imaging findings, underwent neuroSPECT scans with and without acetazolamide injection, both before treatment and during treatment with valproate. Acetazolamide injection induced increased prefrontal perfusion not evident at baseline. Valproate treatment was associated with increased prefrontal perfusion concomitant with clinical improvement and abolished response to acetazolamide challenge. PMID- 15664800 TI - Clinical value of 99mTc-HMPAO SPECT in depressed bipolar I patients. AB - Regional cerebral blood flow was studied in 17 bipolar I depressed patients (DSM IV criteria) with single photon emission computed tomography (SPECT). Visual analysis of images revealed no abnormality in eight patients and abnormal findings in nine patients. In the nine patients with abnormal findings, all showed regional decreases of the uptake of (99m)Tc-D,L-hexamethylpropylene amine oxime (HMPAO, four in the frontal region, two in the basal ganglia, and three in both the frontal region and the basal ganglia). The patients with visible SPECT abnormalities had significantly higher scores on the Hamilton Rating Scale for Depression (HDRS). PMID- 15664801 TI - Enantiomerical excess determination, purification and biological evaluation of (3S) and (3R) alpha,beta-butenolide analogues of isobenzofuranone. AB - The asymmetric synthesis of isobenzofurane analogues, new potential antiviral agents, is reported. High performance liquid chromatography (HPLC) was the technique chosen to separate the enantiomers. We describe this chiral separation and then determine the enantiomerical excess. The biological results of each tested enantiomer are given. PMID- 15664802 TI - Hybridization dependent cleavage of internally modified disulfide-peptide nucleic acids. AB - Selectivity of the cleavage of single stranded over hybridized forms of internally modified disulfide-peptide nucleic acids (PNA) has been optimized using a series of phosphines and thiols, which have different sizes and charges. For the most selective cleaver found (tris-(carboxyethyl)-phosphine), reactivity of single stranded PNA is 33 times higher than that of the PNA-DNA duplex. Selectivity of single stranded disulfide-PNA cleavage has been explained in terms of electrostatic interaction between the substrate and the cleaver. PMID- 15664803 TI - Synthesis of alkyne derivatives of a novel triazolopyrazine as A(2A) adenosine receptor antagonists. AB - A novel [1,2,4]triazolo[1,5-a]pyrazine core was synthesized and coupled with terminal acetylenes. The structure-activity relationship of the alkynes from this novel template was studied for their in vitro and in vivo adenosine A(2A) receptor antagonism. Selected compounds from this series were shown to have potent in vitro and in vivo activities against adenosine A(2A) receptor. Compound 12, in particular, was found to be orally active at 3mg/kg in both a mouse catalepsy model and a 6-hydroxydopamine-lesioned rat model. PMID- 15664804 TI - Identification of putative metabolites of docosahexaenoic acid as potent PPARgamma agonists and antidiabetic agents. AB - We found that putative metabolites of docosahexaenoic acid (DHA) are strong PPARgamma activators and potential antidiabetic agents. We designed DHA derivatives based on the crystal structure of PPARgamma, synthesized them and evaluated their activities in vitro and in vivo. The efficacy of 5E-4-hydroxy-DHA 2a as a PPARgamma activator was about fourfold stronger than that of pioglitazone. Furthermore, the 4-keto derivative (10b) showed antidiabetic activity in animal models without producing undesirable effects such as obesity and hepatotoxicity. PMID- 15664805 TI - Synthesis and acetylcholinesterase inhibition of derivatives of huperzine B. AB - By targeting dual active sites of AChE, a number of new derivatives of HupB have been synthesized and tested as acetylcholinesterase inhibitors. The most potent compound, bis-HupB 5b is 72-fold more potent in AChE inhibition and 79-fold more selective for AChE versus BChE than HupB. PMID- 15664806 TI - Structure-activity relationship studies on ortho-substituted cinnamic acids, a new class of selective EP(3) antagonists. AB - A series of novel ortho-substituted cinnamic acids have been synthesized, and their binding activity and selectivity on the four prostaglandin E(2) receptors evaluated. Many of them are very potent and selective EP(3) antagonists (K(i) 3 10 nM), while compound 9 is a very good and selective EP(2) agonist (K(i) 8 nM). The biological profile of the EP(2) agonist 9 in vivo and the metabolic profile of selected EP(3) antagonists are also reported. PMID- 15664807 TI - Syntheses of 2,4,6-trisubstituted triazines as antimalarial agents. AB - A series of 2,4,6-trisubstituted-1,3,5-triazines (2a-s) were synthesized and evaluated for their in vitro antimalarial activity against P. falciparum. Out of the 19 compounds synthesized eight compounds showed MIC in the range of 1-2 microg/mL. These compounds are in vitro several times more active than cycloguanil. PMID- 15664808 TI - N-Alkylation of phenethylamine and tryptamine. AB - A clean and efficient method for the N-alkylation of tryptamine and phenethylamine, employing alcohols as the alkylating agents, has been developed. The reaction proceeds via catalytic electronic activation, involving an iridium catalyst which activates the alcohol by borrowing hydrogen from the substrate, returning it later in the catalytic cycle. Some examples of N-heterocyclisation have been performed employing a diol as the substrate. PMID- 15664809 TI - Synthesis and biological evaluation of 3-benzyl-1-methyl- and 1-methyl-3-phenyl isothioureas as potential inhibitors of iNOS. AB - Novel benzyl- and phenyl-isothioureidic derivatives have been synthesised and evaluated as inhibitors of nitric oxide synthesis, induced in lipopolysaccharide (LPS)-activated J774.A1 macrophage cell line. The most potent iNOS inhibitor resulting was 1-methyl-3-phenyl-S-methyl isothiourea 5l. PMID- 15664810 TI - New conformationally locked bicyclic N,O-nucleoside analogues of antiviral drugs. AB - In order to obtain rigidity within the sugar moiety of nucleosides, the bicyclic pyrimidine derivatives of N,O-isoxazolidines were designed and synthesized by using 1,3-dipolar cycloaddition of Delta(1)-pyrrolidine-1-oxide and the appropriate vinyl-nucleobases. PMID- 15664811 TI - Unified Markov thermodynamics based on stochastic forms to classify drugs considering molecular structure, partition system, and biological species: distribution of the antimicrobial G1 on rat tissues. AB - To date, molecular descriptors do not commonly account for important information beyond chemical structure. The present work, attempts to extend, in this sense, the stochastic molecular descriptors, incorporating information about the specific biphasic partition system, the biological species, and chemical structure inside the molecular descriptors. Consequently, MARCH-INSIDE molecular descriptors may be identified with time-dependent thermodynamic parameters (entropy and mean free energy) of partition process. A classification function was developed to classify data of 423 drugs and up to 14 different partition systems at the same time. The model has shown a high overall accuracy of 92.1% (293 out of 318 cases) in training series and 90% (36 out of 40 cases) in predicting ones. Finally, we illustrate the use of the model by predicting a high probability (%) for G1 (a novel antibacterial drug) to undergo partition on different biotic systems (rat organs): liver (97.7), spleen (97.5), lung (97.4), and adipose tissue (97.6). These theoretical results coincide with herein reported steady state plasma concentrations (c) and partition coefficients (P) in liver (c=42.25+/-7.86/P=4.75), spleen (11.47+/-4.43/P=1.29), lung (17.04+/ 3.58/P=1.91), and adipose tissue (28.19+/-11.82/P=3.17). All values were relative to (14)C-labeled-radioactive-G1 in plasma (c=8.9+/-3.05) after 3h of oral administration. In closing, the present stochastic forms derive average thermodynamic parameters fitting on a more clearly physicochemical framework with respect to classic vector-matrix-vector forms, which include, as particular cases, quadratic forms such as Wiener index, Randic invariants, Zagreb descriptors, Harary index, Balaban index, and Marrero-Ponce quadratic molecular indices. PMID- 15664812 TI - Synthesis of modified proanthocyanidins: introduction of acyl substituents at C-8 of catechin. Selective synthesis of a C-4-->O-->C-3 ether-linked procyanidin-like dimer. AB - The regioselective introduction of substituents at C-8 of (+)-catechin is described, leading to the synthesis of several catechin derivatives with various substitution patterns to be used for the further synthesis of modified proanthocyanidins. Thereafter, a new 3-O-4 ether-linked procyanidin-like derivative was synthesized. Its formation was selectively achieved through TiCl(4)-catalyzed condensation of 4-(2-hydroxyethoxy)tetra-O-benzyl catechin with the 8-trifluoroacetyl adduct of tetra-O-benzyl catechin. PMID- 15664813 TI - Synthesis of modified proanthocyanidins: easy and general introduction of a hydroxy group at C-6 of catechin; efficient synthesis of elephantorrhizol. AB - A general procedure for the oxidation of catechin derivatives is described, leading to the introduction of a new hydroxy group at C-6. This procedure has been applied for the synthesis of elephantorrhizol, a natural flavan-3-ol exhibiting a fully substituted cycle A. PMID- 15664814 TI - Carbonic anhydrase inhibitors. Inhibition of isozymes I, II, IV, V, and IX with anions isosteric and isoelectronic with sulfate, nitrate, and carbonate. AB - The inhibition of five human carbonic anhydrase (hCA, EC 4.2.1.1) isozymes; the cytosolic hCA I and II, the membrane-bound hCA IV, the mitochondrial hCA V, and the tumor-associated, transmembrane hCA IX, with anions isosteric and isoelectronic with sulfate, nitrate, and carbonate; such as chlorate, perchlorate, bromate, iodate, periodate, silicate, bismuthate, vanadate, molybdate, and wolframate is reported. Apparently, the geometry of the inhibitor (tetrahedral or trigonal) does not influence its binding to the Zn(II) ion of the enzyme active site, but the nature of the central element is the most important factor influencing potency. Isozymes hCA I and II are best inhibited by chlorate, perchlorate, and silicate, together with the anions structurally related to sulfate, sulfamate, and sulfamidate, but sulfate itself is a weak inhibitor (inhibition constant of 74 mM against hCA I and 183 mM against hCA II). Molybdate is a very weak hCA I inhibitor (K(I) of 914 mM) but it interacts with hCA II (K(I) of 27.5mM). Isozyme IV is well inhibited by sulfate (K(I) of 9 mM), sulfamate, and sulfamidate (in the low micromolar range), but not by perchlorate (K(I) of 767 mM). The mitochondrial isozyme V has the lowest affinity for sulfate (K(I) of 680 mM) and carbonate (K(I) of 95 mM) among all the investigated isozymes, suggesting on one hand its possible participation in metabolon(s) with sulfate anion exchanger(s), and on the other hand an evolutionary adaptation to working at higher pH values (around 8.5 in mitochondria) where rather high amounts of carbonate in equilibrium with bicarbonate may be present. Metasilicate, isosteric to carbonate, is also about a 10 times weaker inhibitor of this isozyme as compared to other CAs investigated here (K(I) of 28.2 mM). Surprisingly, the tumor-associated isozyme IX is resistant to sulfate inhibition (K(I) of 154 mM) but has affinity in the low micromolar range for carbonate, sulfamate, and sulfamidate (K(I) in the range of 8.6-9.6 microM). This constitutes another proof that this isozyme best works at acidic pH values present in tumors, being inhibited substantially at higher pH values when more carbonate may be present. Bromate and chlorate are quite weak CA IX inhibitors (K(I) s of 147-274 mM). PMID- 15664815 TI - Carbonic anhydrase inhibitors. Interaction of isozymes I, II, IV, V, and IX with carboxylates. AB - A detailed inhibition study of five carbonic anhydrase (CA, EC 4.2.1.1) isozymes with carboxylates including aliphatic (formate, acetate), dicarboxylic (oxalate, malonate), hydroxy/keto acids (l-lactate, l-malate, pyruvate), tricarboxylic (citrate), or aromatic (benzoate, tetrafluorobenzoate) representatives, some of which are important intermediates in the Krebs cycle, is presented. The cytosolic isozyme hCA I was strongly activated by acetate, oxalate, pyruvate, l-lactate, and citrate (K(A) around 0.1 microM), whereas formate, malonate, malate, and benzoate were weaker activators (K(A) in the range 0.1-1mM). The cytosolic isozyme hCA II was weakly inhibited by all the investigated anions, with inhibition constants in the range of 0.03-24 mM. The membrane-associated isozyme hCA IV was the most sensitive to inhibition by carboxylates, showing a K(I) of 99 nM for citrate and oxalate, of 2.8 microM for malonate and of 14.5 microM for pyruvate among others. The mitochondrial isozyme hCA V was weakly inhibited by all these carboxylates (K(I)s in the range of 1.67-25.9 mM), with the best inhibitor being citrate (K(I) of 1.67 mM), whereas this is the most resistant CA isozyme to pyruvate inhibition (K(I) of 5.5mM), which may be another proof that CA V is the isozyme involved in the transfer of acetyl groups from the mitochondrion to the cytosol for the provision of substrate(s) for de novo lipogenesis. Furthermore, the relative resistance of CA V to inhibition by pyruvate may be an evolutionary adaptation of this mitochondrial isozyme to the presence of high concentrations of this anion within this organelle. The transmembrane, tumor-associated isozyme hCA IX was similar to isozyme II in its slight inhibition by all these anions (K(I) in the range of 1.12-7.42 mM), except acetate, lactate, and benzoate, which showed a K(I)>150 mM. The lactate insensitivity of CA IX also represents an interesting finding, since it is presumed that this isozyme evolved in such a way as to show a high catalytic activity in hypoxic tumors rich in lactate, and suggests a possible metabolon in which CA IX participates together with the monocarboxylate/H(+) co-transporter in dealing with the high amounts of lactate/H(+) present in tumors. PMID- 15664816 TI - Carbonic anhydrase inhibitors: synthesis and inhibition of cytosolic/tumor associated carbonic anhydrase isozymes I, II, and IX with bis-sulfamates. AB - A series of bis-sulfamates incorporating aliphatic, aromatic, or betulinyl moieties in their molecules was obtained by reaction of the corresponding diols/diphenols with sulfamoyl chloride. The library of bis-sulfamates thus obtained was tested for the inhibition of three physiologically relevant human carbonic anhydrase (hCA, EC 4.2.1.1) isozymes, the cytosolic hCA I and II, and the transmembrane, tumor-associated hCA IX. The new compounds reported here inhibited hCA I with K(I) s in the range of 79 nM-16.45 microM, hCA II with K(I) s in the range of 6-643 nM, and hCA IX with K(I) s in the range of 4-5400 nM. Several low nanomolar hCA IX inhibitors were detected, such as 1,8-octylene-bis sulfamate or 1,10-decylene-bis-sulfamate (K(I) s in the range of 4-7 nM), which showed good selectivity ratios (in the range of 3.50-3.85) for hCA IX over hCA II inhibition. The most selective hCA IX inhibitor was phenyl-1,4-dimethylene-bis sulfamate (K(I) of 61.6 nM), which was a 10.43 times better hCA IX than hCA II inhibitor. These derivatives are interesting candidates for the development of novel antitumor therapies targeting hypoxic tumors, since hCA IX is highly overexpressed in such tissues, and its presence is correlated with bad prognosis and unfavorable clinical outcome. PMID- 15664817 TI - Generation of a new class of hNK(2) receptor ligands using the 'fragment approach'. AB - The so called 'fragment approach' was applied in the search for new leads as selective hNK(2) antagonists. A first round of structural space exploration through the use of bond rigidity as scaffold to support the fragments, afforded 27a as 200 nM hNK(2) ligand. Further refinement gave MEN 14933 as a 16 nM hNK(2) ligand, selective versus hNK(1), of a novel class. Conformational analysis was used to study results and plan future work. PMID- 15664818 TI - Synthesis and SAR studies of 3-phenoxypropyl piperidine analogues as ORL1 (NOP) receptor agonists. AB - A series of 3-phenoxypropyl piperidine analogues have been discovered as novel ORL1 receptor agonists. Structure-activity relationships have been explored around the 3-phenoxypropyl region with several potent and selective analogues identified. PMID- 15664819 TI - Novel spiroanellated 1,2,4-trioxanes with high in vitro antimalarial activities. AB - A remarkable increase in antimalarial in vitro activity was achieved by integration of spiroadamantane motifs in 6-alkylidene 1,2,4-trioxanes 3a-h via diastereoselective photooxygenation of allylic alcohols and subsequent BF(3) catalyzed peroxyacetalization with adamantanone to give the active compounds 3e h. PMID- 15664820 TI - Studies on the structure-activity relationship of 2',6'-dimethyl-l-tyrosine (Dmt) derivatives: bioactivity profile of H-Dmt-NH-CH(3). AB - The 2',6'-dimethyl-l-tyrosine (Dmt) enhances receptor affinity, functional bioactivity and in vivo analgesia of opioid peptides. To further investigate its direct influence on these opioid parameters, we developed a series of compounds (H-Dmt-NH-X). Among them, H-Dmt-NH-CH(3) showed the highest affinity (K(i)mu=7.45 nM) equal to that of morphine, partial mu-opioid agonism (E(max)=66.6%) in vitro and a moderate antinociception in mice. PMID- 15664821 TI - Synthesis and in vitro selective anti-Helicobacter pylori activity of pyrazoline derivatives. AB - In order to develop new anti-Helicobacter pylori agents, a series of N1 substituted 3,5-diphenyl pyrazolines P1-P13 was prepared and evaluated for their antibacterial activity. All synthesized compounds showed little or no activity against different species of Gram-positive and Gram-negative bacteria of clinical relevance and against various strains of pathogenic fungi. The same derivatives exhibited a significant degree of activity against a range of H. pylori strains, including those resistant to the reference compound metronidazole. Among the prepared compounds those with an N1-acetyl group and a 4-methoxy substituent in the 5-phenyl ring showed the best activity against H. pylori metronidazole resistant strains in the 1-4 microg/mL MIC range. PMID- 15664822 TI - The discovery of a selective, high affinity A(2B) adenosine receptor antagonist for the potential treatment of asthma. AB - Adenosine has been suggested to play a role in asthma, possibly via activation of A(2B) adenosine receptors on mast cells and other pulmonary cells. We describe our initial efforts to discover a xanthine based selective A(2B) AdoR antagonist that resulted in the discovery of CVT-5440, a high affinity A(2B) AdoR antagonist with good selectivity (A(2B) AdoR K(i)=50 nM, selectivity A(1)>200: A(2A)>200: A(3)>167). PMID- 15664823 TI - A new facile chemoenzymatic synthesis of levamisole. AB - An efficient and facile chemoenzymatic synthesis of levamisole by employing lipase-mediated resolution of 3-hydroxy-3-phenylpropanenitrile followed by its conversion to beta-amino alcohol as the key intermediate is described. PMID- 15664824 TI - Acyl sulfonamide anti-proliferatives. Part 2: activity of heterocyclic sulfonamide derivatives. AB - The anti-proliferative activity of acylated heterocyclic sulfonamides is described in Vascular Endothelial Growth Factor-dependent Human Umbilical Vascular Endothelial Cells (VEGF-HUVEC) and in HCT116 tumor cells in a soft agar diffusion assay. PMID- 15664825 TI - Design and synthesis of a peptide-PEG transporter tool for carrying adenovirus vector into cells. AB - The adenovirus vector is a promising carrier for the efficient transfer of genes into cells via the coxackie-adenovirus receptor (CAR) and integrins (alphavbeta3 and alphavbeta5). The clinical use of the adenovirus vector remains problematic however. Successful administration of this vector is associated with side effects because antibodies to this vector are commonly found throughout the human body. To make the adenovirus vector practicable for clinical use, it is necessary to design an auxiliary transporter. The present study describes the use of Arg-Gly Asp(RGD)-related peptide, a peptide that binds to integrins, as an auxiliary transporter to aid efficient transport of adenovirus vector. Furthermore, poly(ethylene glycol) (PEG) was also used as a tool to modify the adenovirus such that the risk of side effects incurred during clinical application was reduced. The present study describes the design, preparation and use of (acetyl-Tyr-Gly Gly-Arg-Gly-Asp-Thr-Pro-(beta)Ala)(2)Lys-PEG-(beta)Ala-Cys-NH(2)[(Ac YGGRGDTP(beta)A)(2)K-PEG-(beta)AC] as an efficient peptide-PEG transporter tool for carrying adenovirus vector into cells. (Ac-YGGRGDTP(beta)A)(2)K-PEG-(beta)AC was coupled with 6-maleimidohexanoic acid N-hydroxysuccinimide ester and the resulting 6-[(Ac-YGGRGDTP(beta)A)(2)K-PEG-(beta)AC-succinimido]hexanoic acid N hydroxysuccinimide ester reacted with adenovirus. The modified adenovirus with the peptide-PEG hybrid exhibited high gene expression even in a CAR-negative cell line, DC2.4. PMID- 15664826 TI - A new bactericidal lead structure for the protection of materials. AB - Our search for a new broad spectrum bactericide for preserving materials lead to the discovery of a highly active bicyclic amine (1) and a number of derivatives. The synthesis and biological evaluation as well as a first toxicological assessment of these compounds are described. Compound 1 shows strong bactericidal activity down to levels of below 100 ppm but unfortunately increases the number of mutations in Ames tests. PMID- 15664827 TI - Biarylcarboxybenzamide derivatives as potent vanilloid receptor (VR1) antagonistic ligands. AB - Seventeen biarylcarboxybenzamide derivatives were prepared for the study of their agonistic/antagonistic activities to the vanilloid receptor (VR1) in rat DRG neurons. The replacement of the piperazine moiety of the lead compound 1 with phenyl ring showed quite enhanced antagonistic activity. Among the prepared derivatives, N-(4-tert-butylphenyl)-4-pyridine-2-yl-benzamide (2, IC(50)=31 nM) and N-(4-tert-butylphenyl)-4-(3-methylpyridine-2-yl)benzamide (3g, IC(50)=31 nM), showed 5-fold higher antagonistic activity than 1 in (45)Ca(2+)-influx assay. PMID- 15664828 TI - Inhibitors of type I MetAPs containing pyridine-2-carboxylic acid thiazol-2 ylamide. Part 1: SAR studies on the determination of the key scaffold. AB - Systematic SAR studies on the HTS hit pyridine-2-carboxylic acid thiazol-2 ylamide (PACT) analogues revealed that the scaffold of PCAT is indispensable for the inhibition of type I MetAP. For effective inhibition of the enzyme, the most suitable position to modify is the 3-position of the pyridine ring of PCAT, and the best substituents are those containing O or N atoms connected directly with the pyridine ring. These findings provide useful information for the design and discovery of more potent inhibitors of type I MetAPs. PMID- 15664829 TI - Inhibitors of type I MetAPs containing pyridine-2-carboxylic acid thiazol-2 ylamide. Part 2: SAR studies on the pyridine ring 3-substituent. AB - Systematic SAR studies on the pyridine ring 3-substituent of PCAT, an inhibitor of EcMetAP1 and ScMetAP1, revealed that 3-substituents have different selectivity for EcMetAP1 and ScMetAP1. The selective inhibitors of type I MetAP are useful tools for investigating the detailed interactions between the enzymes and their inhibitors. In addition, these findings provide useful information for the design and discovery of more potent inhibitors of type I MetAPs. PMID- 15664830 TI - Synthesis and SAR of 5,6-diarylpyridines as human CB1 inverse agonists. AB - Structure-activity relationship studies for two series of 2-benzyloxy-5-(4 chlorophenyl)-6-(2,4-dichlorophenyl)pyridines having either a 3-cyano or 3 carboxamide moiety resulted in the preparation of the 2-(3,4-difluorobenzyloxy)-3 nitrile analog 10d and the 2-(3,4-difluorobenzyloxy)-3-(N-propylcarboxamide) analog 16c, (hCB1 IC(50)=1.3 and 1.7 nM, respectively) as potent and selective hCB1 inverse agonists. Their synthesis and biological activities are described herein. PMID- 15664831 TI - Truncated azinomycin analogues intercalate into DNA. AB - The design and synthesis of a potentially more therapeutically-viable azinomycin analogue 4 based upon 3 has been completed. It involved coupling of a piperidine mustard to the acid chloride of the azinomycin chromophore. Both the designed azinomycin analogue 4 and the natural product 3 bind to DNA and cause unwinding, supporting an intercalative mode of binding. PMID- 15664832 TI - Pharmacophore identification of alpha(1A)-adrenoceptor antagonists. AB - A chemical feature based pharmacophore model was developed for alpha(1A) adrenoceptor antagonists by HypoGen module implemented in catalyst software package. The best scoring pharmacophore hypothesis, Hypo1, consisted of four important chemical features (one positive ion, one hydrogen-bond donor, one aromatic ring, and one hydrophobic group). The results of our study provide a valuable tool in designing new leads with desired biological activity by virtual screening. PMID- 15664833 TI - Cleavage at 5-methylcytosine in DNA by photosensitized oxidation with 2-methyl 1,4-naphthoquinone tethered oligodeoxynucleotides. AB - Photosensitized one-electron oxidation of 5-methylcytosine in DNA by 2-methyl-1,4 naphthoquinone, attached to 5'-end of an oligodeoxynucleotide strand, produced 5 formylcytosine and led to selective DNA strand cleavage at the original 5 methylcytosine configuration. This specified photoreaction is useful for positive display of 5-methylcytosine in DNA on a sequencing gel. PMID- 15664834 TI - Pyrrolidinohydroquinazolines--a novel class of CCR3 modulators. AB - A novel class of CCR3 modulators is described. Starting with lead compound 4a (K(i): 110 nM), which turned out to be an antagonist of eotaxin at the CCR3 receptor, further optimization led to compound 8b (K(i): 28 nM), which surprisingly proved to be an agonist. PMID- 15664835 TI - Parallel synthesis of pteridine derivatives as potent inhibitors for hepatitis C virus NS5B RNA-dependent RNA polymerase. AB - From compound library screening using an HCV NS5B RNA-dependent RNA polymerase enzymatic assay, we identified a pteridine hit compound with an IC(50) of 15 microM. Our SAR studies were focused on the different groups at the 6- and 7 positions, substitutions at the 4-position, and replacement of N(1) or N(3) with carbon in the pteridine ring. We found that NH or OH at 4-position is critical for the inhibitory activity. Furthermore, a hydrophobic substituent at the 4 position may help compounds permeate through the cell membrane. PMID- 15664836 TI - Synthesis and antifungal activity of noble 5-arylamino- and 6-arylthio-4,7 dioxobenzoselenazoles. AB - 5-Arylamino- and 6-arylthio-4,7-dioxobenzoselenazoles 4 and 5 were synthesized and tested for in vitro antifungal activity against Candida and Aspergillus species. 5-Arylamino-4,7-dioxobenzoselenazoles 4 showed, in general, more potent antifungal activity than 6-arylthio-4,7-dioxobenzoselenazoles 5. The results suggest that 5-arylamino-4,7-dioxobenzoselenazoles 4 would be potent antifungal agents. PMID- 15664837 TI - Stereoselective synthesis of 9-beta-d-arabianofuranosyl guanine and 2-amino-9 (beta-d-arabianofuranosyl)purine. AB - 9-beta-d-Arabianofuranosyl guanine (6) and 2-amino-9-(beta-d arabianofuranosyl)purine (8) were prepared from 2-amino-6-chloro-9-(2,3,5 triphenylmethoxyl-beta-d-arabianofuranosyl)purine (4), a key intermediate which was stereoselectively prepared from 2,3,5-triphenylmethoxyl-d-arabianofuranose and 2-amino-6-chloro-purine. The yield of the intermediate was obviously improved and only beta-isomer was formed by using the activated molecular sieve as environmental friendly catalyst, overcoming the defect that a 1:1 mixture of alpha- and beta-isomers was formed, which was difficult to separate, when toxic mercury cyanide was previously used as catalyst. PMID- 15664838 TI - Novel isoindoline compounds for potent and selective inhibition of prolyl dipeptidase DPP8. AB - DPP8 is a prolyl dipeptidase homologous to DPP-IV, which is a drug target for Type II diabetes. The biological function of DPP8 is not known. To identify potent and selective chemical compounds against DPP8, we have synthesized a series of isoquinoline and isoindoline derivatives and have tested their inhibitory activity against DPP8, DPP-IV and DPP-II. Isoindoline derivatives were found to be more potent DPP8 inhibitors than isoquinoline derivatives. Isoindoline with a 1-(4,4'-difluor-benzhydryl)-piperazine group at the P2 site was observed to be a very potent DPP8 inhibitor, having an IC(50) value of 14nM with at least a 2500-fold selectivity over either DPP-IV or DPP-II. From SAR results, we speculate that the S1 site of DPP8 may be larger than that of DPP-IV, which would allow the accommodation of larger C-terminal residues, such as isoquinoline or isoindoline. PMID- 15664839 TI - A convenient one-pot synthesis of asymmetric 1,3,5-triazine-2,4,6-triones and its application towards a novel class of gonadotropin-releasing hormone receptor antagonists. AB - A convenient one-pot synthetic route was developed for the preparation of asymmetric 1,3-dialkyl-1,3,5-triazine-2,4,6-triones from readily available alkyl- or aryl-isocyanates, primary amines and N-chlorocarbonyl isocyanate in excellent yields. Subsequent alkylation with N-protected amino alcohols afforded the desired 1,3,5-triazine-2,4,6-triones in good yields. This methodology was applied to the synthesis of a chemical library acting as antagonists of the hGnRH receptor. PMID- 15664840 TI - Discovery and structure-activity relationships of novel selective norepinephrine and dual serotonin/norepinephrine reuptake inhibitors. AB - Novel arylthiomethyl morpholines are potent selective norepinephrine reuptake inhibitors (NERIs) and dual serotonin/norepinephrine reuptake inhibitors (SRI/NERIs). The target compounds were prepared using a stereochemically versatile synthesis featuring an aldol condensation as the key step. One enantiomer of the 2-methoxy-substituted analogue was found to be a potent and selective norepinephrine reuptake inhibitor, whereas the opposite enantiomer was a potent dual serotonin/norepinephrine reuptake inhibitor. PMID- 15664841 TI - Pyrenyldiazomethane, a versatile reagent for nucleotide phosphate alkylation. AB - Pyrenyldiazomethane was shown to react quantitatively and selectively at phosphate with 2'-, 3'-, and 5'-nucleotide phosphates incorporating the different nucleic bases. PMID- 15664842 TI - Synthesis of 9-(2-beta-C-methyl-beta-d-ribofuranosyl)-6-substituted purine derivatives as inhibitors of HCV RNA replication. AB - A series of 9-(2'-beta-C-methyl-beta-d-ribofuranosyl)-6-substituted purine derivatives were synthesized as potential inhibitors of HCV RNA replication. Their inhibitory activities in a cell based HCV replicon assay were reported. A prodrug approach was used to further improve the potency of these compounds by increasing the intracellular levels of 5'-monophosphate metabolites. These nucleotide prodrugs showed much improved inhibitory activities of HCV RNA replication. PMID- 15664844 TI - 4-(2-Pyridyl)piperazine-1-benzimidazoles as potent TRPV1 antagonists. AB - A series of 4-(2-pyridyl)piperazine-1-benzimidazole analogues based on compound 1 was synthesized and evaluated for TRPV1 antagonist activity in capsaicin-induced (CAP) and pH5.5-induced (pH) FLIPR assays in a human TRPV1-expressing HEK293 cell line. Potent TRPV1 antagonists were identified through SAR studies. From these studies, several antagonists were found, with IC(50) values ranging from 32 nM to approximately 5000 nM. Among these, 11 [IC(50)=90 nM (CAP) and 104 nM (pH)] was further evaluated and found to be orally available in rats (F%=19.7). PMID- 15664843 TI - Estrogen receptor ligands. Part 11: Synthesis and activity of isochromans and isothiochromans. AB - The ring oxygen and sulfur analogs of lasofoxifene, 1a and 1b, were synthesized in an attempt to impart ERalpha selectivity, as found in the closely related dihydrobenzoxathiin compound I, recently discovered in these laboratories. The resulting isochroman and isothiochroman compounds were found to exhibit equipotent binding affinities to the ER isoforms and were less active in the inhibition of estradiol-triggered uterine growth when compared to I and lasofoxifene. PMID- 15664845 TI - Synthesis of 2'-beta-C-methyl toyocamycin and sangivamycin analogues as potential HCV inhibitors. AB - Coupling reaction of 2-beta-C-methyl-1,2,3,4-tetra-O-benzoyl-d-ribofuranose with 4-amino-6-bromo-5-cyanopyrrolo[2,3-d]pyrimidine, followed by debromination and debenzoylation, gave the 2'-beta-C-methyl toyocamycin in high yield. Based on this result, a series of 2'-beta-C-methyl-4-substituted toyocamycin and sangivamycin analogues were synthesized for biological screening as potential inhibitors of HCV RNA replication. PMID- 15664847 TI - 1-(Phenyl)isoquinoline carboxamides: a novel class of subtype selective inhibitors of thyrotropin-releasing hormone (TRH) receptors. AB - We report the synthesis of and binding to the two subtypes of mouse thyrotropin releasing hormone (TRH) receptors, TRH-R1 and TRH-R2, of several 1 (phenyl)isoquinoline carboxamide analogues. These analogues showed a degree of selectivity for binding at TRH-R2. These are the first ligands reported that show selective binding to these receptors. PMID- 15664848 TI - 3,4-Dihydro-2H-benzoxazinones are 5-HT(1A) receptor antagonists with potent 5-HT reuptake inhibitory activity. AB - Starting from a high throughput screening hit, a series of 3,4-dihydro-2H benzoxazinones has been identified with both high affinity for the 5-HT(1A) receptor and potent 5-HT reuptake inhibitory activity. The 5-(2 methyl)quinolinyloxy derivative combined high 5-HT(1A/1B/1D) receptor affinities with low intrinsic activity and potent inhibition of the 5-HT reuptake site (pK(i)8.2). This compound also had good oral bioavailability and brain penetration in the rat. PMID- 15664846 TI - Active site directed inhibitors of replication-specific bacterial DNA polymerases. AB - 7-Substituted-N(2)-(3,4-dichlorobenzyl)guanines potently and competitively inhibit DNA polymerases IIIC and IIIE from Gram(+) bacteria. Certain derivatives are also competitive inhibitors of DNA polymerase IIIE from Gram(-) bacteria. PMID- 15664849 TI - Design and synthesis of macrocycles active against vancomycin-resistant enterococci (VRE): the interplay between d-Ala-d-Lac binding and hydrophobic effect. AB - A modified vancomycin binding pocket (D-O-E ring) incorporating a CHNHCOR function at the AA4 position is designed and synthesized. Potent bioactivities against both sensitive- and resistant-strain are found for some of these compounds (MIC 4 microg/mL against VREF). From this preliminary SAR studies, it was speculated that the D-Ala-D-Ala binding was required for this series of compounds since the corresponding des-leucine derivative is inactive. The presence of long aliphatic chain was important for the desired activities and such hydrophobic effect is specific as no beneficial effect is observed when the same aliphatic chain was attached to the other part of the molecule. PMID- 15664850 TI - Novel 3-aminochromans as potential pharmacological tools for the serotonin 5 HT(7) receptor. AB - The synthesis of novel C6-aryl substituted derivatives of 3 (dimethylamino)chroman is described. The novel derivatives display 5-HT(7) receptor affinities that varies from nM to muM, indicating that this small set of derivatives constitute a novel and interesting starting point for further structure-serotonin 5-HT(7) activity relationship (SAR) studies. PMID- 15664851 TI - Quinazolinethiones and quinazolinediones, novel inhibitors of inosine monophosphate dehydrogenase: synthesis and initial structure-activity relationships. AB - The development of a series of novel quinazolinethiones and quinazolinediones as inhibitors of inosine monophosphate dehydrogenase (IMPDH) is described. The synthesis, in vitro inhibitory values for IMPDH II and in vitro inhibitory value for PBMC proliferation are discussed. PMID- 15664852 TI - Tellurium-based cysteine protease inhibitors: evaluation of novel organotellurium(IV) compounds as inhibitors of human cathepsin B. AB - New organotellurium(IV) compounds with specific cysteine protease inhibitory activity were synthesized. Serine and aspartic protease activity were not affected by any of these compounds. All Te(IV) compounds tested exhibited high specific second-order constant for cathepsin B inactivation. Tellurium(IV) compound 6 was the best inhibitor of the series, showing a second-order constant of 36,000 M(-1)s(-1). This value is about 100-fold higher than the second-order rate for cysteine protease inactivation shown by the historic Te(IV) compound AS 101 (1). The inhibition was irreversible and time and concentration dependent; no saturation kinetics were observed, suggesting a direct bimolecular reaction. The results described in this paper show that the new organotellurium(IV) compounds are powerful inhibitors of cathepsin B, constituting promising potential anti metastatic agents. PMID- 15664853 TI - Allosteric Akt (PKB) inhibitors: discovery and SAR of isozyme selective inhibitors. AB - This letter describes the development of two series of potent and selective allosteric Akt kinase inhibitors that display an unprecedented level of selectivity for either Akt1, Akt2 or both Akt1/Akt2. An iterative analog library synthesis approach quickly provided a highly selective Akt1/Akt2 inhibitor that induces apoptosis in tumor cells and inhibits Akt phosphorylation in vivo. PMID- 15664855 TI - Non-peptidic small molecule inhibitors of XIAP. AB - Non-peptidic small molecule SMAC mimetics were designed and synthesized that bind to the BIR3 domain of XIAP using structure-based design. Substituted five membered heterocycles such as thiazoles and imidazoles were identified that serve as replacements for peptide fragments of the lead. PMID- 15664854 TI - 1,4-Benzodiazepine-2,5-diones as small molecule antagonists of the HDM2-p53 interaction: discovery and SAR. AB - A library of 1,4-benzodiazepine-2,5-diones was screened for binding to the p53 binding domain of HDM2 using Thermofluor, a miniaturized thermal denaturation assay. The hits obtained were shown to bind to HDM2 in the p53-binding pocket using a fluorescence polarization (FP) peptide displacement assay. The potency of the series was optimized, leading to sub-micromolar antagonists of the p53-HDM2 interaction. PMID- 15664856 TI - Increased catalytic activity of protein disulfide isomerase using aromatic thiol based redox buffers. AB - PDI is an enzyme that acts as a chaperone, shufflase, and oxidase during the folding of disulfide-containing proteins. The ability of aromatic thiols to increase the activity of PDI-catalyzed protein folding over that of the standard thiol glutathione (GSH) was measured. 4-Mercaptobenzoic acid (ArSH) increased the activity of PDI by a factor of three. PMID- 15664857 TI - Triaryl bis-sulfones as a new class of cannabinoid CB2 receptor inhibitors: identification of a lead and initial SAR studies. AB - A novel class of cannabinoid CB2 receptor ligands is described. These triaryl bis sulfones are nanomolar inhibitors of the CB2 receptor and show high selectivity over the cannabinoid CB1 receptor. One example of this new class decreases ligand induced GTPgammaS binding to recombinant CB2 cell membranes, identifying the compound as a CB2-selective inverse agonist. PMID- 15664858 TI - N-Arylalkylpiperidine urea derivatives as CC chemokine receptor-3 (CCR3) antagonists. AB - The synthesis and structure-activity relationships of N-arylalkylpiperidylmethyl ureas as antagonists of the CC chemokine receptor-3 (CCR3) are presented. These compounds displayed potent binding to the receptor as well as functional antagonism of eotaxin-elicited effects on eosinophils. PMID- 15664859 TI - Structure-based design, synthesis and biochemical testing of novel and potent Smac peptido-mimetics. AB - Structure-based design, chemical synthesis and biochemical testing of a series of novel Smac peptido-mimetics as inhibitors of XIAP protein are described. The most potent compound, 6j, has a binding affinity (K(i) value) of 24 nM to XIAP BIR3 protein and is 24 times more potent than the native Smac AVPI peptide. Further optimization of these potent Smac mimetics may ultimately lead to the development of a novel class of anticancer drugs for the treatment of human cancer by overcoming apoptosis-resistance of cancer cells through targeting the inhibitor of apoptosis proteins. PMID- 15664860 TI - Benzimidazole derivatives as novel nonpeptide luteinizing hormone-releasing hormone (LHRH) antagonists. Part 1: Benzimidazole-5-sulfonamides. AB - A new class of benzimidazole-5-sulfonamides has been identified as nonpeptide luteinizing hormone-releasing hormone (LHRH) antagonists. Initial structure activity relationships are presented resulting in compounds 19 and 28 with submicromolar dual functional activity on human and rat receptors. PMID- 15664861 TI - Benzimidazole derivatives as novel nonpeptide luteinizing hormone-releasing hormone (LHRH) antagonists. Part 2: Benzimidazole-5-sulfonamides. AB - The 2-cyclopropyl substituted benzimidazole 2 has been used as a starting point for further optimization of an LHRH antagonist series. SAR studies revealed that a tert-butyl urea fragment connected through a simple carbon chain would improve activity. Further modification of the benzylsulfonamide moiety led to the discovery of 23 (IC(50): 4.2 nM). PMID- 15664862 TI - 1-Hydroxyalkyl-3-phenylthioureas as novel HDL-elevating agents. AB - A series of 1-hydroxyalkyl-3-phenylthiourea analogs were prepared and evaluated as HDL-and Apo A-I-elevating agents. Derivatives 5h, 7j, 7n, and 7o were found to be as effective or superior to gemfibrozil. PMID- 15664863 TI - DNA topoisomerase I inhibitors from Rinorea anguifera. AB - An organic extract prepared from Rinorea anguifera was investigated in order to identify the natural principle(s) responsible for stabilization of a topoisomerase I-DNA covalent binary complex. Bioassay-guided fractionation resulted in the isolation of mauritianin and (+)-syringaresinol as new topoisomerase I inhibitors, and also of the known inhibitor camptothecin. PMID- 15664864 TI - Design of selective phenylglycine amide tissue factor/factor VIIa inhibitors. AB - Proof of concept experiments have shown that tissue factor/factor VIIa inhibitors have antithrombotic activity without enhancing bleeding propensity. Starting from lead compounds generated by a biased combinatorial approach, phenylglycine amide tissue factor/factor VIIa inhibitors with low nanomolar affinity and good selectivity against other serine proteases of the coagulation cascade were designed, using the guidance of X-ray structural analysis and molecular modelling. PMID- 15664865 TI - Novel CDK inhibition profiles of structurally varied 1-aza-9-oxafluorenes. AB - A series of 1-aza-9-oxafluorenes with functionally varied 3-substituents have been prepared from N-phenoxycarbonyl-4-phenyl-1,4-dihydropyridines and p benzoquinone and biologically evaluated as inhibitors of various cyclin-dependant kinases. The absence of a 3-hydrogen bond acceptor function leads to a complete loss of inhibitory activity. Differing hydrogen bond acceptor functions surprisingly cause significant shifts in the selectivity of inhibition profiles. PMID- 15664866 TI - Specific recognition of napthyridine-based ligands toward guanine-containing bulges in RNA duplexes and RNA-DNA heteroduplexes. AB - Mismatched bulges in nucleic acid constructs are important in the recognition event between biological molecules. Herein, it is observed that napthyridine dimer 2 is able to specifically bind G-G mismatches in all nucleic acid constructs comprising of RNA-RNA, RNA-DNA and DNA-DNA duplexes. However, the binding affinity of 2 is strongest toward DNA duplex, followed by RNA-DNA heteroduplex and RNA duplex being the weakest binding partner. Nonetheless, this binding behavior suggests that the binding process primarily occurs between the guanine base pairs and the napthyridine moiety, and is independent of the tertiary structure of the nucleic acid duplexes. PMID- 15664867 TI - Identification of agonists and antagonists of the human melanocortin-4 receptor from piperazinebenzylamines. AB - SAR studies of a series of piperazinebenzylamines resulted in identification of potent agonists and antagonists of the human melanocortin-4 receptor. Thus, the 1,2,3,4-tetrahydroisoquinolin-1-ylacetyl compound 12e and the quinolin-3 ylcarbonyl analogue 12l possessed K(i) values of 6.3 and 4.5 nM, respectively. Interestingly, 12e was a full agonist with an EC(50) value of 31 nM, and 12l was a weak partial agonist (IA=17%) and functioned as an antagonist (IC(50)=300 nM). PMID- 15664868 TI - Crystallographic determination of stereochemistry of biologically active 2'',3'' dibromo-7-epi-10-deacetylcephalomannine. AB - The stereochemistry at C2'' and C3'' of two diastereomers of 2'',3''-dibromo-7 epi-10-deacetylcephalomannine (6 and 7), which were synthesized by reacting 7-epi 10-deacetylcephalomannine (5) with bromine, were assigned unambiguously based on crystallographic studies of 6. The X-ray crystallographic analysis shows that 6 adopts an absolute configuration of (2''S,3''R), and 7 can be assigned as (2''R,3''S) configuration. The side-chain conformation of 6 was revealed to be different with the known hydrophobic collapse and the apolar conformations, as found in solid state and in solution. However, most side-chain torsion angles of 6 were found to be very similar to those of tubulin-bound T-shaped conformation (T-Taxol). Both 6 and 7 showed strong in vitro paclitaxel-like activity. PMID- 15664870 TI - A review of sleeve fractures of the patella in children. AB - Patella fractures are rare in children and the sleeve fracture is a particular form of fracture that only occurs in children where it is the most common patella fracture. It is caused by rapid muscle contraction. Diagnosis may be difficult both clinically and on the X-ray which may look normal if there is no bony fragment. Patella alta is the best sign and ultrasound is very helpful. Awareness of the existence of the injury is all important. This fracture differs from straightforward avulsion because of the "sleeve" of periosteum which is pulled off the patella and will continue to form bone if not treated thus enlarging or even duplicating the patella. Treatment involves prompt reduction and, usually, internal fixation of the disrupted patella tendon. PMID- 15664871 TI - Retropatellar contact characteristics before and after total knee arthroplasty. AB - PURPOSE: Qualitative analysis of the retropatellar contact characteristics after total knee arthroplasty. MATERIAL AND METHODS: Six cadaveric knees were investigated before and after implantation of a Genesis I knee prosthesis without and with patellar resurfacing in different positions. Joint contact characteristics were evaluated with Fuji Prescale film type 'Super Low' and analyzed qualitatively in nine quadrants. The pressure was determined from a 5-s loading duration in four different knee positions between 45 degrees and 120 degrees of flexion. The femoral component of the prosthesis was implanted in neutral as well as in internal and external rotation. A quadriceps force of 280 N with either a predominant medial or lateral pulling direction was applied. RESULTS: Without prosthesis the largest contact area is between 60 degrees and 90 degrees of flexion. A lateral muscle force direction as well as an external rotation increased the frequency of loading in the medial quadrants. After implantation of the prosthesis the central and superior quadrants were predominantly contacted irrespective of the flexion angle. No marked differences between the flexion angles were found. Implantation of the patellar resurfacing led to contact in the three central quadrants. CONCLUSION: Implantation of the endoprosthesis leads to increased contact in slight and extreme flexion angles. Especially the central areas are increasingly loaded. No predominant influence of the rotation of the femoral component or the direction of the muscle pull was found. An improved distribution of the contact area could not be demonstrated after patellar resurfacing. PMID- 15664872 TI - Accuracy of patellar resection in total knee replacement. A study using the medial pivot knee. AB - A series of 80 patellae were randomly allocated to osteotomy by sawing or milling while implanting the medial pivot knee. Three landmarks were used to control the plane of the cut in the coronal plane. The lateral edge of the patellar tendon distally, and both medial and lateral edges of the quadriceps tendon proximally. A line drawn across the widest points if the patella (the patellar horizon) was used to analyse the slope of the cut (P angle) and the orientation of the patella relative to the trochlear groove both pre- and post-operatively (PF angle) on 45 degrees skyline views. No patellae were under resected. The mean thickness of the remnant was 16 mm (range 14-19). There was no difference between sawing and milling, but the former was technically preferable. No patellae were found to be subluxed. The mean P angle was 2 degrees (S.D. 3.2). The maximum P angle was 10 degrees , but in no cases did obliquity of cut with under resection of the medial patellar facet lead to lateral tilt of the patellar horizon. The mean post operative PF angle was 2.3 degrees (S.D. 2.6). The majority of patellae retained a similar orientation to the patellar groove post-operatively. In three patients, the patella tilted laterally with respect to the groove. One resulted from under resection of the lateral patellar facet and two in which the slope of the cut was good, probably resulted from an error in femoral rotation. In all 80 knees, the patellar dome remained in full contact with the groove. Use of three fixed landmarks provides consistent patellar resection in terms of depth and slope. Errors in slope of up to 10 degrees do not cause patellar tilt but may lead to medial overload due to increasing thickness of the patella. Errors in femoral sizing and rotation are more potent causes of lateral tilt and overload. PMID- 15664873 TI - The effects of different dressings on the skin temperature of the knee during cryotherapy. AB - AIM: To assess the effect of different dressing types on the efficiency of cryotherapy. METHODS: Eighteen normal volunteers were divided equally into group 1-no dressing, group 2-thin adhesive dressing (tegaderm), and group 3--bulky dressing ("wool and crepe"). Cryotherapy (cryocuff and autochill) was applied to one knee with the other knee serving as control. Skin temperature was measured bilaterally every 5 min for 2 h. Statistical analysis used temperature differences between control and test knees. RESULTS: The mean baseline skin temperature differences were not statistically different among the three groups (p=0.96). The mean skin temperature decreases at 2 h measured, 17 degrees C (S.D.=0.8) in group 1, 17 degrees C (S.D.=1.9) in group 2, and 5 degrees C (S.D.=1.4) in group 3. "Wool and crepe" significantly impaired the cooling effect of cryotherapy (p<0.001). Tegaderm showed no significant effect (p=0.6). CONCLUSIONS: Wool and crepe dressings following knee surgery would prevent effective cryotherapy, whereas, thin adhesive dressings would not. PMID- 15664874 TI - In-vivo sagittal plane knee kinematics: ACL intact, deficient and reconstructed knees. AB - Sagittal plane video fluoroscopy was used to analyse the bilateral knee kinematics of patients with unilateral ACL deficiency (ACLD) before, and 4 months after, hamstrings graft ACL reconstruction. Kinematics were studied during weight resisted knee extension, passive knee extension, and a step up. Thigh EMG records were also obtained from five subjects. In addition, six uninjured control subjects were analysed to establish normal kinematics. Kinematics were analysed by calculating the angle between the long axis of the tibia and the patella tendon through the range of knee flexion. Shear forces were calculated using a model of knee extension and force plate data. A maximum anterior TT of 7.3 (S.D. 2.1) mm was recorded in ACL intact (ACLI) knees during the weightbearing activity. Hamstrings activity was able to control ATT in the ACLD tibia to within the normal range of translation of the ACLI knee. Despite clinical improvements, ATT increased postoperatively in the reconstructed knees to 11.6 (S.D. 3.8) mm (P<0.048). The findings suggest that the ACL has only a minor role in the control of ATT during activity, and that the extent of ATT is more closely related to the level of quadriceps-generated shear force. A reduction in hamstrings force is proposed as the most likely cause for increased ATT post reconstruction. These findings have implications for the development of ACL rehabilitation regimes, and also may modify graft choice and the surgical aims of the procedure. PMID- 15664875 TI - Changes in arthroscopic findings in the anterior cruciate ligament deficient knee prior to reconstructive surgery. AB - We retrospectively compared the arthroscopic findings of 75 patients at the time of diagnosis of ACL rupture and the findings at the time of ACL reconstruction. We found that in the ACL deficient knee the deterioration in meniscal tears and osteochondral lesions was statistically greater with increased interval between diagnosis of ACL rupture and reconstruction. This study implies that in those patients where ACL reconstruction is indicated, a delay in reconstructive surgery can have a deleterious effect on the articular surface of the knee. PMID- 15664876 TI - Examination of the failure properties of the anterior cruciate ligament during the estrous cycle. AB - The purpose of this study was to determine whether the mechanical properties of the rat anterior cruciate ligament (ACL) vary when tested in vitro at different stages of the estrous cycle. Sixty female rats were allocated to four groups according to their stage of the estrous cycle: diestrus (n=16), proestrus (n=17), estrus (n=13) and metestrus (n=14). Right hindlimbs were harvested for mechanical testing and left hindlimbs were harvested for immunohistochemical staining to confirm the presence of the estrogen receptor. Results from the first relaxation test showed a significant difference between the estrus and proestrus stage, which was not observed in a second subsequent relaxation test. Likewise, no significant differences were found when comparing failure load and stiffness between the different stages of the estrous cycle. These results suggest that normal physiological fluctuations in estrogen during the estrous cycle did not alter the failure properties of the rat ACL. PMID- 15664877 TI - Anterior cruciate ligament reconstruction, hamstring versus bone-patella tendon bone grafts: a systematic literature review of outcome from surgery. AB - The Anterior Cruciate Ligament (ACL) is regarded as critical to the normal functioning of the knee, its disruption causing functional impairment. In recent years central third of the patellar tendon (PT) and combined Semitendinosis and gracilis tendons (HT) have become the most frequently used graft types for anterior cruciate knee ligament reconstruction. For the past two decades, the gold standard in ACL reconstructions has been the PT, but increasingly the HT graft has been used. This shift in popularity has occurred for several reasons, including concerns about damaging the knee extensor apparatus using the PT procedure, but potential complications also exist with HT techniques. Despite the vast amount of literature on ACL reconstruction and its outcome, there are very few controlled randomised studies directly comparing the two most commonly used tissue grafts. This review aimed to examine the data available from randomised trials, in order to combine and evaluate the best available evidence for choice between these two popular tissue grafts for use in ACL reconstruction. A literature search revealed 13 studies, which met the inclusion criteria of the review. The results of the 13 studies included in this review suggest that there is no significant evidence to indicate that one graft is superior. Both the PT and HT grafts appear to improve patients' performance, and therefore both would be good choices for ACL reconstruction. PMID- 15664878 TI - An experimental canine model for subchondral lesions of the knee joint. AB - Aim of the study was to create an animal model for the investigation of the role of subchondral bone damage without initial cartilage lesion in the pathogenesis of osteoarthritis, the mechanical properties of the joints as well as its role in cartilage metabolism. Therefore, after cadaver studies an animal model was created to apply a transarticular load to the femoro-patellar joint under reproducible conditions and produce a pure subchondral damage without affecting the articular cartilage. Following the cadaver studies a first group of four dogs was impacted to identify forces to produce isolated subchondral fractures in the femoral condyle. Then a second group of 12 dogs knee joints was impacted under identical conditions with forces of approximately 2100 N to produce similar subchondral fractures without cartilage damage in one joint under MRI control: T1 weighted SE-sequences. T2-weighted TSE, fat suppressed TIRM-sequences and 3D FLASH fat saturated sequences. FLASH 3D-sequences revealed intact cartilage after impact in all cases and TIRM-sequences showed subchondral fractures representing bleeding, microfractures and fragmented bone trabecules. Turbo spin echo sequences and T1-weighted images revealed other intact intraarticular structures such as ligaments and menisci. The proposed experimental animal model is suitable to investigate the effect of pure subchondral damage on the articular cartilage and on means of treatment of cartilage defects without surgical intervention and without initial cartilage damage. PMID- 15664879 TI - Intra-articular hylan G-F 20 (Synvisc) in the management of patellofemoral osteoarthritis of the knee (POAK). AB - The clinical effect of intra-articular viscosupplementation has been assessed in patients suffering tibiofemoral osteoarthritis and review of the data suggest pain relief extending to 12 months in such patients. There are no prospective trials of viscosupplementation targeting patients with patellofemoral osteoarthritis (POAK) of the knee, which is frequent, associated with anterior knee pain and often disabling. This pilot study addressed the therapeutic and adverse effects of intra-articular (IA) hylan in patients with evidence of lone POAK. Forty-three outpatients were recruited to receive a (three-injection) course of IA hylan G-F 20 (Synvisc). Patient and clinician global rating and pain on stair climbing improved significantly from 4 weeks post initial injection to 26 weeks. Secondary outcome measures (including the Oxford Knee Outcome Score) were also recorded. Adverse knee events occurred in 18.6% of patients within 48 h, but only two failed to complete the course of injections because of adverse knee events. One hundred and four patients would be required for a randomised controlled trial of IA hylan to effectively demonstrate an effect size of 0.5 (a 15-mm difference between placebo and treatment arm over a 100-mm visual analogue pain scale) with 95% confidence and 90% power, to detect a 15-mm change in VAS (effect size 0.5) with 95% confidence and 90% power. This would require a RCT of 104 patients to detect a 15-mm change in VAS (effect size 0.5) with 95% confidence and 90% power. PMID- 15664880 TI - Real-time measurement of patellofemoral kinematics in asymptomatic subjects. AB - The aims of this study were to determine whether patellofemoral kinematics in the anteroposterior (AP), mediolateral (ML) and proximal-distal (PD) directions could be measured in real time with high repeatability using a three-dimensional magnetic tracking system, and to evaluate the symmetry between sides. Data was collected from both knees of 40 asymptomatic subjects while performing knee flexion from 0 degrees to 60 degrees . Measurement of patellar displacement along the three anatomical axes (AP, ML, and PD) with respect to an anatomical coordinate system was collected using the Fastrak system. Data from three trials of each knee was plotted against knee angle and the repeatability of the data determined by examining the similarity of the movement-time curves. Symmetry was determined for maximal displacement and kinematic pattern, and a paired t-test performed to evaluate differences between sides. The data was found to be highly repeatable with mean r values for the three directions over 0.85 for both sides. The pattern of patellar displacement and maximal displacement was comparable to that reported in the literature. Results of paired t-test revealed no significant differences between the knees when comparing maximal displacement in any direction (p>0.05). Overall, there was a highly symmetrical kinematic pattern for ML and PD displacement and a small degree of asymmetry for AP displacement. However, wide variations among the subjects were noted between the left and right patellae in all directions. The results of this study have demonstrated that the use of magnetic tracking sensors provides repeatable and relevant three dimensional kinematic data of in vivo patella tracking from 0 degrees to 60 degrees of knee flexion. PMID- 15664881 TI - Clonidine induces rat aorta relaxation by nitric oxide-dependent and -independent mechanisms. AB - The alpha1- and alpha2-adrenoceptors coexist in vascular smooth muscle cells producing vascular contraction and relaxation. This study was designed to investigate which is the mechanism activated by clonidine in the rat aorta, and the endothelial factors possibly involved in the relaxation induced by clonidine. The alpha2-adrenoceptors agonist clonidine relaxed rat aortas pre-contracted with phenylephrine, with or without endothelium. In non-contracted denuded arteries, clonidine produced contractions instead of relaxation. In intact endothelium aortic rings, clonidine induced greater relaxation than in denuded aortic rings. In aortas with intact endothelium, the NO-synthase inhibitor L-NAME (10 micromol/L) and the NO-scavenger hemoglobin (10 micromol/L) reduced the relaxation to clonidine. On the other hand, indomethacin (10 micromol/L) failed to alter the relaxation induced by clonidine. These results suggest the participation of NO, but not prostacyclin in clonidine-induced relaxation. In aortic rings pre-contracted with KCl (60 mmol/L) the relaxation induced by clonidine was abolished; however, the K+ channel blockers glibenclamide (K(ATP)), tetraethylamonium (K(Ca)), and the combination of apamin and charybdotoxin (K(Ca)) did not change the relaxation induced by clonidine. The relaxation induced by clonidine on PGF2alpha-contracted arteries was not affected by prazosin. However, in the absence of prazosin, clonidine had an additional contractile effect in PGF2alpha-contracted arteries. In conclusion, our results show that in rat aorta clonidine can activate alpha2-adrenoceptors in the smooth muscle cells and alpha2-adrenoceptors in the endothelial cells that activates NO production, but not prostacyclin and/or EDHF. In the absence of phenylephrine and prazosin, clonidine can also activate alpha1-adrenoceptors and rat aorta contraction. PMID- 15664882 TI - Anti-inflammatory effects of a traditional Korean herbal formulation, Silsosangami, consisting of seven medicinal herbs: effect on hemolysis, neutrophil function, and gene expressions of iNOS and COX-2. AB - Silsosangami is a dried decoctum of a mixture of seven Korean herbal medicine, which is consisted of seven herbs (indicated as concentrations) of Typhae Pollen, Pteropi Faeces, Paeoniae Radicis rubra, Cnidii Rhizoma, Persicae Semen, Carthami Flos and Curcumae Tuber. In the present study, the effects of Silsosangami water extract (SSG) on hemolysis in human blood were studied. Using an in vitro system, only Curcumae Tuber, Persicae Semen and Paeoniae Radicis rubra had the strongest effects on hemolysis; Typhae Pollen and Pteropi Faeces had the slight effects; and Cnidii Rhizoma and Carthami Flos had no effect. On the other hand, the SSG inhibited neutrophil functions, including degranulation, superoxide generation, and leukotriene B4 production, without any effect on 5-lipoxygenase activity. This SSG reduced nitric oxide (NO) and prostaglanin E2 (PGE2) production in mouse peritoneal macrophages stimulated with lipopolysaccharide, without the influence on the activity of inducible NO synthase (iNOS), cyclooxygenase COX-2 and COX-1 being observed. SSG significantly reduced mouse paw oedema induced by carrageenan. Western blot analysis showed that SSG reduced the expression of iNOS and COX-2. These results suggested that SSG might be used as a novel antithrombotic therapeutic agents in post-myocardial infarction and also, indicated that SSG exerts anti-inflammatory effects related to the inhibition of neutrophil functions and of NO and PGE2 production, which could be due to a decreased expression of iNOS and COX-2. PMID- 15664883 TI - Varicose disease affects the P2 receptor-mediated responses of human greater saphenous vein. AB - The aim of the present study was to investigate in vitro the differences in P2 receptor mediated responses of human greater saphenous vein (GSV) taken from patients with varicose disease and obliterating atherosclerosis. Samples of the inguinal part of the GSV were taken from the patients who underwent phlebectomia operation due to varicose disease (n=9, VD group) or femoropoplitea bypass operation using auto-vein due to obliterating atherosclerosis of lower extremities (n=11, OA group). The mechanical responses of the isolated segments of GSV to P2 receptor agonists were tested using standard organ-bath technique. ATP (10(-6)-10(-4) M), ADP (10(-6)-10(-4) M) and alpha,betamethyleneATP (10(-8) 10(-5) M) caused concentration-dependent contractions of the veins of both groups, the latter agonist being approximately tenfold more active than first two. ATP at all concentrations tested, alpha,betamethyleneATP at concentrations of 10(-6) and 10(-5) M and ADP at a concentration of 10(-6) M produced significantly higher contractions of the GSV taken from OA group than from VD group. UTP (10(-6)-10(-4) M) caused concentration-dependent contractions of the veins taken from OA group, while in VD group this agonist was virtually without effect. Adenosine (10(-6)-10(-4) M) and 2-methylthio-ATP (10(-7)-10(-5) M) had no significant contractile activity in this tissue in both groups. It is concluded from this study that there are P2 receptor and adrenoceptor mediated contractions in human greater saphenous veins, which are impaired by varicose disease, in contrast to contractions produced by histamine and carbachol which are, if anything, enhanced. PMID- 15664884 TI - Mechanism of vasoselective action of mebudipine, a new calcium channel blocker. AB - In previous studies, mebudipine, a newly synthesized dihydropyridine calcium channel blocker, showed a considerable vasoselective action. To investigate the mechanism of this property, the pattern of inhibitory action on the KCl-induced contraction in isolated rat aortic rings and the effect of changing resting membrane potential on the potency of this compound were considered. In addition, its chronotropic and inotropic actions were also studied. Mebudipine inhibited KCl-induced contractions. Its inhibitory effect was progressive and needed time to reach maximum. Incubation of the aortic rings in depolarizing physiological solution (high potassium, zero calcium) resulted in the augmentation of mebudipine effect. The potency of mebudipine in inhibiting aortic contractions and its time- and voltage-dependent action were significantly greater than those of nifedipine. In comparison with nifedipine, mebudipine showed a greater negative chronotropic effect, but in the case of negative inotropism, the reverse relation was observed. It is concluded that mebudipine has a greater time- and voltage-dependent inhibitory effect, as compared to nifedipine and this property could explain its prominent vasoselective action. It has also marked negative chronotropic effect and minor negative inotropic action. With regard to the above findings, mebudipine might have a selective and protective calcium channel blocking effect in ischemic regions (ischemia-selectivity), and the potential to be used in cardiovascular diseases without causing harmful effects such as reflex tachycardia and heart failure which have sometimes been seen with the older agents. PMID- 15664885 TI - Mechanisms underlying quercetin-induced vasorelaxation in aorta of subchronic diabetic rats: an in vitro study. AB - In this study, the mechanisms involved in vasorelaxant effect of the flavonoid quercetin was investigated in isolated aortic rings from streptozotocin (STZ) diabetic rats. After 4 weeks, addition of quercetin (0.1 microM-1 mM) caused a significant dose-dependent relaxation of noradrenaline (NA)- and KCl preconstricted rings in both control and diabetic groups with a significant inter group difference of P<0.01. Furthermore, both nitro-L-arginine-methyl ester (L NAME, 100 microM) and indomethacin (10 microM) markedly attenuated the vasorelaxant responses following quercetin application. Meanwhile, endothelium removal significantly attenuated the quercetin-induced vasorelaxation. It is concluded that the quercetin can relax the preconstricted rings of aorta in subchronic STZ-diabetic rats through nitric oxide- and -prostaglandin-mediated pathways, which themselves could be considered as endothelium-dependent. PMID- 15664887 TI - Protein structural transitions and their functional role. AB - Living cells are a collection of molecular machines which carry out many of the functions essential for the cell's existence, differentiation and reproduction. Most, though not all, of these machines are made up of proteins. Because of their complexity, an understanding of how they work requires a synergistic combination of experimental and theoretical studies. In this paper we outline our studies of two such protein machines. One is GroEL, the chaperone from Escherichia coli, which aids in protein folding; the other is F(1)-ATPase, a motor protein which synthesizes and hydrolyses ATP. PMID- 15664888 TI - The energy landscape as a unifying theme in molecular science. AB - The potential energy surface (PES) underlies most calculations of structure, dynamics and thermodynamics in molecular science. In this contribution connections between the topology of the PES and observable properties are developed for a coarse-grained model of virus capsid self-assembly. The model predicts that a thermodynamically stable, kinetically accessible icosahedral shell exists for pentameric building blocks of the right shape: not too flat and not too spiky. The structure of the corresponding PES is probably common to other systems where directed searches avoid Levinthal's paradox, such as 'magic number' clusters, protein folding and crystallization. PMID- 15664889 TI - The flight of macromolecular complexes in a mass spectrometer. AB - The discovery that conditions can be found such that non-covalent macromolecular complexes can survive the transition from solution to gas phase and remain intact during their flight in a mass spectrometer is an intriguing observation. While the nature of the interaction between the components, either ionic, hydrophobic or van der Waals, undoubtedly has an effect on the stability of these gas phase species, the role of small molecules in conferring additional stability is often overlooked. Here we review historical aspects of the development of mass spectrometry for macromolecular complexes with particular focus on the role of small molecules in stabilizing gas-phase complexes. Moreover, we demonstrate how the dissociation of small molecules from subunits within a macromolecular complex can be used to probe the topological arrangement. Overall, therefore, we show that mass spectrometry used in this way is capable of addressing features of the energy landscape not readily accessed by traditional structural biology approaches. PMID- 15664890 TI - Multiple tautomers of cytosine identified and characterized by infrared laser spectroscopy in helium nanodroplets: probing structure using vibrational transition moment angles. AB - Infrared laser spectroscopy in helium nanodroplets is used to identify and characterize several distinct tautomers of cytosine. The experimentally observed species correspond to the lowest-energy structures obtained from ab initio calculations, also reported here. The assignment of the infrared vibrational bands in the spectra is aided by the measurement of the corresponding vibrational transition moment angles, which are also calculated using ab initio methods. In the present study we confirm the existence of three primary tautomers and provide tentative assignments for even higher-energy forms of cytosine in helium nanodroplets. PMID- 15664891 TI - Energy landscapes for cooperative processes: nearly ideal glass transitions, liquid-liquid transitions and folding transitions. AB - We describe basic phenomenology in the physics of supercooling liquids at constant volume (most simulations), and at constant pressure (most laboratory experiments) before focusing attention on the exceptional cases that exhibit liquid-liquid phase transitions on constant-pressure cooling. We give evidence for point defects in glasses and liquids near T(g). Models based on defects predict transitions with density gaps in constant-pressure systems. We describe the energy landscape representation of such systems. Water, in these terms, is post-critical, and its nearly ideal glass formation can be related to nucleation free protein 'funnel-folding'. For nucleated folding of proteins, a pseudo-gap should be present. Experimental methods of distinguishing between alternative folding scenarios are described. PMID- 15664892 TI - Towards complete descriptions of the free-energy landscapes of proteins. AB - In recent years increasingly detailed information about the structures and dynamics of protein molecules has been obtained by innovative applications of experimental techniques, in particular nuclear magnetic resonance spectroscopy and protein engineering, and theoretical methods, notably molecular dynamics simulations. In this article we discuss how such approaches can be combined by incorporating a wide range of different types of experimental data as restraints in computer simulations to provide unprecedented detail about the ensembles of structures that describe proteins in a wide variety of states from the native structure to highly unfolded species. Knowledge of these ensembles is beginning to enable the complete free-energy landscapes of individual proteins to be defined at atomic resolution. This strategy has provided new insights into the mechanism by which proteins are able to fold into their native states, or by which they fail to do so and give rise to harmful aggregates that are associated with a wide range of debilitating human diseases. PMID- 15664893 TI - Energy landscapes and solved protein-folding problems. AB - Energy-landscape theory has led to much progress in protein folding kinetics, protein structure prediction and protein design. Funnel landscapes describe protein folding and binding and explain how protein topology determines kinetics. Landscape-optimized energy functions based on bioinformatic input have been used to correctly predict low-resolution protein structures and also to design novel proteins automatically. PMID- 15664894 TI - Experimental configurational landscapes in aqueous solutions. AB - Structures and interactions between molecules in solution are modulated by the solvent. Changes in solvent conditions can lead to structural changes and transitions such as the assembly processes seen in micelle formation and protein folding. In the case of even quite complex liquid systems, we can now explore experimentally the configurational energy landscapes that underlie these processes. Using an aqueous solution of an amphiphile as an example, the structural transitions induced by changes in temperature, concentration and added salt are examined at the molecular level, and some critical regions of the landscape identified. Moreover, the potentials of mean force that quantitatively describe the solvent-modulated interaction between molecules in solution can now be experimentally accessed. PMID- 15664895 TI - A 'first principles' potential energy surface for liquid water from VRT spectroscopy of water clusters. AB - We present results of gas phase cluster and liquid water simulations from the recently determined VRT(ASP-W)III water dimer potential energy surface (the third fitting of the Anisotropic Site Potential with Woermer dispersion to vibration rotation-tunnelling data). VRT(ASP-W)III is shown to not only be a model of high 'spectroscopic' accuracy for the water dimer, but also makes accurate predictions of vibrational ground-state properties for clusters up through the hexamer. Results of ambient liquid water simulations from VRT(ASP-W)III are compared with those from ab initio molecular dynamics, other potentials of 'spectroscopic' accuracy and with experiment. The results herein represent the first time to the authors' knowledge that a 'spectroscopic' potential surface is able to correctly model condensed phase properties of water. PMID- 15664896 TI - Static and dynamic heterogeneities in water. AB - The thermodynamic behaviour of water seems to be related to static heterogeneities. These static heterogeneities are related to the local structure of water molecules and, when properly characterized, may offer an economical explanation of thermodynamic data. 'What matters' most in determining some of the unusual properties of liquid water may be the fact that the local geometry of the liquid molecules is not spherical or oblong, but rather tetrahedral. In respect to static heterogeneities, this local geometry is critical. The dynamic behaviour of water seems to be related to dynamic heterogeneities, which seem to explain the dynamics of supercooled liquid water well. PMID- 15664897 TI - Simulated silica. AB - We review how molecular dynamics computer simulations are providing a comprehensive picture of the behaviour of silica, as modelled by the van Beest Kramer-van Santen (BKS) potential. We have recently evaluated a number of key properties of this model system: the phase diagram, including melting lines of three crystal phases; the equation of state and free energy of the liquid phase; the dynamical equation of state; the average energy of inherent structures, and configurational entropy, associated with the potential energy landscape of the liquid; and a characterization of the local coordination environments in the supercooled liquid. The results reveal the interplay among a number of phenomena, in particular, the relationship between the energy landscape and the fragile-to strong crossover of the liquid dynamics; and the relation of both of these to the possibility of a liquid-liquid phase transition in the supercooled liquid. PMID- 15664898 TI - Exploring the temperature-pressure configurational landscape of biomolecules: from lipid membranes to proteins. AB - Hydrostatic pressure has been used as a physical parameter for studying the stability and energetics of biomolecular systems, such as lipid mesophases and proteins, but also because high pressure is an important feature of certain natural membrane environments and because the high-pressure phase behaviour of biomolecules is of biotechnological interest. By using spectroscopic and scattering techniques, the temperature- and pressure-dependent structure and phase behaviour of lipid systems, differing in chain configuration, headgroup structure and concentration, and proteins have been studied and are discussed. A thermodynamic approach is presented for studying the stability of proteins as a function of both temperature and pressure. The results demonstrate that combined temperature-pressure dependent studies can help delineate the free-energy landscape of proteins and hence help elucidate which features and thermodynamic parameters are essential in determining the stability of the native conformational state of proteins. We also introduce pressure as a kinetic variable. Applying the pressure jump relaxation technique in combination with time-resolved synchrotron X-ray diffraction and spectroscopic techniques, the kinetics of un/refolding of proteins has been studied. Finally, recent advances in using pressure for studying misfolding and aggregation of proteins will be discussed. PMID- 15664899 TI - Kinetic equivalence of the heat and cold structural transitions of lambda6-85. AB - Heat- and cold-denatured proteins are considered separate thermodynamic states because temperature tuning requires the protein to pass through two 'soft' first order phase transitions. When both pressure and temperature changes are allowed, the heat- and cold-denatured states of proteins can be interconverted without going through the native state. This raises the question of whether these states are distinguished from one another by their folding kinetics. For the Tyr22Trp/Ala37Gly/Ala49Gly mutant of the 80 residue five-helix bundle protein lambda(6-85), we show that viscosity-corrected folding rates do not distinguish the cold- and heat-denatured states. We attribute this to a folding mechanism dominated by hydrophobic collapse. Our 'temperature-symmetric' approach offers an alternative to viscosity tuning with solvent additives in such cases. PMID- 15664900 TI - Physical landscapes in biological membranes: physico-chemical terrains for spatio temporal control of biomolecular interactions and behaviour. AB - The evolving complexities of biological membranes are discussed from the point of view of potential roles of the physical constitution of the membrane. These include features of the surface and dipole potentials and membrane 'rafts'. These properties are outlined; they emphasize that protein-lipid and specific lipid environments are influential parameters in how biomolecular interactions may take place with and within membranes. Several fluorescence detection technologies directed towards measurement of these properties are also outlined that permit high-resolution experimental determination of intermolecular interactions with membranes by measuring small changes of these potentials. These point to the possibility that the membrane dipole potential in particular is enormously influential in determining the behaviour of receptor and signalling systems within membrane rafts, and offers the means of a novel mechanism for biological control. PMID- 15664901 TI - X-ray and neutron diffraction studies and MD simulation of atomic configurations in polyamorphic Y2O3-Al2O3 systems. AB - Supercooled liquids in the Y(2)O(3)-Al(2)O(3) system undergo a liquid-liquid phase transition between a high-temperature, high-density amorphous (HDA) polymorph form and one with lower density (LDA form) that is stable at lower temperature. The two amorphous polymorphs have the same chemical composition, but they differ in their density (ca. 4% difference) and in their heat content (enthalpy) and entropy determined by calorimetry. Here we present new results of structural studies using neutron and high-energy X-ray diffraction to study the structural differences between HDA and LDA polyamorphs. The combined datasets show no large differences in the average nearest-neighbour Al-O or Y-O bond lengths or coordination numbers between the low- and high-density liquids. However, the data indicate that substantial changes occur among the packing geometries and clustering of the Al-O and Y-O coordination polyhedra, i.e. within the second-nearest-neighbour shell defined by the metal-metal (i.e. Y-Y, Y-Al, Al Al) interactions. Polarizable ion model molecular dynamics simulations of Y(2)O(3)-Al(2)O(3) liquids are used to help interpret the pair-correlation functions obtained from X-ray and neutron scattering data. Unexpectedly large density fluctuations are observed to occur during the simulation of nominally equilibrated systems. These are interpreted as being due to dynamic sampling of high- and low-density configurations within the single-phase liquid at temperatures above the critical point or phase transition line. Calculated partial radial distribution functions indicate that the primary differences between HDA and LDA configurations occur among the Y-Y correlations. PMID- 15664902 TI - Diffraction, crystallography and microscopy beyond three dimensions: structural dynamics in space and time. AB - In this article we highlight recent developments of ultrafast electron diffraction and crystallography at Caltech. These developments have made it possible to resolve transient structures, both spatially (0.01 A) and temporally (picosecond and now femtosecond), in the gas phase and condensed media--surfaces, interfaces, and crystals--with wide-ranging applications. With the extension to ultrafast electron microscopy, discussed here and elsewhere, we present an overview of one major research area at our centre, the Laboratory for Molecular Sciences. PMID- 15664903 TI - A Clue to the Origin of the Bilateria? PMID- 15664904 TI - Antibody-mediated immunomodulation: a strategy to improve host responses against microbial antigens. PMID- 15664905 TI - EspJ is a prophage-carried type III effector protein of attaching and effacing pathogens that modulates infection dynamics. AB - Enterohemorrhagic Escherichia coli, enteropathogenic E. coli, and Citrobacter rodentium are highly adapted enteropathogens that successfully colonize their host's gastrointestinal tract via the formation of attaching and effacing (A/E) lesions. These pathogens utilize a type III secretion system (TTSS) apparatus, encoded by the locus of enterocyte effacement, to translocate bacterial effector proteins into epithelial cells. Here, we report the identification of EspJ (E. coli-secreted protein J), a translocated TTSS effector that is carried on the 5' end of the cryptic prophage CP-933U. Infection of epithelial cells in culture revealed that EspJ is not required for A/E lesion activity in vivo and ex vivo. However, in vivo studies performed with mice demonstrated that EspJ possesses properties that influence the dynamics of clearance of the pathogen from the host's intestinal tract, suggesting a role in host survival and pathogen transmission. PMID- 15664906 TI - Oral mucosal endotoxin tolerance induction in chronic periodontitis. AB - The oral mucosa is exposed to a high density and diversity of gram-positive and gram-negative bacteria, but very little is known about how immune homeostasis is maintained in this environment, particularly in the inflammatory disease chronic periodontitis (CP). The cells of the innate immune response recognize bacterial structures via the Toll-like receptors (TLR). This activates intracellular signaling and transcription of proteins essential for the induction of an adaptive immune response; however, if unregulated, it can lead to destructive inflammatory responses. Using single-immunoenzyme labeling, we show that the human oral mucosa (gingiva) is infiltrated by large numbers of TLR2(+) and TLR4(+) cells and that their numbers increase significantly in CP, relative to health (P < 0.05, Student's t test). We also show that the numbers of TLR2(+) but not TLR4(+) cells increase linearly with inflammation (r(2) = 0.33, P < 0.05). Double-immunofluorescence analysis confirms that TLR2 is coexpressed by monocytes (MC)/macrophages (mphi) in situ. Further analysis of gingival tissues by quantitative real-time PCR, however, indicates that despite a threefold increase in the expression of interleukin-1beta (IL-1beta) mRNA during CP, there is significant (30-fold) downregulation of TLR2 mRNA (P < 0.05, Student's t test). Also showing similar trends are the levels of TLR4 (ninefold reduction), TLR5 (twofold reduction), and MD-2 (sevenfold reduction) mRNA in CP patients compared to healthy persons, while the level of CD14 was unchanged. In vitro studies with human MC indicate that MC respond to an initial stimulus of lipopolysaccharide (LPS) from Porphyromonas gingivalis (PgLPS) or Escherichia coli (EcLPS) by upregulation of TLR2 and TLR4 mRNA and protein; moreover, IL-1beta mRNA is induced and tumor necrosis factor alpha (TNF-alpha), IL-10, IL-6, and IL-8 proteins are secreted. However, restimulation of MC with either PgLPS or EcLPS downregulates TLR2 and TLR4 mRNA and protein and IL-1beta mRNA and induces a ca. 10-fold reduction in TNF-alpha secretion, suggesting the induction of endotoxin tolerance by either LPS. Less susceptible to tolerance than TNF-alpha were IL-6, IL-10, and IL-8. These studies suggest that certain components of the innate oral mucosal immune response, most notably TLRs and inflammatory cytokines, may become tolerized during sustained exposure to bacterial structures such as LPS and that this may be one mechanism used in the oral mucosa to attempt to regulate local immune responses. PMID- 15664907 TI - Bioluminescence imaging of Toxoplasma gondii infection in living mice reveals dramatic differences between strains. AB - We examined the in vivo growth, dissemination, and reactivation of strains of the protozoan parasite Toxoplasma gondii using a bioluminescence-based imaging system. Two T. gondii strains, one with a highly virulent disease phenotype in mice (S23) and the other with a 1,000-fold-lower virulence phenotype (S22), were engineered to stably express the light-emitting protein luciferase. One clone of each wild-type strain was isolated, and the two clones (S23-luc7 and S22-luc2) were found to express similar levels of luciferase. Mice were infected intraperitoneally with S23-luc7 (50 or 5 parasites) or S22-luc2 (500, 50, or 5 parasites), and the progress of the infections was examined noninvasively following injection of the substrate for luciferase, d-luciferin. In mice infected with 50 S23-luc7 parasites, the parasites grew exponentially within the peritoneal cavity (as measured by light emitted from luciferase-expressing parasites) during days 1 to 10 p.i., and this proliferation continued until there was severe disease. In mice infected with 500 S22-luc2 parasites, the parasites proliferated in a fashion similar to the S23-luc7 proliferation during days 1 to 6, but this was followed by a precipitous drop in the signal to levels below the limit of detection. Using this technique, we were also able to observe the process of reactivation of T. gondii in chronically infected mice. After treatment with dexamethasone, we detected reactivation of toxoplasmosis in mice infected with S23-luc7 and S22-luc2. During reactivation, growth of S23-luc7 was initially detected primarily in the head and neck area, while in S22-luc2 infected mice the parasites were detected primarily in the abdomen. This method has great potential for identifying important differences in the dissemination and growth of different T. gondii strains, especially strains with dramatically different disease outcomes. PMID- 15664908 TI - Class I major histocompatibility complex presentation of antigens that escape from the parasitophorous vacuole of Toxoplasma gondii. AB - The intracellular parasite Toxoplasma gondii, the causative agent of toxoplasmosis, induces a protective CD8 T-cell response in its host; however, the mechanisms by which T. gondii proteins are presented by the class I major histocompatibility complex remain largely unexplored. T. gondii resides within a specialized compartment, the parasitophorous vacuole, that sequesters the parasite and its secreted proteins from the host cell cytoplasm, suggesting that an alternative cross-priming pathway might be necessary for class I presentation of T. gondii antigens. Here we used a strain of T. gondii expressing yellow fluorescent protein and a secreted version of the model antigen ovalbumin to investigate this question. We found that presentation of ovalbumin secreted by the parasite requires the peptide transporter TAP (transporter associated with antigen processing) and occurs primarily in actively infected cells rather than bystander cells. We also found that dendritic cells are a major target of T. gondii infection in vivo and account for much of the antigen-presenting activity in the spleen. Finally, we obtained evidence that Cre protein secreted by T. gondii can mediate recombination in the nucleus of the host cell. Together, these results indicate that Toxoplasma proteins can escape from the parasitophorous vacuole into the host cytoplasm and be presented by the endogenous class I pathway, leading to direct recognition of infected cells by CD8 T cells. PMID- 15664909 TI - HtrA stress protein is involved in intramacrophagic replication of adherent and invasive Escherichia coli strain LF82 isolated from a patient with Crohn's disease. AB - Adherent and invasive Escherichia coli (AIEC) bacteria isolated from Crohn's disease patients are able to greatly replicate within macrophages without escaping from the phagosome and without inducing macrophage death. In the present study, evidence is provided that in AIEC strain LF82 the htrA gene encoding the stress protein HtrA is essential for intracellular replication within J774-A1 macrophages. Deletion of the htrA gene in strain LF82 induced increased sensitivity of the isogenic mutant to oxidative stress caused by hydrogen peroxide and a reduced rate of growth in an acid and nutrient-poor medium partly reproducing the microenvironment of the phagosome. In vitro experiments using an LF82 htrA gene promoter fusion with the lacZ gene revealed a 38-fold activation of the promoter in AIEC LF82 intramacrophagic bacteria. The CpxRA two-component signaling pathway was not involved in this activation. In addition, the activation of the LF82 htrA gene promoter was not observed in the nonpathogenic E. coli K-12 intramacrophagic bacteria, indicating that the AIEC LF82 genetic background is crucial for induction of htrA gene transcription during phagocytosis. PMID- 15664910 TI - Identification of an iron-regulated hemin-binding outer membrane protein, HupO, in Vibrio fluvialis: effects on hemolytic activity and the oxidative stress response. AB - In pathogenic bacteria, iron acquisition is important for colonization and proliferation in the host under iron-limited conditions. The ability of Vibrio spp. to acquire iron is often critical to their virulence, causing gastroenteritis or excessive watery diarrhea in humans. In the study described here, we cloned the 2,100-bp heme utilization protein gene hupO in Vibrio fluvialis. HupO had high homology to iron-regulated outer membrane receptor proteins in Vibrio sp. and contained motifs that are common to bacterial heme receptors, including a consensus TonB box, a FRAP domain, and an NPNL domain. To characterize the hemin-binding activity of HupO, we purified the recombinant HupO protein (rHupO) from Escherichia coli by using an overexpression system. HupO was found to bind to hemin but not to hemoglobin. Sodium dodecyl sulfate polyacrylamide gel electrophoresis and Western blotting demonstrated that the 77 kDa outer membrane protein HupO of V. fluvialis was induced under iron-restricted conditions. We constructed a hupO mutant, HP1, to investigate the biochemical function of HupO in V. fluvialis. The hemolytic activity of HP1 was reduced compared to that of wild-type cells and, when exposed to hydrogen peroxide, significantly lower numbers of HP1 survived than was the case in the wild type. These results suggest that HupO is associated with virulence expression in V. fluvialis through stimulation of hemolysin production and resistance to oxidative stress. In experimentally infected mice, the 50% lethal dose value of the wild type was lower than that of the mutant, HP1. PMID- 15664912 TI - Genetic control of Bordetella pertussis infection: identification of susceptibility loci using recombinant congenic strains of mice. AB - Susceptibility to and severity of Bordetella pertussis infection in infants and children vary widely. The spectrum of clinical symptoms ranges from subclinical infection to mild disease, severe whooping cough, and death. The aims of this study were to examine genetic susceptibilities of mice to B. pertussis and to identify genetic loci in the mouse genome that are involved in susceptibility to B. pertussis infection. For this purpose we screened two sets of recombinant congenic strains (RCS) of mice, HcB and CcS, for differences in the numbers of bacteria in the lung 7 days after inoculation. For both CcS and in HcB mice, a wide range in numbers of bacteria in the lung was found, suggesting that the course of infection is under multigenic control. From both RCS sets of mice, we selected one strain to identify possible susceptibility loci in F(2) hybrid mice. The degree of lung colonization 7 days postinoculation in these F(2) mice was evaluated in relation to genetic markers by linkage analysis. We found three novel loci that are involved in the control of B. pertussis infection. One locus, designated B. pertussis susceptibility locus 1 (Bps-1), was identified on chromosome 12. The presence of the C57BL/10 genome on this locus instead of the C3H genome significantly decreased the number of B. pertussis bacteria in the lung. Bps-1 has a dominant-positive effect on the clearance of B. pertussis from the lung. The function of most genes in this region is unknown. Two other loci, Bps-2 and Bps-3, showed genetic interaction and are located on chromosomes 5 and 11. We aim to identify the gene(s) in these regions which modify susceptibility to B. pertussis. PMID- 15664911 TI - Contribution of the ATP-dependent protease ClpCP to the autolysis and virulence of Streptococcus pneumoniae. AB - The ATP-dependent caseinolytic proteases (Clp) are fundamental for stress tolerance and virulence in many pathogenic bacteria. The role of ClpC in the autolysis and virulence of Streptococcus pneumoniae is controversial. In this study, we tested the role of ClpC in a number of S. pneumoniae strains and found that the contribution of ClpC to autolysis is strain dependent. ClpC is required for the release of autolysin A and pneumolysin in serotype 2 S. pneumoniae strain D39. In vivo, ClpC is required for the growth of the pneumococcus in the lungs and blood in a murine model of disease, but it does not affect the overall outcome of pneumococcal disease. We also report the requirement of ClpP for the growth at elevated temperature and virulence of serotype 4 strain TIGR4 and confirm its contribution to the thermotolerance, oxidative stress resistance, and virulence of D39. PMID- 15664913 TI - Evaluation of the role of the Bvg intermediate phase in Bordetella pertussis during experimental respiratory infection. AB - The BvgAS system of Bordetella pertussis was traditionally considered to mediate a transition between two phenotypic phases (Bvg(+) and Bvg(-)) in response to environmental signals. We characterized a third state, the intermediate (Bvg(i)) phase, which can be induced by introducing a 1-bp substitution into bvgS (the bvgS-I1 mutation) or by growing B. pertussis under conditions intermediate between those leading to the Bvg(+) and Bvg(-) phases. Like B. bronchiseptica, B. pertussis displays in its Bvg(i) phase a characteristic colony morphology and hemolytic activity and expresses a Bvg(i)-phase-specific polypeptide called BipA, whose synthesis is regulated by bvgAS at the transcriptional level. Based on our results, we hypothesize that the Bvg(i) phase of B. pertussis may be involved in facilitating transmission between hosts. Thus, a B. pertussis mutant carrying the bvgS-I1 mutation (GMT1i) persisted at wild-type levels only in the upper murine respiratory tract. Interestingly, a bipA deletion derivative of GMT1i displayed a reduced ability to colonize the nasal cavity of mice compared with GMT1i. However, in experimental mixed infections GMT1i expressing the Bvg(i) phase could establish an initial colonization in the nose and trachea of mice as efficiently as GMT1, but the wild-type strain outcompeted GMT1i at a later time point at all sites of the respiratory tract, suggesting that the Bvg(i) phase does not serve as a phenotypic phase specialized in colonization. Finally, even though B. pertussis expresses in vitro the Bvg(i) phase at the human nasal temperature, anti-BipA antibodies were undetectable in a large collection of sera from pertussis patients. PMID- 15664914 TI - Helicobacter pylori induces transendothelial migration of activated memory T cells. AB - Helicobacter pylori infection is associated with pronounced infiltration of granulocytes and lymphocytes into the gastric mucosa, resulting in active chronic gastritis that may develop into duodenal ulcer disease or gastric adenocarcinoma. Infiltrating T cells play a major role in the pathology of these diseases, but the signals involved in recruitment of T cells from blood to H. pylori-infected tissues are not well understood. We therefore examined H. pylori-induced T-cell transendothelial migration (TEM). The Transwell system, employing a monolayer of human umbilical vein endothelial cells, was used as a model to study TEM. H. pylori induced a significant T-cell migration, compared to spontaneous migration. CD4+ and CD8+ T cells migrated to the same extent in response to H. pylori, whereas there was significantly larger transmigration of memory T cells compared to naive T cells. Both H. pylori culture filtrate and urease induced migration, and the presence of the H. pylori cag pathogenicity island increased TEM. T-cell TEM was mediated by LFA-1-ICAM-1 interactions in accordance with an increased ICAM-1 expression on the endothelial cells after contact with H. pylori. Migrating T cells had increased expression of activation marker CD69 and chemokine receptors CXCR3, CCR4, and CCR9. Furthermore, T cells migrating in response to H. pylori secreted Th1 but not Th2 cytokines upon stimulation. In conclusion, our data indicate that live H. pylori and its secreted products contribute to T-cell recruitment to the gastric mucosa and that the responding T cells have an activated memory Th1 phenotype. PMID- 15664915 TI - Enhanced killing of Candida albicans by human macrophages adherent to type 1 collagen matrices via induction of phagolysosomal fusion. AB - Candida albicans, a component of the normal flora of the alimentary tract and mucocutaneous membranes, is the leading cause of invasive fungal disease in premature infants, diabetics, and surgical patients and of oropharyngeal disease in AIDS patients. As little is known about the regulation of monocyte/macrophage anti-Candida activity, we sought to determine if fungicidal activity might be regulated by extracellular matrix proteins to which monocytes/macrophages are adherent in vivo. Compared to monocyte/macrophages that adhered to plastic, human monocytes and monocyte-derived macrophages that adhered to type 1 collagen matrices, but not to fibronectin, vitronectin, or laminin, demonstrated a significant increase in candidacidal activity. The enhancement of monocyte fungicidal activity was maintained over a 4-h period, whereas macrophage fungicidal activity was maximum at 1 h. Although adherence of monocytes and macrophages to collagen matrices concomitantly enhanced the production of superoxide anion, only the fungicidal activity of collagen-adherent monocytes was partially blocked by superoxide dismutase and catalase. Remarkably, we found that only 10% of the phagosomes in C. albicans-infected macrophages that adhered to plastic fused with lysosomes. In contrast, 80% of yeast-containing phagosomes of collagen-adherent macrophages fused with lysosomes. These data suggest that nonoxidative mechanisms are critical for human macrophage anti-Candida activity and that C. albicans pathogenicity is mediated, in part, by its ability to inhibit phagolysosomal fusion in macrophages. PMID- 15664916 TI - Rhizopus oryzae adheres to, is phagocytosed by, and damages endothelial cells in vitro. AB - Rhizopus oryzae is the most common cause of zygomycosis, a life-threatening infection that usually occurs in immunocompromised patients. A characteristic hallmark of zygomycosis is angioinvasion by the fungus, resulting in thrombosis and subsequent tissue necrosis. Interactions between R. oryzae and vascular endothelial cells are therefore likely of central importance to the organism's pathogenetic strategy. We studied the ability of R. oryzae to adhere to and damage human umbilical vein endothelial cells (HUVECs) in vitro. We report that R. oryzae spores and germ tubes adhere to HUVECs, whereas only spores adhere to subendothelial matrix proteins. Additionally, R. oryzae damages endothelial cells. This endothelial cell damage requires direct contact and subsequent phagocytosis of the fungus. Surprisingly, R. oryzae viability was not required for damage, but phagocytosis was required for dead R. oryzae to cause damage. These results elucidate the nature of R. oryzae-endothelial cell interactions, which are likely central to the angioinvasion and tissue necrosis seen during zygomycotic infections. The fact that dead R. oryzae damage human endothelial cells may, in part, explain the lack of efficacy of fungicidal agents during clinical disease. PMID- 15664918 TI - A high-affinity monoclonal antibody to anthrax protective antigen passively protects rabbits before and after aerosolized Bacillus anthracis spore challenge. AB - We have developed a therapeutic for the treatment of anthrax using an affinity enhanced monoclonal antibody (ETI-204) to protective antigen (PA), which is the central cell-binding component of the anthrax exotoxins. ETI-204 administered preexposure by a single intravenous injection of a dose of between 2.5 and 10 mg per animal significantly protected rabbits from a lethal aerosolized anthrax spore challenge ( approximately 60 to 450 times the 50% lethal dose of Bacillus anthracis Ames). Against a similar challenge, ETI-204 administered intramuscularly at a 20-mg dose per animal completely protected rabbits from death (100% survival). In the postexposure setting, intravenous administration of ETI-204 provided protection 24 h (8 of 10) and 36 h (5 of 10) after spore challenge. Administration at 48 h postchallenge, when 3 of 10 animals had already succumbed to anthrax infection, resulted in the survival of 3 of 7 animals (43%) for the duration of the study (28 days). Importantly, surviving ETI-204-treated animals were free of bacteremia by day 10 and remained so until the end of the studies. Only 11 of 51 ETI-204-treated rabbits had positive lung cultures at the end of the studies. Also, rabbits that were protected from inhalational anthrax by administration of ETI-204 developed significant titers of PA-specific antibodies. Presently, the sole therapeutic regimen available to treat infection by inhalation of B. anthracis spores is a 60-day course of antibiotics that is effective only if administered prior to or shortly after exposure. Based upon results reported here, ETI-204 is an effective therapy for prevention and treatment of inhalational anthrax. PMID- 15664917 TI - Closely related mycobacterial strains demonstrate contrasting levels of efficacy as antitumor vaccines and are processed for major histocompatibility complex class I presentation by multiple routes in dendritic cells. AB - Mycobacteria expressing recombinant antigens are already being developed as vaccines against both infections and tumors. Little is known about how dendritic cells might process such antigens. Two different mycobacterial species, the fast growing Mycobacterium smegmatis and the slow-growing M. bovis M. bovis BCG, were engineered to express a model tumor antigen, the K(b)-restricted dominant cytotoxic T-lymphocyte epitope OVA(257-264). Recombinant M. bovis BCG but not recombinant M. smegmatis conferred protection to mice challenged with the B16-OVA tumor cell line. We went on to investigate whether the contrast in antitumor efficacy could be due to differences in how dendritic cells process antigen from the two mycobacterial strains for class I presentation. Both strains of mycobacteria caused phenotypic maturation of dendritic cells, but recombinant M. smegmatis infection led to a greater degree of dendritic cell maturation than recombinant M. bovis BCG infection. Antigen from recombinant M. smegmatis was processed and presented as OVA(257-264) on K(b) molecules by the dendritic cell line DC2.4 but not by bone marrow-derived dendritic cells (BMDC) or splenic dendritic cells. In contrast, antigen from recombinant M. bovis BCG was presented by all three dendritic cell types as long as the mycobacteria were viable. Such presentation was dependent on proteasome function and nascent major histocompatibility complex (MHC) class I molecules in DC2.4 cells but independent of the proteasome and transporter associated with antigen processings (TAP) in BMDC and splenic dendritic cells. These data demonstrate for the first time that antigen vectored by the slow-growing M. bovis BCG but not that vectored by fast growing, readily destroyed M. smegmatis is processed and presented on MHC class I by in vitro-generated dendritic cells, which has implications for recombinant microbial vaccine development. PMID- 15664919 TI - Chemotaxis plays multiple roles during Helicobacter pylori animal infection. AB - Helicobacter pylori is a human gastric pathogen associated with gastric and duodenal ulcers as well as specific gastric cancers. H. pylori infects approximately 50% of the world's population, and infections can persist throughout the lifetime of the host. Motility and chemotaxis have been shown to be important in the infection process of H. pylori. We sought to address the specific roles of chemotaxis in infection of a mouse model system. We found that mutants lacking cheW, cheA, or cheY are all nonchemotactic and infect FVB/N mice with an attenuated phenotype after 2 weeks of infection. If infections proceeded for 6 months, however, this attenuation disappeared. Histological and culture analysis revealed that nonchemotactic mutants were found only in the corpus of the stomach, while the wild type occupied both the corpus and the antrum. Further analysis showed that nonchemotactic H. pylori isolates had an increased 50% infectious dose and were greatly outcompeted when coinfected with the wild type. If nonchemotactic mutants were allowed to establish an infection, subsequent infection with the wild type partially displaced the nonchemotactic mutants, indicating a role for chemotaxis in maintenance of infection. The data presented here support four roles for chemotaxis in H. pylori mouse infections: (i) establishing infection, (ii) achieving high-level infection, (iii) maintaining an infection when there are competing H. pylori present, and (iv) colonizing all regions of the stomach. PMID- 15664920 TI - Cross-protective efficacy of a prophylactic Leishmania donovani DNA vaccine against visceral and cutaneous murine leishmaniasis. AB - The fucose-mannose ligand (FML) complex of Leishmania donovani is a promising vaccine candidate against murine and canine visceral leishmaniasis, and its main component is a 36-kDa nucleoside hydrolase (NH36). In this study, we tested the immune response and protection induced by the purified FML, the recombinant NH36 (rNH36), and NH36 DNA vaccines against the agents of visceral (L. chagasi) and cutaneous (L. mexicana) leishmaniasis in BALB/c mice. Mice developed weak humoral response to the vaccines alone, except for those immunized with FML. However, all three vaccine groups presented elevated immunoglobulin G (IgG), IgG1, and IgG2a levels after infection with L. chagasi, whereas no differences were observed between vaccine and control groups after infection with L. mexicana. A strong intradermal reaction to L. donovani and L. mexicana antigens was observed in mice immunized with rNH36 or FML, whereas mice immunized with NH36 DNA only reacted against L. donovani antigens. Experimental infection of immunized mice demonstrated that FML and rNH36 induced significant protection against L. chagasi infection with reductions in parasite loads of 79%. FML also conferred partial protection against L. mexicana infection. The best protection was observed in mice immunized with the VR1012-NH36 DNA vaccine, which induced an 88% reduction in L. chagasi parasite load and a 65% reduction in L. mexicana lesion size. Fluorescence-activated cell sorting analysis indicated the DNA vaccine induced a two- to fivefold increase in gamma interferon-producing CD4(+) T cells, indicating a Th1-type immune response. Our results showed that the NH36 DNA vaccine induced a strong immunoprotection against visceral and cutaneous leishmaniasis, suggesting that this DNA vaccine represents a very good candidate for use against several Leishmania species. PMID- 15664921 TI - Mannoproteins from Cryptococcus neoformans promote dendritic cell maturation and activation. AB - Our previous data show that mannoproteins (MPs) from Cryptococcus neoformans are able to induce protective responses against both C. neoformans and Candida albicans. Here we provide evidence that MPs foster maturation and activation of human dendritic cells (DCs). Maturation was evaluated by the ability of MPs to facilitate expression of costimulatory molecules such as CD40, CD86, CD83, and major histocompatibility complex classes I and II and to inhibit receptors such as CD14, CD16, and CD32. Activation of DCs was measured by the capacity of MPs to promote interleukin-12 and tumor necrosis factor alpha secretion. DC-induced maturation and interleukin-12 induction are largely mediated by engagement of mannose receptors and presume MP internalization and degradation. DC activation leads to IkappaBalpha phosphorylation, which is necessary for nuclear factor kappaB transmigration into the nucleus. MP-loaded DCs are efficient stimulators of T cells and show a remarkable capacity to promote CD4 and CD8 proliferation. In conclusion, we have evidenced a novel regulatory role of MPs that promotes their candidacy as a vaccine against fungi. PMID- 15664922 TI - CpG oligodeoxynucleotides adsorbed onto polylactide-co-glycolide microparticles improve the immunogenicity and protective activity of the licensed anthrax vaccine. AB - To reduce the biothreat posed by anthrax, efforts are under way to improve the protection afforded by vaccination. This work examines the ability of immunostimulatory CpG oligodeoxynucleotides (ODN) adsorbed onto cationic polylactide-co-glycolide (PLG) microparticles (CpG ODN-PLG) to accelerate and boost the protective immunity elicited by Anthrax Vaccine Adsorbed (AVA, the licensed human anthrax vaccine). The results indicate that coadministering CpG ODN-PLG with AVA induces a stronger and faster immunoglobulin G response against the protective antigen of anthrax than AVA alone. Immunized mice were protected from lethal anthrax challenge within 1 week of vaccination with CpG ODN-PLG plus AVA, with the level of protection correlating with serum immunoglobulin G anti protective antigen titers. PMID- 15664923 TI - Monoacyl lipoteichoic acid from pneumococci stimulates human cells but not mouse cells. AB - We have developed a method for obtaining pneumococcal lipoteichoic acid (LTA) with none, one, or two acyl chains. Anion-exchange chromatography at pH 9.5 yields pneumococcal LTA (labeled LTA-9.5) that has a mass spectrum identical to that of pre-ion-exchange LTA and loses 500 mass units after deacylation by alkali hydrolysis. Anion exchange at pH 10.5 produces LTA (labeled LTA-10.5) with mass peaks that are 264 mass units lower than those of pre-ion-exchange LTA, and deacylation of LTA-10.5 by alkali hydrolysis reduces the mass by only 239 mass units. This result indicates that LTA-10.5 has lost one of the two acyl chains, whereas LTA-9.5 has both acyl chains. When the biological properties of LTA-9.5 and LTA-10.5 are examined with mouse cells, only LTA-9.5 (and not LTA-10.5) is able to stimulate mouse cells to produce tumor necrosis factor alpha, interleukin 1beta, and nitric oxide. In contrast, both LTA-9.5 and LTA-10.5 can stimulate human cells. LTA became inactive when both acyl chains were removed. Thus, acyl chains are critical for LTA function, and small variations in acyl chains can alter biological properties of LTA. PMID- 15664924 TI - Role of sulfated glycans in adherence of the microsporidian Encephalitozoon intestinalis to host cells in vitro. AB - Microsporidia are obligate intracellular opportunistic protists that infect a wide variety of animals, including humans, via environmentally resistant spores. Infection requires that spores be in close proximity to host cells so that the hollow polar tube can pierce the cell membrane and inject the spore contents into the cell cytoplasm. Like other eukaryotic microbes, microsporidia may use specific mechanisms for adherence in order to achieve target cell proximity and increase the likelihood of successful infection. Our data show that Encephalitozoon intestinalis exploits sulfated glycans such as the cell surface glycosaminoglycans (GAGs) in selection of and attachment to host cells. When exogenous sulfated glycans are used as inhibitors in spore adherence assays, E. intestinalis spore adherence is reduced by as much as 88%. However, there is no inhibition when nonsulfated glycans are used, suggesting that E. intestinalis spores utilize sulfated host cell glycans in adherence. These studies were confirmed by exposure of host cells to xylopyranoside, which limits host cell surface GAGs, and sodium chlorate, which decreases surface sulfation. Spore adherence studies with CHO mutant cell lines that are deficient in either surface GAGs or surface heparan sulfate also confirmed the necessity of sulfated glycans. Furthermore, when spore adherence is inhibited, host cell infection is reduced, indicating a direct association between spore adherence and infectivity. These data show that E. intestinalis specifically adheres to target cells by way of sulfated host cell surface GAGs and that this mechanism serves to enhance infectivity. PMID- 15664925 TI - Invariant Valpha14 chain NKT cells promote Plasmodium berghei circumsporozoite protein-specific gamma interferon- and tumor necrosis factor alpha-producing CD8+ T cells in the liver after poxvirus vaccination of mice. AB - Understanding the protective mechanism in the liver induced by recombinant vaccines against the pre-erythrocytic stages of malaria is important for vaccine development. Most studies in mice have focused on splenic and peripheral blood T cells and identified gamma interferon (IFN-gamma)-producing CD8+ T cells as correlates of protection, which can be induced by prime-boost vaccination with recombinant poxviruses. Invariant natural killer T (Valpha14iNKT) cells can also protect against liver stage malaria, when activated, and are abundant in the liver. Since poxviruses have nonspecific immunomodulating effects, which are incompletely understood, we investigated whether recombinant poxviruses affect the protective properties of hepatic Valpha14iNKT cells and thus vaccine efficacy. We show that intradermal vaccination with recombinant poxviruses activated Valpha14iNKT cells and NK cells in the livers of BALB/c mice while inducing IFN-gamma- and tumor necrosis factor alpha (TNF-alpha)-producing pre erythrocytic stage antigen-specific CD8+ T cells. Greater numbers of hepatic Valpha14iNKT cells secreted interleukin-4 than IFN-gamma. Vaccinated Valpha14iNKT cell-deficient mice had lower, but still protective levels of hepatic and splenic IFN-gamma+ and TNF-alpha+ CD8+ T cells and better protection rates later after challenge with Plasmodium berghei sporozoites. Therefore, vaccine-activated hepatic Valpha14iNKT cells help in generating specific T cells but are not required for protection induced by recombinant poxviruses. Furthermore, double positive INF-gamma+/TNF-alpha+ CD8+ T cells were enriched in protected livers, suggesting that cells expressing both of these cytokines may be most relevant for protection. PMID- 15664926 TI - The M protein is dispensable for maturation of streptococcal cysteine protease SpeB. AB - The streptococcal pyrogenic exotoxin B (SpeB) is an important virulence factor of group A streptococci (GAS) with cysteine protease activity. Maturation of SpeB to a proteolytically active form was suggested to be dependent on cell-wall-anchored M1 protein, the major surface protein of GAS (M. Collin and A. Olsen, Mol. Microbiol. 36:1306-1318, 2000). Collin and Olsen showed that mutant GAS strains expressing truncated M protein secrete a conformationally different form of unprocessed SpeB with no proteolytic activity. Alternatively, we hypothesized that a truncated M protein may interfere with processing of this secreted protease, and therefore we tested cysteine protease activity in genetically defined mutant strains that express either no M protein or membrane-anchored M protein with an in-frame deletion of the AB repeat region. Measurements of SpeB activity by cleavage of a substrate n-benzoyl-Pro-Phe-Arg-p-nitroanilide hydrochloride showed that the proteolytic activities in culture supernatants of both mutants were similar to those from the wild-type strain. In addition, Western blot analysis of culture supernatants showed that SpeB expression and processing to a mature form was unaffected by either deletion mutation. Therefore, we conclude that M protein is not required for maturation of the streptococcal cysteine protease SpeB. PMID- 15664927 TI - Deletion of the SSK1 response regulator gene in Candida albicans contributes to enhanced killing by human polymorphonuclear neutrophils. AB - The isolation and partial functional characterization of the two-component response regulator SSK1 gene of Candida albicans was previously reported. Compared to wild-type (CAF2-1) and gene-reconstituted (SSK23) strains, the ssk1 null strain (SSK21) was avirulent in a murine model of hematogenously disseminated candidiasis and less able to adhere to human esophageal cells. More recent data indicate that SSK21 is sensitive to 4 to 8 mM H(2)O(2) in vitro than CAF2-1 and SSK23. Furthermore, microarray studies indicate that the regulation of two classes of genes, those encoding cell wall functions and stress adaptation, are altered in the ssk1 mutant. In the present study, the susceptibility of strains CAF2-1, SSK21, and SSK23 to killing by human polymorphonuclear neutrophils (PMNs) was assessed. Results are also described for a newly constructed ssk1 mutant (SSK24) in which the URA3 gene is integrated into its native locus. Our results indicate that killing of SSK21 and SSK24 was significantly greater than that of CAF2-1 and SSK23 (P < 0.01). In order to determine why Ssk1p at least partially protects the organism against the killing activity of human PMNs, we compared the signal transduction activity and the inflammatory response gene profiles of PMNs infected with either the wild type or the ssk1 mutant. Phosphorylation of the mitogen-activated protein kinases p42/44 and p38 from neutrophils infected with either CAF2-1 (wild type) or SSK21 (ssk1/ssk1) was similar, while expression and phosphorylation of the JNK mitogen activated protein kinase was not observed following infection with either strain. On the other hand, we observed an upregulation of seven inflammatory response genes in PMNs infected with the SSK21 mutant only, while an increase in interleukin-10 expression was measured in PMNs infected with either strain. Downregulation of interleukin-2 was observed in PMNs infected with either strain. Verification of the transcriptional profiling was obtained by reverse transcription-PCR for three of the genes that were upregulated in neutrophils infected with the ssk1 mutant. Also, the sensitivity of strain SSK21 to human defensin-1, one of the nonoxidative, antimicrobial peptides of PMNs, was greater than that of CAF2-1, demonstrating that nonoxidative killing in PMNs may contribute to the increased susceptibility of the ssk1 mutant. Our results indicate that the Ssk1p response regulator protein may provide at least partial adaptive functions for the survival of C. albicans following its encounter with human neutrophils. PMID- 15664928 TI - Staphylococcus aureus virulence factors identified by using a high-throughput Caenorhabditis elegans-killing model. AB - Staphylococcus aureus is an important human pathogen that is also able to kill the model nematode Caenorhabditis elegans. We constructed a 2,950-member Tn917 transposon insertion library in S. aureus strain NCTC 8325. Twenty-one of these insertions exhibited attenuated C. elegans killing, and of these, 12 contained insertions in different genes or chromosomal locations. Ten of these 12 insertions showed attenuated killing phenotypes when transduced into two different S. aureus strains, and 5 of the 10 mutants correspond to genes that have not been previously identified in signature-tagged mutagenesis studies. These latter five mutants were tested in a murine renal abscess model, and one mutant harboring an insertion in nagD exhibited attenuated virulence. Interestingly, Tn917 was shown to have a very strong bias for insertions near the terminus of DNA replication. PMID- 15664929 TI - Solubility and bioactivity of the Pseudomonas quinolone signal are increased by a Pseudomonas aeruginosa-produced surfactant. AB - Pseudomonas aeruginosa is a gram-negative bacterium that causes serious infections in immunocompromised individuals and cystic fibrosis patients. This opportunistic pathogen controls many of its virulence factors and cellular functions through the activity of three cell-to-cell signals, N-(3-oxododecanoyl) L-homoserine lactone, N-butyryl-L-homoserine lactone, and the Pseudomonas quinolone signal (PQS). The activity of these signals is dependent upon their ability to dissolve in and freely diffuse through the aqueous solution in which P. aeruginosa happens to reside. Despite this, our data indicated that PQS was relatively insoluble in aqueous solutions, which led us to postulate that P. aeruginosa could be producing a PQS-solubilizing factor. In this report, we show that the P. aeruginosa-produced biosurfactant rhamnolipid greatly enhances the solubility of PQS in aqueous solutions. The enhanced solubility of PQS led to an increase in PQS bioactivity, as measured by both a gene induction assay and an apoptosis assay. This is the first demonstration of the importance of a bacterial surfactant in the solubilization and bioactivity of a cell-to-cell signal. PMID- 15664930 TI - A functional virulence complex composed of gingipains, adhesins, and lipopolysaccharide shows high affinity to host cells and matrix proteins and escapes recognition by host immune systems. AB - Arg-gingipain (Rgp) and Lys-gingipain (Kgp) are Porphyromonas gingivalis cysteine proteinases implicated as major virulence factors in pathologies of periodontitis. We purified a 660-kDa cell-associated gingipain complex existing as a homodimer of two catalytically active monomers which comprises their catalytic and adhesin domains. Electron microscopy revealed that the complex was composed of a globular particle with a 10-nm external diameter possessing one or two electron-dense hole-like structures. Two-dimensional gel electrophoresis and immunoblot analyses revealed the association of lipopolysaccharide (LPS) with the catalytic domains and a hemagglutinin domain, Hgp44, of Rgp and Kgp in the complex. The complex significantly degraded human type I collagen and elastin and strongly disrupted viability of human gingival fibroblasts and umbilical vein endotherial cells with an efficiency which was higher than that of the monomeric gingipains. The native complex produced only a small amount of nitrogen dioxide, tumor necrosis factor alpha, and interleukin-6 by macrophages, whereas the heat denatured complex resulted in increased production. Inhibition of the proteolytic activities of the gingipain complex did not up-regulate the cytokine production, indicating that the functional domains in LPS are structurally masked by the complex proteins. These results indicate the importance of the complex in evasion of host defense mechanisms as well as in host tissue breakdown. PMID- 15664931 TI - Contribution of three bile-associated loci, bsh, pva, and btlB, to gastrointestinal persistence and bile tolerance of Listeria monocytogenes. AB - Listeria monocytogenes must resist the deleterious actions of bile in order to infect and subsequently colonize the human gastrointestinal tract. The molecular mechanisms used by the bacterium to resist bile and the influence of bile on pathogenesis are as yet largely unexplored. This study describes the analysis of three genes--bsh, pva, and btlB--previously annotated as bile-associated loci in the sequenced L. monocytogenes EGDe genome (lmo2067, lmo0446, and lmo0754, respectively). Analysis of deletion mutants revealed a role for all three genes in resisting the acute toxicity of bile and bile salts, particularly glycoconjugated bile salts at low pH. Mutants were unaffected in the other stress responses examined (acid, salt, and detergents). Bile hydrolysis assays demonstrate that L. monocytogenes possesses only one bile salt hydrolase gene, namely, bsh. Transcriptional analyses and activity assays revealed that, although it is regulated by both PrfA and sigma(B), the latter appears to play the greater role in modulating bsh expression. In addition to being incapable of bile hydrolysis, a sigB mutant was shown to be exquisitely sensitive to bile salts. Furthermore, increased expression of sigB was detected under anaerobic conditions and during murine infection. A gene previously annotated as a possible penicillin V amidase (pva) or bile salt hydrolase was shown to be required for resistance to penicillin V but not penicillin G but did not demonstrate a role in bile hydrolysis. Finally, animal (murine) studies revealed an important role for both bsh and btlB in the intestinal persistence of L. monocytogenes. PMID- 15664932 TI - The Salmonella enterica serovar typhimurium-encoded type III secretion systems can translocate Chlamydia trachomatis proteins into the cytosol of host cells. AB - Chlamydia trachomatis is an obligate, intracellular pathogen that is a major cause of preventable blindness and infertility worldwide. Although the published genome sequence suggests that C. trachomatis encodes a type III secretion system, the lack of genetic tools for studying Chlamydia has hindered the examination of this potentially important class of virulence genes. We have developed a technique to identify Chlamydia proteins that can be translocated into the host cell cytoplasm by a type III secretion system. We have selected several Chlamydia proteins and tagged them with a multiple peptide motif element called F8M4. Epitopes contained in the F8M4 tag allow us to use tools corresponding to different arms of the adaptive immune system to detect the expression and translocation of these proteins by Salmonella enterica serovar Typhimurium. In particular, CD8(+)-T-cell reactivity can be used to detect the translocation of F8M4-tagged proteins into the cytoplasm of host cells. We have found that CD8(+) T-cell activity assays are sensitive enough to detect translocation of even a small amount of F8M4-tagged protein. We have used CD8(+)-T-cell activity to show that CopN, a Chlamydia protein previously shown to be translocated by Yersinia type III secretion, can be translocated by the Salmonella pathogenicity island 1 (SPI-1) type III secretion system. Additionally, we demonstrate that CopD and Pkn5, two Chlamydia proteins hypothesized to be substrates of a type III secretion system, are translocated via the SPI-2 type III secretion system of serovar Typhimurium. The epitope tag system described here can be used more generally to examine the expression and subcellular compartmentalization of bacterial proteins deployed during the interaction of pathogens with mammalian cells. PMID- 15664933 TI - Probiotic Lactobacillus spp. diminish Helicobacter hepaticus-induced inflammatory bowel disease in interleukin-10-deficient mice. AB - Clinical and experimental evidence has demonstrated the potential role of probiotics in the prevention or treatment of inflammatory bowel disease. Probiotic clones with direct immunomodulatory activity may have anti-inflammatory effects in the intestine. We investigated the roles of tumor necrosis factor alpha (TNF-alpha)-inhibitory Lactobacillus clones with a pathogen-induced murine colitis model. Murine-derived probiotic lactobacilli were selected in vitro for their ability to inhibit TNF-alpha secretion by Helicobacter hepaticus-stimulated macrophages. Interleukin-10 (IL-10)-deficient mice were treated with probiotic Lactobacillus reuteri in combination with Lactobacillus paracasei and then challenged with H. hepaticus. Ten weeks postinoculation, the severity of typhlocolitis was assessed by histologic examination of the cecocolic region. Intestinal proinflammatory cytokine responses were evaluated by real-time quantitative reverse transcriptase PCR and immunoassays, and the quantities of intestinal H. hepaticus were evaluated by real-time PCR. Intestinal colonization by TNF-alpha-inhibitory lactobacilli reduced intestinal inflammation in H. hepaticus-challenged IL-10-deficient mice despite similar quantities of H. hepaticus in cocolonized animals. Proinflammatory colonic cytokine (TNF-alpha and IL-12) levels were lowered in Lactobacillus-treated animals. In this H. hepaticus challenged IL-10-deficient murine colitis model, lactobacilli demonstrated probiotic effects by direct modulation of mucosal inflammatory responses. PMID- 15664934 TI - Large-scale screen highlights the importance of capsule for virulence in the zoonotic pathogen Streptococcus iniae. AB - Zoonotic pathogens have the unique ability to cross the species barrier, causing disease in both humans and specific animal hosts. Streptococcus iniae is a zoonotic pathogen of both fish and humans, and the clinical presentations of S. iniae infections in fish and humans are very similar to those caused by various human-specific streptococcal pathogens. Virulence mechanisms required for infection by this pathogen of either host have yet to be determined. Using the previously reported zebrafish infectious disease model, we performed a large scale screening to determine genes required for systemic infection. Screening 1,128 signature-tagged transposon mutants through the zebrafish model allowed identification of 41 potential mutants that were unable to survive within the host environment. Greater than 50% of the mutants that could be identified through homology searches were highly homologous to genes found in other human specific streptococcal pathogens, while 32% were found to have no homology to any sequences found in the databases, suggesting as yet unknown gram-positive bacterial virulence factors. A large percentage of the insertions were found to be located in several putative capsule synthesis genes, an important virulence component for other systemic pathogens. Density gradient assays demonstrated that several of these putative capsule mutants have dissimilar buoyant densities, suggesting different levels of capsule synthesis. Putative capsule mutants were also less resistant to phagocytosis in whole-blood assays than wild-type S. iniae. Our initial large-scale characterization of S. iniae virulence highlights the importance of the capsule for successful infection. PMID- 15664935 TI - Cytokine profiling of macrophages exposed to Porphyromonas gingivalis, its lipopolysaccharide, or its FimA protein. AB - To characterize the roles of Porphyromonas gingivalis and its components in the disease processes, we investigated the cytokine profile induced by live P. gingivalis, its lipopolysaccharides (LPS), and its major fimbrial protein, fimbrillin (FimA). Using cytokine antibody arrays, we found that P. gingivalis LPS and FimA induced a similar profile of cytokine expression when exposed to mouse peritoneal macrophages but that this profile differed significantly in response to live P. gingivalis. In vitro, mouse peritoneal macrophages were stimulated to produce interleukin-6 (IL-6), granulocyte colony-stimulating factor, and lymphotactin by live P. gingivalis, but not by P. gingivalis LPS or FimA, while RANTES, gamma interferon, IL-17, vascular cell adhesion molecule 1 (VCAM-1), and vascular endothelial growth factor were induced by P. gingivalis LPS or FimA, but not by live P. gingivalis. In vivo, IL-6 mRNA was strongly induced only by live P. gingivalis while monocyte chemoattractant protein 1 mRNA was strongly induced only by P. gingivalis LPS and FimA in mouse calvarial scalp, further confirming the differences of cytokine profile induced in vitro. Cytokine antibody arrays using toll-like receptor 2 (TLR2)- and TLR4-deficient macrophages revealed that most of the cytokines induced by P. gingivalis LPS or FimA signal through TLR2, while most of cytokines induced by live P. gingivalis signal through both TLR2 and TLR4. Interestingly, the activation of TLR2 by live P. gingivalis inhibited the release of RANTES, VCAM-1, and IL-1alpha from mouse peritoneal macrophages. A tumor necrosis factor alpha enzyme-linked immunosorbent assay further confirmed that P. gingivalis LPS and FimA activate mouse peritoneal macrophages via TLR2. These results indicate that host immune cells sense live P. gingivalis and its components differently, which translates into the expression of different inflammatory cytokine profiles. PMID- 15664936 TI - Meningococcal transferrin-binding proteins A and B show cooperation in their binding kinetics for human transferrin. AB - Neisseria meningitidis, a causative agent of bacterial meningitis and septicemia, obtains transferrin-bound iron by expressing two outer membrane-located transferrin-binding proteins, TbpA and TbpB. A novel system was developed to investigate the interaction between Tbps and human transferrin. Copurified TbpA TbpB, recombined TbpA-TbpB, and individual TbpA and TbpB were reconstituted into liposomes and fused onto an HPA chip (BIAcore). All preparations formed stable monolayers, which, with the exception of TbpB, could be regenerated by removing bound transferrin. The ligand binding properties of these monolayers were characterized with surface plasmon resonance and shown to be specific for human transferrin. Kinetic data for diferric human transferrin binding showed that recombined TbpA-TbpB had K(a) and K(d) values similar to those of copurified TbpA TbpB. Individual TbpA and TbpB also displayed K(a) values similar to those of copurified TbpA-TbpB, but their K(d) values were one order of magnitude higher. Chemical cross-linking studies revealed that TbpA and TbpB, in the absence of human transferrin, formed large complexes with TbpA as the predominant species. Upon human transferrin binding, a complex was formed with a molecular mass corresponding to that of a TbpB-human transferrin heterodimer as well as a higher molecular-mass complex of this heterodimer cross-linked to TbpA. This indicates that TbpA and TbpB form a functional meningococcal receptor complex in which there is cooperativity in the human transferrin binding kinetics. However, iron loss from the diferric human transferrin-TbpA-TbpB complex was not greater than that from human transferrin alone, suggesting that additional meningococcal transport components are involved in the process of iron removal. PMID- 15664937 TI - Inverse associations of human leukocyte antigen and malaria parasite types in two West African populations. AB - Differences in allelic associations between populations continue to cause difficulties in the mapping and identification of susceptibility genes for complex polygenic diseases. Although well recognized, the basis of such interpopulation differences is poorly understood. We present an example of an inverse allelic association of an immune response genotype to an infectious disease in two neighboring West African populations. In this case, both the key environmental contributor, i.e., the malaria parasite, and a major biological mechanism are well defined. We show that this surprising result fits well with the predictions of a mathematical model describing the population genetics and dynamics of this interaction. PMID- 15664938 TI - Protective levels of polysaccharide-specific maternal antibodies may enhance the immune response elicited by pneumococcal conjugates in neonatal and infant mice. AB - Maternal antibodies (MatAbs) may protect the offspring against infections but may also interfere with their immune responses to vaccination. We have previously shown that maternal immunization with pneumococcal polysaccharides (PPS) conjugated to tetanus protein (Pnc-TT) protected the offspring against infections caused by three important pediatric serotypes. To study the influence of MatAb on the immune response to Pnc-TT early in life, adult female mice were immunized twice with Pnc-TT of serotype 1 (Pnc1-TT), and their offspring received Pnc1-TT subcutaneously three times at 3-week intervals starting at 1 week (neonatal) or 3 weeks (infant) of age. High levels of PPS-1-specific MatAb (>3 log) in offspring of Pnc1-TT-immunized dams completely inhibited their anti-PPS-1 response elicited by Pnc1-TT. In contrast, low or moderate ( approximately 1 to 2 log) levels of MatAb did not interfere with and even enhanced the immune response of the offspring, and a booster response to a second Pnc1-TT dose was observed. Carrier specific MatAbs had little effect on the response of offspring to the conjugate. All Pnc1-TT-immunized offspring were protected against pneumococcal bacteremia and had reduced lung infection. These results demonstrate that in the presence of MatAb, Pnc1-TT may elicit a protective PPS-1-specific antibody response and prime for PPS-1-specific memory in young offspring. Importantly, low or moderate levels of PPS-1-specific MatAb not only provided protection against pneumococcal infections but also enhanced the immune response elicited by Pnc1-TT in neonatal and infant mice. This murine model will be used to develop novel strategies combining maternal and neonatal immunization to protect against infections caused by encapsulated bacteria in early life. PMID- 15664940 TI - Second-generation recombination-based in vivo expression technology for large scale screening for Vibrio cholerae genes induced during infection of the mouse small intestine. AB - We have constructed an improved recombination-based in vivo expression technology (RIVET) and used it as a screening method to identify Vibrio cholerae genes that are transcriptionally induced during infection of infant mice. The improvements include the introduction of modified substrate cassettes for resolvase that can be positively and negatively selected for, allowing selection of resolved strains from intestinal homogenates, and three different tnpR alleles that cover a range of translation initiation efficiencies, allowing identification of infection induced genes that have low-to-moderate basal levels of transcription during growth in vitro. A transcriptional fusion library of 8,734 isolates of a V. cholerae El Tor strain that remain unresolved when the vibrios are grown in vitro was passed through infant mice, and 40 infection-induced genes were identified. Nine of these genes were inactivated by in-frame deletions, and their roles in growth in vitro and fitness during infection were measured by competition assays. Four mutant strains were attenuated >10-fold in vivo compared with the parental strain, demonstrating that infection-induced genes are enriched in genes essential for virulence. PMID- 15664939 TI - The IrgA homologue adhesin Iha is an Escherichia coli virulence factor in murine urinary tract infection. AB - The role of the Escherichia coli iron-regulated gene homologue adhesin (Iha) in the pathogenesis of urinary tract infections (UTIs) is unknown. We performed a series of complementary analyses to confirm or refute the hypothesis that Iha is a virulence factor in uropathogenic E. coli. Fecal E. coli isolates exhibited significantly lower prevalences of iha (range, 14 to 22%) than did clinical isolates from cases of pediatric cystitis or pyelonephritis, adult pyelonephritis or urosepsis, or bacteremia (range, 38 to 74%). Recombinant Iha from E. coli pyelonephritis isolate CFT073 conferred upon nonadherent E. coli ORN172 the ability to adhere to cultured T-24 human uroepithelial cells. In a well established mouse model of ascending UTI, CFT073 and its derivative UPEC76 (a pap [P fimbriae] mutant version of strain CFT073) each significantly outcompeted their respective iha deletion mutants in CBA/J mice 48 h after bladder challenge (P < 0.03 for urine, both kidneys, and bladders of both constructs, except for bladders of mice challenged with UPEC76 and its deletion mutant, where P = 0.11). These data suggest that Iha(CFT073) is a virulence factor and might be a target for anti-UTI interventions. PMID- 15664941 TI - PppA, a surface-exposed protein of Streptococcus pneumoniae, elicits cross reactive antibodies that reduce colonization in a murine intranasal immunization and challenge model. AB - The multivalent pneumococcal conjugate vaccine is effective against both systemic disease and otitis media caused by serotypes contained in the vaccine. However, serotypes not covered by the present conjugate vaccine may still cause pneumococcal disease. To address these serotypes, and the remaining otitis media due to Streptococcus pneumoniae, efforts have been devoted to identifying protective protein antigens. Immunity to conserved surface proteins important for adhesion, nutrient acquisition, or other functions could result in a reduction of colonization and a lower disease potential. We have been searching for conserved surface-exposed proteins from S. pneumoniae that may be involved in pathogenesis to test as vaccine candidates. Here, an approximately 20-kDa protein that has significant homology to a nonheme iron-containing ferritin protein from Listeria innocua and other bactoferritins was identified as pneumococcal protective protein A (PppA). We expressed and purified recombinant PppA (rPppA) and evaluated its potential as a vaccine candidate. The antibodies elicited by purified rPppA were cross-reactive with PppA from multiple strains of S. pneumoniae and were directed against surface-exposed epitopes. Intranasal immunization of BALB/c mice with PppA protein and either a synthetic monophosphoryl lipid A analog, RC529AF, or a cholera toxin mutant, CT-E29H, used as an adjuvant reduced nasopharyngeal colonization in mice following intranasal challenge with a heterologous pneumococcal strain. PppA-specific systemic and local immunoglobulin G (IgG) and IgA antibody responses were induced. The antisera reacted with whole cells of a heterologous S. pneumoniae type 3 strain. These observations indicate that PppA may be a promising candidate for inclusion in a vaccine against pneumococcal otitis media. PMID- 15664942 TI - Role of sigmaB in the expression of Staphylococcus aureus cell wall adhesins ClfA and FnbA and contribution to infectivity in a rat model of experimental endocarditis. AB - Isogenic Staphylococcus aureus strains with different capacities to produce sigma(B) activity were analyzed for their ability to attach to fibrinogen- or fibronectin-coated surfaces or platelet-fibrin clots and to cause endocarditis in rats. In comparison to the sigma(B)-deficient strain, BB255, which harbors an rsbU mutation, both rsbU-complemented and sigma(B)-overproducing derivatives exhibited at least five times greater attachment to fibrinogen- and fibronectin coated surfaces and showed increased adherence to platelet-fibrin clots. No differences in adherence were seen between BB255 and a DeltarsbUVWsigB isogen. Northern blotting analyses revealed that transcription of clfA, encoding fibrinogen-binding protein clumping factor A, and fnbA, encoding fibronectin binding protein A, were positively influenced by sigma(B). Sigma(B) overproduction resulted in a statistically significant increase in positive spleen cultures and enhanced bacterial densities in both the aortic vegetations and spleens at 16 h postinoculation. In contrast, at 72 h postinoculation, tissues infected with the sigma(B) overproducer had lower bacterial densities than did those infected with BB255. These results suggest that although sigma(B) appears to increase the adhesion of S. aureus to various host cell-matrix proteins in vitro, it has limited effect on pathogenesis in the rat endocarditis model. Sigma(B) appears to have a transient enhancing effect on bacterial density in the early stages of infection that is lost during progression. PMID- 15664943 TI - Vaccination with recombinant N-terminal domain of Als1p improves survival during murine disseminated candidiasis by enhancing cell-mediated, not humoral, immunity. AB - Candida spp. are opportunistic fungal pathogens that are among the most common causes of nosocomial bloodstream infections. The mortality attributable to disseminated candidiasis is 40 to 50% despite antifungal therapy. Clearly, new strategies are needed to prevent this life-threatening infection. Because risk factors for disseminated candidiasis are well defined and frequently of limited duration, vaccination is an appealing prophylactic strategy. We have identified a cell surface protein, Als1p, that mediates adherence of Candida albicans to a variety of human substrates and plastic. Here we report that immunizing BALB/c mice with the recombinant N-terminal domain of Als1p (rAls1p-N) improved survival during a subsequent challenge with a lethal inoculum of C. albicans. The protective 20-mug dose of rAls1p-N significantly increased Candida stimulation of Th1 splenocytes and increased in vivo delayed-type hypersensitivity. In contrast, antibody titers did not correlate with protection. Finally, the vaccine was not protective in T-cell-deficient mice but was protective in B-cell-deficient mice. These data indicate that the mechanism of action of the rAls1p-N vaccine is stimulation of cell-mediated, rather than humoral, immunity against C. albicans. The majority of efforts to date have focused on the development of passive immunization strategies to prevent or treat disseminated candidiasis. In contrast, our results provide proof of principle for vaccination with an adhesin of C. albicans and emphasize the potential for cell-mediated immune modulation as a prophylactic or therapeutic strategy against disseminated candidiasis. PMID- 15664944 TI - Differential PsaA-, PspA-, PspC-, and PdB-specific immune responses in a mouse model of pneumococcal carriage. AB - Larger numbers of pneumococci were detected in the nasal tract compared to the lung, cervical lymph nodes, and spleen 1, 2, 4, 7, 14, and 21 days after nasal challenge with Streptococcus pneumoniae strain EF3030. In this mouse model of pneumococcal carriage, peripheral S. pneumoniae pneumococcal surface adhesin A (PsaA)-specific humoral responses (immunoglobulin G2a [IgG2a] >> IgG1 = IgG2b > IgG3) were significantly higher than pneumococcal surface protein A (PspA) specific, genetic toxoid derivative of pneumolysin (PdB)-specific, or pneumococcal surface protein C (PspC)-specific serum antibody levels. However, PspA-specific mucosal IgA antibody levels were significantly higher than those against PsaA, PdB, and PspC. In general, both PsaA- and PspA-specific lung-, cervical lymph node-, nasal tract-, and spleen-derived CD4(+) T-cell cytokine (interleukin-4, interleukin-6, granulocyte-macrophage colony-stimulating factor, gamma interferon, and tumor necrosis factor alpha) and proliferative responses were higher than those for either PspC or PdB. Taken together, these findings suggest that PsaA- and PspA-specific mucosal responses as well as systemic humoral and T helper cell cytokine responses are predominantly yet differentially induced during pneumococcal carriage. PMID- 15664945 TI - Borrelia burgdorferi, host-derived proteases, and the blood-brain barrier. AB - Neurological manifestations of Lyme disease in humans are attributed in part to penetration of the blood-brain barrier (BBB) and invasion of the central nervous system (CNS) by Borrelia burgdorferi. However, how the spirochetes cross the BBB remains an unresolved issue. We examined the traversal of B. burgdorferi across the human BBB and systemic endothelial cell barriers using in vitro model systems constructed of human brain microvascular endothelial cells (BMEC) and EA.hy 926, a human umbilical vein endothelial cell (HUVEC) line grown on Costar Transwell inserts. These studies showed that B. burgdorferi differentially crosses human BMEC and HUVEC and that the human BMEC form a barrier to traversal. During the transmigration by the spirochetes, it was found that the integrity of the endothelial cell monolayers was maintained, as assessed by transendothelial electrical resistance measurements at the end of the experimental period, and that B. burgdorferi appeared to bind human BMEC by their tips near or at cell borders, suggesting a paracellular route of transmigration. Importantly, traversal of B. burgdorferi across human BMEC induces the expression of plasminogen activators, plasminogen activator receptors, and matrix metalloproteinases. Thus, the fibrinolytic system linked by an activation cascade may lead to focal and transient degradation of tight junction proteins that allows B. burgdorferi to invade the CNS. PMID- 15664946 TI - Sarcocystis neurona merozoites express a family of immunogenic surface antigens that are orthologues of the Toxoplasma gondii surface antigens (SAGs) and SAG related sequences. AB - Sarcocystis neurona is a member of the Apicomplexa that causes myelitis and encephalitis in horses but normally cycles between the opossum and small mammals. Analysis of an S. neurona expressed sequence tag (EST) database revealed four paralogous proteins that exhibit clear homology to the family of surface antigens (SAGs) and SAG-related sequences of Toxoplasma gondii. The primary peptide sequences of the S. neurona proteins are consistent with the two-domain structure that has been described for the T. gondii SAGs, and each was predicted to have an amino-terminal signal peptide and a carboxyl-terminal glycolipid anchor addition site, suggesting surface localization. All four proteins were confirmed to be membrane associated and displayed on the surface of S. neurona merozoites. Due to their surface localization and homology to T. gondii surface antigens, these S. neurona proteins were designated SnSAG1, SnSAG2, SnSAG3, and SnSAG4. Consistent with their homology, the SnSAGs elicited a robust immune response in infected and immunized animals, and their conserved structure further suggests that the SnSAGs similarly serve as adhesins for attachment to host cells. Whether the S. neurona SAG family is as extensive as the T. gondii SAG family remains unresolved, but it is probable that additional SnSAGs will be revealed as more S. neurona ESTs are generated. The existence of an SnSAG family in S. neurona indicates that expression of multiple related surface antigens is not unique to the ubiquitous organism T. gondii. Instead, the SAG gene family is a common trait that presumably has an essential, conserved function(s). PMID- 15664947 TI - Temporal expression of enteropathogenic Escherichia coli virulence genes in an in vitro model of infection. AB - The hallmark of enteropathogenic Escherichia coli (EPEC) infection is the ability of EPEC to cause attaching and effacing (A/E) lesions on intestinal epithelium. This event is reproducible in in vitro tissue culture models of infection. We used real-time PCR to measure transcription from several locus of enterocyte effacement (LEE) operons (LEE1 to LEE5) and from bfp during a 5-h infection of HEp-2 cells with EPEC. We found that after the initial formation of A/E lesions, which occurs as early as 5 min postinfection, EPEC continues to increase transcription from LEE3 to LEE5 as well as from bfp. These levels are maximized by 3 h postinfection and remain constant throughout the course of infection. This increase in transcription from LEE3 to LEE5 occurs when LEE1 (ler) transcription is decreasing. EspA, EspB, intimin, Tir, and bundle-forming pilus expression is detectable during the entire 5-h infection. These results indicate that the EPEC genes involved in localized and intimate adherence are continually expressed after the initial stages of A/E lesion formation on host cells. PMID- 15664948 TI - Expression of major histocompatibility complex class II and CD80 by gingival epithelial cells induces activation of CD4+ T cells in response to bacterial challenge. AB - HLA-DR (major histocompatibility complex [MHC] class II) is often expressed by epithelial cells in gingival tissues with periodontal disease but not by cells in healthy gingival tissues. Confocal microscopic analyses revealed that gingival epithelial cells (GEC) from tissue with periodontal disease express both HLA-DR and B7-1 (CD80) costimulatory molecules. Rat GEC lines were established to elucidate the possible role of MHC class II and B7-1 expression by GEC. Stimulation of a rat GEC line with gamma interferon (IFN-gamma) induced the expression of MHC class II, whereas the cell line constitutively expressed B7-1 costimulatory molecules as determined by reverse transcription-PCR and flow cytometry. Actinobacillus actinomycetemcomitans Omp29-specific CD4(+) Th1 clone cells proliferated in response to pretreatment of GEC with fixed A. actinomycetemcomitans and IFN-gamma. However, the Th1 cells did not respond to pretreatment of GEC with the bacteria alone or IFN-gamma alone. The activation of Th1 clone cells induced by the GEC was inhibited by antibody to MHC class II or by CTLA4 immunoglobulin (CTLA4-Ig). Lymph node T cells did not demonstrate superantigen activity to A. actinomycetemcomitans, although both lymph node T cells and Th1 clone cells were sensitive to superantigen activity of staphylococcal enterotoxin A as cultured in the presence of IFN-gamma-treated GEC. These results suggested that GEC can take up bacterial antigen and consequently process and present the bacterial antigen to CD4(+) T cells by MHC class II in conjunction with B7 costimulation. GEC appeared to play a role in the adaptive immune response by stimulating antigen-specific CD4(+) T cells. PMID- 15664949 TI - Effect of capsulation of opportunistic pathogenic bacteria on binding of the pattern recognition molecules mannan-binding lectin, L-ficolin, and H-ficolin. AB - Mannan-binding lectin (MBL), L-ficolin, and H-ficolin are pattern recognition molecules of the innate immune system. We investigated their ability to bind to different serotypes and noncapsulated variants of two gram-positive bacterial species, Streptococcus pneumoniae and Staphylococcus aureus. MBL did not bind to capsulated S. aureus or capsulated S. pneumoniae but did bind to a noncapsulated S. aureus variant (Wood). L-ficolin bound to some capsulated S. aureus serotypes (serotypes 1, 8, 9, 11, and 12) and capsulated S. pneumoniae serotypes (11A, 11D, and 11F) but not to noncapsulated strains. H-ficolin did not bind to any of the S. pneumoniae and S. aureus serotypes included in this study but did bind to one strain of Aerococcus viridans. The concentrations of the three proteins in 97 plasma samples were estimated. The median concentrations were 0.8 mug per ml for MBL, 3.3 mug per ml for L-ficolin, and 18.4 mug per ml for H-ficolin. PMID- 15664950 TI - Role of contact lens wear, bacterial flora, and mannose-induced pathogenic protease in the pathogenesis of amoebic keratitis. AB - The ocular surface is continuously exposed to potential pathogens, including free living amoebae. Acanthamoeba species are among the most ubiquitous amoebae, yet Acanthamoeba keratitis is remarkably rare. The pathogenesis of Acanthamoeba keratitis is a complex, sequential process. Here we show that Acanthamoeba keratitis is profoundly affected by mannosylated proteins on the ocular surface, which stimulate the amoebae to elaborate a 133-kDa pathogenic protease. The mannose-induced protease (MIP133) mediates apoptosis of the corneal epithelium, facilitates corneal invasion, and degrades the corneal stroma. We show that contact lens wear upregulates mannosylated proteins on the corneal epithelium, stimulates MIP133 secretion, and exacerbates corneal disease. Corynebacterium xerosis, a constituent of the ocular flora, contains large amounts of mannose and is associated with Acanthamoeba keratitis. The present results show that amoebae exposed to C. xerosis produce increased amounts of MIP133 and more severe corneal disease. Oral immunization with MIP133 mitigates Acanthamoeba keratitis and demonstrates the feasibility of antidisease vaccines for pathogens that resist immune elimination. PMID- 15664951 TI - Murine macrophage transcriptional responses to Bacillus anthracis infection and intoxication. AB - Interactions between Bacillus anthracis and host macrophages represent critical early events in anthrax pathogenesis, but their details are not clearly understood. Here we report the first genomewide characterization of the transcriptional changes within macrophages infected with B. anthracis and the identification of several hundred host genes that were differentially expressed during this intracellular stage of infection. These loci included both genes that are known to be regulated differentially in response to many other bacterial pathogens and those that appear to be differentially regulated in response to B. anthracis but not other bacterial species that have been tested. These data provide a transcriptional basis for a variety of physiological changes observed during infection, including the induction of apoptosis caused by the infecting bacteria. The expression patterns underlying B. anthracis-induced apoptosis led us to test further the importance of one very highly induced macrophage gene, that for ornithine decarboxylase. Our data show that this enzyme plays an important and previously unrecognized role in suppressing apoptosis in B. anthracis-infected cells. We have also characterized the transcriptional response to anthrax lethal toxin in activated macrophages and found that, following toxin treatment, many of the host inflammatory response pathways are dampened. These data provide insights into B. anthracis pathogenesis as well as potential leads for the development of new diagnostic and therapeutic options. PMID- 15664952 TI - Murein lipoprotein is a critical outer membrane component involved in Salmonella enterica serovar typhimurium systemic infection. AB - Lipopolysaccharide (LPS) and Braun (murein) lipoprotein (Lpp) are major components of the outer membrane of gram-negative enteric bacteria that function as potent stimulators of inflammatory and immune responses. In a previous paper, we provided evidence that two functional copies of the lipoprotein gene (lppA and lppB) located on the chromosome of Salmonella enterica serovar Typhimurium contributed to bacterial virulence. In this study, we characterized lppA and lppB single-knockout (SKO) mutants and compared them with an lpp double-knockout (DKO) mutant using in vitro and in vivo models. Compared to the lpp DKO mutant, which was nonmotile, the motility of the lpp SKO mutants was significantly increased (73 to 77%), although the level of motility did not reach the level of wild-type (WT) S. enterica serovar Typhimurium. Likewise, the cytotoxicity was also significantly increased when T84 human intestinal epithelial cells and RAW264.7 murine macrophages were infected with the lpp SKO mutants compared to the cytotoxicity when cells were infected with the lpp DKO mutant. The level of interleukin-8 (IL-8) in polarized T84 cells infected with the lppB SKO mutant was significantly higher (two- to threefold higher), reaching the level in cells infected with WT S. enterica serovar Typhimurium, than the level in host cells infected with the lppA SKO mutant. The lpp DKO mutant induced minimal levels of IL-8. Similarly, sera from mice infected with the lppB SKO mutant contained 4.5- to 10-fold-higher levels of tumor necrosis factor-alpha and IL-6; the levels of these cytokines were 1.7- to 3.0-fold greater in the lppA SKO mutant-infected mice than in animals challenged with the lpp DKO mutant. The increased cytokine levels observed with the lppB SKO mutant in mice correlated with greater tissue damage in the livers and spleens of these mice than in the organs of animals infected with the lppA SKO and lpp DKO mutants. Moreover, the lppB SKO mutant infected mice had increased susceptibility to death. Since the lpp DKO mutant retained intact LPS, we constructed an S. enterica serovar Typhimurium triple knockout (TKO) mutant in which the lppA and lppB genes were deleted from an existing msbB mutant (msbB encodes an enzyme required for the acylation of lipid A). Compared to the lpp DKO and msbB SKO mutants, the lpp-msbB TKO mutant was unable to induce cytotoxicity and to produce cytokines and chemokines in vitro and in vivo. These studies provided the first evidence of the relative contributions of Lpp and lipid A acylation to Salmonella pathogenesis. PMID- 15664953 TI - Phagosomal processing of Mycobacterium tuberculosis antigen 85B is modulated independently of mycobacterial viability and phagosome maturation. AB - Control of Mycobacterium tuberculosis infection requires CD4 T-cell responses and major histocompatibility complex class II (MHC-II) processing of M. tuberculosis antigens (Ags). We have previously demonstrated that macrophages process heat killed (HK) M. tuberculosis more efficiently than live M. tuberculosis. These observations suggested that live M. tuberculosis may inhibit Ag processing by inhibiting phagosome maturation or that HK M. tuberculosis may be less resistant to Ag processing. In the present study we examined the correlation between M. tuberculosis viability and phagosome maturation and efficiency of Ag processing. Since heat treatment could render M. tuberculosis Ags more accessible to proteolysis, M. tuberculosis was additionally killed by antibiotic treatment and radiation. Processing of HK, live, radiation-killed (RadK), or rifampin-killed (RifK) M. tuberculosis in activated murine bone marrow macrophages was examined by using an I-A(b)-restricted T-cell hybridoma cell line (BB7) that recognizes an epitope derived from Ag 85B. Macrophages processed HK M. tuberculosis more rapidly and efficiently than they processed live, RadK, or RifK M. tuberculosis. Live, RadK, and RifK M. tuberculosis cells were processed with similar efficiencies for presentation to BB7 T hybridoma cells. Furthermore, phagosomes containing live or RadK M. tuberculosis expressed fewer M. tuberculosis peptide MHC-II complexes than phagosomes containing HK M. tuberculosis expressed. Since only live M. tuberculosis was able to prevent acidification of the phagosome, our results suggest that regulation of phagosome maturation does not explain the differences in processing of different forms of M. tuberculosis. These findings suggest that the mechanisms used by M. tuberculosis to inhibit phagosomal maturation differ from the mechanisms involved in modulating phagosome Ag processing. PMID- 15664954 TI - The capsular polysaccharide of Burkholderia pseudomallei contributes to survival in serum by reducing complement factor C3b deposition. AB - Burkholderia pseudomallei produces an extracellular polysaccharide capsule -3)-2 O-acetyl-6-deoxy-beta-D-manno-heptopyranose-(1- which has been shown to be an essential virulence determinant. The addition of purified capsule was shown to increase the virulence of a capsule mutant strain in the Syrian hamster model of acute melioidosis. An increase in the number of wild-type B. pseudomallei cells in the blood was seen by 48 h, while the number of capsule mutant cells in the blood declined by 48 h. Capsule expression was shown to be induced in the presence of serum using a lux reporter fusion to the capsule gene wcbB. The addition of purified B. pseudomallei capsule to serum bactericidal assays increased the survival of B. pseudomallei SLR5, a serum-sensitive strain, by 1,000-fold in normal human serum. Capsule production by B. pseudomallei contributed to reduced activation of the complement cascade by reducing the levels of complement factor C3b deposition. An increase in phagocytosis of the capsule mutant compared to the wild type was observed in the presence of normal human serum. These results suggest that the production of this capsule contributes to resistance to phagocytosis by reducing C3b deposition on the surface of the bacterium, thereby contributing to the persistence of bacteria in the blood of the infected host. Continued studies to characterize this capsule are essential for understanding the pathogenesis of B. pseudomallei infections and the development of preventive strategies for treatment of this disease. PMID- 15664955 TI - Localized multigene expression patterns support an evolving Th1/Th2-like paradigm in response to infections with Toxoplasma gondii and Ascaris suum. AB - Human infectious diseases have been studied in pigs because the two species have common microbial, parasitic, and zoonotic organisms, but there has been no systematic evaluation of cytokine gene expression in response to infectious agents in porcine species. In this study, pigs were inoculated with two clinically and economically important parasites, Toxoplasma gondii and Ascaris suum, and gene expression in 11 different tissues for 20 different swine Th1/Th2 related cytokines, cytokine receptors, and markers of immune activation were evaluated by real-time PCR. A generalized Th1-like pattern of gene expression was evident in pigs infected with T. gondii, along with an increased anti inflammatory gene expression pattern during the recovery phase of the infection. In contrast, an elevated Th2-like pattern was expressed during the period of expulsion of A. suum fourth-stage larvae from the small intestine of pigs, along with low-level Th1-like and anti-inflammatory cytokine gene expression. Prototypical immune and physiological markers of infection were observed in bronchial alveolar lavage cells, small intestinal smooth muscle, and epithelial cells. This study validated the use of a robust quantitative gene expression assay to detect immune and inflammatory markers at multiple host tissue sites, enhanced the definition of two important swine diseases, and supported the use of swine as an experimental model for the study of immunity to infectious agents relevant to humans. PMID- 15664957 TI - Antibody-mediated protection against Cryptococcus neoformans pulmonary infection is dependent on B cells. AB - The pathogenesis of pulmonary Cryptococcus neoformans infection and the efficacy of passive immunoglobulin G1 (IgG1) administration were investigated in B-cell deficient and C57BL/6J mice. C57BL/6J mice lived longer than B-cell-deficient mice after both intratracheal and intravenous infections. Administration of IgG1 prior to infection prolonged the survival of C57BL/6J mice but had no effect on the survival or numbers of CFU in the lungs of B-cell-deficient mice. C. neoformans infection in B-cell-deficient mice resulted in significantly higher levels of gamma interferon (IFN-gamma), monocyte chemoattractant protein-1 (MCP 1), and macrophage inflammatory protein-1alpha (MIP-1alpha) than in C57BL/6J mice. IgG1 administration reduced IFN-gamma and MCP-1 levels in C57BL/6J mice but not in B-cell-deficient mice. In addition, compared to its effect in C57BL/6J mice, C. neoformans infection in FcRgammaIII-deficient, athymic, and SCID mice significantly increased IFN-gamma and MCP-1 levels. IgG1 administration was associated with reduced IFN-gamma levels in C57BL/6J mice but not in FcRgammaIII deficient, athymic, and SCID mice. These observations suggest that IgG1-mediated protection in this system is a consequence of alterations in the inflammatory response that translate into less damage to the host without directly reducing the fungal burden. For hosts with impaired immunities, the ineffectiveness of passive antibody (Ab) may reflect an inability to down-regulate inflammation and avoid self-damage. The results indicate an important role for B cells in host defense against C. neoformans infection and demonstrate a surprising dependence of Ab-mediated protection on B cells in this system. PMID- 15664956 TI - A549 lung epithelial cells grown as three-dimensional aggregates: alternative tissue culture model for Pseudomonas aeruginosa pathogenesis. AB - A three-dimensional (3-D) lung aggregate model was developed from A549 human lung epithelial cells by using a rotating-wall vessel bioreactor to study the interactions between Pseudomonas aeruginosa and lung epithelial cells. The suitability of the 3-D aggregates as an infection model was examined by immunohistochemistry, adherence and invasion assays, scanning electron microscopy, and cytokine and mucoglycoprotein production. Immunohistochemical characterization of the 3-D A549 aggregates showed increased expression of epithelial cell-specific markers and decreased expression of cancer-specific markers compared to their monolayer counterparts. Immunohistochemistry of junctional markers on A549 3-D cells revealed that these cells formed tight junctions and polarity, in contrast to the cells grown as monolayers. Additionally, the 3-D aggregates stained positively for the production of mucoglycoprotein while the monolayers showed no indication of staining. Moreover, mucin-specific antibodies to MUC1 and MUC5A bound with greater affinity to 3-D aggregates than to the monolayers. P. aeruginosa attached to and penetrated A549 monolayers significantly more than the same cells grown as 3-D aggregates. Scanning electron microscopy of A549 cells grown as monolayers and 3-D aggregates infected with P. aeruginosa showed that monolayers detached from the surface of the culture plate postinfection, in contrast to the 3-D aggregates, which remained attached to the microcarrier beads. In response to infection, proinflammatory cytokine levels were elevated for the 3-D A549 aggregates compared to monolayer controls. These findings suggest that A549 lung cells grown as 3-D aggregates may represent a more physiologically relevant model to examine the interactions between P. aeruginosa and the lung epithelium during infection. PMID- 15664958 TI - The region comprising amino acids 100 to 255 of Neisseria meningitidis lipoprotein GNA 1870 elicits bactericidal antibodies. AB - GNA 1870 is a novel surface-exposed lipoprotein, identified by genome analysis of Neisseria meningitidis strain MC58, which induces bactericidal antibodies. Three sequence variants of the protein were shown to be sufficient to induce bactericidal antibodies against a panel of strains representative of the diversity of serogroup B meningococci. Here, we studied the antigenic and immunogenic properties of GNA 1870, which for convenience was divided into domains A, B, and C. The immune responses of mice immunized with each of the three variants were tested using overlapping peptides scanning the entire protein length and using recombinant fragments. We found that while most of the linear epitopes are located in the A domain, the bactericidal antibodies are directed against conformational epitopes located in the BC domain. This was also confirmed by the isolation of a bactericidal murine monoclonal antibody, which failed to recognize linear peptides on the A, B, and C domains separately but recognized a conformational epitope formed only by the combination of the B and C domains. Arginine in position 204 was identified as important for binding of the monoclonal antibody. The identification of the region containing bactericidal epitopes is an important step in the design of new vaccines against meningococci. PMID- 15664959 TI - Mouse model of enteropathogenic Escherichia coli infection. AB - Enteropathogenic Escherichia coli (EPEC) is an important cause of diarrhea in humans. EPEC infection of cultured intestinal epithelial cells induces attaching and effacing (A/E) lesions, alters intestinal ion transport, increases paracellular permeability, and stimulates inflammation. The lack of a small animal model has restricted in vivo studies examining EPEC-host interactions. The aim of this study was to characterize the C57BL/6J mouse as a model of EPEC infection. We have shown that EPEC can adhere to and colonize the intestinal epithelium of C57BL/6J mice. Animal weight and water intake were not altered during 10 days of EPEC infection. The proximal colon of infected mice contained semisolid stool, with stool pellets forming only in the distal colon. In contrast, the entire colon of control mice contained formed stool. Microvillous effacement and actin rearrangement, characteristic of A/E lesions, were seen in the intestine of infected mice but not control mice. Histological assessment revealed increased numbers of lamina propria neutrophils with occasional crypt abscesses, intraepithelial lymphocytes, and goblet cells in the intestine of EPEC infected mice. Altogether, these data suggest that the C57BL/6J mouse is susceptible to infection by EPEC and will provide a suitable in vivo model for studying the consequences of EPEC infection. PMID- 15664960 TI - Fusobacterium nucleatum increases collagenase 3 production and migration of epithelial cells. AB - Fusobacterium nucleatum is closely associated with human periodontal diseases and may also be a causative agent in other infections, such as pericarditis, septic arthritis, and abscesses of tonsils and liver. Initiation and outcome of infective diseases depend critically on the host cell signaling system altered by the microbe. Production of proteinases by infected cells is an important factor in pericellular tissue destruction and cell migration. We studied binding of F. nucleatum to human epithelial cells (HaCaT keratinocyte line) and subsequent cell signaling related to collagenase 3 expression, cell motility, and cell survival, using a scratch wound cell culture model. F. nucleatum increased levels of 12 protein kinases involved in cell migration, proliferation, and cell survival signaling, as assessed by the Kinetworks immunoblotting system. Epithelial cells of the artificial wound margins were clearly preferential targets of F. nucleatum. The bacterium colocalized with lysosomal structures and stimulated migration of these cells. Of the 13 anaerobic oral bacterial species, F. nucleatum and Fusobacterium necrophorum were among the best inducers of collagenase 3 mRNA levels, a powerful matrix metalloproteinase. Production of collagenase 3 was detected in fusobacterium-infected cells and cell culture medium by immunocytochemistry, immunoblotting, and zymography. The proteinase production involved activation of p38 mitogen-activated protein kinase in the infected cells. The study suggests that F. nucleatum may be involved in the pathogenesis of periodontal diseases (and other infections) by activating multiple cell signaling systems that lead to stimulation of collagenase 3 expression and increased migration and survival of the infected epithelial cells. PMID- 15664961 TI - Long-term survival and intracellular replication of Mycoplasma hominis in Trichomonas vaginalis cells: potential role of the protozoon in transmitting bacterial infection. AB - The existence of a symbiotic relationship between Trichomonas vaginalis and Mycoplasma hominis, which is the first reported example of symbiosis between two obligate human pathogens, has been recently reported by our research group. In this work, we examined the cellular location of M. hominis in respect to T. vaginalis. By using gentamicin protection assays, double immunofluorescence, and confocal microscopy, we obtained strong evidence that M. hominis is located within protozoan cells. 5-Bromodeoxyuridine incorporation assays showed that intracellularly located mycoplasmas actively synthesize DNA. Our results demonstrate that M. hominis has the capability of entering trichomonad cells and of replicating inside the protozoon. These findings suggest that symbiosis might provide the bacteria, during human infection, with the capability to resist to environmental stresses, such as host defense mechanisms and pharmacological therapies. PMID- 15664962 TI - Defective phagocytosis in Anaplasma phagocytophilum-infected neutrophils. AB - Anaplasma phagocytophilum infection induces functional neutrophil changes. Using both Candida albicans and fluorescent-aggregate phagocytosis assays, we examined whether neutrophil and dimethyl sulfoxide-differentiated HL-60 cell infection impairs internalization. A. phagocytophilum infection significantly decreased phagocytosis compared to that of controls (P < 0.05). This further impairment of neutrophil function may promote opportunistic infections and exacerbate disease. PMID- 15664963 TI - Distinct Leishmania braziliensis isolates induce different paces of chemokine expression patterns. AB - Inflammatory events during Leishmania braziliensis infection in mice were investigated. Large lesions were directly correlated with the inflammatory reaction but not with parasite burden. Different L. braziliensis strains induce different paces of chemokine expression patterns, leading to diverse cell recruitment and differential inflammatory responses. PMID- 15664964 TI - Long-term protection against tuberculosis following vaccination with a severely attenuated double lysine and pantothenate auxotroph of Mycobacterium tuberculosis. AB - We report the safety and immunogenicity of a double lysine and pantothenate auxotroph of Mycobacterium tuberculosis in mice. The DeltalysA DeltapanCD mutant is completely attenuated in immunocompromised SCID and gamma interferon knockout mice yet induces short-term and long-term protection in immunocompetent and CD4 deficient mice following single-dose subcutaneous vaccination. PMID- 15664965 TI - Salmonella-induced filament formation is a dynamic phenotype induced by rapidly replicating Salmonella enterica serovar typhimurium in epithelial cells. AB - Salmonella enterica serovar Typhimurium has the fascinating ability to form tubular structures known as Salmonella-induced filaments (Sifs) in host cells. Here, we show that the prevalence of the Sif phenotype in HeLa cells is affected by host cell density, growth, and the multiplicity of infection. Sif formation was observed in cells that displayed rapid intracellular bacterial replication and was found to be dynamic, being maximal 8 to 10 h postinfection and declining thereafter. The virulence factors SpvB and SseJ were found to negatively modulate Sif formation. Our findings demonstrate the complex and dynamic nature of the Sif phenotype. PMID- 15664966 TI - Peroxisome proliferator-activated receptor gamma and retinoid X receptor agonists have minimal effects on the interaction of endothelial cells with Plasmodium falciparum-infected erythrocytes. AB - Peroxisome proliferator-activated receptor gamma-retinoid X receptor (PPARgamma RXR) agonists had minimal effects on the surface levels of CD36, intercellular cell adhesion molecule-1, or platelet-endothelial cell adhesion molecule-1 and had no effect on the cytoadherence of infected erythrocytes to either human umbilical vein endothelial cells or human microvascular endothelial cells or on malaria-induced interleukin-6 secretion from these cells. PPARgamma-RXR agonists do not significantly modify malaria-infected erythrocyte-endothelial cell interactions in vitro. PMID- 15664967 TI - Colonization of gnotobiotic piglets by a luxS mutant strain of Escherichia coli O157:H7. AB - Gnotobiotic piglets inoculated with Escherichia coli O157:H7, its luxS mutant derivative, or nonpathogenic E. coli were evaluated for attaching and effacing lesions. Although no differences in clinical symptoms were seen between pigs inoculated with the parent and those inoculated with the luxS mutant, the luxS mutant-inoculated pigs had a lower frequency of attaching and effacing lesions in the spiral colon than parent strain-inoculated pigs. PMID- 15664968 TI - Modulation of an outer membrane protease contributes to the virulence defect of Shigella flexneri strains carrying a mutation in the virK locus. AB - The Shigella actin assembly protein IcsA is removed from the bacterial surface by the protease IcsP. We show that decreased intracellular spreading of virK::Tn10 mutants is due in part to significant increases in IcsP and IcsP-mediated cleavage of IcsA and that IcsP expression is a critical determinant of Shigella virulence. PMID- 15664969 TI - Identification of new hmwA alleles from nontypeable Haemophilus influenzae. AB - High-molecular-weight proteins of Haemophilus influenzae mediate attachment to epithelial cells. Previous reports describe several allelic versions of hmwA genes that have different adherence properties. Here we report three new alleles of hmwA (hmwA from strain AAr96, hmwA from strain AAr105, and hmwA from strain G822), demonstrating the high degree of DNA variation of these genes among different strains. PMID- 15664970 TI - Down-regulation of the kps region 1 capsular assembly operon following attachment of Escherichia coli type 1 fimbriae to D-mannose receptors. AB - A differential-display PCR procedure identified the capsular assembly gene kpsD after Escherichia coli type 1 fimbrial binding to mannose-coated Sepharose beads. Limiting-dilution reverse-transcribed PCRs confirmed down-regulation of the kpsD gene, and Northern blot and lacZ fusion analyses showed down-regulation of the kpsFEDUCS region 1 operon. KpsD protein levels fell, and an agglutination test showed less K capsular antigen on the surface following the bacterial ligand receptor interaction. These data show that binding of type 1 fimbriae (pili) to d mannose receptors triggers a cross talk that leads to down-regulation of the capsule assembly region 1 operon in uropathogenic E. coli. PMID- 15664971 TI - Age-dependent changes in susceptibility of suckling mice to individual strains of Helicobacter pylori. AB - To model establishment of Helicobacter pylori infection in infants, suckling mice were inoculated with mixtures of strains that preferentially colonize different gastric regions and coexist in vivo. Characterization of H. pylori recovered 2 weeks later showed that susceptibility begins earlier for some strains than for others and that the onset of susceptibility varies among mouse lines. PMID- 15664972 TI - Plasmodium falciparum merozoite surface protein 6 displays multiple targets for naturally occurring antibodies that mediate monocyte-dependent parasite killing. AB - Plasmodium falciparum MSP6 is a merozoite surface antigen that shows organization and sequence homologies similar to those of MSP3. Within its C-terminus conserved region, it presents some epitopes that are cross-reactive with MSP3 and others that are not, both being targets of naturally occurring antibodies that block the P. falciparum erythrocytic cycle in cooperation with monocytes. PMID- 15664973 TI - CaNAT1, a heterologous dominant selectable marker for transformation of Candida albicans and other pathogenic Candida species. AB - A dominant selectable marker for Candida albicans and other Candida species, which confers resistance to nourseothricin, was characterized. In a heterologous promoter system and a recyclable cassette, the marker efficiently permitted deletion and complementation of C. albicans genes. Neither growth nor filamentous development was affected in strains expressing this marker. PMID- 15664974 TI - Interaction of enteropathogenic Escherichia coli with human intestinal mucosa: role of effector proteins in brush border remodeling and formation of attaching and effacing lesions. AB - Enteropathogenic Escherichia coli (EPEC) strains deliver effector proteins Tir, EspB, Map, EspF, EspH, and EspG into host cells to induce brush border remodeling and produce attaching and effacing (A/E) lesions on small intestinal enterocytes. In this study, the role of individual EPEC effectors in brush border remodeling and A/E lesion formation was investigated with an in vitro human small intestinal organ culture model of EPEC infection and specific effector mutants. tir, map, espB, and espH mutants produced "footprint" phenotypes due to close bacterial adhesion but subsequent loss of bacteria; an espB mutant and other type III secretion system mutants induced a "noneffacing footprint" associated with intact brush border microvilli, whereas a tir mutant was able to efface microvilli resulting in an "effacing footprint"; map and espH mutants produced A/E lesions, but loss of bacteria resulted in a "pedestal footprint." An espF mutant produced typical A/E lesions without associated microvillous elongation. An espG mutant was indistinguishable from the wild type. These observations indicate that Tir, Map, EspF, and EspH effectors play a role in brush border remodeling and production of mature A/E lesions. PMID- 15664976 TI - Characterization of an extracellular dipeptidase from Streptococcus gordonii FSS2. AB - PepV, a dipeptidase found in culture fluids of Streptococcus gordonii FSS2, was purified and characterized, and its gene was cloned. PepV is a monomeric metalloenzyme of approximately 55 kDa that preferentially degrades hydrophobic dipeptides. The gene encodes a polypeptide of 467 amino acids, with a theoretical molecular mass of 51,114 Da and a calculated pI of 4.8. The S. gordonii PepV gene is homologous to the PepV gene family from Lactobacillus and Lactococcus spp. PMID- 15664975 TI - Mutagenesis of a novel gene in the prcA-prtP protease locus affects expression of Treponema denticola membrane complexes. AB - A novel gene was identified in the Treponema denticola prcA-prtP protease operon. Strains with mutations in either the prcA-prtP or the msp region showed altered expression of a product(s) of the other locus. Together, these results provide information on the assembly of outer membrane complexes involved in T. denticola interaction with host cells and tissue. PMID- 15664977 TI - A type IV pilin, PilA, Contributes To Adherence of Burkholderia pseudomallei and virulence in vivo. AB - The Burkholderia pseudomallei K96243 genome contains multiple type IV pilin associated loci, including one encoding a putative pilus structural protein (pilA). A pilA deletion mutant has reduced adherence to human epithelial cells and is less virulent in the nematode model of virulence and the murine model of melioidosis, suggesting a role for type IV pili in B. pseudomallei virulence. PMID- 15664979 TI - Split decision: what happens to nucleosomes during DNA replication? PMID- 15664978 TI - Lactobacillus johnsonii La1 antagonizes Giardia intestinalis in vivo. AB - This study describes the in vivo activity of Lactobacillus johnsonii La1 (NCC533) in Giardia intestinalis-infected gerbils (Meriones unguiculatus). Daily administration of lactobacilli in the drinking water from 7 days before inoculation with Giardia trophozoites efficiently prevented G. intestinalis strain WB clone C6 from infecting gerbils. More specifically, shedding of fecal Giardia antigens (GSA65 protein) was diminished in the La1-treated group, and resolution of infection was observed by 21 days postinoculation. Histology and analysis of enzymatic markers of microvillus membrane integrity revealed that probiotic administration also protected against parasite-induced mucosal damage. In addition, a cellular response to Giardia antigens was stimulated in spleen cells from La1-treated gerbils. Results show for the first time the antigiardial effect of probiotic lactobacilli in vivo and provide further insight into the antagonistic properties of lactic acid bacteria against protozoa involved in intestinal infections. PMID- 15664980 TI - Co-translational involvement of the chaperonin GroEL in the folding of newly translated polypeptides. AB - A large fraction of the newly translated polypeptides emerging from the ribosome require certain proteins, the so-called molecular chaperones, to assist in their folding. In Escherichia coli, three major chaperone systems are considered to contribute to the folding of newly synthesized cytosolic polypeptides. Trigger factor (TF), a ribosome-tethered chaperone, and DnaK are known to exhibit overlapping co-translational roles, whereas the cage-shaped GroEL, with the aid of the co-chaperonin, GroES, and ATP, is believed to be implicated in folding only after the polypeptides are released from the ribosome. However, the recent finding that GroEL-GroES overproduction permits the growth of E. coli cells lacking both TF and DnaK raised questions regarding the separate roles of these chaperones. Here, we report the puromycin-sensitive association of GroEL-GroES with translating ribosomes in vivo. Further experiments in vitro, using a reconstituted cell-free translation system, clearly demonstrate that GroEL associates with the translation complex and accomplishes proper folding by encapsulating the newly translated polypeptides in the central cavity formed by GroES. Therefore, we propose that GroEL is a versatile chaperone, which participates in the folding pathway co-translationally and also achieves correct folding post-translationally. PMID- 15664981 TI - In vivo evidence for the specificity of Plasmodium falciparum phosphoethanolamine methyltransferase and its coupling to the Kennedy pathway. AB - Unlike humans and yeast, Plasmodium falciparum, the agent of the most severe form of human malaria, utilizes host serine as a precursor for the synthesis of phosphatidylcholine via a plant-like pathway involving phosphoethanolamine methylation. The monopartite phosphoethanolamine methyltransferase, Pfpmt, plays an important role in the biosynthetic pathway of this major phospholipid by providing the precursor phosphocholine via a three-step S-adenosyl-L-methionine dependent methylation of phosphoethanolamine. In vitro studies showed that Pfpmt has strong specificity for phosphoethanolamine. However, the in vivo substrate (phosphoethanolamine or phosphatidylethanolamine) is not yet known. We used yeast as a surrogate system to express Pfpmt and provide genetic and biochemical evidence demonstrating the specificity of Pfpmt for phosphoethanolamine in vivo. Wild-type yeast cells, which inherently lack phosphoethanolamine methylation, acquire this activity as a result of expression of Pfpmt. The Pfpmt restores the ability of a yeast mutant pem1Deltapem2Delta lacking the phosphatidylethanolamine methyltransferase genes to grow in the absence of choline. Lipid analysis of the Pfpmt-complemented pem1Deltapem2Delta strain demonstrates the synthesis of phosphatidylcholine but not the intermediates of phosphatidylethanolamine transmethylation. Complementation of the pem1Deltapem2Delta mutant relies on specific methylation of phosphoethanolamine but not phosphatidylethanolamine. Interestingly, a mutation in the yeast choline-phosphate cytidylyltransferase gene abrogates the complementation by Pfpmt thus demonstrating that Pfpmt activity is directly coupled to the Kennedy pathway for the de novo synthesis of phosphatidylcholine. PMID- 15664982 TI - Interdimer processing and linearity of procaspase-3 activation. A unifying mechanism for the activation of initiator and effector caspases. AB - Caspase activation during apoptosis occurs in a cascade from the initiator caspase(s) (e.g. caspase-8) to the effector caspases (e.g. caspase-3), which ensures the generation of large amounts of active caspases to dismantle cells. However, the mechanism that safeguards against inadvertent caspase activation is not well understood. Previous studies have suggested that the activation of procaspase-8 is mediated by cross-cleavage of precursor dimers, formed upon apoptosis induction, which are not only enzymatically competent but also highly susceptible to cleavage, and that procaspase-8 activation is a linear process without self-amplification. Effector procaspases constitutively exist as dimers and their activation is started by trans-cleavage by an initiator caspase followed by autocleavage of effector caspases. Here we show that the dimerization of caspase-3 molecules through their protease domains is required for their processing by initiator caspases. The subsequent autoprocessing takes place through cleavage between the dimeric intermediates. Moreover, mature caspase-3 fails to process its own precursor. Thus, despite a marked difference in the generation of active intermediates, the activation of initiator and effector caspases shares the features of interdimer cleavage and lack of self amplification. These features may be important in preventing accidental cell death. PMID- 15664983 TI - Two anti-radial spoke monoclonal antibodies inhibit Chlamydomonas axonemal motility by different mechanisms. AB - In the 9 + 2 axoneme, radial spokes are structural components attached to the A tubules of the nine outer doublet microtubules. They protrude toward the central pair microtubule complex with which they have transient but regular interactions for the normal flagellar motility to occur. Flagella of Chlamydomonas mutants deficient in entire radial spokes or spoke heads are paralyzed. In this study the importance of two radial spoke proteins in the flagellar movement is exemplified by the potent inhibitory action of two monoclonal antibodies on the axonemal motility of demembranated-reactivated Chlamydomonas models. We show that one of these proteins is localized on the stalk of the radial spokes, whereas the other is a component of the head of the same structure and most likely correspond to radial spoke protein 2 and 1, respectively. Fine motility analysis by videomicrography further indicates that these two anti-radial spoke protein antibodies at low concentration affect motility of demembranated-reactivated Chlamydomonas by changing the flagellar waveform without modifying axonemal beat frequency. They also modify wave amplitude differently during motility inhibition. This brings more direct evidence for the involvement of both radial spoke stalk and head in the fine tuning of the waveform during flagellar motility. PMID- 15664984 TI - Paired cysteine mutagenesis to establish the pattern of disulfide bonds in the functional intact secretin receptor. AB - The amino-terminal domain of class B G protein-coupled receptors contains six conserved cysteine residues involved in structurally and functionally critical disulfide bonds. The mapping of these bonds has been unclear, with one pattern based on biochemical and NMR structural characterizations of refolded, nonglycosylated amino-terminal fragments, and another pattern derived from functional characterizations of intact receptors having paired cysteine mutations. In the present study, we determined the disulfide bonding pattern of the prototypic class B secretin receptor by applying the same paired cysteine mutagenesis approach and confirming the predicted bonding pattern with proteolytic cleavage of intact functional receptor. As expected, systematic mutation to serine of the six conserved cysteine residues within this region of the secretin receptor singly and in pairs resulted in loss of function of most constructs. Notable exceptions were single mutations of the 4th and 6th cysteine residues and paired mutations involving the 1st and 3rd, 2nd and 5th, and 4th and 6th conserved cysteines, with secretin eliciting statistically significant cAMP responses above basal levels of activation for each of these constructs. Immunofluorescence microscopy confirmed similar levels of plasma membrane expression for each of the mutated receptors. Furthermore, cyanogen bromide cleaved a series of wild type and mutant secretin receptors, yielding patterns that agreed with our paired cysteine mutagenesis results. In conclusion, these data suggest the same pattern of disulfide bonding as that predicted previously by NMR and thus support a consistent pattern of amino-terminal disulfide bonds in class B G protein-coupled receptors. PMID- 15664985 TI - Gating-enhanced accessibility of hydrophobic sites within the transmembrane region of the nicotinic acetylcholine receptor's {delta}-subunit. A time-resolved photolabeling study. AB - General anesthetics often interact more strongly with sites on open than on closed states of ligand-gated ion channels. To seek such sites, Torpedo membranes enriched in nicotinic acetylcholine receptors (nAChRs) were preincubated with the hydrophobic probe 3-(trifluoromethyl)-3-(m-iodophenyl) diazirine ([125I]TID) and exposed to agonist for either 0 ms (closed state), 1.5 and 10 ms (activated states), 1 s (fast desensitized state), or > or =1 h (equilibrium or slow desensitized state) and then rapidly frozen (<1 ms) and photolabeled. Within 1.5 ms, the fractional change in photoincorporation relative to the closed state decreased to 0.7 in the beta- and gamma-subunits, whereas in the alpha-subunit, it changed little. The most dramatic change occurred in the delta-subunit, where it increased to 1.6 within 10 ms but fell to 0.7 during fast desensitization. Four residues in the delta-subunit's transmembrane domain accounted for the enhanced photoincorporation induced by a 10-ms agonist exposure both when TID was added simultaneously with agonist and when it was preincubated with membranes. In the published closed state structure, two residues (deltaThr274 and deltaLeu278) are situated toward the extracellular end of helix M2, both contralateral to the ion channel and adjacent to the third residue (deltaPhe232) on M1. The fourth labeled residue (deltaIle288) is toward the end of the M2-M3 loop. Contact with these residues occurs on the time scale of a rapid phase of TID inhibition in Torpedo nAChRs, suggesting the formation of a transient hydrophobic pocket between M1, M2, and M3 in the delta-subunit during gating. PMID- 15664986 TI - Transcriptional regulation of the Grp78 promoter by endoplasmic reticulum stress: role of TFII-I and its tyrosine phosphorylation. AB - TFII-I is a signal-induced multi-functional transcription factor that has recently been implicated as a regulatory component of the endoplasmic reticulum (ER) stress response. TFII-I acts through ER stress-induced binding to the ER stress element, which is highly conserved in promoters of ER stress-inducible genes such as Grp78/BiP. Interestingly, its tyrosine phosphorylation sites are required for its activation of the Grp78 promoter. Toward understanding the link between TFII-I, the tyrosine kinase signaling pathway, and Grp78 activation, we discovered that Tg stress induces a dramatic increase of TFII-I phosphorylation at Tyr248 and localization of this form of TFII-I to the nucleus. Chromatin immunoprecipitation analysis further reveals enhanced TFII-I binding to the Grp78 promoter in vivo upon ER stress. Previously, we reported that genistein, a general inhibitor of tyrosine kinase, could suppress ER stress induction of Grp78 by inhibiting complex formation on the ER stress element; however, the mechanism is not known. Consistent with TFII-I being a target of genistein suppression, we observed that genistein could suppress Tg stress-induced phosphorylation of TFII I. We further demonstrate that c-Src, which is one of kinases identified to mediate phosphorylation of TFII-I at Tyr248, is activated by Tg stress and is able to stimulate the Grp78 promoter activity. Lastly, using stable cell lines with suppressed TFII-I levels, we show that TFII-I is required for optimal induction of Grp78 by ER stress. Our studies provide a molecular link that connects the c-Src tyrosine kinase transduction pathway to ER stress-induced transcriptional activation of Grp78 mediated by TFII-I. PMID- 15664987 TI - Substrate specificity, localization, and essential role of the glutathione peroxidase-type tryparedoxin peroxidases in Trypanosoma brucei. AB - Trypanosoma brucei, the causative agent of African sleeping sickness, encodes three nearly identical cysteine homologues of the classical selenocysteine containing glutathione peroxidases. Although one of the sequences, peroxidase III, carries both putative mitochondrial and glycosomal targeting signals, the proteins are detectable only in the cytosol and mitochondrion of mammalian bloodstream and insect procyclic T. brucei. The enzyme is a trypanothione/tryparedoxin peroxidase as are the 2 Cys-peroxiredoxins of the parasite. Hydrogen peroxide, thymine hydroperoxide, and linoleic acid hydroperoxide are reduced with second order rate constants of 8.7 x 10(4), 7.6 x 10(4), and 4 x 10(4) m(-1) s(-1), respectively, and represent probable physiological substrates. Phosphatidylcholine hydroperoxide is a very weak substrate and, in the absence of Triton X-100, even an inhibitor of the enzyme. The substrate preference clearly contrasts with that of the closely related T. cruzi enzyme, which reduces phosphatidylcholine hydroperoxides but not H(2)O(2). RNA interference causes severe growth defects in bloodstream and procyclic cells in accordance with the peroxidases being essential in both developmental stages. Thus, the cellular functions of the glutathione peroxidase-type enzymes cannot be taken over by the 2 Cys-peroxiredoxins that also occur in the cytosol and mitochondrion of the parasite. PMID- 15664988 TI - Latent S49P neuroserpin forms polymers in the dementia familial encephalopathy with neuroserpin inclusion bodies. AB - The serpinopathies result from conformational transitions in members of the serine proteinase inhibitor superfamily with aberrant tissue deposition or loss of function. They are typified by mutants of neuroserpin that are retained within the endoplasmic reticulum of neurons as ordered polymers in association with dementia. We show here that the S49P mutant of neuroserpin that causes the dementia familial encephalopathy with neuroserpin inclusion bodies (FENIB) forms a latent species in vitro and in vivo in addition to the formation of polymers. Latent neuroserpin is thermostable and inactive as a proteinase inhibitor, but activity can be restored by refolding. Strikingly, latent S49P neuroserpin is unlike any other latent serine proteinase inhibitor (serpin) in that it spontaneously forms polymers under physiological conditions. These data provide an alternative method for the inactivation of mutant neuroserpin as a proteinase inhibitor in FENIB and demonstrate a second pathway for the formation of intracellular polymers in association with disease. PMID- 15664990 TI - Critical and distinct roles of amino- and carboxyl-terminal sequences in regulation of the biological activity of the Chp atypical Rho GTPase. AB - Chp (Cdc42 homologous protein) shares significant sequence and functional identity with the human Cdc42 small GTPase, and like Cdc42, promotes formation of filopodia and activates the p21-activated kinase serine/threonine kinase. However, unlike Cdc42, Chp contains unique amino- and carboxyl-terminal extensions. Here we determined whether Chp, like Cdc42, can promote growth transformation and evaluated the role of the amino- and carboxyl-terminal sequences in Chp function. Surprisingly, we found that a GTPase-deficient mutant of Chp exhibited low transforming activity but that deletion of the amino terminus of Chp greatly enhanced its transforming activity. Thus, the amino terminus may serve as a negative regulator of Chp function. The carboxyl terminus of Cdc42 contains a CAAX (where C is cysteine, A is aliphatic amino acid, X is terminal amino acid) tetrapeptide sequence that signals for the posttranslational modification critical for Cdc42 membrane association and biological function. Although Chp lacks aCAAXmotif, we found that Chp showed carboxyl terminus dependent localization to the plasma membrane and to endosomes. Furthermore, an intact carboxyl terminus was required for Chp transforming activity. However, treatment with inhibitors of protein palmitoylation, but not prenylation, caused Chp to mislocalize to the cytoplasm. Thus, Chp depends on palmitoylation, rather than isoprenylation, for membrane association and function. In summary, Chp is implicated in cell transformation, and the unique amino and carboxyl termini of Chp represent atypical mechanisms of regulation of Rho GTPase function. PMID- 15664989 TI - Co-chaperone CHIP associates with expanded polyglutamine protein and promotes their degradation by proteasomes. AB - A major hallmark of the polyglutamine diseases is the formation of neuronal intranuclear inclusions of the disease proteins that are ubiquitinated and often associated with various chaperones and proteasome components. But, how the polyglutamine proteins are ubiquitinated and degraded by the proteasomes are not known. Here, we demonstrate that CHIP (C terminus of Hsp70-interacting protein) co-immunoprecipitates with the polyglutamine-expanded huntingtin or ataxin-3 and associates with their aggregates. Transient overexpression of CHIP increases the ubiquitination and the rate of degradation of polyglutamine-expanded huntingtin or ataxin-3. Finally, we show that overexpression of CHIP suppresses the aggregation and cell death mediated by expanded polyglutamine proteins and the suppressive effect is more prominent when CHIP is overexpressed along with Hsc70. PMID- 15664991 TI - Monooxygenation of an aromatic ring by F43W/H64D/V68I myoglobin mutant and hydrogen peroxide. Myoglobin mutants as a model for P450 hydroxylation chemistry. AB - Myoglobin (Mb) is used as a model system for other heme proteins and the reactions they catalyze. The latest novel function to be proposed for myoglobin is a P450 type hydroxylation activity of aromatic carbons (Watanabe, Y., and Ueno, T. (2003) Bull. Chem. Soc. Jpn. 76, 1309-1322). Because Mb does not contain a specific substrate binding site for aromatic compounds near the heme, an engineered tryptophan in the heme pocket was used to model P450 hydroxylation of aromatic compounds. The monooxygenation product was not previously isolated because of rapid subsequent oxidation steps (Hara, I., Ueno, T., Ozaki, S., Itoh, S., Lee, K., Ueyama, N., and Watanabe, Y. (2001) J. Biol. Chem. 276, 36067 36070). In this work, a Mb variant (F43W/H64D/V68I) is used to characterize the monooxygenated intermediate. A modified (+16 Da) species forms upon the addition of 1 eq of H2O2. This product was digested with chymotrypsin, and the modified peptide fragments were isolated and characterized as 6-hydroxytryptophan using matrix-assisted laser desorption ionization time-of-flight tandem mass spectroscopy and 1H NMR. This engineered Mb variant represents the first enzyme to preferentially hydroxylate the indole side chain of Trp at the C6 position. Finally, heme extraction was used to demonstrate that both the formation of the 6 hydroxytryptophan intermediate (+16 Da) and subsequent oxidation to form the +30 Da final product are catalyzed by the heme cofactor, most probably via the compound I intermediate. These results provide insight into the mechanism of hydroxylation of aromatic carbons by heme proteins, demonstrating that non thiolate-ligated heme enzymes can perform this function. This establishes Mb compound I as a model for P450 type aromatic hydroxylation chemistry. PMID- 15664993 TI - The N-terminal peptide of the Kaposi's sarcoma-associated herpesvirus (KSHV) cyclin determines substrate specificity. AB - Cyclin-dependent kinases (Cdks) are activated by cyclin binding and phosphorylation by the Cdk-activating kinase (CAK). Activation of Cdk6 by the D type cyclins requires phosphorylation of Cdk6 by CAK on threonine 177. In contrast, Cdk6 is activated by the Kaposi's sarcoma-associated herpesvirus (KSHV) cyclin in the absence and presence of CAK phosphorylation. The activity of Cdk6.KSHV-cyclin complexes was investigated here by analyzing mutants of the KSHV cyclin and Cdk6 in vitro as well as in U2OS cells. Deletion of the N terminus of the KSHV-cyclin affects the substrate specificity indicating that the N terminus is required for phosphorylation of histone H1 but not for other substrates. Mutation of residues in the region 180-200 of the KSHV-cyclin decreases the binding affinity to Cdk6 in U2OS cells but increases the activity of Cdk6.KSHV cyclin complexes in vitro indicating that low affinity binding of cyclins to the Cdk subunit might favor increased on- or off-rates of Cdk substrates. Expression of high levels of p16(INK4a) in cells leads to the formation of a heterotrimeric complex composed of Cdk6, KSHV-cyclin, and p16(INK4a). Some of the Cdk6 .KSHV cyclin.p16 complexes were found to be active indicating that there might be different modes of p16 binding to Cdk6.cyclin complexes. PMID- 15664992 TI - The novel Drosophila lysosomal enzyme receptor protein mediates lysosomal sorting in mammalian cells and binds mammalian and Drosophila GGA adaptors. AB - Biogenesis of lysosomes depends in mammalian cells on the specific recognition and targeting of mannose 6-phosphate-containing lysosomal enzymes by two mannose 6-phosphate receptors (MPR46, MPR300), key components of the extensively studied receptor-mediated lysosomal sorting system in complex metazoans. In contrast, the biogenesis of lysosomes is poorly investigated in the less complex metazoan Drosophila melanogaster. We identified the novel type I transmembrane protein lysosomal enzyme receptor protein (LERP) with partial homology to the mammalian MPR300 encoded by Drosophila gene CG31072. LERP contains 5 lumenal repeats that share homology to the 15 lumenal repeats found in all identified MPR300. Four of the repeats display the P-lectin type pattern of conserved cysteine residues. However, the arginine residues identified to be essential for mannose 6-phosphate binding are not conserved. The recombinant LERP protein was expressed in mammalian cells and displayed an intracellular localization pattern similar to the mammalian MPR300. The LERP cytoplasmic domain shows highly conserved interactions with Drosophila and mammalian GGA adaptors known to mediate Golgi endosome traffic of MPRs and other transmembrane cargo. Moreover, LERP rescues missorting of soluble lysosomal enzymes in MPR-deficient cells, giving strong evidence for a function that is equivalent to the mammalian counterpart. However, unlike the mammalian MPRs, LERP did not bind to the multimeric mannose 6 phosphate ligand phosphomannan. Thus ligand recognition by LERP does not depend on mannose 6-phosphate but may depend on a common feature present in mammalian lysosomal enzymes. Our data establish a potential important role for LERP in biogenesis of Drosophila lysosomes and suggest a GGA function also in the receptor-mediated lysosomal transport system in the fruit fly. PMID- 15664994 TI - Signal transducers and activators of transcription 3 augments the transcriptional activity of CCAAT/enhancer-binding protein alpha in granulocyte colony stimulating factor signaling pathway. AB - The Janus kinase (Jak)-Stat pathway plays an essential role in cytokine signaling. Granulocyte colony-stimulating factor (G-CSF) promotes granulopoiesis and granulocytic differentiation, and Stat3 is the principle Stat protein activated by G-CSF. Upon treatment with G-CSF, the interleukin-3-dependent cell line 32D clone 3(32Dcl3) differentiates into neutrophils, and 32Dcl3 cells expressing dominant-negative Stat3 (32Dcl3/DNStat3) proliferate in G-CSF without differentiation. Gene expression profile and quantitative PCR analysis of G-CSF stimulated cell lines revealed that the expression of C/EBPalpha was up-regulated by the activation of Stat3. In addition, activated Stat3 bound to CCAAT/enhancer binding protein (C/EBP)alpha, leading to the enhancement of the transcription activity of C/EBPalpha. Conditional expression of C/EBPalpha in 32Dcl3/DNStat3 cells after G-CSF stimulation abolishes the G-CSF-dependent cell proliferation and induces granulocytic differentiation. Although granulocyte-specific genes, such as the G-CSF receptor, lysozyme M, and neutrophil gelatinase-associated lipocalin precursor (NGAL) are regulated by Stat3, only NGAL was induced by the restoration of C/EBPalpha after stimulation with G-CSF in 32Dcl3/DNStat3 cells. These results show that one of the major roles of Stat3 in the G-CSF signaling pathway is to augment the function of C/EBPalpha, which is essential for myeloid differentiation. Additionally, cooperation of C/EBPalpha with other Stat3 activated proteins are required for the induction of some G-CSF responsive genes including lysozyme M and the G-CSF receptor. PMID- 15664995 TI - Regulation of the transcriptional activity of the IRF7 promoter by a pathway independent of interferon signaling. AB - Genes containing an interferon (IFN)-stimulated response element (ISRE) can be divided into two groups according to their inducibility by IFN and virus infection: one induced only by IFN and the other induced by both IFN and virus infection. Although it is now clear that IFN regulatory factor 7 (IRF7) is a multifunctional gene essential for induction of type I IFNs, regulation of the IRF7 promoter (IRF7p) is poorly understood. The IRF7 gene includes two IFN responsive elements, an IRF-binding element (IRFE) in the promoter region and an ISRE in the first intron, and is induced by the IFN-triggered Jak-STAT pathway by binding of the IFN-stimulated gene factor 3 (ISGF3) complex to the ISRE. In this study, we demonstrate that IRF3 and IRF7, which with the coactivators CREB binding protein and P300 form the virus-activated factor (VAF) complex upon Sendai virus infection, bind to the IRF7 ISRE and IRFE and can directly activate IRF7 transcription. Promoter reporter assays show that both the ISRE and IRFE are responsive to activation by IRF7 and IRF3. In cells transiently expressing IRF7 or/and IRF3, the VAF level and binding of VAF are clearly increased after Sendai virus infection. Studies with Jak1 kinase inactive 293 cells that were stably transfected with a Jak1 kinase dead dominant negative construct, and the mutant cell lines SAN (IFNalpha-/beta-), U2A (IRF9-), U4A (Jak1-), and DKO (IRF1-/IRF2-) show that the IRF7 transcription activated directly by VAF is distinct from and independent of the IFN signaling pathway. Thus, IRF7 transcription is autoregulated by binding of the IRF7-containing VAF to its own ISRE and IRFE. The results show two distinct mechanisms for the activation of the IRF7 promoter, by IFN and by virus infection. A regulatory network between type I IFNs and IRF7 is proposed. The distinct pathways may reflect special roles for an efficient antiviral response at different stages of virus infection. PMID- 15664996 TI - Direct role of ChREBP.Mlx in regulating hepatic glucose-responsive genes. AB - Enzymes required for de novo lipogenesis are induced in mammalian liver after a meal high in carbohydrates. In addition to insulin, increased glucose metabolism initiates an intracellular signaling pathway that transcriptionally regulates genes encoding lipogenic enzymes. A cis-acting sequence, the carbohydrate response element (ChoRE), has been found in the promoter region of several of these genes. ChREBP (carbohydrate response element-binding protein) was recently identified as a candidate transcription factor in the glucose-signaling pathway. We reported that ChREBP requires the heterodimeric partner Max-like factor X (Mlx) to bind to ChoRE sequences. In this study we provide further evidence to support a direct role of Mlx in glucose signaling in the liver. We constructed two different dominant negative forms of Mlx that could dimerize with ChREBP but block its binding to DNA. When introduced into hepatocytes, both dominant negative forms of Mlx inhibited the glucose response of a transfected ChoRE containing promoter. The glucose response was rescued by adding exogenous wild type Mlx or ChREBP, but not MondoA, a paralog of ChREBP that can also form a heterodimer with Mlx. Furthermore, dominant negative Mlx blocked the induction of glucose-responsive genes from their natural chromosomal context under high glucose conditions. In contrast, genes induced by the insulin and thyroid hormone signaling pathways were unaffected by dominant negative Mlx. Mlx was present in the glucose-responsive complex of liver nuclear extract from which ChREBP was purified. In conclusion, Mlx is an obligatory partner of ChREBP in regulating lipogenic enzyme genes in liver. PMID- 15664997 TI - A crucial role of MafA as a novel therapeutic target for diabetes. AB - MafA, a recently isolated pancreatic beta-cell-specific transcription factor, is a potent activator of insulin gene transcription. In this study, we show that MafA overexpression, together with PDX-1 (pancreatic and duodenal homeobox factor 1) and NeuroD, markedly increases insulin gene expression in the liver. Consequently, substantial amounts of insulin protein were induced by such combination. Furthermore, in streptozotocin-induced diabetic mice, MafA overexpression in the liver, together with PDX-1 and NeuroD, dramatically ameliorated glucose tolerance, while combination of PDX-1 and NeuroD was much less effective. These results suggest a crucial role of MafA as a novel therapeutic target for diabetes. PMID- 15664998 TI - Role of Kruppel-like factor 15 (KLF15) in transcriptional regulation of adipogenesis. AB - Kruppel-like zinc finger transcription factors (KLFs) play diverse roles during cell differentiation and development in mammals. We have now shown by microarray analysis that expression of the KLF15 gene is markedly up-regulated during the differentiation of 3T3-L1 preadipocytes into adipocytes. Inhibition of the function of KLF15, either by expression of a dominant negative mutant or by RNA interference, both reduced the expression of peroxisome proliferator-activated receptor gamma (PPARgamma) and blocked adipogenesis in 3T3-L1 preadipocytes exposed to inducers of adipocyte differentiation. However, the dominant negative mutant of KLF15 did not affect the expression of CCAAT/enhancer-binding protein beta (C/EBPbeta) elicited by inducers of differentiation in 3T3-L1 preadipocytes. In addition, ectopic expression of KLF15 in NIH 3T3 or C2C12 cells triggered both lipid accumulation and the expression of PPARgamma in the presence of inducers of adipocyte differentiation. Ectopic expression of C/EBPbeta, C/EBPdelta, or C/EBPalpha in NIH 3T3 cells also elicited the expression of KLF15 in the presence of inducers of adipocyte differentiation. Moreover, KLF15 and C/EBPalpha acted synergistically to increase the activity of the PPARgamma2 gene promoter in 3T3 L1 adipocytes. Our observations thus demonstrate that KLF15 plays an essential role in adipogenesis in 3T3-L1 cells through its regulation of PPAR gamma expression. PMID- 15664999 TI - Oxidative stress-mediated, post-translational loss of MafA protein as a contributing mechanism to loss of insulin gene expression in glucotoxic beta cells. AB - Glucose toxicity in pancreatic islet beta cells causes loss of insulin gene expression, content, and secretion due to loss of binding of transcription factors, most notably PDX-1 and RIPE-3b1 activator, to the promoter region of the insulin gene. Recently, RIPE-3b1 activator was cloned and identified as the mammalian homologue of avian MafA/Maf-L (MafA). This enabled us to carry out more extensive studies of the role of MafA in glucotoxicity than were hitherto possible. Northern analysis of glucotoxic HIT-T15 cells revealed normal amounts of MafA mRNA, but Western analysis demonstrated a 97 +/- 1% reduction in MafA protein (p < 0.0001). The proteasome is a likely site for MafA degradation as lactacystin, an irreversible proteasome inhibitor, caused an accumulation of MafA protein. Antioxidants have previously been shown to prevent the adverse effects of glucose toxicity on beta cell function both in vivo and in vitro. In the current study, chronic culturing of HIT-T15 cells with the antioxidant N acetylcysteine (NAC) prevented loss of MafA protein (late passage = 18.9 +/- 10.4% of early passage, p < 0.001; late passage with NAC = 68.7 +/- 19.7% of early passage, p = not significant) and loss of DNA binding (late passage = 63.7 +/- 9% of early passage, p < 0.02; late passage with NAC = 116 +/- 10% of early passage, p = not significant). Additionally, transient transfection of PDX-1 or MafA cDNA into glucotoxic cells increased PDX-1 and MafA protein levels and individually increased insulin promoter activity (untreated = 34%, PDX-1 = 70%, MafA = 78%; percentage of activity of early passage cells), whereas the combined transfection of MafA and PDX-1 completely restored insulin promoter activity. This recovery of promoter activity following transient transfection had no effect on endogenous insulin mRNA. However, adenoviral infection of MafA and PDX-1 significantly increased endogenous insulin mRNA levels by 93% (121 +/- 9 versus 233 +/- 18 density light units; n = 5, p < 0.001). We conclude that the absence of MafA protein from beta cells via chronic oxidative stress contributes importantly to the loss of endogenous insulin gene expression as glucose toxicity develops. PMID- 15665000 TI - The islet beta cell-enriched MafA activator is a key regulator of insulin gene transcription. AB - The islet-enriched MafA, PDX-1, and BETA2 activators contribute to both beta cell specific and glucose-responsive insulin gene transcription. To investigate how these factors impart activation, their combined impact upon insulin enhancer driven expression was first examined in non-beta cell line transfection assays. Individual expression of PDX-1 and BETA2 led to little or no activation, whereas MafA alone did so modestly. MafA together with PDX-1 or BETA2 produced synergistic activation, with even higher insulin promoter activity found when all three proteins were present. Stimulation was attenuated upon compromising either MafA transactivation or DNA-binding activity. MafA interacted with endogenous PDX 1 and BETA2 in coimmunoprecipitation and in vitro GST pull-down assays, suggesting that regulation involved direct binding. Dominant-negative acting and small interfering RNAs of MafA also profoundly reduced insulin promoter activity in beta cell lines. In addition, MafA was induced in parallel with insulin mRNA expression in glucose-stimulated rat islets. Insulin mRNA levels were also elevated in rat islets by adenoviral-mediated expression of MafA. Collectively, these results suggest that MafA plays a key role in coordinating and controlling the level of insulin gene expression in islet beta cells. PMID- 15665002 TI - PAR1 cleavage and signaling in response to activated protein C and thrombin. AB - Activated protein C (APC), a natural anticoagulant protease, can trigger cellular responses via protease-activated receptor-1 (PAR1), a G protein-coupled receptor for thrombin. Whether this phenomenon contributes to the physiological effects of APC is unknown. Toward answering this question, we compared the kinetics of PAR1 cleavage on endothelial cells by APC versus thrombin. APC did cleave PAR1 on the endothelial surface, and antibodies to the endothelial protein C receptor inhibited such cleavage. Importantly, however, APC was approximately 10(4)-fold less potent than thrombin in this setting. APC and thrombin both triggered PAR1 mediated responses in endothelial cells including expression of antiapoptotic (tumor necrosis factor-alpha-induced a20 and iap-1) and chemokine (interleukin-8 (il-8) and cxcl3) genes, but again, APC was approximately 10(4)-fold less potent than thrombin. The addition of zymogen protein C to endothelial cultures did not alter the rate of PAR1 cleavage at low or high concentrations of thrombin, and PAR1 cleavage was substantial at thrombin concentrations too low to trigger detectable conversion of protein C to APC. Thus, locally generated APC did not contribute to PAR1 cleavage beyond that effected by thrombin in this system. Although consistent with reports that sufficiently high concentrations of APC can cleave and activate PAR1 in culture, our data suggest that a significant physiological role for PAR1 activation by APC is unlikely. PMID- 15665001 TI - Aberrant localization of intracellular organelles, Ca2+ signaling, and exocytosis in Mist1 null mice. AB - Ca2+ signaling and exocytosis are highly polarized functions of pancreatic acinar cells. The role of cellular architecture in these activities and the capacity of animals to tolerate aberrant acinar cell function are not known. A key regulator of acinar cell polarity is Mist1, a basic helix-loop-helix transcription factor. Ca2+ signaling and amylase release were examined in pancreatic acini of wild type and Mist1 null mice to gain insight into the importance of cellular architecture for Ca2+ signaling and regulated exocytosis. Mist1-/- acinar cells exhibited dramatically altered Ca2+ signaling with up-regulation of the cholecystokinin receptor but minimal effect upon expression of the M3 receptor. However, stimulation of inositol 1,4,5-trisphosphate production by cholecystokinin and carbachol was inefficient in Mist1-/- cells. Although agonist stimulation of Mist1-/- cells evoked a Ca2+ signal, often the Ca2+ increase was not in the form of typical Ca2+ oscillations but rather in the form of a peak/plateau-type response. Mist1-/- cells also displayed distorted apical-to-basal Ca2+ waves. The aberrant Ca2+ signaling was associated with mislocalization and reduced Ca2+ uptake by the mitochondria of stimulated Mist1-/- cells. Deletion of Mist1 also led to mislocalization of the Golgi apparatus and markedly reduced digestive enzyme content. The combination of aberrant Ca2+ signaling and reduced digestive enzyme content resulted in poor secretion of digestive enzymes. Yet, food consumption and growth of Mist1-/- mice were normal for at least 32 weeks. These findings reveal that Mist1 is critical to normal organelle localization in exocrine cells and highlight the critical importance of maintaining cellular architecture and polarized localization of cellular organelles in generating a propagating apical-to-basal Ca2+ wave. The studies also reveal the spare capacity of the exocrine pancreas that allows normal growth and development in the face of compromised exocrine pancreatic function. PMID- 15665003 TI - The functional basis of mycophenolic acid resistance in Candida albicans IMP dehydrogenase. AB - Candida albicans is an important fungal pathogen of immunocompromised patients. In cell culture, C. albicans is sensitive to mycophenolic acid (MPA) and mizoribine, both natural product inhibitors of IMP dehydrogenase (IMPDH). These drugs have opposing interactions with the enzyme. MPA prevents formation of the closed enzyme conformation by binding to the same site as a mobile flap. In contrast, mizoribine monophosphate, the active metabolite of mizoribine, induces the closed conformation. Here, we report the characterization of IMPDH from wild type and MPA-resistant strains of C. albicans. The wild-type enzyme displays significant differences from human IMPDHs, suggesting that selective inhibitors that could be novel antifungal agents may be developed. IMPDH from the MPA resistant strain contains a single substitution (A251T) that is far from the MPA binding site. The A251T variant was 4-fold less sensitive to MPA as expected. This substitution did not affect the k(cat) value, but did decrease the K(m) values for both substrates, so the mutant enzyme is more catalytically efficient as measured by the value of k(cat)/K(m). These simple criteria suggest that the A251T variant would be the evolutionarily superior enzyme. However, the A251T substitution caused the enzyme to be 40-fold more sensitive to mizoribine monophosphate. This result suggests that A251T stabilizes the closed conformation, and this hypothesis is supported by further inhibitor analysis. Likewise, the MPA-resistant strain was more sensitive to mizoribine in cell culture. These observations illustrate the evolutionary challenge posed by the gauntlet of chemical warfare at the microbial level. PMID- 15665004 TI - RhoA/ROCK signaling regulates Sox9 expression and actin organization during chondrogenesis. AB - Endochondral ossification is initiated by the differentiation of mesenchymal precursor cells to chondrocytes (chondrogenesis). This process is characterized by a strong interdependence of cell shape, cytoskeletal organization, and the onset of chondrogenic gene expression, but the molecular mechanisms mediating these interactions are not known. Here we investigated the role of the RhoA/ROCK pathway, a well characterized regulator of cytoskeletal organization, in chondrogenesis. We show that pharmacological inhibition of ROCK signaling by Y27632 resulted in increased glycosaminoglycan synthesis and elevated expression of the chondrogenic transcription factor Sox9, whereas overexpression of RhoA in the chondrogenic cell line ATDC5 had the opposite effects. Suppression of Sox9 expression by ROCK signaling was achieved through repression of Sox9 promoter activity. These molecular changes were accompanied by reorganization of the actin cytoskeleton, where RhoA/ROCK signaling suppressed cortical actin organization, a hallmark of differentiated chondrocytes. This led us to analyze the regulation of Sox9 expression by drugs affecting cytoskeletal dynamics. Both inhibition of actin polymerization by cytochalasin D and stabilization of existing actin filaments by jasplakinolide resulted in increased Sox9 mRNA levels, whereas inhibition of microtubule polymerization by colchicine completely blocked Sox9 expression. In conclusion, our data suggest that RhoA/ROCK signaling suppresses chondrogenesis through the control of Sox9 expression and actin organization. PMID- 15665005 TI - Does home based medication review keep older people out of hospital? The HOMER randomised controlled trial. AB - OBJECTIVE: To determine whether home based medication review by pharmacists affects hospital readmission rates among older people. DESIGN: Randomised controlled trial. SETTING: Home based medication review after discharge from acute or community hospitals in Norfolk and Suffolk. PARTICIPANTS: 872 patients aged over 80 recruited during an emergency admission (any cause) if returning to own home or warden controlled accommodation and taking two or more drugs daily on discharge. INTERVENTION: Two home visits by a pharmacist within two weeks and eight weeks of discharge to educate patients and carers about their drugs, remove out of date drugs, inform general practitioners of drug reactions or interactions, and inform the local pharmacist if a compliance aid is needed. Control arm received usual care. MAIN OUTCOME MEASURE: Total emergency readmissions to hospital at six months. Secondary outcomes included death and quality of life measured with the EQ-5D. RESULTS: By six months 178 readmissions had occurred in the control group and 234 in the intervention group (rate ratio = 1.30, 95% confidence interval 1.07 to 1.58; P = 0.009, Poisson model). 49 deaths occurred in the intervention group compared with 63 in the control group (hazard ratio = 0.75, 0.52 to 1.10; P = 0.14). EQ-5D scores decreased (worsened) by a mean of 0.14 in the control group and 0.13 in the intervention group (difference = 0.01, -0.05 to 0.06; P = 0.84, t test). CONCLUSIONS: The intervention was associated with a significantly higher rate of hospital admissions and did not significantly improve quality of life or reduce deaths. Further research is needed to explain this counterintuitive finding and to identify more effective methods of medication review. PMID- 15665006 TI - Access to catheterisation facilities in patients admitted with acute coronary syndrome: multinational registry study. AB - OBJECTIVE: To investigate the relation between access to a cardiac catheterisation laboratory and clinical outcomes in patients admitted to hospital with suspected acute coronary syndrome. DESIGN: Prospective, multinational, observational registry. SETTING: Patients enrolled in 106 hospitals in 14 countries between April 1999 and March 2003. PARTICIPANTS: 28,825 patients aged > or = 18 years. MAIN OUTCOME MEASURES: Use of percutaneous coronary intervention or coronary artery bypass graft surgery, death, infarction after discharge, stroke, or major bleeding. RESULTS: Most patients (77%) across all regions (United States, Europe, Argentina and Brazil, Australia, New Zealand, and Canada) were admitted to hospitals with catheterisation facilities. As expected, the availability of a catheterisation laboratory was associated with more frequent use of percutaneous coronary intervention (41% v 3.9%, P < 0.001) and coronary artery bypass graft (7.1% v 0.7%, P < 0.001). After adjustment for baseline characteristics, medical history, and geographical region there were no significant differences in the risk of early death between patients in hospitals with or without catheterisation facilities (odds ratio 1.13, 95% confidence interval 0.98 to 1.30, for death in hospital; hazard ratio 1.05, 0.93 to 1.18, for death at 30 days). The risk of death at six months was significantly higher in patients first admitted to hospitals with catheterisation facilities (hazard ratio 1.14, 1.03 to 1.26), as was the risk of bleeding complications in hospital (odds ratio 1.94, 1.57 to 2.39) and stroke (odds ratio 1.53, 1.10 to 2.14). CONCLUSIONS: These findings support the current strategy of directing patients with suspected acute coronary syndrome to the nearest hospital with acute care facilities, irrespective of the availability of a catheterisation laboratory, and argue against early routine transfer of these patients to tertiary care hospitals with interventional facilities. PMID- 15665007 TI - Mosaic status in lymphocytes of infertile men with or without Klinefelter syndrome. AB - BACKGROUND: Gonosomal aneuploidies such as Klinefelter syndrome (47,XXY) are the most frequent chromosomal aberration in infertile men. Normally the chromosomal status of patients is detected by karyotyping of up to 20 metaphase spreads of lymphocyte nuclei, whereby low grade mosaicism may be overlooked. To test whether Klinefelter patients with 47,XXY karyotype or infertile men with 46,XY karyotype represent gonosomal mosaicisms, we performed meta- and interphase fluorescence in situ hybridization (FISH) on 45 men. METHODS AND RESULTS: A total of 400 interphase and 40 metaphase lymphocyte nuclei per patient were scored after hybridization with DNA probes specific for chromosomes X and Y, and chromosome 9 as a control. On the basis of conventional karyotype, hormone levels and clinical appearance, patients were subdivided into 18 Klinefelter syndrome patients with 47,XXY (group I), 11 Klinefelter syndrome-like patients with normal karyotype, 46,XY (group II) and six non-Klinefelter-like infertile patients with normal 46,XY karyotype (group III). Ten normal men (group IV) served as controls. Testicular volume in the Klinefelter group I was smaller compared with group II (P = 0.016), group III (P < 0.001) and group IV (P < 0.001). In addition, testicular volumes in group II were lower compared with group III and group IV (P < 0.004). No significant differences between the aneuploidy rate analysed by FISH in interphase nuclei and metaphases were found in either single patients or groups. Patients with Klinefelter syndrome, 47,XXY (group I) or with symptoms similar to those in Klinefelter patients 46,XY (group II) showed a similar aneuploidy rate (group I 7.1 +/- 4.0% and group II 4.6 +/- 3.4%) and two 47,XXY patients with a high prevalence for normal 46,XY lymphocytes had sperm in their ejaculate. However, in general, no correlations between FISH mosaic status and serum hormone parameters, nor with ejaculate parameters were found. CONCLUSIONS: The results suggest that 47,XXY patients with an increased incidence of XY cells (average of 4.2 +/- 2.3) may have a higher probability of germ cells as we found sperm only in the ejaculate of Klinefelter syndrome patients with mosaic 46,XY cells (6.0 and 7.0%). On the other hand, 46,XY patients with mosaic sex chromosome aneuploidies detected by FISH analysis more often show symptoms of hypogonadism phenotypically resembling Klinefelter syndrome. PMID- 15665008 TI - Single embryo transfer: a mini-review. AB - This paper provides a concise review of single embryo transfer (SET) in cycles using fresh embryos as well as in cycles using frozen-thawed embryos. Relevant studies were identified by a computerized search in PubMed for the period 1995 2004. The pregnancy rates, delivery rates and multiple pregnancy/birth rates were evaluated after fresh or frozen embryo transfer as well as cumulative delivery rates after fresh and frozen SET. The results of four randomized controlled trials (RCT) and seven observational studies using fresh embryo transfers are analysed. No RCT with SET in freezing-thawing cycles was identified, while one observational study was identified. The effects of a change in the rules from the National Board of Health and Welfare in Sweden in 2003 regarding the implementation of SET in Sweden are summarized. PMID- 15665009 TI - Volume changes of mature human oocytes on exposure to cryoprotectant solutions used in slow cooling procedures. AB - BACKGROUND: Despite the recent increase in pregnancies from cryopreserved human oocytes, success in terms of births per thawed oocyte is still poor. Modifications to cryopreservation protocols have not been based on measurement of the osmotic response of oocytes, and methodologies are often poorly described or protocols not strictly adhered to, inevitably resulting in variability. METHODS: Volume change of mature human oocytes was measured on exposure to cryoprotectant. Oocytes were exposed to either 0.75 mol/l propane-1,2-diol (PrOH) for 10 min; 1.5 mol/l PrOH for 10 min, having been exposed to 0.75 mol/l PrOH for 7.5 min; or 1.5 mol/l PrOH plus 0.2 or 0.3 mol/l sucrose for 10 min, having been exposed to 1.5 mol/l PrOH for 10 min. RESULTS: On exposure to PrOH alone, oocytes shrank and then re-expanded, having reached 75 and 84% of their starting volume in 0.75 and 1.5 mol/l, respectively. Oocytes shrank continuously in PrOH plus sucrose, reaching 67 or 55% of their initial volume in 0.2 or 0.3 mol/l sucrose, respectively. CONCLUSIONS: To improve consistency following cryopreservation, protocols must be strictly adhered to; small changes in duration of exposure to cryoprotectant can result in drastic changes in cellular hydration and thus the fate of the cell during freezing/thawing. PMID- 15665010 TI - Recombinant luteinizing hormone supplementation to recombinant follicle stimulating hormone induced ovarian hyperstimulation in the GnRH-antagonist multiple-dose protocol. AB - BACKGROUND: Suppression of endogenous LH production by mid-follicular phase GnRH antagonist administration in controlled ovarian hyperstimulation protocol using recombinant (rec) FSH preparations void of LH activity may potentially affect ovarian response and the outcome of IVF treatment. The present study prospectively assessed the effect of using a combination of recFSH and recLH on ovarian stimulation parameters and treatment outcome in a fixed GnRH-antagonist multiple dose protocol. METHODS: 127 infertile patients with an indication for IVF or ICSI were recruited and randomized (using sealed envelopes) to receive a starting dose of either 150 IU recFSH (follitropin alpha) or 150 IU recFSH plus 75 IU recLH (lutropin alpha) for ovarian hyperstimulation. GnRH-antagonist (Cetrorelix) 0.25 mg was administered daily from stimulation day 6 onwards up to and including the day of the administration of recombinant HCG (chorion gonadotropin alpha). Gonadotropin dose adjustments were allowed from stimulation day 6 onwards, HCG was administered as soon as three follicles > or =18 mm were present. The primary outcome parameter was treatment duration until administration of HCG. RESULTS: Exogenous LH did not shorten the time necessary to reach ovulation induction criteria. Serum estradiol (E(2)) and LH levels were significantly higher on the day of HCG administration in the recLH-supplemented group (1924.7 +/- 1256.4 vs 1488.3 +/- 824.0 pg/ml, P < 0.03), and 2.1 +/- 1.4 vs 1.4 +/- 1.5 IU/l, P < 0.01, respectively). CONCLUSIONS: Except for higher E(2) and LH levels on the day of HCG administration, no positive trend in favour of additional LH was found as defined by treatment outcome parameters. PMID- 15665011 TI - Stress and outcome success in IVF: the role of self-reports and endocrine variables. AB - BACKGROUND: The aim of this study was to examine the associations between urinary levels of the stress hormones adrenaline, noradrenaline and cortisol during treatment with self reported stress, in order to investigate the mechanism for the previously observed negative association of anxiety and depression with the outcome of IVF/ICSI. METHODS: In a multicentre prospective cohort study, women entering their first cycle of IVF/ICSI treatment were asked to participate. From each participant nocturnal urine samples were collected; pre-treatment, before oocyte retrieval and before embryo-transfer (ET), to assess hormonal concentrations. Additionally, two questionnaires were administered before the start of the treatment to measure anxiety and depression. RESULTS: 168 women completed the questionnaires and collected at least two urine specimens. A significant positive correlation between urinary adrenaline concentrations at baseline and ET and the scores on depression at baseline were found. In women with successful treatment, lower concentrations of adrenaline at oocyte retrieval and lower concentrations of adrenaline and noradrenaline at ET, compared with unsuccessful women, were found. CONCLUSIONS: The significant positive association of adrenaline concentration with pregnancy and with depression suggested that this adrenal hormone could be one of the links in the complex relationship between psychosocial stress and outcome after IVF/ICSI. PMID- 15665012 TI - The effects of 1-month administration of asoprisnil (J867), a selective progesterone receptor modulator, in healthy premenopausal women. AB - BACKGROUND: Asoprisnil (J867) is a novel selective progesterone receptor modulator (SPRM) that exhibits partial agonist and antagonist activities and tissue selective effects. This double-blind, dose-escalation study was conducted to evaluate the effects of asoprisnil in 60 regularly cycling premenopausal women. METHODS: Asoprisnil or placebo was administered orally for 28 days starting at the beginning of the menstrual cycle in doses of 5 mg once daily (QD), 5 mg twice daily (BID), 10 mg QD, 25 mg QD, 25 mg BID and 50 mg BID. Within each dose group, two women were randomized to placebo and eight to asoprisnil. Progesterone concentrations indicative of luteinization were defined as at least one progesterone measurement during the luteal phase exceeding 3.5 ng/ml. RESULTS: Asoprisnil consistently prolonged the menstrual cycle at doses > or = 10 mg QD. However, the effects on luteal phase progesterone indicative of luteinization were inconsistent and lacked dose dependency. Asoprisnil suppressed periovulatory estradiol but not below follicular phase levels. No significant changes were observed in cortisol and prolactin. Asoprisnil was well tolerated. CONCLUSIONS: Asoprisnil reversibly suppressed menstruation at doses > or = 10 mg QD irrespective of the effect on luteal phase progesterone concentrations indicative of luteinization. It induces amenorrhea primarily by targeting the endometrium in the absence of estrogen deprivation. PMID- 15665013 TI - Sequential FISH analysis using competitive displacement of labelled peptide nucleic acid probes for eight chromosomes in human blastomeres. AB - BACKGROUND: The aim was to introduce a new strategy based on peptide nucleic acid (PNA) probes and competitive displacement for using fluorescence in-situ hybridization (FISH) analysis on human blastomeres. METHODS: Sequential FISH analysis with PNA probes and competitive displacement was performed using three different probe sets. The first set consisted of labelled probe only. The second and third sets included labelled as well as unlabelled probe, corresponding to the labelled probes in the previous cycles. The probes for enumeration were for chromosome 1, 13, 16, 17, 18, 21, X and Y. RESULTS: The performance of PNA probes was similar to the established DNA probes. The strategy of competitive displacement resulted in a destabilization of already bound probe before the next FISH cycle at only 50 degrees C, which allowed for up to five sequential FISH cycles without loss of signal. CONCLUSIONS: PNA probes are a good alternative to DNA probes in the present set-up, since the low temperature required both for binding and destabilization of PNA probes minimizes the loss of signal, and several FISH cycles can therefore be carried out before FISH errors occur. PMID- 15665014 TI - A novel organotypic culture model for normal human endometrium: regulation of epithelial cell proliferation by estradiol and medroxyprogesterone acetate. AB - BACKGROUND: A novel organotypic culture system was established for modelling the hormonal responses of the normal human endometrium in vitro. METHODS: Endometrial epithelial cells were cultured as glandular organoids within reconstituted extracellular matrix (Matrigel) in tissue culture inserts and stromal cells on plastic below the epithelial compartment. The effects of estradiol (E2) and E2 together with medroxyprogesterone acetate (MPA) on cell proliferation and the expression of estrogen receptor alpha (ERalpha) and progesterone receptor (PR) were studied in 10 epithelial-stromal co-cultures and in three parallel monocultures of epithelial organoids. RESULTS: In co-cultures, E2 was shown to increase the percentage of Ki67-positive cells by approximately 2-fold relative to untreated controls. In the presence of MPA, a significant decrease in cell proliferation was detected. Similar results were obtained when the corresponding percentages of Ki67-positive organoids were calculated instead of individual cells. In the absence of stromal fibroblasts, Ki67 epithelial labelling remained below the control value after both hormonal treatments. Epithelial organoids retained their capacity to express estrogen and progesterone receptors in culture. E2 was shown to markedly increase and MPA to down-regulate the expression of PR. The expression of ERalpha was only slightly affected by either hormonal treatment. CONCLUSIONS: The present organotypic model provides a novel in vitro system in which to study the effects of steroids in the normal human endometrium both in terms of cell proliferation and gene expression. The culture system holds promise as a useful method to screen novel steroid compounds and may help to circumvent problems related to the use of animal models. PMID- 15665016 TI - Susceptibility to ovarian endometriosis in Polish population is not associated with HLA-DRB1 alleles. AB - BACKGROUND: Endometriosis is associated with inflammatory autoimmune reactions; however, aetiopathogenesis of the disease is still poorly understood. While autoimmune disorders are often associated with particular HLA alleles, the possible involvement of HLA in the aetiopathogenesis of endometriosis is still a subject of controversy. The aim of the study was to examine the distribution of HLA-DRB1 alleles in women with endometriosis. To ensure homogeneity of the studied group, only women with ovarian endometrial cysts were included. METHODS: The study included 65 Polish patients of Caucasian origin in whom ovarian endometriosis had been confirmed by laparoscopic and histopathological examinations. HLA-DRB1 alleles were typed using a reverse slot blot method. A frequency of particular HLA-DRB1 alleles in patients was compared with that of a control group of 700 unrelated ethnically matched individuals as well as 193 age matched women without endometriosis. RESULTS: No statistically significant differences were found in the distribution of HLA-DRB1 alleles in patients with ovarian endometriosis as compared with control populations. CONCLUSIONS: The results of the present study show that ovarian endometriosis is not associated with particular HLA-DRB1 allele(s). This may suggest that aetiology of this form of endometriosis may be not primarily associated with class II HLA-mediated autoimmune reactions. PMID- 15665015 TI - Basal and stimulation day 5 anti-Mullerian hormone serum concentrations as predictors of ovarian response and pregnancy in assisted reproductive technology cycles stimulated with gonadotropin-releasing hormone agonist--gonadotropin treatment. AB - BACKGROUND: Anti-Mullerian hormone (AMH) has been recently proposed as a marker for ovarian ageing and poor ovarian response to controlled ovarian hyperstimulation in assisted reproduction cycles. The present study was undertaken to investigate the usefulness of baseline cycle day 3 AMH levels and AMH serum concentrations obtained on the fifth day of gonadotropin therapy in predicting ovarian response and pregnancy in women undergoing ovarian stimulation with FSH under pituitary desensitization for assisted reproduction. METHODS: A total of 80 women undergoing their first cycle of IVF/intracytoplasmic sperm injection (ICSI) treatment were studied. Twenty consecutive cycles which were cancelled because of a poor follicular response were initially selected. As a control group, 60 women were randomly selected from our assisted reproduction programme matching by race, age, body mass index, basal FSH and indication for IVF/ICSI to those in the cancelled group. For each cancelled patient, three IVF/ICSI women who met the matching criteria were included. RESULTS: Basal and day 5 AMH serum concentrations were significantly lower in the cancelled than in the control group. Receiver-operating characteristic (ROC) analysis showed that the capacity of day 5 AMH in predicting the likelihood of cancellation in an assisted reproduction treatment programme was significantly higher than that for basal AMH measurement. However, the predictive capacity of day 5 AMH was not better than that provided by day 5 estradiol. In addition, neither basal nor day 5 AMH or estradiol measurements were useful in the prediction of pregnancy after assisted reproductive treatment. CONCLUSIONS: AMH concentrations obtained early in the follicular phase during ovarian stimulation under pituitary suppression for assisted reproduction are better predictors of ovarian response than basal AMH measurements. However, AMH is not useful in the prediction of pregnancy. Definite clinical applicability of AMH determination as a marker of IVF outcome remains to be established. PMID- 15665017 TI - Imprinting diseases and IVF: Danish National IVF cohort study. AB - BACKGROUND: The aim of this study was to compare the frequency of imprinting diseases in children born after IVF with the incidence in naturally conceived children. METHODS: All singleton children born in Denmark from January 1, 1995 through December 31, 2001 were stratified into children born without and after IVF, and were followed from birth until the end of 2002 in the National Register of Patients and the Central Register of Psychiatric Diseases, which include all discharge diagnoses from somatic and psychiatric hospitals/clinics, respectively. Included in the study were malignancies, mental, behavioural and neurological diseases, congenital syndromes, and developmental disturbances. Only diagnosis codes potentially relevant for imprinting diseases were included. RESULTS: During the 7-year study period, 442,349 singleton non-IVF and 6052 IVF children were born. Mean follow-up time was 4.5 and 4.1 years for the two groups, respectively, corresponding to 2 million and 25 000 follow-up years. In the IVF/non-IVF cohort, we detected 0/72 children with cancer, 47/3766 with mental diseases, 72/3654 neurological diseases, 4/287 congenital syndromes and 96/6727 developmental disturbances, in a total of 219/14,506 clinical outcomes. The number of children with specific imprinting diseases in the non-IVF group was 54: 44 kidney cancers, five retinoblastoma, three Prader-Willi syndrome and two Russel-Silver syndrome. Anticipating the same occurrence in IVF children, the total expected number was calculated to be 0.74. The observed number in the IVF group was 0. We found a significantly increased risk of cerebral palsy in the IVF group, with a rate ratio (RR) (IVF:non-IVF) of 1.8 [95% confidence interval (CI) 1.2-2.8; P < 0.01], and of sleeping disturbances, with an RR 2.0 (95% CI 1.2-3.3). The incidence rate of childhood cancers, mental diseases, congenital syndromes and developmental disturbances was equal in the two groups. CONCLUSIONS: We found no indication of an increased risk of imprinting diseases after IVF, but an 80% increased risk of cerebral palsy. We observed equal frequencies of childhood cancers, mental diseases, congenital syndromes and developmental disturbances in the two groups. Danish register data do not support reports of an increased risk of imprinting diseases after IVF. PMID- 15665018 TI - Parental coping with sudden infant death after donor insemination: case report. AB - We report the case of an artificial donor insemination couple experiencing sudden infant death of their 8-month-old child. Six months after the incident, the couple were investigated by means of an extensive interview, a repertory grid investigation and the Family Assessment Measure, as well as at 6 years after the incident by an extensive interview. The results show the importance of the diagnosis of male infertility and the preceding fertility treatment for coping with the death of their child. Six months after the incident, acute feelings associated with bereavement are mixed with feelings of anger and shame, apparently due to the experience of infertility. However, secrecy and shame associated with male infertility and donor insemination make it impossible for the couple to communicate their feelings to each other or to friends and relatives; furthermore, they decline psychological counselling. Repertory grid investigation and the Family Assessment Measure point to significant problems within the partnership. Six years after the incident, the couple's relationship is destabilized and both partners plan to divorce. We suggest a possible link between donor insemination secrecy and difficulties with coping. We discuss implications for couple counselling and emphasize the necessity for an improved legal framework for donor insemination in Germany. PMID- 15665019 TI - Unique t(Y;1)(q12;q12) reciprocal translocation with loss of the heterochromatic region of chromosome 1 in a male with azoospermia due to meiotic arrest: a case report. AB - A de novo reciprocal translocation 46,X,t(Y;1)(q12;q12) was found in an azoospermic male with meiotic arrest. Cytogenetics and fluorescent in situ hybridization (FISH) were used to define the karyotype, translocation breakpoints and homologue pairing. SRY (Yp), Yq11.2-AZF regions, DAZ gene copies and the distal Yq12 heterochromatin were studied by PCR and restriction analysis using sequence-tagged sites and single nucleotide variants. High resolution GTL, CBL and DA-DAPI staining revealed a (Y;1) translocation in all metaphases and a normal karyotype in the patient's father. FISH showed the presence of the distal Yq12 heterochromatic region in der(1) and loss of the heterochromatic region of chromosome 1. PCR demonstrated the intactness of the Y chromosome, including the SRY locus, AZF regions, DAZ genes and distal heterochromatin. A significant decrease (P = 0.005) of Xp/Yp pairing (18.6%), as compared with controls (65.7%), was found in arrested primary spermatocytes, and cell culture and mRNA expression studies confirmed an irreversible arrest at meiosis I, with induction of apoptosis and removal of germ cells by Sertoli cells. We characterized a de novo t(Y;1)(q12;q12) balanced reciprocal translocation with loss of the heterochromatic region of chromosome 1, that caused unpairing of sex chromosomes followed by meiosis I arrest, apoptotic degeneration of germ cells and azoospermia. PMID- 15665021 TI - Assisted reproductive technology in Europe, 2001. Results generated from European registers by ESHRE. AB - European results of assisted reproductive techniques from treatments initiated during 2001 are presented in this fifth report. Data were collected mainly from already existing national registers. From 23 countries, 579 clinics reported 289 690 cycles with: IVF 120 946, ICSI 114 378, frozen embryo transfer (FER) 47 195 and egg donation (ED) 7171. Overall this represents a 4% increase since the year 2000. For the first time, results on European data on intra-uterine inseminations (IUIs) were reported from 15 countries. A total of 67 124 cycles [IUI husband'sperm (IUI-H) 52 949 and IUI donor sperm (IUI-D) 14 185] were included. In 12 countries where all clinics reported to the register, a total of 108 910 cycles were performed in a population of 131.4 million, corresponding to 829 cycles per million inhabitants. For IVF, the clinical pregnancy rate per aspiration and per transfer was 25.1 and 29.0%, respectively. For ICSI, the corresponding rates were 26.2 and 28.3%. These figures are similar to the results from 2000. After IUI-H, the clinical pregnancy rate was 12.8% in women <40 and 9.7% in women > or =40 years of age. After IVF and ICSI, the distribution of transfer of one, two, three and > or =4 embryos was 12.0, 51.7, 30.8 and 5.5%, respectively. Compared with the year 2000, fewer embryos were transferred, but huge differences existed between countries. The distribution of singleton, twin and triplet deliveries for IVF and ICSI combined was 74.5, 24.0 and 1.5%, respectively. This gives a total multiple delivery rate of 25.5%, compared with 26.9% in the year 2000. The range of triplet deliveries after IVF and ICSI differed from 0.0 to 8.2% between countries. After IUI-H in women <40 years of age, 10.2% were twin and 1.1% were triplet gestations. PMID- 15665020 TI - A novel method for chromosome analysis of human sperm using enucleated mouse oocytes. AB - BACKGROUND: Mouse oocytes can be used in conjunction with intracytoplasmic sperm injection (ICSI) as a technique to permit chromosomal analysis of human sperm. However, chromosomes derived both from the human sperm and the mouse oocyte appear simultaneously following ICSI. The present study focused on evaluating whether or not previously enucleated mouse oocytes are usable for the analysis of human sperm chromosomes. METHODS: The metaphase chromosome-spindle complex was removed from a mouse oocyte. Human sperm from a donor with proven fertility were injected into mouse enucleated oocytes or intact oocytes. The presence of pronuclei in the oocytes was confirmed approximately 7-11 h after ICSI, and the oocytes were then fixed so that the nuclei could be observed as chromosome samples at 15-16 h after ICSI. RESULTS: The formation rate of one pronucleus in enucleated oocytes after ICSI was 93.9% (186/198) while that of two pronuclei in intact oocytes after ICSI was 85.4% (88/103). The appearance rate of metaphase chromosomes of human sperm in the enucleated oocytes, 89.4% (160/179), was significantly higher than that in intact oocytes, 78.7% (74/94) (P = 0.017). CONCLUSIONS: An efficient ICSI method using enucleated mouse oocytes was established to allow the visualization of the human sperm chromosome complement without the risk of confusion with mouse oocyte chromosomes. PMID- 15665022 TI - The importance of IRS-1 Gly972Arg polymorphism in evaluating the response to metformin treatment in polycystic ovary syndrome. AB - BACKGROUND: Recent evidence suggests that one of the modes of action of metformin may be through phosphorylation of the insulin receptor and insulin receptor substrates. With this in mind, we supposed that the G972A variant of insulin receptor substrate-1 (IRS-1) may modulate the response to metformin treatment in women with polycystic ovary syndrome (PCOS). METHODS: This preliminary study involved 60 randomly selected women with PCOS. All patients received dietary instructions and metformin 500 mg three times daily for 6 months. Main outcome measures were androgen levels, parameters of glucose and insulin metabolism and anthropometric variables. After a second evaluation of the patients at 6 months, they were genotyped for the Gly972Arg variant of the IRS-1 gene. RESULTS: Metformin had differential effects on fasting insulin levels, insulin resistance as demonstrated by homeostasis model assessment (HOMA), LH, total testosterone, dehydroepiandrosterone sulphate and free testosterone index on the basis of IRS genotype. The response to metformin therapy in other parameters was not different according to IRS genotype. CONCLUSION: There was a differential effect of metformin therapy in PCOS women on the basis of IRS genotype. This study may call attention to the importance of molecular markers in the management of women with PCOS. PMID- 15665023 TI - A prospective evaluation of a single-visit strategy to manage pregnancies of unknown location. AB - BACKGROUND: The aim was to assess whether women with a pregnancy of unknown location (PUL) can be safely excluded from potentially unnecessary multiple clinic visits. METHODS: A single-visit protocol was developed based upon data from 200 consecutive PULs. PULs were divided into groups according to the probable risk of ectopic pregnancy. Those PULs with an initial serum progesterone < or =10 nmol/l or a serum HCG of < or =25 U/l were deemed to be at low risk and classified as resolving or failing PULs. Those PULs with an initial serum progesterone of >50 nmol/l, regardless of serum HCG, were thought to be a probable intra-uterine pregnancy (IUP) and were also classified as being low risk. Those PULs with an initial serum progesterone of 10-50 nmol/l and a serum HCG of >25 U/l were classified as being at high risk. This protocol was then tested prospectively on 318 consecutive PULs. Management was based solely on the basis of an initial transvaginal ultrasound scan (TVS) and a single measurement of HCG and progesterone taken at the time of initial consultation. RESULTS: A total of 5544 consecutive women were scanned, of which 560 (10.1%) women were classified as PULs. Forty-two were lost to follow-up and therefore 518 (9.34%) were analysed. In the training set of 200 PULs, there were 111 (55.5%) failing PULs, 67 (33.5%) IUPs and 22 (11%) ectopic pregnancies. In the test set of 318, there were 189 (59.4%) failing PULs, 114 (35.8%) IUPs and 15 (4.7%) ectopic pregnancies. For the training group, the sensitivity and specificity of a single visit to detect low-risk PULs were 77 and 82%, respectively. The positive (PPV) and negative predictive values (NPV) were 97 and 31%, respectively. For the test group of 318 PULs, the sensitivity and specificity were 84 and 33%, respectively. The PPV and NPV were 96 and 9.4%, respectively. CONCLUSIONS: A single-visit strategy based on commonly used criteria eliminates 84% of non-ectopic pregnancies correctly from the system. However, as 67% of ectopic pregnancies are discharged without adequate follow-up, a single-visit strategy should not be used as an alternative to the current multi-visit strategy used in most units. PMID- 15665024 TI - Antioxidant intake is associated with semen quality in healthy men. AB - BACKGROUND: We seek to determine whether dietary and supplement intake of specific micronutrients (zinc and folate) and antioxidants (vitamins C, E and beta-carotene) is associated with semen quality. METHODS: Ninety-seven healthy, non-smoking men provided semen and were interviewed. Average daily nutrient intake from food and supplements was derived from a self-administered food frequency questionnaire. Intake levels were summarized as low, moderate and high. Semen volume, sperm concentration, total sperm count, motility, progressive motility and total progressively motile sperm count (TPMS) were measured. RESULTS: After controlling for covariates, a high intake of antioxidants was associated with better semen quality but, in almost all cases, there was no clear dose relationship in that moderate intake groups had the poorest semen quality. For example, positive associations were observed between vitamin C intake and sperm number as reflected in the higher mean count (P=0.04), concentration (P=0.05) and TPMS (P = 0.09); between vitamin E intake and progressive motility (P = 0.04) and TPMS (P = 0.05); and between beta-carotene intake and sperm concentration (P = 0.06) and progressive motility (P = 0.06). Folate and zinc intake were not associated with improved semen quality. CONCLUSIONS: In a convenience sample of healthy non-smoking men from a non-clinical setting, higher antioxidant intake was associated with higher sperm numbers and motility. PMID- 15665025 TI - A randomized double-blind controlled study of the efficacy of laser-assisted hatching on implantation and pregnancy rates of frozen-thawed embryo transfer at the cleavage stage. AB - BACKGROUND: Assisted hatching (AH) in fresh embryo transfer (ET) cycles increases the implantation and pregnancy rates, especially in women with a poor prognosis, repeated implantation failures and in older women. Little information exists in the literature regarding the role of AH in frozen-thawed embryo transfer (FET) cycles. METHODS: Embryos were cryopreserved at the cleavage stage. On the day of FET, 160 patients were randomized according to a computer-generated randomization list in sealed envelopes into the AH group and the control group. The patients and the clinicians were blinded to the group assigned. In the AH group, the outer half of the zona pellucida over a quarter of the diameter of zona was removed using a 1480 nm non-contact laser. RESULTS: The two groups were comparable in terms of demographic characteristics, ovarian response of the stimulated cycle and quality of fresh and frozen-thawed embryos. No differences in implantation, pregnancy and multiple pregnancy rates were found between the two groups. There was a non-significant trend of a higher implantation rate in the AH group when the zona thickness was > or = 16 mm. CONCLUSION: Laser AH did not improve the implantation rate of FET cycles and should not be performed routinely in all frozen-thawed embryos at the cleavage stage. PMID- 15665026 TI - Intact HCG, free HCG beta subunit and HCG beta core fragment: longitudinal patterns in urine during early pregnancy. AB - BACKGROUND: Detecting and monitoring early pregnancy depend on the measurement of HCG. Little is known about how production of various forms of HCG may evolve over the earliest weeks of pregnancy, particularly in naturally conceived pregnancies. METHODS: We describe the daily excretion of three urinary HCG analytes during the first 6 weeks post-conception in 37 naturally conceived pregnancies ending in singleton birth. We assayed daily first morning urine samples for intact HCG, free beta subunit and beta?core fragment, plus the combined measurement of these HCG forms. We calculated doubling times for each analyte and the inter- and intra subject day-to-day variation. RESULTS: Intact HCG and the free beta subunit were initially the predominant forms of HCG, with the beta core fragment emerging as the predominant form in the fifth week after conception. Intact HCG and the free beta subunit showed the most day-to-day variability, and were transiently undetectable even 10 days after detection of pregnancy. The most stable estimate of doubling time was provided by the combined measurement of all these forms. CONCLUSIONS: Although intact HCG is usually regarded as the main analyte for detection and monitoring of early pregnancy, it can fluctuate markedly during early pregnancy. This variability could affect pregnancy test results based on early pregnancy urine, and may distort estimates of doubling time. Assays that combine several forms of HCG may be more reliable. PMID- 15665027 TI - Characterization of recombinant influenza B viruses with key neuraminidase inhibitor resistance mutations. AB - OBJECTIVES AND METHODS: An influenza B virus plasmid-based rescue system was used to introduce site-specific mutations, previously observed in neuraminidase (NA) inhibitor-resistant viruses, into the NA protein of six recombinant viruses. Three mutations observed only among in vitro selected zanamivir-resistant influenza A mutants were introduced into the B/Beijing/1/87 virus NA protein, to change residue E116 to glycine, alanine or aspartic acid. Residue E116 was also mutated to valine, a mutation found in the clinic among oseltamivir-resistant viruses. An arginine to lysine change at position 291 (292 N2 numbering) mimicked that seen frequently in influenza A N2 clinical isolates resistant to oseltamivir. Similarly, an arginine to lysine change at position 149 (152 in N2 numbering) was made to reproduce the change found in the only reported zanamivir resistant clinical isolate of influenza B virus. In vitro selection and prolonged treatment in the clinic leads to resistance pathways that require compensatory mutations in the haemagglutinin gene, but these appear not to be important for mutants isolated from immunocompetent patients. The reverse genetics system was therefore used to generate mutants containing only the NA mutation. RESULTS AND CONCLUSIONS: With the exception of a virus containing the E116G mutation, mutant viruses were attenuated to different levels in comparison with wild-type virus. This attenuation was a result of altered NA activity or stability depending on the introduced mutation. Mutant viruses displayed increased resistance to zanamivir, oseltamivir and peramivir, with certain viruses displaying cross resistance to all three drugs. PMID- 15665028 TI - C-peptide prevents glomerular hypertrophy and mesangial matrix expansion in diabetic rats. AB - BACKGROUND: There is accumulating evidence that C-peptide exerts beneficial renal effects in type-1 diabetes by reducing glomerular hyperfiltration, albuminuria and glomerular hypertrophy in the early stage of nephropathy. The aim of this study was to clarify further the effects of C-peptide on renal structural changes in type-1 diabetic rats. METHODS: The effects of C-peptide or placebo on glomerular volume, mesangial expansion, glomerular basement membrane thickness, albuminuria and glomerular filtration rate (GFR) were studied in three groups of rats: a non-diabetic group (N, n=9) and two groups that, during 8 weeks of diabetes, were left untreated for 4 weeks and then given a subcutaneous infusion of either placebo (D, n=11) or C-peptide (DCp, n=11) during the next 4 weeks. Furthermore, GFR was studied after 4 weeks of diabetes in an additional diabetic group (D-early, n=9) and in an age-matched non-diabetic group (N-early, n=9). RESULTS: After 4 weeks, GFR in the D-early group was 102% higher than in the N early group. GFR after 8 weeks did not differ between the study groups. The D group presented with a 33% larger glomerular volume than the N group (P<0.001), while glomerular volume in the DCp group was similar to that in the N-group. Total mesangial and mesangial matrix fractions were increased by 46% (P<0.001) and 133% (P<0.001), respectively, in the D group. The corresponding values in the DCp group did not differ from those for the non-diabetic animals. Neither the thickness of the glomerular basement membrane nor the level of albuminuria differed significantly between the study groups. CONCLUSIONS: C-peptide administration in replacement dose to streptozotocin-diabetic rats serves to limit or prevent the glomerular hypertrophy and the mesangial matrix expansion seen in the post-hyperfiltration phase of early diabetic nephropathy. PMID- 15665029 TI - Chewing gum and a saliva substitute alleviate thirst and xerostomia in patients on haemodialysis. AB - BACKGROUND: Most patients on haemodialysis (HD) have to maintain a fluid restricted diet to prevent a high interdialytic weight gain (IWG). The prevalence of xerostomia (the feeling of a dry mouth) is higher in HD patients than in controls. Recently, we demonstrated that xerostomia and thirst were positively correlated with IWG in HD patients. Thus, this may play a role as a stimulus for fluid intake between dialysis sessions. The aim of the present study was to investigate the effect of chewing gum or a saliva substitute on xerostomia, thirst and IWG. METHODS: This study was a randomized two-treatment crossover design with repeated measures. After the use of chewing gum or saliva substitute for 2 weeks, a wash-out period of 2 weeks was introduced and hereafter the other regimen was carried out. Xerostomia and thirst were assessed by validated questionnaires as xerostomia inventory (XI) and dialysis thirst inventory (DTI), at baseline and after each treatment period, as were IWG and salivary flow rates. RESULTS: Sixty-five HD patients (42 men, 54.6+/-14.1 years; 23 women, 54.7+/-16.3 years) participated in this study. Chewing gum decreased XI from 29.9+/-9.5 to 28.1+/-9.1 (P<0.05). Chewing gum as well as a saliva substitute reduced DTI significantly (P<0.05), but no differences occurred for the average IWG or salivary flow rates. CONCLUSIONS: The use of chewing gum and, to a lesser extent, a saliva substitute may alleviate thirst and xerostomia in some HD patients. PMID- 15665030 TI - Outcome of home haemodialysis patients: a case-cohort study. AB - BACKGROUND: Randomized, controlled comparisons between home haemodialysis (HHD) and centre haemodialysis (CHD) have not been performed to date. Reported survival benefits of HHD as compared with CHD from uncontrolled studies have been attributed largely to patient selection. METHODS: In order to minimize a selection bias, we have compared the outcome of our HHD and CHD patients with a nested case-cohort study. For each patient trained for HHD at our dialysis centre between 1970 and 1995 (n=103), a corresponding match was searched from the CHD patients by retrospective chart analysis. The pairs were matched for sex, age (+/ 5 years), time of dialysis therapy onset (+/-2 years) and renal disease category. For 58 of the 103 HHD patients, a corresponding matched CHD patient was identified. Both treatment groups had the same mean age (50+/-13 years) at dialysis onset and were comparable with respect to the Khan comorbidity index, prevalence and duration of hypertension, smoking habits, history of myocardial infarction, stroke and peripheral vascular disease. In both groups, approximately 50% of the patients were transplanted during the observation period. RESULTS: HHD patients were hospitalized less often and tended to have fewer operations as compared with CHD patients. Survival was significantly longer in HHD as compared with CHD. Five, 10 and 20 year survival rates were 93 (n=55 patients at risk), 72 (41) and 34% (11) with HHD and 64 (38), 48 (26) and 23% (4) with CHD, respectively. This survival difference persisted after adjusting for predictors of mortality, i.e. age at onset of dialysis, year of start of dialysis therapy and Khan comorbidity index. CONCLUSIONS: HHD offers a cheap and valuable alternative to CHD, with no apparent disadvantages. PMID- 15665031 TI - Increased bone resorption in HD patients: is it caused by elevated RANKL synthesis? AB - BACKGROUND: The receptor activator of nuclear factor kappaB ligand (RANKL), produced by osteoblasts/stromal cells, is a member of the RANK/RANKL/OPG system, which regulates bone resorption by osteoclasts. Since RANKL and osteoprotegerin (OPG) production in bone is influenced by parathyroid hormone (PTH), we measured serum RANKL and OPG concentrations in haemodialysis (HD) patients, who commonly hypersecrete PTH. We aimed to determine if clinically demonstrated PTH-enhanced bone resorption is a consequence of increased RANKL synthesis. METHODS: RANKL, OPG, osteocalcin, intact PTH, bone-specific alkaline phosphatase, tartrate resistant acid phosphatase 5b and beta-CrossLaps (CTx) were measured in blood samples from 80 HD patients and 50 age-matched controls. HD patients were stratified to tertiles according to their serum PTH levels: 29.3-103.0, 109.7 263.0 and 262.0-1700.0 pg/ml in the first, second and third tertiles, respectively. RESULTS: Mean serum RANKL levels were 1.6 times higher in HD patients than in age-matched controls (1.36+/-0.39 vs 0.83+/-0.70 pmol/l; P<0.001). All the measured bone markers significantly differed between patients and controls (P<0.001). Spearman's tests of correlation showed a statistically significant association of RANKL with PTH, osteocalcin and CTx (r=0.322, P=0.004; r=0.231, P=0.039; and r=0.230, P=0.040, respectively). Mean serum RANKL levels were significantly different between PTH tertiles (P = 0.003), but serum OPG levels were not (P=0.144). The highest RANKL levels were measured in the upper PTH tertile (1.54+/-0.39 pmol/l) and were significantly higher than in the middle or lower tertiles (1.27+/-0.42 and 1.23+/-0.26 pmol/l, respectively; P=0.003). Both of the measured bone-resorption markers, tartarate-resistant acid phosphatase 5b and CTx, as well as both bone formation markers, osteocalcin and bone-specific alkaline phosphatase were also significantly higher in the upper tertile, indicating that whole-bone remodelling is activated at high PTH and RANKL levels. CONCLUSIONS: Serum RANKL levels were significantly higher in HD patients than in healthy age-matched controls. Moreover, RANKL levels were significantly higher in the upper PTH tertile, indicating enhanced RANKL synthesis in a PTH-dependent fashion. Thus, our clinical findings clearly support published in vitro studies that demonstrated a stimulating effect of PTH on RANKL synthesis. Therefore, the hypothesis that PTH increases bone resorption in HD patients through RANKL appears valid. PMID- 15665032 TI - HGF reduces advancing lung fibrosis in mice: a potential role for MMP-dependent myofibroblast apoptosis. AB - Pulmonary fibrosis is characterized by a loss of lung epithelial cells, replaced by interstitial myofibroblasts to deposit extracellular matrix (ECM) proteins. Previous studies demonstrated that hepatocyte growth factor (HGF) improved lung fibrosis in murine models, whereas molecular mechanisms whereby HGF improved lung fibrosis have yet to be fully understood. When MRC-5 human lung fibroblasts were treated with transforming growth factor-beta1, the cells underwent phenotypic change similar to myofibroblasts and this was associated with up-regulation of c Met/HGF receptor expression. For the myofibroblast-like cells, HGF increased activities of MMP-2/-9, predominant enzymes for breakdown of fibronectin (FN). Under such conditions, HGF induced caspase-dependent apoptosis, linked with a decrease in a FN central cell binding (CCB) domain involved in FAK phosphorylation. When MMI270 (a broad-spectrum MMP inhibitor) was added together with HGF, decreases in FN-CCB domain expression and FAK phosphorylation by HGF were restored, and these events were associated with an inhibition of HGF-induced apoptosis, suggesting that increased activities of MMPs underlie the major mechanism of HGF-mediated apoptosis in myofibroblasts. In bleomycin-treated mice, c-Met expression was found on interstitial myofibroblasts and HGF increased apoptosis in culture of myofibroblasts isolated from bleomycin-treated murine lungs. Furthermore, administration of recombinant HGF to bleomycin-treated mice increased lung MMP activities and enhanced myofibroblast apoptosis, while in vivo MMI270 injections together with HGF inhibited such MMP activation, leading to suppressed myofibroblast apoptosis. In conclusion, we identified HGF as a key ligand to elicit myofibroblast apoptosis and ECM degradation, whereas activation of the HGF/c-Met system in fibrotic lungs may be considered a target to attenuate progression of chronic lung disorders. PMID- 15665033 TI - Expression of caveolin-1 in lymphocytes induces caveolae formation and recruitment of phosphofructokinase to the plasma membrane. AB - Compartmentation of carbohydrate metabolism has been shown in a wide range of tissues including reports of one compartment of glycolysis associated with the plasma membrane of cells. However, only in the erythrocyte has the physical basis for plasma membrane-associated glycolytic pathway been established. We have previously found that phosphofructokinase (PFK) appeared to colocalize with the fairly ubiquitous plasma membrane protein caveolin-1 (CAV-1), consistent with a role for CAV-1 as an anchor for glycolysis to the plasma membrane. To test the hypothesis that CAV-1 functions as a scaffolding protein for PFK, we transfected human lymphocytes (a cell without CAV-1 expression) with human CAV-1 cDNA. We demonstrate that expression of CAV-1 in lymphocytes results in the formation of caveolae at the plasma membrane and affects the subcellular localization of PFK by recruiting PFK to the plasma membrane. Targeting of PFK by CAV-1 also was validated by the significant colocalization between the proteins after transfection, which resulted in a correlation of 0.97 +/- 0.004 between the two fluorophores. This finding is significant in as much as it illustrates the CAV-1 feasibility of generating binding sites for glycolytic enzymes on the plasma membrane. We therefore conclude that CAV-1 functions as a scaffolding protein for PFK and that this may contribute to the elucidation of the basis for carbohydrate compartmentation to the plasma membrane in a wide variety of cell types. PMID- 15665034 TI - Angiotensin AT2 receptor protects against cerebral ischemia-induced neuronal injury. AB - Several lines of clinical and experimental evidence suggest an important role of the renin-angiotensin system in ischemic brain injury although the cellular regulation of the angiotensin AT1 and AT2 receptors and their potential relevance in this condition have not yet been clearly defined. We first assessed the regulation of brain AT1 and AT2 receptors in response to transient unilateral medial cerebral artery occlusion in rats by real-time RT-PCR, Western blot, and immunofluorescence labeling. AT2 receptors in the peri-infarct zone were significantly upregulated 2 days after transient focal cerebral ischemia. Increased AT2 receptors, which were abundantly distributed in a large number of brain regions adjacent to the infarct area including cerebral frontal cortex, piriform cortex, striatum, and hippocampus, were exclusively expressed in neurons. By contrast, AT1 receptors, which remained unaltered, were mainly expressed in astrocytes. In neurons of ischemic striatum, increased AT2 receptors were associated with intense neurite outgrowth. Blockade of central AT2 receptors with PD123177 abolished the neuroprotective effects of central AT1 receptor blockade with irbesartan on infarct size and neurological outcome. In primary cortical neurons, stimulation of AT2 receptors supported neuronal survival and neurite outgrowth. Our data indicate that cerebral AT2 receptors exert neuroprotective actions in response to ischemia-induced neuronal injury, possibly by supporting neuronal survival and neurite outgrowth in peri-ischemic brain areas. PMID- 15665035 TI - Muscle fiber specific apoptosis and TNF-alpha signaling in sarcopenia are attenuated by life-long calorie restriction. AB - Increased tumor necrosis factor-alpha (TNF-alpha) levels have been found with age and are connected to muscle atrophy and cell loss, yet the signaling events that occur in vivo are unknown. Calorie restriction (CR), a robust intervention shown to repeatedly evade the physiological declines associated with aging, has been reported to reduce TNF-alpha and may assist in understanding the mechanisms of muscle sarcopenia. The effects of age and CR on muscle mass, myocyte area, fiber number, myocyte TNF-alpha expression, plasma TNF-alpha levels, and specific elements linked with the TNF-alpha signaling cascade (TNF-R1, IKKgamma, IkappaBalpha, p65, NF-kappaB binding activity, FADD, caspase-8, and DNA fragmentation) were investigated in soleus (predominately Type I fiber), and superficial vastus lateralis (SVL, predominately Type II fiber), of 6-month-old ad libitum fed (6AL), 26-month-old ad libitum fed (26AL), and 26-month-old calorie-restricted (26CR) male Fischer 344 rats (CR = 40% restriction compared with ad libitum). Plasma TNF-alpha was increased with age, and the age-associated rise was attenuated with life-long CR. In soleus muscle, we reported a greater capacity to cultivate inflammatory signaling through the transcription factor NF kappaB compared with that detected in SVL with age. In contrast, in the SVL TNF alpha stimulated apoptotic signaling with age to a much higher extent than was observed in the soleus. Moreover, a reduction in muscle mass, cross-sectional area, and fiber number in the SVL coincided with this age-linked elevation in apoptosis. In agreement with CR's ability, TNF-alpha stimulation of both inflammatory and apoptotic pathways were abrogated. Our results suggest that TNF alpha signals transmitted to specific fiber types determine the decision of selecting life or death signaling pathways and are linked to the extent of fiber loss experienced in the aging muscle. Such a specific potential may constitute a major proponent in the pathogenesis of sarcopenia. PMID- 15665036 TI - ABAD enhances Abeta-induced cell stress via mitochondrial dysfunction. AB - Amyloid-beta peptide (Abeta) binding alcohol dehydrogenase (ABAD), an enzyme present in neuronal mitochondria, is a cofactor facilitating Abeta-induced cell stress. We hypothesized that ABAD provides a direct link between Abeta and cytotoxicity via mitochondrial oxidant stress. Neurons cultured from transgenic (Tg) mice with targeted overexpression of a mutant form of amyloid precursor protein and ABAD (Tg mAPP/ABAD) displayed spontaneous generation of hydrogen peroxide and superoxide anion, and decreased ATP, as well as subsequent release of cytochrome c from mitochondria and induction of caspase-3-like activity followed by DNA fragmentation and loss of cell viability. Generation of reactive oxygen species (ROS) was associated with dysfunction at the level of mitochondrial complex IV (cytochrome c oxidase, or COX). In neurons cultured from Tg mAPP/ABAD mice, COX activity was selectively decreased, and cyanide, an inhibitor of complex IV, exacerbated leakage of ROS, induction of caspase-3-like activity, and DNA fragmentation. In vivo, Tg mAPP/ABAD mice displayed reduced levels of brain ATP and COX activity, diminished glucose utilization, as well as electrophysiological abnormalities in hippocampal slices compared with Tg mAPP mice. In contrast, neither Tg ABAD mice nor nontransgenic (non-TG) littermates showed similar changes in ATP, COX activity, glucose utilization or electrophysiological properties. Each of the genotypes (Tg ABAD, Tg mAPP and Tg mAPP/ABAD mice, and non-TG littermates) displayed normal reproductive fitness, development and lifespan (1) These findings link ABAD-induced oxidant stress to critical aspects of Alzheimer's disease (AD)-associated cellular dysfunction, suggesting a pivotal role for this enzyme in the pathogenesis of AD. PMID- 15665037 TI - Ultrastructure of fibre and parenchyma cell walls during early stages of culm development in Dendrocalamus asper. AB - BACKGROUND AND AIMS: The anatomy of bamboo culms and the multilayered structure of fibre cell walls are known to be the main determinant factors for its physical and mechanical properties. Studies on the bamboo cell wall have focussed mainly on fully elongated and mature fibres. The main aim of this study was to describe the ultrastructure of primary and secondary cell walls in culm tissues of Dendrocalamus asper at different stages of development. METHODS: The development of fibre and parenchyma tissues was classified into four stages based on light microscopy observations made in tissues from juvenile plants. The stages were used as a basis for transmission electron microscopy study on the ultrastructure of the cell wall during the process of primary and early secondary cell wall formation. Macerations and phloroglucinol-HCl staining were employed to investigate fibre cell elongation and fibre cell wall lignification, respectively. KEY RESULTS: The observations indicated that the primary wall is formed by the deposition of two distinct layers during the elongation of the internode and that secondary wall synthesis may begin before the complete cessation of internode and fibre elongation. Elongation was followed by a maturation phase characterized by the deposition of multiple secondary wall layers, which varied in number according to the cell type, location in the culm tissue and stage of shoot development. Lignification of fibre cell walls started at the period prior to the cessation of internode elongation. CONCLUSIONS: The structure of the primary cell wall was comprised of two layers. The fibre secondary cell wall began to be laid down while the cells were still undergoing some elongation, suggesting that it may act to cause the slow-down and eventual cessation of cell elongation. PMID- 15665038 TI - The p38 mitogen-activated protein kinase inhibitor SB203580 reduces glucose turnover by the glucose transporter-4 of 3T3-L1 adipocytes in the insulin stimulated state. AB - Insulin induces a profound increase in glucose uptake in 3T3-L1 adipocytes through the activity of the glucose transporter-4 (GLUT4). Apart from GLUT4 translocation toward the plasma membrane, there is also an insulin-induced p38 MAPK-dependent step involved in the regulation of glucose uptake. Consequently, treatment with the p38 MAPK inhibitor SB203580 reduces insulin-induced glucose uptake by approximately 30%. Pretreatment with SB203580 does not alter the apparent K(m) of GLUT4-mediated glucose uptake but reduces the maximum velocity by approximately 30%. Insulin-induced GLUT4 translocation and exposure of the transporter to the extracellular environment was not altered by pretreatment with SB203580, as evidenced by a lack of effect of the inhibitor on the amount of GLUT4 present in the plasma membrane, as assessed by subcellular fractionation, the amount of GLUT4 that is able to undergo biotinylation on intact adipocytes and the level of extracellular exposure of an ectopically expressed GLUT-green fluorescence protein construct with a hemagglutinin tag in its first extracellular loop. In contrast, labeling of GLUT4 after insulin stimulation by a membrane-impermeable, mannose moiety-containing, photoaffinity-labeling agent [2 N-4(1-azido-2,2,2-trifluoroethyl)benzoyl-1,3-bis(d-mannose-4-yloxy)-2 propylamine] that binds to the extracellular glucose acceptor domain was markedly reduced by SB203580, although photolabeling with this compound in the absence of insulin was unaffected by SB203580. These data suggest that SB203580 affects glucose turnover by the insulin-responsive GLUT4 transporter in 3T3-L1 adipocytes. PMID- 15665039 TI - Alpha1- and beta1-adrenoceptor signaling fully compensates for beta3-adrenoceptor deficiency in brown adipocyte norepinephrine-stimulated glucose uptake. AB - To assess the relative roles and potential contribution of adrenergic receptor subtypes other than the beta3-adrenergic receptor in norepinephrine-mediated glucose uptake in brown adipocytes, we have here analyzed adrenergic activation of glucose uptake in primary cultures of brown adipocytes from wild-type and beta3-adrenergic receptor knockout (KO) mice. In control cells in addition to high levels of beta3-adrenergic receptor mRNA, there were relatively low alpha1A , alpha1D-, and moderate beta1-adrenergic receptor mRNA levels with no apparent expression of other adrenergic receptors. The levels of alpha1A-, alpha1D-, and beta1-adrenergic receptor mRNA were not changed in the beta3-KO brown adipocytes, indicating that the beta3-adrenergic receptor ablation does not influence adrenergic gene expression in brown adipocytes in culture. As expected, the beta3 adrenergic receptor agonists BRL-37344 and CL-316 243 did not induce 2-deoxy-d glucose uptake in beta3-KO brown adipocytes. Surprisingly, the endogenous adrenergic neurotransmitter norepinephrine induced the same concentration dependent 2-deoxy-D-glucose uptake in wild-type and beta3-KO brown adipocytes. This study demonstrates that beta1-adrenergic receptors, and to a smaller degree alpha1-adrenergic receptors, functionally compensate for the lack of beta3 adrenergic receptors in glucose uptake. Beta1-adrenergic receptors activate glucose uptake through a cAMP/protein kinase A/phosphatidylinositol 3-kinase pathway, stimulating conventional and novel protein kinase Cs. The alpha1 adrenergic receptor component (that is not evident in wild-type cells) stimulates glucose uptake through a phosphatidylinositol 3-kinase and protein kinase C pathway in the beta3-KO cells. PMID- 15665040 TI - Acute hypoxia increases intracellular [Ca2+] in pulmonary arterial smooth muscle by enhancing capacitative Ca2+ entry. AB - Hypoxic pulmonary vasoconstriction (HPV) requires influx of extracellular Ca2+ in pulmonary arterial smooth muscle cells (PASMCs). To determine whether capacitative Ca2+ entry (CCE) through store-operated Ca2+ channels (SOCCs) contributes to this influx, we used fluorescent microscopy and the Ca2+-sensitive dye fura-2 to measure effects of 4% O2 on intracellular [Ca2+] ([Ca2+]i) and CCE in primary cultures of PASMCs from rat distal pulmonary arteries. In PASMCs perfused with Ca2+-free Krebs Ringer bicarbonate solution (KRBS) containing cyclopiazonic acid to deplete Ca2+ stores in sarcoplasmic reticulum and nifedipine to prevent Ca2+ entry through L-type voltage-operated Ca2+ channels (VOCCs), hypoxia markedly enhanced both the increase in [Ca2+]i caused by restoration of extracellular [Ca2+] and the rate at which extracellular Mn2+ quenched fura-2 fluorescence. These effects, as well as the increased [Ca2+]i caused by hypoxia in PASMCs perfused with normal salt solutions, were blocked by the SOCC antagonists SKF-96365, NiCl2, and LaCl3 at concentrations that inhibited CCE >80% but did not alter [Ca2+]i responses to 60 mM KCl. In contrast, the VOCC antagonist nifedipine inhibited [Ca2+]i responses to hypoxia by only 50% at concentrations that completely blocked responses to KCl. The increased [Ca2+]i caused by hypoxia was completely reversed by perfusion with Ca2+-free KRBS. LaCl3 increased basal [Ca2+]i during normoxia, indicating effects other than inhibition of SOCCs. Our results suggest that acute hypoxia enhances CCE through SOCCs in distal PASMCs, leading to depolarization, secondary activation of VOCCs, and increased [Ca2+]i. SOCCs and CCE may play important roles in HPV. PMID- 15665041 TI - Chronic hypoxia inhibits Kv channel gene expression in rat distal pulmonary artery. AB - In pulmonary arterial smooth muscle cells (PASMCs), voltage-gated K+ (Kv) channels play an important role in regulating membrane potential, cytoplasmic free Ca2+ concentration, and pulmonary vasomotor tone. Previous studies demonstrated that exposure of rats to chronic hypoxia decreased Kv channel function in PASMCs from distal pulmonary arteries (dPA). To determine whether this decrease in function was due to decreased expression of Kv channel proteins and which Kv proteins might be involved, we analyzed Kv channel gene expression in intact, endothelium-denuded dPAs obtained from rats exposed to 10% O2 for 3 wk. Kv1.1, Kv1.2, Kv1.4, Kv1.5, Kv1.6, Kv2.1, Kv3.1, Kv4.3, and Kv9.3 channel alpha-subunits and Kv1, Kv2, and Kv3 beta-subunits were expressed in rat dPAs. Exposure to chronic hypoxia decreased mRNA and protein levels of Kv1.1, Kv1.5, Kv1.6, Kv2.1, and Kv4.3 alpha-subunits in dPAs but did not alter gene or protein expression of these channels in aorta. Furthermore, chronic hypoxia did not alter the mRNA levels of beta-subunits in dPAs. These results suggest that diminished transcription of Kv alpha-subunits may reduce the number of functional Kv channels in dPAs during prolonged hypoxia, causing the decreased Kv current previously observed in PASMCs and leading to pulmonary artery vasoconstriction. PMID- 15665042 TI - Simvastatin attenuates vascular leak and inflammation in murine inflammatory lung injury. AB - Therapies to limit the life-threatening vascular leak observed in patients with acute lung injury (ALI) are currently lacking. We explored the effect of simvastatin, a 3-hydroxy-3-methylglutaryl (HMG)-CoA reductase inhibitor that mediates endothelial cell barrier protection in vitro, in a murine inflammatory model of ALI. C57BL/6J mice were treated with simvastatin (5 or 20 mg/kg body wt via intraperitoneal injection) 24 h before and again concomitantly with intratracheally administered LPS (2 microg/g body wt). Inflammatory indexes [bronchoalveolar lavage (BAL) myeloperoxidase activity and total neutrophil counts assessed at 24 h with histological confirmation] were markedly increased after LPS alone but significantly reduced in mice that also received simvastatin (20 mg/kg; approximately 35-60% reduction). Simvastatin also decreased BAL albumin (approximately 50% reduction) and Evans blue albumin dye extravasation into lung tissue (100%) consistent with barrier protection. Finally, the sustained nature of simvastatin-mediated lung protection was assessed by analysis of simvastatin-induced gene expression (Affymetrix platform). LPS-mediated lung gene expression was significantly modulated by simvastatin within a number of gene ontologies (e.g., inflammation and immune response, NF-kappaB regulation) and with respect to individual genes implicated in the development or severity of ALI (e.g., IL-6, Toll-like receptor 4). Together, these findings confirm significant protection by simvastatin on LPS-induced lung vascular leak and inflammation and implicate a potential role for statins in the management of ALI. PMID- 15665043 TI - Oncostatin M causes VEGF release from human airway smooth muscle: synergy with IL 1beta. AB - Vascular endothelial growth factor (VEGF), a potent angiogenesis factor, likely contributes to airway remodeling in asthma. We sought to examine the effects and mechanism of action of IL-6 family cytokines on VEGF release from human airway smooth muscle (HASM) cells. Oncostatin M (OSM), but not other IL-6 family cytokines, increased VEGF release, and IL-1beta enhanced OSM-induced VEGF release. OSM increased VEGF mRNA expression and VEGF promoter activity, whereas IL-1beta had no effect. IL-1beta did not augment the effects of OSM on VEGF promoter activity but did augment OSM-induced VEGF mRNA expression and mRNA stability. The STAT3 inhibitor piceatannol decreased both OSM-induced VEGF release and synergy between OSM and IL-1beta, without affecting responses to IL 1beta alone. Piceatannol also inhibited OSM-induced VEGF mRNA expression. In contrast, inhibitors of MAPK pathway had no effect on OSM or OSM plus IL-1beta induced VEGF release. OSM increased type 1 IL-1 receptor (IL-1R1) mRNA expression, as measured by real-time PCR, and piceatannol attenuated this response. Consistent with the increase in IL-1R1 expression, OSM markedly augmented IL-1beta-induced VEGF, MCP-1, and IL-6 release. In summary, our data indicate OSM causes VEGF expression in HASM cells by a transcriptional mechanism involving STAT3. IL-1beta also synergizes with OSM to increase VEGF release, likely as a result of effects of IL-1beta on VEGF mRNA stability as well as effects of OSM on IL-1R1 expression. This is the first description of a role for OSM on IL-1R1 expression in any cell type. OSM may contribute to airway remodeling observed in chronic airway disease. PMID- 15665044 TI - Cytosolic NADH redox and thiol oxidation regulate pulmonary arterial force through ERK MAP kinase. AB - An ERK MAP kinase-mediated contractile mechanism previously reported to be activated by peroxide and stretch in bovine coronary arteries is shown in this study to be present in endothelium-denuded bovine pulmonary arteries and subject to regulation by modulation of cytosolic NAD(H) redox through the lactate dehydrogenase reaction. Although our previous work identified an acute PO2 dependent peroxide-mediated relaxation of bovine pulmonary arteries on exposure to lactate, a 30-min treatment with 10 mM lactate enhanced ERK phosphorylation and increased force generation to 30 mM KCl. Hypoxia inhibited these responses to lactate. Increases in ERK phosphorylation and the enhancement of force generation by lactate and stretch are attenuated in the presence of inhibitors of Nox oxidase (0.1 mM apocynin) or ERK activation (10 microM PD-98059) and by 0.1 mM ebselen. Additionally, incubation of pulmonary arteries with 10 mM pyruvate lowered basal levels of ERK phosphorylation, and it inhibited both the ERK phosphorylation and the enhancement in force generation to 30 mM KCl caused by stretch. Treatment of pulmonary arteries with the thiol oxidant diamide (1 microM) elicited what appears to be a peroxide-independent increase in force and ERK phosphorylation that were both attenuated by PD-98059. Thus pulmonary arteries possess a peroxide-elicited contractile mechanism involving ERK MAP kinase, which is stimulated by stretch and regulated through the control of Nox oxidase activity by the availability of cytosolic NADH. PMID- 15665045 TI - Crystalline and amorphous silica differentially regulate the cyclooxygenase prostaglandin pathway in pulmonary fibroblasts: implications for pulmonary fibrosis. AB - Inhalation of crystalline (CS) and amorphous silica (AS) results in human pulmonary inflammation. However, silicosis develops only following CS exposure, and the pathogenic mechanisms are poorly understood. This report describes the differential abilities of CS and AS to directly upregulate the early inflammatory mediator COX-2, the recently identified prostaglandin E (PGE) synthase and the downstream mediator PGE2 in primary human lung fibroblasts. Increased cyclooxygenase (COX)-2 gene transcription and protein production were demonstrated by ribonuclease protection assay, Western blot analysis, and immunocytochemistry. In each case the ability of AS to induce COX-2 exceeded that of CS. Similarly, downstream of COX-2, production of the antifibrotic prostaglandin PGE2 was induced in a dose-dependent fashion, but AS was significantly more potent (maximal production: CS = 4,710 pg/ml and AS = 7,651 pg/ml). These increases in COX-2 and PGE2 were preceded by induction of the PGE2 synthase protein, demonstrating the potential role of this novel molecule in silica-mediated inflammation. There was specificity of induction of prostaglandins, as PGF2alpha, but not PGD2, was induced. Using specific COX-2 inhibitors, we showed increased PG production to be dependent on the COX-2 enzyme. Furthermore, stimulation of fibroblasts was particle specific, as silica but not carbon black resulted in fibroblast activation. These results demonstrate that silica can directly stimulate human lung fibroblasts to produce key inflammatory enzymes and prostaglandins. Moreover, they suggest a mechanism to explain the differing fibrogenic potential of CS and AS. The molecules COX-2, PGE synthase, and PGE2 are identified as effectors in silicosis. PMID- 15665046 TI - Poor agreement among prothrombin time international normalized ratio methods: comparison of seven commercial reagents. AB - BACKGROUND: Prothrombin time (PT) has long been the most popular test for monitoring oral anticoagulation therapy. The International Normalized Ratio (INR) was introduced to overcome the problem of marked variation in PT results among laboratories and the various recommendations for patient care. According to this principle, all reagents should be calibrated to give identical results and the same patient care globally. This is necessary for monitoring of single patients and for application of the results of anticoagulation trials and guidelines to clinical practice. METHODS: We took blood samples from 150 patients for whom oral anticoagulation had been prescribed. Plasmas were separated and PTs determined by use of seven commercial reagents and four calibrator sets. The differences in results were assessed by plotting, for each possible pair of methods, the differences in INR values for each sample against the mean INR value (Bland Altman difference plots). RESULTS: Mean results differed significantly (P <0.001) for 17 of 21 possible paired comparisons of methods. Only two pairs of methods produced very similar results when assessed for problems of substantial differences in INR values; a significant, systematic increase in the difference with INR; and a significant systematic increase in the variation in difference with increasing INR values. CONCLUSIONS: The agreement among several (and perhaps most) commercial INR methods is poor. The failure of current calibration strategies may severely compromise both the monitoring of individual patients and the application of oral anticoagulation guidelines and trial results to clinical practice. PMID- 15665048 TI - Mechanism of potentiation by polyphenols of contraction in human vein-engineered media. AB - The potential of natural dietary polyphenols in the treatment of vascular diseases originating from veins has been suggested in the literature. However, the mechanisms involved to explain the effects of polyphenols are not yet elucidated. Therefore, the aim of this study was to investigate the mechanisms by which polyphenols from red wine (Provinols) modulated contraction in human veins. We took advantage of a human model previously reported as a new tool for pharmacological research, using tissue-engineered techniques allowing the production of vascular media based exclusively on human smooth muscle cells. Thus human tissue-engineered vascular media (TEVM) were produced with cells originating from umbilical cord vein. TEVM were treated with either vehicle or Provinols. Results showed that treatment of TEVM with Provinols significantly potentiated the contractile responses induced by histamine and bradykinin. The potentiating effect of Provinols was not associated with an enhancement of histamine-induced increase in cytosolic calcium; rather, it implied the presence of a Ca(2+)-independent signaling pathway. Pharmacological studies indicated that action of Provinols took place at the level of phospholipase A(2)-Rho-kinase pathway and was associated with an enhancement of myosin light chain kinase activity. These results, obtained using the human TEVM, bring new insights to explain the regulation of venous contraction by polyphenols. PMID- 15665047 TI - Potential candidates for ischemic preconditioning-associated vascular growth pathways revealed by antibody array. AB - Our understanding of the phenomenon of myocardial vascular growth is very limited even though various studies have been conducted in several different models, because the focus in each has been on a select very few number of proteins as the possible growth factors. In the present study, we used the ischemic preconditioning (IP) model in the form of four in vivo repetitive cycles of coronary artery occlusion, each followed by reperfusion as the model to stimulate vascular growth, and performed the protein profiling using high-throughput antibody array technology. Rats were divided into two groups: control + left anterior descending coronary artery (LAD) occlusion (CMI), and IP+ LAD occlusion (IPMI). The antibody array experiment performed to compare the expression of 512 proteins between the IPMI and CMI samples revealed significant upregulation of growth proteins like TGF-beta, BMX, granulocyte-monocyte colony-stimulating factor, signal transducer and activator of transcription 3, alpha- and beta catenins, ubiquitin-conjugating enzyme UbcH6, nexilin, and PKC-epsilon and lambda. JNK1 and c-Src tyrosine kinase were expectedly found to be downregulated. Western blot experiments validated the changes in expression of these proteins. Therefore, this study puts forward the above-mentioned proteins as valid participants in the vascular growth signals that are known to be triggered by ischemic preconditioning of heart. PMID- 15665049 TI - Alpha1-adrenoceptor-mediated phosphorylation of MYPT-1 and CPI-17 in the uterine artery: role of ERK/PKC. AB - We previously demonstrated that ERK/PKC signaling pathways play a key role in regulation of Ca(2+) sensitivity and contractility of the uterine artery. The present study tested the hypothesis that ERK and PKC differentially regulated myosin light chain phosphatase activity by phosphorylation of myosin phosphatase target protein-1 (MYPT-1) and CPI-17. Agonist-induced contractions and phosphorylation of MYPT-1/Thr(696), MYPT-1/Thr(850), and CPI-17/Thr(38) were measured simultaneously in the same tissues of isolated near-term pregnant ovine uterine arteries. Phenylephrine produced time-dependent concurrent increases in the phosphorylation of ERK(44/42) and MYPT-1/Thr(850) that preceded contractions. In addition, phenylephrine induced phosphorylation of CPI-17/Thr(38) that was concurrent with the contractions. In contrast, phenylephrine did not induce phosphorylation of MYPT-1/Thr(696) in the uterine artery. PD-098059 inhibited phosphorylation of ERK(44/42) and the initial peak phosphorylation of MYPT 1/Thr(850) but did not affect CPI-17/Thr(38) phosphorylation. Activation of PKC by phorbol 12,13-dibutyrate induced a time-dependent phosphorylation of CPI 17/Thr(38) that preceded contractions of the uterine artery. In addition, phorbol 12,13-dibutyrate activated PKC-alpha and induced a coimmunoprecipitation of PKC alpha with caldesmon. The results suggest that phosphorylation of MYPT-1/Thr(850) and CPI-17/Thr(38) play important roles in regulation of agonist-mediated Ca(2+) sensitivity in the uterine artery, in part by ERK and PKC, respectively. In addition, phosphorylated CPI-17 may regulate Ca(2+) sensitivity by interacting with caldesmon and reversing its inhibitory effect on myosin ATPase. PMID- 15665050 TI - Chronic nicotine in hearts with healed ventricular myocardial infarction promotes atrial flutter that resembles typical human atrial flutter. AB - The potential of chronic nicotine exposure for atrial fibrillation (AF) and atrial flutter (AFL) in hearts with and without chronic myocardial infarction (MI) remains poorly explored. MI was created in dogs by permanent occlusion of the left anterior descending coronary artery, and dogs were administered nicotine (5 mg.kg(-1).day(-1) sc) for 1 mo using osmotic minipumps. High-resolution epicardial (1,792 bipolar electrodes) and endocardial Halo catheters were used to map activation during induced atrial rhythms. Nicotine promoted inducible sustained AFL at a mean cycle length of 134 +/- 10 ms in all MI dogs (n = 6) requiring pacing and electrical shocks for termination. No AFL could be induced in MI dogs (n = 6), control (non-MI) dogs (n = 3) not exposed to nicotine, and dogs with no MI and exposed to nicotine (n = 3). Activation maps during AFL showed a single reentrant wavefront in the right atrium that rotated either clockwise (60%) or counterclockwise (40%) around the crista terminalis and through the isthmus. Ablation of the isthmus prevented the induction of AFL. Nicotine caused a significant (P < 0.01) but highly heterogeneous increase in atrial interstitial fibrosis (2- to 10-fold increase in left and right atria, respectively) in the MI group but only a 2-fold increase in the right atrium in the non-MI group. Nicotine also flattened (P < 0.05) the slope of the epicardial monophasic action potential duration (electrical restitution) curve of both atria in the MI but not in non-MI dogs. Two-dimensional simulation in an excitable matrix containing an isthmus and nicotine's restitutional and reduced gap junctional coupling (fibrosis) parameters replicated the experiments. Chronic nicotine in hearts with MI promotes AFL that closely resembles typical human AFL. Increased atrial interstitial fibrosis and flattened electrical restitution are important substrates for the AFL. PMID- 15665051 TI - CHIP, a cochaperone/ubiquitin ligase that regulates protein quality control, is required for maximal cardioprotection after myocardial infarction in mice. AB - Limitation of damage after ischemia and reperfusion injury to the myocardium remains an elusive clinical goal. Previous studies have suggested that molecular chaperones, which include members of the heat shock protein (Hsp) family, may have cardioprotective effects, although the protective role of endogenous chaperones has not been well documented. CHIP (carboxyl terminus of Hsp70 interacting protein) is a cochaperone/ubiquitin ligase that integrates the response to stress at multiple levels. We tested the response of CHIP(-/-) mice to in vivo ischemia and reperfusion injury induced by left anterior descending coronary artery ligation. Compared with wild-type littermates, CHIP(-/-) mice had decreased survival and increased incidence of arrhythmias during reperfusion. The size of myocardial infarction, as assessed by the ratio of infarct area to area at risk, was 50% greater in CHIP(-/-) mice. Increased infarct size was accompanied by impaired upregulation of the chaperone Hsp70 after ischemia reperfusion injury. In situ analysis also indicated that hearts of CHIP(-/-) mice were more prone to develop apoptosis in cardiomyocytes and especially endothelial cells of intramural vessels. Previous studies have found that CHIP plays a central role in maintaining protein quality control and coordinating the response to stress. The present data indicate that these functions of CHIP provide a critical cardioprotective effect in the setting of ischemia-reperfusion injury due in part to increased apoptosis in cardiac cells. Quality control mechanisms therefore may be underappreciated clinical targets for maximizing myocardial protection after injury. PMID- 15665052 TI - Platelet-activating factor increases endothelial [Ca2+]i and NO production in individually perfused intact microvessels. AB - We have demonstrated that inhibition of NO synthase (NOS) in endothelial cells by either the NOS inhibitor N(omega)-monomethyl-l-arginine (l-NMMA) or the internalization of caveolin-1 scaffolding domain attenuated platelet-activating factor (PAF)-induced increases in microvessel permeability (Am J Physiol Heart Circ Physiol 286: H195-H201, 2004) indicating the involvement of an NO-dependent signaling pathway. To investigate whether an increase in endothelial cytoplasmic Ca(2+) concentration ([Ca(2+)](i)) is the initiating event and Ca(2+)-dependent NO production is crucial for permeability increases, PAF (10 nM)-induced changes in endothelial [Ca(2+)](i) and NO production were measured in individually perfused rat mesenteric venular microvessels via fluorescence microscopy. When venular microvessels were exposed to PAF, endothelial [Ca(2+)](i) increased from 69 +/- 8 nM to a peak value of 374 +/- 26 nM within 3 min and then declined to a sustained level at 190 +/- 12 nM after 15 min. Inhibition of NOS did not modify PAF-induced increases in endothelial [Ca(2+)](i). PAF-induced NO production was visualized and quantified at cellular levels in individually perfused microvessels using 4,5-diaminofluorescein diacetate and fluorescence imaging. Increased fluorescence intensity (FI), which is an indication of increased NO production, occurred in 75 +/- 7% of endothelial cells in each vessel. The mean maximum FI increase was 140 +/- 7% of baseline value. This increased FI was abolished by pretreatment of the vessel with l-NMMA and attenuated in the absence of extracellular Ca(2+). These results provide direct evidence from intact microvessels that increased endothelial [Ca(2+)](i) is the initial signal that activates endothelial NOS, and the subsequent increased NO production contributes to PAF-induced increases in microvessel permeability. PMID- 15665053 TI - Impaired resting perfusion in viable myocardium distal to chronic coronary stenosis in rats. AB - Chronic coronary artery stenosis results in patchy necrosis in the dependent myocardium and impairs global and regional left ventricular (LV) function in rats in vivo. The aim of the present study was to compare regional myocardial blood flow (RMBF) and function (F) in poststenotic myocardium by using magnetic resonance imaging (MRI) and to compare MRI blood flow changes to histological alterations to assess whether RMBF in the viable poststenotic tissue remains normal. MRI was performed in 11 anesthetized Wistar rats with 2-wk stenosis of the left coronary artery. Postmortem, the extent of fibrotic tissue was quantified. Poststenotic RMBF was significantly reduced to 2.21 +/- 0.30 ml.g( 1).min(-1) compared with RMBF in the remote myocardium (4.05 +/- 0.50 ml.g( 1).min(-1)). A significant relationship between the poststenotic RMBF (%remote area) and the poststenotic F (%remote myocardium) was calculated (r = 0.61, P < 0.05). Assuming perfusion in scar tissue to be 32 +/- 5% of perfusion of remote myocardium, as measured in five additional rats, and that in remote myocardium to be 114 +/- 25% of that in normal myocardium, as assessed in five sham rats, the calculated perfusion in partially fibrotic tissue samples (35.7 +/- 5.2% of analyzed area) was 2.88 +/- 0.18 ml.g(-1).min(-1), whereas measured MRI perfusion was only 1.86 +/- 0.24 ml.g(-1).min(-1) (P < 0.05). These results indicate that resting perfusion in viable poststenotic myocardium is moderately reduced. Alterations in global and regional LV function are therefore secondary to both patchy fibrosis and reduced resting perfusion. PMID- 15665054 TI - Magnetic resonance stress tagging in ischemic heart disease. AB - High-dose dobutamine magnetic resonance stress testing has been shown to be superior to dobutamine stress echocardiography for diagnosis of coronary artery disease (CAD). We determined the feasibility of quantitative myocardial tagging during low- and high-dose dobutamine stress and tested the ability of global systolic and diastolic quantitative parameters to identify patients with significant CAD. Twenty-five patients suspected of having significant CAD were examined with a standard high-dose dobutamine/atropine stress magnetic resonance protocol (1.5-T scanner, Philips). All patients underwent invasive coronary angiography as the standard of reference for the presence (n = 13) or absence (n = 12) of significant CAD. During low-dose dobutamine stress, systolic (circumferential shortening, systolic rotation, and systolic rotation velocity) and diastolic (velocity of circumferential lengthening and diastolic rotation velocity) parameters changed significantly in patients without CAD (all P < 0.05 vs. rest) but not in patients with CAD. Identification of patients without and with CAD during low-dose stress was possible using the diastolic parameter of "time to peak untwist." At high-dose stress, none of the global systolic or diastolic parameters showed the potential to identify the presence of significant CAD. With myocardial tagging, a quantitative analysis of systolic and diastolic function was feasible during low- and high-dose dobutamine stress. In our study, the diastolic parameter of time to peak untwist as assessed during low-dose dobutamine stress was the most promising global parameter for identification of patients with significant CAD. Thus quantitative myocardial tagging may become a tool that reduces the need for high-dose dobutamine stress. PMID- 15665055 TI - Critical role of endothelial cell activation in hypoxia-induced vasoocclusion in transgenic sickle mice. AB - Activation of vascular endothelium plays an essential role in vasoocclusion in sickle cell disease. The anti-inflammatory agents dexamethasone and adhesion molecule-blocking antibodies were used to inhibit endothelial cell activation and hypoxia-induced vasoocclusion. Transgenic sickle mice, expressing human alpha-, beta(S)-, and beta(S-Antilles)-globins, had an activated vascular endothelium in their liver, lungs, and skin, as exhibited by increased activation of NF-kappaB compared with normal mice. NF-kappaB activation increased further in the liver and skin after sickle mice were exposed to hypoxia. Sickle mice had decreases in red blood cell (RBC) velocities and developed vasoocclusions in subcutaneous venules in response to hypoxia. Dexamethasone pretreatment prevented decreases in RBC velocities and inhibited vasoocclusions and leukocyte-endothelium interactions in venules after hypoxia. Dexamethasone treatment inhibited NF kappaB, VCAM-1, and ICAM-1 expression in the liver, lungs, and skin of sickle mice after hypoxia-reoxygenation. VCAM-1 or ICAM-1 blockade with monoclonal antibodies mimicked dexamethasone by inhibiting vasoocclusion and leukocyte adhesion in sickle mice, demonstrating that endothelial cell activation and VCAM 1 and ICAM-1 expression are necessary for hypoxia-induced vasoocclusion in sickle mice. VCAM-1, ICAM-1, and vasoocclusion increased significantly 3 days after dexamethasone discontinuation, possibly explaining rebounds in vasoocclusive crises observed after withdrawal of glucocorticosteroids in sickle patients. We conclude that anti-inflammatory treatments that inhibit endothelial cell activation and adhesion molecule expression can inhibit vasoocclusion in sickle cell disease. Rebounds in vasoocclusive crises after dexamethasone withdrawal are caused by rebounds in endothelial cell activation. PMID- 15665056 TI - Activation of Rho-associated kinase during augmented contraction of the basilar artery to serotonin after subarachnoid hemorrhage. AB - Delayed cerebral vasospasm after subarachnoid hemorrhage (SAH) may be due, in part, to altered regulation of arterial smooth muscle contraction. Contraction of cerebral arteries to serotonin is augmented after experimental SAH. We hypothesized that activation of Rho-associated kinase (Rho kinase) contributes to augmented contraction of cerebral arteries to serotonin after SAH. Autologous arterial blood (SAH) or artificial cerebrospinal fluid (control) was injected into the cisterna magna of anesthetized rabbits. At 2 days after injection, the basilar artery was excised and isometric contraction of arterial rings was recorded. Maximum contraction of the basilar artery to serotonin was augmented about fourfold in SAH compared with control rabbits (P < 0.01). Contraction to histamine was similar in the two groups. Fasudil hydrochloride (3 mumol/l), an inhibitor of Rho kinase, markedly attenuated serotonin-induced contraction. Fasudil had little effect on contractions induced by histamine or phorbol 12,13 dibutyrate. In addition, phosphorylation of myosin phosphatase, a major target of Rho kinase in regulation of smooth muscle contraction, in the basilar artery was examined by Western blotting. In basilar arteries of SAH, but not control, rabbits, serotonin increased phosphorylation of myosin phosphatase about twofold at Thr(853) of the myosin-targeting subunit. These results suggest that enhanced activation of Rho kinase contributes to augmented contraction of the basilar artery to serotonin after SAH. PMID- 15665057 TI - Protective effect of prenatal water restriction on offspring cardiovascular homeostasis in response to hemorrhage. AB - We determined the cardiovascular and AVP responses of prenatally dehydrated (PreDehy) neonates to intravascular hemorrhage. Ewes with singleton fetuses were subjected to water restriction from 110 days of gestation to full term to achieve hypernatremia of 8-10 meq/l. Water and food were provided ad libitum to control ewes. After delivery, water and food were provided ad libitum to ewes from both groups, and newborns were allowed to nurse ad libitum. At 15 +/- 2 days of age, PreDehy and control lambs were prepared with bladder and femoral catheters and studied at 25 +/- 2 days of age. After a 2-h basal period, lambs were hemorrhaged to 30% of blood volume over 1 h (0.5% of blood volume/min) and monitored 1 h after hemorrhage. Neonatal arterial blood pressure was measured, and blood samples were collected. Basal plasma sodium levels, plasma osmolality, hematocrit, and mean arterial pressure were increased in PreDehy lambs compared with controls. Both groups had similar basal AVP levels and heart rate. In response to hemorrhage, all parameters remained significantly elevated in PreDehy lambs. Blood pressure decreased less in PreDehy lambs than in controls. The hemorrhage-AVP threshold (percent blood volume withdrawal at which plasma AVP values significantly increased) was markedly elevated (20 vs. 15%) and peak hemorrhage-induced AVP plasma levels were lower (5.6 +/- 1.5 vs. 10.1 +/- 1.5 pg/ml, P < 0.01) in PreDehy lambs than in controls. Thus offspring of dehydrated ewes demonstrate enhanced AVP secretory responses to hypotension. Despite potential long-term adverse effects of systemic hypertension, these results suggest a protective effect of prenatal water restriction on offspring cardiovascular homeostasis during blood volume reduction. PMID- 15665058 TI - Regression of pressure overload-induced left ventricular hypertrophy in mice. AB - As a prelude to investigating the mechanism of regression of pressure overload induced left ventricular (LV) hypertrophy (LVH), we studied the time course for the development and subsequent regression of LVH as well as accompanying alterations in cardiac function, histology, and gene expression. Mice were subjected to aortic banding for 4 or 8 wk to establish LVH, and regression was initiated by release of aortic banding for 6 wk. Progressive increase in LV mass and gradual chamber dilatation and dysfunction occurred after aortic banding. LVH was also associated with myocyte enlargement, interstitial fibrosis, and enhanced expression of atrial natriuretic peptide, collagen I, collagen III, and matrix metalloproteinase-2 but suppressed expression of alpha-myosin heavy chain and sarcoplasmic reticulum Ca(2+)-ATPase. Aortic debanding completely or partially reversed LVH, chamber dilatation and dysfunction, myocyte size, interstitial fibrosis, and gene expression pattern, each with a distinct time course. The extent of LVH regression was dependent on the duration of pressure overload, evidenced by the fact that restoration of LV structure and function was complete in animals subjected to 4 wk of aortic banding but incomplete in animals subjected to 8 wk of aortic banding. In conclusion, LVH regression comprises a variety of morphological, functional, and genetic components that show distinct time courses. A longer period of pressure overload is associated with a slower rate of LVH regression. PMID- 15665059 TI - Estimating oxygen consumption rates of arteriolar walls under physiological conditions in rat skeletal muscle. AB - To examine the effects of vascular tone reduction on O2 consumption of the vascular wall, we determined the O2 consumption rates of arteriolar walls under normal conditions and during vasodilation induced by topical application of papaverine. A phosphorescence quenching technique was used to quantify intra- and perivascular PO2 in rat cremaster arterioles with different branching orders. Then, the measured radial PO2 gradients and a theoretical model were used to estimate the O2 consumption rates of the arteriolar walls. The vascular O2 consumption rates of functional arterioles were >100 times greater than those observed in in vitro experiments. The vascular O2 consumption rate was highest in first-order (1A) arterioles, which are located upstream, and sequentially decreased downstream in 2A and 3A arterioles under normal conditions. During papaverine-induced vasodilation, on the other hand, the O2 consumption rates of the vascular walls decreased to similar levels, suggesting that the high O2 consumption rates of 1A arterioles under normal conditions depend in part on the workload of the vascular smooth muscle. These results strongly support the hypothesis that arteriolar walls consume a significant amount of O2 compared with the surrounding tissue. Furthermore, the reduction of vascular tone of arteriolar walls may facilitate an efficient supply of O2 to the surrounding tissue. PMID- 15665060 TI - Preconditioning of salvaged myocardium in conscious rabbits with postinfarction dysfunction. AB - Protection against postinfarction myocardial dysfunction is modest with classic preconditioning (PC). We investigated whether multiple cycles of PC could improve this protection and whether postinfarction dysfunction only depends on the amount of viable tissue. Eighteen rabbits were chronically instrumented with coronary occluders and ultrasonic crystals (segment shortening, SH) in the ischemic zone. A control group underwent 30-min coronary artery occlusion (CAO) with 72-h reperfusion (CAR). In two other groups, PC was induced by six 4-min CAO/4-min CAR cycles (PCx6) or one 5-min CAO/10-min CAR cycle (PCx1). After 72-h CAR, depression in SH was reduced in PCx1 (-68 +/- 7% from baseline) and to a greater extent in PCx6 (-18 +/- 10%) vs. control (-99 +/- 7%; all P < 0.05). Infarct sizes were reduced in PCx1 (15 +/- 2%) and to a greater extent in PCx6 (3 +/- 1%) vs. control (46 +/- 5%; P < 0.05). Contractility of salvaged myocardium was evaluated by calculating the ratio between SH at 72-h CAR and the amount of viable tissue. This index was enhanced in PCx1 (0.39 +/- 0.07, P < 0.05) and to a greater extent in PCx6 (0.82 +/- 0.09) vs. control (0.0 +/- 0.10). This differential effect of PC was not related to changes in apoptosis, endothelial nitric oxide synthase (NOS) expression, or macrophages infiltration but, rather, to blunted inducible NOS expression in PCx6 vs. control and PCx1. Thus multiple cycles of PC induced an almost complete protection against postinfarction dysfunction, potentially involving beneficial effects on salvaged myocardium. PMID- 15665061 TI - Reduced voltage dependence of inactivation in the SCN5A sodium channel mutation delF1617. AB - Deletion of a phenylalanine at position 1617 (delF1617) in the extracellular linker between segments S3 and S4 in domain IV of the human heart Na(+) channel (hH1a) has been tentatively associated with long QT syndrome type 3 (LQT3). In a mammalian cell expression system, we compared whole cell, gating, and single channel currents of delF1617 with those of wild-type hH1a. The half points of the peak activation-voltage curve for the two channels were similar, as were the deactivation time constants at hyperpolarized test potentials. However, delF1617 demonstrated a significant negative shift of -7 mV in the half point of the voltage-dependent Na(+) channel availability curve compared with wild type. In addition, both the time course of decay of Na(+) current (I(Na)) and two-pulse development of inactivation of delF1617 were faster at negative test potentials, whereas they tended to be slower at positive potentials compared with wild type. Mean channel open times for delF1617 were shorter at potentials <0 mV, whereas they were longer at potentials >0 mV compared with wild type. Using anthopleurin A, a site-3 toxin that inhibits movement of segment S4 in domain IV (S4-DIV), we found that gating charge contributed by the S4-DIV in delF1617 was reduced 37% compared with wild type. We conclude that deletion of a single amino acid in the S3-S4 linker of domain IV alters the voltage dependence of fast inactivation via a reduction in the gating charge contributed by S4-DIV and can cause either a gain or loss of I(Na), depending on membrane potential. PMID- 15665062 TI - Effect of pravastatin on development of left ventricular hypertrophy in spontaneously hypertensive rats. AB - Endothelin (ET)-1 has been implicated in the development of cardiac hypertrophy. We investigated the effect of pravastatin on development of ventricular hypertrophy in spontaneously hypertensive rats (SHR) and whether the attenuated hypertrophic effect was via reduced ET-1 expression. Normolipidemic SHR were treated with one of the following therapies for 8 wk: vehicle, the nonselective ET receptor antagonists bosentan, pravastatin, mevalonate, hydralazine, or combination of pravastatin + mevalonate from the age of 8 wk at the very early stage of cardiac hypertrophy. Treatment with bosentan and pravastatin significantly decreased left ventricular mass index for body weight and cardiomyocyte sizes isolated by enzymatic dissociation. The myocardial ET-1 levels and preproET-1 mRNA assessed using real-time quantitative RT-PCR were significantly higher (both P < 0.001) in the SHR compared with Wistar-Kyoto rats. The increased tissue ET-1 levels can be inhibited after pravastatin administration. Immunohistochemical analysis confirmed the changes of ET-1. Left ventricular mass index for body weight correlated positively with tissue ET-1 levels (P = 0.0004). A dissociation between the effects of blood pressure and cardiac structure was noted, because pravastatin and hydralazine reduced arterial pressure similarly. Pravastatin-induced effects were reversed by the addition of mevalonate. In conclusion, these results suggest a crucial role of cardiac endothelin system in the early development of ventricular hypertrophy in the SHR. Pravastatin is endowed with cardiac antihypertropic properties that are independent of its hemodynamic and hypolipidemic effects and appear to be related to their capacity to decrease cardiac ET-1 levels, which is linked to mevalonate metabolism. PMID- 15665063 TI - Sympathetic innervation promotes vascular smooth muscle differentiation. AB - The sympathetic nervous system (SNS) is an important modulator of vascular smooth muscle (VSM) growth and function. Several lines of evidence suggest that the SNS also promotes VSM differentiation. The present study tests this hypothesis. Expression of smooth muscle myosin (SM2) and alpha-actin were assessed by Western analysis as indexes of VSM differentiation. SM2 expression (normalized to alpha actin) in adult innervated rat femoral and tail arteries was 479 +/- 115% of that in noninnervated carotid arteries. Expression of alpha-actin (normalized to GAPDH or total protein) in 30-day-innervated rat femoral arteries was greater than in corresponding noninnervated femoral arteries from guanethidine-sympathectomized rats. SM2 expression (normalized to alpha-actin) in neonatal femoral arteries grown in vitro for 7 days in the presence of sympathetic ganglia was greater than SM2 expression in corresponding arteries grown in the absence of sympathetic ganglia. In VSM-endothelial cell cultures grown in the presence of dissociated sympathetic neurons, alpha-actin (normalized to GAPDH) was 300 +/- 66% of that in corresponding cultures grown in the absence of neurons. This effect was inhibited by an antibody that neutralized the activity of transforming growth factor-beta2. All of these data indicate that sympathetic innervation increased VSM contractile protein expression and thereby suggest that the SNS promotes and/or maintains VSM differentiation. PMID- 15665064 TI - PPAR-alpha activation required for decreased glucose uptake and increased susceptibility to injury during ischemia. AB - The transcription of key metabolic regulatory enzymes in the heart is altered in the diabetic state, yet little is known of the underlying mechanisms. The aim of this study was to investigate the role of peroxisome proliferator-activated receptor-alpha (PPAR-alpha) in modulating cardiac insulin-sensitive glucose transporter (GLUT-4) protein levels in altered metabolic states and to determine the functional consequences by assessing cardiac ischemic tolerance. Wild-type and PPAR-alpha-null mouse hearts were isolated and perfused 6 wk after streptozotocin administration or after 14 mo on a high-fat diet or after a 24-h fast. Myocardial d-[2-(3)H]glucose uptake was measured during low-flow ischemia, and differences in GLUT-4 protein levels were quantified using Western blotting. In wild-type mice in all three metabolic states, elevated plasma free fatty acids were associated with lower total cardiac GLUT-4 protein levels and decreased glucose uptake during ischemia, resulting in poor postischemic functional recovery. Although PPAR-alpha-null mice also had elevated plasma free fatty acids, they had neither decreased cardiac GLUT-4 levels nor decreased glucose uptake during ischemia and, consequently, did not have poor recovery during reperfusion. We conclude that elevated plasma free fatty acids are associated with increased injury during ischemia due to decreased cardiac glucose uptake resulting from lower cardiac GLUT-4 protein levels, the levels of GLUT-4 being regulated, probably indirectly, through PPAR-alpha activation. PMID- 15665065 TI - Theory-based identification of barriers to quality improvement: induced abortion care. AB - BACKGROUND: The UK Royal College of Obstetricians and Gynaecologists published the clinical guideline, The Care of Women Requesting Induced Abortion, to address recognized variations in care. There is little empirical evidence on factors that influence compliance with the guideline. A better understanding of such factors is needed for quality improvement initiatives. OBJECTIVE: To identify factors that influence compliance with two key guideline recommendations: offer of an assessment appointment within 5 days of referral, and supply of contraceptives at discharge. SETTING: Thirteen hospital gynaecology units in Scotland. METHODS: Guideline compliance was measured by a case note review. Barriers and facilitators were identified using a combination of approaches: semi-structured interviews with local gynaecologists and a survey of clinical staff. The questionnaire, based upon constructs from the Theory of Planned Behaviour, measured behavioural intention, attitude, subjective norm (perceived social pressure), and perceived behavioural control. RESULTS: Of 507 cases reviewed, median unit compliance was 46% for the assessment appointment and 59% for contraceptive supplies. Questionnaires were returned by 151 (74%) of 205 staff in 12 units. The interviews and open-ended questions highlighted organizational barriers to guideline implementation. Staff generally had strong intentions and positive attitudes to follow both recommendations. For the assessment appointment, perceived behavioural control was low. The Theory of Planned Behaviour accounted for 27% of the variation in intentions, with subjective norm being the strongest predictor. Intention and perceived behavioural control best explained unit compliance, together explaining 15% of the variation. For contraceptive supplies, the theory accounted for 34% of the variation in intentions, with perceived behavioural control being the strongest predictor. CONCLUSION: Clinical staff were highly motivated to implement the guideline but hindered by organizational constraints. Quality improvement initiatives need to target organizational barriers as well as individual professionals. PMID- 15665066 TI - Integrated care programmes for chronically ill patients: a review of systematic reviews. AB - OBJECTIVE: To investigate effectiveness, definitions, and components of integrated care programmes for chronically ill patients on the basis of systematic reviews. DESIGN: Literature review from January 1996 to May 2004. MAIN MEASURES: Definitions and components of integrated care programmes and all effects reported on the quality of care. RESULTS: Searches in the Medline and Cochrane databases identified 13 systematic reviews of integrated care programmes for chronically ill patients. Despite considerable heterogeneity in interventions, patient populations, and processes and outcomes of care, integrated care programmes seemed to have positive effects on the quality of patient care. No consistent definitions were present for the management of patients with chronic illnesses. In all the reviews the aims of integrated care programmes were very similar, namely reducing fragmentation and improving continuity and coordination of care, but the focus and content of the programmes differed widely. The most common components of integrated care programmes were self-management support and patient education, often combined with structured clinical follow-up and case management; a multidisciplinary patient care team; multidisciplinary clinical pathways and feedback, reminders, and education for professionals. CONCLUSION: Integrated care programmes seemed to have positive effects on the quality of care. However, integrated care programmes have widely varying definitions and components and failure to recognize these variations leads to inappropriate conclusions about the effectiveness of these programmes and to inappropriate application of research results. To compare programmes and better understand the (cost) effectiveness of the programmes, consistent definitions must be used and component interventions must be well described. PMID- 15665067 TI - Waiting for orthopaedic surgery: factors associated with waiting times and patients' opinion. AB - OBJECTIVE: To assess waiting times for three groups of orthopaedic patients in Sweden and to identify factors explaining variations in waiting time. Also examined were factors associated with patients' perceptions that waiting times were too long. DESIGN: Retrospective study. SETTING AND STUDY PARTICIPANTS: Patients from orthopaedic units at 10 Swedish hospitals participated in the study. A questionnaire was sent to 1336 surgical patients (517 hip replacement, 321 back surgery, and 498 arthroscopic knee surgery) 3 months after surgery. Information extracted from the hospitals' patient administrative systems was also used. Outcome measures. Length of waiting time, socio-economic variables, hospital type, health-related quality of life, and opinion about waiting time. The data were analysed mainly using regression analyses. RESULTS: The overall response rate was 79%. In all pre-operative stages, waiting times were longest in the hip replacement group. Socio-economic variables were not consistent determinants of variation in waiting times except for working status in the back surgery group where working patients had shorter waiting times than non-working patients irrespective of phase of waiting time. Admission to a county/district county hospital, compared with a university/regional hospital, was associated with shorter time on the waiting list. Patients with better health-related quality of life had significantly longer waiting times for arthroscopic knee surgery by all waiting time measures. The length of wait was a significant predictor of the patients' acceptance of waiting time. Patients' influence over the date of surgery also appeared to affect their opinion about the waiting time. CONCLUSIONS: Hospital-related factors are more important than patient characteristics as explanations of variations in waiting times for orthopaedic surgery. Patients value short waiting times and the possibility of influencing the date of surgery. PMID- 15665068 TI - Predictors of high quality clinical practice guidelines: examples in oncology. AB - OBJECTIVE: Clinical practice guidelines are widely used as effective tools for improving the management of patients with cancer. However, there is increasing concern about variation in guideline quality. In this study we identified predictors for high-quality guidelines in oncology. DESIGN: The quality scores for 32 oncology guidelines from 13 countries were determined by four independent appraisers using the Appraisal of Guidelines for Research and Evaluation (AGREE) instrument. MAIN MEASURE: : The contribution to the quality score of six characteristics of guidelines and three of guideline developing organizations was then assessed using analysis of variance and stepwise linear regression analysis. RESULTS: Some guideline and organizational characteristics were shown to be responsible for a large part of the variations in quality scores. The availability of background information was the strongest predictor of quality with an explained variance ranging from 17% ("Applicability") to 67% ("Rigour of development"). High-quality guidelines were more often produced by government supported organizations and/or within a structured, coordinated programme. The other characteristics (publication year, type of guideline, format, level of care, and scope) were not independent predictors of quality. CONCLUSIONS: Guidelines should provide more explicit information about the context of their development and methods used in order to improve their quality and thus encourage their use in clinical practice. PMID- 15665069 TI - Development of a set of strategy-based system-level cancer care performance indicators in Ontario, Canada. AB - OBJECTIVES: To develop a set of scientifically sound and managerially useful system-level cancer care performance indicators for public reporting in Ontario, Canada. IMPLEMENTATION: Using a modified Delphi panel method, comprising a systematic literature review and multiple rounds of structured feedback from 34 experts, the Cancer Quality Council of Ontario developed a set of quality indicators spanning cancer prevention through to end-of-life care. To be useful to decision-makers and providers, indicator selection criteria included a clear focus on the cancer system, relevance to a diversity of cancer providers, a strong link to the mission and strategic objectives of the cancer system, clear directionality of indicator results, presence of targets and/or benchmarks, feasibility of populating the indicator, and credibility of the measure as an indicator of quality. To ensure that the selected indicators would measure progress over time against specific and widely accepted goals, we created a strategy map based on the five strategic objectives of the Ontario cancer system: (i) to improve the measurement and reporting of cancer quality, (ii) to increase the use of evidence and innovation in decision-making, (iii) to improve access to cancer services and reduce waiting times, (iv) to increase efficiency across the system, (v) to reduce the burden of cancer. An analysis of the mean indicator ratings by experts, and the strategy mapping exercise resulted in the identification of 36 indicators deemed suitable for routine performance measurement of the Ontario cancer system. LESSONS LEARNED: The resulting instrument incorporates a credible evidence basis for performance measurement aligned to the five strategic goals for the Ontario cancer system. It represents the integrating of a management culture, focused on the implementation of a new strategic direction for the cancer system, with the underlying evidence-based culture of clinicians. PMID- 15665070 TI - Relationship between Bispectral Index, electroencephalographic state entropy and effect-site EC50 for propofol at different clinical endpoints. AB - BACKGROUND: State entropy (SE) is a newly available monitor for depth of anaesthesia. We investigated whether the relationship between predicted effect site propofol concentration and Bispectral Index (BIS) and SE values is useful for predicting loss of verbal contact and loss of consciousness during steady state conditions. METHODS: Twenty unpremedicated patients undergoing elective major abdominal surgery were recruited. A target-controlled infusion of propofol was administered using Schneider's pharmacokinetic model. The propofol infusion was set at an initial site effect concentration of 1.0 microg ml(-1) and increased by 1.0 microg ml(-1) steps every 4 min up to 6.0 microg ml(-1). A 4-min interval was chosen to ensure that steady-state effect-site concentrations were obtained. Propofol site effect concentrations and BIS and SE values were recorded at loss of verbal contact (LVC) and loss of consciousness (LOC). Population values for predicted effect-site concentrations at the clinical endpoints were estimated and correlated with BIS and SE values. RESULTS: For LVC, the effect site concentration for 90% of patients was 1.1 (1.1-3.2) microg ml(-1) and for LOC it was 2.8 (2.8-5.65) microg ml(-1). LVC occurred in 90% of patients at a BIS value of 70.2 (70.2-90.2) and an SE value of 60.3 (60.3-75.5), and LOC occurred at a BIS value of 38.2 (38.2-70.4) and an SE value of 42.2 (42.2-60.4). CONCLUSIONS: LVC and LOC occurred within a defined range of predicted effect-site concentrations. SE had a smaller range than BIS and greater correlation with effect-site concentration and may be more useful than BIS in predicting both LVC and LOC. PMID- 15665071 TI - Hypoalbuminaemia does not impair Diprifusor performance during sedation with propofol. AB - BACKGROUND: About 98% of plasma propofol is bound to albumin. We investigated if severe hypoalbuminaemia may affect the accuracy of a target-controlled infusion (TCI) device, the Diprifusor, during sedation in critically ill patients. METHODS: Ten critically ill hypoalbuminaemic patients (<24 g litre(-1)) and 10 critically ill normoalbuminaemic patients (>32 g litre(-1)) were included in this study. They underwent sedation with propofol, aimed at a Ramsey sedation score of 4-5. The Diprifusor was used to achieve target propofol plasma concentrations that ranged between 0.6 and 1.5 mg litre(-1). Propofol concentration was measured by high-performance liquid chromatography 5 min, 15 min, 30 min, 1 h, 2 h, 4 h, 6 h and 8 h after starting TCI. The accuracy of TCI was evaluated by calculating performance errors [PE=100x(measured concentration-predicted concentration)/predicted concentration], absolute and relative individual median performance errors (MDAPE and MDPE) and divergence (the slope of individual regression lines between PEs and time). RESULTS: PEs [median (range)] were -7 ( 65, 79) in hypoalbuminaemic patients and -2 (-53, 188) in normoalbuminaemic patients; absolute PEs were 21 (1, 79) and 22 (0, 188). No significant difference was observed between the two groups. MDPE, MDAPE and divergence values were also similar. In most patients the accuracy of TCI increased with time because higher PE values were observed during the first 30 min. CONCLUSIONS: Hypoalbuminaemia does not affect the accuracy of Diprifusor during sedation with propofol in critically ill patients. PMID- 15665072 TI - Perioperative myocardial infarction--aetiology and prevention. AB - Perioperative myocardial infarction (PMI) is one of the most important predictors of short- and long-term morbidity and mortality associated with non-cardiac surgery. Prevention of a PMI is thus a prerequisite for an improvement in overall postoperative outcome. The aetiology of PMI is multifactorial. The perioperative period induces large, unpredictable and unphysiological alterations in coronary plaque morphology, function and progression, and may trigger a mismatch of myocardial oxygen supply and demand. With many diverse factors involved, it is unlikely that one single intervention will successfully improve cardiac outcome following non-cardiac surgery. A multifactorial, step-wise approach is indicated. Based on increasing knowledge of the nature of atherosclerotic coronary artery disease, and in view of the poor positive predictive value of non-invasive cardiac stress tests, and the considerable risk of coronary angiography and coronary revascularization in high-risk patients, the paradigm is shifting from an emphasis on extensive non-invasive preoperative risk stratification to a combination of selective non-invasive testing and aggressive pharmacological perioperative therapy. Perioperative plaque stabilization by pharmacological means may be as important in the prevention of PMI as an increase in myocardial oxygen supply or a reduction in myocardial oxygen demand. PMID- 15665073 TI - Reduced activation of immunomodulatory transcription factors during positive end expiratory pressure adjustment based on volume-dependent compliance in isolated perfused rabbit lungs. AB - BACKGROUND: Repeated alveolar collapse and cyclic alveolar overdistension with associated activation of inflammatory signalling cascades contribute to ventilator-induced lung injury (VILI). The appropriate positive end-expiratory pressure (PEEP) which prevents or ameliorates VILI is unknown. In the isolated perfused lung, repeated adjustments of PEEP based on the continuously analysed intratidal compliance-volume curve have previously been shown to result in full end-expiratory alveolar recruitment and low risk of cyclic alveolar overdistension. Accordingly, we tested the hypothesis that such ventilatory management reduces intrapulmonary activation of the immunomodulatory transcription factors nuclear factor kappaB (NF-kappaB), activator protein 1 (AP 1) and cAMP-responsive element binding protein (CREB) which induce the expression of various chemokines and cytokines. METHODS: Isolated perfused rabbit lungs were randomly allocated to one of three groups: zero end-expiratory pressure (ZEEP) to induce repeated alveolar collapse (n=6), high PEEP to induce cyclic alveolar overdistension (n=6) and repeated PEEP adjustments based on intratidal compliance volume curve analysis by the slice method to minimize repeated alveolar collapse and overdistension (n=9). All lungs were ventilated with a tidal volume of 6 ml kg(-1) bodyweight for 120 min. Thereafter, activation of transcription factors NF kappaB, AP-1 and CREB in lung tissue was analysed by electrophoretic mobility shift assay. RESULTS: High PEEP was associated with the highest activation of NF kappaB and AP-1 and repeated PEEP adjustments with the lowest activation when compared with the other two study groups (P<0.001). In contrast, activation of CREB did not differ between groups. Activated NF-kappaB and AP-1 protein complexes consisted mainly of the transactivators p50/p65 and c-Fos/Jun, respectively. CONCLUSIONS: In isolated perfused rabbit lungs, repeated adjustments of PEEP based on the continuously analysed intratidal compliance volume curve were associated with less activation of early steps of inflammatory signalling cascades than ventilation with ZEEP or high PEEP. PMID- 15665074 TI - A peptide inhibitor of cytochrome c/inositol 1,4,5-trisphosphate receptor binding blocks intrinsic and extrinsic cell death pathways. AB - Apoptotic stimuli augment intracellular calcium concentration through inositol 1,4,5-trisphosphate receptors (IP3R) on endoplasmic reticulum calcium stores. We previously discovered an apoptotic cascade wherein cytochrome c binds to IP3R early in apoptosis, resulting in dysregulated calcium release. Here we show that cytochrome c binding to IP3R depends on a cluster of glutamic acid residues within the C terminus of the channel. A cell permeant peptide derived from this sequence displaces cytochrome c from IP3R and abrogates cell death induced by staurosporine treatment of HeLa cells and Fas ligand stimulation of Jurkat cells. Small-molecule inhibitors of cytochrome c/IP3R interactions may prove useful in treating disorders associated with inappropriate intrinsic and extrinsic apoptotic signaling. PMID- 15665075 TI - The diversity of dolichol-linked precursors to Asn-linked glycans likely results from secondary loss of sets of glycosyltransferases. AB - The vast majority of eukaryotes (fungi, plants, animals, slime mold, and euglena) synthesize Asn-linked glycans (Alg) by means of a lipid-linked precursor dolichol PP-GlcNAc2Man9Glc3. Knowledge of this pathway is important because defects in the glycosyltransferases (Alg1-Alg12 and others not yet identified), which make dolichol-PP-glycans, lead to numerous congenital disorders of glycosylation. Here we used bioinformatic and experimental methods to characterize Alg glycosyltransferases and dolichol-PP-glycans of diverse protists, including many human pathogens, with the following major conclusions. First, it is demonstrated that common ancestry is a useful method of predicting the Alg glycosyltransferase inventory of each eukaryote. Second, in the vast majority of cases, this inventory accurately predicts the dolichol-PP-glycans observed. Third, Alg glycosyltransferases are missing in sets from each organism (e.g., all of the glycosyltransferases that add glucose and mannose are absent from Giardia and Plasmodium). Fourth, dolichol-PP-GlcNAc2Man5 (present in Entamoeba and Trichomonas) and dolichol-PP- and N-linked GlcNAc2 (present in Giardia) have not been identified previously in wild-type organisms. Finally, the present diversity of protist and fungal dolichol-PP-linked glycans appears to result from secondary loss of glycosyltransferases from a common ancestor that contained the complete set of Alg glycosyltransferases. PMID- 15665076 TI - Increased activity of cyclin-dependent kinase 5 leads to attenuation of cocaine mediated dopamine signaling. AB - Cocaine, a drug of abuse, increases synaptic dopamine levels in the striatum by blocking dopamine reuptake at axon terminals. Cyclin-dependent kinase 5 (Cdk5) and its activator p35, proteins involved in phosphorylation of substrates in postmitotic neurons, have been found to be up-regulated after chronic exposure to cocaine. To further examine the effects of Cdk5 and p35 induction on striatal dopamine signaling, we generated two independent transgenic mouse lines in which Cdk5 or p35 was overexpressed specifically in neurons. We report here that increased Cdk5 activity, as a result of p35 but not of Cdk5 overexpression, leads to attenuation of cocaine-mediated dopamine signaling. Increased Cdk5-mediated phosphorylation of dopamine and cAMP-regulated phosphoprotein, molecular mass 32 kDa (DARPP-32) at Thr-75, was accompanied by decreased phosphorylation of DARPP 32 at Thr-34. Increased Cdk5-mediated phosphorylation of extracellular signal regulated kinase kinase 1 at Thr-286 was accompanied by decreased activation of extracellular signal-regulated kinase 1/2. These effects contributed to attenuation of cocaine-induced phosphorylation of cAMP response element-binding protein as well as a lesser induction of c-fos in the striatum. These results support the idea that Cdk5 activity is involved in altered gene expression after chronic exposure to cocaine and hence impacts the long-lasting changes in neuronal function underlying cocaine addiction. PMID- 15665077 TI - BDNF modulates GABAA receptors microtransplanted from the human epileptic brain to Xenopus oocytes. AB - Cell membranes isolated from brain tissues, obtained surgically from six patients afflicted with drug-resistant temporal lobe epilepsy and from one nonepileptic patient afflicted with a cerebral oligodendroglioma, were injected into frog oocytes. By using this approach, the oocytes acquire human GABAA receptors, and we have shown previously that the "epileptic receptors" (receptors transplanted from epileptic brains) display a marked run-down during repetitive applications of GABA. It was found that exposure to the neurotrophin BDNF increased the amplitude of the "GABA currents" (currents elicited by GABA) generated by the epileptic receptors and decreased their run-down; both events being blocked by K252A, a neurotrophin tyrosine kinase receptor B inhibitor. These effects of BDNF were not mimicked by nerve growth factor. In contrast, the GABAA receptors transplanted from the nonepileptic human hippocampal uncus (obtained during surgical resection as part of the nontumoral tissue from the oligodendroglioma margins) or receptors expressed by injecting rat recombinant alpha1beta2gamma2 GABAA receptor subunit cDNAs generated GABA currents whose time-course and run down were not altered by BDNF. Loading the oocytes with the Ca2+ chelator 1,2 bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetate-acetoxymethyl ester (BAPTA-AM), or treating them with Rp-8-Br-cAMP, an inhibitor of the cAMP-dependent PKA, did not alter the GABA currents. However, staurosporine (a broad spectrum PK inhibitor), bisindolylmaleimide I (a PKC inhibitor), and U73122 (a phospholipase C inhibitor) blocked the BDNF-induced effects on the epileptic GABA currents. Our results indicate that BDNF potentiates the epileptic GABAA currents and antagonizes their use-dependent run-down, thus strengthening GABAergic inhibition, probably by means of activation of tyrosine kinase receptor B receptors and of both PLC and PKC. PMID- 15665078 TI - Differential proton sensitivity of related G protein-coupled receptors T cell death-associated gene 8 and G2A expressed in immune cells. AB - G2A, T cell death-associated gene 8 (TDAG8), ovarian cancer G protein-coupled receptor 1 (OGR1), and G protein-coupled receptor 4 (GPR4) form a group of structurally related G protein-coupled receptors (GPCRs) originally proposed to bind proinflammatory lipids. More recent studies have challenged the identification of lipid agonists for these GPCRs and have suggested that they function primarily as proton sensors. We compared the ability of these four receptors to modulate pH-dependent responses by using transiently transfected cell lines. In accordance with previously published reports, OGR1 was found to evoke strong pH-dependent responses as measured by inositol phosphate accumulation. We also confirmed the pH-dependent cAMP production by GPR4 and TDAG8. However, we found the activity of the human G2A receptor and its mouse homolog to be significantly less sensitive to pH fluctuations as measured by inositol phosphate and cAMP accumulation. Sequence homology analysis indicated that, with one exception, the histidine residues that were previously shown to be important for pH sensing by OGR1, GPR4, and TDAG8 were not conserved in the G2A receptor. We further addressed the pH-sensing properties of G2A and TDAG8 in a cellular context where these receptors are coexpressed. In thymocytes and splenocytes explanted from receptor-deficient mice, TDAG8 was found to be critical for pH-dependent cAMP production. In contrast, G2A was found to be dispensable for this process. We conclude that members of this GPCR group exhibit differential sensitivity to extracellular protons, and that expression of TDAG8 by immune cells may regulate responses in acidic microenvironments. PMID- 15665079 TI - Inositol pyrophosphates regulate cell death and telomere length through phosphoinositide 3-kinase-related protein kinases. AB - Inositol pyrophosphates physiologically regulate vesicular endocytosis, ribosomal disposition, and directly phosphorylate proteins. Here we demonstrate roles in cell death and regulation of telomere length. Lethal actions of wortmannin and caffeine are selectively abolished in yeast mutants that cannot synthesize inositol pyrophosphates. Wortmannin and caffeine appear to act through the phosphoinositide 3-kinase-related protein kinases Tel1 and Mec1, known regulators of telomere length. Inositol pyrophosphates physiologically antagonize the actions of these kinases, which is demonstrated by the fact that yeast mutants with reduced or elevated levels of inositol pyrophosphates, respectively, display longer and shorter telomeres. PMID- 15665080 TI - A cascade of cytokines mediates mechanical inflammatory hypernociception in mice. AB - The hypernociceptive effects of cytokines [TNF-alpha, keratinocyte-derived chemokine (KC), and IL-1beta] and their participation in carrageenan (Cg)-induced inflammatory hypernociception in mice were investigated. Nociceptor sensitization (hypernociception) was quantified with an electronic version of the von Frey filament test in WT and TNF receptor type 1 knockout mice (TNF-R1-/-). TNF-alpha induced hypernociception was abolished in TNF-R1-/- mice, partially inhibited by pretreatment with IL-1 receptor antagonist (IL-1ra) or indomethacin and unaffected by Ab against KC (AbKC) or guanethidine. IL-1ra and indomethacin pretreatment strongly inhibited the hypernociception induced by IL-1beta, which was not altered by AbKC or guanethidine or by knocking out TNF-R1. KC-induced hypernociception was abolished by AbKC, inhibited by pretreatment with indomethacin plus guanethidine, and partially inhibited by IL-1ra, indomethacin, or guanethidine. In contrast, KC-induced hypernociception was not altered by knocking out TNF-R1. Cg-induced hypernociception was abolished by administration of indomethacin plus guanethidine, diminished in TNF-R1-/- mice, and partially inhibited in WT mice pretreated with AbKC, IL-1ra, indomethacin, or guanethidine. TNF-alpha, KC, and IL-1beta concentrations were elevated in the skin of Cg injected paws. The TNF-alpha and KC concentrations rose concomitantly and peaked before that of IL-1beta. In mice, the cytokine cascade begins with the release of TNF-alpha (acting on TNF-R1 receptor) and KC, which stimulate the release of IL 1beta. As in rats, the final mediators of this cascade were prostaglandins released by IL-1beta and sympathetic amines released by KC. These results extend to mice the concept that the release of primary mediators responsible for hypernociception is preceded by a cascade of cytokines. PMID- 15665081 TI - Fast and reliable prediction of noncoding RNAs. AB - We report an efficient method for detecting functional RNAs. The approach, which combines comparative sequence analysis and structure prediction, already has yielded excellent results for a small number of aligned sequences and is suitable for large-scale genomic screens. It consists of two basic components: (i) a measure for RNA secondary structure conservation based on computing a consensus secondary structure, and (ii) a measure for thermodynamic stability, which, in the spirit of a z score, is normalized with respect to both sequence length and base composition but can be calculated without sampling from shuffled sequences. Functional RNA secondary structures can be identified in multiple sequence alignments with high sensitivity and high specificity. We demonstrate that this approach is not only much more accurate than previous methods but also significantly faster. The method is implemented in the program rnaz, which can be downloaded from www.tbi.univie.ac.at/~wash/RNAz. We screened all alignments of length n > or = 50 in the Comparative Regulatory Genomics database, which compiles conserved noncoding elements in upstream regions of orthologous genes from human, mouse, rat, Fugu, and zebrafish. We recovered all of the known noncoding RNAs and cis-acting elements with high significance and found compelling evidence for many other conserved RNA secondary structures not described so far to our knowledge. PMID- 15665082 TI - Exposure of acidic residues as a danger signal for recognition of fibrinogen and other macromolecules by integrin alphaXbeta2. AB - The structural integrity of tissue proteins is damaged in processes ranging from remodeling of the extracellular matrix to destruction by microbial pathogens. Leukocytes play a prominent role in tissue surveillance and repair. However, it remains enigmatic what features of structurally decayed proteins prompt recognition by leukocyte cell-surface receptors. Here, we report that adhesion of human neutrophil granulocytes to fibrinogen is greatly increased by plasmin digestion in a mode where alphaXbeta2 dominates the integrin-dependent binding. The bacterial protease subtilisin also enhances binding by alphaXbeta2. The alphaX ligand binding domain has an unusually high affinity for carboxyl groups, with KD at approximately 100 microM. Our findings implicate enhanced accessibility of negatively charged residues in structurally decayed proteins as a pattern recognition motif for alphaXbeta2 integrin. Comparisons among integrins show relevance of these findings to the large number of ligands recognized by alphaMbeta2 and alphaXbeta2 but not alphaLbeta2. The observations suggest that the pericellular proteolysis at the leading edge of neutrophils not only facilitates passage through the extracellular matrix but also manufactures binding sites for alphaXbeta2. PMID- 15665083 TI - Peptide 15-mers of defined sequence that substitute for random amino acid copolymers in amelioration of experimental autoimmune encephalomyelitis. AB - Myelin basic protein (MBP) is a major candidate autoantigen in multiple sclerosis (MS). Its immunodominant epitope, MBP 85-99, forms a complex with human leukocyte antigen (HLA)-DR2 with which multiple sclerosis is genetically associated. Copolymer 1 (Copaxone), a random amino acid copolymer [poly (Y,E,A,K)n] as well as two modified synthetic copolymers [poly (F,Y,A,K)n and poly (V,W,A,K)n] also form complexes with HLA-DR2 (DRA/DRB1*1501) and compete with MBP 85-99 for binding. Moreover, two high-affinity synthetic peptide 15-mers that could inhibit binding even more effectively were previously designed. Here, we show that further-modified peptide 15-mers inhibited even more strongly (in order J5 > J3 > J2) both the binding of MBP 85-99 to HLA-DR2 and IL-2 production by two MBP 85-99 specific HLA-DR2-restricted T cells. J5, J3, and J2 also suppressed both MBP 85 99-induced experimental autoimmune encephalomyelitis (EAE) in humanized mice and proteolipid protein 139-151-induced EAE in SJL/J mice. Moreover, none of these previously uncharacterized peptide inhibitors crossreacted with MBP 85-99- or proteolipid protein 139-151-specific T cells. In both cases, spleen and lymph node cultures stimulated with these peptides produced large amounts of Th2 cytokines (IL-4 and IL-10), and adoptive transfer of established T cell lines suppressed disease induction. These peptide 15-mers provide specific, nonrandom sequences that appear to be at least as effective as random copolymers in suppressing EAE in several models. PMID- 15665084 TI - Genes for direct methylation of glycine provide high levels of glycinebetaine and abiotic-stress tolerance in Synechococcus and Arabidopsis. AB - Betaine is an important osmoprotectant, synthesized by many plants in response to abiotic stresses. Almost all known biosynthetic pathways of betaine are two-step oxidations of choline. Recently, a biosynthetic pathway of betaine from glycine, catalyzed by two N-methyltransferase enzymes, was found. Here, the potential role of N-methyltransferase genes for betaine synthesis was examined in a freshwater cyanobacterium, Synechococcus sp. PCC 7942, and in Arabidopsis plants. It was found that the coexpression of N-methyltransferase genes in Synechococcus caused accumulation of a significant amount of betaine and conferred salt tolerance to a freshwater cyanobacterium sufficient for it to become capable of growth in seawater. Arabidopsis plants expressing N-methyltransferase genes also accumulated betaine to a high level in roots, stems, leaves, and flowers and improved seed yield under stress conditions. Betaine levels were higher than those produced by choline-oxidizing enzymes. These results demonstrate the usefulness of glycine N-methyltransferase genes for the improvement of abiotic stress tolerance in crop plants. PMID- 15665085 TI - T-bet deficiency reduces atherosclerosis and alters plaque antigen-specific immune responses. AB - The influence of the immune system on atherosclerosis involves both helper T (Th) cell and antibody responses to plaque antigens. These responses may have proatherogenic and protective effects. T-bet is a transcription factor required for Th1 differentiation and regulates the balance between Th1 and Th2 responses in inflammatory diseases. To clarify how helper T cell subset differentiation influences atherosclerosis, we compared lesion development and immune responses to plaque antigens in low-density lipoprotein receptor-deficient (Ldlr-/-) mice with or without functional T-bet genes. Atherosclerosis was significantly reduced in T-bet-deficient Ldlr-/- mice compared with Ldlr-/- controls, and the lesions that did develop in the absence of T-bet had less smooth muscle cell content. Furthermore, T-bet deficiency caused a Th2 switch in the response to the atherosclerosis-associated antigen heat shock protein-60, and a change in T dependent isotypes of oxidized LDL-specific antibodies. Of particular significance, T-bet deficiency caused a >250% increase in the titer of E06 antibodies, which are known to be atheroprotective and whose production by B-1 B cells is enhanced by IL-5. These findings establish that T cell subset differentiation influences both T cell and antibody responses that modulate atherosclerosis, and validate the therapeutic goal of skewing T responses to atherosclerosis-associated antigens. PMID- 15665087 TI - Regular multicharged transient soft matter in Coulomb explosion of heteroclusters. AB - Nanointerfaces of mobile, thin spherical shells of light ions that expand on the femtosecond time scale, can be produced by Coulomb explosion of extremely ionized molecular heteroclusters consisting of light and heavy ions, e.g., (D+Iq+)n (q = 7-35), which are generated in ultraintense laser fields (intensity, I, = 10(16) to 10(20) W.cm(-2)). Modeling, together with molecular dynamics simulations, reveals the expansion of 2D monolayers with high energies and narrow energy distributions [e.g., Eav approximately 23 keV and DeltaE/Eav = 0.16 for D+ from (D+I25+)(2171)] arising from kinematic run-over effects. The expanding regular, monoionic, spherical nanointerfaces manifest the attainment of transient self organization in complex systems driven by repulsive Coulomb interactions. PMID- 15665086 TI - Bacterial glycolipids and analogs as antigens for CD1d-restricted NKT cells. AB - The CD1 family of proteins binds self and foreign glycolipids for presentation to CD1-restricted T cells. To identify previously uncharacterized active CD1 ligands, especially those of microbial origin, numerous glycolipids were synthesized and tested for their ability to stimulate mouse and human natural killer T (NKT) cells. They included analogs of the well known NKT cell agonist alpha-galactosyl ceramide (alpha-GalCer), bacterial glycolipids, and variations of the self-glycolipid, sulfatide. Bacterial glycolipids, alpha-galacturonosyl ceramides from Sphingomonas wittichii, although structurally similar to alpha GalCer, have significant differences in the sugar head group as well as the ceramide portion. The Sphingomonas glycosphingolipids (GSLs) and sulfatide variants were shown to activate human NKT cells as measured by IL-4 and IFN-gamma secretion. Moreover, CD1d-dimer staining revealed human NKT cell reactivity toward these GSLs and to the sulfatides in a fashion comparable with alpha GalCer. Because alpha-GalCer is a marine-sponge-derived ligand, our study here shows that bacterium-derived antigens are also able to stimulate mouse and human NKT cells. PMID- 15665088 TI - Transient interference with staphylococcal quorum sensing blocks abscess formation. AB - The staphylococcal virulon is controlled largely by the agr locus, a global accessory gene regulator that is autoinduced by a self-coded peptide (AIP) and is therefore a quorum sensor. The agr locus has diverged within and between species, giving rise to AIP variants that inhibit heterologous agr activation, an effect with therapeutic potential against Staphylococcus aureus: a single dose of an inhibitory AIP blocks the formation of an experimental murine abscess. As the AIP is unstable at physiological pH, owing to its essential thiolactone bond, its single-dose efficacy seems paradoxical, which has led us to analyze the in vivo kinetics of agr activation and the consequences of its blockage by a heterologous AIP. Initially, the infecting bacteria grow rapidly, achieving sufficient population density within the first 3 h to activate agr, and then enter a neutrophil-induced metabolic eclipse lasting for 2-3 d, followed by agr reactivation concomitantly with the development of the abscess. The inhibitory AIP prevents agr expression only during its short in vivo lifetime, suggesting that the agr-induced and therefore quorum-dependent synthesis of virulence factors shortly after infection is necessary for the subsequent development of the abscess lesion and bacterial survival. We confirm this finding by showing that a sterile agr+ supernatant causes a sterile abscess similar to the septic abscess caused by live bacteria. These results may provide a biological rationale for regulation of virulence factor expression by quorum sensing rather than by response to specific host signals. PMID- 15665089 TI - Microengineered hydromechanical cochlear model. AB - Micromachined fluid-filled variable impedance waveguides intended to mimic the mechanics of the passive mammalian cochlea have been fabricated and experimentally examined. The structures were microfabricated with dimensions similar to those of the biological system. Experimental tests demonstrate acoustically excited traveling fluid-structure waves with phase accumulations between 1.5 and 3 pi radians at the location of maximum response. The resulting measured frequency-position mapping function, with similarities to that observed in the cochlea, is presented. Results for both isotropic and orthotropic membranes are reported, demonstrating that the achieved orthotropy ratio of 8:1 in tension is insufficient to produce the sharp filtering observed in animal experiments and many computational models that use higher ratios. It is also shown experimentally that high viscosity fluids must be used to provide sufficient damping to avoid the formation of a nonphysiological standing wave pattern. A mathematical model incorporating a thin-layer viscous, compressible fluid approximation coupled to an orthotropic membrane model is validated against experimental results. The work presented herein is motivated by the possibility of producing microfabricated cochlear-like filters, thus the structure is designed for production in a scalable microfabrication process. PMID- 15665090 TI - Alpha-glucan, water dikinase (GWD): a plastidic enzyme with redox-regulated and coordinated catalytic activity and binding affinity. AB - The recently discovered potato tuber (Solanum tuberosum) alpha-glucan, water dikinase (GWD) (formerly known as R1) catalyzes the phosphorylation of starch by a dikinase-type reaction mechanism in which the beta-phosphate of ATP is transferred to either the C-6 or the C-3 position of the glucosyl residue of starch. In the present study, we found that the GWD enzyme is inactive in the oxidized form, which is accompanied by the formation of a specific intramolecular disulfide bond as determined by disulfide-linked peptide mapping. The regulatory properties of this disulfide linkage were confirmed by site-directed mutagenesis studies. Both reduced thioredoxin (Trx) f and Trx m from spinach leaves reduced and activated oxidized GWD at very low concentrations, with Trx f being the more efficient, yielding an S0.5 value of 0.4 microM. Interestingly, GWD displays a reversible and selective binding to starch granules depending on the illumination state of the plant. Here we show that starch granule-bound GWD isolated from dark adapted plants exists in the inactive, oxidized form, which is capable of reactivation upon treatment with reduced Trx. Furthermore, the soluble form of GWD was found in its fully reduced state, providing evidence of a Trx-controlled regulation mechanism linking enzymatic activity and specific binding affinities of a protein to an intracellular surface. The regulatory site sequence, CFATC, of potato GWD is conserved in chloroplast-targeted GWDs from other species, suggesting an overall redox regulation of the GWD enzyme. PMID- 15665091 TI - Iron release from macrophages after erythrophagocytosis is up-regulated by ferroportin 1 overexpression and down-regulated by hepcidin. AB - Ferroportin 1 (FPN1) is transmembrane protein involved in iron homeostasis. In the duodenum, FPN1 localizes to the basolateral surface of enterocytes where it appears to export iron out of the cell and into the portal circulation. FPN1 is also abundantly expressed in reticuloendothelial macrophages of the liver, spleen, and bone marrow, suggesting that this protein serves as an iron exporter in cells that recycle iron from senescent red blood cells. To directly test the hypothesis that FPN1 functions in the export of iron after erythrophagocytosis, FPN1 was stably expressed in J774 mouse macrophages by using retroviral transduction, and release of 59Fe after phagocytosis of 59Fe-labeled rat erythrocytes was measured. J774 cells overexpressing FPN1 released 70% more 59Fe after erythrophagocytosis than control cells, consistent with a role in the recycling of iron from senescent red cells. Treatment of cells with the peptide hormone hepcidin, a systemic regulator of iron metabolism, dramatically decreased FPN1 protein levels and significantly reduced the efflux of 59Fe after erythrophagocytosis. Subsequent fractionation of the total released 59Fe into heme and nonheme compounds revealed that hepcidin treatment reduced the release of nonheme 59Fe by 50% and 25% from control and FPN1-overexpressing cells, respectively, but did not diminish efflux of 59Fe-heme. We conclude that FPN1 is directly involved in the export of iron during erythrocyte-iron recycling by macrophages. PMID- 15665092 TI - Crystal structure of a pH-regulated luciferase catalyzing the bioluminescent oxidation of an open tetrapyrrole. AB - The luciferase of Lingulodinium polyedrum, a marine bioluminescent dinoflagellate, consists of three similar but not identical domains in a single polypeptide. Each encodes an active luciferase that catalyzes the oxidation of a chlorophyll-derived open tetrapyrrole (dinoflagellate luciferin) to produce blue light. These domains share no sequence similarity with any other in the GenBank database and no structural or motif similarity with any other luciferase. We report here the 1.8-A crystal structure of the third domain, D3, at pH 8, and a mechanism for its activity regulation by pH. D3 consists of two major structural elements: a beta-barrel pocket putatively for substrate binding and catalysis and a regulatory three-helix bundle. N-terminal histidine residues previously shown to regulate activity by pH are at the interface of the helices in the bundle. Molecular dynamics calculations indicate that, in response to changes in pH, these histidines could trigger a large molecular motion of the bundle, thereby exposing the active site to the substrate. PMID- 15665093 TI - Kisspeptin directly stimulates gonadotropin-releasing hormone release via G protein-coupled receptor 54. AB - We have recently described a molecular gatekeeper of the hypothalamic-pituitary gonadal axis with the observation that G protein-coupled receptor 54 (GPR54) is required in mice and men for the pubertal onset of pulsatile luteinizing hormone (LH) and follicle-stimulating hormone (FSH) secretion to occur. In the present study, we investigate the possible central mode of action of GPR54 and kisspeptin ligand. First, we show that GPR54 transcripts are colocalized with gonadotropin releasing hormone (GnRH) neurons in the mouse hypothalamus, suggesting that kisspeptin, the GPR54 ligand, may act directly on these neurons. Next, we show that GnRH neurons seem anatomically normal in gpr54-/- mice, and that they show projections to the median eminence, which demonstrates that the hypogonadism in gpr54-/- mice is not due to an abnormal migration of GnRH neurons (as occurs with KAL1 mutations), but that it is more likely due to a lack of GnRH release or absence of GnRH neuron stimulation. We also show that levels of kisspeptin injected i.p., which stimulate robust LH and FSH release in wild-type mice, have no effect in gpr54-/- mice, and therefore that kisspeptin acts directly and uniquely by means of GPR54 signaling for this function. Finally, we demonstrate by direct measurement, that the central administration of kisspeptin intracerebroventricularly in sheep produces a dramatic release of GnRH into the cerebrospinal fluid, with a parallel rise in serum LH, demonstrating that a key action of kisspeptin on the hypothalamo-pituitary-gonadal axis occurs directly at the level of GnRH release. The localization and GnRH release effects of kisspeptin thus define GPR54 as a major control point in the reproductive axis and suggest kisspeptin to be a neurohormonal effector. PMID- 15665094 TI - Metabolic engineering of dhurrin in transgenic Arabidopsis plants with marginal inadvertent effects on the metabolome and transcriptome. AB - Focused and nontargeted approaches were used to assess the impact associated with introduction of new high-flux pathways in Arabidopsis thaliana by genetic engineering. Transgenic A. thaliana plants expressing the entire biosynthetic pathway for the tyrosine-derived cyanogenic glucoside dhurrin as accomplished by insertion of CYP79A1, CYP71E1, and UGT85B1 from Sorghum bicolor were shown to accumulate 4% dry-weight dhurrin with marginal inadvertent effects on plant morphology, free amino acid pools, transcriptome, and metabolome. In a similar manner, plants expressing only CYP79A1 accumulated 3% dry weight of the tyrosine derived glucosinolate, p-hydroxybenzylglucosinolate with no morphological pleitropic effects. In contrast, insertion of CYP79A1 plus CYP71E1 resulted in stunted plants, transcriptome alterations, accumulation of numerous glucosides derived from detoxification of intermediates in the dhurrin pathway, and in loss of the brassicaceae-specific UV protectants sinapoyl glucose and sinapoyl malate and kaempferol glucosides. The accumulation of glucosides in the plants expressing CYP79A1 and CYP71E1 was not accompanied by induction of glycosyltransferases, demonstrating that plants are constantly prepared to detoxify xenobiotics. The pleiotrophic effects observed in plants expressing sorghum CYP79A1 and CYP71E1 were complemented by retransformation with S. bicolor UGT85B. These results demonstrate that insertion of high-flux pathways directing synthesis and intracellular storage of high amounts of a cyanogenic glucoside or a glucosinolate is achievable in transgenic A. thaliana plants with marginal inadvertent effects on the transcriptome and metabolome. PMID- 15665095 TI - Estrogen receptor alpha and imprinting of the neonatal mouse ventral prostate by estrogen. AB - Exposure to estrogen in the neonatal period affects prostatic growth and leads to an increased incidence of prostatic intraepithelial neoplasia in later life. The effects of neonatal estrogen are clearly dependent on estrogen receptor (ER) alpha because they do not occur in ERalpha-knockout mice. Because ERalpha is expressed in the stroma, but not in the epithelium, of the adult ventral prostate, the concept of indirect estrogen effects through stromal signaling has been proposed. Here, we show that during the first 4 weeks of life, there are profound and rapid changes in the ER profile in the mouse ventral prostate. ERalpha is abundant in the stroma during week 1, but by week 2 it is exclusively epithelial, and then by week 4, ERalpha is lost and ERbeta is dominant in the prostatic epithelium. The presence of ERalpha is associated with a high proliferation index, and ERbeta is associated with quiescence. Branching morphogenesis was altered in ERalpha-/-, but not in ERbeta-/-, mice. We conclude that imprinting and branching morphogenesis of the ventral prostate are mediated by estrogen acting directly on epithelial and stromal ERalpha during the first 2 weeks of life. PMID- 15665096 TI - Impaired intranuclear trafficking of Runx2 (AML3/CBFA1) transcription factors in breast cancer cells inhibits osteolysis in vivo. AB - Runx transcription factors comprise a family of proteins that are essential for organogenesis. A unique nuclear matrix-targeting signal in the C terminus directs these factors to their appropriate subnuclear domains. At these sites, they interact with coregulatory proteins and target genes. We have previously shown that aberrant expression of the Runx2 DNA binding domain in metastatic breast cancer cells can prevent production of osteolytic lesions in bone. Here, we show that proper Runx2 subnuclear targeting is required for osteolysis. We have identified point mutations of the Runx2 nuclear matrix-targeting signal sequence that impair its targeting to nuclear matrix sites. These mutations block the invasive and osteolytic properties of MDA-MB-231 breast cancer cells in vivo. Cell lines expressing this Runx2 mutant protein inhibit the osteogenic properties of bone marrow stromal cells in coculture assays. The mutant breast cancer cells also exhibit reduced invasiveness in vitro and do not express genes involved in invasion and angiogenesis (VEGF and MMP13). Our findings suggest that fidelity of Runx2 intranuclear organization is necessary for expression of target genes that mediate the osteolytic activity of metastatic breast cancer cells. PMID- 15665099 TI - Experiments investigating cooperative types in humans: a complement to evolutionary theory and simulations. AB - Unlike other species, humans cooperate in large, distantly related groups, a fact that has long presented a puzzle to biologists. The pathway by which adaptations for large-scale cooperation among nonkin evolved in humans remains a subject of vigorous debate. Results from theoretical analyses and agent-based simulations suggest that evolutionary dynamics need not yield homogeneous populations, but can instead generate a polymorphic population that consists of individuals who vary in their degree of cooperativeness. These results resonate with the recent increasing emphasis on the importance of individual differences in understanding and modeling behavior and dynamics in experimental games and decision problems. Here, we report the results of laboratory experiments that complement both theory and simulation results. We find that our subjects fall into three types, an individual's type is stable, and a group's cooperative outcomes can be remarkably well predicted if one knows its type composition. Reciprocal types, who contribute to the public good as a positive function of their beliefs about others' contributions, constitute the majority (63%) of players; cooperators and free-riders are also present in our subject population. Despite substantial behavioral differences, earnings among types are statistically identical. Our results support the view that our human subject population is in a stable, polymorphic equilibrium of types. PMID- 15665097 TI - Unparalleled complexity of the MHC class I region in rhesus macaques. AB - The highly polymorphic gene products of the classical MHC class I genes in humans (HLA-A, HLA-B, and HLA-C) play a critical role in the immune defense against intracellular infections. Because non-human primates are important models for AIDS vaccine research, rhesus monkeys from a thoroughly pedigreed and serotyped colony were subjected to full-length cDNA analysis of MHC class I gene transcripts. Rhesus macaques express multiple dominant Mamu-A and Mamu-B transcripts (majors) per chromosome, which are characterized by high expression levels. The presence of additional cDNAs with low levels of expression (minors) suggests evidence for transcriptional control of MHC class I genes. Moreover, phylogenetic analyses illustrate that most of the Mamu-A and Mamu-B loci/lineages identified display no or only limited levels of allelic polymorphism. Thus, MHC class I diversity in rhesus macaques is typified by the existence of an unmatched high number of Mamu-A and Mamu-B region configurations that exhibit polymorphism with regard to the number and combination of transcribed loci present per chromosome. PMID- 15665098 TI - Differential contribution of the three Aph1 genes to gamma-secretase activity in vivo. AB - Gamma-secretase is the protease responsible for amyloid beta peptide release and is needed for Notch, N-Cadherin, and possibly other signaling pathways. The protease complex consists of at least four subunits, i.e., Presenilin, Aph1, Pen2, and Nicastrin. Two different genes encode Aph1A and Aph1B in man. A duplication of Aph1B in rodents has given rise to a third gene, Aph1C. Different mixes of gamma-secretase subunits assemble in at least four human and six rodent complexes but it is not known whether they have different activities in vivo. We report here the inactivation of the three Aph1 genes in mice. Aph1A-/- embryos show a lethal phenotype characterized by angiogenesis defects in the yolk sac, neuronal tube malformations, and mild somitogenesis defects. Aph1B-/- or C-/- or the combined Aph1BC-/- mice (which can be considered as a model for total Aph1B loss in human) survive into adulthood. However, Aph1BC-/- deficiency causes a mild but significant reduction in amyloid beta percursor protein processing in selective regions of the adult brain. We conclude that the biochemical and physiological repercussions of genetically reducing gamma-secretase activity via the different Aph1 components are quite divergent and tissue specific. Our work provides in vivo evidence for the concept that different gamma-secretase complexes may exert different biological functions. In the context of Alzheimer's disease therapy, this implies the theoretical possibility that targeting specific gamma-secretase subunit combinations could yield less toxic drugs than the currently available general inhibitors of gamma-secretase activity. PMID- 15665100 TI - Accessing key steps of human tumor progression in vivo by using an avian embryo model. AB - Experimental in vivo tumor models are essential for comprehending the dynamic process of human cancer progression, identifying therapeutic targets, and evaluating antitumor drugs. However, current rodent models are limited by high costs, long experimental duration, variability, restricted accessibility to the tumor, and major ethical concerns. To avoid these shortcomings, we investigated whether tumor growth on the chick chorio-allantoic membrane after human glioblastoma cell grafting would replicate characteristics of the human disease. Avascular tumors consistently formed within 2 days, then progressed through vascular endothelial growth factor receptor 2-dependent angiogenesis, associated with hemorrhage, necrosis, and peritumoral edema. Blocking of vascular endothelial growth factor receptor 2 and platelet-derived growth factor receptor signaling pathways by using small-molecule receptor tyrosine kinase inhibitors abrogated tumor development. Gene regulation during the angiogenic switch was analyzed by oligonucleotide microarrays. Defined sample selection for gene profiling permitted identification of regulated genes whose functions are associated mainly with tumor vascularization and growth. Furthermore, expression of known tumor progression genes identified in the screen (IL-6 and cysteine-rich angiogenic inducer 61) as well as potential regulators (lumican and F-box-only 6) follow similar patterns in patient glioma. The model reliably simulates key features of human glioma growth in a few days and thus could considerably increase the speed and efficacy of research on human tumor progression and preclinical drug screening. PMID- 15665101 TI - Fast decay of iconic memory in observers with mild cognitive impairments. AB - In a previous clinical report, unusually fast decay of iconic memory was obtained from a subject who later developed Alzheimer's disease. By using the partial report paradigm, iconic memory (a form of visual sensory memory) in a group of observers with mild cognitive impairments (MCI) was characterized and compared with that of young college-age adults and older controls. Relatively long stimulus exposures were used for all three groups to ensure that older observers could perceive the stimuli. A set of conventional neuropsychological tests assessed cognitive functions of the MCI and older control groups. We found that iconic memory decayed much faster for observers with MCI than for normal controls, old or young, although the two groups of older observers performed at equivalent levels in precue tests (assay of visibility) and tests cued at long delays (assay of short-term memory). The result suggests that fast decay of iconic memory might be a general characteristic of observers with MCI who are at much higher than average risk of developing Alzheimer's disease later in life. PMID- 15665102 TI - Gating of acetylcholine receptor channels: brownian motion across a broad transition state. AB - Acetylcholine receptor channels (AChRs) are proteins that switch between stable "closed" and "open" conformations. In patch clamp recordings, diliganded AChR gating appears to be a simple, two-state reaction. However, mutagenesis studies indicate that during gating dozens of residues across the protein move asynchronously and are organized into rigid body gating domains ("blocks"). Moreover, there is an upper limit to the apparent channel opening rate constant. These observations suggest that the gating reaction has a broad, corrugated transition state region, with the maximum opening rate reflecting, in part, the mean first-passage time across this ensemble. Simulations reveal that a flat, isotropic energy profile for the transition state can account for many of the essential features of AChR gating. With this mechanism, concerted, local structural transitions that occur on the broad transition state ensemble give rise to fractional measures of reaction progress (Phi values) determined by rate equilibrium free energy relationship analysis. The results suggest that the coarse-grained AChR gating conformational change propagates through the protein with dynamics that are governed by the Brownian motion of individual gating blocks. PMID- 15665103 TI - An intermolecular base triple as the basis of ligand specificity and affinity in the guanine- and adenine-sensing riboswitch RNAs. AB - Riboswitches are highly structured RNA elements that control the expression of many bacterial genes by binding directly to small metabolite molecules with high specificity and affinity. In Bacillus subtilis, two classes of riboswitches have been described that discriminate between guanine and adenine despite an extremely high degree of homology both in their primary and secondary structure. We have identified intermolecular base triples between both purine ligands and their respective riboswitch RNAs by NMR spectroscopy. Here, specificity is mediated by the formation of a Watson-Crick base pair between the guanine ligand and a C residue or the adenine ligand and a U residue of the cognate riboswitch RNA, respectively. In addition, a second base-pairing interaction common to both riboswitch purine complexes involves a uridine residue of the RNA and the N3/N9 edge of the purine ligands. This base pairing is mediated by a previously undescribed hydrogen-bonding scheme that contributes to the affinity of the RNA ligand interaction. The observed intermolecular hydrogen bonds between the purine ligands and the RNA rationalize the previously observed change in specificity upon a C to U mutation in the core of the purine riboswitch RNAs and the differences in the binding affinities for a number of purine analogs. PMID- 15665105 TI - Reconstructing early sponge relationships by using the Burgess Shale fossil Eiffelia globosa, Walcott. AB - The relationships of the sponge classes are controversial, particularly between the calcareous and siliceous sponges. Specimens of the putative calcarean Eiffelia globosa Walcott from the Burgess Shale show the presence of diagnostic hexactinellid spicules integrated into the skeletal mesh. The arrangement of these spicules in Eiffelia is shown to be precisely equivalent to that of early protospongioid hexactinellids, and sponge growth occurred through an identical pattern to produce identical skeletal body morphology. The difference in spicule composition of the classes is interpreted through the observation of taphonomic features of Eiffelia that suggest the presence of at least two mineralogically distinct layers within the spicules. These results support molecular analyses that identify the calcarean-silicisponge transition as the earliest major sponge branch and suggest that the heteractinids were paraphyletic with respect to the Hexactinellida. PMID- 15665104 TI - The Wnt/beta-catenin signaling pathway targets PPARgamma activity in colon cancer cells. AB - Control of colon cell fate in adenocarcinomas is disrupted, in part, due to aberrant Wnt/beta-catenin signaling. The nuclear receptor peroxisome proliferator activated receptor-gamma (PPARgamma) has been implicated in the development of colon cancers. In the adenomatous polyposis coli multiple intestinal neoplasia (APCMin) mouse cancer model, PPARgamma expression in the colonic mucosa is markedly altered. In addition, PPARgamma protein levels are elevated, possibly through sequestration by activated beta-catenin in colon cancer cell lines. Induction of the Wnt/beta-catenin pathway by LiCl also elevated PPARgamma levels and induced PPARgamma-dependent reporter and endogenous target genes. Mechanistically, PPARgamma, through interactions with beta-catenin and T cell transcription factor (Tcf)-4, may be a determinant of cell fate and is likely a target of the Wnt pathway in cancer cells. PMID- 15665106 TI - Attenuation of cardiac remodeling after myocardial infarction by muscle LIM protein-calcineurin signaling at the sarcomeric Z-disc. AB - Adverse left ventricular (LV) remodeling after myocardial infarction (MI) is a major cause for heart failure. Molecular modifiers of the remodeling process remain poorly defined. Patients with heart failure after MI have reduced LV expression levels of muscle LIM protein (MLP), a component of the sarcomeric Z disk that is involved in the integration of stress signals in cardiomyocytes. By using heterozygous MLP mutant (MLP+/-) mice, we explored the role of MLP in post MI remodeling. LV dimensions and function were similar in sham-operated WT and MLP+/- mice. After MI, however, MLP+/- mice displayed more pronounced LV dilatation and systolic dysfunction and decreased survival compared with WT mice, indicating that reduced MLP levels predispose to adverse LV remodeling. LV dilatation in MLP+/- mice was associated with reduced thickening but enhanced elongation of cardiomyocytes. Activation of the stress-responsive, prohypertrophic calcineurin-nuclear factor of activated T-cells (NFAT) signaling pathway was reduced in MLP+/- mice after MI, as shown by a blunted transcriptional activation of NFAT in cardiomyocytes isolated from MLP+/-/NFAT luciferase reporter gene transgenic mice. Calcineurin was colocalized with MLP at the Z-disk in WT mice but was displaced from the Z-disk in MLP+/- mice, indicating that MLP is essential for calcineurin anchorage to the Z-disk. In vitro assays in cardiomyocytes with down-regulated MLP confirmed that MLP is required for stress-induced calcineurin-NFAT activation. Our study reveals a link between the stress sensor MLP and the calcineurin-NFAT pathway at the sarcomeric Z-disk in cardiomyocytes and indicates that reduced MLP-calcineurin signaling predisposes to adverse remodeling after MI. PMID- 15665107 TI - Ancestry and progeny of nutrient amino acid transporters. AB - The biosynthesis of structural and signaling molecules depends on intracellular concentrations of essential amino acids, which are maintained by a specific system of plasma membrane transporters. We identify a unique population of nutrient amino acid transporters (NATs) within the sodium-neurotransmitter symporter family and have characterized a member of the NAT subfamily from the larval midgut of the Yellow Fever vector mosquito, Aedes aegypti (aeAAT1, AAR08269), which primarily supplies phenylalanine, an essential substrate for the synthesis of neuronal and cuticular catecholamines. Further analysis suggests that NATs constitute a comprehensive transport metabolon for the epithelial uptake and redistribution of essential amino acids including precursors of several neurotransmitters. In contrast to the highly conserved subfamily of orthologous neurotransmitter transporters, lineage-specific, paralogous NATs undergo rapid gene multiplication/substitution that enables a high degree of evolutionary plasticity of nutrient amino acid uptake mechanisms and facilitates environmental and nutrient adaptations of organisms. These findings provide a unique model for understanding the molecular mechanisms, physiology, and evolution of amino acid and neurotransmitter transport systems and imply that monoamine and GABA transporters evolved by selection and conservation of earlier neuronal NATs. PMID- 15665109 TI - Anomalous megakaryocytopoiesis in mice with mutations in the c-Myb gene. AB - Mpl(-/-) mice bearing the Plt3 or Plt4 mutations in the c-Myb gene exhibit thrombopoietin (TPO)-independent supraphysiological platelet production accompanied by excessive megakaryocytopoiesis and defective erythroid and lymphoid cell production. To better define the cellular basis for the thrombocytosis in these mice, we analyzed the production and characteristics of megakaryocytes and their progenitors. Consistent with thrombocytosis arising from hyperactive production, the high platelet counts in mice carrying the c-Myb(Plt4) allele were not accompanied by any significant alteration in platelet half-life. Megakaryocytes in c-Myb mutant mice displayed reduced modal DNA ploidy and, among the excessive numbers of megakaryocyte progenitor cells, more mature precursors were particularly evident. Megakaryocyte progenitor cells carrying the Plt3 or Plt4 c-Myb mutations, but not granulocyte-macrophage progenitors, exhibited 200 fold enhanced responsiveness to granulocyte-macrophage colony-stimulating factor (GM-CSF), suggesting that altered responses to cytokines may contribute to expanded megakaryocytopoiesis. Mutant preprogenitor (blast colony-forming) cells appeared to have little capacity to form megakaryocyte progenitor cells. In contrast, the spleens of irradiated mice 12 days after transplantation with mutant bone marrow contained abundant megakaryocyte progenitor cells, suggesting that altered c-Myb activity skews differentiation commitment in spleen colony forming units (CFU-S) in favor of excess megakaryocytopoiesis. PMID- 15665108 TI - The DNA/RNA-binding protein MSY2 marks specific transcripts for cytoplasmic storage in mouse male germ cells. AB - During spermatogenesis, male germ cells temporally synthesize many proteins as they differentiate through meiosis and become spermatozoa. The germ cell Y-box protein, MSY2, constituting approximately 0.7% of total protein in male germ cells, binds to a consensus promoter element, and shows a general lack of RNA binding specificity. Combining immunoprecipitation and suppressive subtractive hybridization, we identified populations of germ cell mRNAs that are not bound or bound by MSY2. The former population is enriched in cell growth and ubiquitously expressed mRNAs, whereas the latter population is enriched for stored or translationally delayed, male gamete-specific transcripts. Chromatin precipitation assays reveal that most of the MSY2 target mRNAs are transcribed from genes containing the Y-box DNA-binding motif in their promoters. In transgenic mice, mRNAs encoding exogenous GFP are directed or not directed into the MSY2-bound fraction by promoters containing or lacking the Y-box motif, respectively. We propose that MSY2 marks specific mRNAs in the nucleus for cytoplasmic storage, thereby linking transcription and mRNA storage/translational delay in meiotic and postmeiotic male germ cells of the mouse. PMID- 15665110 TI - Allogeneic stem-cell transplantation with reduced conditioning intensity as a novel immunotherapy and antiviral therapy for adult T-cell leukemia/lymphoma. AB - Sixteen patients with adult T-cell leukemia/lymphoma (ATL) who were all over 50 years of age underwent allogeneic stem cell transplantation with reduced conditioning intensity (RIST) from HLA-matched sibling donors after a conditioning regimen consisting of fludarabine (180 mg/m2), busulfan (8 mg/kg), and rabbit antithymocyte globulin (5 mg/kg). The observed regimen-related toxicities and nonhematologic toxicities were all found to be acceptable. Disease relapse was the main cause of treatment failure. Three patients who had a relapse subsequently responded to a rapid discontinuation of the immunosuppressive agent and thereafter achieved another remission. After RIST, the human T-cell leukemia virus type 1 (HTLV-1) proviral load became undetectable in 8 patients. RIST is thus considered to be a feasible treatment for ATL. Our data also suggest the presence of a possible graft-versus-ATL effect; an anti-HTLV-1 activity was also found to be associated with this procedure. PMID- 15665111 TI - Platelet homeostasis is regulated by platelet expression of CD47 under normal conditions and in passive immune thrombocytopenia. AB - Interaction between target cell CD47 and the inhibitory macrophage receptor signal regulatory protein alpha (SIRPalpha) counteracts macrophage phagocytosis of CD47-expressing host cells. As platelets also express CD47, we asked whether inhibitory CD47/SIRPalpha signaling regulates normal platelet turnover and clearance of platelets in immune thrombocytopenic purpura (ITP). CD47(-/-) mice had a mild spontaneous thrombocytopenia, which was not due to a decreased platelet half-life as a result of increased expression of P-selectin, CD61, or phosphatidylserine. In contrast, CD47(-/-) platelets were rapidly cleared when transfused into CD47(+/+) recipients, whereas CD47(+/-) platelets had a nearly normal half-life in CD47(+/+) mice under nonautoimmune conditions. CD47(-/-) mice were more sensitive to ITP, as compared with CD47(+/+) mice. In vitro, macrophage phagocytosis of immunoglobulin G (IgG)-opsonized CD47(-/-) platelets was significantly higher than that for equally opsonized CD47(+/+) platelets. However, when SIRPalpha was blocked, phagocytosis of CD47(+/+) platelets increased to the level of CD47(-/-) platelets. Phagocytosis of opsonized CD47(+/ ) platelets was higher than that for CD47(+/+) platelets, but lower than that for CD47(-/-) platelets, suggesting a gene-dose effect of CD47 in this system. In conclusion, we suggest that inhibitory CD47/SIRPalpha signaling is involved in regulating platelet phagocytosis in ITP, and that targeting SIRPalpha may be a new means of reducing platelet clearance in ITP. PMID- 15665112 TI - TEF, an antiapoptotic bZIP transcription factor related to the oncogenic E2A-HLF chimera, inhibits cell growth by down-regulating expression of the common beta chain of cytokine receptors. AB - Gain and/or loss of function mediated by chimeric transcription factors generated by nonrandom translocations in leukemia is a key to understanding oncogenesis. E2A-hepatic leukemia factor (HLF), a chimeric basic region/leucine zipper (bZIP) transcription factor expressed in t(17;19)-positive leukemia cells, contributes to leukemogenesis through its potential to inhibit apoptosis. To identify physiologic counterparts of this chimera, we investigated the function of other bZIP factors that bind to the same DNA sequence recognized by E2A-HLF. Here, we show that thyrotroph embryonic factor (TEF), which shares a high level of sequence identity with HLF and recognizes the same DNA sequence, is expressed in a small fraction of each subset of hematolymphoid progenitors. When TEF was introduced into FL5.12 interleukin 3 (IL-3)-dependent cells, TEF protected the cells from apoptosis due to IL-3 deprivation. Unexpectedly, TEF also almost completely down-regulated expression of the common beta (betac) chain of cytokine receptors. Consequently, TEF-expressing cells accumulated in G(0)/G(1) phase without undergoing apoptosis. These findings suggest that TEF is one of the apoptotic regulators in hematopoietic progenitors and controls hematopoietic-cell proliferation by regulating the expression of the betac chain. In contrast, E2A HLF promoted cell survival more efficiently than TEF but did not down-regulate betac chain expression, suggesting that E2A-HLF retains ideal properties for driving leukemic transformation. PMID- 15665114 TI - P2Y1 and P2Y12 receptors for ADP desensitize by distinct kinase-dependent mechanisms. AB - Adenosine 5'-diphosphate (ADP) plays a central role in regulating platelet function by the activation of the G protein-coupled receptors P2Y(1) and P2Y(12). Although it is well established that aggregation responses of platelets to ADP desensitize, the underlying mechanisms involved remain unclear. In this study we demonstrate that P2Y(1)- and P2Y(12)-mediated platelet responses desensitize rapidly. Furthermore, we have established that these receptors desensitize by different kinase-dependent mechanisms. G protein-coupled receptor kinase (GRK) 2 and GRK6 are both endogenously expressed in platelets. Transient overexpression of dominant-negative mutants of these kinases or reductions in endogenous GRK expression by the use of specific siRNAs in 1321N1 cells showed that P2Y(12), but not P2Y(1), desensitization is mediated by GRKs. In contrast, desensitization of P2Y(1), but not P2Y(12), is largely dependent on protein kinase C activity. This study is the first to show that both P2Y(1) and P2Y(12) desensitize in human platelets, and it reveals ways in which their sensitivity to ADP may be differentially and independently altered. PMID- 15665113 TI - Synergistic interactions between imatinib mesylate and the novel phosphoinositide dependent kinase-1 inhibitor OSU-03012 in overcoming imatinib mesylate resistance. AB - Resistance to the Ableson protein tyrosine (Abl) kinase inhibitor imatinib mesylate has become a critical issue for patients in advanced phases of chronic myelogenous leukemia. Imatinib-resistant tumor cells develop, in part, as a result of point mutations within the Abl kinase domain. As protein kinase B (Akt) plays a pivotal role in Abl oncogene-mediated cell survival, we hypothesize that concurrent inhibition of Akt will sensitize resistant cells to the residual apoptotic activity of imatinib mesylate, thereby overcoming the resistance. Here, we examined the effect of OSU-03012, a celecoxib-derived phosphoinositide dependent kinase-1 (PDK-1) inhibitor, on imatinib mesylate-induced apoptosis in 2 clinically relevant breakpoint cluster region (Bcr)-Abl mutant cell lines, Ba/F3p210(E255K) and Ba/F3p210(T315I). The 50% inhibitory concentration (IC50) values of imatinib mesylate to inhibit the proliferation of Ba/F3p210(E255K) and Ba/F3p210(T315I) were 14 +/- 4 and 30 +/- 2 microM, respectively. There was no cross-resistance to OSU-03012 in these mutant cells with an IC50 of 5 microM irrespective of mutations. Nevertheless, in the presence of OSU-03012 the susceptibility of these mutant cells to imatinib-induced apoptosis was significantly enhanced. This synergistic action was, at least in part, mediated through the concerted effect on phospho-Akt. Together these data provide a novel therapeutic strategy to overcome imatinib mesylate resistance, especially with the Abl mutant T315I. PMID- 15665115 TI - Hereditary hemochromatosis is reflected in the iron isotope composition of blood. AB - It has recently been shown that the iron isotopic composition of blood differs between individuals and sexes, which is supposed to reflect individual differences in iron metabolism. We hypothesized that patients suffering from hereditary hemochromatosis would demonstrate alterations in the iron isotopic composition of blood due to persistent up-regulation of intestinal iron absorption. Blood from 30 patients with homozygous C282Y hemochromatosis was analyzed for iron isotopic composition by a newly developed technique using multicollector inductively coupled plasma mass spectrometry (MC-ICP-MS). Blood of patients with hemochromatosis is characterized by a higher 56Fe/54Fe isotope ratio than blood of healthy individuals, which are either members of an age matched control group (n = 10; P < .001) or young adults (n = 36; P < .001). In patients with hereditary hemochromatosis, the 56Fe/54Fe isotope ratio of blood significantly correlates with total-body iron accumulation, severity of clinical disease, and the need for regular phlebotomies to prevent iron reaccumulation. We conclude that blood of patients with hereditary hemochromatosis contains more of the heavier iron isotopes than blood of healthy individuals. The primary determinant of the iron isotopic composition of blood appears to be isotope sensitive iron absorption in the intestine and the efficiency of this process. PMID- 15665116 TI - Pharmacologic inhibitors of PI3K/Akt potentiate the apoptotic action of the antileukemic drug arsenic trioxide via glutathione depletion and increased peroxide accumulation in myeloid leukemia cells. AB - Treatment for 14 to 24 hours with low concentrations of arsenic trioxide (As2O3, 1-4 microM) caused apoptosis in U-937 promonocytes and other human myeloid leukemia cell lines (HL-60, NB4). This effect was potentiated by cotreatment with the phosphatidylinositol 3-kinase (PI3K) inhibitors LY294002 and wortmannin, and the Akt inhibitor Akt(i)5. However, the inhibitors did not increase the toxicity of the mitochondria-targeting drug lonidamine, and the DNA-specific drugs camptothecin and cisplatin, when used under similar experimental conditions as As2O3. The potentiation of As2O3-provoked apoptosis involved the increased disruption of mitochondrial transmembrane potential, increased caspase-3 activation and cytochrome c release from mitochondria, increased Bax and Bid activation, and attenuation of 27-kDa heat shock protein (HSP27) expression; the potentiation was prevented by Bcl-2 overexpression. The PI3K/Akt inhibitors decreased the intracellular glutathione content, and caused intracellular oxidation, as measured by peroxide accumulation. Cotreatment with subcytotoxic concentrations of hydrogen peroxide increased apoptosis induction by As2O3. On the other hand, the treatments did not significantly affect glutathione S transferase pi expression and activity. These results, which indicate that glutathione is a target of PI3K/Akt in myeloid leukemia cells, may partially explain the selective increase of As2O3 toxicity by PI3K/Akt inhibitors, and may provide a rationale to improve the efficacy of these inhibitors as therapeutic agents. PMID- 15665118 TI - Membrane type 1-matrix metalloproteinase is involved in migration of human monocytes and is regulated through their interaction with fibronectin or endothelium. AB - Membrane type 1-matrix metalloproteinase (MT1-MMP) is involved in endothelial and tumor-cell migration, but its putative role in leukocyte migration has not been characterized yet. Here, we demonstrate that anti-MT1-MMP monoclonal antibody (mAb) impaired monocyte chemotactic protein-1 (MCP-1)-stimulated monocyte migration on fibronectin (FN), vascular cell adhesion molecule-1 (VCAM-1), and intercellular adhesion molecule-1 (ICAM-1). In addition, monocyte transmigration through tumor necrosis factor-alpha (TNF-alpha)-activated endothelium is also inhibited by anti-MT1-MMP mAb. Therefore, regulation of MT1-MMP in human peripheral blood monocytes was investigated. First, MT1-MMP clustering was observed at motility-associated membrane protrusions of MCP-1-stimulated monocytes migrating on FN, VCAM-1, or ICAM-1 and at the leading edge, together with profilin, of monocytes transmigrating through activated endothelial cells. In addition, up-regulation of MT1-MMP expression was induced in human monocytes upon attachment to FN in a manner dependent on alpha4beta1 and alpha5beta1 integrins. Binding of monocytes to TNF-alpha-activated human endothelial cells as well as to VCAM-1 or ICAM-1 also resulted in an increase of MT1-MMP expression. These findings correlated with an enhancement of MT1-MMP fibrinolytic activity in monocytes bound to FN, VCAM-1, or ICAM-1. Our data show that MT1-MMP is required during human monocyte migration and endothelial transmigration and that MT1-MMP localization, expression, and activity are regulated in monocytes upon contact with FN or endothelial ligands, pointing to a key role of MT1-MMP in monocyte recruitment during inflammation. PMID- 15665117 TI - Activation of Src kinase Lyn by the Kaposi sarcoma-associated herpesvirus K1 protein: implications for lymphomagenesis. AB - The K1 gene of Kaposi sarcoma-associated herpesvirus (KSHV) encodes a transmembrane glycoprotein bearing a functional immunoreceptor tyrosine-based activation motif (ITAM). Previously, we reported that the K1 protein induced plasmablastic lymphomas in K1 transgenic mice, and that these lymphomas showed enhanced Lyn kinase activity. Here, we report that systemic administration of the nuclear factor kappa B (NF-kappaB) inhibitor Bay 11-7085 or an anti-vascular endothelial growth factor (VEGF) antibody significantly reduced K1 lymphoma growth in nude mice. Furthermore, in KVL-1 cells, a cell line derived from a K1 lymphoma, inhibition of Lyn kinase activity by the Src kinase inhibitor PP2 decreased VEGF induction, NF-kappaB activity, and the cell proliferation index by 50% to 75%. In contrast, human B-cell lymphoma BJAB cells expressing K1, but not the ITAM sequence-deleted mutant K1, showed a marked increase in Lyn kinase activity with concomitant VEGF induction and NF-kappaB activation, indicating that ITAM sequences were required for the Lyn kinase-mediated activation of these factors. Our results suggested that K1-mediated constitutive Lyn kinase activation in K1 lymphoma cells is crucial for the production of VEGF and NF kappaB activation, both strongly implicated in the development of KSHV-induced lymphoproliferative disorders. PMID- 15665120 TI - Posttranslational modifications of p27kip1 determine its binding specificity to different cyclins and cyclin-dependent kinases in vivo. AB - Using 2-dimensional gel electrophoresis (2D-gel) analysis, we show here that cell cycle entry is associated with a significant increase in p27(kip1) phosphorylation in human primary B cells. A similar pattern of increase in p27(kip1) phosphorylation was also seen in 2 fast-growing tumor cell lines, Burkitt lymphoma cell line BL40 and breast carcinoma cell line Cal51, where inactive p27(kip1) is expressed at high levels. Detailed analysis revealed for the first time that different cyclins and cyclin-dependent kinases (cdk's) interact with distinct posttranslationally modified isoforms of p27(kip1) in vivo. Cyclin E but not cyclin A selectively interacts with phosphorylated p27(kip1) isoforms, while cyclin D1 and D2 favor unphosphorylated p27(kip1) isoforms in vivo. Interestingly, cyclin D3 and cdk4 selectively interact with phosphorylated p27(kip1) in BL40 cells. Among all D-type cyclin/cdk4 and cdk6 complexes, cyclin D3/cdk4 is most active in sequestering the inhibitory activity of p27(kip1) in vitro in a cyclinE/cdk2 kinase assay. This novel feature of the binding specificity of p27(kip1) to cyclins and cdk's in vivo is interpreted in the context of overexpression of cyclin D3 in the presence of high levels of p27(kip1) in human B-cell lymphomas with adverse clinical outcome. PMID- 15665119 TI - A pathobiologic pathway linking thrombopoietin, GATA-1, and TGF-beta1 in the development of myelofibrosis. AB - Idiopathic myelofibrosis (IM) is a disease characterized by marrow fibrosis, abnormal stem/progenitor cell trafficking, and extramedullary hematopoiesis frequently associated with alterations in megakaryocytes (Mks). Mice harboring genetic alterations in either the extrinsic (ectopic thrombopoietin expression, TPO(high) mice) or intrinsic (hypomorphic GATA-1 mutation, GATA-1(low) mice) control of Mk differentiation develop myelofibrosis, a syndrome similar to IM. The relationship, if any, between the pathobiologic mechanism leading to the development of myelofibrosis in the 2 animal models is not understood. Here we show that plasma from GATA-1(low) mice contained normal levels of TPO. On the other hand, Mks from TPO-treated wild-type animals (TPO(high) mice), as those from GATA-1(low) animals, had similar morphologic abnormalities and contained low GATA-1. In both animal models, development of myelofibrosis was associated with high transforming growth factor beta1 (TGF-beta1) content in extracellular fluids of marrow and spleen. Surprisingly, TPO treatment of GATA-1(low) mice restored the GATA-1 content in Mks and halted both defective thrombocytopoiesis and fibrosis. These data indicate that the TPO(high) and GATA-1(low) alterations are linked in an upstream-downstream relationship along a pathobiologic pathway leading to development of myelofibrosis in mice and, possibly, of IM in humans. PMID- 15665121 TI - A mathematical model of protein degradation by the proteasome. AB - The proteasome is the major protease for intracellular protein degradation. The influx rate of protein substrates and the exit rate of the fragments/products are regulated by the size of the axial channels. Opening the channels is known to increase the overall degradation rate and to change the length distribution of fragments. We develop a mathematical model with a flux that depends on the gate size and a phenomenological cleavage mechanism. The model has Michaelis-Menten kinetics with a V(max) that is inversely related to the length of the substrate, as observed in the in vitro experiments. We study the distribution of fragment lengths assuming that proteasomal cleavage takes place at a preferred distance from the ends of a protein fragment, and find multipeaked fragment length distributions similar to those found experimentally. Opening the gates in the model increases the degradation rate, increases the average length of the fragments, and increases the peak in the distribution around a length of 8-10 amino acids. This behavior is also observed in immunoproteasomes equipped with PA28. Finally, we study the effect of re-entry of processed fragments in the degradation kinetics and conclude that re-entry is only expected to affect the cleavage dynamics when short fragments enter the proteasome much faster than the original substrate. In summary, the model proposed in this study captures the known characteristics of proteasomal degradation, and can therefore help to quantify MHC class I antigen processing and presentation. PMID- 15665122 TI - Role of actin DNase-I-binding loop in myosin subfragment 1-induced polymerization of G-actin: implications for the mechanism of polymerization. AB - Proteolytic cleavage of actin between Gly(42) and Val(43) within its DNase-I binding loop (D-loop) abolishes the ability of Ca-G-actin to spontaneously polymerize in the presence of KCl. Here we show that such modified actin is assembled into filaments, albeit at a lower rate than unmodified actin, by myosin subfragment 1 (S1) carrying the A1 essential light chain but not by S1(A2). S1 titration of pyrene-G-actin showed a diminished affinity of cleaved actin for S1, but this could be compensated for by using S1 in excess. The most significant effect of the cleavage, revealed by measuring the fluorescence of pyrene-actin and light-scattering intensities as a function of actin concentration at saturating concentrations of S1, is strong inhibition of association of G-actin S1 complexes into oligomers. Measurements of the fluorescence of dansyl cadaverine attached to Gln(41) indicate substantial inhibition of the initial association of G-actin-S1 into longitudinal dimers. The data provide experimental evidence for the critical role of D-loop conformation in both longitudinal and lateral, cross-strand actin-actin contact formation in the nucleation reaction. Electron microscopic analysis of the changes in filament-length distribution during polymerization of actin by S1(A1) and S1(A2) suggests that the mechanism of S1-induced polymerization is not substantially different from the nucleation elongation scheme of spontaneous actin polymerization. PMID- 15665123 TI - Paradoxical loss of excitation with high intensity pulses during electric field stimulation of single cardiac cells. AB - Transmembrane potential responses of single cardiac cells stimulated at rest were studied with uniform rectangular field pulses having durations of 0.5-10 ms. Cells were enzymatically isolated from guinea pig ventricles, stained with voltage sensitive dye di-8-ANEPPS, and stimulated along their long axes. Fluorescence signals were recorded with spatial resolution of 17 microm for up to 11 sites along the cell. With 5 and 10 ms pulses, all cells (n = 10) fired an action potential over a broad range of field amplitudes (approximately 3-65 V/cm). With 0.5 and 1 ms pulses, all cells (n = 7) fired an action potential for field amplitudes ranging from the threshold value (approximately 4-8 V/cm) to 50 60 V/cm. However, when the field amplitude was further increased, five of seven cells failed to fire an action potential. We postulated that this paradoxical loss of excitation for higher amplitude field pulses is the result of nonuniform polarization of the cell membrane under conditions of electric field stimulation, and a counterbalancing interplay between sodium current and inwardly rectifying potassium current with increasing field strength. This hypothesis was verified using computer simulations of a field-stimulated guinea pig ventricular cell. In conclusion, we show that for stimulation with short-duration pulses, cells can be excited for fields ranging between a low amplitude excitation threshold and a high amplitude threshold above which the excitation is suppressed. These results can have implications for the mechanistic understanding of defibrillation outcome, especially in the setting of diseased myocardium. PMID- 15665125 TI - The influence of protein-protein interactions on the organization of proteins within thylakoid membranes. AB - The influence of attractive protein-protein interactions on the organization of photosynthetic proteins within the thylakoid membrane was investigated. Protein protein interactions were simulated using Monte Carlo techniques and the influence of different interaction energies was examined. It was found that weak interactions led to protein clusters whereas strong interactions led to ramified chains. An optimum curve for the relationship between interaction energy and the number of contact sites emerged. With increasing particle densities the effect decreased. In a mixture of interacting and noninteracting particles the distance between the noninteracting particles was increased and there seemed to be much more free space around them. In thylakoids, this could lead to a more homogeneous distribution of the noninteracting but rate-limiting cytochrome bf complexes. Due to the increased free space between cytochrome bf, obstruction of binding sites- occurring unavoidably in a random distribution--may be drastically reduced. Furthermore, protein-protein interactions in thylakoids may lead to a decrease in plastoquinone diffusion. PMID- 15665124 TI - Probing the viscoelastic behavior of cultured airway smooth muscle cells with atomic force microscopy: stiffening induced by contractile agonist. AB - Complex rheology of airway smooth muscle cells and its dynamic response during contractile stimulation involves many molecular processes, foremost of which are actomyosin cross-bridge cycling and actin polymerization. With an atomic force microscope, we tracked the spatial and temporal variations of the viscoelastic properties of cultured airway smooth muscle cells. Elasticity mapping identified stiff structural elements of the cytoskeletal network. Using a precisely positioned microscale probe, picoNewton forces and nanometer level indentation modulations were applied to cell surfaces at frequencies ranging from 0.5 to 100 Hz. The resulting elastic storage modulus (G') and dissipative modulus (G'') increased dramatically, with hysteresivity (eta = G''/G') showing a definitive decrease after stimulation with the contractile agonist 5-hydroxytryptamine. Frequency-dependent assays showed weak power-law structural damping behavior and universal scaling in support of the soft-glassy material description of cellular biophysics. Additionally, a high-frequency component of the loss modulus (attributed to cellular Newtonian viscosity) increased fourfold during the contractile process. The complex shear modulus showed a strong sensitivity to the degree of actin polymerization. Inhibitors of myosin light chain kinase activity had little effect on the stiffening response to contractile stimulation. Thus, our measurements appear to be particularly well suited for characterization of dynamic actin rheology during airway smooth muscle contraction. PMID- 15665126 TI - Surface changes of the mechanosensitive channel MscS upon its activation, inactivation, and closing. AB - MscS is a bacterial mechanosensitive channel that shows voltage dependence. The crystal structure of MscS revealed that the channel is a homoheptamer with a large chamber on the intracellular site. Our previous experiments indicated that the cytoplasmic chamber of the channel is not a rigid structure and changes its conformation upon the channel activation. In this study, we have applied various sized cosolvents that are excluded from protein surfaces. It is well known that such cosolvents induce compaction of proteins and prevent thermal fluctuations. It is also known that they shift channel equilibrium to the state of lower volume. We have found that large cosolvents that cannot enter the channel interior accelerate channel inactivation when applied from the cytoplasmic side, but they slow down inactivation when applied from the extracellular side. We have also found that small cosolvents that can enter the channel cytoplasmic chamber prevent the channel from opening, unlike the large ones. These data support our idea that the channel cytoplasmic chamber shrinks upon inactivation but also give new clues about conformational changes of the channel upon transitions between its functional states. PMID- 15665127 TI - Molecular dynamics simulations indicate a possible role of parallel beta-helices in seeded aggregation of poly-Gln. AB - The molecular structures of amyloid fibers characterizing neurodegenerative diseases such as Huntington's or transmissible spongiform encephalopathies are unknown. Recently, x-ray diffraction patterns of poly-Gln fibers and electron microscopy images of two-dimensional crystals formed from building blocks of prion rods have suggested that the corresponding amyloid fibers are generated by the aggregation of parallel beta-helices. To explore this intriguing concept, we study the stability of small beta-helices in aqueous solution by molecular dynamics simulations. In particular, for the Huntington aggregation nucleus, which is thought to be formed of poly-Gln polymers, we show that three-coiled beta-helices are unstable at the suggested circular geometries and stable at a triangular shape with 18 residues per coil. Moreover, we demonstrate that individually unstable two-coiled triangular poly-Gln beta-helices become stabilized upon dimerization, suggesting that seeded aggregation of Huntington amyloids requires dimers of at least 36 Gln repeats (or monomers of approximately 54 Gln) for the formation of sufficiently stable aggregation nuclei. An analysis of our results and of sequences occurring in native beta-helices leads us to the proposal of a revised model for the PrP(Sc) aggregation nucleus. PMID- 15665128 TI - Exploring the helix-coil transition via all-atom equilibrium ensemble simulations. AB - The ensemble folding of two 21-residue alpha-helical peptides has been studied using all-atom simulations under several variants of the AMBER potential in explicit solvent using a global distributed computing network. Our extensive sampling, orders of magnitude greater than the experimental folding time, results in complete convergence to ensemble equilibrium. This allows for a quantitative assessment of these potentials, including a new variant of the AMBER-99 force field, denoted AMBER-99 phi, which shows improved agreement with experimental kinetic and thermodynamic measurements. From bulk analysis of the simulated AMBER 99 phi equilibrium, we find that the folding landscape is pseudo-two-state, with complexity arising from the broad, shallow character of the "native" and "unfolded" regions of the phase space. Each of these macrostates allows for configurational diffusion among a diverse ensemble of conformational microstates with greatly varying helical content and molecular size. Indeed, the observed structural dynamics are better represented as a conformational diffusion than as a simple exponential process, and equilibrium transition rates spanning several orders of magnitude are reported. After multiple nucleation steps, on average, helix formation proceeds via a kinetic "alignment" phase in which two or more short, low-entropy helical segments form a more ideal, single-helix structure. PMID- 15665129 TI - Surface density dependence of PCR amplicon hybridization on PNA/DNA probe layers. AB - Surface plasmon field-enhanced fluorescence spectroscopy was employed to extensively investigate the hybridization behaviors of polymerase chain reaction (PCR) amplicons on a peptide nucleic acid (PNA) or DNA probe layer that was previously attached on a streptavidin-modified gold surface via biotin/streptavidin interaction. Despite the neutral backbone of PNA, the hybridization reactions were strongly influenced by the variation of ionic strength. The association rates exhibited a monotonic decrease with ionic strength increase and the maximum hybridization signal was achieved at an intermediate sodium concentration (approximately 100 mM). These effects were mainly ascribed to the electrostatic cross talk among the hybridized DNA molecules and the secondary structure of PCR amplicons. For the negatively charged DNA probes, the hybridization reaction was subjected additionally to the DNA/DNA electrostatic barrier, particularly in lower ionic strength range (e.g., 10 approximately 150 mM Na(+)). The electrostatic cross talk was shown to be largely reduced if the PNA probe layer was sufficiently diluted by following a strategic templated immobilization method. As a consequence, a pseudo-first-order kinetic model was applicable to describe the hybridization kinetics, and affinity constants were derived for evaluating the influence of single nucleotide polymorphisms (SNPs). PMID- 15665130 TI - Measuring forces between protein fibers by microscopy. AB - We propose a general scheme for measuring the attraction between mechanically frustrated semiflexible fibers by measuring their thermal fluctuations and shape. We apply this analysis to a system of sickle hemoglobin (HbS) fibers that laterally attract one another. These fibers appear to "zip" together before reaching mechanical equilibrium due to the existence of cross-links into a dilute fiber network. We are also able to estimate the rigidities of the fibers. These rigidities are found to be consistent with sickle hemoglobin "single" fibers 20 nm in diameter, despite recent experiments indicating that fiber bundling sometimes occurs. Our estimate of the magnitude of the interfiber attraction for HbS fibers is in the range 8 +/- 7 kBT/microm, or 4 +/- 3 k(B)T/microm if the fibers are assumed, a priori to be single fibers (such an assumption is fully consistent with the data). This value is sufficient to bind the fibers, overcoming entropic effects, although extremely chemically weak. Our results are compared to models for the interfiber attraction that include depletion and van der Waals forces. This technique should also facilitate a similar analysis of other filamentous protein assembles in the future, including beta-amyloid, actin, and tubulin. PMID- 15665131 TI - Structure of fully hydrated fluid phase DMPC and DLPC lipid bilayers using X-ray scattering from oriented multilamellar arrays and from unilamellar vesicles. AB - Quantitative structures of the fully hydrated fluid phases of dimyristoylphosphatidylcholine (DMPC) and dilauroylphosphatidylcholine (DLPC) were obtained at 30 degrees C. Data for the relative form factors F(q(z)) for DMPC were obtained using a combination of four methods. 1), Volumetric data provided F(0). 2), Diffuse x-ray scattering from oriented stacks of bilayers provided relative form factors |F(q(z))| for high q(z), 0.22 < q(z) < 0.8 A(-1). 3), X-ray scattering from extruded unilamellar vesicles with diameter 600 A provided |F(q(z))| for low q(z), 0.1 < q(z) < 0.3 A(-1). 4), Previous measurements using a liquid crystallographic x-ray method provided |F(2 pi h/D)| for h = 1 and 2 for a range of nearly fully hydrated D-spacings. The data from method 4 overlap and validate the new unilamellar vesicles data for DMPC, so method 4 is not required for DLPC or future studies. We used hybrid electron density models to obtain structural results from these form factors. Comparison of the model electron density profiles with that of gel phase DMPC provides areas per lipid A, 60.6 +/- 0.5 A(2) for DMPC and 63.2 +/- 0.5 A(2) for DLPC. Constraints on the model provided by volume measurements and component volumes obtained from simulations put the electron density profiles rho(z) and the corresponding form factors F(q(z)) on absolute scales. Various thicknesses, such as the hydrophobic thickness and the steric thickness, are obtained and compared to literature values. PMID- 15665132 TI - Protein self-association induced by macromolecular crowding: a quantitative analysis by magnetic relaxation dispersion. AB - In the presence of high concentrations of inert macromolecules, the self association of proteins is strongly enhanced through an entropic, excluded-volume effect variously called macromolecular crowding or depletion attraction. Despite the predicted large magnitude of this universal effect and its far-reaching biological implications, few experimental studies of macromolecular crowding have been reported. Here, we introduce a powerful new technique, fast field-cycling magnetic relaxation dispersion, for investigating crowding effects on protein self-association equilibria. By recording the solvent proton spin relaxation rate over a wide range of magnetic field strengths, we determine the populations of coexisting monomers and decamers of bovine pancreatic trypsin inhibitor in the presence of dextran up to a macromolecular volume fraction of 27%. Already at a dextran volume fraction of 14%, we find a 30-fold increase of the decamer population and 510(5)-fold increase of the association constant. The analysis of these results, in terms of a statistical-mechanical model that incorporates polymer flexibility as well as the excluded volume of the protein, shows that the dramatic enhancement of bovine pancreatic trypsin inhibitor self-association can be quantitatively rationalized in terms of hard repulsive interactions. PMID- 15665134 TI - MSP dynamics drives nematode sperm locomotion. AB - Most eukaryotic cells can crawl over surfaces. In general, this motility requires three sequential actions: polymerization at the leading edge, adhesion to the substrate, and retraction at the rear. Recent in vitro experiments with extracts from spermatozoa from the nematode Ascaris suum suggest that retraction forces are generated by depolymerization of the major sperm protein cytoskeleton. Combining polymer entropy with a simple kinetic model for disassembly we propose a model for disassembly-induced retraction that fits the in vitro experimental data. This model explains the mechanism by which disassembly of the cytoskeleton generates the force necessary to pull the cell body forward and suggests further experiments that can test the validity of the models. PMID- 15665133 TI - Determination of the activation volume of PLCbeta by Gbeta gamma-subunits through the use of high hydrostatic pressure. AB - Activation of phospholipase Cbeta (PLCbeta) by G-proteins results in increased intracellular Ca(2+) and activation of protein kinase C. We have previously found that activated PLCbeta-Gbetagamma complex can be rapidly deactivated by Galpha(GDP) subunits without dissociation, which led to the suggestion that Galpha(GDP) binds to PLCbeta-Gbeta gamma and perturbs the activating interaction without significantly affecting the PLCbeta-Gbeta gamma binding energy. Here, we have used high pressure fluorescence spectroscopy to determine the volume change associated with this interaction. Since PLCbeta and G-protein subunits associate on membrane surfaces, we worked under conditions where the membrane surface properties are not expected to change. We also determined the pressure range in which the proteins remain membrane bound: PLCbeta binding was stable throughout the 1-2000 bars range, Gbeta gamma binding was stable only at high membrane concentrations, whereas Galpha(s)(GDP) dissociated from membranes above 1 kbar. High pressure dissociated PLCbeta-Gbeta gamma with a DeltaV = 34 +/- 5 ml/mol. This same volume change is obtained for a peptide derived from Gbeta which also activates PLCbeta. In the presence of Galpha(s)(GDP), the volume change associated with PLCbeta-Gbeta gamma interaction is reduced to 25 +/- 1 ml/mol. These results suggest that activation of PLCbeta by Gbeta gamma is conferred by a small (i.e., 3-15 ml/mol) volume element. PMID- 15665136 TI - Functional activity of the M2 and M4 receptor subtypes in the spinal cord studied with muscarinic acetylcholine receptor knockout mice. AB - Stimulation of spinal muscarinic acetylcholine receptors (mAChRs) produces potent analgesia. Both M(2) and M(4) mAChRs are coupled to similar G proteins (G(i/o) family) and play a critical role in the analgesic action of mAChR agonists. To determine the relative contribution of M(2) and M(4) subtypes to activation of G(i/o) proteins in the spinal cord, we examined the receptor-mediated guanosine 5'-O-(3-[(35)S]thio)triphosphate ([(35)S]GTPgammaS) binding in M(2) and M(4) subtype knockout (KO) mice. Basal [(35)S]GTPgammaS binding in the spinal cord was similar in the wild-type controls, M(2) and M(4) single-KO, and M(2)/M(4) double KO mice. The spinal [(35)S]GTPgammaS binding stimulated by either muscarine or oxotremorine-M was not significantly different among three groups of wild-type mouse strains. In M(2) single-KO and M(2)/M(4) double-KO mice, the agonist stimulated [(35)S]GTPgammaS binding was completely abolished in the spinal cord. Furthermore, the agonist-stimulated [(35)S]GTPgammaS binding in the spinal cord of M(4) single-KO mice was significantly reduced ( approximately 15%), compared with that in wild-type controls. On the other hand, the spinal [(35)S]GTPgammaS binding stimulated by a mu-opioid agonist was not significantly different between wild-type and M(2) and M(4) KO mice. This study provides complementary new evidence that M(2) is the most predominant mAChR subtype coupled to the G(i/o) proteins in the spinal cord. Furthermore, these data suggest that a small but functionally significant population of M(4) receptors exists in the mouse spinal cord. The functional activity of these M(4) receptors seems to require the presence of M(2) receptors. PMID- 15665135 TI - Ligand-induced conformational change in the alpha7 nicotinic receptor ligand binding domain. AB - Molecular dynamics simulations of a homology model of the ligand binding domain of the alpha7 nicotinic receptor are conducted with a range of bound ligands to induce different conformational states. Four simulations of 15 ns each are run with no ligand, antagonist d-tubocurarine (dTC), agonist acetylcholine (ACh), and agonist ACh with potentiator Ca(2+), to give insight into the conformations of the active and inactive states of the receptor and suggest the mechanism for conformational change. The main structural factor distinguishing the active and inactive states is that a more open, symmetric arrangement of the five subunits arises for the two agonist simulations, whereas a more closed and asymmetric arrangement results for the apo and dTC cases. Most of the difference arises in the lower portion of the ligand binding domain near its connection to the adjacent transmembrane domain. The transfer of the more open state to the transmembrane domain could then promote ion flow through the channel. Variation in how subunits pack together with no ligand bound appears to give rise to asymmetry in the apo case. The presence of dTC expands the receptor but induces rotations in alternate directions in adjacent subunits that lead to an asymmetric arrangement as in the apo case. Ca(2+) appears to promote a slightly greater expansion in the subunits than ACh alone by stabilizing the C-loop and ACh positions. Although the simulations are unlikely to be long enough to view the full conformational changes between open and closed states, a collection of different motions at a range of length scales are observed that are likely to participate in the conformational change. PMID- 15665137 TI - RH1 induces cellular damage in an NAD(P)H:quinone oxidoreductase 1-dependent manner: relationship between DNA cross-linking, cell cycle perturbations, and apoptosis. AB - Structure-based development of NAD(P)H:quinone oxidoreductase (NQO1)-directed antitumor quinones resulted in development of RH1 [2,5-diaziridinyl-3 (hydroxymethyl)-6-methyl-1,4-benzoquinone], a methyl-substituted diaziridinyl quinone. We conducted experiments to evaluate the mechanism of RH1-induced cytotoxicity and the inter-relationship between DNA cross-linking, cell cycle changes, and apoptosis using an isogenic cell line pair developed from the human breast cancer cell line MDA-MB-468 differing only in expression of wtNQO1 (NQ16 cells). Statistically significant DNA cross-linking was detected using a modified comet assay in cells with wtNQO1 within 1 h of dosing, whereas in parental cells, only marginal DNA cross-linking was observed and required a concentration up to 50 times higher. Cross-linking in NQ16 cells could be abrogated with 5-methoxy 1,2-dimethyl-3-[(4-nitrophenoxy)methyl]indole-4,7-dione, a mechanism-based inhibitor of NQO1. RH1 prolonged S phase and caused a G(2)/M block. Cell cycle changes were observed up to 10-fold lower in RH1 concentrations in NQ16 cells relative to parental cells. Apoptosis was similarly observed morphologically in both cell lines after RH1 treatment but was induced preferentially in NQ16 cells at lower concentrations and earlier time points. Marked cleavage of caspase-3 was observed in NQ16 cells relative to parental cells using lower concentrations of RH1. Temporally, low doses of RH1-induced rapid DNA cross-linking in NQ16 cells followed by induction of apoptosis at times when a G(2)/M block was not observed. This suggests that cell cycle arrest is not required for RH1-induced apoptosis and that DNA damage may directly initiate apoptotic events. In summary, RH1 induced preferential DNA cross-linking, cell cycle changes, and apoptosis in an NQO1-dependent manner. PMID- 15665138 TI - Glycogen synthase kinase-3 mediates acetaminophen-induced apoptosis in human hepatoma cells. AB - The mild analgesic drug acetaminophen (AAP) induces severe hepatic injury when taken at excessive doses. Recent evidence shows that the initial form of damage is through apoptosis, but this fails to go to completion and degenerates into necrosis. The aim of this study was to elucidate the mechanism through which AAP induces apoptosis using human HuH7 hepatoma cells as an in vitro model system to investigate the initial phase of AAP-induced hepatic injury. AAP-induced apoptosis in HuH7 cells as evidenced by chromatin condensation was preceded by the translocation of Bax to mitochondria and the cytoplasmic release of the proapoptotic factors cytochrome c and Smac/DIABLO. A concomitant loss of mitochondrial membrane potential occurred. Activation of the mitochondrial pathway of apoptosis led to the activation of execution caspases-3 and -7. AAP induced apoptosis and cell death was blocked by inhibitors of caspases but not by inhibitors of calpains, cathepsins, and serine proteases. Apoptosis was unaffected by inhibitors of the mitochondrial permeability transition pore and by inhibitors of Jun NH(2)-terminal kinases, p38 mitogen-activated protein kinase, or mitogen-activated protein kinase kinase 1/2. However, pharmacological inhibition of glycogen synthase kinase-3 (GSK-3) delayed and decreased the extent of AAP-induced apoptosis. In comparison, endoplasmic reticulum stress-induced but not prooxidant-induced apoptosis of HuH7 cells was sensitive to GSK-3 inhibition. It is concluded that AAP-induced apoptosis involves the mitochondrial pathway of apoptosis that is mediated by GSK-3 and most likely initiated through an endoplasmic reticulum stress response. PMID- 15665139 TI - Vectorial transport of the peptide CCK-8 by double-transfected MDCKII cells stably expressing the organic anion transporter OATP1B3 (OATP8) and the export pump ABCC2. AB - CCK-8 (L-aspartyl-L-tyrosyl-L-methionylglycyl-L-tryptophyl-L-methionyl-L-aspartyl L-phenylalaninamide hydrogen sulfate ester), a derivative of the gastrointestinal peptide hormone cholecystokinin, is specifically taken up into human hepatocytes by the organic anion transporter OATP1B3 (OATP8). So far it was unknown which transporter mediates the excretion of CCK-8 into bile. Double-transfected Madin Darby canine kidney strain II cells, expressing recombinant human OATP1B3 in the basolateral membrane together with human ABCC2 (multidrug resistance protein 2, MRP2) in the apical membrane, represent a valuable model system to study vectorial transport. The importance of an appropriate filter support for optimized protein localization and substrate transport was demonstrated by the comparison of filter pore densities of 2 x 10(6) and 1 x 10(8) per cm(2). At the high pore density, immunofluorescence microscopy showed an intense OATP1B3 signal in the basolateral membrane of all cells, and 82 +/- 8% of cells expressed ABCC2 in the apical membrane. Uptake and efflux of radiolabeled CCK-8 in the double transfected cells grown at high pore density was enhanced 3.5- and 5.6-fold, respectively, compared with cells grown at lower pore density. Higher transport rates were also observed with [(3)H]bromosulfophthalein. The high-affinity ATP dependent transport of CCK-8 by ABCC2 was directly demonstrated in ABCC2 containing membrane vesicles with a K(m) value of 8.1 microM. The uptake by OATP1B3 and hence the vectorial transport of CCK-8 was inhibited by cyclosporin A (K(i) 1.2 microM) and by MK571 [(3-(3-(2-(7-chloro-2-quinolinyl)ethenyl)phenyl) ((3-dimethylamino-3-oxopropyl)thio)methyl)thiopropanoic acid] (K(i) 0.6 microM); the respective K(i) values for the ABCC2-mediated transport were 24 and 8.5 microM. Thus, using an optimized filter support, we demonstrate vectorial transport of CCK-8 by OATP1B3 and by the apical export pump ABCC2. PMID- 15665141 TI - Mechanism of bradykinin-induced Ca2+ mobilization in murine proximal tubule epithelial cells. AB - Despite the recognized physiological role of bradykinin (BK) in the kidney in maintaining glomerular and tubule function and its role in pathological states such as endotoxemia, diabetes, and other diseases, relatively little is known about the mechanisms by which BK can impact kidney function. Furthermore, the signaling of BK receptors in the murine nephron has not been fully characterized. The present studies were undertaken to examine BK-stimulated Ca(2+) signaling using Fura-2 in the murine proximal tubule epithelial cell line TKPTS. BK produced a concentration-dependent rise in intracellular Ca(2+) ([Ca(2+)])(i) (pEC(50) = 8.39 +/- 0.04). Selective antagonists showed the rise in [Ca(2+)](i) was mediated through B2 receptors. The rise in [Ca(2+)](i) was rapid and reversible and was maximally stimulated at 1 microM (697 +/- 70 nM above basal level of 115 +/- 6 nM). Studies with thapsigargin and EGTA showed Ca(2+) mobilization was dependent on two events: release and influx. Both U73122 (1-[6 [[17-beta-3-methoxyestra-1,3,5(10)-trien-17-yl]amino]hexyl]-1H-pyrrole-2,5-dione) [a phospholipase C (PLC) inhibitor] and genistein (a tyrosine kinase inhibitor) partially inhibited BK-stimulated rise in [Ca(2+)](i). When combined, both agents produced a further decrease, suggesting multiple pathways for PLC activation may be involved. The ability of Ni(2+) to inhibit influx indicated the activation of a Ca(2+) release-activated channel (CRAC). Ca(2+) mobilization did not seem to be affected by cyclic nucleotides or protein kinase C. In summary, the TKPTS murine proximal tubule cell line expresses functional B2 receptors linked to Ca(2+) mobilization that is dependent on phospholipase C and activation of CRAC. PMID- 15665140 TI - Specific antibody modulates absorptive transport and metabolic activation of benzo[a]pyrene across Caco-2 monolayers. AB - It has been shown that oral anticarcinogen antibodies can decrease intestinal absorption of procarcinogens. So far, no attempts have been made to understand the potential modulatory effect of such antibodies on metabolic activation at mucosal surfaces. Moreover, the influence of naturally induced serosal-specific antibodies on absorptive transport of carcinogens remains unknown. In this study, the prototype food carcinogen benzo[a]pyrene (B[a]P) and a specific monoclonal antibody were used to address these questions in a bicompartmental model of polarized intestinal cells (Caco-2). Apical (i.e., luminal) administration of a 30-fold molar excess antibodies increased about 25-fold recovery of unmetabolized B[a]P, concomitantly with a decrease of 80% in both absorptive transport and formation of phenol metabolites. Interestingly, when metabolism was slowed down by antibodies, cross-reactive antibodies also increased at least 5-fold the extracellular levels of the 7,8-diol-B[a]P, interrupting subsequent activation steps. On the other hand, basolateral antibodies changed by 8-fold the rate of carcinogen appearance in the basolateral compartment, albeit without interfering with the apical cellular uptake or sequestration of either B[a]P or 7,8-diol B[a]P by apical antibodies. Furthermore, basolateral antibodies reduced exposure of Caco-2 monolayers to B[a]P as demonstrated by a 50% decrease in apical efflux of 3-OH-B[a]P. These data show for the first time that both luminal and serosal antibodies may reduce the carcinogenic process by preventing high local concentrations, which would overload DNA repair mechanisms. This study also sheds light on the relevance of both naturally induced and immunoprophylactic antibodies against polycyclic aromatic hydrocarbon carcinogens. PMID- 15665142 TI - Mechanism of myricetin stimulation of vascular L-type Ca2+ current. AB - An in-depth analysis of the mechanism of the L-type Ca(2+) current [I(Ca(L))] stimulation induced by myricetin was performed in rat tail artery myocytes using the whole-cell patch-clamp method. Myricetin increased I(Ca(L)) in a frequency-, concentration-, and voltage-dependent manner. At holding potentials (V(h)) of -50 and -90 mV, the pEC(50) values were 4.9 +/- 0.1 and 4.2 +/- 0.1, respectively; the latter corresponded to the drug-apparent dissociation constant for resting channels, K(R), of 67.6 microM. Myricetin shifted the maximum of the current voltage relationship by 10 mV in the hyperpolarizing direction but did not modify the threshold for I(Ca(L)) or the T-type Ca(2+) current. The Ca(2+) channel blockers nifedipine, verapamil, and diltiazem antagonized I(Ca(L)) in the presence of myricetin. Myricetin increased the time to peak of I(Ca(L)) in a voltage- and concentration-dependent manner. Washout reverted myricetin effect on both current kinetics and amplitude at V(h) of -90 mV while reverting only current kinetics at V(h) of -50 mV. At the latter V(h), myricetin shifted the voltage dependence of inactivation and activation curves to more negative potentials by 6.4 and 13.0 mV, respectively, in the mid-potential of the curves. At V(h) of -90 mV, myricetin shifted, in a concentration-dependent manner, the voltage dependence of the inactivation curve to more negative potentials with an apparent dissociation constant for inactivated channels (K(I)) of 13.8 muM. Myricetin induced a frequency- and V(h)-dependent block of I(Ca(L)). In conclusion, myricetin behaves as an L-type Ca(2+) channel agonist that stabilizes the channel in its inactivated state. PMID- 15665143 TI - Rosuvastatin attenuates monocyte-endothelial cell interactions and vascular free radical production in hypercholesterolemic mice. AB - One of the earliest observable events in atherogenesis is enhanced monocyte adhesion to the endothelium. In addition to reducing circulating levels of cholesterol, 3-hydroxy-3-methylglutaryl-CoA reductase inhibitors (statins) are thought to have direct salutary effects upon vascular cells. We hypothesized that the new statin, rosuvastatin, would have anti-inflammatory effects on the vessel wall. Eight-week-old apolipoprotein E-deficient mice were fed a normal chow diet for a period of 12 weeks. During this time mice were administered vehicle or rosuvastatin at a dose of 0, 1, 5, or 20 mg/kg by subcutaneous injection at the same time daily for a period of 2 or 6 weeks prior to sacrifice. At the end of the study, rosuvastatin-treated animals displayed lower plasma total cholesterol levels, whereas showing little change in high-density lipoprotein cholesterol or triglycerides. Using a functional binding assay, we also demonstrated that endothelial adhesiveness for monocytes was significantly attenuated after 2 weeks of treatment with rosuvastatin. Quantitative real-time polymerase chain reaction determined that rosuvastatin reduced the expression of vascular cell adhesion molecule-1, monocyte chemotactic protein-1, and metalloproteinase-9 in the vessel wall. In addition, rosuvastatin inhibited vascular expression of p22(phox) and superoxide production, as well as diminishing plasma 8-isoprostanes concentrations. Thus, treatment with rosuvastatin has acute anti-inflammatory actions that likely participate in its beneficial actions during atherogenesis. PMID- 15665144 TI - The nonthiazolidinedione tyrosine-based peroxisome proliferator-activated receptor gamma ligand GW7845 induces apoptosis and limits migration and invasion of rat and human glioma cells. AB - Despite new approaches, treatment options for malignant gliomas are still limited, calling for further development of therapeutic strategies. The peroxisome proliferator-activated receptor (PPAR)gamma, a member of the nuclear hormone receptor family, represents a possible new target for neoplastic therapies. Synthetic PPARgamma agonists were developed and are already in clinical use for the treatment of type II diabetes, since PPARgamma plays a crucial role in lipid metabolism and regulation of insulin sensitivity. Beyond these metabolic effects, PPARgamma agonists exhibit antineoplastic effects in various malignant tumor cells. Here, we investigated the antineoplastic effects of the nonthiazolidinedione tyrosine-based PPARgamma ligand (S)-2-(1-carboxy-2-{4 [2-(5-methyl-2-phenyloxazol-4-yl)ethoxy]phenyl}ethylamino)benzoic acid methyl ester (GW7845) in rat and human glioma cells. GW7845 reduced cellular viability of rat C6 glioma and human glioma cells in a time-dependent manner. Analysis of GW7845-treated tumor cells revealed induction of apoptotic cell death as determined by terminal deoxynucleotidyl transferase dUTP nick-end labeling staining and cleaved caspase-3 activation. Furthermore, GW7845 reduced proliferation of C6 glioma cells as measured by Ki-67 immunore-activity. There was also a reduction of migration and invasion, assessed by Boyden chamber and spheroid experiments. Together, these data indicate that the PPARgamma agonist GW7845 may be of potential use in treatment of malignant gliomas. PMID- 15665146 TI - R-R interval-blood pressure interaction in subjects with different tolerances to orthostatic stress. AB - In addition to the gain, the time delay in the input-output response in a feedback system is crucial for the maintenance of its stability. Patients with posturally related (vasovagal) syncope have inadequate control of blood pressure and one possible explanation for this could be prolonged latency of the baroreflex. We studied 14 patients with histories of syncope and poor orthostatic tolerance (assessed by a progressive orthostatic stress test) and 16 healthy controls. We performed spontaneous sequence analysis of the fluctuations of R-R period (ECG) and systolic arterial pressure (SAP, Finapres) recorded during a 20 min supine period and during 20 min 60 deg head-up tilt (HUT). The baroreflex latency was determined by identifying the lag between the changes in SAP and in R R interval from which the highest correlation coefficient was obtained. During the supine period, 74% of sequences in control subjects and 54% in patients occurred with zero beats of delay (i.e. R-R interval changed within the same R-R interval). The remaining sequences occurred with delays of up to four beats. HUT shifted the baroreflex delay to be approximately one heartbeat slower and again patients showed more sequences with prolonged response. The delay in heartbeats was transformed into delay in time. In control subjects, 75% of baroreflex responses occurred within 1 s. In patients, 75% of baroreflex responses took more than 2 s to occur. The results showed that syncopal patients with poor orthostatic tolerance have increased baroreflex latency. This may lead to instability and inadequate blood pressure control and may predispose to vasovagal syncope. PMID- 15665147 TI - The initial phase of exercise hyperpnoea in humans is depressed during a cognitive task. AB - Increased wakefulness is known to suppress the initial ventilatory response to passive movement and the steady-state ventilatory response to exercise. However, the effect of increased wakefulness upon the integrated ventilatory response at the onset of exercise is not known. We hypothesized that increasing wakefulness via a cognitive task would attenuate the initial ventilatory response to exercise, and so we examined the response to active leg extensions under two conditions: with and without concurrently solving a puzzle. At rest before exercise, subjects demonstrated greater minute ventilation while solving a puzzle (mean +/- S.E.M., 12.38 +/- 0.55 versus 10.12 +/- 0.51 l min(-1), P < 0.001), due to a higher mean breathing frequency (mean +/- S.E.M., 17.1 +/- 0.93 versus 13.6 +/- 0.59 breaths min(-1), P < 0.001). At the start of exercise, subjects did not increase their ventilation significantly while solving the puzzle (P = 0.170), but did by a mean +/-s.e.m. of 6.16 +/- 1.12 l min(-1) (P < 0.001) when not puzzle solving. The ventilation achieved at the start of exercise in absolute terms was also lower while solving the puzzle (14.6 +/- 1.1 versus 16.3 +/- 1.3 l min(-1), P = 0.047). Despite differences in the rapid ventilatory response to exercise between conditions, the steady-state responses were not different. We conclude that the performance of a cognitive task decreases the initial phase of exercise hyperpnoea, and suggest that this might occur because of either a competitive interaction between drives to breathe or a behavioural distraction from the 'task' of exercise. PMID- 15665150 TI - A new paradigm for developing drugs in children: atomoxetine as a model. AB - When prescribing drugs for children, it is fundamental to acknowledge that they are distinct from adults with a different physiology and metabolism. They are also still maturing. The development of drugs with a primarily paediatric indication thus requires the trialling of these drugs in a paediatric population to assess safety, tolerability, and efficacy as appropriately as possible. When designing and running a paediatric clinical trial, a number of complexities must be addressed to ensure a successful study, including practical considerations, ethical issues, and tailoring communication appropriately to study participants and parents. The drug development process for atomoxetine, a novel, non-stimulant treatment for attention deficit/hyperactivity disorder (ADHD), encompassed a preclinical programme, initial trials in healthy adults, and a proof of concept trial in adults with ADHD. Open label and placebo controlled studies in paediatric patients followed, thus establishing the drug's safety and efficacy in children with ADHD. Further trials have addressed, and continue to address, wider aspects of the atomoxetine response in a paediatric setting. PMID- 15665149 TI - Attention deficit/hyperactivity disorder and ethnicity. AB - Despite the large volume of research dedicated to attention deficit/hyperactivity disorder (ADHD), the influence of ethnicity on the condition is still to be adequately addressed. Previous studies have shown differing rates of ADHD between different countries (although this can be affected by the assessment tool used). Cultural environment may affect a child's behaviour, but the differing attitudes of parents, clinicians, and society around the world towards acceptable behaviour may also influence diagnosis. If assessment criteria could be consistently applied across different ethnic groups, the prevalence of ADHD would probably be similar. PMID- 15665151 TI - Off licence and off label prescribing in children: litigation fears for physicians. AB - So-called "off label" and unlicensed prescribing refers to the use of medicines outside of the indications for which they are licensed by national regulatory bodies. Off label prescribing is quite common in children, as most drugs are developed only on the basis of trials with adults. Nevertheless, physicians and hospitals can be wary of using medicines in this way for fear of litigation if adverse events occur. Given this unsatisfactory state of affairs, regulatory bodies are beginning to request robust data from pharmaceutical companies with regard to the use of their products in children. In the meantime, off label prescribing remains acceptable if there is no suitable alternative and physicians are confident that they are using agents in accordance with the body of respected medical opinion. PMID- 15665153 TI - The effect of ADHD on the life of an individual, their family, and community from preschool to adult life. AB - Attention deficit/hyperactivity disorder (ADHD) may affect all aspects of a child's life. Indeed, it impacts not only on the child, but also on parents and siblings, causing disturbances to family and marital functioning. The adverse effects of ADHD upon children and their families changes from the preschool years to primary school and adolescence, with varying aspects of the disorder being more prominent at different stages. ADHD may persist into adulthood causing disruptions to both professional and personal life. In addition, ADHD has been associated with increased healthcare costs for patients and their family members. PMID- 15665154 TI - Atomoxetine: a new pharmacotherapeutic approach in the management of attention deficit/hyperactivity disorder. AB - Atomoxetine is a novel, non-stimulant, highly selective noradrenaline reuptake inhibitor that has been studied for use in the treatment of attention deficit/hyperactivity disorder (ADHD). Data from clinical trials show it to be well tolerated and effective in the treatment of ADHD in children, adolescents, and adults. Improvements were seen not only in core symptoms of ADHD, but also in broader social and family functioning and self esteem. Once-daily dosing of atomoxetine has been shown to be effective in providing continuous symptom relief. Atomoxetine does not appear to have abuse potential and is associated with a benign side effect profile. The development of atomoxetine thus represents an important advance in the pharmacological management of ADHD. PMID- 15665152 TI - Managing attention deficit/hyperactivity disorder: unmet needs and future directions. AB - The management of attention deficit/hyperactivity disorder (ADHD) requires a multimodal approach. Although shared-care protocols are suggested, uptake by general practice has been tentative. Behavioural management, both on its own and in combination with pharmacotherapy, reduces core symptoms and associated behaviours, such as oppositional defiant disorder (ODD). Links with education and social support are also important for helping children and their families. Psychostimulants have shown to be effective in the treatment of ADHD, however, their short duration of action highlights potential limitations. Other drugs used in the management of ADHD and comorbid disorders include antidepressants, clonidine and its analogues, newer atypical neuroleptics, and atomoxetine, a novel, non-stimulant therapy that has potential to fulfil some unmet treatment needs. Further research is needed in preschool children, girls, those with epilepsy, young adults, and in those with associated sleep disturbance. Selected rating scales and careful history taking, allied to close links with schools, are vital for the initial and ongoing assessment of ADHD and its comorbidities. PMID- 15665155 TI - The parent's perspective. AB - "A wonderful son of whom I am very proud": the mother of a 14 year old boy with attention deficit/hyperactivity disorder describes her experiences. PMID- 15665156 TI - Readmission of neonates. PMID- 15665157 TI - Congenital hypothyroidism. PMID- 15665158 TI - The evolution of paediatric hospitals. PMID- 15665159 TI - Maternal phenylketonuria: the importance of early control during pregnancy. PMID- 15665160 TI - Babies of a pandemic. PMID- 15665161 TI - Early discharge and readmission to hospital in the first month of life in the Northern Region of the UK during 1998: a case cohort study. AB - AIMS: To study the frequency and associations of early postpartum discharge and infant readmission to hospital. METHODS: Infants readmitted to hospital during the first 28 days of life in 1998 in the Northern Region of the UK were studied. RESULTS: A total of 4743 of 11,338 (42%) babies were discharged on or before the first postnatal day. Rates of early discharge varied significantly between hospitals. Infants <2500 g at birth (adjusted odds ratio (AOR) 0.44, 95% CI 0.29 to 0.66), infants 35-37 weeks gestation at birth (AOR 0.65, 95% CI 0.49 to 0.86), and firstborn infants (AOR 0.09, 95% CI 0.08 to 0.10) were less likely to be discharged early. Women from more deprived areas were more likely to be discharged early (AOR 1.37, 95% CI 1.12 to 1.67). A total of 907 of 32,015 (2.8%) babies liveborn in the region were readmitted to hospital during 1998. Readmission rates varied significantly by hospital of birth but not by timing of discharge. Babies <2500 g at birth (AOR 1.95, 95% CI 1.16 to 3.28) and babies born at 35-37 weeks gestation (AOR 1.72, 95% CI 1.15 to 2.57) were more likely to be readmitted. Breast fed babies were less likely to be readmitted (AOR 0.69, 95% CI 0.53 to 0.90). Infants initially discharged early were not more likely to be readmitted. CONCLUSIONS: Early discharge occurred variably in the Northern Region in 1998. It is not associated with readmission to hospital. Breast feeding is associated with lower rates of readmission to hospital. PMID- 15665162 TI - Rehospitalisation after birth hospitalisation: patterns among infants of all gestations. AB - AIM: To analyse rehospitalisation of newborns of all gestations. METHODS: A total of 33,276 surviving infants of all gestations born between 1 October 1998 and 31 March 2000 at seven Kaiser Permanente Medical Care Program (KPMCP) delivery services were studied retrospectively. RESULTS: Rehospitalisation rates within two weeks after nursery discharge ranged from 1.0% to 3.7%. The most common reason for rehospitalisation was jaundice. Among babies > or =34 weeks, the most important factor with respect to rehospitalisation was use of home phototherapy. Among babies who were not rehospitalised for jaundice, African-American race (adjusted odds ratio (AOR) = 0.56), and having a scheduled outpatient visit (AOR = 0.73) or a home visit (AOR = 0.59) within 72 hours after discharge were protective. Factors associated with increased risk were: being small for gestational age (AOR = 1.83), gestational age of 34-36 weeks without admission to the neonatal intensive care unit (AOR = 1.65), Score for Neonatal Acute Physiology, version II, > or =10 (AOR = 1.95), male gender (AOR = 1.24), having both a home as well as a clinic visit within 72 hours after discharge (AOR = 1.84), and birth facility (range of AORs = 1.52-2.36). Asian race was associated with rehospitalisation (AOR = 1.49) when all hospitalisations were considered, but this association did not persist if hospitalisations for jaundice were excluded. CONCLUSIONS: In this insured population with access to integrated care, rehospitalisation rates for jaundice were strongly affected by availability of home phototherapy and by follow up. For other causes, moderate prematurity and follow up visits played a large role, but variation between centres persisted even after controlling for multiple factors. Future research should include development of better process measures for evaluation of follow up strategies. PMID- 15665164 TI - Hospital based alternatives to acute paediatric admission: a systematic review. AB - AIMS: To synthesise published evidence of the impacts of introducing hospital based alternatives to acute paediatric admission. METHODS: Systematic review of studies of interventions for children with acute medical problems. Main outcome measures were: admission or discharge, unscheduled returns to hospital, satisfaction of parents and general practitioners, effects on health service activity, and costs. RESULTS: Twenty five studies were included: one randomised controlled trial, 23 observational or cross-sectional studies, and one qualitative study. Many studies were of uncertain quality or were open to significant potential bias. About 40% of children attending acute assessment units in paediatric departments, and over 60% of those attending acute assessment units in A&E departments, do not require inpatient admission. There is little evidence of serious clinical consequences in children discharged from these units, although up to 7% may subsequently return to hospital. There is some evidence that users are satisfied with these services and that they are associated with reductions in inpatient activity levels and certain hospital costs. Evidence about the impact of urgent outpatient clinics is very limited. CONCLUSIONS: Current evidence supports a view that acute paediatric assessment services are a safe, efficient, and acceptable alternative to inpatient admission, but this evidence is of limited quantity and quality. Further research is required to confirm that this type of service reorganisation does not disadvantage children and their families, particularly where inpatient services are withdrawn from a hospital. PMID- 15665163 TI - Congenital hypothyroidism: no adverse effects of high dose thyroxine treatment on adult memory, attention, and behaviour. AB - BACKGROUND: In congenital hypothyroidism (CH) it has been questioned whether high dose thyroxine replacement therapy has detrimental effects on memory, attention, and behaviour. AIMS: To describe memory, attention, and behaviour problems in young adults with CH, and to study possible negative effects of high dose thyroxine replacement therapy. METHODS: A cohort based follow up study of 49 young adults (mean age 20 years) with early treated CH, and sibling controls (n = 41). RESULTS: Controlled for age and sex, the CH group attained significantly lower scores than sibling controls on some tests of memory (Wechsler Logical Memory part II: 12.9 versus 17.8; difference 5.2, 95% CI 3.6 to 6.8) and attention (Wechsler Freedom From Distractibility factor: 95.6 versus 104.8; difference 9.9, 95% CI 6.4 to 13.4). They rated themselves with more behaviour problems than did sibling controls (52.7 versus 44.7; difference -7.6, 95% CI 11.2 to -4.0) on the Achenbach Self Report. A high thyroxine starting dose, high serum thyroxine treatment levels during the first six childhood years, and high levels at assessment had no adverse effects on outcome measures at age 20. On the contrary, the results suggest better outcome with higher childhood treatment levels. CONCLUSIONS: Long term outcome revealed deficits in some aspects of memory, attention, and behaviour in young adults with CH relative to sibling controls. No adverse effects of high dose thyroxine therapy were found on measures of memory, attention, and behaviour problems. PMID- 15665165 TI - Maternal phenylketonuria: report from the United Kingdom Registry 1978-97. AB - BACKGROUND: The effects of maternal phenylalanine on the fetus include facial dysmorphism, microcephaly, intrauterine growth retardation, developmental delay, and congenital heart disease. AIMS: To evaluate the impact of phenylalanine restricted diet in pregnant women with phenylketonuria (PKU) on their offspring. METHODS: Data on virtually all pregnancies of women with PKU in the United Kingdom between 1978 and 1997 were reported to the United Kingdom PKU Registry. The effect of the use and timing in relation to pregnancy of a phenylalanine restricted diet on birth weight, birth head circumference, the presence or absence of congenital heart disease (CHD), 4 year developmental quotient, and 8 year intelligence quotient were examined. RESULTS: A total of 228 pregnancies resulted in live births (seven twin pregnancies were excluded). In 110 (50%), diet started before conception. For this group mean (SD) birth weight was 3160 (612) g, birth head circumference 33.6 (1.9) cm, 4 year DQ 108.9 (13.2), 8 year IQ 103.4 (15.6), and incidence of CHD was 2.4%. In comparison, for those born where treatment was started during pregnancy (n = 91), birth weight was 2818 (711) g, birth head circumference 32.7 (2.0) cm, 4 year DQ 96.8 (15.0), 8 year IQ 86.5 (13.0), and incidence of CHD was 17%. Month-by-month regression analyses suggested that metabolic control by 12-16 weeks gestation had most influence on outcome. CONCLUSIONS: Many features of the maternal PKU syndrome are preventable by starting a phenylalanine restricted diet. Women with PKU and their carers must be aware of the risks and should start the diet before conception, or as soon after as possible. PMID- 15665166 TI - Risk of Wilms' tumour with multicystic kidney disease: a systematic review. AB - BACKGROUND: Children with multicystic kidney disease (MCKD) are increasingly managed conservatively and are followed up throughout childhood because they are perceived to be at increased risk of developing Wilms' tumour. With this risk still poorly defined and somewhat controversial, the strategy and the duration of follow up do not seem to be based on evidence. METHODS: Systematic review of the literature for all published cohort studies (prospective and retrospective) of children diagnosed to have unilateral MCKD and managed conservatively. EXCLUSION CRITERIA: bilateral MCKD, nephrectomy (not for malignancy) during the follow up period. We estimated for children with MCKD the probability of developing Wilms' tumour during the follow up period, with 95% CI using the Poisson distribution. RESULTS: From 26 reviewed studies, no cases of Wilms' tumour developed in 1041 eligible children. The mean probability of a child with unilateral MCKD to develop Wilms' was therefore nil, with a 97.5% upper CI estimated at 0.0035 (or 3.5 per 1000 children). CONCLUSION: The development of a national or a European registry for children with MCKD would increase the precision of their risk estimate to develop Wilms' tumour. In the meantime, there is no evidence to support any of the different modalities for following up these children by ultrasound, if indeed such a strategy is necessary. PMID- 15665167 TI - Hyperopia and educational attainment in a primary school cohort. AB - BACKGROUND: Vision screening addresses the visual impairments that impact on child development. Tests of long-sightedness are not found in most school screening programmes. The evidence linking mild-moderate hyperopia and lack of progress in school is insufficient, although strengthened by recent findings of developmental problems in infants. AIMS: To report on the relation between hyperopia and education test results in a cohort of primary school children. METHODS: A total of 1298 children, aged 8 years, were screened for hyperopia on the basis of fogging test results. School test results (NFER and SATs) were compared between groups categorised by referral status and refractive error. RESULTS: A total of 166 (12.8%) fogging test failures were referred for ophthalmic assessment. Ophthalmic tests on 105 children provided an accurate diagnosis of vision defects, for reference to their education scores. Fifty per cent of the children examined by optometrists required an intervention (prescription change, glasses prescribed, or referral). Mean (95% CI) NFER scores of children with refractive errors (summed for both eyes) >+3D (98.4, 93.0-103.8, n = 32) or >+1.25D (best eye) (99.3, 93.0-105.6, n = 26) were lower than the respective scores of children with a less positive refractive state (104.8, 100.7 108.9, n = 43) (103.6, 99.7-107.4, n = 49), the non-referred group, and total sample. The SATs results followed a similar trend. A high proportion of the fogging test failures (16%) and confirmed hyperopes (29%) had been referred to an educational psychologist, and the latter group contributed substantially to the poor education scores. CONCLUSIONS: The results of this study provide further evidence for a link between hyperopia and impaired literacy standards in children. PMID- 15665168 TI - School hearing screening programme in the UK: practice and performance. AB - BACKGROUND: Paediatric audiology services and screening programmes are currently under review. AIMS AND METHODS: To investigate current practice and performance of the school hearing screening programme (SHSP) by means of a questionnaire. RESULTS: SHSP was found to detect previously unrecognised hearing loss at low cost. Wide variation in practice was shown, and the majority of services had no computerised system for data collection. CONCLUSION: There is a need for nationally agreed protocols and quality assurance procedures. PMID- 15665169 TI - Changing socioeconomic inequality in infant mortality in Cumbria. AB - AIMS: To investigate infant deaths in Cumbria, 1950-93, in relation to individual and community level socioeconomic status. METHODS: Retrospective birth cohort study of all 283,668 live births and 4889 infant deaths in Cumbria, 1950-93. Community deprivation (Townsend score) and individual social class were used to estimate socioeconomic status. Logistic regression was used to investigate risk of infant death (early neonatal, neonatal, and postneonatal) in relation to social class and Townsend deprivation score, adjusting for year of birth, birth order, multiple births, and stratified by time period, 1950-65, 1966-75, 1976-85, 1986-93. RESULTS: The risk of infant death in all categories was higher in the lower social classes and more deprived communities, although inequality in risk of neonatal death declined after 1975 to such an extent that there was no significant difference in neonatal death rates by socioeconomic status in the most recent time period. By contrast, there was no narrowing in socioeconomic inequality in postneonatal death risk over the study period. Community deprivation was associated with a significant increased risk of postneonatal death after adjusting for individual level socioeconomic status. CONCLUSIONS: Postneonatal deaths remain higher in the most deprived communities and in the more disadvantaged social classes. The social, lifestyle, and environmental determinates of adverse health outcomes for children need to be fully understood, and interventions should be designed and targeted at the more socially deprived sectors of our community. PMID- 15665170 TI - Plasma phenylalanine in patients with phenylketonuria self-managing their diet. PMID- 15665171 TI - First day back at school: an unusual presentation of a foreign body in the mouth. PMID- 15665173 TI - Children, rights, and responsibilities. PMID- 15665172 TI - Portfolios, appraisal, revalidation, and all that: a user's guide for consultants. AB - From April 2005 all doctors in the UK will be expected to be able to demonstrate their fitness to practice as part of the GMC revalidation procedures. The revalidation process is explicitly linked to the consultant appraisal process implemented in 2001. Central to both processes is the development of a folder (portfolio) of supporting evidence. Many consultants have no experience of developing portfolios and are unclear about how to meaningfully do so and what sort of evidence is suitable for revalidation. Furthermore, they are uncertain about whether there is any evidence to support their use for appraisal or assessment. This paper describes what a portfolio is, summarises the evidence for their use in appraisal and assessment, and provides guidance on the collection of evidence for revalidation purposes. In addition, it explores the distinction between appraisal and revalidation. Some evaluation data on perceived benefits and drawbacks by participants in appraisal in a paediatric setting is also included to inform conclusions and thoughts on planning for the future. PMID- 15665175 TI - Children's rights. PMID- 15665176 TI - Challenges and outcomes of working from a rights based perspective. PMID- 15665177 TI - A general paediatrician's practice in children's rights. PMID- 15665178 TI - Are there patterns of bruising in childhood which are diagnostic or suggestive of abuse? A systematic review. AB - AIMS: To investigate what patterns of bruising are diagnostic or suggestive of child abuse by means of a systematic review. METHODS: All language literature search 1951-2004. Included: studies that defined patterns of bruising in non abused or abused children <18 years. Excluded: personal practice, review articles, single case reports, inadequate confirmation of abuse. Two independent full text reviews using standardised data extraction and critical appraisal forms. Studies ranked by study design and definition of abuse used. RESULTS: Twenty three studies included: seven non-abusive bruising, 14 abusive bruising, and two both. Non-abusive: The prevalence, number, and location of bruises is related to increased motor development. Bruising in non-independently mobile babies is very uncommon (<1%). Seventeen per cent of infants who are starting to mobilise, 53% of walkers, and the majority of schoolchildren have bruises. These are small, sustained over bony prominences, and found on the front of the body. Abuse: Bruising is common in children who are abused. Any part of the body is vulnerable. Bruises are away from bony prominences; the commonest site is head and neck (particularly face) followed by the buttocks, trunk, and arms. Bruises are large, commonly multiple, and occur in clusters. They are often associated with other injury types that may be older. Some bruises carry the imprint of the implement used. CONCLUSION: When abuse is suspected, bruising must be assessed in the context of medical, social, and developmental history, the explanation given, and the patterns of non-abusive bruising. Bruises in non-mobile infants, over soft tissue areas, that carry the imprint of an implement and multiple bruises of uniform shape are suggestive of abuse. Quality research across the whole spectrum of children is urgently needed. PMID- 15665179 TI - Can you age bruises accurately in children? A systematic review. AB - AIMS: To investigate whether it is possible to determine the age of a bruise in a child in clinical practice by means of a systematic review. METHODS: An all language literature search up to 2004. Included studies assessed the age of bruises in live children less than 18 years old. Excluded: review articles, expert opinion, and single case reports. Standardised data extraction and critical appraisal forms were used. Two reviewers independently reviewed studies. RESULTS: Of 167 studies reviewed, three were included: two studies described colour assessment in vivo and one from photographs. Although the Bariciak et al study showed a significant association between red/blue/purple colour and recent bruising and yellow/brown and green with older bruising, both this study and Stephenson and Bialas reported that any colour could be present in fresh, intermediate, and old bruises. Results on yellow colouration were conflicting. Stephenson and Bialas showed yellow colour in 10 bruises only after 24 hours, Carpenter after 48 hours, and Bariciak et al noted yellow/green/brown within 48 hours. Stephenson and Bialas reported that red was only seen in those of one week or less. The accuracy with which clinicians correctly aged a bruise to within 24 hours of its occurrence was less than 40%. The accuracy with which they could identify fresh, intermediate, or old bruises was 55-63%. Intra- and inter observer reliability was poor. CONCLUSION: A bruise cannot accurately be aged from clinical assessment in vivo or on a photograph. At this point in time the practice of estimating the age of a bruise from its colour has no scientific basis and should be avoided in child protection proceedings. PMID- 15665180 TI - Pubertal stage and hypoglycaemia counterregulation in type 1 diabetes. AB - AIMS: To compare physiological and autonomic responses to acute hypoglycaemia in diabetic children in pre-, mid-, and post-pubertal stages of development. METHODS: Twenty seven children (8 pre-pubertal, 7 mid-pubertal, 12 post-pubertal) with type 1 diabetes were studied. Hypoglycaemia was induced by insulin infusion until an autonomic reaction (R) was identified. Counterregulatory hormone levels were measured at baseline, R, R+15, and R+30 minutes. Haemodynamic changes and sweat production were measured. RESULTS: The mean blood glucose level at R was lower in pre-pubertal than mid-pubertal children (2.0 v 2.5 mmol/l), and was positively correlated with HbA1c. Glucagon and noradrenaline responses to hypoglycaemia were minimal in all children. A brisk increase in pancreatic polypeptide (PP) concentration only occurred in post-pubertal children. Only two children showed a sweating response to hypoglycaemia. CONCLUSIONS: The blood glucose level at which sympatho-adrenal responses to hypoglycaemia were activated was associated with glycaemic control, and varied with pubertal stage. As in adults, the glucagon response to hypoglycaemia was deficient within a few years of developing diabetes. However, sweating and secretion of PP in response to hypoglycaemia did not occur until after puberty, indicating some qualitative differences from adults. PMID- 15665181 TI - Partially hydrolysed guar gum supplemented comminuted chicken diet in persistent diarrhoea: a randomised controlled trial. AB - BACKGROUND: Partially hydrolysed guar gum (Benefiber) added to a diet is fermented in the colon, producing short chain fatty acids, which improve intestinal function, including colonic salt and water absorption. AIMS: To evaluate the effect of Benefiber supplemented comminuted chicken diet in the treatment of persistent diarrhoea. METHODS: One hundred and sixteen children (aged 5-24 months), presenting to Dhaka Hospital with a history of watery diarrhoea for more than 14 days (persistent diarrhoea), were randomised to receive either: (1) comminuted chicken diet with Benefiber (study diet); or (2) comminuted chicken diet without Benefiber (control diet). The study period was seven days. RESULTS: Of 116 children, 57 received the study diet and 59 received the control diet. Diarrhoea resolved in a greater number of children with the study than with the control diet (46/55 (84%) v 36/58 (62%); odds ratio 3.12, 95% CI 1.19 to 8.4). Survival analysis for the duration of diarrhoea also showed a reduced duration of diarrhoea in children receiving the study diet. There was also a trend in daily stool reduction in children receiving the study diet, significant on days 4-7. CONCLUSION: Results show that Benefiber supplemented comminuted chicken diet enhances recovery of children with persistent diarrhoea, indicating its therapeutic potential. PMID- 15665182 TI - The management of sick young infants at primary health centres in a rural developing country. AB - AIMS: To investigate the epidemiology of illness among young infants at remote health clinics in a rural developing country, and to determine risk factors for mortality that might be used as triggers for emergency treatment or referral. METHODS: Multi-site 12 month observational study of consecutive presentations of infants less than 2 months, and an investigation of neonates who died in one district without accessing health care. RESULTS: Forty per cent of 511 young infant presentations occurred in the first week of life and most of these in the first 24 hours. Twenty five deaths were recorded: 18 in the health facilities and seven in villages. In addition there were eight stillbirths. Clinical signs predicting death were: not able to feed, fast respiratory rate, apnoea, cyanosis, "too small", "skin-cold", and severe abdominal distension. Signs indicating severe respiratory compromise were present in 25% of young infants; failure to give oxygen therapy was a modifiable factor in 27% of deaths within health facilities. A high proportion of seriously ill young infants were discharged from health facilities early without adequate follow up. A common reason for not seeking care for fatally ill neonates was the perception by parents that health staff would respond negatively to their social circumstances. CONCLUSIONS: Clinical signs with moderate positive predictive value for death may be useful triggers for emergency treatment and longer observation or urgent referral. The results of this study may be useful in planning strategies to address high neonatal mortality rates in developing countries. PMID- 15665183 TI - Visual failure without headache in idiopathic intracranial hypertension. AB - AIM: To determine the differences in outcome in a group of children with idiopathic intracranial hypertension (IIH) who do not present with headaches. METHODS: Differences in epidemiological and aetiological factors, clinical presentation, and visual outcome were investigated in children with a diagnosis of IIH presenting with and without headaches to the Paediatric Neurology and Paediatric Ophthalmology Services at Guy's & St Thomas' Hospitals NHS Trust between 1997 and 2002. RESULTS: Compared to the 29 children with headaches, the 12 children in the non-headache group were younger (7.3 v 9.5 years), presented with more neurological signs (33% v 10%), and were more likely to present with severe visual failure (33% v 4%), with a tenfold increased risk of an enlarged blind spot or field defects (50% v 5%). Permanent visual failure affected a third of all children in the non-headache group, but was rare in children presenting with headaches (33% v 3%), with one patient registered blind and two severely visually impaired. CONCLUSION: The management of IIH is difficult in the absence of headache. Visual surveillance is vital. These children were treated with an aggressive management programme to reduce cerebrospinal fluid pressure by repeated lumbar puncture, medication, and early surgical intervention if required. Non-invasive monitoring techniques might contribute to a better understanding of the natural history of IIH, improved management, and visual outcome. PMID- 15665184 TI - Reliability of PRISM and PIM scores in paediatric intensive care. AB - AIMS: To assess the reliability of mortality risk assessment using the Paediatric Risk of Mortality (PRISM) score and the Paediatric Index of Mortality (PIM) in daily practice. METHODS: Twenty seven physicians from eight tertiary paediatric intensive care units (PICUs) were asked to assess the severity of illness of 10 representative patients using the PRISM and PIM scores. Physicians were divided into three levels of experience: intensivists (>3 years PICU experience, n = 12), PICU fellows (6-30 months of PICU experience, n = 6), and residents (<6 months PICU experience, n = 9). This represents all large PICUs and about half of the paediatric intensivists and PICU fellows working in the Netherlands. RESULTS: Individual scores and predicted mortality risks for each patient varied widely. For PRISM scores the average intraclass correlation (ICC) was 0.51 (range 0.32 0.78), and the average kappa score 0.6 (range 0.28-0.87). For PIM scores the average ICC was 0.18 (range 0.08-0.46) and the average kappa score 0.53 (range 0.32-0.88). This variability occurred in both experienced and inexperienced physicians. The percentage of exact agreement ranged from 30% to 82% for PRISM scores and from 28 to 84% for PIM scores. CONCLUSION: In daily practice severity of illness scoring using the PRISM and PIM risk adjustment systems is associated with wide variability. These differences could not be explained by the physician's level of experience. Reliable assessment of PRISM and PIM scores requires rigorous specific training and strict adherence to guidelines. Consequently, assessment should probably be performed by a limited number of well trained professionals. PMID- 15665185 TI - My daughter. PMID- 15665186 TI - Expert witnesses: opinion and dogma are pitfalls in medical journalism as well as in reports. PMID- 15665187 TI - Induction of human sperm capacitation and protein tyrosine phosphorylation by endometrial cells and interleukin-6. AB - In order to become fully competent at fertilizing the oocyte, spermatozoa must undergo the maturational process of capacitation during their journey in the female reproductive tract. Endometrial cells secrete an array of growth factors that can affect spermatozoa. Among these factors, it has been previously demonstrated that interleukin-6 (IL-6) affects the fertilizing capacity of human spermatozoa. As the expression of this cytokine varies throughout the menstrual cycle and increases during the periovulatory period, the involvement of IL-6 in human sperm capacitation was investigated, with emphasis on the signal transduction cascade triggered by this agent in sperm cells. Spermatozoa were treated with recombinant human IL-6. Protein phosphotyrosine content and localization of the phosphotyrosine containing proteins were evaluated by western blot and indirect immunofluorescence, respectively, using a monoclonal anti phosphotyrosine antibody. The acrosomal status was evaluated on IL-6 treated spermatozoa before or after challenge with the ionophore A23187 according to the fluorescent pattern observed upon binding to the Pisum sativum agglutinin conjugated to fluorescein isothiocyanate. In the present study, it is shown that, as for endometrial cell-conditioned media, IL-6 induces human sperm capacitation. The IL-6 effects most likely occur through binding to its receptor, IL-6Ralpha, whose presence in the sperm is also reported in this study. As for the IL-6 receptor, this is the first report on the presence of the tyrosine kinase JAK1 in the spermatozoa. Moreover, this kinase becomes phosphorylated on tyrosine residues upon sperm treatment with recombinant IL-6, which suggests its activation by the cytokine. Taken together, our results demonstrate that the IL-6 intracellular signalling machinery is present in human spermatozoa and might be involved in the acquisition of sperm fertilizing ability, also known as the capacitation process. PMID- 15665188 TI - Prevalence of headache in Australian footballers. AB - OBJECTIVE: To survey the prevalence and risk factors for headache in a population of elite professional Australian footballers. METHODS: A prospective questionnaire based survey was performed on elite Australian footballers participating in a national competition. The survey was designed to assess the prevalence and risk factors for headache using standardised International Headache Society (HIS) criteria. Headache prevalence was compared with that of an age and sex matched community control population. RESULTS: A total of 160 questionnaires were analysed. Headaches were reported by 80% of subjects, with 49% of respondents reporting headaches during competitive match play and 60% during training. There was no significant difference between the footballers and the community sample in the number of headaches ever; however, they did report more headaches in the three months before the survey--that is, during the competitive playing season. In the footballers, 22% of headaches conformed to the strict IHS definition of migraine headaches. When the relaxed definition of "footballer's migraine" was used, 34% of headaches met these criteria. Footballers were at significantly increased risk of footballer's migraine than community controls. CONCLUSIONS: Headaches are common in Australian footballers, and the prevalence of migraine is increased in relation to community prevalence studies. Furthermore, the strict IHS criteria may not adequately identify the specific subtype of football related headache. This finding has important implications in the management of headache in this setting. PMID- 15665189 TI - Effects of training period on haemorheological variables in regularly trained footballers. AB - OBJECTIVE: To investigate the effects of one football training period on haemorheological variables in regularly trained footballers. METHOD: Ten subjects were randomly selected from the reserve team of a football club in the Turkish Premier League. During the last week of the football season, one day before a standard training session and two days after the previous league match, venous blood samples were taken (pre-exercise). After 90 minutes of standard training, further blood samples were taken (post-exercise). Blood lactate, blood viscosity, plasma fibrinogen, blood clotting time, acid-base variables, and plasma Na+, K+, and Ca2+ were determined. RESULTS: Haemoglobin, packed cell volume, and mean corpuscular volume were all significantly decreased, whereas white blood cells and platelets were both increased after training. Blood viscosity decreased but the reduction was not significant. Blood lactate, plasma glucose, and Na+ content were significantly increased, but standard bicarbonate, actual bicarbonate, and Ca2+ were significantly decreased. Blood clotting time had shortened significantly after training. Blood viscosity was inversely correlated with plasma glucose concentration (r = -0.48 and p = 0.032). CONCLUSIONS: The results show that blood viscosity tends to decrease as the result of this type of training. This is due to a reduction in packed cell volume and mean corpuscular volume. The increased blood lactate does not have an adverse effect on the blood viscosity of these subjects because protective mechanisms develop with regular training throughout the season. PMID- 15665190 TI - Differences in sole arch indices in various sports. AB - BACKGROUND: There are controversial data about the relation between foot morphology and athletic injuries of the lower extremity. Studies in soldiers have shown some relationship, whereas those involving athletes have not shown any significant relationship. The reason for these differences is not clear. OBJECTIVE: To determine the effect of various sports on sole arch indices (AIs). METHOD: A total of 116 elite male athletes (24 soccer players, 23 wrestlers, 19 weightlifters, 30 handball players, and 20 gymnasts) and 30 non-athletic men were included in this cross sectional study. Images of both soles were taken in a podoscope and transferred to a computer using a digital still camera. AIs were calculated from the stored images. RESULTS: The AI of the right sole of the gymnasts was significantly lower than that of the soccer players, wrestlers, and non-athletic controls (p<0.01). The AI of the right sole of the wrestlers was significantly higher than that of the soccer players, handball players, weightlifters, gymnasts, and non-athletic controls (p<0.03). The AI of the left sole of the gymnasts was significantly lower than that of the wrestlers and non athletic controls (p<0.001). The AI of the left sole of the wrestlers was significantly higher than that of the soccer players, handball players, and gymnasts (p<0.007). The AI of both soles in handball players was significantly lower than those of the non-athletic subjects (p = 0.049). The correlation between the AI of the left and right foot was poor in the soccer players, handball players, and wrestlers (r = 0.31, 0.69, and 0.56 respectively), but was high in the gymnasts, weightlifters, and non-athletic controls (r = 0.96, 0.88, and 0.80 respectively). CONCLUSION: The AIs of the gymnasts and wrestlers were significantly different from those of other sportsmen studied, and those of the gymnasts and handball players were significantly different from those of non athletic controls. PMID- 15665191 TI - Propagation of stress fracture of the patella. AB - Anterior knee pain in athletes is common and usually self limited. There should be a high index of suspicion and low threshold for special imaging in cases with acute onset and specific tenderness. The risk of propagation of stress fracture of the patella in athletes is highlighted. The case report presented illustrates the potential sequence of events. PMID- 15665192 TI - Kleine-Levin syndrome: a unique cause of fatigue in an athlete. AB - Kleine-Levin syndrome (KLS) is a rare disorder characterised, most notably, by periodic episodes of hypersomnolence and hyperphagia. Associated features of the disorder include a lack of concentration, mood changes, and anxiety. Laboratory tests may show slight changes in the electroencephalogram. However, clinical presentation and laboratory tests are normal during asymptomatic intervals. KLS most often presents in adolescent males, with complete recovery by the 3rd to 4th decade of life. Possible precipitating factors include excessive workload, febrile illness, and respiratory infections. Presented is a classical case of KLS in an adolescent male athlete. The patient's history, complete laboratory results, and symptoms are discussed. Possible treatments for this disorder are also mentioned, along with diagnostic criteria. PMID- 15665193 TI - False aneurysm of the common femoral vein in a footballer. AB - Traumatic false aneurysm of the femoral vein has never been reported in the English literature. The case is here reported of a footballer with a traumatic false aneurysm of the common femoral vein which was initially misdiagnosed as an arterial pseudoaneurysm. This is a very rare clinical condition, but this diagnosis should be among those considered for post-traumatic unexplained thigh pain after trauma. PMID- 15665194 TI - Quadrilateral space syndrome: a case study and review of the literature. AB - Quadrilateral space syndrome is an uncommon injury. The true prevalence is unknown because of a lack of literature and possible misdiagnosis. Prevalence may increase as knowledge of the syndrome increases. The case is presented of a recreational triathlete who had a spontaneous onset of quadrilateral space syndrome. The diagnosis was made by physical examination and confirmed with magnetic resonance imaging. A conservative, yet aggressive rehabilitation programme resulted in functional improvement within six weeks. Results have been maintained for eight weeks. PMID- 15665195 TI - Aqua vita. PMID- 15665196 TI - Investigate, study, explore, examine, enquire, seek. PMID- 15665197 TI - Do non-steroidal anti-inflammatory drugs adversely affect stress fracture healing? A short review. AB - A literature search was performed to determine whether non-steroidal anti inflammatory drugs (NSAIDs) adversely affect the healing of stress fractures. Evidence exists from laboratory studies and animal subjects that NSAIDs can affect fracture healing. This link has not been proved or disproved in human subjects, particularly for stress fractures. In view of the high usage of NSAIDs in treating musculoskeletal disorders, research is required to investigate whether the healing of stress fractures is affected by these drugs. PMID- 15665201 TI - Cost effectiveness of adding magnetic resonance imaging to the usual management of suspected scaphoid fractures. AB - OBJECTIVE: To determine the cost effectiveness of a magnetic resonance imaging scan (MRI) within 5 days of injury compared with the usual management of occult scaphoid fracture. METHODS: All patients with suspected scaphoid fractures in five hospitals were invited to participate in a randomised controlled trial of usual treatment with or without an MRI scan. Healthcare costs were compared, and a cost effectiveness analysis of the use of MRI in this scenario was performed. RESULTS: Twenty eight of the 37 patients identified were randomised: 17 in the control group, 11 in the MRI group. The groups were similar at baseline and follow up in terms of number of scaphoid fractures, other injuries, pain, and function. Of the patients without fracture, the MRI group had significantly fewer days immobilised: a median of 3.0 (interquartile range 3.0-3.0) v 10.0 (7-12) in the control group (p = 0.006). The MRI group used fewer healthcare units (median 3.0, interquartile range 2.0-4.25) than the control group (5.0, 3.0-6.5) (p = 0.03 for the difference). However, the median cost of health care in the MRI group (594.35 dollars AUD, 551.35-667.23 dollars) was slightly higher than in the control group (428.15 dollars, 124.40-702.65 dollars) (p = 0.19 for the difference). The mean incremental cost effectiveness ratio derived from this simulation was that MRI costs 44.37 dollars per day saved from unnecessary immobilisation (95% confidence interval 4.29 dollars to 101.02 dollars). An illustrative willingness to pay was calculated using a combination of the trials measure of the subjects' individual productivity losses and the average daily earnings. CONCLUSIONS: Use of MRI in the management of occult scaphoid fracture reduces the number of days of unnecessary immobilisation and use of healthcare units. Healthcare costs increased non-significantly in relation to the use of MRI in this setting. However, when productivity losses are considered, MRI may be considered cost effective, depending on the individual case. PMID- 15665199 TI - Ocular complications of boxing. AB - OBJECTIVES: To investigate the prevalence of ocular injuries in a large population of boxers over a period of 16 years, in particular, the most severe lesions that may be vision threatening. METHODS: Clinical records of the medical archive of the Italian Boxing Federation were analysed. A total of 1032 boxers were examined from February 1982 to October 1998. A complete ophthalmological history was available for 956, who formed the study population (a total of 10 697 examinations). The following data were collected: age when started boxing; duration of competitive boxing career (from the date of the first bout); weight category; a thorough ocular history. The following investigations were carried out: measurement of visual acuity and visual fields, anterior segment inspection, applanation tonometry, gonioscopy, and examination of ocular fundus. Eighty age matched healthy subjects, who had never boxed, formed the control group. RESULTS: Of the 956 boxers examined, 428 were amateur (44.8%) and 528 professional (55.2%). The median age at first examination was 23.1 (4.3) years (range 15-36). The prevalence of conjunctival, corneal, lenticular, vitreal, ocular papilla, and retinal alterations in the study population was 40.9% compared with 3.1% in the control group (p< or =0.0001). The prevalence of serious ocular findings (angle, lens, macula, and peripheral retina alterations) was 5.6% in boxers and 3.1% in controls (NS). CONCLUSIONS: Boxing does not result in a higher prevalence of severe ocular lesions than in the general population. However, the prevalence of milder lesions (in particular with regard to the conjunctiva and cornea) is noteworthy, justifying the need for adequate ophthalmological surveillance. PMID- 15665202 TI - A nomogram for assessment of breathing patterns during treadmill exercise. AB - OBJECTIVE: To assess the breathing patterns of trained athletes under different conditions. The hypothesis is that the breathing pattern during a progressive treadmill exercise is independent of the protocol, at least in healthy people, and can be assessed using a nomogram. METHODS: A total of 43 male and 21 female athletes from different sports were studied. They performed one of two different protocols (steps or ramp) on a treadmill. The two protocols started at the same speed and had the same rate of increase in work. During the test, the expired air was analysed for CO2 and O2. Ventilation (VE) was continuously recorded, and tidal volume (Vt) and breathing frequency (BF) at the same intensity were analysed for both protocols, as well as Vt/T(i) and T(i)/T(tot). RESULTS: No significant differences were observed in Vt and BF between the two protocols in either the men or women at any level (confidence intervals up to 0.958 in all the groups). T(i)/T(tot) remained constant, and all increases in VE were strongly related to the respective increases in Vt/T(i). Plots of data for men and women showed a curvilinear relation between Vt and BF which could be fitted with an exponential function with a strong correlation (R2 = 0.98 for men and 0.97 for women). CONCLUSIONS: Graphic expression of Vt v BF is a useful nomogram for the routine assessment of ventilatory response during exercise in healthy trained subjects. PMID- 15665204 TI - Management of ankle sprains: a randomised controlled trial of the treatment of inversion injuries using an elastic support bandage or an Aircast ankle brace. AB - BACKGROUND: Lateral ligament ankle sprains are the single most common sports injury. OBJECTIVE: To determine the functional outcome of the ankle joint after a moderate or severe inversion injury, comparing standard treatment with an elastic support bandage against an Aircast ankle brace. DESIGN: Prospective, randomised controlled trial. SETTING: Two accident and emergency departments. METHOD: Fifty patients presenting consecutively were randomised into two equal groups: one group was treated with an elastic support bandage and the other with an Aircast ankle brace. All patients were given a standardised advice sheet referring to rest, ice, compression, and elevation. Patients were reviewed after 48-72 hours, 10 days, and one month. PRIMARY OUTCOME MEASURE: Ankle joint function assessed at 10 days and one month using the modified Karlsson scoring method (maximum score 90). SECONDARY OUTCOME MEASURE: The difference in ankle girth (swelling) and pain score at 10 days. RESULTS: Seventeen patients in the elastic support bandage group (six defaulted, two excluded) and 18 patients in the Aircast ankle brace group (six defaulted, one excluded) completed the study. There were no significant differences between the two groups at presentation in terms of age (mean 35.3 and 32.6 years respectively), sex, dominant leg, left or right ankle injured, previous injury, time to presentation (median three and four hours respectively), difference in ankle girth (mean 14.5 and 14.3 mm respectively), and pain scores (mean 6.2 and 5.8 respectively). The Karlsson score was significantly higher in the Aircast ankle cast group than in the elastic bandage group at 10 days (mean 50 v 35, p = 0.028, 95% confidence interval (CI) 1.7 to 27.7) and one month (mean 68 v 55, p = 0.029, 95% CI 1.4 to 24.8) (Student's t test). There was no difference between the groups in the secondary outcome measures (swelling, p = 0.09; pain, p = 0.07). When hierarchical multiple regression analysis was used to correct for possible baseline confounding factors, the Aircast ankle brace group was significantly associated with higher Karlsson scores at 10 days (p = 0.009) and one month (p = 0.024). CONCLUSION: The use of an Aircast ankle brace for the treatment of lateral ligament ankle sprains produces a significant improvement in ankle joint function at both 10 days and one month compared with standard management with an elastic support bandage. PMID- 15665203 TI - Effect of dry needling of gluteal muscles on straight leg raise: a randomised, placebo controlled, double blind trial. AB - OBJECTIVES: To use a randomised, double blind, placebo controlled trial to establish the effect on straight leg raise, hip internal rotation, and muscle pain of dry needling treatment to the gluteal muscles in athletes with posterior thigh pain referred from gluteal trigger points. METHODS: A randomised, double blind, placebo controlled trial of 59 male runners was performed during the 2002 Australian Rules football season. Subjects were thoroughly screened and had magnetic resonance imaging of their hamstring muscles to exclude local pathology. The inclusion criterion was reproduction of recognisable posterior thigh pain with the application of digital pressure to the gluteal trigger points. Subjects randomly received either therapeutic or placebo needle treatment on one occasion at their gluteal trigger points. Range of motion and visual analogue scale data were collected immediately before, immediately after, 24 hours after, and 72 hours after the intervention. Range of motion was measured with passive straight leg raise and hip internal rotation. Visual analogue scales were completed for hamstring and gluteal pain and tightness at rest and during a running task. RESULTS: Magnetic resonance imaging scans revealed normal hamstring musculature in most subjects. Straight leg raise and hip internal rotation remained unchanged in both groups at all times. Visual analogue scale assessment of hamstring pain and tightness and gluteal tightness after running showed improvements immediately after the intervention in both groups (p = 0.001), which were maintained at 24 and 72 hours. The magnitude of this improvement was the same for therapeutic and placebo interventions. Resting muscle pain and tightness were unaffected. CONCLUSIONS: Neither dry needling nor placebo needling of the gluteal muscles resulted in any change in straight leg raise or hip internal rotation. Both interventions resulted in subjective improvement in activity related muscle pain and tightness. Despite being commonly used clinical tests in this situation, straight leg raise and hip internal rotation are not likely to help the therapist assess response to treatment. Patient reports of response to such treatment are better indicators of its success. The mechanisms by which these responses occur and the reasons for the success of the placebo needling treatment are areas for further investigation. PMID- 15665205 TI - Appropriate interpretation of aerobic capacity: allometric scaling in adult and young soccer players. AB - OBJECTIVE: To compare aerobic capacity of young and adult elite soccer players using appropriate scaling procedures. METHODS: Twenty four male adult (mean (SD) age 24 (2) years, weight 75.7 (7.2) kg, VO2max 66.6 (5.2) ml/lbm/min, where lbm is lean body mass in kg) and 21 youth (14 (0.4) years, 60.2 (7.3) kg, 66.5 (5.9) ml/lbm/min) elite soccer players took part in the study. Allometric equations were used to determine the relation between maximal and submaximal oxygen cost of running (running economy) and body mass. RESULTS: Maximal and submaximal oxygen uptake increased in proportion to body mass raised to the power of 0.72 (0.04) and 0.60 (0.06) respectively. The VO2max of adult players was similar to that of the youth players when expressed in direct proportion to body mass--that is, ml/kg/min--but 5% higher (p<0.05) when expressed using appropriate procedures for scaling. Conversely, compared with seniors, youth players had 13% higher (p<0.001) energy cost of running--that is, poorer running economy--when expressed as ml/kg/min but not when expressed according to the scaling procedures. CONCLUSIONS: Compared with the youth soccer players, VO2max in the seniors was underestimated and running economy overestimated when expressed traditionally as ml/lbm/min. The study clearly shows the pitfalls in previous studies when aerobic capacity was evaluated in subjects with different body mass. It further shows that the use of scaling procedures can affect the evaluation of, and the resultant training programme to improve, aerobic capacity. PMID- 15665207 TI - Eccentric decline squat protocol offers superior results at 12 months compared with traditional eccentric protocol for patellar tendinopathy in volleyball players. AB - BACKGROUND: Conservative treatment of patellar tendinopathy has been minimally investigated. Effective validated treatment protocols are required. OBJECTIVES: To investigate the immediate (12 weeks) and long term (12 months) efficacy of two eccentric exercise programmes for the treatment of patellar tendinopathy. METHODS: This was a prospective randomised controlled trial of 17 elite volleyball players with clinically diagnosed and imaging confirmed patellar tendinopathy. Participants were randomly assigned to one of two treatment groups: a decline group and a step group. The decline group were required to perform single leg squats on a 25 degrees decline board, exercising into tendon pain and progressing their exercises with load. The step group performed single leg squats on a 10 cm step, exercising without tendon pain and progressing their exercises with speed then load. All participants completed a 12 week intervention programme during their preseason. Outcome measures used were the Victorian Institute of Sport Assessment (VISA) score for knee function and 100 mm visual analogue scale (VAS) for tendon pain with activity. Measures were taken throughout the intervention period and at 12 months. RESULTS: Both groups had improved significantly from baseline at 12 weeks and 12 months. Analysis of the likelihood of a 20 point improvement in VISA score at 12 months revealed a greater likelihood of clinical improvements in the decline group than the step group. VAS scores at 12 months did not differ between the groups. CONCLUSIONS: Both exercise protocols improved pain and sporting function in volleyball players over 12 months. This study indicates that the decline squat protocol offers greater clinical gains during a rehabilitation programme for patellar tendinopathy in athletes who continue to train and play with pain. PMID- 15665208 TI - Risk factors for hamstring injuries in community level Australian football. AB - OBJECTIVES: To identify risk factors for hamstring injury at the community level of Australian football. METHODS: A total of 126 community level Australian football players participated in this prospective cohort study. To provide baseline measurements, they completed a questionnaire and had a musculoskeletal screen during the 2000 preseason. All were monitored over the season. Injury surveillance and exposure data were collected for the full season. Survival analysis was used to identify independent predictors of hamstring injury. RESULTS: A hamstring injury was the first injury of the season in 20 players (16%). After adjustment for exposure, increasing age and decreased quadriceps flexibility were identified as significant independent predictors of the time to sustaining a hamstring injury. Older age (> or =23 years) was associated with an increased risk of hamstring injury (RR 3.8; 95% confidence interval (CI) 1.1 to 14.0; p = 0.044). Players with increased quadriceps flexibility (as measured by the modified Thomas test) were less likely to sustain a hamstring injury (RR 0.3; 95% CI 0.1 to 0.8; p = 0.022). CONCLUSIONS: The findings of this study can be used in the development of hamstring injury prevention strategies and to identify Australian football players at increased risk of hamstring injury. PMID- 15665211 TI - A simple method for removal of fish hooks in the emergency department. PMID- 15665210 TI - An economic evaluation of a proprioceptive balance board training programme for the prevention of ankle sprains in volleyball. AB - OBJECTIVES: To evaluate the cost effectiveness of a proprioceptive balance board training programme for the prevention of ankle sprains in volleyball. METHODS: A total of 116 volleyball teams participated in this study which was carried out during the 2001-2002 volleyball season. Teams were randomly allotted to an intervention group (66 teams, 628 players) or a control group (52 teams, 494 players). Intervention teams followed a prescribed balance board training programme as part of their warm up. Control teams followed their normal training routine. An ankle sprain was recorded if it occurred as a result of volleyball and caused the subject to stop volleyball activity. The injured player completed a cost diary for the duration of the ankle sprain. Analyses were performed according to the intention to treat principle. Mean direct, indirect, and total costs were calculated and were compared between the two groups. RESULTS: The total costs per player (including the intervention material) were significantly higher in the intervention group (36.99 (93.87)) than in the control group (18.94 (147.09)). The cost of preventing one ankle sprain was approximately 444.03. Sensitivity analysis showed that a proprioceptive balance board training programme aimed only at players with previous ankle sprains could be cost effective over a longer period of time. CONCLUSIONS: Positive effects of the balance board programme could only be achieved at certain costs. However, if broadly implemented, costs associated with the balance board programme would probably be lower. PMID- 15665212 TI - Paediatric sport related concussion pilot study. PMID- 15665214 TI - Brain. Editorial. PMID- 15665213 TI - From catastrophe to complexity: a novel model of integrative central neural regulation of effort and fatigue during exercise in humans: summary and conclusions. AB - It is hypothesised that physical activity is controlled by a central governor in the brain and that the human body functions as a complex system during exercise. Using feed forward control in response to afferent feedback from different physiological systems, the extent of skeletal muscle recruitment is controlled as part of a continuously altering pacing strategy, with the sensation of fatigue being the conscious interpretation of these homoeostatic, central governor control mechanisms. PMID- 15665215 TI - Facial reflexes. PMID- 15665216 TI - 'A beautiful creature that jumps'. PMID- 15665218 TI - Atrial fibrillation: multi-detector row CT of pulmonary vein anatomy prior to radiofrequency catheter ablation--initial experience. AB - PURPOSE: To evaluate multi-detector row computed tomographic (CT) depiction of pulmonary veins to provide a road map for radiofrequency catheter ablation. MATERIALS AND METHODS: For patients, institutional review board (IRB) approval was not required, and consent was obtained for treatment. Control subjects were part of an IRB-approved research protocol at the institution, in which they had consented to participate. Multi-detector row CT was performed in 23 patients (17 men, six women; mean age, 48 years +/- 11 [standard deviation]) with atrial fibrillation who were admitted for isolation of pulmonary veins by means of radiofrequency catheter ablation. Pulmonary vein anatomy was evaluated, and diameters of pulmonary vein ostia were measured. To determine the shape of ostia, a venous ostium index was calculated for all veins by dividing anterior-posterior measurements by superior-inferior measurements. Results were compared with those in a control group of 11 patients (eight men, three women; mean age, 56 years +/- 11) without atrial fibrillation. Images were evaluated by two observers in consensus. RESULTS: Pulmonary veins additional to the four main veins were found in seven (30%) of 23 patients. Common ostia of left and right pulmonary veins were detected in 19 (83%) and nine (39%) patients, respectively. Early branching occurred more often with right than with left veins (19 [83%] vs three [13%] cases, P <.05) in both patients and control subjects. Anterior-posterior diameter of ostia was 12.8 mm +/- 3.3 for left veins, 16.2 mm +/- 3.8 for right veins, and 18.8 mm +/- 7.7 and 28.7 mm +/- 5.1 for left and right common ostia, respectively. Ostia of right pulmonary veins were more round than were ostia of left pulmonary veins (venous ostium index in patients, 0.91 +/- 0.21 vs 0.75 +/- 0.17, P <.05; in control subjects, 0.93 +/- 0.12 vs 0.82 +/- 0.17, P <.05). The CT data were used to determine ablation strategy and guide catheters during radiofrequency ablation. CONCLUSION: Multi-detector row CT provides a valuable road map for pulmonary vein anatomy prior to radiofrequency catheter ablation. Variations in number and insertion of pulmonary veins were observed in a considerable number of patients and control subjects. PMID- 15665219 TI - Imaging of macrophages in soft-tissue infection in rats: relationship between ultrasmall superparamagnetic iron oxide dose and MR signal characteristics. AB - PURPOSE: To describe dose-dependent signal intensity (SI) characteristics of experimentally induced soft-tissue abscesses on 1.5-T T1- and T2*-weighted magnetic resonance (MR) images obtained 24 hours after administration of ultrasmall superparamagnetic iron oxide (USPIO) and to describe the relationship between SI and amount of USPIO uptake and macrophage iron content. MATERIALS AND METHODS: Local institutional review committee on animal care approved the experiments, which were performed according to the guidelines of the National Institutes of Health and the committee on animal research at our institution. Unilateral calf muscle abscesses were induced in 21 rats with an injection of a Staphylococcus aureus suspension. The rats were divided into three groups of seven animals each: low USPIO dose (50 micromol of iron per kilogram of body weight), high USPIO dose (150 micromol Fe/kg), and control (saline solution). All rats were imaged before and 24 hours after USPIO administration at 1.5 T (transverse T1-weighted spin-echo, T2*-weighted fast gradient-echo, and short inversion time inversion-recovery sequences). Images were analyzed quantitatively and qualitatively with regard to SI and signal pattern. Temporal variation of calculated contrast-to-noise ratios was analyzed with the Wilcoxon signed rank test. MR findings were correlated with histopathologic findings, including those of electron microscopy. RESULTS: Twenty-four hours after USPIO administration in the high-dose group, susceptibility effects were present in abscess periphery on postcontrast T2*-weighted images (P=.04), and SI enhancement was noted on postcontrast T1-weighted images within both abscess wall and abscess center (P=.04 for both). In the low-dose group, SI enhancement was noted in entire abscess on T1-weighted postcontrast images (P=.03). Neither significant SI loss (P=.09) nor susceptibility effects were detected in periphery or center of any abscess on postcontrast T2*-weighted images. There was no obvious difference in total amount of macrophages among the groups, but there was a clear difference with regard to individual iron content of iron-positive macrophages between the USPIO dose groups. CONCLUSION: At 1.5 T, SI characteristics of abscesses on T1- and T2*-weighted images obtained 24 hours after USPIO injection strongly depend on administered dose of the contrast agent. At low doses, T1 effects were stronger than T2* effects. PMID- 15665220 TI - Posterior cul-de-sac obliteration associated with endometriosis: MR imaging evaluation. AB - PURPOSE: To retrospectively evaluate the accuracy of magnetic resonance (MR) imaging in depicting posterior cul-de-sac obliteration in patients with endometriosis. MATERIALS AND METHODS: Institutional review board approval was not required for this retrospective study, but informed consent was obtained from all patients. MR images obtained between January 1989 and December 2000 in 57 women (mean age, 39 years; age range, 26-52 years) with histologically confirmed endometriosis were retrospectively evaluated by four radiologists independently. All patients underwent laparotomy or laparoscopy less than 1 month after MR imaging. MR images were evaluated for the presence and location of endometrial implants and adhesions. MR images were also scored for the presence of five findings: retroflexed uterus, elevated posterior vaginal fornix, intestinal tethering or tethered appearance of rectum in direction of uterus, faint strands between uterus and intestine, and fibrotic plaque or nodule covering serosal surface of the uterus. Interobserver agreement for each of the five findings and for the overall diagnosis of cul-de-sac obliteration was calculated. Sensitivity, specificity, accuracy, positive and negative predictive values, and kappa statistics were determined. RESULTS: Laparotomy or laparoscopy revealed posterior cul-de-sac obliteration in 30 patients. Overall, the four radiologists had mean accuracies of 89.0% and 76.3% for diagnosing endometrial implants and adhesions, respectively, at MR imaging. Overall, the radiologists achieved mean sensitivity, specificity, accuracy, and positive and negative predictive values of 68.4%, 76.0%, 71.9%, 76.6%, and 68.5%, respectively, in diagnosing posterior cul-de-sac obliteration. The best accuracy (mean value, 64.5%) was obtained with the finding of fibrotic plaque in the uterine serosal surface. Readers agreed on the observations 63.2%-91.2% of the time. For the impression of the presence or absence of posterior cul-de-sac obliteration, interobserver agreement varied between substantial and moderate: Mean interobserver agreement was 78.4% (range, 70.2%-84.2%), and mean kappa was 0.57 (range, 0.40-0.67). Mean accuracy of MR imaging for diagnosing posterior cul-de-sac obliteration was 71.9%. CONCLUSION: These results suggest that use of the described MR imaging findings may enable diagnosis of posterior cul-de-sac obliteration. PMID- 15665221 TI - Coronary MR angiography at 3.0 T versus that at 1.5 T: initial results in patients suspected of having coronary artery disease. AB - PURPOSE: To prospectively evaluate the feasibility, image quality, and accuracy of coronary magnetic resonance (MR) angiography at 3.0 T in patients suspected of having coronary artery disease and to prospectively compare these results with those of coronary MR angiography performed at 1.5 T. MATERIALS AND METHODS: The study was approved by the institutional review board, and informed consent was obtained from all patients. Eighteen patients (11 men, seven women; mean age, 63 years; age range, 45-76 years) suspected of having coronary artery disease who were scheduled to undergo elective conventional coronary angiography (reference standard) were included. For coronary MR angiography at 3.0 and 1.5 T, a vector electrocardiographically gated three-dimensional segmented k-space gradient-echo imaging sequence was combined with real-time respiratory navigator gating and tracking. Signal-to-noise ratios (SNRs), contrast-to-noise ratios (CNRs), scores of image quality and sensitivity and specificity for the detection of coronary artery stenosis on a segment-by-segment basis were assessed at 3.0 and 1.5 T. Data were analyzed for statistical differences by using the Wilcoxon matched pairs test and the McNemar test. RESULTS: The average increase in SNR at 3.0 T with respect to that at 1.5 T was 29.5% for the left coronary artery (LCA) and 31.2% for the right coronary artery (RCA) (P < .001), and the average increase in CNR was 21.8% for the LCA and 23.5% for the RCA (P < .001). Scores of image quality (P = .77) and diagnostic accuracy for the detection of coronary artery stenoses (sensitivity and specificity: 82% and 89%, respectively, at 3.0 T vs 82% and 88% at 1.5 T; P > .99) were identical or almost identical at both field strengths. CONCLUSION: Coronary MR angiography at 3.0 T is feasible in patients suspected of having coronary artery disease and yields significant increases in SNR and CNR, although current techniques do not result in significantly improved image quality and diagnostic accuracy compared with the quality and accuracy at 1.5 T. (c) RSNA, 2005. PMID- 15665222 TI - Transgluteal approach for draining pelvic fluid collections in pediatric patients. AB - PURPOSE: To evaluate a transgluteal approach for draining pelvic fluid collections in pediatric patients. MATERIALS AND METHODS: Institutional review board approval and informed consent for interventional procedures were obtained. From September 1995 to December 2002, 140 abdominopelvic fluid collections were drained at two institutions. Retrospective analysis of a prospectively gathered procedural database revealed that a transgluteal approach was used to drain 53 of these 140 collections (38%) in 51 patients (29 female, 22 male; mean age, 11.2 years; age range, 0.2-19 years). All patients received antibiotics before the procedure. A retrospective analysis was conducted of the patients' medical records and diagnostic imaging and interventional procedure findings, including the reason for referral; location, volume, and character of collections; method and equipment used for evacuation; duration of therapy; evidence of complications; results of microbiologic examination of specimens; and success rates. RESULTS: Transgluteal drainage was performed with computed tomographic (CT) guidance in 45 of the 53 collections (85%), with fluoroscopic guidance in three (6%), and with a combination of both modalities in five (9%). A drainage catheter was successfully placed in 49 collections; four small collections were aspirated without drain placement. Infected fluid was obtained from 41 collections, and serosanguineous fluid was obtained from 12 collections. The mean volume aspirated was 80 mL (2-600 mL). A positive culture was obtained at 28 of the 53 procedures. The mean duration of catheter placement was 4 days (range, 2 14 days). There were no major complications. CONCLUSION: The transgluteal approach to the drainage of abdominopelvic fluid collections with imaging guidance is safe and effective. PMID- 15665223 TI - CT of deep venous thrombosis and pulmonary embolus: does iso-osmolar contrast agent improve vascular opacification? AB - PURPOSE: To prospectively compare the vascular attenuation achieved with the iso osmolar dimeric contrast agent iodixanol with that achieved with the nonionic monomeric contrast agent iohexol for computed tomographic (CT) venography after CT pulmonary angiography. MATERIALS AND METHODS: Institutional review board approval and informed consent were obtained, and 51 consecutive patients undergoing CT pulmonary angiography and CT venography were recruited. A 130-mL dose of iodixanol 320 was injected intravenously at a rate of 4 mL/sec and followed by injection of 50 mL of saline. CT venography was performed after 3.5 minutes. From prior studies, 51 patients of similar weight were picked as control subjects. They received a similar iodine load with iohexol 300 and were studied with a similar technique. Section thickness was 1.25 mm for pulmonary emboli and 5 mm for deep venous thrombosis. Test and control group characteristics (ie, sex, age, and weight) were not significantly different (P >.05). Additionally, in test patients who had undergone CT pulmonary angiography and CT venography during the two preceding years, current and previously obtained CT scans were compared (ie, paired studies). Regions of interest were measured in four pulmonary artery and four lower extremity vein locations by two independent observers. RESULTS: Iodixanol increased average attenuation by 7 HU (P <.05) in the lower extremities and decreased average attenuation by 42 HU (P <.05) in the pulmonary arteries. In the 11 paired studies, similar results were obtained. CONCLUSION: Iodixanol caused a modest but statistically significant improvement in venous attenuation and a decrease in arterial attenuation. The diagnostic importance of this small increase in venous attenuation is not clear. PMID- 15665224 TI - Diffusion-tensor MR tractography of somatotopic organization of corticospinal tracts in the internal capsule: initial anatomic results in contradistinction to prior reports. AB - The goal of this study was to use diffusion-tensor magnetic resonance (MR) imaging to define the location and organization of corticospinal tracts (CSTs) in the posterior limb of the internal capsule (PLIC). The Institutional Review Board approved the study, and informed consent was obtained from all subjects. Eight volunteers and two patients with brain tumor were imaged at 3 T. All CSTs were found to lie in a compact area in one part of the PLIC: If the PLIC is divided into four equal quarters from anterior to posterior, the CST was shown to be in the third quarter. Seventeen of 20 CSTs were organized somatotopically, with hand fibers anterolateral to foot fibers, not anteromedial as is currently believed. In three of 20, hand and foot fibers were intermixed. Classically, it was thought that the CST was located in the anterior third of the PLIC. The present data confirm recent results that the CST is located more posteriorly. In the majority of cases, however, the CST is organized somatotopically. PMID- 15665225 TI - Appropriateness of a donor liver with respect to macrosteatosis: application of artificial neural networks to US images--initial experience. AB - PURPOSE: To retrospectively compare performance of artificial neural networks (ANNs) applied to ultrasonographic (US) images with that of radiologists for prediction of appropriateness of a donor liver with respect to macrosteatosis before liver transplantation. MATERIALS AND METHODS: Institutional ethics committee approved study; written informed consent was obtained. ANNs, constructed with three-layered 15-neuron back-propagation algorithm, were trained to predict appropriateness of a donor liver with respect to macrosteatosis by using statistically significant laboratory and US parameters derived from univariate analyses, together with correct diagnosis. Input variables for ANNs were alkaline phosphatase, glutamic oxaloacetic transaminase, glutamic pyruvate transaminase, gamma-glutamyltransferase, hepatorenal ratio of echogenicity, and tail area ratio and tail length of portal vein wall echogenicity. Three radiologists graded US images in 94 potential donors (71 men and 23 women) on the basis of four degrees of hepatic steatosis. After training and testing of ANNs, performance of ANNs and radiologists in predicting appropriateness of potential donors was evaluated with receiver operating characteristic (ROC) analysis and compared by means of univariate z score test. RESULTS: Among 94 potential donor livers, 76 were normal or had mild steatosis, and 18 had moderate or severe macrosteatosis at histopathologic examination. Area under ROC curve (Az) of ANNs (Az=0.9673) was significantly greater than that of radiologists (faculty, Az=0.9106, P=.048; fellow, Az= 0.9038, P=.044; resident, Az=0.8931, P=.038). No statistically significant difference in sensitivity for predicting appropriateness as a liver donor with respect to macrosteatosis was found between ANNs (88.9%) and radiologists (P >.05). However, specificity of ANNs (96.1%) was significantly better than that of radiologists (P <.003). CONCLUSION: ANNs might be a useful tool to categorize whether a donor liver is appropriate for transplantation with respect to macrosteatosis on the basis of multiple variables related to laboratory and US features. Further study is needed. PMID- 15665226 TI - Early-stage hepatocellular carcinoma in patients with cirrhosis: long-term results of percutaneous image-guided radiofrequency ablation. AB - PURPOSE: To perform a prospective, intention-to-treat clinical trial to determine the long-term survival rates of patients with hepatic cirrhosis and early-stage hepatocellular carcinoma (HCC) in whom percutaneous image-guided radiofrequency (RF) ablation was used as the sole first-line anticancer treatment. MATERIALS AND METHODS: The study was performed with approval of the ethics committee, and written informed consent was obtained for all patients. From June 1, 1996, to January 1, 2003, 206 patients (143 men, 63 women; age range, 51-81 years; mean age, 67 years +/- 7) who were excluded from surgery and who had Child class A or B cirrhosis with either a single HCC less than or equal to 5 cm in diameter or multiple (as many as three) HCCs less than or equal to 3 cm in diameter each were enrolled. RF ablation was performed in 187 (91%) of 206 patients; 19 (9%) were excluded from RF treatment because of unfavorable tumor location. Follow-up ranged from 3 to 78 months (mean, 24 months +/- 21) and included measurement of alpha-fetoprotein level, ultrasonography at 3-month intervals, and spiral computed tomography at 6-month intervals. Patients were observed for recurrence of the treated tumor and for the emergence of new HCC tumors. Survival probabilities were estimated with the Kaplan-Meier method, and differences between survival curves were evaluated with the log-rank test. RESULTS: At the end of the study, 145 patients were alive, and 61 were dead. In the intention-to treat analysis, overall survival rates were 97% at 1 year, 67% at 3 years, and 41% at 5 years. Median survival was 49 months. In the 187 patients treated with RF ablation, overall survival rates were 97% at 1 year, 71% at 3 years, and 48% at 5 years. Median survival was 57 months. The difference between the two survival curves was not statistically significant (P=.5094). Survival of patients treated with RF ablation was dependent on Child class (P=.0006) and tumor multiplicity (P=.0133). Patients who had Child class A cirrhosis with solitary HCC (n=116) had 1-, 3-, and 5-year survival rates of 100%, 89% and 61%; median survival was 65 months. The 1-, 3-, and 5-year recurrence rates were 14%, 49%, and 81% for the emergence of new tumors and 4%, 10%, and 10% for local tumor progression. CONCLUSION: RF ablation is an effective first-line treatment for cirrhotic patients with early-stage HCC who were excluded from surgery. PMID- 15665227 TI - Three-dimensional dynamic susceptibility-weighted perfusion MR imaging at 3.0 T: feasibility and contrast agent dose. AB - PURPOSE: To prospectively investigate if T2*-weighted dynamic susceptibility weighted first-pass perfusion magnetic resonance (MR) imaging is feasible at 3.0 T and which dose of contrast agent is suitable for high-field-strength imaging. MATERIALS AND METHODS: Informed consent was obtained from all participants; study protocol was approved by the institutional review board. Study included three volunteers (two men, one woman aged 35, 39, and 52 years) and 26 patients (mean age, 49 years +/- 12.8 [standard deviation]; range, 19-76 years). Volunteers underwent 3.0-T perfusion MR imaging with 0.20, 0.10, and 0.05 mmol per kilogram body weight of gadopentetate dimeglumine; patients underwent imaging with 0.10- and 0.05-mmol doses. Perfusion MR imaging was performed with three-dimensional echo-shifted echo-planar imaging (repetition time msec/echo time msec, 14/21; isotropic 4 mm3 voxels; 50 dynamic volumes with 30 sections each, covering entire brain at temporal resolution of 1.5 seconds per MR image). Quality of source echo planar images and perfusion maps was assessed; perfusion maps obtained at studies with different contrast media doses were compared. Quantitative perfusion values and diagnostic sensitivity of perfusion studies with 0.10-mmol dose were compared with results with 0.05-mmol dose. Image quality scores were compared with marginal homogeneity test for multinomial variables (Mantel-Haenszel statistics for ordered categorized values). Signal-to-noise ratio and baseline signal intensity in perfusion studies were tested (Student t test for paired samples). Mean transit time (MTT), negative integral (NI), and maximum T2* effect from region-of-interest analysis were compared (one-tailed Student t test for paired samples). Quantitative data on number of gamma-fitted pixels were compared (t test for paired samples). Difference with P=.05 (t test for paired samples) was considered significant. RESULTS: Perfusion image quality was satisfactory even in areas close to skull base (47 of 52 images, minor distortions; remaining images, marked distortions). Perfusion imaging with 0.20-mmol dose caused almost complete signal cancellation during first pass, particularly in cortical gray matter, since mean maximum T2* effect of 98%, 99%, and 98% for gray matter was reached such that the accurate calculation of perfusion maps was impossible. With 0.10 mmol dose, the NI and maximum T2* effect were comparable to published data for 1.5-T perfusion imaging with 0.20- and 0.05-mmol doses; perfusion maps of sufficient diagnostic quality were obtained. For gray matter, mean maximum T2* effect was 25.4% +/- 9.8 with 0.10-mmol dose and 17.5% +/- 9.0 with 0.05-mmol dose. For white matter, mean maximum T2* effect was 15.2% +/- 4.5 with 0.10-mmol dose and 7.7% +/- 2.9 with 0.05-mmol dose. Difference in maximum signal intensity decrease was significant (P <.01). For NI, the difference between 0.10- and 0.05 mmol doses was significant: For gray matter, mean NI was 3.1 +/- 1.60 for 0.10 mmol dose and 1.56 +/- 1.16 for 0.05-mmol dose. For white matter, mean NI was 1.35 +/- 0.59 with 0.1-mmol dose and 0.59 +/- 0.30 with 0.05-mmol dose. CONCLUSION: With echo-shifted multishot echo-planar imaging, dynamic susceptibility-weighted perfusion MR imaging at high field strength is feasible without relevant image distortions. Compared with contrast agent dose for 1.5 T imaging, the dose for 3.0 T can be reduced to 0.10 mmol. PMID- 15665228 TI - Minimum number of basis projections for caries detection with local CT. AB - OBJECTIVES: To investigate the relationship between the number of basis projections for local computed tomography (CT) and the detection of proximal caries and to find a minimum number of projections needed to maintain diagnostic accuracy. METHODS: We presented observers (n = 12) with stacks of both axial and vertical CT slices of 23 extracted teeth placed in a dry human mandible. The slices were prepared with 14, 20, 33 and 100 basis projections. The observers scored the proximal surfaces for the presence of caries on a 1-5 confidence scale. The performance of the varying number of projections was compared with conventional digital radiographs. RESULTS: The performance of all four CT modalities was significantly better than conventional radiographs (P = 0.005 to P = 0.021) and showed a consistent increase with the number of projections. Diagnostic performance depended significantly on lesion depth (P = 0.00), but not on observer. CONCLUSIONS: We conclude that the number of CT projections used can be reduced at least to 20 with the diagnostic performance still markedly better than that of conventional film, provided that the observer can make use of both axial and vertical stacks of CT slices. PMID- 15665229 TI - Effect of number of projections on image quality of local CT. AB - OBJECTIVES: To determine the image quality of the local CT imaging procedure, in terms of resolution, contrast, and noise as a function of the number of projections. METHODS: The contrast and resolution of the images was determined with a phantom object consisting of three rods of different materials, as well as a phantom human head embedded in soft tissue equivalent material. In addition, slices reconstructed from computed sinograms were used for comparison. RESULTS: Sharpness, contrast and noise were determined as a function of the number of projections. The number of projections was found to affect the contrast and the noise most, and had much less influence on resolution. CONCLUSIONS: Judging from the images of the phantom head and the numerical data, it seems that the minimum number of projections needed to obtain images of useful quality in the geometry used is about 33. Improved image quality (at any number of projections) can best be achieved through noise suppression. PMID- 15665230 TI - Prediction of premolar tooth lengths based on their panoramic radiographic lengths. AB - OBJECTIVE: Establishing a linear regression model for the determination of first premolar lengths based on panoramic radiographs. MATERIALS AND METHODS: The real lengths of 112 first premolars extracted for orthodontic reasons were measured and compared with their panoramic lengths. The teeth were divided into four groups according to their intraoral quadrant locations (T14, T24, T34, T44) and regression analysis was conducted for each group. RESULTS: A linear regression model for the prediction of tooth length (mm) based on the panoramic length was established (P < 0.0001): For group T14 the predicted length = (panoramic length x 0.698) + 2.61. For group T24 the predicted length = (panoramic length x 0.5056) + 7.844. For group T34 the predicted length = (panoramic length x 0.5075) + 9.282. For group T44 the predicted length = (panoramic length x 0.436) + 11.298. CONCLUSION: Prediction of all first premolar lengths using their panoramic images is both feasible and reliable. PMID- 15665231 TI - Simple bone cyst versus odontogenic keratocyst: differential diagnosis by digitized panoramic radiography. AB - OBJECTIVES: To identify the contour and pixel grey levels of simple bone cyst (SBC) and odontogenic keratocyst (OKC) images that might contribute to differentiating between the two lesions. METHODS: Panoramic radiographs of 32 SBC and 20 OKC were digitized and analysed by six examiners. The contours of the images were classified as indistinct, distinct without a sclerotic border, and distinct with a sclerotic border. The presence or absence of scalloping and the pixel grey levels of the radiolucent part of the images were also determined. RESULTS: The sclerotic border was more frequent in the OKC, especially in the posterior segment (P = 0.0301). The presence of scalloping was more frequent in the superior segment of the SBC (P = 0.0295). The pixel values were higher in the OKC than in the SBC images (P = 0.00134). CONCLUSIONS: Details about the contour and pixel grey levels of OKC and SBC assist in the diagnosis of these lesions. PMID- 15665232 TI - A digital subtraction radiography scheme based on automatic multiresolution registration. AB - OBJECTIVES: To establish a digital subtraction radiography scheme for aligning clinical in vivo radiographs based on the implementations of an automatic geometric registration method and a contrast correction technique. METHODS: Thirty-five pairs of in vivo dental radiographs from four clinical studies were used in this work. First, each image pair was automatically aligned by applying a multiresolution registration strategy using the affine transformation followed by the implementation of the projective transformation at full resolution. Then, a contrast correction technique was applied in order to produce subtraction radiographs and fused images for further clinical evaluation. The performance of the proposed registration method was assessed against a manual method based on the projective transformation. RESULTS: The qualitative assessment of the experiments based on visual inspection has shown advantageous performance of the proposed automatic registration method against the manual method. Furthermore, the quantitative analysis showed statistical difference in terms of the root mean square (RMS) error estimated over the whole images and specific regions of interest. CONCLUSIONS: The proposed automatic geometric registration method is capable of aligning radiographs acquired with or without rigorous a priori standardization. The methodology is pixel-based and does not require the application of any segmentation process prior to alignment. The employed projective transformation provides a reliable model for registering intraoral radiographs. The implemented contrast correction technique sequentially applied provides subtraction radiographs and fused images for clinical evaluation regarding the evolution of a disease or the response to a therapeutic scheme. PMID- 15665233 TI - Evaluation of the usefulness of the limited cone-beam CT (3DX) in the assessment of the thickness of the roof of the glenoid fossa of the temporomandibular joint. AB - OBJECTIVES: The purpose of this study was to evaluate the usefulness of the limited cone-beam X-ray CT (3DX) (Morita Co., Japan) in measuring the thickness of the roof of the glenoid fossa (RGF) of the temporomandibular joint (TMJ). MATERIALS AND METHODS: Twenty-one TMJs removed at autopsy from 21 cadavers were investigated macroscopically using dissection and 3DX imaging. A Digimatic Outside Micrometer and a 3DX-image tool were used to measure the minimum thickness of the RGF. Multiple measurements were made to identify the thinnest area. Once the thinnest areas had been identified, three linear measurements were made and the average value was used for statistical analysis. RESULTS: The average macroscopic examination measurement was 1.37 mm (range 0.55-3.6 mm) and the average 3DX image measurement was 1.22 mm (range: 0.51-3.0 mm). There was no significant difference between these two groups using the Mann-Whitney U-test (P < 0.05). The Spearman's correlation coefficient by rank between these two groups was r = 0.93(P < 0.001). CONCLUSION: These results suggest that bone thickness measurements of the RGF by 3DX imaging was effective. PMID- 15665234 TI - The nasopalatine canal revisited using 2D and 3D CT imaging. AB - OBJECTIVES: To assess the location, morphology and dimensions of the nasopalatine canal on two-dimensional and three-dimensional (2D and 3D) CT images. METHODS: Material included 34 spiral CT scans for pre-operative planning of implant placement in the maxilla. Scanning was performed using a standard exposure and patient positioning protocol. 2D and 3D spiral CT images were carefully examined for the location, morphology and dimensions of the nasopalatine canal by two independent observers. A comparison was made between 2D observations and a 2D/3D combined observation strategy (paired t-tests). RESULTS: The nasopalatine canal typically appeared as a canal with a mean (standard deviation (SD)) length of 8.1 (3.4) mm. Its palatal opening is the incisive foramen with a mean (SD) inner O of 4.6 (1.8) mm. At the level of the nasal floor often 2 (Y-canal morphology), but sometimes 3 or 4 openings could be observed. In particular cases, the canal showed up as a cylinder with only one nasal opening. The average (SD) maximum width of the nasopalatine canal structure at the level of the nasal floor was 4.9 (1.2) mm. The buccopalatal width of the jaw, anterior to the canal was 7.4 (2.6) mm. Interpretation of canal morphology was significantly different when comparing 2D image observation with a 2D/3D combined observation strategy. However, dimensional measurements of the canal were not significantly different for a 2D and a combined 2D/3D approach. CONCLUSIONS: The nasopalatine canal may show important anatomical variations, both with regard to morphology and dimensions. To avoid any potential complications during surgical procedures such as implant placement, a careful pre-operative observation is required. Cross-sectional imaging may be advocated to determine canal morphology and dimensions and to assess anterior bone width for potential implant placement buccally to the canal. PMID- 15665236 TI - Ameloblastomatous calcifying odontogenic cyst in the mandible. AB - Ameloblastoma is one of the well-known odontogenic tumours that can be associated with calcifying odontogenic cysts (COCs), but only a few reports include detailed clinical and radiographic features. In this paper we demonstrate a case of large ameloblastomatous COC in the mandible. The radiographic examination revealed the presence of a multilocular cystic lesion in the right posterior part of the mandible containing the impacted lower second molar with remarkable expansion toward both lingual and buccal side. This feature was different from the general findings of COC and rather resembled that of ameloblastomas. PMID- 15665235 TI - The WEBD project: a research of new methodologies for a distant-learning 3D system prototype. AB - OBJECTIVES: To create and to spread a new interactive multimedia instrument, based upon virtual reality technologies, that allows both the running simulation of machines and equipment and the reproduction via Web of complex three dimensional (3D) anatomical models such as the skull. METHODS: There were two main aspects of the project, one of design engineering and the other biomedical engineering, for the creation of "artificial" and anatomical objects. The former were made with 3D Studio Max R4 by Autodesk, San Rafael, CA, while the latter were created starting from real bones scanned with a CT system or a surface scanner and elaborated with different programs (3D Studio Max R4, Scenebuilder by Viewpoint, New York, NY and Spinfire by Actify, San Francisco, CA). The 3D models were to be integrated into web modules and had to respect file limits while preserving a sufficient definition. Two systems of evaluation were used, a questionnaire on a selected sample and an external evaluation by a different university. RESULTS: The Viewpoint format offers the best interactivity and size reduction (up to 96% from the original 3D model). The created modules included production of radiological images, rapid prototyping, and anatomy. The complete "3D Distant Learning Prototype" is available at www.webd.etsii.upm.es. CONCLUSIONS: The software currently available permits the construction of interactive modules. The verification on the selected sample and the evaluation by the University of Naples show that the structure is well organized and that the integration of the 3D models meets the requirements. PMID- 15665237 TI - Unusual combination of presenting features in multiple myeloma. AB - Initial oral signs of multiple myeloma may involve pain, paraesthesia, swelling, tooth mobility and radiolucency. A 62-year-old female patient presented with the chief complaint of numbness in the right lower jaw. Intraoral examination revealed no abnormalities and her history was non-contributory. A panoramic radiograph revealed an irregular, small radiopacity in the premolar region and small, multiple and diffuse osteolytic alterations in the mandible. Computed tomography revealed maxillary and mandibular cortical destruction. An incisional biopsy was performed. The histopathological diagnosis was plasma cell myeloma and further investigations lead to the diagnosis of multiple myeloma. Dysfunction of a nerve coursing through diseased bone is an ominous sign and may be an indication that a malignant process is present. PMID- 15665238 TI - Digitized conventional radiograph. PMID- 15665240 TI - Reductions of Rubisco activase by antisense RNA in the C4 plant Flaveria bidentis reduces Rubisco carbamylation and leaf photosynthesis. AB - To function, the catalytic sites of Rubisco (EC 4.1.1.39) need to be activated by the reversible carbamylation of a lysine residue within the sites followed by rapid binding of magnesium. The activation of Rubisco in vivo requires the presence of the regulatory protein Rubisco activase. This enzyme is thought to aid the release of sugar phosphate inhibitors from Rubisco's catalytic sites, thereby influencing carbamylation. In C3 species, Rubisco operates in a low CO2 environment, which is suboptimal for both catalysis and carbamylation. In C4 plants, Rubisco is located in the bundle sheath cells and operates in a high CO2 atmosphere close to saturation. To explore the role of Rubisco activase in C4 photosynthesis, activase levels were reduced in Flaveria bidentis, a C4 dicot, by transformation with an antisense gene directed against the mRNA for Rubisco activase. Four primary transformants with very low activase levels were recovered. These plants and several of their segregating T1 progeny required high CO2 (>1 kPa) for growth. They had very low CO2 assimilation rates at high light and ambient CO2, and only 10% to 15% of Rubisco sites were carbamylated at both ambient and very high CO2. The amount of Rubisco was similar to that of wild-type plants. Experiments with the T1 progeny of these four primary transformants showed that CO2 assimilation rate and Rubisco carbamylation were severely reduced in plants with less than 30% of wild-type levels of activase. We conclude that activase activity is essential for the operation of the C4 photosynthetic pathway. PMID- 15665241 TI - beta-Maltose is the metabolically active anomer of maltose during transitory starch degradation. AB - Maltose is the major form of carbon exported from the chloroplast at night as a result of transitory starch breakdown. Maltose exists as an alpha- or beta anomer. We developed an enzymatic technique for distinguishing between the two anomers of maltose and tested the accuracy and specificity of this technique using beta-maltose liberated from maltoheptose by beta-amylase. This technique was used to investigate which form of maltose is present during transitory starch degradation in bean (Phaseolus vulgaris), wild-type Arabidopsis (Arabidopsis thaliana), two starch deficient Arabidopsis lines, and one starch-excess mutant of Arabidopsis. In Phaseolus and wild-type Arabidopsis, beta-maltose levels were low during the day but were much higher at night. In Arabidopsis plants unable to metabolize maltose due to a T-DNA insertion in the gene for the cytosolic amylomaltase, (Y. Lu, T.D. Sharkey [2004] Planta 218: 466-473) levels of alpha- and beta-maltose were high during both the day and night. In starchless mutants of Arabidopsis, total maltose levels were low and almost completely in the alpha form. We also found that the subcellular concentration of beta-maltose at night was greater in the chloroplast than in the cytosol by 278 microm. We conclude that beta-maltose is the metabolically active anomer of maltose and that a sufficient gradient of beta-maltose exists between the chloroplast and cytosol to allow for passive transport of maltose out of chloroplasts at night. PMID- 15665243 TI - Enhanced photosynthetic performance and growth as a consequence of decreasing mitochondrial malate dehydrogenase activity in transgenic tomato plants. AB - Transgenic tomato (Solanum lycopersicum) plants expressing a fragment of the mitochondrial malate dehydrogenase gene in the antisense orientation and exhibiting reduced activity of this isoform of malate dehydrogenase show enhanced photosynthetic activity and aerial growth under atmospheric conditions (360 ppm CO2). In comparison to wild-type plants, carbon dioxide assimilation rates and total plant dry matter were up to 11% and 19% enhanced in the transgenics, when assessed on a whole-plant basis. Accumulation of carbohydrates and redox-related compounds such as ascorbate was also markedly elevated in the transgenics. Also increased in the transgenic plants was the capacity to use L-galactono-lactone, the terminal precursor of ascorbate biosynthesis, as a respiratory substrate. Experiments in which ascorbate was fed to isolated leaf discs also resulted in increased rates of photosynthesis providing strong indication for an ascorbate mediated link between the energy-generating processes of respiration and photosynthesis. This report thus shows that the repression of this mitochondrially localized enzyme improves both carbon assimilation and aerial growth in a crop species. PMID- 15665242 TI - Arabidopsis sphingosine kinase and the effects of phytosphingosine-1-phosphate on stomatal aperture. AB - Sphingolipids are a major component of membrane lipids and their metabolite sphingosine-1-phosphate (S1P) is a potent lipid mediator in animal cells. Recently, we have shown that the enzyme responsible for S1P production, sphingosine kinase (SphK), is stimulated by the phytohormone abscisic acid in guard cells of Arabidopsis (Arabidopsis thaliana) and that S1P is effective in regulating guard cell turgor. We have now characterized SphK from Arabidopsis leaves. SphK activity was mainly associated with the membrane fraction and phosphorylated predominantly the Delta4-unsaturated long-chain sphingoid bases sphingosine (Sph) and 4,8-sphingadienine, and to a lesser extent, the saturated long-chain sphingoid bases dihydrosphingosine and phytosphingosine (Phyto-Sph). 4 Hydroxy-8-sphingenine, which is a major sphingoid base in complex glycosphingolipids from Arabidopsis leaves, was a relatively poor substrate compared with the corresponding saturated Phyto-Sph. In contrast, mammalian SphK1 efficiently phosphorylated Sph, dihydrosphingosine, and 4,8-sphingadienine, but not the 4-hydroxylated long-chain bases Phyto-Sph and 4-hydroxy-8-sphingenine. Surface dilution kinetic analysis of Arabidopsis SphK with Sph presented in mixed Triton X-100 micelles indicated that SphK associates with the micellar surface and then with the substrate presented on the surface. In addition, measurements of SphK activity under different assay conditions combined with phylogenetic analysis suggest that multiple isoforms of SphK may be expressed in Arabidopsis. Importantly, we found that phytosphingosine-1-phosphate, similar to S1P, regulates stomatal apertures and that its action is impaired in guard cells of Arabidopsis plants harboring T-DNA null mutations in the sole prototypical G protein alpha-subunit gene, GPA1. PMID- 15665244 TI - Multiple protein regions contribute to differential activities of YABBY proteins in reproductive development. AB - Members of the YABBY family of putative transcription factors participate in abaxial-adaxial identity determination in lateral organs in Arabidopsis (Arabidopsis thaliana). Two YABBY genes specifically expressed in reproductive structures, CRABS CLAW (CRC) and INNER NO OUTER (INO), have additional activities, with CRC promoting nectary development and carpel fusion, and INO responding to spatial regulation by SUPERMAN during ovule development. All YABBY coding regions, except YABBY5, were able to restore outer integument growth in ino-1 mutants when expressed from the INO promoter (PRO(INO)). However, INO was the only YABBY family member that responded correctly to SUPERMAN to maintain the wild-type gynoapical-gynobasal asymmetry of the outer integument. By contrast, INO, FILAMENTOUS FLOWER, and YABBY3 failed to complement crc-1 when expressed from PRO(CRC). Roles of individual regions of CRC and INO in these effects were assessed using chimeric proteins with PRO(INO) and PRO(CRC) and the relatively constitutive cauliflower mosaic virus PRO(35S). Regions of CRC were found to contribute additively to CRC-specific functions in nectary and carpel formation, with a nearly direct relationship between the amount of CRC included and the degree of complementation of crc-1. When combined with INO sequences, the central and carboxyl-terminal regions of CRC were individually sufficient to overcome inhibitory effects of SUPERMAN within the outer integument. Reproductive phenotypes resulting from constitutive expression were dependent on the nature of the central region with some contributions from the amino terminus. Thus, the YABBY family members have both unique and common functional capacities, and residues involved in differential activities are distributed throughout the protein sequences. PMID- 15665245 TI - Alterations in tocopherol cyclase activity in transgenic and mutant plants of Arabidopsis affect tocopherol content, tocopherol composition, and oxidative stress. AB - Tocopherol belongs to the Vitamin E class of lipid soluble antioxidants that are essential for human nutrition. In plants, tocopherol is synthesized in plastids where it protects membranes from oxidative degradation by reactive oxygen species. Tocopherol cyclase (VTE1) catalyzes the penultimate step of tocopherol synthesis, and an Arabidopsis (Arabidopsis thaliana) mutant deficient in VTE1 (vte1) is totally devoid of tocopherol. Overexpression of VTE1 resulted in an increase in total tocopherol of at least 7-fold in leaves, and a dramatic shift from alpha-tocopherol to gamma-tocopherol. Expression studies demonstrated that indeed VTE1 is a major limiting factor of tocopherol synthesis in leaves. Tocopherol deficiency in vte1 resulted in the increase in ascorbate and glutathione, whereas accumulation of tocopherol in VTE1 overexpressing plants led to a decrease in ascorbate and glutathione. Deficiency in one antioxidant in vte1, vtc1 (ascorbate deficient), or cad2 (glutathione deficient) led to increased oxidative stress and to the concomitant increase in alternative antioxidants. Double mutants of vte1 were generated with vtc1 and cad2. Whereas growth, chlorophyll content, and photosynthetic quantum yield were very similar to wild type in vte1, vtc1, cad2, or vte1vtc1, they were reduced in vte1cad2, indicating that the simultaneous loss of tocopherol and glutathione results in moderate oxidative stress that affects the stability and the efficiency of the photosynthetic apparatus. PMID- 15665247 TI - Beyond complementation. Map-based cloning in Chlamydomonas reinhardtii. AB - Chlamydomonas reinhardtii is an excellent model system for plant biologists because of its ease of manipulation, facile genetics, and the ability to transform the nuclear, chloroplast, and mitochondrial genomes. Numerous forward genetics studies have been performed in Chlamydomonas, in many cases to elucidate the regulation of photosynthesis. One of the resultant challenges is moving from mutant phenotype to the gene mutation causing that phenotype. To date, complementation has been the primary method for gene cloning, but this is impractical in several situations, for example, when the complemented strain cannot be readily selected or in the case of recessive suppressors that restore photosynthesis. New tools, including a molecular map consisting of 506 markers and an 8X-draft nuclear genome sequence, are now available, making map-based cloning increasingly feasible. Here we discuss advances in map-based cloning developed using the strains mcd4 and mcd5, which carry recessive nuclear suppressors restoring photosynthesis to chloroplast mutants. Tools that have not been previously applied to Chlamydomonas, such as bulked segregant analysis and marker duplexing, are being implemented to increase the speed at which one can go from mutant phenotype to gene. In addition to assessing and applying current resources, we outline anticipated future developments in map-based cloning in the context of the newly extended Chlamydomonas genome initiative. PMID- 15665246 TI - Faster Rubisco is the key to superior nitrogen-use efficiency in NADP-malic enzyme relative to NAD-malic enzyme C4 grasses. AB - In 27 C4 grasses grown under adequate or deficient nitrogen (N) supplies, N-use efficiency at the photosynthetic (assimilation rate per unit leaf N) and whole plant (dry mass per total leaf N) level was greater in NADP-malic enzyme (ME) than NAD-ME species. This was due to lower N content in NADP-ME than NAD-ME leaves because neither assimilation rates nor plant dry mass differed significantly between the two C4 subtypes. Relative to NAD-ME, NADP-ME leaves had greater in vivo (assimilation rate per Rubisco catalytic sites) and in vitro Rubisco turnover rates (k(cat); 3.8 versus 5.7 s(-1) at 25 degrees C). The two parameters were linearly related. In 2 NAD-ME (Panicum miliaceum and Panicum coloratum) and 2 NADP-ME (Sorghum bicolor and Cenchrus ciliaris) grasses, 30% of leaf N was allocated to thylakoids and 5% to 9% to amino acids and nitrate. Soluble protein represented a smaller fraction of leaf N in NADP-ME (41%) than in NAD-ME (53%) leaves, of which Rubisco accounted for one-seventh. Soluble protein averaged 7 and 10 g (mmol chlorophyll)(-1) in NADP-ME and NAD-ME leaves, respectively. The majority (65%) of leaf N and chlorophyll was found in the mesophyll of NADP-ME and bundle sheath of NAD-ME leaves. The mesophyll-bundle sheath distribution of functional thylakoid complexes (photosystems I and II and cytochrome f) varied among species, with a tendency to be mostly located in the mesophyll. In conclusion, superior N-use efficiency of NADP-ME relative to NAD-ME grasses was achieved with less leaf N, soluble protein, and Rubisco having a faster k(cat). PMID- 15665248 TI - Microarray analysis confirms the specificity of a Chlamydomonas reinhardtii chloroplast RNA stability mutant. AB - The expression of chloroplast and mitochondrial genes depends on nucleus-encoded proteins, some of which control processing, stability, and/or translation of organellar RNAs. To test the specificity of one such RNA stability factor, we used two known Chlamydomonas reinhardtii nonphotosynthetic mutants carrying mutations in the Mcd1 nuclear gene (mcd1-1 and mcd1-2). We previously reported that these mutants fail to accumulate the chloroplast petD mRNA and its product, subunit IV of the cytochrome b6/f complex, which is essential for photosynthesis. Such mutants are generally presumed to be gene specific but are not tested rigorously. Here, we have used microarray analysis to assess changes in chloroplast, mitochondrial, and nuclear RNAs, and since few other RNAs were significantly altered in these mutants, conclude that Mcd1 is indeed specifically required for petD mRNA accumulation. In addition, a new unlinked nuclear mutation was discovered in mcd1-2, which greatly reduced chloroplast atpA mRNA accumulation. Genetic analyses showed failure to complement mda1-ncc1, where atpA containing transcripts are similarly affected (D. Drapier, J. Girard-Bascou, D.B. Stern, F.-A. Wollman [2002] Plant J 31: 687-697), and we have named this putative new allele mda1-2. We conclude that DNA microarrays are efficient and useful for characterizing the specificity of organellar RNA accumulation mutants. PMID- 15665249 TI - Comparative genomics of the pennate diatom Phaeodactylum tricornutum. AB - Diatoms are one of the most important constituents of phytoplankton communities in aquatic environments, but in spite of this, only recently have large-scale diatom-sequencing projects been undertaken. With the genome of the centric species Thalassiosira pseudonana available since mid-2004, accumulating sequence information for a pennate model species appears a natural subsequent aim. We have generated over 12,000 expressed sequence tags (ESTs) from the pennate diatom Phaeodactylum tricornutum, and upon assembly into a nonredundant set, 5,108 sequences were obtained. Significant similarity (E < 1E-04) to entries in the GenBank nonredundant protein database, the COG profile database, and the Pfam protein domains database were detected, respectively, in 45.0%, 21.5%, and 37.1% of the nonredundant collection of sequences. This information was employed to functionally annotate the P. tricornutum nonredundant set and to create an internet-accessible queryable diatom EST database. The nonredundant collection was then compared to the putative complete proteomes of the green alga Chlamydomonas reinhardtii, the red alga Cyanidioschyzon merolae, and the centric diatom T. pseudonana. A number of intriguing differences were identified between the pennate and the centric diatoms concerning activities of relevance for general cell metabolism, e.g. genes involved in carbon-concentrating mechanisms, cytosolic acetyl-Coenzyme A production, and fructose-1,6-bisphosphate metabolism. Finally, codon usage and utilization of C and G relative to gene expression (as measured by EST redundance) were studied, and preferences for utilization of C and CpG doublets were noted among the P. tricornutum EST coding sequences. PMID- 15665250 TI - Tonoplast intrinsic proteins AtTIP2;1 and AtTIP2;3 facilitate NH3 transport into the vacuole. AB - While membrane transporters mediating ammonium uptake across the plasma membrane have been well described at the molecular level, little is known about compartmentation and cellular export of ammonium. (The term ammonium is used to denote both NH3 and NH4+ and chemical symbols are used when specificity is required.) We therefore developed a yeast (Saccharomyces cerevisiae) complementation approach and isolated two Arabidopsis (Arabidopsis thaliana) genes that conferred tolerance to the toxic ammonium analog methylammonium in yeast. Both genes, AtTIP2;1 and AtTIP2;3, encode aquaporins of the tonoplast intrinsic protein subfamily and transported methylammonium or ammonium in yeast preferentially at high medium pH. AtTIP2;1 expression in Xenopus oocytes increased 14C-methylammonium accumulation with increasing pH. AtTIP2;1- and AtTIP2;3-mediated methylammonium detoxification in yeast depended on a functional vacuole, which was in agreement with the subcellular localization of green fluorescent protein-fusion proteins on the tonoplast in planta. Transcript levels of both AtTIPs were influenced by nitrogen supply but did not follow those of the nitrogen-derepressed ammonium transporter gene AtAMT1;1. Transgenic Arabidopsis plants overexpressing AtTIP2;1 did not show altered ammonium accumulation in roots after ammonium supply, although AtTIP2;1 mRNA levels in wild-type plants were up-regulated under these conditions. This study shows that AtTIP2;1 and AtTIP2;3 can mediate the extracytosolic transport of methyl-NH2 and NH3 across the tonoplast membrane and may thus participate in vacuolar ammonium compartmentation. PMID- 15665251 TI - Functional genomics of the regulation of the nitrate assimilation pathway in Chlamydomonas. AB - The existence of mutants at specific steps in a pathway is a valuable tool of functional genomics in an organism. Heterologous integration occurring during transformation with a selectable marker in Chlamydomonas (Chlamydomonas reinhardtii) has been used to generate an ordered mutant library. A strain, having a chimeric construct (pNia1::arylsulfatase gene) as a sensor of the Nia1 gene promoter activity, was transformed with a plasmid bearing the paramomycin resistance AphVIII gene to generate insertional mutants defective at regulatory steps of the nitrate assimilation pathway. Twenty-two thousand transformants were obtained and maintained in pools of 96 for further use. The mutant library was screened for the following phenotypes: insensitivity to the negative signal of ammonium, insensitivity to the positive signal of nitrate, overexpression in nitrate, and inability to use nitrate. Analyses of mutants showed that (1) the number or integrated copies of the gene marker is close to 1; (2) the probability of cloning the DNA region at the marker insertion site is high (76%); (3) insertions occur randomly; and (4) integrations at different positions and orientations of the same genomic region appeared in at least three cases. Some of the mutants analyzed were found to be affected at putative new genes related to regulatory functions, such as guanylate cyclase, protein kinase, peptidyl-prolyl isomerase, or DNA binding. The Chlamydomonas mutant library constructed would also be valuable to identify any other gene with a screenable phenotype. PMID- 15665252 TI - Wound-inducible biosynthesis of phytoalexin hydroxycinnamic acid amides of tyramine in tryptophan and tyrosine decarboxylase transgenic tobacco lines. AB - The wound-activated biosynthesis of phytoalexin hydroxycinnamic acid amides of tyramine was compared in untransformed and transgenic tobacco (Nicotiana tabacum) lines that express tryptophan decarboxylase (TDC), tyrosine decarboxylase (TYDC), or both activities. Transgenic in vitro-grown tobacco lines expressing TDC activity accumulated high levels of tryptamine but not hydroxycinnamic amides of tryptamine. In contrast, transgenic tobacco lines expressing TYDC accumulated tyramine as well as p-coumaroyltyramine and feruloyltyramine. The MeOH-soluble and cell wall fractions showed higher concentrations of wound-inducible p coumaroyltyramine and feruloyltyramine, especially at and around wound sites, in TYDC and TDC xTYDC tobacco lines compared to wild-type or TDC lines. All the enzymes involved in the biosynthesis of hydroxycinnamic acid amides of tyramine were found to be similarly wound inducible in all tobacco genotypes investigated. These results provide experimental evidence that, under some circumstances, TYDC activity can exert a rate-limiting control over the carbon flux allocated to the biosynthesis of hydroxycinnamic acid amides of tyramine. PMID- 15665254 TI - Molecular interactions between matrilysin and the matrix metalloproteinase inhibitor doxycycline investigated by deuterium exchange mass spectrometry. AB - Matrix metalloproteinases (MMPs) play an essential role in normal and pathological extracellular matrix degradation. Deuterium exchange mass spectrometry (DXMS) was used to localize the binding regions of the broad spectrum MMP inhibitor doxycycline on the active form of matrilysin (residues 95 267) and to assess alterations in structure induced by doxycycline binding. DXMS analyses of inhibitor-bound versus inhibitor-free forms of matrilysin reveal two primary sites of reduced hydrogen/deuterium exchange (residues 145-153; residues 193-204) that flank the structural zinc binding site. Equilibrium dialysis studies of doxycycline-matrilysin binding yielded a K(d) of 73 microM with a binding stoichiometry of 2.3 inhibitor molecules per protein, which compares well with DXMS results that show principal reduction in deuterium exchange at two sites. Lesser changes in deuterium exchange evident at the amino and carboxyl termini are attributed to inhibitor-induced structural fluctuations. Tryptophan fluorescence quenching experiments of matrilysin with potassium iodide suggest changes in conformation induced by doxycycline binding. In the presence of doxycycline, tryptophan quenching is reduced by approximately 17% relative to inhibitor-free matrilysin. Examination of the X-ray crystal structure of matrilysin shows that the doxycycline-binding site at residues 193 to 204 is positioned within the structural metal center of matrilysin, adjacent to the structural zinc atom and near both calcium atoms. These results suggest a mode of matrilysin inhibition by doxycycline that could involve interactions with the structural zinc atom and/or calcium atoms within the structural metal center of the protein. PMID- 15665255 TI - Magnesium deficiency in critical illness. AB - Magnesium (Mg) deficiency commonly occurs in critical illness and correlates with a higher mortality and worse clinical outcome in the intensive care unit (ICU). Magnesium has been directly implicated in hypokalemia, hypocalcemia, tetany, and dysrhythmia. Moreover, Mg may play a role in acute coronary syndromes, acute cerebral ischemia, and asthma. Magnesium regulates hundreds of enzyme systems. By regulating enzymes controlling intracellular calcium, Mg affects smooth muscle vasoconstriction, important to the underlying pathophysiology of several critical illnesses. The principle causes of Mg deficiency are gastrointestinal and renal losses; however, the diagnosis is difficult to make because of the limitations of serum Mg levels, the most common assessment of Mg status. Magnesium tolerance testing and ionized Mg2+ are alternative laboratory assessments; however, each has its own difficulties in the ICU setting. The use of Mg therapy is supported by clinical trials in the treatment of symptomatic hypomagnesemia and preeclampsia and is recommended for torsade de pointes. Magnesium therapy is not supported in the treatment of acute myocardial infarction and is presently undergoing evaluation for the treatment of severe asthma exacerbation, for the prevention of post-coronary bypass grafting dysrhythmias, and as a neuroprotective agent in acute cerebral ischemia. PMID- 15665253 TI - Targeting effector memory T cells with a selective peptide inhibitor of Kv1.3 channels for therapy of autoimmune diseases. AB - The voltage-gated Kv1.3 K(+) channel is a novel target for immunomodulation of autoreactive effector memory T (T(EM)) cells that play a major role in the pathogenesis of autoimmune diseases. We describe the characterization of the novel peptide ShK(L5) that contains l-phosphotyrosine linked via a nine-atom hydrophilic linker to the N terminus of the ShK peptide from the sea anemone Stichodactyla helianthus. ShK(L5) is a highly specific Kv1.3 blocker that exhibits 100-fold selectivity for Kv1.3 (K(d) = 69 pM) over Kv1.1 and greater than 250-fold selectivity over all other channels tested. ShK(L5) suppresses the proliferation of human and rat T(EM) cells and inhibits interleukin-2 production at picomolar concentrations. Naive and central memory human T cells are initially 60-fold less sensitive than T(EM) cells to ShK(L5) and then become resistant to the peptide during activation by up-regulating the calcium-activated K(Ca)3.1 channel. ShK(L5) does not exhibit in vitro cytotoxicity on mammalian cell lines and is negative in the Ames test. It is stable in plasma and when administered once daily by subcutaneous injection (10 mug/kg) attains "steady state" blood levels of approximately 300 pM. This regimen does not cause cardiac toxicity assessed by continuous EKG monitoring and does not alter clinical chemistry and hematological parameters after 2-week therapy. ShK(L5) prevents and treats experimental autoimmune encephalomyelitis and suppresses delayed type hypersensitivity in rats. ShK(L5) might prove useful for therapy of autoimmune disorders. PMID- 15665256 TI - Use of cellular and plasma apheresis in the critically ill patient: part 1: technical and physiological considerations. AB - Apheresis is the process of separating the blood and removing or manipulating a cellular or plasma component for therapeutic benefit. An apheresis procedure, or series of procedures, may be indicated in the critical care setting as primary or adjunctive therapy for certain hematologic, neurologic, renal, and autoimmune/ rheumatologic disorders. Optimal management of severely ill patients undergoing apheresis requires a working knowledge of the technical, methodological, and therapeutic considerations. These considerations include instrument hardware and separation methods, vascular access requirements, hemodynamic and hemostatic effects of the procedures, exposure to anticoagulants and homologous blood products, physiological variables affecting blood/plasma processing efficiency, and therapeutic endpoints for specific indications. Part 1 of this review will discuss each of those technological considerations and the basic physiological principles that guide this form of therapy. Part 2 of this series will deal with the clinical indications and applications for specific disorders that are most likely to affect patients in the intensive care unit. PMID- 15665257 TI - CVP and PAoP measurements are discordant during fluid therapy after traumatic brain injury. AB - The objective of the study was to compare measurements of central venous pressure (CVP) and pulmonary artery occlusion pressures (PAoP) as estimates of intravascular volume during the first 96 hours of fluid therapy after traumatic brain injury (TBI). One thousand five hundred ten simultaneous CVP and PAoP measurements from 31 patients entered into the National Acute Brain Injury Study: Hypothermia (NABISH:H) protocol were retrospectively compared. The effect of fluid administration and body temperature upon the paired measurements was statistically assessed. Agreement between CVP and PAoP values was poor. The CVP and PAoP were equal in only 11% of paired values. The CVP was always higher than PAoP in 1 patient, whereas PAoP always exceeded the CVP in 5 others. In 74% of the pairs, the PAoP was higher than the CVP, whereas in 15%, CVP was greater than PAoP. For any CVP measurement, the PAoP was either 3 mm Hg above or below the CVP in 67% of the pairs and at least 5 mm Hg above or below the CVP in 21% of the pairs. In 21 (68%) patients, PAoP was > or = 5 mm Hg above CVP in more than 4 readings, a clinically important difference. Discordance was not attributed to the fluid administered or to the temperature protocol. Neurological outcome appears affected by the volume of fluid administration. However, during initial therapy, estimates of intravascular volume provided by the CVP and PAoP are discordant. Although documented in other clinical conditions, the disparity noted here after TBI has not been previously reported. Assessment of intravascular volume to avoid hypovolemia should utilize other measurement techniques. PMID- 15665259 TI - Acute effects of upright position on gas exchange in patients with acute respiratory distress syndrome. AB - Patients with acute respiratory distress syndrome (ARDS) have dorsal atelectasis of the lungs. This is probably caused by several mechanisms: compression on dependent lung zones, purulent secretions in alveoli, and upward shift of the diaphragm. An upright position (UP) of the patient (the whole body in a straight line at 40 to 45 degrees) can theoretically ameliorate these mechanisms. The objective was to evaluate whether there was an improvement of gas exchange during UP of ARDS patients and to evaluate the hemodynamic effects. A prospective interventional study was performed in the surgical and medical ICUs and the burn unit of the Ghent University Hospital, a tertiary care center. Included were ARDS patients with onset of ARDS within 48 hours before start of the study. Patients were excluded when there was hemodynamic instability or when the PaO2/FiO2 ratio deteriorated during the 2 hours preceding UP. After a 2-hour observation period in a semirecumbent position, patients were put in UP for 12 hours. Respiration and hemodynamic data were recorded at the start and end of the 2-hour observation period, and after 1, 4, and 12 hours in UP. Eighteen patients were included in the study. There was a significant increase of the PaO2/FiO2 ratio during UP (P < .001). Except for the need for volume resuscitation in 5 patients (27.8%), there was no significant change in the hemodynamic profile of the patients. Upright positioning of patients with ARDS, a relatively simple maneuver, resulted in an improvement of gas exchange and was tolerated hemodynamically relatively well during a 12-hour observation period. PMID- 15665258 TI - A prospective multicenter study to evaluate the feasibility and safety of aggressive antihypertensive treatment in patients with acute intracerebral hemorrhage. AB - The authors performed a multicenter prospective observational study to evaluate the feasibility and safety of intravenous antihypertensive protocol for acute hypertension in patients with intracerebral hemorrhage (ICH). Twenty-seven patients with ICH and acute hypertension (mean age 61.37 +/- 14.27; 10 were men) were treated to maintain the systolic blood pressure (BP) below 160 mm Hg and diastolic BP below 90 mm Hg within 24 hours of symptom onset. Neurological deterioration (defined as a decrease in initial Glasgow Coma Scale score > or = 2) was observed in 2 (7.4%) of 27 patients during treatment. Among patients who underwent follow-up computed tomography, hematoma expansion (more than 33% increase in hematoma size at 24 hours) was observed in 2 (9.1%) of 22 patients. Patients treated within 6 hours of symptom onset were more likely to be functionally independent (modified Rankin scale < or = 2) at 1 month compared with patients who were treated between 6 and 24 hours (8 of 18 versus 0 of 9,P = .03). Aggressive pharmacological treatment of acute hypertension in patients with ICH can be initiated early with a low rate of neurological deterioration and hematoma expansion. PMID- 15665263 TI - Beyond concept analysis in holistic nursing. PMID- 15665260 TI - Isn't it the time to consider blood glucose concentration as part of the severity assessment of critically ill patients? PMID- 15665265 TI - Navigating the journey to menses cessation: a study of change in an emancipatory context. AB - PURPOSE: This article explores contemporary women's perceptions of the experience of the menopausal process within the Western industrialized culture, describes and synthesizes the processes used by women to evaluate available information, and explores and facilitates processes used by women to envision and create change in their life world. METHOD: An emancipatory group process was designed to facilitate dialogue between the investigator and nine women who met as a group eight times during a 10-week period in the home of one of the participants. FINDINGS: Participants identified menses cessation as a time of change in all aspects of their lives. They employed decision making to cope with these changes and, as a result of the awareness that came through the research process, invited other women to celebrate the collective wisdom of women at all stages of life. CONCLUSION: Women experiencing menses cessation need information and time to process that information internally and in relation with others. IMPLICATIONS: Nurses can facilitate the process by providing knowledge, assisting women in decision making, and intervening in ways that contribute to a holistic quality of life for women. PMID- 15665264 TI - Attributes of spiritual care in nursing practice. AB - Nurses are increasingly being called on to engage in spiritual care with their patients. A diverse body of theoretical and empirical literature addresses spirituality as it relates to nursing practice, yet there is little consensus about what spiritual nursing care entails. The purpose of this article is to conceptualize spiritual care in relation to nursing practice. A brief historical review indicates that our current understandings of spiritual nursing care have been shaped by three eras characterized by particular approaches: the religious approach, the scientific approach, and the existential approach. We draw elements from each of these approaches to propose attributes of spiritual care in the context of nursing practice. We propose that spiritual nursing care is an intuitive, interpersonal, altruistic, and integrative expression that is contingent on the nurse's awareness of the transcendent dimension of life but that reflects the patient's reality. PMID- 15665267 TI - Development and preliminary evaluation of the existential meaning scale. AB - PURPOSE: The aim of this study was to develop and examine the psychometric properties of the Existential Meaning Scale (EMS). FINDINGS: Construct validity of the EMS was examined through factor analysis and correlational analyses with theoretically related instruments. After several weak items were deleted, the 20 item scale had a Cronbach's alpha coefficient of > .80 in an overall sample of 418 individuals. Scores on the EMS were significantly lower in a sample of persons with HIV-1 infection than in the general population samples. CONCLUSION: Although its initial psychometric properties were satisfactory, additional validation of the EMS is necessary in other clinical populations to examine further the psychometric properties of the EMS. In addition, further examination of the responsiveness of the EMS over time is needed to evaluate its potential utility in longitudinal trials. PMID- 15665269 TI - Being involved and making a difference: empowerment and well-being among women living in poverty. AB - BACKGROUND: The relationships between poverty, empowerment, and health are of theoretical and practical interest to nurses, yet few researchers have examined volunteerism in poor communities from a holistic health perspective. PURPOSE: This study explores the experiences of women engaged in community volunteer work in the context of economic poverty. METHOD: Individual, in-depth interviews were conducted with 8 women, ages 21 to 77 years, who qualified for federal assistance or self-identified as low-income and were currently involved in volunteer community work. FINDINGS: Participants' stories of being involved and making a difference illustrated women making connections, developing relationships, gaining knowledge and skills, increasing self-esteem and confidence, reaching out to help others, and strengthening communities. CONCLUSION: At both the individual and community level, well-being was enhanced through women's community volunteerism in the context of economic poverty. IMPLICATIONS: At the individual and community levels, well-being was enhanced through women's community volunteerism in the context of economic poverty. From a holistic nursing perspective, the findings provide support for community volunteerism as a health promoting strategy among women living in economic poverty. PMID- 15665270 TI - Intentionality: evolutionary development in healing: a grounded theory study for holistic nursing. AB - Although intentionality has been implicated as a causal variable in healing research, its definition has been inconsistent and vague. The objective of this grounded theory study is to develop a substantive theory of intentionality in a naturalistic encounter between nurse-healers and their healee-clients, and to consider the implications for practice and research. Six expert nurse-healers and six healee-clients were interviewed as individuals and in dyads before and after treatments. Interviews and observational data were analyzed using the constant comparative method and synthesized analysis. Participants described their experience of intentionality in healing as an evolutionary process characterized by distinctive shifts. The theory of intentionality: the matrix for healing (IMH) includes definitions of intentionality and a conceptual framework of three developmental phases of intentionality (generic, healing, and transforming intentionalities). The predominant attribute, development, is described. The theory contributes to knowledge about healing and intentionality and has implications for practice and future research. PMID- 15665272 TI - A spliced isoform of interleukin 6 mRNA produced by renal cell carcinoma encodes for an interleukin 6 inhibitor. AB - Interleukin 6 (IL-6) is a multifunctional and pleiotropic cytokine and in renal cell carcinoma (RCC), this cytokine exerts proinflammatory, immunosuppressive and growth stimulating properties. A spliced isoform of IL-6 mRNA has been described in peripheral blood mononuclear cells and encodes for a potential protein lacking IL-6 activity. In the present study, a novel spliced form of IL-6 mRNA was found detectable in RCC cell lines, normal renal cells, but not in other tumor cells. This splicing resulted in a frameshift and the generation of multiple stop codon in the spliced IL-6 mRNA. However, two ATG of the third IL-6 exon were identified as translation initiation sites and two truncated IL-6 (tIL-6) with the expected molecular weight were recovered from transfected cell supernatant. The cDNA of a spliced form of IL-6 mRNA detected in RCC lines was cloned, and expressed in a baculovirus expression vector. The functional properties of the tIL-6 were investigated and this protein blocked IL-6 bioactivity, including mitogenic activity on tumor cells. In conclusion, this spliced form of IL-6 mRNA detected in RCC encodes for a truncated IL-6 with IL-6 antagonist properties. PMID- 15665273 TI - Down-regulation of BRCA1-BARD1 ubiquitin ligase by CDK2. AB - BRCA1, a breast and ovarian tumor suppressor, is a phosphoprotein whose cellular expression level is regulated in a cell cycle-dependent manner. BRCA1 interacts with BARD1 to generate significant ubiquitin ligase activity which catalyzes nontraditional Lys-6-linked polyubiquitin chains. However, it is not clear how the activity is regulated and how this affects BRCA1's multiple cellular functions. Here we show that the ubiquitin ligase activity of BRCA1-BARD1 is down regulated by CDK2. During the cell cycle, BARD1 expression can largely be categorized into three patterns: moderately expressed in a predominantly unphosphorylated form in early G(1) phase, expressed at low levels in both phosphorylated and unphosphorylated forms during late G(1) and S phases, and highly expressed in its phosphorylated form during mitosis coinciding with BRCA1 expression. CDK2-cyclin A1/E1 and CDK1-cyclin B1 phosphorylate BARD1 on its NH(2) terminus in vivo and in vitro. Intriguingly, the BRCA1-BARD1-mediated in vivo ubiquitination of nucleophosmin/B23 (NPM) and autoubiquitination of BRCA1 are dramatically disrupted by coexpression of CDK2-cyclin A1/E1, but not by CDK1 cyclin B1. The inhibition of ubiquitin ligase activity is not due to the direct effect of the kinases on BARD1 because an unphosphorylatable mutant of BARD1, S148A/S251A/S288A/T299A, is still inhibited by CDK2-cyclin E1. Alternatively, BRCA1 and BARD1 are likely exported to the cytoplasm and their expressions are remarkably reduced by CDK2-cyclin E1 coexpression. Recognizing the importance of cyclin E1 overexpression in breast cancer development, these results suggest a CDK2-BRCA1-NPM pathway that coordinately functions in cell growth and tumor progression pathways. PMID- 15665274 TI - Decreased fidelity in replicating CpG methylation patterns in cancer cells. AB - The unmethylated or methylated status of individual CpG sites is faithfully copied into daughter cells. Here, we analyzed the fidelity in replicating their methylation statuses in cancer cells. A single cell was clonally expanded, and methylation statuses of individual CpG sites were determined for an average of 12.5 DNA molecules obtained from the expanded population. By counting the deviation from the original methylation patterns inferred, the number of errors was measured. The analysis was done in four gastric cancer cell lines for five CpG islands (CGI), and repeated six times (total 1,495 clones sequenced). HSC39 and HSC57 showed error rates <1.0 x 10(-3) errors per site per generation (99.90 100% fidelity) for all the five CGIs. In contrast, AGS showed significantly elevated error rates, mainly due to increased de novo methylation, in three CGIs (1.6- to 3.2-fold), and KATOIII showed a significantly elevated error rate in one CGI (2.2-fold). By selective amplification of fully methylated DNA molecules by methylation-specific PCR, those were stochastically detected in KATOIII and AGS but never in HSC39 and HSC57. When methylation of entire CGIs was examined for eight additional CGIs, KATOIII and AGS had frequent methylation, whereas HSC39 and HSC57 had few. KATOIII and AGS had four and eight times, respectively, as high expression levels of DNMT3B as HSC39. These data showed that some cancer cells have decreased fidelity in replicating methylation patterns in some CGIs, and that the decrease could lead to methylation of the entire CGIs. PMID- 15665275 TI - The dual ErbB1/ErbB2 inhibitor, lapatinib (GW572016), cooperates with tamoxifen to inhibit both cell proliferation- and estrogen-dependent gene expression in antiestrogen-resistant breast cancer. AB - Effective treatment of estrogen receptor (ER)-positive breast cancers with tamoxifen is often curtailed by the development of drug resistance. Antiestrogen resistant breast cancers often show increased expression of the epidermal growth factor receptor family members, ErbB1 and ErbB2. Tamoxifen activates the cyclin dependent kinase inhibitor, p27 to mediate G(1) arrest. ErbB2 or ErbB1 overexpression can abrogate tamoxifen sensitivity in breast cancer lines through both reduction in p27 levels and inhibition of its function. Here we show that the dual ErbB1/ErbB2 inhibitor, lapatinib (GW572016), can restore tamoxifen sensitivity in ER-positive, tamoxifen-resistant breast cancer models. Treatment of MCF-7(pr), T-47D, and ZR-75 cells with lapatinib or tamoxifen alone caused an incomplete cell cycle arrest. Treatment with both drugs led to a more rapid and profound cell cycle arrest in all three lines. Mitogen-activated protein kinase and protein kinase B were inhibited by lapatinib. The two drugs together caused a greater reduction of cyclin D1 and a greater p27 increase and cyclin E-cdk2 inhibition than observed with either drug alone. In addition to inhibiting mitogenic signaling and cell cycle progression, lapatinib inhibited estrogen stimulated ER transcriptional activity and cooperated with tamoxifen to further reduce ER-dependent transcription. Lapatinib in combination with tamoxifen effectively inhibited the growth of tamoxifen-resistant ErbB2 overexpressing MCF 7 mammary tumor xenografts. These data provide strong preclinical data to support clinical trials of ErbB1/ErbB2 inhibitors in combination with tamoxifen in the treatment of human breast cancer. PMID- 15665276 TI - Inactivation of imprinted genes induced by cellular stress and tumorigenesis. AB - Cellular proliferation under stressful conditions may result in permanent genetic and epigenetic changes. Using primary mouse embryonic fibroblasts, we have completed a screening test to identify gene expression changes triggered when cells proliferate under stress. In this manner, we have discovered a novel phenomenon that consists of the rapid and coordinated silencing of genes subject to imprinting, including Cdkn1c, Igf2, H19, Ndn1, Grb10, and Meg3. This generalized silencing of imprinted genes is independent of the stress-responsive tumor suppressors p53, p19(Arf), and p16(Ink4a), and it is also independent of the oxidative culture conditions and the stress response known as "mouse embryonic fibroblast senescence". In the case of Cdkn1c and H19, their silencing is associated with unscheduled de novo methylation of the normally expressed allele at their corresponding CpG island promoters, thus resulting in biallelic methylation. Finally, we provide evidence for frequent de novo methylation of Cdkn1c in a variety of murine cancer types. Altogether, our data support the concept that silencing of imprinted genes, including methylation of Cdkn1c, constitutes an epigenetic signature of cellular stress and tumorigenesis. PMID- 15665277 TI - High-density single nucleotide polymorphism array defines novel stage and location-dependent allelic imbalances in human bladder tumors. AB - Bladder cancer is a common disease characterized by multiple recurrences and an invasive disease course in more than 10% of patients. It is of monoclonal or oligoclonal origin and genomic instability has been shown at certain loci. We used a 10,000 single nucleotide polymorphism (SNP) array with an average of 2,700 heterozygous SNPs to detect allelic imbalances (AI) in 37 microdissected bladder tumors from 17 patients. Eight tumors represented upstaging from Ta to T1, eight from T1 to T2+, and one from Ta to T2+. The AI was strongly stage-dependent as four chromosomal arms showed AI in > 50% of Ta samples, eight in T1, and twenty two in T2+ samples. The tumors showed stage-dependent clonality as 61.3% of AIs were reconfirmed in later T1 tumors and 84.4% in muscle-invasive tumors. Novel unstable chromosomal areas were identified at chromosomes 6q, 10p, 16q, 20p, 20q, and 22q. The tumors separated into two distinct groups, highly stable tumors (all Ta tumors) and unstable tumors (2/3 muscle-invasive). All 11 unstable tumors had lost chromosome 17p areas and 90% chromosome 8 areas affecting Netrin 1/UNC5D/MAP2K4 genes as well as others. AI was present at the TP53 locus in 10 out of 11 unstable tumors, whereas 6 had homozygous TP53 mutations. Tumor distribution pattern reflected AI as seven out of eight patients with additional upper urinary tract tumors had genomic stable bladder tumors (P < 0.05). These data show the power of high-resolution SNP arrays for defining clinically relevant AIs. PMID- 15665279 TI - Xenoestrogen action in prostate cancer: pleiotropic effects dependent on androgen receptor status. AB - Androgen is critical for prostate development, growth, and survival. Therapies for advanced prostate cancer aim to block androgen receptor (AR) action. However, recurrent tumors ultimately arise, which harbor restored AR activity. One mechanism of such reactivation occurs through AR mutations, rendering the receptor responsive to noncanonical ligands. We have shown previously that a known xenoestrogen, bisphenol A (BPA), activates a tumor-derived AR mutant (T877A), leading to androgen-independent prostate cancer cell proliferation. Here, we show that BPA cooperates with androgen to activate AR-T877A as shown by both reporter assays and increased levels of prostate-specific antigen expression. Further investigations using both yeast and mammalian model systems revealed that multiple AR alleles are responsive to BPA, thus expanding the potential influence of xenoestrogens on prostate cancer. Moreover, in vitro radioligand binding assay revealed that BPA alters 5alpha-dihydrotestosterone binding to AR-T877A likely through noncompetitive inhibition. We also show that higher concentrations of BPA block proliferation of AR-positive, androgen dependent prostate adenocarcinoma cells (LNCaP and LAPC-4), with a more modest inhibitory effect on androgen-independent cells (22Rv-1). By contrast, AR negative prostate cancer cells failed to show growth inhibition after exposure to high BPA dose. Together, these data show that BPA can serve as a potential "hormone sensitizer" of the mutant ARs present in advanced prostate adenocarcinomas, thereby possibly contributing toward therapeutic relapse in advanced prostate cancer patients and supporting the notion that nonsteroidal environmental compounds can alter the function of nuclear receptor complexes. PMID- 15665278 TI - A functional common polymorphism in a Sp1 recognition site of the epidermal growth factor receptor gene promoter. AB - The epidermal growth factor receptor (EGFR) plays a prominent role in cell growth and development. Its regulation in humans is complex and incompletely understood. In this study, 12 new polymorphisms were discovered in the 5'-regulatory region of EGFR gene and 2 common single nucleotide polymorphisms (-216G/T and -191C/A) were found in the essential promoter area, one of which is located in a Sp1 recognition site (-216). Transient transfection in human cancer and primary cell lines showed significantly different promoter activity between the two most common haplotypes (-216G-191C and -216T-191C). The replacement of G by T at position -216 increases the promoter activity by 30%. A transient transfection assay in the Sp1-deficient cell line (Schneider cell line 2) showed a strong dependence of EGFR promoter activity on Sp1 and confirmed the effect of the aforementioned polymorphisms. Electrophoretic mobility shift assay also showed a significantly higher binding efficiency of nuclear protein or pure Sp1 protein to the T allele compared with the G allele. We then investigated the allelic imbalance of EGFR transcription in fibroblast cell lines with heterozygous genotype at -216G/T but C/C homozygous genotype at -191C/A. The expression of mRNA carrying T-C haplotype was significantly stronger compared with that of G-C haplotype (P < 0.02). Thus, we successfully showed that a common polymorphism in the EGFR promoter was associated with altered promoter activity and gene expression both in vitro and in vivo. Our findings have implications for cancer etiology and therapy and may also be relevant to the inherited susceptibility of other common diseases. PMID- 15665280 TI - Loss of heterozygosity analysis and DNA copy number measurement on 8p in bladder cancer reveals two mechanisms of allelic loss. AB - Many epithelial tumors show deletion of the short arm of chromosome 8 that is related to aggressive disease or adverse prognosis. In undissected samples of urothelial cell carcinoma of the bladder, at least two regions of loss of heterozygosity (LOH) were identified previously within a small region of 8p11 p12. LOH analysis on a panel of pure tumor DNA samples confirmed this and identified tumors with allelic imbalance, some with clear breakpoints in 8p12. This suggests either that these samples contained genetically distinct subclones or that breakpoints in 8p12 may confer a selective advantage without LOH. To assess the mechanism of LOH and to map breakpoints precisely, a panel of bladder cancer cell lines was examined. Microsatellite analysis of 8p markers identified regions of contiguous homozygosity that coincided with regions of LOH in tumors. Fluorescence in situ hybridization analysis was carried out on seven cell lines predicted to have 8p LOH using a chromosome 8 paint, a chromosome 8 centromeric probe, and a series of single-copy genomic probes. This revealed overall underrepresentation of 8p and overrepresentation of 8q. Several breakpoints and one interstitial deletion were identified in 8p12. Two cell lines with extensive interstitial regions of homozygosity showed no reduction in DNA copy number by fluorescence in situ hybridization analysis, indicating that, in addition to large deletions and rearrangements of 8p, small regions of interstitial LOH on 8p12 may be generated by mitotic recombination. This implicates both major DNA double-strand break repair mechanisms in the generation of 8p alterations. PMID- 15665281 TI - Expression analysis of juvenile pilocytic astrocytomas by oligonucleotide microarray reveals two potential subgroups. AB - Juvenile pilocytic astrocytoma (JPA) is one of the most common brain tumors in children. The expression profiles of 21 JPAs, determined using Affymetrix GeneChip U133A, were compared with subjects with normal cerebella. The genes involved in neurogenesis, cell adhesion, synaptic transmission, central nervous system development, potassium ion transport, protein dephosphorylation, and cell differentiation were found to be significantly deregulated in JPA. These 21 JPAs were further clustered into two major groups by unsupervised hierarchical clustering using a set of 848 genes with high covariance (0.5-10). Supervised analysis with Significance Analysis of Microarrays software between these two potential subgroups identified a list of significant differentially expressed genes involved in cell adhesion, regulation of cell growth, cell motility, nerve ensheathment, and angiogenesis. Immunostaining of myelin basic protein on paraffin sections derived from 18 incompletely resected JPAs suggests that JPA without myelin basic protein-positively stained tumor cells may have a higher tendency to progress. PMID- 15665282 TI - Heterozygosity for p53 (Trp53+/-) accelerates epithelial tumor formation in fanconi anemia complementation group D2 (Fancd2) knockout mice. AB - Fanconi anemia (FA) is an autosomal recessive disease characterized by progressive bone marrow failure and an increased susceptibility to cancer. FA is genetically heterogeneous, consisting of at least 11 complementation groups, FA-A through L, including FA-D1 (BRCA2) and D2. We have previously reported an increased incidence of epithelial tumors in Fancd2 knockout mice. To further investigate the role of the FA pathway in tumor prevention, Fancd2 mutant mice were crossed to mice with a null mutation in the tumor suppressor gene, Trp53. The tumor spectrum in Fancd2(-/-)/Trp53(+/-) mice included sarcomas expected in Trp53 heterozygotes, as well as mammary and lung adenocarcinomas that occur rarely in Trp53 heterozygotes. These tumors occurred earlier than in Fancd2(-/-) control mice. Therefore, the Fancd2(-/-)/Trp53(+/-) mice represent an improved model for the study of adenocarcinoma in FA. In addition, it was found that Fancd2(-/-) mouse embryonic fibroblasts but not Fancd2(-/-)/Trp53(-/-) mouse embryonic fibroblasts arrest following DNA damage. Therefore, Trp53 is required for the S phase checkpoint activation observed in Fancd2 mutant cells. Fancd2(-/ )/Trp53(-/-) cells showed an increase in aneuploidy and had multiple gross chromosomal rearrangements. PMID- 15665283 TI - Tumor susceptibility of Rassf1a knockout mice. AB - The human Ras association domain family 1 (RASSF1) gene is located at 3p21.3 in an area that is believed to harbor at least one important tumor suppressor gene. The two major isoforms of RASSF1, RASSF1A and RASSF1C, are distinguished by alternative NH(2)-terminal exons and the two transcripts initiate in two separate CpG islands. RASSF1A is one of the most frequently inactivated genes described thus far in human solid tumors. Inactivation of RASSF1A most commonly involves methylation of the promoter and CpG island associated with the RASSF1A isoform. In contrast, RASSF1C is almost never inactivated in tumors. Here, we have derived Rassf1a knockout mice in which exon 1-alpha of the Rassf1 gene was deleted, leading to specific loss of Rassf1a but not Rassf1c transcripts. Rassf1a-targeted mice were viable and fertile. Rassf1a(-/-) mice were prone to spontaneous tumorigenesis in advanced age (18-20 months). Whereas only two tumors developed in 48 wild-type mice, six tumors were found in 35 Rassf1a(+/-) mice (P < 0.05) and thirteen tumors were found in 41 Rassf1a(-/-) mice (P < 0.001). The tumors in Rassf1a-targeted mice included lung adenomas, lymphomas, and one breast adenocarcinoma. Rassf1a(-/-) and wild-type mice were treated with two chemical carcinogens, benzo(a)pyrene and urethane, to induce skin tumors and lung tumors, respectively. Rassf1a(-/-) and Rassf1a(+/-) mice showed increased tumor multiplicity and tumor size relative to control animals. The data are consistent with the tumor-suppressive role of Rassf1a, which may explain its frequent epigenetic inactivation in human tumors. PMID- 15665284 TI - Expression-based discovery of variation in the human glutathione S-transferase M3 promoter and functional analysis in a glioma cell line using allele-specific chromatin immunoprecipitation. AB - Discovery and functional evaluation of biologically significant regulatory single nucleotide polymorphisms (SNP) in carcinogen metabolism genes is a difficult challenge because the phenotypic consequences may be both transient and subtle. We have used a gene expression screening approach to identify a functional regulatory SNP in glutathione S-transferase M3 (GSTM3). Anttila et al. proposed that variation in GSTM3 expression was affected by exposure to cigarette smoke and inheritance of the GSTM1-null genotype. To investigate the mechanism of GSTM3 expression was affected by exposure to cigarette smoke and inheritance of the GSTM1-null genotype. To investigate the mechanism of GSTM3 expression variation, we measured GSTM3 expression in lymphoblast cells from a human Centre d'Etude du Polymorphisme Humain family and observed a low expression phenotype. Promoter sequencing revealed two novel GSTM3 promoter SNPs: A/C and A/G SNPs, 63 and 783 bp upstream of the codon 1 start site, respectively. In this pedigree, the two children homozygous for the -63C/C genotype had 8-fold lower GSTM3 expression relative to the two children with the -63A/A genotype, with no association between A-783G SNP and GSTM3 expression. Further evaluation using genotyped glioma cell lines and with luciferase reporter constructs showed that the -63C allele was associated with lower GSTM3 expression (P < 0.0001 and P < 0.003). RNA pol II chromatin immunoprecipitation was combined with quantitative probed-based allelic discrimination genotyping to provide direct evidence of a 9-fold reduced RNA pol II binding capacity for the -63C allele. These results show that the GSTM3 -63C allele strongly affects gene expression in human cell lines and suggests that individuals who carry the low expression allele may be deficient in glutathione transferase catalyzed biological functions. PMID- 15665285 TI - Down-regulation of RAB6KIFL/KIF20A, a kinesin involved with membrane trafficking of discs large homologue 5, can attenuate growth of pancreatic cancer cell. AB - To identify novel molecular targets for treatment of pancreatic ductal adenocarcinoma (PDAC), we generated precise gene expression profiles of PDACs on a genome-wide cDNA microarray after populations of tumor cells were purified by laser microdissection. Through functional analysis of genes that were transactivated in PDACs, we identified RAB6KIFL as a candidate for development of drugs to treat PDACs at the molecular level. Knockdown of endogenous RAB6KIFL expression in PDAC cell lines by small interfering RNA drastically attenuated growth of those cells, suggesting an essential role for the gene product in maintaining viability of PDAC cells. RAB6KIFL belongs to the kinesin superfamily of motor proteins, which have critical functions in trafficking of molecules and organelles. Proteomics analyses using a polyclonal anti-RAB6KIFL antibody identified one of the cargoes transported by RAB6KIFL as discs, large homologue 5 (DLG5), a scaffolding protein that may link the vinexin-beta-catenin complex at sites of cell-cell contact. Like RAB6KIFL, DLG5 was overexpressed in PDACs, and knockdown of endogenous DLG5 by small interfering RNA significantly suppressed the growth of PDAC cells as well. Decreased levels of endogenous RAB6KIFL in PDAC cells altered the subcellular localization of DLG5 from cytoplasmic membranes to cytoplasm. Our results imply that collaboration of RAB6KIFL and DLG5 is likely to be involved in pancreatic carcinogenesis. These molecules should be promising targets for development of new therapeutic strategies for PDACs. PMID- 15665286 TI - Particle irradiation suppresses metastatic potential of cancer cells. AB - Particle radiotherapy such as proton and carbon ion has been producing promising clinical results worldwide. The purpose of this study was to compare metastatic capabilities of malignant tumor cells after irradiation with photon, proton, and carbon ion beams to clarify their ion beam-specific biological effects. We examined the biological properties of highly aggressive HT1080 human fibrosarcoma cells to assess their metastatic processes in terms of cell adhesion capability to extracellular matrix, expression of integrins, cell migration, cell invasive capability, and matrix metalloproteinase-2 activity in vitro. We then assessed the metastatic capabilities of LM8 mouse osteosarcoma irradiated with carbon ion or photon beam in the syngeneic mice. Both proton and carbon ion irradiation decreased cell migration and invasion in a dose-dependent manner and strongly inhibited matrix metalloproteinase-2 activity. On the other hand, lower X-ray irradiation promoted cell migration and invasion concomitant with up-regulation of alphaVbeta3 integrin. For cancer cells treated with carbon ion irradiation, the number of pulmonary metastasis was decreased significantly in vivo. These findings suggest that particle irradiation suppresses metastatic potential even at lower dose, whereas photon irradiation promotes cell migration and invasive capabilities at lower dose level, and provide preclinical evidence that ion beam radiotherapy may be superior to conventional photon beam therapy in possible preventive effects on metastases of irradiated malignant tumor cells. PMID- 15665287 TI - Flat dysplastic aberrant crypt foci are related to tumorigenesis in the colon of azoxymethane-treated rat. AB - We evaluated the role of aberrant crypt foci (ACF) as biomarkers of colon cancer by studying the sequential development (6-28 weeks) from early lesion to tumor in the colon of azoxymethane-exposed F344 rats (15 mg/kg bw x 2). Surface examination of unsectioned methylene blue-stained colon preparations, transilluminated in the inverse light microscope, revealed two types of early lesions: classic elevated ACF and small flat lesions, which we denoted flat ACF and which were characterized by bright blue staining, compressed crypt openings, and crypts not elevated above the surrounding mucosa. At a later stage, the crypts surrounding large flat ACF became enlarged, a change that slightly raised the structure; principally, large flat ACF and nascent tumors displayed the same surface morphology. Furthermore, flat ACF with 18.6 +/- 10.6 crypt/focus and tumors showed a uniform picture of severe dysplasia with frequent presence of Paneth cells, compressed crypts, cytoplasmic/nuclear overexpression of beta catenin, and nuclear overexpression of cyclin D1. In contrast, classic elevated ACF with 5.3 +/- 2.5 crypts/focus did not display such changes: they showed mainly hyperplasia, mild or moderate dysplasia but never severe dysplasia. Along the time course, the number of flat ACF + tumors, including microscopic and macroscopic, was virtually constant, approximately 2.5 lesions/rat. The number of classic elevated ACF was initially approximately 180 lesions/rat and terminally approximately 80 lesions/rat. Flat ACF grew significantly faster than classic elevated ACF. In conclusion, our data indicate a continuous developmental growth from small flat dysplastic ACF to the stage of a tumor. In contrast, classic elevated ACF do not seem to be as closely related to tumorigenesis. PMID- 15665289 TI - Thymosin {beta}(10) inhibits angiogenesis and tumor growth by interfering with Ras function. AB - Thymosin beta(10) is a monomeric actin sequestering protein that regulates actin dynamics. Previously, we and others have shown that thymosin beta(10) acts as an actin-mediated tumor suppressor. In this study, we show that thymosin beta(10) is not only a cytoskeletal regulator, but that it also acts as a potent inhibitor of angiogenesis and tumor growth by its interaction with Ras. We found that overexpressed thymosin beta(10) significantly inhibited vascular endothelial growth factor-induced endothelial cell proliferation, migration, invasion, and tube formation in vitro. Vessel sprouting was also inhibited ex vivo. We further show that thymosin beta(10) directly interacted with Ras. This interaction resulted in inhibition of the Ras downstream mitogen-activated protein kinase/extracellular signal-regulated kinase kinase signaling pathway, leading to decreased vascular endothelial growth factor production. Thymosin beta(10) injected into a xenograft model of human ovarian cancer in nude mice markedly inhibited tumor growth and reduced tumor vascularity. In contrast, a related thymosin family member, thymosin beta(4), did not bind to Ras and showed positive effects on angiogenesis. These findings show that the inhibition of Ras signal transduction by thymosin beta(10) results in antiangiogenic and antitumor effects, suggesting that thymosin beta(10) may be valuable in anticancer therapy. PMID- 15665288 TI - Matrix metalloproteinase-2 contributes to cancer cell migration on collagen. AB - Matrix metalloproteinases (MMP) are central to tissue penetration by cancer cells, as tumors expand and form metastases, but the mechanism by which MMP-2 contributes to cancer cell migration is not well understood. In the present experiments, both a broad-spectrum MMP inhibitor and the isolated collagen binding domain (CBD) from MMP-2 inhibited cell migration on native type I collagen. These results verified the involvement of MMPs in general and showed that MMP-2, specifically, contributes to cell migration by a mechanism involving MMP-2 interaction with collagen. To exclude potential overlapping effects of MMP 9, additional experiments showed that MMP-2 also contributed to migration of MMP 9-/- cells. To investigate whether the homologous CBD from human fibronectin also inhibited cell migration, we first showed that fragmentation of fibronectin is a feature of breast cancer tumors and that several fragments contained the CBD. However, the recombinant fibronectin domain did not alter cell migration on collagen. This lack of effect on cell migration was explored in competitive protein-protein binding assays, which showed that the affinity of MMP-2 for collagen exceeds that of fibronectin. Furthermore, whereas the isolated MMP-2 CBD inhibited the gelatinolytic activities of MMP-2 and tumor extracts, such an inhibition was not characteristic of the corresponding fibronectin domain. Together, our results provide evidence that MMP-2 is an important determinant of cancer cell behavior but is not inhibited by the collagen binding segment of fibronectin. PMID- 15665290 TI - Anastellin, a fragment of the first type III repeat of fibronectin, inhibits extracellular signal-regulated kinase and causes G(1) arrest in human microvessel endothelial cells. AB - The formation of a microvascular endothelium plays a critical role in the growth and metastasis of established tumors. The ability of a fragment from the first type III repeat of fibronectin (III(1C)), anastellin, to suppress tumor growth and metastasis in vivo has been reported to be related to its antiangiogenic properties, however, the mechanism of action of anastellin remains unknown. Utilizing cultures of human dermal microvascular endothelial cells, we provide evidence that anastellin inhibits signaling pathways which regulate the extracellular signal-regulated (ERK) mitogen-activated protein kinase pathway and subsequent expression of cell cycle regulatory proteins. Addition of anastellin to primary microvascular endothelial cells resulted in a complete inhibition of serum-dependent proliferation. Growth inhibition correlated with a decrease in serum-dependent expression of cyclin D1, cyclin A and the cyclin-dependent kinase, cdk4, key regulators of cell cycle progression through G(1) phase. Consistent with a block in G(1)-S transition, anastellin inhibited serum dependent incorporation of [(3)H]-thymidine into S-phase nuclei. Addition of anastellin to serum-starved microvessel cells resulted in a time-dependent and dose-dependent decrease in basal levels of phosphorylated MEK/ERK and blocked serum-dependent activation of ERK. Adenoviral infection with Ad.DeltaB-Raf:ER, an inducible estrogen receptor-B-Raf fusion protein, restored levels of active ERK in anastellin-treated cells, rescued levels of cyclin D1, cyclin A, and cdk4, and rescued [(3)H]-thymidine incorporation. These data suggest that the antiangiogenic properties of anastellin observed in mouse models of human cancer may be due to its ability to block endothelial cell proliferation by modulating ERK signaling pathways and down-regulating cell cycle regulatory gene expression required for G(1)-S phase progression. PMID- 15665291 TI - Acceleration of Smad2 and Smad3 phosphorylation via c-Jun NH(2)-terminal kinase during human colorectal carcinogenesis. AB - Conversion of normal epithelial cells to tumors is associated with a shift in transforming growth factor-beta (TGF-beta) function: reduction of tumor suppressor activity and increase of oncogenic activity. However, specific mechanisms of this functional alteration during human colorectal carcinogenesis remain to be elucidated. TGF-beta signaling involves Smad2/3 phosphorylated at linker regions (pSmad2/3L) and COOH-terminal regions (pSmad2/3C). Using antibodies specific to each phosphorylation site, we herein showed that Smad2 and Smad3 were phosphorylated at COOH-terminal regions but not at linker regions in normal colorectal epithelial cells and that pSmad2/3C were located predominantly in their nuclei. However, the linker regions of Smad2 and Smad3 were phosphorylated in 31 sporadic colorectal adenocarcinomas. In particular, late stage invasive and metastatic cancers typically showed a high degree of phosphorylation of Smad2/3L. Their extent of phosphorylation in 11 adenomas was intermediate between those in normal epithelial cells and adenocarcinomas. Whereas pSmad2L remained in the cytoplasm, pSmad3L was located exclusively in the nuclei of Ki-67-immunoreactive adenocarcinomas. In contrast, pSmad3C gradually decreased as the tumor stage progressed. Activated c-Jun NH(2)-terminal kinase in cancers could directly phosphorylate Smad2/3L. Although Mad homology 2 region sequencing in the Smad4 gene revealed a G/A substitution at codon 361 in one adenocarcinoma, the mutation did not correlate with phosphorylation. No mutations in the type II TGF-beta receptor and Smad2 genes were observed in the tumors. In conclusion, pSmad3C, which favors tumor suppressor activity of TGF-beta, was found to decrease, whereas c-Jun NH(2)-terminal kinase tended to induce the phosphorylation of Smad2/3L in human colorectal adenoma-carcinoma sequence. PMID- 15665292 TI - Transcriptional profiles of intestinal tumors in Apc(Min) mice are unique from those of embryonic intestine and identify novel gene targets dysregulated in human colorectal tumors. AB - The adenomatous polyposis coli (APC) tumor suppressor is a major regulator of the Wnt signaling pathway in normal intestinal epithelium. APC, in conjunction with AXIN and GSK-3beta, forms a complex necessary for the degradation of beta catenin, thereby preventing beta-catenin/T-cell factor interaction and alteration of growth-controlling genes such as c-MYC and cyclin D1. Inappropriate activation of the Wnt pathway, via Apc/APC mutation, leads to gastrointestinal tumor formation in both the mouse and human. In order to discover novel genes that may contribute to tumor progression in the gastrointestinal tract, we used cDNA microarrays to identify 114 genes with altered levels of expression in Apc(Min) mouse adenomas from the duodenum, jejunum, and colon. Changes in the expression of 24 of these 114 genes were not observed during mouse development at embryonic day 16.5, postnatal day 1, or postnatal day 14 (relative to normal adult intestine). These 24 genes are not previously known Wnt targets. Seven genes were validated by real-time reverse transcription-PCR analysis, whereas four genes were validated by in situ hybridization to mouse adenomas. Real-time reverse transcription-PCR analysis of human colorectal cancer cell lines and adenocarcinomas revealed that altered expression levels were also observed for six of the genes Igfbp5, Lcn2, Ly6d, N4wbp4 (PMEPA1), S100c, and Sox4. PMID- 15665293 TI - Glycolytic enzymes can modulate cellular life span. AB - An unbiased screen for genes that can immortalize mouse embryonic fibroblasts identified the glycolytic enzyme phosphoglycerate mutase (PGM). A 2-fold increase in PGM activity enhances glycolytic flux, allows indefinite proliferation, and renders cells resistant to ras-induced arrest. Glucosephosphate isomerase, another glycolytic enzyme, displays similar activity and, conversely, depletion of PGM or glucosephosphate isomerase with short interfering RNA triggers premature senescence. Immortalized mouse embryonic fibroblasts and mouse embryonic stem cells display higher glycolytic flux and more resistance to oxidative damage than senescent cells. Because wild-type p53 down-regulates PGM, mutation of p53 can facilitate immortalization via effects on PGM levels and glycolysis. PMID- 15665294 TI - Wavelet transformations of tumor expression profiles reveals a pervasive genome wide imprinting of aneuploidy on the cancer transcriptome. AB - Aneuploidy is frequently observed in many human cancers, but its global effects on the cancer transcriptome are controversial. We did a systematic and unbiased genome-wide survey to determine the extent a tumor's abnormal karyotype (chromosomal amplifications and deletions) is detectably "imprinted" onto that tumor's gene expression profile. By using a novel methodology employing wavelet transform signal-processing algorithms to identify genomic regions of coordinated gene expression (wavelet variance scanning), we analyzed a series of gastric cancer cell lines and identified >100 genomic regions exhibiting distinct patterns of subtle but significant coordinated transcription, ranging from tens to hundreds of genes. A large majority (80%) of these regions could be specifically localized to a site of detectable genomic amplification or deletion; reciprocally, up to 47% of the total aneuploidy in each of the individual cell lines could be directly inferred from the gene expression data. Genome-wide portraits of tumor aneuploidy can thus be successfully reconstructed solely from gene expression data, implying that the effects of aneuploidy must be pervasively and globally imprinted within the cancer transcriptome. Aneuploidy may contribute to tumor behavior not just by affecting the expression of a few key oncogenes and tumor suppressor genes but also by subtly altering the expression levels of hundreds of genes in the oncogenome. PMID- 15665295 TI - Implication of STAT3 signaling in human colonic cancer cells during intestinal trefoil factor 3 (TFF3) -- and vascular endothelial growth factor-mediated cellular invasion and tumor growth. AB - Signal transducer and activator of transcription (STAT) 3 is overexpressed or activated in most types of human tumors and has been classified as an oncogene. In the present study, we investigated the contribution of the STAT3s to the proinvasive activity of trefoil factors (TFF) and vascular endothelial growth factor (VEGF) in human colorectal cancer cells HCT8/S11 expressing VEGF receptors. Both intestinal trefoil peptide (TFF3) and VEGF, but not pS2 (TFF1), activate STAT3 signaling through Tyr(705) phosphorylation of both STAT3alpha and STAT3beta isoforms. Blockade of STAT3 signaling by STAT3beta, depletion of the STAT3alpha/beta isoforms by RNA interference, and pharmacologic inhibition of STAT3alpha/beta phosphorylation by cucurbitacin or STAT3 inhibitory peptide abrogates TFF- and VEGF-induced cellular invasion and reduces the growth of HCT8/S11 tumor xenografts in athymic mice. Differential gene expression analysis using DNA microarrays revealed that overexpression of STAT3beta down-regulates the VEGF receptors Flt-1, neuropilins 1 and 2, and the inhibitor of DNA binding/differentiation (Id-2) gene product involved in the neoplastic transformation. Taken together, our data suggest that TFF3 and the essential tumor angiogenesis regulator VEGF(165) exert potent proinvasive activity through STAT3 signaling in human colorectal cancer cells. We also validate new therapeutic strategies targeting STAT3 signaling by pharmacologic inhibitors and RNA interference for the treatment of colorectal cancer patients. PMID- 15665296 TI - A mutation in the SDHC gene of complex II increases oxidative stress, resulting in apoptosis and tumorigenesis. AB - Intracellular oxidative stress from mitochondria is thought to be important in carcinogenesis and tumorigenesis, but direct experimental proof is limited. In this study, a transgenic mouse cell line (SDHC E69) with a mutated SDHC gene (a subunit of complex II in the electron transport chain) was constructed to test this question. The SDHC E69 cells overproduced superoxide anion (O(2)(-)) from mitochondria, had elevated cytoplasmic carbonyl proteins and 8-OH-deoxyguanine in their DNA as well as significantly higher mutation frequencies than wild type. There were many apoptotic cells in this cell line, as predicted by the observed increase in caspase 3 activity, decrease in mitochondrial membrane potential, and structural changes in their mitochondria. In addition, some cells that escaped from apoptosis underwent transformation, as evidenced by the fact that SDHC E69 cells caused benign tumors when injected under the epithelium of nude mice. These results underscore the notion that mitochondrially generated oxidative stress can contribute to nuclear DNA damage, mutagenesis, and ultimately, tumorigenesis. PMID- 15665297 TI - cIAP1 Localizes to the nuclear compartment and modulates the cell cycle. AB - We explored the location and function of the human cIAP1 protein, a member of the inhibitor of apoptosis protein (IAP) family. Unlike family member X-linked IAP (XIAP), which was predominantly cytoplasmic, the cIAP1 protein localized almost exclusively to nuclei in cells, as determined by immunofluorescence microscopy and subcellular fractionation methods. Interestingly, apoptotic stimuli induced nuclear export of cIAP1, which was blocked by a chemical caspase inhibitor. In dividing cells, cIAP1 was released into the cytosol early in mitosis, then reaccumulated in nuclei in late anaphase and in telophase, with the exception of a pool of cIAP1 that associated with the midbody. Survivin, another IAP family member, and cIAP1 were both localized on midbody microtubules at telophase, and also interacted with each other during mitosis. Cells stably overexpressing cIAP1 accumulated in G(2)-M phase and grew slower than control-transfected cells. These cIAP1-overexpressing cells also exhibited cytokinesis defects over 10 times more often than control cells and displayed a mitotic checkpoint abnormality with production of polyploid cells when exposed to microtubule-targeting drugs nocodazole and paclitaxel (Taxol). Our findings demonstrate a role for overexpressed cIAP1 in genetic instability, possibly by interfering with mitotic functions of Survivin. These findings may have important implications for cancers in which cIAP1 overexpression occurs. PMID- 15665298 TI - Identification of brain-derived neurotrophic factor as a novel functional protein in hepatocellular carcinoma. AB - This study aims to identify a novel molecule that may contribute to hepatocarcinogenesis in a rat orthotopic hepatocellular carcinoma model. The hepatocellular carcinoma model was generated by injection of tumor cells into the left lobe of the liver. Proteomic approaches, including ProteinChip and two dimensional electrophoresis, were used to identify proteins from serially collected rat serum samples. By both ProteinChip and two-dimensional electrophoresis techniques, the level of a 27-kDa protein was found to be augmented in serum samples during tumor development, decreased after left lobectomy, and reincreased at the time of tumor recurrence. The protein was identified to be brain-derived neurotrophic factor (BDNF). By using specific primers and monoclonal antibody, the expression pattern of BDNF was confirmed in tumor tissue but not in the adjacent nontumorous liver tissue. In addition, the truncated isoform of BDNF receptor-tyrosine protein kinase receptor B was only found in tumor tissue. An in vitro study showed that exogenous BDNF could induce tumor cell proliferation predominantly in relatively small numbers of inoculated cells. Administration of BDNF to tumor cell lines induced significantly increased expression of heat shock protein 90 (Hsp90) and cyclin D1, and blocking the activity of Hsp90 could reverse the up-regulation of cyclin D1 induced by BDNF. The present study revealed that BDNF and its receptor were uniquely expressed in tumor tissue and cell lines of hepatocellular carcimona but not in nontumorous liver tissue and normal cell line. BDNF could stimulate tumor cell proliferation in a Hsp90-dependent manner. PMID- 15665299 TI - Aberrant epidermal growth factor receptor signaling and enhanced sensitivity to EGFR inhibitors in lung cancer. AB - Epidermal growth factor receptor (EGFR) is occasionally amplified and/or mutated in non-small cell lung cancer (NSCLC) and can be coexpressed with other members of the HER receptor family to form functional heterodimers. We therefore investigated lung cancer cell lines for alterations in EGFR gene copy number, enhanced expression of EGFR and other HER family members, and EGFR coding sequence mutations and correlated these findings with response to treatment with the EGFR inhibitors and the kinetics of ligand-induced signaling. We show here that somatic deletions in the tyrosine kinase domain of EGFR were associated with increased EGFR gene copy number in NSCLC. Treatment with the specific EGFR tyrosine kinase inhibitors (TKI) gefitinib or erlotinib or the EGFR inhibitory antibody cetuximab induced apoptosis of HCC827, a NSCLC cell line with EGFR gene amplification and an exon 19 deletion. H1819, a NSCLC cell line that expresses high levels of EGFR, ErbB2, and ErbB3 but has wild-type EGFR, showed intermediate sensitivity to TKIs. In both cell lines, ligand-induced receptor tyrosine phosphorylation was delayed and prolonged and AKT was constitutively phosphorylated (but remained inhibitable by EGFR TKI). Thus, in addition to EGFR mutations, other factors in NSCLC cells, such as high expression of ErbB family members, may constitutively activate AKT and sensitize cells to EGFR inhibitors. PMID- 15665300 TI - Glioma formation in neurofibromatosis 1 reflects preferential activation of K-RAS in astrocytes. AB - Children with the tumor predisposition syndrome, neurofibromatosis 1 (NF1), develop optic pathway gliomas. The NF1 gene product, neurofibromin, functions as a negative regulator of RAS, such that NF1 inactivation results in RAS hyperactivation. Recent studies have highlighted the divergent biological and biochemical properties of the various RAS isoforms, which prompted us to examine the consequence of Nf1 inactivation in astrocytes on RAS isoform activation in vitro and in vivo. In this report, we show that only K-RAS is activated in Nf1-/- astrocytes and that activation of K-RAS, but not H-RAS, accounts for the proliferative advantage and abnormal actin cytoskeleton-mediated processes observed in Nf1-/- astrocytes in vitro. Moreover, dominant inhibitory K-RAS corrects these abnormalities in Nf1-/- astrocytes invitro. Lastly, we show that Nf1+/- mice with astrocyte-specific activated K-RAS expression in vivo develop optic pathway gliomas, similar to our previously reported Nf1+/- mice with astrocyte Nf1 inactivation. Collectively, our results show that K-RAS is the primary target for neurofibromin GTPase-activating protein activity in vitro and in vivo and that K-RAS activation in astrocytes recapitulates the biochemical, biological, and tumorigenic properties of neurofibromin loss. PMID- 15665301 TI - Perinucleolar compartment prevalence has an independent prognostic value for breast cancer. AB - The perinucleolar compartment (PNC) is a multicomponent nuclear structure enriched with RNAs transcribed by RNA pol III and RNA binding proteins. Studies in cultured cells showed an association between PNC and transformed phenotype. To evaluate the relationship between structure and malignancy in vivo, we examined PNC prevalence (the percentage of cells containing at least one PNC) in normal and cancerous paraffin-embedded breast tissues using immunohistochemistry against a PNC-associated protein. Five hundred nuclei in the most active area of each sample were scored for PNC prevalence. The results show that PNC prevalence significantly correlates with the progression of breast cancer (by the criteria of staging). PNC prevalence in primary tumors, lymph nodes, and distant metastases shows a stepwise increase from a median of 23% in primary tumors to approximately 100% in distant metastases. In addition, univariate and multivariate (controlling for tumor size and grade) analyses show that early stage patients with invasive ductal carcinomas containing a higher PNC prevalence have a significantly poorer prognosis. These findings link PNC prevalence with the progression of breast cancer in vivo and suggest that PNC-containing cells have metastatic advantages. These findings also show the potential of PNC prevalence as a prognostic marker for breast cancer. PMID- 15665302 TI - Gene transfer-mediated pre-mRNA segmental trans-splicing as a strategy to deliver intracellular toxins for cancer therapy. AB - Virus-mediated transfer of genes coding for intracellular toxins holds promise for cancer therapy, but the inherent toxicity of such vectors make them a risk to normal tissues and a challenge to produce due to the intrinsic dilemma that expression of toxin molecules kills producer cells. We employed pre-mRNA segmental trans-splicing (STS), in which two engineered DNA fragments coding for 5' "donor" and 3' "acceptor" segments of a toxin gene, respectively, are expressed by viral vectors. When co-delivered to target cells, the two vectors generate two toxin pre-mRNA fragments which are spliced by the target cell machinery to produce functional mRNA and toxin. To test this approach, we used an enzymatic fragment of Shigatoxin1A1 (STX1A1) known to provoke apoptotic cell death. Two adenovirus vectors, Shigatoxin1A1 donor (AdStx1A1Do) and Shigatoxin1A1 acceptor (AdStx1A1Ac), respectively, were used to deliver the Stx1A1 gene fragments. HeLa, HEp2, and A549 cells transfected with AdStx1A1Do and AdStx1A1Ac had a dose-dependent reduction in viability and inhibition of protein synthesis. Intratumoral injection of AdStx1A1Do and AdStx1A1Ac into preexisting HeLa, Hep2, and A549 tumors in immunodeficient mice revealed significant inhibition of tumor growth. There was no evidence of liver damage, suggesting that there was no leakage of vector or toxin from the site of injection following intratumoral injection of AdStx1A1Do and AdStx1A1Ac. These results suggest that the obstacles preventing gene transfer of intracellular toxins for local cancer therapy could be overcome by pre-mRNA segmental trans-splicing. PMID- 15665303 TI - Rituximab (chimeric anti-CD20 monoclonal antibody) inhibits the constitutive nuclear factor-{kappa}B signaling pathway in non-Hodgkin's lymphoma B-cell lines: role in sensitization to chemotherapeutic drug-induced apoptosis. AB - The chimeric anti-CD20 antibody rituximab (Rituxan, IDEC-C2B8) is widely used in the clinical treatment of patients with non-Hodgkin's lymphoma (NHL). Rituximab sensitizes NHL B-cell lines to drug-induced apoptosis via down-regulation of Bcl x(L) expression. We hypothesized that the mechanism by which rituximab down regulates Bcl-x(L) may be, in part, due to inhibition of constitutive nuclear factor-kappaB (NF-kappaB) activity that regulates Bcl-x(L) expression. This hypothesis was tested in CD20(+) drug-resistant Ramos (Bcl-2(-)/Bcl-x(L)(+)) and Daudi (Bcl-2(+)/Bcl-x(L)(+)) cell lines. Rituximab decreased the phosphorylation of NF-kappaB-inducing kinase, IkappaB kinase, and IkappaB-alpha, diminished IKK kinase activity, and decreased NF-kappaB DNA binding activity. These events occurred with similar kinetics and were observed 3 to 6 hours post-rituximab treatment. Rituximab significantly up-regulated Raf-1 kinase inhibitor protein expression, thus interrupting the NF-kappaB signaling pathway concomitant with Bcl-x(L) and Bfl-1/A1 down-regulation. The role of NF-kappaB in the regulation of Bcl-x(L) transcription was shown using promoter reporter assays in which deletion of the two-tandem NF-kappaB binding sites in the upstream promoter region significantly reduced the luciferase activity. This was further corroborated by using IkappaB superrepressor cells and by NF-kappaB-specific inhibitors. The direct role of Bcl-x(L) in drug resistance was assessed by using Bcl-x(L) overexpressing cells, which exhibited higher drug resistance that was partially reversed by rituximab. Rituximab-mediated inhibition of the NF-kappaB signaling pathway and chemosensitization was corroborated by the use of specific inhibitors. These findings reveal a novel pathway mediated by rituximab through Raf-1 kinase inhibitor protein induction that negatively regulates the constitutive NF-kappaB pathway and chemosensitization of the NHL B-cells. PMID- 15665304 TI - Attenuation of proteasome-induced proteolysis in skeletal muscle by {beta} hydroxy-{beta}-methylbutyrate in cancer-induced muscle loss. AB - Loss of skeletal muscle is an important determinant of survival in patients with cancer-induced weight loss. The effect of the leucine metabolite beta-hydroxy beta-methylbutyrate (HMB) on the reduction of body weight loss and protein degradation in the MAC16 model of cancer-induced weight loss has been compared with that of eicosapentaenoic acid (EPA), a recognized inhibitor of protein degradation. HMB was found to attenuate the development of weight loss at a dose greater than 0.125 g/kg accompanied by a small reduction in tumor growth rate. When EPA was used at a suboptimal dose level (0.6 g/kg) the combination with HMB seemed to enhance the anticachectic effect. Both treatments caused an increase in the wet weight of soleus muscle and a reduction in protein degradation, although there did not seem to be a synergistic effect of the combination. Proteasome activity, determined by the "chymotrypsin-like" enzyme activity, was attenuated by both HMB and EPA. Protein expression of the 20S alpha or beta subunits was reduced by at least 50%, as were the ATPase subunits MSS1 and p42 of the 19S proteasome regulatory subunit. This was accompanied by a reduction in the expression of E2(14k) ubiquitin-conjugating enzyme. The combination of EPA and HMB was at least as effective or more effective than either treatment alone. Attenuation of proteasome expression was reflected as a reduction in protein degradation in gastrocnemius muscle of cachectic mice treated with HMB. In addition, HMB produced a significant stimulation of protein synthesis in skeletal muscle. These results suggest that HMB preserves lean body mass and attenuates protein degradation through down-regulation of the increased expression of key regulatory components of the ubiquitin-proteasome proteolytic pathway, together with stimulation of protein synthesis. PMID- 15665305 TI - Darbepoietin alfa potentiates the efficacy of radiation therapy in mice with corrected or uncorrected anemia. AB - Darbepoietin alfa (DA) is a long-acting analogue of erythropoietin that has reduced receptor affinity and enhanced biological activity. Experiments were done to test the hypothesis that correction of anemia in tumor-bearing mice by DA would increase tumor oxygenation and potentiate radiation-induced tumor cell killing. A SCC VII tumor model was used to study tumor responses to fractionated radiation therapy in mice with anemia induced by total body irradiation. Administration of DA reduced the extent and duration of anemia and associated tumor hypoxia, protected the bone marrow cells and prevented the body weight loss from the effect of irradiation, and facilitated the recovery in a time-dependent manner, with the administration of DA prior to total body irradiation having the greatest protective effect. When combined with fractionated radiation therapy, DA increased the tumor growth delay time from 2.7 days for irradiation alone to 7.3 to 10.6 days for combination of DA and irradiation. The effect of DA on tumor responses to fractionated radiation therapy was observed when DA was given 18 to 4 days before starting radiation therapy, but DA was also equally effective as a radiosensitizer when given only 2 hours before fractionated irradiation therapy. Weekly dosing of DA was as efficacious for the enhancement of radiation responses of tumors as biweekly dosing. Similar results were obtained in the RIF-1 fibrosarcoma tumor model. These studies show that DA can effectively correct anemia in tumor-bearing mice and sensitize tumor cells to fractionated radiation therapy. Importantly, DA was also able to sensitize tumors to radiation in mice with uncorrected anemia and hypoxia, suggesting that the effect of DA on radiosensitivity was independent of these factors and a different mechanism of action may be responsible for this effect. PMID- 15665306 TI - A genome-wide view of the in vitro response to l-asparaginase in acute lymphoblastic leukemia. AB - To investigate the effect of l-asparaginase on acute lymphoblastic leukemia (ALL), we used cDNA microarrays to obtain a genome-wide view of gene expression both at baseline and after in vitro exposure to l-asparaginase in cell lines and pediatric ALL samples. In 16 cell lines, a baseline gene expression pattern distinguished l-asparaginase sensitivity from resistance. However, for 28 pediatric ALL samples, no consistent baseline expression pattern was associated with sensitivity to l-asparaginase. In particular, baseline expression of asparagine synthetase (ASNS) was not predictive of response to l-asparaginase. After exposure to l-asparaginase, 5 cell lines and 10 clinical samples exhibited very similar changes in the expression of a large number of genes. However, the gene expression changes occurred more slowly in the clinical samples. These changes included a consistent increase in expression of tRNA synthetases and solute transporters and activating transcription factor and CCAAT/enhancer binding protein family members, a response similar to that observed with amino acid starvation. There was also a consistent decrease in many genes associated with proliferation. Taken together, the changes seem to reflect a consistent coordinated response to asparagine starvation in both cell lines and clinical samples. Importantly, in the clinical samples, increased expression of ASNS after l-asparaginase exposure was not associated with in vitro resistance to l asparaginase, indicating that ASNS-independent mechanisms of in vitro l asparaginase resistance are common in ALL. These results suggest that targeting particular genes involved in the response to amino acid starvation in ALL cells may provide a novel way to overcome l-asparaginase resistance. PMID- 15665309 TI - Oncogenic transformation of human mammary epithelial cells by autocrine human growth hormone. AB - The human growth hormone (hGH) gene is expressed in the normal human mammary epithelial cell and its expression increases concomitant with the acquisition of proliferative lesions. Herein we demonstrate that autocrine production of hGH in human mammary carcinoma cells dramatically enhances anchorage-independent growth in a Janus kinase 2-dependent manner. Forced expression of the hGH gene in immortalized human mammary epithelial cells increased proliferation, decreased apoptosis, altered the cellular morphology and resulted in oncogenic transformation. Autocrine hGH was therefore sufficient to support anchorage independent growth of immortalized human mammary epithelial cells and tumor formation in vivo. Moreover, autocrine hGH disrupted normal mammary acinar architecture with luminal filling and deregulated proliferation in three dimensional epithelial cell culture. Autocrine hGH utilized homeobox A1 to govern the transcriptional program required for autocrine hGH-stimulated oncogenic transformation of human mammary epithelial cells, including transcriptional up regulation of c-Myc, cyclin D1, and Bcl-2. Forced expression of a single orthotopically expressed wild-type gene is therefore sufficient for oncogenic transformation of the immortalized human mammary epithelial cell. PMID- 15665307 TI - Androgens repress the expression of the angiogenesis inhibitor thrombospondin-1 in normal and neoplastic prostate. AB - In order to understand why the angiogenesis inhibitor thrombospondin-1 (TSP1) is often, although not always, associated with prostatic tumors, we have investigated its relationship with the testosterone and the vasculature on which both normal and tumorigenic prostatic epithelia depend. In vivo, androgen withdrawal led to increased TSP1 production and decreased vascularization in the normal rat prostate which was reversed by androgen replacement. Androgen repression of TSP1 production occurred at the transcriptional level and was dependent on the presence of the first intron of the TSP1 gene. In an experimental model of prostate tumorigenesis, TSP1, when delivered by admixed stromal fibroblasts, markedly delayed LNCaP tumor growth and limited tumor vascularization. However, prolonged exposure to TSP1 resulted in the growth of tumors secreting high levels of vascular endothelial growth factor in the bloodstream of tumor-bearing animals and tumor growth was no longer sensitive to TSP1 inhibitory effects. Clinical evidence also suggested that prostate carcinomas are able to adapt to escape the antiangiogenic effects of TSP1. In human androgen-dependent localized prostate carcinomas, TSP1 expression was inversely correlated with blood vessel density. Androgen deprivation in patients with hormone-responsive tumors led to increased TSP1 expression and vascular regression. In contrast, despite a sustained expression in the tumor bed, TSP1 was no longer associated with decreased vascularization in hormone-refractory prostate tumors. Overall, these results suggest that the high in situ TSP1 exposure triggered by androgen deprivation in patients with prostate cancer could lead to early tumor resistance. Such patients could benefit from a combination of androgen deprivation and antiangiogenic therapy in order to minimize the induction of such tumor escape. PMID- 15665308 TI - Modification of professional antigen-presenting cells with small interfering RNA in vivo to enhance cancer vaccine potency. AB - RNA interference using small interfering RNA (siRNA) is an effective means of silencing gene expression in cells. Intradermal administration of nucleic acids via gene gun represents an efficient method for delivering nucleic acids to professional antigen-presenting cells in vivo. In this study, we show that the coadministration of DNA vaccines encoding human papillomavirus type 16 E7 with siRNA targeting key proapoptotic proteins Bak and Bax prolongs the lives of antigen-expressing dendritic cells in the draining lymph nodes, enhances antigen specific CD8(+) T-cell responses, and elicits potent antitumor effects against an E7-expressing tumor model in vaccinated mice. Our data indicate that intradermal administration of siRNA to manipulate gene expression represents a plausible strategy for modification of the properties of professional antigen-presenting cells in vivo to enhance cancer vaccine potency. PMID- 15665310 TI - Up-regulation of the error-prone DNA polymerase {kappa} promotes pleiotropic genetic alterations and tumorigenesis. AB - It is currently widely accepted that genetic instability is key to cancer development. Many types of cancers arise as a consequence of a gradual accumulation of nucleotide aberrations, each mutation conferring growth and/or survival advantage. Genetic instability could also proceed in sudden bursts leading to a more drastic upheaval of structure and organization of the genome. Genetic instability, as an operative force, will produce genetic variants and the greater the instability, the larger the number of variants. We report here that the overexpression of human DNA polymerase kappa, an error-prone enzyme that is up-regulated in lung cancers, induces DNA breaks and stimulates DNA exchanges as well as aneuploidy. Probably as the result of so many perturbations, excess polymerase kappa favors the proliferation of competent tumor cells as observed in immunodeficient mice. These data suggest that altered regulation of DNA metabolism might be related to cancer-associated genetic changes and phenotype. PMID- 15665311 TI - Apolipoprotein E is required for cell proliferation and survival in ovarian cancer. AB - Apolipoprotein E (ApoE) has been recently identified as a potential tumor associated marker in ovarian cancer by serial analysis of gene expression. ApoE has long been known to play a key role in lipid transport, and its specific isoforms may participate in atherosclerogenesis. However, its role in human cancer is not known. In this study, apoE expression was frequently detected in ovarian serous carcinomas, the most common and lethal type of ovarian cancer. It was not detected in serous borderline tumors and normal ovarian surface epithelium. Inhibition of apoE expression using an apoE-specific siRNA led to G(2) cell cycle arrest and apoptosis in an apoE-expressing ovarian cancer cell line, OVCAR3, but not in apoE-negative cell lines. Furthermore, the phenotype of apoE siRNA-treated OVCAR3 cells was reversed by expressing engineered mutant apoE with introduced silent mutations in the siRNA target sequence. Expression of apoE in nuclei was significantly associated with a better survival in patients who presented peritoneal effusion at the time of diagnosis (5-year follow-up, P = 0.004). This study suggests a new role of apoE in cancer as apoE expression is important for the proliferation and survival in apoE-expressing ovarian cancer cells. PMID- 15665313 TI - Cell cycle checkpoints, DNA damage/repair, and lung cancer risk. AB - Given that defects in cell cycle control and DNA repair capacity may contribute to tumorigenesis, we hypothesized that patients with lung cancer would be more likely than healthy controls to exhibit deficiencies in cell cycle checkpoints and/or DNA repair capacity as gauged by cellular response to in vitro carcinogen exposure. In an ongoing case-control study of 155 patients with newly diagnosed lung cancer and 153 healthy controls, we used the comet assay to investigate the roles of cell cycle checkpoints and DNA damage/repair capability in lung tumorigenesis. The median gamma-radiation-induced and benzo(a)pyrene diol epoxide induced Olive tail moments, the comet assay parameter for measuring DNA damage, were significantly higher in the case group (5.31 and 4.22, respectively) than in the control group (4.42 and 2.83, respectively; P < 0.001). Higher tail moments of gamma-radiation and benzo(a)pyrene diol epoxide-induced comets were significantly associated with 2.32- and 4.49-fold elevated risks, respectively, of lung cancer. The median gamma-radiation-induced increases of cells in the S and G(2) phases were significantly lower in cases (22.2% and 12.2%, respectively) than in controls (31.1% and 14.9%, respectively; P < 0.001). Shorter durations of the S and G(2) phases resulted in 4.54- and 1.85-fold increased risks, respectively, of lung cancer. Also observed were joint effects between gamma radiation-induced increases of S and G(2) phase frequencies and mutagen-induced comets. In addition, we found that in controls, the S phase decreased as tail moment increased. This study is significant because it provides the first molecular epidemiologic evidence linking defects in cell cycle checkpoints and DNA damage/repair capacity to elevated lung cancer risk. PMID- 15665312 TI - Novel prognostic value of nuclear epidermal growth factor receptor in breast cancer. AB - Epidermal growth factor receptor (EGFR) has been detected in the nucleus of cancer cells and primary tumors for decades. While localized in the nucleus, EGFR functions as a transcriptional regulator resulting in the activation of the cyclin D1 gene. Despite nuclear accumulation of EGFR is linked to increased DNA synthesis and proliferative potential, the pathological significance of nuclear EGFR, however, remains uninvestigated. Furthermore, expression of EGFR has not provided a consistent predictive value for survival of breast cancer patients. Here, we analyzed 130 breast carcinomas via immunohistochemical analyses for the levels of nuclear and non-nuclear EGFR. We found 37.7% of the cohort immunostained positively for nuclear EGFR and 6.9% with high levels of expression. Importantly, Kaplan-Meier survival analysis and log-rank test revealed a significant inverse correlation between high nuclear EGFR and overall survival (P = 0.009). Expression of nuclear EGFR correlated positively with increased levels of cyclin D1 and Ki-67, both are indicators for cell proliferation. In contrast, expression of non-nuclear EGFR did not significantly correlate with those of cyclin D1 and Ki-67 or the overall survival rate. In addition, we analyzed 37 oral squamous carcinomas for EGFR expression and found 24.3% of the cases to contain moderate/high levels of nuclear EGFR. Taken together, our findings indicate pathological significance of nuclear EGFR and may have important clinical implication. PMID- 15665314 TI - Association of fetal hormone levels with stem cell potential: evidence for early life roots of human cancer. AB - Intrauterine and perinatal factors have been linked to risk of childhood leukemia, testicular cancer, and breast cancer in the offspring. The pool of stem cells in target tissue has been suggested as a critical factor linking early life exposures to cancer. We examined the relation between intrauterine hormone levels and measurements of stem cell potential in umbilical cord blood. Cord blood donors were 40 women, ages >/=18 years, who delivered, from August 2002 to June 2003, a singleton birth after a gestation of at least 37 weeks. We assayed plasma concentrations of estradiol, unconjugated estriol, testosterone, progesterone, prolactin, sex hormone binding globulin, insulin-like growth factor-I (IGF-I), and IGF binding protein-3. For stem cell potential, we measured concentrations of CD34(+) and CD34(+)CD38(-) cells and granulocyte-macrophage colony-forming unit (CFU-GM). We applied linear regression analysis and controlled for maternal and neonatal characteristics. We found strong positive associations between IGF-I and stem cell measures, 1 SD increase in IGF-I being associated with a 41% increase in CD34(+) (P = 0.008), a 109% increase in CD34(+)CD38(-) (P = 0.005), and a 94% increase in CFU-GM (P = 0.01). Similar associations were observed for IGF binding protein-3. Among steroid hormones, estriol and testosterone were significantly positively associated with CD34(+) and CFU-GM. These findings indicate that levels of growth factors and hormones are strongly associated with stem cell potential in human umbilical cord blood and point to a potential mechanism that may mediate the relationship between in utero exposure to hormones and cancer risk in the offspring. PMID- 15665315 TI - Inhibition of nuclear translocation of nuclear factor-{kappa}B contributes to 3,3'-diindolylmethane-induced apoptosis in breast cancer cells. AB - Dietary indole-3-carbinol (I3C), a natural compound present in vegetables of the genus Brassica, showed clinical benefits and caused apoptosis in breast cancer cells. Our laboratory and others have shown that I3C induces apoptosis in breast cancer cells mediated by inactivation of Akt and nuclear factor-kappaB (NF kappaB) pathway. 3,3'-Diindolylmethane (DIM), a major in vivo acid-catalyzed condensation product of I3C, also showed some benefit in breast cancer. However, the precise molecular mechanism(s) by which DIM induces apoptosis in breast cancer cells has not been fully elucidated. Hence, we investigated whether DIM induced apoptosis of breast cancer cells could also be mediated by inactivation of Akt and NF-kappaB. We found that DIM induces apoptotic processes in MCF10A derived malignant (MCF10CA1a) cell lines but not in nontumorigenic parental MCF10A cells. DIM specifically inhibits Akt kinase activity and abrogates the epidermal growth factor-induced activation of Akt in breast cancer cells, similar to those observed for I3C. We also found that DIM reduces phosphorylation of IkappaBalpha, an inhibitor of NF-kappaB. Our confocal microscopy study clearly showed that DIM blocks the translocation of p65, a subunit of NF-kappaB to the nucleus. DNA binding analysis and transfection studies with IkappaB kinase cDNA revealed that overexpression of IkappaB kinase mediates IkappaBalpha phosphorylation, which activates NF-kappaB, and this activation was completely abrogated by DIM treatment. Taken together, these results showed for the first time that the inactivation of Akt and NF-kappaB activity also plays important roles in DIM-induced apoptosis in breast cancer cells, which seems to be more relevant to in vivo situations. PMID- 15665316 TI - Glypican-3 as a serum marker for hepatocellular carcinoma. PMID- 15665317 TI - Cheyne-Stokes respiration in stroke: relationship to hypocapnia and occult cardiac dysfunction. AB - BACKGROUND: Central sleep apnea (CSA) and Cheyne-Stokes respiration have been reported in association with stroke, but their pathophysiologic correlates have not been well described. OBJECTIVE: To test the hypotheses that (1) CSA in patients with stroke is associated with nocturnal hypocapnia and (2) in those stroke patients with CSA and with left ventricular (LV) systolic dysfunction, periodic breathing (PB) will have a Cheyne-Stokes respiration pattern in which cycle duration is greater than in those without LV systolic dysfunction. METHODS: We prospectively performed polysomnography and echocardiography in 93 patients with stroke. CSA was defined as central apneas and hypopneas occurring at a rate of 10 or more per hour of sleep. In patients with CSA, we compared PB cycle duration between those with normal and impaired LV systolic function (LV ejection fraction [LVEF] > 40% and < or = 40%, respectively). RESULTS: CSA was found in 19% of subjects who had lower nocturnal transcutaneous PCO2 (39.3 +/- 0.9 vs. 42.8 +/- 0.8 mmHg, p = 0.015) and a higher prevalence of LVEF of 40% or less (22 vs. 5%, p = 0.043) than stroke patients without CSA. There was no significant difference in stroke location or type between the two groups. In patients with CSA, those with LVEF of 40% or less had a longer PB cycle than those with an LVEF of more than 40% (66.6 +/- 5.6 vs. 46.6 +/- 2.9 seconds, p = 0.006), but had no symptoms of heart failure. CONCLUSION: In patients with stroke, CSA is associated with hypocapnia and occult LV systolic dysfunction but is not related to the location or type of stroke. The presence of LV systolic dysfunction is associated with a Cheyne-Stokes pattern of hyperpnea. PMID- 15665318 TI - Transbronchial versus transesophageal ultrasound-guided aspiration of enlarged mediastinal lymph nodes. AB - RATIONALE: Transesophageal and transbronchial, ultrasound-guided, fine-needle aspiration of enlarged mediastinal lymph nodes have become popular, but have never been compared directly. OBJECTIVES: To compare the relative diagnostic yield and ability of the transesophageal and transbronchial approaches to reach abnormal mediastinal lymph nodes. METHODS: A total of 160 patients with enlarged lymph nodes in one of eight mediastinal lymph node stations underwent transbronchial and transesophageal biopsies in a crossover design. Each of the eight stations was allocated 20 patients. Two needle punctures were done with each approach. MEASUREMENTS: Percentage of successful biopsies, percentage of patients diagnosed, and biopsy time were measured from when the lymph node was identified with ultrasound. MAIN RESULTS: Among the 106 men and 54 women (mean age 53.2 years), transbronchial aspiration was successful in 85%, and transesophageal aspiration was successful in 78% (p = 0.2). For each station, the number of positive samples for the transbronchial/transesophageal approaches was: 2R: 19/13; 2L: 16/19; 3: 17/15; 4R: 19/12; 4L: 17/20; 7: 19/20; 10R: 18/9; and 10L: 17/18. Combining both approaches produced successful biopsies in 97% and diagnoses in 94% of patients. Mean biopsy times were 3.2 minutes for the transbronchial approach and 4.1 minutes for the transesophageal approach. The transbronchial approach was superior in nodes 2R, 4R, and 10R. No complications were encountered. CONCLUSIONS: In experienced hands, enlarged mediastinal lymph nodes may be aspirated with either the transbronchial or transesophageal approach. These nonsurgical approaches have similar diagnostic yields, although the transbronchial approach is superior for right-sided lymph nodes. Combining both approaches provides results similar those of mediastinoscopy. PMID- 15665319 TI - Risk of post-traumatic stress symptoms in family members of intensive care unit patients. AB - RATIONALE: Intensive care unit (ICU) admission of a relative is a stressful event that may cause symptoms of post-traumatic stress disorder (PTSD). OBJECTIVES: Factors associated with these symptoms need to be identified. METHODS: For patients admitted to 21 ICUs between March and November 2003, we studied the family member with the main potential decision-making role. MEASUREMENTS: Ninety days after ICU discharge or death, family members completed the Impact of Event Scale (which evaluates the severity of post-traumatic stress reactions), Hospital Anxiety and Depression Scale, and 36-item Short-Form General Health Survey during a telephone interview. Linear regression was used to identify factors associated with the risk of post-traumatic stress symptoms. MAIN RESULTS: Interviews were obtained for family members of 284 (62%) of the 459 eligible patients. Post traumatic stress symptoms consistent with a moderate to major risk of PTSD were found in 94 (33.1%) family members. Higher rates were noted among family members who felt information was incomplete in the ICU (48.4%), who shared in decision making (47.8%), whose relative died in the ICU (50%), whose relative died after end-of-life decisions (60%), and who shared in end-of-life decisions (81.8%). Severe post-traumatic stress reaction was associated with increased rates of anxiety and depression and decreased quality of life. CONCLUSION: Post-traumatic stress reaction consistent with a high risk of PTSD is common in family members of ICU patients and is the rule among those who share in end-of-life decisions. Research is needed to investigate PTSD rates and to devise preventive and early detection strategies. PMID- 15665320 TI - Graft ischemic time and outcome of lung transplantation: a multicenter analysis. AB - RATIONALE: The effect of graft ischemic time on early graft function and long term survival of patients who underwent lung transplantation remains controversial. Consequently, graft ischemic time has not been incorporated in the decision-making process at the time of graft acceptance. OBJECTIVES: To investigate the relationship between graft ischemic time and (1) early graft function and (2) long-term survival after lung transplantation. MEASUREMENTS AND MAIN RESULTS: The data from 752 patients who underwent single lung transplantation (n = 258), bilateral lung transplantation (n = 247), and heart lung transplantation (n = 247) in seven French transplantation centers during a 12-year period were reviewed. Independent data quality control was done to ensure the quality of the collected variables. Mean graft ischemic time was 245.8 +/- 96.4 minutes (range 50-660). After adjustment on 11 potential confounders, graft ischemic time was associated with the recipient Pa(O2)/FI(O2) ratio recorded within the first 6 hours and with long-term survival in patients undergoing single or double lung transplantation but not in patients undergoing heart-lung transplantation. The relationship between graft ischemic time and survival appears to be of cubic form with a cutoff value of 330 minutes. These results were unaffected by the preservation fluid employed. CONCLUSIONS: The results of this large cohort of patients suggest a close relationship between graft ischemic time and both early gas exchange and long-term survival after single and double lung transplantation. Such relationship was not found in patients undergoing heart-lung transplantation. The expected graft ischemic time should be incorporated in the decision-making process at the time of graft acceptance. PMID- 15665322 TI - Effects of high versus low positive end-expiratory pressures in acute respiratory distress syndrome. AB - A recent study by the Acute Respiratory Distress Syndrome Network compared the traditional lower end-expiratory pressure strategy with a higher end-expiratory pressure strategy in patients with the acute respiratory distress syndrome ventilated with low tidal volumes. Clinical outcomes were similar whether lower or higher positive end-expiratory pressure (PEEP) levels were used. We applied both the lower (9 +/- 2 cm H2O) and higher (16 +/- 1 cm H2O) PEEP strategy in 19 patients. In nine recruiters, the higher end-expiratory pressure strategy resulted in significant alveolar recruitment (587 +/- 158 ml), improvement in arterial oxygen partial pressure/inspired oxygen fraction ratio (from 150 +/- 36 to 396 +/- 138), and reduction in static lung elastance (from 23 +/- 3 to 20 +/- 2 cm H2O/L). In 10 nonrecruiters, alveolar recruitment was minimal, oxygenation did not improve, and static lung elastance significantly increased (from 26 +/- 5 to 28 +/- 6 cm H2O/L). The increase in oxygenation, the reduction in static lung elastance, and the shape of the volume-pressure curve during the lower PEEP strategy were independently associated with alveolar recruitment. In conclusion, the protocol proposed by the Acute Respiratory Distress Syndrome Network, lacking solid physiologic basis, frequently fails to induce alveolar recruitment and may increase the risk of alveolar overinflation. PMID- 15665321 TI - Inhibition of the Src and Jak kinases protects against lipopolysaccharide-induced acute lung injury. AB - The cascade of cellular and molecular pathways mediating acute lung injury is complex and incompletely defined. Although the Src and Jak family of kinases is upregulated in LPS-induced murine lung injury, their role in the development of lung injury is unknown. Here we report that systemic inhibition of these kinases using specific small molecule inhibitors (PP2, SU6656, tyrphostin A1) significantly attenuated LPS-induced lung injury, as determined by histologic and capillary permeability assays. These inhibitors blocked LPS-dependent cytokine and chemokine production in the lung and in the serum. In contrast, lung-targeted inhibition of these kinases in the airway epithelium via adenoviral-mediated gene transfer of dominant negative Src or of suppressor of cytokine signaling (SOCS-1) disrupted lung cytokine production but had no effect on systemic cytokine production or lung vascular permeability. Mice were significantly protected from lethal LPS challenge by the small molecule inhibitors of Jak and Src kinase. Importantly, this protection was still evident even when the inhibitors were administered 6 hours after LPS challenge. Taken together, these observations suggest that Jak and Src kinases participate in acute lung injury and verify the potential of this class of selective tyrosine kinase inhibitors to serve as novel therapeutic agents for this disease. PMID- 15665323 TI - Closed-loop control of respiratory drive using pressure-support ventilation: target drive ventilation. AB - By using diaphragm electrical activity (multiple-array esophageal electrode) as an index of respiratory drive, and allowing such activity above or below a preset target range to indicate an increased or reduced demand for ventilatory assistance (target drive ventilation), we evaluated whether the level of pressure support ventilation can be automatically adjusted in response to exercise-induced changes in ventilatory demand. Eleven healthy individuals breathed through a circuit (18 cm H2O/L/second inspiratory resistance at 1 L/second flow; 0.5-1.0 L/second expiratory flow limitation) connected to a modified ventilator. Subjects breathed for 6-minute periods at rest and during 20 and 40 W of bicycle exercise, with and without target drive ventilation (the target was set to 60% of the increase in diaphragm electrical activity observed between rest and 20 W of unassisted exercise). With target drive ventilation during exercise, the level of pressure-support ventilation was automatically increased, reaching 13.3 +/- 4.0 and 20.3 +/- 2.8 cm H2O during 20- and 40-W exercise, respectively, whereas diaphragm electrical activity was reduced to a level within the target range. Both diaphragmatic pressure-time product and end-tidal CO2 were significantly reduced with target drive ventilation at the end of the 20- (p < 0.01) and 40-W (p < 0.001) exercise periods. Minute ventilation was not altered. These results demonstrate that target drive ventilation can automatically adjust pressure support ventilation, maintaining a constant neural drive and compensating for changes in respiratory demand. PMID- 15665324 TI - Characteristics of physical activities in daily life in chronic obstructive pulmonary disease. AB - Quantification of physical activities in daily life in patients with chronic obstructive pulmonary disease has increasing clinical interest. However, detailed comparison with healthy subjects is not available. Furthermore, it is unknown whether time spent actively during daily life is related to lung function, muscle force, or maximal and functional exercise capacity. We assessed physical activities and movement intensity with the DynaPort activity monitor in 50 patients (age 64 +/- 7 years; FEV1 43 +/- 18% predicted) and 25 healthy elderly individuals (age 66 +/- 5 years). Patients showed lower walking time (44 +/- 26 vs. 81 +/- 26 minutes/day), standing time (191 +/- 99 vs. 295 +/- 109 minutes/day), and movement intensity during walking (1.8 +/- 0.3 vs. 2.4 +/- 0.5 m/second2; p < 0.0001 for all), as well as higher sitting time (374 +/- 139 vs. 306 +/- 108 minutes/day; p = 0.04) and lying time (87 +/- 97 vs. 29 +/- 33 minutes/day; p = 0.004). Walking time was highly correlated with the 6-minute walking test (r = 0.76, p < 0.0001) and more modestly to maximal exercise capacity, lung function, and muscle force (0.28 < r < 0.64, p < 0.05). Patients with chronic obstructive pulmonary disease are markedly inactive in daily life. Functional exercise capacity is the strongest correlate of physical activities in daily life. PMID- 15665325 TI - Upregulation of phosphodiesterase-4 in the lung of allergic rats. AB - Inhibitors of phosphodiesterase-4 (PDE4) are efficacious for allergic asthma in animal models and have shown some efficacy in human asthma. Regulation of PDE4 in allergy and asthma has been widely investigated in blood leukocytes, with discrepant results. This study investigated PDE4 regulation in the lung in a rat model of allergic asthma. Ovalbumin sensitization and challenge significantly increased pulmonary resistance and lung interleukin (IL)-4 production. The increases in pulmonary resistance and IL-4 production were both suppressed by the PDE4-selective inhibitor rolipram or the corticosteroid drug dexamethasone. Furthermore, cAMP-PDE enzyme activity in the lung was also significantly increased by the sensitization and challenge. mRNA analysis confirmed that PDE4 gene expression was increased in the lung of the allergic rats. A highly significant correlation was observed between the increases in PDE activity and IL 4 production. Our data suggest, for the first time, that PDE4 may be upregulated in the lung and play a role in the pathogenesis of allergic asthma. PMID- 15665326 TI - Double-blind, placebo-controlled trial of pirfenidone in patients with idiopathic pulmonary fibrosis. AB - Idiopathic pulmonary fibrosis (IPF) is a fatal disorder without an effective therapy to date. In a double-blind, randomized, placebo-controlled trial, 107 patients were prospectively evaluated for efficacy of a novel compound, pirfenidone. The difference in the change in the lowest oxygen saturation by pulse oximetry (SpO2) during a 6-minute exercise test, the primary endpoint, from baseline to 6 months was not significant between the two groups (p = 0.0722). In a prespecified subset of patients who maintained a SpO2 greater than 80% during a 6-minute exercise test at baseline, the lowest SpO2 improved during a 6-minute exercise test in the pirfenidone group at 6 and 9 months (p = 0.0069 and 0.0305, respectively). Positive treatment effect was demonstrated in secondary endpoints: (1) change in VC measurements at 9 months (p = 0.0366) and (2) episodes of acute exacerbation of IPF occurring exclusively in the placebo group during the 9 months (p = 0.0031). Significant adverse events were associated with pirfenidone; however, adherence to treatment regimen was similar between pirfenidone and placebo groups. In conclusion, treatment with pirfenidone improved VC and prevented acute exacerbation of IPF during the 9 months of follow-up. Future long term studies are needed to clarify the overall safety and efficacy of pirfenidone in IPF. PMID- 15665328 TI - Proline metabolism in procyclic Trypanosoma brucei is down-regulated in the presence of glucose. AB - Proline metabolism has been studied in procyclic form Trypanosoma brucei. These parasites consume six times more proline from the medium when glucose is in limiting supply than when this carbohydrate is present as an abundant energy source. The sensitivity of procyclic T. brucei to oligomycin increases by three orders of magnitude when the parasites are obliged to catabolize proline in medium depleted in glucose. This indicates that oxidative phosphorylation is far more important to energy metabolism in this latter case than when glucose is available and the energy needs of the parasite can be fulfilled by substrate level phosphorylation alone. A gene encoding proline dehydrogenase, the first enzyme of the proline catabolic pathway, was cloned. RNA interference studies revealed the loss of this activity to be conditionally lethal. Proline dehydrogenase defective parasites grew as wild-type when glucose was available, but, unlike wild-type cells, they failed to proliferate using proline. In parasites grown in the presence of glucose, proline dehydrogenase activity was markedly lower than when glucose was absent from the medium. Proline uptake too was shown to be diminished when glucose was abundant in the growth medium. Wild type cells were sensitive to 2-deoxy-D-glucose if grown using proline as the principal carbon source, but not in glucose-rich medium, indicating that this non catabolizable glucose analogue might also stimulate repression of proline utilization. These results indicate that the ability of trypanosomes to use proline as an energy source can be regulated depending upon the availability of glucose. PMID- 15665329 TI - A novel metabolic pathway for degradation of 4-nonylphenol environmental contaminants by Sphingomonas xenophaga Bayram: ipso-hydroxylation and intramolecular rearrangement. AB - Several nonylphenol isomers with alpha-quaternary carbon atoms serve as growth substrates for Sphingomonas xenophaga Bayram, whereas isomers containing hydrogen atoms at the alpha-carbon do not. Three metabolites of 4-(1-methyloctyl)-phenol were isolated in mg quantities from cultures of strain Bayram supplemented with the growth substrate isomer 4-(1-ethyl-1,4-dimethyl-pentyl)-phenol. They were unequivocally identified as 4-hydroxy-4-(1-methyl-octyl)-cyclohexa-2,5-dienone, 4 hydroxy-4-(1-methyl-octyl)-cyclohex-2-enone, and 2-(1-methyl-octyl)-benzene-1,4 diol by high pressure liquid chromatography-mass spectrometry and nuclear magnetic resonance spectroscopy. Furthermore, two metabolites originating from 4 n-nonylphenol were identified as 4-hydroxy-4-nonyl-cyclohexa-2,5-dienone and 4 hydroxy-4-nonyl-cyclohex-2-enone by high pressure liquid chromatography-mass spectrometry. We conclude that nonylphenols were initially hydroxylated at the ipso-position forming 4-alkyl-4-hydroxy-cyclohexa-2,5-dienones. Dienones originating from growth substrate nonylphenol isomers underwent a rearrangement that involved a 1,2-C,O shift of the alkyl moiety as a cation to the oxygen atom of the geminal hydroxy group yielding 4-alkoxyphenols, from which the alkyl moieties can be easily detached as alcohols by known mechanisms. Dienones originating from nongrowth substrates did not undergo such a rearrangement because the missing alkyl substituents at the alpha-carbon atom prevented stabilization of the putative alpha-carbocation. Instead they accumulated and subsequently underwent side reactions, such as 1,2-C,C shifts and dihydrogenations. The ipso-hydroxylation and the proposed 1,2-C,O shift constitute key steps in a novel pathway that enables bacteria to detach alpha branched alkyl moieties of alkylphenols for utilization of the aromatic part as a carbon and energy source. PMID- 15665330 TI - Maize phosphoenolpyruvate carboxylase. Mutations at the putative binding site for glucose 6-phosphate caused desensitization and abolished responsiveness to regulatory phosphorylation. AB - Phosphoenolpyruvate carboxylases (PEPC, EC 4.1.1.31) from higher plants are regulated by both allosteric effects and reversible phosphorylation. Previous x ray crystallographic analysis of Zea mays PEPC has revealed a binding site for sulfate ion, speculated to be the site for an allosteric activator, glucose 6 phosphate (Glc-6-P) (Matsumura, H., Xie, Y., Shirakata, S., Inoue, T., Yoshinaga, T., Ueno, Y., Izui, K., and Kai, Y. (2002) Structure (Lond.) 10, 1721-1730). Because kinetic experiments have also supported this notion, each of the four basic residues (Arg-183, -184, -231, and -372' on the adjacent subunit) located at or near the binding site was replaced by Gln, and the kinetic properties of recombinant mutant enzymes were investigated. Complete desensitization to Glc-6-P was observed for R183Q, R184Q, R183Q/R184Q (double mutant), and R372Q, as was a marked decrease in the sensitivity for R231Q. The heterotropic effect of Glc-6-P on an allosteric inhibitor, l-malate, was also abolished, but sensitivity to Gly, another allosteric activator of monocot PEPC, was essentially not affected, suggesting the distinctness of their binding sites. Considering the kinetic and structural data, Arg-183 and Arg-231 were suggested to be involved directly in the binding with phosphate group of Glc-6-P, and the residues Arg-184 and Arg-372 were thought to be involved in making up the site for Glc-6-P and/or in the transmission of an allosteric regulatory signal. Most unexpectedly, the mutant enzymes had almost lost responsiveness to regulatory phosphorylation at Ser-15. An apparent lack of kinetic competition between the phosphate groups of Glc-6-P and of phospho-Ser at 15 suggested the distinctness of their binding sites. The possible roles of these Arg residues are discussed. PMID- 15665327 TI - Flow shear stress stimulates Gab1 tyrosine phosphorylation to mediate protein kinase B and endothelial nitric-oxide synthase activation in endothelial cells. AB - Fluid shear stress generated by blood flow modulates endothelial cell function via specific intracellular signaling events. We showed previously that flow activated the phosphatidylinositol 3-kinase (PI3K), Akt, and endothelial nitric oxide synthase (eNOS) via Src kinase-dependent transactivation of vascular endothelial growth factor receptor 2 (VEGFR2). The scaffold protein Gab1 plays an important role in receptor tyrosine kinase-mediated signal transduction. We found here that laminar flow (shear stress = 12 dynes/cm2) rapidly stimulated Gab1 tyrosine phosphorylation in both bovine aortic endothelial cells and human umbilical vein endothelial cells, which correlated with activation of Akt and eNOS. Gab1 phosphorylation as well as activation of Akt and eNOS by flow was inhibited by the Src kinase inhibitor PP2 (4-amino-5-(4-chlorophenyl)-7-(t butyl)pyrazolo[3,4-d]pyrimidine) and VEGFR2 kinase inhibitors SU1498 and VTI, suggesting that flow-mediated Gab1 phosphorylation is Src kinase-dependent and VEGFR2-dependent. Tyrosine phosphorylation of Gab1 by flow was functionally important, because flow stimulated the association of Gab1 with the PI3K subunit p85 in a time-dependent manner. Furthermore, transfection of a Gab1 mutant lacking p85 binding sites inhibited flow-induced activation of Akt and eNOS. Finally, knockdown of endogenous Gab1 by small interference RNA abrogated flow activation of Akt and eNOS. These data demonstrate a critical role of Gab1 in flow-stimulated PI3K/Akt/eNOS signal pathway in endothelial cells. PMID- 15665331 TI - Crystal structure of vinorine synthase, the first representative of the BAHD superfamily. AB - Vinorine synthase is an acetyltransferase that occupies a central role in the biosynthesis of the antiarrhythmic monoterpenoid indole alkaloid ajmaline in the plant Rauvolfia. Vinorine synthase belongs to the benzylalcohol acetyl-, anthocyanin-O-hydroxy-cinnamoyl-, anthranilate-N-hydroxy-cinnamoyl/benzoyl-, deacetylvindoline acetyltransferase (BAHD) enzyme superfamily, members of which are involved in the biosynthesis of several important drugs, such as morphine, Taxol, or vindoline, a precursor of the anti-cancer drugs vincaleucoblastine and vincristine. The x-ray structure of vinorine synthase is described at 2.6 angstrom resolution. Despite low sequence identity, the two-domain structure of vinorine synthase shows surprising similarity with structures of several CoA dependent acyltransferases such as dihydrolipoyl transacetylase, polyketide associated protein A5, and carnitine acetyltransferase. All conserved residues typical for the BAHD family are found in domain 1. His160 of the HXXXD motif functions as a general base during catalysis. It is located in the center of the reaction channel at the interface of both domains and is accessible from both sides. The channel runs through the entire molecule, allowing the substrate and co-substrate to bind independently. Asp164 points away from the catalytic site and seems to be of structural rather than catalytic importance. Surprisingly, the DFGWG motif, which is indispensable for the catalyzed reaction and unique to the BAHD family, is located far away from the active site and seems to play only a structural role. Vinorine synthase represents the first solved protein structure of the BAHD superfamily. PMID- 15665332 TI - The small heat shock protein IbpA of Escherichia coli cooperates with IbpB in stabilization of thermally aggregated proteins in a disaggregation competent state. AB - The small heat shock proteins are ubiquitous stress proteins proposed to increase cellular tolerance to heat shock conditions. We isolated IbpA, the Escherichia coli small heat shock protein, and tested its ability to keep thermally inactivated substrate proteins in a disaggregation competent state. We found that the presence of IbpA alone during substrate thermal inactivation only weakly influences the ability of the bi-chaperone Hsp70-Hsp100 system to disaggregate aggregated substrate. Similar minor effects were observed for IbpB alone, the other E. coli small heat shock protein. However, when both IbpA and IbpB are simultaneously present during substrate inactivation they efficiently stabilize thermally aggregated proteins in a disaggregation competent state. The properties of the aggregated protein substrates are changed in the presence of IbpA and IbpB, resulting in lower hydrophobicity and the ability of aggregates to withstand sizing chromatography conditions. IbpA and IbpB form mixed complexes, and IbpA stimulates association of IbpB with substrate. PMID- 15665333 TI - Kinetic analysis of oxidation of coumarins by human cytochrome P450 2A6. AB - Human cytochrome P450 (P450) 2A6 catalyzes 7-hydroxylation of coumarin, and the reaction rate is enhanced by cytochrome b5 (b5). 7-Alkoxycoumarins were O dealkylated and also hydroxylated at the 3-position. Binding of coumarin and 7 hydroxycoumarin to ferric and ferrous P450 2A6 are fast reactions (k(on) approximately 10(6) m(-1) s(-1)), and the k(off) rates range from 5.7 to 36 s(-1) (at 23 degrees C). Reduction of ferric P450 2A6 is rapid (7.5 s(-1)) but only in the presence of coumarin. The reaction of the ferrous P450 2A6 substrate complex with O2 is rapid (k > or = 10(6) m(-1) s(-1)), and the putative Fe2+.O2 complex decayed at a rate of approximately 0.3 s(-1) at 23 degrees C. Some 7 hydroxycoumarin was formed during the oxidation of the ferrous enzyme under these conditions, and the yield was enhanced by b5. Kinetic analyses showed that approximately 1/3 of the reduced b5 was rapidly oxidized in the presence of the Fe2+.O2 complex, implying some electron transfer. High intrinsic and competitive and non-competitive intermolecular kinetic deuterium isotope effects (values 6 10) were measured for O-dealkylation of 7-alkoxycoumarins, indicating the effect of C-H bond strength on rates of product formation. These results support a scheme with many rapid reaction steps, including electron transfers, substrate binding and release at multiple stages, and rapid product release even though the substrate is tightly bound in a small active site. The inherent difficulty of chemistry of substrate oxidation and the lack of proclivity toward a linear pathway leading to product formation explain the inefficiency of the enzyme relative to highly efficient bacterial P450s. PMID- 15665334 TI - The crystal structure of archaeal nascent polypeptide-associated complex (NAC) reveals a unique fold and the presence of a ubiquitin-associated domain. AB - Nascent polypeptide-associated complex (NAC) was identified in eukaryotes as the first cytosolic factor that contacts the nascent polypeptide chain emerging from the ribosome. NAC is highly conserved from yeast to humans. Mutations in NAC cause severe embryonically lethal phenotypes in mice, Drosophila, and Caenorhabditis elegans. NAC was suggested to protect the nascent chain from inappropriate early interactions with cytosolic factors. Eukaryotic NAC is a heterodimer with two subunits sharing substantial homology with each other. All sequenced archaebacterial genomes exhibit only one gene homologous to the NAC subunits. Here we present the first archaebacterial NAC homolog. It forms a homodimer, and as eukaryotic NAC it is associated with ribosomes and contacts the emerging nascent chain on the ribosome. We present the first crystal structure of a NAC protein revealing two structural features: (i) a novel unique protein fold that mediates dimerization of the complex, and (ii) a ubiquitin-associated domain that suggests a yet unidentified role for NAC in the cellular protein quality control system via the ubiquitination pathway. Based on the presented structure we propose a model for the eukaryotic heterodimeric NAC domain. PMID- 15665335 TI - Otto Fritz Meyerhof and the elucidation of the glycolytic pathway. PMID- 15665336 TI - Retinopathy of prematurity's turning point. PMID- 15665337 TI - Radiotherapy and intracranial meningiomas causing visual disturbance. PMID- 15665338 TI - Fractionated stereotactic radiotherapy for parasellar meningiomas: a preliminary report of visual outcomes. AB - BACKGROUND/AIM: Fractionated stereotactic radiotherapy (FSRT) is a new treatment for brain tumours that are close to critical structures, such as the visual apparatus. This study aims to assess the visual outcomes for patients with parasellar meningioma following FSRT. METHODS: A retrospective, non-comparative case series of 13 patients with parasellar meningiomas who were treated in one institution with FSRT between January 1995 and January 2001. RESULTS: 13 patients (26 eyes) were followed for a mean of 2 years. Visual acuity improved in four eyes (12.5%), remained stable in 18 eyes (75%), and worsened in three eyes (12.5%). Visual field improved in 15 eyes (57%), remained stable in six eyes (23%), and worsened in four eyes (15%). No adverse visual outcome occurred as a result of radiation. CONCLUSION: These preliminary findings suggest that FSRT is a safe and effective treatment for parasellar meningiomas. PMID- 15665339 TI - Transplantation of corneal endothelium with Descemet's membrane using a hyroxyethyl methacrylate polymer as a carrier. AB - AIMS: To evaluate the histology and function of Descemet's membrane transplanted with intact endothelium. METHODS: Japanese white rabbits and human eyebank eyes were used as donors and recipients of Descemet's membrane transplantation. Donor endothelium was hydrodissected by injecting indocyanine green from a limbal incision, and then processed as a corneal scleral button. A 6 mm diameter donor sheet was trephined, and folded in half using a 6 mm diameter polymer as a carrier. Recipient endothelium was also hydrodissected from the limbus using trypan blue to stain the Descemet's membrane. Continuous curvilinear descemetorhexis (CCD) was performed to remove a circular section of the Descemet's membrane using a 27 gauge cystotome. Donor tissue was inserted into the anterior chamber through a 5 mm limbal incision and apposed to the host stroma. Polymers were removed following transplantation. Similar surgical procedures were performed in both rabbits and eyebank eyes. Haematoxylin eosin stains were performed after 28 days in rabbits, and eyebank eyes were fixed immediately following surgery for endothelial cell counts. RESULTS: Rabbit control eyes demonstrated stromal oedema caused by loss of Descemet's membrane, whereas transplanted eyes had clear corneas. The mean (standard deviation) pachymetry of operated eyes was 376.6 (SD 32.5) mum compared with 389.6 (SD 25.1) mum in the unoperated eye. Mean endothelial density immediately following surgery in eyebank eyes was 2749 (SD 288) cells/mm(2). CONCLUSIONS: Transplantation of Descemet's membrane by CCD produces a functional graft with an optically clear interface similar to control cornea. PMID- 15665340 TI - Posterior polar cataract is the predominant consequence of a recurrent mutation in the PITX3 gene. AB - BACKGROUND: The authors recently identified three large genetically unrelated families with an identical 17 base pair duplication mutation in exon 4 of the PITX3 gene. Here, they report the detailed clinical phenotype. METHODS: Affected and unaffected individuals in the three families with autosomal dominant posterior polar cataract underwent full clinical examination and donated blood samples for DNA extraction and molecular genetic studies. RESULTS: In all three families, an identical 17 base pair duplication mutation in PITX3 was identified which co-segregated with disease status in the family. All affected individuals had bilateral progressive posterior polar cataracts. In one family, posterior polar cataract was the only clinical abnormality but in the other two families, one of 10 affected individuals and four of 11 affected individuals also had anterior segment mesenchymal dysgenesis (ASMD). CONCLUSION: Mutations in the PITX3 gene in humans result in posterior polar cataract and variable ASMD. The gene encodes a transcription factor which has a key role in lens and anterior segment development. The mechanism by which the mutant protein gives rise to such a regional pattern of lens opacity remains to be elucidated. PMID- 15665341 TI - Exaggerated relative nasal-temporal asymmetry of macular capillary blood flow in patients with clinically significant diabetic macular oedema. AB - AIMS: To compare the macular capillary blood flow of patients with clinically significant diabetic macular oedema (DMO) with that of non-diabetic subjects and to determine the relation between blood flow and capillary leakage in patients with DMO. METHODS: The sample comprised 45 non-diabetic subjects (mean age 59 years) and 18 type 2 patients with clinically significant DMO (mean age 60 years). Macular capillary blood flow measurements were acquired using the Heidelberg retina flowmeter (HRF) and a 10 degrees x2.5 degrees scan field centred on the fovea. Fluorescein angiography was undertaken on each of the diabetic patients after the completion of HRF measurements. RESULTS: Temporal macular capillary blood flow was significantly lower for the patients with clinically significant DMO compared with age matched non-diabetic subjects (ANCOVA, p = 0.0011) while relative nasal-temporal asymmetry of macular capillary blood flow was significantly higher (p = 0.0125). Nasal-temporal asymmetry of macular capillary blood flow was significantly higher for the patients with DMO and capillary leakage within the scan area (two tailed t test, p = 0.0071). Macular capillary blood flow was always lower in areas of DMO and capillary leakage. CONCLUSION: Capillary blood flow was reduced in areas of DMO and capillary leakage, suggesting the presence of a localised perturbation of capillary blood flow regulation. PMID- 15665342 TI - Photodynamic therapy for inflammatory choroidal neovascularisation unresponsive to immunosuppression. AB - AIM: To report on visual and angiographic outcomes of a consecutive series of patients with inflammatory choroidal neovascular membranes (CNV) unresponsive to systemic immunosuppression treated with photodynamic therapy (PDT). METHODS: The medical records of six consecutive patients with inflammatory CNVs that failed to respond to systemic immunosuppression and that later underwent PDT were retrospectively reviewed. Patient demographics, visual acuity, and fluorescein angiographic findings were evaluated. RESULTS: There were five females and one male with a mean age of 40.8 years (range 35-58 years). Four patients had clinical features consistent with punctate inner choroidopathy and two with presumed ocular histoplasmosis. In all cases clinical signs of CNV activity, including subretinal fluid, subretinal blood, hard exudates, and/or recent decrease in visual acuity were present prior to PDT. All patients had been treated with high dose systemic immunosuppressants, which failed to induce regression of the CNV and/or to improve vision. The CNVs were subfoveal in five patients and juxtafoveal in one; all were classified as predominantly classic. Following PDT an improvement in vision occurred in all cases (median improvement of 18 letters, range 3-42 letters). At last follow up, signs of decreased activity in the CNV were detected in all cases. Patients were followed for a median of 10 months (range 9-20 months). CONCLUSION: PDT appears to be a useful option in the management of patients with inflammatory CNVs unresponsive to immunosuppressive therapies. PMID- 15665343 TI - Multifocal electroretinogram in children on atropine treatment for myopia. AB - AIM: To assess retinal function by multifocal electroretinogram (mfERG) in children on atropine eye drops for the treatment of myopia. METHODS: mfERGs were recorded in children receiving atropine eye drops (n = 48) once daily for 2 years and in those receiving placebo eye drops (n = 57) for a similar time. All recordings were performed between the second and third month of cessation of atropine/placebo treatment by a masked investigator. The amplitude and implicit time of the first order kernel (k1) and first slice of the second order kernel (k21) of mfERG responses were used to study the outer and inner retinal function, respectively. RESULTS: There was no significant reduction in k1 response amplitudes of the atropine group compared to that of the placebo group (N1, p = 0.181; P1, p = 0.150). No significant difference in the k1 response implicit times between the groups was found (N1, p = 0.767; P1, p = 0.849). The differences in the k21 amplitudes and implicit times between the groups were not statistically significant (k21 amplitude, p = 0.058; k21 implicit time, p = 0.156). CONCLUSIONS: Daily atropine usage over 2 years for the treatment of myopia has no significant effect on retinal function as demonstrated by recordings of mfERG. PMID- 15665344 TI - Treated threshold stage 3 versus spontaneously regressed subthreshold stage 3 retinopathy of prematurity: a study of motility, refractive, and anatomical outcomes at 6 months and 36 months. AB - AIM: To compare the visual acuity (VA), spherical equivalent refractive error, motility, and anatomical outcomes in children with treated regressed threshold stage 3 retinopathy of prematurity (ROP) and those with spontaneously regressed subthreshold stage 3 ROP. METHOD: 6 month and 3 year data collected from infants examined between 1989 and 1999 with regressed stage 3 ROP, with or without treatment were retrospectively reviewed. RESULTS: 85 infants were included in this study. 40 eyes received cryotherapy, 81 eyes laser photocoagulation, and 34 eyes had spontaneously regressed subthreshold stage 3 ROP. Grating acuity score > or =2 cycles/degree (c/d) at 6 months was predictive of optotype acuity > or =6/9 in 69% of eyes and a score <2 c/d at 6 months was predictive of acuity < 6/9 in 88% of eyes. Eyes with subthreshold stage 3 ROP were twice as likely to have VA of 6/9 or better at 36 months than the treated eyes. The mean spherical equivalent refractive error at 36 months was -6.5 dioptres (D) (-21.5D to +1.38D) in cryotherapy treated eyes, -2.4D (-13D to +4D) in the laser group, and -0.22D ( 9D to +2.25D) in the subthreshold group. Eyes within the treated groups were more myopic than the eyes within the spontaneously regressed group (p = 0.005). At 36 months, 42 out of the 85 infants (that is, 49%) had strabismus (44% in the cryotherapy group, 26% in the laser group, and 25% in the subthreshold group). There was a statistically significant association between the presence of strabismus and anisometropia (p = 0.016) and strabismus and intraventricular haemorrhage (IVH) (p = 0.005). There was a statistically significant difference in the incidence of strabismus between mild and moderate and severe grade IVH (p = 0.01). Eight out of 40 eyes in the cryotherapy group and six out of the 81 eyes in the laser group developed macular ectopia. None of the eyes in the spontaneously regressed group had macular dragging. CONCLUSIONS: In this study, the grating acuity at 6 months was a good predictor of the 3 year optotype acuity in all groups. Eyes with spontaneously regressed subthreshold stage 3 ROP were associated with better vision at 3 years of age and a lesser degree of myopia compared to the treated groups. Strabismus developed predominantly in the treated groups and was frequently associated with neurological damage and/or anisometropia. The spontaneously regressed subthreshold stage 3 group had a better anatomical outcome compared to the groups in which the retinopathy regressed following treatment. PMID- 15665345 TI - The preoperative intraocular pressure level predicts the amount of underestimated intraocular pressure after LASIK for myopia. AB - AIMS: To evaluate the statistical significance of the parameters that affect underestimation of intraocular pressure (IOP) after laser in situ keratomileusis (LASIK) for myopia. METHODS: In this prospective case series study, patient age, axial length, preoperative corneal curvature, preoperative central corneal thickness (CCT), preoperative IOP, and ablation depth were studied to determine whether they affect the underestimation of IOP in the right eyes of 100 consecutive patients who underwent LASIK. RESULTS: The preoperative IOP was the most important parameter for an amount of underestimated Goldmann applanation tonometric IOP (GAT) and non-contact tonometric IOP (ncIOP) at 1 month (r = 0.654, p<0.0001, R(2) = 0.427, and r = 0.694, p<0.0001, R(2) = 0.481, respectively) and 3 months (r = 0.637, p<0.0001, R(2) = 0.406, and r = 0.726, p<0.0001, R(2) = 0.527, respectively). Patient age was statistically significant for underestimating the GAT at 1 month, and both the ablation depth and CCT were statistically significant parameters for underestimating the ncIOP at 1 month and at 3 months by stepwise multiple regression analysis (F>4.000). However, these parameters had small bivariate correlation coefficients, and were considered as minor parameters. CONCLUSION: Preoperative IOP is the most important parameter that affects an underestimation of IOP after LASIK for myopia. Eyes with a higher true IOP have a larger underestimation of the IOP after LASIK for myopia. From these results, the importance of the modulus of elasticity on IOP measurements was discussed. PMID- 15665346 TI - Augmented trabeculectomy in paediatric glaucoma. AB - AIMS: To report the experience with trabeculectomy augmented with mitomycin C and 5-fluorouracil for the treatment of paediatric glaucoma. METHODS: Retrospective, interventional case series design was used. The sample included 17 children (29 eyes) with primary (19 eyes) or secondary (10 eyes) glaucoma who were treated with augmented trabeculectomy as the primary procedure between 1990 and 2002. Data were collected on age and family history, preoperative and end of follow up intraocular pressure, cup/disc ratio (evaluated by drawing), visual acuity, complications, and post-surgery treatment. RESULTS: Patient age at surgery ranged from 1 month to 8 years; most patients (n = 14, 82.3%) were aged less than 1 year (range 1 month-8 months, mean 3.95 (SD 2.56) months); three patients (17.7%) were aged 3, 5, and 8 years. The duration of follow up was 3-120 months (mean 46 months). Intraocular pressure significantly improved from 21 mm Hg to 60 mm Hg (mean 33.1 (10) mm Hg) before surgery to 6-26 mm Hg (mean 17.1 (6) mm Hg) after, (p <0.0001). There was no significant change in cup/disc ratio: 0.1-0.8 (mean 0.42 (0.26)) before and 0.1-1.0 (mean 0.511 (0.27)) after (p = 0.45). In 22 eyes (75.8%), intraocular pressure was controlled at less than 20 mm Hg and the cup/disc ratio remained stable or improved. The life table success rate for intraocular pressure control remained stable at 86% at the 12, 24, and 36 months and after 48 months decreased to 53%. There was no significant difference in the life table results between primary and secondary glaucoma. 14 eyes (48.2%) had a visual acuity better than 20/120 by the end of follow up. Repeated surgery was necessary in eight eyes (27.5%), and additional antiglaucoma treatment in 13 (44.8%). Complications included retinal detachment 1 year after surgery, choroidal detachment, and blebitis (one eye each). CONCLUSIONS: Augmented trabeculectomy with mitomycin C and 5-fluorouracil may serve as the primary procedure in a selected group of paediatric patients with glaucoma. PMID- 15665347 TI - Elevated aqueous humour tissue inhibitor of matrix metalloproteinase-1 and connective tissue growth factor in pseudoexfoliation syndrome. AB - BACKGROUND/AIMS: Pseudoexfoliation syndrome (PXF) was recently found to be associated with increased expression of transforming growth factor beta(1) (TGFbeta(1)) in the aqueous humour. As concern has been raised regarding anti TGFbeta therapy, which can potentially disrupt the maintenance of anterior chamber associated immune deviation, the authors explored the levels of tissue inhibitor of matrix metalloproteinase-1 (TIMP-1), matrix metalloproteinase-9 (MMP 9), and connective tissue growth factor (CTGF) in aqueous humour to determine if these may represent alternative therapeutic targets. METHODS: Aqueous humour samples were collected from patients who underwent routine cataract surgery. All patients were categorised into three main groups-PXF, uveitis, and control. The PXF group was further subcategorised into three grades based on the density of the exfoliative material observed on biomicroscopy, as well as the presence or absence of glaucoma. TIMP-1, MMP-9, and CTGF levels were measured using specific enzyme immunoassays (ELISA). RESULTS: Eyes with PXF had significantly higher aqueous humour TIMP-1 concentration (n = 56, mean (SE), 9.76 (1.10) ng/ml) compared with controls (n = 112, 5.73 (0.43) ng/ml, p<0.01). Similarly, the CTGF level in PXF eyes (n = 36, 4.38 (0.65) ng/ml) was higher than controls (n = 29, 2.35 (0.46) ng/ml, p<0.05). Further, the CTGF concentration in the PXF glaucoma group is significantly higher compared with PXF eyes without glaucoma (6.03 (1.09) ng/ml v 2.73 (0.45) ng/ml, p<0.01). The MMP-9 levels were low and below detection limit in all PXF and control samples with no statistical difference between groups. CONCLUSION: A raised TIMP-1 level and a low MMP-9 level in aqueous humour of PXF eyes may imply a downregulation in proteolytic activity. The increased CTGF concentration supports the proposed fibrotic pathology of PXF. Regulation of MMP/TIMP expression and anti-CTGF therapy may offer potential therapeutic avenues for controlling PXF associated ocular morbidity. PMID- 15665349 TI - Probing glaucoma visual damage by rarebit perimetry. AB - AIM: To compare rarebit perimetry (RBP) with standard achromatic perimetry (SAP) in detecting early glaucomatous functional damage. METHODS: 43 patients with ocular hypertension (OH), 39 with early primary open angle glaucoma (POAG), and 41 controls were considered. Visual fields were assessed using the Humphrey field analyser (HFA) 30-2 and RBP tests. Differences among the groups were evaluated using Student-Newman-Keuls and chi(2) tests. Correlation between HFA and RBP parameters was assessed using the Pearson's correlation coefficients and regression analysis. Sensitivity and specificity of RBP in detecting early glaucomatous visual damage were calculated with different algorithms. RESULTS: RBP-mean hit rate (MHR) was respectively 88.6% (SD 4.8%) in controls; 79.1% (10.9%) in the OH group; 64.3% (13.8%) in the POAG group (differences statistically significant). Good correlation in the POAG group was found between HFA-mean deviation and RBP-MHR. Largest AROC (0.95) and optimal sensitivity (97.4%) were obtained when an abnormal RBP test was defined as having (at least 1): MHR <80%; >15 areas with a non-hit rate of >10%; > or =2 areas with a non-hit rate of >50%; at least one area with a non-hit rate of > or =70%. CONCLUSIONS: The RBP appeared to be a rapid, comfortable, and easily available perimetric test (requiring only a PC device), showing a high sensitivity and specificity in detecting early glaucomatous visual field defects. PMID- 15665350 TI - Effect of trabeculectomy on ocular blood flow. AB - BACKGROUND/AIM: Current evidence suggests that vascular insufficiencies in the optic nerve head play an important part in the pathogenesis of glaucomatous optic neuropathy. Trabeculectomy is the most common operative procedure for the treatment of medically uncontrolled glaucoma. This study was conducted to investigate whether trabeculectomy may improve ocular haemodynamics. METHODS: 30 patients with primary open angle glaucoma about to undergo trabeculectomy were included in the study. Patients were evaluated before surgery and at 2 and 10 weeks after trabeculectomy. Optic nerve head blood flow (OnhBF) was assessed with scanning laser Doppler flowmetry. Fundus pulsation amplitude (FPA) measurements were obtained with laser interferometry. RESULTS: Because of the decrease in intraocular pressure there was a significant increase in ocular perfusion pressure (OPP) following trabeculectomy (18.5% (SD 12.0%) and 19.0% (17.1%) at 2 and 10 weeks postoperatively; p <0.001). A significant increase in OnhBF was observed after trabeculectomy (11.6% (16.4%) and 16.2% (20.2%) for each postoperative visit, respectively; p <0.001). FPA was also significantly higher compared with baseline values (17.2% (17.3%) and 17.4% (16.3%), respectively; p <0.001). A significant association between the increase in OPP and the increase in OnhBF and FPA was observed 10 weeks after surgery (r = 0.47; p = 0.009, and r = 0.50; p = 0.005, respectively). CONCLUSION: The results of this study suggest that trabeculectomy improves ocular blood flow in patients with chronic open angle glaucoma. PMID- 15665348 TI - Optic nerve compression by normal carotid artery in patients with normal tension glaucoma. AB - AIM: To determine whether compression of the optic nerve by the intracranial carotid artery (ICA) can be a causative factor of normal tension glaucoma (NTG). METHODS: The medical records of 103 eyes of 54 Japanese patients with NTG and 104 eyes of 52 age matched control patients were reviewed. The neuroradiological findings of magnetic resonance images (MRI) were evaluated to determine the relation between the optic nerve and ICA. The clinical characteristics and general medical conditions, such as diabetes and systemic hypertension, were also compared between the two groups. RESULTS: The prevalence of optic nerve compression by the ICA in patients with NTG was 49.5%, which was significantly higher than that in control group with 34.6% (p = 0.035). Bilateral compression of the optic nerve was detected in 22 patients with NTG (40.7%), and this was also significantly higher (p = 0.029) than that in the control group (11 patients, 21.2%). In the NTG group, eyes with cup/disc ratio (C/D ratio) > or =0.7 showed a higher percentage of compression (52.6%) compared with eyes with C/D ratio of <0.7 (12.5%; p = 0. 042). The presence of diabetes and hypertension did not affect the incidence of optic nerve compression by ICA significantly. CONCLUSIONS: The significantly higher percentage of NTG patients who had optic nerve compression by the ICA suggests that compression of the optic nerve by ICA may be a possible causative factor or a risk factor for optic nerve damage in some patients with NTG. PMID- 15665351 TI - Macular microholes: pathogenesis and natural history. AB - AIMS: To study the natural history and evaluate optical coherence tomography (OCT) and the retinal thickness analyser (RTA) in patients with macular microholes. METHODS: The medical records of 22 patients with a well demarcated red intraretinal foveal or juxtafoveal defect were reviewed. Fluorescein angiography (FA), RTA, and OCT were performed. The main outcome measures were visual acuity (VA), and OCT and RTA characteristics of microholes. Long term follow up was available in 13 eyes of 12 patients. RESULTS: The patients had a mean age of 50 years and a mean refractive error of -0.93 dioptres. The presenting symptom was a central scotoma in 14 eyes and metamorphopsia in eight eyes. All patients had a corrected VA ranging from 20/16 to 20/125, with 20 out of 24 eyes (83%) having a VA > or =20/40. Symptoms remained stable or improved in 16 out of 22 patients (72%). OCT 2 findings were normal but an abnormality of the outer retina and/or a defect of the retinal pigment epithelium (RPE) were demonstrated on OCT 3 in 15 of 18 eyes (83%). The RTA topographic map demonstrated a defect at the site of the microhole in two out of 12 eyes. CONCLUSION: Although biomicroscopic examination suggested an inner foveal defect, the OCT 3 scans demonstrated a localised abnormality of the outer retina and/or RPE which could not be resolved using OCT 2. Macular microholes have a favourable long term prognosis with stable VA. Bilateral involvement is uncommon. PMID- 15665352 TI - Further evidence of genetic heterogeneity in familial exudative vitreoretinopathy; exclusion of EVR1, EVR3, and EVR4 in a large autosomal dominant pedigree. AB - BACKGROUND/AIMS: Familial exudative vitreoretinopathy (FEVR) is an inherited blinding condition characterised by abnormal development of the retinal vasculature. The aim of this study was to perform linkage analysis in a large family affected with FEVR to determine whether the mutation involved was in one of the three known autosomal dominant FEVR loci or in another as yet unidentified gene. METHODS: Genomic DNA samples from family members were polymerase chain reaction (PCR) amplified with fluorescently tagged microsatellite markers spanning the EVR1/EVR4 locus (11q13-14) and the EVR3 locus (11p12-13). The resulting PCR products were resolved using an automated DNA sequencer and the alleles sized. These data were used to construct haplotypes across each locus and linkage analysis was performed to prove or exclude linkage. RESULTS: The clinical evaluation in this family suggested features typical of FEVR, with deficient peripheral retinal vascularisation being the common phenotype in all affected individuals. However, linkage analysis proved that this family has a form of FEVR genetically distinct from the EVR1, EVR3 and EVR4 loci. CONCLUSION: The exclusion of linkage in this family to any of the known FEVR loci proves the existence of a fourth locus for autosomal dominant FEVR and shows that this rare disorder is far more heterogeneous than previously thought. PMID- 15665354 TI - Enhanced optical coherence tomography imaging by multiple scan averaging. AB - AIMS: To describe a method for computerised alignment and averaging of sequences in optical coherence tomography (OCT) B-scans and to present selected clinical observations based on the resulting improvement in retinal imaging. METHODS: A methodological study and retrospective investigation of selected cases. Five human subjects were included, one healthy subject, two patients with central serous chorioretinopathy, one patient with branch retinal vein occlusion, and one patient with cilioretinal artery pseudo-occlusion. Based on computerised alignment of sets of B-scans obtained at identical retinal locations, average OCT images were produced and displayed in false colour or grayscale. These enhanced tomograms were compared with other morphological and functional characteristics. RESULTS: Improved retinal imaging enabled assignment of the OCT image to retinal anatomy particularly at the outer layer of the photoreceptors and the retinal pigment epithelium, both in the healthy eye and in pathology. Identification of both post-oedematous structural disorganisation as well as post-ischaemic attenuation of the inner retina was superior to standard OCT images. CONCLUSIONS: Averaging of multiple OCT B-scans enhances the quality of retinal imaging sufficiently to reveal new details of retinal pathophysiology. Using the technique on OCT3 scans enables visualisation of details comparable with the results obtained using ultra high resolution OCT. PMID- 15665353 TI - A detailed study of the phenotype of an autosomal dominant cone-rod dystrophy (CORD7) associated with mutation in the gene for RIM1. AB - AIM: To characterise the phenotype of an autosomal dominant cone-rod dystrophy (CORD7) associated with the Arg844His mutation in RIM1. METHODS: Eight members of a four generation, non-consanguineous British family were examined clinically and underwent electrophysiological testing, automated dark adapted perimetry, dark adaptometry, colour vision assessment, colour fundus photography, fundus fluorescein angiography (FFA), and fundus autofluorescence (AF) imaging. RESULTS: The majority of affected individuals described a progressive deterioration of central vision, night vision, and peripheral visual field usually between the third and fourth decades. The visual acuity ranged from 6/6 to 3/60. Colour vision testing showed mild to moderate dyschromatopsia in the majority of individuals. Fundus changes comprised a range of macular appearances varying from mild retinal pigment epithelial (RPE) disturbance to extensive atrophy and pigmentation. In some individuals retinal vessels were attenuated and in two subjects peripheral areas of retinal atrophy were present. An absent or severely reduced PERG was detected in all subjects, indicative of marked macular dysfunction. Full field ERG showed abnormal rod and cone responses. AF imaging revealed decreased macular AF centrally surrounded by a ring of increased AF in the majority of individuals. "Bull's eye" lesions were present in two individuals, comprising of a ring of decreased perifoveal AF bordered peripherally and centrally by increased AF. Photopic sensitivity testing demonstrated elevated central visual field thresholds with additional superior greater than inferior peripheral field loss. There were rod and cone sensitivity reductions in the central and peripheral visual fields, with the inferior retina being more affected than the superior. CONCLUSIONS: The detailed phenotype is described of the autosomal dominant cone-rod dystrophy, CORD7, which is associated with a point mutation in RIM1, a gene encoding a photoreceptor synaptic protein. The pattern of disease progression and long term visual outcome facilitates improved genetic counselling and advice on prognosis. Such phenotypic data will be invaluable in the event of future therapy. PMID- 15665355 TI - Health services experiences of parents of recently diagnosed visually impaired children. AB - AIM: To investigate the health service experiences and needs of parents in the period around diagnosis of ophthalmic disorders in their children. METHODS: Parents of children newly diagnosed with visual impairment and/or ophthalmic disorders at a tertiary level hospital in London participated in a questionnaire survey, using standard instruments, followed by in-depth individual interviews, to elicit their views about the processes of care, their overall level of satisfaction, and their unmet needs. RESULTS: 67% (147) of eligible families (135 mothers, 76 fathers) participated. Overall satisfaction with care was high, being greater among parents of children with milder visual loss or isolated ophthalmic disorders than those with more severe visual loss or multiple impairments. Nevertheless, parents' reported greatest need was the provision of general information, including about their child's ophthalmic disorder and educational and social services and support. Mothers reported greater information needs than fathers, as did white parents compared to those from ethnic minorities. White parents also regarded the processes of care to be less comprehensive and coordinated, as well as less enabling, than did parents from ethnic minorities. CONCLUSIONS: Although parents reported high overall satisfaction with services, improving the medium, content, and scope of general information provided by professionals to parents of visually impaired children emerges as a priority. Equitable planning and provision of health services for families of children with visual impairment needs to take into account that informational and other needs vary by whether the parent is the primary carer or not and their ethnicity, as well as by the severity and complexity of their child's visual loss. PMID- 15665356 TI - Incidence of host site complications in periocular full thickness skin grafts. AB - AIM: To evaluate the complications of periocular full thickness skin grafts (FTSG) in patients treated with Mohs' micrographic surgery (MMS) for periocular malignancy. METHOD: This prospective, multicentre case series included all patients in Australia treated with MMS for periocular malignancy followed by reconstruction with FTSG, who were monitored by the Skin and Cancer Foundation, between 1993 and 1999. The parameters recorded were patient demographics, reason for referral, histological classification of malignancy and evidence of perineural invasion, duration of tumour, site, recurrences prior to MMS, preoperative tumour size, and postoperative defect size. FTSG donor sites included upper lid, preauricular, retroauricular, inner brachial, and supraclavicular. The primary outcome measures were FTSG host site complications (partial/complete graft failure, graft infection, acute bleeding/haematoma, graft hypertrophy, and graft contracture). RESULTS: 397 patients (229 males, 168 females), mean age 60 (SD 15) years (range 20-91 years). 92.7% were diagnosed with basal call carcinoma, 2.0% with Bowen's disease, and 3.3% with squamous cell carcinoma. Medial canthus was involved in 66.5% of patients, lower eyelid in 28.0%, and upper eyelid in 5.5%. Postoperative complications were recorded in 62 patients (15.6% of all patients), and consisted of graft hypertrophy (45.1% of complications), graft contraction (29.1%), and partial graft failure (12.9%). The only statistically significant association found was a higher rate of graft hypertrophy in medial canthal reconstruction (p = 0.007). CONCLUSION: Host site complications of periocular FTSG are not common. Graft hypertrophy accounted for most complications and was more common in the medial canthal area. No other variables such as patient demographics, tumour characteristics, or donor site factors were associated with a higher risk of complications. PMID- 15665357 TI - Enhanced endothelium derived hyperpolarising factor activity in resistance arteries from normal pressure glaucoma patients: implications for vascular function in the eye. AB - BACKGROUND/AIMS: Endothelial cell dysfunction in the ocular circulation may contribute to normal pressure glaucoma (NPG). This study aimed to investigate the contributions made by endothelium derived relaxing factors to relaxation of (1) subcutaneous resistance arteries from patients with NPG, and (2) porcine ciliary arteries. METHODS: Human gluteal resistance arteries were isolated from seven patients with NPG and matched controls. Human and porcine arteries produced endothelium dependent relaxation when exposed to acetylcholine (ACh) (10(-9)-3 x 10(-5)M) or bradykinin (10(-10)-3 x 10(-6)M). Pharmacological agents were used to inhibit the nitric oxide pathway (l-arginine analogues, soluble guanylate cyclase inhibitor), endothelium derived hyperpolarising factor (EDHF) activity (potassium channel antagonists), and prostaglandin synthesis (cyclo-oxygenase inhibitors). RESULTS: In all arteries, endothelium dependent relaxation was attenuated by nitric oxide (NO) inhibition or potassium channel blockade, but not by cyclo oxygenase inhibition. Inhibition of ACh mediated relaxation by potassium channel antagonists was greater (p<0.05) in patients with NPG (Emax, 55.4% (SD 8.16%) relaxation, n = 4) than controls (Emax, 81.8% (6.0%), n = 5). In contrast, combined inhibition of NO synthase (NOS) and cyclo-oxygenase produced similar inhibition of ACh mediated relaxation in both groups. CONCLUSIONS: The enhanced contribution of EDHF to ACh mediated relaxation in systemic resistance arteries from NPG patients may contribute to the maintained endothelium mediated relaxation in these vessels. EDHF also contributes significantly to bradykinin mediated relaxation in porcine ocular ciliary arteries. Therefore, similar changes in the balance of relaxing factors in the ocular circulation could influence the response of the eye to vascular endothelial dysfunction in NPG. PMID- 15665359 TI - Demonstration of identical clonal derivation in a case of "oculocerebral" lymphoma. PMID- 15665358 TI - Why cotton wool spots should not be regarded as retinal nerve fibre layer infarcts. AB - Cotton wool spots (CWSs) comprise localised accumulations of axoplasmic debris within adjacent bundles of unmyelinated ganglion cell axons. Their formation is widely held to reflect focal ischaemia from terminal arteriolar occlusion, but credible evidence supporting this view is lacking. CWSs are here purported to be nothing more than sentinels of retinal nerve fibre layer pathology, hence their recommended redesignation "cotton wool sentinels." After branch arteriolar occlusion, CWSs evolve as boundary sentinels of infarction, their uniform width suggesting a glial constraint to axonal expansion. In pre-proliferative diabetic retinopathy, CWSs form a C-shaped chain nasal to the disc and around the macula where they constitute sentinels of ischaemia affecting the entire retinal mid periphery. The polymorphous CWSs evolving during acute panretinal hypoperfusion represent sentinels of an ischaemic penumbra. Those surrounding the disc in Purtscher's traumatic angiopathy are sentinels of neuronal damage from transient venous hyperdistension that overwhelms the protection afforded by peripapillary axonal decompartmentalisation. PMID- 15665361 TI - Subconjunctival migration of silicone oil through a Baerveldt pars plana glaucoma implant. PMID- 15665360 TI - The prevalence of pseudoexfoliation syndrome in Chinese people: the Tanjong Pagar Survey. PMID- 15665362 TI - Hoarse voice and visual loss. PMID- 15665363 TI - Rosai-Dorfman disease: isolated epibulbar masses in two adult patients. PMID- 15665364 TI - Systemic FK506 improved tear secretion in dry eye associated with chronic graft versus host disease. PMID- 15665365 TI - 3D ultrasound coronal C-scan imaging for optic nerve sheath meningioma. PMID- 15665366 TI - An infected hydrogel buckle with Corynebacterium pseudotuberculosis. PMID- 15665367 TI - A deficit in visits to the optometrist by preschool age children: implications for vision screening. PMID- 15665368 TI - "Only rarely seen in dreams"--visual experiences during cataract surgery. PMID- 15665369 TI - Spontaneous closure of microaneurysms in diabetic retinopathy with treatment of co-existing anaemia. PMID- 15665370 TI - Ciprofloxacin in endophthalmitis: an alternative to ceftazidime and amikacin! PMID- 15665371 TI - In answer to "Who is Ivan Schwab?". PMID- 15665372 TI - Severe ocular trauma caused by an ostrich. PMID- 15665373 TI - Identification of silicone oil in ocular tissues. PMID- 15665374 TI - No evidence for severe retinopathy of prematurity following sildenafil. PMID- 15665375 TI - Cost accounting in cost utility analysis of screening and treatment. PMID- 15665377 TI - Quantitative phosphoproteomics applied to the yeast pheromone signaling pathway. AB - Cellular processes such as proliferation, differentiation, and adaptation to environmental changes are regulated by protein phosphorylation. Development of sensitive and comprehensive analytical methods for determination of protein phosphorylation is therefore a necessity in the pursuit of a detailed molecular view of complex biological processes. We present a quantitative modification specific proteomic approach that combines stable isotope labeling by amino acids in cell culture (SILAC) for quantitation with IMAC for phosphopeptide enrichment and three stages of mass spectrometry (MS/MS/MS) for identification. This integrated phosphoproteomic technology identified and quantified phosphorylation in key regulator and effector proteins of a prototypical G-protein-coupled receptor signaling pathway, the yeast pheromone response. SILAC encoding of yeast proteomes was achieved by incorporation of [(13)C(6)]arginine and [(13)C(6)]lysine in a double auxotroph yeast strain. Pheromone-treated yeast cells were mixed with SILAC-encoded cells as the control and lysed, and extracted proteins were digested with trypsin. Phosphopeptides were enriched by a combination of strong cation exchange chromatography and IMAC. Phosphopeptide fractions were analyzed by LC-MS using a linear ion trap-Fourier transform ion cyclotron resonance mass spectrometer. MS/MS and neutral loss-directed MS/MS/MS analysis allowed detection and sequencing of phosphopeptides with exceptional accuracy and specificity. Of more than 700 identified phosphopeptides, 139 were differentially regulated at least 2-fold in response to mating pheromone. Among these regulated proteins were components belonging to the mitogen-activated protein kinase signaling pathway and to downstream processes including transcriptional regulation, the establishment of polarized growth, and the regulation of the cell cycle. PMID- 15665378 TI - Cellular fate of truncated slow skeletal muscle troponin T produced by Glu180 nonsense mutation in amish nemaline myopathy. AB - A nonsense mutation at codon Glu180 in exon 11 of slow skeletal muscle troponin T (TnT) gene (TNNT1) causes an autosomal-recessive inherited nemaline myopathy. We previously reported the absence of intact or prematurely terminated slow TnT polypeptide in Amish nemaline myopathy (ANM) patient muscle. The present study further investigates the expression and fate of mutant slow TnT in muscle cells. Intact slow TnT mRNA was readily detected in patient muscle, indicating unaffected transcription and RNA splicing. Sequence of the cloned cDNAs revealed the single nucleotide mutation in two alternatively spliced isoforms of slow TnT mRNA. Mutant TNNT1 cDNA is translationally active in Escherichia coli and non muscle eukaryotic cells, producing the expected truncated slow TnT protein. The mutant mRNA was expressed at significant levels in differentiated C2C12 myotubes, but unlike intact exogenous TnT, truncated slow TnT protein was not detected. Transfective expression in undifferentiated myoblasts produced slow TnT mRNA but not a detectable amount of truncated or intact slow TnT proteins, indicating a muscle cell-specific proteolysis of TnT when it is not integrated into myofilaments. The slow TnT-(1-179) fragment has substantially lower affinity for binding to tropomyosin, in keeping with the loss of one of two tropomyosin binding sites. Our findings suggest that inefficient incorporation into myofilament is responsible for the instability of mutant slow TnT in ANM muscle. Rapid degradation of the truncated slow TnT protein, rather than instability of the nonsense mRNA, provides the protective mechanism against the potential dominant negative effect of the mutant TnT fragment. PMID- 15665379 TI - The EB1 homolog Mal3 stimulates the ATPase of the kinesin Tea2 by recruiting it to the microtubule. AB - Tea2 is a kinesin family member from Schizosaccharomyces pombe that is targeted to microtubule tips and cell ends in a process that depends on Mal3. Constructs of Tea2 containing the motor domain only or the motor domain plus the N-terminal extension are monomeric, whereas a construct including the first predicted coiled coil region is dimeric. These constructs have a low basal rate of ATP hydrolysis of <0.1 s(-1), but microtubules stimulate the rate of ATP hydrolysis to a maximum of approximately 15 s(-1). Hydrodynamic analysis of Mal3 indicates that it is dimeric. Mal3 is known to associate with Tea2, and analysis with the above Tea2 constructs indicates that the principal site of interaction of Mal3 with Tea2 is the N-terminal extension, although a weaker interaction is also observed with the motor domain alone. In parallel to the binding studies, Mal3 strongly stimulates the ATPase of constructs containing the N-terminal extension by decreasing the K0.5(MT) for stimulation by microtubules but only weakly stimulates motor domains without the N-terminal extension. Mal3 reduces the K0.5(MT) values without affecting the k(cat) value at saturating microtubule level. Binding of Mal3 to microtubules induces an increase in the binding of Tea2 and a reciprocal stimulation of Mal3 binding by Tea2 is also observed. Tea2 is a plus end directed motor that drives sliding of axonemes when adsorbed to a glass surface. The sliding rate is initially unaffected by Mal3, but axonemes stop moving on continued exposure to Mal3. PMID- 15665380 TI - Monastrol inhibition of the mitotic kinesin Eg5. AB - Monastrol is a small, cell-permeable molecule that arrests cells in mitosis by specifically inhibiting Eg5, a member of the Kinesin-5 family. We have used steady-state and presteady-state kinetics as well as equilibrium binding approaches to define the mechanistic basis of S-monastrol inhibition of monomeric human Eg5/KSP. In the absence of microtubules (Mts), the basal ATPase activity is inhibited through slowed product release. In the presence of microtubules, the ATPase activity is also reduced with weakened binding of Eg5 to microtubules during steady-state ATP turnover. Monastrol-treated Eg5 also shows a decreased relative affinity for microtubules under equilibrium conditions. The Mt.Eg5 presteady-state kinetics of ATP binding and the subsequent ATP-dependent isomerization are unaffected during the first ATP turnover. However, monastrol appears to stabilize a conformation that allows for reversals at the ATP hydrolysis step. Monastrol promotes a dramatic decrease in the observed rate of Eg5 association with microtubules, and ADP release is slowed without trapping the Mt.Eg5.ADP intermediate. We propose that S-monastrol binding to Eg5 induces a stable conformational change in the motor domain that favors ATP re-synthesis after ATP hydrolysis. The aberrant interactions with the microtubule and the reversals at the ATP hydrolysis step alter the ability of Eg5 to generate force, thereby yielding a nonproductive Mt.Eg5 complex that cannot establish or maintain the bipolar spindle. PMID- 15665381 TI - Caveolae targeting and regulation of large conductance Ca(2+)-activated K+ channels in vascular endothelial cells. AB - The vascular endothelium is richly endowed with caveolae, which are specialized membrane microdomains that facilitate the integration of specific cellular signal transduction processes. We found that the large conductance Ca(2+)-activated K+ (BK) channels are associated with caveolin-1 in bovine aortic endothelial cells (BAECs). OptiPrep gradient cell fractionation demonstrated that BK channels were concentrated in the caveolae-rich fraction in BAECs. Immunofluorescence imaging showed co-localization of caveolin-1 and BK channels in the BAEC membrane. Immunoprecipitation and glutathione S-transferase pull-down assay results indicated that caveolin-1 and BK channels are physically associated. However, whole cell patch clamp recordings could not detect BK (iberiotoxin-sensitive) currents in cultured BAECs under baseline conditions, even though the presence of BK mRNA and protein expression was confirmed by reverse transcription-PCR and Western blots. Cholesterol depletion redistributed the BK channels to non caveolar fractions of BAECs, resulting in BK channel activation (7.3 +/- 1.6 pA/picofarad (pF), n = 5). BK currents were also activated by isoproterenol (ISO, 1 microM, 6.9 +/- 2.4 pA/pF, n = 6). Inclusion of a caveolin-1 scaffolding domain peptide (10 microM) in the pipette solution completely abrogated the effects of ISO on BK channel activation, whereas inclusion of the scrambled control peptide (10 microM) did not inhibit the ISO effects. We have also found that caveolin-1 knockdown by small interference RNA activated BK currents (5.3 +/- 1.4 pA/pF, n = 6). We conclude that: 1) BK channels are targeted to caveolae microdomains in vascular endothelial cells; 2) caveolin-1 interacts with BK channels and exerts a negative regulatory effect on channel functions; and 3) BK channels are inactive under control conditions but can be activated by cholesterol depletion, knockdown of caveolin-1 expression, or ISO stimulation. These novel findings may have important implications for the role of BK channels in the regulation of endothelial function. PMID- 15665382 TI - The effect of estrogens and phytoestrogenic lignans on macrophage uptake of atherogenic lipoproteins. AB - The present study examined the effect of estrogens and compounds with estrogenic activity on the uptake of atherogenic lipoproteins into macrophages, thought to be the initiating step in the development of atherosclerotic lesions. Isolated low density lipoprotein (LDL) and lipoprotein(a) (Lp(a)) were radiolabelled with (3)H-cholesterol linoleate, and incubated with J774 macrophages for 24 hours in the presence of pharmacological doses of estrogens and phytoestrogens. At a concentration of 0.1 microM, the estrogen 17beta-estradiol significantly reduced LDL uptake by macrophages by 14% (p < 0.05), but estrone did not have any effect. At 10 microM, both estrogens significantly reduced macrophage LDL uptake, but the phytoestrogenic-lignans enterodiol and enterolactone had no effect on LDL uptake. Lp(a) uptake into cells was significantly reduced by both estrone and estradiol, and by enterolactone and enterodiol at concentrations of 10 microM (p < 0.01), with enterodiol being most effective. The results of this study suggest that the uptake of these structurally similar lipoproteins is regulated differently. Macrophage Lp(a) uptake appears more phytoestrogen sensitive than does LDL uptake. PMID- 15665383 TI - Oxidative status of human milk and its variations during cold storage. AB - Breastfeeding and human milk are widely accepted as optimal for human infants' nutrition. Nowadays lifestyle often makes it difficult to maintain or even initiate human lactation. This situation is mostly related to the workload of women away from home. New approaches are needed to enable maternal lactation under these circumstances. Human breastmilk storage for differed use is one possibility. The aim of this study was to assess changes in glutathione peroxidase (GPx) activity and in the concentration of the lipid peroxidation marker, malondialdehyde (MDA), when human milk was kept refrigerated or frozen. Thirty-two human milk samples were assayed for GPx activity and MDA concentration. Samples were divided in three aliquot portions, the first to be immediately analysed, the second to be refrigerated at 4 degrees C and analysed 24 h thereafter, and the third to be frozen at -20 degrees C and assayed after 10 days. GPx activity was significantly decreased in refrigerated and in frozen milk, when compared to their control samples. MDA was increased only in refrigerated milk but not in frozen samples. Thus, freezing seems better than refrigeration in order to prevent lipid peroxidation in stored human milk samples. PMID- 15665384 TI - Study of factors affecting binding of zinc with albumin at physiological zinc concentrations. AB - Albumin has very high affinity for many organic and inorganic compounds that may influence albumin bound Zn (ABZn). To get insight of these molecular interactions, the effect of riboflavin, nicotinic acid, thiamine, folic acid, pyruvic acid and glucose on ABZn were studied. The ABZn was separated from the unbound zinc using equilibrium dialysis and estimated using atomic absorption spectrometer. At therapeutic zinc concentrations, folic acid and thiamine significantly enhanced the ABZn (p < 0.010), while nicotinic acid inhibited zinc binding to albumin. Folic acid was found to enhance the ABZn also at lower zinc concentrations representing physiological levels of plasma zinc (138-150 micromoles) (p < 0.05). PMID- 15665385 TI - In vitro protective effect of Rhodiola rosea extract against hypochlorous acid induced oxidative damage in human erythrocytes. AB - Rhodiola rosea L. (Crassulaceae) is a plant living at high altitudes in Europe and Asia. Its roots have long been used in the traditional medical system of these geographical areas to increase the organism resistance to physical stress; today, it has become an important component of many dietary supplements. In this study we investigate the antioxidant capacity of the R. rosea aqueous extract evaluating its ability to counteract some of the main damages induced by hypochlorous acid (HOCl), a powerful oxidant generated by activated phagocytes, to human erythrocytes. Ascorbic acid was used as a reference substance because of its physiological HOCl-scavenging ability. Our study demonstrates that R. rosea is able to significantly protect, in a dose-dependent manner, human RBC from glutathione (GSH) depletion, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) inactivation and hemolysis induced by the oxidant. Furthermore, we demonstrate that R. rosea aqueous extract acts from the inside of the erythrocyte suggesting a probable involving of cell components. The protection on GSH afforded by the R. rosea extract with respect to ascorbic acid, occurred also if added 2 or 5 min. later than the oxidant, suggesting a more rapid or powerful effect. PMID- 15665386 TI - Dietary fat composition modifies the effect of boron on bone characteristics and plasma lipids in rats. AB - Female and male rats weighing about 170 g and 200 g, respectively, were fed diets (approximately 70 microg boron/kg) in a factorial arrangement with supplemental boron at 0 (deficient) and 3 (adequate) mg/kg and canola oil or palm oil at 75 g/kg of diet as variables. After 5 weeks, six females in each treatment were bred. Dams and pups continued on their respective dietary treatments through gestation, lactation and post-weaning. Thirteen weeks after weaning, plasma and bones were collected from 12 male and 12 female offspring in each treatment. Boron supplementation increased femur strength measured by the breaking variable bending moment; tibial calcium and phosphorus concentrations; and plasma alkaline phosphatase. Femur breaking stress was greatest in boron-supplemented rats fed canola oil, and lowest in boron-deprived females fed canola oil; this group also exhibited the lowest femur bending moment. Minerals associated with bone organic matrix, zinc and potassium, were increased by boron supplementation in tibia. Plasma phospholipids were decreased by boron deprivation in females, but not males. Plasma cholesterol was decreased in boron-supplemented males by replacing canola oil with palm oil. The findings suggest that a diet high in omega-3 alpha linolenic acid promotes femur strength best when the dietary boron is adequate. PMID- 15665387 TI - Plasma levels and redox status of coenzyme Q10 in infants and children. AB - INTRODUCTION: Increased attention has been paid to the role of lipophilic antioxidants in childhood nutrition and diseases during recent years. The lipophilic antioxidant coenzyme Q10 (CoQ10) is known as an effective inhibitor of oxidative damage. In contrast to other lipophilic antioxidants like alpha tocopherol the plasma concentrations of CoQ10 in childhood are poorly researched. The aim of this study was to determine plasma level and redox status (oxidized form in total CoQ10 in %) of CoQ10 in clinically healthy infants, preschoolers and school-aged children. METHODS: Plasma level and redox status of CoQ10 were measured by HPLC in 199 clinically healthy children, three groups of infants [1st 4th month (n = 35), 5th-8th month (n = 25), 9th-12th month (n = 25) ], preschoolers (n = 60) and school-aged children (n = 54). The CoQ10 plasma levels were related to plasma cholesterol concentrations. The median and the 5th and 95th percentile were calculated. RESULTS: Plasma levels and redox status of CoQ10 in infants were significantly higher than in preschoolers and school-aged children. The CoQ10 redox status in the 1st-4th month was significantly increased when compared to the remaining subgroups of infants. In elder children the CoQ10 redox status stabilized. CONCLUSIONS: This is the first study concerning age related values of plasma level and redox status of CoQ10 in apparently healthy children. Decreased CoQ10 values could be involved in various pathological conditions affecting childhood. Therefore, the application of age-adjusted reference values may provide more specific criteria to define threshold values for CoQ10 deficiency in plasma. PMID- 15665388 TI - The matrix metalloproteinase-3 (MMP-3) 5A/6A promoter polymorphism is not associated with ischaemic heart disease: analysis employing a family based approach. AB - Matrix metalloproteinase-3 (MMP-3) has been proposed as an important mediator of the atherosclerotic process. The possible role of the functional -1612 5A/6A polymorphism of the MMP-3 gene in the susceptibility to ischaemic heart disease (IHD) was investigated in a well-defined Irish population using two recently described family based tests of association. One thousand and twelve individuals from 386 families with at least one member prematurely affected with IHD were genotyped. Using the combined transmission disequilibrium test (TDT)/sib-TDT and the pedigree disequilibrium test (PDT), no association between the MMP-3 -1612 5A/6A polymorphism and IHD was found. Our data demonstrate that, in an Irish population, the MMP-3 -1612 5A/6A polymorphism is not associated with IHD. PMID- 15665389 TI - Measurement reproducibility in the early stages of biomarker development. AB - Biomarker discovery and development requires measurement reproducibility studies in addition to case-control studies. Parallel pursuit of reproducibility studies is especially important for emerging technologies such as protein biomarkers based on time-of-flight mass spectrometry, the case considered in this paper. For parallel studies, a way to improve reproducibility prior to identification of protein species is necessary. One approach is use of functional principal components analysis (PCA) as the basis for assessing measurement reproducibility. Reproducibility studies involve repeated measurement of a reference material such as a human serum standard. Measurement in our example is by SELDI-TOF (surface enhanced laser desorption and ionization time-of-flight) mass spectrometry. Reproducibility is defined in reference to a source of variation, which in our example is associated with a type of commercially available protein biochip. We obtained spectra for 8 spots on each 11 chips. Two spectra are generally more alike when obtained from the same chip rather than different chips. Thus, our experiment indicates potential improvements from reducing variation in chip manufacture and chip handling during measurement. Our analysis involves careful registration of the spectra and characterization of the spectral differences. As shown by our example, a metrological analysis may enhance case-control studies by guiding optimization of the measurements underlying the biomarker. PMID- 15665390 TI - HLA B-27 subtypes in Turkish patients with spondyloarthropathy and healthy controls. AB - The frequency and the distribution of HLA-B27 subtypes in spondylarthropathy (SpA) patients and controls were investigated in a sample Turkish population. B27 subtyping was performed by PCR-SSP method in two groups: 49 unrelated HLA-B27 positive Turkish patients with the diagnosis of SpA according to the European Spondyloarthropathy Study Group Criteria, and 55 HLA-B27 positive healthy controls. The frequency of HLA-B*27 was 2.6% in the Turkish population, and B*2705 was the predominant allele among patients with SpA. The difference was mainly between male patients and male controls The proportion of B*2705 among B27 positive patients and controls was significantly different (P=0.02). Our study supports other reports from different populations which showed that B*2705 and B*2702 were more frequent in Caucasian patients with SpA. PMID- 15665391 TI - Mutation rate at commonly used forensic STR loci: paternity testing experience. AB - Paternity tests are carried out by the analysis of hypervariable short tandem repeat DNA loci. These microsatellite sequences mutate at a higher rate than that of bulk DNA. The occurrence of germline mutations at STR loci posses problems in interpretation of resulting genetic profiles. We recently analyzed 59-159 parent/child allele transfers at 13 microsatellite loci. We identified 12 mutations in 7 microsatellite loci. No mutations were occurred in other 6 loci. The highest mutation rate was observed with 5 mutations at D8S1179 locus at different alleles. The event was always single repeat related. The mutation rate was between 0 and 1.5 x 10(-2) per locus per gamete per generation. The mutation event is very crucial for forensic DNA testing and accumulation of STR mutation data is extremely important for genetic profile interpretation. PMID- 15665392 TI - Evaluation on the use of beta-lactamase and aminoglycoside modifying enzyme gene sequences as markers for the early detection of antibiotic resistance profile of Pseudomonas aeruginosa. AB - Pseudomonas aeruginosa is one of the major causes of infections including the hospital acquired (Nosocomial) infections. Detection of them and their antibiotic resistance profile by conventional method takes about three days. Recently, DNA based diagnostic methods are being used for the identification of the pathogens. Hence we have tested a rapid and sensitive method using DNA sequences as markers for detecting the presence of three genes coding for the enzymes that inactivate the two most commonly used Anti-pseudomonadal drugs such as beta-lactam antibiotics (Penicillin, and its derivatives) and Aminoglycosides such as Gentamicin, Tobramycin, Amikacin, Streptomycin. The internal region of these genes were used for designing and synthesizing primers and these primers were used in Polymerase Chain Reaction (PCR) to screen for the presence of these genes in the clinical isolates and to label them non-radioactively with Biotin. They in turn were used to detect the presence of the antibiotic resistance genes in the clinical isolates by hybridization. The specificity (ratio of positive results obtained in both methods and the sensitivity (the minimum amount of sample DNA and the labeled probe required for the tests) were evaluated. PMID- 15665393 TI - Mutations in transglutaminase 1 gene in autosomal recessive congenital ichthyosis in Egyptian families. AB - Autosomal recessive congenital ichthyosis (ARCI) is a rare heterogeneous keratinization disorder of the skin. It is clinically divided into 2 subtypes, lamellar ichthyosis (LI) and congenital ichthyosiformis erythroderma (CIE). We investigated forty-three ARCI Egyptian individuals in 16 severe LI, and 10 CIE families. We identified 5 alleles in two Egyptian families as having intron 5/exon-6 splice acceptor mutation recognized by the MspI restriction endonuclease. This promoted to a frequency of 9.6% for this mutation (5 splice mutation alleles/52 alleles tested). We extended our previous dataset to update the detection of R142H mutation in 4 CIE Egyptian families and one LI phenotype (frequency of 28.8%; 15/52), whereas we still had no R141H among our Egyptian population. There was no correlation between phenotype and genotype in our study. Surprisingly, the mutant alleles detected in intron-5 acceptor splice-site were associated with the other extreme of CIE phenotypes rather than the severe LI form. We clearly demonstrated that the ARCI Egyptian families in Upper Egypt was ethnically pure and had a tendency not to be a hybrid with other populations in Lower Egypt, Delta zone and Cairo city. PMID- 15665394 TI - Matrix metalloproteinase-2, squamous cell carcinoma antigen, and tissue polypeptide-specific antigen expression in Egyptian patients with cervical carcinoma: Relationship with prognosis. AB - Matrix metalloproteinases (MMPs), a family of proteolytic enzymes produced by both stromal and tumor cells, appear to have a key role in the events leading to local invasion and metastasis by malignant neoplasms. In the present study, we evaluated the role of MMP-2, squamous cell carcinoma antigen (SCCA), and tissue polypeptide - specific antigen (TPS) in cervical neoplasia. Using Western blotting and enzyme immunoassay (EIA), we analyzed 50 patients with cervical carcinoma (CC) and 25 normal controls for expression of MMP-2 in tissue cell lysates. We also quantified SCCA and TPS with microparticle immunoassay and EIA, respectively. The results were correlated with human papilloma virus (HPV) infection, clinicopathological findings, and disease outcome. The cutoff point for each marker was estimated from receiver operating characteristic curves. Logistic regression analysis was performed to estimate the odds ratio (OR) and 95% confidence interval (CI) for each marker. MMP-2, SCCA, and TPS protein expression were significantly higher in patients with CC than in normal controls. While TPS was the best marker for discriminating between patients and controls, MMP-2 was associated with an advanced tumor stage (OR, 13.9 [95% CI, 1.4-133.9]) and poor histological grade (OR, 10.2 [95% CI, 1.7-60.5]). Moreover, independent of the effect of an advanced CC stage and grade, the patients' age, and the presence of HPV infection, MMP-2 was considered a strong predictor for CC recurrence (OR, 8.1 [95% CI, 1.3- 49.1]). Tissue markers may be used to select high-risk patients for early detection of and adjuvant therapy for recurrence. Our MMP-2 findings are particularly relevant to the development of protease inhibitors as a new cancer therapy approach. PMID- 15665395 TI - Microparticles and a P-selectin-mediated pathway of blood coagulation. PMID- 15665396 TI - Tissue factor in the myocardium: evidence of roles in haemostasis and inflammation. AB - The interaction between cell-surface tissue factor (TF) and the plasma coagulation factor VII (FVII) initiates the coagulation network that leads to the generation of thrombin and the formation of a fibrin clot. Thrombin also activates cellular protease activated receptors (PARs) through which it activates components of the inflammatory pathway. TF is expressed constitutively by cardiomyocytes and evidence from mice transgenic for a human TF mini-gene that express very low levels of human TF suggests that the TF-FVII interaction is critical for haemostasis within the heart. Pathological contact between TF and FVII may occur in the heart during ischaemia-reperfusion (I-R) injury and this may lead to activation of coagulation and thrombin generation. Evidence from animal models now suggests that thrombin is an important mediator of inflammation in I-R injury. The coagulation pathway therefore represents a novel therapeutic target for intervention in the prevention of I-R injury. PMID- 15665397 TI - (+/-)-huprine Y, (-)-huperzine A and tacrine do not show neuroprotective properties in an apoptotic model of neuronal cytoskeletal alteration. AB - Acetylcholinesterase inhibitors (AChEI) are among the drugs most widely used in the treatment of Alzheimer's disease. They increase the levels of acetylcholine and thus improve the cognitive symptoms that are impaired. We tested whether specific AChEI show additional neuroprotective properties against colchicine induced apoptosis in cerebellar granule neurons (CGNs), a well established apoptotic model mediated by neuronal cytoskeleton alteration. Colchicine-induced apoptosis is due to an increase in the activity of GSK-3beta and CDK5, two enzymes involved in cytoskeletal alteration. Furthermore, the intrinsic apoptotic pathway is activated by colchicines, as revealed by cytochrome c release and Bax translocation. Tacrine, (-)-huperzine A and (+/-)-huprine Y, the AChEI tested in the study, did not reverse the loss of neuronal viability induced by colchicine. Moreover, the increase in apoptotic features induced by colchicine treatment, as measured by flow cytometry and nuclear chromatin condensation, was not prevented by these AChEI. Although some of these drugs are of interest to treat Alzheimer's disease, their lack of efficacy in the prevention of colchicine-induced apoptosis in CGNs suggests that they cannot prevent neuronal loss due to cytoskeleton alteration. PMID- 15665398 TI - Is there a characteristic lipid profile in Alzheimer's disease? AB - OBJECTIVE: To characterize the lipid profile in Alzheimer's Disease (AD) and to determine whether it differs from the cardiac risk profile. BACKGROUND: Links between hypercholesterolemia and AD development continue to grow. Presently, limited information exists about the lipid profile characteristics in AD. METHODS: We examined the lipid profiles (total cholesterol (TC), high-density lipoprotein (HDL), lower-density lipoprotein (LDL), TC/HDL ratio, and triglyceride (TG) levels) of 153 subjects with probable/possible AD (mean age 77.2 +/- 8.6 years, mean MMSE 19.9 +/- 5.6) and 25 non-demented subjects with atherosclerotic heart disease (ASHD) (mean age 73.8 +/- 7.2 years); neither on lipid lowering therapy. RESULTS: Subjects with TC > 200 mg/dl composed 69% of AD and 72% of ASHD groups. Mean TC was 218.9 +/- 38.9 mg/dl and 218.5 +/- 9.2 mg/dl for AD and ASHD subjects respectively. AD subjects exhibited significantly higher HDL and lower TG and TC/HDL ratios. MMSE did not correlate with any lipid parameters in AD. DISCUSSION: Elevated TC, LDL and TG with normal HDL and TC/HDL ratio characterize the lipid profile in AD, which somewhat overlaps with but may be distinct from the cardiac risk profile. MMSE does not correlate with lipid parameters suggesting no interaction between cholesterol and cognition in AD. PMID- 15665399 TI - Werner helicase polymorphism is not associated with Alzheimer's disease. AB - Alzheimer disease (AD) is the most common neurodegenerative disorder in the elderly and is also considered a progeroid genetic syndrome. The etiology of AD is complex and the mechanisms underlying its pathophysiology remains to be clarified. Werner syndrome (WS) is a rare autosomal recessive disorder characterized as a segmental progeroid syndrome. The gene (WRN) was recently identified. Its product acts as a DNA helicase and exonuclease. This study investigates the association of AD with the WRN 1367 polymorphisms in samples of 67 DA patients, 56 elderly healthy and 66 young healthy controls. DNA was isolated from blood cells, amplified by PCR and digested with PmaCI. We observed that the genotype distributions of WRN 1367 variants were within Hardy-Weinberg equilibrium in all subject samples. Furthermore, chi-square test comparison for genotype distributions and allele frequencies did not reveal any significant difference among the three groups of subjects (P>0.05). These results support the idea that these variants are not involved as a risk factor for developing AD. PMID- 15665400 TI - Associations of cortical astrogliosis with cognitive performance and dementia status. AB - We examined 204 decedents of the autopsy component of the Honolulu-Asia Aging Study, a longitudinal cohort study, who had been clinically assessed for dementia. A sensitive ELISA technique was used to quantify glial fibrillary acidic protein (GFAP), a marker for astrogliosis, in four specific cortical brain regions and assess associations between GFAP and 1) a measure of cognitive function, 2) several clinical dementia conditions, and 3) neuritic plaque (NP) and neurofibrillary tangle (NFT) formation. Cognitive function was inversely associated with GFAP in the occipital, parietal and temporal lobes, but not in the frontal lobe. This relationship remained significant when the contribution of NP and NFT counts was removed. Further, compared to brain samples from non demented individuals, significantly greater GFAP levels were found in samples from individuals diagnosed with Alzheimer's disease, mixed dementia, and vascular mediated dementia. Because elevated levels of GFAP reflect astroglial responses to even subtle forms of neural damage, our data indicate that increments in GFAP may provide independent, supporting evidence for the damage underlying dementia, even in the absence of other evidence of neuropathology such as the presence of NPs or NFTs. Our findings underscore the need to look beyond standard neuropathological measures putatively linked to specific neuropathological conditions in efforts to identify common cellular and molecular processes that contribute to dementia. PMID- 15665401 TI - Assembly of filamentous tau aggregates in human neuronal cells. AB - Intraneuronal deposition of microtubule-associated protein tau in filamentous aggregates constitutes a pathological hallmark of neurofibrillary degeneration that is characteristic of Alzheimer's disease (AD) and related disorders known collectively as tauopathies. Formation of such fibril inclusions, consisting of hyperphosphorylated tau in multiple isoforms, correlates with the severity of cognitive decline in AD. How neurofibrillary pathology evolves in tauopathy remains unclear at present, but availability of a cellular model with robust tau aggregation will permit experimental scrutiny of the mechanistic process leading to such neurodegeneration. Through the use of a serial transfection strategy in conjunction with a tau minigene construct, we succeeded in generating conditional transfectants of human neuronal lineage that overproduce wild-type human brain tau in isoforms 4R0N, 3R1N and 4R1N via TetOff and ecdysone inducible expression mechanisms. Such transgenic overexpression of tau in multiple isoforms facilitated the assembly of filamentous tau aggregates that exhibit immunoreactivities, physicochemical properties, and ultrastructural attributes reminiscent of those found in human tauopathies. The conditional tau transfectants thus provide us with a useful tool to elucidate the molecular and cellular events leading to neurofibrillary degeneration and a convenient means to test hypothetical mechanisms implicated in the etiopathogenesis of AD and related tauopathies. PMID- 15665402 TI - Correlation between astrocyte apoptosis and Alzheimer changes in gray matter lesions in Alzheimer's disease. AB - The relationships between astrocytic apoptosis and both senile plaques and neurofibrillary tangles (NFT) in gray matter lesions were examined quantitatively in Alzheimer's disease (AD) brains. Seven cortical regions were examined in seven AD brains by terminal dUTP nick end-labeling and immunolabeling with antibodies to glial fibrillary acidic protein, phosphorylated tau protein (AT180), apoptosis related proteins (caspase-3, bcl-2, and CD95), and beta amyloid protein. Senile plaques showed the lowest density in the cornu ammonis. The density of apoptotic astrocytes was significantly correlated with the density of uncored and cored senile plaques. Neuronal caspase-3 and CD95 expression levels were too low to allow statistical assessment, but Bcl-2 was expressed strongly in the astrocytes and neurons with and without NFT. The correlation of the density of apoptotic astrocytes with apoptotic neurons and NFT was not statistically significant. The density of Bcl2-positive neurons correlated significantly with those of NFT and cored senile plaques, but Bcl2-positive astrocyte density showed no correlation with density of senile plaques or apoptotic astrocytes. These observations suggest that senile plaques may be a cause of astrocytic apoptosis in the gray matter, and that Bcl-2 protein is associated with NFT formation. PMID- 15665403 TI - Amyloid-beta induced cell death is independent of free radicals. AB - Acutely dissociated rat cerebellar granule cell neurons were incubated with amyloid-beta (1-42) and studied by flow cytometry. Amyloid-beta caused a dose dependent loss of viability, as determined by intracellular accumulation of propidium iodide (PI),and that was not accompanied by significant elevation of intracellular calcium, measured by Fluo-3 or reactive oxygen species (ROS), measured by 2',7'-dihydro-dichlorfluorescein diacetate (DCF). Carnosine, a ROS scavenger and an inhibitor of non-enzymatic glycosylation, partially reduced cell death caused by amyloid-beta. We conclude that amyloid-beta causes a relatively acute loss of cell viability in cerebellar granule cell neurons, which does not result from either elevation of intracellular calcium concentration or generation of ROS. PMID- 15665404 TI - Borrelia burgdorferi persists in the brain in chronic lyme neuroborreliosis and may be associated with Alzheimer disease. AB - The cause, or causes, of the vast majority of Alzheimer's disease cases are unknown. A number of contributing factors have been postulated, including infection. It has long been known that the spirochete Treponema pallidum, which is the infective agent for syphilis, can in its late stages cause dementia, chronic inflammation, cortical atrophy and amyloid deposition. Spirochetes of unidentified types and strains have previously been observed in the blood, CSF and brain of 14 AD patients tested and absent in 13 controls. In three of these AD cases spirochetes were grown in a medium selective for Borrelia burgdorferi. In the present study, the phylogenetic analysis of these spirochetes was made. Positive identification of the agent as Borrelia burgdorferi sensu stricto was based on genetic and molecular analyses. Borrelia antigens and genes were co localized with beta-amyloid deposits in these AD cases. The data indicate that Borrelia burgdorferi may persist in the brain and be associated with amyloid plaques in AD. They suggest that these spirochetes, perhaps in an analogous fashion to Treponema pallidum, may contribute to dementia, cortical atrophy and amyloid deposition. Further in vitro and in vivo studies may bring more insight into the potential role of spirochetes in AD. PMID- 15665405 TI - Alzheimer's disease, oxidative injury, and cytokines. AB - Alzheimer's disease is infrequently a genetically driven disease. Rather it is the product of free radical injury inflicted over decades after an initial insult to the central nervous system (CNS). The brain is uniquely sensitive to oxidative injury. A variety of insults to the CNS are now associated with Alzheimer's disease. These include hypertension, diabetes, and head trauma. These then cause a cytokine cascade and microlocalized inflammation in the CNS, that in time results in clinical Alzheimer's disease. By the ninth decade of life over half of the population manifests Alzheimer's disease. Prevention or reversal of this pathophysiology will lie in administration of effective antioxidant therapy with specific treatments when etiologies are known. PMID- 15665406 TI - Induction of hyperphosphorylated tau in primary rat cortical neuron cultures mediated by oxidative stress and glycogen synthase kinase-3. AB - Neurofibrillary tangles (NFT) containing paired helical filaments (PHF) composed of abnormally phosphorylated tau are one of the hallmark lesions of the Alzheimer's disease (AD) brain. Although phosphorylation of tau is thought to precede the formation of PHF, the kinases/phosphatases involved remain poorly understood. Here we report that treatment of primary rat cortical neuron cultures with cuprizone, a copper chelator, in combination with oxidative stress (Fe(2+)/H(2)O(2)), significantly increased aberrant tau phosphorylation identified by TG3 immunochemistry. To determine the potential contribution of glycogen synthase kinase-3 (GSK-3) to the phosphorylation of tau in this model, activity of GSK-3 was determined. Cultures treated with cuprizone/Fe(2+)/H(2)O(2) showed significantly increased GSK-3 activity compared with control cultures or cultures treated with cuprizone, or Fe(2+)/H(2)O(2) alone. Concomitant treatment of cultures with lithium, a GSK-3 inhibitor, significantly decreased GSK-3 activity and reduced TG3 staining. Together these data suggest a culture model of hyperphosphorylated tau that implicates increased GSK-3 activity. PMID- 15665407 TI - New directions in neuroprotection: basic mechanisms, molecular targets and treatment strategies. PMID- 15665409 TI - Neurogenesis as a potential therapeutic strategy for neurodegenerative disorders. PMID- 15665408 TI - Neurotrophin-based strategies for neuroprotection. AB - Neurotrophins activate a number of signaling pathways relevant to neuroprotection; however, their poor pharmacological properties and their pleiotropic effects resulting from interaction with the p75(NTR)-Trk-sortilin three-receptor signaling system limit therapeutic application. While local application of neurotrophin proteins addresses some of the pharmacological challenges, selective targeting of neurotrophin receptors might allow for more selective application of neurotrophin receptor signaling modulation. Recent studies have supported the feasibility of developing non-peptidyl small molecules that mimic specific domains of neurotrophins and modulate signaling of specific neurotrophin receptors. The expression of p75(NTR) by populations of neurons most vulnerable in Alzheimer's disease and the linkage of p75(NTR) signaling to aberrant signaling mechanisms occurring in this disorder, point to potential applications for p75(NTR)-based small molecule strategies. Small molecules targeted to p75(NTR) in the settings of neurodegenerative disease and other forms of neural injury might serve to inhibit death signaling, block proNGF-mediated degenerative signaling and minimize deleterious effects promoted by pharmacologically upregulated Trk signaling. PMID- 15665411 TI - Toward a mechanistic taxonomy of cell death programs. PMID- 15665410 TI - Requirements of a brain selective estrogen: advances and remaining challenges for developing a NeuroSERM. AB - Our goal is to develop therapeutic agents that prevent age-associated neurodegenerative disease such as Alzheimer's. To achieve this goal, we are building on extensive knowledge regarding mechanisms of estrogen action in brain and the epidemiological human data indicating that estrogen/hormone therapy reduces the risk of developing Alzheimer's disease when administered at the time of the menopause and continued over several to many years. The mechanisms of estrogen action in neurons provides a systematic mechanistic rationale for determining why estrogen therapy is efficacious for prevention of Alzheimer's disease and why it is not efficacious for long-term treatment of the disease. Our preclinical research plan is a hybrid of both discovery and translational research to develop a brain selective estrogen receptor modulator (SERM). We have termed such molecules NeuroSERMs to denote their preferential selectivity for activating estrogen mechanisms in brain. Our strategy to develop NeuroSERMs is threefold: (1) determine the target of estrogen action in brain, specifically the estrogen receptor in hippocampal and cortical neurons required for the neurotrophic and neuroprotective actions of estrogen; (2) develop NeuroSERM candidate molecules using three in silico discovery and design strategies and (3) determine the neurotrophic and neuroprotective efficacy of candidate molecules using neuronal responses predictive of clinical efficacy. Using an academic translational research model, a team of scientists with expertise in molecular biology, computational chemistry, synthetic chemistry, proteomics, neurobiology and mitochondrial function have been assembled along with state of the art technologies required to develop candidate NeuroSERM molecules. PMID- 15665412 TI - The femtomolar-acting NAP interacts with microtubules: Novel aspects of astrocyte protection. AB - Activity-dependent neuroprotective protein (ADNP), a gene product essential for brain formation, contains a short octapeptide sequence NAPVSIPQ (NAP) that protects neurons against a wide variety of insults. At the pico-molar concentration range, NAP provides neuroprotection by direct interaction with neurons. At the femtomolar concentration range, NAP requires the presence of glial cells to provide neuroprotection. To further understand the mechanism of neuroprotection afforded by NAP, specific binding proteins were searched for. Tubulin, the major subunit protein of microtubules, was identified as a NAP binding molecule. NAP structure allows membrane penetration, followed by tubulin binding and facilitation of microtubule assembly toward cellular protection in astrocytes. NAP (10(-15) M) promoted microtubule assembly in vitro and protected astrocytes against zinc intoxication which is associated with microtubule disruption. A two hour incubation period of astrocytes with femtomolar concentrations of NAP resulted in microtubule re-organization and transient increases in immunoreactive non-phosphorylated tau. Microtubules are the key component of the neuronal and glial cytoskeleton that regulates cell division, differentiation and protection, while tau pathology is a major contributor to Alzheimer's disease and other dementias. The findings described here may open up new horizons in research and development of neuroprotective compounds. PMID- 15665413 TI - Redefining neuroprotective gene therapeutic strategies: Lessons learned from caloric restriction and NAD(+) metabolism. AB - Herein a case is made for the development of novel cytoprotective approaches based upon molecular mechanisms thought to underlie the caloric restriction phenomenon. This analysis leads to the prediction that molecular genetic perturbations affecting the metabolism of nuclear NAD(+) and metabolites will be neuroprotective. PMID- 15665414 TI - Iron dysregulation and Parkinson's disease. AB - We have recently demonstrated that chelation of in vivo brain iron in a form which is not available to participate in oxidative events protects against a toxin-induced form of Parkinsonism in rodents, the well-established MPTP model [32]. These data strongly suggest that iron elevations observed in the Parkinsonian substantia nigra (SN), the brain region which undergoes selective neurodegeneration in the disease, are actively involved in subsequent neurodegenerative events. However the mechanism(s) by which iron levels become elevated in the Parkinsonian SN are still unclear. We hypothesize that increased oxidative stress associated with the disease may result in dysregulation of iron homeostasis in midbrain dopaminergic neurons via alterations in binding of iron regulatory proteins (IRPs). This would mechanistically explain the noted increase in cellular iron levels in the Parkinsonian SN which appear to contribute to subsequent neurodegeneration. PMID- 15665416 TI - Paradigm shift in NMDA receptor antagonist drug development: molecular mechanism of uncompetitive inhibition by memantine in the treatment of Alzheimer's disease and other neurologic disorders. AB - Alzheimer's disease (AD) is the most common form of dementia, as least in western countries. It has been estimated that the cost to society for caring for AD patients will consume the entire gross national product of the U.S.A. by the middle of this century if left unabated. Until recently, the only available drugs for this condition were cholinergic treatments, which symptomatically enhance cognitive state to some degree, but they were not neuroprotective. In fact, many potential neuroprotective drugs tested in clinical trials failed because they were poorly tolerated. However, after our discovery of its clinically-tolerated mechanism of action, one neuroprotective drug, memantine, was recently approved by the European Union and the U.S. Food and Drug Administration (FDA) for the treatment of Alzheimer's disease. Recent phase 3 clinical trials have shown that memantine is effective in the treatment of moderate-to-severe Alzheimer's disease and possibly vascular dementia (multi-infarct dementia). Here we review the molecular mechanism of memantine's action and also the basis for the drug's use in these neurological diseases, which are mediated at least in part by excitotoxicity. Excitotoxicity is defined as excessive exposure to the neurotransmitter glutamate or overstimulation of its membrane receptors, leading to neuronal injury or death. Excitotoxic neuronal cell death is mediated in part by overactivation of N-methyl-d-aspartate (NMDA)-type glutamate receptors, which results in excessive Ca(2+) influx through the receptor's associated ion channel. Physiological NMDA receptor activity, however, is also essential for normal neuronal function. This means that potential neuroprotective agents that block virtually all NMDA receptor activity will very likely have unacceptable clinical side effects. For this reason many previous NMDA receptor antagonists have disappointingly failed advanced clinical trials for a number of neurodegenerative disorders. In contrast, studies in our laboratory have shown that the adamantane derivative, memantine, preferentially blocks excessive NMDA receptor activity without disrupting normal activity. Memantine does this through its action as an uncompetitive, low-affinity, open-channel blocker; it enters the receptor associated ion channel preferentially when it is excessively open, and, most importantly, its off-rate is relatively fast so that it does not substantially accumulate in the channel to interfere with normal synaptic transmission. Clinical use has corroborated the prediction that memantine is thus well tolerated. Besides Alzheimer's disease, memantine is currently in trials for additional neurological disorders, including other forms of dementia, depression, glaucoma, and severe neuropathic pain. A series of second-generation memantine derivatives are currently in development and may prove to have even greater neuroprotective properties than memantine. These second-generation drugs take advantage of the fact that the NMDA receptor has other modulatory sites in addition to its ion channel that potentially could also be used for safe but effective clinical intervention. PMID- 15665415 TI - Quantifying age-related myelin breakdown with MRI: novel therapeutic targets for preventing cognitive decline and Alzheimer's disease. AB - Myelin plays an essential role in brain structure and function and the human brain is uniquely dependent on the elaboration of this late invention of evolution. Our brain has the most extensive and protracted process of myelination that extends to approximately age 50 in cortical regions that have the highest risk for developing Alzheimer's disease (AD) pathology. This myelin-centered model of the human brain asserts that unique vulnerabilities of myelin, especially late-developed myelin, and the oligodendrocytes that produce it are directly pertinent to many uniquely human neuropsychiatric diseases including late-life neurodegenerative disorders such as AD. Magnetic resonance imaging (MRI) technology permits the in vivo assessment of the roughly quadratic (inverted U) lifelong trajectory of human myelin development and its subsequent breakdown. There is close agreement between neuropsychology, neuropathology, and imaging measures suggesting that the process of myelin breakdown begins in adulthood, accelerates as aging progresses, and underlies both age-related cognitive declines and the most powerful risk factor of dementia-causing disorders such as AD: age. This myelin-centered model together with the technology that makes it possible to measure the trajectory of myelin breakdown provide a framework for developing novel treatments, as well as assessing efficacy of currently available treatments, intended to slow or reverse the breakdown process in both clinically healthy as well as symptomatic populations. Such treatments can be expected to have a wide spectrum of efficacy and impact multiple human disease processes including potentially slowing brain aging and thus provide opportunities for primary prevention of age-related degenerative disorders such as AD. PMID- 15665417 TI - Neurotrophin survival signaling mechanisms. PMID- 15665418 TI - Novel nitrates as NO mimetics directed at Alzheimer's disease. AB - GT 1061 is a novel therapeutic agent that is in Phase 1 clinical studies for Alzheimer's disease. GT 1061 is one of a family of novel nitrates that have demonstrated neuroprotective properties and cognition- and memory-enhancing properties in animal models. The prototype of this family, GT 715, has been reported effectively to dissociate the neuromodulatory and the systemic hypotensive effects of nitrates, the latter seriously limiting the therapeutic use of classical nitrates. Further data on the novel nitrates, GT 715 and GT 061, are presented in (a) the malonate-lesion rat model of excitotoxic neurodegeneration, and (b) the reversal of a scopolamine-induced cognition deficit in the Morris water task which tests spatial memory. These data exemplify and reinforce the combined neuroprotective and cognition enhancing properties observed in this family of NO mimetic therapeutic agents. NO mimetics, that mimic the biological activity of NO, will bypass cholinergic receptor activation and are anticipated to provide multiple pathways of treating and circumventing dementia. NO mimetic activation of soluble guanylyl cyclase and cGMP formation in the brain represents one element of an effective neuroprotective strategy. Substantial evidence suggests that NO mimetics may display cGMP-dependent and cGMP-independent activity and may operate via multiple biochemical signaling pathways, both to ensure the survival of neurons subjected to stress and also to provide cognition-enabling pathways to circumvent dementia, providing a combined neuroprotective and cognition-enabling approach to anti-neurodegenerative therapy. PMID- 15665419 TI - Novel analogs of choline as potential neuroprotective agents. AB - The ability of choline to serve as a full, though low potency agonist for the alpha7 nicotinic receptor has provided the impetus to develop analogs that exhibit levels of potency and effectiveness suitable for use as therapeutic agents. Seven analogs of choline were synthesized based on previous work with the analog pyrrolidinecholine. The drugs were administered to differentiated PC-12 cells 24 hr prior to growth factor withdrawal which itself induced cytotoxicity in 30-40% of the cells. Three of 7 choline analogs exhibited potency and efficacy similar to that for nicotine as cytoprotective agents. Despite being tertiary amines, 4 of the choline analogs were more potent than choline in inhibiting [(3)H]choline uptake into cultured fibroblasts transfected with the high affinity, sodium-dependent choline transporter. One of the most effective analogs JAY 2-21-29 was shown to produce a potent (EC50 approximately 30 nM) cytoprotective action that was blocked by pretreatment with the alpha7 nicotinic receptor selective antagonist methyllycaconitine, but not by thealpha2 subtype preferring antagonist dihydro-beta-erythroidin. These preliminary studies support the further neurochemical characterization of these compounds 1) as selectivealpha7 nicotinic receptor agonists and, 2) based on their interaction with the choline transporter, as potential cholinergic false neurotransmitters as has been demonstrated for pyrrolidinecholine. PMID- 15665421 TI - Prolonged recirculation is required to detect secondary metabolic and hemodynamic deterioration after superior mesenteric artery occlusion. AB - We evaluated late (4 hrs) effects of reperfusion on hemodynamics after 30 or 60 min occlusion of the superior mesenteric artery (SMA) in a rat model. Spontaneously breathing animals (n=30) underwent occlusion of the SMA for 0 (sham), 30 (SMAO_30) or 60 min (SMAO_60) followed by reperfusion with normal saline. Abdominal blood flow (ABF), SMA blood flow (SBF), arterial blood pressure and heart rate were recorded continuously. Systemic vascular resistance (SVR) and SMA vascular resistance (MVR) were calculated at baseline and after 240 min reperfusion (240R). All animals survived in SMAO_30 and sham, two died in SMAO_60 after 120R. ABF remained constant in all groups. SVR increased in SMAO_30 and sham and decreased in SMAO_60 at 240R. SBF was significantly lower after reperfusion in ischemia groups as compared to sham. After 120R, SBF had increased significantly in SMAO_60 versus SMAO_30. MVR increased significantly in SMAO_30 but not in SMAO_60 and sham at 240R. 60 minutes SMA occlusion revealed early hemodynamic changes of septic circulation with increased blood flow in the SMA, decreased SVR, and pseudo-normalization of MVR. Prolonged observation periods are required to detect these significant changes which are overlooked when only studying 120 minutes of reperfusion as usually done. PMID- 15665420 TI - Novel small peptides with neuroprotective and nootropic properties. AB - The tripeptide thyrotropin-releasing hormone (TRH) and/or related analogues have shown neuroprotective activity across multiple animal trauma models as well as in a small clinical trial of spinal cord injury. The metabolic product of TRH (cyclo his-pro) retains physiological activity. We have developed a number of novel cyclic dipeptides that are structurally similar to cyclo-his-pro, and have examined their neuroprotective activity across multiple in vitro models of neuronal injury and after traumatic brain injury (TBI) in rodents. Four such compounds were found to reduce cell death after trophic withdrawal or traumatic injury in primary neuronal cultures; two also protected against glutamate or beta amyloid neurotoxicity. All compounds significantly improved motor and cognitive recovery after controlled cortical impact injury in mice, and markedly reduced lesion volumes as shown by high field magnetic resonance imaging. Further, compound 35b, which is being developed for clinical trials, also showed considerable neuroprotection after fluid percussion induced TBI in rats, and improved cognitive function after daily administration in chronically brain injured rats. At a mechanistic level, the drugs attenuate both apoptotic and necrotic cell death in primary neuronal cultures, markedly reduce intracellular calcium accumulation after injury, and limit changes in mitochondrial membrane potential and associated cytochrome c release. In addition, microarray studies show that 35b reduces transcriptional changes after injury for a number of genes (and proteins) that may be associated with secondary injury, including cell cycle genes, aquaporins and cathepsins. It also upregulates brain-derived neurotrophic factor (BDNF), heat shock proteins (HSP) and hypoxia inducible factor (HIF). Thus, these novel dipeptides have multipotential actions that make them candidates for the treatment of both acute and chronic neurodegeneration. PMID- 15665422 TI - Fibrinolytic activity in Nigerian diabetics with renal insufficiency. AB - Diabetics have reduced fibrinolytic activity while chronic renal failure patients are prone to both haemorrhagic diathesis and reduced fibrinolysis. In the quest to find out what obtains when diabetics develop renal impairment, fibrinolytic activity was studied in 51 known diabetics consisting of 9 diabetics with normal renal function, 13 with mild renal impairment and 29 in chronic renal failure. No patient had end-stage renal disease and none was on dialysis therapy. Plasma fibrinogen concentration (PFC) was measured using clot weight method and euglobulin lysis time (ELT) as a measure of fibrinolytic activity was also measured. Our results show that PFC and ELT did not differ significantly in the different groups of diabetics. There was also no correlation between the serum creatinine of all diabetics and PFC on one hand and ELT on the other hand. We conclude that Nigerian diabetics probably still benefit from enhanced fibrinolysis before they reach end-stage renal disease. PMID- 15665423 TI - Effects of Compound Dan-shen Root Dropping Pill on hemorheology in high-fat diet induced hyperlipidemia in dogs. AB - The purpose of this study was to examine the effects of Compound Dan-shen Root Dropping Pill (CDRDP) (Tasly Group, Tianjing, China) on hemorheology and biorheology of dogs suffering from hyperlipidemia induced by high-fat diet. Eighteen dogs were randomly divided into two groups: the high-fat diet group (H group); the control group (C group), fed with a standard laboratory diet. Six month later, six dogs in the H group were chosen as the drug-taking group (D group), to which CDRDP was administered, fed with the same diet as H group. In the 4th month, blood was taken from the veins of the dogs, and blood triglyceride (TG), total cholesterol (TC), RBC hemorheological indexes as well as malondialdehyde (MDA), glutathione transferase (GSH-ST) and superoxide dismutase (SOD) activities in plasma and erythrocytes were measured. Compared with H group, TC, TG, plasma MDA levels, the whole blood viscosity, RBC osmotic fragility and the value of CHOL (cholesterol)/PL (phospholipid) of the membrane of D group decreased, however, erythrocyte GSH-ST, histopathological changes in liver, deformation index (DI), orientation index (DI)or, small deformation index (DI)d, electrophoresis ratio and microfluidity of the membrane lipid bilayer of RBCs, increased distinctly. CDRDP can improve micro-hemorheological characteristics, therefore has a significant therapy application of hyperlipidemia. PMID- 15665424 TI - Whole blood, plasma viscosity, and erythrocyte aggregation as a determining factor of competitiveness in standard bred trotters. AB - In situations where capillary perfusion in skeletal muscles is limited, changes in blood flow play an important role. Especially alterations in intrinsic erythrocyte factors like red cell aggregation and deformability would increase blood flow resistance. In our study we investigated whether whole blood and plasma viscosity influence exercise tolerance during submaximal training and whether a difference can be realized between trained and untrained standardbred trotters. Venous blood from 42 healthy adult trotters (20 horses at the beginning of their training and 22 well trained horses) was investigated before, immediately afterwards and 30 minutes after submaximal exercise. In both groups whole blood viscosity (WBV; LS30, Contraves, Switzerland) increased significantly (p<0.001) at all shear rates (94 s-1, 2.4 s-1, 0.7 s-1) and decreased after 30 minutes to baseline, as did plasma viscosity (PV; OCR-D, Paar, Austria; p<0.001). No changes in WBV could be seen in hematocrit (40%) standardized samples. Erythrocyte aggregation (EA) indices increased (Myrenne, Germany; p<0.001). Creatin-kinase (p<0.001), lactate (p<0.001), hemoglobine (p<0.001), heart rate and oxygen saturation (p<0.05) increased, while PH (p<0.05), and BE (p<0.001) decreased during the race. In our study, submaximal exercise was related to remarkable changes in hemorheologic variables in the single animal. It was also shown that exercise resulted in a more extensive change of fluidity in well trained horses in comparison to untrained animals, at least for low shear WBV and EA (p<0.05). PMID- 15665425 TI - Polymorphonuclear leukocyte integrins in chronic renal failure. AB - Patients with chronic renal failure (CRF), in comparison with general population, show a higher cardiovascular mortality, not fully explained by the "traditional" risk factors. Among the new factors that have been hypothesized, leukocytes might play an important role. In a group of patients with mild CRF we determined, at baseline and after in vitro activation with 4-phorbol-12-myristate-13-acetate (PMA) and N-formyl-methionyl-leucyl-phenylalanine (fMLP), the polymorphonuclear leukocytes (PMN) beta2-integrin pattern (CD11a, CD11b, CD11c and CD18) by using indirect immunofluorescence with a flow cytometer. At baseline we observed an increase in the phenotypical expression of CD11b, CD11c and CD18 in CRF patients. In normal subjects, after activation with both agents, we noted an increase of all adhesion molecules, while in CRF patients we found an increase in the expression of CD11b, CD11c and CD18 but not of CD11a. The altered behaviour of the PMN integrin pattern in mild CRF patients, likely reflecting a state of PMN activation, might have a pathophysiological significance, considering the high incidence of cardiovascular events in CRF. PMID- 15665426 TI - Comparison of cation-osmotic haemolysis in normal and low birth weight newborns during the first month of life. AB - The present study was designed to compare cation-osmotic haemolysis (COH) in normal versus low birth weight newborns during the first month of postnatal life. COH was assessed in 80 normal (NBW) and 45 low birth weight newborns (LBW). A significant decrease in COH in the solutions with low ionic strength (15.4-30.8 mmol.l-1 NaCl) was found in LBW in the early period after birth when compared with NBW (p<0.01; p<0.001). In the solutions with high ionic strength (123.2 154.0 mmol.l-1 NaCl), COH again was significantly lower in LBW than in NBW neonates (p<0.01; p<0.001). The differences in COH were still present after both two weeks and one month of postnatal life but only in the solutions with the highest ionic strength (138.6-154.0 mmol.l-1 NaCl) (p<0.01; p<0.001). To conclude, our study demonstrates that COH is significantly lower in LBW than in NBW neonates. Furthermore, the latter show still lower values than adults. Finally, the relationship between COH and erythrocyte deformability is discussed. PMID- 15665427 TI - Tubular dysfunction and hemodynamic alteration in normoalbuminuric type 2 diabetes. AB - Altered renal function has been encountered in normoalbuminuric patient with type 2 diabetes. A search for alternative index that is more sensitive than microalbuminuria for early detection of diabetic nephropathy has been performed. In the present paper, compartmental functions of nephron namely creatinine clearance (CCr) reflecting glomerular function, fractional excretion of magnesium (FE Mg) reflecting tubular function and intrarenal hemodynamics reflecting vascular function were assessed in 40 type 2 diabetic patients with normoalbuminuria and in 10 type 2 diabetic patients with albuminuria. In normoalbuminuric patients, glomerular function showed a low, normal or high CCr due to hyperfiltration. Tubular function revealed a significantly elevated FE Mg. Vascular function was altered with preferential constriction of the efferent arteriole, intraglomerular hypertension and profound reduction in peritubular capillary flow. A greater degree of functional defect was observed in albuminuric patients. Defective functions of the nephron implies that renal tissue injury has already been present in normoalbuminuric state. FE Mg appears to be a sensitive marker for early detection of diabetic nephropathy. Significant reduction in peritubular capillary flow correlates inversely with the altered FE Mg. Such correlation favors the chronic ischemic concept of tubulointerstitial injury and therefore supports the tubular functional defect observed in type 2 diabetes. PMID- 15665428 TI - Plasma markers of platelet and polymorphonuclear leukocyte activation in young adults with acute myocardial infarction. AB - Our goal was to evaluate some plasma markers of platelet and polymorphonuclear leukocyte (PMN) activation in a group of young adults with acute myocardial infarction (AMI) at the initial stage and after three months. We enrolled 49 AMI subjects aged<45 years and examined plasmatic levels of platelet factor 4 (PF4), beta-thromboglobulin (beta-TG), elastase and myeloperoxidase (MPO) using ELISA methods. PF4 and beta-TG were increased, compared to control subjects, both at the initial stage and after 3 months. In control subjects and in AMI patients, at both times of observation, there was a significant and positive correlation between the two platelet parameters, while no correlation was present between each parameter and platelet count. In AMI patients there was an increase in elastase levels in comparison with the control group; this increase was evident at the initial stage and after 3 months. There was no difference in MPO levels between control subjects and AMI patients. In control subjects and in AMI patients there was a significant and positive correlation between elastase and MPO level, whereas no relationship was found between each marker and PMN count. Our data show that in young AMI patients the discharge treatment including antiplatelet drugs did not modify platelet activation and suggest the association of molecules able to inhibit PMN activation to the conventional therapy of these AMI patients. PMID- 15665429 TI - Down with dogma. PMID- 15665430 TI - Clinical management of the Herbst Occlusal Hinge appliance. PMID- 15665431 TI - A new indirect bonding technique. PMID- 15665432 TI - Recycling debonded brackets with an acid bath. PMID- 15665433 TI - Functional treatment of mild skeletal asymmetry. PMID- 15665434 TI - Movement of ankylosed permanent teeth with a distraction device. PMID- 15665435 TI - Stabilizing a pontic with wax. PMID- 15665436 TI - Intrusion of overerupted upper second molars with a modified lingual arch. PMID- 15665437 TI - Managing orthodontic pain. PMID- 15665438 TI - Lower molar distalization with the unilateral Frozat appliance. PMID- 15665439 TI - A new indirect bonding technique for lingual retainers. PMID- 15665440 TI - Late-forming supernumerary teeth. PMID- 15665441 TI - JCO Roundtable: women in orthodontics. Interview. PMID- 15665442 TI - Passive and active overlay systems. PMID- 15665443 TI - A new nickel titanium rapid maxillary expansion screw. PMID- 15665444 TI - Physiology and pathophysiology of proteinase-activated receptors (PARs): proteinases as hormone-like signal messengers: PARs and more. AB - Proteinases like thrombin and trypsin, long known for their ability to activate the coagulation cascade or to act as digestive enzymes for many protein targets, are now recognized as hormone-like regulators of cell function. These serine proteinases activate cell signaling by triggering a novel family of G-protein coupled receptors, termed proteinase-activated receptors (PARs). This article summarizes the unique mechanisms involved in PAR activation and outlines the many different settings in which the PARs act to regulate tissue function. The PARs can be seen to play a role in inflammatory processes in large part via a neurogenic mechanism. Apart from activating PARs to cause their physiological effects in tissues, proteinases can also mediate cell signaling via a number of other mechanisms, including the activation of growth factor receptors, like the one for insulin. Thus, this article also points out the non-PAR mechanisms whereby proteinases can have hormone-like actions in cells and tissues. PMID- 15665445 TI - Potentiation of T cell function by a marine algae-derived sulfated polymannuroguluronate: in vitro analysis of novel mechanisms. AB - Marine algae-derived sulfated polymannuroguluronate (SPMG), a candidate drug for AIDS treatment, was intraperitoneally injected into normal mice for 6 weeks, and the in vivo and in vitro mechanisms of SPMG for immunomodulation were investigated in isolated lymphocytes by MTT assay, flow cytometry, and surface plasmon resonance assay. SPMG treatment at 5 and 10 mg/kg enhanced concanavalin A (ConA)-induced T cell proliferation, cellular levels of CD69, interleukin-2 (IL 2), and interferon-gamma (IFN-gamma), as well as CD4/CD8 ratio, while decreasing tumor necrosis factor-alpha (TNF-alpha) level in T cells of peripheral blood mononuclear cells. In addition, 1 molecule of SPMG bound to 2/3 molecules of IL-2 with a K(D) of 9.53 x 10(-7) M. Heparin prevented SPMG binding to IL-2 by 72.2%; thus, to a large extent, SPMG and heparin share common binding sites on IL-2. In contrast, other glycosaminoglycans (e.g., chondroitin sulfate and dermatan sulfate) had little effect on SPMG and IL-2 interaction, suggesting the requirement of a defined sequence within the sugar chain for specific recognition of IL-2. Concomitant treatment of IL-2 and SPMG augmented lymphocyte proliferation, compared with IL-2 alone; in contrast, SPMG alone had no proliferative effect. Taken together, our findings demonstrated for the first time that SPMG exerted its immunomodulation by direct activation of T cell function, accompanied by simultaneous modulation of cytokine function, which suggests that SPMG would show great promise for use in anti-AIDS therapy. PMID- 15665446 TI - Ability of fourteen chemical agents used in dental practice to induce chromosome aberrations in Syrian hamster embryo cells. AB - To assess the genotoxicity of 14 chemical agents used in dental practice, the ability of these agents to induce chromosome aberrations was examined using Syrian hamster embryo (SHE) cells. Statistically significant increases in the frequencies of chromosome aberrations were induced in SHE cells treated with 7 of 10 chemical agents used as endodontic medicaments, that is, carbol camphor, m cresol, eugenol, guaiacol, zinc oxide, hydrogen peroxide, and formaldehyde. The other 3 chemical agents, that is, thymol, glutaraldehyde, and iodoform, did not increase the levels of chromosome aberrations. Of the 4 chemical agents that are used as an antiseptic on the oral mucosa, chromosome aberrations were induced by iodine, but not by the other 3 antiseptics, benzalkonium chloride, benzethonium chloride, and chlorhexidine. Among the 6 chemical agents exhibiting a negative response in the assay, only thymol induced chromosome aberrations in the presence of exogenous metabolic activation. Our results indicate that chemical agents having a positive response in the present study are potentially genotoxic to mammalian cells and need to be studied further in detail. PMID- 15665447 TI - Yersinia enterocolitica serovar O:8 infection in breeding monkeys in Japan. AB - In the period from December 2002 to January 2003, 5 of 50 squirrel monkeys (Saimiri sciureus) housed at a Zoological Garden in the Kanto region of Japan died following a few days' history of diarrhea. After this outbreak had ended in the squirrel monkeys, 1 of 2 dark-handed gibbons (Hylobates agilis) died in April of 2003, showing similar clinical signs. We examined the organs of 3 of the dead squirrel monkeys and of the dark-handed gibbon, and Yersinia enterocolitica serovar O:8, which is the most pathogenic serovar of Y. enterocolitica, was isolated. In order to determine the source and the transmission route of infection, 98 fecal samples (45 from squirrel monkeys, 20 from other monkeys of 18 different species, and 33 from black rats captured around the monkey houses) and 7 water samples were collected in the Zoological Garden, and were examined for the prevalence of Y. enterocolitica serovar O:8. Serovar O:8 was isolated from 21 of 65 monkeys (32.3%) and 5 of 33 (15.2%) black rats (Rattus rattus). Furthermore, we examined the 30 isolates using molecular typing methods, pulsed field gel electrophoresis (PFGE), ribotyping using the RiboPrinter system, and restriction endonuclease analysis of virulence plasmid DNA (REAP), and compared the isolates in this outbreak with Japanese O:8 isolates previously identified. Genotyping showed that almost all the isolates were identical, and the genotype of the isolates was highly similar to that from wild rodents captured in Niigata Prefecture. This is the first report of fatal cases of Y. enterocolitica serovar O:8 infection in monkeys anywhere in the world. PMID- 15665448 TI - Detection of Tannerella forsythia and/or Prevotella intermedia might be useful for microbial predictive markers for the outcome of initial periodontal treatment in Koreans. AB - A proportion of diseased sites in periodontal disease do not respond to the initial treatment, which might be due in part to the presence of specific microbial pathogens. The aim of this study was to clarify the value of microbial screening for predicting the outcome of periodontal treatment in Koreans using a polymerase chain reaction (PCR). This study enrolled 32 adults with periodontal disease. Microbial and clinical examinations were performed at the baseline and after the initial treatment (professional toothbrushing, scaling, and root planing). Subgingival plaque samples were taken from four sites in each subject (total 128 samples). PCR was used to detect the four putative pathogenic bacteria. There was an improvement in the average of each clinical measurement after the initial treatment. However, approximately half of the sites exhibiting bleeding upon probing (BOP) at the baseline still exhibited bleeding after treatment. There was a close association between the presence of BOP and the presence of Tannerella forsythia (formerly Bacteroides forsythus) and/or Prevotella intermedia. Furthermore, the sites harboring both T. forsythia and P. intermedia at the baseline had a poorer response to treatment than the sites where these two species were not detected. Therefore, microbial screening for T. forsythia and P. intermedia might be useful for predicting the treatment outcome in Koreans. PMID- 15665449 TI - Seroepidemiology of spotted fever group and typhus group rickettsioses in humans, South Korea. AB - The prevalence of spotted fever group (SFG) and typhus group (TG) rickettsioses was investigated in 3,362 sera by immunofluorescence assay. The serum samples were obtained from patients with acute febrile episodes in South Korea from December 1992 to November 1993. The number of polyvalent positive sera against SFG rickettsial agents at the level of 1: 40 dilution was 269 (8%) in Rickettsia sibirica, 482 (14.34%) in R. conorii, and 546 (16.24%) in R. akari. Many of the positive sera contained immunoglobulin (Ig) M antibodies rather than IgG antibodies. These results strongly suggest that SFG rickettsioses are prevalent in Korea. For TG rickettsial agents, the number of positive sera was 1,096 (32.60%) in R. typhi and 951 (28.29%) in R. prowazekii. Only a few epidemic typhus positive sera contained IgM antibodies. The result suggests that recent and/or primary infections of epidemic typhus were very rare in Korea during the said period. Among seven patients who had high titers (1:5,120) of IgG antibody to R. prowazekii, six were over 50 years old. The result suggests that Brill Zinsser disease was prevalent in Korea. PMID- 15665450 TI - Quantitative analysis of cereulide, an emetic toxin of Bacillus cereus, by using rat liver mitochondria. AB - An emetic toxin cereulide, produced by Bacillus cereus, causes emetic food poisonings, but a method for quantitative measurement of cereulide has not been well established. A current detection method is a bioassay method using the HEp-2 cell vacuolation test, but it was unable to measure an accurate concentration. We established a quantitative assay for cereulide based on its mitochondrial respiratory uncoupling activity. The oxygen consumption in a reaction medium containing rat liver mitochondria was rapid in the presence of cereulide. Thus uncoupling effect of cereulide on mitochondrial respiration was similar to those of uncouplers 2,4-dinitrophenol (DNP), carbonylcyanide m-chlorophenylhydrazone (CCCP), and valinomycin. This method gave constant results over a wide range of cereulide concentrations, ranging from 0.05 to 100 microg/ml. The minimum cereulide concentration to detect uncoupled oxygen consumption was 50 ng/ml and increased dose-dependently to the maximum level. Semi-log relationship between the oxygen consumption rate and the cereulide concentration enables this method to quantify cereulide. The results of this method were highly reproducible as compared with the HEp-2 cell vacuolation test and were in good agreement with those of the HEp-2 cell vacuolation test. The enterotoxin of B. cereus or Staphylococcus aureus did not show any effect on the oxygen consumption, indicating this method is specific for the identification of cereulide as a causative agent of emetic food poisonings. PMID- 15665451 TI - Anaerospora hongkongensis gen. nov. sp. nov., a novel genus and species with ribosomal DNA operon heterogeneity isolated from an intravenous drug abuser with pseudobacteremia. AB - A bacterium was isolated from the blood culture of an intravenous drug abuser with pseudobacteremia. The cells were strictly anaerobic, straight or slightly curved, sporulating, Gram-negative rods. It grew on sheep blood agar as non hemolytic, pinpoint colonies after 48 hr of incubation at 37 C in an anaerobic environment. It was motile but did not produce catalase or cytochrome oxidase. 16S ribosomal DNA (rDNA) sequencing revealed three different copies of 16S rDNA sequences. More than 90% of the differences among them were due to differences in the lengths of the sequences. Phylogenetically, the bacterium is clustered with Dendrosporobacter, Sporomusa, and Propionispora, the other three genera of anaerobic, sporulating, Gram-negative rods. There were 8.6-11.1% differences between the 16S rDNA sequences of the bacterium and that of D. quercicolus, 4.7 15.1% differences between the 16S rDNA sequences of it and those of S. acidovorans, S. aerivorans, S. malonica, S. ovata, S. paucivorans, S. silvacetica, S. spaeroides, and S. termitida, and 7.6-13.1% differences between the 16S rDNA sequences of it and those of P. hippei and P. vibrioides. The G+C content of the bacterium (mean +/- SD) was 46.8 +/- 3.2%. For these reasons, a new genus and species, Anaerospora hongkongensis gen. nov. sp. nov., is proposed, for which HKU15T is the type strain. PMID- 15665453 TI - A calcium binding protein, S100A4, mediates T cell dependent cytotoxicity as a transformation-associated antigen. AB - The nature of the target molecule of TCR gamma delta T cell-mediated lysis remains to be determined. As we previously reported, #067 monoclonal antibody (mAb) recognizes one of the transformation-associated antigens, designated as #067 antigen. This antigen is expressed on the cell surface of rat fibrosarcoma W31 cells, which are established by transformation of fetal fibroblastic WFB cells with H-ras oncogene. It has been suggested that the #067 antigen is a target molecule for TCR gamma delta T cells since #067 mAb inhibited TCR gamma delta T cell-mediated lysis against #067 positive cells. In this study we attempted to identify the protein sequence of the #067 antigen. By using molecular cloning techniques, we demonstrated that a calcium binding protein, S100A4, was possibly one and the same molecule as the #067 antigen. It was shown that the expression of S100A4 was higher in W31 cells than in WFB cells at transcription and protein level. Flow cytometry and immunocytochemical studies showed that #067 antigen partially co-localized with S100A4 on the cell surface as well as the cytoplasm of W31 cells. Moreover, rabbit anti-S100A4 polyclonal antibodies (pAb) inhibited TCR gamma delta T cell-mediated lysis against #067 positive cells. Our results indicated that S100A4 may play a role as a possible target molecule for TCR gamma delta T cell-mediated lysis although how S100A4 is involved in TCR gamma delta T cell-mediated lysis remains to be determined. PMID- 15665452 TI - Induction of inducible nitric oxide (NO) synthase mRNA and NO production in macrophages infected with influenza A/PR/8 virus and stimulated with its ether split product. AB - We investigated the inductive activity of infective influenza A/PR/8/34 (PR8) virus and its ether-split product (ESP) on the expression of inducible nitric oxide (NO) synthase (iNOS) and NO production in RAW264.7 (RAW) cells, a murine macrophage (M psi) cell line, and thioglycolate-elicited peritoneal M psi (TPM). In both cells, PR8 virus infection induced iNOS mRNA between 4 hr and 24 hr, attaining a peak value at 12 hr. In correlation with induction of iNOS mRNA, NO amounts increased significantly from 12 to 24 hr. Moreover, this study demonstrated that ESP with the same hemagglutination titer as PR8 virus could induce iNOS mRNA and NO production, although the inductive activity of ESP was weaker than that of PR8 virus. Considering the dual role (beneficial and detrimental roles) of NO on certain inflammatory disorders and virus infections, the inductive activity of influenza virus on the iNOS-mediated NO production independent of its infectivity might contribute to a modification of influenza virus infection. PMID- 15665455 TI - Sequence analysis of the hypervariable regions of the 56 kDa immunodominant protein genes of Orientia tsutsugamushi strains in Malaysia. AB - The DNA sequences encompassing two hypervariable regions, VD II and III of the 56 kDa immunodominant protein gene of 21 Malaysian strains of Orientia tsutsugamushi were determined. Two strains demonstrated a 100% DNA homology with the Gilliam prototype strain, and one with TH1817 strain and TA678 strain respectively. High percentages of DNA similarity (95-99%) were observed with Karp (4 strains), Gilliam (2 strains), TH1817 (4 strains), TC586 (3 strains) and TA763 (1 strain). The remaining strains demonstrated the highest DNA similarity with TA763 (1 strain, 89%), TA678 (1 strain, 86%) and TA686 (1 strain, 87%). Our study provides additional evidence on the existence and the genetic heterogeneity of TA strains of the Southeast Asia and their closely related strains in Malaysia. PMID- 15665454 TI - Autoantibodies to peroxiredoxin I and IV in patients with systemic autoimmune diseases. AB - Anti-oxidative enzymes protect living bodies from various oxidative stresses. In the systemic autoimmune diseases, autoantibodies to oxidized molecules and to anti-oxidative enzymes have been reported. To promote understanding of the relationships between autoimmunity and oxidative stress, we here investigate whether autoimmunity to the anti-oxidative peroxiredoxin (Prxs) enzymes exists in patients with systemic autoimmune diseases. Specifically, we detected autoantibodies to recombinant Prx I and Prx IV respectively by ELISA and western blotting. Next, clinical parameters were compared between the anti-Prx I or IV positive and-negative patients. We found that 33% of the 92 patients with autoimmune diseases tested possessed autoantibodies to Prx I (57% in systemic lupus erythematosus (SLE), 19% in rheumatoid arthritis (RA), 5% in Behcet disease, and 46% in primary vasculitis syndrome). In contrast, autoantibodies to Prx IV were detected in only 17% of the same patients. No significant correlation was found between occurrence of the two autoantibodies. Clinically, possession of anti-Prx I autoantibodies correlated with lower serum levels of CH50, C3, and C4. Taken together, our data demonstrate the existence of autoantibodies to Prxs for the first time. The autoantibodies to Prx I may be involved in the pathophysiology of systemic autoimmune diseases such as SLE and vasculitis. PMID- 15665456 TI - A hemolysin of Vibrio mimicus (VMH) stimulates cells to produce ATP and cyclic AMP which appear to be secretory mediators. AB - The hemolysin of Vibrio mimicus(VMH) is a pore-forming toxin with both enterotoxic and hemolytic activity. The hemolysis by VMH is induced by creation of pores in the membrane of erythrocyte; however, the mechanism for the enterotoxic action of VMH has remained unclear. In order to clarify the mechanism, we incubated T84 cells (a human colon carcinoma cell line) with VMH and found that the levels of ATP and cyclic AMP of culture medium increased after exposure of the cells to VMH. Subsequently, we found that the fluid accumulating activity of VMH in a mouse internal loop assay was reduced by administration of glibenclamide, an inhibitor of cyclic AMP-dependent chloride channels, into the intestinal loop. These results suggest that the stimulation of cells to produce nucleotides by VMH is linked to the enterotoxic activity of the toxin. PMID- 15665457 TI - Recombinant cholera toxin B subunit activates dendritic cells and enhances antitumor immunity. AB - Activation of dendritic cells (DC) is crucial for priming of cytotoxic T lymphocytes (CTL), which have a critical role in tumor immunity, and it is considered that adjuvants are necessary for activation of DC and for enhancement of cellular immunity. In this study, we examined an adjuvant capacity of recombinant cholera toxin B subunit (rCTB), which is non-toxic subunit of cholera toxin, on maturation of murine splenic DC. After the in vitro incubation of DC with rCTB, the expression of MHC class II and B7-2 on DC was upregulated and the secretion of IL-12 from DC was enhanced. In addition, larger DC with longer dendrites were observed. These data suggest that rCTB induced DC maturation. Subsequently, we examined the induction of tumor immunity by rCTB-treated DC by employing Meth A tumor cells in mice. Pretreatment with subcutaneous injection of rCTB-treated DC pulsed with Meth A tumor lysate inhibited the growth of the tumor cells depending on the number of DC. Moreover, intratumoral injection of rCTB treated DC pulsed with tumor lysate had therapeutic effect against established Meth A tumor. Immunization with DC activated by rCTB and the tumor lysate increased number of CTL precursor recognizing Meth A tumor. The antitumor immune response was significantly inhibited in CD8+ T cell-depleted mice, although substantial antitumor effect was observed in CD4+ T cell-depleted mice. These results indicated that rCTB acts as an adjuvant to enhance antitumor immunity through DC maturation and that CD8+ T cells play a dominant role in the tumor immunity. Being considered to be safe, rCTB may be useful as an effective adjuvant to raise immunity for a tumor in clinical application. PMID- 15665458 TI - B1-B cells are the main antigen presenting cells in CpG-ODN-stimulated peritoneal exudate cells. AB - Peritoneal exudate cells (PEC) have long been used as antigen presenting cells (APC), because they have been considered to contain mainly macrophages. However, it is still unclear specifically which cells of the peritoneal exudate function as APC. Herein, we focused on macrophages and B1-B cells of the PEC and examined their APC function and cytokine production. B1-B cells purified from PEC functioned effectively as APC after CpG-stimulation and mainly produced IL-10. In contrast, macrophages purified from PEC were not able to present incorporated antigens to T cells, despite the production of IL-12 and expression of co stimulatory molecules after CpG stimulation. These results suggest that previously held ideas regarding the functions of the mixture of cells in the PEC need to re-evaluated. In summary, the antigen presenting function of PEC was mainly attributed to B1-B cells and immunoenhancing cytokine production was dominantly derived from peritoneal macrophages. PMID- 15665459 TI - Coumarin and flavone derivatives from estragon and thyme as inhibitors of chemical mediator release from RBL-2H3 Cells. AB - Estragon and thyme extracts showed potent inhibitory activities against chemical mediator release from rat basophilic leukemia RBL-2H3 cells. 7-Methoxycoumarin was isolated from estragon, and 5,4'-dihydroxy-6,7,3'-trimethoxyflavone, 5,4' dihydroxy-6,7,8,3'-tetramethoxyflavone, 5-hydroxy-6,7,8,3',4'-pentamethoxyflavone and luteolin were isolated from thyme as active components. Structure-activity relationship studies among the active isolates and their related compounds indicated that the oxygen-containing functional group at the 7-position of the coumarin structure was advantageous for the inhibitory activity and that methylation of the hydroxyl group at the 4'-position of the flavone structure was disadvantageous. It was also found that coumarin derivatives inhibited an earlier step than intracellular calcium release and proteinkinase C activation, while flavones inhibited a later step or both earlier and later steps. PMID- 15665460 TI - Interaction between trehalose and alkaline-earth metal ions. AB - We investigated the interaction between trehalose and alkaline-earth metal ions. The nuclear relaxation times of carbon atoms of trehalose were shortened by addition of the alkaline-earth chloride salts, MgCl2, CaCl2, and SrCl2, indicating that trehalose formed metal-complexes with the alkaline-earth metal chlorides. From the data of the 1H-1H coupling constants of trehalose in the presence of the alkaline-earth chlorides, it appeared that trehalose formed complexes with MgCl2, and CaCl2 at the various complexing sites: Mg2+ was coordinated to O-4 and O-4' of trehalose, and Ca2+ to O-2 and O-3. We succeeded in the preparation of two types of crystals of the trehalose/CaCl2. One was a crystal consisting of trehalose, CaCl2, and water in a ratio of 1:1:1. The other was an anhydrous crystal containing trehalose and CaCl2 in a ratio of 1:2. Several applications of the complexing between trehalose and the metal ions for food processing are proposed. PMID- 15665461 TI - Comparative effects of non-gelatinized corn and rice starches on the life-span of ICR mice. AB - We compared the effects of non-gelatinized rice and corn starches on the life span of ICR mice. Six groups of male ICR mice consisting of 30 animals each were maintained on purified experimental diets containing either corn or rice starch and different amounts of soybean oil (6, 12 or 24%) throughout their life-time. Plots of the survival rates of the mice indicate that rice compared to corn starch conferred a longer life-span to ICR mice, although a significant difference due to the starch type was only observed in the mice fed on the 24% fat diet (p=0.012). A divergent effect of rice and corn starches on the survival rate was apparent when observations were combined with respect to the starch type regardless of the dietary fat level (p=0.005). In addition, two-way ANOVA data indicate that the mean survival time was longer for the mice given rice starch (593-645 days) than for those fed corn starch (538-580 days) (p=0.011). However, no significant difference in these parameters due to dietary fat levels was observed. The results of our study indicate that starch type is one of the determinants of longevity in mice. PMID- 15665462 TI - Immunological effects of partially hydrolyzed arabinoxylan from corn husk in mice. AB - The effect of oral administration of partially hydrolyzed water-soluble corn husk arabinoxylan (CHAX), the average molecular weight of which is about 53 kDa, on immunopotentiating activity was investigated in mice. Oral administration of CHAX to healthy mice significantly augmented the production of interleukin (IL)-2 and interferon (IFN)-gamma with a slight increase in IL-4 in mitogen-induced proliferation of spleen cells. Natural killer (NK) cell activity in spleen cells from mice, which were transplanted tumor and administrated CHAX, was augmented about 2-fold. In model mice of atopic dermatitis, the average ear thickness of mice administrated CHAX was induced by dinitrophenyl-fluorobenzene (DNFB) after injection of an anti-dinitrophenyl (DNP)-IgE monoclonal antibody was much smaller than that in control animals. These results suggest that CHAX has the ability to increase the level of immunopotentiating activity without causing over response of immunological reaction even if it is administrated orally to mice. PMID- 15665463 TI - Characterization of a metal-chelating substance in coffee. AB - A metal-chelating substance in brewed coffee was separated and characterized by its chemical structure. This substance was a brown polymer. The contents of sugars, amino acids and phenolics in the substance were evaluated. This polymer contained small amounts of sugars and amino acids in its partial structure. After being decomposed by alkaline fusion, the decomposition products were identified by HPLC and GC-MS. Several phenolics were detected in the decomposed products. To characterize this substance, various types of model compounds were prepared by roasting chlorogenic acid, sucrose, and (or) protein with cellulose powder. Among these model compounds, the polymer-forming ability was highest in the model prepared from all four of materials, but the metal-chelating ability was the highest in the model prepared from chlorogenic acid and cellulose. These results suggest that this metal-chelating substance was a melanoidin-like polymer formed by the decomposition and polymerization of sugars, amino acids and phenolics. PMID- 15665464 TI - Effects of dietary protein of Korean foxtail millet on plasma adiponectin, HDL cholesterol, and insulin levels in genetically type 2 diabetic mice. AB - We examined the effects of intake of Korean foxtail millet protein (FMP) on plasma levels of lipid, glucose, insulin, and adiponectin in genetically type 2 diabetic KK-Ay mice. When mice were fed a normal FMP diet or a high-fat-high sucrose diet containing FMP for 3 weeks, in both experiments plasma concentrations of high-density lipoprotein cholesterol (HDL-cholesterol) and adiponectin increased remarkably in comparison with a casein diet group, whereas concentrations of insulin decreased greatly and that of plasma glucose was comparable to that in the casein diet group. Considering the role of adiponectin, insulin, and HDL-cholesterol in diabetes, atherosclerosis, and obesity, it appears likely that FMP may improve insulin sensitivity and cholesterol metabolism through an increase in adiponectin concentration. Therefore, FMP would serve as another beneficial food component in obesity-related diseases such as type 2 diabetes and cardiovascular diseases. PMID- 15665465 TI - Purification and characterization of serine proteinase from a halophilic bacterium, Filobacillus sp. RF2-5. AB - In order to find a unique proteinase, proteinase-producing bacteria were screened from fish sauce in Thailand. An isolated moderately halophilic bacterium was classified and named Filobacillus sp. RF2-5. The molecular weight of the purified enzyme was estimated to be 49 kDa. The enzyme showed the highest activity at 60 degrees C and pH 10-11 under 10% NaCl, and was highly stable in the presence of about 25% NaCl. The activity was strongly inhibited by phenylmethane sulfonyl fluoride (PMSF), chymostatin, and alpha-microbial alkaline proteinase inhibitor (MAPI). Proteinase activity was activated about 2-fold and 2.5-fold by the addition of 5% and 15-25% NaCl respectively using Suc-Ala-Ala-Phe-pNA as a substrate. The N-terminal 15 amino acid sequence of the purified enzyme showed about 67% identity to that of serine proteinase from Bacillus subtilis 168 and Bacillus subtilis (natto). The proteinase was found to prefer Phe, Met, and Thr at the P1 position, and Ile at the P2 position of peptide substrates, respectively. This is the first serine proteinase with a moderately thermophilic, NaCl-stable, and NaCl-activatable, and that has a unique substrate specificity at the P2 position of substrates from moderately halophilic bacteria, Filobacillus sp. PMID- 15665466 TI - Mode of action of cellulases on dyed cotton with a reactive dye. AB - Cotton woven fabrics which were previously dyed with a reactive dye were treated with a commercial cellulase preparation. Dyeing with a reactive dye for cotton apparently inhibited the weight loss activity and saccharification activity of cellulase. In addition, dyed cotton was treated with highly purified cellulases which were exo-type cellulases (Cellobiohydrolase I (CBH I) and Cellobiohydrolase II (CBH II)) and endo-type cellulase (Endoglucanase II (EG II)). Exo-type cellulases were inhibited more than endo-type cellulase by dyeing in the case of saccharification activity. CBH I was severely inhibited by dyeing as compared with CBH II or EG II from the viewpoint of morphological changes in the fiber surface. Dyes on the cellulose substrates severely influenced CBH I in spite of the rare modification, because CBH I hydrolyzed cellulose with true-processive action. The change in the activity of each cellulase component on dyed cotton can affect the synergistic action of cellulases. PMID- 15665467 TI - Enhanced expression of apoptin by the Myc-Max binding motif and SV40 enhancer for SCLC gene therapy. AB - Apoptin is derived from chicken anemia virus (CAV) and known to induce tumor specific apoptosis but not normal cells. The aim of this study was to use increased expression of apoptin by the Myc-Max response element (MMRE) and SV40 enhancer in small-cell lung cancer (SCLC) gene therapy. To investigate the possibility of the utilization of the MMRE, apoptin, and SV40 promoter/enhancer in targeted cancer gene therapy, adenovirus vector expressing apoptin controlled by the MMRE, and SV40 promoter/enhancer was constructed. Ad-MMRE-apoptin-enh infected SCLC cells were significantly suppressed and induced apoptosis more than those of Ad-apoptin or Ad-apoptin-enh. Infection with Ad-MMRE-apoptin-enh of normal cells did not increase apoptosis. About 85% of SCLC tumors show overexpression of the myc family, so the increased expression of apoptin by MMRE and SV40 enhancer can be used in targeted SCLC gene therapy. These results indicate that apoptin expression was increased by the MMRE and SV40 promoter/enhancer, and that this strategy can be used in SCLC targeted cancer gene therapy. PMID- 15665468 TI - Adenovirus-mediated suicide SCLC gene therapy using the increased activity of the hTERT promoter by the MMRE and SV40 enhancer. AB - Telomerase is a ribonucleoprotein complex of which the function is to add telomeric repeats to chromosomal ends. Telomerase consists of two essential components, the telomerase RNA template (hTR) and the catalytic subunit (hTERT). hTERT is expressed only in cells and tissues positive for telomerase activity, i.e., tumor or stem cells. The aim of this study was to use increased telomerase promoter activity in small-cell lung cancer (SCLC) gene therapy. The hTERT promoter and Myc-Max response elements (MMRE) in pGL3-Control vector containing SV40 enhancer resulted in strong expression of the luciferase gene only in telomerase positive and myc overexpressing SCLC cell line but not in normal human cell line. To investigate the possibility of the utilization of the MMRE, hTERT promoter, and SV40 enhancer in targeted SCLC gene therapy, adenovirus vector expressing HSV-TK controlled by the MMRE, hTERT promoter, and SV40 enhancer for the induction of telomerase positive and myc-overexpressing cancer specific cell death was constructed. SCLC cells infected with Ad-MMRE-hT-TK-enh were significantly suppressed and induced apoptosis more than those of Ad-hT-TK or Ad hT-TK-enh infected cells. Telomerase and c-myc are activated in 60 approximately 80% of SCLC, so the increased activity of telomerase promoter can be used for targeted SCLC gene therapy. These results show that the MMRE, hTERT promoter, and SV40 enhancer can be used in SCLC targeted cancer gene therapy. PMID- 15665469 TI - Transient RNA silencing of scoulerine 9-O-methyltransferase expression by double stranded RNA in Coptis japonica protoplasts. AB - RNAi (RNA interference, RNA silencing) is a powerful tool for functional genomics, but the construction of an RNAi vector(s) and the establishment of stable transformants are time-consuming and laborious. Here we report the transient RNAi of endogenous biosynthetic genes involved in isoquinoline alkaloid biosynthesis in Coptis japonica protoplasts. Double stranded (ds) RNA fragments of various lengths prepared from several different positions of the coding sequence of scoulerine 9-O-methyltransferase (SMT) were introduced into C. japonica protoplasts by polyethylene glycol-mediated transformation, and their effects were monitored by reverse transcription-polymerase chain reaction. Substantial silencing of SMT gene expression was obtained by the introduction of these SMT dsRNAs. A significant reduction in SMT protein levels was also observed. The potentials of this transient RNAi system to evaluate the functions of biosynthetic genes in Coptis alkaloid research are discussed. PMID- 15665471 TI - Purification and characterization of a yeast carbonyl reductase for synthesis of optically active (R)-styrene oxide derivatives. AB - Optically active styrene oxide derivatives are versatile chiral building blocks. Stereoselective reduction of phenacyl halide to chiral 2-halo-1-phenylethanol is the key reaction of the most economical synthetic route. Rhodotorula glutinis var. dairenensis IFO415 was discovered on screening as a potent microorganism reducing a phenacyl halide to the (R)-form of the corresponding alcohol. An NADPH dependent carbonyl reductase was purified to homogeneity through four steps from this strain. The relative molecular mass of the enzyme was estimated to be 40,000 on gel filtration and 30,000 on SDS-polyacrylamide gel electrophoresis. This enzyme reduced a broad range of carbonyl compounds in addition to phenacyl halides. Some properties of the enzyme and preparation of a chiral styrene oxide using the crude enzyme are reported herein. PMID- 15665470 TI - Isolation and characterization of a cDNA encoding phytochrome A in the non photosynthetic parasitic plant, Orobanche minor Sm. AB - In this study, the isolation and characterization of a phytochrome A (PHYA) homologous cDNA (OmPHYA) in the non-photosynthetic holoparasitic plant Orobanche minor are described. The present findings provide the first report of the presence of a PHYA homolog in the holoparasite. This study found that OmPHYA is of similar size to the other PHYAs of green plants and shows 72, 77, and 77% amino acid sequence identity with PHYA in Arabidopsis, potato, and tobacco respectively. The OmPHYA contains a conserved chromophore attachment cysteine at position 323. Although OmPHYA shows high sequence identity with other PHYAs in green plants, 13 amino acid substitutions located in both the N and C-terminal domains are observed (a total of 26 amino acids). OmPHYA is encoded by a single gene within the O. minor genome. The abundance of the OmPHYA transcript as well as nuclear translocation of OmphyA occurs in a light-dependent manner. PMID- 15665472 TI - The class V chitin synthase gene csmA is crucial for the growth of the chsA chsC double mutant in Aspergillus nidulans. AB - chsA and chsC are genes encoding class II and I chitin synthases of Aspergillus nidulans respectively. In a previous study, chsA chsC double mutants showed various growth defects, suggesting that their cell wall architecture was disorganized and their cell wall integrity diminished. Here, we constructed chsA chsC chsD triple mutants and chsA chsC csmA triple mutants to investigate the role of the class IV and class V chitin synthases, ChsD and CsmA respectively, in maintaining the cell wall structure of the chsA chsC double mutant. The former triple mutant grew a little slower than the chsA chsC double mutant, but the two showed similar phenotypes. In contrast, the latter triple mutant exhibited severe growth defects, particularly under low osmotic conditions. The levels of the csmA transcript of the wild-type strain and chsA or chsC single mutants were markedly elevated under low osmotic conditions, while that of the chsA chsC double mutants was high even under such conditions. These and other results suggest that the function of csmA is important for the maintenance of cell wall integrity and the polarized growth of the chsA chsC double mutant. PMID- 15665473 TI - Confirmation and quantification of strigolactones, germination stimulants for root parasitic plants Striga and Orobanche, produced by cotton. AB - The germination stimulants for root parasitic plants Striga and Orobanche produced by cotton (Gossypium hirsutum L.) were examined in detail. Seeds of cotton were germinated and grown on glass wool wetted with sterile distilled water in sterile filter units. The root exudate was collected daily and extracted with ethyl acetate. Each of these ethyl acetate extracts was analyzed directly by high-performance liquid chromatography linked with tandem mass spectrometry (LC/MS/MS). The results demonstrate that cotton roots exuded strigol and strigyl acetate, but no other known strigolactones such as orobanchol and alectrol. The production of strigol was detected even in the root exudate collected during the first 24 h of incubation and reached a maximum 5-7 days later. The average exudation of strigol and strigyl acetate during the incubation period was ca. 15 and 2 pg/plant/day, respectively, indicating that strigol mainly contributed to germination stimulation by the cotton root exudate. PMID- 15665474 TI - Enzymatic properties of rhea lysozyme. AB - Rhea lysozyme was analyzed for its enzymatic properties both lytic and oligomer activities to reveal the structural and functional relationships of goose type lysozyme. Rhea lysozyme had the highest lytic activity at pH 6, followed by ostrich and goose at pH 5.5-6, whereas the optimum of cassowary was at pH 5. pH profile was correlated to the net charge of each molecule surface. On the other hand, the pH optimum for oligomer substrate was found to be pH 4, indicating the mechanism of rhea catalysis as a general acid. The time-course of the reaction was studied using beta-1,4-linked oligosaccharide of N-acetylglucosamine (GlcNAc) with a polymerization degree of n ((GlcNAc)n) (n=4, 5, and 6) as the substrate. This enzyme hydrolyzed (GlcNAc)6 in an endo-splitting manner, which produced (GlcNAc)3+(GlcNAc)3 predominating over that to (GlcNAc)2+ (GlcNAc)4. This indicates that the lysozyme hydrolyzed preferentially the third glycosidic linkage from the nonreducing end. Theoretical analysis has shown the highest rate constant value at 1.5 s-1 with (GlcNAc)6. This confirmed six substrate binding subsites as goose lysozyme (Honda, Y., and Fukamizo, T., Biochim. Biophys. Acta, 1388, 53-65 (1998)). The different binding free energy values for subsites B, C, F, and G from goose lysozyme might responsible for the amino acid substitutions, Asn122Ser and Phe123Met, located at the subsite B. PMID- 15665475 TI - Synthesis and antioxidant activity of olivil-type lignans. AB - Olivil-type lignans, an enantiomeric type of natural olivil, were synthesized for the first time to evaluate the relationship between the structure of olivil and its antioxidant activity. A comparison of the antioxidant activity with that of other synthesized tetrahydrofuran lignans indicated reduced activity with the tertiary hydroxy group. A different effect of the two phenolic groups of olivil on the antioxidant activity was also observed. PMID- 15665476 TI - Stimulation of mycelia growth in several mushroom species by rice husks. AB - When supplemented to the culture medium of mushroom Coprinus cinereus, rice husks soaked beforehand in methanol stimulated mycelia growth up to a concentration of 80 mg/ml dose-dependently, whereas the non-treated stimulated mycelia growth up to 20 mg/ml. This result suggests the existence of both stimulatory and inhibitory substances in rice husks. Since momilactone A (MLA) is recognized as one of the phytoalexins in rice husks, its biological activity against mycelia growth was tested. Momilactone A inhibited mycelia growth at 5 microg/disc, whereas the methanol extract of husks did so at 1 mg/disc, wherein 0.2 microg of MLA was estimated by LC/MS/MS. Thus the phytoalexins including MLA should inhibit mycelia growth. Rice husks stimulated mycelia growth in some edible mushroom species such as Grifola frondosa (maitake), Lentinus edodes (shiitake), Pleurotus eryngii (eringi), and P. ostreatus (hiratake). Our findings might lead to the development of new profitable cultivation methods for mushrooms using rice husks. PMID- 15665477 TI - Preparation and properties of gelatin-immobilized beta-glucosidase from Pyrococcus furiosus. AB - Hyperthermostable beta-glucosidase from Pyrococcus furiosus was enclosed in gelatin gel by cross-linking with transglutaminase. Gelatin-immobilized beta glucosidase was considerably more thermostable than the native enzyme. Lyophilized immobilisate was stored at 90 degrees C for 1 month without loss of activity. The immobilized beta-glucosidase catalyzed transglucosylation of 5 phenylpentanol with 10.0 equivalent of cellobiose at pH 5.0 and 70 degrees C for 12 h to afford 5-phenylpentyl beta-D-glucopyranoside in 41% yield. The immobilized enzyme was more effective than the native one in transglucosylation. The gelatin-immobilized Pfu-beta-glucosidase recovered from the first run of the reaction was reusable on successive runs. PMID- 15665478 TI - Occurrence of Lewis a epitope in N-glycans of a glycoallergen, Jun a 1, from mountain cedar (Juniperus ashei) pollen. AB - We have determined the structures of N-glycans linked to major allergens in the mountain cedar (Juniperus ashei) pollen, Jun a 1. First, two kinds of the pollen glycoallergen (Jun a 1-A and Jun a 1-B) were purified from partially purified Jun a 1 by cation exchange chromatography. The N-glycans were liberated by hydrazinolysis from the two glycoallergens and the resulting sugar chains were N acetylated and then coupled with 2-aminopyridine. Three pyridylaminated sugar chains were purified by reversed-phase HPLC and size-fractionation HPLC from Jun a 1-A and Jun a 1-B respectively. The structures were determined by a combination of exo- and endo-glycosidase digestions, two dimensional sugar chain mapping, and electrospray ionization mass spectrometry (ESI-MS) analysis. Structural analysis indicated that Lewis a epitope (Galbeta1-3(Fucalpha1-4)GlcNAcbeta1-) occurs in the N-glycans of the pollen allergens. PMID- 15665479 TI - Short-step and scalable synthesis of (+/-)-cytoxazone. AB - A five-step and scalable synthesis of racemic cytoxazone, a novel cytokine modulator, was accomplished in a total yield of 51% from p-methoxycinnamyl alcohol without any protective groups. The keystep was the new one-pot azidohydroxylation procedure by the combined use of NaN3-H2O2-CH3CN. The epoxidation of an olefin by means of an in situ-formed iminohydroperoxide worked well, accompanied by the concomitant regioselective ring opening reaction of the resulting highly reactive epoxide with an azide ion. PMID- 15665480 TI - Electron transfer ability from NADH to menaquinone and from NADPH to oxygen of type II NADH dehydrogenase of Corynebacterium glutamicum. AB - Type II NADH dehydrogenase of Corynebacterium glutamicum (NDH-2) was purified from an ndh overexpressing strain. Purification conferred 6-fold higher specific activity of NADH:ubiquinone-1 oxidoreductase with a 3.5-fold higher recovery than that previously reported (K. Matsushita et al., 2000). UV-visible and fluorescence analyses of the purified enzyme showed that NDH-2 of C. glutamicum contained non-covalently bound FAD but not covalently bound FMN. This enzyme had an ability to catalyze electron transfer from NADH and NADPH to oxygen as well as various artificial quinone analogs at neutral and acidic pHs respectively. The reduction of native quinone of C. glutamicum, menaquinone-2, with this enzyme was observed only with NADH, whereas electron transfer to oxygen was observed more intensively with NADPH. This study provides evidence that C. glutamicum NDH-2 is a source of the reactive oxygen species, superoxide and hydrogen peroxide, concomitant with NADH and NADPH oxidation, but especially with NADPH oxidation. Together with this unique character of NADPH oxidation, phylogenetic analysis of NDH-2 from various organisms suggests that NDH-2 of C. glutamicum is more closely related to yeast or fungal enzymes than to other prokaryotic enzymes. PMID- 15665481 TI - Functions of family-22 carbohydrate-binding module in Clostridium thermocellum Xyn10C. AB - Clostridium thermocellum xylanase Xyn10C (formerly XynC) is a modular enzyme, comprising a family-22 carbohydrate-binding module (CBM), a family-10 catalytic module of the glycoside hydrolases, and a dockerin module responsible for cellulosome assembly consecutively from the N-terminus. To study the functions of the CBM, truncated derivatives of Xyn10C were constructed: a recombinant catalytic module polypeptide (rCM), a family-22 CBM polypeptide (rCBM), and a polypeptide composed of the family-22 CBM and CM (rCBM-CM). The recombinant proteins were characterized by enzyme and binding assays. Although the catalytic activity of rCBM-CM toward insoluble xylan was four times higher than that of rCM toward the same substrate, removal of the CBM did not severely affect catalytic activity toward soluble xylan or beta-1,3-1,4-glucan. rCBM showed an affinity for amorphous celluloses and insoluble and soluble xylan in qualitative binding assays. The optimum temperature of rCBM-CM was 80 degrees C and that of rCM was 60 degrees C. These results indicate that the family-22 CBM of C. thermocellum Xyn10C not only was responsible for the binding of the enzyme to the substrates, but also contributes to the stability of the CM in the presence of the substrate at high temperatures. PMID- 15665482 TI - Design and facile synthesis of neoglycolipids as lactosylceramide mimetics and their transformation into glycoliposomes. AB - Neoglycolipids composed of disaccharide glycoside and phospholipid were designed and prepared as mimetics of lactosylceramide. The lactosyl- and N acetyllactosaminyl-phospholipids (Lac-DPPA and LacNAc-DPPA) were enzymatically synthesized from lactose and LacNAc respectively by cellulase-mediated condensation with 1,6-hexanediol, followed by conjugation of the resulting glycosides and dipalmitoylphosphatidyl choline (DPPC) mediated by Streptomyces phospholipase D. Alternatively, allyl beta-lactoside was ozonolyzed to give an aldehyde, which was condensed with dipalmytoyl phosphatidyl ethanolamine to afford a second type of glycolipid (Lac-DPPE). NMR spectroscopy indicated that the neoglycolipids behave differently in different solvent systems. X-ray diffraction clearly showed that multilamellar vesicles (MLVs) of Lac-DPPE and Lac DPPA-MLV are in the bilayer gel phase at 20 degrees C, whereas those of Lac-DPPE MLV were in the lamellar liquid-crystalline phase at 50 degrees C. Differential scanning calorimetry showed that Lac-DPPE-MLV had complex thermotropic behavior depending on the incubation conditions. After a long incubation at 10 degrees C, endothermic transitions are observed at 39.6, 42.3 degrees C, and 42.9 degrees C. These neoglycolipids have the ability to trap calcein, a chelating derivative of fluorescein, in MLVs and showed specific binding to lectin in plate assays using fluorescently labeled compounds. PMID- 15665483 TI - Modulating effect of sesamin, a functional lignan in sesame seeds, on the transcription levels of lipid- and alcohol-metabolizing enzymes in rat liver: a DNA microarray study. AB - Sesamin, a major lignan in sesame seeds, has multiple functions such as cholesterol-lowering and anti-hypertensive activities. To investigate the effect of sesamin on gene expression in the liver, a DNA microarray analysis was carried out. The ingestion of sesamin dissolved in olive oil up-regulated the expression of 38 genes, 16 of which encode proteins possessing a lipid-metabolizing function, and 16 of which encode proteins possessing a xenobiotic/endogenous substance metabolizing function. In particular, sesamin significantly increased the expression of beta-oxidation-associated enzymes in peroxisomes and auxiliary enzymes required for degradation, via the beta-oxidation pathway, of unsaturated fatty acids in mitochondria. The ingestion of sesamin also resulted in an increase in the gene expression of acyl-CoA thioesterase involved in acyl-CoA hydrolase and very-long-chain acyl-CoA thioesterase. Interestingly, it induced the expression of the gene for aldehyde dehydrogenase, an alcohol-metabolizing enzyme. These results suggest that sesamin regulates the metabolism of lipids, xenobiotics, and alcohol at the mRNA level. PMID- 15665484 TI - Purification, characterization, and antifungal activity of chitinases from pineapple (Ananas comosus) leaf. AB - Three chitinases, designated pineapple leaf chitinase (PL Chi)-A, -B, and -C were purified from the leaves of pineapple (Ananas comosus) using chitin affinity column chromatography followed by several column chromatographies. PL Chi-A is a class III chitinase having a molecular mass of 25 kDa and an isoelectric point of 4.4. PL Chi-B and -C are class I chitinases having molecular masses of 33 kDa and 39 kDa and isoelectric points of 7.9 and 4.6 respectively. PL Chi-C is a glycoprotein and the others are simple proteins. The optimum pHs of PL Chi-A, -B, and -C toward glycolchitin are pH 3, 4, and 9 respectively. The chitin-binding ability of PL Chi-C is higher than that of PL Chi-B, and PL Chi-A has lower chitin-binding ability than the others. At low ionic strength, PL Chi-B exhibits strong antifungal activity toward Trichoderma viride but the others do not. At high ionic strength, PL Chi-B and -C exhibit strong and weak antifungal activity respectively. PL Chi-A does not have antifungal activity. PMID- 15665485 TI - Screening for tyrosinase inhibitors among extracts of seashore plants and identification of potent inhibitors from Garcinia subelliptica. AB - The tyrosinase inhibitory activity of methanol extracts of the leaves of 39 plant species growing on the seashore of Iriomote island (Okinawa, Japan) was investigated. The extracts of Hibiscus tiliaceus, Carex pumila, and Garcinia subelliptica showed potent activity among them. The inhibitors in the extract of Garcinia subelliptica were purified by assay-guided fractionation to give two biflavonoids. These were known compounds (2R,3S-5,7,4',5'',7'',3''',4''' heptahydroxy flavanone[3-8''] flavone and 5,7,4',5'',7'',3''',4'''-heptahydroxy[3 8''] biflavanone), although their strong inhibitory activity toward tyrosinase is revealed for the first time in this work. One of these biflavonoids (2R,3S 5,7,4',5'',7'',3''',4'''-heptahydroxy flavanone[3-8''] flavone) showed much stronger activity (IC50 2.5 microM) than that of kojic acid (IC50 9.1 microM) when L-tyrosine was used as the substrate. PMID- 15665486 TI - Stimulation of IL-8 production by Aralia cordate lectin in human colon carcinoma Caco-2 cells. AB - Some lectins are known to stimulate interleukin-8 (IL-8) productions in human colon carcinoma Caco-2 cells. Since IL-8 may cause deleterious effects, we examined this stimulatory activity of Aralia cordate lectin (ACL) and Wasabia japonica lectin (WJL), both of which we isolated recently. The results indicate that ACL exhibited strong stimulatory activity for IL-8 protein production, while WJL showed marginal activity. The activity of ACL was associated with high enhancement of IL-8 gene expression. The effect of ACL was abolished almost completely in the presence of brefeldin A, indicating that internalization into cells is necessary for ACL to exert activity. The findings suggest that ingestion of a large amount of raw vegetable Aralia cordate might cause unfavorable effects on the colon. PMID- 15665487 TI - Improvement of the Aspergillus oryzae enolase promoter (P-enoA) by the introduction of cis-element repeats. AB - We constructed a protein expression vector with an improved enoA promoter that harbored 12 tandem repeats of the cis-acting element (region III) of Aspergillus oryzae. The improved promoter yielded reporter beta-glucuronidase (GUS) activity approximately 30-fold of the original promoter. Northern blot analysis confirmed that GUS expression was increased at the transcriptional level. The transformant harboring seven copies of the novel vector showed more than 100,000 U/mg GUS protein, which was approximately 30% of all the cell-free soluble proteins. PMID- 15665488 TI - Y-700, a novel inhibitor of xanthine oxidase, suppresses the development of colon aberrant crypt foci and cell proliferation in 1,2-dimethylhydrazine-treated mice. AB - Y-700, 1-[3-cyano-4-(2,2-dimethylpropoxy)phenyl]-1H-pyrazole-4-carboxylic acid, is a newly synthesized inhibitor of xanthine oxidase. This study found that feeding of Y-700 suppressed the development of colonic aberrant crypt foci, precursor lesions of colon cancer, and cell proliferation in 1,2 dimethylhydrazine-treated mice, accompanied by reduced serum urate. These results suggest that Y-700 is a useful agent for the prevention of colon tumorigenesis and that xanthine oxidase plays an important role in the development of colon cancer. PMID- 15665489 TI - Ergosterol peroxide, an apoptosis-inducing component isolated from Sarcodon aspratus (Berk.) S. Ito. AB - Among the lipophilic extracts of seven traditional edible mushrooms, the acetone extract of Sarcodon aspratus markedly inhibited the growth of HL60 human leukemia cells and induced apoptosis after 24 h incubation. The major active component was identified as ergosterol peroxide by NMR and ESI-MS analysis. Ergosterol peroxide completely inhibited growth and induced apoptosis of HL60 cells at a concentration of 25 microM. PMID- 15665490 TI - Identification of insertion sequence from a gamma-hexachlorocyclohexane degrading bacterium, Sphingomonas paucimobilis UT26. AB - Tn5-derived mutants of the gamma-hexachlorocyclohexane-degrading bacterium Sphingomonas paucimobilis UT26 were genetically characterized, and an endogenous insertion sequence (IS) which belongs to the IS1380 family was identified. The IS, named ISsp1, existed as multi copies in UT26, and its transposition appeared to be activated during the process of Tn5-mutagenesis. It was found that transposon mutagenesis can cause endogenous mutations. PMID- 15665491 TI - Contribution of conserved Asn residues to the inhibitory activities of Kunitz type protease inhibitors from plants. AB - Plant Kunitz-type protease inhibitors contain a conserved Asn residue in the N terminal region. To investigate the role of Asn residue in protease inhibitory activities, Erythrina variegata trypsin inhibitor a (ETIa), E. variegata chymotrypsin inhibitor (ECI), and their mutants, ETIa-N12A and ECI-N13A, were used. Both mutants exhibit weaker inhibitory activities toward their cognate proteases than the wild-type proteins and were readily cleaved at reactive sites. Furthermore, kinetic analysis of the interactions of the mutated proteins with their cognate proteases by surface plasmon resonance (SPR) measurement indicated that replacements of the Asn residue mainly affected dissociation rate constants. The conserved Asn residues of Kunitz-type inhibitors play an important role in exhibiting effective inhibitory activity by stabilizing the structures of the primary binding loop and protease-inhibitor complex. PMID- 15665492 TI - Characterization of apoptosis induced by fucoxanthin in human promyelocytic leukemia cells. AB - Apoptosis induced by fucoxanthin in HL-60 cells was associated with a loss of mitochondrial membrane potential at an early stage, but not with an increase in reactive oxygen species. Fucoxanthin treatment caused cleavages of procaspase-3 and poly (ADP-ribose) polymerase without any effect on the protein level of Bcl 2, Bcl-X(L), or Bax. Apoptosis induction by fucoxanthin may be mediated via mitochondrial membrane permeabilization and caspase-3 activation. PMID- 15665494 TI - Enantioselective synthesis of four isomers of 3-hydroxy-4-methyltetradecanoic acid, the constituent of antifungal cyclodepsipeptides W493 A and B. AB - Four possible stereoisomers of 3-hydroxy-4-methyltetradecanoic acid were enantioselectively synthesized by using Sharpless epoxidation and a subsequent epoxide-ring opening reaction with trimethylaluminum as the key steps. The absolute configuration of the beta-oxyacid component of antifungal cyclodepsipeptides W493 A and B was consequently determined as 3S,4R. PMID- 15665493 TI - Optimum culture conditions for the production of N-substituted formamide deformylase by Arthrobacter pascens F164. AB - We investigated the optimum culture conditions for the production of a novel enzyme, N-substituted formamide deformylase, which acts mainly on N benzylformamide, in Arthrobacter pascens F164. The highest enzyme activity was obtained when this strain F164 was cultivated in a synthetic medium with N benzylformamide as sole nitrogen source. This deformylase was found to be an inducible enzyme depending on N-benzylformamide. PMID- 15665495 TI - Transformation of Escherichia coli mediated by natural phospholipids. AB - Transformation system for Escherichia coli based upon introduction of plasmid DNA by natural phospholipids has been developed. Transformants are easily obtained by treatment with natural phospholipids such as phosphatidylethanolamine, phosphatidylcoline, and phosphatidylserine, where the presence of MgCl2 or CaCl2 is essential. This method of transformation is applicable not only for small plasmid pHSG399 (2.3 Kb) but also for giant plasmid R6K (100 Kb). PMID- 15665496 TI - Pradimicin resistance of yeast is caused by a mutation of the putative N glycosylation sites of osmosensor protein Sln1. AB - Pradimicin, a mannose-binding antifungal antibiotic, induces apoptosis-like cell death in Saccharomyces cerevisiae. Previously we found that the substitution of the 74th amino acid from glycine to cysteine in Ypd1 yields a mutant resistant to pradimicin. In this study, the involvement of a membrane-spanning osomosensor, Sln1, which is located upstream of Ypd1, was investigated. A mutant, sln1 DeltaNG, that lacks the putative N-glycosylation sites in the extracellular domain became resistant to pradimicin. On the other hand, the null mutants of Ssk1, Pbs2, and Hog1, which are located downstream of the Sln1 cascade, were sensitive to pradimicin as well as the wild-type strain. In conclusion, pradimicin exerts its fungicidal action with the involvement of Sln1, but the downstream branch, Ssk1 and the HOG pathway, is not involved. PMID- 15665497 TI - Sugar responsible and tissue specific expression of a gene encoding AtCIPK14, an Arabidopsis CBL-interacting protein kinase. AB - Expression of AtCIPK14 (an Arabidopsis CBL-interacting protein kinase 14) was induced by metabolic sugars. Two A/T-rich sequences similar to elements involved in sugar-inducible expression of other genes were found within the -183 bp 5' region of the AtCIPK14 promoter that was responsible for the sugar induction. Histochemical analysis using a reporter gene indicated vascular-specific expression of AtCIPK14. PMID- 15665499 TI - Oral administration of freeze-dried kefir reduces intestinal permeation of and oral sensitization to ovalbumin in mice. AB - An increase in plasma ovalbumin concentrations after intragastric administration of ovalbumin was suppressed by concomitant freeze-dried kefir in BALB/c mice. Serum levels of ovalbumin-specific immunoglobulin G and proliferation of splenic mononuclear cells in mice immunized orally with ovalbumin were suppressed by feeding freeze-dried kefir. We propose that kefir reduces intestinal permeation of food antigen, which contributes to suppression of oral sensitization. PMID- 15665498 TI - Interaction of modified cyclodextrins with cytochrome P-450. AB - The effects of modified cyclodextrins (CDs) hydroxypropyl-beta-CD and methyl-beta CD were studied in vitro on cDNA-expressed human cytochrome P-450 (CYP) activities (CYP1A2, CYP2C9, CYP2C19, CYP2D6, and CYP3A4). The modified CDs inhibited the activities of CYP2C19 and CYP3A4 while enhancing CYP2C9 activity by 140 to 176% relative to the control values at lower concentrations. In addition, methyl-beta-CD inhibited CYP1A2 and CYP2D6 at higher concentrations. PMID- 15665500 TI - Regeneration of bacteriorhodopsin from thermally unfolded bacterio-opsin and all trans retinal at high temperatures. AB - The temperature dependence of regeneration of bacteriorhodopsin (bR) from its apoprotein, bacterio-opsin (bO), and all-trans retinal was investigated using two different procedures to probe the structural properties of bO at high temperatures. Regeneration experiments performed at 25 degrees C after incubation of bO within the temperature range of 35-75 degrees C indicate that irreversible thermal unfolding begins at 50 degrees C. When bO is incubated for one hour and mixed with retinal at the same elevated temperatures, however, a greater extent of regeneration to bR occurs, even at temperatures ranging from 50 to 65 degrees C. These experimental results indicate that regeneration of bR occurs from thermally unfolded bO and suggest dynamic structural fluctuation of bO in the unfolded state. PMID- 15665501 TI - Acid hydrolysis of protein in a microcapillary tube for the recovery of tryptophan. AB - The acid hydrolysis of proteins was miniaturized and simplified by employing microcapillary tubes (100 microl in volume) with 6 M HCl containing 1% 2 mercaptoethanol and 3% phenol for an amino acid compositional analysis. The method not only eliminated the laborious evacuation step for the hydrolysis tube but also decreased the destruction of tryptophan during hydrolysis. The recovery of tryptophan was 79% by acid hydrolysis at 145 degrees C for 4 h. Since the acid mixture could be removed under vacuum, the hydrolysate was subjected to an amino acid analysis without neutralization or dilution. PMID- 15665502 TI - Occurrence of fructosyl-amino acid oxidase-reactive compounds in fungal cells. AB - Fructosyl-amino acid oxidase (FAOD)-reactive fraction (FRY) was found in commercial yeast extract. FRY showed very hydrophilic property and was adsorbed to phenylboronate silica gel, indicating that it contained the Amadori compound. TLC and amino acid analyses revealed that glucosone, lysine, and arginine were produced from FRY after incubation with FAOD. TOF-MS analysis confirmed that FRY is a mixture of fructosyl lysine and fructosyl arginine. These compounds were also detected in mycelial extract of an FAOD-producer, Aspergillus terreus GP1, grown on the minimum medium, suggesting that a glycation reaction occurs in fungal cells and that FAOD acts toward the resultant Amadori compounds. PMID- 15665503 TI - Functional characterization of contractile vacuole isolated from Amoeba proteus. AB - Contractile vacuoles (CVs) released from cells of Amoeba proteus were used to analyze its function in vitro. When CV was transferred to a hypertonic medium, its volume decreased within 10 sec. When it was subsequently returned to its original medium, it quickly started swelling. However, it ruptured before recovering its initial volume. These results suggested that the CV membrane is semi-permeable and that the fluid is collected by the osmotic gradient in vivo. The water permeability of membrane of isolated CV was calculated from the rate of osmotic volume change to be 0.94 microm/sec . OsM. This high value suggested that CV membrane is equipped with water channel. CV contracted (or burst) quickly upon addition of 1 mM ATP. Contraction was induced by ATP, but not by other nucleotides, GTP, ITP, ADP, or the analogues of ATP, AMP-PNP and ATPgammaS. It was suggested that the contraction of isolated CV was caused by increase in the tension of its membrane by ATP. PMID- 15665504 TI - Immunohistochemical localization of Klotho protein in brain, kidney, and reproductive organs of mice. AB - Klotho mutant mouse (kl-/-), a mouse model for human aging, exhibits various phenotypes in a wide range of organs including arteriosclerosis, neural degeneration, skin and gonadal atrophy, pulmonary emphysema, calcification of soft tissues, and cognition impairment. Klotho mRNA, however, is expressed only in brain, kidney, reproductive organs, pituitary gland, and parathyroid gland. Therefore it remains to be elucidated how lack of Klotho protein in these limited organs leads to the variety of phenotypes. To shed light on mechanisms by which Klotho protein acts on distant targets, we examined localization of Klotho protein in brain, kidney, and reproductive organs, and analyzed brain and kidney in kl-/- mice searching for changes in target regions in these organs. In brain, Klotho proteins were localized at choroid plexus, where the proteins were dominantly localized at the apical plasma membrane of ependymal cells. In kl-/- brain, reduction of synapses was evident in hippocampus, suggesting a role of Klotho as a humoral factor in cerebrospinal fluid. Klotho proteins in kidney localized at distal renal tubules. Interestingly, in kl-/-mice, type IIa Na/phosphate (Pi) cotransporters, which function at the proximal renal tubules in reabsorption of phosphate ions, were translocated. This suggests that Klotho protein in kidney is implicated in calcium homeostasis which regulates localization of type IIa Na/Pi cotransporters via parathyroid hormone (PTH). Klotho proteins in reproductive organs were expressed only in mature germ cells, although in kl-/- mice germ cell maturation was arrested at earlier stages. Thus, Klotho proteins not only function as a humoral factor, but also are implicated in hormonal regulation, which may explain why mutation of klotho gene results in a variety of phenotypes. PMID- 15665505 TI - Adenosine A2A receptor facilitates calcium-dependent protein secretion through the activation of protein kinase A and phosphatidylinositol-3 kinase in PC12 cells. AB - Adenosine modulates a variety of cellular functions including calcium-dependent exocytosis. Activation of adenosine A(2A) receptor (A(2A)-R) facilitates neurotransmitter release in some cell types, although the underlying mechanisms are not fully understood. In this study, we found that treatment of PC12 cells with the A(2A)-R agonist CGS21680 promotes calcium-evoked secretion of the fusion protein between neuropeptide Y and modified yellow fluorescence protein (NPY Venus). CGS21680 treatment of PC12 cells transiently increased the phosphorylation of p38 and JNK MAP kinases and Akt, as well as that of ATF2 and CREB, reaching maximal levels at around 10-15 min of CGS21680 treatment. Importantly, pretreatment of PC12 cells with the PI3K inhibitor LY294002, together with the protein kinase A (PKA) inhibitor KT5720, significantly inhibited CGS21680 enhancement of calcium-dependent NPY-Venus release. Moreover, expression of a dominant-negative form of Akt and the PKA inhibitory polypeptide protein kinase inhibitor (PKI) co-operatively inhibited the facilitating effect of CGS21680 on secretion of NPY-Venus. These data suggest that the PI3K-Akt and PKA pathways play a critical role in A(2A)-R-mediated facilitation of calcium dependent secretion. We also found that CGS21680 treatment promoted recruitment of the NPY-Venus-containing vesicles to the proximity of the plasma membrane at around 10-15 min of CGS21680 treatment, which may in part account for the facilitated secretion by A(2A)-R activation. PMID- 15665506 TI - Body composition in diabetic subjects with chronic kidney disease: interest of bio-impedance analysis, and anthropometry. AB - BACKGROUND/AIMS: Lean body mass (LBM) is reduced in uremia, but this has not been reported in diabetic nephropathy. SUBJECTS AND METHODS: We compared predicted % LBM to DEXA measurements in 10 non-diabetic uremic, 10 non-uremic diabetic and 10 uremic diabetic subjects matched for age, gender and BMI. We also measured % LBM by anthropometry, bio-impedance analysis (BIA) and compared them with DEXA in 49 diabetic subjects with a wide range of renal failure. The results were compared and a Bland & Altman procedure was performed. Associations between glomerular filtration rate (GFR) and % LBM were tested. RESULTS: In matched groups, predicted % LBM values were overestimated in non-diabetic uremic subjects, and underestimated in non-uremic diabetic subjects. In uremic diabetic subjects, the error was intermediary. As compared to DEXA (% LBM: 69.0 +/- 7.1%), measurement of % LBM by anthropometry (71.4 +/- 8.0%, p < 0.05) and BIA (67.2 +/- 7.6%, p < 0.05) were biased in the 49 diabetic subjects. The mean of anthropometric and BIA (Ant+BIA) were similar to DEXA results (69.3 +/- 6.8%, p = 0.64), with best correlation coefficients and Bland & Altman plots. GFR was correlated to % LBM assessed by DEXA, BIA and Ant+BIA. CONCLUSION: In diabetic subjects with chronic kidney disease, LBM should be measured, rather than predicted. A good evaluation is possible, even without DEXA. PMID- 15665508 TI - Normative data of penile length for term Chinese newborns. AB - Careful examination and assessment of penile length in the newborn is important because micropenis is associated with hypothalamic disorders. This prospective cross-sectional study was designed to establish the norm of penile length for term Hong Kong Chinese newborns. 4,628 full-term healthy male newborns were enrolled in the study. Penile length was measured from pubic ramus to the tip of the glans penis by placing the end of a straight edge ruler against the pubic ramus. A gestation-associated graph of 3rd, 10th, 25th, 50th, 75th, 90th and 97th centiles for the penile length was created. The penile length was significantly correlated with gestational age, birth weight and body length. When compared with published data for other populations, Hong Kong newborns have significantly shorter penile length. The norm of the penile length is useful in diagnostic and therapeutics for the ethnic Chinese male newborns. PMID- 15665507 TI - Vitamin and mineral supplements use in German children and adolescents between 1986 and 2003: results of the DONALD Study. AB - BACKGROUND/AIMS: Dietary supplements may contribute to a considerable proportion to micronutrient intake. However, little is known about the consumption of supplements in children and adolescents, especially in Germany. We therefore examined patterns and time trends in supplement consumption in healthy children and adolescents. METHODS: A total of 5,990 3-day records from 931 subjects 2-18 years of age from the DONALD Study between 1986 and 2003 were examined. RESULTS: (a) Supplement type: A total of 166 different supplements were reported: 49% vitamin-mineral combinations, 31% vitamin, 13% mineral, 7% fluorine supplements. 12% (vitamin) and 13% (mineral) were single nutrient supplements. Vitamin C (72%), B(1) (57%), B(2) (54%), calcium (44%), magnesium (31%) and phosphorus (20%) were the most frequent added nutrients. (b) Users: In 25.8% (males 13.2%, females 12.6%) of the records, supplement usage was documented. Fluorine supplements were by far the most often consumed items (18.1%) followed by vitamins (4.5%), vitamin-mineral combinations (3.6%), minerals (2.4%), and multiple usage (2.6%). (c) Time trend: We found a marked time trend in supplement consumption in the past 18 years with a peak between 1994 and 1996 and lower usage before and after that time (independent of age and gender). (d) Associated factors: Supplement usage was influenced by age, year of study, season, smoking and number of persons in families, education level and employment of mothers, whereas gender or the number of children per family failed to have any effect. CONCLUSION: Supplement usage is a common behaviour in German children and adolescents and changing with time. Type and frequency of supplement usage is age dependent. Those nutrients found mostly in supplements are not the critical ones. In evaluations of children's diet it is mandatory to separate fluorine from other supplements. PMID- 15665509 TI - Low-density lipoprotein cholesterol: association with mortality and hospitalization in hemodialysis patients. AB - BACKGROUND/AIMS: Hypocholesterolemia is a common finding in hospitalized elderly people, critically ill surgical patients, septic patients and end-stage renal disease patients. The different effect of lipid subfractions on patients with end stage renal disease has never been demonstrated. We aim to study the effect of lipid subfractions on hospitalization and mortality in maintenance hemodialysis (MHD) patients. METHODS: Lipid subfractions, including total cholesterol (TC), triglycerides (TG), high-density lipoprotein cholesterol (HDL-C) and low-density lipoprotein cholesterol (LDL-C) were measured in 210 patients with MHD in a single dialysis center. Patients were stratified into three groups based on the tertiles of lipid levels, and differences in patient characteristics and survival were evaluated. RESULTS: Of a total of 22 deceased patients in our MHD cohort, infection-related mortality (50%) was higher than cardiovascular-related mortality (18.2%). Significant differences (p < 0.05) in the duration and frequency of hospitalization and in mortality events were observed when patients were divided into different subgroups according to the tertiles of baseline TC and LDL-C levels. Patients with lower LDL had significantly lower levels of albumin, TC and TG. The LDL-C tertiles were similar in terms of age, hypertension, diabetes, biochemical results, hematocrit, adequacy of hemodialysis and normalized protein catabolism rate. Both TC and LDL-C predicted survival (p < 0.001), but not TG and HDL-C in the Kaplan-Meier model. The Cox proportional hazard model demonstrated that baseline serum LDL-C was the best lipid subfraction in predicting all-cause death with an adjusted hazard ratio (95% confidence interval) for each 10 mg/dl of 0.752 (0.631-0.898; p = 0.002). CONCLUSIONS: We firstly demonstrated that lipid subfractions, including TC and LDL-C, predict poor outcomes in a MHD cohort with high infection-related mortality. PMID- 15665510 TI - Systemic administration of argatroban reduces secondary brain damage in a rat model of intracerebral hemorrhage: histopathological assessment. AB - This study investigated the effects of argatroban, a thrombin inhibitor, on brain edema and inflammation in a rat intracerebral hemorrhage (ICH) model. ICH was induced by injecting collagenase IV into the right caudate nucleus. Argatroban was administered intraperitoneally. Argatroban reduced brain edema from 44.6 to 14.3 microl at 72 h. Infiltration of polymorphonuclear leukocytes at 24 h and monocyte/macrophage at 24 and 72 h was significantly suppressed by argatroban. Argatroban did not increase the volume of hematoma. Systemic administration of argatroban reduced secondary brain damage including edema and inflammation in a rat ICH model. PMID- 15665511 TI - Erythrocyte signal transduction pathways, their oxygenation dependence and functional significance. AB - Erythrocytes play a key role in human and vertebrate metabolism. Tissue O2 supply is regulated by both hemoglobin (Hb)-O2 affinity and erythrocyte rheology, a key determinant of tissue perfusion. Oxygenation-deoxygenation transitions of Hb may lead to re-organization of the cytoskeleton and signalling pathways activation/deactivation in an O2-dependent manner. Deoxygenated Hb binds to the cytoplasmic domain of the anion exchanger band 3, which is anchored to the cytoskeleton, and is considered a major mechanism underlying the oxygenation dependence of several erythrocyte functions. This work discusses the multiple modes of Hb-cytoskeleton interactions. In addition, it reviews the effects of Mg2+, 2,3-diphosphoglycerate, NO, shear stress and Ca2+, all factors accompanying the oxygenation-deoxygenation cycle in circulating red cells. Due to the extensive literature on the subject, the data discussed here, pertain mainly to human erythrocytes whose O2 affinity is modulated by 2,3-diphosphoglycerate, ectothermic vertebrate erythrocytes that use ATP, and to bird erythrocytes that use inositol pentaphosphate. PMID- 15665512 TI - Protein phosphatase 2A regulatory subunit Bbeta promotes MAP kinase-mediated migration of A431 cells. AB - BACKGROUND/AIMS: Phosphatases are involved in regulation of MAP kinase (MAPK). A431 cells migrate on collagen after EGF stimulation using MAPK. To clarify the involvement of PP2A in this MAPK-dependent migration, the expression of an isoform of the B regulatory subunit was inhibited. METHODS: An antisense sequence corresponding to Bbeta cDNA was transfected into A431 cells. Their migratory activity on collagen was examined using Transwell, and MAPK phosphorylation and phosphatase activity were measured, and the results were compared with those obtained with mock-transfected cells. RESULTS: Antisense-transfected cells showed less Bbeta protein and phosphatase activity than mock-transfected controls. Migration of antisense-transfected cells showed a low response to EGF. The response of MAPK phosphorylation of antisense-transfected cells to EGF stimulation and adhesion to collagen in the presence or absence of EGF were markedly decreased. Phosphatase activity of PP2A-Bbeta also did not respond to EGF, collagen or EGF plus collagen, and remained at low levels. CONCLUSION: These results suggested that PP2A-Bbeta promotes cell migration through the MAPK cascade. PMID- 15665513 TI - TNF-alpha activates MUC2 transcription via NF-kappaB but inhibits via JNK activation. AB - The molecular mechanisms responsible for TNF-alpha-mediated MUC2 intestinal mucin up-regulation in HM3 colon adenocarcinoma cells were analyzed using promoter reporter assays of the 5'-flanking region of the MUC2 gene. Chemical inhibitors, mutant reporter constructs, and EMSA confirmed I-kappaB/NF-kappaB pathway involvement. Wortmannin, LY294002 and dominant negative Akt, as well as dominant negative NF-kappaB-inducing kinase (NIK) inhibited MUC2 reporter transcription, indicating that both phosphatidylinositol-3-OH kinase (PI3K)/Akt signaling pathway and NIK pathways mediate the effects of TNF-alpha. Wortmannin inhibited NF-kappaB binding and transcriptional activity without inhibiting NF-kappaB translocation to the nucleus, indicating that PI3K/Akt signaling activates NF kappaB transcriptional activity directly. Our results demonstrate that TNF-alpha up-regulates MUC2 in human colon epithelial cells via several signaling pathways, involving both NIK and PI3K/Akt, which converge at the common IKK/I-kappaB/NF kappaB pathway. TNF-alpha activated JNK, but JNK inhibitor SP600125 and dominant negative cJun consistently activated transcription, revealing a negative role for this signaling pathway. Thus TNF-alpha causes a net up-regulation of MUC2 gene expression in cultured colon cancer cells because NF-kappaB transcriptional activation of this gene is able to counter-balance the suppressive effects of the JNK pathway. However, the existence of this inhibitory JNK pathways suggests a mechanism whereby--in the absence of NF-kappaB activation--TNF-alpha production during inflammation in vivo could actually inhibit MUC2 production, giving rise to the defective mucosal protection which characterizes inflammatory bowel disease. PMID- 15665514 TI - The antidiabetic agent rosiglitazone upregulates SERCA2 and enhances TNF-alpha- and LPS-induced NF-kappaB-dependent transcription and TNF-alpha-induced IL-6 secretion in ventricular myocytes. AB - Positive hemodynamic effects of the antidiabetic agent rosiglitazone on perfused whole hearts have recently been described, but the mechanisms regulating these effects are not well understood. This study reports the effects of rosiglitazone on calcium regulation in isolated neonatal rat ventricular myocytes by measurement of Ca2+ transient decay rates and SERCA2 gene expression, and shows that rosiglitazone enhances known cardioprotective signaling pathways. Myocyte treatment with 10 micromol/L rosiglitazone accelerated Ca2+ transient decay rates by approximately 30%, enhanced SERCA2 mRNA levels by approximately 1.5-fold and SERCA2 production by approximately 3-fold. Rosiglitazone treatment (1, 5, and 10 micromol/L) also led to a dose-dependent increase (approximately 1.2-1.5-fold) in SERCA2 promoter activity. Comparable levels of cardiac SERCA overexpression have been associated with physiologically relevant and compensatory effects in vivo. These data link thiazolidinedione-induced improvement in cardiac myocyte function to an upregulation of SERCA2 gene expression. Since NF-kappaB-dependent pathways, including the upregulation of IL-6 secretion, were shown to protect neonatal rat ventricular myocytes from apoptosis upon TNFalpha stimulation, additional experiments were designed to determine whether rosiglitazone enhances TNFalpha induced NF-kappaB-dependent transcription and IL-6 secretion. Because the endotoxin stress response in ventricular myocytes involves the upregulation of TNFalpha, and the activation of NF-kappaB, the effects of rosiglitazone on lipopolysaccharide-induced NF-kappaB-dependent transcription were also investigated. Treatment of neonatal rat ventricular myocytes with 10 micromol/L rosiglitazone enhanced TNF-alpha- and lipopolysaccharide-induced NF-kappaB dependent transcription by approximately 1.8- and approximately 1.4-fold respectively, and TNF-alpha-induced IL-6 secretion by n1.5-fold. Rosiglitazone had no significant effects on basal levels of NF-kappaB-dependent transcription and IL-6 secretion. Thus, cardioprotective effects of rosiglitazone may be partly mediated by NF-kappaB. PMID- 15665515 TI - Insulin receptor (IR) and glucose transporter 2 (GLUT2) proteins form a complex on the rat hepatocyte membrane. AB - The hepatic glucose transporter, GLUT2, facilitates bidirectional glucose transport across the hepatocyte plasma membrane under insulin regulation. We studied the interactions of IR and GLUT2 proteins to determine whether they are physically coupled in a receptor-transporter complex. By comparing endosome and plasma membrane IR and GLUT2 ratios before and after feeding, it was determined that IR and GLUT2 are internalized in a fixed ratio. When solubilized hepatocytes were immunoprecipitated with antibodies against either IR or GLUT2, both proteins co-precipitated. The association of IR and GLUT2 was further assessed by confocal microscopy. Sections of fed liver were incubated with fluorescein-tagged anti GLUT2 or Texas Red-tagged anti-IR. Colocalization was observed both at the plasma membrane and in the cytosol. Fluorescence-resonance energy transfer studies further confirmed this association. We conclude that IR and GLUT2 form a receptor transporter complex in hepatocytes, which forms a mechanism of insulin-mediated hepatic glucose regulation. PMID- 15665518 TI - Atrial natriuretic peptide effects on intracellular pH changes and ROS production in HEPG2 cells: role of p38 MAPK and phospholipase D. AB - AIMS: The present study was performed to evaluate Atrial Natriuretic Peptide (ANP) effects on intracellular pH, phospholipase D and ROS production and the possible relationship among them in HepG2 cells. Cancer extracellular microenvironment is more acidic than normal tissues and the activation of NHE-1, the only system able to regulate pHi homeostasis in this condition, can represent an important event in cell proliferation and malignant transformation. METHODS: The ANP effects on pHi were evaluated by fluorescence spectrometry. The effects on p38 MAPK and ROS production were evaluated by immunoblots and analysis of DCF DA fluorescence, respectively. RT-PCR analysis and Western blotting were used to determine the ANP effect on mRNA NHE-1 expression and protein levels. PLD catalyzed conversion of phosphatidylcholine to phosphatydilethanol (PetOH), in the presence of ethanol, was monitored by thin layer chromatography. RESULTS: A significant pHi decrease was observed in ANP-treated HepG2 cells and this effect was paralleled by the enhancement of PLD activity and ROS production. The ANP effect on pHi was coupled to an increased p38 MAPK phosphorylation and a down regulation of mRNA NHE-1 expression and protein levels. Moreover, the relationship between PLD and ROS production was demonstrated by calphostin-c, a potent inhibitor of PLD. At the same time, all assessed ANP-effects were mediated by NPR-C receptors. CONCLUSION: Our results indicate that ANP recruits a signal pathway associated with p38 MAPK, NHE-1 and PLD responsible for ROS production, suggesting a possible role for ANP as novel modulator of ROS generation in HepG2 cells. PMID- 15665516 TI - Regulation of choline deficiency apoptosis by epidermal growth factor in CWSV-1 rat hepatocytes. AB - Previous studies show that acute choline deficiency (CD) triggers apoptosis in cultured rat hepatocytes (CWSV-1 cells). We demonstrate that prolonged EGF stimulation (10 ng/mL x 48 hrs) restores cell proliferation, as assessed by BrdU labeling, and protects cells from CD-induced apoptosis, as assessed by TUNEL labeling and cleavage of poly(ADP-ribose) polymerase. However, EGF rescue was not accompanied by restoration of depleted intracellular concentrations of choline, glycerphosphocholine, phosphocholine, or phosphatidylcholine. In contrast, we show that EGF stimulation blocks apoptosis by restoring mitochondrial membrane potential (Delta Psi(m)), as determined using the potential-sensitive dye chloromethyl-X-rosamine, and by preventing the release and nuclear localization of cytochrome c. We investigated whether EGF rescue involves EGF receptor phosphorylation and activation of the down-stream cell survival factor Akt. Compared to cells in control medium (CT, 70 micromol choline x 48 hrs), cells in CD medium (5 micromol choline) were less sensitive to EGF-induced (0-300 ng/mL x 5 min) receptor tyrosine phosphorylation. Compared to cells in CT medium, cells in CD medium treated with EGF (10 ng/mL x 5 min) exhibited higher levels of phosphatidylinositol 3-kinase (PI3K)-dependent phosphorylation of AktSer473. Inactivation of PI3K was sufficient to block EGF-stimulated activation of Akt, restoration of mitochondrial Delta Psi(m), and prevention of cytochrome c release. These studies indicate that stimulation with EGF activates a cell survival response against CD-apoptosis by restoring mitochondrial membrane potential and preventing cytochrome c release and nuclear translocation which are mediated by activation of Akt in hepatocytes. PMID- 15665517 TI - Thyroid status affects rat liver regeneration after partial hepatectomy by regulating cell cycle and apoptosis. AB - In rats, various growth factors and hormones, as well as partial hepatectomy (PH) are able to trigger the proliferative response of hepatocytes. Although recent evidence highlights the important role of thyroid hormones and thyroid status in regulating the growth of liver cells in vitro and in vivo models, the mechanism involved in the pro-proliferative effects of thyroid hormones is still unclear. Here we have investigated how in rats made hypo- and hyperthyroid after prolonged treatment respectively with propylthiouracil (PTU) and triiodothyronine (T3), the thyroid status affects liver regeneration after PH by regulating cell cycle and apoptosis proteins. Our results show that both in control and partially hepatectomized animals hyperthyroidism increases the cyclin D1, E and A levels and the activity of cyclin-cdk complexes, and decreases the levels of cdk inhibitors such as p16 and p27. On the contrary hypothyroidism induces a down regulation of the activity of cyclin cdk complexes decreasing cyclin levels. Thyroid hormones control also p53 and p73, two proteins involved in apoptosis and growth arrest which are induced by PH. In particular, hypothyroidism increases and T3 treatment decreases p73 levels. The analysis of the phosphorylated forms of p42/44 and p38 MAPK revealed that they are induced during hepatic regeneration in euthyroid and hyperthyroid rats whereas they are negatively regulated in hypothyroid rats. In conclusion our data demonstrate that thyroid status can affects liver regeneration, altering the expression and the activity of the proteins involved in the control of cell cycle and growth arrest. PMID- 15665519 TI - Enzymatic oxidation of vanillin, isovanillin and protocatechuic aldehyde with freshly prepared Guinea pig liver slices. AB - BACKGROUND/AIMS: The oxidation of xenobiotic-derived aromatic aldehydes with freshly prepared liver slices has not been previously reported. The present investigation compares the relative contribution of aldehyde oxidase, xanthine oxidase and aldehyde dehydrogenase activities in the oxidation of vanillin, isovanillin and protocatechuic aldehyde with freshly prepared liver slices. METHODS: Vanillin, isovanillin or protocatechuic aldehyde was incubated with liver slices in the presence/absence of specific inhibitors of each enzyme, followed by HPLC. RESULTS: Vanillin was rapidly converted to vanillic acid. Vanillic acid formation was completely inhibited by isovanillin (aldehyde oxidase inhibitor), whereas disulfiram (aldehyde dehydrogenase inhibitor) inhibited acid formation by 16% and allopurinol (xanthine oxidase inhibitor) had no effect. Isovanillin was rapidly converted to isovanillic acid. The formation of isovanillic acid was not altered by allopurinol, but considerably inhibited by disulfiram. Protocatechuic aldehyde was converted to protocatechuic acid at a lower rate than that of vanillin or isovanillin. Allopurinol only slightly inhibited protocatechuic aldehyde oxidation, isovanillin had little effect, whereas disulfiram inhibited protocatechuic acid formation by 50%. CONCLUSIONS: In freshly prepared liver slices, vanillin is rapidly oxidized by aldehyde oxidase with little contribution from xanthine oxidase or aldehyde dehydrogenase. Isovanillin is not a substrate for aldehyde oxidase and therefore it is metabolized to isovanillic acid predominantly by aldehyde dehydrogenase. All three enzymes contribute to the oxidation of protocatechuic aldehyde to its acid. PMID- 15665520 TI - VEGF production by primary human renal proximal tubular cells: requirement of HIF 1, PI3-kinase and MAPKK-1 signaling. AB - Renal proximal tubular epithelial cells (PTEC) respond to hypoxia exposure or interleukin-1beta (IL-1beta) treatment with increased vascular endothelial growth factor (VEGF) production. With respect to O2 deprivation, the hypoxia-inducible factor 1alpha/ beta (HIF-1) is the most important transcription factor driving VEGF mRNA expression. HIF-1 is also activated by IL-1beta and may thus be involved in the stimulation of VEGF production by this cytokine. However, the molecular mechanisms of HIF-1 dependent VEGF synthesis are poorly understood. Herein, human PTEC in primary culture were challenged by hypoxic incubation and/or IL-1beta treatment in absence or presence of specific phosphatidylinositol 3-kinase (PI3K) or mitogen activated protein kinase kinase-1 (MAPKK-1) inhibitors for assay of VEGF protein, VEGF mRNA and detection of HIF-1alpha by Western Blotting, EMSA and fluorescence microscopy. In addition, the activities of PI3K and MAPKK-1 were studied following hypoxia and IL-1beta treatment of the cultures. The study shows that PI3K but not MAPKK-1 inhibition resulted in the loss of hypoxic and IL-1beta induced HIF-1alpha accumulation, whereas VEGF synthesis was reduced by either intervention. Thus, PI3K signaling is required for HIF-1alpha accumulation and VEGF synthesis, whereas MAPKK-1 signaling is required for VEGF synthesis only. Furthermore, hypoxia alone was sufficient to activate PI3K in PTEC in contrast to MAPKK-1, whose activity was lowered in hypoxia. PMID- 15665521 TI - Down-regulation of calpain 9 is linked to hypertensive heart and kidney disease. AB - Calpains are a family of 14 intracellular calcium-dependent proteases, which have been implicated in cardiovascular diseases. We aimed to analyze specifically the expressional regulation of the different calpain isoforms in hypertensive target organ damage. Using real-time PCR, we found calpain 6 and 9 down-regulated by more than 50% and the endogenous calpain inhibitor calpastatin up-regulated by 225%, respectively, in the hearts of Dahl salt-sensitive rats on a high salt (4% NaCl) compared to normal salt diet. On the protein level, calpain 9 but not calpastatin was regulated in the hypertensive target organs heart and kidney. Moreover, the myocardial expression of calpain 9 protein was inversely linked to left ventricular mass (r= -0.93, p<0.01), and renal expression of calpain 9 protein correlated inversely with albuminuria (r= -0.82, p<0.05). In the aorta, there was no regulation of calpain 9 on the protein level. We conclude that differential regulation of calpain 9 may play a role in hypertensive target organ damage. PMID- 15665522 TI - Adrenomedullin impairs the profibrotic effects of transforming growth factor beta1 through recruiting Smad6 protein in human renal tubular cells. AB - Adrenomedullin (AM) was originally identified as a vasodilator peptide, and has recently been shown to be an antiproliferative factor in renal mesangial cells, suggesting that adrenomedullin may impair the progression of glomerulosclerosis. This study was to investigate the effect of adrenomedullin on transforming growth factor-beta1 (TGF-beta1)-stimulated cell growth, synthesis of extracellular matrix (ECM) components and the related molecular mechanism in a human tubular epithelial cell line HK-2. TGF-beta1 inhibited cell proliferation induced by fetal bovine serum, but neither AM itself affectted cell proliferation, nor did AM influence TGF-beta1-caused cell growth arrest. However, AM beginning at 10(-8) M alleviated the action of TGF-beta1-stimulated cellular collagen synthesis and secretion of fibronectin into cell culture supernatant. Activation of Smad proteins is known to be the key signaling pathway of the profibrotic effect of TGF-beta1, AM at 10(-8) M exerted no effect on TGF-beta1-induced Smad2 phosphorylation, but prevented the suppression of the inhibitory Smad6 protein by TGF-beta1 and restored Smad2-Samd6 complex formation. Our results suggest that AM can attenuate TGF-beta1-mediated renal tubulointerstitial ECM turnover via an antagonistic mechanism of inhibitory Smad in TGF-beta1-elicited signaling. PMID- 15665523 TI - The nephrotoxin ochratoxin A induces key parameters of chronic interstitial nephropathy in renal proximal tubular cells. AB - Ochratoxin A (OTA) is a nephrotoxic and cancerogenic mycotoxin. There is epidemiological evidence that OTA exposition leads to cortical interstitial nephropathies in humans. However, virtually no data are available investigating the effect of OTA on renal cortical cells with respect to induction of nephropathy. Thus, we investigated whether OTA is able to induce changes of cellular properties potentially leading to interstitial nephropathy, using proximal tubular cell lines (OK, NRK-52E). OTA decreased cell number and cell protein time and dose dependently. Accordingly we investigated the effect of 100 nM or 1000 nM OTA. The decline of cell number after OTA exposure is due to necrosis and apoptosis, as measured by LDH release or DNA ladder formation and caspase-3 activation, respectively. OTA incubation of proximal tubular cells also resulted in a loss of epithelial tightness as determined by diffusion of FITC labeled inulin. Inflammation, fibrosis and epithelial-to-mesenchymal transition are described in chronic interstitial renal disease. Therefore, we also investigated the effect of OTA on NFkappaB activity, collagen secretion and generation of alpha smooth muscle actin. OTA alone was sufficient to induce the latter parameters in proximal tubular cells. Finally, OTA is a nephrotoxcic substance and elevated activity of mitogen activated protein kinases (MAPK) is described in nephropathies. As we investigated the effect of OTA on activity of ERK, JNK and p38 by ELISA, we found that OTA activates the MAPK measured dose dependently. In summary, OTA induced phenomena typical for chronic interstitial nephropathy, like loss of cells and epithelial tightness, necrosis and apoptosis as well as markers of inflammation, fibrosis and epithelial-to-mesenchymal transition in proximal tubular cells. Thus, we could show for the first time that OTA is able to induce key parameters of nephropathy in proximal tubular cells in culture. Moreover OTA interacts with MAPK and thus may exert its specific toxic actions. PMID- 15665524 TI - Crosstalk between mesangial and endothelial cells: angiotensin II down-regulates endothelin-converting enzyme 1. AB - OBJECTIVE: Since mesangial and endothelial cells interact in the kidney, the present experiments were designed to analyze the ability of human mesangial cells (HMC) to modulate endothelin-1 (ET-1) synthesis by human umbilical vein endothelial cells (HuVEC). METHODS AND RESULTS: The supernatants of HuVEC/HMC contained significantly lower amounts of ET-1 than those of HuVEC alone. This effect was not due to a decreased prepro-ET-1 mRNA expression and was only partially the consequence of HMC-dependent ET-1 degradation. Therefore, we tested the influence of the coculture on endothelin-converting enzyme-1 (ECE-1), and found a significant reduction of its mRNA and protein levels as well as a decreased activity in HuVEC/HMC as compared to HuVEC alone. Using a pharmacological blockade approach (sulotrobam, BN52021, losartan or catalase), losartan was shown to completely abolish down-regulation of ECE-1 observed in HuVEC/HMC. Angiotensin II (AII) induced a dose and time-dependent inhibition of ECE-1 expression in HuVEC. CONCLUSIONS: These results support the importance of cross-talk among different cell types in the regulation of vascular or renal function. ET-1, and particularly ECE-1, might constitute a target in this regulation. In addition, locally synthesized AII could be one of the mediators involved in the down-regulation of ECE-1. PMID- 15665525 TI - Ca2+ induced surfactant secretion in alveolar type II cultures isolated from the H-2Kb-tsA58 transgenic mouse. AB - BACKGROUND/AIMS: There is a need for the development of transgenic mice to elucidate molecular mechanisms in surfactant secretion. However at present very little is known about the regulation of surfactant exocytosis in murine alveolar type II (AT II) cells. METHODS: We brought AT II cells isolated from the Immorto mouse into culture at 33 degrees C, in the presence of interferon, to generate immortal mouse AT II cells (iMAT II). Surfactant secretion was measured using real-time fluorescence imaging. RESULTS: iMAT II cells stained with lysotracker green (LTG), a dye specific for lamellar body related vesicles in rat AT II cells. Expression of densely layered structures, characteristic of LBs, was confirmed by transmission electron microscopy. Flash photolysis of caged Ca2+, which specifically elevates intracellular Ca2+ concentration ([Ca2+]i), resulted in LB fusion to the plasma membrane, as analysed using the lipid staining dye FM 1-43. Purinergic stimulation with ATP (10 microM), also resulted in a rise in [Ca2+]i (measured by fura-2), which was followed by LB fusion. CONCLUSIONS: iMATII cells maintain the expression of LBs over several passages. Surfactant secretion in these cells is regulated by [Ca2+]i, and exhibits similar characteristics to that of rat AT II cells. These cells will be beneficial in studying the impact of genetic modifications on regulated surfactant secretion. PMID- 15665526 TI - Presentation of ICAM-1 protein at the cell surface of oral keratinocytes in the presence of adrenomedullin and corticotrophin. AB - Increasing evidence suggests that adrenomedullin (AM) and corticotrophin (ACTH) are immunomodulatory. Intercellular adhesion molecule-1 (ICAM-1) plays an important role in the recruitment of leukocytes not only from peripheral blood into inflamed tissues but also into epithelia. We have investigated the effects of AM and ACTH on the expression of ICAM-1 by human oral keratinocytes. The human oral keratinocyte cell line H357 was incubated with either AM or ACTH for up to 8 hrs and ICAM-1 expression was measured by cell surface ELISA. ICAM-1 was up regulated by both peptides and this was attenuated by the adenylyl cyclase inhibitor SQ22,536 and the NF-kappaB inhibitor SN-50. H357 cells constitutively express ICAM-1 mRNA and expression of this gene was significantly modulated by AM and ACTH. Furthermore AM caused translocation of NF-kappaB from the cytoplasm to the nucleus. This is the first report describing up regulation of ICAM-1 in oral keratinocytes by AM and ACTH and the results suggest both cAMP and NF-kappaB may play a role. These results further suggest both peptides may have an immunostimulatory role in oral muocsa and skin. PMID- 15665528 TI - The temperature-sensitive ion channel TRPV2 is endogenously expressed and functional in the primary sensory cell line F-11. AB - In sensory neurons heat is transduced by a subfamily of TRP channels sharing sequence homology with the capsaicin-sensitive vanilloid receptor subtype 1 (TRPV1), but differing in their thermal response thresholds. To identify a neuronal cell line endogenously expressing noxious heat-transducing ion channels, we examined F-11 cells, a hybridoma derived from rat dorsal root ganglia and mouse neuroblastoma. Using RT-PCR, transcripts homologous to TRPV2 and TRPV4, but not to TRPV1 or TRPV3, were found. We isolated a full-length cDNA of 2.4 kb coding for a 757-amino acid protein corresponding to mouse TRPV2, which was further characterized by immunocytochemistry and electrophysiology. Using the whole-cell patch-clamp technique, we observed a heat-evoked increase in outward and inward currents with a threshold of 51.6 +/- 0.2 degrees C. The current voltage relationship stimulated by a temperature of 52 degrees C was characterized by an outward rectification with a reversal potential close to -10 mV. Heat-evoked currents could be inhibited by ruthenium red. There was no activation by stimulation with capsaicin or 2-aminoethoxydiphenyl borate. Our results indicate that F-11 cells express functional noxious heat-sensitive TRPV2 channels. Thus, we propose that F-11 cells represent a valuable in vitro model to characterize the properties of TRPV2 in a native neuronal environment. PMID- 15665527 TI - Requirement of PDZ domains for the stimulation of the epithelial Ca2+ channel TRPV5 by the NHE regulating factor NHERF2 and the serum and glucocorticoid inducible kinase SGK1. AB - Renal calcium reabsorption involves the epithelial calcium channel ECaC1 (TRPV5) which is tightly regulated by 1,25(OH)2D3. As shown recently, TRPV5 is activated by the serum and glucocorticoid inducible kinase SGK1, a kinase transcriptionally upregulated by 1,25(OH)2D3. This stimulatory effect is due to enhanced TRPV5 abundance in the plasma membrane and requires the presence of the scaffold protein NHERF2 (sodium hydrogen exchanger regulating factor 2). The present study aims to define the molecular requirements for the interaction of TRPV5 with SGK1 and NHERF2. Pull-down experiments and overlay assays revealed that the TRPV5 C tail interacts in a Ca2+-independent manner with NHERF2. Deletion of the second but not of the first PDZ domain in NHERF2 abrogates the stimulating effect of SGK1/NHERF2 on TRPV5 protein abundance in the plasma membrane as quantified by chemiluminescence and electrophysiology. Thus, the second PDZ domain in NHERF2 is required for stabilization at or TRPV5 targeting to the plasma membrane. The experiments demonstrate the significance of SGK1 and NHERF2 as TRPV5 modulators which are likely to participate in the regulation of calcium homeostasis by 1,25(OH)2D3. PMID- 15665529 TI - Safety of cardiac rehabilitation in a medically supervised, community-based program. AB - The time to occurrence of cardiovascular complications after the beginning of an exercise rehabilitation program is variable. It is not clear whether such complications are related to the duration in the program. The aim of the present study was to assess the timing of cardiovascular events occurring during the activity and the long-term safety of a medically supervised cardiac rehabilitation program performed in the community, in a large cohort. We retrospectively evaluated 3,511 patients with a history of myocardial infarction, coronary artery bypass grafting and risk factors for coronary artery disease, participating in exercise training, for 69 months. The total number of patient hours was 338,688 with an event rate of 1/58,902 patient-hours/year (0.02%). Non fatal events occurred in 11 patients and fatal cardiovascular events in 2 patients; 1 was successfully resuscitated. Most of the non-fatal events (62%) occurred during the first 4 weeks from the beginning of the exercise program. One third of the patients who experienced cardiovascular events, resumed the exercise program with no further complications. Medically supervised cardiac rehabilitation program is accompanied by a very low incidence of cardiovascular events. Nevertheless, special caution should be undertaken during the first sessions of the program. PMID- 15665530 TI - Electrocardiographic features of hypothermia. PMID- 15665532 TI - Detection of Chlamydia pneumoniae DNA in the coronary arteries and bypass in three patients with diffuse coronary artery disease. PMID- 15665531 TI - Serum B-Type natriuretic peptide in STEMI patients treated with PCI. PMID- 15665533 TI - Self-expanding coronary stent (Radius) implantation with cutting balloon angioplasty. AB - To evaluate the effect of cutting balloon (CB) angioplasty on the outcome of self expanding coronary stent implantation, we compared the results of Radius stent implantation using two predilation strategies. Seventy de novo lesions in 70 patients (60 men) were included in the study. Stenting was performed after conventional angioplasty (n = 38, group 1) or CB angioplasty (n = 32, group 2). In-stent restenosis (ISR) rates of each group were 39 and 10% at 6-month follow up (p = 0.02). Major adverse cardiac events occurred in 20 and 11%, respectively (p = 0.25). Multivariate analysis revealed that ISR was predicted only by lesion length > or = 20 mm. In conclusion, CB angioplasty did not reduce ISR after Radius stent implantation, when other independent risk factors, such as lesion length, were taken into consideration. PMID- 15665534 TI - ST-segment elevation pattern and myocardial injury induced by acute pancreatitis. PMID- 15665535 TI - Metabolic syndrome in a screened worksite sample: prevalence and predictors. AB - Relatively limited contemporary information is available about the magnitude of, and factors associated with, the metabolic syndrome in adult men and women. The purpose of our observational study was to describe the prevalence and predictors of the metabolic syndrome in a sample of employed adults attending a worksite cardiovascular screening program. The study sample consisted of 871 men and women between the ages of 21 and 77 years from 6 locations of the parent company. These individuals attended an employer-sponsored cardiovascular screening and wellness program during 2003. A standardized questionnaire was administered to all study participants and a number of different coronary risk factors were measured. Approximately 27% of the study sample was classified as having the metabolic syndrome. Men, persons with a history of hypertension, heart disease, or stroke, sedentary individuals, and those with an increased heart rate and higher levels of C-reactive protein were associated with presence of the metabolic syndrome. A relatively similar risk factor profile was noted in persons without a self reported history of prior cardiovascular disease. The results of our cross sectional observational study suggest that the prevalence of the metabolic syndrome is considerable. A number of demographic, comorbid, and other factors are associated with this syndrome. Increased attention to the metabolic syndrome, and modification of predisposing factors, remains of considerable public health and clinical importance. PMID- 15665536 TI - Mitochondrial dysfunction as an initiating event in atherogenesis: a plausible hypothesis. AB - It is now widely accepted that oxidant stress and the ensuing endothelial dysfunction play a key role in the pathogenesis of atherosclerosis and cardiovascular diseases. The mitochondrial respiratory chain is the major source of reactive oxygen species as byproducts of normal cell respiration. Mitochondria may also be important targets for reactive oxygen species, which may damage mitochondrial lipids, enzymes and DNA with following mitochondrial dysfunction. Free cholesterol, oxidized low-density lipoprotein and glycated high-density lipoprotein are further possible causes of mitochondrial dysfunction and/or apoptosis. Moreover, in patients with mitochondrial diseases, vascular complications are commonly observed at an early age, often in the absence of traditional risk factors for atherosclerosis. We propose that mitochondrial dysfunction, besides endothelial dysfunction, represents an important early step in the chain of events leading to atherosclerotic disease. PMID- 15665537 TI - Shunt obstruction after an air travel in an adult patient with cyanotic congenital heart disease. PMID- 15665538 TI - Delayed diagnosis and lower resection rate of adenocarcinoma of the distal duodenum. AB - BACKGROUND: Carcinoma of the duodenum is a rare disease that can present with varied symptoms and is often misdiagnosed. Prolonged survival following resection of the primary tumour is possible, whilst irresectable disease has a very poor prognosis. The factors determining resectability of the primary tumour have not been addressed. AIMS: We reviewed 45 consecutive cases of duodenal carcinoma to investigate factors which influence the operative outcome of patients with this condition. PATIENTS AND METHODS: Details of symptoms, diagnoses, surgical procedures and pathology were retrieved from patient records. There were 29 male and 16 female patients aged 24-79 years (median = 64 years). RESULTS: The frequency of tumours in the proximal and distal duodenum was 27 and 18. Failure to diagnose the tumours at endoscopy occurred in 10 of 27 tumours of the proximal duodenum and 15 of 18 tumours of the distal duodenum. The duration of symptoms prior to diagnosis was correspondingly longer for tumours in the distal duodenum (20 weeks) than the proximal duodenum (12 weeks). Of 27 patients with tumours in the proximal duodenum, 18 underwent a potentially curative resection, whereas only 6 of the 18 tumours in the distal duodenum were resectable with curative intent. The reasons for irresectability of lesions in the distal duodenum included malignant lymphadenopathy affecting the small bowel mesentery in ten cases, which was not noted in any patient with adenocarcinoma of the proximal duodenum. CONCLUSIONS: Our results suggest that lesions of the distal duodenum are inadequately investigated by endoscopy, and that distal duodenal tumours are less curable by resectional surgery due to invasion of the small bowel mesentery. PMID- 15665539 TI - A prospective audit of the complications of loop ileostomy construction and takedown. AB - AIM: A prospective review of the complications of ileostomy construction and takedown. MATERIALS AND METHODS: One hundred twenty-seven consecutive patients undergoing construction of a loop ileostomy were included in a prospective nonrandomized computer database. Complications of the loop ileostomy were assessed prior to and after closure. Three closure techniques were performed [enterotomy suture (25.7%), resection and hand sewn (31.2%) or stapled anastomosis (43.1%)] and compared. RESULTS: One hundred twenty-seven (73 male, 54 female) patients, mean age 54 years were included from 1992 to 2002. Seventy-two patients underwent anterior resection for low rectal carcinoma, 30 an ileoanal pouch for ulcerative colitis and 25 for miscellaneous conditions. Fifty-nine pre takedown complications occurred in 50 (39.4%) patients. The most common were dermatitis (12.6%) and erythema (7.1%). The most severe were dehydration in 1 patient and stomal prolapse in 4 patients. Closure was associated with a complication rate of 33.1% and a mortality rate of 0.9%. Wound infection occurred in 18.3% and small bowel obstruction in 4.6%. Anastomotic leak requiring reanastomosis occurred in 2.8% and enterocutaneous fistula treated conservatively in 5.5%. There were no statistically significant differences in morbidity between closure techniques (p = 0.892). There were no statistically significant differences in complications (p = 0.516) between patients with ulcerative colitis and those with neoplasia (39.29% vs. 32.2%). CONCLUSIONS: Loop ileostomy construction and takedown is associated with considerable morbidity, mostly minor. No differences exist between technique used for closure or the baseline pathology of the patient. PMID- 15665540 TI - David Erlik (1909-1995). A founder of surgery in modern Israel. PMID- 15665541 TI - Sclerosant therapy as first-line treatment for solitary liver cysts. AB - AIM: The aim was to determine the outcome from percutaneous sclerosing treatment of solitary non-parasitic hepatic cysts. METHODS: The results of treatment of patients with symptomatic solitary non-parasitic hepatic cysts treated between 1995 and 2000 were reviewed. RESULTS: There were 23 women and one man with a median (range) age of 59 (34-79) years. The median (range) diameter of the cysts was 10 (5-24) cm. Five patients were treated by laparoscopic fenestration ab initio as they also required a cholecystectomy because of gallstones. The remaining 19 patients underwent percutaneous sclerotherapy. In one just aspiration was successful without further treatment. In six contrast leaked from the cyst and five of these had laparoscopic fenestration. Twelve patients had sclerosant treatment with good results at a median (range) follow-up of 35 (6-60) months in 10 patients. Good results were also obtained in 10 of the 12 patients who had fenestration. PMID- 15665542 TI - Rectal cancer with paraneoplastic nephropathy: association of vascular endothelial growth factor. AB - A patient with advanced rectal cancer was complicated by progressing proteinuria and hypoproteinemia. Low anterior resection was a procedure of choice. A surgical specimen obtained by intraoperative renal biopsy showed the findings of minimal change nephrotic syndrome. After surgery, nephropathy remitted promptly and completely. Her pre/postoperative serum level of vascular endothelial growth factor was 1,880/52.3 pg/ml, suggesting its elevation was associated with the nephropathy. Immunohistochemistry revealed strongly expressed tumor vascular endothelial cell growth factor. Minimal change nephrotic syndrome is a rare type of paraneoplastic nephropathy, and successful remission may require therapeutic resection of the underlying tumor, or administration of a vascular endothelial growth factor antagonist if the tumor is unresectable. PMID- 15665543 TI - Formation and regrowth of intra-abdominal adhesions after adhesiolysis: the paradox of surgical adhesion-reduction strategies. PMID- 15665545 TI - Atrial natriuretic peptide preserves endothelial function during intimal hyperplasia. AB - BACKGROUND: Atrial and C-type natriuretic peptides (ANP and CNP), acting through different receptors, have antiproliferative effects in vitro. Beneficial effects of CNP in vivo on early atherosclerosis have been described, but it is not known if ANP is antiproliferative in vivo. In the present study, the effects of chronic in vivo ANP were tested and compared with CNP on endothelial dysfunction and intimal thickening caused by peri-arterial collars. METHODS: Non-occlusive collars were placed bilaterally around the common carotid arteries of rabbits. One collar was filled with saline vehicle. The contralateral collar was filled with ANP or CNP (1 or 10 microM, n = 5-7) with slow replacement of peptide via mini-pump (1 or 10 fmol/h). RESULTS: After 7 days, endothelium-dependent vasorelaxation in saline-collared arteries was 33 +/- 3% of maximum [averaged over 0.03-1 muM acetylcholine (ACh)] compared to 64 +/- 2% in normal (uncollared) arteries (p < 0.05, n = 23). In vivo ANP restored the ACh relaxation to normal (e.g., 57 +/- 6%, 1 microM ANP), similar to effects seen with CNP in vivo. Endothelium-independent vasorelaxation of collared-vessels was not altered by either peptide. Intimal hyperplasia induced by the collars was not prevented by peri-arterial natriuretic peptides. In additional rabbits (n = 6), CNP (100 pmol/h) given directly into the lumen of collared carotid arteries for 7 days reduced neointima formation by 16 +/- 5% (p < 0.05), whereas ANP given intraluminally (100 pmol/h; n = 6) did not. CONCLUSIONS: The more potent actions of CNP on vascular smooth muscle cell migration and proliferation (established in vitro) may explain differences between the peptides on intimal hyperplasia in vivo. The major hallmark of atherosclerosis and restenosis, endothelial dysfunction, was prevented by chronic, peri-arterial administration of ANP or CNP. PMID- 15665546 TI - A new ex vivo model to study venous angiogenesis and arterio-venous anastomosis formation. AB - Explants of rat inferior vena cava embedded in collagen gel and cultured under serum-free conditions produced microvascular outgrowths composed of endothelial cells and pericytes. Exogenous vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) dose-dependently stimulated angiogenesis and induced the formation of complex networks of highly branched microvessels. VEGF and the VEGF/bFGF combination also promoted pericyte recruitment. Medium conditioned by untreated vena cava cultures contained endogenous VEGF, and a blocking antibody against VEGF significantly reduced the spontaneous angiogenic response of the explants. Vena cava explants exhibited a greater capacity to form neovessels than aortic rings when tested in parallel cultures from the same animal. When compared with aorta-derived microvessels, neovessels of vena cava origin were longer and had fewer pericytes. Vena cava-aorta cocultures produced extensive anastomosing networks of microvessels, which were primarily contributed by the venous explants. Because of its florid angiogenesis and exquisite sensitivity to angiogenic factor stimulation, the vena cava model may provide novel insights into the regulation of the angiogenic process, which typically initiates from the venous side of the vascular bed. Combined with the aortic ring model, this new assay may also enhance our understanding of the mechanisms of anastomosis formation between the arterial and the venous circulations. PMID- 15665547 TI - Structural identification of oxidized acyl-phosphatidylcholines that induce platelet activation. AB - Oxidation of low-density lipoprotein (LDL) generates proinflammatory and prothrombotic mediators that may play a crucial role in cardiovascular and inflammatory diseases. In order to study platelet-activating components of oxidized LDL 1-stearoyl-2-arachidonoyl-sn-glycero-3-phosphocholine, a representative of the major phospholipid species in LDL, the 1-acyl phosphatidylcholines (PC), was oxidized by CuCl(2) and H(2)O(2). After separation by high-performance liquid chromatography, three compounds were detected which induced platelet shape change at low micromolar concentrations. Platelet activation by these compounds was distinct from the pathways stimulated by platelet-activating factor, lyso-phosphatidic acid, lyso-PC and thromboxane A(2), as evidenced by the use of specific receptor antagonists. Further analyses of the oxidized phospholipids by electrospray ionization mass spectrometry structurally identified them as 1-stearoyl-2-azelaoyl-sn-glycero-3-phosphocholine (m/z 694; SAzPC), 1-stearoyl-2-glutaroyl-sn- glycero-3-phosphocholine (m/z 638; SGPC), and 1-stearoyl-2-(5-oxovaleroyl)-sn-glycero-3-phosphocholine (m/z 622; SOVPC). These observations demonstrate that novel 1-acyl-PC which had previously been found to stimulate interaction of monocytes with endothelial cells also induce platelet activation, a central step in acute thrombogenic and atherogenic processes. PMID- 15665548 TI - Decreased whole body endogenous nitric oxide production in patients with primary pulmonary hypertension. AB - Impaired pulmonary release of nitric oxide (NO) is one of the characteristic phenotypic changes of vascular cells in pulmonary hypertension. The aim of this study was to determine nitric oxide synthase (NOS)-dependent whole body NO production in patients with primary pulmonary hypertension. NOS-dependent whole body NO production was assessed by giving an intravenous infusion of L-[(15)N](2) arginine (50 micromol/min for 30 min) and measuring isotopic urinary enrichment of (15)N-nitrite and (15)N-nitrate. Four female patients with no signs of infection were recruited and compared with 6 age-matched control subjects. Mean 12-hour excretion of (15)N-nitrite and (15)N-nitrate in the total urine over 36 h was smaller in patients than in control subjects (57.2 +/- 27.6 vs. 229.1 +/- 65.2 nmol/mmol creatinine, p < 0.01, Mann-Whitney U test, respectively). Neither mean 12-hour excretion of (14)N-nitrite and (14)N-nitrate (51.6 +/- 10.0 vs. 72.4 +/- 10.0 micromol/mmol creatinine, p = 0.3) nor glomerular filtration rates (84.5 +/- 15.8 vs. 129.7 +/- 16.0 ml/min, p = 0.1) were different between patients and control subjects. Our results suggest that either basal NOS-dependent whole body NO production is impaired or excess NO metabolism occurs in patients with primary pulmonary hypertension. PMID- 15665549 TI - Comparison of the effect of pentoxifylline and captopril on proteinuria in patients with type 2 diabetes mellitus. AB - OBJECTIVES: To compare the relative efficacy of pentoxifylline (PTX) and angiotensin-converting enzyme (ACE) inhibitor, captopril in the treatment of proteinuria of type 2 diabetic patients. DESIGN: A randomized open, crossover, clinical trial conducted from October 2000 to March 2001. SETTING AND PARTICIPANTS: 39 patients with type 2 diabetes age 34-75 years were randomly allocated to the two treatment groups. The first group received PTX (400 mg three times a day) orally for a total of 2 months. The second group received captopril (25 mg three times a day) for 2 months. Response to treatment was assessed at 1, 2, 4, and 8 weeks after start of therapy. RESULTS: Captopril appeared to be equivalent in efficacy and safety to PTX. A significant decrease in proteinuria occurred in both groups. Of the 20 patients treated with PTX, the mean (SD) of 24 h urinary protein decreased from 1.4 (0.7) to 1.0 (0.7) g/24 h (p < 0.05). Correspondingly, in the 19 patients treated with captopril, the mean (SD) of 24 h urinary protein decreased from 1.3 (0.7) to 0.8 (0.7) g/24 h (p < 0.01). CONCLUSION: This study demonstrates that treatment with PTX and captopril both significantly reduce overt proteinuria in patients with type 2 diabetes. This effect of ACE inhibition has previously been shown to slow progression to renal failure and we postulate that treatment with PTX may have a similar benefit. PMID- 15665550 TI - Serum cross-linked N-terminal telopeptide of type I collagen for evaluation of renal osteodystrophy in hemodialysis patients. AB - BACKGROUND: Useful markers of bone resorption are needed for hemodialysis patients with renal osteodystrophy. This study investigated the use of a new immunoassay for cross-linked N-terminal telopeptide of type 1 collagen to assess bone changes in hemodialysis patients. METHODS: Radial bone mineral density was examined in 178 hemodialysis patients at baseline and after 12 months. Serum levels of N-terminal telopeptide and other markers were measured. RESULTS: The annual percent change of radial bone mineral density was negatively correlated with the levels of N-terminal telopeptide, intact osteocalcin, tartrate-resistant acid phosphatase, bone-specific alkaline phosphatase, and intact parathyroid hormone. The annual percent change of radial bone mineral density showed a stronger correlation with N-terminal telopeptide levels than with the other markers, except for intact parathyroid hormone. Also, intact parathyroid hormone and N-terminal telopeptide levels showed a stronger correlation than that of either tartrate-resistant acid phosphatase or cross-linked carboxyterminal telopeptide of type 1 collagen with intact parathyroid hormone. CONCLUSION: Serum N-terminal telopeptide may be the most useful bone resorption marker in renal osteodystrophy and its use combined with bone formation markers may improve the management of this condition. PMID- 15665551 TI - Blood pressure control before and after starting dialysis. AB - BACKGROUND/AIM: We have previously reported a study on the adequacy of management of hypertension in patients developing end-stage renal failure (ESRF) over a period of 3 years from a single health district in the UK (n = 107). There were significant shortcomings in all aspects of management of hypertension including blood pressure (BP) control. METHODS: In this report, we have compared BP control in the same cohort of patients before and after starting renal replacement therapy (RRT). RESULTS: BP control improved significantly after the patients were established on dialysis (mean 158/ 87 mm Hg pre-RRT vs. 152/82 mm Hg post-RRT; p < 0.0001), and fewer antihypertensive agents were prescribed to control BP (mean 2.45 vs. 1.74) in this period (p < 0.0001). Moreover, patients on continuous ambulatory peritoneal dialysis (n = 50) had a better systolic BP control compared with the haemodialysis patients (n = 57; p = 0.03). CONCLUSIONS: This study shows significant improvement in BP control in a cohort of patients with ESRF following the start of dialysis. PMID- 15665552 TI - Rituximab induced remission in a patient with Wegener's granulomatosis. AB - BACKGROUND: Wegener's granulomatosis is a form of systemic vasculitis typically involving the kidneys and upper and lower respiratory tract. Treatment employing cyclophosphamide and prednisone has improved prognosis, but relapses and treatment-induced side effects still cause severe morbidity and frequent mortality. There is therefore an urgent need to find new treatment modalities that are efficient and cause few side effects. Chimeric anti-CD20 (rituximab) may be one such treatment, apparently working in part by suppressing anti-PR3 production. PATIENTS AND METHODS: We describe a 26-year-old man with two relapses while on azathioprine and mycophenolate necessitating a high cumulative dose of cyclophosphamide. He was treated with a single standard course of rituximab while continuing steroids and mycophenolate. RESULTS: After 4 months, rituximab led to resolution of pulmonary lesions and also rapidly caused normalization of elevated anti-PR3. He still remains in complete remission 11 months later with a negative anti-PR3 and normal kidney function. There were no side effects to rituximab. CONCLUSION: Rituximab may be effective and seems to be safe in inducing remission in Wegener's granulomatosis. Studies are needed to establish the long-term benefit of this expensive treatment. PMID- 15665553 TI - Interleukin-1beta-induced transdifferentiation of renal proximal tubular cells is mediated by activation of JNK and p38 MAPK. AB - Interleukin (IL)-1beta induces renal tubular epithelial cells to transdifferentiate to myofibroblasts, which express alpha-smooth muscle actin (alpha-SMA). To understand the signal transduction mechanisms involved in transdifferentiation, we examined the roles of mitogen-activated protein kinases (MAPKs) in IL-1beta-stimulated alpha-SMA expression and cell migration in the HK 2 human renal proximal tubular cell line. IL-1beta induced the transdifferentiation of renal proximal tubular cells, which was characterized by upregulated expression of alpha-SMA and increased cell migration. In addition, IL 1beta increased the activity of the three members of the MAPK family, ERK, JNK and p38 MAPK, in these cells. Both SP600125, a specific inhibitor of JNK, and SB203580, a specific inhibitor of p38 MAPK, suppressed the IL-1beta-induced expression of alpha-SMA and cell migration, but these effects were not observed with PD98059, a specific inhibitor of ERK. These results suggest that IL-1beta induced HK-2 cell transdifferentiation is mediated, at least in part, through the activation of the JNK and p38 MAPK signaling pathways. PMID- 15665554 TI - Comparative study on the effects of cyclosporin a in renal cells in culture. AB - BACKGROUND: Although cyclosporin A (CSA) inhibits P-glycoprotein (ABCB1), the relationship between this inhibition and CSA-induced nephrotoxicity is not established. METHODS: Three renal cell lines were used to investigate the effects of CSA in cellular viability and accumulation of rhodamine 123 (Rho123): LLC-PK1, which does not express ABCB1 substantially; MDCK, expressing moderate amounts of this protein, and Ma104 cells, which express high amounts of ABCB1. RESULTS: The viability was significantly reduced in the three cell lines after treatment with CSA concentrations >10 microM. Ma104 was the more resistant and LLC-PK1 the more sensitive. CSA increased Rho123 accumulation in the three cell lines when incubated simultaneously, MDCK presenting the higher increase. However, different results were achieved when the periods of incubation with Rho123 and CSA were disconnected: a post-incubation with CSA was more effective in Ma104 cells, while MDCK and LLC-PK1 showed no difference between pre-, co- and post-incubation with CSA. CONCLUSIONS: Our results suggest that the effects of CSA may be divided into two groups: ABCB1-independent (direct injury), and ABCB1-dependent toxicity, due to modulation of its activity. This could result in increased accumulation of noxious ABCB1 substrates, contributing to CSA-induced nephrotoxicity. Furthermore, the mechanisms of ABCB1 modulation by CSA may be different for different cell lines. PMID- 15665555 TI - Possible effects of hepatocyte growth factor for the prevention of peritoneal fibrosis. AB - OBJECTIVE: Some patients who had carried out long-term continuous ambulatory peritoneal dialysis discontinued the treatment because of progressive peritoneal fibrosis. It has been previously reported that transforming growth factor-beta1 (TGF-beta1) is one of the factors that induces peritoneal fibrosis. Also, hepatocyte growth factor (HGF) plays a role in the prevention of fibrosis and in inhibiting TGF-beta1 production. In this study, we examined the effects of HGF on peritoneal fibrosis by TGF-beta1 induced by high concentrations of D-glucose. DESIGN: We transfected a full-length human HGF cDNA in an expression vector into human peritoneal mesothelial cells (HPMCs) using the calcium phosphate method. Transfected HPMCs were cultured with high concentrations of D-glucose solution and co-cultured with fibroblasts using a transwell system. Cell proliferation was determined using the Tetra Color One method. TGF-beta1 and HGF protein were measured by enzyme-linked immunosorbent assay. RESULTS: In addition to recombinant HGF, the growth inhibition of HPMCs by high concentration D-glucose or TGF-beta1 was significant. By transfecting HGF cDNA into HPMCs, growth inhibition by high concentration D-glucose was completely restored. Furthermore, the production of TGF-beta1 was also significantly decreased. CONCLUSION: These results suggested that exogenous HGF could possibly prevent peritoneal fibrosis. PMID- 15665557 TI - Zinc and cadmium interactions in a renal cell line derived from rabbit proximal tubule. AB - BACKGROUND: The aim of this work was to characterize the relationship between zinc (Zn(2+)) and cadmium (Cd(2+)) and the toxic effects of Cd(2+) in immortalized renal proximal tubule cells RP1. METHODS: An RP1 cell line was developed from primary cultures of microdissected S1 and S2. Uptakes of (65)Zn and (109)Cd and competitive experiments with Cd(2+) and Zn(2+) were performed and kinetic parameters were determined. Oxygen consumption, metallothionein synthesis, and necrotic and apoptotic phenomena were studied. RESULTS: Kinetic parameters indicate that (65)Zn (Km = 71.8 +/- 10.6 microM) and (109)Cd (Km = 23.3 +/- 2.0 microM) were both transported by a saturable carrier-mediated process. Competition between Cd(2+) and Zn(2+) uptake was reciprocal. Cd(2+) induced an increase in necrosis and apoptosis, and a decrease in oxygen consumption, depending on Cd(2+) concentrations. Concomitant addition of Zn(2+) (10 microM) reduced the number of necrotic and apoptotic cells and maintained oxygen consumption at control levels. Cd(2+) alone, or in the presence of Zn(2+), increased metallothionein levels, whereas Zn(2+) alone did not. CONCLUSION: Zn(2+) and Cd(2+) probably share the same transporter in the proximal tubule. Cd(2+) caused necrotic and apoptotic cell death. Cd(2+) toxicity may occur through an effect on the mitochondrial electron transport chain and not on metallothionein synthesis. Zn(2+) protects against the renal cell toxicity of Cd(2+). PMID- 15665556 TI - Kisspeptin activation of gonadotropin releasing hormone neurons and regulation of KiSS-1 mRNA in the male rat. AB - The KiSS-1 gene codes for a family of neuropeptides called kisspeptins which bind to the G-protein-coupled receptor GPR54. To assess the possible effects of kisspeptins on gonadotropin secretion, we injected kisspeptin-52 into the lateral cerebral ventricles of adult male rats and found that kisspeptin-52 increased the serum levels of luteinizing hormone (p < 0.05). To determine whether the kisspeptin-52-induced stimulation of luteinizing hormone secretion was mediated by gonadotropin-releasing hormone (GnRH), we pretreated adult male rats with a GnRH antagonist (acyline), then challenged the animals with intracerebroventricularly administered kisspeptin-52. The GnRH antagonist blocked the kisspeptin-52-induced increase in luteinizing hormone. To examine whether kisspeptins stimulate transcriptional activity in GnRH neurons, we administered kisspeptin-52 intracerebroventricularly and found by immunocytochemistry that 86% of the GnRH neurons coexpressed Fos 2 h after the kisspeptin-52 challenge, whereas fewer than 1% of the GnRH neurons expressed Fos following injection of the vehicle alone (p < 0.001). To assess whether kisspeptins can directly act on GnRH neurons, we used double-label in situ hybridization and found that 77% of the GnRH neurons coexpress GPR54 mRNA. Finally, to determine whether KiSS-1 gene expression is regulated by gonadal hormones, we measured KiSS-1 mRNA levels by single-label in situ hybridization in intact and castrated males and found significantly higher levels in the arcuate nucleus of castrates. These results demonstrate that GnRH neurons are direct targets for regulation by kisspeptins and that KiSS-1 mRNA is regulated by gonadal hormones, suggesting that KiSS-1 neurons play an important role in the feedback regulation of gonadotropin secretion. PMID- 15665558 TI - Intraluminal tumour thrombus of a mixed non-seminomatous germ cell tumour of testis within the inferior vena cava. AB - BACKGROUND: Intracaval tumour thrombus developed per continuitatem from a primary testicular tumour is rare. CASE REPORT: A patient with metastatic mixed non seminomatous germ cell tumour of the testis extending into the inferior vena cava (IVC) is presented. He belonged to the intermediate-risk group according to the IGCCCG (International Germ Cell Cancer Collaborative Group) classification. The 26-year-old man underwent right inguinal orchiectomy. Computed tomography revealed the tumour thrombus as filling defect in the IVC extending nearly to the right renal vein. Duplex sonography detected a partial thrombosis of the IVC. Combination chemotherapy led to regression of pulmonal metastases and the intraluminal tumour thrombus. 5 months later, retroperitoneal lymphadenectomy was performed and the intraluminal thrombus was extracted by cavotomy. The thrombus originated from the ostium of the right testicularis vein in the IVC. Histological examination revealed no vital tumour tissue. CONCLUSION: In patients with testicular cancer information about pathological processes of the IVC is important for therapeutic management. Testicular tumours seldom extend up the IVC. PMID- 15665559 TI - Raf kinase inhibitors in oncology. AB - The importance of the MAP kinase pathway, which includes the kinases Raf, MEK1/2, and ERK1/2, for the proliferation and survival of tumor cells recently increased with the discovery of activating BRAF mutations in human tumors. Therefore, in addition to a role in controlling tumors with Ras mutations and activated growth factor receptors, inhibitors of Raf kinase may harbor therapeutic potential in tumors carrying a BRAF oncogene. A variety of agents have been discovered to interfere with Raf kinase, including antisense oligonucleotides and small molecules. These inhibitors prevent the expression of Raf protein, block Ras/Raf interaction, or obstruct its kinase activity. Raf inhibitors that are currently undergoing clinical evaluation show promising signs of anti-cancer efficacy with a very tolerable safety profile. Clinically most advanced is the Raf inhibitor BAY 43-9006, which recently entered phase III clinical testing. This review addresses the rationale for targeting Raf kinase and the current status of various pharmacological approaches. PMID- 15665560 TI - The efficacy of linear accelerator radiosurgery in the management of patients with Cushing's disease. AB - We identified 35 patients who had undergone stereotactic radiosurgery (SRS) for their biochemically proven Cushing's disease in order to assess the efficacy of SRS with regard to control of hypercortisolism, improvement of clinical features and prevention of tumor progression, and subsequent incidence of hypopituitarism. Seventeen (49%) patients achieved control of their cortisol levels following SRS; the mean time to normalization was 7.5 months (range: 1-33). Four (19%) patients experienced recurrent hypercortisolism at a mean time of 35.5 months following therapy (range: 17-64). Control of tumor progression was achieved in 91% patients. Fourteen (40%) patients demonstrated a new pituitary deficiency following SRS. Our results suggest that cortisol levels are normalized more efficiently and with a lower recurrence rate with SRS than with conventional fractionated external beam radiotherapy (EBT). We have confirmed the near 100% tumor control rate reported with SRS. The percentage of patients developing pituitary insufficiency following SRS is less than that of patients having undergone EBT; however, deficits occurred up to 10 years posttreatment. We advocate the use of SRS as the primary therapeutic modality in those patients who are poor surgical candidates, or as the adjunct treatment to microsurgery in eliminating residual tumor cells or disease that is not easily amenable to resection. PMID- 15665561 TI - [Gastric heterotopia in rectum]. PMID- 15665562 TI - [Indications and limitations of endoscopic mucosal resection in gastric cancer]. AB - Endoscopic mucosal resection (EMR) has come to play an increasingly important role in treatment of early cancer in gastrointestinal tract. Recent advances in EMR are very remarkable. These allow minimally invasive treatment of diseases that would otherwise require major surgery. The most important factors of EMR are accuracy and safety. Further improvement in both staging and resection technologies, as well as safety and short procedure time will ultimately conspire to make this an even more effective tool in the management of early cancer in gastrointestinal tract. EMR must prove to be safe for the majority of patients when performed by competent endoscopist. The new techniques will continue to solve the limitations of endoscopic treatment and its use will also continue to expand increasingly. Also, further studies are required to refine and standardize EMR. As EMR technology becomes more complex, necessitating the use of multiple accessories simultaneously, technical ease may be enhanced by simple adjunct devices. The future of EMR depends on extending its boundaries safely in a controlled setting of prospective clinical trials. I believe that current EMR techniques and devices are only the beginning of a new age in therapeutic endoscopy, the age of minimal invasive endosurgery, the next frontier. PMID- 15665563 TI - [Laparoscopic surgery for gastric cancer]. AB - Recently, the detection of early gastric cancer (EGC) has increased because of the development of diagnostic techniques. Accordingly, new surgical procedures with minimal invasiveness including laparoscopic gastrectomy have been developed. Since the first laparoscopic-assisted distal gastrectomy (LADG) for EGC was performed, various new laparoscopic procedures, such as laparoscopic wedge resection (LWR) and intragastric mucosal resection (IGMR) have been developed for the treatment of EGC. Laparoscopic approaches to gastric cancer provide for minimal invasion, early recovery and decreased morbidity and mortality according to several short-term results. In the future, laparoscopic procedures for gastric cancer will be widely accepted in Korea, if the advantages of laparoscopic approaches are confirmed in randomized controlled trials of long-term outcomes. PMID- 15665564 TI - [Conventional surgery for gastric cancer]. AB - In Korea, gastric cancer ranks second among all the causes of death from cancer, and is the most frequently diagnosed cancer. Surgery is the only possible curative treatment, and results of gastrectomy have improved throughout the years with respect to survival, morbidity, and postoperative mortality. Extent of gastric resection and type of reconstruction can affect the survival and postoperative quality of life (QOL) in gastric cancer patients. Various kinds of reconstruction, especially after total gastrectomy, has been applied to improve the QOL. Eastern, especially Korean and Japanese, surgeons differ in their approaches to lymph node dissection during surgery for stomach cancer compared to Western surgeons. D2 lymph node dissection has not gained widespread popularity in the West, because improved survival has never been demonstrated in randomized trials. However, Korean surgeons believe that D2 dissection is not a dangerous procedure if done by specialists in specialized centers. It is still a matter of question if chemoradiotherapy after D2 dissection can improve the results of D2 dissection alone. However, the more urgent issue is to establish the standard adjuvant chemotherapy protocol after the good local control by surgery (D2 or more). PMID- 15665565 TI - [The relationship between intake of vegetables and fruits and colorectal adenoma carcinoma sequence]. AB - BACKGROUND/AIMS: Evidence for the effect of dietary factors on colorectal carcinogenesis is yet inconsistent. Few studies have been conducted to investigate whether dietary factors were associated with the development of colorectal adenoma-carcinoma sequence in Korea. We evaluated the relationship between the intake of vegetables and fruits and the degree of dysplasia of the colorectal adenoma and cancer. METHODS: For this study, 539 cases with histopathologically confirmed incidental colorectal adenoma, 162 cases with colorectal cancer and 2,576 controls were collected from Our Lady of Mercy Hospital, The Catholic University of Korea during 1994-1999. Informations on demographic characteristics, life style habits and dietary intake were obtained by interviewed questionnaire before the colonoscopy. Adjusted odds ratio (OR) and 95% confidence interval (CI) were estimated by using polytomous logistic regression model. RESULTS: In female, the high intake of raw green and yellow vegetables were found to be negatively associated with the risk of colorectal cancer (adjusted OR: 0.54, 95% CI = 0.32-0.93) and the high intake of persimmon, mandarin and strawberry among fruits were negatively associated with the risk of adenoma with mild dysplasia (adjusted OR: 0.43, 95% CI = 0.20-0.94). In male, the high intake of banana, pear, apple and watermelon among fruits were negatively associated with the risk of colorectal cancer (adjusted OR: 0.36, 95% CI = 0.16 0.84). CONCLUSIONS: These findings suggest that the intake of vegetables and fruits may act differently in developmental steps of colorectal adenoma-carcinoma sequence. PMID- 15665566 TI - [Granulocyte and monocyte adsorption apheresis in Korean conventional treatment refractory patients with active ulcerative colitis: a prospective open-label multicenter study]. AB - BACKGROUND/AIMS: In chronic inflammatory conditions such as ulcerative colitis (UC), the migration of granulocytes and monocytes/macrophages from the circulation into the colonic mucosa is especially important in maintaining inflammation. The aim of this trial was to assess safety and efficacy of granulocyte and monocyte adsorption apheresis in patients with moderate-to-severe UC refractory to conventional drug therapies. METHODS: Twenty-seven patients with moderate (55.6%) to severe (44.4%) active UC refractory to conventional drug therapies who had no changes in their conventional therapy regimen in the past two weeks before the recruitment were enrolled in an open-label trial. Concomitant medications were allowed, and steroids were tapered down according to the clinical activity during the course. We used an adsorptive type extracorporeal column (Adacolumn; JIMRO, Takasaki, Japan), which selectively adsorb granulocytes and monocytes. Patients took five apheresis sessions, each with 60 minutes duration for 5 consecutive weeks. The primary efficacy variables were clinical disease activity, short inflammatory bowel disease questionnaire (SIBDQ), C-reactive protein (CRP), and endoscopic scores. These variables were scored at regular intervals, and analyzed at week 7 on an intention-to-treat (ITT) principles. RESULTS: At 7 weeks, 70.4% of patients showed overall improvement. Clinical disease activity (p < 0.0001), endoscopic score (p < 0.001), and the quality of life as assessed by SIBDQ (p < 0.0001) were significantly improved after the therapy. In 56.3% of concomitant steroid users, tapering down or discontinuation of steroids was possible. Treatment was well tolerated, and no severe adverse events were observed. CONCLUSIONS: Adacolumn was very efficacious in patients with moderate-to-severe active UC refractory to conventional drug therapy, but further assessment is needed. PMID- 15665567 TI - [Is the prevalence of cryptogenic hepatocellular carcinoma increasing in Korea?]. AB - BACKGROUND/AIMS: Nonalcoholic fatty liver disease (NAFLD) has been characterized by a wide spectrum of liver damages that span from steatosis to cryptogenic liver cirrhosis and even to hepatocellular carcinoma (HCC). The aims of this study were to determine whether the prevalence of HCC arising from cryptogenic cirrhosis has increased during the last ten years and to characterize the clinical features of cryptogenic HCC in Korea. METHODS: A retrospective and hospital-based analysis of the clinical data was done in 1,145 HCC patients; group A (Jan. 1993-Dec. 1995), group B (Jan. 2000-Dec. 2002). The etiologies of HCC with liver cirrhosis in group A and group B were analyzed. The risk factors of NAFLD such as obesity, type 2 diabetes mellitus, hypertriglyceridemia and hypertension between cryptogenic HCC and HCC with well-defined etiologies were compared. RESULTS: The major leading causes of HCC in each group were hepatitis B virus infection, followed by alcohol, hepatitis C virus and cryptogenic. There was a significant increase in the proportion of cryptogenic HCC in group B (A: 2.3%, B: 5.4%, p < 0.05). In the case of HCV, it was 5.3% in group A and 9.9% in group B (p < 0.05). Although the prevalence of cyptogenic HCC was significantly increased at an interval of seven years apart, there was no significant difference in the proportions of risk factors of NAFLD between cryptogenic HCC group and well defined etiology group. CONCLUSIONS: The prevalence of cryptogenic HCC was significantly increased in Korea during the last decade. Although statistically insignificant, there was a trend toward the higher proportion of risk factors with NAFLD in patients with cryptogenic HCC. This suggests that increased proportion of risk factors associated for NAFLD may have contributed to the development of cryptogenic HCC. PMID- 15665568 TI - [Expression of epidermal growth factor receptor, ErbB2 and matrix metalloproteinase-9 in hepatolithiasis and cholangiocarcinoma]. AB - BACKGROUND/AIMS: Hepatolithiasis is a common disease in East Asia and presents as a histological feature of proliferative glands containing mucin. 5-10% of hepatolithiasis is known to be associated with cholangiocarcinoma. Recent studies reported that epidermal growth factor receptor (EGFR) could be activated through heparin binding- EGF cleavage by metalloproteinases. Matrix metalloproteinases (MMPs) which digest the extracellular matrix are required for cancer cell invasion and the expression of MMP-9 is known to be increased in cholangiocarcinoma. However, there has been few studies on the expressions and roles of EGFR and MMP in hepatolithiasis. This study was performed to clarify and compare the expressions of EGFR, erbB2 and MMP-9 in hepatolithiasis and cholangiocarcinoma. METHODS: Surgically resected liver tissues with hepatolithiasis (n = 14), cholangiocarcinoma (n = 20) and trauma (n = 2 as controls) were included. The expressions of EGFR, erbB2 and MMP-9 in tissue samples were examined by immunohistochemistry using respective monoclonal antibodies. RESULTS: In traumatic livers, the expressions of EGFR, erbB2 and MMP 9 were all negative. The expression of EGFR was increased in hepatolithiasis group (79%, 11/14) compared with cholangiocarcinoma group (25%, 5/20) (p < 0.05). The expression of erbB2 was detected only in cholangiocarcinoma (25%, 5/20). MMP 9 was increased in both hepatolithiasis (79%, 11/14) and cholangiocarcinoma (95%, 19/20) (p > 0.05). CONCLUSIONS: EGFR expression appears to be the dominant component in periductular hyperplasia of hepatolithiasis and MMP-9 is upregulated not only in cholangiocarcinoma but also in hepatolithiasis. This study suggests that EGFR and MMP-9 are associated with cholangiocarcinoma and hepatolithiasis. PMID- 15665569 TI - [A case of intestinal tuberculosis presenting massive hematochezia controlled by endoscopic coagulation therapy]. AB - The clinical manifestations of intestinal tuberculosis are non-specific. But, abdominal pain, low grade fever, weight loss, anorexia, and diarrhea are major symptoms of intestinal tuberculosis. Massive bleeding has been reported as a rare manifestation of intestinal tuberculosis. Massive hematochezia from intestinal tuberculosis has rarely been reported in the medical literature. Also, most of them were treated with anti-tuberculosis medication only or with surgery. We treated a case of intestinal tuberculosis presenting massive hematochezia with colonoscopic coagulation therapy and anti-tuberculosis medication. Here, we report a Korean man who presented with massive hematochezia from ileal tuberculosis and treated by endoscopic coagulation therapy. PMID- 15665570 TI - [Two cases of histopathologically advanced (stage IV) early gastric cancer]. AB - Various minimally invasive surgical techniques in some cases of early gastric cancer are becoming common practice. However, there are rare cases of advanced cancer with distant metastasis although the invasion of the gastric wall is limited to the mucosa and/or submucosa (defined as early gastric cancer according to UICC-TNM classification). We report two cases of early gastric cancer with distant metastasis (stage IV). Both tumors were defined as early cancer because they were confined to the submucosa. One was a type IIa early cancer, histologically classifiable as a signet ring cell carcinoma (according to the Japanese Classification of Gastric Carcinoma and UICC-TNM classification); the other was a surface spreading type IIb + IIc, classifiable as a signet ring cell carcinoma, too. Stage IV factors were ovarian metastasis (Krukenberg tumor) in the former and N3 in the latter case. PMID- 15665571 TI - [Chemoprevention of colorectal cancer: can it be possible by food?]. PMID- 15665573 TI - [Treatment of chronic hepatitis C]. AB - Chronic infection with HCV represents second most common cause of end-stage liver diseases and hepatocellular carcinoma in Korea. The introduction of new agents and regimens for the treatment of chronic hepatitis C, such as pegylated forms of interferon-alpha (Peg-IFN) and combination with oral ribavirin has resulted in substantial improvement in sustained virologic response (SVR) rates. SVR rate of Peg-IFN and ribavirin combination therapy can be 40-46% of individuals infected with genotype 1 and approximately 75-85% with genotype 2 and 3. Peg-IFN/ribavirin combination therapy represents current standard therapy of chronic hepatitis C. This article reviews the treatment objectives, outcomes, optimal regimens, efficacy and predictors of response, monitoring during treatment, adverse events, retreatment of persons who failed to respond to previous treatments, and treatment of special patient groups in chronic hepatitis C. PMID- 15665574 TI - [Altered colonic transit in TNBS-induced experimental colitis in guinea pig and distribution of nitric oxide synthase in the colonic wall]. AB - BACKGROUND/AIMS: Inflammation-induced alterations in smooth muscle contractility may be due to the effects on smooth muscle itself, neurotransmitters or enteric nerves. In dextran sulfate sodium-induced colitic rat, the delay in colonic transit was caused by decreased activity and production of neuronal nitric oxide synthase (nNOS) in the myenteric plexus of the distal colon. The aim of this study was to investigate the relationship between the delay in colonic transit and the distribution of inducible NOS (iNOS) and nNOS immunoreactive cells in the myenteric plexus of trinitrobenzene sulfonic acid (TNBS)-induced colitic guinea pig. METHODS: Sacrificed and their colonic tissues of forty-five TNBS-induced colitic guinea pigs were used to measure the colonic transit, and analyzed by immunohistochemistry. RESULTS: Colonic transit was delayed significantly at 3, 7 and 14 days after administration of TNBS. In control, nNOS immunoreactivity was present in the mucosa, submucosa, lamina propria, and ganglion cells of the myenteric plexus, while after TNBS treatment, reduced nNOS cells were found. However, the number of nNOS ganglion cells in the myenteric plexus was similar to those seen in controls. After administration of TNBS, iNOS immunoreactivity was increased in the mucosa and submucosa, but the number of iNOS positive ganglion cells in the myenteric plexus was not changed compared to control. CONCLUSIONS: It is suggested that in TNBS-induced guinea pig colitis, delayed colonic transit is not associated with the expression of nNOS nor iNOS in the myenteric plexus. PMID- 15665575 TI - Cyclin E, p27 and mutant p53 do not predict the prognosis in AJCC stage II colorectal carcinomas. AB - BACKGROUND/AIMS: Carcinogenesis is characterized by the abnormal regulation of cell cycle. The abnormal expression of the regulators of cell cycle may be related to the prognosis. Since the clinical significance of the expression of the three proteins in colorectal carcinomas is still controversial, we evaluated the prognostic value of the expression of cyclin E, p27 and mutant p53 in stage II colorectal cancer. METHODS: The expression levels of cyclin E, p27 and mutant p53 proteins in 41 patients with stage II colorectal carcinomas were analyzed by immunohistochemistry. RESULTS: In the univariate analysis, the level of CEA at diagnosis was associated with disease relapse. In the multivariate analysis, the clinicopathological variables such as age, gender, site of primary tumor, tumor size, state of tumor differentiation and preoperative plasma CEA level were not associated with disease relapse. When Kaplan-Meier survival curves were constructed to determine the prognosis, cyclin E, p27 and mutant p53 expressions did not predict poor prognosis. CONCLUSIONS: Our results suggested that the expression of cyclin E, p27 and mutant p53 proteins did not predict the clinical outcome in the stage II colorectal carcinomas. PMID- 15665576 TI - [Clinical significance of erosive and/or small ulcerative lesions in the colon and terminal ileum--short-term follow-up study]. AB - BACKGROUND/AIMS: Various etiologies and diseases may be related to erosions and/or small ulcers without gross inflammatory changes in the surrounding mucosa found in the colon and terminal ileum during colonoscopy. However, studies on follow-up of these lesions are rare. Thus, we investigated the clinical significance of these lesions and their characteristics helpful for differential diagnosis. METHODS: We reviewed the data of 183 patients with colonoscopically observed erosive or small ulcerative lesions (<2 cm), and analyzed them according to the location, number, and size of lesions, histopathologic findings, chief complaints, laboratory findings, changes of symptoms, and changes in lesions during 4-12 week follow-up period. RESULTS: Histopathologic findings of these lesions included acute nonspecific inflammation, chronic nonspecific inflammation, Crohn's disease, tuberculous colitis, ischemic colitis, Behcet's disease, cytomegalovirus infection, eosinophilic colitis, ulcerative colitis or pseudomembranous colitis, but most of them were nonspecific (84%). In patients with nonspecific inflammation, histopathologic findings, symptoms, location and multiplicity of the lesions were not prognostic factors for the persistency of symptoms and lesions during follow-up period. Two patients with acute inflammation, who showed no improvement in symptoms and lesions, were later diagnosed as Crohn's disease. CONCLUSIONS: Erosive or small ulcerative lesions without macroscopic inflammatory changes in the surrounding mucosa during colonoscopy, are mainly nonspecific. However, careful follow-up is required when the symptoms and/or lesions are not improved. PMID- 15665577 TI - [Oxysterol (3,5-cholestadien-7-one, 5 beta-cholestan-3-one, 5,24-cholestadien-3 beta-OL) induced cytotoxicity and apoptosis in gallbladder epithelial cells]. AB - BACKGROUND/AIMS: Biliary epithelial cells are exposed to highly concentrated oxysterols. Therefore, oxysterols may play a role in pathogenesis of biliary tract diseases. We investigated the cytotoxic effect and apoptosis inducing effect of oxysterol on gallbladder epithelial cells. METHODS: We studied the cytotoxic effect of 3,5- cholestadien-7-one, 5 beta-cholestan-3-one and 5,24 cholestadien-3 beta-OL which are identified in human bile and pigment gallstones on dog gallbladder epithelial cells (DGBE) and mouse gallbladder epithelial cells (MGBE). We used model bile to dissolve oxysterols as in vitro experiment and also used MTT, cell count, Diff-Quick stain, and flow cytometry to investigate cytotoxicity and apoptosis. RESULTS: Oxysterols dissolved in model bile have cytotoxic effects in a dose dependent fashion. In oxysterol containing model bile, viable cells are 51% in 500 microM 5 beta-cholestan-3-one (cholesterol:oxysterol 50:50) and 47% in 5 mM 3,5-cholestadien-7-one (90:10) on MGBE, and are 129% and 38% in 500 microM (50:50) 3,5-cholestadien-7-one and 5 beta-cholestan-3-one on DGBE, and are 74% and 71.5% in 5 mM (90:10) 3,5 cholestadien-7-one and 5 beta-cholestan-3-one on DGBE, respectively. 500 microM (50:50) 3,5- cholestadien-7-one, 5 beta-cholestan-3-one, and 5,24-cholestadien-3 beta-OL treated on DGBE increase the apoptotic cell number as 22.0+/-8.8, 30.2+/ 12.6, and 45.5+/-13.2%, respectively, compared with control (14.6+/-10.0%). 500 microM (50:50) 3,5-cholestadien-7-one, 5 beta-cholestan-3-one, and 5,24 cholestadien-3 beta-OL also affect the changes in cell cycles compared with the control. CONCLUSIONS: We concluded that oxysterol containing model bile is useful as an in vitro experiment as model to analyze the effects of oxysterols on biliary epithelial cells and that adequate concentration of oxysterols can induce the cytotoxic effect and the apoptosis on gallbladder epithelial cells. PMID- 15665578 TI - [A case of reactivated tuberculous colitis after 9 months of anti-tuberculous therapy]. AB - Tuberculous colitis, an important extra-pulmonary tuberculosis, is still prevalent in the developing countries and has been resurging in the Western world. The duration and dose of anti-tuberculous therapy have not yet been clarified in the tuberculous colitis. We experienced a case of tuberculous colitis, which relapsed after 9 months of therapy. A 28-year-old man presented with hematochezia and was diagnosed as tuberculous colitis on the basis of colonoscopic findings. He was treated with anti-tuberculous agents for 9 months successfully. Three months later, however, he complained of hematochezia again, suggesting the relapse of tuberculous colitis. He had taken anti-tuberculous therapy for another 15 months and showed no evidence of relapse. Although anti tuberculous therapy is efficient for tuberculous colitis, rare cases of reactivation should be reminded. PMID- 15665580 TI - [Cell cycle regulators as prognostic predictor of colorectal cancers]. PMID- 15665579 TI - [A case of acute hepatitis E with the travel history to an endemic area]. AB - Hepatitis E is a self-limited and enterically transmitted acute viral hepatitis that occurs from epidemic outbreaks of developing countries and sporadic hepatitis in non-endemic areas. In endemic areas, hepatitis E virus (HEV) is one of the most common causes of acute hepatitis and hepatic failure in pregnancy. Its mortality rate has been reported up to 20%. In non-endemic areas, HEV infection without any travel history is very rare. In Korea, only one case of simple hepatitis E without any travel history to endemic areas was reported. Recently, we experienced a case of acute hepatitis. The patient who had a travel history to India, showed watery diarrhea and high fever. Transaminase level and total bilirubin were increased, and prothrombin time was prolonged. It was positive for IgM anti-HEV and IgG anti-HEV, and showed no evidence of other viral infections or drug ingestion history. In spite of absence of useful test such as seroconversion of IgM anti-HEV and HEV RNA PCR, we diagnosed the case as an acute hepatitis E from his symptom, travel history and initial serologic marker. We report this as a case of hepatitis E infected from endemic areas. PMID- 15665582 TI - The potential roles of p53 tumor suppressor in nucleotide excision repair (NER) and base excision repair (BER). AB - The p53 tumor suppressor has long been envisaged to preserve genetic stability by the induction of cell cycle checkpoints and apoptosis. More recently, p53 has been implicated to play roles in DNA repair responses to genotoxic stresses. UV damage and the damage caused by certain chemotherapeutics including cisplatin and nitrogen mustards are known to be repaired by the nucleotide excision repair (NER) pathway which is reportedly regulated by p53 and its downstream genes. There are evidences to suggest that the base excision repair (BER) induced by the base-damaging agent methyl methanesulfonate (MMS) is partially deficient in cells lacking functional p53. This result suggests that the activity of BER might be also dependent on the p53 status. In this review, we discuss the possibilities that p53 regulates BER as well as NER; these are one of the most significant potentials of p53 tumor suppressor for repairing the vast majority of DNA damages that is incurred from various environmental stresses. PMID- 15665583 TI - The upstream sequence of Mycobacterium leprae 18-kDa gene confers transcription repression activity in orientation-independent manner. AB - In order to understand the role of the upstream region of the Mycobacterium leprae 18-kDa gene on the gene regulation, the region was divided into two at the -50 position from the first start codon of the gene and their effect on transcription was examined by using a LacZ transcriptional reporter gene assay. The presence of each of these two regions conferred transcription repression not only on its cognate M. lepraerae 18-kDa gene promoter, but also on a heterologous promoter such as the Mycobacterium bovis BCG hsp65 gene promoter. Moreover, it was found that these regions could confer transcription repression activity in both cases in an orientation-independent manner. Thus, these results indicate that the upstream region of the M. leprae 18-kDa gene harbors transcription repression responsive element(s) acting as an operator and can be further divided into two separately functional regions, suggesting a bipartite structure of the element(s). The identification of transcription repression activity of the upstream region in the M. leprae 18-kDa gene will contribute greatly for the understanding of the 18-kDa gene regulation mechanism, and provide also useful information for the manipulation of mycobacterium gene expression. PMID- 15665584 TI - 5-azacytidine induces cardiac differentiation of P19 embryonic stem cells. AB - The P19 embryonal carcinoma cell line is a useful model cells for studies on cardiac differentiation. However, its low efficacy of differentiation hampers its usefulness. We investigated the effect of 5-azacytidine (5-aza) on P19 cells to differentiate into a high-efficacy cardiomyocytes. Embryoid-body-like structures were formed after 6 days with 1 mM of 5-aza in a P19 cell monolayer culture, beating cell clusters first observed on day 12, and, the production of beating cell clusters increased by 80.1% (29 of 36-wells) after 18 days. In comparison, the spontaneous beating cells was 33.3% (12 of 36-wells) for the untreated control cells. In response to 1 mM of 5-aza, P19 cells expressed bone morphogenetic protein-2 (BMP-2), BMP-4, Bmpr1a and Smad1 at day 6 or 9, and also cardiac markers such as GATA-4, Nkx2.5, cardiac troponin I, and desmin were up regulated in a time-dependent manner after induction of BMP signaling molecules. Immunocytochemistry revealed the expression of smooth muscle a-actin, sarcomeric a-actinin, cardiac myosin heavy chain, cardiac troponin T and desmin, respectively. The proportion of sarcomeric a-actinin positive cells accounted for 6.48% on day 15 after 5-aza exposure as measured by flow cytometry. This study has demonstrated that 5-aza induces differentiation of P19 cells into cardiomyocytes in a confluent monolayer culture in the absence of prior embryoid formation and dimethyl sulfoxide exposure, depending in part on alteration of BMP signaling molecules. These results suggest that 5-aza treatment could be used as a new method for cardiac differentiation in P19 cells. PMID- 15665585 TI - Ongoing angiogenesis in blood vessels of the abdominal aortic aneurysm. AB - Pathogenesis of the abdominal aortic aneurysm has been attributed to neovascularization of the aortic wall. However, it is not clear whether angiogenesis persists in the aneurysm. In sections of aneurysms, we determined the immunohistochemical distributions of the avb3 integrin, tenascin and endothelial nitric oxide synthase (eNOS), which are markers respectively, of angiogenesis, matrix remodeling and vasoregulatory function. In addition, we used reverse transcription followed by in situ PCR, to determine the distribution of av mRNA. All aneurysm specimens exhibited extensive increases of wall vascularization as compared with the control aortic wall, and showed the presence of perivascular inflammatory exudates containing macrophages and lymphocytes. The neovascularization consisted of thick-walled vessels in the media and adventitia, and capillaries in the subintima. The majority of vessels stained positively for the avb3 antigen and eNOS. Tenascin was deposited as bands that circumscribed thick-walled vessels. The distribution of av mRNA was extensive and was positive even in those vessels that failed to stain for the avb3 protein. No staining was evident in control aortas for the avb3 antigen, tenascin or av mRNA. The upregulation of av mRNA and the avb3 integrin in blood vessels surrounded by a matrix expressing tenascin, indicates that angiogenesis is an ongoing process in the mature aortic aneurysm. PMID- 15665586 TI - Ox-LDL suppresses PMA-induced MMP-9 expression and activity through CD36-mediated activation of PPAR-g. AB - During chronic inflammatory response, mono- cytes/macrophages produce 92-kDa matrix metalloproteinase-9 (MMP-9), which may contribute to their extravasation, migration and tissue remodeling. Activation of peroxisome proliferator- activated factor receptor-g (PPAR-g) has been shown to inhibit MMP-9 activity. To evaluate whether ox-LDL, a PPAR-g activator, inhibits PMA-induced MMP-9 expression and activity, and if so, whether CD36 and PPAR-g are involved in this process, we investigated the effect of ox-LDL on MMP-9 expression and activity in PMA activated human monocytic cell line U937. PMA-induced MMP-9 expression and activity were suppressed by the treatment with ox-LDL (50 mg/ml) or PPAR-g activators such as troglitazone (5 mM), ciglitazone (5 mM), and 15d- PGJ2 (1 mM) for 24 h. This ox-LDL or PPAR-g activator-mediated inhibition of MMP-9 activity was diminished by the pre-treatment of cells with a blocking antibody to CD36, or PGF2a (0.3 mM), which is a PPAR-g inhibitor, as well as overexpression of a dominant-negative form of CD36. Taken together, these results suggest that ox-LDL suppresses PMA-induced MMP-9 expression and activity through CD36-mediated activation of PPAR-g. PMID- 15665587 TI - Corn silk induces nitric oxide synthase in murine macrophages. AB - Corn silk has been purified as an anticoagulant previously and the active component is a polysaccharide with a molecular mass of 135 kDa. It activates murine macrophages to induce nitric oxide synthase (NOS) and generate substantial amounts of NO in time and dose-dependent manners. It was detectable first at 15 h after stimulation by corn silk, peaked at 24 h, and undetectable by 48 h. Induction of NOS is inhibited by pyrolidine dithiocarbamate (PDTC) and genistein, an inhibitor of nuclear factor kappa B (NF-kappaB) and tyrosine kinase, respectively, indicating that iNOS stimulated by corn silk is associated with tyrosine kinase and NF-kappaB signaling pathways. IkappaB-alpha degradation was detectible at 10 min, and the level was restored at 120 min after treatment of corn silk. Corn silk induced nuclear translocation of NF-kappaB by phosphorylation and degradation of IkappaB-alpha. PMID- 15665588 TI - Pseudolaric acid B induces apoptosis via activation of c-Jun N-terminal kinase and caspase-3 in HeLa cells. AB - Pseudolaric acid B was isolated from Pseudolarix kaempferi Gordon (Pinaceae) and was evaluated for the anti-cancer effect in HeLa cells. We ob-served that pseudolaric acid B inhibited cell proliferation and induced apoptosis in a time- and dose-dependent manner. HeLa cells treated with pseudolaric acid B showed typical characteristics of apoptosis including the morphological changes and DNA fragmentation. JNK inhibitor, SP600125,markedly inhibited pseudolaric acid B induced celldeath. In addition, Bcl-2 expression was down-regulated while Bax protein level was up-regulated.Caspase-3 inhibitor, z-DEVD-fmk, partially blocked pseudolaric acid B-induced cell death, and the expression of two classical caspase substrates,PARP and ICAD, were both decreased in a time-dependent manner, indicative of downstream cas-pase activation. PMID- 15665589 TI - p21Cip/WAF1 activation is an important factor for the ERK pathway dependent anti proliferation of colorectal cancer cells. AB - p21Cip/WAF1, an important regulator of cell proliferation, is induced by both p53 and extracellular signal regulated kinase (ERK) pathways. The induction of p21Cip/WAF1 occurs by prolonged activation of the ERKs caused by extracellular stimuli, such as zinc. However, not all the cells appeared to respond to ERK pathway dependent p21Cip/WAF1 induction. Here we investigated the cause of such difference using colorectal cancer cells. p21Cip/WAF1 induction and concomitant reduction of bromodeoxyuridine (BrdU) incorporation were observed by zinc treatment within HT-29 and DLD-1. However, HCT-116 cells with high endogenous p21Cip/WAF1 levels did not show any additional increment of p21Cip/WAF1 levels by zinc treatment and did maintain high BrdU incorporation level. The p21Cip/WAF1 induction by zinc depended upon prolonged activation of extracellular signal regulated kinase (ERK) was not observed in HCT-116 cells. The percentage of BrdU positive cells was 50% higher in p21Cip/WAF1 -/- HCT-116 cells compared to p21Cip/WAF1 +/+ HCT- 116 cells, and no cells induced p21Cip/WAF1 incorporated BrdU in its nucleus, yet confirming the importance of p21Cip/WAF1 induction in anti- proliferation. These results again support that p21Cip/WAF1 induction is a determinant in the regulation of colonic proliferation by the ERK pathway. PMID- 15665590 TI - Association of the XRCC1 gene polymorphisms with cancer risk in Turkish breast cancer patients. AB - The X-ray repair cross-complementing group 1 (XRCC1) gene is believed to play an important role in base excision repair and displays genetic polymorphisms. Data on the role of XRCC1 polymorphisms in cancer susceptibility is inconsistent. In the present study, we investigated the effect of two XRCC1 polymorphisms, Arg194Trp and Arg399Gln, on breast cancer risk in a case- control study involving Turkish breast cancer patients and healthy women. Both alleles exhibited a similar distribution among cases and controls leading to lack of any significant association between the XRCC1 polymorphisms and breast cancer risk, either in homozygotes and heterozygotes or combined. The allele frequency of the codon 194 variant was very low in cases and healthy individuals (5.3 and 3.9%, respectively) compared to that of the variant 399Gln allele (39.7 and 37.4%). Our results do not support evidence for a role of the XRCC1 polymorphism in developing breast cancer. PMID- 15665592 TI - Gel-based application of siRNA to human epithelial cancer cells induces RNAi dependent apoptosis. AB - Gene silencing by RNA interference (RNAi) operates at the level of mRNA that is targeted for destruction with exquisite sequence specificity. In principle, any disease-related mRNA sequence is a putative target for RNAi-based therapeutics. To develop this therapeutic potential, it is necessary to develop ways of inducing RNAi by clinically acceptable delivery procedures. Here, we ask if inducers of RNAi can be delivered to human cells via a gel-based medium. RNAi was induced using synthetic small interfering RNAs (siRNAs), which bypass the need for expression vectors and carry the added bonus of high potency and immediate efficacy. Established cultures of human cells of normal and tumor origin were overlaid with an agarose/liposome/siRNA gel formulation without adverse effects on cell viability or proliferation. Epithelial cancer cells (but not normal human fibroblasts) proved vulnerable to specific siRNAs delivered via the agarose/liposome/siRNA formulation. Moreover, proapoptotic siRNAs induced apoptosis of cervical carcinoma cells (treated with human papillomavirus [HPV] E7 siRNA) and of colorectal carcinoma cells (treated with Bcl-2 siRNA). Thus, we demonstrate successful topical gel-based delivery of inducers of RNAi to human epithelial cancer cells. Topical induction of RNAi opens an important new therapeutic approach for treatment of human diseases, including cervical cancer and other accessible disorders. PMID- 15665591 TI - bPAK-interacting exchange factor may regulate actin cytoskeleton through interaction with actin. AB - p21-activated kinase (PAK)-interacting exchange factor (PIX) is known to be involved in regulation of Cdc42/Rac GTPases and PAK activity. PIX binds to the proline-rich region of PAK, and regulates biological events through activation of Cdc42/Rac GTPase. To further investigate the role of PIX we produced monoclonal antibodies (Mab) against bPIX. Three clones; N-C6 against N-terminal half and C A3 and C-B7 against C- terminal half of bPIX were generated and characterized. N C6 Mab detected bPIX as a major band in most cell lines. C-A3 Mab recognizes GIT binding domain (GBD), but it does not interfere with GIT binding to bPIX. Using C A3 Mab possible bPIX interaction with actin in PC12 cells was examined. bPIX Mab (C-A3) specifically precipitated actin of the PC12 cell lysates whereas actin Mab failed to immunoprecipitate bPIX. Co-sedimentation of PC12 cell lysates with the polymerized F-actin resulted in the recovery of most of bPIX in the cell lysates. These results suggest that bPIX may not interact with soluble actin but with polymerized F-actin and revealed that bPIX constitutes a functional complex with actin. These data indicate real usefulness of the bPIX Mab in the study of bPIX role(s) in regulation of actin cyoskeleton. PMID- 15665593 TI - Sequence-specific cleavage of hepatitis X RNA in cis and trans by novel monotarget and multitarget hammerhead motif-containing ribozymes. AB - We constructed two monoribozymes and a diribozyme against the conserved region of the X RNA of hepatitis B virus (HBV). All the ribozymes (Rzs) possessed sequence specific cleavage activities under standard and simulated physiologic conditions. Specific cleavage was also obtained when the same Rzs were placed in cis configuration with respect to X gene in multiple combinations. Rz-expressing cells were able to specifically interfere with the functional expression of X RNA and protein production in a liver-specific cell line HepG2. Potential applications of these novel Rzs are discussed. PMID- 15665594 TI - Telomerase reverse transcriptase (hTERT) mRNA and telomerase RNA (hTR) as targets for downregulation of telomerase activity. AB - Telomerase is expressed in cancer cells but not in most normal cells, leading to the hypothesis that telomerase inhibitors may be a powerful approach to cancer therapy. It is possible that telomerase plays roles in the cell other than telomere elongation and that blocking telomerase expression may have consequences that differ from simply blocking the active site through competitive inhibition. Here, we test this hypothesis by comparing the effects of antisense oligonucleotides and small interfering RNAs (siRNAs) that target the telomerase reverse transcriptase (hTERT) mRNA with the effects of oligonucleotides that target the telomerase RNA component (hTR). We find that the use of anti-hTR oligomers is more effective in blocking telomerase expression than strategies that target hTERT mRNA. Anti-hTR compounds are active on addition to cells in the absence of lipid, whereas antisense oligonucleotides are not. The modest inhibition of hTERT expression caused by antisense oligonucleotides or siRNAs does not persist, suggesting development of resistance. These data suggest that strategies for telomerase inhibition that require downregulation of hTERT mRNA may be less straightforward than those that target hTR. In addition, we have not seen evidence for a role for hTERT other than in telomere maintenance. PMID- 15665595 TI - Double displacement loops (double d-loops) are templates for oligonucleotide directed mutagenesis and gene repair. AB - Appreciable levels of gene repair result from the hybridization of two oligonucleotides at a specific site in a mutated gene and subsequent correction by a form of oligonucleotide-directed mutagenesis known as gene repair. The incorporation of the two oligonucleotides into superhelical plasmid DNA leads to the formation of double d-loops, structures shown to be templates for the repair of both frameshift and point mutations. Structural limitations placed on the template indicate that correction is influenced significantly by the positioning of the second oligonucleotide, known as the annealing oligonucleotide. Complexes constructed with two oligonucleotides directly opposite each other exhibit the highest levels of gene repair activity. Blocking the 3'-end of either oligonucleotide with an amino C7 group does not diminish the performance of the double d-loop as a template for correction of the point mutation, suggesting that primer extension does not play a pivotal role in the mechanism of gene repair. PMID- 15665596 TI - Binding and uptake of immunostimulatory CpG oligodeoxynucleotides by human neuroblastoma cells. AB - Oligodeoxynucleotides (ODNs) that contain unmethylated CpG dinucleotides (CpG ODN) trigger a strong innate immune response in vertebrates. They have been used to eradicate experimental neuroblastoma, but a direct interaction of CpG-ODN with neuroblastoma cells has not been investigated. We have analyzed uptake, binding, and intracellular distribution of CpG-ODN in the neuroblastoma cells line SKNSH. Our results indicate that cellular uptake of CpG-ODN is dose, time, temperature, and energy dependent but independent of the CpG motif. After internalization, CpGODN localized to the cytoplasm and showed a typical speckled distribution pattern. The intracellular distribution pattern and binding proteins are CpG motif independent as well. Thus, CpG-ODNs are taken up by neuroblastoma cells by a nonspecific transfer mechanism for oligonucleotides and interact with intracellular proteins. These mechanisms might help us to understand the biodistribution of oligo within tumors and might be helpful in evaluating the therapeutic effects of oligonucleotides and rational drug design. PMID- 15665597 TI - Distribution and excretion of a phosphorothioate oligonucleotide in rats with experimentally induced renal injury. AB - The effects of renal injury on the urinary excretion and tissue distribution of a 20-mer phosphorothioate oligonucleotide were investigated in male Sprague-Dawley rats. Renal injury was produced by treating the rats with either 5.0 mg/kg cisplatin or 2.5 mg/kg of a monoclonal antibody (mAb) directed toward Thy1.1. Controls received saline. Three days after cisplatin treatment or 2 days after anti- Thy1.1 treatment, the rats received 10 mg/kg ISIS 3521. Blood was collected at various times to assess the plasma concentrations of ISIS 3521, and rats were killed at various times from 6 to 48 hours after intravenous (i.v.) infusion of oligonucleotide to assess tissue concentrations by capillary gel electrophoresis (CGE). Cisplatin and anti-Thy1.1 antibody produced histologic and biochemical changes consistent with proximal tubular damage and glomerular damage, respectively. Urinary excretion of oligonucleotides was increased 2- to 4-fold of control; however, this amount accounted for only 1% to 2% of dose compared to 0.5% in controls. Proximal tubular damage reduced renal accumulations of ISIS 3521 and other oligonucleotide metabolites, but there were no obvious compensatory increases in concentrations in other organs except for a slight increase in spleen levels of total oligonucleotide. Glomerular damage was not associated with any change in oligonucleotide disposition. Immunohistochemical studies showed no evidence of alterations in the pattern of distribution within the injured kidney. The data suggest that acute renal dysfunction, either renal tubular or glomerular, does not markedly alter the urinary elimination and tissue deposition of a phosphorothioate oligonucleotide. PMID- 15665598 TI - Mechanism of gene repair open for discussion. AB - During the last decade, chimeric RNA-DNA oligonucleotides (RDOs) and single stranded oligodeoxynucleotides have been used to make permanent and specific sequence changes in the genome, with the ultimate goal of curing human genetic disorders caused by mutations. There have been large variations observed in the rate of gene repair in these studies. This has been due, at least in part, to the lack of standardized assay conditions and the paucity of mechanistic studies in the early developmental stages. Previously, it was proposed that strand pairing is the rate-limiting step and mismatch DNA repair is involved in the gene repair process. We propose an alternative model, in which an oligonucleotide is assimilated to the target DNA during active transcription, leading to formation of a transient D-loop. The trafficking of RNA polymerase is interrupted by the D loop, and the stalled RNA polymerase complex may signal for recruitment of DNA repair proteins, including transcription-coupled DNA repair and nucleotide excision repair. Thus, oligonucleotides can be considered as a class of DNA damaging agents that cause a transient but major structural change in DNA. Understanding of the recognition and repair pathways to process this unusual DNA structure may have relevance in physiologic processes, transcription, and DNA replication. PMID- 15665601 TI - Translational neurochemical research in acute human brain injury: the current status and potential future for cerebral microdialysis. AB - Microdialysis (MD) was introduced as an intracerebral sampling method for clinical neurosurgery by Hillered et al. and Meyerson et al. in 1990. Since then MD has been embraced as a research tool to measure the neurochemistry of acute human brain injury and epilepsy. In general investigators have focused their attention to relative chemical changes during neurointensive care, operative procedures, and epileptic seizure activity. This initial excitement surrounding this technology has subsided over the years due to concerns about the amount of tissue sampled and the complicated issues related to quantification. The interpretation of mild to moderate MD fluctuations in general remains an issue relating to dynamic changes of the architecture and size of the interstitial space, blood-brain barrier (BBB) function, and analytical imprecision, calling for additional validation studies and new methods to control for in vivo recovery variations. Consequently, the use of this methodology to influence clinical decisions regarding the care of patients has been restricted to a few institutions. Clinical studies have provided ample evidence that intracerebral MD monitoring is useful for the detection of overt adverse neurochemical conditions involving hypoxia/ischemia and seizure activity in subarachnoid hemorrhage (SAH), traumatic brain injury (TBI), thromboembolic stroke, and epilepsy. There is some data strongly suggesting that MD changes precede the onset of secondary neurological deterioration following SAH, hemispheric stroke, and surges of increased ICP in fulminant hepatic failure. These promising investigations have relied on MD-markers for disturbed glucose metabolism (glucose, lactate, and pyruvate) and amino acids. Others have focused on trying to capture other important neurochemical events, such as excitotoxicity, cell membrane degradation, reactive oxygen species (ROS) and nitric oxide (NO) formation, cellular edema, and BBB dysfunction. However, these other applications need additional validation. Although these cerebral events and their corresponding changes in neurochemistry are important, other promising MD applications, as yet less explored, comprise local neurochemical provocations, drug penetration to the human brain, MD as a tool in clinical drug trials, and for studying the proteomics of acute human brain injury. Nevertheless, MD has provided new important insights into the neurochemistry of acute human brain injury. It remains one of very few methods for neurochemical measurements in the interstitial compartment of the human brain and will continue to be a valuable translational research tool for the future. Therefore, this technology has the potential of becoming an established part of multimodality neuro-ICU monitoring, contributing unique information about the acute brain injury process. However, in order to reach this stage, several issues related to quantification and bedside presentation of MD data, implantation strategies, and quality assurance need to be resolved. The future success of MD as a diagnostic tool in clinical neurosurgery depends heavily on the choice of biomarkers, their sensitivity, specificity, and predictive value for secondary neurochemical events, and the availability of practical bedside methods for chemical analysis of the individual markers. The purpose of this review was to summarize the results of clinical studies using cerebral MD in neurosurgical patients and to discuss the current status of MD as a potential method for use in clinical decision-making. The approach was to focus on adverse neurochemical conditions in the injured human brain and the MD biomarkers used to study those events. Methodological issues that appeared critical for the future success of MD as a routine intracerebral sampling method were addressed. PMID- 15665602 TI - Lateral fluid percussion brain injury: a 15-year review and evaluation. AB - This article comprehensively reviews the lateral fluid percussion (LFP) model of traumatic brain injury (TBI) in small animal species with particular emphasis on its validity, clinical relevance and reliability. The LFP model, initially described in 1989, has become the most extensively utilized animal model of TBI (to date, 232 PubMed citations), producing both focal and diffuse (mixed) brain injury. Despite subtle variations in injury parameters between laboratories, universal findings are evident across studies, including histological, physiological, metabolic, and behavioral changes that serve to increase the reliability of the model. Moreover, demonstrable histological damage and severity dependent behavioral deficits, which partially recover over time, validate LFP as a clinically-relevant model of human TBI. The LFP model, also has been used extensively to evaluate potential therapeutic interventions, including resuscitation, pharmacologic therapies, transplantation, and other neuroprotective and neuroregenerative strategies. Although a number of positive studies have identified promising therapies for moderate TBI, the predictive validity of the model may be compromised when findings are translated to severely injured patients. Recently, the clinical relevance of LFP has been enhanced by combining the injury with secondary insults, as well as broadening studies to incorporate issues of gender and age to better approximate the range of human TBI within study design. We conclude that the LFP brain injury model is an appropriate tool to study the cellular and mechanistic aspects of human TBI that cannot be addressed in the clinical setting, as well as for the development and characterization of novel therapeutic interventions. Continued translation of pre clinical findings to human TBI will enhance the predictive validity of the LFP model, and allow novel neuroprotective and neuroregenerative treatment strategies developed in the laboratory to reach the appropriate TBI patients. PMID- 15665603 TI - Changes in white matter in long-term survivors of severe non-missile traumatic brain injury: a computational analysis of magnetic resonance images. AB - The aim of this study was to investigate long-term consequences of severe non missile traumatic brain injury (nmTBI) in patients without macroscopic focal brain lesions (>1.6 cm(3)) on regional white-matter density (WMd), and possible correlations with days of coma and memory performances. T1-weighted magnetic resonance images (MRI) were acquired in 19 nmTBI patients, 3-113 months following the injury, and in 19 control subjects matched for age and gender. In addition, nmTBI patients underwent a battery of standardised memory tests. The MRIs were processed in a fully automatic system using voxel-by-voxel methods. Corpus callosum, fornix, anterior limb of the internal capsule, superior frontal gyrus, para-hippocampal gyrus, optic radiation and chiasma showed significant WMd reduction in nmTBI when compared to control subjects. None of the correlations between days of coma and memory performance scores with nmTBI voxels value that showed WMd reduction reached significance, with the exception of a significant negative correlation between WMd in the mid body of corpus callosum and short story delayed recall. We detected reductions in WM density in several brain locations similar to those described in previous post mortem investigations. In addition, we observed WMd reduction in the optic chiasma and in the optic radiations; this finding may reflect transneural degeneration along the visual pathway. The weak correlations between specific anatomical sites of the reduced WMd and behavior may reflect the diffuse nature of the brain damage and/or the different time of onset between behavioral manifestations and neuropathological modifications occurring in nmTBI. PMID- 15665604 TI - Cleaved-tau: a biomarker of neuronal damage after traumatic brain injury. AB - Previous studies from our laboratory indicate that traumatic brain injury (TBI) in humans results in proteolysis of neuronally-localized, intracellular microtubule associated protein (MAP)-tau to produce cleaved tau (C-tau). The present study evaluated the utility of C-tau to function as a biomarker of neuronal injury and as a biomarker for evaluating neuroprotectant drug efficacy in a controlled cortical impact model of rat TBI. Brain C-tau was determined in rats subjected to controlled cortical impact-induced mild, moderate or severe levels of TBI. A significant severity-dependent increase in C-tau levels was observed in the cortex and hippocampus (1.5-8-fold) of TBI rats compared to shams 72 h after impact. C-tau rat brain and serum time course was determined by measuring levels at 0.25, 6, 24, 48, 72 and 168 h after TBI. A significant time dependent increase in C-tau levels was observed in ipsilateral cortex (5-16-fold) and hippocampus (2-40-fold) compared to sham animals. C-tau levels increased as early as 6 h after TBI with peak C-tau levels observed 168 h after injury. Elevated brain C-tau levels were associated with TBI-induced tissue loss, which was histologically determined. The effect of cyclosporin-A (CsA), previously demonstrated to be neuroprotective in rat TBI, on brain C-tau levels was examined. CsA (20 mg/kg i.p., 15 min and 24 h after TBI) significantly attenuated the TBI-induced increase in hippocampal C-tau levels observed in vehicle-treated animals confirming CsA's neuroprotectant effect. CsA treatment also lowered ipsilateral cortical C-tau levels, although it did not reach statistical significance. CsA's neuroprotectant effect was confirmed utilizing histologic measures of TBI-induced tissue loss. In addition, serum C-tau levels were significantly increased 6 h after TBI but not at later time points. These results suggest that C-tau is a reliable, quantitative biomarker for evaluating TBI induced neuronal injury and a potential biomarker of neuroprotectant drug efficacy in the rat TBI model. Serum data suggests that C-tau levels are dependent both on a compromised blood-brain barrier as well as release of TBI biomarkers from the brain, which has implications for the study of human serum TBI biomarkers. PMID- 15665605 TI - Cell proliferation and neuronal differentiation in the dentate gyrus in juvenile and adult rats following traumatic brain injury. AB - It is well known that the cognitive functions of juveniles recover to a greater extent than adult patients following traumatic brain injury (TBI). The exact mechanisms underlying this age-related disparity are unknown; however, we speculate that this improved recovery in juveniles following TBI may be associated with an endogenous neurogenic response in the hippocampus. We, therefore, examined the effects of TBI on cellular proliferation and differentiation in the dentate gyrus (DG) of the hippocampus in juvenile and adult rats following lateral fluid percussion injury (FPI). The temporal profile of the injury-induced proliferative response was determined using BrdU labeling at varying survival times. The differentiation of these newly generated cells was investigated using cell-type specific markers. We found that, following injury, there was a significant increase in cell proliferation in the DG in both injured juveniles and adults at 2 days post injury when compared to shams. When comparing the extent of cell proliferation between juveniles and adults following TBI, the absolute number of cells generated in the subgranular zone (SGZ) was far greater in the juveniles. Moreover, the percentage of newly generated cells in the SGZ that differentiated into neurons was nearly two times higher in the juveniles as compared to adults. Conversely, more glial differentiation was observed in the DG of adult rats. These findings provide compelling evidence that age-related differences in the neurogenic response to injury may underlie the differences observed in cognitive recovery in juvenile mammals as compared to adults following TBI. PMID- 15665607 TI - The effect of poloxamer-188 on neuronal cell recovery from mechanical injury. AB - Neuronal injury resulting from mechanical deformation is poorly characterized at the cellular level. The immediate structural consequences of the mechanical loading lead to a variety of inter- and intra-cellular signaling events that interact on multiple time and length scales. Thus, it is often difficult to establish cause-and-effect relationships such that appropriate treatment strategies can be devised. In this report, we showed that treating mechanically injured neuronal cells with an agent that promotes the resealing of disrupted plasma membranes rescues them from death at 24 h post-injury. A new in vitro model was developed to allow uniform mechanical loading conditions with precisely controlled magnitude and onset rate of loading. Injury severity increased monotonically with increasing peak shear stress and was strongly dependent on the rate of loading as assessed with the MTT cell viability assay, 24 h post-injury. Mechanical injury produced an immediate disruption of membrane integrity as indicated by a rapid and transient release of LDH. For the most severe injury, cell viability decreased approximately 40% with mechanical trauma compared to sham controls. Treatment of cells with Poloxamer 188 at 15 min post-injury restored long-term viability to control values. These data establish membrane integrity as a novel therapeutic target in the treatment of neuronal injury. PMID- 15665606 TI - The neurosteroids progesterone and allopregnanolone reduce cell death, gliosis, and functional deficits after traumatic brain injury in rats. AB - This report compares the effects of progesterone and its metabolite, allopregnanolone, on the early injury cascade (apoptosis) and long-term functional deficits after TBI. Progesterone (16 mg/kg) or allopregnanolone (4, 8, or 16 mg/kg) were injected at 1 h, 6 h, and then for 5 consecutive days after bilateral contusions of the frontal cortex in adult male rats. Within one day after injury, progesterone and allopregnanolone reduced both the expression of pro-apoptotic proteins caspase-3 and Bax, and apoptotic DNA fragmentation. Progesterone and allopregnanolone also reduced the size of glial fibrillary acid protein (GFAP)-positive astrocytes at the lesion site 24 h after injury. Compared to sham-operated controls at 19 days after injury, injured rats given either progesterone or any of three doses of allopregnanolone had equivalent numbers of ChAT-positive cells in the nucleus basalis magnocellularis. At 19 days post injury, rats given progesterone or allopregnanolone (8 mg/kg) showed improved performance in a spatial learning task compared to injured rats given only the vehicle. These results provide evidence of the anti-apoptotic and anti astrogliotic effects of progesterone and allopregnanolone and help to explain why better cognitive performance is observed after injury when animals are given either neurosteroid. PMID- 15665608 TI - The cervico-ocular reflex is increased in whiplash injury patients. AB - Whiplash-associated disorders (WAD) are a major problem in the Western world, which put a formidable financial burden on modern society and which evoke an emerging debate on the true nature of their origin. To date there is no generally accepted test that allows us to diagnose WAD objectively. Because whiplash injury causes dysfunction of proprioception in the neck, we investigated the characteristics of the cervico-ocular reflex (COR) of presumptive WAD patients. These patients and age-matched healthy controls were rotated at different stimulus peak velocities in the dark while their head was fixed in space. The gain values of the COR were significantly increased in the patient population at a wide range of stimulus peak velocities with maximum difference at the lower frequencies (p = 0.037, ANOVA). Hence, although larger numbers of patients should be measured, the COR gain appears to be a parameter that may permit an objective diagnosis of WAD. PMID- 15665610 TI - Voluntary wheel running improves recovery from a moderate spinal cord injury. AB - Recently, locomotor training has been shown to improve overground locomotion in patients with spinal cord injury (SCI). This has triggered renewed interest in the role of exercise in rehabilitation after SCI. However, there are no mouse models for voluntary exercise and recovery of function following SCI. Here, we report voluntary wheel running improves recovery from a SCI in mice. C57Bl/10 female mice received a 60-kdyne T9 contusion injury with an IH impactor after 3 weeks of voluntary wheel running or 3 weeks of standard single housing conditions. Following a 7-day recovery period, running mice were returned to their running wheels. Weekly open-field behavior measured locomotor recovery using the Basso, Beattie and Bresnahan (BBB) locomotor rating scale and the Basso Mouse Scale (BMS) locomotor rating scale, a scale recently developed specifically for mice. Initial experiments using standard rung wheels show that wheel running impaired recovery, but subsequent experiments using a modified flat-surface wheel show improved recovery with exercise. By 14 days post SCI, the modified flat surface running group had significantly higher BBB and BMS scores than the sedentary group. A repeated measures ANOVA shows locomotor recovery of modified flat-surface running mice was significantly improved compared to sedentary animals (p < 0.05). Locomotor assessment using a ladder beam task also shows a significant improvement in the modified flat-surface runners (p < 0.05). Finally, fibronectin staining shows no significant difference in lesion size between the two groups. These data represent the first mouse model showing voluntary exercise improves recovery after SCI. PMID- 15665609 TI - Alginate encapsulated BDNF-producing fibroblast grafts permit recovery of function after spinal cord injury in the absence of immune suppression. AB - Encapsulation of cells has the potential to provide a protective barrier against host immune cell interactions after grafting. Previously we have shown that alginate encapsulated BDNF-producing fibroblasts (Fb/BDNF) survived for one month in culture, made bioactive neurotrophins, survived transplantation into the injured spinal cord in the absence of immune suppression, and provided a permissive environment for host axon growth. We extend these studies by examining the effects of grafting encapsulated Fb/BDNF into a subtotal cervical hemisection on recovery of forelimb and hindlimb function and axonal growth in the absence of immune suppression. Grafting of encapsulated Fb/BDNF resulted in partial recovery of forelimb usage in a test of vertical exploration and of hindlimb function while crossing a horizontal rope. Recovery was significantly greater compared to animals that received unencapsulated Fb/BDNF without immune suppression, but similar to that of immune suppressed animals receiving unencapsulated Fb/BDNF. Immunocytochemical examination revealed neurofilament (RT-97), 5-HT, CGRP and GAP 43 containing axons surrounding encapsulated Fb/BDNF within the injury site, indicating axonal growth. BDA labeling however showed no evidence of regeneration of rubrospinal axons in recipients of encapsulated Fb/BDNF, presumably because the amounts of BDNF available from the encapsulated grafts are substantially less than those provided by the much larger numbers of Fb/BDNF grafted in a gelfoam matrix in the presence of immune suppression. These results suggest that plasticity elicited by the BDNF released from the encapsulated cells contributed to reorganization that led to behavioral recovery in these animals and that the behavioral recovery could proceed in the absence of rubrospinal tract regeneration. Alginate encapsulation is therefore a feasible strategy for delivery of therapeutic products produced by non-autologous engineered fibroblasts and provides an environment suitable for recovery of lost function in the injured spinal cord. PMID- 15665611 TI - Changes in distribution of serotonin induced by spinal injury in larval lampreys: evidence from immunohistochemistry and HPLC. AB - Larval lampreys are known to successfully recover normal behavior following spinal cord injury. More recently, we showed temperature can influence functional recovery, with colder temperatures more likely producing behavioral abnormality despite the cold being the animals' normal temperature. Here we analyze the differences associated with temperature effects. We examine serotonergic expression along the spinal cord following midbody lesions considering time from injury, temperature during recovery and fiber location. We also examine the relationship between regeneration and locomotion, insofar as the pattern of muscle potentials during unrestrained swimming is normal or abnormal. At 26 weeks after spinal cord injury in all groups of animals, immunohistochemistry and HPLC for serotonin and serotonin expression above and below the lesion can be significantly changed in all regional sources of serotonin independent of the temperature animals recover from their injuries. Animals from warmer tanks recover serotonin expression in the segment immediately caudal to the lesion site with little further away from the lesion; animals from the cold room aquaria have significantly less recovery of expression caudal to the lesion and none further away. There was no apparent relationship between the distribution of serotonin and recovery. The changes suggest that some intraspinal reorganization has occurred. We propose a relationship between the observed results and functional recovery, but it remains conjectural. The fact that some animals recover normal function suggests plasticity must occur in animals successful in recovering normal function. Thus, the lamprey can be used as a model system to study the adaptive changes that permit or prevent functional recovery. PMID- 15665614 TI - Determination of antibody immunoreactive fraction. PMID- 15665612 TI - Astroglial nitration after postnatal excitotoxic damage: correlation with nitric oxide sources, cytoskeletal, apoptotic and antioxidant proteins. AB - Oxygen free radicals and nitric oxide (NO) participate in the pathogenesis of acute central nervous system (CNS) injury by forming peroxynitrite, which promotes oxidative damage and tyrosine nitration. Neuronal nitration is associated with cell death, but little is known of the characteristics and cell fate of nitrated astrocytes. In this study, we have used a postnatal excitotoxic lesion model (intracortical NMDA injection) and our aims were (i) to evaluate the temporal and spatial pattern of astroglial nitration in correlation with the neuropathological process and the sources of NO; and (ii) to establish, if any, the correlation among astrocyte nitration and other events such as expression of cytoskeletal proteins, antioxidant enzymes, and cell death markers to cope with nitration and/or undergo cell death. Our results show that after postnatal excitotoxic damage two distinct waves of nitration were observed in relation to astrocytes. At 24 h post-lesion, early-nitrated astrocytes were found within the neurodegenerating area, coinciding with the time of maximal cell death. These early-nitrated astrocytes are highly ramified protoplasmic cells, showing diffuse glial fibrillary acidic protein (GFAP) content and expressing inducible NOS. At later time-points, when astrogliosis is morphologically evident, nitrated hypertrophied reactive astrocytes are observed in the penumbra and the neurodegenerated area, displaying increased expression of GFAP and vimentin cytoskeletal proteins and of metallothionein I-II and Cu/Zn superoxide dismutase antioxidant proteins. Moreover, despite revealing activated caspase-3, they do not show TUNEL labeling. In summary, we show that nitrated astrocytes in vivo constitute a subpopulation of highly reactive astrocytes which display high resistance towards oxidative stress induced cell death. PMID- 15665615 TI - The immunoreactivity of radiolabeled antibodies--its impact on tumor targeting and strategies for preservation. PMID- 15665616 TI - The evolution of accelerated, partial breast irradiation as a potential treatment option for women with newly diagnosed breast cancer considering breast conservation. AB - Breast conservation therapy (BCT) is a safe, effective alternative to mastectomy for many women with newly diagnosed breast cancer. This approach involves local excision of the malignancy with tumor-free margins, followed by 5-7 weeks of external beam whole breast (WB) radiotherapy (XRT) to minimize the risk of an in breast tumor recurrence (IBTR). Though clearly beneficial, the extended course of almost daily postoperative radiotherapy interrupts normal activities and lengthens care. Additional options are now available that shorten the radiotherapy treatment time to 1-5 days (accelerated) and focus an increased dose of radiation on just the breast tissue around the excision cavity (partial breast). Recent trials with accelerated, partial breast irradiation (APBI) have shown promise as a potential replacement to the longer, whole breast treatments for select women with early-stage breast cancer. Current APBI approaches include interstitial brachytherapy, intracavitary (balloon) brachytherapy, and accelerated external beam (3-D conformal) radiotherapy, all of which normally complete treatment over 5 days, while intraoperative radiotherapy (IORT) condenses the entire treatment into a single dose delivered immediately after tumor excision. Each approach has benefits and limitations. This study covers over 2 decades of clinical trials exploring APBI, discusses treatment variables that appear necessary for successful implementation of this new form of radiotherapy, compares and contrasts the various APBI approaches, and summarizes current and planned randomized trials that will shape if and how APBI is introduced into routine clinical care. Some of the more important outcome variables from these trials will be local toxicity, local and regional recurrence, and overall survival. If APBI options are ultimately demonstrated to be as safe and effective as current whole breast radiotherapy approaches, breast conservation may become an even more appealing choice, and the overall impact of treatment may be further reduced for certain women with newly diagnosed breast cancer. PMID- 15665617 TI - Determination of immunoreactive fraction of radiolabeled monoclonal antibodies: what is an appropriate method? AB - Determination of the immunoreactive fraction (IF) of radiolabeled monoclonal antibodies (MAb) is essential to the understanding of the effects of radiolabeling and subsequent target-specific tumor localization. There has been generally no accepted method of determining the IF of MAbs. The conventional method is based on a radioimmunoassay technique in which the fraction of radiolabeled MAb bound to antigen under conditions of "antigen excess" is determined. Lindmo et al. introduced a modified method in which the IF is determined by extrapolation to conditions representing "infinite antigen excess." Although the Lindmo method, in principle, is insensitive to experimental parameters, it does not always provide a reliable estimate of IF. We, therefore, evaluated an alternate method in which percent cell bound fraction is measured under conditions of fixed antigen concentration and various dilutions of radiolabeled MAb. We developed a mathematical equation to estimate immunoreactivity. J591 MAb specific for prostate-specific membrane antigen was radiolabeled with (111)In, (90)Y and (177)Lu to specific activities of 1-20 mCi/mg. We compared the effect of several experimental conditions on the determination of IF using all three different methods. The Lindmo method requires careful optimization of experimental conditions for each radiolabeled MAb. The alternate method, based on a fixed antigen concentration, appears to be practical and may provide a more reliable measure of immunoreactivity. PMID- 15665618 TI - The 17th International IRIST Congress Proceedings. AB - The 17th IRIST (International Research Group on Immunoscitigraphy and Therapy) meeting was held July 27-29, 2004, at the Ghent University Hospital, Ghent, Belgium. Cochairs Christophe Van de Wiele and Alberto Signore brought together basic researchers, clinicians, radiopharmacists, and nuclear medicine specialists to discuss progress in single photon emission computed tomography (SPECT) and positron emission tomography (PET), using radiolabeled antibodies and peptides for imaging biologic properties related to the effectiveness of available treatment options and for treatment of cancer patients. The format of the conference was the presentation of research that focused on the basic and translational biology of cancer and on current state-of-the-art radiopharmacy and related SPECT and PET imaging techniques for the purposes described above. This report summarizes the 17th International IRIST Congress. PMID- 15665619 TI - Safety, pharmacokinetics, immunogenicity, and biodistribution of (186)Re-labeled humanized monoclonal antibody BIWA 4 (Bivatuzumab) in patients with early-stage breast cancer. AB - The aim of this prospective study was to evaluate the safety, pharmacokinetics, immunogenicity, and biodistribution of (186)Re-labeled humanized anti-CD44v6 monoclonal antibody (MAb( BIWA 4 (Bivatuzumab( in 9 patients with early-stage breast cancer. Radioimmunoscintigraphy (RIS( was performed within 1, 24, and 72 hours after administration. BIWA 4 concentration in plasma (ELISA and radioactivity measurements( and the development of human antihuman antibody (HAHA( responses was determined. The biodistribution of (186)Re-BIWA 4 was determined by radioactivity measurements in tumor and normal tissue biopsies obtained during surgery 1 week after administration. Administration of (186)Re BIWA 4 was well tolerated by all patients and no HAHA responses were observed. The mean t(1/2) in plasma of BIWA 4 (ELISA( was 81 hours (range, 67-97(, whereas the mean radioactivity t(1/2) tended to be longer, at 105 hours (range, 90-114(. RIS unmistakably showed the tumor in 3 patients. Less clear identifications were established in 3 additional patients. In 2 patients, the tumor was wrongly identified in the contralateral breast. Median tumor CD44v6 expression, as determined by immunohistochemistry, was 70% (range, 10-90%). Mean tumor uptake was 2.96% ID/kg (range, 0.92-6.27(, with no apparent correlation with either tumor CD44v6 expression, tumor-cell cellularity, or tumor diameter. Tumor-to nontumor ratios were unfavorable for blood, bone marrow, mammary gland tissue, and skin. CONCLUSIONS: The (186)Re-labeled humanized MAb BIWA 4 can safely be administered to patients with early-stage breast cancer. Tumorto- nontumor ratios were unfavorable, with no apparent correlation with CD44v6 expression, tumor-cell cellularity, or tumor diameter. BIWA 4, therefore, appears to have limitations as a vehicle for radioimmunotherapy in patients with breast cancer. PMID- 15665620 TI - Tumor-infiltrating lymphocytes and interleukin-2: dose and schedules of administration in the treatment of metastatic cancer. AB - PURPOSE: The potential for therapeutic use of tumor-infiltrating lymphocytes (TIL), as adoptive cellular therapy has been touted for many years with some encouraging reports in patients with metastatic melanoma. MATERIALS AND METHODS: We previously described methodologies for TIL production and phenotypic characterization of TIL generated in our laboratory between 1991 and 1995 in semipermeable bags and between 1996 and 2000 in bioreactors. Patients treated in the earlier era were to have received a hybrid bolus and a 12-hour continuous infusion of interleukin (IL)-2 (total, 48 MIU), while in the latter era 4 days of interferon- alpha preceded the TIL and IL-2; which was given by a hybrid schedule that included bolus and 72- hour continuous IL-2 (total, 96 MIU). There were 55 patients, including 23 patients with melanoma, 9 patients with renal cell carcinoma, and 8 patients with colorectal cancer. There was only 1 objective tumor response, which was noted in a patient with renal cell carcinoma. The 55 patients who received these products were grouped in cohorts by treatment era, quantity of TIL received, amount of IL-2 intended, and different combinations of TIL and IL-2. RESULTS: There was no difference in survival by production method (treatment era), or amount of IL-2 given with TIL, but 33 patients who received an intermediate or higher dose of TIL (mean = 54.4 x 10(9)) had a median survival of 11.8 months, compared to 6.4 months for 22 patients who received 1 low-dose TIL (mean = 6.48 x 10(9)) (p = 0.059, log rank test). The objective response rate in this heterogeneous group of patients was not encouraging. The data suggest there may be a dose/benefit relationship between the total number of TIL infused and survival. PMID- 15665621 TI - Comparison of radiation dose estimation for myeloablative radioimmunotherapy for relapsed or recurrent mantle cell lymphoma using (131)I tositumomab to that of other types of non-Hodgkin's lymphoma. AB - Patients with relapsed or refractory mantle cell lymphoma (MCL) demonstrate poor survival after standard treatment. Myeloablative radioimmunotherapy (RIT) using (131)I tositumomab (anti-CD20) has the ability to deliver specific radiation absorbed dose to antigen-bearing tumor. We reviewed normal organ radiation- absorbed doses in MCL patients. METHODS: Records of patients with MCL (n =25), who received myeloablative RIT between January 1996 and December 2003 were reviewed. Individual patient radiation dosimetry was performed on all patients after a trace-labeled infusion of (131)I tositumomab (mean = 348 MBq), to calculate the required amount of radioactivity for therapy, based on medical internal radiation dose (MIRD) schema. RESULTS: Mean organ residence times (hour) corrected for computed tomography (CT) derived organ volumes for MCL, were as follows: Lungs: 9.0; Liver: 12.4; Kidneys: 1.7; Spleen: 2.17; Whole Body: 62.4 and mean radiation absorbed doses mGy/Mbq were: Lungs: 1.2; Liver: 1.1; Kidneys: 0.85; Spleen: 1.7; Whole Body: 0.21. This is similar to patients with other non Hodgkin's lymphoma (NHL). Patients received a mean activity of 21 GBq of (131)I (range, 11.5-41.4) for therapy estimated to deliver 25 Gy to the normal organ receiving the highest radiation-absorbed dose. CONCLUSION: Myeloablative RIT using (131)I tositumomab results in normal organ radiation-absorbed doses similar to those in patients with other non-Hodgkin's lymphoma, and is suitable for treating patients with relapsed or refractory MCL. PMID- 15665623 TI - Continuous infusion interleukin-2 and antihistamines in melanoma: a retrospective review showing activity of this combination. AB - A recent randomized trial suggests that there may be an advantage in terms of survival with the combination of histamine and subcutaneous interleukin-2 (IL-2), compared to IL-2 alone. It has been postulated, then, that antihistamines may actually be antagonistic to IL-2 and, therefore, interfere with its antitumor activity. Because antihistamines such as cimetidine and ranitidine are commonly used as prophylaxis against gastrointestinal toxicity commonly seen with IL-2, and, because antihistamines may increase natural killer cell activity, it is reasonable to examine the response rate for this combination. An OVID Medline literature search between 1985 and 2003 was done. Continuous infusion (CIV) interleukin- 2 was used as the reference therapy because of the relatively constant IL-2 levels generated by this approach. Included studies were those in which either cimetidine, ranitidine, or famotidine were regularly scheduled and administered concurrently with IL-2, typically for gastrointestinal ulcer prophylaxis. Six (6) studies were identified. A total of 21 patients responded to therapy. Total response rate was 11%, with a 95% Confidence Interval: 7-17%. Four (4) complete responses were noted. Complete response rate was 2%, with a 95% Confidence Interval: 1-6%. These response rates are consistent with previously noted IL-2 response rates. In this retrospective review of CIV IL-2 and antihistamines, this combination appears to be active in melanoma. There appears to be no deleterious effect of routine antihistamine on the CIV IL-2 response rate. PMID- 15665622 TI - The efficacy of tamoxifen in estrogen receptor-positive breast cancer cells is enhanced by a medical nutriment. AB - Avemar, a fermented wheat germ extract, has been applied in the supplementary therapy of human cancers. Because tamoxifen is commonly used in the therapy of ER+ breast cancer, in this study the combined effect of tamoxifen and Avemar treatment was investigated on MCF-7 breast cancer cells, in order to detect a possible agonistic or antagonistic action. Cytotoxicity was measured by MTT assay, the percentage of mitoses and apoptotic cells was determined morphologically, apoptosis and S-phase was measured by flow cytometry, and estrogen-receptor activity was determined by semiquantitative measurement of the estrogen-responsive pS2 gene mRNA production. Tamoxifen (1 nM) alone had no effect on the percentage of the apoptotic cell fraction and significantly reduced the percentage of the S-phase, compared to untreated cells. Avemar (625 microg/mL) significantly increased apoptosis after 48 hours of treatment. Tamoxifen together with Avemar significantly increased apoptosis already 24 hours after starting treatment but had only a slight (not significant) effect on mitosis and S-phase. Estrogen-receptor activity of MCF-7 cells was enhanced by Avemar, decreased by tamoxifen, and was further decreased by combined tamoxifen and Avemar treatment. As apoptosis increased when Avemar was added to tamoxifen treatment, the use of supplementary therapy with Avemar in the case of ER+ breast tumors may enhance the therapeutic effects of tamoxifen. PMID- 15665624 TI - Individualized synthetic peptide vaccines with GM-CSF in locally advanced melanoma patients. AB - We report on 10 patients with resected American Joint Committee on Cancer (AJCC)stage IIA-IIIC melanoma receiving individualized adjuvant peptide vaccinations derived from the melanosomal antigens MelanA/MART1, gp100 and tyrosinase, according to patient tumor associated HLA restricted antigen expression, in combination with granulocyte-macrophage colony-stimulating factor (GM-CSF). Except for 1 patient, all patients had received systemic pretreatment with immunotherapy (n = 8), chemoimmunotherapy (n = 1), chemotherapy (n = 1), or cefalectin therapy (n = 1). Upon prior therapy, 7 of 10 patients had progressed with subcutaneous/cutaneous (n = 2), lymph node (n = 3), or subcutaneous/cutaneous and lymph node (n = 2)metastases, which were subsequently resected prior to vaccination. After a mean of 6.5 vaccination cycles, progression-free survival was 6 months (median, range 2-10). Five patients were relapse-free for 1+ up to 21+ months, 3 patients developed a solitary cutaneous metastasis, and 2 patients developed multiple metastases during vaccination. Overall, vaccine treatment was well tolerated, with no severe side-effects. Eight of 10 patients developed local delayed type hypersensitivity (DTH)reactions to synthetic peptides after the first or second injection. In 2 patients, transient fever, nausea, diarrhea, and muscle pain of National Cancer Institute (NCI)Grade I occurred. In summary, individualized synthetic peptide vaccination, combined with GM-CSF, was feasible and warrants further clinical investigation. PMID- 15665625 TI - Characterization of cytokine-encapsulated controlled-release microsphere adjuvants. AB - Controlled-release, injectable polymer microspheres provide a clinically feasible alternative to gene-modification for the local, sustained delivery of cytokines to tumors for cancer immunotherapy. Long-term release kinetics, bioactivity profiles, and stability of interleukin-2 (IL-2), interleukin-12 (IL-12), and granulocyte- macrophage colony-stimulating factor (GM-CSF)-encapsulated microspheres prepared by phase inversion nanoencapsulation (PIN) were evaluated. While all formulations released physiologically relevant quantities of cytokine for up to 30 days, the individual release kinetics were different. Recovery of specific activity after encapsulation was 40%, 60%, and 90%-that of pre encapsulation levels for IL-2, GM-CSF and IL-12, respectively. Upon storage, the IL-12 microspheres rapidly lost activity, whereas IL-2 and GM-CSF microspheres remained stable for at least 9 weeks. These studies demonstrate that biochemical properties of microsphere formulations vary depending on the cytokine, and rigorous characterization of formulations is a prerequisite to in vivo testing. PMID- 15665626 TI - High-dose continuous infusion plus pulse interleukin-2 and famotidine in melanoma. AB - High-dose, continuous infusion interleukin-2 (IL-2) regimens generate greater Lymphokine Activated Killer cell (LAK) cytotoxicity in vitro and a higher rebound lymphocytosis in vivo than do bolus IL-2 regimens. Lymphocytes initially activated by continuous infusion IL-2 then subsequently pulsed with IL-2 have increased cytotoxicity against cancer cells. Famotidine may enhance the lysis of tumors by cytotoxic lymphocytes. Fourteen patients with melanoma were treated with famotidine 20 mg intravenously twice per day and continuous infusion IL-2 (18 MIU/sq m/24 hours) for 72 hours, followed by a 24-hour rest, then IL-2 18 MIU/sq m over 15-30 minutes for 1 dose (12 patients) or daily for 3 doses (2 patients). Most common toxicities were fever, nausea/emesis, hypophosphatemia, hypomagnesemia, and rigors. Nine partial responses (64% response rate; 95% Confidence Interval: 39%-84%) have been seen. Median survival has not been reached at greater than 10 months. Two patients responding to therapy showed an increase in detectable CD 56(+) cells in serial subcutaneous or lymph node biopsies, while 1 patient undergoing progression of disease had no such infiltrate. High-dose, 72-hour continuous infusion plus pulse interleukin-2 with famotidine has activity in melanoma. CD 56(+) cells may play a role in responding patients. PMID- 15665627 TI - Biodistribution and dosimetry of (99m)Tc-depreotide (P829) in patients suffering from breast carcinoma. AB - OBJECTIVE: This paper reports on the biodistribution and dosimetry of (99m)Tc depreotide in patients. METHODS: Whole body planar images were acquired 30 minutes, 1, 2, 4, 9, and 24 hours after intravenous injection of 555-740MBq (99m)Tc-depreotide in 5 breast cancer patients. Urine was collected up to 24 hours after injection, allowing for a calculation of renal clearance and an interpretation of whole body clearance. Time activity curves were generated for the thyroid, lungs, liver, spleen, kidneys, colon, thoracic vertebrae/sternum, and whole body by fitting the organ-specific geometric mean counts, obtained from regions of interest (ROIs). The Medical Internal Radiation Dose (MIRD) formulation was applied to calculate the absorbed radiation dose for various organs. RESULTS: The whole body images show most of the activity distributed in the liver, spleen, and kidneys. Nearly all excretion of activity occurred by the renal system, and hepatobiliary excretion was negligible. Elimination of administered activity occurred predominantly through physical decay. The mean cumulative measured urinary excretion at 24 hours postinjection was 14.0% (standard deviation; 11.8%) of the administered activity. The highest absorbed dose was received by the kidneys, thyroid, and spleen. The average effective dose was estimated to be 1.15E-02mSv/MBq (standard deviation; 1.41E-03mSv/MBq). CONCLUSION: The biodistribution of (99m)Tc-depreotide demonstrated low lung and myocardial uptake allowing early imaging of the supradiaphragmatic region and this with a dosimetry favorable for clinical whole body and single photon emission computed tomography (SPECT) imaging. PMID- 15665628 TI - Prospective evaluation of factors influencing success rates of sentinel node biopsy in 814 breast cancer patients. AB - OBJECTIVE: This prospective multicenter study was performed to assess the reliability of sentinel lymph node (SLN) biopsy in breast cancer and to analyze factors potentially influencing success rates. METHODS: In 21 departments, SLN biopsy and consecutive axillary lymph node dissection were performed in 814 breast cancer patients. The 80 surgeons involved were free in the choice of lymphography technique. The detection rate and the sensitivity, as well as the impact of lymphography technique, patient selection, technical procedure and learning curves, were evaluated. RESULTS: The blue dye technique was used in 137 patients, radiocolloid in 169 patients, and combined blue dye/radiocolloid in 508 patients. The identification rate for the sentinel node was 83.9% for the entire group and showed a significant dependence on the lymphography technique (blue dye, 71.6%; radiocolloid, 78.8%; combined blue dye and radiocolloid, 89.6%). The overall sensitivity in detecting lymph node metastases was 91.3%. Immunostaining for cytoceratine revealed micrometastases in 19 (5.1%) of 374 patients in whom H/E staining was negative. The combined subdermal/peritumoral injection of the colloid showed a significantly higher identification rate than subdermal or peritumoral injection alone (96.8%, 84.6%, 78.6%; p < 0.001). There was also a significant higher detection rate in cases of SLN biopsy performed prior to lumpectomy, compared to SLN biopsy following lumpectomy (94.7% versus 82.8%; p < 0.001). Furthermore, there was a close correlation between the number of performed examinations and the detection rate. CONCLUSION: SLN mapping predicts the axillary lymph node status accurately. Learning curves and several technical features influence the detection rate significantly. However, the false negative rate was independent of experience and injection technique. PMID- 15665629 TI - The experts speak. Studying the potential of tenofovir to prevent sexual transmission of HIV: first steps. Interview by Vicki Glaser. AB - Melanie Thompson, M.D., is Founder and Principal Investigator of the AIDS Research Consortium of Atlanta (ARCA), one of the first non-profit community based HIV/AIDS research centers in the United States and the primary resource for HIV clinical research in Georgia. Dr. Thompson is also a private practitioner in Atlanta and serves on the International AIDS Society- USA's Antiretroviral Guidelines Panel. She is the past chair of the National Institutes of Health Office of AIDS Research Therapeutics Research Working Group. Dr. Thompson has served as ARCA's principal investigator for more than 200 studies involving more than 12,000 patients. To date, ARCA's studies have contributed to the FDA approval of 21 treatments for HIV and its complications. PMID- 15665630 TI - Accessibility and longevity of Internet citations in a clinical AIDS journal. AB - Little is known about the accessibility and longevity of Internet references in medical and scientific journals. This is particularly problematic in the ever changing field of HIV/AIDS. We examined all issues of AIDS Patient Care and STDs between 2001 and 2004 to test the use and accessibility of Internet citations during this period. The results were compared to findings from a similar evaluation by Hester and colleagues in 2004 of oncology journals. We found that Internet referencing increased from a mean of 0.7% (range, 0.0%-2.1%) of all citations in 2001 to a mean of 3.5% (range, 0.0%-11.2%) in 2004. As the period of time increased from an article's publication, the proportion of inactive Internet references increased (21.3% [1 year] to 41.7% [4 years]). These findings demonstrate the need to adapt new citation policies to facilitate the accessibility of referenced Internet information. PMID- 15665631 TI - Abacavir compared to protease inhibitors as part of HAART regimens for treatment of HIV infection: patient satisfaction and implications for adherence. AB - The purpose of this study was to compare treatment satisfaction with triple nucleoside reverse transcriptase inhibitor (NRTI) highly active antiretroviral treatment (HAART) regimens including abacavir (ABC) to HAART regimens that include protease inhibitors (PIs) and to estimate the relationship between patient satisfaction and adherence to HAART. Three open-label clinical trials comparing ABC-including HAART regimens with PI-including HAART regimens were completed, two with patients previously untreated with antiretroviral therapy and one with patients successfully treated with PI-including HAART regimens. The HIV Treatment Satisfaction Questionnaire (HIVTSQ) was completed at several time points during each trial. Levels of patient satisfaction with the ABC and PI regimens were compared for all three trials. The correlation between adherence and patient satisfaction scores was measured using data from an adherence questionnaire in one of the studies. In all three clinical trials, patient satisfaction scores were significantly higher with an ABC-including triple NRTI HAART regimen than with a PI-including HAART regimen. The difference was apparent by week 4 of the trial and was maintained throughout the trial time period. Inspection of the item responses in the patient satisfaction questionnaire indicated that treatment convenience, flexibility, impact on lifestyle, and side effects were key factors in the difference in satisfaction between the treatment groups. In addition, patient satisfaction was shown to be significantly correlated with adherence defined as taking 95% or more of prescribed doses. Greater satisfaction was reported by patients given an ABC-including HAART regimen than those given a PI-including HAART regimen. Patient satisfaction may be an indicator for better treatment adherence. PMID- 15665632 TI - Body habitus in a cohort of HIV-seropositive and HIV-seronegative injection drug users. AB - We determined anthropometric measurements (including height, weight, circumferences, and skinfolds) and self-reported symptoms related to body habitus changes in 324 HIV-seropositive and HIV-seronegative inner city injection drug users (IDUs) who participated in a substudy from the ALIVE (AIDS Linked to Intravenous Experiences) cohort. Participants who reported lipoatrophy in body parts had consistently lower anthropometric measurements and those reporting adiposity had correspondingly higher anthropometric measurements than participants who did not report these changes. Peripheral lipoatrophy was more common among all HIV-seropositive than HIV-seronegative participants, however, it was not associated with highly active antiretroviral therapy (HAART) (39% HIV seronegatives; 58% HIV-seropositive not receiving HIV treatment [No Tx]; 49% HAART, p = 0.04). Central adiposity was more common among HAART (52%) than No Tx (26.6%) and HIV-seronegative (42%) participants (p = 0.001). However, waist circumference, while somewhat higher among HAART than No Tx participants, did not differ significantly from HIV-seronegative participants (85.2 cm HIV seronegatives; 83.3 cm No Tx; 85.8 cm HAART). A large proportion of those who reported peripheral lipoatrophy also reported central lipoatrophy (76.9% HIV seronegatives; 69.6% No Tx; 66.2% HAART). A large proportion of those who reported central adiposity also reported adiposity of the peripheral sites (88.1% HIV seronegatives; 66.7% No Tx; 74.3% HAART). The combination of lipoatrophy and adiposity was associated with HAART treatment (6% HIV-seronegatives; 3% No Tx; 16% HAART, p = 0.002), but may be driven by the association with adiposity. These data suggest validity of self-reports for body habitus changes among injection drug users. PMID- 15665633 TI - Integration of motivational interviewing and cognitive behavioral therapy to improve HIV medication adherence and reduce substance use among HIV-positive men and women: results of a pilot project. AB - Directly addressing HIV medication adherence and substance use together is ideal in light of the research on the complex connections between these two behaviors. This paper describes the development of a pilot program in which a combined motivational interviewing and cognitive behavioral therapy intervention was used to increase adherence to highly active antiretroviral therapy (HAART) and reduce substance use in HIV-infected adults. The main goals of the pilot study were (1) to confirm the ability to recruit HIV-positive substance users taking HAART; (2) to demonstrate the ability to retain participants over the course of an eight session intervention; and (3) to examine changes in substance use and HAART adherence from pretreatment to posttreatment. Twelve HIV-positive adults with a substance use disorder participated in an 8-week intervention, which consisted of weekly individual sessions with a trained therapist. Results showed that despite the small sample size, there was a significant reduction in substance use from pretreatment to posttreatment. No statistically significant differences were found for changes in HIV medication adherence, but the trends suggest the potential for positive results with a larger sample. Retention for all eight sessions of the treatment was 73.3%. Eighty percent completed the 3-month follow up assessment. Participants reported high therapeutic alliance reflecting comfort with their therapists and in the treatment. Overall, the pilot demonstrated feasibility and acceptability of the treatment. Furthermore, results suggest that this may, in fact, be an effective intervention to reduce substance use and improve HIV medication adherence. PMID- 15665634 TI - STD and HIV counseling practices of British Columbia primary care physicians. AB - This study evaluated British Columbian primary care physicians' use of sexually transmitted disease (STD) and HIV counseling guidelines. The authors sought to assess barriers to integrated HIV and STD testing and counseling, with the objective of developing and revising practice guidelines to meet the needs of primary care physicians in diverse settings. The BC College of Physicians and Surgeons membership of general practice/family medicine practitioners was stratified by practitioner location; 1200 physicians were randomly selected to receive a mailed self-administered questionnaire. Four hundred seven completed questionnaires were returned. Female physicians reported better adherence to practice guidelines on many items than their male counterparts. A minority of primary care physicians routinely integrated HIV and STD testing and counseling. Physicians in nonurban areas were more likely to report that they lacked sufficient information on HIV and STD risk and prevention. Although male primary care physicians in British Columbia were more likely to provide care for people with HIV infection, female physicians were more likely to undertake ongoing screening for sexual risk behavior and sexually transmitted infections (STIs) among their HIV-positive patients. Clinical practice guidelines are theoretically equally available to all physicians, but they are not universally used. Physicians may not receive adequate training in sexual health during their medical education; continuing education opportunities may be limited to physicians outside of urban centres. Accessible and relevant continuing medical education in novel formats are needed to address the gap between ideal and actual practice in HIV and STD prevention, treatment, and care in Canada. PMID- 15665635 TI - Risk factors for sexually transmitted disease among rural-to-urban migrants in China: implications for HIV/sexually transmitted disease prevention. AB - The objective of the study was to identify risk factors associated with sexually transmitted diseases (STDs) among rural-to-urban migrants in Beijing in 2002. Migrants with STDs consisted of 432 migrants who sought STD care in two public STD clinics. Migrants without STDs included 892 migrants recruited from 10 occupational clusters. Multiple logistic regression was used for data analysis. Compared to migrants without STDs, migrants with STDs were more likely to report having engaged in commercial sex (selling or buying sex) (odds ratio [OR] = 2.70, 95% confidence interval [CI]: 1.71-4.25), multiple sex partners in the previous month (OR = 6.50, 95% CI: 3.73-11.32) and higher perceived HIV-related stigma (OR = 1.89, 95% CI: 1.30-2.75). Being a migrant with an STD was also associated with female gender (OR = 4.10, 95% CI: 2.89-5.82), higher education (OR = 2.92, 95% CI: 1.40-6.06), and higher monthly salary (OR = 1.68. 95% CI: 1.23-2.29). Migrants with STDs visited their hometowns more frequently and had more stable jobs than migrants without STDs. Approximately 10% of the migrants with STDs and 7.7% of the migrants without STDs always used condoms. This study suggests that among migrants, acquisition of an STD is associated with higher participation in risk behaviors as would be expected, but also with higher perceived stigma, education, stable jobs, salary, and with female gender. Appropriate behavioral intervention programs are advocated to reduce the risk and stigma among the special population. PMID- 15665636 TI - RESIST-1 results released. PMID- 15665637 TI - Internet helps prostitutes. PMID- 15665638 TI - High prevalence of HTLV-I and HTLV-II among blood donors in Argentina: a South American health concern. AB - After an HTLV-I endemic area was discovered in Jujuy, a northwestern province in Argentina, to detect other possible foci of HTLV-I/II we started in 1997 a multicentric study to estimate the seroprevalence of HTLV-I/II including blood banks from different regions of the country. Out of 123,233 samples from nine different provinces, 68 proved seropositive with a final prevalence of 0.05% (95% CI = 0.0432-0.0704%). The seroprevalence ranged from 0.03 to 0.16% depending on the geographic location. An HTLV-I [odds ratio (OR) 12.7, 95% confidence interval (CI), 3.43-47.60] prevalence significantly greater was found in the northwest (Salta Province) and a nonsignificant but higher prevalence of HTLV-II in the North (Chaco). Two (0.001%) of the seropositive blood donors reported risk factors due to receiving transfusions and 19 (0.01%) were born in HTLV-I/II South American endemic areas. Although Argentina has been considered a nonendemic country for HTLV-I/II, the need for systematic screening for such infection became imperative based on the present study. PMID- 15665639 TI - Predictors of HIV serostatus among HIV discordant couples in Lusaka, Zambia and female antenatal clinic attendants in Kigali, Rwanda. AB - Clinical manifestations of HIV disease in Africa are nonspecific and easily confused with other endemic diseases. Several studies have compared the prevalence of HIV-related signs and symptoms in infected versus uninfected populations, but little is known about differences in HIV disease manifestations between African men and women across geographic areas. We conducted a cross sectional study to define predictors of HIV status and assess their differences by gender and country in two African cohorts: 1351 heterosexual couples recruited from a voluntary HIV counseling and testing center in Lusaka, Zambia, and 1458 women recruited from antenatal and pediatric clinics in Kigali, Rwanda. HIV positive Zambian men and women differed most with respect to prevalence of wasting syndrome (48.1% vs. 35.5%, p < 0.01). Zambian women were more likely to have a disseminated adenopathy than Rwandan women (33.2% vs. 7.8%, p < 0.01) and had a much higher median erythrocyte sedimentation rate (ESR) than either of the two other groups (78 mm/hr vs. 47 mm/hr, p < 0.01). Multivariable logistic regression modeling showed a history of tuberculosis [odds ratio (OR): 2.8-20.7], adenopathy on examination (OR: 4.0-6.3), and an ESR of >65 mm/hr (OR: 3.1-5.9) to be strongly predictive of HIV status in all groups. These screening tools, though highly predictive of HIV infection, were insensitive, as most infected persons were asymptomatic. Given these differences in HIV disease manifestation, screening tools based on signs and symptoms should be adapted accordingly. Additional studies are required to evaluate clinical markers as predictors of HIV disease progression and adjust them according to regional and gender differences. PMID- 15665640 TI - Longitudinal assessment of immune response and viral characteristics in HIV infected patients with prolonged CD4(+)/viral load discordance. AB - Although suppression of HIV-1 RNA below the limit of detection is associated with optimal outcomes, many patients can maintain or increase their CD4(+) count for prolonged time periods in the presence of persistent low-level viremia. We followed seven patients with prolonged (>5 years) discordant CD4(+)/viral load (VL) responses on protease inhibitor (PI)-based highly active antiretroviral therapy (HAART) prospectively for 1 year to assess evolution of immune function, viral phenotype, replication capacity (RC), and resistance profile. Immune function was assessed by qualitative and quantitative measurement of cellular activation (CD38(+)HLA-DR(+) and CD38 antibodies bound per cell), and the interferon (IFN)-() ELISpot assay. Presence of syncytium-inducing (SI) or nonsyncytium-inducing (NSI) viral strains was determined by MT-2 cell culture. RC was measured by a modified rapid recombinant virus assay. The resistance profile was characterized by both genotypic and phenotypic analysis. Over the year of follow-up, IFN-() production to gag persisted, responses to other HIV antigens increased, and markers of cellular activation did not change. NSI virus predominated. The genotypic (GSS) and phenotypic (PSS) susceptibility scores remained stable. Evolution of RC was variable over the year of follow-up, but the RC of viruses remained well below that of wild-type clinical isolates. Thus, CD4(+)/VL discordance can be maintained for periods exceeding 5 years in some patients receiving PI-based HAART without significant evolution of HIV resistance. PMID- 15665641 TI - Nigerian HIV type 2 subtype A and B from heterotypic HIV type 1 and HIV type 2 or monotypic HIV type 2 infections. AB - The presence of HIV-2 in Nigeria has been confirmed serologically, but not genetically. To determine the frequency of HIV-2 infections and the dynamics between HIV-1 and HIV-2 in 35 of 36 Nigerian states, 420 blood samples were collected in 1999. Antibodies to HIV-1 and HIV-2 were detected by EIA and seroreactivity was confirmed with the INNO-LIA HIV Line Assay. The frequency of HIV-2 was 4.3% (18 of 420), with 3.8% (16 of 420) HIV-1 and HIV-2 (HIV-1/2) heterotypic and 0.5% (2 of 420) HIV-2 homotypic infections. The presence of HIV-2 subtype B in the two monotypic HIV-2 infections and subtype A in 11 (68.8%) of 16 HIV-1/2 dually seropositive samples was established by sequencing and phylogenetic analysis. HIV-2 subtype B viruses were not found in any of the HIV 1/2 dual infections, and HIV-2 subtype A strains were not identified in either of the two monotypic HIV-2 infections. Since our sample size was small and represented only convenience samples, larger randomized studies will be needed to better understand the dynamics of infection between HIV-1 and different HIV-2 subtypes and to determine whether significant biological differences exist among the HIV- 2 subtypes. PMID- 15665642 TI - New insights in HTLV-I phylogeny by sequencing and analyzing the entire envelope gene. AB - The HTLV-I envelope plays a major role in the process of target cell infection. It is implied in the recognition of the viral receptor(s), penetration of the viral genetic material, and induction of host immunity to the virus. It is thus important to study the genetic variability of the viral env gene as well as its variation in terms of evolution. In a new approach to these features, we sequenced the entire env gene of 65 HTLV-I isolates originating from Gabon, French Guiana, West Indies, and Iran, such isolates representing all major HTLVI phylums but the Australo-Melanesian one. The sequences obtained and all PTLV-I (HTLV-I and STLV-I) env sequences available in the literature were analyzed. Phylogenetic studies using different algorithms (minimum evolution, neighbor joining, maximum parsimony, and maximum likelihood) gave the same clear-cut results. Newly sequenced HTLV-I isolates described in this report allocated in three well-defined subtypes: Cosmopolitan, Central African, and a new distinct one that we termed "Maroni" subtype (present in the Maroni Basin, French Guiana, and West Indies). Clearly, the most divergent PTLV-I strains present in Asia- Australo-Melanesia as well as African and Asian STLV-I derived from the same node in the phylogenetic tree as isolates of the Central African subtype. In addition, we showed that within each PTLV-I subtype, groups of isolates may be characterized by nonrandom and systematically associated mutations. PMID- 15665644 TI - HIV type 1 protease inhibitors enhance bone marrow progenitor cell activity in normal subjects and in HIV type 1-infected patients. AB - HIV-1 protease inhibitors (PIs) may improve hematopoietic functions owing to their direct effects on bone marrow (BM) progenitor cells. In this study we investigated this hypothesis evaluating the effect of adding ritonavir (RTV) and indinavir (IND) on hematopoietic colony formation assays by colony-forming cell (CFC) and long-term culture-initiating cell (LTC-IC) assays, on apoptosis, on cytokine production and stromal cells, in subjects with HIV-1 infection, and in seronegative controls. After PI addition, CFC and LTC-IC assays in HIV-1-infected patients showed levels of colony growth significantly higher than those observed at baseline; the same PI activity on colony formation was observed in healthy subjects. No significant modifications on Fas, the membrane form of Fas (mFas) and Fas-ligand (FasL) expression, and on cytokine production were observed at BM level after the addition of PIs. At baseline, in HIV-1-infected patients, the majority of the stromal cells appeared as large and rounded, whereas after the addition of RTV or IND the stromal cells exhibited a "fibroblast-like" morphology and produced higher stem cell factor (SCF) and lower MIP-1alpha levels when compared with the stromal production without the addition of IND. RTV and IND increased colony growth of BM obtained either from HIV-1-infected patients or from normal individuals, in parallel with the normalization of functional and morphological characteristics of stromal cells. PMID- 15665645 TI - Comparison of HIV Type 1 ADA gp120 monomers versus gp140 trimers as immunogens for the induction of neutralizing antibodies. AB - Designing an immunogen for effective neutralizing antibody induction against diverse primary isolates of human immunodeficiency virus type 1 (HIV-1) is a high priority for HIV-1 vaccine development. Soluble gp120 envelope (Env) glycoprotein subunit vaccines elicit high titers of antibodies that neutralize T cell line adapted (TCLA) strains but the antibodies possess poor neutralizing activity against many primary isolates. Previously, we generated soluble trimeric recombinant gp140 from the HIV-1 primary isolate ADA. Here we compared monomeric ADAgp120 and trimeric ADAgp140 as immunogens for neutralizing antibody responses in guinea pigs. Both immunogens generated a neutralizing antibody response that was detectable against the vaccine strain and several heterologous strains. The magnitude of this response was significantly greater in ADAgp140-immunized animals when measured against the TCLA strain, MN, and the R5 primary isolate, Bal. Two additional isolates (SS1196 and Bx08) were neutralized equally by sera from both groups of animals whereas other isolates were neutralized weakly or not at all. Despite equal titers of V3 loop specific binding antibodies in sera from both groups of animals, neutralization of ADA by sera from gp140-immunized animals was insensitive to the presence of ADA-V3 peptide, whereas addition of this peptide to sera from gp120- immunized animals blocked all detectable neutralizing activity against ADA. These results support the idea that trimeric gp140 is an improved immunogen compared to monomeric gp120 but that additional improvements are required to afford broad protection against a spectrum of heterologous primary HIV-1 isolates. This ADAgp140 immunogen may be considered a starting point from which to engineer additional improvements for cross-reactive neutralizing antibody induction. PMID- 15665643 TI - Cholesterol dependence of HTLV-I infection. AB - Cholesterol-rich plasma membrane microdomains are important for entry of many viruses, including retroviruses. Depletion of cholesterol with 2-hydroxypropyl beta-cyclodextrin inhibits entry of human T cell leukemia virus type I (HTLV-1) and HTLV-I envelope pseudotyped lentivirus particles. Using a soluble fusion protein of the HTLV-I surface envelope protein with the immunoglobulin Fc domain, the HTLV-I receptor was found to colocalize with a raft-associated marker and to cluster in specific plasma membrane microdomains. Depletion of cholesterol did not alter receptor binding activity, suggesting a requirement for cholesterol in a postbinding virus entry step. PMID- 15665646 TI - Results of an ELISPOT proficiency panel conducted in 11 laboratories participating in international human immunodeficiency virus type 1 vaccine trials. AB - We used an external quality assurance (EQA) panel to assess laboratory competency and comparability when performing ELISPOT assays in support of human immunodeficiency virus type 1 (HIV-1) vaccine trials. Cell recovery, viability, and frequency of interferon-gamma (IFN-gamma)-secreting cells after antigen stimulation were obtained from 11 laboratories on a coded panel of 11 peripheral blood mononuclear cell samples. The median recovery and viability before plating for all samples were 35% and 86%, respectively, with notable interlaboratory and intrasample variability. Empirical as well as statistical analysis methods were used to define positive ELISPOT responses. Remarkable concordance between laboratories was obtained in defining a qualitative assessment of responder/nonresponder status to antigens, but the frequency of responding cells varied among the laboratories. This study highlights the need for better standardization of protocols and reagents to obtain reliable and reproducible data that may support immunogenicity studies, vaccine regulatory submissions, and licensure. PMID- 15665647 TI - Immunogenicity of HIV type 1 gp120 CD4 binding site phage mimotopes. AB - The conserved domain of the CD4 binding site (CD4bs) on the human immunodeficiency virus type 1 (HIV- 1) envelope represents a potential target for vaccine development. Here we describe selection of peptide mimotopes by panning a phage peptide library on the HIV-1 CD4bs-specific, broadly neutralizing anti-HIV 1 monoclonal antibody, IgG(1) b12. We identified an initial consensus sequence for IgG1 b12 binding (M/VThetaSD, where Theta represents an aromatic amino acid). A molecular evolution approach, using second- and third-generation libraries, led us to identify a refined consensus sequence (GLLVWSDEL). The resulting IgG1 b12 phage mimotopes compete with gp160 for the IgG1 b12 antigen-binding site, but the phage coat protein (pIII) may play an important structural role, since both free peptides and KLH-conjugated peptides have no detectable binding activity. Mice immunized with IgG1 b12 phage mimotopes elicited a weak but persistent humoral response directed against the HIV-1 envelope. An antibody fragment was isolated from the antibody repertoires of these animals. It is noteworthy that while it has a relatively low affinity for HIV-1 gp160, the antibody targets an epitope that overlaps with that of IgG1 b12. Our data therefore suggest that engineered IgG1 b12 mimotopes share immunogenic features with the CD4bs. However, these peptidic structures will require further improvement in order to generate broad specificity neutralizing antibodies like IgG1 b12. PMID- 15665649 TI - A modified bootscan algorithm for automated identification of recombinant sequences and recombination breakpoints. AB - We have developed a modified BOOTSCAN algorithm that may be used to screen nucleotide sequence alignments for evidence of recombination without prior identification of nonrecombinant reference sequences. The algorithm is fast and includes a Bonferroni corrected statistical test of recombination to circumvent the multiple testing problems encountered when using the BOOTSCAN method to explore alignments for evidence of recombination. Using both simulated and real datasets we demonstrate that the modified algorithm is more powerful than other phylogenetic recombination detection methods and performs almost as well as one of the best substitution distribution recombination detection methods. PMID- 15665648 TI - Limited evolution in the HIV type 1 pol region among acute seroconverters in Pune, India. AB - India has the second largest burden of HIV-1-infected persons worldwide. Access to antiretroviral drugs in India is increasing. We analyzed HIV-1 protease and reverse transcriptase sequences in 12 acute seroconverters from Pune, India, and evaluated HIV-1 evolution in these individuals over time. HIV-1 genotyping was performed with the ViroSeq HIV-1 Genotyping System. Baseline samples, collected between 1999 and 2001, had viral loads from 3,523 to 8,556,280 copies/ml. All subjects had subtype C HIV-1. None of the samples had primary drug resistance mutations. The sequence identity between baseline and 1-year samples ranged from 99.7% to 99.9%, and between baseline and 2-year samples ranged from 99.4% to 100%. Most of the nucleotide changes were silent (synonymous). Amino acid substitutions were rare, and varied from subject to subject. In this cohort, drug resistance was not observed and evolution in the pol region was very limited during the first 2 years of infection. PMID- 15665650 TI - Characterization of human immunodeficiency virus type 1 from a previously unexplored region of South Africa with a high HIV prevalence. AB - HIV prevalence in the Limpopo Province has increased rapidly within the past 10 years, as in other parts of South Africa. Little is known about the genetic and biological properties of HIV circulating in this region including the baseline drug resistance profiles. We therefore collected blood samples from 42 HIV-1 infected patients residing in this region for analysis. All samples were shown to belong to HIV-1 subtype C by env and gag heteroduplex mobility assay (HMA). Viral isolates from 14 of these patients were shown to use the CCR5 coreceptor exclusively and had gp120 V3 loop sequences consistent with this phenotype. Sequence analysis of both protease and reverse transcriptase genes showed that none of 13 isolates harbored primary resistance mutations. These data suggest that HIV-1 subtype C is the predominant subtype circulating in the Limpopo Province, and that viral strains from this region are indistinguishable from those found in other parts of South Africa. PMID- 15665652 TI - Report from the CDC. The National Public Health Initiative on Diabetes and Women's Health: leading the way for women with and at risk for diabetes. AB - Diabetes is a serious public health problem in the United States. The burden of the disease has had a significant impact on individuals, communities, and society at large, and the number of people with diabetes is expected to double by the year 2025. In response to the growing burden of the disease and the profound health consequences for women and their offspring, the National Public Health Initiative on Diabetes and Women's Health convened a call-to-action conference to form a collective effort to address diabetes and women's health issues. This paper documents the process of developing a call-to-action conference and identifying potential stakeholders and presents results from the conference and the accomplishments of the National Public Health Initiative on Diabetes and Women's Health. PMID- 15665653 TI - Toward optimal health: the experts discuss chronic obstructive pulmonary disease in women. Interview by Jody Godfrey. PMID- 15665654 TI - The impact of the Women's Health Initiative on discontinuation of postmenopausal hormone therapy: the Minnesota Heart Survey (2000-2002). AB - BACKGROUND: The Women's Health Initiative (WHI) Data Safety and Monitoring Board terminated the trial arm of the estrogen plus progestin combined hormone therapy early after observing an excess of harm relative to benefit of this therapy. METHODS: To learn how postmenopausal hormone therapy (PMT) use in a community setting was affected by the WHI results, we resurveyed current users of PMT in August 2002 to January 2003 after publication of the WHI findings from our Minnesota Heart Survey (MHS) cohort (2000-2002), a cross-sectional epidemiological survey in the Minneapolis-St. Paul, Minnesota, area. We evaluated women's interpretation of the WHI results, reports of their physicians' recommendations, and the impact of these on PMT use in this group. RESULTS: We obtained an excellent response rate (97.9%, 185 of 189). Almost a third of women with an intact uterus but few women with a hysterectomy discontinued their PMT regimen within the first 6 months after the WHI research results were released. The majority of women in our study consulted their physician about whether to discontinue PMT. Women in the intact uterus group who started their PMT regimen for menopausal symptoms were less likely to discontinue PMT. Of women who continued their PMT regimen, the majority cited relief of menopausal symptoms or physician recommendation for continuation of PMT or both. Almost a third of the women interviewed reported being confused, worried, or nervous about the media reports of the WHI results. CONCLUSIONS: These results suggest effective dissemination of WHI information by the media and physicians and significant changes in patterns of PMT use in the community. PMID- 15665655 TI - The impact of the Women's Health Initiative on hormone replacement therapy in a Medicaid program. AB - BACKGROUND: Observational studies suggest that hormone replacement therapy (HRT) reduces women's risk of cardiovascular disease (CVD). However the Women's Health Initiative (WHI) HRT arm was stopped early because of increased risk of CVD, stroke, pulmonary embolism, and breast cancer. Evaluating HRT-prescribing patterns in relation to the release of the WHI will help determine if these study results have influenced the use of HRT. METHODS: This is a descriptive time series analysis to primarily evaluate utilization trends of HRT in women > or =50 years enrolled in the Pennsylvania Medicaid Program. HRT was categorized as follows: conjugated equine estrogen (CEE) plus medroxyprogesterone acetate (MPA), CEE only, and other estrogens/progesterone combinations. RESULTS: The overall prevalence of HRT decreased significantly post-WHI study release for all age and racial groups. The prevalence of CEE plus MPA decreased throughout the study; however, the prevalence of HRT declined at a faster rate after the WHI study release among women in their 50s and 60s compared with women in their 70s and 80s. There was a statistically significant reduction in all types of HRT use. CONCLUSIONS: The WHI influenced all types of HRT use among postmenopausal women in a Medicaid program. Administrative claims data can be a useful tool for monitoring an immediate impact of national guidelines or a national level outcomes trial. PMID- 15665656 TI - The relationship between serum dehydroepiandrosterone sulfate concentration and bone mineral density, lipids, and hormone replacement therapy in premenopausal and postmenopausal women. AB - OBJECTIVE: To investigate the relationship between serum dehydroepiandrosterone sulfate (DHEAS) concentration and bone mineral density (BMD), lipids, and hormone replacement therapy (HRT) in postmenopausal women. METHODS: Two hundred twenty seven women aged >40 years were admitted to the study and divided into three groups: 61 premenopausal normally menstruating women, 108 postmenopausal women who were not receiving HRT, and 58 postmenopausal women receiving HRT. DHEAS levels and lipid patterns were measured. BMD measurements of the lumbar spine (L1 L4) were performed by dual-energy x-ray absorptiometry (DXA). RESULTS: A positive correlation between DHEAS levels and BMD was found in all three groups. A linear regression model was used to assess the effect of age, body mass index (BMI), parity, and BMD of lumbar vertebrae (L1-L4) on changes in the serum level of DHEAS and found that aging and BMD at the spine had a significant association with serum level of DHEAS and only aging after adjustment for age, BMI, parity, and lipid patterns among the three groups. Age (RR 0.80), DHEAS (RR 0.98), and osteoporosis/osteopenia rate (RR 24.94) were also found to be independent influencing factors for HRT use. CONCLUSIONS: A positive correlation between DHEAS levels and BMD was found in all three groups. Our study confirms earlier reports that DHEAS levels decrease with age in premenopausal and, especially, postmenopausal women regardless of estrogen treatment. DHEAS levels in postmenopausal women were not associated with atherogenic lipid patterns in the present study. PMID- 15665657 TI - Decreased bone resorption with soy isoflavone supplementation in postmenopausal women. AB - OBJECTIVE: To investigate the effect of soy isoflavone supplementation on bone mineral density (BMD) and markers of bone turnover in postmenopausal women. METHODS: In this randomized, placebo-controlled clinical trial, we used a crossover design to test the effect of soy isoflavone (110 mg/day) (1.3:1.0:0.22 ratio of genistein/daidzein/ glycitein) on bone formation, bone resorption, bone mineral content (BMC), and BMD for 6 months. RESULTS: Postmenopausal women (n = 19), mean age 70.6 +/- 6.3 years and mean time since menopause 19.1 +/- 5.5 years, were given isoflavone supplements for 6 months. There was a 37% decrease in urinary concentrations of type 1 collagen alpha1-chain helical peptide (HP), a marker of bone resorption, during the isoflavone supplementation compared with baseline (p < 0.05) and a significant difference in mean (SE) HP excretion levels when isoflavone was compared with placebo (43.4 +/- 5.2 vs. 56.3 +/- 7.2 microg/mmol creatinine [cr], p < 0.05). With isoflavone supplementation, mean spine BMD at L2 and L3 was significantly greater when treatment was compared with control, with a difference between means of 0.03 +/- 0.04 g and 0.03 +/- 0.04 g (p < 0.05), respectively. There were nonsignificant increases from baseline for total spine BMC (3.5%), total spine BMD (1%), total hip BMC (3.6%), and total hip BMD (1.3%) with the isoflavone treatment. CONCLUSIONS: Soy isoflavone, in isolated form, was effective in this study to significantly decrease bone resorption in postmenopausal women. Further investigation needs to be done to evaluate the long-term effects of soy isoflavone on bone mass and fracture risk. PMID- 15665658 TI - Can hyperbaric oxygen therapy reduce breast cancer treatment-related lymphedema? A pilot study. AB - OBJECTIVE: Arm lymphedema after surgery or radiation for breast cancer is common, causing pain and limitation of activities. Previous reports of hyperbaric oxygen (HBO) therapy for breast edema led us to consider the use of HBO therapy for arm lymphedema. METHODS: Ten healthy postmenopausal women (age 58 +/- 5.7 years) with persistent (9.4 years +/- 9.1 years) arm lymphedema following breast cancer surgery and radiation (n = 10) plus chemotherapy (n = 7) received 20 HBO treatments (90 minutes at 2.0 ATA five times a week for 4 weeks). End points included changes in upper extremity volume, platelet counts, plasma levels of vascular endothelial growth factor (VEGF), and lymph angiogenic-associated vascular endothelial growth factor-C (VEGF-C). Lymphedema volume (LV) was defined as the volume of the unaffected arm subtracted from the volume of the affected arm. RESULTS: We observed a 38% average reduction in hand lymphedema (-7.4 ml, 11.6 SD, range -30-+8 ml, p = 0.076, 95% confidence interval -15.7-0.9 ml) at the end of HBO, which was independent of changes in body weight. For those who benefited (n = 8), the reduction was persistent from the end of treatment to a final measurement an average of 14.2 months after the last HBO treatment. However, total LV did not change significantly. VEGF-C increased from baseline (p = 0.004) before treatment 20, suggesting HBO had begun to stimulate this growth factor. CONCLUSIONS: Future studies should explore the effects of a greater number of HBO treatments on lymphedema, with more patients. PMID- 15665659 TI - Health and sociodemographic factors associated with body weight and weight objectives for women: 2000 behavioral risk factor surveillance system. AB - BACKGROUND: The increasing body mass index (BMI) of women in the United States gives rise to concerns about associated comorbid conditions and decreases in life expectancy. Also of concern are underweight women, especially as the result of an eating disorder or undernutrition. METHODS: Data from a national sample of women aged > or =18 years (n = 98,387) are used to examine the relationship between health and sociodemographic factors (diabetes, physical activity, self-rated health, smoking status, weight loss attempts, age, and education) and body weight (BMI, desired weight). Models are stratified by race. RESULTS: Roughly 70% of the women in each race/ethnic group (72.0% white women, 68.3% black women, 69.4% Hispanic women) wanted to weigh less, and just under one half of the women were actively trying to lose weight. A notable percentage of women who were classified as obese indicated that they were at their ideal weight and desired no weight change. Most women had not received advice from a health professional in the past year regarding their weight, and most were not engaging in the optimally recommended level of physical activity. CONCLUSIONS: Results document the range of satisfaction with current weight among adult women and capture low levels of health practitioner involvement in issues of weight. Perception of weight, combined with BMI, will need to be assessed to determine how best to proceed toward an ideal weight and satisfaction with that weight. PMID- 15665661 TI - Vaginal douching: personal practices and public policies. AB - BACKGROUND: Vaginal douching is associated with adverse reproductive health outcomes, yet both health providers and women are often poorly informed about details of this practice. METHODS: We searched the English language articles in the MEDLINE database (1965-March 2004) to describe vaginal douching products, policies of professional organizations, predictors of douching practice, douching methods used, timing of use, and motivation. A key report was obtained from the Food and Drug Administration (FDA) via the Freedom of Information Act. Additional product information was obtained from manufacturers. Primary key terms for the literature search included (vagina OR vaginal) and (douche OR douching). Health effects of douching are reviewed briefly; personal practices and public policies are highlighted. RESULTS: From the literature search, we identified 432 papers, of which 150 were reviewed in detail. Contrary to the assumptions of many health professionals, douching products are only loosely regulated by the FDA. Few professional organizations have clearly stated policy statements regarding douching. In the United States, the prevalence of douching varies considerably by race (more common among African Americans) and age cohort (more common in women born earlier). Internationally, vaginal douching is common in some cultures and is rare in others. Opinions of mothers, peers, and health professionals, in addition to marketing of commercial products, affect douching behavior. CONCLUSIONS: Regulation of vaginal douching products and public education efforts on douching behavior need to be reassessed. Because of the preponderance of evidence that suggests an association between vaginal douching and adverse reproductive health outcomes, professional and public health associations should consider educational and policy activities to discourage women from douching. PMID- 15665660 TI - Sexual orientation, health risk factors, and physical functioning in the Nurses' Health Study II. AB - OBJECTIVES: To examine associations between sexual orientation and breast cancer risk factors, cardiovascular disease (CVD) risk factors, mental health status, and health-related functioning. METHODS: We compared participants in the Nurses' Health Study II (NHSII) reporting a lesbian or bisexual orientation with those reporting a heterosexual orientation, with heterosexuals serving as the reference group for all comparisons. Prevalence of health behaviors and conditions was adjusted for differences in the distribution of age, ancestry, and region of residence by standardizing to the distribution of the overall cohort. Multivariate prevalence ratios were calculated to compare lesbians and bisexuals with heterosexuals using binomial regression with the log link function. Means of health conditions were measured using continuous scales standardized to the distribution of the overall cohort. Differences in means comparing lesbians and bisexuals with heterosexuals were tested by multivariate linear regression. All comparisons were adjusted for age, ancestry, and region of residence. RESULTS: Based on information from 90,823 women aged 32-51 in 1995, those reporting a sexual orientation of lesbian (n = 694) had a higher prevalence of risk factors for breast cancer, including nulliparity and high daily alcohol intake, compared with heterosexual women. Lesbians also had a higher prevalence of several risk factors for CVD, including higher body mass index (BMI) and elevated prevalence of current smoking. Lesbians were more likely to report depression and the use of antidepressants. Key results for health risk factors were similar for lesbians and bisexual women (n = 317). CONCLUSIONS: Lesbian and bisexual women were found to have a higher prevalence of several important risk factors for breast cancer, CVD, and poor mental health and functioning outcomes. Most of these risk factors are modifiable, and appropriate interventions could play an important role in improving the health status of lesbian and bisexual women. PMID- 15665665 TI - Lid wiper epitheliopathy and dry eye symptoms. AB - OBJECTIVES: The lid wiper is defined as that portion of the marginal conjunctiva of the upper eyelid that wipes the ocular surface during blinking. The purpose of this study was to investigate whether lid wiper epitheliopathy occurred with patients who reported dry eye symptoms, yet had normal fluorescein breakup time (FBUT) and Schirmer test values and an absence of fluorescein corneal staining. METHODS: One hundred patients were divided into two groups based on the presence or absence of dry eye symptoms, as determined with the Standard Patient Evaluation of Eye Dryness questionnaire. Other criteria for admission to both groups were FBUT of 10 seconds or more, Schirmer test value of 10 mm or more, and absence of fluorescein corneal staining. After instillation of fluorescein and rose bengal dyes, the lid wipers of 50 asymptomatic and 50 symptomatic patients were graded for staining from grade 0 (absent) to grade 3 (severe). RESULTS: Of the symptomatic patients, 76% had staining of the lid wiper: 44%, grade 1; 22%, grade 2; and 10%, grade 3. Of the asymptomatic patients, 12% had staining; 8%, grade 1; 4%, grade 2; and 0%, grade 3. The difference in prevalence of lid wiper staining between the symptomatic and asymptomatic groups was significant (P<0.0001). CONCLUSIONS: Lid wiper epitheliopathy, diagnosed by staining with fluorescein and rose bengal dyes, is a frequent finding when symptoms of dry eye are experienced in the absence of routine clinical dry eye findings. PMID- 15665666 TI - Corneal ulcers in two children wearing paragon corneal refractive therapy lenses. AB - OBJECTIVES: To report the development of corneal ulcers in two children wearing Paragon Corneal Refractive Therapy (CRT) lenses. METHODS: Two case reports of children who developed corneal ulcers when fit with Paragon CRT contact lenses, which were worn nightly, are discussed. Each patient presented with a bacterial corneal ulcer after wearing CRT contact lenses for less than 6 months. In the first patient, Pseudomonas aeruginosa was identified as the causative organism. In the second patient, Haemophilus influenza was cultured from the ulcer. RESULTS: Both patients were treated with intensive, fortified antibiotics with rapid resolution of the corneal ulcers and preservation of vision. No surgical intervention was required. CONCLUSIONS: Paragon CRT lenses have been approved for use in patients of all ages. When used in children, these lenses may present unique problems. The absolute incidence of bacterial corneal ulcers in patients with CRT lenses are unknown. Severe caution should be used before prescribing CRT lenses for children and informed consent should include potential sight threatening corneal ulcers. PMID- 15665667 TI - Therapeutic effects of contact lenses after refractive surgery. AB - PURPOSE: Although highly successful, refractive surgery can have adverse effects, such as loss of best spectacle-corrected visual acuity, glare, haloes, monocular polyopia, and loss of contrast. Further surgical intervention may no longer be an adequate or appropriate response. The efficacy of rigid gas-permeable contact lenses and their therapeutic effects in managing these conditions are studied. METHODS: A retrospective review of the records of 67 eyes of 37 patients fitted with contact lenses after different refractive surgeries was performed. The most successful lens design for each patient was evaluated. Klyce corneal statistics of surface regularity index and surface asymmetry index, mean keratometric values, and optical zone diameter were measured before and after contact lens wear using the Tomey TMS-2 topographer. Comparative wavefront aberrometry was obtained for five eyes of three patients. RESULTS: The cohort had a mean uncorrected visual acuity of 20/40, a mean best spectacle-corrected visual acuity of 20/26, and a mean visual acuity with contact lenses of 20/20. Contact lens wear resulted in significant improvements in corneal regularity and symmetry, as indicated by Klyce corneal statistics, with concomitant improvements in visual acuity between wearing periods. The mean visual acuity on lens removal was 20/24. This lens molding effect lasted from several hours to several days. The lenses also had the effects of decreasing higher-order aberrations, as measured by wavefront aberrometry in three patients. CONCLUSIONS: Rigid gas-permeable contact lenses can dramatically improve visual acuity and higher-order aberrations in eyes of patients with unsatisfactory refractive surgery outcomes. Contact lenses can also have a measurable therapeutic effect on vision when the lenses are not being worn or cannot be worn full time. PMID- 15665668 TI - Corneal oxygen deficiency. AB - OBJECTIVES: To compare the ocular effects of low Dk hydrogel and high Dk silicone hydrogel contact lenses. METHODS: A review of recent literature was conducted to determine the effects of wearing low and high Dk soft lenses on corneal swelling, vascular response, refractive error and the corneal epithelium. RESULTS: High Dk silicone hydrogel contact lenses became available for research and clinical practice almost 10 and 5 years ago respectively. During that time many studies have been conducted and it has been demonstrated repeatedly how much improvement there has been to the physiological response of the eye particularly when wearing the lenses on an overnight basis. CONCLUSIONS: Although silicone hydrogels only constitute about 2% of current wearers, their positive effects on the eye compared with low Dk hydrogel lenses should make them the most dominant lenses in the near future. PMID- 15665669 TI - Keratocyte density: comparison of two confocal microscopes. AB - PURPOSE: The ConfoScan 3 clinical confocal microscope provides sharper images of cells in the stroma than the Tandem Scanning confocal microscope does. In this study, we compared volumetric densities of stromal cells determined from images recorded by these two instruments. METHODS: Fifty corneas of 25 normal subjects were examined by confocal microscopy, first by using a Tandem Scanning confocal microscope and then by using a ConfoScan 3 confocal microscope. Bright objects, assumed to represent keratocytes, were counted in a known area of two frames selected from the mid-stroma. The effective depth of the sample volume represented by each frame was estimated from the number of consecutive frames and the corresponding distance that selected cells were visible and countable during a scan. Density was the number of visible cells in the sample area divided by the effective sample volume. RESULTS: The effective focal depth of the Tandem Scanning microscope was 11.9 +/- 2.6 microm (mean +/- SD), and was 25.9 +/- 7.1 microm for the ConfoScan 3. Mean cell density at mid stroma was 23,013 +/- 4,420 cells/mm(3) with the Tandem Scanning microscope and 23,996 +/- 2,898 cells/mm(3) with the ConfoScan 3. This difference was not significant (P = 0.15). CONCLUSIONS: In normal corneas, the ConfoScan 3 and the Tandem Scanning confocal microscopes indicate stromal cell densities that are not significantly different from each other. Estimates of cell density from both instruments require an accurate estimate of the effective depth of the sample volume; this depth is approximately 2.2 times greater with the ConfoScan 3. This difference must be considered when comparing results from studies that use one instrument with results from studies that use the other. PMID- 15665671 TI - Effect of a multipurpose contact lens solution on the survival and binding of Acanthamoeba species on contact lenses examined with a no-rub regimen. AB - PURPOSE: To determine the effect of a multipurpose contact lens solution (ReNu MultiPlus Multi-Purpose Solution [RMP]) on the relative survival and binding of trophozoites and cysts of Acanthamoeba on hydrogel lenses with a no-rub regimen. METHODS: A stand-alone test procedure with RMP was conducted with and without the presence of organic soil (1 x 10(7) colony-forming units/mL heat-killed cells of Saccharomyces cerevisiae in heat-inactivated fetal bovine serum). Survival of amoebae on hydrogel contact lenses exposed to RMP was determined with a no-rub care regimen. RESULTS: ReNu MultiPlus Multi-Purpose Solution reduced the number of recoverable amoebae by more than 95% within 4 hours of inocula of 10(5) trophozoites and cysts, regardless of the presence or absence of an organic soil. Amoebae, particularly cysts, were readily rinsed from contact lenses, including silicone hydrogels, without rubbing after exposure to RMP. CONCLUSIONS: The efficacy of RMP for Acanthamoeba was not appreciably altered in the presence of organic soil in a no-rub protocol. The antimicrobial activity, in part, appeared to be a combination of reducing the capacity for binding of representative Acanthamoeba to the lens by alteration of morphology, often followed by lysis of the amoebae. PMID- 15665670 TI - Effect of preservative-free artificial tears on the antimicrobial activity of human beta-defensin-2 and cathelicidin LL-37 in vitro. AB - PURPOSE: Human beta-defensin-2 (hBD-2) and cathelicidin LL-37 are salt-sensitive cationic antimicrobial peptides expressed by ocular surface epithelia. The goal of this study was to investigate the effect of preservative-free artificial tears on hBD-2 and LL-37 antimicrobial activity against Pseudomonas aeruginosa. METHODS: P. aeruginosa was incubated with hBD-2 or LL-37 in the absence or presence (70% vol/vol) of different preservative-free artificial tears--Visine Tears (300 mOsm/kg), Tears Naturale Free (261 mOsm/kg), TheraTears (185 mOsm/kg), and Refresh Plus (325 mOsm/kg)--for 2 hours at 37 degrees C. In some experiments, P. aeruginosa was incubated with hBD-2 or LL-37 and Visine Tears or Tears Naturale Free with or without carboxymethylcellulose (0.5% vol/vol final concentration). Plates were inoculated with samples of each reaction mixture and then incubated for 24 hours at 37 degrees C. RESULTS: Visine Tears and Tears Naturale Free had little or no effect on the antimicrobial activity of 100 microg/mL hBD-2 or LL-37. In the presence of Refresh Plus and TheraTears, the activity of 100 microg/mL hBD-2 or LL-37 was reduced by 90% to 100%. Carboxymethylcellulose, at a concentration comparable to that present in Refresh Plus, reduced the effectiveness of hBD-2 or LL-37 by 40% to 90% in the presence of Tears Naturale Free and Visine Tears. CONCLUSION: Human beta-defensin-2 and cathelicidin LL-37 inhibit the growth of P. aeruginosa in vitro, but this activity is markedly reduced in the presence of Refresh Plus and TheraTears. These results suggest that carboxymethylcellulose-containing artificial tears may reduce the activity of the endogenously produced antimicrobial peptides. PMID- 15665672 TI - Microdot stromal degenerations. PMID- 15665674 TI - Computed tomography coronary angiography with 370-millisecond gantry rotation time: evaluation of the best image reconstruction interval. AB - OBJECTIVE: To evaluate the best reconstruction window for noninvasive coronary angiography when using a 16-detector row computed tomography (CT) scanner with a gantry rotation time of 370 milliseconds. METHODS: In a pilot study, 189 coronary artery segments of 21 patients with a mean heart rate of 65 beats per minute (bpm, maximum: 45-94 bpm) were investigated using a 16-detector row CT scanner. Raw data were reconstructed in 10% increments from 40% to 70% of the RR interval. Two experienced observers independently evaluated the image quality of the coronary arteries in a segmental fashion. A 5-point ranking scale was applied, with 1 being very poor (no evaluation possible); 2, poor; 3, moderate; 4, good; and 5, very good. RESULTS: In the mean of all patients, the best reconstruction window was found to be at 60% of the RR interval. In patients with higher heart rates, the best reconstruction window was found to be at an earlier stage of the R wave-to-R wave interval. CONCLUSIONS: Initial results show that good diagnostic image quality could be achieved for all evaluated segments of the coronary tree with image reconstructions at 60% of the R wave-to-R wave interval in patients with heart rates of 70 bpm or less. Using a 16-detector row CT scanner with a gantry rotation time of 370 milliseconds, the need for adapting the reconstruction window to each segment for the best image quality was overcome in those cases. In patients with heart rates faster than 70 bpm, reconstructions at an earlier stage within the cardiac cycle were necessary. PMID- 15665676 TI - Dynamic magnetic resonance imaging of solitary pulmonary nodules: utility of kinetic patterns in differential diagnosis. AB - OBJECTIVE: The aim of this study was to evaluate the clinical feasibility of dynamic magnetic resonance (MR) imaging with a 3-dimensional (3D) gradient recalled echo (GRE) volumetric interpolated breath-hold examination (VIBE) sequence to differentiate between benign and malignant solitary pulmonary nodules (SPNs). METHODS: Dynamic 3D GRE VIBE was performed in 45 patients with SPNs. For each lesion, the morphologic parameters, including the edge configuration, presence of peripheral enhancement (PE), and an internal signal on T2-weighted images, and the kinetic enhancement parameters were evaluated. RESULTS: All 29 of the malignant SPNs had internal enhancement, whereas 13 (81%) of the benign SPNs did not exhibit internal enhancement. A washout pattern was only observed in the malignant SPNs. The presence of PE was found in 56% of the benign SPNs and in 50% of the malignant SPNs. The lesion size was significantly different between malignant SPNs with PE and those without PE (P <0.01). The positive predictive value for malignancy was 91% (29 of 32 malignant SPNs). The negative predictive value was 100% (13 of 13 benign SPNs). CONCLUSIONS: A combination of morphologic criteria and kinetic information is useful for differentiating between benign and malignant SPNs. In particular, internal enhancement with PE and positive visual washout is thought be a useful tool. PMID- 15665675 TI - Clinical usefulness of computed tomography study without contrast injection in the evaluation of acute pulmonary embolism. AB - OBJECTIVE: To evaluate the percentage of cases in which emboli can be detected in unenhanced scans and to identify the cases in which they appear hyperattenuating or hypoattenuating in comparison to the circulating blood. METHOD: An angio computed tomography (CT) scan was performed before and after contrast injection in 140 consecutive patients after clinical suspicion of pulmonary embolism. A radiologist analyzed the examination results thus obtained. The enhanced scan was analyzed first, and after detecting the thrombus, the unenhanced scan was evaluated. RESULTS: Fifty-one examinations were positive for a pulmonary embolism; in 21 cases, it was possible to identify the embolus even in the unenhanced scans. In 10 cases, the clots were hyperattenuating in comparison to the circulating blood; in 5 cases, they were hypoattenuating; and in 6 cases, they were mixed hyper-hypoattenuating. CONCLUSION: In a relatively high percentage of cases, particularly those of central thromboembolism, it is possible to identify and characterize the clots even in unenhanced scans. PMID- 15665677 TI - Postoperative multidetector computed tomography angiography after aneurysm clipping: comparison with digital subtraction angiography. AB - OBJECTIVES: To evaluate the usefulness of multidetector computed tomography angiography (MDCTA) as a noninvasive diagnostic modality for the detection of aneurysm remnants after clipping of intracranial aneurysms. METHODS: Thirty-eight patients who had aneurysm clipping and had undergone MDCTA and digital subtraction angiography (DSA) were enrolled in this study. In 16 of the 38 patients, MDCTA was performed with 16-channel MDCTA, in 20 patients with 4 channel MDCTA, and in 2 with both. Two neuroradiologists evaluated the image quality of MDCTA with a 3-point rating scale and the presence of the residual aneurysm sac with a 5-point rating scale. Receiver operating characteristic analysis was used to measure the diagnostic performance of MDCTA. RESULTS: Forty nine surgically clipped aneurysms were evaluated in this study. The overall diagnostic accuracy of MDCTA was 88.1% (95% confidence interval: 0.761-0.954). The ability of 16-channel MDCTA to discriminate between surgical clips and parent vessels was superior to that of 4-channel MDCTA (P=0.005). CONCLUSION: Multidetector computed tomography angiography is a valuable noninvasive diagnostic modality for the assessment of aneurysm remnants in patients after aneurysm clipping. PMID- 15665678 TI - Volumetric cine imaging for cardiovascular circulation using prototype 256 detector row computed tomography scanner (4-dimensional computed tomography): a preliminary study with a porcine model. AB - This is a preliminary demonstration of volumetric cine imaging of cardiovascular circulation in domestic pigs using a prototype 256-detector row computed tomography (CT) scanner. The scan range is approximately 120 mm in the craniocaudal direction, with a 0.5-mm slice thickness. The thin sections can be used to create cine loops in multiple planes. Thus, the 256-detector row CT scanner overcomes some of the limitations of present helical CT methods. PMID- 15665680 TI - Transient left ventricular apical ballooning: magnetic resonance imaging evaluation. AB - The magnetic resonance imaging characteristics of transient left ventricular apical ballooning are described in this report, and the features that distinguish it from acute myocardial infarction are emphasized. PMID- 15665679 TI - Hydatid cysts of the bilateral pulmonary arteries and left ventricle wall: computed tomography and magnetic resonance imaging findings. AB - Echinococcal cysts located inside the pulmonary artery are extremely rare. The cause is usually rupture of intracardiac cysts or, more rarely, dissemination from a hepatic focus. The case of a 39-year-old patient with multiple hydatid cysts in the right and left pulmonary arteries and left ventricle wall is reported. The patient had undergone surgery for a hepatic hydatid cyst 10 years ago. Multidetector computed tomography angiography and magnetic resonance imaging were performed for the diagnostic evaluation. PMID- 15665681 TI - Imaging findings of intestinal tuberculosis. AB - Intestinal tuberculosis (TB) has 3 main forms: ulcerative, hypertrophic or ulcerohypertrophic, and fibrous stricturing. In the ulcerative form, barium examination reveals thickened folds, spasticity, and shallow ulcers involving the cecum and terminal ileum. Computerized tomography shows preferential thickening of the ileocecal valve and medial wall of the cecum as well as a few small regional nodes. In the hypertrophic or ulcerohypertrophic form, a hyperplastic reaction is seen in the exophytic masses around the ulcerated lumen on computed tomography. An inflammatory mass that extends into adjacent muscle suggests TB. In the sclerotic form, the main reaction is fibrosis with single or multiple short strictures. The cecum classically becomes amputated, conical, shrunken, and retracted. In comparison, Crohn's disease (CD) has a rather uniform and lesser thickening of the bowel wall. Mural stratification, vascular jejunization or the comb sign, and mesenteric fibrofatty proliferation are seen only in CD. The hypertrophic form may also mimic malignant neoplasms, such as lymphoma or carcinoma. Cecal carcinoma rarely extends beyond the ileocecal valve, however. In lymphoma, it can be seen as a greater degree of wall thickness with aneurysmatic dilation of the intestinal lumen. Single or multiple strictures are also seen as a CD complication. Advanced skip lesions adjacent to the stricture are usually diagnostic for CD. PMID- 15665683 TI - Dynamic subtraction magnetic resonance imaging of cirrhotic liver: assessment of high signal intensity lesions on nonenhanced T1-weighted images. AB - PURPOSE: The purpose of this work was to determine the technical feasibility and value of dynamic subtraction (postcontrast-precontrast) magnetic resonance (MR) imaging for the assessment of hyperintense lesions on precontrast T1-weighted images in the cirrhotic liver. METHODS: One hundred four hyperintense lesions on T1-weighted precontrast and arterial phase postcontrast images were subjected to analysis of their subtraction qualities depending on the lesion size, location and/or the degree of misregistration between the source images in 27 different MR imaging sets. RESULTS: The quality of subtraction images was always diagnostic for lesions larger than 2 cm in diameter (n=8) but not diagnostic for 73% (40 of 55 lesions) of small subcentimetric lesions. Thirty-one subcapsular lesions always showed a variable degree of coregistration artifact. Only 3 of 35 lesions with a slice misregistration of 3 mm or more gave rise to subtraction images of diagnostic quality. For determining the contrast enhancement, the area under the receiver operating characteristic curve of 30 verified lesions was significantly larger (P <0.001) for subtraction images than for conventional arterial phase images. CONCLUSION: Depending on the lesion size and/or location or the degree of misregistration between the source images, dynamic subtraction MR imaging can be useful for the characterization of hyperintense lesions on precontrast T1 weighted imaging. PMID- 15665682 TI - Detecting postsurgical recurrent hepatocellular carcinoma with multiphasic helical computed tomography: intrahepatic metastasis or multicentric occurrence? AB - OBJECTIVE: The aim of this study was to assess the computed tomography (CT) patterns of postoperative recurrent hepatocellular carcinoma (HCC). METHODS: Of 84 patients with histologically proven HCC by hepatectomies, multiphasic helical CT demonstrated 54 HCC lesions of intrahepatic recurrence in 31 (37%) patients. The initial and final appearances of HCC on hepatic arterial phase images were retrospectively determined by the serial CT scans, which were compared with appearances of primary HCC. RESULTS: The initial appearances of 54 recurrent HCCs were identical to the appearances of primary HCC in 41 (76%) lesions. Serial changes from the initial appearance to the final appearance of recurrent HCC were observed in 10 (42%) of 24 lesions. The 13 discordant lesions and the 10 lesions with altered lesion vascularity in our series implied that at least 43% were of multicentric occurrence. CONCLUSION: More than 40% of postoperative recurrent HCCs show intranodular hemodynamic changes. The incompatibility between CT findings of primary and recurrent HCCs implies that these tumors contain nodules of multicentric occurrence. PMID- 15665684 TI - Quantitative colorectal cancer perfusion measurement using dynamic contrast enhanced multidetector-row computed tomography: effect of acquisition time and implications for protocols. AB - OBJECTIVE: To determine the effect of acquisition time on quantitative colorectal cancer perfusion measurement. METHODS: Dynamic contrast-enhanced computed tomography (CT) was performed prospectively in 10 patients with histologically proven colorectal cancer using 4-detector row CT (Lightspeed Plus; GE Healthcare Technologies, Waukesha, WI). Tumor blood flow, blood volume, mean transit time, and permeability were assessed for 3 acquisition times (45, 65, and 130 seconds). Mean values for all 4 perfusion parameters for each acquisition time were compared using the paired t test. RESULTS: Significant differences in permeability values were noted between acquisitions of 45 seconds and 65 and 130 seconds, respectively (P=0.02, P=0.007). There was no significant difference for values of blood volume, blood flow, and mean transit time between any of the acquisition times. CONCLUSIONS: Scan acquisitions of 45 seconds are too short for reliable permeability measurement in the abdomen. Longer acquisition times are required. PMID- 15665685 TI - Preoperative staging of renal cell carcinoma with inferior vena cava thrombus using multidetector CT and MRI: prospective study with histopathological correlation. AB - OBJECTIVE: To evaluate the accuracy of multidetector computed tomography (CT) and magnetic resonance imaging (MRI) in staging and estimating renal carcinomas with caval thrombus. METHODS: Initially, 23 patients with suspected caval thrombi were admitted into this prospective study. Triphasic CT imaging was performed using a multidetector CT with a reconstructed slice thickness of 2 mm. 3D CT reconstructions were used to improve surgical planning. MRI protocol included: a transversal T1-weighted GE sequence with and without Gd-DTPA, a transversal T2 weighted respiratory-gated TSE, and a coronal T1-weighted GE sequence with Gd DTPA and fat saturation. In addition, a multiphase 3D angiography was performed after Gd-DTPA injection. Patients were divided into 3 groups: caval thrombus below the insertion of the hepatic veins, within the intrahepatic vena cava, and intra-atrial extension. The results the tumor thrombus extension and staging results of 2 independent readers were correlated with surgical and histopathological staging. RESULTS: Of the 23 patients admitted, CT and MR scans of 14/13 patients respectively were correlated with histopathological workup. CT thrombus detection sensitivity and specificity for both readers was 0.93 and 0.8 respectively. MRI sensitivity and specificity for both readers was 1.0/0.85 and 0.75. Readers I and II evaluated the uppermost extension of the cranial tumor thrombus by both CT and MRI. CT and MR accuracy was 78% and 72%, 88% and 76% respectively. CONCLUSION: In cases of a suspected tumor thrombus, MRI and multidetector CT imaging showed similar staging results. Consequently, these staging modalities can be used to assess the extension of the tumor thrombus. PMID- 15665686 TI - Computed tomography findings of ovarian metastases from colon cancer: comparison with primary malignant ovarian tumors. AB - OBJECTIVE: The computed tomography (CT) findings of ovarian metastases from colon cancer were evaluated and were compared with those of primary malignant ovarian tumors. METHODS: Sixteen patients with 21 masses from colon cancer and 20 patients with 31 primary malignant ovarian tumors were included in this study. The CT findings (laterality, size, margin, shape, mass characteristic, strong enhancement of cyst wall, enhancement of solid portion, amount of ascites, peritoneal seeding, lymph node enlargement, and metastasis) and ages of the patients in both groups were compared. Univariate analysis, the Pearson chi test, and the independent-samples t test were used to distinguish them. RESULTS: A smooth margin of the tumor (odds ratio=24.3, 95% confidence interval: 2.9-204.2) and cystic nature of the mass (Pearson chi=12.96, P=0.005) were strong predictors of ovarian metastasis from colon cancer. CONCLUSION: Ovarian metastases from colon cancer show a smooth margin and more cystic nature on CT compared with primary malignant ovarian tumors. PMID- 15665687 TI - Extraovarian cystadenomas: ultrasound and MR findings in 7 cases. AB - OBJECTIVE: To describe the ultrasound and MR appearance of paraovarian cystadenomas. METHODS: We reviewed retrospectively the radiologic findings in 7 patients with surgically proven paraovarian cystic neoplasms, including 6 serous cystadenomas and 1 borderline seromucinous cystadenoma. All had ultrasound and 4 had MR preoperatively. RESULTS: On ultrasound, the ipsilateral ovary was visualized in six cases, in contact with the cyst in five and separate from it in one. On MR, the ovary and the cyst were visible in four cases, in contact in three and separate in one. Internal papillary excrescences, present at pathology in all cysts, were seen in five on ultrasound and in four on MR. CONCLUSION: Although the extraovarian location of these neoplasms is difficult to determine preoperatively by ultrasound and MR, these imaging modalities are more reliable in predicting the histology of these rare lesions and differentiating them from simple paraovarian cysts. PMID- 15665688 TI - Magnetic resonance imaging of adrenal angiomyolipoma. AB - Angiomyolipoma is a benign mesenchymal neoplasm that typically occurs in the kidney sporadically or in patients with tuberous sclerosis complex. Extrarenal angiomyolipomas are uncommon, and the liver is the most common extrarenal site. Other sites reported include the bone, colon, heart, lung, parotid gland, skin, spermatic cord, gynecologic regions, and retroperitoneum. A case of an angiomyolipoma involving the right adrenal gland in a 49-year-old female patient with tuberous sclerosis is reported. The magnetic resonance (MR) imaging features are also discussed. Only 2 cases describing adrenal angiomyolipoma were found in the English literature. Neither of these 2 cases was documented in the radiology literature or discussed the MR features of adrenal angiomyolipoma. PMID- 15665690 TI - Osteonecrosis of the patella: diagnostic imaging perspective. AB - Disability from knee dysfunction has important implications on a patient's performance of simple day-to-day tasks, work-related duties, and leisure activities, including sports. Diagnostic imaging is of fundamental importance in the clinical evaluation of the patient with knee pain. The use of magnetic resonance imaging (MRI) for the assessment of knee pain and dysfunction has increased over the past few years as orthopedic surgeons and radiologists have familiarized themselves with the unique advantages of this modality. Magnetic resonance imaging provides a comprehensive evaluation of a wide spectrum of intra articular and extra-articular pathologic conditions involving the knee. One of the most common indications for MRI of the knee is osteonecrosis. Osteonecrosis of the patella is unusual, and it is a well-documented cause of knee pain. After a brief overview of the anatomy of the patella, the diagnosis of osteonecrosis at this site by conventional radiography, MRI, and other imaging methods is focused on in this review. PMID- 15665689 TI - Assessment of response of uterine fibroids and myometrium to embolization using diffusion-weighted echoplanar MR imaging. AB - Our objective was to evaluate the role of echoplanar diffusion-weighted imaging (DWI) and apparent diffusion coefficient (ADC) maps in the assessment of response of uterine fibroids and myometrium to uterine fibroid embolization (UFE). We evaluated 32 fibroids in 11 patients treated with UFE. Treated lesions had low ADC values compared to untreated lesions. Furthermore, ADC values of the myometrium did not significantly change after embolization, suggesting maintained myometrial viability after embolization. DWI and ADC maps, by providing functional information at a cellular level, may serve as imaging tools for assessing treatment response after UFE. PMID- 15665691 TI - Three-dimensional computed tomography imaging: its applicability in the evaluation of extensor tendons in the hand and wrist. AB - OBJECTIVE: The purpose of this study was to determine whether 3-dimensional (3D) computed tomography (CT) imaging with volume rendering can accurately evaluate extensor tendons in the hand and wrist. METHODS: Three-dimensional CT imaging was used to evaluate 14 cases. The location where the rupture was suspected was the wrist level in all cases. The accuracy of the image diagnosis was confirmed during surgery in 13 cases. RESULTS: Three-dimensional CT could identify all extensor tendons proximal to the metacarpophalangeal joint. In the cases with rupture, the location of the distal stump could be identified, whereas that of the proximal stump was uncertain. The tendon stump location confirmed during surgery correlated well with the images. CONCLUSIONS: This study revealed the usefulness of 3D CT imaging in defining extensor tendons proximal to the metacarpophalangeal joint. This method increased the accuracy and ease of diagnosis and was always useful in surgical planning and patient education. PMID- 15665692 TI - Is epidural fat associated with body habitus? AB - OBJECTIVE: To evaluate, in the spine, the relationship between the amount of epidural fat to body weight, height, body mass index (BMI), presence of obesity, depth of posterior subcutaneous fat, as well as gender and age. METHODS: At 1.5 T, 101 random patients were analyzed. In the lumbar spine, we calculated the depth of the anterior and posterior epidural fat, as well as posterior subcutaneous fat, separately at the L3-4, L4-5, and L5-S1 levels. Obtained via questionnaire was patient's age, gender, body weight, height, with a calculated BMI (body weight/height, kg/m). Statistical analysis was performed to assess the correlation between epidural and subcutaneous fat with age and gender, weight, height, BMI, and presence of obesity (BMI > 27.5 kg/m). RESULTS: There were 66 men and 45 women, age ranged 14-83 years old (mean 43). The weight range was 47.6 135.4 (mean 79.5 kg); height range was 124.5-208.3 (mean 169.7 cm); BMI range was 16.9-42.8 (mean 27.8). Female gender correlated only with subcutaneous fat depth (sum, r=-0.31, P=0.002). Younger patients had more anterior epidural fat (sum, r= 0.22, P=0.024) but not posterior epidural fat (sum, r=0.01, P=0.954) or subcutaneous fat (sum, r=0.09, P=0.0357). Weight correlated with posterior epidural fat (sum, r=0.21, P=0.037) and subcutaneous fat (sum, r=0.51, P <0.0001). Height showed correlation negatively with posterior subcutaneous fat thickness (sum, r=-0.25, P=0.014), but had no significant correlation with epidural fat. BMI showed a correlation with posterior subcutaneous fat (r=0.71, P <0.0001 for sum). Somewhat unexpectedly, BMI had no correlation with either posterior (r=0.12, P=0.221 for sum) or anterior epidural fat (r=0.11, P=0.271 for sum), and most importantly the presence of obesity was associated only with subcutaneous fat (P <0.0001), but not with any specific or summated epidural fat measurement (P=0.0801-0.7692). CONCLUSION: Weight but not body habitus is associated with specific, usually posterior, patterns of epidural fat deposition. Overall obesity is unrelated to epidural fat. PMID- 15665693 TI - Magnetic resonance appearance of bone marrow edema associated with hydroxyapatite deposition disease without cortical erosion. AB - The authors report the magnetic resonance (MR) appearance of bone marrow edema associated with hydroxyapatite deposition disease without cortical erosion. Hydroxyapatite deposition disease may have bone marrow edema on MR imaging without radiographic evidence of cortical erosion, mimicking the appearance of fracture, neoplasm, or infection. Awareness of this association can prevent unnecessary additional imaging evaluation or biopsy. PMID- 15665694 TI - Proliferative myositis: value of imaging. AB - Imaging findings in a 52-year-old woman with proven proliferative myositis are described. US revealed preservation of continuous muscle bundles and patchy areas of hyperechogenicity, containing hypoechoic lines. MRI showed at T2-w sequences an ill-defined, hyperintense, intramuscular lesion, containing isointense lines. Subtotal enhancement, a nonenhancing geometrical web, and fascial enhancement were noted. In patients with painful growing masses, US and MRI correlation may suggest the diagnosis of proliferative myositis leading to biopsy, thus avoiding mutilating surgery. PMID- 15665695 TI - Differential diagnosis between cerebral tuberculosis and neurocysticercosis by magnetic resonance spectroscopy. AB - Single enhancing brain lesions (SELs), mostly as a result of neurocysticercosis or tuberculosis, are a common cause of seizures. Ten patients with SELs caused by neurocysticercosis (n=6) or tuberculosis (n=4) were examined by proton magnetic resonance spectroscopy. Tuberculomas had a high peak of lipids, more choline, and less N-acetylaspartate and creatine. The choline/creatine ratio was greater than 1 in all tuberculomas but in none of the cysticerci. Magnetic resonance spectroscopy differentiates SELs caused by cysticercosis or tuberculosis and may avoid brain biopsies or unnecessary antituberculosis treatments. PMID- 15665696 TI - Evaluation of extraocular muscles using dynamic contrast enhanced MRI in patients with chronic thyroid orbitopathy. AB - OBJECTIVE: The purpose of the current study is to investigate the underlying pathophysiological changes of extraocular muscle (EOM) in Graves orbitopathy using dynamic contrast MR imaging and to correlate these MR functional changes with the anatomic abnormalities. METHODS: EOMs of 16 patients with Graves disease and 12 normal volunteers were examined by dynamic enhanced MRI. We quantified the peak enhancement ratio of EOMs and calculated the ratio versus temporalis muscle and the ratio of maximum upslope versus temporalis muscle. We compared the ratios between normal volunteers and patient groups. RESULTS: Mean of peak enhancement ratio values for the EOMs in patients with Graves disease tends to decrease according to the severity of the anatomic and clinical changes. The mean maximum upslope also decreased according to the severity of the disease for EOMs. CONCLUSION: Hemodynamic information obtained by dynamic contrast enhanced MRI is useful in evaluating the clinical course of thyroid orbitopathy. PMID- 15665697 TI - Acinic cell carcinoma of the head and neck: radiologic-pathologic correlation. AB - OBJECTIVE: To describe and correlate the imaging and pathologic findings of acinic cell carcinoma (ACC) in the head and neck. METHODS: We reviewed the radiologic findings of 12 patients with pathologically proven ACC in the head and neck. They were 6 males and 6 females (ages: 5-75 years, mean 36 years) who undergoing computed tomography (CT, n=9) and CT with magnetic resonance (MR) imaging (n=3). RESULTS: The lesions in the superficial lobe of the parotid gland were solid (n=7), cystic (n=1), and cystic mass with mural nodule (n=1) on CT. A parapharyngeal lesion was cystic mass with mural nodule, and a submandibular and a palate tumor were cystic lesions on CT. All solid masses in the parotid gland (n=7) included focal low-attenuating portions on CT, which were microcyst, hemorrhage, or necrosis on pathologic examination. We could not find intratumoral calcifications or metastatic lymphadenopathy on imaging and histologic studies in all 12 cases. Internal hemorrhage on the MR images was seen in a parapharyngeal and a parotid lesion. CONCLUSION: Although ACC appears to have nonspecific imaging findings, familiarity with some imaging features can be helpful for differential diagnosis of head and neck tumors. PMID- 15665698 TI - Diffusion tensor tractography of gliomatosis cerebri: fiber tracking through the tumor. AB - In previous reports, tracts obtained by diffusion tensor (DT) fiber tracking were terminated or deviated by the brain tumors or surrounding edema. There has been no report showing diffusion tensor tractography penetrating through the tumor. A case of glioma is reported, whose DT fiber tract passing through the tumor was observed by changing the threshold of fractional anisotropy. PMID- 15665699 TI - Rhabdomyoma of the head and neck demonstrated by prenatal magnetic resonance imaging. AB - A case of fetal rhabdomyoma (myxoid type) of the head and neck demonstrated on prenatal magnetic resonance imaging (MRI) is presented. This benign tumor of skeletal muscle is uncommon and should not be confused with its malignant counterpart-rhabdomyosarcoma. With the increasing use of ultrafast MRI, the radiologist is more likely to encounter head and neck masses in the fetus. PMID- 15665700 TI - Cervical radiculopathy caused by vertebral artery loop formation: multislice computed tomography angiography findings. AB - Vertebral artery tortuosity and loop formation are rare causes of cervical radiculopathy. The criterion standard for the detection of vertebral artery loop formation is digital subtraction angiography. Computed tomography (CT) and magnetic resonance imaging provide good accuracy in the evaluation of vertebral arteries. A case of vertebral artery loop formation is reported. The diagnosis was confirmed by 16-row multisection CT angiography findings of vertebral artery loop formation. PMID- 15665701 TI - Propylene glycol is essential in the LCModel basis set for pediatric 1H-MRS. AB - Propylene glycol (PG) is an organic solvent used in a variety of parenterally injected pharmaceutical preparations. In this current study, we analyzed neonatal brains following routine MRS evaluation at 1.5 T, using the LCModel software, with and without the addition of PG to the analysis basis-set. We suggest including PG in the LCModel basis-set for neonatal brain H-MRS to improve the discrimination and quantification of lactate and inositol. PMID- 15665702 TI - Dose reduction in multislice computed tomography. AB - OBJECTIVE: Evaluation of the attenuation-based on-line modulation of tube current on multislice computed tomography (CT) to explore the potential of this dose saving technique. METHODS: Fifty-five patients with follow-up CT examinations were scanned without and with a CARE Dose (Siemens Medical Solutions, Erlangen, Germany). The applied dose, image noise, and subjective image quality were evaluated. The reduction in patient exposure was determined by obtaining the effective milliamperes per second for each reconstructed scan and the absolute dosage requirement for the whole scan. RESULTS: The dose reduction achieved by applying the attenuation-based on-line modulation of tube current was 29.4% +/- 3.1% (P=0.002) for all scans, 30.5% +/- 3.2% (P=0.002) for the thorax scans, 29.7% +/- 2.9% (P=0.002) for the abdomen scans, and 28.7% +/- 2.7% (P=0.003) for the thorax and abdomen scans together. No significant restrictions in image quality were observed. CONCLUSIONS: Remarkable dose reduction can be obtained using the attenuation-based on-line modulation of tube current on multislice CT without compromising image quality. PMID- 15665703 TI - Six revolutions in vaccinology. AB - The history of vaccine development can be divided into 5 waves, produced by revolutions in technology. They are attenuation, inactivation, cell culture of viruses, genetic engineering and methods to induce cellular immune responses. This division is somewhat artificial, and all of the past strategies continue to be useful. I discuss the candidates for the sixth revolution, which include combination vaccines, new adjuvants, proteomics, reverse vaccinology and vaccines for noninfectious diseases, among others. I propose new delivery systems as the most likely to succeed, although humbly admitting that prediction is always subject to error. PMID- 15665704 TI - Impact of pneumococcal conjugate vaccine and the severity of winter influenza like illnesses on invasive pneumococcal infections in children and adults. AB - BACKGROUND: Pneumococcal conjugate vaccine (PCV7), recommended in July 2000 for routine use in infants, has resulted in a reduction in the rate of invasive pneumococcal disease (IPD) in young children. We studied the impact of PCV7 and the possible contribution of the severity of influenza-like respiratory infection season on the rate of IPD on children and adults. METHODS: In 7 hospitals of a health system, episodes of IPD were identified by the microbiology laboratories during the 2-year period before July 2000 and the 4-year period after July 2000 during routine use of PCV7, and patient records were reviewed. Episodes of influenza-like illnesses during each winter in a local county were prospectively measured by reports from all acute care hospitals and episodes of absenteeism resulting from influenza-like illnesses from all schools. RESULTS: There were 720 patients with blood and/or cerebrospinal fluid isolates of Streptococcus pneumoniae. There were significant reductions in cases of IPD in children younger than 2 years, 68% reduction; children 2-4 years of age, 70%; adults 18-49 years of age, 42%; and adults older than 64 years, 30%. Annually, during the PCV7 period, there were significantly fewer episodes of influenza-like illnesses than during the pre-PCV7 years. CONCLUSIONS: PCV7 efficacy and resultant herd-type immunity resulted in a reduction in the rate of IPD not only in young children but also in young and elderly adults. Milder winter respiratory viral seasons may possibly have contributed to the observed reduction in the rate of IPD. PMID- 15665705 TI - Population-based surveillance for childhood invasive pneumococcal disease in the era of conjugate vaccine. AB - BACKGROUND: Heptavalent pneumococcal conjugate vaccine was licensed in the United States in February 2000 and, following national guidelines, universally distributed in Massachusetts starting in July 2000 to children younger than 2 years of age and selected children 2-5 years of age. We performed statewide surveillance for all cases of invasive pneumococcal disease (IPD) in children younger than 18 years of age to determine risk features and contribution of vaccine failure to ongoing pneumococcal invasive disease. METHODS: Massachusetts pediatric IPD cases were identified via enhanced passive surveillance of microbiology laboratory reports of pneumococcal isolates from sterile body sites of children younger than 18 years for 2 years starting in October 2001. Serotyping was performed on isolates of Streptococcus pneumoniae from normally sterile body fluid. Case demographic and clinical data (including dates of prior doses of PCV7) were collected via follow-up telephone interviews with case primary care providers and/or parents. RESULTS: Between October 1, 2001 and September 30, 2003, 191 cases of IPD were identified statewide (138 in children younger than 5 years). Annual incidence rate for IPD was 17.4 per 100,000 children younger than 5 years, representing a decline of 69% when compared with annual incidence rate of 56.9 per 100,000 from Massachusetts statewide active surveillance performed 1990-1991. In 2001-2003, 30% of cases occurred in the first year of life (36.5 per 100,000), representing a 7.8-fold increased risk compared with children older than 1 year of age. Race-specific annual incidence rates in blacks and Hispanics were 2.3-fold (95% confidence interval, 1.21-4.42) and 1.9-fold (95% confidence interval, 1.06-3.37), greater than in whites. Fifty nine cases were reported to have underlying comorbid conditions. Serotyping was available for 136 of 191 (71%) cases younger than 18 years; of isolates available for serotyping, 40 (29%) were vaccine serotype (ST), 31 (23%) vaccine-related ST and 65 (48%) nonvaccine ST. Seven of 40 cases with IPD caused by vaccine ST received at least 3 doses of PCV7 vaccine before IPD. CONCLUSIONS: Universal administration of PCV7 to children younger than 2 years of age and selective administration to children 2-5 years of age has resulted in a significant decline in IPD in Massachusetts. Children younger than 1 year of age, African American and Hispanic children and those with recognized comorbid illnesses (malignancy, human immunodeficiency virus, immune deficiency, nephrotic syndrome, etc.) continue to remain at risk for IPD. These risk features should be considered when evaluating febrile infants and children. PMID- 15665706 TI - Impact of amoxicillin on pneumococcal colonization compared with other therapies for acute otitis media. AB - BACKGROUND: This study compared the effects of 4 outpatient antibiotic regimens on colonization by penicillin-susceptible and -nonsusceptible pneumococci to assess their relative potential to promote colonization with Streptococcus pneumoniae with reduced susceptibility to penicillin. METHODS: Children presenting with acute otitis media were randomized to receive amoxicillin, cefprozil, ceftriaxone or azithromycin. Nasopharyngeal specimens were collected on days 0, 3-5, 10-14 and 28-30 and assessed for the presence of S. pneumoniae. At each visit, the proportions of penicillin-susceptible and -nonsusceptible pneumococci were compared among treatment groups. RESULTS: Among 1009 enrollees, the prevalence of colonization by S. pneumoniae at baseline was 23.5%, of which 41.1% were penicillin-nonsusceptible. Colonization by nonsusceptible pneumococci was unaltered during the observation period in all treatment groups, with no detectable differences among groups at each visit. By contrast, there was a substantial reduction in the prevalence of colonization by penicillin-susceptible organisms, most notably in subjects treated with amoxicillin. This resulted in a proportional shift toward resistant organism colonization in all groups, with this shift being significantly more pronounced among amoxicillin recipients than in the other groups at 10-12 days (P < 0.02 for each comparison with amoxicillin). CONCLUSIONS: Treatment with amoxicillin for acute otitis media resulted in a larger shift toward nonsusceptible organism colonization among those children still colonized postexposure than did treatment with 3 comparison agents. This phenomenon raises theoretical concerns that at the population level, amoxicillin produces conditions that promote the dissemination of the nonsusceptible phenotype more readily than other outpatient antibiotics. Confirmation of these results requires further study. PMID- 15665708 TI - Pharmacokinetics of intravenously administered azithromycin in pediatric patients. AB - BACKGROUND: The objective of this study was to characterize the pharmacokinetics and tolerance of a single intravenous (IV) azithromycin dose in children. METHODS: Subjects were stratified into 4 age groups: 0.5-2 years; >2-<6 years; 6 <12 years; and 12-<16 years. Each subject received a single 10 mg/kg dose (500 mg maximum) infused in 1 hour. Serial venous blood samples were obtained for a 168 hour period, and laboratory safety evaluations were performed immediately preceding azithromycin administration and at the conclusion of the study. Serum azithromycin concentrations were quantified with a validated high performance liquid chromatography method with mass spectrometric detection. Pharmacokinetic indices were calculated for each subject by noncompartmental techniques. RESULTS: Thirty-two subjects (6.7 +/- 5.0 years, 11 boys) participated. Mean serum concentration-time data were comparable for the 4 age groups. For all subjects with evaluable data, the mean area under the curve from 0 to 72 hours (AUC0-72) was 8.2 microg . h/mL (n = 26), the maximum concentration (Cmax) was 2.4 microg/mL and the elimination half-life (t1/2) was 65.2 hours (n = 25). The AUC0 72 and Cmax were not associated with age. The dose was well-tolerated with no serious adverse events. CONCLUSION: The disposition of azithromycin after a single 10-mg/kg IV dose (maximum labeled adult dose of 500 mg) is comparable in pediatric patients between 0.5 and 16 years of age. These pharmacokinetic data can be used to guide dose selection for future therapeutic trials of IV azithromycin in pediatric patients. PMID- 15665707 TI - Quantitation of adenovirus genome during acute infection in normal children. AB - BACKGROUND: Adenovirus infection causes a wide range of clinical illness in normal children. New molecular techniques allow quantitation of viral genome to study the natural history of adenovirus infection and viral load in normal children. METHODS: Clinical samples were collected from 38 previously healthy, febrile children, and viral cultures were performed. Quantitative polymerase chain reaction (PCR) was used to detect adenovirus genome and to determine viral load. Adenovirus isolates were genotyped with a PCR-based assay. RESULTS: Adenovirus culture was positive in 6 children who were diagnosed with acute adenovirus infection. Throat swabs contained high copy numbers of adenovirus genome (1.6 x 10(6)-6 x 10(7) copies/swab) from 4 of 4 adenovirus culture positive children. Only 2 of 32 adenovirus culture-negative children had detectable adenovirus genome from throat swabs, but with a lower copy number (8 x 10(2) copies/swab). Adenovirus genome was not detected in blood samples from 5 of 6 adenovirus culture-positive children with uncomplicated upper respiratory tract infection and from all adenovirus culture-negative children. High level viremia (1.8 x 10(8)/ml) was detected in an adenovirus culture-positive 6-month-old infant with fever, pneumonia, conjunctivitis and hepatitis. Subsequent reduction in viral load paralleled her clinical recovery. Adenovirus viruria (1 x 10(9) copies/ml) with normal urinanalysis was detected in another adenovirus culture positive child. All 6 adenovirus isolates were genotyped as adenovirus type 7h. CONCLUSION: Viral load assessment in clinical samples determined by quantitative PCR can be useful in the diagnosis of adenovirus infection in immunocompetent, febrile children. PMID- 15665709 TI - Clinical features and genotyping analysis of community-acquired methicillin resistant Staphylococcus aureus infections in Taiwanese children. AB - BACKGROUND: Community-acquired methicillin-resistant Staphylococcus aureus (CA MRSA) infection in children without health care-associated risk factors has emerged. To evaluate the clinical features and genotyping analysis of CA-MRSA in children in Taiwan, we conducted this study. METHODS: Between July 2000 and June 2001, 198 episodes of S. aureus infection were identified in 191 children hospitalized at Chang-Gung Children's Hospital. The medical records of these children were retrospectively reviewed. The MRSA clinical isolates from each episode of infection, if available, were collected for pulsed-field gel electrophoresis (PFGE) typing and staphylococcal cassette chromosome mec (SCCmec) type determination. RESULTS: Among the 198 episodes of S. aureus infection, MRSA accounted for 47% of 114 CA infections and 62% of 84 hospital-acquired (HA) infections, respectively. Among 54 CA-MRSA infections, 32 episodes (59%) occurred in children without risk factors. Similar to HA-MRSA isolates, these CA-MRSA isolates were also multiresistant, with a high rate of resistance to clindamycin (93%) and erythromycin (94%), but 91% of them were susceptible to trimethoprim/sulfamethoxazole. Superficial soft tissue infection was the most common presentation, accounting for 65% of CA-MRSA infections. All 54 children with CA-MRSA infection recovered without sequelae, though 63% of 35 children with superficial soft tissue infection were treated with in vitro nonsusceptible antibiotics. Of the 83 clinical isolates (40 CA, 22 with no identified risk) available for analysis, 6 genotypes with 30 subtypes were identified. Three major PFGE patterns were identified, accounting for 94% of the isolates (92.5% for CA isolates, 95% for HA isolates). Type IV SCCmec was carried by 25 and 40% of CA and HA isolates, respectively. CONCLUSION: In Taiwan, MRSA was prevalent among pathogens of CA infections in children, and these CA isolates were multiresistant and genetically associated with HA isolates. In an area with a high prevalence of methicillin resistance, for children with putative S. aureus infection, even community-acquired, a glycopeptide-containing regimen should be considered for initial empirical therapy in the case of serious infection. PMID- 15665711 TI - Comparative trial of the safety and immunogenicity of quadrivalent (A, C, Y, W 135) meningococcal polysaccharide-diphtheria conjugate vaccine versus quadrivalent polysaccharide vaccine in two- to ten-year-old children. AB - BACKGROUND: A quadrivalent meningococcal diphtheria conjugate vaccine (MCV-4) has been developed to provide T-cell dependent immune responses against 4 major disease-causing serogroups (A, C, Y, W-135). METHODS: In a comparative, randomized, modified double blind, controlled study in healthy 2- to 10-year-old U.S. children, safety and immunogenicity profiles of MCV-4 (n = 696) were compared with those of a licensed quadrivalent polysaccharide vaccine, Menomune A/C/Y/W-135 (PSV-4, n = 702). Vaccine-related adverse reactions were assessed for 28-day and 6-month follow-up periods. Serum bactericidal activity (SBA) was assayed in prevaccination, day 28 and 6-month postvaccination sera samples. RESULTS: Both vaccines were well-tolerated, with no vaccine-related serious adverse events and similar rates of mostly mild local and systemic reactions. Functional antibody (SBA) seroconversion percentages were significantly higher for all 4 serogroups in the MCV-4 group. The SBA geometric mean titers against serogroups A, C, Y and W-135 with MCV-4 were 1700, 354, 637 and 750, respectively, compared with PSV-4 (893, 231, 408 and 426) 28 days postvaccination (P < 0.001 for all comparisons). This significant difference persisted through 6 months. CONCLUSIONS: In 2- to 10-year-old children MCV-4 had a safety profile similar to that of PSV-4 and elicited significantly higher and more persistent serum bactericidal antibody responses against meningococcal serogroups A, C, Y and W-135 than did the licensed polysaccharide vaccine. PMID- 15665710 TI - Morbidity and mortality during the first two years of life among uninfected children born to human immunodeficiency virus type 1-infected women: the women and infants transmission study. AB - OBJECTIVE: We evaluated morbidity and mortality during the first 2 years of life among children born to human immunodeficiency virus-(HIV) type 1-infected women enrolled in the Women and Infants Transmission Study (WITS) during an 11-year period (1990-2001). DESIGN AND METHODS: As part of WITS, evaluations were performed at birth and at 1, 2, 4, 6, 9, 12, 18 and 24 months of age. Growth, hospitalization and the incidence of clinical disease were assessed regularly. RESULTS: Data regarding 1118 children born to HIV-infected women (955 HIV uninfected children and 163 HIV-infected children) were analyzed. Fewer changes in the caretaker of the child and fewer in utero exposures to drugs, tobacco and alcohol occurred in the latter periods of the study (all P values for time trend analyses <0.01). The percentages of HIV-uninfected children with poor weight gain (44 of 767; 5.7%), short stature (32 of 703; 4.5%) and wasting (27 of 792; 3.4%) were higher than expected for the general population. Two or more changes in caretaker were associated with all growth deficiencies except wasting, and fetal exposure to tobacco was associated with height abnormalities. Anemia was common and was associated with receipt of zidovudine prophylaxis. Morbidity and mortality decreased during the study period. For the uninfected children, a decrease in class A events (Kaplan-Meier rates: group 1, 22.3%; group 2, 6.8%; group 3, 4.2%; P < 0.001) and class C events and death (Kaplan- Meier event rates: group 1, 2.0%; group 2, 1.7%; group 3, 0.2%; P = 0.062) during the first 2 years of life account for the differences in the curves over time. CONCLUSIONS: During an 11-year period, morbidity and mortality during the first 24 months of life decreased substantially for children born to HIV-infected women. PMID- 15665712 TI - Oculorespiratory syndrome after influenza immunization in children. AB - BACKGROUND: During the 2000-2001 season, an oculorespiratory syndrome (ORS) was identified in association with 1 manufacturer's influenza vaccine in Canada. ORS included bilateral red eyes, facial edema or respiratory symptoms beginning within 24 hours of influenza immunization. Few reports involved children. We assessed the rate of ORS in a pediatric cohort. METHODS: In February 2001, invitation to participate in an influenza vaccine safety survey was sent to the parents of children who attend a diabetes clinic. A questionnaire elicited influenza immunization history and adverse event experience beginning within 1 day of vaccination for the child with diabetes and up to 3 siblings. Follow-up telephone interviews collected information from those who failed to return a questionnaire by mail. RESULTS: Of the 959 households sent a questionnaire, 780 participated (81%). Among 780 children with diabetes, 418 (54%) received influenza vaccine in 2000. Adverse event experience was collected from an additional 242 immunized siblings. Among immunized children, 13% (95% confidence interval, 10-16%) experienced ORS, consisting primarily of respiratory symptoms. The majority (72%) resolved within 2 days. There was association between ORS and first time receipt of influenza vaccine (OR 2.7; 95% confidence interval, 1.6 4.4). Fewer parents of ORS-affected compared with unaffected children indicated that they were likely or very likely to have their child revaccinated (87 versus 97%; P < 0.001). CONCLUSIONS: In 2000-2001, children experienced ORS in Canada at a rate comparable with that of adults. ORS should be incorporated into influenza vaccine safety monitoring and discussed with parents, especially those contemplating influenza vaccine for their child for the first time. PMID- 15665713 TI - Immunogenicity and reactogenicity of four doses of diphtheria-tetanus-three component acellular pertussis-hepatitis B-inactivated polio virus-Haemophilus influenzae type b vaccine coadministered with 7-valent pneumococcal conjugate Vaccine. AB - BACKGROUND: The 7-valent pneumococcal (7vPn) conjugate vaccine is licensed for primary and booster vaccination according to the same immunization schedules as routinely recommended diphtheria-tetanus-pertussis-based childhood vaccines and can be coadministered during the same vaccination visit. METHODS: An open, randomized study evaluated the immunogenicity and safety of a hexavalent diphtheria-tetanus-acellular pertussis-hepatitis B-inactivated polio virus Haemophilus influenzae type b (DTPa-HBV-IPV/Hib) vaccine and a 7vPn conjugate vaccine when coadministered at 2, 3 and 4 months and 12-23 months of age, compared with the administration of the hexavalent DTPa-HBV-IPV/Hib vaccine given alone. Serum antibody titers were measured before and 1 month after the primary course and before and 1 month after the booster dose. Solicited local and general adverse events were recorded for 4 days and unsolicited adverse events were recorded for 30 days after each vaccine dose. RESULTS: A total of 345 subjects were enrolled for primary vaccination with the hexavalent vaccine (170 without and 175 with the 7vPn vaccine coadministered) and 266 returned for booster vaccination (122 without and 144 with coadministration of the 7vPn vaccine). After primary vaccination, antibody responses against the common antigens were similar in both groups, with seroprotection rates of 93.6-100% and with similar antibody decay before booster vaccination. The fourth dose induced a vigorous booster response, with seroprotection/vaccine response rates of 96.8-100%. Response to the 7vPn primary and booster vaccination was within previously reported ranges. Differences in reactogenicity resulted from higher incidences of symptoms after concomitant vaccination. Rectal temperature >39.5 degrees C was observed after 1.2% of the coadministered vaccine doses during primary vaccination and after 2.8% of the booster vaccine doses. CONCLUSION: Coadministration of the DTPa-HBV-IPV/Hib and 7vPn vaccines at separate injection sites during the same vaccination visit was effective and safe. PMID- 15665714 TI - Staphylococcus aureus decolonization. PMID- 15665716 TI - Encephalopathy after bone marrow transplantation. PMID- 15665715 TI - Human herpesvirus-8: clinical features of an emerging viral pathogen. PMID- 15665717 TI - Reduced immunogenicity of diphtheria and tetanus toxoids when combined with pertussis toxoid. AB - The effect of pertussis toxoid on the immunogenicity of diphtheria and tetanus toxoids (DT) was studied during a double blind efficacy trial of an acellular pertussis vaccine. Infants received DT with or without pertussis toxoid at 3, 5 and 12 months of age. Geometric mean concentrations were higher in the DT than in the DT-pertussis toxoid group 1 month (diphtheria toxoid 4.76 versus 3.58 IU/mL, P = 0.009; tetanus toxoid 4.42 versus 2.66 IU/mL, P < 0.0001) and 2 years after the third injection (diphtheria toxoid 0.15 versus 0.10 IU/mL, P < 0.0001; tetanus toxoid 0.38 versus 0.18 IU/mL, P < 0.0001). Pertussis toxoid causes a small but significant reduction of the immunogenicity of diphtheria toxoid and tetanus toxoid. PMID- 15665718 TI - Cryptosporidium infection in diarrheic children in southeastern Iran. AB - In a cross-sectional study conducted in children referred to Bandar Abbas Pediatric Hospital in southeastern Iran, the prevalence of Cryptosporidium infection was 7%. Diarrhea lasted significantly longer in children infected with Cryptosporidium. There were also a significant association between Cryptosporidium infection and underweight children and no association with parent's occupation, breast-feeding, source of drinking water, vicinity or presence of sewage or animal exposure. PMID- 15665719 TI - Vertical transmission of human echovirus 11 at the time of Bornholm disease in late pregnancy. AB - We report a case of human echovirus 11 infection in a neonate at the time of Bornholm disease in late pregnancy of the mother. Mother and baby were found to be infected with the same virus, with a combination of diagnostic virologic techniques, demonstrating likely transmission from mother to baby around the time of delivery. PMID- 15665720 TI - A boy with nosocomial malaria tropica contracted in a Dutch hospital. AB - A 12-year-old Dutch boy was admitted because of severe neurotrauma after a traffic accident. On day 33 after admission, a Plasmodium falciparum infection was found in a routine blood smear. Most probably he was infected by blood of a patient next to him, a girl with severe malaria tropica. The genotype of the P. falciparum in both patients was identical. PMID- 15665721 TI - Candida epiglottitis in an adolescent with acquired immunodeficiency syndrome. AB - We report a case of an adolescent with human immunodeficiency virus infection who presented with epiglottitis caused by Candida albicans. This patient's symptom was severe oropharyngeal thrush without the classic manifestations of epiglottitis. Atypical organisms can cause epiglottitis in immunocompromised adolescents and they can have mild symptoms. PMID- 15665722 TI - Soft tissue infection caused by Kluyvera species. PMID- 15665723 TI - Calcium/calmodulin-dependent kinase II is required for platelet-activating factor priming. AB - Platelet-activating factor (PAF) primes the macrophage proinflammatory response to inflammatory stimuli, such as lipopolysaccharide (LPS). The cellular events responsible for this priming or reprogramming remain unresolved, but may occur through an increase in cytosolic calcium, inducing calcium/calmodulin-dependent kinase (CaMK) activation. To study this, differentiated THP-1 cells were used to study the effect of CaMK II and IV inhibition on PAF-induced reprogramming of TLR4-mediated events. LPS induced p38, ERK 1/2, and JNK/SAPK phosphorylation, NF kappaB and AP-1 activation, and TNF-alpha and IL-10 production. PAF pretreatment selectively increased LPS-induced ERK 1/2, JNK/SAPK, NF-kappaB and AP-1 activation, and TNF-alpha production. Inhibition of CaMK II prevented PAF-induced priming of these events. Inhibition of CaMK IV prevented LPS-induced ERK 1/2, JNK/SAPK, NF-kappaB and AP-1 activation, and TNF-alpha production, but increased IL-10 production with or without PAF pretreatment. Neither CaMK II nor IV inhibition had any affect on p38 activity. These data suggest that the function of CaMK II is essential for PAF-induced macrophage priming. This priming event is mediated in part by modulation of ERK 1/2, JNK/SAPK, NF-kappaB, and AP-1 activation. CaMK IV, on the other hand, is not specific for priming by PAF and appears to have a direct link in TLR4-mediated events. PMID- 15665724 TI - Inhibitors of NADPH oxidase reduce the organ injury in hemorrhagic shock. AB - Reactive oxygen species contribute to the multiple organ dysfunction syndrome in hemorrhagic shock. Here, we investigate the effects of two chemically distinct inhibitors of NADPH oxidase on the circulatory failure and the organ dysfunction and injury associated with hemorrhagic shock in the anesthetized rat. Hemorrhage (sufficient to lower mean arterial blood pressure of 45 mmHg for 90 min) and subsequent resuscitation with shed blood resulted (within 4 h after resuscitation) in a delayed fall in blood pressure and in renal dysfunction and liver injury. Treatment of rats upon resuscitation with the NADPH oxidase inhibitors diphenylene iodonium (DPI, 1 mg/kg i.v.) reduced renal dysfunction and liver injury, whereas apocynin (3 mg/kg i.p.) did reduce the liver injury, but not the renal dysfunction caused by hemorrhagic shock. DPI and apocynin also attenuated the lung and intestinal injury (determined by histology) caused by hemorrhage and resuscitation. In the liver, DPI and apocynin abolished the increase in the formation of superoxide anions associated with hemorrhagic shock. However, neither DPI nor apocynin had a significant effect on the delayed circulatory failure caused by hemorrhage and resuscitation. In addition, DPI and apocynin did not reduce the increase in nitric oxide synthesis caused by hemorrhagic shock. Moreover, DPI reduced the activation of the transcription factor activator protein-1 caused by severe hemorrhage and resuscitation in the liver. Thus, we propose that an enhanced formation of superoxide anions by NADPH oxidase contributes to the liver injury caused by hemorrhagic shock, and that inhibitors of NADPH oxidase may represent a novel therapeutic approach for the therapy of hemorrhagic shock. PMID- 15665725 TI - Effect of thermal injury on splenic myelopoiesis. AB - Thermal injury increases the number of macrophage progenitors in the bone marrow but leads to a decrease in the number of granulocyte progenitors. In the spleen, thermal injury increases the numbers of myeloid progenitors, but the lineage commitment of these cells is unknown. In this study mice were given a scald burn, and the number of splenic myeloid progenitors as well as their progeny was determined. BrdU uptake was used to monitor the de novo production of splenocytes for 8 days after the burn. Burn injury increased the numbers of splenic granulocyte-macrophage (GM), granulocyte (G), and macrophage (M) progenitors at postburn day 8 by 12-, 11-, and 18-fold, respectively. Scald injury increased the number of mature PMN (CD11b GR1(bright)) in the spleen and increased the number of white pulp monocyte/macrophages. Increased numbers of BrdU-positive PMN and monocyte/macrophages were seen after injury. Burn macrophages produced increased levels of the anti-inflammatory hematopoietic cytokine G-CSF. Our work clearly shows that the increased myelopoiesis observed postinjury leads to the production of mature myeloid cells. However, the effects of thermal injury on progenitors in the spleen and marrow are not equivalent. PMID- 15665726 TI - Genetic variability in the immune-inflammatory response after major burn injury. AB - Thermal injury induces immune dysfunction and alters numerous physiological parameters. Studies have proposed that genetics influence the outcome after traumatic injury and/or sepsis, however, the contribution of genetics to the immune-inflammatory response postburn has not been investigated. In this study, mice of three distinct genetic backgrounds (C57BL/6NCrlBR, BALB/cAnNCrlBR, and 129S6/SvEvTac) were subjected to thermal injury or a sham procedure, and 3 days later, blood and splenic immune cells (splenocytes and macrophages) were isolated for analysis. Splenocytes from the C57BL/6NCrlBR strain displayed suppressed splenic T cell proliferation postinjury, whereas the other strains were unaffected. Burn injury also induced a shift toward a Th2-type T-cell response (suppressed IFN-gamma production) in the C57BL/6NCrlBR strain, but not in the other strains. Macrophages from C57BL/6NCrlBR and 129S6/SvEvTac mice were highly proinflammatory with elevated productive capacity for TNF-alpha and nitric oxide, whereas no such changes were observed in macrophages for BALB/cNCrlBR mice. C57BL/6NCRLBR macrophages produced increased IL-10 levels postburn, and BALB/cNCrlBR macrophages had suppressed IL-10 production postinjury. No differences in fasting blood glucose and insulin were observed after thermal injury. However, significant postburn weight loss was observed in the BALB/cNCrlBR and 129S6/SvEvTac strains, but not in the C57BL/6NCrlBR strain. In summary, these findings support the concept that the immune-inflammatory response postburn is influenced by genetic make-up. Further elucidation of the influence of genetics under such conditions is likely to contribute to the improvement in existing, and development of new, therapeutic regimes for burn patients. PMID- 15665727 TI - The development of acute respiratory distress syndrome after gut ischemia/reperfusion injury followed by fecal peritonitis in pigs: a clinically relevant model. AB - Numerous clinical trials using anti-inflammatory agents for patients with acute respiratory distress syndrome (ARDS) have failed despite efficacy in acute animal models. This underscores the necessity of developing a clinically relevant model of ARDS. Initially, we attempted to induce lung injury in pigs by fecal peritonitis only. When this was unsuccessful, we designed a two-hit model of ischemia/reperfusion (I/R) injury followed by fecal peritonitis to create a clinically applicable model of ARDS. The initial study consisted of Yorkshire swine [group 1, fecal clot (FC), n = 4] that were followed clinically after intraperitoneal placement of a fecal (0.5 mL/kg) blood (2 mL/kg) clot. Blood was sampled daily for cultures, a complete blood count, a lactate level, and various cytokine expression determined by enzyme-linked immunosorbent assay (ELISA). Pigs were treated with antibiotics and fluids, placed on a ventilator before sacrifice to obtain hemodynamic and pulmonary parameters, and underwent histologic lung assessment. Additionally, bronchoalveolar lavage fluid was obtained for protein concentration and cytokine levels. Once it was evident that no lung injury had occurred, we designed a more severe model. A second group of Yorkshire swine [group 2, superior mesenteric artery (SMA) + FC, n = 4] underwent SMA occlusion for 30 min (I/R) followed by intraperitoneal placement of a FC as in the initial group. These pigs were monitored more invasively and continuously in an intensive care setting for 48 h and followed, treated, and assessed in a similar fashion to group 1. Group 1 (FC) pigs survived 9 days and showed signs of sepsis (bacteremia with polymicrobial organisms), an inflammatory response in the form of elevated cytokines, yet no physiologic or histologic evidence of lung injury. Group 2 (SMA + FC) pigs demonstrated more severe sepsis, a significantly increased cytokine response compared with animals in the FC group, and physiologic signs of progressive pulmonary injury. Pigs in the SMA + FC group were sacrificed at 48 h after clinical deterioration (significant decline in oxygenation) and demonstrated pathologic evidence of lung injury indicated by increased bronchoalveolar lavage fluid protein, diffuse and thickened alveolar septae, hyaline membrane formation, and pulmonary edema. The addition of a second "hit" (SMA occlusion, I/R) to a FC sepsis model resulted in severe lung injury that developed within a 3-day period. To our knowledge, this is the first large animal experiment that definitively and consistently causes insidious onset ARDS in pigs. By closely paralleling the clinical development of pulmonary injury, this model should prove invaluable in the study of human ARDS. PMID- 15665728 TI - Effects of manganese superoxide dismutase nebulization on pulmonary function in an ovine model of acute lung injury. AB - Smoke inhalation injury is a major cause of morbidity and mortality in thermally injured individuals. There is evidence of increased oxygen free radical activity, e.g., superoxide, in association with smoke inhalation injury. Because superoxide dismutase converts the reactive superoxide radical to peroxide, we hypothesized that nebulization of manganese superoxide dismutase (Mn-SOD) into the airway might attenuate pulmonary dysfunction secondary to smoke inhalation injury. The present study was designed as a prospective, controlled, and randomized laboratory experiment to determine the effects of aerosolized Mn-SOD on lung fluid balance, as indexed by changes in pulmonary microvascular permeability, lung lymph flow (Q(L)), and gas exchange in an established and clinically relevant ovine model of smoke inhalation injury. Fifteen female Merino sheep were chronically instrumented with a femoral arterial, a Swan-Ganz, and a left atrial catheter. In addition, the right caudal mediastinal lymph node was cannulated to measure Q(L) (mL.h(-1)). Pneumatic occluders were placed around the right pulmonary veins for the determination of the reflection coefficient (sigma). After 7 days of recovery, sheep were randomly allocated to (a) an untreated control group (4 groups of 12 breaths of cotton smoke), (b) an injured group treated with nebulized Mn-SOD (5 mg/kg), and (c) an injured group that received only the vehicle (nebulized saline). Nebulization was performed 1 h and 12 h after smoke inhalation. Mn-SOD nebulization attenuated the increase in both filtration coefficient and sigma and significantly decreased lung tissue conjugated dienes. However, there were no differences in Q(L), PaO2/FiO2 ratio, and bloodless lung wet/dry weight ratio between groups. Although Mn-SOD nebulization attenuated the loss of protein, it failed to improve lung edema and pulmonary gas exchange, thereby limiting its clinical use. PMID- 15665729 TI - Suppression of hepatic cytochrome p450-mediated drug metabolism during the late stage of sepsis in rats. AB - The effects of polymicrobial sepsis on the activity and gene expression of hepatic microsomal cytochrome P450 (CYP) were examined. Rats were subjected to polymicrobial sepsis by cecal ligation and puncture (CLP). Liver and blood samples were taken 2, 6, and 24 h after CLP. The serum aminotransferase levels and lipid peroxidation increased 24 h after CLP. The hepatic concentrations of reduced glutathione and total CYP content decreased 24 h after CLP. The CYP1A1 activity and its protein level decreased 24 h after CLP. The CYP1A2 activity decreased 2 h and 24 h after CLP. Although the CYP2B1 mRNA expression level decreased 6 h and 24 h after CLP, the CYP2B1 activity and its protein level did not change in any of the experimental groups. The CYP2E1 activity and its protein level decreased 24 h after CLP. The CYP2E1 mRNA levels were lower at both 6 h and 24 h after CLP. The TNF-alpha mRNA expression level increased 2, 6, and 24 h after CLP. The iNOS mRNA expression level increased 24 h after CLP. These findings suggest that sepsis causes abnormalities in the microsomal drug metabolizing function, particularly in the late stage, which is associated with higher level of oxidant stress and lipid peroxidation. PMID- 15665730 TI - Effects of inhaled nitric oxide on lung injury after intestinal ischemia reperfusion in rats. AB - Splanchnic ischemia/reperfusion (I/R) induces a systemic inflammatory response with acute lung injury. Impaired production of endothelial nitric oxide (NO) plays a key role in this process. We evaluated the effects of early treatment with inhaled NO (iNO) on lung microcirculatory inflammatory changes during splanchnic I/R. I/R was induced in rats by occlusion of the superior mesenteric artery (SMA; 40 min) and reperfusion (90 min). Four groups were studied: Control, anesthesia only; Sham, all surgical procedures without I/R, ventilated with air; Air, SMA I/R, ventilation with air; and NO, SMA I/R, ventilation with NO (20 ppm) starting 10 min before reperfusion. Intravital video microscopy was used to monitor pulmonary macromolecular flux and capillary flow velocity (CFV). Leukocyte infiltration was determined by morphometry. SMA I/R decreased mean arterial blood pressure, capillary CFV (P < 0.01), and shear rate (P < 0.01), and increased pulmonary macromolecular leak by 138% +/- 8% (P < 0.001). iNO markedly attenuated the increase in macromolecular leak (P < 0.01), blunted the decrease in capillary CFV (P < 0.05) and shear rate (P < 0.05), and prevented the increase in leukocyte infiltration of the lungs after SMA I/R (P < 0.05). The direct, real time, in vivo data suggest that early institution of low-dose iNO therapy effectively ameliorates the acute remote pulmonary inflammatory response after splanchnic I/R. PMID- 15665731 TI - Sivelestat, a neutrophil elastase inhibitor, attenuates neutrophil priming after hepatoenteric ischemia in rabbits. AB - Neutrophils play an important role in ischemia-reperfusion injury. The neutrophil elastase not only causes tissue damage, but also mediates neutrophil priming. In the present study, we use a rabbit model of hepatoenteric ischemia-reperfusion to test the hypothesis that neutrophil elastase inhibition ameliorates an ischemia reperfusion injury by attenuating neutrophil priming and suppressing enzymatic activity. Twenty-four Japanese white rabbits underwent 30 min of supraceliac aortic cross-clamping and 180 min of reperfusion under isoflurane anesthesia. The rabbits randomly received the neutrophil elastase inhibitor, sivelestat (n = 10), or saline (n = 14). Neutrophil priming was then assayed with luminol-dependent neutrophil chemiluminescence. Hepatic, intestinal, renal, and pulmonary damages were assessed with serum transaminase, lactate dehydrogenase concentrations, urinary N-acetyl glucosaminidase activity, and protein concentration in post mortem bronchoalveolar lavage fluid. We discovered that neutrophil elastase inhibition suppressed plasma neutrophil elastase, and that lipid peroxide concentrations increased after reperfusion. It improved ischemia-reperfusion injuries in the liver, intestine, kidney, and lung. Furthermore, inhibition of neutrophil elastase with sivelestat significantly attenuated post-reperfusion neutrophil priming. The results of this study demonstrate that neutrophil elastase inhibition could effectively attenuate an ischemia-reperfusion injury caused by supraceliac aortic cross-clamping, most likely from the attenuation of neutrophil priming. PMID- 15665732 TI - Heat shock pretreatment may protect against heatstroke-induced circulatory shock and cerebral ischemia by reducing oxidative stress and energy depletion. AB - The mechanisms underlying the protective effects of heat shock pretreatment on heatstroke remain unclear. Here we attempted to ascertain whether the possible occurrence of oxidative stress and energy depletion exhibited during heatstroke can be reduced by heat shock preconditioning. In the present study, colonic temperature, mean arterial pressure, heart rate, striatal levels of heat shock protein 72 (HSP72), local Po2, brain temperature, cerebral blood flow, cellular ischemia and damage markers, dihydroxybenzoic acid (DHBA), lipid peroxidation, glutathione, glutathione peroxidase and reductase activities, and ATP were assayed in normothermic control rats and in heatstroke rats with or without preconditioning 16 or 96 h before initiation of heatstroke. Heatstroke was induced by exposing the anesthetized rats to a high ambient temperature (Ta = 43 degrees C) until the moment at which MAP decreased from its peak level. Sublethal heat shock pretreatment 16 h before initiation of heatstroke, in addition to increasing striatal HSP72 levels, conferred significant protection against heatstroke-induced arterial hypotension, striatal ischemia and damage, increment of hydroxyl radical formation, lipid peroxidation, glutathione oxidation, and decrement of glutathione peroxidase activity and ATP. However, at 96 h after heat shock, when striatal HSP72 expression returned to basal levels, the above responses that occurred during onset of heatstroke were indistinguishable between the two groups. These results suggest that heat shock pretreatment induces HSP72 overexpression in striatum and confers protection against heatstroke-induced striatal ischemia and damage by reducing oxidative stress and energy depletion. PMID- 15665733 TI - Macrophage arginase regulation by CCAAT/enhancer-binding protein beta. AB - Arginase activity is expressed by macrophages in healing wounds and other sites of inflammation and has been shown to modulate the synthesis of nitric oxide, polyamines, and collagen. The role of CCAAT/enhancer-binding protein beta (C/EBPbeta) in the regulation of macrophage arginase by different agonists was investigated using C/EBPbeta-/- and +/+ macrophage cell lines. 8-Bromo-cyclic adenosine monophosphate (8-Br-cAMP, 0.5 mM), recombinant murine interleukin 4 (rmIL-4, 20 U/mL), Escherichia coli lipopolysaccharide (100 ng/mL), and hypoxia (1% O2) induced arginase activity in C/EBPbeta+/+ macrophages, where enzyme activity correlated with arginase I protein. Only rmIL-4 increased arginase activity in C/EBPbeta-/- cells. Arginase II protein was expressed constitutively in wild-type and C/EBPbeta-/- cell lines and was unaltered by 8-Br-cAMP or rmIL 4. rmIL-4-stimulated immortalized C/EBPbeta-/- macrophages demonstrated higher nuclear signal transducer and activator of transcription-6 (STAT6) and phospho STAT6 content than their +/+ counterparts. Validating the biological relevance of findings with the cell lines, additional experiments examined wound fluids and peritoneal macrophages from C/EBPbeta-/- mice and demonstrated that both contained less arginase activity than those from wild-type controls. Wounds in C/EBPbeta-/- animals showed signs of delayed maturation, as manifested by the persistence of neutrophils in the inflammatory infiltrate. Peritoneal macrophages from C/EBPbeta+/+ animals responded to 8-Br-cAMP and rmIL-4 with increased arginase activity, whereas those from C/EBPbeta-/- mice did not respond to cAMP. Results demonstrate a key mechanistic role for C/EBPbeta in the modulation of macrophage arginase I expression in vivo and in vitro. PMID- 15665734 TI - Mechanisms of increased survival after lipopolysaccharide-induced endotoxic shock in mice consuming olive oil-enriched diet. AB - We examined the impact of dietary fatty acid intake on lipopolysaccharide (LPS) induced endotoxic shock. C57Bl/6J mice were fed for 6 weeks with a commercial laboratory chow (CC) or with test chows containing 7% (w/w) canola oil (CO), sesame oil (SeO), soybean oil (SO), or virgin olive oil (OO). The increase in body weight and energy consumption were similar for all diets tested. In the sixth week, mice were injected intraperitoneally with 400 microg of bacterial LPS to induce endotoxic shock. LPS induced a massive neutrophil infiltration into the peritoneal cavity and an increase in lipid body (LB) formation in leukocytes recovered from the peritoneal fluid of mice fed with CC, CO, SeO, or SO. In addition, there were increases in prostaglandin E(2) (PGE(2)), leukotriene B4 (LTB(4)), and cytokines IL-6, IL-10, and MCP-1 in peritoneal lavage, as well as in plasma TNF-alpha. In contrast, mice fed with OO exhibited reduced neutrophil accumulation and LB formation, and also had lower levels of PGE(2), LTB(4), MCP 1, and TNF-alpha. All mice fed with CC, CO, SeO, or SO died within 48 to 72 h after LPS injection. Interestingly, mice fed with the OO diet were resistant to endotoxic shock, with 60% survival at 168 h. These data indicate that intake of OO may have a beneficial role, reducing the magnitude of the inflammatory process triggered by endotoxic shock through modulation of LB formation and of the production of inflammatory mediators. PMID- 15665735 TI - Nitric oxide-cyclic GMP contributes to abnormal activation of Na+-K+-ATPase in the aorta from rats with endotoxic shock. AB - We examined pharmacologically the influence of nitric oxide (NO), guanosine 3':5' cyclic monophosphate (cyclic GMP), adenine 3':5'-cyclic monophosphate (cyclic AMP), and protein kinase C-linked signaling pathways on relaxation to potassium in aortic segments isolated from rats treated for 6 h with bacterial endotoxin (lipopolysaccharide). Endotoxemia for 6 h was associated with a severe hypotension and vascular hyporeactivity to norepinephrine (NE), and an increase in plasma NO in vivo and aortic NO ex vivo. The NE-induced contraction was attenuated and the potassium-induced relaxation was accentuated in the aorta of rats with endotoxic shock. Ouabain inhibited the potassium-induced relaxation in aortae from normal and endotoxemic rats. 8-Bromo-cyclic GMP significantly enhanced the potassium-induced relaxation in control aortae, whereas 1H [1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) abolished this difference between normal and endotoxemic rats. In contrast, inhibition of potassium-induced relaxation was observed in aortae from normal and endotoxemic rats treated with 8 bromo-cyclic AMP or phorbol 12-myristate 13-acetate. Individually, inhibitors of protein kinase A or protein kinase C did not significantly alter relaxation to potassium; however, in combination, these inhibitors significantly potentiated relaxation in aortae from control rats. These results suggest that activity of Na(+)-K(+)-ATPase is enhanced in the vascular bed of animals with endotoxic shock and that this elevation in activity is mediated by NO-cyclic GMP, but not by cyclic AMP-protein kinase A or protein kinase C. PMID- 15665736 TI - Superoxide potentiates NF-kappaB activation and modulates endotoxin-induced cytokine production in alveolar macrophages. AB - Gram-negative bacterial infection predisposes to the development of shock and acute lung injury with multiple organ dysfunction in the critically ill. Although overexpression of proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) and interleukin (IL)-1beta, IL-6, IL-8, and other mediators is causally implicated in the pathogenesis of shock and lung injury, the underlying mechanisms following cellular exposure to gram-negative endotoxin remain unclear. De novo generation of reactive oxygen species (ROS) by monocytes/macrophages in particular has been proposed as a pivotal regulatory mechanism by which enhanced transactivation of redox-sensitive genes culminates in augmented cytokine expression within the lower respiratory tract. Here we sought to characterize the mechanism of action of a synthetic, nonpeptide, low-molecular-weight, Mn containing superoxide dismutase mimetic (SODm), M40403, in modulating E. coli lipopolysaccharide serotype 0111:B4 (LPS)-induced cytokine production by cultured rat alveolar macrophages. Intracellular superoxide (O2) ion generation was measured using hydroethidine (HE) dye, and the dose-dependent effects of M40403 on TNF-alpha and IL-6 biosynthesis by ELISAs. Upstream redox-sensitive signaling events involving the pleiotropic transcription factor NF-kappaB were determined in nuclear extracts by electrophoretic mobility shift assays (EMSAs) and p65 subunit Western blot. The levels of the cytosolic inhibitory protein IkappaB alpha were also assessed by Western analysis. We found that M40403 potently suppressed the production of superoxide, TNF-alpha, and IL-6 in LPS-stimulated alveolar macrophages, suggesting a key role for superoxide in endotoxin-induced cytokine production in the distal air spaces. In addition, M40403 decreased E. coli LPS-induced activation of NF-kappaB, and this effect was associated with modest suppression of cytoplasmic IkappaB-alpha degradation. Together, these results suggest that removal of superoxide by M40403 inhibits endotoxin-induced production of TNF-alpha and IL-6 in alveolar macrophages by a mechanism involving suppression of redox-sensitive NF-kappaB transactivation or signaling. PMID- 15665738 TI - Seeking optimal prescription of cyclosporine ME. AB - Optimal use of cyclosporine microemulsion (CsA ME) in transplant recipients is still a matter of debate. Therapeutic drug monitoring of CsA ME is needed because high variability of inter- and intraindividual exposure to this drug has been reported. Thus, consensus guidelines have been recommended, but the ideal method of follow-up has not yet been found. Pharmacokinetic studies have shown that the 2-hour postdose (C2) sample concentration is more closely correlated with the risk of acute rejection and with toxicity in solid organ recipients compared with the trough (C0) concentration within the first 3 months posttransplantation. For some years, this time point has therefore been used to monitor patients treated with CsA ME. However, there are still some technical (accurate dilution) and practical concerns (education of the patient). Moreover, the target levels of C2, in particular after the first year posttransplantation, are less well defined. Pharmacogenetic studies of the MDR-1 gene as well as the CYP3A4 and CYP3A5 gene polymorphisms could not yet demonstrate any clear influence on the interindividual variations on the pharmacokinetic profile of CsA ME. PMID- 15665737 TI - When a cocaine-using mother wishes to breastfeed: proposed guidelines. PMID- 15665739 TI - Improved renal function in de novo renal transplant patients on sirolimus maintenance therapy following discontinuation of cyclosporine. AB - In the presirolimus era, cyclosporine withdrawal in de novo renal transplant patients was associated with increased rates of rejections; thus, no improvement in long-term graft survival was achieved. However, those patients who did not reject exhibited longer graft survival than did those on cyclosporine. In recent years sirolimus has been introduced into clinical practice and, so far, 4 randomized cyclosporine withdrawal trials in de novo patients after renal transplantation have been published, together with 3 smaller studies directly comparing cyclosporine with sirolimus. The main finding in all these studies was better renal function after cyclosporine withdrawal. In the largest trial with the longest follow-up there was even a trend toward higher rates of graft survival at 3 years and fewer histopathological findings in protocol biopsies at that time. Whether this success in renal function and graft survival will project to prolonged patient survival remains to be determined. In summary, with the introduction of sirolimus as a potent immunosuppressive but nonnephrotoxic drug, a considerable improvement in graft and patient survival might be possible. PMID- 15665740 TI - Cost-effectiveness of therapeutic drug monitoring: a systematic review. AB - There are a number of effective but highly toxic drugs that exhibit a narrow therapeutic index and marked interpatient pharmacokinetic variability. Individualized therapy with such drugs requires therapeutic drug monitoring (TDM) to obtain the desired clinical effects safely. Cost-effectiveness analysis in health care is still at an early stage of development, especially for TDM. A systematic review was carried out to document studies that have addressed the cost-effectiveness of TDM. The Cochrane database and Medline were searched. References identified by this approach were then searched manually for relevant articles. Very few studies have been performed that document the cost effectiveness of TDM, and TDM has been demonstrated to be cost-effective only for aminoglycosides. For the other classes of drugs that are monitored, the rationale for TDM has been supported, but appropriate cost-effectiveness analyses have not been performed. Because the use of many of these drugs without TDM would increase the risk of under- or overdosing, emphasis should not be placed solely on cost effectiveness but rather on how such interventions can be applied in the most cost-effective and clinically useful manner. PMID- 15665741 TI - Evaluation of Bayesian estimation of pharmacokinetic parameters. AB - The validity of pharmacokinetic parameters estimated by the maximum a posteriori probability (MAP) Bayesian method was investigated by simulation studies. A 1 compartment model with bolus intravenous administration was used as a pharmacokinetic model, and the coefficients of variation for the parameters and residual error were set at 30% and 10%, respectively. The accuracy of the posterior modes of pharmacokinetic parameters estimated by the MAP Bayesian method was assessed by the difference between the true value and the estimated value. The results showed that the accuracy of the Bayesian estimation depended on sampling times and on the differences between the prior means and individual true parameter values. For assessing the reliability and accuracy of the Bayesian estimation, the authors suggest using the whole posterior distribution of the pharmacokinetic parameters to describe the 95th percentile range for predicted blood concentration profiles. The authors believe that the proposed procedures provide helpful information for evaluating the Bayesian estimation of pharmacokinetic profiles. PMID- 15665742 TI - A simple and sensitive fully validated HPLC-UV method for the determination of 5 fluorouracil and its metabolite 5,6-dihydrofluorouracil in plasma. AB - The authors developed a simple and sensitive, fully validated HPLC-UV method for the determination of both 5-FU and its metabolite DHFU in small-volume plasma samples. The analytes were separated on a 4.6 x 250 mm ID Atlantis dC18 5-microm column with isocratic elution at room temperature. Chlorouracil was used as internal standard. The analytes were detected with an UV diode array detector. DHFU was detected at 205 nm, 5-FU at 266 nm, and chlorouracil at both wavelengths. The limits of quantification in plasma were 0.040 mug /mL for 5-FU and 0.075 microg/mL for DHFU. Linearity, accuracy, precision, recovery, dilution, freeze-thaw stability, and stability in the sample compartment were evaluated. The method appeared linear over a range from 0.04 to 15.90 microg/mL for 5-FU and from 0.075 to 3.84 microg/mL for DHFU. The method appeared very suitable for therapeutic drug monitoring and pharmacokinetic studies of 5-FU because of its simple extraction and small sample volume. Problems in earlier published methods with interfering peaks and variable retention times were overcome. The method appeared also to be suitable for detection of uracil and its metabolite dihydrouracil in plasma. PMID- 15665743 TI - Analytic performance evaluation of a new turbidimetric immunoassay for carbamazepine on the ADVIA 1650 analyzer: effect of carbamazepine 10,11-epoxide. AB - Carbamazepine, an anticonvulsant, requires therapeutic drug monitoring. Recently Bayer HealthCare, Diagnostics Division released a turbidimetric immunoassay of carbamazepine on the ADVIA 1650 analyzer. We evaluated the analytic performance of this assay by comparing values obtained with this new assay in sera of 54 patients receiving carbamazepine with the values obtained by using a widely used fluorescence polarization immunoassay (FPIA) and a chemiluminescent immunoassay (CLIA). The new turbidimetric immunoassay for carbamazepine showed excellent precision. The low control showed a total CV of 4.9% (mean 2.86, SD 0.14 microg/mL), the medium control demonstrated a total CV of 3.5% (mean 7.79, SD 0.27 microg/mL), and the high control showed a total CV of 4.8% (mean 16.15, SD 0.78 microg/mL). The assay was linear up to a carbamazepine concentration of 20 microg/mL. The assay showed excellent dilution recovery and recovery of samples supplemented with carbamazepine (mean recovery 102.2%). We observed an excellent correlation between the values obtained by the FPIA (x-axis) assay and the new turbidimetric (y-axis) assay (y = 0.96 x - 0.46, r = 0.99, n = 54). We also observed excellent correlation between the values obtained by the CLIA (x-axis) and the turbidimetric (y-axis) assay (y = 1.10 x -0.32, r = 0.99, n = 54). However, the slope of 1.10 was higher than the slope of 0.96 observed with the regression equation obtained by using values obtained by the FPIA and the turbidimetric assay. The positive bias obtained with the new turbidimetric assay compared with the CLIA assay resulted from lower cross reactivity of carbamazepine 10,11-epoxide, the active metabolite of carbamazepine, with CLIA. On the other hand, the cross reactivity of the metabolite is similar between the new turbidimetric assay and the FPIA assay. We conclude that the new turbidimetric assay can be used for routine monitoring of carbamazepine in clinical laboratories. PMID- 15665744 TI - The portal immunosuppressive storm: relevance to islet transplantation? AB - Outcomes in clinical islet transplantation improved substantially with the introduction of combined sirolimus and tacrolimus immunosuppression. However, multiple islet preparations are often required to achieve insulin independence, suggesting that islet engraftment may not be optimal when these agents are absorbed via the portal vein. The current study was designed to assess the differential concentrations of immunosuppressive drugs within the portal and systemic circulations of a large animal model, to assess the local concentrations of drugs to which islets are exposed early after implantation. Chronic catheters were placed in the portal vein and carotid artery of 6 mongrel dogs, and immunosuppressants were administered orally. Blood samples were drawn simultaneously from portal and systemic catheters, and drug concentrations were analyzed. Peak immunosuppressant levels as well as area under the curve were dramatically elevated in portal blood relative to systemic levels for all drugs tested. This "portal storm" of immunosuppression may be relevant to intrahepatic islet transplantation. PMID- 15665746 TI - Quantitative determination of the calcium channel antagonists amlodipine, lercanidipine, nitrendipine, felodipine, and lacidipine in human plasma using liquid chromatography-tandem mass spectrometry. AB - A sensitive and specific liquid chromatography-tandem mass spectrometric method has been developed and validated for the quantification of the five 1,4 dihydropyridine calcium channel antagonists amlodipine, lercanidipine, nitrendipine, felodipine, and lacidipine in human plasma. Sample preparation involved solid-phase extraction on RP-C18 cartridges with good recovery for all the compounds. Sample analysis was performed on a Luna RP-C18 analytical column (15 mm x 2 mm ID, 3.0 microm) with a Sciex API 365 triple quadrupole mass spectrometer with turboionspray source and multiple reaction monitoring. The method is sensitive with a limit of detection below 1 ng/mL for each drug in plasma, with good linearity (r(2) > 0.998), over the therapeutic concentration range (1 to 40 ng/mL). All the validation data, such as accuracy, precision, and interday repeatability, were within the required limits. The method can be used for pharmacokinetic studies and therapeutic drug monitoring of the compounds in humans. PMID- 15665745 TI - A sensitive HPLC method for the determination of fluoxetine and norfluoxetine in human plasma with fluorescence detection. AB - A selective, sensitive, and precise HPLC method for the simultaneous determination of fluoxetine (FL) and its N-demethylated metabolite norfluoxetine (NFL) in human plasma has been developed. Following extraction with n-hexane, FL, NFL, and fluvoxamine (internal standard) were derivatized with 7-chloro-4 nitrobenzofurazan (NBD-Cl) under weakly alkaline conditions. NBD derivatives were extracted with chloroform after acidification and chromatographed on a reversed phase column with gradient elution using acetonitrile and 0.1 mol/L nitric acid (pH 3) solution. Calibration curves were linear over the range of 1.0-100.0 ng/mL and 0.1-50.0 ng/mL for FL and NFL, respectively, with inter- and intraassay precision given by a relative standard deviation (RSD%) of less than 9.2%. The lower limits of quantification were 1.0 ng/mL for FL and 0.1 ng/mL for NFL. Recoveries of FL and NFL from plasma at three different concentrations were assessed. Average recovery was about 100% for both substances. The assay was applied to pharmacokinetic study in 2 healthy volunteers after a single oral administration of 40 mg of FL. PMID- 15665747 TI - A multidisciplinary approach to the toxicologic problems associated with the use of herbal medicines. AB - In Hong Kong, a multidisciplinary team consisting of a pharmacist, a chemical pathologist, a scientific officer, and a physician has jointly provided an advisory service on herbal safety to healthcare professionals of all public hospitals since August 2000. This paper reports the first 2 years of experience of this team to emphasize the importance of a multidisciplinary approach to herbal poisoning. Twenty referrals received from the public hospitals from August 2000 to June 2002 were reviewed. The diagnosis of herbal toxicity was made after consideration of the temporal relationship between exposure to the herb and the occurrence of the adverse event and the exclusion of other causes. The causal relationship was seen as much stronger if the herb's pharmacological effects could fully explain the adverse event, if there were similar previous reports, and if the toxicological analysis findings were supportive. In 10 patients, the adverse events were deemed unlikely to be related to the use of herbal medicines. In 3 patients, it was concluded that the adverse events were possibly related to the use of herbal medicines. In 7 patients, the causal relationship was seen to be much stronger because the adverse events in these patients simply reflected the pharmacological effects of the herbs consumed and specific toxic ingredients were isolated from herbal residues and patients' urine or serum. The clinical problems arising from the use of herbal medicines included the Aristolochia species causing acute renal failure (n = 1), aconite roots causing aconitine poisoning (n = 2), the Datura species causing anticholinergic poisoning (n = 1), and "yulan" (Stephania sinica) causing tetrahydropalmatine poisoning (n = 3). The 4 patients with acute renal failure and tetrahydropalmatine poisoning received these toxic herbs, which were not listed in the prescriptions, as a result of poor dispensing practice or for other reasons. Toxicological problems associated with the use of herbal medicines are complex and may be easily overlooked. A multidisciplinary team of experts should be made available to provide advice to frontline healthcare professionals. PMID- 15665748 TI - Factors influencing the overestimation of plasma vancomycin concentrations measured by the Abbott TDx technique. AB - The Abbott TDx technique (TDx) has been reported to overestimate the plasma concentration of vancomycin (VCM) in patients with renal failure and also in those with normal renal function. The aim of this study was to investigate factors influencing the overestimation of plasma VCM concentrations measured by TDx compared with high-performance liquid chromatography (HPLC) as a reference technique. First, the precision and accuracy of TDx and HPLC were compared using 5 weighed-in concentrations of VCM. The coefficients of variation (CV) for both TDx and HPLC were less than 3% at weighed-in concentrations of 3.0, 7.5, 15.0, 30.0, and 60.0 microg/mL. The authors did not find overestimation of VCM concentrations by TDx in any weighed-in sample. Next, VCM concentrations measured by TDx were compared with those measured by HPLC in 253 plasma samples obtained from 83 patients receiving VCM. Regression analysis showed that VCM concentrations measured by TDx were closely correlated with those measured by HPLC, with a correlation coefficient of 0.963. Then, the authors divided plasma samples into 2 groups based on the time of sampling, ie, pre- and postdose. The mean TDx/HPLC ratio was significantly higher in the predose group (1.23 +/- 0.22) than in the postdose group (1.11 +/- 0.16) (P < 0.01). An Eksborg plot showed that the TDx/HPLC ratio decreased slightly with increased total bilirubin concentrations in the predose samples. These results indicated that plasma VCM concentrations measured by TDx were influenced by the time of sampling and the plasma bilirubin concentration. The overestimation of VCM concentrations in the predose group may have resulted from metabolites of VCM because the relative concentrations of the metabolites were likely to be greater in the predose group than in the postdose group. Thus, VCM postdose samples may provide useful information in cases in which a good clinical outcome has not been obtained using monitoring of the predose concentration alone. PMID- 15665749 TI - Indinavir plasma concentration and adherence score are codeterminant of early virologic response in HIV-infected patients of the APROCO cohort. AB - To study the respective roles of indinavir concentrations and treatment adherence as predictors of early virologic response, we analyzed the patients of the APROCO cohort treated by indinavir 800 mg TID during the first 4 months. Minimum (Cmin), maximum (Cmax), and the ratio of the measured to expected concentrations (CR) were estimated for each patient at M4, from a population pharmacokinetic analysis of all data. The relationship among virologic success at M4 [plasma HIV RNA (VL) <500 copies/mL], baseline characteristics, estimated indinavir concentrations, and adherence score measured by a self-administered questionnaire, was analyzed by multivariate logistic regression. In the 216 studied patients, baseline median HIV RNA was 4.4 log10 copies/mL, and CD4 cell count was 309/mm. Virologic success was achieved in 195 (90%) patients; it was independently related to baseline viral load (OR = 0.524, CI 0.29-0.93; P = 0.03), antiretroviral treatment naive status (OR = 3.89, CI 1.29-11.76; P = 0.01), and indinavir Cmin (OR = 1.06, CI 1.02-1.10; P = 0.004) when adherence score was not included in the model, whereas full adherence was the only independent related factor when included in the model (OR = 8.8, 95% CI 2.85-27.3; P < 10). In the 168 fully adherent patients, virologic success was more frequent in patients with shorter duration of antiretrovirals at baseline (P = 0.03), lower baseline HIV RNA (P = 0.03), and higher indinavir CR (P < 10); the most discriminating Cmin cut-off was 194 ng/mL. Data on the relationship between indinavir plasma concentration and virologic success are therefore misleading without a concomitant assessment of adherence. These data suggest that any strategy of therapeutic drug monitoring must imply first a combined evaluation of plasma concentrations and adherence level and second an intervention target based on the results of both assessments. PMID- 15665750 TI - The pH dependency of the binding of drugs to plasma proteins in man. AB - An analysis of pH-induced changes of drug binding may contribute to the understanding of the mechanisms involved and the clinical relevance. A literature search was performed, and acceptance criteria set up, to select reported data for quantitative evaluation. The relationship between percentage of unbound drug, fu, and pH was analyzed, and the relevance of physicochemical characteristics of the ligand drugs and the importance of hydrogen ion-induced changes in plasma proteins for the pH sensitivity of the binding were evaluated. With all basic and the majority of acidic drugs, fu depended linearly on pH. Basic drugs showed a consistent behavior with fu decreasing with increasing pH. Acidic compounds behaved differently: With some, fu increased, and with others fu decreased, with pH, and with a third group of acids fu was pH independent. Large differences in the pH sensitivity of the plasma protein binding among individual compounds were found. The fu in plasma for some bases and acids increased up to 136% and 95%, respectively, at pH values seen in severe acidemia or alkemia. These changes in fu could be clinically relevant with narrow-therapeutic-range drugs. Physicochemical properties and other characteristics of the ligands affect the pH sensitivity of the interaction with plasma proteins, but there was clear evidence indicating that pH-induced changes in the plasma proteins are also involved in the observed pH-dependent interaction with ligands. It is generally accepted that the unbound, free fraction in whole blood or plasma is an important determinant of the pharmacokinetics and pharmacodynamics of drugs. pH-dependent protein binding and consequent changes in the free fraction have been reported for many drugs. From a basic science point of view, the systematic study of pH-induced perturbations of the drug-protein interaction may provide insight into the mechanism and forces involved in the binding of drugs to plasma proteins. From a clinical viewpoint it may be of interest to know the extent of pH-induced changes in the unbound fraction of drugs under extreme acidemic or alkalemic conditions. Arterial blood pH values compatible with life reportedly range between 6.7 and 8.0. pH values as low as 6.3 have been measured in survivors of drowning accidents. To the best knowledge of the authors, a review and interpretation of pH-associated changes in the protein binding of drugs has not been attempted to date. The goals of this investigation were to (1) review published results of studies that determined the impact of pH changes on the protein binding of drugs in man, (2) select representative data using predetermined criteria, (3) determine relevant factors impacting the pH sensitivity of the drug-protein interaction, and (4) attempt to interpret the results and their clinical relevance. PMID- 15665751 TI - Adulteration of South African traditional herbal remedies. AB - Traditional herbal remedies are used as alternative medicine by a large proportion of people worldwide. Herbals are generally perceived as safe, harmless, and free from side effects, but there have been reports of adulteration of Asian herbal remedies with Western pharmaceuticals, a practice that has not hitherto been observed in Africa. The authors present 2 cases from South Africa in which herbal remedies, consumed by patients admitted to hospital, were found to be adulterated with commercially available Western medicines. The adulterants were identified using high-performance liquid chromatography with a photodiode array detector and gas chromatography-mass spectrometry. The authors alert clinicians to the fact that adulteration of herbal remedies may become more common and should be considered, especially in cases where the symptoms are confusing in light of the history. PMID- 15665752 TI - Evaluation of the immunotech cyclosporine direct radioimmunoassay for the determination of whole-blood cyclosporine in renal transplant patients. AB - Therapeutic drug monitoring (TDM) of cyclosporine (CsA) has been an accepted as an essential tool in the management of solid organ transplant recipients. The authors evaluated a new CsA method, Immunotech cyclosporine direct radioimmunoassay (Beckman Coulter, Prague, Czech Republic), for the measurement of whole-blood CsA concentrations. The performance was compared with CEDIA Plus method as well as group mean data for HPLC and other immunoassays available from the International CsA Proficiency Testing Program (www.bioanalytics.co.uk). Regression analysis of patient samples gave a relationship of RIA = 1.0822 CEDIA(+) + 69.84 (r(2) = 0.933). External CsA-spiked proficiency-testing (PT) samples gave a regression equation of RIA = 0.9672 CEDIA(+) + 4.99 (r(2) = 0.996). The correlation with the CEDIA Plus method using patient specimens (hence, including CsA metabolites) suggested that the test RIA method possibly had slightly inferior specificity for parent CsA. The results suggest that the Immunotech cyclosporine direct RIA kit is suitable for the measurement of whole blood CsA concentrations and maintained clinically acceptable analytic precision and accuracy, displaying CVs of less than 15% and biases of less than 10%. The PT program CsA-metabolite-free samples showed that calibration between methods was comparable with the possible exception of mFPIA/TDx. PMID- 15665753 TI - Effects of hematocrit value on microparticle enzyme immunoassay of tacrolimus concentration in therapeutic drug monitoring. AB - The effects of hematocrit (Ht) value on microparticle enzyme immunoassay (MEIA) of tacrolimus concentration were examined in 1063 whole-blood samples from 42 transplant recipients (13 liver, 20 kidney, and 9 bone marrow transplantations). MEIA guarantees the test's assay quality for blood tacrolimus in samples with Ht values of 25% to 45%. However, 129 samples (29.3%) obtained from liver transplant recipients and 107 samples (61.5%) from bone marrow transplant recipients had lower Ht (<25%). Further, 81 blood samples (18.1%) with Ht > 45% were observed in kidney transplant patients. Twenty-five whole-blood samples with low Ht were tested by 3 assay methods for tacrolimus: MEIA, modified, corrected MEIA (cMEIA), and enzyme-linked immunosorbent assay (ELISA). MEIA gave higher blood concentrations of tacrolimus than ELISA (16.1 versus 11.0 ng/mL, P < 0.001). This difference was generated by overestimation in MEIA and was not observed in samples with normal Ht. This overestimation was eliminated by using cMEIA on samples with low Ht values: there was no difference in blood tacrolimus concentration between cMEIA and ELISA (12.3 versus 11.0 ng/mL). ELISA or cMEIA should be used for tacrolimus assay in samples obtained from bone marrow transplant recipients with anemia and from liver and kidney transplant recipients with unstable Ht values. PMID- 15665754 TI - Pharmacodynamics of midazolam in pediatric intensive care patients. AB - The aim of the study was to study a possible pharmacokinetic-pharmacodynamic (PK PD) relationship for midazolam in pediatric intensive care patients and to determine how adequate sedation could be reached using the COMFORT scale as sedation scale. Twenty-one pediatric intensive care patients (2 days to 17 years) received a midazolam infusion (0.05-0.4 mg/kg/h, 3.8 hours to 25 days). Sedation levels were determined using the COMFORT scale as well as plasma concentrations of midazolam and metabolites. An evident PK-PD relationship was not found. In 20 of the 21 patients midazolam dosing could be effectively titrated to the desired level of sedation, assessed by the COMFORT scale. Based on our findings that there is no relationship between pharmacokinetic parameters and pharmacodynamic outcome, we recommend that midazolam dosing should be titrated according to the desired clinical effect in combination with a validated assessment instrument, eg, the COMFORT scale. PMID- 15665755 TI - Serum concentrations of risperidone and 9-hydroxyrisperidone after administration of the long-acting injectable form of risperidone: evidence from a routine therapeutic drug monitoring service. AB - There is limited documentation on the pharmacokinetics of long-acting intramuscular risperidone in a naturalistic setting. The objective of this study was to investigate the concentrations of risperidone and its active metabolite 9 hydroxyrisperidone as well as the concentration/dose (C/D) ratios achieved after intramuscular depot administration in a routine therapeutic drug monitoring setting. Thirty samples were collected from 10 female and 20 male patients receiving depot injections of risperidone. For 6 of the patients, the results could be compared with corresponding data available after previous oral administration of risperidone. In addition, data from a group of 278 patients using oral risperidone were retrieved. The median serum concentrations of risperidone plus 9-hydroxyrisperidone were 38 nmol/L 12-14 days after a intramuscular dose of 25 mg/14 days, 67 nmol/L after a dose of 37.5 mg/14 days, 99 nmol/L after a dose of 50 mg/14 days, and 148 nmol/L after a dose of 75 mg/14 days. The median C/D ratio for risperidone plus 9-hydroxy-risperidone was 22.2 (nmol/L)/(mg/d). In the group on oral medication, the median C/D ratio was 18.6 (nmol/L)/(mg/d). In the 6 patients previously using oral risperidone, switching to depot injections gave an average increase of 33% (range 12%-68%) in the C/D ratio. In conclusion, the authors' data indicate that there are great interindividual differences in the extent to which the daily dose has to be reduced when switching from oral to intramuscular depot administration to achieve the same serum concentrations of risperidone plus 9-hydroxyrisperidone. Because the degree of dose adjustment required for the individual patient so far cannot be predicted, guidance by therapeutic drug monitoring might be helpful. PMID- 15665756 TI - Pharmacokinetic interaction between tacrolimus and clarithromycin in a heart transplant patient. PMID- 15665757 TI - A population pharmacokinetic-metabolism model for individualizing ciprofloxacin therapy in ophthalmology. PMID- 15665758 TI - Age and immune response in organ transplantation. AB - The immune system undergoes a complex and continuous remodeling as the result of aging. These changes have a major impact on allorecognition and alloresponse. In addition, immunosuppression in the elderly is challenging as a consequence of an increased incidence of associated comorbidities and altered pharmacokinetics. Both advanced donor and recipient age should be considered independent risk factors for poor patient and graft survival rates, albeit acting in a synergistic manner. Consequently, modifications of the immune system because of aging may request an age-adapted allocation and immunosuppression in parallel with close patient monitoring. Interventions to selectively target changes associated with the senescence process seem to be promising therapeutic strategies to improve transplantation outcome. Here, we are going to review the immunologic changes associated with the aging process relevant for transplantation and their impact on immunosuppressive protocols, organ allocation policies, and transplantation outcome. PMID- 15665759 TI - Tolerance or accommodation: the lesson from leflunomide. PMID- 15665761 TI - Fetal CD34+ cells in the maternal circulation and long-term microchimerism in rhesus monkeys (Macaca mulatta). AB - BACKGROUND: Studies in humans have shown that during pregnancy fetal cells can enter the maternal circulation and persist for many years. While we have previously reported the presence of cell-free fetal DNA during pregnancy in rhesus monkeys, it is unknown whether cells circulate and persist long term in maternal tissues. In this study, we asked whether fetal CD34 cells can be found in the maternal circulation and if male fetal cells persist in maternal tissues postdelivery. METHODS: The presence of the Y chromosome in maternal blood and tissues was assessed using real-time PCR assays for the sex determining region Y (SRY) and testes specific protein Y (TSPY) genes. Analysis was done on CD34 and CD34 cells isolated from maternal blood collected at select time points during gestation from gravid animals with male or female fetuses, and tissues were analyzed from nongravid animals that had previously delivered male offspring. RESULTS: All animals with male fetuses tested positive for the Y chromosome in CD34 cells (0-30 cells/50,000 genome equivalents). Y sequences were also found in one or more maternal tissues collected up to 3-years postdelivery (thyroid, heart, spleen, liver, pituitary, adrenals, skin, inguinal lymph nodes). CONCLUSION: These studies suggest transfer of fetal CD34 cells during pregnancy and persistent fetal microchimerism in the rhesus model. Thus, rhesus monkeys can be used to further our understanding of fetal:maternal microchimerism and the role of fetal cells in maternal health and disease. PMID- 15665760 TI - Effects of a short course of leflunomide on T-independent B-lymphocyte xenoreactivity and on susceptibility of xenografts to acute or chronic rejection. AB - BACKGROUND: Leflunomide is a novel immunosuppressive agent with promising activity for xenotransplantation. It is not clear yet which mechanisms of action of leflunomide are responsible for that. METHODS: In a hamster-to-C57BL/6 nude mouse heart transplantation model, a 2-week course of leflunomide was used after transplantation or for pretreating donors. Nontolerant B lymphocytes were transferred to recipients after transplantation of first or second xenogeneic heart grafts that were transplanted with or without leflunomide treatment. RESULTS: Hamster xenogeneic hearts transplanted into athymic C57BL/6 nude mice receiving leflunomide did not induce immunoglobulin (Ig) M xenoantibodies (XAb) and survived without signs of chronic rejection. Second xenogeneic hearts transplanted 4 weeks after withdrawal of leflunomide survived without induction of XAb but developed chronic vascular lesions. After injection of naive B lymphocytes at 6 weeks after grafting a first or second hamster heart, only in the latter case were XAb induced. These were deposited in, and provoked acute rejection of, only the second grafts. Pretreatment of donors with leflunomide decreased the ex vivo xenoantibody deposition on the xenogeneic heart endothelia. CONCLUSIONS: A short posttransplant course of leflunomide induces T-independent B lymphocyte xenotolerance. Leflunomide treatment also influences xenoantigen expression, as nontolerant B lymphocytes provoke IgM XAb formation and rejection of only second xenografts (transplanted without leflunomide) and not of first xenografts (transplanted with leflunomide treatment). The ex vivo experiments that show that XAb deposition is decreased in leflunomide-pretreated xenografts further confirm this. The latter may also explain the resistance of first and not second xenografts against chronic rejection. PMID- 15665762 TI - Tolerizing effects of co-stimulation blockade rest on functional dominance of CD4+CD25+ regulatory T cells. AB - BACKGROUND: Clinical tolerance is the net result of regulatory and effector functions. In this article, the authors show that tolerance induction by co stimulation blockade preferentially works through CD4CD25 regulatory T-cell mediated suppression that is effectively achieved by selective reduction of the effector T-cell load. Anti-CD86 and anti-CD40L monoclonal antibody treatment during in vitro mixed lymphocyte reaction (MLR) typically results in the induction of a suppressive polyclonal T-cell population. This induced suppressive capacity was found to be dependent on the presence of CD4CD25 T cells at the start of MLR. METHODS: Using a CFSE-based strategy, the authors show that within the polyclonal T-cell population, the suppressive effect was exerted by a nondividing CD4CD25 T-cell subset. RESULTS: The cells exclusively originated from preexisting CD4CD25 regulatory T cells and proved anergic and highly suppressive on isolation. They carried the CD45RB and CD62L phenotype and expressed GITR. There was no indication of de novo induction of regulatory T cells by co stimulation blockers. Instead, the authors observed, both in vitro and in vivo, that co-stimulation blockade shifted the ratio between alloreactive effectors and regulatory T cells in favor of the latter. CONCLUSION: The authors therefore conclude that co-stimulation blockade contributes to functional dominance of regulatory T cells by preventing expansion of alloreactive effector T cells. Tolerance-inducing protocols should ideally facilitate this phenomenon. PMID- 15665763 TI - The efficacy of CD40 ligand blockade in discordant pig-to-rat islet xenotransplantation is correlated with an immunosuppressive effect of immunoglobulin. AB - BACKGROUND: The authors' aim was to evaluate the efficacy of immunosuppression with monoclonal anti-CD40 ligand antibodies (aCD40L) or nonspecific polyclonal intravenous immunoglobulin (IVIG) in the pig-to-rat islet xenotransplantation model. METHODS: Fetal porcine islet-like cell clusters were transplanted under the kidney capsule of nondiabetic rats. All antibodies were administered alone or in combination with cyclosporine A (CsA). In addition, some animals were administered antibodies plus tacrolimus (TAC) or sirolimus (SIR). Twelve days after transplantation, islet xenograft survival and rejection were evaluated using immunohistochemistry. RESULTS: aCD40L plus CsA had a pronounced inhibitory effect on islet xenograft rejection for up to 12 days after transplantation. Unexpectedly, treatment with a monoclonal control antibody (anti-keyhole limpet hemocyanin [aKLH]) plus CsA had a similar inhibitory effect. Furthermore, a similar inhibition of islet xenograft rejection was observed also in animals administered IVIG plus CsA. Monotherapy with aCD40L, aKLH, IVIG, or CsA had no effect on the rejection process. Also, when aCD40L or aKLH was administered together with TAC, islet xenograft rejection was inhibited. There was no marked difference compared with rats treated with aCD40L or aKLH and CsA. Immunosuppression with aCD40L or aKLH in combination with SIR also inhibited pig to-rat islet xenograft rejection, but the protective effect was not as pronounced. CONCLUSIONS: Immunosuppression with high doses of antibodies, monoclonal or polyclonal, in combination with CsA or TAC inhibits pig-to-rat islet xenograft rejection. No specific effect of co-stimulatory blockade with aCD40L could be observed. Instead, the results indicate a nonspecific immunosuppressive effect of high doses of antibodies in this model. PMID- 15665764 TI - Reduction of postischemic immune inflammatory response: an effective strategy for attenuating chronic allograft nephropathy. AB - BACKGROUND: Ischemia added to the allogeneic background accelerates the cellular mechanisms involved in alloresponsiveness, supporting the influence of early nonspecific inflammatory injury on chronic allograft nephropathy (CAN). The authors hypothesize that reinforcing initial immunosuppressive regimens may prevent immunogenicity derived from postischemic inflammatory responses, attenuating CAN. METHODS: Lewis rats engrafted with Fischer kidneys received for 15 days overimmunosuppressive doses of rapamycin, a standard cyclosporine regimen, or both, and were followed functionally for 24 weeks. Animals were grouped according to the initial immunosuppressant or cold-ischemia period. Grafts were evaluated for acute inflammatory response at 1 week and for chronic histologic damage at 24 weeks. RESULTS: Rats under cyclosporine alone displayed the highest mortality, which was decreased in the long term by reducing cold ischemia or by strengthening immunosuppression. At 24 weeks, all rapamycin treated groups displayed much less severe tubulointerstitial and vascular damage. The combination of both immunosuppressants offered better functional outcome and a global reduction in chronic histologic damage. After 1 week, ATN and profibrotic features appeared in all 5-hr ischemic animals, indicating that cyclosporine and rapamycin co-treatment did not induce further nephrotoxicity. Treatment with rapamycin, alone or combined with cyclosporine, greatly reduced the severe immune-inflammatory damage, including vessels, shown in cyclosporine treated ischemic grafts. CONCLUSIONS: Strengthening initial immunosuppression attenuates the intensity and extent of the early postischemic immune-inflammatory response as well as later function and structure of renal allografts. Severe CAN may be prevented by reducing cold ischemia or strengthening immunosuppression. Because the former approach is not always possible, reinforcement of early immunosuppression constitutes an excellent alternative. PMID- 15665765 TI - Multiple negative feedbacks on CD152 expression in allograft tolerance. AB - BACKGROUND: CD152 has been implicated in tolerance induction. This study investigated how CD80 and CD86 regulated CD152 expression in a low-responding cardiac transplant model with CD152-mediated long-term graft acceptance. METHODS: A low-responding cardiac transplant model from BALB/c to B10.A was used. Donor specific stimulation and multiple antibody blockade of the CD80/CD86:CD28/CD152 co-stimulatory pathway was applied to the splenic T cells from B10.A recipients with 100-day grafts (B10.A-100). Proliferation assays, quantitative (Q) real-time polymerase chain reaction (PCR), flow cytometric analyses, and fluorescence microscopy were conducted to examine the roles of CD80 and CD86 in CD152 expression. RESULTS: B10.A-100 splenic T cells were hyporesponsive to donor specific stimulation, and anti-CD80, anti-CD86, or anti-CD152 treatment significantly enhanced the proliferation response of the B10.A-100 splenic T cells. Proliferation assays and Q-PCR revealed that CD152 inhibited T-cell proliferation and, at the same time, decreased CD152 expression by secluding CD80 and CD86 from CD28 engagement. Flow cytometric analyses and fluorescence microscopy showed that CD28 engagement facilitated intracellular accumulation of CD152. Besides, CD152 engagement by CD80 decreased CD152 mRNA transcription, and CD152 engagement by CD86 inhibited surface expression of CD152. CONCLUSIONS: CD80 and CD86 controlled CD152-mediated allograft tolerance by multiple negative feedbacks on CD152 mRNA and surface expression. PMID- 15665766 TI - Combined vascular endothelial growth factor and platelet-derived growth factor inhibition in rat cardiac allografts: beneficial effects on inflammation and smooth muscle cell proliferation. AB - BACKGROUND: Perivascular inflammation and subsequent smooth muscle cell (SMC) proliferation are central in the development of cardiac allograft arteriosclerosis. We examined the effect of combined inhibition of proinflammatory vascular endothelial growth factor (VEGF) and SMC mitogen platelet-derived growth factor (PDGF) in rat cardiac allografts. METHODS: Heterotopic cardiac transplantations were performed between fully major histocompatibility mismatched rat strains receiving cyclosporine A immunosuppression. In situ hybridization and immunohistochemistry were performed to examine VEGF and PDGF ligand and receptor (R) expression. Protein tyrosine kinase inhibitors PTK787 and imatinib were used to inhibit VEGFR and PDGFR activity, respectively. Rat coronary artery SMC migration and proliferation assays were used to examine the effect of VEGF and PDGF and tyrosine kinase inhibitors in vitro. RESULTS: Both ligand and receptor expression of VEGF and PDGF were detected in chronically rejecting allografts. In vitro, PDGF-BB mediated rat coronary artery SMC migration and proliferation was completely inhibited with imatinib and partially with PTK787. In vivo, combined treatment with PTK787 and imatinib significantly reduced the formation of neointimal lesions in arteries of cardiac allografts at 8 weeks, producing a greater effect than either drug alone. PTK787, in contrast with imatinib, reduced the number of ED1 macrophages and PDGF-B immunoreactivity in the allografts at 4 weeks. CONCLUSIONS: Blocking VEGF and PDGF receptor signaling in cardiac allografts has distinctive effects on inflammation and SMC proliferation, suggesting that targeting both inflammation and pathologic vascular remodeling may be needed to inhibit cardiac allograft arteriosclerosis. PMID- 15665767 TI - Immunologic benefits of longer graft in rat allogenic small bowel transplantation. AB - BACKGROUND: The effect of graft length on rejection reaction in small bowel transplantation (SBT), which is poorly understood, is tested using rat allogenic SBT models with a short course of tacrolimus. MATERIALS AND METHODS: Inbred Brown Norway rats (major histocompatibility complex: RT1) and Lewis rats (RT1) were used as donors and recipients, respectively. The intestinal tract of the recipient was partially or totally replaced by segmental (15 cm) or whole (70 cm) donor intestine, using two different SBT models. With tacrolimus treatment (0.64 mg/kg per day, 0-13 postoperative days, intramuscularly), recipients' body weights and their survival were evaluated. To compare the extent of peripheral chimerism, donor passenger leukocytes were followed using flow cytometry with a donor-specific monoclonal antibody, OX-27. For the periodical histologic analysis, heterotopic SBT and protocol biopsies of the graft were also performed with short or long intestinal grafts. RESULTS: In a classical Monchik and Russell orthotopic SBT model, whole SBT recipients survived more than 60 days. However, all of the allogenic segmental SBT recipients died within 14 days without histologic sign of graft rejection. In the modified orthotopic SBT model using a cuff technique without systemic clamping, all recipients with segmental allograft survived longer than 29 days. However, recipients with whole graft tended to survive longer than those with segmental graft. The suffering period, lasting from the onset of rejection to death, was significantly shorter in the segmental group than in the whole group. Flow cytometric analysis showed that recipients with whole intestinal grafts had significantly higher ratio of donor passenger leukocytes in peripheral blood. Histologic studies of the protocol biopsies showed that the shorter graft tended to be more severely rejected than the longer graft. CONCLUSIONS: We have demonstrated experimentally that long intestinal grafts have immunologic advantage over short grafts. PMID- 15665768 TI - Ethyl pyruvate ameliorates liver ischemia-reperfusion injury by decreasing hepatic necrosis and apoptosis. AB - BACKGROUND: Hepatic ischemia-reperfusion injury (I/R) occurs in the settings of transplantation, trauma, and elective liver resections. Reactive oxygen species (ROS) have been shown to play a major role in organ I/R injury. Pyruvate, a key intermediate in cellular metabolism, is an effective scavenger of ROS. The purpose of this study was to test the hypothesis that ethyl pyruvate (EP), a soluble pyruvate derivative, is effective in preventing hepatic I/R injury. METHODS: Lewis rats underwent 60 minutes of partial warm hepatic ischemia. Three doses of EP dissolved in lactated Ringer's solution or lactated Ringer's solution (LR) alone were given by intravenous injection. Serum and tissue samples were obtained at 1 to 24 hours postreperfusion. RESULTS: Serum transaminases, degree of hepatic necrosis, and neutrophil infiltration were all significantly decreased in the EP-treated rats compared with control animals. The amount of hepatic lipid peroxidation was also significantly decreased in EP-treated animals. Both circulating levels and hepatic expression of inflammatory cytokines were significantly decreased in the EP-treated animals. Furthermore, EP inhibited activation of extracellular signal-regulated kinase, p38, and c-Jun N-terminal kinase mitogen-activated protein kinases, as well as nuclear factor-kappaB, signaling pathways involved in cytokine release. Treatment with EP also inhibited hepatic apoptosis. CONCLUSION: EP has a protective effect on hepatic I/R injury, mediated in part by decreasing lipid peroxidation, down-regulation of inflammatory mediators, and inhibition of apoptosis. Strategies using this additive to LR solution should be considered in clinical settings of ischemic liver injury to decrease organ damage. PMID- 15665769 TI - Risk factors for reaching renal endpoints in the assessment of Lescol in renal transplantation (ALERT) trial. AB - BACKGROUND: The aim of the study was to identity risk factors for long-term renal transplant function and development of chronic allograft nephropathy (CAN) in renal transplant recipients included in the Assessment of Lescol in Renal Transplantation (ALERT) trial. METHODS: The ALERT trial was a randomized, double blind, placebo-controlled study of the effect of fluvastatin, 40 and 80 mg/day, in renal transplant recipients who were randomized to receive fluvastatin (Lescol) (n = 1,050) or placebo (n = 1,052) over 5 to 6 years of follow-up. Renal endpoints including graft loss or doubling of serum creatinine or death were analyzed by univariate and multivariate regression analysis in the placebo group. RESULTS: There were 137 graft losses (13.5%) in the placebo group, mainly caused by CAN (82%). Univariate risk factors for graft loss or doubling of serum creatinine were as follows: serum creatinine, proteinuria, hypertension, pulse pressure, time since transplantation, donor age, human leukocyte antigen-DR mismatches, treatment for rejection, low high-density lipoprotein cholesterol, and smoking. Multivariate analysis revealed independent risk factors for graft loss as follows: serum creatinine (relative risk [RR], 3.12 per 100-microM increase), proteinuria (RR, 1.64 per 1-g/24 hr increase), and pulse pressure (RR, 1.12 per 10 mm Hg), whereas age was a protective factor. With patient death in the composite endpoint, diabetes mellitus, smoking, age, and number of transplantations were also risk factors. CONCLUSIONS: Independent risk factors for graft loss or doubling of serum creatinine or patient death are mainly related to renal transplant function, proteinuria, and blood pressure, which emphasizes the importance of renoprotective treatment regimens in this population. PMID- 15665770 TI - Self-organizing maps can determine outcome and match recipients and donors at orthotopic liver transplantation. AB - BACKGROUND: There is a relative lack of donor organs for liver transplantation. Ideally, to maximize the utility of those livers that are offered, donor and recipient characteristics should be matched to ensure the best possible posttransplant survival of the recipient. METHODS: With prospectively collected data on 827 patients receiving a primary liver graft for chronic liver disease, we used a self-organizing map (SOM) (one form of a neural network) to predict outcome after transplantation using both donor and recipient factors. The SOM was then validated using a data set of 2622 patients undergoing transplantation in the United Kingdom at other centers. RESULTS: SOM analysis using 72 inputs and two survival intervals (3 and 12 months) yielded three neurons with either higher or lower probabilities of survival. The model was validated using the independent data set. With 20 patients on the waiting list and 10 sequential donor livers, it was possible to demonstrate that the model could be used to identify which potential recipients were likely to benefit most from each liver offered. CONCLUSIONS: With this approach to matching donor livers and recipients, it is possible to inform transplant clinicians about the optimum use of donor livers and thereby effectively make the best use of a scarce resource. PMID- 15665771 TI - Impaired dendritic cell immunophenotype and function in heart transplant patients undergoing active cytomegalovirus infection. AB - BACKGROUND: Human cytomegalovirus (HCMV) infects dendritic cells (DCs) in vitro and inhibits their maturation properties and their ability to stimulate T-cell proliferation and cytotoxicity. This study analyzed HCMV infection of DCs in vivo. METHODS: We compared blood DCs and monocyte-derived DCs from heart transplant patients undergoing an acute HCMV infection with DCs obtained from HCMV-negative transplant patients. Diagnosis of active HCMV infection was established by antigenemia test. RESULTS: We detected viral RNA and antigens in defined DC subsets obtained from patients undergoing an active HCMV infection. In addition, we found an impaired immunophenotype in immature DCs from HCMV-positive subjects and a reduced ability of mature DCs from the same group of patients to stimulate allogenic T-cell proliferation. CONCLUSIONS: The impaired immunophenotype and function detected in DCs from transplant patients undergoing an active HCMV infection may be a mechanism used by the virus to interfere with early immune functions and thereby contributing to the HCMV-induced immunosuppression in these patients. PMID- 15665772 TI - Implications of immunohistochemical detection of C4d along peritubular capillaries in late acute renal allograft rejection. AB - BACKGROUND: Immunohistochemical detection of the C4d complement product along peritubular capillaries (PC) may indicate humoral rejection of renal allografts. We examined the frequency of PC C4d expression in renal-allograft biopsies with acute rejection (AR) arising more than 6 months after transplantation and the impact of this finding. METHODS: C4d was detected by immunoperoxidase in 2-micron paraffin sections of consecutive biopsies obtained over a 3-year period. The extent was classified as diffuse (> or =50% PC C4d+), focal (<50% C4d+), and negative (C4d-). Clinical data were obtained by retrospective chart review. Fifty five AR episodes with Banff 97 types 1A (n = 13), 1B (n = 26), 2A (n = 11), 2B (n = 3), and 3 (n = 2) met inclusion criteria. RESULTS: PC C4d expression was diffuse in 23 (42%), focal in 9 (16%), and negative in 23 (42%) biopsies. AR episodes with focal and diffuse C4d expression had higher proportionate elevation of serum creatinine at biopsy and 4 weeks after diagnosis (P< or =0.05). Biopsies with diffuse PC C4d had interstitial hemorrhage (56.5%) and plasmacytic infiltrates (52%) more frequently than C4d- biopsies (22% and 16%), P = 0.02, but had no other distinctive histologic features. Graft loss was greater in diffuse (65%) compared with focal C4d+ (33%) and C4d- (33%) groups 1 year after diagnosis, P = 0.03. Other clinical and pathologic parameters did not differ significantly, including treatment received for AR. CONCLUSION: Evidence of acute cellular with occult humoral rejection is identified in more than 40% of late AR episodes. Late acute humoral rejection may be associated with interstitial hemorrhage and plasma cells and contributes significantly to graft loss. PMID- 15665773 TI - Measurement of chimerism in cynomolgus monkeys using human-specific short tandem repeat-based assay. AB - Preclinical testing of a mixed chimerism mediated organ transplant tolerance strategy, in a cynomolgus macaque model, would be facilitated by the establishment of a reliable technique for quantitative assessment of chimerism. Among various techniques used for measurement of chimerism in humans, microsatellite DNA profiling has been considered the most versatile one that can discriminate between two individuals. We adopted a commercially available short tandem repeat profiling methodology to cynomolgus monkeys using two human specific alleles, TPOX and CSF1PO. Polymerase chain reaction (PCR) was used to amplify these alleles, and the analysis of the PCR products was performed by capillary electrophoresis. Of 54 cynomolgus macaques investigated, only one pair with the same ABO blood type demonstrated identity at both alleles. This implies that this technique should interfere minimally with the assignment of donor recipient pairs based upon molecular tissue typing or mixed lymphocyte cultures. PMID- 15665774 TI - Gene silencing in rat-liver and limb grafts by rapid injection of small interference RNA. AB - Genetic modification is a promising therapeutic strategy for organ transplantation. In previous studies, we introduced a nonviral method of gene transfer to donor grafts using an organ-selective injection technique to up regulate gene expression. Based on the attractive methodology of RNA interference for silencing a particular gene expression, we applied our catheter-based injection method to transfer small interference RNA (siRNA)-green fluorescence protein (GFP) into liver and limb grafts. We first quantified the interfering activity after the systemic delivery of siRNA in the liver of Alb-DsRed2 transgenic (Tg) rats using in vivo bioimaging system. Then, using GFP Tg Lewis rats as donors, transient down-regulation of the GFP expression was achieved both in liver- and limb-transplantation models by the preoperative rapid injection of siRNA. Genetic modification by siRNA may provide new therapeutic options for down regulation of endogenous antigenicity. PMID- 15665776 TI - Cytomegalovirus and thromboembolism in renal transplantation. PMID- 15665775 TI - Differences between early and late posttransplant lymphoproliferative disorders in solid organ transplant patients: are they two different diseases? AB - BACKGROUND: The objective of the authors' study was to characterize the clinical and pathologic differences between patients who develop posttransplant lymphoproliferative disorders (PTLD) early or late after transplantation and to assess the overall survival in these two groups. METHODS: One hundred seven adult solid organ transplant patients were identified at the Mayo Clinic between December 1970 and May 2003. RESULTS: Forty-nine patients developed PTLD within the first year (early PTLD, 1-11.8 months) and 58 patients developed PTLD after 1 year (late PTLD, 14 months-17 years). Patients with early PTLD more commonly had the following characteristics: positive Epstein-Barr virus (EBV) in situ hybridization status (P < 0.0001), CD20-positive status (P = 0.002), and involvement of the grafted organ (P = 0.02). Overall survival did not differ between the two groups (P = 0.25). PTLD may occur in two different settings with different characteristics. CONCLUSIONS: Early PTLD is more commonly EBV in situ hybridization-positive and CD20-positive, and more commonly involves the grafted organ. PMID- 15665777 TI - Venous thromboembolism in renal transplant recipients. PMID- 15665778 TI - Human hepatocyte isolation for liver cell therapy: whole marginal livers or healthy segments from splitting? PMID- 15665779 TI - Is the application of HLAMatchmaker relevant in kidney transplantation? PMID- 15665781 TI - Sirolimus and ACE-inhibitors: a note of caution. PMID- 15665782 TI - Cyclosporine and low-dose ketoconazole in renal transplant recipients: a single center experience. PMID- 15665783 TI - Role of ketoconazole in renal transplant recipients. PMID- 15665784 TI - Aspirin promotes kidney allograft survival and function. PMID- 15665785 TI - The T-tube approach to biliary strictures in liver transplant recipients. PMID- 15665786 TI - Hematopoietic stem cell transplantation and subsequent 80% skin exchange by grafts from the same donor in a patient with Herlitz disease. PMID- 15665787 TI - Intravenous immunoglobulin treatment in a kidney transplant patient with chronic allograft nephropathy. PMID- 15665788 TI - Laparoscopic nephrectomy donor death due to cerebral gas embolism in a specialized transplant center: risk zero does not exist. PMID- 15665789 TI - Hypophosphatemia and the live liver donor. PMID- 15665790 TI - Native kidney BK nephropathy post cardiac transplantation. PMID- 15665791 TI - Registration of clinical trials. PMID- 15665792 TI - Complex coronary intervention in the drug-eluting stent era. Introduction. PMID- 15665793 TI - Choosing a drug-eluting stent: a comparison between CYPHER and TAXUS. AB - For patients with coronary artery disease undergoing percutaneous intervention, drug-eluting stents (DESs) have rapidly become the standard of care. This article reviews the currently available delivery-platform/drug-carrier-vehicle combinations and those expected to become available in the future. It also evaluates and compares current DES platforms in terms of the drug involved, the delivery platform, efficacy, and safety. Currently, 2 DES platforms are available: 1 eluting sirolimus and 1 eluting paclitaxel. Sirolimus is a macrolide antibiotic with a cytostatic mechanism and an anti-inflammatory effect. Paclitaxel is a chemotherapeutic (cytotoxic) agent. The delivery platform is composed of the balloon catheter, the stent, and the drug-carrier vehicle. The carrier vehicle offers controlled drug release and enhances drug distribution. It can be a polymer that serves as a diffusion barrier or a matrix (either durable or degradable) for drug loading. Alternatively, a structural modification on the surface of the stent itself, such as a groove or well in which the drug is placed, can serve as carrier. With respect to efficacy, major trials have shown that the sirolimus platform has a lower late luminal loss rate than does the paclitaxel stent. Moreover, less intimal proliferation and obstruction occurs with the sirolimus platform than with the paclitaxel platform. Also, compared to bare metal stents, the sirolimus platform reduces late luminal loss in challenging subsets of patients. Both stents offer excellent short-term safety. To improve our understanding of these stents, a head-to-head comparison is needed. PMID- 15665794 TI - Safety and efficacy of drug-eluting stents. AB - This review characterizes the relative effectiveness and safety of drug-eluting stents (DES) compared to bare metal stents. The data evaluated will include clinical and angiographic outcomes from randomized clinical trials as well as observational databases and registries including RESEARCH, e-Cypher, and DYNAMIC, which typically include a much broader spectrum of patients. In addition, specific patient subsets, including those patients with multivessel coronary disease, will be evaluated. PMID- 15665795 TI - Drug-eluting stents: clinical perspectives on drug and design differences. AB - Although the introduction of coronary stents has significantly improved the treatment of patients with coronary artery disease, restenosis, due to neointimal proliferation following stent deployment and associated with a return of ischemic symptoms, has remained a critical concern. Recent studies have shown that the use of drug-eluting stents to deliver antiproliferative agents directly to the vessel wall dramatically reduces the rate of restenosis. However, important differences exist among stent designs, drug-delivery vehicles, and choices of pharmacologic agents that can significantly affect the safety and efficacy of each device. Although engineers, vascular biologists, and clinicians all agree that clinical success of drug-eluting stents requires careful integration of the individual system components, the optimal combination remains to be determined. PMID- 15665796 TI - The approach to small vessels in the era of drug-eluting stents. AB - The treatment of small-vessel disease will occupy an increasingly important part of interventional cardiology practice and this raises several issues. The definition of "small vessels" has great implications for device size selection, and knowledge of "normal" small-vessel dimensions is important. Stents have been applied in the setting of smaller-vessel disease and future iterations of small vessel stents will need to address several design factors. Stent strut thickness might impact on subsequent restenosis as well as late lumen loss. There has been great interest in the use of drug-eluting stents for small vessels. Randomized clinical trials of sirolimus-eluting versus bare metal stents in the treatment of small-vessel disease have shown significant improvements in the rates of target lesion revascularization and restenosis with sirolimus-eluting stents. These improvements in restenosis rates are attributable to the low levels of late loss with the drug-eluting stent. PMID- 15665797 TI - Drug-eluting stents for emerging treatment strategies in complex lesions. AB - The use of drug-eluting stents (DES) has rapidly expanded from lower-risk, single lesion procedures to include a broad spectrum of high-risk patients and complex lesions. For 4 complex patient subgroups, emerging data suggest that DES might offer an advantage for reducing late clinical restenosis. In ST elevation myocardial infarction, early registry reports are promising, with no evidence to date for an increased incidence of subacute stent thrombosis and significant trends for less restenosis. For chronic total occlusions, early, small clinical series show that DES might have unprecedented long-term patency. Initial registries of DES for in-stent restenosis reveal striking reductions in late loss and restenosis, compared with brachytherapy historical controls. The use of DES in saphenous vein graft lesions is increasing, and early registry results show a very acceptable incidence of thromboembolic complications and major adverse cardiac events. Important data regarding much larger groups of these patient cohorts will emerge over the next year, to help guide the broader application of DES in "real-world" practice. PMID- 15665798 TI - Percutaneous coronary revascularization of diabetic patients in the era of drug eluting stents. AB - Patients with diabetes have worse clinical outcomes following both surgical revascularization and percutaneous coronary intervention (PCI). Although coronary stenting has improved late outcomes (versus balloon angioplasty) following PCI, both angiographic restenosis and the requirement for repeat revascularization are increased in diabetics versus nondiabetics and limit the durability of PCI compared with surgery. Polymer-based drug-eluting stents (DES) have markedly reduced late coronary lumen loss and angiographic restenosis as well as the need for repeat revascularization when compared with conventional (non-drug-eluting) coronary stent deployment. Specifically, the CYPHER sirolimus-eluting stent (Cordis Cardiology, Miami Lakes, FL) has demonstrated durable clinical and angiographic benefit for diabetic patients in both randomized clinical trials and postmarket surveillance registries. Data on the more recently approved paclitaxel eluting TAXUS (Boston Scientific, Natick, MA) stent suggest similar efficacy for the treatment of diabetic patients. By markedly reducing restenosis, DES significantly improve or eliminate the major limitation of conventional stenting/PCI in diabetic patients. The advent of DES promises a paradigm shift from surgical revascularization in diabetic patients (especially those with multivessel disease) to PCI. Nevertheless, continued improvement in DES delivery as well as optimal adjunctive pharmacotherapy and control of hyperglycemia will be required to achieve the best clinical outcomes following PCI with DES in patients with diabetes. PMID- 15665799 TI - Mechanism of ipamorelin-evoked insulin release from the pancreas of normal and diabetic rats. AB - OBJECTIVE: To examine the effect of ipamorelin (IPA), a novel pentapeptide with a strong growth hormone releasing potency, on insulin secretion from pancreatic tissue fragments of normal and diabetic rats. MATERIALS AND METHODS: Diabetes mellitus was induced by streptozotocin (60 mg kg(-1)). Four weeks after the induction of diabetes, pancreatic tissue fragments of normal and diabetic rats were removed and incubated with different concentrations (10(-12) - 10(-6) M) of IPA. Insulin release from the pancreas was measured by radioimmunoassay. RESULTS: Ipamorelin evoked significant (p<0.04) increases in insulin secretion from the pancreas of normal and diabetic rats. Either diltiazem or yohimbine or propranolol or a combination of atropine, propranolol and yohimbine inhibited IPA evoked insulin secretion significantly (p<0.03) from the pancreas of normal and diabetic rats. Atropine caused a significant (p<0.007) reduction in the IPA induced insulin secretion in diabetic but not in normal rats. CONCLUSION: IPA stimulates insulin release through the calcium channel and the adrenergic receptor pathways. This is the first study to examine the effect of ipamorelin on insulin secretion in the pancreas. PMID- 15665800 TI - Paradoxical firing of thalamic neurons under neuropathic pain state in rats. AB - OBJECTIVES: A novel evaluative approach was used to determine single unit activities of non-bursting intralaminar thalamic nuclei under neuropathic pain state following dorsal rhizotomy. SETTINGS AND DESIGN: Extensive dorsal rhizotomy at cervicothoracic level in rats was used as a model of central pain. After rhizotomy, rats were divided into two groups: rats without any signs of self mutilation, and those presenting self-mutilation. Spontaneous single unit activities of neurons of intralaminar thalamic nuclei were recorded and interspike intervals (ISIs) of non-bursting cells were counted for both groups and compared with that of non-rhizotomized control rats. Chaodynamic methods were applied for the evaluation of the ISIs. RESULTS: In control rats Lyapunov exponents, Shannon entropy and mutual information average values were significantly higher than those of rhizotomized rats without any signs of self mutilation. Paradoxically, in animals presenting self-mutilation following rhizotomy the evaluated parameters were similar to those of controls. Further, Lyapunov exponents were positive values in all animals indicating chaotic pattern of the neuronal firing. MAIN FINDINGS: 1. Neurons behave in chaotic way in all animals, 2. The most regular firing was found in non-mutilating rhizotomized animals, 3. Patterns of the firing in selfmutilating rats were similar to those in controls. CONCLUSIONS: It is concluded that pain feeling is not executed neither by changes of chaotic dynamics of non-bursting intralaminar thalamic neurons. On the other hand, the paradoxical firing of the neurons under pathological brain matrix might participate in modification pain feeling. PMID- 15665801 TI - Melatonin circadian rhythm in women with idiopathic hyperprolactinemia. AB - OBJECTIVES: Relationship between melatonin and prolactin has been suggested on the basis of both experimental and clinical studies. However, there are scarce and controversial data concerning melatonin concentrations in hyperprolactinemic patients. Therefore, the aim of the present study was to evaluate the circadian rhythm of melatonin in female patients suffering from hyperprolactinemia. MATERIAL AND METHODS: The study was performed on 28 women aged 17-58 years (mean+/-SEM: 30.5+/-1.8 years) suffering from idiopathic hyperprolactinemia and 14 healthy volunteers aged 20-50 years (mean+/-SEM: 36.4+/-3.0 years) with normal prolactin levels. Blood samples for measurements of serum melatonin and prolactin concentrations were collected at 08:00, 12:00, 16:00, 20:00, 24:00, 02:00, 04:00, 06:00, and 08:00 h. Melatonin and prolactin concentrations were measured by enzyme immunoassay. RESULTS: Significant increase in melatonin serum concentrations was also observed in hyperprolactinemic patients in comparison with healthy volunteers during the night. Similar increase was also observed in the area under curve of melatonin concentrations. However, no correlation was found between prolactin and melatonin concentrations in any examined time points. CONCLUSION: The results of the present study confirm suggestions of the presence of the relationship between melatonin and prolactin secretion. PMID- 15665802 TI - Declining melatonin levels and older people. How old is old? AB - OBJECTIVES: The purpose of this study was to investigate whether melatonin levels in older cohorts within the 'aged' population were significantly lower than in younger 'aged' individuals and hence point to a possible confound in cross sectional studies which group individuals over 55 in one category. SETTING AND DESIGN: Melatonin levels of 35 North Queensland residents over 55 years of age living in an aged-care facility, a retirement village or the general community were compared across three age groups. METHODS: Subjects were ten 56-65 year olds, eighteen 66-75 year olds and seven over-75 year olds. Information was obtained on sleep, awakening, medical conditions and medications, with subsequent exclusion of those with known medical conditions and/or medications. Melatonin was collected by salivary samples at 2200 hours and concentrations were determined by immunoassay. RESULTS: Mean melatonin levels were significantly lower (p=.03) in the 'oldest' (over 75 yrs) group compared to the 'youngest' (56 65 yrs) group. MAIN FINDINGS: The results of this preliminary study indicate that within the older population, melatonin levels appear to decline significantly with age. CONCLUSIONS: Future studies of melatonin and ageing may benefit from a longitudinal approach, with older subjects sampled across time. PMID- 15665803 TI - Hyperprolactinemia after low dose of amisulpride. AB - OBJECTIVES: Amisulpride in antipsychotic doses can induce hyperprolactinemia. The aim of this study was to prove whether the same is true for low doses of amisulpride. METHODOLOGY: Plasma prolactin levels were measured in 5 males and 5 females with depressive symptoms who were treated with 50 mg of amisulpride per day as an augmentation to antidepressants (n=5), benzodiazepine anxiolytics (n=8) or in monotherapy (n=1). Six of these patients were assessed prior to onset of amisulpride treatment and after 10 days of amisulpride use. Four patients had been using amisulpride for more than a month. RESULTS: There was a significant increase of prolactin levels from mean 16+/-6 ng/ml to 113+/-65 ng/ml (median 14.5 ng/ml to median 92 ng/ml; Wilcoxon matched pair test, p=0.027). All patients had hyperprolactinemia (30-200 ng/ml). The prolactinemia was significantly higher in females (mean 160+/-50 ng/ml; median 198 ng/ml) than in males (mean 48+/-12 ng/ml; median 48 ng/ml; Mann-Whitney U test, p=0.041). CONCLUSION: Even low doses of amisulpride used as an augmentation to antidepressant treatment, benzodiazepines or in monotherapy seem to be associated with hyperprolactinemia. The co-medication of antidepressants and benzodiazepines can potentially increase intensity of prolactinemia. PMID- 15665804 TI - The presence of heme-oxygenase and biliverdin reductase in human cranial ganglia indicates a role for carbon monoxide in neural transmission. AB - OBJECTIVES: The purpose of this study was to localize in cranial ganglia of man the occurrence of the putative gaseous neural messenger carbon monoxide (CO) and the biliverdin degrading enzyme biliverdin reductase (BVR). METHODS: Immunocytochemistry with antibodies against the CO-inducing enzymes HO-1, HO-2, BVR and calcitonin gene-related peptide was used. RESULTS: In the trigeminal ganglion about 60% of the cell bodies exhibited HO-2- and about 40% BVR immunoreactivity. HO-2- as well as BVR-immunoreactivity was predominantly (78%) expressed in medium-sized cells (30-60 microm). In the superior cervical ganglion about 40% of the cell bodies exhibited HO-2-immunoreactivity. In the sphenopalatine and otic ganglia only a few cell bodies were HO-2 immunoreactive. HO-1, the inducible isoform of heme oxygenase, gave only very weak immunoreactivity in all ganglia examined. Double immunostaining revealed that in the human trigeminal ganglion HO-2 and BVR co-localized with calcitonin gene related peptide. CONCLUSIONS: The finding suggests that CO might serve as a modulator of synaptic transmission in man. PMID- 15665805 TI - Influence of the operative treatment of leiomyomas on lipid profile. AB - OBJECTIVE: The aim of this study was to asses the influence of operative treatment of leiomyomas on lipid profile. The study was conducted from April 1, 2000, through April 30, 2002 in The Department of Septic Gynecology and Obstetrics at Collegium Medicum of the Jagiellonian University. METHODOLOGY: Most hysterectomies are performed because of leiomyomas, which are the most common benign tumors of the female reproductive organ. Pathogenesis and evolution of uterine leiomyomas are not well understood. Treatment of leiomyomas is divided into operative and inoperative. Operative methods include generally laparoscopic myomectomy, conservative myomectomy and hysterectomy. 130 women with diagnosed leiomyomas were analyzed in four groups. CONCLUSION: Regardless of the type of performed surgery, decrease in cholesterol, HDL and LDL are observed 10 days after operation, with return to preoperative values at 6 months after surgical procedure. The highest level of cholesterol and LDL (6 moths from surgery) is observed in patients who underwent hysterectomy with bilateral oophorectomy. PMID- 15665806 TI - 5-Hydroxyindoleacetic acid and homovanillic acid concentrations in cerebrospinal fluid in patients with Alzheimer's disease, depression and mild cognitive impairment. AB - In this study we investigated the cerebrospinal fluid (CSF) concentrations of 5 hydroxyindoleacetic acid (5-HIAA) and homovanillic acid (HVA) in Alzheimer (AD) patients (n=75), patients with mild cognitive impairment (MCI, n=9) and patients with depression (n=7). CSF HVA was significantly elevated in AD with depression (Geriatric Depression Scale, 15 point version GDS>5) in comparison to AD without depression (p<0.05, ANOVA) and CSF HVA showed a significant positive correlation with the GDS score of AD-patients (p=0.03, Spearman Rho: 0.38, Spearman Rank Correlation). In the group of AD patients CSF 5-HIAA was positively correlated with cerebrospinal fluid beta-amyloid 1-42 (Abeta42), p<0.05, Spearman Rho: 0.3, Spearman Rank Correlation, but not with CSF tau. Additionally, there was a significant positive correlation between cerebrospinal fluid 5-HIAA and HVA in the group of AD patients (p<0.0001, Rho: 0.47, Spearman Rank correlation). Neither 5-HIAA nor HVA in CSF could differentiate between mild cognitive impairment, depression and AD. The results of this study support the hypothesis that the serotonergic system plays a role in the course of AD. They further suggest an important role of dopamine metabolism in depression within AD patients. PMID- 15665807 TI - The naloxone test in Klinefelter syndrome. AB - AIM TO THE STUDY: The investigation of the influence of anti-opioid drug on hypophyseal and gonadal hormones secretion in case of Klinefelter syndrome. MATERIAL AND METHODS: The naloxone test (0,4 mg iv) was performed in 14 patients with Klinefelter syndrome aged from 19 to 32 and in 12 age matched control subjects with azoospermia and normal spermatogenesis in testicular histology. The plasma levels of FSH, LH, prolactin, testosterone and estradiol were established before and after 30, 60, 60 and 120 minutes respectively, following the drug administration. RESULTS: Basal FSH, prolactin and estradiol levels were significantly higher whereas basal testosterone was significantly lower in patients with Klinefelter syndrome than in the control group. After the naloxone administration the mean plasma prolactin level decreased significantly (p=0.01) in Klinefelter subjects. The respective diminution in control group was not significant. The levels of FSH and LH as well as testosterone and estradiol did not change during the naloxone test in both Klinefelter and control subjects. CONCLUSIONS: The naloxone administration in Klinefelter syndrome caused the decrease in plasma prolactin levels but did not affect the plasma level of another hypophyseal and gonadal hormones. The opioid controlled gonadotropin secretion is altered in case of Klinefelter syndrome. PMID- 15665815 TI - DNA repair in post-mitotic neurons: a gene-trapping strategy. PMID- 15665816 TI - Inhibition of histone deacetylase 2 increases apoptosis and p21Cip1/WAF1 expression, independent of histone deacetylase 1. AB - Histone deacetylases (HDACs) 1 and 2 share a high degree of homology and coexist within the same protein complexes. Despite their close association, each possesses unique functions. We show that the upregulation of HDAC2 in colorectal cancer occurred early at the polyp stage, was more robust and occurred more frequently than HDAC1. Similarly, while the expression of HDACs1 and 2 were increased in cervical dysplasia and invasive carcinoma, HDAC2 expression showed a clear demarcation of high-intensity staining at the transition region of dysplasia compared to HDAC1. Upon HDAC2 knockdown, cells displayed an increased number of cellular extensions reminiscent of cell differentiation. There was also an increase in apoptosis, associated with increased p21Cip1/WAF1 expression that was independent of p53. These results suggest that HDACs, especially HDAC2, are important enzymes involved in the early events of carcinogenesis, making them candidate markers for tumor progression and targets for cancer therapy. PMID- 15665817 TI - Apoptosis-based therapies and drug targets. AB - The pathogenesis of many diseases is most closely connected with aberrantly regulated apoptotic cell death. The past 15 years have witnessed an explosion in the basic knowledge of mechanisms that regulate apoptosis and the mediators that either trigger or inhibit cell death. Consequently, great interest has emerged in devising therapeutic strategies for modulating the key molecules of life-and death decisions. Numerous novel approaches are currently being followed employing gene therapy and antisense strategies, recombinant biologics or classical organic and combinatorial chemistry in order to target specific apoptotic regulators. Although drug development is still in its infancy, several therapeutics have progressed to clinical testing or have even been approved in record time. This review outlines the recent advances in the field of apoptosis-based therapies and explores some highlights of a very active field of drug development. PMID- 15665819 TI - p38 kinase mediates nitric oxide-induced apoptosis of chondrocytes through the inhibition of protein kinase C zeta by blocking autophosphorylation. AB - This study investigated the molecular mechanisms underlying inhibition of protein kinase C (PKC) zeta by p38 kinase during nitric oxide (NO)-induced apoptosis of chondrocytes. Coimmunoprecipitation experiments showed that activation of p38 kinase following addition of an NO donor resulted in a physical association between PKCzeta and p38 kinase. Direct interaction of p38 kinase with PKCzeta was confirmed in vitro using p38 kinase and PKCzeta recombinant proteins. p38 kinase interacts with the regulatory domain of PKCzeta and its association blocked PKCzeta autophosphorylation. Micro LC-MS/MS analysis using recombinant proteins indicated that the interaction of p38 kinase with PKCzeta blocked autophosphorylation of PKCzeta on Thr-560, which is required for PKCzeta activation. Collectively, our results demonstrate a novel mechanism of PKCzeta regulation: following activation by the production of NO, p38 kinase binds directly to the PKCzeta regulatory domain, preventing PKCzeta autophosphorylation on Thr-560, thereby inhibiting PKCzeta activation. PMID- 15665818 TI - Akt inhibition upregulates FasL, downregulates c-FLIPs and induces caspase-8 dependent cell death in Jurkat T lymphocytes. AB - In T lymphocytes, the role of Akt in regulating Fas/Fas ligand (FasL)-mediated apoptotic signaling and death is not clearly understood. In this study, we observed that inhibition of Akt causes enhanced expression of FasL mRNA and protein and increased death-inducing signaling complex (DISC) formation with Fas associated death domain (FADD) and procaspase-8 recruitment. Also, caspase-8 was activated at the DISC with accompanying decrease in c-FLIPs expression. FasL neutralizing antibody significantly decreased apoptotic death in the Akt inhibited T cells. Additionally, Akt inhibition-induced Fas signaling was observed to link to the mitochondrial pathway via Bid cleavage. Further, inhibition of caspase-8 activity effectively blocked the loss of mitochondrial membrane potential and DNA fragmentation, suggesting that DISC formation and subsequent caspase-8 activation are critical initiating events in Akt inhibition induced apoptotic death in T lymphocytes. These data demonstrate yet another important survival function governed by Akt kinase in T lymphocytes, which involves the regulation of FasL expression and consequent apoptotic signaling. PMID- 15665820 TI - Antitumor immune responses mediated by adenoviral GDEPT using nitroreductase/CB1954 is enhanced by high-level coexpression of heat shock protein 70. AB - Gene-directed enzyme prodrug therapy (GDEPT) is a promising approach to local management of cancer through targeted chemotherapy. Killing localized tumors by GDEPT in a manner that induces strong antitumor cellular immune responses might improve local management and allow benefit in disseminated cancer. Here we evaluated the combination of nitroreductase (NTR)/CB1954 GDEPT with high-level expression of heat shock protein 70 (HSP70, a stress protein that can shuttle cytosolic peptides into antigen-presenting cells) for induction of antitumor immunity using adenovirus gene delivery in an aggressive and nonimmunogenic BALB/c syngeneic 4T1 breast cancer model. The mechanism of cell death and spectrum of stress proteins induced are likely to be important determinants of the resulting immune responses. We showed that NTR/CB1954 treatment of 4T1 cells gave both apoptotic and nonapoptotic killing. In vivo killing of 4T1 cells expressing NTR gave weak antitumor immunity and very limited induction of stress proteins including HSP70. High-level coexpression of HSP70 during NTR/CB1954 mediated killing of 4T1 cells in vivo gave much greater protection from tumor challenge (67% long-term survivors compared to 17%) and induced 4T1-specific cytotoxic T-cell responses. The enhancement of antitumor responses resulting from HSP70 coexpression was similar to that conferred by coexpression of GM-CSF. PMID- 15665821 TI - Immune characterization of clinical grade-dendritic cells generated from cancer patients and genetically modified by an ALVAC vector carrying MAGE minigenes. AB - Gene delivery into dendritic cells (DC) is most efficiently achieved by viral vectors. Recombinant canarypox viruses (ALVAC) were validated safe and efficient in humans. We aimed firstly to evaluate DC transduction by ALVAC vectors, then to investigate if such infection induced or not the maturation of the DC, and finally to assess the efficiency of ALVAC-MAGE-transduced DC to activate specific CTL clones. Clinical grade DC from melanoma patients were generated from blood monocytes and infected with a recombinant ALVAC virus encoding either a marker gene (EGFP) or the MAGE-1-MAGE-3 minigenes. According to the patient-donor, 22+/ 16% of immature DC were successfully transduced. Flow cytometry analysis of surface markers expressed on DC after ALVAC infection did not reveal a mature phenotype. Moreover, ALVAC transduction did not interfere with the capacity of the DC to further mature under poly:IC stimulation. But most importantly, our results demonstrated that DC from HLA-A1 patient-donors infected with the recombinant ALVAC MAGE-1-MAGE-3 minigenes virus were capable of activating a MAGE 3/A1 CTL clone more efficiently than same DC loaded with MAGE 3/A1 peptide, as shown by increased IFN-gamma secretion. These results could be the basis for the development of a new clinical strategy in melanoma patient's immunotherapy. PMID- 15665822 TI - Widespread intratumoral virus distribution with fractionated injection enables local control of large human rhabdomyosarcoma xenografts by oncolytic herpes simplex viruses. AB - Novel methods of local control for sarcomas are needed. We investigated the antitumor effect of two related herpes simplex virus (HSV) mutants, NV1020 and NV1066, on human rhabdomyosracoma cells and xenografts. Cell death correlated with virus replication and apoptosis in cultured cells and tumors. Complete regression was seen in all tumors <250 mm(3) following a single injection, yet only half of tumors >250 mm(3) showed a complete response. Fractionation of the virus dose into five injection sites did not increase transduction efficiency, transgene expression, or virus production, but did yield more widespread intratumoral distribution. Despite the same total dose of virus, improved control of large tumors was seen using fractionated injections as all large tumors (500 700 mm(3)) had durable, complete regression. Our data suggest that oncolytic HSVs may be useful for local control of bulky rhabdomyosarcoma tumors and that fractionated virus administration results in a more widespread virus infection and better tumor control. Therefore, strategies to maximize intratumoral virus distribution at initial delivery should be sought. PMID- 15665823 TI - Integral role of IRF-5 in the gene induction programme activated by Toll-like receptors. AB - The activation of Toll-like receptors (TLRs) is central to innate and adaptive immunity. All TLRs use the adaptor MyD88 for signalling, but the mechanisms underlying the MyD88-mediated gene induction programme are as yet not fully understood. Here, we demonstrate that the transcription factor IRF-5 is generally involved downstream of the TLR-MyD88 signalling pathway for gene induction of proinflammatory cytokines, such as interleukin-6 (IL-6), IL-12 and tumour necrosis factor-alpha. In haematopoietic cells from mice deficient in the Irf5 gene (Irf5-/- mice), the induction of these cytokines by various TLR ligands is severely impaired, whereas interferon-alpha induction is normal. We also provide evidence that IRF-5 interacts with and is activated by MyD88 and TRAF6, and that TLR activation results in the nuclear translocation of IRF-5 to activate cytokine gene transcription. Consistently, Irf5-/- mice show resistance to lethal shock induced by either unmethylated DNA or lipopolysaccharide, which correlates with a marked decrease in the serum levels of proinflammatory cytokines. Thus, our study identifies IRF-5 as a new, principal downstream regulator of the TLR-MyD88 signalling pathway and a potential target of therapeutic intervention to control harmful immune responses. PMID- 15665824 TI - Complete genome sequence of the acetic acid bacterium Gluconobacter oxydans. AB - Gluconobacter oxydans is unsurpassed by other organisms in its ability to incompletely oxidize a great variety of carbohydrates, alcohols and related compounds. Furthermore, the organism is used for several biotechnological processes, such as vitamin C production. To further our understanding of its overall metabolism, we sequenced the complete genome of G. oxydans 621H. The chromosome consists of 2,702,173 base pairs and contains 2,432 open reading frames. In addition, five plasmids were identified that comprised 232 open reading frames. The sequence data can be used for metabolic reconstruction of the pathways leading to industrially important products derived from sugars and alcohols. Although the respiratory chain of G. oxydans was found to be rather simple, the organism contains many membrane-bound dehydrogenases that are critical for the incomplete oxidation of biotechnologically important substrates. Moreover, the genome project revealed the unique biochemistry of G. oxydans with respect to the process of incomplete oxidation. PMID- 15665825 TI - Admixture mapping for hypertension loci with genome-scan markers. AB - Identification of genetic variants that contribute to risk of hypertension is challenging. As a complement to linkage and candidate gene association studies, we carried out admixture mapping using genome-scan microsatellite markers among the African American participants in the US National Heart, Lung, and Blood Institute's Family Blood Pressure Program. This population was assumed to have experienced recent admixture from ancestral groups originating in Africa and Europe. We used a set of unrelated individuals from Nigeria to represent the African ancestral population and used the European Americans in the Family Blood Pressure Program to provide estimates of allele frequencies for the European ancestors. We genotyped a common set of 269 microsatellite markers in the three groups at the same laboratory. The distribution of marker location-specific African ancestry, based on multipoint analysis, was shifted upward in hypertensive cases versus normotensive controls, consistent with linkage to genes conferring susceptibility. This shift was largely due to a small number of loci, including five adjacent markers on chromosome 6q and two on chromosome 21q. These results suggest that chromosome 6q24 and 21q21 may contain genes influencing risk of hypertension in African Americans. PMID- 15665826 TI - Human telomeric protein TRF2 associates with genomic double-strand breaks as an early response to DNA damage. AB - DNA damage surveillance networks in human cells can activate DNA repair, cell cycle checkpoints and apoptosis in response to fewer than four double-strand breaks (DSBs) per genome. These same networks tolerate telomeres, in part because the protein TRF2 prevents recognition of telomeric ends as DSBs by facilitating their organization into T loops. We now show that TRF2 associates with photo induced DSBs in nontelomeric DNA in human fibroblasts within 2 s of irradiation. Unlike gammaH2AX, a common marker for DSB damage, TRF2 forms transient foci that colocalize closely with DSBs. The TRF2 DSB response requires the TRF2 basic domain but not its Myb domain and occurs in the absence of functional ATM and DNA PK protein kinases, MRE11/Rad50/NBS1 complex and Ku70, WRN and BLM repair proteins. Furthermore, overexpression of TRF2 inhibits DSB-induced phosphorylation of ATM signaling targets. Our results implicate TRF2 in an initial stage of DSB recognition and processing that occurs before association of ATM with DSBs and activation of the ATM-dependent DSB response network. PMID- 15665827 TI - System-level identification of transcriptional circuits underlying mammalian circadian clocks. AB - Mammalian circadian clocks consist of complexly integrated regulatory loops, making it difficult to elucidate them without both the accurate measurement of system dynamics and the comprehensive identification of network circuits. Toward a system-level understanding of this transcriptional circuitry, we identified clock-controlled elements on 16 clock and clock-controlled genes in a comprehensive surveillance of evolutionarily conserved cis elements and measurement of their transcriptional dynamics. Here we report the roles of E/E' boxes, DBP/E4BP4 binding elements and RevErbA/ROR binding elements in nine, seven and six genes, respectively. Our results indicate that circadian transcriptional circuits are governed by two design principles: regulation of E/E' boxes and RevErbA/ROR binding elements follows a repressor-precedes-activator pattern, resulting in delayed transcriptional activity, whereas regulation of DBP/E4BP4 binding elements follows a repressor-antiphasic-to-activator mechanism, which generates high-amplitude transcriptional activity. Our analysis further suggests that regulation of E/E' boxes is a topological vulnerability in mammalian circadian clocks, a concept that has been functionally verified using in vitro phenotype assay systems. PMID- 15665828 TI - Integration of Notch and Wnt signaling in hematopoietic stem cell maintenance. AB - A fundamental question in hematopoietic stem cell (HSC) biology is how self renewal is controlled. Here we show that the molecular regulation of two critical elements of self-renewal, inhibition of differentiation and induction of proliferation, can be uncoupled, and we identify Notch signaling as a key factor in inhibiting differentiation. Using transgenic Notch reporter mice, we found that Notch signaling was active in HSCs in vivo and downregulated as HSCs differentiated. Inhibition of Notch signaling led to accelerated differentiation of HSCs in vitro and depletion of HSCs in vivo. Finally, intact Notch signaling was required for Wnt-mediated maintenance of undifferentiated HSCs but not for survival or entry into the cell cycle in vitro. These data suggest that Notch signaling has a dominant function in inhibiting differentiation and provide a model for how HSCs may integrate multiple signals to maintain the stem cell state. PMID- 15665829 TI - Uridine adenosine tetraphosphate: a novel endothelium- derived vasoconstrictive factor. AB - Beyond serving as a mechanical barrier, the endothelium has important regulatory functions. The discovery of nitric oxide revolutionized our understanding of vasoregulation. In contrast, the identity of endothelium-derived vasoconstrictive factors (EDCFs) remains unclear. The supernatant obtained from mechanically stimulated human endothelial cells obtained from dermal vessels elicited a vasoconstrictive response in an isolated perfused rat kidney. A purinoceptor blocker had a greater effect than an endothelin receptor blocker in decreasing endothelially derived vasoconstriction in the isolated perfused rat kidney. The nucleotide uridine adenosine tetraphosphate (Up(4)A) was isolated from the supernatant of stimulated human endothelium and identified by mass spectrometry. Up(4)A is likely to exert vasoconstriction predominantly through P2X1 receptors, and probably also through P2Y2 and P2Y4 receptors. Plasma concentrations of Up(4)A that cause vasoconstriction are found in healthy subjects. Stimulation with adenosine 5'-triphosphate (ATP), uridine 5'-triphosphate (UTP), acetylcholine, endothelin, A23187 and mechanical stress releases Up(4)A from endothelium, suggesting that Up(4)A contributes to vascular autoregulation. To our knowledge, Up(4)A is the first dinucleotide isolated from living organisms that contains both purine and pyrimidine moieties. We conclude that Up(4)A is a novel potent nonpeptidic EDCF. Its vasoactive effects, plasma concentrations and its release upon endothelial stimulation strongly suggest that Up(4)A has a functional vasoregulatory role. PMID- 15665830 TI - Re-engineered CD40 receptor enables potent pharmacological activation of dendritic-cell cancer vaccines in vivo. AB - Modest clinical outcomes of dendritic-cell (DC) vaccine trials call for the refinement of DC vaccine design. Although many potential antigens have been identified, development of methods to enhance antigen presentation by DCs has lagged. We have engineered a potent, drug-inducible CD40 (iCD40) receptor that permits temporally controlled, lymphoid-localized, DC-specific activation. iCD40 is comprised of a membrane-localized cytoplasmic domain of CD40 fused to drug binding domains. This allows it to respond to a lipid-permeable, high-affinity dimerizer drug while circumventing ectodomain-dependent negative-feedback mechanisms. These modifications permit prolonged activation of iCD40-expressing DCs in vivo, resulting in more potent CD8(+) T-cell effector responses, including the eradication of previously established solid tumors, relative to activation of DCs ex vivo (P < 0.01), typical of most clinical DC protocols. In addition, iCD40 mediated DC activation exceeded that achieved by stimulating the full-length, endogenous CD40 receptor both in vitro and in vivo. Because iCD40 is insulated from the extracellular environment and can be activated within the context of an immunological synapse, iCD40-expressing DCs have a prolonged lifespan and should lead to more potent vaccines, perhaps even in immune-compromised patients. PMID- 15665831 TI - Cathepsin L is required for endothelial progenitor cell-induced neovascularization. AB - Infusion of endothelial progenitor cells (EPC), but not of mature endothelial cells, promotes neovascularization after ischemia. We performed gene expression profiling of EPC and endothelial cells to identify genes that might be important for the neovascularization capacity of EPC. Notably, the protease cathepsin L (CathL) was highly expressed in EPC as opposed to endothelial cells and was essential for matrix degradation and invasion by EPC in vitro. CathL-deficient mice showed impaired functional recovery following hind limb ischemia, supporting the concept of a crucial role for CathL in postnatal neovascularization. Infused CathL-deficient progenitor cells neither homed to sites of ischemia nor augmented neovascularization. Forced expression of CathL in mature endothelial cells considerably enhanced their invasive activity and sufficed to confer their capacity for neovascularization in vivo. We concluded that CathL has a critical role in the integration of circulating EPC into ischemic tissue and is required for EPC-mediated neovascularization. PMID- 15665832 TI - Angiotensin-converting enzyme is a GPI-anchored protein releasing factor crucial for fertilization. AB - The angiotensin-converting enzyme (ACE) is a key regulator of blood pressure. It is known to cleave small peptides, such as angiotensin I and bradykinin and changes their biological activities, leading to upregulation of blood pressure. Here we describe a new activity for ACE: a glycosylphosphatidylinositol (GPI) anchored protein releasing activity (GPIase activity). Unlike its peptidase activity, GPIase activity is weakly inhibited by the tightly binding ACE inhibitor and not inactivated by substitutions of core amino acid residues for the peptidase activity, suggesting that the active site elements for GPIase differ from those for peptidase activity. ACE shed various GPI-anchored proteins from the cell surface, and the process was accelerated by the lipid raft disruptor filipin. The released products carried portions of the GPI anchor, indicating cleavage within the GPI moiety. Further analysis by high-performance liquid chromatography-mass spectrometry predicted the cleavage site at the mannose-mannose linkage. GPI-anchored proteins such as TESP5 and PH-20 were released from the sperm membrane of wild-type mice but not in Ace knockout sperm in vivo. Moreover, peptidase-inactivated E414D mutant ACE and also PI-PLC rescued the egg-binding deficiency of Ace knockout sperms, implying that ACE plays a crucial role in fertilization through this activity. PMID- 15665833 TI - Experimental autoimmune encephalomyelitis repressed by microglial paralysis. AB - Although microglial activation occurs in inflammatory, degenerative and neoplastic central nervous system (CNS) disorders, its role in pathogenesis is unclear. We studied this question by generating CD11b-HSVTK transgenic mice, which express herpes simplex thymidine kinase in macrophages and microglia. Ganciclovir treatment of organotypic brain slice cultures derived from CD11b HSVTK mice abolished microglial release of nitrite, proinflammatory cytokines and chemokines. Systemic ganciclovir administration to CD11b-HSVTK mice elicited hematopoietic toxicity, which was prevented by transfer of wild-type bone marrow. In bone marrow chimeras, ganciclovir blocked microglial activation in the facial nucleus upon axotomy and repressed the development of experimental autoimmune encephalomyelitis. We conclude that microglial paralysis inhibits the development and maintenance of inflammatory CNS lesions. The microglial compartment thus provides a potential therapeutic target in inflammatory CNS disorders. These results validate CD11b-HSVTK mice as a tool to study the impact of microglial activation on CNS diseases in vivo. PMID- 15665835 TI - Controlling the catalytic bond-breaking selectivity of Ni surfaces by step blocking. AB - The reactivity of catalytic surfaces is often dominated by very reactive low coordinated atoms such as step-edge sites. However, very little knowledge exists concerning the influence of step edges on the selectivity in reactions involving multiple reaction pathways. Such detailed information could be very valuable in rational design of new catalysts with improved selectivity. Here we show, from an interplay between scanning tunnelling microscopy experiments and density functional theory calculations, that the activation of ethylene on Ni(111) follows the trend of higher reactivity for decomposition at step edges as compared with the higher-coordinated terrace sites. The step-edge effect is considerably more pronounced for the C-C bond breaking than for the C-H bond breaking, and thus steps play an important role in the bond-breaking selectivity. Furthermore, we demonstrate how the number of reactive step sites can be controlled by blocking the steps with Ag. This approach to nanoscale design of catalysts is exploited in the synthesis of a new high-surface-area AgNi alloy catalyst, which is tested in hydrogenolysis experiments. PMID- 15665834 TI - Chronic inhibition of cyclic GMP phosphodiesterase 5A prevents and reverses cardiac hypertrophy. AB - Sustained cardiac pressure overload induces hypertrophy and pathological remodeling, frequently leading to heart failure. Genetically engineered hyperstimulation of guanosine 3',5'-cyclic monophosphate (cGMP) synthesis counters this response. Here, we show that blocking the intrinsic catabolism of cGMP with an oral phosphodiesterase-5A (PDE5A) inhibitor (sildenafil) suppresses chamber and myocyte hypertrophy, and improves in vivo heart function in mice exposed to chronic pressure overload induced by transverse aortic constriction. Sildenafil also reverses pre-established hypertrophy induced by pressure load while restoring chamber function to normal. cGMP catabolism by PDE5A increases in pressure-loaded hearts, leading to activation of cGMP-dependent protein kinase with inhibition of PDE5A. PDE5A inhibition deactivates multiple hypertrophy signaling pathways triggered by pressure load (the calcineurin/NFAT, phosphoinositide-3 kinase (PI3K)/Akt, and ERK1/2 signaling pathways). But it does not suppress hypertrophy induced by overexpression of calcineurin in vitro or Akt in vivo, suggesting upstream targeting of these pathways. PDE5A inhibition may provide a new treatment strategy for cardiac hypertrophy and remodeling. PMID- 15665836 TI - Field-effect electroluminescence in silicon nanocrystals. AB - There is currently worldwide interest in developing silicon-based active optical components in order to leverage the infrastructure of silicon microelectronics technology for the fabrication of optoelectronic devices. Light emission in bulk silicon-based devices is constrained in wavelength to infrared emission, and in efficiency by the indirect bandgap of silicon. One promising strategy for overcoming these challenges is to make use of quantum-confined excitonic emission in silicon nanocrystals. A critical challenge for silicon nanocrystal devices based on nanocrystals embedded in silicon dioxide has been the development of a method for efficient electrical carrier injection. We report here a scheme for electrically pumping dense silicon nanocrystal arrays by a field-effect electroluminescence mechanism. In this excitation process, electrons and holes are both injected from the same semiconductor channel across a tunnelling barrier in a sequential programming process, in contrast to simultaneous carrier injection in conventional pn-junction light-emitting-diode structures. Light emission is strongly correlated with the injection of a second carrier into a nanocrystal that has been previously programmed with a charge of the opposite sign. PMID- 15665837 TI - Ferroelectricity near room temperature in co-crystals of nonpolar organic molecules. AB - The research on ferroelectric materials-mostly inorganic compounds or organic polymers-is increasingly motivated by both basic scientific concerns and the potential for practical applications in electronics and optics. Ferroelectricity in organic solids would be important for the development of all-organic electronic and photonic devices. The conventional approach to making organic ferroelectrics is based on the use of polar molecules. Here we report that through supramolecular assembly of nonpolar conjugated molecules, a remarkable ferroelectric response can be obtained in co-crystals of low-molecular-weight organic compounds. Co-crystals of phenazine and chloranilic acid reveal large spontaneous polarization and sizable room-temperature dielectric constants exceeding 100. The present findings provide an approach to making potentially useful organic ferroelectric materials. PMID- 15665838 TI - Novel chain structures in group VI elements. AB - Recent developments in high-pressure methods and advances in X-ray crystallography have led to a new level of understanding of phase diagrams and structures of materials under pressure. Recently discovered phenomena such as complex phases of alkali metals, incommensurate host-guest structures, and incommensurately modulated structures have rendered obsolete our conventional wisdom about the range of structures possible in the elements. Using new in situ diffraction techniques, we have resolved the long-standing problem of the phase transition sequence of sulphur in its non-metallic state. We demonstrate that it is very different from that previously proposed, with only two phases stable between 1.5 GPa and 83 GPa (the pressure of metallization), and temperatures from 300 K to 1,100 K. The phases have a triangular chain and a squared chain structure. The same squared chain structure is found in the heavier group VI element selenium. PMID- 15665839 TI - Impurities block the alpha to omega martensitic transformation in titanium. AB - Impurities control phase stability and phase transformations in natural and man made materials, from shape-memory alloys to steel to planetary cores. Experiments and empirical databases are still central to tuning the impurity effects. What is missing is a broad theoretical underpinning. Consider, for example, the titanium martensitic transformations: diffusionless structural transformations proceeding near the speed of sound. Pure titanium transforms from ductile alpha to brittle omega at 9 GPa, creating serious technological problems for beta-stabilized titanium alloys. Impurities in the titanium alloys A-70 and Ti-6Al-4V (wt%) suppress the transformation up to at least 35 GPa, increasing their technological utility as lightweight materials in aerospace applications. These and other empirical discoveries in technological materials call for broad theoretical understanding. Impurities pose two theoretical challenges: the effect on the relative phase stability, and the energy barrier of the transformation. Ab initio methods calculate both changes due to impurities. We show that interstitial oxygen, nitrogen and carbon retard the transformation whereas substitutional aluminium and vanadium influence the transformation by changing the d-electron concentration. The resulting microscopic picture explains the suppression of the transformation in commercial A-70 and Ti-6Al-4V alloys. In general, the effect of impurities on relative energies and energy barriers is central to understanding structural phase transformations. PMID- 15665840 TI - Value of surveillance blood culture for early diagnosis of occult bacteremia in patients on corticosteroid therapy following allogeneic hematopoietic stem cell transplantation. AB - Bloodstream infection (BSI) is a significant complication following allogeneic hematopoietic stem cell transplantation (allo-SCT). Corticosteroids mask inflammatory responses, delaying the initiation of antibiotics. We reviewed medical records of 69 allo-SCT patients who had been on >0.5 mg/kg prednisolone to investigate the efficacy of weekly surveillance blood cultures. A total of 36 patients (52%) had positive cultures, 25 definitive BSI and 11 probable BSI. Pathogens in definitive BSI were Staphylococcus epidermidis (n=7), S. aureus (n=4), Entrococcus faecalis (n=3), Pseudomonas aeruginosa (n=5), Acenitobacter lwoffii (n=4), and others (n=10). The median interval from the initiation of corticosteroids to the first positive cultures was 24 days (range, 1-70). At the first positive cultures, 15 patients with definitive BSI were afebrile. Four of them remained afebrile throughout the period of positive surveillance cultures. Patients with afebrile BSI tended to be older (P=0.063), and had in-dwelling central venous catheters less frequently than febrile patients (P<0.0001). Bloodstream pathogens were directly responsible for death in two patients with afebrile BSI. This study demonstrates that cortisosteroid frequently masks inflammatory reactions in allo-SCT recipients given conrticosteroids, and that surveillance blood culture is only diagnostic clue for 'occult' BSI. PMID- 15665841 TI - Circulating dendritic cell subset levels after allogeneic stem cell transplantation in children correlate with time post transplant and severity of acute graft-versus-host disease. AB - We examined the recovery of circulating monocytoid (Lin- CD11+ HLA-DR+) and plasmacytoid (Lin- CD123+ HLA-DR+) precursor (pre) dendritic cell (DC) subsets after allogeneic stem cell transplantation (SCT) in 39 children, using age matched healthy children as controls. The frequencies of DCs in peripheral blood samples were determined by flow cytometry. The initial recovery of DC occurred simultaneously with myeloid engraftment. However, with time, DC subset values declined, being very low 40-50 days after SCT. Low monocytoid and plasmacytoid DC values were associated significantly with the development of severe acute graft versus-host disease (aGVHD) (P=0.042 and 0.017, respectively). Plasmacytoid DC values were lower than in the age-matched controls for the entire follow-up period (range 102-2569 days), although, with time, values approached normal levels. Normal monocytoid DC numbers were observed within 300-400 days post SCT. The severity of chronic GVHD did not correlate with quantitative recovery of DC. We conclude that in pediatric SCT, initial recovery of DC production is concurrent with that of myelopoiesis, yet with time, DC subset values decline and low counts are associated with severe aGVHD. Monocytoid DC numbers approach normal levels within a year of SCT, but plasmacytoid DC counts recover very slowly. PMID- 15665842 TI - Predictive factors for hematopoietic engraftment after autologous peripheral blood stem cell transplantation for AL amyloidosis. AB - Treatment of patients with AL amyloidosis with high-dose melphalan and autologous peripheral blood stem cells (PBSC) produces hematologic remissions in approximately 40% of evaluable patients, accompanied by improvements in organ disease and quality of life. These patients, who frequently have amyloid deposits in bone marrow blood vessels and interstitium and impaired function of kidneys, liver, spleen, and heart, represent an unusual population for stem cell transplantation, with unique problems. To identify factors influencing engraftment rates after chemotherapy and autologous granulocyte colony stimulating factor (G-CSF)-mobilized PBSC reinfusion, we studied a group of 225 patients. The median time to neutrophil engraftment was 10 days (range, 8-17 days). In a multivariate analysis, the factors positively affecting the rate of neutrophil engraftment were CD34+ stem cell dose, female gender, and minimal prior alkylator therapy. The median time to platelet engraftment was 13 days (range, 7-52 days). Factors positively affecting platelet engraftment, in addition to CD34+ cell dose, included preserved renal function and the absence of neutropenic fever. The conditioning dose of intravenous melphalan was not found to be an independent predictive factor for hematopoietic recovery. Thus, in this patient population, organ function and host and hematopoietic factors influence engraftment after PBSC rescue. PMID- 15665843 TI - Successful treatment of severe hemorrhagic cystitis with selective vesical artery embolization. PMID- 15665844 TI - In utero transplantation: baby steps towards an effective therapy. AB - In utero transplantation (IUT) offers the potential to treat a large number of diseases by transplantation of healthy cells into a fetus with a birth defect. Prenatal diagnosis is feasible for many diseases prior to the full development of the fetal immune system offering the opportunity to introduce foreign cells and antigens into the developing fetus. At least 45 cases of IUT have been performed for a variety of diseases. IUT has successfully treated severe combined immunodeficiency and there are indications that it may be effective in treating some nonhematopoietic diseases. However, many diseases remain resistant to fetal therapy owing to the low levels of chimerism that can be achieved. Promising efforts to improve the levels of engraftment are focusing on optimizing the graft and developing donor-specific tolerance in the fetal recipient. Mounting evidence suggests that donor T cells can aid in achieving clinically significant levels of chimerism. The use of fetal donor cells may also offer some benefit. Animal experiments suggest that even low-level chimerism can lead to tolerance, which can be exploited by booster transplants in the neonate. Continued research appears likely to succeed in developing IUT into an effective form of therapy for a variety of diseases. PMID- 15665846 TI - CD34+ selected haematopoietic stem cell (HSC) not preceded by any immunosuppressive therapy as effective treatment for graft failure. PMID- 15665847 TI - Medium dose long-wavelength ultraviolet A (UVA1) phototherapy for the treatment of acute and chronic graft-versus-host disease of the skin. AB - Long-wavelength ultraviolet A (340-400 nm UVA1) phototherapy has been reported to be effective in atopic dermatitis, localized scleroderma and other T-cell-derived skin diseases. UVA1 as an adjunct to systemic immunosuppressive treatment was found to be safe, and effective in 10 patients with chronic cutaneous (seven lichenoid and three sclerodermoid) graft-versus-host disease (GVHD) after stem cell transplantation. Complete and partial responses were achieved in six (60%), and in three (30%) patients, respectively. One patient had improvement of sclerotic skin lesions. At a median follow-up of 14 months, two patients with lichenoid lesions relapsed. Both responded to another treatment cycle. Furthermore, we treated seven patients with UVA1 as primary therapy for acute cutaneous GVHD grades II and III in a pilot experience. Five patients had a complete response with no relapse at a median follow-up of 9 months after UVA1. Two patients showed no response and systemic steroids had to be started. UVA1 therapy is feasible, well tolerated and can be effective in treating chronic as well as acute GVHD confined to the skin thereby avoiding systemic steroids. Our results should be confirmed in larger studies and the effectiveness of UVA1 compared to other established treatment modalities. PMID- 15665849 TI - Comparative analysis of a double primer PCR assay with plasma, leukocytes and antigenemia for diagnosis of active human cytomegalovirus infection in bone marrow transplant patients. AB - The aim of the study was to determine the prognostic value of a double primer PCR assay to detect human cytomegalovirus (HCMV) infection or disease in bone marrow transplant (BMT) recipients. A total of 209 blood samples including peripheral blood mononuclear cells (PBMN), polymorphonuclear (PMN) leukocytes and plasma from 26 BMT recipients were tested by PCR assay. To discriminate between latent and active HCMV infection, 177 blood samples were also tested by a quantitative antigenemia assay. HCMV serology status of donors and recipients was determined before transplantation by an enzyme immunosorbent assay method. Using the double primer PCR assay, the number of positive samples increased by an average of 11.6%. Symptomatic active HCMV infection was diagnosed in 14 (53.8%) out of 26 BMT patients. There was a good association between double primer PCR assay of PMN leukocytes and antigenemia assays for detection of active HCMV infection in all patients. Detection of HCMV DNA in PMN leukocytes of BMT patients by double primer PCR assay can be an alternative method for antigenemia assay. However, quantitative PCR methods will be necessary for monitoring antiviral treatment. PMID- 15665848 TI - Marrow transplants from matched unrelated donors for aplastic anaemia using alemtuzumab, fludarabine and cyclophosphamide based conditioning. AB - Graft failure, regimen-related toxicity and graft-versus-host disease (GVHD) are the critical barriers to unrelated donor transplants for aplastic anaemia (AA). We investigated the use of a novel conditioning regimen consisting of alemtuzumab (humanized CD52 antibody), fludarabine and cyclophosphamide in seven patients with AA, who underwent bone marrow transplant procedure using matched unrelated donors. The aetiology of AA was acquired (n=3), Fanconi's (n=3) and congenital (n=1). Median age was 13 years (range 8-35). All the donors were fully matched for HLA class I and II antigens using high-resolution typing. All the patients engrafted at a median of 18 days (range 13-35). Two patients died of transplant related complications: one of adenovirus disease and the other developed extensive chronic GVHD of skin followed by cytomegalovirus (CMV) disease. Three patients developed Grade II acute GVHD disease (GVHD); none had Grade III-IV acute GVHD. Of the six evaluable patients, only one developed chronic GVHD. We conclude that this conditioning regimen for unrelated donor transplants for AA is sufficiently immunosuppressive to allow stable engraftment and appears to have a favourable impact on the incidence and severity of GVHD, warranting further investigation. PMID- 15665850 TI - Non-radiotherapy conditioning with stem cell transplantation from alternative donors in children with refractory severe aplastic anemia. AB - Conditioning including total body/lymphoid irradiation is widely used to prevent graft rejection in patients with refractory severe aplastic anemia (SAA) undergoing hemopoietic cell transplantation (HCT) from alternative donors and or after graft manipulation. To reduce regimen-related toxicity we transplanted three children with refractory SAA after conditioning with radiotherapy-free regimens. Conditioning included fludarabine 175-180 mg/m2 in all patients. In addition, patient 1 (failing two previous grafts) received thiotepa 10 mg/kg and Campath-1H 60 mg/m2; patient 2 cyclophosphamide 120 mg/kg, thiotepa 15 mg/kg and OKT-3 0.1 mg/kg/day for 4 weeks; and patient 3 cyclophosphamide 120 and ATG 90 mg/kg. Stem cell source was unmanipulated marrow from the same unrelated donor as for the two previous transplantations in patient 1 and CD34+-purified peripheral blood stem cells from an HLA-matched unrelated donor and from the haploidentical mother in patients 2 and 3. Only patient 1 received graft-versus-host disease (GVHD) prophylaxis with cyclosporine A and mycophenolate mofetil. Follow-up is now 30, 51, and 15 months. None of the patients developed GVHD. All patients have normal counts with complete donor chimerism. Fludarabine-based conditioning is powerfully immunosuppressive and may be used for children with refractory SAA undergoing HCT from alternative donors even after rejection following previous HCT. PMID- 15665851 TI - EBV transformation of cells from cord blood donations: relevance to future retrospective studies on cord blood transplants. PMID- 15665852 TI - A prognostic model for prolonged event-free survival after autologous or allogeneic blood or marrow transplantation for relapsed and refractory Hodgkin's disease. AB - There are several prognostic models for Hodgkin's disease (HD) patients, but none evaluating patient characteristics at time of blood and marrow transplantation (BMT). We developed a prognostic model for event-free survival (EFS) post-BMT based on HD patient characteristics measured at the time of autologous (auto) or allogeneic (allo) BMT. Between 1/1991 and 12/2001, 64 relapsed or refractory HD patients received an auto (n=46) or allo (n=18) BMT. A multivariate prognostic model was developed measuring time to relapse, progression or death. Median follow-up was 51.7 months; median EFS for auto and allo BMT was 36 and 3 months, respectively (P=0.001). Significant multivariate predictors of shorter EFS were chemotherapy-resistant disease, KPS <90 and > or =3 chemotherapy regimens pre BMT. Patients with two to three adverse factors had significantly shorter EFS at 2 years (58 vs 11% in auto; 38 vs 0% in allo BMT patients). Despite a selection bias favoring auto BMT, the model was valid in both auto and allo BMT groups. We were able to differentiate patients at high vs low risk for adverse outcomes post BMT. This prognostic model may prove useful in predicting patient outcomes and identifying high-risk patients for novel treatment strategies. Validation of this model in a larger cohort of patients is warranted. PMID- 15665853 TI - SAS-6 defines a protein family required for centrosome duplication in C. elegans and in human cells. AB - The mechanisms that ensure centrosome duplication are poorly understood. In Caenorhabditis elegans, ZYG-1, SAS-4, SAS-5 and SPD-2 are required for centriole formation. However, it is unclear whether these proteins have functional homologues in other organisms. Here, we identify SAS-6 as a component that is required for daughter centriole formation in C. elegans. SAS-6 is a coiled-coil protein that is recruited to centrioles at the onset of the centrosome duplication cycle. Our analysis indicates that SAS-6 and SAS-5 associate and that this interaction, as well as ZYG-1 function, is required for SAS-6 centriolar recruitment. SAS-6 is the founding member of an evolutionarily conserved protein family that contains the novel PISA motif. We investigated the function of the human homologue of SAS-6. GFP-HsSAS-6 localizes to centrosomes and its overexpression results in excess foci-bearing centriolar markers. Furthermore, siRNA-mediated inactivation of HsSAS-6 in U2OS cells abrogates centrosome overduplication following aphidicolin treatment and interferes with the normal centrosome duplication cycle. Therefore, HsSAS-6 is also required for centrosome duplication, indicating that the function of SAS-6-related proteins has been widely conserved during evolution. PMID- 15665854 TI - TRPC1 forms the stretch-activated cation channel in vertebrate cells. AB - The mechanosensitive cation channel (MscCa) transduces membrane stretch into cation (Na(+), K(+), Ca(2+) and Mg(2+)) flux across the cell membrane, and is implicated in cell-volume regulation, cell locomotion, muscle dystrophy and cardiac arrhythmias. However, the membrane protein(s) that form the MscCa in vertebrates remain unknown. Here, we use an identification strategy that is based on detergent solubilization of frog oocyte membrane proteins, followed by liposome reconstitution and evaluation by patch-clamp. The oocyte was chosen because it expresses the prototypical MscCa (>or=10(7)MscCa/oocyte) that is preserved in cytoskeleton-deficient membrane vesicles. We identified a membrane protein fraction that reconstituted high MscCa activity and showed an abundance of a protein that had a relative molecular mass of 80,000 (M(r) 80K). This protein was identified, by immunological techniques, as the canonical transient receptor potential channel 1 (TRPC1). Heterologous expression of the human TRPC1 resulted in a >1,000% increase in MscCa patch density, whereas injection of a TRPC1-specific antisense RNA abolished endogenous MscCa activity. Transfection of human TRPC1 into CHO-K1 cells also significantly increased MscCa expression. These observations indicate that TRPC1 is a component of the vertebrate MscCa, which is gated by tension developed in the lipid bilayer, as is the case in various prokaryotic mechanosensitive (Ms) channels. PMID- 15665855 TI - OASIS, a CREB/ATF-family member, modulates UPR signalling in astrocytes. AB - Endoplasmic reticulum (ER) stress transducers IRE1, PERK and ATF6 are well known to transduce signals from the ER to the cytoplasm and nucleus when unfolded proteins are accumulated in the ER. Here, we identified OASIS as a novel ER stress transducer. OASIS is a basic leucine zipper (bZIP) transcription factor of the CREB/ATF family with a transmembrane domain that allows it to associate with the ER. The molecule is cleaved at the membrane in response to ER stress, and its cleaved amino-terminal cytoplasmic domain, which contains the bZIP domain, translocates into the nucleus where it activates the transcription of target genes that are mediated by ER stress-responsive and cyclic AMP-responsive elements. Intriguingly, OASIS was induced at the transcriptional level during ER stress in astrocytes of the central nervous system, but not in other cell types examined. Furthermore, overexpression of OASIS resulted in induction of BiP and suppression of ER-stress-induced cell death, whereas knockdown partially reduced BiP levels and led to ER stress in susceptible astrocytes. Our results reveal pivotal roles for OASIS in modulating the unfolded protein response in astrocytes, and the possibility that cell type-specific UPR signalling also exists in other cells. PMID- 15665856 TI - The cell-cycle checkpoint kinase Chk1 is required for mammalian homologous recombination repair. AB - The essential checkpoint kinase Chk1 is required for cell-cycle delays after DNA damage or blocked DNA replication. However, it is unclear whether Chk1 is involved in the repair of damaged DNA. Here we establish that Chk1 is a key regulator of genome maintenance by the homologous recombination repair (HRR) system. Abrogation of Chk1 function with small interfering RNA or chemical antagonists inhibits HRR, leading to persistent unrepaired DNA double-strand breaks (DSBs) and cell death after replication inhibition with hydroxyurea or DNA damage caused by camptothecin. After hydroxyurea treatment, the essential recombination repair protein RAD51 is recruited to DNA repair foci performing a vital role in correct HRR. We demonstrate that Chk1 interacts with RAD51, and that RAD51 is phosphorylated on Thr 309 in a Chk1-dependent manner. Consistent with a functional interplay between Chk1 and RAD51, Chk1-depleted cells failed to form RAD51 nuclear foci after exposure to hydroxyurea, and cells expressing a phosphorylation-deficient mutant RAD51(T309A) were hypersensitive to hydroxyurea. These results highlight a crucial role for the Chk1 signalling pathway in protecting cells against lethal DNA lesions through regulation of HRR. PMID- 15665857 TI - The histamine H3 receptor: from gene cloning to H3 receptor drugs. AB - Since the cloning of the histamine H(3) receptor cDNA in 1999 by Lovenberg and co workers, this histamine receptor has gained the interest of many pharmaceutical companies as a potential drug target for the treatment of various important disorders, including obesity, attention-deficit hyperactivity disorder, Alzheimer's disease, schizophrenia, as well as for myocardial ischaemia, migraine and inflammatory diseases. Here, we discuss relevant information on this target protein and describe the development of various H(3) receptor agonists and antagonists, and their effects in preclinical animal models. PMID- 15665858 TI - Metabotropic glutamate receptors as novel targets for anxiety and stress disorders. AB - Anxiety and stress disorders are the most commonly occurring of all mental illnesses, and current treatments are less than satisfactory. So, the discovery of novel approaches to treat anxiety disorders remains an important area of neuroscience research. Glutamate is the major excitatory neurotransmitter in the mammalian central nervous system, and G-protein-coupled metabotropic glutamate (mGlu) receptors function to regulate excitability via pre- and postsynaptic mechanisms. Various mGlu receptor subtypes, including group I (mGlu(1) and mGlu(5)), group II (mGlu(2) and mGlu(3)), and group III (mGlu(4), mGlu(7) and mGlu(8)) receptors, specifically modulate excitability within crucial brain structures involved in anxiety states. In addition, agonists for group II (mGlu(2/3)) receptors and antagonists for group I (in particular mGlu(5)) receptors have shown activity in animal and/or human conditions of fear, anxiety or stress. These studies indicate that metabotropic glutamate receptors are interesting new targets to treat anxiety disorders in humans. PMID- 15665859 TI - Erythromycin exerts in vivo anti-inflammatory activity downregulating cell adhesion molecule expression. AB - 1. Macrolides have long been used as anti-bacterial agents; however, there is some evidence that may exert anti-inflammatory activity. Therefore, erythromycin was used to characterize the mechanisms involved in their in vivo anti inflammatory activity. 2. Erythromycin pretreatment (30 mg kg(-1) day(-1) for 1 week) reduced the lipopolysaccharide (LPS; intratracheal, 0.4 mg kg(-1))-induced increase in neutrophil count and elastase activity in the bronchoalveolar lavage fluid (BALF) and lung tissue myeloperoxidase activity, but failed to decrease tumor necrosis factor-alpha and macrophage-inflammatory protein-2 augmented levels in BALF. Erythromycin pretreatment also prevented lung P-selectin, E selectin, intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) mRNA upregulation in response to airway challenge with LPS. 3. Mesentery superfusion with LPS (1 mug ml(-1)) induced a significant increase in leukocyte-endothelial cell interactions at 60 min. Erythromycin pretreatment abolished the increases in these parameters. 4. LPS exposure of the mesentery for 4 h caused a significant increase in leukocyte rolling flux, adhesion and emigration, which were inhibited by erythromycin by 100, 93 and 95%, respectively. 5. Immunohistochemical analysis showed that LPS exposure of the mesentery for 4 h caused a significant enhancement in P-selectin, E-selectin, ICAM-1 and VCAM-1 expression that was downregulated by erythromycin pretreatment. 6. Flow cytometry analysis indicated that erythromycin pretreatment inhibited LPS induced CD11b augmented expression in rat neutrophils. 7. In conclusion, erythromycin inhibits leukocyte recruitment in the lung and this effect appears mediated through downregulation of CAM expression. Therefore, macrolides may be useful in the control of neutrophilic pulmonary diseases. PMID- 15665860 TI - Imidapril treatment improves the attenuated inotropic and intracellular calcium responses to ATP in heart failure due to myocardial infarction. AB - 1. Adenosine 5'-triphosphate (ATP) is known to augment cardiac contractile activity and cause an increase in intracellular Ca(2+) concentration ([Ca(2+)](i)) in isolated cardiomyocytes. However, no information regarding the ATP-mediated signal transduction in the myocardium in congestive heart failure (CHF) is available. 2. CHF due to myocardial infarction (MI) in rats was induced by the occlusion of the left coronary artery for 8 weeks. The positive inotropy due to ATP was depressed in failing hearts. Treatment of 3 weeks infarcted animals with imidapril (1 mg kg(-1) day(-1)) for a period of 5 weeks improved the left ventricle function and decreased the attenuation of inotropic response to ATP. 3. ATP-induced increase in [Ca(2+)](i) was significantly depressed in cardiomyocytes isolated from the failing heart and this change was partially attenuated by imidapril treatment. However, the binding characteristics of (35)S labeled adenosine 5'-(gamma-thio) triphosphate in sarcolemma isolated from the failing heart remained unaltered. 4. ATP-induced increase in [Ca(2+)](i) was depressed by verapamil and cibacron blue in both control and failing heart cardiomyocytes; however, the ATP response in the failing hearts, unlike the control preparations, was not decreased by ryanodine. This insensitivity to ryanodine was attenuated by imidapril treatment. 5. Treatment of infarcted rats with enalapril and losartan produced effects similar to imidapril. 6. These findings indicate that the positive inotropic response to ATP and ATP-induced increase in [Ca(2+)](i) in cardiomyocytes are impaired in heart failure. Furthermore, blockade of renin angiotensin system prevented the impairment of the ATP-mediated inotropic and [Ca(2+)](i) responses in the failing heart. PMID- 15665861 TI - TNF alpha reduces tachykinin, PGE2-dependent, relaxation of the cultured mouse trachea by increasing the activity of COX-2. AB - 1. Chronic inflammation is a central feature of asthma. The inflammatory cytokine tumour necrosis factor alpha (TNFalpha) has been implicated in this disease, and is known to alter airway smooth muscle functionally. 2. The aim of this study was to investigate the influence of TNFalpha on tachykinin-induced airway relaxation. Mouse tracheae were cultured in the absence and presence of TNFalpha for 1 or 4 days. 3. In the absence of TNFalpha, substance P (SP) and neurokinin A (NKA) induced comparable levels of relaxation in fresh and cultured segments. Functional studies with selective antagonists/inhibitors indicated that the relaxation was mediated by the NK(1) receptor coupled to cyclooxygenase (COX)-2 activation and subsequent release of prostaglandin E(2) (PGE(2)). TNFalpha attenuated SP- and NKA-induced relaxation in a time- and concentration-dependent manner, decreasing the ability of PGE(2) to relax tissues. 4. Further studies indicated that TNFalpha elevated COX-2 activity and that concomitant inhibition of COX-2 reversed TNFalpha-attenuated PGE(2) relaxation. Culture with PGE(2) decreased SP- and PGE(2)-mediated relaxation, further implicating the activity of COX-2 in the attenuation of tachykinin signalling. 5. Gene expression analysis demonstrated that TNFalpha increased the expression of smooth muscle COX-2, PGE(2) synthase and EP(2) receptor mRNA, and decreased the expression of the EP(4) receptor. 6. Overall, these results show that NK(1) receptor-mediated relaxation induced by PGE(2) is attenuated by prolonged TNFalpha stimulation. Increased COX-2 activity induced by TNFalpha appears to be central to this process. PMID- 15665862 TI - A comparative study on the acute and long-term effects of MDMA and 3,4 dihydroxymethamphetamine (HHMA) on brain monoamine levels after i.p. or striatal administration in mice. AB - 1. This study investigated whether the immediate and long-term effects of 3,4 methylenedioxymethamphetamine (MDMA) on monoamines in mouse brain are due to the parent compound and the possible contribution of a major reactive metabolite, 3,4 dihydroxymethamphetamine (HHMA), to these changes. The acute effect of each compound on rectal temperature was also determined. 2. MDMA given i.p. (30 mg kg( 1), three times at 3-h intervals), but not into the striatum (1, 10 and 100 microg, three times at 3-h intervals), produced a reduction in striatal dopamine content and modest 5-HT reduction 1 h after the last dose. MDMA does not therefore appear to be responsible for the acute monoamine release that follows its peripheral injection. 3. HHMA does not contribute to the acute MDMA-induced dopamine depletion as the acute central effects of MDMA and HHMA differed following i.p. injection. Both compounds induced hyperthermia, confirming that the acute dopamine depletion is not responsible for the temperature changes. 4. Peripheral administration of MDMA produced dopamine depletion 7 days later. Intrastriatal MDMA administration only produced a long-term loss of dopamine at much higher concentrations than those reached after the i.p. dose and therefore bears little relevance to the neurotoxicity. This indicates that the long-term effect is not attributable to the parent compound. HHMA also appeared not to be responsible as i.p. administration failed to alter the striatal dopamine concentration 7 days later. 5. HHMA was detected in plasma, but not in brain, following MDMA (i.p.), but it can cross the blood-brain barrier as it was detected in the brain following its peripheral injection. 6. The fact that the acute changes induced by i.p. or intrastriatal HHMA administration differed indicates that HHMA is metabolised to other compounds which are responsible for changes observed after i.p. administration. PMID- 15665863 TI - Deoxycholic acid inhibits pacemaker currents by activating ATP-dependent K+ channels through prostaglandin E2 in interstitial cells of Cajal from the murine small intestine. AB - 1. We investigated the role of deoxycholic acid in pacemaker currents using whole cell patch-clamp techniques at 30 degrees C in cultured interstitial cells of Cajal (ICC) from murine small intestine. 2. The treatment of ICC with deoxycholic acid resulted in a decrease in the frequency and amplitude of pacemaker currents and increases in resting outward currents. Also, under current clamping, deoxycholic acid produced the hyperpolarization of membrane potential and decreased the amplitude of the pacemaker potentials. 3. These observed effects of deoxycholic acid on pacemaker currents and pacemaker potentials were completely suppressed by glibenclamide, an ATP-sensitive K(+) channel blocker. 4. NS-398, a specific cyclooxygenase-2 (COX-2) inhibitor, significantly inhibited the deoxycholic acid-induced effects. The treatment with prostaglandin E(2) (PGE(2)) led to a decrease in the amplitude and frequency of pacemaker currents and to an increase in resting outward currents, and these observed effects of PGE(2) were blocked by glibenclamide. 5. We next examined the role of deoxycholic acid in the production of PGE(2) in ICC, and found that deoxycholic acid increased PGE(2) production through the induction of COX-2 enzyme activity and its gene expression. 6. The results suggest that deoxycholic acid inhibits the pacemaker currents of ICC by activating ATP-sensitive K(+) channels through the production of PGE(2). PMID- 15665864 TI - Effects of minocycline on Fas-mediated fulminant hepatitis in mice. AB - 1. Minocycline has anti-inflammatory and antiapoptotic effects on cartilage, neurons and periodontal tissues, and both properties are central to the pharmaceutical treatment of liver diseases. We investigated the effects of minocycline on fulminant hepatitis in C57BL/6J mice induced by lethal challenge of the activating anti-Fas antibody, Jo2. 2. Intraperitoneal injection of Jo2 (0.6 microg g(-1)) to mice resulted in fulminant hepatitis, as evidenced by increase of serum alanine/aspartate transaminase activities and histopathological alterations in liver sections, as well as animal death. Nevertheless, mice pretreated with three doses of minocycline (5 mg kg(-1)) resisted this lethal effect significantly. Minocycline treatment improved the survival kinetics, although to a lesser extent, when mice were challenged simultaneously with Jo2 or even treated 30 min after the lethal challenge. 3. Jo2-induced activation of caspase-3 or -9 in liver tissues was inhibited by minocycline pretreatment, and yet the direct addition of minocycline to liver extracts from Jo2-challenged mice failed to block caspase activation in vitro. Moreover, minocycline efficiently suppressed the release of cytochrome c from mitochondria of the liver tissues from Jo2-challenged mice. In contrast, caspase-8 activation and Bid truncation triggered by Jo2 were not diminished by minocycline pretreatment in mouse livers. 4. Our results suggest that easing of Fas-triggered fulminant hepatitis by minocycline may involve a mitochondrial apoptotic pathway, probably through preventing cytochrome c release and thereby blocking downstream caspase activation. PMID- 15665865 TI - Extracellular protons differentially potentiate the responses of native AMPA receptor subtypes regulating neurotransmitter release. AB - 1. The effects of pH changes on the basal and evoked release of [(3)H]noradrenaline ([(3)H]NA) and [(3)H]5-hydrohytryptamine ([(3)H]5-HT) from hippocampal synaptosomes and of [(3)H]dopamine ([(3)H]DA) and [(3)H]acetylcholine ([(3)H]ACh) from striatal and cortical synaptosomes were investigated in rat brain. 2. Changing pH between 6.4 and 8.0 did not affect the spontaneous release of the four [(3)H]neurotransmitters; alkalinization to pH 8.8 significantly enhanced release. Acidification to pH 6.4 augmented the AMPA-evoked overflows of [(3)H]NA, [(3)H]5-HT and [(3)H]DA, but not that of [(3)H]ACh. In contrast, lowering pH to 6.4 decreased the K(+)-evoked overflows of [(3)H]NA, [(3)H]5-HT, [(3)H]DA and [(3)H]ACh. 3. AMPA released transmitters in a Ca(2+)-dependent, exocytotic manner since its effects, at pH 7.4 or 6.4, were abolished by omitting external Ca(2+) or by depleting vesicular transmitter stores with bafilomycin A1. AMPA did not evoke carrier-mediated release because the uptake blockers nisoxetine, 6-nitroquipazine, GBR12909 and hemicholinium-3 could not inhibit the AMPA-induced release of [(3)H]NA, [(3)H]5-HT, [(3)H]DA and [(3)H]ACh. 4. Extraterminal acidification to pH 6.4 prevented the potentiating effect of cyclothiazide on the AMPA-evoked release of [(3)H]NA, [(3)H]DA and [(3)H]5-HT, whereas the proton-insensitive AMPA-evoked release of [(3)H]ACh, previously found to be cyclothiazide-insensitive at pH 7.4 was cyclothiazide-resistant also at pH 6.4. 5. To conclude, the cyclothiazide-sensitive AMPA receptors mediating release of NA, 5-HT and DA, but not the cyclothiazide-insensitive AMPA receptors mediating the release of ACh, become more responsive when external pH is lowered to pathophysiologically relevant values. The results with cyclothiazide suggest that H(+) ions may prevent desensitization of some AMPA receptor subtypes. PMID- 15665866 TI - Pivotal role of the glycine-rich TM3 helix in gating the MscS mechanosensitive channel. AB - The crystal structure of an open form of the Escherichia coli MscS mechanosensitive channel was recently solved. However, the conformation of the closed state and the gating transition remain uncharacterized. The pore-lining transmembrane helix contains a conserved glycine- and alanine-rich motif that forms a helix-helix interface. We show that introducing 'knobs' on the smooth glycine face by replacing glycine with alanine, and substituting conserved alanines with larger residues, increases the pressure required for gating. Creation of a glycine-glycine interface lowers activation pressure. The importance of residues Gly104, Ala106 and Gly108, which flank the hydrophobic seal, is demonstrated. A new structural model is proposed for the closed-to-open transition that involves rotation and tilt of the pore-lining helices. Introduction of glycine at Ala106 validated this model by acting as a powerful suppressor of defects seen with mutations at Gly104 and Gly108. PMID- 15665867 TI - Motility of myosin V regulated by the dissociation of single calmodulin. AB - Myosin V is a calmodulin-binding motor protein. The dissociation of single calmodulin molecules from individual myosin V molecules at 1 microM Ca(2+) correlates with a reduction in sliding velocity in an in vitro motility assay. The dissociation of two calmodulin molecules at 5 microM Ca(2+) correlates with a detachment of actin filaments from myosin V. To mimic the regulation of myosin V motility by Ca(2+) in a cell, caged Ca(2+) coupled with a UV flash system was used to produce Ca(2+) transients. During the Ca(2+) transient, myosin V goes through the functional cycle of reduced sliding velocity, actin detachment and reattachment followed by the recovery of the sliding velocity. These results indicate that myosin V motility is regulated by Ca(2+) through a reduction in actin-binding affinity resulting from the dissociation of single calmodulin molecules. PMID- 15665868 TI - Dissection of COPII subunit-cargo assembly and disassembly kinetics during Sar1p GTP hydrolysis. AB - COPII coat proteins are required for direct capture of cargo and SNARE proteins into transport vesicles from the endoplasmic reticulum (ER). Cargo and SNARE capture occurs during the formation of a 'prebudding complex' comprising a cargo, Sar1p-GTP and the COPII subunits Sec23/24p. The assembly and disassembly cycle of the prebudding complex on ER membranes is coupled to the Sar1p GTPase cycle. Using FRET to monitor a single round of Sec23/24p binding and dissociation from SNAREs in reconstituted liposomes, we show that Sec23/24p dissociates from v SNARE and complexed t-SNARE with kinetics slower than Sar1p-GTP hydrolysis. Once Sec23/24p becomes associated with v-SNARE or complexed t-SNARE, the complex remains assembled during multiple rounds of Sar1p-GTP hydrolysis mediated by the GDP-GTP exchange factor Sec12p. These data suggest a model for the maintenance of kinetically stable prebudding complexes during the Sar1p GTPase cycle that regulates cargo sorting into transport vesicles. PMID- 15665869 TI - The snake venom protein botrocetin acts as a biological brace to promote dysfunctional platelet aggregation. AB - Botrocetin is a snake venom protein that enhances the affinity of the A1 domain of plasma von Willebrand factor (vWF) for the platelet receptor glycoprotein Ibalpha (GPIbalpha), an event that contributes to bleeding and host death. Here we describe a kinetic and crystallographic analysis of this interaction that reveals a novel mechanism of affinity enhancement. Using high-temporal-resolution microscopy, we show that botrocetin decreases the GPIbalpha off-rate two-fold in both human and mouse complexes without affecting the on-rate. The key to this behavior is that, upon binding of GPIbalpha to vWF-A1, botrocetin prebound to vWF A1 makes no contacts initially with GPIbalpha, but subsequently slides around the A1 surface to form a new interface. This two-step mechanism and flexible coupling may prevent adverse alterations in on-rate of GPIbalpha for vWF-A1, and permit adaptation to structural differences in GPIbalpha and vWF in several prey species. PMID- 15665870 TI - Structural basis for mRNA recognition by elongation factor SelB. AB - In bacteria, incorporation of selenocysteine, the 21(st) amino acid, into proteins requires elongation factor SelB, which has the unusual property of binding to both transfer RNA (tRNA) and mRNA. SelB binds to an mRNA hairpin formed by the selenocysteine insertion sequence (SECIS) with extremely high specificity, the molecular basis of which has been unknown. We have determined the crystal structure of the mRNA-binding domain of SelB in complex with SECIS RNA at a resolution of 2.3 A. This is the first example of a complex between an RNA and a winged-helix (WH) domain, a motif found in many DNA-binding proteins and recently discovered in RNA-binding proteins. Notably, RNA binding does not induce a major conformational change in the WH motif. The structure reveals a new mode of RNA recognition with a geometry that allows the complex to wrap around the small ribosomal subunit. PMID- 15665871 TI - Mapping the interaction of DNA with the Escherichia coli DNA polymerase clamp loader complex. AB - Sliding clamps are loaded onto DNA by ATP-dependent clamp loader complexes. A recent crystal structure of a clamp loader-clamp complex suggested an unexpected mechanism for DNA recognition, in which the ATPase subunits of the loader spiral around primed DNA. We report the results of fluorescence-based assays that probe the mechanism of the Escherichia coli clamp loader and show that conserved residues clustered within the inner surface of the modeled clamp loader spiral are critical for DNA recognition, DNA-dependent ATPase activity and clamp release. Duplex DNA with a 5'-overhang single-stranded region (corresponding to correctly primed DNA) stimulates clamp release, as does blunt-ended duplex DNA, whereas duplex DNA with a 3' overhang and single-stranded DNA are ineffective. These results provide evidence for the recognition of DNA within an inner chamber formed by the spiral organization of the ATPase domains of the clamp loader. PMID- 15665872 TI - Structure of the p115RhoGEF rgRGS domain-Galpha13/i1 chimera complex suggests convergent evolution of a GTPase activator. AB - p115RhoGEF, a guanine nucleotide exchange factor (GEF) for Rho GTPase, is also a GTPase-activating protein (GAP) for G12 and G13 heterotrimeric Galpha subunits. The GAP function of p115RhoGEF resides within the N-terminal region of p115RhoGEF (the rgRGS domain), which includes a module that is structurally similar to RGS (regulators of G-protein signaling) domains. We present here the crystal structure of the rgRGS domain of p115RhoGEF in complex with a chimera of Galpha13 and Galphai1. Two distinct surfaces of rgRGS interact with Galpha. The N-terminal betaN-alphaN hairpin of rgRGS, rather than its RGS module, forms intimate contacts with the catalytic site of Galpha. The interface between the RGS module of rgRGS and Galpha is similar to that of a Galpha-effector complex, suggesting a role for the rgRGS domain in the stimulation of the GEF activity of p115RhoGEF by Galpha13. PMID- 15665873 TI - Key features of the interaction between Pcf11 CID and RNA polymerase II CTD. AB - The C-terminal domain (CTD) of the large subunit of RNA polymerase II is a platform for mRNA processing factors and links gene transcription to mRNA capping, splicing and polyadenylation. Pcf11, an essential component of the mRNA cleavage factor IA, contains a CTD-interaction domain that binds in a phospho dependent manner to the heptad repeats within the RNA polymerase II CTD. We show here that the phosphorylated CTD exists as a dynamic disordered ensemble in solution and, by induced fit, it assumes a structured conformation when bound to Pcf11. In addition, we detected cis-trans populations for the CTD prolines, and found that only the all-trans form is selected for binding. These data suggest that the recognition of the CTD is regulated by independent site-specific modifications (phosphorylation and proline cis-trans isomerization) and, probably, by the local concentration of suitable binding sites. PMID- 15665874 TI - Prefrontal white matter volume is disproportionately larger in humans than in other primates. AB - Determining how the human brain differs from nonhuman primate brains is central to understanding human behavioral evolution. There is currently dispute over whether the prefrontal cortex, which mediates evolutionarily interesting behaviors, has increased disproportionately. Using magnetic resonance imaging brain scans from 11 primate species, we measured gray, white and total volumes for both prefrontal and the entire cerebrum on each specimen (n = 46). In relative terms, prefrontal white matter shows the largest difference between human and nonhuman, whereas gray matter shows no significant difference. This suggests that connectional elaboration (as gauged by white matter volume) played a key role in human brain evolution. PMID- 15665875 TI - Bistability of cerebellar Purkinje cells modulated by sensory stimulation. AB - A persistent change in neuronal activity after brief stimuli is a common feature of many neuronal microcircuits. This persistent activity can be sustained by ongoing reverberant network activity or by the intrinsic biophysical properties of individual cells. Here we demonstrate that rat and guinea pig cerebellar Purkinje cells in vivo show bistability of membrane potential and spike output on the time scale of seconds. The transition between membrane potential states can be bidirectionally triggered by the same brief current pulses. We also show that sensory activation of the climbing fiber input can switch Purkinje cells between the two states. The intrinsic nature of Purkinje cell bistability and its control by sensory input can be explained by a simple biophysical model. Purkinje cell bistability may have a key role in the short-term processing and storage of sensory information in the cerebellar cortex. PMID- 15665876 TI - Invariant computations in local cortical networks with balanced excitation and inhibition. AB - Cortical computations critically involve local neuronal circuits. The computations are often invariant across a cortical area yet are carried out by networks that can vary widely within an area according to its functional architecture. Here we demonstrate a mechanism by which orientation selectivity is computed invariantly in cat primary visual cortex across an orientation preference map that provides a wide diversity of local circuits. Visually evoked excitatory and inhibitory synaptic conductances are balanced exquisitely in cortical neurons and thus keep the spike response sharply tuned at all map locations. This functional balance derives from spatially isotropic local connectivity of both excitatory and inhibitory cells. Modeling results demonstrate that such covariation is a signature of recurrent rather than purely feed-forward processing and that the observed isotropic local circuit is sufficient to generate invariant spike tuning. PMID- 15665877 TI - The site of action potential initiation in cerebellar Purkinje neurons. AB - Knowledge of the site of action potential initiation is essential for understanding how synaptic input is converted into neuronal output. Previous studies have shown that the lowest-threshold site for initiation of action potentials is in the axon. Here we use recordings from visualized rat cerebellar Purkinje cell axons to localize the site of initiation to a well-defined anatomical structure: the first node of Ranvier, which normally forms at the first axonal branch point. PMID- 15665878 TI - Using visual direction in three-dimensional motion perception. AB - The eyes receive slightly different views of the world, and the differences between their images (binocular disparity) are used to see depth. Several authors have suggested how the brain could exploit this information for three-dimensional (3D) motion perception, but here we consider a simpler strategy. Visual direction is the angle between the direction of an object and the direction that an observer faces. Here we describe human behavioral experiments in which observers use visual direction, rather than binocular information, to estimate an object's 3D motion even though this causes them to make systematic errors. This suggests that recent models of binocular 3D motion perception may not reflect the strategies that human observers actually use. PMID- 15665879 TI - Cyclic AMP controls BDNF-induced TrkB phosphorylation and dendritic spine formation in mature hippocampal neurons. AB - Synaptic actions of brain-derived neurotrophic factor (BDNF) are 'gated' by cyclic AMP (cAMP), but the underlying molecular mechanisms remain unclear. Here we report that cAMP regulates BDNF function in mature hippocampal neurons by modulating the signaling and trafficking of its receptor TrkB. cAMP gated the TrkB tyrosine kinase with three characteristic features: BDNF-induced TrkB phosphorylation was attenuated by inhibitors of cAMP signaling, it was potentiated by cAMP analogs, and activation of the cAMP pathway alone had no effect. In addition, cAMP facilitated trafficking of TrkB to dendritic spines, possibly by promoting its interaction with synaptic scaffolding protein PSD-95. Norepinephrinergic and dopaminergic agonists, which elevate intracellular cAMP concentration, also enhanced TrkB phosphorylation and its translocation to spines. cAMP gated long-term modulation by BDNF of spine density, but not the number of primary dendrites. These results reveal a specific role of cAMP in controlling BDNF actions in the brain, and provide new insights into the molecular mechanism underlying cAMP gating. PMID- 15665880 TI - The voices of wrath: brain responses to angry prosody in meaningless speech. AB - We report two functional magnetic resonance imaging experiments showing enhanced responses in human middle superior temporal sulcus for angry relative to neutral prosody. This emotional enhancement was voice specific, unrelated to isolated acoustic amplitude or frequency cues in angry prosody, and distinct from any concomitant task-related attentional modulation. Attention and emotion seem to have separate effects on stimulus processing, reflecting a fundamental principle of human brain organization shared by voice and face perception. PMID- 15665881 TI - [Pure routine]. PMID- 15665882 TI - [Vision rehabilitation after brain injury]. PMID- 15665883 TI - [From science fiction to clinica routine]. PMID- 15665884 TI - [Buprenorphine and methadone to opiate addicts--a randomized trial]. AB - BACKGROUND: There are approximately 12,000 opioid dependants in Norway. Methadone assisted treatment was approved in Norway in 1998, buprenorphine in 2000. This study compares the efficacy of methadone (n = 25) and buprenorphine (n = 25) assisted maintenance treatment in a group of long-term (> 10 years) opioid dependant. MATERIAL AND METHODS: After randomisation patients received either 16 mg sublingual buprenorphine or individually adjusted methadone (mean 106 mg, range 80 - 160) for 26 weeks, with a rehabilitation programme run in parallel. RESULTS: After 180 days, patient retention was highest in the methadone group (85 % vs. 36 %, p < 0.0005). Days in treatment were 167 vs. 114 (95 % CI for difference 53 days (26-80), p < 0.001). Positive urine test rates for opiates (20 % vs 24 %, p < 0.01) and cannabis (33 % vs 45 %, p < 0.001) were lower in the methadone group which also had lower self-reported risk behaviour and psychological distress. However, only those on buprenorphine reported significant improvement in physical health. For older, long-term opioid dependants with significant co-morbidity and unsuccessful medication-free treatment, high-dose methadone maintenance appears to be the treatment of choice. However, in cases where methadone is poorly tolerated, buprenorphine therapy may be a good alternative. PMID- 15665885 TI - [Life following acute subarachnoid haemorrhage]. AB - BACKGROUND: We studied the outcome after aneurysmal subarachnoid haemorrhage from a patient perspective. We compared outcomes of microsurgical and endovascular aneurysm repair and assessed the impact of gender and age on quality of life. MATERIAL AND METHODS: 60 patients undergoing repair of a ruptured intracranial aneurysm from January to July 2001 were included consecutively (35 women). Endovascular repair was performed on 39 and microsurgery on 21. The mean age was 55 years. Quality of life was assessed 3 and 12 months after the bleed using WHOQOL-BREF and Rankin score questionnaires, and through interviews (n = 9). RESULTS AND INTERPRETATION: A total of 40 of 58 patients returned both questionnaires; two died. Twelve months after the bleed, 26 of 44 patients reported a "good" or "very good" subjective quality of life, 3 out of 44 reported a "poor", and 15 of out 44 a "neither good nor poor" subjective quality of life. Males reported a significantly better quality of life (p=0.008). Women reported feeling more depressed and also significantly more symptoms of asthenia, less energy and working capacity. We found no significant differences pertaining to age. Despite moderate disabilities, the respondents generally scored high on quality of life after subarachnoid haemorrhage. The gender differences clearly deserving further investigations. PMID- 15665886 TI - [Primary adrenal failure--causes, diagnostics and therapy]. AB - BACKGROUND AND METHODS: An overview of primary adrenal failure with emphasis on replacement therapy is presented. The article is based on a review of recent literature and authors' personal experience. RESULTS AND CONCLUSIONS: Addison's disease is usually caused by an autoimmune destruction of the adrenal cortex. It is relatively rare (prevalence 14 per 100,000 inhabitants), but often considered as a differential diagnosis when evaluating fatigue, tiredness and loss of weight. Addison's disease is treated by repletion of glucocorticoids and mineralocorticoids. The recommended starting dose is 25 mg cortisone acetate per day divided into three (12.5 + 6.25 + 6.25 mg) or two doses (12,5 mg x 2). Mineralocorticoid replacement is given as fludrocortisone 0.05 - 0.2 mg in one dose per day. Treatment with 20 - 50 mg dehydroepiandrosterone has been studied in adrenal failure, but the evidence for positive effects is weak, and it can not be recommended as standard treatment in primary adrenal failure. PMID- 15665887 TI - [Do vitamins C and E protect against the development of carotid stenosis and cardiovascular disease?]. AB - BACKGROUND: Some observational and randomized, clinical interventional studies have indicated that the antioxidative vitamins C (ascorbic acid) and E (alpha tocopherol) can reduce intima-media thickness of the carotid arteries. It is, however, not clarified whether these vitamins may have a preventive effect against cardiovascular events. MATERIAL AND METHODS: The literature on the effects in relation to several cardiovascular endpoints of vitamins C and E, also used in combination, has been evaluated. The literature has been continuously and systematically collected over many years and supplemented by recent studies retrieved from Medline. RESULTS: Observational studies including mainly healthy individuals have shown a favourable relationship between intake of vitamins C and E, also taken in combination, and subsequent cardiovascular events. However, most randomized, clinical interventional studies including patients with manifest atherosclerotic disease have not been able to document such a relationship. INTERPRETATION: The discrepancy found between observational studies and clinical interventional studies may be due to different study populations (healthy/ill individuals) and differences regarding age, sex, diet, smoking, degree of oxidative stress and other probable confounding factors. The significance of vitamins C and E in preventing cardiovascular disease is in our opinion not yet clarified. PMID- 15665889 TI - [Treatment of cervical intraepithelial neoplasia before and after introduction of laser conization]. AB - BACKGROUND: Cervical intraepithelial neoplasia (CIN) is an established precursor of invasive cervical cancer. Excision procedures such as cold-knife conization, electrodiathermy or laser conization of the cervix are major surgical treatment modalities of CIN. MATERIAL AND METHODS: Women who were treated for CIN 2/3 or suspected invasive cancer with cold-knife conization between 1977 and 1980 (n=212) were compared with women treated with laser conization between 1987 and 1990 (n=439). Outcome parameters were method of anaesthesia, duration of hospital stay, treatment efficacy and postoperative complications such as bleeding, infection or cervical stenosis. RESULTS: General anaesthesia was used in 88 % of women treated with cold-knife conization, while paracervical block anaesthesia was used in 97 % of women treated with laser conization. Mean hospital stay was 7.6 days after cold-knife conization, while laser conization was performed as an outpatient procedure. The overall complication rate was 36.8 % after cold-knife conization and 8.4 % after laser conization. Significantly higher rates of postoperative bleeding (21.1 % v. 5.0 %), infections (2.6 % v. 0.5 %) and cervical stenosis (11.8 % v. 1.6 %) were found after cold-knife conization compared to laser conization. Treatment efficacy was equally high (98 %) with both methods. CONCLUSION: Laser conization was found to be a significantly less resource consuming procedure and with fewer postoperative complications compared to cold-knife conization. PMID- 15665888 TI - [Capsule endoscopy--a new method for the diagnosis of diseases of the small intestine]. AB - BACKGROUND: Diseases of the small intestine are rare and often difficult to diagnose because of a lack of sensitive methods. A new technique called wireless capsule endoscopy has recently been developed; it allows visualisation of the entire small intestinal surface. MATERIAL AND METHODS: The patient swallows a capsule containing a small digital camera, a battery and a radio transmitter. The capsule takes 2 pictures per second during its passage through the intestines; pictures are transferred by radio signals that are detected by sensors on the abdominal skin, stored on a data recorder and later analysed on a PC. RESULTS: By the end of November 2003 we had performed 80 capsule endoscopies in 69 patients. All patients were referred because of suspected small bowel disease and had been through an extensive diagnostic workup before capsule endoscopy was performed. Most of them were referred because of obscure gastrointestinal bleeding or iron deficiency anaemia. Small bowel pathology was found in one third of the patients. INTERPRETATION: Capsule endoscopy seems to be a major improvement in the diagnosis of small intestinal diseases. It is a non-invasive and painless procedure. PMID- 15665890 TI - [Brace treatment of idiopathic scoliosis]. AB - BACKGROUND: Idiopathic scoliosis is a complex three-dimensional deformity of the spine. In order to prevent a progressive course of the disorder, brace treatment is started in selected patients. The aim of this study was to evaluate the effect of brace treatment on the structural components in patients with idiopathic scoliosis. MATERIAL AND METHODS: We selected 54 patients and analysed standing radiographs at different stages. Cobb angle, lateral deviation, wedge angle and axial rotation were calculated. The patients were divided into two groups: group A, characterised by an initial correction with a following increase, and group B, in which the patients' disorder took a progressive course. RESULTS: 43 out of the 54 patients (79.6 %) had an initial correction followed by a gradual increase of the analysed variables. Cobb angle and the lateral deviation demonstrated a maximum significant correction of 23.8 % and 23.2 % respectively. After treatment the Cobb angle was 93.4 % and the lateral deviation was 106.1 % compared to the values measured before treatment. Axial rotation and wedge angle did not show any significant differences during follow up. Despite brace treatment, the disorder took a progressive course in the 11 patients (20.4 %) in group B. INTERPRETATION: This study indicates that brace treatment gives an initial correction of the analysed structural components of the scoliotic spine in four out of five patients. This is followed by a slow increase in the variables. This effect is most pronounced on Cobb angle and lateral deviation. In one out of five patients the disorder took a progressive course in spite of the brace treatment. PMID- 15665891 TI - [Bias from dependent errors in observational studies]. AB - Measurement errors in two variables are dependent when the degree of error in one of them correlates with the degree of error in the other. When dependent error affects measured exposure and measured outcome, the estimated association between the two is likely to be falsely inflated. Such information bias is probably not uncommon in cross-sectional studies providing data on both variables from questionnaires. This often occurs in published studies, but there seems to be limited awareness of the problem. The basic source of dependent error is usually normal variation in certain personality traits, but it may also be in more transitional moods in the study population or inadequate measurement tools. The major precaution that should be taken in order to eliminate bias from dependent error is to break the bond between information on exposure and outcome by gathering data from two separate sources. We should also recognise that not all study designs and not all data are suited for establishing etiology. PMID- 15665892 TI - [Visual problems in cerebral stroke]. AB - BACKGROUND: It is estimated that every year 13,000 persons in Norway have a cerebral stroke. Visual problems after cerebral stroke are frequent but visual rehabilitation is rarely performed. We wanted to estimate the frequency of a visual field defect being detected in patients with cerebral stroke during their stay in a medical department. MATERIAL AND METHODS: Twenty consecutive patients referred to the hospital with cerebral stroke had their visual field examined in the department of medicine with Donders' method. Eighteen of the patients could cooperate in an examination by eye clinic personnel in which visual acuity, eye motility and fundus of the eye were also examined. RESULTS: Visual acuity was mostly within normal limits. No patient had a visual acuity of less than 0.4. Two patients had homonymous visual field defects. Both were detected in the department of medicine. Disturbances of eye motility were detected in 10 patients. INTERPRETATION: Visual problems are common in patients with cerebral stroke. Homonymous defects in the visual field can be detected in the medical departments and visual training should be given. The nature of the disturbances in eye motility is not understood. PMID- 15665893 TI - [Drugs for smoking cessation]. PMID- 15665894 TI - [Telephone accessibility in general practices]. AB - BACKGROUND: We wanted to chart telephone accessibility in 100 randomly selected general practices in Norway and to define a standard against which others could compare themselves. MATERIAL AND METHODS: During one week, the telephone company recorded all calls to each practice and how many were answered, unanswered, or blocked. Mean waiting time for answer and mean conversation time were also registered. Each practice received a report, comparing its own results to those of other practices. An accessibility index was calculated by dividing the percentage of answered calls with mean holding time on the line (waiting time plus conversation time). Background variables about the practices were collected by questionnaires. RESULTS: There were 266 calls (95 % confidence interval 219 - 312) per week per 1000 patients on the list. 66 % (61 - 71) were answered. Mean waiting time was 25 seconds (20 - 30), conversation time 119 seconds (111 - 127). It is estimated that 7.0 calls (6.4 - 7.5) are answered per patient on a list per year. The accessibility index showed large variations, mostly independent of background variables. INTERPRETATION: Compared with earlier registrations, telephone accessibility has improved despite a strong increase in traffic. Structural aspects of the practices do not explain the variation in telephone accessibility; interest and attitudes are probably just as important. PMID- 15665896 TI - [Australia at the cross-roads]. PMID- 15665899 TI - [Doping preparations list and exceptions based on medical reasons]. PMID- 15665900 TI - [Treatment of obstructive sleep apnea]. PMID- 15665903 TI - [Don't antioxidants protect against cancer?]. PMID- 15665904 TI - Effect of stimulation of subthalamic nucleus on grooming movements and their rhythmicity in pups and mature rats. AB - The effect of rhythmic stimulation of subthalamic nucleus on grooming movements and their rhythmicity was studied on mature rats and on 13- and 14-day rat pups. Unilateral monopolar stimulation of subthalamic nucleus was performed on unrestrained animals via implanted electrodes. This stimulation activated virtually all grooming movements (scratching, hair-plucking, licking, and body shaking); the effect was most pronounced in pups. Acceleration of the rhythm of grooming movements was observed only in rat pups for hair-plucking and scratching on the contralateral side relatively to the stimulated nucleus. In mature rats stimulation of the subthalamic nucleus prolonged grooming movements in most cases, but did not increase their number and rhythm. PMID- 15665905 TI - Effect of blood reticulocyte concentration on electrophoretic mobility of erythrocytes in rats. AB - In intact rats electrokinetic characteristics of erythrocytes weakly depended on the intensity of erythropoiesis. This dependence became significant when the degree of erythropoiesis approached the upper limit of variations. Electrophoretic mobility of erythrocytes progressively decreased with increasing blood reticulocyte count. Subpopulations of erythrocytes with low mobility in electric field accumulated in the blood. Our results indicate that erythrocytes whose electrokinetic potential is similar to the mean mobility of circulating cells migrate from the bone marrow into the circulation under normal conditions. PMID- 15665906 TI - Heart resistance to oxidative stress in rats of different genetic strains. AB - In August rats reperfusion after regional myocardial ischemia in situ or intracoronary administration of hydrogen peroxide less significantly suppressed contractile activity of the heart compared to Wistar rats. Activities of catalase and superoxide dismutase in the myocardium during reperfusion remained unchanged in August rats. In Wistar rats a profound inhibition of cardiac function was accompanied by a decrease in enzyme activity. PMID- 15665907 TI - Effect of hypoxia on metabolism and contractile function of the heart in rats with type 2 diabetes mellitus and abdominal obesity. AB - Type 2 diabetes mellitus was modeled in newborn albino rat pups. Metabolism and contractile activity of isolated heart under conditions of hypoxia were studied on adult rats. Contractile activity of the myocardium in animals with type 2 diabetes mellitus and abdominal obesity decreased during hypoxia. It was manifested in a decrease in systolic and developed pressure and disturbances in diastolic relaxation of the myocardium. Damage to cell membranes and increased secretion of aspartate transaminase into the coronary circulation were observed under conditions of energy deficit and activation of the anaerobic pathway of energy production. These changes became more pronounced with increasing the period of hypoxic exposure. PMID- 15665908 TI - Reactivity of alveolar macrophages during granulomatous inflammation of the lungs under conditions of acute massive blood loss. AB - Reactivity of mouse alveolar macrophages by their ability to phagocytize killed St. aureus bacteria and production of reactive oxygen metabolites (nitro blue tetrazolium test) in response to zymosan administration was studied under normal conditions and after acute massive blood loss. Zymosan-induced granulomatous inflammation of the lungs during acute massive blood loss 2-fold inhibited the increase in oxidative metabolism of alveolar macrophages. Suppressed production of toxic oxygen radicals in alveolar macrophages was accompanied by accelerated recovery of cells on the surface of the respiratory tract. PMID- 15665910 TI - Transcriptional activation of cytochrome P450 1A1 with alpha-tocopherol. AB - Peroral administration of alpha-tocopherol in a daily dose of 150 mg/kg for 1, 4, 8, and 12 days leads to induction of cytochromes P450 1A in male rats. Activity of CYP1A1 and CYP1A2 increased most significantly one day after alpha-tocopherol administration (by 2.6 and 2.7 times, respectively). CYP1A1 was immunohistochemically detected in rat liver microsomes during this period. The content of CYP1A1 mRNA significantly increased in the liver. The amount of CYP1A2 mRNA and regulatory proteins for signal activation of CYP1A1 (AhR and Arnt) remained unchanged after treatment with alpha-tocopherol. PMID- 15665909 TI - Caspase inhibitor Z-VAD-FMK potentiates heat shock-induced apoptosis and HSP70 synthesis in macrophages. AB - Caspase inhibitor Z-VAD-FMK potentiated heat shock-induced apoptosis in macrophages. Z-VAD-FMK did not activate HSP70 synthesis, but significantly increased the intensity of this process during heat shock. It cannot be excluded that caspases abolish HSP70 accumulation under these conditions. The HSP70 synthesis inhibitor quercetin potentiated DNA fragmentation in macrophages cocultured with Z-VAD-FMK after heat shock. HSP70 play an important role in the protection of macrophages from caspase-independent apoptosis. PMID- 15665911 TI - Effect of long-term cold exposure on activities of cytochrome P450-containing monooxygenases and glutathione S-transferase in rat liver microsomes. AB - Long-term cold exposure (5 degrees C) was followed by induction of rat liver monooxygenases. We revealed an increase in activity of NADPH-cytochrome C reductase, total content of cytochrome P450 (CYP), and activities of its molecular forms CYP1A1, 1A2, 2B1/B2, 2E1, and 3A1/A2 in microsomes. These indexes reached maximum by the 10th day, but decreased with lengthening of cold exposure. Glutathione S-transferase activity decreased under these conditions. Changes in enzyme activity could be related to the increase in blood corticosterone concentration. PMID- 15665912 TI - Activity of cytochrome P450 in children. AB - The 6beta-hydroxycortisol/cortisol ratio was measured in 52 children aging 1.1 14.0 years. The maximum increment in this ratio occurred in the age interval of 1.1-2.0 years. During this period, the regression coefficients in the linear (r=0.57; p=0.044) and nonlinear logarithmic models (r=0.56; p=0.049) were similar. At the age of 10-14 years, the examined ratio attained 19.17+/-17.79. PMID- 15665913 TI - Changes in intracellular potassium concentration in a one-cell mouse embryo after enucleation. AB - Comparative analysis of potassium concentrations in the cytoplasm of intact and enucleated one-cell mouse embryos of showed that microsurgical manipulations during collection of pronuclei disordered potassium homeostasis in the embryonic cell. PMID- 15665914 TI - Effect of oxidized fibrinogens on blood coagulation. AB - We studied the effect of UV-irradiated fibrinogen on blood coagulation. Fibrinogen with oxidation degree of 10% moderately activated the intrinsic pathway, but inhibited the extrinsic pathway of blood coagulation. Fibrinogen with oxidation degree of 20% inhibited both the extrinsic and intrinsic blood coagulation pathways. We revealed disturbances in the formation of fibrin clot with oxidized fibrinogen, suppression of platelet aggregation mediated by collagen receptors, and inhibition of aggregation associated with von Willebrand factor activity. ADP initiated platelet aggregation, which was in direct proportion to the degree of fibrinogen oxidation. Our results indicate that oxidized fibrinogen produces a dysregulatory effect on platelets. PMID- 15665915 TI - Low ionic strength promotes association of circulating modified LDL in human blood. AB - The resistance to association of circulating multiply-modified low-density lipoproteins (LDL) isolated from human blood and characterized by a decreased content of sialic acids in comparison with native LDL was studied by analysing light transmission fluctuations. LDL association was stimulated by decreasing environmental ionic strength. It is established that circulating modified LDL are less resistant to association than native LDL. Association of LDL in a medium with low ionic strength was irreversible. Probably, increased capacity to irreversible association determines the atherogenic properties of circulating modified LDL subfraction. PMID- 15665916 TI - Stromal cell strains derived from suspension fraction of bone marrow cultures of tumor necrosis factor-deficient mice. AB - Culturing of suspension fraction of a long-term bone marrow culture derived from tumor necrosis factor (TNF)-deficient mice for 75 days produced cells forming adherent cell strains. The cells of all strains expressed RNA of various stromal differentiating markers in various combinations. The cells of many strains simultaneously expressed genes encoding products characteristic of different differentiation lineages. The derived strains maintained hemopoiesis for 10 weeks. RNA-analysis of gene expression by cells of these strains showed that they express a set of various growth factors and cytokines. It was hypothesized that suspension fraction of long-term bone marrow culture derived from TNF-deficient mice includes immature stromal precursor cells, which were never detected in long term bone marrow culture derived from wild-type mice. PMID- 15665917 TI - Effect of carnosine on immunocompetent cells from alcoholic patients. AB - We studied the effect of natural dipeptide carnosine on phagocytosis, respiratory burst in neutrophils, and subpopulation composition of lymphocytes from healthy donors and alcoholic patients. Carnosine in vitro produced different effects on immunocompetent cells from healthy donors and patients with alcoholism. In patients with alcoholism phagocytic activity of leukocytes and generation of reaction oxygen species increased under the influence of carnosine in a concentration of 0.01 mM, but decreased after treatment with this compound in a concentration of 1 mM. Carnosine in both concentrations stimulated the respiratory burst, but had no effect on the count of phagocytic cells in healthy donors. Carnosine in a concentration of 0.01 mM increased the number of lymphocytes carrying apoptosis markers (CD95+) in patients with alcoholism not receiving therapy. Our results indicate that carnosine holds much promise for the therapy of alcoholism. PMID- 15665918 TI - Search for potent modulators of cytokine production by macrophages. AB - We compared the effects of Tamerit, Polyoxidony, and Licopid on spontaneous and lipopolysaccharide-stimulated production of interleukin-1 and tumor necrosis factor by mouse peritoneal macrophages in vitro. The test preparations were equally potent in stimulating nonactivated cells. Licopid produced a costimulatory effect on macrophages primed with endotoxin. Tamerit in different doses suppressed cytokine production by cells. Polyoxidony in low doses activated, but in high doses suppressed this process. PMID- 15665919 TI - Stimulatory effect of green tea extract on the growth of neurites in the rat spinal ganglion culture. AB - Addition of green tea extract in concentrations of 0.004-0.006% to the nutrient medium markedly stimulated the growth of spinal ganglion neurites of 1-2-day-old rats. PMID- 15665920 TI - Preventive and therapeutic effect of complex antioxidant preparation in rats with burn trauma. AB - The production of blood radicals and activity of superoxide dismutase in erythrocytes increased in rats with contact burn trauma (20%). In animals with burn trauma antioxidant activity of the plasma was much lower, while myeloperoxidase content in the lung tissue and epidermis was higher than in control rats. The complex of antioxidants (Immudzhen) inhibited radical generation at the peak of inflammation (day 4), increased antioxidant activity of the plasma, and normalized myeloperoxidase content in the lung tissue. PMID- 15665921 TI - Status of the progeny of rats treated with platinum-containing cytostatics. AB - Wistar rat pups from rats treated with first- and second-generation platinum containing cytostatics (platidiam, carboplatin) 1, 3, and 6 months before mating with intact partners had similar disorders. The severity of these disorders depended in many cases on the chemical structure of the drug and sex of the parent treated with the cytostatic. The severity of toxic effects in the progeny of intact females mated with cytostatic-treated males decreased with prolongation of the period elapsing between the treatment and mating. Carboplatin produced a more potent toxic effect on the reproductive function of rats compared to platidiam. PMID- 15665922 TI - Effects of low ethanol doses on heart rhythm in rabbits. AB - Effects of low ethanol doses on the vagosympathetic mechanisms of heart rhythm regulation were studied in rabbits. Analysis of heart rhythm variability showed that single intragastric administration of 0.5 mg/kg ethanol caused tachycardia in animals with initial predominance of vagal activity and bradycardia in animals with predominating sympathetic mechanisms. This was associated with general activation of all regulatory effects on the heart rhythm and a drastic increase in power spectrum for all frequency ranges, though with a certain deficiency of vagal effects. However, after 24 h the vagal component of the spectrum drastically increased in animals of both groups, while other parameters did not differ from the control. Presumably, this rebound can be used as a physiological marker, ethanol tolerance measure, formation of the abstinence syndrome and liability to alcoholism. PMID- 15665923 TI - Effect of human serum on bioluminescence of natural and recombinant luminescent bacteria. AB - Biphasic modification of bacterial bioluminescence by human serum was revealed: bioluminescence was inhibited at high concentrations of the serum and stimulated at low concentrations. Effects of temperature and duration of exposure on bioluminescence manifested in stimulation of the inhibitory effect at higher temperature and longer exposure. The degree of inhibition of bioluminescence under in the presence of serum depends on species characteristics of the microorganism and nature of the luminescent system. PMID- 15665924 TI - Detection of stages of autoimmune hemolytic anemia by evaluating erythrocyte deformability and density. AB - Study of erythrocyte density and deformability in patients with hemolytic anemia, including long-term monitoring of 5 patients, helped us to characterize the pathological processes leading to changes in the erythrocyte population at different terms of the disease and to detect its main stages (agglutination, pathological dehydration, combination of pathological dehydration and microvesiculation, hemolytic crisis, and remission). PMID- 15665925 TI - Damage to cell DNA in the bone marrow and testes of mice with experimental trichinosis. AB - Metabolites of Trichinella spiralis produced genotoxic and cytotoxic effects on somatic and generative cells of the host organism. They increased the number of single-chain breaks, alkaline-labile sites in nuclear DNA, and count of apoptotic cells in the bone marrow and testes of infected mice. These effects depended on the stage of parasite development in the host organism and became more pronounced with increasing invasion intensity. PMID- 15665926 TI - Effects of extracts from medicinal plants on the development of metastatic process. AB - The possibility of inhibiting metastases by preparations from plants growing in Siberia and Far East after removal of the tumor node was proven in experiments on animals with transplanted tumors. Plant preparations stimulated the antitumor resistance of the organism. PMID- 15665927 TI - Sensitivity of Lewis lung carcinoma to cisplatin undergoes considerable variations during growth and metastasizing. AB - Growth and metastasizing of Lewis lung carcinoma were accompanied by complex changes in tumor sensitivity to cisplatin. On day 17 of growth, tumor sensitivity was much lower than immediately after transplantation. Further growth of Lewis lung carcinoma was accompanied by a progressive increase in its sensitivity to cytostatic treatment. The study of migration activity of the tumor and dynamics of metastasizing showed that metastatically active carcinoma cells dominated on day 17 of tumor growth, but then the number of these cells progressively decreased to the 29th day of tumor growth. Growth-related changes in the sensitivity of Lewis lung carcinoma were probably due to complex variations in the number of metastatically active tumor cells with high resistance to cisplatin. PMID- 15665928 TI - Contractile function of the smooth-muscle wall and its adrenergic regulation in megaureter. AB - We demonstrated disordered contractile capacity of megaureters formed against the background of reflux, organic or functional obstruction. The rhythmic and tonic contractions were suppressed in the megaureters with organic obstruction in comparison with megaureters caused by reflux. Norepinephrine stimulation induced pronounced pressor responses in obstructive megaureters. The state of the smooth muscle walls of megaureters with functional obstruction was characterized by high tone, weak phase contractions, and increased capacity to pressor tonic reactions induced by norepinephrine. PMID- 15665929 TI - Treatment of acute coronary syndromes in patients with type 2 diabetes mellitus with beta-adrenoblockers and angiotensin-converting enzyme inhibitors: cardiohemodynamic effects and impact for prognosis. AB - Long-term oral treatment of patients with acute coronary syndromes and type 2 diabetes mellitus with beta-adrenoblockers and angiotensin-converting enzyme inhibitors is associated with positive, though ambiguous changes in the left ventricular structure and function. These changes should be the reason for choosing optimal therapy ensuring better prognosis in this patient population. PMID- 15665930 TI - Crystallization of oral fluid components in patients with type 2 diabetes mellitus. AB - Crystal aggregations of oral fluid from normal subjects and patients with type 2 diabetes mellitus were examined. Morphological signs characterizing crystal aggregations of salivary pools from patients with type 2 diabetes mellitus are described and classified. PMID- 15665931 TI - Regeneration of the spleen in intact animals and radiation chimeras. AB - Regeneration of the splenic tissue after partial splenectomy is incomplete in adult non-irradiated mice and lethally irradiated animals reconstituted with donor syngeneic bone marrow. Transplantation of the splenic tissue to intact adult animals after partial splenectomy resulted in virtually complete regeneration of the spleen. In chimeras recovery of the splenic tissue was decreased; autotransplantation of the whole spleen or its part did not lead to appreciable changes in the weight and cellularity of this organ. No more than 30% splenic tissue is restored after complete splenectomy and transplantation of the splenic tissue in intact and chimeric mice. PMID- 15665932 TI - Effects of new wound dressings on healing of thermal burns of the skin in acute radiation disease. AB - Effects of new wound dressing bacterial cellulose impregnated with SOD and poviargol (Procel-Super and Procel-PA) and Inerpan hydrogel dressing on the reparative processes in deep dermal burns (IIIa-IIIb degree) in rats exposed to total even irradiation in a dose of 4 Gy were studied. Inerpan and Procel-Super dressings proved to be the most effective under these conditions: they accelerated healing of burn wounds by 17.0 and 5.5%, respectively. PMID- 15665933 TI - In vitro regulation of cell behavior by calcium phosphates synthesized by the mechanochemical method. AB - Biological activity of eight hydroxyapatite powders synthesized by the mechanochemical method was studied in vitro. Introduction of copper and zinc atoms into synthetic hydroxyapatite structure promotes cell adhesion. Hydroxyapatites obtained by the mechanochemical method are promising base for a new class of materials and coatings with regulated bioactivity for orthopedics and traumatology. PMID- 15665934 TI - Ability of L-histidine to decrease desensitization of the myometrium to epinephrine. AB - Experiments were performed with 62 longitudinal strips of the uterine horns from 10 nonpregnant rats. The ability of continuously infused epinephrine in high concentration (10(-6) g/ml, 30 min) to suppress spontaneous contractions due to activation of beta-adrenoceptors progressively decreased, which was associated with receptor desensitization. Histidine in a concentration of 3 x 10(-11) g/ml had no effect, while in concentrations of 3 x 10(-8), 3 x 10(-7), and 3 x 10(-6) g/ml decreased the degree of desensitization. Our results indicate that histidine not only potentiates beta-adrenoceptor activation, but also prevents the development of desensitization. These data should be taken into account during therapy with beta-adrenoceptor agonists. PMID- 15665935 TI - Effect of serotoninergic and cholinergic nervous systems and corresponding neurotransmitters on heart function and motor activity of pelvic smooth muscle organs. AB - Serotonin potentiated vagal negative chronotropic effect in rabbits and increased the synergistic action of autonomic nervous structures on cardiac function in frogs. Acetylcholine in low doses produced a positive chronotropic effect in frogs, which was related to activation of intracardiac adrenergic neurons. Simultaneous activation of the serotoninergic and cholinergic system suppressed heart function, but increased motor activity of pelvic smooth muscle organs. PMID- 15665936 TI - Effect of MK-801 on sensitivity of the respiratory system to oxygen deficiency and organisms resistance to hypoxia. AB - We studied changes in organism's sensitivity and resistance to hypoxic hypoxia under conditions of NMDA receptor blockade with MK-801. Breathing hypoxic gas mixture after administration of MK-801 sharply decreased the mean blood pressure in the systemic circulation and slightly improved lung ventilation in animals. Respiratory arrest was observed in 40% animals. NMDA receptor blockade decreased organism's sensitivity and resistance to hypoxic hypoxia. PMID- 15665937 TI - Mechanisms of regulation of erythropoiesis during hemolytic anemia. AB - We studied changes in the erythroid hemopoietic stem during phenylhydrazine induced hemolytic anemia. Stimulation of erythropoiesis was associated with increased functional activity of erythroid precursors, which resulted from changes in feeder capacity of hemopoiesis-inducing microenvironmental cells and erythropoietic activity of the plasma. The development of encephalopathy induced by a hemolytic poison was accompanied by a decrease in hyperplasia of bone marrow erythropoiesis. It was related to a decrease in the number of proliferating erythroid precursor cells. These changes accompanied the increase in the secretory function of adherent myelokaryocytes, rise in erythropoietic activity of the plasma, enhanced formation of erythroid hemopoietic islets, and accelerated maturation of hemopoietic cells. PMID- 15665938 TI - Effect of in vivo hypoxic preconditioning on changes in intracellular calcium content induced by long-term anoxia in rat brain slices. AB - We studied changes in intracellular calcium content induced by 10-min anoxia in olfactory cortex slices from rats exposed to single or 3-fold moderate hypobaric hypoxia ("ascend" at 5000 m). Repeated preconditioning with moderate hypobaric hypoxia produced a neuroprotective effect. This treatment abolished pathological calcium overload in brain slices induced in vitro by 10-min test anoxia. PMID- 15665939 TI - Free radical lipid oxidation in brain cortex neurons and neuroglia during convulsions. AB - The amount of lipid peroxidation products (conjugated dienes and trienes and Schiff bases) in cortical neurons of the cerebral hemispheres decreased by 30% at the peak of convulsions observed 10-15 min after intraperitoneal injection of picrotoxin. In neuroglial cells of control animals the intensity of lipid peroxidation in was 1.7-2.0 times lower. Picrotoxin had no effect on this parameter. PMID- 15665940 TI - Selective accumulation of monoclonal antibodies against neurospecific enolase in brain tissue of rats with middle cerebral artery occlusion. AB - Preparations of I(125)-labeled monoclonal antibodies against neurospecific enolase and mouse plasma IgG1 were injected intravenously to rats immediately after unilateral occlusion of the middle cerebral artery. Radioactivity of I(125) labeled monoclonal antibodies against neurospecific enolase in the brain tissue progressively increased, reached a maximum by the 48th hour, and remained practically unchanged after 72 h. At the same time radioactivity of labeled IgG1 in the brain tissue and radioactivity of both preparations in the blood, liver, spleen, kidneys, heart, and lungs decreased over 72 h. Selective accumulation of I(125)-labeled monoclonal antibodies against neurospecific enolase was less significant in the brain tissue of the contralateral hemisphere and cerebellum not exposed to ischemia. PMID- 15665941 TI - Age-related differences in the response of blood stroke volume to stimulation of the sympathetic ganglion in rats with beta-adrenoceptor blockade. AB - Blood stroke volume in rats aging 21 and 56 days decreased during beta adrenoceptor blockade with propranolol, but increased again by the 15th minute after treatment. Suprathreshold stimulation of the stellate ganglion decreased the stroke volume and increased the heart rate in control animals. Electrical stimulation after beta-adrenoceptor blockade was followed by a further decrease in stroke volume in young rats. In 100-day-old animals this parameter remained unchanged, while the cardiac output improved. PMID- 15665942 TI - Effect of perftoran on macroglobulin content in the plasma and peritoneal exudate of rats with acute exudative inflammation (peritonitis). AB - The effect of blood substitute perftoran on the content of alpha(1)- and alpha(2) macroglobulins in the plasma and exudate was studied in rats with acute exudative inflammation. After intravenous injection of perftoran macroglobulin content increased in the plasma, but remained unchanged in the peritoneal fluid. PMID- 15665944 TI - Recovery of contractile function of cryodamaged rat myocardium after transplantation of fetal cardiomyocytes and predifferentiated bone marrow stromal stem cells. AB - The effect of cell transplantation into cryodamaged rat myocardium was studied on isolated hearts by increasing functional load to the left ventricle. Transplantation of allogeneic fetal cardiomyocytes improved the function of the left ventricle under conditions of considerably increased preload. Transplantation of autologous mesenchymal stem cells repaired left-ventricular function under conditions of increased pre- and afterload. PMID- 15665943 TI - Phagocytic activity of neutrophils and lymphocytes at various periods of storage in the state of cold anabiosis (-40 degrees C). AB - We studied changes in phagocytic activity of neutrophils and lymphocytes subjected to cold anabiosis. The proposed effective method for introduction of human blood leukocytes into cold anabiosis at moderately low temperature (-40 degrees C) in the presence of a cryopreserving agent does not require washout of the biological material after defrosting. PMID- 15665945 TI - Natural antioxidant L-carnosine inhibits LPO intensification in structures of the auditory analyzer under conditions of chronic exposure to aminoglycoside antibiotics. AB - Intragastric administration of L-carnosine suspension to Wistar-Kyoto rats 3 days before and after 7-day course of intraperitoneal injections of ototoxic aminoglycoside antibiotic kanamycin compensated expenditures of tissue antioxidant systems and significantly eliminated kanamycin-induced intensification of MDA production in tissues of the membrane part of the cochlea and in the auditory cortex of the temporal lobe. L-NAME (competitive NO synthase inhibitor) also inhibited LPO, increased total antioxidant activity, and decreased ototoxicity of kanamycin, which confirms the contribution of NO into LPO intensification under conditions of aminoglycoside treatment. Inhibition of pathological intensification of LPO processes and increase in total antioxidant activity under conditions of induced acute aminoglycoside ototoxicity characterizes L-carnosine as a highly effective otoprotector. PMID- 15665946 TI - Initiation of stress protein synthesis in the myocardium of coronary patients. AB - We studied myocardial biopsy specimens from the right atrium of cardiological patients with different degree of cardiac ischemia obtained during surgery. No inducible HSP70 stress proteins were detected in atrial cardiomyocytes of patients with the WPW syndrome without signs of ischemic injuries of the heart. These proteins were detected in the myocardium of coronary patients. Their expression was more intense in patients with coronary disease paralleled by the development of myocardial dyskinesia. Two-dimensional electrophoresis showed only acid HSP70 but no alkaline isoforms in coronary patients even with pronounced dyskinesia. Presumably, alkaline HSP70 isoforms are present in the myocardium directly involved in the dyskinesia zone. PMID- 15665947 TI - Stimulation of immunotoxicity of chemicals metabolizing in vivo into highly toxic compounds by the monooxygenase system inductors. AB - Oral treatment of experimental random-bred albino rats with inductors of the monooxygenase system phenobarbital (50 mg/kg) and benzenal (70 mg/kg) for 3 days until acute poisoning with toxins (methanol, ethylene glycol, and dichloroethane in doses of 1.0 LD50) metabolizing in the body to compounds with higher toxicity (phenomenon of lethal synthesis) increased immunotoxicity of these inductors. PMID- 15665948 TI - Role of phosphatidylinositol-3-kinase and protein kinase C zeta in the adenylate cyclase signal mechanism of action of relaxin in muscle tissues of rats and mollusks. AB - We showed that phosphatidylinositol-3-kinase and protein kinase C zeta are involved in the adenylate cyclase signal mechanism of relaxin action. A selective inhibitor of phosphatidylinositol-3-kinase wortmannin blocked the stimulatory effect of relaxin on adenylate cyclase in rat skeletal muscles and Anodonta cygnea smooth muscles. Antibodies against protein kinase C zeta abolished the relaxin-induced stimulation of adenylate cyclase in rat muscles, but not in mollusk muscles. Our results indicate that phosphatidylinositol-3-kinase and protein kinase C zeta play a role in the adenylate cyclase signal mechanism of relaxin action. PMID- 15665949 TI - Delta-opioid receptor antagonists exhibit properties of partial delta-receptor agonists in isolated perfused heart. AB - Perfusion of the isolated intact rat heart with Krebs-Henseleit solution containing agonists ((-)-TAN-67, DPDPE, and dalargin) or antagonists of delta opioid receptors (naltrindole, TIPP[psi], and ICI 174,864) in a final concentration of 0.1 mg/liter was followed by a decrease in the heart rate, end diastolic pressure, contraction rate, relaxation rate, and left ventricular developed pressure. Perfusion with a solution containing the delta-opioid receptor agonist DPDPE or delta-antagonists naltrindole, TIPP[psi], and ICI 174,864 before modeling of global ischemia increased the severity of reperfusion induced contractile dysfunction in the myocardium. Our results suggest that delta opioid receptor antagonists in vitro exhibit properties of partial delta-receptor agonists. PMID- 15665950 TI - Resistance of native and circulating modified low-density lipoproteins in human blood to association. AB - The resistance of native and circulating modified low-density lipoproteins from human blood to spontaneous and polyethylene glycol-induced association was studied by recording light transmission fluctuations. Circulating modified low density lipoproteins were less resistant to association than native low-density lipoproteins. Polyethylene glycol-induced association of low-density lipoproteins was irreversible. Our results suggest that atherogenic activity of circulating modified low-density lipoproteins is associated with their increased predisposition to irreversible association. PMID- 15665951 TI - Postirradiation volatile secretions of mice: syngeneic and allogeneic immune and behavioral effects. AB - The immune response in immunologically and olfactorily high- and low-reactive CBA and C57Bl/6 mice is almost similarly decreased after exposure to volatile secretions of syngeneic animals exposed to ionizing radiation in a dose of 4 Gy. In the preference/avoidance test intact animals prefer secretions of irradiated syngeneic and allogeneic animals to those of intact animals, while without irradiation animals of both strains prefer syngeneic secretions. C57Bl/6 mice differ from CBA animals by lower sensitivity. PMID- 15665952 TI - Effect of dehydroepiandrosterone on radioligand binding of [3H]-testosterone by androgen receptors in rat hypothalamus. AB - Intramuscular injections of dehydroepiandrosterone in a dose of 0.7 mg/kg for 10 days significantly increased nuclear and cytoplasmic fractions of androgen receptors in the preoptic/anterior hypothalamic area. Presumably, the effect of the neurosteroid is mediated by 5 alpha-reductase transformation of dehydroepiandrosterone into 5 alpha-dehydroepiandrosterone, which initiates the synthesis of androgen receptors. PMID- 15665953 TI - Activating effect of Tanacetum vulgare L. pectin polysaccharide on ionic channels of neuronal membrane. AB - The membranotropic effects of TVF tanacetan pectin polysaccharide derived from Tanacetum vulgare L. was studied by the voltage-clamp method on isolated neurons of Lymnaea stagnalis mollusk. TVF in concentrations of 0.1-10.0 microg/ml nonselectively activated the outward potassium and total inward (sodium and calcium) ionic currents (slightly dose-dependently and reversibly increased their amplitude by 5-10%) and decreased nonspecific leakage current. PMID- 15665954 TI - Effects of dimephosphone, xydiphone, and ionol on the content and activities of rat liver cytochromes P-450 during long-term treatment with phenobarbital. AB - Effects of dimephosphone, xydiphone, and ionol administered in parallel with phenobarbital on the content of cytochromes P-450 in rat liver and on the rate of C-hydroxylation of diazepam, haloperidol, and prednisolone by rat liver microsomal enzymes were studied in vitro. Dimephosphone, xydiphone, and ionol exhibited similar inductive effects on C-hydroxylation reactions in the CYP P-450 system during treatment with phenobarbital. Xydiphone and ionol in a dose of 1 mmol/kg canceled phenobarbital-induced increase in P-450 cytochrome content in rat liver. Sex-dependent cytochromes P-450 are involved in the prednisolone and haloperidol C-hydroxylation reactions in rats. PMID- 15665955 TI - Effect of NO synthase inhibitor 2-amino-5,6-dihydro-4H-1,3-thiazine on endotoxin induced changes in hemodynamic parameters and respiration in rats. AB - We studied the effect of NO synthase inhibitor 2-amino-5,6-dihydro-4H-1,3 thiazine (2-ADT) on the cardiovascular system in rats with endotoxic shock. Blood pressure, heart rate, and respiratory rate were recorded. E. coli lipopolysaccharide decreased blood pressure and heart rate. 2-ADT in a dose of 5 mg/kg normalized these hemodynamic parameters. The normalizing effect of 2-ADT decreased with increasing the dose of this preparation. 2-ADT in high doses (10, 20, and 30 mg/kg) and repeated injections of the preparation caused death of experimental animals. PMID- 15665956 TI - Chronic administration of imipramine decreases freezing time in rats genetically predisposed to catalepsy. AB - The effects of acute and chronic imipramine treatment on the degree of catalepsy were compared in GC rats genetically predisposed to catalepsy. We recorded the time over which the rats remained in a vertical position they were placed. As differentiated from acute treatment, chronic administration of imipramine dose dependently decreased the time of freezing in GC rats. PMID- 15665957 TI - MDR1 Gene C1236T and C6+139T polymorphisms in the Russian population: associations with predisposition to lymphoproliferative diseases and drug resistance. AB - Study of MDR1 polymorphism in intron 6 and exon 12 of healthy individuals and patients with chronic lymphoproliferative diseases showed that the presence of mutant 6+139T allele is a factor determining resistance to lymphoproliferative diseases. Comparison of genotyping results in 53 patients and the data on the efficiency of drug therapy showed no significant associations of C(6+139)T and C(1236)T genotypes with drug resistance. PMID- 15665958 TI - Kinetic characteristics of biochemical parameters during consupren therapy after allogenic transplantation of the kidney. AB - Changes in biochemical and common clinical parameters of the blood were detected in patients treated with consupren (cyclosporin) as the main immunosuppressant after allogenic transplantation of the kidney. Kinetic model for evaluation of treatment adequacy was based on therapeutic drug monitoring and monitoring of blood biochemistry in recipients of the primary kidney transplant. The k1, t1/2, C0, clearance (for hemoglobin), conditioned volume of distribution (for cyclosporin) of processes of biochemical parameters (toxicity markers) stabilization within the framework of a single-part kinetic model were determined. PMID- 15665959 TI - Efficiency and mechanisms of the antioxidant effect of standard therapy and refracterin in the treatment of chronic heart failure in elderly patients with postinfarction cardiosclerosis. AB - Refracterin therapy of patients with chronic heart failure caused by coronary heart disease and postinfarction cardiosclerosis markedly promoted improvement in the pulmonary and systemic circulation in comparison with patients receiving traditional therapy. The mean functional class of chronic cardiac failure decreased by 43% under the effect of refracterin vs. 27% decrease in the group receiving traditional therapy. After 1-month refracterin course the end-systolic and end-diastolic sizes of the left ventricle decreased by 12 and 7%, respectively, ejection fraction increased by 7.2% in comparison with the initial level, total oxidant activity and MDA content in the plasma decreased significantly, while total antioxidant activity, catalase and SOD activities, cytochrome C, NADH, and NADPH levels increased. The prooxidant-antioxidant system was shifted towards antioxidants, which attests to activation of the defense and adaptive mechanisms after administration of refracterin, which is especially important in elderly patients with initially decreased reserve potentialities of the antioxidant defense system. PMID- 15665960 TI - Immunomorphological characteristics of animals with different levels of orientation and exploratory behavior. AB - We revealed some relationships between the level of orientation and exploratory behavior, functional activity of the immune system, and structural and functional organization of the CNS in animals. Significant differences in brain morphology and expression of cytokine IL-1 beta, type I IL-1 receptor, and erythropoietin receptor genes in brain cells were detected in (CBA x C57Bl/6)F1 mice with different initial levels of orientation and exploratory behavior. Immunocompetent cells of mice with high and low levels of exploratory behavior differ by spontaneous and mitogen-induced proliferative activity. The formation of humoral and cellular immune response in these animals causes opposite changes in exploratory behavior and the type of these changes depends on the initial level of this behavioral reaction. PMID- 15665961 TI - Structural and functional changes in gastric epithelium in Helicobacter pylori associated chronic gastroduodenal pathologies. AB - Complex structural analysis of the gastric mucosa was carried out in patients with Helicobacter pylori-associated chronic gastroduodenal ulcers, chronic gastritis, and vibration gastropathy. Microscopic examination showed stereotypical changes in the epithelium in all diseases: degeneration, focal intestinal metaplasia, dysplasia, and glandular atrophy. The severity of these changes depended on the disease entity. The most typical ultrastructural modifications of epitheliocytes were damage to the apical plasmalemma, heterogeneity of the secretory compartment of the cytoplasm, dilatation of the cytoplasmic reticulum, vacuolation, and signs of cytolysis. Plastic reactions of the gastric epithelium reflected disproportional changes in all cell metabolites caused by increased proliferative activity of the epithelium under conditions of uneven inhibition of intracellular protein synthesis. PMID- 15665962 TI - Comparative study of vasaprostan and alprostan in spontaneous platelet aggregation test in mice. AB - We propose a method for evaluation of spontaneous aggregation of mouse platelets based on routine clinical blood tests for humans. Comparative analysis of Vasaprostan and Alprostan drugs showed that this test can be used for testing of new drugs on animals. PMID- 15665963 TI - Hereditary muscular dystrophy in MDX mice as a homologous model for introduction of cell technologies in the treatment of progressive muscular dystrophies in humans. AB - Life-time monitoring of the main clinical and laboratory manifestations of hereditary muscular dystrophy in mdx mice confirmed the presence of mutation in exon 23 of dystrophin gene and the absence of this protein in skeletal muscles of mutant animals. Muscular dystrophy in mice was similar to human progressive muscle disorder, which allows the use of this model for the development of cell technologies for the treatment of hereditary muscular diseases in humans. PMID- 15665964 TI - SELEX: just another separation? AB - The potential for quickly isolating high affinity, highly selective ligands generated much excitement when SELEX was first described. Fifteen years later, SELEX has still not achieved widespread acceptance due to limitations in aptamer affinity, stability and throughput. Recent research is beginning to address these limitations though and SELEX is quickly regaining its standing as a hot topic in areas as diverse as drug discovery, chemical analysis, and even genomics and proteomics. PMID- 15665965 TI - Novel ion chromatographic stationary phases for the analysis of complex matrices. AB - Ion chromatography (IC) has a proven track record in the determination of inorganic and organic anions and cations in complex matrices. Recently, application of IC to the separation and determination of bio-molecules such as amino acids, carbohydrates, nucleotides, proteins and peptides has also received much attention. The key to the determination of all of the above species in the most analytically challenging complex matrices is the ability to manipulate selectivity through control of stationary phase chemistry, mobile phase chemistry and the choice of detection method. This Tutorial Review summarises some of the most significant recent advances made in IC stationary phase technology. In particular, the review details stationary phases specifically designed for ion analysis in complex sample matrices, and considers in which direction future stationary phase development might proceed. PMID- 15665966 TI - Spurious serotonin dimer formation using electrokinetic injection in capillary electrophoresis from small volume biological samples. AB - One normally assumes that the analytical measurement process does not introduce spurious compounds. Capillary electrophoresis is a separation method frequently used for small-volume biological measurements. We demonstrate the potential for creating new peaks in a capillary electropherogram when using electrokinetic injections and illustrate the potential deleterious effects with biological samples involving serotonin and nitric oxide measurements. Specifically, when measuring the serotonin content from individual neurons using electrokinetic injections from 360 nL stainless steel vials, we detect a new peak that we identify as a serotonin dimer. We do not observe this peak when using hydrodynamic injections. PMID- 15665967 TI - Headspace gas chromatography-mass spectrometry: a fast approach to the identification and determination of 2-alkyl-3- methoxypyrazine pheromones in ladybugs. AB - Static headspace sampling technique coupled with gas chromatography and mass spectrometry was used to investigate the presence of volatile 2-alkyl-3 methoxypyrazines in three different species of ladybugs of the Coccinellidae family. The species investigated were Coccinella septempunctata, Harmonia axyridis and Hippodemia convergens. 2-isopropyl-3-methoxypyrazine (IPMP) was identified in all three species with detectable levels of 2-sec-butyl-3 methoxypyrazine (SBMP) and 3-isobutyl-2-methoxypyrazines (IBMP) in only Hippodemia convergens and Harmonia axyridis species. Relative amounts of 2-alkyl 3-methoxypyrazines based on body mass showed that Hippodemia convergens had the highest levels of all three methoxypyrazines and Coccinella septempunctata the least. PMID- 15665968 TI - Raman spectroscopy of endoliths from Antarctic cold desert environments. AB - Six endolithic communities from Antarctic cold desert environments have been analysed by Raman spectroscopy. The extreme conditions that the organisms have to withstand in cold environments leads to the adoption of different survival strategies and adaptation of the geological environment. Production of radiation- and desiccation-protective biomolecules is identifiable but the displacement of potentially protective minerals onto the rock surface has also been detected as a protective mechanism against UV-radiation. In this work, Raman spectroscopy is demonstrated as a valuable technique to determine the organic and inorganic compounds used by microorganisms as protective mechanisms against extreme stress conditions. The data from this study will be useful for construction of molecular recognition biomarkers and remote Raman spectral sensing experiments proposed for terrestrial extremophiles in stressed environments. PMID- 15665969 TI - Optical calcium sensors: development of a generic method for their introduction to the cell using conjugated cell penetrating peptides. AB - Probes Encapsulated By Biologically Localised Embedding (PEBBLEs) are optical sensors with nanometer dimensions fabricated by microemulsion polymerisation. The most beneficial characteristic of these sensors is the protection offered by the sensor matrix which decreases interaction between the fluorophore and the cell. These sensors have been introduced to the cell by a number of methods; however this paper discusses the development of a generic method to facilitate inclusion of this type of sensor in the cell by a simple incubation step. This was achieved by covalent linkage of a synthetic Cell Penetrating Peptide (CPP) based on the Human Immuno-deficiency Virus (HIV) -1 Tat, to the external sensor matrix. Calcium sensors were used to demonstrate this approach to incorporate the sensors within the cell. Characterisation revealed the calcium sensors were approximately 30 +/- 7 nm in diameter with a slightly negative zeta potential. The sensors demonstrated a linear range of 0-50 microM with negligible interference from a range of cellular ions and protein. Leaching of entrapped dyes from the calcium sensors was determined as 3% in a 24 h period, while photobleaching of the entrapped dye was minimal over a 40 min period. The sensors ability to cross the cell membrane using the covalently attached synthetic Tat peptide is demonstrated. Cellular inclusion of the sensors occurred within a 30 min incubation period. PMID- 15665970 TI - Application of the lag-after-pulsed-separation (LAPS) flow meter to different protein solutions. AB - A lag after pulsed separation (LAPS) meter was previously developed to measure flow rates of protein solutions. The LAPS meter operates on the time-of-flight principle. An upstream event (electrophoretic concentration of the particles in one section of the device) is detected downstream (by change in ac resistance). The time lag between the event and its detection is inversely proportional to the fluid flow rate. We demonstrate the ability of the LAPS meter to measure the flow rate of solutions containing one or more charged biomacromolecules or particles. A prototype of the LAPS meter was used to measure flow rates of solutions of model proteins [bovine serum albumin (BSA), lysozyme and hemoglobin] and mixtures of BSA and lysozyme. Flow rates of 10-50 microl min(-1)(average velocities of 0.24-1.2 mm s(-1)) were measured. When a single ac measurement was used, the results were solution-dependent, which we attribute to the interface between the protein solution and the ac electrodes. A differential mode, in which the signal from a positive and a negative dc pulse were subtracted from each other, eliminated interfacial effects and led to a single universal (solution independent) calibration curve. The LAPS meter can be used as a non-invasive, no moving-parts flow sensor in any microfluidic system (such as drug delivery devices or micro-reactor arrays) where one needs to measure the flow rate of a solution or a suspension containing charged species such as proteins or cells. PMID- 15665971 TI - Imprinted polymers for chiral resolution of (+/-)-ephedrine: understanding the pre-polymerisation equilibrium and the action of different mobile phase modifiers. AB - A thorough study has been made of the ephedrine-methacrylic acid (MAA) system for molecular imprinting, involving NMR studies of the pre-polymerisation equilibria, modelling and HPLC enantioseparations with different mobile phases. When dimerisation of MAA is accounted for, NMR titrations demonstrate there is a very strong ('stoichiometric', K approximately 10000 M(-1)) interaction between ephedrine and a single MAA monomer. Polymers prepared with a 1:1 monomer:template ratio are capable of enantioseparation, indicating, in combination with the NMR results, that the 1:1 interaction probably involves the carboxylic acid acting as a chelating monomer, forming hydrogen bonds to both the template amine and hydroxyl moieties. Higher monomer:template ratios cause further changes in the NMR signals, suggesting at least one further MAA can interact with the amine group, with a weaker association constant (K approximately 80 M(-1)). Polymers prepared with a 4:1 monomer:template ratio are thus proposed to contain a mixture of 1-monomer binding sites and 2-monomer binding sites, the latter being of enhanced acidity. In HPLC, better results are obtained with the 4:1 polymer using acetic acid as a modifier, while better results are obtained for the 1:1 polymer using butylamine as a modifier. We propose a model whereby acetic acid exerts its effect by reducing binding to the 1-monomer sites, while butylamine works largely through blocking the most acidic, 2-monomer sites. For preparative chromatography we suggest the 1:1 polymer, with a larger population of weaker but more uniform binding sites, is most promising. PMID- 15665972 TI - Determination of phthalates in water using fiber introduction mass spectrometry. AB - Fiber introduction mass spectrometry (FIMS)-a direct coupling of SPME and MS using selective ion monitoring (SIM) was used to detect and quantify dimethylphthalate (DMP), diethylphthalate (DEP) and dipropylphthalate (DPP) in mineral water. In FIMS, a chromatographic silicone septum is the only barrier between ambient and the high-vacuum mass spectrometer, permitting direct introduction of the SPME fiber into the ionization region of the equipment. After their thermal desorption and ionization and dissociation, the extracted phthalates are detected and quantitated by MS. Three types of SPME fibers were screened for best analyte sorption/desorption behaviors: 100 microm polydimethylsiloxane (PDMS), 65 microm polydimethylsiloxane/divinylbenzene (PDMS/DVB) and 65 microm Carbowax/divinylbenzene (CW/DVB). The PDMS/DVB and CW/DVB fibers were then evaluated for precision, and quantitative figures of merit were assessed for extractions using the PDMS/DVB fiber, which displayed the best overall performance. FIMS with the PDMS/DVB fiber allows simple extraction and MS detection and quantitation of DMP in water with good linearity and precision, and at concentrations as low as 3.6 microg L(-1). The LD and LQ of FIMS are below the maximum phthalate concentration allowed by the USEPA for drinking water (6 microg L(-1)). PMID- 15665973 TI - Capillary electrophoresis coupled to mass spectrometry from a polymer modified poly(dimethylsiloxane) microchip with an integrated graphite electrospray tip. AB - Hybrid capillary-poly(dimethysiloxane)(PDMS) microchips with integrated electrospray ionization (ESI) tips were directly fabricated by casting PDMS in a mould. The shapes of the emitter tips were drilled into the mould, which produced highly reproducible three-dimensional tips. Due to the fabrication method of the microfluidic devices, no sealing was necessary and it was possible to produce a perfect channel modified by PolyE-323, an aliphatic polyamine coating agent. A variety of different coating procedures were also evaluated for the outside of the emitter tip. Dusting graphite on a thin unpolymerised PDMS layer followed by polymerisation was proven to be the most suitable procedure. The emitter tips showed excellent electrochemical properties and durabilities. The coating of the emitter was eventually passivated, but not lost, and could be regenerated by electrochemical means. The excellent electrochemical stability was further confirmed in long term electrospray experiments, in which the emitter sprayed continuously for more than 180 h. The PolyE-323 was found suitable for systems that integrate rigid fused silica and soft PDMS technology, since it simply could be applied successfully to both materials. The spray stability was confirmed from the recording of a total ion chromatogram in which the electrospray current exhibited a relative standard deviation of 3.9% for a 30 min run. CE-ESI-MS separations of peptides were carried out within 2 min using the hybrid PDMS chip resulting in similar efficiencies as for fused silica capillaries of the same length and thus with no measurable band broadening effects, originating from the PDMS emitter. PMID- 15665974 TI - A backscattering light detection assembly for sensitive determination of analyte concentrated at the liquid/liquid interface using the interaction of quercetin with proteins as the model system. AB - We report on the construction of a backscattering light (BSL) detection assembly based on detecting angle-dependent light scattering signals, by changing the sample chamber of a common spectrofluorometer. The BSL detection assembly was used to detect, with high sensitivity, the analyte concentrated at the liquid/liquid interface. We applied this assembly to study the interaction of proteins with quercetin in the presence of cationic surfactant. The species resulting from the interaction of quercetin with proteins, when concentrated at the H2O/CCl4 interface, generate enhanced BSL signals characterized at 376.0 nm which were found to be proportional to human serum albumin (HSA) and bovine serum albumin (BSA) in the range of 1-1250 ng mL(-1) and 2-1250 ng mL(-1), respectively. Limits of determination (3sigma) of 75 and 180 pg mL(-1) are reported for the two proteins. PMID- 15665975 TI - Sol-gel derived nitric oxide-releasing oxygen sensors. AB - An amperometric sol-gel derived sensor that both releases nitric oxide (NO) and measures physiologically relevant concentrations of oxygen (PO2) is described. The sensor consists of a platinum electrode coated with an aminosilane/ethyltrimethoxysilane hybrid xerogel film. Hydrophilic polyurethane (HPU) is doped into the hybrid film to reduce sensor hydration time and increase oxygen permeability. Diazeniumdiolate NO donors are formed within the polymer matrix by exposing the cured film to high pressures of NO. These coatings release up to 7.2 pmol s(-1) cm(-2) of NO over the first 12 h and maintain detectable levels of NO release through 48 h. Sensors modified with HPU-doped, NO-releasing xerogels exhibit a linear response to O2 within 30 min of polarization at -0.65 V vs. Ag/AgCl, and have a sensitivity of approximately 6 nA/mmHg O2. The xerogel coating is stable in buffer solution with minimal fragmentation over 48 h. In vitro biocompatibility studies indicate that these materials effectively reduce platelet adhesion. PMID- 15665976 TI - Surface properties and electromagnetic excitation of a piezoelectric gallium phosphate biosensor. AB - The surface properties of GaPO4 have been studied by secondary ion mass spectrometry, X-ray photoelectron spectroscopy and electromagnetic acoustic wave excitation in order to explore the potential of this relatively new piezoelectric material as a biosensor. The X-ray photoelectron spectrum of the substrate shows a Ga-rich surface (Ga:P = 1.4), while the negative secondary ion mass spectrum is similar to that of other phosphates, with PO3- and PO2- being the main fragments derived from the substrate. Surface analysis reveals that the linker protein for biotinylated moieties, neutravidin, is both readily chemisorbed to bare gallium phosphate at pH 7.5 and attached to p-hydroxy benzaldehyde-treated devices, establishing the possibility to exploit the surface chemistry of the phosphate for the fabrication of an electrode-free acoustic wave biosensor. Preliminary results regarding the detection of the adsorption of neutravidin with an electromagnetic field-excited GaPO4 device incorporated in a FIA configuration showed comparable results with those obtained with a quartz-sensor equivalent. The frequency shift for the adsorbed protein layer at the device fundamental frequency was 200 Hz and the noise was routinely around 13 Hz. The possibility to use the electrodeless acoustic GaPO4 device at higher harmonics in the liquid phase has also been confirmed. PMID- 15665977 TI - Cobalt oxide/tetraruthenated cobalt-porphyrin composite for hydrogen peroxide amperometric sensors. AB - A new composite material constituted by mu-{5,10,15,20-tetra(4 pyridyl)porphyrinato cobalt(iii)}-tetrakis-{chloro-bis-(2,2' bipyridine)ruthenium(ii)} complex (or CoTRP) and cobalt oxide, exhibiting high stability and sensitivity for the quantification of hydrogen peroxide, was obtained by the electrochemical polymerization of the tetraruthenated cobalt porphyrin in alkaline medium. The optimized experimental conditions for the preparation of the modified glassy carbon electrodes and for analysis of H2O2 were carefully determined. Fast sequential analysis (120 determinations h(-1)) in a wide linear dynamic range (5.0 x 10(-7) mol L(-1) to 2.0 x 10(-3) mol L(-1)), with high sensitivity and low detection limit (2.0 x 10(-7) mol L(-1)), was achieved by using these electrodes and the batch injection analysis (BIA) technique. Such characteristics allied to a good stability were explored for the specific determination of hydrogen peroxide in six commercial cosmetics and pharmaceutical product samples, giving results in excellent agreement with those obtained by the spectrophotometric method. PMID- 15665978 TI - Determination of chemical oxygen demand in fresh waters using flow injection with on-line UV-photocatalytic oxidation and spectrophotometric detection. AB - A flow injection manifold incorporating UV-photocatalytic oxidation for the determination of chemical oxygen demand (COD) in freshwater is reported. The method utilises the UV-photocatalytic oxidation of organic compounds instead of conventional heating (used in the standard method), with acidified potassium permanganate as the oxidant. Sodium oxalate, d-glucose and potassium hydrogen phthalate were used as COD standards. A 100 microL sample solution was injected into a 0.3 mol L(-1) H2SO4 carrier stream containing 0.1 mol L(-1) (NH4)2SO4, merged with a permanganate solution (8 x 10(-4) mol L(-1)) and passed through a 250 cm FEP (fluoroethylene polymer) photo-reaction coil wound around a 15 W UV lamp. The sample throughput was 30 h(-1), with an LOD (blank plus 3sigma) of 0.5 mg COD L(-1) and a linear range of 0.5-50 mg COD L(-1) (D-glucose, r2 = 0.9966). The method had good precision with relative standard deviations of 2.7% at 5 mg COD L(-1) and 1.2% at 20 mg COD L(-1) (n = 12) for glucose. Results for a COD certified reference material (QC Demand Quality Control Standard) were in good agreement with the certified COD value. Recovery from Tamar River water samples for all three COD standards was 83.0-111.0% and the COD values determined were in good agreement with those of a permanganate index reference method. PMID- 15665979 TI - Determination of the enantiomeric composition of phenylalanine samples by chemometric analysis of the fluorescence spectra of cyclodextrin guest-host complexes. AB - A novel strategy for determining the enantiomeric composition of phenylalanine samples that combines ordinary fluorescence spectroscopy, guest-host cyclodextrin chemistry, and multivariate regression modeling is investigated. Partial-least squares regression (PLS-1) models were developed from fluorescence spectral data obtained with a series of samples containing cyclodextrin guest-host complexes of phenylalanine with different known enantiomeric compositions. The regression models were subsequently validated by determining the enantiomeric composition of a set of independently prepared phenylalanine samples. The ability of the models to correctly predict the enantiomeric compositions of future samples was evaluated in terms of the root-mean-square percent relative error (RMS%RE). The RMS%RE in the mol fraction of D-phenylalanine ranged from 1.3% to 3.0% when beta cyclodextrin was used as the host molecule for different guest-host concentrations. The RMS%RE in the mol fraction of D-phenylalanine obtained in a similar validation study conducted with gamma-cyclodextrin ranged between 1.8% and 4.0% for different guest-host concentrations. Compared with previous studies done in absorption, fluorescence data were found to be more sensitive and the spectral differences observed as a function of enantiomeric composition were more uniformly spaced, making regression modeling more reliable. As a result, good regression models could be made at lower concentrations than were possible previously when absorption measurements were used. PMID- 15665980 TI - Determination of environmentally important metal ions by fluorescence quenching in anionic micellar solution. AB - This work describes the effect of a variety of metal ions as quenchers of the fluorescence of naphthalene, in aqueous micellar solutions of sodium dodecyl sulfate (SDS). The quenching by the metal ions can be adequately described by the Stern-Volmer equation and the best signal to noise ratios are obtained with low micellized detergent concentrations. Apparent Stern-Volmer constants decrease in the order: Fe3+ > Cu2+ > Pb2+ > Cr3+ > Ni2+ and directly reflect the relative sensitivity of the method for these ions. Detection limits (defined as three times the standard deviation of the blank for n= 10) for the fluorescence quenching of naphthalene by the metal ions in aqueous micellar SDS are in the range of 1.0 x 10(-6) to 1.0 x 10(-5) mol dm(-3). The proposed fluorescence quenching method shows good repeatibility for a variety of added quencher metal ions, indicating that anionic micelle-enhanced fluorescence quenching by metal ions constitutes an analytical method of rather general application. PMID- 15665981 TI - L1 sequence of a new human papillomavirus type-58 variant associated with cervical intraepithelial neoplasia. AB - The present study on molecular characterization of a human papillomavirus (HPV) isolated in Central Brazil describes the L1 gene sequence from a new variant of HPV-58, the isolate Bsb-02. The sample was from a smear obtained from a woman with cervical intraepithelial neoplasia grade II. The whole L1 gene from isolate Bsb-02 was sequenced automatically, showing 99.1% nucleotide identity with the gene from the HPV-58 reference. The clustering between Bsb-02 and HPV-58 reference sequence was also supported by phylogenetic analysis. Fourteen nucleotide substitutions were observed: eight were synonymous and six were associated with amino acid substitutions. A10V and V144I have not been previously described. At GenBank, the only complete L1 sequence from HPV-58 in addition to the HPV-58 reference one is that of Bsb-02. These data provide information that may be relevant to HPV diagnosis and to rational vaccine strategies. HPV variants may also be associated with host immune responses and with the risk of cervical neoplasia. PMID- 15665982 TI - Activation of a P2Y4-like purinoceptor triggers an increase in cytosolic [Ca2+] in the red blood cells of the lizard Ameiva ameiva (Squamata, Teiidae). AB - An increasing number of pathophysiological roles for purinoceptors are emerging, some of which have therapeutic potential. Erythrocytes are an important source of purines, which can be released under physiological and physiopathological conditions, acting on purinergic receptors associated with the same cell or with neighboring cells. Few studies have been conducted on lizards, and have been limited to ATP agonist itself. We have previously shown that the red blood cells (RBCs) of the lizard Ameiva ameiva store Ca2+ in the endoplasmic reticulum (ER) and that the purinergic agonist ATP triggers a rapid and transient increase of [Ca2+]c by mobilization of the cation from internal stores. We also reported the ability of the second messenger IP3 to discharge the ER calcium pool of the ER. Here we characterize the purinoceptor present in the cytoplasmic membrane of the RBCs of the lizard Ameiva ameiva by the selective use of ATP analogues and pyrimidine nucleotides. The nucleotides UTP, UDP, GTP, and ATPgammaS triggered a dose-dependent response, while interestingly 2MeSATP, 2ClATP, alpha, ss-ATP, and ADP failed to do so in a 1- to 200-microm con- centration. The EC50 obtained for the compounds tested was 41.77 microM for UTP, 48.11 microM for GTP, 53.11 microM for UDP, and 30.78 microM for ATPgammaS. The present data indicate that the receptor within the RBCs of Ameiva ameiva is a P2Y4-like receptor due to its pharmacological similarity to the mammalian P2Y4 receptor. PMID- 15665983 TI - Prevalence of deltaF508, G551D, G542X, and R553X mutations among cystic fibrosis patients in the North of Brazil. AB - Cystic fibrosis (CF) is the most common genetic disease among Caucasians and is rare among sub-Saharan Africans. The Brazilian population is not ethnically homogeneous but it is the result of three-way ethnic admixture of Europeans, Africans and Amerindians in varying proportions, depending on the region. In the present study, we investigated 33 patients who had been diagnosed and are currently under treatment for CF at the University Hospital Joao de Barros Barreto, Belem, Para State. The molecular analysis for G542X, G551D and R553X mutations was performed by PCR followed by RFLP using BstNI, HincII and MboI, respectively, in polyacrylamide gel eletrophoresis and stained with AgNO3. ThedeltaF508 mutation (a deletion of 3 bp) was only analyzed by polyacrylamide gel electrophoresis and stained with AgNO3. Each sample was analyzed for regions of interest in the CFTR gene using amplified by PCR and specific primers. The deltaF508 and G551D mutations presented frequencies of 22.7 and 3%, respectively. In 74.3% of the remaining patients, none of the mutations investigated was found. The present study characterized in a sample of patients with an established clinical diagnosis of CF (asthma, repeated bronchopneumonia, disorders of nutritional status, etc.) the most frequent mutation (deltaF508) in the North region of Brazil and is also the first report of the G551D mutation. In spite of the wide spectrum of CF mutations and the heterogeneous ethnic origin of the Amazon population, the molecular diagnosis is a helpful additional tool for the diagnosis and treatment of CF patients. PMID- 15665985 TI - Expression of dorsal-ventral genes during early development of Rhynchosciara americana embryos. AB - The establishment of dorsal-ventral polarity in Drosophila is a complex process which involves the action of maternal and zygotically expressed genes. Interspecific differences in the expression pattern of some of these genes have been described in other species. Here we present the expression of dorsal-ventral genes during early embryogenesis in the lower dipteran Rhynchosciara americana. The expression of four genes, the ventralizing genes snail (sna) and twist (twi) and the dorsalizing genes decapentaplegic (dpp) and zerknullt (zen), was investigated by whole-mount in situ hybridization. Sense and antisense mRNA were transcribed in vitro using UTP-digoxigenin and hybridized at 55 degrees C with dechorionated fixed embryos. Staining was obtained with anti-digoxigenin alkaline phosphatase-conjugated antibody revealed with NBT-BCIP solution. The results showed that, in general, the spatial-temporal expression of R. americana dorsal ventral genes is similar to that observed in Drosophila, where twi and sna are restricted to the ventral region, while dpp and zen are expressed in the dorsal side. The differences encountered were subtle and probably represent a particular aspect of dorsal-ventral axis determination in R. americana. In this lower dipteran sna is expressed slightly later than twi and dpp expression is expanded over the lateral ectoderm during cellular blastoderm stage. These data suggest that the establishment of dorsal-ventral polarity in R. americana embryos follows a program similar to that observed in Drosophila melanogaster. PMID- 15665984 TI - Mutations in SRY and WT1 genes required for gonadal development are not responsible for XY partial gonadal dysgenesis. AB - The WT1 transcription factor regulates SRY expression during the initial steps of the sex determination process in humans, activating a gene cascade leading to testis differentiation. In addition to causing Wilms' tumor, mutations in WT1 are often responsible for urogenital defects in men, while SRY mutations are mainly related to 46,XY pure gonadal dysgenesis. In order to evaluate their role in abnormal testicular organogenesis, we screened for SRY and WT1 gene mutations in 10 children with XY partial gonadal dysgenesis, 2 of whom with a history of Wilms' tumor. The open reading frame and 360 bp of the 5' flanking sequence of the SRY gene, and the ten exons and intron boundaries of the WT1 gene were amplified by PCR of genomic DNA. Single-strand conformation polymorphism was initially used for WT1 mutation screening. Since shifts in fragment migration were only observed for intron/exon 4, the ten WT1 exons from all patients were sequenced manually. No mutations were detected in the SRY 5' untranslated region or within SRY open-reading frame sequences. WT1 sequencing revealed one missense mutation (D396N) in the ninth exon of a patient who also had Wilms' tumor. In addition, two silent point mutations were found in the first exon including one described here for the first time. Some non-coding sequence variations were detected, representing one new (IVS4+85A>G) and two already described (-7ATG T>G, IVS9-49 T>C) single nucleotide polymorphisms. Therefore, mutations in two major genes required for gonadal development, SRY and WT1, are not responsible for XY partial gonadal dysgenesis. PMID- 15665986 TI - Skin morphology of the mutant hairless USP mouse. AB - The morphology of the skin of the mutant hairless USP mouse was studied by histological, histochemical and immunohistochemical methods and compared to the skin of BALB/c mice. Representative sections of the dorsal skin from mice of both strains aged 18 days, and 1, 3, 6, and 8 months were studied. Sections stained with hematoxylin and eosin showed cystic formations called utricles and dermal cysts in the dermis that increased in size and number during growth. Skin thickness increased significantly at 8 months. Sections stained with picrosirius and examined with polarized light, displayed different colors, suggesting different thicknesses of dermal collagen fibers (probably types I and III). Weigert, Verhoeff and resorcin-fuchsin stains revealed fibers of the elastic system. The PAS and Alcian blue methods revealed neutral and acid glycosaminoglycans in the skin ground substance of both mouse strains. Immunohistochemical staining for fibronectin and laminin did not show differences between the mutant and BALB/c mice. Mast cells stained by the Gomori method and macrophages positive for HAM 56 antibodies were observed in both mouse strains. Except for the presence of enlarged cysts in the hairless strain, no qualitative differences were found during development of the skin of BALB/c and the mutant hairless mice. PMID- 15665987 TI - Geographic distribution of hepatitis C virus genotypes in Brazil. AB - Brazil is a country of continental dimension with a population of different ethnic backgrounds. Thus, a wide variation in the frequencies of hepatitis C virus (HCV) genotypes is expected to occur. To address this point, 1,688 sequential samples from chronic HCV patients were analyzed. HCV-RNA was amplified by the RT-PCR from blood samples collected from 1995 to 2000 at different laboratories located in different cities from all Brazilian States. Samples were collected in tubes containing a gel separator, centrifuged in the site of collection and sent by express mail in a refrigerated container to Laboratorio Bioquimico Jardim Paulista, Sao Paulo, SP, Brazil. HCV-RNA was extracted from serum and submitted to RT and nested PCR using standard procedures. Nested PCR products were submitted to cycle sequencing reactions without prior purification. Sequences were analyzed for genotype determination and the following frequencies were found: 64.9% (1,095) for genotype 1, 4.6% (78) for genotype 2, 30.2% (510) for genotype 3, 0.2% (3) for genotype 4, and 0.1% (2) for genotype 5. The frequencies of HCV genotypes were statistically different among Brazilian regions (P = 0.00017). In all regions, genotype 1 was the most frequent (51.7 to 74.1%), reaching the highest value in the North; genotype 2 was more prevalent in the Center-West region (11.4%), especially in Mato Grosso State (25.8%), while genotype 3 was more common in the South (43.2%). Genotypes 4 and 5 were rarely found and only in the Southeast, in Sao Paulo State. The present data indicate the need for careful epidemiological surveys throughout Brazil since knowing the frequency and distribution of the genotypes would provide key information for understanding the spread of HCV. PMID- 15665988 TI - Th-1 and Th-2 cytokine production in infants with virus-associated wheezing. AB - Wheezing associated with respiratory viral infections in infancy is very common and results in high morbidity worldwide. The Th1/Th2 pattern of immune response in these patients remains unclear and previous studies have shown controversial results. The aim of the present study was to compare the type of Th1/Th2 cytokine response between infants with acute bronchiolitis, recurrent wheezing and upper respiratory infections from a developing country. Infants younger than 2 years of age admitted to Hospital Sao Lucas, Porto Alegre, RS, Brazil, between May and November 2001, with an acute episode of wheezing associated with viral respiratory infection were selected. Subjects with upper respiratory infections from the emergency department were selected for the control group. Interferon gamma (IFN-gamma) and interleukin-4 (IL-4) levels from nasal aspirates were determined by ELISA from peripheral mononuclear cell cultures. Twenty-nine subjects with acute bronchiolitis, 18 with recurrent wheezing and 15 with upper respiratory infections were enrolled. There were no differences in family history of atopy or parental smoking between groups. Oxygen requirement was similar for the acute bronchiolitis and recurrent wheezing groups. The percentage of positive tests for the cytokines studied and the IFN-gamma/IL-4 ratio was similar for all groups. Comparison of the polarized Th1/Th2 cytokine results for the various groups showed no specific pattern of cytokine production. Infants with wheezing from a developing country do not show any specific predominant pattern of Th1/Th2 cytokine production, suggesting that multiple factors may be involved in the pathogenesis of this illness. PMID- 15665989 TI - Differential role of entorhinal and hippocampal nerve growth factor in short- and long-term memory modulation. AB - We studied the effects of infusion of nerve growth factor (NGF) into the hippocampus and entorhinal cortex of male Wistar rats (250-300 g, N = 11-13 per group) on inhibitory avoidance retention. In order to evaluate the modulation of entorhinal and hippocampal NGF in short- and long-term memory, animals were implanted with cannulae in the CA1 area of the dorsal hippocampus or entorhinal cortex and trained in one-trial step-down inhibitory avoidance (foot shock, 0.4 mA). Retention tests were carried out 1.5 h or 24 h after training to measure short- and long-term memory, respectively. Immediately after training, rats received 5 microl NGF (0.05, 0.5 or 5.0 ng) or saline per side into the CA1 area and entorhinal cortex. The correct position of the cannulae was confirmed by histological analysis. The highest dose of NGF (5.0 ng) into the hippocampus blocked short-term memory (P < 0.05), whereas the doses of 0.5 (P < 0.05) and 5.0 ng (P < 0.01) NGF enhanced long-term memory. NGF administration into the entorhinal cortex improved long-term memory at the dose of 5.0 ng (P < 0.05) and did not alter short-term memory. Taken as a whole, our results suggest a differential modulation by entorhinal and hippocampal NGF of short- and long-term memory. PMID- 15665990 TI - Changes of ongoing activity in Cebus monkey perirhinal cortex correlate with behavioral performance. AB - A Cebus apella monkey weighing 4 kg was trained in a saccadic eye movement task and while the animal performed the task we recorded the extracellular activity of perirhinal cortical neurons. Although the task was very simple and maintained at a constant level of difficulty, we observed considerable changes in the performance of the monkey within each experimental session. The behavioral states responsible for such variation may be related to arousal, motivation or attention of the animal while engaged in the task. In approximately 20% (16/82) of the units recorded, long-term direct or inverse correlations could be demonstrated between the monkey's behavioral state and the cells' ongoing activity (independent of the visual stimulation or of the specific behavior along a trial). The perirhinal cortex and other medial temporal structures have long been associated with normal memory function. The data presented here were interpreted in terms of recent reports focusing on the subcortical afferents to temporal lobe structures and their possible role in controlling arousal, motivation, or attention. PMID- 15665991 TI - Fecal androgen levels in common marmoset (Callithrix jacchus) males living in captive family groups. AB - In captive common marmoset groups, the reproductive inhibition observed in subordinate female seems to be a result of olfactory, visual and behavioral cues from the dominant female. However, few studies have examined the relationship among adult males living in the same social group. These studies have shown that reproductive failure among peer males seems to be based on hormonal and behavioral mechanisms. New insights on sexual strategies in primates have been shown using fecal steroids, but so far no information is available for common marmoset males. In the present study, we evaluated the influence of light-dark cycle, age and reproductive condition on the profile of fecal androgens in males living in the same family group. Feces were collected from six fathers and six sons for androgen determination during the light phase of the 24-h cycle for eight days randomly distributed over a 4-week period. Androgen levels were determined by enzyme immunoassay technique. Adult sons showed higher androgen levels (166.97 +/- 22.95 ng/g) than fathers (80.69 +/- 44.38 ng/g) and juveniles (49.06 +/- 23.15 ng/g; P < 0.05). No diurnal variation (P > 0.05) in fecal androgen profile was observed in adults or juveniles. No indication of androgen mediated social competition between fathers and adult sons was demonstrable. These results provide basic information on fecal androgen profile useful to investigate the socioendocrinology of free-ranging common marmoset males and verify that, in contrast to daughters, the reproductive suppression of sons is not based on physiological inhibition of their gonads. PMID- 15665992 TI - The terrestrial Gastropoda Megalobulimus abbreviatus as a useful model for nociceptive experiments: effects of morphine and naloxone on thermal avoidance behavior. AB - We describe the behavior of the snail Megalobulimus abbreviatus upon receiving thermal stimuli and the effects of pretreatment with morphine and naloxone on behavior after a thermal stimulus, in order to establish a useful model for nociceptive experiments. Snails submitted to non-functional (22 degrees C) and non-thermal hot-plate stress (30 degrees C) only displayed exploratory behavior. However, the animals submitted to a thermal stimulus (50 degrees C) displayed biphasic avoidance behavior. Latency was measured from the time the animal was placed on the hot plate to the time when the animal lifted the head-foot complex 1 cm from the substrate, indicating aversive thermal behavior. Other animals were pretreated with morphine (5, 10, 20 mg/kg) or naloxone (2.5, 5.0, 7.5 mg/kg) 15 min prior to receiving a thermal stimulus (50 degrees C; N = 9 in each group). The results (means +/- SD) showed an extremely significant difference in response latency between the group treated with 20 mg/kg morphine (63.18 +/- 14.47 s) and the other experimental groups (P < 0.001). With 2.5 mg/kg (16.26 +/- 3.19 s), 5.0 mg/kg (11.53 +/- 1.64 s) and 7.5 mg/kg naloxone (7.38 +/- 1.6 s), there was a significant, not dose-dependent decrease in latency compared to the control (33.44 +/- 8.53 s) and saline groups (29.1 +/- 9.91 s). No statistically significant difference was found between the naloxone-treated groups. With naloxone plus morphine, there was a significant decrease in latency when compared to all other groups (minimum 64% in the saline group and maximum 83.2% decrease in the morphine group). These results provide evidence of the involvement of endogenous opioid peptides in the control of thermal withdrawal behavior in this snail, and reveal a stereotyped and reproducible avoidance behavior for this snail species, which could be studied in other pharmacological and neurophysiological studies. PMID- 15665993 TI - Long-term ethanol intoxication reduces inflammatory responses in rats. AB - The anti-inflammatory effects of long-term ethanol intoxication were determined during ethanol treatment and withdrawal on the basis of neutrophil and eosinophil migration, hind paw edema and mast cell degranulation. Male Wistar rats (180-200 g, around 2 months of age) were exposed to increasing concentrations of ethanol vapor over a 10-day period. One group was evaluated immediately after exposure (treated group - intoxicated), and another was studied 7 h later (withdrawal group). Ethanol inhalation treatment significantly inhibited carrageenan--(62% for the intoxicated group, N = 5, and 35% for the withdrawal group, N = 6) and dextran-induced paw edema (32% for intoxicated rats and 26% for withdrawal rats, N = 5 per group). Ethanol inhalation significantly reduced carrageenan-induced neutrophil migration (95% for intoxicated rats and 41% for withdrawn rats, N = 6 per group) into a subcutaneous 6-day-old air pouch, and Sephadex-induced eosinophil migration to the rat peritoneal cavity (100% for intoxicated rats and 64% for withdrawn rats, N = 6 per group). A significant decrease of mast cell degranulation was also demonstrated (control, 82%; intoxicated, 49%; withdrawn, 51%, N = 6, 6 and 8, respectively). Total leukocyte and neutrophil counts in venous blood increased significantly during the 10 days of ethanol inhalation (leukocytes, 13, 27 and 40%; neutrophils, 42, 238 and 252%, respectively, on days 5, 9 and 10, N = 7, 6 and 6). The cell counts decreased during withdrawal, but were still significantly elevated (leukocytes, 10%; neutrophils, 246%, N = 6). These findings indicate that both the cellular and vascular components of the inflammatory response are compromised by long-term ethanol intoxication and remain reduced during the withdrawal period. PMID- 15665994 TI - Participation of ATP-sensitive K+ channels in the peripheral antinociceptive effect of fentanyl in rats. AB - We examined the effect of several K+ channel blockers such as glibenclamide, tolbutamide, charybdotoxin (ChTX), apamin, tetraethylammonium chloride (TEA), 4 aminopyridine (4-AP), and cesium on the ability of fentanyl, a clinically used selective micro-opioid receptor agonist, to promote peripheral antinociception. Antinociception was measured by the paw pressure test in male Wistar rats weighing 180-250 g (N = 5 animals per group). Carrageenan (250 microg/paw) decreased the threshold of responsiveness to noxious pressure (delta = 188.1 +/- 5.3 g). This mechanical hyperalgesia was reduced by fentanyl (0.5, 1.5 and 3 microg/paw) in a peripherally mediated and dose-dependent fashion (17.3, 45.3 and 62.6%, respectively). The selective blockers of ATP-sensitive K+ channels glibenclamide (40, 80 and 160 microg/paw) and tolbutamide (80, 160 and 240 microg/paw) dose dependently antagonized the antinociception induced by fentanyl (1.5 microg/paw). In contrast, the effect of fentanyl was unaffected by the large conductance Ca2+-activated K+ channel blocker ChTX (2 microg/paw), the small conductance Ca2+-activated K+ channel blocker apamin (10 microg/paw), or the non specific K+ channel blocker TEA (150 microg/paw), 4-AP (50 microg/paw), and cesium (250 microg/paw). These results extend previously reported data on the peripheral analgesic effect of morphine and fentanyl, suggesting for the first time that the peripheral micro-opioid receptor-mediated antinociceptive effect of fentanyl depends on activation of ATP-sensitive, but not other, K+ channels. PMID- 15665995 TI - Effect of the GABAB agonist baclofen on dipyrone-induced delayed gastric emptying in rats. AB - Dipyrone administered intravenously (iv) or intracerebroventricularly (icv) delays gastric emptying (GE) in rats. Gamma-aminobutyric acid (GABA) is the most potent inhibitory neurotransmitter of the central nervous system. The objective of the present study was to determine the effect of icv baclofen, a GABAB receptor agonist, on delayed GE induced by dipyrone. Adult male Wistar rats received a saline test meal containing phenol red as a marker. GE was indirectly evaluated by determining the percent of gastric retention (%GR) of the meal 10 min after orogastric administration. In the first experiment, the animals were injected iv with vehicle (Civ) or 80 mg/kg (240 micromol/kg) dipyrone (Dpiv), followed by icv injection of 10 microl vehicle (bac0), or 0.5 (bac0.5), 1 (bac1) or 2 microg (bac2) baclofen. In the second experiment, the animals were injected icv with 5 microl vehicle (Cicv) or an equal volume of a solution containing 4 micromol (1333.2 microg) dipyrone (Dpicv), followed by 5 microl vehicle (bac0) or 1 microg baclofen (bac1). GE was determined 10 min after icv injection. There was no significant difference between control animals from one experiment to another concerning GR values. Baclofen at the doses of 1 and 2 microg significantly reduced mean %GR induced by iv dipyrone (Dpivbac1 = 35.9% and Dpivbac2 = 26.9% vs Dpivbac0 = 51.8%). Similarly, baclofen significantly reduced the effect of dipyrone injected icv (mean %GR: Dpicvbac1 = 30.4% vs Dpicvbac0 = 54.2%). The present results suggest that dipyrone induces delayed GE through a route in the central nervous system that is blocked by the activation of GABAB receptors. PMID- 15665996 TI - Electrophysiological evidence for the presence of NR2C subunits of N-methyl-D aspartate receptors in rat neurons of the nucleus tractus solitarius. AB - The nucleus tractus solitarius (NTS) plays an important role in the control of autonomic reflex functions. Glutamate, acting on N-methyl-D-aspartate (NMDA) and non-NMDA ionotropic receptors, is the major neurotransmitter in this nucleus, and the relative contribution of each receptor to signal transmission is unclear. We have examined NMDA excitatory postsynaptic currents (NMDA-EPSCs) in the subpostremal NTS using the whole cell patch clamp technique on a transverse brainstem slice preparation. The NMDA-EPSCs were evoked by stimulation of the solitary tract over a range of membrane potentials. The NMDA-EPSCs, isolated pharmacologically, presented the characteristic outward rectification and were completely blocked by 50 microM DL-2-amino-5-phosphonopentanoic acid. The I-V relationship of the NMDA response shows that current, with a mean (+/- SEM) amplitude of -41.2 +/- 5.5 pA, is present even at a holding potential of -60 mV, suggesting that the NMDA receptors are weakly blocked by extracellular Mg2+ at near resting membrane potentials. This weak block can also be inferred from the value of 0.67 +/- 0.17 for parameter delta obtained from a fit of the Woodhull equation to the I-V relationship. The maximal inward current measured on the I-V relationship was at -38.7 +/- 4.2 mV. The decay phase of the NMDA currents was fitted with one exponential function with a decay time constant of 239 +/- 51 and 418 +/- 80 ms at a holding potential of -60 and +50 mV, respectively, which became slower with depolarization (e-fold per 145 mV). The biophysical properties of the NMDA receptors observed in the present study suggest that these receptors in the NTS contain NR2C subunits and may contribute to the synaptic signal integration. PMID- 15665997 TI - Inotropic effect of Citrus sinensis (L.) Osbeck leaf extracts on the guinea pig atrium. AB - The objective of the present investigation was to determine the contractile effect of crude and acetone leaf extracts of Citrus sinensis (L.) Osb. on mammalian myocardium. Crude leaf extracts have been used in folk medicine to treat neurological disorders. Some flavonoids isolated from this plant presented a positive inotropic effect on myocardium. This motivated us to test the extracts on the atria of guinea pigs of both sexes (300-500 g) and surprisingly we observed inotropic depression instead of an increase in force. The maximum effect of the crude extract was 79.4 +/- 8.1% of the control force amplitude (N = 5 hearts, 10 trials, 27 +/- 0.1 degrees C, stimulus: 2 Hz, 400 V, 0.5 ms). The EC50 for crude, ethanol, acetic, aqueous, and acetone extracts was 300, 300, 600, 1000, and 140 microg/ml, respectively, with a Hill constant of 1.8, 2.0, 2.5, 2.0, and 1.4, respectively. Blockade of cholinergic, beta-adrenergic, or opioid membrane receptors with 1.5 microM atropine sulfate, 1 microM propranolol, and 10 microM naloxone, respectively, did not change the effect of the crude extract. The acetone extract abolished the Bowditch positive staircase phenomenon (N = 5 hearts, 10 trials, 27 +/- 0.1 degrees C), suggesting a possible reduction of the calcium inward current, and also promoted the so-called Woodworth phenomenon. The effect was concentration-dependent and indicated the existence of another inhibitory contractile mechanism such as the simultaneous activation of some of the membrane potassium channels reducing the myocardial action potential duration and further decreasing the cellular calcium entry. PMID- 15665998 TI - Blood flow measurements in rats using four color microspheres during blockade of different vasopressor systems. AB - The use of colored microspheres to adequately evaluate blood flow changes under different circumstances in the same rat has been validated with a maximum of three different colors due to methodological limitations. The aim of the present study was to validate the use of four different colors measuring four repeated blood flow changes in the same rat to assess the role of vasopressor systems in controlling arterial pressure (AP). Red (150,000), white (200,000), yellow (150,000), and blue (200,000) colored microspheres were infused into the left ventricle of 6 male Wistar rats 1) at rest and 2) after vasopressin (aAVP, 10 microg/kg, iv), 3) renin-angiotensin (losartan, 10 mg/kg, iv), and 4) sympathetic system blockade (hexamethonium, 20 mg/kg, iv) to determine blood flow changes. AP was recorded and processed with a data acquisition system (1-kHz sampling frequency). Blood flow changes were quantified by spectrophotometry absorption peaks for colored microsphere components in the tissues evaluated. Administration of aAVP and losartan slightly reduced the AP (-5.7 +/- 0.5 and -7.8 +/- 1.2 mmHg, respectively), while hexamethonium induced a 52 +/- 3 mmHg fall in AP. The aAVP injection increased blood flow in lungs (78%), liver (117%) and skeletal muscle (>150%), while losartan administration enhanced blood flow in heart (126%), lungs (100%), kidneys (80%), and gastrocnemius (75%) and soleus (94%) muscles. Hexamethonium administration reduced only kidney blood flow (50%). In conclusion, four types of colored microspheres can be used to perform four repeated blood flow measurements in the same rat detecting small alterations such as changes in tissues with low blood flow. PMID- 15665999 TI - Evaluation of two experimental models of hepatic encephalopathy in rats. AB - The serious neuropsychological repercussions of hepatic encephalopathy have led to the creation of several experimental models in order to better understand the pathogenesis of the disease. In the present investigation, two possible causes of hepatic encephalopathy, cholestasis and portal hypertension, were chosen to study the behavioral impairments caused by the disease using an object recognition task. This working memory test is based on a paradigm of spontaneous delayed non matching to sample and was performed 60 days after surgery. Male Wistar rats (225 250 g) were divided into three groups: two experimental groups, microsurgical cholestasis (N = 20) and extrahepatic portal hypertension (N = 20), and a control group (N = 20). A mild alteration of the recognition memory occurred in rats with cholestasis compared to control rats and portal hypertensive rats. The latter group showed the poorest performance on the basis of the behavioral indexes tested. In particular, only the control group spent significantly more time exploring novel objects compared to familiar ones (P < 0.001). In addition, the portal hypertension group spent the shortest time exploring both the novel and familiar objects (P < 0.001). These results suggest that the existence of portosystemic collateral circulation per se may be responsible for subclinical encephalopathy. PMID- 15666000 TI - Electrical parameters and water permeability properties of monolayers formed by T84 cells cultured on permeable supports. AB - T84 is an established cell line expressing an enterocyte phenotype whose permeability properties have been widely explored. Osmotic permeability (POSM), hydraulic permeability (PHYDR) and transport-associated net water fluxes (JW transp), as well as short-circuit current (ISC), transepithelial resistance (RT), and potential difference (deltaVT) were measured in T84 monolayers with the following results: POSM 1.3 +/- 0.1 cm.s-1 x 10-3; PHYDR 0.27 +/- 0.02 cm.s-1; RT 2426 +/- 109 omega.cm2, and deltaVT 1.31 +/- 0.38 mV. The effect of 50 microM 5,6 dichloro-1-ethyl-1,3-dihydro-2H-benzimidazol-2-one (DCEBIO), a "net Cl- secretory agent", on T84 cells was also studied. We confirm the reported important increase in ISC induced by DCEBIO which was associated here with a modest secretory deltaJW-transp. The present results were compared with those reported using the same experimental approach applied to established cell lines originating from intestinal and renal epithelial cells (Caco-2, LLC-PK1 and RCCD-1). No clear association between PHYDR and RT could be demonstrated and high PHYDR values were observed in an electrically tight epithelium, supporting the view that a "water leaky" barrier is not necessarily an "electrically leaky" one. Furthermore, the modest secretory deltaJW-transp was not consistent with previous results obtained with RCCD-1 cells stimulated with vasopressin (absorptive fluxes) or with T84 cells secreting water under the action of Escherichia coli heat stable enterotoxin. We conclude that, while the presence of aquaporins is necessary to dissipate an external osmotic gradient, coupling between water and ion transport cannot be explained by a simple and common underlying mechanism. PMID- 15666001 TI - [Cognitive preservation: an agenda for the 21th Century]. PMID- 15666009 TI - [Late-onset male hypogonadism or andropause]. PMID- 15666010 TI - [Noncompaction of the myocardium]. PMID- 15666012 TI - [Ureteral colic and emotional stress]. PMID- 15666013 TI - [Lung volume reduction surgery]. PMID- 15666014 TI - [Febrile neutropenia in patients with breast cancer submitted to chemotherapy: a 12-year experience]. AB - PURPOSE: To identify the characteristics of patients with breast cancer who developed febrile neutropenia and to establish risk factors for its incidence and parameters for an unfavorable evolution. PATIENTS AND METHODS: A case-control study was performed and included 65 patients: 13 patients presented febrile neutropenia and four controls per case matched by: date and number of previous chemotherapy cycles, drugs and doses used. The clinical and laboratory data were obtained from medical records. We utilized the odds ratio (OR) and the 95% confidence interval (CI) to estimate the significance of risk factors. RESULTS: We identified two risk factors associated to occurrence of febrile neutropenia: use of chemotherapy within the first 24 hours post surgery (OR: 159.9 95% CI: 9.5 to 2699), and the concomitant use of chemotherapy and breast radiotherapy (OR: 108.3 95%CI: 4.9 to 2391). We did not observe a significant difference between cases and controls regarding age, body mass index, neutrophils and monocytes count prior to chemotherapy. Three patients died (23.1%), two of them were more than 60 years old, had no comorbidities, had received the first CMF within the first post surgery day and had surgical site infection. CONCLUSIONS: The main risk factors associated with febrile neutropenia in patients with breast cancer were: chemotherapy within the first 24 hours post surgery, and concomitant chemotherapy and breast radiotherapy. As such, our study shows that these situations must be avoided. PMID- 15666015 TI - [Health and aging study: prevalence of chronic joint symptoms among the elderly in Bambui]. AB - BACKGROUND: Arthritis is one of the more prevalent chronic conditions and a leading cause of disability in the elderly. The objective of this study is to assess the prevalence and to identify the factors associated with arthritis in the elderly population of Bambui, Brazil. METHODS: A population based cross sectional study was carried out among 1606 senior citizens ( > or = 60 years of age). Arthritis was defined by: a) report of medical diagnosis of arthritis and b) report of chronic hand and knee symptoms (CHKS). Multiple logistic regression was used to investigate the independent association between arthritis and selected factors. RESULTS: Prevalence of medical diagnosis of arthritis was of 25.3% (15.3% in men and 31.9% in women). Prevalence of CHKS was of 44.2%. CHKS was negatively associated with gender (masculine) and years of education ( > or = 8) and positively associated with Body Mass Index (25-29, 30-34, > or = 35 kg/m2), report of myocardial infarct, stroke symptoms and Chaga's disease. CONCLUSIONS: Our results are consistent with other studies concerning higher prevalence of arthritis among obese and less educated women. Association of CHKS with the report of some chronic conditions may be related to greater utilization of health services motivated by these conditions. This finding requires further investigation in future studies. Identification of these characteristics of the senior citizens living in the community, with a higher prevalence of arthritis may subsidize the organization of healthcare programs for this age group. PMID- 15666016 TI - [Bacteremia -- prevalence and associated factors in patients with acute calculous cholecystitis]. AB - BACKGROUND: Bacteremia seems to increase the morbidity and mortality in clinical and surgical conditions, however the importance and factors associated with it during acute calculous cholecystitis are not well established. The purpose of this study was to determine pre-operative prevalence, associated factors and prognosis of bacteremia in patients with acute calculous cholecystitis submitted to an emergency cholecystectomy. METHODS: A prospective study of 51 patients with histological diagnosis of acute calculous cholecystitis was designed. The Bactec System was used to detect bacteremia. Mean results of clinical and laboratory analyses were related to the presence of bacteremia. RESULTS: The pre-operative prevalence of bacteremia was of 15.68%. Age (P=0.024), heart rate (P=0.026), respiratory rate (P=0.028), serum creatinine (P=0.028) and presence of systemic inflammatory response syndrome (P=0.016) were positively associated with bacteremia. Bacteremic patients had statistically higher overall (P=0.045) and infectious (P=0.039) complication rates and longer hospitalization (P<0.005) including one death. CONCLUSIONS: Patients with acute calculous cholecystitis have a considerable prevalence of bacteremia in the pre-operative period. It is associated with old age, heart rate, respiratory rate, serum creatinine and systemic inflammatory response syndrome. In patients with acute calculous cholecystitis, presence of bacteremia may be associated to severity of the condition and poor outcome related to pos-operative complications. PMID- 15666017 TI - [Ketogenic diet for intractable epilepsy in children and adolescents: report of six cases]. AB - BACKGROUND: This study aims to report on use of the ketogenic diet in a group of six children and adolescents with intractable epilepsy. METHODS: Authors reviewed the medical records of every patient under 15 years of age who received the ketogenic diet between April 1999 and July 2003. A comparison is made between treatment results, adverse events and beneficial effects with the pertinent medical literature. RESULTS: The ketogenic diet was administered to six patients, whose median age was 7.0 years (range, 1.8-12.2). Average duration of diet application was of 9.7 months (range, 7 days-4 years). A reduction equal to or greater than 50% in seizure frequency was observed in half of the cases. Complications included neutropenia, constipation, dehydration, priapism, and seizure recurrence. CONCLUSIONS: The ketogenic diet was effective and safe in three out of six patients with intractable epilepsy. Neutropenia was the most common complication. PMID- 15666018 TI - [Secular trends of growth of schoolchildren from Paulinia, Sao Paulo-Brazil (1979/80 - 1993/94)]. AB - BACKGROUND: The purpose of this study was to evaluate the secular trends in height, weight and weight/height of schoolchildren from the city of Paulinia, Sao Paulo, Brazil between 1979/80 and 1993/94. METHODS: Anthropometric measurements (height, weight and weight/height) of 1,903 children (6.5-12.5 y); 51.5% M: 48.5% F, from Paulinia public schools were compared with data from a previous study carried out in the same city 15 years earlier. Decade increments were calculated and data was smoothed by the technique of means and medians (3H3H3). RESULTS: Height and weight mean values were always greater than those of the previous study, with positive increments. Height increments ranged from 1.13 to 5.0 cm in boys and from 1.2 to 4.33 cm in girls. Weight increments ranged from 0.53 to 4.13 kg in males and from 0.87 to 3.0 kg in females. In the two studies, weight/height means were very similar for both genders. CONCLUSIONS: Increments in height and weight during this period are an indicator of development on the economical and health levels. A positive secular trend was also observed in developed countries after the 2nd World War and in the Brazilian people. PMID- 15666019 TI - [Predictors of choledocholithiasis in patients sustaining acute biliary pancreatitis]. AB - BACKGROUND: To assess the role of alkaline phosphatase (AP), gamil glutamyltransferase (gammaGT) and abdominal ultrasound (US) as predictors of choledocholithiasis in patients sustaining acute biliary pancreatitis. METHODS: Data was prospectively collected during a period of 31 months. Forty patients were included, 30 were female and the mean age was 49 +/- 16. All patients sustaining acute biliary pancreatitis were enrolled. Patients with clinical jaundice and severe pancreatitis were excluded. Serum content of AP and gGT as well as US were assessed at admission and 48 hours before cholecistectomy. All patients underwent intra-operative cholangiography (IOC) or pre-operative endoscopic retrograde cholangiography (ERCP), which was indicated based on the odds of choledocholithiasis. In order to identify the predictors of choledocholithiasis, variables were compared between patients sustaining or not such alteration in cholangiography. Student t, Fisher and chi square tests were used for statistical analysis, considering p<0.05 as significant. Positive (PPV) and negative predictor values (NPV) were calculated for each variable. RESULTS: Upon admission, 15 (37%) patients sustained biliary tract dilatation and 5 (12%) choledocholithiasis at the US. Forty eight hours before the operation, 34 (85%) patients had altered levels of gGT and 16 (40%) of AP. Pre-operative US showed biliary tract dilatation in nine patients and choledocholithiasis in three. ERCP was performed in 15 (37%) cases. Higher PPV (55%) was attributed to pre-operative US, which had also a NPV of 96%. CONCLUSION: The best predictor of choledocholithiasis in patients sustaining mild acute pancreatitis was the biliary tract dilatation in pre-operative US. PMID- 15666020 TI - [Effects on hearing due to the occupational noise exposure of marble industry workers in the Federal District, Brazil]. AB - OBJECTIVE: To evaluate the prevalence of hearing loss, its degree and type, among workers in the marble industry in the Brazilian Federal District (FD). METHOD: Workers from eight marble industries in the FD were evaluated by means of a cross sectional epidemiological study. An audiometry screening test (air conduction) was performed. Workers with hearing loss were submitted to liminal tonal audiometry air & bone conduction and speech audiometry tests using an audiometer AD-28 (Interacoustics). All subjects studied were submitted to a visual inspection of the external acoustic meatus. RESULTS: One hundred and fifty two workers were examined; mean age was 32 years (SD = 8.6); average occupational noise exposure was of 8.3 years (SD = 6.8). Audiometries demonstrated that 48.0% (n = 73) had some type of hearing loss. Among the workers with hearing loss, 50.0% had results compatible with noise-induced hearing loss (NIHL); 41.0% with incipient noise induced hearing loss, 5.0% with sensorineural hearing loss (all except NIHL) and 4.0% with conductive and mixed hearing losses. Among workers with NIHL, 57.1% had bilateral involvement, 17.1% in the right ear and 25.7% in the left ear. Among those with incipient NIHL, 13.9% were bilateral, 19.4% were only in the right ear and 66.7% were only in the left ear. CONCLUSION: Abnormal audiograms were found in 48.0% of the sample. Among those with hearing loss, the predominant cause was NIHL, followed by those classified as having incipient noise induced hearing loss. Hearing loss usually started at 6 kHz, frequently in the left ear. PMID- 15666021 TI - [The Pediatrician's ability to recognize the presenting signs and symptoms of retinoblastoma]. AB - BACKGROUND: Retinoblastoma is the most frequent primary intraocular tumor in children. The most common presenting sign is leukocoria and the second one is strabism. In the developing countries extra ocular disease is seen more frequently because of delayed referral. METHODS: Two hundred and thirty-nine patients with retinoblastoma admitted at the Pediatrics and Ophthalmology Departments of the Hospital do Cancer AC Camargo from 1986 to 1995 were evaluated regarding lag-time and tumor extension. RESULTS: Patients referred within less than 6 months presented a higher rate of intraocular disease compared with patients having a lag-time greater than 6 months (75% vs. 25%, p<0.001). No statistically significant difference was noted for patients with extra-ocular tumors. CONCLUSION: Early diagnosis is the gold standard for patients with Retinoblastoma and the pediatrician's ability to recognize the presenting signs and symptoms can lead to early diagnosis and efficient treatment. PMID- 15666022 TI - [Comparison of imaging methods for diagnosis of renal tumors and their calcifications]. AB - BACKGROUND: To establish the best methodology for diagnosis and management of patients with solid and complex renal masses by comparing the costs and benefits of different imaging methods and to improve differential diagnosis of these benign and malignant lesions, particularly by investigating tumour calcifications. METHODS: We performed a prospective study on 31 patients with solid or complex masses by submitting them to Abdominal Ultrasonography (US), Doppler Ultrasonography of the renal mass (US Dop), Computed Tomography (CT), and Magnetic Resonance Imaging (MRI). RESULTS: We found 28 patients with malignant and three with benign masses. Of the 28 malignant, 17 showed calcifications at CT; 16 central and one was of the pure peripheral curvilinear type (egg shell). Excretory Urography (IVP) had a significantly lower detection rate for central calcifications than both US and CT. Benign and malignant masses appeared as described in literature, with US, CT and MRI showing high sensitivity and specificity in renal tumor diagnosis. The exception was US Dop where we obtained lower sensitivity for the characterization of malignant tumor flow. CONCLUSIONS: In this series we were surprised to find that CT revealed central calcifications in 51.6% of patients, all with malignant lesions, while, literature reports a frequency of calcification in renal cell carcinoma between 8 and 22%, in studies using abdominal films and EU (IVP). This finding is of great importance when we consider that these calcifications occur particularly in malignant neoplasms. As a result of comparing these different imaging methods we have developed a better methodology for renal tumor investigation. PMID- 15666024 TI - [Epidemiological profile of mandible fractures treated at the Federal University of Sao Paulo-Paulista Medical School]. AB - BACKGROUND: Mandible fractures can result in esthetic, functional and financial problems and their epidemiological patterns have changed in many locations. This study was carried out to detect these changes, aiming to compare data of patients with mandible fractures treated at the Sao Paulo Hospital (UNIFESP-EPM) from June 1999 to March 2002 with data of patients treated from January 1991 to March 1996. METHODS: Information on most affected gender and age, most often fractured mandible segment, associated injuries, treatment and complications of 98 victims of mandible fracture admitted from June 1999 to March 2002 were compared to the same data of 166 patients treated from January 1991 to March 1996. RESULTS: the most affected gender and age ranges remain the same. Aggressions surpassed traffic accidents as the main etiology. Incidence of associated injuries and multiple fractures in the mandible decreased, a fact probably related to the change in etiology. The most affected segment is still the body of the mandible. The most used type of treatment in both samples was internal rigid fixation with miniplates and the number of complications decreased, due to the higher standard of patient care. CONCLUSION: Mandible fractures in the Sao Paulo population have undergone epidemiological changes and this knowledge enables local authorities to establish adequate measures for prevention and treatment. PMID- 15666023 TI - [Prevalence of ectopic pregnancy liable to surgical treatment in a public hospital from 1995 through 2000]. AB - OBJECTIVES: To determine the prevalence of women with ectopic pregnancy submitted to surgical treatment at the HMCP from 1995 to 2000; as well as to evaluate some characteristics of these women and the treatment performed. METHODS: The study was retrospective. Data was obtained from medical register of women submitted to laparotomy with the diagnosis of ectopic pregnancy. Variables studied were: patient's age, place of birth and occupation, status of ectopic pregnancy (with or without rupture), type of surgery, complications and duration of hospital stay. Data was analyzed in a descriptive way, followed by logistic regression analysis. RESULTS: Ninety six cases were studied. Age of the majority of women (65%) ranged from 25 to 35 years; 6% were adolescents. Of all patients 30% were employed. Distribution according to status of pregnancy: the study showed that 66% presented rupture; 79% of the patients were submitted to salpingectomy with or without ooforectomy. COMPLICATIONS: The most frequent complication was acute anemia, including one death due to DIVC (disseminated intravascular coagulation), and 12% of the patients had to receive blood transfusion. The average time of hospital stay was of 3.63 days (SD+/-0.81). Prevalence of ectopic pregnancies did not change during the period of study and was responsible for 11.2% of the total number of laparotomies. CONCLUSIONS: For the period of this study, the number of women submitted to laparotomy due to ectopic pregnancy remained quite high and unchanged For control, this situation requires preventive measures at different levels. Ectopic pregnancy is a severe pathology and induces high rates of morbidity. It remains a challenge for gynecologists. PMID- 15666025 TI - [HLA polymorphism in a racially admixed sample of the population of Teresina, Piaui]. AB - OBJECTIVE: To establish the frequencies of HLA-A, B, DRB1 and DQB1 specificities in a racially admixed sample of the city of Teresina, Piaui to characterize its genetic composition. METHODS: Polymerase chain reaction-sequence specific primers (PCR-SSP) were used to determine HLA-A, B, DRB1 and DQB1 specificities of 97 unrelated healthy racially admixed people of Teresina. The genotypic frequencies were estimated and compared to those described in samples of Brazilian Caucasian, Portuguese, Black and Amerindian populations using Principal Component Analysis (PCA) and Hierarchical Cluster Analysis (HCA). RESULTS: The frequencies of HLA-A, B, DRB1 and DQB1 specificities observed in the study sample were intermediate between Blacks and Caucasians and the typical elevation of HLA-specificities seen in the Amerindian race was not observed in the study population. The PCA and HCA analysis revealed that Teresina's racially admixed are very close to both Black and Caucasian and do not show similarities with the Amerindians. CONCLUSION: The genetic composition of Teresina's racially admixed is predominantly bi-hybrid of genes originated from Blacks and Caucasians with little contribution from Amerindian genes. PMID- 15666026 TI - [Sensitivity and specificity of hysterosonography in endometrial abnormalities in asymptomatic postmenopausal women]. AB - BACKGROUND: To estimate sensitivity and specificity of hysterosonography for diagnosis of endometrial cavity abnormalities. The gold-standard was hysteroscopy; to compare the agreement between ultrasonographic, hysterosonographic and hysteroscopic findings using the KIA (Kappa Index Agreement). METHODS: Fifty asymptomatic postmenopausal women that had a suspicion of endometrial abnormalities based upon transvaginal ultrasonography were studied. Hysterosonography, diagnostic hysteroscopy and oriented biopsy were performed and the Kia was used to compare results. RESULTS: The most frequent abnormalities were polyps (58%), synechiae (20%), submucous myoma (12%) and endometrial thickening (6%). The uterine cavity was considered normal in 4% of the evaluations by hysteroscopy. The sensitivity of hysterosonography to diagnose polyps was of 89.7%, the specificity of 81.0% and the KIA of 71.1%. For synechia sensitivity of hysterosonography was of 80%, specificity of 100% and the KIA of 86.5%; for submucous myoma sensitivity was of 83.3%; specificity of 97.7% and the KIA of 81.1%, and for endometrial thickening, sensitivity was of 33.3%, specificity of 89.4% and the KIA of 15.5%. CONCLUSION: Hysterosonography showed very good agreement with hysteroscopy for the diagnosis of synechiae and submucous myomas; good agreement for polyps and poor agreement for endometrial thickening. Based upon this data hysterosonography may be deemed a simple, efficient, and accurate method for the evaluation of the uterine cavity in the postmenopausal period. PMID- 15666027 TI - [Retrospective analysis of toxicity of eardrops, topical nasal and oropharyngeal medicines, documented in Sao Paulo, Brazil]. AB - BACKGROUND: Retrospective analysis of human toxicity files involving topical medicines for treatment of upper airways diseases (eardrops, topical nasal medicines, lozenges, drops and sprays for oropharyngeal affections). METHODS: Thirty-four brands of eardrops, 48 of topical nasal medicines and 22 of tablets, lozenges and sprays for oropharyngeal affections were selected, from a total of 104 products available in Brazil. We analyzed the registries in the electronic database from the Poison Control Centre of Sao Paulo (CCI-Jabaquara), Brazil, for the period from January 1996 through December 2000. The cases related to selected pharmaceuticals were collected. RESULTS: 10,823 cases of human toxicity caused by medicines were voluntarily reported to CCI-Jabaquara. Topical medicines for treatment of upper airways diseases accounted for 291 cases (2.68%), from which 240 (82.5%) represented poisoning; 12 (4.1%) involved ear drops, 268 (92%), topical nasal medicines and 11 (3.9%), topical medicines for oropharyngeal affections. Among topical nasal medicines, vasoconstrictors predominated (233 cases), and among medicines for oropharyngeal affections, it was tetracaine (four cases). Considering age distribution, toxicity predominated significantly in children aged from 1 to 4 years (p=0.0003). The main causes of toxicity were: accidental intake of medicines (43%) and error in drug administration (14.8%). Hypereflexia and vomiting were the most frequent symptoms related to toxicity. CONCLUSIONS: There was significant incidence of systemic toxicity due to eardrops, topical nasal and oropharyngeal medicines in children 1 to 4 years-old, whose main cause was accidental intake of these medicines. PMID- 15666028 TI - [Severe sepsis and septic shock in children with cancer]. AB - BACKGROUND: This report describes the clinical characteristics of children and adolescents bearers of oncological disease who were admitted to PICU with severe sepsis and septic shock. The predicting factors for mortality and for need of pulmonary mechanical ventilation were also determined. METHODS: Thirty-three children diagnosed with severe sepsis and septic shock were evaluated prospectively at the PICU of Hospital do Cancer between June and December of 2001. RESULTS: Thirty-three admissions were analyzed during this period; ages ranged from 1 to 23 years; 16 (48%) were boys and 17 (52%) were girls. Twenty patients had leukemia/lymphoma and 13 patients had solid tumors. Twenty-eight patients had a diagnosis of infectious diseases. In 73% of the patients, infection germs were isolated and gram negative organisms were responsible for 67% of the samples. Respiratory support was necessary for 18 patients (54%), inotropic support for 22 (67%) and dialysis for four patients. The mortality rate was of 41% for patients who needed inotropic support, of 69% for those requiring respiratory support and of 100% for those with an indication for dialysis. The overall mortality rate was of 27%. CONCLUSIONS: This research suggests that early intensive treatment for children with cancer exhibiting severe sepsis and/or septic shock could be an important factor to influence the mortality rate of these patients. Moreover, that noninvasive ventilation could be an option to reduce endotracheal intubation and invasive ventilation. PMID- 15666029 TI - [Assessment of orgasmic capacity of postmenopausal women]. AB - OBJECTIVE: To correlate the capacity of healthy postmenopausal women to have orgasms (during intercourse or by solitary masturbation) with psychosocial, behavioral, climacteric, hormonal and interpersonal factors. METHODS: Nine hundred and ninety-nine women (aged from 41 to 60 years) underwent physical and supplementary tests and answered questionnaires regarding sexual behaviour. Sixty healthy women, sexually active, with one or more years of amenorrhea, without hormone therapy and with a partner capable of intercourse were chosen from this group. A Logistic Regression Model with one dependent variable -- orgasmic capacity -- and seventeen independent variables -- psychosocial, behavioral, interpersonal, climacteric and hormonal factors -- was developed. RESULTS: The orgasmic capacity correlated significantly with the practice of masturbation (p=0.000), with pleasure in embracing and caressing the partner's body (p=0.036) and with the presence of vaginal dryness (p=0.021). CONCLUSIONS: This study shows that the most important factors were interpersonal and behavioral and that the other parameters considered were not statistically significant. Women with vaginal dryness, who however engage in masturbation and maintain an affective relationship with their partner, obtain an equal or a greater number of orgasms when compared to the frequency of sexual intercourse. PMID- 15666030 TI - [Research on contraception: opinion of researchers, subjects and advocates of women rights]. AB - OBJECTIVE: To survey the opinion of researchers, women -- subjects of research-- and of advocates of women's rights about the meaning of "research" and "guinea pigs". METHODS: Researchers (11), women (18) and advocates (10) were interviewed. The study was qualitative and a thematic analysis was carried out. RESULTS: Participants experienced difficulty to explain what research meant and some were unable to do so. For the researchers and most of the women, subjects are not guinea pigs, because they have freedom of choice. Most of the advocates, however, said that subjects are guinea pigs, even when they consent to participate in a study. CONCLUSIONS: The difficulty experienced by researchers when explaining the meaning of research was surprising. The different opinions about "guinea pig" reflect the controversy about its meaning in research. Certainly the expression "guinea pig" does not apply to all studies however the different meanings identified should make researchers cautious about the ethical issues involved in their studies. PMID- 15666032 TI - [Institutional Review Boards: compliance with Resolution 196/96]. AB - PURPOSE: This article intends to evaluate the structure, functioning and performance of 17 Institutional Review Boards (IRB), from the viewpoint of their presidents, in relation to the instructions of Resolution 196/96 of the National Council of Health, Ministry of Health, Brazil. METHODS: Presidents of 33 IRBs, that evaluated proposals for research on fertility control, were indicated by the professionals responsible for the area of gynecology in 46 Brazilian medical schools and by the directors of four research centers on human reproduction . A letter with a questionnaire were sent to the presidents, asking them to volunteer for the study. RESULTS: Seventeen presidents completed the questionnaire. Results disclosed a series of violations of Resolution 196/96. Three IRBs did not include representatives of the community among their members; four took over a month to release the Board's decision on proposals and 13 did not follow-up the studies. Composition and filing of the research proposals were in agreement with the Resolution. However, in eight IRBs time of mandate differed from that set forth in the Resolution. Almost all presidents (about 14 of the 17) thought that the composition and activities of their IRBs were adequate. Eleven considered the Resolution appropriate but difficult to comply with. CONCLUSION: These results suggest that an extensive debate on the operational viability of the Resolution would be timely. This endeavor could bring forth valuable suggestions for the improvement and applicability of the Resolution contributing to the improvement of the scientific and ethical quality of research. PMID- 15666033 TI - Cytidine deaminase in polymyalgia rheumatica and elderly onset rheumatoid arthritis. AB - Serum cytidine deaminase (CD) as a marker of inflammatory disease was assessed in 44 patients and 47 controls to differentiate polymyalgia rheumatica (PMR) from elderly onset rheumatoid arthritis (EORA). The patients were divided into four groups: PMR with and without synovitis and seropositive and seronegative EORA. No statistically significant differences were found when serum CD levels of seropositive EORA patients were compared with serum CD of PMR patients without synovitis, neither when serum CD levels of all PMR patients were compared with a seronegative EORA group, nor when serum CD levels of PMR patients with synovitis were compared with those with EORA. Nevertheless, statistically significant differences were detected between EORA's serum CD levels and the control group (p=0.023). This difference was 10% when comparing CD levels of PMR patients with the control group (p=0.070). We did not demonstrate that serum CD levels could be a useful tool to differentiate PMR from EORA, but these findings could nevertheless reflect the presence of an inflammatory disease. PMID- 15666034 TI - Greater superficial petrosal nerve schwannoma. AB - Greater superficial petrosal nerve (GSPN) schwannoma is a very rare type of facial nerve schwannoma. Including our case, only 6 schwannomas have been reported to originate from the GSPN. Clinical features, imaging, diagnosis, differential diagnosis and treatment are discussed reviewing other cases in the pertinent literature. PMID- 15666035 TI - Antioxidant capacity of the neurohormone melatonin. AB - The aim of this study was to elucidate the antioxidant behaviour of melatonin (M) and determine its activity-structure relationship. M or 5-metoxy-N acetyltriptamine is a neurohormone secreted by the pineal gland, which plays a proven role in maintaining sleep-wake rhythms. The antioxidant capacity of M was analysed using the oxygen radical absorbance capacity (ORAC) assay. Furthermore, spectral measurements for aerobic photolytic reaction of neutral red (NR) and degree of inhibition of photolysis with M, glutathione (GSH), ascorbic acid (AA) and vitamin E analogue Trolox were studied at room temperature 25 degrees C, using visible (VIS) and ultra-violet (UV) radiations. In the ORAC assay 2,2 azobis (2-amidino-propane)dihydrochloride (AAPH) a peroxyl radical generator, ROO degrees ; H2O2-Cu2+, mainly a hydroxyl radical generator, degrees OH; and Cu2+ a transition metal were used. Although some studies indicated that M is a powerful antioxidant, no one has compared its antioxidant capacities with GSH, E-vitamin and AA, using three free radical (FR) generators in an assay which utilizes an area-under curve technique and thus combines both inhibition time and inhibition degree of FR action by an antioxidant into a single quantity. In the current study, we used ORAC assay with three FR generators. The assay is based on propensity of the fluorescence emitted by the protein beta-phycoerythrin (beta PE) from porphyridium cruentum to be quenched when exposed to FR action. M in our experiments acted as a universal antioxidant against ROO degrees and degrees OH radicals. Also, M served as an antioxidant in the presence of Cu2+. M, which is a lipid-soluble compound, was a twice more powerful antioxidant than vitamin E, and four times than AA or GSH. Furthermore, M inhibited aerobic photolysis of NR photoinduced with VIS and UV rays faster and more effectively, than AA, GSH or vitamin E. AA with NR, under aerobic conditions during irradiation with VIS and UV acted as a pro-oxidant. M may be the premier molecule to protect the cells from oxidative stress. PMID- 15666036 TI - Serotonin transporter promoter polymorphism genotype is associated with temperament, personality traits and illegal drugs use among adolescents. AB - Serotonin transporter promoter polymorphism (5-HTTLPR) genotype was previously found associated with substance use disorders, particularly in the subjects with comorbid antisocial behavior, and with temperament and personality traits at risk for substance abuse. Aim of the present study was to evaluate the possible association between 5-HTTLPR genotype and the availability to experiment illegal drugs among adolescents, in relationship with psychological characteristics. 216 caucasian high school students (aged 14-19 ys), 125 abstinent subjects, who have never experimented psychotropic drugs, and 91 experimenters of illegal drugs have been genotyped. Aggressiveness levels and temperamental traits were measured in both abstinent subjects and experimenters utilizing respectively Buss-Durkee Hostility-Inventory (BDHI) and Cloninger Three-dimensional Personality Questionnaire (TPQ). Data about school performance have been also collected. The short-short (SS) genotype frequency was significantly higher among experimenters compared with abstinent subjects (p = 0.001). The odds ratio for the SS genotype vs the long-long (LL) genotype frequency was 4.67, 95% Cl (1.97-11.04), when experimenters were compared with abstinent students. The SS genotype frequency was significantly higher among aggressive/novelty seeker (NS) experimenters with poor school achievements, compared with drugs experimenters without aggressiveness and school failure (p = 0.02). When evaluated on the entire sample, BDHI mean total scores, NS scores at TPQ and school failure frequency were significantly higher in SS individuals, in comparison with LL subjects. Our data suggest that a decreased expression of the gene encoding the 5-HTT transporter, due to "S" promoter polymorphism, may be associated with an increased availability to experiment illegal drugs among adolescents, particularly in the subjects with more consistent aggressiveness, NS temperament and learning disabilities. PMID- 15666038 TI - Changes in vascularization in substantia nigra pars compacta of monkeys rendered parkinsonian. AB - The degeneration of nigral dopaminergic neurons in Parkinson's disease is believed to be associated with a glial reaction and inflammatory changes. In turn, local factors may induce changes in vascularization and contribute to neuronal vulnerability. Among these factors, Vascular Endothelial Growth Factor (VEGF) is released in adults under pathological conditions and is thought to induce angiogenesis. In order to determine whether changes in brain vasculature are observed in the affected brain regions in parkinsonism, we quantitatively analysed the VEGF-expressing cells and blood vessels in the substantia nigra of monkeys rendered parkinsonian by MPTP injection and compared the results with those obtained in control monkeys. Using stereological methods, we observed an increase in the number of VEGF-expressing neurons and an increase of the number of blood vessels and their volume occupying the substantia nigra pars compacta of monkeys rendered parkinsonian by chronic MPTP intoxication. These changes in vascularization may therefore modify the neuronal availability of blood nutrients, blood cells or toxic substances and neuronal susceptibility to parkinsonism. PMID- 15666039 TI - Startle response in inpatients with borderline personality disorder vs. healthy controls. AB - Startle response is found to be enhanced in conditions that are accompanied by intense emotions, high levels of anxiety in particular. We assessed the autonomous and behavioral component of the startle reflex in a sample of 28 unmedicated inpatients with borderline personality disorder (BPD) compared to 28 controls. Amplitudes of electrodermal and startle response as well as habituation were measured in response to 15 bursts of white noise (100-db, 40-ms). We assessed prepulse inhibition to study gating capacities in BPD, with each prepulse (70-db, 30-msec) delivered 100 ms before the startling tone. In contrast to our hypothesis, our study revealed neither enhanced amplitudes of autonomic and electromyographic startle responses nor differences in habituation compared to healthy controls. In addition, no deficit in prepulse inhibition was found in BPD. In summary, our data indicate normal response to unconditioned threatening stimuli as well as normal gating capacities in BPD. PMID- 15666037 TI - Influence of peroxisome proliferator-activated receptor gamma gene polymorphism on 24S-hydroxycholesterol levels in Alzheimer's patients. AB - The peroxisome proliferator-activated receptor gamma (PPARgamma) is a nuclear hormone receptor, that is involved in lipid and glucose metabolism, which both seem to influence the risk of Alzheimer's disease (AD). 24S-Hydroxycholesterol is the major cholesterol elimination product of the brain and plasma and CSF 24S hydroxycholesterol levels are altered in patients with neurodegenerative diseases. We investigated the effect of the common Pro12Ala variant of the PPARgamma gene on plasma cholesterol levels and 24S-hydroxycholesterol/ cholesterol ratios in 124 AD patients and 77 healthy controls. Furthermore, the influence of PPARgamma polymorphism on the risk of AD in 247 AD patients and 324 healthy controls was investigated. We found that PPARgamma Pro12Ala polymorphism influenced plasma 24S-hydroxycholesterol/ cholesterol ratios in AD patients in that carriers of the Ala allele presented with higher ratios than homozygote carriers of the Pro-allele. PPARgamma polymorphism did not influence the risk of AD. These results might point to an influence of PPARgamma Pro12Ala polymorphism on the elimination of 24S-hydroxycholesterol. PMID- 15666040 TI - Brain SPECT imaging in children and adolescents with obsessive-compulsive disorder. AB - OBJECTIVE: To evaluate the patterns of regional cerebral blood flow (rCBF) in cortical and subcortical regions by Brain SPECT imaging, in children and adolescents with obsessive-compulsive disorder (OCD) before and after treatment. METHOD: Fourteen OCD patients (6 to 17 years old) underwent brain SPECT; ten of those subjects were reexamined after successful treatment. rCBF ratios were correlated with clinical parameters on the 14 patients in symptomatic state, and we compared rCBF ratios of the ten patients before and after treatment. RESULTS: There was no statistically significant difference in average ratios of rCBF before and after treatment. There were significant clinical correlations between current age and age of onset of OCD and rCBF in the bilateral superior frontal, and bilateral parietal cortical regions. CONCLUSIONS: Further investigations on abnormal neurodevelopment of cortical-subcortical circuits possibly involved in symptomatology of paediatric OCD are warranted. PMID- 15666041 TI - CNS Targets for multi-functional drugs in the treatment of Alzheimer's and Parkinson's diseases. AB - Patients with mild forms of dementia and age-related memory impairment have just begun to benefit from pharmacotherapy developed over the last several years. However, current approaches do not significantly modify the course of neurodegeneration or of the aging process, and they offer limited and transient benefit to many patients. The goal of this review is to summarize new potential approaches in which molecules have been developed expressly to target multiple brain systems for the treatment of memory and cognition impairment. Some of these approaches include the development of single molecular entities that combine activity as cholinesterase inhibitors, muscarinic cholinergic M2 receptor antagonists, nicotinic acetylcholine receptor agonists, alpha(2)-adrenergic agonists, or monoamine oxidase inhibitors. Many of the bi-functional compounds discussed have improved efficacy as cognitive enhancing agents and/or they offer potential for neuroprotection and disease modification. It is likely that syndromes such as Alzheimer's disease will require multiple drug therapy to address the varied pathological aspects of the disease. Even if the strategy of combining drugs with different therapeutic targets is workable, the development of multi-functional compounds will obviate the challenge of administering multiple single drug entities with potentially different degrees of bioavailability, pharmacokinetics, and metabolism. Also, the simplification of the therapeutic regimen for individuals with AD who have difficulty with compliance is important. PMID- 15666042 TI - The application of mesh support in periareolar breast surgery: clinical and mammographic evaluation. AB - BACKGROUND: Numerous techniques have been described for the treatment of breast hypertrophy and ptosis. Unfortunately, recurrent ptosis after mammaplasty can occur regardless of the technique used. To avoid this problem, different kinds of supporting devices have been described with variable rates of success. However, the true implications of incorporating prosthetic materials into breast surgery have never been clarified. Therefore, surgeons have traditionally been reluctant to apply any kind of prosthetic material to the breast, fearing inflammation, an unfavorable aesthetic outcome, palpable or visible deformities, and interference with the mammographic evaluation of breast cancer. This study analyzed the aesthetic, clinical, and mammographic implications of using mesh as a supportive device in periareolar breast surgery. METHODS: For this study, 18 patients (mean age, 42 years) with breast hypertrophy, ptosis, or both were managed with the double-skin periareolar mammaplasty technique, with placement of mixed (60% Polyglactine and 40% polyester) mesh. Clinical assessment was performed by three breast surgeons actively working on cancer surveillance who knew that the patients had experienced mesh application. After a mean follow-up period of 30 months, a standard mammogram was performed for each patient and analyzed by both the surgeons and an expert radiologist. The evaluated factors were hyperemia, calcifications, contour irregularities, capsular contraction, thickening or widening of the scar with extrusion of the mesh, and any palpable or hardened areas. RESULTS: According to the authors' clinical observations, there were no mesh-related abnormalities in the breast; the mesh was not palpable after the operation; and there was no recurrent ptosis. In terms of mammographic imaging, the mesh was visible as a very fine line in the periphery of the breast's parenchyma (measuring 0.2 mm on the lateral views) in three patients (17%). The mesh did not interfere with the visualization and analysis of the breast's parenchyma. In seven patients (39%), benign localized microcalcifications were detected in the breast and no further investigation was performed. In two patients (11%), grouped calcifications were detected and biopsied, with histopathologic analysis demonstrating epithelial hyperplasia with atypia. In two patients (11%), nodules smaller than 1 cm were detected and biopsied, with histopathologic analysis demonstrating a fibroadenoma in one patient and an invasive ductal carcinoma in the other. CONCLUSIONS: The use of mesh support in breast surgery can enhance the aesthetic results without inducing visible or palpable deformities or mammographic abnormalities. In terms of surveillance mammograms, the presence of the mesh did not interfere with the diagnosis and treatment of minute lesions such as calcifications and small nodules. PMID- 15666043 TI - Gynecomastia treatment with subareolar glandular pedicle. Discussion, 287. AB - BACKGROUND: Gynecomastia is enlargement of the male breast. Although treatment is not indicated in most cases, aesthetic reconstructive surgery is commonly performed for psychological reasons. The goals in surgical treatment are to restore the breast contour with minimal scar and to protect areolar anatomy and sensation. This clinical study investigates the results of the subareolar glandular pedicle technique, in which the pedicle is dissected 2 mm wider than the areola with a circumareolar incision and the breast tissue is excised en bloc. The technique differs from the classical circumareolar approach with its thinner pedicle and excision of the breast without leaving prepectoral tissue. METHODS: We operated on nine patients with grades 1 and 2 gynecomastia using the subareolar glandular pedicle. Exposure was excellent with the circumareolar incision. Neither hematoma nor seroma formation was seen in any of the patients. Partial areola necrosis, which caused suture separation, was the only early postoperative complication seen, and this in a patient who smoked heavily. Patients were followed for at least 6 months. RESULTS: Eight patients achieved a good aesthetic contour of the chest, and one patient needed a contour revision for the residual mass because of a bulky pedicle. Circumareolar scars were satisfactory for all the patients, including the patient with partial areola necrosis. Circumareolar hyperpigmentation developed in one patient, and areola sensation was preserved in seven patients. CONCLUSIONS: The subareolar glandular pedicle is indicated for grades 1 and 2 gynecomastia. Circumareolar incision provides perfect exposure. The technique is reliable if the pedicle is dissected 2 mm wider than the areola and dissection under the pedicle is avoided. Postoperative circumareolar scarring is minimal and nipple-areola sensation is preserved in most cases. However, experience is needed to determine the pedicle girth because a wide pedicle leads to subareolar bulk, whereas a thin pedicle may cause partial areola necrosis. PMID- 15666044 TI - A technique to decrease breast shape deformity during muscle contraction in submuscular augmentation mammaplasty. AB - The main objective of this study was to decrease breast shape distortion during pectoralis muscle contraction following submuscular augmentation mammaplasty. We followed 348 patients who had retromuscular augmentation mammoplasty: 251 (72.1%) had polyurethane-covered gel-filled, 97 (27.9%) had textured-silicone gel-filled implants. Among the 348, 46 had surgery following the Regnault technique and 302 had surgery by the below-mentioned technique. Periareolar incision, bipartision of breast parenchyma down to the fascia, undermining of breast base from the fascia downward to the inframammary sulcus or a little below it, detaching of muscle off the thoracic cage, disinsertion of abdomino-costal pectoralis attachments. Full thickness incision of pectoralis muscle on a vertical line on the nipple projection for 2-5 inches. Placing of prosthesis. Drainage. Closure. No objective evaluation was used, only clinical judgments by three observers--the surgeon, a nurse and the patient herself. The results showed a definite decrease of the dynamic deformity among patients in whom the author's technical variation was carried out. This technique allows also, for decreasing the upward pushing of the implant during pectoralis muscle contraction and facilitates stretch of the breast tissue in patients with tighter breast envelopes. Despite lack of precise measurements, conclusions drawn from clinical judgments, taken as objectively as possible, suggest that the use of this technique may offer the solution of an otherwise disturbing collateral effect, frequently seen after this operation. PMID- 15666045 TI - Direct skin excision fat reshaping and repositioning for correction of prominent nasolabial fold. AB - In addition to all the consequences mentioned elsewhere in the literature, the aging process causes the most dramatic changes in the midface and nasolabial complex. Genetic predisposition also plays a role in the final appearance of this area. The nasolabial complex in the aged face generally is accepted as a "hard-to treat area" for which many different techniques have been described. Among the techniques described to date, direct excision still keeps its popularity for selected patients. However, some controversies in the technique are open to discussion. To overcome these problems, we did skin excision, reshaping and repositioning the fibrofatty tissue lateral to the fold, which is contrary to fat excision in the classical technique. It seems to be a better approach to reshape and reposition the ptotic tissue so as not to obliterate but rather to smooth out the nasolabial area. Excision of fat would decrease the amount of tissue in the already atrophic and ptotic face. This technique offers a new approach to nasolabial complex problems. The most significant drawback is the scar, which usually is imperceptible in selected patients. PMID- 15666046 TI - External rhinoplasty for the Arabian nose: a columellar scar analysis. AB - This study aimed to evaluate columellar scar problems after external rhinoplasty in the Arabian population, and to analyze the technical factors that help prevent such problems and maximize the scar cosmesis. The investigation was conducted in university and private practice settings of the author in Alexandria, Egypt. A total of 600 Arab patients who underwent external rhinoplasty were included in the study. All the patients underwent surgery using the external rhinoplasty approach, in which bilateral alar marginal incisions were connected by an inverted V-shaped transcolumellar incision. At completion of the procedure, a two layer closure of the columellar incision was performed. At a minimum of 1 year postoperatively, the columellar scar was evaluated subjectively by means of a patient questionnaire, and objectively by clinical examination and comparison of the close-up pre- and postoperative basal view photographs. Objectively, anything less than a barely visible, leveled, thin, linear scar was considered unsatisfactory. Subjectively, 95.5% of the patients rated the scar as unnoticeable, 3% as noticeable but acceptable, and 1.5% as unacceptable. Objectively, the scar was unsatisfactory in 7% of the cases. This was because of scar widening with or without depression (5%), hyperpigmentation (1.5%), and columellar rim notching (0.5%). The use of a deep 6/0 polydioxanon (PDS) suture significantly decreased the incidence of scar widening (p < 0.005).The columellar incision can be used safely in the Arab population regardless of their thick, dark, and oily skin. Technical factors that contributed to the favorable outcome of the columellar scar included proper planning of location and design of the incision used, precise execution, meticulous multilayered closure, and good postoperative care. PMID- 15666048 TI - Low-volume tumescent liposuction does not change plasma malondialdehyde levels: a preliminary study. AB - To evaluate the effects of low-volume tumescent liposuction on plasma malondialdehyde (MDA) levels, a controlled clinical study was conducted. Venous blood samples were taken just before and 10 min after the completion of tumescent suction-lipectomy for 15 patients. The means +/- standard deviations for patients' data were as follows: of patients' age (29.81 +/- 13.2 years) aspirate volume (1,250 +/- 450 ml), and body mass index (24.5 +/- 1.2). There was no significant difference between the preoperative and postoperative plasma malondialdehyde levels (p > 0.05). The results show no evidence of increased lipoperoxidation in the systemic circulation during low-volume tumescent liposuction. PMID- 15666047 TI - Cosmetic lateral canthoplasty. AB - BACKGROUND: Although there are patients who want cosmetic lateral canthoplasty to lengthen the lateral palpebral fissure of the lateral canthal area, no recommendable operational procedure exists that results in an acceptable scar formation. Because of the round eyeball shape, lengthening and widening of the lateral palpebral fissure should not be in the lateral direction, but rather in the posterior direction to keep the surface between the bulbar conjunctiva and the palpebral conjunctiva in contact. METHODS: The authors believed that using an upper crus flap of the upper eyelid margin as a transposition flap would enable lengthening of the lower eyelid width and posterior deepening of the lateral canthal area. This was thought possible by transverse back-cutting of the palpebral conjunctiva on the lateral fornix area. RESULTS: The authors have performed cosmetic lateral canthoplasty in more than 200 cases to increase the width of the lateral palpebral fissure. All the results were satisfactory and aesthetically acceptable. CONCLUSIONS: The reported surgery makes the eyes appear larger and brighter. The authors think this procedure is a good and convenient method although it is not perfect. PMID- 15666049 TI - The use of polarized light in aesthetic surgery. AB - This article presents a clinical investigation of polarized light therapy after aesthetic surgery procedures. The study included patients who the authors underwent face-lifts, blepharoplasties, and various facial ancillary procedures, at center during the past 2 1/2 years. One side of the surgically treated area was managed with polarized light, whereas the other side served as a control. The results were compared using clinical examination only including signs of recovery such as resolution of swelling and bruises. The results in most cases showed a significant difference between the treated and untreated sides. PMID- 15666050 TI - Long-term results of ultrapulsed carbon dioxide laser resurfacing of the Mediterranean face. AB - BACKGROUND: In Mediterranean countries, increased exposure to sunlight accelerates aging of the skin and the formation of wrinkles. The long-term follow up results for the patients who underwent resurfacing with ultrapulsed carbon dioxide (CO(2)) laser are presented. METHODS: All 47 patients who underwent ultrapulsed CO(2) laser between 1994 and 1996 were included in the study. The patients were photographed with their permission before the laser treatment, at postoperative month 6, and at year 5, with comparison of the results. RESULTS: In this study, with its 5-year follow-up period, it was found that the ultrapulsed CO(2) laser achieved long-lasting and acceptable facial resurfacing. CONCLUSIONS: The ultrapulsed CO(2) laser enables damage to the abnormal dermal region with high-energy transfer in a very short period. The recent development of high energy pulsed CO(2) laser has generated much renewed enthusiasm for cutaneous resurfacing, especially over the long term. PMID- 15666051 TI - The fate of intramuscularly injected fat autografts: an experimental study in rabbits. AB - An experimental study was designed to assess the viability and revascularization of intramuscularly injected fat autografts. For the study, 18 rabbits were divided into two groups. In the first group, fat was injected intramuscularly (12 rabbits). Autologous fat was obtained from the inguinal area and subsequently injected into the thigh muscle. In the second group, physiologic saline was injected intramuscularly to determine the effects of cannulation and pressure on muscle tissue (6 rabbits). Fat autografts were performed on the right side of the animal, and the left side was used as the control. Scintigraphic imaging and histopathologic examination of the limbs were performed after injection of adipose tissue on days 15, 30, 45, 60, 90, and 120. On the technetium-99m ((99m)Tc) hexamethylpropylene amine oxime scintigraphy, whereas similar activity distribution was observed between the left and right thigh on days 15, 30, and 45, there was increased uptake at the right thigh on days 60, 90, and 120. This increased uptake indicates that there is viable fat tissue in this region. Histopathologic evaluation showed that microcysts resulting from degeneration of some adipocytes and inflammatory changes on day 15 additionally increased vascularity and fibrosis in some animals on day 30, as well as fibrosis, microcysties, and focal calcification areas in adipose tissue on day 45 and later. It was observed that adipose tissue survived in more than 50% of the graft area in all the animals. These findings show that fat autografts can survive in muscle tissue with less than 50% fibrotic change. PMID- 15666052 TI - What makes buttocks beautiful? A review and classification of the determinants of gluteal beauty and the surgical techniques to achieve them. AB - BACKGROUND: No reports can be found in medical literature describing the anatomic details that make the buttocks beautiful, nor the defects in this region that can help the surgeon decide whether gluteal prostheses are necessary or not, and, if so, which type, design, and volume of prostheses should be used, in which plane they should be placed, and so forth. METHODS: The authors studied 1,320 photographs of nude women and measured 132 female patients ages 16 to 62 years. They found that four characteristics determine attractive buttocks. They also found the balanced anatomic dimension of the maximal gluteal projection. RESULTS: On the basis of the information obtained, the defects of the gluteal region were classified into five types, resulting in surgical recommendations intended to achieve buttocks as close as possible to the standard of beauty, with the right volume and projection. CONCLUSIONS: The authors concluded that surgery for correction of the buttocks involves more than projection and volume. The surgeon must be familiar with the "signs" of beauty so an attempt can be made to recreate them. Most importantly, the surgeon should achieve buttocks with projection and volume. However, these must be in a balanced proportion with the rest of the body. PMID- 15666060 TI - An explanation of the unexpected efficacy of L-DOPS in pure autonomic failure. PMID- 15666061 TI - Sympathetic genes, baroreflexes, and hypertension. PMID- 15666062 TI - To tilt or not to tilt: what is the question? PMID- 15666063 TI - Clinical pharmacokinetics of the norepinephrine precursor L-threo-DOPS in primary chronic autonomic failure. AB - BACKGROUND: Oral L-threo-3,4-dihydroxyphenylserine (L-DOPS), a synthetic catechol amino acid, increases standing blood pressure and improves standing ability in patients with neurogenic orthostatic hypotension, by conversion of L-DOPS to norepinephrine (NE) outside the brain. This study assessed the pharmacokinetics of L-DOPS, NE, and dihydroxyphenylglycol (DHPG), the main neuronal metabolite of NE, in patients with primary chronic autonomic failure from pure autonomic failure (PAF) or multiple system atrophy (MSA). METHODS: In 5 MSA and 4 PAF patients, antecubital venous blood was drawn during supine rest and plasma levels of catechols measured at various times for 48 hours after a single oral dose of 400 mg of L-DOPS. RESULTS: Plasma L-DOPS peaked at 1.9 microg/ml (9 micromol/L) about 3 hours after drug administration, followed by a monoexponential decline with a half-time of 2-3 hours in both patient groups. Plasma NE and DHPG also peaked at about 3 hours, but at much lower concentrations (4 and 42 nmol/L). Compared to the MSA group, the PAF group had a smaller calculated volume of distribution of L-DOPS and up to 10-fold lower plasma NE levels at all time points. Plasma NE was above baseline in MSA even at 48 hours after L-DOPS. CONCLUSIONS: The relatively long half-time for disappearance of L-DOPS compared to that of NE explains their very different attained plasma concentrations. The similar NE and DHPG responses in PAF and MSA suggests production of NE from LDOPS mainly in non-neuronal cells. Persistent elevation of plasma NE in MSA suggests residual release of NE from sympathetic nerves. PMID- 15666064 TI - Autonomic nervous system activity in normotensive subjects with a family history of hypertension. AB - This study was designed to address alterations in autonomic nervous system activity in normotensive subjects with a family history of hypertension. We compared the autonomic nervous system activity in 59 normotensives with a family history of hypertension and 46 normotensives with no family history of hypertension. Skin blood flow was measured using laser-Doppler method on the nailfold skin in the resting condition, during systemic cooling and during upright tilting. Finger blood pressure, pulse and ECG were monitored by a finapres device. Heart rate, systolic pressure and microvascular flow power spectral analyses were performed using fast Fourier transformation. Baroreflex sensitivity was estimated with the sequence method. Compared to the control group, normotensives with a family history of hypertension showed significantly higher systolic pressure, decreased proportion and area of the high-frequency band of the heart rate variability power spectrum and reduced baroreflex sensitivity in the resting condition as well as a decreased proportion and area of the high-frequency band of the heart rate variability power spectrum during systemic cooling. We also proved a different time course of baroreflex sensitivity during upright tilting in the two groups. In contrast, we did not find any significant differences in the parameters of systolic pressure and microvascular variability power spectra between the two groups. Our results indicate that even normotensives with a family history of hypertension exhibit an increased ratio of sympathetic to parasympathetic activity at the cardiac level; however, they do not show any alteration of the vascular sympathetic reactivity. PMID- 15666065 TI - Comparison of the active standing test and head-up tilt test for diagnosis of syncope in childhood and adolescence. AB - We examined 51 children and adolescents with orthostatic symptoms using two orthostatic tests, the active standing test (the AS test) and head-up tilt test (HUT), and compared circulatory responses, autonomic function in addition to the induction rate of syncope during short-time orthostasis. Syncope was induced in eight patients with both tests, in only six patients with the AS test and in only one patient with HUT. The induction rate was significantly higher with the AS test (p<0.0001). In addition, the AS test is common and daily postural motion and does not require a tilt table. We calculated percent changes in systolic blood pressure at the initial drop (DeltaID-SBP), in systolic blood pressure (DeltaSBP), in diastolic blood pressure (DeltaDBP), in heart rate (DeltaHR), component coefficient variation LF/HF (DeltaLF/HF) from supine to upright. DeltaHR were significantly larger in fainters than in non-fainters with both tests, although there was no difference in DeltaSBP and in DeltaDBP. In six fainters only with the AS test, DeltaHR was significantly larger with the AS test than with HUT. With the AS test DeltaID-SBP were correlative with DeltaLF/HF, and DeltaLF/HF were correlative with DeltaHR, whereas these relations were not clear in HUT. These results indicated the AS test caused cardiac sympathetic activation associated with an initial pressure drop, and was more prone to induce syncope with a greater HR increase in some patients. We conclude the AS test is as potential as HUT as a diagnostic test for syncope. PMID- 15666066 TI - Effect of local heating and vasodilation on the cutaneous venoarteriolar response. AB - The cutaneous venoarteriolar response (CVAR) is a local non-adrenergic vasoconstrictor reflex that is engaged via increases in local transmural pressure. The purpose of this study was to determine if local temperature alters the vasoconstrictor ability of the CVAR. Twelve (5 male, 7 female) subjects performed a CVAR maneuver at local temperatures of 30+/-1, 34, 38, and 42 degrees C. CVAR was also engaged after vasodilation via intradermal perfusion of sodium nitroprusside or the attenuation of local heating-induced vasodilation via intradermal perfusion of N(G)-nitro-L: -arginine methyl ester (L: -NAME) in five subjects (2 male, 3 female). CVAR was elicited by rapid cuff inflation to 45mmHg proximal to two dorsal forearm sites for 2 min in both protocols. Local heating caused a progressive increase in skin blood flow (8+/-1, 18+/-4, 43+/-11, and 78+/-2% maximal skin blood flow for 30+/-1, 34, 38, and 42 degrees C, respectively). Engagement of the CVAR decreased skin blood flow by 53+/-2, 57+/ 3, and 51+/-4%, for 30+/-1, 34, and 38 degrees C, respectively. In contrast, local heating to 42 degrees C significantly attenuated the CVAR (16+/-11 %). Local administration of sodium nitroprusside during neutral temperature and L: NAME during local heating also significantly attenuated the vasoconstrictor response of the CVAR by 27+/-8 and 38+/-4%, respectively. These data indicate that CVAR is attenuated at high (42 degrees C) local skin temperatures and that this attenuation is likely due to an effect of both local heating-induced vasodilation and a direct temperature effect. PMID- 15666068 TI - Effect of unilateral forced nostril breathing on tonic accommodation and intraocular pressure. AB - BACKGROUND: Unilateral forced nostril breathing (UFNB) has specific measurable effects on the autonomic nervous system. Ocular accommodation, which is controlled by the autonomic nervous system, would be expected to be under the influence of UFNB when it is applied. The purpose of this study was to investigate the effect of UFNB on the resting state of the accommodation system, i. e. tonic accommodation (TA), along with measures of intraocular pressure (IOP), blood pressure and heart rate. METHODS: TA levels were measured using the Shin-Nippon autorefractor before and after 20 minutes of UFNB. IOP, blood pressure and heart rate, which are known to be affected by UFNB, were also measured with a non-contact tonometer and an automated blood pressure monitor respectively. RESULTS: Right and left UFNB produced slight, but not significant changes in TA. However, there was a tendency for left UFNB to produce a greater decrease in TA in subjects with higher base-line TA levels. Right UFNB produced a statistically significant decrease in IOP while the effect of left UFNB on IOP was not significant. CONCLUSION: UFNB produced changes in IOP consistent with previous reports. As studied in this trial, UFNB did not have any significant effect on TA. Further studies using a larger sample size are required to investigate the effect of UFNB on the autonomic inputs to the ciliary muscle of the eye and the subsequent measures of tonic accommodation. PMID- 15666067 TI - Birth prematurity determines prolonged autonomic nervous system immaturity. AB - Because of its multiple involvement in physiological processes, autonomic nervous system (ANS) activity, a key regulator of homeostatic control, demonstrates a progressive increase during pregnancy. The profile of its maturation, mainly in the parasympathetic arm, in premature or full term infants, may help us to better understand its pathophysiological role. We prospectively evaluated ANS maturity in a group of 23 premature (PREM) infants at their theoretical term age and in 8 full term (FT) newborns. All recordings were registered close to the theoretical full term period (from 38 to 41 weeks) for the PREM group and during the first week of life for the FT newborns. Polygraphic recordings, EEG monitoring associated with visual clinical control, and Holter ECG, were performed simultaneously. ANS indices were then calculated during quiet sleep periods, using Wavelet transform of RR (beat to beat) intervals. High frequency components were found to be significantly lower in the PREM than in the FT group (p<0.05). Furthermore, at theoretical full term age, the greater the prematurity, the lower was parasympathetic activity. Because it is easy, monitoring of parasympathetic activity may help us to understand autonomic maturation and its clinical prognostic implications. PMID- 15666069 TI - Distal site testing of sympathetic skin response (big toe) in diabetic polyneuropathy. AB - The aim of this study was to determine whether the sympathetic skin response (SSR) recorded from the big toe is more sensitive than standard SSR recorded from the sole for the detection of sudomotor fiber dysfunction in diabetic neuropathy. We recorded big toe SSR (SSRBT) and plantar SSR (SSRP) in 17 diabetic patients with non-disabling neuropathy (group A), 13 patients with disabling neuropathy (group B) and 30 age-matched normal controls. With regard to controls, SSRP amplitude was reduced only in group B. In contrast, SSRBT amplitude was reduced in both groups of patients (p<0.0001). In 8 patients in group B, SSRBT was not recordable while the SSRP still persisted. Our results suggest that SS-RBT is a more sensitive test than SSRP in detecting distal sudomotor failure in patients with diabetic neuropathy. PMID- 15666079 TI - Medical humanities and cultural studies: lessons learned from an NEH Institute. AB - In this essay, the directors of an NEH Institute on "Medicine, Literature, and Culture" consider the lessons they learned by bringing humanities scholars to a teaching hospital for a month-long institute that mingled seminar discussions, outside speakers and clinical observations. In an exchange of letters, they discuss the productive tensions inherent in approaching medicine from multiple perspectives, and they argue the case for a broader conception of medical humanities that incorporates the methodologies of cultural studies. PMID- 15666071 TI - Comparison of QT interval duration and dispersion in elderly population versus healthy young subjects. AB - We compared QT dispersion (QTd) in 75 elderly and 36 young subjects and found that those over the age of 75 years had higher QTd than those younger than 75. In addition those older than 75 years had higher QTd values compared to the younger subjects. We concluded that QTd increased especially over the age of 75. PMID- 15666070 TI - Postprandial hypotension treated with acarbose in a patient with type 1 diabetes mellitus. AB - Treatment of postprandial hypotension (PPH) is often unsuccessful. We report a case of a type 1 diabetic patient suffering from severely symptomatic PPH. The patient was treated with acarbose and showed definite improvement of both glycemic control and PPH. PMID- 15666080 TI - Coming out: considering the closet of illness. AB - This essay explores key concerns surrounding "coming out" as a person with illness and addresses important professional and social considerations for those who are closeted in various kinds of illness. Using central tenets of Queer Theory and Disability and Cultural Studies as a theoretical base, I examine the politics of coming out in the specific context of my lived experience during the 2002 NEH Summer Institute, "Medicine, Literature, and Culture" While such an environment might foster unusual candor about personal illness experience, I discovered that the choice to come out as a person with chronic, non-infectious disease (Inflammatory Bowel Disease) was nevertheless complicated in interesting ways. PMID- 15666081 TI - Foucault on the case: the pastoral and juridical foundation of medical power. AB - This paper employs Foucault's concept of "governmentality" to examine critically the efforts by medical humanists to reform the medical case. I argue that these reform efforts contribute to the individualizing dimensions of medical power through the development of a "pastoral" technique that medicine has taken over from religious authority. Clinical experiences at this NEH Institute also revealed a juridical dimension of the medical case that treats a patient's statements as suspect and in need of corroboration by evidence provided by the patient's body. The combination of these pastoral and juridical dimensions of the case contributes to the normalizing power of modern medicine, and medical humanists need to be aware of their own contribution to this form of power as they reform the case. PMID- 15666082 TI - Power at work: navigating hierarchies, teamwork and webs. AB - I explore tensions between the dynamics observed during a shadowing experience in a clinic and a hospital and theories of power and organizations. Commenting on what the tensions reveal about "power at work" (in the dual sense of how power works and power in the workplace), I suggest that some of the most gratifying experiences of workplace collaboration for healthcare professionals may occur as a result of their having successfully navigated complex coalitions of hierarchies, teams and webs rather than dutifully adhering to any single organizational structure. PMID- 15666083 TI - (In/Out)side AIDS activism: searching for a critically engaged politics. AB - Experience has always been a hallmark of activist work; my work in AIDS activism began with my family's role as caretakers for two children whose parents died of HIV-related complications. Previously, my scholarly work critiqued political and medical establishments and their policies surrounding HIV/AIDS. At the NEH institute, I interacted with the medical world, shadowing nurses and doctors. Through this experience, I discovered the importance of interactivity as a crucial element of the critically engaged AIDS activist experience, creating a more thorough understanding of the medical establishment and a more humanized portrait of hospitals and their staff. PMID- 15666085 TI - Enzymes of sphingolipid metabolism in Drosophila melanogaster. AB - Sphingolipids are important structural components of membranes that delimit the boundaries of cellular compartments, cells and organisms. They play an equally important role as second messengers, and transduce signals across or within the compartments they define to initiate physiological changes during development, differentiation and a host of other cellular events. For well over a century Drosophila melanogaster has served as a useful model organism to understand some of the fundamental tenets of development, differentiation and signaling in eukaryotic organisms. Directed approaches to study sphingolipid biology in Drosophila have been initiated only recently. Nevertheless, earlier phenotypic studies conducted on genes of unknown biochemical function have recently been recognized as mutants of enzymes of sphingolipid metabolism. Genome sequencing and annotation have aided the identification of homologs of recently discovered genes. Here we present an overview of studies on enzymes of the de novo sphingolipid biosynthetic pathway, known mutants and their phenotypic characterization in Drosophila. PMID- 15666084 TI - Protein kinase C regulation of GABAA receptors. AB - Pharmacological studies with drugs that activate or inhibit several protein kinase C (PKC) isozymes have identified the PKC family of serine-threonine kinases as important in the regulation of gamma-aminobutyric acid type A (GABA(A)) receptor function. PKC modulates GABA(A) receptor surface density, chloride conductance and receptor sensitivity to positive allosteric modulators such as neurosteroids, ethanol, benzodiazepines and barbiturates. Recent studies using PKC isozyme-selective reagents and gene-targeted mice have begun to identify critical roles for three isozymes, PKCbetaII, PKCvarepsilon and PKCgamma, in various aspects of GABA(A) receptor regulation. Progress in this field touches upon therapeutic areas that are of great clinical importance such as anxiety and addiction. Increased understanding of how PKC regulates GABA(A) receptors and which PKC isozymes are involved holds promise for development of new treatments for diverse neuropsychiatric disorders. PMID- 15666086 TI - Aluminium in Alzheimer's disease: are we still at a crossroad? AB - Aluminium, an environmentally abundant non-redox trivalent cation has long been implicated in the pathogenesis of Alzheimer's disease (AD). However, the definite mechanism of aluminium toxicity in AD is not known. Evidence suggests that trace metal homeostasis plays a crucial role in the normal functioning of the brain, and any disturbance in it can exacerbate events associated with AD. The present paper reviews the scientific literature linking aluminium with AD. The focus is on aluminium levels in brain, region-specific and subcellular distribution, its relation to neurofibrillary tangles, amyloid beta, and other metals. A detailed mechanism of the role of aluminium in oxidative stress and cell death is highlighted. The importance of complex speciation chemistry of aluminium in relation to biology has been emphasized. The debatable role of aluminium in AD and the cross-talk between aluminium and genetic susceptibility are also discussed. Finally, it is concluded based on extensive literature that the neurotoxic effects of aluminium are beyond any doubt, and aluminium as a factor in AD cannot be discarded. However, whether aluminium is a sole factor in AD and whether it is a factor in all AD cases still needs to be understood. PMID- 15666087 TI - Serotonin reuptake inhibitors and cardiovascular diseases: a platelet connection. AB - Selective serotonin reuptake inhibitors (SSRIs) are a heterogeneous group of new antidepressants that cause a well documented acquired but reversible serotonin deficiency in blood platelets. Platelets are small, anucleate cells and are the only blood cells specialized in storing peripheral serotonin. Platelets are also an integral part of the hemostatic process that is initiated during pathologic thrombus formation in cardiovascular diseases. Serotonin release from platelets is important for functional hemostasis as indicated by congenital diseases with serotonin-deficient platelets that can lead to life-threatening bleeding problems. The postulate that SSRIs should have an impact on cardiovascular diseases is therefore well founded. Cardiovascular effects of SSRIs have indeed been shown in a number of studies investigating the effect of SSRIs in patients with psychosomatic comorbidity. SSRIs reduce the incidence of recurrent myocardial infarction (MI) in patients suffering from post-MI depression. In addition, SSRIs inhibit tight clot formation of platelets in vitro, which points to a direct anti-thrombotic or pro-fibrinolytic effect of SSRIs. PMID- 15666088 TI - alpha-1,3-Fucosyltransferase-VII stimulates the growth of hepatocarcinoma cells via the cyclin-dependent kinase inhibitor p27Kip1. AB - After the transfection of alpha-1,3-fucosyltransferase (FucT)-VII cDNA into H7721 human hepatocarcinoma cells, the protein expression of some cyclins, cyclin dependent kinases (CDKs) and cyclin-dependent kinase inhibitors (CDIs) p16INK4 and p21waf1/Cip1 were unchanged. However, CDI p27Kip1 protein, both the total amount and the amount that bound to CDK2, but not its mRNA, was significantly reduced. The de-inhibited CDK2 stimulated the phosphorylation of retinoblastoma (Rb) protein and facilitated the G1/S transition and growth rate of the cells. The decrease of p27Kip1 protein, the increase of CDK2 activity and Rb phosphorylation, as well as the cell growth and percentage of S phase cells were correlated to the increased amount of cell surface sialyl Lewis X (SLe(x)) antigen in cells with different alpha-1,3-FucT-VII expression. The reduction in p27Kip1 and the difference in its expression among different transfected cells were blocked by the SLe(x) antibody KM93 in a dose-dependent manner, indicating that p27Kip1 expression was influenced by alpha-1,3-FucT-VII and its product SLe(x). The MEK/MAPK signaling pathway was more important than the PI-3K pathway in the regulation of p27Kip1 expression. PMID- 15666089 TI - Mycobacterial lipoarabinomannans modulate cytokine production in human T helper cells by interfering with raft/microdomain signalling. AB - Lipoarabinomannans (LAMs) are major lipoglycans of the mycobacterial envelope and constitute immunodominant epitopes of mycobacteria. In this paper, we show that mannose-capped (ManLAM) and non-mannose-capped (PILAM) mycobacterial lipoglycans insert into T helper cell rafts without apparent binding to known receptors. T helper cells modified by the insertion of PILAM responded to CD3 cross-linking by decreasing type 1 (IL-2 and IFN-gamma) and increasing type 2 (IL-4 and IL-5) cytokine production. Modification by the mannose-capped ManLAMs had similar, but more limited effects on T helper cell cytokine production. When incorporated into isolated rafts, PILAMs modulated membrane-associated kinases in a dose-dependent manner, inducing increased phosphorylation of Src kinases and Cbp/PAG in Th1 rafts, while decreasing phosphorylation of the same proteins in Th2 rafts. Mycobacterial lipoglycans thus modify the signalling machineries of rafts/microdomains in T helper cells, a modification of the membrane organization that eventually leads to an overall enhancement of type 2 and inhibition of type 1 cytokine production. PMID- 15666090 TI - Sp1-associated activation of macrophage inflammatory protein-2 promoter by CpG oligodeoxynucleotide and lipopolysaccharide. AB - Macrophage inflammatory protein-2 (MIP-2) is a C-X-C chemokine that is important in recruiting neutrophils to inflammatory sites. Our previous reports demonstrated that lipopolysaccharide (LPS) or CpG-oligode-oxynucleotide (CpG-ODN) rapidly induce MIP-2 gene expression in the macrophage cell line, RAW 264.7. Here, we show that the DNA sequence of the MIP-2 promoter between -114 and +14 is sufficient for strong promoter activity in LPS- or CpG-ODN-stimulated RAW 264.7 cells. Importantly, comprehensive mutant analysis reveals that an Sp1 element in the promoter region between -114 and -94 is essential for synergistic MIP-2 promoter activation by NF-kappaB and c-Jun regardless of the presence of an AP-1 site. By combining deletion or site-specific mutant analysis with immunocomplex assays, we also confirmed that Sp1 mediates the recruitment of transcription factors NF- kappaB and c-Jun in LPS- or CpG-ODN-treated RAW 264.7 cells. Several lines of experimental evidence imply that the Sp1-binding element is an important determinant of MIP-2 promoter activity, and that NF-kappaB, c-Jun and Sp1 can functionally cooperate to elicit maximal activation of the promoter. PMID- 15666092 TI - Formation of the B cell synapse: retention or recruitment? AB - Interaction of B cells with membrane antigen results in the formation of the B cell synapse: the B cell receptor (BCR) and antigen concentrate in the contact zone while CD45/B220 and the phosphatase SHP-1 are excluded. This study shows that, unlike in T cells, synapse formation does not require active transport processes (while subsequent antigen extraction and IgM downregulation do). The synapse architecture depends on the available protein ligands in the contact zone. Thus Syk, IgM and Fc receptor accumulation require the presence of ITAM bearing BCRs, membrane antigen and membrane (IgG-containing) immune complexes, respectively. Remarkably, non-bound proteins are frequently not only homogeneously distributed but excluded from the contact zone. These results suggest that proteins mainly reach the contact zone by undirected diffusion, and in order not to be expelled by molecular crowding they require capture by and fixation to a binding protein. PMID- 15666091 TI - Evidence for a novel racemization process of an asparaginyl residue in mouse lysozyme under physiological conditions. AB - We examined chemical reactions in mouse lysozyme after incubation under physiological conditions (pH 7 and 37 degrees C). After incubation for 8 weeks, racemization was observed specifically at Asn127 among the 19 Asp/Asn residues in mouse lysozyme. Furthermore, analysis of the primary structure showed that the racemized residue was not Asp, but Asn, which demonstrates that deamidation and isomerization did not occur. These results mean that this racemization occurs without forming a succinimide intermediate. This is the first example of D asparaginyl formation in a protein occurring during the racemization process under physiological conditions. PMID- 15666093 TI - The transcription factor profile of human mast cells in comparison with monocytes and granulocytes. AB - Expression profiles of mRNAs and proteins for various transcription factors were determined for human skin mast cells (sMCs), leukemic HMC-1 MCs, monocytes and granulocytes. By quantitative RT-PCR, sMCs expressed lower levels of c-fos, PU.1, C/EBPalpha, and C/EBPepsilon than monocytes and granulocytes, but higher levels of MITF, SCL, GATA-1 and GATA-2. At the protein level, MITF, SCL, GATA-2, Elf-1 and c-fos were clearly detectable in sMCs. With the exception of c-fos, these proteins were absent or expressed only slightly in monocytes and granulocytes. The expression of NF-E2p45, GATA-1, PU.1, Ets-1, C/EBPalpha and C/EBPepsilon was below the detection limit in sMCs, but detectable in other myelocytes. The high expression of SCL and GATA-2 in sMCs is reminiscent of stem cells. The absence of C/EBPvarepsilon in sMCs, but strong expression in HMC-1, suggests it may impair MC maturation. In summary, mature human MCs can be characterized as C/EBPalpha(low), C/EBPvarepsilon-, PU.1(low), GATA-1(low), GATA-2+, SCL+, MITF(high). PMID- 15666096 TI - Another view of the role of FMRP in translational regulation. PMID- 15666097 TI - Genoa Update on Colorectal Polyp. Proceedings of the advanced course in coloproctology: the colorectal polyps, from genetics to surgery. Genoa, Italy, 11 12 March 2004. PMID- 15666095 TI - Oestrogen-modulated gene expression in the human endometrium. AB - To identify key regulatory mechanisms in the growth and development of the human endometrium, microarray analysis was performed on uncultured human endometrium collected during menstruation (M) and the late-proliferative (LATE-P)-phase of the menstrual cycle, as well as after 24 h incubation in the presence of oestradiol (17beta-E2). We demonstrate the expression of novel gene transcripts in the human endometrium. i.e. mucin-9, novel oestrogen-responsive gene transcripts, i.e. gelsolin and flotillin-1, and genes known to be expressed in human endometrium but not yet shown to be oestrogen responsive, i.e. connexin-37 and TFF1/pS2. Genes reported to be expressed during the implantation window and implicated in progesterone action, i.e. secretoglobin family 2A, member 2 (mammaglobin) and homeobox-containing proteins, were up-regulated in uncultured LATE-P-phase endometrium compared to M-phase endometrium. Some gene transcripts are regulated directly by 17beta-E2 alone, others are influenced by the in vivo environment as well. These observations emphasise that the regulation of endometrium maturation by oestrogen entails more then just stimulation of cell proliferation. PMID- 15666098 TI - Genetics of colorectal polyps. AB - Currently, genetic investigation of human tumours starts from the analysis of advanced cancers. Once a given genetic alteration has been found in advanced tumours, this same alteration is investigated in the pre-neoplastic lesions. The aim of this approach is to assess the significance of the genetic alteration during the carcinogenic process. This review is focused on alterations that have proven to be present in pre-neoplastic lesions that are associated to colorectal cancer (ACF and early adenoma). Alterations that are present at the early stages are likely to play a crucial role in colorectal tumorigenesis. Colorectal tumorigenesis is extremely heterogeneous from a genetic point of view: tumours follow alternative molecular pathways and show different phenotypes (CIN, MIN and CIMP). Tumours are genetically heterogeneous from their early stages: the sequence of genetic events that accumulate within cells during progression to malignancy appears to be determined by the first events. These events have been investigated in ACF and in early adenomas. The understanding of the molecular mechanisms underlying genesis and progression of colorectal tumours will allow the development of new tools for cancer prevention and early diagnosis, as well as for therapeutic approaches specific for different molecular targets. PMID- 15666094 TI - Compensatory caspase activation in MPP+-induced cell death in dopaminergic neurons. AB - Many have hypothesized that cell death in Parkinson's disease is via apoptosis and, specifically, by the mitochondrial-mediated apoptotic pathway. We tested this hypothesis using a mouse dopaminergic cell line of mesencephalic origin, MN9D, challenged with the Parkinsonism-causing neurotoxin MPP+ (1-methyl-4 phenylpyridinium ion). Apoptosis was the main mode of cell death when the cells were subjected to MPP+ treatment under serum-free conditions for 24 h. Caspase-3 and caspase-9, however, were not activated, thus indicating the existence of alternate or compensatory cell death pathway(s) in dopaminergic neuronal cells. Using caspase inhibitors, we demonstrated that these pathways involve caspase-2, 8, -6 and -7. A time-course study indicated that activation of caspase-2 and -8 occurred upstream of caspase-6 and caspase-7. Upon MPP+ challenge, the apoptosis inducing factor was translocated from the mitochondria into the MN9D cytosol and nucleus. These results suggest the existence of alternative apoptotic pathways in dopaminergic neurons. PMID- 15666099 TI - Epidemiology of colorectal polyps. AB - The prevalence of colorectal adenomatous polyps varies widely from country to country. Among asymptomatic, average-risk patients, adenoma prevalence averages approximately 10% in sigmoidoscopy studies and more than 25% in colonoscopy studies, whereas the prevalence of colorectal cancer among these patients is less than 1%. These data may change in the future due to the advent of new technological approaches and, in particular, chromo- and magnifying endoscopy as well as confocal laser endoscopy. The cumulative incidence of new adenomas within 3 years after normal endoscopy averages about 7% by flexible sigmoidoscopy and 27% by colonoscopy. As far as risk factors for colorectal adenomas are concerned, several data are now available on the potential role of various diet items. Tobacco smoking may be important in the early stages of adenoma formation, but not necessarily in the later stages. Alcohol consumption elevates the risk of adenomatous colorectal polyps and this seems increased by ADH3 polymorphism. Another gene-environment relationship of interest in colorectal tumorigenesis may be based on folate's effects on K-ras mutations. PMID- 15666100 TI - Chemoprevention of colorectal cancer: an update. AB - Colorectal cancer (CRC) is the leading cause of cancer-related mortality in western countries. Adjuvant treatment does not seem to be highly effective and recurrent or metastatic disease occurs in half of the new cases within one year of diagnosis and median survival does not exceed 18 months. CRC represents an optimal model for primary and secondary prevention, given the availability of effective screening procedures and of a well defined multi-step carcinogenic pathway. Colon cancer is supposed to arise as the result of a series of genetic mutations, which parallel histopathologic and molecular changes, from normal colonic epithelium to invasive carcinoma, with adenomatous polyps as an intermediate step. A growing body of evidence has shown a wide variety of effective compounds, in vitro in animal models and in human clinical trials. The more studied agents are the non-steroidal anti-inflammatory drugs. Among those, aspirin has been shown, in two recent randomised trials, to lower the incidence on polyps vs. placebo. Intervention studies on diet showed disappointing results, but diet micronutrients are promising agents in CRC prevention. Calcium, vitamin D and folic acid in different proportions in different populations have been shown to have a certain degree of action in preventing cancer development in epidemiological studies and in randomised trials. Also oestrogens or, rather, hormone replacement therapy for the menopause can protect against CRC. In conclusion, the rapid growth of information and knowledge in chemoprevention, especially for CRC, is very encouraging and gives us hope that soon this approach will be applicable in a larger scale population. PMID- 15666101 TI - Malignant adenoma: diagnosis, staging, risk factors, lymph node involvement and problems of sampling. AB - Colorectal adenoma containing invasive carcinoma corresponds to a carcinoma invading the submucosa and represents the earliest form of clinically relevant colon cancer. Despite the generally favourable course after endoscopic removal, a limited risk of developing lymph node metastases still exists and the correct histologic assessment of malignant polyps can help in defining the evolutive potential. At present, histopathologic parameters alone determine whether a high (35%) or low (7%) risk of nodal metastases exists and the most relevant diagnostic criteria are the grade of differentiation of invasive adenocarcinoma, vascular invasion, the level of invasion of carcinomatous cells and the status of the resection margin. PMID- 15666102 TI - Colonic neoplastic polyps: biopsy is not efficient to exclude malignancy. The Trafford experience. AB - The biological behaviour of the polyp depends entirely on its histological subtype. Adenomas are the classic precursors to carcinoma. Polyp biopsy can differentiate neoplastic (adenoma and carcinoma) from non-neoplastic polyps (hyperplastic, inflammatory, hamartomatous). Due to potential sampling error, biopsy does not always differentiate between adenoma and carcinoma. Our aim was to investigate the degree of this potential sampling error. We carried out a study to compare the results of biopsies vs. total tissue examinations in a cohort of 433 neoplastic polyps. In this study 18.5% of carcinomatous polyps show definitive malignant elements only when the entire polyp is presented to the pathologist. We have therefore adopted a special reporting template if the polypoidal tumour is either biopsied or incompletely excised. The study also showed that within adenomas there could be variance of architectural patterns between the biopsy and the whole specimen. In this series, biopsying malignant colonic polyps gives rise to 18.5% false negative results and therefore it is significantly less efficient in the diagnosis of malignancy compared to total polyp examination. PMID- 15666103 TI - Flat colorectal neoplasms: endoscopic detection, clinical relevance and management. AB - The prevalence of flat neoplasms in Western countries appears to be similar to Japan. These lesions are difficult to detect but careful examination of the colonic mucosa with the aid of targeted dye spray reveals subtle mucosal changes characteristic of these lesions. Management of flat neoplasms depends on their size, site and on whether the lesions lift after submucosal injection. In this article, we highlight the typical features of flat neoplasms and discuss their optimal management in routine practice. PMID- 15666104 TI - Colorectal carcinoma: is screening possible? AB - Colorectal cancer (CRC) is a major health problem in western countries. It is the most frequently diagnosed cancer in EU countries with about 220,000 new cases per year; the number of CRC deaths approaches 112,000. Most cancer arises from adenomatous polyps through the adenoma-carcinoma sequence; the natural history of the disease is long: the risk of invasive cancer in adenoma is about 2% per year. Given the natural history of CRC, early diagnosis represents the most appropriate tool to reduce the disease-related mortality. Several tests are available to screen healthy subjects at average risk for CRC: faecal occult blood tests, flexible sigmoidoscopy, combined faecal occult blood tests and flexible sigmoidoscopy, total colonoscopy and double contrast barium enema. Issues related to the introduction on a large scale of population-based screening programmes for CRC are discussed. PMID- 15666105 TI - Sensitivity of diagnostic examinations for colorectal polyps. AB - The removal of adenomatous polyps of the large bowel reduces mortality from colorectal cancer (CRC). Faecal occult blood testing only reveals 20.40% of polyps. The flexible rectosigmoidoscope explores less than half of the large bowel. Its use should always be coupled with faecal occult blood testing which, if positive, requires a total colonoscopy. The sensitivity of double-contrast barium enema for the search of polyps is 35%. Colonoscopy does not reach the caecum in about 10% of cases. It misses 15-20% of polyps with diameter <10 mm and about 6% of polyps with diameter >10 mm. Virtual colonoscopy has substantially the same sensitivity as optical colonoscopy for polyps > or =7 mm in diameter. PMID- 15666106 TI - Advanced diagnostic tools: virtual colonoscopy. AB - Regarding new prospects in detecting polyps and cancers of the colon and the rectum, it is impossible to neglect the rising role of CT colonography, also called virtual colonoscopy (VC). The aim of this brief paper is to show the state of the art of VC as a relatively new radiological diagnostic tool, discussing techniques, applications, problems, results in the literature and prospects. PMID- 15666107 TI - Advanced diagnostic tools: virtual colonoscopy and magnifying chromoendoscopy. AB - Early endoscopic diagnosis of colorectal cancers (CRCs) represents the best tool for the reduction of CRC mortality, but the conventional colonoscopic view seems unable to clarify faint changes of colorectal mucosa. For these reasons, during the last few decades, two new techniques have been developed in order to better define, diagnose and treat colorectal lesions: chromoendoscopy (CE) and magnifying chromoendoscopy (MC). The authors herein highlight the benefits of both and compare the results of conventional endoscopy and CE with 0.2% indigo carmine solution, evaluating 995 consecutive selected patients referred for colonoscopy between January 1999 and March 2003 at the International Health Union of Rome. CE showed new neoplastic patterns not detectable with conventional endoscopy in 102 patients (10.2%). The incidence of high-grade dysplasia and early carcinomas was 9.9% for protruded, 13.1% for flat and 60% for depressed lesions. Our comparative study on 995 consecutive selected patients confirmed the existence of flat and depressed neoplasms in Italy and shows the high accuracy rate of CE over conventional endoscopy for the detection of non-polypoid lesions. CE should be used routinely in order to enhance the early diagnosis of CRCs, while MC in experienced hands represents a significant advance in colonoscopic practice improving the diagnosis of every single lesion, enhancing therapeutic efficacy of colorectal tumours. PMID- 15666108 TI - Endoscopic polypectomy: techniques, complications and follow-up. AB - Colorectal cancer is one of the most common cancers in developed countries. Increasing numbers of early stage colorectal cancers and precancerous adenomatous lesions, polypoidal type lesions, and flat and depressed type lesions can be visualised and treated endoscopically thanks to endoscopic ultrasonography, high magnification chromoendoscopy and other technical advances. Polypectomy is generally considered for protuberant lesions, with both sessile and pedunculate morphology, and endoscopic mucosal resection (EMR) is indicated for superficial, flat or depressed types of lesions. Endoscopic therapy for colonic adenoma with dysplasia and early colorectal cancer is more advantageous than the conventional operative treatment, in that it is a relatively non-invasive and less costly method. However, endoscopic therapy is completely ineffective in lesions with lymph node or distant metastasis. Therefore, it is becoming more and more important to understand the correct indications and limitations of endoscopic polypectomy/mucosectomy as well as complication rates and the correct follow-up schedule. PMID- 15666109 TI - Indications for colorectal resection for adenoma before and after polypectomy. AB - Adenomas presenting in colon and rectum require different strategies. Those in the rectum are accessible to local endoanal excision while those in the colon can be removed by endoscopic polypectomy. Essentially the decision in both cases centres around the risk of malignancy. Does the tumour itself contain a focus of malignancy? Where invasion has been demonstrated in the tumour, are the regional lymph nodes involved? Rectal adenomas can be assessed by clinical examination, rectal ultrasound and magnetic resonance imaging. Colonic adenomas on the other hand, are assessed by endoscopic appearances amplified by chromo- and zoom endoscopy techniques and by endoscopic endosonography. In both locations the decision for surgical resection may be necessary in two clinical circumstances: the preoperative assessment of invasion and following the histopathology report where invasion has been demonstrated. With any adenoma with malignant change, a surgical opinion should be obtained. The decision for surgery depends on the risk of failure of the local excision balanced by the risk of morbidity and mortality following major surgery. Patients' wishes in the light of discussion of clinical and pathological prognostic factors are crucial in making the decision. Surgery is more frequently applied for early colonic than rectal cancers. This is due to two main factors: resective colonic surgery has less complications and mortality than anterior resection and transanal local excision may be more radical than endoscopic colonic polypectomy because it excises the full-thickness rectal wall. PMID- 15666110 TI - Locating polyps by endoscopy with or without videolaparoscopy, radioguided occult colonic lesion identification or magnetic endoscopic imaging: the way forward to complete polyp removal. AB - Endoscopic polypectomy is the gold standard for the treatment of colorectal polyps. In the case of non-palpable lesions or to complete polyp removal, the lesions are located intra-operatively. With the advent of laparoscopy, identifying their position is even more important because there is no opportunity for intestinal palpation. Several methods of preoperative endoscopic marking have been proposed using different types of tattooing and recently using clips followed by ultrasonography detection. Innovative methods are analysed; magnetic endoscopic imaging is a reliable and accurate method for determining the anatomical position of the tip of the endoscope during colonoscopy. Radioguided colonic lesion identification needs a gamma detection probe. Endoscopic removal can be converted to endo-laparoscopic rendezvous, failing which, laparoscopic resection is a reliable and safe choice, offering all the advantages of minimally invasive surgical techniques. PMID- 15666111 TI - Local excision of rectal polyp: indications and techniques. AB - The local excision of a rectal polyp is often wrongly considered to be a minor surgical procedure. In reality, the malignant potential of adenomas and the not infrequent presence of cancer in larger polyps, require, for their removal, an oncologically correct operation with strict indication and accurate execution. Despite an increasing inclination to extend the indications of endoscopic polypectomies to polyps of larger size and villous configuration, the local surgical approach remains the preferred treatment in most cases. Here the indications and the results of different surgical techniques proposed for the local excision of a rectal polyp are reported. Among these procedures, transanal endoscopic microsurgery is gaining a primary role in many circumstances. PMID- 15666112 TI - Familial adenomatous polyposis: genetics and epidemiology. AB - Familial adenomatous polyposis (FAP) is a rare genetic disease characterised by the development of hundreds to thousands of adenomatous polyps along the colon rectum leading to cancer at a young age, if left untreated. In 1991, the gene responsible for the vast majority of FAP cases, the adenomatous polyposis coli (APC) gene, was identified. In 5-30% of FAP patients, no APC mutation is identifiable by current genetic testing. In 2003, it was shown that 'APC negative' FAP patients may carry biallelic mutations in a different gene, the MYH gene. Genetics of FAP will be discussed in relation to its present clinical applications. If the hereditable mutation(s) is/are known in a family, it is possible to plan endoscopic surveillance only for those who actually inherited the mutation(s). Also, genetic testing may be of help in the diagnosis of atypical adenomatous polyposis cases and in the clinical management of affected individuals. PMID- 15666113 TI - Familial adenomatous polyposis. Surgical treatment: when and how. AB - Familial adenomatous polyposis is an autosomal dominated inherited disease, caused by the mutation of the tumour suppressor gene adenomatous polyposis coli on chromosome 5. Despite being a rare disorder, accounting for some 1% of colorectal cancers, it represents an interesting model of hereditary disease, because of its intrinsic characteristics, conventionally defined by the presence of more than 100 colorectal polyps, as well as extra-colon manifestations, the attenuated form of the disease, genetic aspects, the inevitable progression to colorectal cancer and hence the correct therapy to treat or prevent the fatal evolution of the disease. Surgical treatment is based above all on two techniques: ileorectal anastomosis, which requires careful surveillance of rectal remnant, and ileal pouch-anal anastomosis, which totally eradicates the disease. The suitability of using these two techniques is discussed in view of new genetic and clinical findings, acquired from personal experience and from the literature. PMID- 15666115 TI - Functional diagnosis using multimodal spinal cord evoked potentials in cervical myelopathy. AB - The electrophysiological measurement of spinal cord evoked potentials (SCEPs) has been established as a tool for diagnosing the spinal level responsible for cervical myelopathy. Only a few studies, however, employed multimodal SCEPs for this purpose. The objective of this study was to investigate the correlation between multimodal SCEPs recorded during cervical laminoplasty and magnetic resonance imaging (MRI) abnormalities in 18 patients aged 70 years and older versus 32 patients less than 65 years of age. Both the SCEPs and MRIs showed higher incidences of multiple-level cord involvement in the older group than in the younger group. Discrepancies in the spinal levels involved between the SCEPs and the MRIs were found in 12 patients (67%) for the older group and 6 patients (19%) for the younger group. The accuracy of the MR images in localizing the lesion site was significantly lower in the older group than in the younger group, indicating that MR images tend to show clinically silent cord compression in elderly patients. Because spinal cord compression can appear without functional change in elderly patients, electrophysiological evaluations with intraoperative multimodal SCEP studies are a useful addition to MRI for understanding the pathology of myelopathy. PMID- 15666116 TI - Results of total hip arthroplasty with bone graft and support ring for protrusio acetabuli in rheumatoid arthritis. AB - We evaluated 19 hips of 15 rheumatoid arthritis patients with protrusio acetabuli that had been surgically treated with total hip arthroplasty (THA) using a support ring; four hips were replaced with a TACT cup supporter, one hip with a Kerboull plate, six hips with a Ganz ring, and eight hips with a Muller ring. The average age was 60.2 years (range 48-75 years), and the average follow-up was 38 months (range 6-74 months). Two patients (three hips) died before the final follow-up, and two patients (two hips) underwent revision owing to loosening of the Ganz ring. Hip functions were rated according to the evaluation chart of hip joint functions of the Japanese Orthopaedic Association (JOA score). The mean total JOA score was 25.2 +/- 8.6 preoperatively and 55.6 +/- 8.7 postoperatively. Radiographically, 17 hips (minus the two revised hips) showed satisfactory incorporation of bone graft and no loosening of the support ring. The mean depth of protrusio acetabuli for these 17 hips was 3.5 +/- 4.1 mm before operation, 2.8 +/- 5.1 mm just after operation, and 2.3 +/- 3.9 mm at the final follow-up (no significant difference at the critical rate of 5%). Our study indicated the usefulness of THAs using a bone graft and a support ring in RA patients with protrusio acetabuli. PMID- 15666117 TI - Surgical treatment of developmental dysplasia of the hip in the periadolescent period. AB - The surgical management of patients with neglected developmental dysplasia of the hip (DDH) after the age of 6 years has been the subject of controversy. We present 11 cases (16 hips) of neglected DDH that were treated operatively by means of open reduction and derotational subtrochanteric osteotomy. Patient age ranged between 10 and 17 years (mean, 12 years). Follow-up ranged from 5 to 13 years (mean, 8.7 years). The results have been satisfactory both clinically (evaluated using the modified Harris hip score) and radiographically (evaluated using Severin's classification). Our data suggest that neglected DDH cases, not only during early childhood, but also in the periadolescent period, should be considered for surgical treatment. PMID- 15666118 TI - Clinical results in severe carpal tunnel syndrome and motor nerve conduction studies. AB - The purposes of this study were to assess the value of motor nerve conduction studies as a prognostic indicator for severe carpal tunnel syndrome (CTS) and to investigate residual nerve disorder after surgery. Fifty hands in 46 patients with severe CTS were followed for at least 6 months after surgery for open carpal tunnel release. Compound muscle action potential (CMAP) from the abductor pollicis brevis (APB) muscle and the second lumbrical (SL) muscle were recorded before and after surgery, and distal latency (DL) and amplitude were analyzed. APB-CMAP was not recordable in 43 hands while SL-CMAP was recordable in all 50 hands before surgery. Results were excellent in 28 hands, good in 16 hands, fair in 6 hands, with no poor results after surgery. The ratio of excellent result was 65% in patients in whom DL of preoperative SL-CMAP was less than 10 ms, and 20% in patients whose DL was 10 ms or more. Delayed DL was seen in postoperative APB CMAP and SL-CMAP, especially in the patients with good or fair results. We consider that SL-CMAP was valuable as a prognostic indicator for severe CTS, and if DL was 10 ms or more, myelinization of fibers would not be sufficient after surgery. PMID- 15666119 TI - Percutaneous leverage pinning in the treatment of Bennett's fracture. AB - Twelve patients (eleven males, one female; mean age 36 years) with Bennett's fracture having a gap or a step-off of more than 2 mm and in whom maintenance of the reduced position was difficult were treated by percutaneous leverage pinning. Bone union was obtained in all patients, and the wire was removed 37 days on average after the initial treatment. On the final follow-up examination, mild pain with motion was noted in two patients, and moderate to severe pain was noted in one patient. The reduction of the articular surface of the carpometacarpal joint of the thumb was confirmed by radiography; it was less than 1 mm in nine patients, less than 2 mm in two, and more than 2 mm in one. Because reduction and fixation are performed utilizing the leverage force of the wire inserted into the trapezium, percutaneous pinning has advantages, such as technical simplicity and the ability to apply tension to the fractured site. PMID- 15666120 TI - Efficacy of different dosing schedules of etidronate for stress shielding after cementless total hip arthroplasty. AB - Forty-four women who had undergone cementless total hip arthroplasty (THA) were selected for determination of the optimum dosage of etidronate in the treatment of stress shielding after surgery. Patients were 55-86 years of age. The patients were randomized into three groups. The control group (n = 17) was not treated with osteoactive drugs. The low-dose group (n = 12) and the high-dose group (n = 15) received 200 mg or 400 mg etidronate daily for 2 weeks, followed by 12 weeks of calcium supplementation of 500 mg/day. These patients received four cycles of therapy over 1 year postoperatively. Periprosthetic bone mineral density (BMD) was measured with dual-energy X-ray absorptiometry at 3 weeks, 6 months, and 12 months. At 12 months, bone loss in the low-dose and high-dose groups was significantly lower compared with the control group in Gruen zones 1 and 7. There were additional significant differences with regard to bone loss between the control group and the high-dose group in zones 2, 4, and 6. Our data suggest that high dosages are more effective in reducing postoperative bone loss after cementless THA. PMID- 15666121 TI - Effect of flexion angle on coronal laxity in patients with mobile-bearing total knee arthroplasty prostheses. AB - Proper soft tissue tension is one of the important factors in mobile-bearing total knee arthroplasty (TKA). We evaluated varus/valgus laxities, particularly at flexion, which is a key factor in reducing the risk of subluxation and dislocation of bearings to assess the effect that the flexion angle and the presence or absence of the posterior cruciate ligament (PCL) have on laxity in patients with low-contact stress (LCS) prostheses of the PCL-retaining (24 patients, 24 knees) and PCL-sacrificing (24 patients, 24 knees) type designs during extension and flexion. Both types of prosthesis had about 4 degrees laxity at extension and 3 degrees at flexion. PCL-retaining prostheses had significantly less laxity at flexion than at extension (P = 0.0004 in varus, P = 0.0043 in valgus). For good clinical outcomes following TKA, 3 degrees-4 degrees laxity in the varus and valgus orientations is recommended. In addition, the PCL might be involved in flexion and could affect varus/valgus laxity in PCL-retaining prostheses. PMID- 15666122 TI - Importance of the lateral anatomic tibial slope as a guide to the tibial cut in total knee arthroplasty in Japanese patients. AB - Using three-dimensional computed tomography in 50 osteoarthritic knees, we simulated at various cutting angles the tibial cut for total knee arthroplasty. Cutting angles of 0 degrees, 3 degrees, 5 degrees, 7 degrees, 9 degrees, and 11 degrees were used. We then calculated the anterior and posterior thicknesses, the medial-lateral widths, and the medial and lateral condylar depths of the resected tibial bone at each cutting angle. Each set of measurements was evaluated according to a comparison between the cutting angle and the anatomic posterior slopes. The cutting angles showing the smallest anterior-posterior difference at the medial and lateral plateaus were 9 degrees and 7 degrees, respectively. The mean anatomic posterior slopes at the medial and lateral plateaus were 9.0 degrees and 8.1 degrees, respectively. When the tibia was cut at the cutting angle closest to the medial anatomical posterior slope in each knee, the thickness of the lateral posterior plateaus resected from 12 knees (24%) was more than 10 mm. Among these 12 knees, the cutting angle was more than 9 degrees in 9 knees (75%). In contrast, when the tibia was cut at the cutting angle closest to the lateral anatomical posterior slope, only one resected medial posterior plateau was more than 10 mm. The cutting angle of this case was 7 degrees. Therefore, in consideration of the thickness of bone resection from anatomic posterior slope, we demonstrated the importance of using the lateral anatomic posterior tibial slope as a guide to the tibial cut. There was no significant difference with respect to resected bone morphology at any cutting angle. PMID- 15666123 TI - Topographical histology of the posterolateral corner of the knee, with special reference to laminar configurations around the popliteus tendon: a study of elderly Japanese and late-stage fetuses. AB - We investigated the histology of laminar configurations at the posterolateral corners of 40 knees (18 late-stage fetuses, 22 elderly persons). In the fetuses, the deeply located popliteal fascia and the superficially located inferior geniculate vascular sheath were evident in a space between the popliteus tendon (PT) and the biceps femoris and gastrocnemius. Along, and deep to, the popliteal fascia, the popliteus muscle parenchyma developed into both the PT and another dense connective tissue mass. The PT attached to the fibula (i.e., the fetal popliteofibular ligament) and to the base of the lateral meniscus (i.e., the fetal popliteomeniscal fasciculus). This laminar configuration was essentially maintained in the elderly. The fetal dense connective tissue of the popliteus origin seemed to correspond to the adult arcuate ligament. However, because a connective tissue complex (including the PT and the arcuate and popliteofibular ligaments) was often well developed, the complex involved the inferior geniculate vascular sheath and popliteal fascia. A spectrum of variations was found in how thickly the complex developed. Thus, clear separation of the PT from the surrounding tissues often became difficult. Notably, the connective tissue complex could be peeled in different manners, depending on the site and the individual. Therefore, macroscopic variation in the posterolateral corner, including the apparent absence of some ligaments, seemed to be related to the thickness and number of layers developed by the connective tissue complex, possibly due to an adaptation to the individual's mechanical environment during growth and aging. PMID- 15666124 TI - Efficacy of novel-concept pedicle screw fixation augmented with calcium phosphate cement in the osteoporotic spine. AB - Pedicle screw instrumentation has become increasingly popular for rigid internal stabilization of the thoracolumbar spine. However, when pedicle screws are used in elderly osteoporotic patients, the screw-bone interface is stripped easily. Therefore, the risk of screw loosening and backing-out after surgery has increased. The purpose of this study was to evaluate the efficacy of the novel concept pedicle screw fixation augmented with calcium phosphate cement (CPC) in the osteoporotic spine. The novel-concept screw has the same shape as the ordinary screw, but it is hollow and fabricated with 20 small holes (1.3 mm in diameter) leading to the hollow part on the bottom of the thread. Fifteen embalmed cadaveric lumbar vertebrae were instrumented with two types of pedicle screw (the ordinary screw and the novel-concept screw) in each pedicle. Only the novel-concept screws were augmented with CPC after insertion. Seven days later, axial pull-out testing was performed at a crosshead speed of 10 mm/min. The mean maximal pull-out strength of the ordinary screws was 258 N, and that of the novel concept screws was 637 N. These results suggest that the novel-concept screw augmented with CPC can be useful for pedicle screw fixation of the osteoporotic spine. PMID- 15666125 TI - Gene expression of bone-resorbing cytokines in rat osteolysis model. AB - Osteolysis after single-injection polyethylene (PE) administration has been analyzed, but this experimental model did not reflect the clinical postoperative condition. Therefore, we investigated the bone-resorbing cytokines interleukin and tumor necrosis factor (IL-1alpha, IL-6, TNFalpha) produced by continuous PE administration using histomorphologic findings and the reverse transcription polymerase chain reaction. TNFalpha mRNA was continuously detected in the PE continuous-infusion group, whereas it was not expressed in the PE single injection group. IL-1alpha and IL-6 mRNA were detected in both groups. As for histomorphological findings, in the PE continuous-infusion group proliferating fibrous tissue penetrated subchondral bone and the interface membrane around the K-wire inserted into the femur. Numerous osteoclasts were observed in the vicinity of resorbed bone. In the PE single-injection group, proliferation of fibrous tissue with bone resorption was not observed. TNFalpha may play a major role in fibrous tissue formation and osteoclastic bone resorption caused by PE debris. This osteolysis model is useful for investigating osteolysis associated with bone-resorbing cytokines under conditions similar to those seen with a human prosthesis. PMID- 15666126 TI - Ovariectomy fails to augment bone resorption and marrow B lymphopoiesis in granulocyte colony-stimulating factor transgenic mice. AB - The mechanism of pathological bone loss induced by estrogen deficiency has not been fully elucidated. It has been shown in recent animal studies that increased B lymphopoiesis induced by estrogen deficiency is involved in the mechanism of stimulated bone resorption. Mice transgenic for granulocyte colony-stimulating factor (G-CSF) (G-Tg) exhibit generalized osteopenia with an increase in osteoclast number and enhancement of bone resorption, which coexists with enhanced hematopoiesis. When ovariectomy was performed on G-Tg, it did not further reduce bone mass as revealed by radiography, dual-energy X-ray absorptiometry, and peripheral quantitative computed tomography. Ovariectomy increased the amount of colony-forming units of interleukin 7 (CFU-IL-7) by threefold in the marrow of normal mice in association with an increase in the number of B220-positive cells expressing the receptor activator of nuclear factor kappaB ligand (RANKL). In contrast, the number of B220-positive cells expressing RANKL and CFU-IL-7 remarkably decreased in the marrow of G-Tg. Ovariectomy induced neither CFU-IL-7 nor B220-positive cells expressing RANKL in the marrow of G-Tg. Strong inhibition of B lymphopoiesis by G-CSF resulted in depletion of B cells expressing RANKL from the marrow, which may lead to resistance to bone loss due to ovariectomy. This observation suggests that B lymphopoiesis plays a possible role in bone loss in a condition of acute estrogen deficiency. PMID- 15666127 TI - Development of heating method by microwave for sterilization of bone allografts. AB - The purpose of this study was to develop a disinfection method using a microwave apparatus to treat large bone allografts. Heating of a bone allograft is an effective method for the disinfection of bacteria or inactivation of viruses. However, the size of the bone we can treat is limited, and following the popular method of using a bathtub is a lengthy process. The experimental system described here was designed using a microwave oven, an optical-fiber thermometer, and a power regulator. Large and small specimens, a femoral head, and a metatarsal were harvested from a bovine femur. The influence of size and the electrical or thermal characteristics of the specimens were assessed regarding temperature distribution after microwave irradiation. The effects of humidity or hot-air supply were also assessed. The average temperature of the bovine femoral head became 80 degrees C throughout the 15 min of microwave irradiation, although the temperature in the metatarsal did not attain uniformity. Microwave irradiation with a hot-air supply realized a uniform distribution of temperature at 83.0 degrees +/- 0.4 degrees C in the metatarsal within 15 min. Use of microwave irradiation enables quick heating for disinfection of large allograft bones when a hot-air supply was used as well. PMID- 15666128 TI - Differential sensitivity to NO-induced apoptosis between anterior cruciate and medial collateral ligament cells. AB - It is well known that the anterior cruciate ligament (ACL) of the knee joint has poorer healing responses than the medial collateral ligament (MCL). Nitric oxide (NO) induces free radicals and plays a key role in the induction of apoptosis in various wound-healing models. We hypothesized that the poor healing response of the ACL may be ascribed to high susceptibility to apoptosis, and we investigated the difference in susceptibility to apoptosis between ACL and MCL cells after treatment with sodium nitroprusside, a NO donor. Apoptosis was evaluated by phase contrast microscopy, electron microscopy, DNA gel electrophoresis, and flow cytometric analysis. Although morphological changes and DNA ladders were observed in both ACL and MCL cells after 2 mM sodium nitroprusside treatment, ACL cells were more prone to apoptosis at 1 mM. Based on flow cytometric analysis, DNA fragmentation at 1 mM sodium nitroprusside was significantly greater in ACL cells than in MCL cells (58.6% +/- 1.6% vs. 11.9% +/- 2.2%). Caspase-3 inhibitor (Ac Asp-Glu-Val-Asp-CHO) and caspase-9 inhibitor (Ac-Leu-Glu-His-Asp-CHO) completely inhibited this DNA fragmentation. In conclusion, the ACL and MCL cells exhibit essential differences, and the differential sensitivity to NO-induced apoptosis between the ACL and MCL cells may be a reflection of these differences. PMID- 15666129 TI - Extension of lumbar spine infection into osteoarthritic hip through psoas abscess. AB - We present a case of pyogenic lumbar discitis and septic hip arthritis, accompanied by a psoas abscess and pyogenic iliopsoas bursitis, for which the correct diagnosis was delayed. The patho-mechanism was speculated to be initial hematogenous infection in the lumbar spine that spread along the psoas muscle as a psoas abscess and then extended into the hip joint via the iliopsoas bursa. For an early correct diagnosis, clinicians should be aware that the lumbar spine and hip joint regions communicate through the psoas muscle space and iliopsoas bursa, making it possible for infection to spread. PMID- 15666130 TI - Arthroscopic treatment for septic arthritis of the shoulder in an infant. AB - We report a case of septic arthritis in the shoulder of an infant treated with a combination of arthroscopic irrigation, debridement, and synovectomy. The results were encouraging. The patient was a boy who was 2 years 6 months of age. His first symptoms were a body temperature of 40 degrees C and right upper arm pain. Five days after the onset, he was brought to our institution. His shoulder was swollen and erythematous. Yellowish fluid (7 ml) was aspirated from his shoulder joint; the causative organism was group A Streptococcus. On admission, arthroscopic surgery was performed. His temperature gradually decreased, and inflammatory markers including the white blood cell count and C-reactive protein level improved. At his 1-year follow-up there was no sign of infection, and a full range of motion in the shoulder was maintained. The interval between the onset and the start of treatment is an important factor affecting the prognosis. The patient was brought to our institution 5 days after onset and underwent arthroscopic surgery on the same day. This early treatment was one of the factors that contributed to a successful result. Arthroscopic surgery, including irrigation and debridement, may be one of the options for treating septic arthritis of an infant's shoulder. PMID- 15666131 TI - Incomplete scapholunate and trapeziotrapezoid coalitions with an accessory carpal bone. AB - Coalitions of the carpal bones associated with accessory carpal bones are relatively uncommon. We present a case of scapholunate and trapeziotrapezoid incomplete coalition with a dorsally situated accessory bone between trapezium and trapezoideum. X-ray, computed tomography, and magnetic resonance imaging findings of the patient are discussed in light of the pertinent literature. PMID- 15666132 TI - Radial head dislocation with acute plastic bowing of the ulna. AB - Five radial head dislocations with acute plastic bowing of the ulna in patients aged 6-12 years were reviewed. Closed reduction was successful in two, and open reduction was required in three patients in whom treatment was started more than 2 weeks after injury. In one child who presented 2 months after injury, realignment by osteotomy of the ulna as well as open reduction of the radial head was necessary. Follow-up evaluations at 6-24 months revealed good clinical outcomes in all patients. Awareness of this type of radial head dislocation is important to avoid delays in diagnosis and treatment. PMID- 15666133 TI - Solitary eosinophilic granuloma of the sternum. AB - We report a rare case of eosinophilic granuloma of the sternum in a 25-year-old woman, who presented with anterior chest pain and a tender mass over the sternum. Total-body bone scintigraphy and computed tomography scanning of the thorax revealed an isolated lytic lesion of the manubrium. An open biopsy showed the typical histologic appearance of an eosinophilic granuloma. Surgical curettage of the solitary lesion was performed, and the sternal defect was filled with a bone replacement material. At the 2-year follow-up, no local recurrence was found, and the patient was in good health. PMID- 15666134 TI - New strategies for disc repair: novel preclinical trials. AB - Degeneration of lumbar intervertebral discs is a major cause of low back complaints, an irreversible occurrence with no currently available treatment. Furthermore, various surgical procedures can accelerate disc degeneration. On the other hand, recent experimental studies on disc cells have demonstrated an important role for the nucleus pulposus in preserving overall disc structure. The author's group has already found that nucleus pulposus cells activated annulus fibrous cells, and reinsertion of nucleus pulposus cells slowed further disc degeneration. We have designed three subsequent studies that were designed to examine further possibilities for clinical transplantation: (1) activation of nucleus pulposus cells by mesenchymal stem cells; (2) focus on the multilineage differentiation potential of mesenchymal stem cells as an alternative cell source for cell transplantation therapy of disc degeneration; (3) the possibility of a human nucleus pulposus cell line as a cell source for cell transplantation therapy. Activation of nucleus pulposus could be achieved by co-culture with autogenous mesenchymal stem cells allowed to have direct cellular interaction. This would be a useful clinical cell source. Induction of nucleus pulposus cells by autogenous mesenchymal stem cells also would be an important subject for a clinical trial. Clinical application of the cells derived from a human nucleus pulposus cell line is an important project to be undertaken in the near future. PMID- 15666135 TI - The hyperbolic law--a 25-year perspective. PMID- 15666136 TI - [Pathological findings in tissue surrounding revised metal/metal articulations]. AB - INTRODUCTION: The aim of the current study was to elucidate the incidence of allergic reactions to metal/metal articulations in revised total hip arthroplasties. MATERIALS AND METHODS: Between 1 January 1997 and 31 January 2002 a consecutive series of tissue samples from 13 revised total hip arthroplasties with metal/metal articulations were histopathologically examined for signs of delayed type hypersensitivity (DTH). Mean age at the time of revision of the eight women and five men was 58.7 years. The prostheses were revised after a mean follow-up of 45 months. Indications for revision were progressive osteolysis of the proximal femur in 12 cases and instability in one case. All patients were clinically and radiologically evaluated after a mean follow-up of 52 months (min. 22, max. 74) after revision. RESULTS: No signs of infection were found in either histopathological or microbiological examinations. In ten cases, perivascular lymphocytic infiltrates could be found as a sign of DTH. After revision and changing of the articulation all osteolyses healed. CONCLUSION: In 10/13 cases (76.9%) signs of DTH could be detected. The fact that all osteolyses healed after changing the articulation may give a strong hint that there is an immunological contribution to this radiological changes. Metal/metal articulations cannot be recommended as the optimum implant for young patients, as the number of patients with allergic reactions to nickel, chrome or cobalt is increasing continuously. PMID- 15666137 TI - [Development of the spine after traumatic spinal cord injury in children and adolescents]. AB - In the 1970s, there was growing concern about the risk of secondary deformations of the spine as these seemed to endanger the otherwise improving prognosis for the life of paraplegic subjects, especially of paralyzed children and adolescents. According to the literature, the level and extent of the spinal cord injury and the age at the time of injury are determinants of the development of scoliosis, hyperlordosis or global kyphosis. Correction of the deformity by brace orthosis is not indicated except for children. The indications for surgical intervention in terms of the extent of the scoliosis and technical performance corresponds to the well known situation for idiopathic scoliosis except for the length of fusion. A special form of scoliosis, the so called "collapsing spine", allows good surgical correction because it is usually not rigid. Early, substantial degenerative processes such as segmental intervertebral instability at the level of the paraplegia, as well as distinct uncarthrosis proximally distant from the innervated zones with secondary radicular damage, are observed. By means of modern surgical procedures, the appearance of the patient's body, as well as the quality of life, can be favorably influenced. PMID- 15666138 TI - [Heterotopic ossification spinal cord injury. Management through early diagnosis and therapy]. AB - Heterotopic ossification is a frequent and potentially disastrous complication of acute spinal cord injury. Pathogenesis and etiology are not well described, initial clinical symptoms are uncharacteristic, specific laboratory findings do not exist. Between March 1997 and May 2000 all 290 patients admitted to our facility with acute spinal cord injury underwent standardized sonographic examinations of the soft tissue around the hip joint every three weeks, starting as early as two weeks after injury. In 12% of the patient population characteristic sonographic findings for heterotopic ossification were present while the regular x-ray examination was still unremarkable. Laboratory findings (alkaline phosphatase, C-reactive protein, anorganic phosphate) were unspecific. Clinical findings were present only in a few patients. All patients underwent radiotherapy consisting of the administration of 5 times 3 Gy to the area as soon as possible. Follow up demonstrated no progression of the heterotopic bone formation in these cases. In conclusion, regular ultrasound examination proved to be a secure, fast and reproducible method for the very early diagnosis of heterotopic ossification after acute spinal cord injury. PMID- 15666139 TI - Vacuum-assisted closure in the treatment of large skin defects due to necrotizing fasciitis. PMID- 15666141 TI - Withdrawal of feeding-tubes from incompetent patients: the Terri Schiavo case raises new issues regarding who decides in end-of-life decision making. PMID- 15666140 TI - One-year mortality of bloodstream infection-associated sepsis and septic shock among patients presenting to a regional critical care system. AB - OBJECTIVE: The long-term mortality outcome associated with sepsis and septic shock has not been well defined in a nonselected critically ill population. This study investigated the occurrence and the role of bloodstream infection (BSI) associated sepsis and septic shock at time of intensive care unit (ICU) admission on the 1-year mortality of patients admitted to a regional critical care system. DESIGN AND SETTING: Population-based inception cohort in all adult multidisciplinary and cardiovascular ICUs in the Calgary Health Region (population approx. 1 million) between 1 July 1999 and 31 March 2002. PATIENTS AND PARTICIPANTS: Adults (>/=18 years; n=4,845) who had at least one ICU admission to CHR ICUs. RESULTS: In 251 (5%) patients there was BSI-associated sepsis at presentation to ICU, and 159 of these also had septic shock. The 28 day, 90-day, and 1-year mortality rates overall were 18%, 21%, and 24%: 23%, 30%, and 36% for BSI-associated sepsis without shock, and 51%, 57%, and 61% with shock, respectively. Surgical diagnosis, BSI-associated sepsis, and increasing age were independently associated with late (28-day to 1-year) mortality whereas higher APACHE II and TISS scores were associated with reduced odds in logistic regression analysis. CONCLUSIONS: BSI-associated sepsis and septic shock are associated with increased risk of mortality persisting after 28-days up to 1 year or more. Follow-up duration beyond 28 days better defines the burden of illness associated with these syndromes. PMID- 15666142 TI - End-of-life decisions in The Netherlands: false euthanasia and false murder. PMID- 15666143 TI - Towards a fast-responding, label-free electrochemical DNA biosensor. AB - DNA sensors and sensor arrays (biochips) have become an important tool in molecular biology and biotechnology in recent years. For low-throughput, easy-to use devices it is desirable that they be of low cost, reagentless, and label free. Displacement sensors with electrochemical detection offer these advantages, and therefore the development of such a detection principle is show in this work. An HRP-labeled oligonucleotide was sub-optimally pre-hybridized with a capture probe and was displaced upon introduction of the fully complementary probe target, producing a decrease in signal that was proportional to the sample concentration. This detection scheme has been demonstrated colorimetrically and electrochemically, obtaining a total signal displacement of 55% only 5 min after introduction of the sample. PMID- 15666144 TI - Hydrogen production by Rhodobacter sphaeroides strain O.U.001 using spent media of Enterobacter cloacae strain DM11. AB - Combined dark and photo-fermentation was carried out to study the feasibility of biological hydrogen production. In dark fermentation, hydrogen was produced by Enterobacter cloacae strain DM11 using glucose as substrate. This was followed by a photo-fermentation process. Here, the spent medium from the dark process (containing unconverted metabolites, mainly acetic acid etc.) underwent photo fermentation by Rhodobacter sphaeroides strain O.U.001 in a column photo bioreactor. This combination could achieve higher yields of hydrogen by complete utilization of the chemical energy stored in the substrate. Dark fermentation was studied in terms of several process parameters, such as initial substrate concentration, initial pH of the medium and temperature, to establish favorable conditions for maximum hydrogen production. Also, the effects of the threshold concentration of acetic acid, light intensity and the presence of additional nitrogen sources in the spent effluent on the amount of hydrogen produced during photo-fermentation were investigated. The light conversion efficiency of hydrogen was found to be inversely proportional to the incident light intensity. In a batch system, the yield of hydrogen in the dark fermentation was about 1.86 mol H(2) mol(-1) glucose; and the yield in the photo-fermentation was about 1.5-1.72 mol H2 mol-1 acetic acid. The overall yield of hydrogen in the combined process, considering glucose as the preliminary substrate, was found to be higher than that in a single process. PMID- 15666145 TI - Construction of minimum size cellulase (Cel5Z) from Pectobacterium chrysanthemi PY35 by removal of the C-terminal region. AB - Pectobacterium chrysanthemi PY35 secretes the endoglucanase Cel5Z, an enzyme of the glycoside hydrolase family 5. Cel5Z is a 426 amino acid, signal peptide (SP) containing protein composed of two domains: a large N-terminal catalytic domain (CD; 291 amino acids) and a small C-terminal cellulose binding domain (CBD; 62 amino acids). These two domains are separated by a 30 amino acid linker region (LR). A truncated cel5Z gene was constructed with the addition of a nonsense mutation that removes the C-terminal region of the protein. A truncated Cel5Z protein, consisting of 280 amino acid residues, functioned as a mature enzyme despite the absence of the SP, 11 amino acid CD, LR, and CBD region. In fact, this truncated Cel5Z protein showed an enzymatic activity 80% higher than that of full-length Cel5Z. However, cellulase activity was undetectable in mature Cel5Z proteins truncated to less than 280 amino acids. PMID- 15666146 TI - Prediction of growth and biotransformation rates of hexahydro-1,3,5-trinitro 1,3,5-triazine (RDX) in the presence of barium. AB - The biotransformation of explosives has been investigated by many researchers. Bioremediation of soil and water contaminated with hexahydro-1,3,5-trinitro-1,3,5 triazine (RDX) is becoming the method of choice for clean-up of a variety of sites. In this study, we investigated biotransformation of RDX in the presence of barium. Ba is a metal commonly found in combination with RDX at sites requiring remediation. RDX was biotransformed by both a consortium of bacteria and an isolate from the consortium under anoxic conditions using a rich medium. However, Ba inhibited cell growth under both aerobic and anoxic conditions and slowed biotransformation rates by 40%. RDX and Ba inhibited growth of the isolate more than growth of the consortium. An additive inhibition model is proposed that accurately predicts the reduced growth rates observed. PMID- 15666147 TI - Fine measurement of ergosterol requirements for growth of Saccharomyces cerevisiae during alcoholic fermentation. AB - Yeasts can incorporate a wide variety of exogenous sterols under strict anaerobiosis. Yeasts normally require oxygen for growth when exogenous sterols are limiting, as this favours the synthesis of lipids (sterols and unsaturated fatty acids). Although much is known about the oxygen requirements of yeasts during anaerobic growth, little is known about their exact sterol requirements in such conditions. We developed a method to determine the amount of ergosterol required for the growth of several yeast strains. We found that pre-cultured yeast strains all contained similar amounts of stored sterols, but exhibited different ergosterol assimilation efficiencies in enological conditions [as measured by the ergosterol concentration required to sustain half the number of generations attributed to ergosterol assimilation (P(50))]. P(50) was correlated with the intensity of sterol synthesis. Active dry yeasts (ADYs) contained less stored sterols than their pre-cultured counterparts and displayed very different ergosterol assimilation efficiencies. We showed that five different batches of the same industrial Saccharomyces cerevisiae ADY exhibited significantly different ergosterol requirements for growth. These differences were mainly attributed to differences in initial sterol reserves. The method described here can therefore be used to quantify indirectly the sterol synthesis abilities of yeast strains and to estimate the size of sterol reserves. PMID- 15666148 TI - Pleading for the use of biodegradable polymers in favor of marine environments and to avoid an asbestos-like problem for the future. AB - Research results about the movement and accumulation of floating marine debris drifting throughout the world's oceans are reviewed in this paper. A mechanism for this accumulation and movement is strongly associated with surface currents consisting of the Ekman drift and the geostrophic current, because all floating marine debris is passive to surface currents. The basic published mechanism for the North Pacific is common across the world's ocean. After marine debris accumulates in the narrow Ekman convergence zone, it is moved to the east by geostrophic currents. The most important thing is that floating marine debris concentrates in some specific regions, independent of the initial quantity of marine debris. In order to resolve this problem and to avoid an asbestos-like problem, the use of biodegradable polymers is important in our daily life. PMID- 15666149 TI - Combined carbon and nitrogen removal from acetonitrile using algal-bacterial bioreactors. AB - When compared with Chlorella vulgaris, Scenedesmus obliquus and Selenastrum capricornutum, C. sorokiniana presented the highest tolerance to acetonitrile and the highest O(2) production capacity. It also supported the fastest acetonitrile biodegradation when mixed with a suitable acetonitrile-degrading bacterial consortium. Consequently, this microalga was tested in symbiosis with the bacterial culture for the continuous biodegradation of acetonitrile at 2 g l(-1) in a stirred tank photobioreactor and in a column photobioreactor under continuous illumination (250 microE m(-2) s(-1)). Acetonitrile removal rates of up to 2.3 g l(-1) day(-1) and 1.9 g l(-1) day(-1) were achieved in the column photobioreactor and the stirred-tank photobioreactor, respectively, when operated at the shortest retention times tested (0.4 days, 0.6 days, respectively). In addition, when the stirred-tank photobioreactor was operated with a retention time of 3.5 days, the microbial culture was capable of assimilating up to 71% and nitrifying up to 12% of the NH(4) (+) theoretically released through the biodegradation of acetonitrile, thus reducing the need for subsequent nitrogen removal. This study suggests that complete removal of N-organics can be combined with a significant removal of nitrogen by using algal-bacterial systems and that further residual biomass digestion could pay-back part of the operation costs of the treatment plant. PMID- 15666152 TI - [Ambiguous unilateral visual impairment with hyperopia]. PMID- 15666150 TI - Understanding the response to immunotherapy in humans. AB - Whether the efforts of the last decade aimed at the development of vaccines against tumor-specific antigens encountered success or failure is a matter of expectations. On the bright side, we could optimistically observe that anti cancer-vaccines stand as an outstanding example of the successful implementation of modern biotechnology tools for the development of biologically sound therapeutics. In particular, vaccines against melanoma (the prototype model of tumor immunology in humans) can reproducibly induce cytotoxic T cell (CTL) responses exquisitely specific for cancer cells. This achievement trespasses the specificity of any other anti-cancer therapy. The skeptics, on the other end, might point out that immunization only rarely leads to cancer regression, labeling, therefore, this approach is ineffective. In our opinion this judgment stems from the naive expectation that CTL induction is sufficient for an effective immune response. Here we propose that more needs to be understood about the mechanisms required for the induction of a therapeutically relevant immune response in humans. In particular, we will discuss the variables related to cancer heterogeneity, the weight of individual patients' polymorphism(s), the role of the T cell activation and differentiation and, finally, the complex relationship between immune and cancer cells within the tumor microenvironment. PMID- 15666153 TI - [The influence of transport on organ cultured corneas. An experimental study of porcine corneal endothelium]. AB - BACKGROUND: To investigate the influence of transport on the quality of corneas in organ culture, based on the vitality of the corneal endothelium. METHODS: Transport was simulated for 222 porcine corneal disks. These were placed in standard transportation containers filled with organ culture medium II, kept in an incubator and then shaken on a laboratory shaker. RESULTS: Agitated corneas at all acceleration rates always showed less endothelial cell damage than corneas kept motionless. The best condition was found after maximum accelerations of 0.10 g and 0.72 g. Damage consisted predominantly of disseminated cell loss and circular cell damage. A storage temperature of 37 degrees C caused most harm to the endothelium. CONCLUSION: The reduction in endothelial damage found in corneas after agitation can be attributed to a better distribution of nutrients in the transportation container. Assuming transferability, a better quality of human grafts can be achieved by inducing slight motions of corneas in organ culture. PMID- 15666151 TI - Tumor immunity via homeostatic T cell proliferation: mechanistic aspects and clinical perspectives. AB - Efforts to develop effective anti-tumor immunotherapies are hampered by the difficulty of overcoming tolerance against tumor antigens, which in most instances are normal gene products that are over-expressed, preferentially expressed or re-expressed in cancer cells. Considering that lymphopenia-induced homeostatic T cell proliferation is mediated by self-peptide/MHC recognition and that the expanded cells acquire some effector functions, we hypothesized that this process could be used to break tolerance against tumor antigens. Studies by us and others in several mouse models demonstrated that availability of tumor antigens during homeostatic T cell proliferation indeed leads to effective anti tumor autoimmunity with specificity and memory. This effect appears to be mediated by reduction in the activation threshold of low-affinity tumor-specific T cells, leading to their preferential engagement and expansion. In its simplicity, this approach is likely to have application in humans, since it relies on conventional lymphopenia-inducing cancer therapies, infusion of autologous lymphocytes and, optimally, tumor-specific vaccination. PMID- 15666154 TI - Can three-dimensional endoanal ultrasonography detect external anal sphincter atrophy? A comparison with endoanal magnetic resonance imaging. AB - PURPOSE: Anal sphincter atrophy is associated with a poor clinical outcome of sphincter repair in patients with faecal incontinence. Preoperative assessment of the sphincters is therefore relevant. External anal sphincter (EAS) atrophy can be detected by endoanal magnetic resonance imaging (MRI), but not by conventional endoanal ultrasonography (EUS). Three-dimensional EUS allows multiplanar imaging of the anal sphincters and thus enables more reliable anal sphincter measurements. The aim of the present study was to establish whether 3D EUS measurements can be used to detect EAS atrophy. For this purpose 3D EUS measurements were compared with endoanal MRI measurements. METHODS: Patients with symptoms of faecal incontinence underwent 3D EUS and endoanal MRI. Internal anal sphincter (IAS) and EAS defects were assessed on 3D EUS and endoanal MRI. EAS atrophy was determined on endoanal MRI. The following measurements were performed: EAS length, thickness and area. Furthermore, EAS volume was determined on 3D EUS and compared with EAS thickness and area measured on endoanal MRI. RESULTS: Eighteen parous women (median age 56 years, range 32-80) with symptoms of faecal incontinence were included. Agreement between 3D EUS and endoanal MRI was 61% for IAS defects and 88% for EAS defects. EAS atrophy was seen in all patients on endoanal MRI. Correlation between the two methods for EAS thickness, length and area was poor. In addition, correlation was also poor for EAS volume determined on 3D EUS, and EAS thickness and area measured on endoanal MRI. CONCLUSION: Three-dimensional EUS and endoanal MRI are comparable for detecting EAS defects. However, correlation between the two methods for EAS thickness, length and area is poor. This is also the case for EAS volume determined on 3D EUS and EAS thickness and area measured on endoanal MRI. Three-dimensional EUS can be used for detecting EAS defects, but no 3D EUS measurements are suitable parameters for assessing EAS atrophy. PMID- 15666157 TI - Lactic acidosis caused by sodium nitroprusside in a newborn with congenital heart disease. PMID- 15666156 TI - The developmental relationship between the deciduous dentition and the oral vestibule in human embryos. AB - In humans, there is no consensus about the developmental relationship between the deciduous dentition and the oral vestibule separating the teeth from the lips and cheeks. The classical concept assumes that two horseshoe-shaped epithelial structures exist: the dental lamina, giving rise to single tooth primordia, and the vestibular lamina running parallel and externally to it, giving rise to the oral vestibule. The aim of this study was to investigate the development of the dental and vestibular laminae in the upper jaw and to determine their developmental relationship in humans from embryonic week 6 to 9. Although a thickening of the vestibular epithelium was always present on serial histological sections, computer-aided three-dimensional reconstructions did not show any continuous vestibular lamina. Several discontinuous epithelial structures (bulges and ridges) occurred transiently at different stages of oral vestibule development. Along the mesiodistal axis, the dental and vestibular epithelia were regionalized in parallel: in the incisive, canine, and 1st and 2nd molar regions. The vestibular ridges fused with the dental lamina distally to the deciduous canine, 1st molar and 2nd molar. These interactions between the developing teeth and vestibular structures are reminiscent of the situation in some reptiles, where single teeth are paired one-to-one with single tooth glands. PMID- 15666155 TI - Low temperature promotes intron retention in two e-cor genes of durum wheat. AB - Following the screening of a suppression subtractive library developed from durum wheat plants exposed to low temperature for 6 h, two early cold-regulated (e-cor) genes have been isolated. These genes, coding putatively for a ribokinase (7H8) and a C3H2C3 RING-finger protein (6G2), were characterized by the stress-induced retention of a subset of introns in the mature mRNA. This feature was dependent on cold for 7H8 and on cold and dehydration for 6G2. When other genes, such as the stress-related gene WCOR410c, coding for a dehydrin (one intron), or a gene coding for a putative ATP binding cassette transporter (16 introns) were analyzed, no cold-dependent intron retention was observed. Cold-induced intron retention was not observed in mutants defective in the chloroplast development; nevertheless treatment with cycloheximide in the absence of cold was able to promote intron retention for the 7H8 e-cor gene. These results suggest that the cold-induced intron retention reflects the response of the spliceosoma to specific environmental signals transduced to the splicing protein factors through a chloroplast-dependent pathway. Notably, when the 7H8 Arabidopsis orthologous gene was analyzed, no stress induction in terms of mRNA abundance and no cold dependent intron retention was detected. Otherwise, 6G2 Arabidopsis homologous sequences sharing the same genomic structure of the durum wheat 6G2 showed a similar intron retention event although not strictly dependent on stress. PMID- 15666158 TI - Isolated myocardial non-compaction in an infant with distal 4q trisomy and distal 1q monosomy. PMID- 15666159 TI - Vascular rings--presentation, investigation and outcome. AB - Our aim was to determine the presentation of patients with vascular rings and evaluate the effectiveness of investigations. Surgical outcomes and respiratory sequelae were also examined. The design was a retrospective case note study over a 13-year period set in a tertiary children's hospital. Children below the age of 16 years presenting with a vascular ring to the Royal Hospital for Sick Children, Glasgow were studied. Demographic data at presentation, including symptoms, were recorded. The ability of diagnostic investigations to identify the presence of a vascular ring was evaluated. Surgical outcomes were determined by measuring surgical complications and mortality. Respiratory sequelae were recorded by the presence of persistent symptoms or the need for tracheostomy or long-term ventilation following surgery. A total of 24 patients were identified with a median age at presentation of 4.5 months. Stridor was the commonest presenting symptom (14/24). Angiography, chest CT scanning and MRI were the most accurate imaging modalities (accurate in 100% of cases used). Chest X-ray films and echocardiography had the lowest detection rates. Surgical complications (4/24) and mortality (1/24) were low. A substantial number of patients available to follow-up (7/20) were still experiencing stridor 3 months post-operatively. CONCLUSION: Vascular rings are rare, however, often present with common symptoms. Most children present in early infancy, but a minority presents much later. The investigation of choice is a barium swallow followed by high-resolution computed tomography. Surgery is safe although a number of patients will have persisting symptoms. PMID- 15666160 TI - Multidrug-resistant tuberculosis in Russia: clinical characteristics, analysis of second-line drug resistance and development of standardized therapy. AB - The aim of the study presented here was to identify patients with multidrug resistant tuberculosis (MDRTB) in the Samara region of Russia and to analyze the susceptibility of their isolates to second-line drugs in order to develop an empirical, standard, second-line treatment regimen. Treatment of MDRTB can be individualized based on in vitro laboratory analysis or standardized. In the latter case there is still a need to ascertain local second-line drug-resistance patterns. Described here are the clinical characteristics of 251 MDRTB patients identified in the study and the second-line drug susceptibility of 69 MDRTB isolates obtained from them. Antimicrobial resistance to the following agents was detected in the isolates: rifabutin (88.2%), streptomycin (42.8%), amikacin (7.2%), doxycycline (7.4%), ciprofloxacin (4.3%), clofazimine (2.9%), cycloserine (7.4%), and prothionamide (1.5%). The results of the study indicate it is possible to develop a standard, effective, clinical treatment regimen using ethambutol, pyrazinamide, prothionamide, a fluoroquinolone and amikacin. PMID- 15666161 TI - Position-dependent sensitivity and density of taste receptors on the locust leg underlies behavioural effectiveness of chemosensory stimulation. AB - Chemical stimulation of contact chemoreceptors located on the legs of locusts evokes withdrawal movements of the leg. The likelihood of withdrawal depends on the site of stimulation, in addition to the identity and concentration of the chemical stimulus. A significantly higher percentage of locusts exhibit leg avoidance movements in response to stimulation of distal parts of the leg with any given chemical stimulus compared to proximal sites. Moreover, the percentage of locusts exhibiting avoidance movements is correlated with the density and sensitivity of chemoreceptors on different sites of an individual leg. The effectiveness of chemical stimulation also differs between the fore and hind legs, with NaCl evoking a higher probability of leg withdrawal movements on the foreleg. Moreover, sucrose was less effective than NaCl at evoking withdrawal movements of the foreleg, particularly at low concentrations. The gradients in behavioural responses can be partially attributed to differences in the responsiveness and density of the contact chemoreceptors. These results may reflect the different specialization of individual legs, with the forelegs particularly involved in food selection. PMID- 15666162 TI - Analogues of virus resistance genes map to QTLs for resistance to sharka disease in Prunus davidiana. AB - Plum pox virus (PPV), the causative agent of sharka disease in Prunoideae, is one of the most serious problems affecting stone fruit production in Europe and America. Resistance to PPV was previously described in a Prunus davidiana clone, P1908, and introduced into peach (Prunus persica) genotypes. Genetic resistance to PPV displays a complex pattern of quantitative inheritance. An analysis of quantitative trait loci (QTLs) for resistance was performed on an F1 interspecific peach population obtained from a cross between the susceptible nectarine cultivar Summergrand and P. davidiana. The hybrids were graft inoculated with PPV in duplicate following a classical procedure. The incidence of infection was evaluated four times, over two vegetative cycles, by symptom observation and enzyme-linked immunoadsorbent assays (ELISA). Restriction of systemic downward movement of the PPV virus was also evaluated by testing the susceptible rootstocks. Using both analysis of variance and non-parametric tests, six genomic regions involved in PPV resistance were detected. Depending on the scoring data considered, between 22 and 51% of the phenotypic variance could be explained by the quantitative model. One QTL, located in the distal region of linkage group 1, maps in a genomic region that is syntenic to the location of a resistance gene previously identified in the apricot cv. Goldrich. Some QTLs appeared to be temporally specific, reflecting the environmental dependence of PPV-resistance scoring. Candidate gene fragments were amplified by PCR, isolated and mapped on the peach interspecific linkage map. We report here the co localization of three analogues of virus resistance genes with two distinct genomic regions linked to PPV resistance in P. davidiana. PMID- 15666164 TI - Prevalence of C1/C2 involvement in Czech rheumatoid arthritis patients, correlation of pain intensity, and distance of ventral subluxation. AB - The aim of this study was to determine the prevalence of C1/C2 involvement in rheumatoid arthritis (RA) in Czech patients and to identify typical pain symptoms and their relationship to radiologic findings at the C1/C2 level. Four hundred patients with RA were selected randomly and examined by plain X-ray. Cervical spine involvement was found in 45.8% of these patients. Cervicocranial syndrome was the most common symptom of any spine involvement at the C1/C2 level and was present in 54.6%. Cervicocranial syndrome was typical for ventral subluxation 3-6 mm and was found in 52.9%. The distance of 8 mm or more was associated with mild pain. The pain intensity at the C1/C2 level decreased with increasing distance of ventral atlantoaxial subluxation (P < 0.0001). PMID- 15666163 TI - Crosstalk between on-line and off-line processing of visual features. AB - Three dual-task experiments were conducted to investigate the relation between immediate, "on-line" judgments about visual features and delayed, "off-line" judgments. One hypothesis ("selective interference") predicted that dual-task performance would be challenged specifically within a visual dimension, as both tasks compete for the same resources. Another hypothesis ("cost of switching") made the opposite prediction. In Experiment 1, participants performed either color or shape discriminations in the on-line and off-line visual tasks, with systematic variation of feature similarity between the on-line and off-line features. In Experiment 2, participants performed either color or shape discriminations in the off-line task and color discriminations in the on-line task, with no overlap between the on-line and off-line features. In Experiment 3, participants performed color discriminations in both the on-line and off-line tasks, with partially overlapping stimulus sets. Altogether, the data from the three experiments provided evidence in favor of the hypothesis of cost of switching. Stimulus-stimulus compatibility effects between features in the off line task and those in the on-line task further underscored the perceptual nature of the crosstalk. PMID- 15666165 TI - FPF1 transgene leads to altered flowering time and root development in rice. AB - AtFPF1 (FLOWERING PROMOTING FACTOR 1) is a gene that promotes flowering in Arabidopsis. An expression vector containing AtFPF1 driven by a Ubi-1 promoter was constructed. The gene was introduced into rice callus by Agrobacterium mediated transformation and fertile plants were obtained. The presence of AtFPF1 in rice plants was confirmed by PCR, Southern and Northern blot analyses, as well as by beta-glucuronidase assay. The results showed that, as in Arabidopsis, AtFPF1 reduced flowering time in rice. Furthermore, introduction of AtFPF1 enhanced adventitious root formation but inhibited root growth in rice during the seedling stage. The results suggest that AtFPF1 promotes flowering time in both dicots and monocots, and plays a role in the initiation of adventitious roots in rice. PMID- 15666166 TI - Agrobacterium-mediated transformation of Solanum phureja. AB - A population of transgenic plants was produced by the transformation of internodal explants of Solanum phureja, DB337/37 (the cultivar Mayan Gold) using an Agrobacterium tumefaciens LBA4404-based vector containing a phytoene synthase gene (crtB). The regeneration strategy utilised a two-step protocol, with a 12 day callus induction stage mediated by 1.07 microM alpha-napthaleneacetic acid (NAA), 7.10 microM zeatin riboside and 0.06 microM gibberellic acid (GA3), followed by a prolonged (up to 90 day) shoot induction stage on medium containing 0.11 microM NAA, 7.10 microM zeatin riboside and 0.06 microM GA3 supplemented with kanamycin at 50 mg l(-1) as the selection agent. Southern analysis of the transgenic population revealed that the transgene copy number varied between one and five in the lines tested. Northern blot analysis showed significant expression of the introduced crtB gene in some lines during tuber development. Cytological analysis of the material showed a high incidence of chromosome doubling in the transgenic population with over 80% of all lines tested having doubled their chromosome complement during the transformation process. PMID- 15666167 TI - [Botulism caused by consumption of smoked salmon]. AB - Fresh fish products are exquisitely prone to be the source for food botulism because of primary contamination with Clostridium botulinum spores, lack of heat inactivation of C. botulinum spores, frequently used air-tight packaging, lack of preservatives, lack of heat inactivation of already produced botulinum toxin and because of substantial low temperature resistance of C. botulinum type E typically found in fish botulism. We can reporting a 25-year-old patient with food botulism presenting with an anticholinergic syndrome with mydriasis, accommodation difficulties, dryness of mouth, reduced sweating, constipation and reduced heart rate variance and only discrete involvement of striate muscles with ptosis, diplopia and generalised fatigue, all of which developed 12 h after the consumption of airtight sealed smoked salmon. The salmon was consumed 3 days after the 'use by' date had expired. PMID- 15666169 TI - [Early diagnosis of language development disorders in children]. AB - In a prospective study, we examined whether it is possible to diagnose language disorders in children in their second and third years. Twenty patients aged 17-35 months (average 26 months) were examined from August 2002 to June 2004. Language development was tested on the basis of a questionnaire for parents (ELFRA-1 or ELFRA-2) and on a detailed language assessment. After 3-8 months, the children's language was examined again by the same speech pathologist. In 12 children, in whom we had suspected a language disorder, the diagnosis was confirmed. Eight patients showed age-appropriate language development, which was confirmed in six patients at follow-up. In two cases, the follow-up revealed a language disorder. On the basis of a parent questionnaire and examination by a speech therapist, language disorders can be diagnosed in the second and third years of life. However, control assessments are necessary in order to reveal language development disturbances beginning at later ages. PMID- 15666168 TI - [Vagus nerve stimulation, repetitive transcranial magnetic stimulation, and electroconvulsive therapy in the treatment of depressive disorders]. AB - Affective disorders, especially major depression, are the most common psychiatric disorders. Although well treatable, a number of patients do not or do not sufficiently respond to antidepressant pharmacotherapy. Therefore there is a need for safe and efficient alternative therapeutic strategies. Neurostimulatory therapies such as electroconvulsive therapy, repetitive transcranial magnetic stimulation, and vagus nerve stimulation belong to these alternatives. In this article we review their mechanisms of action and summarize efficacy and adverse effects. PMID- 15666170 TI - In vivo antitumor activity by 2',4'-dihydroxy-6'-methoxy-3',5'-dimethylchalcone in a solid human carcinoma xenograft model. AB - Previously, we showed that 2',4'-dihydroxy-6'-methoxy-3',5'-dimethylchalcone (DMC), isolated from the buds of Cleistocalyx operculatus, significantly inhibited the growth of human liver cancer SMMC-7721 cells and could induce SMMC 7721 cells apoptosis in vitro. Here, we report the antitumor effects of DMC in vivo, using a solid human tumor xenograft model with a human liver cancer SMMC 7721 cell line. Our results revealed that the average tumor weight in a control group and a 150-mg/kg DMC injection group was 1.42+/-0.11 g and 0.59+/-0.12 g, respectively. Flow cytometric analysis of the tumor cell population demonstrated the existence of an aneuploid peak (representing 33.60+/-0.80% of the total in the 150-mg/kg DMC injection group). To our knowledge, this is the first time that chalcone compounds were applied to a human tumor xenograft model. PMID- 15666171 TI - The role of bisphosphonates in hormone-refractory prostate cancer. AB - Men with advanced prostate cancer are at high risk for developing bone metastases, which result in clinically significant skeletal morbidity. Treatments that prevent skeletal complications in these patients could considerably improve their quality of life. Therefore, this paper reviews the role of bisphosphonates in the treatment of hormone-refractory prostate cancer (HRPC). Published studies were identified through MEDLINE searches, review of bibliographies of relevant articles, and review of abstracts from scientific meetings. Metastatic bone disease in men with HRPC results in skeletal complications such as pathologic fractures, spinal cord compression, and debilitating bone pain. First- and second generation bisphosphonates, clodronate and pamidronate, had transient palliative effects that were not durable and did not prevent skeletal events in these patients. A small open-label study of ibandronate demonstrated significant reductions in pain, but these results have not been confirmed in a larger, randomized, controlled trial. To date, zoledronic acid is the only bisphosphonate that has demonstrated a statistically significant reduction in skeletal morbidity in this patient population. In a large, multicenter, randomized, placebo controlled trial, treatment of men with HRPC and bone metastases with zoledronic acid significantly reduced skeletal-related events and provided a durable reduction of bone pain over 24 months compared with placebo. Zoledronic acid is the only bisphosphonate that has demonstrated efficacy for preventing skeletal complications in patients with HRPC and bone metastases. Therefore, initiation of zoledronic acid therapy should be considered to prevent skeletal morbidity and improve the quality of life of these patients. PMID- 15666172 TI - Antipsychotic prescribing pattern among Italian general practitioners: a population-based study during the years 1999-2002. AB - OBJECTIVES: To assess the antipsychotic use and the prevalence/incidence of antipsychotic drug users in Italy during the years 1999-2002. To estimate the persistence with antipsychotic medications, and to measure their off-label use. METHODS: We selected 465,061 individuals registered by June 2002 in the lists of 320 general practitioners, homogeneously distributed throughout Italy, from the Health Search Database. We measured the antipsychotic drug consumption, calculated as defined daily dose (DDD) per 1000 inhabitants per day. We also calculated the number of individuals receiving at least one antipsychotic prescription, to estimate the annual prevalence and incidence of antipsychotic users. Among incident users, we evaluated the percentage of patients adherent to drug label indications and the average duration of treatment, estimated as medical possession ratio (MPR). RESULTS: Atypical antipsychotic use has continuously increased from 1999 to 2002. Women, older people and patients affected by psychotic disorders other than schizophrenia were seen to be more likely to receive antipsychotic prescriptions. Persistence with atypical drug treatment (MPR=0.213 in 2002) appeared longer than that of typical drugs (0.169). The percentage of patients adherent to drug label indications was significantly higher among typical antipsychotic users (P<0.001). The most common off-label use for atypical drug was senile dementia. CONCLUSION: Atypical drug use has been continuously expanding over the years 1999-2002, particularly in older people with dementia. The rapidly increasing use of this new class of antipsychotics highlights the need for a better evaluation regarding their safety profile and a better definition of their role in psychiatric treatments. PMID- 15666173 TI - Milk thistle and indinavir: a randomized controlled pharmacokinetics study and meta-analysis. AB - OBJECTIVES: To determine whether ingestion of milk thistle affects the pharmacokinetics of indinavir. METHODS: We conducted a three-period, randomized controlled trial with 16 healthy participants. We randomized participants to milk thistle or control. All participants received initial dosing of indinavir, and baseline indinavir levels were obtained (AUC(0-8)) (phase I). The active group were then given 450 mg milk-thistle extract capsules to be taken t.i.d. from day 2 to day 30. The control group received no plant extract. On day 29 and day 30, indinavir dosing and sampling was repeated in both groups as before (phase II). After a wash-out period of 7 days, indinavir dosing and sampling were repeated as before (phase III). RESULTS: All participants completed the trial, but two were excluded from analysis due to protocol violation. There were no significant between-group differences. Active group mean AUC(0-8) indinavir decreased by 4.4% (90% CI, -27.5% to -26%, P=0.78) from phase I to phase II in the active group, and by 17.3% (90% CI, -37.3% to +9%, P=0.25) in phase III. Control group mean AUC(0-8) decreased by 21.5% (90% CI, -43% to +8%, P=0.2) from phase I to phase II and by 38.5% (90% CI, -55.3% to -15.3%, P=0.01) of baseline at phase III. To place our findings in context, milk thistle-indinavir trials were identified through systematic searches of the literature. A meta-analysis of three milk thistle-indinavir trials revealed a non-significant pooled mean difference of 1% in AUC(0-8) (95% CI, -53% to 55%, P=0.97). CONCLUSIONS: Indinavir levels were not reduced significantly in the presence of milk thistle. PMID- 15666174 TI - [Medieval scenes of ritual circumcision as a reflection of sociopolitical circumstances]. AB - Ritual circumcision in males is regarded as one of the oldest surgical procedures. Whereas their medieval illustrations are mostly interpreted within a religious context, this study tries to prove the influence of the political and social situation of the above-mentioned period.Selected iconography of ritual male circumcision in the Middle Ages from Germany, France, Italy, and the Byzantine Empire was critically examined. Special attention was paid to the stained glass windows recently returned to St. Mary's Church in Frankfurt/Oder, where circumcision of the so-called Antichrist is also shown. Up to now we could not find any medical historical information about this subject. Clerical fine art of the fifteenth and sixteenth centuries shows more frequently than before illustrations of ritual circumcision, which evidently demonstrate the political, economic, and social tensions and controversies of that period. In many cases this iconography indicates a rejection of this old Jewish tradition and its confessors. Also the stained glass image of the Antichrist posthetomy could be interpreted as criticism or aversion.This study expands our approach to medieval illustrations of ritual circumcision that have hitherto mostly been interpreted in religious terms. The influence of changing political and social situations in the Middle Ages is evident. PMID- 15666175 TI - Direct targeting and rapid isolation of BAC clones spanning a defined chromosome region. AB - To isolate genes of interest in plants, it is essential to construct bacterial artificial chromosome (BAC) libraries from specific genotypes. Construction and organisation of BAC libraries is laborious and costly, especially from organisms with large and complex genomes. In the present study, we developed the pooled BAC library strategy that allows rapid and low cost generation and screening of genomic libraries from any genotype of interest. The BAC library is constructed, directly organised into a few pools and screened for BAC clones of interest using PCR and hybridisation steps, without requiring organization into individual clones. As a proof of concept, a pooled BAC library of approximately 177,000 recombinant clones has been constructed from the barley cultivar Cebada Capa that carries the Rph7 leaf rust resistance gene. The library has an average insert size of 140 kb, a coverage of six barley genome equivalents and is organised in 138 pools of about 1,300 clones each. We rapidly established a single contig of six BAC clones spanning 230 kb at the Rph7 locus on chromosome 3HS. The described low-cost cloning strategy is fast and will greatly facilitate direct targeting of genes and large-scale intra- and inter-species comparative genome analysis. PMID- 15666176 TI - Moderate hypoxia does not affect the zone of thermal comfort in humans. AB - The zone of thermal comfort was determined during normoxia and hypoxia in 15 healthy normothermic young subjects. Subjects dressed only in shorts/shorts and bikini top donned a water-perfused suit and assumed a supine position on a bench. The ambient temperature was maintained at a mean (SD) of 25.7 (0.3) degrees C. The thermal comfort zone was determined by increasing the temperature of the water perfusing the suit from cool to warm. During the heating process, subjects were instructed to report when their perception of the thermal stimulus provided by the suit changed from unpleasant to pleasant, and again from pleasant to unpleasant. The boundaries of the thermal comfort zone were assumed to be the temperatures of the water perfusing the suit at the time the subjects reported a change in the affective component of their thermal perception. In normoxia, subjects inspired room air and in hypoxia a gas mixture containing 10% O(2) in N(2). Tympanic temperature was similar in the normoxia and hypoxia conditions (P>0.05). The average (SD) lower and upper limits of the thermal comfort zone were 30.5 (1.5) and 34.7 (3.3) degrees C, respectively, during normoxia, and 30.5 (1.7) and 35.1 (3.4) degrees C, respectively, during hypoxia. No significant differences were observed between the normoxia and hypoxia conditions (P>0.05). Also, no gender-related differences were observed in the characteristics of the thermal comfort zone. The results of the present study indicate that acute hypoxic exposure simulated in the present study does not affect the zone of thermal comfort in humans. PMID- 15666177 TI - Comparison of D&C and hysterectomy pathologic findings in endometrial cancer patients. AB - OBJECTIVE: The objective of this study is to compare the accuracy of tumor grade in endometrial cancer between fractional dilatation and curettage (D&C) and postoperative hysterectomy specimen findings. METHODS: From January 2000 to November 2002, 52 women with abnormal vaginal bleeding and diagnosed or suspected endometrial cancer were treated in the Department of Obstetrics and Gynecology, Renji Hospital, Shanghai Second Medical University. The comparison of tumor grades was performed on both D&C and hysterectomy specimens, the relationship between the concordance rate of grade, and the depth of tumor invasion into the myometrium. The expression of estrogen receptor (ER), progesterone receptor (PR), p53, Bcl2, and proliferation cell nuclear antigen (PCNA) in tumor samples were analyzed. RESULTS: The concordance rates were 20% in grade 1, 61.5% in grade 2, and 77.8% in grade 3. The concordance rates for grade 2 and grade 3 were higher than grade 1 significantly (grade 2 approximately grade 1, P=0.010; grade 3 approximately grade 1, P=0.005). Fourteen out of 52 (26.9%) patients diagnosed with atypical endometrial hyperplasia by D&C had their diagnosis changed to endometrial cancer after being ascertained by hysterectomy specimen. The accuracy of tumor grade diagnosis by D&C was 50%, and 48% of patients were downgraded after comparison of postoperative specimens. The concordance rate of tumor invasion into the outer half of myometrium was higher than in tumors localized in the endometrium (P= 0.030). No association between high concordance rate and the expression of ER, PR, p53, Bcl-2 and PCNA was found. CONCLUSION: We concluded that D&C had a high accuracy in the diagnosis of endometrial cancer in grade 3 tumors (77.8%) and tumor invasion into the outer half of the myometrium (75%). Overall, 48% of endometrial cancer patients' tumor grades seemed to be decreased by D&C evaluation; thus, the diagnosis of endometrial disorders by D&C should not be overlooked. PMID- 15666178 TI - Hemispherotomy: description of surgical technique. AB - INTRODUCTION: Hemispherectomy constitutes an established surgical method in the management of patients with medically intractable epilepsy, secondary to severe unilateral hemisphere damage. The well-established association of the anatomical hemispherectomy initially described with severe complications such as late hydrocephalus has led to the development of less resective and more disconnecting procedures. All these technical variations of hemispherotomy carry less favorable outcomes compared with anatomic hemispherectomy, but significantly fewer complications. METHODS: In our current communication, we outline the indications and the surgical technique of hemispherotomy and report our experience of the clinical application of this surgical procedure. RESULTS: In our clinical series, the 5-year follow-up shows that 66.6% of our patients (6 out of 9) had class I outcome according to Engel's classification system, 22.2% (2 out of 9) class II outcome, while 11.1% (1 out of 9) had class III outcome. No mortality occurred in the current series and operative blood loss was significantly lowered. CONCLUSION: Hemispherotomy represents a less efficacious technique compared with anatomic hemispherectomy, but is a safe and technically simple surgical alternative for the management of patients with medically intractable seizures. PMID- 15666179 TI - Bilateral abducent and facial nerve palsies following fourth ventricle shunting: two case reports. AB - CASE REPORTS: Treatment of isolated fourth ventricle syndrome is difficult and there is no widely agreed method. Fourth ventriculo-peritoneal shunting is the most commonly utilized procedure for the management of this syndrome. Complications from shunting are common and are usually related to malfunction, infection, dislocation and overdrainage. We present two unusual cases in which both patients developed bilateral abducens and facial nerve palsies following shunting of an isolated fourth ventricle. Magnetic resonance imaging (MRI) in both cases revealed collapse of the fourth ventricles with downward displacement of the brain stem. In the first case the trans-tentorial pressure difference was equilibrated with the aid of a "Y" connector between the supratentorial and infratentorial shunts, with full recovery of the neurological deficits; in the second case this approach failed and following a complicated neurosurgical course successful endoscopic aqueductal stenting was performed. DISCUSSION: Pathogenesis of cranial nerve palsies following fourth ventricle shunting and the rationale of treatment are discussed and the literature is reviewed. PMID- 15666181 TI - The role of alpha-synuclein in the pathogenesis of multiple system atrophy. AB - The discovery of glial cytoplasmic inclusions (GCIs) in 1989 helped to define multiple system atrophy (MSA) as a clinicopathological entity, and drew attention to the prominent role played by these inclusions in the pathogenesis of the disorder. Subsequently, GCIs were shown to be highly positive for alpha synuclein, a neuronal protein that is normally absent in oligodendroglia except during embryonic development. The source of oligodendroglial alpha-synuclein aggregation in MSA is unknown. Since genetic overexpression has been excluded, active uptake from dying neurons remains a possibility. The similar topography of oligodendroglial and neuronal pathology in MSA suggests a fundamental disturbance of the functional unit between oligodendroglia, axon, and neuron. Transgenic MSA mouse models are now available to determine these aspects of cellular disturbance experimentally. PMID- 15666180 TI - Contractile properties of inner and outer smooth muscle bundles from pig urinary detrusor. AB - Like in the human detrusor, the pig urinary detrusor muscle consists of two layers: compactly arranged smooth muscle bundles on the mucosal side (inner layer) and loosely arranged smooth muscle bundles on the serosal side (outer layer). The contractile properties of muscle bundles of both layers were measured using the stop test followed by an isometric contraction. Total and passive forces were measured in ten muscle bundles from the inner and outer muscle layers. Active force was defined as the difference between total and passive force. The curvature and the unloaded shortening velocity of the force-velocity relation were calculated from the shortening forces measured during the stop test. The rate of force development was calculated from the isometric contraction. Differences in contractile properties between both layers were pairwise tested using the Wilcoxon Signed Ranks test. Percentage wise, the outer layer muscle bundles produced the highest active isometric force. The shortening forces were also higher in the outer layer bundles. As a result, both the curvature and the unloaded shortening velocity, derived from the average force velocity relations fitted to the data sets, were higher in the muscle bundles from this layer. Finally, the outer layer muscle bundles contracted significantly faster than those of the inner layer. Muscle bundles from the outer layer of pig detrusor were found to be faster and stronger (more phasic) than the weaker and slower (more tonic) bundles from the inner layer, suggesting that during bladder contraction the outer layer of the detrusor does more work than the inner layer. PMID- 15666185 TI - Intra-strain clonal phenotypic variation of Tritrichomonas foetus is related to the cytotoxicity exerted by the parasite to cultured cells. AB - As observed in most of the investigated trichomonads, a strain of Tritrichomonas foetus includes different parasite subpopulations. Such population diversity might account for important properties such as the ability of the parasite to destroy host cells. The aim of this study was to characterize the cytotoxicity exerted by subpopulations (named as K1, K2, K3, K4 and K5) of an isolate of T. foetus on epithelial cultured cells. The five populations studied here destroyed epithelial monolayers at different rates (from 25% to 55%), even though the cytoadhesion level and whole-cell protease activity were closely related among them. We were also able to detect differences in contact-dependent and contact independent cytotoxicity mechanisms among the five populations. An extracellular parasite protease had varying activity among the parasite populations. The intensity of contact-independent cytotoxicity was strictly related to the degree of enzyme activation, suggesting that such a protease might be involved in the cytotoxicity mediated by T. foetus. PMID- 15666187 TI - Spinal intramedullary ependymal cyst: a case report and review of the literature. AB - We report a rare case of spinal intramedullary ependymal cyst in a 44-year-old female and reviewed 12 cases reported in the literature. The patient presented with slowly progressive lower limb paresis. She underwent biopsy of the cyst wall and placement of a cysto-subarachnoid shunt with complete recovery at the follow up examination 18 months after surgery. This is a benign lesion and appropriate management should be performed at an early stage of the disease. PMID- 15666186 TI - A molecular analysis of the subgenus Transphlebotomus Artemiev, 1984 (Phlebotomus, Diptera, Psychodidae) inferred from ND4 mtDNA with new northern records of Phlebotomus mascittii Grassi, 1908. AB - A comparative molecular study was performed on the three species of Phlebotomine sandflies belonging to the subgenus Transphlebotomus Artemiev whose morphological differentiation is difficult. All three species have been suspected, but never proven, to be vectors of Leishmania infantum. The ND4 gene from mtDNA was sequenced from specimens of five populations of Phlebotomus mascittii Grassi from Belgium (the first records of Phlebotomine sandflies from this country), France, and Germany. Additionally, specimens from one population of Lebanese P. canaaniticus Adler and Theodor and of one population of Cypriot P. economidesi Leger, Depaquit and Ferte topotypes were examined. The results reveal the validity of the three taxa inferred from molecular variation and a molecular homogeneity shared by all of the populations of P. mascittii included into the present study. The epidemiological consequences are discussed. PMID- 15666189 TI - Present and future of deep brain stimulation for refractory epilepsy. PMID- 15666188 TI - Hydrocephalus due to superior sagittal sinus thrombosis. AB - The great variability of clinical appearance is one of the main features of superior sagittal sinus thrombosis. However, hydrocephalus associated with cerebral sinus thrombosis is rare. We report on a patient presented with thunderclap headache, accompanied by nausea, vomiting, and drowsiness. Lumbar puncture ruled out subarachnoid hemorrhage, whereas CT revealed marked hydrocephalus. In addition, magnetic resonance venography then confirmed the diagnosis of cerebral sinus thrombosis. It is a rare occurrence but clinically important, since it entails disastrous sequels if unrecognized, and hydrocephalus is treated in the usual fashion with ventricular drainage. PMID- 15666190 TI - Biweekly administration of gemcitabine and vinorelbine as first line therapy in elderly advanced breast cancer. AB - BACKGROUND: Gemcitabine and Vinorelbine both as a single agent or associated are active in advanced breast cancer patients as second line therapy, with low toxicity. In the elderly patients polichemotherapy is difficult for co-morbidity, but results with monotherapy are fewer. The use of this association as first line could be of help. PATIENTS AND METHODS: Thirty-four patients over 65 were treated with 1000 mg/m2 of gemcitabine and 25 mg/m2 of vinorelbine on days 1 and 8 every 21 days. An analysis of toxicities, TTP and OS were performed. RESULTS: The ORR was 53%: a CR was obtained in five patients (15%) and a PR in 13 patients (38%). Moreover, seven patients (21%) had a stable disease maintained for 6 months. The mean duration of CR and PR were, respectively, of 10 (range 7-19) and 7 (range: 4 14), months. Toxicity was low, mainly haematological: grade 3-4 neutropenia occurred only in 7 (20%) cases without febrile neutropenia. CONCLUSIONS: The gemcitabine and vinorelbine combination shows significant activity in elderly metastatic breast cancer patients. The treatment is well tolerated and has an acceptable toxicity profile. PMID- 15666192 TI - Polymorphisms in DNA repair gene XRCC1 and increased genetic susceptibility to breast cancer. AB - X-ray repair cross-complementing 1 (XRCC1) gene encodes for a scaffolding protein, which plays an important role in base excision DNA repair by bringing together DNA polymerase beta, DNA ligase III and poly(ADP-Ribose) polymerase (PARP) at the site of DNA damage. Three polymorphisms of the XRCC1 gene at codons 194, 280 and 399 leading to amino acid changes at evolutionary conserved regions are found to alter the efficiency of the resulting protein and may therefore constitute potential breast cancer risk. In the present study we sought to determine whether these genetic variants of the XRCC1 gene was associated with any increased risk of breast cancer among the South Indian women in a hospital based case control study using PCR-RFLP and DNA sequencing techniques. Our data showed a positive association between the polymorphisms of codons 194 (OR = 1.98, 95% CI = 1.13-3.48 for Trp allele) and 399 (OR = 2.14, 95% CI = 1.29-3.58 for Gln allele) and breast cancer risk. However, XRCC1 codon 280 genotype analysis showed no evidence for an association with increased risk of breast cancer. A combined analysis of the effect of XRCC1 codon 194 and 399 revealed the highest risk (OR = 3.64, 95% CI = 1.57-8.46) for carriers of the polymorphic alleles in both these codons. In conclusion, the present study suggested involvement of XRCC1 codon 194 and 399 polymorphisms in the genetic predisposition to breast cancer among South Indian women. Our preliminary results based on the analysis of functionally relevant polymorphisms in XRCC1 low penetrance gene may provide a better model that would exhibit additive effects on individual susceptibility to breast cancer. PMID- 15666191 TI - SHP-2 promoting migration and metastasis of MCF-7 with loss of E-cadherin, dephosphorylation of FAK and secretion of MMP-9 induced by IL-1beta in vivo and in vitro. AB - Shp-2, an src homology (SH) two-containing phosphotyrosine phosphatase, appears to be involved in cytoplasmic signaling downstream of a variety of cell surface receptors. It also plays an important role in the control of cell spreading, migration, and cytoskeletal architecture. In our study, abrogation of SHP-2 catalytic activity with a'dominant-negative mutant (SHP-2C > S) displayed an increased number of focal adhesion, high expression of E-cadhenrin and phosphorylation of the focal adhesion kinase (FAK). Interestingly, the cells expressing SHP-2C > S showed reduced IL-1beta-stimulated chemotaxis compared with either mock- or SHP-2 wild type-transfected cells. We also found that SHP-2-GFP transfected cell lines did not express E-cadherin nearly and produced high level of the matrix metalloproteinase MMP-9 in the supernatants. The loss of E-cadherin mediated adhesion and the increase of MMP-9-induced migration had been shown to play an important role in the transition of epithelial tumors from a benign to an invasive state. These findings have raised the possibility that SHP-2 can promote the cancer cell to invasion the distant tissues. To determine whether SHP-2 promotes invasion and metastasis, we transfected MCF-7 breast cancer cell lines with SHP-2-GFP, SHP-2C > S-GFP and analyzed the effects of the SHP-2 on cell migration, invasion, and metastasis. In vitro, SHP-2-GFP-transfected cells migrated more efficiently, showed an increased invasion of Matrigel, and adhered less efficiently to monolayers of fibroblast cells. When injected into the abdominal cavity of nude mice, SHP-2-GFP-transfected cells metastasized widely to the lung, kidney, but MCF-7 with SHP-2C > S-GFP was not observed in the these organs. These results demonstrate that SHP-2 promotes invasion and metastasis of MCF-7 with the loss of E-cadherin, the dephosphorylation of FAK and the secretion of MMP-9 induced by IL-1beta. PMID- 15666193 TI - Relationship of depression to patient satisfaction: findings from the barriers to breast cancer study. AB - OBJECTIVE: Examine the association between depressive symptoms and patient satisfaction for older women with a diagnosis of breast cancer. METHODS: Prospective study of 210 women aged 65 or older from southeast Texas newly diagnosed with breast cancer. Baseline (<2 months after diagnosis) and 12 month follow-up interviews were conducted face-to-face to collect information on sociodemographic characteristics, physical and emotional health, use of health services and satisfaction with medical care. Data analyses included descriptive statistics, chi2 analysis, and multivariate logistic regression analysis. RESULTS: Average age at baseline interview was 71.8 years (SD 6.6). The sample was 70.5% non-Hispanic white, 61.0% were unmarried, and 85.2% reported no ADL limitations. Logistic regression analysis showed a significant association between increasing depressive symptoms at baseline and lower patient satisfaction at follow-up. Each unit increase in depressive symptoms at baseline was associated with a 6% decrease in the predicted odds of being "very satisfied" with medical care at follow up (OR = 0.94, 95% CI = 0.89, 0.99), after adjusting for age, marital status, race/ethnicity, stage of diagnosis and other relevant factors. Similarly, patients who had an increase in CES-D score between baseline and follow-up interview were less likely to be satisfied with their medical care at follow up. CONCLUSION: Increasing depressive symptoms are associated with lower patient satisfaction. Early recognition and treatment of depressive symptoms may improve patients' well being and perception about quality of medical care. PMID- 15666194 TI - High preoperative serum TIMP-1 is a prognostic indicator for survival in breast carcinoma. AB - The amount of the immunoreactive protein for the tissue inhibitor of the matrix metalloproteinase-1 (TIMP-1) was studied prospectively from the pretreatment sera of 71 breast carcinoma patients using an enzyme-linked immunoassay (ELISA). The study consisted of patients with a primary breast carcinoma diagnosed between 1988 and 1991. The median follow-up time was more than 10 years, and routine adjuvant treatment was not used in the primary treatment. High TIMP-1 (> 196 ng/ml) was found to correlate with a poor relapse-free survival (RFS) in primary node-negative breast carcinoma. After 10 years of the follow-up only 42% of the patients with a high preoperative serum TIMP-1-level were free of the relapse, whereas 82% of the patients with a low serum TIMP-1 enjoyed a long RFS-time (log rank p =0.009). High serum TIMP-1 also indicated poor RFS (p = 0.02) and overall survival (p = 0.05) in stage I breast carcinoma. In Kaplan-Meier analysis the RFS was 89% in patients with a low serum level of TIMP-1 compared to 52% in patients with a high serum concentration of TIMP-1. In conclusion, preoperative high serum TIMP-1 levels predict poor outcome in primary breast carcinoma. In multivariate analysis preoperative high serum TIMP-1 increases the risk of relapse 3.4-fold during the first 10 years of follow-up in primary node-negative breast carcinoma. PMID- 15666195 TI - Prognostic factors affecting the natural history of node-negative breast cancer. AB - PURPOSE: We undertook a natural history investigation of a broad selection of prognostic factors in a cohort of women with node-negative breast cancer. PATIENTS AND METHODS: The cohort consisted of 415 consecutive histologic node negative (T1-3, M0) patients, operated on for primary breast cancer at Women's College Hospital, Toronto, Canada, between 1977 and 1986. Only 7% of these patients were given adjuvant systemic therapy; further, for the 48% of women who underwent lumpectomy, only 29% received adjuvant radiotherapy to the breast. Paraffin-embedded tumour tissue was available for the majority of patients. The following factors were examined for their univariate and multivariate effects on time to recurrence outside the breast (DFI) and survival from breast cancer (DSS): age, weight, tumour size, estrogen receptor, progesterone receptor, histologic type, tumour grade, nuclear grade, lymphovascular invasion, overexpression of neu oncoprotein, DNA ploidy, % cells in S-phase, and adjuvant therapy. Multivariate analyses utilized a Cox model with a step-wise factor selection for the 260 patients with complete information. RESULTS: A worse prognosis was indicated when there was lymphovascular invasion (for DFI, p < 0.001; for DSS, p = 0.0046), high %S-phase (for DFI, p = 0.08; for DSS, p = 0.02), high tumour grade (for DFI, p = 0.02; for DSS, p = 0.03), and overexpression of neu oncoprotein (for DSS, p = 0.07). CONCLUSIONS: In our natural history investigation, two factors, lymphovascular invasion and tumour grade, are of particular interest since they may be readily incorporated into clinical practice. Overexpression of neu oncoprotein may also play a role in determining prognosis for women administered adjuvant systemic therapy. PMID- 15666196 TI - The effect of age, race, tumor size, tumor grade, and disease stage on invasive ductal breast cancer survival in the U.S. SEER database. AB - PURPOSE: To examine the effect of patient and tumor characteristics on breast cancer survival as recorded in the U.S. National Cancer Institute's Surveillance, Epidemiology, and End Results (SEER) database from 1973 to 1998. METHODS: A sample of 72,367 female cases from 1973 to 1998 aged 21-90 years with invasive ductal breast cancer were examined with Cox proportional hazards regression to determine the effect of age at diagnosis, race, tumor size, tumor grade, disease stage, and year of diagnosis on disease-specific survival. RESULTS: Larger tumor size and higher tumor grade were found to have large negative effects on survival. Blacks had a 47 % greater risk of death than whites. Year of diagnosis had a positive effect, with a 15 % reduction in risk for each decade in the time period under study. The effects of patient age and disease stage violated the proportional hazards assumption, with distant disease having much poorer short term survival than one would expect from a proportional hazards model, and younger age groups matching or even falling below the survival rate of the oldest group over time. CONCLUSION: Tumor size, grade, race, and year of diagnosis all have significant constant effects on disease-specific survival in breast cancer, while the effects of age at diagnosis and disease stage have significant effects that vary over time. PMID- 15666197 TI - Florence-Sydney Breast Biopsy Study: sensitivity of ultrasound-guided versus freehand fine needle biopsy of palpable breast cancer. AB - BACKGROUND: Fine needle aspiration biopsy (FNAB) is widely used in the diagnosis of breast cancer. It is unknown whether, for palpable cancers, ultrasound-guided FNAB is more accurate than freehand FNAB, and practice varies between physicians, services and countries. METHODS: From consecutive women attending a major cancer centre in Florence, we prospectively recruited subjects who had a definitely palpable lump which was solid on ultrasound and suspicious of malignancy (n = 102). All subjects were investigated using both ultrasound-guided and freehand FNAB (one aspirate with each method). Radiologists skilled in both sampling techniques performed all clinical examinations and aspirations, and for each subject the same radiologist obtained both FNAB samples. Sequence of aspiration method was randomised. Cytological interpretation was blinded to method of sampling. Comparative sensitivity (and insufficiency) for FNAB using the two methods was calculated in all cancers (n = 97). RESULTS: Ultrasound-guided FNAB resulted in 13.6% (5-22%) less insufficient aspirates than freehand FNAB (chi2 = 7.58; p = 0.006). When insufficient aspirates are included and considered as negative, ultrasound-guided FNAB has a 14.6% (5.8-23%) or a 16.5% (7.6-25.4%) significantly better sensitivity than freehand FNAB (for cytology 3-5 positive or cytology 4-5 positive respectively). When insufficient aspirates are excluded from the analysis, ultrasound-guided FNAB has a 1.4% (-1.2 to 3.9%) or a 2.6% ( 2.5 to 7.8%) higher sensitivity than freehand FNAB (for cytology 3-5 positive or cytology 4-5 positive respectively) but this difference in the sensitivity of the two methods is not statistically significant. CONCLUSION: Our data suggest that ultrasound-guided FNAB has better sensitivity than freehand FNAB in palpable breast cancer, which is predominantly an effect of a significant reduction in insufficient aspirates, but in part an effect of 'upgrading' cytological classification of cancers. PMID- 15666198 TI - Reduced rates of cancer-related worries and mortality associated with guideline surveillance after breast cancer therapy. AB - OBJECTIVE: Guidelines have been developed for appropriate post-therapy surveillance for breast cancer recurrence. Two objectives of post-therapy surveillance are to support and counsel patients and to detect potentially curable local recurrences and new cancers in the opposite breast. The objective of this investigation was to assess the impact of guideline surveillance (history, physical examination, and annual mammography) on cancer-related worries and all-cause mortality. STUDY DESIGN AND SETTING: We collected data on a cohort of 303 Massachusetts women with stages I or II breast cancer diagnosed between 1992 and 1994. Cases were women with increasing cancer-related worries or decedents. We used risk-set sampling to match five controls to each case on follow-up time. Cases and members of their matched risk set were characterized with respect to receipt of guideline surveillance and covariates preceding the date of their outcomes. RESULTS: The adjusted odds ratio associating guideline surveillance in the preceding year with an increase in cancer-related worries equaled 0.37 (95% CI = 0.14-0.99). The adjusted odds ratio associating continuous guideline surveillance with all-cause mortality equaled 0.66 (95% CI = 0.51 0.86). CONCLUSION: The results are consistent with the stated objectives of surveillance follow-up of breast cancer patients after the completion of their primary therapy. PMID- 15666199 TI - The sentinel lymph node biopsy under local anesthesia in breast carcinoma: experience of the European Institute of Oncology and impact on quality of life. AB - BACKGROUND: Sentinel lymph node biopsy performed under local anesthesia currently represents one of the main fields of investigation in breast cancer surgery because of its positive impact on the management and organisation of treatment. We previously published our preliminary experience with 115 patients; here we present an update of our experience with particular attention devoted to quality of life. PATIENTS AND METHODS: From September 2000 to April 2003 we treated 244 patients with this technique. We developed a 'questionnaire of impact on quality of life - LA' to be administered to these patients, to assess their reactions to the procedure: we administered the questionnaire to 243 patients. Our aim was to verify whether the advantages in terms of treatment planning are accompanied by an improvement in quality of life and a good degree of acceptance by the patients. RESULTS: The questionnaire was administered to the patients during follow-up and we obtained positive results; the impact of the procedure under local anesthesia was positive for 81% of patients, while the remaining 19% reported that it had 'no impact at all'. We can conclude that axillary sentinel node biopsy, when performed under local anesthesia, meets with a very high degree of acceptance by the patients and exerts an optimal impact on quality of life. PMID- 15666200 TI - The impact of having relatives affected with breast cancer on psychological distress in women at increased risk for hereditary breast cancer. AB - PURPOSE: Being at hereditary risk of breast cancer (BC) may lead to elevated levels of distress because of the impact of the BC-process in relatives. OBJECTIVE: Determine the association between psychological distress and BC in relatives. We studied: kind of kinship with the affected relative(s), degree of involvement with the relative's BC, time elapsed since the BC diagnosis of the relative, and loss of a relative as a consequence of BC. METHODS: The study cohort consisted of women at increased risk of developing BC, adhering to regular surveillance and participating in the Dutch MRISC-study. Two months prior to the surveillance appointment, demographics, general and BC specific distress and experience with BC in the family were assessed. RESULTS: 347 out of 351 participants (mean age 40 1/2) had at least one relative affected with BC. The following variables were significantly, positively related to BC specific distress: having at least one affected sister (n = 105; p < 0.04); close involvement in a sister's BC process (n = 94; p < 0.03); and a recent (less than three years ago) BC diagnosis in a sister (n = 30; p < 0.03). General distress did not show any significant associations with the experience of BC in the family. CONCLUSION: These findings show the impact of a BC diagnosis in a sister, particularly a recent diagnosis, on psychological distress. Women who have experienced BC in their sister may be in need of additional counselling or of more attention during the surveillance process. PMID- 15666201 TI - Locally advanced breast cancer in octogenarian women. AB - Elderly patients are more likely to present with locally advanced breast cancer than younger patients. Furthermore, due to the accelerated aging of the western population, the incidence of breast cancer in this population is expected to steadily rise in the coming decades. So far, no guidelines are available for the management of octogenarian patients presenting with inoperable disease, what frequently results in a dilemma for the treating physician. For the time being, these patients should be ideally treated within the context of a clinical trial. In all other cases, the treatment has to be individualised, frequently based on data extrapolated from different population of patients, or retrospective series. This article reviews the current evidence, options, and most promising approaches for these patients. PMID- 15666202 TI - The 3020insC allele of NOD2 predisposes to early-onset breast cancer. AB - The NOD2 gene has been associated with susceptibility to Crohn's disease, and more recently with carcinoma of the colon as well. NOD2 is involved in the inflammatory response and the activation of the NFkB pathway. The range of cancer types associated with NOD2 has not been well studied. The 3020insC allele results in a truncated NOD2 protein and is present in approximately 7% of the population. We studied a possible association between the 3020insC allele of the NOD2 gene and breast cancer using 462 cases and 1910 controls from Poland. Patients were diagnosed with invasive breast cancer at are of two Szczecin regional hospitals between 2002 and 2004. Pathology specimens were reviewed for histological subtype and for the presence of ductal carcinoma in situ (DCIS). Overall there was no association between breast cancer and NOD2 (OR = 1.1; p = 0.76), but significant associations were observed between the presence of the allele and early-onset breast cancer (OR = 1.9; p = 0.01) and between the allele and ductal breast cancer with an in situ component (OR = 2.2; p = 0.006). PMID- 15666205 TI - Benign paroxysmal positional vertigo and post-treatment quality of life. AB - Benign paroxysmal positional vertigo is probably the most common cause of vertigo. It is characterized by acute short-lived episodes of severe vertigo in association with change in the position of the head. This condition is benign, and after the repositioning procedure, the cure rate is between 70 to 80%. Numerous studies describe the association between vertigo and social handicap and emotional disturbance. In our study, we report for the first time the level of anxiety and social consequences, tested with the Hamilton anxiety scale and vertigo handicap questionnaire, in a group of patients who had suffered only from positional vertigo and were clinically cured when the questionnaires and test were administered. The important role played by psychological factors in maintaining or increasing the social consequences and perception of vertigo is revealed in this study. PMID- 15666206 TI - The glycine decarboxylase complex is not essential for the cyanobacterium Synechocystis sp. strain PCC 6803. AB - In order to investigate the metabolic importance of glycine decarboxylase (GDC) in cyanobacteria, mutants were generated defective in the genes encoding GDC subunits and the serine hydroxymethyl-transferase (SHMT). It was possible to mutate the genes for GDC subunits P, T, or H protein in the cyanobacterial model strain Synechocystis sp. PCC 6803, indicating that GDC is not necessary for cell viability under standard conditions. In contrast, the SHMT coding gene was found to be essential. Almost no changes in growth, pigmentation, or photosynthesis were detected in the GDC subunit mutants, regardless of whether or not they were cultivated at ambient or high CO2 concentrations. The mutation of GDC led to an increased glycine/serine ratio in the mutant cells. Furthermore, supplementation of the medium with low glycine concentrations was toxic for the mutants but not for wild type cells. Conditions stimulating photorespiration in plants, such as low CO2 concentrations, did not induce but decrease the expression of the GDC and SHMT genes in Synechocystis. It appears that, in contrast to heterotrophic bacteria and plants, GDC is dispensable for Synechocystis and possibly other cyanobacteria. PMID- 15666207 TI - The Linderniaceae and Gratiolaceae are further lineages distinct from the Scrophulariaceae (Lamiales). AB - The Lamiales are one of the largest orders of angiosperms, with about 22,000 species. The Scrophulariaceae, as one of their most important families, has recently been shown to be polyphyletic. As a consequence, this family was re classified and several groups of former scrophulariaceous genera now belong to different families, such as the Calceolariaceae, Plantaginaceae, or Phrymaceae. In the present study, relationships of the genera Craterostigma, Lindernia and its allies, hitherto classified within the Scrophulariaceae, were analyzed. Sequences of the chloroplast trnK intron and the matK gene (approximately 2.5 kb) were generated for representatives of all major lineages of the Lamiales and the former Scrophulariaceae. Bayesian and parsimony analyses revealed two isolated lineages, one of which consists of Lindernia and its allies, the other of Gratiola and allies. Gratiola was previously assumed to be related to Lindernia and was therefore included here. It is proposed to treat the two clades as separate families, Linderniaceae and Gratiolaceae. For the Linderniaceae, several morphological synapomorphies exist in addition to molecular data, such as conspicuous club-shaped stamen appendages. PMID- 15666209 TI - Sucrose carrier RcSCR1 is involved in sucrose retrieval, but not in sucrose unloading in growing hypocotyls of Ricinus communis L. AB - The transport of assimilates from source to sink tissues is mediated by the phloem. Along the vascular system the phloem changes its physiological function from loading phloem to transport and unloading phloem. Sucrose carrier proteins have been identified in the transport phloem, but it is unclear whether the physiological role of these transporters is phloem unloading of sucrose or retrieval of apoplasmic sucrose back into the sieve element/companion cell complex. Here, we describe the dynamic expression of the Ricinus communis sucrose carrier RcSCR1 in the hypocotyl at different sink strengths. Our results indicate that phloem unloading in castor bean is not catalysed by the phloem loader RcSCR1. However, this sucrose carrier represents the molecular basis of the sucrose retrieval mechanism along the transport phloem, which is dynamically adjusted to the sink strength. As a consequence, we assume that other release carrier(s) exist in sink tissues, such as the hypocotyl, in R. communis. PMID- 15666208 TI - Transport of isoprenoid intermediates across chloroplast envelope membranes. AB - The common precursor for isoprenoid biosynthesis in plants, isopentenyl diphosphate (IPP), is synthesized by two pathways, the cytosolic mevalonate pathway and the plastidic 1-deoxy-D-xylulose 5-phosphate/methylerythritol phosphate (DOXP/MEP) pathway. The DOXP/MEP pathway leads to the formation of various phosphorylated intermediates, including DOXP, 4-hydroxy-3-methylbutenyl diphosphate (HMBPP), and finally IPP. There is ample evidence for metabolic cross talk between the two biosynthetic pathways. The present study addresses the question whether isoprenoid intermediates could be exchanged between both compartments by members of the plastidic phosphate translocator (PT) family that all mediate a counter-exchange between inorganic phosphate and various phosphorylated compounds. Transport experiments using intact chloroplasts, liposomes containing reconstituted envelope membrane proteins or recombinant PT proteins showed that HMBPP is not exchanged between the cytosol and the chloroplasts and that the transport of DOXP is preferentially mediated by the recently discovered plastidic transporter for pentose phosphates, the xylulose 5 phosphate translocator. Evidence is presented that transport of IPP does not proceed via the plastidic PTs although IPP transport is strictly dependent on various phosphorylated compounds on the opposite side of the membrane. These phosphorylated trans compounds are, in part, also used as counter-substrates by the plastidic PTs but appear to only trans activate IPP transport without being transported. PMID- 15666210 TI - Let's talk about...chloroplast import. PMID- 15666212 TI - Endogenous cytokinin oscillations control cell cycle progression of tobacco BY-2 cells. AB - The significance of cytokinins for the progression of the cell cycle is well known. Cytokinins contribute to the control of the expression of D-cyclins and other cell cycle genes, but knowledge as to how they affect the progression of the cell cycle is still limited. Highly synchronized tobacco BY-2 cells with clearly defined cell cycle stages were employed to determine cytokinin patterns in detail throughout the entire cycle. Concentrations of trans-zeatin, and of some other cytokinins, oscillated during the course of the cell cycle, increasing substantially at all four phase transitions and decreasing again to a minimum value during the course of each subsequent phase. Addition of exogenous cytokinins or inhibition of cytokinin biosynthesis promoted the progression of the cell cycle when the effects of these manipulations intensified the endogenous fluctuations, whereas the progression of the cycle was retarded when the amplitude of the fluctuations was decreased. The results show that the attainment of low concentrations of cytokinins is as important as the transient increases in concentration for a controlled progression from one phase of the cell cycle to the next. Cytokinin oxidase/dehydrogenase activity also showed fluctuations during the course of the cell cycle, the timing of which could at least partly explain oscillations of cytokinin levels. The activities of the enzyme were sufficient to account for the rates of cytokinin disappearance observed subsequent to a phase transition. PMID- 15666213 TI - Specificity patterns indicate that auxin exporters and receptors are the same proteins. AB - A study of transport and action of synthetic auxin analogues can help to identify transporters and receptors of this plant hormone. Both aspects--transportability and action on growth--were tested with 2-naphthoxyacetic acid (2-NOA) and compared across several plant species. 2-NOA stimulates elongation effectively at low concentrations in petioles of the gymnosperm Ginkgo biloba L., in hypocotyls or internodes of the dicot legumes, mung bean (Vigna mungo L.) and pea (Pisum sativum L.), in cotyledons of onion (Allium cepa L.) and in leaf bases of chive (Allium schoenoprasum L.), the latter two of the monocot order Asparagales. In contrast, elongation of coleoptile segments of maize (Zea mays L.) is poorly responsive to 2-NOA. Significant auxin-like transport of 2-NOA was observed in segments of mung bean hypocotyls, pea internodes, and chive leaf bases, but not in segments of the grass coleoptiles. Thus, for the two assays, elongation and polar transportability, the same difference in ligand specificity was observed between the grass and all other species assayed. This finding supports the hypothesis that a common protein mediates auxin efflux as well as auxin action on elongation. PMID- 15666214 TI - Changes in the subcellular distribution of glutathione during virus infection in Cucurbita pepo (L.). AB - Changes in the subcellular distribution and quantification of glutathione were studied with electron microscopic immunogold cytochemistry in Zucchini yellow mosaic virus (ZYMV)-infected Styrian pumpkin plants (Cucurbita pepo L. ssp. pepo var. styriaca Greb.) two weeks after inoculation. The amount of gold particles bound to glutathione was statistically evaluated for different cell structures, including mitochondria, plastids, nuclei, peroxisomes, and cytosol. In general, ZYMV-infected plants showed higher gold labelling density in intact mesophyll cells of the 5th (older leaves) and the youngest fully developed leaves (younger leaves), and decreased levels of glutathione within root tip cells when compared to the control. In general, within older and younger leaves the highest amount of gold particles was found in mitochondria and the lowest amount in plastids. In ZYMV-infected older leaves, an increase in glutathione was found in peroxisomes (1.7-fold), the cytosol (1.6-fold), mitochondria (1.4-fold), and nuclei (1.2 fold), whereas glutathione levels in plastids did not differ significantly when compared to control cells. In ZYMV-infected younger leaves elevated glutathione contents were found in the cytosol (3-fold), nuclei (2.1-fold), peroxisomes (1.8 fold), and plastids (1.5-fold), whereas mitochondria showed an insignificant decrease in glutathione levels in comparison to the control. In root tip cells of ZYMV-infected plants the amount of gold particles bound to glutathione was decreased in all investigated cell structures by between 0.7- to 0.8-fold. Additionally, total glutathione contents were determined in older and younger leaves using high-performance liquid chromatography (HPLC), which revealed no significant differences between control and ZYMV-infected leaves. The relevance of the results of both methods were compared and are discussed. PMID- 15666215 TI - The quantum yield of CO2 fixation is reduced for several minutes after prior exposure to darkness. Exploration of the underlying causes. AB - Previous work has shown that the apparent quantum yield of CO2 fixation can be reduced for up to several minutes after prior exposure to darkness. In the work reported here, we investigated this phenomenon more fully and have deduced information about the underlying processes. This was done mainly by concurrent measurements of O2 and CO2 exchange in an oxygen-free atmosphere. Measurements of O2 evolution indicated that photochemical efficiency was not lost through dark adaptation, and that O2 evolution could proceed immediately at high rates provided that there were reducible pools of photosynthetic intermediates. Part of the delay in reaching the full quantum yield of CO2 fixation could be attributed to the need to build up pools of photosynthetic intermediates to high enough levels to support steady rates of CO2 fixation. There was no evidence that Rubisco inactivation contributed towards delayed CO2 uptake (under measurement conditions of low light). However, we obtained evidence that an enzyme in the reaction path between triose phosphates and RuBP must become completely inactivated in the dark. As a consequence, in dark-adapted leaves, a large amount of triose phosphates were exported from the chloroplast over the first minute of light rather than being converted to RuBP for CO2 fixation. That pattern was not observed if the pre-incubation light level was increased to just 3-5 micromol quanta m(-2) s(-1). The findings from this work underscore that there are fundamental differences in enzyme activation between complete darkness and even a very low light level of only 3-5 micromol quanta m(-2) s(-1) which predispose leaves to different gas exchange patterns once leaves are transferred to higher light levels. PMID- 15666216 TI - Molecular relationships in Encephalartos (Zamiaceae, Cycadales) based on nucleotide sequences of nuclear ITS 1&2, rbcL, and genomic ISSR fingerprinting. AB - The cycad genus Encephalartos is restricted to Africa and is threatened with extinction in most of its range. Total DNA was extracted from 51, i.e., 78 %, of the described species of Encephalartos. The accessions were sampled from the furthest western occurrence of the genus in Nigeria, via Sudan and Uganda, to southern South Africa. The sequences of nuclear ribosomal internal transcribed spacer regions 1 and 2 (ITS 1&2), the chloroplast encoded rbcL gene, and ISSR genomic fingerprinting were employed to resolve the molecular history and the relationships within the genus. Sequence alignment, as well as ISSR fingerprinting, data show low genetic variation among all analysed accessions, indicating diversification within the Pliocene/Pleistocene. ITS 1&2 data agree well with morphological and geographical characters and resolved three major genetic clusters with overlapping distribution ranges in eastern South Africa. This area, that contains the largest diversity of genotypes of Encephalartos, may have served as a Pliocene/Pleistocene refugium. PMID- 15666217 TI - Inheritance of a recessive transgene-associated character controlling albinism in transgenic bean (Phaseolus vulgaris L.). AB - We identified a transgenic line exhibiting albinism during our work to introduce genes through genetic engineering in dry bean (Phaseolus vulgaris). The transgenic mother plant (R0) presented a normal phenotype and generated albino and normal green plants in the first generation (R1). The segregation ratio of the albino character in the R1 and R2 generations fitted the expected ratio for a character controlled by a single recessive gene linked to a foreign gus gene, suggesting that albinism could be a consequence of insertional mutation caused by introduction of the exogenous gene. Analysis by electron microscope revealed that the albino cells possessed no chloroplasts and a greater number of mitochondria when compared to normal green plants. This transgenic bean line may be used in understanding the genetic control of chloroplast genesis, for acquiring additional knowledge of genomic structure or in physiological studies. This is the first described transgene-associated mutant bean plant. PMID- 15666218 TI - [Did we make progress in the treatment of anorexia nervosa?]. PMID- 15666211 TI - ESTs from the fibre-bearing stem tissues of flax (Linum usitatissimum L.): expression analyses of sequences related to cell wall development. AB - In order to learn more about the diversity of genes expressed during flax fibre cell wall formation, expressed sequence tags (ESTs) were obtained from a cDNA library derived from the outer fibre-bearing tissues of flax (Linum usitatissimum) stems (cv Hermes) harvested at the mid-flowering stage. After elimination of vector and unreadable sequences, 927 ESTs were grouped into 67 clusters and 754 singletons. The flax ESTs have been submitted to the dbEST and GenBank databases with the accession numbers 25939634 - 25940560 (dbEST) and CV478070 - CV478996 (GenBank). Functional analysis allowed the grouping of ESTs into 13 functional categories and revealed that 62 % of ESTs were similar to known sequences, while 12.4 % of ESTs presented no similarity to any known sequences and 25.6 % of ESTs corresponded to proteins of unknown function. The most highly expressed transcripts belonged to four functional categories: protein maturation and metabolism (31 ESTs), signalling (22 ESTs), the cell wall (21 ESTs) and photosynthesis (19 ESTs). 4.4 % (41) of the total ESTs were potentially related to cell wall formation and maturation. The most highly expressed cell wall EST (15 ESTs) corresponded to a beta-xylosidase gene--potentially involved in cell wall remodelling during growth and development. Other cell wall-related ESTs corresponded to cellulose synthase, xyloglucan endotranglucosylase/hydrolase (XTH), beta-galactosidases, and peroxidases. The expression patterns of different cell wall-related ESTs were determined at different developmental stages in flax plants grown under different field conditions. The potential roles of gene products associated with cell wall related ESTs in fibre cell wall development is discussed. PMID- 15666219 TI - [Involuntary hospitalization of patients with anorexia nervosa: clinical issues and empirical findings]. AB - Avoidance and refusal of treatment by patients with anorexia nervosa are eminent features of the disorder. A subgroup of severely ill patients are affected by involuntary hospitalizations. Based on empirical studies, this review analyzes whether patients with involuntary hospitalization differ from patients with voluntary treatment with regard to clinical characteristics and treatment outcomes. Furthermore, reasons for treatment refusal and diminished treatment motivation are outlined based on a selective review of the literature. Patients who where admitted involuntarily had significantly more previous hospitalizations and a longer duration of illness. Other predisposing factors for an involuntary admission were a history of childhood abuse and self-mutilative behavior, whereas a history of comorbid substance abuse or clinical depression was not predictive. Empirical findings suggest that weight restoration was similarly effective for both patient groups. Because catamnestic studies of patient groups with involuntary admissions are insufficient, it is presently difficult to assess the treatment response and the risk of mortality in the long-term. Future research is mandatory to identify risk factors in this special group of patients in order to establish specific interventions. PMID- 15666220 TI - [Preliminary recidivism rates of the Essener prognosis study]. AB - The prediction of dangerousness in mentally disordered offenders -- and here mainly the issue of safety -- is still of primary interest for forensic experts and the public. The effectiveness of treatment and therefore of the whole German forensic system is generally measured by recidivism rates of this clientele. This article presents topical recidivism rates of 255 mentally ill offenders after a minimum time at risk of 2 years (mean 4 years). Data are collected in the Essener prospective multicenter study funded by the German research association which began in 1997 and in which 23 forensic hospitals in Germany took part. Main objective of the study is the determination of valid criteria for predicting recidivism of mentally disordered offenders. Currently the rates for general recidivism are 21.6 % and for severe reoffences 7.5 % which are in comparison to other recent studies relatively low. PMID- 15666221 TI - [Horner's syndrome -- update on neuroanatomy, topographic diagnosis and etiology]. AB - Due to the complex neuroanatomy of the sympatho-excitatory pathway, Horner's syndrome represents a clinical sign that may result from a variety of lesions in the central and peripheral nervous system. The purpose of the present communication is to help the reader to localize the site of the lesion and to demonstrate the most common etiologic mechanisms resulting in Horner's syndrome. The functional anatomy of the sympathetic supply to the iris, eyelids, facial sweat glands and blood vessels is reviewed and in particular the structure of the central pathway updated. Moreover, pharmacological testing and tests of sudomotor function are described that may help to guide the decision regarding useful additional diagnostic, especially neuroimaging procedures. Finally, a schematic overview is given on the most common pathology, considering additional clinical signs and symptoms. PMID- 15666222 TI - [The suffering of Heinrich Heine]. AB - The analysis of clinical neurological symptoms on the basis of available letters and other documents confirms beyond a doubt the long standing diagnosis of neurosyphilis in the form of chronic meningitis with cranial polyneuritis and wide spread polyradiculitis, formerly called Lues cerebro-spinalis. The symptoms which recurred over 25 years include multiple severe paralysis of cranial nerves, dramatic radicular neuralgias and radicular paralysis in addition to a partial transverse lesion of the spinal cord and terminal coughing and regurgation fits in the absence of symptoms in the cerebrum and brain stem, these symptoms hardly permit different diagnosis. Tuberculosis, lead poisoning, multiple sclerosis or even a myatrophic lateral sclerosis do not explain his neurological symptoms or the cause of his death. PMID- 15666223 TI - [Migraine]. PMID- 15666224 TI - Inversion prepared coronary MR angiography: direct visualization of coronary blood flow. AB - PURPOSE: Visualization of coronary blood flow by means of a slice-selective inversion pre-pulse in concert with bright-blood coronary MRA. MATERIALS AND METHODS: Coronary magnetic resonance angiography (MRA) of the right coronary artery (RCA) was performed in eight healthy adult subjects on a 1.5 Tesla MR system (Gyroscan ACS-NT, Philips Medical Systems, Best, NL) using a free breathing navigator-gated and cardiac-triggered 3D steady-state free-precession (SSFP) sequence with radial k-space sampling. Imaging was performed with and without a slice-selective inversion pre-pulse, which was positioned along the main axis of the coronary artery but perpendicular to the imaging volume. Objective image quality parameters such as SNR, CNR, maximal visible vessel length, and vessel border definition were analyzed. RESULTS: In contrast to conventional bright-blood 3D coronary MRA, the selective inversion pre-pulse provided a direct measure of coronary blood flow. In addition, CNR between the RCA and right ventricular blood pool was increased and the vessels had a tendency towards better delineation. Blood SNR and CNR between right coronary blood and epicardial fat were comparable in both sequences. CONCLUSION: The combination of a free-breathing navigator-gated and cardiac-triggered 3D SSFP sequence with a slice-selective inversion pre-pulse allows for direct and directional visualization of coronary blood flow with the additional benefit of improved contrast between coronary and right ventricular blood pool. PMID- 15666225 TI - [Endovenous treatment of primary varicose veins: an effective and safe therapeutic alternative to stripping?]. AB - Endovenous laser therapy (EVLT) is a new, minimally invasive therapeutic option for treating primary varicose veins and provides an effective and safe alternative to conventional surgical management (stripping). Short-term and intermediate-term outcome is comparable to surgical stripping in terms of elimination of venous reflux (90 % - 98 %), resolution of visible varices (85 %), and improvement of subjective complaints such as sensations of heaviness and tension (96 %). Complications occur in 1 % - 3 % of cases, which is markedly below the rate of conventional surgical management (up to 30 %). The intermediate term incidence of recurrent varicosis in a vein treated by EVLT depends on the laser fluence applied and is reported to range from 7 % - 9 % compared to 10 % - 20 % after surgical intervention. Based on a review of the current literature and our own experience, this survey article presents an overview of the indications and contraindications, the technique and pathophysiology of laser-induced venous occlusion, and the results and possible complications of EVLT. PMID- 15666226 TI - [Computer-aided diagnosis and volumetry of pulmonary nodules: current concepts and future perspectives]. AB - For computer-aided detection (CAD) and volumetry of small pulmonary nodules, a number of algorithms have been developed for multislice CT data sets in recent years, with the goal of improving the diagnostic work-up and the follow-up of findings. Recent data show that the detection of small lesions may improve with CAD, suggesting that especially experienced readers may benefit from using CAD systems. This has lead to the recommendation of CAD as a replacement of the second reader in clinical practice. Furthermore, computer-aided volumetry of pulmonary nodules allows a precise determination of nodular growth rates as a prerequisite for a better classification of nodules as benign or malignant. In this article, we review recent developments of CAD and volumetry tools for pulmonary nodules, and address open questions regarding the use of these software tools in clinical routine. PMID- 15666227 TI - [Influence of body constitution on bone mineral density measured by digital radiogrammetry]. AB - PURPOSE: In addition to many established osteodensitometric techniques, digital radiogrammetry (DXR) is considered to be a reliable method for measuring the cortical bone mineral density (DXR-BMD). This study investigates the influence of body constitution on BMD of healthy adults as calculated by DXR. MATERIALS AND METHODS: In a prospective study, 246 adults without bone affecting diseases in their clinical history underwent DXR for analysis and calculations of bone mineral density and determination of metacarpal index (MCI) and porosity index (PI). Height, weight and body mass index (BMI) were recorded for each patient. RESULTS: For all individuals and for all BMI subgroups, both height (0.55 < R < 0.70, p < 0.01) and body weight (0.56 < R < 0.78, p < 0.01) correlated closely with DXR-BMD. Only in the over-weight group, no significant correlation was found between body weight and DXR-BMD. In addition, a significant reduction of the relative DXR-BMD and MCI values was observed between the over-weight and the under-weight group as well as between normal-weight and under-weight individuals (p < 0.01). Otherwise, cortical porosity decreased with increasing body weight. CONCLUSION: Similar to Dual Energy X-ray Absorptiometry-based studies (DXA), digital radiogrammetry measures an increase in BMD with increasing body weight. Therefore DXR, which provides a precise technique without influence of soft tissue, seems to be a promising technique for quantifying marginal alterations in cortical BMD as well for following the course of osteoporosis. PMID- 15666228 TI - [Computed tomographic examination of muscle volume, cross-section and density in patients with dysgnathia]. AB - PURPOSE: The individual jaw position is determined by the masticatory muscle among other factors. Before surgical treatment of malocclusions, thorough evaluation of the muscles is required to estimate the relapse risk. MATERIALS AND METHODS: By means of computer tomography, lateral radiographs of the skull and denture models, the relationships between morphological parameters of the masticatory muscles and the jaw bone were analyzed. Furthermore, possible causes for the extent of the malocclusion are described. RESULTS: A patient group with deep overbite was found to have significantly higher muscle densities (measured in Hounsfield units [HU]) in the medial pterygoideus muscle (59.89 +/- 3.91 HU to 48.94 +/- 4.14 HU, p < 0.01), masseter muscle, and genioglossus muscle (p < 0.05) in comparison to open bite patients. Significant differences of the muscle cross section were measured in the masseter muscle between patients with retroclined maxillary incisors and with an open bite (5.4 +/- 0.7 cm (2) to 3.8 +/- 0.4 cm (2), p < 0.05). CONCLUSION: The results show a correlation between different jaw positions and masticatory muscles. They also suggest that the function of each muscle may be different. Additional examinations of the muscle structures are required for verification of the influence of the masticatory muscles on facial morphology. PMID- 15666229 TI - [The role of projectional radiography in the detection of primary malignant and indeterminate bone neoplasms]. AB - PURPOSE: To evaluate the role of projectional radiography in the detection of primary and potentially malignant bone neoplasms. MATERIALS AND METHODS: We retrospectively reviewed the images and medical records of 80 patients (42 male, 38 female; range: 6 to 73 years; mean: 33 years) with pathologically proven primary or indeterminate bone tumors. RESULTS: The most common symptom was local pain (84 %). In 69 of 80 patients (86 %), projectional radiography was the first imaging modality, which correctly identified the malignant character of the bone lesion in 60 of these 69 patients (87 %). In 2 patients, bone neoplasms were overlooked due to diagnostically inadequte image quality. The false-diagnosis rate was not significantly different between board-certified radiologists working in hospitals and radiologists in outpatient facilities (p > 0.05). Bone tumors predominantly affecting adults, such as chondrosarcomas, needed a longer time for the diagnosis than bone tumors mainly found in childhood, such as osteosarcomas. The median age of patients with delayed imaging work-up because of attempted conservative therapy (45 years +/- 20 years) was significant higher than the median age of patients with immediate radiographic work-up (27 years +/- 16 years). CONCLUSION: Projectional radiography is the most common initial imaging method for bone pain and continues to play an important role to reduce the time gap between first onset of symptoms and definitive treatment. In unremitting focal bone pain, bone neoplasms should be taken into consideration not only in children but also in patients beyond adolescence. PMID- 15666230 TI - [Morphometric analysis of the temporomandibular joint with MRI in 320 joints]. AB - PURPOSE: To compare several morphometric parameters in MRI with the functional status of the articular disc in a large patient group suffering from internal derangement (ID) of the temporomandibular joint (TMJ). MATERIALS AND METHODS: In a retrospective study, 320 analyzable high resolution MRI examinations of the TMJs obtained in a 1.5 T unit were evaluated in 184 patients with clinically suspected ID. The analysis included the anatomical structures and a number of morphometric parameters previously described in the literature. The parameters were compared with the position of the articular disc. RESULTS: The disc position was categorized as "normal" (NDP, 21.9 %, n = 70), "anterior displacement with reduction on opening" (AMR, 51.6 %, n = 165) and "anterior displacement without reduction on opening" (AOR, 26.6 %, n = 85). With increasing disc displacement, significant configurational changes of the disc were observed. Disc displacement was associated with changes of the condyle consisting of increasing deformity and other degenerative changes. A large tuberculum and marked inclination of the eminence can be seen as predisposition for the development of ID. With increasing severity of the ID, the position of the condyle moved from a centric position of the condylar center to an excentric (dorsal and cranial) position. CONCLUSION: MRI demonstrated that increasing disc displacement is associated with changes of the disc, condyles and condylar position in the fossa. PMID- 15666231 TI - [Sonography of the optical nerve -- experience in 483 children]. AB - INTRODUCTION: In case of increased intracranial pressure (IICP), the inflow of cerebrospinal fluid widens the space between the optic nerve (ON) itself and the surrounding dura mater leading to the sonographic appearance of increased diameter of the ON. The purpose of the study was to gain clinical experience in children and to determine (a) the mean values for patients without proven IICP and (b) pathologic values of those with proven IICP. MATERIALS AND METHODS: Ultrasound of the optic nerve was performed in 483 children with symptoms consistent with IICP, comprising 287 males and 196 females at an age ranging from 4 days to 24 years with a mean age of 7.5 + 5.1 years. The measured diameter of the ON of both eyes and the morphologic criteria concerning nerve sheath and papilla were evaluated retrospectively. RESULTS: Most of the investigated patients (n = 466) had no IICP as confirmed by neurological examination, EEG, sometimes CCT and/or MRI and clinical follow-up. The typical morphological findings in patients with normal intracranial pressure (ICP) were a clear and longitudinally extended demarcation of the ON with a well-delimited nerve sheath and without prominent papilla. The mean diameter of the ON in patients with normal ICP was 3.4 mm + 0.7 mm. In 17 patients with proven IICP, the mean diameter of the ON increased to 5.6 mm + 0.9 mm. Typically, the nerve sheath was indistinguishable from the hypoechoic ON, often but not invariably associated with a prominent papilla. In patients with IICP, the diameter of the ON alone matches the diameter of the ON plus nerve sheath in patients with normal ICP. CONCLUSION: IICP was detected with a high sensitivity by ultrasound using the diameter and the morphological criteria of an indistinguishable nerve sheath. A diameter of more than 4.5 mm is definitely pathologic and requires further investigation. Prominence of the papilla is an unreliable criterion for acute IICP. PMID- 15666232 TI - [Duplex ultrasound of external carotid artery branches for the detection of dural arteriovenous fistulae]. AB - PURPOSE: To evaluate duplex ultrasonography of external carotid artery branches for the non-invasive diagnosis of dural AV fistulae (DAVFs). MATERIALS AND METHODS: A total of 126 patients suffering from pulsatile tinnitus underwent duplex ultrasonography of the cervicocephalic vasculature including external carotid artery branches. DAVFs were identified by increased flow velocity and reduced pulsatility. Patients with ultrasound findings of DAVFs were considered for selective angiography. RESULTS: Of the 23 patients that fulfilled the ultrasound criteria of AV fistula, 5 did not undergo angiography and were excluded from the study. The ultrasound findings of AV fistula were confirmed in all remaining patients (17 DAVFs, 1 extracranial AV fistula). Duplex ultrasound detected a DAVF of the common carotid artery in 29 %, of the external carotid artery in 65 % and of the external carotid artery branches in 100 % of the cases. CONCLUSION: Duplex ultrasonography of external carotid artery branches improves the non-invasive detection of DAVFs. For the evaluation of the venous drainage, however, selective angiography is necessary. PMID- 15666233 TI - [Prospective randomized trial of a modified standard multislice CT protocol for the evaluation of multiple trauma patients]. AB - PURPOSE: To evaluate whether modification of a standard multislice CT (MSCT) protocol might improve the diagnostic work flow in patients with multiple trauma without relevant loss of image quality. MATERIALS AND METHODS: Between September 2002 and September 2003, 80 multiple trauma patients underwent 4-slice CT encompassing head, thorax, abdomen/pelvis and spine. All patients were randomly assigned to either protocol A or B: Protocol A included serial scanning of the head (collimation 1 mm, 350/380 mAs/120 kV) and spiral scans of thorax, abdomen/pelvis and spine (collimation 2.5 mm, 220 mAs/120 kV) with gantry angulation and arm elevation; protocol B included spiral scanning of all body regions (collimation 2.5 mm, 300/150 mAs/120 kV) without gantry angulation or arm elevation. Time intervals, radiation exposure and results of the initial and final analysis were documented. RESULTS: In the investigated 64 male and 16 female patients (mean age 41.7 years), 88.7 % of the 407 pathologic findings were correctly identified on the initial images. Protocol B revealed a significant decrease in scan time (6.4 vs. 16.8 min., p < 0.001), time in the CT examination room (22.9 vs. 32.8 min.; p < 0.001), time until initial (25.3 vs. 35.8 min.; p < 0.001) and final image analysis (93.7 vs. 112.9 min; p < 0.005). No significant difference was found for patient transport time and image reconstruction time. Protocol B has a significantly lower effective radiation dose compared to protocol A (10.2 vs. 12.7 mSv, p < 0.001). CONCLUSIONS: Applying a modified MSCT protocol without gantry angulation and arm elevation can significantly decrease radiation exposure and examination time in multiple trauma patients without relevant loss of diagnostic image information and, consequently, has the potential of improving the diagnostic process and prognosis in multiple trauma patients. PMID- 15666234 TI - [Recommendations for the selection of storage media for archiving digital radiological image data based on the comparison of retrieval times at different PACS archive levels]. AB - PURPOSE: Recommendations for archiving digital radiological image data based on the comparison of retrieval times for different PACS archive levels. MATERIALS AND METHODS: For a large PACS installation (Agfa Impax, Release 4.1), image retrieval times for radiological standard examinations (chest radiographs with 2, MRI with 250, CT with 100 and 1000 images; n = 120, each) from hard disk array, magneto-optical disk (MOD), and magnetic tape archives (TAPE) were examined in high and low network traffic load. RESULTS: Even large CT examinations (1000 images) were available from hard disk arrays within 4.0 +/- 0.8 s, smaller studies within 1.8 +/- 0.3 s. Radiographic image retrieval from MOD (30 +/- 4.7 s) was more then 50 % faster than from TAPE. For typical cross-sectional studies, the velocity gain amounted to 19 %. For both technologies, no significant difference was found for large CT examinations (651 +/- 144 s). For high and low network traffic load scenarios, image retrieval times from hard disk, MOD, and TAPE archives increased by 87 %, 7 %, and 22 %, respectively. CONCLUSION: Hard disk arrays are specifically suited as departmental intermediate storage media because they allow fast access to current and previous examinations within a short time. Performance properties enable both MOD and TAPE systems to serve as long-term archives. However, MOD archives are less flexible in the expansion of storage capacity and at present the medium costs per memory unit are about 2 - 3 times higher than for tape archives. The use of existing MOD-archives may be adequate as intermediate archives. For new PACS installations or system expansions, however, it is recommended to combine a sufficiently large local data memory (RAID) with data storage on tape archives outside the radiological unit that can be used by other departments as well. Future development of hard disk prices will show whether archiving for the whole data retention period may be handled by RAID systems. In any case, prefetching problems and waiting periods for demanded pre-studies would not occur any more. PMID- 15666235 TI - [Comparison of radiation dose and image quality of Siremobil-IsoC(3D) with a 16 slice spiral CT for diagnosis and intervention in the human pelvic bone]. AB - PURPOSE: To compare the image quality of 16-slice computed tomography with the image quality of Siremobil-IsoC (3D) of the pelvic region and to measure simultaneously the radiation dose before and after implantation of a sacroiliac screw (SI-screw) MATERIALS AND METHODS: The pelvic region of 8 human cadavers was examined in the Siremobil-IsoC (3D) at five different levels. We used a standard protocol for the 16-slice CT of the complete pelvic region before and after insertion of a pelvic screw, followed by stepwise reduction of the tube current to find the tube current that equalizes the image quality of both modalities. We controlled the image quality by judging important structures such as neuroforamen, nerves, sacroiliacal joint space, intervertebral space, osteophytes, iliopsoas muscle, acetabular surface, fovea centralis, hip joint and os pubis. The image quality was judged by three radiologists and three trauma surgeons using a ranking from 1 to 5. The dose was measured with an endorectally placed NOMEX Dosimeter, to obtain the gonadal dose. RESULTS: The medium score for all viewers of the Siremobil-IsoC (3D) examinations was between 3 and 4.3. The medium score for all CT-examinations with a tube current of 250 mA was between 1.3 and 2.2. The reduction of tube current down to 80 mA hardly influenced the marks for the analyzed structures. Under 80 mA, bony structures, even after implantation of a SI-screw, were still marked as good, but soft tissue differentiation was getting worse. For the examination of the pelvis, the average dose-length product for the IsoC (3D) was 41.2 mGy x cm. The medium dose-length product for CT was 389 mGy x cm for 250 mA, 125 mGy x cm for 80 mA and 82 mGy x cm for 60 mA. CONCLUSION: The Siremobil-IsoC (3D) is sufficient for therapeutic intraoperative purpose, but the image quality is not sufficient for diagnostic purpose. The higher dose-length product of a CT examination is justifiable because of a better overview, shorter examination time and qualitative superiority. An advantage of the Siremobil-IsoC (3D) is the intraoperative availability with acceptable 3D image quality compared to conventional fluoroscopy. PMID- 15666236 TI - [Dose optimization for multislice computed tomography protocols of the midface]. AB - PURPOSE: To optimize multislice computed tomography (MSCT) protocols of the midface for dose reduction and adequate image quality. MATERIALS AND METHODS: MSCT (Somatom Volume Zoom, Siemens) of the midface was performed on 3 cadavers within 24 hours of death with successive reduction of the tube current, applying 150, 100, 70 and 30 mAs at 120 kV as well as 40 and 21 mAs at 80 kV. At 120 kV, a pitch of 0.875 and collimation of 4x1 mm were used, and at 80 kV, a pitch of 0.7 and collimation of 2x0.5 mm. Images were reconstructed in transverse and coronal orientation. Qualitative image analysis was separately performed by two radiologists using a five-point scale (1 = excellent; 5 = poor) applying the following parameters: image quality, demarcation and sharpness of lamellar bone, overall image quality, and image noise (1 = minor; 5 = strong). The effective body dose [mSv] and organ dose [mSv] of the ocular lens (using the dosimetry system "WINdose") were calculated, and the interobserver agreement (kappa coefficient) was determined. RESULTS: For the evaluation of the lamellar bone, adequate sharpness, demarcation and image quality was demonstrated at 120 kV/30 mAs, and for the overall image quality and noise, 120 kV/40 mAs was acceptable. With regard to image quality, the effective body dose could be reduced from 1.89 mSv to 0.34 mSv and the organ dose of the ocular lens from 27.2 mSv to 4.8 mSv. Interobserver agreement was moderate (kappa = 0.39). CONCLUSION: Adequate image quality was achieved for MSCT protocols of the midface with 30 mAs at 120 kV, resulting in a dose reduction of 70 % in comparison to standard protocols. PMID- 15666237 TI - [Comparison of four digital and one conventional radiographic image systems for the chest in a patient study with subsequent system optimization]. AB - PURPOSE: Using a patient study to prove the clinical relevance of a comparison of five different radiographic systems for the chest conducted with an anthropomorphic chest phantom. Depending on the results, it was tested whether the performance of a modern digital system with a transparent imaging plate can be improved by changing the post-processing of the image. METHOD: Chest radiographs of patients were taken with a CsI/aSi-flat panel detector (FDR), transparent imaging plate (tDLR), selenium drum detector (DSR), conventional storage phosphor plate (DLR) and asymmetrical screen-film-system (aFFS), and compared using image criteria scoring (ICS) and visual grading analysis (VGA) for anatomical structures (modified criterions of the EUR 16 260 EN guidelines). After optimizing the post processing, the images of the tDLR-system were evaluated once more in a phantom ROC study and patient VGA study. RESULTS: The flat panel detector-system proved to meet best the anatomical image quality criteria, followed by DSR, tDLR, aFFS and DLR. The modified post processing of the tDLR-images resulted in a significantly better detection of simulated pathological lung-structures, but improved the perceptibility of anatomical structures only slightly. CONCLUSIONS: The results of the patient VGA study and the phantom ROC study are similar and considered valid. The new digital imaging systems with flat panel detector and transparent imaging plate provide the best image quality of the tested radiographic devices for chest imaging, assuming that all system components are attuned and optimized for the type of structure to be detected. Image processing is of primary importance for system optimization. PMID- 15666238 TI - [Swallowing disorder in diffuse idiopathic bone hyperostosis in the area of the cervical vertebrae]. PMID- 15666239 TI - [Hyperdontia--diagnosis in the increased sight of spiral CT]. PMID- 15666240 TI - [Acute anterior cerebral artery infarctS: correlation with early indication of infarct by CT X RAY with CT-perfusion, CT-angiography and conventional angiography]. PMID- 15666241 TI - [Thrombosis of the right sinus transversus with consecutive closing of the ipsilateral vena cava]. PMID- 15666243 TI - Year's comments for 2004. PMID- 15666244 TI - Organic matter in meteorites. AB - Some primitive meteorites are carbon-rich objects containing a variety of organic molecules that constitute a valuable record of organic chemical evolution in the universe prior to the appearance of microorganisms. Families of compounds include hydrocarbons, alcohols, aldehydes, ketones, carboxylic acids, amino acids, amines, amides, heterocycles, phosphonic acids, sulfonic acids, sugar-related compounds and poorly defined high-molecular weight macromolecules. A variety of environments are required in order to explain this organic inventory, including interstellar processes, gas-grain reactions operating in the solar nebula, and hydrothermal alteration of parent bodies. Most likely, substantial amounts of such organic materials were delivered to the Earth via a late accretion, thereby providing organic compounds important for the emergence of life itself, or that served as a feedstock for further chemical evolution. This review discusses the organic content of primitive meteorites and their relevance to the build up of biomolecules. PMID- 15666242 TI - Antenatal presentation of Bardet-Biedl syndrome may mimic Meckel syndrome. AB - Bardet-Biedl syndrome (BBS) is a multisystemic disorder characterized by postaxial polydactyly, progressive retinal dystrophy, obesity, hypogonadism, renal dysfunction, and learning difficulty. Other manifestations include diabetes mellitus, heart disease, hepatic fibrosis, and neurological features. The condition is genetically heterogeneous, and eight genes (BBS1-BBS8) have been identified to date. A mutation of the BBS1 gene on chromosome 11q13 is observed in 30%-40% of BBS cases. In addition, a complex triallelic inheritance has been established in this disorder--that is, in some families, three mutations at two BBS loci are necessary for the disease to be expressed. The clinical features of BBS that can be observed at birth are polydactyly, kidney anomaly, hepatic fibrosis, and genital and heart malformations. Interestingly, polydactyly, cystic kidneys, and liver anomalies (hepatic fibrosis with bile-duct proliferation) are also observed in Meckel syndrome, along with occipital encephalocele. Therefore, we decided to sequence the eight BBS genes in a series of 13 antenatal cases presenting with cystic kidneys and polydactyly and/or hepatic fibrosis but no encephalocele. These fetuses were mostly diagnosed as having Meckel or "Meckel like" syndrome. In six cases, we identified a recessive mutation in a BBS gene (three in BBS2, two in BBS4, and one in BBS6). We found a heterozygous BBS6 mutation in three additional cases. No BBS1, BBS3, BBS5, BBS7, or BBS8 mutations were identified in our series. These results suggest that the antenatal presentation of BBS may mimic Meckel syndrome. PMID- 15666245 TI - Biocontrol mechanisms of Trichoderma strains. AB - The genus Trichoderma comprises a great number of fungal strains that act as biological control agents, the antagonistic properties of which are based on the activation of multiple mechanisms. Trichoderma strains exert biocontrol against fungal phytopathogens either indirectly, by competing for nutrients and space, modifying the environmental conditions, or promoting plant growth and plant defensive mechanisms and antibiosis, or directly, by mechanisms such as mycoparasitism. These indirect and direct mechanisms may act coordinately and their importance in the biocontrol process depends on the Trichoderma strain, the antagonized fungus, the crop plant, and the environmental conditions, including nutrient availability, pH, temperature, and iron concentration. Activation of each mechanism implies the production of specific compounds and metabolites, such as plant growth factors, hydrolytic enzymes, siderophores, antibiotics, and carbon and nitrogen permeases. These metabolites can be either overproduced or combined with appropriate biocontrol strains in order to obtain new formulations for use in more efficient control of plant diseases and postharvest applications. PMID- 15666246 TI - How are gene sequence analyses modifying bacterial taxonomy? The case of Klebsiella. AB - Bacterial names are continually being changed in order to more adequately describe natural groups (the units of microbial diversity) and their relationships. The problems in Klebsiella taxonomy are illustrative and common to other bacterial genera. Like other bacteria, Klebsiella spp. were isolated long ago, when methods to identify and classify bacteria were limited. However, recently developed molecular approaches have led to taxonomical revisions in several cases or to sound proposals of novel species. PMID- 15666247 TI - Virulence genes and intimin types of Shiga-toxin-producing Escherichia coli isolated from cattle and beef products in Argentina. AB - A total of 153 Shiga-toxin-producing Escherichia coli (STEC) isolates from feces of cattle and beef products (hamburgers and ground beef) in Argentina were characterized in this study. PCR showed that 22 (14%) isolates carried stx1 genes, 113 (74%) possessed stx2 genes and 18 (12%) both stx1 and stx2. Intimin (eae), enterohemolysin (ehxA), and STEC autoagglutinating adhesin (saa) virulence genes were detected in 36 (24%), 70 (46%) and in 34 (22%) of the isolates, respectively. None of 34 saa-positive isolates carried the gene eae, and 31 were ehxA-positive. Fourteen (7 of serotype O26:H11 and 4 of serotype O5:H-) isolates had intimin b1, 16 isolates possessed intimin g1 (11 of serotype O145:H- and 5 of serotype O157:H7), 5 isolates had intimin type e1 (4 of serotypes O103:H- and O103:H2), and one isolate O111:H- showed intimin type q/g2. Although the 153 STEC isolates belonged to 63 different seropathotypes, only 12 accounted for 58% of isolates. Seropathotype ONT:H- stx2 (18 isolates) was the most common, followed by O171:H2 stx2 (12 isolates), etc. The majority (84%) of STEC isolates belonged to serotypes previously found in human STEC and 56% to serotypes associated with STEC isolated from patients with hemolytic uremic syndrome (HUS). Thus, this study confirms that cattle are a major reservoir of STEC pathogenic for humans. To our knowledge, this is the first study that described the presence of saa gene in STEC of serotypes O20:H19, O39:H49, O74:H28, O79:H19, O116:H21, O120:H19, O141:H7, O141:H8, O174:H21, and ONT:H21. The serotypes O120:H19 and O185:H7 were not previously reported in bovine STEC. PMID- 15666248 TI - Comparative effect of the fungicide Prochloraz-Mn on Agaricus bisporus vegetative mycelium and fruit-body cell walls. AB - Fungicides to control mycopathogens of commercial Agaricus bisporus, a mushroom cultivated for human consumption, are a major field of study, since these chemicals are toxic to both the host and its fungal parasites. The fungicide Prochloraz-Mn, used at its LD50 for A. bisporus, partially inhibited protein biosynthesis in the vegetative mycelial cell walls of this mushroom and caused significant changes in cell-wall polysaccharide structure, as deduced by methylation analysis and gas liquid chromatography-mass spectrometry (GLC-MS). Furthermore, the aggregated mycelial walls showed distinct alterations in their overall chemical composition following the administration of Prochloraz-Mn at the LD50 and the LD50 x1000. As expected, GLC-MS studies indicated that the latter dose caused more appreciable differences in polysaccharide structure. The decrease in mushroom crop yields obtained from industrial cultures treated with Prochloraz-Mn to control V. fungicola infection depended on the dose of the fungicide employed, whereas fruit-body morphology was only slightly affected at the highest Prochloraz-Mn concentration used. PMID- 15666250 TI - Infectious disease and environment: cholera as a paradigm for waterborne disease. PMID- 15666251 TI - Photokeratoconjunctivitis caused by different light sources. AB - To assess the current state of photokeratoconjunctivitis (PKC) and compare patients caused by different light sources, we recruited 106 consecutive cases from August 2001 to March 2002 through the emergency department at a teaching hospital in Taiwan and conducted questionnaire interviews and telephone follow ups. Whereas 85.8% of the cases received exposures from welding, only 3.8% were professional welders, and most cases were of other occupations with occasional use of welding. A substantial number of cases were attributable to ultraviolet (UV) lamps and sparkles from short circuits (7 each). Cases caused by different light sources (welding, UV lamps, and sparkles) had significant differences in occupations, exposure durations, and distances to light sources. In comparison with cases caused by welding, cases caused by short circuit sparkles had a more acute course, but those caused by UV lamps had a less acute course. Few cases used protection equipment, which highlighted the importance of education. PMID- 15666252 TI - Predicted peak expiratory flow: differences across formulae in the literature. AB - The study objectives were to examine the differences between Peak Expiratory Flow (PEF) formulae in the literature and to assess the potential impact of those differences on the interpretation of clinical guidelines for asthma management. We calculated 100% PEF values for hypothetical patients at the 50(th) percentile for height and weight and classified the percent predicted PEF into severity groups according to national asthma guidelines. Choosing different formulae could give an 18 year old man a 100% predicted PEF as low as 501 L/min and as high as 730 L/min (delta = 229 L/min); and a 35 year old woman a classification of severe (46%) using one, but moderate (57%) using another. Predicted PEF varied widely across formulae and choice of formula may alter guideline-based care. We propose recently published population-based equations as the reference standard for future asthma guidelines. PMID- 15666253 TI - Clinical impact of advanced trauma life support. AB - The Advanced Trauma Life Support (ATLS) course sponsored by the American College of Surgeons Committee On Trauma (ACSCOT) presents a standardized method of initial trauma care. This study attempted to measure any changes in morbidity and mortality in trauma patients after the introduction of ATLS training. Over a 3 year period (May 1996 to September 1997-pre-ATLS period; December 1997 to April 1999-post-ATLS period), 63 trauma patients with an Injury Severity Scale (ISS) > or =16 (n = 31, pre-ATLS and n = 32, post-ATLS) were prospectively studied in two community teaching hospitals. There was no significant difference in mortality rate between groups (48% [15 of 31] pre-ATLS vs. 30% [10 of 32] post-ATLS; P = .203, Fisher exact test). Mortality rates within the ISS range of 16 to 25 were 64% (nine of 14 pre-ATLS) versus 29% (five of 17 post-ATLS), and for the ISS 26 to 35 subgroup, 40% (four of 10 pre-ATLS) versus 25% (two of eight post-ATLS), and within the ISS 36 to 75 subgroup, 29% (two of seven pre-ATLS) versus 43% (three of seven post-ATLS). There was a significant difference in mortality during the first 60 minutes after admission: 0.0% post-ATLS versus 24.2% pre-ATLS (P = .002, Fisher exact test (95% confidence interval ranged from 12-45% in the pre-ATLS group and 0-11% in the post-ATLS group). According to the TRISS methodology (a worldwide-accepted mathematical method to calculate chances of survival through logistical regression),ATLS improved outcome from sub-"Major Trauma Outcome Study" (MTOS) standard results (z = -2.9 to a MTOS standard result z = -0.49). Our data demonstrate that introduction of the ATLS program significantly improved trauma patient outcome in the first hour after admission, as well as improvement from sub-MTOS standard to MTOS standard levels. PMID- 15666254 TI - The accuracy of visual estimation of body weight in the ED. AB - Preventable medical errors may be due to incorrect drug dosage based on poor weight estimation. This study was to examine the accuracy of patient weight estimations in an emergency medical setting. This prospective study enrolled a convenience sample of medically stable adults. The patient's attending physician, resident physician, nurse, a paramedic, and the patient estimated the patient's weight. Of 394 patients enrolled, patients erred in the estimation of their weight by greater than 20% only 1.5% of the time. The group values were 14.7% for attending physicians, 13.4% for resident physicians, 15.9% for nurses, and 17.4% for paramedics. Our study suggests that emergency department staff estimation of a patient's weight is often inaccurate. When available, the patient's own estimate can be used as their actual weight. When the patient is incapacitated, measurement of the patient's weight is the proven method to avoid this type of dosage error. PMID- 15666255 TI - Infections and severe sepsis in solid-organ transplant patients admitted from a university-based ED. AB - The objective was to provide a descriptive analysis of infectious processes in transplant patients admitted from the emergency department (ED). A database of all adult transplant patients at a university medical center was cross-referenced with a computerized record of all ED visits over an 18-month period. ED charts, inpatient records, and microbiology data were retrospectively reviewed. Final diagnoses and outcomes were analyzed. There were 352 ED visits by transplant patients (kidney 66%, kidney/pancreas 15%, liver 13%, lung 3%, heart 3%). Infections were the most common indications for admission (77/217, 35%). Urinary tract infection and pneumonia were the most common infections. Nine of 77 patients (11.7%) with documented infections developed severe sepsis, which was the most common reason for ICU utilization. Thirty-five percent of transplant patients admitted from the ED had acute infections, and 11.7% of these patients had severe sepsis. The emergency physician must have a high index of suspicion for infections when evaluating organ transplant recipients. PMID- 15666256 TI - The effect of limited English proficiency on admission rates from a pediatric ED: stratification by triage acuity. AB - The objectives of this study were to determine the effect of Limited English Proficiency (LEP) visits and acuity status on admission rates from a pediatric ED. A retrospective cohort study was performed using a fully computerized medical record, which includes information on language spoken, triage acuity, and disposition. Data was collected on all patient visits from July 2002 to November 2002 from a tertiary-care pediatric ED. Admission rates and acuity status for LEP and non-LEP patients were compared. A total of 13,585 patient visits were identified, of which 12,416 fit the study criteria. There were 244 LEP patient visits, of which 206 were Spanish-speaking. There were 12,172 English-speaking patient visits. Compared with English-speaking visits, LEP visits were more likely to be triaged as high acuity (25.8% vs. 16.1%, P < .001). LEP patients were more likely to be admitted to the hospital (22.1% vs. 13%, P < .001). For high- and low-acuity patients, no significant differences in admission rates were seen between LEP and English-speaking patients. In contrast, moderate-acuity LEP visits showed a significantly increased admission rate compared to moderate acuity English visits (22.5% vs. 12.4%, P = .005). Similar trends were seen among Spanish-speaking LEP patients. Differences in medical disposition from the ED were found between English-speaking and LEP patient visits. There were higher rates of admission for LEP patients, particularly among moderate-acuity visits. This highlights disparities of care for this vulnerable population. PMID- 15666257 TI - Efficacy and safety in patients on a resuscitator, Oxylator EM-100, in comparison with a bag-valve device. AB - The purpose of this study was to compare the Oxylator EM-100, a ventilator with a fixed flow and working pressure of 25 to 50 cm H(2)O, with a bag-valve device with respect to safety, efficiency, and efficacy when used by professionals. We conducted a prospective, controlled, caregiver-blind single center study. Induction was followed by ventilation with a bag-valve device oxylator in manual and automatic modes. Steps were repeated under hypnosis, after muscle relaxation with mask, and with an endotracheal tube. Forty patients, aged 48.8 +/- 13.5 years weighing 50 to 111 kg were studied. Airway management was consistently better with the bag-valve (P <.0001). Normocapnia was maintained with the bag valve in 88%, oxylator (manual) in 73%, but only 14% of cases in automatic mode with tidal volumes of 1200 mL and more. This device was associated with the potential for rapid hyperventilation in patients when used by professionals. Simplicity of airway management was not improved. PMID- 15666258 TI - Effects of potassium supplementation on the recovery of thyrotoxic periodic paralysis. AB - Potassium supplements have been recommended to hasten recovery and prevent cardiopulmonary complications in patients with thyrotoxic periodic paralysis (TPP). However, this recommendation has not yet been proven efficacious. Thirty two patients with acute attacks of TPP over a 3-year-period were divided into 2 groups. Group A (n = 12) was a control group treated with normal saline infusion 125 mL/hr only. Group B (n = 20) received intravenous KCl administration at a rate of 10 mmol/hr in normal saline 125 mL/hr. During the attack and for 6 hours after muscle recovery, hemodynamics were continuously recorded and muscle strength and plasma K(+) concentration were measured hourly. The sex, age, muscle strength, thyroid function, biochemical values including plasma K(+) levels, as well as the time from attack to therapy (3.6 +/- 1.6 v 3.3 +/- 1.0 hr) were not significant between the 2 groups. However, recovery time was significantly shorter in the KCl group than the control (6.3 +/- 3.8 v 13.5 +/- 7.5 hr, P < .01). Rebound hyperkalemia greater than 5.5 mmol/L occurred in 40% patients receiving KCl. The dose of KCl administered and peak K(+) concentration were positively correlated (r = 0.85, P < .001). In conclusion, KCl therapy proves to help the recovery of paralysis in TPP associated with rebound hyperkalemia. KCl supplementation should be given as small as possible (<10 mmol/hr) to avoid rebound hyperkalemia unless there are cardiopulmonary complications. PMID- 15666260 TI - Influence of circadian rhythm on mortality after myocardial infarction: data from a prospective cohort of emergency calls. AB - Myocardial infarction (MI) occurs more frequently in the morning as a result of the concomitant unfavorable timing of several physiological parameters and/or biochemical conditions. However, little is known about the possible influence of this circadian pattern on prognosis. To evaluate whether the time of symptom onset could potentially influence mortality from acute MI, this prospective study considered all consecutive MIs admitted to the ED of Ferrara, Italy, after a call to the Emergency Coordinating Unit from January 1, 1998, to December 31, 2001. The total sample consisted of 442 MIs (mean age, 68.7 years; males, 72%). Eighty patients (males, 82.5%) died in the ED; the remaining 362 were admitted to the hospital. Of these, 50 (males, 60%) died during their hospital stay. Based on the timing of their symptom onset, cases were categorized both into 24 1-hour intervals and four 6-hour intervals (midnight to 5:59 am, 6:00 am to 11:59 am, noon to 5:59 pm, and 6:00 pm to 11:59 pm), and the circadian distributions of fatal versus nonfatal MIs were compared. The circadian variation of MI peaked between 6:00 am and noon (P < .001), and in this period, there was a trend toward a higher frequency of fatal cases (41.5% vs. 35.2%; chi(2) = 1.911, P = .167). To verify whether this higher frequency of fatal events in the morning hours could be related to possible higher severity of cases observed in that hours, a further separate analysis considering age, infarct site, and peak levels of MB was made. Again, no significant temporal differences among the four 6-hour intervals were found between fatal and nonfatal Mis, although a trend toward older age was observed in morning MIs. Not only the frequency, but also the mortality, of acute MI could be increased in the morning hours. This could be of practical interest for emergency doctors and could have significant implications for acute treatment, because several studies have reported a lowered efficacy of thrombolytic drugs in the morning hours. PMID- 15666259 TI - Prolonged gastric emptying half-time and gastric hypomotility after drug overdose. AB - A prospective study was undertaken to determine if gastric motility and emptying are altered by the ingestion of overdoses of tricyclic antidepressants, acetaminophen, opioid-acetaminophen mixtures, carbamazepine or phenytoin. Gastric scintigraphy was used to measure gastric emptying half-time and assess gastric motility in 104 patients at initial presentation and again at follow-up (n = 85). Patients were imaged for 5 hours after being given 20 MBq of 99mTc tin colloid to drink. Drug serum levels were measured on all patients at initial presentation and at follow-up. We observed markedly prolonged gastric emptying half-times and severe hypomotility at initial presentation compared with follow-up in the vast majority of patients, except for a small group of patients with phenytoin poisoning. Twelve patients had gastric emptying half-times of over 300 minutes, a further 14 had half-times of over 200 minutes and 21 others had half-times of over 120 minutes. Poisoning is associated with hypomotility and a marked delay in gastric emptying that could influence the clinical course and patient management. These abnormalities may not be due to a direct effect of the ingested drug and factors such as stress may play a role. PMID- 15666261 TI - Effect of season, age, and gender on renal colic incidence. AB - Our objective was to examine the effect of ambient temperature, age, and gender on the incidence of emergency department (ED) renal colic visits. We retrospectively analyzed a database of 15 New Jersey EDs from January 1, 1996 to December 31, 2002. We analyzed the number of renal colic visits as a fraction of total visits in monthly intervals. We used the Chi-squared test and Pearson's correlation coefficient, with P<.05 taken as statistically significant. Of the 3.5 million patient visits in the database, 30,358 (0.9%) had renal colic. Renal colic visits were 16% more likely in warmer than colder months (P<.001) and this effect was greatest in older patients and males. We conclude that higher ambient temperature, older age and male gender are associated with increased incidence of ED renal colic visits. Advice to patients, especially older males, to avoid dehydration particularly during hot weather may help prevent bouts of renal colic. PMID- 15666262 TI - The ED is an efficient place to treat ED patients. AB - The purpose of this study was to determine what percentage of ED patients would be more efficiently treated in an offsite clinic. A stratified sample consisting of 650 ED visits were reviewed. Patient care was classified as more efficiently delivered in ED or clinic using prospectively developed criteria. Five hundred fifty-three (85%; 95% confidence interval [CI], 82-88%) visits met the definition of efficient ED utilization. One (0.15%; 95% CI, 0.01-0.99%) met the criteria for efficient clinic utilization. The other 96 (15%; 95% CI, 12-18%) failed to meet the definition of efficient for either site; 92 of these failed 2 or more clinic criteria. Only 25 (3.9%; 95% CI, 2.6-5.7%) were seen for nonacute problems. Few ED patients would be more efficiently seen in a clinic. PMID- 15666263 TI - Clinical features, triage, and outcome of patients presenting to the ED with suspected acute coronary syndromes but without pain: a multicenter study. AB - We studied the impact on triage and outcome of patients presenting to the emergency department (ED) with symptoms suggestive of an acute coronary syndromes (ACS) but without a complaint of pain. Data from a prospective clinical trial of patients with symptoms suggesting an ACS in the EDs of 10 US hospitals comparing patient demographics, clinical variables, and outcomes was used to perform a secondary analysis. Of 10,783 subjects, a final diagnosis of an ACS was confirmed in 24% of which 35% had acute myocardial infarction (AMI) and 65% unstable angina pectoris (UAP). Pain was absent in 6.2% of patients with acute ischemia and in 9.8% of those with AMI. Compared to similar patients who presented with pain, patients with painless ischemia were older, were more commonly women, had more cardiac and related diseases. Among patients with AMI, fewer patients without pain were admitted to critical care units compared to similar patients with pain. Among patients with AMI, logistic regression predicting lack of pain identified age, heart failure and diabetes, with only age and heart failure among all with ACS. After controlling for clinical features, lack of pain during acute ischemia predicted increased hospital mortality. We concluded that age and heart failure are independently associated with painless ACS, in addition to diabetes among those with AMI. Lack of pain predicts increased hospital mortality in patients with ACI through mechanisms that remain to be elucidated. There is a need for greater awareness in the general public of the different manifestations of ACS to enhance the recognition of and prompt response to their symptoms. PMID- 15666264 TI - The state of ED on-call coverage in California. AB - The ED provides initial treatment, but failure of specialists to respond unravels the safety net. To assess the scope of problems with on-call physicians in California. A mailed anonymous survey to all CAL/ACEP physician members (1876) asking patient, physician and ED demographics, specialist availability for consultation, insurance profile, and availability of follow-up care. 608/1876 physicians responded (32.4%), representing 320/353 California EDs (90.6%). The seven specialties in which the greatest proportion of EDs reported trouble with specialty response were: plastic surgery (37.5%), ENT (35.9%), dentistry (34.9%), psychiatry (26.0%), neurosurgery (22.9%), ophthalmology (18.4%) and orthopedics (18.0%). 71.6% of responder EDs reported that their medical staff rules required ED on-call coverage. However, the percentage of responders who stated that hospitals paid each specialty for call was low: neurosurgery (37.3%), orthopedics (34.4%), ENT (17.9%), plastic surgery (15.1%) and ophthalmology (13.1%). On-call problems were more acute at night (77.2%) or on weekends (72.4%). Patient insurance negatively affected (69.9%) willingness of on-call physicians to consult for at least a quarter of patients. Regarding follow-up, 91% reported some trouble, whereas 64% reported a problem at least half the time. Surgical sub specialists are the most problematic on-call physicians. Insurance status has a major negative effect on ED and follow-up care. The on-call situation in California has reached crisis proportions. PMID- 15666265 TI - Mandated pain scales improve frequency of ED analgesic administration. AB - A retrospective study design was used to determine the effect of introducing a mandated verbal numeric pain scale on the incidence and timing of analgesic administration in the ED. Consecutive patients presenting with renal colic, extremity trauma, headache, ophthalmologic trauma, and soft tissue injury were included. 521 encounters were reviewed before and 479 encounters after the introduction of the pain scale. Groups were similar in baseline characteristics. Analgesic use increased from 25% to 36% (p < 0.001), and analgesics were administered more rapidly after the scale was introduced (113 minutes vs. 152 minutes, p = 0.09). Analgesic use correlated with pain severity. Patients undergoing diagnostic testing were less likely to receive analgesics, especially when presenting with a headache (p < 0.001). We conclude that use of a pain scale at triage significantly increases use of analgesia, and shortens the time till its administration. Patients undergoing diagnostic workups were less likely to receive analgesia. PMID- 15666266 TI - Control of massive bleeding from facial gunshot wound with a compact elastic adhesive compression dressing. AB - The issue of immediate control of acute external traumatic hemorrhage is poorly dealt with in the medical literature. A compact unit incorporating the desired components capable of applying significant compression over diverse body areas has been suggested in the past but not formally demonstrated effective in practice. In this report we describe the treatment of a young man presenting with a very large, complex, profusely bleeding facial gunshot wound. The calvarium remained intact. The upper airway was diverted and secured by performing a cricothyroidotomy. A transparent elastic adhesive dressing was then applied by covering the anterior aspect of the face by a contact pad followed by sequential wrapping of the roll covering all structures between the forehead and the neck. This procedure successfully controlled the hemorrhage and maintained the victim's condition hemodynamically stable until definitive surgical intervention at the level 1 medical center. We attribute the survival of this victim to the innovative dressing technique and excellent cooperation between the trauma team and hospital staff. This case demonstrates the contribution of elastic adhesive compression dressing towards saving the lives of those inflicted by severely challenging bleeding wounds. We suggest this technique be considered by Emergency personnel working in the prehospital arena in selected cases. PMID- 15666267 TI - Water bath evaluation technique for emergency ultrasound of painful superficial structures. AB - Researchers have described the use of bedside emergency ultrasound as an effective way to evaluate for and accurately drain potential abscesses. Similarly, descriptions exist of long bone fracture evaluation in the wrist and hands. Tendon injury can also be detected with ultrasound and exploration can be obviated or at least focused. Sonographic examination of painful extremity pathology such as abscesses or lacerations involving the hand or foot can be challenging. Patients may be uncooperative if they experience significant pain when the transducer is placed on the area of interest. While ample amounts of ultrasound gel can decrease the need for firm transducer contact with the skin it is still difficult to obtain a good evaluation without causing any discomfort. The solution may lie in an old technique that has been recently brought back to life for use in hand evaluation in which the patient's extremity is placed in a water bath. The water bath replaces the need for ultrasound gel or contact between the ultrasound transducer and the patient's skin, thus eliminating discomfort. We describe 7 cases in which, despite aggressive attempts at pain control, adequate evaluation of extremity pathology was not possible without the use of the water bath technique. Patients reported no discomfort and superior quality images were obtained due to the water bath properties. Emergency sonologists should keep this technique in mind when contact between skin and the ultrasound transducer is likely to cause a patient significant discomfort. PMID- 15666268 TI - Preventing tourniquet effect when dressing finger wounds in children. AB - Hand and finger injuries are common injuries in children seen in emergency departments. Many of these are soft tissue injuries, which are often caused by household items, such as doors and exercise equipment. Once these injuries are repaired, usually they are covered with a simple dressing using a material such as Coban (3M, Minneapolis, MN). This dressing often is applied by winding it circumferentially around the digit. However, with very little manipulation, this dressing can be lifted and rolled up the digit in a distal direction, creating a tourniquet effect, which can cause hypoxia and tissue necrosis. To prevent the tourniquet effect, the dressing must include the hand and wrist. PMID- 15666270 TI - Bedside emergency ultrasonographic diagnosis of diaphragmatic rupture in blunt abdominal trauma. AB - Abdominal injury from significant blunt trauma can include injury to bowel, kidneys, liver, and spleen. In approximately 5% of all injuries one of the diaphragms is ruptured. Diaphragmatic rupture may not be easily detected and this can lead to significant morbidity and even mortality. Rupture may be suggested on chest X-ray film especially with abnormal nasogastric tube location but the accuracy of this method is modest only. Abdominal computed tomography is not accurate and magnetic resonance imaging, although very sensitive and specific, is not feasible in most trauma situations. Surgeons have often resorted to exploratory laparotomy or laparoscopy to make the diagnosis. Although not typically part of the basic Focused Abdominal Sonography for Trauma (FAST) examination, ultrasonographic diagnosis of diaphragmatic rupture is possible with little added time to the examination. We present 3 cases of diaphragmatic rupture discovered shortly after the patients' arrival, on initial trauma evaluation with the FAST. A discussion of previous literature and ultrasound technique for diagnosis follows the cases. PMID- 15666269 TI - Orthopedic pitfalls in the ED: Achilles tendon rupture. AB - Achilles tendon rupture is a relatively uncommon occurrence in a general ED population. The history can be subtle, and physical findings may not be clear cut. Prompt diagnosis and treatment of these injuries, however, is important to improved clinical outcome. The emergency physician needs to remain vigilant for this diagnosis to avoid this orthopedic pitfall. This review article examines the clinical presentation, diagnostic technique, and management options applicable to the emergency physician in the treatment of Achilles tendon rupture. PMID- 15666271 TI - CT diagnosis of fat embolism. PMID- 15666272 TI - Orthopedic pitfalls in the ED: calcaneal fractures. AB - Fractures of the calcaneus are usually associated with a high-force mechanism, which frequently can involve associated injuries and prolonged disability. Due to distracting injury and variations in clinical findings, calcaneal fractures may be initially missed or misdiagnosed. This review article examines the clinical presentation, diagnostic techniques, and management of calcaneal fractures applicable to the emergency practitioner. PMID- 15666273 TI - A new pepsin assay to detect pulmonary aspiration of gastric contents among newly intubated patients. AB - Aspiration of gastric contents by endotracheally intubated patients is associated with significant morbidity and mortality. Previous studies suggest that pepsin in tracheal aspirates may be a valuable marker of occult aspiration. We sought to show the sensitivity and specificity of a new, pepsin-specific assay in humans. A prospective, case-controlled study was conducted with subjects serving as their own controls. After planned endotracheal and nasogastric intubation for elective surgery, 20 participants had tracheal and gastric aspirates withdrawn. A blinded investigator tested samples for the presence of pepsin using the assay. Positive samples were then tested with pepstatin, a specific pepsin inhibitor, to ensure that positive results were due to pepsin. All tracheal aspirates tested negative and all gastric aspirates tested positive for pepsin. Pepstatin halted pepsin activity in all positive samples, ensuring that positive results were due to pepsin. A pepsin-specific assay is extremely reliable for detecting gastric contents in humans. PMID- 15666274 TI - An uncommon life-threatening complication after chest tube drainage of pneumothorax in the ED. AB - Re-expansion pulmonary oedema (REPO) after chest tube drainage of pneumothorax is uncommon. We contrast one patient with life threatening against another mildly symptomatic REPO occurring in our emergency department (ED). The mechanism and management of REPO differs distinctly from that of cardiogenic pulmonary oedema. We discuss the predictors of REPO, review clinical details of reported fatalities and suggest preventive measures. PMID- 15666275 TI - Methanol poisoning and heparin: a dangerous couple? PMID- 15666276 TI - Endometrioma of the abdominal wall masquerading as hernia. PMID- 15666277 TI - Janos Balassa and the first CPR. PMID- 15666278 TI - Osborne wave in a patient with tachycardia. PMID- 15666279 TI - Commentary on: Severe manifestations of coricidin intoxication. PMID- 15666280 TI - Intractable vomiting as the initial presentation of pericardial effusion. PMID- 15666281 TI - Serotonin syndrome resulting from an herbal detox cocktail. PMID- 15666282 TI - Traumatic medial cerebral artery occlusion in a 4-year-old child. PMID- 15666283 TI - Acute viral myocarditis presenting as sudden cardiac arrest and refractory ventricular tachycardia. PMID- 15666284 TI - The spontaneous splenic rupture as first manifestation of mantle cell lymphoma, a dangerous rarity. PMID- 15666286 TI - Allergic reactions from insect bites. PMID- 15666285 TI - Cortical blindness in preeclampsia. PMID- 15666287 TI - Fresh frozen plasma for acute exacerbations of hereditary angioedema. PMID- 15666290 TI - Rapid pKa estimation using vacuum-assisted multiplexed capillary electrophoresis (VAMCE) with ultraviolet detection. AB - A rapid approach for estimating the pK(a) value of small organic molecules was developed using vacuum-assisted multiplexed capillary electrophoresis (VAMCE) with ultraviolet detection. The VAMCE method employed a 96-capillary array, arranged in a standard 8 x 12 microtiter plate configuration, with each row of capillaries filled with 12 individual running buffers of equal ionic strength (I = 50 mM) covering a pH range from 2.2 to 10.7. A separate compound was injected hydrodynamically into each row of capillaries allowing the estimation of pK(a) values for eight compounds in a single run. The application of a vacuum during the separation generated a bulk fluid flow and allowed the electrophoretic separation to be completed within 5 min. The complete VAMCE method, conditioning, and electrophoretic separation was optimized to allow the pK(a) estimation for between 128 to 168 compounds in an 8-h period. The VAMCE method provided a reliable approach for estimating pK(a) values both within- and between-day. The pK(a) values for a series of 96 compounds estimated by VAMCE agreed well with some of literature pK(a) values with an average absolute difference of 0.22 log units. PMID- 15666291 TI - Reductase-mediated metabolism of motexafin gadolinium (Xcytrin) in rat and human liver subcellular fractions and purified enzyme preparations. AB - The biotransformation of motexafin gadolinium (MGd, Xcytrin) was investigated in subcellular rat and human liver fractions. Microsomal MGd metabolism was dependent on NADPH in both species. Cytosolic metabolism in rat and human livers was dependent on NADPH or NADH. Under anaerobic conditions, MGd metabolism increased in liver microsomes and purified enzyme preparations. Cytochrome P450 (CYP450) inhibitors ketoconazole, proadifen, and carbon monoxide increased NADPH dependent MGd metabolism in microsomes. Treatment of rats with beta naphthoflavone increased cytosolic metabolism of MGd twofold, but had no effect on microsomal metabolism. Conversely, in liver preparations from phenobarbital treated rats microsomal metabolism of MGd was enhanced twofold, but not in cytosolic preparations. Purified CYP450 reductase from phenobarbital-treated rabbit or untreated human livers metabolized MGd suggesting involvement of CYP450 reductase. Dicumarol, a potent DT-diaphorase inhibitor, inhibited MGd metabolism in both rat and human liver cytosol. These data suggest MGd metabolism in rat liver involves CYP450 reductase and/or DT-diaphorase. In human liver preparations only CYP450 reductase is directly involved in MGd metabolism. A metabolite identified in microsomes and cytosol is a metal-free, reduced form of MGd, indicating that both enzymes generate metabolite 1, which appears to be PCI-0108, a synthetic precursor to MGd. PMID- 15666293 TI - Carrier mediated transport of chlorpheniramine and chlorcyclizine across bovine olfactory mucosa: implications on nose-to-brain transport. AB - Delivery to the CNS via the nasal cavity has been pursued as a means to circumvent the blood-brain barrier (BBB), yet the mechanism of drug transport across this novel route is not well understood. Hydroxyzine and triprolidine have been reported to readily reach the CNS following nasal administration, whereas no measurable amounts of chlorcyclizine or chlorpheniramine, structurally similar antihistamines, were observed in the CSF. The permeation of chlorpheniramine and chlorcyclizine in vitro across the bovine olfactory mucosa was studied to investigate the biological and physicochemical characteristics that contribute to the limited CNS disposition of these compounds following nasal administration. The submucosal to mucosal fluxes (J(s-m)) of chlorcyclizine and chlorpheniramine across the olfactory mucosa were significantly greater than the mucosal to submucosal fluxes (J(m-s)). Moreover, the submucosal-mucosal permeability of both compounds was temperature dependent and saturable. In the presence of metabolic inhibitors (ouabain and 2,4-dinitrophenol) and P-glycoprotein (P-gp)/multidrug resistance protein 1 (MRP1) inhibitors (quinidine and verapamil), the J(m-s) increased and J(s-m) decreased significantly. These results indicate that chlorpheniramine and chlorcyclizine are effluxed from the olfactory mucosa by efflux transporters such as P-gp and MRP1. Transport studies across inert polymeric membranes demonstrated that the permeability of chlorpheniramine and chlorcyclizine decreased at donor concentrations higher than 3 mM suggesting that physicochemical properties such as self-aggregation also play a role in the reduced olfactory mucosal permeability of these compounds at higher concentrations. PMID- 15666292 TI - Pharmacodynamic interactions between recombinant mouse interleukin-10 and prednisolone using a mouse endotoxemia model. AB - The pharmacodynamic interactions between recombinant mouse interleukin-10 (IL-10) and prednisolone were examined in lipopolysaccharide (LPS)-induced experimental endotoxemia in Balb/c mice. Treatment phases consists of single doses of IL-10 (10 microg/kg i.p.), prednisolone (25 (mg/kg i.p.), IL-10 (2.5 microg/kg i.p.) with prednisolone (6.25 mg/kg i.p.), or placebo (saline). Measurements included plasma steroid kinetics and IL-10 concentrations and responses to LPS including proinflammatory cytokines (TNF-alpha, IFN-gamma) and circulatory NO measured as plasma nitrate/nitrite concentrations. The intraperitoneal dosing of LPS produced large and transient elevations of plasma TNF-alpha, IFN-gamma, and NO concentrations. Noncompartmental and model fitting using extended indirect response models based on drug inhibition of multiphase stimulation of biomarkers by LPS were used to describe the in vivo pharmacodynamics and drug interactions. Dosing with prednisolone, IL-10, or their combinations produced strong inhibition of cytokine and NO production. The IC50 values of prednisolone ranged from 54 to 171 ng/mL, and IC50 values for IL-10 ranged from 0.06 to 0.69 ng/mL. The production of NO was described as a cascading consequence of the TNF-alpha and IFN-gamma plasma concentrations. The joint dosing of IL-10 with prednisolone produces moderately synergistic immunosuppressive effects in this system. Both drugs were sufficiently protective in suppressing the inflammatory mediators when administered prior to the LPS trigger, while such effects were modest when administered after the inflammatory stimulus was provoked. The integrated and complex pharmacokinetic/pharmacodynamic models well capture the in vivo processes, drug potencies, and interactions of IL-10 and prednisolone. PMID- 15666294 TI - Kinetics of glycine crystallization during freezing of sucrose/glycine excipient systems. AB - Isothermal calorimetry and low-field nuclear magnetic resonance were used to measure crystallization of glycine during annealing of glycine/sucrose mixtures, a commonly-used excipient system for freeze-dried proteins. Kinetics of crystallization of glycine were consistent between the two methods, although the NMR method was significantly more sensitive. By the calorimetric method used here, sensitivity was lost when the total solute concentration was below about 20% (w/v) and the relative glycine concentration below about 35% of the total solids. By the NMR method, total solute concentrations as low as 5% (w/v) could be studied. When the relative concentration of glycine is below about 30% of total solids, the time course of crystallization becomes excessively long for practical freeze-drying applications. A good fit of the crystallization data was obtained with the Johnson-Mehl-Avrami (JMA) equation. The Avrami exponent of 2.5 is consistent with diffusion-limited spherulitic growth of glycine. PMID- 15666295 TI - A factor analysis for complex systems containing nimesulide. AB - Binary systems containing Nimesulide and PEG 4000 were prepared by the melting method in the concentration range 3-25% w/w of the drug. The systems are homogeneous in the molten state, while, after cooling, two phases were formed of different density. They were manually separated and separately studied. Upper phases are richer in PEG 4000, while the lower ones contain the drug at levels even higher than those of the starting mixtures. The two phases were examined by DSC and UV techniques; high dissolution rates were observed with upper phases, while lower phases did not display improvement with respect to a physical mixture or micronized drug. With the aim to avoid phase separation, a third component was added to the binary system containing 5% w/w drug, during the melting. The ternary systems were prepared containing sodium dodecyl sulfate, triethanolamine, polysorbate 80, poloxamer, and cetomacrogol: a homogeneous phase was obtained only in two cases (with the addition of sodium dodecyl sulfate and triethanolamine), but only in the presence of triethanolamine dissolution rate was improved. Finally, a factor analysis was performed for complex systems containing a combination of the four additives, each one at two concentrations (1.25 and 2.5% w/w), to evaluate the optimum system in terms of both kinetic and composition parameters. Results suggest that additives affect mainly the physical aspect of the formulation rather than the kinetic behavior, which appears little improved only in a few cases. PMID- 15666296 TI - Structural analysis of polymorphism and solvation in tranilast. AB - Five polymorphic forms of tranilast were characterized by thermal, diffractometric, and spectroscopic techniques. The crystal structures of the most stable anhydrous form (Form I), a chloroform solvate, and a dichloromethane solvate were determined from single-crystal X-ray analysis. Two additional anhydrous forms of tranilast (Forms II and III) were also studied, but were not amenable to SCXRD. All five forms were also analyzed using solid-state nuclear magnetic resonance, Fourier transform infrared, and Fourier transform-Raman spectroscopy, and thermal methods. From the trends observed in the crystal structures and the spectral data, some conclusions can be made about hydrogen bonding, molecular conformation, and crystal packing differences in the polymorphs and solvates. Form II was found to be a spectroscopically distinctive polymorph that is probably missing an important intramolecular hydrogen bond coupled with a conformational change. In contrast, Form III was found to be more similar to the crystallographically characterized forms, and is more likely a packing and hydrogen-bonding polymorph with a weakened intermolecular hydrogen bonding interaction relative to the other forms. From a pharmaceutical development perspective, it is shown that although the anhydrous forms of tranilast have similar thermal properties, they can be reliably distinguished by spectroscopic methods. PMID- 15666297 TI - Determination of fluidized bed granulation end point using near-infrared spectroscopy and phenomenological analysis. AB - Simultaneous real-time monitoring of particle size and moisture content by near infrared spectroscopy through a window into the bed of a fluidized bed granulator is used to determine the granulation end point. The moisture content and particle size determined by the near-infrared monitor correlates well with off-line moisture content and particle size measurements. The measured particle size is modeled using a population balance approach, and the moisture content is shown to follow accepted models during drying. Given a known formulation, with predefined parameters for peak moisture content, final moisture content, and final granule size, the near-infrared monitoring system can be used to control a fluidized bed granulation by determining when binder addition should be stopped and when drying of the granules is complete. PMID- 15666298 TI - Inter- and intralaboratory variation of in vitro diffusion cell measurements: an international multicenter study using quasi-standardized methods and materials. AB - In vitro measurements of skin absorption are an increasingly important aspect of regulatory studies, product support claims, and formulation screening. However, such measurements are significantly affected by skin variability. The purpose of this study was to determine inter- and intralaboratory variation in diffusion cell measurements caused by factors other than skin. This was attained through the use of an artificial (silicone rubber) rate-limiting membrane and the provision of materials including a standard penetrant, methyl paraben (MP), and a minimally prescriptive protocol to each of the 18 participating laboratories. "Standardized" calculations of MP flux were determined from the data submitted by each laboratory by applying a predefined mathematical model. This was deemed necessary to eliminate any interlaboratory variation caused by different methods of flux calculations. Average fluxes of MP calculated and reported by each laboratory (60 +/- 27 microg cm(-2) h(-1), n = 25, range 27-101) were in agreement with the standardized calculations of MP flux (60 +/- 21 microg cm(-2) h(-1), range 19-120). The coefficient of variation between laboratories was approximately 35% and was manifest as a fourfold difference between the lowest and highest average flux values and a sixfold difference between the lowest and highest individual flux values. Intralaboratory variation was lower, averaging 10% for five individuals using the same equipment within a single laboratory. Further studies should be performed to clarify the exact components responsible for nonskin-related variability in diffusion cell measurements. It is clear that further developments of in vitro methodologies for measuring skin absorption are required. PMID- 15666299 TI - DLX3 mutation associated with autosomal dominant amelogenesis imperfecta with taurodontism. AB - Amelogenesis imperfecta hypoplastic-hypomaturation with taurodontism (AIHHT) is an autosomal dominant (AD) trait associated with enamel defects and enlarged pulp chambers. In this study, we mapped an AIHHT family to human chromosome 17 q21-q22 (lod score 3.3) and identify a two basepair deletion (CT) at nucleotide 560 in DLX3 associated with the disease. This mutation causes a frameshift altering the last two amino acids of the DNA-binding homeodomain introducing a premature stop codon truncating the protein by 88 amino acids. This is the first report of a mutation within the homeodomain of DLX3. Previous studies have shown a DLX3 mutation outside the homeodomain associated with tricho-dento-osseous syndrome (TDO) suggesting TDO and some forms of AIHHT are allelic. PMID- 15666300 TI - GJB2 mutations: passage through Iran. AB - Hereditary hearing loss (HHL) is a very common disorder. When inherited in an autosomal recessive manner, it typically presents as an isolated finding. Interestingly and unexpectedly, in spite of extreme heterogeneity, mutations in one gene, GJB2, are the most common cause of congenital severe-to-profound deafness in many different populations. In this study, we assessed the contributions made by GJB2 mutations and chromosome 13 g.1777179_2085947del (the deletion more commonly known as del (GJB6-D13S1830) that includes a portion of GJB6 and is hereafter called Delta(GJB6-D13S1830)) to the autosomal recessive non syndromic deafness (ARNSD) genetic load in Iran. Probands from 664 different nuclear families were investigated. GJB2-related deafness was found in 111 families (16.7%). The carrier frequency of the 35delG mutation showed a geographic variation that is supported by studies in neighboring countries. Delta(GJB6-D13S1830) was not found. Our prevalence data for GJB2-related deafness reveal a geographic pattern that mirrors the south-to-north European gradient and supports a founder effect in southeastern Europe. PMID- 15666301 TI - Interstitial deletion 11(p11.12p11.2) and analphoid marker formation results in inherited Potocki-Shaffer syndrome. AB - We report a family with inherited Potocki-Shaffer syndrome. The phenotypically normal mother has an interstitial deletion of 11(p11.12p11.2) with neocentric marker chromosome formation. The marker chromosome contains the deleted material on 11p11.2 and is likely a ring. The patient inherited a maternal deleted chromosome 11 but not the marker chromosome, thus resulting in an unbalanced karyotype along with the phenotype of Potocki-Shaffer syndrome. The deleted region in our case-11p11.12p11.2-is a newly reported site of constitutional neocentromere formation. This is also the first report describing deletion of 11p11.12-p11.2 and neocentromere formation resulting in inherited Potocki-Shaffer syndrome. PMID- 15666302 TI - CHARGE syndrome "behaviors": challenges or adaptations? PMID- 15666303 TI - Trisomy 15q25.2-qter in an autistic child: genotype-phenotype correlations. AB - We report on the case of a male child with autistic disorder, postnatal overgrowth, and a minor brain malformation. Karyotyping and fluorescent in situ hybridization (FISH) analysis showed the presence of an extra copy of the distal portion of chromosome 15q (15q25.2-qter) transposed to chromosome 15p leading to 15q25.2-qter pure trisomy. This karyotype-phenotype study further supports the evidence for a specific phenotype related to trisomy 15q25 or 26-qter and suggests that distal chromosome 15q may be implicated in specific behavioral phenotypes. PMID- 15666304 TI - Craniofacial features in osteogenesis imperfecta: a cephalometric study. AB - Osteogenesis imperfecta (OI) is a heterogeneous group of connective tissue diseases that mainly manifest as bone fragility and skeletal deformity. In most families it segregates as a dominant trait and results from mutations in type I collagen genes. In this study we analyzed the size and form of the bony structures in heads of 59 consecutive patients with OI types I, III, or IV (Sillence classification), using lateral radiographs. Paired controls were matched for gender and age. The purpose was to obtain baseline information of craniofacial development in OI patients that have not received bisphosphonate treatment. In OI type I we found smaller than normal linear measurements, indicating a general growth deficiency, but no remarkable craniofacial deformity. In OI types III and IV, the growth impairment was pronounced, and the craniofacial form was altered as a result of differential growth deficiency and bending of the skeletal head structures. We found strong support both for an abnormally ventral position of the sella region due to bending of the cranial base, and for a closing mandibular growth rotation. Vertical underdevelopment of the dentoalveolar structures and the condylar process were identified as the main reasons for the relative mandibular prognathism in OI. Despite of the widespread intervention with bisphosphonates, the facial growth impairment will probably remain characteristic for many OI patients, and their orthodontic treatment should be further developed. PMID- 15666305 TI - Familial neurocardiogenic (vasovagal) syncope. AB - Vasovagal syncope (VSS) is an exaggerated tendency towards the common faint caused by a sudden and profound hypotension with or without bradycardia. The etiology of VVS is unknown though several lines of evidence indicate central and peripheral abnormalities of sympathetic function. Studies however indicate a strong heritable component to the etiology of VVS in over 20% of cases. Here, we report the findings from a family that shows apparently autosomal dominant VVS in at least three generations. Clinical findings included an absence of any discernible cardiac or autonomic abnormalities and reproducible hypotension on tilt table testing in affected family members. PMID- 15666306 TI - Psychological benefit of diagnostic certainty for mothers of children with disabilities: lessons from Down syndrome. AB - Diagnostic and prognostic uncertainty is one of the major psychological stressors for patients in acute and chronic illness, as well as for parents of children with disabilities or chronic disease. Whereas the parents' feeling of uncertainty is undoubtedly very strong shortly after the birth of a child with disabilities, the long-term effects on the parents of having or not having a precise genetic diagnosis, in terms of emotional stress, remain unclear. In this study, mothers of non-disabled children are compared to mothers of children with Down syndrome, and to mothers of children with a diagnostically unassigned mental retardation with regard to the level of anxiety, feelings of guilt, and emotional burden. While the mothers of children with Down syndrome score comparably to the mothers of non-disabled children, the results show broad psychoemotional disadvantages for mothers of children with a mental retardation of unknown etiology. Consequently, the value of genetic diagnosis of infantile disabilities encompasses, beyond clinical considerations like therapy planning and assignment of the recurrence risk for siblings, significant and long-lasting emotional relief for the parents. PMID- 15666307 TI - Isolated elevated sweat chloride concentrations in the presence of the rare mutation S1455X: an extremely mild form of CFTR dysfunction. AB - Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) have been shown to cause typical cystic fibrosis (CF) and several milder phenotypes. We report on two asymptomatic sisters who had isolated increased sweat chloride concentrations, and in whom systematic scanning of the whole coding region of the CFTR gene revealed the F508del/S1455X genotype. PMID- 15666308 TI - Updated diagnostic criteria for CHARGE syndrome: a proposal. PMID- 15666309 TI - Nevo syndrome is allelic to the kyphoscoliotic type of the Ehlers-Danlos syndrome (EDS VIA). AB - We report on seven patients affected with Nevo syndrome, a rare, autosomal recessive disorder characterized by increased perinatal length, kyphosis, muscular hypotonia, and joint laxity. Since its first description by Nevo et al. [1974], only a few cases have been reported. Because some of these patients present clinical features similar to those of the kyphoscoliotic type of Ehlers Danlos syndrome (EDS VIA), an inherited connective tissue disorder characterized by a deficiency of lysyl hydroxylase due to mutations in PLOD1, we studied seven patients with Nevo syndrome, three of whom have previously been reported, and four of whom are new. In the five patients from whom urine was available, the ratio of total urinary lysyl pyridinoline (LP) to hydroxylysyl pyridinoline (HP) was elevated (8.2, 7.8, 8.6, 3.5, and 4.8, respectively) compared with that in controls (0.20 +/- 0.05, range 0.10-0.38), and similar to that observed in patients with EDS VIA (5.97 +/- 0.99, range 4.3-8.1). Six patients were homozygous for a point mutation in exon 9 of PLOD1 causing a p.R319X nonsense mutation, while one patient was homozygous for a large deletion comprising exon 17 of PLOD1. We conclude that the Nevo syndrome is allelic to and clinically indistinguishable from EDS VIA, and present evidence that increased length at birth and wristdrop, in addition to muscular hypotonia and kyphoscoliosis, should prompt the physician to consider EDS VIA earlier than heretofore. PMID- 15666310 TI - A chicken consultation with ramifications. AB - In Madison I once worked with two postdoctoral fellows who had spent their youth in New York City and who, when asked what birds they knew both responded "why, pigeons and LBJ's!" (little brown jobbies). Despite their undoubted brilliance, they clearly had an educational deficiency not fixed by buying eggs and poultry at a grocery store. Though of enormous economic and nutritional importance to humans, turkeys and chickens constitute only a minute fraction of the disappearing avian life in our ecology. One could easily teach an entire middle or high school biology course around the reproduction, embryology, evolution, genetics, anatomy, special adaptations, virology, bacteriology, taxonomy, behavior, and extinctions of birds, as paradigmatic of all of life. Where would developmental or evolutionary biology be without the Galapagos finches, chick embryo, or neurobiology without the Zebra Finch? The modifications of the original red jungle fowl of India and South East Asia into hundreds of races through artificial selection and breeding practices provide as beautiful an example of developmental plasticity, well-known to Darwin, as the domestic dog, cat, laboratory mice, and guinea pigs. In what follows I have begun to repay my indebtedness to my mentor Emil Witschi who introduced me to developmental biology, physiology, and genetics and its historical study on the basis of birds (and amphibians); and to Mark Leppert, Professor of Human Genetics at the University of Utah for collaborative support, and bird-watching fieldtrips. PMID- 15666311 TI - Supernumerary ring chromosome 20 in a mother and her child. AB - The authors report a familial case of mosaicism for an extra ring 20, identified by fluorescence in situ hybridization (FISH), in a mother and her child. In spite of the fact that both patients had clinical abnormalities, the more severe phenotype present in the child was probably due to the higher percentage of abnormal cells. To our knowledge, this is the first report of a familial extra ring 20 mosaicism. PMID- 15666313 TI - A dominantly inherited spondylometaphyseal dysplasia with "corner fractures" and congenital scoliosis. AB - Spondylometaphyseal dysplasia (SMD) is a term applied to a varied group of skeletal dysplasias that principally involve the spine and the metaphyses of long bones. SMD Sutcliffe or "Corner Fracture" type is characterized by short stature, developmental coxa vara, fragmented appearance of the metaphyses ("corner fractures"), abnormally shaped vertebrae, odontoid hypoplasia, and dominant inheritance. We report a family with a dominantly inherited SMD with "corner fractures" and severe, congenital scoliosis but neither coxa vara nor odontoid abnormalities. This could either represent phenotypic variability in SMD-"Corner Fracture" type, or be a new, dominantly inherited SMD. The presence of severe, congenital scoliosis and short stature is present in all members of this family, and not typically seen in SMD-"Corner Fracture" type, supporting our hypothesis that this might represent a new, dominantly inherited SMD. PMID- 15666312 TI - Mosaic monosomy of a neocentric ring chromosome maps brachyphalangy and growth hormone deficiency to 13q31.1-13q32.3. AB - We describe a boy with moderate intellectual disability associated with distinctive hand malformations (hypoplastic and angel-shaped middle phalanges) and partial growth hormone (GH) deficiency associated with mosaic deletion of 13q31.1-13q32.3. The deleted segment was mapped to a 20-Mb region bounded by BACs RP11-1143C2 and RP11-139C1, narrowing the previously described locus for hand malformations at this region and suggesting that a locus for GH deficiency is also present at this location. The deleted segment contains at least three candidate genes, glypican-5, FARP1 and SOX21, that may be contributing to the phenotype in this boy. In a significant proportion (approximately 50%) of cells, the deleted region is present as a supernumerary ring chromosome stabilized by the formation of a neocentromere at 13q31-q32, within a region with a known propensity for neocentromere formation. The ring chromosome appears to be prone to low-level misdivision and loss in vitro which, in vivo, must be countered by selection for the balanced karyotype because the level of mosaicism has remained stable over 13 years. PMID- 15666314 TI - Lateral meningocele syndrome: vertical transmission and expansion of the phenotype. AB - Lateral meningoceles were first described by Lehman et al. [(1977); J Pediatr 90: 49-54] in a patient with other skeletal findings and distinctive craniofacial features. Subsequently, six more patients with the so-called lateral meningocele syndrome (LMS) have been reported. We describe the findings in three new cases and expand the phenotype. The existence of an affected mother and daughter supports the hypothesis that LMS is a dominant disorder affecting primarily the connective tissue. PMID- 15666315 TI - Comparison of primary monoatomic with primary polyatomic ions for the characterisation of polyesters with static secondary ion mass spectrometry. AB - Static secondary ion mass spectrometry (S-SIMS) emerges as one of the most adequate methods for the surface characterisation of polymers with an information depth of essentially one monolayer. The continuing search for increased analytical sensitivity and specificity has led to exploring the use of polyatomic primary ions as an alternative to the traditionally applied monoatomic projectiles. As part of a systematic investigation on polyatomic bombardment of organic and inorganic solids, this paper focuses on selected polyesters. Mass spectra and ion yields are compared for layers deposited on silicon wafers by spincoating solutions with different concentrations of poly(epsilon-caprolactone) (PCL), poly(butylene adipate) (PBA) and poly(ethylene adipate) (PEA). Accurate mass measurements have been used to support the assignment of the ions and link the composition of the detected ions to the analyte structure. Use of polyatomic projectiles increases the yield of structural ions with a factor of +/-15, +/-30 and +/-10 for PCL, PBA and PEA, respectively, in comparison to bombardment with Ga+ primary ions, while the molecular specificity is improved by the detection of additional high m/z ions. PMID- 15666316 TI - Investigation of the metabolic fate of 2-, 3- and 4-bromobenzoic acids in bile duct-cannulated rats by inductively coupled plasma mass spectrometry and high performance liquid chromatography/inductively coupled plasma mass spectrometry/electrospray mass spectrometry. AB - Inductively coupled plasma mass spectrometry (ICPMS) has been used to determine the rate and routes of excretion of bromine following the intraperitoneal administration (50 mg kg(-1)) of 2-, 3- and 4-bromobenzoic acids to male bile duct-cannulated rats. Analysis of urine and bile for (79/81)Br using ICPMS showed that all three bromobenzoic acids were rapidly excreted (82-98%) within 48 h of dosing, primarily via the urine. High-performance liquid chromatography/inductively coupled plasma mass spectrometry (HPLC/ICPMS) was then used to obtain metabolite profiles for bile and urine. These profiles revealed that extensive metabolism had taken place, with the unchanged bromobenzoic acids forming a minor part of the total of compound-related material detected. Concomitant MS studies, supplemented by alkaline hydrolysis, enabled the identification of the major metabolite of all three of the bromobenzoic acids as a glycine conjugate. Ester glucuronide conjugates were also identified, but formed only a small proportion of total. PMID- 15666317 TI - A selective unsaturated hydrocarbon subtraction technique for stable carbon isotopic analysis of atmospheric methyl chloride, methyl bromide, and C2-C5 saturated hydrocarbons using continuous-flow isotope ratio mass spectrometry. AB - Using continuous-flow isotope ratio mass spectrometry, we have developed a new analytical system which enables us to determine the stable carbon isotopic composition of CH3Cl, CH3Br, and C2-C5 saturated hydrocarbons in gas samples even if they contain substantial amounts of unsaturated hydrocarbons, using an I2O5 reagent for their selective subtraction. The analytical precision of the delta13C determinations is better than 0.5 per thousand for >300 pmolC injections and better than 5 per thousand for 20 pmolC injections. Using the system, delta13C values for CH3Cl and CH3Br were found in burning exhaust that contain a substantial quantity of unsaturated hydrocarbons. CH3Cl and CH3Br measured in exhaust from burning rice plants exhibit highly 13C-depleted values of -56.6 +/- 1.3 per thousand and -48.6 +/- 3.9 per thousand, respectively, while saturated hydrocarbons exhibit delta13C values (-26.4 to -28.9 per thousand) that are comparable with the total delta13C value of the parent material (rice plant; 28.0 per thousand). Using the system, we can determine the delta13C values of methyl halides and hydrocarbons in many kinds of gas samples. PMID- 15666318 TI - Accurate mass measurement in nano-electrospray ionization mass spectrometry by alternate switching of high voltage between sample and reference sprayers. AB - An electrospray dual sprayer, which generates separate sample and reference sprays by alternately switching the high voltage between the two sprayers, is described. The technique permits accurate mass measurements in nano-electrospray ionization mass spectrometry (ESI-MS) to be obtained using a quadrupole/orthogonal acceleration time-of-flight mass spectrometer (Q-TOF). Similar to the method employed with a dual ESI source (Wolff JC et al., Anal. Chem. 2001; 73: 2605), the two sprays are orthogonal with respect to each other, but can be independently sampled without any baffle between these sprays. The reference sprayer is used in the original configuration of the ESI source and was optimized for a 1-2 muL/min flow, whereas the sample sprayer can be either a conventional glass capillary or a borosilicate tip of the type used for nano-ESI. Both sprayers can be positioned close to the cone so as to give maximum ion currents. The sample and reference sprays are independently generated by raising the potentials on the sample and reference sprayers to 1.4 and 3.0 kV, respectively; the high voltages can be rapidly turned on and off in ca. 1 ms. A nano-ESI-MS or nano-flow LC/ESI-MS experiment using a Q-TOF coupled with the above system gave mass accuracies within 3 ppm for measurements of ions up to m/z 1000 using subpicomole samples. PMID- 15666319 TI - Determination of chemical agent optical clearing potential using in vitro human skin. AB - BACKGROUND AND OBJECTIVES: Optical clearing is a method that temporarily reduces the optical scattering of biological tissues. STUDY DESIGN/MATERIALS AND METHODS: To determine the optical clearing potential (OCP) of various chemical agents, we examined the change in reduced scattering coefficients of in vitro human skin after application directly to the dermal side. RESULTS/CONCLUSIONS: We found that the mean agent OCP did not correlate with refractive index or osmolality. Lipophilic hyperosmotic agents had a mean OCP less than unity, indicating increased optical scattering. PMID- 15666320 TI - Differentiation of organ availability by sequential and simultaneous analyses: intestinal conjugative metabolism impacts on intestinal availability in humans. AB - The impact of intestinal conjugative metabolism on oral bioavailability was assessed by sequential and simultaneous analyses of the reported data in humans. The data were retrieved from reports on drugs that are metabolized by sulfate conjugation, and the organ availabilities affecting oral bioavailability were differentiated. Sequential analysis gave the following results. The intestinal availability (Fg) of salbutamol was 0.700, whereas hepatic availability (Fh) and bioavailability (F) were 0.893 and 0.493, respectively. Fg of (+)-terbutaline, ( )-terbutaline, and (+/-)-terbutaline was 0.128, 0.254, and 0.250, respectively. In contrast, Fh of (+)-terbutaline, (-)-terbutaline, and (+/-)-terbutaline was 0.979, 0.971, and 0.946, respectively. Fg and Fh of ethynylestradiol were 0.536 and 0.780, respectively. Simultaneous analysis also gave similar results, although the sequential analysis overestimated the intestinal availability. These results indicate that intestinal sulfation metabolism has more impact on intestinal availability than on hepatic availability, resulting in low bioavailability in humans. PMID- 15666322 TI - Loss of E-cadherin and p27 expression is associated with head and neck squamous tumorigenesis. AB - BACKGROUND: In neoplastic head and neck lesions, it has been found that the loss or reduction in E-cadherin expression is a late event and is associated with invasion. Low p27 levels have been associated with a poor prognosis in many different tumors, including laryngeal carcinoma. The authors investigated p27 and E-cadherin protein expression in the early stages of head and neck tumorigenesis and evaluated their predictive roles individually and in association with carcinogenesis. METHODS: Tissue biopsies from 46 patients who were participants in 3 chemoprevention trials were analyzed for E-cadherin expression, and 40 samples were analyzed for p27 expression using immunohistochemistry. RESULTS: The data suggested that loss of both E-cadherin expression and p27 expression occurred early during the preneoplastic steps of head and neck carcinogenesis, and loss of p27 protein expression alone (P=0.02) and in combination with loss of E-cadherin expression (P=0.04) was a significant predictor of the risk for head and neck carcinoma. CONCLUSIONS: The loss of p27 expression may be useful in the construction of a risk model for head and neck carcinogenesis and may represent a potential target for chemopreventive interventions. Longer follow-up of the high percentage of low-risk preneoplastic lesions in the current study and validation in a larger sample size may be required to establish the predictive role of these abnormalities. PMID- 15666321 TI - Prospective evaluation of major vascular events in patients with nonsmall cell lung carcinoma treated with cisplatin and gemcitabine. AB - BACKGROUND: Cancer and cisplatin-based chemotherapy both are well recognized risk factors for coagulation disorders and thrombosis. However, vascular events (VEs) seldom are considered adverse effects of treatment and may not even be taken into account in reports of chemotherapy trials. METHODS: VEs were recorded prospectively in a population of patients with nonsmall-cell lung carcinoma (NSCLC) who were treated consecutively with cisplatin and gemcitabine using a diagnostic flow chart based on a thorough clinical examination, hematologic and coagulative parameters, and imaging assessments when appropriate. RESULTS: From January, 2000 to January 2003, 108 patients with Stage III-IV NSCLC underwent chemotherapy and were evaluated. Overall, 22 VEs occurred in 19 patients (17.6%; 95% confidence interval [95% CI], 10.3-24.8%), including 10 arterial VEs (2 myocardial infarctions, 7 lower limb arterial thrombosis, and 1 ischemic stroke) and 12 venous VEs (3 catheter-related upper limb VEs, 6 venous thrombosis of the lower limb, and 3 pulmonary embolisms). The cumulative proportion of VEs at 1 year after the start of chemotherapy was 22.0% (95% CI, 12.7-31.3%). Four patients died due to the VE (overall mortality, 3.7%), and 3 patients needed surgical revascularization. In the other patients, conservative medical treatment was effective. Baseline patient-related and disease-related characteristics of the patients with VEs did not differ significantly from the characteristics of patients without VE; liver and brain metastases were more frequent in patients with VE, although the difference did not reach statistical significance. Response rates were similar in the two groups. A double VE was detected in three patients who were given further chemotherapy after resolution of the first event. CONCLUSIONS: VEs were a common finding in chemotherapy-treated NSCLC patients. Chemotherapy itself seem to be a powerful risk factor for VE. Strategies to predict the occurrence of VEs should be developed to spare this life-threatening toxicity. PMID- 15666323 TI - Combined chemotherapy with carmustine, doxorubicin, etoposide, vincristine, and cyclophosphamide plus mitoxantrone, cytarabine and methotrexate with citrovorum factor for the treatment of aggressive non-Hodgkin lymphoma: a long-term follow up study. AB - BACKGROUND: The standard treatment for patients with aggressive non-Hodgkin lymphoma (NHL) is cyclophosphamide, doxorubicin, vincristine, and prednisone (CHOP). Since 1989, the authors have used a new chemotherapy regimen with combined carmustine, doxorubicin, etoposide, vincristine, and cyclophosphamide plus mitoxantrone, cytarabine and methotrexate with citrovorum factor called BAVEC-MiMA. The objective of the current study was to explore, after a long follow-up period, the impact of this third-generation regimen for the treatment of aggressive NHL. METHODS: One hundred and one consecutive patients (median age, 41 years) with either B-cell (n=94 patients) or non-B-cell (n=7 patients), aggressive lymphoma were diagnosed and treated between 1989 and 1999 with the BAVEC-MiMA regimen. RESULTS: The complete response rate was 74%, and the overall response rate was 89%. Eleven patients with refractory disease died rapidly after a median period of 5 months. The major toxicity was Grade 4 neutropenia (according to World Health Organization criteria), which was observed in 15 patients (15%). There were four toxicity-related deaths. The overall survival rate was 63% at 9 years. In multivariate analysis, factors that were associated with advantage in overall survival were response to induction therapy, bulky disease, and high score on the International Prognostic Index (IPI). The disease free survival rate was 77% at 9 years. In multivariate analysis, the IPI was the most important variable for the definition of disease-free survival. CONCLUSIONS: The BAVEC-MiMA regimen was feasible on an outpatient basis, it was tolerated well, and it showed a low toxicity-related mortality. The long follow-up in patients with NHL, which is a rapidly fatal disease, led the authors to observe that, with this regimen, a cure was obtained in > 50% of patients who had low risk or low-to-intermediate-risk, aggressive NHL. PMID- 15666324 TI - Determinants of quality of life in patients with cancer. AB - BACKGROUND: Because health-related quality of life (QOL) is an important outcome in cancer management, the authors sought to better understand its determinants. To address this subject, they analyzed QOL, as measured with the Functional Assessment of Cancer Therapy-General questionnaire (FACT-G), Spanish Version 4, and depicted the complex relations among physical, psychological, social, and cultural factors, including spirituality. METHODS: A cross-sectional study design was used with a sample of 309 patients with cancer. The influence of several possible determinants was first studied by univariate regression analysis. Variables showing an association were included in a forward stepwise multivariate regression model. RESULTS: Five regression models were studied, for the FACT-G total score and its four subscales. Five variables explained 32.1% of the variance of the FACT-G total score: tumor stage, spiritual well-being, income, mood disorders, and mode of questionnaire administration. The type and relevance of the explanatory variables differed among the various dimensions of QOL. CONCLUSIONS: The authors underlined the entwining of biologic, psychosocial, and spiritual factors as determinants of the QOL of patients with cancer, thus supporting the multidimensional definition and modeling of the construct. PMID- 15666325 TI - The persistence of isolated tumor cells in bone marrow from patients with breast carcinoma predicts an increased risk for recurrence. AB - BACKGROUND: The prognostic significance of isolated tumor cells (ITCs) in bone marrow (BM) from patients with breast carcinoma at the time of their primary diagnosis recently was been confirmed by a large pooled analysis. If the persistence of ITCs after adjuvant therapy confers a similar risk for recurrence, then it would be an indication to consider secondary adjuvant therapy. METHODS: The authors analyzed BM aspirates from 228 patients during recurrence-free follow up at a median interval +/- standard deviation (SD) of 21.3 +/- 29.1 months after a primary diagnosis of breast carcinoma (pathologic T1 [pT1]-pT2, pN0-pN3, pM0). Carcinoma cells were detected using a standardized immunoassay with monoclonal antibody A45-B/B3 directed against cytokeratin (CK). Patients were followed for a median +/- SD of 49.8 +/- 32.1 months after their primary diagnosis. RESULTS: Persistent ITCs in BM were detected in 12.7% of patients (n=29 patients). Positive BM status was more frequent (15.7%) within the first 21 months after primary diagnosis than after a follow-up > 21 months (9.7%). The Kaplan-Meier estimate for mean recurrence-free survival was 149.7 months (95% confidence interval [95% CI], 139.6-159.8 months) in patients with negative BM status and 86.5 months (95% CI, 65.7-107.4 months; P=0.0003) in patients with positive BM status at the time patients underwent follow-up BM aspiration. Patients who were without evidence of persistent ITCs had a significantly longer overall survival (162.1 months; 95% CI, 152.1-172.0 months) compared with patients who had positive BM status (overall survival, 98.7 months; 95% CI, 79.7-117.9 months; P=0.0008). In multivariate Cox regression analysis that included BM status, tumor size, lymph node status, and histopathologic grade, evidence of ITCs was an independent significant predictor for reduced disease-free survival (relative risk [RR], 4.57; P <0.0001) and overall survival (RR, 5.57; P=0.002). Persistent ITCs had the greatest prognostic relevance when they were detected between 25 months and 42 months after primary diagnosis (RR, 7.68). CONCLUSIONS: Evidence of persistent ITCs in BM from patients with breast carcinoma indicated an increased risk for subsequent recurrence. Prospective trials should investigate the benefit of secondary adjuvant treatment on the basis of BM marrow status. PMID- 15666326 TI - Fiberoptic ductoscopy findings in women with and without spontaneous nipple discharge. AB - BACKGROUND: Fiberoptic ductoscopy (FD), which allows direct visualization of the breast ductal lumen, is performed in women with and without spontaneous nipple discharge (SND). Previous reports suggested that cytologic evaluation of SND may be falsely interpreted as containing malignancy. The purpose of the current study, which was performed prospectively, was to determine whether ductoscopic findings were different in women with versus without SND, and to assess the implications of the differences in SND versus non-SND samples regarding the role of FD in assessing whether a woman has breast carcinoma. METHODS: Data were collected on the distance traveled by the ductoscope, visual observations, pathology, cytology, epithelial and foam cell quantity, and image analysis for ploidy, hypertetraploidy, and S-phase fraction. RESULTS: Of 100 FD specimens, 60 were from breasts without SND and 40 were from breasts with SND. Intraductal visual observations (P < or = 0.0002), pathologic findings in the resected specimen (P < or = 0.001), and quantity of epithelial cells (P=0.03) were influenced by the presence or absence of SND. Although one specimen from a benign breast was interpreted as cytologically malignant, every breast with both malignant cytology and aneuploidy contained cancer cells. A model incorporating cytology and SND was 92% sensitive and 60% specific in predicting which women had breast carcinoma. CONCLUSIONS: There were pronounced differences in FD samples from women with and without SND. FD biologic parameters can be chosen to optimize breast carcinoma predictive sensitivity and specificity. SND cytology can present a diagnostic problem, suggesting the need for histologic confirmation before the initiation of therapy. PMID- 15666327 TI - Is older age associated with a worse prognosis due to different patterns of care? A long-term study of 1346 patients with glioblastomas or brain metastases. AB - BACKGROUND: The objective of this study was to find out whether the worse prognosis of older patients with primary or metastatic brain tumors can be explained by different patterns of care compared with younger patients. METHODS: A data base that included 430 patients with glioblastomas and 916 patients with brain metastases who underwent radiotherapy at the author's hospital between 1980 and 2000 was analyzed. Patterns of care were compared for different age groups using the chi-square test. RESULTS: In both patient groups, age turned out to be an independent risk factor. Older age was associated with worse overall survival. Independent of the cut-off age (< 50 years vs. > or = 50 years, < 60 years vs. > or = 60 years, < 65 years vs. > or = 65 years, and < 70 years vs. > or = 70 years), there were no statistically significant differences between the age groups concerning the use of different imaging modalities (computed tomography scans vs. magnetic resonance imaging), type of surgery (none vs. biopsy vs. resection), waiting time for radiotherapy (< median vs. > or = median), radiotherapy treatment planning (simulator-based vs. computer-based), use of radiation sources (cobalt unit vs. linear accelerator), and fractionation protocols (conventional vs. modified). When the recruitment period of 21 years was divided into 3 intervals, impressive changes with regard to the patterns of care became apparent. However, the changes were seen similarly throughout the different age groups. CONCLUSIONS: Older age did not limit access to state-of-the art patterns of care in neurooncology. Patients participated in medical progress irrespective of their age. The worse prognosis of older patients with glioblastomas or brain metastases was not determined by age-related differences in access to health care. PMID- 15666328 TI - Perspectives for the management of febrile neutropenic patients with cancer in the 21st century. AB - Over the past several decades, there has been substantial progress in the management of patients with febrile neutropenia. However, the ever-changing patterns of infection, ecology, and antibiotic-resistance trends do not allow the development of treatment guidelines that could be applied universally. Hence, the institution's predominant pathogens and resistance patterns should guide the empirical choice of antimicrobials. Prompt initiation of antimicrobial therapy remains the gold standard. Monotherapy with the newer broad-spectrum antimicrobials has tended to replace the classic combination therapy. Empirical administration of glycopeptides, such as vancomycin, without documentation of a gram-positive infection is not favored. The development of risk-stratification models has allowed for identification of low-risk patients with additional treatment options, such as early discharge and exclusively outpatient treatment with oral antimicrobials. The initiation of empirical antifungal therapy in persistently febrile neutropenic patients has become common practice, especially recently, since the introduction of new, effective, less toxic antifungal drugs. It is hoped that the development of new nonculture-based diagnostic methods will allow for the early detection of invasive fungal infections and, thus, the replacement of empirical antifungal therapy with pathogen-specific, preemptive therapy. PMID- 15666329 TI - Receptor fragment approach to the binding between CCK8 peptide and cholecystokinin receptors: a fluorescence study on type B receptor fragment CCK(B)-R (352-379). AB - Fluorescence titrations in a membrane mimetic solvent system allowed us to estimate that the dissociation constant of the bimolecular complex between CCK8 peptide and cholecystokinin type B receptor fragment CCK(B)-R (352-379) is in the micromolar range. When considered in the context of the full receptor/ligand model, these experiments demonstrate that the receptor fragment chosen on the basis of previous structural studies represents a reliable model system to monitor the ability of CCK8 or CCK8 analogs to bind the cholecystokinin receptor. Together with previous studies, this confirms that the receptor fragment approach adopted to define the binding mode of the CCK8 fragment of cholecystokinin with its two receptors, CCK(A) and CCK(B,) can be used to characterize the binding from the equilibrium standpoint. In this context, fluorescence spectroscopy proves to be the favored technique to measure dissociation constants in the nanomolar to micromolar range. PMID- 15666330 TI - Crystal structure of cyclic (APGVGV)2, an analog of elastin, and a suggested mechanism for elongation/contraction of the molecule. AB - Tropoelastin is a complex polymeric protein composed primarily of repeating segments of Val-Pro-Gly-Gly, Val-Pro-Gly-Val-Gly, and Ala-Pro-Gly-Val-Gly-Val that occurs in connective tissue and arteries. It has rubber-like extensible properties. A synthetic cyclic dodecapeptide, with a double repeat of the hexapeptide sequence, has been shown to undergo a reversible inverse temperature transition; that is, crystals grow at 60 degrees C and dissolve in the mother liquor upon cooling. An x-ray crystal structure analysis established that the cyclic backbone formed an elongated loop with a Pro-Gly, type II beta turn at both ends. Six internal cross strand NH...OC hydrogen bonds form between six NH donors and four O=C acceptors where two of the carbonyl O atoms are bifurcated acceptors. As a result, the molecule is pulled up into a corrugated profile. The corrugated loops form extended beta-sheets by additional intermolecular hydrogen bonds. An analysis of the dome region in a corrugated sheet suggests a reversible mechanism for extending and contracting the length of the whole molecule, akin to the motion of opening and closing an umbrella, caused by the motion of a water molecule with its associated hydrogen bonds acting as spokes. Crystal parameters: C44H72N12O12.3H2O, sp. gr. P2(1)2(1)2(1), a = 9.212 angstroms, b = 19.055 angstroms, c = 32.247 angstroms, d = 1.157 g/cm3. PMID- 15666331 TI - Thermal stability and energetics of 15-mer DNA duplex interstrand crosslinked by trans-diamminedichloroplatinum(II). AB - The effect of the location of the interstrand cross-link formed by trans diamminedichloroplatinum(II) (transplatin) on the thermal stability and energetics of 15-mer DNA duplex has been investigated. The duplex containing single, site-specific cross-link, thermodynamically equivalent model structures (hairpins) and nonmodified duplexes were characterized by differential scanning calorimetry, temperature-dependent uv absorption, and circular dichroism. The results demonstrate that the formation of the interstrand cross-link of transplatin does not affect pronouncedly thermodynamic stability of DNA: the cross-link induces no marked changes not only in enthalpy, but also in "reduced" (concentration independent) monomolecular transition entropy. These results are consistent with the previous observations that interstrand cross-links of transplatin structurally perturb DNA only to a relatively small extent. On the other hand, constraining the duplex with the interstrand cross-link of transplatin results in a significant increase in thermal stability that is primarily due to entropic effects: the cross-link reduces the molecularity of the oligomer system from bimolecular to monomolecular. Importantly, the position of the interstrand cross-link within the duplex modulates cooperativity of the melting transition of the duplex and consequently its thermal stability. PMID- 15666332 TI - Weight gain associated with the -759C/T polymorphism of the 5HT2C receptor and olanzapine. AB - BACKGROUND: Weight gain from atypical antipsychotic use has become a significant problem. Recent reports have liked the -759 polymorphism of the 5HT2C receptor and obesity as well as weight gain from chlorpromazine, risperidone, and clozapine. AIM: To determine associations between weight gain during olanzapine treatment and the -759C/T polymorphism of the 5HT2C receptor gene. METHODS: This study included 42 acutely ill patients with schizophrenia (DSM-IV). Weekly assessments included Brief Psychiatric Rating Scale (BPRS), Scale for Assessment of Negative Symptoms (SANS), and weight measurements. Olanzapine was titrated to a fixed dose (7.5-20 mg/day) for 2-6 weeks. A 24 hr plasma level was obtained at the endpoint visit. Genomic DNA was isolated from a whole blood sample and analyzed for the -759C/T polymorphism of the 5HT2C receptor. RESULTS: A chi square analysis was conducted comparing the distribution of T and C alleles in subjects grouped as gaining more or less than 5, 7, and 10% of their baseline weight during treatment with olanzapine. A threshold of 10% was found to be significant. The distribution of T alleles was higher in subjects not gaining 10% of more of their body weight compared who did gain significant weight (11/27 (40.7%) vs. 0/15 (100%), chi2 = 11.805, P = 0.0035). CONCLUSIONS: Subjects with a T allele of the 5HT2C receptor -759C/T polymorphism may have a lower incidence of weight gain from olanzapine over a 6 week period compared to those with the C allele. These results need to be replicated. PMID- 15666333 TI - Autism and 15q11-q13 disorders: behavioral, genetic, and pathophysiological issues. AB - New insights into biological factors that underlie autism may be gained by comparing autism to other neurodevelopmental disorders that have autistic features and relatively well-delineated genetic etiologies or neurobiological findings. This review moves beyond global diagnoses of autism and instead uses an endophenotypic approach to compare specific clusters of autistic symptomatology to features of chromosome 15q11-q13 disorders. Paternally or maternally derived deficiencies of 15q11-q13 result in Prader-Willi or Angelman syndromes, and we first use a global approach to review potential autism susceptibility genes in the 15q11-q13 region. We then use a more trait-based approach to suggest possible ties between specific phenotypic characteristics of autism and Prader-Willi syndrome, namely savant-like skills. We conclude with insights from pathophysiological studies that implicate altered development of specific neuron types and circuits in the cerebral cortex as part of the pathophysiological processes associated with autism and mental retardation. PMID- 15666334 TI - Toward a developmental neurobiology of autism. AB - Autism is a complex, behaviorally defined, developmental brain disorder with an estimated prevalence of 1 in 1,000. It is now clear that autism is not a disease, but a syndrome with a strong genetic component. The etiology of autism is poorly defined both at the cellular and the molecular levels. Based on the fact that seizure activity is frequently associated with autism and that abnormal evoked potentials have been observed in autistic individuals in response to tasks that require attention, several investigators have recently proposed that autism might be caused by an imbalance between excitation and inhibition in key neural systems including the cortex. Despite considerable ongoing effort toward the identification of chromosome regions affected in autism and the characterization of many potential gene candidates, only a few genes have been reproducibly shown to display specific mutations that segregate with autism, likely because of the complex polygenic nature of this syndrome. Among those, several candidate genes have been shown to control the early patterning and/or the late synaptic maturation of specific neuronal subpopulations controlling the balance between excitation and inhibition in the developing cortex and cerebellum. In the present article, we review our current understanding of the developmental mechanisms patterning the balance between excitation and inhibition in the context of the neurobiology of autism. PMID- 15666335 TI - Designing mouse behavioral tasks relevant to autistic-like behaviors. AB - The importance of genetic factors in autism has prompted the development of mutant mouse models to advance our understanding of biological mechanisms underlying autistic behaviors. Mouse models of human neuropsychiatric diseases are designed to optimize (1) face validity, i.e., resemblance to the human symptoms; (2) construct validity, i.e., similarity to the underlying causes of the disease; and (3) predictive validity, i.e., expected responses to treatments that are effective in the human disease. There is a growing need for mouse behavioral tasks with all three types of validity for modeling the symptoms of autism. We are in the process of designing a set of tasks with face validity for the defining features of autism: deficits in appropriate reciprocal social interactions, deficits in verbal social communication, and high levels of ritualistic repetitive behaviors. Social approach is tested in an automated three chambered apparatus that offers the subject a choice between a familiar environment, a novel environment, and a novel environment containing a stranger mouse. Preference for social novelty is tested in the same apparatus, with a choice between the start chamber, the chamber containing a familiar mouse, and the chamber containing a stranger mouse. Social communication is evaluated by measuring the ultrasonic distress vocalizations emitted by infant mouse pups and the parental response of retrieving the pup to the nest. Resistance to change in ritualistic repetitive behaviors is modeled by forcing a change in habit, including reversal of the spatial location of a reinforcer in a T-maze task and in the Morris water maze. Mouse behavioral tasks that may model additional features of autism are discussed, including tasks relevant to anxiety, seizures, sleep disturbances, and sensory hypersensitivity. Applications of these tests include (1) behavioral phenotyping of transgenic and knockout mice with mutations in genes relevant to autism, (2) characterization of mutant mice derived from random chemical mutagenesis, (3) DNA microarray analyses of genes in inbred strains of mice that differ in social interaction, social communication and resistance to change in habit, and (4) evaluation of proposed therapeutics for the treatment of autism. PMID- 15666337 TI - Preface. PMID- 15666336 TI - Neuroanatomical substrates of social cognition dysfunction in autism. AB - In this review article, we summarize recent progress toward understanding the neural structures and circuitry underlying dysfunctional social cognition in autism. We review selected studies from the growing literature that has used the functional neuroimaging techniques of cognitive neuroscience to map out the neuroanatomical substrates of social cognition in autism. We also draw upon functional neuroimaging studies with neurologically normal individuals and individuals with brain lesions to highlight the insights these studies offer that may help elucidate the search for the neural basis of social cognition deficits in autism. We organize this review around key brain structures that have been implicated in the social cognition deficits in autism: (1) the amygdala, (2) the superior temporal sulcus region, and (3) the fusiform gyrus. We review some of what is known about the contribution of each structure to social cognition and then review autism studies that implicate that particular structure. We conclude with a discussion of several potential future directions in the cognitive neuroscience of social deficits in autism. PMID- 15666339 TI - Identifying environmental contributions to autism: provocative clues and false leads. AB - The potential role of environmental factors in autism spectrum disorders (ASD) is an area of emerging interest within the public and scientific communities. The high degree of heritability of ASD suggests that environmental influences are likely to operate through their interaction with genetic susceptibility during vulnerable periods of development. Evaluation of the plausibility of specific neurotoxicants as etiological agents in ASD should be guided by toxicological principles, including dose-effect dependency and pharmacokinetic parameters. Clinical and epidemiological investigations require the use of sufficiently powered study designs with appropriate control groups and unbiased case ascertainment and exposure assessment. Although much of the existing data that have been used to implicate environmental agents in ASD are limited by methodological shortcomings, a number of efforts are underway that will allow more rigorous evaluation of the role of environmental exposures in the etiology and/or phenotypic expression of the disorder. Surveillance systems are now in place that will provide reliable prevalence estimates going forward in time. Anticipated discoveries in genetics, brain pathology, and the molecular/cellular basis of functional impairment in ASD are likely to provide new opportunities to explore environmental aspects of this disorder. PMID- 15666338 TI - Early detection of core deficits in autism. AB - The goal of this review of the research literature is to discuss approaches to the early detection of autism in infancy. Early detection would enable diagnoses to be made before 18 months of age rather than at 24-30 months, the age where diagnoses start to be made now. After summarizing the criteria for a deficit to be considered "core" to the disorder, the literature on research strategies used in early detection is examined. In order to guide the design of future studies, the review then turns to an overview of what is known about the processes of early social development in typically developing children that underlie the domains in which core deficits are manifested in young children with autism. The social domains covered in the review are those that show development in typically developing infants below 18 months of age: dyadic interaction and imitation; emotion discrimination; and attachment. The review concludes that all of these areas are worthy of investigation in young children, particularly those at higher risk of showing some of the core deficits of autism such as the infant siblings of children with autism. PMID- 15666340 TI - Treating the core features of autism: are we there yet? AB - A wide variety of nonestablished treatments have been proposed as "cures" for the core features of autism and are used frequently despite having largely escaped scientific scrutiny. In contrast, a growing body of empirical evidence supports the use of a few forms of theory-based and empirically validated treatment for some aspects of the core features of autism. These include behavioral/psychoeducational interventions and specific forms of medication treatment, which can produce significant improvements in communication, social interaction, and problem behaviors that both maintain over time and generalize across settings. While there is no doubt that treatment and educational services for persons with autism have improved over the past 6 decades, it also appears that significant issues remain with respect to (1) the routine application of validated treatments for the majority of cases with autism, (2) the resistance to even validated forms of treatment for a substantial minority of cases with autism, and (3) the extent to which validated treatments effectively treat the specific core features of autism that are most disabling for persons with autism and their families. PMID- 15666341 TI - Trajectory of development in adolescents and adults with autism. AB - This article seeks to elucidate the trajectory of development in adolescents and adults with autism. Prospective, retrospective, and cross-sectional studies are reviewed to reveal the manifestation of and changes in the core symptoms of autism in adolescence and adulthood. Comparing children with adolescents and adults, modest degrees of symptom abatement and improvement in skills have been documented in multiple studies, as are increases in verbal and decreases in performance IQ. Nevertheless, most individuals do not attain normative outcomes in adulthood and continue to manifest significant degrees of symptomatology and dependency. However, a small sub-group (about 15%) has more favorable adult outcomes. PMID- 15666342 TI - The search for autism disease genes. AB - Autism is a heritable disorder characterized by phenotypic and genetic complexity. This review begins by surveying current linkage, gene association, and cytogenetic studies performed with the goal of identifying autism disease susceptibility variants. Though numerous linkages and associations have been identified, they tend to diminish upon closer examination or attempted replication. The review therefore explores challenges to current methodologies presented by the complexities of autism that might underlie some of the current difficulties, and finishes by describing emerging phenotypic, statistical, and molecular investigational approaches that offer hope of overcoming those challenges. PMID- 15666343 TI - More on sperm acrosomal exocytosis. PMID- 15666345 TI - Distributed robustness versus redundancy as causes of mutational robustness. AB - A biological system is robust to mutations if it continues to function after genetic changes in its parts. Such robustness is pervasive on different levels of biological organization, from macromolecules to genetic networks and whole organisms. I here ask which of two possible causes of such robustness are more important on a genome-wide scale, for systems whose parts are genes, such as metabolic and genetic networks. The first of the two causes is redundancy of a system's parts: A gene may be dispensable if the genome contains redundant, back up copies of the gene. The second cause, distributed robustness, is more poorly understood. It emerges from the distributed nature of many biological systems, where many (and different) parts contribute to system functions. I will here discuss evidence suggesting that distributed robustness is equally or more important for mutational robustness than gene redundancy. This evidence comes from the functional divergence of redundant genes, as well as from large-scale gene deletion studies. I also ask whether one can quantify the extent to which redundancy or distributed robustness contribute to mutational robustness. PMID- 15666346 TI - Rising starlet: the starlet sea anemone, Nematostella vectensis. AB - In recent years, a handful of model systems from the basal metazoan phylum Cnidaria have emerged to challenge long-held views on the evolution of animal complexity. The most-recent, and in many ways most-promising addition to this group is the starlet sea anemone, Nematostella vectensis. The remarkable amenability of this species to laboratory manipulation has already made it a productive system for exploring cnidarian development, and a proliferation of molecular and genomic tools, including the currently ongoing Nematostella genome project, further enhances the promise of this species. In addition, the facility with which Nematostella populations can be investigated within their natural ecological context suggests that this model may be profitably expanded to address important questions in molecular and evolutionary ecology. In this review, we explore the traits that make Nematostella exceptionally attractive as a model organism, summarize recent research demonstrating the utility of Nematostella in several different contexts, and highlight a number of developments likely to further increase that utility in the near future. PMID- 15666347 TI - Mechanisms of germ-cell specification in mouse embryos. AB - The mode and timing of germ-cell specification has been studied in diverse organisms, however, the molecular mechanism regulating germ-cell-fate determination remains to be elucidated. In some model organisms, maternal germ cell determinants play a key role. In mouse embryos, some germ-line-specific gene products exist as maternal molecules and play critical roles in a pluripotential cell population at preimplantation stages. From those cells, primordial germ cells (PGCs) are specified by extracellular signaling mediated by tissue, as well as cell-cell interaction during gastrulation. Thus, establishment of germ-cell lineage in mammalian embryos appears to be regulated by a multistep process, including formation and maintenance of a pluripotential cell population, as well as specification of PGCs. PGCs can be generated from pluripotential embryonic stem (ES) cells in a simple monolayer culture in which tissue interaction does not occur. This raises the possibility that ES cells, as well as, possibly, pluripotential cells in preimplantation embryos, are more closely related to the PGC precursors than pluripotential cells after implantation. PMID- 15666348 TI - Do protein motifs read the histone code? AB - The existence of different patterns of chemical modifications (acetylation, methylation, phosphorylation, ubiquitination and ADP-ribosylation) of the histone tails led, some years ago, to the histone code hypothesis. According to this hypothesis, these modifications would provide binding sites for proteins that can change the chromatin state to either active or repressed. Interestingly, some protein domains present in histone-modifying enzymes are known to interact with these covalent marks in the histone tails. This was first shown for the bromodomain, which was found to interact selectively with acetylated lysines at the histone tails. More recently, it has been described that the chromodomain can be targeted to methylation marks in histone N-terminal domains. Finally, the interaction between the SANT domain and histones is also well documented. Overall, experimental evidence suggests that these domains could be involved in the recruitment of histone-modifying enzymes to discrete chromosomal locations, and/or in the regulation their enzymatic activity. Within this context, we review the distribution of bromodomains, chromodomains and SANT domains among chromatin modifying enzymes and discuss how they can contribute to the translation of the histone code. PMID- 15666349 TI - Turning it up a Notch: cross-talk between TGF beta and Notch signaling. AB - Signaling through both the transforming growth factor beta (TGF beta) superfamily of growth factors and Notch play crucial roles during embryonic pattern formation and cell fate determination. Although both pathways are able to exert similar biological responses in certain cell types, a functional interaction between these two signaling pathways has not been described. Now, three papers provide evidence of both synergy and antagonism between TGF beta and Notch signaling. These reports describe a requirement for Notch signal transducers in TGF beta- and BMP-induced expression of Notch target genes, as well as in BMP-controlled cell differentiation and migration. These papers uncover a direct link between the Notch and TGF beta pathways and suggest a critical role for Notch in some of the biological responses to TGF beta family signaling. PMID- 15666350 TI - Retrotransposons and regulatory suites. AB - Cellular differentiation and multicellular development require the programmed expression of coregulated suites of genetic loci dispersed throughout the genome. How do functionally diverse loci come to share common regulatory motifs? A new paper finds that retrotransposons (RTEs) may play a role in providing common regulation to a group of functions expressed during the development of oocytes and preimplantation embryos. Examining cDNA libraries, Peaston et al. find that 13% of all processed transcripts in full-grown mouse oocytes contain RTE sequences, mostly from the MT family of retroviral-like elements. Smaller but still significant percentages of RTE sequences are found in cDNA libraries from 2 cell embryos and blastocysts. A quarter of these RTE sequences are at the 5' ends of chimeric transcripts that also contain exons from endogenous mouse loci. These chimeric transcripts display restricted expression in oocytes and preimplantation embryos and presumably originate from developmentally regulated LTR promoters. Some, but not all, chimeric transcripts encode novel protein products. PMID- 15666351 TI - Interview with Mike Berridge. PMID- 15666352 TI - Petromyzonol sulfate and its derivatives: the chemoattractants of the sea lamprey. AB - Petromyzonol sulfate (PZS) and 3 keto-PZS are bile alocohol derivatives that serve as chemoattractants during the life cycle of sea lamprey (Petromyzon marinus). The sulfonate moiety is crucial perhaps conferring the required solubility for the pheromone that is released into the streams and for the specificity to bind to its receptor. During the life cycle of lamprey, larvae produce copious amounts of 5 alpha-cholan-PZS, and trace amounts of allocholic acid (ACA), which attracts adults to the same breeding ground. Later the spermeating males produce 3keto-PZS, and trace amounts of 3-keto-ACA, which attracts the ovulating females, signaling both its reproductive status and its nesting location for successful reproduction. In both stages, a mixture of components serves as pheromone plume, similar to insects. The receptors for the migratory and the reproductive pheromones need to be molecularly cloned and characterized in order to understand the molecular biology of olfaction in the sea lamprey. PMID- 15666353 TI - Leukocytes on the move with phosphoinositide 3-kinase and its downstream effectors. AB - Cell signalling mediators derived from membrane phospholipids are frequent participants in biological processes. The family of phosphoinositide 3-kinases (PI3Ks) phosphorylate the membrane lipid phosphatidylinositol, generating second messengers that direct diverse responses. These PI3K products are fundamental for leukocyte migration or chemotaxis, a pivotal event during the immune response. This system is therefore of significant biomedical interest. This review focuses on the biochemistry and signalling pathways of PI3K, with particular emphasis on chemokine (chemotactic cytokine)-directed responses. The key objectives of chemotaxis are motility and direction. The latter--direction--requires distinct events at the front and back of a cell. In light of this, the coordinated localisation of signalling factors, an event choreographed by a sharp intracellular gradient of PI3K-derived products, is a common theme. PMID- 15666354 TI - Are there non-catalytic functions of acetylcholinesterases? Lessons from mutant animal models. AB - Acetylcholinesterase (AChE) hydrolyses acetylcholine (ACh) ensuring the fast clearance of released neurotransmitter at cholinergic synapses. Many studies led to the hypothesis that AChE and the closely related enzyme butyrylcholinesterase (BChE) may play other, non-hydrolytic roles during development. In this review, we compare data from in vivo studies performed on invertebrate and vertebrate genetic models. The loss of function of ache in these systems is responsible for the appearance of several phenotypes. In all aspects so far studied, the phenotypes can be explained by an excess of the undegraded substrate, ACh, leading to misfunction and pathological alterations. Thus, the lack of AChE catalytic activity in the mutants appears to be solely responsible for the observed phenotypes. None of them appears to require the postulated adhesive or other non-hydrolytic functions of AChE. PMID- 15666355 TI - PAR proteins and the establishment of cell polarity during C. elegans development. AB - Cells become polarized to develop functional specializations and to distribute developmental determinants unequally during division. Studies that began in the nematode C. elegans have identified a group of largely conserved proteins, called PAR proteins, that play key roles in the polarization of many different cell types. During initial stages of cell polarization, certain PAR proteins become distributed asymmetrically along the cell cortex and subsequently direct the localization and/or activity of other proteins. Here I discuss recent findings on how PAR proteins become and remain asymmetric in three different contexts during C. elegans development: anterior-posterior polarization of the one-cell embryo, apicobasal polarization of non-epithelial early embryonic cells, and apicobasal polarization of epithelial cells. Although polarity within each of these cell types requires PAR proteins, the cues and regulators of PAR asymmetry can differ. PMID- 15666356 TI - Spectrin repeat proteins in the nucleus. AB - Spectrin repeat sequences are among the more common repeat elements identified in proteins, typically occurring in large structural proteins. Examples of spectrin repeat-containing proteins include dystrophin, alpha-actinin and spectrin itself- all proteins with well-demonstrated roles of establishing and maintaining cell structure. Over the past decade, it has become clear that, although these proteins display a cytoplasmic and plasma membrane distribution, several are also found both at the nuclear envelope, and within the intranuclear space. In this review, we provide an overview of recent work regarding various spectrin repeat containing structural proteins in the nucleus. As well, we hypothesize about the regulation of their nuclear localization and possible nuclear functions based on domain architecture, known interacting proteins and evolutionary relationships. Given their large size, and their potential for interacting with multiple proteins and with chromatin, spectrin repeat-containing proteins represent strong candidates for important organizational proteins within the nucleus. Supplementary material for this article can be found on the BioEssays website (http://www.interscience.wiley.com/jpages/0265-9247/suppmat/index.html). PMID- 15666357 TI - Effect of varied release kinetics of the osteogenic thrombin peptide TP508 from biodegradable, polymeric scaffolds on bone formation in vivo. AB - This study was designed to assess the influence of varied release kinetics of the osteogenic thrombin peptide TP508 from osteoconductive poly(propylene fumarate) based (PPF) composite scaffolds on bone formation in vivo. Four classes of scaffolds were constructed with different TP508 dosages (200, 100, or 0 microg) and release kinetics (large burst release, minimal burst release, or no release) and implanted in 15.0 mm segmental defects in rabbit radii. The animals were euthanized at 12 weeks and the implants were analyzed by light microscopy, histological scoring analysis, and histomorphometric analysis. Samples from all classes displayed bone growth within the pores of the scaffold near the edges of the defect. In areas where bone was not observed, the pores were filled with mostly fibrous tissue and exhibited minimal inflammatory response for all classes. In contrast to other scaffold classes, scaffolds containing a total dose of 200 microg TP508 and exhibiting a large burst release profile showed statistically more bone formation guided along the surface of the scaffold, with these scaffolds averaging 80% of the defect length bridged with bone compared to 10% or less bridged for the other scaffold classes. These results demonstrate that the extent of in vivo bone formation in response to controlled release from PPF-composite scaffolds is determined by the release kinetics of the incorporated osteogenic peptide. PMID- 15666358 TI - To SIR with Polycomb: linking silencing mechanisms. AB - Yeast SIR2, the most evolutionarily conserved deacetylase, plays an essential role in epigenetic silencing at the silent mating type loci and telomeres. SIR2 has been implicated in chromatin silencing and lifespan determination in several organisms. Discovery that Drosophila SIR2 is also involved in epigenetic silencing mediated by the Polycomb group proteins and is physically associated with a complex containing the E(Z) histone methyltransferase has wide implications. These findings suggest possible link of Polycomb system to diverse cellular processes including senescence. PMID- 15666359 TI - Receptor for advanced glycation end products (RAGE) and its ligand, amphoterin are overexpressed and associated with prostate cancer development. AB - BACKGROUND: Advanced glycation end products (AGE) are produced with normal aging. Recently, some reports indicated that the interaction between AGE and the cognate receptor (RAGE) has a role in cancer dependent. METHODS: We investigated RAGE and amphoterin mRNA expression in prostate cancer cell lines (DU145, PC-3, and LNCaP cells), hormone-refractory prostate cancer tissues, and paired untreated primary prostate cancer and normal prostate (including benign prostatic hypertrophy (BPH)) tissues using real-time quantitative PCR. Moreover, to confirm the AGE RAGE interaction in prostate cancer, DU145 cells stimulated with AGE-bovine serum albumin (AGE-BSA) were examined by in vitro matrigel assay, cell viability assay, MTT assay, reverse transcription-polymerase chain reaction (RT-PCR), and Western blot. RESULTS: DU145 cells, a hormone-independent prostate cancer cell line, showed the highest RAGE mRNA expression. Amphoterin mRNA was expressed in all three cell lines. In prostate tissues, untreated prostate cancer tissue and hormone-refractory prostate cancer tissue showed higher RAGE and amphoterin mRNA expression than normal prostate tissue. The AGE-RAGE interaction induced the invasion and growth in DU145 cells stimulated with AGE-BSA. CONCLUSIONS: The AGE RAGE interaction is important in prostate cancer development, and inhibition of this interaction has potential as a new molecular target for cancer therapy or prevention. PMID- 15666361 TI - Temporal effects of a COX-2-selective NSAID on bone ingrowth. AB - The effects of a short course of a COX-2 inhibitor on bone healing when the drug is discontinued are unknown. We examined the effects of rofecoxib on bone ingrowth over a 6-week period using a well-defined animal model. The Bone Harvest Chamber was implanted bilaterally in mature rabbits. After osseointegration of the chamber, the following treatments were given for 6 weeks each, followed by a harvest in each case: control-no drug; oral rofecoxib (12.5 mg/day) for the first 2 of 6 weeks; washout period-no drug; oral rofecoxib for the last 2 of 6 weeks; washout period-no drug; rofecoxib given continuously for all 6 weeks. Harvested specimens were snap-frozen, cut into serial 6-microm sections, and stained with hematoxylin and eosin and alkaline phosphatase (osteoblast marker), and processed using immunohistochemistry to identify the vitronectin receptor (osteoclast-like cells). Rofecoxib given continuously for 6 weeks yielded statistically less bone ingrowth compared to the control treatment. Rofecoxib given during the initial or final 2 weeks of a 6-week treatment did not appear to interfere with bone ingrowth. This suggests that the effects of COX-2 inhibitors on bone are less profound when the drug is administered for a short period of time. PMID- 15666362 TI - Evidence for downregulation of calcium signaling proteins in advanced mouse adenocarcinoma. AB - BACKGROUND: Prostate cancer (PCa) is the leading cancer related death in America. Gleason grading is currently the predominant method for prediction, with only few biomarkers available. More biomarkers, especially as they relate to cancer progression are desirable. METHODS: The abundance of several important proteins in prostate tissue was compared between wild-type mouse dorsal prostate and well differentiated transgenic adenocarcinoma mouse prostate (TRAMP) mouse dorsal prostates, and between wild-type mouse dorsal prostate and poorly-differentiated TRAMP mouse tumor tissue. 2DIGE method in conjunction with MALDI-ToF and Western blots was used to determine differential expression. RESULTS: In TRAMP dorsal prostates with well-differentiated adenocarcinoma, there were few significant changes in the protein abundances compared to wild-type dorsal prostates, with the exception of increases in proliferating cell nuclear antigen (PCNA) and beta tubulin, two proteins implicated in cell proliferation, and a more than 2-fold increase in Hsp60, a protein involved in the suppression of apoptosis. In the poorly-differentiated tumors, the changes in protein abundance were substantial. While some of those changes could be related to the disappearance of stromal tissue or the appearance of epithelial tissue, other changes in protein abundance were more significant to the cancer development itself. Most notable was the overall decrease in calcium homeostasis proteins with a 10-fold decrease in calreticulin and Hsp70 and a 40-fold decrease in creatine kinase bb in the cancerous tissue. CONCLUSIONS: Proteomics of TRAMP mice provide an excellent method to observe changes in protein abundance, revealing changes in pathways during cancer progression. PMID- 15666363 TI - Cellular evaluation of oral chemotherapy carriers. AB - Hydrogel nanospheres composed of methacrylic acid and poly(ethylene glycol) were loaded with bleomycin and tested as a potential oral delivery system for chemotherapeutic agents. The gastrointestinal epithelium was modeled through the use of Caco-2 monolayers for studies of permeation enhancement by the carriers as well as bleomycin transport. Bleomycin efficacy following release from the carrier was evaluated with a DLD-1 tumor cell model. The nanospheres release bleomycin in response to a pH increase similar to that seen when passing from the stomach into the upper small intestine. These carriers can also increase the permeability of a model of the epithelial barrier, which would hopefully improve drug transport into the bloodstream. Efficacy studies using a tumor cell model showed retention of activity for bleomycin following loading and release from the nanospheres. The carriers described performed well during in vitro evaluation and can hopefully expand the spectrum of chemotherapeutic agents capable of being administered orally. PMID- 15666364 TI - Mechanical performance and osteoblast-like cell responses of fluorine-substituted hydroxyapatite and zirconia dense composite. AB - A fluorine-substituted hydroxyapatite (FHA) and zirconia (ZrO(2)) dense composite (50:50 by volume) was fabricated, and its feasibility for hard tissue applications was investigated in terms of its mechanical properties and osteoblast-like cell (MG63) responses in vitro. The incorporation of fluorine into the hydroxyapatite (HA) structure was highly effective in producing a completely dense apatite-ZrO(2) composite through a pressureless sintering route, by preventing the thermal degradation of the apatite and ZrO(2). The resultant FHA-ZrO(2) dense composite had excellent mechanical properties, such as flexural strength (310 MPa), fracture toughness (3.4 MPam(1/2)), hardness (10 GPa), and elastic modulus (160 GPa). The flexural strength and fracture toughness of the composite showed a noticeable improvement by a factor of approximately 4 with respect to the pure apatites (HA and FHA). The MG63 cellular responses to the composite were assessed in terms of the cell proliferation (cell number and [(3)H]-thymidine incorporation) and differentiation (alkaline phosphatase activity, osteocalcin, and collagen production). The cells on the FHA-ZrO(2) composite spread and grew well, and proliferated actively during the culture period. The expression of alkaline phosphatase, osteocalcin, and collagen by the cells on the composite showed a similar trend to that on the pure apatites, although slight down-regulations were observed, implying that the FHA-ZrO(2) 50:50 composite retains the osteoblastic functionality and traits of the pure HA ceramics to a high degree. This finding, in conjunction with the considerable improvements in mechanical properties, supports the extended use of this composite for hard tissue applications. PMID- 15666365 TI - Adherent apatite coating on titanium substrate using chemical deposition. AB - Plasma-sprayed "HA" coatings on commercial orthopedic and dental implants consist of mixtures of calcium phosphate phases, predominantly a crystalline calcium phosphate phase, hydroxyapatite (HA) and an amorphous calcium phosphate (ACP) with varying HA/ACP ratios. Alternatives to the plasma-spray method are being explored because of some of its disadvantages. The purpose of this study was to deposit an adherent apatite coating on titanium substrate using a two-step method. First, titanium substrates were immersed in acidic solution of calcium phosphate resulting in the deposition of a monetite (CaHPO4) coating. Second, the monetite crystals were transformed to apatite by hydrolysis in NaOH solution. Composition and morphology of the initial and final coatings were identified using X-ray diffraction (XRD), Scanning Electron Microscopy, and Energy Dispersive Spectroscopy (EDS). The final coating was porous and the apatite crystals were agglomerated and followed the outline of the large monetite crystals. EDS revealed the presence of calcium and phosphorous elements on the titanium substrate after removing the coating using tensile or scratching tests. The average tensile bond of the coating was 5.2 MPa and cohesion failures were observed more frequently than adhesion failures. The coating adhesion measured using scratch test with a 200-microm-radius stylus was 13.1N. Images from the scratch tracks demonstrated that the coating materials were squashed without fracturing inside and/or at the border of the tracks until the failure point of the coating. In conclusion, this study showed the potential of a chemical deposition method for depositing a coating consisting of either monetite or apatite. This method has the advantage of producing a coating with homogenous composition on even implants of complex geometry or porosity. This method involves low temperatures and, therefore, can allow the incorporation of growth factors or biogenic molecules. PMID- 15666366 TI - Splicing mutation in the fibrillin-1 gene associated with neonatal Marfan syndrome and severe pulmonary emphysema with tracheobronchomalacia. AB - Neonatal Marfan syndrome is an autosomal-dominant connective tissue disease with unique clinical manifestations and mutations. We describe the clinical course of an infant with neonatal Marfan syndrome that had the novel IVS31-2A > G splice site mutation in fibrillin-1. This mutation affects the second base of the acceptor consensus splice site of intron 31, and probably leads to abnormal splicing events. The patient presented with respiratory distress and heart murmur in early neonatal life. Cardiac evaluation revealed pulmonic stenosis, atrioventricular regurgitation, and a dilated aortic root that were controlled by balloon dilatation of the pulmonic stenosis and medications for congestive heart failure. At age 3 months, he presented with severe respiratory distress caused by upper and lower airway obstruction. Imaging studies showed severe pulmonary emphysema, and a bronchoscopy demonstrated megatracheobronchomalacia, an unusual finding in this syndrome. Subsequently, the patient developed recurrent hyperinflation of the right and left lungs, with emphysematous changes and mediastinal shift. After discussing with his parents the grave prognosis for neonatal Marfan syndrome, he was discharged home with oxygen treatment and died at home at age 4.5 months. This case report demonstrates and discusses pulmonary involvement in neonatal Marfan syndrome and the difficult therapeutic challenges created by the severe cardiopulmonary abnormalities in this invariably fatal condition. PMID- 15666367 TI - Comparison of blood pressure measurements in children with and without habitual snoring. AB - Higher or similar systolic and/or diastolic blood pressure has been recorded in children with sleep apnea compared to subjects with primary snoring or in those with primary snoring compared to controls. To investigate the association between blood pressure and habitual snoring, we studied children in four randomly selected schools in central Greece. A symptom questionnaire was answered by parents, and children's weight, height, and blood pressure were measured. Seven hundred and sixty children (4-14 years old; 352 female) were recruited. Fifty of 760 (6.6%) participants were snoring more than 3 nights/week (habitual snorers). Mean (+/- SD) systolic blood pressure was 106.9 (+/-10.6) mmHg in habitual snorers vs. 107 (+/- 12) in nonhabitual snorers (P > 0.05). Mean diastolic blood pressure was 61.9 (+/- 7.6) in the former vs. 61.8 (+/- 6.8) in the latter (P > 0.05). While age, gender, and body mass index were significant predictors of systolic blood pressure in a general linear model, snoring was not. Similarly, that gender and body mass index but not snoring were significant predictors of diastolic blood pressure. In a community sample of children, habitual snorers do not have higher morning systolic or diastolic blood pressure than nonhabitual snorers. PMID- 15666368 TI - Intramuscular triamcinolone for difficult asthma. AB - We treated a selected group of children attending a difficult asthma clinic with intramuscular triamcinolone acetonide. This study retrospectively reviews markers of asthma severity in those who received one or more monthly doses for three periods: 1) 3 months preceding the first injection (pretreatment), 2) from the first injection to 1 month after the last injection (treatment period), and 3) 3 months after the treatment period (follow-up period). Severity markers during the treatment and follow-up periods were compared with the pretreatment period by paired t-test. Five children (5-13 years old) received a single dose, and 8 children (12-15 years old) received multiple doses. Multiple doses of triamcinolone (n = 3-5) were associated with a fall in the number of asthma exacerbations (P < 0.01) and hospital admissions (P < 0.01) in both the treatment and follow-up periods. A single dose reduced exacerbations (P < 0.05, treatment vs. pretreatment) but not hospital admissions. We conclude that intramuscular triamcinolone is a useful short-term therapy in difficult asthma. Whether its efficacy is due to improved compliance, or an improved anti-inflammatory profile compared with oral steroids, remains unclear. PMID- 15666370 TI - Detection of Pseudomonas aeruginosa type III antibodies in children with tracheostomies. AB - Pseudomonas aeruginosa is often cultured from the airways of children with tracheostomies. P. aeruginosa produces exotoxin A (ETA) and type III cytotoxins. This study tested the hypothesis that children with tracheostomies are colonized by P. aeruginosa that express these virulence factors and will have antibodies directed against these virulence factors, indicating infection rather than only colonization. A convenience sample of 30 patients, ranging in age from 2 months 22 years, was recruited. Serum was tested for the presence of antibodies to ETA and components of the type III system by Western blot analysis. Twenty-one of 39 patients (70%) had antibodies to components of the type III system. Fifteen of 30 (50%) were seropositive for ETA. Sera from patients who were antibody-positive for ETA were also seropositive for either ExoS or ExoU. Nine of 30 patients (30%) did not possess antibodies to ETA or components of the type III system. In conclusion, these data identified a seropositive reaction to P. aeruginosa cytotoxins in some patients with tracheostomies, suggestive of infection by cytotoxic strains of P. aeruginosa. Future studies will determine the utility of measuring seroconversion to these cytotoxins as an early indication of infection in children with tracheostomies. PMID- 15666371 TI - Resurgence of biliary cast syndrome. PMID- 15666372 TI - Reporting outcomes on living donor liver transplant. PMID- 15666373 TI - Amelioration of hypertrophic cardiomyopathy using nonsurgical septal ablation in a cirrhotic patient prior to liver transplantation. AB - A 53-year-old male with hepatitis C cirrhosis, who had been refused liver transplantation because of hypertrophic cardiomyopathy (HC), underwent nonsurgical septal ablation using alcohol with resolution of his ventricular outflow obstruction. This patient was able to subsequently undergo a successful deceased donor liver transplantation. This is the first reported case of alcohol induced septal ablation being performed in a cirrhotic patient with HC. Such nonsurgical procedures may be attractive in cirrhotic patients who are refused access to liver transplantation because of high surgical risk. PMID- 15666374 TI - Transmission of West Nile virus by organ transplantation. PMID- 15666375 TI - Liver transplantation with simultaneous removal of an intracardiac transjugular intrahepatic portosystemic shunt and a vena cava filter without the utilization of cardiopulmonary bypass. AB - Transjugular intrahepatic shunts (TIPSs) are widely used in the management of portal hypertension complications including variceal bleeding, refractory ascites, and hepatic hydrothorax. Vena cava filters (VCFs) are an important therapeutic modality in the prevention of pulmonary emboli in patients suffering deep venous thrombosis and clinical contraindications for anticoagulation. Stent and filter misplacement or migration may occur, complicating liver transplantation (LT) surgery. We describe the intraoperative management of a patient with cirrhosis, who had a TIPS extending into the right atrium (RA) and a retrohepatic VCF. Stent and filter removals were deferred until the time of LT. Both procedures were performed successfully by complete cava and portal reconstruction. In conclusion, careful assessment and surgical management of patients with stent and filters permits successful LT. PMID- 15666376 TI - Compression of the brachial plexus during right lobe liver donation as a cause of brachial plexus injury: a case report. AB - We present a case of brachial plexus injury in a living-related liver donor, most likely caused by compression of the plexus between the 1st rib and clavicle, the result of rib retraction for surgical exposure. PMID- 15666378 TI - Liver transplantation for HELLP syndrome. AB - Hemolysis, elevated liver enzymes, and low platelet count (HELLP) syndrome is a rare complication of pregnancy that is associated with preeclampsia and may result in rupture of the liver. Although there have been case reports of liver transplantation for HELLP syndrome, the outcomes of transplantation for this rare indication have not been reported. Furthermore, the optimal management of complicated HELLP syndrome and indications for liver transplantation are unclear. Our objective was to review the national experience with liver transplantation for HELLP syndrome and to develop a comprehensive algorithm for the management of liver complications of HELLP syndrome, including indications for transplantation. A recent case from our institution is reported and the literature is reviewed. The results of liver transplantation for HELLP syndrome were analyzed from the United Network for Organ Sharing database. Between October 1987 and November 2003 there have been 8 deceased donor liver transplants performed for complications related to HELLP syndrome. As of the most recent follow-up, 6 of the 8 patients are alive, with both deaths occurring within 1 month of transplantation, and 2 patients have required retransplantation. This review supports that good results can be obtained with liver transplantation for patients with complicated HELLP syndrome that have either ongoing, uncontrolled hemorrhage or liver necrosis and failure. Patients with complicated HELLP syndrome are best managed at a center with expertise in liver transplantation. PMID- 15666377 TI - Colonization with methicillin-resistant Staphylococcus aureus after liver transplantation. AB - Methicillin-resistant Staphylococcus aureus (MRSA) is a frequent cause of infection after orthotopic liver transplantation (OLT). Colonization with MRSA is associated with a higher risk of infection. Previous studies have shown a high prevalence of MRSA colonization among OLT candidates. However, the risk of colonization with MRSA after OLT is still unclear. The objective of this study was to estimate the incidence and the factors associated with colonization with MRSA after OLT. This was a prospective cohort study including patients submitted to OLT between the years 2000 and 2002. Surveillance cultures of nasal swab specimens were performed within the 1st 72 hours of hospital admission and, subsequently, on weeks 2, 6, 13, and 26. Patients whose baseline cultures revealed nasal carriage of MRSA were excluded. A total of 60 patients were included in the study. The median follow-up was 72 days. A total of 9 patients (15%) became colonized. In multiple logistic regression analyses, the use of a urinary catheter for > or =5 days (P = .006), postoperative bleeding at the surgical site (P = .009), and preoperative use of fluoroquinolones (P = .08) were associated with a higher risk of colonization. Patients without any of these risk factors did not become colonized. In conclusion, nasal carriage of MRSA is frequently acquired after OLT. Periodic postoperative screening for MRSA carriage should be an integral component in programs designed to reduce nosocomial MRSA transmission in these patients. Further studies are needed to set up and validate a predictive model that could allow targeting postoperative screening to high risk OLT recipients. PMID- 15666379 TI - Noninvasive ventilation for pediatric patients including those under 1-year-old undergoing liver transplantation. AB - Pulmonary complications are an important cause of the mortality associated with liver transplantation. The efficacy of noninvasive ventilation (NIV) in pediatric patients following transplantation is unknown. The purpose of this retrospective study is to investigate the effects of NIV for pediatric patients undergoing liver transplantation. Of 102 pediatric patients who underwent liver transplantation, 15 patients (aged 73 months; range 2.5-179) were supported by NIV because of atelectasis, hypercapnia, hypoxemia, pneumonia, massive effusion, or postextubation ventilatory support. Of 15 patients, 5 were under the age of 1 year (range 2.5-12 months). Of the 15 patients, 7 had required multiple intubations before NIV treatment because of pulmonary complications. NIV treatment was administered to 6 patients because of hypercapnia. Partial pressure of arterial carbon dioxide (PaCO(2)) levels improved from 56.9 (95% confidence interval [CI]: 48.4-65.4) to 41.5 (95% CI: 36.8-46.2) mmHg (P = .028) within 2 days. NIV treatment was very effective for patients with atelectasis with and without other pulmonary complications. Mean inspiratory positive pressure (IPAP) was 7.2 (95% CI: 6.0-8.3) cm H(2)O and expiratory positive pressure (EPAP) was 3.5 (95% CI: 3.2-3.9) cm of H(2)O. Mean duration of NIV was 18.5 (95% CI: 8.6 28.4) days. IPAP and EPAP levels were closely and significantly correlated with height (IPAP: r = .65, P = .016; EPAP: r = .77, P = .004). A total of 13 patients recovered and 2 patients died. However, no patient died of respiratory complications. In conclusion, NIV is effective in pediatric patients undergoing liver transplantation with subsequent pulmonary complications. The IPAP and EPAP levels may be predicted by the height of the patient. PMID- 15666380 TI - Ischemic preconditioning in deceased donor liver transplantation: a prospective randomized clinical trial of safety and efficacy. AB - Ischemic preconditioning (IPC) has the potential to decrease graft injury and morbidity after liver transplantation. We prospectively investigated the safety and efficacy of 5 minutes of IPC induced by hilar clamping in local deceased donor livers randomized 1:1 to standard (STD) recovery (N = 28) or IPC (N = 34). Safety was assessed by measurement of heart rate, blood pressure, and visual inspection of abdominal organs during recovery, and efficacy by recipient aminotransferases (aspartate aminotransferase [AST] and alanine aminotransferase [ALT], both measured in U/L), total bilirubin, and international normalized ratio of prothrombin time (INR) after transplantation. IPC performed soon after laparotomy did not cause hemodynamic instability or visceral congestion. Recipient median AST, median ALT, and mean INR, in STD vs. IPC were as follows: day 1 AST 696 vs. 841 U/L; day 3 AST 183 vs. 183 U/L; day 1 ALT 444 vs. 764 U/L; day 3 ALT 421 vs. 463 U/L; day 1 INR 1.7 +/- .4 vs. 2.0 +/- .8; and day 3 INR 1.3 +/- .2 vs. 1.4 +/- .3; all P > .05. No instances of nonfunction occurred. The 6 month graft and patient survival STD vs. IPC were 82 vs. 91% and median hospital stay was 10 vs. 8 days; both P > .05. In conclusion, deceased donor livers tolerated 5 minutes of hilar clamping well, but IPC did not decrease graft injury. Further trials with longer periods of preconditioning such as 10 minutes are needed. PMID- 15666381 TI - Preharvest donor hyperoxia predicts good early graft function and longer graft survival after liver transplantation. AB - A total of 44 donor/recipient perioperative and intraoperative variables were prospectively analyzed in 89 deceased-donor liver transplantations classified as initial good graft function (IGGF) or initial poor graft function (IPGF) according to a scoring system based on values obtained during the 1st 72 postoperative hours from the serum alanine aminotransferase (ALT) concentration, bile output, and prothrombin activity. The IGGF compared with the IPGF group showed: 1) longer graft (P = .002) and patient (P = .0004) survival; 2) at univariate analysis, a higher (mean [95% confidence interval]) preharvest donor arterial partial pressure of oxygen (PaO(2)) (152 [136-168] and 104 [91-118] mmHg, respectively; P = .0008) and arterial hemoglobin oxygen saturation (97.9 [97.2-98.7] and 96.7 [95.4-98.0]%, respectively; P = .0096), a lower percentage of donors older than 65 years (13 and 33%, respectively; P = .024), a lower percentage of donors treated with noradrenaline (16 and 41%, respectively; P = .012). At multivariate analysis, IGGF was associated positively with donor PaO(2) and negatively with donor age greater than 65 years and with donor treatment with noradrenaline. Independently from the grouping according to initial graft function, graft survival was longer when donor PaO(2) was >150 mmHg than when donor PaO(2) was < or =150 mmHg (P = .045). In conclusion, preharvest donor hyperoxia predicts IGGF and longer graft survival. PMID- 15666382 TI - How common is delayed cyclosporine absorption following liver transplantation? AB - The mean time to peak absorption of cyclosporine (CsA) in liver transplant patients is approximately 2 hours, but in some patients the peak occurs later. The goal of this study was, therefore, to investigate the incidence of delayed absorption in 27 de novo liver transplant recipients receiving CsA > or =10 mg/kg/day (C(2) monitoring) and in 15 maintenance patients. Patients were categorized as 'normal' absorbers (C(2) exceeding C(4) and C(6)) or 'delayed' absorbers (C(4) or C(6) exceeding C(2)), and as 'good' (>800 ng/mL at C(0), C(2), C(4), or C(6)) or 'poor' absorbers (C(0), C(2), C(4) and C(6) <800 ng/mL) on the day of study. Among de novo patients, 15 (56%) had 'normal' CsA absorption and 12 (44%) 'delayed' absorption. Good CsA absorption occurred in 16 patients (59%) and poor absorption in 11 (41%). The proportion of poor absorbers was similar in patients with normal (6/15, 40%) or delayed (5/12, 42%) absorption. Among the 12 delayed absorbers, 11 had peak CsA concentration at C(4). Mean C(0) level was significantly higher in delayed absorbers (282 +/- 96 ng/mL) than in normal absorbers (185 +/- 88 ng/mL; P = .01). Delayed absorbers reverted to normal absorption (C(2) > C(4)) after a median of 6 days from the day of study, and no cases of delayed absorption were found among maintenance patients. In conclusion, almost 50% of the patients had delayed CsA absorption early posttransplant; around half of these exhibited normal CsA exposure. Measurement of C(4) in addition to C(2) differentiates effectively between delayed and poor absorbers of CsA such that over- or underimmunosuppression can be avoided. PMID- 15666383 TI - MELD and the quality of life. PMID- 15666384 TI - Understanding the meaning of fat in the liver. PMID- 15666386 TI - Management of pre-liver transplantation patients--part 1. PMID- 15666387 TI - Use of thymosin beta15 as a urinary biomarker in human prostate cancer. AB - BACKGROUND: Additional prostate cancer (CaP) biomarkers are needed to increase the accuracy of diagnosis and to identify patients at risk of recurrence. In tissue-based assays, thymosin beta15 (Tbeta15) has been linked to an aggressive CaP phenotype and correlated with future tumor recurrence. We hypothesized that Tbeta15 may have clinical utility in biological fluids. METHODS: Tbeta15 was measured in urine from CaP patients; untreated (N = 61), prostatectomy (RP, N = 46), androgen deprivation therapy (ADT, N = 14) and control groups; normal (N = 52), genitourinary carcinoma (N = 15), non-malignant prostate disease (N = 81), and other urology (N = 73). We evaluated the utility of urinary Tbeta15 for CaP diagnosis, alone or in combination with prostate-specific antigen (PSA), and the relationship to CaP progression. RESULTS: A normal threshold of 40 (ng/dl)/(mug_protein/mg_creatinine) was defined using receiver operating characteristic analysis and marked the 19th centile for age-matched controls. The proportion of untreated CaP patients with urinary Tbeta15 above the threshold was significantly higher than normal and genitourinary disease controls (P < 0.001). RP caused urinary Tbeta15 to drop significantly (P = 0.005). Pre-surgery Tbeta15 concentrations greater than the normal threshold may confer greater risk of CaP recurrence. Relative to normal controls, patients receiving ADT for aggressive CaP were 12 times more likely to have elevated urinary Tbeta15 (P = 0.001, 95% CI = 2.8, 51.8). Combining PSA and Tbeta15 (PSA > 4, or PSA > 2.5, Tbeta15 > 40, or PSA = 2.5, Tbeta15 > 90) provided the same sensitivity as a 2.5 ng/ml PSA cutoff, but markedly improved diagnostic specificity. CONCLUSIONS: We report that Tbeta15 is a urinary biomarker for CaP and suggest that Tbeta15, in combination with PSA, can be used to improve both the sensitivity and specificity of CaP diagnosis. PMID- 15666389 TI - Raf-1 expression may influence progression to androgen insensitive prostate cancer. AB - BACKGROUND: Recent evidence has implicated the MAP kinase pathway with the development of androgen insensitive prostate cancer (AIPC). We have previously reported gene amplification of critical members of this pathway with the development of androgen insensitive disease. METHODS: Protein expression of Raf-1 was analyzed using immunohistochemistry (IHC) in a database of 65 paired tumor specimens obtained before and after the development of AIPC and correlated with other members of the pathway. RESULTS: Patients whose Raf-1 expression rose with development of AIPC had a significantly shorter median time to biochemical relapse compared to those whose expression fell or remained unchanged (1.16 vs. 2.62 years, P = 0.0005). In AIPC tumors, expression of Raf-1 correlated significantly with expression of HER2 and with expression of c-fos. CONCLUSIONS: We conclude that the HER2/Raf-1/AP-1 axis may promote the development of AIPC, leading to early relapse. Members of the pathway may act as novel therapeutic targets for patients. PMID- 15666390 TI - Basal growth hormone concentrations in blood and the risk for prostate cancer: a case-control study. AB - OBJECTIVE: To assess the relationship between basal serum growth hormone (GH) levels and prostate cancer risk. METHODS: We conducted a population-based case control study; cases included 68 men, aged 45-85 years, diagnosed with incident, primary, histologically confirmed, and clinically apparent (stage B and higher) prostate cancer. Controls included 240 men, frequency matched on age and residential area. Age, race, BMI, waist circumference, history of enlarged prostate, education, and current smoking status, were all considered as possible confounders. RESULTS: We found a statistically significant trend of decreasing prostate cancer risk across increasing GH quintiles, in both crude (OR: 0.31, 95% CI: 0.12-0.83, P for trend 0.01) and adjusted models (OR: 0.35, 95% CI: 0.12 1.05, P for trend 0.03), in the highest compared to the lowest quintile, respectively. CONCLUSIONS: Lower basal levels of GH in serum are associated with increased prostate cancer risk. The inverse association may be explained by the negative feedback loop generated by IGF-1 produced by the tumor on GH secretion. PMID- 15666391 TI - Negative impact of systemic catecholamine administration on hepatic blood perfusion after porcine liver transplantation. AB - Catecholamines are often administered during and after liver transplantation (LTx) to support systemic perfusion and to increase organ oxygen supply. Some vasoactive agents can compromise visceral organ perfusion. We followed the hypothesis that the vasculature of transplanted livers presents with a higher sensitivity, which leads to an increased vulnerability for flow derangement after application of epinephrine (Epi) or norepinephrine (NorEpi). Hepatic macroperfusion and microperfusion during systemic Epi or NorEpi infusion were measured by Doppler flow and thermodiffusion probes in porcine native, denervated, and transplanted livers (n = 16 in each group). Epi or NorEpi were infused (n = 8 in each subgroup) in predefined dosages (low dose = 5 microg/kg/minute and high dose = 10 microg/kg/minute) over 240 minutes. Systemic cardiocirculatory parameters were monitored continuously. Hepatic perfusion data were compared between all groups at comparable time points and dosages. In all native, denervated, and transplanted liver groups, Epi and NorEpi induced an inconsistent rise of mean arterial pressure and heart rate shortly after onset of infusion in both dosages compared with baseline. No significant differences of cardiovascular parameters at comparable time points were observed. In native livers, Epi and NorEpi induced only temporary alterations of hepatic macrocirculation and microcirculation, which returned to baseline 2 hours after onset of infusion. No significant alterations of hepatic blood flow were detected after isolated surgical denervation of the liver. By contrast, transplanted livers showed a progressive decline of hepatic macrocirculation (33-75% reduction) and microcirculation (39-58% reduction) during catecholamine infusions in a dose-dependent fashion. Characteristics of liver blood flow impairment were comparable for both vasoactive agents. In conclusion, pronounced disturbances of hepatic macrocirculation and microcirculation were observed during systemic Epi and NorEpi infusion after LTx compared with native and denervated livers. Microcirculation disturbances after LTx might be explained by impairment of hepatic blood flow regulation caused by an increased sensitivity of hepatic vasculature after ischemia-reperfusion and by lengthening of vasopressor effects caused by reduced hepatocyte metabolism. Clinicians should be aware of this potentially hazardous effect. Therefore, application of catecholamines after clinical LTx should be indicated carefully. PMID- 15666393 TI - Presence of multiple bile ducts in the liver graft increases the incidence of biliary complications in pediatric liver transplantation. AB - We studied the impact of multiple bile duct anastomosis on the development of biliary complications after liver transplantation in children. A total of 101 patients received a primary liver transplant and were divided into 2 groups: those with a single bile duct (n = 77) and those with multiple bile ducts (n = 24). Mean follow-up was 39.8 +/- 20.8 months. A total of 27 patients presented with biliary complications (26.7%), 18 patients (18.7%) presented with early complications (12 leaks and 6 strictures), and 9 patients (8.9%) had late strictures. Hepatic artery thrombosis (HAT) and multiple bile ducts were significant risk factors for the development of biliary complications, and the presence of multiple bile ducts was an independent risk factor. Patients with multiple bile ducts had a significantly greater incidence of total biliary complications compared to those with single ducts. Patients with multiple ducts had a higher incidence of leaks when compared to those in the single duct group, but the incidence of strictures, both early and late, was similar in both groups. One-year patient and graft survivals were not statistically different in the 2 groups. In conclusion, the presence of more than one bile duct in the graft is an independent risk factor for the development of biliary complications after pediatric liver transplantation. PMID- 15666392 TI - MELD fails to measure quality of life in liver transplant candidates. AB - Previous studies have demonstrated an association between Child Turcotte-Pugh (CTP) class and impaired quality of life. However, the relationship between the model for end-stage liver disease (MELD) score and quality of life (QOL) has not been well studied. In this study, quality of life questionnaires (Medical Outcomes Short Form 36 [SF-36] and the Chronic Liver Disease Questionnaire [CLDQ]) were administered to 150 adult patients awaiting liver transplantation. We also collected demographic data and laboratory results and recorded manifestations of hepatic decompensation. The study found that all domains of the SF-36 and CLDQ were significantly lower in our patient cohort than in normal controls (P < .001). There was a moderate negative correlation between CPT class and physical components of the SF-36 (r = -.30), while there was a weak negative correlation (r = -.10) between CPT class and the mental component. There was a negative moderate correlation between CPT class and overall CLDQ (r = -.39, P < .001) and a weak correlation (r = -.20) between MELD score and overall CLDQ score. Both encephalopathy (correlation coefficient = -.713, P = .004) and ascites (correlation coefficient = -.68, P = .006) were predictive of the QOL using CLDQ (adjusted R(2) = .1494 and f = 0.000). In conclusion, in liver transplant candidates, the severity of liver disease assessed by the MELD score was not predictive of QOL. The presence of ascites and/or encephalopathy was significantly associated with poor quality of life. CTP correlates better to QOL, probably because it contains ascites and encephalopathy. PMID- 15666394 TI - The hepatic regeneration power of mild steatotic grafts is not impaired in living donor liver transplantation. AB - The aim of this study was to assess histologic changes in steatotic grafts, regenerative capacity, and the outcome of steatotic grafts in living-donor liver transplantation (LDLT). Between September 2002 and February 2004, 55 cases of LDLT with a liver biopsy performed on the 10th postoperative day were enrolled. Patients were grouped according to the intraoperative histologic degree of macrovesicular steatosis (MaS) as follows: Group 1, <5% (n = 24); Group 2, 5 to 15% (n = 24); and Group 3, 15 to 30% (n = 7). The intraoperative microscopic findings and the findings on the 10th postoperative day were compared. Immunohistochemistry was performed using antibody of proliferating cell nuclear antigen (PCNA) and Ki-67 to assess the regeneration power of grafts on the 10th postoperative day. The histologic degree of MaS on postoperative day 10 decreased from 5.22 +/- 1.04% (mean +/- standard deviation) to 2.17 +/- 1.90 in Group 2 (P < .001) and from 21.4 +/- 8.02 to 4.43 +/- 2.70 in Group 3 (P = .003). The number of positively stained hepatocytes in 10 high power fields was 48.0 +/- 17.1, 53.8 +/- 14.4, and 51.5 +/- 4.1 in each group by PCNA (P = .681), and 24.0 +/- 14.0, 25.5 +/- 11.8, and 21.6 +/- 6.8 by Ki-67 (P = .825), respectively. No primary graft nonfunction (PNF) or delayed graft function (DGF) occurred. Major complications were comparable among groups. In conclusion, in LDLT, steatosis disappeared immediately after transplantation and hepatic regeneration power was not impaired in grafts with less than 30% of MaS. Furthermore, a mildly steatotic graft did not increase the risk of graft dysfunction or morbidity in LDLT. PMID- 15666395 TI - Long-term outcome of pediatric liver transplantation for biliary atresia: a 10 year follow-up in a single center. AB - The aim of this study was to review our experience in orthotopic liver transplantation (OLT) for biliary atresia (BA) in children and analyze the survival and prognostic factors, and long-term outcome. We reviewed 332 OLTs performed in 280 children between the years 1986 and 2000. Univariate and multivariate analysis were performed on patient and graft survivals according to recipients' and donors' characteristics as well as intraoperative data. The long term outcome among the 80 children living at 10 years after OLT was studied according to growth, immunosuppressive therapy, and liver and renal functions. Liver graft status was eventually documented by liver biopsy. Status of rehabilitation was assessed by reviewing school performance and employment. Overall patient survival rates at 1, 5, and 10 years were 85, 82, and 82%, respectively, and the corresponding overall graft survival rates were 77, 73, and 71%. In the multivariate analysis, we identified 4 independent prognostic factors: polysplenia syndrome (P = .03), United Network for Organ Sharing (UNOS) status (P = .05), donor's age (P = .01), and perioperative surgical complications (P = .03). At 10 years after transplant, 80 children were alive and had normal growth rates. Liver histology was abnormal in 73% of these long-term survivors, mainly due to chronic rejection and centrilobular fibrosis. A total of 63 of the 80 children attended normal school and in 55 children (69%) school performance was not delayed. In conclusion, we discovered that a good long-term survival could be achieved after liver transplantation for BA, with a 82% survival rate at 10 years with normal scholastic studies in the majority of recipients. PMID- 15666397 TI - Central brightening due to constructive interference with, without, and despite dielectric resonance. AB - PURPOSE: To aid in discussion about the mechanism for central brightening in high field magnetic resonance imaging (MRI), especially regarding the appropriateness of using the term dielectric resonance to describe the central brightening seen in images of the human head. MATERIALS AND METHODS: We present both numerical calculations and experimental images at 3 T of a 35-cm-diameter spherical phantom of varying salinity both with one surface coil and with two surface coils on opposite sides, and further numerical calculations at frequencies corresponding to dielectric resonances for the sphere. RESULTS: With two strategically placed surface coils it is possible to create central brightening even when one coil alone excites an image intensity pattern either bright on one side only or bright on both sides with central darkening. This central brightening can be created with strategic coil placement even when the resonant pattern would favor central darkening. Results in a conductive sample show that central brightening can similarly be achieved in weakly conductive dielectric materials where any true resonances would be heavily damped, such as in human tissues. CONCLUSION: Constructive interference and wavelength effects are likely bigger contributors to central brightening in MR images of weakly conductive biological samples than is true dielectric resonance. PMID- 15666396 TI - Interstudy reproducibility of three-dimensional volume-selective fast spin echo magnetic resonance for quantifying carotid artery wall volume. AB - PURPOSE: To assess the interstudy reproducibility of a three-dimensional volume selective, fast spin echo (FSE) magnetic resonance technique for the assessment of carotid artery wall volume, which is a marker for total carotid plaque volume. MATERIALS AND METHODS: Interstudy reproducibility was evaluated in 10 subjects with evidence of carotid artery atherosclerotic disease on carotid Doppler ultrasonography. Subjects were scanned twice with an interscan time of one hour to four days. The carotid artery was imaged in cross-section, and the total carotid arterial wall volume (TWV) was calculated by subtraction of the total carotid lumen volume from the total outer carotid vessel volume. RESULTS: The mean carotid TWV for the scans was 741 and 734 mm3, respectively, with no significant difference (mean difference 7 mm3; P = 0.5). The time for each study was approximately 20 minutes. The standard deviation of the differences between the measurements was 33 mm3, yielding an interstudy coefficient of variation of 4.4%. Sample size calculations showed that 16 patients would enable this difference in plaque volume over time to be detected with 80% power at a P value of 0.05. CONCLUSION: Volumetric analysis with CMR of carotid artery plaques using a three-dimensional volume-selective FSE is efficient with good interstudy reproducibility, and is well suited for longitudinal studies of progression of carotid atheroma with reasonable sample sizes. PMID- 15666398 TI - MR imaging of the normal appendix and acute appendicitis. AB - PURPOSE: To describe the MR appearance of the normal appendix and the MR imaging characteristics of acute appendicitis with correlation to pathological severity. MATERIALS AND METHODS: A total of 20 volunteers participated in this study to demonstrate normal appendices by MR imaging. A total of 37 consecutive patients with clinically diagnosed acute appendicitis were also scanned. T1-weighted (T1WI) spin-echo images, T2-weighted (T2WI) fast spin-echo, and fat-suppressed spectral presaturation inversion recovery T2-weighted (T2SPIR) fast spin-echo images were obtained. The MR criteria for considering acute appendicitis were as follows: 1) thickening of the appendiceal wall with high intensity on T2WI or T2SPIR; 2) dilated lumen filled with high intensity material on T2WI or T2SPIR; and 3) increased intensity of periappendiceal tissue on T2WI or T2SPIR. RESULTS: The visibility of a normal appendix on MR imaging was 90% (18/20). It appeared as a cord-like structure of medium intensity without fluid collection in the lumen. A total of 30 cases with clinically diagnosed acute appendicitis had positive MR findings and all except one were pathologically proven. The one had cecal diverticulitis. These cases demonstrated filled lumen, with a hypointense wall on T1WI and slightly hyperintense on T2WI or T2SPIR. MR findings correlated well with pathological severity, especially a thicker wall, periappendiceal high intensity, and ascites were useful for suspecting severe appendicitis. CONCLUSION: Correct diagnosis of acute appendicitis was obtained with MRI, and correlated well with its pathological severity. MRI is a powerful alternative for diagnosing acute appendicitis especially for the patients in whom the radiation is major concern. PMID- 15666399 TI - Myometrial invasion and cervical invasion by endometrial carcinoma: evaluation by CO2-volumetric interpolated breathhold examination (VIBE). AB - PURPOSE: To assess myometrial invasion and cervical invasion by endometrial carcinoma, using CO2-volumetric interpolated breathhold examination (VIBE) enabling more precise evaluation of depth of tumor invasion. MATERIALS AND METHODS: CO2-VIBE was performed in 21 cases of endometrial carcinoma (Stage Ia IIb) prior to treatment. The images were interpreted by performing multiplanar reconstruction (MPR), and the findings obtained from the images (degree of myometrial invasion and presence or absence of cervical invasion) were assessed in comparison with the histopathological diagnosis. RESULTS: The sites of the endometrial carcinoma lesions were clearly visualized by the CO2-VIBE method. Evaluation of the degree of myometrial invasion enabled a high correct diagnosis rate of 90.5%, and evaluation for the presence of cervical invasion also allowed a high correct diagnosis rate of 90.5%. CONCLUSION: VIBE permits evaluation of any plane desired by means of thin slices, and it is a truly revolutionary method for preoperative evaluation of depth of invasion of endometrial carcinoma that enables highly accurate determination of the extent of lesion sites and degree of invasion before treatment. PMID- 15666400 TI - Coronary arteries at 3.0 T: Contrast-enhanced magnetization-prepared three dimensional breathhold MR angiography. AB - PURPOSE: To evaluate the efficacy of contrast-enhanced coronary magnetic resonance angiography (MRA) at 3.0 T. MATERIALS AND METHODS: Nine healthy human volunteers were studied on a 3.0-T whole-body MR system. A three-dimensional, breathhold, magnetization-prepared, segmented, gradient-echo sequence was used, with injection of 20 mL gadopentetate dimeglumine for each three-dimensional slab. Imaging parameters were optimized based on computer simulations. Signal-to noise ratio (SNR), contrast-to-noise ratio (CNR), depicted coronary artery length, lumen diameter, and imaging sharpness with contrast agent were evaluated. SNR and CNR were compared to the results from a previous 1.5-T study. RESULTS: A 53% increment in SNR and a 305% enhancement in CNR were measured with contrast. Vessel length and sharpness depicted were higher and the lumen diameter was lower (all P values < 0.05) in postcontrast images. Compared to previous results from 1.5-T, the SNR, CNR, and vessel sharpness were enhanced at 3.0 T with higher spatial resolution. CONCLUSION: Contrast-enhanced, three-dimensional, coronary MRA at 3.0 T is a promising technique for diagnosing coronary artery diseases. Patient studies are necessary to evaluate its clinical utility. PMID- 15666402 TI - 3.0-Tesla MR angiography of intracranial aneurysms: comparison of time-of-flight and contrast-enhanced techniques. AB - PURPOSE: To determine whether 3.0-T elliptical-centric contrast-enhanced (CE) magnetic resonance (MR) angiography is superior to 3.0-T elliptical-centric time of-flight (TOF) MR angiography in the detection and characterization of intracranial aneurysms, and to determine whether increasing the acquisition matrix size in 3.0-T CE MR angiography improves image quality. MATERIALS AND METHODS: A total of 50 consecutive patients referred for MR angiographic evaluation of a known or suspected intracranial aneurysm underwent MR angiography, including three-dimensional TOF and elliptical-centric CE techniques at 3.0 T. The 3.0-T three-dimensional TOF and 3.0-T CE examinations were graded for image quality. A blind review identified the presence and location of aneurysms. RESULTS: A total of 28 aneurysms were identified in 23 of the 50 patients. The 3.0-T TOF MR angiography had a higher mean score for image quality than the 3.0-T elliptical-centric CE MR angiography (P < 0.0001). A total of 14 patients with aneurysms had conventional angiography for comparison. The 3.0-T TOF showed all the aneurysms, whereas 3.0-T CE MR angiography did not show 1 of 19 aneurysms when conventional angiography was the reference standard. CONCLUSION: For imaging intracranial aneurysms, 3.0-T TOF MR angiography offers better image quality than 3.0-T CE MR angiography using the elliptical-centric technique. PMID- 15666401 TI - Assessment of left ventricular function by breath-hold cine MR imaging: Comparison of different steady-state free precession sequences. AB - PURPOSE: To compare steady-state free precession (SSFP) sequence protocols with different acquisition times (TA) and temporal resolutions (tRes) due to the implementation of a view sharing technique called shared phases for the assessment of left ventricular (LV) function by breath-hold cine magnetic resonance (MR) imaging. MATERIALS AND METHODS: End-diastolic and end-systolic volumes (EDV, ESV) were measured in contiguous short-axis slices with a thickness of 8 mm acquired in 10 healthy male volunteers. The following true fast imaging with steady-state precession (TrueFISP) sequence protocols were compared: protocol A) internal standard of reference, segmented: tRes 34.5 msec, TA 18 beats per slice; protocol B) segmented, shared phases: tRes 34.1 msec, TA 10 beats per slice; and protocol C) real-time, shared phases, parallel acquisition technique: tRes 47.3 msec, TA 24 beats for 12 slices covering the entire left ventricle. RESULTS: Phase sharing leads to a significant decrease in EDV, stroke volume (SV), and ejection fraction (EF) (median difference -7.0 mL [*], -9.6 mL, and -3.4%, respectively, for protocol B; -15.3 mL, -13.3 mL, and -2.4% for protocol C; P = 0.002, *P = 0.021). The observed median difference of real-time EDV and SV estimates is of clinical relevance. Real-time cine MR imaging shows a greater variability of EDV and SV. No relevant differences in ESV were observed. CONCLUSION: The true cine frame duration of both shared phases sequence protocols exceeds the period of isovolumetric contraction (IVCT) of the left ventricle resulting in a systematic and significant underestimation of EDV and consequently SV and EF. SSFP sequence protocol parameters, particularly tRes and use of view sharing techniques, should therefore be known at follow-up examinations in order to be able to assess LV remodeling in patients with heart failure. PMID- 15666403 TI - Coronary artery wall imaging: initial experience at 3 Tesla. AB - PURPOSE: To assess the feasibility of black-blood turbo spin-echo imaging of the left anterior descending coronary artery wall at 3 Tesla under free-breathing and breath-hold conditions. MATERIALS AND METHODS: Proton density-weighted black blood turbo spin-echo imaging of the left anterior descending coronary artery was performed on 15 volunteers on a 3 T whole body scanner with an eight channel phased array coil. Volunteers were imaged during free-breathing (with navigators, N = 5), or with breath-hold (N = 5), or both (N = 2). Imaging was not possible in three volunteers due to either gradient or radiofrequency (RF) coupling with the electrocardiogram (ECG). Images were analyzed to determine coronary artery wall thickness, wall area, lumen diameter, and lumen area. Signal-to-noise and contrast-to-noise ratios were calculated. RESULTS: Coronary artery wall thickness, wall area, lumen diameter, and lumen area measurements were consistent with previous magnetic resonance (MR) measurements of the coronary wall at 1.5 Tesla. CONCLUSION: Coronary wall imaging using free-breathing and breath-hold two dimensional black-blood TSE is feasible at 3 T. Further improvement in resolution and image quality is required to detect and characterize coronary plaque. PMID- 15666404 TI - Magnetic resonance imaging of in vivo kinematics after total knee arthroplasty. AB - PURPOSE: To improve the quality of magnetic resonance imaging (MRI) on knees after total knee arthroplasty (TKA) by minimizing image artifacts caused by metallic implants, and to establish a method determining in vivo kinematics of TKA knees using MRI. MATERIALS AND METHODS: Two knee implants made of cobalt chrome and oxidized zirconium were tested with different pulse sequences and imaging parameters. Then, in vivo kinematic MRI was performed on five well functioning TKAs under simulated weight-bearing conditions. Kinematic measurements were made and a linear correlation test was run between the tibio- and patellofemoral measurements. RESULTS: The best images with minimum metallic artifacts were observed using oxidized zirconium implants, a fast spin echo sequence (FSE), thin slice thickness, and high readout gradient. TKA kinematics exhibited a large deviation from the normal kinematics and considerable patient to-patient variability. However, significant linear correlations between tibiofemoral and patellofemoral kinematics were observed (R = -0.96, 0.92, 0.88). CONCLUSION: Metallic artifacts due to orthopedic implants can be reduced in MR images for some materials, appropriate pulse sequence, and imaging parameters selection, enabling MR quantification of knee kinematics. Tibiofemoral kinematics appears to affect patellofemoral position after total knee arthroplasty. PMID- 15666405 TI - A modified projection reconstruction trajectory for reduction of undersampling artifacts. AB - PURPOSE: To reduce undersampling artifacts for a given number of repetitions of the projection reconstruction (PR) sequence by modifying its k-space trajectory to sample more mid-frequencies while reducing the sampling coverage of the peripheral spatial frequencies. MATERIALS AND METHODS: The single k-space spoke measured per repetition in the standard PR was modified so that one complete and two partial spokes were measured per repetition but with decreased k-space extent. The point spread functions (PSFs) and undersampling artifacts of the modified PR were compared with those of the standard PR for various numbers of projections. Phantom and in vivo images were used to assess the relative performance. RESULTS: PSF analysis indicated that the modified PR method provided reduced undersampling artifacts with somewhat reduced spatial resolution. The phantom and in vivo images corroborated this. CONCLUSION: The modified PR trajectory provides reduced undersampling artifact vs. the standard PR, particularly when the number of projections is limited and the artifact level is high. PMID- 15666406 TI - Correlation of R2 with total iron concentration in the brains of rhesus monkeys. AB - PURPOSE: To estimate the relationship between R2 = 1/T2 as measured with a double echo spin echo sequence and total iron concentration in gray matter structures in the brains of aging rhesus monkeys. MATERIALS AND METHODS: Using a 1.5-T magnetic resonance (MR) imager, we collected double echo spin echo images of the brains of 12 female rhesus monkeys aged between 9 and 23 years. From the double echo images, the transverse relaxation rate R2 = 1/T2 was calculated in selected gray matter regions. After the animals were euthanized, their brains were excised and tissue punches were taken of the substantia nigra, globus pallidus, and gray matter regions of the cerebellum. Some of the tissue punches were assayed for total iron using atomic absorption spectroscopy. RESULTS: The range of tissue iron concentration spanned from 15 to 450 microg/g wet weight, with the highest levels in the globus pallidus and the lowest levels in the cerebellum. The results show that R2 was highly correlated with the total iron concentration and that the relationship between R2 and tissue iron concentration appeared to depend upon the iron concentration. For concentrations above approximately 150 microg/g wet weight, R2 increased with a sensitivity of 0.0484 +/- 0.0023 second( 1)(microg/g)(-1). In contrast, where the iron concentration was below 150 microg/g, R2 increased at 0.0013 +/- 0.0073 second(-1)(microg/g)(-1). The bilinear behavior may reflect changes with age in the relative amounts of iron distributed diffusely and in granular form in the globus pallidus and substantia nigra. Histological sections of the tissues stained for iron and ferritin support this hypothesis and indicate that the distribution of ferritin is similar to the distribution of iron. CONCLUSION: This study reaffirms the value of measuring the MR relaxation rate R2 for a noninvasive estimate of iron content in the brain and identified limitations in the relationship at low tissue iron concentrations. PMID- 15666407 TI - Indirect detection of lung perfusion using susceptibility-based hyperpolarized gas imaging. AB - PURPOSE: To address the problem of inadequate signal-to-noise ratio (SNR) encountered in lung perfusion magnetic resonance imaging (MRI) by developing an indirect detection based on the strong hyperpolarized (HP) gas signal. MATERIALS AND METHODS: Our model is based on detecting the effects of gadolinium (Gd) flowing through lung capillaries by recording the phase of the nearby alveolar HP gas. In a HP gas 3He phantom we imaged gas phases before and after removing tubes containing paramagnetic solution away from the phantom. We also imaged HP gas phases in pig lungs before and after injection of Gd. Finally, parenchymal spin phase in excised lungs was measured as a function of Gd concentration. RESULTS: In the phantom, the differential phase map displayed a pattern characteristic of a susceptibility-induced dipole field, showing the possibility of an indirect detection. In vivo, the differential phase map showed homogeneous appearance, as expected for uniform perfusion in healthy lungs. Ex vivo, the parenchymal spin phases were shown to depend linearly on Gd concentration. CONCLUSION: Our method should allow indirect perfusion (Q) and direct ventilation (V) to be assessed simultaneously, thus allowing for diagnosis of V/Q mismatches. The linear dependency of parenchymal spin phase vs. Gd concentration may allow for quantification of lung perfusion. PMID- 15666409 TI - Disparity of activation onset in sensory cortex from simultaneous auditory and visual stimulation: Differences between perfusion and blood oxygenation level dependent functional magnetic resonance imaging. AB - PURPOSE: To compare the temporal behaviors of perfusion and blood oxygenation level-dependent (BOLD) functional magnetic resonance imaging (fMRI) in the detection of timing differences between distinct brain areas, and determine potential latency differences between stimulus onset and measurable fMRI signal in sensory cortices. MATERIALS AND METHODS: Inversion recovery (IR) spin-echo echo-planar imaging (EPI) and T2*-weighted gradient-echo EPI sequences were used for perfusion- and BOLD-weighted experiments, respectively. Simultaneous auditory and visual stimulations were employed in an event-related (ER) paradigm. Signal time courses were averaged across 40 repeated trials to evaluate the onset of activation and to determine potential differences of activation latency between auditory and visual cortices and between these scanning methods. RESULTS: Temporal differences between visual and auditory areas ranged from 90-200 msec (root-mean-square (RMS) = 134 msec) and from -80 to 930 msec (RMS = 604 msec) in perfusion and BOLD measurements, respectively. The temporal variability detected with BOLD sequences was larger between subjects and was significantly greater than that in the perfusion response (P < 0.04). The measured time to half maximum (TTHM) values for perfusion imaging (visual, 3260 +/- 710 msec; auditory, 3130 +/ 700 msec) were earlier than those in BOLD responses (visual, 3770 +/- 430 msec; auditory, 3360 +/- 460 msec). CONCLUSION: The greater temporal variability between brain areas detected with BOLD could result from differences in the venous contributions to the signal. The results suggest that perfusion methods may provide more accurate timing information of neuronal activities than BOLD based imaging. PMID- 15666408 TI - The role of edema and demyelination in chronic T1 black holes: a quantitative magnetization transfer study. AB - PURPOSE: To use quantitative magnetization transfer imaging (qMTI) in an investigation of T1-weighted hypointensity observed in clinical magnetic resonance imaging (MRI) scans of multiple sclerosis (MS) patients, which has previously been proposed as a more specific indicator of tissue damage than the more commonly detected T2 hyperintensity. MATERIALS AND METHODS: A cross sectional study of 10 MS patients was performed using qMTI. A total of 60 MTI measurements were collected in each patient at a resolution of 2 x 2 x 7 mm, over a range of saturation pulses. The observed T1 and T2 were also measured. qMT model parameters were estimated using a voxel-by-voxel fit. RESULTS: A total of 65 T2-hyperintense lesions were identified; 53 were also T1 hypointense. In these black holes, the qMTI-derived semisolid pool fraction F correlated negatively with T(1,obs) (r2 = 0.76; P < 0.0001). The water pool absolute size (PDf) showed a weaker correlation with T(1,obs) (positive, r2 = 0.53; P < 0.0001). The magnetization transfer ratio (MTR) showed a similarly strong correlation with F and a weaker correlation with PDf (r2 = 0.18; P < 0.04). CONCLUSION: T1 increases in chronic black holes strongly correlated with the decline in semisolid pool size, and somewhat less to the confounding effect of edema. MTR was less sensitive than T(1,obs) to liquid pool changes associated with edema. PMID- 15666410 TI - Quantification of the kinetics and thermodynamics of protein adsorption using atomic force microscopy. AB - Both in situ and ex situ methods for quantifying area fraction coverage of protein on a surface using atomic force microscopy were developed. The in situ method used a continuous fluid flow system to observe the kinetics of adsorption in real time. The ex situ method required immersing the sample in solution, drying the sample, and imaging in an ambient environment to obtain kinetic and isothermal data. These methods were developed using the plasma protein fibrinogen in a phosphate-buffered saline solution on grade IV muscovite mica and highly ordered pyrolytic graphite (HOPG) substrates. Kinetic and quasiisothermal data were obtained and a Langmuir model was fit to the data. An adsorption rate constant of 2.2 x 10(-4) mL . microg(-1)s(-1) and a desorption rate constant of 8.3 x 10(-5) s(-1) were found on an HOPG surface. Completely irreversible adsorption was found on the mica surface with an adsorption rate constant of 2.7 x 10(-4) mL . microg(-1)s(-1). Additionally, protein conformation and assembly orientation on these surfaces were documented where fibrinogen on HOPG formed a network-like structure, whereas fibrinogen on mica was more random. Also, nano topographical factors (ledges) were seen as sites of preferential adsorption. PMID- 15666411 TI - Human brain derived neurotrophic factor (BDNF) genes, splicing patterns, and assessments of associations with substance abuse and Parkinson's Disease. AB - Potential roles for variants in the human BDNF gene in human brain disorders are supported by findings that include: (a) influences that this trophic factor can exert on important neurons, brain regions, and neurotransmitter systems, (b) changes in BDNF expression that follow altered neuronal activity and drug treatments, and (c) linkages or associations between genetic markers in or near BDNF and human traits and disorders that include depression, schizophrenia, addictions, and Parkinson's disease. We now report assembly of more than 70 kb of BDNF genomic sequence, delineation of 7 noncoding and 1 coding human BDNF exons, elucidation of BDNF transcripts that are initiated at several alternative promoters, identification of BDNF mRNA splicing patterns, elucidation of novel sequences that could contribute to activity-dependent BDNF mRNA transcription, targeting and/or translation, elucidation of tissue-specific and brain-region specific use of the alternative human BDNF promoters and splicing patterns, identification of single nucleotide polymorphism (SNP), and simple sequence length polymorphism (SSLP) BDNF genomic variants and identification of patterns of restricted haplotype diversity at the BDNF locus. We also identified type 2 BDNF-locus transcripts that are coded by a novel gene that is overlapped with type 1 BDNF gene and transcribed in reverse orientation with several alternative splicing isoforms. Association studies of BDNF variants reveal no associations with Parkinson's disease. Comparisons between substance abusers and controls reveal modest associations. These findings increase interest in this diverse human gene. PMID- 15666413 TI - Highly reactive, general, and long-lived catalysts for coupling heteroaryl and aryl chlorides with primary nitrogen nucleophiles. PMID- 15666412 TI - Bromoallenes as allyl dication equivalents in the absence of palladium(0): synthesis of bicyclic sulfamides by tandem cyclization of bromoallenes. PMID- 15666414 TI - Stereocontrolled total synthesis of (-)-aurisides A and B. PMID- 15666415 TI - Easy access to aryl- and heteroaryl-annulated[a]carbazoles by the indium catalyzed reaction of 2-arylindoles with propargyl ethers. PMID- 15666416 TI - Enantioselective copper-catalyzed conjugate addition to trisubstituted cyclohexenones: construction of stereogenic quaternary centers. PMID- 15666417 TI - Highly enantioselective epoxidation of 2,4-diarylenones by using dimeric cinchona phase-transfer catalysts: enhancement of enantioselectivity by surfactants. PMID- 15666418 TI - Highly rigid diphosphane ligands with a large dihedral angle based on a chiral spirobifluorene backbone. PMID- 15666420 TI - Direct, highly enantioselective pyrrolidine sulfonamide catalyzed Michael addition of aldehydes to nitrostyrenes. PMID- 15666419 TI - Recognizing a single base in an individual DNA strand: a step toward DNA sequencing in nanopores. PMID- 15666421 TI - Aromatic boron wheels with more than one carbon atom in the center: C2B8, C3B9(3+), and C5B11+. PMID- 15666423 TI - Alkyne carbonylation by radicals: tin-radical-catalyzed synthesis of alpha methylene amides from 1-alkynes, carbon monoxide, and amines. PMID- 15666422 TI - cis-Bromination of alkynes without cationic intermediates. PMID- 15666424 TI - Cognitive- and movement-related potentials recorded in the human basal ganglia. AB - Sources of potentials evoked by cognitive processing of sensory and motor activities were studied in 9 epilepsy surgery candidates with electrodes implanted in the basal ganglia (BG), mostly in the putamen. Several contacts were also located in the pallidum and the caudate. The recorded potentials were related to a variety of cognitive and motor activities (attentional, decisional, time estimation, sensory processing, motor preparation, and so on). In five different tests, we recorded P3-like potentials evoked by auditory and visual stimuli and sustained potential shifts in the Bereitschaftspotential and Contingent Negative Variation protocols. All of the studied potentials were generated in the BG. They were recorded from all over the putamen. Various potentials on the same lead or nearby contacts were recorded. A functional topography in the BG was not displayed. We presume that the cognitive processes we studied were produced in clusters of neurons that are organized in the basal ganglia differently than the known functional organization, e.g., of motor functions. The basal ganglia, specifically the striatum, may play an integrative role in cognitive information processing, in motor as well as in nonmotor tasks. This role seems to be nonspecific in terms of stimulus modality and in terms of the cognitive context of the task. PMID- 15666425 TI - The architecture of protein-ligand binding sites revealed through template assisted intramolecular peptide-peptide interactions. PMID- 15666426 TI - A clue to the origin of the Bilateria? PMID- 15666427 TI - Partial withdrawal of levothyroxine to stimulate serum thyroglobulin for thyroid cancer monitoring. PMID- 15666428 TI - Marine-Lenhart syndrome. PMID- 15666430 TI - You can quit smoking. PMID- 15666429 TI - Hb F-Porto Torres [Agamma75(E19)Ile-->Thr, 136(H14)Ala-->Ser]: a novel variant of the Agamma chain having two substitutions, one being that of Hb F-Sardinia. AB - The abnormal Hb F-Porto Torres [Agamma75(E19)Ile-->Thr, 136(H14)Ala-->Ser] was observed during a cord blood survey for hemoglobinopathies in North Sardinia. This silent variant showed the same mobility as Hb F-Sardinia in isoelectric focusing (IEF) of the tetramers, whereas the abnormal globin chain was clearly separated by acid-urea-Triton polyacrylamide gel electrophoresis (AUT-PAGE) from the normal Ggamma- and Agamma-globin chains. Separation of the globin chains by reversed phase high performance liquid chromatography (HPLC) indicated the following percentages: Ggamma 68.4, Agamma 14.0, Xgamma 17.6, that strongly suggested the abnormal chain as being a variant of the Agamma-globin. Sequencing of the gamma-globin genes indicated that the mutated gene was in fact an Agamma with two nucleotide replacements, one being the ATA-->ACA (Ile-->Thr) at codon 75 (the so-called AgammaT of the rather common Hb F-Sardinia) and the second the GCA ->TCA (Ala-->Ser) at codon 136. This new variant is the seventh having the sequence of the AgammaT chain with an additional mutation so far described and the third characterized by gene sequencing. PMID- 15666432 TI - Yellow fever. PMID- 15666433 TI - Fluocinonide. PMID- 15666434 TI - Welcome to improving working lives. PMID- 15666435 TI - Caring for children with burn injuries. PMID- 15666436 TI - [Protection mechanism or risk factor. How much anxiety can the heart stand?]. PMID- 15666437 TI - [A scientific chip hunt]. PMID- 15666438 TI - [Endothelin receptor antagonist bosentan. Breakthrough in the treatment of pulmonary hypertension]. PMID- 15666440 TI - FMD exercise demonstrates 'substantial advances' in contingency planning...but Royal Society still has concerns. PMID- 15666441 TI - Healthy animals, safe food: the role of veterinary medicines. PMID- 15666442 TI - Pick the right IT product. PMID- 15666444 TI - Sustaining the sustainability in waste management. PMID- 15666443 TI - Reproducibility of urinary cadmium, alpha1-microglobulin, and beta2-microglobulin levels in health screening of the general population. AB - The present study examined whether levels of cadmium, and alphal- and beta2 microglobulin in urine (Cd-U, ac-MG-U, and beta2-MG-U, respectively) were reproducible in urine samples collected from the same subjects on multiple occasions. For this purpose, two databases on background exposure to cadmium in Japan-one from study I between 2000 and 2001 and the other from study II in 2002 were revisited to find 231 apparently healthy, nonpregnant, nonlactating adult women who participated in both studies and thus had provided two urine samples. The databases contained information on Cd-U, alphal,-MG-U, and beta2-MG-U, creatinine (CR), and specific gravity (SG) as well as smoking and other lifestyle factors. Of the 231 women, 195 who had never smoked were selected for the present analysis. Cd-U as well as alpha1-MG-U were reproducible (e.g., with correlation coefficients [r] between study I and II results of 0.4 to 0.6) when measured on two occasions 9 to 10 months apart. The r values were lower for beta2-MG-U (r0.3). Exclusion of urine samples with inadequate urine density(i.e., CR <0.5 or >3.0 g/L or SG <1.010 or >1.030) resulted in substantial improvement of the agreements between the two measures (e.g., r = 0.6 to 0.7 for Cd-U and alpha1-MG U). CR and SG correlated closely with each other, especially in low-density urine samples (r >0.9), and therefore the effects of CR and SG could not be evaluated separately. In the overall evaluation,single determination (i.e., without repeated urine sampling) of Cd-U and alpha1-MG-U should be acceptable, and it may also be acceptable for beta2-MG-U. Use of samples with adequate urine density rather than application of density correction to low-density urine samples in recommended. PMID- 15666445 TI - The contribution of waste management to the reduction of greenhouse gas emissions with applications in the city of Bucharest. AB - Waste management is a key process to protect the environment and conserve resources. The contribution of appropriate waste management measures to the reduction of greenhouse gas (GHG) emissions from the city of Bucharest was studied. An analysis of the distribution of waste flows into various treatment options was conducted using the material flows and stocks analysis (MFSA). An optimum scenario (i.e. municipal solid waste stream managed as: recycling of recoverable materials, 8%; incineration of combustibles, 60%; landfilling of non combustibles, 32%) was modelled to represent the future waste management in Bucharest with regard to its relevance towards the potential for GHG reduction. The results indicate that it can contribute by 5.5% to the reduction of the total amount of GHGs emitted from Bucharest. PMID- 15666446 TI - New tool for landfill location. AB - In the present paper a decision-making process for the potential location of new landfill areas with wide community participation and acceptance is suggested. The main scientific contribution of this work is the elaboration of an independent decision-making tool, which can be used in landfill site selection. Specifically, at a first level it acts as an intermediary between experts (i.e. engineers, technical advisors) and decision-makers (i.e. elected representatives, appointive advisors), helping decision-makers to use experts' knowledge. At a higher level, it acts as an independent processor of decision-makers judgments thereby giving a result that is in accordance with pre-chosen criteria. In this way, the local authorities can effectively participate in the decision-making process and avert the violation of possible agreements. Furthermore, the criteria (pre-selection - evaluation) and the methodology of multicriteria analysis for new landfill location are presented. PMID- 15666447 TI - Measured gas emissions from four landfills in south africa and some implications for landfill design and methane recovery in semi-arid climates. AB - The magnitude of annual global emissions of methane from municipal solid waste landfills without landfill gas control systems implies that these landfills are significant contributors to the atmospheric load of greenhouse gases. There have been a number of field studies undertaken internationally to measure actual fluxes of methane and carbon dioxide from landfills, with a view to corroborating modelled predictions of the contribution of landfills to the global greenhouse gas budget. The vast majority of these studies have been undertaken in more temperate climates and in developed countries. This paper reports a study of landfill gas emissions from four large landfills located in the semi-arid interior of South Africa. A static accumulation chamber was used and measurements were made at each site over a period of two to three days. The results were analysed by three different methods, all of them leading to the same general conclusion that landfill gas emission rates were lower than expected. A common conclusion based on results from all four sites was that capping of landfills in semi-arid climates with low permeability covers would probably significantly retard the already low rate of waste degradation and thus gas generation. While this may be regarded as advantageous in the short term, it cannot be relied upon in perpetuity as clayey landfill covers will inevitably desiccate and crack in a semiarid environment. In addition, reasonable after-care periods for such landfills are likely to extend well beyond the currently stipulated 30-year period, and efforts to encourage energy recovery from landfills may be hampered because gas generation rates decrease as the waste dries out under conditions of minimal recharge from precipitation. A landfill cover that allows small amounts of percolation of rainfall into the waste may therefore in fact be beneficial in semiarid climates, although care would need to be taken to carefully regulate this infiltration. PMID- 15666448 TI - Laboratory-scale measurements of N2O and CH4 emissions from hybrid poplars (Populus deltoides x Populus nigra). AB - The purpose of this study was to determine whether or not young hybrid poplar (Populus deltoides x Populus nigra) could transport landfill biogas internally from the root zone to the atmosphere, thereby acting as conduits for landfill gas release. Fluxes of methane (CH4) and nitrous oxide (N2O) from the seedlings to the atmosphere were measured under controlled conditions using dynamic flux chambers and a tunable diode laser trace gas analyser (TDLTGA). Nitrous oxide was emitted from the seedlings, but only when extremely high soil N2O concentrations were applied to the root zone. In contrast, no detectable emissions of CH4 were measured in a similar experimental trial. Visible plant morphological responses, characteristic of flood-tolerant trees attempting to cope with the negative effects of soil hypoxia, were observed during the CH4 experiments. Leaf chlorosis, leaf abscission and adventitious roots were all visible plant responses. In addition, seedling survival was observed to be highest in the biogas 'hot spot' areas of a local municipal solid waste landfill involved in this study. Based on the available literature, these observations suggest that CH4 can be transported internally by Populus deltoides x Populus nigra seedlings in trace amounts, although future research is required to fully test this hypothesis. PMID- 15666449 TI - A model to minimize joint total costs for industrial waste producers and waste management companies. AB - The model LINKopt is a mixed-integer, linear programming model for mid- and long term planning of waste management options on an inter-company level. There has been a large increase in the transportation of waste material in Germany, which has been attributed to the implementation of the European Directive 75/442/EEC on waste. Similar situations are expected to emerge in other European countries. The model LINKopt has been developed to determine a waste management system with minimal decision-relevant costs considering transportation, handling, storage and treatment of waste materials. The model can serve as a tool to evaluate various waste management strategies and to obtain the optimal combination of investment options. In addition to costs, ecological aspects are considered by determining the total mileage associated with the waste management system. The model has been applied to a German case study evaluating different investment options for a co operation between Daimler-Chrysler AG at Rastatt, its suppliers, and the waste management company SITA P+R GmbH. The results show that the installation of waste management facilities at the premises of the waste producer would lead to significant reductions in costs and transportation. PMID- 15666450 TI - Deposits on single use containers--a social cost-benefit analysis of the Danish deposit system for single use drink containers. AB - This study compares the social costs and environmental benefits of collecting single use drink containers through the Danish deposit system with the social costs and benefits of treating the containers as part of the municipal waste disposal system. It focuses on single use polyethylene terphthalate and glass bottles, and steel and aluminium cans. The social costs of handling these containers in the deposit system includes the costs of collection, sorting, and transportation, adjusted against the profit from selling the collected material for recycling. The social cost of incinerating these containers as municipal waste consists of the expenses for the collection, incineration of the containers and disposal of ashes. If there is any income from energy generation accompanying incineration, this is adjusted against costs. The main environmental effects related to both strategies are quantified, valuated and included in the assessment. The results of the analysis show that there are significant social costs compared to the benefits connected with the new deposit system. This is true for all four types of single use drink containers examined. All in all, Denmark bears a net social cost of 6.7 to 8.1 million Euros per year compared to a baseline of incineration with energy recovery. PMID- 15666451 TI - Transformation of nitrogen during vermicomposting of fly ash. AB - In view of environmental problems generated by large-scale production of fly ash, increasing attention is now being paid to the recycling fly ash as a source of plant nutrients in agriculture. However, the low amount of nitrogen in such materials forms a major constraint for such application. In the present study, the possibility of improving the N status in mixtures of fly ash and organic matter was investigated by adopting vermicomposting technology. Different combinations of fly ash and cow (Bos taurus) dung; namely fly ash alone, cow dung alone and fly ash + cow dung at 1:1, 1:3 and 3:1 ratios were incubated with and without epigeic earthworms (Eisenia fetida) for 50 days. The occurrence of N in different bio-available forms; namely easily mineralizable, NH4+ and NO3- tended to increase considerably in the series treated with earthworms. This behaviour was attributed primarily to increased microbiological activity in the vermicomposted samples and also to a considerable rise in the concentration of nitrogen-fixing bacteria in this series. Among the three combinations of vermicomposted fly ash and cow dung, the 1:1 mixture appeared to exhibit the highest availability of nitrogen. PMID- 15666452 TI - Biopurification of industrial waste gas from rubber regeneration. AB - By using the biopurifying technology, the organic waste gas in low concentrations emitted from the rubber-regeneration process was purified in this research. The result of the 100-day continuous running test of the industrial test device indicated that the purification efficiency of toluene in the rubber-regeneration waste gas could be maintained at about 90% for a long period of time and the treated waste gas could meet the China National Emission Standard. The cost of waste gas biotreatment was about 0.12-0.14% of rubber-regeneration production value of the factory. The biopurifying technology of waste gas displayed its excellent technical advancement and economic rationale. The following industrialized device was run continuously and passed the examination and acceptance by the local EPA. PMID- 15666453 TI - Smoking and alcohol drinking during pregnancy. The reliability of retrospective maternal self-reported information. AB - BACKGROUND: The reliability of retrospective maternal self-reported information regarding smoking and alcohol drinking during pregnancy seemed necessary to be checked. METHODS: Two groups of congenital abnormalities were selected for the study from the population based Hungarian Congenital Abnormality Registry. The prevalence of smoking and drinking during the study pregnancy was measured in the mothers of 81 cases affected with isolated orofacial cleft and 537 cases with congenital limb deficiencies, and in their matched control pairs without any defect by maternal self-reported retrospective information through a mailed structured questionnaire. In the second step the latter data were checked by an independent personal interview of fathers and other family members at the home visit or in our department using the same structured questionnaire. RESULTS: The family consensus indicated the low reliability of retrospective maternal self reported information concerning smoking in the mothers of cases but not in mothers of controls. However, the comparison of retrospective maternal self reported data and information of close relatives indicated a reported bias concerning the drinking of alcohol beverages during pregnancy in both controls and cases. CONCLUSIONS: Since retrospective maternal self-reported data had low reliability, therefore, data of smoking and alcohol drinking were not collected for the data set of the Hungarian Case-Control Surveillance of Congenital Abnormalities between 1980 and 1996. PMID- 15666454 TI - Effectiveness of school programs in tobacco control. AB - The authors reviewed published data dealing with the effectiveness of school programs in tobacco control. Most of the evaluated school programs showed at least partial effect including namely improved knowledge level, decreased prevalence of smoking initiation and continuation. Less successful was achieving of behavioural changes and social resistance. Effect of the school programs can be significantly amplified by combination with other interventions such as mass media campaigns, parent involvement and extracurricular activities. The main problem of the studies in this field is a relatively short follow-up time not allowing considering findings as relevant evidences for long-term effects of school programs. However, even assuming only time limited decrease of prevalence of smoking among intervened students, such temporary effect leads to the decrease of a lifetime cigarette exposure having beneficial health effects. Considering social, demographic and cultural aspects of the epidemiology of smoking habit, evidence based data in this field, relevant for Central and Eastern Countries, are required. Such situation calls for authentic trials and studies respecting specific conditions in these countries. PMID- 15666455 TI - The level of nickel in smoker's blood and urine. AB - General population is exposed to nickel from various sources. Smoking presents a significant form of exposure. The research was conducted in period 2000--2003 in Institute of Public Health in Nis. The samples of tobacco and cigarettes (127 samples) were both domestic and imported, and samples of biological material (123 blood samples and 147 urine samples) were taken from occupationally unexposed persons (smokers and non-smokers). The analyses were performed by electrothermal atomization technique, by Perkin Elmer AAS M-1100. The results obtained, revealed a high content of nickel in cigarettes (2.32-4.20 mg/kg) and in tobacco (2.20 4.91 mg/kg) regardless of the kind and the origin of tobacco. Nickel content in the blood of smokers (0.01-0.42 microg/l, median 0.07 microg/l) was higher than in the blood of non-smokers (0.01-0.26 microg/l, median 0.06 microg/l) although this difference was not statistically significant (p>0.05). In the urine of smokers (<0.01-8.20 microg/l, median 1.20 microg/l) there was a significantly higher concentration of nickel than in the urine of non-smokers (<0.01-4.60 microg/l, median 0.50 microg/l), p<0.05. The exposure of smokers to nickel through tobacco smoke was high regardless of the kind and the origin of tobacco and cigarettes. The content of nickel in tissue fluids established by biomonitoring shows that smokers can be far more exposed to this carcinogenic substance than non-smokers and that health risks for smokers are higher in this context. PMID- 15666456 TI - Tobacco industry efforts to erode tobacco advertising controls in Hungary. AB - OBJECTIVE: To review strategies of transnational tobacco companies (TTCs) at creating a favourable advertising environment for their products in Hungary, with special regard to efforts resulting in the liberalisation of tobacco advertising in 1997. METHOD: Analysis of internal tobacco industry documents relevant to Hungary available on the World Wide Web. Transcripts of speeches of members of the Parliament during the debate of the 1997 advertising act were also reviewed. CONCLUSIONS: The tobacco companies not only entered the Hungarian market by early participation in the privatisation of the former state tobacco monopoly, but also imported theirsophisticated marketing experiences. Evasion and violation of rules in force, creation of new partnerships, establishment and use of front groups, finding effective ways for influencing decision makers were all parts of a well orchestrated industry effort to avoid a strict marketing regulation for tobacco products. PMID- 15666458 TI - Epidemiology of aids defining conditions in Greece. AB - OBJECTIVE: To examine the secular trends of all AIDS opportunistic infections to occur first (OIs) in Greece, by year, by gender and by mode of transmission. METHODS: The study included all AIDS defining conditions reported among Greek residents diagnosed with AIDS from 1981 to June 2003 and notified to the Hellenic Centre of Infectious Diseases Control. The analysis of trends in AIDS defining conditions in Greece has been performed only for the period 1993--2003. RESULTS: From 1981 to the first six months of 2003, 2,394 AIDS cases, 2,361 adults and 33 children, have been reported. HIV wasting syndrome was the most frequent OI to occur first followed by PCP pneumonia and Kaposi sarcoma. The frequency at which OIs occurred first varied by sex. Kaposi sarcoma was more frequent in males while tuberculosis and oesophageal candidiasis were more frequent in females. The frequency at which OIs occurred first varied also by exposure mode. Kaposi sarcoma was more frequent among men who have sex with men but that was not the case for the remaining transmission categories. From 1993 to the first six months of 2003 a downward trend was noticed only for chronic simplex disease. Since the introduction of HAART, an increasing trend was noticed for CMV disease, recurrent pneumonia, oesophageal candidiasis, Burkitt and immunoblastic lymphoma. CONCLUSION: Further epidemiological studies are needed to assess the OIs trends in coming years in order to plan prevention strategies and future medical care needs. PMID- 15666457 TI - HIV infection and risk behaviour of commercial sex workers and intravenous drug users in Slovakia. AB - INTRODUCTION: Aim of the study was to determine risk behaviour and HIV prevalence among commercial sex workers (CSWs) and intravenous drug users (IDUs) in streets of Bratislava and B. Bystrica, SR. METHODS: HIV antibodies were tested from saliva using ELISA test. Anonymous questionnaire was completed. RESULTS: 121 persons (61 men and 60 women) were involved in the sociological study. Mean age of the participants was 21.9 years. 185.1% of subjects were from Bratislava. 108 participants were tested for the presence of HIV-antibodies, one was confirmed HIV-positive (0.82%). In the past 47.9% of participants and 22.3% of their partners were tested for the presence of HIV-antibodies. 10.8% of subjects proclaimed that they suffered from other sexually transmitted infection (STI) in the past HIV testing of participants significantly correlated with the testing for other STI (p<0.002) as well as with HBV/HCV (p<0.001). 58 participants were using tattooing (47.9%). 46.3% of all participants never used condoms with partners. 31.4% of respondents proclaimed disruption of condom during sexual intercourse. Significant correlation was found between testing of participants for other STI and usage of condoms with their partners (p<0.013). Women used condoms more often by sexual contacts with partners than men used condoms (p<0.094). They were also significantly more tested for other STI in the past (p<0.021) and they suffered from other STI more often than men (p<0.033). 26.5% of person--only women--were involved in commercial sex work. 93.5% of them were taking drugs as well, 21.8% suffered for other STI in the past They were working in sex business on average for 26 months. The average number of their clients per week was 12.3. CSWs used condoms more often with clients than with partners. 98.2% of all participants were taking drugs, 93% of them intravenously. 24.6% of IDUs always used new or their own needles and syringes, while 69.4% shared equipments with the other users. IDUs drug users used condoms significantly less often with their partners than did CSWs (p<0.006). CSWs were significantly more often tested for other STI (p<0.001) and they also more often suffered for other STI than IDUs (p<0.045). CONCLUSION: More effort should be done to decrease risk behaviour revealed in the groups of CSWs and drug users. PMID- 15666459 TI - Prevalence of self-reported myocardial infarction in a Greek sample: findings from a population-based study in an urban area (medical express 2002). AB - During the 1970s and 1980s, Greece was known as a country with low prevalence and incidence of coronary heart disease, compared to Western populations. However, during the past decades, the Greek population has experienced marked but uneven socio-economic development, as well as change in lifestyle habits. We assessed the prevalence of self-reported myocardial infarction (MI) in a sample of the general population, aged 20-94 years. The overall prevalence of self-reported MI was 4.1% (6.3% in men and 1.9% in women). The age-adjusted prevalence was found to be 3.6%, showing a threefold increase compared to 1980s. Age, gender (male), low educational level, obesity/overweight, hypercholesterolemia, diabetes, hypertension, smoking and origin were strongly associated with prevalence of MI. Our findings indicate that the prevalence of MI increased dramatically during the past years, reflecting the change in lifestyle habits that have gradually given way to "Western"-type diets and a more sedentary lifestyle. Therefore, the need for urgent intervention is considered essential in order to prevent a further increase of disease burden. PMID- 15666460 TI - Functional ability after hip fracture among patients home-dwelling at the time of fracture. AB - The aim of the study was to assess the change in function and residential status four months after hip fracture in patients over 50 years of age and living independently at home at the time of fracture. All consecutive hip fracture patients treated at Oulu University Hospital during 1989--1999 were followed up. Data collection was done on standardized hip fracture forms, which were filled in at admission and at four months postoperatively. The forms included demographic data, place of residence, locomotor ability, use of walking aids, data on the fracture and its treatment, hospital stay, place of discharge and pain. At four months, 16.0% of the men and 14.3% of the women were permanently institutionalized. Preoperatively, 81.1% of the patients had been able to walk out of doors either alone or accompanied, while at 4 months, less than half of the patients (149 men, 391 women) were able to do so. Two thirds of the hip fracture patients had been able to walk without walking aids before the fracture, the corresponding proportion being one fifth at four months after the fracture. Cumulative mortality at 4 months was 9.9%, being higher among the male (15.5%, n=53) than the female patients (7.9%, n=75). The original study population was also divided into two subgroups operated at different period of time (1989--92 and 1993--99), the functional results seemed to improve with time. Hip fracture leads to the institutionalization of every seventh patient able to live at home at the time of fracture and impairs markedly one's functional capacity. To restore the pre-fracture status as well as possible and to reduce the burden of the consequences of hip fracture, it might be beneficial to evaluate and improve the existing rehabilitation system. PMID- 15666462 TI - Determination of n-methylcarbamates in foods. AB - The multiresidue method using multisolvent extraction, SPE cleanup of the extract, HPLC with the use of OPA post-column reaction and fluorescence detection for the determination of N-methylcarbamate pesticides in food products was used. A matrix solid phase dispersion method of the isolation and extraction of carbamates was alternatively applied. In the introductory study 44 items of the food basket for the Czech Republic were analysed. In the major part of the studied samples, the considerable part of which was culinary treated, the concentration of the target carbamates was below the limit of the used detection method. In the concentration range of 10-100 microg/kg in the analysed samples of the studied analytes, aldicarb and its metabolites, e.g. methomyl and methiocarb were being found most frequently. PMID- 15666461 TI - Homocysteine and its nutritional determinants in two ethnic groups of Slovakia. AB - Determinants of total homocysteine involve demographic (age, sex, ethnic origin), genetic (enzymatic defects of metabolic homocysteine pathways) and acquired factors (deficiency of B-group vitamins, state of health, lifestyle). Plasma levels of total homocysteine and serum levels of vitamin B12, folic acid, vitamin B6 were measured in adult apparently healthy Romany minority subjects (n=119) and compared with those levels in apparently healthy subjects of majority Slovak population (n=146). Mean homocysteine level was similar in both groups (9.92 micromol/l in the Romany vs. 9.61 micromol/l in majority group). Hyperhomocysteinemia was only observed in 3% of both ethnic probands. Vitamin B12 level was equal (301 micromol/l in the Romany and 311 micromol/ in majority group). Deficient levels were found in 4% of the Romany subjects and 9% of the majority subjects. Folic acid level was significantly lower in Romany group (11.3 nmol/l vs. 14.8 nmol/l) with deficiency in 42% of the Romany subjects vs. 28% in the majority subjects. This finding is a consequence of lower consumption of fruit, vegetables, pulses and whole grain products in the Romany group. Vitamin B6 deficiency was found in 68% of the Romany subjects and 40% of majority subjects. This vitamin is homocysteine determinant under excessive methionine intake (overnutrition with predominance of animal protein intake). As was demonstrated in a dietetic questionnaires, the Romany subjects are more frequent consumers of meat and eggs. This nutrition regime should indicate an increased homocysteine level under vitamin B6 deficiency. The results of normal homocysteine levels in the Romany population under condition of higher folic acid and vitamin B6 deficiencies, smoking and higher alcohol consumption may suggest a more effective homocysteine metabolism in relation to different ethnic origin. PMID- 15666463 TI - Emergency medical service in Poland--current state and a need of reform. AB - The article focuses on the question of reorganisation of Emergency Medical Service in Poland. First part of the paper contains a short description of a project of the Integrated Rescue System, which have been included in the National Emergency Medical Service Act enacted by Parliament in 2001. Considering to the fact, that implementation of this reform has been stopped after general elections in autumn 2001, in the second part of the paper some arguments supporting the postulate of urgent realisation of this project are discussed. The arguments refer to five spheres: epidemiological, social, political, legal and economical. The conclusions of the discussion are, that in every of those spheres negative consequences of blocking the reform may be observed. The final conclusion is, that reorganisation of ineffective Emergency Medical Service in Poland is still a challenge, which public authorities have to manage. PMID- 15666464 TI - Obesity in byzantine and in modern era from a public health perspective. AB - Obesity is one of today's most serious and amplified public health problems. Surprisingly, obesity constituted a health problem through the Byzantine Empire (3rd to 15th century AD) as well; the extent of the problem was then very much alike to that one seen in modem industrialized and developing countries of today. In this report we perform an historical throwback in Byzantine years in order to explore the link regarding the aspects of obesity in these years and in modem era. PMID- 15666465 TI - Theoretical and observational assessments of flare efficiencies. AB - Flaring of waste gases is a common practice in the processing of hydrocarbon (HC) materials. It is assumed that flaring achieves complete combustion with relatively innocuous byproducts such as CO2 and H2O. However, flaring is rarely successful in the attainment of complete combustion, because entrainment of air into the region of combusting gases restricts flame sizes to less than optimum values. The resulting flames are too small to dissipate the amount of heat associated with 100% combustion efficiency. Equations were employed to estimate flame lengths, areas, and volumes as functions of flare stack exit velocity, stoichiometric mixing ratio, and wind speed. Heats released as part of the combustion process were then estimated from a knowledge of the flame dimensions together with an assumed flame temperature of 1200 K. Combustion efficiencies were subsequently obtained by taking the ratio of estimated actual heat release values to those associated with 100% complete combustion. Results of the calculations showed that combustion efficiencies decreased rapidly as wind speed increased from 1 to 6 m/sec. As wind speeds increased beyond 6 m/sec, combustion efficiencies tended to level off at values between 10 and 15%. Propane and ethane tend to burn more efficiently than do methane or hydrogen sulfide because of their lower stoichiometric mixing ratios. Results of theoretical predictions were compared to nine values of local combustion efficiencies obtained as part of an observational study into flaring activity conducted by the Alberta Research Council (ARC). All values were obtained during wind speed conditions of less than 4 m/sec. There was generally good agreement between predicted and observed values. The mean and standard deviation of observed combustion efficiencies were 68 +/- 7%. Comparable predicted values were 69 +/- 7%. PMID- 15666466 TI - Carbon bed fires and the use of carbon canisters for air emissions control on fixed-roof tanks. AB - Fixed-roof tanks are used extensively at manufacturing, waste management, and other facilities to store or process liquids containing volatile organic compounds. Federal and state air standards require the control of organic air emissions from many of these tanks. A common practice used for some fixed-roof tanks that are required to use controls is to vent the tank through an activated carbon canister. When organic vapors are adsorbed on activated carbon, heat is released. Under certain conditions, the temperature of the carbon bed can increase to a level at which the carbon or organic vapors spontaneously ignite, starting a fire in the carbon bed. Bed fires in carbon canisters are not uncommon and can present a significant safety hazard at facilities if proper safety measures are not implemented. This article discusses how carbon adsorber bed fires occur and presents general guidance on safety measures for carbon canisters installed on fixed-roof tanks to reduce the likelihood of a carbon bed fire and to minimize the impact in the event of a fire. PMID- 15666467 TI - Microwave process for volatile organic compound abatement. AB - The CHA Corporation has completed the U.S. Air Force Phase II Small Business Innovation Research program to investigate the feasibility of using a novel microwave-based process for the removal and destruction of volatile organic compounds (VOCs) in effluents from noncombustion sources, such as paint booth ventilation streams. Removal of solvents by adsorption, followed by the regeneration of saturated granular activated carbon (GAC) by microwave energy, was achieved in a single fixed-bed reactor. Microwave regeneration of the fixed bed-saturated carbon restored the original GAC adsorption capacity. After 20 adsorption/regeneration cycles, the adsorption capacity dropped from 13.5 g methyl ethyl ketone (MEK)/100 g GAC to 12.5 g MEK/100 g GAC. During microwave regeneration of the GAC fixed bed, the concentrated desorbed paint solvent was oxidized by passing the solvent mixture through a fixed bed of an oxidation catalyst mixed with silicon carbide in a microwave reactor. A 98% oxidation efficiency was consistently achieved from the oxidation of VOCs in the microwave catalytic reactor. PMID- 15666468 TI - Combined PCDD/F destruction and particulate control in a baghouse: experience with a catalytic filter system at a medical waste incineration plant. AB - Phoenix Services, Inc., owns and operates the Baltimore Regional Medical Waste Incinerator in Baltimore, MD. New regulations for dioxins and furans imposed a limit that was considerably below historical emission levels. To determine a method to comply with the new dioxin/furan regulations, Phoenix Services performed trials with powdered activated carbon (PAC). Although the results with carbon were acceptable, Phoenix Services decided to replace their woven fiberglass filter bags with catalytic filters that simultaneously destroy dioxins and furans and collect particulate matter (PM). The catalytic filter system offered several advantages to Phoenix Services, including destruction of dioxins and furans instead of adsorption on carbon. The catalytic filters also offered a passive solution that did not require new carbon injection equipment. In January 2000, a campaign to measure dioxins/furans and PM was undertaken. The measurements allowed the catalytic filter system to be evaluated. Some of the key findings of this investigation are The dioxin/furan emission was less than 0.1 ng toxicity equivalents (TEQ)/Nm3 at 11% O2. This concentration is approximately 2 orders of magnitude less than historical averages and it is well below the new regulatory limits, for both existing and new sources of this type; the amount of dioxin/furans destroyed by the catalytic filters was approximately 1.73 ng TEQ/Nm3 at 11% O2; and the particulate emission was 12-17 times less than the regulatory limit. PMID- 15666469 TI - A cost-effective weighing chamber for particulate matter filters. AB - Particulate matter (PM) is a ubiquitous air pollutant that has been receiving increasing attention in recent years due in part to the association between PM and a number of adverse health outcomes, including mortality and increases in emergency room visits and respiratory symptoms, as well as exacerbation of asthma and decrements in lung function. As a result, the ability to accurately sample ambient PM has become important, both to researchers and to regulatory agencies. The federal reference method for the determination of fine PM as PM2.5 in the atmosphere recommends that particle-sampling filters be conditioned and weighed in an environment with constant temperature and relative humidity (RH). It is also recommended that vibration, electrostatic charges, and contamination of the filters from laboratory air be minimized to reduce variability in filter weight measurements. These controls have typically been maintained in small, environmentally controlled "cleanrooms." As an alternative to constructing an elaborate cleanroom, we have designed, and presented in this paper, an inexpensive weighing chamber to maintain the necessary level of humidity control. PMID- 15666471 TI - Biofiltration of a mixture of volatile organic emissions. AB - Air biofiltration is now under active consideration for the removal of the volatile organic compounds (VOCs) from polluted airstreams. To optimize this emerging environmental technology and to understand compound removal mechanisms, a biofilter packed with peat was developed to treat a complex mixture of VOCs: oxygenated, aromatic, and chlorinated compounds. The removal efficiency of this process was high. The maximum elimination capacity (ECmax) obtained was approximately 120 g VOCs/m3 peat/hr. Referring to each of the mixture's components, the ECmax showed the limits in terms of biodegradability of VOCs, especially for the halogenated compounds and xylene. A stratification of biodegradation was observed in the reactor. The oxygenated compounds were metabolized before the aromatic and halogenated ones. Two assumptions are suggested. There was a competition between bacterial communities. Different communities colonized the peat-based biofilter, one specialized for the elimination of oxygenated compounds, the others more specialized for elimination of aromatic and halogenated compounds. There was also substrate competition. Bacterial communities were the same over the height of the column, but the more easily biodegradable compounds were used first for the microorganism metabolism when they were present in the gaseous effluent. PMID- 15666470 TI - Indoor pollutant levels from the use of unvented natural gas fireplaces in Boulder, Colorado. AB - High CO and NO2 concentrations have been documented in homes with unvented combustion appliances, such as natural gas fireplaces. In addition, polycyclic aromatic hydrocarbons (PAH) are emitted from incomplete natural gas combustion. The acute health risks of CO and NO2 exposure have been well established for the general population and for certain high-risk groups, including infants, the elderly, and people with heart disease or asthma. Health effects from PAH exposure are less well known, but may include increased risk of cancer. We monitored CO emissions during the operation of unvented natural gas fireplaces in two residences in Boulder, CO, at various times between 1997 and 2000. During 1999, we expanded our tests to include measurements of NO2 and PAH. Results show significant pollutant accumulation indoors when the fireplaces were used for extended periods of time. In one case, CO concentrations greater than 100 ppm accumulated in under 2 hr of operation; a person at rest exposed for 10 hr to this environment would get a mild case of CO poisoning with an estimated 10% carboxyhemoglobin level. Appreciable NO2 concentrations were also detected, with a 4-hr time average reaching 0.36 ppm. Similar time-average total PAH concentrations reached 35 ng/m3. The results of this study provide preliminary insights to potential indoor air quality problems in homes operating unvented natural gas fireplaces in Boulder. PMID- 15666472 TI - Tetrachloroethylene emissions and exposure in dry cleaning. AB - Tetrachloroethylene (PCE) emissions and the exposure of workers in six commercial and three industrial dry-cleaning establishments that use dry-to-dry machines were determined. The personal samples and area samples [8-hr time-weighted average (TWA) and short-term exposure] were collected with charcoal tubes and passive monitors. The temporal variation of PCE concentration in the workplace air was monitored using a Fourier transform infrared analyzer (FTIR). The PCE emission rates were determined by multiplying the average PCE concentration in the room and the total airflow rate in the room. The PCE emissions were related to the cleaning rate in units of kg/hr. The operators' mean TWA exposure in commercial shops and industrial establishments was 28 (4.1 ppm) and 32 mg/m3 (4.6 ppm), and the pressers' exposure was 3.4 (0.5 ppm) and 7.7 mg/m3 (1.1 ppm), respectively. The customer service personnel had the lowest TWA exposure with a mean value of 0.8 mg/m3 (0.1 ppm). The highest peak concentration (2300 mg/m3; 334 ppm) was observed during cleaning of the lint and button trap, during which operation respirators were used. The PCE emission rates ranged from 4 to 118 g/hr corresponding to emission factors (mass of solvent evaporated per mass of cleaned cloths) of 0.3-3.6 g/kg. The workers' exposure to PCE was below the occupational limit values in the United States [according to the American Conference of Governmental Industrial Hygienists (ACGIH)] and in Finland. The outdoor PCE emissions were clearly below the limit values given in the European Union volatile organic compound (VOC) directive requirements. PMID- 15666473 TI - Flue gas desulfurization: the state of the art. AB - Coal-fired electricity-generating plants may use SO2 scrubbers to meet the requirements of Phase II of the Acid Rain SO2 Reduction Program. Additionally, the use of scrubbers can result in reduction of Hg and other emissions from combustion sources. It is timely, therefore, to examine the current status of SO2 scrubbing technologies. This paper presents a comprehensive review of the state of the art in flue gas desulfurization (FGD) technologies for coal-fired boilers. Data on worldwide FGD applications reveal that wet FGD technologies, and specifically wet limestone FGD, have been predominantly selected over other FGD technologies. However, lime spray drying (LSD) is being used at the majority of the plants employing dry FGD technologies. Additional review of the U.S. FGD technology applications that began operation in 1991 through 1995 reveals that FGD processes of choice recently in the United States have been wet limestone FGD, magnesium-enhanced lime (MEL), and LSD. Further, of the wet limestone processes, limestone forced oxidation (LSFO) has been used most often in recent applications. The SO2 removal performance of scrubbers has been reviewed. Data reflect that most wet limestone and LSD installations appear to be capable of approximately 90% SO2 removal. Advanced, state-of-the-art wet scrubbers can provide SO2 removal in excess of 95%. Costs associated with state-of-the-art applications of LSFO, MEL, and LSD technologies have been analyzed with appropriate cost models. Analyses indicate that the capital cost of an LSD system is lower than those of same capacity LSFO and MEL systems, reflective of the relatively less complex hardware used in LSD. Analyses also reflect that, based on total annualized cost and SO2 removal requirements: (1) plants up to approximately 250 MWe in size and firing low- to medium-sulfur coals (i.e., coals with a sulfur content of 2% or lower) may use LSD; and (2) plants larger than 250 MWe and firing medium- to high-sulfur coals (i.e., coals with a sulfur content of 2% or higher) may use either LSFO or MEL. PMID- 15666475 TI - [Physiopathological mechanisms in type 1 and type 2 diabetes]. AB - Type 1 and type 2 diabetes have different clinical presentations in their common forms. The absence of biological tests for diagnosis of type 2 diabetes, for reliable evaluation of insulin secretion and, beta-cell mass or for diagnosis of insulitis and the limited sensitivity of antibody tests in type 1 diabetes explain the difficulty of classifying a large number of cases of diabetes and the identification of questionable clinical entities: LADA (Latent Autoimmune Diabetes in Adult), non-insulin dependant diabetes in coloured people. Hyperglycaemia may also be observed in genetic diseases affecting insulin secretion (MODY, mutations of mitochondrial DNA), complex genetic diseases, pancreatic, endocrine and iatrogenic affections. Type 1 diabetes is characterised by the destruction of, beta-cells while type 2 diabetes combines abnormal insulin secretion and insulin resistance. The reduction of the beta-cell mass in type 2 diabetes remains controversial. These different conditions are examples of multifactorial diseases during which several physiopathological processes are disturbed in association with the expression of multiple genes, of normal structure and function, but variants of which copy for a number of partial phenotypes the association of which causes the disease. Some hypotheses suggest that type 2 diabetes is the consequence of the selection of variants of genes involved in energetic storing and responsible for defence against infection and of the recent very rapid changes in the environment. PMID- 15666474 TI - Ethylbenzene removal in a multiple-stage biofilter. AB - In practice, biofilters are often conceived as entire, single-unit systems. However, the activity of a biofilter varies greatly over its depth. For a given period, each stage of the biofilter dominates ethylbenzene removal. Ethylbenzene was continuously removed in a mixed-medium biofilter. The overall removal efficiency of the ethylbenzene ranged from 70% to greater than 99%. In the upflow biofilter, the most dominant ethylbenzene degrading stage shifted consecutively from the bottom to the top of the reactor. Average water content throughout the biofilter media was relatively consistent. However, the water content of each stage fluctuated dramatically and was correlated with the ethylbenzene removal rate. Without any water addition, the biofilter was operated for 62 days above the target removal efficiency of 80%. A 9-month slow-release fertilizer, mixed with composting media, was an effective way to eliminate the nutrient deficiency in the biofilter operation. PMID- 15666476 TI - [Physiopathology of atherosclerosis in diabetics]. AB - Vascular diseases, especially atherosclerosis, are the main cause of morbidity and mortality in diabetics. Diabetes greatly increases the risk of developing coronary heart disease, cerebral vascular accident and lower limb arteritis. The physiopathology of vascular disease in the diabetic patient involves endothelial and smooth muscle cell abnormalities. Metabolic disturbances which are characteristic of diabetes, such as hyperglycaemia or AGE accumulation, contribute to endothelial dysfunction and augment the inflammatory response at the vascular level. Atherosclerotic plaques in diabetics are more inflammatory than in non-diabetics, with an accumulation of macrophages and T lymphocytes, a larger lipid core and the presence of a greater number of macrophages and smooth muscle cells in apoptosis, which makes them more vulnerable. PMID- 15666477 TI - [Optimising the equilibrium of diabetes and atheromatous progression: myth or reality?]. AB - Is chronic hyperglycaemia, which defines diabetes of whatever type, a simple marker of cardiovascular risk, with no effect on atheroma independent of the other factors with which it is associated such as dyslipidaelia or hypertension? Whatever the case, its treatment, although fundamental for preventing the progression of microangiopathy and neuropathy, has another significance regarding the progression of atheromatous disease. Chronic hyperglycaemia acts as an independent factor for the genesis and progression of atheromatous disease and the beneficial effect of treating it aggressively could add to that obtained by treatment of the classic atherogenic factors such as hypercholesterolaemia or hypertension. This controversy, renewed by the interpretation of the UKPDS results, still remains. There is also a 2nd question: how does chronic hyperglycaemia, which is generally well evaluated in the medium term by HbA1c, overlap with other physiological situations? In particular, does post-prandial hyperglycaemia have greater atherogenic effects than the same level of fasting hyperglycaemia? Whatever the case, greater attention must be paid to post prandial glycaemic control, which has been rather neglected up to now in favour of fasting glycaemic control, taken to mean overall glycaemic control. PMID- 15666478 TI - [Specificity of the management of atheromatous disease in diabetic renal failure]. AB - Atheromatous disease is particularly common and severe in diabetics with renal failure. The prognosis depends on specific therapeutic measures: in this article, the authors will limit the discussion to the management of coronary artery disease. The cardiovascular risk factors must be treated intensively and corrected. The outlook of these patients also depends on the diagnosis of silent ischaemia with a stress test every two years and the prevention of acute renal failure after coronary angiography. Myocardial revascularisation improves the prognosis of high risk coronary patients. The choice of mode of revascularisation depends on the anatomy of the coronary lesions, the surgical risk and the presence of associated valve disease. The results of coronary angioplasty have been improved by the optimisation of anti-thrombotic treatment and the use of active stents. Despite revascularisation, short and medium-term mortality remains much higher than that of non-diabetics with normal renal function. Non-invasive investigations for residual ischaemia are justified 6 months after angioplasty for diagnosing restenosis and, thereafter, every year to detect progression of the atherosclerosis. PMID- 15666479 TI - [Cerebral vascular pathology in diabetes]. AB - Diabetes is a major risk factor for cerebral vascular accidents (CVA) and the prevalence of diabetes in the population of patients presenting with CVA varies from 13 to 36% in studies. On the other hand, it is not a risk factor for haemorrhagic CVA. The two principal causes of CVA in diabetic patients are small artery disease and atherosclerosis of cervical and intracranial arteries. Diabetics differ from non-diabetics with a higher prevalence of hypertension. The data from the literature suggest a worse prognosis for CVA in diabetics. While the initial stroke severity seems comparable, the acute and especially late mortality is increased in diabetics. In those who survive, diabetics have a slower recovery and greater handicap at 3 months post CVA. Management in the acute phase of cerebral infarction is identical in the diabetic and non-diabetic. In secondary prevention, carotid surgery is indicated for stenoses of more than 50%; treatment of risk factors is the same. On the other hand, clopidogrel seems to be of more benefit than aspirin in diabetics. In conclusion, although there are aetiological and prognostic characteristics of diabetics presenting with a CVA, the treatment and its benefits in the acute phase and in secondary prevention are comparable. PMID- 15666480 TI - [Critical limb ischaemia: endovascular treatment in diabetic patients?]. AB - Endovascular treatment has an increasing role in the treatment of patients with critical limb ischemia, particularly in diabetic patients with a majority of infrapopliteal lesions. The aim of the procedure is to obtain a "straight-line flow to the foot" by treating all the significant stenoses and short occlusions that impair distal vascularization. Stents are indicated when there is a suboptimal results following balloon angioplasty (recoil or dissection). Restenosis rate after primary stenting for long lesion is high. Angioplasty is a safe and effective procedure, allowing limb salvage rate in a majority of the cases with a low mortality and morbidity rate. PMID- 15666481 TI - [Strategy of investigation of coronary artery disease in diabetes: from screening to suspicion of acute coronary syndromes]. AB - Coronary artery disease is a common and serious condition in diabetes and the prognosis of the diabetic without a history of cardiovascular disease is either the same or nearly as serious as that of a non-diabetic patient with a history of coronary disease. This is particularly true in women. The prognosis is even worse in the presence of silent myocardial ischaemia. Conversely, anti-ischaemic and anti-thrombotic therapy and myocardial revascularisation of most severely affected patients are effective. This justifies the recent recommendations (as those of the working group of the French Society of Cardiology and the ALFEDIAM) for the diagnosis of coronary artery disease in diabetes, even in asymptomatic patients. This is a two stage process: --First, the identification of patients who should be screened for ischaemia, diabetics with a priori an intermediate or high risk of the presence of CAD, with respect to the presence of markers easily identified on initial examination, like the presence of clinical macroangiopathy (femoral, carotid), of renal disease or ECG changes or the presence of several classical risk factors; --The second stage is the demonstration of myocardial ischaemia in patients identified to be at risk. This article reviews the advantages and limits of the tests available: ECG stress test, myocardial perfusion imaging on effort or under dipyridamole, stress echocardiography. Coronary angiography in asymptomatic patients is only recommended in the presence of significant ischaemia or with a poor prognosis (affecting over 20% of the myocardium or several myocardial territories). This should precede a myocardial revascularisation procedure. The prescription of coronary angiography may be more direct in some symptomatic patients. PMID- 15666482 TI - [Predictive value of glycemia in acute coronary syndromes]. AB - Intravenous insulin therapy is used in diabetic patients at the acute phase of coronary syndrome (ACS). However, hyperglycemia in diabetic patients is a powerful predictive factor for patient outcome as it is associated with a doubling of in-hospital mortality and poor long-term prognosis. Recent studies involving non-diabetic patients show that even mild hyperglycemia in the setting of ACS is also a predictive factor of in-hospital mortality. Moreover, the new entity called impaired fasting glucose (IFG) (6.1 to 7 mmol/L) is not only an independent factor of mortality for coronary patients, but very recently has also been associated with a doubling of the risk of in-hospital mortality in the setting of ACS. Admission as well as follow-up glycaemia are fundamental parameters in ACS on the one hand for their prognostic value, and on the other end as a diagnostic tool in determining the presence of diabetes or IFG. PMID- 15666483 TI - [Does diabetes affect the choice of pharmaceutical treatment in coronary artery disease?]. AB - Coronary artery disease is the principal cause of death in diabetic patients. The choice of pharmaceutical treatment used in diabetic patients with coronary artery disease should be based on that of non-diabetic coronary patients (BASIC). However, diabetes has its own specificities. First of all, dietetic measures and physical exercise should be continually encouraged. The questions about the basic treatment concern mainly the choice of oral platelet antiaggregants and the need to associate these drugs systematically and definitively. With regards to the management of acute coronary syndromes, the important point is the improved short and long-term benefits of GP IIb/IIIa on the survival of diabetic coronary patients, especially when angioplasty is performed. Finally, intravenous insulin therapy is promising in the initial phase of treatment of acute coronary syndromes with better defined blood glucose objectives. PMID- 15666484 TI - [Does diabetes affect therapy in coronary patients? Revascularisation]. AB - Techniques of myocardial revascularisation are very commonly employed in diabetic patients. In unstable angina, the reported data suggests a very significant benefit of an invasive strategy for the diabetic patient. In stable angina, subgroups analysis of randomised clinical trials carried out over ten years ago showed a better outcome after surgery in cases of multivessel disease. What has been the impact of technical advances archived since then? Active stents and antiplatelet drugs have considerably improved the outcome of diabetics treated by angioplasty. Cardiac surgery has also made considerable progress with the systematic use of arterial grafts. New randomised clinical trials are underway to determine the modern options of myocardial revascularisation for diabetic patients. PMID- 15666485 TI - [Insulin in acute coronary syndromes]. AB - There is a state of major stress during acute coronary syndromes with insulin resistance which results in cardiac cellular metabolism becoming almost completely dependant on fatty acids. This increases cellular oxygen demand with consequent cellular necrosis and apoptosis. Under normal conditions, the metabolism in type 2 diabetes is almost exclusively based on fatty acids. During ischaemia, these phenomena are accentuated which partially explains the higher morbid-mortality after myocardial infarction. It has been demonstrated that insulin administration (GIK) creates conditions for an "optimal milieu" protecting the myocardium during the ischaemic and reperfusion phases by the effects of glucose, the anti-inflammatory action of insulin and its action on NO synthesis. Its efficacy in reduction in post-infarction mortality has been demonstrated in type 2 diabetes in the DIGAMI trial and also in all coronary patients and after cardiac surgery, especially in the ECLA, DUTCH, Texas Heart Institute trials. The results were convergent with respect to insulin dosage, blood sugar levels and volumes of solute, also in the context of cardiac failure. However, in type 2 diabetes, the GIK protocol remains underused in our acute cardiac units and insulin therapy is often prescribed in conditions which do not provide a real benefit. Very few centres use the GIK protocol in non-diabetics in acute coronary syndromes with a risk of loss of benefit for these patients. Therapeutic protocols which can be used by paramedical personnel should be developed. PMID- 15666486 TI - [Does coffee consumption represent a coronary risk factor?]. AB - Aim of this paper is to critically review the evidence available regarding the relationship between coffee consumption and coronary risk. Large prospective studies do not support the hypothesis that moderate (< 5 cups of coffee a day) caffeine consumption significantly increases the risk of coronary heart disease. Data on higher coffee consumption are scanty and do not allow the acquisition of established conclusions, even if they suggest a statistically significant trend in coronary risk with increasing dose. PMID- 15666487 TI - [Diagnostic efficiency on digital snapshots of the standard radiology imaging]. AB - The authors had experienced the telediagnosis on digital snapshots of standard radiology imaging (chest, abdomen, and bones), sent by e-mailing, to support the medical doctors working in remote areas of developing countries. In order to validate the overall procedure, the authors have set up a simulating model and estimated some parameters of accuracy of the diagnosis on digital snapshots against the golden standard of the diagnosis by direct look. The study concerned the standard X-ray tests of one hundred randomly-selected patients out of a hospital archive. Four years later the diagnosis by direct look, the team of radiologists carried out the blind cross check on the digital snapshots of the radiograms and stated their second diagnosis. Sensibility, specificity, predictive value of positives, predictive value of negatives and efficiency of the whole series have been 83.0, 95.1, 96.1, 79.6 and 88.0%. By breaking up the series by apparatus, the skeleton test shows similar data of the whole series. The chest test shows a specificity and predictive value of positives of 100.0%. Although the number of cases is low, the abdomen test apparently shows a sensibility and predictive value of negatives as high as 100%, but a lower specificity and predictive value of negatives (85.7 and 87.5%). Though this data is supportive to the validation of the procedure, even better results are supposedly achieved by increasing the quality of the snapshots and by improving the skills of using the software. PMID- 15666488 TI - [Falls in hospitalized elderly patients: incidence and risk factors]. AB - Aim of the study was to determine the incidence and risk factors for falls in hospitalized elderly patients. In a prospective observational study we investigated the characteristics of 620 elderly patients. We examined demographic variables, cognitive and functional status, main pathologies and drugs. Balance and gait were evaluated by Tinetti's scale. Seventy patients (11.3%) fell during hospitalization. The incidence rate of first falls was 5.2 per 1000 patient-days. Five independent variables were significantly and independently associated to falls: age, balance impairment, acute cognitive impairment, diabetes, and use of tricyclic antidepressants. Falls are common in hospitalized elderly patients and risk factors identification can be useful for a targeted multiple intervention falls prevention programme. PMID- 15666489 TI - [Turnover of bone metabolism and prevalence of vertebral fractures in adrenal incidentalomas]. AB - Aim of the study was to evaluate, in a homogeneous group of patients with adrenal incidentalomas (AI), calcium-phosphorus metabolism alterations, bone mineral density (BMD) and the prevalence of vertebral fractures. We selected 46 patients with adrenal incidentalomas (26M, 20F; age: M = 61 +/- 14, F = 65 +/- 10 years, BMI: M = 26.2 +/- 4 Kg/m2, F = 28.8 +/- 4 Kg/m2) compared with 84 normal subjects (NS) (44M, 40F; age: M = 60 +/- 10, F = 62 +/- 8 years; BMI: M = 27 +/- 2 Kg/m2, F = 28.1 +/- 4.5 Kg/m2). In all subjects we estimated calcium-phosphorus parameters. Our results showed that in 46 patients with AI there were a significant reduction of BMD-LS (0.915 +/- 0.176 g/cm2; p = 0.01) and of BMD-FN (0.710 +/- 0.129; p = 0.034) respect to those of NS (respectively: 0.994 +/- 0.14 9 g/cm2; 0.758 +/- 0.117 g/cm2). BMD-LS of the 20 women with AI (0.864 +/- 0.157 g/cm2, p = 0.01) was significantly reduced compared with the 40 female normal subjects (0.904 +/- 0.148 g/cm2); BMD-LS of the 26 men with AI (0.967 +/- 0.187 g/cm2; p = 0.048) was significantly reduced compared with the male normal subjects (1.048 +/- 0.133 r/cm2; p = 0.048). The MXA showed vertebral fractures in 15 (75%) of 20 patients with AI (2 patients were osteoporotic, 9 osteopenic and 4 normal at the MXA scans). In 20 women with AI, compared with female NS, we found a significant reduction of serum 25 OH D3 (p = 0.024) levels and a significant increase of plasma i-PTH (P = 0.04) value; and we found a negative correlation between plasma i-PTH and 25 OH D3 values (r = -0.451; p > 0.045). In conclusion, we demonstrated that patients with non-functioning adrenal incidentalomas present calcium-phosphorus metabolism alterations, associated at a reduction of BMD and an increase of vertebral fractures. PMID- 15666490 TI - [Correlation between subjective well-being and pharmacological therapy in patients with psychotic disorders]. AB - AIMS: The present study aims to investigate whether exists a meaningful relation between quality of life and subjective well being with regard to the pharmacological treatment (antipsychotic typical versus atypical) in a sample of people with psychotic disorders integrated in a Community Residential Rehabilitation Centre; to examine whether the different antipsychotic treatment is correlated to a different answer to the psychosocial rehabilitation intervention in terms of significant improvement in the positive and negative symptomatology, subjective well-being and quality of life. METHOD: All patients, who suffer from schizophrenia and schizoaffective disorder according to DSM-IV criteria, treated with antipsychotic and stabilized from at least one month, were enrolled in the study. RESULTS: 32 patients have participated in the study: 22 patients treated with atypical drugs and 10 with typical. The analysis of the collected data didn't show any significant statistical difference at baseline with regard to symptomatology, subjective well-being and quality of life. From the statistical analysis of the data to the endpoint, after a month of psychosocial rehabilitation, we found a statistically meaningful improvement in all the areas inquired in the group of the patients dealt with antipsychotic atypical drugs. CONCLUSION: The results confirm that the atypical antipsychotics are more effective, than the typical, to improve symptomatology, subjective well being and quality of life of psychiatric patients. PMID- 15666491 TI - [Low incidence of streptococcal pharyngitis in young adults]. PMID- 15666492 TI - [New insights on the molecular mechanisms of type-1 angiotensin II receptor blockers and their contribution to atherosclerotic plaque stabilization]. AB - Clinical trials have demonstrated that agents inhibiting the angiotensin II pathway confer benefit beyond the reduction of blood pressure alone. However, the molecular mechanism underlying this effect has yet to be investigated. Recently, we have demonstrated enhanced expression of inducible COX and PGE synthase (COX 2/mPGES-1) in human symptomatic plaques, and provided evidence that it is associated with plaque rupture induced by metalloproteinases (MMPs), proteolytic enzymes of matrix structural components. In a recent study, we hypothesized that angiotensin II could promote plaque instability through induction of the COX 2/mPGES-1 pathway. We analyzed atherosclerotic plaques from symptomatic patients who were randomized to treatment with irbesartan (300 mg/die) or chlortalidone (50 mg/die) for 4 months before endarterectomy. We found that plaques from the irbesartan group had reduced inflammatory infiltration, less immunoreactivity for COX-2/mPGES-1 and MMPs, reduced gelatinolytic activity and increased collagen content. It is worth noting that COX-2/mPGES-1 inhibition was observed after incubation in vitro with irbesartan but not with the selective AT2 blockade PD123,319. These findings suggest that AT1 receptor blockers could represent a novel form of therapy for plaque stabilization in patients with atherosclerotic disease. PMID- 15666493 TI - [Infective endocarditis. Guidelines for diagnosis and treatment]. AB - After careful review of evidence-based literature, clinical and laboratory criteria for diagnosis of bacterial and fungal endocarditis are examined. The choice criteria for therapy of bacterial endocarditis, both empiric and directed against a specific pathogen, are reviewed, on the basis of the clinical and epidemiological context (prosthetic or native valve, left or right heart, drug addiction). Different treatment options are proposed, based on results of antibiotic resistance testing. Indications and contraindications for a parenteral home treatment and those for surgical treatment are examined, also according to the results of ultrasonography. PMID- 15666494 TI - [A mere strip of skin]. PMID- 15666495 TI - A chronology of dental education in the United States. AB - Formal dental education in the United States began in 1840 with the opening of the Baltimore College of Dental Surgery. Other dental schools slowly began to emerge, gradually displacing the traditional preceptorship method of training for dentistry. The period of the late 1800s saw a surge in the number of dental colleges in the United States. These schools were largely proprietary in nature, meaning that they were not affiliated with major universities, were private, and were of a commercial nature and, usually, established to benefit their owners. As the trend toward affiliation of dental schools with universities gained impetus at the beginning of the 20th Century, and with the establishment of the Dental Educational Council of America, the trend continued. Finally, the Carnegie Foundation for the Advancement of Teaching formed a committee, under the direction of Dr. William J. Gies, to study the entire system of dental education in America. Out of this landmark study came the strong recommendation that all dental schools become affiliated with major universities. Shortly after that study, in the early 1930s, the last proprietary school was abolished. This article traces the histories of dental schools, past and present, in the United States. PMID- 15666496 TI - Dentistry on stamps. PMID- 15666497 TI - Dr. Lytle Adams' incendiary "bat bomb" of World War II. AB - On December 7, 1941, a 60-year old dentist from Irwin, Pennsylvania, Dr. Lytle S. Adams, was driving home from a vacation at Carlsbad Caverns in New Mexico. Hours earlier, he had been gripped with amazement as he witnessed millions of bats exiting the caves of Carlsbad. Listening to his car radio on his return trip, he was shocked to hear that Japan had just attacked Pearl Harbor. Dr. Adams, outraged over this travesty, began to mentally construct a plan for U.S. retaliation. As his thoughts returned to the countless bats that had awed him, he formed a tentative plan: millions of these small, flying mammals could be connected to tiny, time-fused incendiary bombs, and then released to land on the flimsily constructed structures which dotted the cities of Japan. Within a few minutes, the bombs would explode and enflame the entire urban areas. He postulated that these immeasurable numbers of fires, spreading their devastation over such vast areas within Japanese cities would result in the enemy's speedy surrender. This article documents the futile efforts of Dr. Adams, his team and the U.S. government to develop and employ an effective, incendiary bat bomb. The recently developed atom bomb, a far more deadly weapon was used in its place. PMID- 15666498 TI - Dentistry postcards XXVII. "Do you need filling madam"? PMID- 15666499 TI - Gleanings about dentistry from the world of literature (thirty-fourth in a series). PMID- 15666500 TI - The Stromeyer hook--life and work of the man behind the eponym. AB - Georg Friedrich Stromeyer (1804-1876) is generally known as one of the founders of orthopedics and orthopedic surgery and a pioneer of modern military medicine. Every oral maxillofacial surgeon in Germany knows him because of the "Stromeyer hook" for the elevation of zygomatic arch fractures. The present paper focuses on this particular aspect of maxillofacial surgery as well as on Stromeyer's biography. PMID- 15666501 TI - Dental collections XXI. PMID- 15666503 TI - 19th century dentistry advertising trade cards. PMID- 15666502 TI - Dental writings in a medical self-help book of 1650. AB - Only one copy exists of the very first medical self-help book written in the Yiddish language. The author, a well respected. barber-surgeon in Prague, states that he wrote the book for those too poor to consult a physician. Physicians were not as knowledgable or experienced as the barber-surgeons who were the principal providers of medical care for the masses. The book follows the format of discussing illnesses from head to toe, and includes numerous prescriptions for toothache, loose teeth, foul mouth and sore tongue. We derive an interesting picture of what dental treatment was like in the mid-seventeenth century. PMID- 15666505 TI - Extinguishing the embers: medical-waste incineration and the community. PMID- 15666504 TI - Dental, oral and facial limericks of yesterday and today. AB - The limerick is a popular form of light and poetic verse which was originally recited and sung in the local inns and taverns of Ireland and England. It may be anecdotal, nonsensical and/or ribald in nature. As a form of satire, it confronts society's "sacred cows" and/or foibles. During the mid-1800s, Edward Lear popularized this mode of expression in England. From there it spread throughout the entire world. By 1900, two branches of limerick writing were formed--the decent and the "smutty." In reviewing 7,000 traditional limericks, the authors have located about 50 significant dental, oral and/or facial limericks which are socially acceptable. In addition, we have produced 50 original poems of this nature. This article offers 33 traditional and new limerick "gems" for your reading pleasure. PMID- 15666506 TI - [HIV and AIDS in Spain]. PMID- 15666507 TI - [HIV prevalence among individuals having had serology performed in eight Spanish cities following heterosexual exposure, 1992-2003]. AB - BACKGROUND: The human immunodeficiency virus (HIV) epidemic in Spain was characterized throughout the first few years by the predominance of infections among injected drug users (IDU's), but everything currently points to a progressive predominance of sexual transmission. This study is aimed at describing the trend in HIV infection among several heterosexual populations groups and at characterizing the situations in which the infections occurred. METHODS: Descriptive study of the individualls who had the test conducted for the first time at nine HIV diagnosis centres within the 1992-2003 period due to high risk heterosexual exposures, not including injected drug users. RESULTS: A total of 47,870 individuals had serology performed during the period under study. The HIV prevalence among the population under study dropped from 3.2% to 1.0% (p< 0.001). Among the individuals with an HIV infected partners, the prevalence remained at around 10%, the male users of prostitution having dropped from 1.9% to 1.0% (p=0.049); among women involved in prostitution, from 1.4% to 0.7% (p=0.008); and among all other heterosexuals, having dropped from 1.6% to 1.0% among males (p=0.014) and from 1.1% to 0.7% among women (p=0.012). CONCLUSIONS: The results indicate advances in the control of heterosexual human immunodeficiency virus transmission, mainly among women, although it is necessary to still continue stressing prevention. PMID- 15666508 TI - [Intervention priorities in the acute stage of complex emergencies drafted by nine humanitarian aid agencies]. AB - BACKGROUND: Complex Emergencies are an international Public Health problem currently becoming increasingly more frequent and of growing proportions which lead to major death and disease rates, especially during the acute stage thereof. This study is aimed at identifying and analyzing the top-priority areas of intervention in the acute stage of a complex emergency drafted in the operating manuals of the main aid agencies, as well as the degree of development and structuring of the activities proposed in each area on which priority has been placed. METHODS: The intervention manuals drafted by nine major aid agencies were used as study material. A quantitative analysis was then made of the 16 intervention priorities set out, as well as of the degree to which each priority was defined based on the development of 73 variables of aspects of the proposals set out in the manuals. RESULTS: The ACNUR manual includes 90% of the 73 variables for further expansion upon the priorities, the UN Humanitarian Affaire Coordination Office manual including 35% of the 73 variables. ACNUR better expands upon the non-healthcare variables, followed by MSF and USAID. Doctors without borders shows a 97.3% degree of expansion of the healthcare variables) followed by ACNUR (94.7%), USAID (92.1%). CONCLUSIONS: ACNUR has been found to have the most integral proposal, the UN Humanitarian Affairs Coordination Office having the most discreet. There is a general trend towad further expanding upon and unifying the health indicators, whilst other aspects are not further expanded upon. PMID- 15666509 TI - [Eating-nutritional interculturality in the Wixarika ethnic group of Mexico]. AB - BACKGROUND: The high prevalence of eating-nutritional problems and the low degree of impact the food programs have, mainly because the cosmovision of this people has been overlooked warrants the recovery of the eating-nutritional culture of the Wixarika ethnic group in Mexico. This research is aimed at providing elements for constructing a sustainable, intercultaral, participation-based eating and nutrition model bringing together modernity and this people's ancestral taditions. METHODS: The participative action based on the Sociocritical epistemology was employed as the research methodology with the in-depth survey and participating ethnography techniques. This research was conducted in the Wixarika of Santa Catarina Cuexcomatitlan community in the municipality of Mezquitic, Jalisco, Mexico. RESULTS: The foods eaten by the Wixarika people have a religious meaning, in which corn is the main connecting force of their food related cosmovision. As most Mesoamerican cultures, the basis of food production and consumption is comprised by the "three sisters: corn, beans and squash, to which jitomate and chile have been added, as well as the gathering of foods from the surrounding environment such as fungus, chelites and nopal, foods which, on being produced and eaten in sufficient quantities and properly combined may provide for this ethic group being properly fed. CONCLUSIONS: Food comprises a central aspect in the way in which the cosmovision of the Wixarika people is set out, these being representations and meanings which must be integrated in order to model which will ensure the eating-nutritional soundness of this ethnic group. PMID- 15666510 TI - [Hepatitis B case grouping serological study among six chinese families in Almeria, Spain]. AB - BACKGROUND: Following the detection of two cases of members of 6 Chinese families having tested positive for the hepatitis B virus, a study of those living in these families was begun for the purpose of knowing the spread of the infection within the family environment of the cases detected. METHODS: Descriptive study. Population under study: 24 members of six Chinese families. VARIABLES: Age, sex, serological diagnosis, risk factors, healthcare-related attitude. INFORMATION SOURCES: Clinical records, serological data, epidemiological survey and immunization cards. A family focus was employed and the genogram used. DATA ANALYSIS: Distribution Binomial spread for calculating probability of occurrence of the process to be studied. RESULTS: A total of 14 males (58.3%) and 10 females (41.7%) ranking from 1 to 54 years of age were studied. The age group having the largest number of subjects studied was the age 21-30 group (37.5%). Twelve chronic hepatitis B infections were recorded (50%). No relationship was found to exist with the risk factors studied in the epidemiological survey conducted. The probability of this number of chronic hepatitis cases occurring was 0.066 x 10( 6). CONCLUSIONS: It was concluded that the prevalence of infection found was probable due to intra-family transmission. Given the low probability of occurrence of a process of this type, the case grouping found is considered to be high. PMID- 15666511 TI - [Influence of standard population on age-adjusted mortality rates: an example using cervical cancer in Spain]. PMID- 15666512 TI - [The future of oncological urology. Reflections for one program]. PMID- 15666513 TI - [Knowledge of the history of urology: a teaching commitment]. PMID- 15666514 TI - [Experimental study about viability of autologous free graft in vitro cultivated urinary epithelium]. AB - OBJECTIVE: The purpose of this study is to apply the in vitro keratinocyte culture techniques and the tissue engineering principles to urothelium, to obtain a three-dimensional autologous tissue suitable for grafting. We also showed the viability of free graft cultured urothelium in an experimental model. MATERIAL AND METHODS: An animal experimental model was designed to apply the techniques of cellular culture and tissue engineering. Biopsy specimens of bladder mucosa were obtained, in vitro cultured and posteriorly implanted in each animal. We established three groups based on different follow-up periods (7, 14 and 30 days), and made a final histomorphological study to demonstrate the viability of the graft at the end of its respective follow-up period. RESULTS: A three dimensional in vitro tissue was obtained, composed of a bio-artificial submucosa (fibrin gel and fibroblast) where the uroepithelial cells were seeding; a biodegradable polyglycolic acid mesh was used to facilitate the tissue manipulation and implantation. In the morphological study all the implants appeared viable, but the grafts with longer implantations periods were better conformed, showing a tisular structure with multiple cellular layers. CONCLUSIONS: In vitro keratinocyte culture techniques could be applied to other epithelial tissues as the urothelium. We obtained a three-dimensional in vitro tissue suitable for grafting in a relatively short time. The histological study demonstrated that free autologous urothelial graft is totally viable, opening future clinics applications. PMID- 15666515 TI - [Sacral root neuromodulation. Experience in our site: 1998-2003, concerning 18 definite neuromodulation implants]. AB - OBJECTIVE: Sacral root neuromodulation is an effective technique for the treatment of Chronic Micturition Dysfunction (CMD) refractory to conventional therapy. New indications such as chronic pelvic pain and interstitial cystitis are currently making their way within the urologic setting. Several groups of gastric surgeons are now choosing this technique for the management of rectal diseases (constipation, sphincter dysfunction). This paper contributes our experience in the treatment of patients with CMD and the comparative results at one year from implant in all patients. MATERIAL AND METHODS: From December 1998 through July 2003, 18 neuromodulation definite implants were performed. The main indication was CMD in 62.5% patients. Follow up of patients was done at one month and three months, and every six months thereafter. Follow up is conducted through a micturition diary and QoL questionnaire. Electrode implant in the first 16 patients was achieved by open surgery. The next 2 patients had the electrode placed by a percutaneous technique using the "Tined Lead" kit. RESULTS: Results at one year after implant were compared in all patients. The total number of neuromodulation implants placed was 18, 14 (77.7%) of which were women and 4 (22.3%) men. Mean age was 52.56 years. Implant indication was CMD in 72.2% patients, mixed incontinence (urinary and faecal) in 22.1%, and interstitial cystic disease in 5.5%. Symptoms improvement at one year from implant, as determined by a micturition diary and QoL questionnaire was 76.4%. Clinical improvement was greater in patients with urgency symptoms than in patients with predominance of voiding symptoms. PMID- 15666516 TI - [Prostate cancer recurrence after radical prostatectomy and salvage radiotherapy]. AB - OBJECTIVE: To evaluate the efficacy of the radiotherapy to prostatic bed in patients with biochemical recurrence for prostate cancer after radical prostatectomy. MATERIAL AND METHODS: We analyse the results of 292 patients underwent radical prostatectomy for localized prostate cancer T1-T2 between January 1992 and June 2003, with an average folow-up of 36 months (range 6 months to 12 years). We detect biochemical recurrence (PSA >0.20 ng/ml) in 75 (26%) patients. Of 75 patients with biochemical recurrence, 9 (12%) was diagnosed of local recurrence by the following criteria: a) The first PSA obtained 6 weeks after radical prostatectomy <0.20 ng/ml. b) The time to biochemical recurrence >6 months. c) The prostate specific antigen doubling time >6 months. d) The prostate specific antigen velocity after radical prostatectomy <0.75 ng/ml/year. e) The prostate specific antigen level after radical prostatectomy <2.5 ng/ml. The 9 patients diagnosed of local recurrence received an average dose of 56.42 Gy in the prostate bed. RESULTS: Of all 9 patients with local recurrence, 7 (77.7%) has complete response with an average time of follow-up of 25 months (6-30 months). The time between the radiotherapy and the response, in patients with complete response, was lower than 3 months. Were not observed significant adverse effects associated to radiotherapy. CONCLUSIONS: The salvage radiotherapy may be beneficial in select patients with local recurrence. The characteristics of prostate specific antigen elevation are useful in distinguishing men with local recurrence from those with distant metastases. PMID- 15666517 TI - [SAFYRE. A new concept for adjustable minimally invasive sling for female urinary stress incontinence]. AB - INTRODUCTION: SAFYRE is a new readjustable and minimally invasive sling for the treatment of stress urinary incontinence (SUI). Attempts to restore the normal suburethral hammock using an anatomical approach have been made in recent years. The authors report their experience with this device, which associates the efficacy of slings with readjustability. MATERIAL AND METHODS: A total of 100 consecutive patients with clinical and urodynamic diagnosis of SUI underwent SAFYRE sling procedure. The age range was from 40 to 71, mean age 63 years. Seventy-five patients (75%) presented previous failed anti-incontinence procedures. Physical clinical examination, stress test, pad use and a urodynamic study were performed before the surgery. All the patients presented symptoms of SUI and 30% also reported mild urgency. RESULTS: The average follow up period was 14 months (12-30 months). The mean operative time was of 25 minutes. Dystopia repair was performed whenever necessary, during the same procedure. The average hospital stay was 24 hours. In 3% of the implants, bladder perforation occurred. During the postoperative period, 26 patients developed urgency symptoms. During that follow up period, 92% were found to be continent, 3% reported an improvement and 5% were dissatisfied. CONCLUSION: SAFYRE is a safe and quick procedure that allows for postoperative readjustment. This technique may be an attractive alternative if the good result obtained so far proves to be long lasting. PMID- 15666518 TI - [Orgasm and its impact on quality of life after radical prostatectomy]. AB - INTRODUCTION: Orgasm is a neurophysiological event, which produces bulbous cavernous muscle contraction that usually coincided with ejaculation. The aim of this study was to assess the orgasm's presence and quality in patients treated with radical prostatectomy, as well as its impact on quality of life of these patients. METHOD: The medical records of 152 patients with radical prostatectomy were reviewed retrospectively. Patients were operated between january 1999 and december 01, with an average age of 64.4 (44-75) years and the follow-up period was 33 (21-45) months. 42 (31.6%) patients showed erectile dysfunction (ED) previous to surgery. The research was performed by a personal interview through a questionnaire. RESULTS: 134 patients (96.4%) treated showed post operative ED, 91.6% of patients had stable relationship and 44.4% have sexual intercourse, 23.3% masturbation only and 32.3% no sexual activity. 84 patients (55.2%) were not interested in receiving treatment and 25 (16.4%) referred a reduced libido. Concerning to orgasm sensation, 140 patients (92.1%) preserved a subjectively normal orgasm, 4 (2.6%) referred lack of it and 8 (5.2%) a weakened or anomalous sensation. Furthermore 24 patients (15.7) had urine loss during orgasm. CONCLUSIONS: After radical prostatectomy, both the orgasmic function and libido were kept by the majority of patients despite the neuro vascular bundle damaged caused. Only a minority of patients having urine loss, as a consequence of surgical procedure. PMID- 15666519 TI - [On the need of pubic region shaving in patients undergoing endoscopic urologic surgery]. AB - OBJECTIVE: The objective of this study is to assess the effects of preoperative shaving of the pubic region on post-operative bacteriuria after endoscopic urological surgery. MATERIALS AND METHOD: The study was carried out distributing the patients undergoing endoscopic urological surgery in a controlled randomized way in two groups. In a group the pubic region was shaved, according to the habitual techniques, while the other group was not shaved; the rest of the preparation was the same for both groups. Urine samples were collected for their culture before the administration of the prophylactic antibiotic and a week after the removal of the Foley catheter, yet without antibiotic treatment. Special attention was paid to the postoperative incidences, mainly, those of infectious nature. RESULTS: They were included a total of 449 patients, of which 149 were removed from the study by different causes. Of these, 149 were shaved and 151 were not it. In the group of unshaved patients a 19.5% of postoperative bacteriurias was observed, while in the other group it was of 16.6%. Differences in both groups were not statistically significant. CONCLUSIONS: We conclude that there is no an increase of postoperative bacteriurias in the unshaved patients undergoing endoscopic urological surgery, compared with the group of patients shaved with disposable bladders. PMID- 15666520 TI - [Ureteral triplication, an unusual isolated presentation]. AB - We report the case of a 26 years old female who presented, as a casual finding, a right complete ureteral triplicity (probably Campbell's tipe I) with no other associated abnormalities. Despite ureteral duplication is a frequent urological anomaly, the presence of three ureters occurring in the same side, in a complete or incomplete form, is a true rarity, with less than 100 cases publicated up to date. The present case is singular, not only because of its exceptional presentation but also because no other urological or extraurological anomalies were associated and no clinical manifestations were present. The most relevant literature is reviewed. PMID- 15666521 TI - [Epithelioid hemangioendothelioma of penis]. AB - A case of EH of the penis in a sixtyfour-year-old patient is described. This is a rare--in this location--vascular tumor of borderline malignant potential between hemangioma and angiosarcoma. Local excision of the tumor with a cut-off of normal tissue and careful follow-up of the patient is the accepted procedure. Immunohistochemistry is essential for the diagnosis. PMID- 15666522 TI - [Brucellar orchiepididymitis]. AB - Brucellosis is a cause of human disease, especially in regions whose brucellosis in animals is enzootic. Brucella orchitis is a rare complication of systemic brucellosis (2-14%). Necrotizing orchitis is posible and it must be distinguished from necrotizing involvement arising from other pathogens (TB or Salmonella). Treatment with specific antimicrobial medication helps to avoid unnecessary orchiectomy and prevent infectious disease in the contralateral testis as well as systemic manifestations. We report a case of brucellar epididymo-orchitis presenting as an acute scrotal mass in a 19 years old male who lived in a city. The anamnesis was crucial in the diagnosis of this rare testicular infection. We review the diagnosis and management of this patology. PMID- 15666523 TI - [Resolution of a case of arterial priapism secondary to bilateral arteriocavernous fistula with selective embolization using reabsorbible material]. AB - Priapism is defined as persistent erection without sexual stimulus. The new classifications make a distinction between venooclusive low flow priapism, isquemic and arterial high flow priapism, non isquemic. The perineal or penis trauma are responsible the most of cases arterial priapism, formation of an arteriocavernous fistula. The diagnosis is clinical, blood gas analysis and colour Doppler ultrasonography of the corpora cavernosa confirmed the diagnosis. The arteriography with selective embolization constitutes a safe and effective therapeutic method. We report one case of arterial priapism secondary to bilateral arteriocavernous fistula resolved with selective embolization using reabsorbible material. PMID- 15666524 TI - [Epidermoid carcinoma of the urethra and glans penis: two independent tumors?]. AB - A 64 years-old man is treated in our hospital presenting a big tumor in glans penis. During the surgical act, another neoplasm is detected in the urethra. The microscopic study showed a concomintant epidermoid carcinoma of glans penis and urethra with non-affected tissue between both tumors. PMID- 15666525 TI - [Adrenal gland hematoma due to rupture of myelolipoma. A case report]. AB - The myelolipoma is a benign tumour of the adrenal cortex, non functioning and normally asymptomatic, which can be diagnosed by chance in imaging test. Rupture and bleeding of the myelolipoma is an infrequent complication, which can conclude with the formation of a haematoma or, less usually, result in a massive retroperitoneal haemorrhage. The compression of adjacent structures by retroperitoneal haematomas with adrenal origin is a possible but not frequent eventuality. Indications for surgery of haematoma retroperitoneal continue to be, nowadays, a difficult and controversial decision, because the problem is normally resolved with conservative treatment and a watchful approach. We are presented with a 69 year old male who came to casualty with abdominal pain of 7 days duration, whose origin coincided with a light blow. The image test revealed the existence of a retroperitoneal haematoma with adrenal gland origin of 16 cm, with active bleeding, which was displaced and compressed the cava vein. We have only found one case, in the literature, of compression of the cava vein coused by adrenal gland haematoma. In the case under our observation, urgent surgery was needed. PMID- 15666527 TI - [False postvoid residual volume diagnosed by videourodynamics]. AB - We present a case report of a young male patient, with a bilateral vesico renal reflux. The urodynamic study findings suggested the possibility of a non neurogenic bladder-external spincter dissinergya producing a valuable residual volume. After biofeedback treatment, the dissinergia disappeaed, but residual volume persisted. The videourodynamic assessment allowed us the accurate diagnosis of a false residual volume, produced by the voiding of the refluxed urine from the ureters into the bladder. PMID- 15666526 TI - [Vesical endometriosis: report of two cases]. AB - Endometriosis affecting the urinary tract is very rare and the most common site of involvement is urinary bladder. The clinical features are urgency and frequency, hipogastric pain and hematuria. Cistoscopic examination is the most valuable diagnostic test but definitive diagnosis requires histologic confirmation. We report 2 cases of endometriosis in 2 young women, one with previously cesarean section, in which surgical treatment was effective. After 1 and 3 years of follow-up respectively the patients remain assymptomatic. PMID- 15666529 TI - [Double-J catheters: 4 years of evolution]. PMID- 15666531 TI - [Prostate rhabdomyosarcoma]. PMID- 15666532 TI - Flavonoids as chemotaxonomic markers for Erythroxylum australe. AB - Methanolic leaf extracts of Erythroxylum australe F. Muell. produced eight O conjugated flavonoids. Six of the flavonoid aglycones were dihydroisoflavones (all dihydro-orobol derivatives), one a flavanone, eriodictyol, and one a flavonol, quercetin. The major glycosides of the flavonoids included mono glucosyl-rhamnosyls and dirhamnosyl-glucosides with either 3, 5, 7 or 3', 4' linkage or a combination thereof The array of flavonoids present in E. australe suggests kinship to E. ulei and linkage to the four cultivated alkaloid-bearing Erythroxylum, especially the ancestral E. coca var. coca. Because of the uniqueness of the flavonoids present in leaf tissue of E. australe they are unambiguously useful as chemotaxonomic markers for the taxon. PMID- 15666533 TI - Antimicrobial activity, essential oil composition and micromorphology of trichomes of Satureja laxiflora C. Koch from Iran. AB - The antimicrobial activity, essential oil composition and micromorphology of trichomes of Satureja laxiflora C. Koch, a native plant from Iran, were studied. The essential oil was obtained from the aerial parts at the flowering stage by hydrodistillation, and analyzed by GC and GC/MS. Thirty-three compounds representing 99.1% of the total oil were characterized. The major compounds were thymol (63.9%) and gamma-terpinene (11.9%) followed by carvacrol (4.8%), p-cymene (3.9%), geraniol (3.2%) and geranyl acetate (3.1%). Furthermore, the essential oil and its three main components were tested against two bacteria and three fungi. The result of the bioassays has been shown that the oil possesses potent antimicrobial property. Chemical studies confirmed that a major portion of this antimicrobial activity is due to thymol present in the oil. Micromorphological analysis by SEM of both vegetative and reproductive organs revealed the presence of abundant sessile capitate and sparse short-stalked glandular trichomes along with retrorse eglandular hairs, giving useful diagnostic characters for identification of this medicinal plant. PMID- 15666535 TI - Evaluation of in vivo biological activity profile of isoorientin. AB - Anti-nociceptive, anti-inflammatory and gastroprotective activities of the known C-glycosyl flavonoid, isoorientin, were studied in rats and mice. For the anti nociceptive activity assessment the p-benzoquinone-induced writing test, for the anti-inflammatory activity the carrageenan-induced hind paw edema model in mice, and for the gastroprotective activity the EtOH-induced ulcerogenesis model in rats were used. Isoorientin was shown to possess significant anti-nociceptive and anti-inflammatory activities at 15 mg/kg and 30 mg/kg doses, without inducing any apparent acute toxicity as well as gastric damage. However, the compound did not possess any significant gastroprotective activity against EtOH-induced ulcerogenesis. PMID- 15666536 TI - Antifungal activity of the carrot seed oil and its major sesquiterpene compounds. AB - Carrot seed oil is the source of the carotane sesquiterpenes carotol, daucol and beta-caryophyllene. These sesquiterpenic allelochemicals were evaluated against Alternaria alternata isolated from the surface of carrot seeds cultivar Perfekcja, a variety widely distributed in horticultural practise in Poland. Alternaria alternata is one of the most popular phytotoxic fungi infesting the carrot plant. The strongest antifungal activity was observed for the main constituent of carrot seed oil, carotol, which inhibited the radial growth of fungi by 65% at the following concentration. PMID- 15666534 TI - Antimicrobial activity of N-alkoxycarbonylmethyl-N-alkyl-piperidinium chlorides. AB - The aim of the study was to assay antibacterial and antifungal activity of newly synthesised N-alkoxycarbonylmethyl-N-alkyl-piperidinium chlorides. The compounds tested were found to inhibit the growth of some Gram-negative bacteria, Gram positive strains and some representatives of yeast-type Candida. From microbiological experiments two of the compounds tested, N dodecyloxycarbonylmethyl-N-methyl-piperidinium chloride (3) and N-dodecyl-N ethoxycarbonylmethyl-piperidinium chloride (6), emerged as more active than the other compounds. Since the resistance of biofilms to biocides should be noted during the design and testing of new antimicrobial agents therefore, we have analysed antibacterial properties of the most active compounds towards biofilms. Our study focused on strains of Pseudomonas aeruginosa and Staphylococcus aureus that served as main model organisms for the biofilm studies. PMID- 15666537 TI - Constituents of Quinchamalium majus with potential antitubercular activity. AB - Antitubercular bioassay-guided fractionation of the dichloromethane extracts of the above-ground biomass and roots of Quinchamalium majus led to the identification of six known constituents, betulinic acid (1), daucosterol (2), 5,7-dihydroxyflavone (3), oleanolic acid (4), (-)-2S-pinocembrin (5), and ursolic acid (6), for the first time in this species. Their chemical structures were determined on the basis of spectroscopic evidence and chemical transformation methods. All of these compounds along with additional 11 analogues were evaluated for their antitubercular potential against Mycobacterium tuberculosis in a microplate alamar blue assay, and the primary structure-activity relationships (SARs) for 4 and 6 were discussed. In addition, all the isolates were tested for cytotoxicity against African green monkey Vero cells in order to evaluate for their selectivity potential. PMID- 15666538 TI - Phytogrowth-inhibitory lactones derivatives of glaucolide B. AB - The sesquiterpene lactone glaucolide B (1), isolated from Vernonia fruticulosa (Asteraceae), was transformed into six lactones (2-7). The structures of the products were elucidated by spectroscopic analysis. A series of solutions of compounds 1-7, at 200 microM, were tested on the germination and on the root and shoot growth of the dicotyledons Physalis ixocarpa and Trifolium alexandrinum and of the monocotyledons Lolium multiflorum and Amaranthus hypochondriacus. Lactone 5 exhibited clear selectivity towards dicotyledonous species at 200 microM, with an average inhibition of 90% on the germination of P. ixocarpa. Lactones 1, 3 and 4 had a greater effect on root length of monocotyledonous species, inhibiting around 70% at 200 microM in L. multiflorum. It seems that the diol function is required in lactones 4-6 to increase the activity, the polarity in the molecule might be required to reach its target. PMID- 15666539 TI - DPPH free radical scavenger components from the fruits of Alpinia rafflesiana Wall. ex. Bak. (Zingiberaceae). AB - The methanol extract of the dried ripe fruits of Alpinia rafflesiana was investigated for its DPPH free radical scavenger constituents. 2',3',4',6' Tetrahydroxychalcone (7), which has never been isolated from natural sources was found to be most active as a DPPH free radical scavenger with the IC50 value of 55 microM. Other known compounds isolated from this species include 5,6 dehydrokawain (1), flavokawin B (2). 1,7-diphenyl-5-hydroxy-6-hepten-3-one (3), ( )-pinocembrin (4), cardamonin (5) and (-)-pinostrobin (6). The DPPH free radical scavenger compounds were detected using TLC autographic analysis. The percentage inhibition of DPPH free radical scavenging activity was measured on isolates (5 7) using colorimetric analysis. PMID- 15666540 TI - Antibiotics in the chemical communication of fungi. AB - In dual cultures Oudemansiella mucida and Xerula melanotricha (basidiomycetes) react to the presence of living Penicillium notatum or P. turbatum with an increased production of strobilurin A (1) or X (2). P. notatum in turn reacts to the two basidiomycetes or their antibiotic strobilurin A alone with the production of N-(2-hydroxypropanoyl)-2-aminobenzoic acid amide (3) or chrysogine (4). P. melinii and P. urticae overgrow O. mucida due to complete resistance to strobilurin A. P. brevicompactum, P. citrinum, P. janczewskii and the other Penicillium strains are all sensitive but apparently do not induce O. mucida to produce the amounts of strobilurin A needed to inhibit their growth. PMID- 15666541 TI - pH-dependent quenching of the fluorescence of tryptophan residues in class A beta lactamase from E. coli (TEM-1). AB - We performed an investigation of the pH-dependent quenching of the fluorescence of tryptophan residues of TEM-1 beta-lactamase from E. coli by uncharged and charged quenchers. pH-dependent Stern-Volmer constants (Ksv/pH) of tryptophan residues allowed us to determine subtle but discrete structurally and functionally important processes. PMID- 15666542 TI - Effects of theobroxide, a natural product, on the level of endogenous jasmonoids. AB - The natural potato microtuber inducing substance, theobroxide, strongly induces the formation of tuber of potato (Solanum tuberosum L.) and flower bud of morning glory (Pharbitis nil) plants under non-inducing conditions (long days) (Yoshihara et al., 2000). In the present study, theobroxide was evaluated for its effect on the level of endogenous jasmonoids in different tissues of such two plants. An in vitro bioassay using cultures of single-node segments of potato stems was performed with the supplement of theobroxide in the medium. The endogenous jasmonic acid (JA) and its analogue tuberonic acid (TA, 12-hydroxyjasmonic acid) in segments and microtubers were quantitatively analyzed. The increase in the endogenous JA level caused by theobroxide was observed in both segments and microtubers. Endogenous TA was only detected in segments, and the content increased with the concentration of theobroxide. As for morning glory, the whole plant was sprayed with theobroxide for 1 approximately 5 weeks under different photoperiods and endogenous JA in the leaves was quantitatively analyzed. Theobroxide spraying increased the level of endogenous JA in the leaves of the plants grown under both long and short days. PMID- 15666543 TI - Quantification and fatty acid profiles of sulfolipids in two halophytes and a glycophyte grown under different salt concentrations. AB - This study was aimed at understanding the role of sulfolipids in salt tolerance mechanisms of the halophytes Aster tripolium L., Compositae, and Sesuvium portulacastrum L., Aizoaceae, and of the glycophyte Arabidopsis thaliana (L.) Heynh., Brassicaceae. In Aster and Sesuvium the sulfolipid contents increased significantly under salt stress conditions (517 mM or 864 mM). In Arabidopsis, changes in sulfolipid contents were not observed (NaCl up to 100 mM). The fatty acid profile of sulfoquinovosyldiacylglycerol (SQDG) in Aster was modified with increasing NaCl concentrations. LC-MS analyses of sulfolipids from Aster and Sesuvium revealed the presence of 18:3/18:3 and 16:0/18:3 molecules. Obviously, the function of sulfolipids during salt stress differs between halophytic species and between halophytes and glycophytes where sulfolipid accumulation was not observed. PMID- 15666544 TI - Up-regulation of pathogenesis-related proteins in the apoplast of Malus domestica after application of a non-pathogenic bacterium. AB - The intercellular washing fluid (IWF) of Malus domestica cv. Holsteiner Cox before and after application of the non-pathogenic bacterium Pseudomonas fluorescens Bk3 to the leaves was investigated in a comparative manner. SDS-PAGE in combination with ESI Q-ToF mass spectrometry, and homology search in relevant data bases revealed the highly up-regulated expression of several pathogenesis related plant proteins in the apoplast of the leaves treated with P. fluorescens. These proteins were beta3-1,3-glucanase, chitinase, thaumatin-like protein, ribonuclease-like protein, and a hevein-like protein. Moreover, a 9 kDa non specific lipid transfer protein was significantly reduced after the application of P. fluorescens. The possible relevance of a pre-treatment of apple cultivars with the non-pathogenic bacterium P. fluorescens Bk3, as an alternative method to the treatment with fungicides, for increasing the resistance of susceptible apple cultivars against an infection with the fungus Venturia inaequalis is discussed. PMID- 15666545 TI - Evaluation of the oxidative burst in suspension cell culture of Phaseolus vulgaris. AB - Plants respond to the attack of pathogens with the oxidative burst, a production of reactive oxygen species (ROS). In this work a cell culture suspension of Phaseolus vulgaris was used to investigate the oxidative burst triggered by a conidia suspension of different races of Colletotrichum lindemuthianum. As a defence response of the cells a two-phase peak was observed with all used races of Colletotrichum lindemuthianum, varying only in the produced amounts of hydrogen peroxide. Findings with additives such as superoxide dismutase (SOD), diphenyleneiodonium (DPI) and catalase gave rise to the conclusion that more superoxide radicals were produced than be detectable with Amplex Red as hydrogen peroxide. It is assumed that the conversion of the superoxide radical is spontaneous and not driven via a cell-derived superoxide dismutase. The addition of low-molecular cell wall components (ergosterol, glucosamine, galactosamine) showed clearly that compounds like this act as elicitors and thus are involved in triggering the burst. Furthermore, an evaluation of the metabolizing capacities of hydrogen peroxide of the suspension culture cells revealed the enormous capacity of the cells to detoxify this ROS. PMID- 15666546 TI - Role of salicylic acid in tomato defense against cotton bollworm, Helicoverpa armigera Hubner. AB - We investigated the role of the salicylic acid (SA) signaling pathway in defense responses of tomato plants to the herbivore, cotton bollworm. After exposure to the cotton bollworm, tomato leaves rapidly accumulated a high level of SA. The transcription of PR1 and BGL2 genes, the marker genes of SA pathway, was up regulated. An enhanced endogenous SA level was accompanied by an increase in the endogenous H2O2 level as compared with controls. Spraying tomato plants with a solution containing either SA or methyl salicylic acid (Me-SA), the H2O2 level dramatically increased. These data proved that the SA pathway was involved in the tomato plant defense responses to the herbivore. PMID- 15666547 TI - Induction of tropane alkaloid formation in transformed root cultures of Brugmansia suaveolens (Solanaceae). AB - Hairy root cultures of Brugmansia suaveolens were set up by infection of root tips with Agrobacterium rhizogenes. The successful transformation was confirmed by analysing rolC and virC genes using polymerase chain reaction (PCR). Hairy root cultures were employed to study the formation of tropane alkaloids, such as hyoscyamine. The transformed cultures were incubated with potential elicitors, such as methyljasmonate, quercetin and salicylic acid in order to stimulate the biosynthesis of tropane alkaloids. Profile and amounts of tropane alkaloids were analysed using capillary GLC-MS. At least 18 different tropane alkaloids could be identified. Treatment of the cultures with 200 microM methyljasmonate increased the alkaloid accumulation 25-fold up to a level of 1 mg/g fresh weight as compared to untreated controls. Quercetin enhanced the alkaloid production 10 fold (0.4 mg/g fresh weight) within 24 h. In contrast 100 microM salicylic acid decreased alkaloids to a level of 1 microg/g fresh weight. PMID- 15666548 TI - Genome relationship among nine species of Millettieae (Leguminosae: Papilionoideae) based on random amplified polymorphic DNA (RAPD). AB - Random amplified polymorphic DNA (RAPD) marker was used to establish intergeneric classification and phylogeny of the tribe Millettieae sensu Geesink (1984) (Leguminosae: Papilionoideae) and to assess genetic relationship between 9 constituent species belonging to 5 traditionally recognized genera under the tribe. DNA from pooled leaf samples was isolated and RAPD analysis performed using 25 decamer primers. The genetic similarities were derived from the dendrogram constructed by the pooled RAPD data using a similarity index, which supported clear grouping of species under their respective genera, inter- and intra-generic classification and phylogeny and also merger of Pongamia with Millettia. Elevation of Tephrosia purpurea var. pumila to the rank of a species (T. pumila) based on morphological characteristics is also supported through this study of molecular markers. PMID- 15666549 TI - A 1.3 kb satellite DNA from Bubalus bubalis not conserved evolutionarily is transcribed. AB - A 1.3 kb satellite DNA from a size defined genomic library of mammal Bubalus bubalis was cloned and sequenced. The clone pSB1 is AT rich with 447 A (33.6%), 262 C (19.7%), 240 G (19.0%) and 383 T (28.8%). There were about 1400 copies of contig in the bubaline genome but it did not uncover allele length variation when used as probe in conjunction with a number of restriction enzymes. The contig pSB1 is not conserved evolutionarily and cross hybridizes only with the Bovideae family. A set of primers from 5' (nt 422 to 441) and 3' (nt 962 to 947) deduced from the clone used for PCR amplification with four members of the Bovideae family gave the expected 530 bp band of equal intensity indicating a similar number of copies in all the four species namely Bos indicus, Capra hircus, Ovis aries and Bubalus bubalis. Expression studies with pSB1 following slot-blot hybridization with total RNA isolated from ovary, testes, kidney, lung and spleen revealed varying signal intensities in all the tissues with a most prominent signal in spleen but a faint one in ovary. Further sequence analysis revealed the presence of several eukaryotic transcriptional elements such as NF-E1, Poly-A signal, lariat consensus sequences, and CTF/NF1 binding sites. Blast search showed 90% sequence similarity with the reverse transcriptase gene of Bos taurus and sequences from nt 283 to 636 within the contig showed highly conserved reverse transcriptase like signatures along with N-glycosylation and protein kinase C phosphorylation sites. From the data we conclude that the pSB1 representing satellite DNA is associated with transcribing sequences. The prospect of identifying functional genes linked with the satellite fraction in higher vertebrates is discussed. PMID- 15666550 TI - Thiol-reactive clenbuterol analogues conjugated to bovine serum albumin. AB - Several novel thiol-reactive clenbuterol analogues were coupled in high yield with bovine serum albumin (BSA). After labelling of unreacted cysteines with maleimide spin label (MiSL), the yield of the coupling reaction was determined by electron paramagnetic resonance (EPR) spectroscopy and spectral analysis. Two spin-probe populations with different mobility states were quantitatively determined. Molecular dynamics was used to model the structure of clenbuterol analogues and spin label conjugated to BSA and recognition of conjugates by anti clenbuterol antibodies was demonstrated. The recognition of BSA-A, BSA-C and BSA S conjugates with monoclonal and polyclonal anti-clenbuterol (mCLB-Ab and rCLB Ab) antibodies was an indication, that chlorine substituents on the aromatic ring of clenbuterol derivatives are not necessary for the binding of antibodies to the conjugates. These results confirmed the importance of the tert-butylamino group as a part of the epitope and contribute to the understanding of the recognition process with anti-clenbuterol antibodies. PMID- 15666551 TI - Activity and stability of catalase in nonionic micellar and reverse micellar systems. AB - Catalase activity and stability in the presence of simple micelles of Brij 35 and entrapped in reverse micelles of Brij 30 have been studied. The enzyme retains full activity in aqueous micellar solution of Brij 35. Catalase exhibits "superactivity" in reverse micelles composed of 0.1 M Brij 30 in dodecane, n heptane or isooctane, and significantly lowers the activity in decaline. The incorporation of catalase into Brij 30 reverse micelles enhances its stability at 50 degrees C. However, the stability of catalase incubated at 37 degrees C in micellar and reverse micellar solutions is lower than that in homogeneous aqueous solution. PMID- 15666552 TI - Quantification of the particle method for chemotactic bioassay using Peronosporomycete zoospores. AB - We estimated the amount of test solution absorbed by each Chromosorb W AW particle (60-80 mesh) using an isotopic technique to quantitate the particle method. 14C-Labeled standard compounds like carbendazim (MBC), 5-O methylcochliophilin A, sucrose and proline were dissolved in several solvents, and Chromosorb carrier particles were treated with the solution to coat the particle with these test compounds. The ratios of the radioactivity of 5 microl of the test solution to that of 2 mg of carrier particles treated with the solution at some different concentrations were measured. It was found that each carrier particle holds approx. 3.8 nl of the test solution within a range of 2 x 10(-3) to 1 x 10(-7) M concentrations. Accordingly, it is now possible to widely use the particle method as a quantitative procedure to assay chemotaxis of Peronosporomycete zoospores. PMID- 15666553 TI - An improved micromethod for tyrosine estimation. AB - A modified and improved micromethod for tyrosine determination has been developed. The method is sensitive, economic and applicable for estimation of tyrosine released in enzymatic reactions and in tissue. A range of Folin Ciocalteu (FC) reagent was used to optimize the conditions for the development of blue color. Thus in 1.5 ml of the assay system, the suitably diluted FC reagent at the final concentration of 0.2 N gave a rapid optimum color development with an absorption maximum at 750 nm. Color development showed a linear relationship in the range of 2 to 16 microg tyrosine for a described assay system under optimized conditions. Thus, the method is 3-fold more sensitive in terms of its estimation range than a conventional method. The blue color formed was stable up to 24 h. The applicability of the method for tyrosine determination in the assay of lysosomal cathepsin D and in tissue was checked by comparison to the conventional procedure. Under both systems the results obtained by the micromethod were identical to those obtained by the conventional method. In general the method that produces quantitatively a blue color, not only is rapid and economical in terms of chemical usage but also has application for routine biochemical analysis. PMID- 15666554 TI - Duty of care owed to parents of child with genetic disorder. PMID- 15666555 TI - Tortious interference claim remanded for trial. PMID- 15666557 TI - Marketing your medical practice--part VI. PMID- 15666558 TI - SORT: the strength-of-recommendation taxonomy. PMID- 15666560 TI - Measurement of 15N csa/dipolar cross-correlation rates by means of Spin State Selective experiments. AB - We propose a method for the determination of (15)N csa/dipolar cross-correlation rates based on the measurement of the two apparent transverse (or longitudinal) relaxation rates associated with each component of the nitrogen doublet (N(alpha) and N(beta)). This is achieved by inserting a spin state selective scheme in conventional inverse Carr-Purcell-Meiboom-Gill (or inversion-recovery) pulse sequence which allows for the edition of a HSQC-type spectrum for each of the spin states. Transverse cross-correlation rates necessitate two independent sets of measurements (for N(alpha) and N(beta), respectively), whereas for longitudinal cross correlation rates, besides N(alpha) and N(beta) measurements, the method requires the knowledge of both the (15)N longitudinal auto-relaxation rate and the longitudinal two-spin order (2NzHz) auto-relaxation rate. These additional parameters are mandatory because of the non-exponential behavior of the N(alpha) and N(beta) longitudinal decays. Conversely, the present method does not require any complex manipulation of 2D spectra, the cross-correlation rates being obtained from the difference of the two (N(alpha) and N(beta)) apparent relaxation rates. This approach is applied to (15)N-labelled ubiquitin at two different magnetic fields (9.4 T and 14.1 T). PMID- 15666559 TI - Improved simulation of NOESY spectra by RELAX-JT2 including effects of J coupling, transverse relaxation and chemical shift anisotrophy. AB - RELAX-JT2 is an extension of RELAX, a program for the simulation of 1H 2D NOESY spectra and (15)N or (13)C edited 3D NOESY-HSQC spectra of biological macromolecules. In addition to the already existing NOE-simulation it allows the proper simulation of line shapes by the integrated calculation of T(2) times and multiplet structures caused by J-couplings. Additionally the effects of relaxation mediated by chemical shift anisotropy are taken into account. The new routines have been implemented in the program AUREMOL, which aims at the automated NMR structure determination of proteins in solution. For a manual or automatic assignment of experimental spectra that is based on the comparison with the corresponding simulated spectra, the additional line shape information now available is a valuable aid. The new features have been successfully tested with the histidine-containing phosphocarrier protein HPr from Staphylococcus carnosus. PMID- 15666561 TI - An evaluation of chemical shift index-based secondary structure determination in proteins: influence of random coil chemical shifts. AB - Random coil chemical shifts are commonly used to detect protein secondary structural elements in chemical shift index (CSI) calculations. Though this technique is widely used and seems reliable for folded proteins, the choice of reference random coil chemical shift values can significantly alter the outcome of secondary structure estimation. In order to evaluate these effects, we present a comparison of secondary structure content calculated using CSI, based on five different reference random coil chemical shift value sets, to that derived from three-dimensional structures. Our results show that none of the reference random coil data sets chosen for evaluation fully reproduces the actual secondary structures. Among the reference values generally available to date, most tend to be good estimators only of helices. Based on our evaluation, we recommend the experimental values measured by Schwarzinger et al.(2000), and statistical values obtained by Lukin et al. (1997), as good estimators of both helical and sheet content. PMID- 15666562 TI - Quantitative J correlation methods for the accurate measurement of 13C'-13Calpha dipolar couplings in proteins. AB - Methods are described for the precise and accurate measurement of one-bond dipolar (13)C'-(13)C(alpha) couplings in weakly aligned proteins. The experiments are based on the principle of quantitative J correlation, where (1)J(C'C(alpha)) (or (1)J(C'C(alpha)) + 1D(C'C(alpha)) is measured from the relative intensity of two interleaved 3D TROSY-HN(CO)CA or 3DTROSY-HNCO spectra recorded with dephasing intervals of zero (reference spectrum) and approximately 3/(2(1)J(C'C(alpha)) (attenuated spectrum). In analogy to other quantitative J correlation techniques, the random error in the measured (1)J(C'C(alpha)) value is inversely proportional to the signal-to-noise ratio in the reference spectrum. It is shown that for weakly aligned proteins, with the magnitude of the alignment tensor of D(a)(NH) < or = 10-15 Hz, the systematic errors are typically negligible. The methods are demonstrated for the third IgG-binding domain of protein G (GB3) and a-synuclein in complex with a detergent micelle, where errors in (1)D(C'C(alpha)) of less than 0.1 Hz and ca. 0.2 Hz,respectively, are estimated. Remarkably, the dipolar couplings determined for GB3 are in even better agreement with the recently refined 1.1-angstroms X-ray structure than the input (13)C'-(13)C(alpha) couplings used for the refinement. PMID- 15666563 TI - Can traumatic brain injury surveillance systems be used to link individuals with TBI to services? AB - PRIMARY OBJECTIVE: This study was conducted to determine the feasibility of using Colorado Traumatic Brain Injury (TBI) Surveillance System data to link individuals to information and services in their communities. METHODS AND PROCEDURES: Using a qualitative exploratory approach, the investigators conducted focus groups of individuals with TBI and family members (n = 29) and individual interviews with state agency, medical and community services representatives (n = 15). MAIN OUTCOMES: The results showed that the participants saw many current problems with linking persons to services and with accessing care. The participants supported using TBI surveillance data to link persons to information and services, offered suggestions, discussed confidentiality and consent issues, described possible cultural competence issues and addressed cost feasibility. CONCLUSIONS: Overall persons with TBI and their family members overwhelmingly supported using the Colorado TBI Surveillance System to link persons to services. One major concern, however, was how to link persons who were not included in the surveillance data because their TBI happened before the surveillance system was implemented or because their injury did not result in hospitalization. This concern is addressed in a Linkage Model. PMID- 15666564 TI - Exploring the service needs and experiences of persons with TBI and their families: the South Carolina experience. AB - PRIMARY OBJECTIVE: This article presents the results of four focus groups with persons with traumatic brain injury (TBI) and their families living in South Carolina. The objective was to learn what participants perceive their service needs to be and where they experience service gaps in the existing system of TBI services. METHODS AND PROCEDURES: Four focus groups were conducted. In each group, a convenience sample of persons < 5 years post-injury and family members responded to 10 semi-structured questions. MAIN OUTCOMES: Qualitative content analysis revealed overwhelming consensus regarding the need for (1) early, continuous, comprehensive service delivery; (2) information/education; (3) formal/informal advocacy; (4) empowerment of persons with TBI/families; and (5) human connectedness/social belonging. CONCLUSIONS: Persons with TBI and families in South Carolina experience the service system as unorganized, uneducated, unresponsive and uncaring. Effective strategies are needed that link services into an ongoing continuum of TBI care, increase TBI-specific education and awareness and foster social re-integration. PMID- 15666565 TI - The role of sub-cortical brain structures in emotion recognition. AB - PRIMARY OBJECTIVE: This study investigated the role of sub-cortical brain structures in emotion recognition. METHODS AND PROCEDURES: Fourteen patients (eight left, six right) with sub-cortical brain damage (SS) and 14 matched healthy volunteers (HV) were recruited. A brief neuropsychological battery was administered to measure working memory, visual inattention, Stroop effect and visual organization. A facial and prosodic emotion recognition battery previously developed was used. RESULTS: SS patients were generally impaired on emotion recognition, with the exception of facial emotion discrimination and tasks involving happy expressions, relative to HV. Preliminary analyses also showed no statistical difference between patients with left- and right-sub-cortical brain damage in terms of emotion recognition. CONCLUSIONS: The findings provide further support for the role of sub-cortical brain structures (and the damage thereof) as well as probable frontal-limbic neural networks in recognizing basic emotions. PMID- 15666566 TI - A comparison of attention, processing and strategy use by adults with and without acquired brain injuries. AB - PRIMARY OBJECTIVE: To compare the visual attention, processing and strategy use of adults with and without acquired brain injuries (ABI). DESIGN AND METHODS: A repeated-measures, matched groups design was used with 30 adults. Pre-test/ post test measures included three functional computer tasks. Weekly measures included a computerized version of PASAT and two computerized matching tasks. INTERVENTIONS: Six weekly sessions consisted of computerized tasks designed to challenge visual attention and processing. RESULTS: Both groups improved on all measures. The participants without neurological impairments performed better on all measures. Both groups used similar strategies to complete the visual attention and processing tasks. Strategy use was much more prolific among participants without neurological deficits. CONCLUSIONS: ABI results in reduced visual attention and processing during functional computerized tasks and in the ability to use strategies for these deficit areas. Despite better visual attention and processing skills, participants without neurological deficits used more strategies to complete functional tasks. PMID- 15666567 TI - Effects of creatine and cyclocreatine supplementation on kainate induced injury in pre-pubescent rats. AB - PURPOSE: To investigate if energy precursor supplementation is neuroprotective in two neuroexcitotoxicity models; the kainate and the kainate followed by chronic phenobarbital models. METHODS: Rats in experiment 1 received 1% creatine or cyclocreatine chow from age (P) 21-65 days, underwent kainate induced status epilepticus on P35 and were compared, as adults, to kainate alone rats and to normal controls. Rats in experiment 2 received 1% creatine chow (P21-P85), underwent kainate status epilepticus on P35, received daily phenobarbital (or saline) injections (P36-P85) and were compared, as adults, to kainate, kainate phenobarbital and to normal control rats that received regular chow. RESULTS: In experiment 1, the cyclocreatine-kainate group had increased emotionality and visuospatial learning deficits on the handling and watermaze tests as compared to all other groups. Creatine supplementation did not have any effects. In experiment 2, creatine supplementation did not prevent spontaneous recurrent seizures, aggressivity on the handling test or hippocampal histologic injury. CONCLUSION: Energy precursor supplementation in the doses used did not have neuroprotective effects in the kainate or kainate-phenobarbital models in pre pubescent rats. PMID- 15666568 TI - Cerebral anoxia and disability. AB - OBJECTIVES: To describe the sequelae of cerebral anoxia following out-of-hospital cardiac arrest, to study the functional outcome and to seek a link between the acute stage and the disability. METHOD: A retrospective study was performed. The initial findings and the neurological and neuropsychological status are recorded of 12 patients admitted to the rehabilitation unit for after-effects of cerebral anoxia following out-of-hospital cardiac arrest. RESULTS: After clinical and neuropsychological assessment, all patients displayed cognitive impairment. Two groups of patients appeared: seven patients were severely disabled with a dysexecutive and behavioural frontal lobe syndrome and memory deficit; five out of the seven also presented an extra-pyramidal syndrome; the other five patients presented behavioural dysfunction related to frontal lobe disorder but were independent in daily life activities. No correlation was found between the acute stage data and the outcome. CONCLUSION: Neurological and neuropsychological impairment after cerebral anoxia may be severe but seems difficult to predict. A dysexecutive syndrome was noted in all 12 patients. PMID- 15666569 TI - The Sensory Modality Assessment and Rehabilitation Technique (SMART): a valid and reliable assessment for vegetative state and minimally conscious state patients. AB - PRIMARY OBJECTIVE: To establish the reliability and validity of the Sensory Modality Assessment and Rehabilitation Technique (SMART) as a tool for discriminating awareness in patients with profound brain damage. RESEARCH DESIGN: A comparative prospective study was conducted. METHOD AND PROCEDURES: Sixty subjects diagnosed in vegetative state (VS) on admission were assessed at 2 monthly intervals. Rancho level ratings derived from referring physicians, SMART and Western Neuro Sensory Stimulation Profile (WNSSP) scores were compared. MAIN OUTCOMES AND RESULTS: The intra-observer intra class correlation (ICC) was 0.97 and inter-observer ICC was 0.96, implying very little within and between observer scoring variations. A modest, although significant correlation was established between SMART and either physician or WNSSP scores. However, the correlation between the WNSSP and SMART was higher (r = 0.70) than that between WNSSP and physicians scores (r = 0.451) or between SMART and physicians (r= 0.474). CONCLUSION: SMART is a valid and reliable assessment for discriminating awareness in VS and Minimally Conscious State (MCS). PMID- 15666570 TI - The Potential Unlimited Programme: an outdoor experiential education and group work approach that facilitates adjustment to brain injury. AB - Two pilot Potential Unlimited Programmes (PUPs) were examined to determine participant outcomes related to adjustment to acquired brain injury (ABI) and to investigate the contribution of the group work component of the programme. A mixed qualitative and quantitative longitudinal design was employed with 14 participants and a demographically matched comparison group of eight respondents. Results showed significant and sustained improvement in participants' subjective quality of life. Analysis of interview data provided insights into how the programmes provided opportunities for participants to engage in key tasks of adjustment to injury. Participants who regularly attended the follow-up group stages of the programmes made significantly greater gains than non-attendees. In conclusion, the combination of outdoor experiential education with extensive group work found in the PUPs is likely to represent a unique and powerful approach for addressing many of the issues associated with the process of psychosocial adjustment to injury and restoring quality of life, following ABI. PMID- 15666571 TI - Video analysis of head blows leading to concussion in competition Taekwondo. AB - PRIMARY OBJECTIVES: To analyse the situational and contextual factors surrounding concussions and head blows in Taekwondo. METHODS AND PROCEDURES: Prospective design. Direct observation, subject interview and videotape recording used. A total of 2328 competitors participated in the 2001 tournament, South Korea. All matches were recorded on videotape. All recipients of head blows were interviewed by athletic therapists and the researcher immediately after the match. The videotapes of concussions and head blows were analysed. RESULTS: A total of 1009 head blows including concussions were analysed. Head blows and concussions were most evident when the attacker was situated in a closed stance and received a single roundhouse kick. The most frequent anatomical site of the head impact was the temporal region. CONCLUSIONS: The frequency of head blows and concussions is high in Taekwondo. Development of blocking skills, safety education, rigorous enforcement of the competition rules and improvement of head-gear are recommended. PMID- 15666572 TI - The therapeutic challenges of dual diagnosis: TBI/SCI. AB - Rehabilitation centres provide therapeutic intervention to both patients with traumatic brain injury (TBI) and spinal cord injury (SCI). Patients with a dual diagnosis of TBI and SCI present a challenge to the rehabilitation professional. Previous studies indicate an incidence of concomitant TBI and SCI to be between 24-59%. Many research articles discuss how a diagnosis of TBI is often missed during the medical examination of a person with a suspected SCI. This article will focus on the implications of dual diagnosis encountered during the rehabilitation process. A case study is used to highlight the challenges presented by the cognitive impairments of a TBI in combination with the motoric deficits of SCI. Management strategies utilised by the rehabilitation team are discussed. PMID- 15666574 TI - Type 2 diabetes mellitus is becoming the most common type of diabetes in school children. AB - Obesity, now an epidemic in the USA, northern Europe, and Italy, is associated with several co-morbidities that shorten life expectancy, in particular type 2 diabetes mellitus (T2DM), arterial hypertension, and hyperlipidemia. The impact of obesity on mortality is evident in all ages, and is especially strong in young persons. Obesity, especially visceral obesity, associated with a sedentary lifestyle, is among the strongest risk factors for T2DM, and a diagnosis of T2DM seems to increase linearly as a function of duration of obesity. The pathogenesis of T2DM is based on a dual defect, i.e. increased insulin resistance coupled with defective insulin release. The main abnormality in obesity is increased insulin resistance, while insulin release, even though defective compared with body needs, is usually abundant. The incidence of obesity among children aged 6-16 years is now even greater than that among adults: in Italy, figures up to 30% have been reported. As in adults, obesity is a cause, among children, of arterial hypertension, left ventricular hypertrophy, hyperlipidemia, non-alcoholic-steato hepatitis, sleep apnea syndrome (SAS), and orthopedic, psychological, and social problems. Together with an increase in body weight, there is an increase of visceral fat. Obesity in children has also led to a tremendous increase in the prevalence of impaired glucose tolerance (IGT); the percentages span from 25% in a multiethnic cohort in the USA, to 4% in Italian Caucasians. Management of obesity and of T2DM in children has to face the issue of poor compliance; there is consensus that dietary treatment of obese T2DM children is a failure, so that drugs are required; the only drug evaluated in a formal trial is metformin, that behaves in terms of efficacy and of minor side effects as in adults. In conclusion, obesity in children is not pure obesity, but is accompanied by co morbidities that cluster to form the "metabolic syndrome" just like in the adults. If this epidemics continues and is not properly challenged, in the next decades we will face an epidemic of early cardiovascular morbidity and mortality. PMID- 15666573 TI - Amantadine for neurobehavioural deficits following delayed post-hypoxic encephalopathy. AB - Delayed post-hypoxic encephalopathy is an uncommon but potentially debilitating consequence of hypoxic-ischemic brain injury. This condition is characterized by delayed neurological deterioration days-to-weeks after an initial partial or complete recovery from hypoxic-ischemic brain injury. The course of recovery from this condition is highly variable, ranging from rapid and fatal progression over several weeks to delayed but occasionally complete recovery. There are no reports describing neurorehabilitative, including neuropharmacologic, interventions for persons with persistent neurological and/or neurobehavioural deficits following delayed post-hypoxic encephalopathy. This study describes the case of a 24-year old male who developed delayed post-hypoxic encephalopathy following an unintentional methadone and diazepam overdose and who demonstrated cognitive and neurobehavioural improvements during treatment with amantadine HCl hydrochloride in a single-case, open-label design. A brief review of the literature regarding this condition, its treatment and suggestions for further study are presented. PMID- 15666575 TI - What do herbalists suggest to diabetic patients in order to improve glycemic control? Evaluation of scientific evidence and potential risks. AB - In the course of 12 continuing education seminars given in different regions of Italy in 2001, we distributed a questionnaire to all the attending herbalists asking information about the herbal remedy and dietary supplement they mainly recommended to subjects who required a "natural" treatment to control glycemia. We distributed 720 questionnaires and we received 685 completed ones. We have compiled a short review on the efficacy and safety of the 10 most frequently advised products for each category. The 10 more frequently suggested herbal remedies were gymnema, psyllium, fenugreek, bilberry, garlic, Chinese ginseng, dandelion, burdock, prickly pear cactus, and bitter melon. The 10 most frequently recommended dietary supplements were biotin, vanadium, chromium, vitamin B6, vitamin C, vitamin E, zinc, selenium, alpha-lipoic acid, and fructooligosaccharides. The majority of the products recommended by Italian herbalists may be efficacious in reducing glycemia. If a diabetic patient is already assuming products that even slightly reduce glycemia, we risk to underestimate the level of glucose intolerance, while if the patient stops the complementary treatment after initiating pharmaceutical therapy, in the subsequent visit we may underestimate the effect of our prescription. Therefore, if doctors are to have a role in gate-keeping or advising patients about complementary and alternative medicine, they need to be familiar with this type of medicine. If they choose otherwise, then the provision of complementary and alternative medicine will continue to be patchy and largely outside the conventional care framework, perhaps through a growing network of parallel care providers involving a large number of non-medically qualified practitioners, who patients will continue to access directly. PMID- 15666576 TI - Techniques to evaluate erythrocyte deformability in diabetes mellitus. AB - Using several rheological techniques, we examined erythrocyte deformability in different groups of diabetic subjects. The macrorheological techniques used for this evaluation were respectively whole-blood filtration, filtration of erythrocyte suspensions, polyviscosimetry and diffractometry. Whole-blood filterability, at a negative pressure of 20 cm water, was decreased in type 2 diabetics; no difference was evident at a negative pressure of 10 cm water. The filtration of erythrocyte suspensions at low haematocrit (5%) did not show differences between normal and diabetic subjects. Polyviscosimetry, which explores the filterability of erythrocyte suspensions at high haematocrit (80%) through wide pores, demonstrated an impaired behaviour especially in type 2 diabetics. Diffractometry, which measures erythrocyte elongation induced by a defined shear stress through the diffraction pattern of a laser beam, showed an alteration in type 1 diabetic subjects. The microrheological methods employed for this evaluation were those based on fluorescence spectroscopy. Labeling intact red blood cells with fluorescent probes, we determined the membrane dynamic properties and using these techniques we found a reduction of erythrocyte membrane fluidity and a decrease of red cell membrane protein lateral mobility. PMID- 15666577 TI - Effects of leptin, acetylcholine and vasoactive intestinal polypeptide on insulin secretion in isolated ob/ob mouse pancreatic islets. AB - Obesity is often accompanied by hyperleptinemia, hyperinsulinemia, and an increased parasympathetic tone. Obese-hyperglycemic mice (Umea ob/ob) have functional leptin receptors and a raised parasympathetic tone. We studied insulin release in islets isolated from 9-month-old severely obese ob/ob mice. Leptin (0.5-18 nM) did not affect insulin release together with 2.8-20 mM glucose. Leptin (18 microM) had no effect in the presence of low glucose (2.8-5.5 mM), but increased insulin secretion in islets challenged with 11.1 or 16.7 mM glucose. Leptin at 18 microM increased insulin secretion stimulated by the parasympathetic neurotransmitters acetylcholine (ACh; 10 microM) or vasoactive intestinal peptide (VIP; 10 nM), and by 5 mM theophylline or 2.5 microM forskolin. Overnight culture increased the effect of 18 microM leptin, but no effects were observed with 18 nM leptin. Pretreatment of islets with phorbol 12-myristate 13-acetate (PMA) did not suggest any involvement of protein kinase C. In summary, a high concentration of leptin stimulates insulin release in the presence of stimulatory concentrations of glucose alone and with parasympathetic neurotransmitters. Hyperleptinemia and increased parasympathetic stimulation may in part cause the hyperinsulinemia observed in obesity. This may aggravate insulin resistance and the abnormal metabolism in diabetes mellitus. PMID- 15666578 TI - Evaluation of visceral adipose accumulation in Japanese women and establishment of a predictive formula. AB - Abdominal obesity is a known risk factor for diabetes-related diseases. This study aimed to establish a formula to predict visceral abdominal fat area on the basis of simple clinical and anthropomorphic parameters easily measured in the clinic. We determined visceral fat (V) and subcutaneous fat (S) areas in 115 Japanese women using the standard procedure based on computed tomography (CT) at umbilical level. Furthermore, we measured clinical and anthropometric parameters including height, weight, waist circumference, hip circumference, skin fold thickness and body fat percentage. In 115 subjects, V area was 87.8+/-52.5 cm2 and S area was 221.1+/-99.7cm2. Abdominal obesity is diagnosed in Japan as a V area > or =100 cm2; on this basis 42 women (37%) had abdominal obesity. The prevalences of diabetes and related diseases were significantly higher among women with abdominal obesity. By simple regression analysis, V and S areas significantly correlated with anthropometric parameters: in particular, V area correlated with waist circumference (r=0.745, p<0.01) and S area with body mass index (r=0.793, p<0.01). However, these parameters were not sufficient to predict V area. By multiple regression analysis using simple parameters, we established the following formula to predict visceal fat: V area = 159.475 + 1.023(age) - 2.119(height) + 1.454(body weight) + 2.841(waist circumference) - 1.208(hip circumference) (r=0.812, p<0.01). The V area calculated by formula correlated (r=0.761) with that determined by CT in a second age-matched group of 31 Japanese women. The present study confirms that visceral adipose tissue is closely associated with type 2 diabetes mellitus, dyslipidemia and hypertension, and generated a formula to predict visceral adipose tissue accumulation. PMID- 15666579 TI - Soluble adhesion molecules are not involved in the development of retinopathy in type 2 diabetic patients. AB - Raised serum levels of adhesion molecules are believed to reflect endothelial activation and may contribute to the development of diabetic vascular complications. The aim of this study was to clarify the association between soluble adhesion molecules levels and retinopathy in type 2 diabetic patients. Levels of soluble E-selectin, ICAM-1 and VCAM-1 were measured by enzyme-linked immunosorbent assay (ELISA) in 47 type 2 diabetic patients classified in two subgroups according to the presence (n=34) or absence (n=13) of retinopathy as determined by fundus ophthalmoscopy; 22 control subjects were also studied. Soluble E-selectin levels were significantly elevated in both diabetic subgroups compared to control subjects (p<0.01), while no significant difference was found in sICAM-1 and sVCAM-1 levels. However, sE-selectin, sICAM-1 and sVCAM-1 levels were comparable in diabetic subgroups. The progression of retinopathy was not associated with an increase in soluble adhesion molecules levels. Stepwise multiple regression analysis revealed that only diabetes duration and microalbuminuria were independent determinants of retinopathy (p<0.01). Our results confirm the contribution of endothelial activation in the development of diabetic complications as indicated by increased levels of soluble adhesion molecules. However, a direct implication of adhesion molecules in the pathogenesis or progression of type 2 diabetic retinopathy cannot be supported. PMID- 15666581 TI - Pattern and determinants of dyslipidaemia in type 2 diabetes mellitus patients in Kuwait. AB - We conducted a clinical study to assess the pattern of dyslipidaemia in type 2 diabetic patients and to examine the demographic and clinical factors associated with dyslipidaemia. The study population comprised 206 consecutive type 2 diabetic patients attending the out-patient clinic at a major hospital in Kuwait. Clinical history and physical examination were done and fasting blood samples were taken to determine HbA1c and lipid levels. American Diabetes Association criteria were applied to define clinical targets for lipid levels and coronary heart disease risk categories. Stepwise multiple linear regression was conducted to identify the demographic and clinical factors associated with lipid levels outside of the clinical target. The large majority of the patients were either over-weight (32%) or obese (57%); the mean BMI was 32.6 kg/m2. Serum total cholesterol, LDL-cholesterol, and triglycerides were above optimal levels in 67%, 86%, and 25% of patients, respectively. For HDL-cholesterol, 63% of men and 71% of women had values below the corresponding optimal level. Only 14 patients (6.8%) had all three lipid values within the respective target level. The percentages of patients with one, two, or all three lipid values outside of target were 31%, 46%, and 16%, respectively. The most frequent (41%) pattern of dyslipidaemia was a combination of LDL-cholesterol level above target with HDL cholesterol level below target; the second most common pattern was an isolated increase in LDL-cholesterol, observed in 21% of the patients. In the stepwise regression analyses, glycaemic control was strongly associated with dyslipidaemia (i.e. high total- and LDL-cholesterol and triglycerides); female gender were associated with low HDL-cholesterol. Kuwaiti type 2 DM patients have a high prevalence of dyslipidaemia and obesity. Weight reduction, increased physical activity, improved glycaemic control, and increased HDL-cholesterol levels, along with reduced LDL-cholesterol, should be important goals of therapy in these patients to reduce the risk of coronary heart disease. PMID- 15666580 TI - Early changes in glial reactivity and lipid peroxidation in diabetic rat retina: effects of melatonin. AB - Diabetes mellitus, the most common serious metabolic disorder, is characterized by functional and structural changes in the peripheral and central nervous systems. Glial cells provide structural and metabolic support for retinal neurons. During diabetes, one of the early pathogenic events is retinal glial reactivity. We studied the effects of melatonin, which is known to reduce oxidation-based neurotoxicity, on glial reactivity and lipid peroxidation in the retina of diabetic rats. Diabetes was induced by a single injection of streptozotocin (STZ), and these diabetic rats were treated daily either with melatonin (10 mg/kg) or saline vehicle. After 6 weeks of diabetes, we determined the extents of lipid peroxidation and glial reactivity in retina. Lipid peroxidation, measured on the basis of malondialdehyde and 4-hydroxyalkenals concentrations, was increased in diabetic rats (p<0.01) and this increase was prevented by melatonin treatment (p<0.05). Furthermore, gial reactivity, determined immunohistochemically from the levels of glial fibrillary acid protein (GFAP), was also increased significantly (p<0.01). Melatonin administration partially prevented this increase in GFAP content (p<0.05). In conclusion, glial reactivity is an early pathogenic event in diabetic retina and both reactive gliosis and accumulation of malondialdehyde and 4-hydroxyalkenals are prevented by melatonin supplementation. PMID- 15666582 TI - From genomic advances to public health benefits: the unbearable lightness of being stuck. AB - Genetic determinants of common human diseases are still poorly understood. Due to large investments, many small successes have been made and the research field is rapidly expanding. However, genetic susceptibility variants showing repeatable associations with common diseases are usually of small effect. They are therefore unlikely to individually explain substantial share of disease burden in any community or provide new insights into disease pathogenesis that could lead to development of new drugs effective in considerable portion of the disease cases in a population. Genetic architecture of common diseases is beginning to reveal an incredible diversity of potential genetic causes that act through somewhat limited number of mechanisms with important contribution of environmental interactions. In light of these findings, we present current understanding of genetic architecture of a spectrum of human diseases. We address the encountered problems in susceptibility gene identification, review the success of leading gene identification strategies and discuss current prospects for translating genomic advances into measurable public health benefits. PMID- 15666583 TI - Environmental stress and adaptational responses: consequences for human health outcomes. AB - With the dramatic pace of modernization of the world's population, human adaptation as a theoretical construct and paradigm will likely become a focal scientific issue involving scientists from many disciplinary areas during the 21st Century. Macro and micro environments are in rapid flux and human populations are exposed to rapid change. The concept of adaptation, at least in the field of biological anthropology and human biology, will likely remain tied to evolutionary processes and concepts of selection and fitness. In this paper, we discuss the theoretical constructs of adaptation and adaptability and select three current examples from our ongoing research that involve studies of adaptation and evolutionary processes in modernizing populations in different locations worldwide. PMID- 15666584 TI - Regional and social differences in body mass index, and the prevalence of overweight and obesity among 18 year old men in Austria between the years 1985 and 2000. AB - The BMI data of four birth cohorts of totally 180,716 male 18 year old Austrian conscripts were documented in 5 year intervals starting 1985 and ending 2000 in order to analyze regional and social variety and a time trend of stature, body weight and BMI as well as the prevalence rates of overweight (defined as a BMI > 25.00) and obesity (BMI > 30.00). At first a marked time trend in BMI and overweight/obesity prevalence rates was found. Over the 15 years of investigation BMI increased significantly and the variation of BMI distribution increased too. The impact of educational level on BMI and the prevalence of overweight and obesity was also statistically significant. With increasing educational level BMI and the prevalence rates of overweight and obesity decreased significantly. Furthermore a significant regional variety in BMI and the prevalence rates of overweight and obesity were found. BMI and the prevalence rates of obesity and overweight decreased significantly from the Eastern part of Austria to the Western part. Both observations, e.g. the social and regional variety of obesity and overweight prevalence, were true of all 4 birth cohorts. From these results we can conclude that obesity and overweight represent an increasing health problem among young Austrian males. This is especially true of young men of low social status living in the eastern part of Austria. PMID- 15666585 TI - Age and sex variation in measures of body composition among the elderly Bengalee Hindus of Calcutta, India. AB - In the present cross-sectional study we examined 332 (171 men and 161 women) elderly (60 years and above) urban Bengalee Hindu resident in south Calcutta, India. Individuals were selected by random sampling procedure using local voter's registration list. Skin folds measures were used to compute body composition measures among them. There existed significant sex differences in various anthropometric body composition measures. Age had significant (p < 0.001) negative association with all anthropometric body composition measures namely percentage of body fat (PBF), fat mass (FM), arm muscle circumference (AMC), arm muscle area (AMA) and arm fat area (AFA) in both sexes. Fat free mass (FFM) in contrast had negative but not significant age impact. Regression analyses demonstrated that age had explained substantial amount of variance of PBF (men = 32%; women = 18.2%), FM (men = 18.2%; women = 12.8%), AMC (men = 23.4%; women = 19.2%), AMA (men = 22.2%; women = 10.2%) and AFA (men = 34%; women = 31%) in both sexes. Two-way ANOVA revealed age-sex interaction only had significant effect on FFM. The present investigation vindicated that there is a significant inverse age trends in anthropometric body composition measures among the Bengalee Hindus. Moreover, there existed sexual dimorphism in the effect of age on various anthropometric body composition measures. PMID- 15666586 TI - Mother's age effect on the boys' anthropometric properties in the Region of Tuzla (Bosnia and Herzegovina). AB - Our objective was to establish if mother's age affects the anthropometric properties their 11 and 16 years old sons, and if the boys' BMI is satisfactory. Cross-sections, questionnaires, and statistics were used for data processing BMI (kg/m2) was presented according to the scale Quetelet's index nutritional status. Twelve anthropometric properties were researched. It was established that mother's age affects anthropometric properties in adolescent boys. The younger mothers' boys come to puberty with significantly higher mean values for most anthropometric parameters. In the course of adolescent jump, older mothers' boys have significantly higher anthropometric measures in comparison to their age boys delivered by younger mothers. After the completion of their intensive growth and development anthropometric measures in both category boys are equal or insignificantly higher in younger mothers' sons. Mother's age affects their sons' anthropometric properties of growth and development, particularly in prepuberty and puberty. After puberty that effect is not significant. According to BMI, the boys coming from the researched region make the group of underfed children. That is probably the outcome of bad living conditions, irregular and insufficient diet, stress, etc. in the course of four year aggression on Bosnia and Herzegovina. PMID- 15666587 TI - Secular trend in stature and age at menarche among Punjabi Aroras residing in New Delhi, India. AB - The study was conducted on Punjabi Arora girls (n = 159) and their mothers to see the phenomenon of secular trend in stature and age at menarche. An increase in stature and decrease in age at menarche was reported when the data of daughters was compared with that of their mothers thereby indicating secular trend in these two parameters. Better living conditions, improved nutrition and medical facilities, changes in environmental and socio-economic factors may account for increase in stature and early biological maturation. PMID- 15666588 TI - Twin seasonality in a rural Catalonian population. AB - The seasonality of twinning in the Spanish populations has not been studied until now. Differences between seasonal distribution of the twin conceptions and those of the single births have been observed in other populations. The aim of this work is to explore the frequency of twinning in a rural population from Catalonia during the nineteenth century, as well as the seasonality patterns characterizing each of the twinning types. Data corresponding to all births recorded at Tortosa (South Catalonia) from 1801 to 1900 have been analyzed in order to study the twinning distribution. The distribution of the moving averages of the monthly rates of twins shows a peak in autumn. Twinning distribution differs from the total births' distribution in Tortosa. This fact is very clear in the case of unlike-sexed twins that have their greater incidence in the last quarter of the year, while the total maternities have their peak in the first one. PMID- 15666589 TI - Inbreeding and susceptibility to osteoporosis in Croatian island isolates. AB - The aim of this study was to investigate a recessive genetic component in susceptibility to osteoporosis (OP) by comparing its prevalence in isolated villages of three Croatian islands: Brac, Hvar and Korcula with different levels of inbreeding. A random sample of 20-30% adults from 14 villages was obtained, including a total of 1,389 examinees. The average inbreeding coefficient (F) of examinees from each village population was estimated using Wright's path method (based on genealogical information). The morphometry of the metacarpal bones was performed on hand-wrist radiographs of both hands in all examinees. OP was defined as values of cortical index smaller than 2 standard deviations based on distribution of values in examinees of the same sex under 45 years of age. Mean values of cortical index (CI) and prevalence of OP (both standardized by age and weighted for the sample size) in each village were correlated to the mean inbreeding coefficient (F). The coefficient of correlation (r) between F values and CI was -0.28 in males (p = 0.08) and -0.42 in females (p = 0.005), and between F and OP prevalence 0.32 in males (p < 0.001) and 0.43 in females (p < 0.001). These results indicate a trend of increased susceptibility to osteoporosis with increasing level of inbreeding in isolated communities of Croatian islands. PMID- 15666591 TI - The -429 T/C and -374 T/A gene polymorphisms of the receptor of advanced glycation end products gene (RAGE) are not risk factors for coronary artery disease in Slovene population with type 2 diabetes. AB - Receptor for advanced glycation end products (RAGE) plays a role in atherosclerosis in diabetics. There are two functional polymorphisms in the promoter of the RAGE gene (-429T/C and -374T/A). The aim of this study was to look for a relationship between the -429T/C and the -374T/A gene polymorphisms of the RAGE gene and the development of coronary artery disease (CAD) in the Slovene population with type 2 diabetes of duration longer than 10 years. One hundred and sixty-eight subjects with diabetes and CAD were compared to 241 diabetic subjects without CAD. The -429T/C and the -374T/A RAGE genotype distributions in patients with CAD (-429T/C: CC: 3%, TC: 31%, TT: 66.0%; -374T/A:AA: 7.7%, TA: 48.2%, TT: 44.1%) were not significantly different from those in patients without CAD (-429 T/C: CC: 1.7%, TC: 26.1%, TT: 72.2%; -374T/A: AA: 11.2%, TA: 43.2%, TT: 45.6%). Our study failed to demonstrate an association between either the -429T/C or the 374T/A gene polymorphism of the RAGE gene and CAD in the Slovene population with type 2 diabetes of duration longer than 10 years. PMID- 15666590 TI - Ace Alu insertion polymorphism in Croatia and its isolates. AB - Alu elements are a family of interspersed repeats in the genome propagating by retroposition into new chromosomal locations. Alu insertion in Ace gene is known to be polymorphic (presence/absence of Alu element) in worldwide populations and as such serves as marker for population structure analyses. In this study we examined the distribution of genotypes and allele frequencies of this polymorphism in general Croatian population and its two isolates (the island of Hvar and the coastal region of the Middle Dalmatia) and related them to the level of endogamy as an indicator of inbreeding in these populations. Results showed that these three population groups are different with respect to Ace Alu polymorphism. The endogamy was highest on the island of Hvar. With the increase of endogamy a decrease in heterozigosity was observed. The same trend was observed for the frequency of insertion allele. Its frequencies in the village subpopulations of two studied isolates are subject to genetic drift due to small population sizes and high levels of endogamy. This in turn causes genetic differentiation among villages that is observed to be higher on the island of Hvar than in the coastal region. In the worldwide perspective, the Ace Alu insertion allele frequency of 50.6% in the general Croatian population falls within the range of other European populations. PMID- 15666592 TI - The ScaI gene polymorphism of the atrial natriuretic factor and essential arterial hypertension in childhood. AB - In order to investigate the contribution of the atrial natriuretic factor (ANF) gene in pathogenesis of essential arterial hypertension (EAH), we analyzed the ScaI gene polymorphism of the ANF gene in a group of children with EAH. Fifty eight children, aged 8-19 years, with the diagnosis of EAH were included in the association study and were compared to 57 subjects with normal blood pressure (the control group). Arterial hypertension was defined as systolic/diastolic blood pressure higher than the 95th age-gender-height percentile of the adopted reference values. We failed to demonstrate an association between the ScaI ANF gene polymorphism and EAH in childhood (OR = 2; 95% CI 0.9-4.2; p = 0.07), however, we provided evidence of an interaction between the ScaI ANF gene polymorphism and obesity defined as BMI over the 85th percentile (OR = 13.1; 95% CI 1.6-106; p < 0.001). PMID- 15666593 TI - The effect of 3-weeks stationary cardiac rehabilitation on plasma lipids level in 444 patients with coronary heart disease. AB - The aim of this study was to investigate the effect of 3-weeks stationary cardiac rehabilitation on plasma lipids level in patients with CHD. The study included 444 consecutive patients (364 male and 80 female, mean age 58 +/- 9 year) with CHD who underwent 3-weeks stationary cardiac rehabilitation. Patients were divided into groups depending on their baseline levels of cholesterol and medication therapy: patients with normal (< 5 mmol/L, group I, 129 patients) and elevate plasma level of Total cholesterol (> 5 mmol/L, group II, 315 patients) and subgroups Ia and IIa (with statin in therapy), Ib and IIb (without statin in therapy). After 3-weeks cardiac rehabilitation, the levels of Total cholesterol 5.75 +/- 1.34 vs. 5.17 +/- 1.08 mmol/l; p < 0.001, triglycerides 2.04 +/- 1.33 vs. 1.81 +/- 1.06 mmol/L; p = 0.004, LDL-cholesterol 3.77 +/- 1.14 vs. 3.21 +/- 0.96 mmol/L; p < 0.001 were significantly lower while the level of HDL cholesterol 0.94 +/- 0.28 vs. 0.99 +/- 0.27 mmol/L; p = 0.008 were significantly higher in comparison with the baseline values. Furthermore, we found significant changes in lipid profile at the end of rehabilitation in each group of patients compared with the baseline values. There were no significant differences in plasma lipids level between group of patients with or without statin in therapy at the end of rehabilitation. The results of this study suggest that moderate regular physical activity and diet alone or in combination with hypolipidemic drugs already after 3 weeks have a favourable effect on plasma lipids level and should be propagate in the prevention of CHD. PMID- 15666594 TI - Effects of troglitazone and insulin on glucose production in cultured hepatocytes isolated from rats on high fat diet. AB - Increased dietary fat intake in general, and saturated fat specifically, will lead to the impairment of insulin action. The aim of this study was to find out the changes in hepatic glucose output in dependence of fat diet and a possible direct action of insulin and trogitazone in hepatocytes. Hepatocytes were isolated by a collagenase perfusion technique and cultured for 24 h in M 199 serum-free medium. The glucose production in hepatocytes isolated from rats on high fat diet (unsaturated fat) was 79% higher compared to control and even 139% higher than in rats on high-fat diet (saturated fat). Troglitazone significantly decreased the glucose production in hepatocytes obtained from rats on unsaturated fat diet. The troglitazone in presence of insulin totally normalized glucose production but also only in hepatocytes obtained from rats on unsaturated-fat diet. The troglitazone showed an insulinomimetic as well as insulin-sensitizing effect but only in rats on unsaturated-fat diet. PMID- 15666595 TI - Effects of rHuEPO treatment on red blood cell osmotic resistance. AB - Red blood cell osmotic resistance (RBCOR) is defined as resistance to osmotic changes in cell integrity after their exposure to hypotonic saline solution. The investigation examined the effect of rHuEPO on RBCOR in hemodialysed patients. The study included 58 patients aged 49 +/- 14 years, treated by hemodialysis for 59 +/- 43 months on average. Half of the patients received rHuEPO for anemia correction. RBCOR was determined in all patients as 3 values: hemolysis start point (HSP), hemolysis end point (HEP) and middle osmotic resistance (MOR). The patients underwent laboratory checkup for parameters characteristically changed in the uremic syndrome. In the control group of healthy subjects (n = 16) RBCOR was only determined. No differences were found in the average values of HSP, HEP and MOR between the rHuEPO treated group of patinets and the untreated group. Compared to healthy individuals, the hemodialysed patients displayed significantly higher values of HSP, HEP and MOR. The only one significant correlation of RBCOR and routine laboratory features was found between MOR and predialytic serum concentrations of calcium (r = 0.28, p < 0.05) and hydrogen ions (r = 0.37, p < 0.05). Our results suggest that the administration of rHuEPO does not affect RBCOR in hemodialysed patients, that RBCOR is not always reduced in this population and that it correlates with a small number of laboratory parameters characteristic for the uremic syndrome. PMID- 15666596 TI - 5, 10-Methylenetetrahydrofolate reductase (MTHFR) 677 C --> T genetic polymorphism in 228 Croatian volunteers. AB - 5, 10-Methylenetetrahydrofolate Reductase (MTHFR) is one of the key enzymes in the metabolism of homocysteine, where it catalyses its remethylation. The autosomal recessive bp 677 C --> T mutation in the MTHFR gene leads to the substitution of valine for alanine. Individuals who are homozygous for this C677T mutation exhibit a decreased specific activity and increased thermolability of this enzyme. This leads to increased plasma levels of homocysteine, which is a known risk factor for atherosclerosis and various manifestations of the atherosclerotic disease. The aim of this study was to find out the distribution and frequency of this mutation in the general Croatian population. A group of 228 volunteers (175 males and 53 females) has been analyzed for the MTHFR polymorphism, which revealed the following distribution: 105 (46.05%) individuals were without mutation (C/C), 102 (44.74%) were heterozygous (C/T) and 21 (9.21%) homozygous (T/T). These findings are within the results of studies on other European populations. PMID- 15666597 TI - Metabolic evaluation of urolithiasis patients from eastern Croatia. AB - Metabolic parameters were determined in fasting blood serum, fasting first morning urine, and 24-hour urine of male patients with recurrent calcium oxalate stones (N = 26, age 39.1 +/- 6.2 years) as well as in male healthy controls (N = 18, age 35.0 +/- 7.1 years), recruited from the eastern part of Croatia. The 24 hour urinary calcium excretion was significantly higher (p < 0.01) for patients (5.6 +/- 2.5 mmol) than for controls (3.7 +/- 1.9 mmol), but potassium excretion was higher (p < 0.01) for controls (74.5 +/- 33.8 mmol) than for patients (49.2 +/- 15.7 mmol). The mean ionic activity product of calcium and oxalate ions, IAP(CaOx), calculated from the fasting first morning urine parameters, was 25% higher for patients than for controls, but the difference was not statistically significant (p > 0.05). Very strong correlation (r = 0.97) was obtained between IAP(CaOx) values and calculated Ogawa indices that were recommended for estimating the potential risk for calcium oxalate stone formation. PMID- 15666598 TI - Renal vascular resistance in glomerular diseases--correlation of resistance index with biopsy findings. AB - Duplex Doppler sonography has been recognized as a noninvasive method to evaluate hemodynamic features of renal blood in renal and intrarenal arteries in patients with various renal diseases. The significance of duplex Doppler sonography in the evaluation of renal vascular resistance in glomerular diseases has not yet been clearly determined. The aim of the present study was to evaluate renal vascular resistance in patients with glomerular diseases by measuring intrarenal arterial resistance (RI) and to correlate RI with renal functional tests and other clinical and laboratory data. The Doppler parameters were also correlated with histopathological findings in the kidney which underwent the percutaneous biopsy. Duplex Doppler sonography was used to measure RIs in intrarenal arteries in 50 patients with glomerular diseases and 60 age-matched control subjects. The renal vascular resistance index (RI) was determined by the use of Doppler sonography. The mean RI in 50 patients with glomerular diseases was 0.68 +/- 0.09, which was statistically significantly higher than in 60 control subjects (the mean RI was 0.596 +/- 0.035). In a group of patients with membranoproliferative glomerulonephritis the mean RI was 0.817 +/- 0.624 which was statistically significantly higher than in other groups of glomerulonephritis. The renal vascular (resistance) RI significantly correlated with serum creatinine, creatinine clearance and beta2 microglobulin. Qualitative duplex sonography measure of renal arterial resistance-resistive index does not appear to be reliable in distinguishing different types of glomerulonephritis. PMID- 15666599 TI - Adherence ability of Pseudomonas aeruginosa strains isolated from patients with cystic fibrosis to two different epithelial cell lines. AB - The ability of 59 wild-type strains of Pseudomonas aeruginosa to adhere to the HeLa and Buffalo Green Monkey Kidney (BGMK) cells was investigated. Twenty strains were isolated from sputa of cystic fibrosis patients, while 19 strains were isolated from tracheal aspirates and 20 from bronchial secretions of patients without cystic fibrosis, and they were used as a control group of strains. The statistically significant difference between adherence ability of strains was observed (p < 0.01). While most of the tracheal and bronchial isolates were hyperadhesive (51-110 bacteria per cell) most of the cystic fibrosis isolates adhered poorly to the HeLa and BGMK cells (1-10 bacteria per cell). The bacterial binding to the cells was blocked when bacteria were incubated at 80 degrees C for 20 min before the adherence assay. These results indicate that alginate is not involved in the adherence of P. aeruginosa to the used epithelial cell lines, and, because of that, mucoid strains isolated from persistently colonized cystic fibrosis patients showed poor adherence ability. PMID- 15666600 TI - Developmental potential of mouse embryos without extraembryonic membranes in modified organ culture. AB - The long-term stationary culture of postimplanatation embryos without extraembryonic membranes is a method to assess their developmental potential in vitro. The method was almost exclusively used on rat embryos, while mouse embryos were considered unsuitable due to their poor differentiation. In present study the postimplantation mouse embryos were used to verify potential of this method in mice. In addition, the course of in vitro differentiation was compared to embryo development in situ. Embryos were cultivated for maximum of 14 days and morphology and differentiation was analysed on serial semithin sections. Although anatomical relationships were lost from the beginning of the cultivation, the differentiation was only delayed, and the developmental potential after long-term culture was comparable to those observed in rats. Therefore the advantages of long-term cultivation could be utilized to analyse the differentiation of numerous lines of genetically modified mice with impaired postimplantation development. PMID- 15666601 TI - Smoking habits, signs of chronic diseases and survival in inland and coastal regions of Croatia: a follow-up study. AB - Aim of the study was to estimate, the relationship between survival, smoking habits, and the results of medical examinations in inland and coastal regions of Croatia. Age and sex stratified sample of general population (1,571 men and 1,793 women, 37-56 years old in 1972; followed in 1982: N = 1,090 men; 1,325 women and/or 1972-1999 controlling vital status). Relative risks and 95% confidence limits were estimated using Cox regression in the model with time dependent covariates, separately by sex. In all regions, the proportion of male smokers decreased between 1972 and 1982. The proportion of female smokers increased, differently in urban and rural regions. During the follow-up between 1972 and 1999, 568 deaths were recorded among men and 382 among women. In men, in addition to age, significant hazards of death were the number of smoked cigarettes per day, body mass index, sedative intake, vital lung capacity (FVC), 100FEV1/VC, systolic blood pressure, electrocardiogram, history of heart attack, and region. In women, in addition to age, significant predictors were the number of smoked cigarettes per day, systolic blood pressure, electrocardiogram, history of heart disease (excluding coronary diseases), and region. Survival relative risk increased with each additional smoked pack of cigarettes by 2.4% in women and 1.3% in men. Regional differences vs. smoking habit were observed. These data emphasize the need for prevention of smoking. PMID- 15666602 TI - The ventilatory patterns of the left upper lobe of lung. AB - Having knowledge on models of the bronchial tree branching, is of a special interest for clinical and surgical pulmology, because the hemilobectomy, segmentectomy and subsegmentectomy are always determined by intralobar, intrasegmental and intrasubsegmental bronchial ramification. Investigations were performed on 100 lungs of children and adults of both sexes, one day to 85 years old, randomly chosen. There are two main types of branching of the left upper lobe bronchus: the bifurcation pattern as dominant model in 74% and the trifurcation model found in 26%. Out of 100 lungs studied, 21 lungs had the ventilatory variations of the bronchopulmonary segments. The classification and categorization of the ventilatory of bronchopulmonary segments of the left upper lobe of lung were made. This classification contains 5 categories and 8 subcategories. PMID- 15666603 TI - Lung function changes in pleural asbestosis. AB - The aim of this study was to examine the relationship between radiographically detectable pleural changes and lung function in pleural asbestosis. One hundred and twenty chrysotile asbestos-exposed workers were enrolled in this retrospective study. For each examinee the length of asbestos exposure and the degree of dust cover at the workplace were assessed as well as the radiological and functional tests has been performed. The examinees were divided into two groups based on radiologically detectable changes: a) group with pleural changes (29%) and b) group without perceived pleural changes (71%). The obtained results indicate association between the length of asbestos exposure, pleural changes and the impairment of lung function. PMID- 15666604 TI - Respiratory findings in art students. AB - Art students are exposed to many noxious agents during their training. We studied respiratory findings in a cohort of the 117 art students in order to investigate the potential effects of these toxic agents in the art student's environment. A group of 88 medical students matched for age, sex and smoking, not exposed to known environmental pollutants were studied as controls for respiratory symptoms. Respiratory symptoms acute and chronic were evaluated by modifying the British Medical Research Council questionnaire. Lung function studies were performed with a spirometer (Jaeger, Germany) measuring maximum expiratory flow-volume (MEFV) curves. Significantly higher prevalences of most of the chronic respiratory symptoms were recorded in art compared to medical students (p < 0.05). Art students who were smokers had significantly higher prevalences of many of the chronic respiratory symptoms than nonsmoking art students. High prevalences of acute symptoms related to the study environment were recorded for art students. Odds ratios in male art students were significant for chronic cough, chronic phlegm and chronic bronchitis for the risk factors of exposure and smoking. Significantly decreased lung function was recorded for FVC, FEF50 and FEF25 in male and FVC, and FEF25 in female art students. Smokers and nonsmokers had similar reductions of lung function. Our data indicate that art students may be at risk of developing chronic respiratory symptoms and lung function changes as a result of their exposure and their smoking habits. PMID- 15666605 TI - Some anthropologic factors of performance in rhythmic gymnastics novices. AB - The aim of the study was to determine motor and morphological factors, and to assess their impact on specific motor skill performance in rhythmic gymnastics (RG). Experimental training process aimed at learning and improving basic movement structures of rhythmic gymnastics was performed for nine months in a sample of 50 female rhythmic gymnastics novices (mean age 7.1 +/- 0.3 years). Seven dimensions in total were isolated by factorial analysis of 13 motor, 11 morphological, and 20 specific rhythmic gymnastics tests. The factors of flexibility (Beta = 0.26; p < 0.05), explosive strength (Beta = 0.25; p < 0.05) and adipose voluminosity (Beta = -0.42; p < 0.001) explains 41% of the success in performing RG basic body elements--jumps, rotations, balance and flexibility (R = 0.64), while the frequency of movement (Beta = 0.44; p < 0.001) and non-adipose voluminosity (Beta = 0.26; p < 0.05) explains 26% of RG-specific manipulations with the apparatus--club, ribbon and ball wrist manipulation (R = 0.52; p < 0.01). According to study results, the RG-training process intended for rhythmic gymnastics novices should be programmed, with preset objectives for the development of flexibility and explosive strength, speed and peripheral joint strength and adipose tissue reduction. PMID- 15666606 TI - The contribution of collective attack tactics in differentiating handball score efficiency. AB - The prevalence of 19 elements of collective tactics in score efficient and score inefficient teams was analyzed in 90 First Croatian Handball League--Men games during the 1998-1999 season. Prediction variables were used to describe duration, continuity, system, organization and spatial direction of attacks. Analysis of the basic descriptive and distribution statistical parameters revealed normal distribution of all variables and possibility to use multivariate methods. Canonic discrimination analysis and analysis of variance showed the use of collective tactics elements on attacks to differ statistically significantly between the winning and losing teams. Counter-attacks and uninterrupted attacks predominate in winning teams. Other types of attacks such as long position attack, multiply interrupted attack, attack with one circle runner attack player/pivot, attack based on basic principles, attack based on group cooperation, attack based on independent action, attack based on group maneuvering, rightward directed attack and leftward directed attack predominate in losing teams. Winning teams were found to be clearly characterized by quick attacks against unorganized defense, whereas prolonged, interrupted position attacks against organized defense along with frequent and diverse tactical actions were characteristic of losing teams. The choice and frequency of using a particular tactical activity in position attack do not warrant score efficiency but usually are consequential to the limited anthropologic potential and low level of individual technical-tactical skills of the players in low-quality teams. PMID- 15666607 TI - Differences in physical activity between non-overweight, overweight and obese children. AB - The purpose of the study was to find out differences in moderate to vigorous physical activity among non-overweight, overweight and obese children, and between boys and girls. The sample included 364 children (179 boys and 185 girls), aged 6.4 years (+/- 0.3 SD). Physical activity was assessed by 7-day questionnaire. Age adapted BMI was used as overweight and obesity indicator. The children were divided into non-overweight, overweight and obese groups. It was found out, that there are significant differences in non-overweight, overweight and obese children (p < 0.05). Boys were significantly (p < 0.05) less moderate to vigorous physical active than girls, especially in indoor activities. There were also significant differences (p < 0.05) in moderate to vigorous physical activity among non-overweight and obese boys and among overweight and obese boys in weekends and total weekly activity. In girls there are significant differences (p < 0.05) in non-overweight, overweight and groups in weekends and total weekly activity. It is possible to conclude, that obese boys and overweight and obese girls, are prone to less physical activity. PMID- 15666608 TI - Prevalence of the three categories of handedness among Malawian school children. AB - A cross sectional study of prevalence of left, right and mixed handedness was made on 512 Malawian school children (240 boys and 272 girls) aged 6-17 years. Handedness was assessed using questionnaire incorporating questions on hand preferred for eight unimanual activities. Interviews with guardians, pupils and teachers were conducted to assess the pressure experienced by children to use right hand for unimanual activities. The overall prevalence of left-, right- and mixed-handers was 3.9%, 90.4% and 5.7%, respectively. Association between handedness and age or gender of children was nonsignificant. 96.2% of guardians and 92.7% of teachers were insistent upon use of right hand for unimanual tasks. Most of non-right handed children indicated that they had experienced pressure to use right hand for unimanual activities but they were still using left hand for one or more manual tasks. 28 pupils indicated that they stopped using left hand under the pressure from guardians and teachers. PMID- 15666609 TI - Duplex-Doppler ultrasonography in the detection of lower extremities deep venous thrombosis and in the detection of alternative findings. AB - The diagnoses observed in patients referred for the Doppler ultrasonographic examination of peripheral and iliac veins for suspected deep venous thrombosis (DVT) are presented in this study. During 48 months 2,610 patients were examined by duplex Doppler ultrasonography (US). Among these, 1,879 were women (72%) and 731 men (28%), with the age-range 16-91 (mean 56, 2) years. Ultrasonic scanners Acuson 128 XP 10, ATL HDI 5000, GE Logiq 7, and GE Logiq 9 were used, with transducers in the frequency range from 2.5-14 MHz. Findings were categorized into four main categories: (1) deep venous thrombosis (DVT); (2) pathology predominantly related to superficial veins without DVT, (3) pathology of adjacent structures; (4) normal findings. 562 patients had DVT (21.5%). 1,108 patients (42.5%) had predominant pathology of superficial veins: postthrombotic syndrome, superficial thrombophlebitis and varicose veins. 390 patients (14.9%) had pathology of surrounding structures, unrelated to veins, the most common pathology being popliteal cysts and muscular hematomas. These lesions must be properly diagnosed by US to avoid erroneous anticoagulant treatment. PMID- 15666610 TI - Electromyographic study of the anal sphincter in women. AB - The objectives of this study were to identify the electromyographic (EMG) changes in the anal sphincters, and to determine whether there are myogenic or neurogenic damages to the muscles in stress incontinence in women (SI). Possible damage was expected while the number of EMG studies has reported evidence of denervation as a result of certain aethiologic factors. The intention was to determine risk factors for development of fecal incontinence. 110 women with SI and 91 women with spinal lesion (SL) were examined. The anal sphincters were examined in both groups by means of standarized EMG technique with concentric needle electrode. The results demonstrated predominatly neurogenic lesion of the anal sphincters in SL women and predominatly normal findings in SI women. The contribution of genitourinary or pelvic surgeries to the fecal incontinence in SI group has been established by means of discriminatory analysis. PMID- 15666611 TI - Diagnostic value of a peroral sucrose permeability test in children with recurrent upper abdominal pain. AB - The access of ingested sucrose into blood and urine indicates the presence of mucosal lesions in the upper gastrointestinal tract. The study involved 42 children, aged 5-15, having recurrent upper abdominal pain and 43 peers with minor extra-abdominal complaints. Sucrose in urine was determined by thin layer chromatography. The test was positive in 27 out of 42 children having recurrent abdominal pain (64.3%) and in none of the control children (chi2 = 37.6, p < 0.0001). When correlated with endoscopic findings it was falsely negative in 12 out of 38 patients with endoscopically verified lesions of the stomach or duodenum and falsely positive in 1 out of 4 without lesions. Sensitivity of the test was 68.4%, specificity 97.9%, positive predictive value 96.3%. The test cannot be used as an alternative to endoscopy, but may serve for screening of candidates for it. PMID- 15666612 TI - Palliative treatment of hepatocellular carcinoma with percutaneous ethanol injection using tumor's feeding artery occlusion under the ultrasonic color Doppler guidance. AB - We evaluate the efficacy of PEIT in patients with HCC using duplex color Doppler US. The study included 27 HCC patients admitted to the University Hospital Centre Zagreb, between 1993 and 1997. PEIT was performed for ablation of tumor supplying vessels in HCCs of < 5 cm in diameter, and as a palliative measure for tumor feeding vessel obliteration in larger tumors. The efficacy of PEIT was evaluated with duplex color Doppler US, and controlled by dynamic CT scan (16 patients) or selective angiography of hepatic artery (11 patients). All patients had well vascularized tumors before PEIT, and after therapy 25 of them showed absent or minimal tumor vascularization. Recanalization of the tumor feeding vessel was detected with Doppler US within 9 months after therapy. Study results suggested that duplex color Doppler US should be the method of choice in the evaluation of PEIT as well as in the follow-up of HCC patients after PEIT. PMID- 15666613 TI - Uterine artery embolization for the treatment of uterine fibroids. AB - Uterine artery embolization can be regarded as a less invasive procedure for the treatment of fibroids compared with myomectomy, hysterectomy, and laparoscopic myolysis. The aim of this study was the evaluation of safety and efficacy of uterine artery embolization and of womens' opinion about this treatment. After gynecological examination sixty-nine premenopausal women underwent uterine artery embolization. All procedures but four were technically successful; three women underwent unilateral embolization because of vascular malformation and one of them had an allergic reaction to contrast medium. Of the 69 patients: 58 went home the day after embolization, and 11 within first week. The follow-up examinations after 3, 6 and 12 month showed a significant reduction of uterine and fibroid volume with significant improvement of bleeding. Therefore, according to this report, uterine artery embolization is a successful, minimal invasive treatment of myoma that preserves the uterus and requires shorter hospitalization and recovery times than surgery. PMID- 15666614 TI - Influence of the partial denture on the articulation of dental and postalveolar sounds. AB - Dental prosthesis is a foreign body in oral cavity and thus necessarily interferes with speech articulation. The purpose of this study was to examine influence of partial denture on speech quality and to show eventual differences in pronunciation of dental sounds c[ts], z [z], s [s] and postalveolar sounds c [t], z [3] and s [integral of]. We have examined differences in pronunciation between subjects with removable partial dentures, the same group without partial dentures and a control group. The study was performed on 30 subjects with removable partial dentures and 30 subjects with complete dental arch. All subjects were recorded while reading six Croatian words containing the examined sounds. Recordings were analyzed with Multispeech Program (Kay Elemetrics Inc.). Acoustic analysis--LPC (linear prediction coding) provided formant peaks (Hz) for each examined sound, its intensity (dB) and formant bandwidths (Hz). Results showed that subjects with partial dentures had 50% less distorted variables and that prostheses did not completely restore articulation of postalveolar sounds. Groups with and without prostheses had lower formant peaks intensities and wider formant bandwidths in comparison to the control group. Partial dentures have not significantly interfered with resonance frequency. At the same time, pronunciation of the examined sounds was significantly improved. However, precision of the articulation movements has deteriorated. PMID- 15666615 TI - Axillar compression syndrome: anatomical and clinical study. AB - In order to evaluate the possibility of compression of axillar artery by medial and lateral fascicle of brachial plexus, authors performed 26 axillar dissections on cadavers. Second part included analysis of 24 selective angiograms of axillar artery of patients with diagnosis of TOS. Third part included the use of modified hyperabduction test for determination of vascular bruit as safe test for diagnosis of axillar compression. Macroscopic changes of axillar artery by compression of medial and lateral fascicle of brachial plexus were present in 11.5%. Specific angiographic horizontal spike-shaped stop of contrast behind the surgical neck of humerus was present in 12.5%. Use of modified hyperabduction test revealed vascular bruit 29.5%. Specific relation of axillar artery and medial and lateral fascicle of brachial plexus revealed another possible etiologic factor in hyperabduction syndrome as a part of TOS. Use of modified hyperabduction test revealed subclinical phase of possible syndrome. PMID- 15666616 TI - Clinical importance of significant asimptomatic bacteriuria in newborns and infants during early postnatal period. AB - The aim of the study was to detect newborns at risk for developing renal impairment, and to point out the importance of significant asimptomatic bacteriuria in perinatal period and early infancy. Severe urinary tract anomalies are very often accompanied only by asimptomatic bacteriuria in perinatal period. Three urinalysis ware done after delivery. 212 newborns with significant asimptomatic bacteriuria underwent ultrasound examination, and were followed up to three months. Those with normal findings and with passing bacteriuria in the first 2 months were excluded. Group of 52 newborns underwent radioisotope examination. Frequency of urinary tract anomalies in newborns was 34.6%. Increased risk for renal impairment had children with urinary tract anomalies in close family, urinary tract infection or bacteriuria, EPH gestosis and prepartal symptoms of febrile infection in mother, children with IUGR, strangulated umbilical cord, prolonged jaundice and attacks of peripheral cyanosis in perinatal period. PMID- 15666617 TI - Lymphocyte subpopulations and the expression of intercellular adhesion molecules in chronic periodontitis. AB - Immunological responses to invading bacteria play a major role in the course of inflammatory periodontal diseases, such as CP. It was suggested that one of the major elements in determining the course of the disease is the expression of cellular adhesion molecules. We therefore investigated the expression of cellular adhesion molecules, ICAM-1 and beta-1 integrins, capillary density and lymphocyte subpopulations in gingival biopsies obtained from 20 patients with CP who responded and 21 patient who failed to respond to initial treatment using immunohistochemical methods. We found no differences between the two groups in capillary density, ICAM-1 and beta-1 integrin expression. Patients who responded to treatment had a lymphocytic inflammatory infiltrate consisting predominantly of T cells, while those who failed to respond had an approximately equal number of T and B cells. Our findings support the role of host immunological mechanisms in determining the outcome of CP and argue against a major role of differential cellular adhesion molecule expression. PMID- 15666618 TI - Reasons for extraction of permanent teeth in urban and rural populations of Croatia. AB - The survey aimed to determine the reasons for extraction of permanent teeth by general dental practitioners in urban and rural population of the Senj region, Adriatic coast, Croatia. During a two-year period (1998-9), a total of 2006 teeth were extracted in both regions, in patients aged 15+. The causes were defined as follows: (1) decay or root without a crown (radix relicta), (2) periodontal disease, (3) endodontic or periapical diseases and (4) other reasons- orthodontics/prosthodontics and dental trauma. The statistical Chi-square-test was used to determine the significant difference between the populations and the sexes. Dental caries was the most frequent cause for extraction (over 50%), followed by endodontic and periapical diseases (23%) as the result of untreated caries and at the end periodontal disease (21%). Urban population more often lose teeth due to periodontal disease (22.75%) than rural (18.93%, p < 0.05). Similarly, this is more frequent in the urban male population (25.61%) than the female urban population (20%, p < 0.05). In rural areas, people more often lost teeth as a result of endodontic and periapical disease (25.85%) than in the urban locations (19.07%, p < 0.01) and this is more frequent in women from rural areas (28.37%) than the rural men (22.44%, p < 0.05). Periodontal disease was not the main cause of tooth loss in either the rural or the urban population. Dental caries and its sequel remain the most important challenge for the dental service. It also reveals the inadequacy of dental services. Education of both the population and the general dental practitioners must be conducted in order to improve oral hygiene and to insist on conservative rather than extraction therapy. PMID- 15666619 TI - Palatal and dental arch morphology in Down syndrome. AB - The analysis of palatal vault morphology and maxillary dental arch shape was carried out in the sample of 42 Down syndrome (DS) patients with trisomy 21. The data were compared to those of healthy controls from the same population matched for age and sex. Palatal morphology and upper dental arch shape were studied on hard plaster casts of the patients and controls. No sexual dimorphism in palatal and dental arch shape was observed in DS and controls. Normal palatal shape was more frequent in controls than in DS subjects (52.38% vs. 28.57%; p < 0.05). DS patients displayed significantly higher frequency of shelf-like or "stair palate" (38.1%) than controls (11.9%) (p < 0.02). The younger age group (3-14 year) showed much higher frequency of "stair palate" than controls (26.19% vs. 2.38%; c2 = 9.72; p = 0.003). The older group of DS patients did not show increased frequency of such shape of the palatal vault. There was no significant difference in dental arch shape between DS patients and controls. High frequency of shelf like palate in DS subjects is decreasing by age. The obtained results indicate that palatal vault morphology is subjected to the age related changes. These changes can be attributed to the growth of caraniofacial structures and increased tonus of tongue and other orofacial muscles. PMID- 15666620 TI - The sequence in appearance and disappearance of impressiones gyrorum cerebri and cerebelli. AB - We investigated the sequence and the intensity in the appearance and the disappearance of the impressiones gyrorum cerebri and cerebelli, of juga cerebralia and cerebellaria and of juga cerebellaria interlobularia in the collection of 34 macerated and disarticulated skull bones from the newborn to 30 years of age (68 specimens/halves of skulls) and 19 skulls in the period from 30 to 80 years of age (38 specimens). Juga cerebralia on the squama of the temporal bone and cerebral lamina of the frontal bone appeared already in the course of the first year of life, much earlier than cited in the literature. The intensity of the development of juga cerebralia increased to the third decade. After that age, the intensity decreased gradually, and the juga cerebralia disappeared completely in parietal bones, in the cerebral fossae of the occipital bones and finally in most cases also on the cerebral lamina of the frontal bones. Juga cerebellaria and impressiones gyrorum cerebelli appeared in the middle of the second year of age and persisted to the ten years of age, which coincides with the closure of the fissures among the parts of the occipital bone. Jugum cerebellare intersemilunare appeared in the first year of life and persisted in its complete length, or interrupted in different sections of its course, during the whole life. The intensity in appearance of juga is partly influenced by the increasing thickness of the diploe. PMID- 15666621 TI - Use of digital photographs for artificial tooth selection. AB - Digital photography has become available to everybody. The aim of this study was to examine possibility of calculating the width of a missing central incisor using digital photographs. Digital photographs were obtained from 51 dentate subjects using a 3.1 Megapixel digital camera from various distances: 35 cm, 70 cm, 1 m and 1.5 m. For the calculation of the width of maxillary left central incisor (MLI), the following equation was used: MLI(calculated) = Photographic width of MLI x IPD / photographic IPD. Statistical analysis was made (Kolmogorov Smirnov test, dependent sample t-test, correlation and frequencies) using SPSS 10.0 for Windows. Results revealed no significant difference between the calculated MLI (70 cm, 1 m and 1.5 m distance) and actual MLI, however calculated MLI from 35 cm distance was significantly different from the actual MLI value (p < 0.01). The highest correlation was between calculated MLI (70 cm distance) and actual MLI. However, the highest percentage of results from a distance of 70 cm also fitted within +/- 0.3 mm; +/- 0.5 mm and +/- 1 mm difference from the actual MLI values. However, the results obtained from 1 m distance were also satisfactory. The technique of use of digital photography is of proven value in calculating the width of a maxillary central incisor. The photographs using a simple digital camera should be taken from a distance from 70 cm to 1 m. Saving such photographs in a dental office may eventually be helpful for calculating dimensions of artificial teeth. PMID- 15666622 TI - Temperament and character in cross-cultural comparisons between Swedish and Iranian people and Iranian refugees in Sweden--personality in transition? AB - The aim of the study was a cross-cultural comparison of personality traits between individuals from two very different cultures and refugees who resettled several years before from one to the other. Four hundred forty four Swedish individuals of the normal population; and 100 Iranian refugees in Sweden, and a group of 335 individuals from Tehran, capital of Iran, were investigated by means of the Temperament and Character Inventory, a questionnaire to assess temperament and character Iranians are those that are most frequently correctly classified followed by the Swedish based on temperament scores by means of a Discriminance analyses. Iranian refugees in Sweden were classified to about 50 per cent as Swedish and to slightly more then one-third as Iranians. Especially concerning character, 4 per cent only could be correctly classified as refugees. The results give some perspective on the adaptation process and personality changes in refugees several years after resettlement in another country with a complete different culture. PMID- 15666623 TI - Breastfeeding practices of health professionals and care workers in Turkey. AB - Breastfeeding has a significant positive impact on the health of infants and is the best nutrition source. Health professionals/care workers play a considerable role in the promotion of breast-feeding. The knowledge and attitude level of health professionals/care workers and its influence on their own breastfeeding behaviour was investigated. Two hundred sixty three of 345 health care workers (response rate = 76.2%) working in Children and Birth Hospitals and primary health care centres in a rural city (Isparta) in south Turkey participated in this study. A questionnaire was administered. Descriptive statistics and chi2 test have been used (alpha = 0.05) to analyse the data. Seventy four (41.1%) of the respondents with children (n = 180) exclusively nursed their children for less than 4 months; 62 (34.4%) nursed beyond 6 months. Physicians with children (n = 14, 53.8%) were used to exclusively breastfeed less than 4 months (chi2 (4) = 9.76, p = 0.045). This study showed a difference between health professionals/care workers attitudes and knowledge level and their or their spouses breastfeeding behaviour. The reason why knowledge level of health professionals did not translate into their own or their spouses' breastfeeding practices needs to be evaluated in further studies. PMID- 15666624 TI - Chinese women in Italy--menarche, pregnancy and maternity. AB - The purpose of this study was to assess the well-being and health status of Chinese immigrant women in Italy. The subjects (mean 29.5 years) frequented the "Health Center for Foreign Women and their Children" in Bologna, Italy. Information on living conditions, biological characteristics and pregnancy or maternity was obtained through interviews of the women in the Center during normal medical visits. Data on menarche age were based on the retrospective method since all the women were in China at the onset of menarche. Social factors influencing the health and behavior of Chinese women in Italian society are discussed. The analysis on children sex ratio at birth indicates that the old cultural Chinese tradition of son preference still continues to exist. These women have achieved a socio-economic improvement and generally good health status in Italy. The difficult living conditions they faced in China are indirectly reflected in their delayed puberty (14.4 +/- 1.4 years). The delayed sexual development was probably due to the same situation of severe poverty that forced them to migrate. PMID- 15666625 TI - What the Croatian doctors want to be learning about? AB - The aim of the study was to examine doctor's attitude about topics in continuing medical education (CME) using anonymous questionnaire that was given to the members of the Croatian Medical Association in Rijeka. The questions concerned doctor's interest of certain medical fields, influence of CME to their everyday practice, and importance of getting credits for re-licensure as a motive to participate in CME. The highest interest was shown for CME in emergency/intensive medicine and the lowest for transplantation medicine. The doctors in primary care showed statistically significantly higher interest for CME in family medicine, pulmology, rheumatology and rehabilitation medicine than hospital doctors. The influence of CME in everyday practice and the importance of getting credits for re-licensure as a reason to participate in CME, in the most cases, have been graded with medium grade 3. The results indicated the existence of specific needs in CME and stressed the importance of having CME with topics from clinical practice. PMID- 15666626 TI - Make love to forget: two cases of transient global amnesia triggered by sexual intercourse. AB - Transient global amnesia (TGA) is characterized by a sudden onset and by a typical resolution within several hours. Several precipitating events have been proposed: physical exertion, emotional experiences, etc. The aim of this paper was to present two cases of TGA triggered by sexual intercourse and to suggest a possible mechanism for the development of TGA. In both patients, clinical examination revealed elevated blood pressure. Laboratory examinations and brain CTs were normal. EEG demonstrated diffuse dysrhythmia and slow spike-waves, respectively. SPECT revealed hypoperfusion in the left frontal and right medial temporal regions. Various explanations of the mechanism of TGA are discussed. Based on the observed hypoperfusion in the medial temporal regions, a new hypothesis is advanced, suggesting the possibility that TGA occurs due to a pathologically changed or less adaptable anterior chorioid artery, initially constricted by hypotension following a blood shift from the center towards periphery. PMID- 15666627 TI - The race concept in six regions: variation without consensus. AB - Race, once the central concept in physical anthropology worldwide, now varies in the degree of support it receives in different regions. We present the currently available information on the status of the concept in the United States, the Spanish language areas, Poland, Europe, Russia, and China. Rejection of race ranges from high to low with the highest rejection occurring among anthropologists in the United States (and Canada). Rejection of race is moderate in Europe, sizeable in Poland and Cuba, and lowest in Russia and China. A discussion on the scientific and contextual reasons influencing these variations is presented. The tension between scientific evidence and social influences varies from region to region. The methods used in the studies reported here included questionnaires and content analysis. Response rates to questionnaires were often around 50 percent (with exception of the Polish studies). We discuss reasons for the low rates. Although a uniform method of data gathering is desirable, it may not suit scientists working in different traditions of theory and research. We conclude that it is once again timely to discuss the race concept in international meetings where all scientific and political changes occurring throughout the world in recent past decades are taken into account. PMID- 15666628 TI - Posttraumatic locked-in syndrome with an unusual three day delay in the appearance. AB - We report a rare case of posttraumatic locked-in syndrome (LIS) that appeared after an unusual three day delay. LIS was diagnosed according to clinical status and diagnostic methods (roentgenograms, computerized tomography, transcranial Doppler, electroencephalography, magnetic resonance imaging). A fourteen-year-old girl had a cervical spine injury during floor exercises that provoked LIS 72 hours after trauma. A rapid diagnosis of basilar thrombosis followed by antioedema and continuous anticoagulant therapy significantly improved the neurological status. In conclusion, posttraumatic locked-in syndrome can have sometimes a prolonged three-day delay in the clinical appearance. PMID- 15666629 TI - Bilateral amaurosis caused by Salmonella enteritidis infection. AB - The aim of this paper was to show the potential of Salmonella enteritidis infection to eventually result in visual impairment. A case of salmonellosis in a 6-year-old boy, caused by intake of a cake made from eggs infected with Salmonella enteritidis, is presented. Prolonged duration of the disease was followed by complete remission of neurologic complications and persistent amaurosis with bilateral optic nerve atrophy. A severe form of Salmonella enterocolitis with neurologic involvement can lead to optic nerve lesion with consequential loss of vision. PMID- 15666630 TI - Primary pulmonary botryomycosis: case report. AB - A case is presented of pulmonary botryomycosis in a 61-year-old man with a massive right-side pulmonary infiltrate which looked like a tumor (on X-ray). Microscopic examination of a transbronchial biopsy specimen revealed chronic suppurative inflammation, which did not regress despite intensive antibiotic therapy for a period of two months. Histological analysis of specimens taken during surgery for hemoptysis revealed pulmonary botryomycosis. The disease was diagnosed on the basis of characteristic eosinophilic granules in which the bacteria are surrounded by protein material (Splendore-Hoeppli phenomenon). Pulmonary actinomycosis was excluded. The case demonstrates that pulmonary botryomycosis can have the appearance of a mass which resembles pulmonary carcinoma on X-ray, and may also be mistaken for pulmonary actinomycosis. For this reason, pulmonary botryomycosis, although rare, should be excluded during differential diagnosis of hemoptysis or pulmonary infiltrates. PMID- 15666631 TI - Spontaneously ruptured gastrointestinal stromal tumor (GIST) of the jejunum mimicking acute appendicitis. AB - Gastrointestinal stromal tumors (GISTs) are characterized with diverse clinical presentations, including acute and chronic gastrointestinal bleeding, abdominal pain, presence of an intra-abdominal mass, anorexia, and intestinal obstruction. A 60-year-old obese woman presented as an acute abdominal emergency with right lower quadrant (RLQ) pain and tenderness, nausea and leukocytosis, all mimicking acute appendicitis. Laparotomy revealed a spontaneously ruptured GIST of the jejunum, which was localized to the RLQ due to postoperative adhesions following previous two cesarean sections and cholecystectomy. Complete surgical resection was performed, followed by an uneventful early postoperative course. PMID- 15666632 TI - Five reasons why inbreeding may have considerable effect on post-reproductive human health. AB - As the genetic architecture of common complex diseases of late onset is emerging through intensive research, it is intriguing to assess the predicted effect of inbreeding on those diseases. In this paper, we propose five reasons why we believe inbreeding may have a considerable effect on post-reproductive human health. (i) The joint effect of inbreeding depression on all polygenic quantitative phenotypes that confer risk for late-onset diseases is predicted to be multiplicative rather than additive. (ii) The "genetic load" of rare "Mendelian" variants with large deleterious effects in post-reproductive adults is unknown, but could be much greater than expected as these variants were invisible to selection through human history. (iii) Deleterious effects resulting from autozygosity in hundreds of affected rare recessive variants of small effect under common disease/rare variant (CD/RV) hypothesis could result in epistatic effects that could jointly impair capacity to compensate against environmental risks. (iv) Heterozygote advantage in loci under balancing selection could be reduced by inbreeding. (v) Published empirical evidence in animals and humans consistently report large inbreeding effects on late-onset traits. Since inbreeding is common in many populations and the effects of inbreeding depression could substantially contribute to disease burden and reduced life expectancy we believe there is now a clear need for further genetic epidemiological research in humans to investigate this issue. PMID- 15666633 TI - 200 years of Lithuanian anthropology. PMID- 15666634 TI - [Contribution to the experimental study of artificial respiration by the rocking bed]. PMID- 15666635 TI - [On the destruction processes induced by androgenic hormones, in the embryonic rudiments of the mouse]. PMID- 15666636 TI - [Problem of breath analyser]. PMID- 15666637 TI - [Study of morbidity related to age in patients who have died in hospital]. PMID- 15666638 TI - [High density lipoproteins of blood plasma]. PMID- 15666639 TI - [Autoimmunity and diabetes mellitus. Recent developments]. PMID- 15666640 TI - [On the auditory prosthesis using telestimulation. History perspectives]. PMID- 15666641 TI - [About of the therapy with the bacteriophages]. PMID- 15666642 TI - [New trends in therapeutic use of bacteriophages]. PMID- 15666643 TI - [Measure of red cells deformability in sonic hemolytic anemias]. PMID- 15666644 TI - [A comparative study of the activity of chlorpromazine and meprobamate in healthy mice and those made diabetic with alloxan]. PMID- 15666645 TI - [Problems about pituitary tumors in Cushing's disease after surrenalectomy (one case)]. PMID- 15666646 TI - [Regarding the physical state of lipids in atherosclerotic lesions]. PMID- 15666647 TI - [1879-1979. It is now one hundred years since Victor Galtier, a professor of Veterinary School in Lyon, presented a paper on the prophylaxis of rabies to the Academy of Sciences]. PMID- 15666648 TI - [Results of a campaign for the early detection of cancer prevention of gynecological and mammary cancers]. PMID- 15666649 TI - [Precipitation of low density lipoproteins by polyanions. Mechanisms, clinical interest and practical applications]. PMID- 15666650 TI - [Surgical treatment of "severe" congenital cardiopathies in babies]. PMID- 15666651 TI - [Problems raised by long-term accidental irradiations]. PMID- 15666653 TI - Nursing and CAP: caregivers in action. PMID- 15666654 TI - Quit smoking: for good. PMID- 15666655 TI - Fibrin sealants in surgical or traumatic hemorrhage. AB - PURPOSE OF REVIEW: Fibrin sealants have been used to control surgical hemorrhage for three decades, and numerous articles have reported their use in a variety of surgical procedures in both animal models and humans. This article reviews the recent literature on fibrin sealants with the specific aim of highlighting the use of fibrin sealants in planned and simulated trauma to provide background for clinicians who may consider using fibrin sealants in specific cases. An overview of the mechanisms of action of fibrin sealants, their indications, and current commercial formulations is also provided. RECENT FINDINGS: Recent studies have evaluated the use of fibrin sealants in vascular surgery, including aortic anastomosis in an animal model, gastrointestinal anastomoses, plastic surgery, urologic procedures including heminephrectomy, and other procedures. SUMMARY: Fibrin sealants continue to be used and evaluated in animal models and surgery. Their use in military settings and in civilian trauma centers, explored for several years, is not reported extensively in the literature. Recent events and the current tempo of military operations dictate that many potential scenarios exist for using fibrin sealants to control traumatic hemorrhage. PMID- 15666656 TI - Immunoadsorption of factor VIII inhibitors. AB - PURPOSE OF REVIEW: Extracorporeal immunoadsorption is being increasingly applied in a variety of disorders. This approach is particularly suited to removal of antibodies or inhibitors to coagulation factor VIII and may be particularly useful before the administration of large amounts of expensive replacement or bypass therapy for patients with hemophilia who are bleeding, or patients undergoing immune tolerance therapy. RECENT FINDINGS: In patients with inhibitors to factor VIII, several types of immunoadsorption therapy have been used, although reports are mainly anecdotal, consisting of relatively small numbers of persons. Nonetheless, the findings suggest that immunoadsorption may be clinically effective and cost-effective and should be considered early in the treatment of appropriate patients. New immunoadsorption technologies are being described for a variety of disorders, including hemophilia, and a new synthetic matrix of polystyrene beads functionalized with sulfonate and tyrosyl methylester groups for immunoadsorption removal of factor VIII inhibitors is intriguing. SUMMARY: Although immunoadsorption was shown to be clinically effective in patients with inhibitors to factor VIII more than two decades ago, recent papers have emphasized the desirability of early implementation of the modality in the treatment plan. Immunoadsorption is relatively easy to perform with few adverse effects, but specialized equipment is required, and it should be performed in an experienced setting. Although potentially less costly than other (bypass) therapies, immunoadsorption is itself not inexpensive, and its comparative effectiveness with plasmapheresis and other management options for the dangerously bleeding patient with antibodies to factor VIII should be determined by multicenter randomized controlled trials. Interesting recent novel technical developments in the field may facilitate increased use of the procedure. PMID- 15666657 TI - Barcode identification for transfusion safety. AB - PURPOSE OF REVIEW: Errors related to blood transfusion in hospitals may produce catastrophic consequences. This review addresses potential solutions to prevent patient misidentification including the use of new technology, such as barcoding. RECENT FINDINGS: A small number of studies using new technology for the transfusion process in hospitals have shown promising results in preventing errors. The studies demonstrated improved transfusion safety and staff preference for new technology such as bedside handheld scanners to carry out pretransfusion bedside checking. They also highlighted the need for considerable efforts in the training of staff in the new procedures before their successful implementation. SUMMARY: Improvements in hospital transfusion safety are a top priority for transfusion medicine, and will depend on a combined approach including a better understanding of the causes of errors, a reduction in the complexity of routine procedures taking advantage of new technology, improved staff training, and regular monitoring of practice. The use of new technology to improve the safety of transfusion is very promising. Further development of the systems is needed to enable staff to carry out bedside transfusion procedures quickly and accurately, and to increase their functionality to justify the cost of their wider implementation. PMID- 15666658 TI - Blood donor complications after whole-blood donation. AB - PURPOSE OF REVIEW: This review focuses on the common and uncommon adverse events that occur during or after whole-blood donation. The incidence of seeking outside medical care for an adverse event is at least 1 in 3400 blood donations. Physicians may encounter blood donors with adverse events and should be familiar with their prognosis and treatment. RECENT FINDINGS: There are 15 million whole blood donations per year in the United States, and one third of the donors develop one or more adverse events. A recent study defined the common complications based on a postdonation interview as bruise (23%), sore arm (10%), fatigue (80/%), and vasovagal reaction (7%). A 2003 American Red Cross national donor complication database from 7,000,000 whole-blood donations classified reported donor complications and the incidence of outside medical care. Descriptive studies have been published on vasovagal syncopal reactions, antecubital nerve injuries (irritations), and arterial punctures, and the medical outcomes for these entities have been more precisely defined. The effect of blood donor adverse events on donor retention rates is a new area that is just beginning to be studied. SUMMARY: The incidence of complications and types of complications is more precisely defined today. Medical outcomes are better defined for vasovagal reactions, nerve injuries (irritations), and arterial puncture. These data should help physicians who encounter these entities. PMID- 15666659 TI - Solvent/detergent-treated plasma: composition, efficacy, and safety. AB - PURPOSE OF REVIEW: Recent reports on adverse events associated with solvent/detergent-treated plasma have raised concerns about the efficacy and safety of this type of therapeutic plasma. Comparisons of various brands of solvent/detergent-treated plasma have revealed substantial differences in their composition. RECENT FINDINGS: Retrospective analyses and case reports suggest an association between the low protein S activity levels found in solvent/detergent treated plasma and venous thromboembolism when solvent/detergent-treated plasma is given for thrombotic thrombocytopenic purpura or transfused massively during liver transplantation surgery. Augmented bleeding caused by hyperfibrinolysis, possibly caused by the low plasmin inhibitor (also termed alpha2-antiplasmin) activity of solvent/detergent-treated plasma, has also been a matter of concern. The efficacy and safety of various types of solvent/detergent-treated plasma have been studied intensively for all indications for therapeutic plasma by at least five prospective trials, four observational studies, and post-licensure monitoring. A review of the literature shows that the clinical significance of transient protein S and plasmin inhibitor deficiency has been considerably overstated, even when such deficiency might be aggravated by the administration of large amounts of solvent/detergent-treated plasma. However, different brands of solvent/detergent-treated plasma differ substantially with respect to their composition and hemostatic balance. It cannot be completely excluded that the very low citrate and protein S concentrations versus higher clotting factor concentrations found in a given brand of solvent/detergent-treated plasma could contribute to coagulation activation in the settings of thrombotic thrombocytopenic purpura and liver transplantation. SUMMARY: Future research should seek to optimize the composition of solvent/detergent-treated plasma. Prospective trials and prospective hemovigilance studies are required to determine the rate of adverse events occurring after treatment with solvent/detergent-treated plasma and other types of therapeutic plasma. PMID- 15666660 TI - The modern landscape of transfusion-related iatrogenic Creutzfeldt-Jakob disease and blood screening tests. AB - The idea that blood in naturally occurring transmissible spongiform encephalopathies is not infectious has imploded in the face of recent transmissions from the blood of naturally occurring scrapie in sheep and of variant Creutzfeldt-Jakob disease in humans. Although donor exclusion criteria ensure that the number of any further iatrogenic cases will be small, the risk of future blood-borne disease transmissions could be entirely eliminated by a diagnostic preclinical screening test. A variety of methodological approaches to blood testing are under development, with different levels of success, but no method has yet achieved the critical goal of discriminating transmissible spongiform encephalopathy-infected from healthy uninfected humans. PMID- 15666661 TI - Testing for parentage and kinship. AB - PURPOSE OF REVIEW: Parentage analyses are of interest to workers in health care, law enforcement, immigration and other fields. This review describes recent applications, technical advances, and quality improvements. RECENT FINDINGS: Mutations at short tandem repeat sequence loci confound interpretations of genetic data used to assess all blood relationships. Rates of the usual mutation type (change in repeat number) are probably related to specific alleles at each locus as well as to allele length, locus, and gender. Short tandem repeat sequences have relatively limited information content per locus. Intermediate tandem repeat sequence loci may be better. In immigration proceedings, probabilities can be calculated for excluding parentage in blood relatives who might impersonate the biologic parent. Unrelated immigrants from a subpopulation may appear to be related, but it is now possible to statistically determine the effect of population substructure on kinship determinations. In forensic analyses, sex chromosomal (X and Y) short tandem repeat sequences and mitochondrial DNA sequence variations have helped identify the parental lineages of human remains. Recent laboratory quality improvements include a way to estimate the frequency of common mother-child specimen mislabeling in routine paternity cases. In prenatal testing there are now methods for avoiding erroneous assignment of contaminant maternal alleles to the fetus. False paternity exclusions can be avoided by adhering to a standard of the American Association of Blood Banks requiring duplicate DNA isolation and retesting of excluded men. SUMMARY: Laboratory technology and quality have advanced, but genetic tests with greater information content are needed. Better communication is highly desirable between persons requesting tests and parentage laboratories. PMID- 15666662 TI - [Where is nursing quality?]. PMID- 15666663 TI - [Lack of clinical competence]. PMID- 15666664 TI - [Fewer falls thanks to nursing research]. PMID- 15666666 TI - ["Always a second too soon"]. PMID- 15666665 TI - ["Participation" is the magic word]. PMID- 15666667 TI - [The entire department went "fishing"]. PMID- 15666668 TI - ["We are observed like a colorful bird in a cage" (interview by Urs Luthi)]. PMID- 15666669 TI - [Improved quality lowers costs]. PMID- 15666670 TI - ["Being recognized as a human being who suffers"]. PMID- 15666671 TI - [The role of ethics committees]. PMID- 15666672 TI - [A record about the patient]. PMID- 15666673 TI - [Perhaps a small thumb poke is enough]. PMID- 15666674 TI - ["I ask myself how the family will react"]. PMID- 15666675 TI - ["Team work, it is always a learning experience" (interview by Francoise Taillens)]. PMID- 15666676 TI - [Custodian of silence]. PMID- 15666677 TI - Long-term interactions between migratory caribou, wildfires and Nunavik hunters inferred from tree rings. AB - Barren-ground caribou (Rangifer tarandus) herds in North America may reach considerable size and undertake large-scale seasonal migrations from the Arctic tundra to the boreal forest. To test the caribou decline hypothesis associated with native harvesting and fire, we have documented the long-term trends of caribou activity based on a novel approach which uses tree-ring dated trampling scars produced by caribou hooves in the extensive trails distributed over the summer and winter ranges of the Riviereaux-Feuilles herd (RAF herd, east of Hudson Bay in northern Quebec). The age structure data of trampling scars from lichen woodlands distributed over the entire RAF range confirmed the overall trends of caribou activity from the late 1700s to present time. Over the last 200 years, the RAF herd has undergone two highs in the late 1700s and 1900s separated by a moderate activity pattern in the late 1800s. Native harvesting was possibly involved in the early 1900s decline, although at a moderate level. The reduced magnitude of caribou activity during this period has not modified the natural cycle of highs and lows, which suggests that other demographic factors were controlling the changing caribou abundance. Our data also show that only exceptionally large fires may have a minor, short-lived impact on caribou migrations but not on caribou numbers. PMID- 15666678 TI - Declining extent of open-water refugia for top predators in Baffin Bay and adjacent waters. AB - Global climate change is expected to severely impact Arctic ecosystems, yet predictions of impacts are complicated by region-specific patterns and nonuniform trends. Twentyfour open-water overwintering areas (or "microhabitats") were identified to be of particular importance for eight seabird and marine mammal species in the eastern Canadian High Arctic and Baffin Bay. Localized trends in the available fraction of open-water were examined in March during 1979--2001, derived from approximate sea ice concentrations from satellite-based microwave telemetry. Declines in the fraction of open-water were identified at microhabitats in Baffin Bay, Davis Strait, coastal West Greenland, and Lancaster Sound. Increases in open-water were observed in Hudson Bay, Hudson Strait, and Foxe Basin. The biological importance of each microhabitat was examined based on species distribution and abundance. Potential consequences of reduced open-water for top marine predators include impacts on foraging efficiency and oxygen and prey availability. PMID- 15666679 TI - Reduction of organochlorine residues in Goshawk eggs (Accipiter gentilis) from Northern Germany (1971-2002) and increasing eggshell index. AB - We investigated unhatched eggs (n = 57) of Goshawks (Accipiter gentilis) from the period 1988--2002 and compared them with eggs (n = 134) from 1971--1978. Mean concentrations of PCBs, sigmaDDT and HCB in the earlier period were (mg kg(-1) lipid weight) 50.1, 126.6, and 117.7, while the values from the later period were statistically significantly lower: 27.7, 4.1, 0.1, respectively. Nevertheless, the concentration of PCB is relatively constant during the last 8-10 years. The variability of residues was lower within clutches than among clutches. No age dependent accumulation in eggs could be demonstrated. A statistically significant lower shell index was measurable for the period 1971--1978, while this index during 2001--2002 was relatively high as it was during the late 18th century. We are suggesting a threshold value for effect concentrations on shell index. Residues of the DDT-group had an influence on the number of fledged young per nest site. PMID- 15666680 TI - Declining biodiversity in agricultural landscapes and the effectiveness of agri environment schemes. AB - Agricultural intensification, greatly accelerated as a result of the EU Common Agricultural Policy (CAP), has led to drastic reductions in the populations of many wild plant and animal species that used to be characteristic of farmland. In 1992, the EU provided the member states with its Agri-environment Regulation 2078/92 to help member states reverse these developments by means of agri environment schemes. The question is: will the implementation of these schemes be sufficient to restore the biological diversity on farmland? Most studies that have examined the effectiveness of agri-environment schemes have focussed on farmland birds in Great Britain and The Netherlands. So far, the positive effects appear to be limited. Continuous evaluation and adaptation of these schemes is needed to enable the biodiversity on farmland to recover from the EU's former policy. PMID- 15666681 TI - Pesticide pollution remains severe after cleanup of a stockpile of obsolete pesticides at Vikuge, Tanzania. AB - High levels of DDT residues and hexachlorocyclohexanes (HCHs) were found in soil, well water, and surface water around a collapsed pesticide storage shed at Vikuge Farm, Tanzania. Residues of DDT and HCHs were found at three soil depths down to 50 cm. Surface soil samples contained up to 28% total DDT and 6% total HCH residues. Water samples had concentrations of up to 30 microg L(-1) of organochlorine pesticides. Other compounds detected were aldrin, azinphos-methyl, carbosulfan, gamma-chlordane, chlorprofam, heptachlor, hexazinone, metamitron, metazachlor, pendimethalin, and thiabendazole. Although the visible remains of pesticides have been removed, the remaining soil is itself hazardous waste and poses a risk to the environment and the inhabitants of the surrounding villages. These findings show the necessity to follow up the environmental situation at former storage sites of obsolete stocks of pesticides, and that the environmental problems are not necessarily solved by removing the visible remains. PMID- 15666682 TI - Tracking the anthropogenic drivers of ecological impacts. AB - Despite the pivotal role human factors (anthropogenic drivers) are presumed to play in global environmental change, substantial uncertainties and contradictory conclusions about them continue. We attempt to further discipline the human factors issue by estimating the effects of two anthropogenic drivers, population and affluence, on a wide variety of global environmental impacts, including greenhouse gas emissions, emissions of ozone depleting substances, and the ecological footprint. Population proportionately increases all types of impacts examined. Affluence typically increases impacts, but the specific effect depends on the type of impact. These findings refocus attention on population and material affluence as principal threats to sustainability and challenge predictions of an ameliorating effect of rising affluence on impacts. PMID- 15666683 TI - Integrating protected area management with local needs and aspirations. AB - Conservationists recognize that many protected areas have limited future prospects without the cooperation and support of local people, especially in developing countries. Since the 1980s Integrated Conservation and Development Projects (ICDPs) have attempted to reconcile park management with local needs and aspirations, usually with disappointing results. Achieving local cooperation and support without jeopardizing conservation goals remains a top priority for parks, however. Fortunately, the lessons from the ICDP experience provide an important opportunity to inform the next generation of biodiversity conservation programs, including those concerned with poverty alleviation as well as those working at ecosystem and landscape scales. More recent and more promising approaches have started to incorporate elements of adaptive management, new partnership models with stakeholders and the vertical integration of site-level work with policy initiatives and institutional development. PMID- 15666684 TI - Globalization and multi-spatial trends in the coverage of protected-area conservation (1980-2000). AB - This study is focused on the global expansion of protected-area coverage that occurred during the 1980--2000 period. We examine the multi-scale patterning of four of the basic facets of this expansion: i) estimated increases at the world regional and country-level scales of total protected-area coverage; ii) transboundary protected areas; iii) conservation corridor projects; and iv) type of conservation management. Geospatial patterning of protected-area designations is a reflection of the priorities of global conservation organizations and the globalization of post-Cold War political and economic arrangements. Local and national-level factors (political leadership and infrastructure) as well as international relations such as multilateral and bilateral aid combine with these globalization processes to impact the extent, type, and location of protected area designations. We conclude that the interaction of these factors led to the creation and reinforcement of marked spatial differences (rather than tendencies toward worldwide evenness or homogenization) in the course of protected-area expansion during the 1980--2000 period. PMID- 15666685 TI - Soil functional responses to excess nitrogen inputs at global scale. AB - There is little evidence that nitrogen (N) cycling in the highly weathered, low phosphorus (P), acidic soils found in Southern Hemisphere continents will differ greatly from that in North America and Europe. Evidence from the 'south' shows: the similarity in forms and temporal patterns in losses of N from different land uses; that the C:N ratios of the forest floor/litter layer from different continents are strongly predictive of a range of processes on a global scale; that generalizations based on Northern Hemisphere experience of the impact of N additions to 'P-limited' ecosystems are likely to fail for southern ecosystems where anatomical and physiological adaptation of native plants to low-P soils makes questionable the concept of 'P-limitation'; that the greatest threats in the 'south' are probably changes in land use that may greatly increase N inputs and turnover; that localized increases in N inputs produce similar effects to those seen in the 'north'. PMID- 15666686 TI - Bacteria, biomineralization, and biomarker molecules--biogeochemical aspects of the evolution and impacts of life on earth. PMID- 15666687 TI - A perspective on the role of minerals in prebiotic synthesis. AB - The possible role of minerals in prebiotic chemistry is discussed. Reactions involving the transformation of inorganic forms of the biogenic elements into simple organic molecules are emphasized. Three central issues are presented in detail: i) the types of reactions minerals could possibly have promoted; ii) the availability of minerals with catalytic potential on early Earth; and iii) the available research strategy and methods to evaluate the roles minerals may have played in prebiotic chemistry on early Earth. PMID- 15666688 TI - The microbial role in hot spring silicification. AB - Recent experimental studies indicate that microorganisms play a passive role in silicification. The organic functional groups that comprise the outer cell surfaces simply serve as heterogeneous nucleation sites for the adsorption of polymeric and/or colloidal silica, and because different microorganisms have different cell ultrastructural chemistry, species-specific patterns of silicification arise. Despite their templating role, they do not appear to increase the kinetics of silicification, and at the very most, they contribute only marginally to the magnitude of silicification. Instead, silicification is due to the polymerization of silica-supersaturated hydrothermal fluids upon discharge at the surface of the hot spring. Microorganisms do, however, impart an influence on the fabric of the siliceous sinters that form around hot spring vents. Different microorganisms have different growth patterns, that in turn, affect the style of laminations, the primary porosity of the sinter and the distribution of later-stage diagenetic cementation. PMID- 15666689 TI - Are picoplankton responsible for calcite precipitation in lakes? AB - Deposits of lacustrine calcite are important records of environmental changes. In order to interpret these archives, knowledge about the origin of the calcite is essential. It has been accepted that calcite precipitation can be induced by bacteria and algae. However, the detailed mechanisms are still unclear. This review summarizes what is known about the interactions between calcite precipitation and the autotrophic picoplankton. We consider findings from both field and laboratory studies. Field studies show that calcite precipitation in oligotrophic lakes is strongly linked with picocyanobacteria blooms. Laboratory experiments led to the formulation of the mechanism of precipitation induced by microalga. Experiments also showed that precipitation induced by picocyanobacteria is influenced by various factors including the uptake of inorganic carbon and the structure of the cell walls. Recent studies indicate that the influence of environmental conditions like the composition of lake water has to be taken into account as well. We conclude that in situ observations of precipitation processes at picoplankton cells under controlled conditions are needed to improve our understanding of mineral bacteria interaction. PMID- 15666690 TI - Particulate organic matter in the sea: the composition conundrum. AB - As organic matter produced in the euphotic zone of the ocean sinks through the mesopelagic zone, its composition changes from one that is easily characterized by standard chromatographic techniques to one that is not. The material not identified at the molecular level is called "uncharacterized". Several processes account for this transformation of organic matter: aggregation/disaggregation of particles resulting in incorporation of older and more degraded material; recombination of organic compounds into geomacromolecules; and selective preservation of specific biomacromolecules. Furthermore, microbial activities may introduce new cell wall or other biomass material that is not easily characterized, or they may produce such material as a metabolic product. In addition, black carbon produced by combustion processes may compose a fraction of the uncharacterized organic matter, as it is not analyzed in standard biochemical techniques. Despite these poorly-defined compositional changes that hinder chemical identification, the vast majority of organic matter in sinking particles remains accessible to and is ultimately remineralized by marine microbes. PMID- 15666692 TI - Hemichorea-hemiballism associated with hyperintense putamen on T1-weighted MR images: an update and a hypothesis. AB - In 1998 some patients with hyperglycemia-related hemichorea-hemiballism have been reported with a hyperintense putamen on T1-weighted MR images, presumably resulting from petechial hemorrhage. I questioned this explanation from my experience because (1) the areas of hyperintense lesions and their time evolutions did not match with those of the high density lesions on CT, (2) these hyperintense lesions persisted for years, and (3) the hyperintense lesions extended inferiorly to the midbrain. Therefore, a biopsy was performed in one patient and disclosed a fragment of gliotic brain tissue with abundant gemistocytes, which I proposed was sufficient to explain the shortening of T1 relaxation time. In addition, because two of our patients were associated with cortical infarcts and without hyperglycemia, I have suggested that cerebral ischemia might be a more important cause. In 1999 Fujioka et al reproduced the MR finding in animals 7 days after 15-minute occlusion of the middle cerebral artery. Therefore, both studies have suggested that the MRI finding resulted from a progressive pathological reaction in an incomplete infarction. In 2003 Fujioka et al further reported that the hyperintensity on T1-weighted MR images after mild ischemia may involve a paramagnetic effect resulting from tissue manganese accumulation in reactive astrocytes. PMID- 15666691 TI - Clinical advances in the preventive treatment of migraine. AB - Evidence is emerging on the physiological processes underlying episodic migraine and the rationale for preventive treatment. Objectives for preventive treatment include limiting future pain and disability and potentially modulating the course of disease progression. Many factors influence medication choice for patients such as migraine type, patient preference, co-existing conditions, and medication side effects. In this paper, frequently prescribed treatment options for migraine prevention are reviewed. Most headache preventive medications treat other medical disorders and are found serendipitously to be beneficial in migraine or other headache disorders. A new treatment option is topiramate, with proven safety and efficacy across the largest controlled migraine prevention trials conducted to date. New clinical information is enhancing patient care and enabling clinicians to ease the burden of migraine worldwide. PMID- 15666694 TI - Is the stimulation frequency of the repetitive nerve stimulation test that you choose appropriate? AB - The repetitive nerve stimulation test (RNST) has been a useful method in the diagnosis of myasthenia gravis (MG). In clinical practice, a short train of repetitive stimulation is usually given at 3 Hz. Although it was documented that lower stimulation frequencies could offer a greater sensitivity, no study has been done to testify the most sensitive stimulation frequency for RNST. To find out an optimal stimulation frequency, we performed RNST at 0.5, 1, 3, 5, 7, 10, 15 and 20 Hz in 15 MG patients and 5 healthy subjects. The results showed that the decremental response was most often seen at 7 Hz rather than at 3 Hz. To augment the sensitivity in the diagnosis of MG, RNST should be performed stimulation not only at 3Hz but also at other frequencies, preferably 7 Hz. PMID- 15666693 TI - Immunohistochemical study of skin nerve regeneration after toe-to-finger transplantation: correlations with clinical, quantitative sensory, and electrophysiological evaluations. AB - Cutaneous nerve regeneration following toe-to-finger transplantation was studied by immunohistochemical technique using antibody to protein gene product 9.5 (PGP 9.5) which is a specific neuronal marker. By this technique, epidermal and dermal nerves were semi-quantified and the Meissner's corpuscles were quantified. There were also quantitative sensory tests (QST) including pinprick, pressure and temperature, as well as electrophysiological studies including digital nerve sensory conduction, digital nerve somatosensory evoked potentials and sympathetic skin response at the pulp of the transplanted toes. The opposite corresponding normal finger and normal toe served as controls. Study subjects were 20 adult patients with toe-to-finger transplantation for at least one year. A score system was used to quantify the results of histochemical, psychophysiological and electrophysiological studies. Clinically 7 patients had good recovery and 13 patients had poor recovery. Cutaneous nerve regeneration in the transplanted toes was incomplete with epidermal nerve, dermal nerve and the Meissner's corpuscle significantly reduced. The nerve regeneration was correlated with clinical recovery, QST and electrophysiological data. These findings indicate that immunohischemical technique is useful to evaluate skin nerve regeneration following toe-to-finger transplantation, and that although nerve regeneration did occur, it was incomplete and correlated with the severity of hand injury. PMID- 15666695 TI - Paraparesis as the major initial presentation of aortic dissection: report of four cases. AB - Aortic dissection, which typically manifests as sudden tearing or migratory pain, is a well-known medical emergency. However, in 5% of aortic dissection patients, there is no pain. In these patients, the diagnosis depends on the development of neurologic complications. After analyzing the initial symptoms of a series of patients with aortic dissection, we found 4/211 (1.9%) patients suffered from paraparesis. We suggested that the mid- or low thoracic cord be most vulnerable site during acute aortic dissection. This report highlighted the importance of considering the diagnosis of aortic dissection in a patient with a history of acute onset of transient or permanent neurological symptoms in the lower limbs. Whether paraparesis can be an indicator of the prognosis of aortic dissection requires further researches. PMID- 15666696 TI - Fibromuscular dysplasia of the vertebral artery presenting with lateral medullary syndrome: a case report. AB - We have recently encountered a rare case of fibromuscular dysplasia (FMD) of the vertebral artery (VA) presenting as lateral medullary syndrome. A 39-year-old male was admitted to our hospital due to vertigo, dysarthria and numbness of the left face and the right limbs. A magnetic resonance imaging (MRI) and magnetic resonance angiography (MRA) of brain revealed lateral medullary infarction in the territory of the left posterior inferior cerebellar artery (PICA). The angiography of the VA revealed tubular stenosis of the left extracranial VA and a focal vascular kinking as well as web in the right extracranial VA, confirming the diagnosis of FMD. We present this rare case to emphasize that FMD could be one of the risk factors causing lateral medullary syndrome in young people. PMID- 15666697 TI - [Basic mechanism for generation of brain rhythms]. AB - Study of the basic mechanism of brain rhythms adds to our understanding of the underlying processes of neuronal network within the human brain. Electroencephalography (EEG) records summated extracellular field potentials from large pyramidal neurons in the cerebral cortex. The characteristic rhythmic oscillation of brain rhythms are beststudied by sleep spindles, which are generated within the thalamus through a network of thalamic reticular cells, thalamocortical projection cells and cortical pyramidal cells. Fast rhythms including beta and gammaare present during arousal and focused attention. Besides a reflection of activation from the brainstem reticular activating system, fast rhythms also represent an activated state of the underlying neuronal network. Alpha waves are readily recognized brain waves during relaxed wakefulness; however, the origins of alpha rhythm are not well understood. It is presumed that alpha rhythms are generated by contingent pyramidal cell, and by intracortical connections, spread in the cortical layers. Rhythmic alpha spindles probably represent a de-activated state. PMID- 15666698 TI - [Legendary Hwa Tuo's surgery under general anesthesia in the second century China]. AB - In traditional Chinese medicine, Hwa Tuo (110 ? - 208 A.D.) is one of the most famous doctors. He used only few herbs in drug treatment or applied few points in acupuncture, and achieved excellent results. His ultimate fame came from his remarkable surgical skills and his discovery of general anesthesia. According to the Chronicle of the Three Kingdoms (ca. 270 A.D.) and the Annals of the Later Han Dynasty (ca. 430 A.D.), Hwa Tuo performed operations under general anesthesia and the operations even included major ones such as dissection of gangrenous intestines. Before the surgery, he gave patient an anesthetic to drink to become drunk, numb and insensible. The anesthetic was called " foamy narcotic powder" and probably dissolved in wine. Because Confucian teachings regarded the body sacred, surgery as a form of body mutilation was not encouraged, or even became a taboo. Despite his great achievement, practice of surgery could hardly take off and the death of Hwa Tuo marked the end of Chinese surgery. Unfortunately, the composition of the anesthetic powder was not mentioned in those two books or other Chinese medical writings. The herb has been thought to be datura flower, aconite root, rhododendron flower, or jasmine root. Furthermore, Hwa Tuo's operations under general anesthesia were not described in details. Therefore, his remarkable achievement needs to be further documented. In Western medicine, the first operation under general anesthesia occurred at the Massachusetts General Hospital in 1846 when William Morton demonstrated the effectiveness of ether. How could Hwa Tuo accomplish such scientific achievement in the second century has remained a mystery. Even so, it seems quite remarkable that Hwa Tuo had come up with the idea of performing surgery under general anesthesia using the "foamy narcotic powder". PMID- 15666699 TI - Progressive facial and cerebral hemiatrophy: Parry-Romberg syndrome or scleroderma "en coup de sabre"? PMID- 15666700 TI - [Exclusivity of data in drug registration files and Israel's international status with regard to intellectual property rights protection]. AB - Protecting intellectual property rights encourages creativity and innovation, which are key measures for research and development of new drugs and also promotes the introduction of drugs to new markets. Over the past years, many countries have enacted laws implementing the protection of data in drug registration files. Until recently, Israel has not provided any form of data protection. Current moves focus on new legislation granting exclusivity in the marketing of innovative drugs. Although controversial, this legislation aims to secure balance in Israel between the innovative pharmaceutical companies and the public's interest in competitive generic drugs. PMID- 15666701 TI - [Performance of screening tests in the preventive services for infants]. AB - Early detection is one of the aims of the surveillance by the preventive services in infancy. OBJECTIVES: This article presents the performance of screening for anemia in infancy, compliance with iron supplementation and treatment, screening for hearing, and abnormalities of the hip joint, and referral to ear, nose and throat (E.N.T.) and orthopedic specialists according to the screening results. METHODS: The sample consisted of Jewish and Arab women who gave birth during March 2000. Mothers were interviewed after the infants reached the age of 15-19 months (667 Jewish and 211 Arab). All the women utilized the preventive services of the Ministry of Health, municipalities or H.M.O's. Arab women from East Jerusalem utilized the services of non-governmental organizations (NGO's). RESULTS: A total of 61% of the Jewish and 86% of the Arab infants had a hemoglobin examination. Eighty-five percent of the Jewish and 75% of the Arab infants received iron as a supplement or treatment for anemia. The performance of hearing tests was 88% and 77%, respectively. Among the infants referred to E.N.T. no hearing loss was detected. Fourteen percent were not screened nor referred by the family physician/pediatrician. Among the self referrals, one child was detected with a hearing loss. Percent referrals to the orthopedic surgeon was higher among Jewish than Arab infants. Among those not examined through the preventive services, the rate of pathology detected by the orthopedic surgeon was higher than those referred. RECOMMENDATIONS: Findings highlight the need to improve the quality of screening and increase the percent of infants screened in the Arab and Jewish population, particularly those of the N.G.O.'s, with emphasis on high risk groups. Furthermore, efforts should be directed to define and recommend routines for detection of abnormalities of the hip joint, and integrate the procedures within the health services. PMID- 15666702 TI - [The effect of acute physical activity on blood glucose levels of children with insulin-dependent diabetes mellitus]. AB - BACKGROUND: Physical activity has an important role in the management of insulin dependent diabetes mellitus (IDDM). However, the effect of acute physical activity on blood glucose levels of children with IDDM has not been studied in depth although this type of activity is typical of children. PURPOSE: To evaluate the effect of acute physical activity on blood glucose, growth hormone and cortisol levels in children with IDDM with relation to their hemoglobin A1 (HbA1) level. METHODS: Eleven children with IDDM performed both 20 minutes of cycling and 20 minutes of free physical exercise, on two different occasions. Blood glucose, growth hormone and cortisol levels were measured before and after each activity. HbA1 level was used to assess their metabolic control. RESULTS: Both cycling and physical exercise induced similar elevation in growth hormone and cortisol levels. However, while cycling induced a significant decrease in glucose levels (p < 0.001), physical exercise induced its increase in 6 children. The decrease in glucose level following cycling was correlated with baseline glucose levels (r = 0.64) and with the duration of diabetes (r = 0.87). The changes in glucose level following physical activity did not correlate with HbA1 levels. CONCLUSIONS: The effect of acute physical activity on children with IDDM probably depends on the type of activity and not its intensity or the metabolic control. It is suggested that cycling has an advantage over free exercise in reducing blood glucose levels. It is advisable that children with IDDM and their patents should be aware of the implications of acute physical activity. PMID- 15666704 TI - [Development and assessment of simulated patient based curricular unit in order to improve clinical skills in adolescent medicine of pediatric and family practice residents]. AB - OBJECTIVE: This study describes the development and evaluation of a curricular program aimed at improving the clinical and communication skills required for managing the clinical encounter with adolescents, among pediatric and family practice residents using the technique of standardized patients. METHODS: A daylong curricular unit which included: clinical interviews with simulated patients presenting typical clinical problems occurring among adolescents; a teaching unit explaining the principles of communication with adolescents and a clinical discussion and feedback session using video clips. Forty-eight residents in pediatrics and family practice undertook the curricular unit. A total of 120 doctor-simulated patient interviews were documented using six patients' case scenarios representing somatization disorder, eating disorder, chronic illness and non-compliance, pregnancy, drug abuse and menstrual disorder. RESULTS: The doctors' self-estimation of clinical skills associated with adolescent medicine that was documented at the beginning of the curricular unit was generally reported as low. In most of the cases the participants did not identify the main problem and diagnoses that were presented. The problems least correctly diagnosed were eating disorder and non-compliant chronic illness. Only in the cases of somatization and pregnancy more than half of the residents identified the problems that were presented. At the end of the curricular unit the doctors expressed a very high level of satisfaction with the program. CONCLUSION: The present study developed and evaluated a short and structured curricular unit dealing with the principles of communication with adolescent patients using the technique of simulated/standardized patients. It was found that the lack in these skills, which was identified in pediatric and family practice residents, could be bridged using this educational technique. PMID- 15666703 TI - [Is there an association between PUPPP and preeclampsia, abruptio placentae and fetal death?]. AB - This is a case study of a 28-year old woman, gravida 1 para 0, presented at 37+4 weeks gestation due to purpura and fetal death. A 3700 gram dead male fetus without gross malformation was delivered by forceps. Post delivery abruptio placentae, diffuse intravascular coagulation, fulminant preeclampsia and acute renal failure followed. Skin biopsy revealed pruritic urticarial papules and plaques of pregnancy. The patient fully recovered one week postpartum. This case report discusses a possible association between pruritic urticarial papules, plaques of pregnancy and severe preeclampsia with fetal death. PMID- 15666705 TI - [Informed consent--a simulation workshop for surgeons and gynaecologists]. AB - BACKGROUND: Informed consent as part of the Israeli Patient's Rights Act (1996) is poorly known and only partly followed among physicians in Israel. PURPOSE: A one-day workshop on informed consent was designed with a dual purpose: a) to instruct on the legal requirements for obtaining consent to surgery for physicians in the surgical disciplines, and b) to practice effective and humane physician-patient relationships. METHODS: Sixty-one residents and senior staff took part in 6 sessions of a one-day workshop. Participants watched twice and performed twice videotaped simulated consent conversations with standardized patients. Six professional actors personified patients with common surgical problems and different characters and behaviours. Each participant was challenged with medical problems in her/his discipline. Following the first simulation, the participants attended a group discussion concerning legal requirements for informed consent, and expectations of patients and physicians in terms of communication skills and patient-physician relationships. Three types of feed back were employed: a) non-professional personal feedback from the actors; b) personal feedback from a communication specialist using one of the videotaped encounters; and c) group discussions of each of the 6 types of simulations with examples from the encounters. RESULTS: Feedback from the participants was positive and showed interest and need of knowledge of the law as well as of communicative skills. CONCLUSIONS: There is a place, among residents and senior physicians, for experiential learning of how to effectively and humanely obtain informed consent. PMID- 15666706 TI - [Family health centers in Israel: to change or not to be]. AB - As reported in this issue, the Ministry of Health Family Health Centers (FHC) in Israel is not fulfilling some of the requirements of the Ministry of Health. For example, hemoglobin levels are not checked in all of the babies and at least 31% of the babies do not receive supplemental iron. Universal neonatal screening for hearing loss by objective methods is not conducted nor recommended by the Ministry of Health, as advised by pediatric professional organizations, while screening for hearing loss is conducted in the second half of the first year by a method which is not currently recommended. FHC physicians conduct initial screening for suspected developmental dysplasia for only some of the children, while too many children go directly to the orthopedic specialist. Previous studies have shown that only 20% of Jewish women residing in larger townships in Israel use the FHC for prenatal care. Based on all the above the government policy makers are facing hard decisions regarding FHC services. One option is to transfer all services to the health maintenance organizations. A second option is to maintain the FHC with the following modifications: 1) revise and update the health management guidelines to meet universal recommendations; 2) improve compliance to guidelines; 3) improve outreach and acceptance by the public of the services of the FHC. PMID- 15666707 TI - [Primary congenital glaucoma]. AB - Primary congenital glaucoma (PCG) is a rare genetic disease usually diagnosed during the first year of life. It occurs because of developmental anomalies of the chamber angle that prevents drainage of aqueous humor, thereby elevating intraocular pressure. Its incidence is 1 in 10,000 live newborns in Western societies and 1 in 1,200 live newborns in the Arab-Bedouin population of the Negev region in Israel. Most cases of PCG appear to be sporadic. The cytochrome P4501B1 gene located within the GLC3A locus on chromosome 2p21 is mutated in individuals with PCG. The triad of epiphora, photophobia, and blepharospasm is classical for PCG. General anesthesia is usually required for an adequate examination of intraocular pressure, corneal diameter, optic disc, and axial length in young children. Congenital glaucoma is almost always managed surgically, with medical therapy being used only as a temporizing measure before surgery or when surgical intervention has repeatedly failed. At least 50% of eyes with PCG presenting at birth will become legally blind (visual acuity < 6/60). Patients with PCG require follow-up examinations throughout their lives. PMID- 15666708 TI - [Cerebral CT perfusion in the diagnosis of acute brain infarction]. AB - In recent years medical imaging is directed towards functional evaluation of tissues. Perfusion CT is an established functional method providing information on brain perfusion, enlightening the differentiation between irreversibly injured tissue and reversibly impaired "tissue at risk". With the new generation of multislice CT scanners, the performance of three examinations in a very short time is now feasible: Non-enhanced CT, CT angiography (CTA) and perfusion CT. Since these technologies are available in most hospitals 24 hours daily, comprehensive evaluation of the extent of ischemic event is now possible even in the emergency room. This information allows intravenous tissue plasminogen activator therapy within 3 hours of stroke onset in cases of acute ischemic stroke. We present the technique of CT perfusion and discuss its advantages and limitations as well as future applications. PMID- 15666709 TI - [Vitamin B6 (Pyridoxine)--excessive dosage in food supplements and OTC medications]. AB - Vitamin B6 (Pyridoxine) is sold in Israel as a supplement and is available over the-counter (OTC) without regulation. High intake of this vitamin is found in patients with premenstrual syndrome, carpal tunnel syndrome, pregnancy associated nausea and vomiting, decreasing homocysteine levels and improving cognitive function. Mega-doses of this vitamin may result in intoxication. In this review we will outline vitamin B6 function, daily recommended intake, deficiency signs and patients in deficiency risk, and the clinical spectrum of vitamin B6 intoxication. PMID- 15666710 TI - [Cranberry juice and urinary tract infection]. AB - Cranberries have long been the focus of interest for their beneficial effects in preventing urinary tract infections (UTIs). Cranberries contain two compounds with anti-adherence properties, which prevent fimbriated E. coli from adhering to uroepithelial cells in the urinary tract. Approximately a dozen clinical trials have been performed testing the effects of cranberries on the urinary tract. However, these trials have a number of apparent limitations. Most importantly, the trials have used a wide variety of cranberry products, such as cranberry juice concentrate, juice cocktail, and cranberry capsules, and have employed different dosing regimens. Further research is required to clarify unanswered questions regarding the role of cranberries in protecting against UTI in general and in women with anatomical abnormalities in particular. PMID- 15666711 TI - [Pancreas transplantation in insulin dependent diabetic patients]. AB - Successful transplantation of the pancreas as a whole organ, or as isolated islet cells, is the only treatment that achieves a stable normoglycemia as a result of insulin secretion and renewal of serum glucose levels control systems. Nowadays, one year patient survival after pancreas transplantation is above 90%, while functioning grafts are observed in 84% of combined pancreas kidney and in 70% of isolated pancreas transplantations. Type I insulin dependent diabetic patients aged 45 or less with severe diabetic nephropathy, without immediate life saving cardiovascular risk, highly motivated and well informed achieve the best results. Namely, these results include euglycemia without exogenous insulin, improvement of secondary complications of diabetes, protection of the kidney graft, and longer life expectancy with better quality of life, as compared to dialyzed diabetic patients. PMID- 15666712 TI - [The foundation meeting of the Israeli Society for Medical and Research Chronobiology (ISMRC)]. PMID- 15666713 TI - [The pelvic inflammatory snake]. PMID- 15666714 TI - Seymour Hutner's soup kitchen for protozoa. PMID- 15666715 TI - Fourteen morphotypes of Entodinium ovumrajae (Ophryoscolecidae, Entodiniomorphida) found in the Dromedary camel of Egypt. AB - During a survey of the ciliate protozoal composition of the stomach contents of nine dromedary camels of Egypt, fourteen morphotypes of Entodinium ovumrajae, which has been considered as a species peculiar to camels, were found in six camels. Except for five morphotypes including one originally described as an independent species and its forms, these were newly detected. These morphotypes, divided into three groups, can be identified mainly by the morphology of their ectoplasmic processes. Each camel had on average, about five morphotypes of this species. PMID- 15666716 TI - Analyses of the ribosomal DNA region in Nosema bombycis NIS 001. AB - Ribosomal DNA (rDNA) containing small subunit (SSU) rDNA and both flanking regions in the entomopathogenic microsporidian Nosema bombycis NIS 001 was amplified from genomic DNA with a primer set based on the sequence of an inverse polymerase chain reaction (PCR)-derived fragment. In this fragment, SSU rDNA was divided by a 618-bp insert at nt 599, and 5S rDNA was located downstream of the SSU rDNA, fragmented by 284-bp intergenic spacer. In addition, the 48-bp 3'-end of large subunit (LSU) rDNA was located 118 bp upstream of the fragmented SSU rDNA. In the amplicon, the region upstream of the LSU rDNA was a homologue of the C-terminal CHARLIE8 transposon-like element of human GTF2IRD2. In this organism, another fragmented SSU rDNA, which was divided by a 231-bp insert at nt 50, was also detected. Both the intact (insertless) and fragmented SSU rDNAs clustered with LSU rDNA and 5S rDNA and the intergenic sequences between SSU rDNA and 5S rDNA were divergent in an organism. Reverse transcription (RT)-PCR assay indicated that not only the intact SSU rDNA but also the fragmened SSU rDNA were transcribed in N. bombycis. PMID- 15666717 TI - Body, nuclear, and ciliary changes during conjugation of Protospathidium serpens (Ciliophora, Haptoria). AB - We studied cell size and shape, nuclear changes, and the ciliary pattern during conjugation of Protospathidium serpens, using protargol impregnation and morphometry. Preliminary data were gathered from Epispathidium ascendens and Apertospathula armata. Conjugation of P. serpens is temporary, isogamic, and without preconjugation divisions. Pair formation is heteropolar, and the partners unite obliquely with the oral bulge. The body becomes smaller and broader during conjugation, but no basic changes occur in the ciliary pattern. Conjugation and nuclear reconstruction follow the usual mode of ciliates. However, some peculiarities occur: only two of the four synkaryon derivatives of the second synkaryon division enter the third division and generate four macronuclear anlagen, which fuse to a single, long macronucleus strand. During conjugation, E. ascendens unites obliquely as P. serpens, while A. armata can pair dorsal-to dorsal surface, ventral-to-dorsal surface, or obliquely as P. serpens. The nuclear processes of these three species are also rather different, showing a considerable diversity in union modes and nuclear events of spathidiids; E. ascendens even has preconjugation division. Confirming previous data, the present study shows convincingly that most of the spathidiid nuclear variability is caused by reconstruction processes occurring in post-dividers, exconjugants and, possibly, exautogamonts. When these specimens are removed from the populations, spathidiid species are as stable (or variable) as other ciliate species. PMID- 15666718 TI - Establishment of a new amphileptid genus, Apoamphileptus nov. gen. (Ciliophora, Litostomatea, Pleurostomatida), with description of a new marine species, Apoamphileptus robertsi nov. spec. from Qingdao, China. AB - A new pleurostomatid genus Apoamphileptus is described, which is diagnosed as: Belonging to the Amphileptidae with spica on right side; on each side of the cell, a single perioral kinety, which encircles the cytostome and does not extend to the posterior end of the cell; somatic kineties of both sides near ventral margin shortened and forming a postoral suture; two to several extra fragments with densely arranged dikinetids located in anterior portion of left side. As the type species, the morphology and infraciliature of Apoamphileptus robertsi nov. spec., isolated from a shrimp-farming pond near Qingdao (Tsingtao), China, have been investigated using living observations and the protargol silver impregnation method. The diagnosis for this new species is: Apoamphileptus 90-180 x 30-60 microm in vivo, body elongate pyriform-shaped and slightly flattened; with one cross-striated band along the cytostome; 2-6 (generally 4) large macronuclear nodules, one micronucleus; 33-43 right somatic kineties; left side 6-8 kineties; two extra anterior fragments on left side; about 13 contractile vacuoles dispersed throughout whole body; extrusomes absent or not recognizable; marine habitat. Some morphologically related morphotypes are discussed and tabulated. Regarding the pattern of infraciliature and other morphological features, the well-described fresh-water species, Amphileptus claparedii Stein, 1867 is believed to be a member of this new genus, hence a new combination is suggested: Apoamphileptus claparedii (Stein, 1867) nov. comb. PMID- 15666719 TI - Unexpected effects of prey dimensions and morphologies on the size selective feeding by two bacterivorous flagellates (Ochromonas sp. and Spumella sp.). AB - Current models on protistan size-selective feeding assume that contact probability is the factor that largely explains observed food preferences. Contact probability is generally expected to be positively correlated with prey size and therefore to explain observed food selection for larger prey items. We critically tested these basic assumptions on size-selective feeding using the interception-feeding chrysomonad nanoflagellates Ochromonas sp. and Spumella sp. Mechanisms of differential feeding were studied during distinct stages of the selection process (i.e. contact probability, capture efficiency, ingestion efficiency, and differential digestion) by means of high-resolution video microscopy. Food selection was investigated using a mixture of microspheres ranging from 0.3-2.2 microm in diam., as well as a mixed bacterial community. In contrast to current model assumptions, the contact probability was highest for microspheres of intermediate size (0.9-1.2 microm), but was not generally positively correlated with prey size over the whole prey size range. Capture and ingestion also proved to be involved in size selection: these patterns were also independent of the food concentration (p = 0.968 for Ochromonas, p = 0.971 for Spumella). Even though the capture rate was significantly higher for attached flagellates than for swimming flagellates (p < 0.001), size selectivity was not affected (p > 0.05). Our results indicate that: (i) size selection is not actively regulated by these flagellates, but is a passive process; (ii) contact probability is not generally positively correlated with prey size, but shows a maximum for intermediate-sized prey in the prey size spectrum of 0.3-2.2 microm; and (iii) selection steps other than contact probability are crucial for size selection and should be integrated in models on size selection. PMID- 15666720 TI - Sterol composition of Pneumocystis jirovecii with blocked 14alpha-demethylase activity. AB - Several drugs that interact with membrane sterols or inhibit their syntheses are effective in clearing a number of fungal infections. The AIDS-associated lung infection caused by Pneumocystis jirovecii is not cleared by many of these therapies. Pneumocystis normally synthesizes distinct C28 and C29 24 alkylsterols, but ergosterol, the major fungal sterol, is not among them. Two distinct sterol compositional phenotypes were previously observed in P. jirovecii. One was characterized by delta7 C28 and C29 24-alkylsterols with only low proportions of higher molecular mass components. In contrast, the other type was dominated by high C31 and C32 24-alkylsterols, especially pneumocysterol. In the present study, 28 molecular species were elucidated by nuclear magnetic resonance analysis of a human lung specimen containing P. jirovecii representing the latter sterol profile phenotype. Fifteen of the 28 had the methyl group at C 14 of the sterol nucleus and these represented 96% of the total sterol mass in the specimen (excluding cholesterol). These results strongly suggest that sterol 14alpha-demethylase was blocked in these organisms. Twenty-four of the 28 were 24 alkylsterols, indicating that methylation of the C-24 position of the sterol side chain by S-adenosyl-L-methionine:sterol C-24 methyl transferase was fully functional. PMID- 15666721 TI - Exuviotrophic apostome ciliates from crustaceans of St. Andrew Bay, Florida, and a description of Gymnodinioides kozloffi n. sp. AB - Gymnodinioides kozloffi n. sp. is described from the eelgrass broken-back shrimp Hippolyte zostericola. The species is distinct from others in the apostome genus Gymnodinioides in that the trophont ciliature has a small group of kinetosomes located to the right of Kinety 9a, and Kinety 1 and 2 are divided. Other apostome morphologies are described from many decapod crustaceans from St. Andrew Bay, Florida, including Gymnodinioides inkystans, Hyalophysa chattoni, and variants of both H. chattoni and G. kozloffi. All of these apostome ciliates are exuviotrophic, found feeding on exuvial fluid within the exoskeleton of the host after ecdysis. The hosts surveyed for this study are the following: Callinectes sapidus, Eurypanopeus depressus, Hippolyte zostericola, Farfantepenaeus spp., Palaemonetes intermedius, Palaemon floridanus, Portunus spp., Tozeuma carolinense, and Sicyonia laevigata, which revealed a number of new host-apostome records. PMID- 15666722 TI - HSP90, tubulin and actin are retained in the tertiary endosymbiont genome of Kryptoperidinium foliaceum. AB - The dinoflagellate Kryptoperidinium foliaceum has replaced its ancestral peridinin-containing plastid with a fucoxanthin-containing diatom plastid via tertiary endosymbiosis. The diatom endosymbiont of K. foliaceum is much less reduced than well-studied endosymbiotic intermediates, such as cryptophytes and chlorarachniophytes, where relict nuclear genomes are retained in secondary endosymbionts. The K. foliaceum endosymbiont retains a prominent nucleus, multiple four-membrane plastids, and mitochondria, all within a relatively large volume of cytoplasm that is separated from the host cytoplasm by a single membrane. Here we report the first protein-coding gene sequences from the K. foliaceum endosymbiont and host nuclear genomes. We have characterised genes for nucleus-encoded cytosolic proteins, actin (from endosymbiont), alpha-tubulin (from both), beta-tubulin (from host), and HSP90 (from both), in addition to homologues from pennate diatoms Nitzschia thermalis and Phaeodactylum tricornutum. Phylogenetic reconstruction shows that the actin is diatom-derived, the beta-tubulin dinoflagellate-derived, while both diatom- and dinoflagellate derived alpha-tubulin and HSP90 genes were found. The base composition biases of these genes co-varied with their phylogenetic position, suggesting that the genes still reside in their respective genomes. The presence of these genes implies they are still functional and more generally indicates that the endosymbiont is less genetically reduced than those of cryptophytes or chlorarachniophytes, raising the interesting question of whether any genes have transferred between the two nuclear genomes. PMID- 15666723 TI - Phylogenetic comparison of the myxosporea based on an actin cDNA isolated from Myxobolus cerebralis. AB - The full-length actin gene from Myxobolus cerebralis (McerAct-1), the first characterized from representatives in the phylum Myxozoa, encodes a 378-amino acid polypeptide with an estimated molecular weight of 41,580-Da. A phylogenetic comparison found M. cerebralis to branch outside the metazoans. This finding contrasts with previous reports that suggest an evolutionary affinity of the Myxozoa with either the Bilateria or Cnidaria. PMID- 15666724 TI - Identification of single nucleotide mutations that prevent developmentally programmed DNA elimination in Paramecium tetraurelia. AB - The excision of internal eliminated sequences (IESs) occurs during the differentiation of a new somatic macronuclear genome in ciliated protozoa. In Paramecium tetraurelia, IESs show few conserved features with the exception of an invariant 5'-TA-3' dinucleotide that is part of an 8-bp inverted terminal repeat consensus sequence with similarity to the ends of mariner/Tc1 transposons. We have isolated and analyzed two mutant cell lines that are defective in excision of individual IESs in the A-51 surface antigen gene. Each cell line contains a mutation in the flanking 5'-TA-3' dinucleotide of IES6435 and IES1835 creating a 5'-CA-3' flanking sequence that prevents excision. The results demonstrate that the first position of the 5'-TA-3' is required IES excision just as previous mutants have shown that the second position (the A residue) is required. Combining these results with other Paramecium IES mutants suggests that there are few positions essential for IES excision in Paramecium. Analysis of many IESs reveals that there is a strong bias against particular nucleotides at some positions near the IES termini. Some of these strongly biased positions correspond to known IES mutations, others correlate with unusual features of excision. PMID- 15666725 TI - Urotricha psenneri n. sp. and Amphileptus piger (Vuxanovici, 1962) n. comb., two planktonic ciliates (Protozoa, Ciliophora) from an oligotrophic lake in Austria. AB - Two euplanktonic ciliates, Urotricha psenneri n. sp. (Prostomatida) and Amphileptus piger (Vuxanovici, 1962) n. comb. (Pleurostomatida), were discovered in the surface plankton of the oligotrophic Lake Traunsee in Austria. Their morphology and infraciliature were studied in live cells as well as in specimens impregnated with protargol and silver nitrate. Urotricha psenneri is a small urotrichid, less than 50 microm length and with a single caudal cilium. It is unique in having (i) a massive oral basket projecting as a conspicuous bulge with cylindrical microfibrillar annulus and (ii) a curved brosse row 1 in the broad, barren circumoral area. Amphileptus piger (Vuxanovici, 1962) is about 55 x 13 microm in vivo, has two macronuclear nodules with a single micronucleus in between in the posterior body half, has a single contractile vacuole with a terminal excretory pore, and few, but thick and thus highly conspicuous extrusomes. The amphileptid ciliary pattern (spica) is difficult to recognise due to the widely spaced basal bodies. PMID- 15666727 TI - Potential for escape of live boll weevils (Coleoptera: Curculionidae) into cottonseed, motes, and cleaned lint at the cotton gin. AB - Reintroductions of the boll weevil, Anthonomus grandis grandis Boheman, into areas of the United States where it has been eradicated or suppressed are very expensive to mitigate. There is concern that a cotton gin in an eradication zone may serve as a site of boll weevil reintroductions when processing cotton harvested in a neighboring infested zone. Similarly, there is a question whether weevil-free areas can safely import gin products, such as cottonseed and baled lint, from infested areas without risking an introduction. Many countries require fumigation of imported U.S. cotton bales to protect against boll weevil introductions, costing the U.S. cotton industry millions of dollars annually. In previously reported experiments, we quantified the potential for boll weevils to survive passage through precleaning machinery in the gin. In this study, we quantified survival potential of boll weevils passing through the gin stand and segregating into the cottonseed, mote, or lint fractions. We also examined boll weevil survival when passed with ginned lint through a lint cleaner. We present a flow chart of experimentally determined survival potentials of boll weevils passing through the various subprocesses of the gin, from which one can calculate the risk of a live boll weevil reaching any point in the process. Our data show that there is virtually no chance of a boll weevil being segregated alive into the cottonseed or of one surviving in the lint to approach the bale press. Therefore, quarantine or fumigation of cottonseed and cotton bales to guard against boll weevil introductions is unnecessary. PMID- 15666726 TI - An analysis of the microsporidian genus Brachiola, with comparisons of human and insect isolates of Brachiola algerae. AB - The genus Brachiola is the newest microsporidian genus established for a human infection with the type species being B. vesicularum in skeletal muscle. Subsequently, the microsporidium, Nosema algerae, identified from mosquitoes, was added to this genus because of morphological and physiological similarities. The present report illustrates a confirmed case of Brachiola algerae infecting skeletal muscle in a 56-year-old woman who was being treated for rheumatoid arthritis with immunosuppressive drugs. In the following study, these two human infecting microsporidian species are ultrastructurally compared from human biopsy tissue. Additionally, Brachiola algerae from mosquitoes as reference B. algerae, was grown in athymic mice and compared to the human isolate in vivo, and in culture. B. algerae is morphologically identical in the host situations presented and different from B. vesicularum in human skeletal muscle. B. algerae has a consistently, slightly longer spore that typically contains one row of polar filament coils, while B. vesicularum typically contains two rows of polar filament coils and occasionally, one or three rows. In proliferative development, B. vesicularum forms protoplasmic extensions which do not occur on B. algerae, nor have they been reported on any other microsporidium. This report demonstrates that B. vesicularum and B. algerae are two different species of Brachiola that infect human skeletal muscle. PMID- 15666728 TI - The case and opportunity for public-supported financial incentives to implement integrated pest management. AB - Food, water, and worker protection regulations have driven availability, and loss, of pesticides for use in pest management programs. In response, public supported research and extension projects have targeted investigation and demonstration of reduced-risk integrated pest management (IPM) techniques. But these new techniques often result in higher financial burden to the grower, which is counter to the IPM principle that economic competitiveness is critical to have IPM adopted. As authorized by the 2002 Farm Bill and administered by the U.S. Department of Agriculture (USDA) Natural Resources Conservation Service (NRCS), conservation programs exist for delivering public-supported financial incentives to growers to increase environmental stewardship on lands in production. NRCS conservation programs are described, and the case for providing financial incentives to growers for implementing IPM is presented. We also explored the opportunity and challenge to use one key program, the Environmental Quality Incentives Program (EQIP), to aid grower adoption of IPM. The EQIP fund distribution to growers from 1997 to 2002 during the last Farm Bill cycle totaled approximately 1.05 billion dollars with a portion of funds supporting an NRCS designed pest management practice. The average percentage of allocation of EQIP funds to this pest management practice among states was 0.77 +/- 0.009% (mean +/- SD). Using Michigan as an example, vegetable and fruit grower recognition of the program's use to implement IPM was modest (25% of growers surveyed), and their recognition of its use in aiding implementation of IPM was improved after educational efforts (74%). Proposals designed to enhance program usefulness in implementing IPM were delivered through the NRCS advisory process in Michigan. Modifications for using the NRCS pest management practice to address resource concerns were adopted, incentive rates for pest management were adjusted, and an expanded incentive structure for IPM technique adoption was tabled for future consideration. The case is strong for using public-supported financial incentives offered by the EQIP to aid grower adoption of IPM as a means to address resource concerns, but current use of the EQIP for this purpose is modest to meager. With appropriate program adjustments and increased grower awareness, USDA NRCS conservation programs, and the EQIP in particular, may provide an important opportunity for growers to increase their use of IPM as a resource conservation and farm management tool. PMID- 15666729 TI - Variation in barley yellow dwarf virus transmission efficiency by Rhopalosiphum padi (Homoptera: Aphididae) after acquisition from transgenic and nontransformed wheat genotypes. AB - The effects of different acquisition access periods (AAPs) and inoculation access periods (IAPs) on the transmission efficiency of barley yellow dwarf luteovirus (BYDV) by Rhopalosiphum padi (L.) (Homoptera: Aphididae) after feeding on transgenic or nontransformed wheat, Triticum aestivum L., genotypes were studied. Three wheat genotypes were tested as virus sources: virus-susceptible 'Lambert' and 'Lambert'-derived transgenic lines 103.1J and 126.02, which express the BYDV PAV coat protein gene. Lower virus titers were measured in BYDV-infected transgenic plants compared with Lambert. No significant differences in transmission efficiency were detected for R. padi after varying IAPs, regardless of genotype. Transmission efficiency increased with an increase in AAP in all genotypes tested. However, AAPs ranging from 6 to 48 h on Lambert resulted in significantly greater transmission efficiency than similar periods on transgenic 103.1J. Maximum transmission efficiency (70%) was observed after a 48-h AAP on Lambert, whereas the same AAP on 103.1J and 126.02 resulted in a significantly lower transmission efficiency (57%). Contrasts were used to compare the rates of transmission and the theoretical maximum transmission percentage among the different wheat genotypes serving as virus sources. Both parameters were significantly different among genotypes, indicating that viral acquisition from each genotype resulted in a unique pattern of virus transmission by R. padi. The lowest rate of virus transmission after an AAP was observed on 103.1J compared with 126.02 or Lambert. This is likely associated with a lower virus titer in 103.1J. This is the first report of transgenic virus resistance in wheat affecting the transmission efficiency of a virus vector. PMID- 15666730 TI - Efficacy of native and recombinant Cry1B protein against experimentally induced and naturally acquired ovine myiasis (fly strike) in sheep. AB - Several hundred strains of Bacillus thuringiensis (Bt), isolated in New Zealand from samples of soil and sheep fleece, were tested for toxicity to larvae of the blowfly Lucilia cuprina Wiedemann. Characterization of the Bt strains revealed that three of the more active strains produced Cry1Ba (an insecticidal protein present in Bt mother cell crystal inclusion) that was toxic to blowflies. These strains were evaluated for the ability to prevent experimentally induced fly strike in a bioassay by using first instars. Results with undiluted spore/crystal preparations were variable, but they generally prevented fly strike on sheep maintained on pasture for 3-6 wk. Spore viability was satisfactory throughout the trials and environmental factors (e.g., precipitation and UV radiation) seemed to have minimal effect on persistence. The loss of fly strike protection in these experiments correlated with the movement of spore/crystal toxicity away from the skin as a result of wool growth. Solubilized protein preparations were not as potent as spore/crystal preparations and fly strike protection lasted only from 1 to 3 wk. Vegetative forms of the Cry1Ba-producing strains of Bt did not establish on the fleece of sheep, did not produce significant sporulation, and no protection against fly strike was achieved. Escherichia coli expressing recombinant Cry1Ba protein was toxic to larvae in vitro but did not effectively protect sheep from fly strike because blowfly larvae were able to establish readily 8 d posttreatment. In a single field experiment involving 80 sheep per group, a spore/crystal preparation from a Bt strain expressing Cry1Ba provided less protection from naturally acquired fly strike than afforded by a commercially available dip. PMID- 15666731 TI - A strategy for shuffling numerous Bacillus thuringiensis crystal protein domains. AB - Bacillus thuringiensis that produce Cry1Ba are toxic to Lucilia cuprina Wiedemann blow fly maggots in vivo, and when applied in quantity to sheep fleece, provide up to 6 wk protection against flystrike in the field. These strains also are toxic to Epiphyas postvittana (Walker) light brown apple moth caterpillars. B. thuringiensis expressing Cry1Db are toxic only to E. postvittana. When Cry1Ba and Cry1Db proteins are expressed within Escherichia coli, the recombinant bacteria have the same toxicity profile as the wild-type B. thuringiensis strain. In an effort to develop a Cry protein with improved blow fly toxicity, three different internal regions of Cry1Ba coding DNA, encoding all or part of domains I, II and III respectively were systematically exchanged with the corresponding region from a pool of other Cry protein coding DNAs. The chimeric products were then expressed in recombinant E. coli, and the resulting bacteria assayed for toxicity on L. cuprina and E. postvittana. Clones having insecticide bioactivity were characterized to identify the source of the replacement Cry domain. Despite successfully expressing a large number and variety of chimeric proteins within E. coli, many with measurable insecticidal activity, none of the chimeras had greater potency against L. cuprina than the wild-type Cry1Ba. Chimeric replacements involving domains I and II were rarely active, whereas a much higher proportion of domain III chimeras had some bioactivity. We conclude that shuffling of Cry coding regions through joining at the major conserved sequence motifs is an effective means for the production of a diverse number of chimeric Cry proteins but that such toxins with enhanced bioactive properties will be rare or nonexistent. PMID- 15666732 TI - Experimental analysis of the influence of pest management practice on the efficacy of an endemic arthropod natural enemy complex of the diamondback moth. AB - Maximizing the contribution of endemic natural enemies to integrated pest management (IPM), programs requires a detailed knowledge of their interactions with the target pest. This experimental field study evaluated the impact of the endemic natural enemy complex of Plutella xylostella (L.) (Lepidoptera: Yponomeutidae) on pest populations in commercial cabbage crops in southeastern Queensland, Australia. Management data were used to score pest management practices at experimental sites on independent Brassica farms practicing a range of pest management strategies, and mechanical methods of natural enemy exclusion were used to assess the impact of natural enemies on introduced cohorts of P. xylostella at each site. Natural enemy impact was greatest at sites adopting IPM and least at sites practicing conventional pest management strategies. At IPM sites, the contribution of natural enemies to P. xylostella mortality permitted the cultivation of marketable crops with no yield loss but with a substantial reduction in insecticide inputs. Three species of larval parasitoids (Diadegma semiclausum Hellen [Hymenoptera: Ichneumonidae], Apanteles ippeus Nixon [Hymenoptera: Braconidae], and Oomyzus sokolowskii Kurdjumov [Hymenoptera: Eulophidae]) and one species of pupal parasitoid Diadromus collaris Gravenhorst (Hymenoptera: Ichneumonidae) attacked immature P. xylostella. The most abundant groups of predatory arthropods caught in pitfall traps were Araneae (Lycosidae) > Coleoptera (Carabidae, Coccinelidae, Staphylinidae) > Neuroptera (Chrysopidae) > Formicidae, whereas on crop foliage Araneae (Clubionidae, Oxyopidae) > Coleoptera (Coccinelidae) > Neuroptera (Chrysopidae) were most common. The abundance and diversity of natural enemies was greatest at sites that adopted IPM, correlating greater P. xylostella mortality at these sites. The efficacy of the natural enemy complex to pest mortality under different pest management regimes and appropriate strategies to optimize this important natural resource are discussed. PMID- 15666733 TI - Infection and pathogenicity of the mosquito densoviruses AeDNV, HeDNV, and APeDNV in Aedes aegypti mosquitoes (Diptera: Culicidae). AB - These studies compared three genetically distinct mosquito densoviruses Aedes aegypti (AeDNV), Hemagogus equinus (HeDNV), and Aedes Peruvian (APeDNV) densoviruses in a laboratory investigation to begin to evaluate their potential as mosquito control agents. A real-time polymerase chain reaction (PCR) assay for quantification of viral genomes and a standardized mosquito infection protocol were developed. Mortality associated with exposure to AeDNV increased in a dose dependent manner, with the maximum mortality of 75.1% occurring in those organisms exposed to the highest dose of virus. The majority of death occurred as larvae. Similar results were observed with AeDNV produced from ground larvae and AeDNV produced from cell culture. Exposure of mosquitoes to HeDNV and APeDNV resulted in lower mortality, with values peaking at 33.5% for HeDNV and 27.8% for APeDNV. AeDNV-exposed larvae develop at a slower rate than nonexposed and HeDNV- and APeDNV-exposed larvae. Decreased virulence does not reflect a decrease in virus replication. PCR analysis of infectivity rates and titers in adults revealed reproduction of all three viruses, with an average viral titer of approximately 10 logs/mosquito after exposure to the highest dose of each virus. Accumulation of virus in the larval-rearing water was also observed with values approaching 10-11 logs/ml for each virus. These data indicate that there are dramatic differences in the pathogenicity among mosquito densoviruses. PMID- 15666734 TI - Bacillus thuringiensis variety kurstaki x aizawai applied to spruce flowers reduced Dioryctria abietella (Lepidoptera: Pyralidae) infestation without affecting seed quality. AB - We investigated the effects of Bacillus thuringiensis variety kurstaki x aizawai (Bt) on infestation levels of two lepidopteran insects as well as on seed quality in Norway spruce, Picea abies L. (Karst.) in central Sweden. Spruce flowers (female strobili) were sprayed with a 0.2% suspension (wt:wt) of the Bt preparation Turex 50 WP, 25,000 IU/mg in water. To expose even those lepidopteran larvae that feed exclusively embedded within the cone tissue, the Bt treatment was applied to open flowers, before they closed and developed into cones. The experimental design included three main factors: treatment (untreated control, water, or Bt), spruce genotype (three clones), and spraying time (spraying before, during, and after the phase of highest pollen receptivity). The Bt treatment reduced the proportion of cones infested by the cone worm Dioryctria abietella Den. et Schiff. (Lepidoptera: Pyralidae) from approximately 30 to 15%. There was no statistically significant treatment effect on the infestation rate of Cydia strobilella (L.) (Lepidoptera: Tortricidae). The Bt variety kurstaki x aizawai treatment caused no reduction in seed quality as measured by seed weight or percentage of nonfilled seeds. There was no difference in number of seeds per cone between the Bt-treated and untreated control cones. There was a significant effect of genotype on insect infestation rates, as well as on number of seeds per cone and seed weight. Neither level of insect damage nor any seed quality parameters were affected by time of application of the treatments. PMID- 15666735 TI - Effect of white grub developmental stage on susceptibility to entomopathogenic nematodes. AB - The pathogenicity of the entomopathogenic nematodes Heterorhabditis bacteriophora Poinar and Steinernema scarabaei Stock & Koppenhofer against different developmental stages of the Japanese beetle, Popillia japonica Newman, and the oriental beetle, Anomala (=Exomala) orientalis Waterhouse, were studied under laboratory conditions. The efficacy of S. scarabaei did not differ between second and third instars in P. japonica or A. orientalis or between small (young) and large (older) third instars in A. orientalis. However, H. bacteriophora efficacy decreased from first over second to third instar and also from small third instars to large third instars in A. orientalis but did not differ significantly between P. japonica larval stages. Once A. orientalis third instars had purged their intestines in preparation for pupation, no significant mortality by S. scarabaei and H. bacteriophora was observed. In contrast, P. japonica susceptibility to both nematode species gradually decreased from stage to stage from actively feeding third instars to pupae. In two additional experiments, we found no difference in Steinernema glaseri (Steiner) susceptibility between second and third instars of A. orientalis but an increase in S. scarabaei susceptibility from the second to third instar of Asiatic garden beetle, Maladera castanea (Arrow). Our observations combined with those of previous studies with other nematode and white grub species show that nematode efficacy against white grub developmental stages varies with white grub and nematodes species, and no generalization can be made. PMID- 15666736 TI - Quantifying individual fruit fly consumption with Anastrepha suspensa (Diptera: Tephritidae). AB - We needed a technique to compare the consumption of baits by individual Carribbean fruit fly, Anastrepha suspensa (Loew). By improving consumption and determining individual dose, we could lower pesticide concentration while retaining bait/pesticide efficacy and potentially reduce the environmental impact of fruit fly bait/pesticide eradication methods. We report here a precise dye based technique for the quantification of consumption by individual adult A. suspensa fruit flies. Fluorescein, measured at 491 nm, and cresol red, measured at 573 nm, were efficiently extracted with 0.1 M NaOH and quantified with a spectrophotometer. The lower limit for this method with 0.1% dye concentration is 300 nl consumed by an individual fly. Dye movement to the hindgut and possible defecation occurred in approximately 4 h; maximum ingestion occurred in approximately 1 h. Maximum experimental time is limited to 4 h. Flies preferred feeding upside down compared with right side up when given a choice; consumption was equal when flies were given no choice of feeding position. Thus, maximum bait/pesticide efficacy might be achieved with an upside-down presentation. Regurgitation led to a 100% overestimation of actual consumption with the J-tube presentation of food. Our individual fly consumption technique will be useful in comparing consumption in phagostimulant studies, estimating dose in oral toxicity tests, differentiating behavioral and physiological resistance in toxicant studies, ultimately leading to improved bait/pesticide methods and reduced environmental impact of area wide fruit fly eradication programs. This technique could be applied to studies of tephritid consumption, to the consumption of other insects, and to regurgitation studies. PMID- 15666737 TI - A novel nonchemical method for quarantine treatment of fruits: California red scale on citrus. AB - A process for removing or killing California red scale, Aonidiella aurantii (Maskell), from citrus fruit as a postharvest treatment was evaluated. The process subjects the fruit to vacuum, steam, and vacuum that physically removes red scale from the fruit and kills those scales that are not removed from the fruit. Different numbers of cycles and steam temperatures were compared for efficacy in removing scale from lemons or killing those that remained. Multiple (two to three) cycles removed up to 96% of first molt scales on the fruit, but they were much less effective in removing other stages, especially those that had advanced beyond the second instar. However, it was extremely effective in killing the scales remaining on the fruit. Although this process does not eliminate cosmetic damage caused by scale presence, it might be used in combination with high-pressure washers currently used in packing houses to allow importers and exporters to meet the most stringent quarantine requirements. Because of its killing power, this technique should be tried on other insects and commodities to see whether it can be substituted for certain uses of methyl bromide. PMID- 15666738 TI - Thermal death kinetics of red flour beetle (Coleoptera: Tenebrionidae). AB - While developing radio frequency heat treatments for dried fruits and nuts, we used a heating block system developed by Washington State University to identify the most heat-tolerant life stage of red flour beetle, Tribolium castaneum (Herbst), and to determine its thermal death kinetics. Using a heating rate of 15 degrees C/min to approximate the rapid heating of radio frequency treatments, the relative heat tolerance of red flour beetle stages was found to be older larvae > pupae and adults > eggs and younger larvae. Lethal exposure times for temperatures of 48, 50, and 52 degrees C for the most heat-tolerant larval stage were estimated using a 0.5th order kinetic model. Exposures needed for 95% mortality at 48 degrees C were too long to be practical (67 min), but increasing treatment temperatures to 50 and 52 degrees C resulted in more useful exposure times of 8 and 1.3 min, respectively. Red flour beetle was more sensitive to changes in treatment temperature than previously studied moth species, resulting in red flour beetle being the most heat-tolerant species at 48 degrees C, but navel orangeworm, Amyelois transitella (Walker), being most heat tolerant at 50 and 52 degrees C. Consequently, efficacious treatments for navel orangeworm at 50 52 degrees C also would control red flour beetle. PMID- 15666739 TI - Northward migration of Helicoverpa armigera (Lepidoptera: Noctuidae) and other moths in early summer observed with radar in northern China. AB - The northward migration of Helicoverpa armigera (Hubner) and other moths in early summer was observed with radar in 2001 and 2002 at Langfang, Hebei province, China. Migratory flights typically occurred at heights up to 1.2 km above ground level (AGL), and high density layer concentrations frequently formed at 200-300 m AGL. Adult moths of local populations took off at dusk and ceased flight approximately 0.5 h before sunrise with area density peaking approximately 35 min after dusk. A strong dumb-bell pattern of echoes on the plan position indicator screen, indicating collective orientation of the targets, was evident in 2001, when targets were typically moving toward the northeast and when layering was associated with a temperature inversion and maximum wind speed. By contrast, there was no notable dumb-bell pattern in 2002, when targets were moving toward the northwest. However, orientations calculated from target and wind velocities showed that downwind common orientation also was occurring in 2002 and that the direction varied with the wind direction. The probable sources of the H. armigera observed at Langfang were deduced to be Henan province in 2001 and Shandong province in 2002, and the destination regions were Liaoning and Inner Mongolia. PMID- 15666740 TI - Laboratory evaluation of six algal species for larval nutritional suitability of the pestiferous midge Glyptotendipes paripes (Diptera: Chironomidae). AB - Glyptotendipes paripes Edwards midge larval growth, development, survival, emerging adult size, and food digestibility when provided with six species of algae as food were studied in the laboratory. For the study, eggs from G. paripes adults maintained in the laboratory were reared to the adult stage at 30 degrees C for 60 d on pure culture of each algal species at densities of 0.4, 0.1, and 0.02 mg of algae (fresh weight) per milliliter, as a sole food source. All larvae reared on Microcystis sp., Botryoccocus braunii, and Scenedesmus quadricauda died before completing development. The only larvae to complete development to adult were those reared on 0.4 mg/ml Lyngbia cf. aeruginosa (44.0 d), Anabaena flos aquae (29.7 d), and Chlorella keslerii (44.8 d). No significant differences in body size of the adults achieving complete development on the three algal species were found. Algal digestion, measured by comparing amounts of live and dead algal cells in remains of cultures used for feeding and in larval excrement, revealed that >95% of all L. cf. aeruginosa, A. flos-aquae, and Microcystis sp. cells were digested; for C. keslerii, 13% of cells were digested, whereas little or no digestion of B. braunii and S. quadricauda was observed. To evaluate the effects of algal species on larval growth, laboratory-reared (on artificial food) late third/early fourth instars of G. paripes were fed individual algal species, and 10 d later, body mass changes were recorded and compared with nonfed larvae. Body mass of larvae reared on L. cf. aeruginosa and A. flos-aquae significantly increased, whereas those provided Microcystis sp. and the nonfed larvae showed significant body mass reductions. Overall, B. braunii and S. quadricauda were not suitable as larval food, probably due to their low digestibility, and Microcystis sp. because of its toxicity. This study identifies some algae that do and others that do not support G. paripes larval growth. The information is useful in understanding the feeding habits of G. paripes larvae as some of these algae occur in the larval environments of this pestiferous midge. PMID- 15666741 TI - Effect of rearing strategy and gamma radiation on field competitiveness of mass reared codling moths (Lepidoptera: Tortricidae). AB - We compared the field competitiveness of sterile codling moth, Cydia pomonella (L.), males mass-reared through diapause or standard production protocols and treated with either 150 or 250 Gy of gamma radiation. Evaluations were performed during spring and summer 2003 by using field release-recapture tests. Released males were recaptured using traps baited with synthetic pheromone or with virgin females. In addition, mating tables baited with virgin females were used in the summer to assess the mating competitiveness of the released moths. Field performance of released males was significantly improved by rearing through diapause and by lowering the dose of radiation used to treat the insects. These effects were observed during spring when evening temperatures were relatively cool and in summer when evening temperatures were high. These effects were observed regardless of the sampling method (i.e., capture in pheromone-baited traps, virgin female-baited traps, or in mating tables). There were significant interactions between larval rearing strategy and radiation dose with respect to day of recapture. The effect of rearing strategy on male performance was observed immediately after release, whereas the effect of dose of radiation was usually delayed by 2-3 d. In general, the best treatment for improving codling moth male field performance was a combination of rearing through diapause and using a low dose of radiation (150 Gy). The difference in performance when insects were treated with 150 or 250 Gy was greater when males had been reared using standard (nondiapause) rearing protocols, suggesting that diapause rearing may attenuate some of the negative effects of the higher doses of radiation. PMID- 15666742 TI - Phenology and infestation patterns of plum curculio (Coleoptera: Curculionidae) on four highbush blueberry cultivars. AB - The plum curculio, Conotrachelus nenuphar (Herbst), is a well known pest in apple and peach orchards, but it also is capable of having an economic impact in highbush blueberries. Host phenology and plum curculio oviposition patterns were determined on four highbush blueberry cultivars differing in fruit maturation period. Numbers of oviposition scars were higher on early- ('Weymouth') and mid season ('Duke' and 'Bluecrop') blueberries than on late-season 'Elliott' in 2001, 2002, and 2003. In 2002, eggs were first present on the three earliest cultivars 21 d before those on 'Elliott', whereas eggs were found on 'Elliott' >40 d after the last sample with eggs for the other three cultivars. The pattern of host phenology and infestation levels suggested that plum curculio oviposition synchronizes well with the availability of suitable fruit for oviposition on early and mid-season cultivars compared with a late-season cultivar of highbush blueberries. The implications of a transition to use of reduced-risk insecticides are discussed in relation to plum curculio management. PMID- 15666743 TI - Effects of two biorational insecticides, spinosad and methoxyfenozide, on Spodoptera littoralis (Lepidoptera: Noctuidae) under laboratory conditions. AB - The toxicity of two biorational insecticides, spinosad (Tracer) and methoxyfenozide (RH-2485), was tested against eggs, larvae, and pupae of the noctuid Spodoptera littoralis (Boisduval). In the first experiment, filter paper circles containing egg masses of two different age classes, young (<24 h old) and old (24-48 h old), were dipped in different concentrations of each insecticide diluted in either water or acetone. No ovicidal activity was recorded when insecticides were diluted in water. In contrast, when insecticides were diluted in acetone, both egg age classes generally showed a concentration-dependent response for both compounds. Mortality of larvae that hatched from both egg age classes was significantly increased, compared with control larvae, at all concentrations of both insecticides when diluted in water or acetone alike. The prevalence of mortality was similar with each insecticide. In the second experiment, third instars of S. littoralis were fed semisynthetic diet containing different concentrations of both insecticides. According to LC50 values, no significant differences were observed between spinosad (2.11 mg [AI]/kg diet) and methoxyfenozide (3.98 mg [AI]/kg diet) after 48 h of treatment, based on the overlap of 95% CL. Toxic effects on the mortality of pupae, adult emergence, and the prevalence of deformed adults after topical application on young pupae also were examined. Only methoxyfenozide caused pupal mortality and deformed adults. Our results suggest that spinosad and methoxyfenozide are potentially potent compounds for control of S. littoralis. PMID- 15666744 TI - Acaricidal activities of extracts of Stellera chamaejasme against Tetranychus viennensis (Acari: Tetranychidae). AB - Extracts of a perennial and poisonous weed, Stellera chamaejasme L. (Thymelaeaceae) were bioassayed to determine their acaricidal activities against Tetranychus viennensis Zacher (Acari: Tetranychidae) in the laboratory. Extracts had both contact and systemic toxicity to the mite. After liquid chromatography and thin layer chromatography, the extract was further concentrated, separated, and bioassayed. This study is the first to report the acaricidal property of S. chamaejasme and its potential as a botanical pest control agent. PMID- 15666745 TI - Changes of zeatin riboside content in rice plants due to infestation by Nilaparvata lugens (Homoptera: Delphacidae). AB - The effect of Nilaparvata lugens (Stal) (Homoptera: Delphacidae), infestation on the content of zeatin ribosides (ZR) in rice plants was investigated with enzyme linked immunosorbent assay. Hydroponics experiments were conducted on 'Zhendao 2' rice, in which plants were subjected to N. lugens infestation at three nonhopperburn-causing densities (15, 30, and 60 nymphs per hill) for 2, 4, 6, and 8 d and at one hopperburn-causing density (240 nymphs per hill) for 2, 4, and 6 d, respectively. When rice plants were infested at the nonhopperburn-causing densities, ZR content in leaves varied significantly with the infestation density. Compared with the control plants, ZR content in rice leaves decreased significantly after infestation by 60 nymphs per hill for 2 d, but it tended to increase due to prolonged infestation at all the nonhopperburn-causing densities. In contrast, ZR content in rice roots significantly reduced after the plants being infested at the density of 15 nymphs for 2 d and at all densities for prolonged duration, except for the plants infested by 60 nymphs for 6 and 8 d, in which the ZR content increased or did not change significantly. However, infestation at the hopperburn-causing density caused significant reduction in ZR content in rice roots, regardless of infestation duration, and in rice leaves from the plants subjected to 2-d infestation. These results are discussed in relation to the possible physiological reaction of rice plants to N. lugens infestation and the resultant severe damage or hopperburn. PMID- 15666746 TI - Effect of rice bug Leptocorisa oratorius (Hemiptera: Alydidae) on rice yield, grain quality, and seed viability. AB - The effect of Leptocorisa oratorius (F.) on the yield, grain quality, and seed viability of four rice, Oryza sativa L., lines was studied. Three of the lines, C2, IR64, and PSBRc20, are grown in the Philippines. The fourth, IR72164-201-1 is an unreleased experimental line of an O. sativa japonica x O. sativa indica cross. Each line was exposed to four infestation densities for 21 d. L. oratorius feeding produced unfilled and partially filled grains, resulting in a negative correlation of yield to rice bug density. When filled grains were sown, germination rates were negatively correlated with rice bug densities. The percentage of discolored grains was positively correlated with L. oratorius density on all rice lines. At the same infestation rates, PSBRc20 and IR64 had higher yields, less damaged grain, and higher germination rates than IR72164-201 1 and C2, suggesting host plant tolerance to rice bug feeding. The economic injury levels (EILs) currently used for rice bug management are based solely on yield loss estimates. The results of this study suggest that EIL for rice bugs should be revised to take into account reductions in grain quality and seed germination rates in addition to yield loss. PMID- 15666747 TI - Boll injury and yield losses in cotton associated with brown stink bug (Heteroptera: Pentatomidae) during flowering. AB - Brown stink bug, Euschistus servus (Say), was infested on cotton, Gossypium hirsutum L., plants during reproductive stages to determine the effects on boll injury and seedcotton yield. During each week in 2002 and 2003, significantly more bolls with > or = 1 injured locule, bolls with > or = 2 injured locules, and bolls with discolored lint were recorded on stink bug-infested plants compared with that on noninfested plants. Significantly fewer bolls displayed external injury on the boll exocarp compared with bolls with only internal locule injury. Boll injury was significantly underestimated by the presence of external symptomology. The boll population increased 6.6- and 5.1-fold from weeks 1-5 in 2002 and 2003, respectively. There was a corresponding 6.2- and 4.6-fold increase in 2002 and 2003, respectively, for total bolls injured from weeks 1-5. Percentage of boll injury ranged from 10.7 (week 4) to 27.4 (week 2) in 2002 and from 9.2 (week 3) to 16.0 (week 2) in 2003. Percentage of injury was greatest during weeks 1 and 2 in both years and also in week 5 in 2002. Brown stink bug significantly reduced seedcotton yield of bolls present on cotton plants during weeks 1, 2, and 5 in 2002 and in weeks 4 and 5 in 2003. However, total seedcotton yield, as a function of bolls exposed to brown stink bug and subsequent bolls produced on plant in the absence of stink bugs, was not significantly different for plots infested during weeks 1-4 in 2002 and weeks 1-3 in 2003. Flowering period and boll population influence the severity of stink bug injury on seedcotton yield. Infestation timing and number of bolls should be considered, in addition to insect densities, when initiating treatments against brown stink bug. PMID- 15666748 TI - Evaluation of the potential role of glufosinate-tolerant rice in integrated pest management programs for rice water weevil (Coleoptera: Curculionidae). AB - The impact of a herbicide-tolerant rice, Oryza sativa L., variety was assessed for its resistance to rice water weevil, Lissorhoptrus oryzophilus Kuschel (Coleoptera: Curculionidae), and its place in current integrated pest management (IPM) programs. Greenhouse experiments were conducted to evaluate the resistance of a glufosinate-tolerant rice variety and its glufosinate-susceptible parent line Bengal to the rice water weevil in the presence and absence of glufosinate applications. The LC50 dose-response and behavioral effects of glufosinate on adult rice water weevils also were studied. Field studies investigated the impacts of glufosinate-tolerant rice on rice water weevil management in the presence and absence of glufosinate under early and delayed flood conditions. Greenhouse studies demonstrated that in the absence of glufosinate, oviposition was 30% higher on the glufosinate-tolerant rice line than on Bengal rice or on glufosinate-tolerant line treated with recommended rates of commercially formulated glufosinate. Applications of glufosinate to glufosinate-tolerant rice resulted in a 20% reduction in rice water weevil larval densities compared with nontreated glufosinate-tolerant rice. The LC50 of glufosinate against adult rice water weevil was nearly 2 times the concentration recommended for application to glufosinate-tolerant rice. There was no difference in the amount of leaf area consumed by adult rice water weevils on glufosinate-treated and nontreated foliage. The absence of direct toxicity of glufosinate to rice water weevil at recommended glufosinate use rates and lack of behavioral effects suggest that the reduction in rice water weevil densities observed after glufosinate applications resulted from herbicide-induced plant resistance. Field experiments showed that neither rice variety nor herbicide use affected larval densities; however, delaying flood and applying insecticide effectively reduced numbers of rice water weevil larvae. PMID- 15666749 TI - Landscape features and spatial distribution of adult northern corn rootworm (Coleoptera: Chrysomelidae) in the South Dakota areawide management site. AB - The northern corn rootworm, Diabrotica barberi Smith & Lawrence (Coleoptera: Chrysomelidae), creates economic and environmental concerns in the Corn Belt. To supplement the population control tactics of the Areawide Pest Management Program in Brookings, SD, geographic information systems were used from 1997 to 2001 to examine the spatial relationships between D. barberi population dynamics and habitat structure, soil texture, and elevation. Using the inverse distance weighted interpolation technique, D. barberi population density maps were created from georeferenced emergence and postemergence traps placed in maize, Zea mays L., fields. For each year, these maps were overlaid with vegetation, soil, and elevation maps to search for quantitative relationships between pest numbers and landscape structure. Through visual interpretation and correlation analysis, shifts in landscape structure, such as size, number, and arrangement of patches were shown to associate with D. barberi population abundance and distribution in varying degrees. D. barberi were found in greater proportions than expected on loam and silty clay loam soils and on elevations between 500 and 509 m. An understanding of the interactions between D. barberi population dynamics and landscape variables provides information to pest managers, which can be used to identify patterns in the landscape that promote high insect population density patches to improve pest management strategies. PMID- 15666750 TI - Biology of Hylobitelus xiaoi (Coleoptera: Curculionidae), a new pest of slash pine, Pinus elliottii. AB - The biology of the weevil Hylobitelus xiaoi Zhang was studied in both field and laboratory in Shangyou, Jiangxi Province, China. This species required 2 yr to complete one generation with overwintering by adults in pupal chambers and larvae in galleries in the bark of host trees. Adults emerged from early March to early April and fed on the inner bark of branches of the trees. Adults fly little. Adults exhibited a diel periodicity, climbing up the trees around sunset and returning to the tree base the next morning. The mean preovipostion period was 46 d. Oviposition commenced in early May and ended in late August. The average fecundity per female was 36 eggs. Overwintered adult females and males lived 208 and 227 d, respectively. At 25 degrees C, the mean egg incubation period was 13 d. In the field, egg hatch occurred in 12-15 d with 83% survival. There were five to seven instars. At 25 degrees C, duration of the larval stage averaged 129 d. Pupation commenced in late August. At 25 degrees C, pupation averaged 20 d. In the field, pupation required 20-26 d. Transformation to adults occurred from late September to October. New adults remained in the pupal chambers until the next year. Infection by Beauveria bassiana Vuill. occurred in 0.8% of the pupae and 8.8% of the overwintered adults. PMID- 15666751 TI - Differential susceptibility of poplar hybrids to the aphid Chaitophorus leucomelas (Homoptera: Aphididae). AB - After its recent introduction to Chile, the aphid Chaitophorus leucomelas Koch is becoming a serious pest affecting commercial poplar, Populus spp., plantations. The pattern of natural infestation of C. leucomelas among poplar hybrids with different pedigrees and the aphid intrinsic rate of increase (r(m)), of C. leucomelas were assessed in the field. In most of the hybrids, aphid abundance peaked in March (late summer). Among 12 types of poplar crosses, [(P. trichocarpa Torr. & Gray x P. deltoides Bartram ex Marshall) x (P. trichocarpa x P. deltoides)] and [(P. trichocarpa x P. maximowiczii Henry) x P. maximowiczii] showed the highest and lowest aphid densities, respectively. A trend to find more aphids in branch bases was apparent. The intrinsic rate of C. leucomelas increase was higher in [(P. trichocarpa x P. deltoides) x P. deltoides] hybrids, and lower in [(P. trichocarpa x P. maximowiczii) x P. trichocarpa] hybrids. Aphid density and performance were higher in hybrids with P. deltoides parentage, whereas hybrids with P. maximowiczii parentage showed lower aphid densities and performance. Hybrids with P. nigra L. parentage, namely, [P. trichocarpa x P. nigra], also had high aphid density, but aphid performance was lower compared with hybrids with P. deltoides parentage. These results suggest that among poplar hybrids studied, susceptibility to C. leucomelas is inherited through P. deltoides, whereas resistance seems to be inherited through P. maximowiczii. Thus, P. maximowiczii hybrids are recommended for commercial or ornamental planting programs in zones where there is a high risk of aphid infestation. PMID- 15666752 TI - Long-term benefits to the growth of ponderosa pines from controlling southwestern pine tip moth (Lepidoptera: Tortricidae) and weeds. AB - The southwestern pine tip moth, Rhyacionia neomexicana (Dyar) (Lepidoptera: Tortricidae), is a native forest pest that attacks seedlings and saplings of ponderosa pine, Pinus ponderosa Dougl. ex Laws, in the southwestern United States. Repeated attacks can cause severe deformation of host trees and significant long-term growth loss. Alternatively, effective control of R. neomexicana, vegetative competition, or both in young pine plantations may increase survival and growth of trees for many years after treatments are applied. We test the null hypothesis that 4 yr of R. neomexicana and weed control with insecticide, weeding, and insecticide plus weeding would not have any residual effect on survival and growth of trees in ponderosa pine plantation in northern Arizona 14 yr post-treatment, when the trees were 18 yr old. Both insecticide and weeding treatment increased tree growth and reduced the incidence of southwestern pine tip moth damage compared with the control. However, weeding alone also significantly increased tree survival, whereas insecticide alone did not. The insecticide plus weeding treatment had the greatest tree growth and survival, and the lowest rate of tip moth damage. Based on these results, we rejected our null hypothesis and concluded that there were detectable increases in the survival and growth of ponderosa pines 14 yr after treatments applied to control R. neomexicana and weeds. PMID- 15666753 TI - Feeding and attraction of Agelastica coerulea (Coleoptera: Chrysomelidae) to Betulaceae plants. AB - The feeding and attraction of Agelastica coerulea (Baly) to nine species of Betulaceae in four genera (Alnus, Betula, Carpinus, and Corylus) were examined in the laboratory by using choice and no-choice as well as olfactometer bioassays. In no-choice feeding bioassays with A. coerulea larvae, Alnus hirsuta (Spach) Ruprecht, Alnus japonica Steudel, Corylus sieboldiana variety mandshurica (Max.) c. k. Schneider, and Corylus heterophylla variety thunbergii Blume were the most preferred, whereas Betula davurica Pallas, Betula schmidtii Regel, and Carpinus cordata Blume were the least preferred. The larvae showed moderate preference for Alnus maximowiczii Callier and Carpinus tschonoskii variety brevicalycina Nakai. In choice feeding bioassays, no differences in preference between A. hirsuta and A. japonica were observed. However, there were significant differences in preference between A. hirsuta and each of the other seven plant species. In olfactometer bioassays with adult female A. coerulea, fresh leaf odor from A. hirsuta attracted significantly more adults than that from A. japonica and C. sieboldiana variety mandshurica. These results indicate that A. coerulea differed in its ability to discriminate among the Betulaceae plants at the feeding preference and attraction of host selection. PMID- 15666754 TI - Optimization of a valine:isoleucine methyl ester pheromone blend and comparison of Robbins and Trece traps for capture of Phyllophaga anxia (Coleoptera: Scarabaeidae) in Rhode Island. AB - Eight ratios of L-valine:L-isoleucine methyl esters were tested in Robbins traps for capture of Phyllophaga anxia (LeConte) adult males. The 90:10, 80:20, and 60:40 ratios of valine:isoleucine were the most effective blends for capture of beetles in Rhode Island. Females were captured in small numbers in some traps but not consistently to any particular blend. Other male Phyllophaga species captured included Phyllophaga fusca (Frolich), Phyllophaga forsteri (Burmeister), P. hirsuta (Knoch), and P. marginalis (LeConte). The number of these species collected was low, and it was not possible to determine whether they were attracted to any particular pheromone blend. Peak captures of P. anxia males occurred 31 May in 1999 and 2002 in Kingston, RI. The standard Japanese beetle trap manufactured by Trece (Adair, OK) captured significantly more beetles than the Robbins trap. Because the Trece trap is already marketed for Japanese beetles, a lure and trapping system can be adopted for P. anxia. PMID- 15666755 TI - Rainfastness of a microencapsulated sex pheromone formulation for codling moth (Lepidoptera: Tortricidae). AB - The rainfastness of a microencapsulated sex pheromone formulation for codling moth, Cydia pomonella (L.), was evaluated in a series of laboratory experiments with detached apple, pear, and walnut leaves. Increasing the intensity and duration of simulated rainfall significantly increased the removal of microcapsules from both the top and bottom of apple leaves. The removal of microcapsules was significantly higher from the top versus the bottom of leaves at all rates tested. Leaf angle was a significant factor affecting the removal of microcapsules from the top surface of apple leaves with fewer microcapsules removed, because leaves were oriented with a steeper downward angle. Both leaf surfaces of apple and pear retained a higher proportion of microcapsules than walnut leaves, and the bottom surface of apple leaves retained significantly more than pear leaves. Three spray adjuvants were evaluated as stickers for microcapsules. No difference was found in the number of microcapsules deposited on apple leaves among three stickers tested at rates from 0.06 to 0.25%. However, in a second test a latex sticker significantly increased the deposition of microcapsules on apple leaves compared with a polyvinyl polymer and a pine resin sticker at a rate of 0.06%. Significantly more microcapsules were retained on the bottom versus the top of apple leaves with all stickers. The latex and polyvinyl stickers significantly increased the retention of microcapsules versus the pine resin sticker and the control on apple leaves. In another test, the addition of 0.06% latex sticker did not increase the deposition of microcapsules on any of the three leaf types. However, the addition of the latex sticker significantly increased the retention of microcapsules on the top of apple and pear leaves and the bottom of apple leaves. The addition of a latex sticker did not affect the retention of microcapsules on walnut leaves. PMID- 15666756 TI - Species and cultivar influences on survival and parasitism of fall armyworm. AB - Interactions between host plant resistance and biological control may benefit or hinder pest management efforts. Turfgrass cultivars have rarely been tested for extrinsic resistance characteristics such as occurrence and performance of beneficial arthropods on plant genotypes with resistance to known turf pests. Parasitism of fall armyworm, Spodoptera frugiperda (J.E. Smith), among six turfgrass genotypes was evaluated. The six grasses tested [Sea Isle-1 and 561-79 seashore paspalum, Paspalum vaginatum Swartz; TifSport and TifEagle hybrid Bermuda grass, Cynodon dactylon (L.) x C. transvaalensis (Burtt-Davy); and Cavalier and Palisades zoysiagrass, Zoysia japonica von Steudel and Z. matrella (L.) Merrill, respectively] represented a range in resistance to S. frugiperda. Differential recovery of larvae released as first instars reflected this gradient in resistance of Cavalier > or = Palisades > or = TifSport = TifEagle > or = 561- = Sea Isle-1 Larval recovery (percentage of initial number released) was greatest in May, less in July and August, and least in October, probably reflecting the increase in activity of on-site predators and disease pressure. Parasitism of the fall armyworm by the braconid Aleiodes laphygmae Viereck varied among turfgrass genotypes. Parasitism was greatest during July. In total, 20,400 first instars were placed in the field; 2,368 were recovered; 468 parasitoids were subsequently reared; 92.2% were A. laphygmae. In the field, the greatest percentage of reduction in S. frugiperda larvae by A. laphygmae occurred on the armyworm susceptible seashore paspalums (51.9% on Sea Isle-1 in July). Cotesia marginiventris Cresson and Meteorus sp. also were reared from collected larvae. No parasitoids were reared from larvae collected from resistant Cavalier zoysiagrass. A. laphygmae and C. marginiventris were reared from larvae collected from the other five grass cultivars. No parasitoids of older larvae or pupae were observed. PMID- 15666757 TI - Comparison of management strategies for squash bugs (Hemiptera: Coreidae) in watermelon. AB - Two watermelon pest management practices, a squash trap crop and a standard recommendation using soil-applied carbofuran, were compared using large-scale field plots to assess trap crop suitability as a replacement for the standard in 2000, 2001, and 2002. In both systems, foliar insecticide applications were used to control squash bugs when populations exceeded threshold levels. During 2001 and 2002, a treatment of untreated watermelon was used. Early season adult insects, from seedling to fruit set, are most critical for watermelon. Significantly fewer early adult bugs were found on watermelon in the trap crop than in the standard recommended practice in 1 of 3 yr. In both years, significantly fewer adult squash bugs were found in watermelon in the trap crop than in untreated fields. The standard recommended practice significantly reduced adult squash bugs in watermelon compared with the untreated in 1 of 2 yr. There was no significant correlation of watermelon yield and squash bug density, indicating that squash bug densities were too low to impact yield. Although squash bugs were reduced significantly by the trap crop, marketable watermelon yields were lower in the squash trap crop than in untreated watermelon, suggesting that pest management treatments may interfere with crop productivity factors other than squash bug colonization. Results suggest that mid-season production squash bug should be managed by monitoring populations and using insecticides as needed rather than using at-plant treatment. Further research is needed to compare treatments during early-season production. PMID- 15666758 TI - Evaluation of traps and toxicants in an attract-and-kill system for Rhagoletis mendax (Diptera: Tephritidae). AB - A series of laboratory and field assays were performed to evaluate new methods for an attract-and-kill approach to control blueberry maggot, Rhagoletis mendax Curran. In laboratory assays, fly mortality was similar among insecticide-treated matted paper, plastic, and biodegradable sphere traps. Plastic and biodegradable traps baited with ammonium acetate captured significantly more flies than baited matted paper traps in field assays, whereas fly captures were similar in baited matted paper traps and unbaited plastic sphere traps. Treatments containing imidacloprid resulted in the highest amount of fly knockdown, and spheres coated with this insecticide still had significant knockdown after 6 wk of field exposure. Fly mortality was comparable on sphere traps coated with fipronil and imidacloprid, with both resulting in significantly more fly mortality than the control when flies were exposed to traps that had been weathered for 4 wk in the field. Spheres coated with acetamiprid resulted in fly mortality, but further evaluation is necessary to determine the potential of this compound. Deltamethrin was ineffective in causing fly knockdown at the rate tested. Our results indicate that fipronil and imidacloprid have potential as insecticidal coatings on either plastic or biodegradable spheres in an attract-and-kill system for control of R. mendax. PMID- 15666759 TI - Disruption of reproductive activity of Coptotermes formosanus (Isoptera: Rhinotermitidae) primary reproductives by three chitin synthesis inhibitors. AB - Effects of the chitin synthesis inhibitors (CSIs) diflubenzuron, hexaflumuron, and lufenuron on the Formosan subterranean termite, Coptotermes formosanus Shiraki, primary reproductives were studied in the laboratory. Incipient colonies were established by collecting and pairing C. formosanus alates and placing them in dishes containing an artificial diet. Three groups of 56 colonies each were fed with a diet containing 10 ppm of one of each of the CSIs and were compared with a control group fed with an untreated diet. All eggs oviposited by treated young queens failed to hatch at the end of 6 mo. Estimated queen fecundity was significantly lower in the lufenuron treatment compared with the control group. Fecundity of hexaflumuron-treated queens did not differ significantly from that of queens from the control group and the other treatments. Adult mortality was significantly higher in the diflubenzuron and lufenuron treatments than in the control group at the end of 6 mo., but not in the hexaflumuron treatment. All the pairs died within 8 mo. in the diflubenzuron and lufenuron treatments, even after treatment was suspended at the end of 6 mo. Mortality in the hexaflumuron treatment was significantly higher than in the control group by the end of 9 mo. The three CSIs tested eliminated reproduction in C. formosanus by preventing egg hatching and induced adult mortality. Possible mechanisms by which CSIs induce termite adult death are discussed. PMID- 15666761 TI - Use of a bait impact index to assess effects of bait application against populations of Formosan subterranean termite (Isoptera: Rhinotermitidae) in a large area. AB - The effects of bait applications on the overall activity of the Formosan subterranean termite, Coptotermes formosanus Shiraki, in a town of 1.2 km2 were assessed between 2001 and 2003 by using a bait impact index. The index incorporated the amount of baits applied and the distance between bait application loci and the monitoring stations from which termite activity was measured. Even with the collaboration of only 35% of the residents and incomplete treatment records, the bait impact index demonstrated that more baits applied in proximity to monitoring loci significantly caused the decline of C. formosanus activity. PMID- 15666760 TI - Effect of delayed toxicity of chemical barriers to control Argentine ants (Hymenoptera: Formicidae). AB - Brief exposures of Argentine ants to four different insecticide treatments, bifenthrin, beta-cyfluthrin, bifenthrin + beta-cyfluthrin, and fipronil, were conducted to determine Kaplan-Meier product limit survivorship percentiles (SPs) at 21-23 and 27-29 degrees C. Bifenthrin, beta-cyfluthrin, and bifenthrin + beta cyfluthrin provided rapid kill at 21-23 degrees C with SP10 values ranging from 11.2 to 33.7 min. Fipronil provided delayed toxicity at 21-23 degrees C with SP10 values ranging from 270 to 960 min. At 27-29 degrees C, all of the SP10 values significantly decreased. Field tests in which Argentine ants were induced to forage across insecticide-treated surfaces were used to determine the effect that speed of action has on foraging and recruitment ability, and whether these insecticides are repellent. The slower-acting fipronil allowed a greater amount of foraging and consequently a greater fraction of the colony to be exposed, whereas fast-acting bifenthrin, beta-cyfluthrin, and bifenthrin + beta-cyfluthrin inhibited recruitment, resulting in fewer ants being exposed and killed. Implications for controlling ants by using perimeter barrier treatments are discussed. PMID- 15666762 TI - Contamination affects the performance of insecticidal baits against German cockroaches (Dictyoptera: Blattellidae). AB - The effects of contamination of insecticidal bait formulations, by using mint oil and silica aerogel, were evaluated in a series of laboratory experiments against the German cockroach, Blattella germanica (L.). Bait consumption at 3 d for uncontaminated baits ranged from 0.01 g for Avert dust to 0.399 g for Combat dry bait with hydramethylnon presented in a station. LT50 values for uncontaminated gel bait deposits ranged from 0.4 d for PreEmpt containing imidacloprid to 4.1 d for Maxforce containing hydramethylnon. As a group, significantly more gel bait was consumed than solid formulations even when both formulations had similar concentrations of the same active ingredient. As a result, gel baits were significantly more toxic than solid formulations. Application of mint oil directly to bait deposits significantly decreased bait consumption and increased overall LT50 values. When bait formulation types were examined individually, there was no difference in consumption or toxicity between contaminated and uncontaminated gel formulations. Contaminated solid baits, however, had significantly greater LT50 values and less consumption than uncontaminated solid baits. Gel formulations probably absorbed the contaminants and removed them from the surface of the bait deposits resulting in normal bait consumption and toxicity. Gel and solid bait deposits, inside plastic exposure stations or alone with no station and placed onto mint oil-contaminated substrates, had significantly lower bait consumption and greater LT50 values than baits placed on uncontaminated substrates. Contamination of a baited area is more likely than contamination of just the bait deposit and therefore a more realistic test of the effects of contamination on bait performance. The importance of contamination to the performance of cockroach baits is discussed. PMID- 15666763 TI - Fipronil resistance in the diamondback moth (Lepidoptera: Plutellidae): inheritance and number of genes involved. AB - Bioassays (at generation 1, G1) using fipronil, spinosad, indoxacarb, and Bacillus thuringiensis toxins Cry1Ac and Cry1Ca with a newly collected field population of Plutella xylostella (L.) from farmers fields in the Cameron Highlands, Malaysia, indicated a resistance ratio of approximately 400-, 1,170-, 330-, 2,840-, and 1,410-fold, respectively, compared with a laboratory susceptible population of P. xylostella (ROTH). At G3, the field-derived population was divided into two subpopulations, one was selected (G3 to G7) with fipronil (fip-SEL), whereas the second was left unselected (UNSEL). Bioassays at G8 found that selection with fipronil gave a resistance ratio of approximately 490 compared with UNSEL and approximately 770 compared with ROTH. The resistance ratio for fipronil, spinosad, indoxacarb, Cry1Ac, and Cry1Ca in the UNSEL population declined significantly by G8. Logit regression analysis of F1 reciprocal crosses between fip-SEL (at G8) and UNSEL indicated that resistance to fipronil in the fip-SEL population was inherited as an autosomal, incompletely recessive (D(LC) = 0.37) trait. At the highest dose of fipronil tested, resistance was completely recessive, whereas at the lowest dose it was incompletely recessive. A direct test of monogenic inheritance based on a backcross of F1 progeny with fip-SEL suggested that resistance to fipronil was controlled by a single locus. The fip-SEL population at G8 showed little change in its response to spinosad and indoxacarb compared with G1, whereas its susceptibility to Cry1Ac and Cry1Ca increased markedly over the selection period. This suggests that there may be some low level of cross-resistance between fipronil, spinosad, and indoxacarb. PMID- 15666764 TI - Dynamics of resistance to the neonicotinoids acetamiprid and thiamethoxam in Bemisia tabaci (Homoptera: Aleyrodidae). AB - The dynamics of resistance in the sweetpotato whitefly, Bemisia tabaci (Gennadius), to the neonicotinoids acetamiprid and thiamethoxam was studied extensively in cotton fields in Israel during the cotton-growing seasons 1999 2003. Whitefly strains were collected in early and late seasons mainly in three locations in northern, central, and southern Israel. The whiteflies were assayed under laboratory conditions for susceptibility to neonicotinoids, as part of the Israeli cotton insecticide resistance management strategy. Selections to both acetamiprid and thiamethoxam and cross-resistance between them also were conducted in the laboratory. Although no appreciable resistance to acetamiprid was observed up to 2001, a slight increase of approximately five-fold resistance was detected during 2002 and 2003. However, from 2001 to 2003 thiamethoxam resistance increased >100-fold in the Ayalon Valley and Carmel Coast cotton fields. In cross-resistance assays with both neonicotinoids, the strain that had been selected with thiamethoxam for 12 generations demonstrated almost no cross resistance to acetamiprid, whereas the acetamiprid-selected strain exhibited high cross-resistance of >500-fold to thiamethoxam. PMID- 15666765 TI - Effects of synergists on toxicity of six insecticides in parasitoid Diaeretiella rapae (Hymenoptera: Aphidiidae). AB - The resistance to and the effects of synergists on the toxicity of six insecticides in Diaeretiella rapae (M'Intosh) (Hymenoptera: Aphidiidae), a parasitoid of vegetable aphid collected in Jianxin at Fuzhou-City, Fujian, China, were studied. In comparison with susceptible F21 progeny, the resistance ratios in resistant F0 parents were 27.6 for methamidophos, 20.8 for fipronil, 47.5 for avermectin, 3.3 for fenvalerate, 4.5 for cypermethrin, and 74.7 for imidacloprid. Piperonyl butoxide (PB), triphenyl phosphate (TPP), and diethyl maleate (DEM) were chosen to be applied in susceptible F21 progeny, as well as in resistant F11 progeny and F0 parents. Significant synergistic effects on the toxicity of the six insecticides were found by using PB, TPP, and DEM in F0 parents; on methamidophos, avermectin, and imidacloprid by PB, TPP, and DEM in F11 progeny; on fipronil by PB and DEM in F11 progeny; and on fenvalerate and cypermethrin by PB in F11 progeny. PB also showed significant synergism on the six insecticides in susceptible F21 progeny, although the synergism was far less in F21 progeny than those in resistant F0 parents. TPP and DEM showed little or no synergistic effects on the toxicity of the six insecticides in F21 progeny. Compared with TPP and DEM, the highest synergistic ratios of PB for methamidophos, fipronil, avermectin, fenvalerate, cypermethrin, and imidacloprid were observed in F0 parents, and F11 and F21 progeny. The resistance levels to methamidophos, fipronil, avermectin, fenvalerate, and cypermethrin could be inhibited strongly by applying PB in F0 parents. From the results, oxidative degradation is believed to play a critical role in resistance to methamidophos, fipronil, avermectin, fenvalerate, and cypermethrin in D. rapae. To a lesser extent, hydrolytic reactions also were partially involved in the resistance to these five insecticides by using the synergists PB, TPP, and DEM. However, although high synergism of PB, TPP, and DEM on imidacloprid was found, the resistance levels to imidacloprid remained high in the presence of PB, TPP, and DEM. The mediated detoxification of oxidative degradation and hydrolytic reactions was thought to be involved in the resistance to imidacloprid in F0 parents. PMID- 15666766 TI - Behavioral and physiological resistance of the German cockroach to gel baits (Blattodea: Blattellidae). AB - A gel bait-resistant German cockroach, Blattella germanica (L.), strain Cincy was collected in Cincinnati, OH. This strain exhibited a high level of behavioral resistance to Avert (0.05% abamectin) and Maxforce FC (0.01% fipronil) gel baits. Topical application assays indicated moderate levels of physiological resistance of the Cincy strain to abamectin and fipronil. Resistance ratios (based on LD50 values from topical applications) to abamectin and fipronil were 2.5 and 8.7, respectively. The Cincy strain of had a significantly lower LD50 value to abamectin than a nonaverse field strain (Dorie) and similar LD50 values to fipronil as the Dorie strain. The aversion behavior (avoidance of gel baits) was therefore caused by food ingredients in the gel baits. The Cincy strain showed avoidance of agar containing fructose, glucose, maltose, and sucrose, which are phagostimulants to the laboratory strain. Modifications of the inert ingredients in the Maxforce FC gel bait significantly improved the efficacy against the Cincy strain. The Cincy strain produced significantly smaller oothecae and lower numbers of eggs in each egg capsule than the nonaverse Jwax and Dorie strains of cockroaches, suggesting fitness costs are associated with resistance. PMID- 15666767 TI - Inheritance of resistance to Bacillus thuringiensis Cry1Ac toxin in a greenhouse derived strain of cabbage looper (Lepidoptera: Noctuidae). AB - A population of cabbage looper, Trichoplusia ni (Hubner), collected from commercial greenhouses in the lower mainland of British Columbia, Canada, in 2001 showed a resistance level of 24-fold to Dipel, a product of Bacillus thuringiensis (Bt) subspecies kurstaki. This population was selected with Cry1Ac, the major Bt Cry toxin in Dipel, to obtain a homogenous population resistant to Cry1Ac. The resulting strain of T. ni, named GLEN-Cry1Ac, was highly resistant to Cry1Ac with a resistance ratio of approximately 1000-fold. The larvae from the GLEN-Cry1Ac strain could survive on Cry1Ac-expressing transgenic broccoli plants that were highly insecticidal to T. ni and diamondback moth, Plutella xylostella (L.). The inheritance of Cry1Ac resistance in this T. ni strain was autosomal and incompletely recessive. The degree of dominance of the resistance was -0.402 and 0.395, respectively, for the neonates in reciprocal crosses between the GLEN Cry1Ac and a laboratory strain of T. ni. Using chi2 goodness-of-fit test, we demonstrated that the inhibition of larval growth resulting from testing 12 toxin doses in the progeny of the backcross fit the predicted larval responses based on a monogenic inheritance model. Therefore, we conclude that the inheritance of the resistance to Cry1Ac in the T. ni larvae is monogenic. PMID- 15666768 TI - Rapid method to screen resistance of potato plants against Myzus persicae (Homoptera: Aphididae) in the laboratory. AB - With the objective to develop a potato, Solanum tuberosum L., resistance program against aphids, we propose a rapid screening method with Myzus persicae (Sulzer) in the laboratory. We aimed to optimize the duration of the whole procedure and to decrease the frequency of measurements. In a first experiment, intrinsic rate of natural increase (r(m)) values were compared between adult aphids reared throughout their entire life and adults reared only during a period equivalent to their prereproductive period. No significant differences were observed. In a second experiment, four groups of aphids were distinguished according to the sampling frequency, i.e., those whose biological parameters were evaluated every single, second, third, and fourth day. Except for the fourth-day experiment, the r(m) values estimated on aphids reared on the three potato lines were not significantly different whatever sampling frequency of single, second, or third day used to check aphids. Thus, screening efforts in laboratory can be largely optimized by evaluating adult aphids only during a period equivalent to their prereproductive period and assessing M. persicae populations every third day. Our method is reliable and adapted to screen a large number of potato plants against M. persicae because it allows an average 70% reduction in the time required for the whole experimental process. PMID- 15666769 TI - Impact of Bemisia argentifolii (Homoptera: Auchenorrhyncha: Aleyrodidae) infestation and squash silverleaf disorder on zucchini yield and quality. AB - Fruit yield and quality of zucchini, Cucurbita pepo L., plants infested with Bemisia argentifolii Bellows & Perring were evaluated in a screenhouse under spring and fall growing conditions by using closely related sister lines that were either susceptible (ZUC61) or tolerant (ZUC76-SLR) to squash silverleaf disorder. Our objective was to test separately the effects of level of whitefly infestation and expression of silverleaf symptoms on zucchini yield and quality. In a second experiment, yield and quality of fruit produced by silverleaf tolerant zucchini genotypes incorporating two different sources of tolerance (ZUC76-SLR and ZUC33-SLR/PMR) were compared with that of 'Zucchini Elite', a silverleaf-susceptible commercial hybrid. Zucchini fruit yield was reduced in plants exposed to repeated infestations of whiteflies in spring and fall of both experiments. In addition, fruit grew to harvestable size more slowly under the highest whitefly infestations. Fruit quality was reduced at high infestations because of uneven and reduced pigmentation. The fruit yield and quality of ZUC61 and ZUC76-SLR were similarly affected by whitefly infestation despite differences in their susceptibility to squash silverleaf disorder. Fruit from infested plants showed decreased levels of chlorophyll and carotenoids causing the "blanching" of the fruit that is associated with loss of quality and reduced marketability. Leaves of infested plants of all genotypes had reduced levels of photosynthetic and photoprotectant pigments, possibly leading to reduced photosynthesis and consequently reduced yield. We conclude that feeding by high whitefly populations rather than expression of squash silverleaf disorder is responsible for yield and quality reduction in zucchini. PMID- 15666770 TI - Soyacystatin N inhibits proteolysis of wheat alpha-amylase inhibitor and potentiates toxicity against cowpea weevil. AB - Genetic engineering may be used to introduce multiple insect resistance genes with different modes of action into crop plants. We explored the possible interactions of two differing gene products fed in the diet of cowpea weevil, Callosobruchus maculates (F.), a stored grain pest. The soybean cysteine protease inhibitor soyacystatin N (scN) and alpha-amylase inhibitor (alphaAI) from wheat have defensive function against this coleopteran. When artificial seeds containing both scN and alpha(AI) were infested with eggs of C. maculatus, the delays in larval development were longer than was predicted by summing the developmental delays seen when larvae were fed a diet containing the individual proteins, indicating that the effects of scN and alpha(AI) are synergistic. Alpha(AI) was readily hydrolyzed when incubated with insect gut extract. This proteolytic degradation was inhibited by scN, but not by Kunitz inhibitor (a serine protease inhibitor). Thus, degradation of alpha(AI) was due to proteolysis by insect digestive cysteine proteases. These data suggest that C. maculatus uses digestive enzymes not only to function in food protein digestion but also to defend the insects themselves by helping reduce the concentration of a toxic dietary protein. PMID- 15666771 TI - Buffalograss germplasm resistance to Blissus occiduus (Hemiptera: Lygaeidae). AB - Plant germplasm collections may offer genetic variability useful in identifying insect resistance. The goal of this project was to evaluate buffalograss genotypes [Buchloe dactyloides (Nutt.) Engelm.] for resistance to the chinch bug, Blissus occiduus Barber (Hemiptera: Lygaeidae), and to relate resistance to ploidy level, chinch bug number, and pubescence. Forty-eight buffalograss genotypes from diverse geographic locations were evaluated in replicated studies under greenhouse conditions. Of the genotypes studied, four were highly resistant, 22 were moderately resistant, 19 were moderately susceptible, and three were highly susceptible to chinch bug damage. The mean number of chinch bugs was significantly different among the 48 genotypes. There was no significant correlation between chinch bug resistance and ploidy level or chinch bug resistance and pubescence. These results indicate the genetic source of resistance to chinch bugs exists in buffalograss germplasm. Highly resistant genotypes can be used in breeding programs to further improve buffalograss cultivars. PMID- 15666772 TI - Management of feeding damage and survival of southwestern corn borer and sugarcane borer (Lepidoptera: Crambidae) with Bacillus thuringiensis transgenic field corn. AB - The efficacy of transgenic corn hybrids expressing an insecticidal Bacillus thuringiensis (Bt) delta-endotoxin from different transformation events was evaluated in field corn, Zea mays L., against the southwestern corn borer, Diatraea grandiosella Dyar, and sugarcane borer, Diatraea saccharalis (F.). Susceptibilities of neonates and third instars were determined on Bt and non-Bt corn plants (V6 and R1 stages) in field plots and corn leaf tissue feeding exposure in laboratory bioassays. Bt corn hybrids associated with MON810 and CBH351 transformation events sustained significantly less injury by southwestern corn borer and sugarcane borer during mid-whorl stage infestations compared with their respective non-Bt hybrid equivalents. Southwestern corn borer and sugarcane borer feeding injury to ear leaf-sheath and husk tissues during the silking stage of corn was significantly reduced in MON810 and CBH351 Bt corn compared with their respective non-Bt hybrids. However, resistance levels to feeding injury in Bt hybrids associated with the MON810 event were significantly higher than that in the hybrid associated with the CBH351 event. Southwestern corn borer and sugarcane borer caused more feeding injury to husk tissue than to ear leaf-sheath tissue in both Bt and non-Bt hybrids infested during the silking stage. Laboratory performance of the MON810 event against southwestern corn borer and sugarcane borer varied among hybrids associated with the same event. Third instars of southwestern corn borer were highly susceptible to MON810 Bt corn hybrids in leaf tissue experiments. However, sugarcane borer larvae were susceptible to the MON810 event only in one of the Bt hybrids evaluated. Sugarcane borer mortality was significantly lower after 96 h of feeding exposure on CBH351 Bt corn leaf tissue than on MON810 Bt corn leaf tissue. Plant resistance to southwestern corn borer and sugarcane borer increased as plants matured, independent of the presence of a Bt construct. These results are essential to estimate the importance of Bt transgenic corn in areas of southern United States and other areas where mixed populations of southwestern corn borer and sugarcane borer are predominant and cause severe damage to corn production. PMID- 15666773 TI - Lost P1 allele in sh2 sweet corn: quantitative effects of p1 and a1 genes on concentrations of maysin, apimaysin, methoxymaysin, and chlorogenic acid in maize silk. AB - In the United States, insecticide is used extensively in the production of sweet corn due to consumer demand for zero damage to ears and to a sweet corn genetic base with little or no resistance to ear-feeding insects. Growers in the southern United States depend on scheduled pesticide applications to control ear-feeding insects. In a study of quantitative genetic control over silk maysin, AM-maysin (apimaysin and methoxymaysin), and chlorogenic acid contents in an F2 population derived from GE37 (dent corn, P1A1) and 565 (sh2 sweet corn, p1a1), we demonstrate that the P1 allele from field corn, which was selected against in the development of sweet corn, has a strong epistatic interaction with the a1 allele in sh2 sweet corn. We detected that the p1 gene has significant effects (P < 0.0001) not only on silk maysin concentrations but also on AM-maysin, and chlorogenic acid concentrations. The a1 gene also has significant (P < 0.0005) effects on these silk antibiotic chemicals. Successful selection from the fourth and fifth selfed backcrosses for high-maysin individuals of sweet corn homozygous for the recessive a1 allele (tightly linked to sh2) and the dominant P1 allele has been demonstrated. These selected lines have much higher (2 to 3 times) concentrations of silk maysin and other chemicals (AM-maysin and chlorogenic acid) than the donor parent GE37 and could enhance sweet corn resistance to corn earworm and reduce the number of applications of insecticide required to produce sweet corn. PMID- 15666774 TI - Enumerative and binomial sequential sampling plans for soybean aphid (Homoptera: Aphididae) in soybean. AB - Since the discovery of the soybean aphid, Aphis glycines Matsumura, in midwestern U.S. soybean, Glycine max L., in 2000, the aphid has become a significant economic pest. Basic information about estimating population density within fields is unknown. Therefore, we developed two sampling plans to efficiently characterize A. glycines densities. Enumerative and binomial sequential plans were developed using 89 data sets collected from 10 commercial fields sampled during 2001-2003. Re-sampling software was used to validate the enumerative plan on whole plant counts, based on Taylor's power law parameters (a = 9.157 and b = 1.543). For research applications, the enumerative plan was modified to provide an actual precision level of 0.10 (SE/mean), which resulted in an average sample number of 310 individual plants. For integrated pest management (IPM) purposes, we developed an enumerative plan with an actual precision of 0.25, which resulted in an average sample number of 38 individual plants. For IPM applications, the binomial plan will likely be more practical. Binomial plans were developed using two tally thresholds at five action thresholds. Final analysis of the operating characteristic curve for each plan indicated that the tally threshold of > or = 40 aphids per plant, and an action threshold of 0.837 (84% of the plants infested) provided the most correct treat (4%) and no-treat (95%) decisions, with very low incorrect treat (0.5%) and no-treat (0.5%) decisions. A tally threshold of > or = 40 aphids per plant and action thresholds of 84% of plants infested is equivalent to a mean density of 250 aphids per plant, a recently recommended economic threshold. Using this threshold, the minimum required sample number for the binomial plan was 11 plants. PMID- 15666776 TI - Mites and fungi in heavily infested stores in the Czech Republic. AB - Toxigenic and allergen-producing fungi represent a serious hazard to human food and animal feed safety. Ninety-four fungal species were isolated from mite infested samples of seeds taken from Czech seed stores. Fungi were isolated from the surface of four kinds of seeds (wheat, poppy, lettuce, and mustard) and from the gut and external surface of five species of mites (i.e., Acarus siro L., 1758, Caloglyphus rhizoglyphoides (Zachvatkin, 1973), Lepidoglyphus destructor (Schrank, 1781), Tyrophagus putrescentnae (Schrank, 1781) and Cheyletus malaccensis Oudemans 1903) separately. Multivariate analysis of fungi complex composition showed that the frequency of fungal was species significantly influenced by the kind of seed. Fungal frequencies differed between mites gut and exoskeleton surface and between the surfaces of mites and seeds. Three groups of fungal species were recognized: 1) mite surface-associated fungi: Penicillium brevicompactum, Alternaria alternata, and Aspergillus versicolor; 2) mite surface and seed-associated fungi: Aspergillus niger, Penicillium crustosum, Penicillium aurantiogriseum, Penicillium chrysogenum, and Aspergillus flavus; and 3) seed associated fungi: Cladosporium herbarum, Mucor dimorphosporus f. dimorphosporus, Botrytis cinerea, Penicillium griseofulvum, and Eurotium repens. Mite-carried species of microfungi are known to produce serious mycotoxins (e.g., aflatoxin B1, cyclopiazonic acid, sterigmatocystin, ochratoxin A, and nephrotoxic glycopeptides) as well as allergen producers (e.g., A. alternata and P. brevicompactum). Storage mites may play an important role in the spread of some medically hazardous micromycetes. In addition, these mite-fungi associations may heighten the risk of occurrence of mycotoxins in food and feed stuffs and cause mixed contamination by fungal and mite allergens. PMID- 15666775 TI - Capture of Mediterranean fruit flies (Diptera: Tephritidae) in dry traps baited with a food-based attractant and Jackson traps baited with trimedlure during sterile male release in Guatemala. AB - Captures of the Mediterranean fruit fly, Ceratitis capitata (Wiedemann), in Jackson traps baited with trimedlure were compared with captures in cylindrical open-bottom dry traps baited with a food-based synthetic attractant (ammonium acetate, putrescine, and trimethylamine). Tests were conducted in Guatemala during a sterile male release program in an area where wild flies were present in low numbers. More wild and sterile females were captured in food-based traps, and more wild and sterile males were captured in trimedlure traps. The food-based traps captured almost twice as many total (male plus female) wild flies as the trimedlure traps, but the difference was not significant. Females made up approximately 60% of the wild flies caught in the food-based attractant traps; the trimedlure traps caught no females. The ratio of capture of males in trimedlure traps to food-based traps was 6.5:1 for sterile and 1.7:1 for wild flies. Because fewer sterile males are captured in the food-based traps, there is a reduction in the labor-intensive process of examining flies for sterility. The results indicate that traps baited with food-based attractants could be used in place of the Jackson/trimedlure traps for C. capitata sterile release programs because they can monitor distributions of sterile releases and detect wild fly populations effectively; both critical components of fruit fly eradication programs by using the sterile insect technique. PMID- 15666777 TI - Susceptibility of laboratory and field strains of four stored-product insect species to spinosad. AB - Two field strains of the Indianmeal moth, Plodia interpunctella (Hubner); red flour beetle, Tribolium castaneum (Herbst); and lesser grain borer, Rhyzopertha dominica (F.), and one field strain of the rusty grain beetle, Cryptolestes ferrugineus (Stephens), were collected from hard red winter wheat stored on farms in northeastern Kansas. Fifty eggs of P. interpunctella and 25 beetle adults of each species were exposed to 100 g of untreated wheat or wheat treated with various rates of spinosad, to determine susceptibility of the field and corresponding insecticide-susceptible laboratory strains. Mortality of beetle adults and P. interpunctella larvae was assessed after 7 and 21 d postinfestation, respectively. Field strains of P. interpunctella, C. ferrugineus, and T. castaneum were less susceptible to spinosad than the corresponding laboratory strains. The LD50 and LD95 values for P. interpunctella and C. ferrugineus field strains were 1.7-2.5 times greater than values for corresponding laboratory strains. Adults of both laboratory and field strains of T. castaneum were tolerant to spinosad, resulting in <88% mortality at 8 mg/kg. The LD50 and LD95 values for the field strains of T. castaneum were 2.0-7.5 times greater compared with similar values for the laboratory strain. The field and laboratory strains of R. dominica were highly susceptible to spinosad, and one of the field strains was relatively less susceptible to spinosad than the laboratory strain. Our results confirm a range of biological variability in field populations, which is consistent with findings for other compounds, and underscores the need to adopt resistance management programs with stored grain insect pests. The baseline data generated on the susceptibility of the four insect species to spinosad will be useful for monitoring resistance development and for setting field rates. PMID- 15666778 TI - Insecticidal effect of three diatomaceous earth formulations against adults of Sitophilus oryzae (Coleoptera: Curculionidae) and Tribolium confusum (Coleoptera: Tenebrionidae) on oat, rye, and triticale. AB - Bioassays were conducted in the laboratory to assess the effect of the diatomaceous earth (DE) formulations Insecto, SilicoSec, and PyriSec, on stored oat, rye, and triticale, against adults of the rice weevil, Sitophilus oryzae (L.), and the confused flour beetle, Tribolium confusum Jacquelin du Val. The DEs were tested at three dose rates, 0.75, 1, and 1.5 g of DE/kg of grain. Adults of the two aforementioned species were exposed to all combinations of grain formulation-dose rate, at 26 degrees C and 60% RH. Mortality in DE-treated commodities was recorded after 24 h, 48 h, 7 d, and 14 d of exposure for S. oryzae and T. confusum and after 21 d for T. confusum. In S. oryzae, adult mortality was almost 100% after 7 d of exposure in all three grains examined. The mortality of T. confusum adults in DE-treated grains did not reach 100%, even after 21 d of exposure. Generally, the application of DE in rye caused higher adult mortality of T. confusum than in the other two products. All three dose rates tested provided the same mortality level of S. oryzae adults after 7 d of exposure. In contrast, 1.5 g of DE resulted in significant higher adult mortality of T. confusum, in comparison with the other dose rates, even after 21 d of exposure. All formulations were equally effective after 7 d of exposure against S. oryzae, but at 48 h of exposure, PyriSec caused significantly higher mortality than the other two formulations. For both species, progeny production in the treated grains was significantly reduced in comparison with the untreated grains, whereas significant differences were noted among commodities, formulations, and dose rates. No progeny were recorded in the treated rye for either species or in the treated triticale for S. oryzae. PMID- 15666779 TI - Relative susceptibility of Tribolium confusum life stages exposed to elevated temperatures. AB - Methyl bromide, a space fumigant used in food-processing facilities, may be phased out in the United States by 2005. The use of elevated temperatures or heat treatment is gaining popularity as a methyl bromide alternative. During heat treatment, the temperature of the whole food-processing facility, or a portion of it, is raised and held between 50 and 60 degrees C for 24-36 h to kill stored product insects. We determined time-mortality responses of the confused flour beetle, Tribolium confusum (Jacquelin du Val), eggs, young larvae, old larvae, pupae, and adults exposed to six constant temperatures between 46 and 60 degrees C. Responses of all five insect stages also were measured using exposure times of 160, 40, and 12 min at 46, 50, and 60 degrees C, respectively. Time-mortality responses of all T. confusum life stages increased with an increase in exposure time and temperature. Both time-mortality and fixed time responses showed eggs and young larvae to be most susceptible at elevated temperatures and old larvae to be least susceptible. Our results suggest that old larvae should be used as test insects to gauge heat treatment effectiveness, because heat treatment aimed at controlling old larvae should be able to control all other T. confusum life stages. Besides providing baseline data for successful use of heat treatments, time-mortality data collected at the six temperatures can be used for developing thermal death kinetic models for this species to predict mortality during actual facility heat treatments. PMID- 15666780 TI - Molecular mechanisms of cell cycle control in the mouse Y1 adrenal cell line. AB - Y1 adrenocortical tumor cells possess amplified and overexpressed c-Ki-ras proto oncogene, displaying chronic high levels of the c-Ki-Ras-GTP protein. Despite this oncogenic lesion, we previously reported that Y1 cells retain tight regulatory mechanisms of cell cycle control typified by the mitogenic response triggered by FGF2 in G0/G1-arrested cells. ACTH, on the other hand, elicits cAMP/PKA-mediated antimitogenic mechanisms involving Akt/PKB dephosphorylation/deactivation and c-Myc protein degradation, blocking G1 phase progression stimulated by FGF2. In this paper we report that ACTH does not directly antagonize any of the early or late sequential steps comprising the mitogenic response triggered by FGF2. In effect, ACTH targets deactivation of constitutively phosphorylated-Akt, restraining the potential of c-Ki-Ras-GTP to subvert Y1 cell cycle control. Thus, we can consider ACTH a tumor suppressor rather than an antimitogenic hormone. In addition, we present initial results showing that high constitutive levels of c-Ki-Ras-GTP render Y1 cells susceptible to dye upon FGF2 treatment. This surprising FGF2 death-effect, that is independent of the well known FGF2-mitogenic activity, might involve a natural unsuspected mechanism for restraining oncogene-induced proliferation. PMID- 15666781 TI - Growth factors from mesonephros implicated in gonadal and adrenal differentiation. PMID- 15666782 TI - Validity of the 21-OH/LacZ transgenic mouse as a model for studying adrenocortical cell lineage. AB - Mosaic beta-galactosidase reporter staining patterns in the adult adrenal cortex of 21-OH/LacZ transgenic mice were compared to those observed in mouse chimeras and X-inactivation mosaics, which are known to have a lineage basis. This revealed similar patterns of blue and white radial stripes in all three experimental groups. Each blue stripe may contain one or more blue coherent clones of cells but this was taken into account by correcting the observed stripe numbers for the effects of different proportions of LacZ-positive (blue) and LacZ negative (unstained) cells between adrenals. The corrected stripe numbers were similar in all three experimental groups, which supports the hypothesis that the stripes in the adrenals of 21-OH/LacZ transgenic mice are formed in a similar way to those in chimeras and X-inactivation mosaics (i.e., they have a lineage basis). This suggests that the 21-OH/LacZ transgenic mouse is likely to be a valid model for studying steroidogenic cell lineage in the adrenal cortex, thereby providing additional support for the centripetal migration hypothesis of adrenocortical cytogenesis. PMID- 15666783 TI - Function and regulation of steroidogenic genes in development. PMID- 15666784 TI - Mesonephric Wnt signaling associate with a formation of an adreno-gonadal primordium in chick embryos. PMID- 15666785 TI - Angiotensin II inhibits cell proliferation and stimulates protein synthesis in rat glomerulosa cells. PMID- 15666786 TI - Effects of anencephaly on adrenocortical cell proliferation. PMID- 15666787 TI - Is reduced cell size the mechanism for shrinkage of the adrenal zona reticularis in aging? AB - In aging humans, corticosteroid production is preserved, or even increased, but there is an unexplained reduction in adrenal androgen secretion that likely has significant health implications. Preliminary analyses on adrenocortical morphology have revealed an age-associated reduction in the thickness of the zona reticularis (ZR), the cortical zone responsible for the majority of DHEA/DHEA sulfate production in the adult human, but no change in the overall thickness of the adrenal cortex. The ZR width could decrease in aging due to loss of ZR cells and/or to shrinkage of ZR cells. In the current study, we investigated whether there was a relation between thickness of the zona reticularis in young and old humans and the cell density in this zone. Paraffin-embedded sections of the adrenal cortex of 10 young (21-35 yr old) and 10 old (54-89 yr old) adults who had died suddenly as the result of trauma were stained with hematoxylin and eosin. These specimens were chosen from a larger cohort of samples for having a broad ZR in the young group and a narrower ZR in the older group. After determining the overall cortical thickness and the width of the ZR by use of computerized image analysis software, we counted the number of adrenocortical cells in two random high power fields of the ZR of each specimen. The ZR width of the older group (57 +/- 7 arbitrary units, Mean +/- SE) was significantly reduced compared to that of the young group (124 +/- 21), P < 0.001. On the other hand, the overall cortical width in the old group (232 +/- 17) was similar to that of the young adults (249 +/- 38). In the old group, the ZR comprised 24.7 +/- 3% of the total cortical width, whereas it was 50 +/- 2% of the cortical width in the young adrenals, P < 0.001. The cell density (cell number/60 x high power field) of the ZR of old adults (83 +/- 9) was similar to that of the young group (87 +/- 5). In summary, although the width of the ZR regresses with aging, cell size in this zone is preserved. Therefore, loss of trophic support for ZR cells would not appear to be the explanation for zonal shrinkage in aging. Rather, it is likely that aging effects may be due to increased cell loss in the ZR or else reduced rates of differentiation/migration of cells into this cortical zone. PMID- 15666788 TI - Metyrapone infusion stimulates adrenal growth without activating the cell cycle or the IGF system in the late gestation fetal sheep. AB - Recently, we have demonstrated that administration of metyrapone, to suppress cortisol synthesis and decrease negative feedback at the pituitary, results in an increase in circulating ACTH and adrenal growth in the late gestation sheep fetus. In these studies, we demonstrated a 2-fold increase in adrenocortical growth using morphometric techniques. To elucidate the potential molecular mechanisms leading to the increase in adrenal growth, we examined adrenal expression of the cell cycle regulatory proteins (cyclin D1) and cyclin-kinase inhibitory proteins (p16ink, p21Cip), and insulin-like growth factor-II (IGF-II), IGF-binding protein 2 (IGFBP-2) and IGF-I type 1 Receptor (IGF1R) from fetuses infused with metyrapone or vehicle for 15 days. There was a significant decrease in adrenal expression of cyclin D1 in metyrapone-(472.0 +/- 29.7) compared with vehicle-infused (662.7 +/- 29.2) fetuses. There was no significant difference, however, in the adrenal expression of the cyclin-kinase inhibitory proteins (p16ink or p21Cip) or in the IGF system (IGF-II, IGFBP-2 or IGF1R) mRNA between metyrapone- and vehicle-infused. In summary, in this model of metyrapone activated adrenal cortical hypertrophy, growth occurs with a suppression of the rate-limiting cell cycle protein and without activation of the IGF system. PMID- 15666789 TI - Ontogeny of cytochrome B5 and cytochrome P450 C17 in the human fetal adrenal gland during normal development. PMID- 15666790 TI - Coordinated regression of adrenocortical endocrine and endothelial compartments under adrenocorticotropin deprivation. AB - The adrenal cortex is a complex tissue comprising different cell types, including endocrine, stromal, nerve, perivascular, and endothelial cells. The pituitary hormone ACTH controls the coordinated development of the vasculature and the endocrine tissue mass. This suggests that paracrine secretions between steroidogenic cells and capillary endothelial cells participate in the control of adrenocortical homeostasis. In this paper, we present data demonstrating that ACTH deprivation-induced adrenal atrophy results from alterations of both endocrine and endothelial compartments. These data support the concept that, in addition to its protective effect on endocrine cells, ACTH controls adult adrenal cortex trophicity through an additional paracrine mechanism implying maintenance of the vasculature by VEGF. PMID- 15666791 TI - Tpit-/-NeuroD1-/- mice reveal novel aspects of corticotroph development. PMID- 15666792 TI - Telomerase is not required for experimental tumorigenesis of human and bovine adrenocortical cells. AB - Telomerase has often been thought to be essential for tumorigenesis of human cells. Adrenocortical cancers, like other cancers, typically have telomerase activity. We reinvestigated the requirement for telomerase in the conversion of normal human and bovine adrenocortical cells to cancer cells. When primary adrenocortical cells were transduced with retroviruses encoding SV40 large T antigen and Ha-RasG12V and immediately transplanted into immunodeficient mice they produced invasive and metastatic tumors. Cells had negligible telomerase activity before transplantation and after recovery from tumors. However, these tumors were not immortal and cells entered crisis, limiting further growth of the tumor as well as invasion and metastasis. Infection of these tumor cells with a retrovirus encoding hTERT restored growth in culture and restored the malignant properties of the cells in immunodeficient animals. These experiments differ from previous studies in which telomerase was found to be essential for tumorigenicity: 1) we used tissue reconstruction techniques for introduction of cells into host animals and 2) we infected primary cells with retroviruses and immediately transplanted them without drug selection. PMID- 15666794 TI - Clinical and molecular genetic studies of bilateral adrenal hyperplasias. AB - ACTH-independent, cortisol-producing hyperplasia is caused by 2 distinct disorders, primary pigmented nodular adrenocortical disease (PPNAD) and ACTH independent macronodular adrenal hyperplasia (AIMAH). We will review recent findings on the clinical and molecular aspects of PPNAD and AIMAH. Inactivating mutations of PRKAR1A on 17q22-24, which codes for the type 1A regulatory subunit of protein kinase A, have been found in a subgroup of patients with PPNAD with and without Carney complex. AIMAH is a rare condition in which cortisol secretion may be mediated by non-ACTH circulating hormones such as gastric inhibiting polypeptide (leading to food-dependent Cushing's syndrome), vasopressin, catecholamines, luteinizing hormone, serotonin, angiotensin-II or leptin. The primary etiology of AIMAH remains unclear. Recently, we studied the expression profile of AIMAH by genomic cDNA microarray analysis. Several candidate genes were identified, suggesting pathways that affect the cell cycle, adhesion and transcription as possible mediators of adrenocortical hyperplasia. PMID- 15666793 TI - A role for the NGFI-B family in adrenal zonation and adrenocortical disease. AB - The three zones of the human adrenal cortex are functionally distinct with the glomerulosa producing aldosterone, the fasciculata producing cortisol, and the reticularis producing DHEA/DHEAS. This functional zonation is largely due to the zone-specific expression of steroidogenic enzymes. Recent evidence suggests a role for the NGFI-B family of orphan nuclear receptors (particularly NURR1 and NGFI-B) in the zone-specific expression of two key steroidogenic enzymes, aldosterone synthase (CYP11B2) and 3beta-hydroxysteroid dehydrogenase (HSD3B2). Herein we discuss the evidence that suggests a role for NURR1 (NR4A2) in the expression of CYP11B2 in the glomerulosa as well as in the dysregulation of CYP11B2 gene expression as is seen in aldosterone-producing adenoma (APA), a major cause of endocrine hypertension. NURR1 appears to be important for CYP11B2 transcription and is found at higher levels in glomerulosa and in APA. Its expression in adrenal cells is also readily increased by angiotensin II treatment. HSD3B2 is a steroid-metabolizing enzyme that is essential for adrenal production of mineralocorticoids and glucocorticoids. Thus, HSD3B2 is expressed at high levels in the glomerulosa and fasciculata where these steroids are produced but at low levels in the adrenal reticularis, which produces mainly DHEA. We recently demonstrated that NGFI-B (nur77 or NR4A1) plays an important role in the regulation of HSD3B2 transcription and may play an important role in the functional zonation of the adrenal gland. Immunohistochemistry confirmed that, within adult and fetal adrenal gland, NGFI-B expression paralleled expression of HSD3B2. Transient transfections demonstrated that NGFI-B family members enhanced HSD3B2 reporter activity but had no effect on a 17alpha hydroxylase (CYP17) promoter construct. Taken together these results suggest that the NGFI-B family of transcription factors plays a role in establishing the functional zonation of the human adrenal by regulating CYP11B2 and HSD3B2 gene transcription. PMID- 15666795 TI - Mechanistic roles of inhibin as a tumor suppressor in the adrenal cortex. PMID- 15666796 TI - The melanocortin pathway: effects of voluntary exercise on the melanocortin-4 receptor knockout mice and ACTH(1-24) ligand structure activity relationships at the melanocortin-2 receptor. AB - The melanocortin pathway consists of endogenous agonists, antagonists, G-protein coupled receptors, and ancillary proteins that mediate the function of the endogenous antagonists. The melanocortin-4 receptor (MC4R) is involved in the regulation of obesity and the melanocortin-2 receptor (MC2R) is involved in the regulation of steroidogenesis. Herein, we present the effects of voluntary exercise on the MC4R knockout mice in terms of bypassing the morbid obesity and hyperphagia phenotypes associated with this genetic obesity model. Additionally, a systematic truncation study of the adrenocorticotropin hormone (ACTH 1-24) has been performed and characterized at the cloned MC2R. PMID- 15666797 TI - Arachidonic acid regulation of steroid synthesis: new partners in the signaling pathway of steroidogenic hormones. AB - Although the role of arachidonic acid (AA) in trophic hormone-stimulated steroid production in various steroidogenic cells is well documented, the mechanism responsible for AA release remains unknown. We have previously shown evidence of an alternative pathway of AA generation in steroidogenic tissues. Our results are consistent with the hypothesis that, in steroidogenic cells, AA is released by the action of a mitochondrial acyl-CoA thioesterase (MTE-I). We have shown that recombinant MTE-I hydrolyses arachidonoyl-CoA to release free AA. An acyl-CoA synthetase specific for AA, acyl-CoA synthetase 4, has also been described in steroidogenic tissues. In the present study we investigate the new concept in the regulation of intracellular levels of AA, in which trophic hormones can release AA by mechanisms different from the classical PLA2-mediated pathway. Inhibition of ACS4 and MTE-I activity by triacsin C and NDGA, respectively results in a reduction of StAR mRNA and protein abundance. When both inhibitors are added together there is a synergistic effect in the inhibition of StAR mRNA, StAR protein levels and ACTH-stimulated steroid synthesis. The inhibition of steroidogenesis produced by the NDGA and triacsin C can be overcome by the addition of exogenous AA. In summary, results shown here demonstrate a critical role of the acyl-CoA synthetase and the acyl-CoA thioesterase in the regulation of AA release, StAR induction, and steroidogenesis. This further suggests a new concept in the regulation of intracellular distribution of AA through a mechanism different from the classical PLA2-mediated pathway that involves a hormone induced acyl-CoA synthetase and a hormone-regulated acyl-CoA thioesterase. PMID- 15666798 TI - NGFI-B is important for induction of SF-1 dependent transcription in response to cAMP. PMID- 15666799 TI - Role of extracellular matrix components in adult rat adrenal gland homeostasis. PMID- 15666800 TI - Proteasome-mediated mineralocorticoid receptor degradation attenuates transcriptional response to aldosterone. AB - The ubiquitin-proteasome pathway regulates the turnover of many nuclear hormone receptors, such as the estrogen receptor. For estrogen receptor, proteasome inhibition decreases ligand-mediated transcription. We provide evidence that the mineralocorticoid receptor (MR) is degraded by the ubiquitin-proteasome pathway in a ligand-dependent manner and that proteasomal inhibition results in increased accumulation of the MR with enhancement of transcriptional response to aldosterone. Examination of the primary sequence of human and rat MR has identified two candidate PEST degradation motifs. Mutation of lysine 715 and/or 367 within this PEST element failed to prevent degradation of MR protein or transcriptional activity mediated by aldosterone, indicating that other lysine residues are targeted by proteasomal degradation of MR. These findings demonstrate a coupling between MR up-regulation and transcriptional hyperactivity. PMID- 15666801 TI - FHL2, UBC9, and PIAS1 are novel estrogen receptor alpha-interacting proteins. AB - Estrogen plays important roles in the pathophysiology of atherosclerosis and cardiovascular diseases mediated by estrogen receptor alpha (ERalpha). To elucidate the molecular mechanisms, we screened ERalpha-interacting proteins from a human heart cDNA library using a yeast two-hybrid system, and identified the four and a half of LIM-only protein 2 (FHL2). FHL2 interacted with ERalpha in the presence of 17beta-estradiol, but not of tamoxifen or raloxifene in yeast. FHL2 mainly interacted with N-terminal A/B domain of ERalpha but not C-terminal ligand binding domain. However, overexpression of full-length FHL2 did not affect ERalpha-dependent transcriptional activities of a reporter containing 3 copies of estrogen response element in COS-1 cells. Since tissue distribution of FHL2 was highly restricted to the heart, the function of FHL2 may be observed in a cell type- or promoter-specific manner. We have also detected strong interactions of ERalpha with Ubc9 and PIAS1 in yeast. Ubc9 and PIAS1, small ubiquitin-related modifier-1 (SUMO-1) conjugating enzyme and ligase, respectively, markedly interacted with ERalpha in a 17beta-estradiol-dependent manner. These proteins mainly interacted with the DNA-binding and ligand-binding domains of ERalpha. Overexpression of Ubc9 or PIAS1 potentiated ERalpha-mediated transcriptional activities in COS-1 cells in a dose-dependent manner, indicating that both Ubc9 and PIAS1 function as coactivators of ERalpha. In addition, the SUMOylation defective mutant, Ubc9 (C93S) continued to enhance ERalpha-dependent transcriptional activities. These findings suggest that coactivator abilities and SUMOylation capacities of Ubc9 and PIAS1 are separable and distinct. The present studies indicate that ERalpha exhibit tissue-specific functions utilizing multiple tissue-restricted receptor-interacting proteins. PMID- 15666802 TI - Tyrosine phosphates act on steroidogenesis through the activation of arachidonic acid release. AB - The ACTH signaling pathway includes both PKA activation as well as PKA-dependent tyrosine phosphatase activation. In addition, the action of this hormone also includes the regulation of the intracellular levels of arachidonic acid (AA) by the concerted action of two enzymes: an acylCoA-thioesterase and an acyl-CoA synthetase (ACS4). This work describes the production and characterization of a specific ACS4 antibody, which was used to analyze the effect of ACTH on ACS4 protein level in Y1 adrenocortical cells and the putative relationship between tyrosine phosphatases and ACS4. The antiserum was obtained from rabbits immunized with the recombinant ACS4. This immunogen was produced in bacteria and eluted from an acrylamide gel after SDS-PAGE separation of a partially purified bacteria lysate. When used in Western blot analysis, the antibody obtained specifically recognized only one protein of the molecular mass corresponding to ACS4, in Y1 cells and in several rat tissues. Using the antibody described here, we analyzed the effect of ACTH stimulation on ACS4 protein level. The hormone produced an increase of this acyl-CoA synthetase in Y1 adrenocortical cells. Moreover, this effect was mimicked by cAMP and partially reduced by a tyrosine phosphatase inhibitor. We propose that ACTH regulates ACS4 protein levels through a PKA dependent mechanism that could involve also PTP activity. PMID- 15666803 TI - The role of the melanocortin 3 receptor in mediating the effects of gamma-MSH peptides on the adrenal. AB - The pro-opiomelanocortin (POMC)-derived peptides, pro-gamma-MSH (16K fragment), and Lys-gamma3-MSH, have been shown to potentiate the steroidogenic action of corticotrophin (ACTH) on the adrenal cortex. Using a continuously perfused adrenal cell column system, we have tested the hypothesis that gamma-MSH peptides exert their effect through the Melanocortin 3 Receptor (MC3-R), since this is the only known receptor to have high affinity for gamma-MSH peptides and has been suggested to be expressed in the rat adrenal. To investigate this hypothesis we tested whether the MC3-R agonist MTII and antagonist SHU9119 could mimic or block the actions of pro-gamma-MSH. We found that MTII could not mimic, and SHU9119 could not block pro-gamma-MSH mediated potentiation of ACTH-induced steroidogenesis. These results suggest that the MC3-R is not involved in mediating the potentiation effect, adding further evidence to the argument that another melanocortin receptor exists. PMID- 15666804 TI - Multiple contributions from long-chain fatty acid metabolism in Y-1 and MA-10 cells. AB - Acute steroidogenesis in either Y-1 or MA-10 cells is sensitive to different effects of fatty acids compare to a chronic stimulation. A 3-h stimulation of StAR expression in both cell types was completely blocked by NDGA and AA861, each functioning as lipoxygenase inhibitors. However, the acute 15-min stimulation in Y-1 cells was inhibited by these agents by distinct mechanisms. The inhibition by NDGA was reversed by arachidonic, linoleic, and oleic acids. The inhibition by AA861 was insensitive to reversal by these acids. StAR expression was not affected by these short-term inhibitor treatments. These more rapid fatty acid reversible effects of NDGA are consistent with previously reported inhibition of mitochondrial Acyl CoA ligase. This may function in cooperation with StAR and PBR in providing fatty acid regulation of mitochondrial cholesterol transport. The acute effect of AA861 supports the involvement of an NDGA-insensitive lipoxygenase in the acute stimulation of mitochondrial cholesterol metabolism. The activity of MA-10 cells during prolonged treatments with cAMP appears to utilize each of these processes, which depend on different metabolism of fatty acids. PMID- 15666805 TI - Possible participation of outer mitochondrial membrane cytonchrome B5 in steroidogenesis in zona glomerulosa of rat adrenal cortex. AB - Outer mitochondrial membrane cytochrome b5 (OMb) originally found in rat liver is an isoform of cytochrome b5 (b5) of the endoplasmic reticulum. In contrast to accumulated data on the physiological roles of b5, functions of OMb have not been well characterized except for its involvement in regeneration of ascorbic acid [i.e., in a semidehydroascorbate reductase (SDAR) system]. By using highly specific antibodies against rat OMb, we found immunohistochemically that OMb in the rat adrenal gland was most abundant in the zona glomerulosa (zG) among the three cortical zones, and the expression level was enhanced on angiotensin II stimulation. SDAR activity was found in zG and inhibited by anti-OMb antibody. Moreover, the increase in plasma aldosterone concentration under Na+ -deficiency was suppressed by limited ascorbic acid (Asc) availability in rat mutants unable to synthesize Asc, while plasma corticosterone concentration was not affected. These data suggest that OMb, present abundantly in zG, participates in aldosterone formation in zG of rat under angiotensin II-stimulation through regeneration of Asc. PMID- 15666806 TI - POMC peptides in adrenal maintenance and function. PMID- 15666807 TI - Endocytosis of connexin protein in adrenal cells. AB - The ability of adrenocorticotropin (ACTH) to affect gap junctions was examined in adrenal cells in vivo and in vitro. Treatment with ACTH increased the size and number of gap junction plaques on the cell membranes in hypophysectomized animals and in adrenal culture. Intracellular (cytoplasmic) annular gap junctions were observed in cells of the inner adrenal cortical zones and in adrenal cell cultures. To investigate the relationship of annular gap junctions to surface junctions, adrenal cells in culture were transfected with cDNA encoding a green fluorescent protein tagged connexin 43 construct (Cx43-GFP), and subsequently studied by time-lapse video microscopy, immunocytochemistry, and transmission electron microscopy (TEM). Internalization of part or all of a surface gap junction plaque resulted in annular gap junction formation. These studies support the hypothesis that cytoplasmic vesicles, initially described with TEM methods, can result from removal of gap junction plaques from the cell surface. It is suggested that hormones can play a regulatory role in cell-cell communication by influencing the availability of gap junction protein at the cell surface and that hormonally-sensitive processes might serve as a means of altering intercellular communication. PMID- 15666808 TI - Molecular events triggered by heat shock in Y1 adrenocortical cells. AB - Several stimuli, including stress conditions, promote the activation of MAP kinases family members (ERK1/2, JNK, p38). In turn, these enzymes regulate several cellular functions. Given that MAPK activation requires the phosphorylation of these proteins, their inactivation depends on the activity of specific phosphatases. MAPK phosphatase-1 (MKP-1), a phosphatase specifically involved in the inactivation of MAPK family members, is induced by mitogenic stimuli and stress conditions. Here we describe the effect of heat shock (HS), 10 min, 45 degrees C, on MAPKs activities and MKP-1 mRNA and protein levels in Y1 adrenocortical cells. Western blot analysis performed with antibodies against the phosphorylated forms of ERK1/2 and JNK revealed that HS produced the rapid activation of these kinases. Their inactivation was also a rapid event and occurred together with the increase of MKP-1 protein levels detected by Western blot analysis. In addition, the effect of HS on MKP-1 protein levels seems to be exerted at the transcriptional level, since the amount of its mRNA in heat shocked cells was higher than in nonheated cells. Comparison of the temporal profiles of MKP-1 protein induction and MAPKs phospho-dephosphorylation suggests that MKP-1 induction could contribute to ERK1/2 and JNK inactivation after HS. PMID- 15666809 TI - ACTH modulates ERK phosphorylation in the adrenal gland in a time-dependent manner. AB - ACTH is known to act through the activation of cAMP/PKA in adrenocortical cells, but it has been suggested that it could also act via other pathways such as the ERK 1/2 cascade. To determine the effects of ACTH administration at sequential time points on the activation of ERKs 1/2, groups of rats (n = 6/group) were subjected to i.p. injections of either ACTH (Synacthen Depot-0.2 mg/Kg), or saline (Ct). The animals were sacrificed and the adrenal glands collected at different timings after ACTH injection (2 h, 18 h and 24 h). Two additional groups were injected daily until sacrifice (3 days and 15 days). Blood was collected for analysis and the adrenals were used for immunohistochemistry or Western Blot (WB) analysis. Immunoreactivity was scored by counting the mean number of zonae fasciculata (ZF) and reticularis (ZR) positive cells/section (mean +/- SEM). Adrenal weight was increased by ACTH in comparison with Ct. Corticosterone levels, as expected, were higher in ACTH treated animals than in Ct. The number of pERK positive cells increased in a time-dependent manner until 3d, and declined although not significantly in the 15 days animals (Control- 48.13 +/- 9.0; ACTH 2 h--125.93 +/- 14.5; ACTH 18 h-139.46 +/- 10.0; ACTH 24 h- 185.28 +/- 13.3; ACTH 3 days--198.47 +/- 18.6; ACTH 15 days--158.58 +/- 15.1). Comparable results were obtained with WB analysis. Our data shows that ACTH induces the activation of the MAPK/ERKs 1/2 cascade, especially in the ZF, consistent with this zone being more responsive to ACTH. PMID- 15666810 TI - Dehydroepiandrosterone sulphate (DHEAS) inhibits growth of human vascular endothelial cells. AB - Dehydroepiandrosterone sulphate (DHEAS) is a steroid product of the adrenal gland, which circulates in high concentrations, but whose functions are largely unknown. There is evidence for antiproliferative effects of DHEAS in neoplastic tissue. The present study was designed to investigate the effects of DHEAS on vascular endothelial cell proliferation. It was found that DHEAS at physiological concentrations (10 microM) caused inhibition of cellular growth, which was reversible following removal of the steroid. This effect was not mimicked by other steroids, suggesting that it is not mediated by androgen or estrogen receptors. Uptake of 3H-thymidine was not altered by DHEAS, suggesting that this steroid may induce apoptosis in vascular endothelial cells. PMID- 15666811 TI - In search of the function of the peripheral-type benzodiazepine receptor. AB - The peripheral-type benzodiazepine receptor (PBR) is a mitochondrial protein, involved in the regulation of cholesterol transport from the outer to the inner mitochondrial membrane, the rate-determining step in steroid hormone biosynthesis. Molecular modeling of PBR suggested that it might function as a channel for cholesterol. Indeed, cholesterol uptake and transport by bacteria cells was induced upon PBR expression. Amino acid deletion, site-directed mutagenesis, and structural studies identified a cholesterol recognition/interaction amino acid consensus sequence in the cytoplasmic carboxy terminus of the receptor. In vitro reconstitution experiments demonstrated that the 18 kDa PBR protein binds with high affinity both drug ligands and cholesterol. In situ and in vitro studies indicated that in steroidogenic cells the StAR-induced cholesterol import into mitochondria was mediated by the outer mitochondrial membrane PBR. In search of the tissue specificity of PBR expression it was shown that the high levels of PBR expression in steroidogenic cells are due, at least in part, to the expression of Sp1/Sp3 transcription factors. Moreover, PBR's function in cholesterol transport was found to be conserved across kingdoms because a PBR-homologous Arabidopsis sequence when expressed in bacteria protoplasts caused a ligand-induced uptake of cholesterol suggesting that the Arabidopsis PBR homologue is involved in steroid import in plant mitochondria. In conclusion, these studies suggest that PBR's ability to bind and transport cholesterol is a well-conserved function of this ubiquitous protein. Expression of specific transcription factors results in the overexpression of PBR and increased cholesterol transport into mitochondria associated with a specialized function (steroidogenesis). In other tissues, PBR expression might be part of the mitochondrial membrane biogenesis process involved in increased cell proliferation (cancer, gliosis) and tissue repair (nerve damage and ischemia reperfusion injury). PMID- 15666812 TI - Janus kinase 2 signaling in the angiotensin II-dependent activation of StAR expression. AB - Angiotensin II (Ang II)-stimulated aldosterone production in adrenocortical glomerulosa cells requires de novo expression of the steroidogenic acute regulatory protein (StAR). We previously reported that StAR mRNA levels and promoter-reporter gene activity in transiently transfected H295R human adrenocortical cells were stimulated by Ang II and the goals for the current study were to identify signaling pathways activated by Ang II that contribute to StAR transcriptional activation. Using StAR promoter-reporter gene activity and pharmacological inhibition of signaling pathways, we have shown that Ang II stimulated StAR transcription in H295R cells is dependent upon both influx of external Ca2+ and tyrosine kinase signaling and is enhanced by protein kinase C and mitogen-activated protein kinase (ERK1/2) activation. In particular, Janus tyrosine kinase-2 (Jak2) activation was increased with Ang-II treatment of H295R cells and the select Jak2 inhibitor, AG490, blocked Ang II-dependent Jak2 activation, StAR reporter gene activity, and steroid production. The Ang II dependent, but not (Bu)2cAMP-dependent, induction of StAR mRNA was also blocked by AG490 and shown to be sensitive to cycloheximide treatment. Together our data support Jak2 as a novel pathway in the Ang II-dependent activation of StAR expression and steroidogenesis in adrenocortical cells and indicate a requirement for ongoing protein synthesis in Ang II-mediated StAR transcription. PMID- 15666813 TI - Role of StAR binding protein in steroidogenic cells. PMID- 15666814 TI - SR-BI and HDL cholesteryl ester metabolism. AB - Scavenger receptor class B, type I (SR-BI) is the receptor for high density lipoprotein (HDL) that mediates cellular uptake of HDL cholesteryl ester (CE) and is a major route for cholesterol delivery to steroidogenic pathways. SR-BI is localized in specialized microvillar channel plasma membrane compartments that retain HDL and are sites for HDL CE selective uptake. In fact, adrenal gland microvillar channel formation is regulated by adrenocorticotropin hormone and requires SR-BI expression. SR-BI-mediated uptake of HDL CE is a two-step process requiring high affinity HDL binding followed by transfer of CE to the membrane. SR-BI delivers HDL CE to sites in the membrane where it is readily metabolized to free cholesterol by cell type-specific neutral CE hydrolases. The most likely candidate for the hydrolysis of HDL CE delivered via SR-BI in the adrenal gland is hormone sensitive lipase. New data in adrenocortical cells as well as the study of a mutant SR-BI receptor lend insight into the mechanism of cholesterol transfer from plasma HDL to the steroidogenic pathway in endocrine cells. PMID- 15666815 TI - Promiscuous 3beta-hydroxysteroid dehydrogenases: testosterone 17beta hydroxysteroid dehydrogenase activities of mouse type I and VI 3beta hydroxysteroid dehydrogenases. AB - 3Beta-hydroxysteroid dehydrogenase (3beta-HSD) activity is essential for the synthesis of all classes of steroid hormones, converting various delta5-3beta hydroxysteroids into hormonally active delta4-3-ketosteroids in NAD+ -dependent reactions. Certain 3beta-HSD isoforms have been reported to exhibit additional dehydrogenase character (e.g., 17-hydroxysteroid dehydrogenase/reductase). We have investigated whether mouse type I (adrenal/gonadal) and type VI 3beta-HSDs (uterine/embryonic) display significant 17beta-HSD-like activity. Nonsteroidogenic HEK 293T cells were transiently transfected with pCMV-based expression vectors containing mouse type I and type VI 3beta-HSDs. Transfected cells expressing either mouse type I or type VI 3beta-HSD converted testosterone to androstenedione, albeit at rates one-tenth of those of pregnenolone to progesterone in similarly transfected 293T cells. Our findings demonstrate that the mouse 3beta-HSD I and VI isoforms can inactivate testosterone within an intact cell milieu. These findings are important not only in establishment of structure-function relationships, but also whenever murine systems are used for developmental/reproductive paradigms associated with human disorders. PMID- 15666816 TI - Enzymatic phenotyping and evolutionary relatedness of species specific cytochromes P450 17alpha. PMID- 15666817 TI - Cellular specific expression of the androgen-conjugating enzymes UGT2B15 and UGT2B17 in the human prostate epithelium. AB - In humans, 3beta-hydroxysteroid dehydrogenase (3beta-HSD), 17beta-HSD, and 5alpha reductase enzymes convert dehydroepiandrosterone (DHEA), androstenedione, and testosterone into the most potent natural androgen dihydrotestosterone (DHT) in the prostate. This androgen is transformed mainly in situ to two Phase I metabolites, androsterone (ADT) and androstane-3alpha,17beta-diol (3alpha-DIOL), which can, however, be back-converted to DHT. Here, we report recent findings on the characterization of specific anti-UDP-glucuronosyltransferases (UGT) 2B15 and 2B17 antibodies and their use to identify UGT2B expressing-cells in the human prostate epithelium. We found that UGT2B17 is expressed in basal cells where DHEA is converted into 3alpha-DIOL and ADT. By contrast, the expression of UGT2B 15 was observed only in luminal cells, where DHT is formed from testosterone. These results demonstrate that, in the human prostate, UGT2B15 and UGT2B17 genes have complementary roles, and are expressed in cells where their specific substrates are synthesized. This reinforces the hypothesis that UGT enzymes catalyze an important mechanism for modulating the action of steroids and protecting the steroid-sensitive tissues from deleteriously high steroid concentrations. PMID- 15666818 TI - Niemann pick type C disease as a model for defects in neurosteroidogenesis. AB - Many functions have been attributed to neurosteroids including actions as anxiolytics, roles in myelination, inhibitors of neuronal toxicity and ischemia, and roles in neuronal growth and differentiation. To understand the functions of neurosteroids during nervous system development, we used two mouse models: one, in which the cyp17 gene was ablated, thus ablating synthesis of the neurosteroid DHEA, and a second, in a mouse model of a human childhood fatal neurodegenerative disease, Niemann-Pick Type C (NP-C). Cyp17-/- mice died unexpectedly approximately embryonic day 7. Cyp17 was expressed in the embryonic endoderm at E7, where 17alpha hydroxylase and c17,20 lyase activities were found. Hormonal replacement was ineffective in rescuing the embryos. The function of P450c17 and/or its steroid products in early mouse development is unknown. In the second model, we used a naturally-occurring NP-C mutant mouse. Mutations in the npc1 gene results in lysosomal accumulation of cholesterol and gangliosides in humans and in the mouse, which also recapitulates the onset of neurological deficits, neuronal loss and death typical of the most severe form of the human disease. We showed that there is a substantial reduction in the synthesis of the neurosteroid allopregnanolone (ALLO) at birth, which may lead to abnormal neural development. ALLO treatment was highly effective; ALLO-treated NP-C mice had substantially increased survival and delays in neurologic impairments, coinciding with marked improvements in neuronal survival, and reduction of gangliosides. These data suggest that neurosteroids play an important role in brain development and maturation and may be an effective therapy for NP-C and perhaps other lysosomal storage diseases. PMID- 15666819 TI - Isolation of marmoset P450c17 cDNA and gene regulation in vitro. AB - Marmosets express a fetal zone during adrenal development but fail to produce significant amounts of C19 steroids in adulthood. It is not clear however if P450c17 regulation is different from that in humans/rhesus or the primary sequence is altered. To this end we isolated marmoset and rhesus adrenocortical cells, and treated the cells with known regulators of P450c17 expression for 48 h. P450c17 protein increased with Forskolin (F) treatment, but was marginally inhibited by AII (A) and TPA (T) alone. Combined A + F and T + F dramatically ablated the F response. Cortisol levels (EIA) increased upon F treatment and were inhibited by A and T. Combination of treatments partially inhibited the F-induced response. The protein-coding region of marmoset and rhesus P450c17 cDNAs were then isolated from adrenals using RT-PCR/TA cloning. Marmoset P450c17 shows one amino acid deletion but otherwise shares 90.6% and 91.4% homologies with the human and rhesus cDNA sequences, and 82.4% and 85% homologies with the human and rhesus predicted AA sequences, respectively. Since marmoset adrenocortical cells exhibit similar endocrine function to rhesus, impaired 17,20-lyase activity in the adult marmoset adrenal may in part be due to differences in the primary sequence. PMID- 15666820 TI - The effect of Sutherlandia frutescens on steroidogenesis: confirming indigenous wisdom. AB - Sutherlandia frutescens (Cancer bush), a Southern African indigenous plant, is traditionally used to treat stress related maladies linked to the endocrine system. Extracts of the shrub were used to investigate the claimed stress relieving properties of the shrub. Dysregulation of the stress response is associated with elevated glucocorticoid levels. A model of chronic intermittent immobilization stress was investigated in 40 adult male Wistar rats to determine the effect of Sutherlandia. Immobilization stress resulted in increased corticosterone levels in the control group while rats receiving Sutherlandia extract showed significantly decreased corticosterone levels (P < 0.005). Since the biosynthesis of glucocorticoids in the adrenals is catalyzed by the cytochrome P450-dependent enzymes, the influence of Sutherlandia extracts on adrenal steroidogenesis was determined in ovine adrenocortical microsomes and mitochondria, using spectral binding and enzyme conversion assays. Water extracts showed inhibition of substrate binding to cytochrome P450 21-hydroxylase (CYP21) by 38% and cytochrome P450 11beta-hydroxylase (CYP11B1) by 60%. The conversion of progesterone and pregnenolone was inhibited by 34% and 30%, respectively. Subsequent extractions with chloroform and methanol showed inhibition of substrate binding and conversion with hydrophobic compounds exhibiting a greater inhibitory effect on deoxycorticosterone binding to CYP11B1 (30%) and on progesterone binding to CYP21 (50%). The inhibition of binding of pregnenolone to CYP17 by the chloroform extract was 62%, with negligible inhibition by the methanol extract. The chloroform extract showed a greater inhibitory effect than the methanol extract on progesterone and pregnenolone metabolism (20%-50%). PMID- 15666821 TI - Expression of AT1-R in marmoset whole adrenal glands and adrenocortical cells in culture. AB - The marmoset adrenal is interesting as it is developmentally similar to humans and other primates, but in adulthood 17,20-lyase activity is very low. One possible explanation is an altered regulation of P450c17 expression by AT1-R. We investigated the expression and zonal distribution of the AT1-R in marmoset adrenal glands and adrenocortical cells in culture since it is known that AT1-R is a regulator of P450c17 expression and activity. AT1-R was expressed strongly in the ZG and to diminishing degrees through the remainder of the cortex. There was negative expression in a putative ZI. Dispersed adrenocortical cells retained AT1-R protein as detected by ICC. Cells cultured for several days and treated in serum-free media for 48 h maintained AT1-R expression, as measured by western. However, at that 48 h time point, treatment with Forsk, AII, TPA, or the combination of A+F or T+F did not appear to effect AT1-R expression. We conclude that marmoset adrenals express AT1-R similarly to humans and other higher mammals, adrenocortical cells retain AT1-R expression in culture, and consistent with other adrenal cell culture models, AT1-R expression is not lost in response to agonists long term. PMID- 15666822 TI - Lack of hepatic conversion of dehydroepiandrosterone sulfate (DHEAS) to DHEA. PMID- 15666823 TI - The influence of the amino acid substitution I98K on the catalytic activity of baboon cytochrome P450 side-chain cleavage (CYP11A1). AB - Cytochrome P450 side-chain cleavage (CYP11A1) catalyzes the first and "rate limiting" step in steroidogenesis, the conversion of cholesterol to pregnenolone. In an effort to gain further insight into the structure/function relationship of this key enzyme, CYP11A1 was characterized in the Cape baboon (Papio ursinus), a species closely related to humans. Baboon cDNA was isolated from adrenal tissue and direct sequence analysis showed mature baboon and human CYP11A1 share 98% deduced amino acid homology. The cDNA was subsequently amplified and two recombinant constructs, CYP11A1a and CYP11A1b, were cloned. Sequence analyses of the constructs revealed four amino acid substitutions. The constructs were expressed in nonsteroidogenic mammalian COS-1 cells with 25-hydroxycholesterol as substrate. Apparent Km values of 1.62 and 4.53 microM were determined for CYP11A1a and CYP11A1b, respectively. Homology modeling revealed that the lower substrate affinity of CYP11B1b could be attributed to an I98K substitution, which lies between the B and C helices, providing further evidence for the importance of this domain in the catalytic activity of CYP11A1. PMID- 15666824 TI - Effective StAR expression in mouse MA-10 and Y-1 cells: extensive overexpression does not increase basal or cAMP-induced steroidogenesis. PMID- 15666825 TI - Generation and characterization of human steroidogenic factor 1 LBD crystals with and without bound cofactor peptide. AB - The nuclear receptor Steroidogenic Factor 1 (SF1) plays a critical role in the development of the adrenal gland and gonads, and in sexual differentiation. SF1 performs this pivotal function through the regulation of hormone expression that is essential for organogenesis and endocrine homeostasis. SF1 is a member of a nuclear receptor subclass that contains LRH1 and the Drosophila receptor FTZ-F1. To date, a natural ligand has not been reported for any member of this subfamily. Here we report the crystallization and characterization of the ligand-binding domain (LBD) of human SF1 from two different crystal forms: a binary complex with fortuitous ligand and a ternary complex with the same ligand and a peptide containing a motif of a nuclear receptor cofactor. The structural determination of the binary complex required the use of sulfur SAD phasing, a relatively new technique that uses anomalous diffraction from the endogenous sulfur atoms present in the protein. The structure of the ternary complex was determined by multiple wavelength anomalous diffraction (MAD) using seleno-methionine substituted SF1. Preliminary analysis suggested SF1 contained a fortuitous ligand in the binding pocket. This ligand may account for the relatively high basal activity observed for SF1 in cofactor recruitment and cell-based assays. PMID- 15666826 TI - ACTH regulates steroidogenic gene expression and cortisol biosynthesis in the human adrenal cortex via sphingolipid metabolism. AB - Sphingolipids are a diverse family of phospholipids and glycolipids that mediate cell-cell interactions, participate in signal transduction pathways and modulate the activity of various cellular proteins and receptors. The objective of the present studies was to characterize the role of the sphingolipid biosynthetic pathway in adrenocorticotropin (ACTH)-dependent steroidogenic gene expression and cortisol production. H295R human adrenocortical cells were treated with ACTH or dibutyryl cAMP (Bt2cAMP) for various time periods and the content of sphingolipids was quantified by mass spectrometry. Treatment of H295R cells with ACTH and Bt2cAMP activated sphingolipid metabolism within five minutes. Decreases were found in the cellular levels of several sphingolipids, including sphingomyelin (SM) and glucosylceramide. ACTH/cAMP rapidly decreased levels of the signaling molecules ceramide, sphingosine and sphingosine-1-phosphate (S1P). The effect of these bioactive sphingolipids on steroidogenic gene expression was also examined. Both sphingosine and S1P were found to increase endogenous CYP17 mRNA and activate the transcriptional activity of CYP17-luciferase reporter constructs. Further, sphingosine and S1P rapidly increase cortisol biosynthesis in H295R cells. In summary, our studies establish a link between ACTH/cAMP dependent steroidogenesis and sphingolipid metabolism in the human adrenal cortex. Finally, these findings suggest that sphingolipids may serve as signaling mediators in ACTH-stimulated cortisol biosynthesis. PMID- 15666828 TI - Parallel early development of zebrafish interrenal glands and pronephros: differential control by wt1 and ff1b. PMID- 15666827 TI - COUP-TF and transcriptional co-regulators in adrenal steroidogenesis. AB - Chicken ovalbumin upstream promoter-transcription factors (COUP-TFs) and steroidogenic factor-1 (SF-1) play key roles in the transcriptional regulation of steroidogenic P450 genes. Transfection studies showed that SF-1 activated bovine CYP17 promoter activity, whereas COUP-TFs repressed it from the CRS2 element in a mutually exclusive manner in mouse adrenocortical Y-1 cells. COUP-TFI and SF-1 competitively bind to the Ad5 element of the human CYP11B2 gene promoter. Unexpectedly, overexpression of COUP-TFI increased the CYP11B2 promoter activity, whereas overexpression of SF-1 repressed it in human adrenocortical H295R cells. In cortisol-producing adrenal cortical adenomas, down-regulation of nuclear receptors, including COUP-TFs was found. We therefore screened for COUP-TFI interacting proteins using a yeast two-hybrid system and have identified Ubc9 and PIAS1, SUMO-1 conjugating enzyme and ligase, respectively. Coexpression of Ubc9 and PIAS1 synergistically enhanced COUP-TFI-mediated trans-repression of CYP17 gene as well as transactivation of CYP11B2 gene. The SUMOylation-defective mutants of these proteins continued to function as co-regulators of COUP-TFI. These findings indicate that Ubc9 and PIAS1 can function as transcriptional co regulators of COUP-TFI to modulate adrenal cortical steroidogenesis in a SUMOylation-independent manner. PMID- 15666829 TI - The Sp transcription factor family is involved in the cellular expression of the human GIP-R gene promoter. PMID- 15666830 TI - On the control of the hCYP11B2 gene expressing cytochrome P450 aldosterone synthase. AB - We have previously reported that the protein kinase C ligand 12-O tetradecanoyphorbol-13-acetate (TPA) inhibited the angiotensin II (AII) stimulated CYP11B2 gene expression in the adrenocortical H295R cell line. Here we report that TPA increased the level of phospho-p44/42 MAPK but AII did not. The MEK1 inhibitor PD98059 was found to increase the level of aldosterone synthase mRNA and the activity of a human CYP11B2(-2023 bp)-promoter construct. The cotransfection of H295R with ERK 1 and the hCYP11B2 promoter resulted in the inhibition of the promoter activity. TPA but not AII increased the level of the transcription factor JunB in nuclear extracts and the increase was partially abolished by the MEK1 inhibitor PD98059. The cotransfection of H295R with JunB and the hCYP11B2 promoter abolished the AII stimulating effect. Taken together these results suggest that TPA inhibits the AII-dependent activation of CYP11B2 via the p44/42 MAPK signaling pathway leading to an increase of the level of nuclear JunB. PMID- 15666831 TI - Study of the function of proximal SF-1 binding sites on Cyp11A1 promoter. PMID- 15666832 TI - Fetal programming of the neuroendocrine response to stress: links between low birth weight and the metabolic syndrome. AB - There is now substantial agreement that small size at birth is associated with increased rates of the metabolic syndrome (glucose intolerance, high blood pressure, and dyslipidaemia) and related pathologies including cardiovascular disease in adult life. Evidence is also emerging that suggests programming of hormonal systems in response to an adverse fetal environment may be one of the mechanisms underlying these long-term consequences of early life events. In particular, alterations in the neuroendocrine response to stress may play an important part. Recent research suggests that increased adrenocortical and sympathoadrenal responses are associated with small size at birth. Evidence from epidemiological studies shows that subtle alterations in these neuroendocrine systems appear to exert a powerful influence on the levels of cardiovascular risk factors including plasma glucose and lipid concentrations and blood pressure. PMID- 15666833 TI - Fetal glucocorticoid exposure and hypothalamo-pituitary-adrenal (HPA) function after birth. AB - The fetus may be exposed to increased endogenous glucocorticoid or synthetic glucocorticoid in late gestation. Indeed, 7-10% of pregnant women in Europe and North America are treated with synthetic glucocorticoid to promote lung maturation in fetuses at risk of preterm delivery. Such therapy is effective in reducing respiratory complications. However, very little is known about the mechanisms by which synthetic glucocorticoid or prenatal stress influence neurodevelopment in the human, or whether specific time windows of increased sensitivity exist. Glucocorticoids are essential for many aspects of normal brain development. However, there is growing evidence that exposure of the fetal brain to excess glucocorticoid can have lifelong effects on neuroendocrine function and behavior. We have shown that both endogenous glucocorticoid and synthetic glucocorticoid exposure has a number of rapid effects in the fetal brain in late gestation, including modification of neurotransmitter systems and transcriptional machinery. Such fetal exposure permanently alters hypothalamo-pituitary-adrenal (HPA) function in prepubertal, postpubertal, and aging offspring, in a sex dependent manner. These effects are linked to changes in central glucocorticoid feedback machinery after birth. Prenatal glucocorticoid manipulation also leads to modification of HPA-associated behaviors, brain and organ morphology, as well as altered regulation of other endocrine systems. Permanent changes in endocrine function will have a long-term impact on health, since elevated cumulative exposure to endogenous glucocorticoid is linked to the premature onset of pathologies associated with aging. PMID- 15666834 TI - Direct inhibitory effects of leptin on the neonatal adrenal and potential consequences for brain glucocorticoid feedback. AB - Leptin is most studied for its primary role in the CNS control of energy balance and food intake in humans and rodents, yet it has functions on multiple target sites including the adrenal gland. In adult rodents, leptin has been shown to inhibit adrenal steroidogenesis and we have recently demonstrated that some of the mechanisms responsible for leptin-induced inhibition of adrenal glucocorticoid production, namely a reduction of StAR protein expression are already present in the neonatal adrenal gland. The effect of leptin on the neonatal adrenal gland integrates well with the previously demonstrated effect of this protein to inhibit stress responses, enhance glucocorticoid receptor expression in the CNS and sensitivity to glucocorticoid inhibitory feedback in neonates. The leptin receptor isoform and intracellular mechanisms involved in regulation of the adrenocortical activity at multiple levels might differ between target tissues (CNS vs periphery) and age (neonates vs adult). Neonatal leptin represents an important regulator of adrenocortical function during a critical period of brain development, which is exquisitely sensitive to circulating glucocortcoid concentrations. Since circulating leptin levels in neonates vary according to maternal diet, this protein can be viewed as a critical link between environmental and maternal factors and the developing physiology of the infant. PMID- 15666835 TI - Early embryonic environment, the fetal pituitary-adrenal axis and the timing of parturition. AB - It is well established in the sheep, that the normal timing of parturition is dependent on a prepartum activation of the fetal pituitary-adrenal axis. We have recently demonstrated for the first time that embryo number, embryo sex, and alterations in the environment of the early embryo, including exposure to maternal undernutrition during the periconceptional period, alter the timing and level of activation of the pituitary-adrenal axis in the sheep fetus during late gestation. There is a delay in activation of the fetal HPA axis in twin fetuses and we speculate that the diminished adrenocortical responsiveness in the twin fetus may be an adaptive response, which counters the impact of the potential enhanced intrauterine stress experienced by a twin fetus, thereby reducing the possibility of preterm delivery. We have also reported that a moderate restriction of maternal nutrition to during the periconceptional period (from 60 days before and for one week after conception) resulted in an earlier activation of the pituitary-adrenal axis of twin, but not singleton, fetuses during late gestation. A series of studies using assisted reproductive technologies have also found that perturbation of the early embryonic environment results in a dysregulation of placental and fetal growth and development and in the timing of normal parturition. In summary, after several decades of work focussed on events in late gestation associated with the prepartum activation and stress responsiveness of the fetal HPA axis, our recent studies indicate that the environment of the early embryo may have a significant role to play in determining the timing and level of the prepartum activation of this axis and potentially on the functional capacity of the axis to respond to acute or chronic stress in later life. PMID- 15666836 TI - Tissue-specific knockouts of steroidogenic factor 1. PMID- 15666837 TI - Genetic dissection of stress response pathways in vivo. AB - A number of lines of evidence suggest that alterations in forebrain glucocorticoid receptor (GR)-mediated regulation of the hypothalamic-pituitary adrenal (HPA) axis may be involved in the etiology of depression. The level of expression of GR in the hippocampus is highly correlated with HPA axis activity, and a number of animal models of depression are associated with altered forebrain GR expression. We have generated a line of mice with a conditional, forebrain specific deletion of GR (FBGRKO) to determine if a primary deficit in forebrain GR signaling is an etiologic factor in the pathogenesis of depression. These mice should prove to be valuable for identifying GR target genes in major depressive disorder (MDD) and testing pharmacological agents for efficacy in this disorder. PMID- 15666838 TI - Fat cells may be the obesity-hypertension link: human adipogenic factors stimulate aldosterone secretion from adrenocortical cells. AB - Obesity has become an epidemic problem in Western societies contributing to several disease processes including metabolic diseases, hypertension, and cardiovascular disease. Overweight and obesity are frequently associated with increased plasma levels of aldosterone suggesting a direct link between obesity hypertension and increased mineralocorticoid levels. The adipocyte has long been suggested to be directly involved in the regulation of the body's homeostasis and recent evidence now proves that human fat is a highly active endocrine tissue. We therefore tested the hypothesis that adipocyte secretory products directly stimulate adrenocortical aldosterone secretion. Indeed, secretory products from isolated human adipocytes strongly stimulated steroidogenesis in human adrenocortical cells (NCI-H295R), as well as in bovine adrenocortical cells with a predominant effect on mineralocorticoid secretion. In conclusion, a possible direct link exists between fat tissue metabolism and adrenal mineralocorticoid secretion that may be responsible for obesity-related hypertension. PMID- 15666839 TI - Vitamin C is an important cofactor for both adrenal cortex and adrenal medulla. AB - The adrenal gland is among the organs with the highest concentration of vitamin C in the body. Interestingly, both the adrenal cortex and the medulla accumulate such high levels of ascorbate. Ascorbic acid is a cofactor required both in catecholamine biosynthesis and in adrenal steroidogenesis. Here we provide an overview on the role of vitamin C in the adrenal cortex and medulla derived from in vitro and in vivo studies. In addition, recent insights gained from an animal model with a deletion in the gene for the ascorbic acid transporter will be summarized. Mutant mice lacking the plasma membrane ascorbic acid transporter (SVCT2) have severely reduced tissue levels of ascorbic acid and die soon after birth. There is a significant decrease of tissue catecholamine levels in the adrenals. On the ultrastructural level, adrenal chromaffin cells in SVCT2 null mice show depletion of catecholamine storage vesicles, signs of apoptosis, and increased glycogen storage. Decreased plasma levels of corticosterone and altered morphology of mitochondrial membranes indicate additional effects of the deficiency on adrenal cortical function. The data derived from these animal models and various cell culture studies confirm a crucial role for vitamin C for both the adrenal cortex as well as the adrenal medulla further underlining the interdependence of the two endocrine systems united in one gland. PMID- 15666840 TI - P450 oxidoreductase deficiency: a new disorder of steroidogenesis affecting all microsomal P450 enzymes. AB - Combined partial deficiency of 17alpha-hydroxylase and 21-hydroxylase activities was first described in 1985; however the genes for P450c17 and P450c21 in these patients lack mutations. In 1986 we postulated that this disorder might be due to mutations in P450 oxidoreductase (POR), the flavoprotein that donates electron to these and all other microsomal P450 enzymes, but this hypothesis was not tested until the POR gene sequence became available through the genome database. We found five POR missense mutations in our first four patients. In vitro assays of the activities of these mutations showed that the standard assay of POR activity, reduction of cytochrome c, correlated poorly with the patients' phenotypes, but that assays of POR-supported 17alpha-hydroxylase and 17,20 lyase activities correlated well. POR deficiency is a new disorder of adrenal and gonadal steroidogenesis that affects all microsomal cytochrome P450 enzymes, hence may have important implications for genetic differences in drug metabolism. PMID- 15666841 TI - Mutations in a novel gene, encoding a single transmembrane domain protein are associated with familial glucocorticoid deficiency type 2. PMID- 15666842 TI - The triple A syndrome is due to mutations in ALADIN, a novel member of the nuclear pore complex. AB - The triple A syndrome (MIM#231550) is a rare autosomal recessive disorder characterized by adrenocorticotropic hormone (ACTH) resistant adrenal failure, achalasia, alacrima, and a variety of neurological and dermatological features. The triple A syndrome is caused by mutations in the AAAS gene, which encodes a protein known as ALADIN (ALacrima Achalasia aDrenal Insufficiency Neurologic disorder). ALADIN is a new WD-repeat protein that has no significant homology to any previously identified WD-repeat protein. It has been shown that it colocalizes with nuclear pore complexes (NPCs), a finding that strongly suggests an involvement of ALADIN in nucleocytoplasmic transport. An investigation of 110 families with triple A syndrome disclosed mutation hot spots including Q15K (exon 1), and S293P (exon 8), which occur in 17 and 21 families from different geographical regions, respectively. The variable phenotype of all patients cannot be correlated with the localization and the nature of the ALADIN mutations. Thus, modifying genes/factors may be involved in the progression of this neurodegenerative disease. The lack of AAAS mutations in eight patients and negative linkage to chromosome 12q13 in three families are suggestive of genetic heterogeneity. To examine the cellular localization of ALADIN mutants causing triple A syndrome, we investigated nine different ALADIN-mutants: 2 nonsense (W84X, Q456X), 2 frameshift (F157fsX171, G397fsX414) and 5 point mutations (Q15K, L25P, H160R, S263P, L381R) by transfection experiments with green fluorescence protein. Mutants were predominantly localized in the cytoplasm, but also found in the nucleus indicating that ALADIN is essential for NPC targeting. To investigate physiological functions of ALADIN in vivo, we generated and analysed Aaas-/- knockout mice by homologous recombination in embryonic stem cells. Surprisingly, required animals lack any gross abnormality in adrenal and nervous system function. Further studies have to investigate the role of ALADIN at NPCs and to identify interacting proteins. Functional analyses of ALADIN may permit further understanding of its role for adrenocortical function and neurodevelopment. PMID- 15666843 TI - A mouse model for Carney complex. AB - Mice with complete inactivation of the type Ialpha regulatory subunit (RIalpha) of cyclic (c) AMP-dependent protein kinase (PKA) (coded by the Prkar1a gene) die early in embryonic life. To bypass the early embryonic lethality of Prkar1a-/- mice, we established transgenic mice carrying an antisense transgene for Prkar1a exon 2 (X2AS) under the control of a tetracycline-responsive promoter. Mice developed thyroid follicular hyperplasia and adenomas, adrenocortical hyperplasia, and other features reminiscent of PPNAD, and histiocytic and epithelial hyperplasias, lymphomas, and other mesenchymal tumors. This mouse provides a useful tool for the investigation of cAMP, RIalpha, and PKA functions and confirms Prkar1a's critical role in tumorigenesis in endocrine and other tissues. PMID- 15666844 TI - Nude mice as a model for gonadotropin-induced adrenocortical neoplasia. AB - Certain inbred mice (e.g., DBA/2J, CE) develop sex steroid producing adrenocortical tumors following gonadectomy. This adrenal response is thought to result from an unopposed increase in circulating gonadotropins and/or a decrease in factor(s) of gonadal origin. To differentiate between these two possibilities, we utilized the NU/J strain of nude mice, which are immunologically compromised and therefore permissive to xenografts. One group of female nude mice was gonadectomized, while another group of females received xenografts of CHO cells stably transfected with human chorionic gonadotropin (hCG). After 1-2 months, subcapsular adrenocortical neoplasms containing sex steroid-producing cells were observed in both groups. We conclude that high levels of circulating gonadotropins are sufficient to induce adrenocortical tumorigenesis, even in the presence of intact gonads. PMID- 15666845 TI - Transcription factors GATA-4 and GATA-6 in human adrenocortical tumors. AB - Transcription factors GATA-4 and GATA-6 are expressed during normal adrenocortical development in mice and humans, and in vitro studies have linked them to adrenal steroidogenesis. GATA-4 is highly expressed in the adrenocortical tumors of gonadectomized mice, whereas GATA-6 is down-regulated in the tumor area. Based on these findings we studied GATA-4 and GATA-6 expression in 39 human adrenocortical tumors using RT-PCR, Northern analysis and immunohistochemistry. 6/18 adenomas and 4/11 carcinomas were positive for GATA-4 mRNA. GATA-6 mRNA was expressed in 19/19 adenomas and 9/10 carcinomas, and GATA-6 immunoreactivity was remarkably lower in adrenocortical carcinomas than in adenomas (p < 0.05). Some of the steroidogenically active human adrenocortical cells (NCI-H295R) were weakly positive for GATA-4, whereas steroidogenically inactive cells (ACT-1) were totally GATA-4 negative. In contrast, both cell lines expressed GATA-6. GATA expression patterns similar to the animal models can thus be observed in human adrenocortical tumors, but the pathophysiological significance of these findings remains to be elucidated. PMID- 15666846 TI - Family of two patients with congenital lipoid adrenal hyperplasia due to StAR mutation. AB - We are reporting the case of two sisters born to nonrelated French Canadian parents. Patient A is of female phenotype with 46,xy, and patient B with 46,xx. The children had severe manifestations of mineralocorticoid deficiency at the age of 11 and 4.5 months, respectively. Residual cortisol secretion seemed present until the age of 3 years for patient A and until 15 months in the case of her sister. Both patients responded to glucocorticoid and Florinef treatment. Patient A did not show any androgen secretion and gonadectomy was performed at the age of 13.4 years; estrogen therapy was started at the age of 14 years resulting in a good breast development and an increase of growth velocity. In patient B, a progressive development of secondary sex characters occurred at 11.6 years of age followed at 14 years by menarche associated with a normal secretion of LH, FSH and estradiol; regular menstruations continued up to her last visit at the age of 25 years. We identified a homozygous L275P mutation on the StAR gene of both patients and a heterozygous L275P mutation on that of their mother and father. In transfection analysis in COS-1 cells, the mutant L275P was well-expressed, but its StAR activity was 87% impaired. The remaining activity of the L275P StAR mutant is consistent with the moderate severity of clinical onset of manifestations. PMID- 15666847 TI - Immunohistochemical determination of somatostatin receptor subtypes 1, 2A, 3, 4, and 5 in various adrenal tumors. AB - While octreotide binds with high affinity to sst2a only, the new analogue SOM230 demonstrates high affinity for sstl, 3, and 5, in addition. We examined the immunohistochemical expression of somatostatin receptor subtypes (sst) in 7 pheochromocytomas (PHEO), 5 Conn adenomas (CONN), 9 Cushing adenomas (CUSH), 7 nonfunctioning tumors (NFA), and 4 adrenal carcinomas (CA). About one third of the PHEO were positive for sst1, 2a, and 5. Less than 30% of cells were stained in the majority of these tumors. Each of the PHEO expressed sst3 with more than 60% of cells stained. Two thirds of the NFA revealed positive staining for sst1, 2a, and 3 with less than 30% of cells affected. Sst5 was expressed in nearly all of the NFA with positive staining in 30-60% of tumor cells. Nearly all CUSH and CONN were positive for the subtypes evaluated. In the majority of these tumors, less than 30% of cells were positively stained. Fifty percent of CA expressed sst2a and 3 with positive staining in 30-100% of cells. None of them expressed sst1. Somatostatin receptors are expressed in adrenal tumors with a tumor specific distribution pattern. This may offer new diagnostic and therapeutic possibilities. PMID- 15666848 TI - The higher affinity of human type 1 3beta-hydroxysteroid dehydrogenase (3beta HSD1) for substrate and inhibitor steroids relative to human 3beta-HSD2 is validated in MCF-7 tumor cells and related to subunit interactions. AB - Two distinct genes encode the tissue-specific expression of the two isoforms of human 3beta-hydroxysteroid dehydrogenase: 3beta-HSD1 (placenta, mammary gland, breast tumors) and 3beta-HSD2 (gonads, adrenals). Purified 3beta-HSD1 utilizes DHEA as a substrate with 13-fold lower Km than 3beta-HSD2. Using homogenates of human MCF-7 Tet-off breast tumor cells stably transfected with human 3beta-HSD1 or 3beta-HSD2, DHEA is utilized as substrate by 3beta-HSD1 (Km = 4.8 microM) much more avidly than by 3beta-HSD2 (Km = 43 microM). In addition, the 3beta-HSD inhibitor, epostane, binds to purified 3beta-HSD1 with a 17-fold higher affinity compared to 3beta-HSD2. In the MCF-7 cells, two 3beta-HSD inhibitors block 3beta HSD1 activity (Ki = 0.06 microM, epostane; 0.08 microM, trilostane) with 12- to 14-fold higher affinities compared to the inhibition of 3beta-HSD2 (Ki = 0.85 microM, epostane; 1.01 microM, trilostane). Thus, the substantially higher affinities of human 3beta-HSD1 for substrate and inhibitor steroids measured using the pure isoenzymes have been validated using microsome-bound 3beta-HSD1 and 3beta-HSD2 in the MCF-7 cells. Similar to our previously reported H156Y mutant of 3beta-HSD1, the Q105M mutant of 3beta-HSD1 shifts the substrate and inhibitor kinetic profiles to those of wild-type 3beta-HSD2. However, the Q105M mutant of 3beta-HSD2 retains the substrate and inhibitor kinetic profiles of wild type 3beta-HSD2. Our structural homology model of human 3beta-HSD predicts that Gln105 on one enzyme subunit hydrogen-binds to His156 on the other subunit of the enzyme homodimer. The higher affinity of 3beta-HSD1 for the steroids may be related to different subunit interactions in the quaternary structures of the two isoenzymes. It may be possible to exploit these kinetic differences to selectively inhibit the conversion of DHEA ultimately to estradiol by 3beta-HSD1 and slow the growth of breast tumor cells. PMID- 15666849 TI - A neonatal form of isolated ACTH deficiency frequently associated with Tpit gene mutations. PMID- 15666850 TI - ACTH and PRL sensitivity of highly differentiated cell lines obtained by adrenocortical targeted oncogenesis. AB - We established cell lines from adrenal tumors of transgenic mice harboring the large T-antigen of simian virus 40 under the control of the adrenocortical specific promoter of the scavenger aldose reductase-like akr1b7 gene. Mass spectrometry analyses of serum-supplemented or serum-free culture media showed that ATC1 line secreted only corticosterone. These cells, propagated over 25 passages, were characterized with regard to ACTH and PRL responsiveness, as measured by increased corticosterone production, induction of genes involved in the different steps of steroidogenesis (cholesterol delivery, steroid biosynthesis and detoxification of by-products) and expression of transcriptional regulators (SF-1 and DAX1). Corticosterone secretion (RIA) in serum-free medium was stimulated over 12-fold after 6 h treatment with either 10(-9)M ACTH or PRL and both hormones seemed equivalent in promoting this secretion (149 +/- 14 ng and 145 +/- 18 ng/10(6) cells/6 h, respectively). As expected, Northern blots indicate that ATC1 cells expressed mRNAs for the enzymes of corticosterone metabolism CYP11B1 and CYP21A, as well as those for the proteins SIK, SRB1, StAR, CYP11A1, and AKR1B7. Interestingly, these cells have maintained not only the expression of SF-1 but also that of DAX1. No expression of the zona glomeruloza specific cyp11b2 gene was detected. With the exception of cyp21a and mc2r genes which were constitutively expressed, most of the genes above mentioned were induced in a time- and dose-dependent fashion in response to ACTH or PRL while DAX1 was repressed. Importantly, hormone-mediated repression of DAX1 gene expression was also observed in vivo in mice adrenals. Altogether these data demonstrate that ATC1 line provided an unique model of well differentiated zona fasciculata immortalized cells suitable for the dissection of molecular events leading to ACTH and PRL regulation of adrenal functions. PMID- 15666851 TI - Paradoxical inhibitory effect of serotonin on cortisol production from adrenocortical lesions causing Cushing's syndrome. AB - In the human adrenal gland, serotonin (5-HT) stimulates cortisol production through a paracrine mechanism involving 5-HT4 receptors positively-coupled to adenylyl cyclase. A hyperresponsiveness of adrenocortical tissue to 5-HT has also been described in several cases of ACTH-independent bilateral macronodular adrenal hyperplasias (AIMAHs) and adenomas causing Cushing's syndrome. In the present study, we report two cases of cortisol-producing adrenocortical lesions, i.e., one AIMAH (case 1) and one adenoma (case 2), whose secretory activity was inhibited in vitro by 5-HT. The potencies (pIC50) and efficacies (Emax) of 5-HT to inhibit cortisol secretion were 8.2 +/- 0.4 and -64.1% +/- 7.5% in case 1, and 9.2 +/- 0.5 and -32.3% +/- 3.8% in case 2. The specific 5-HT4 antagonist GR 113808 failed to influence the 5-HT-induced decrease in cortisol production by the two tissues, indicating that the paradoxical inhibitory effect of 5-HT could not be accounted for by activation of eutopic 5-HT4 receptors. These results suggest that the tissues expressed aberrant 5-HT receptors. In conclusion, the present study provides the first evidence for an inhibitory effect of 5-HT on cortisol secretion in adrenocortical lesions causing Cushing's syndrome. Our data also suggest that expression of illegitimate membrane receptors by cortisol producing adrenal hyperplasias and/or adenomas may convert a paracrine stimulatory factor into an inhibitory signal. PMID- 15666852 TI - Effect of anticonvulsants on activities of P450c17 and 3betaHSDII. PMID- 15666853 TI - A male twin infant with skull deformity and elevated neonatal 17 hydroxyprogesterone: a prismatic case of P450 oxidoreductase deficiency. AB - We report on a male twin infant who presented with brachy-turri-cephaly, frontal bossing, large anterior fontanelle, low set and malformed ears, and mild arachnodactyly. He had normal male genitalia. There was no evidence for maternal virilization during pregnancy. The pattern of malformations resembled Antley Bixler-Syndrome (ABS). However, sequencing analysis of the fibroblast growth factor receptor 2 gene (FGFR2) did not reveal mutations. The boy's twin sister did not show any somatic or endocrine abnormalities. In the boy, neonatal screening for congenital adrenal hyperplasia was positive with moderately elevated 17-hydroxyprogesterone. Sequence analysis of his CYP21 gene did not reveal any mutations. The short synacthen test revealed an exaggerated 17 hydroxyprogesterone and a blunted cortisol response. Urinary steroid profiling by gas chromatography-mass spectrometry (GC-MS) revealed a unique steroid metabolome suggestive of impaired activity of both 17-hydroxylase and 21-hydroxylase. Clinical and metabolic findings therefore were compatible with the recently described variant of congenital adrenal hyperplasia, P450 oxidoreductase deficiency (ORD). Subsequently, sequencing analysis of CPR, the gene encoding P450 oxidoreductase (OR), revealed a homozygous mutation in the patient, resulting in an amino acid exchange in position 284 of the OR protein (A284P). Both the female twin sister and the parents were heterozygous for the A284P mutation. P450 oxidoreductase deficiency represents a novel autosomal recessively inherited form of congenital adrenal hyperplasia. Its characteristic steroid metabolome can readily be detected by GC-MS analysis of spot urine. Clinical features may include an ABS phenotype, ambiguous genitalia (virilization in girls, feminization in boys), and glucocorticoid deficiency. If required, hydrocortisone replacement should be provided. PMID- 15666854 TI - Chicken infectious anemia virus: an example of the ultimate host-parasite relationship. AB - Chicken infectious anemia virus (CIAV) is a resistant and ubiquitous virus of chickens causing disease in young chickens and immunosuppression in all birds. This paper reviews the current knowledge of CIAV with a focus on new findings indicating that immunosuppressive effects have not been fully appreciated, especially as they relate to the development of antigen-specific cytotoxic T cells. A more complete understanding of the immunosuppressive effects of CIAV emphasizes the need for better vaccines, especially for the broiler industry. In addition, a new model is proposed for the control of viral replication in the reproductive tract of specific-pathogen-free chickens, which may be latently infected. This model suggests that virus transcription is controlled by viral enhancer and repressor elements, which are regulated by different hormones. As a consequence, CIAV has a well-adapted relationship with its host, avoiding immune detection, ensuring passage of virus to the next generation, and eliciting limited pathology to the host. PMID- 15666855 TI - Infection of turkeys with Histomonas meleagridis by the cloacal drop method. AB - The infection of turkeys with Histomonas meleagridis was attempted in the absence of its normal vector Heterakis gallinarum, using several experimental techniques. Battery-reared poults were inoculated at 2 wk of age with histomonads cultured in vitro, by several routes, including (a) per os (PO), (b) intradoacal (CI), and (c) cloacal drop (CD). Feed restriction was also studied as a predisposing factor. Intracloacal inoculation (CI) consistently produced severe infections in all experiments. In several experiments, turkeys did not become infected after inoculation PO with 1 x 10(5) cultured histomonads. Feed restriction prior to inoculation did not make turkeys susceptible to infection inoculated PO. However, when liquid cultures containing histomonads were applied to the vent (CD) and the dorsal lip stimulated to initiate cloacal drinking, the histomonads were taken into the cloaca and transported to the ceca by retrograde peristalsis. Heavy infections were produced by this method, with severe liver and cecal lesions recorded when birds were necropsied 12 days later. These results suggest that CD may provide ready entry into the lower intestinal tract for these parasites and may facilitate spread of infection through flocks. PMID- 15666856 TI - Comparison of several challenge models for studies in avian colibacillosis. AB - In previous studies, the embryo lethality assay (ELA) discriminated between virulent and avirulent avian Escherichia coli isolates, and also proved to be highly correlated with mortality and morbidity results of the intravenous (IV) challenge model. In the current study, the same 20 avian E. coli isolates were used in subcutaneous (subQ) and intratracheal (IT) chicken challenge models in order to determine whether the results from the prior ELA challenges and/or the IV challenge model correlate with these models. The correlation observed between the two previous ELA trials and the combined mortality/morbidity percentages of the subQ challenge model were r = 0.792, P > 0.0001 for the first ELA trial and r = 0.738, P = 0.0002 for the second ELA trial. The IV challenge results were more highly correlated with the subQ challenge results (mortality/morbidity comparison, r = 0.894, P < 0.0001). The IV challenge mortality results were slightly correlated (r = 0.4810, P=0.0319) with the IT challenge results. Several of the isolates differed in their ability to produce mortality and/or morbidity with the different challenge models. The mortality/morbidity results of the IV and subQ challenges and the mortality results of the ELA were all positively correlated with the ability of an E. coli isolate to produce Colicin V (ColV) (r = 0.7131, P = 0.0004). The IT mortality results were slightly correlated with the production of ColV (r = 0.455, P = 0.049). The IT challenge results were only slightly correlated with resulting IV mortality and ColV production. Previous results indicate that the ELA correlates extremely well with the IV challenge model. The current study demonstrates that ELA also correlates well with the subQ challenge model. Overall, the conclusion of this study is that the ELA, IV, and subQ challenge models similarly demonstrate the ability to discriminate between virulent and avirulent avian E. coli isolates. PMID- 15666857 TI - Survey of parasites and bacterial pathogens from free-living waterfowl in zoological settings. AB - To determine if free-living waterfowl residing in a zoological setting pose health risks for its animal collections, visitors, and employees, 450 fecal samples were collected and cultured for the presence of Campylobacter jejuni, Escherichia coli, Salmonella spp., and Pasteurella multocida. A survey of endoparasites infecting the waterfowl was also conducted. Sixty-seven percent, 42%, and 1.7% of the samples tested positive for E. coli, C. jejuni, and Salmonella spp., respectively. No P. multocida was isolated from the sampled population. Antimicrobial susceptibility testing for the bacterial isolates demonstrated that a majority of the isolates were susceptible to the antibiotics tested. A survey for parasites revealed 16% of the samples had coccidia oocytes; 8% of the sample had spirurid ova; and 17% of the sample had strongylate-type nematode ova. Ascaris spp. ova, Capillaria spp. ova, oxyurid ova, and mites were also noted in some fecal samples. PMID- 15666858 TI - Serotype 1 viruses modified by backpassage or insertional mutagenesis: approaching the threshold of vaccine efficacy in Marek's disease. AB - Improved vaccines to control Marek's disease (MD) in chickens are desired by the poultry industry but have been difficult to develop. Studies were conducted to evaluate strategies for deriving MD vaccines of high protective efficacy, irrespective of virulence. Candidate viruses from parent strains representing v and vv+ pathotypes were modified by cell culture passage, backpassage in chickens, or insertional mutagenesis following cocultivation with retroviruses. Ten strains considered most likely to exhibit high protective efficacy were selected for further study. The ability of these modified viruses to protect commercial or maternal antibody-positive (ab+) chickens against virulent MD virus (MDV) challenge was compared with that of strain CVI988, the standard commercial MD vaccine. Modified strains were also evaluated for the ability to induce lymphomas or other pathologic changes in ab+ and antibody-negative (ab-) chickens. Two of the 10 modified viruses, strains RM1 and CVI988/BP5, provided high levels of protection against highly virulent MDV challenge. The magnitude of protection was greater than that of one laboratory and two commercial preparations of CV1988, but was approximately equal to that of two other commercial preparations of CVI988 in laboratory and field tests. Three of the strains, including RMI and CVI988/BP5, induced lymphoid organ atrophy in ab chicks but not in ab+ commercial chicks, a property designated here as L phenotype. Seven strains, including two L+ strains, were mildly oncogenic for ab- chicks, a property designated here as O phenotype. Five of these strains caused no tumors in ab+ chickens. The two fully attenuated strains induced neither lymphomas nor lymphoid organ atrophy. The L and O phenotypes appeared not to be linked, and both (especially the L phenotype) appeared associated with high levels of protection. These studies also illustrated differences in the protective efficacy of different preparations of CVI988 vaccine, indicating the need to choose carefully the most protective strains as controls for efficacy studies. A new vv+ strain, designated as 686, is described and appears useful as a challenge virus; it is the most virulent of the 48 field isolates of MDV thus far pathotyped in this laboratory. These findings support the conclusion that new virus strains with high levels of protective immunity comparable to that of CVI988 can be developed. However, the question of whether strains can be developed that exceed the efficacy of current CVI988-based vaccines remains unanswered. After more than 30 years of unsuccessful endeavor by many laboratories toward this goal, it now may be useful to consider whether the efficacy of MD vaccines is limited by some type of biologic threshold. PMID- 15666859 TI - In vivo effects of CpG oligodeoxynucleotide on Eimeria infection in chickens. AB - Poultry coccidiosis is the major parasitic disease of poultry and, until now, no recombinant vaccine has been developed. Short oligodeoxynucleotides (ODNs) containing unmethylated CpG motifs (CpG ODNs) have been shown to be effective immunoprotective agents and vaccine adjuvants in mammalian systems. Their use in poultry to protect against intracellular parasites has not been reported to date. The present work investigated the effects of CpG ODN treatment on host susceptibility to Eimeria infection in two chicken strains with different genetic background, SC and TK. The data show that CpG ODN enhanced the birds' resistance to coccidiosis in a normally susceptible chicken strain (TK), as shown by reduced oocyst shedding and improved weight gain. CpG treatment had a differential effect on body weight gains and serum antibody responses, depending on the chicken strain and ODN dose, delivery route, and backbone. This study shows for the first time that CpG ODNs could be used as immunoprotective agents in Eimeria-infected chickens to enhance resistance to the pathogen and improve performance. Future research is needed to optimize their use alone and as vaccine adjuvants that may lead to better and more efficient vaccine applications. PMID- 15666860 TI - Serotyping and virulence genes detection in Escherichia coli isolated from fertile and infertile eggs, dead-in-shell embryos, and chickens with yolk sac infection. AB - Escherichia coli is a common avian pathogen mainly associated with extraintestinal infections such as yolk sac infection (YSI). The aim of this study was to determine the serotypes and the presence of some virulence genes of E. coli strains isolated from different samples in a vertically integrated poultry operation in Mexico. Two hundred sixty-seven E. coli isolates from different samples were serotyped using rabbit serum against the 175 somatic (O) and 56 flagellar (H) antigens of the typing schema. Virulence genes were determined by colony blot hybridization, using DNA probes for st, eae, agg1, agg2, bfp, lt, cdt, slt, and ipaH diarrhea-associated virulence factors. The serogroup of 85% of the strains was determined; O19 (12%), 084 (9%), 08 (6%), and 078 (5%) were the most common. Using the complete antigenic formula (O and H), O19:NM (n = 31) was the serotype most frequently isolated from dead-in-shell embryos and in broilers that had died on the fourth, fifth, sixth, and seventh days after hatch. One hundred ten strains (41.2%) hybridized with one or more of the used probes. Of these, ipaH (72%), eae (30%), and cdt (27%) were the most common. Considering the origin of the respective isolates, 40% of the broiler farm strains were positive for at least one probe. Results show that some avian E. coli strains isolated in Mexico are included in avian pathogenic E. coli serotypes not previously reported, suggesting that they could be specific for this geographic area. The wide distribution of the ipaH gene among nonmotile strains suggests that this invasiveness trait could be important in YSI pathogenesis. On the other hand, some other genes could contribute to E. coli virulence during YSI. PMID- 15666861 TI - Quail cell lines supporting replication of Marek's disease virus serotype 1 and 2 and herpesvirus of turkeys. AB - Marek's disease virus (MDV), a highly cell-associated alphaherpesvirus, can be isolated and propagated in chicken kidney cells (CKC) and chicken or duck embryo fibroblast cells (CEF or DEF, respectively). Two recently developed cell lines, CU447 and CU453, developed from methylcholanthrene-induced tumors in Japanese quail, were examined for their suitability to propagate the three serotypes of MDV. The MDV strain RB-1B (serotype 1) was passaged for more than 30 passages in CU447 without causing cytopathic effects (CPE). Polymerase chain reaction analysis of RB-1B-infected CU447 cells demonstrated the presence of MDV DNA using primers specific for ICP4, pp38, and gB. The 132-bp direct repeats within the BamH1-H and -D fragments were amplified to the same level as RB-1B that was passaged in CKC or CEF. Different passages of RB-1B in CU447 were examined for expression of gB and pp38 transcripts, and pp38, gB, gE, and VP22 protein expression. Irrespective of the passage level, these transcripts and proteins were detected in the RB-1B-infected CU447 cells. Infectious virus was rescued by cocultivation of RB-1B-infected CU447 with CKC. Herpes virus of turkeys was propagated in CU447 and CU453 causing CPE in both cell lines. SB-1 did not cause CPE in either cell line but a few SB-1-infected cells could be detected using a monoclonal antibody specific for serotype 2 MDV. PMID- 15666862 TI - Protection of neonatal chicks against a lethal challenge of Escherichia coli using DNA containing cytosine-phosphodiester-guanine motifs. AB - Oligodeoxynucleotides (ODN) containing cytosine-phosphodiester-guanine (CpG) motifs have been shown to be effective immunoprotective agents in murine models for a variety of viral, intracellular bacterial, and protozoan infections. We recently have shown that CpG ODN protects against extracellular bacterial infections in mature chickens. The objective of this study was to investigate the effect of CpG ODN on Escherichia coli septicemia in neonatal broiler chicks. Two day-old chicks, or embryonated eggs that had been incubated for 18 or 19 days, received 50 microg CpG ODN. Three days after exposure to CpG ODN, a virulent isolate of E. coli was inoculated subcutaneously in the neck of each bird. Birds were examined for 7 days post-E. coli challenge and dinical, pathologic, and bacteriologic assessments were conducted. The control group of birds that received no CpG ODN had a survival rate of 0% to 20%. In contrast, groups that received CpG ODN, either by intramuscular or in ovo routes, had significantly higher survival rates (P < 0.0001). Bacterial counts in air sacs were significantly lower when birds or embryos were treated with CpG ODN as compared with controls. A dose as low as 10 microg of CpG ODN, administered intramuscularly, was able to protect birds significantly against E. coli challenge. Formulation of CpG ODN with 30% Emulsigen did not enhance the protection. This study demonstrates that CpG ODN has systemic protective effects in broiler chicks against E. coli infections. This is the first time that CpG ODN has been demonstrated to have an immunoprotective effect against a bacterial infection in chicks following in ovo delivery. PMID- 15666863 TI - Prevalence of vancomycin resistant enterococci on poultry farms established after the ban of avoparcin. AB - Fecal samples from poultry on farms established after the ban of avoparcin (study farms) and from poultry on farms previously exposed to avoparcin (control farms) were examined for the presence of vancomycin-resistant enterococci (VRE). The samples were collected during the autumn and winter of 2001-2002. One isolate from each positive sample was selected, identified to species level, and examined for the presence of the vanA gene. The concentration of VRE and generic enterococci in the samples were also determined. In addition, the susceptibility to the ionophoric coccidiostat narasin was examined in a number of enterococcal isolates from poultry and in some enterococci of porcine origin that had not been exposed to narasin. VanA-type VRE was detected in samples from 64% of the study farms and 96% of the control farms. However, the concentration of VRE in the control samples was about six times larger than in the samples from the study farms. The minimum inhibitory concentration values for narasin differed between the poultry (1-4 mg/liter) and the porcine (0.25-0.5 mg/liter) isolates, indicating a decreased susceptibility towards narasin among enterococci from poultry. PMID- 15666864 TI - A very virulent genotype of infectious bursal disease virus predominantly associated with recurrent infectious bursal disease outbreaks in Tunisian vaccinated flocks. AB - Outbreaks of infectious bursal disease (IBD) still continue to afflict the Tunisian poultry industry even in those flocks where the vaccination program is strictly applied. To characterize the viruses that circumvent protection provided by vaccination, field isolates of infectious bursal disease virus (IBDV) obtained from vaccinated flocks that have repeatedly experienced IBDV outbreak episodes were analyzed from bursal samples by reverse transcription coupled with polymerase chain reaction and dideoxynucleotide sequencing of the VP2 hypervariable region. Although sequence data were obtained from samples collected from three distinct flocks over a period of 3 years, only limited sequence variation has been observed. The few nucleotide changes were silent and the deduced amino acid sequences were identical. Thus, the virus population that predominates in the field seems to represent a homogeneous antigenic pool. Compared with the VP2 sequences of several IBDV strains, this predominant pool was found to be closely related to the very virulent (vv) IBDV viruses described in Europe and Asia. Sequence and phylogenetic analysis of the precursor polyprotein coding sequence of a representative Tunisian isolate further confirmed its assignment to the vv genotype. The deduced amino acid sequence of the whole polyprotein of the Tunisian isolate was found to be identical to a South Korean IBDV strain. Alignment of the polyprotein amino acid sequence of 35 IBDV strains identified additional mutations outside the VP2 variable domain and which occur frequently in vv strains. Based on this comparative analysis, the set of amino acid residues that should represent a typical vv profile involves Ala222, Ile242, Ile256, Ile294, Leu451, Tyr680, N685, Ser715, Asp751, Val990, and Ala1005. Such a combination of amino acid changes was observed for the majority of vvIBDV strains that define a distinct phylogroup. PMID- 15666865 TI - Inactivation, storage, and PCR detection of Mycoplasma on FTA filter paper. AB - We evaluated the feasibility of using Flinders Technology Associates (FTA) filter paper for the inactivation and storage of mycoplasma DNA templates and their detection by the polymerase chain reaction (PCR). FTA paper is a cotton-based cellulose membrane containing lyophilized chemicals that lyses most types of bacteria and viruses. Mycoplasma gallisepticum (MG) and Mycoplasma synoviae (MS) cultures were spotted at various volumes on the filter paper and stored at different temperatures for various periods of time before performing PCR. MG and MS were readily detected at all time frames (1-60 days) independent of the volume applied (1-100 microl) or storage temperature (4 C-41 C). Sensitivity and specificity of the FTA-PCR were comparable to the standard diagnostic PCR, allowing the detection of MG/MS in field samples without interference by nontargeted mycoplasma. Analysis of 193 field samples by both methods showed nearly 100% agreement with serology and culture results. The long-term DNA stability at a wide range of temperatures makes the FTA cards a good alternative for collecting and simultaneously inactivating mycoplasma. It also offers the convenience of storage and transport of DNA in a cost-effective manner for further molecular analysis, such as restriction enzyme length polymorphism and nucleotide sequencing. PMID- 15666866 TI - Heteroduplex mobility assay for genotyping infectious bursal disease virus. AB - A heteroduplex mobility assay (HMA) was developed to genotype infectious bursal disease virus (IBDV). This method analyzed 390-base pair (bp) polymerase chain reaction (PCR) products, encompassing the hypervariable region of the VP2 gene. IBDV strains from the United States and other countries were analyzed. The HMA was able to differentiate standard, antigenic variants and very virulent strains of IBDV. Minor differences between different strains from the same subtype were also detected. Close relationships between field IBDV with vaccines prepared with Delaware E strain were determined by HMA. The results obtained by HMA were confirmed by restriction fragment length polymorphism (RFLP) and phylogenetic analysis of nucleotide sequences. The HMA proved to be a useful technique to rapidly genotype different field strains of IBDV and should prove to be a useful tool in epidemiologic studies. PMID- 15666867 TI - Colonization of reproductive organs and internal contamination of eggs after experimental infection of laying hens with Salmonella heidelberg and Salmonella enteritidis. AB - Internal contamination of eggs laid by hens infected with Salmonella enteritidis has been a prominent international public health issue since the mid-1980s. Considerable resources have been committed to detecting and controlling S. enteritidis infections in commercial laying flocks. Recently, the Centers for Disease Control and Prevention also reported a significant association between eggs or egg-containing foods and S. heidelberg infections in humans. The present study sought to determine whether several S. heidelberg isolates obtained from egg-associated human disease outbreaks were able to colonize reproductive tissues and be deposited inside eggs laid by experimentally infected hens in a manner similar to the previously documented behavior of S. enteritidis. In two trials, groups of laying hens were orally inoculated with large doses of four S. heidelberg strains and an S. enteritidis strain that consistently caused egg contamination in previous studies. All five Salmonella strains (of both serotypes) colonized the intestinal tracts and invaded the livers, spleens, ovaries, and oviducts of inoculated hens, with no significant differences observed between the strains for any of these parameters. All four S. heidelberg strains were recovered from the interior liquid contents of eggs laid by infected hens, although at lower frequencies (between 1.1% and 4.5%) than the S. enteritidis strain (7.0%). PMID- 15666868 TI - Development of enzyme-linked immunosorbent assay with nucleoprotein as antigen for detection of antibodies to avian influenza virus. AB - During the avian influenza outbreak of 2003-04 in Southeast Asia, two avian influenza viruses (AIV), one of H5N1 subtype and the other H9N2 subtype, were isolated and identified from local farms. The nudeoprotein (NP) gene of the H5N1 AI isolate was cloned, and the segment encoding amino acid 47-384, which covers its major antigenic domains, was subcloned and expressed in E. coli. Subsequently, the NP (47-384) expression product was purified and used as the diagnostic antigen to develop a NP-based type-specific indirect enzyme-linked immunosorbent assay (ELISA) for detecting antibodies to AI from chicken sera. The ELISA is shown to be specific for AIV and does not cross-react with chicken sera that has antibodies to other avian viruses. The NP(47-384)-ELISA was compared with a hemagglutination inhibition test and a commercial AIV ELISA kit in evaluating 150 sera samples from experimentally AIV-infected or vaccinated specific-pathogen-free (SPF) chickens. Our NP(47-384)-ELISA was more sensitive than the two tests and showed an 82% agreement ratio with the HI test and an 80.67% agreement ratio with the commercial kit. The NP(47-384)-ELISA and the commercial AIV ELISA were used to evaluate 448 field sera samples from diseased chickens or vaccinated chickens during the 2003-04 AI outbreak in China. The two ELISA tests had a 95% agreement ratio. We conclude that the NP(47-384)-ELISA developed in our laboratory was specific and sensitive and it has great application potential in China's long-term prevention and control of AI. PMID- 15666869 TI - Effects of Mycoplasma gallisepticum on reproductive success in house finches. AB - Long known as a pathogen of poultry, Mycoplasma gallisepticum (MG) was first detected in house finches in 1994. The disease rapidly spread throughout the eastern United States and Canada and was associated with debilitating disease and high mortality in house finches. However, in the late 1990s, the proportion of infected finches dying as a result of infection with MG decreased, and asymptomatic infection was more common among wild birds than in the past. We documented MG infections in breeding house finches and concluded that adults of both sexes transmit the infection to dependent young, probably after hatch. MG infections of breeding adults occurred late in the breeding season and were found in birds completing significantly more nests than birds that never tested positive for MG, implying that higher rates of reproduction carry a cost in the form of increased risk of infection. We found evidence of an MG-induced delay in dispersal of nestlings from their natal area and demonstrated a significant impact of infection on nestling growth. PMID- 15666870 TI - Virulence of the nine serovar reference strains of Haemophilus paragallinarum. AB - The virulence of the reference strains of the nine currently recognized Kume serovars of Haemophilus paragallinarum was investigated. The capacity of the H. paragallinarum strains to cause the typical clinical signs of upper respiratory tract disease associated with infectious coryza in unvaccinated, nasal-challenged chickens was assessed. Differences in virulence were assessed by means of a standardized scoring system for clinical signs. All nine strains were pathogenic to chickens, producing typical clinical signs of infectious coryza. The highest clinical signs score was obtained for serovar C-1 (1.72), while the lowest clinical signs score was obtained for serovar C-4 (0.32). Our results indicate that virulence differences exist among the serovars of H. paragallinarum. PMID- 15666871 TI - Typing of Haemophilus paragallinarum strains by using enterobacterial repetitive intergenic consensus-based polymerase chain reaction. AB - The enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) technique was used for fingerprinting of reference strains and Mexican isolates of Haemophilus paragallinarum. A total of nine ERIC patterns were given by the nine serovar reference strains of this bacteria. Two Modesto (C-2) reference strains from different sources showed the same ERIC pattern. Seventeen ERIC patterns were obtained among 29 Mexican isolates included in the study, belonging to serovars prevalent in Mexico (A-1, A-2, B-1, and C-2). Obtained results indicate that the ERIC-PCR technique could be used as a molecular laboratory tool for subtyping of H. paragallinarum. PMID- 15666872 TI - Antigenicity of two turkey astrovirus isolates. AB - Astroviruses are positive-sense single-stranded RNA viruses. These viruses cause gastroenteritis in humans and in a variety of animal species, including turkey poults. Only human astroviruses are well characterized antigenically. In the current study, two turkey astrovirus isolates, TAstV1987 and TAstV2001, were antigenically compared using cross-neutralization tests in turkey embryos, as well as cross-reactivity of the two isolates by an enzyme-linked immunosorbent assay (ELISA). The antigenic relatedness values (R) were calculated using the Archetti and Horsfall formula. The R value based on the cross-neutralization tests was 0.56%, which indicates that TAstV1987 and TAstV2001 belong to different serotypes; the R value of the two viruses based on ELISA was 70.7%, which suggests these two viruses share common antigen(s). PMID- 15666873 TI - Evidence of avian pneumovirus spread beyond Minnesota among wild and domestic birds in central North America. AB - To detect avian pneumovirus (APV) in central North America, nasal turbinates or choanal deft tissues from domestic turkeys and wild birds were examined for the presence of APV RNA by reverse transcriptase-polymerase chain reaction (RT-PCR), whereas serum samples from domestic turkeys were analyzed for APV antibodies by enzyme-linked immunosorbent assay (ELISA). In 2002, the seroprevalence of disease in domestic turkeys in Minnesota remained high (42.3% of the flocks). In addition, there is evidence the disease has spread to turkey flocks in North Dakota (8.2%), South Dakota (7%), Iowa (10%), and Wisconsin (8.6%) as detected by RT-PCR and/or ELISA. House sparrows and ring-billed gulls sampled in Minnesota and snow geese from Saskatchewan, Canada, were found to harbor APV RNA. Sequence analysis of wild bird APV strains showed high amino acid sequence identity among wild bird isolates (<97%) and between wild bird and turkey viral isolates (93.2% 99.3%). This study demonstrated that APV infections were present in domestic turkey flocks and wild birds outside the state of Minnesota; however, the role of wild birds in spreading APV to domestic turkeys remains unclear. PMID- 15666874 TI - Avian infectious bronchitis virus: a possible cause of reduced fertility in the rooster. AB - The formation of epididymal stones in the rooster epididymis is a widespread problem that has detrimental effects on sperm production and fertility. The cause of epididymal stones is unknown, but an infectious agent, the avian infectious bronchitis virus (AIBV), has been implicated. The goal of this study was to determine if administering the live attenuated AIBV vaccine to male chicks increases the incidence of stones in the epididymal region of the adult rooster. Specific pathogen free (SPF) Leghorn roosters were divided into two groups: a vaccine-free group (n = 7) and a group vaccinated with AIBV (n = 12). The vaccine was administered orally at 2, 4, 10, and 14 wk of age. Blood was drawn weekly to monitor antibodies to AIBV. At 26 wk of age, blood was obtained to determine testosterone concentrations, and reproductive tracts were removed to analyze daily sperm production and to detect epididymal stones. Nine of 12 vaccinated roosters developed stones, whereas those not given the vaccine did not develop stones. Serum testosterone concentrations were significantly (P < 0.05) reduced in vaccinated roosters with epididymal stones (3.6 +/- 0.30 ng/ml) when compared with nonvaccinated roosters that did not have epididymal stones (7.0 +/- 1.63 ng/ml). Testis weight was significantly (P < 0.05) reduced in vaccinated roosters with epididymal stones (12.1 +/- 0.76 g), as compared with nonvaccinated roosters without epididymal stones (15.2 +/- 0.81 g). Daily sperm production was significantly (P < 0.05) decreased in vaccinated roosters with epididymal stones (5.03 +/- 0.31 x 10(8) sperm/testis/day) when compared with nonvaccinated roosters without epididymal stones (7.43 +/- 0.52 x 10(8) sperm/testis/day). Comparing daily sperm production on a per gram basis, vaccinated roosters with epididymal stones had 4.38 +/- 0.14 x 10(7) sperm/g of testis, which was significantly (P < 0.05) smaller than nonvaccinated roosters without epididymal stones, which had 5.17 +/- 0.17 x 10(7) sperm/g of testis. We conclude that the use of a live attenuated AIBV vaccine increases the incidence of epididymal stones in roosters, resulting in decreased sperm production and decreased serum testosterone concentrations. PMID- 15666875 TI - Pathogenicity of Mycoplasma meleagridis for chicken cells. AB - Mycoplasma meleagridis (MM) is a known pathogen for turkeys only. In this study, MM was used to inoculate chicken embryos and tumor cells to assess its pathogenic potential for chickens. In chicken embryos, it caused abnormal-shaped toes and severely denuded tracheae. In chicken tumor cells, MM reduced the cellular capacity to release a chemoattractant that causes the migration of heterophils. MM also caused death and/or a reduced growth rate in chicken HD-11 cells, a macrophage-monocyte-derived cell line. Thus, the data show that MM is a potential pathogen for chicken embryos and chickens cells. Further exploration to determine the pathogenicity in chickens may be warranted. PMID- 15666876 TI - Tissue tropism and bursal transformation ability of subgroup J avian leukosis virus in White Leghorn chickens. AB - In Experiment 1, a monoclonal antibody against the envelope glycoprotein (gp85) of subgroup J avian leukosis virus (ALV-J) was used to study the distribution of ALV-J in various tissues of White Leghorn chickens inoculated as embryos with the strain ADOL-Hcl of ALV-J. At 2 and 6 wk of age, various tissues from infected and control uninfected chickens were tested for the presence of ALV-J gp85 by immunohistochemistry. In Experiment 2, using the methyl green-pyronine (MGP) stain, sections of bursa of Fabricius (BF) from chickens of line 15I5 x 7(1), inoculated with ALV-J or Rous-associated virus-1 (RAV-1), a subgroup A ALV, at hatch were examined for transformation of bursal follicles at 4 and 10 wk of age. In Experiment 1, specific staining indicative of the presence of ALV-J gp85 was noted at both 2 and 6 wk of age in the adrenal gland, bursa, gonads, heart, kidney, liver, bone marrow, nerve, pancreas, proventriculus, spleen, and thymus. In Experiment 2, by 10 wk of age, transformed bursal follicles were detected in MGP-stained sections of BF in only one of five (20%) chickens inoculated with ALV J at hatch, compared with five of five (100%) chickens inoculated with RAV-1. The data demonstrate distribution of ALV-J gp85 in various tissues of White Leghorn chickens experimentally inoculated as embryos with the virus. The data also confirm our previous observation that ALV-J is capable of inducing transformation of bursal follicles, albeit the incidence is less frequent than that induced by subgroup A ALV. PMID- 15666877 TI - Virus characterization, clinical presentation, and pathology associated with H7N3 avian influenza in British Columbia broiler breeder chickens in 2004. AB - Low-pathogenicity avian influenza (LPAI) subtype H7N3 was diagnosed on a two-age broiler breeder farm in Abbotsford, British Columbia (BC), in early February 2004. The presenting complaint in the older index flock was feed refusal, with 0.5% mortality over 72 hr that resolved over the following week Ten days after the initial complaint in the index flock, a younger flock in an adjacent barn experienced an abrupt spike in mortality (25% in 48 hr). The gross lesions of tracheal hyperemia and hilar pulmonary consolidation were subtle and nonspecific, and the diagnosis of avian influenza required laboratory confirmation. Two different viruses were isolated from the index farm: a LPAI (H7N3) was isolated from the older flock and a high-pathogenicity avian influenza (HPAI) (H7N3), which had an additional 21 base insertion at the hemagglutinin-cleavage site, was isolated from the younger flock. The presence of this insertion sequence and the similarity of adjacent sequences indicate that the LPAI had mutated into HPAI at some point between the first and second barn. Despite enhanced on-farm biosecurity measures, the virus was not contained on the index farm and eventually spread to over 40 commercial poultry facilities before massive depopulation efforts enabled its eradication. PMID- 15666878 TI - Selenium toxicity in breeding ring-necked pheasants (Phasianus colchicus). AB - A flock of breeding ring-necked pheasants received feed with a high selenium content. Within 4 days of eating the toxic feed, the rate of egg production began to decrease, and bird aggression increased. Approximately 12% of the hens died within a week. Necropsy of the hens revealed colorless fluid around the heart and a friable, but otherwise normal, liver. The rapid onset of the problem and signs noted at necropsy suggested toxicosis. Based on analysis, the feed contained 9.3 ppm of selenium. Selenium toxicity was consistent with the histologic diagnosis of degenerative cardiomyopathy, vacuolar degeneration of hepatocytes, and centrilobular hepatic necrosis. After 8 days, the toxic feed was removed and replaced with fresh feed. Egg production, which had dropped to 50%, returned to normal within 10 days of feed replacement. Hatchability of eggs laid from days 8 to 14 after delivery of the toxic feed was 35%. Approximately 10% of the chicks that hatched had deformed beaks and abnormal eyes. Many of the chicks that died in the shell had deformities, bringing the total to more than 50% of all embryos that developed. The selenium content of eggs that had no embryonic development was 2.05 ppm. Hatchability of eggs laid from days 21 to 28 after the toxic feed was delivered was almost 80%, which was slightly lower than normal. The selenium content of these eggs was 0.30 ppm. These results show the rapid onset and correction of selenium toxicity and suggest that specific embryologic defects are diagnostic for selenium toxicity. PMID- 15666879 TI - An epizootic of subcutaneous tumors associated with subgroup A avian leukosis/sarcoma virus in young layer chickens. AB - An outbreak of subcutaneous tumors in young layer chickens in a flock in Japan was investigated. Tumors appeared as extensive swelling or bulbous protrusions of the integument and were observed in the head or wing of chickens approximately 9 wk old, with a prevalence of 0.4% (157 of 42,000) in the affected flock. Histologically, two types of tumor were observed: myxoma containing abundant hyaluronic acid and neurofibroma with hyperplasia of the Herbst corpuscles. Ultrastructurally, type C retroviruses, such as viral particles, were found in the tumors. The tumors were specifically stained by immunohistochemistry using monoclonal antibodies against the subgroup A avian leukosis/sarcoma virus (ALSV) and yielded a positive reaction to primers specific for subgroup A ALSV by reverse transcriptase-polymerase chain reaction assay. The virus was isolated from the tumors. Seventeen of 20 clinically normal chickens in the affected flock showed antibodies against ALSV. These results suggest that subcutaneous tumors are associated with subgroup A ALSV infection. PMID- 15666880 TI - How to think about stemming an insurgency. PMID- 15666881 TI - Medicine and war. PMID- 15666882 TI - Neuroethics: toward broader discussion. PMID- 15666883 TI - Neuroethics: toward broader discussion. PMID- 15666884 TI - On "humility": the limited effect of disability. PMID- 15666885 TI - Living wills: are durable powers of attorney better? PMID- 15666886 TI - "Demoralization" and decisionmaking: psychiatry again at the forefront. PMID- 15666887 TI - Organs.com: new commercially brokered organ transfers raise questions. PMID- 15666888 TI - Dialysis for a prisoner of war. PMID- 15666889 TI - Medically assisted nutrition and hydration: a contribution to the dialogue. PMID- 15666890 TI - One loss may hide another. PMID- 15666891 TI - Lullabies for Sophia. PMID- 15666892 TI - Bioethics and armed conflict: mapping the moral dimensions of medicine and war. AB - Medical ethics in times of war are fundamentally different from those in times of peace. War brings military and medical values into conflict, often overwhelming other moral obligations, such as a doctor's charge to relieve suffering, in the face of military necessity. PMID- 15666896 TI - Matching patients with therapy. AB - Multiple options are available for the treatment of actinic keratosis (AK). Each therapy has its own idiosyncrasies and risks, and not every modality is appropriate for every patient. Physician experience and clinical factors such as patient demographics, concerns, reason for the office visit, and lifestyle, as well as the pros and cons of each treatment modality, must be considered when developing customized rational treatment programs. PMID- 15666893 TI - Enhancing reflection: an interpersonal exercise in ethics education. AB - There are no moral cookbooks--no algorithms for whipping up moral confections to suit every occasion. But more modest and flexible tools might still be useful for practical ethics. One team describes how professionals can be taught to use a framework for understanding moral problems. PMID- 15666897 TI - Treatment options in the management of actinic keratosis. AB - Successful treatment of actinic keratosis (AK) requires an understanding of available treatments and matching those treatments with each patient's disease severity, lifestyle, insurance coverage, and primary reason for the physician visit. Cryotherapy and other destructive strategies are optimal for hypertrophic and clearly delineated lesions, but they may not address subclinical lesions and are associated with risk of scarring, infection, and pigmentary changes. Topical therapies such as the fluorouracil creams are appropriate for treating large regions of affected skin but are associated with application-site irritation. A combination of destructive and topical approaches may benefit patients with multiple lesion pathologies. PMID- 15666899 TI - Auditory memory and proficiency of second language speaking: a latent variable analysis approach. AB - Previous studies of second language aptitude have mainly used verbal stimuli in memory tasks. Memory for musical stimuli has not been used in aptitude studies although music and language have structural similarity. In this study, 30 Japanese university students who speak English as a second language (19 men, M=21.3 yr., SD=1.8) participated in the experiment as volunteers. They performed verbal memory tasks, musical memory tasks, and English pronunciation tasks. Factor analysis indicated that verbal and musical memory abilities are better represented as a unitary factor rather than two independent factors. Further, a path analysis supported the hypothesis that the memory for both verbal and musical tasks affects proficiency of second language pronunciation, including prosodic features such as stress in word or intonation through a couple of sentences. The memory factor was interpreted as reflecting the performance of "auditory working memory." PMID- 15666898 TI - Fluorouracil 5% and 0.5% creams for the treatment of actinic keratosis: equivalent efficacy with a lower concentration and more convenient dosing schedule. AB - Several formulations and concentrations of topical fluorouracil have received US Food and Drug Administration (FDA) approval for the treatment of actinic keratosis (AK). The most commonly used are the fluorouracil 5% and 0.5% creams. In clinical trials, these formulations have demonstrated a marked ability to partially and completely eradicate AK lesions. Application site irritation, erythema, and burning are common side effects of both formulations, but comparative data suggest that the fluorouracil 0.5% cream is more cost-effective and may be safer, more tolerable, and as efficacious as fluorouracil 5% cream. PMID- 15666900 TI - Development of a cognitive behavioral group intervention programme for patients with multiple sclerosis: an exploratory study. AB - A substantial group of patients with multiple sclerosis (MS) has difficulty coping with their disease. Cognitive behavioral group interventions may help these patients cope more effectively with MS. We developed an 8-session group intervention programme for patients recently diagnosed with MS to help them cope more effectively with MS and to overcome negative thoughts and beliefs about the disease to improve health-related quality of life. We tested the feasibility of the group intervention programme and health-related quality of life in a sample of 11 patients recently diagnosed with MS [mean age: 38 (+/-7.9) yr.; 8 women and 3 men]. All patients were recruited through direct referral by their neurologist or by an MS nurse specialist. The programme was conducted in two small groups of 7 patients each, and each group was led by two psychologists. Cognitive behavioral therapy was an important ingredient in each group session as well as sharing of personal experiences and discussing homework assignments. Each session was formatted the same way but addressed a different MS-specific theme, for example, 'coping with physical impairments' or 'communication with medical staff'. Participants experienced a significant improvement in the health-related quality of life domains of psychological status and vitality, as measured by subscales of Disability and Impact Profile and the Short Form-36 Health Survey. Although further studies are warranted, it appears that a short group intervention programme based on cognitive behavioral techniques for patients with MS might have a positive influence on health-related quality of life. PMID- 15666901 TI - Legalized gambling and crime in Canada. AB - In the 10 provinces and 2 territories of Canada in 2000, but not in 1990, the total number of types of gambling activities was positively associated with rates of robbery (p<.05). Controls for other social variables did not eliminate these associations. With so many correlations in the present study the likelihood of a Type I error was quite large. Alpha was adjusted to control that likelihood. Statistical analysis now required even stronger evidence before concluding that there were significant relationships between crime and gambling variables or among gambling variables. In the 10 provinces of Canada in 1999/2000, the total numbers of electronic gambling machines for each province was associated with rates of theft over $5000 (p<.01). In 1990 there were positive associations found for burglary with off-track betting and race/sportsbooks; motor vehicle theft with off-track betting, and race/sportsbooks; rate of theft with casinos; quarter horse racing with thoroughbred racing. In 2000 there were positive associations for robbery with casinos and slot machines; casinos with slot machines; scratch tickets with raffles, break-open tickets, sports tickets, and charitable bingo; raffles with break-open tickets, sports tickets, and charitable bingo; break-open tickets with sports tickets; charitable bingo with break-open tickets and sports tickets. PMID- 15666902 TI - Patronizing and policing elderly people. AB - 426 students rated a vignette wherein an older person was patronized (more or less) or not by a police officer. Trait attributions were linearly related to extent of patronization: predictably negative for the patronizers, yet positive for the recipients. Visual appearances of patronizers and patronizees did not influence these patterns. PMID- 15666903 TI - Failure of a traffic control "fatality" sign to affect pedestrians' and motorists' behavior. AB - The behavior of 643 pedestrians and 1749 motorists at two signal-controlled intersections was observed over a 2-mo. period of observation in March and November. One intersection had a coffin-shaped traffic sign with the wording "Fatality" erected during the first month of observation (March). The second intersection was identical in traffic and pedestrian volume but lacked a "Fatality" sign in either March or November. "Fatality" signs are erected and maintained by the City of Edmonton for 6 mo. whenever a pedestrian death has occurred, and they are intended to raise the awareness of both pedestrians and motorists concerning the risks at dangerous intersections. This sign had been removed for nearly 3 mo. when a follow-up observation was made in November. While female pedestrians and pedestrians over the age of 50 years showed greater caution at both intersections, the presence or absence of the "Fatality" sign had no statistically significant influence on safety. Similarly, the presence or absence of a sign did not significantly influence motorists' behavior. Of motorists 7.6% ran either amber or red traffic control lights at the two intersections. The "Fatality" sign did not affect the rates of these violations. PMID- 15666904 TI - Relations of sex, age, perceived fitness, and aerobic activity with social physique anxiety in adults sixty years and older. AB - Self-reported physical activity, and perceived fitness were examined to assess their effects on social physique anxiety in a sample of older individuals (N= 249; M age=70.4 yr., SD=8.2). Participants reported their fitness as "average" to "above average." There were no significant effects of perceived fitness or age on social physique anxiety scores. The women had significantly higher social physique anxiety scores than the men. PMID- 15666905 TI - A longitudinal study: some preliminary results of association of prenatal maternal stress and fetal movements, temperament factors in early childhood and behavior at age 2 years. AB - Niederhofer (1994) found that maternal stress during pregnancy was significantly associated with development of personality in early childhood. This study examined the correlation between maternal stress (self-report) during pregnancy for 22 women, its ultrasound objectification by observation of intrauterine fetal movements, child's temperament in early childhood, and child's Independence and social behavior at the age of 2 years while controlling for possible confounding variables. Only intrauterine fetal movements (head/arm/leg) were not associated with stress during pregnancy, temperament, or independence. PMID- 15666906 TI - How is stigmatization affected by the "layering" of stigmatized conditions, such as serious mental illness and HIV? AB - Vignettes were used to examine the effect of labeling a person with two stigmatized illnesses, HIV disease and serious mental illness (schizophrenia). The additive model predicted that stigma associated with combined HIV and serious mental illness would resemble the simple sum of those for the two conditions. The discounting model predicted that the presence of serious mental illness would lead subjects to view the target individual as less responsible for infection, resulting in less stigmatization than given for HIV alone. Data collected from 244 participants at a public northeastern university supported the additive model. Stigmatization was highest in the group labeled with both HIV and serious mental illness, while stigmatization associated with someone with only HIV was less than those associated with someone with only serious mental illness. Possible explanations are discussed. PMID- 15666907 TI - Emotional regulation strategies and negotiation. AB - This study examined the relationship between profit achievement and emotional regulation strategies, using Kelley's Negotiation Game to measure profit achievement. The game involves bargaining for the prices of three products. Emotional Regulation Strategies were measured by The Emotional Regulation Questionnaire. Scores were obtained from 104 lower level managers of a bank in Turkey. Their average age was 32.0 yr. (SD=3.7), (39 women and 65 men). A correlation of .65 (p<.01) was obtained between scores on profit achievement with scores on Cognitive Reappraisal strategy and -.50 (p<.01) with scores on Suppression strategy. PMID- 15666908 TI - English letter frequencies and their applications: part II--digraph frequencies. AB - This article continues the presentation of new data regarding the frequencies of English letters organized by word length and letter position. Digraphs (defined here as all two-letter combinations) were the objects of study. The frequencies of digraphs were derived from a sample of 320,780 English words (including 6505 different words), which were credibly demonstrated by Whissell to be a parsimonious representation of modern English word usage. A total of 997,380 digraphs were counted and sorted by locations according to word length and digraph positions within words. As assessed by the Whissell source, the data about digraphs presented accurately represent the frequencies with which digraphs occur in modern English. How these data can provide a resource for reading research and practice is explored. PMID- 15666909 TI - Relations between big five traits and fundamental motives. AB - Relations were examined between configurations of Big Five Traits (Extraversion, Agreeableness, Conscientiousness, Neuroticism, Openness to Experience) and 16 fundamental motives (Social Contact, Curiosity, Honor, Power, Order, Idealism, Independence, Status, Vengeance, Romance, Family, Activity, Saving, Acceptance, Eating, Tranquility) in 138 university students (93 women, 45 men; M age= 20.3 yr., SD=4.5). Big Five traits were measured with the NEO-PI-R and motives were measured with the Reiss Profile of Fundamental Goals and Motivation Sensitivities. The traits were significantly related with all the motives (adjusted R2=.06 to .43) except Physical Activity. Four motives were related with only one trait and nine configurations of two or more traits were correlated with the remaining 11 motives. Total motive scores across all participants, an index of the strength of overall motivation, were positively correlated with Extraversion and Neuroticism and negatively with Agreeableness. PMID- 15666910 TI - A laboratory study of satisfaction effects on mood state, withdrawal intentions, and organizational citizenship behavior. AB - 154 undergraduates role-played a restaurant server experiencing positive, neutral, or negative Job or Life Satisfaction. Positive Satisfaction resulted in ratings of more positive reported mood state, lower expected absenteeism and turnover intentions, and greater expected Organizational Citizenship Behavior than did Neutral Satisfaction and Negative Satisfaction. Life Satisfaction was associated with reported mood states and intentions similarly to Job Satisfaction, except Negative Job Satisfaction was more negatively influential on expectations of quitting than Negative Life Satisfaction. Both Job and Life Satisfaction may be antecedents to affect and its consequences in work settings. PMID- 15666911 TI - Clinical correlates of insight in schizophrenia. AB - 111 schizophrenic patients diagnosed in accordance with DSM-III were rated on Landmark's symptom checklist, on demographic variables, and on variables descriptive of the course of illness. Of the 111 patients, 108 (97.3%) showed poor insight into their illness at some time in the past and 65 (58.6%) at the time of assessment. Those presently showing poor insight were significantly (Pearson rs, p< .01, 2-tailed) more frequently rated as currently displaying poor judgement also in other matters (r =.50), as showing social withdrawal (r=.42) and poor rapport (r=.33), and as being preoccupied with their delusions or hallucinations (r=.31) and as being unreliable informants (r=.41). They usually had lower education (r=.33), their income in the last taxation year was lower (r = .47), and their work functioning was less adequate (r = .30). PMID- 15666912 TI - Obsessiveness and a thematic apperception test-based measure of aggression. AB - Freud (1909/1955) hypothesized a conflict between love and hate in obsessive neurosis. To test this relationship, we compared a Thematic Apperception Test based measure of aggressive fantasies in college men who scored either high or low on the Obsessive-Compulsive Inventory-Revised. 64 undergraduate men from beginning classes in psychology participated. Their mean age was 19.4 yr. (SD= 1.7). 16 men with high scores had significantly higher scores on a TAT-based measure of aggressive fantasies toward parents, partners, and others than 15 men with low scores, which is consistent with Freud's hypothesis. PMID- 15666913 TI - Did the defeat of Saddam Hussein reduce suicide bombing casualties and attacks in Israel? A statistical analysis. AB - It was hypothesized that one of the reasons for the U.S. invasion of Iraq was to stop the payments being made by Saddam Hussein to the families of suicide (homicide) bombers in Israel. The consequences of suicide (homicide) bombing attacks against Israel between March 2001 and August 2004 were evaluated as related to the time of the U.S. invasion of Iraq. On average per month during this period, there were fewer overall casualties after the invasion than before it. As many as nearly 1,100 casualties may have been prevented in Israel as a consequence of the U.S. invasion of Iraq, suggesting that at least one possible goal of the U.S. invasion may have been achieved, at least as averaged over the first 17 months after the invasion. PMID- 15666914 TI - Challenge for studying metacognitive experiences: a rejoinder to Akama and Yamauchi. AB - This commentary focuses on three points of the Akama and Yamauchi paper (2004) that raise possibilities for research. These points regard (a) the nature of metacognitive experiences, (b) their relations with performance, and (c) their role in self-regulation. PMID- 15666915 TI - Comparing professional schools and traditional clinical program faculty on measures of professional and scientific achievement. AB - 474 faculty in 56 professional clinical psychology programs were compared with 972 faculty from 157 traditional clinical psychology programs. The faculty in the professional programs were more likely to be licensed (93% to 85%) and to be diplomates of the American Board of Professional Psychology (15% to 11%, respectively). The faculty in traditional programs averaged more publications (35 vs 17) and citations, and they were more likely to be fellows of the American Psychological Association (26% to 11%). These differences appear congruent with the different missions and objectives of the two types of programs. There were no differences in the number of years since the doctorate, used as a measure of professional experience. PMID- 15666916 TI - The irrational beliefs inventory: cross-cultural comparisons between South African and previously published Dutch and American samples. AB - The Irrational Beliefs Inventory gives a measure of irrational beliefs, as postulated by Ellis's Rational Emotive Behavior therapy. Given the increasing cross-cultural use of psychometric scales, it is important to assess whether the psychometric properties of the inventory are consistent across cultures. In the present study cross-cultural applicability, in terms of internal consistency and independence of subscales, was investigated for an ad hoc sample of White (n= 100, M age = 21.3 yr., SD=4.0) and Black (n=82, M age=19.8 yr., SD=2.2) undergraduate South African university students. Cronbach coefficients alpha for the subscales and Pearson correlations between subscales for American and Dutch students, as reported by Bridges and Sanderman, were compared with those indices for the South African students. The magnitude and rank order of Cronbach alpha, as well as the correlations between subscales for the three groups showed strong similarities. Values of alpha for the Black South African students were lower in magnitude on all subscales than those for American, Dutch, and White South African samples, but intercorrelations between subscale scores were consistent. Findings in the present study are supportive of the cross-cultural applicability of the Irrational Beliefs Inventory to White South African students but not to South African Black students. PMID- 15666917 TI - The increasing English homicide rate. AB - In England and Wales from 1980 to 1987, the increasing homicide rate did not appear to be associated with an increase in the proportion of 15- to 24-yr.-olds. PMID- 15666918 TI - Relations between family structure and students' health-related attitudes and behaviors. AB - This study examined the relations between family structure and students' health related behaviors. Participants were 6,130 Greek students, ages 11 to 16 years. They responded to questionnaires based on the Planned Behavior model assessing attitudes, perceived behavioral control, intentions and behavior regarding healthy and unhealthy habits. Students who reported growing up with one or no parent reported more unhealthy lifestyle attitudes and behaviors (smoking, drug use, violence, exercise, nutrition) than students growing up with both parents. The findings suggest that family structure is related to students' healthy or unhealthy lifestyle, indicating that in health education programs all members of the close environment within which the children live should be involved, especially for children who do not live with both parents. PMID- 15666919 TI - Documentation of ethnicity. AB - The comparison of the ethnic composition of an intermediate care facility with several Hispanic residents and the general population was hindered by the absence of categorization of ethnicity according to the United States Census. If all Hispanic residents of the facility were white, then 55% of the facility population were white, a proportion comparable to the 58.2% white population of the general population. On the other hand, if all the Hispanic residents were not white, then 27.5% of the facility residents were white. In that case, the proportion of white residents of the facility is much less than in the general population. Therefore, a Demographic Coding Form was developed to capture the essential data to make direct comparisons and contrasts with the general population recorded by the United States Census. Since the United States Census records Hispanic ethnic minority status as a separate category independent from all other ethnic groups, the design of experiments to investigate the possible effects of ethnicity on populations wisely incorporates the administration of a Demographic Coding Form to capture the key ethnic data to permit direct comparison with the general population. PMID- 15666920 TI - Sex differences in attitudes toward computers and the internet. AB - A study of 137 undergraduate students yielded no sex differences in attitudes toward computers or the Internet. PMID- 15666921 TI - Did Freud mislead patients to confabulate memories of abuse? A reply to Gleaves and Hernandez (1999). AB - Gleaves and Hernandez have argued that skepticism about the validity of Freud's seduction theory, including by Powell and Boer, is largely unjustified. This paper contends that their analysis is in many ways both inaccurate and misleading. For example, we did not, as they implied, reject the possibility that some of Freud's early patients were victims of childhood sexual abuse. We also maintain that the weight of the available evidence indicates that false memories of traumatic events probably can be implanted, and that Freud's (1896/1962a) original evidence for the validity of his patients' recovered memories remains lacking in several respects-particularly in view of the extremely suggestive procedures he often used to elicit such memories. PMID- 15666922 TI - Buying online: textbooks versus books. AB - Predictors of purchasing textbooks and other types of books by college and university students were found to be different. PMID- 15666923 TI - Reaction to "Comments on 'The effects of leader-member exchange and differential treatment on work unit commitment': distinguishing between neutralizing and moderating effects". AB - Ye and Barbuto in 2004 made a number of interesting observations about use in 2002 by Van Breukelen, Konst, and Van der Vlist of the term 'neutralizer'. They advised caution when employing the term 'neutralizer', preferring instead the designation 'moderator'. Their comments underscore the importance of discussion on the nature of interaction effects in studies of leadership. Here we clarify our procedures in more detail and present the results of further analyses of our data. PMID- 15666924 TI - Autonomy and job satisfaction for a sample of Greek teachers. AB - Analysing the relation between Job Satisfaction and Autonomy in a sample of 300 Greek teachers (114 men and 186 women, 28 to 59 years old) from primary and secondary schools, showed statistically significant positive correlations between Job Satisfaction and Autonomy. Particularly, Autonomy was correlated with Job Itself (.21), Supervision (.22), and the Organizational as a Whole (.27), aspects of Job Satisfaction. Findings are in line with previous studies conducted in different cultural contexts. Percent common variance accounted for is small. PMID- 15666925 TI - A phenomenological and perceptual research methodology for understanding hypnotic experiencing. AB - Phenomenology and perceptual psychology opens up the essential meanings of hypnosis by presenting a qualitative method as an alternative to the current predominant quantitative method in the study of hypnosis. Scales that measure susceptibility from behavioral and cognitive aspects abound in the hypnosis literature, but understanding the structure of hypnotic experiencing is yet to come. A new qualitative approach to researching hypnotic experiencing by combining aspects of phenomenological research as in work of Giorgi, Moustakas, and Wertz, familiarity with Husserl's philosophy, and a perceptual psychological research method (cf. work by Combs, Richards, & Richards and by Wasicsko). The author utilized this combined methodology to formulate the theory of Perceptually Oriented Hypnosis. This methodology enables the therapist or professional and patient or client to share benefits from the effects of their hypnotic experiencing in its intersubjective sense. This method can be applied in numerous life situations such as teaching and therapy in addition to the experimental situation. PMID- 15666926 TI - Suicide and homicide in rural areas of California. AB - Analysis of 1989-1998 age-adjusted death rates from personal violence in California did not show significantly different suicide and homicide rates in rural counties. No urban vs rural differences in rates of personal violence were noted; however, numerically there were more rural than urban suicides and homicides. PMID- 15666927 TI - On feeling negative past as a part of current self: subjective temporal organization of autobiographical memories. AB - When a past event is remembered, the subjective elapsed time is often different from the objective elapsed time, even if the exact date of an event is known. In this study, Feeling of Time Discrepancy between objective elapsed time and subjective elapsed time of turning point events was examined. Participants remembered one turning point event in life, dated the event, and estimated the Feeling of Time Discrepancy. Two groups, formed on their judgement of whether their present understanding of the event is the same or different from understanding when the event occurred (the Same assessment group and the Different assessment group). Analysis showed the Different assessment group felt the turning point event to be more recent than the Same assessment group did, although there was no difference between actual elapsed time and memory vividness for the two groups. In addition, the events which changed from negative to positive were felt subjectively to be more recent than those for which affect was consistently positive. These results suggest that, even if we experience negative events, the event might become a part of the present self and be felt temporally closer to now, when temporal organization of autobiographical memories allow recognition of the event as good experience. Finally, two types of temporal organization of such memory were discussed, the subjective temporal organization of autobiographical memory which is organized only for oneself and the social objective temporal organization of autobiographical memory which becomes organized in sharing it with another person. PMID- 15666928 TI - Reliabilities of the WAIS-III for discrepancy scores: generalization to a clinical sample. AB - Internal consistency reliabilities for WAIS-III IQ, Index, and subtest discrepancy scores are provided for 100 men in a treatment program for substance abuse disorders. There were 63 Euro-Americans and 37 African Americans. Means for age, education, and Full Scale IQ were 46.1 yr. (SD=8.8), 12.7 yr. (SD=1.5), and 93.8 (SD=14.0), respectively. Reliabilities ranged from .34 to .86. Compared with the WAIS-III standardization sample, discrepancy score reliability coefficients differed only for the Verbal Comprehension Index vs Perceptual Organization Index contrast, which was higher in the patient sample. Interpretation of discrepancy scores for the WAIS-III appears to be useful in a clinical sample with substance abuse as in a purportedly normal sample. PMID- 15666929 TI - Enterprise failures correlate positively with suicide rate for both sexes in Japan. AB - Correlations among suicide rate, unemployment rate, and enterprise failure were examined from 1976 to 1994 by using a Loess smoothing method. Significant positive correlations were found between suicide rate in men and unemployment and between suicide rate in both sexes and enterprise failure (p < .001). PMID- 15666930 TI - Self-control, deviant peers, and software piracy. AB - Tests of self-control theory have examined a substantial number of criminal behaviors, but no study has examined the correlation of low self-control with software piracy. Using data collected from 302 students in this university, this study examined the correlation of low self-control with software piracy and the moderating role of associating with deviant peers in this correlation. Low self control correlated with software piracy more strongly for those who had high associations with deviant peers than for students with low associations with deviant peers. Analysis indicated differential links for lack of moral attitude in relation to software piracy and favorable attitudes for software piracy for varying association with deviant peers. PMID- 15666931 TI - Depression, optimism, and positive health practices in young adolescents. AB - Correlations among depression, optimism, and positive health practices were studied in young adolescents. A sample of 131 boys and girls, aged 12 to 14 (M=13.2, SD=.8) completed the Revised Personal Lifestyle Questionnaire, the Center for Epidemiological Studies-Depression Scale, and the Life Orientation Test. A negative correlation (-.47) was found between scores on Depression and Positive Health Practices, and the correlation for scores on Optimism and Positive Health Practices was .52. PMID- 15666932 TI - Evidence for aging as the cause of Alzheimer's disease. AB - Part 1 presents the results of a meta-analytic study on the effects of aging on intelligence. Analysis of a total of 20 longitudinal samples shows that most of the intelligence scores rose before the age of 50 and fell at a progressively increasing rate after the age of 50. An equation describing this rise and fall in intelligence was derived. Part 2 shows the relationship between the predicted prevalence of Alzheimer's disease (from the equation derived in Part 1) and the prevalence of the disease obtained from 10 studies. The predictive curve fit so well with the observed prevalence data that the results can be interpreted as evidence that Alzheimer's is a manifestation of aging. PMID- 15666933 TI - Use of paradoxical intention in the context of acceptance and commitment therapy. AB - The purpose of this paper is to describe the use of therapeutic paradox within the contextual-behavioral treatment called Acceptance and Commitment Therapy. Emphasis is on the difference between inherent and constructed paradox, expanding on previous work and emphasizing Acceptance and Commitment Therapy's relationship to Logotherapy. PMID- 15666934 TI - Theories of intelligence in children with reading disabilities: a training proposal. AB - A recent trend in the study of reading difficulties promotes multidimensional intervention, focusing on the reciprocal influences exerted by cognitive and emotional-motivational variables. This study evaluated improvements in reading performance as a function of metacognitive training in 36 children (M age = 8.7 yr.) with different representations of intelligence. Posttest evaluations show significantly more improvement in reading comprehension in children with an incremental theory of intelligence. These results indicate the importance of treatment programmes that take into account both the specificity of deficits and factors relating to the domain of motivation. PMID- 15666935 TI - Smoking and depressive symptoms among children ages 11 to 16 years. AB - Relationships between cigarette smoking and depressive symptoms were analysed in a representative random sample of 1447 secondary-school children aged 11 to 16 years in Galicia (NW Spain). The sample comprised 797 boys (55.1%) and 650 girls (44.9%). Their mean age was 12.8 yr. (SD=1.2). Depressive symptoms, evaluated with the Children's Depression Inventory, were reported by 11.0% of children who responded they had never smoked, versus 23.7% of those who said they sometimes smoked, and 44.2% of those who identified themselves as current daily smokers. These results indicate an association between smoking and depression among children in this age group. PMID- 15666936 TI - A measure of parents' assessments of their children's abilities. AB - The present study assessed the psychometric properties of a multidimensional measure of parents' assessments of their children's abilities, including problem solving skills, social skills, dexterity, creativity, and learning motivation. A nationwide sample of Finnish parents (N=432), representing two educational groups, both mothers (64%) and fathers (36%) whose mean age was 37.8 yr. (SD=5.3), were asked to assess their children's abilities during preschool and at the end of the first and the third school year. A set of analyses of this longitudinal data indicated that the scales had adequate internal and temporal consistency, the factor structure showed invariability across time, and the scales related meaningfully to the parents' perceptions of their children's competencies in different school subjects. PMID- 15666937 TI - Sex differences in severity of suicidal behavior in an Iranian sample. AB - A nonsignificant tendency was found for 48 females in Iran to make more serious suicide attempts using opium than did 152 males. PMID- 15666938 TI - Attitudes toward extrarelational sex as a function of participant's and third party's gender. AB - An experiment was conducted to examine whether attitudes toward extrarelational sex, i.e., "swinging," differed as a function of participant's gender and gender of the third party, i.e., the "swinging" partner. Participants were asked to imagine that their current romantic partner had expressed an interest in "swinging" with another individual (male or female, randomly assigned). Analysis yielded several significant differences by participants' gender. Specifically, men expressed greater interest than did women in joining a swinger's club, reported a higher likelihood than did women of actually joining such a club, and believed more than women that their sex life with their partner would improve after joining a swinger's club. Participants also preferred a female more than a male swinging partner, although this comparison was not statistically significant. PMID- 15666940 TI - Moderation of automatic achievement goals by conscious monitoring. AB - The aim of this study was to test the assumption that nonconscious goal pursuit guides behavior more efficiently and produces better performance, only when the conscious monitoring of the task is low. Some studies have documented that goals can be activated and guide behaviors outside of awareness. 66 students, 22 men and 44 women with a mean age of 18.3 yr. (SD=0.9) years, were randomly assigned to Achievement Goal Priming or Neutral Priming conditions. In the Achievement Goal Priming condition, concepts associated with high performance were activated outside of their awareness via a scrambled sentence task. All participants were then given a calculation task to complete. To manipulate conscious monitoring, half of the participants were presented the task as a fun filler task, and the rest were presented the task as a measure of ability and were encouraged to monitor their performance on the task consciously. Analysis showed, among participants who were presented the task as a fun filler task, the Achievement Goal Priming group completed more tasks relative to the Neutral Priming group. Those who were encouraged to monitor their performance consciously did not benefit from Achievement Goal priming. The results implied that consciously monitored behavior might be counterproductive. PMID- 15666939 TI - Exploring relationships between college students' learning styles and motivation. AB - A pilot study explored the relationships between learning styles and motivation for a sample of 103 undergraduate and graduate students who completed both the Approaches to Studying Inventory and the Motivation Sources Inventory online. A significant positive but small correlation was found between scores on Surface Apathetic Approach and on Self-concept Internal motivation (r =.32, p< .01). Significant negative correlations were found between scores on Surface Apathetic Approach and on Self-concept External motivation (r = -.32, p < .01), and for Strategic Approach and Self-concept Internal (r =-.37, p<.01). Implications and further research are discussed. PMID- 15666941 TI - A study orientation questionnaire in mathematics for use in a tertiary environment. AB - The Study Orientation Questionnaire in Mathematics Tertiary is being developed as a diagnostic measure for South African lecturers and counselors to help students improve their orientation toward the study of mathematics. In this pilot study subjects were freshman black students registered for the Five-year Study Programme in the School of Engineering at the University of Pretoria. During the first phase of the standardisation in 2000, 33 students (28 men; 5 women, M age=19.6, SD=.9) were assessed, and in 2001, 40 students (27 men, 13 women, M age=19.2, SD=.9). In 2002 the questionnaire was administered to 51 students (39 men, 12 women, M age= 19.1, SD= .9). Analysis showed satisfactory reliability coefficients and item-field correlations. Step-wise linear regression in 2001 indicated that a combination of two fields, namely, Information Processing and Problem-solving Behaviour in mathematics, contributed significantly (R2=46.9%) to predicting achievement in calculus. In 2002 one field, Mathematics Confidence, contributed significantly (R2= 25.2%) to that prediction. PMID- 15666942 TI - Association between neuroticism and introversion. AB - In three samples of subjects (ns=67, 115, 77) introversion and neuroticism scores on one of Eysenck's tests were positively associated. PMID- 15666943 TI - Marital satisfaction and relational attachment in a sample of newly married couples. AB - The purpose was to investigate possible correlates [perception of parental marital satisfaction (Relationship Assessment Scale), marital status of parents, participation in marital enrichment, current stress, and sex] of marital satisfaction (Kansas Marital Satisfaction Scale) and relational attachment (Revised Adult Attachment Scale). 71 young couples (M age=25.3 yr., SD=3.2) in their first ten years of marriage completed the measures. Backward regression analysis indicated that marital status of parents, participation in marital enrichment, current stress, and sex accounted for a significant portion of the variance for the Anxiety subscale of the Revised Adult Attachment Scale. None of the models significantly accounted for the variance on the measure of marital satisfaction. PMID- 15666944 TI - Assigning blame in British and American students. AB - British university students (n = 142) obtained a higher mean external locus of control score than did American university students (n = 185) overall and, in particular, on items related to politicians and government. PMID- 15666945 TI - Faith maturity and doctrinal orthodoxy: a validity study of the faith maturity scale. AB - This study investigated the factor structure of the 12-item Faith Maturity Scale and the association of Protestant Faith Maturity and Doctrinal Orthodoxy. The sample was taken from 207 urban evangelical church members in southern California. Analysis indicated the Faith Maturity Scale incorporates two factors and Faith Maturity has a weak, positive association with Doctrinal Orthodoxy as tapped by the Christian Doctrinal Orthodoxy Scale. Protestant faith maturity seems to be more independent of doctrinal orthodoxy than generally thought. PMID- 15666946 TI - Religion and perceived stress among undergraduates during fall 2001 final examinations. AB - To examine the relationship of religion and perceived stress, 97 undergraduate college students responded to the Perceived Stress Scale, the Spiritual Well being Scale, and the Intrinsic/Extrinsic-Revised scale during a period of extreme national stressors during Fall 2001, namely, the September 11th terrorist attacks, anthrax scare, and war in Afghanistan, in addition to the local stressor of pending final college examinations. Scores on the Perceived Stress Scale were negatively correlated to scores on Existential Well-being (-.66), Religious Well being (-.43), and Intrinsic Religious Orientation (-.44). Age was unrelated to all variables. Both the quality (well-being) of students' religious experience and the orientation of that experience were related to their perception of stress. PMID- 15666947 TI - Depression and intrinsic religiosity. AB - A sample of 895 adults completed the Center for Epidemiological Studies Depression scale and a test of intrinsic religious motivation. The correlation of .06 between scores on the two scales was not significant. PMID- 15666948 TI - Validation of the lie/bet screen for pathological gambling on two normal population data sets. AB - The validity of the Lie/Bet Screen was tested on two community population samples, one adult (n=2,014) and one adolescent sample (n=3,237), in Norway. With positive responses on at least one of the questions on Lie/Bet Screen used as the cutoff point the screen showed high both sensitivity and specificity. The negative predictive value was also high, but the positive predictive value was comparatively lower. A prediction of probable pathological gambling or "At-risk gambling" based on both Lie/Bet questions identified a valid screening in the two samples (0.54% in adults, 5.6% in adolescents). Compared to the use of the full DSM-IV this is pretty close, with the figures 0.45% and 5.22%. It is concluded that the Lie/Bet Screen may function as a good screening device for pathological gambling plus At-risk gambling in normal community samples. PMID- 15666949 TI - Correlation of national internet use with extraversion and neuroticism. AB - Using data from 23 nations, scores on a measure of Extraversion by nation were positively associated with more Internet use per capita. PMID- 15666950 TI - Application and revision of the Kansas marital satisfaction scale for use with Korean couples. AB - To apply the Kansas Marital Satisfaction Scale to Korean couples, one item about the parenting role was added to the original scale. Using survey data from 350 married couples from the urban area of Seoul, Korea, this study investigated the availability of the original and revised forms in Korea. Values of internal consistency of responding, means, standard deviations and Pearson rs for both scales were remarkably consistent in the Korean sample and were good assessments of key dimensions of the marital process of Korean couples with children. PMID- 15666951 TI - Reliability of single-item ratings of quality in higher education: a replication. AB - Single-item ratings of the quality of instructors or subjects are widely used by higher education institutions, yet such ratings are commonly assumed to have inadequate psychometric properties. Recent research has demonstrated that reliability of such ratings can indeed be estimated, using either the correction for attenuation formula or factor analytic methods. This study replicates prior research on the reliability of single-item ratings of quality of instruction, using a different, more student-focussed approach to teaching and learning evaluation than used by previous researchers. Class average data from 1,097 classes, representing responses from 59,815 students, were analysed. At the "class" level of analysis, both methods of estimation suggested the single item of quality had high reliability: .96 using the correction for attenuation formula, and .94 using the factor analytic method. An alternative method of calculating reliability, which takes into account the hierarchical nature of the data, likewise suggested high estimated reliability (.92) of the single-item rating. These results indicate the suitability of the overall class rating for quality improvement in higher education, with a large sample. PMID- 15666952 TI - Convergent validity of the Larocque Obesity Questionnaire and self-reported behavior during obesity treatment. AB - A validation study of the Larocque Obesity Questionnaire designed for use in obesity treatment was performed. Unlike other measures of obesity, this questionnaire includes scales measuring general emotional state as well as eating behavior. Subscales measured uncontrolled eating, physical stress responses, depression, and perfectionism. Subjects were 458 women and 79 men in treatment for obesity by general practitioners. The subscales showed acceptable internal consistency and related in predictable ways to measures of eating behavior, depression, self-criticism, stress, physical complaints and weight-control motivation. Subjects in the heaviest weight category (Body Mass Index > or = 40) showed higher scores on Stress Response and Depression subscales. There were no significant sex differences, after controlling for weight. All four subscales showed significant improvement after 5 wk., which indicates their sensitivity to changes during treatment. PMID- 15666953 TI - Using the range to calculate the coefficient of variation. AB - In this research a coefficient of variation (CVhigh-low) is calculated from the highest and lowest values in a set of data. Use of CVhigh-low when the population is normal, leptokurtic, and skewed is discussed. The statistic is the most effective when sampling from the normal distribution. With the leptokurtic distributions, CVhigh-low works well for comparing the relative variability between two or more distributions but does not provide a very "good" point estimate of the population coefficient of variation. With skewed distributions CVhigh-low works well in identifying which data set has the more relative variation but does not specify how much difference there is in the variation. It also does not provide a "good" point estimate. PMID- 15666954 TI - Cross-cultural examination of social interactions during a one-week dousa-hou (Japanese psychorehabilitation) camp. AB - Dousa-hou is a Japanese psychological rehabilitation method which is widely used in Japan for children with mental retardation, cerebral palsy, and autism. The primary focus of the rehabilitation method is to improve bodily movements and posture as well as to introduce social support to patients and their first-degree relatives. Analysis showed mothers got more social support interacting with their child's trainer and supervisor during Dousa-hou. Trainers were more interactive than mothers in the Indian group, followed by the Japanese and Korean cultural groups. PMID- 15666955 TI - Effect of the September 11, 2001 terrorist attacks on death anxiety in university students. AB - A sample of 440 undergraduate university students completed the Templer Death Anxiety Scale 2 wk. prior to and 2 wk. after the September 11, 2001 terrorist attacks. Women comprised 66% of the sample, and 79% of the sample identified themselves as 18 to 21 years of age and either freshmen or sophomores. There was no significant mean difference in the pre- and postterrorist attack Death Anxiety scores. Differences were found on two individual scale items. PMID- 15666956 TI - Long-term effect of social relationships on mortality in patients with congestive heart failure. AB - Congestive heart failure is a major cause of morbidity and mortality within the Western world, and yet psychosocial impact of this disease is under-researched. The aim of this study was to evaluate the possible effects of social relationships (perceived social support and perceived social isolation) on mortality risk in 119 patients with stable, symptomatic congestive heart failure. Fifty-one deaths were registered during the six-year follow-up period, all from cardiac causes. Analysis using proportional hazard models indicated that social isolation was a significant predictor of mortality (relative risk, 1.36; confidence interval, 1.04 to 1.78; p < 0.03), controlling for neuroticism, heart failure severity, functional status, gender, and age. The small sample size was a limitation of the study; therefore, further research is required in order to confirm these findings and to illuminate the mechanisms behind the relationships between social isolation and mortality. PMID- 15666957 TI - Mental health treatment preferences of older and younger primary care patients. AB - OBJECTIVE: To compare mental health treatment history and preferences in older and younger primary care patients. METHOD: We surveyed 77 older (60+) and 312 younger adult primary care patients from four outpatient medical clinics about their mental health treatment history and preferences. RESULTS: Older adults were less likely than younger adults to report a history of mental health treatment (29% vs. 51%) or to be currently receiving treatment (11% vs. 23%). They were also less likely to indicate that they currently desire help with emotional problems (25% vs. 50%). Older adults were more likely to hold a belief in self reliance that could limit their willingness to accept treatment for mental health problems, although they were less likely than younger adults to identify other barriers to treatment. Older adults reported that they were less likely to attend programs in primary care targeting mental health issues (counseling, stress management) than younger adults, although they were as willing as younger adults to attend programs targeting physical health issues (healthy living class, fitness program). Age remained a significant predictor of mental health treatment history and preferences even after controlling for other demographic variables. CONCLUSION: These results suggest that older adults in the primary care setting may be less willing to accept mental health services than younger adults. Results further suggest that perceived barriers may differ for older and younger patients, which may indicate the need for age-specific educational messages and services targeted to older adults in primary care. PMID- 15666958 TI - Hispanic ethnicity, language, and depression: physician-patient communication and patient use of alternative treatments. AB - OBJECTIVE: To examine the relationship between Hispanic ethnicity and language spoken with physician communication about depression and patient use of alternative treatments for depression. METHOD: This is a secondary data analysis from a trial of depression screening conducted in four primary care clinics. Patients with Hispanic or non-Hispanic White ethnic backgrounds and those meeting DSM-III-R criteria for current major depression, minor depression, dysthymia as well as those that screened positive on a depression screening instrument (n = 141) are included in this analysis. We labeled those who screened positive for depression but did not meet DSM-IIIR criteria for a current depressive disorder as "distressed." Clinicians' use of counseling and patient use of alternative treatments were based on patient self-report. RESULTS: Forty-two percent (n = 59) of the sample stated that their physician had either told them that they had depression, treated them for depression, or recommended that they seek help for depression. Over 40% of patients spent time talking with their physicians at their current visit about what was making them depressed and 34% received counseling about depression. Hispanic ethnicity and language were not significantly related to physician communication with patients about how to overcome depression. Thirty-six percent of patients reported talking with a minister or other religious person about feelings of depression or sadness. Seventeen percent of patients had used herbal remedies or non-prescription medications and 5% had seen a curandero for feelings of depression or sadness. Hispanic ethnicity and language were not significantly related to patient use of alternative treatments for depression. CONCLUSIONS: Hispanic ethnicity and language were not significantly related to physician-patient communication about depression or patient use of alternative treatments for depression. Physicians should make sure to ask patients about all of the types of things they are doing to help overcome their depression. PMID- 15666959 TI - Who benefits from more structured depression treatment? AB - OBJECTIVE: This article uses data from two studies that have demonstrated the overall effectiveness of Collaborative Care interventions to evaluate factors associated with poor outcomes overall (general prognostic factors) and factors associated with greater or lesser effects of treatment (differential treatment effects). METHODS: Adult primary care patients initiating antidepressant treatment for major depression were randomized to usual care or to Collaborative Care, a structured depression treatment program that included planned, proactive and coordinated care with a health care team and informed, activated patients (n = 156, mean age = 43, 85% white). Response to treatment was defined as a 50% or greater reduction in depression at four months. RESULTS: High neuroticism and a history of recurrent major depression (3+ episodes) or dysthymia predicted poor outcomes in general. While the magnitude of the intervention effects differed, frequently cited predictors of persistence of depression (age, gender, depression severity, medical and psychiatric comorbidity) were not significantly associated with greater or lesser benefit from Collaborative Care (no differential treatment effects). Results demonstrate the robustness of intervention effects across numerous groups at risk for persistence of depression. CONCLUSIONS: These findings suggest that at the time of diagnosis it is not possible to predict who is most likely to benefit from Collaborative Care. Instead, outcomes of treatment should be routinely monitored among depressed patients to ensure optimal response. PMID- 15666960 TI - Quetiapine in the successful treatment of psychosis and comorbid brittle diabetes mellitus: a case report. AB - BACKGROUND: As evidence of a biologic determinant of schizophrenia has been elaborated, an interest in the relationship between schizophrenia and autoimmune disorders has become increasingly more developed over the last decade. Pedigree analysis has shown that schizophrenia, like autoimmune disorders, is likely a heritable phenomenon, and a genetic liability in this disorder is hardly disputed. Research has indicated that physiologic connections between IFN-gamma and TNF-alpha are suggestive of a connection between the symptoms associated with schizophrenia and those of hypoglycemic events in IDDM. Autoimmune pathogeneses of schizophrenia have been hypothesized; however, the clinical delineation of a potentially corresponding subset of patients is rarely addressed. CASE REPORT: We treated a 22-year-old white female who carried the concomitant diagnoses of Schizophrenia, IDDM, and Hypothyroidism with quetiapine and risperidone on an acute basis at our inpatient facility, and observed an apparent resolution of her brittle diabetes with the successful treatment of her psychotic disorder. CONCLUSIONS: The well documented link between antipsychotic agents and changes in blood glucose may be of benefit in a subset of patients who suffer from both psychotic and diabetic disorders. PMID- 15666962 TI - Mesial temporal sclerosis and psychiatric symptoms: a case report. AB - We present the history of a woman with mesial temporal sclerosis (MTS) who was diagnosed with a psychotic depression before the correct diagnosis was made. The atypical course and some specific features raised doubt on the initial diagnosis. This case demonstrates the difficulty making a diagnosis of MTS. PMID- 15666961 TI - A case of bipolar disorder and typhoid fever. AB - Many medical disorders presentwith psychiatric symptoms. Teasing out what is caused by the medical problem and what is underlying psychopathology can be a tremendous challenge. We report the case of a young man with no documented past psychiatric history that developed symptoms consistent with bipolar disorder with mood-congruent psychotic features as well as typhoid fever. In this report, we summarize the biological, psychological, and social underpinnings of this rare and interesting case. PMID- 15666963 TI - A malignant neuroleptic spectrum: review of diagnostic criteria and treatment implications in three case reports. AB - Prompt identification of cases of Neuroleptic Malignant Syndrome (NMS) pose a diagnostic and treatment challenge. Three hospital cases of neuroleptic toxicity were reviewed and compared across five published diagnostic criteria sets for NMS. Of these criteria sets, only Levenson's criteria led to the detection of NMS in two out of our three patients. This criteria set supports a NMS spectrum concept, allowing earlier diagnosis and intervention. Lorazepam was used as initial treatment, which provided significant improvement. Use of Levenson's criteria for early diagnosis of NMS and lorazepam for its treatment may be useful tools for the early management of NMS. PMID- 15666964 TI - Development of a toolbox to assess microbial contamination risks in small water systems. AB - Individual and small water systems account for the majority of waterborne disease outbreaks recorded in the United States each year. To address this problem a project was undertaken to develop a comprehensive self-assessment toolbox that could be used by small water system personnel to determine where their system has the greatest potential risks from microbial contamination. The toolbox components consist of: (1) a survey that asks specific questions; (2) a ranking tool that computes numerical scores for water system components based on survey answers; (3) comments and results from the ranking tool; (4) a guidance document to help the user understand why certain conditions may represent a risk; and (5) instructions for using the toolbox. A unique feature of the ranking tool is the ability to input expert opinion in the form of scores for each answer and weighting factors. Weighting factors are derived using ranked, pairwise comparisons, and then used to determine numerical scores for system components. Toolbox administrators are allowed to modify weighting factors used by the ranking tool application, thus allowing input of expert opinion. PMID- 15666965 TI - Chlorination by-products (CBPs) in drinking water and adverse pregnancy outcomes in Italy. AB - Chlorination by-products (CBPs) in drinking water have been associated with an increased risk of adverse pregnancy outcomes, including small for gestational age at term (term-SGA) and preterm delivery. Epidemiological evidence is weakened by a generally inaccurate exposure assessment, often at an ecological level. A case control study with incident cases was performed in nine Italian towns between October 1999 and September 2000. A total of 1,194 subjects were enrolled: 343 preterm births (26th-37th not completed week of pregnancy), 239 term-SGA (from 37th completed week, and weight less than the lowest 10th percentile) and 612 controls. Exposure was assessed both by applying a questionnaire on mothers' personal habits during pregnancy and by water sampling directly at mothers' homes. Levels of trihalomethanes (THMs) were low (median: 1.10 microg l(-1)), while chlorite and chlorate concentrations were relatively high (median: 216.5 microg l(-1) for chlorites and 76.5 microg l(-1) for chlorates). Preterm birth showed no association with CBPs, while term-SGA, when chlorite levels > or =200 microg l(-1) combined with low and high levels of inhalation exposure are considered, suggested a dose-response relationship (adjusted-Odds Ratios (ORs): 1.52, 95%CI: 0.91-2.54 and 1.70, 95%CI: 0.97-3.0, respectively). A weak association with high exposure levels of either THMs (> or =30 microg l(-1)), or chlorite or chlorate (> or =200 microg l(-1)) was also found (adjusted-OR: 1.38, 95%CI: 0.92-2.07). Chlorine dioxide treatment is widespread in Italy; therefore, chlorite levels should be regularly and carefully monitored and their potential effects on pregnancy further evaluated and better understood. PMID- 15666966 TI - Tracking the origin of faecal pollution in surface water: an ongoing project within the European Union research programme. AB - The objectives of this study are to generate knowledge about methods to track the sources of faecal pollution in surface waters, with the aim of having one or a few easy procedures applicable to different geographic areas in Europe. For this, a first field study using already proposed methods (genotypes of F-specific RNA bacteriophages, bacteriophages infecting Bacteroides fragilis, phenotypes of faecal coliforms and enterococci, and sterols) has been done in five areas representing a wide array of conditions in Europe. The present faecal indicators (faecal coliforms, enterococci, sulfite reducing clostridia and somatic coliphages) have also been included in this first field study. At the same time some emerging methods have been settled or adapted to water samples and assayed in a limited number of samples. The results of this first field study indicate that no single parameter alone is able to discriminate the sources, human or non human, of faecal pollution, but that a 'basket' of 4 or 5 parameters, which includes one of the present faecal indicators, will do so. In addition, numerical analysis of the data shows that this 'basket' will allow the successful building of predictive models. Both the statistical analyses and the studied predictive models indicate that genotype II of F-specific RNA bacteriophages, the coprostanol and the ratio coprostanol: coprostanol+epicoprostanol are, out of the studied parameters, those with a greater discriminating power. Either because unsuccessful adaptation of the methods to water samples or because the preliminary assays in water samples indicated low discriminating capability, only three (sorbitol-fermenting bifidobacteria, some species of bifidobacteria detected by PCR with specific primers and phages infecting Bacteroides tethaiotaomicron) of the newly assayed methods have been considered for a second field study, which is currently underway. Expectations are that these new tools will minimize the number of parameters in the 'basket', or at least minimize the difficulty in assaying them. PMID- 15666967 TI - Comparison of bacteriophages for use in iodine inactivation: batch and continuous flow studies. AB - Inactivation rates in batch studies for four commonly used surrogate bacteriophages were measured in stable aqueous iodine solutions for the purpose of determining which was the most suited to evaluate iodine disinfection efficacy in batch and continuous flow conditions. Two types of group Leviviridae bacteriophages were used, Type I (MS2) and Type II (GA), along with group Microviridae, Phi-X174, and group Tectiviridae, PRD1. Inactivation was compared at iodine doses of 1.0-1.5 mg l2/l. MS2 was the most susceptible to iodine inactivation of the four phages tested. Inactivation of naked, icosahedral bacteriophages, MS2 and Phi-X174 demonstrated removals to below detection limits (>99.99%) in less than 10 min. Lipid-containing PRD1 and F+ssRNA GA bacteriophages demonstrated the greatest iodine resistance in batch experiments with an average of 1.82 logs of inactivation (98.5%) after 60 min and 1.05 logs of inactivation (91.1%) after 30 min respectively. Similarly, in continuous flow studies through pentaiodide quaternary ammonium strong base resin, MS2, GA and Phi-X174 were more strongly inactivated than PRD1. The lipid component of PRD1 is thought to enhance resistance to iodine over non-lipid-containing bacteriophages by protecting easily oxidized groups on the protein capsid, but further research is needed before proving this hypothesis. The results from this research may provide a surrogate standard for more rigorous and developed research into the mode of iodine disinfection and its inactivation kinetics. PMID- 15666968 TI - The effectiveness of a multi-spark electric discharge system in the destruction of microorganisms in domestic and industrial wastewaters. AB - The aim of this work was to investigate the effectiveness of a high voltage multi spark electric discharge, with pulse energy of 1 Joule, in killing microorganisms in wastewater. Wastewater from primary treated effluent arising from domestic and industrial sources was abstracted for continuous pulsed discharge disinfection. The wastewater contained a large mixed population of microorganisms (approximately 10(7) CFU ml(-1) [10(9) CFU 100 ml(-1)] total aerobic heterotrophic bacteria) including vegetative cells and spores. The electrical conductivity of the wastewater ranged from 900-1400 microS cm(-1) and it was shown that a specific energy of 1.25-1.5 J cm(-3) was required to achieve 1 log reduction in bacterial (faecal coliforms/total aerobic heterotrophs) content. This is higher than that previously shown to reduce the population of E. coli in tap water of low conductivity, demonstrating the role of total wastewater constituents, including dissolved and particulate substances, water colour and the presence of microbial spores, in effective disinfection. The system can be engineered to eradicate microbial populations to levels governed by legislation by increasing treatment time or energy input. PMID- 15666969 TI - [A century of the The German Society of Forensic Medicine or Legal Medicine]. AB - On 20 September 1904 the Section of Forensic Medicine of the former Society of German Natural Scientists and Physicians decided in Breslau to found an independent "German Society of Forensic Medicine". The constituent assembly was held in Meran on 25 September 1905, with Fritz Strassmann from Berlin being elected as the first president. According to section 1 of the first statutes the object of the association was to "offer a platform for joint scientific work". Later the name of the association was changed several times; the last change (to "German Society of Legal Medicine") was made in 1968. The close relationship between legal medicine and criminalistics is demonstrated on the background of the historical development. PMID- 15666970 TI - [Differentiation of ballpoint pen inks by thermodesorption and gas chromatography mass spectrometry]. AB - Differentiation and classification of ink entries with dated samples of a reference collection are important aspects in the examination of questioned documents. Classification of writing inks is presently achieved by analysis of dyes and colorants contained in the ink. This technique has its limitations in newly developed ink formulations with identical dye composition but differing in their solvents and binder resins. This paper introduces a method for the determination of solvents and binder resins of an ink sample directly from paper without sample preparation. This aim is accomplished by thermodesorption of the sample followed by gas chromatography/mass spectroscopy. The method was tested on numerous samples of ballpoint pen inks, which were subsequently grouped into several solvent and resin subgroups. A case study shows the applicability of the newly developed method. PMID- 15666971 TI - [Medicolegal aspects of witnessed suicide due to gunshot to the head. II. Legal medicine aspects and examination of the firing hand]. AB - The second part of the paper on suicides by gunshots to the head in the presence of witnesses focuses on relevant morphological autopsy findings such as entrance site, signs of close range or contact shots, bullet path etc. and also discusses selected aspects of ascertaining gunshot residues. For the identification of the shooter an integral medicolegal assessment of all the facts including the investigation results and the autopsy findings is essential. However, the morphological findings alone do not allow safe diagnosis, as for example in a homicide the temporal region, which was affected in all our cases, may have been deliberately chosen by the perpetrator as a localization typical of suicide. Thus methods to ascertain gunshot residues on the firing hand (by means of adhesive films and the polyvinyl-alcohol collection method--PVAL) are of great practical importance. In seven cases adhesive films and/or the polyvinyl-alcohol collection method were used. In one case the gunshot residues (GSR) were analysed by means of tape lifts and subsequent scanning electron microscopy (SEM). It was found that especially the combined application of topographical (adhesive tape/PVAL) and cumulative (SEM) methods allowed for the doubtless identification of the shooter, thus usually confirming the suicide. By the example of one case it is demonstrated that without the immediate collection of evidence at the scene objective reconstruction of the event becomes impossible. On the basis of the reported cases recommendations are finally given for a differentiated approach in the medicolegal evaluation of alleged witnessed suicide by gunshot (to the head). PMID- 15666972 TI - [Fatal electric arc accidents due to high voltage]. AB - The frequency of electric arc accidents has been successfully reduced owing to preventive measures taken by the professional association. However, the risk of accidents has continued to exist in private setting. Three fatal electric arc accidents caused by high voltage are reported with reference to the autopsy findings. PMID- 15666973 TI - [Paternity analysis in deficiency cases with related putative fathers: simulation of a deficiency analysis in 27 families]. AB - During the last few years, the number of privately ordered paternity investigations has increased considerably. Probably due to financial reasons in more and more cases only the putative father and the child are investigated. Additionally, very often only one method, such as STR analysis, is employed. This raises the question whether such a reduced analysis leads to reliable and clear results when investigating cases with related putative fathers. We investigated 165 individuals from 27 families using the AmpFlSTRIdentifiler multiplex PCR and calculated the paternity probabilities of the children to their biological fathers, uncles, grand fathers and other relatives. In more than 30% less than three exclusions between child and relative were detected. In five cases no exclusions were found between child and uncle, always leading to paternity probabilities >99.9%. These results show that the calculation of high probabilities (>99.9%) does not necessarily lead to the accurate conclusion of fatherhood. In many of our cases misleadingly the brother of the real father or another close relative would have been declared to be the biological father. PMID- 15666974 TI - Effect of probiotic supplementation on laying hen diets on yield performance and serum and egg yolk cholesterol. AB - The effects of dietary supplementation of a commercial probiotic (BioPlus 2B) on daily feed consumption, egg yield, egg weight, specific gravity, body weight, feed conversion ratio, serum and egg yolk cholesterol, and serum trigylceride in layer hens were investigated. In 12 replicates, 480 27-week-old Brown-Nick layers were fed with diets containing 0, 250, 500 or 750 mg kg(-1) probiotic for 90 days. When compared with the controls, supplementation of 250, 500 and 750 mg kg( 1) probiotic increased egg production, but decreased the damaged egg ratio (p < 0.05), egg yolk cholesterol and serum cholesterol (p < 0.001) levels. In addition, serum triglyceride levels were reduced by using 500 and 750 mg kg(-1) probiotic supplementation (p < 0.001). Feed conversion ratios were positively affected by supplementation of 250 and 500 mg kg(-1) probiotic compared with controls (p < 0.05). There was no statistically significant difference between the control and all treatment groups on feed consumption, egg weight, specific gravity, body weight, and egg yolk weight. PMID- 15666975 TI - Novel source of semicarbazide: levels of semicarbazide in cooked crayfish samples determined by LC/MS/MS. AB - Nitrofuran antibiotics were previously used in animal healthcare but are now prohibited. Semicarbazide is a breakdown product of 5-nitrofurazone and protein bound semicarbazide is used as a marker residue for the illegal use of 5 nitrofurazone. However, the presence of the prohibited semicarbazide has been reported in some food items of animal origin. A novel observation is reported that semicarbazide can be detected in Finnish crayfish samples, i.e. crustacea, never medicated with nitrofurazone. The origin of the semicarbazide is presently unknown. Positive identification was undertaken by liquid chromatography coupled with tandem mass spectrometry detection. The level of semicarbazide was determined as the protein-bound form as well as the total amount of semicarbazide in the sample. The average levels of total semicarbazide and the protein-bound form were 4.2 and 0.5 ng g(-1) fresh crayfish meat, respectively. All the tested samples (n = 18) contained traces of semicarbazide, the highest amount being 12 ng g(-1) fresh crayfish meat. PMID- 15666976 TI - Prevalence of zeranol, taleranol and Fusarium spp. toxins in urine: implications for the control of zeranol abuse in the European Union. AB - There is currently little information concerning the prevalence of zeranol and taleranol in animal urine following metabolism of the naturally occurring Fusarium spp. toxins. An epidemiological study is described which involves four European Union control laboratories in which 8008 urine samples were screened for the presence of zeranol using a time-resolved fluoroimmunoassay (TR-FIA). Of these samples, 93.6% screened negative for zeranol. All samples testing positive for zeranol were then analysed with a confirmatory method. Based on the confirmatory results, the TR-FIA-positive samples were then categorized as false positive, true-positive or 'equivocal' (zeranol/taleranol and the Fusarium spp. toxins detected). The true-positive samples represented only 0.05% of the total number of samples (n = 4). After statistical analysis, 170 of 174 equivocal samples proved to belong to a 'normal' population in which the amount of zeranol/taleranol could be related to the total amount of Fusarium spp. toxins through a linear regression with a 99% prediction interval. This suggested that the presence of zeranol in these samples might be due to in vivo metabolism of the Fusarium spp. toxins. The presence of zeranol in the four remaining 'outliers' might be attributable to zeranol abuse rather than to natural contamination. The results are of interest for control laboratories as they might provide an analytical tool to help distinguish between abuse and natural contamination in zeranol testing. PMID- 15666977 TI - Detection of 3-amino-2-oxazolidinone (AOZ), a tissue-bound metabolite of the nitrofuran furazolidone, in prawn tissue by enzyme immunoassay. AB - Furazolidone, a nitrofuran antibiotic, is banned from use in food animal production within the European Union. Increasingly, compliance with this ban is monitored by use of analytical methods to detect a stable tissue-bound metabolite, 3-amino-2-oxazolidinone (AOZ). Widespread use of furazolidone in poultry and prawns imported into Europe highlighted the urgent need for development of nitrofuran immunoassay screening tests. The first enzyme-linked immunoabsorbant assay for detection of AOZ residues in prawns (shrimps) is now described. Prawn samples were derivatized with o-nitrobenzaldehyde, extracted into ethyl acetate, washed with hexane and applied to a competitive enzyme immunoassay based on a rabbit polyclonal antiserum. Assay limit of detection (LOD) (mean + 3 s) calculated from the analysis of 20 known negative cold and warm water prawn samples was 0.1 microg kg(-1). Intra- and interassay relative standard deviations were determined as 18.8 and 38.2%, respectively, using a negative prawn fortified at 0.7 microg kg(-1). The detection capability (CCbeta), defined as the concentration of AOZ at which 20 different fortified samples yielded results above the LOD, was achieved at fortification between 0.4 and 0.7 microg kg(-1). Incurred prawn samples (n = 8) confirmed by liquid chromatography coupled with tandem mass spectrometry detection to contain AOZ concentrations between 0.4 and 12.7 microg kg(-1) were all screened positive by this enzyme linked immunoabsorbant assay. Further data are presented and discussed with regard to calculating assay LOD based on accepting a 5% false-positive rate with representative negative prawn samples. Such an acceptance improves the sensitivity of an ELISA and in this case permitted an LOD of 0.05 microg kg(-1) and a CCbeta of below 0.4 microg kg(-1). PMID- 15666978 TI - Methylmercury levels in predatory fish species marketed in Canada. AB - Mercury was detected in all analysed samples of swordfish, marlin, shark and tuna purchased from major supermarket outlets and fish retailers in three cities across Canada. Total mercury and methylmercury levels ranged up to 3845 and 2346 ng g(-1), respectively. Swordfish contained the highest levels, followed by shark, fresh/frozen tuna and marlin. Levels in canned tuna were considerably less than the other examined samples. Methylmercury was extracted with toluene from enzymatically hydrolysed samples after the addition of sulphuric acid and potassium bromide. An L-cysteine back-extraction was used to separate the methylmercury from most organic co-extractives. Analysis of methylmercury (as methylmercury bromide) was by gas chromatography with pulsed discharge detection. PMID- 15666979 TI - Effects of refining predicted chronic dietary intakes of pesticide residues: a case study using glyphosate. AB - A chronic consumer risk assessment based on a worst-case scenario, conducted as part of the European Union review leading to Annex I inclusion for glyphosate, was evaluated and refined. An extensive database of information on the effects of processing on the levels of glyphosate residues in food is available. This database together with refined consumption data from the UK's surveys of adults and toddlers and extensive monitoring data of glyphosate residues in mainly cereal products conducted in the UK were combined to examine the potential overestimates of dietary intakes that are predicted using the current regulatory methodology developed by the Food and Agricultural Organization/World Health Organization and applied as part of the European Union regulatory process. Analysis focussed on the chronic exposure from treated cereals, the crop group contributing significantly to the dietary intake of glyphosate residues. A steep reduction of predicted intake was seen when progressively realistic measures of residues were incorporated into the models, giving a strong indication of the conservative nature of current regulatory procedures. Calculations using even the most unrefined methodology gave rise to intakes of up to 11% of the acceptable daily intake, this was reduced to 0.6% of the acceptable daily intake when justifiable refinements based on extensive monitoring data collected in the UK were made. Consumption data for processed foods abstracted from the UK Food Standard Agency's database were used to refine further the predicted dietary intakes as a result of residue reductions or concentration from processing. The current regulatory model used in the UK generally only has the potential to use a single value for consumption of a particular food. The Pesticides Safety Directorate model consistently predicted the highest intakes with the exception of intakes by adults using the supervised trials median residue and median monitoring data. This suggests that conservatism in the regulatory model exists particularly where specific processing factors cannot be applied to individual fractions of the diet. PMID- 15666980 TI - Fumonisin contamination of maize in Burkina Faso, West Africa. AB - Maize throughout the world is frequently contaminated with a family of mycotoxins, the fumonisins, produced by species of Fusaria. The study investigated the level of fumonisin contamination of maize samples from village farms and large market traders in Burkina Faso, West Africa. Maize samples (5 kg) from each of five to six large storage barns from farms in five villages in the district of N'Dorola, Kenedougou province, western Burkina Faso, were sampled (n = 26) in Jan 1999 (> 1 year storage), and a further 26 maize samples from the same farms were collected directly from the field in October 1999. In addition, 72 maize samples were obtained in July 1999 from large markets in Bobo Dioulasso. Fumonisins were extracted from dried maize, derivatized with o-phthaldialdehyde and quantified by reversed-phase high-performance liquid chromatography with fluorescence detection. All 26 samples from the first (mean 1170 microg kg(-1), range 110-3120 microg kg(-1)) and from the second (mean 130 microg kg(-1), range 10-450 microg kg(-1)) village collection were fumonisin positive. All 72 maize samples from the large markets were also positive for fumonisins, and had the highest levels of contamination (mean 2900 microg kg(-1), range 130-16,040 microg kg(-1)). As fumonisins were a ubiquitous contaminant of maize and given that this crop is a dietary staple in this region, chronic exposure is likely. PMID- 15666981 TI - Effect and stability of gliotoxin, an Aspergillus fumigatus toxin, on in vitro rumen fermentation. AB - Aspergillus fumigatus is a toxicogenic fungus usually found in contaminated animal feeds, especially in conserved forages where it can produce several mycotoxins. Gliotoxin, one of the most important toxic metabolites produced by this fungus, has antibacterial, immunosuppressive and apoptotic effects. Ruminants due to the high proportion of forages they receive in the ration would be particularly exposed to gliotoxin. The objective of this work was (1) to assess the effect of gliotoxin on in vitro rumen fermentation and (2) to determine the effect of fermentation on gliotoxin stability. Gliotoxin did not affect rumen fermentation at concentrations found in naturally contaminated feeds. No effects were observed up to a concentration of 20 microg toxin ml(-1) and an extremely high toxin concentration (80 microg ml(-1)) was necessary to affect dry matter degradation, gas and total volatile fatty acids production by 24, 37 and 18%, respectively (p < 0.01). In addition, the toxin was unstable in the rumen environment with 90% disappearance at 6 h of incubation (p < 0.05). In contrast, extracts of A. fumigatus cultures containing gliotoxin at concentrations several times lower than that used for experiments with pure toxin had a negative effect on fermentations indicating the toxicity and possible synergism of other metabolites produced by this fungus. Extracts containing 8.8 microg gliotoxin ml(-1) decreased dry matter degradation, gas and volatile fatty acids production by 28, 46 and 35%, respectively (p < 0.01). Identification of these toxic metabolites and assessment of the rate of passage of gliotoxin to the lower intestinal tract is necessary to evaluate the potential risk of these toxins to ruminants. PMID- 15666982 TI - Azaspiracid poisoning, the food-borne illness associated with shellfish consumption. AB - Azaspiracid poisoning (AZP) is a recently discovered toxic syndrome that was identified following severe gastrointestinal illness from the consumption of contaminated mussels (Mytilus edulis). The implicated toxins, azaspiracids, are polyethers with unprecedented structural features. Studies toward total toxin synthesis revealed that the initial published structures were incorrect and they have now been revised. These toxins accumulate in bivalve molluscs that feed on toxic microalgae of the genus Protoperidinium, previously considered to be toxicologically benign. Although first identified in shellfish from Ireland, azaspiracid contamination of several types of bivalve shellfish species has now been confirmed throughout the western coastline of Europe. Toxicological studies have indicated that azaspiracids can induce widespread organ damage in mice and that they are probably more dangerous than previously known classes of shellfish toxins. The exclusive reliance on live animal bioassays to monitor azaspiracids in shellfish failed to prevent human intoxications. This was a consequence of poor sensitivity of the assay and the fact that azaspiracids are not exclusively found in the shellfish digestive glands used for toxin testing. The strict regulatory control of azaspiracids in shellfish now requires frequent testing of shellfish using highly specific and sensitive methods involving liquid chromatography-mass spectrometry. PMID- 15666983 TI - Relative hopane content confirming the mineral origin of hydrocarbons contaminating foods and human milk. AB - Hopanes, triterpenoid hydrocarbons formed under geological conditions, were analysed to confirm the mineral origin of the unresolved complex mixtures of hydrocarbons observed in the gas chromatography with flame ionization detection chromatograms of human milk and certain foodstuffs. The 'relative hopane content' (RHC) is introduced, i.e. it is the area ratio of the sum of the hopanes and the paraffins in the same segment of the chromatogram. The RHC in various mineral oil products (motor oils, hydraulic oils, lubricating oils, Vaseline) was 3.4%, with a relative standard deviation of 19%. The RHC determined in samples of vegetable oils, mussels and clams as well as of human milk containing an unresolved complex mixture of hydrocarbons was in the same range, confirming that these samples were contaminated by mineral oil material. PMID- 15666985 TI - M.D. Anderson, Ball cancer centers focus on research, care, prevention. AB - The folks at M.D. Anderson Cancer Center, Houston, win first place in this category with their traditionally excellent report. The Cancer Center at Ball Memorial Hospital, Muncie, Ind., receives the second place award with a report that uses unusual design elements to "shatter the myths" about cancer research. PMID- 15666984 TI - Review of the toxicology, human exposure and safety assessment for bisphenol A diglycidylether (BADGE). AB - BADGE (whose chemical names are bisphenol A diglycidylether and 2,2-bis(4-(2,3 epoxypropyl)phenyl)propane) is the lowest molecular weight oligomer in commercial epoxy resins and the major component in commercial liquid epoxy resins. The major application areas for epoxy resins are protective coatings and civil engineering. Additional applications include printed circuit boards, composites, adhesives and tooling, while a relatively small amount of epoxy resins (< 10%) finds use in protective coatings inside food and drink cans. The use of BADGE in food-contact applications was first regulated through EC Directive 2002/16/EC and amended in EC Directive 2004/13/EC with migration levels in food-contact applications being generally well below the regulatory thresholds. The paper discusses the commercial use of BADGE focusing on the current knowledge of human exposure from canned food applications. To assess the safety of this application, the exposure data are compared with no adverse effect levels (NOAEL) from various toxicological investigations with BADGE including reproductive and developmental assays, endocrine toxicity investigations, and sub-chronic and chronic assays. Consumer exposure to BADGE is almost exclusively from migration of BADGE from can coatings into food. Using a worst-case scenario that assumes BADGE migrates at the same level into all types of food, the estimated per capita daily intake for a 60-kg individual is approximately 0.16 microg kg(-1) body weight day(-1). A review of one- and two-generation reproduction studies and developmental investigations found no evidence of reproductive or endocrine toxicity, the upper ranges of dosing being determined by maternal toxicity. The lack of endocrine toxicity in the reproductive and developmental toxicological tests is supported by negative results from both in vivo and in vitro assays designed specifically to detect oestrogenic and androgenic properties of BADGE. An examination of data from sub-chronic and chronic toxicological studies support a NOAEL of 50 mg kg( 1) body weight day(-1) from the 90-day study, and a NOAEL of 15 mg kg(-1) body weight day(-1) (male rats) from the 2-year carcinogenicity study. Both NOAELS are considered appropriate for risk assessment. Comparing the estimated daily human intake of 0.16 microg kg(-1) body weight day(-1) with the NOAELS of 50 and 15 mg kg(-1) body weight day(-1) shows human exposure to BADGE from can coatings is between 250,000 and 100,000-fold lower than the NOAELs from the most sensitive toxicology tests. These large margins of safety together with lack of reproductive, developmental, endocrine and carcinogenic effects supports the continued use of BADGE for use in articles intended to come into contact with foodstuffs. PMID- 15666986 TI - Shining the light on their donors. Anne Arundel Medical Center and Palo Alto Medical Foundation. AB - The first-place report from Anne Arundel Medical Center, Annapolis, Md. and second-place winner Palo Alto Medical Foundation, Palo Alto, Calif., succeed in their common objectives. Each provides a memorable portrait of the organization, likely to impress donors and build additional support. PMID- 15666987 TI - Fact-based information scores high. Regional reports address their neighbors in Iowa and Vermont. AB - They're in far different parts of the country, but first place awardee St. Anthony Regional Hospital and Nursing Home, Carroll, Iowa; and second place winner Rutland Regional Medical Center, Rutland, Vt., share a great deal. Both acknowledge the interests and preferences of their own local populations. Both use their pages effectively to deliver information within budgetary constraints. PMID- 15666988 TI - Community hospital reports demonstrate positive attitudes. AB - First place winner in the community hospital category is Windham Community Memorial Hospital, Willimantic, Conn. Windham created an annual with a distinctive format, then expanded its use by reprinting the pages separately for strategic distribution. Second place Evangelical Community Hospital, Lewisburg, Pa., epitomizes the optimistic approach seen in most of this year's entries. This colorful report was distributed with local newspapers as well as being mailed to donors. PMID- 15666989 TI - Large system reports meet BIG task, with costly, lavish productions. AB - HealthONE, Denver, takes first place in this category, with a richly printed, 56 page book that personalizes each hospital in its own section. Second place winner, North Shore-Long Island Jewish Health System, Great Neck, N.Y., effectively combines a progress report and a financial report. Kaleida Health, Buffalo, N.Y. takes third place honors with a well-designed, upbeat presentation. Judges also awarded special recognition to Baystate Health System, Springfield, Mass., for its combined printed/CD-ROM report. Each system in this category serves more than 700 beds. PMID- 15666990 TI - They are smaller, but these systems produce mighty reports. AB - The first place winner, Commonwealth Health Corporation, Bowling Green, Ky., has a successful cost-saving story. Designed in-house, with donated printing, it cost a mere 54 cents per unit. Little Company of Mary Hospital, Evergreen Park, Ill., wins second place with a publication that enlisted the help of many personnel. University Health Care System, Augusta, Ga., third place winner, uses dramatic graphics to observe its 185th anniversary. Princeton HealthCare System, Princeton, N.J., receives special recognition for the clarity and effectiveness of its four-page report. PMID- 15666991 TI - Quinone formation as dopaminergic neuron-specific oxidative stress in the pathogenesis of sporadic Parkinson's disease and neurotoxin-induced parkinsonism. AB - Parkinson's disease (PD) is a progressive neurodegenerative disease characterized by dopaminergic neuron-specific degeneration in the substantia nigra. A number of gene mutations and deletions have been reported to play a role in the pathogenesis of familial PD. Moreover, a number of pathological and pharmacological studies on sporadic PD and dopaminergic neurotoxin-induced parkinsonism have hypothesized that mitochondrial dysfunction, inflammation, oxidative stress, and dysfunction of the ubiquitin-proteasome system all play important roles in the pathogenesis and progress of PD. However, these hypotheses do not yet fully explain the mechanisms of dopaminergic neuron-specific cell loss in PD. Recently, the neurotoxicity of dopamine quinone formation by auto oxidation of dopamine has been shown to cause specific cell death of dopaminergic neurons in the pathogenesis of sporadic PD and dopaminergic neurotoxin-induced parkinsonism. Furthermore, this quinone formation is closely linked to other representative hypotheses in the pathogenesis of PD. In this article, we mainly review recent studies on the neurotoxicity of quinone formation as a dopaminergic neuron-specific oxidative stress and its role in the etiology of PD, in addition to several neuroprotective approaches against dopamine quinone-induced toxicity. PMID- 15666992 TI - Intrapulmonary lymph nodes: thin-section CT findings, pathological findings, and CT differential diagnosis from pulmonary metastatic nodules. AB - We compared the thin-section CT findings of 11 intrapulmonary lymph nodes with pathological findings and evaluated the possibility of CT scan differential diagnosis from pulmonary metastatic nodules. First, we retrospectively reviewed CT scan and pathological findings of intrapulmonary lymph nodes. The median size of these nodules was 6.2 mm. The nodules appeared round (n=3) or angular (n=8) in shape with a sharp border, and they were found below the level of the carina. The median distance from the nearest pleural surface was 4.6 mm, and 3 of the 11 nodules were attached to the pleura. On thin-section CT scan, linear densities extending from the intrapulmonary lymph nodes were frequently visualized, and were pathologically proven to be ectatic lymphoid channels. We then compared the thin-section CT findings of 8 metastatic nodules less than 1 cm in diameter with those of the 11 intrapulmonary lymph nodes. The median size of these nodules was 6.8 mm, and the median distance from the nearest pleural surface was 16 mm. All nodules appeared round in shape. None of the nodules had linear densities extending from the nodules. The linear densities on thin-section CT scan may be the most useful characteristic of intrapulmonary lymph nodes, when differential diagnosis from metastatic nodules is necessary. PMID- 15666993 TI - Characteristics and outcomes of school refusal in Hiroshima, Japan: proposals for network therapy. AB - The authors conducted a study on children undergoing treatment at major school refusal treatment centers in Hiroshima Prefecture. On the whole, school refusal in the prefecture was found to peak between 13 and 14 years of age. By age group, the main reason for school refusal in elementary school group was parent-child relationship with separation anxiety. Given additional problems such as neglect at home and complicated social situations in their schools, junior high school students were found to present diverse symptoms from introversion and self analysis to extroversion, neglect of studies, and delinquency. Among high school students, there were more cases suffering withdrawal and schizophrenia spectrum disorders. The major task regarding treatment seems to lie in how to treat complex cases combining different problems. We summarized herein the studies we have carried out and propose a model for a network therapy system based on functional liaisons between treatment centers. With this system, a child psychiatric medical facility plays the part of a liaison center for the overall network system. PMID- 15666994 TI - A thoracic-epidural granulocytic sarcoma case that was diagnosed preceding the onset of and that recurred co-incidental to acute promyelocytic leukemia, which developed after surgical treatment. AB - Granulocytic sarcoma or chloroma is a tumor seen in myelocytic leukemia. Spinal epidural onset is rare and is generally seen before or together with the onset of myelocytic leukemia. An epidural mass located at the 2nd-5th thoracic levels in an 18-year-old male patient was pathologically diagnosed as granulocytic sarcoma. Radiotherapy was performed after surgical intervention. Ten months later, he was re-admitted with abdominal pain. At this time, an epidural mass at the 6th-9th thoracic levels was detected on magnetic resonance imaging, and acute promyelocytic leukemia was diagnosed. After systemic chemotherapy, partial remission was achieved. We aimed to present this rare case with its remarkable follow-up findings. PMID- 15666995 TI - Copper chelation as an antiangiogenic therapy. AB - Angiogenesis is now recognized as a crucial process in tumor development. Copper appears to act as an essential cofactor for several angiogenic growth factors, and both copper metabolism and ceruloplasmin expression are upregulated in many tumors. The role of copper chelators has been investigated in animal models with promising results. New therapies for Wilson's disease (a disease of copper accumulation) have enabled clinical trials of copper chelation to be undertaken. Here we discuss the evidence for a role of copper in angiogenesis and possible mechanisms of action of anticopper agents. PMID- 15666996 TI - Design and construction of a phosphorylatable chimeric monoclonal antibody with a highly stable phosphate. AB - A recognition site for the cAMP-dependent protein kinase was introduced into the MAb-chCC49 by site-directed mutation of the coding sequence to make a variant of MAb-chCC49 containing a highly stable phosphate. To design this monoclonal antibody (MAb) without changing its immunoreactivity or biological properties, molecular modeling was used to locate appropriate regions for introduction of the cAMP-dependent phosphorylation site with desirable properties. We selected one position to mutate on the heavy chain based on molecular dynamics study of the solvated antibody. A vector expressing the mutant was constructed and transfected into mouse myeloma NS0 cells that expressed a high level of the resultant MAb WW5. MAb-WW5 contained the cAMP-dependent phosphorylation site at the hinge region of the heavy chain, could be phosphorylated by the catalytic subunit of cAMP-dependent protein kinase with [gamma-32P]ATP to high specific activity, and retained the phosphate stably. Compared with MAb-chCC49K1, another phosphorylatable variant of MAb-chCC49, the phosphate attached to MAb-WW5 showed much improved stability: about a 10-fold increase in resistance to hydrolysis. MAb-WW5 exhibited the same binding specificity to the TAG-72 antigen on MCF-7 4C10 breast cancer cells as we observed with MAb-chCC49K1. The improved stability of the attached phosphate provides a MAb with potential to be used in diagnosis and therapy of adenocarcinomas. PMID- 15666997 TI - Radiation-induced sensitizing effect of folic acid (vitamin B11) and its synergistic action to mitomycin C: in vitro experiments and radiolysis. AB - Folic acid (FA; vitamin B11) is found to exhibit very strong cytostatic effects upon gamma-irradiation in neutral media. This effect depends on the FA concentration, pH of the media, and on the presence of air. The largest cytostatic efficiency of FA was observed in an air-free environment (pH 7.4), where deltaD37 = -90 for 10(-4) mol/dm3 FA and deltaD37 = -160 for 10(-3) mol/dm3 FA was found. FA also acts as a very efficient electron donor and is therefore able, in air-free neutral media, to enhance the efficiency of 7.5 x 10(-7) mol/dm3 mitomycin C (MMC) from deltaD37 = -80 up to deltaD37 = -200 in the presence of 5 x 10(-5) mol/dm3 FA, respectively. This synergistic effect offers a new pathway for more efficient radiation therapy by joint implementation of MMC and FA. Additionally, FA radiolysis was studied in the absence and presence of air as well as in media saturated with N20 (conversion of e(aq)- into OH). As a result of these investigations, various products were detected: ammonia, aldehydes, and a mixture of carboxylic acids, in addition to some not yet identified compounds. Their individual yields could not be identified at present. PMID- 15666998 TI - Resistance of ovarian carcinoma cells to docetaxel is XIAP dependent and reversible by phenoxodiol. AB - Although several pathways have been proposed to explain chemoresistance, all lead to some specific defect in the mechanism of apoptosis. The objective of this study was to characterize the molecular mechanisms of drug resistance to docetaxel in epithelial ovarian cancer cells (EOC) and the use of phenoxodiol as a chemosensitizer. Four established and 12 primary cultures of ovarian carcinoma cell lines (EOC) were treated with docetaxel (5-500 ng/ml) for 24 and/or 48 h. In all the studied cell lines, the best response was seen using 500 ng/ml of docetaxel. Sensitive cell lines were identified as those with IC50 < 100 ng/ml for 48 h while resistant cell lines were identified as those with IC50 > 100 ng/ml. The morphological features of apoptosis and the activation of caspases were seen only in the sensitive cell lines determined by Hoechst staining and Caspase Glo assay. Although X-linked inhibitor of apoptosis protein (XIAP) was expressed in all EOC cells, it was only inactivated in chemosensitive cells. We confirmed the role of XIAP in docetaxel resistance by downregulation of XIAP expression using RNA interference (RNAi) as well as by pretreatment with phenoxodiol. Our results indicate that 1) docetaxel induces its cytotoxic effect through the activation of apoptosis; 2) caspase activation relies on the removal of XIAP; and 3) phenoxodiol restores sensitivity in docetaxel-resistant EOC cells. We demonstrate that phenoxodiol, by interfering with XIAP activity, functions as a chemosensitizer to docetaxel and could provide a more effective treatment for refractory ovarian cancer. PMID- 15666999 TI - The unique biological features of the marine product Yondelis (ET-743, trabectedin) are shared by its analog ET-637, which lacks the C ring. AB - It was previously suggested that the peculiar mechanism of action of the novel anticancer drug Yondelis (ET-743, trabectedin) was due to part of the molecule, units A and B, binding to DNA in the minor groove, causing an alkylation at the N2 of guanine, while unit C protrudes out of DNA, possibly interacting with transcription factors or other DNA binding proteins. To test this hypothesis, we have compared the biological activity and the mode of action of Yondelis with its analogue ET-637, which has the same chemical structure except for the lack of the C ring. Yondelis and ET-637 showed similar cytotoxic potency and cell cycle perturbations. As already reported for Yondelis, the UV-96 cell line, deficient in ERCC-1, was less sensitive to ET-637 than the parental cell line. The binding of Yondelis or ET-637 to DNA-oligonucleotides was demonstrated by gel shift assay and SDS did not reverse the binding. Both compounds blocked the temperature induced activation of the HSP40 promoter in the range of 1-10 nM. This study indicates that ET-637 acts similarly to Yondelis and demonstrates that the C ring of Yondelis may not be required for its biological activity. PMID- 15667000 TI - secErbB4-26/549 antagonizes ligand-induced ErbB4 tyrosine phosphorylation. AB - ErbB4 is a member of the ErbB family of receptor tyrosine kinases. Because of a paucity of appropriate pharmacologic tools, little is known about ErbB4 functions in vivo. In response to this need, we hypothesized that a recombinant form of the extracellular domain of ErbB4 would antagonize ligand-induced receptor tyrosine phosphorylation and subsequent downstream signaling and could be used to probe ErbB4 function. Indeed, we show here that one such ErbB4 protein, secErbB4 26/549, is a potent inhibitor of ligand-induced ErbB4 tyrosine phosphorylation and of ligand-induced ErbB4 coupling to biological responses. Furthermore, we demonstrate that secErbB4-26/549 antagonizes ligand-induced ErbB4 signaling by acting as a ligand sink. Thus, secErbB4-26/549 is suitable for elucidating the effects of ErbB4 ligand-induced ErbB signaling in a variety of biological contexts. PMID- 15667001 TI - Serum levels of soluble E-selectin are associated with the clinical course of metastatic disease in patients with liver metastases from breast cancer. AB - Increasing evidence indicates that the expression of the endothelial adhesion molecule E-selectin is associated with progression and metastasis of breast cancer. Patients with liver metastases also show increased serum levels of the soluble form of E-selectin. It was our aim to compare serum levels of soluble E selectin (sES) in such patients with the biology of the primary tumor and the course of the metastatic disease under therapy. We examined 69 patients with liver metastases from breast cancer who were selected to receive systemic tumor therapy because of progressive disease (n = 44) or newly detected liver metastases (n = 25). Serum concentrations of sES were measured before each therapy cycle using a specific ELISA. Serum concentrations of sES before the start of therapy were compared to clinical parameters and histopathological findings referring to the primary tumor. Secondly, serum levels of sES were compared to serum concentrations of the corresponding tumor markers. We observed a possible trend for certain unfavorable prognostic parameters (e.g., young women, low-graded tumors, human epidermal growth factor receptor 2 overexpression) to be related to higher serum levels of sES. Serum levels of sES were correlated with tumor marker levels in a logarithmical relation (r = 0.44, P < 0.0005). In some cases it could be demonstrated that serum levels of sES changed similarly to the course of tumor marker levels. We conclude that serum levels of sES are associated with the clinical course of liver metastases from breast cancer. Further investigations are needed to clarify if serum levels of sES may serve as tumor marker in certain clinical situations. E-selectin should be evaluated as a possible target for antimetastatic therapy studies. PMID- 15667002 TI - Concentrations of VEGF and VEGFR1 in paired tumor arteries and veins in patients with rectal cancer. AB - Increased plasma concentrations of vascular endothelial growth factor (sVEGF) are associated with poor prognosis of colorectal cancer patients. The aim was to investigate the contribution of the tumor to plasma concentrations of VEGF and VEGF receptor 1 (VEGFR1). Preoperative blood samples from a peripheral vein and intraoperative blood samples from a tumor artery, a tumor vein, and from a peripheral vein were drawn from 28 patients undergoing elective surgical resection of primary rectal cancer. Plasma concentrations of VEGF and VEGFR1 were determined by ELISA. Counts of white blood cells and platelets were performed in all samples. No significant difference between plasma VEGF levels in the obtained blood samples was found (0.35 < P < 0.86). Plasma sVEGFR1 concentrations were significantly increased in tumor veins compared with tumor arteries. In addition, a significant reduction in plasma sVEGFR1 concentrations from preoperative to intraoperative samples was observed. There was a significant efflux of neutrophils to the tumor, but none of the observed changes in plasma VEGF or VEGFR1 levels correlated to changes in counts of white blood cells or platelets (sVEGF: 0.33 < P < 0.73 and sVEGFR1: 0.32 < P < 0.98). No changes in sVEGF plasma concentrations from tumor arteries to tumor veins were demonstrated, whereas there was a significant increase in sVEGFR1 from tumor arteries to tumor veins. Changes in sVEGF or sVEGFR1 from tumor arteries to tumor veins were not associated with changes in counts of white blood cells or platelets. PMID- 15667003 TI - Rural health in Iraqi Kurdistan. PMID- 15667004 TI - Title VII funding and physician practice in rural or low-income areas. AB - CONTEXT: Whether Title VII funding enhances physician supply in underserved areas has not clearly been established. PURPOSE: To determine the relation between Title VII funding in medical school, residency, or both, and the number of family physicians practicing in rural or low-income communities. METHODS: A retrospective cross sectional analysis was carried out using the 2000 American Academy of Family Physicians physician database, Title VII funding records, and 1990 U.S. Census data. Included were 9,107 family physicians practicing in 9 nationally representative states in the year 2000. FINDINGS: Physicians exposed to Title VII funding through medical school and residency were more likely to have their current practice in low-income communities (11.9% vs 9.9%, P< or =.02) and rural areas (24.5% vs 21.8%, P< or =.02). Physicians were more likely to practice in rural communities if they attended medical schools (24.2% vs 21.4%; P =.009) and residencies (24.0% vs 20.3%; P =.011) after the school or program had at least 5 years of Title VII funding vs before. Similar increases were not observed for practice in low-income communities. In a multivariate analysis, exposure to funding and attending an institution with more years of funding independently increased the odds of practicing in rural or low-income communities. CONCLUSIONS: Title VII funding is associated with an increase in the family physician workforce in rural and low-income communities. This effect is temporally related to initiation of funding and independently associated with effect in a multivariate analysis, suggesting a potential causal relationship. Whereas the absolute 2% increase in family physicians in these underserved communities may seem modest, it can represent a substantial increase in access to health care for community members. PMID- 15667005 TI - Capitated Medicaid managed care in a rural area: the impact of Minnesota's PMAP program. AB - CONTEXT: Although states have had difficulty extending Medicaid managed care (MMC) to rural areas, rural models of capitated MMC are expected to grow in response to new federal regulations and the serious budget problems facing nearly all states. As such, understanding the effects of capitated MMC in rural settings is important for policy considerations. PURPOSE: To evaluate the effects of capitated MMC on beneficiary access and use in rural Minnesota. METHODS: We took advantage of delays in the timing of the introduction of MMC across rural counties in Minnesota to estimate the effects of managed care on adults and children under Medicaid using a difference-in-differences framework. FINDINGS: We found that Minnesota's shift from fee-for-service Medicaid to MMC in its rural counties had little effect on access to health care for either adults or children. CONCLUSIONS: Because Minnesota reports that Medicaid costs under MMC are below expected costs under FFS Medicaid, it appears that the primary accomplishment of Minnesota's rural MMC initiative is one of cost savings: MMC provides the same access to care as FFS Medicaid, but at lower cost. With steep budget deficits in nearly all states, other states may want to consider Minnesota's rural MMC model as a mechanism for reducing their Medicaid costs. PMID- 15667006 TI - Rural-urban differences in employment-related health insurance. AB - CONTEXT: Rural residents are disproportionately represented among the uninsured in the United States. PURPOSE: We compared nonelderly adult residents in 3 types of nonmetropolitan areas with metropolitan workers to evaluate which characteristics contribute to lack of employment-related insurance. RESEARCH DESIGN AND ANALYSIS: Data were obtained from the Medical Expenditure Panel Survey, pooled across 3 panels (1996--1998) to enhance the rural sample size. Econometric decomposition was used to quantify the contribution of employment structure to differences in the probability of being offered employment-related health insurance. FINDINGS: The most rural workers are 10.4 percentage points less likely to be offered insurance compared with urban workers; the difference is smaller for residents of other rural areas. In rural counties not adjacent to urban areas, lower wages and smaller employers each account for about one-third of the total difference. CONCLUSIONS: Health insurance disparities associated with rural residence are related to the structure of employment. Major factors include smaller employers, lower wages, greater prevalence of self-employment, and sociodemographic characteristics. PMID- 15667007 TI - Access to transportation and health care utilization in a rural region. AB - CONTEXT: Access to transportation to transverse the large distances between residences and health services in rural settings is a necessity. However, little research has examined directly access to transportation in analyses of rural health care utilization. PURPOSE: This analysis addresses the association of transportation and health care utilization in a rural region. METHODS: Using survey data from a sample of 1,059 households located in 12 western North Carolina counties, this analysis tests the relationship of different transportation measures to health care utilization while adjusting for the effects of personal characteristics, health characteristics, and distance. FINDINGS: Those who had a driver's license had 2.29 times more health care visits for chronic care and 1.92 times more visits for regular checkup care than those who did not. Respondents who had family or friends who could provide transportation had 1.58 times more visits for chronic care than those who did not. While not significant in the multivariate analysis, the small number who used public transportation had 4 more chronic care visits per year than those who did not. Age and lower health status were also associated with increased health care visits. The transportation variables that were significantly associated with health care visits suggest that the underlying conceptual frameworks, the Health Behavior Model and Hagerstrand's time geography, are useful for understanding transportation behavior. CONCLUSIONS: Further research must address the transportation behavior related to health care and the factors that influence this behavior. This information will inform policy alternatives to address geographic barriers to health care in rural communities. PMID- 15667008 TI - Cancer incidence in Kentucky, Pennsylvania, and West Virginia: disparities in Appalachia. AB - CONTEXT: Composed of all or a portion of 13 states, Appalachia is a heterogeneous, economically disadvantaged region of the eastern United States. While mortality from cancer in Appalachia has previously been reported to be elevated, rates of cancer incidence in Appalachia remain unreported. PURPOSE: To estimate Appalachian cancer incidence by stage and site and to determine if incidence was greater than that in the United States. METHODS: Using 1994--1998 data from the central registries of Kentucky, Pennsylvania, and West Virginia, age-adjusted incidence rates were calculated for the rural and nonrural regions of Appalachia. These state rates were compared to rates from the Surveillance, Epidemiology, and End Results (SEER) program for the same years by calculating the adjusted rate ratio (RR) and a 95% confidence interval (CI). FINDINGS: Both the entire and rural Appalachian regions had an adjusted incidence rate for all cancer sites similar to the SEER rate (RR = 1.00 [95% CI, 1.00-1.01] and RR = 0.99 [95% CI, 0.99-1.00], respectively). However, incidence of cancer of the lung/ bronchus, colon, rectum, and cervix in Appalachia was significantly elevated (RR = 1.22 [95% CI, 1.20-1.23], 1.13 [95% CI, 1.11-1.14], 1.19 [95% CI, 1.16-1.22], and 1.12 [95% CI, 1.07-1.17], respectively). Incidence of cancer of the lung/bronchus and cervix in rural Appalachia was even more elevated (RR = 1.34 [95% CI, 1.31-1.36] and 1.29 [95% CI, 1.21-1.38], respectively). Incidence of unstaged disease for all cancer sites in Appalachia (RR = 1.06 [95% CI, 1.05 1.08]), particularly rural Appalachia (RR = 1.28 [95%CI, 1.25-1.301), was elevated. CONCLUSIONS: Cancer incidence in Appalachia was not found to be elevated. However, incidence of cancer of the lung/bronchus, colon, rectum, and cervix was elevated in Appalachia. The rates of unstaged cancer of every examined site were elevated in rural Appalachia, suggesting a lack of access to cancer health care. PMID- 15667009 TI - The social/sexual environment of gay men residing in a rural frontier state: implications for the development of HIV prevention programs. AB - CONTEXT: The incidence and prevalence of HIV/AIDS is increasing among rural men who have sex with men (MSM). Yet little is known about the social/ sexual environment of rural frontier areas. PURPOSE: The purpose of this study was to assess the social/sexual environment of gay men living in rural areas and how this environment contributes to the development of HIV/AIDS prevention programs. METHODS: A qualitative study was conducted in Wyoming. In-depth guided interviews were conducted with 39 self-identified gay men. Data were analyzed for emergent themes using constant comparative analysis. FINDINGS: Four broadly related themes emerged. Participants perceive that they live in a hostile social environment in which the potential for becoming a target of violence is present. In order to cope with this social reality, men adopt strategies to assimilate into the predominant heterosexual culture and to look for sex partners. These, in turn, are related to their attitudes about HIV/AIDS and prevention activities. Notably, the Internet was discussed by participants as a means for men to connect to a larger gay society and look for sex partners and as a potential venue to HIV/AIDS prevention programs. CONCLUSIONS: Data provided a number of implications for developing HIV/AIDS prevention programs targeting rural MSM. Especially apparent was the need for programs to be mindful of the desire to keep one's sexual preferences shielded from public knowledge and the effect this may have on recruiting rural MSM to participate in prevention activities. The Internet, because men can access it privately, might provide a venue for prevention projects. PMID- 15667010 TI - Predictors of HIV testing and intention to test among Hispanic farmworkers in South Florida. AB - CONTEXT AND PURPOSE: This study examined the predictors of HIV testing and factors associated with intention to accept a free HIV test among 244 Hispanic migrant/seasonal farmworkers in South Florida. METHODS: Time and space sampling procedures were used to recruit participants in public venues. Bilingual staff interviewed eligible respondents in these settings. FINDINGS: Despite high rates of sexual risk, only 21% of respondents had been tested for HIV. The majority of those tested were females tested during prenatal care. In multivariable logistic regression analyses, being female (odds ratio [OR] = 3.73), having at least 12 years of education (OR = 4.46), earning more than $201 per week (OR = 2.76), and ever having used marijuana (OR = 3.31) were positively associated with having been tested for HIV, while not being documented (OR = 0.24) and having rated one's health as "very good" or "good" (OR = 0.42) were negatively associated with testing. The multivariable predictors of intention to accept a free HIV test were having visited a health care provider and/or an emergency room in the past 12 months (OR = 1.97), having been tested for HIV (OR = 2.36), preferring an HIV test that used a finger stick for specimen collection with results given in 30 minutes (OR = 4.47), and worrying "some" or "a lot" about getting HIV (OR = 3.64). Women (OR = 0.52) were less likely than men to intend to accept a free HIV test. CONCLUSIONS: Our findings highlight the importance of routinely offering HIV testing to sexually active individuals in high HIV prevalence areas. They also suggest the need to make testing more accessible to migrant and seasonal farmworkers. PMID- 15667011 TI - Analysis of prior health system contacts as a harbinger of subsequent fatal injury in American Indians. AB - CONTEXT: Many American Indian nations, tribes, and bands are at an elevated risk for premature death from unintentional injury. Previous research has documented a relationship between alcohol-related injury and subsequent injury death among predominately urban samples. The presence or nature of such a relationship has not been documented among American Indians living in the northern plains. PURPOSE: The purpose of this study was to identify and characterize any association between prior injury and/or alcohol use contacts with the Indian Health Service (IHS) and subsequent alcohol-related injury death that may suggest opportunities for mitigation. METHODS: Death certificates of American Indians who died from injury (ICD-9-E 800-999) in a rural IHS area over 6 consecutive years were linked to IHS acute-care facility records and toxicology reports. Deaths and prior IHS contacts were stratified by alcohol use as a contributing factor. Of the 526 injury deaths involving American Indians in the IHS area studied, 411 (78%) were successfully linked to IHS records. One hundred fifty-two of these cases met the inclusion criteria, with an additional 98 cases identified as a comparison group. FINDINGS: No differences in alcohol use at time of death between groups with and without prior health care contact (for injury or alcohol) could be determined (81% vs 73%). A significant relationship was found between previous visits for acute or chronic alcohol use and subsequent alcohol-related fatalities (P =.01). CONCLUSIONS: Based on these findings, injury-prevention activities in the population studied should be initiated at the time of any health-system contact in which alcohol use is identified. Intervention strategies should be developed that convey the immediate risk of death from injury in these patients. PMID- 15667012 TI - A comparison of ATV-related behaviors, exposures, and injuries between farm youth and nonfarm youth. AB - CONTEXT: All terrain vehicles (ATVs) are a popular form of transportation and recreation for youth. ATVs are also convenient for farm-related activities. However, the impact of the farming environment on ATV-related injuries is not clear. PURPOSE: To determine differences in ATV-related behaviors, exposures, risk factors, and injuries between farm youth and their nonfarm peers. METHODS: A cross-sectional study design was used. A survey was administered to 652 youths in agricultural education programs throughout the state of Arkansas. RESULTS: A majority (60%) of students have operated ATVs within the past month. Cross tabulations found that farm youth who rode ATVs were more likely to be white and male, to own a 3-wheel ATV, and to ride more often with a single rider. Risk factors for sustaining an ATV-related injury were frequency of use and the number of persons on the ATV. CONCLUSIONS: Study findings suggest that ATV use among farm youth does differ from their nonfarm peers. ATV use among all youth is a safety concern in Arkansas because of the behaviors and exposures that the youth cited. PMID- 15667014 TI - Perceptions of local health care quality in 7 rural communities with telemedicine. AB - CONTEXT: Rural health services are difficult to maintain because of low patient volumes, limited numbers of providers, and unfavorable economies of scale. Rural patients may perceive poor quality in local health care, directly impacting the sustainability of local health care services. PURPOSE: This study examines perceptions of local health care quality in 7 rural, underserved communities where telemedicine was implemented. This study also assesses factors associated with travel outside of local communities for health care services. METHODS: Community-based pretelemedicine and posttelemedicine random telephone surveys were conducted in 7 northern California rural communities assessing local residents' perceptions of health care quality and the frequency of travel outside their community for health care services. Five-hundred rural residents were interviewed in each of the pretelemedicine and posttelemedicine surveys. Between surveys, telemedicine services were made available in each of the communities. FINDINGS: Residents aware of telemedicine services in their community had a significantly higher opinion of local health care quality (P =.002). Satisfaction with telemedicine was rated high by both rural providers and patients. Residents with lower opinions of local health care quality were more likely to have traveled out of their community for medical care services (P =.014). CONCLUSIONS: The introduction of telemedicine into rural communities is associated with increases in the local communities' perception of local health care quality. Therefore, is it possible that telemedicine may result in a decrease in the desire and need for local patients to travel outside of their community for health care services. PMID- 15667013 TI - Chronic hepatitis C infection in a rural Medicaid HMO. AB - CONTEXT: Chronic hepatitis C infection (CHCI) is an increasingly common problem, affecting about 2% of the US population. The cost and complexity of treatment and difficulties in communicating with the infected population are of concern to insurers and health planners. PURPOSE: To describe the clinical features of patients with CHCI in a rural Medicaid-covered population and to describe a method developed for treating CHCI in an underserved rural community. METHODS: We developed a disease management approach to patients with CHCI receiving insurance coverage through a Medicaid HMO in rural Oregon. A locally based multidisciplinary hepatitis committee was formed to develop a management protocol and a process for selecting patients for treatment. The committee met monthly to develop the treatment plan for individual patients. Day-to-day treatment was provided by a nurse under the supervision of the committee. FINDINGS: One hundred forty-three adults with CHCI were identified by their primary care physicians. About half the patients had a type 1 genotype. Treatment with pegylated interferon and ribavirin was completed on 21 persons, 11 (52%) of whom had a virologic cure. Problems with treatment toxicity were common. Patient satisfaction with the treatment by the nurse was high. CONCLUSIONS: CHCI is common in this rural, nonminority Medicaid-insured population. A locally based disease management model was developed that was well received by patients and was successful in delivering a high quality of care for people with CHCI in a rural area. PMID- 15667017 TI - Overcoming ageism, one student at a time. PMID- 15667015 TI - Recruiting and retaining mental health professionals to rural communities: an interdisciplinary course in Appalachia. AB - CONTEXT: Faculty from 5 disciplines (health administration, nursing, psychology, social work, and special education) collaborated to develop and teach a distance learning course designed to encourage undergraduate and graduate students to seek mental health services employment in rural areas and to provide the skills, experience, and knowledge necessary for successful rural practice. METHODS: The primary objectives of the course, developed after thorough review of the rural retention and recruitment literature, were to (1) enhance interdisciplinary team skills, (2) employ technology as a tool for mental health practitioners, and (3) enhance student understanding of Appalachian culture and rural mental health. Didactic instruction emphasized Appalachian culture, rural mental health, teamwork and communication, professional ethics, and technology. Students were introduced to videoconferencing, asynchronous and synchronous communication, and Internet search tools. Working in teams of 3 or 4, students grappled with professional and cultural issues plus team process as they worked through a hypothetical case of a sexually abused youngster. The course required participants to engage in a nontraditional manner by immersing students in Web based teams. FINDINGS: Student evaluations suggested that teaching facts or "content" about rural mental health and Appalachian culture was much easier than the "process" of using new technologies or working in teams. CONCLUSIONS: Given that the delivery of mental health care demands collaboration and teamwork and that rural practice relies increasingly more on the use of technology, our experience suggests that more team-based, technology-driven courses are needed to better prepare students for clinical practice. PMID- 15667016 TI - Discussion suppers as a means for community engagement. AB - PURPOSE: This paper describes how Lehigh Valley Hospital and Health Network (LVHHN), a large tertiary care urban hospital, used discussion suppers as a means for community engagement designed to lead to community health improvement. An overview of the implementation of the project is described. PROJECT: In 1996, with an awareness of the need to address population-based health improvement, the Dorothy Rider Pool Health Care Trust and LVHHN undertook a multiyear, multidimensional effort to improve health and quality of life in the Lehigh Valley of Pennsylvania. Data were obtained via a series of community and health assessments. Action-Oriented Community Diagnosis and the Behavioral Risk Factor Surveillance System survey, a national instrument, are 2 assessments discussed. The community was engaged through a series of discussion suppers in which community data were shared in a friendly, interactive fashion. The process included community definition of priorities from the data and the subsequent determination of corresponding actions (programs). CONCLUSIONS: The success of these activities demonstrates the discussion suppers were an effective approach and that data can be shared with rural areas in ways that build partnerships and provide a basis for joint actions. This is increasingly important as communities expect our health care systems to provide care both within the hospital as well as outside its walls. PMID- 15667018 TI - Advances in antiepileptic drug treatments. A rational basis for selecting drugs for older patients with epilepsy. AB - Incidence of epilepsy increases rapidly after age 65; recent studies indicate that approximately 10% of nursing home residents are being treated with antiepileptic drugs (AEDs). Almost all are being treated with first generation AEDs. The average nursing home patient receives six medications, has age-related changes in protein binding, decreases in hepatic and renal clearance, and may have alterations in gastrointestinal absorption. AEDs that do not have drug-drug interactions, are not metabolized by the liver, and are readily absorbed may offer benefits in this population. New studies are demonstrating that the first generation AEDs have a number of shortcomings for treating older patients, whereas some of the newer AEDs may overcome these limitations. This paper reviews the present knowledge base and compares properties of the first generation AEDs with newer agents to develop a more rational approach for drug selection in older adults. PMID- 15667019 TI - Inflammatory disease in older adults. Cranial arteritis. AB - Cranial arteritis (CA), also called giant cell arteritis or temporal arteritis, is a vasculitis primarily affecting adults over age 50. It is a large vessel vasculitis, and giant cells classically can be identified on histopathologic examination of temporal arteries, but are not essential for diagnosis. Patients typically present with severe headaches, fatigue, polymyalgia-like symptoms, or ischemic complaints such as jaw claudication. Visual loss is the major feared irreversible outcome and can occur in up to 50% of those with untreated disease. Glucocorticoids, typically high dose prednisone (> or = 60 mg/d) is the first line treatment and successfully controls the inflammatory disease in the vast majority of patients. Most patients can be tapered off steroids within 6 months to 2 years. PMID- 15667020 TI - Behavioral disturbances in dementia: finding the cause(s). PMID- 15667021 TI - The Flying Doctor Service of Australia founded by John Flynn. PMID- 15667022 TI - The future of breast cancer diagnosis: molecular breast imaging. PMID- 15667023 TI - On improving hypertension treatment success. PMID- 15667024 TI - An overview of transforming changes at Mayo Clinic Proceedings during 2005. AB - For the past 6 years, the Editorial Board and staff of Mayo Clinic Proceedings have dedicated ourselves to the progressive evolution of this journal. During this period, and for the foreseeable future, we have embraced change and improvement as themes for our daily efforts. As such, the changes identified in this communication are not a leap forward for the journal, they merely represent the next logical step in the growth of the journal. Given the momentum that has developed during the past few years, readers should expect to see further changes, all intended to better communicate with our principal readers, practicing physicians. In the final analysis, we are driven by a rededication of the journal to the primary value of Mayo Clinic: "The needs of the patient come first." While the journal's leadership has long looked to this statement for guidance, our approach to addressing this value in 2005 will require a considerable number of innovations. We look forward to sharing with you, our readers, these next steps in the journal's history. We trust that we will be able to learn and grow together so that, in the final analysis, the journal will build on its reputation as an authoritative and respected source of information. PMID- 15667025 TI - Molecular breast imaging: a new technique using technetium Tc 99m scintimammography to detect small tumors of the breast. AB - OBJECTIVE: To determine the sensitivity of molecular breast imaging (MBI) to detect small cancers of the breast. PATIENTS AND METHODS: A cadmium-zinc telluride gamma camera with a field of view of 20 x 20 cm was used. The detector elements were 2.5 x 2.5 mm. The gamma camera was mounted on a modified mammographic gantry. Between November 2001 and March 2004, we performed MBI on patients who were scheduled to undergo biopsy for a lesion suggestive of malignancy that was smaller than 2 cm on a mammogram. Patients were injected with 20 mCi of technetium Tc 99m sestamibi and underwent imaging immediately after injection. Using light pain-free compression, we obtained craniocaudal and mediolateral oblique views of each breast. RESULTS: Of the 40 women included in the study, 26 had a total of 36 malignant lesions confirmed at surgery. Of these 36 lesions, 33 were detected by MBI (overall sensitivity, 92%). Of the 22 malignant lesions 1 cm or smaller in diameter, 19 were detected by MBI (sensitivity, 86%). Two patients had false-negative MBI results. Of the 14 malignant lesions larger than 1 cm in diameter, all were identified correctly by MBI. In 4 patients, MBI identified additional lesions not seen on mammography that were confirmed subsequently on magnetic resonance imaging and were true positive cases at surgery. Three of these patients had lesions in the breast contralateral to the breast containing the initial mammographic finding suggestive of malignancy. Of 14 patients with no evidence of cancer at biopsy or surgery, 9 had true-negative (normal) scans and 5 had false-positive scans on MBI. False-positive results included benign fibroadenoma (2 patients), inflammatory fat necrosis (1 patient), benign breast parenchyma (1 patient), and complex sclerosing lesion (1 patient). CONCLUSION: This prototype gamma camera system for MBI reliably detects malignant breast lesions smaller than 2 cm. Furthermore, we obtained the highest sensitivity (86%) yet reported for the detection of lesions smaller than 1 cm. These results suggest an important role for MBI, particularly for women in whom the sensitivity of mammography is reduced by the density of the breast parenchyma. PMID- 15667026 TI - Improved blood pressure control with a physician-nurse team and home blood pressure measurement. AB - OBJECTIVE: To assess whether a physician-nurse team model could improve long-term hypertension control rates by active intervention and modification of antihypertensive drug regimens based on home blood pressure (BP) measurements. PATIENTS AND METHODS: This study consisted of patients referred to a hypertension specialty clinic between July 1999 and June 2002 for the evaluation and management of uncontrolled hypertension. Patients were evaluated initially by a physician. A treatment plan was designed and implemented subsequently by a hypertension nurse specialist. Each patient was given an automated digital home BP monitor and requested to provide 42 BP readings taken during 7 days at intervals of 1, 3, 6, 9, and 12 months after dismissal from the clinic. The mean of these weekly values was reviewed by the physician-nurse team, and the treatment regimen was adjusted to achieve a goal BP of less than 135/85 mm Hg. RESULTS: One hundred six consecutively referred patients were enrolled in the study (mean+/-SD age, 64+/-14 years; 58% female; baseline BP, 156+/-16/85+/-11 mm Hg). Ninety-four patients submitted BP data after 1 month, and 78 patients completed the entire 12-month study period. Overall, mean BP decreased to 138+/ 17/78+/-8 mm Hg at 1 month and to 131+/-9/75+/-7 mm Hg at 12 months (P<.01 vs baseline). The percentage of patients who achieved BP control to less than 135/85 mm Hg increased from 0% at baseline to 63% at 12 months. Intensification of antihypertensive drug therapy was required, on average, in 24% of patients at each study interval. The mean number of drugs increased from 1.2 at baseline to 2.0 at 12 months (P<.01). CONCLUSION: The use of home BP measurement by a physician-nurse team has the potential to significantly improve long-term hypertension control rates in a geographically dispersed patient population. This model should reduce both cost and inconvenience associated with the treatment of hypertension. PMID- 15667027 TI - Idiopathic pulmonary fibrosis in patients younger than 50 years. AB - OBJECTIVE: To assess clinical, radiological, histopathologic, and prognostic differences in younger patients with idiopathic pulmonary fibrosis (IPF). PATIENTS AND METHODS: This study consisted of patients younger than 50 years with IPF who were seen at the Mayo Clinic in Rochester, Minn, from January 1, 1994, to December 31, 2000. Clinical, radiological, and histopathologic data were abstracted from clinical records. Total lung capacity using plethysmography, vital capacity, diffusing capacity for carbon monoxide using the single-breath method, and alveolar volume using single-breath neon wash-in were measured. RESULTS: Our study population was composed of 16 men and 6 women with a median age of 45 years (range, 28-49 years). Median survival was 2.1 years, with 1- and 2-year survival rates of 68% and 53%, respectively. Of the 22 patients, 2 were current smokers, 14 were former smokers, and 6 had never smoked. Three patients had familial pulmonary fibrosis, 21 had bibasilar inspiratory crackles, and 10 had digital clubbing. Median total lung capacity was 56.2%, vital capacity was 51.0%, diffusing capacity was 45.5%, and alveolar volume was 65.0%. CONCLUSION: Although previous studies have suggested that younger age is a favorable prognostic factor in patients with IPF, we found that younger patients have the same poor prognosis as do older patients with this disorder. We observed no distinguishing differences in the clinical, radiological, and histopathologic features compared with those in older patients in whom the disorder is more common. Lung transplantation should be considered early in the treatment of younger patients with IPF. PMID- 15667028 TI - Effectiveness of intravenous immunoglobulin therapy for skin disease other than toxic epidermal necrolysis: a retrospective review of Mayo Clinic experience. AB - OBJECTIVE: To examine retrospectively the use and effectiveness of intravenous immunoglobulin (IVIg) treatment of various skin diseases, primarily immunobullous disease. PATIENTS AND METHODS: We identified patients who had received IVIg therapy for skin disease between 1996 and 2003 at the Mayo Clinic in Rochester, Minn, Scottsdale, Ariz, and Jacksonville, Fla, and retrospectively reviewed their medical records. RESULTS: Eighteen patients were treated with IVIg for various skin diseases: immunobullous disease in 11 adults (pemphigus vulgaris [7 patients], bullous pemphigold [3], and cicatricial pemphigoid [1]); dermatomyositis (2); mixed connective tissue disease (1); chronic urticaria (1); scleromyxedema (1); leukocytoclastic vasculitis (1); and linear IgA bullous disease (1). Responses of patients by type of disease were as follows: pemphigus vulgaris, 1 partial response (PR) and 6 no response (NR); bullous pemphigoid, 1 complete response (CR) and 2 NR; cicatricial pemphigoid, 1 NR; dermatomyositis, 1 CR and 1 PR; mixed connective tissue disease, 1 CR; chronic urticaria, 1 CR; scleromyxedema, 1 CR; leukocytoclastic vasculitis, 1 PR; and linear IgA bullous disease, 1 CR. Six patients (33%) experienced CR, 3 (17%) had PR, and 9 (50%) had NR to IVIg therapy. All 9 nonresponders were adult patients with immunobullous disease. CONCLUSION: Although this was a retrospective study of a small cohort of a mixture of patients, the findings emphasize that our experience with IVIg treatment for skin disease, particularly immunobullous disease, is less favorable than that reported previously. Further studies are needed to verify the efficacy of IVIg for skin disease. PMID- 15667029 TI - Ethnicity and peripheral arterial disease. AB - OBJECTIVE: To determine whether race/ethnicity is an independent risk factor for peripheral arterial disease (PAD). PATIENTS AND METHODS: From September 2000 through August 2001, we screened patients (age > or = 55 years) for PAD within 4 primary care clinics located in the Houston, Tex, area. Variables that were bivariately associated with PAD (P< or = .05) were selected for entry into a multivariate logistic regression model to determine the independent risk factors for PAD. RESULTS: Among 403 patients (136 white, 136 African American, and 131 Latino patients, 81 of whom were Spanish speaking), the prevalence of PAD was 22.8% among African American patients, 13.7% among Latino patients, and 13.2% among white patients (P = .06). Within the multivariate model, adjusting for age, smoking status (odds ratio [OR], 2.58; 95% confidence interval [CI], 1.27-5.25), diabetes mellitus (OR, 2.98; 95% CI, 1.58-5.63), hypertension (OR, 2.58; 95% CI, 1.12-5.95), and education, African American and Latino patients were not more likely than white patients to have a diagnosis of PAD (OR 1.89, 95% CI 0.89-3.99 and OR 1.54, 95% CI 0.59-4.06, respectively). CONCLUSION: After adjusting for atherosclerotic risk factors and level of education, ethnicity was not an independent risk factor for PAD. When determining ethnic variation in outcomes among patients with PAD, efforts are needed to better understand the role of the primary care setting to reduce the burden of social inequality on health. PMID- 15667030 TI - Use of complementary and alternative medical therapies by patients referred to a fibromyalgia treatment program at a tertiary care center. AB - OBJECTIVE: To evaluate the frequency and pattern of complementary and alternative medicine (CAM) use in patients referred to a fibromyalgia treatment program at a tertiary care center. PATIENTS AND METHODS: Patients referred to the Mayo Fibromyalgia Treatment Program between February 2003 and July 2003 were invited on their initial visit to participate in a survey regarding CAM use during the previous 6 months. An 85-question survey that addressed different CAM domains was used. RESULTS: Of the 304 patients invited to participate, 289 (95%) completed the survey (263 women and 26 men). Ninety-eight percent of the patients had used some type of CAM therapy during the previous 6 months. The 10 most frequently used CAM treatments were exercise for a specific medical problem (48%), spiritual healing (prayers) (45%), massage therapy (44%), chiropractic treatments (37%), vitamin C (35%), vitamin E (31%), magnesium (29%), vitamin B complex (25%), green tea (24%), and weight-loss programs (20%). CONCLUSION: CAM use is common in patients referred to a fibromyalgia treatment program. PMID- 15667031 TI - Genetic testing: practical, ethical, and counseling considerations. AB - Genetic testing is becoming a much more common practice in medicine today. This presents a unique set of challenges for medical professionals in virtually all specialties. The practical aspects of determining which test to order, and in interpreting the result accurately in the context of the family history, can be difficult. Additionally, the ethical conundrums that frequently present themselves when genetic risk assessment and/or genetic testing is being considered can be daunting. These challenges present real concerns for medical professionals and patients alike. Included in this article is a review of some of the practical and ethical complexities associated with genetic testing. Pretest and posttest genetic counseling is also emphasized as an important and essential process in today's medical practice. PMID- 15667032 TI - Group B streptococcus endocarditis with endophthalmitis. PMID- 15667033 TI - Blood eosinophilia: a new paradigm in disease classification, diagnosis, and treatment. AB - Acquired blood eosinophilia is considered either a primary or a secondary phenomenon. Causes of secondary (ie, reactive) eosinophilia include tissue invasive parasitosis, allergic or inflammatory conditions, and malignancies in which eosinophils are not considered part of the neoplastic process. Primary eosinophilia is classified operationally into 2 categories: clonal and idiopathic. Clonal eosinophilia stipulates the presence of either cytogenetic evidence or bone marrow histological evidence of an otherwise classified hematologic malignancy such as acute leukemia or a chronic myeloid disorder. Idiopathic eosinophilia is a diagnosis of exclusion (ie, not secondary or clonal). Hypereosinophilic syndrome is a subcategory of idiopathic eosinophilia; diagnosis requires documentation of both sustained eosinophilia (absolute eosinophil count > or = 1500 cells/microL for at least 6 months) and target organ damage (eg, involvement of the heart, lung, skin, or nerve tissue). Genetic mutations involving the platelet-derived growth factor receptor genes (PDGFR alpha and PDGFR-beta) have been pathogenetically linked to clonal eosinophilia, and their presence predicts treatment response to imatinib. Accordingly, cytogenetic and/or molecular investigations for the presence of an imatinib sensitive molecular target should accompany current evaluation for primary eosinophilia. In the absence of such a drug target, specific treatment is dictated by the underlying hematologic malignancy in cases of clonal eosinophilia; however, the initial treatment of choice for symptomatic patients with hypereosinophilic syndrome is prednisone and/or interferon alfa. PMID- 15667034 TI - Role of endothelin in the pathogenesis of hypertension. AB - In 1985, investigators characterized a potent vasoconstrictor of endothelial origin called endothelin (ET). Subsequently, 3 peptides were recognized that had a comparable molecular structure but different receptors that mediate potent vasoconstrictive and mild vasodilative effects. The renal effects are characterized by natriuresis despite renal vasoconstriction. This effect, along with the stimulation of ET by high sodium intake, suggests that ET may be responsible for maintaining sodium balance when the renin-angiotensin system is depressed. Endothelin is activated in desoxycorticosterone acetate salt hypertension models and salt-sensitive hypertension. However, ET involvement with spontaneous hypertension models and renovascular hypertension in rats appears minimal. In humans, the role of ET appears similar to that in experimental animals; in both, ET regulates salt metabolism. Salt-sensitive patients exhibit a blunted renal ET-1 response during sodium load. The role of ET in humans has been investigated using nonspecific ET receptor blockers that inhibit the vasoconstrictive and vasodilative components of ET. However, the effects of ET blockade should be investigated with ET subtype A receptor blockers that mediate vasoconstriction alone. Effects of ET blockade also should be evaluated with respect to stimulation of oxidative stress and tissue damage, important mechanisms responsible for tissue fibrosis. This review offers the clinician a balanced view on the hypertensive mechanisms involved with activation of ET and associated clinical implications. PMID- 15667035 TI - The indolent natural history of essential thrombocythemia: a challenge to new drug development. PMID- 15667037 TI - Atopic dermatitis. PMID- 15667036 TI - Thyrotoxic periodic paralysis. AB - Thyrotoxic periodic paralysis (TPP), a hyperthyroidism-related hypokalemia and muscle-weakening condition resulting from a sudden shift of potassium into cells, has been seen increasingly in Western countries. Failure to recognize TPP may lead to improper management. Many patients with TPP have no obvious symptoms related to hyperthyroidism. Therefore, several important clues may help in diagnosing and managing TPP: presentation in an adult male with no family history of periodic paralysis; presence of systolic hypertension, tachycardia, high QRS voltage, first-degree atrioventricular block on electrocardiography; presence of low-amplitude electrical compound muscle action potential on electromyography and no notable changes in amplitudes after low doses of epinephrinine; and typical acid-base and electrolyte findings such as normal blood acid-base state, hypokalemia with low urinary potassium excretion, hypophosphatemia associated with hypophosphaturia, and hypercalciuria. Immediate therapy with potassium chloride supplementation may foster a rapid recovery of muscle strength, but with a risk of rebound hyperkalemia. Nonselective beta-blockers may provide an alternative choice. Long-term therapy with definite control of hyperthyroidism completely abolishes attacks. Early diagnosis and prompt treatment of TPP prevent life-threatening complications of this treatable and curable disorder. PMID- 15667038 TI - 38-year-old man with priapism and a history of paranoid schizophrenia. PMID- 15667039 TI - Large cell carcinoma with calcitonin and vasoactive intestinal polypeptide associated Verner-Morrison syndrome. AB - Verner-Morrison syndrome, characterized by diarrhea, hypokalemia, and hypochlorhydria, is caused most commonly by vasoactive intestinal polypeptide secreting islet cell tumors of the pancreas. Verner-Morrison syndrome has not been described as a paraneoplastic syndrome in non-small cell lung cancer. We describe a 38-year-old man with metastatic non-small cell lung cancer of large cell carcinoma with neuroendocrine differentiation who presented with bone metastasis and intractable secretory diarrhea that was unresponsive to pharmacological treatment, including octreotide. Laboratory evaluation indicated elevated serum calcitonin and vasoactive intestinal polypeptide levels. Chemotherapy resulted in a transient response in the patient's diarrhea and neuroendocrine markers. The patient did not respond to further therapy and died 5 months after onset of back pain. To our knowledge, this is the first published case of large cell carcinoma with neuroendocrine differentiation associated with treatment-responsive paraneoplastic Verner-Morrison syndrome. PMID- 15667040 TI - Genetics of cerebrovascular disorders. AB - Physicians must be able to recognize stroke caused by a mendelian or mitochondrial disorder. Some genetic disorders such as sickle cell anemia and Fabry disease have proven disease-specific treatments, whereas others have no effective treatment, including cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL) and mitochondrial encephalopathy, lactic acidosis, and stroke-like episodes (MELAS). Proper diagnosis of a genetic disorder has prognostic value and prevents patient exposure to unnecessary and potentially harmful therapeutic agents and diagnostic tests. This article reviews the clinical and genetic features of some mendellan and mitochondrial disorders associated with ischemic stroke, hemorrhagic stroke, and cerebrovascular malformations. PMID- 15667041 TI - Thalidomide--cost and development. PMID- 15667042 TI - Plastic surgery after bariatric surgery and massive weight loss. PMID- 15667043 TI - Neurologic complications of bariatric surgery. PMID- 15667044 TI - How internet users find, evaluate, and use online health information: a cross cultural review. AB - The Internet has become a favored source to find health information. Worldwide, about 4.5% of all Internet searches are for health-related information. However, research has found that the quality of online health information is mixed, which raises serious concerns about the impact of this information. This paper reviews relevant research to understand how health information on the Internet is retrieved, evaluated, and used. Most users of online health information are looking for information about specific health conditions because they or someone they know was diagnosed with a medical condition. They typically use general search engines to find online health information and enter short phrases, often misspelled. They seldom go beyond the first page of a search. Both their search and evaluation skills are limited although they are concerned about the quality of online health information. They avoid sites with overt commercialism, but often do not pay attention to indicators of credibility. Online health information is used to fill an information void which can enhance coping and self efficacy, affects health-related decisions and behavior of users and their friends and family, and is often discussed with health care providers. There are cross-cultural differences in the types of sites used as well as how online information is used. Based on the research reviewed in this paper, three major recommendations are suggested. Professionals should recommend sites. Professionals should promote more effective search and evaluation techniques. Professionals should be involved in developing and promoting uniform standards for health and mental health sites. PMID- 15667045 TI - Online data collection from video game players: methodological issues. AB - The paper outlines the advantages and disadvantages of using the Internet to collect data concerning both online and offline gamers. Drawing from experience of a number of studies carried out online by the authors and by reviewing the available literature, the authors discuss the main issues concerning data collected from video game players. The paper examines a number of areas, including recruiting and utilizing participants, validity, suitable methods of data collection (i.e., questionnaire studies, online tests, participant observation, online interviews), and ethical issues. It is concluded that online research methods can be a useful way of examining the psychosocial aspects of video game playing. PMID- 15667046 TI - Neurofeedback training with virtual reality for inattention and impulsiveness. AB - In this research, the effectiveness of neurofeedback, along with virtual reality (VR), in reducing the level of inattention and impulsiveness was investigated. Twenty-eight male participants, aged 14-18, with social problems, took part in this study. They were separated into three groups: a control group, a VR group, and a non-VR group. The VR and non-VR groups underwent eight sessions of neurofeedback training over 2 weeks, while the control group just waited during the same period. The VR group used a head-mounted display (HMD) and a head tracker, which let them look around the virtual world. Conversely, the non-VR group used only a computer monitor with a fixed viewpoint. All participants performed a continuous performance task (CPT) before and after the complete training session. The results showed that both the VR and non-VR groups achieved better scores in the CPT after the training session, while the control group showed no significant difference. Compared with the other groups, the VR group presented a tendency to get better results, suggesting that immersive VR is applicable to neurofeedback for the rehabilitation of inattention and impulsiveness. PMID- 15667047 TI - Clinically significant virtual environments for the treatment of panic disorder and agoraphobia. AB - The aim of this work is the description of virtual environments designed to apply exposure therapy in the treatment of panic disorder and agoraphobia. The program allows the simultaneous exposure to two different kinds of stimuli usually avoided by panic disorder and agoraphobia sufferers (external and interoceptive). The characteristics of the virtual environments are described, as well as the target behaviors that can be simulated with these virtual environments. PMID- 15667048 TI - Internet addiction disorder and chatting in the Czech Republic. AB - This study analyzes connections between online chatting and the Internet Addiction Disorder (IAD). In the research are compared two groups of respondents- Czech online chat users and Czech university students. The set of answers designed by Kimberly Young, of the University of Pittsburgh, was the common base for research. The research showed that IAD is known among university students, that some of them spend an enormous time on the Internet, and that many of them feel it is a problem. The research also clearly showed that the most serious chat users more often have problems with IAD than other university students. PMID- 15667049 TI - Presence within a mixed reality environment. AB - Mixed reality environments represent a new approach to creating technology mediated experiences. However, there is a lack of empirical research investigating users' actual experience. The aim of the current exploratory, non experimental study was to establish levels of and identify factors associated with presence, within the framework of Schubert et al.'s model of presence. Using questionnaire and interview methods, the experience of the final performance of the Desert Rain mixed reality environment was investigated. Levels of general and spatial presence were relatively high, but levels of involvement and realness were not. Overall, intrinsic motivation, confidence and intention to re-visit Desert Rain were high. However, age was negatively associated with both spatial presence and confidence to play. Furthermore, various problems in navigating the environment were identified. Results are discussed in terms of Schubert's model and other theoretical perspectives. Implications for system design are presented. PMID- 15667051 TI - Shyness and locus of control as predictors of internet addiction and internet use. AB - The new psychological disorder of Internet addiction is fast accruing both popular and professional recognition. Past studies have indicated that some patterns of Internet use are associated with loneliness, shyness, anxiety, depression, and self-consciousness, but there appears to be little consensus about Internet addiction disorder. This exploratory study attempted to examine the potential influences of personality variables, such as shyness and locus of control, online experiences, and demographics on Internet addiction. Data were gathered from a convenient sample using a combination of online and offline methods. The respondents comprised 722 Internet users mostly from the Net generation. Results indicated that the higher the tendency of one being addicted to the Internet, the shyer the person is, the less faith the person has, the firmer belief the person holds in the irresistible power of others, and the higher trust the person places on chance in determining his or her own course of life. People who are addicted to the Internet make intense and frequent use of it both in terms of days per week and in length of each session, especially for online communication via e-mail, ICQ, chat rooms, newsgroups, and online games. Furthermore, full-time students are more likely to be addicted to the Internet, as they are considered high-risk for problems because of free and unlimited access and flexible time schedules. Implications to help professionals and student affairs policy makers are addressed. PMID- 15667050 TI - Internet use and attitudes towards illicit internet use behavior in a sample of Russian college students. AB - This study assessed Internet use and attitudes toward illicit use of the Internet in a sample of Russian college students. A sample comprised of 198 students was recruited from a university in Moscow. Each participant completed a survey assessing demographic characteristics, Internet use, and attitudes towards engaging in illicit behaviors over the Internet. About half of the students reported that they used the Internet at least several times a year, with 8% reporting daily use of the Internet. Among Internet users, most reported having Internet access either at home or at a friends' home, and 16 % reported having Internet access from work, school, or a computer center. Among Internet users, the main purpose was for school-related activities (60%), followed by e-mail (55%), entertainment (50%), chatting (24%), and searching for pornography (6%). Although most students thought it was inappropriate to read someone else's e mail, use someone else's password or credit card information without their permission, or break into someone's computer, many students did endorse those illicit behaviors. Over a fifth of the students reported that they knew hackers. Forty three percent of students agreed that people make too much fuss about watching videos, movies or downloading music on the Internet without paying. Males were more likely than females to report using the Internet for entertainment purposes (p = 0.006) and were more likely to agree that it was okay to break into someone's computer (p = 0.04). The results of this study suggest that these Russian college students predominately use the Internet to help with their schoolwork, to communicate with others, and for entertainment. These results also suggest that interventions may be useful to change attitudes endorsing illicit uses of the Internet. PMID- 15667053 TI - Factors that influence presence in educational virtual environments. AB - The present article is a part of a project for the measurement of presence in educational virtual environments (VEs), since presence is correlated with higher levels of cognitive performance and emotional development, factors that contribute to knowledge construction. The aim of our study was to investigate the sense of presence of 12-year-old pupils within an educational VE representing an ancient Greek house through a sense of embodiment and the ability to handle task performance, while using various peripheral devices. This is the first report on presence measurement with children, based on the indication that children and adults may apply very unrelated criteria. Our results showed statistically significant differences on the level of tiredness and ease of use in using six different input devices. The combination of the keyboard and mouse and the keyboard on its own were the least tedious and easiest input devices, giving a sense of presence as stated by the pupils. Environmental richness and the high level of interactivity within the VE resulted in a high degree of presence for almost all the pupils. The majority of them felt a sense of presence whilst driving the avatar, indicating that presence is significantly correlated with pupils' degree of association with their virtual bodies. All the pupils felt a sense of presence when wearing the head-mounted display. Our findings are in line with those of other researchers and show evidence of personal, social and environmental presence. PMID- 15667052 TI - Video game addiction in children and teenagers in Taiwan. AB - Video game addiction in children and teenagers in Taiwan is associated with levels of animosity, social skills, and academic achievement. This study suggests that video game addiction can be statistically predicted on measures of hostility, and a group with high video game addiction has more hostility than others. Both gender and video game addiction are negatively associated with academic achievement. Family function, sensation seeking, gender, and boredom have statistically positive relationships with levels of social skills. Current models of video game addiction do not seem to fit the findings of this study. PMID- 15667054 TI - Lifestyles of virtual world residents: living in the on-line game "lineage". AB - This study was conducted to explore the lifestyles of online game players who have adopted the virtual world as part of their life. An online survey was conducted on players of an Internet-based game, Lineage. Lineage is the largest online game where people assume new identities and play various roles in a virtual environment, accommodating over 6 million users worldwide. A total of 4,786 game players participated in this survey, and their lifestyles were compared with their values and attitudes in the virtual world. Upon classification of their real-world lifestyles, their tendencies and desires were compared to lifestyles in the virtual world. This study showed that game players have developed their own distinctive lifestyles, and their lifestyles were a strong criterion for explaining behavior patterns and desires in the virtual world. Lifestyles were classified into three general categories: (1) single oriented player, (2) community-oriented player, and (3) off-Real world player. Each group displayed distinct differences in their values and game activities, as well as in their anti-social behavior tendencies. The differences reflected not only their personality but also their socio-economic status within the virtual world, which is constructed through game activities. This study serves as a model to understand how players from different real-life backgrounds will behave in various game features and how they adopt the virtual world for their new social identities. PMID- 15667055 TI - Bronchial auscultation: an effective adjunct to speech and language therapy bedside assessment when detecting dysphagia and aspiration? AB - Detection of aspiration by bedside examination has frequently been found to be clinically inadequate when compared with videofluoroscopy (VF) as the gold standard. In Doncaster, UK, a new multidisciplinary approach to bedside assessment was devised using physiotherapists (PT) performing bronchial auscultation (BA) in combination with the speech and language therapists' (SLT) clinical examination of dysphagia. In this study 105 patients referred for VF examination of dysphagia were first tested by the BA team. Comparison was made between the results of the VF team and the results of the BA team in classifying the patients as "aspirating" or at "risk of aspirating." A high degree of agreement was found for risk of aspiration (sensitivity 87%), although specificity was low (37%). BA was highly specific (88%) when confirming the absence of aspiration, but sensitivity to the presence of aspiration was 45%. From the 105 patients tested, the BA team would have failed to modify the diet in only one subject who was aspirating and would have unnecessarily modified the diet of 17 subjects. In conclusion, in the sample population of individuals with complex dysphagia, the BA team approach reliably detected patients identified by VF as at risk of aspiration. In the group of patients identified by VF as aspirating, the BA team proved unreliable in detecting the presence of aspiration, although it did reliably identify patients who were not aspirating. BA is a potentially useful clinical tool which requires further research. PMID- 15667056 TI - Effects of motor and sensory stimulation in stroke patients with long-lasting dysphagia. AB - Dysphagia is a common poststroke symptom with negative effects on recovery and rehabilitation. However, the orofacial regulation therapy, developed by Castillo Morales, comprising body regulation and orofacial regulation in combination with a palatal plate application has shown promising results in stroke patients. This therapy is based not only on muscle exercises but also on an improvement of the entire sensory-motor reflex arc involved in normal deglutition, and on the knowledge that the function of face and oropharynx at deglutition is closely interrelated with the entire body posture as well as with appropriate breathing. The treatment concept is relatively unknown to caregivers, partly due to lack of scientific evaluation of treatment results. The present investigation aimed to assess the effect of motor and sensory stimulation in stroke patients with dysphagia persisting for more than six months. Seven patients were evaluated with respect to orofacial and pharyngeal motility and sensory function before and two weeks after a five-week treatment period. The evaluation comprised a swallowing capacity test, a meal observation test, clinical examination of oral motor and sensory function, a velopharyngeal closure test, and videofluoroscopy. In addition, the symptoms were scored by the patients. An overall single-blind estimation showed objective and self-assessed swallowing improvement in all seven patients. Kappa coefficients are calculated on all reliability data, both inter- and intrarater reliabilities. Sensory and motor stimulation seems to be a promising therapy in stroke patients with long-lasting and persistent oropharyngeal dysphagia. PMID- 15667057 TI - Reliability and validity of cervical auscultation: a controlled comparison using videofluoroscopy. AB - Cervical auscultation is experiencing a renaissance as an adjunct to the clinical swallowing assessment. It is a controversial technique with a small evidence base. We have aimed to establish whether cervical auscultation interpretation is based on the actual sounds heard or, in practice, influenced by information gleaned from other aspects of the clinical assessment, medical notes, or previous knowledge. We sought to determine (a) rater reliability and its impact on the clinical value of cervical auscultation and (b) how judgments compare with the "gold standard": videofluoroscopy. Swallow sounds were computer recorded via a Littmann stethoscope. Sounds were sampled from 10 healthy control swallows with no aspiration/penetration and 10 patient swallows with aspiration/penetration, all recorded during simultaneous videofluoroscopy. The system generated sound quality similar to "live" bedside listening, a feature rarely seen in cervical auscultation studies. The 20 sound clips were classified as "normal" or "abnormal" by 19 volunteer speech-language pathologists with experience in cervical auscultation. After at least four weeks, 11 of these judges rated the sounds rerandomized on a new CD. Intrarater reliability kappa ranged from -0.12 to 0.71. Individual reliability did not correlate with years of experience, practice pattern, or frequency of use. Interrater reliability kappa = 0.17. Comparison with radiologically defined aspiration/penetration yielded 66% specificity, 62% sensitivity, and majority consensus gave 90% specificity, 80% sensitivity. There was a significant relationship between individual reliability and true positive rate (r(s) = 0.623, p = 0.040). The reliability of individual judges varied widely and thus, inevitably, agreement between judges was poor. Validity is dependent upon reliability: Improving the poor raters would improve the overall accuracy of this technique in predicting abnormality in swallowing. The group consensus correctly identified 17 of the 20 clips so we may speculate that the swallow sound contains audible cues that should in principle permit reliable classification. PMID- 15667058 TI - Age-related remodeling of the hypopharyngeal constrictor muscle and its subneural apparatuses: a scanning electron microscopical study in rats. AB - Age-related remodeling of the hypopharyngeal constrictor muscle was studied by comparing the morphological features of the subneural apparatus (SNA) of the thyropharyngeal (TP) and cricopha ryngeal (CP) muscles in young and old rats. Scanning electron microscopy revealed that the TP and CP muscles had both gutter- and depression-type SNAs, although different proportions of the two types of apparatus were found in each muscle. In young-adult rats, the gutter-type SNA was predominant in the TP muscle, whereas in the CP muscle the depression type was predominant. By contrast, in old rats, the depression-type SNA was predominant in the TP muscle, while no such transformation of the dominant SNA was noted in the CP muscle. In addition, the number of type IIb and type I muscle fibers was lower and greater, respectively, in older animals compared with those of younger animals. Furthermore, there were more type IIc fibers in the TP muscle of old rats, but no difference in the CP muscle between young and old animals. These findings suggest that the transformation of SNAs and the muscle fibers of the hypopharyngeal constrictor muscle takes place during aging and that their remodeling processes differ in the TP and CP muscles. PMID- 15667059 TI - Laryngopharyngeal reflux: trends in diagnostic interpretation criteria. AB - Laryngopharyngeal reflux (LPR) is becoming recognized as a clinical entity with a variety of presentations distinct from those of gastroesophreflux disease (GERD). However, much uncertainty remains as to what is considered pathologic versus physiologic reflux. The aim of the study was to determine the normal range of pharyngeal reflux (PR) occurring in healthy adults based on pH-monitoring parameters utilized in the DeMeester scoring system for GERD. We have reviewed the current pool of prospective literature examining ambulatory dual-channel pH monitoring study data derived from hypopharyngeal proximal probes in normal adults. From our review we have identified trends in several monitoring parameters based on the DeMeester scoring system for GERD. Our discussion recognizes and accepts the limitations imposed by small sample sizes and the number of healthy individuals that would be required to determine the general adult physiologic range of PR. We also explore the possible need for separate normal PR reference intervals based on age or gender disparities. Additional discussion and the summary address future directions for LPR research notably, (1) identification of the most appropriate research paradigm for LPR (i.e., pH 4 vs. 5), (2) establishing reproducibility for the appropriate LPR research paradigm, and (3) complementary modalities to ambulatory dual-channel pH monitoring for the study of acid and nonacid bolus movement within the esophagus PMID- 15667060 TI - Conservative management of a large postlaryngectomy neopharyngeal diverticulum. AB - Dysphagia is common following total laryngectomy. Postlaryngectomy neopharyngeal diverticulae are known to cause postlaryngectomy dysphagia but are more frequently asymptomatic. We report a case presenting with late-onset postlaryngectomy dysphagia and a suprastomal swelling secondary to such a diverticulum. The patient was managed conservatively. PMID- 15667061 TI - Pattern of tongue pressure on hard palate during swallowing. AB - Contact of the tongue against the hard palate plays an important role in swallowing. This study aimed to clarify the pattern of contact between the tongue and hard palate by analyzing tongue pressure produced in swallowing 15 ml of water by healthy subjects wearing an experimental palatal plate with seven pressure sensors. Tongue pressure was generated initially by close contact with the anteriomedian part of the hard palate, then with the circumferential part, and finally softly with the posteriomedian part. Tongue pressure reached a peak quickly, then decreased gradually before disappearing almost simultaneously at each measured part of the hard palate. Magnitude and duration were significantly larger in the anteriomedian part compared to the other six parts measured, and was significantly smaller in the posteriomedian part. No laterality was found in tongue pressure produced at the circumferential parts of the hard palate. Our findings indicate that the order of tongue contact against each part of the hard palate as well as duration and magnitude of tongue pressure are coordinated precisely during swallowing. These findings could aid assessment of the tongue movement of dysphagic patients during rehabilitation. PMID- 15667062 TI - Improved methodology for measuring timing and pressure of tongue-hard palate apposition in the initiation of swallowing. PMID- 15667063 TI - Prevalence of dysphagia among community-dwelling elderly individuals as estimated using a questionnaire for dysphagia screening. AB - The objective of this cross-sectional study was to determine the prevalence of dysphagia among elderly persons living at home in a community using a questionnaire for dysphagia screening. As the reliability of the questionnaire had not yet been confirmed in an epidemiological study, it was also verified. The relationship between dysphagia and the level of daily living competence was also clarified. The subjects consisted of 1313 elderly persons 65 years and older (575 males and 738 females) living at home in a community. The questionnaire included questions concerning the individual's past history of stroke, and questions for dysphagia screening, competence scoring, and perceived ill health. The reliability of the questionnaire was confirmed after calculating the Cronbach's alpha coefficient to be 0.83. The results of a factor analysis showed that the cumulative contribution rate was 61.8%. The prevalence rate of dysphagia was 13.8%. After matching for age and sex, there were significant differences in the competence scores, history of stroke, and perceived ill health status observed between the group with dysphagia and the group without dysphagia. PMID- 15667065 TI - Ontario rejects Great Lakes diversion proposal. PMID- 15667064 TI - A little string music. PMID- 15667066 TI - Global warming's other effects on the oceans. PMID- 15667067 TI - No silver bullet to replace methyl bromide. PMID- 15667068 TI - Monitoring the response to changing mercury deposition. PMID- 15667069 TI - Methyl tert-butyl ether occurrence and related factors in public and private wells in southeast New Hampshire. AB - The occurrence of methyl tert-butyl ether (MTBE) in water from public wells in New Hampshire has increased steadily over the past several years. Using a laboratory reporting level of 0.2 microg/L, 40% of samples from public wells and 21% from private wells in southeast New Hampshire have measurable concentrations of MTBE. The rate of occurrence of MTBE varied significantly for public wells by establishmenttype; for example, 63% of public wells serving residential properties have MTBE concentrations above 0.2 microg/L, whereas lower rates were found for schools (21%). MTBE concentrations correlate strongly with urban factors, such as population density. Surprisingly, MTBE was correlated positively with well depth for public supply wells. Well depth is inversely related to yield in New Hampshire bedrock wells, which may mean that there is less opportunity for dilution of MTBE captured by deep wells. Another possibility is that the source(s) of water to low-yield wells may be dominated by leakage from potentially contaminated shallow groundwater through near-surface fractures or along the well casing. These wells may also have relatively large contributing areas (due to low recharge at the bedrock surface) and therefore have a greater chance of intersecting MTBE sources. This finding is significant because deep bedrock wells are often considered to be less vulnerable to contamination than shallow wells, and in southeast New Hampshire, wells are being drilled deeper in search of increased supply. PMID- 15667070 TI - Regional comparisons of coastal sediment contamination detected by a biomarker (P450 HRGS; EPA Method 4425). AB - Pollution investigations by the Center for Coastal Monitoring and Assessment of the National Oceanic and Atmospheric Administration (NOAA) have been conducted since 1984 and have incorporated extensive biological and chemical analyses. Since 1993, one of the biological tests utilized in these studies has been the biomarker P450HRGS, which is more recently described as EPA Method 4425. Extracts of sediments are applied to a human cell line with a reporter gene (firefly luciferase) at the CYP1A1 site. Light produced by the extracts is a function of the concentrations and potencies of those compounds with an affinity for Ah receptor (certain polycyclic aromatic hydrocarbons, polychlorinated biphenyls, and dioxins/ furans). These compounds are carcinogenic and can produce chronic toxicity, and those containing chlorine are persistent and bioaccumulated. Nineteen coastal regions and 1309 samples from the three U.S. coasts have been evaluated as part of the NOAA investigations. The stratified random sampling approach used by NOAA provides estimates of the areas (km2) of each region containing levels of the compounds above thresholds. From analysis of the database, sediments with concentrations at or below 11 microg benzo[a]pyrene equivalents (B[a]PEq)/g would not be expected to produce effects on the benthos. At 32 microg B[a]PEq/g and above there is the potential for impacts on the biota, and above 60 microg/g, the degradation of the benthic community has been observed. Several of the regional surveys found no samples at or above 60 microg B[a]PEq/g, but 60% of the samples from New York Harbor (280 km2) were above this level. Analyses of data from Puget Sound and Chesapeake Bay demonstrate an increase in samples above 32 microg B[a]PEq/g in more populated and industrial regions. Method 4425 serves as both a biomarker, simulating the response of an organism (with CYP1A) exposed to inducing compounds, and as a bioanalytical technique measuring the levels of these chemicals in the samples. A targeted investigation of the distribution of the three important classes of compounds identified by Method 4425 would be more cost-effective by first screening samples by this method before expending substantial funds in the detailed chemical analysis of all samples. PMID- 15667071 TI - Impact of the 2002 Canadian forest fires on particulate matter air quality in Baltimore city. AB - With increasing evidence of adverse health effects associated with particulate matter (PM), the exposure impact of natural sources, such as forest fires, has substantial public health relevance. In addition to the threat to nearby communities, pollutants released from forest fires can travel thousands of kilometers to heavily populated urban areas. There was a dramatic increase in forest fire activity in the province of Quebec, Canada, during July 2002. The transport of PM released from these forest fires was examined using a combination of a moderate-resolution imaging spectroradiometer satellite image, back trajectories using a hybrid single-particle Lagrangian integrated trajectory, and local light detection and ranging measurements. Time- and size-resolved PM was evaluated at three ambient and four indoor measurement sites using a combination of direct reading instruments (laser, time-of-flight aerosol spectrometer, nephelometer, and an oscillating microbalance). The transport and monitoring results consistently identified a forest fire related PM episode in Baltimore that occurred the first weekend of July 2002 and resulted in as much as a 30-fold increase in ambientfine PM. On the basis of tapered element oscillating microbalance measurements, the 24 h PM25 concentration reached 86 microg/m3 on July 7, 2002, exceeding the 24 h national ambient air quality standard. The episode was primarily comprised of particles less than 2.5 microm in aerodynamic diameter, highlighting the preferential transport of the fraction of PM that is of greatest health concern. Penetration of the ambient episode indoors was efficient (median indoor-to-outdoor ratio 0.91) such that the high ambient levels were similarly experienced indoors. These results are significant in demonstrating the impact of a natural source thousands of kilometers away on ambient levels of and potential exposures to air pollution within an urban center. This research highlights the significance of transboundary air pollution and the need for studies that assess the public health impacts associated with such sources and transport processes. PMID- 15667072 TI - Levels and distribution of polybrominated diphenyl ethers in water, surface sediments, and bivalves from the San Francisco Estuary. AB - Polybrominated diphenyl ethers (PBDEs) were found in water, surface sediments, and bivalve samples that were collected from the San Francisco Estuary in 2002. sigmaPBDE concentrations in water samples ranged from 3 to 513 pg/ L, with the highest concentrations found in the Lower South Bay (range 103-513 pg/L) region, which receives approximately 26% of the Estuary's wastewater treatment plant effluents. The sigmaPBDEs in sediments ranged from below detection limits to 212 ng/g dry wt, with the highest concentration found at a South Bay station (212 ng/g dry wt), which was up to 3 orders of magnitude higher than other stations. The sigmaPBDE concentrations ranged from 9 to 64 ng/g dry wt in oysters (Crassostrea gigas), from 13 to 47 ng/g dry wt in mussels (Mytilus californianus), and from 85 to 106 ng/g dry wt in clams (Corbicula fluminea). Only three PBDE congeners were detected in bivalves, BDE-47, BDE-99, and BOE-100; these are the most bioaccumulative congeners from the commercial Penta-BDE mixture. PMID- 15667073 TI - Field deployment of thin film passive air samplers for persistent organic pollutants: a study in the urban atmospheric boundary layer. AB - This paper reports on the first field deployment of rapidly equilibrating thin film passive air samplers under ambient conditions. The POlymer-coated Glass (POG) samplers have a coating of ethylene vinyl acetate (EVA) less than 1 microm thick applied to a glass surface. This can be dissolved off after exposure and prepared for the quantification of persistent organic pollutants (POPs) that have partitioned into the film during field exposure. In this study, POGs were deployed at various heights on the CN Tower in Toronto, Canada, to investigate the vertical distribution of selected compounds (PCBs, PAHs, organochlorine pesticides) in the atmospheric boundary layer of an urban area. The feasibility of the method to detect POPs from a few cubic meters of air was demonstrated, indicating the potential for rapid, low-volume sampling of air for ambient levels of POPs. PAH levels declined sharply with height, confirming ground-level emissions in urban areas as sources of these compounds; PCBs did the same, although less strongly. Different sampling events detected different vertical distributions of OC pesticides which could be related to local or distantsources, and variations in POPs on the samplers in these different events/heights demonstrate the dynamic nature of sources and atmospheric mixing of POPs. PMID- 15667074 TI - Abundances, depositional fluxes, and homologue patterns of polychlorinated biphenyls in dated sediment cores from the Pearl River Delta, China. AB - Despite the recent efforts to investigate the distribution and fate of persistent organic pollutants in the tropical and subtropical regions of Asia, very little was known about the temporal change of polychlorinated biphenyls (PCBs) in the environmental ecosystem of China. In this study, three dated sediment cores collected from the Pearl River Delta of southern China were analyzed for a large suite of PCB congeners, from which the temporal profiles of PCB abundances, fluxes, and homologue patterns were constructed. The sedimentary inventories of total PCBs at the sampling sites ranged from 480 to 1310 ng/cm2, at the low end of the worldwide figures. Although production and use of PCBs have been banned or highly restricted in China since the early 1980s, the fluxes of total PCBs continued to increase in the Pearl River Delta sediments. There was a concurrent increase of PCB fluxes and gross domestic product per capita in the region from 1980 to 1997, and a decline of agricultural land use was evident at the same time. Apparently, large-scale land transform since the early 1980s as well as emissions from the PCB-containing electrical equipments were responsible for the sharp rise of PCB fluxes in the recent sediments. The difference in the PCB homologue patterns from 1940 to the mid-1970s was probably indicative of the different timelines of PCB usage in Macao/Hong Kong and mainland China and the differenttypes of technical PCBs commercially used. PCBs were detectable in sediments deposited well before the time frame when production of PCBs began (before 1930) and were relatively enriched in the less chlorinated homologue groups (3Cl and 4Cl PCBs), suggesting the downward mobility of lightly chlorinated PCB congeners in the sediment column. PMID- 15667075 TI - Milwaukee, WI, as a source of atmospheric PCBs to Lake Michigan. AB - A field study of atmospheric PCBs in Milwaukee, WI, U.S.A. was conducted on the shore of Lake Michigan. We believe this is the first report of atmospheric PCBs in Milwaukee, although PCBs are well-known to contaminate the sediments of the Milwaukee River and Outer Harbor. Concentrations of PCBs collected during the June 2001 study are similar to concentrations in other urban-industrial areas and higher than PCBs in background air. The average sigmaPCB (sum of 88 congener groups) gas-phase concentration in Milwaukee was 1.9 ng m(-3)+/-standard deviation 0.78 ng m(-3). The average and standard deviation for the particulate associated PCBs are 0.05+/-0.02 ng m(-3). Particulate-phase PCBs account for less than 5% of the total atmospheric concentration. PCBs in Milwaukee air are a source of PCBs to Lake Michigan. Calculated net gas exchange fluxes predicted forthe Milwaukee sampling period ranged from -60 to -400 ng m(-2) d(-1), where net deposition is indicated by the negative sign. Calculated particle-associated PCB deposition ranged from 80 to 500 ng m(-2) d(-1). Most of the particle-phase deposition flux is a result of coarse particle deposition and decreases rapidly with distance from shore. Under typical meteorological conditions, particle associated PCBs depositional flux to the lake surface decreases by 90% within 40 km. For net gas-exchange, the flux reaches zero at about the same distance. At greater distances, particle-phase PCB deposition is negligible, and PCBs are volatilizing at a higher rate than they are being deposited. We calculated that Milwaukee air contributes about 120 kg of PCBs to Lake Michigan each year. This is about 10 times larger than the discharge of PCBs from the Milwaukee River. PMID- 15667076 TI - Atmospheric reactions influence seasonal PAH and nitro-PAH concentrations in the Los Angeles basin. AB - Ambient measurements of polycyclic aromatic hydrocarbons (PAHs) and nitro-PAHs were carried out during August 2002 and January 2003 in Los Angeles, CA, a source site and in Riverside, CA, a downwind receptor site approximately 90 km to the east of Los Angeles. Atmospheric concentrations of PAHs and nitro-PAHs are of interest because both of these compound classes include potent mutagens and carcinogens. To augment our current understanding of atmospheric formation of nitro-PAHs, four sampling periods were employed to study the diurnal variations of these compounds. The PAH concentrations were highest in Los Angeles during January, as a result of traffic input at this source site undertightwintertime atmospheric inversions. In contrast, nitro-PAH levels were highest in Riverside during August, as a result of enhanced summertime photochemistry. Hydroxyl radical-initiated reactions produced nitro-PAHs in both seasons, while in winter little evidence for nitrate radical chemistry was seen. For the August samples, nitrate radical-initiated formation of nitro-PAHs is suggested by nitro-PAH isomer profiles not only at the downwind location as anticipated, but also atthe source site. In southern California, the contribution of atmospheric formation through gas-phase radical-initiated PAH reactions to the ambient burden of nitro PAHs is dominant, with the semi-volatile nitro-PAHs being the most abundant and 2 nitrofluoranthene being the major particle-associated nitro-PAH. PMID- 15667077 TI - Mass budget of perfluorooctane surfactants in Lake Ontario. AB - Perfluorooctane surfactants have been reported in biota, water, and air samples worldwide. Despite these reports, the main environmental sources of these compounds remain undefined. To address this gap in knowledge, an annual lake-wide mass budget of eight perfluorooctane surfactants was developed for Lake Ontario. To determine the atmospheric contribution to the mass budget, over-the-lake gas phase air concentrations for N-EtFOSE and N-EtFOSA and particulate-phase air concentrations for PFOS in any air sample are reported for the first time, with mean concentrations when present of 0.5+/-0.32 (N-EtFOSE gas-phase), 1.1+/-0.9 (N EtFOSA gas-phase), and 6.4+/-3.3 (PFOS particulate-phase) pg/m3. The mass budget finds inflow from Lake Erie (14 361+/-4489 kg sigma perfluorooctane surfactants) and wastewater discharge (1762+/-2697 kg sigma perfluorooctane surfactants) to be the major sources, while outflow through the St. Lawrence River is the dominant loss mechanism (22,727+/-7060 kg/year sigma perfluorooctane surfactants). Using the mass budget data, the steady state and measured mean concentrations in the lake water are the same at the 95% confidence level. PMID- 15667078 TI - Temporal trends of PFOS and PFOA in guillemot eggs from the Baltic Sea, 1968- 2003. AB - Perfluorooctane sulfonic acid (PFOS) and perfluorooctanoic acid (PFOA) have recently been identified as ubiquitous environmental contaminants. Although they have been produced for 50 years, little is known about when they first appeared in the environment and how their concentrations have changed over time, particularly in response to the phase-out of PFOS, which began in 2000. In this study temporal trends in the concentrations of PFOS and PFOA in the Baltic Sea marine environmentwere measured using archived guillemot eggs. Samples collected from Stora Karlso (Sweden) between 1968 and 2003 were received from an environmental specimen bank and concentrations of PFOS and PFOA were analyzed using HPLC coupled to ESI-MS/MS. PFOA was not detected in any of the samples (LOD 3 ng/g), but there was an almost 30-fold increase in PFOS concentrations in the guillemot eggs during the time period, from 25 ng/g in 1968 to 614 ng/g in 2003 (wet weight). Regression analysis indicated a significant trend, increasing on average between 7 and 11% per year. A sharp peak in PFOS concentrations was observed in 1997 followed by decreasing levels up to 2002, but this cannot be linked to the PFOS phase-out, which occurred at the end of this period. PMID- 15667079 TI - Occurrence, profiles, and photostabilities of chlorinated polycyclic aromatic hydrocarbons associated with particulates in urban air. AB - The atmospheric levels of 12 chlorinated polycyclic aromatic hydrocarbons (ClPAHs) associated with particulates at an urban site in Japan were investigated. Only 7 of the 12 species studied were detected in air samples collected monthly during 2002. 1-Chloropyrene (1-ClPy) was detected at the highest concentration (7.5 pg m(-3) (annual mean)), followed by 6 chlorobenzo[a]pyrene (6-ClBaP; 5.6 pg m(-3)) and 9,10-dichlorophenanthrene (5.1 pg m(-3)). The concentrations of the ClPAHs tended to be higher in winter than in summer, with the exception of the 6-ClBaP concentration, which was high in both summer and winter. Correlation analysis also indicated that the behavior of 6 CIBaP differed significantly from that of the other ClPAHs. Comparison of the atmospheric ClPAH concentration profile normalized to 1-CIPy concentration with that for a traffic air sample revealed significant differences between the profiles. The behavior of the atmospheric ClPAHs was also influenced by the origin of the associated particulates, which varied according to season. The positions of chlorination in the detected ClPAHs were consistent with those where the frontier electron density was high. This means that the atmospheric ClPAHs were formed by secondary reactions with chlorine atoms. The photostabilities of the ClPAHs were also investigated in laboratory experiments using a chemical model system. The ClPAHs decayed according to first-order reaction rate kinetics, with photostabilities increasing in the order 6-ClBaP < 1-ClPy < 7-ClBaA < ClPhe < ClFluor, consistent with the trend for the parent PAHs. The photolyses of chlorophenanthrenes and 7-chlorobenz[a]anthracene were confirmed to proceed by initial abstraction of chlorine, followed by oxidative degradation. PMID- 15667080 TI - Pu and U atom ratios and concentration factors in reservoir 11 and Asanov Swamp, Mayak PA: an application of accelerator mass spectrometry. AB - Mayak Production Association, East Ural, Russia, was established to produce weapons-grade plutonium. Routine discharges and accidents at Mayak PA contaminated large areas, including the Techa River. The objectives of the present work were to study atom ratios for plutonium and, for the first time to our knowledge, uranium isotopes in water, soil, grass, and aquatic biota samples from Reservoir 11 and the Asanov Swamp, downstream from Mayak PA. Atom ratios (240Pu/239Pu, 236U/235U, 235U/238U) were determined using accelerator mass spectrometry to confirm radionuclide source characteristics and calculate activities and concentration factors for the studied samples. The lowest 240Pu/239Pu atom ratios were consistently found in Asanov Swamp samples (approximately 0.019), indicating a major contribution from early discharges of weapons-grade Pu. 240Pu/239Pu atom ratios in Reservoir 11 were higher, indicating influence from more recent civil reprocessing. The presence of 236U is usually indicative of fuel irradiation; 236U/235U ratios increase from weapons to civil sources. Our new data show that Asanov samples had lower 236U/ 235U ratios than Reservoir 11 samples (0.0005-0.0045 for Asanov compared with 0.0074-0.0153 for Reservoir 11) in agreement with Pu results. Pu and U concentration factors calculated for vegetation and biota samples at Mayak were comparable with corresponding values found in the literature. PMID- 15667081 TI - Physiological basis for large differences in resistance to nitrite among freshwater and freshwater-acclimated euryhaline fishes. AB - Uptake of environmental NO2- by most freshwater fishes occurs at the gills where NO2- is actively transported into the blood by the Cl- uptake pathway. Some freshwater fishes do not concentrate NO2- in their plasma, regardless of environmental NO2- exposure and exhibit a high degree of resistance to NO2-. Recent studies indicate that freshwater-adapted killifish (Fundulidae: Fundulus heteroclitus) and European eel (Anguillidae: Anguilla anguilla) have no or minimal Cl- uptake activity at the gills relative to most freshwater fishes; rather, Cl- requirements are met in other ways (probably dietary). We hypothesized that different rates of Cl- uptake by the gill may explain the observed differences in NO2- uptake and consequent toxicity among freshwater fishes. Cl- influx rates of channel catfish (Ictaluridae: Ictalurus punctatus), a species that concentrates NO2- in the plasma and is sensitive to NO2-, and bluegill (Centrarchidae: Lepomis macrochirus), a species that does not concentrate NO2- in the plasma and is resistant to NO2-, were determined over a range of environmental Cl- concentrations. Channel catfish actively transported chloride into the plasma (Km = 155.6+/-101.2 micromol/L Cl-; Jmax = 414.9+/-51.4 nmol/g/h; +/-SEM). In contrast, bluegill exhibited no observable Cl- uptake. We placed our results and previously reported results in a phylogenetic context and concluded that differences in Cl- uptake mechanisms among groups of freshwater fishes may explain, in large part, the wide range of sensitivity to environmental NO2-. NO2- uptake determinations may also prove to be an easy screening method when studying the phylogenetic distribution and nature of Cl- uptake mechanisms in the gills of fishes. PMID- 15667082 TI - Carbon isotopic fractionation during anaerobic biotransformation of methyl tert butyl ether and tert-amyl methyl ether. AB - The fuel oxygenate methyl tert-butyl ether (MTBE) has been frequently detected in groundwater and surface water. Since contaminated sites are often subsurface, anaerobic degradation of MTBE will likely be significant for remediation. As traditional approaches to evaluate biodegradation generally involve laboratory microcosm studies which require time and resources, innovative approaches are needed to demonstrate active in situ biodegradation of MTBE. This study was conducted to gather information at the laboratory level to evaluate the potential of applying carbon isotope fractionation as an indicator for in situ biodegradation of the fuel oxygenates MTBE and tert-amyl methyl ether (TAME). In this study, MTBE utilization was observed in a methanogenic sediment microcosm after a lengthy lag period of about 400 days. MTBE utilization was sustained upon refeeding and subculturing. tert-Butyl alcohol (TBA) was found to accumulate after propagation of cultures. The MTBE-grown cultures also utilized TAME and produced tert-amyl alcohol (TAA). The detection of TBA and TAA indicated that ether bond cleavage was the initial step in degradation for both compounds. Carbon isotope fractionation during anaerobic MTBE and TAME degradation was studied, and isotopic enrichment factors (epsilon) with 95% confidence intervals of -15.6 +/-4.1% and -13.7+/-4.5% were estimated for anaerobic MTBE and TAME degradation, respectively. Addition of 2-bromoethanesulfonic acid, an inhibitor of methanogenesis, substantially prolonged the lag period before transformation, but did not influence carbon isotope fractionation. Our experiment provided strong evidence of significant carbon isotope fractionation during anaerobic MTBE and TAME degradation, demonstrating that this technique can be used as an indicator for in situ MTBE and TAME degradation. PMID- 15667083 TI - Dark oxidation of dissolved and liquid elemental mercury in aquatic environments. AB - Elemental mercury (Hg0) can be found in liquid or dissolved forms in aquatic systems. Whereas dissolved Hg0 is measured in virtually all aquatic systems, liquid Hg0 droplets are mainly observed at poorly lit sediment/water interfaces of ecosystems with local point sources such as hydro-thermal vents, gold extraction sites, and near industrial facilities. Here, we report that, in the dark, liquid and dissolved forms of Hg behave differently with respect to their oxidation. Liquid Hg0 is rapidly oxidized in oxygenated solution in the presence of chloride. Liquid Hg0 oxidation rates are positively correlated with chloride concentrations and droplet surface area. When liquid Hg is removed from solution, the oxidation stops even though the solution is still saturated with dissolved Hg0. Liquid Hg0 droplets in oxygenated marine or brackish environments should be oxidized and release Hg2+ to solution. In freshwaters or anoxic seawater, liquid Hg will dissolve releasing Hg(aq)0 which, itself, will slowly oxidize. PMID- 15667084 TI - High plankton densities reduce mercury biomagnification. AB - This study tests the hypothesis that differences in the densities of plankton across lakes lead to significantvariation in Hg accumulation by the biota. The study comprised (i) a field program to measure and compare total Hg concentrations in phytoplankton (0.4-45 microm), two size classes of zooplankton (45-202 and >202 microm), and fish to phytoplankton and zooplankton densities from 20 lakes in the northeastern United States and (ii) a statistical analysis of fish Hg burdens and plankton densities from 38 additional EMAP lakes in the northeastern United States. We found that phytoplankton density was negatively correlated with Hg concentrations within both the phytoplankton and their primary consumers (small zooplankton). We also found a negative correlation between zooplankton density and Hg concentrations in zooplankton and in both herbivorous and predatory fish. Zooplankton density alone explained more than 40% of the variation in predatory fish Hg levels across lakes. We discuss several mechanisms to explain the negative relationship of plankton density to Hg concentrations in lower trophic levels and to the biomagnification of Hg to fish. Our results suggest that measures of plankton density, which are relatively easy and inexpensive to estimate, may be effective predictors of Hg biomagnification potential across lakes. PMID- 15667085 TI - Mechanisms of dioxin formation from the high-temperature oxidation of 2 chlorophenol. AB - The homogeneous, gas-phase oxidative thermal degradation of 2-chlorophenol was studied in a 1 cm i.d., fused silica flow reactor at a concentration of 88 ppm, reaction time of 2.0 s, over a temperature range of 300 to 1000 degrees C. Observed products in order of yield were as follows: 4,6-dichlorodibenzofuran (4,6-DCDF) > dibenzo-p-dioxin (DD) > 1-monochlorodibenzo-p-dioxin (1-MCDD), 4 chlorodibenzofuran (4-MCDF), dibenzofuran (DF), naphthalene, chloronaphthalene, 2,4-dichlorophenol, 2,6-dichlorophenol, phenol, chlorobenzene, and benzene. In contrast to pyrolysis, 4,6-DCDF is the major product rather than DD, and 1-MCDD and naphthalene are formed at temperatures as low as 400 degrees C. Under oxidative conditions, .OH and Cl. are the major carriers, which favors 4,6-DCDF formation over DD or 1-MCDD through abstraction of H. through diketo- and ether- intermediates. It is proposed that below 500 degrees C, unimolecular tautomerization/HCI elimination and CO elimination/isomerization reactions result in the formation of 1-MCDD and naphthalene, respectively. PMID- 15667086 TI - When aerosol sulfate goes up, so does oxalate: implication for the formation mechanisms of oxalate. AB - Oxalic acid is often the single most abundant water-soluble organic compound identified in ambient aerosols, but its precursors have not been identified, and its formation mechanism is not well understood. On the contrary, sulfate as a major aerosol component, its formation pathways have been established, and in cloud processing is recognized as its major production pathway. Our measurements of aerosol sulfate and oxalate collected across a wide geographical span in the East Asia region, up to Beijing in the north and down to Hong Kong in the south, indicate that the two species are highly correlated among samples collected at the same location and among samples collected at different locations in the Pearl River Delta region. This good correlation is also found in measurements made elsewhere by other researchers. We argue that a common dominant formation pathway, likely in-cloud processing, explains the close tracking of the two chemically distinct species. This also highlights the potential importance of in cloud processing as a pathway leading to formation of secondary organic aerosols. PMID- 15667087 TI - Phenanthrene sorption to sequentially extracted soil humic acids and humins. AB - Humic substances strongly influence the environmental fate of hydrophobic organic chemicals in soils and sediments. In this study, the sorption of phenanthrene by humic acids (HAs) and humins was examined. HAs were obtained from progressively extracting a soil, eight times with 0.1 M Na4P207 and two times with 0.1 M NaOH solution, and then the residue was separated into two humin fractions by their organic carbon contents. The chemical and structural heterogeneity of the HAs and humins were characterized by elemental analysis, ultraviolet-visible spectroscopy, diffuse reflectance infrared Fourier transform spectroscopy, and solid-state 13C NMR. There were significant chemical and structural differences among the HA fractions and humins; the later extracted HAs had relatively high aliphatic carbons content. All sorption data were fitted to a Freundlich equation, S = K(F)C(N), where S and C are the sorbed and solution-phase concentrations, respectively, and K(F) and N are constants. All of the phenanthrene sorptions were nonlinear, and the nonlinearity decreased with further extractions from 0.90 (first extracted HA) to 0.96 (ninth HA) and was the lowest (0.88) for the higher organic carbon content humin. Phenanthrene sorption coefficient by HAs significantly increased with progressive extractions, being the highest for the humins. For HAs isotherms, a positive trend was observed between the sorption coefficient and the aliphaticity, but a negative relation was shown between the nonlinearity and the aliphaticity and between the sorption capacity and polarity of HAs. Phenanthrene sorption was greatly affected by chemical structure and composition of humic substances, even from a same soil. In addition, polarity of humic substances seems to mainly regulate the magnitude of phenanthrene sorption rather than structure. PMID- 15667088 TI - Importance of black carbon to sorption of native PAHs, PCBs, and PCDDs in Boston and New York harbor sediments. AB - The solid-water distribution ratios (Kd values) of "native" PAHs, PCBs, and PCDDs in Boston and New York Harbor sediments were determined using small passive polyethylene samplers incubated for extended times in sediment-water suspensions. Observed solid-water distribution coefficients exceeded the corresponding f(oc)Koc products by 1-2 orders of magnitude. It was hypothesized that black carbon (fBC), measured in the Boston harbor sediment at about 0.6% and in the New York harbor sediment at about 0.3%, was responsible for the additional sorption. The overall partitioning was then attributed to absorption into the organic carbon and to adsorption onto the black carbon via Kd = f(oc)Koc + f(BC)K(BC)C(w)n-1 with Cw in microg/L. Predictions based on published Koc, K(BC), and n values for phenanthrene and pyrene showed good agreement with observed Kd,obs values. Thus, assuming this dual sorption model applied to the other native PAHs, PCBs, and PCDDs, black carbon-normalized adsorption coefficients, K(BC)S, were deduced forthese contaminants. Log K(BC) values correlated with sorbate hydrophobicity for PAHs in Boston harbor (log K(BC) approximately 0.83 log gamma w(sat) - 1.6; R2 = 0.99, N= 8). The inferred sorption to the sedimentary BC phase dominated the solid-water partitioning of these compound classes, and its inclusion in these sediments is necessary to make accurate estimates of the mobility and bioavailability of PAHs, PCBs, and PCDDs. PMID- 15667089 TI - Analysis of column tortuosity for MnCl2 and bacterial diffusion using magnetic resonance imaging. AB - Subsurface bacteria often have to travel significant distances through tortuous porous media for purposes of groundwater remediation. In modeling such processes, motile bacteria are often represented as suspended colloids, ignoring their individual swimming or diffusive properties. In fact, bacterial migration is much more profoundly affected by the presence of porous media than is that of a chemical contaminant. In this study, we use magnetic resonance imaging (MRI) to perform noninvasive measurements of changes in bacterial concentration distributions across a packed column at a spatial resolution of 330 microm as a function of time. We analyze the diffusive behavior of Pseudomonas putida F1 under static conditions and compare that behavior to the diffusion of a chemical solute and of Escherichia coli NR50. Results indicate that P. putida cells experience a column tortuosity 50 times higher than that predicted from solute diffusion experiments. E. coli cells, which display shorter swimming run lengths in bulk solution than P. putida, seem to be less affected by the constricted pore space. Knudsen diffusion, or reductions in run length because of interactions between the diffusing bacteria and the porous media, may help to explain some of this discrepancy. PMID- 15667090 TI - Relative leaching and aquatic toxicity of pressure-treated wood products using batch leaching tests. AB - Size-reduced samples of southern yellow pine dimensional lumber, each treated with one of five different waterborne chemical preservatives, were leached using 18-h batch leaching tests. The wood preservatives included chromated copper arsenate (CCA), alkaline copper quaternary, copper boron azole, copper citrate, and copper dimethyldithiocarbamate. An unpreserved wood sample was tested as well. The batch leaching tests followed methodology prescribed in the U.S. Environmental Protection Agency toxicity characteristic leaching procedure (TCLP). The wood samples were first size-reduced and then leached using four different leaching solutions (synthetic landfill leachate, synthetic rainwater, deionized water, and synthetic seawater). CCA-treated wood leached greater concentrations of arsenic and copper relative to chromium, with copper leaching more with the TCLP and synthetic seawater. Copper leached at greater concentrations from the arsenic-free preservatives relative to CCA. Arsenic leached from CCA-treated wood at concentrations above the U.S. federal toxicity characteristic limit (5 mg/L). All of the arsenic-free alternatives displayed a greater degree of aquatic toxicity compared to CCA. Invertebrate and algal assays were more sensitive than Microtox. Examination of the relative leaching of the preservative compounds indicated that the arsenic-free preservatives were advantageous over CCA with respect to waste disposal and soil contamination issues but potentially posed a greater risk to aquatic ecosystems. PMID- 15667092 TI - Electrochemical study of 2,3-dihydroxybenzoic acid and its interaction with Cu(II) and H2O2 in aqueous solutions: implications for wood decay. AB - The electrochemical behavior of 2,3-dihydroxybenzoic acid (2,3-DHBA) and the electron-transfer characteristics between Cu(II) and 2,3-DHBA were studied in aqueous solutions using cyclic voltammetry (CV). The overall electrochemical oxidation process of 2,3-DHBA by Cu(II) may be classified as a chemical reaction involving one-electron oxidation of 2,3-DHBA to its semiquinone radical in solution, followed by an electron-transfer reaction involving the oxidation of the semiquinone radical to a quinone at the electrode surface. In the presence of H2O2, oxidation of 2,3-DHBA by Cu(II) is enhanced due to the regeneration of Cu(II) by H2O2 oxidizing Cu(I). The redox cycling between Cu(I)/Cu(II) and H2O2 also produces hydroxyl radicals (OH). Even though the presence of OH may not be detected at the surface of a glassy carbon electrode, production of electroactive dissolved oxygen (O2) suggests the presence of OH. The production of O2 is dependent on Cu(II):H2O2 concentration ratio. At the electrode surface and when the initial Cu(II):H2O2 is less than 1, O2 is produced, suggesting that H2O2 may act as a scavenger for OH; at initial Cu(II):H2O2 > 1, the production of O2 is not favored, and OH will be involved in the oxidation of Cu(I) and the organic ligand. The reaction mechanisms proposed in this study indicate that OH production by chelator-mediated Fenton reactions is favorable under conditions found in the wood cell wall. PMID- 15667091 TI - Semiempirical model for organic aerosol growth by acid-catalyzed heterogeneous reactions of organic carbonyls. AB - Aerosol growth by heterogeneous reactions of diverse carbonyls in the presence and absence of acidified seed aerosols was studied in a 4 m long flow reactor (2.5 cm i.d.) and a 2-m3 indoor Teflon film chamber under darkness. The acid catalytic effects on heterogeneous aerosol production were observed for diverse carbonyls in various ranges of humidities and compositions of seed inorganic aerosols. Particle population data measured by a scanning mobility particle sizer were used to calculate organic aerosol growth. To accountforthe aerosol growth contributed by heterogeneous reactions, the increase in organic aerosol mass was normalized bythe organic mass predicted by partitioning or the square of predicted organic mass. The carbonyl heterogeneous reactions were accelerated in the presence of acid catalysts (H2SO4), leading to higher aerosol yields than in their absence. The experimental data from aerosol yields in the flow reactorwere semiempirically fitted to the model parameters to predict the organic aerosol growth. The model parameters consist of environmental characteristics and molecular structure information of organic carbonyls. Basicity constants of carbonyls were used to describe the proton affinity of carbonyls for the acid catalysts. Particle environmental factors, such as humidity, temperature, and inorganic seed composition, were expressed by excess acidity and the parameters obtained from an inorganic thermodynamic model. A stepwise regression analysis of the aerosol growth model for the experimental data revealed that either the chemical structure information of carbonyls or characteristic environmental parameters are statistically significant in the prediction of organic aerosol growth. It was concluded thatthis model approach is applicable to predict secondary organic aerosol formation by heterogeneous reaction. PMID- 15667093 TI - Zinc adsorption effects on arsenite oxidation kinetics at the birnessite-water interface. AB - Arsenite is more toxic and mobile than As(V) in soil and sediment environments, and thus it is advantageous to explore factors that enhance oxidation of As(III) to As(V). Previous studies showed that manganese oxides, such as birnessite (delta-MnO2), directly oxidized As(III). However, these studies did not explore the role that cation adsorption has on As(III) oxidation. Accordingly, the effects of adsorbed and nonadsorbed Zn on arsenite (As(III)) oxidation kinetics at the birnessite-water interface were investigated using batch adsorption experiments (0.1 g L(-1); pH 4.5 and 6.0; I= 0.01 M NaCl). Divalent Zn adsorption on synthetic delta-MnO2 in the absence of As(II) increased with increasing pH and caused positive shifts in electrophoretic mobility values at pH 4-6, indirectly suggesting inner-sphere Zn adsorption mechanisms. Arsenite was readily oxidized on birnessite in the absence of Zn. The initial As(III) oxidation rate constant decreased with increasing pH from 4.5 to 6.0 and initial As(III) concentrations from 100 to 300 microM. Similar pH and initial As(III) concentration effects were observed in systems when Zn was present (i.e., presorbed Zn prior to As(III) addition and simultaneously added Zn-As(III) systems), but As(III) oxidation reactions were suppressed compared to the respective control systems. The suppression was more pronounced when Zn was presorbed on the delta-MnO2 surfaces as opposed to added simultaneously with As(III). This study provides further understanding of As(III) oxidation reactions on manganese oxide surfaces under environmentally applicable conditions where metals compete for reactive sites. PMID- 15667094 TI - Speciation of iron and sulfate in acid waters: aqueous clusters to mineral precipitates. AB - Acid mine drainage (AMD) contaminates surface water bodies, groundwater, soils, and sediments at innumerable locations around the world. AMD usually originates by weathering of pyrite (FeS2) and is rich in Fe and sulfate. In this study, we investigated speciation of FeII, FeIII, and SO4 in acid waters by Fourier transform infrared and X-ray absorption spectroscopy. The molalities of sulfate (15 mmol/ kg) and iron (10, 20, and 50 mmol/kg), and pH (1, 2, and 3) were chosen to mimic the concentration of ions in AMD waters. Sulfate and FeII either associate in outer-sphere complexes or do not associate at all. In contrast, sulfate interacts strongly with FeIII. The predominating species in FeIII-SO4 solutions are hydrogen-bonded complexes; inner-sphere complexes account only for 10+/-10% of the total sulfate. Our results show that the mode of interaction between FeIII and sulfate is similar in aqueous phase and in nanocrystalline precipitate schwertmannite (approximately FeO(OH)3/4(SO4)1/8). Because of this similarity, schwert-mannite should be the phase that controls solubility and availability of FeIII, SO4, and indirectly also other components in the AMD solutions. PMID- 15667095 TI - Kinetics and inhibition of reductive dechlorination of chlorinated ethylenes by two different mixed cultures. AB - Kinetic studies with two different anaerobic mixed cultures (the PM and the EV cultures) were conducted to evaluate inhibition between chlorinated ethylenes. The more chlorinated ethylenes inhibited the reductive dechlorination of the less chlorinated ethylenes, while the less chlorinated ethylenes weakly inhibited the dechlorination of the more chlorinated ethylenes. Tetrachloroethylene (PCE) inhibited reductive trichloroethylene (TCE) dechlorination but not cis dichloroethylene (c-DCE) dechlorination, while TCE strongly inhibited c-DCE and VC dechlorination. c-DCE also inhibited vinyl chloride (VC) transformation to ethylene (ETH). When a competitive inhibition model was applied, the inhibition constant (K(I)) for the more chlorinated ethylene was comparable to its respective Michaelis-Menten half-velocity coefficient, K(S). Model simulations using independently derived kinetic parameters matched the experimental results well. k(max) and K(S) values required for model simulations of anaerobic dechlorination reactions were obtained using a multiple equilibration method conducted in a single reactor. The method provided precise kinetic values for each step of the dechlorination process. The greatest difference in kinetic parameters was for the VC transformation step. VC was transformed more slowly by the PM culture (k(max) and K(S) values of 2.4+/-0.4 micromol/mg of protein/day and 602+/-7 microM, respectively) compared to the EV culture (8.1+/-0.9 micromol/mg of protein/day and 62.6+/-2.4 microM). Experimental results and model simulations both illustrate how low K(S) values corresponded to efficient reductive dechlorination for the more highly chlorinated ethylenes but caused strong inhibition of the transformation of the less chlorinated products. Thus, obtaining accurate K(S) values is important for modeling both transformation rates of parent compounds and their inhibition on daughter product transformation. PMID- 15667096 TI - Adsorption of ethylene glycol vapor on (alpha-AI2O3 (0001) and amorphous SiO2 surfaces: observation of molecular orientation and surface hydroxyl groups as sorption sites. AB - Vapor adsorption is an important process influencing the migration and the fates of many organic pollutants in the environment. In this study, adsorption of ethylene glycol (EG) vapor onto single crystal alpha-Al2O3 (0001) and fused SiO2 (amorphous) surfaces was studied with sum frequency generation spectroscopy, a well-suited surface specific technique for probing interfacial phenomena atthe molecular scale. Air-aqueous EG solutions were also investigated to compare to the adsorption at the air-solid interface in the presence of water vapor. The gauche conformer of EG molecules dominates the air-aqueous EG solution interface, and EG molecules act as hydrogen acceptors at the air-liquid interface. Water and surface hydrophilic/ hydrophobic properties play important roles for the adsorption of EG onto silica and alumina surfaces. The adsorbed EG molecules interact in different ways at the two different oxide surfaces. EG molecules weakly physisorb onto the alpha-Al2O3 (0001) surface by forming relatively weak hydrogen bonds with surface water molecules. On the silica surface, the suppression of the silanol OH stretching peak indicates that EG molecules form hydrogen bonds with silanol OH groups. PMID- 15667097 TI - Enrichment of stable carbon and hydrogen isotopes during anaerobic biodegradation of MTBE: microcosm and field evidence. AB - The conventional approach to evaluate biodegradation of organic contaminants in groundwater is to demonstrate an increase in the concentration of transformation products. This approach is problematic for MTBE from gasoline spills because the primary transformation product (TBA) can also be a component of gasoline. Compound-specific stable isotope analysis may provide a useful alternative to conventional practice. Changes in the delta13C and deltaD of MTBE during biodegradation of MTBE in an anaerobic enrichment culture were compared to the delta13C and deltaD of MTBE in groundwater at nine gasoline spill-sites. The stable isotopes of hydrogen and carbon were extensively fractionated during anaerobic biodegradation of MTBE. The stable isotope enrichment factor for carbon (epsilonC) in the enrichment cultures was -13 (-14.1 to -11.9 at 95% confidence level), and the hydrogen enrichment factor (epsilonH) was -16 (-21 to -11 at 95% confidence level). The isotope enrichment factors for carbon and hydrogen during anaerobic biodegradation indicate that the first reaction is enzymatic hydrolysis of the O-Cmethyl bond. The ratio of epsilonH to epsilonC was consistent between the enrichment culture and the field site that provided the inoculum, and with the other eight sites, suggesting a common degradation pathway. Compound-specific isotope evidence is discussed in terms of its utility for monitoring in situ biodegradation, in particular, for measuring how much MTBE was degraded. For the studied field sites, significant biodegradation of the original mass of MTBE is suggested, in some cases exceeding 90%. PMID- 15667098 TI - Effects of Fe(II) and hydrogen peroxide interaction upon dissolving UO2 under geologic repository conditions. AB - Iron redox cycling is supposed to be one of the major mechanisms that control the geochemical boundary conditions in the near field of a geologic repository for UO2 spent nuclear fuel. This work investigates the impact of reactions between hydrogen peroxide (H2O2) and iron (Fe2+/Fe3+) on UO2 dissolution. The reaction partners were contacted with UO2 in oxygen-free batch reactor tests. The interaction in absence of UO2 gives a stoichiometric redox reaction of Fe2+ and H2O2 when the reactants are present in equal concentration. Predomination of H202 results in its delayed catalytic decomposition. With UO2 present, its dissolution is controlled by either a slow mechanism (as typical for anoxic environments) or uranium peroxide precipitation, depending strongly on the reactant ratio. Uranium peroxide (UO4 x nH2O, m-studtite), detected on UO2 surfaces after exposure to H2O2, was not found on the surfaces exposed to solutions with stoichometric Fe(II)/ H2O2 ratios. This suggests that H2O2 was deactivated in redox reactions before a formation of UO4 took place. ESR measurements employing the spin trapping technique revealed only the DMPO-OH adduct within the first minutes after the reaction start (high initial concentrations of the OH radical); however, in the case of Fe(II) and H2O2 reacting at 10(-4) mol/L with UO2, dissolved oxygen and Fe2+ concentrations indicate the participation of further Fe intermediates and, therefore, Fenton redox activities. PMID- 15667099 TI - Reduction of 2,4,6-trinitrotoluene by iron metal: kinetic controls on product distributions in batch experiments. AB - The reaction kinetics and product distributions for the reduction of 2,4,6 trinitrotoluene (TNT) by granular iron metal (Fe0) were studied in batch experiments under a variety of initial concentrations of TNT and Fe0. Although the kinetics of TNT disappearance were found to behave in accord with the standard theory for surface-mediated reactions, a complex relationship was found between the initial concentrations of TNT and Fe0 and the appearance of the expected nitro reduction product, 2,4,6-triaminotoluene (TAT). TNT was completely converted to TAT only when the initial concentration of TNT was low and/or the initial concentration of Fe0 was high. Mathematical analysis of a range of generic reaction schemes that produce stable end products in addition to TAT showed that (i) surface complexation of TAT is insufficient to describe all of our data and (ii) polymerization reactions involving TAT and/or various reaction intermediates are the likely source of the incomplete conversion of TNT to TAT at high initial TNT concentration and low Fe0 concentration. The relationship between TAT production and reaction conditions is shown to imply that passivation due to reaction products is more likely when the ratio of initial TNT concentration to Fe0 concentration is high and, therefore, that passivation rates observed at the laboratory scale are likely to be faster than those which would be observed at the field scale. PMID- 15667100 TI - Modeling of porous filter penneability via image-based stochastic reconstruction of spatial porosity correlations. AB - A methodology for producing a pore-scale, 3D computational model of porous filter permeability is developed that is based on the analysis of 2D images of the filter matrix and first principles. The computationally reconstructed porous filter model retains statistical details of porosity and the spatial correlations of porosity within the filter and can be used to calculate permeability for either isotropic or 1D anisotropic porous filters. In the isotropic case, validation of the methodology was conducted using 0.2 and 0.8 microm ceramic membrane filters,forwhich it is shown that the image-based computational models provide a viable statistical reproduction of actual porosity characteristics. It is also shown that these models can predict water flux directly from first principles with deviations from experimental measurements in the range of experimental error. In the anisotropic case, validation of the methodology was conducted using a natural river sand filter. For this case, it is shown that the methodology yields predictions of filtration velocity that are similar or better than predictions offered by existing filtration models. It was found for the sand filter that the deviation between observation and prediction was mostly due to swelling during the preparation of the sand filter for imaging and can be reduced significantly using alternative methods reported in the literature. On the basis of these results, it is concluded that the computational reconstruction methodology is valid for porous filter modeling, and given that it captures pore scale details, it has potential application to the investigation of permeability decline underthe influence of pore-scale fouling mechanisms. PMID- 15667101 TI - X-ray absorption near-edge structure analysis of arsenic species for application to biological environmental samples. AB - Arsenic is an element that is ubiquitous in the environment and is known to form compounds with toxic, even carcinogenic properties. Arsenic toxicity is a function of its chemical form (species). Identification of arsenic species is necessary to accurately determine the transformation and fate of arsenicals as well as the actual risk posed by arsenic contamination. We report X-ray absorption near-edge structure (XANES) measurements of 16 biologically important arsenic compounds. Solid and aqueous standards were studied for differences in XANES spectral features, white line positions, stability during exposure to the beam, and stability between two beam exposures separated by 48 h. Samples containing As(III) (11870.0-11871.7+/-0.5 eV) and As(V) (11872.6-11875.3+/-0.5 eV) were easily distinguished by white line energies and could be further subdivided into a total of seven groups. Valuable examples include As(III)-sulfur compounds (11870.0+/-0.5 eV), arsenobetaine and arsenocholine (11872.6+/-0.5 eV), and a dimethyl arsinyl riboside (11873.3+/-0.5 eV). A growing number of environmental and biological studies use X-ray absorption spectroscopy (XAS) results to complement their more traditional analyses. Results provided here are intended to help make XAS more accessible to new users interested in the study of arsenic in the environment. PMID- 15667102 TI - Chemical fingerprinting of petroleum biomarkers using time warping and PCA. AB - A new method for chemical fingerprinting of petroleum biomakers is described. The method consists of GC-MS analysis, preprocessing of GC-MS chromatograms, and principal component analysis (PCA) of selected regions. The preprocessing consists of baseline removal by derivatization, normalization, and alignment using correlation optimized warping. The method was applied to chromatograms of m/z 217 (tricyclic and tetracyclic steranes) of oil spill samples and source oils. Oil spill samples collected from the coastal environment in the weeks after the Baltic Carrier oil spill were clustered in principal components 1 to 4 with oil samples from the tank of the Baltic Carrier (source oil). The discriminative power of PCA was enhanced by deselecting the most uncertain variables or scaling them according to their uncertainty, using a weighted least squares criterion. The four principal components were interpreted as follows: boiling point range (PC1), clay content (PC2), carbon number distribution of sterols in the source rock (PC3), and thermal maturity of the oil (PC4). In summary, the method allows for analyses of chromatograms using a fast and objective procedure and with more comprehensive data usage compared to other fingerprinting methods. PMID- 15667103 TI - Field calibration of rapidly equilibrating thin-film passive air samplers and their potential application for low-volume air sampling studies. AB - This paper reports on a field calibration and ambient deployment study with rapidly equilibrating thin-film passive air samplers. POlymer-coated Glass (POG) samplers have a coating of ethylene vinyl acetate (EVA) less than 1 microm thick coated on to glass, which can be dissolved off after exposure and prepared for quantification of persistent organic pollutants (POPs) that have partitioned into the film during field exposure. In this study, POGs were exposed for up to 18 d, in a study to assess compound uptake rates and their time to approach equilibrium. Results confirmed theoretical predictions, with time to equilibrium varying between a few hours to ca. 20 d for PCB-18 and PCB-138, respectively. Performance reference compounds and contaminated POGs were used to investigate depuration kinetics, confirming that lighter congeners behave extremely dynamically with substantial losses from the films over periods of a few hours. Repeated deployments of the samplers for different3-d periods yielded detectable levels of a range of PCB congeners, which had partitioned from as little as approximately 2 to 10 m3 air. This highlights the potential utility of POGs for extremely sensitive and dynamic passive air sampling in the future to help improve understanding of sources, environmental fate, and behavior of POPs. Recommendations are made for future improvements/refinements in POG sampling and handling procedures. PMID- 15667104 TI - Real-time visualization and quantification of PAH photodegradation on and within plant leaves. AB - Vegetation plays a key role in the environmental cycling and fate of many organic chemicals. A compound's location on or within leaves will affect its persistence and significance; retention in surface compartments (i.e., the epicuticular wax and cuticle) renders the compound more susceptible to photodegradation and volatilization, while penetration into the epidermal cell walls or cytoplasm will enhance susceptibility to metabolism. Here, for the first time, methodologies which combine plant and PAH autofluorescence with two-photon excitation microscopy (TPEM) are used to visualize and quantify compound photodegradation on and within living plant leaves. Anthracene,fluoranthene, and phenanthrene were introduced to living leaves of Zea mays and monitored in real time, in control treatments, and when subject to UV-A radiation. Compound photodegradation was observed directly; different degradation rates occurred for different compounds (anthracene > fluoranthene > phenanthrene) and in different locations (at the leaf surface > within the epidermal cells). Results suggest that photodegradation on vegetation may be a more important loss mechanism for PAHs than previously thought. Compound fate in vegetation is potentially highly complex, influenced by diffusion into and location within leaf structures, the rates of supply/loss with the atmosphere, exposure to sunlight, and other environmental conditions. The techniques described here provide a real-time tool to advance insight into these issues. PMID- 15667105 TI - New device and method for flux-proportional sampling of mobile solutes in soil and groundwater. AB - The importance of monitoring the transport of organic contaminants in soil and groundwater, and the pros and cons of existing sampling methods, are outlined. A new, alternative sampling method is proposed, using a passive sampler that functions as a water-permeable, semi-infinite sink for passing solutes of interest. Tracers integrated in the device store information on the volume of water passing through the sampler during the installation period. The conceptual basis of the sampling method is described. This device enables flux-proportional monitoring of the concentrations of mobile contaminants in the soil and groundwater. 14C-labeled phenanthrene (PHEN) and glyphosate (GLY) are used as case study compounds in laboratory experiments. The sorption capacities and uptake kinetics of 13 adsorbents are screened and compared, as well as the dissolution kinetics of three tracer salts: calcium citrate, calcium fluoride (CaF2), and calcium hydrogen phosphate (CaHPO4). The application of the passive sampler is then demonstrated in long-term laboratory experiments, using large soil columns under steady-state hydraulic conditions. The accumulated flux of PHEN was sampled with an accuracy of 3.6%-17.8%, using graphitized carbon, hexagonal mesoporous silica, and cross-linked polymers as adsorbents. The accumulated flux of GLY was sampled with an accuracy of 12.4%, using gamma alumina as an adsorbent. The advantages and limitations of this new environmental monitoring method are discussed. PMID- 15667106 TI - Potential use of a host associated molecular marker in Enterococcus faecium as an index of human fecal pollution. AB - Several genotypic and phenotypic microbial source tracking (MST) methods have been proposed and utilized to differentiate groups of microorganisms, usually indicator organisms, for the purpose of tracking sources of fecal pollution. Targeting of host-specific microorganisms is one of the approaches currently being tested. These methods are useful as they circumvent the need to isolate individual microorganisms and do not require the establishment of reference databases. Several studies have demonstrated that the presence and distribution of Enterococcus spp. in feces seems to be influenced by the host species. Here, we present a method for detection of genetic sequences in culturable enterococci capable of identifying human sources of fecal pollution in the environment. The human fecal pollution marker designed in this study targets a putative virulence factor, the enterococcal surface protein (esp), in Enterococcus faecium. This gene was detected in 97% of sewage and septic samples but was not detected in any livestock waste lagoons or in bird or animal fecal samples. Epidemiological studies in recreational and groundwaters have shown enterococci to be useful indicators of public health risk for gastroenteritis. By identifying the presence of human fecal pollution, and therefore the possible presence of human enteric pathogens, this marker allows for further resolution of the source of this risk. PMID- 15667107 TI - Selective quantification of trace palladium in road dusts and roadside soils by displacement solid-phase extraction online coupled with electrothermal atomic absorption spectrometry. AB - There is a growing concern about the effect of palladium on human health because of the toxicity and increasing occurrence of palladium as a result of its extensive use in automotive catalytic converters. Development of reliable analytical methodologies for the determination of palladium in environmental materials is of great importance for critical evaluation of the possible risks for human health. In this work, a displacement solid-phase extraction technique was developed and online coupled to electrothermal atomic absorption spectrometry (ETAAS) for selective and sensitive determination of trace palladium in environmental samples without need of any special selective complexing agents, selective sorbents, and masking agents. The developed methodology involved the online formation of copper pyrrolidine dithiocarbamate (Cu-PDC), and the resultant Cu-PDC was extracted onto a microcolumn packed with the sorbent from a cigarette filter. Trace Pd(II) was selectively preconcentrated through loading the sample solution onto the microcolumn by online displacement reaction between Pd(II) and the extracted Cu-PDC on the microcolumn. The retained analyte was subsequently eluted with 40 microL of ethanol for online ETAAS determination. Interferences from coexisting heavy metal ions with lower stability of their PDC complexes relative to Cu-PDC were minimized. The tolerable concentrations of Cd (II), Fe(III), Co(II), Mn(II), Cr(III), and Zn(II) were up to 2, 6, 40, 2, 1.5, and at least 100 mg L(-1), respectively. Compared with conventional solid-phase extraction methodology, the developed displacement solid-phase extraction protocol gave 2-4 orders of magnitude improvement in the maximum tolerable concentrations of coexisting heavy metal ions. With the consumption of only 2.8 mL of sample solution, an enhancement factor of 52 and a detection limit (3sigma) of 18 ng L(-1) were achieved at a sample throughput of 30 samples h(-1). The precision (RSD, n = 13) was 2.5% at the 1 microg L(-1) level. The present methodology was successfully applied to selective determination of trace palladium in local road dusts and roadside soils. PMID- 15667108 TI - Bioaccumulation of organic chemicals in contaminated soils: evaluation of bioassays with earthworms. AB - Earthworms live in close contact with the soil and can thus be considered representative for the bioavailability of chemicals at contaminated sites. Bioavailability can either be assessed by analyzing earthworms from contaminated locations or by exposing laboratory-reared specimens to soil samples from the field (bioassays). In this study, we investigate the relevance of bioassays by using an extended experimental design (to identify signs of depletion of the bioavailable phase by the earthworms) and by using two species of earthworm (the standard test species Eisenia andrei and the field-relevant Aporrectodea caliginosa). Furthermore, bioassay results are compared to body residues of worms collected from the field site: a heavily polluted polder, amended with dredge spoil. We focused on telodrin, dieldrin, hexachlorobenzene, and eight PCBs. With our bioassay design, it was shown that depletion was unlikely, although more subtle effects could have occurred (e.g., changes in sorption during the experiments). E. andrei is a good choice for bioassays because its body residues correlate well to those in A. caliginosa, as well as to those in the field collected worms. Nevertheless, E. andrei accumulated slightly more than the other species and appeared to be more sensitive to the conditions in soil from one of our sites. PMID- 15667109 TI - Reliability of a commercial kit to test groundwater for arsenic in Bangladesh. AB - A comparison of field and laboratory measurements of arsenic in groundwater of Araihazar, Bangladesh, indicates that the most widely used field kit correctly determined the status of 88% of 799 wells relative to the local standard of 50 microg/L As. Additional tests showthatthe inconsistencies, mainly underestimates in the 50-100 microg/L As range, can be avoided by increasing the reaction time from 20 to 40 min. Despite this limitation, the field data already compiled for millions of wells by the Bangladesh Arsenic Mitigation and Water Supply Project, in combination with information on well location and depth, should prove to be extremely useful to prioritize interventions in thousands of affected villages. PMID- 15667110 TI - New field method: gas push-pull test for the in-situ quantification of microbial activities in the vadose zone. AB - Quantitative information on microbial processes in the field is important. Here we propose a new field method, the "gas push-pull test" (GPPT) for the in-situ quantification of microbial activities in the vadose zone. To evaluate the new method, we studied microbial methane oxidation above an anaerobic, petroleum contaminated aquifer. A GPPT consists of the injection of a gas mixture of reactants (e.g., methane, oxygen) and nonreactive tracer gases (e.g., neon, argon) into the vadose zone and the subsequent extraction of the injection gas mixture together with soil air from the same location. Rate constants of gas conversion are calculated from breakthrough curves of extracted reactants and tracers. In agreement with expectations from previously measured gas profiles, we determined first-order rate constants of 0.68 h(-1) at 1.1 m below soil surface and 2.19 h(-1) at 2.7 m, close to the groundwater table. Co-injection of a specific inhibitor (acetylene) for methanotrophs showed that the observed methane consumption was microbially mediated. This was confirmed by increases of stable carbon isotope ratios in methane by up to 42.6 %. In the future, GPPTs should provide useful quantitative information on a range of microbial processes in the vadose zone. PMID- 15667111 TI - Metal speciation in anoxic sediments: when sulfides can be construed as oxides. AB - Metal speciation in aquatic sediments is often characterized using wet chemical sequential extraction techniques. However, these methods are operationally defined and subject to artifacts, particularly when dealing with anoxic sediments, in which metal sulfide precipitates are likely to occur. Using X-ray absorption spectroscopy (XAS) and acid-volatile sulfide (AVS) analysis, we evaluated the effectiveness of one of the most widely used sequential extraction protocols, the Tessier method, at determining Zn and Pb speciation in anoxic wetland sediments. Sequential extraction results significantly underestimated the amount of Zn associated with sulfide phases as compared to the other two approaches. XAS analysis of ZnS amended sediments indicates that the most likely source of this conflict is an early dissolution of amorphous metal sulfide phases during the sequential extraction step corresponding to the extraction of iron and manganese oxides. The reagent mixture used in this step, hydroxylamine hydrochloride-HCl, is widely used in other sequential extraction protocols, including the BCR method, limiting their application to anoxic sediments. For this reason, current sequential extraction techniques should only be used on anoxic sediments with caution, and/or in conjunction with complementary approaches to assess metal speciation. PMID- 15667112 TI - Solvent release into a sandy aquifer. 2. Estimation of DNAPL mass based on a multiple-component dissolution model. AB - A chlorinated solvent mixture (2.0 L of trichloroethylene, 0.5 L of chloroform, and 2.5 L of tetrachloroethylene) was released into a sandy aquifer to create a heterogeneously distributed DNAPL (dense nonaqueous-phase liquid) source. The dissolution and dissolved-phase plume development from this source were studied in detail along a cross-section downgradient of the source for a period of approximately 1 year. At the conclusion of the experiment, the site was excavated to map the actual distribution of solvent residuals in the subsurface. Multiple component dissolution theory provides a tool for the estimation of the mass of a multiple-component DNAPL source present in the groundwater. Concentration ratios between the compounds change with time, and those changes can be used to estimate the mass of DNAPL upgradient of the monitoring point(s) or well(s). The method is independent of the dilution occurring in the groundwater and only requires observations of time series of the contaminants in one or more monitoring points. For the field experiment, the method was applied using the measured concentrations of individual sampling points, the depth-integrated concentrations, the area-integrated concentrations, and the effluent concentrations of the cell. The experiment showed that multiple-component dissolution theory may be a valuable tool for the estimation of the mass of multiple-component DNAPL residuals in the saturated zone. PMID- 15667113 TI - Bioremediation of diethylhexyl phthalate contaminated soil: a feasibility study in slurry- and solid-phase reactors. AB - The aim of the research was to verify the possibility of applying bioremediation as a treatment strategy on a poly(vinyl chloride) (PVC) manufacturing site in the north of Italy contaminated by diethylhexyl phthalate (DEHP) at a concentration of 5.51 mg/g of dry soil. Biodegradation kinetic experiments with DEHP contaminated soil samples were performed in both slurry- and solid-phase systems. The slurry-phase results showed that the cultural conditions, such as N and P concentrations and the addition of a selected DEHP degrading strain, increased the natural DEHP degradation rate. On the basis of these data, experiments to simulate bioventing on contaminated soil columns were performed. The DEHP concentration reached 0.63 mg/g of dry soil in 76 days (89% of degradation). A kinetic equation was developed to fit the experimental data and to predict the concentration of contaminant after treatment. The data obtained are encouraging for a future in situ application of the bioventing technology. PMID- 15667114 TI - Benzene: a secondary pollutant formed in the three-way catalyst. AB - Benzene emissions from a relevant proportion of today's gasoline-driven passenger cars and light-duty vehicles can increase by up to 2 orders of magnitude when driving at high engine load (e.g., on highways). Under such conditions, post catalyst benzene levels exceeded those found pre-catalyst. As a consequence, formation of benzene in the catalyst was postulated. To further reduce ambient air concentrations of benzene,these critical operating conditions must be carefully avoided. Here, we report in detail to what extent and at what operating conditions catalyst-induced benzene and toluene formation can occur. For that purpose, a EURO-1 passenger car (1.8 L, model year 1995)fulfilling the valid regulations, equipped with a new, two-layered, Pd-CeO2-Al2O3/Rh-ZrO2-Al2O3 three way catalyst was operated at steady state on a chassis dynamometer at 100, 125, and 150 km/h at variable air to fuel ratios. Pre- and post-catalyst exhaust gas concentrations of benzene, toluene, C2-, and C3-benzenes were monitored at a time resolution of 0.5 Hz by means of chemical ionization mass spectrometry. A net benzene formation window, ranging from pre-catalyst exhaust gas temperatures of 600-730 degrees C and lambda-values of 0.83-0.95, with a pronounced minimum at 0.87, was observed. Dealkylation reactions of aromatic hydrocarbons are assumed to be the major pathway leading to benzene. PMID- 15667115 TI - Bacterial inactivation in open air by the afterglow plume emitted from a grounded hollow slot electrode. AB - Escherichia coli, Bacillus atrophaeus, and Bacillus atrophaeus spores were exposed to a downstream plasma afterglow plume emitted from a slotted plasma device operating in open air at atmospheric pressure. The reactor electrodes were RF powered at 13.56 MHz to excite a mixture of argon and oxygen gases by a capacitive discharge as it flowed past the electrodes into open air. Bacterial inactivation experiments on surfaces exposed to the plasma afterglow were conducted with varying plasma exposure times. Experimental results demonstrated a colony forming unit (CFU) reduction of almost 5 log10 of E. coli with only 1 s of exposure per unit area. One log CFU reduction was observed in B. atrophaeus with the same treatment time of 1 s per unit area. B. atrophaeus spores showed a reduction of 3 log10 with exposure time of 10 min. Comparison on various growth media suggests that cells are killed rather than sublethally injured, while the mechanistic action of the plasma appears to affect both nucleic acids as well as the cell wall structure. These results present a promising means of inactivation of harmful microbes in a practical environment with an electrically grounded device that is handheld, much like a wand applicator. Results are applicable to the development of plasma sterilization tools for various environmental purposes. PMID- 15667116 TI - Characterization and preliminary assessment of a sorbent produced by accelerated mineral carbonation. AB - This study shows that calcium silicate/aluminate-based materials can be carbonated to produce sorbents for metal removal. The material chosen for investigation, cement clinker, was accelerated carbonated, and its structural properties were investigated using X-ray diffraction (XRD), scanning electron microscopy, thermal gravimetric and differential thermal analysis, nuclear magnetic resonance spectroscopy, and nitrogen gas adsorption techniques. The principal carbonation reactions involved the transformation of dicalcium silicate, tricalcium silicate, and tricalcium aluminate into a Ca/Al-modified amorphous silica and calcium carbonate. It was found that carbonated cement had high acid buffering capacity, and maintained its structural integrity within a wide pH range. The uptake of Pb(II), Cd(II), Zn(II), Ni(II), Cr(II), Sr(II), Mo(VI), Cs(II), Co(II), and Cu(II) from concentrated (1000 mg L(-1)) single-metal solutions varied from 35 to 170 mg g(-1) of the carbonate cement. The removal of metals was hardly effected by the initial solution pH due to the buffering capability of the carbonated material. The kinetics of Pb, Cd, Cr, Sr, Cs, and Co removal followed a pseudo-second-order kinetic model, whereas the equilibrium batch data for Cu fitted the pseudo-first-order rate equation. PHREEQC simulation supported by XRD analysis suggested the formation of metal carbonates and silicates, calcium molybdate, and chromium (hydro)oxide. Cesium was likely to be adsorbed by Ca/Al-modified amorphous silica. PMID- 15667117 TI - Generalized first-order kinetic model for biosolids decomposition and oxidation during hydrothermal treatment. AB - The main objective of this study was to develop generalized first-order kinetic models to represent hydrothermal decomposition and oxidation of biosolids within a wide range of temperatures (200-450 degrees C). A lumping approach was used in which oxidation of the various organic ingredients was characterized by the chemical oxygen demand (COD), and decomposition was characterized by the particulate (i.e., nonfilterable) chemical oxygen demand (PCOD). Using the Arrhenius equation (k = k(o)e(-Ea/RT)), activation energy (Ea) levels were derived from 42 continuous-flow hydrothermal treatment experiments conducted at temperatures in the range of 200-450 degrees C. Using predetermined values for k(o) in the Arrhenius equation, the activation energies of the various organic ingredients were separated into 42 values for oxidation and a similar number for decomposition. The activation energy values were then classified into levels representing the relative ease at which the organic ingredients of the biosolids were oxidized or decomposed. The resulting simple first-order kinetic models adequately represented, within the experimental data range, hydrothermal decomposition of the organic particles as measured by PCOD and oxidation of the organic content as measured by COD. The modeling approach presented in the paper provide a simple and general framework suitable for assessing the relative reaction rates of the various organic ingredients of biosolids. PMID- 15667118 TI - Long life modified lead dioxide anode for organic wastewater treatment: electrochemical characteristics and degradation mechanism. AB - Recent studies have shown that the lack of ideal anodes with both good activity and stability is still one of the critical problems in electrochemical oxidation for organic wastewater treatment. The electrochemical properties, the activity and stability for anodic oxidation of various phenolic compounds, and the degradation mechanism on a novel beta-PbO2 electrode modified with fluorine resin were investigated. The anode life after modification was greatly improved to be more than 10 yr in common electrochemical current conditions. Such an anode was effective for partial degradation of phenolic compounds, but selective because reactive activities were varied with different substituents. Characterized by SEM and XRD, the crystal form of the anode was verified to be mainly beta-PbO2, and it hardly changed when used for p-nitrophenol degradation for around 320 h although there existed slow electrode corrosion. The active species generated during anodic oxidation were determined to be mainly hydroxyl radical and little ozone. The reactions between hydroxyl radical and phenolic compounds were proved to be electrophilic reactions, based on which a general electrochemical degradation mechanism for aromatic compounds was proposed. In general, such a novel anode has a good performance for organics degradation with perfect electrode life, showing potential for environmental application. PMID- 15667119 TI - Methodology for predicting OEL from rodent LD50 values for metals and metallic compounds. AB - The relationship between the occupational exposure limits (OEL) and the lethal dose 50 (LD50) values of rats or mice for metals and metallic compounds was statistically analyzed by a stepwise multivariate regression method. The OEL values were predicted from LD50 values and metallic compensation coefficients (MCC), which were developed as the regression coefficients of dummy variables that represented the metallic element contained in the substance of interest. The value of the MCC indicated the extent of the adverse health effects of the metal in the substance. Smaller values of the MCC were assigned to metals that would have the more severe adverse health effects, such as carcinogenesis, while larger values were given to the less toxic metals. The Health Index (HI) based on the OEL values was proposed as a convenient measure of the toxicity of industrial products. The prediction method could be applied to toxicity risk assessments by using the HI when a designer of consumer products wants to use substances for which OEL values have not been determined. Two case studies were conducted to estimate the potential toxicity of materials used in solders and in rechargeable batteries. PMID- 15667120 TI - Comment on "Global assessment of polybrominated diphenyl ethers in farmed and wild salmon". PMID- 15667121 TI - Comment on "Modeling maximum adsorption capacities of soot and soot-like materials for PAHs and PCBs". PMID- 15667123 TI - ArQiologist: an integrated decision support tool for lead optimization. AB - This paper describes ArQiologist, a Web-based tool that integrates chemical, analytical, biological, and computational data to facilitate decision support for lead optimization at ArQule. It features an easy-to-use graphical query builder that allows queries to be saved, reused, and shared by researchers. Query results can be viewed with built-in data browsers or exported with structures to external applications such as Microsoft Excel or Spotfire for further analysis. PMID- 15667124 TI - Virtual screening of novel noncovalent inhibitors for SARS-CoV 3C-like proteinase. AB - The SARS coronavirus 3C-like proteinase is considered as a potential drug design target for the treatment of severe acute respiratory syndrome (SARS). Owing to the lack of available drugs for the treatment of SARS, the discovery of inhibitors for SARS coronavirus 3C-like proteinase that can potentially be optimized as drugs appears to be highly desirable. We have built a "flexible" three-dimensional model for SARS 3C-like proteinase by homology modeling and multicanonical molecular dynamics method and used the model for virtual screening of chemical databases. After Dock procedures, strategies including pharmocophore model, consensus scoring, and "drug-like" filters were applied in order to accelerate the process and improve the success rate of virtual docking screening hit lists. Forty compounds were purchased and tested by HPLC and colorimetric assay against SARS 3C-like proteinase. Three of them including calmidazolium, a well-known antagonist of calmodulin, were found to inhibit the enzyme with an apparent K(i) from 61 to 178 microM. These active compounds and their binding modes provide useful information for understanding the binding sites and for further selective drug design against SARS and other coronavirus. PMID- 15667126 TI - Generalized fragment-substructure based property prediction method. AB - The need for fast and accurate predictors of pharmaceutically important properties has been increasing due to pressure from high-throughput screening, in silico screening, and the need to more rapidly identify potential pharmacokinetic issues before drugs advance to the more expensive clinical development stages. A novel method for making predictive models based on decomposing 2D structure into component structural fragments is used to model logP, water solubility, and melting point. The fragment orientation of the method facilitates understanding of how molecules might be altered to improve the desired properties. The 2D structure-based descriptor is computed by analysis of the target molecules with a substructure searching algorithm and a set of fragments selected for chemical and pharmaceutical relevance. These are combined with partial least squares to create predictive models. The correlation coefficients achieved are 0.86 for logP (SE = 0.68), 0.73 for logS (SE = 0.89), and 0.64 (SE = 48.9 degrees) for melting point over diverse data sets of 11,447, 2427, and 5598 molecules, respectively. The models were verified via test sets of compounds not included in the training set. PMID- 15667125 TI - Application of the genetic algorithm joint with the Powell method to nonlinear least-squares fitting of powder EPR spectra. AB - The application of the stochastic genetic algorithm (GA) in conjunction with the deterministic Powell search to analysis of the multicomponent powder EPR spectra based on computer simulation is described. This approach allows for automated extraction of the magnetic parameters and relative abundances of the component signals, from the nonlinear least-squares fitting of experimental spectra, with minimum outside intervention. The efficiency and robustness of GA alone and its hybrid variant with the Powell method was demonstrated using complex simulated and real EPR data sets. The unique capacity of the genetic algorithm for locating global minima, subsequently refined by the Powell method, allowed for successful fitting of the spectra. The influence of the population size, mutation, and crossover rates on the performance of GA was also investigated. PMID- 15667127 TI - Molecular modeling study of leflunomide and its active metabolite analogues. AB - Leflunomide is known as a compound with various sorts of biological activity, which found a practical application in medicine. Search of current literature revealed an active metabolite of Leflunomide together with its eight analogues synthesized as protein tyrosine kinase inhibitors with potential anticancer activity. Accurate description of the molecular structure of these compounds is valuable. The detailed geometrical parameters description was performed using DFT theory. The conformational analysis and intramolecular proton transfer were considered. Using the most stable conformation the detailed electronic structure description was obtained by analysis of electron density and electrostatic potential distribution in the first step. Next, the topological analysis of the electron density by AIM method and electron localization function (ELF) theories supplemented this study. The AIM and ELF theories were applied to study the topology of the molecules, atomic charges distribution, and details of bonding. The theoretical investigations were performed in the gas phase and by using SCRF/PCM solvent reaction field. In this study the molecular modeling results for Leflunomide and the analogues of its active metabolite are presented. PMID- 15667128 TI - Enrichment of ligands for the serotonin receptor using the Shape Signatures approach. AB - Shape Signatures, a new 3-dimensional molecular comparison method, has been adapted to rank ligands of the serotonin receptors. A set of 825 agonists and 400 antagonists together with approximately 10,000 randomly chosen compounds from the NCI database were used in this study. Both 1D and 2D Shape Signature databases were created, and enrichment studies were carried out. Results from these studies reveal that the 1D Shape Signature approach is highly efficient in separating agonists from a mixture of molecules which includes compounds randomly selected from the NCI database taken as inactives. It is also equally effective at separating agonists and antagonists from a pool of active ligands for the serotonin receptor. Parallel enrichment studies using 2D shape signatures showed high selectivity with more restricted coverage due to the high specificity of 2D signatures. The influence of conformational variation of the shape signature on enrichment was explored by docking a subset of ligands into the crystal structure of serotonin N-acetyltransferase. Enrichment studies on the resulting "docked" conformations produced only slightly improved results compared with the CORINA generated conformations. PMID- 15667129 TI - Quantum mechanical study of the potential energy surface of the ClO + NO2 reaction. AB - In the study of the reaction pathways of the ClO + NO2 reaction including reliable structures of the reactants, products, intermediates, and transition states as well as energies the MP2/6-311G(d), B3LYP/6-311G(d), and G2(MP2) methods have been employed. Chlorine nitrate, ClONO2, is formed by N-O association without an entrance barrier and is stabilized by 29.8 kcal mol(-1). It can undergo either a direct 1,3 migration of Cl or OCl rotation to yield an indistinguishable isomer. The corresponding barriers are 45.8 and 7.1 kcal mol( 1), respectively. ClONO2 can further decompose into NO3 + Cl with an endothermicity of 46.4 kcal mol(-1). The overall endothermicity of the NO2 + ClO -> NO3 + Cl reaction is calculated to be 16.6 kcal mol(-1). The formation of cis perp and trans-perp conformer of chlorine preoxynitrite, ClOONO(cp) and ClOONO(tp), are exothermic by 5.4 and 3.8 kcal mol(-1), respectively. Calculations on the possible reaction pathways for the isomerization of ClOONO to ClONO2 showed that the activation barriers are too high to account for appreciable nitrate formation from peroxynitrite isomerization. All quoted relative energies are related to G2(MP2) calculations. PMID- 15667130 TI - Determining the validity of a QSAR model--a classification approach. AB - The determination of the validity of a QSAR model when applied to new compounds is an important concern in the field of QSAR and QSPR modeling. Various scoring techniques can be applied to specific types of models. We present a technique with which we can state whether a new compound will be well predicted by a previously built QSAR model. In this study we focus on linear regression models only, though the technique is general and could also be applied to other types of quantitative models. Our technique is based on a classification method that divides regression residuals from a previously generated model into a good class and bad class and then builds a classifier based on this division. The trained classifier is then used to determine the class of the residual for a new compound. We investigated the performance of a variety of classifiers, both linear and nonlinear. The technique was tested on two data sets from the literature and a hand built data set. The data sets selected covered both physical and biological properties and also presented the methodology with quantitative regression models of varying quality. The results indicate that this technique can determine whether a new compound will be well or poorly predicted with weighted success rates ranging from 73% to 94% for the best classifier. PMID- 15667131 TI - QSPR using MOLGEN-QSPR: the challenge of fluoroalkane boiling points. AB - By means of the new software MOLGEN-QSPR, a multilinear regression model for the boiling points of lower fluoroalkanes is established. The model is based exclusively on simple descriptors derived directly from molecular structure and nevertheless describes a broader set of data more precisely than previous attempts that used either more demanding (quantum chemical) descriptors or more demanding (nonlinear) statistical methods such as neural networks. The model's internal consistency was confirmed by leave-one-out cross-validation. The model was used to predict all unknown boiling points of fluorobutanes, and the quality of predictions was estimated by means of comparison with boiling point predictions for fluoropentanes. PMID- 15667132 TI - What is the smallest saturated acyclic alkane that cannot be made? AB - Saturated acyclic alkanes may show a high degree of strain if they have many branches close to each other. We report calculations which indicate how strained a molecule may become before it falls apart rapidly at room temperature and so allow us to identify the simplest alkane which cannot be made. PMID- 15667133 TI - New Born radii deriving method for Generalized Born model. AB - Here we report a method to calculate Born radii, an important parameter used in a Generalized Born model. Traditional methods to derive Born radii are mostly based on a complicated formula, while our method is easier and more direct. Atoms are classified according to their atom type, and the Born radii of each type are obtained by fitting to experimental solvation free energy. The SMARTS language is used for the exact definition of atoms types, and Ullmann's subgraph isomorphism algorithm is used to deduce the environment. A generic algorithm is used for the parameter fitting because of its efficiency in searching a huge phase space, and its results are then optimized by using the conjugate gradient method. The final parameter set is fitting from a training set containing 357 molecules and is tested using a test set of 44 small organic molecules, and the average error is 0.58 kcal/mol for 36 neutral molecules and is 1.67 kcal/mol for 8 ions. The model is further tested under organic molecules, biopolymers, and a protein-inhibitor complex and yields reliable results in all these cases. This method can be used to accelerate molecular docking calculations. PMID- 15667134 TI - QSPR treatment of the soil sorption coefficients of organic pollutants. AB - In this study, general and class-specific QSPR models for soil sorption, logK(OC), of 344 organic pollutants (0 < logK(OC) < 4.94) were developed using a large variety of theoretical molecular descriptors based only on molecular structure. Two general models were obtained. The first model was derived for a structurally representative set of 68 chemicals (R2=0.76, s=0.44), whereas the second involved a total of 344 compounds (R2=0.76, s=0.41). The first was validated using the data for the remaining 276 pollutants (R2=0.70, s=0.45). An additional validation of both models was performed using an independent set of 48 pollutants. Both models predict the logK(OC) at the level of experimental precision, while the theoretical molecular descriptors appearing in the QSPR models give further insight into the mechanisms of soil sorption. The analysis of the distribution of the residuals of the logK(OC) values calculated by both general models indicated the need and possible advantages of modeling soil sorption for smaller data sets related to individual classes of chemicals. Accordingly, QSPR models were also developed for 14 chemical classes. The descriptors appearing in these models were discussed as related to the possible interaction mechanisms in soil sorption. PMID- 15667135 TI - Description of the electronic structure of organic chemicals using semiempirical and ab initio methods for development of toxicological QSARs. AB - The quality of quantitative structure-activity relationship (QSAR) models depends on the quality of their constitutive elements including the biological activity, statistical procedure applied, and the physicochemical and structural descriptors. The aim of this study was to assess the comparative use of ab initio and semiempirical quantum chemical calculations for the development of toxicological QSARs applied to a large and chemically diverse data set. A heterogeneous collection of 568 organic compounds with 96 h acute toxicity measured to the fish fathead minnow (Pimephales promelas) was utilized. A total of 162 descriptors were calculated using the semiempirical AM1 Hamiltonian, and 121 descriptors were compiled using an ab initio (B3LYP/6-31G**) method. The QSARs were derived using multiple linear regression (MLR) and partial least squares (PLS) analyses. Statistically similar models were obtained using AM1 and B3LYP calculated descriptors supported by the use of the logarithm of the octanol water partition coefficient (log K(ow)). The main difference between the models derived by both MLR and PLS with the two sets of quantum chemical descriptors was concentrated on the type of descriptors selected. It was concluded that for large scale predictions, irrespective of the mechanism of toxic action, the use of precise but time-consuming ab initio methods does not offer considerable advantage compared to the semiempirical calculations and could be avoided. PMID- 15667136 TI - New invariant of DNA sequences. AB - For a DNA sequence with n bases, one can always associate it with an n x n nonnegative real symmetric matrix whose diagonal entries are zero. Once the matrix is given, its leading eigenvalue is usually calculated and used as an invariant to characterize the DNA sequence. Let M be such a matrix, and lambda1 its leading eigenvalue. Then (1/n)//M//m1 and sqrt [(n-1)/n]//M//F are the lower and upper bounds of lambda1, respectively. Since their arithmetic average is an approximate value of lambda1 and simpler for calculation, we can use it as an alternative invariant to characterize the DNA sequence. The utility of the new parameter is illustrated on the DNA sequences of five species: human, chimpanzee, mouse, rat, and gallus. PMID- 15667138 TI - Homology modeling of a human glycine alpha 1 receptor reveals a plausible anesthetic binding site. AB - The superfamily of ligand-gated ion channels (LGICs) has been implicated in anesthetic and alcohol responses. Mutations within glycine and GABA receptors have demonstrated that possible sites of anesthetic action exist within the transmembrane subunits of these receptors. The exact molecular arrangement of this transmembrane region remains at intermediate resolution with current experimental techniques. Homology modeling methods were therefore combined with experimental data to produce a more exact model of this region. A consensus from multiple bioinformatics techniques predicted the topology within the transmembrane domain of a glycine alpha one receptor (GlyRa1) to be alpha helical. This fold information was combined with sequence information using the SeqFold algorithm to search for modeling templates. Independently, the FoldMiner algorithm was used to search for templates that had structural folds similar to published coordinates of the homologous nAChR (1OED). Both SeqFold and Foldminer identified the same modeling template. The GlyRa1 sequence was aligned with this template using multiple scoring criteria. Refinement of the alignment closed gaps to produce agreement with labeling studies carried out on the homologous receptors of the superfamily. Structural assignment and refinement was achieved using Modeler. The final structure demonstrated a cavity within the core of a four-helix bundle. Residues known to be involved in modulating anesthetic potency converge on and line this cavity. This suggests that the binding sites for volatile anesthetics in the LGICs are the cavities formed within the core of transmembrane four-helix bundles. PMID- 15667137 TI - A new approach for the tissue-blood partition coefficients of neutral and ionized compounds. AB - A nonlinear model equation based on tissue composition (a content of lipids, proteins, and water) for the tissue-blood partition coefficients of compounds was further developed in this paper to account for the neutral and ionized forms of the compounds. The nonlinear model equation may be considered as the expressive form of the Hansch equation in nonlinear spaces (or multiphase system). Based on this model our nonlinear regression analysis for neutral and ionized compounds partitioning into kidney, brain, muscle, lung, liver, heart, and fat resulted in equations with high fitting power (training set: n=201, r=0.905, s=0.291, Q=0.890) and strong predictive power (test set: n=64, r=0.906, s=0.247). These results showed that the equilibrium distribution of a compound in a variety of tissues is essentially the equilibrium distribution of the compound in tissue (chemical) compositions. Neutral and ionic forms of a compound as well as in different tissue (chemical) compositions have a different mechanism of action in vivo. PMID- 15667139 TI - CoMFA and docking studies on glycogen phosphorylase a inhibitors as antidiabetic agents. AB - Glycogen phosphorylase (GP(a)) is a specific target for the design of inhibitors and may prevent glycogenolysis under high glucose conditions in type II diabetes. The carboxamides first reported by Hoover D. J. et al. (J. Med. Chem. 1998, 41, 2934-2938) are one of the major classes of GP(a) inhibitors other than glucose derivatives. The recent, X-ray crystallographic analyses (Oikonomakos et al. Biochim. Biophys. Acta 2003, 1647, 325-332) have revealed a distinct mechanism of action for these inhibitors, which bind at a new allosteric site away from the inhibitory and catalytic sites. To elucidate the essential structural and physicochemical requirements responsible for binding to the GP(a) enzyme and to develop predictive models, CoMFA and docking studies have been carried out on a series of indole-2-carboxamide derivates. The CoMFA model developed using pharmacophoric alignments and hydrogen-bonding fields demonstrated high predictive ability against the training (r2 = 0.98, q2 = 0.68) and the test set (r2pred = 0.85). Further the superimposition of PLS coefficient contour maps from CoMFA with the GP(a) active site (PDB: 1lwo) has shown a high level of compatibility. PMID- 15667140 TI - Pharmacophore identification, in silico screening, and virtual library design for inhibitors of the human factor Xa. AB - Factor Xa inhibitors are innovative anticoagulant agents that provide a better safety/efficacy profile compared to other anticoagulative drugs. A chemical feature-based modeling approach was applied to identify crucial pharmacophore patterns from 3D crystal structures of inhibitors bound to human factor Xa (Pdb entries 1fjs, 1kns, 1eqz) using the software LIGANDSCOUT and CATALYST. The complex structures were selected regarding the criteria of high inhibitory potency (i.e. all ligands show K(i) values against factor Xa in the subnanomolar range) and good resolution (i.e. at least 2.2 A) in order to generate selective and high quality pharmacophore models. The resulting chemical-feature based hypotheses were used for virtual screening of commercial molecular databases such as the WDI database. Furthermore, a ligand-based molecular modeling approach was performed to obtain common-feature hypotheses that represent the relevant chemical interactions between 10 bioactive factor Xa inhibitors and the protein, respectively. In a next step a virtual combinatorial library was designed in order to generate new compounds with similar chemical and spatial properties as known inhibitors. The software tool ILIB DIVERSE was used for this procedure in order to provide new scaffolds of this group of anticoagulants. Finally we present the combination of these two techniques, hence virtual screening was performed with selective pharmacophore models in a focused virtual combinatorial database. De novo derived molecular scaffolds that were able to adequately satisfy the pharmacophore criteria are revealed and are promising templates for candidates for further development. PMID- 15667141 TI - LigandScout: 3-D pharmacophores derived from protein-bound ligands and their use as virtual screening filters. AB - From the historically grown archive of protein-ligand complexes in the Protein Data Bank small organic ligands are extracted and interpreted in terms of their chemical characteristics and features. Subsequently, pharmacophores representing ligand-receptor interaction are derived from each of these small molecules and its surrounding amino acids. Based on a defined set of only six types of chemical features and volume constraints, three-dimensional pharmacophore models are constructed, which are sufficiently selective to identify the described binding mode and are thus a useful tool for in-silico screening of large compound databases. The algorithms for ligand extraction and interpretation as well as the pharmacophore creation technique from the automatically interpreted data are presented and applied to a rhinovirus capsid complex as application example. PMID- 15667142 TI - Linear and nonlinear methods in modeling the aqueous solubility of organic compounds. AB - Solubility data for 930 diverse compounds have been analyzed using linear Partial Least Square (PLS) and nonlinear PLS methods, Continuum Regression (CR), and Neural Networks (NN). 1D and 2D descriptors from MOE package in combination with E-state or ISIS keys have been used. The best model was obtained using linear PLS for a combination between 22 MOE descriptors and 65 ISIS keys. It has a correlation coefficient (r2) of 0.935 and a root-mean-square error (RMSE) of 0.468 log molar solubility (log S(w)). The model validated on a test set of 177 compounds not included in the training set has r2 0.911 and RMSE 0.475 log S(w). The descriptors were ranked according to their importance, and at the top of the list have been found the 22 MOE descriptors. The CR model produced results as good as PLS, and because of the way in which cross-validation has been done it is expected to be a valuable tool in prediction besides PLS model. The statistics obtained using nonlinear methods did not surpass those got with linear ones. The good statistic obtained for linear PLS and CR recommends these models to be used in prediction when it is difficult or impossible to make experimental measurements, for virtual screening, combinatorial library design, and efficient leads optimization. PMID- 15667143 TI - ZINC--a free database of commercially available compounds for virtual screening. AB - A critical barrier to entry into structure-based virtual screening is the lack of a suitable, easy to access database of purchasable compounds. We have therefore prepared a library of 727,842 molecules, each with 3D structure, using catalogs of compounds from vendors (the size of this library continues to grow). The molecules have been assigned biologically relevant protonation states and are annotated with properties such as molecular weight, calculated LogP, and number of rotatable bonds. Each molecule in the library contains vendor and purchasing information and is ready for docking using a number of popular docking programs. Within certain limits, the molecules are prepared in multiple protonation states and multiple tautomeric forms. In one format, multiple conformations are available for the molecules. This database is available for free download (http://zinc.docking.org) in several common file formats including SMILES, mol2, 3D SDF, and DOCK flexibase format. A Web-based query tool incorporating a molecular drawing interface enables the database to be searched and browsed and subsets to be created. Users can process their own molecules by uploading them to a server. Our hope is that this database will bring virtual screening libraries to a wide community of structural biologists and medicinal chemists. PMID- 15667144 TI - Structure-related statistical singularities along protein sequences: a correlation study. AB - A data set composed of 1141 proteins representative of all eukaryotic protein sequences in the Swiss-Prot Protein Knowledge base was coded by seven physicochemical properties of amino acid residues. The resulting numerical profiles were submitted to correlation analysis after the application of a linear (simple mean) and a nonlinear (Recurrence Quantification Analysis, RQA) filter. The main RQA variables, Recurrence and Determinism, were subsequently analyzed by Principal Component Analysis. The RQA descriptors showed that (i) within protein sequences is embedded specific information neither present in the codes nor in the amino acid composition and (ii) the most sensitive code for detecting ordered recurrent (deterministic) patterns of residues in protein sequences is the Miyazawa-Jernigan hydrophobicity scale. The most deterministic proteins in terms of autocorrelation properties of primary structures were found (i) to be involved in protein-protein and protein-DNA interactions and (ii) to display a significantly higher proportion of structural disorder with respect to the average data set. A study of the scaling behavior of the average determinism with the setting parameters of RQA (embedding dimension and radius) allows for the identification of patterns of minimal length (six residues) as possible markers of zones specifically prone to inter- and intramolecular interactions. PMID- 15667145 TI - Toward an optimal procedure for PC-ANN model building: prediction of the carcinogenic activity of a large set of drugs. AB - The performances of the three novel QSAR algorithms, principal component artificial neural network modeling method combining with three factor selection procedures named eigenvalue ranking, correlation ranking, and genetic algorithm (ER-PC-ANN, CR-PC-ANN, PC-GA-ANN, respectively), are compared by application of these model to the prediction of the carcinogenic activity of a large set of drugs (735 drugs) belonging to a diverse type of compounds. A total number of 1350 theoretical descriptors are calculated for each molecule. The matrix of calculated descriptors (with 735 x 1350 dimension) is subjected to PCA. 95% of the variances in the matrix are explained by the first 137 principal components (PC's). From the pool of 137 PC's, the factor selection methods (ER, CR, and GA) are employed to select the best set of PC's for PC-ANN modeling. In the ER-PC ANN, the PC's are successively entered into the ANN based on their decreasing eigenvalue. In the CR-PC-ANN, the ANN is first employed to model the nonlinear relationship between each one of the PC's and the carcinogen activity separately. Then, the PC's are ranked based on their decreasing correlating ability and entered to the input layer of the network one after another. Finally, a search algorithm (i.e. genetic algorithm) is used to find the best set of PC's. Both the external and cross-validation methods are used to validate the performances of the resulting models. One is able to see that the results obtained by the PC-GA ANN and CR-PC-ANN procedures are superior to those resulted from the EV-PC-ANN. Comparison of the results reveals that the results produced by the PC-GA-ANN algorithm are better than those produced by CR-PC-ANN. However, the difference is not significant. PMID- 15667146 TI - Use of structure descriptors to discriminate between modes of toxic action of phenols. AB - Two classification models were developed based on a data set of 220 phenols with four associated Modes of Toxic Action (MOA). Counter-propagation neural networks (CPG NN) and multinomial logistic regression (multinom) were used as classification methods. The combination of topological autocorrelation of empirical pi-charge and sigma-electronegativity and of surface autocorrelation of hydrogen-bonding potential resulted in a 21-dimensional model that successfully discriminated between the four MOAs. Its overall predictive power was estimated to 92% using 5-fold cross-validation. Subsequently, a simple score for the distance to the training data was used to determine the prediction space of the model and used in an exploratory study on the phenols contained in the open NCI database. The use of a prediction space metric proved indispensable for the screening of such a diverse database. The prediction space covered by the proposed model is still of rather local nature which is either caused by the limited diversity and size of the training set or by the high dimensionality of the descriptors. PMID- 15667150 TI - Phenomenon observed at the onset of micellization using static light scattering. AB - A phenomenon was observed near the critical micelle concentration (cmc) of surfactants using static light scattering. This consists of an unexpected peak in light scattering as the concentration varies between zero and above the cmc. This work studied three different surfactants: the two ionic surfactants hexadecyltrimethylammonium bromide (CTABr) and sodium dodecyl sulfate (SDS) and the nonionic surfactant Triton X-100. Peaks were observed for all three under different conditions such as varying ionic strengths and different concentration paths. These peaks are real, are reproducible, and appear to have static properties. PMID- 15667151 TI - Lateral heterogeneity of dipalmitoylphosphatidylethanolamine-cholesterol Langmuir Blodgett films investigated with imaging time-of-flight secondary ion mass spectrometry and atomic force microscopy. AB - To better understand the influence of cholesterol (CH) on dipalmitoylphosphatidylethanolamine (DPPE), Langmuir-Blodgett (LB) model membranes of DPPE with varying amounts of cholesterol were imaged by time-of flight secondary ion mass spectrometry (ToF-SIMS) and atomic force microscopy (AFM). Cholesterol has a condensing effect on DPPE that at low cholesterol concentrations results in lateral heterogeneity of the LB monolayer. At 4:1 DPPE/CH, islands of DPPE/CH phase exist with a connected DPPE phase. As the concentration of cholesterol is increased, the percolation threshold is crossed and the DPPE/CH phase islands connect to separate the DPPE phase (2:1 DPPE/CH). Finally, at 50 mol % cholesterol a single homogeneous DPPE/CH phase LB monolayer exists. ToF-SIMS of the DPPE/CH phase provides a lower ion signal for the characteristic lipid fragments and substrate apparently owing to the higher molecular density induced by cholesterol. AFM data indicate that the DPPE/CH phase is lower in height than the DPPE phase. As phosphatidylethanolamine is predominant in the inner lipid leaflet of cellular membranes, this work has implications for the understanding of cholesterol domains in the inner leaflet of cells. PMID- 15667152 TI - 2D spatially periodic architectures via the drying of 1D holographically photopatterned polymer solutions. AB - We demonstrate an efficient and versatile method for selectively generating 1D and 2D periodic polymer structures in the submicron to tens of micron range by the directed drying of 1D photopatterned polymer solutions. Ultraviolet (UV) holographic lithography (Xia et al. Chem. Rev. 1999, 99, 1823-1848) is initially used to create 1D periodic cross-link and density variations in the polymer/volatile solvent solutions. These variations act as anisotropic barriers (walls) that direct the subsequent solvent evaporation process. Somewhat akin to directional drying (Allain and Limat Phys. Rev. Lett. 1995, 74, 2981-2984) and directional solidification (Pelc, P. Dynamics of Curved Fronts; Academic Press: San Diego, CA, 1988) experiments, the drying exhibits channel-like interface propagation behavior. The combination of the instabilities and minimization of the interface area during drying can be effectively used to produce larger scale 2D pseudohexagonal polymer-strut structures, or by the addition of a monomeric component to the polymer solution, the instabilities can be suppressed, resulting in the formation of 1D linear gratings. PMID- 15667153 TI - Novel polymer patterns formed by lithographically induced self-assembly (LISA). AB - A variety of self-assembly patterns, e.g., concentric rings, rods, and pillars, in polymer thin film have been achieved by lithographically induced self-assembly (LISA) in this study. The variations of the LISA patterns are controlled by many operation factors, such as the choice of the polymers, mask topology, process temperatures, surface tension, and so forth. It was found that as the inter- and intramolecular hydrogen bond interactions were incorporated into the polymer [poly(methyl methacrylate-co-methacrylic acid)], novel LISA patterns such as rods and corresponding arrays (concentric ring, triangle, hexagonal, etc.) were formed, in addition to the pillar arrays formed in poly(methyl methacrylate) under the same experimental conditions. The origins of the rod array are determined by the topology of the masks. Under a plain mask, the patterns developed from any nonuniform defects (spots) on the mask or polymer thin film and propagated outward. However, under a mask with protruding flat patterns, the rod patterns started along the edge of the protrusions and propagated inward. By increasing the process temperature, those novel rods and corresponding array patterns could transform back to pillar or pillar arrays. PMID- 15667154 TI - Solvent-adaptable silver nanoparticles. AB - A simple and efficient way of obtaining silver nanoparticles that are dispersible both in organic and in aqueous solvents using a single capping agent is described. The silver nanoparticles are initially prepared in water in the presence of aerosol OT [sodium bis(2-ethylhexyl)-sulfosuccinate, AOT]. Thereafter, transfer of the AOT-capped silver nanoparticles to an organic phase is induced by the addition of a small amount of orthophosphoric acid during shaking of the biphasic mixture. The AOT-stabilized silver nanoparticles could be separated out from the organic phase in the form of a powder. The hydrophobic nanoparticles thus prepared are stable and are readily resuspended in a variety of other polar (including water) and nonpolar solvents without further surface treatment. The amphiphatic nature of the silver surface is brought about by a small orientational change in the AOT monolayer on the silver surface in response to the polarity of the solvent. PMID- 15667155 TI - Controlling diffusion in sol-gel derived monoliths. AB - Redox probes were trapped within a silica monolith prepared in part with organoalkoxysilanes containing a quaternary ammonium functional group. The diffusion coefficients of the entrapped molecules were measured as the gels were slowly dried using chronoamperometry and cyclic voltammetry with ultramicroelectrodes. Gel-entrapped cobalt(II) tris(bipyridine) (Co(bpy)(3)(2+)) diffuses at rates similar to that measured in the sols by incorporating a small amount of the positively charged functional group in the matrix. In comparison, the diffusion coefficient of gel-entrapped ferricyanide (Fe(CN)(6)(3-)) drops an order of magnitude relative to its value in the sol soon after gelation. These results demonstrate the ease at which diffusion in hydrated gels can be easily controlled by simply changing the charge on the walls of the silica host. PMID- 15667156 TI - Micropatterning of cells using modulated magnetic fields. AB - A new technique of cell micropatterning was presented. Mouse osteoblast cells (MC3T3-E1) were seeded on a substrate whose surface was exposed to a periodically modulated magnetic field (a line pattern with a 200- or 600-microm pitch) produced by a field modulator inserted into a homogeneous magnetic field of 1 T generated by an electromagnet. The cells were trapped consistent with the line profile of the modulated field. The trapping efficiency was enhanced by adding Mn(II)EDTA (paramagnetic) to the cultivation medium. The cells were subsequently incubated in the magnetic field. The same technique was applied to whole blood to pattern red blood cells. PMID- 15667157 TI - Facile synthesis of polyaniline nanofibers using chloroaurate acid as the oxidant. AB - This work demonstrated a facile route to the synthesis of polyaniline (PANI) nanofibers by polymerization of aniline using chloroaurate acid (HAuCl(4)) as the oxidant. The reduction of AuCl(4)(-) is accompanied by oxidative polymerization of aniline, leading to uniform PANI nanofibers with a diameter of 35 +/- 5 nm and aggregated gold nanoparticles which can precipitate from the liquid phase during the reaction. The resultant PANI nanofibers and gold particles were characterized by means of different techniques, such as UV-vis, FTIR spectroscopy, and scanning and transmission electron microscopy methods. It is found that the gold aggregates are capped with polyaniline, and the conductivity of the fibers is around 0.16 S/cm. PMID- 15667158 TI - Necessity of aromatic carboxylate anions to be planar to induce growth of cationic micelles. AB - It is well-known that some aromatic anions have the ability to induce viscoelastic transformation in aqueous solutions of cationic surfactants even at added salt concentrations as low as 10-20 mM. This behavior is associated with the formation of an entangled network of elongated micelles. However, the effect of aromatic ring substituents on the anion's ability to promote rapid micelle growth is not well-understood. We have performed ab initio calculations of the carbonyl group rotation barriers in a series of substituted benzoate and naphthoate anions at the MP2/STO-3G level of theory. It was found that aromatic carboxylates, known to be particularly effective in causing sphere-rod transition in cationic micelles, preferably adopt conformations with the COO(-) group in the same plane as the ring(s). This structural preference can be attributed to either intramolecular hydrogen bonding (o-hydroxyl derivatives) or pi-conjugation effects (m- and p-halogenated derivatives). In the former case the barrier to rotation is 40-50 kcal/mol, whereas in the latter case the threshold value is around 3.0 kcal/mol. Propensity for the planar conformation correlates with a greater depth of counterion penetration into the micelle surface, as inferred from NMR experiments, compared to the anions with less hindered carbonyl rotation. This points to favorable hydrophobic interactions between the surfactant methylene groups and the aromatic ring(s) of the anion as a possible explanation for the rapid growth of cationic micelles observed upon addition of certain aromatic carboxylates. PMID- 15667159 TI - Breakdown of colloid filtration theory: role of the secondary energy minimum and surface charge heterogeneities. AB - The mechanisms and causes of deviation from the classical colloid filtration theory (CFT) in the presence of repulsive Derjaguin-Landau-Verwey-Overbeek (DLVO) interactions were investigated. The deposition behavior of uniform polystyrene latex colloids in columns packed with spherical soda-lime glass beads was systematically examined over a broad range of physicochemical conditions, whereby both the fluid-phase effluent particle concentration and the profile of retained particles were measured. Experiments conducted with three different-sized particles in a simple (1:1) electrolyte solution reveal the controlling influence of secondary minimum deposition on the deviation from CFT. In a second series of experiments, sodium dodecyl sulfate (SDS) was added to the background electrolyte solution with the intent of masking near-neutrally charged regions of particle and collector surfaces. These results indicate that the addition of a small amount of anionic surfactant is sufficient to reduce the influence of certain surface charge inhomogeneities on the deviation from CFT. To verify the validity of CFT in the absence of surface charge heterogeneities, a third set of experiments was conducted using solutions of high pH to mask the influence of metal oxide impurities on glass bead surfaces. The results demonstrate that both secondary minimum deposition and surface charge heterogeneities contribute significantly to the deviation from CFT generally observed in colloid deposition studies. It is further shown that agreement with CFT is obtained even in the presence of an energy barrier (i.e., repulsive colloidal interactions), suggesting that it is not the general existence of repulsive conditions which causes deviation but rather the combined occurrence of "fast" and "slow" particle deposition. PMID- 15667160 TI - Association of octyl-modified poly(acrylic acid) onto unilamellar vesicles of lipids and kinetics of vesicle disruption. AB - Water-soluble polymers containing a few hydrophobic anchors are known to bind onto lipid vesicles and are used as stabilizers of liposome-based formulas. In contrast, polymers with high hydrophobicity destabilize the lipid bilayers. With macromolecules of intermediate hydrophobic/hydrophilic balance, a gradual sweep of the stabilization-destabilization capacity can be achieved and is considered as promising triggered systems for drug release, although the mechanism of permeabilization and membrane breakage using polymers is essentially conjectural to date. As a model system, we used short octyl-modified poly(acrylic acid)s (MW 8000 g/mol) sensitive to pH, temperature, and ionic strength in conjunction with small unilamellar vesicles mainly comprised of DPPC or egg-PC. Kinetics of vesicle fragmentation was followed using static and dynamic light scattering. Polymer adsorption was studied by nonradiative energy transfer between pyrene labeled lipids and a naphthalene-modified polymer. The permeability of the vesicles was characterized by calcein leakage experiments. The key findings were (i) the lack of coupling between the density of bound polymer and the rate of disruption and (ii) the qualitative difference depending on whether the polymer contains or not isopropyl side groups. Point i relates to the increase of the rate of polymer adsorption with increasing bulk polymer concentration, while the breakage is essentially unaffected. Point ii relates to the stabilization of large membrane fragments (Stokes radius ca. 40 nm) in the presence of a polymer with no isopropyl side groups, while micelle-like assemblies (Stokes radius 8 nm) containing the lipids are obtained with an isopropyl-containing polymer of similar hydrophobicity. Both polymers display similar efficiency at disrupting small vesicles. The mechanism of polymer-induced disruption appears to differ markedly from the disruption steps now recognized for conventional (molecular) surfactant and is discussed on the basis of data obtained with different membrane fluidity, polymer structure, concentration, and hydrophilicity. PMID- 15667161 TI - Morphological evolution of inorganic crystal into zigzag and helical architectures with an exquisite association of polymer: a novel approach for morphological complexity. AB - The morphology of potassium sulfate (K(2)SO(4)) crystals grown in a viscous polymer solution of poly(acrylic acid) (PAA) was remarkably changed from the tilted columnar assembly into zigzag and helical architectures with increasing PAA concentration. The habit modification of orthorhombic K(2)SO(4) with adsorption of PAA molecules on a specified crystal face fundamentally led to the formation of tilted unit crystals. Concurrently with the habit modification, a diffusion-limited condition controlling the assembly of tilted units was achieved in the presence of PAA molecules in the matrix. Various complex morphologies, including zigzag and helical assembly, emerged through the formation of twinned crystals with the variation of the diffusion condition. Understanding the morphogenesis observed in this report would provide a novel approach for sophisticated crystal design by using an exquisite association of organic and inorganic materials. PMID- 15667163 TI - Comparison of Si-O-C interfacial bonding of alcohols and aldehydes on Si(111) formed from dilute solution with ultraviolet irradiation. AB - Aliphatic alcohols and aldehydes were reacted with the Si(111)-H surface to form Si-O-C interfacial bonds from dilute solution by using ultraviolet light. The resulting monolayers were characterized by using transmission infrared spectroscopy, spectroscopic ellipsometry, and contact angle measurements. The effect of different solvents on monolayer quality is presented. The best monolayers were formed from CH(2)Cl(2). The optimized monolayers were thoroughly characterized to determine the film structure and monolayer stability. The UV promoted, alcohol-functionalized, and aldehyde-functionalized monolayers are of comparable quality to those previously prepared by other means. Although both molecules are tethered through a Si-O-C bond, the film reactivity is distinctly different with the aldehyde films being more chemically resistant. The differences in chemical reactivity, vibrational spectra, hydrophobicity, and ellipsometric thickness between the alcohol and aldehyde monolayers are attributed to a difference in molecular coverage and monolayer formation. PMID- 15667162 TI - Plasma deposition and surface characterization of oligoglyme, dioxane, and crown ether nonfouling films. AB - Plasma-deposited PEG-like films are emerging as promising materials for preventing protein and bacterial attachment to surfaces. To date, there has not been a detailed surface analysis to examine the chemistry and molecular structure of these films as a function of both precursor size and structure. In this paper, we describe radio-frequency plasma deposition of a series of short-chain oligoglymes, dioxane, and crown ethers onto glass cover slips to create poly(ethylene glycol)-like coatings. The resultant films were characterized by X ray photoelectron spectroscopy (XPS), time-of-flight secondary ion mass spectrometry (TOF-SIMS), dynamic contact angle goniometry, and radiolabeled fibrinogen adsorption. Detailed analysis of the high-mass (120-300 m/z) TOF-SIMS oligoglyme film spectra revealed six classes of significant fragments. Two new models are proposed to describe how these fragments could be formed by distinct film-building processes: incorporation of intact and fragmented precursor molecules. The models also provide for the incorporation of hydrocarbon--a species that is not present in the precursors but is evidenced in XPS C(1s) spectra of these films. Two additional models describe the effects of incorporating intact and fragmented cyclic precursors. PMID- 15667164 TI - Intercalation and photophysical characterization of 1-pyrenemethylamine in zirconium phosphate layered materials. AB - The ion exchange of the luminescent probe 1-pyrenemethylamine (PYMA) into zirconium phosphate (ZrP) layered materials has been accomplished. The matrices used were the hexahydrated 10.3 A phase of ZrP (10.3 A ZrP, where 10.3 A represents the interlayer distance) and butylammonium-exchanged ZrP (BAZrP) with an expanded 18.6 A interlayer distance. The XRPD patterns for the 10.3 A ZrP after PYMA exchange (PYMA-exchanged ZrP), at high PYMA concentrations, show an increase in the interlayer distance from 10.3 A in unexchanged 10.3 A ZrP to 23.5 A in PYMA-exchanged ZrP, indicating PYMA intercalation. The luminescence spectrum for the PYMA-exchanged ZrP exhibits an excimer band at 458 nm that is absent in the luminescence spectrum of PYMA in aqueous solution at low concentrations. The intensity of the excimer emission increased at low PYMA concentrations. These results are in contrast to experiments using the BAZrP matrix. The XRPD patterns for PYMA-exchanged BAZrP do not show changes in the interlayer distance, which suggests that PYMA is not being intercalated and is only surface bound. The luminescence spectrum for PYMA-exchanged BAZrP exhibits a lower emission intensity in its excimer band, at different PYMA concentrations, compared with the PYMA-exchanged ZrP excimer band. For PYMA-exchanged ZrP, we propose a process in which exchange at low PYMA concentrations occurs at external surface sites with clustering promoting excimer formation followed by exchange at high PYMA concentrations occurring at interior sites reducing excimer formation. PMID- 15667165 TI - Structural characterization of single-walled carbon nanotube bundles by experiment and molecular simulation. AB - A procedure, combining molecular simulation, Raman spectroscopy, and standard nitrogen adsorption, is developed for structural characterization of single walled carbon nanotube (SWNT) samples. Grand canonical Monte Carlo simulations of nitrogen adsorption are performed on the external and internal adsorption sites of homogeneous arrays of SWNTs of diameters previously determined by Raman spectroscopy of the sample. The results show the importance of the peripheral grooves of a nanotube bundle at low relative pressure and the insensitivity of nanotube diameter toward adsorption on the external surface of the bundle at higher pressures. Simulations also reveal that samples containing thin nanotubes have less internal adsorption capacity that saturates at lower pressure than those comprising large diameter nanotubes. The fraction of open-ended nanotubes in a sample can be estimated by scaling the simulated internal adsorption inside nanotubes to obtain a near perfect fit between simulated and experimental isotherms. This procedure allows extrapolation of adsorption properties to conditions in which all nanotubes in the sample are open-ended. PMID- 15667166 TI - Dependence of the molecular aggregation state of octadecylsiloxane monolayers on preparation methods. AB - The molecular aggregation state of octadecylsiloxane monolayers on Si-wafer substrate surfaces prepared from octadecyltrimethoxysilane (OTMS) or octadecyltrichlorosilane (OTS) was investigated on the basis of grazing incidence X-ray diffraction (GIXD), Fourier transform infrared spectroscopy (FT-IR), contact angle measurement, field emission scanning electron microscopy (FE-SEM), and scanning force microscopy (SFM). The OTMS monolayer was prepared by using the chemical vapor adsorption (CVA) method, and the OTS monolayers, which were used as reference samples, were prepared either by chemisorption (OTS-S) or by the water-cast method (OTS-W). The GIXD, FT-IR, lateral force microscopic (LFM) measurements, and FE-SEM observation revealed that the alkyl chains in the OTMS monolayers prepared using the CVA method are in an amorphous state at room temperature. According to the LFM measurement, the transition temperature from the hexagonal crystalline phase to the amorphous phase was found to be ca. 333 K for the OTS-S monolayer prepared by the chemisorption method. However, the phase transition was not observed in the OTMS monolayer prepared by the CVA method. Also, the atomic force microscopic (AFM) observation and the contact angle measurement showed that the OTMS monolayer prepared by the CVA method has a uniform surface when compared to the OTS monolayers. These results indicated that organosilane compounds in the monolayer prepared by the CVA method were immobilized on the Si-wafer substrate surface in an amorphous state, which was quite different from the hexagonal crystalline state obtained by the chemisorption and water-cast methods. PMID- 15667167 TI - Macroscopic-wetting anisotropy on the line-patterned surface of fluoroalkylsilane monolayers. AB - Micropatterned fluoroalkylsilane monolayer surfaces with liquidphilic/liquidphobic area (line width 1-20 microm) were prepared with few defects by vacuum ultraviolet (VUV) photolithography. The anisotropic wetting of a macroscopic droplet with a 0.5-5 mm diameter on the micropatterned surfaces was investigated. The strong anisotropy of the contact angle and the sliding angle and droplet distortion for fluoroalkylsilane/silanol patterned surfaces was attributed to the difference in the energy barrier of wetting between parallel and orthogonal lines. The wetting anisotropy decreased with decreases in the liquidphilic area. Fluoroalkylsilane/alkylsilane patterned surfaces with small differences in the surface free energies of the components showed anisotropic wetting only for the low-surface-tension liquids. PMID- 15667168 TI - Supramolecular structures of coronene and alkane acids at the Au(111)-solution interface: a scanning tunneling microscopy study. AB - Scanning tunneling microscopy (STM) is utilized to study the solution-solid interface formed between Au(111) and solutions of coronene in hexanoic, heptanoic, and octanoic acids. In all three cases adsorbed coronene is observed and lays flat on the metal surface. Heptanoic and hexanoic acid solutions produce a hexagonal symmetry monolayer. For the heptanoic and hexanoic cases, dipole image dipole interactions and H bonding stabilize a surface structure in which 12 acid molecules surround each coronene and produce a coronene spacing of 1.45 nm. In the case of octanoic acid as solvent, the incorporation of the solvent into the monolayer is not as strongly favored. The coronene spacing can range from close-packed (1.2 nm) with no solvent presumed present in the monolayer, to 1.50 nm with up to 12 solvent molecules surrounding each coronene. The close-packed regions have hexagonal symmetry, as do those with the largest (1.5 nm) spacing. Heptanoic acid solutions give the clearest STM images and are associated with the most stable two-component monolayer. The present paper demonstrates that non covalent interactions at the solution-metal interface can lead to complex multicomponent monolayer structures. PMID- 15667169 TI - Effect of temperature on carbon-black agglomeration in hydrocarbon liquid with adsorbed dispersant. AB - Suspensions of carbon black in oil, stabilized with adsorbed polyisobutylene succinimide (PIBSI) dispersant, are commonly used as model systems for investigating the soot-handling characteristics of motor oils. The structure of the carbon-black agglomerates changes dramatically with temperature; this results in a concomitant change in the suspension rheology. Linear and nonlinear rheological experiments indicate a large increase of the interparticle attractions as the temperature is raised. To elucidate the origin of this behavior, we investigate the effect of temperature on the stabilizing effect of the dispersant. Measurements of adsorption isotherms of the dispersant on carbon black indicate that there is little variation of the binding energy with temperature. Intrinsic viscosity measurements of PIBSI dispersants in solution clearly exhibit an inverse dependence of the dispersant chain dimension with temperature. These results suggest that the temperature-dependent changes in the chain conformation of the PIBSI dispersant are primarily responsible for the changes in the dispersion rheology, and we propose a simple model to account for these data. PMID- 15667170 TI - Local packing environment strongly influences the frictional properties of mixed CH3- and CF3-terminated alkanethiol SAMs on Au(111). AB - Compositionally mixed, self-assembled monolayers (SAMs) derived from 16,16,16 trifluorohexadecanethiol and a normal alkanethiol, either hexadecanethiol or pentadecanethiol, were formed on Au(111) substrates. The relative composition of the films was determined using X-ray photoelectron spectroscopy and was found to approximately equal the equimolar composition of the isooctane solution from which they were formed. The frictional properties of the mixed films were measured on the nanometer scale using atomic force microscopy and were observed to decrease when the chain length of the CH(3)-terminated component was shortened by one methylene unit (i.e., when hexadecanethiol was replaced by pentadecanethiol). For comparison, the frictional properties of a mixed-chain length CH(3)-terminated SAM derived from hexadecanethiol and pentadecanethiol in a 1:1 ratio was also examined. In contrast to the mixed CF(3)/CH(3) system, the latter mixed-chain-length system exhibited relatively higher friction when compared to single-component SAMs derived solely from either hexadecanethiol or pentadecanethiol. For both types of mixed films, the change in frictional properties that occurs as a result of modifying the position of neighboring terminal groups with respect to the surface plane is discussed in terms of the influence of local packing environments on interfacial energy dissipation (friction). PMID- 15667171 TI - Wetting and wetting transitions on copper-based super-hydrophobic surfaces. AB - Rough and patterned copper surfaces were produced using etching and, separately, using electrodeposition. In both of these approaches the roughness can be varied in a controlled manner and, when hydrophobized, these surfaces show contact angles that increase with increasing roughness to above 160 degrees . We show transitions from a Wenzel mode, whereby the liquid follows the contours of the copper surface, to a Cassie-Baxter mode, whereby the liquid bridges between features on the surface. Measured contact angles on etched samples could be modeled quantitatively to within a few degrees by the Wenzel and Cassie-Baxter equations. The contact angle hysteresis on these surfaces initially increased and then decreased as the contact angle increased. The maximum occurred at a surface area where the equilibrium contact angle would suggest that a substantial proportion of the surface area was bridged. PMID- 15667172 TI - Lateral intermolecular forces in the physisorbed state: surface field polarization of benzene and n-hexane at the water/ and mercury/vapor interfaces. AB - The available experimental data on the dependence of the surface tensions of water and mercury on the adsorption of benzene and hexane from the vapor phase are critically analyzed and interpreted to obtain the two-dimensional second virial coefficients [B(2)(T)] for these adsorbed nonpolar molecules. Calculations based on the unperturbed Lennard-Jones (L-J) 12-6 formalism for benzene and the related 12-5 Salem formalism for long chains in two dimensions for hexane require that B(2)(T) should be negative for both adsorbates. On water, the experimental data indicate that B(2)(T) for both molecules is less negative than expected from the unperturbed L-J and Salem estimates, and on mercury the B(2)(T) values from experiment are positive. These findings are analyzed first in terms of a possible reduction in the attractive component of the potential of mean force between physisorbed molecules arising from their frequency-dependent interaction with their electrostatic images in the bulk phases, as described by McLachlan. It is concluded that the McLachlan corrections are small for these molecules and surfaces. A second analysis considers the effect of an extra repulsion between the adsorbed molecules arising from the induction of dipoles normal to the interface by the surface electric field. Surface field polarization (SFP) accounts reasonably well for the experimental results, leading to estimates of the surface fields at the mercury and water surfaces which are consistent with estimates from contact potentials for mercury and computation from modeling the water surface. SFP may have a wide impact in determining the form of physisorption isotherms. PMID- 15667173 TI - Adsorption kinetics of p-nitrobenzenethiol self-assembled monolayers on a gold surface. AB - The adsorption kinetics of octadecanethiol (ODT) and p-nitrobenzenethiol (NBT) from ethanol solutions has been studied by means of contact angle, optical ellipsometry, angle-resolved X-ray photoelectron spectroscopy (ARXPS), and grazing angle attenuated total reflection Fourier transform infrared (FTIR) measurements. ODT data were used as a reference for the kinetics studies of film growth. The growth of self-assembled monolayers from dilute solutions follows Langmuir isotherm adsorption kinetics. A saturated film is formed within 5 h after immersion in solutions of concentrations ranging from 0.0005 to 0.01 mM. The density of the monolayer depends on the concentration of the solution. PMID- 15667174 TI - Wetting and self-cleaning properties of artificial superhydrophobic surfaces. AB - The wetting and the self-cleaning properties (the latter is often called the "Lotus-Effect") of three types of superhydrophobic surfaces have been investigated: silicon wafer specimens with different regular arrays of spikes hydrophobized by chemical treatment, replicates of water-repellent leaves of plants, and commercially available metal foils which were additionally hydrophobized by means of a fluorinated agent. Water droplets rolled off easily from those silicon samples which had a microstructure consisting of rather slender spikes with narrow pitches. Such samples could be cleaned almost completely from artificial particulate contaminations by a fog consisting of water droplets (diameter range, 8-20 microm). Some metal foils and some replicates had two levels of roughening. Because of this, a complete removal of all particles was not possible using artificial fog. However, water drops with some amount of kinetic impact energy were able to clean these surfaces perfectly. A substrate where pronounced structures in the range below 5 microm were lacking could not be cleaned by means of fog because this treatment resulted in a continuous water film on the samples. PMID- 15667175 TI - Hydrogen chemisorption on gallium oxide polymorphs. AB - The chemisorption of H(2) over a set of gallia polymorphs (alpha-, beta-, and gamma-Ga(2)O(3)) has been studied by temperature-programmed adsorption equilibrium and desorption (TPA and TPD, respectively) experiments, using in situ transmission infrared spectroscopy. Upon heating the gallium oxides above 500 K in 101.3 kPa of H(2), two overlapped infrared signals developed. The 2003- and 1980-cm(-1) bands were assigned to the stretching frequencies of H bonded to coordinatively unsaturated (cus) gallium cations in tetrahedral and octahedral positions [nu(Ga(t)-H) and nu(Ga(o)-H), respectively]. Irrespective to the gallium cation geometrical environment, (i) a linear relationship between the integrated intensity of the whole nu(Ga-H) infrared band versus the Brunauer Emmett-Teller surface area of the gallia was found and (ii) TPA and TPD results revealed that molecular hydrogen is dissociatively chemisorbed on any bulk gallium oxide polymorph following two reaction pathways. An endothermal, homolytic dissociation occurs over surface cus-gallium sites at T > 450 K, giving rise to Ga-H(I) bonds. The heat and entropy of this type I hydrogen adsorption were determined by the Langmuir's adsorption model as Deltah(I) = 155 +/- 25 kJ mol(-1) and Deltas(I) = 0.27 +/- 0.11 kJ mol(-1) K(-1). In addition, another exothermic, heterolytic adsorption sets in already in the low-temperature region. This type of hydrogen chemisorption involves surface Ga-O-Ga species, originating GaO-H and Ga-H(II) bonds which can only be removed from the gallia surface after heating under evacuation at T > 650 K. The measured desorption energy of this last, second-order process was equal to 77 +/- 10 kJ mol(-1). The potential of the H(2) chemisorption as a tool to measure or estimate the specific surface area of gallia and to discern the nature and proportion of gallium cation coordination sites on the surface of bulk gallium oxides is also analyzed. PMID- 15667176 TI - Influence of coadsorbed hydrogen on ethylene adsorption and reaction on a (radical3 x radical3)R30 degrees-Sn/Pt(111) surface alloy. AB - The effect of surface-bound hydrogen adatoms on adsorption, desorption, and reaction of ethylene (CH(2)=CH(2)) on a (radical3 x radical3)R30 degrees Sn/Pt(111) surface alloy with theta(Sn) = 0.33 was investigated by using temperature-programmed desorption (TPD) and Auger electron spectroscopy (AES). Preadsorbed H decreased the saturation coverage of chemisorbed ethylene and less H was required to completely block ethylene chemisorption on this alloy than that on Pt(111). This is also the first report of extensive H site-blocking of ethylene chemisorption on Pt(111). Preadsorbed H also decreased the desorption activation energy of ethylene on the alloy surface. The reaction chemistry of ethylene on this Sn/Pt(111) alloy is dramatically different than on the Pt(111) surface: the H-addition reaction channel taking ethylene to ethane on Pt(111) is totally inhibited on the alloy. This is important information for advancing understanding of the surface chemistry involved in hydrogenation and dehydrogenation catalysis. PMID- 15667177 TI - Piezoluminescence at the air-water interface through dynamic molecular recognition driven by lateral pressure application. AB - The steroid cyclophanes with a cyclic core consisting of a 1,6,20,25 tetraaza[6.1.6.1]paracyclophane connected to four steroid moieties (cholic acid or cholanic acid) through a flexible l-lysine spacer were spread on water as Langmuir monolayers. The pi-A isotherm of the cholic-type steroid cyclophane includes a transition to the condensed phase with a limiting area of approximately 2 nm(2). This value is close to the cross-sectional area of the steroid cyclophane with a standing-up conformation of the cholic acid moieties, strongly suggesting that the cavity converts from a two-dimensional cavity to a three-dimensional cavity upon compressing the monolayer. Surface-reflective fluorescence spectroscopy of the monolayer using an aqueous fluorescent probe (6 (p-toluidino)naphthalene-2-sulfonate (TNS)) showed an abrupt increase in the TNS fluorescence intensity at a molecular area of 2 nm(2). Efficient binding of the guest probe would occur upon the completion of the three-dimensional cavity. Repeated compression and expansion induces periodic changes in the fluorescence intensity. This indicates a piezoluminescence effect through the catch and release of the TNS guest upon dynamic cavity formation. Analyses of the binding behavior of TNS to the steroid cyclophane resulted in binding constants in the range of approximately (5-9) x 10(4) M(-1) which are similar to that observed in lipid bilayer media (K = 5.1 x 10(4) M(-1)). The fluorescence intensity within the condensed phase was significantly increased with increasing pressure, suggesting that suppression of the molecular motion of the bound TNS may retard the nonemission process. Similar monolayer experiments were carried out with the monolayer of the cholanic-type steroid cyclophane that cannot form an open conformation on water. Both the phase transition in the pi-A isotherm and the change in the fluorescence intensity were negligible, confirming that the dynamic characteristic of the cavity is indispensable for the efficient pressure-induced binding of the guest and the consequent luminescence. PMID- 15667178 TI - Characteristic self-motion of a camphor boat sensitive to ester vapor. AB - As a simple example of an autonomous motor, the self-motion of a camphor boat on water with changes in chemical stimuli was investigated. The nature of the self motion of a camphor boat changed characteristically with the addition of ester vapor (methyl n-butyrate) to a circular water chamber. Thus, continuous motion changed to oscillatory motion, and its period increased depending on the location between an ester droplet and the chamber, L. The surface tension in the water chamber was measured to clarify how the velocity of the self-motion changed with L. The nature of the self-motion is discussed in relation to the surface tension as the driving force. We believe that the present results may be useful for realizing artificial chemotaxis systems under nonlinear and isothermal conditions. PMID- 15667179 TI - Electrocapillary at contact: potential-dependent adhesion between a gold electrode and a mica surface. AB - Using the electrochemical surface forces apparatus, we investigated adhesion (from pull-off measurements) between gold and mica as the potential of the gold surface was changed externally. Measurements were performed at different concentrations of KClO(4) in a potential window where the gold electrode is ideally polarizable. At applied potentials where the gold-mica interactions are repulsive, we obtain double layer forces that are predictable by the Derjaguin Landau-Verwey-Overbeek (DLVO) theory of colloid stability but deviate from the theory at short range. At applied potentials where the gold-mica interactions are attractive, we observed a very strong dependence of adhesion on the applied potential, a result that cannot be directly related to DLVO theory. We show, however, that an approach based on electrocapillary thermodynamics can be employed to model the potential dependence of adhesion seen in our measurements. This electrocapillary approach presents evidence of charging at the gold-mica interface and stresses the relation between the charge within and outside of the contact area. PMID- 15667180 TI - Fractal analysis methods for solid alkane monolayer domains at SiO2/air interfaces. AB - A systematic evaluation of various fractal analysis methods is essential for studying morphologies of finite and noisy experimental patterns such as domains of long chain alkanes at SiO(2)/air interfaces. The derivation of trustworthy fractal dimensions crucially relies on the definition of confidence intervals for the assumed scaling range. We demonstrate that the determination of the intervals can be improved largely by comparing the scaling behavior of different morphological measures (area, boundary, curvature). We show that the combination of area and boundary data from coarse-grained structures obtained with the box counting method reveals clear confidence limits and thus credible morphological data. This also holds for the Minkowski density method. It also reveals the confidence range. Its main drawback, the larger swing-in period at the lower cutoff compared to the box-counting method, is compensated by more details on the scaling behavior of area, boundary, and curvature. The sandbox method is less recommendable. It essentially delivers the same data as box-counting, but it is more susceptible to finite size effects at the lower cutoff. It is found that the domain morphology depends on the surface coverage of alkanes. The individual domains at low surface coverage have a fractal dimension of approximately 1.7, whereas at coverages well above 50% the scaling dimension is 2 with a large margin of uncertainty at approximately 50% coverage. This change in morphology is attributed to a crossover from a growth regime dominated by diffusion-limited aggregation of individual domains to a regime where the growth is increasingly affected by annealing and the interaction of solid growth fronts which approach each other and thus compete for the alkane supply. PMID- 15667181 TI - Electrochemical nucleation of gold nanoparticles in a polymer film at a liquid liquid interface. AB - Simultaneous nucleation of gold nanoparticles and polymerization of tyramine has been carried out at an immiscible electrolyte interface. By transferring the gold ion of tetraoctylammoniumtetracloroaurate (TOAAuCl(4)) from the organic to the aqueous phase, a fast homogeneous electron transfer from the tyramine monomer reduces the gold ion. Electropolymerization then proceeds, and gold nanoparticles form. The newly formed nanoparticles act as nucleation sites for the deposition of the oligomers/polymer (and possibly vice versa). This results in gold nanoparticles stabilized in a polytyramine matrix. The size of the nanoparticles is controlled by the concentration of oligomers/polymer in solution. The polymer nanoparticle composite film was analyzed with TEM, XPS, and AFM. PMID- 15667182 TI - Electrochemistry of cytochrome C in aqueous and mixed solvent solutions: thermodynamics, kinetics, and the effect of solvent dielectric constant. AB - The solvent dielectric constant is considered an important factor in determining the redox potential of the heme-containing protein cytochrome c in solution. In this study, we investigate the electrochemical response of cytochrome c in aqueous/organic solvent mixtures (100% aqueous buffer, 30% acetonitrile, 40% dimethyl sulfoxide, and 50% methanol), reporting the redox potential (E degrees'), enthalpy, and entropy of reduction. The temperature dependence of the solvent dielectric constant (epsilon) was also measured. The results show that epsilon alone cannot regulate the E degrees' of cytochrome c in mixed solvent systems. The implications of the temperature dependence of epsilon on the validity of the thermodynamic data are also discussed. The effect of solvent and temperature on the electron-transfer rate constant, k(s), was determined in each solvent mixture. A substantial increase in the activation energy for electron transfer was observed in 40% DMSO. PMID- 15667183 TI - Electrochemical properties of mixed self-assembled monolayers on gold electrodes containing mercaptooctylhydroquinone and alkylthiols. AB - Mixed self-assembled monolayers of 2-(mercaptooctyl)hydroquinone (QH2) and alkylthiols were formed on gold electrodes in EtOH and the redox process of the hydroquinone moiety of QH2 was characterized by cyclic voltammetry (CV) in 0.1 M H(2)SO(4). The monolayers were formed at a series of QH2:alkylthiol ratios and the QH2:alkylthiol ratio in solution was compared to the electrochemical response from QH2 in the obtained monolayer. Mixed monolayers of QH2 with hexylthiol, dodecylthiol, and octadecylthiol were studied. The length of the alkylthiol is crucial for the electrochemical response from QH2 in the monolayer. The total concentration of thiols during monolayer formation and incubation times were also studied and low concentrations of < 2.5 mM and long incubation times gave rise to lower peak separation, lower peak half widths in the CVs of the mixed monolayers, and lower background current. The stability of a pure QH2 monolayer and a 1:4 QH2:hexylthiol monolayer toward high potentials of up to 1.5 V versus Ag/AgCl was also studied and it was observed that the mixed monolayer is significantly more stable than the pure QH2 monolayer. PMID- 15667184 TI - Chemical control of electrode functionalization for detection of DNA hybridization by electrochemical impedance spectroscopy. AB - We report sensitive label-free detection of DNA oligonucleotide sequences using ac impedance measurements. The surface attachment chemistry is critical, and using mixed self-assembled monolayers on a gold electrode results in much better performance than homogeneous self-assembled monolayers. Contrary to expectations, binding of the target sequence reduces rather than increases the charge transfer resistance. Similar behavior is observed on indium tin oxide electrodes, and we ascribe it to the hydrophilicity and rigidity of the DNA duplex that cause it to reside further from the electrode surface and facilitate the approach of negatively charged redox moieties to the interface. PMID- 15667185 TI - Modifying calf lung surfactant by hexadecanol. AB - Monolayer properties such as phase behavior, collapse pressure, and surface viscosity are determined by monolayer composition. Learning how to control these properties through simple additives is important to understanding lung surfactant monolayers and to designing optimal replacement surfactants for treatment of disease. The properties of Infasurf, a replacement lung surfactant derived from calf lung lavage organic extract, can be modified in a controlled fashion by adding hexadecanol (HD). Grazing incidence X-ray diffraction shows that the HD preferentially interacts with dipalmitoylphosphatidylcholine (DPPC), the main phospholipid component of Infasurf, in the same way as in binary mixtures of DPPC and HD. HD intercalates between the DPPC chains, which leads to a tighter packing of the two-dimensional lattice and greater stability of the solid phase. This molecular reorganization triggers changes at the macroscopic scale, leading to a greatly increased solid-phase fraction at a given surface pressure and order of magnitude increases in the surface shear viscosity. However, the collapse pressure decreases, and, hence, the minimum surface tension of the film increases. PMID- 15667186 TI - Protein resistance of surfaces prepared by sorption of end-thiolated poly(ethylene glycol) to gold: effect of surface chain density. AB - Nonspecific protein adsorption generally occurs at the biomaterial-tissue interface and usually has adverse consequences. Thus, surfaces that are protein resistant are eagerly sought with the expectation that these materials will exhibit improved biocompatibility. Surfaces modified with end-tethered poly(ethylene oxide) (PEO) have been shown to be protein-resistant to some degree. Although the mechanisms are unclear, it has been suggested that chain length, chain density, and chain conformation are important factors. To investigate the effects of PEO chain density, we selected a model system based on the chemisorption of chain-end thiolated PEO to a gold substrate. Chain density was varied by varying PEO solubility (proximity to cloud point) and incubation time in the chemisorption solution. The adsorption of fibrinogen and lysozyme to these surfaces was investigated. It was found that for 750 and 2000 MW PEO layers, resistance to fibrinogen increased with chain density and was maximal at a density of approximately 0.5 chains/nm(2) (80% decrease in adsorption compared to unmodified gold). As PEO chain density increased beyond 0.5/nm(2) adsorption increased. For PEO of 5000 MW the optimal chain density was 0.27/nm(2) and gave only a 60% reduction in fibrinogen adsorption. It is suggested that, at high chain density, the chemisorbed PEO is dehydrated giving a surface that is no longer protein resistant. The PEO-modified surfaces were found also to be resistant to lysozyme adsorption with reductions similar to, if somewhat less than, those for fibrinogen. The fibrinogen to lysozyme molar ratios were within the expected range for close-packed layers of these proteins in their native conformation and were relatively insensitive to PEO chain density and MW. This may suggest that such adsorption as did occur, even at chain densities giving minimum adsorption, may have been on patches of unmodified gold. PMID- 15667187 TI - Molecular gradients: an efficient approach for optimizing the surface properties of biomaterials and biochips. AB - A variety of molecular gradients of alkanethiols with the structure HS-(CH(2))(m) X (m = 15; X = COOH, CH(2)NH(2), or CH(3)) and oligo(ethylene glycol)-terminated alkanethiols with the structures HS-(CH(2))(15)-CO-NH-Eg(n) (n = 2, 4, or 6), HS (CH(2))(15)-CO-NH-Eg(2)-(CH(2))(2)-NH-CO-(CH(2))(4)-biotin, and HS-(CH(2))(15)-CO NH-Eg(6)-CH(2)-COOH were prepared on polycrystalline gold films. These gradients were designed to serve as model surfaces for fundamental studies of protein adsorption and immobilization phenomena. Ellipsometry, infrared spectroscopy, and X-ray photoelectron spectroscopy, operating in scanning mode, were used to monitor the layer composition, gradient profiles, tail group conformation, and overall structural quality of the gradient assemblies. The gradient profiles were found to be 4-10 mm wide, and they increased in width with increasing difference in molecular complexity between the thiols used to form the gradient. The oligo(ethylene glycol) thiols are particularly interesting because they can be used to prepare so-called conformational gradients, that is, gradients that display a variation in oligo(ethylene glycol) chain conformation from all trans on the extreme Eg(2,4) sides, via an amorphous-like phase in the mixing regimes, to helical at the extreme Eg(6) sides. We demonstrate herein a series of experiments where the above gradients are used to evaluate nonspecific binding of the plasma protein fibrinogen, and in agreement with previous studies, the highest amounts of nonspecifically bound fibrinogen were observed on all-trans monolayers, that is, on the extreme Eg(2,4) sides. Moreover, gradients between Eg(2) and a biotinylated analogue have been prepared to optimize the conditions for the immobilization of streptavidin. Ellipsometry and infrared spectroscopy reveal high levels of immobilization over a fairly broad range of compositions in the gradient regime, with a maximum between 50 and 60% of the biotinylated analogue in the monolayer. A pI gradient composed of (NH(3)(+)/COO(-))-terminated thiols was also prepared and evaluated with respect to its ability to separate differently charged proteins, pepsin, and lysozyme, on a solid surface. PMID- 15667188 TI - Hydrolysis reaction analysis of L-alpha-distearoylphosphatidylcholine monolayer catalyzed by phospholipase A2 with polarization-modulated infrared reflection absorption spectroscopy. AB - The hydrolysis reaction of L-alpha-distearoylphosphatidylcholine (DSPC) monolayers catalyzed by phospholipase A2 (PLA(2)) has been studied using polarization-modulated infrared reflection absorption spectroscopy (PM-IRRAS) with film balance measurements. The PM-IRRAS analysis provides quantitative information about the reaction efficiency at different surface pressures. It was found that the reaction efficiency of L-DSPC monolayer hydrolysis catalyzed by PLA(2) decreased with increasing surface pressure. At zero pressure (lift-off point), the hydrolysis reaction efficiency has the highest value of 45%. Increasing surface pressure leads to the decrease of the hydrolysis efficiency. Since the surface pressure is above 20 mN/m, the hydrolysis reaction nearly stopped. PM-IRRAS technique provides a powerful means to study the hydrolysis process catalyzed by phospholipase A2 at the air/water interface. PMID- 15667189 TI - Polymer-assisted dispersion of single-walled carbon nanotubes in alcohols and applicability toward carbon nanotube/sol-gel composite formation. AB - Single-walled carbon nanotubes (SWCNTs) were directly dispersed into various alcohols by sonicating the nanotubes in the presence of poly(4-vinylpyridine) (P4VP). Depending upon the alcohol, it was possible to disperse up to 0.3 g of SWCNTs per liter of alcohol using only 0.6 g of P4VP, and with solution stability greater than 6 weeks. Scanning electron microscopy of "bucky" paper prepared from the polymer-treated nanotubes revealed reduced bundle size compared to the corresponding untreated nanotube paper. Additionally, the applicability of the dispersion system in the formation of SWCNT/silica composites is demonstrated. PMID- 15667190 TI - Spatially correlated metallic nanostructures on self-assembled diblock copolymer templates. AB - Polymeric complexes based on diblock copolymers (polystyrene-block-4 vinylpyridine) hydrogen bonded with pentadecylphenol self-assemble under oscillatory shear flow into a highly ordered lamellar structure (Ikkala et al. Science 2002). Microtomed films of the lamellar structure form an array of "nanosheets"following immersion in methanol. We have exploited this nanosheet array as an extremely effective template to direct the spatial organisation of metallic (Pd) nanoclusters. The electroless deposition metal on the nanotemplates leads to morphologically complex nanostructured metallic films which were observed using atomic force microscopy and scanning electron microscopy. PMID- 15667191 TI - In situ detection of birefringent mesoscopic H and J aggregates of thiacarbocyanine dye in solution. AB - Polarized-light microscopy, fluorescence microscopy, atomic force microscopy as well as absorption and fluorescence spectroscopy were used to characterize mesoscopic structures of both supramolecular H and J aggregates of 3,3' disulfopropyl-5,5'-dichloro-9-methyl thiacarbocyanine dye in aqueous solution. Polarized-light microscopy visualizes in situ the mesoscopic morphology of the H and J aggregates and distinguishes between them by their own colors. The H aggregate having a fibrous structure showed negative birefringence, namely, the refractive index along the fiber short axis was higher than that of the long axis, so that pi-electron chromophores of the dye molecule are likely to orient along the short axis of the elongated fibers. The degree of birefringence of the H aggregate fiber was approximately -0.3. Investigations on the concentration dependence of the absorption spectra showed that the amount of J aggregates increased at the expense of a decrease in the amount of H aggregates. With respect to the J aggregates, a small dot morphology was observed at a relatively low dye concentration of 3.0 mM. With an increase of the dye concentration up to 10 mM, the morphology changed into mesoscopic fibers. In contrast, fluorescence microscopy for the fibrous J aggregates reveals that the constituent molecules are approximately aligned along the long axis of the fibers. PMID- 15667192 TI - Formation of colloidal CuO nanocrystallites and their spherical aggregation and reductive transformation to hollow Cu2O nanospheres. AB - In this work, we demonstrate that cuprous oxide Cu(2)O nanospheres with hollow interiors can be fabricated from a reductive conversion of aggregated CuO nanocrystallites without using templates. A detailed process mechanism has been revealed: (i) formation of CuO nanocrystallites; (ii) spherical aggregation of primary CuO crystallites; (iii) reductive conversion of CuO to Cu(2)O; and (iv) crystal aging and hollowing of Cu(2)O nanospheres. In this template-free process, Ostwald ripening is operative in (iv) for controlling crystallite size in shell structures and thus for precisely tuning the optical band gap energy (E(g)) of resultant semiconductor nanostructures. For the first time, a wealth of colorful Cu(2)O hollow nanospheres (outer diameters in 100-200 nm), with variable E(g) in the range of 2.405-2.170 eV, has been fabricated via this novel chemical route. Considering their unique hollow structure and facile tuning in band gap energy, the prepared Cu(2)O hollow spheres can be potentially useful for harvesting solar energy in the visible range. Possibility of fabrication of Cu-Cu(2)O nanocomposites has also been discussed. PMID- 15667193 TI - Atomic force microscopy study of beta-substituted-T7 oligothiophene films on mica: mechanical properties and humidity-dependent phases. AB - The structural and mechanical properties of Langmuir-Blodgett monolayer and multilayer films of 3",4""-didecyl-5,2'; 5',2"; 5",2'''; 5''',2""; 5"",2'''''; 5''''',2"""-heptathiophene-4'''-acetic acid on mica have been studied by atomic force microscopy (AFM) as a function of humidity, temperature, and applied force. The molecules orient with the carboxylic acid group pointing toward the mica surface and expose the alkyl side chains to the air interface. As the load applied by the AFM tip increases, the film is compressed easily from an initial height of 2 to 1.2 nm. After compression the films can support much higher loads without loss of height. The state of aggregation of the molecules was found to be sensitive to the environmental humidity, which induced reversible changes. Annealing the samples with monolayer or multilayer films resulted in irreversible changes when the temperature exceeded approximately 100 degrees C. PMID- 15667194 TI - Synthesis and optical properties of thiol-stabilized PbS nanocrystals. AB - Thiol-capped water-soluble PbS nanocrystals (NCs) stabilized with 1-thioglycerol, dithioglycerol, or a mixture of 1-thioglycerol/dithioglycerol (TGL/DTG) were prepared via one-stage synthesis at room temperature. We found that NCs stabilized with a TGL/DTG mixture show efficient and stable infrared photoluminescence centered in the second "biological window" (1050-1200 nm). Under optimized conditions, full width at half-maximum of the PL emission peak was from 70 to 100 nm. PbS NCs were stable to precipitation and aggregation for the time period from 2 to 3 months when stored in the dark under room temperature. Room-temperature photoluminescence quantum efficiency of NCs was from 7 to 10%. When NCs were stored at 37 degrees C, their PL emission red shifted, consistent with the NC growth. PMID- 15667196 TI - Modeling of composite latex particle morphology by off-lattice Monte Carlo simulation. AB - Composite latex particles have shown a great range of applications such as paint resins, varnishes, water borne adhesives, impact modifiers, etc. The high performance properties of this kind of materials may be explained in terms of a synergistical combination of two different polymers (usually a rubber and a thermoplastic). A great variety of composite latex particles with very different morphologies may be obtained by two-step emulsion polymerization processes. The formation of specific particle morphology depends on the chemical and physical nature of the monomers used during the synthesis, the process temperature, the reaction initiator, the surfactants, etc. Only a few models have been proposed to explain the appearance of the composite particle morphologies. These models have been based on the change of the interfacial energies during the synthesis. In this work, we present a new three-component model: Polymer blend (flexible and rigid chain particles) is dispersed in water by forming spherical cavities. Monte Carlo simulations of the model in two dimensions are used to determine the density distribution of chains and water molecules inside the suspended particle. This approach allows us to study the dependence of the morphology of the composite latex particles on the relative hydrophilicity and flexibility of the chain molecules as well as on their density and composition. It has been shown that our simple model is capable of reproducing the main features of the various morphologies observed in synthesis experiments. PMID- 15667195 TI - Surface fabrication of hollow microspheres from N-methylated chitosan cross linked with glutaraldehyde. AB - We have successfully prepared biocompatible and biodegradable hollow microspheres with sizes between 2 and 5 mum using cyclohexane droplets as a template and the N methylated chitosan (NMC) cross-linked with glutaraldehyde (GA) as the shell. The structure, morphology, and formation process of the hollow microspheres were characterized by FT-IR, (1)H and (13)C NMR, scanning electron microscopy (SEM), and transmission electron microscopy (TEM). The results revealed that the microspheres exhibited a very smooth and hollow structure. This work confirmed that the hollow microspheres were accomplished by fabricating on the basis of chemical cross-linking on the surface of the emulsion droplets and by removing cyclohexane as core. The results from SEM and TEM indicated that the emulsion droplets covered with cross-linked NMC in the oil-in-water system aggregated together to form a precipitate of microspheres by coagulating with acetone. Moreover, the cross-linked NMC on the surface of the microspheres continuously cured to form the tight shell, whereas the inner area became a cavity with increase of the aging time, leading to the hollow microspheres. In addition, an anti-infective drug, ofloxacin (Floxin), encapsulated in the microspheres more rapidly released to reach 90 wt % at pH 7.4 within 8 h than at pH 1.2. PMID- 15667197 TI - Molecular simulation to characterize the adsorption behavior of a fibrinogen gamma-chain fragment. AB - Implants invoke inflammatory responses from the body even if they are chemically inert and nontoxic. It has been shown that a crucial precedent event in the inflammatory process is the spontaneous adsorption of fibrinogen (Fg) on implant surfaces, which is typically followed by the presence of phagocytic cells. Interactions between the phagocyte integrin Mac-1 and two short sequences within the fibrinogen gamma chain, gamma190-202 and gamma377-395, may partially explain phagocyte accumulation at implant surfaces. These two sequences are believed to form an integrin binding site that is inaccessible when Fg is in its soluble state structure but then becomes available for Mac-1 binding following adsorption, presumably due to adsorption-induced conformational changes. The objective of this research was to theoretically investigate this possibility by using molecular dynamics simulations of the gamma-chain fragment of Fg over self assembled monolayer (SAM) surfaces presenting different types of surface chemistry. The GROMACS software package was used to carry out the molecular simulations in an explicit solvation environment over a 5 ns period of time. The adsorption of the gamma-chain of fibrinogen was simulated on five types of SAM surfaces. The simulations showed that this protein fragment exhibits distinctly different adsorption behavior on the different surface chemistries. Although the trajectory files showed that significant conformational changes did not occur in this protein fragment over the time frame of the simulations, it was predicted that the protein does undergo substantial rotational and translational motions over the surface prior to stabilizing in various preferred orientations. This suggests that the kinetics of surface-induced conformational changes in a protein's structure might be much slower than the kinetics of orientational changes, thus enabling the principles of adsorption thermodynamics to be used to guide adsorbing proteins into defined orientations on surfaces before large conformational changes can occur. This finding may be very important for biomaterial surface design as it suggests that surface chemistry can potentially be used to directly control the orientation of adsorbing proteins in a manner that either presents or hides specific bioactive sites contained within a protein's structure, thereby providing a mechanism to control cellular responses to the adsorbed protein layer. PMID- 15667198 TI - Molecular dynamics simulations of polyelectrolyte multilayering on a charged particle. AB - Molecular dynamics simulations of polyelectrolyte multilayering on a charged spherical particle revealed that the sequential adsorption of oppositely charged flexible polyelectrolytes proceeds with surface charge reversal and highlighted electrostatic interactions as the major driving force of layer deposition. Far from being completely immobilized, multilayers feature a constant surge of chain intermixing during the deposition process, consistent with experimental observations of extensive interlayer mixing in these films. The formation of multilayers as well as the extent of layer intermixing depends on the degree of polymerization of the polyelectrolyte chains and the fraction of charge on its backbone. The presence of ionic pairs between oppositely charged macromolecules forming layers seems to play an important role in stabilizing the multilayer film. PMID- 15667199 TI - Tradeoff between mixing and transport for electroosmotic flow in heterogeneous microchannels with nonuniform surface potentials. AB - Electroosmotic flow (EOF) is a phenomenon associated with the movement of an aqueous solution induced by the application of an electric field in microchannels. The characteristics of EOF depend on the nature of the surface potential, i.e., whether it is uniform or nonuniform. In this paper, a lattice Boltzmann model (LBM) combined with the Poisson-Boltzmann equation is used to simulate flow field in a rectangular microchannel with nonuniform (step change) surface potentials. The simulation results indicate that local circulations can occur near a heterogeneous region with nonuniform surface potentials, in agreement with those by other authors. Largest circulations, which imply a highest mixing efficiency due to convection and short-range diffusion, were found when the average surface potential is zero, regardless of whether the distribution of the heterogeneous patches is symmetric or asymmetric. In this work, we have illustrated that there is a trade-off between the mixing and liquid transport in EOF microfluidics. One should not simply focus on mixing and neglect liquid transport, as performed in the literature. Excellent mixing could lead to a poor transport of electroosmotic flow in microchannels. PMID- 15667200 TI - Surfactant-assisted large-scale preparation of crystalline CeO2 nanorods. PMID- 15667201 TI - Biomimetic nanostructured materials: inherent reversible stabilization of polypeptide microcapsules. PMID- 15667202 TI - Intramolecular occlusion of the diacylglycerol-binding site in the C1 domain of munc13-1. AB - Protein kinase C (PKC) isozymes and other receptors of diacylglycerol (DAG) bind to this widespread second messenger through their C(1) domains. These alternative DAG receptors include munc13-1, a large neuronal protein that is crucial for DAG dependent augmentation of neurotransmitter release. Whereas the structures of several PKC C(1) domains have been determined and have been shown to require little conformational changes for ligand binding, it is unclear whether the C(1) domains from other DAG receptors contain specific structural features with key functional significance. To gain insight into this question, we have determined the three-dimensional structure in solution of the munc13-1 C(1) domain using NMR spectroscopy. The overall structure includes two beta-sheets, a short C-terminal alpha-helix, and two Zn(2+)-binding sites, resembling the structures of PKC C(1) domains. However, the munc13-1 C(1) domain exhibits striking structural differences with the PKC C(1) domains in the ligand-binding site. These differences result in occlusion of the binding site of the munc13-1 C(1) domain by a conserved tryptophan side chain that in PKCs adopts a completely different orientation. As a consequence, the munc13-1 C(1) domain requires a considerable conformational change for ligand binding. This structural distinction is expected to decrease the DAG affinity of munc13-1 compared to that of PKCs, and is likely to be critical for munc13-1 function. On the basis of these results, we propose that augmentation of neurotransmitter release may be activated at higher DAG levels than PKCs as a potential mechanism for uncoupling augmentation of release from the multitude of other signaling processes mediated by DAG. PMID- 15667203 TI - Structural and kinetic characterization of active-site histidine as a proton shuttle in catalysis by human carbonic anhydrase II. AB - In the catalysis of the hydration of carbon dioxide and dehydration of bicarbonate by human carbonic anhydrase II (HCA II), a histidine residue (His64) shuttles protons between the zinc-bound solvent molecule and the bulk solution. To evaluate the effect of the position of the shuttle histidine and pH on proton shuttling, we have examined the catalysis and crystal structures of wild-type HCA II and two double mutants: H64A/N62H and H64A/N67H HCA II. His62 and His67 both have their side chains extending into the active-site cavity with distances from the zinc approximately equivalent to that of His64. Crystal structures were determined at pH 5.1-10.0, and the catalysis of the exchange of (18)O between CO(2) and water was assessed by mass spectrometry. Efficient proton shuttle exceeding a rate of 10(5) s(-)(1) was observed for histidine at positions 64 and 67; in contrast, relatively inefficient proton transfer at a rate near 10(3) s( )(1) was observed for His62. The observation, in the crystal structures, of a completed hydrogen-bonded water chain between the histidine shuttle residue and the zinc-bound solvent does not appear to be required for efficient proton transfer. The data suggest that the number of intervening water molecules between the donor and acceptor supporting efficient proton transfer in HCA II is important, and furthermore suggest that a water bridge consisting of two intervening water molecules is consistent with efficient proton transfer. PMID- 15667204 TI - Kinetic analysis of the zinc-dependent deacetylase in the lipid A biosynthetic pathway. AB - The first committed step of lipid A biosynthesis in Gram-negative bacteria is catalyzed by the zinc-dependent hydrolase LpxC that removes an acetate from the nitrogen at the 2' '-position of UDP-3-O-acyl-N-acetylglucosamine. Recent structural characterization by both NMR and X-ray crystallography provides many important details about the active site environment of LpxC from Aquifex aeolicus, a heat-stable orthologue that displays 32% sequence identity to LpxC from Escherichia coli. The detailed reaction mechanism and specific roles of active site residues for LpxC from A. aeolicus are further analyzed here. The pH dependencies of k(cat)/K(M) and k(cat) for the deacetylation of the substrate UDP 3-O-[(R)-3-hydroxymyristoyl]-GlcNAc are both bell-shaped. The ascending acidic limb (pK(1)) was fitted to 6.1 +/- 0.2 for k(cat) and 5.7 +/- 0.2 for k(cat)/K(M). The descending basic limb (pK(2)) was fitted to 8.0 +/- 0.2 for k(cat) and 8.4 +/- 0.2 for k(cat)/K(M). The pH dependence of the E73A mutant exhibits loss of the acidic limb, and the mutant retains only 0.15% activity versus the wild type. The pH dependencies of the other active site mutants H253A, K227A, H253A/K227A, and D234N remain bell-shaped, although their significantly lower activities (0.25%, 0.05%, 0.007%, and 0.57%, respectively) suggest that they contribute significantly to catalysis. Our cumulative data support a mechanism for LpxC wherein Glu73 serves as the general base for deprotonation and activation of the zinc-bound water. PMID- 15667205 TI - Refined solution structure of the LpxC-TU-514 complex and pKa analysis of an active site histidine: insights into the mechanism and inhibitor design. AB - Lipopolysaccharide, the major constituent of the outer monolayer of the outer membrane of Gram-negative bacteria, is anchored into the membrane through the hydrophobic moiety lipid A, a hexaacylated disaccharide. The zinc-dependent metalloamidase UDP-3-O-acyl-N-acetylglucosamine deacetylase (LpxC) catalyzes the second and committed step in the biosynthesis of lipid A. LpxC shows no homology to mammalian metalloamidases and is essential for cell viability, making it an important target for the development of novel antibacterial compounds. Recent NMR and X-ray studies of the LpxC from Aquifex aeolicus have provided the first structural information about this family of proteins. Insight into the catalytic mechanism and the design of effective inhibitors could be facilitated by more detailed structural and biochemical studies that define substrate-protein interactions and the roles of specific residues in the active site. Here, we report the synthesis of the (13)C-labeled substrate-analogue inhibitor TU-514, and the subsequent refinement of the solution structure of the A. aeolicus LpxC TU-514 complex using residual dipolar couplings. We also reevaluate the catalytic role of an active site histidine, H253, on the basis of both its pK(a) as determined by NMR titration and pH-dependent kinetic analyses. These results provide a structural basis for the design of more potent LpxC inhibitors than those that are currently available. PMID- 15667206 TI - Evidence for two different mechanisms triggering the change in quaternary structure of the allosteric enzyme, glucosamine-6-phosphate deaminase. AB - The generation and propagation of conformational changes associated with ligand binding in the allosteric enzyme glucosamine-6-phosphate deaminase (GlcN6P deaminase, EC 3.5.99.6) from Escherichia coli were analyzed by fluorescence measurements. Single-tryptophan mutant forms of the enzyme were constructed on the basis of previous structural and functional evidence and used as structural change probes. The reporter residues were placed in the active-site lid (position 174) and in the allosteric site (254 and 234); in addition, signals from the natural Trp residues (15 and 224) were also studied as structural probes. The structural changes produced by the occupation of either the allosteric or the active site by site-specific ligands were monitored through changes in the spectral center of mass (SCM) of their steady-state emission fluorescence spectra. Binding of the allosteric activator produces only minimal signals in titration experiments. In contrast, measurable spectral signals were found when the active site was occupied by a dead-end inhibitor. The results reveal that the two binary complexes, enzyme-activator (R(A)) and enzyme-inhibitor (R(S)) complexes, have structural differences and that they also differ from the ternary complex (R(AS)). The mobility of the active-site lid motif is shown to be independent of the allosteric transition. The active-site ligand induces cooperative SCM changes even in the enzyme-activator complex, indicating that the propagation pathway of the conformational relaxation triggered from the active site is different from that involved in the heterotropic activation. Analysis of the complete set of mutants shows that the occupation of the active site generates structural perturbations, which are propagated to the whole of the monomer and extend to the other subunits. The accumulative effect of these propagated changes should be responsible for the change in the sign of the DeltaG degrees ' of the T to R transition associated with the progression of the active site occupation, resulting in the predominance of the R over the T forms in the population of deaminase hexamers. PMID- 15667207 TI - Crystal structures of pyruvate phosphate dikinase from maize revealed an alternative conformation in the swiveling-domain motion. AB - Pyruvate phosphate dikinase (PPDK) reversibly catalyzes the conversion of ATP, phosphate, and pyruvate into AMP, pyrophosphate, and phosphoenolpyruvate (PEP), respectively. Since the nucleotide binding site (in the N-terminal domain) and the pyruvate/PEP binding site (in the C-terminal domain) are separated by approximately 45 A, it has been proposed that an intermediary domain, called the central domain, swivels between these remote domains to transfer the phosphate. However, no direct structural evidence for the swiveling central domain has been found. In this study, the crystal structures of maize PPDK with and without PEP have been determined at 2.3 A resolution. These structures revealed that the central domain is located near the pyruvate/PEP binding C-terminal domain, in contrast to the PPDK from Clostridium symbiosum, wherein the central domain is located near the nucleotide-binding N-terminal domain. Structural comparisons between the maize and C. symbiosum PPDKs demonstrated that the swiveling motion of the central domain consists of a rotation of at least 92 degrees and a translation of 0.5 A. By comparing the maize PPDK structures with and without PEP, we have elucidated the mode of binding of PEP to the C-terminal domain and the induced conformational changes in the central domain. PMID- 15667208 TI - Disulfide bond mutagenesis and the structure and function of the head-to-tail macrocyclic trypsin inhibitor SFTI-1. AB - SFTI-1 is a novel 14 amino acid peptide comprised of a circular backbone constrained by three proline residues, a hydrogen-bond network, and a single disulfide bond. It is the smallest and most potent known Bowman-Birk trypsin inhibitor and the only one with a cyclic peptidic backbone. The solution structure of [ABA(3,11)]SFTI-1, a disulfide-deficient analogue of SFTI-1, has been determined by (1)H NMR spectroscopy. The lowest energy structures of native SFTI-1 and [ABA(3,11)]SFTI-1 are similar and superimpose with a root-mean-square deviation over the backbone and heavy atoms of 0.26 +/- 0.09 and 1.10 +/- 0.22 A, respectively. The disulfide bridge in SFTI-1 was found to be a minor determinant for the overall structure, but its removal resulted in a slightly weakened hydrogen-bonding network. To further investigate the role of the disulfide bridge, NMR chemical shifts for the backbone H(alpha) protons of two disulfide deficient linear analogues of SFTI-1, [ABA(3,11)]SFTI-1[6,5] and [ABA(3,11)]SFTI 1[1,14] were measured. These correspond to analogues of the cleavage product of SFTI-1 and a putative biosynthetic precursor, respectively. In contrast with the cyclic peptide, it was found that the disulfide bridge is essential for maintaining the structure of these open-chain analogues. Overall, the hydrogen bond network appears to be a crucial determinant of the structure of SFTI-1 analogues. PMID- 15667209 TI - Role of water in aging of human butyrylcholinesterase inhibited by echothiophate: the crystal structure suggests two alternative mechanisms of aging. AB - Organophosphorus poisons (OP) bind covalently to the active-site serine of cholinesterases. The inhibited enzyme can usually be reactivated with powerful nucleophiles such as oximes. However, the covalently bound OP can undergo a suicide reaction (termed aging) yielding nonreactivatable enzyme. In human butyrylcholinesterase (hBChE), aging involves the residues His438 and Glu197 that are proximal to the active-site serine (Ser198). The mechanism of aging is known in detail for the nerve gases soman, sarin, and tabun as well as the pesticide metabolite isomalathion. Aging of soman- and sarin-inhibited acetylcholinesterase occurs by C-O bond cleavage, whereas that of tabun- and isomalathion-inhibited acetylcholinesterase occurs by P-N and P-S bond cleavage, respectively. In this work, the crystal structures of hBChE inhibited by the ophthalmic reagents echothiophate (nonaged and aged) and diisopropylfluorophosphate (aged) were solved and refined to 2.1, 2.25, and 2.2 A resolution, respectively. No appreciable shift in the position of the catalytic triad histidine was observed between the aged and nonaged conjugates of hBChE. This absence of shift contrasts with the aged and nonaged crystal structures of Torpedo californica acetylcholinesterase inhibited by the nerve agent VX. The nonaged hBChE structure shows one water molecule interacting with Glu197 and the catalytic triad histidine (His438). Interestingly, this water molecule is ideally positioned to promote aging by two mechanisms: breaking either a C-O bond or a P-O bond. Pesticides and certain stereoisomers of nerve agents are expected to undergo aging by breaking the P-O bond. PMID- 15667210 TI - Limited proteolysis and biophysical characterization of the lipovitellin homology region in apolipoprotein B. AB - Apolipoprotein B (apoB) is the essential nonexchangeable protein in chylomicrons and very low-density lipoprotein-derived lipoprotein particles, including low density lipoprotein (LDL). ApoB has been a key target for cardiovascular research because of its essential role in the assembly, secretion, delivery, and receptor binding of LDL. The three-dimensional structure of apoB has not been determined. However, the N-terminal region of apoB is homologous to the lipid storage protein lipovitellin, which allows the modeling of this region based on the X-ray structure of lipovitellin. The model of the N-terminal 17% of apoB (B17) suggests that, like lipovitellin, B17 consists of an N-terminal beta-barrel domain, a helical domain, and a beta-sheet domain (C-sheet). Here we test the validity of this model by limited proteolysis of B17 and the characterization of individual domains expressed in Escherichia coli and insect cell systems that are consistent with the model and proteolysis data. Circular dichroism studies of the individual domains indicate that they are folded and their secondary structures are in agreement with the model. We find that the helical domain and C-sheet of apoB interact with each other in vitro, suggesting a strong interaction between these two domains, even without a covalent peptide bond linkage. Our data suggest that the three lipovitellin-like domains exist in B17. Furthermore, the domains fold independently with secondary structures and stabilities like those of intact B17. PMID- 15667211 TI - Crystallographic and thermodynamic analysis of the binding of S-octylglutathione to the Tyr 7 to Phe mutant of glutathione S-transferase from Schistosoma japonicum. AB - Glutathione S-transferases are a family of multifunctional enzymes involved in the metabolism of drugs and xenobiotics. Two tyrosine residues, Tyr 7 and Tyr 111, in the active site of the enzyme play an important role in the binding and catalysis of substrate ligands. The crystal structures of Schistosoma japonicum glutathione S-transferase tyrosine 7 to phenylalanine mutant [SjGST(Y7F)] in complex with the substrate glutathione (GSH) and the competitive inhibitor S octylglutathione (S-octyl-GSH) have been obtained. These new structural data combined with fluorescence spectroscopy and thermodynamic data, obtained by means of isothermal titration calorimetry, allow for detailed characterization of the ligand-binding process. The binding of S-octyl-GSH to SjGST(Y7F) is enthalpically and entropically driven at temperatures below 30 degrees C. The stoichiometry of the binding is one molecule of S-octyl-GSH per mutant dimer, whereas shorter alkyl derivatives bind with a stoichiometry of two molecules per mutant dimer. The SjGST(Y7F).GSH structure showed no major structural differences compared to the wild-type enzyme. In contrast, the structure of SjGST(Y7F).S-octyl-GSH showed asymmetric binding of S-octyl-GSH. This lack of symmetry is reflected in the lower symmetry space group of the SjGST(Y7F).S-octyl-GSH crystals (P6(3)) compared to that of the SjGST(Y7F).GSH crystals (P6(3)22). Moreover, the binding of S-octyl-GSH to the A subunit is accompanied by conformational changes that may be responsible for the lack of binding to the B subunit. PMID- 15667212 TI - Conformational changes in the alpha-subunit coupled to binding of the beta 2 subunit of tryptophan synthase from Escherichia coli: crystal structure of the tryptophan synthase alpha-subunit alone. AB - When the tryptophan synthase alpha- and beta(2)-subunits combine to form the alpha(2)beta(2)-complex, the enzymatic activity of each subunit is stimulated by 1-2 orders of magnitude. To elucidate the structural basis of this mutual activation, it is necessary to determine the structures of the alpha- and beta subunits alone and together with the alpha(2)beta(2)-complex. The crystal structures of the tryptophan synthase alpha(2)beta(2)-complex from Salmonella typhimurium (Stalpha(2)beta(2)-complex) have already been reported. However, the structures of the subunit alone from mesophiles have not yet been determined. The structure of the tryptophan synthase alpha-subunit alone from Escherichia coli (Ecalpha-subunit) was determined by an X-ray crystallographic analysis at 2.3 A, which is the first report on the subunits alone from the mesophiles. The biggest difference between the structures of the Ecalpha-subunit alone and the alpha subunit in the Stalpha(2)beta(2)-complex (Stalpha-subunit) was as follows. Helix 2' in the Stalpha-subunit, including an active site residue (Asp60), was changed to a flexible loop in the Ecalpha-subunit alone. The conversion of the helix to a loop resulted in the collapse of the correct active site conformation. This region is also an important part for the mutual activation in the Stalpha(2)beta(2)-complex and interaction with the beta-subunit. These results suggest that the formation of helix 2'that is essential for the stimulation of the enzymatic activity of the alpha-subunit is constructed by the induced-fit mode involved in conformational changes upon interaction between the alpha- and beta-subunits. This also confirms the prediction of the conformational changes based on the thermodynamic analysis for the association between the alpha- and beta-subunits. PMID- 15667214 TI - Effects of metal ions in the CuB center on the redox properties of heme in heme copper oxidases: spectroelectrochemical studies of an engineered heme-copper center in myoglobin. AB - The electrochemical properties of an engineered heme-copper center in myoglobin have been investigated by UV-visible spectroelectrochemistry. In the cyanide bridged, spin-coupled heme-copper center in an engineered myoglobin, the presence of Zn(II) in the Cu(B) center raises the heme reduction potential from -85 to 49 mV vs NHE. However, in the cyanide-free, spin-decoupled derivative of the same protein, the presence of Zn(II) in the Cu(B) center exerts little influence on the heme reduction potentials (77 and 80 mV vs NHE, respectively, in the absence and in the presence of Zn(II)). Similar trends have also been observed when copper ion is present in the Cu(B) center, although on a smaller scale, due to reduction of Cu(II) to Cu(I) prior to heme reduction. These results show that the presence of a metal ion in the designed Cu(B) center has a significant effect on the redox potential of heme Fe only when the two metal centers are coupled through a bridging ligand between the two metal centers, indicating that spin coupling plays an important role in redox potential regulation. In addition, the presence of a single positively charged Cu(I) center in the Cu(B) center resulted in a much lower increase (16 mV) in heme reduction potential than that of two positively charged Zn(II) (118 mV). Therefore, the heme reduction potential must be lowered after the first electron transfer to reduce heme Fe(3+)-Cu(B)(2+) to Fe(3+)-Cu(B)(+). To raise the heme reduction potential to make the second electron transfer (i.e., reduction of Fe(3+)-Cu(B)(+) to Fe(2+)-Cu(B)(+)) to be favorable, most likely a proton or decoupling of the heme-copper center is needed in the heme-copper site. These findings provide a strong argument for a thermodynamic driving force basis for redox-regulated proton transfer in heme copper oxidases. PMID- 15667213 TI - Exploring the binding site structure of the PPAR gamma ligand-binding domain by computational solvent mapping. AB - Solvent mapping moves molecular probes, small organic molecules containing various functional groups, around the protein surface, finds favorable positions, clusters the conformations, and ranks the clusters based on the average free energy. Using at least six different solvents as probes, the probes cluster in major pockets of the functional site, providing detailed and reliable information on the amino acid residues that are important for ligand binding. Solvent mapping was applied to 12 structures of the peroxisome proliferator activated receptor gamma (PPARgamma) ligand-binding domain (LBD), including 2 structures without a ligand, 2 structures with a partial agonist, and 8 structures with a PPAR agonist bound. The analysis revealed 10 binding "hot spots", 4 in the ligand-binding pocket, 2 in the coactivator-binding region, 1 in the dimerization domain, 2 around the ligand entrance site, and 1 minor site without a known function. Mapping is a major source of information on the role and cooperativity of these sites. It shows that large portions of the ligand-binding site are already formed in the PPARgamma apostructure, but an important pocket near the AF-2 transactivation domain becomes accessible only in structures that are cocrystallized with strong agonists. Conformational changes were seen in several other sites, including one involved in the stabilization of the LBD and two others at the region of the coactivator binding. The number of probe clusters retained by these sites depends on the properties of the bound agonist, providing information on the origin of correlations between ligand and coactivator binding. PMID- 15667215 TI - Light-induced structural changes of apoprotein and chromophore in the sensor of blue light using FAD (BLUF) domain of AppA for a signaling state. AB - AppA is a new class blue-light receptor controlling photosynthesis gene expression in the purple bacterium Rhodobacter sphaeroides and retains a characteristic flavin adenine dinucleotide (FAD)-binding domain named the "sensor of blue light using FAD" (BLUF). AppA functions as an antirepressor controlling transcription of photosynthesis genes through the direct association with a transcriptional repressor PpsR in a blue-light-dependent manner [Masuda and Bauer (2002) Cell 110, 613-623]. Illumination of AppA induces a red shift in the UV visible absorption of FAD, which results in a signaling state of AppA. Light induced Fourier transform infrared (FTIR) difference spectrum of the AppA BLUF domain showed relatively simple features, which were mainly composed of two sets of derivative-shaped sharp bands at 1709(-)/1695(+) and 1632(+)/1619(-) cm(-)(1). We have developed an in vitro reconstitution method, by which a fully functional BLUF domain was reconstituted from free FAD and an apoprotein for the BLUF domain of AppA. An AppA BLUF domain that consisted of an apoprotein isotopically labeled with (13)C and unlabeled FAD was constituted using this method, and hydrated and deuterated samples were applied to FTIR spectroscopic analyses. When the spectra for the reconstituted domain were compared with those for uniformly (15)N- and (13)C-labeled or deuterated domains as well as for the unlabeled domain, the IR bands responsible for the light-induced changes in the FAD chromophore and apoprotein were identified. Unexpectedly, the light-induced spectrum of the unlabeled BLUF domain of AppA was predominantly composed of multiple apoprotein bands, while a C(4)=O stretching of an isoalloxazine ring was the only band exclusively assigned to FAD. The results showed that relatively large structural changes occur in the protein backbone of the BLUF domain of AppA upon illumination. These changes were discussed in relation to the mechanistic role of the BLUF domain in the process of blue-light perception by AppA. PMID- 15667216 TI - Dynamics of the fragment of thrombomodulin containing the fourth and fifth epidermal growth factor-like domains correlate with function. AB - Thrombomodulin (TM) forms a 1:1 complex with thrombin. Whereas thrombin alone cleaves fibrinogen to make the fibrin clot, the thrombin-TM complex cleaves protein C to initiate the anticoagulant pathway. The fourth and fifth EGF-like domains of TM together form the minimal fragment with anticoagulant cofactor activity. A short linker connects the fourth and fifth EGF-like domains of TM, and Met 388 in the middle of the linker interacts with both domains. Several different structures of TMEGF45 variants are now available, and these show that mutation of Met 388 alters the structure of the fifth domain, as well as the connectivity of the two domains. To probe this phenomenon more thoroughly, NMR backbone dynamics experiments have been carried out on the individual fourth and fifth domains as well as on the wild type, the Met 388 Leu mutant, and the variant in which Met 388 is oxidized. The results presented here show that changes at Met 388 cause significant changes in backbone dynamics in both the fourth and fifth EGF-like domains of TM. Backbone dynamics within the small loop of the fourth domain Tyr 358 correlate with anticoagulant cofactor activity. Backbone dynamics of the thrombin-binding residues Tyr 413 and Ile 414 are inversely correlated with thrombin binding. The preordering of the backbone of Tyr 413 and Ile 414 only occurs in the two-domain fragments, revealing a role for the fourth domain in thrombin binding as well as in anticoagulant cofactor activity. PMID- 15667217 TI - Structural evidence of a fourth Gla residue in fish osteocalcin: biological implications. AB - Osteocalcin is a small (45 amino acids) secreted protein found to accumulate in bone and dentin of many organisms by interacting with calcium and hydroxyapatite, through the presence of three gamma-carboxylated residues. In this work, we describe the first X-ray crystal structure for a nonmammalian osteocalcin, obtained at 1.4 A resolution, purified from the marine teleost fish Argyrosomus regius. The three-dimensional fit between the A. regius structure and that of the only other known X-ray structure, the porcine osteocalcin, revealed a superposition of the Calpha atoms of their metal chelating residues, Gla and Asp, showing that their spatial distribution is consistent with the interatomic distances of calcium cations in the hydroxyapatite crystals. In both structures, the protein forms a tight globular arrangement of their three alpha-helices while the remaining residues, at N- and C-terminal regions, have essentially no secondary structure characteristics. This study revealed the presence of a fourth gamma-carboxylation at Glu(25), not previously detected in the structure of the porcine osteocalcin or in any other of the sequentially characterized mammalian osteocalcins (human, cow, and rat). A confirmation of the fourth Gla residue in A. regius osteocalcin was achieved via LC-MS analysis. These four doubly charged residues are, together with Asp(24), concentrated in a common surface region located on the same side of the molecule. This further suggests that the known high affinity of osteocalcin for bone mineral may be derived from the clustering of calcium binding sites on this surface of the molecules. PMID- 15667218 TI - Apparent Debye-Huckel electrostatic effects in the folding of a simple, single domain protein. AB - We have monitored the effects of salts and denaturants on the folding of the simple, two-state protein FynSH3. As predicted by Debye-Huckel limiting law, both the stability and (log) folding rate of FynSH3 increase nearly perfectly linearly (r(2)> 0.99) with the square root of ionic strength upon increasing concentrations of the relatively nonchaotropic salt sodium chloride. The stability of FynSH3 is also linear in square root ionic strength when the relatively nonchaotropic salts sodium bromide, potassium bromide, and potassium chloride are employed. Comparison of the kinetic and equilibrium effects of sodium chloride suggests that the electrostatic interactions formed in the folding transition state are approximately 50% as destabilizing as those formed in the native state, presumably reflecting the more compact nature of the latter. In contrast, the relationship between concentration and folding kinetics is more complex when the highly chaotropic salt guanidine hydrochloride (GuHCl) is employed. At moderate to high GuHCl concentrations the net effect of the linear, presumably chaotrope-induced deceleration and the presumed, square root-dependent ionic strength-induced acceleration is well approximated as linear, thus accounting for the observation of "chevron behavior" (log folding rate linear in denaturant concentration) typically reported for the folding of single domain proteins. At very low GuHCl concentrations, however, significant kinetic rollover is observed. This rollover is reasonably well fitted as a sum of a linear, presumably chaotropic effect and a square root-dependent, presumably electrostatic effect. These results thus not only provide insight into the nature of the folding transition state but also suggest that caution is in order when extrapolating GuHCl-based chevrons to estimate folding rates in the absence of denaturant and in interpreting deviations from chevron linearity as evidence for non-two-state kinetics. PMID- 15667219 TI - Kinetic mechanism of rat polymerase beta-dsDNA interactions. Fluorescence stopped flow analysis of the cooperative ligand binding to a two-site one-dimensional lattice. AB - Kinetics of cooperative binding of rat polymerase beta to a double-stranded DNA has been studied using the fluorescence stopped-flow techniques. The data have been analyzed by an approach developed to examine complete kinetics of cooperative large ligand binding to a one-dimensional lattice. The method is based on using the smallest possible system that preserves key ingredients of cooperative binding; i.e., at saturation, the lattice can accept only two ligand molecules. It allows the identification of collective amplitudes as well as amplitudes describing particular normal modes of the reaction. The mechanism of the intrinsic binding of pol beta to the dsDNA is different from the analogous mechanism for the ssDNA. The difference originates from different enzyme orientations in the corresponding complexes. Intrinsic binding to the dsDNA includes only two sequential steps: a very fast bimolecular association followed by an energetically favorable conformational transition of the complex. The transition following the bimolecular step does not facilitate the engagement of the enzyme in cooperative interactions. Its role seems to be reinforcing the affinity of the bimolecular step. Salt and magnesium cations affect both the bimolecular step and the conformational transition. As a result, the bimolecular step is less sensitive to the increased salt concentration, allowing the enzyme to preserve its initial dsDNA affinity. The changing character of cooperative interactions between bound enzyme molecules as a function of NaCl concentration and MgCl(2) concentration does not affect the binding mechanism. The engagement in cooperative interactions is approximately 3-4 orders of magnitude slower than the conformational transition of the DNA-bound polymerase. The importance of the obtained results for the pol beta activities is discussed. PMID- 15667220 TI - Tryptophan residues flanking the second transmembrane helix (TM2) set the signaling state of the Tar chemoreceptor. AB - The chemoreceptors of Escherichia coli are homodimeric membrane proteins that cluster in patches near the cell poles. They convert environmental stimuli into intracellular signals that control flagellar rotation. The functional domains of a receptor are physically separated by the cell membrane. Chemoeffectors bind to the extracellular (periplasmic) domain, and the cytoplasmic domain mediates signaling and adaptation. These two domains communicate through the second transmembrane helix (TM2) that connects them. In the high-abundance receptors Tar and Tsr, TM2 is flanked by tryptophan residues, which should localize preferentially to the interfacial zone between the polar and hydrophobic layers of the phospholipid bilayer. To investigate the functional significance of the Trp residues that flank TM2 of Tar, we used site-directed mutagenesis to generate the W192A and W209A substitutions. The W192A protein retains full activity in vivo and in vitro, but it increases the K(i) for aspartate in the in vitro assay 3-fold. The W209A replacement eliminates receptor-mediated stimulation of CheA in vitro, and it leads to an increased level of adaptive methylation in vivo. This phenotype in some respects mimics the changes seen upon binding aspartate. Since the W209A substitution may cause the C-terminus of TM2 to protrude farther into the cytoplasm, these results reinforce the hypothesis that aspartate binding causes a similar displacement. Moving Trp to each position from residue 206 to residue 212 generated a wide variety of Tar signaling states that are generally consistent with the predictions of the piston model of transmembrane signaling. None of these receptors was completely locked in one signaling mode, although most showed pronounced signaling biases. Our findings suggest that the Trp residues flanking TM2, especially Trp-209, are important in setting the baseline activity and ligand sensitivity of the Tar receptor. We also conclude that the Tyr-210 residue plays at least an auxiliary role in this control. PMID- 15667221 TI - Identification of residues that contribute to receptor activation through the analysis of compensatory mutations in the G protein-coupled alpha-factor receptor. AB - The alpha-factor receptor (Ste2p) stimulates mating of the yeast Saccharomyces cerevisiae. Ste2p belongs to the large family of G protein-coupled receptors that are characterized by seven transmembrane alpha-helices. Receptor activation is thought to involve changes in the packing of the transmembrane helix bundle. To identify residues that contribute to Ste2p activation, second-site suppressor mutations were isolated that restored function to defective receptors carrying either an F204S or Y266C substitution which affect residues at the extracellular ends of transmembrane domains 5 and 6, respectively. Thirty-five different suppressor mutations were identified. On their own, these mutations caused a range of phenotypes, including hypersensitivity, constitutive activity, altered ligand binding, and loss of function. The majority of the mutations affected residues in the transmembrane segments that are predicted to face the helix bundle. Many of the suppressor mutations caused constitutive receptor activity, suggesting they improved receptor function by partially restoring the balance between the active and inactive states. Analysis of mutations in transmembrane domain 7 implicated residues Ala281 and Thr282 in receptor activation. The A281T and T282A mutants were supersensitive to S. cerevisiae alpha-factor, but were defective in responding to a variant of alpha-factor produced by another species, Saccharomyces kluyveri. The A281T mutant also displayed 8.7-fold enhanced basal signaling. Interestingly, Ala281 and Thr282 are situated in approximately the same position as Lys296 in rhodopsin, which is covalently linked to retinal. These results suggest that transmembrane domain 7 plays a role in receptor activation in a wide range of G protein-coupled receptors from yeast to humans. PMID- 15667223 TI - Evidence for an EPR-detectable semiquinone intermediate stabilized in the membrane-bound subunit NarI of nitrate reductase A (NarGHI) from Escherichia coli. AB - Nitrate reductase A (NRA, NarGHI) is expressed in Escherichia coli by growing the bacterium in anaerobic conditions in the presence of nitrate. This enzyme reduces nitrate to nitrite and uses menaquinol (or ubiquinol) as the electron donor. The location of quinones in the enzyme, their number, and their role in the electron transfer mechanism are still controversial. In this work, we have investigated the spectroscopic and thermodynamic properties of a semiquinone (SQ) in membrane samples of overexpressed E. coli nitrate reductase poised in appropriate redox conditions. This semiquinone is highly stabilized with respect to free semiquinone. The g-values determined from the numerical simulation of its Q-band (35 GHz) EPR spectrum are equal to 2.0061, 2.0051, 2.0023. The midpoint potential of the Q/QH(2) couple is about -100 mV, and the SQ stability constant is about 100 at pH 7.5. The semiquinone EPR signal disappears completely upon addition of the quinol binding site inhibitor 2-n-nonyl-4-hydroxyquinoline N-oxide (NQNO). A semiquinone radical could also be stabilized in preparations where only the NarI membrane subunit is overexpressed in the absence of the NarGH catalytic dimer. Its thermodynamic and spectroscopic properties show only slight variations with those of the wild-type enzyme. The X-band continuous wave (cw) electron nuclear double resonance (ENDOR) spectra of the radicals display similar proton hyperfine coupling patterns in NarGHI and in NarI, showing that they arise from the same semiquinone species bound to a single site located in the NarI membrane subunit. These results are discussed with regard to the location and the potential function of quinones in the enzyme. PMID- 15667222 TI - Critical balance of electrostatic and hydrophobic interactions is required for beta 2-microglobulin amyloid fibril growth and stability. AB - Investigation of factors that modulate amyloid formation of proteins is important to understand and mitigate amyloid-related diseases. To understand the role of electrostatic interactions and the effect of ionic cosolutes, especially anions, on amyloid formation, we have investigated the effect of salts such as NaCl, NaI, NaClO(4), and Na(2)SO(4) on the amyloid fibril growth of beta(2)-microglobulin, the protein involved in dialysis-related amyloidosis. Under acidic conditions, these salts exhibit characteristic optimal concentrations where the fibril growth is favored. The presence of salts leads to an increase in hydrophobicity of the protein as reported by 8-anilinonaphthalene-1-sulfonic acid, indicating that the anion interaction leads to the necessary electrostatic and hydrophobic balance critical for amyloid formation. However, high concentrations of salts tilt the balance to high hydrophobicity, leading to partitioning of the protein to amorphous aggregates. Such amorphous aggregates are not competent for fibril growth. The order of anions based on the lowest concentration at which fibril formation is favored is SO(4)(2)(-) > ClO(4)(-) > I(-) > Cl(-), consistent with the order of their electroselectivity series, suggesting that preferential anion binding, rather than general ionic strength effect, plays an important role in the amyloid fibril growth. Anion binding is also found to stabilize the amyloid fibrils under acidic condition. Interestingly, sulfate promotes amyloid growth of beta(2)-microglobulin at pH between 5 and 6, closer to its isoelectric point. Considering the earlier studies on the role of glycosaminoglycans and proteoglycans (i.e., sulfated polyanions) on amyloid formation, our study suggests that preferential interaction of sulfate ions with amyloidogenic proteins may have biological significance. PMID- 15667224 TI - Effect of substrate on the diiron(III) site in stearoyl acyl carrier protein delta 9-desaturase as disclosed by cryoreduction electron paramagnetic resonance/electron nuclear double resonance spectroscopy. AB - The diiron center in stearoyl-acyl carrier protein (ACP) desaturase (DS) from castor plant Ricinus communis catalyzes the dioxygen- and NADPH-dependent introduction of a cis double bond between C9 and C10 of stearoyl-ACP. Radiolytic reduction of diferric DS at 77 K produces an electron paramagnetic resonance (EPR)-detectable mixed-valence center (or [DS(ox)](mv)) that is trapped in the conformation of the diferric precursor and thus provides a sensitive EPR/electron nuclear double resonance (ENDOR) probe of the structure of the diamagnetic diiron(III) state. The cryoreduced DS shows two distinct EPR signals, suggesting the presence of two diiron(III) states: the mu-oxo (major)- and mu-hydroxo (minor)-bridged diiron centers. ENDOR studies show that in the dominant oxo bridged diferric state each iron(III) coordinates a histidine and a water along with other ligands. Samples containing stoichiometric amounts of stearoyl-ACP show pronounced changes in the EPR and (1)H ENDOR spectra of cryoreduced DS. EPR spectra of the cryoreduced DS-substrate complex reveal two distinct substates of the parent. EPR and ENDOR studies show that both major conformers of the diferric cluster have a mu-oxo bridge. ENDOR shows that the major conformer has a histidine and a water bound to both Fe ions. In the minor conformer, one of the irons has lost the terminal water ligand. The structure of the trapped [DS(ox)](mv) state relaxes upon annealing to 170 K: the mu-oxo bridge in the major cryoreduced DS species protonates on annealing to 170 K; this does not occur for the major DS-substrate complex, even upon annealing to 230 K. The relaxed Fe(II)Fe(III) center in cryoreduced DS and DS-substrate show much less intense and resolved (14)N ENDOR spectra than those of the structurally similar cryoreduced diiron center in ribonucleotide reductase (RNR) protein R2. This difference may reflect some differences in His-Fe bonds. The alterations in the diferric site of DS induced by substrate are suggested to be mediated by conformational changes in the polypeptide chain produced by substrate binding. These structural alterations may provide DS with an additional mechanism for tuning the redox potential of the diferric site. The mixed-valence states of DS are unstable at temperatures above 230 K. PMID- 15667225 TI - Oligomerization and phase transitions in aqueous solutions of native and truncated human beta B1-crystallin. AB - Human betaB1-crystallin is a major eye-lens protein that undergoes in vivo truncation at the N-terminus with aging. By studying native betaB1 and truncated betaB1DeltaN41, which mimics an age-related in vivo truncation, we have determined quantitatively the effect of truncation on the oligomerization and phase transition properties of betaB1 aqueous solutions. The oligomerization studies show that the energy of attraction between the betaB1DeltaN41 proteins is about 10% greater than that of the betaB1 proteins. We have found that betaB1DeltaN41 aqueous solutions undergo two distinct types of phase transitions. The first phase transition involves an initial formation of thin rodlike assemblies, which then evolve to form crystals. The induction time for the formation of rodlike assemblies is sensitive to oligomerization. The second phase transition can be described as liquid-liquid phase separation (LLPS) accompanied by gelation within the protein-rich phase. We refer to this process as heterogeneous gelation. These two phase transitions are not observed in the case of betaB1 aqueous solutions. However, upon the addition of poly(ethylene glycol) (PEG), we observe heterogeneous gelation also for betaB1. Our PEG experiments allow us to estimate the difference in phase separation temperatures between betaB1 and betaB1DeltaN41. This difference is consistent with the increase in energy of attraction found in our oligomerization studies. Our work suggests that truncation is a cataractogenic modification since it favors protein condensation and the consequent formation of light scattering elements, and highlights the importance of the N-terminus of betaB1 in maintaining lens transparency. PMID- 15667226 TI - Thermodynamics of a protein acylation: activation of Escherichia coli hemolysin toxin. AB - HlyC, hemolysin-activating lysine-acyltransferase, catalyses the acylation (from acyl-acyl carrier protein [ACP]) of Escherichia coli prohemolysin (proHlyA) on the epsilon-amino groups of specific lysine residues, 564 and 690 of the 1024 amino acid primary structure, to form hemolysin (HlyA). Isothermal titration calorimetry was used to measure the thermodynamic properties of the protein acylation of proHlyA-derived structures, altered by substantial deletions and separation of the acylation sites into two different peptides and site directed mutation analyses of acylation sites. Acylation of proHlyA-derived proteins catalyzed by HlyC was overall an exothermic reaction driven by a negative enthalpy. The reaction, whose kinetics are compatible to a ping-pong mechanism, is composed of two partial reactions. The first, the formation of an acyl-HlyC intermediate, was entropically driven, most likely by noncovalent complex formation between acyl-ACP and HlyC; enthalpy-driven acyl transfer followed, resulting in acyl-HlyC and ACPSH product formation. The second partial reaction was an energetically unfavorable acyl transfer from acyl-enzyme intermediate to the final acyl acceptor, a proHlyA derivative. Overall the acylation of proHlyA derived proteins catalyzed by HlyC was driven by the energetics of the acyl enzyme intermediate reaction. Of the two acylation sites, intactness of the site equivalent to proHlyA K564 was more important for acylation reaction thermodynamic stability. PMID- 15667227 TI - Specific triazine resistance in bacterial reaction centers induced by a single mutation in the QA protein pocket. AB - We report here the first example of a reaction center mutant from Rhodobacter sphaeroides, where a single mutation (M266His --> Leu) taking place in the primary quinone protein pocket confers selective resistance to triazine-type inhibitors (terbutryn, ametryn, and atrazine), which bind in the secondary quinone protein pocket, at about 13 A from the mutation site. The M266His --> Leu mutation involves one of the iron atom ligands. Interestingly, neither the secondary quinone nor the highly specific inhibitor stigmatellin binding affinities are affected by the mutation. It is noticeable that in the M266His --> Ala mutant a nativelike behavior in observed. We suggest that the long side chain of Leu in position M266 may lack space to accommodate in the Q(A) pocket therefore transferring its hindrance to the Q(B) pocket. This may occur via the structural feature formed by the Q(A)-M219His-Fe-L190His-inhibitor (or Q(B)) connection, pushing L189Leu and/or L229Ile in closer contact to the triazine molecules, therefore decreasing their bindings. This opens the possibility to finely tune, in reaction center proteins, the affinity for herbicides by designing mutations distant from their binding sites. PMID- 15667228 TI - Role of Arc1p in the modulation of yeast glutamyl-tRNA synthetase activity. AB - Yeast methionyl-tRNA synthetase (MetRS) and glutamyl-tRNA synthetase (GluRS) possess N-terminal extensions that bind the cofactor Arc1p in trans. The strength of GluRS-Arc1p interaction is high enough to allow copurification of the two macromolecules in a 1:1 ratio, in contrast to MetRS. Deletion analysis from the C terminal end of the GluRS appendix combined with previous N-terminal deletions of GluRS allows restriction of the Arc1p binding site to the 110-170 amino acid region of GluRS. This region has been shown to correspond to a novel protein protein interaction domain present in both GluRS and Arc1p but not in MetRS [Galani, K., Grosshans, H., Deinert, K., Hurt, E. C., and Simos, G. (2001) EMBO J. 20, 6889-6898]. The GluRS apoenzyme fails to show significant kinetics of tRNA aminoacylation and charges unfractionated yeast tRNA at a level 10-fold reduced compared to Arc1p-bound GluRS. The K(m) values for tRNA(Glu) measured in the ATP PP(i) exchange were similar for the two forms of GluRS, whereas k(cat) is increased 2-fold in the presence of Arc1p. Band-shift analysis revealed a 100 fold increase in tRNA binding affinity when Arc1p is bound to GluRS. This increase requires the RNA binding properties of the full-length Arc1p since Arc1p N domain leaves the K(d) of GluRS for tRNA unchanged. Transcripts of yeast tRNA(Glu) were poor substrates for measuring tRNA aminoacylation and could not be used to clarify whether Arc1p has a specific effect on the tRNA charging reaction. PMID- 15667229 TI - The thermodynamic landscape of testosterone binding to cytochrome P450 3A4: ligand binding and spin state equilibria. AB - Human cytochrome P450 (CYP) 3A4 catalyzes the oxygen-dependent metabolism of greater than 60% of known drugs. CYP3A4 binds multiple ligands simultaneously, and this contributes to complex allosteric kinetic behavior. Substrates that bind to this enzyme change the ferric spin state equilibrium of the heme, which can be observed by optical absorbance and electron paramagnetic resonance (EPR) spectroscopy. The ligand-dependent spin state equilibrium has not been quantitatively understood for any ligands that exhibit multiple binding. The CYP3A4 substrate testosterone (TST) has been shown previously by absorbance spectroscopy to induce spin state changes that are characteristic of a low spin to high spin conversion. Here, EPR was used to examine the equilibrium binding of TST to CYP3A4 at [CYP3A4] > K(D), which allows for characterization of the singly occupied state (i.e., CYP3A4.TST). We also have used absorbance spectroscopy to examine equilibrium binding, where [CYP3A4] < K(D), which allows for determination of K(D)'s. The combination of absorbance and EPR spectroscopy at different CYP3A4 concentrations relative to K(D) and curve fitting of the resultant equilibrium binding titration curves to the Adair-Pauling equations, and modifications of it, reveals that the first equivalent of TST binds with higher affinity than the second equivalent of TST and its binding is positively cooperative with respect to ligand-dependent spin state conversion. Careful analysis of the EPR and absorbance spectral results suggests that the binding of the second TST induces a shift to the high spin state and thus that the second TST binding causes displacement of the bound water. A model involving six thermodynamic states is presented and this model is related to the turnover of the enzyme. PMID- 15667230 TI - Prescribing of antipsychotic medication in a medicaid population: use of polytherapy and off-label dosages. AB - OBJECTIVES: To describe the use of atypical antipsychotic medications in a Medicaid-enrolled population composed primarily of elderly and disabled patients. Our analyses focused upon the frequency of use of polytherapy with multiple antipsychotic medications and the prescribing of off-label dosages. METHODS: We conducted a cross-sectional retrospective analysis of oral antipsychotic medication use, as prescribed for this population in 2003. The unit of analysis was the patient. We determined the prevalence of use of each type of antipsychotic medication according to gender and age group and determined the extent of use of combination therapies with multiple oral antipsychotic medications. Using the dosage ranges described in the product labeling, we identified the percentage of patients prescribed in-range dosages, overall and for each atypical antipsychotic medication studied. Those identified as receiving out-of-range (off-label) dosages were further stratified by gender and age group. The statistical significance of differences between these proportions was assessed using the chi-square test. RESULTS: Of the 8,616 patients meeting our inclusion criteria, 7,748 (90%) received monotherapy with an oral antipsychotic medication and 868 patients (10%) received polytherapy with multiple oral antipsychotic medications. Approximately 2 of 3 patients receiving atypical antipsychotic medications were prescribed a dosage that was within the range recommended in the product labeling. Dosages lower than recommended in the product labeling were prescribed for 27% of patients receiving atypical antipsychotics, while 6% of patients received an above-range dosage. The frequency of patients receiving in-range dosages varied substantially among medications. Younger patients and male patients were more frequently prescribed above-range dosages while older patients and female patients were more frequently prescribed below-range dosages of these medications (P<0.001 for both findings). CONCLUSION: In this subpopulation of Medicaid enrollees who were prescribed antipsychotic medications, we found a 10% incidence of use of antipsychotic polytherapy and a 33% incidence of prescribing of dosages outside the range listed in the product labeling. These findings suggest that physicians commonly prescribe antipsychotic medications in a manner that differs from the recommendations described in the prescribing information. The off-label use of atypical antipsychotic medications raises important questions regarding the purpose and applicability of the product labeling and the role and ability of the pharmacist to provide information regarding the risks and benefits of therapy as commonly prescribed. PMID- 15667231 TI - The impact of chronic obstructive pulmonary disease on long-term disability costs. AB - OBJECTIVES: Chronic obstructive pulmonary disease (COPD) as a cause of disability with subsequent costs remains poorly recognized. The small, growing body of literature on COPD shows that it is one of the leading causes of missed work.greater than asthma or diabetes. However, much less is known about the impact of COPD on long-term disability (LTD). Because the health care burden for disabled, working-age patients will fall heavily on managed care organizations, better estimates of the economic and pharmacoeconomic costs of COPD are required. We seek to improve understanding of the burden of COPD on several national LTD programs. METHODS: We reviewed occupational health and disability literature and government statistics to determine how long-term, respiratory-related disability is addressed by disability pension programs in 8 developed countries (Canada, France, Germany, Italy, Japan, Spain, the United Kingdom, and the United States). We then applied respiratory-specific disability definitions to country-specific population and pension information to estimate the potential burden of COPD on LTD insurance programs in each country. RESULTS: Comprehensive, relevant data to evaluate respiratory-related disability are lacking. Of the study countries, only the United States has explicit respiratory specific criteria for disability eligibility, which are based solely on spirometry. We estimate that the total burden of COPD in the study countries may range from 5 billion dollars to as high as 25 billion dollars per year if all persons who met U.S. eligibility criteria for respiratory-related disability were granted compensation. CONCLUSION: The potential burden of COPD on LTD programs may be large. The lack of standard criteria for respiratory-related disability may lead to underrecognition of COPD's true potential impact. Further work is needed to develop consistent and cost-effective ways to measure the impact of COPD and to assist in disability determination for COPD patients. PMID- 15667232 TI - Clinical monograph for drug formulary review: systemic agents for psoriasis/psoriatic arthritis. AB - BACKGROUND: Significant advances in the pharmacologic treatment of psoriasis, most notably the introduction of the biologic agents efalizumab and alefacept, have occurred recently. In addition, another biologic agent, etanercept, was recently approved for the treatment of psoriasis and psoriatic arthritis, thus adding to the list of biologic agents approved for the treatment of these disease states. A review was conducted by the Drug Information Service of a pharmacy benefits manager (PBM) to determine the relative merits and place in therapy of commonly used systemic agents for the treatment of psoriasis and psoriatic arthritis. OBJECTIVE: To provide readers with a comprehensive clinical monograph on psoriasis and psoriatic arthritis agents, written with a managed care perspective, as used in actual drug formulary decision making by a PBM. METHODS: The drug formulary of this PBM is designed to provide health plans with an evidence-based review of drugs, therapeutic classes, and disease states with a managed care focus. For each therapeutic class or disease review, an extensive and thorough literature search of MEDLINE is conducted for efficacy, safety, effectiveness, and humanistic and economic data. Drug/disease-state databases (UpToDate online, MICROMEDEX), U.S. Food and Drug Administration clinical reviews, key Internet sites, medical/pharmacy-related news sites, clinical guidelines, and AMCP dossiers are also reviewed. Formulary drug monographs produced by the Drug Information Service of the PBM include a critical analysis and summary of disease-oriented and patient-oriented clinical outcomes, effectiveness, and humanistic data. Additional data considered and included in the formulary review process are clinical attributes, patent expirations/generic competition, off-label or pending indications, and pharmacoeconomic data. RESULTS: The biologic agents do not appear to be as efficacious as traditional systemic therapies but are associated with fewer long-term toxicities that often limit treatment duration with traditional systemic agents. Although no head-to head comparisons between alefacept and efalizumab exist, efalizumab appears to offer slightly higher efficacy rates, while alefacept has a longer duration of action. Etanercept at the higher approved dose appears more efficacious compared with efalizumab or alefacept for the treatment of psoriasis, and it is the only biologic currently approved for the treatment of psoriatic arthritis. Efalizumab and alefacept are generally well tolerated, but rebound flare of psoriasis is associated with efalizumab, thus requiring continuous treatment to avoid a flare in disease. Efalizumab and etanercept can be self-administered by the patient, while alefacept and infliximab require administration by a health care professional. CONCLUSIONS: Systemic therapy is reserved for patients with moderate-to-severe psoriasis or patients with psoriatic arthritis. The biologic agents are not as efficacious as traditional therapies but, due to better tolerability, are gaining acceptance in the treatment of psoriasis and psoriatic arthritis. The biologic agents differ in efficacy rates and are generally well tolerated. Clinical attributes, overall efficacy, and economic costs associated with the biologic agents will be significant factors in selecting agents for the treatment of psoriasis and psoriatic arthritis. PMID- 15667233 TI - Formulary review of therapeutic alternatives for atopic dermatitis: focus on pimecrolimus. AB - OBJECTIVE: Atopic dermatitis (AD), often called eczema, is characterized by intense pruritus, erythema, dry skin, and inflammation. The condition is chronic and relapsing, and often occurs in patients with a family history of the atopic triad (asthma, allergic rhinitis, and AD). Use of topical steroids has been the mainstay of medical treatment for AD. Steroid-free treatments for AD, with a more favorable safety profile, have become available within the past 2 years. Tacrolimus ointment, a topical immunomodulator, became available in early 2001 and is indicated for moderate-to-severe AD. A similar but highly skinselective cytokine inhibitor, pimecrolimus cream 1%, became available in March 2002. Pimecrolimus is indicated for mild-to-moderate AD. The objective of this article is to review the key characteristics that differentiate pimecrolimus from steroids and tacrolimus in the treatment of AD. METHODS: Using secondary resources, the clinical aspects and conventional treatment strategies for AD are reviewed as are the pivotal clinical studies with pimecrolimus and literature on quality of life and economic burden of disease for AD patients and families. SUMMARY: Pimecrolimus is an effective, steroid-sparing therapy for mild tomoderate AD. Early treatment prevents flares, the agent works quickly to reduce signs and symptoms of more advanced AD, and it is safe and appropriate for intermittent long-term therapy. Pimecrolimus has fewer side effects than topical steroids and a better side-effect profile than tacrolimus. It can also be used as a first-line therapy. In studies with patients aged 2 to 17 years, it has been shown to be particularly effective in improving eczema of the face and neck, and its use may improve quality of life for many patients, especially children. A single-strength dose (1%) is safe and medically beneficial for pediatric, adolescent, and adult patients. The direct drug cost of pimecrolimus compares favorably with tacrolimus, but it is significantly more expensive than generic topical steroid creams. PMID- 15667234 TI - Health plan budget impact analysis for pimecrolimus. AB - OBJECTIVE: Budget impact models are useful tools for managed care organizations to make drug formulary decisions. The objective of this study was to estimate the incremental budgetary change in per-member-per-month (PMPM) medical and pharmacy costs for atopic dermatitis (AD) or eczema after the introduction of pimecrolimus cream 1%, a topical calcineurin inhibitor. METHODS: Estimates of the percentage of patients seeking care, treatment patterns, and quantities of medications dispensed for AD were measured using 2001 and 2002 medical and pharmacy records in a proprietary database for health plans distributed throughout the United States. Approximately 2.5 million health plan members had continuous health insurance coverage during the study period. Costs for medications were assigned using the 2003 wholesale acquisition cost, and costs for physician visits were based on average 2003 Medicare reimbursement rates. Efficacy data from clinical trials were used to model the impact of pimecrolimus on subsequent physician visits. Sensitivity analyses were performed to evaluate the impact of varying the percentage of patients seeking care, practice patterns, medication quantities, percentage of pimecrolimus users, and levels of patient cost sharing. RESULTS: The estimated percentage of health plan members seeking care for AD in 2001 was 3.2%. The estimated total cost PMPM for AD treatment prior to introduction of pimecrolimus was 0.362 dollars for all covered lives, assuming no patient cost sharing. In the year after its introduction, 5.2% of the AD population filled a prescription for pimecrolimus. The incremental increase in pharmacy benefit cost was 0.008 dollars PMPM in 2003 dollars, but the total incremental medical and pharmacy cost was 0.002 dollars PMPM after accounting for the projected reduction in physician visit costs, representing a 0.7% increase in all AD-related costs. Based on sensitivity analyses, the incremental total cost PMPM after the introduction of pimecrolimus ranged from -0.004 dollars to 0.026 dollars. CONCLUSION: Using claims data for the medical treatment of AD in 2001-2002 and the utilization of pimecrolimus, the addition of pimecrolimus as a treatment option for AD had a minimal impact on PMPM costs for AD-related care in 2003 dollars. As with all pharmacoeconomic models, health plans should perform their own budget forecasting using assumptions derived from their own pharmacy and medical claims data. PMID- 15667235 TI - Older adults' drug benefit beliefs: a focus group study. AB - OBJECTIVES: The Medicare Prescription Drug Improvement and Modernization Act will provide drug benefits for a large proportion of persons aged 65 years and older in the United States. Few studies have examined the beliefs and attitudes of older adults with respect to prescription drug insurance programs. The objective of this study was to better understand the nature and range of older adults. beliefs regarding prescription drug benefits. METHODS: This study employed a qualitative, focus group design. Three focus groups with a total of 19 community dwelling adults aged 65 years and older were conducted in June 2003. The participants were members of the Minnesota Seniors Federation and included persons with and without prescription drug insurance. Discussions were structured and guided by an interview schedule developed a priori. The focus groups were audiotaped and transcribed verbatim. Thematic textual analysis was used to identify codes and categories from the language and ideas of the group participants. RESULTS: Study participants identified a variety of important drug benefit facets. The common themes identified from the 3 groups were: (1) prescription drug access, (2) drug benefit comprehensibility, (3) powerful others, (4) affordability, and (5) equity. CONCLUSION: Older adults view drug benefits as complex entities composed of at least 5 dimensions. In addition to more commonly discussed issues such as access and affordability, seniors evaluate several other aspects of drug insurance programs such as fairness, the ease with which plan terms can be understood, and the degree to which outside actors influence plan policies. PMID- 15667236 TI - Bipolar disorder pills in perspective: questions from peer review. PMID- 15667237 TI - COPD-whose burden, how large, and what to do about it? PMID- 15667238 TI - P&T committees--black boxes versus AMCP format, and what is the true cost and value of pimecrolimus. PMID- 15667239 TI - Prescription-equivalent over-the-counter drugs for allergy, heartburn, and cholesterol reduction. PMID- 15667240 TI - Benchmarking as a management tool in decision making. PMID- 15667241 TI - Essential steps and practical applications for database studies. AB - As the information that is collected from health care encounters becomes more available, managed care pharmacy will have greater insights on the impact of pharmaceutical policy on patient outcomes. Database studies provide valuable information that depicts actual health care consumption and provides a tool to help manage the health care benefit. As compared with clinical trials, one of the strengths of database studies is that they are nonintrusive to patients and providers. However, the integrity of the data and any subsequent analysis are dependent on accurate and consistent coding practices at the time of data entry into the system. This article describes the 6 main steps required to complete a database study. PMID- 15667243 TI - Applications of disease benchmarks and case presentations. AB - A large dataset of integrated pharmacy and medical claims, extracted from independent third-party databases, is being combined with disease benchmarking technology to facilitate analysis of inpatient, outpatient, ancillary services, and pharmaceutical utilization and costs. The Disease Benchmarks Program was developed to create opportunities for health care decision makers to evaluate the entire health care continuum in a disease-specific fashion. The Benchmarks program is valuable because of its flexibility and because it depicts what is occurring in clinical practice. It can be customized and also show regional variations in treatment. The possible applications of benchmarking applications are discussed in the case presentations of otitis externa, acute otitis media with tympanostomy tubes, and Sjorgren's syndrome. PMID- 15667242 TI - Usefulness of database studies as applied to managed care. AB - Benchmarking, an industry comparison tool and a well-known business technology, offers health care a method of establishing standards for health care use with clinical governance. Benchmarking can lead to practice innovations necessary for survival in a managed care environment that has a need for decreasing cost and increasing quality. Information gleaned from the benchmarking dataset can be used to determine where limited resources for disease management programs should be directed. It can also be used to help decision makers manage a drug formulary by providing a basic knowledge about the environment in which a drug will be used and prescribed. This article describes the relationship of database studies and benchmarking and the usefulness as applied to managed care. PMID- 15667247 TI - High-throughput metabolic state analysis: the missing link in integrated functional genomics of yeasts. AB - The lack of comparable metabolic state assays severely limits understanding the metabolic changes caused by genetic or environmental perturbations. The present study reports the application of a novel derivatization method for metabolome analysis of yeast, coupled to data-mining software that achieve comparable throughput, effort and cost compared with DNA arrays. Our sample workup method enables simultaneous metabolite measurements throughout central carbon metabolism and amino acid biosynthesis, using a standard GC-MS platform that was optimized for this purpose. As an implementation proof-of-concept, we assayed metabolite levels in two yeast strains and two different environmental conditions in the context of metabolic pathway reconstruction. We demonstrate that these differential metabolite level data distinguish among sample types, such as typical metabolic fingerprinting or footprinting. More importantly, we demonstrate that this differential metabolite level data provides insight into specific metabolic pathways and lays the groundwork for integrated transcription metabolism studies of yeasts. PMID- 15667248 TI - Bio-hydrogen: microbes have their own agenda. AB - The international hydrogenase community stages a meeting every 3-4 years, and this time it was held in the University of Reading. Each congress has revealed new aspects of the complex nature of hydrogenases: their structure, molecular mechanisms and biosynthesis are all yielding up their secrets. This following introduction provides background information, and the highlights of the meeting are presented in the papers that follow. PMID- 15667249 TI - Electrocatalytic proton reduction by dithiolate-bridged diiron carbonyl complexes: a connection to the H-cluster? AB - Spectroscopic and electrochemical investigation of electrocatalytic proton reduction by Fe(2)(mu-pdt)(CO)(6), 1, have been interpreted in terms of a reaction scheme involving sequential electron-proton reactions to give a two electron, two-proton product that undergoes rate-limiting dihydrogen elimination. Further reduction, at slightly higher negative potentials, gives a more reactive product and this process dominates reactions conducted at higher acid concentrations. Inhibition of the electrocatalytic reaction by CO is due to the more efficient loss of catalyst and this is best modelled by a reaction that is second order in terms of 1(-). During electrocatalytic proton reduction a new species is observed, which features a bridging CO group and the wavenumbers of the nu(CO) modes of the terminally bound carbonyl groups are similar to those of the carbonyl groups bound to the oxidized form of the H-cluster. PMID- 15667250 TI - EPR experiments to elucidate the structure of the ready and unready states of the [NiFe] hydrogenase of Desulfovibrio vulgaris Miyazaki F. AB - Isolation and purification of the [NiFe] hydrogenase of Desulfovibrio vulgaris Miyazaki F under aerobic conditions leads to a mixture of two states, Ni-A (unready) and Ni-B (ready). The two states are distinguished by different activation times and different EPR spectra. HYSCORE and ENDOR data and DFT calculations show that both states have an exchangeable proton, albeit with a different (1)H hyperfine coupling. This proton is assigned to the bridging ligand between Ni and Fe. For Ni-B, a hydroxo ligand is found. For Ni-A, either a hydroxo in a different orientation or a hydroperoxo-bridging ligand is present. PMID- 15667251 TI - On the relationship between affinity for molecular hydrogen and the physiological directionality of hydrogenases. AB - The physiological significance of the generic reaction H(2)<-->2[H] is not always clear because hydrogenases may function in the breakdown of molecular hydrogen or in its synthesis or in both directions. Fe-hydrogenases have nevertheless been most often associated with proton reduction and NiFe-hydrogenases with hydrogen oxidation. A re-determination of the K(M) of H(2) oxidation by Pyrococcus furiosus NiFe-hydrogenase-I and by Desulfovibrio vulgaris Fe-hydrogenase suggests that affinity for hydrogen has been seriously underestimated and that the kinetics of hydrogen activation in relation to the directionality of hydrogenases should be re-evaluated. PMID- 15667252 TI - Treatment of spin-coupled metal-centres in pulsed electron-electron double resonance experiments. AB - In applications of ELDOR (electron-electron double-resonance) spectroscopy to metal centres, significant g-anisotropy and spin-coupling within multinuclear clusters have to be considered. We show the difficulties and the advantages arising from these effects. PMID- 15667253 TI - A semi-empirical molecular orbital scheme to study electron transfer in iron sulphur proteins. AB - We describe the use of the semi-empirical molecular orbital method PM3 (parametric method 3) to study the electronic structure of iron-sulphur proteins. We first develop appropriate parameters to describe models of the redox site of rubredoxins, followed by some preliminary calculations of multinuclear iron systems of relevance to hydrogenases. PMID- 15667254 TI - Hydrogen metabolism in the hyperthermophilic bacterium Aquifex aeolicus. AB - Aquifex aeolicus is a microaerophilic, hydrogen-oxidizing, hyperthermophilic bacterium containing three [NiFe] hydrogenases. Two of these three enzymes (one membrane-bound and one soluble) have been purified and characterized. The Aquifex hydrogenases are thermostable and tolerant to oxygen. A cellular function for the three hydrogenases has been proposed. The two membrane-bound periplasmic hydrogenases may function in energy conservation, whereas the soluble cytoplasmic hydrogenase is probably involved in the CO(2) fixation pathway. PMID- 15667255 TI - Non-standard structures of the Ni-Fe cofactor in the regulatory and the NAD reducing hydrogenases from Ralstonia eutropha. AB - Spectroscopy on two oxygen-insensitive Ni-Fe hydrogenases from Ralstonia eutropha (NAD-reducing, soluble hydrogenase; hydrogen sensor, regulatory hydrogenase) reveals non-standard catalytic behaviour and unique structures of their Ni-Fe cofactors. Possible mechanistic implications are briefly discussed. PMID- 15667256 TI - Transcriptional regulation of the uptake [NiFe]hydrogenase genes in Rhodobacter capsulatus. AB - Transcription of the hupSL genes, which encode the uptake [NiFe]hydrogenase of Rhodobacter capsulatus, is specifically activated by H(2). Three proteins are involved, namely the H(2)-sensor HupUV, the histidine kinase HupT and the transcriptional activator HupR. hupT and hupUV mutants have the same phenotype, i.e. an increased level of hupSL expression (assayed by phupS::lacZ fusion) in the absence of H(2); they negatively control hupSL gene expression. HupT can autophosphorylate its conserved His(217), and in vitro phosphotransfer to Asp(54) of its cognate response regulator, HupR, was demonstrated. The non-phosphorylated form of HupR binds to an enhancer site (5'-TTG-N(5)-CAA) of phupS localized at 162/-152 nt and requires integration host factor to activate fully hupSL transcription. HupUV is an O(2)-insensitive [NiFe]hydrogenase, which interacts with HupT to regulate the phosphorylation state of HupT in response to H(2) availability. The N-terminal domain of HupT, encompassing the PAS domain, is required for interaction with HupUV. This interaction with HupT, leading to the formation of a (HupT)(2)-(HupUV)(2) complex, is weakened in the presence of H(2), but incubation of HupUV with H(2) has no effect on the stability of the heterodimer/tetramer, HupUV-(HupUV)(2), equilibrium. HupSL biosynthesis is also under the control of the global two-component regulatory system RegB/RegA, which controls gene expression in response to redox. RegA binds to a site close to the 35 promoter recognition site and to a site overlapping the integration host factor DNA-binding site (5'-TCACACACCATTG, centred at -87 nt) and acts as a repressor. PMID- 15667257 TI - Biodiversity of uptake hydrogenase systems from legume endosymbiotic bacteria. AB - Uptake hydrogenases in legume endosymbiotic bacteria recycle hydrogen produced during the nitrogen fixation process in legume nodules. Despite the described beneficial effect on plant productivity, the hydrogen oxidation capability is not widespread in the Rhizobiaceae family. Characterization of hydrogenase gene clusters in strains belonging to Rhizobium, Bradyrhizobium and Azorhizobium reveals a similar overall genetic organization along with important differences in gene regulation. In addition, phylogenetic analysis of hup genes indicates distinct evolutionary origins for hydrogenase genes in Rhizobia. PMID- 15667258 TI - Anaerobic regulation of hydrogenase transcription in different bacteria. AB - Hydrogen metabolism is closely related to other important metabolic and energetic processes of bacterial cells, such as photosynthesis, anaerobic respiration and sulphur metabolism. Even small environmental changes influence these networks through different regulatory systems. The presence or absence of oxygen is one of the most important signals; how the cascades evolved to transmit this signal in different bacteria is summarized. In many instances, hydrogen is released only under anoxic conditions, because of bioenergetic considerations. Most [NiFe] hydrogenases are inactivated by oxygen, but many of them can be re-activated under reducing conditions. In addition to direct inactivation of the hydrogenases, oxygen can also regulate their expression. The global regulatory systems [FNR (fumarate and nitrate reduction regulator), ArcAB (aerobic respiratory control) and RegAB], which respond to alterations in oxygen content and redox conditions of the environment, have an important role in hydrogenase regulation of several bacteria. FNR-like proteins were shown to be important for the regulation of hydrogenases in Escherichia coli, Thiocapsa roseopersicina and Rhizobium leguminosarum, whereas RegA protein modulates the expression of hupSL genes in Rhodobacter capsulatus. PMID- 15667259 TI - The exceptional photofermentative hydrogen metabolism of the green alga Chlamydomonas reinhardtii. AB - The photosynthetic green alga Chlamydomonas reinhardtii is capable of performing a complex fermentative metabolism which is related to the mixed acid fermentation of bacteria such as Escherichia coli. The fermentative pattern includes the products formate, ethanol, acetate, glycerol, lactate, carbon dioxide and molecular hydrogen (H(2)). H(2) production is catalysed by an active [Fe] hydrogenase (HydA) which is coupled with the photosynthetic electron-transport chain. The most important enzyme of the classic fermentation pathway is pyruvate formate-lyase, which is common in bacteria but seldom found in eukaryotes. An interaction between fermentation, photosynthesis and H(2) evolution allows the algae to overcome long periods of anaerobiosis. In the absence of sulphur, the cells establish a photofermentative metabolism and accumulate large amounts of H(2). PMID- 15667260 TI - Formate and its role in hydrogen production in Escherichia coli. AB - The production of dihydrogen by Escherichia coli and other members of the Enterobacteriaceae is one of the classic features of mixed-acid fermentation. Synthesis of the multicomponent, membrane-associated FHL (formate hydrogenlyase) enzyme complex, which disproportionates formate into CO(2) and H(2), has an absolute requirement for formate. Formate, therefore, represents a signature molecule in the fermenting E. coli cell and factors that determine formate metabolism control FHL synthesis and consequently dihydrogen evolution. PMID- 15667261 TI - Eukaryotic Fe-hydrogenases -- old eukaryotic heritage or adaptive acquisitions? AB - All eukaryotes seem to possess proteins that most probably evolved from an ancestral Fe-hydrogenase. These proteins, known as NARF or Nar, do not produce hydrogen. Notably, a small group of rather unrelated unicellular anaerobes and a few algae possess Fe-hydrogenases, which produce hydrogen. In most, but not all organisms, hydrogen production occurs in membrane-bounded organelles, i.e. hydrogenosomes or plastids. Whereas plastids are monophyletic, hydrogenosomes evolved repeatedly and independently from mitochondria or mitochondria-like organelles. A systematic analysis of the various hydrogenosomes and their hydrogenases will contribute to an understanding of the evolution of the eukaryotic cell, and provide clues to the evolutionary origin(s) of the Fe hydrogenase. PMID- 15667262 TI - Interacting regulatory networks in the facultative photosynthetic bacterium, Rhodobacter sphaeroides 2.4.1. AB - Regulation of photosynthetic membrane synthesis in Rhodobacter sphaeroides 2.4.1 is dependent on the interactions of numerous regulatory elements, with two of the most important being the cbb(3) terminal oxidase and the PrrBAC two-component regulatory system. Here, we reveal that the cbb(3) terminal oxidase possesses extensive, additional regulatory activities under anaerobic conditions, and that the PrrBAC system is further involved in the regulation of the expression of more than 20% of the R. sphaeroides genome under anaerobic conditions, extending well beyond functions related to redox gene expression. PMID- 15667263 TI - High-level transcription of large gene regions: a novel T(7) RNA-polymerase-based system for expression of functional hydrogenases in the phototrophic bacterium Rhodobacter capsulatus. AB - High-level synthesis of complex enzymes like bacterial [NiFe] hydrogenases, in general, requires an expression system that allows concerted expression of a large number of genes. So far, it has not been possible to overproduce a hydrogenase in a stable and active form by using a customary expression system. Therefore we started to establish a new, T(7)-based expression system in the phototrophic bacterium Rhodobacter capsulatus. The beneficial properties of this bacterial host in combination with the unique capacity of T(7) RNA polymerase to synthesize long transcripts will allow the high-level synthesis and assembly of active hydrogenase as well as other complex enzymes in the near future. PMID- 15667264 TI - Construction of a [NiFe]-hydrogenase deletion mutant of Desulfovibrio vulgaris Hildenborough. AB - A mutant of Desulfovibrio vulgaris Hildenborough lacking a gene for [NiFe] hydrogenase was generated. Growth studies, performed for the mutant in comparison with the wild-type, showed no strong differences during the exponential growth phase. However, the mutant cells died more rapidly in the stationary growth phase. PMID- 15667265 TI - The hydrogenases of Thiocapsa roseopersicina. AB - The purple sulphur phototrophic bacterium, Thiocapsa roseopersicina BBS, contains several NiFe hydrogenases. One of these enzymes (HynSL) is membrane associated, remarkably stable and can be used for practical applications. HupSL is also located in the photosynthetic membrane, its properties are similar to other known Hup-type NiFe hydrogenases. A third hydrogenase activity was located in the soluble fraction and was analogous to the NAD-reducing hydrogenases of cyanobacteria. The hoxEFUYH genes are transcribed together. HoxE is needed for the in vivo electron flow to and from the soluble hydrogenase. Some of the accessory genes were identified using random mutagenesis, and sequencing of the T. roseopersicina genome is in progress. The HupD, HynD and HoxW gene products corresponded to the proteases processing the C-termini of the three NiFe hydrogenases respectively. HypF and HupK mutants displayed significant in vivo H(2) evolution, which could be linked to the nitrogenase activity for the DeltahypF and to the bidirectional Hox activity in the DeltahupK strain. Both HypC proteins are needed for the biosynthesis of each NiFe hydrogenase. The hydrogenase expression is regulated at the transcriptional level through distinct mechanisms. The expression of hynSL is up-regulated under anaerobic conditions with the participation of an FNR (fumarate and nitrate reduction regulator)-type protein, FnrT. Although the genes encoding a typical H(2) sensor (hupUV) and a two-component regulator (hupR and hupT) are present in T. roseopersicina, the system is cryptic in the wild-type BBS strain. The hupR gene was identified in the gene cluster downstream from hupSL. Introduction of actively expressed hupT repressed the hupSL gene expression as expected by analogy with other bacteria. PMID- 15667266 TI - Molecular characterization of uptake hydrogenase in Frankia. AB - A molecular characterization of uptake hydrogenase in Frankia was performed by using two-dimensional gel electrophoresis, matrix-assisted laser-desorption ionization-time-of-flight mass spectrometry, PCR amplification and Southern blotting. A polypeptide of approx. 60 kDa was recognized in Frankia UGL011102, AVCI1 and KB5 on the two-dimensional gel by blotting with Ralstonia eutropha (Hox G) antibody. Further analysis by MS resulted in a peptide 'fingerprint', which was similar to the membrane-bound hydrogenase 2 large subunit (HYD2) in Escherichia coli. In addition, a 127 bp PCR fragment could also be amplified from Frankia AVCI1, which gave a 76% similarity with the large subunit of hydrogenase in, e.g., Azotobacter chrococcum, Bradyrhizobium japonicum and Rhizobium leguminosarum. Although immunological similarity between the small subunit of Frankia hydrogenase and that of other organisms has not yet been found, a PCR product of 500 bp could be amplified from the local source of Frankia, the analysis of which gave 69 and 67% identity with the small subunit of hydrogenases in B. japonicum and R. leguminosarum respectively. A Southern-blot analysis further indicated evidence for the presence of the small hydrogenase subunit in other Frankia strains, i.e. KB5, AvcI1 and CcI3. PMID- 15667267 TI - Uptake hydrogenase in cyanobacteria: novel input from non-heterocystous strains. AB - Most of the research to date on cyanobacterial uptake hydrogenases has been performed on filamentous heterocystous strains. However, recent results on the hup gene cluster organization and its transcriptional regulation in non heterocystous strains has contributed to the widening of knowledge in this field. In the present study, we outline the recent findings on uptake hydrogenases from non-heterocystous cyanobacteria, comparing it with the presently available data from heterocystous strains, and draw attention to potential areas for future research. PMID- 15667268 TI - Approaches to developing biological H(2)-photoproducing organisms and processes. AB - The development of efficient biological systems for the direct photoproduction of H(2) gas from water faces several challenges, the more serious of which is the sensitivity of the H(2)-evolving enzymes (hydrogenases) to O(2), an obligatory by product of photosynthesis. This high sensitivity is common to both FeFe and NiFe hydrogenases, and is caused by O(2) binding to their respective metallocatalytic sites. This overview describes approaches to (i) molecular engineering of algal FeFe-hydrogenase to prevent O(2) access to its catalytic site; (ii) transform a cyanobacterium with an O(2)-tolerant bacterial NiFe hydrogenase or (c) partially inactivate algal O(2)-evolution activity to create physiologically anaerobiosis and induce hydrogenase expression. PMID- 15667269 TI - Hydrogenase electrodes for fuel cells. AB - Considering crucial problems that limit use of platinum-based fuel cells, i.e. cost and availability, poisoning by fuel impurities and low selectivity, we propose electrocatalysis by enzymes as a valuable alternative to noble metals. Hydrogenase electrodes in neutral media achieve hydrogen equilibrium potential (providing 100% energy conversion), and display high activity in H(2) electrooxidation, which is similar to that of Pt-based electrodes in sulphuric acid. In contrast with platinum, enzyme electrodes are highly selective for their substrates, and are not poisoned by fuel impurities. Hydrogenase electrodes are capable of consuming hydrogen directly from microbial media, which ensures their use as fuel electrodes in treatment of organic wastes. PMID- 15667270 TI - Applications of bacterial hydrogenases in waste decontamination, manufacture of novel bionanocatalysts and in sustainable energy. AB - Bacterial hydrogenases have been harnessed to the removal of heavy metals from solution by reduction to less soluble metal species. For Pd(II), its bioreduction results in the deposition of cell-bound Pd(0)-nanoparticles that are ferromagnetic and have a high catalytic activity. Hydrogenases can also be used synthetically in the production of hydrogen from sugary wastes through breakdown of formate produced by fermentation. The Bio-H(2) produced can be used to power an electrical device using a fuel cell to provide clean electricity. Production of hydrogen from confectionery wastes by one organism (Escherichia coli) can be used as the electron donor for the production of Bio-Pd(0) from soluble Pd(II) by a second organism. The resulting Bio-Pd(0) can then be used as a bioinorganic catalyst in the remediation of Cr(VI)-contaminated solutions or polychlorinated biphenyls at the expense of Bio-H(2), as a hydrogenation catalyst for industry or as a component of a fuel cell electrode. PMID- 15667271 TI - Molecular dynamics and experimental investigation of H(2) and O(2) diffusion in [Fe]-hydrogenase. AB - The [Fe]-hydrogenase enzymes are highly efficient H(2) catalysts found in ecologically and phylogenetically diverse microorganisms, including the photosynthetic green alga, Chlamydomonas reinhardtii. Although these enzymes can occur in several forms, H(2) catalysis takes place at a unique [FeS] prosthetic group or H-cluster, located at the active site. Significant to the function of hydrogenases is how the surrounding protein structure facilitates substrate product transfer, and protects the active site H-cluster from inactivation. To elucidate the role of protein structure in O(2) inactivation of [Fe] hydrogenases, experimental and theoretical investigations have been performed. Molecular dynamics was used to comparatively investigate O(2) and H(2) diffusion in CpI ([Fe]-hydrogenase I from Clostridium pasteurianum). Our preliminary results suggest that H(2) diffuses more easily and freely than O(2), which is restricted to a small number of allowed pathways to and from the active site. These O(2) pathways are located in the conserved active site domain, shown experimentally to have an essential role in active site protection. PMID- 15667272 TI - Use of molecular hydrogen as an energy substrate by human pathogenic bacteria. AB - Molecular hydrogen is produced as a fermentation by-product in the large intestine of animals and its production can be correlated with the digestibility of the carbohydrates consumed. Pathogenic Helicobacter species (Helicobacter pylori and H. hepaticus) have the ability to use H(2) through a respiratory hydrogenase, and it was demonstrated that the gas is present in the tissues colonized by these pathogens (the stomach and the liver respectively of live animals). Mutant strains of H. pylori unable to use H(2) are deficient in colonizing mice compared with the parent strain. On the basis of available annotated gene sequence information, the enteric pathogen Salmonella, like other enteric bacteria, contains three putative membrane-associated H(2)-using hydrogenase enzymes. From the analysis of gene-targeted mutants it is concluded that each of the three membrane-bound hydrogenases of Salmonella enterica serovar Typhimurium are coupled with an H(2)-oxidizing respiratory pathway. From microelectrode probe measurements on live mice, H(2) could be detected at approx. 50 muM levels within the tissues (liver and spleen), which are colonized by Salmonella. The half-saturation affinity of whole cells of these pathogens for H(2) is much less than this, so it is expected that the (H(2)-utilizing) hydrogenase enzymes be saturated with the reducing substrate in vivo. All three enteric NiFe hydrogenase enzymes contribute to virulence of the bacterium in a typhoid fever-mouse model, and the combined removal of all three hydrogenases resulted in a strain that is avirulent and (in contrast with the parent strain) one that is not able to pass the intestinal tract to invade liver or spleen tissue. It is proposed that H(2) utilization and specifically its oxidation, coupled with a respiratory pathway, is required for energy production to permit growth and maintain efficient virulence of a number of pathogenic bacteria during infection of animals. These would be expected to include the Campylobacter jejuni, a bacterium closely related to Helicobacter, as well as many enteric bacteria (Escherichia coli, Shigella and Yersinia species). PMID- 15667273 TI - Nar1p, a conserved eukaryotic protein with similarity to Fe-only hydrogenases, functions in cytosolic iron-sulphur protein biogenesis. AB - The genome of the yeast Saccharomyces cerevisiae encodes the essential protein Nar1p that is conserved in virtually all eukaryotes and exhibits striking sequence similarity to bacterial iron-only hydrogenases. Previously, we have shown that Nar1p is an Fe-S protein and that assembly of its co-factors depends on the mitochondrial Fe-S cluster biosynthesis apparatus. Using functional studies in vivo, we demonstrated that Nar1p has an essential role in the maturation of cytosolic and nuclear, but not of mitochondrial, Fe-S proteins. Here we provide further spectroscopic evidence that Nar1p possesses two Fe-S clusters. We also show that Nar1p is required for Fe-S cluster assembly on the P loop NTPase Nbp35p, another newly identified component of the cytosolic Fe-S protein assembly machinery. These data suggest a complex biochemical pathway of extra-mitochondrial Fe-S protein biogenesis involving unique eukaryotic proteins. PMID- 15667274 TI - NifU and NifS are required for the maturation of nitrogenase and cannot replace the function of isc-gene products in Azotobacter vinelandii. AB - In recent years, it has become evident that [Fe-S] proteins, such as hydrogenase, nitrogenase and aconitase, require a complex machinery to assemble and insert their associated [Fe-S] clusters. So far, three different types of [Fe-S] cluster biosynthetic systems have been identified and these have been designated nif, isc and suf. In the present work, we show that the nif-specific [Fe-S] cluster biosynthetic system from Azotobacter vinelandii, which is required for nitrogenase maturation, cannot functionally replace the isc [Fe-S] cluster system used for the maturation of other [Fe-S] proteins, such as aconitase. The results indicate that, in certain cases, [Fe-S] cluster biosynthetic machineries have evolved to perform only specialized functions. PMID- 15667275 TI - Genetics and biotechnology of the H(2)-uptake [NiFe] hydrogenase from Rhizobium leguminosarum bv. viciae, a legume endosymbiotic bacterium. AB - A limited number of strains belonging to several genera of Rhizobiaceae are capable of expressing a hydrogenase system that allows partial or full recycling of hydrogen evolved by nitrogenase, thus increasing the energy efficiency of the nitrogen fixation process. This review is focused on the genetics and biotechnology of the hydrogenase system from Rhizobium leguminosarum bv. viciae, a frequent inhabitant of European soils capable of establishing symbiotic association with peas, lentils, vetches and other legumes. PMID- 15667276 TI - A hydrogen-sensing multiprotein complex controls aerobic hydrogen metabolism in Ralstonia eutropha. AB - H(2) is an attractive energy source for many microorganisms and is mostly consumed before it enters oxic habitats. Thus aerobic H(2)-oxidizing organisms receive H(2) only occasionally and in limited amounts. Metabolic adaptation requires a robust oxygen-tolerant hydrogenase enzyme system and special regulatory devices that enable the organism to respond rapidly to a changing supply of H(2). The proteobacterium Ralstonia eutropha strain H16 that harbours three [NiFe] hydrogenases perfectly meets these demands. The unusual biochemical and structural properties of the hydrogenases are described, including the strategies that confer O(2) tolerance to the NAD-reducing soluble hydrogenase and the H(2)-sensing regulatory hydrogenase. The regulatory hydrogenase that forms a complex with a histidine protein kinase recognizes H(2) in the environment and transmits the signal to a response regulator, which in turn controls transcription of the hydrogenase genes. PMID- 15667277 TI - Identification of genes required for hydrogenase activity in Chlamydomonas reinhardtii. AB - The eukaryotic green alga, Chlamydomonas reinhardtii, produces H(2) under anaerobic conditions, in a reaction catalysed by an [FeFe]-hydrogenase. To identify genes that influence H(2) production in C. reinhardtii, a library of 6000 colonies on agar plates was screened with sensitive chemochromic H(2)-sensor films for clones defective in H(2) production. Two mutants of particular interest were fully characterized. One mutant, hydEF-1, is unable to assemble an active [FeFe]-hydrogenase. This is the first reported C. reinhardtii mutant that is not capable of producing any H(2). The second mutant, sta7-10, is not able to accumulate insoluble starch and has significantly lowered H(2)-photoproduction rates in comparison with the wild-type. In hydEF-1, anaerobiosis induces transcription of the two reported C. reinhardtii hydrogenase genes, HydA1 and HydA2, indicating a normal transcriptional response to anaerobiosis. In contrast, the transcription of both hydrogenase genes in sta7-10 is significantly attenuated. PMID- 15667278 TI - Common principles in the biosynthesis of diverse enzymes. AB - A subset of bacterial periplasmic enzymes are transported from the cytoplasm by the twin-arginine transport apparatus. Such proteins contain distinctive N terminal signal peptides containing a conserved SRRXFLK 'twin-arginine' amino acid motif and often bind complex cofactors before the transport event. It is important that assembly of complex cofactor-containing, and often multi-subunit, enzymes is complete before export. Studies of the unrelated [NiFe] hydrogenase, DMSO reductase and trimethylamine N-oxide reductase systems from Escherichia coli have enabled us to define a chaperone-mediated 'proofreading' mechanism involved in co-ordinating assembly and export of twin-arginine transport-dependent enzymes. PMID- 15667279 TI - [NiFe]-Hydrogenase maturation endopeptidase: structure and function. AB - Hydrogenase maturation endopeptidases catalyse the terminal step in the maturation of the large subunit of [NiFe]-hydrogenases. They remove a C-terminal extension from the precursor of the subunit, triggering a conformational switch that results in the bridging of the Fe and Ni atoms of the metal centre via the thiolate of a cysteine residue and in closure of the centre. This review summarizes what is known about the structure of the protein, its substrate specificity and its possible reaction mechanism. PMID- 15667280 TI - The emerging molecular structure of the nitrogen cycle: an introduction to the proceedings of the 10th annual N-cycle meeting. AB - Over the last 10 years, during the lifetime of the nitrogen cycle meetings, structural biology, coupled with spectroscopy, has had a major impact of our understanding enzymology of the nitrogen cycle. The three-dimensional structures for many of the key enzymes have now been resolved and have provided a wealth of information regarding the architecture of redox active metal sites, as well as revealing novel structural folds. Coupled with structure-based spectroscopic analysis, this has led to new insight into the reaction mechanisms of the diverse chemical transformations that together cycle nitrogen in the biosphere. An overview of the some of the key developments in field over the last decade is presented. PMID- 15667281 TI - 1994-2004: 10 years of research on the anaerobic oxidation of ammonium. AB - The obligately anaerobic ammonium oxidation (anammox) reaction with nitrite as primary electron acceptor is catalysed by the planctomycete-like bacteria Brocadia anammoxidans, Kuenenia stuttgartiensis and Scalindua sorokinii. The anammox bacteria use a complex reaction mechanism involving hydrazine as an intermediate. They have a unique prokaryotic organelle, the anammoxosome, surrounded by ladderane lipids, which exclusively contains the hydrazine oxidoreductase as the major protein to combine nitrite and ammonia in a one-to one fashion. In addition to the peculiar microbiology, anammox was shown to be very important in the oceanic nitrogen cycle, and proved to be a very good alternative for treatment of high-strength nitrogenous waste streams. With the assembly of the K. stuttgartiensis genome at Genoscope, Evry, France, the anammox reaction has entered the genomic and proteomic era, enabling the elucidation of many intriguing aspects of this fascinating microbial process. PMID- 15667282 TI - Chaperones involved in assembly and export of N-oxide reductases. AB - Controlled targeting and transport of redox enzymes to and across the bacterial cytoplasmic membrane is essential for bacterial respiration. A subset of bacterial redox enzymes is exported as folded proteins on the Tat (twin-arginine transport) pathway. Protein export is the point-of-no-return for passenger proteins on the Tat pathway and it is crucial that complex, cofactor-containing enzymes are fully assembled before export is attempted. Using the Escherichia coli trimethylamine N-oxide reductase system as a model, we discuss here the molecular processes governing assembly and export of Tat-dependent enzymes. PMID- 15667283 TI - Molybdate-dependent expression of the periplasmic nitrate reductase in Bradyrhizobium japonicum. AB - The napEDABC genes of Bradyrhizobium japonicum encode the periplasmic nitrate reductase, an Mo-containing enzyme which catalyses the reduction of nitrate to nitrite when oxygen concentrations are limiting. In this bacterium, another set of genes, modABC, code for a high affinity ABC-type Mo transport system. A B. japonicum modA mutant has been obtained that is not capable of growing anaerobically with nitrate and lacks nitrate reductase activity. Under nitrate respiring conditions, when Mo concentrations are limiting, the B. japonicum modA mutant lacked both the 90 kDa protein corresponding to the NapA component of the periplasmic nitrate reductase, and the membrane-bound 25 kDa c-type cytochrome NapC. Regulatory studies using a napE-lacZ fusion indicated that napE expression was highly reduced in the modA mutant background when the cells were incubated anaerobically with nitrate under Mo-deficient conditions. PMID- 15667284 TI - Membrane-bound denitrification in the Gram-positive bacterium Bacillus azotoformans. AB - Gram+ bacteria are capable of complete denitrification just like Gram- (Gram negative) bacteria. However, Gram+ (Gram-positive) bacteria have a very small periplasmic-like space. This leads to the question of whether those enzymes and electron carriers involved in denitrification, which are normally located in the periplasmic space in Gram- bacteria, are located in the periplasmic-like space in Gram+ bacteria or have been modified as membrane-bound proteins. Using Bacillus azotoformans as a Gram+ bacterial model, our study demonstrates that anaerobic denitrification is catalysed by four membrane-bound enzymes and that the electron carriers are membrane-bound c-type cytochromes and menaquinol. NADH dehydrogenase is coupled with the denitrification pathway providing menaquinol. In addition, the cytochrome b(6)f complex forms part of the denitrification pathway, oxidizing menaquinol and reducing at least three different membrane-bound c-type cytochromes. We determined that the NO reductase, qCu(A)NOR (where NOR stands for nitric oxide reductase), can accept electrons from two donors, a specific cytochrome c(551) and menaquinol. Similarly, nitrite reductase, a copper enzyme, and nitrous oxide reductase may be bifunctional enzymes. Regarding the bifunctionality of qCu(A)NOR, we propose that the menaquinol-linked pathway is involved in the detoxification of NO. PMID- 15667285 TI - Microaerobic denitrification in Neisseria meningitidis. AB - The major aetiological agent of human bacterial meningitis is Neisseria meningitidis. During the course of disease and host colonization, the bacterium has to withstand limited oxygen availability. Nitrogen oxide and nitrogen oxyanions are thought to be present, which may constitute an alternative sink for electrons from the N. meningitidis respiratory chain. A partial denitrification pathway is encoded by the aniA nitrite reductase gene and the norB nitric oxide reductase gene. Analysis of the completed genome sequences of two N. meningitidis strains is used to generate a model for the membrane-associated respiratory chain of this organism. Analysis of aniA expression indicates it to be controlled primarily by oxygen and secondarily by nitrite. The ability of N. meningitidis to denitrify relies on microaerobic growth conditions. Here we show that under microaerobic conditions nitrite supplements oxygen as an alternative respiratory substrate. PMID- 15667286 TI - Cytochrome c nitrite reductase: from structural to physicochemical analysis. AB - The recent structural characterization of the NrfA from Escherichia coli provides a framework to rationalize the spectroscopic and functional properties of this enzyme. Analyses by EPR and magnetic CD spectroscopies have been complemented by protein-film voltammetry and these are discussed in relation to the essential structural features of the enzyme. PMID- 15667287 TI - The complete denitrification pathway of the symbiotic, nitrogen-fixing bacterium Bradyrhizobium japonicum. AB - Denitrification is an alternative form of respiration in which bacteria sequentially reduce nitrate or nitrite to nitrogen gas by the intermediates nitric oxide and nitrous oxide when oxygen concentrations are limiting. In Bradyrhizobium japonicum, the N(2)-fixing microsymbiont of soya beans, denitrification depends on the napEDABC, nirK, norCBQD, and nosRZDFYLX gene clusters encoding nitrate-, nitrite-, nitric oxide- and nitrous oxide-reductase respectively. Mutational analysis of the B. japonicum nap genes has demonstrated that the periplasmic nitrate reductase is the only enzyme responsible for nitrate respiration in this bacterium. Regulatory studies using transcriptional lacZ fusions to the nirK, norCBQD and nosRZDFYLX promoter region indicated that microaerobic induction of these promoters is dependent on the fixLJ and fixK(2) genes whose products form the FixLJ-FixK(2) regulatory cascade. Besides FixK(2), another protein, nitrite and nitric oxide respiratory regulator, has been shown to be required for N-oxide regulation of the B. japonicum nirK and norCBQD genes. Thus nitrite and nitric oxide respiratory regulator adds to the FixLJ-FixK(2) cascade an additional control level which integrates the N-oxide signal that is critical for maximal induction of the B. japonicum denitrification genes. However, the identity of the signalling molecule and the sensing mechanism remains unknown. PMID- 15667288 TI - Complexity and diversity in c-type cytochrome biogenesis systems. AB - c-Type cytochromes contain haem covalently attached to protein by thioether bonds formed post-translationally and requiring a dedicated biogenesis apparatus. Three biogenesis systems, found in different cell types, are well known. Here we discuss emerging evidence for at least one additional system, for unanticipated diversity in the location of the systems and for the co-existence of multiple systems in some cells. PMID- 15667289 TI - The enigma of Paracoccus pantotrophus cytochrome cd(1) activation. AB - Paracoccus pantotrophus cytochrome cd(1) nitrite reductase is isolated under aerobic conditions from anaerobically grown cells in an inactive form. This state requires reductive activation to make it catalytically competent for nitrite reduction. In this work, we discuss the methods of this reductive activation and its consequences for the cell. PMID- 15667290 TI - The role of ResA in type II cytochrome c maturation. AB - Numerous bacterial proteins involved in the nitrogen cycle, and other processes, require c-type haem as a cofactor. c-type cytochromes are formed by covalent attachment of haem to the conserved CXXCH motif. Here, we briefly review what is presently known about cytochrome c maturation in Bacillus subtilis with particular emphasis on the crystal structures of ResA. PMID- 15667291 TI - Nitrogen fixation: key genetic regulatory mechanisms. AB - The necessity to respond to the level of fixed nitrogen and external oxygen concentrations and to provide sufficient energy for nitrogen fixation imposes common regulatory principles amongst diazotrophs. The NifL-NifA system in Azotobacter vinelandii integrates the signals of redox, fixed-nitrogen and carbon status to regulate nif transcription. Multidomain signalling interactions between NifL and NifA are modulated by redox changes, ligand binding and interaction with the signal-transduction protein GlnK. Under adverse redox conditions (excess oxygen) or when fixed nitrogen is in excess, NifL forms a complex with NifA in which transcriptional activation is prevented. Oxidized NifL forms a binary complex with NifA to inhibit NifA activity. When fixed nitrogen is in excess, the non-covalently modified form of GlnK interacts with NifL to promote the formation of a GlnK-NifL-NifA ternary complex. When the cell re-encounters favourable conditions for nitrogen fixation, it is necessary to deactivate the signals to ensure that the NifL-NifA complex is dissociated so that NifA is free to activate transcription. This is achieved through interactions with 2-oxoglutarate, a key metabolic signal of the carbon status, which binds to the N-terminal GAF (cGMP specific and stimulated phosphodiesterases, Anabaena adenylate cyclases and Escherichia coli FhlA) domain of NifA. PMID- 15667292 TI - Plasmid replicons of Rhizobium. AB - Rhizobium spp. are found in soil. They are both free-living and found symbiotically associated with the nodules of leguminous plants. Traditionally, studies have focused on the association of these organisms with plants in nitrogen-fixing nodules, since this is regarded as the most important role of these bacteria in the environment. Rhizobium sp. are known to possess several replicons. Some, like the Rhizobium etli symbiotic plasmid p42d and the plasmid pNGR234b of Rhizobium NGR234, have been sequenced and characterized. The plasmids from these organisms are the focus of this short review. PMID- 15667293 TI - Nitrate and nitrite transport in Escherichia coli. AB - Two polytopic membrane proteins, NarK and NarU, are involved in nitrate and nitrite uptake and nitrite extrusion by Escherichia coli. A third polytopic membrane protein, NirC, functions only in nitrite transport. During exponential growth, the quantity of NarU in membrane fractions was <0.01% of the quantity of NarK. During the stationary phase of growth, the ratio of NarU to NarK increased to 0.1%. However, in the exponential phase of growth, the strain expressing only NarK transports and reduces nitrate and nitrite at a rate only slightly higher than that of the strain expressing only NarU, indicating that, in a NarK(+) strain, the rate of nitrate reduction is not limited by the rate of nitrate transport. By measuring nitrate and nitrite transport abilities of strains expressing only narK or expressing both narK and nirC, we hypothesized that NarK might function as a primary nitrate-nitrite antiporter. After nitrate is imported by NarK and reduced to nitrite, some nitrite is expelled from the cell and then reimported for reduction to ammonia. Two highly conserved positively charged residues, Arg-87 and Arg-303 of NarU, were shown by site-directed mutagenesis to play a key role in anion transport. This result indicates that NarU might form a single channel for nitrate and nitrite transport. PMID- 15667294 TI - Function of the Rhizobium etli CFN42 nirK gene in nitrite metabolism. AB - Rhizobium etli CFN42 is not capable of growing anaerobically with nitrate but it grows with nitrite as a terminal electron acceptor. This bacterium contains the nirK gene encoding the copper-containing Nir (nitrite reductase), which is located on the cryptic plasmid pCFN42f. Mutational analysis has demonstrated that a nirK deficient mutant was not capable of growing under nitrite-respiring conditions. Moreover, microaerobic growth of this mutant was inhibited by the presence of nitrite. Nir activity and nitrite uptake were highly diminished in a nirK mutant, compared with the wild-type levels after incubation under anaerobic conditions. Our results suggest that the copper-containing Nir may have both a respiratory and a nitrite-detoxifying role in R. etli. PMID- 15667295 TI - Nitrogen assimilation and nitrogen control in cyanobacteria. AB - Nitrogen sources commonly used by cyanobacteria include ammonium, nitrate, nitrite, urea and atmospheric N(2), and some cyanobacteria can also assimilate arginine or glutamine. ABC (ATP-binding cassette)-type permeases are involved in the uptake of nitrate/nitrite, urea and most amino acids, whereas secondary transporters take up ammonium and, in some strains, nitrate/nitrite. In cyanobacteria, nitrate and nitrite reductases are ferredoxin-dependent enzymes, arginine is catabolized by a combination of the urea cycle and arginase pathway, and urea is degraded by a Ni(2+)-dependent urease. These pathways provide ammonium that is incorporated into carbon skeletons through the glutamine synthetase-glutamate synthase cycle, in which 2-oxoglutarate is the final nitrogen acceptor. The expression of many nitrogen assimilation genes is subjected to regulation being activated by the nitrogen-control transcription factor NtcA, which is autoregulatory and whose activity appears to be influenced by 2-oxoglutarate and the signal transduction protein P(II). In some filamentous cyanobacteria, N(2) fixation takes place in specialized cells called heterocysts that differentiate from vegetative cells in a process strictly controlled by NtcA. PMID- 15667296 TI - Alkaline cyanide biodegradation by Pseudomonas pseudoalcaligenes CECT5344. AB - Pseudomonas pseudoalcaligenes CECT5344 uses cyanide, cyanate, beta-cyanoalanine, and other cyanoderivatives as nitrogen sources under alkaline conditions, which prevents volatile HCN (pK(a) 9.2) formation. The cyanide consumed by this strain is stoichiometrically converted into ammonium. In addition, this bacterium grows with the heavy metal, cyanide-containing waste water generated by the jewellery industry, and is also a cyanide-resistant strain which induces an alternative oxidase and a siderophore-based mechanism for iron acquisition in the presence of cyanide. The detection of cyanase and beta-cyanoalanine nitrilase activities in cyanide-induced cells suggests their implication in the cyanide degradation pathway. PMID- 15667297 TI - Complex formation between AmtB and GlnK: an ancestral role in prokaryotic nitrogen control. AB - Ammonium transport proteins belonging to the Amt family are ubiquitous in prokaryotes. In Escherichia coli, the AmtB protein and the associated P(II) signal transduction protein (GlnK) have recently been recognized as an ammonium sensory system that effectively couples the intracellular nitrogen regulation (Ntr) system to external changes in ammonium availability. Given the almost invariant coupling of AmtB and GlnK in bacteria and archaea it seems probable that these two proteins may constitute an ancestral nitrogen-responsive system that has been coupled with a variety of unrelated nitrogen regulatory processes, which are now found in prokaryotes. The multiplicity of P(II) proteins could therefore be considered to have evolved from an ancestral GlnK-like protein and to have subsequently been adapted to control many other aspects of nitrogen metabolism. PMID- 15667298 TI - Microbial reduction of selenate and nitrate: common themes and variations. AB - A number of biochemically distinct systems have been characterized for the microbial reduction of the oxyanions, selenate (SeO(4)(2-)) and nitrate (NO(3)( )). Two classes of molybdenum-dependent nitrate reductase catalyse the respiratory-linked reduction of nitrate (NO(3)(-)) to nitrite (NO(2)(-)). The main respiratory nitrate reductase (NAR) is membrane-anchored, with its active site facing the cytoplasmic compartment. The other enzyme (NAP) is water-soluble and located in the periplasm. In recent years, our understanding of each of these enzyme systems has increased significantly. The crystal structures of both NAR and NAP have now been solved and they provide new insight into the structure, function and evolution of these respiratory complexes. In contrast, our understanding of microbial selenate (SeO(4)(2-)) reduction and respiration is at an early stage; however, similarities to the nitrate reductase systems are emerging. This review will consider some of the common themes and variations between the different classes of nitrate and selenate reductases. PMID- 15667299 TI - Nitric oxide and nitrosative stress tolerance in bacteria. AB - Nitric oxide is not only an obligatory intermediate in denitrification, but also a signalling and defence molecule of major importance. However, the basis of resistance to NO and RNS (reactive nitrogen species) is poorly understood in many microbes. The cellular targets of NO and RNS [e.g. metalloproteins, thiols in proteins, glutathione and Hcy (homocysteine)] may themselves serve as signal transducers, sensing NO and RNS, and resulting in altered gene expression and synthesis of protective enzymes. The properties of a number of such protective mechanisms are outlined here, including globins, flavorubredoxin, diverse enzymes with NO- or S-nitrosothiol-reducing properties and other redox proteins with poorly defined roles in protection from nitrosative stresses. However, the most fully understood mechanism for NO detoxification involves the enterobacterial flavohaemoglobin (Hmp). Aerobically, Hmp detoxifies NO by acting as an NO denitrosylase or 'oxygenase' and thus affords inducible protection of growth and respiration, and aids survival in macrophages. The flavohaemoglobin-encoding gene of Escherichia coli, hmp, responds to the presence of NO and RNS in an SoxRS independent manner. Nitrosating agents, such as S-nitrosoglutathione, deplete cellular Hcy and consequently modulate activity of the MetR regulator that binds the hmp promoter. Regulation of Hmp synthesis under anoxic conditions involves nitrosylation of 4Fe-4S clusters in the global transcriptional regulator, FNR. The foodborne microaerophilic pathogen, Campylobacter jejuni, also expresses a haemoglobin, Cgb, but it does not possess the reductase domain of Hmp. A Cgb deficient mutant of C. jejuni is hypersensitive to RNS, whereas cgb expression and holoprotein synthesis are specifically increased on exposure to RNS, resulting in NO-insensitive respiration. A 'systems biology' approach, integrating the methodologies of bacterial molecular genetics and physiology with post-genomic technologies, promises considerable advances in our understanding of bacterial NO tolerance mechanisms in pathogenesis. PMID- 15667300 TI - DNA binding properties of the Escherichia coli nitric oxide sensor NorR: towards an understanding of the regulation of flavorubredoxin expression. AB - Nitric oxide is an intermediate of denitrification, and is one of the radical species deployed by macrophages against invading pathogens, therefore bacterial responses to NO are of considerable importance. The Escherichia coli flavorubredoxin and its associated oxidoreductase reduce NO to nitrous oxide under anaerobic conditions, and are encoded by the norVW transcription unit. Expression of norVW requires the NO sensing regulatory protein NorR and is dependent on RNA polymerase containing the alternative sigma factor, sigma(54). We have purified NorR and shown that it binds to three sites in the norVW promoter region, located 75-140 bp upstream of the experimentally verified transcription start site. We have also identified two binding sites for the integration host factor, one between the NorR sites and the sigma(54)-RNA polymerase binding site, and a second downstream of the norVW transcription start site. Comparison of the norVW promoters of enteric bacteria along with known and putative NorR-regulated promoters from Vibrio, Ralstonia and Pseudomonas species suggests that NorR binding sites contain an invariant GT(N7)AC motif flanking an AT-rich central region. The identification of a consensus for NorR binding sites will help to elucidate additional members of the NorR regulon. PMID- 15667301 TI - N-oxide sensing in Pseudomonas aeruginosa: expression and preliminary characterization of DNR, an FNR-CRP type transcriptional regulator. AB - In denitrifying bacteria, the concentration of NO is maintained low by a tight control of the expression and activity of nitrite and NO reductases. Regulation involves redox-linked transcription factors, such as those belonging to the CRP FNR (cAMP receptor protein-fumarate and nitrate reductase regulator) superfamily, which act as oxygen and N-oxide sensors. Given that few members of this superfamily have been characterized in detail, we have cloned, expressed and purified the dissimilative nitrate respiration regulator from Pseudomonas aeruginosa. To gain insights on the structural properties of the dissimilative nitrate respiration regulator, we have also determined the aggregation state of the purified protein and its ability to bind hydrophobic compounds such as 8 anilino-1-naphthalenesulphonic acid. PMID- 15667302 TI - Purification and characterization of cytochrome c' from Neisseria meningitidis. AB - Cytochrome c', a c-type cytochrome with unique spectroscopic and magnetic properties, has been characterized in a variety of denitrifying and photosynthetic bacteria. Cytochrome c' has a role in defence and/or removal of NO but the mechanism of action is not clear. To examine the function of cytochrome c' from Neisseria meningitidis, the protein was purified after heterologous overexpression in Escherichia coli. The electronic spectra of the oxidized c' demonstrated a pH-dependent transition (over the pH range of 6-10) typical of known c'-type cytochromes. Interestingly, the form in which NO is supplied determines the redox state of the resultant haem-nitrosyl complex. Fe(III)-NO complexes were formed when Fe(II) or Fe(III) cytochrome c' was sparged with NO gas, whereas an Fe(II)-NO complex was generated when NO was supplied using DEA NONOate (diazeniumdiolate). PMID- 15667304 TI - Transcriptional regulation of nitric oxide reduction in Ralstonia eutropha H16. AB - Nitric oxide reduction in Ralstonia eutropha H16 is catalysed by the quinol dependent NO reductase NorB. norB and the adjacent norA form an operon that is controlled by the sigma(54)-dependent transcriptional activator NorR in response to NO. A NorR derivative containing MalE in place of the N-terminal domain binds to a 73 bp region upstream of norA that includes three copies of the putative upstream activator sequence GGT-(N(7))-ACC. Mutations altering individual bases of this sequence resulted in an 80-90% decrease in transcriptional activation by wild-type NorR. Similar motifs are present in several proteobacteria upstream of genes encoding proteins of NO metabolism. The N-terminal domain of NorR contains a GAF module and is hypothesized to interact with a signal molecule. A NorR derivative lacking this domain activates the norAB promoter constitutively. Amino acid exchanges within the GAF module identified a cysteine residue that is essential for promoter activation by NorR. Signal sensing by NorR is negatively modulated by the iron-containing protein NorA. PMID- 15667303 TI - Electron transport through nitrate and nitrite reductases in Campylobacter jejuni. AB - Campylobacter jejuni is a small genome pathogen that is incapable of growing strictly anaerobically due to its dependence on an oxygen-requiring ribonucleotide reductase for DNA synthesis. Nevertheless, it has a complex branched respiratory chain, which allows the use of several alternative electron acceptors for growth under oxygen-limited conditions. C. jejuni is equipped with both nitrate reductase (Nap) and nitrite reductase (Nrf) located in the periplasm, each predicted to receive electrons from menaquinol through distinct redox proteins. The pathways of electron transport to nitrate and nitrite are reviewed in this paper. Nitrate is considered as a potential in vivo electron acceptor and a role for nitrite reductase in NO detoxification is suggested. PMID- 15667305 TI - Transcriptional regulation of a hybrid cluster (prismane) protein. AB - HCP (hybrid-cluster protein) contains two Fe/S clusters, one of which is a hybrid [4Fe-2S-2O] cluster. Despite intensive study, its physiological function has not been reported. The Escherichia coli hcp gene is located in a two-gene operon with hcr, which encodes an NADH-dependent HCP reductase. E. coli HCP is detected after anaerobic growth with nitrate or nitrite: possible roles for it in hydroxylamine or nitric oxide reduction have been proposed. To study the regulation and role of HCP, an hcp::lacZ fusion was constructed and transformed into fnr, arcA and norR mutant strains of E. coli. Transcription from the hcp promoter was induced during anaerobic growth. Only the fnr mutant was defective in hcp expression. Nitrate- and nitrite-induced transcription from the hcp promoter was activated by the response regulator proteins NarL and NarP. Gel retardation assays were used to show that FNR (fumarate-nitrate regulation) and NarL form a complex with the hcp promoter. Transcription of the hcp-hcr operon initiates at a thymine nucleotide located 31 bp upstream of the translation-initiation codon. HCP has been overexpressed from a recombinant plasmid for physiological studies. PMID- 15667306 TI - Detoxification of nitric oxide by the flavorubredoxin of Salmonella enterica serovar Typhimurium. AB - Salmonella possesses multiple enzymes that utilize NO as a substrate, and could therefore contribute to the organism's ability to resist nitrosative killing by macrophages. Flavorubredoxin is an oxygen-sensitive enzyme that reduces NO to nitrous oxide. The Salmonella enterica serovar Typhimurium norV gene encoding flavorubredoxin was disrupted and the NO sensitivity of the mutant was determined. The norV mutant showed a greater sensitivity to NO than wild-type S. Typhimurium, but did recover growth after a transient inhibition. The mutant phenotype suggests that multiple enzymes are employed by S. Typhimurium to detoxify NO under anaerobic conditions, one of which is flavorubredoxin. PMID- 15667307 TI - Tracking nitrate reducers and denitrifiers in the environment. AB - The ability to respire nitrate when oxygen is limited has been described in taxonomically diverse microorganisms including members of the alpha-, beta-, gamma- and epsilon-proteobacteria, high and low GC Gram-positive bacteria and even Archaea. Respiratory nitrate reduction is the first step of the denitrification pathway, which is important since it is the main biological process responsible for the return of fixed nitrogen to the atmosphere, thus completing the nitrogen cycle. During the last decade, considerable knowledge has been accumulated on the biochemistry and genetics of the nitrate reductases. In this paper, we summarize the recent progress in molecular approaches for studying the ecology of the nitrate-reducing community in the environment. PMID- 15667308 TI - Complete conversion of nitrate into dinitrogen gas in co-cultures of denitrifying bacteria. AB - In the past 10 years many molecular aspects of microbial nitrate reduction have been elucidated, but the ecophysiology of this process is hardly understood. In this contribution, our efforts to study the complex microbial communities and interactions involved in the reduction of nitrate to dinitrogen gas are summarized. The initial work concentrated on emission of the greenhouse gas nitrous oxide during incomplete denitrification by Alcaligenes faecalis. As more research methods became available, the fitness of A. faecalis could be tested in mixed cultures with other denitrifying bacteria, most notably with the nitrate reducing bacterium Pseudomonas G9. Finally, the advancement of molecular diagnostic tools made it possible to survey complex microbial communities using specific primer sets for/and antibodies raised against the various NO(x) reductases. Given the enormous complexity of substrates and environmental conditions, it is evident that mixed cultures rather than single species are responsible for denitrification in man-made and natural ecosystems. However, it is surprising that even for the breakdown of a single compound, such as acetate, mixed cultures are responsible, and that the consecutive denitrification steps are commonly performed by mutualistic co-operating species. Our observations also indicate that we seldom know the identity of the major key players in the nitrogen cycle of these ecosystems. PMID- 15667309 TI - Nitrate respiration in the actinomycete Streptomyces coelicolor. AB - Streptomyces coelicolor is an obligate aerobic, filamentous soil-dwelling bacterium. Remarkably, the genome of S. coelicolor has three copies of the narGHJI operon that encodes respiratory nitrate reductase. This review summarizes our current views on the requirements for multiple nitrate reductases in S. coelicolor. PMID- 15667310 TI - Transport regulation by the serum- and glucocorticoid-inducible kinase SGK1. AB - The serum- and glucocorticoid-inducible kinase SGK1 is an ubiquitously expressed kinase with the ability to regulate a variety of transport systems. Recent observations point to a role of SGK1 in the regulation of diverse physiological functions such as epithelial transport and cardiac and neuronal excitability. At least partially through its effect on transport, SGK1 contributes to a number of pathophysiological conditions including metabolic syndrome and fibrosing disease. PMID- 15667311 TI - The concentrative nucleoside transporter family (SLC28): new roles beyond salvage? AB - The concentrative nucleoside transporter (CNT) family (SLC28) has three members: SLC28A1 (CNT1), SLC28A2 (CNT2) and SLC28A3 (CNT3). The CNT1 and CNT2 transporters are co-expressed in liver parenchymal cells and macrophages, two suitable models in which to study cell cycle progression. Despite initial observations suggesting that these transporter proteins might contribute to nucleoside salvage during proliferation, their subcellular localization and regulatory properties suggest alternative roles in cell physiology. In particular, CNT2 is a suitable candidate for modulation of purinergic responses, since it is under the control of the adenosine 1 receptor. Increasing evidence also suggests a role for CNT2 in energy metabolism, since its activation relies on the opening of ATP-sensitive K(+) channels. Animal and cell models genetically modified to alter nucleoside transporter expression levels may help to elucidate the particular roles of CNT proteins in cell physiology. PMID- 15667313 TI - Growth factors regulate cell survival by controlling nutrient transporter expression. AB - Growth factors provide permission signals that allow mammalian cells to grow, proliferate and survive. One mechanism by which growth factors maintain this control is through the regulation of cell surface nutrient transporter expression. Following growth factor withdrawal, nutrient transporters are endocytosed and degraded in the lysosome, effectively terminating the cell's ability to obtain nutrients. This results in a state of pseudostarvation in which cells atrophy and initiate a catabolic metabolic programme in the midst of abundant extracellular nutrients. Oncogenic forms of Akt can support growth factor-independent nutrient transporter expression through a mechanism that depends upon mTOR (mammalian target of rapamycin). The ability of activated Akt to support nutrient transporter expression is an essential component of its prosurvival function. When the destruction of nutrient transporters is inhibited, cells are capable of long-term growth-factor-independent cell survival in the absence of receptor-dependent signal transduction. These results imply that proteins involved in nutrient transporter turnover in response to growth factor withdrawal are components of a novel tumour suppressor pathway. Preliminary data suggest that Rab7, a GTPase required for transporter degradation, functions as a tumour suppressor protein, as inhibiting Rab7 activity promotes colony formation in soft agar. These studies indicate that drugs affecting this pathway might have utility as anti-cancer chemotherapeutic agents. PMID- 15667312 TI - Evolution of the bacterial phosphotransferase system: from carriers and enzymes to group translocators. AB - The bacterial phosphotransferase system (PTS) is a structurally and functionally complex system with a surprising evolutionary history. The substrate-recognizing protein constituents of the PTS (Enzymes II) derive from at least four independent sources. Some of the non-PTS precursor constituents have been identified, and evolutionary pathways taken have been proposed. Our analyses suggest that two of these independently evolving systems are still in transition, not yet having acquired the full-fledged characteristics of PTS Enzyme II complexes. The work described provides detailed insight into the process of catalytic protein evolution. PMID- 15667314 TI - Thyroid hormone transporters. AB - Thyroid hormone is important for development of various tissues, in particular brain, and for regulation of metabolic processes throughout life. The follicular cells of the thyroid gland produce predominantly T4 (thyroxine), but the biological activity of thyroid hormone is largely exerted by T3 (3,3',5-tri iodothyronine). The deiodinases involved in T4-to-T3 conversion or T4 and T3 degradation, as well as the T3 receptors, are located intracellularly. Therefore the action and metabolism of thyroid hormone require transport of iodothyronines across the cell membrane via specific transporters. Recently, a number of transporters capable of cellular uptake of iodothyronines have been identified. The most specific transporters identified so far are OATP1C1 and MCT8, which appear to be involved in T4 transport across the blood-brain barrier, and in T3 transport into brain neurons, respectively. The MCT8 gene is located on human chromosome Xq13, and mutations in MCT8 are associated with X-linked severe psychomotor retardation and elevated serum T3 levels. PMID- 15667315 TI - Neutral amino acid transport in epithelial cells and its malfunction in Hartnup disorder. AB - Hartnup disorder is an autosomal recessive abnormality of renal and gastrointestinal neutral amino acid transport. A corresponding transport activity has been characterized in kidney and intestinal cells and named system B(0). The failure to resorb amino acids in this disorder is thought to be compensated by a protein-rich diet. However, in combination with a poor diet and other factors, more severe symptoms can develop in Hartnup patients, including a photosensitive pellagra-like skin rash, cerebellar ataxia and other neurological symptoms. Homozygosity mapping in a Japanese family and linkage analysis on six Australian pedigrees placed the Hartnup disorder gene at a locus on chromosome 5p15. This fine mapping facilitated a candidate gene approach within the interval, which resulted in the cloning and characterization of a novel member of the sodium dependent neurotransmitter transporter family (B(0)AT1, SLC6A19) from mouse and human kidney, which shows all properties of system B(0). Flux experiments and electrophysiological recording showed that the transporter is Na(+) dependent and Cl(-) independent, electrogenic and actively transports most neutral amino acids. In situ hybridization showed strong expression in intestinal villi and in the proximal tubule of the kidney. Expression of B(0)AT1 was restricted to kidney, intestine and skin. A total of ten mutations have been identified in SLC6A19 that co-segregate with disease in the predicted recessive manner, with the majority of affected individuals being compound heterozygotes. These mutations lead to altered neutral amino acid transport function compared to the wild-type allele in vitro. One of the mutations occurs in members of the original Hartnup family described in 1956, thereby defining SLC6A19 as the 'Hartnup'-gene. PMID- 15667316 TI - Biological functions of SLC5A8, a candidate tumour suppressor. AB - SLC5A8 is a candidate tumour suppressor gene that is silenced in colon cancer, gastric cancer and possibly other cancers in humans. This gene codes for a transporter belonging to the Na(+)/glucose co-transporter gene family (SLC5). The cancer-associated silencing of the gene involves hypermethylation of CpG islands present in exon 1 of the gene. SLC5A8 is expressed in colon, ileum, kidney and thyroid gland. The protein coded by the gene mediates the Na(+)-coupled and electrogenic transport of a variety of monocarboxylates, including short-chain fatty acids, lactate and nicotinate. It may also transport iodide. The normal physiological function of this transporter in the intestinal tract and kidney is likely to facilitate the active absorption of short-chain fatty acids, lactate and nicotinate. One of the short-chain fatty acids that serves as a substrate for SLC5A8 is butyrate. This fatty acid is an inhibitor of histone deacetylases and is known to induce apoptosis in a variety of tumours including colonic tumour. Since butyrate is produced in the colonic lumen at high concentrations by bacterial fermentation of dietary fibre, we speculate that the ability of SLC5A8 to mediate the entry of this short-chain fatty acid into colonic epithelial cells underlies the potential tumour suppressor function of this transporter. PMID- 15667317 TI - ABC transporters in the balance: is there a role in multidrug resistance? AB - Drug resistance can occur at several levels and is the major cause of treatment failure in oncology. The ABC (ATP-binding cassette) transporters, beginning with the discovery of P-glycoprotein (Pgp) almost 30 years ago, have been intensively studied as potential mediators of drug resistance. Although we understand that drug resistance is almost certainly multifactorial, investigators have attempted to link anticancer drug resistance to overexpression of ABC transporters and the consequent reduction in drug accumulation. A body of evidence implicated Pgp as being important in clinical outcome; however, critical studies aimed at proving the hypothesis using Pgp inhibitors in clinical trials have to date failed. Identification of the MRP (multidrug resistance protein)/ABCC subfamily expanded the possible mechanisms of reduced drug accumulation, and the discovery of ABCG2 added a new chapter in these investigations. Correlative studies examining ABCG2 and the ABCC subfamily members in clinical drug resistance have been less avidly pursued, while basic molecular studies of structure and function have proceeded briskly. Recently, studies have focused on how single nucleotide polymorphism in multidrug transporters might affect the pharmacokinetics and pharmacodynamics of anticancer agents. These studies suggest an important role for ABC transporters in pharmacology, independent of the ultimate determination of their role in multidrug resistance. PMID- 15667318 TI - Glucose as a hormone: receptor-mediated glucose sensing in the yeast Saccharomyces cerevisiae. AB - Because glucose is the principal carbon and energy source for most cells, most organisms have evolved numerous and sophisticated mechanisms for sensing glucose and responding to it appropriately. This is especially apparent in the yeast Saccharomyces cerevisiae, where these regulatory mechanisms determine the distinctive fermentative metabolism of yeast, a lifestyle it shares with many kinds of tumour cells. Because energy generation by fermentation of glucose is inefficient, yeast cells must vigorously metabolize glucose. They do this, in part, by carefully regulating the first, rate-limiting step of glucose utilization: its transport. Yeast cells have learned how to sense the amount of glucose that is available and respond by expressing the most appropriate of its 17 glucose transporters. They do this through a signal transduction pathway that begins at the cell surface with the Snf3 and Rgt2 glucose sensors and ends in the nucleus with the Rgt1 transcription factor that regulates expression of genes encoding glucose transporters. We explain this glucose signal transduction pathway, and describe how it fits into a highly interconnected regulatory network of glucose sensing pathways that probably evolved to ensure rapid and sensitive response of the cell to changing levels of glucose. PMID- 15667319 TI - Nutrient sensing systems for rapid activation of the protein kinase A pathway in yeast. AB - The cAMP-protein kinase A (PKA) pathway in the yeast Saccharomyces cerevisiae controls a variety of properties that depend on the nutrient composition of the medium. High activity of the pathway occurs in the presence of rapidly fermented sugars like glucose or sucrose, but only as long as growth is maintained. Growth arrest of fermenting cells or growth on a respiratory carbon source, like glycerol or ethanol, is associated with low activity of the PKA pathway. We have studied how different nutrients trigger rapid activation of the pathway. Glucose and sucrose activate cAMP synthesis through a G-protein-coupled receptor system, consisting of the GPCR Gpr1, the Galpha protein Gpa2 and its RGS protein Rgs2. Glucose is also sensed intracellularly through its phosphorylation. Specific mutations in Gpr1 abolish glucose but not sucrose signalling. Activation of the PKA pathway by addition of a nitrogen source or phosphate to nitrogen- or phosphate-starved cells, respectively, is not mediated by an increase in cAMP. Activation by amino acids is triggered by the general amino acid permease Gap1, which functions as a transporter/receptor. Short truncation of the C-terminus results in constitutively activating alleles. Activation by ammonium uses the ammonium permeases Mep1 and Mep2 as receptor. Specific point mutations in Mep2 uncouple signalling from transport. Activation by phosphate is triggered a.o. by the Pho84 phosphate permease. Several mutations in Pho84 separating transport and signalling or triggering constitutive activation have been obtained. PMID- 15667320 TI - Glucose sensing via the protein kinase A pathway in Schizosaccharomyces pombe. AB - The fission yeast Schizosaccharomyces pombe primarily detects glucose via a cAMP signalling pathway. Components of this pathway include the Git3 G-protein-coupled receptor and a heterotrimeric G-protein, from which the Gpa2 Galpha subunit activates adenylate cyclase (Git2/Cyr1). Three additional proteins, Git1, Git7 and Git10 are required to generate a cAMP response even in a strain expressing an activated form of Gpa2, which is capable of bypassing the loss of the GPCR and Gbetagamma dimer. Therefore, Git1, Git7 and Git10 either act in a G-protein independent manner or are required to stabilize or assemble a functional signalling complex. Although prior data suggested that the Cgs2 cAMP phosphodiesterase (PDE) does not regulate the cAMP response, we now have evidence that along with adenylate cyclase regulation, PDE activation is important for limiting the response to glucose. Finally, regulation of protein kinase A activation appears to involve both traditional post-translational regulation of the function of the components of the cAMP pathway and glucose-dependent transcriptional regulation of some of these cAMP pathway genes. PMID- 15667321 TI - Amino acid sensing by Ssy1. AB - Saccharomyces cerevisiae senses extracellular amino acids using two members of the family of amino acid transporters, Gap1 or Ssy1; aspects of the latter are reviewed here. Despite resemblance with bona fide transporters, Ssy1 appears unable to facilitate transport. Exposure of yeast to amino acids results in Ssy1 dependent transcriptional induction of several genes, in particular some encoding amino acid transporters. Amino acids differ strongly in their potency, leucine being the most potent one known. Using a selection system in which potassium uptake was made dependent on amino acid signalling, our laboratory has obtained and described gain-of-function mutations in SSY1. Some alleles conferred inducer independent signalling; others increased apparent affinity for inducers. These results revealed that amino acid transport is not required for signalling and support the notion that sensing by Ssy1 occurs via its direct interaction with extracellular amino acids. Current work includes development of quantitative assays of sensing. We use the finding by Per Ljungdahl's laboratory that the signal transduction from Ssy1 involves proteolytic removal of an inhibitory part of the transcriptional activator Stp1. Protein-A Z-domain fused to the C-terminus of Stp1 and Western analysis using antibody against horseradish peroxidase allow quantification of sensing. PMID- 15667322 TI - Glucose sensing through the Hxk2-dependent signalling pathway. AB - In this work, we describe the hexokinase 2 (Hxk2) signalling pathway within the yeast cell. Hxk2 and Mig1 are the two major factors of glucose repression in Saccharomyces cerevisiae. The functions of both proteins have been extensively studied but there is no information about possible interactions among them in the repression pathway. Our results demonstrate that Hxk2 interacts directly with Mig1 in vivo and in vitro and that the ten amino acids motif between K6 and M15 is required for their interaction. This interaction has been detected at the DNA level both in vivo by chromatin immunoprecipitation experiments and in vitro using purified proteins and a DNA fragment containing the MIG1 site of the SUC2 promoter. This demonstrates that the interaction is of physiological relevance. Our findings show that the main role of Hxk2 in the glucose signalling pathway is the interaction with Mig1 to generate a repressor complex located in the nucleus of S. cerevisiae. PMID- 15667323 TI - Sugar sensing and signalling networks in plants. AB - Plant sugar signalling operates in a complex network with plant-specific hormone signalling pathways. Hexokinase was identified as an evolutionarily conserved glucose sensor that integrates light, hormone and nutrient signalling to control plant growth and development. PMID- 15667324 TI - Sucrose-induced translational repression of plant bZIP-type transcription factors. AB - Sugars as signalling molecules exert control on the transcription of many plant genes. Sugar signals also alter mRNA and protein stability. Increased sucrose concentrations specifically repress translation of the S-class basic region leucine zipper (bZIP) type transcription factor AtbZIP11/ATB2. This sucrose induced repression of translation (SIRT) depends on translation of a highly conserved upstream open reading frame (uORF) in the 5' UTR of the gene. This conserved uORF is exclusively encoded in 5' UTRs of several plant S-class bZIP transcription factors. Arabidopsis homologues of ATB2/AtbZIP11, which harbour the conserved uORF, also show SIRT. Therefore, SIRT emerges as a general sucrose translational control mechanism of a group of transcription factors. SIRT might be part of a sucrose-specific signalling pathway, controlling expression of plant bZIP transcription factor genes. PMID- 15667326 TI - The role of trehalose-6-phosphate synthase in Arabidopsis embryo development. AB - We previously showed that trehalose-6-phosphate synthase 1 (TPS1), which catalyses the first step in trehalose synthesis, is essential for embryo maturation in Arabidopsis. The tps1 mutant embryos develop more slowly than wild type. Patterning in the tps1 embryos appears normal but they do not progress past the torpedo stage to cotyledon stage, which is when storage reserves start to accumulate in the expanding cotyledons. Our initial data led to the hypothesis that trehalose metabolism plays a key role in regulating storage reserve accumulation by allowing the embryo to respond to the dramatic increase in sucrose levels that occurs at the torpedo stage of embryo development. More recent data demonstrate that while the tps1 mutant is blocked in the developmental progression of embryos from torpedo to cotyledon stage the expression of genes involved in the accumulation of storage reserves proceeds in a similar fashion to wild type. Thus it appears that induction of metabolic processes required for accumulation of storage reserves in tps1 occurs independently of the developmental stage and instead follows a temporal programme similar to wild-type seeds in the same silique. PMID- 15667325 TI - Trehalose metabolism and glucose sensing in plants. AB - Plants sense and respond to changes in carbon and nitrogen metabolites during development and growth according to the internal needs of their metabolism. Sugar sensing allows plants to switch off photosynthesis when carbohydrates are abundant. These processes involve regulation of gene and protein activity to allow plants the efficient use of energy storage. Besides being a key element in carbon metabolism, glucose (Glc) has unravelled as a primary messenger in signal transduction. It has been proved that hexokinase (HXK) is a Glc sensor. An unusual disaccharide named trehalose is present in very low levels in most plants except for the desiccation-tolerant plants known as 'resurrection' plants where trehalose functions as an osmoprotectant. We have shown that overexpression of the Arabidopsis trehalose-6-phosphate synthase gene (AtTPS1) in Arabidopsis promotes trehalose and trehalose-6-phosphate (T6P) accumulation. Seedlings expressing AtTPS1 displayed a Glc-insensitive phenotype. Transgenic lines germinated normally on Glc, in contrast to wild-type seedlings showing growth retardation and absence of chlorophyll and root elongation. Gene-expression analysis in transgenic plants showed up-regulation of several genes involved in sugar signalling and metabolism. These data suggest that AtTPS1 and accordingly T6P and trehalose play an important role in the regulation of Glc sensing and signalling genes during plant development. PMID- 15667327 TI - Nitrate and glutamate sensing by plant roots. AB - The architecture of a root system plays a major role in determining how efficiently a plant can capture water and nutrients from the soil. Growth occurs at the root tips and the process of exploring the soil volume depends on the behaviour of large numbers of individual root tips at different orders of branching. Each root tip is equipped with a battery of sensory mechanisms that enable it to respond to a range of environmental signals, including nutrients, water potential, light, gravity and touch. We have previously identified a MADS (MCM1, agamous, deficiens and SRF) box gene (ANR1) in Arabidopsis thaliana that is involved in modulating the rate of lateral root growth in response to changes in the external NO(3)(-) supply. Transgenic plants have been generated in which a constitutively expressed ANR1 protein can be post-translationally activated by treatment with dexamethasone (DEX). When roots of these lines are treated with DEX, lateral root growth is markedly stimulated but there is no effect on primary root growth, suggesting that one or more components of the regulatory pathway that operate in conjunction with ANR1 in lateral roots may be absent in the primary root tip. We have recently observed some very specific effects of low concentrations of glutamate on root growth, resulting in significant changes in root architecture. Experimental evidence suggests that this response involves the sensing of extracellular glutamate by root tip cells. We are currently investigating the possible role of plant ionotropic glutamate receptors in this sensory mechanism. PMID- 15667328 TI - Development and use of fluorescent nanosensors for metabolite imaging in living cells. AB - To understand metabolic networks, fluxes and regulation, it is crucial to be able to determine the cellular and subcellular levels of metabolites. Methods such as PET and NMR imaging have provided us with the possibility of studying metabolic processes in living organisms. However, at present these technologies do not permit measuring at the subcellular level. The cameleon, a fluorescence resonance energy transfer (FRET)-based nanosensor uses the ability of the calcium-bound form of calmodulin to interact with calmodulin binding polypeptides to turn the corresponding dramatic conformational change into a change in resonance energy transfer between two fluorescent proteins attached to the fusion protein. The cameleon and its derivatives were successfully used to follow calcium changes in real time not only in isolated cells, but also in living organisms. To provide a set of tools for real-time measurements of metabolite levels with subcellular resolution, protein-based nanosensors for various metabolites were developed. The metabolite nanosensors consist of two variants of the green fluorescent protein fused to bacterial periplasmic binding proteins. Different from the cameleon, a conformational change in the binding protein is directly detected as a change in FRET efficiency. The prototypes are able to detect various carbohydrates such as ribose, glucose and maltose as purified proteins in vitro. The nanosensors can be expressed in yeast and in mammalian cell cultures and were used to determine carbohydrate homeostasis in living cells with subcellular resolution. One future goal is to expand the set of sensors to cover a wider spectrum of metabolites by using the natural spectrum of bacterial periplasmic binding proteins and by computational design of the binding pockets of the prototype sensors. PMID- 15667329 TI - Carbon source induced yeast-to-hypha transition in Candida albicans is dependent on the presence of amino acids and on the G-protein-coupled receptor Gpr1. AB - Yeast-to-hypha transition in Candida albicans can be induced by a wide variety of factors, including specific nutrients. We have started to investigate the mechanism by which some of these nutrients may be sensed. The G-protein-coupled receptor Gpr1 is required for yeast-to-hypha transition on various solid hypha inducing media. Recently we have shown induction of Gpr1 internalization by specific amino acids, e.g. methionine. This suggests a possible role for methionine as a ligand of CaGpr1. Here we show that there is a big variation in methionine-induced hypha formation depending on the type of carbon source present in the medium. In addition high glucose concentrations repress hypha formation whereas a concentration of 0.1%, which mimics the glucose concentration present in the bloodstream, results in maximal hypha formation. Hence, it remains unclear whether Gpr1 senses sugars, as in Saccharomyces cerevisiae, or specific amino acids like methionine. PMID- 15667330 TI - Glucose modulation of cell size in yeast. AB - Saccharomyces cerevisiae cells grown in glucose have larger average size than cells grown in ethanol. Besides, yeast must reach a carbon source-modulated critical cell size in order to enter S phase at Start. This control is of outmost physiological relevance, since it allows us to coordinate cell growth with cell cycle progression and it is responsible for cell size homeostasis. The cell sizer mechanism requires the overcoming of two sequential thresholds, involving Cln3 and Far1, and Clb5,6 and Sic1, respectively. When both thresholds are non functional, carbon source modulation of cell size at Start is completely abolished. Since inactivation of extracellular glucose sensing through deletion of either the GPR1 or the GPA2 gene causes a marked, but partial, reduction in the ability to modulate cell size and protein content at Start, it is proposed that both extracellular and intracellular glucose signalling is required for properly setting the cell sizer in glucose media. PMID- 15667331 TI - Acquired glucose sensitivity of k-ras transformed fibroblasts. AB - Mutational activation of the ras gene is critical for the onset of different malignant phenotypes. We constructed a dominant negative mutant (GEF-DN) of a Ras activator protein (guanine nucleotide-exchange factor) that upon over-expression in k-ras transformed NIH 3T3 fibroblasts strongly reduces intracellular Ras*GTP, reverting these cells to wild-type phenotype for morphology, anchorage independent growth and reduction of tumour formation in nude mice. Here we review evidence showing that the enhanced proliferation potential of NIH-ras cells requires high initial glucose concentration in the medium and sustained Ras pathway activation. The exquisite sensitivity of NIH-ras fibroblasts to a shortage in nutrient and energy supply highlights an acquired fragility of cancer cells that may be exploited for therapeutic purposes. PMID- 15667332 TI - Nutrient sensing in pancreatic beta cells suppresses mitochondrial superoxide generation and its contribution to apoptosis. AB - Excessively high glucose concentrations have been shown to damage tissues through stimulation of mitochondrial superoxide generation. This effect has therefore been considered as a potential cause for dysfunction and death of pancreatic beta cells in diabetes. We have examined whether the rate of glucose metabolism in isolated rat beta cells is correlated with their formation of oxygen radicals. It was found that high rates of glucose metabolism did not stimulate the formation of superoxide and H(2)O(2) but suppressed it. The higher rates of superoxide production in beta cells with lower mitochondrial metabolic activity contributed to the susceptibility of these cells to apoptosis. PMID- 15667333 TI - Expression of sweet taste receptors of the T1R family in the intestinal tract and enteroendocrine cells. AB - The composition of the intestinal luminal content varies considerably with diet. It is important therefore that the intestinal epithelium senses and responds to these significant changes and regulates its functions accordingly. Although it is becoming evident that the gut epithelium senses and responds to luminal nutrients, little is known about the nature of the nutrient sensing molecule and the downstream cellular events. A prototype example is the modulation in the capacity of the gut to absorb monosaccharides via the intestinal luminal membrane Na(+)/glucose cotransporter, SGLT1. The experimental evidence suggests that luminal sugar is sensed by a glucose sensor residing on the luminal membrane of the gut epithelium and linked to a G-protein-coupled receptor, cAMP/PKA (protein kinase A) pathway, resulting ultimately in modulation of intestinal monosaccharide absorption. Here we report the expression, at mRNA and protein levels, of members of the T1R sweet taste receptors, and the alpha-subunit of the G-protein gustducin, in the small intestine and the enteroendocrine cell line, STC-1. In the small intestine, there is a highly coordinated expression of sweet taste receptors and gustducin, a G-protein implicated in intracellular taste signal transduction, throughout the gut. The potential involvement of these receptors in sugar sensing in the intestine will facilitate our understanding of intestinal nutrient sensing, with implications for better nutrition and health maintenance. PMID- 15667335 TI - CD36-dependent fatty acid uptake regulates expression of peroxisome proliferator activated receptors. AB - CD36 is an important regulator of lipid metabolism in vivo due to its role in the facilitated uptake of long-chain FAs (fatty acids). CD36-deficient mice display reduced TAG (triacylglycerol) in muscle, but elevated hepatic TAG. Also, insulin sensitivity is enhanced peripherally, while it appears impaired in the liver. Tissues such as muscle, which normally express high levels of CD36, shift to high glucose utilization in CD36 deficiency, so we hypothesized that this shift must involve adaptive changes in the PPAR (peroxisome-proliferator-activated receptor) transcription factors which regulate FA metabolism. To test this, we examined mRNA levels for the three PPAR isoforms in tissues of WT (wild-type) and CD36 deficient mice following the administration of saline, glucose or olive oil by intragastric gavage. Compared with WT mice, CD36-null mice had 5-10-fold increased PPAR mRNA in adipose tissue in the basal state, and did not exhibit diet-induced changes. Correlations between adipose PPAR mRNA abundance and plasma lipids were observed in WT mice, but not in CD36-null mice. The opposite was true for hepatic PPAR mRNA levels, which correlated with plasma FA, TAG and/or glucose only in CD36-null mice. No significant differences were observed in PPAR mRNA levels in the intestine, where CD36 does not impact on FA uptake. The data suggest that CD36 and the PPARs are components of the FA-sensing machinery to respond to changes in FA flux in a tissue-specific manner. PMID- 15667334 TI - Glucokinase and glucose homeostasis: proven concepts and new ideas. AB - The enzyme GK (glucokinase), which phosphorylates glucose to form glucose 6 phosphate, serves as the glucose sensor of insulin-producing beta-cells. GK has thermodynamic, kinetic, regulatory and molecular genetic characteristics that are ideal for its glucose sensor function and allow it to control glycolytic flux of the beta-cells as indicated by control-, elasticity- and response-coefficients close to or larger than 1.0. GK operates in tandem with the K(+) and Ca(2+) channels of the beta-cell membrane, resulting in a threshold for glucose stimulated insulin release of approx. 5 mM, which is the set point of glucose homoeostasis for most laboratory animals and humans. Point mutations of GK cause 'glucokinase disease' in humans, which includes hypo- and hyper-glycaemia syndromes resulting from activating or inactivating mutations respectively. GK is allosterically activated by pharmacological agents (called GK activators), which lower blood glucose in normal animals and animal models of T2DM. On the basis of crystallographic studies that identified a ligand-free 'super-open' and a liganded closed structure of GK, on thermostability studies using glucose or mannoheptulose as ligands and studies showing that mannoheptulose alone or combined with GK activators induces expression of GK in pancreatic islets and partially preserves insulin secretory competency, a new hypothesis was developed that GK may function as a metabolic switch per se without involvement of enhanced glucose metabolism. Current research has the goal to find molecular targets of this putative 'GK-switch'. The case of GK research illustrates how basic science may culminate in therapeutic advances of human medicine. PMID- 15667336 TI - The calcium-sensing receptor as a nutrient sensor. AB - Critical to cell fate in many cell types is the ability to sense and respond to acute changes in free ionized extracellular calcium concentration ([Ca(2+)](o)). Such tight control is mediated by the activation of a protein known as the extracellular-calcium-sensing receptor (CaR). CaR belongs to the 'family C' of G protein-coupled receptors and was the first G-protein-coupled receptor to be identified to have an inorganic cation, calcium, as its ligand. While calcium is the physiological agonist of the receptor, several other polyvalent cations and polycations can also modulate CaR function as do certain L-aromatic amino acids, polyamines, salinity and pH. This feature renders the CaR uniquely capable of generating cell- and tissue-specific responses, and of integrating inputs deriving from changes in the Ca(2+)(o) concentration with signals deriving from the local metabolic environment. Here we address the role of the CaR in physiology and disease, the range of CaR modulators and the potential roles of the CaR as a metabolic sensor in a variety of physiological (and pathological) scenarios. PMID- 15667337 TI - Angiotensin II modulates frizzled-2 receptor expression in rat vascular smooth muscle cells. AB - Ang II (angiotensin II) has multiple effects on vascular smooth muscle cells through the modulation of different classes of genes. Using the mRNA differential display method to investigate gene expression in rat aortic smooth muscle cells in culture in response to 3 h of Ang II stimulation, we observed that Ang II down regulated the expression of a member of the family of transmembrane receptors for Wnt proteins that was identified as Fzd2 [Fzd (frizzled)-2 receptor]. Fzds are a class of highly conserved genes playing a fundamental role in the developmental processes. In vitro, time course experiments demonstrated that Ang II induced a significant increase (P<0.05) in Fzd2 expression after 30 min, whereas it caused a significant decrease (P<0.05) in Fzd2 expression at 3 h. A similar rapid up regulation after Ang II stimulation for 30 min was evident for TGFbeta1 (transforming growth factor beta1; P<0.05). To investigate whether Ang II also modulated Fzd2 expression in vivo, exogenous Ang II was administered to Sprague Dawley rats (200 ng.kg(-1) of body weight.min(-1); subcutaneously) for 1 and 4 weeks. Control rats received normal saline. After treatment, systolic blood pressure was significantly higher (P<0.01), whereas plasma renin activity was suppressed (P<0.01) in Ang II- compared with the saline-treated rats. Ang II administration for 1 week did not modify Fzd2 expression in aorta of Ang II treated rats, whereas Ang II administration for 4 weeks increased Fzd2 mRNA expression (P<0.05) in the tunica media of the aorta, resulting in a positive immunostaining for fibronectin at this time point. In conclusion, our data demonstrate that Ang II modulates Fzd2 expression in aortic smooth muscle cells both in vitro and in vivo. PMID- 15667338 TI - Expression of human IL-1alpha after intramarrow gene transfer into healthy non human primate by adenoviral vector. AB - Interleukin-1alpha (IL-1alpha) is a multifunctional cytokine that stimulates myelopoiesis in macaque. However, daily systemic injections of IL-1alpha are associated with severe side effects. We therefore investigated the feasibility of a gene therapy strategy aimed at increasing the IL-1alpha local production in bone marrow with limited release of the vector into the blood circulation. Intra medullar administration of adenoviral vector containing human IL-1alpha (huIL 1alpha) gene resulted in enhanced neutrophil, monocyte and platelet counts during the two first weeks after injection. The DNA vector, the transgene expression and the huIL-1alpha production was detected in treated bone marrow without significant detection of huIL-1alpha in the peripheral blood. Associated with huIL-1alpha production, we observed concomitant plasma C reactive protein and IL 1Ra peaks in the acellular fraction of treated bone marrow at days 3 and 7. No abnormal clinical side effects were observed in any of the animals following the adenoviral vector injection. PMID- 15667339 TI - Herpes B-virus seroreactivity in a colony of Macaca mulatta: data from the Sabana Seca Field Station, a new specific pathogen-free program. AB - The demand for B-virus-free animals for biomedical research is increasing, while at the same time the availability of such animals is decreasing. The establishment of Specific Pathogen-Free (SPF) breeding macaque colonies is a priority of the National Institutes of Health. Nevertheless, it is well known that seroreactivity to B-virus can be difficult to interpret, particularly as it can vary over time in a single animal. The aim of the present study was to implement a reliable algorithm to examine B-virus reactivity among the rhesus monkey population of the Caribbean Primate Research Center. The sensitivity and specificity of our assay were determined using reports from two different laboratories as references. Whereas 95.4% of animals showed consistent serological status and 4.6% of animals recruited to this SPF program showed serovariability to B-virus over the initial 2 years of examination. Implications for all SPF programs are discussed in this article. PMID- 15667340 TI - The significance of platelet-activating factor and fertility in the male primate: a review. AB - Since its discovery nearly 30 years ago platelet-activating factor (PAF) has emerged as one of the more important lipid mediators known. PAF (1-O-alkyl-2-O acetyl-sn-glycero-3-phosphorylcholine) exists endogenously as a mixture of molecular species with structural variants of the alkyl moiety. PAF is a novel potent signaling phospholipid that has unique pleiotropic biological properties in addition to platelet activation. PAF also plays a significant role in reproduction and is present in the sperm of a number of primate species. PAF content in squirrel monkey sperm is significantly higher during the breeding season than the non-breeding season. PAF content in rhesus sperm has a significant relationship with sperm motility. PAF content in human sperm has a positive correlation with seminal parameters and pregnancy outcomes. The enzymes (lyso-PAF-acetyltransferase and PAF-acetylhydrolase) necessary for PAF activation and deactivation are present in primate sperm. PAF-acetylhydrolase may act as a "decapacitation factor". Removal of this enzyme during capacitation promotes PAF synthesis increasing primate motility and fertilization. PAF also plays a significant role in the fertilization process, enhancing the fertilization rates of oocytes. Enhanced embryo development has also been reported in oocytes fertilized with PAF-treated sperm. Exogenous PAF will also significantly improve primate artificial insemination pregnancy outcomes. PAF antagonists inhibit sperm motility, acrosome reaction, and fertilization thus suggesting the presence of receptors for PAF. The PAF-receptor is present on primate sperm, with altered transcript levels and distribution patterns on abnormal cells. Whereas, the exact mechanism of PAF in sperm function and reproduction is uncertain, its importance in normal primate fertility is substantial. PMID- 15667341 TI - Contraception in the chimpanzee: 12-year experience at the CIRMF Primate Centre, Gabon. AB - The Primate Centre of the International Center for Medical Research in Franceville, Gabon (CIRMF) has had a chimpanzee (Pan troglodytes) colony for more than 20 years. A contraceptive policy was started in 1990, following a rapid increase in the number of individuals in the 1980s. Intrauterine devices were first used in 24 females; 10 failures occurred over a period of about 4 years. Depo-provera was then used in 28 chimpanzees for between 4 months and 4 years; 10 failures occurred, the animals gained weight, and cyclic swelling of the external genitalia was markedly modified. Starting in late 2000, 25 females received progestin implants; only one failure has so far occurred, and the main side effect is a complete abrogation of cyclic sexual swelling. PMID- 15667342 TI - Normal hematological indices, blood chemistry and histology and ultrastructure of pancreatic islets in the wild Indian bonnet monkeys (Macaca radiata radiata). AB - The present study is aimed at determining some haematological and biochemical parameters in the wild Indian bonnet monkeys as also the microscopic and ultrastructural characteristics of their pancreatic islets. Adult wild Indian bonnet monkeys (Macaca radiata radiata) of both sexes weighing between 2.5 and 4 kg were used in these experiments. Their platelet, reticulocyte and total leukocyte counts and the blood concentrations of hemoglobin and plasma proteins and the serum concentrations of aspartate amino transferase, alanine amino transferase and calcium are similar to the values reported for M. mulatta. Plasma glucose is lower when compared with reported values of M. mulatta and M. fascicularis. Insulin levels are comparable with those of M. mulatta and M. nigra. Histology of islets is similar to that of humans. Ovoid cell collections of islet cells are scattered throughout the pancreas. Ultrastructure of A, B and D cells is similar to humans. These findings suggest that this relatively underutilized macaques may be a suitable model for biomedical research. PMID- 15667343 TI - Congenital bronchiolo-alveolar airway malformation in a cynomolgus macaque (Macaca fascicularis). AB - Pulmonary congenital anomalies in animals are rare. Previously reported malformations include accessory lung formation, pulmonary hypoplasia, pulmonary agenesis, and various forms of hamartoma. Congenital bronchiolo-alveolar airway malformation, a new entity, is described in a 1-day-old male cynomolgus macaque. This neonate experienced breathing difficulties shortly after birth and died while therapy was being administered. Grossly, the right lung was markedly increased in size, firm, and pink. Histopathologically, sections of right lung showed irregular bronchiole-like and alveolus-like structures. There was marked widening of alveolar septae by loosely arranged mesenchymal cells and many centrally located capillaries. Alveoli were lined by cuboidal epithelial cells. There were scattered islands of immature cartilage. A grossly enlarged lung containing bronchiole-like and alveolus-like structures, immature cartilage islands, and many capillaries within alveolar septae on histopathologic examination, is inconsistent with previously described congenital pulmonary anomalies in animals and humans. PMID- 15667344 TI - Fatal inflammatory heart disease in a bonobo (Pan paniscus). AB - We report the first probable identification of encephalomyocarditis virus (EMCV) in a bonobo (Pan paniscus) that had been part of a forest re-introduction programme. Clinical presentation was of episodic acute on chronic heart failure and cerebral infarction with end-stage renal failure rather than sudden death which is more commonly associated with EMCV infection. A postmortem diagnosis of probable EMCV was made using gross pathological and histopathological examination. Findings included acute on chronic heart failure combined with the unusual but characteristic histopathological features of non-suppurative necrotizing myocarditis with mononuclear, inflammatory infiltration of the brain. PMID- 15667346 TI - Trajectories of physical aggression from toddlerhood to middle childhood: predictors, correlates, and outcomes. PMID- 15667354 TI - The stability of young children's physical aggression: relations with child care, gender, and aggression subtypes. PMID- 15667355 TI - [Editorial. ]. PMID- 15667356 TI - [The significance of Candida serology based on cell wall antigens]. AB - The article reviews the state of the art in detecting anti-Candida antibodies directed against cell wall mannans. Experience in this field was collected over more than four decades. The mannan antigens of Candida spp. are well characterized serologically and biochemically. Candida mannans are immunogenically potent antigens to which the host's immune system reacts regularly. The human host is not immunologically naive towards Candida albicans. Clinical and laboratory evaluation of methodologies detecting anti-Candida antibodies must essentially regard the pathogenetic particularities of candidaemia and candidosis with their opportunistic characters. PMID- 15667357 TI - [Diagnostic value of antibody detection against Candida germ tube antigens in a patient with hepatosplenic candidosis]. AB - In the case of a 53-year-old woman with acute myeloid leukaemia and fever in late aplasia after chemotherapy, invasive mycosis with characteristic involvement of liver and spleen was diagnosed. For the serological identification of Candida in the early phase of the infection, methods for the detection of antibodies against Candida antigens were compared. By ELISA-based detection of IgM and IgG antibodies against a mixture of Candida antigens (ESR 117G and 117M, Virion Serion, Wurzburg, Germany) evidence for invasive candidosis was obtained significantly earlier (22 days) when compared with the immunofluorescence detection of IgG antibodies against Candida albicans germ tube antigens (Vircell, Granada, Spain). In the case of this patient, the detection of a humoral response against Candida germ tube antigens was of little diagnostic value. PMID- 15667358 TI - [Mycoserologic tests in surgical intensive care patients and patients with Candida oesophagitis]. AB - We retrospectively analysed the results of Candida serological testing of 126 surgical intensive care patients. There were high numbers of patients which were mycoserologically positive (haemagglutination test HAT 86.4%, immunoglobulin subclasses 85.3%). Positive microbiological cultures of Candida spp. were found in 43.7% of the patients. Patients with positive cultural findings had an increased HAT titre in 83.9% and increased immunoglobulins in 74.2%. These patients were suspected to have a Candida infection. The correlation to higher titres was much closer in patients with an invasive Candida infection like candidaemia and oesophagitis. PMID- 15667359 TI - [Candida-specific antibodies in intensive care and non-intensive care patients]. AB - The titres of indirect anti-Candida haemagglutination test (C-HAT) and specific immunoglobulins C-IgM, C-IgG and C-IgA in 328 intensive care (IT) patients and 166 non-intensive care (NIT) patients were compared by statistical test methods. Positive correlations were found between values of C-HAT and all three Candida specific immunoglobulins. At an interval of < or =7 days in the first patient group only, the changes in the course of titres were statistically highly significant. For C-IgM the short-term increase of titres can be interpreted in the sense of happened mycotic infection in connection with the clinical picture. In IT patients we found an average increase of C-IgM at 259.3, C-IgG at 174.7, C IgA at 59.8 U ml(-1) and of three titre steps in C-HAT. In our experience of diagnostics of Candida infections in intensive care area with non-neutropenic patients the short-term determination of the anti-Candida immunoglobulin subclasses C-IgM and C-IgG and of C-HAT as screening test should not be neglected. PMID- 15667360 TI - [Performance of the Candida mannan antigen detection in patients with fungemia]. AB - For several years, the Platelia Candida mannan antigen enzyme immunoassay (Candida EIA) has been commercially available as a diagnostic test for invasive candidosis. We evaluated the Candida EIA with patients with proven fungemia caused by yeasts from which at least one serum sample was available. Fifty-nine patients with 121 serum samples were included in the study. Sixty-one different yeast strains were isolated from positive blood-cultures. The Candida EIA was positive (n = 35) or borderline positive (n = 8) in 43 of 59 patients with fungemia, resulting in an overall sensitivity of 73%. For the different yeast species, the following sensitivities were calculated: Candida albicans 30 of 39 (77%), Candida glabrata 7 of 11 (64%), Candida parapsilosis 1 of 3, Candida tropicalis 2 of 2, Candida kefyr 2 of 2, Candida lipolytica 0 of 1, Candida lusitaniae 1 of 1, Candida krusei 1 borderline positive of 1, Saccharomyces cerevisiae 1 of 1. In six patients the antigen levels over time were evaluable. In three cases the antigen was positive 3-4 days before the day the blood culture was drawn, in one case on the same day, and in two cases 2 and 5 days afterwards. In conclusion, the Candida EIA was suitable for the detection of fungemia due to the major facultatively pathogenic yeast species. The test was positive in about half of the patients before blood cultures became positive. In these cases, it contributed to an early diagnosis of invasive candidiasis. PMID- 15667361 TI - [Experience with the Platelia Candida ELISA for the diagnostics of invasive candidosis in neutropenic patients]. AB - Invasive Candida infections (IC) belong to the most important opportunistic fungal infections in immunocompromised patients. IC is difficult to diagnose, because clinical symptoms are nonspecific and cultural methods lack sensitivity or specificity. We evaluated the Platelia Candida enzyme immunoassay (Candida EIA) for the diagnosis of IC in patients with haematological malignancies. A total of 62 neutropenic patients with 469 serum samples were included in the study. Candida colonization was monitored by weekly cultures of mouth washings, urine, and stool samples. Yeasts were grown from samples of 42 patients (68%), mainly Candida albicans (50%), followed by Candida glabrata (23%) and Candida krusei (20%). According to the criteria of the EORTC/NIH, the patients were categorized: (1) 3 patients with proven IC; (2) 6 patients with probable IC; (3) 34 patients colonized with Candida; (4) 19 patients without Candida colonization and without IC. In the patient categories (1) to (4), 3/3 (100%), 3/6 (50%), 20/34 (59%), and 7/19 (37%) patients were Candida EIA positive (>0.5 ng/ml) in at least one serum sample. The sensitivity of the assay for the detection of proven IC was 100%, for proven and probable IC 67%, the specificity was 49% for both groups. An increase of the cut-off value to 2.0 ng/ml raised the specificity to 61%, but lowered the sensitivity to 56%. In conclusion, the Platelia Candida EIA does not discriminate between Candida colonization and probable invasive infection in haematological patients. PMID- 15667362 TI - [Diagnosis of systemic Candida infections. Evaluation of serology, molecular biology and D-arabinitol detection]. AB - In order to diagnose systemic Candida infection in ICU patients, several methods were compared. In the first round, antigens and antibodies of 104 patients were investigated. Seventeen patients were infected; in nine of these patients a positive antigen was detected; elevated antibodies were detected in 11 patients. Only 42 patients were colonized. In this group, one positive antigen and six elevated antibody titres were detected. Combining these results, the sensitivity of the antigen only (58.5%) or the antibodies only (52.9%) increased to 100%. In the second round (n = 83 patients) the D-arabinitol/L-arabinitol quotient was also determined in the urine. 18.1% of the patients had elevated antibodies and 26.5% elevated D-arabinitol/L-arabinitol quotients. In the third round, PCR was also applied (n = 27 patients). Five patients had elevated D-arabinitol/L arabinitol quotients and one of these had a positive PCR result. In conclusion, serological methods (antigen and antibody detection) should only be applied in strictly selected patients. In uncertain cases, the addition of PCR or determination of D-arabinitol/L-arabinitol might be helpful. PMID- 15667363 TI - [The significance of interleukins and of Candida-IgE in chronic recurrent vulvovaginal candidosis]. AB - We examined 104 patients with chronic recurrent vulvovaginal candidosis; 41 healthy women were selected for the control group. Vaginal samples were taken, and yeasts were grown and tested for Candida strains: 29.8% of samples contained Candida spp.; two of the control women were infected. We also identified interleukin IL-4, 5 and 13, but there was only significant increase in IL-4. In addition, prostaglandin E2, whole IgE and Candida-specific-IgE was identified. Also here prostaglandin E2 and the Candida IgE were significantly higher in comparison with the control group, while whole IgE showed no significant increase. This resulted in an allergic component in the chronic recurrent vulvovaginal candidosis, which suggested that therapy should be reconsidered. PMID- 15667364 TI - [Candida- and Aspergillus-antibody-Elisa--expected positive antibody results in sera of the clinical-mycological routine diagnostics]. AB - Based on the ELISA results of more than 15000 serum samples of clinical mycological routine diagnostics, the expected frequency of positive antibody ELISA results within the immunoglobulin classes IgM, IgG and IgA was determined, to optimize the diagnostic assessment of first or single result of Candida or Aspergillus antibody ELISA. In general diagnostics the expected frequency of positive antibody ELISA results of the first sample within the immunoglobulin classes were as follows: Candida antibody IgM 6.1%; IgG 6.0%; IgA 2.1% and Aspergillus antibody IgM 11.4%; IgG 22.1% and IgA 5.1%, respectively. Using the Candida antibody ELISA as confirmation test only, percentages of positive antibody results in the first sample were 2.5 to 3 times higher than in general diagnostics. In follow-up examinations the Candida antibodies showed different kinetics within the immunoglobulin classes compared to those of the Aspergillus antibodies. PMID- 15667365 TI - [Antigen detection in invasive aspergillosis and candidosis: the Austrian experience]. AB - In order to assess the performance of different methods for the detection of fungal antigens, data of five Austrian hospitals were evaluated. The enzyme immunoassay (EIA) Platelia Aspergillus Antigen (Bio-Rad, USA) was used for the diagnosis of invasive aspergillosis and compared with clinical data. It could be shown that it is more effective to investigate at least two sequential sera than only one sole serum. For several high-risk patients diagnosis could be improved or confirmed by investigating other body fluids such as cerebrospinal fluid or bronchoalveolar lavage fluid in addition to sera. For invasive Candida infections several diagnostic kits detecting antigens are commercially available. The performance of the EIA Platelia Candida Antigen (Bio-Rad, USA), the latexagglutination test Cand-Tec (Ramco, USA) and the detection of (1-3)-Beta-D Glucan using the test kit Glucatell (CAPE COD, USA) were evaluated. The detection of (1-3)-Beta-D-Glucan by means of Glucatell showed the highest sensitivity as all sera of 15 patients with invasive candidosis showed positive results. The other tests showed positive and negative results independently of the occurrence of an invasive infection. However, the number of tested sera is too small to decide which test is the most appropriate for establishing an early and definite diagnosis. Prospective studies in combination with clinical data are still needed for definite evaluation. PMID- 15667366 TI - [Antibody detection in patients with invasive aspergillosis]. AB - The clinical significance of Aspergillus antibody assays for the diagnosis of invasive aspergillosis (IA) is unclear. In two studies, three different antibody assays were evaluated with patients suffering from proven IA: (i) a commercial haemagglutination test (HAT), (ii) a commercial enzyme immunoassay (EIA) for IgG, IgM, and IgA, and (iii) an experimental mitogillin enzyme immunoassay for IgG, IgM, and IgA. In the first study, 99 serum samples from 26 patients with IA and 22 serum samples from 22 control patients were tested with all the three tests. Ten of the 26 patients (38%) reacted positively in at least one antibody assay. The highest sensitivity was generated by the detection of IgG using the EIA formats (22 and 21%, respectively), the HAT had a sensitivity of 8%. IgM type antibodies were detected in only two patients; no IgA type antibodies were detected. The specificities of the IgG EIA and the HAT were 72 and 85%, respectively. Antibody detection was the single positive laboratory test in two patients with proven and probable IA. In the second study, antibody test results of 60 patients with proven IA were retrospectively evaluated. Fourteen patients (23%) tested positive in the EIA and/or in the HAT. Investigations of the antibody levels in individual immunocompromised patients over time revealed that IgG production started after a mean of 10.8 days after diagnosis of IA. To conclude, antibodies against Aspergillus were detected in 23% of patients with IA. The antibody production started in successfully treated immunosuppressed patients after a mean of 10.8 days after the onset of infection. In particular, the detection of IgG-antibodies with an EIA can be useful for the confirmation of the diagnosis of IA and for the monitoring of the treatment of IA. PMID- 15667367 TI - [Early signals of systemic mycoses--guidelines for therapeutic decisions?]. AB - Clinical applications and in vitro studies show that all antimycotics are most effective against infection when applied as early as possible. An early diagnosis is, therefore, essential. At this time for aspergillosis and particularly candidosis, a statement is only possible by combining several diagnostic methods. Inflammatory parameters like procalcitonin, C-reactive protein and proinflammatory cytokines are most important evidence of infection. Antigen tests are more significant by higher sensitivity. Attention should be focused on the detection of mannan by ELISA test, beta-glucan and D-arabinitol. Given the present research level in the field of proteomics, the diagnostic importance of transmembranal receptor proteins or other regulatory proteins seems promising. PMID- 15667368 TI - Five more years--thank you! PMID- 15667369 TI - Functional-task exercise versus resistance strength exercise to improve daily function in older women: a randomized, controlled trial. AB - OBJECTIVES: To determine whether a functional-task exercise program and a resistance exercise program have different effects on the ability of community living older people to perform daily tasks. DESIGN: A randomized, controlled, single-blind trial. SETTING: Community leisure center in Utrecht, the Netherlands. PARTICIPANTS: Ninety-eight healthy women aged 70 and older were randomly assigned to the functional-task exercise program (function group, n=33), a resistance exercise program (resistance group, n=34), or a control group (n=31). Participants attended exercise classes three times a week for 12 weeks. MEASUREMENTS: Functional task performance (Assessment of Daily Activity Performance (ADAP)), isometric knee extensor strength (IKES), handgrip strength, isometric elbow flexor strength (IEFS), and leg extension power were measured at baseline, at the end of training (at 3 months), and 6 months after the end of training (at 9 months). RESULTS: The ADAP total score of the function group (mean change 6.8, 95% confidence interval (CI)=5.2-8.4) increased significantly more than that of the resistance group (3.2, 95% CI=1.3-5.0; P=.007) or the control group (0.3, 95% CI=-1.3-1.9; P<.001). Moreover, the ADAP total score of the resistance group did not change significantly compared with that of the control group. In contrast, IKES and IEFS increased significantly in the resistance group (12.5%, 95% CI=3.8-21.3 and 8.6%, 95% CI=3.1-14.1, respectively) compared with the function group (-2.1%, 95% CI=-5.4-1.3; P=.003 and 0.3%, 95% CI=-3.6-4.2; P=.03, respectively) and the control group (-2.7%, 95% CI=-8.6-3.2, P=.003 and 0.6%, 95% CI=-3.4-4.6; P=.04, respectively). Six months after the end of training, the increase in ADAP scores was sustained in the function group (P=.002). CONCLUSION: Functional-task exercises are more effective than resistance exercises at improving functional task performance in healthy elderly women and may have an important role in helping them maintain an independent lifestyle. PMID- 15667370 TI - Executive control mediates memory's association with change in instrumental activities of daily living: the Freedom House Study. AB - OBJECTIVES: To assess the relative independent contribution of changes in executive control function (ECF) and memory to changes in functional status. DESIGN: Three-year longitudinal cohort study. SETTING: A comprehensive care retirement community. PARTICIPANTS: Five hundred forty-seven noninstitutionalized people aged 70 and older. MEASUREMENTS: The California Verbal Learning Test (CVLT) and Executive Interview (EXIT25). Functional status was assessed using instrumental activities of daily living (IADLs). Latent growth curves of CVLT, EXIT25, and IADLs were modeled. The rate of change in IADLs (adjusted for baseline IADLs and cognition) was regressed on the rate of change in each cognitive measure. Models were also adjusted for baseline age, level of care, and comorbid illnesses. RESULTS: There was significant variability around the baseline means and slopes for all variables. The rate of change in EXIT25 was independently correlated with the rate of change in IADLs (correlation coefficient (r)=-0.52, P<.001). This remained significant after adjusting for baseline EXIT25 scores, IADLs, age, comorbid disease, and level of care. The EXIT25's effect on the rate of change in IADLs was stronger than those of age, baseline IADLs, comorbid disease, or level of care. The rate of change in CVLT scores was not significantly associated with the rate of change in IADLs. CONCLUSION: ECF is a strong, significant, and independent correlate of functional status in normal aging. In contrast, decline in memory, as measured using the CVLT, has no independent association with the rate of change in functional status. This suggests that amnestic mild cognitive impairment can be associated with dementia only though the subsequent or comorbid development of ECF impairment. PMID- 15667371 TI - A multifactorial intervention to reduce prevalence of delirium and shorten hospital length of stay. AB - OBJECTIVES: To improve outcomes for cognitively impaired and delirious older adults. DESIGN: Pretest, posttest. SETTING: A university-affiliated hospital. PARTICIPANTS: Physicians and nurses in the emergency department (ED) and on an acute geriatric unit (AGU). INTERVENTION: Multifactorial and targeted to the processes of care for cognitively impaired and delirious older adults admitted to medicine service from the ED. MEASUREMENTS: Prevalence of delirium, admission to AGU, psychotropic medication use, hospital length of stay. RESULTS: Patient characteristics did not differ between baseline and the two outcome cohorts 4 and 9 months postintervention. Prevalence of delirium was 40.9% at baseline, 22.7% at 4 months (P<.002), and 19.1% at 9 months (P<.001). More delirious patients were admitted to the AGU than to non-AGU units at 4 months (P<.01) and 9 months (P<.01). Postintervention medication use in the hospital differed from baseline. Antidepressant use was greater at 4 months (P<.05). Benzodiazepine and antihistamine use were lower at 9 months (P>.01). Antidepressant and neuroleptic use were higher (P<.02) and antihistamine use was lower (P<.02) at 4 months on the AGU than for the baseline group. Benzodiazepine (P<.01) and antihistamine (P<.05) use were lower at 9 months. Each case of delirium prevented saved a mean of 3.42 hospital days. CONCLUSION: A multifactorial intervention designed to reduce delirium in older adults was associated with improved psychotropic medication use, less delirium, and hospital savings. PMID- 15667372 TI - Behavioral and mood effects of snoezelen integrated into 24-hour dementia care. AB - OBJECTIVES: To investigate the effectiveness of snoezelen, integrated in 24-hour daily care, on the behavior and mood of demented nursing home residents. DESIGN: Quasiexperimental pre- and posttest design. SETTING: Twelve psychogeriatric wards of six nursing homes, spread over different parts of the Netherlands. PARTICIPANTS: One hundred twenty-five patients with moderate to severe dementia and care dependency were included in the pretest and 128 in the posttest; 61 were completers (included in both pre- and posttest). INTERVENTION: Experimental subjects received an individual 24-hour snoezel program, based on family history taking and stimulus preference screening. Caregivers were trained, and (organizational) adaptations were made to fulfill the conditions for resident oriented snoezel care. The control group received usual nursing home care. MEASUREMENTS: Observations were made on the wards using subscales of the Dutch Behavior Observation Scale for Psychogeriatric Inpatients, the Dutch version of the Cohen-Mansfield Agitation Inventory, and the Cornell Scale for Depression in Dementia. Independent assessors observed video recordings of morning care and rated residents' behavior and mood using INTERACT and FACE, respectively. RESULTS: Residents receiving snoezel care demonstrated a significant treatment effect with respect to their level of apathetic behavior, loss of decorum, rebellious behavior, aggressive behavior, and depression. During morning care, the experimental subjects showed significant changes in well-being (mood, happiness, enjoyment, sadness) and adaptive behavior (responding to speaking, relating to caregiver, normal-length sentences). CONCLUSION: Snoezel care particularly seems to have a positive effect on disturbing and withdrawn behavior. The results suggest that a 24-hour integrated snoezel program has a generalizing effect on the mood and behavior of demented residents. PMID- 15667374 TI - Sex differences in the risk of frailty for mortality independent of disability and chronic diseases. AB - OBJECTIVES: To determine the effect of static and dynamic frailty on mortality in older men and women. DESIGN: A prospective cohort study with three 3-year measurement cycles. SETTING: Population based. PARTICIPANTS: The sample was derived from the Longitudinal Aging Study Amsterdam and consisted of respondents who participated in two cycles (T1: 1992/1993 and T2: 1995/1996) and for whom there was complete data on disability and chronic diseases (N=2,257). MEASUREMENTS: Nine frailty markers were assessed at T1 and T2. The frailty markers were defined in two ways: low functioning at T2 (static frailty) and change in functioning between T1 and T2 (dynamic frailty). Survival time, calculated in days from T2 to January 1, 2000, was used as the outcome variable. Predictive ability was examined using Cox proportional hazards analyses separately for men and women. RESULTS: Women were frailer than men. Static frailty was significantly associated with mortality in men (relative risk (RR)=2.4) and in women (RR=2.6). Dynamic frailty was also associated with mortality in women (RR=2.6), but it was not significantly associated with mortality in men (RR=1.3). When disability and chronic diseases were included in the model as possible mediators, these RRs dropped to 1.6, 2.0, 2.1, and 1.2, respectively, of which the first three were still significant. CONCLUSION: Frailty was associated with mortality to a greater extent in women than in men, and this effect was independent of disability and chronic disease. In men, the static definition of frailty was more predictive of mortality than the dynamic definition. PMID- 15667373 TI - The influence of anxiety on the progression of disability. AB - OBJECTIVES: To determine the influence of anxiety on the progression of disability and examine possible mediators of the relationship. DESIGN: Community based observational study. SETTING: Women's Health and Aging Study I, a prospective observational study with assessments every 6 months for 3 years. PARTICIPANTS: One thousand two functionally limited women aged 65 and older. MEASUREMENTS: Anxiety symptoms were assessed using four questions from the Hopkins Symptom Checklist (nervous or shaky, avoidance of certain things, tense or keyed up, fearful). Participants who reported experiencing two or more of these symptoms at baseline were considered anxious. Anxiety as a predictor of the onset of four types of disability was examined using Cox proportional hazards models. Three models were tested: an unadjusted model, a model adjusted for confounding variables (age, race, vision, number of diseases, physical performance, depressive symptoms), and a mediational model (benzodiazepine and psychotropic medication use, physical activity, emotional support). RESULTS: Nineteen percent of women reported two or more symptoms of anxiety at baseline. Unadjusted models indicate that anxiety was associated with a greater risk of worsening disability: activity of daily living (ADL) disability (relative risk (RR)=1.40, 95% confidence interval (CI)=1.10-1.79), mobility disability (RR=1.41, 95% CI=1.06-1.86), lifting disability (RR=1.54, 95% CI=1.20-1.97), and light housework disability (RR=1.77, 95% CI=1.32-2.37). After adjusting for confounding variables, anxiety continued to predict the development of two types of disability: ADL disability (RR=1.41, 95% CI=1.08-1.84) and light housework disability (RR=1.56, 95% CI=1.14-2.14). Finally, benzodiazepine and psychotropic medication use, physical activity, and emotional support were not significant mediators of the effect of anxiety on the development of a disability. CONCLUSION: Anxiety is a significant risk factor for the progression of disability in older women. Studies are needed to determine whether treatment of anxiety delays or prevents disability. PMID- 15667375 TI - Effects of a nap on nighttime sleep and waking function in older subjects. AB - OBJECTIVES: To examine, in older subjects, the effect on waking function of increasing 24-hour sleep amounts by providing a nap opportunity; to assess what effects an afternoon nap may have on subsequent nighttime sleep quality and composition. DESIGN: Two-session, within-subject laboratory design. SETTING: The study was conducted in the Laboratory of Human Chronobiology at Weill Cornell Medical College. PARTICIPANTS: Thirty-two healthy men and women aged 55 to 85. MEASUREMENTS: Polysomnography (sleep electroencephalogram), cognitive and psychomotor performance, body core temperature. RESULTS: Napping had little effect on subsequent nighttime sleep quality or duration, resulting in a significant increase in 24-hour sleep amounts. Such increased sleep resulted in enhanced cognitive and psychomotor performance immediately after the nap and throughout the next day. CONCLUSION: A behavioral approach that adds daytime sleep to the 24-hour sleep quota seems worthy of consideration when presented with a situation in which physiological changes associated with aging may limit the duration of nighttime sleep. PMID- 15667376 TI - Relationship between self-assessed masticatory disability and 9-year mortality in a cohort of community-residing elderly people. AB - OBJECTIVES: To examine the relationship between self-assessed masticatory disability and mortality. DESIGN: Prospective. SETTING: Community based. PARTICIPANTS: Total of 1,405 randomly selected people aged 65 and older living in Settsu, Osaka Prefecture, in October 1992. MEASUREMENTS: Data on health status as indicated by disability scores, history of health management, self-assessed masticatory ability, and psychosocial conditions were collected by means of interviews during home visits at the time of enrollment. Nine-year follow-up was completed for 1,245 (88.6%; 398 deceased and 847 alive). RESULTS: Self-assessed masticatory disability was significantly associated with being 75 and older, having overall disability, not using dental health checks or general health checks, not participating in social activities, not feeling that life is worth living (no ikigai), and finding relationships with people difficult. As for the association between self-assessed masticatory disability and mortality, the estimated survival rate for those with self-assessed masticatory disability was lower than that for those without for each group stratified by sex and age (65-74 and >or=75), and the equality of survival curves according to self-assessed masticatory disability was significant for each group. After controlling for potential predictors of mortality, self-assessed masticatory disability remained as a significant predictor of mortality (adjusted hazard ratio=1.63, 95% confidence interval=1.30-2.03, P<.001). CONCLUSION: These results indicate that self-assessed masticatory disability may be associated with a greater risk of mortality in community-residing elderly people. PMID- 15667378 TI - Qualitative analysis of Medicare claims in the last 3 years of life: a pilot study. AB - OBJECTIVES: To study end-of-life care of a representative sample of older people using qualitative interpretation of administrative claims by clinicians and to explore whether this method yields insights into patient care, including continuity, errors, and cause of death. DESIGN: Random, stratified sampling of decedents and all their Medicare-covered healthcare claims in the 3 years before death from a 5% sample of elderly fee-for-service beneficiaries, condensation of all claims into a chronological clinical summary, and abstraction by two independent clinicians using a standardized form. SETTING: United States. PARTICIPANTS: One hundred Medicare fee-for-service older people without disability or end-stage renal disease entitlement who died in 1996 to 1999 and had at least 36 months of continuous Part A and Part B enrollment before death. MEASUREMENTS: Qualitative narrative of the patient's medical course; clinician assessment of care continuity and apparent medical errors; cause, trajectory, and place of death. RESULTS: The qualitative narratives developed by the independent abstracters were highly concordant. Clinicians felt that 75% of cases lacked continuity of care that could have improved the quality of life and the way the person died, and 13% of cases had a medical error identified by both abstracters. Abstracters disagreed about assignment of a single cause of death in 28% of cases, and abstracters and the computer algorithm disagreed in 43% of cases. CONCLUSION: Qualitative claims analysis illuminated many problems in the care of chronically ill older people at the end of life and suggested that traditional vital statistics assignation of a single cause of death may distort policy priorities. This novel approach to claims review is feasible and deserves further study. PMID- 15667377 TI - Unmet desire for caregiver-patient communication and increased caregiver burden. AB - OBJECTIVES: To examine the adequacy of caregiver-patient communication in serious illness and its relationship to caregiver burden. DESIGN: Cross-sectional cohort study. SETTING: Participants' homes. PARTICIPANTS: One hundred ninety-three persons aged 60 and older seriously ill with cancer, congestive heart failure, or chronic obstructive pulmonary disease and their caregivers. MEASUREMENTS: Communication concerns, measured in terms of agreement with statements regarding desire for and difficulty with communication about the patient's illness. Caregiver burden, measured using a 10-item subset of the Zarit Burden Inventory, with scores ranging from 0 to 40 and higher scores indicating greater burden. RESULTS: Of caregivers, 39.9% desired more communication, and 37.3% reported that communication was difficult. Of patients, 20.2% desired more communication, and 22.3% reported that communication was difficult. Disagreement regarding communication concerns was frequent in caregiver-patient pairs; of caregivers who desired more communication, 83.1% of patients did not, and of patients who desired more communication, 66.7% of caregivers did not. Caregivers who desired more communication had significantly higher caregiver burden scores than did caregivers who did not (9.2 vs 4.7, P<.001), even after adjusting for patient's diagnosis, income, and functional status and caregivers' age, sex, and relationship to the patient. CONCLUSION: A large proportion of caregivers and seriously ill older persons had an unmet desire for increased communication, although they frequently disagreed with each other about this desire. Caregivers' desire for increased communication may be a modifiable determinant of caregiver burden. PMID- 15667379 TI - Drop attacks in older adults: systematic assessment has a high diagnostic yield. AB - OBJECTIVES: To investigate the causes of recurrent drop attacks in older patients with a comprehensive battery of investigations in the largest series reported to date. DESIGN: Observational with mean followup of 18 months. SETTING: Inner city emergency department and tertiary facility in Newcastle upon Tyne, United Kingdom. PARTICIPANTS: Ninety-three consecutive patients aged 55 and older with three or more drop attacks in the 6 months before evaluation. METHODS: Subjects underwent a comprehensive diagnostic evaluation, with particular attention to traditional (e.g., gait and balance abnormalities, medications) and more recently identified (e.g., carotid sinus hypersensitivity) risk factors for drop attacks. RESULTS: Subjects tended to be older (mean age+/-standard deviation 77.4+/-9.0) and female (70; 75%) and to have a mean of 10.4 drop attacks before evaluation. Fifty-three (57%) had suffered soft tissue injuries needing medical attention and 32 (34%) fractures secondary to drop attacks. An attributable diagnosis was achieved in all but nine subjects (90%). Cardiovascular diagnoses (49; 53%) were most commonly implicated, with neurological (27; 29%) and gait and balance abnormalities (17; 18%) and drug-related causes (11; 12%) providing the majority of the remaining diagnoses. CONCLUSION: Drop attacks in older subjects are associated with high levels of morbidity and healthcare resource utilization. Attributable diagnoses are achievable in the majority of cases with a systematic investigative approach. The high diagnostic yield more than justifies the approach described. PMID- 15667380 TI - Genetic variant of lactase-persistent C/T-13910 is associated with bone fractures in very old age. AB - OBJECTIVES: To determine the relation between the C/T(-13910) single-nucleotide polymorphism residing 13,910 base pairs from the 5' end of the lactase gene associated with lactase persistence and the occurrence of bone fractures in elderly people. DESIGN: Population-based study. SETTING: Vantaa 85+ population based study, including all 601 subjects born before April 1, 1906, who were living in the city of Vantaa, Finland, on April 1, 1991. PARTICIPANTS: Four hundred eighty-three people aged 85 and older (106 men and 377 women). MEASUREMENTS: Genotype determination was made using a polymerase chain reaction minisequencing technique. RESULTS: The frequency of the genotype C/C(-13910) associated with adult-type hypolactasia (low lactase enzyme activity or primary lactose malabsorption (LM)) was significantly greater in individuals with hip fractures, with an adjusted odds ratio (OR) of 3.7 (95% confidence interval (CI)=1.8-7.8), wrist fractures with an adjusted OR of 2.5 (95% CI=1.2-5.2), and hip and wrist fractures combined with an adjusted OR of 4.1 (95% CI=2.0-8.3). CONCLUSION: The C/C(-13910) genotype associated with primary LM could represent a genetic risk factor for bone fractures for elderly people. PMID- 15667381 TI - Alteration of a clinically meaningful outcome in the natural history of Alzheimer's disease by cholinesterase inhibition. AB - OBJECTIVES: To describe the effect of cholinesterase inhibitors (CEIs) on the natural course of Alzheimer's disease (AD) using clinically meaningful outcomes. DESIGN: Cross-sectional and longitudinal study. SETTING: Referral dementia clinic. PARTICIPANTS: One hundred thirty-five matched pairs of patients with probable AD. MEASUREMENTS: The risk of AD patients being classified as slow progressors (Mini-Mental State Examination (MMSE) score change 3.5). MS1 and fellows had significantly more favorable attitudes scores than more advanced students and residents. CONCLUSION: The results suggest that the UCLA Attitudes Scale and Knowledge Test can be used reliably to assess attitudes and knowledge level across all levels of medical education and training. The information from this study will be used to implement a more structured and comprehensive geriatrics curriculum across all trainee levels to improve attitudes and knowledge in the care of the geriatric patient. PMID- 15667385 TI - Association between mild vascular cognitive impairment and impaired activities of daily living in older stroke survivors without dementia. AB - OBJECTIVES: To determine the relationship between mild vascular cognitive impairment (mVCI) and functional disability in older stroke survivors without dementia. DESIGN: Cohort study. SETTING: Stroke patients from representative hospital-based stroke registers. PARTICIPANTS: Three hundred thirty-nine stroke survivors without dementia, aged 75 and older. MEASUREMENTS: Neuropsychological assessments were completed 3 months poststroke. Activities of daily living (ADLs) were evaluated using the Bristol scale. Operationalized criteria, including cognitive impairment no dementia (CIND), were applied for mVCI. RESULTS: Significant impairments of ADLs were evident in mVCI (CIND vs no CIND; basic care: z=3.2; P=.001, intermediate care: z=3.6; P<.001, complex management: z=4.5; P<.001) but varied according to the profile of cognitive impairments. Patients with attentional or global impairments had more severe functional disability than patients with isolated memory deficits, with an intermediate level of dysfunction in patients with executive impairments. CONCLUSION: The findings emphasize the importance of identifying early cognitive impairments to optimize rehabilitation, care planning, and secondary prevention of dementia in stroke patients. PMID- 15667386 TI - Health-related quality of life, functional impairment, and healthcare utilization by veterans: veterans' quality of life study. AB - OBJECTIVES: To describe the health status of veterans receiving care in a veterans integrated service network (VISN). DESIGN: Cross-sectional survey with prospective follow-up. SETTING: Former Upper Midwest VISN 13 (now a part of VISN 23), a regional Veterans Affairs (VA) network comprising five inpatient facilities and associated outpatient clinics. PARTICIPANTS: All veterans in VISN 13 who had at least one inpatient or outpatient encounter between October 1, 1997, and March 31, 1998. MEASUREMENTS: Health-related quality of life (HRQOL) assessed using subscales and component summaries from the 36-item short form for veterans (SF36-V), functional status assessed according to limitations in activities of daily living (ADLs), healthcare utilization assessed according to outpatient visits and hospitalizations, and death. RESULTS: Of 70,334 eligible veterans, 40,508 responded and reported baseline HRQOL significantly lower than that of the general U.S. population for the physical (35.6, P<.001) and mental (46.4, P<.001) component summary scores (PCS and MCS, respectively) of the SF36 V. Many reported complete inability or some difficulty in completing ADLs such as getting in and out of a chair (35.1%) and walking (45.3%). More than 58% indicated some degree of difficulty with at least one of the ADLs. In multivariate analysis, PCS and MCS were significantly associated with subsequent use of inpatient and outpatient care and with mortality. CONCLUSION: The low quality of life and associated high rates of health services utilization in VA patients imply a need for innovative strategies to improve the HRQOL and functional status of this population. PMID- 15667387 TI - Treatment of vasodepressor carotid sinus syndrome with midodrine: a randomized, controlled pilot study. AB - OBJECTIVES: To evaluate the efficacy of treatment of the vasodepressor form of carotid sinus hypersensitivity (carotid sinus syndrome (CSS)) with midodrine. DESIGN: A prospective, double-blind, randomized, controlled trial of crossover design. SETTING: A dedicated outpatient facility with access to tilt-table, digital arterial photoplethysmography, and 24-hour ambulatory blood pressure (BP) monitoring equipment. PARTICIPANTS: Ten older adults (4 male, 6 female, mean age 75, range 66-86 years) with a history of unexplained syncope who displayed an asymptomatic decrease in systolic BP (SBP) of more than 50 mmHg or a symptomatic decrease of more than 30 mmHg within 30 seconds of carotid sinus massage (CSM). MEASUREMENTS: Symptom reproduction and BP and heart rate changes were evaluated after CSM in supine and semierect positions on the right and then left sides. These measurements were performed on the final day of placebo and active treatment phases. Ambulatory 24-hour BP monitoring took place on the penultimate and final days of each treatment phase. RESULTS: Eight patients were symptomatic after their initial CSM. The mean+/-standard deviation SBP decrease after initial CSM was 54+/-22 mmHg. Initial mean 24-hour ambulatory BP was 127/70+/-7/5 mmHg. Eight patients reported symptoms after CSM at the end of the placebo phase. The mean SBP decrease at the end of the placebo phase was 49+/-12 mmHg. The mean 24 hour ambulatory BP was 127/69+/-9/7 mmHg. One patient reported symptoms after CSM at the end of the active-treatment phase. The mean SBP decrease at the end of the active-treatment phase was 36+/-9 mmHg. The mean 24-hour ambulatory BP at the end of the treatment phase was 133/75+/-7/6 mmHg. The differences in symptom reporting and mean SBP decrease after CSM were both significant (P<.01 and P=.03, respectively). CONCLUSION: The results of this pilot study suggest that treatment of vasodepressor CSS with midodrine significantly reduced the rate of symptom reporting and attenuated SBP decreases after CSM but increased mean 24-hour ambulatory BP. PMID- 15667388 TI - From aspiration to achievement: assessment and noninvasive treatment of erectile dysfunction in aging men. AB - More than 70% of elderly men (>or=65) remain sexually active, and more than 40%, according to one estimate, are dissatisfied with their sex lives. Declining sexual function and a reluctance to seek medical attention with advancing age are cross-cultural observations. Normal erection is largely dependent on intact function of the central and peripheral nervous systems and the penile vascular endothelium. Consequently, chronic conditions (e.g., cardiovascular disease, diabetes mellitus) and lifestyle factors (e.g., smoking) that have adverse effects on the vascular endothelium and central and peripheral nervous systems, as well as on endocrine function or connective tissues within the corpus cavernosum of the penis, can attenuate erectile function. Because of these associations, assessment of sexual function in elderly men often reveals not only erectile dysfunction (ED) but also other reversible conditions. An expanding array of noninvasive options is available to assist the clinician in individualizing ED therapy to the unique health and lifestyle needs of each elderly ED patient and his partner. These treatment alternatives include the phosphodiesterase type 5 inhibitors sildenafil, vardenafil, and tadalafil, as well as other oral medications, such as alpha-adrenoceptor antagonists and topical vasoactive or testosterone therapy. PMID- 15667389 TI - Attitudes of Asian-Indian Hindus toward end-of-life care. AB - The population of older adults in the United States is increasing in cultural diversity. Thus, an appreciation of cultural perspectives regarding end-of-life care may benefit practicing physicians. This article addresses end-of-life care issues from the perspective of Hinduism--a faith embraced by a growing demographic of the U.S. population-and also reviews basic Hindu philosophical concepts salient to caring for patients at the end of life. PMID- 15667390 TI - Geriatric content in medical school curricula: results of a national survey. AB - Despite recent gains in establishing academic sections, divisions, and departments of geriatrics in medical schools, much remains to be done to meet the medical needs of an aging population. To better understand how medical schools are educating students in geriatric-related topics, all U.S. allopathic and osteopathic medical schools were surveyed in two waves, in 1999 and 2000, using a questionnaire based on recommendations from the Education Committee of the American Geriatrics Society. Responding schools were more likely to address diseases and conditions of aging, psychosocial issues, and ethical issues and less likely to cover anatomic changes, nutrition, knowledge of healthcare financing, outcome measurement, and cultural aspects of aging. Although limited, the results indicate that medical schools have increased coverage of aging related material, although further expansion of geriatric content will be necessary to meet the needs of an aging society. PMID- 15667391 TI - Emergency medicine management of the geriatric patient: an educational program for medical students. AB - This article describes the experience of fourth-year medical students participating in a geriatric education program integrated into a 4-week emergency medicine student clerkship. Between July 2002 and April 2003, all students in this required clerkship participated in a geriatric educational program consisting of a small group discussion of medical and psychosocial issues of older adult emergency department (ED) patients. Students used learned skills to evaluate older adult ED patients for medical and psychosocial issues and later followed up with these patients by telephoning them at their homes or visiting them in the hospital. Students tracked their evaluations of the medical problems, functional abilities, and social supports of patients in the ED. Students also noted when their assessments resulted in the acquisition of new skills or knowledge and when their evaluation of geriatric syndromes resulted in a change of the patient care plan. Seventy-seven students evaluated 217 patients in the ED, of whom 167 (77%) received a follow-up visit or phone call. Students documented learning new skills while caring for 80 (48%) of the older adult patients. Qualitative survey responses from students indicated that students had increased understanding of the importance of assessing functional status and social supports and providing interdisciplinary care. Integrating geriatric education modules into existing emergency medicine clerkships is an effective method to expand the geriatric curriculum in medical schools and to emphasize the importance of geriatric assessment and syndromes in emergency care. PMID- 15667392 TI - Differences in amount of informal care received by non-Hispanic whites and latinos in a nationally representative sample of older Americans. AB - The objective of this study was to evaluate informal (unpaid) care and its broad determinants for Latinos in a nationally representative sample. A cross-sectional analysis of the 1993 Asset and Health Dynamics Study, a national probability sample of 7,443 older adults aged 70 and older, was performed to determine the independent effect of Latino ethnicity on the receipt of informal care by disabled older individuals. Self-reported race/ethnicity was used to predict the mean daily hours of informal care received for activity of daily living (ADL) or instrumental activity of daily living (IADL) assistance after adjustment for predisposing, need, and enabling variables. There was a significant association between informal home care and ethnic group, with 44.3% of Latinos receiving informal care, compared with 33.9% of African Americans and 24.6% of non-Hispanic whites (P<.001). After adjustment, Latinos received 11.0 weekly hours of informal care, compared with 7.5 hours for non-Hispanic whites and 6.3 hours for African Americans (P<.001). The results from this nationally representative sample indicate that Latinos receive significantly more hours of informal care on average than African Americans or non-Hispanic whites for ADL and IADL disability. Clinicians should be alert to the significant amount of informal care and possible associated strain in caregivers of older Latinos. PMID- 15667393 TI - A nursing home in Arab-Israeli society: targeting utilization in a changing social and economic environment. AB - This article is a case study of the first 10 years of operation (1992-2002) of the Dabouriya Home for the Aged, the first publicly funded culturally adapted nursing home for Israeli citizens of Arab descent. Although 44% of Arab Israelis and 26% of Jewish Israelis aged 65 and older are disabled, in 1999, 4.3% of the Jewish population but only 0.7% of the Arab-Israeli population aged 65 or older lived in long-term care institutions; disabled Arab-Israeli elderly were mainly cared for by families. As Arab-Israeli society modernizes and traditional caregiving is reduced, alternatives must be found for this growing, disabled population. Medical and administrative records of 404 people admitted consecutively to a 136-bed facility over 10 years were analyzed. Two distinct segments of the needy population were served: people with independent activity of daily living (ADL) function but little or no family to provide help with intermediate ADLs and those dependent in ADLs and with health problems, especially dementia. Economic, demographic, and social changes in Arab-Israeli society may mean that traditional caregivers will not be able to adequately care for this highly disabled population. Administrators of the public health system in Israel should be aware of the underutilization of publicly funded long-term care by disabled Arab Israelis and the lack of care alternatives for the population that does use nursing homes, because there may be severe consequences in terms of caregiver burden and social stress when disabled elderly people remain in unsuitable environments. PMID- 15667395 TI - Drop attacks revisited: yet another manifestation of cardiovascular disease? PMID- 15667396 TI - The emperor has no clothes: dementia treatment on the eve of the aging era. PMID- 15667398 TI - Tramadol/acetaminophen combination tablets for the treatment of pain associated with osteoarthritis. PMID- 15667399 TI - Aromatase inhibitors and mental function. PMID- 15667400 TI - All patient refined diagnosis related groups: a new administrative tool for identifying elderly patients at risk of high resource consumption. PMID- 15667401 TI - Physical and cognitive functioning and resuscitation preferences of aged patients. PMID- 15667402 TI - Carrying habits into old age: prescription drug use without medical advice by older American adults. PMID- 15667403 TI - Chronic colonic pseudo-obstruction associated with recurrent congestive heart failure. PMID- 15667404 TI - Safety of tirofiban therapy in very old patients with acute coronary syndrome or non-Q-wave myocardial infarction. PMID- 15667405 TI - Diagnosis related groups and hospital costs in elderly dependent patients. PMID- 15667406 TI - Alcohol consumption and the risk of dementia: biological and psychological mechanisms. PMID- 15667409 TI - Teaching of neuroepidemiology in Europe: time for action. AB - Many epidemiological and clinical studies in Europe, especially in Eastern Europe and countries in transition, are of poor methodological quality because of lack of background knowledge in clinical epidemiology methods and study designs. The only way to improve the quality of epidemiological studies is to provide adequate undergraduate and/or postgraduate education for the health professionals and allied health professions. To facilitate this process, the European Federation of Neurological Societies (EFNS) Task Force on teaching of clinical epidemiology in Europe was set up in October 2000. Based on analyses of the current teaching and research activities in neuroepidemiology in Europe, this paper describes the Task Force recommendations aimed to improve these activities. PMID- 15667410 TI - A systematic review of the epidemiology of status epilepticus. AB - Population-based data on the incidence, aetiology, and mortality associated with status epilepticus (SE) are required to develop preventative strategies for SE. Through a systematic review, we aimed to assess the methodological quality as well as similarities, and differences between available population based studies in order to arrive at conclusions on the epidemiology of SE. All population-based studies where primary outcome was incidence, aetiology or mortality of SE were identified through a systematic search and synthesized. Methodological quality of studies were independently rated by two examiners using a unique scoring system. Seven population-based projects on SE yielding nine published reports and five abstracts were reviewed. Quality scores were in the range of 19-34 with a possible maximum of 40 (kappa scores 0.67-1.0). The incidence of SE has a bimodal distribution with peaks in children aged less than a year and the elderly. Most SE were acute symptomatic. Short-term mortality was 7.6-22% and long-term mortality was 43%. Age and aetiology were the major determinants of mortality. There are few population-based studies on SE but most are of good quality. Most studies are primarily or exclusively based on adult populations. There is limited information on the association of ethnicity and socio-economic status and SE. PMID- 15667411 TI - Stress lactate in mitochondrial myopathy under constant, unadjusted workload. AB - As it is under debate if determination of lactate during cycle ergometry (lactate stress testing, LST) under a continuous, unadjusted, low workload is a valuable diagnostic tool for mitochondrial myopathy (MMP), the present study aimed to investigate how sensitive the LST is in a large cohort of patients with indications for MMP (MMP patients). Serum lactate was determined once before, three times during, and once after a 15-min, constant 30 W-workload on a bicycle ergometer in 115 healthy controls, 166 patients with neurological disorders other than MMP, and 291 MMP patients. Serum lactate's upper reference limit at rest, 5, 10, 15 min after starting, and 15 min after finishing the exercise was 2.0, 2.0, 2.1, 2.0 and 1.7 mmol/l, respectively. Resting lactate was increased in 75 MMP patients (26%). The specificity of resting lactate determination was 84%. The sensitivity of the LST was 66% and the specificity 84%. Among the 192 MMP patients with abnormal LST, 120 (63%) had a normal resting lactate. The LST is abnormal in two thirds of the MMP patients. The sensitivity of the LST is higher than that of resting lactate determination. The LST is a simple and cheap but effective and reliable screening method for detecting the impaired oxidative metabolism in MMP. PMID- 15667413 TI - Medial temporal lobe atrophy and memory deficits in elderly stroke patients. AB - Medial temporal lobe atrophy (MTA) and its role in memory deficits have been studied extensively in patients with various dementias and non-degenerative neurologic diseases. In stroke patients MTA is a significant risk factor for dementia. However, its role in memory decline in non-demented stroke patients is not yet known. Our aim was to evaluate the relationship between MTA and cognitive functions in a large cohort of elderly patients, who underwent a comprehensive neuropsychologic examination and magnetic resonance imaging 3 months after an ischemic stroke. The study sample (n = 260) was divided into three groups according to the severity of MTA. After adjusting for age, volume of infarcts and cortical atrophy, we found that patients with moderate to severe MTA performed significantly worse in tests of learning, story recall, visual reproduction, block design and mental speed. In contrast, the groups did not differ in tests of digit span, flexibility, verbal fluency and conceptualization. Our conclusion is that in aged stroke patients, MTA is associated with poor performance in specific cognitive domains. The most vulnerable domains are memory and visuospatial functions, whereas verbal and executive functions seem to be unrelated to MTA. PMID- 15667412 TI - Hereditary spastic paraplegia with cerebellar ataxia: a complex phenotype associated with a new SPG4 gene mutation. AB - Complex forms of hereditary spastic paraplegia (HSP) are rare and usually transmitted in an autosomal recessive pattern. A family of four generations with autosomal dominant hereditary spastic paraplegia (AD-HSP) and a complex phenotype with variably expressed co-existing ataxia, dysarthria, unipolar depression, epilepsy, migraine, and cognitive impairment was investigated. Genetic linkage analysis and sequencing of the SPG4 gene was performed and electrophysiologic investigations were carried out in six individuals and positron emission tomography (PET) in one patient. The disease was linked to the SPG4 locus on chromosome 2p as previously reported for pure HSP. Sequence analysis of the SPG4 (spastin) gene identified a novel 1593 C > T (GLN490Stop) mutation leading to premature termination of exon 12 with ensuing truncation of the encoded protein. However, the mutation was only identified in those individuals who were clinically affected by a complex phenotype consisting of HSP and cerebellar ataxia. Other features noted in this kindred including epilepsy, cognitive impairment, depression, and migraine did not segregate with the HSP phenotype or mutation, and therefore the significance of these features to SPG4 is unclear. Electrophysiologic investigation showed increased central conduction time at somatosensory evoked potentials measured from the lower limbs as the only abnormal finding in two affected individuals with the SPG4 mutation. Moreover, PET of one patient showed significantly relatively decreased regional cerebral blood flow in most of the cerebellum. We conclude that this kindred demonstrates a considerable overlap between cerebellar ataxia and spastic paraplegia, emphasizing the marked clinical heterogeneity of HSP associated with spastin mutations. PMID- 15667414 TI - Plasma viral load of Epstein-Barr virus and risk of multiple sclerosis. AB - Elevated antibody titers to Epstein-Barr virus (EBV) have been found prior to the onset of multiple sclerosis (MS) and EBV has been found in serum of patients with MS exacerbations. We conducted a prospective, nested case-control study in the Nurses' Health Study and Nurses' Health Study II cohorts to determine whether plasma EBV viral load in healthy adults predicts the risk of MS. MS cases with blood collected before onset (n = 18) or diagnosis (n = 13) and 62 healthy controls were matched by age and time of blood collection. EBV viral load in plasma was measured by real time polymerase chain reaction. Presence of EBV in plasma was associated with an increased risk of MS (relative risk = 2.5, 95% CI 0.78-7.8, P = 0.12). Adjusting for smoking, ancestry, and latitude of residence at birth did not materially change this result. However, no association was found between the EBV viral load and risk of MS. These results support a role for EBV in the etiology of MS, but need to be confirmed in a larger study. PMID- 15667415 TI - Skin vasodilator response to local heating in human chronic spinal cord injury. AB - Local heating evokes an increase in skin blood flow (SkBF), which consists of an initial peak (axon-reflex mediated) followed by a brief nadir and a secondary rise to a plateau. The aim of this study was to investigate whether heat provoked vasodilatation detects sympathetic vasomotor dysfunction and completeness of injury in patients with spinal cord injury (SCI). Twelve (seven complete, and five incomplete; level C4-L4) SCI patients, and nine healthy subjects as controls were studied. Thermostatic laser Doppler probes, which heat the skin locally, were placed on the dorsum of the hand and foot. SkBF was measured by laser Doppler flowmetry at baseline and at the first peak of vasodilatation (SkBF(max)). On the hand, SkBF at baseline and SkBF(max) were similar between the three groups. On the foot, SkBF at baseline was similar between the three groups but SkBF(max) was significantly diminished in complete SCI patients compared with controls (P < 0.01). In conclusion, heat provoked axon-reflex vasodilatation was diminished in the foot, below the level of lesion, in complete SCI. This test, that evaluates localized sympathetic vasomotor dysfunction, may be a useful non invasive technique to detect completeness of autonomic disruption after SCI. PMID- 15667416 TI - EEG findings in dementia with Lewy bodies causing diagnostic confusion with sporadic Creutzfeldt-Jakob disease. AB - Two patients with pathologically confirmed dementia with Lewy bodies (DLB) had electroencephalogram recordings reported to show periodic discharges suggestive or typical of Creutzfeldt-Jakob disease. These findings caused diagnostic confusion and necessitated appropriate precautions for prion disease at postmortem examination. Periodic sharp wave complexes have occasionally been reported in DLB and should not therefore dissuade one from the diagnosis when other clinical and neuropsychological features are consistent with validated diagnostic criteria. PMID- 15667417 TI - Early sympathetic hyperactivity in Huntington's disease. AB - To investigate whether Huntington's disease (HD) affects autonomic nervous system (ANS) functioning 33 subjects with positive genetic test results were studied. The subjects were classified according to Shoulson and Fahn (S&F) HD disability scale into three subgroups: subgroup 1 (eight asymptomatic gene carriers), subgroup 2 (13 mildly disabled HD patients) and subgroup 3 (eight moderately and four severely disabled HD patients). A battery of cardiovascular autonomic tests (Valsalva maneuver, deep breathing test, sustained handgrip test, orthostatic test) and the spectral analysis of heart rate variability (HRV) were performed. The results were compared with a group of matched controls. In subgroup 1, there was a higher power of low-frequency band (LFB) (P < 0.05). In subgroup 2 a higher power of LFB was detected, LFB/high-frequency band (HFB) coefficient was increased and the blood pressure response to sustained handgrip was elevated (P < 0.05). Subgroup 3 showed significantly lower blood pressure response to sustained handgrip, lower respiratory (P < 0.05) and orthostatic ratio (P < 0.01). Our results suggest that sympathetic hyperfunction is present in asymptomatic gene carriers and mildly disabled HD patients. Contrary to that, ANS hypofunction was found in advanced HD patients. PMID- 15667426 TI - Subjective effects of antipsychotic treatment. PMID- 15667418 TI - Headache attributed to spontaneous low CSF pressure: report of three cases responsive to corticosteroids. AB - The therapy of headache attributed to spontaneous low CSF pressure (previously defined as spontaneous intracranial hypotension) is still a matter of debate. Epidural blood patch is considered the most effective treatment. However, pharmacological strategies may be considered before blood patch. We report three patients with headache attributed to spontaneous low CSF pressure that were successfully treated with oral prednisone. Additional studies may be useful to prove the effectiveness of corticosteroids in this syndrome. PMID- 15667427 TI - Cultural identities and cultural congruency: a new model for evaluating mental distress in immigrants. AB - OBJECTIVE: Societies and cultures have been described as being individualistic or collectivist (also called ego-centric or socio-centric). Similarly individuals are idiocentric or allocentric. METHOD: Using migration, migrants, ethnic minorities, collectivism and individualism, four databases of Embase, Medline, PsychInfo and Social Sciences abstracts were searched. Other key words included mental disorder, mental distress, psychiatric and psychological disorders. In addition, hand searches were conducted from the relevant books, monographs and secondary references. RESULTS: Migration, cultural identity and mental distress are linked. In addition, social support can provide a buffer against mental illness. Other vulnerability factors in migrants include the type of society they originate from and the type of society they settle in. CONCLUSION: When individuals migrate from one type of culture to another it is likely that depending upon their own personality traits (along with their biopsycho-social vulnerabilities) may develop psychiatric disorders. The cognitions and idioms of distress will be influenced by cultural factors. The clinicians must take into account cultural background when planning any interventions to enable a stronger therapeutic alliance. PMID- 15667428 TI - Structural and functional models of depression: from sub-types to substrates. AB - OBJECTIVE: To present a functional model of depression facilitating research and clinical understanding. METHOD: The authors conducted a systematic literature search and reviewed articles pertaining to the neurochemistry and pathophysiology of depressive disorders, focusing on the contribution made by the principal monoamines to three differing depressive structural sub-types (i.e. psychotic, melancholic and non-melancholic). RESULTS: We suggest that the three structural depressive subtypes appear functionally underpinned by differential contributions of serotonergic, noradrenergic and dopaminergic neurotransmitters, so influencing phenotypic distinction (our structural model) and allowing an aetiological model to be derived with treatment specificity implications. CONCLUSION: The functional model logically iterates with the structural model of depression and provides a useful framework for conceptualizing the depressive disorders. This model provides a logic for distinguishing between principal depressive subtypes, pursuing their functional underpinnings and explaining treatment differential effects across the three sub-types. PMID- 15667429 TI - Randomized double blind comparison of olanzapine vs. clozapine on subjective well being and clinical outcome in patients with schizophrenia. AB - OBJECTIVE: This randomized double-blind multicenter trial evaluated the effects of olanzapine vs. clozapine on subjective well-being, quality of life (QOL) and clinical outcome. METHOD: The primary objective was to demonstrate non inferiority of olanzapine, mean dosage 16.2 +/- 4.8 (5-25 mg/day) vs. clozapine, mean dosage 209 +/- 91 (100-400 mg/day) regarding improvement on the 'Subjective Well-Being under Neuroleptic Treatment' (SWN) Scale after 26 treatment weeks in 114 patients with schizophrenia. Secondary outcome parameters included: Munich QOL Dimension List (MLDL), Positive and Negative Symptom Scale (PANSS), Clinical Global Impression (CGI). RESULTS: SWN scores improved significantly in both groups, olanzapine was non-inferior to clozapine (group difference 3.2 points in favor of olanzapine; 95% CI: 4.2;10.5). MLDL-satisfaction, PANSS and CGI-S improved similarly, olanzapine yielded a higher CGI Therapeutic Index. Individual SWN and PANSS changes correlated only moderately (r = -0.45). CONCLUSION: Olanzapine was non-inferior to clozapine. The lack of a marked correlation between PANSS and SWN improvements indicates that patients and psychiatrists perceive treatment differently. PMID- 15667430 TI - Psychosocial predictors of depression in patients with acute coronary syndrome. AB - OBJECTIVE: To describe the prevalence of depression according to ICD-10 criteria using a self-completed questionnaire and to identify psychosocial predictors of depression at discharge in patients with acute coronary syndrome. METHOD: A total of 899 patients with acute coronary syndrome completed the Major Depression Inventory at discharge and a questionnaire regarding previous depression and family history of depression. Information concerning civil status was obtained from the Civil Person Registry. RESULTS: Ninety patients (10%) were depressed according to ICD-10 criteria at discharge with 7.2% having a moderate to severe depression at discharge. Women were significantly more frequently and severely depressed than men. Patients with and without depression reported primarily somatic symptoms of depression. Cardiovascular risk factors or treatment did not differ between patients with and without depression. Previous depression (OR 2.9, 95% CI 1.4-6.0 adjusted) and female gender (OR 2.5, 95% CI 1.5-4.3 adjusted) predicted depression at discharge in a logistic regression model. CONCLUSION: Somatic symptoms of depression are prevalent in patients with acute coronary syndrome. The use of self-completed non-diagnostic questionnaires assessing symptoms of depression therefore is cautioned as patients may wrongly be identified as depressed. In patients with acute coronary syndrome depression is predicted by well-known psychosocial risk factors. PMID- 15667431 TI - Recovery from major depression: the role of support from family, friends, and spiritual beliefs. AB - OBJECTIVE: Many of the risk factors for major depression are not amenable to change. The present study was designed to identify factors associated with recovery from depression that could be targets for clinical intervention. METHOD: Sixty-two psychiatric in-patients who met diagnostic criteria for major depression were interviewed while hospitalized and re-interviewed 3 months after discharge. Analyses examined the relationship between depression and three sources of emotional support: family, friends, and spiritual beliefs. RESULTS: Depression severity at baseline was the most consistent predictor of depression severity and diagnosis at follow-up. Patients who had recovered from depression by the time of the follow-up assessment reported higher perceived emotional support from family and friends at baseline. Support from friends, support from family and a composite of emotional support were significant predictors of depression beyond the effects of initial depression severity. CONCLUSION: Aspects of emotional support were significantly associated with depression outcome. PMID- 15667432 TI - Predicting the onset of major depression in subjects with subthreshold depression in primary care: a prospective study. AB - OBJECTIVE: That subjects with subthreshold depression have an increased probability of developing major depression has been confirmed by many studies. However, the factors which may predict the onset of major depression have yet to be fully examined. METHOD: We examined the control group of a randomized trial in primary care patients with subthreshold depression (N = 109), of whom 20 had developed major depression 1 year later. Using the vulnerability-stress theory, we examined which factors predicted the onset of major depression. RESULTS: In both univariate and multivariate analyses, family history and chronic illnesses predicted the onset of major depression. CONCLUSION: It is possible to predict to a certain degree whether a subject with subthreshold depression will develop major depression within a year. PMID- 15667433 TI - Progressing a spectrum model for defining non-melancholic depression. AB - OBJECTIVE: To further develop a 'spectrum model' for non-melancholic disorders that encompasses underlying personality styles and clinical patterning. METHOD: In a sample of patients with non-melancholic depression, we studied four personality constructs influencing risk to depression, assessing associational strength and specificity between personality scores and symptom and coping response patterns. RESULTS: Analyses refined four personality dimensions (anxious worrying, irritability, social inhibition, and self-centredness) for testing the model. For all dimensions, personality style was specifically linked with a mirroring 'coping' response. Quantification of specific links allowed development of a spectrum model for the non-melancholic depressive disorders in which underpinning personality style showed some specific links with the clinical 'pattern' of symptoms and coping repertoires. CONCLUSION: The model has the capacity to assist clinical assessment, identify aetiological personality influences and allow specific treatment effects for the heterogeneous non melancholic depressive disorders to be determined. PMID- 15667434 TI - Validation of the Bech-Rafaelsen Melancholia Scale and the Hamilton Depression Scale in patients with major depression; is the total score a valid measure of illness severity? AB - OBJECTIVE: Evaluation of antidepressant drug efficacy requires adequate rating scales for measuring the severity of depression. However, to measure the illness severity by such a total score, the scale needs to fulfil criteria of unidimensionality. On this background, we aimed at comparing the unidimensionality of the Bech-Rafaelsen Melancholia Scale (MES) and the 17-item Hamilton Depression Rating Scale (HAM-D(17)). METHOD: A total of 1629 patients aged between 18 and 65 years with a major depressive episode were treated openly with sertraline at a fixed oral dose of 50 mg daily during 4 weeks. The HAM-D(17) and the MES were applied at baseline and at weeks 2 and 4. Unidimensionality was tested with Mokken and Rasch analysis. RESULTS: Unidimensionality of the HAM D(17) could not be confirmed. However, the 6-item Hamilton Depression Subscale (HAM-D(6)), was accepted by the Rasch analysis both at baseline and after 2 and 4 weeks of therapy. For the MES (as well as for the HAM-D(6)), a Loevinger coefficient of homogeneity above 0.40 (suggesting acceptance) was found at week 4. CONCLUSION: The HAM-D(6) and the MES did fulfil criteria for unidimensionality while the HAM-D(17) did not. Therefore, the extended use of the HAM-D(17) in drug trials may be questioned. PMID- 15667435 TI - Selective cognitive deficits in obsessive-compulsive disorder compared to panic disorder with agoraphobia. AB - OBJECTIVE: Visual-spatial and executive functions deficits have been reported in obsessive-compulsive disorder (OCD). We investigated their specificity comparing cognitive function in OCD, panic disorder with agoraphobia (PD/A) and controls by a comprehensive neuropsychological battery. METHOD: Fifty-five subjects (25 OCD, 15 PD/A, 15 controls) without current depressive episode underwent structured clinical interview for DSM-IV, Yale-Brown Obsessive Compulsive Scale, Hamilton Anxiety Rating Scale, Hamilton Depression Rating Scale. Neuropsychological battery assessed: executive functions, visual discrimination, spatial memory and learning, verbal memory, general intellectual functioning. RESULTS: OCD showed controlled fluency, visual-spatial construction, learning and memory deficits; PD/A spatial learning impairment. OCD was discriminated from PD/A and controls by three tests scores, predicting group membership for 76.4% of the cases. CONCLUSION: Visual-constructive and controlled fluency deficits seem specific in OCD, while the spatial learning deficit, shared with PD patients, may not be disorder-specific, but anxiety-related. Results support the proposed ventral frontal-striatal circuit involvement in OCD. PMID- 15667436 TI - Obsessive-compulsive symptoms in adults with history of rheumatic fever, Sydenham's chorea and type I diabetes mellitus: preliminary results. AB - OBJECTIVE: Rheumatic fever (RF) associated with Sydenham's chorea (a neurological variant of RF), but not RF without chorea, has been acutely related to obsessive compulsive symptomatology/disorder (OCS/OCD). This study investigated the presence of OCS in adults who had RF with or without chorea in childhood. METHOD: The Yale-Brown Obsessive-Compulsive Scale (Y-BOCS) was used to evaluate OCS in 38 adults with history of RF (13 with chorea; 25 without chorea) or diabetes (controls; n = 19). RESULTS: The OCS was similar in both groups, although the intensity of symptoms was not clinically relevant. Moreover, subjects with RF with or without chorea did not score differently in the Y-BOCS. CONCLUSION: The similar occurrence of OCS in patients with history of RF and diabetes suggests that the development of this symptomatology, triggered by group A beta-haemolytic streptococcus infections, is restricted to the RF acute phase, occurred during infancy, and did not seem to predispose the appearance of OCS in adulthood. PMID- 15667438 TI - Lycanthropy and delusional misidentification. PMID- 15667440 TI - Effectiveness of amisulpride augmentation of clozapine in a non-responder to either drug alone: a case report. PMID- 15667449 TI - Intracerebroventricular injections of prolactin counteract the antagonistic effect of bromocriptine on rabbit maternal behaviour. AB - To investigate the participation of prolactin in nest-building and maternal behaviour in rabbits, we administered (from pregnancy day 26 to parturition) rabbit prolactin (rbPRL; or vehicle) intracerebroventricularly (i.c.v.) to primiparous animals injected with bromocriptine subcutaneously (s.c.). Control females (given vehicle s.c. and i.c.v.) built a maternal nest (of straw and body hair) in 77% of cases. This proportion decreased to 19% in the bromocriptine-only group (P < 0.05) and increased to 93% in the group given bromocriptine plus rbPRL (P > 0.05). Maternal behaviour (i.e. the adoption of a crouching posture over the litter inside the nest box) was expressed by 77% of control rabbits, 19% of bromocriptine-only animals (P < 0.05) and 57% of females given bromocriptine plus rbPRL (P > 0.05). Values of nonmaternal activities (i.e. scent-marking, ambulation in an open field) were similar among the three studied groups. These results suggest that prolactin, acting in late pregnancy, plays a major role in the stimulation of nest-building and maternal behaviour in rabbits. PMID- 15667450 TI - Sex differences in the distribution and abundance of androgen receptor mRNA containing cells in the preoptic area and hypothalamus of the ram and ewe. AB - Rams and ewes show a negative-feedback response to peripheral treatment with testosterone, with both sexes having a similar degree of suppression in luteinizing hormone (LH) secretion during the breeding season. At least part of the action of testosterone to suppress gonadotropin-releasing hormone/LH secretion is exerted via interaction with an androgen receptor. The distribution of androgen receptor-containing cells in the hypothalamus has been described for the ram, but similar studies have not been performed in the ewe. In the present study, we tested the hypothesis that levels of androgen receptor mRNA expression in the preoptic area and hypothalamus would be similar in rams and ewes. Perfusion-fixed brain tissue was obtained from adult Romney Marsh ewes (luteal phase) and rams during the breeding season (n = 4/sex). Androgen receptor mRNA expression was quantified in hypothalamic sections by in situ hybridization using an (35)S-labelled riboprobe and image analysis. Hybridizing cells were found in the medial preoptic area, bed nucleus of the stria terminalis, anterior hypothalamic area, ventromedial nucleus, arcuate nucleus and premamillary nucleus. The level of androgen receptor mRNA expression was higher in rams than ewes in the rostral preoptic area, caudal preoptic area and rostral portion of the bed nucleus of the stria terminalis, with no sex difference in other regions. The preoptic area and bed nucleus of the stria terminalis are important for reproductive behaviour and the sex differences in androgen receptor mRNA expression at these levels may relate to this. The high level of androgen receptor mRNA expression in the basal hypothalamus, with no sex difference, is consistent with the role of this region in the regulation of gonadotropin secretion. PMID- 15667451 TI - Sex and region-specific regulation of oestrogen receptor beta in the rat hypothalamus. AB - Sexual dimorphism in the expression of oestrogen receptor (ER)beta mRNA and protein was characterized in the rostral forebrain of the rat and its dependence on the neonatal endocrine environment was revealed. We present novel data demonstrating, in gonadectomized adult rats, that the amount of oestrogen caused a significant reduction in the number of ERbeta messages and protein in the ventromedial nucleus in both sexes, but no such effects were detected in the preoptic area or the amygdala. In gonadectomized females, more so than in males, the ventromedial nucleus of the adult rat contained a significantly larger number of ERbeta-positive neurones both in terms of ERbeta mRNA and protein. In the juvenile rat on day 14, sex difference in ERbeta expression was already observed in the ventromedial nucleus. Treatment of neonatal females with oestrogen from days 1-10 or neonatal orchidectomy of males reversed the sex difference in the ventromedial nucleus when observed on day 14, showing that the neonatal presence of oestrogen had caused irreversible masculinization of this structure. Our results suggest that sex-specific expression of ERbeta is patterned by perinatal hormone exposure: down-regulation of ERbeta caused by oestrogen in a region specific manner. PMID- 15667452 TI - Fos expression is selectively and differentially regulated by endogenous glucocorticoids in the paraventricular nucleus of the hypothalamus and the dentate gyrus. AB - We examined the extent to which basal levels of corticosterone, which vary in a circadian fashion, influence the pattern of Fos protein expression in the paraventricular nucleus of the hypothalamus (PVN), the hippocampal formation and three different functional cortical areas. Basal and poststress (1 h of restraint) Fos expression, as determined by immunohistochemistry, was examined in male rats with either no previous surgical manipulation or in rats 5 days after: (i) sham adrenalectomy; (ii) adrenalectomy with no corticosterone replacement; or (iii) adrenalectomy with corticosterone (25 microg/ml) in the drinking water replacement. In adrenal-intact rats, restraint produced similar patterns of Fos expression in the PVN, cortical areas and hippocampus (CA1-CA3), with peak levels of expression attained 60-90 min after restraint onset. Surprisingly, in the dentate gyrus, there was a dissociation between the two blades in the pattern of Fos expression after restraint. In the inner blade (suprapyramidal), there was a delayed induction that occurred between 60 and 90 min after restraint onset and, in the outer blade (infrapyramidal), there was a steady decline in Fos expression after restraint. Adrenalectomy had an effect on Fos expression only in the PVN and dentate gyrus, and the nature of the effect was quite different for both brain regions. In the PVN, adrenalectomy had no effect on Fos expression in unstressed rats, but resulted in an enhanced number of Fos positive cells after restraint. In the dentate gyrus, adrenalectomy resulted in an overall reduction of Fos positive cells in both blades, and this reduction was present in unstressed and stressed rats. Corticosterone replacement normalized the adrenalectomy effect on Fos expression in both brain regions. Thus, Fos expression in the rat brain displays specific patterns of dependency on the permissive effects of glucorticoids, and this dependency varies between brain regions. PMID- 15667453 TI - Plasma and hypothalamic peptide-hormone levels regulating somatotroph function and energy balance in fed and fasted states: a comparative study in four strains of rats. AB - Both growth hormone (GH)/insulin growth factor (IGF)-1 axis and energy balance have been implicated in longevity independently. The aim of the present study was to characterize the effect of a 72-h fasting period at 3 months of age in four different rat strains: (i) Wistar and (ii) Fischer 344 rats, which develop obesity with age, and (iii) Brown Norway and (iv) Lou C rats, which do not. Wistar rats ate more, were significantly bigger, and presented with higher plasma leptin and lower ghrelin levels and hypothalamic growth hormone-releasing hormone (GHRH) content than rats from the three other strains. Plasma insulin and IGF-1 levels were lower in Brown Norway and Lou C rats, and somatostatin content was lower in Brown Norway rats only. Glycaemia was lower in Lou C rats that displayed a lower relative food intake compared to Fischer and Wistar rats. Brown Norway rats showed a greater caloric efficiency than the three other strains. Concerning major hypothalamic neuropeptides implicated in feeding, similar amounts were detected in the four strains for neuropeptide Y, agouti-related peptide, galanin, melanin-concentrating hormone, alpha-melanocortin-stimulating hormone (alpha-MSH) and corticotropin-releasing hormone. Orexin A appeared to be slightly elevated in Fischer rats and cocaine amphetamine-regulated transcript (CART)(55-102) diminished in Brown Norway. At the mRNA level, orexin A, GHSR1, alpha-MSH and CART expression were higher in Wistar and Lou C rats. Principal component analysis confirmed the presence of two main factors in the ad libitum rat population; the first being associated with growth-related parameters and the second being associated with food intake regulation. Hypothalamic GHRH and somatostatin content were positively correlated with feeding-related neuropeptides such as alpha-MSH for GHRH, and orexin A and CART for both peptides. Plasma ghrelin levels were negatively correlated with leptin and IGF-1 levels. Finally, a 72-h fasting period affected minimally body weight, plasma IGF 1 and leptin levels in Lou C rats compared to the three other strains, and plasma insulin levels were less affected in Brown Norway rats. In conclusion, Wistar shorter life span is consistent with its already fatter phenotype at 3 months of age. In terms of IGF-1, glycaemia and leptin responses to fasting, the Lou strain, which presents with a low food intake/body weight and caloric efficiency, is the least affected. The link between food intake regulation, GH axis and ageing is further demonstrated by principal component analysis, where GHRH and somatostatin were found to be strongly associated with energy homeostasis parameters. PMID- 15667454 TI - Gonadal steroid replacement reverses gonadectomy-induced changes in the corticosterone pulse profile and stress-induced hypothalamic-pituitary-adrenal axis activity of male and female rats. AB - We investigated the effects of gonadal hormone replacement on the pulsatile parameters underlying basal circadian corticosterone secretion in castrated male and ovariectomized female rats using an automated sampling system. Blood was collected from freely moving, unanaesthetized rats every 10 min over a 24-h period and sampling was continued during a noise stress and after lipopolysaccharide (LPS) administration. Castrated male rats had markedly higher corticosterone levels than intact controls. This was reflected by increased number and frequency of pulses in addition to an increase in the pulse height and amplitude under both basal circadian and stress conditions. Hormone replacement with either testosterone or dihydrotestosterone returned these corticosterone levels and circadian profile to those found in intact males, confirming an androgen-mediated effect. Ovariectomized females had significantly lower basal and stress-induced corticosterone levels with lower frequency and amplitude of corticosterone pulses than intact females. 17beta-oestradiol replacement returned basal levels, pulsatile measurements and stress-induced corticosterone levels to those found in intact females. Three hours post-LPS administration, castrated males demonstrated significantly higher values of parvocellular paraventricular nucleus (PVN) arginine vasopressin and corticotrophin-releasing factor and anterior pituitary pro-opiomelanocortin mRNA while ovariectomized females showed significantly lower levels of all three transcripts compared to intact controls. PVN glucocorticoid receptor mRNA levels 3 h post-LPS administration were significantly decreased in castrated males and significantly increased in ovariectomized female rats. Replacement of gonadal steroids resulted in a return to the levels found in intact controls after LPS. Gonadal steroid replacement is sufficient to reverse changes in the pulsatile characteristics of corticosterone release after gonadectomy. In addition, gonadal steroid replacement reverses stress-induced alterations in hypothalamic-pituitary-adrenal (HPA) activity. These data demonstrate a major contribution of gonadal steroids to the regulation of HPA axis activity and to the pulsatile characteristics of corticosterone release. PMID- 15667455 TI - Gonadotrophin inhibitory hormone depresses gonadotrophin alpha and follicle stimulating hormone beta subunit expression in the pituitary of the domestic chicken. AB - Studies performed in vitro suggest that a novel 12 amino acid RF amide peptide, isolated from the quail hypothalamus, is a gonadotrophin inhibitory hormone (GnIH). The aim of the present study was to investigate this hypothesis in the domestic chicken. Injections of GnIH into nest-deprived incubating hens failed to depress the concentration of plasma luteinizing hormone (LH). Addition of GnIH to short-term (120 min) cultures of diced pituitary glands from adult cockerels depressed follicle-stimulating hormone (FSH) and LH release and depressed common alpha and FSHbeta gonadotrophin subunit mRNAs, with no effect on LHbeta subunit mRNA. Hypothalamic GnIH mRNA was higher in incubating (out-of-lay) than in laying hens, but there was no significant difference in the amount of hypothalamic GnIH mRNA in out-of-lay and laying broiler breeder hens at the end of a laying year. It is concluded that avian GnIH may play a role in controlling gonadotrophin synthesis and associated constitutive release in the domestic chicken. PMID- 15667456 TI - The neurobiology of social bonds. AB - When released in the brain through giving birth or mating, the neuropeptides oxytocin and vasopressin are involved in promoting parent-offspring and monogamous bonds in animals such as sheep and voles. Bonds are only formed in species where receptors for these neuropeptides are highly expressed in dopamine producing reward centres. In humans, dysfunctions in these same systems can be associated with autism and, when we see people we love, these systems become activated. PMID- 15667459 TI - Cochrane Collaboration: more than systematic reviews? PMID- 15667462 TI - Landmark intensive care study. PMID- 15667463 TI - Engaging Australian physicians in evidence-based medicine: a representative national survey. AB - AIM: To assess Australian adult physicians' views about evidence-based medicine (EBM) and quality improvement (QI). BACKGROUND: Cross-sectional postal survey of two hundred and forty-four randomly selected Australian physicians (78.5% response rate). METHOD: Physicians' views about the promotion of EBM and QI and their impact on patient care, strategies to support better clinical practice and self-reported understanding of EBM terms. RESULTS: Sixty-eight per cent (95% confidence interval (CI): 62-74%) of physicians had a positive view of the current promotion of EBM in Australia. Significantly fewer (45%; 95% CI: 38-51%) were so positive about QI (P <0.001). Although 74% (95% CI: 68-79%) indicated that EBM improves patient care, significantly fewer (46%; 95% CI: 40-53%) held this view with respect to QI (P <0.001). Although 70% (95% CI: 64-76%) of physicians agreed they had adequate skills to search for evidence, significantly fewer (21%; 95% CI: 17-27%) agreed they had sufficient time to do so (P <0.001). EBM skills were positively associated with involvement in teaching. Physicians were significantly more positive about approaches exclusive to themselves compared with those involving other disciplines. CONCLUSIONS: Although Australian physicians have positive views of EBM, QI appears less well regarded. Initiatives to improve quality and safety that incorporate principles and language of EBM will likely be received better by physicians than isolated QI. Further enhancement of EBM requires concomitant attention to physician training, workplace infrastructure and supportive professional development. PMID- 15667464 TI - Haemopoietic stem cell transplantation in Australia and New Zealand, 1992-2001: progress report from the Australasian Bone Marrow Transplant Recipient Registry. AB - BACKGROUND: Bone marrow and blood stem cell transplantation is now used as curative therapy for a range of haematological malignancies and other conditions. The Australasian Bone Marrow Transplant Recipient Registry (ABMTRR) has recorded transplant activity in Australia since 1992; transplant centres in New Zealand have corresponded with the Registry since 1998. AIM: To describe allogeneic and autologous bone marrow and blood stem cell transplantation activity and outcomes in Australia and New Zealand from 1992 to 2001. METHODS: Each haemopoietic stem cell transplant centre in Australia and New Zealand contributes information to the Registry via a single information form compiled when a transplant is performed. An annual follow-up request is then sent from the Registry to the contributing centre at the anniversary of each individual transplant. RESULTS: Haemopoietic stem cell transplants in Australia have increased in number from 478 in 1992 to 937 in 2001, whereas in New Zealand the number has grown from 91 in 1998 to 105 in 2001, mainly as a result of an increase in autologous blood stem cell transplants. The number of hospitals contributing to the ABMTRR has grown from 20 in 1992 to 37 in 2001. The most common indication for autologous transplantation in 2001 was non-Hodgkin's lymphoma, whereas for allogeneic transplants it was acute myeloid leukaemia. The 9-year actuarial disease-free survival probability for patients aged 16 and above between 1992 and 2000 was 37% for autologous, 39% for allogeneic related donor and 30% for allogeneic unrelated donor transplants. Recurrence of the underlying disease was the main cause of death post-transplant after both allogeneic (26.3% of deaths in the first year and 68.0% of deaths in the second year) and autologous transplants (59.0% and 86.2%). Treatment-related mortality was 16.9% after allogeneic transplantation and 2.1% after autologous transplantation in 2000. CONCLUSIONS: The ABMTRR provides a comprehensive source of information on the use of bone marrow transplant, and allows for continuing analysis of changes in the application of this high-cost technology and the outcome of patients undergoing these procedures. Registry data provide a means for directing future clinical research into perceived areas of priority for improvement of outcome, such as the reduction in the risk of disease recurrence post-transplant. PMID- 15667465 TI - Isolated pulmonary hypertension in scleroderma. AB - BACKGROUND: Isolated pulmonary hypertension (PHT) is now the most frequent cause of disease-related death in limited cutaneous scleroderma, the commonest disease variant of this disabling connective tissue disorder. Endothelin-1 receptor antagonists provide symptomatic benefit but to date have not been shown to prolong survival. AIM: To determine the frequency, disease characteristics and survival of symptomatic patients with isolated PHT in our cohort of scleroderma patients. METHODS: Systematic review of the clinical course of all patients registered on the South Australian Scleroderma Register, a population-based register of 374 living and 234 deceased patients with scleroderma. RESULTS: Thirty-four patients were identified with isolated PHT, the majority with limited scleroderma. From our deceased register, we estimate that >11% of patients with this limited variant will develop this complication. Isolated PHT occurs as a late-stage complication approximately 20 years after the first symptoms of scleroderma. Patients with isolated PHT were characterized by the presence of multiple telangiectasia, reduced nailfold capillary density, digital ulceration, gross reduction of diffusing capacity for carbon monoxide and echocardiographic evidence of elevated pulmonary artery pressure. Survival was significantly shortened as compared with those patients without this complication (P=0.002), with a mean survival of 2.5 years from symptomatic onset of PHT. CONCLUSION: Isolated PHT occurs as a late-stage complication in > or =11% of patients with limited cutaneous scleroderma and leads to rapid death from right heart failure. The early use of endothelin-1 receptor antagonists may change the natural history of this fatal complication. PMID- 15667466 TI - Outpatient treatment with subcutaneous interleukin-2, interferon alpha and fluorouracil in patients with metastatic renal cancer: an Australian experience. AB - BACKGROUND: Metastatic renal cell cancer has a poor prognosis and survival. Conventional cytotoxic chemotherapy has no impact on survival and response rates are low. Biologic agents are the most active in treating this disease. We report the feasibility of administering a combination of interferon alpha, subcutaneous interleukin-2 and 5Fluorouracil in the outpatient setting to patients with metastatic renal cell cancer. RESULTS: Between September 1996 and August 2003, fourteen patients were treated with this combination: ten males and four females with a median age of 50 (42-66). Thirteen patients had Eastern Cooperative Oncology Group performance scores of 0 or 1. Ten patients had had nephrectomies. Six patients had undergone prior treatments with chemotherapy or hormonal therapy. Twenty-two cycles were administered (median 1, range of 1-4). Three patients achieved partial response, eight patients had stable disease, and three had progressive disease. The duration of response in patients with stable disease was (3, 3+, 4, 4+, 5+, 6, 10, 11 months) and for the patients with a partial response was 2+, 11 and 12 months. Toxicities with this combination were predictable. There were no treatment-related deaths and no episodes of febrile neutropenia. One patient ceased treatment as a result of toxicity. Fatigue was the most common side-effect. Myalgias, fever and rigors occurred within 6-12 h of administration of interleukin-2, and resolved within 12 h. Grade 1-2 nausea and vomiting occurred in most patients. Four patients had transient asymptomatic transaminitis, which resolved spontaneously. As a result of toxicity, one patient had treatment ceased in his 6th week. CONCLUSION: This combination was feasible, well tolerated and manageable in an outpatient setting. PMID- 15667467 TI - Prospective audit of short-term concurrent ketamine, opioid and anti-inflammatory ('triple-agent') therapy for episodes of acute on chronic pain. AB - AIM: This prospective audit was undertaken in order to document the analgesic response and adverse effects of concurrent short-term ('burst') triple-agent analgesic (ketamine, an opioid and an anti-inflammatory agent--either steroidal or non-steroidal) administration, for episodes of acute on chronic pain. The clinical hypothesis in this study is that better pain control may be obtained by simultaneous multiple target receptor blockade. METHOD: The response of 18 patients is reported. The pain and analgesic requirement data for the 24 h before starting triple-agent therapy were compared with the last 24 h on the triple agent therapy. Patients were then classified as responders or non-responders. RESULTS: According to stringent clinical criteria, 12 out of the 18 patients were classified as responders. The response rate was highest for somatic pain (7/9) and appeared to decrease with duration of prior uncontrolled pain. Only four out of the 18 patients reported adverse effects and all of these were minor. CONCLUSIONS: The results suggest that this 'burst' triple-agent approach is safe and effective in an inpatient palliative care population during episodes of poorly controlled acute on chronic pain, and warrants further investigation to ascertain whether it gives superior results compared to the 'gold-standard' WHO ladder approach. PMID- 15667468 TI - Modern management of portal hypertension. AB - The development of portal hypertension plays a major role in the pathogenesis of many of the complications of chronic liver disease. In developed countries, most patients with portal hypertension have cirrhosis, and, in this condition, portal pressure is elevated as a result of both an increase in hepatic resistance to portal perfusion and increased mesenteric blood flow. Bleeding from oesophageal varices is a major cause of mortality in patients with significant portal hypertension. This review concentrates on the recognition, prevention and acute management of this life threatening complication of cirrhosis. PMID- 15667469 TI - Respiratory infections during air travel. AB - An increasing number of individuals undertake air travel annually. Issues regarding cabin air quality and the potential risks of transmission of respiratory infections during flight have been investigated and debated previously, but, with the advent of severe acute respiratory syndrome and influenza outbreaks, these issues have recently taken on heightened importance. Anecdotally, many people complain of respiratory symptoms following air travel. However, studies of ventilation systems and patient outcomes indicate the spread of pathogens during flight occurs rarely. In the present review, aspects of the aircraft cabin environment that affect the likelihood of transmission of respiratory pathogens on airplanes are outlined briefly and evidence for the occurrence of outbreaks of respiratory illness among airline passengers are reviewed. PMID- 15667470 TI - Mediating telemedicine: ethics at a distance. PMID- 15667471 TI - Radiocontrast anaphylaxis with failure of premedication. AB - Radiocontrast media (RCM) is used commonly in clinical practice, and can be associated with significant adverse effects. We report a patient who experienced severe anaphylaxis after being given multiple drugs. Challenge testing established allergy to both RCM and ceftriaxone. Premedication did not prevent recurrence of anaphylaxis on repeat challenge with RCM. The haemodynamic and serum tryptase consequences of the challenges are discussed, and a summary of RCM allergy is provided. PMID- 15667473 TI - Manifestations of familial hypercholesterolaemia. PMID- 15667472 TI - Scleroderma in Australian aborigines. AB - Scleroderma (systemic sclerosis) has not been reported before in Australian Aborigines. We describe in detail a community middle-aged Aboriginal woman whose diffuse scleroderma terminated fatally with a renal crisis. Moreover, we have identified a further five Aboriginal patients on the South Australian Scleroderma Register (two with diffuse, two with limited and one with overlap scleroderma), a number consistent with that expected from the 2001 census data for our state. However, an analysis of all antinuclear antibody (ANA) requests from the Top End of Australia over a 6-year period revealed only two Aborigines with low titre anticentromere antibody (despite frequent occurrence of ANA with other specificities). Neither of these Aborigines had features of scleroderma. In conclusion, scleroderma does occur in indigenous Australians but further studies are needed to confirm the apparent infrequency of centromere-associated limited scleroderma (which is the commonest form of scleroderma in our Caucasian population). PMID- 15667474 TI - These are a few of my favourite things: reflections on the doctor-doctor relationship. AB - The doctor-patient relationship is often spoken about and is typically characterized by one of trust and compassion. However, the doctor-doctor relationship is rarely discussed openly as it is too hurtful to reveal the truth. Lack of compassion, competitiveness and even cruelty abounds in the opinion of the author who exemplifies this by some personal anecdotes and is based on 30 years as a physician. Unfortunately the lessons to be learned occur too late for most individuals. Selection of medical graduates may also play an important role in the whole malady. PMID- 15667475 TI - Exogenous glucocorticoid excess as a result of ritonavir-fluticasone interaction. PMID- 15667476 TI - Acalculous cholecystitis, multifocal gastrointestinal infarction and pancreatitis resulting from Varicella-zoster virus. PMID- 15667477 TI - Perirectal cellulitis caused by Streptococcus pyogenes. PMID- 15667478 TI - Gender differences in gastroenterology. AB - Balancing professional and personal life can be particularly challenging for young women at the prime of their reproductive years. Young female gastroenterologists were found to work for larger groups, take fewer calls, and receive less compensation than their male counterparts. Getting more women into gastroenterology will be an achievable goal if the needs of female trainees are understood and met. PMID- 15667479 TI - Gender disparity in the practice of gastroenterology: the first 5 years of a career. AB - BACKGROUND: Despite an unprecedented demand for gastroenterology services, the number of gastroenterology trainees has decreased over 50% since 1993. Women comprise nearly 50% of the U.S. medical school student population; yet only occupy 16% of gastroenterology fellowship positions. In order to recruit the best candidates to gastroenterology we must be able to demonstrate the attractiveness of a career in the field. A prospective study was performed to identify the career choices of graduates from gastroenterology fellowship programs using a prospective study model and to identify whether gender differences exist in the practice of gastroenterologists up to 5 yr after completion of training. METHODS: A survey gathering information on demographics, practice pattern, and income was mailed to two cohorts of gastroenterology fellows 3 and 5 yr after graduation. RESULTS: A total of 247 subjects completed the 3 yr and 220 subjects responded to the 5-yr survey. At 3 yr, men reported higher income (p < 0.001), worked longer hours per week (p < 0.002), and were more likely to be part owner of the practice (p= 0.027). Females reported fewer children (p < 0.007), lower board certification rates (p < 0. 002), worked for larger, multispecialty practices (p < 0.001), and practiced more internal medicine. These differences were still present at 5 yr into gastroenterology practice. CONCLUSIONS: Significant differences in practice type, earnings, board certification, professional standing, and alterations in family planning are noted between male and female gastroenterologists in the initial 5 yr of their practice. PMID- 15667480 TI - Reproducibility of multichannel intraluminal electrical impedance monitoring of gastroesophageal reflux. AB - OBJECTIVE: Esophageal impedance measurement is a novel method for gastroesophageal reflux monitoring. Reproducibility is an important aspect of every biomedical test. The aim of this study was therefore to asses the reproducibility of gastroesophageal reflux monitoring using impedance measurements. METHODS: Impedance and pH signals were recorded in 20 healthy volunteers during 90-min postprandial periods on two separate days. Hourly rates of gas, liquid, and mixed gas-liquid reflux episodes were measured in each recording period as well as percentage of time with pH < 4 and rate of acid reflux episodes. As a quantitative description of inter- and intraindividual variation for each variable, the mean percentage of covariation (100 x SD/mean: %COV) was calculated. As a second measure for reproducibility, Kendall's coefficients of concordance (W values) were calculated. RESULTS: For all variables, interindividual %COV was at least 50% higher than intraindividual %COV. Statistically significant concordances were found for gas reflux (W = 0.81, p= 0.04) and mixed reflux (W = 0.85, p= 0.03) while concordance for liquid reflux tended to be significant (W = 0.75, p= 0.08). This was comparable to the reproducibility of the number of acid reflux episodes and percentage of time with pH < 4 (W = 0.78, p= 0.05 and W = 0.88, p= 0.02, respectively). CONCLUSIONS: Postprandial gastroesophageal reflux data assessed with impedance monitoring are as reproducible as assessed with pH monitoring. PMID- 15667481 TI - Long-term outcome of pneumatic dilation in the treatment of achalasia. AB - OBJECTIVE: Achalasia is a well-defined esophageal motor disorder for which pneumatic dilation is an established therapeutic method. Even though it has been used for several years, there are limited data on the long-term outcomes of patients treated with this procedure. Hence, we aimed to evaluate the long-term efficacy of pneumatic dilation to control the symptoms of achalasia. METHODS: The medical records of all patients treated in our unit for achalasia with pneumatic dilation were reviewed. We identified the long-term result of the initial procedure, the date of the first dilation, and the time interval between dilation and retreatment. RESULTS: Of 260 patients who were treated with pneumatic dilation, 153 (67 men, 86 women) were followed up for more than 5 yr. The mean follow-up period was 11.09 +/- 3.91 yr, and the success rate of the dilation was 75.8%. Among these patients, 35 (19 men, 16 women) had follow-up periods of more than 15 yr. The mean follow-up time of those patients was 16.56 +/- 1.09 yr, and the success rate was 51.4%. Kaplan-Meier survival analysis showed that, overall, 50% of patients develop recurring symptoms after 10.92 yr. CONCLUSIONS: Although 51.4% of patients continued to be in clinical remission more than 15 yr after the initial pneumatic dilation, the long-term success rate of pneumatic dilation seems to drop progressively with time. PMID- 15667482 TI - Lugol's dye spray chromoendoscopy establishes early diagnosis of esophageal cancer in patients with primary head and neck cancer. AB - OBJECTIVE: Patients with primary head and neck cancer show a predisposition to develop esophageal cancer. The aim of this study was to investigate in these patients: the prevalence of esophageal cancer comparing the value of chromoendoscopy using Lugol's solution examination to standard endoscopy, in the early diagnosis of esophageal cancer. METHODS: Prospective observational study at a state general university hospital in Sao Paulo, Brazil. 326 consecutive adult patients with primary head and neck cancer were evaluated. A standard endoscopy was performed, followed by a 2% Lugol's dye spray chromoendoscopy and histopathologic study. The prevalence of esophageal cancer was defined. The results of the two endoscopic methods were compared. RESULTS: Twenty-four patients with esophageal cancer and high-grade intraepithelial neoplasia were detected and had a prevalence of 7.36%. Chromoendoscopy and standard endoscopy were equivalent to the diagnosis of advanced and invasive esophageal cancer. However, standard endoscopy diagnosed 55% of high-grade intraepithelial neoplasia, in comparison to chromoendoscopy that detected 100%. CONCLUSIONS: Patients with primary head and neck cancer should be considered as high risks for the presence of esophageal cancer. Lugol's dye chromoendoscopy diagnosed high grade intraepithelial neoplasia, which went unnoticed with standard endoscopy. It permits a more exact detection of lesion boundaries and facilitates a more precise targeting of biopsy fragments. PMID- 15667483 TI - The role of esophageal pH monitoring in symptomatic patients on PPI therapy. AB - BACKGROUND: Ambulatory pH monitoring while on therapy is often recommended in gastroesophageal reflux disease (GERD) patients with continued symptoms. However, to date, little data exist to justify this indication. AIM: To assess the role of pH monitoring in symptomatic patients despite aggressive therapy with typical or extra esophageal GERD. METHODS: Retrospective review of 2,291 ambulatory pH tracings (1999-2003) identified subgroup of studies performed on proton pump inhibitor (PPI) therapy. Patients with prior fundoplication or Barrett's esophagus were excluded. Patients grouped on predominant presenting GERD symptoms: typical (heartburn and regurgitation) or extra esophageal (chest pain, cough, hoarseness, sore throat, shortness of breath, asthma). The distribution of abnormal pH parameters in each group calculated and univariate analyses assessed the probability of abnormal pH in each group. Abnormal cutoff values traditionally used in clinical practice and more stringent cutoff values used to determine distribution of abnormality as a function of cutoff values. RESULTS: A total of 250 patients (mean age 54.3 yrs, 59% female) underwent pH monitoring on either daily (b.i.d.) or twice daily (q.d.) on PPI therapy: 115 (46%) with extra esophageal and 135 (54%) with typical GERD symptoms. Extra esophageal GERD patients were more likely to undergo pH monitoring on b.i.d. PPIs (OR = 2.7; 95% CI = 1.6-4.4; p < 0.01). 52 (93%) of typical and 74 (99%) of extra esophageal GERD patients on b.i.d. PPIs tested normal. The odds of a normal pH values were 11 times higher for patients on b.i.d. PPIs (OR = 11.4; 95% CI = 4.3-30.1, p < 0.01) than those on q.d. PPIs. CONCLUSIONS: 1) The likelihood of an abnormal esophageal pH for symptomatic GERD patients on b.i.d. PPI is very small. 2) In this group of patients failing b.i.d. PPIs causes other than GERD should be sought. PMID- 15667484 TI - TNF gene polymorphisms and Helicobacter Pylori infection in gastric carcinogenesis in Chinese population. AB - BACKGROUND AND AIMS: Helicobacter pylori (H. pylori) is a major cause of chronic gastritis and peptic ulcer disease, and a definite carcinogen for gastric adenocarcinoma. However, the underlying pathogenic mechanisms have not been fully understood although the interactions between environmental, bacterial, and multiple genetic components are likely to be involved. Tumor necrosis factor (TNF) is a key cytokine involved in H. pylori-induced gastric inflammation. The present study aimed to determine the di-allelic polymorphisms of TNF gene and their association with H. pylori infection and gastroduodenal diseases in Chinese population of Han nationality. METHODS: Two hundred and ten patients with gastroduodenal diseases (73 chronic gastritis, 78 duodenal ulcer, and 59 noncardia gastric cancer) and 264 healthy controls were genotyped by the PCR-RFLP method for TNF-alpha 308, lymphotoxin-alpha (LT-alpha) NcoI, and AspHI gene polymorphisms. H. pylori infection status was determined by a validated serological test. RESULTS: H. pylori infection was detected in 90.5% of 210 patients and 62.1% of 264 healthy controls (p < 0.0001; odds ratio [OR]= 5.793; 95%CI: 3.431-9.780). Frequency of LT-alphaNcoI A/G genotype in patients with noncardia gastric cancer with H. pylori infection was significantly higher than that in H. pylori-positive healthy controls (64.0%vs 46.0%; p= 0.0297; OR = 2.026; 95%CI: 1.080-3.803). There were no other associations between TNF-alpha 308, LT-alphaNcoI, and AspHI gene polymorphisms and H. pylori infection in gastroduodenal diseases. CONCLUSIONS: LT-alphaNcoI A/G heterozygous genotype was associated with H. pylori infection in patients with noncardia gastric cancer in Chinese Han population. PMID- 15667485 TI - Marked elevation in serum transaminases: an atypical presentation of choledocholithiasis. AB - BACKGROUND: Choledocholithiasis causes elevations in levels of alkaline phosphatase out of proportion to aspartate aminotransferase (AST) and alanine aminotransferase (ALT). Isolated marked elevation in AST and ALT levels over 1,000 IU/L has been reported infrequently in patients with choledocholithiasis. METHODS: The charts of 18 patients who presented between 1971 and 2002 with documented choledocholithiasis and AST or ALT levels greater than 1,000 IU/L were retrospectively reviewed. An extensive work-up for coexisting disease processes to account for the abnormal AST and ALT levels was negative. RESULTS: Eighteen patients (16 women, 16 Hispanics, age 38 +/- 3 yr) presented with symptoms of choledocholithiasis and marked transaminase elevation. Peak levels of AST and ALT were 1,062 +/- 129 and 1,119 +/- 90, respectively. Following successful management of gallstone disease, AST and ALT levels fell rapidly to 129 +/- 22 and 268 +/- 61, respectively, within 3-14 days. There was also a concomitant improvement in the levels of bilirubin and alkaline phosphatase. CONCLUSIONS: In the absence of other hepatobiliary or pancreatic disease, choledocholithiasis can result in elevations in AST and/or ALT greater than 1,000 IU/L. These levels fall markedly once the gallstone disease is appropriately managed. PMID- 15667486 TI - Erythropoietic response to anemia in chronic hepatitis C patients receiving combination pegylated interferon/ribavirin. AB - OBJECTIVES: In hepatitis C virus (HCV)-infected patients receiving pegylated interferon (PEG-IFN)/ribavirin (RBV) combination therapy, anemia is a well-known side effect. The purpose of this study was to describe the time course and extent of hemoglobin (Hb) changes and the erythropoietic response to PEG-IFN/RBV-induced anemia. METHODS: In this multicenter, observational, 8-wk study, laboratory parameters were measured weekly for 8 wk or until early withdrawal. Primary endpoints included changes in Hb and serum erythropoietin (sEPO) from baseline to week 8; other measures were changes in reticulocytes and RBV dose. The predictive value of baseline factors for maximum Hb decline was assessed. RESULTS: In the 97 evaluable patients, mean Hb decreased from 14.4 +/- 1.4 g/dl (baseline) to 11.9 +/- 1.3 g/dl (week 8). Twenty-one percent of patients withdrew before week 8. The estimated erythropoietic response was lower than that seen in two historic control populations of iron deficiency anemia patients. Mean RBV dose decreased from 986 +/- 190 mg/day (baseline) to 913 +/- 228 mg/day (week 8). Fifty-seven out of 77 (74%) patients who completed the study maintained their initial prescribed RBV dose. Patients maintained on the initial dose of RBV who had a higher baseline Hb and viral load showed a trend toward larger Hb declines. Platelets and white blood cells (WBCs) also declined during the study. CONCLUSIONS: HCV-infected patients receiving PEG-IFN/RBV therapy have reductions in Hb, platelets, and WBCs, possibly due to bone marrow suppression. They also have diminished endogenous sEPO production for their degree of anemia. PMID- 15667487 TI - Mycophenolate mofetil for the treatment of primary sclerosing cholangitis. AB - Primary sclerosing cholangitis (PSC) is a chronic cholestatic liver disease of unknown etiology. Despite advances in understanding the pathophysiology underlying this disorder, no effective medical therapy has been identified for halting disease progression. The aim of this investigation was to determine the safety and estimated efficacy of mycophenolate mofetil (MMF) for the treatment of PSC. Thirty patients with PSC received MMF 1 g daily to a maximum of 3 g daily for 1 yr. Liver tests were determined at 3-month intervals with the Mayo risk score calculated at baseline and at the end of therapy. Twenty-three (77%) patients completed 1 yr of therapy. Significant but clinically marginal improvement in serum alkaline phosphatase level after 1 yr of therapy was observed (1135 +/- 581 U/L vs 912 +/- 463 U/L, p= 0.02). No other significant changes in liver biochemistries or Mayo risk score was observed. Seven patients (23%) discontinued MMF due to adverse events possibly related to therapy. Adverse reactions resolved spontaneously or with dose reduction in 10 (33%) patients. One patient developed pancreatitis, bacterial cholangitis, and sepsis during the eighth month of MMF therapy. No patient developed cytopenia on therapy. In conclusion, MMF does not appear to have clinically important benefits for PSC despite being tolerated by most patients. The results of this pilot study do not support further study of MMF as a single agent in the treatment of PSC. PMID- 15667488 TI - The spectrum of chronic hepatitis C virus infection in the Virginia Correctional System: development of a strategy for the evaluation and treatment of inmates with HCV. AB - BACKGROUND AND OBJECTIVE: Chronic hepatitis C virus (HCV) is common in the inmate population of the United States. Long-standing HCV can progress to cirrhosis, which can contribute to significant morbidity and mortality. However, those inmates with histologically mild disease are unlikely to develop liver-related morbidity or mortality during their period of incarceration. Our objective was to develop an economic strategy for evaluation and treatment of inmates with chronic HCV. METHODS AND MEASURES: A retrospective cohort analysis of 302 inmates within the Virginia Department of Corrections (VDOC) who underwent liver biopsy for chronic HCV at the Virginia Commonwealth University Health System between 1998 and 2002 was performed. The data from this analysis was to utilized to develop a cost model for treatment of chronic HCV in this population based upon biochemical or histologic criteria. We used the perspective of the VDOC using actual costs paid to providers, hospitals, and pharmacies. The primary endpoint was cost effectiveness of HCV treatment. RESULTS: Eighty percent of inmates with chronic HCV were genotype 1, 49% had a normal value for serum ALT at the time of evaluation, 30% had no fibrosis, and 24% had bridging fibrosis or cirrhosis. The cost to evaluate and treat 100 consecutive inmates with peginterferon and ribavirin regardless of serum ALT and liver histology was calculated to be $1,775,900 or $35,500 per sustained virologic response (SVR). Although the cost declined by 50% if only those patients with an elevated serum ALT were treated, 45% of those inmates with varying degrees of fibrosis, and 21% with cirrhosis would not have received therapy utilizing this scenario. In contrast, the cost of performing liver biopsy and treating only those patients with any degree of fibrosis was $1,367,043; a savings of slightly more than $400,000 per 100 patients evaluated. The overall cost of treatment was most influenced by the price of peginterferon and ribavirin, which declined as the histologic criteria utilized for treatment increased. CONCLUSIONS: A strategy in which inmates with chronic HCV are evaluated and a decision regarding treatment is based upon either biochemical or histologic criteria, which appears to balance both the health-care rights of the inmate and the impact of treating this disease on the financial and other resources of the correctional system. PMID- 15667489 TI - Pyogenic liver abscess with a focus on Klebsiella pneumoniae as a primary pathogen: an emerging disease with unique clinical characteristics. AB - OBJECTIVES: Pyogenic liver abscess is a common intraabdominal infection. Historically, Escherichia coli (E. coli) has been the predominant causative agent. Klebsiella liver abscess (KLA) was first reported in Taiwan and has surpassed E. coli as the number one isolate from patients with hepatic abscesses in that country and reports from other countries, including the United States, have increased. We examined the microbiologic trends of pyogenic liver abscess at our institution to determine if a similar shift in etiologic agents was occurring. METHODS: We examined all cases of liver abscess at our institution from 1999 to 2003 via a retrospective chart review of inpatient records and reviewed the English literature via a MEDLINE search for all U.S. cases of KLA. RESULTS: Since 1966, only 12 cases of KLA have been reported in the United States. We report six cases of KLA at our institution alone; 2 patients were not Asian, and 4 were not diabetic. Klebsiella pneumoniae (K. pneumoniae) was the most common cause of pyogenic hepatic abscess at our institution over the last 5 yr period. When comparing Klebsiella versus other causes of pyogenic liver abscess, there were no significant differences in demographics or laboratory findings; however, most of our Klebsiella cases occurred among Filipinos. Review of the 18 cases of K. pneumoniae liver abscess in the United States showed that Klebsiella cases occurred predominantly among middle-aged men; 83% had concurrent bacteremia and 28% had metastatic complications. An increasing number of cases were reported from the United States since the mid-1990s. CONCLUSIONS: These data suggest that KLA may represent an emerging disease in Western countries, such as the United States. The diagnosis of K. pneumoniae should be considered in all cases of liver abscess, and appropriate antibiotic therapy and a diagnostic work up for metastatic complications should be employed. PMID- 15667490 TI - Relationships between hepatic iron content and virologic response in chronic hepatitis C patients treated with interferon and ribavirin. AB - OBJECTIVES: The aims of this study were to determine the effect of pretreatment hepatic iron concentration (HIC) on response to combination therapy with interferon and ribavirin, and to examine the change in HIC associated with this treatment. METHODS: Patients with hepatitis C who underwent liver biopsy before and after combination therapy were studied retrospectively. HIC was measured from paired pre- and posttreatment liver biopsy specimens, and histologic grade and stage were recorded. RESULTS: Sixty of 112 (54%) patients achieved sustained virologic response (SVR); response varied by genotype (genotype 1 (44%), genotype 2 or 3 (85%)). There was no difference in pretreatment median HIC between responders and nonresponders (404 microg/g and 394 microg/g, respectively; p= 0.31); patients with HIC > or = 500 microg/g were not less likely to achieve SVR (OR = 1.1; 95% CI 0.5-2.3). In a multivariate analysis, factors associated with SVR included genotype 2 or 3 (OR = 12.2; 95% CI 3.1-47.8) and viral load < 2 million copies/ml (OR = 3.6; 95% CI 1.3-10.0). HIC > or = 500 microg/g did not decrease the likelihood of SVR (OR = 0.8; 95% CI 0.3-2.1). There was no significant change in HIC after combination therapy (median increase in HIC = 29.5 microg/g), and the change in HIC did not differ between responders and nonresponders (p= 0.73). CONCLUSIONS: Pretreatment HIC is not an independent predictor of response to therapy with interferon and ribavirin. Combination therapy does not significantly change HIC regardless of baseline histology or virologic response. PMID- 15667491 TI - The familial Mediterranean fever (MEVF) gene as a modifier of Crohn's disease. AB - OBJECTIVES: Crohn's disease (CD) has been reported to be more frequent among non Ashkenazi Jewish patients suffering from familial Mediterranean fever (FMF). Interestingly, functional similarities between the CD susceptibility gene (NOD2/CARD15) and the FMF gene (MEFV) have been described: both belong to the death domain containing protein family, important in the regulation of apoptosis, cytokine processing and inflammation. AIMS: To investigate the prevalence of MEFV mutations in Jewish non-Ashkenazi CD patients and its putative effect on CD presentation. METHODS: Germline DNA of 105 Israeli CD patients of non-Ashkenazi and mixed Ashkenazi-non-Ashkenazi ethnic background was analyzed for three most common MEFV mutations: M694V, V726A, and E148Q. Five patients (4.7%) with a clinical diagnosis of FMF were included. Data obtained from each patient included: age of onset, disease location, and behavior, the presence of extraintestinal manifestations of CD and therapeutic regimens. RESULTS: The overall prevalence of mutation carriers among non-FMF-CD patients was 13% (13/100). A stricturing disease pattern was observed in 56% (10/18) of all carriers, FMF-CD, and non-FMF-CD patients, and in 25% (22/87) of noncarriers (OR: 3.7, 95% CI: 1.3-10.5, p= 0.015). The prevalence of fistulas was comparable in both groups. Extraintestinal manifestations were significantly more frequent among carriers than noncarriers (65%vs 32%, OR 3.9, 95% CI = 1.3-11.5, p= 0.015). No differences were observed in disease location and disease severity. CONCLUSIONS: MEFV mutations are not associated with CD susceptibility, yet the presence of these mutations appears to be associated with a stricturing disease pattern and extraintestinal disease manifestations of CD. PMID- 15667492 TI - Technetium-99m-HMPAO labeled leukocyte single photon emission computerized tomography (SPECT) for assessing Crohn's disease extent and intestinal infiltration. AB - OBJECTIVES: Scintigraphy using radiolabeled leukocytes is a useful technique for assessing intestinal infiltration in Crohn's disease (CD). However, limits of planar images include overlapping activity in other organs and low specificity. To investigate the usefulness of (99m)Tc-HMPAO (hexametyl propylene amine oxime) labeled leukocyte single photon emission computerized tomography (SPECT) for assessing CD lesions, in comparison with planar images. METHODS: Twenty-two inflammatory bowel disease patients (19 CD; 2 ulcerative colitis, UC; 1 ileal pouch) assessed by conventional endoscopy or radiology were enrolled. Leukocytes were labeled with (99m)Tc-HMPAO. SPECT images were acquired at 2 h and planar images at 30 min and 2 h. Bowel uptake was quantitated in nine regions (score 0 3). RESULTS: Both SPECT and planar images detected a negative scintigraphy (score 0) in the UC patient with no pouchitis and a positive scintigraphy (score 1-3) in the 21 patients showing active inflammation by conventional techniques. SPECT showed a higher global score than planar images (0.71 +/- 0.09 vs 0.30 +/- 0.05; p < 0.001), and in particular in the right iliac fossa (p= 0.003), right and left flank (p < 0.001; p= 0.02), hypogastrium (p= 0.002), and mesogastrium (p < 0.001). SPECT provided a better visualization and a higher uptake than planar images in patients with ileal and ileocolonic CD (6.45 +/- 0.82 vs 2.8 +/- 0.55, p < 0.001; 5.5 +/- 1.6 vs 2.6 +/- 0.7, p= 0.03), and with perianal CD (6.6 +/- 1.6 vs 3.4 +/- 1.2; p= 0.03). CONCLUSIONS: (99m)Tc-HMPAO labeled leukocyte SPECT provides a more detailed visualization of CD lesions than planar images. This technique may better discriminate between intestinal and bone marrow uptake, thus being useful for assessing CD lesions within the pelvis, including perianal disease. PMID- 15667493 TI - Is endoscopy necessary for the measurement of disease activity in ulcerative colitis? AB - OBJECTIVES: Many disease activity indices are used to measure ulcerative colitis. Invasive indices incorporate an endoscopic score, while noninvasive indices do not require endoscopy. In clinical practice, many patients are treated based on their symptoms without endoscopic evaluation. However, invasive indices are commonly used in clinical research. Our objective was to determine whether endoscopy is necessary for the assessment of disease activity in patients with ulcerative colitis. METHODS: Sixty-six consecutive ulcerative colitis patients were evaluated with invasive indices: the St. Mark's index and the Ulcerative Colitis Disease Activity Index (UCDAI); and noninvasive indices: the Simple Clinical Colitis Activity Index (SCCAI) and the Seo index. The correlations between the indices were measured. The contribution of the endoscopic items was measured with linear regression modeling. The overlap of endoscopy with other items in the standard indices was determined through factor analysis. RESULTS: The two noninvasive indices correlated well with the invasive St. Mark's Index (SCCAI 0.86, Seo 0.70). After adjusting for the three noninvasive UCDAI items, the UCDAI endoscopy item predicted only 0.04% of the variance in the St. Mark's index. Factor analysis demonstrated that this is because the endoscopy items in the invasive indices correlate with stool frequency and stool blood items. CONCLUSIONS: Endoscopy items contribute little additional information to indices of disease activity in ulcerative colitis. The clinical practice of treating patients based on reported symptoms is appropriate. The use of noninvasive indices in clinical trials could lower study costs and may increase subjects' willingness to participate. PMID- 15667494 TI - Tegaserod for the treatment of chronic constipation: a randomized, double-blind, placebo-controlled multinational study. AB - OBJECTIVES: Chronic constipation is a common, persistent disorder with limited effective treatment options. This study investigated the efficacy, safety, and tolerability of tegaserod in the treatment of chronic constipation. METHODS: After a 2-wk baseline period, patients were randomized to double-blind treatment of 12 wk with tegaserod (2 or 6 mg b.i.d.) or placebo. Response during weeks 1-4 (primary variable) was defined as an increase in complete spontaneous bowel movement (CSBM)/wk. Secondary variables included response during weeks 1-12, patient evaluation of individual symptoms, and global assessment of bowel habits and constipation. RESULTS: One thousand two hundred and sixty-four patients were randomized to tegaserod or placebo. Responder rates for the primary efficacy variable were 35.6% for tegaserod 2 mg b.i.d. (p= 0.0059 vs placebo), 40.2% for 6 mg b.i.d. (p < 0.0001 vs placebo) and 26.7% for placebo. The number needed to treat was 7.3 for the 6 mg b.i.d. dose compared with 11.1 for tegaserod 2 mg b.i.d. Tegaserod 6 mg b.i.d. reduced straining, abdominal bloating/distension, and abdominal pain/discomfort during the 12-wk treatment period compared with placebo (p < 0.05 for all symptoms). Significant improvements were also seen in stool form and in global assessment of bowel habits and constipation. The most common adverse events, headache and abdominal pain, were more frequent with placebo than with tegaserod. CONCLUSIONS: Tegaserod was efficacious in relieving symptoms of chronic constipation and was well tolerated. PMID- 15667495 TI - Analysis of the fecal microbiota of irritable bowel syndrome patients and healthy controls with real-time PCR. AB - OBJECTIVE: The gut microbiota may contribute to the onset and maintenance of irritable bowel syndrome (IBS). In this study, the microbiotas of patients suffering from IBS were compared with a control group devoid of gastrointestinal (GI) symptoms. METHODS: Fecal microbiota of patients (n = 27) fulfilling the Rome II criteria for IBS was compared with age- and gender-matched control subjects (n = 22). Fecal samples were obtained at 3 months intervals. Total bacterial DNA was analyzed by 20 quantitative real-time PCR assays covering approximately 300 bacterial species. RESULTS: Extensive individual variation was observed in the GI microbiota among both the IBS- and control groups. Sorting of the IBS patients according to the symptom subtypes (diarrhea, constipation, and alternating predominant type) revealed that lower amounts of Lactobacillus spp. were present in the samples of diarrhea predominant IBS patients whereas constipation predominant IBS patients carried increased amounts of Veillonella spp. Average results from three fecal samples suggested differences in the Clostridium coccoides subgroup and Bifidobacterium catenulatum group between IBS patients (n = 21) and controls (n = 15). Of the intestinal pathogens earlier associated with IBS, no indications of Helicobacter spp. or Clostridium difficile were found whereas one case of Campylobacter jejuni was identified by sequencing. CONCLUSIONS: With these real-time PCR assays, quantitative alterations in the GI microbiota of IBS patients were found. Increasing microbial DNA sequence information will further allow designing of new real-time PCR assays for a more extensive analysis of intestinal microbes in IBS. PMID- 15667496 TI - Visceral sensitivity and symptoms in patients with constipation- or diarrhea predominant irritable bowel syndrome (IBS): effect of a low-fat intraduodenal infusion. AB - BACKGROUND: Visceral hypersensitivity is common in Irritable Bowel Syndrome (IBS) patients, and symptoms exacerbate postprandially. Yet the effects of nutrients on visceral sensitivity and symptoms in these patients have not been fully explored. AIMS: To evaluate the differences of visceral sensitivity and symptoms in healthy subjects and IBS patients during fasting and intraduodenal lipids infusion. METHODS: Graded rectal distensions at fixed tension levels were performed in 16 IBS patients (8 IBS-C and 8 IBS-D) and 6 healthy subjects before and during intraduodenal lipids infusion at 0.5 kcal/min. Tension levels were increased in 4 gr increments up to 64 gr or discomfort during both conditions. At each step, perception and symptoms were measured by means of a validated questionnaire. RESULTS: In basal conditions, perception thresholds in IBS patients and health were, respectively, 8 +/- 2 gr versus 32 +/- 9 gr (p < 0.001) with no changes during lipids. Intraduodenal lipids infusion significantly lowered threshold of discomfort in IBS patients in comparison to fasting (24 +/- 6 gr vs 34 +/- 4 gr; p < 0.05), while health tolerated all distension without discomfort. No differences of compliance, perception, or discomfort were observed between the two subgroups of patients at each tension step. The predominant symptom elicited in patients with IBS-C was abdominal pain (54%), while patients with IBS-D exhibited urgency (63%, p < 0.005); this pattern was maintained during lipids. CONCLUSIONS: Intraduodenal lipids increase visceral sensitivity in both IBS-C and IBS-D; symptoms specificity in response to rectal distension is maintained in the postprandial period. Lipids may be responsible for the postprandial symptoms exacerbation in IBS. PMID- 15667497 TI - Role of supplemental calcium in the recurrence of colorectal adenomas: a metaanalysis of randomized controlled trials. AB - BACKGROUND: Colorectal adenomas are neoplastic growths that are important targets for chemoprevention. Dietary calcium is thought to play an important role in chemoprevention. However, the role of calcium supplementation for preventing recurrence of adenomas is controversial. We performed a systematic review and meta-analysis to study the role of calcium supplementation in preventing recurrence of adenomas. METHODS: We searched electronic bibliographic databases (Cochrane Database of Systematic Reviews, Cochrane Controlled Trials Register, CINAHL, EMBASE, and MEDLINE) and contacted authors to identify potentially eligible studies. RESULTS: We identified three trials including 1,485 subjects with previously removed adenomas who were randomized to calcium versus placebo supplementation. The study endpoint was recurrence of adenomas at the end of 3-4 yr in 1,279 patients who completed the trials. We found that the recurrence of adenomas was significantly lower in subjects randomized to calcium supplementation (RR: 0.80, CI: 0.68, 0.93; p-value = 0.004). CONCLUSIONS: This systematic review and meta-analysis suggest that calcium supplementation prevents recurrent colorectal adenomas. PMID- 15667498 TI - Calcium supplements to prevent colorectal adenomas. AB - Calcium supplements have been shown to decrease the risk of colorectal adenomas. In this issue of the Journal, Shaukat et al. report the results of a systematic review and meta-analysis of randomized controlled trials of calcium supplementation. The authors statistically combined the data from the three trials that met strict eligibility criteria. The overall relative risk was 0.80 (95% CI: 0.68-0.93) and the number needed to treat was 14. The results of this meta-analysis support a preventive role for calcium supplements. PMID- 15667499 TI - Effectiveness of devices purported to reduce flatus odor. AB - OBJECTIVE: A variety of charcoal-containing devices are purported to minimize problems with odoriferous rectal gas; however, the evidence supporting the efficacy of these products is virtually all anecdotal. We objectively evaluated the ability of these devices to adsorb two malodorous, sulfide gases (hydrogen sulfide and methylmercaptan) instilled at the anus. METHODS: Via a tube, 100 ml of nitrogen containing 40 ppm of sulfide gases and 0.5% H(2) was instilled at the anus of six healthy volunteers who wore gas impermeable Mylar pantaloons over their garments. Since H(2) is not adsorbed by charcoal, the fraction of the sulfide gases removed could be determined from the concentration ratio of sulfide gas: H(2) in the pantaloon space relative to the ratio in instilled gas. RESULTS: Measurements with no device in place showed that subjects' garments removed 22.0 +/- 5.3% of the sulfide gases, and results obtained with each device were corrected for this removal. The only product that adsorbed virtually all of the sulfide gases was briefs constructed from an activated carbon fiber fabric. Pads worn inside the underwear removed 55-77% of the sulfide gases. Most cushions were relatively ineffective, adsorbing about 20% of the gases. CONCLUSIONS: The ability of charcoal-containing devices to adsorb odoriferous rectal gases is limited by incomplete exposure of the activated carbon to the gases. Briefs made from carbon fiber are highly effective; pads are less effective, removing 55-77% of the odor; cushions are relatively ineffective. PMID- 15667500 TI - Bowel habits in hemorrhoid patients and normal subjects. AB - OBJECTIVES: Bleeding, pain, soiling, and prolapse are the classic symptoms in hemorrhoid disease, but the patients sometimes report a variety of other symptoms. Little is known about functional bowel symptoms in patients with hemorrhoids and few studies have previously addressed this subject. The aim of this study was to compare patients with hemorrhoids with a control population regarding functional bowel symptoms and anorectal complaints. METHODS: One hundred consecutive patients who participated in a randomized study on hemorrhoidectomy completed a validated questionnaire on bowel and anorectal functional symptoms. Two hundred age- and gender-matched population based control subjects, and 100 gender-matched consecutive patients undergoing an orthopedic procedure served as two control groups, and completed the same questionnaire. RESULTS: Bowel frequency was the same in all three groups, but only 37% of the patients described their bowel movements as normal, compared to 55 and 67% of the controls (p < 0.001). Up to 37% of the patients reported bloating, compared to 18 and 26% in the control groups. Abdominal pain associated with bowel evacuation was experienced by 34% of the patients but in 3 and 5% of the controls (p < 0.001). Excessive straining, feeling of incomplete evacuation, and repeated toilet visits were significantly more usual in the patients. Reduced feeling of well being and disturbed social life caused by bowel symptoms was often reported by patients but rarely in the control groups. CONCLUSIONS: Beside hemorrhoidal symptoms, many patients with Grade 3-4 hemorrhoids have concomitant functional bowel symptoms, possibly associated with the irritable bowel syndrome. This knowledge might be important while selecting therapy for patients with hemorrhoids. PMID- 15667501 TI - A polymorphism in the TNF-alpha promoter gene is associated with pediatric onset and colonic location of Crohn's disease. AB - OBJECTIVES: Studies suggest that pediatric onset of Crohn's disease (CD) may demonstrate more frequent upper intestinal and colonic location and in male gender, in comparison to adults. Variability in age of onset (AOO) and location of disease have not been adequately explained to date. NOD2/CARD15 is highly expressed in the ileum, while TNF-alpha expression is distributed throughout the gastrointestinal tract. We hypothesized that polymorphisms that affect TNF-alpha function may influence variability of disease location and AOO of CD. METHODS: We evaluated two CD cohorts based on AOO (pediatric and adult onset) and 100 ethnically matched healthy controls. Patients were evaluated for AOO, disease location, and genotyped for the presence of polymorphisms in NOD2/CARD15 and in the TNF-alpha promoter region. RESULTS: Early AOO was associated with male gender, upper intestinal involvement, and a polymorphism in the binding site for NF-kappaB (TNF-863A polymorphism). NOD2 mutations and TNF-863A polymorphism had equivalent but opposite effects on disease location, with a strong combined effect (p= 0.004 corrected for multiple testing). NOD2/CARD15 was associated with ileal involvement, while presence of TNF-863A was inversely associated with ileal disease (OR = 0.42, p= 0.008) and positively associated with isolated colitis (OR = 2.16, p= 0.008, OR = 2.12, p= 0.03 corrected) and familial disease (p= 0.004). CONCLUSIONS: Pediatric onset of CD in our population was associated with a frequent polymorphism in the binding site for NF-kappaB in TNF-alpha promoter but not to defined NOD2/CARD15 disease-associated mutations. This polymorphism is associated with colitis and familial disease. NOD2/CARD15 mutations and the TNF 863C/A polymorphism have equivalent but opposite effects on disease location. These findings may help explain differences in CD phenotype. PMID- 15667502 TI - Protein microarray analysis of disease activity in pediatric inflammatory bowel disease demonstrates elevated serum PLGF, IL-7, TGF-beta1, and IL-12p40 levels in Crohn's disease and ulcerative colitis patients in remission versus active disease. AB - OBJECTIVES: Cytokines and growth factors play a major role in the dysregulated immune response in inflammatory bowel disease (IBD). We hypothesized that significant differences exist between the serum cytokine and growth factor profiles of pediatric IBD patients with active disease (AD) and those in remission, and that levels of some of these soluble mediators may be used to define regulators in IBD and determine disease activity. METHODS: Eighty-eight consecutive patients with confirmed Crohn's disease (CD) and ulcerative colitis (UC) seen at the Duke Children's Hospital were prospectively enrolled and a serum sample was obtained. Data were recorded at the time of serum collection to calculate disease activity indices. The relative expression of 78 cytokines, growth factors, and soluble receptors was determined using proprietary antibody based protein microarrays amplified by rolling circle amplification. SPSS 8 (SPSS Inc., Chicago, IL) was used to compare protein profiles for CD and UC patients in clinical remission (CR) versus AD. RESULTS: Sixty-five CD patients and 23 UC patients were enrolled. Forty-one CD patients had available samples and PCDAI results. Twenty-two patients were in remission PCDAI < or = 12.5 (median 5), 19 patients had disease activity >15 (median 30). Univariate analysis revealed that PLGF, IL-7, IL-12p40, and TGF-beta1 cytokine levels were significantly elevated for patients in CR versus AD (p < 0.01). Twelve UC serum samples had Seo/Truelove Witt AI for analysis. Five patients were in remission by TW AI and Seo AI < or =110 and 7 patients had active mild-to-severe disease by TW and Seo AI >110. Only one cytokine, IL12p40, showed significance between CR versus AD (p < 0.02). CONCLUSIONS: Surprisingly, we found no differences in circulating levels of proinflammatory cytokines but found that pediatric IBD patients in remission compared to those with AD had higher levels of specific circulating cytokines, including the regulatory cytokines IL-12p40 and TGF-beta1. It may be that these cytokines directly regulate intestinal inflammation in IBD or reflect the activity of T regulatory cells in negatively regulating the inflammatory response. Further studies will be needed to validate our results to define the molecular pathways involved in the intestinal immune response in man. PMID- 15667503 TI - Apoptosis resistance in Barrett's esophagus: ex vivo bioassay of live stressed tissues. AB - BACKGROUND AND AIMS: Barrett's esophagus (BE) is a premalignant lesion of the distal esophagus in which squamous epithelial cells are replaced by metaplastic intestinal-like columnar epithelium that contains goblet cells. The factors that contribute to the progression from normal squamous mucosa to BE, Barrett's dysplasia, and adenocarcinoma are not well understood at the molecular level. Since reflux of bile acids is associated with BE development, we speculate that cells with an apoptosis-resistant phenotype are selected after long-term repeated exposure to pulses of bile acids. This will result in the survival of cells with unrepaired DNA damage, and a consequent increase in genomic instability leading to cancer progression. The major goal of this study is to compare sensitivity to apoptosis induced by the bile acid, deoxycholate (DOC), a known inducer of apoptosis, in normal esophageal squamous epithelium, normal colon epithelium, and BE. METHODS: Thirteen patients with a confirmed diagnosis of BE and four patients who had undergone clinically indicated colectomy were included in the present study. Freshly obtained biopsies were incubated with control medium or medium supplemented with 1 mM DOC for 3 h and then evaluated for apoptotic changes using transmission electron microscopy and immunohistochemical staining for two apoptotic markers, cleaved caspase 3 and cleaved cytokeratin 18. RESULTS: Our results indicate that BE is resistant to apoptosis induced by DOC compared to esophageal squamous epithelium and normal colon epithelium. In addition, electron micrographs revealed mitochondrial swelling in squamous epithelial cells treated ex vivo with DOC, which was absent in epithelial cells of BE. Formation of swollen mitochondria is an early marker of apoptotic cell death. Altogether, the data indicate that reduced apoptosis capability in BE tissue may contribute to progression to esophageal adenocarcinoma. PMID- 15667505 TI - What route to feed patients with severe acute pancreatitis: vein, jejunum, or stomach? PMID- 15667504 TI - A randomized study of early nasogastric versus nasojejunal feeding in severe acute pancreatitis. AB - BACKGROUND: After 50 yr in which nasoenteric feeding was considered contraindicated in acute pancreatitis (AP), several clinical studies have shown that early nasojejunal (NJ) feeding can be achieved in most patients. A pilot study of early nasogastric (NG) feeding in patients with objectively graded severe AP proved that this approach was also feasible. A randomized study comparing NG versus NJ feeding has been performed. METHODS: A total of 50 consecutive patients with objectively graded severe AP were randomized to receive either NG or NJ feeding via a fine bore feeding tube. The end points were markers of the acute phase response APACHE II scores and C-reactive protein (CRP) measurements, and pain patterns by visual analogue score (VAS) and analgesic requirements. Complications were monitored and comparisons made of both total hospital and intensive-care stays. RESULTS: A total of 27 patients were randomized to NG feeding and 23 to NJ. One of those in the NJ group had a false diagnosis, thereby reducing the number to 22. Demographics were similar between the groups and no significant differences were found between the groups in APACHE II score, CRP measurement, VAS, or analgesic requirement. Clinical differences between the two groups were not significant. Overall mortality was 24.5% with five deaths in the NG group and seven in the NJ group. CONCLUSIONS: The simpler, cheaper, and more easily used NG feeding is as good as NJ feeding in patients with objectively graded severe AP. This appears to be a useful and practical therapeutic approach to enteral feeding in the early management of patients with severe AP. PMID- 15667506 TI - Symptom-based outcome measures for dyspepsia and GERD trials: a systematic review. AB - BACKGROUND: Symptom assessment using questionnaires has been recommended as the primary outcome measure in clinical gastroesophageal reflux and dyspepsia trials. Questionnaires should have proven reliability, validity, and responsiveness, and may assess the frequency and/or severity of dyspepsia symptoms. Although a number of measures have been developed, it remains unclear which of these should be used in new trials. OBJECTIVE: To describe existing questionnaire outcome measures that assess symptoms of gastroesophageal reflux dyspepsia for use in clinical trials. METHODS: Studies were identified from Medline, Embase, the Cochrane library, and reference lists. The inclusion criterion was that the study assessed a questionnaire, which measured the frequency or severity of dyspepsia or gastroesophageal reflux symptoms, in a sample of patients. RESULTS: No direct comparison between questionnaires was possible due to methodological heterogeneity. Thirty-seven studies describing 26 questionnaires met the inclusion criteria. Twelve were unidimensional (assessed symptoms only) and 14 were multidimensional (also assessed quality of life). Eleven questionnaires assessed both frequency and severity of dyspepsia, and 10 had proven reliability, validity, and responsiveness. No studies compared different questionnaires. CONCLUSIONS: Future gastroesophageal reflux and dyspepsia clinical trials should use unidimensional or multidimensional outcome measures that assess both the frequency and severity of symptoms, and have proven reliability, validity, and responsiveness. Further research is necessary to compare existing outcome measures to determine which are the most reliable, valid, and responsive instruments. PMID- 15667507 TI - Iron-deficiency anemia and Helicobacter pylori infection: a review of the evidence. PMID- 15667508 TI - Hepatorenal syndrome: a dreaded complication of end-stage liver disease. AB - Hepatorenal syndrome is the dreaded complication of end-stage liver disease characterized by functional renal failure due to renal vasoconstriction in the absence of underlying kidney pathology. The pathogenesis of hepatorenal syndrome is the result of an extreme underfilling of the arterial circulation secondary to an arterial vasodilation located in the splanchnic circulation. This underfilling triggers a compensatory response with activation of vasoconstrictor systems leading to intense renal vasoconstriction. The diagnosis is based on established diagnostic criteria aimed at excluding nonfunctional causes of renal failure. The prognosis of patients with hepatorenal syndrome is extremely poor especially in those who have a rapidly progressive course. Liver transplantation is the best option in suitable candidates, but it is not always applicable due to the short survival expectancy and donor shortage. Pharmacological therapies based on the use of vasoconstrictor drugs (terlipressin, midodrine, octreotide, or noradrenline) are the most promising in the aim of successfully offering a bridge to liver transplantation. Other treatments such as transjugular intrahepatic portosystemic shunts and albumin dialysis are effective but experience is very limited. Although there is limited information on the prevention of hepatorenal syndrome, intravenous albumin infusion in patients with spontaneous bacterial peritonitis and with oral pentoxifylline in patients with acute alcoholic hepatitis seems to effectively prevent hepatorenal syndrome in these two settings. PMID- 15667509 TI - Emerging indications for albumin dialysis. AB - The accumulation of albumin-bound toxins in liver failure is believed to be responsible for the development of associated end-organ dysfunctions (kidney, circulation, brain). Albumin dialysis utilizes the scavenging functions of albumin for the removal of toxins. The Molecular Adsorbents Recirculating System (MARS) is one such extracorporeal liver support device where blood is dialyzed across an albumin-impregnated membrane against 20% albumin. Charcoal and anion exchange resin columns in the circuit cleanse and regenerate the albumin dialysate. Clinical studies in the last decade have demonstrated proven reduction in hyperbilirubinemia, along with an improvement in encephalopathy in liver failure patients, as well as apparent improvement in survival. Some studies have also reported improvement of systemic hemodynamics and renal function in these patients. Amelioration of intractable pruritus and treatment of toxicities with albumin-bound substances are some of the newer indications emerging. However, the specific underlying pathophysiological mechanisms are still not clear. Two other systems based on the removal of albumin-bound toxins, the Prometheus (using the principle of fractionated plasma separation and adsorption [FPSA]), and the single pass albumin dialysis (SPAD) are also currently under development but available clinical data are limited. PMID- 15667510 TI - The hamartomatous polyposis syndromes: a clinical and molecular review. AB - Inherited forms of gastrointestinal cancer have been a major focus of study and advancement over the past decade. Familial adenomatous polyposis and hereditary nonpolyposis colon cancer are the two most common heritable colon cancer syndromes. Inherited polyposis syndromes are characterized by the dominant type of polyp (whether adenomatous or hamartomatous) present and by the polyp's location within the gastrointestinal tract. The hamartomatous polyposis syndromes are characterized by an overgrowth of cells native to the area in which they normally occur. They represent a small but appreciable number of the gastrointestinal inherited cancer predisposition syndromes; it is now known that many of these syndromes carry a substantial risk for developing colon cancer as well as other gastrointestinal and pancreatic cancers. Patients afflicted with these syndromes are also at significant risk for extraintestinal malignancies. Seven inherited hamartomatous polyposis syndromes have been described: familial juvenile polyposis syndrome, Cowden's syndrome, Bannayan-Ruvalcaba-Riley syndrome, Peutz-Jeghers syndrome, basal cell nevus syndrome, neurofibromatosis 1, and multiple endocrine neoplasia syndrome 2B. Hereditary mixed polyposis syndrome is a variant of juvenile polyposis characterized by both hamartomatous and adenomatous polyps. The hamartomatous syndromes occur at approximately 1/10th the frequency of the adenomatous syndromes and account for <1% of colorectal cancer in Northern America. While the diagnosis of these inherited syndromes is primarily clinical, genetic testing is now available for all six syndromes. However, there are a significant number of spontaneous mutations seen in each of the syndromes. The management of these patients necessitates a coordinated multidisciplinary approach. The purpose of this review is to characterize the clinical and pathological features of these syndromes and to review the targets of cancer surveillance. The molecular alterations responsible for the inherited hamartomatous polyposis syndromes will also be discussed. PMID- 15667511 TI - Painful cystic duct remnant diagnosed by endoscopic ultrasound. AB - Postcholecystectomy pain may remain unexplained and difficult to treat. This report describes three patients with constant postcholecystectomy abdominal pain that may have arisen from the cystic duct remnant or a neuroma of the cystic duct stump. In each case pain was exacerbated by pushing on cystic duct surgical clips with an EUS-guided needle, and temporarily abolished by an EUS-guided injection of bupivicaine and triamcinolone. Two patients underwent surgical resection of the cystic duct remnant and the third did not require further treatment. Two of the three patients had long-term improvement. EUS is a novel modality for assessing the cystic duct remnant and performing a therapeutic trial. PMID- 15667512 TI - EUS assessment of response to chemoradiation in esophageal cancer patients. PMID- 15667513 TI - Infliximab for the treatment of chronic anemia in Crohn's disease. PMID- 15667515 TI - Chemokines as attractive targets in liver carcinogenesis. PMID- 15667516 TI - Reflux esophagitis and ectopic columnar epithelium (Barrett's esophagus) in the esophageal stump after cervical gastroplasty. PMID- 15667517 TI - 5-ASA therapy and renal function in inflammatory bowel disease. PMID- 15667520 TI - Analysis of the contributions of L-selectin and CXCR3 in mediating leukocyte homing to pregnant mouse uterus. AB - PROBLEM: Dynamic changes occur in endometrial immune cell populations during pregnancy but regulatory events promoting cell recruitment to the uterus are not established. Ovarian steroid hormones promote l-selectin and alpha4-integrin mediated interactions between human peripheral natural killer (NK) cells and uterine endothelium while CXCR3, CXCR4 and their ligands are implicated in homing of human uNK cells to decidua. METHOD OF STUDY: Mice genetically-ablated for l selectin or CXCR3 were studied. Morphometric analyses of implantation sites and assays of cell function (in vitro adhesion; in vivo homing following adoptive cell transfer) were undertaken. RESULTS: Leukocytes home to anti-mesometrial decidua with l-selectin making an early (<6 hr in vivo) contribution. Unexpectedly, CXCR3 deletion had no effect on homing but reduced the ability of uNK cells to modify placental spiral arterioles. CONCLUSIONS: Redundant mechanisms underlie homing of leukocytes to the uterus and their importance can be evaluated by an in vivo approach described herein. PMID- 15667521 TI - Loss of surface CD200 on stored allogeneic leukocytes may impair anti-abortive effect in vivo. AB - PROBLEM: Prevention of spontaneous abortion by allogeneic mononuclear leukocyte immunotherapy has proven ineffective in the CBA x DBA/2 murine abortion model if the leukocytes are stored overnight before inoculation. The mechanism and generality of the phenomenon has not been elucidated. METHODS: As prevention of recurrent abortion in the CBA x DBA/2 model requires allogeneic BALB/c lymphoid cells bearing paternal antigens and the tolerance-signaling molecule CD200 (OX 2), we evaluated effects of cell storage on cell surface CD200 expression using flow cytometry of fresh or stored cells stained with monoclonal anti-CD200 antibody. Release of putative CD200 molecules into culture supernatant during storage was tested by the ability of supernatants to block binding of anti-CD200 to freshly isolated cells. Similar studies were done using human peripheral blood mononuclear leukocytes. Possible binding of soluble CD200 to immunoglobulin G (IgG) molecules in plasma as a basis for the anti-abortive effect of intravenous immunoglobulin G (IVIG) was tested using the standard peripheral blood lymphocyte (PBL) natural killer (NK) cell lysis of (51)Cr-labeled K562 cells and monoclonal anti-human CD200 antibodies. RESULTS: Loss of anti-abortive effect of BALB/c cells with overnight storage at 4 degrees C and blocking of protective effect of freshly isolated cells with anti-CD200 antibody was confirmed. Supernatants of stored cells acquired a low level of protective activity against abortion in the CBA x DBA/2 model. Cell surface CD200 was lost with overnight storage at 4 or 22 degrees C, and supernatants acquired the ability to block binding of anti-CD200 antibody to fresh cells. Similar results were obtained using human PBL. However, if cells were stored overnight in IgG containing plasma, binding was not blocked. Suppression of NK cell lysis by PBL was abrogated if anti-CD200 antibody was added to the assay. CONCLUSIONS: Loss of the tolerance signal CD200 from allogeneic cells occurs with storage overnight, and their ability to protect against abortion is lost. CD200 appears to be shed into the supernatant, and may associate with IgG molecules rendering IVIG suppressive. PMID- 15667522 TI - Lymphocyte sub-populations in gestational diabetes. AB - PROBLEM: We hypothesize that the normal immunologic responses by the maternal immune system during pregnancy are not as well-regulated in gestational diabetes (GD) patients as in healthy pregnant women. METHOD OF STUDY: Using two-color flow cytometry we evaluated frequencies of peripheral blood lymphocytes in 20 GD patients being treated with insulin; 43 GD patients treated with dietary therapy but no insulin; 44 women experiencing normal pregnancies; and 48 non-pregnant women. RESULTS: When compared with healthy pregnant women, both GD cohorts showed higher percentages CD4(+)CD25(+) (P < 0.05), CD4(+)CD45RO(+) (P < 0.05) and CD4(+)CD29(+) (P < 0.01) but lower percentages of CD4(+)CD45RA(+) (P < 0.05). Higher percentages of the activated phenotypes CD8(+)CD25(+) and CD8(+)HLA-DR(+) cells in the diet-treated cohort and CD4(+)HLA-DR(+) cells in insulin-treated GB cohort, were observed compared with healthy pregnant subjects (P < 0.05). CONCLUSIONS: Expanded populations of activated peripheral blood T cells are associated with GD, suggesting that normal maternal immunosuppression is less effective in GD-afflicted women. PMID- 15667523 TI - Gonadotropin-releasing hormone agonist administration affects the thymopoiesis in adult female rats independently on gonadal hormone production. AB - PROBLEM: In addition to having an indirect effect on the T-cell development by controlling the production of ovarian steroids, an accumulating body of evidence suggest that GnRH analogue (GnRH-A) administration may exert a thymopoietic regulatory effect that is not mediated by ovarian hormones. METHOD OF STUDY: In non-ovariectomized (non-OVX) and OVX adult female AO rats treated s.c. with GnRH A or saline (controls), over 14 days, were estimated the thymic cellularity and thymocyte expression of CD4/CD8/TCRalphabeta by stereological analysis and three color flow cytometry, respectively. RESULTS: GnRH-A in both groups of rats diminished the thymic cellularity. In non-OVX rats GnRH-A increased the relative numbers of immature cells (CD4-8-TCRalphabeta(-), CD4-8-TCRalphabeta(low) and CD4+8-TCRalphabeta(low)), and reduced those of positively selected CD4+8+TCRalphabeta(high) and mature (CD4-8+TCRalphabeta(high), CD4(+8) TCRalphabeta(high)) cells, suggesting decelerated expression of TCRalphabeta followed by less efficient positive selection and further maturation of the selected cells. Differently, in OVX rats GnRH-A decreased the percentage of immature (CD4-8-TCRalphabeta(-), CD4+8+ TCRalphabeta(-)) cells and increased those of all TCRalphabeta(high) subsets, suggesting an increased rate of early thymocyte differentiation, more efficient positive selection and further maturation of the selected cells. CONCLUSIONS: The effect of GnRH-A administration is affected by the presence of ovarian steroids. PMID- 15667524 TI - Monoclonal antibodies against sperm intra-acrosomal antigens as markers for male infertility diagnostics and estimation of spermatogenesis. AB - PROBLEM: To determine the ability of monoclonal antibodies (MoAbs) against acrosomal antigens to detect physiology and pathology of human spermatozoa and to detect spermatids in ejaculates of infertile male with azoospermia. METHOD OF STUDY: Sperm antigens detected with prepared MoAbs were partially characterized by biochemical and immunocytochemical methods. The acrosomal status of spermatozoa was compared in men with normal and pathological spermiograms and in sperm before and after induced acrosome reaction (AR). Ejaculates from patients were tested for the presence of spermatids. RESULTS: MoAbs specifically bind to intra-acrosomal sperm antigens with quantitative difference between ejaculates with normal and pathological spermiograms. These antigens are released from the acrosome after induced AR. MoAbs labeled acrosomal proteins in round and elongated spermatids in the ejaculates of patients with azoospermia. CONCLUSION: MoAbs against intra-acrosomal sperm antigens are useful for human sperm diagnosis and prediction of spermatogenesis. The spermatids can be utilized in assisted reproduction. PMID- 15667525 TI - Pregnancy outcome in recurrent aborters is not influenced by Chlamydia IgA and/or G. AB - PROBLEM: It is unclear whether chlamydia infection influences the miscarriage rate and immunological factors in patients with recurrent miscarriage. METHOD OF STUDY: Chlamydia DNA, IgA and IgG to Chlamydia trachomatis, natural killer cell activity, complement 3 (C3), C4, hemolytic complement, antinuclear antibodies, antiphospholipid antibodies, prolactin, activated partial thromboplastin time, prothrombin time and fibrinogen were examined in 504 patients with a history of two or more consecutive first-trimester miscarriages. Subsequent pregnancy outcomes were compared between cases with and without antibodies to C. trachomatis. RESULTS: Totals of 10 of 30 and 48 of 201 patients receiving no medication miscarried subsequently with and without chlamydia infection. Chlamydia IgA and/or IgG were associated with a high level of C3 but not other immunological and coagulatory parameters. CONCLUSION: Antibodies to C. trachomatis do not influence subsequent pregnancy outcome in patients with a history of recurrent miscarriage. PMID- 15667526 TI - Changes in immune cell distribution and IL-10 production are regulated through endometrial IP-10 expression in the goat uterus. AB - PROBLEM: Changes in distribution or redistribution of immune cells are required for the establishment and maintenance of pregnancy, but these changes during early pregnancy have been poorly understood in the ruminant ungulates. Expression of a chemokine, interferon-gamma (IFN-gamma)-inducible protein 10 kDa (IP-10, CXCL10), was identified in the endometrium of pregnant goats. Population and/or distribution of endometrial immune cells and their cytokine productions could be regulated by IP-10 during the period of pregnancy establishment. METHOD OF STUDY: Using reverse transcriptase-polymerase chain reaction (RT-PCR), expression of IP 10, IFN-gamma, tumor necrosis factor-alpha, interleukin-10 (IL-10), CXCR3 mRNA and leukocyte cell surface markers, CD4, CD8, CD11b and CD45 mRNA during the caprine early pregnancy was investigated. The ability of IP-10 to stimulate peripheral blood mononuclear cells (PBMCs) migration was demonstrated using a chemotaxis assay. Changes in migration of PBMCs' immune cell population and cytokine expressions with IP-10 stimulation were investigated using flow cytometry and RT-PCR respectively. RESULTS: Levels of IP-10, IL-10, CD4 and CD11b mRNA, and the number of CD4 and CD11b positive cells in pregnant goat endometrium were higher than those of cyclic goat endometrium. Migration of PBMCs was stimulated by recombinant caprine IP-10, and the effect was significantly reduced by neutralization with the use of an anti-IP-10 antibody. In the flow cytometric and RT-PCR analyses, migrated cells stimulated by IP-10 increased the expression of IL-10 and CD11b mRNA. Furthermore, IP-10 could stimulate the expression of IL 10 mRNA from PBMCs. CONCLUSION: Endometrial chemokine IP-10 could regulate IL-10 production by resident and possibly migrated cells expressing CD11b, probably natural killer cells, and these changes may result in immune environments of the uterus suitable for conceptus implantation in ruminants. PMID- 15667527 TI - Essential thrombocythaemia causing massive intra-abdominal thrombosis. PMID- 15667528 TI - Atypical haemolytic-uraemic syndrome at the onset of acute promyelocytic leukaemia. PMID- 15667529 TI - Thrombosis and haemorrhage in polycythaemia vera and essential thrombocythaemia. AB - Despite decades of clinical and laboratory research, relatively little has been accomplished concerning the pathogenesis as well as the identification of risk factors for thrombosis and bleeding in myeloproliferative disorders. In polycythaemia vera, the pro-thrombotic effect of an elevated haematocrit is well established. In contrast, thrombocytosis per se has not been similarly incriminated in essential thrombocythaemia. In both conditions, advanced age and the presence of a prior event identify thrombosis-prone patients. There is increasing evidence to suggest an additional role by leucocytes that might partly explain the antithrombotic effects of myelosuppressive therapy. A substantial minority of affected patients display reduced levels of high molecular weight von Willebrand protein in the plasma during extreme thrombocytosis and it is believed that this might explain the bleeding diathesis of such patients. Recent controlled studies support the therapeutic value of hydroxyurea and aspirin in essential thrombocythaemia and polycythaemia vera, respectively. The current communication will address the incidence, phenotype, pathogenesis, risk factors, prevention, and treatment of both thrombosis and haemorrhage in these disorders. PMID- 15667530 TI - Management of thrombosis in cancer: primary prevention and secondary prophylaxis. AB - Although traditional anticoagulant regimens are highly effective and safe in most patients with venous thromboembolism (VTE), the aggressive natural history of VTE and the high risk of serious bleeding in cancer patients can complicate the management of VTE. In addition, because few clinical trials have focused on the prevention and treatment of VTE in this unique patient population, many clinical questions regarding the care of cancer patients with VTE remain unanswered. Currently, low-molecular-weight heparins and oral vitamin K antagonists are the most commonly used agents for the primary and secondary prophylaxis of VTE in patients with or without cancer, but their use is associated with pharmacological and practical limitations. This review will provide an up-to-date summary of the clinical trials that have addressed the management of VTE in patients with cancer. A brief discussion of the potential application of novel anticoagulants in these clinical settings is also included. PMID- 15667531 TI - The crucial role of vascular permeability factor/vascular endothelial growth factor in angiogenesis: a historical review. AB - Angiogenesis is a biological process by which new capillaries are formed and it occurs in many physiological and pathological conditions. It is controlled by the net balance between molecules that have positive and negative regulatory activity and this concept had led to the notion of the 'angiogenic switch', depending on an increased production of one or more of the positive regulators of angiogenesis. Numerous inducers of angiogenesis have been identified and this review offers a historical account of the relevant literature concerning the discovery of one of the best characterized angiogenic factors, namely vascular endothelial growth factor (VEGF)/vascular permeability factor. Moreover, different strategies, designed to stimulate and to inhibit VEGF production in the context of several potential therapeutical implications, are discussed. PMID- 15667532 TI - Gemtuzumab ozogamicin (Mylotarg) has therapeutic activity against CD33 acute lymphoblastic leukaemias in vitro and in vivo. AB - Gemtuzumab ozogamicin (GO) is a humanized anti-CD33 antibody conjugated with the cytotoxic drug calicheamicin and approved for the treatment of relapsed acute myeloid leukaemia. As approximately 18% of acute lymphoblastic leukaemias (ALL) are also CD33 positive, we have investigated the cytotoxic activity of GO on CD33+ ALL cells in vitro and in vivo. 10 ng/ml GO induced 30-95% inhibition of thymidine uptake and 30-70% cell death in four freshly isolated and one in vivo passaged CD33+ ALL-cell cultures. Furthermore, an in vivo model of a CD33+ ALL carrying the Philadelphia chromosome [t(9;22)] was established. 5 x 10(6) ALL-2 cells inoculated in the tail vein of severe combined immunodeficient mice engrafted into haematopoietic organs, reaching a mean of 70%, 61% and 69% human CD45+ cells in bone marrow, spleen and liver, respectively, at 35 d. To test the therapeutic activity of GO, 50 or 100 microg immunotoxin was inoculated i.p. on days 7, 11 and 15 following tumour-cell inoculation. GO treatment dramatically inhibited expansion of ALL-2 cells in all tested organs and increased survival of tumour-injected animals by 28-41 d, relative to controls. These data demonstrated that GO is active both in vitro and in vivo against CD33+ ALL cells. PMID- 15667533 TI - Development of a human acute myeloid leukaemia screening panel and consequent identification of novel gene mutation in FLT3 and CCND3. AB - A study was undertaken to develop an acute myeloid leukaemia (AML) screening panel to uncover novel recurring gene mutations. Analysis was performed on six genes known to be mutated in AML (RUNX1, FLT3, KIT, CEBPA, PTPN11 and NRAS) and an additional two candidate genes (CCND3 and FES) in a panel of 175 primary human AML samples that included all French-American-British types except M3, and all cytogenetic risk groups. One hundred and fifteen mutations were identified in 97 (55%) patients comprising 81 patients (46%) with one mutation, 14 patients (8%) with two mutations and two patients (1%) with three mutations. Fifty-five of 88 (63%) patients with normal karyotype AML had at least one mutation. Correlation was observed between KIT mutation and 'favourable risk' cytogenetics (P <0.001), CEBPA mutation and 'intermediate risk' cytogenetics (P=0.045), and PTPN11 mutation and 'poor risk' disease (P <0.001). The frequency of individual gene mutation was in accordance with previously published studies. Three novel mutations of FLT3 were detected (Y589D, D839G, Y842H) that would have been overlooked by conventional gel electrophoresis. A 51-bp deletion was detected in CCND3 in a patient with normal karyotype AML. This validated panel now provides an important tool to evaluate other candidate genes in the genesis of myeloid malignancy. PMID- 15667534 TI - MDR1 and MRP1 gene expression are independent predictors for treatment outcome in adult acute myeloid leukaemia. AB - Multi drug resistance (MDR) is a major obstacle for cancer therapy. The three major candidates accounting for the development of MDR in acute myeloid leukaemia (AML) are multi drug resistance gene (MDR1), multi drug resistance-related protein gene (MRP1) and lung resistance protein gene (LRP). So far, the differential impact of resistance gene expression on treatment outcome in AML is not clear. Therefore, we examined MDR1, MRP1 and LRP gene expression at diagnosis in 331 adult AML patients in the context of other known prognostic factors, such as age, disease status, cytogenetics and FMS-like tyrosine kinase 3 (FLT3) internal tandem duplication mutational status. Median observation time was longer than 5 years [64.1 months (40.0-87.6)]. MDR1 expression proved to be an independent prognostic factor for outcome of induction therapy (P <0.001) and overall survival (P=0.02), whereas MRP1 expression was an independent predictor for disease-free survival (P=0.01) in the multivariate analysis. This prognostic impact of both resistance genes was also found in patients with intermediate risk cytogenetics. LRP expression, however, had no impact on treatment outcome in AML. Our study shows that resistance gene expression should be considered together with age, cytogenetics and FLT3 mutational status for risk-adapted treatment strategies in AML in the future. PMID- 15667535 TI - The NFATc1 transcription factor is widely expressed in white cells and translocates from the cytoplasm to the nucleus in a subset of human lymphomas. AB - Stimulation of lymphoid cells via their surface receptors triggers signalling pathways that terminate in the nucleus, where they induce alterations in gene transcription. Nuclear factor of activated T cells (NFAT) transcription factors, involved in a major Ca2+-dependent signalling pathway, normally reside in the cytoplasm but re-locate to the nucleus when activation of the pathway (e.g. following ligation of antigen receptors) leads to their dephosphorylation. This study found that one member of the NFAT family (NFATc1/NFAT2) can be detected in routine biopsy samples, where it is seen in essentially all lymphoid cells, but is absent from the great majority of non-haematopoietic cells. An immunohistological evaluation of NFATc1 in almost 300 lymphomas showed that most neoplastic lymphoid cells also express NFATc1 as a cytoplasmic constituent, although it is absent in classical Hodgkin's disease and plasma cell proliferations. Of particular interest was the finding that NFATc1 was relocated to the nucleus in a minority of lymphoid neoplasms (usually diffuse large B-cell lymphomas or Burkitt lymphoma), presumably reflecting activation of the NFAT pathway. It would be of interest to correlate this feature with patterns of gene expression and also with prognosis, since it may identify a subset of human lymphoma that is distinct in its molecular and clinical features. PMID- 15667536 TI - Preclinical studies on NSC290205 aza-steroid alkylator activity in combination with adriamycin against lymphoid leukaemia. AB - Summary NSC290205 (A) is an hybrid synthetic antineoplastic ester that is a combination of a d-lactam derivative of androsterone and an alkylating derivative of N,N-bis(2-chloroethyl)aniline. We tested NSC290205 for synergistic antileukaemic activity with adriamycin (ADR), (i) in vitro against the human lymphoid leukaemia cell lines: CCRF-CEM, MOLT-4, and RPMI-8226, (ii) in vivo against P388 lymphocytic and L1210 lymphoid murine leukaemias (at incipient and advanced phase). Our results indicated significant cytostatic and cytotoxic synergy of NSC290205 and ADR in vitro. We further examined these results in vivo by replacing cyclophosphamide in the standard CHOP (cyclophosphamide, hydroxydaunomycin, Oncovin, prednisone) regimen with NSC290205 (AHOP) and comparing the efficiency of these two regimens in vivo. Although treatment of P388 and L1210 with cyclophosphamide or NSC290205 alone yielded equivalent results, AHOP produced a clear benefit for survival compared with CHOP against advanced leukaemias, confirming the in vitro observations [higher percentage increase in median lifespan of treated animals over the untreated (control): 188% and 239% in L1210, 308% and 353% in P388, P < 0.01, for CHOP and AHOP respectively]. AHOP also proved to be more genotoxic and cytostatic than CHOP, inducing higher sister chromatid exchange levels and cell division delays on P388 cells in vivo. NSC290205 showed superior antineoplastic potential against lymphoid leukaemia and significant synergy with ADR, producing an excellent therapeutic outcome. PMID- 15667537 TI - Anti-tumour response despite loss of donor chimaerism in patients treated with non-myeloablative conditioning and allogeneic stem cell transplantation. AB - Based on a murine model, we conducted a series of trials of m-myeloablative human leucocyte antigen (HLA)-matched or mismatched related donor stem cell transplantation (SCT) with the intention of inducing mixed chimaerism (MC), then administering prophylactic donor lymphocyte infusions (DLIs), for the treatment of advanced haematologic malignancies. Preparative therapy consisted of cyclophosphamide, equine anti-thymocyte globulin (ATG) or MEDI-507 (an anti-CD2 monoclonal antibody) for in-vivo T-cell depletion, thymic irradiation on day -1 and cyclosporine alone for graft-versus-host disease (GVHD) prophylaxis. DLIs were given as early as 5 weeks post-SCT in patients with MC without evidence of GVHD. Twenty-two patients ultimately lost their graft (<1% donor cells) that could no be rescued by DLIs. Nine of 22 (41%) patients who lost donor chimaerism achieved an objective response, including three patients who showed evidence of disease regression following DLI, despite continued absence of macrochimaerism. Six patients were alive at 2.5-5.5 years following SCT, including four in continuous complete remission. In summary, it is possible to achieve sustained remission in patients with chemorefractory malignancies following non myeloablative allogeneic SCT, even in the absence of sustained donor macrochimaerism; DLI may contribute to an ongoing anti-tumour effect in these patients. Immunological mechanisms that correlated with rejection of the graft may have a role in anti-tumour responses via a cell or cytokine-mediated pathway. PMID- 15667538 TI - First description of somatic mosaicism in MYH9 disorders. AB - MYH9 disorders are characterized by giant platelets, thrombocytopenia, and Dohle body-like cytoplasmic granulocyte inclusion bodies that result from mutations in MYH9, which encodes non-muscle myosin heavy chain-A (NMMHCA). These disorders are known to be transmitted in an autosomal dominant manner, although about 20% of cases are considered to be sporadic. We report here the first case of a MYH9 disorder because of somatic mosaicism. The patient was the father of a male with typical May-Hegglin anomaly. The father had normal platelet counts, however, both normal-sized and giant platelets were observed on his peripheral blood smears. In addition, 14% of neutrophils contained inclusion bodies and the rest showed a normal morphology. Quantitative fluorescent polymerase chain reaction analysis showed that only 6% of DNA from peripheral blood leucocytes harboured the mutation. The mosaicism was demonstrated at a similar rate in different tissues, buccal mucosa cells and hair bulb cells, implying that the mutation had occurred before gastrulation. Mosaicism might account for some de novo mutations in MYH9 disorders. PMID- 15667539 TI - Antiphospholipid antibodies in immune thrombocytopenic purpura tend to emerge in exacerbation and decline in remission. AB - Although the presence of antiphospholipid antibodies (APLA) in immune thrombocytopenic purpura (ITP) has been reported, their clinical significance is not clear. The present study investigated APLA profiles in relation to the clinical stages of ITP. We studied APLA in 40 patients in three stages of ITP: exacerbation/relapse (n=7), stable (n=14) and remission (n=19). Both IgG and IgM APLA to six target antigens were measured by enzyme-linked immunosorbent assay: beta2-glycoprotein 1 (beta2GP1), cardiolipin, phosphatidylcholine, phosphatidylserine, phosphatidylethanolamine and factor VII/VIIa. The central finding was that APLA were common in ITP but differed significantly in disease stages, being highest in exacerbation (86% positive), intermediate in stable disease (57%) and lowest in remission (42%). In exacerbations, APLA were predominantly of IgG class, while in stable disease, IgM predominated. During remission, APLA often became undetectable. Both the frequency and titres of APLA were significantly higher during exacerbation than remission. An inverse correlation was found between platelet count and nearly all APLA (except beta2GP1). Sequential study of six patients revealed that APLA tended to emerge and rise with exacerbation, concurrently with new episodes of bleeding and became undetectable during remission. These findings raise the possibility that APLA may play a role in the exacerbation and remission of ITP or they may be a consequence of platelet destruction. PMID- 15667540 TI - Thrombotic thrombocytopenic purpura in patients with retroviral infection is highly responsive to plasma infusion therapy. AB - We prospectively studied presentation biological differences and the response to therapy in patients with thrombotic thrombocytopenic purpura (TTP) associated with, or unrelated to human immunodeficiency virus (HIV) infection. TTP patients underwent standard evaluations and were treated with prednisone 1 mg/kg in addition to infusions of fresh frozen plasma (FFP; 30 ml/kg/d) until normalization of the platelet count. Unresponsive patients were referred for plasma exchange. Compared with HIV- TTP patients (n=23), in HIV+ subjects (n=21) microangiopathy was dominant among Black females, who had lower presentation Hb (median 5.8 g/dl; P=0.03), platelet count (13 x 10(9)/l; P=0.05) and a CD4 count of 0.096 x 10(9)/l. HIV+ individuals responded to FFP faster than HIV- patients and none of them required apheresis. Ten HIV- TTP patients required apheresis (P=0.03) and four died. Responses in the HIV+ and HIV- groups occurred after treatment with a median of 33 and 55 units (one unit=320 ml) of FFP (P=0.004) respectively. Response to this protocol was seen in 84% (95% response in HIV+ patients). Regression analysis showed that survival was associated with younger age (P=0.001), rapid platelet (P=0.001) and Hb (P=0.0009) recovery, and fewer FFP units to normal lactate dehydrogenase levels (P=0.006). We conclude that in HIV+ individuals, microangiopathy is highly responsive to plasma infusions. This observation is important particularly when apheresis is not available. PMID- 15667541 TI - Severe FVII deficiency caused by a new point mutation combined with a previously undetected gene deletion. AB - A 3-week-old Caucasian female presented with severe unprovoked parenchymal cerebral haemorrhage. Her plasma factor VII (FVII) activity was <0.01 units/ml. FVII activities for her mother and sister were 0.65 units/ml and 0.51 units/ml, respectively, while her father's level was normal. These results indicated that the mother was heterozygous for a non-functional F7 gene that had also been inherited by the proband's sister. The proband's severe FVII deficiency was caused by a new mutation in her paternal F7 gene coupled with the inheritance of the non-functional maternal F7 gene. DNA sequence analysis revealed that the proband had apparent homozygosity for a novel single point mutation (g.3907G >A) changing the codon for Glu29 to Lys (E29K); neither parent had the E29K mutation. Because of the unlikelihood that the proband was homozygous for two identical new point mutations, the DNA sequence abnormality was more likely to have arisen from a single mutated gene on one allele and a F7 gene deletion on the other allele. Real time polymerase chain reaction (PCR) analysis confirmed that the proband had inherited a gene deletion that was present in the maternal side of the family. Subsequent clotting assays and real time PCR revealed that the maternal deletion also included the closely linked F10 gene. PMID- 15667542 TI - Factor V Leiden, prothrombin 20210A and the risk of venous thrombosis among cancer patients. AB - Cancer patients have an increased risk of venous thrombosis (VT). The association of factor V Leiden (FVL) and the prothrombin 20210A variant with VT in cancer patients is not established. We genotyped 101 cancer patients with VT and 101 cancer patients without VT for these polymorphisms. Five cases and three controls were heterozygous for FVL, yielding an odds ratio of 1.7 (95% confidence interval (CI) 0.3-10.7). Five cases and no controls were heterozygous for prothrombin 20210A, for an odds ratio of 6.7 (95% CI 0.9-infinity). Prothrombin 20210A may be associated with VT risk among cancer patients. PMID- 15667543 TI - Tolerance to intravenous heparin in patients with delayed-type hypersensitivity to heparins: a prospective study. AB - Delayed-type hypersensitivity to subcutaneously injected heparin is relatively common. Particularly, extensive cross-reactivity between different heparins and heparinoids often occurs. Delayed-type hypersensitivity to heparin implies the risk of a generalized eczema when heparin is administered intravenously. However, case reports demonstrated a tolerance to intravenous heparin in patients with delayed-type hypersensitivity to subcutaneous heparin, but prospective studies have not been performed. Our study group, of 28 patients with a proven delayed type hypersensitivity to subcutaneous heparin, was challenged with intravenous heparin, which was well tolerated in all 28 patients. Therefore, in case of therapeutic necessity, the shift from subcutaneous to intravenous heparin administration is justified. PMID- 15667544 TI - Pre-transfusion management of children with severe malarial anaemia: a randomised controlled trial of intravascular volume expansion. AB - Symptomatic severe malarial anaemia (SMA) has a high fatality rate of 30-40%; most deaths occur in children awaiting blood transfusion. Blood transfusion services in most of Africa are not capable of delivering adequate supplies of safe blood in a timely manner to critically ill children with SMA. Contrary to widely held belief, hypovolaemia, rather than heart failure, has emerged as a common complication in such children. We examined the safety of pre-transfusion management (PTM) by volume expansion, aimed at stabilizing children and obviating the urgency for blood transfusion. Kenyan children with severe falciparum anaemia (haemoglobin <5 g/dl) and respiratory distress were randomly assigned to 20 ml/kg of 4.5% albumin or 0.9% saline or maintenance only (control) while awaiting blood transfusion. PTM was apparently safe since it did not lead to the development of pulmonary oedema or other adverse events. There was no significant difference in the primary outcome [mean percentage reduction in base excess between admission and 8 h (95% confidence interval)] between the control group 42% (19-66%) albumin group 44% (32-57%) and saline group 36% (16-57%); adjusted analysis of variance F=0.31, P=0.7. However, the number of children requiring emergency interventions was significantly greater in the control group, four of 18 (22%) than the saline group 0 of 20 (P=0.03). We have established the safety of this PTM in children with SMA whilst awaiting blood transfusion at a hospital with an adequate blood banking program. The impact on mortality should be assessed where blood transfusion services are unable to supply emergency transfusions. PMID- 15667545 TI - Inactivation of parvovirus B19 by liquid heating incorporated in the manufacturing process of human intravenous immunoglobulin preparations. AB - Several reports have suggested the possible transmission of human parvovirus B19 (B19) through the administration of plasma derivatives that had undergone virus inactivation by various types of heat treatment. However, none of the reports evaluated and discussed the inactivation of B19 by the heat treatment that is implemented in the individual manufacturing processes of such products. The present study evaluated the ability to inactivate B19 of liquid-heat treatment at 60 degrees C for 10 h that was incorporated in the manufacturing process of intravenous human immunoglobulin preparations. The results showed that B19 was rapidly inactivated under the conditions used for the liquid-heat treatment. PMID- 15667546 TI - Serum free light chains for monitoring multiple myeloma. PMID- 15667550 TI - Should we eradicate Helicobacter pylori infection in patients receiving nonsteroidal anti-inflammatory drugs or low-dose aspirin? AB - Whether Helicobacter pylori infection alters the risk of ulcer disease in patients receiving nonsteroidal anti-inflammatory drugs (NSAIDs) or low-dose aspirin is one of the most controversial topics in peptic ulcer research. This is an important management issue, particularly in countries where peptic ulcer disease is common and the prevalence of H. pylori infection is high. Current evidence shows that H. pylori infection increases the ulcer risk associated with NSAIDs or low-dose aspirin. Eradication of H. pylori reduces the subsequent risk of endoscopic and complicated ulcers in patients who are about to start long-term NSAIDs. Among patients with H. pylori infection and a history of ulcer bleeding who continue to use low-dose aspirin, 1 week of eradication therapy prevents recurrent ulcer bleeding. Failure of eradication and concomitant use of NSAIDs, however, account for most cases of recurrent bleeding with low-dose aspirin. The apparent protective effect of H. pylori in long-term NSAIDs users reported in some studies was actually the weeding out of susceptible patients who were intolerant to NSAIDs. There is no convincing evidence that eradication of H. pylori has any clinically important adverse effect on the healing and prevention of ulcers in NSAIDs users. PMID- 15667548 TI - The co-expression of PML/RAR alpha and AML1/ETO fusion genes is associated with ATRA resistance. PMID- 15667551 TI - Inflammatory bowel disease pathogenesis: therapeutic implications. AB - The pathogenesis of inflammatory bowel disease (IBD) is complex, involving environmental, genetic, microbial, and immune factors. Therefore, treatment should target components that either predispose to or mediate the chronic inflammatory response of IBD. At the moment it is assumed that all components are necessary to have the typical manifestations of IBD but, in reality, it is unclear to what extent each factor contributes to the disease process, and whether some are more important than others. In addition, some factors are not practical targets; for example, environmental factors are poorly defined, too numerous, and require changes that cannot be implemented by the physician or the patient alone. The same is true for genetic factors that are still not amenable to therapeutic manipulations for technical and ethical reasons. This leaves microbial and immune factors as the two categories that can be selected for therapeutic intervention and where all current treatments are focused. The commensal gut flora can be qualitatively or quantitatively modified with antibiotics, probiotics, or diet, and a better characterization of enteric bacteria strains should help greatly in developing more effective therapies. Most current drugs are focused on inhibiting pro-inflammatory molecules produced by immune cells, including biological agents that block specific cytokines such as tumor necrosis factor-alpha. It is anticipated that combination therapies targeting multiple pathogenic components will prove more effective than those blocking single components of IBD pathogenesis. PMID- 15667552 TI - Low expression of XIAP-associated factor 1 in human colorectal cancers. AB - OBJECTIVE: Eight cellular homologs of the inhibitors-of-apoptosis proteins (IAP) have been identified in humans and of them, the X-linked IAP (XIAP) is the most potent. XIAP-associated factor 1 (XAF1) is a newly discovered XIAP-binding protein that negatively regulates the caspase-inhibiting activity of XIAP. It is either not expressed or present at extremely low levels in many cancer cell lines. The aims of the present study were: (i) to investigate the expression of XAF1 in human colorectal cancers (CRC) both in vitro and in vivo, and (ii) to evaluate the possibility of XAF1 as a new tumor marker. METHODS: The expression of XAF1 in four human colon cancer cell lines (Colo205, Colo320, SW1116, LoVo) and in samples from 70 patients with CRC was analyzed by reverse transcriptase polymerase chain reaction. XAF1 concentrations were also detected in the peripheral circulation of the 70 patients, as well as three traditional circulating cancer-associated antigens. RESULTS: A low concentration of XAF1 mRNA was detectable in the three colon cancer cell lines other than Colo205, which showed the strongest expression of XAF1. The expression of XAF1 in tissue was relatively lower in primary CRC compared with a relatively higher level in benign colorectal tumors (P < 0.01). Although the XAF1 expression in circulation of those with CRC was also lower than in those with benign tumors, there was no statistical significance (P > 0.05). CONCLUSIONS: The present results suggest that the low expression of XAF1 in tumor tissue coincides with a similar level in the peripheral circulation, which contributes at least part to the malignant behavior of CRC. Integrating the XAF1 relative expression value with the other three traditional tumor biomarkers created a four-parameter assay that significantly improved the rate of diagnosis of CRC. PMID- 15667553 TI - Experimental study of the killing effects of oxymatrine on human colon cancer cell line SW1116. AB - OBJECTIVE: To study the killing effects of oxymatrine (OM) on a human colon cancer cell line, SW1116, and evaluate its antineoplastic mechanism. METHODS: Methyl thiazolyl tetrazolium (MTT) analysis, flow cytometry, polymerase chain reaction (PCR)-enzyme linked immunosorbent assay (ELISA) and RT-PCR methods were used respectively to determine the killing effects of OM and its influence on cell cycle distribution, telomerase activity and the expressions of hTERT, c-myc, p53 and mad1 in SW1116 cells. RESULTS: Oxymatrine exhibited dose-dependent killing effects on SW1116 cells and induced G1/G0-phase arrest. It suppressed the telomerase activity of the cells in a dose- and time-dependent manner. After OM administration, the expression of hTERT in the SW1116 cells decreased, those of p53 and mad1 increased, and the expression of c-myc was unchanged. CONCLUSIONS: Oxymatrine has dose-dependent killing effects on SW1116 cells and its antineoplastic activity might be attributed to inhibition of telomerase activity by means of its effects on hTERT and the upstream regulating genes. PMID- 15667554 TI - Antinociceptive property of tegaserod in a rat model of chronic visceral hypersensitivity. AB - OBJECTIVE: To establish a model of chronic visceral hypersensitivity in rats and to investigate the effect of tegaserod, a partial 5-hydroxytryptamine-4 receptor agonist, on visceral hypersensitivity. METHODS: Neonate Sprague-Dawley rats at 8 21 days after birth underwent colorectal distension once daily. Adult rats aged 8 10 postnatal weeks underwent colorectal distension and the abdominal withdrawal reflex (AWR) during the distension was determined. The AWR score was recorded before and after intraperitoneal administration of either tegaserod (treatment group: 0.3 mg/kg) or vehicle (control group). RESULTS: Changes in the AWR score were dependent on the pressure intensity of the colorectal distension (P < 0.01). At pressures of 40, 60 and 80 mmHg, the AWR scores in the model rats with visceral hypersensitivity were significantly higher than those recorded in the control group (1.95 +/- 0.16 vs 1.35 +/- 0.15, 2.82 +/- 0.12 vs 2.17 +/- 0.13, 3.20 +/- 0.14 vs 2.59 +/- 0.14, P < 0.01). Compared with the controls, tegaserod significantly decreased the AWR scores at the distension pressures of 40, 60 and 80 mmHg (1.95 +/- 0.50 vs 1.32 +/- 0.55, 3.05 +/- 0.48 vs 2.32 +/- 0.54, 3.25 +/- 0.63 vs 2.77 +/- 0.51, P < 0.05). CONCLUSIONS: Adult rats can develop chronic visceral hypersensitivity after transient colorectal mechanical irritation during their postnatal period. Tegaserod increases the pain threshold to noxious stimuli, suggesting an antinociceptive property in its effect on visceral hypersensitivity. PMID- 15667555 TI - Comparative analysis of patients with clinically diagnosed colorectal cancer and those detected by mass screening. AB - OBJECTIVE: To analyze the differences and similarities between patients with colorectal cancer (CRC) who had been clinically diagnosed (CD) and those detected by mass screening (MS), and thus clarify the practical significance of CRC screening in China. METHODS: All the data for the CRC patients in the CD group were collected by registration through a cohort study, and the patients in the MS group were selected from mass screening of a natural population of 19 852 in Beijing Metropolitan area. CRC was confirmed pathologically in both groups. RESULTS: Patients over the age of 50 years with CRC comprised 81.9% of the CD group and 83.3% of the MS group; the percentage of Dukes A + B was 41.1% and 91.7%, respectively; 0.28% of individuals with risk factors in the MS group were found to have CRC, whereas 0.05% did not (P < 0.05). Endoscopically, 44.7% of the protruding type CRC were Dukes A or B compared with only 10% of the ulcerated type (chi(2) = 22.304, P < 0.001). The positivity rate of the fecal occult blood (FOB) test was 79.9% in Dukes A + B and 82.0% in Dukes C + D, showing no difference between the FOB test and Dukes staging in diagnosing CRC (chi(2) = 0.087, P = 0.768). The percentage of patients with increased serum concentration of carcinoembryonic antigen in Dukes C + D was significantly higher than that in Dukes A + B (chi(2) = 5.547, P = 0.019). Among the moderately and well differentiated CRC, 51.8% (159/307) were Dukes A + B, resulting in a significant difference (chi(2) = 14.504, P < 0.001). There was a familial history of malignancies in 17.9% of CRC cases and 14.6% of their first-degree relatives suffered from malignant tumors. CONCLUSIONS: The incidence of CRC in Beijing Metropolitan area has been rising significantly. The staging and prognosis of CRC involves symptomatology, laboratory tests, endoscopic findings and pathologic characteristics. Mass screening is effective for detecting early CRC in China. PMID- 15667556 TI - Metal-induced oxidative damage in cultured hepatocytes and hepatic lysosomal fraction: beneficial effect of a curcumin/absinthium compound. AB - OBJECTIVE: Metals undergo redox cycling and there is increasing evidence of free radical generation and oxidative injury in the pathogenesis of liver injury and fibrosis in metal storage diseases. The aim of the present study was to test a natural hepatoprotective compound in metal-induced liver injury. METHODS: Hepatocytes were isolated from Wistar rats by collagenase perfusion method and cultured as such and also with alpha-linolenic acid (LNA)-bovine serum albumin (BSA). Hepatocytes were then cultured with a graded dilution of PN-M001 (100 microg/mL and 200 microg/mL), which is a curcuma/absinthium-containing compound, or sylibin (100 microg/mL) dissolved in dimethyl sulfoxide for 10 min before the addition of metallic salts (iron, copper and vanadium). Lysosomal fractions were prepared for lysosome fragility tests in which beta-galactosidase activity and lactate dehydrogenase (LDH) leakage were measured, as well as oxidative damage tests in the presence of hydrophilic and lipophilic free radical generators. Quenching activity by 1,1-diphenyl-2-picrylhydrazyl (DPPH) was also assessed. RESULTS: Malonildialdehyde accumulation in the medium showed a direct time-course increase with incubation time. Both PN-M001 and sylibin showed a significant protective effect against all challenge metal ions, as expressed by the half inhibition concentration (IC(50)) against lipid peroxidation. However, on a molar ratio, sylibin seemed to be more effective than PN-M001 in Fe-induced peroxidative damage (P < 0.05). Both test compounds, irrespective of the concentration, significantly reduced the LDH and beta-galactosidase concentration in the lysosomal fractions. As compared with untreated lysosomal fractions challenged with the two peroxide radicals generators, either PN-M001 or sylibin exerted significant protection However, PN-M001 was significantly better than sylibin in suppressing acid phosphatase enzyme activity. Both compounds showed comparable and significant DPPH radical-scavenging activity. CONCLUSION: These data support the potential clinical application of curcumin-containing compounds. PMID- 15667557 TI - Modulation of the molecular conformation of a hepatocyte-targeting gene drug in order to improve its expression efficiency in vitro. AB - AIM: To construct different conformations of a plasmid DNA/vector complex (pcDNA3.1/IFN-gamma-ASOR-PLL) and transfect cells of the hepatoma cell line BEL7402 to investigate the optimal conformation of the complex for improved expression efficiency in the target cell. METHODS: Double-distilled water and adjuvant were added to the naked pcDNA3.1/IFN-gamma, target vector ASOR-PLL and the ASOR-PLL-pcDNA3.1/IFN-gamma complex to create different conformations; molecules that were transfected into BEL7402 cells and the expression efficiency was determined by measuring the IFN-g concentration in the culture supernatant by ELISA. RESULTS: Naked pcDNA3.1/IFN-gamma DNA distributed linearly in double distilled water and condensed into a mica configuration in adjuvant; ASOR-PLL had a net-like distribution without adjuvant and a spider-like form in the adjuvant treated group; the ASOR-PLL-pcDNA3.1/IFN-g complex had a divaricate form without adjuvant, but a bead-like or granular conformation in 0.1 and 0.2 mol/L of adjuvant, a homogeneous bacilliform or chromatoid-shaped conformation in 0.3 mol/L adjuvant, and varied shapes in 0.4 and 0.5 mol/L adjuvant. The supernatant IFN-gamma expression in the bacilliform/chromatoid conformation complex group was the highest among the different conformation groups and controls. When chloroquine was added the supernatant IFN-gamma concentration increased in the liposome group and decreased in the bacilliform/chromatoid conformation group . CONCLUSIONS: The two structural molecules and their complex, ASOR-PLL pcDNA3.1/IFN-gamma, were adjustable in the liquid mode. The specific bacilliform/chromatoid conformation of complex was lysosome enzyme-resistant and could play an active role in improving the efficiency of gene expression. The hypothesis that a chromosome-like conformation of the target gene molecule is involved in enhancing exogenous gene expression is proposed. PMID- 15667558 TI - Clinical characteristics of acute pancreatitis patients with elevated serum triglyceride concentration. AB - OBJECTIVE: To evaluate the clinical characteristics of acute pancreatitis (AP) patients with elevated serum triglyceride (TG) concentration. METHODS: Ninety nine cases of AP admitted from January 2000 to January 2002 were analyzed: 28 cases comprised the TG-elevated group (serum TG >1.7 mmol/L) and 71 cases were the TG-normal group (serum TG 3500 cloned VDJs (from 11 animals at three time-points, using three to five animals per time-point) that were expressed with immunoglobulin (Ig)M, IgD, IgG, IgA and IgE in thymus, mesenteric lymph nodes (MLN) and peripheral blood B cells (PBB) of newborn piglets and 5-week-old isolator piglets maintained germfree (GF) or colonized with Escherichia coli. The results showed that the repertoire expressed with IgM, IgD, IgG and IgA in PBB and MLN remained polyclonal, undiversified and unselected in piglets maintained GF for 5 weeks, that age and colonization resulted in significant repertoire diversification of IgG and IgA in the MLN and of IgG in blood, that the thymic B-cell repertoire was polyclonal, unaffected by colonization and showed no clonal selection in any isotype, and that the thymic IgA and IgE repertoires were more diverse at birth than the repertoire of any isotype in MLN or PBB. IgD was seldom recovered from the PBB of newborn piglets or at any time-point in thymus, but was recovered in the MLN of all 11 animals examined. The IgD and IgM repertoires in all tissues remained polyclonal and unselected, although V(H) usage by IgD transcripts did not always parallel that of IgM in the same tissue. Therefore, isotype-switched B cells in the thymic medulla cannot be accounted for by immigration of B cells diversified by colonization of the gut, and thymic B cells undergo switch recombination and repertoire diversification before birth without clonal selection. PMID- 15667563 TI - Comparison of the expressed porcine Vbeta and Jbeta repertoire of thymocytes and peripheral T cells. AB - Transcripts of more than 300 unique T-cell receptor-beta (TCR-beta) V-D-J rearrangements recovered from porcine thymocytes and peripheral T cells were compared. We identified 19 groups (families) of porcine Vbeta genes in seven supergroups and provisionally named 17 groups based on their sequence similarity with recognized human Vbeta gene families. TRBV4S, 5S, 7S and 12S accounted for >80% of all Vbeta usage, and usage of these groups by thymocytes and peripheral T cells was highly correlated. No TRBV group was uniquely expressed in significant numbers in thymocytes, although small numbers of TRBV groups 2S, 9S and 15S were only recovered from T cells. Usage of Jbeta segments from the 5' D-J-C duplicon in thymocytes and peripheral T cells directly correlated with their 5' position in the locus, and Jbeta1.1, 1.2 and 1.3 accounted for >or= 35% of all Jbeta usage in both cell types. This contrasts with the usage of Jbeta2 segments in that Jbeta2.4, 2.5 and 2.7 accounted for approximately 30% of Jbeta usage by T cells and thymocytes. Jbeta2.7 was threefold more frequent among T cells than thymocytes. The Vbeta/Jbeta combination was not random. Jbeta1.1 and 1.2 were used in 29% of rearrangements with high frequency among the major Vbeta groups. Combinations of TRBV4 and V12 with Jbeta2.7 were only found in T cells and accounted for half of all Jbeta2.7 usage. These studies show that unlike porcine heavy chain V(H) genes, the occurrence and relative usage of porcine TCR-Vbeta groups resembles that of humans. Thus, highly related gene systems can individually diverge within a species. PMID- 15667564 TI - Major histocompatibility class II molecules prevent destructive processing of exogenous peptides at the cell surface of macrophages for presentation to CD4 T cells. AB - We studied factors affecting major histocompatibility complex class II (MHC-II) restricted presentation of exogenous peptides at the surface of macrophages. We have previously shown that peptide presentation is modulated by surface associated proteolytic enzymes, and in this report the role of the binding of MHC II molecules in preventing proteolysis of exogenous synthetic peptides was addressed. Two peptides containing CD4 T-cell epitopes were incubated with fixed macrophages expressing binding and non-binding MHC-II, and supernatants were analysed by high-performance liquid chromatography and mass spectrometry to monitor peptide degradation. The proportion of full-length peptides that were degraded and the number of peptide fragments increased when non-binding macrophages were used, leading to reduction in peptide presentation. When MHC-II molecules expressed on the surface of fixed macrophages were blocked with monoclonal antibody and incubated with peptides and the supernatants were transferred to fixed macrophages, a significant reduction in peptide presentation was observed. Peptide presentation was up-regulated at pH 5.5 compared to neutral pH, and the latter was found to be the pH optimum of the proteolytic activity of the surface enzymes involved in the degradation of exogenous peptides and proteins. The data suggest that MHC-II alleles that bind peptides protect them from degradation at the antigen-presenting cell surface for presentation to CD4 T cells and we argue that this mechanism could be particularly pronounced at sites of inflammation. PMID- 15667565 TI - Human monocyte isolation methods influence cytokine production from in vitro generated dendritic cells. AB - There is growing interest in the in vitro generation of dendritic cells (DC) from peripheral blood monocytes, but the effect of the method chosen to isolate CD14+ monocytes for subsequent DC generation is poorly documented. The method used to isolate monocytes may have an impact on the subsequent function of DC by affecting their ability to express costimulatory molecules (CD80/86), maturation marker (CD83) and/or to produce important immunomodulatory cytokines. In this study, we show that the positive selection of monocytes by anti-CD14-coated microbeads inhibits the lipopolysaccharide (LPS)-induced production of interleukin (IL)-12, IL-10 and tumour necrosis factor-alpha (TNF-alpha) from human DC. However, when DC were grown from monocytes isolated by plastic adherence, LPS induced the production of much higher levels of these cytokines. DC derived from adherence-isolated monocytes induced the development of potent cytotoxic T lymphocytes of the Tc1 subset specific for influenza matrix protein, as confirmed by interferon-gamma (IFN-gamma) enzyme-linked immunosorbent spot forming cell assay (ELISPOT), cytotoxicity assay, major histocompatibility complex (MHC)-peptide tetrameric complexes and T helper 1/T helper 2 (Th1/Th2) cytokine production assays. PMID- 15667566 TI - Mucosal delivery of CpG oligodeoxynucleotides expands functional dendritic cells and macrophages in the vagina. AB - Antigen-presenting cells (APC) are specialized sentinel cells that sense pathogens within tissues and then activate appropriate immune effector cells in lymphoid organs. Recent evidence, however, suggests that APC can also induce effector cells in non-lymphoid organs. The purpose of this study was to determine the effect of intravaginal (IVAG) delivery of CpG-oligodeoxynucleotide (ODN) on expansion of resident genital APC. Our results show that delivery of CpG-ODN to the murine genital tract induced a rapid and significant, but transient expansion of genital APC in situ. As early as 12 hr post CpG-ODN delivery, we observed an enhanced level of F4/80+ major histocompatibility complex (MHC) class II-negative macrophages in the genital tissue. This was followed by increased levels of F4/80/MHC class II double-positive cells, as well as MHC class II, CD11c and CD86 triple-positive dendritic cells (DC) at 48 hr. Expanded APC levels at 48 hr post CpG-ODN resulted in increased ability of genital cells to induce an allogenic mixed leucocyte reaction. By 72 hr after CpG-ODN treatment, APC levels were not distinguishable from naive levels. Therefore, these results clearly show that administration of CpG-ODN to the genital tract induced a marked but transient enhancement of APC within the genital tissue, and that these APC appear to possess functional capacity. Furthermore, these results indicate that IVAG-CpG ODN may be an important factor for the enhancement of local antigen presentation in the genital tract through increased DC numbers. PMID- 15667567 TI - Follicular dendritic cell dedifferentiation reduces scrapie susceptibility following inoculation via the skin. AB - Transmissible spongiform encephalopathies (TSEs) are a group of subacute infectious neurodegenerative diseases that are characterized by the accumulation in affected tissues of PrP(Sc), an abnormal isoform of the host prion protein (PrPc). Following peripheral exposure, TSE infectivity and PrP(Sc) usually accumulate in lymphoid tissues prior to neuroinvasion. Studies in mice have shown that exposure through scarified skin is an effective means of TSE transmission. Following inoculation via the skin, a functional immune system is critical for the transmission of TSEs to the brain, but until now, it has not been known which components of the immune system are required for efficient neuroinvasion. Temporary dedifferentiation of follicular dendritic cells (FDCs) by treatment with an inhibitor of the lymphotoxin-beta receptor signalling pathway (LTbetaR Ig) 3 days before or 14 days after inoculation via the skin, blocked the early accumulation of PrP(Sc) and TSE infectivity within the draining lymph node. Furthermore, in the temporary absence of FDCs before inoculation, disease susceptibility was reduced and survival time significantly extended. Treatment with LTbetaR-Ig 14 days after TSE inoculation also significantly extended the disease incubation period. However, treatment 42 days after inoculation did not affect disease susceptibility or survival time, suggesting that the infection may have already have spread to the nervous system. Together these data show that FDCs are essential for the accumulation of PrP(Sc) and infectivity within lymphoid tissues and subsequent neuroinvasion following TSE exposure via the skin. PMID- 15667568 TI - Tick saliva inhibits differentiation, maturation and function of murine bone marrow-derived dendritic cells. AB - Haematophagous arthropod vectors such as mosquitoes, tsetse flies, sandflies and ticks have evolved salivary immunomodulatory factors that prevent the vertebrate host from rejecting them meanwhile enhancing pathogen transmission. As dendritic cells (DC) play a major role in host immune responses, we studied the effects of Rhipicephalus sanguineus tick saliva on DC differentiation and maturation. Flow cytometry analysis revealed that the addition of saliva to bone marrow cells inhibits the differentiation of DC and decreased the population of differentiated immature DC, increasing the levels of major histocompatibility complex (MHC) class II while not altering the expression of costimulatory (CD40, CD80 and CD86) and adhesion (CD54) molecules. Furthermore, maturation of DC stimulated by lipopolysaccharide (LPS) in the presence of saliva resulted in a lower expression of costimulatory molecules, but did not alter the up-regulation of MHC class II and CD54. The lipopolysaccharide (LPS)-matured DC cultured with saliva also presented reduced production of interleukin-12, whereas interleukin-10 production was unaltered. Assessment of the function of DC cultured with tick saliva revealed them to be poor stimulators of cytokine production by antigen-specific T cells. Our data indicate a novel modulatory role for the saliva of arthropod vectors at an initial step of the immune response through the inhibition of differentiation and maturation of DC into functional antigen-presenting cells. PMID- 15667569 TI - Intranasal inoculation of Chlamydia trachomatis mouse pneumonitis agent induces significant neutrophil infiltration which is not efficient in controlling the infection in mice. AB - Previous studies have shown that chlamydial infection is accompanied by significant infiltration of neutrophils at the site of infection. However, the role of neutrophils in host defence against chlamydial infection is not clearly understood. Using genetically different inbred mouse strains and CXCR-2 gene knockout (KO) mice, we examined the mechanism for neutrophil recruitment and the role of neutrophils during chlamydial lung infection. Our data showed that C3H mice exhibited significantly higher and more persistent neutrophil infiltration in the lung than did C57BL/6 mice following Chlamydia trachomatis mouse pneumonitis infection. The massive neutrophil infiltration in C3H mice was paralleled by high-level expression of CXCR-2 and its ligands, CXC chemokines (macrophage inflammatory protein 2, cytokine-induced neutrophil attractant (KC) and lipopolysaccharide-induced CXC chemokine), and proinflammatory cytokines (tumour necrosis factor-alpha, interleukin-1 and interleukin-6) in the lung. Although much greater infiltration of neutrophils was observed in C3H mice than in C57BL/6 mice, the former mice had more severe disease and higher in vivo chlamydial growth than the latter. Moreover, CXCR-2 KO mice, which revealed a dramatic reduction in neutrophil activity, showed comparable chlamydial infection to wild-type mice. These results suggest that neutrophils are not efficient for controlling chlamydial lung infection. PMID- 15667570 TI - Significance of Fas and Fas ligand in tuberculous lymphadenitis. AB - The Fas/Fas-ligand (FasL) system plays an important role in regulation of apoptosis and the immune response, and is exploited by mycobacteria to evade the immune response. This study was performed to investigate the distribution and levels of FasL and Fas in lymph node granulomas and sera of tuberculous lymphadenitis patients by immunohistochemistry and enzyme-linked immunosorbent assay. The validity of soluble Fas (sFas) or soluble FasL (sFasL) as a diagnostic tool was also examined. Levels of sFasL in serum were elevated among patients. The numbers of FasL stained cells in lymph node granulomas were higher than Fas. Children had significantly higher levels of sFasL as compared to adults. The human immunodeficiency virus (HIV)-tuberculosis (TB)-coinfected patients displayed no differences in the levels of sFasL or sFas compared with HIV negative patients. The healthy controls from a high endemic tuberculosis country (having latent TB) had significantly higher levels of sFasL than from a country with no TB transmission. The sensitivity and specificity of the FasL and Fas test were low when compared with the culture results as the gold standard. However, by using histology as the gold standard, the sensitivity and specificity of the FasL test were increased to 66.7% and 100%, respectively, but for the Fas test remained low. In conclusion, sFasL and sFas cannot be used as diagnostic tests for tuberculous lymphadenitis. However, its utility in detecting latent TB and childhood tuberculous lymphadenitis remains to be evaluated. FasL seems to play a role in immune modulation and pathogenesis of TB. Modulators of Fas/FasL-mediated apoptosis may therefore be clinically useful. PMID- 15667571 TI - Haptoglobin dampens endotoxin-induced inflammatory effects both in vitro and in vivo. AB - We report that haptoglobin, an acute-phase protein produced by liver cells in response to interleukin-6 (IL-6), can modulate the inflammatory response induced by endotoxins. We provide evidence that haptoglobin has the ability to selectively antagonize lipopolysaccharide (LPS) effects in vitro by suppressing monocyte production of tumour necrosis factor-alpha, IL-10 and IL-12, while it fails to inhibit the production of IL-6, IL-8 and IL-1 receptor antagonist. In two animal models of LPS-induced bronchopulmonary hyperreactivity and endotoxic shock, haptoglobin knockout mice were more sensitive to LPS effects compared to their wild-type counterparts. The present data suggest that haptoglobin regulates monocyte activation following LPS stimulation. The increase in haptoglobin levels during an acute-phase reaction may generate a feedback effect which dampens the severity of cytokine release and protects against endotoxin-induced effects. PMID- 15667572 TI - LIGHT is involved in the pathogenesis of rheumatoid arthritis by inducing the expression of pro-inflammatory cytokines and MMP-9 in macrophages. AB - Macrophages play a crucial role in the perpetuation of inflammation and irreversible cartilage damage during the development of rheumatoid arthritis (RA). LIGHT (TNFSF14) and its receptor TR2 (TNFRSF14) are known to have pro inflammatory activities in foam cells of atherosclerotic plaques. We tested a hypothesis that LIGHT and TR2 are involved in activation of monocyte/macrophages in RA synovium. Immunohistochemical analysis of RA synovial tissue samples revealed that both LIGHT and TR2 are expressed in CD68 positive macrophages. In contrast, synovial tissue samples from osteoarthritis (OA) patients failed to reveal the expression of LIGHT. Expression of TR2 in RA synovial macrophages was also detected using flow cytometry analysis. To identify the role of LIGHT in the functioning of macrophages in RA, we isolated macrophage enriched cells from RA synovial fluid and stimulated them with LIGHT. LIGHT induced expression of matrix metalloproteinase-9 and pro-inflammatory cytokines such as tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, and IL-8. These data indicate that LIGHT and TR2 expressed in macrophages are involved in the pathogenesis of RA by inducing the expression pro-inflammatory cytokines and matrix degrading enzymes. PMID- 15667573 TI - Glycation of CD59 impairs complement regulation on erythrocytes from diabetic subjects. AB - Type 1 diabetes is associated with anaemia. Although the underlying mechanisms remain unclear, the accompanying reticulocytosis implies that erythrocyte lifespan in the circulation is shortened. Among the factors that permit prolonged survival of erythrocytes are the membrane complement regulators. In conditions such as paroxysmal nocturnal haemoglobinuria, where erythrocyte expression of these regulators is reduced, erythrocyte survival is compromised and anaemia follows. Recent in vitro evidence indicates that one of the key membrane complement regulators, CD59, is inactivated by glycation in the presence of high concentrations of glucose or other glycating sugars. To ascertain whether glycation-induced inactivation of CD59 occurrs in vivo we examined CD59 surface expression and function on erythrocytes from a cohort with poorly controlled type 1 diabetes (hyperglycaemic) and from matched normoglycaemic controls. Although expression of CD59, assessed using polyclonal anti-CD59 antiserum, was similar in the two groups, erythrocytes from hyperglycaemic individuals were more susceptible to lysis by complement, entirely as a result of the loss of functional CD59. These data implicate glycation-induced inactivation of CD59 as a factor contributing to anaemia in type 1 diabetes. PMID- 15667575 TI - Asymmetric dimethylarginine (ADMA): the silent transition from an 'uraemic toxin' to a global cardiovascular risk molecule. AB - Endothelial dysfunction as a result of reduced bioavailability of nitric oxide (NO) plays a central role in the process of atherosclerotic vascular disease. In endothelial cells NO is synthesized from the amino acid l-arginine by the action of the NO synthase (NOS), which can be blocked by endogenous inhibitors such as asymmetric dimethylarginine (ADMA). Acute systemic administration of ADMA to healthy subjects significantly reduces NO generation, and causes an increase in systemic vascular resistance and blood pressure. Increased plasma ADMA levels as a result of reduced renal excretion have been associated with atherosclerotic complications in patients with terminal renal failure. However, a significant relationship between ADMA and traditional cardiovascular risk factors such as advanced age, high blood pressure and serum LDL-cholesterol, has been documented even in individuals without manifest renal dysfunction. As a consequence, the metabolism of ADMA by the enzyme dimethylarginine dimethylaminohydrolase (DDAH) has come into the focus of cardiovascular research. It has been proposed that dysregulation of DDAH with consecutive increase in plasma ADMA concentration and chronic NOS inhibition is a common pathophysiological pathway in numerous clinical conditions. Thus, ADMA has emerged as a potential mediator of atherosclerotic complications in patients with coronary heart disease, peripheral vascular disease, stroke, etc., being the culprit and not only an innocent biochemical marker of the atherosclerotic disease process. PMID- 15667577 TI - Ninth edition of the G.B. Morgagni Awards Program. PMID- 15667574 TI - Aberrant regulation of interleukin-12 receptor beta2 chain on type 1 cytokine stimulated T lymphocytes in type 1 diabetes. AB - An aberrant mitogen-induced polarization of peripheral blood T cells has been associated with type 1 diabetes (T1D). We studied, in T1D, type 1 and 2 cytokine induced expression of the interleukin-12 receptor beta2 chain (IL-12Rbeta2 chain), which plays a critical role in regulating T-cell polarization. Peripheral blood lymphocytes from children with newly diagnosed T1D (n=10; mean age 10 years), from children with longstanding T1D (n=8; mean age 12.9 years) and from healthy children (n=15; mean age 11.5 years) were stimulated with phytohaemagglutinin (PHA) in a type 1 (IL-12 and anti-IL-4) or a type 2 (IL-4 and anti-IL-12) cytokine environment. Secretion of interferon-gamma (IFN-gamma), IL-5 and IL-13, as detected by enzyme-linked immunosorbent assay (ELISA), and expression of the IL-12Rbeta2 chain on CD4 and CD8 cells by flow cytometry, were analysed. Children with newly diagnosed and longstanding T1D had lower expression levels of the IL-12Rbeta2 chain on IL-12Rbeta2 chain-positive CD4 T cells (for a type 1 or a type 2 cytokine environment: P=0.01 and P=0.002 or P=0.02 and P=0.01, respectively) and on IL-12Rbeta2 chain-positive CD8 T cells (for a type 1 or a type 2 cytokine environment: P=0.007 and P=0.0007 or P=0.003 and P=0.01, respectively) when compared to healthy children. A decreased percentage of IL 12Rbeta2 chain-expressing CD4 T cells (P=0.07 and P=0.03) and CD8 T cells (P=0.004 and P=0.01) and increased secretion of IL-13 (P=0.006 and P=0.04) in a type 1 cytokine environment was seen in both groups of patients. Peripheral blood T cells from patients with both newly diagnosed and longstanding T1D showed poor polarization towards type 1 cells. PMID- 15667576 TI - The G.B. Morgagni Medal 2004. PMID- 15667578 TI - Peroxisome proliferator-activated receptor gamma: the more the merrier? AB - The consequence of activating the nuclear hormone receptor, peroxisome proliferator-activated receptor gamma (PPARgamma), which coordinates adipocyte differentiation, validates the concept, 'you are what you eat'. Excessive caloric intake leads to fat formation if the energy from these nutrients is not expended. However, this evolutionary adaptation to store energy in fat, which can be released under the form of fatty acids, potent PPARgamma agonists, has become a disadvantage in today's affluent society as it results in numerous metabolic imbalances, collectively known as the metabolic syndrome. With the surge of human and genetic studies on PPARgamma function, the limitations to the benefits of PPARgamma signalling have been realized. It is now evident that the most effective strategy for resetting the balance of this thrifty gene is through its modulation rather than full activation, with the goal to improve glucose homeostasis while preventing adipogenesis. Finally, as more PPARgamma targeted pathways are revealed such as bone homeostasis, atherosclerosis and longevity, it is most certain that the PPARgamma thrifty gene hypothesis will evolve to incorporate these. PMID- 15667579 TI - Prognostic impact of low-shear whole blood viscosity in hypertensive men. AB - BACKGROUND: The role of blood viscosity as a marker for discriminating cardiovascular risk in essential hypertension remains uncertain. The aim of this study was to assess whether whole blood viscosity (WBV) could be useful in assessing cardiovascular risk in men with a first diagnosis of hypertension. DESIGN: A total of 331 middle-aged men with newly diagnosed essential hypertension (age at entry 40-64 years, average blood pressure 151/95 mmHg) underwent low-shear-rate (0.94 s(-1)) and high-shear-rate (94.5 s(-1)) WBV determination and were then followed for a mean of 4.8 +/- 3 years (range 0-12 years). RESULTS: Cardiovascular event rates in the bottom, middle and top tertiles of the distribution of low-shear WBV were 1.10, 2.13 and 4.43 per 100 patient-years, respectively (log-rank test, P < 0.001). After taking into account several established cardiovascular risk factors in a Cox survival analysis, a raised low-shear WBV conferred an increased risk for cardiovascular events (top vs. bottom tertile hazard ratio = 3.42, 95% confidence interval = 1.4-8.4, P = 0.006; middle vs. bottom tertile hazard ratio = 2.25, 95% confidence interval = 0.9-5.6, P = 0.09). The independent association between high-shear-rate WBV and cardiovascular events bordered statistical significance (P = 0.07). Inclusion in the survival model of low-shear-rate resulted in a significantly greater chi(2) improvement (P < 0.05) than inclusion of high-shear-rate WBV. CONCLUSIONS: In hypertensive men, an increased WBV at low shear rate is a predictor of cardiovascular events independently from the effect of several traditional risk factors. Low-shear WBV is a better discriminator of cardiovascular risk than high shear WBV. PMID- 15667580 TI - Left ventricular and cardiac autonomic function in survivors of testicular cancer. AB - BACKGROUND: Following cisplatin-based chemotherapy, survivors of testicular cancer have a high prevalence of cardiovascular risk factors and an increased risk of cardiovascular disease. Cardiac function has not been extensively studied and no comparisons have been made with men from the general population. DESIGN: Left ventricular and cardiac autonomic function were evaluated in chemotherapy treated testicular cancer patients, in stage I patients after orchidectomy only, and in healthy men using Doppler echocardiography [wall motion score index, peak early (E) and atrial filling (A) velocities across the mitral valve, E/A-ratio, isovolumetric relaxation time, and deceleration time of the early peak flow] and measurements of N-terminal pro-brain natriuretic peptide and baroreflex sensitivity. Furthermore, 24-h ambulatory blood pressure was measured. RESULTS: Ninety chemotherapy-treated patients (median age 37 years, range 20-65; median follow up of 7 years, range 3-13) were compared with 44 stage I patients (median age 36 years, range 24-63) and 47 healthy controls (median age 37 years, range 22 55). Wall motion score index was less than 1.5 in all participants. Chemotherapy treated patients had a higher peak A-wave and a lower E/A-ratio than stage I patients and controls. Isovolumetric relaxation and deceleration times did not differ between groups. Age, 24-h diastolic blood pressure and treatment with chemotherapy were significantly associated with E/A-ratio. Natriuretic peptide levels were normal. Baroreflex sensitivity was similar in the three groups. CONCLUSIONS: Chemotherapy-treated testicular cancer survivors have a lower E/A ratio than healthy subjects from the general population, which may indicate impaired relaxation of the left ventricle and reflect the high prevalence of cardiovascular risk factors previously reported in these men. PMID- 15667581 TI - High levels of C-reactive protein with low total cholesterol concentrations additively predict all-cause mortality in patients with coronary artery disease. AB - BACKGROUND: This study aimed to investigate independent and additive predictive effects of raised C-reactive protein (CRP) levels and decreased total cholesterol levels on mortality in patients with chronic coronary artery disease (CAD). Low total cholesterol (TC) levels are associated with worsened survival in chronic and acute diseases. Elevated CRP level is an important predictor of vascular events and mortality in patients with CAD. Potential inhibition of immune activation by circulating lipoproteins could be a link between cholesterol and inflammatory markers. MATERIALS AND METHODS: A group of 387 patients (median age 59 years) with CAD and with or without severe heart failure (HF) were followed for a median of 5.06 years. Serum total cholesterol and CRP concentrations were measured at enrollment. RESULTS: The relationship between lipoproteins, CRP and survival was explored. High CRP concentrations were in significant association with severity of HF and predicted worsened survival in patients with CAD (hazard ratio 5.214, 95% CI 1.762-15.427). The association between CRP levels and mortality was independent of potential confounding factors such as age, body-mass index, severity of HF, smoking habits, hypertension and TC levels. The prediction of mortality by low TC levels was significant (hazard ratio 2.932, 95% CI 1.021 8.422). Furthermore, patients with increased CRP and decreased TC (additive predictive effect) phenotype had 11.714-times higher risk (95% CI 2.619-52.385) of being nonsurvivors than patients with low CRP/high TC. CONCLUSIONS: High CRP levels and low TC concentrations are independent and additive predictors of mortality in patients with CAD. Our data indicate that joint analysis of circulating lipoproteins and inflammatory biomarkers may improve prediction of survival in patients with CAD. PMID- 15667582 TI - Improved early risk stratification and diagnosis of myocardial infarction, using a novel troponin I assay concept. AB - BACKGROUND: We evaluated the clinical performance of a novel cardiac troponin I (cTnI) assay specifically designed to improve the very early risk stratification in acute coronary syndromes. SUBJECTS AND METHODS: Serum and plasma samples (taken 0, 6-12 h and 24 h after admission) from 531 patients with suspected acute coronary syndrome were studied using a novel investigational cTnI assay, reference cTnI assay and myoglobin. The lowest cTnI concentration giving a total assay imprecision of 10% was used as the positive myocardial infarction (MI) cut off value. RESULTS: At the time of admission, the investigational assay was positive in 27.9% of the patients, the reference cTnI assay was positive in only 17.5% (P < 0.001) and myoglobin in 24.1% (P = 0.067). Receiver operating characteristic (ROC) curve analysis for the detection of myocardial injury on admission gave area-under-curve (AUC) values of 0.937, 0.775 and 0.762, respectively (P < 0.001). Of those MI patients who presented within 3 h of symptom onset, 50.0% were identified by the investigational assay at the time of presentation, compared with 44.2% by myoglobin (P = 0.791) but only 11.5% by the reference assay (P < 0.001). CONCLUSIONS: The novel cTnI assay considerably improves the performance of cTnI as an early rule-in biomarker for MI. PMID- 15667583 TI - Is postprandial hypertriglyceridaemia in relatives of type 2 diabetic subjects a consequence of insulin resistance? AB - BACKGROUND: Higher postprandial triglyceride responses reported in first degree relatives of people with type 2 diabetes (REL) were postulated to be the result of an early, possibly intrinsic, defect in oral lipid handling. The postprandial triglyceride response to high fat meals (HFM) in normal subjects is reduced by the insulin response to dietary carbohydrate (CHO) in the meal. The aims of this study were to examine whether (1) insulin resistance is associated with an intrinsic defect in triglyceride handling in insulin-resistant REL and (2) insulin resistance is associated with altered triglyceride handling after HFM with high CHO content. MATERIALS AND METHODS: Postprandial responses to a HFM in normolipidaemic, normoglycaemic REL were compared with subjects without a family history of diabetes mellitus (CON). Over 6 h, the insulin, glucose, triglyceride and nonesterified fatty acid (NEFA) responses after a high fat (80 g fat), low CHO (HFM-LC; 20 g CHO, 4250 kJ) meal and a high fat, high CHO (HFM-HC; 100 g CHO, 5450 kJ) meal were examined. RESULTS: The 10 (7F/3M) REL were significantly more insulin-resistant, determined by glucose infusion during a hyperinsulinaemic euglycaemic clamp than the 10 (5F/5M) CON (glucose infusion rate 44.6 +/- 4.9 vs. 60.0 +/- 4.8 micromol min(-1) kg FFM(-1), P = 0.037). Subjects were similar for age and body mass index (BMI). The triglyceride increments after the HFM-LC were similar in both, peaking at 180-240 min (Delta0.77 +/- 0.11 mmol L(-1)), demonstrating no postprandial defect in REL, despite insulin resistance. There was a significantly lower postprandial triglyceride response in CON following the HFM-HC compared with the HFM-LC, but not in REL. In contrast, the higher insulin level during the HFM-HC was associated with significantly greater NEFA level suppression than in the HFM-LC (2.13 +/- 0.51 vs. 0.70 +/- 0.35 mmol L(-1), P = 0.03), only in the REL. CONCLUSIONS: These results are inconsistent with a primary aetiological role for postprandial hypertriglyceridaemia in already insulin resistant type 2 diabetic REL, but raise the possibility that this potentially atherogenic manifestation is secondary to insulin resistance lessening VLDL production and/or release from the liver. PMID- 15667584 TI - Autoantibodies against pituitary proteins in patients with adrenocorticotropin deficiency. AB - BACKGROUND: An autoimmune cause of adrenocorticotropin (ACTH)-deficiency is presented, as it is known to be a characteristic feature of lymphocytic hypophysitis, a disease of the pituitary gland considered to be autoimmune. MATERIALS AND METHODS: The aim of this study was twofold: (1) to evaluate the occurrence of pituitary autoantibodies and (2) to correlate it to clinical and immunological features in a large group of patients with ACTH-deficiency of possible autoimmune aetiology. Sixty-five patients with ACTH-deficiency and 57 healthy subjects participated in the study. Pituitary autoantibodies were measured by an immunoblotting assay with human pituitary cytosol as antigen. RESULTS: Autoantibodies to a novel 36-kDa pituitary autoantigen were seen in sera from 18.5% (12/65) patients and only 3.5% (2/57) of control subjects (P = 0.0214). When taking only those subjects with strong immunoreactivity into account, the significance was lost; P = 0.3642. Immunoreactivity to a 49-kDa pituitary autoantigen was observed in 21.5% (14/65) of ACTH-deficient patients compared with 8.8% (5/57) of control subjects (P = 0.0910). This 49-kDa pituitary autoantigen has recently been identified as neurone-specific enolase and a candidate marker for neuroendocrine autoimmunity. Clinical parameters in patients with positive versus those with negative pituitary immunoreactivity did not differ. However, autoantibodies to thyroglobulin were positively correlated to immunoreactivity against the 36-kDa pituitary autoantigen (P = 0.014). CONCLUSIONS: Our findings of pituitary autoantibodies in patients' sera support the theory that an autoimmune destruction of corticotrophs may be the underlying cause of hormonal deficit in some patients with ACTH-deficiency. PMID- 15667585 TI - Tensegrity and type 1 dermal dendrocytes in acromegaly. AB - BACKGROUND: Skin is enlarged and doughy in acromegaly. Alterations have been reported in the structure of the extracellular matrix (ECM) and in the mechanical properties of the dermis. It is recognized that internal tensions in the dermis give rise to active cell-ECM and cell-cell mechanical interactions. This mechanobiological aspect can lead to functional and conformational changes in the cells. Such alterations in connective tissue cells have not been explored so far in acromegaly. The aim of the study was to assess structural changes in the dermal ECM, and in the number and shape of factor XIIIa-positive type I dermal dendrocytes (DDs) in acromegaly. MATERIALS AND METHOD: A total of 32 patients with acromegaly (three untreated, 17 with active disease under somatostatin analogues and 12 cured) were examined. Both the maximum and the most recent IGF-1 (insulin-like growth factor) dosages recorded in the patients' files were retrieved. Skin biopsies were taken from their forearms. Histochemistry, immunohistochemistry, morphometry and electron microscopy were used. RESULTS: Collagen-bundle coarsening and acidic glycosaminoglycan deposits were found irrespective of the current endocrine status. Dermal dendrocytes were often markedly reduced in numbers, but those present were plump with few dendrites. A negative correlation was found between DD numbers and the maximum IGF-1 dosages recorded in the patients' files. A similar although weaker trend was found with the most recent IGF-1 dosages. CONCLUSIONS: Collagen, glycosaminoglycans and DDs appeared affected in acromegaly, even when the hormonal status was controlled after surgery and/or somatostatin analogue treatment. The persisting conformational change of DDs suggests a modification in cellular tensegrity, perhaps initiated by hormonal effects and probably maintained by alterations in the ECM interactions. PMID- 15667586 TI - Molecular events associated with accelerated proliferative response in rat livers when partial hepatectomy is preceded by a sham operation. AB - BACKGROUND: When a sham operation is performed 6 h before partial hepatectomy (PH), the regenerative response is accelerated suggesting that sham operation itself contributes to cellular events leading to proliferation. MATERIALS AND METHODS: In order to examine the mechanisms implicated in this acceleration, we compared the activation of several factors associated with the progression through the cell cycle at various times after PH and after PH preceded by sham operation (S6 h + PH). The effect of a single sham (S) and two combined sham operations (S6 h + S) was also examined. Nonoperated rats were used as controls (C). RESULTS: The early factors NF-kappaB and Stat3 were activated after S6 h + PH and S6 h + S. C-jun expression was increased 0.5 h and 2 h after PH and 6 h after sham. There was no further increase in S6 h + PH and S6 h + S. In contrast, c-myc expression returned to baseline levels after S6 h and a new increase was observed 2 h after S6 h + PH but not after S6 h + S. P53 mRNA was significantly expressed 6 h after S6 h + PH, but at a level similar than that observed 6 and 12 h after PH alone. An earlier increase in c-Ha-ras mRNA and cyclin E protein was found in S6 h + PH, in comparison with PH alone. CONCLUSIONS: The first divergent response between the two combined models involved c-myc expression. However, major differences related to the accelerated liver regenerative response observed after S6 h + PH were found at late time points associating an earlier expression of c-Ha-ras and nuclear cyclin E. PMID- 15667587 TI - Anti-oxidants reverse uraemia-induced down-regulation of mitochondrial membrane potential and interleukin-10 production. AB - BACKGROUND: The anti-inflammatory cytokine, interleukin-10 (IL-10), has potent immunomodulatory effects. We hypothesized that previously reported defective synthesis of IL-10 by immunocompetent cells exposed to a uraemic milieu may be due to impaired mitochondrial membrane potential (MMP). MATERIALS AND METHODS: The human promonocytic THP-1 cell line was differentiated to monocytes and incubated with pooled control or uraemic plasma with and without catalase or N acetyl L-cysteine (NAC). Basal hydrogen peroxide (H(2)O(2)) production was measured by flow cytometry. To measure MMP, cells were stained with rhodamine 123 (Rh123) and the uptake of Rh123 assessed by flow cytometry. To assess the relative contribution of the NADPH oxidase and mitochondrial electron transport chain (ELT) to endotoxin (ET)-stimulated IL-10 production among monocytic cells, cells were incubated with and without a selective NADPH oxidase inhibitor, apocynin and mitochondrial ELT inhibitors, diphenyliodinium and rotenone, washed and ET-stimulated IL-10 production was measured. In other experiments, cells were incubated with pooled control or uraemic plasma in the presence or absence of antioxidants followed by overnight incubation with ET. IL-10 production by monocytes in the cell supernatant was then quantified. RESULTS: Basal H(2)O(2) production was significantly higher among differentiated THP-1 cells exposed to uraemic plasma compared with normal plasma (180.57 +/- 10.24 vs. 41.57 +/- 8.98 MCI; P = 0.02). Uraemic plasma also down regulated MMP (4.60 +/- 1.28 vs. 8.00 +/ 1.59 MCI with normal plasma; P = 0.03). Both diphenyliodinium and rotenone, selective inhibitors of the mitochondrial ELT, inhibited ET-stimulated IL-10 production. In contrast, apocynin, a selective NADPH oxidase inhibitor, did not inhibit ET-stimulated IL-10 production. Further, ET-stimulated IL-10 production by cells incubated with uraemic plasma was significantly lower when compared to cells exposed to normal plasma. Pre-incubation with catalase and NAC restored uraemia-induced down regulation of MMP. In addition, ET-stimulated IL-10 production by cells incubated with uraemic plasma was also restored by both catalase and NAC. CONCLUSIONS: Our observations suggest that ET-stimulated IL-10 synthesis by monocytic cells is mitochondrial ELT-dependent and NADPH oxidase independent. Monocytic cells exposed to a uraemic environment exhibit higher basal ROS production, lower MMP, and impaired ET-stimulated IL-10 synthesis. Anti oxidants restore MMP and up-regulate ET-stimulated IL-10 synthesis. PMID- 15667588 TI - Effects of free fatty acids and a triglyceride-rich fat emulsion on endothelial nitric oxide synthase. PMID- 15667590 TI - Herbal or natural medicines as modulators of peroxisome proliferator-activated receptors and related nuclear receptors for therapy of metabolic syndrome. AB - The use of herbal or natural medicines for the treatment of various disorders has a long and extensive history. Many of these herbal medicines are finding their way onto the world market as alternatives to prescribed drugs currently available to treat various disorders/ailments. In particular, hyperlipidaemia is a major risk factor for atherosclerotic coronary vascular disease, which can culminate in mortality in diabetes mellitus. There is overwhelming evidence that patients with type 2 diabetes mellitus often have metabolic syndrome and require a multifactorial intervention including aggressive treatment of arterial hypertension and dyslipidaemia to prevent cardiovascular complications. One of the most active areas of metabolic research into potential treatments is in the role of nuclear receptors as therapeutic targets for both glucose and lipid metabolism. The purpose of this review is to highlight the recent advances made by pharmaceutical and research organizations in identifying biologically active compounds from natural plant products capable of modulating nuclear receptors such as peroxisome proliferator-activated receptors and, to a lesser extent, liver X receptor and farnesoid X receptor. The specific features presented by these receptors provide an in-depth insight into the pathogenesis of metabolic disease and thus, a means of establishing potential mechanisms of action with traditional medicine. In hindsight, the review offers valuable information for rational drug design using known active compounds of plant origin. Further research may ultimately lead to a reduction in both the chronic microvascular complications of type 2 diabetes mellitus and the risk of cardiovascular disease and metabolic syndrome with the use of traditional medicine. PMID- 15667591 TI - Oestrogen receptors and selective oestrogen receptor modulators: molecular and cellular pharmacology. AB - The early termination of the two arms of the Women's Health Initiative Trials has led to an increased interest and demand for selective oestrogen receptor modulators because of their potential to retain the benefits of hormone replacement therapy (oestrogen plus a gestagen) and at the same time avoid most of its severe adverse events. Selective oestrogen receptor modulators are a class of oestrogen receptor binding, small organic molecules that take advantage of the plasticity of the oestrogen receptors (alpha and beta, respectively), modulating the surface conformation of the oestrogen receptors upon binding in the respective ligand binding cavity. By doing so they affect the binding of various co-factors to the surface of the oestrogen receptors that, at least in part, explains why selective oestrogen receptor modulators may mimic the activity of oestrogen in some tissues where so desired, while opposing its activity in tissues where oestrogen-like activity is undesirable. Although selective oestrogen receptor modulators have many properties in common they also display unique activities including oestrogen receptor surface modulation and regulation of target gene expression. Selective oestrogen receptor modulators therefore offer the opportunity to develop pharmaceuticals with very distinct pharmacology and mechanism of action. Furthermore, these modulators offer the advantage of decreased risk for the development of breast and endometrial cancer and circumvent the need for combination with a gestagen. Most selective oestrogen receptor modulators in development bind with roughly equal affinity to both oestrogen receptor alpha and beta (balanced) and our view is that it is unlikely that a balanced selective oestrogen receptor modulator will inherit all desired effects of oestrogen (e.g. 17beta-oestradiol) and at the same time be devoid of all undesired effects. We therefore propose that the development of oestrogen receptor-subtype (alpha and beta, respectively) selective pharmaceuticals for specific applications (designer drugs) would better provide the benefits of hormone replacement therapy without its associated risks. PMID- 15667592 TI - In vitro assessment of renal toxicity and inflammatory events of two protein phosphatase inhibitors cantharidin and nor-cantharidin. AB - In China, cantharidin has been reported to be active against various human cancers, but with severe side effects such as nephrotoxicity. In order to reduce this toxicity, its demethylated analogue nor-cantharidin has been synthesized and used in cancer therapy, but with only few data regarding safety assessment. The aim of this study was to compare the in vitro effects of cantharidin and nor cantharidin on renal toxicity and on inflammatory events associated with tumoural process where protein phosphatases could be involved (energy status, prostanoid production, glutathione and nitrite contents) on RAW 264.7 and LLC-PK1 cells. In macrophages, both cantharidin and nor-cantharidin decreased cell viability, in a concentration- and time-dependent manner. However, IC50 was lower with cantharidin than with nor-cantharidin. These two drugs significantly decreased the ATP level after 24 hr incubation. However, ATP decreased much more with cantharidin (up to 4 times) than with nor-cantharidin. When control macrophages were activated with lipopolysaccharide+interferon-gamma for 24 hr a significant increase in nitrite content and in prostanoids were observed. Addition of either drug decreased nitrite generation and prostanoids, however these decreases were greater with cantharidin than with nor-cantharidin. In LLC-PK1 cells, incubated with either cantharidin or nor-cantharidin, our results show significant differences between the two drugs, similar to those observed in peritoneal macrophages, except for GSH content with opposite variations in both cells. We provide a better understanding of the various mechanisms of cantharidin side effects, allowing an easier comparison with nor-cantharidin which could be an attractive therapeutic potential in cancer chemotherapy in western countries. PMID- 15667593 TI - In vitro activity of phenothiazine derivatives in Enterococcus faecalis and Enterococcus faecium. AB - The antimicrobial activity of the phenothiazine derivatives thioridazine and prochlorperazine have been evaluated with 11 Enterococcus faecalis strains and 9 Enterococcus faecium strains, originating from human infections and animal faecal flora. We found that all E. faecalis and E. faecium strains, regardless of their susceptibility to commonly used antibiotics, were inhibited by thioridazine at a concentration of 16-32 microg/ml and by prochlorperazine at a concentration of 32 64 microg/ml. Combinations of the antibiotics vancomycin or ampicillin and thioridazine and prochlorperazine at subinhibitory concentrations, could render vancomycin- or ampicillin-resistant bacteria sensitive to each of the antibiotics. Verapamil and reserpine, inhibitors of P-glycoprotein-mediated multidrug resistance, did not reduce resistance. Our results outline modification of resistance in enterococci induced by phenothiazine derivatives unrelated to P glycoprotein-mediated multidrug resistance. PMID- 15667594 TI - Evaluation of chondroitin sulfate bioactivity in hippocampal neurones and the astrocyte cell line U373: influence of position of sulfate groups and charge density. AB - Chondroitin sulfates are linear polysaccharides of alternating glucuronic acid and N-acetylgalactosamine, sulfated in varying positions. They form the extracellular framework providing the information for the structural establishment of tissues in multicellular organisms. Growth cones of neurones modulate their outgrowth according to signals received from proteoglycans. The exact molecular structures behind these functions are not fully understood, but structural details of the carbohydrate backbone are crucial. In this report we have employed quantitative cytometry on hippocampal neurite outgrowth in the presence of chondroitin sulfate added in solution to determine the influence of the position and density of the sulfate groups of the N-acetyl-D-galactosamine residues of chondroitin sulfates. It is of profound interest whether externally added chondroitin sulfates can compete with core protein bound chondroitin sulfate to modulate the effects of tissue-synthesized matrix. In series of microscopic images 3 parameters of neuritic outgrowth activity, neurite length, number of neurites and fasciculation (thickness of neurites) are analyzed at concentrations occurring in intact tissues. Fasciculation increased and number of neurites decreased with high di-sulfation. No significant differences on process length reduction were found between the isotypes. Specificity of effects found is emphasized, as no influence on cell proliferation with U373 human astrocyte cell line is detectable, while neurones clearly are inhibited. The IC30 and IC50 values of chondroitin sulfates isoforms are presented for neurones. The data indicate that the soluble fragments from chondroitin sulfate are actively modulating cell development. Besides dosage, sulfation density and position are relevant for effects of chondroitin sulfate in neuronal regenerative activity. PMID- 15667595 TI - All-trans retinoic acid induction of sulfotransferases. AB - All-trans retinoic acid is the bioactive form of vitamin A (retinol). Retinoids have been used clinically as therapeutic agents against a number of cancers. Retinoids have been reported to induce the phase I drug metabolizing enzymes, cytochrome P-450s. In contrast, effects of retinoids on sulfotransferases have not been as well studied. The present investigation evaluates the role of retinoic acid on the expression of aryl sulfotransferase IV and hydroxysteroid sulfotransferase a in male and female Sprague-Dawley rat liver and intestine. Cultured human hepatic carcinoma cells (Hep G2) and intestinal carcinoma cells (Caco-2) were also used to study retinoic acid's effect on simple phenol sulfating sulfotransferase, dehydroepiandrosterone sulfotransferase and oestrogen sulfotransferase. Enzyme assay and Western blot were used to determine sulfotransferase protein expression. Retinoic acid induced aryl sulfotransferase IV in liver of female rats and sulfotransferase a in liver of male rats. Intestinal rat aryl sulfotransferase IV and sulfotransferase a in male rats and intestinal aryl sulfotransferase IV in female rats were also induced after retinoic acid treatment. In Hep G2 and Caco-2 cells, retinoic acid differentially induced the three human sulfotransferase isoforms. In general, intestinal sulfotransferases were found to be more responsive than hepatic sulfotransferases to retinoic acid treatment. mRNA expressions were investigated using reverse transcription polymerase chain reaction with gene specific primers. Reverse transcription polymerase chain reaction results are in good agreement with enzyme activity and Western blot results. This suggests that retinoic acid induction of sulfotransferases is at the transcriptional level. PMID- 15667596 TI - Effect of sodium azulene sulfonate on capsaicin-induced pharyngitis in rats. AB - Sodium azulene sulfonate is a water-soluble derivative of azulene which is an antiinflammatory component of chamomile of the family of Asteraceae. Sodium azulene sulfonate is clinically used as a therapeutic agent in the treatment of pharyngitis as well as other inflammatory diseases such as tonsillitis, stomatitis and conjunctivitis. There has been no documentation on the effect of sodium azulene sulfonate on pharyngitis in laboratory models, probably because of no availability of such models. We recently established a pharyngitis model using capsaicin application on pharyngeal mucosa in rats. The present study investigated the antipharyngitis activity of sodium azulene sulfonate comparing with those of ruthenium red (vanilloid receptor antagonist, 8.5 and 85 mg/ml), ascorbic acid (antioxidative compound, 100 microg/ml), povidone iodine (gargle as disinfectant, oxidative compound, 5 and 20 mg/ml) and diclofenac sodium (cyclooxygenase inhibitor, 0.1 and 1 mg/ml). As an antipharyngeal effect, the capsaicin-induced plasma exudation in the pharyngeal mucosa of the rat was evaluated. The capsaicin-induced plasma exudation in the pharyngeal mucosa was inhibited by sodium azulene sulfonate (100 and 200 microg/ml) as well as ruthenium red and ascorbic acid, but not by povidone iodine and dicrofenac sodium; povidone iodine rather promoted the plasma exudation. In conclusion, the antipharyngitis effect of sodium azulene sulfonate was demonstrated for the first time in a laboratory model. Although the mechanism by which sodium azulene sulfonate inhibited the capsaicin-induced pharyngitis is not yet unraveled, antioxidative effect, but not inhibitory effect on cyclooxygenase pathway, might be involved. PMID- 15667597 TI - Leishmanicidal activity of edelfosine, miltefosine and ilmofosine. AB - The anti-proliferative action of three alkyl-lysophospholipid derivatives, edelfosine (ET-OCH), miltefosine (Hexadecylphosphocholine), and ilmofosine (BM 14.440) has been studied on the promastigotes and amastigotes of Leishmania donovani. The effect of the three drugs has previously been studied, but the action mode was not clearly elucidated. In this study the effect on the intracellular amastigote forms was evaluated by two different methods: the traditional method, counting the amastigotes within the macrophages stained with Giemsa; and by a new method, staining the nuclear macrophages and amastigotes with ethidium bromide and counting the different population by flow cytometry. This new method, based on the flow cytometry, shows an advantage for evaluating the anti-proliferative effects in intracellular parasites. The ED50 were calculated for the drug activity after 72 hr, and for the three alkyl lysophospholipid derivatives it were in the range of 26.73-33.31 microM against promastigotes and in the range of 16.46-23.16 against amastigotes. Also, studying the effect against macrophages J774A1, the ED50 were in the range of 24.28-26.38 microM. The effect of the alkyl-lysophospholipids in the macromolecular biosynthesis of the Leishmania donovani, was studied comparing the incorporation of labelled analogues ([3H] thymidine, [3H] uridine and [3H] leucine), respectively, in the DNA, RNA, and proteins of the flagellates treated. Miltefosine was the most active of the alkyl-lysophospholipids, especially in the inhibition of the RNA synthesis. The three compounds studied show high in vitro activity against L. donovani promastigotes and amastigotes. PMID- 15667598 TI - Comparison of carcinogen hazard characterisation based on animal studies and epidemiology. AB - Recently we have described a simple method for quantitative risk assessment of non-threshold carcinogens based on the dose descriptor T25. In the present report quantitative hazard estimates calculated with the T25 method have been compared with results obtained using quantitative methods based on epidemiological studies. "Known" and "Likely/Probably" human carcinogens were identified from the US EPA database IRIS. In cases were the hazard characterisation was performed on the basis of epidemiological studies, the IARC monographs were used to identify animal studies by oral or inhalation exposure suitable for hazard characterisation by the T25 method. Six agents were identified: benzene, benzidine, 1,3-butadiene, cadmium, nickel subsulfide and vinyl chloride for which US EPA had made their hazard estimation based on epidemiological data. Animal data suitable for hazard characterisation were also available. For comparing hazard characterizations based on epidemiological and animal data, it was pragmatically decided to do this by comparing the chronic doses expressed as those representing a lifetime cancer hazard of 10(-3). In all cases the difference between the chronic doses determined from animal studies by the T25 method differed from those determined from epidemiological studies by a factor of less than three. Although a limited number of carcinogens were studied, the results demonstrate a very good agreement between the hazard characterisation based on epidemiological data and animal experiments over a range of more than 10(4). PMID- 15667599 TI - Population pharmacokinetics of caffeine and its metabolites theobromine, paraxanthine and theophylline after inhalation in combination with diacetylmorphine. AB - The stimulant effect of caffeine, as an additive in diacetylmorphine preparations for study purposes, may interfere with the pharmacodynamic effects of diacetylmorphine. In order to obtain insight into the pharmacology of caffeine after inhalation in heroin users, the pharmacokinetics of caffeine and its dimethylxanthine metabolites were studied. The objectives were to establish the population pharmacokinetics under these exceptional circumstances and to compare the results to published data regarding intravenous and oral administration in healthy volunteers. Diacetylmorphine preparations containing 100 mg of caffeine were used by 10 persons by inhalation. Plasma concentrations of caffeine, theobromine, paraxanthine and theophylline were measured by high performance liquid chromatography. Non-linear mixed effects modelling was used to estimate population pharmacokinetic parameters. The model was evaluated by the jack-knife procedure. Caffeine was rapidly and effectively absorbed after inhalation. Population pharmacokinetics of caffeine and its dimethylxanthine metabolites could adequately and simultaneously be described by a linear multi-compartment model. The volume of distribution for the central compartment was estimated to be 45.7 l and the apparent elimination rate constant of caffeine at 8 hr after inhalation was 0.150 hr(-1) for a typical individual. The bioavailability was approximately 60%. The presented model adequately describes the population pharmacokinetics of caffeine and its dimethylxanthine metabolites after inhalation of the caffeine sublimate of a 100 mg tablet. Validation proved the stability of the model. Pharmacokinetics of caffeine after inhalation and intravenous administration are to a large extent similar. The bioavailability of inhaled caffeine is approximately 60% in experienced smokers. PMID- 15667600 TI - Screening of antioxidants from medicinal plants for cardioprotective effect against doxorubicin toxicity. AB - Doxorubicin is an important and effective anticancer drug widely used for the treatment of various types of cancer but its clinical use is limited by dose dependent cardiotoxicity. Elevated tissue levels of cellular superoxide anion/oxidative stress are a mechanism by which doxorubicin-induced cardiotoxicity. Selected medicinal plant extracts were tested for their antioxidant capacity and cardioprotective effect against doxorubicin-induced cardiotoxicity. The cardiac myoblasts H9c2 were incubated with the antioxidants ascorbic acid, trolox, N-acetylcysteine or selected medicinal plant extracts including; 1) ethanolic extracts from Curcuma longa L-EtOH Phyllanthus emblica L EtOH, and Piper rostratum Roxb-EtOH; and 2) water extracts from Curcuma longa L H2O and Morus alba L-H2O. The cardioprotective effects of these extracts were evaluated by crystal violet cytotoxicity assay. IC50s of doxorubicin were compared in the presence or absence of ascorbic acids, trolox, N-acetylcysteine or plant extracts. Morus alba L-H2O showed the highest antioxidant properties evaluated by ferric reducing/antioxidant power assay. Ascorbic acid and N acetylcysteine had modest effects on the protection of doxorubicin-induced cytotoxicity while trolox showed insignificant protective effect. All plant extracts protected cardiac toxicity at different degrees except that Curcuma longa L-EtOH had no protective effect. Phyllanthus emblica-EtOH (100 microg/ml) showed the highest cardioprotective effect (approximately 12-fold doxorubicin IC50 increase). The data demonstrate that antioxidants from natural sources may be useful in the protection of cardiotoxicity in patients who receive doxorubicin. PMID- 15667601 TI - Preliminary study of the in vitro interaction between alcohol, high-dose flunitrazepam and its three metabolites using human liver microsomes. PMID- 15667602 TI - Intravenous iron therapy in pediatric transplant recipients: what do we really know? PMID- 15667603 TI - Extended family searches for hematopoietic stem cell donors. PMID- 15667604 TI - . . . And it drags on -- ascites after liver transplantation. PMID- 15667605 TI - Basic mechanisms of humoral rejection. AB - Humoral rejection is among the most vexing problems afflicting organ transplants. Triggered by antibodies predominantly against donor human leukocyte (HLA), humoral rejection can now be understood through consideration of basic mechanisms of immunity to foreign antigens and impact of humoral immunity on blood vessels. Basic considerations may also shed light on mechanisms by which various treatments have recently brought about vastly improved outcomes. PMID- 15667606 TI - Transplantation tolerance in pediatric recipients: lessons and challenges. AB - Clinical transplantation tolerance has remained an elusive goal in the 50 yr since it was first described in experimental animals. Greater understanding of the molecular mechanisms responsible for allorecognition have allowed for the development of promising immunosuppressive strategies that may bring us closer to reproducible induction of tolerance; consideration of past successes and failures from both clinical and basic science is required to define future challenges facing this field. This article reviews mechanisms of self and transplantation tolerance, translation of basic science research to clinical protocols in animals and human beings, the changing role of immunosuppression, complications following tolerance induction and controversies surrounding the choice of patients for tolerance trials with a focus on issues relevant to pediatric patients. The role of the Immune Tolerance Network is discussed along with realistic goals for tolerance induction in human beings over the next decade. PMID- 15667607 TI - Intra-individual variation of cystatin C and creatinine in pediatric solid organ transplant recipients. AB - There is controversy about the feasibility of cystatin C (CysC) as a marker of glomerular filtration rate (GFR) post-transplant (Tx). We studied intra-patient variability of CysC in comparison with serum creatinine (SCr) in 20 children (11 males, mean age 11.5 +/- 6.4 yr) with solid organ transplants (14 kidney, four liver, and two combined liver + kidney transplants). The mean age at Tx was 7.0 +/- 5.6 yr. A total of 178 simultaneous SCr and CysC measurements (median 8 per patient) were analyzed. In addition, GFR was calculated using the Schwartz and a novel CysC-based formula. Intra-individual coefficient of variations (CV) was calculated as ratio of standard deviation over mean. The mean CV was significantly lower for SCr (7.71 +/- 4.16%) when compared with CysC (10.27 +/- 4.87, p = 0.04), but was no longer significantly different when excluding patients with a bladder augment. The CV of the GFR estimated by Schwartz formula (7.44 +/- 3.77) was significantly lower than GFR calculated from CysC (12.52 +/- 7.37), p = 0.001. The mean ratio between the Schwartz GFR and the GFR calculated from CysC was 102.6 +/- 12.8%, not significantly different from 100% (p = 0.3796). The only potential confounding factors to explain increased CV after Tx were gender and bladder augmentation, whereas calcineurin inhibitors or steroids did not influence CV. With the limitation of a small number of subjects, our data suggest that the CysC and the CysC-calculated GFR is equivalent but not better than SCr and Schwartz formula. We therefore conclude that measurement of CysC can be used for longitudinal intra-individual follow-up of renal function post-Tx. PMID- 15667608 TI - Surgical complications in live-donor pediatric and adolescent renal transplantation: study of risk factors. AB - To report the surgical complications among our pediatric and adolescent renal transplants and to analyze the different factors that may influence the occurrence of such complications. METHODS: A total of 250 pediatric and adolescent renal transplants were included in this study. Of these patients, there were 154 boys and 96 girls with a mean age of 15.4+/-3.7 yr (range 5-20 yr). All patients received their kidneys from living donors. Four patients underwent renal retransplantation. Surgical complications were reported and their incidence was correlated with several pretransplant, technical and post transplant risk factors by both univariate and multivariate analyses. The impact of surgical complications on graft and patient survival was computed using the Kaplan-Meier technique. RESULTS: Among the 250 patients, 35 surgical complications were encountered in 33 patients. These complications included urinary leakage in 10 patients, ureteric stricture in 11 patients, complicated lymphocele in eight patients, hematoma necessitating surgical exploration in two patients, wound dehiscence in one patient, stone of the graft ureter in one patient and renal artery stenosis in two patients. The incidence of urological complications (ureteral stenosis, urinary leakage and stone disease) was 8.8% and vascular complications 0.8%. Small ureteric leakage (four patients) was treated by endourologic techniques, whereas leaks associated with ureteral necrosis required open revision. Endourologic treatment was attempted in early and mild cases of ureteric obstruction. In late and severe cases, surgery was performed. On univariate analysis, the factors that significantly affected the incidence of surgical complications were recipient's age, lower urinary tract abnormalities, the type of primary urinary recontinuity, the time to diuresis, and height and weight of the patients. On multivariate analysis, the type of primary urinary continuity was the only factor that sustained statistical significance. Neither patient nor graft survival was affected by the occurrence of surgical complications. CONCLUSION: Primary urinary continuity is the only risk factor that affects the incidence of surgical complications among pediatric and adolescent live donor renal transplants, with the extravesical technique of Lich Gregoir providing the best results. Surgical complications in pediatric and adolescent renal transplantation can be minimized if basic principles of careful transplant techniques are used. Prompt identification and treatment of any complication is paramount to graft and patient survival. PMID- 15667609 TI - Excellent outcome in infants and small children with thrombophilias undergoing kidney transplantation. AB - One of the most common causes of early graft failure in children undergoing renal transplantation is vascular thrombosis. Numerous risk factors for graft thrombosis have been previously described. Children with various types of thrombophilias such as protein C, protein S and factor V Leiden deficiencies are at an increased risk for vascular thrombosis. Infants and small children with these disorders undergoing renal transplantation have not been well documented in the literature. We reviewed our experience in the diagnosis, peri-operative management and follow up of these patients at our institution. A retrospective analysis of all children undergoing renal transplantation at our institution, using data obtained from the Pediatric Transplant Registry at our institution since May 2000 was performed. The indications for renal transplant included focal segmental glomerulosclerosis, renal dysplasia and reflux nephropathy. One patient had factor V Leiden mutation and two patients had protein S deficiency. Patients were anticoagulated in the peri-operative and post-transplant period. All index transplants were performed with living donor kidneys. There were no adverse outcomes in children with thrombophilias despite having significantly lower weight at the time of transplant vs. children without thrombophilia. The incidence of graft thrombosis in the pediatric renal transplant recipients is high. We identify a potential cause of thrombosis in children not well documented in the literature. A high index of suspicion combined with preoperative screening and diagnosis of thrombophilias and an appropriate treatment plan may decrease the incidence of graft thrombosis in infants and small children undergoing renal transplantation. PMID- 15667610 TI - Sodium ferric gluconate for post-transplant anemia in pediatric and young adult renal transplant recipients. AB - Post-transplant anemia is a widespread problem among pediatric renal transplant recipients. Many clinicians treat post-transplant anemia in a manner similar to treatment of anemia in dialysis patients, including the use of intravenous iron, such as sodium ferric gluconate (SFG). Data on pediatric dosing of SFG are limited to rare small series containing few or no renal transplant recipients. We conducted a retrospective chart review of practice patterns at our institution to evaluate doses used, hemoglobin response and adverse events. We identified 15 renal transplant recipients who received SFG during the 28-month study period. Data from 14 of these patients were available for analysis. Patients received between one and six doses of SFG to yield a total dose of 100-1000 mg or 2.7-23.7 mg/kg. The largest doses given during a single infusion ranged from 1.9 to 6.4 mg/kg. The mean hemoglobin level increased from 101 +/- 16 to 114 +/- 21 g/L (p = 0.0092) following SFG therapy. Adverse events were recorded for three patients. Treatment with SFG appears to yield some improvement in anemia in renal transplant recipients, but the paucity of published information on this topic highlights the need for stronger data, particularly with respect to pediatric patients. PMID- 15667611 TI - Liver transplantation utilizing pediatric cadaver donor livers. AB - This is a review of a unit's experience with the use of pediatric (up to 15 yr of age), cadaver donor livers for orthotopic liver transplantation (OLTx). The records of 86 patients who underwent 90 OLTx with pediatric donor allografts from January 1986 to September 2002 were reviewed. A total of 69 of 90 (77%) allografts from pediatric donors were used for pediatric (P) recipients, and 21 of 90 (23%) for adult (A) recipients. Donor demographics were: mean donor age 8.5 (0-15) yr; donor sex 53 males/37 females; cause of death head injury 55/90 (61%), cerebral ischaemia 14 of 90 (15%), cerebrovascular 13 of 90 (15%), other eight of 90 (9%). Whole OLTx were performed in 44 of 69 (64%) P group and 20 of 21 (95%) A group. Reduced-size OLTx (13 left lateral lobe, 10 left lobe, one right lobe, one split left lateral lobe) were performed in 25 of 69 (36%) P group vs. in one of 21 (5%) A group, (right lobe split) (p < 0.005). The median follow up was 8.8 yr. The graft loss in the P recipients was 19 of 69 (27.5%) vs. eight of 21 (38.1%) in A recipients. Graft loss for reduced size P OLTx was nine of 25 (36%) vs. 10/44 (22.7%) for whole allograft P OLTx (p = NS). Underlying cause of P graft loss were: chronic rejection six (32%); patient mortality six (32%); vascular four (21%); other three (15%). Underlying cause of A graft loss were: patient mortality four (50%), acute rejection two (25%) and vascular two (25%). OLTx utilizing pediatric cadaver donor allografts provides acceptable outcomes, including use of reduced-size grafts in pediatric recipients. Vascular complications and chronic rejection tend to be more common in pediatric recipients, whilst patient mortality and graft loss from acute rejection are more common in adult recipients. PMID- 15667612 TI - Extended family studies for the identification of allogeneic stem cell transplant donors in Jewish and Arabic patients in Israel. AB - HLA-identified donors are the best source of allogeneic hematopoietic stem cell transplants, and are available in approximately 40% of cases. If no HLA-identical core family member is found, an extended family search may be performed. The aim of the study was to summarize the 10-year (1990-1999) experience of our tertiary care center with extended family donor search. During this period, 356 patients and 2659 of their family members were tissue-typed; 239 patients were Jewish (67%) and 117 were Arabic (33%). An HLA-identical core-family donor was identified for 168 patients (47%): 95 Jewish (40%) and 73 Arabic (62%) (p < 0.0001); 49 patients (14%) had more than one potential donor. An extended family search (grandmother/grandfather, aunts, uncles, etc.) was performed in 38 of the remaining families, which were found to be consanguineous: five Jewish and 33 Arabic. One HLA match was found in the Jewish families (20%) and 21 in the Arabic families (64%). The odds ratio for an Arabic patient to find a donor in the extended family search was 8.75, as opposed to a Jewish patient. Overall, HLA matched donors were found by core and extended family search for 53% of the patients. The rate for Arabic patients was 80% and for Jewish patients, 40% (p < 0.001). This difference may be explained by the greater number of siblings and higher rate of consanguinity in the Arabic population. In conclusion, an extended family search for potential HLA-matched donors is worthwhile, especially in distinct ethnic populations with high consanguinity, such as Israeli Arabs. PMID- 15667613 TI - Pediatric renal transplantation: single center experience. AB - Although renal transplantation (RTx) is actually the first choice of treatment for children with end-stage renal disease, the number of transplanted children remains low in comparison with adults. The experience of the individual pediatric transplant center is very important in the outcome of pediatric transplant recipients. In this study, our pediatric renal transplantation experience is presented. We retrospectively analyzed the results of 72 pediatric renal transplants performed at Ege University Pediatric Nephrology Transplantation Center between June 1989 and May 2003. They were 40 girls, 32 boys and their mean RTx age was 13.27+/-3.73 yr (range 3-20 yr). Thirty-eight (52.8%) of the transplanted kidneys came from a living related donor, and 34 (47.2%) from a cadaveric donor. Preemptive RTx was performed in one patient and a second RTx was performed in one patient after two-period hemodialysis. Hypertension (31.9%), acute rejection (27.8%) and chronic rejection (13.9%) were the most common complications. Cytomegalovirus (CMV) infection occurred in 15 children (20.8%), none of whom died or lost their graft as a result of the infection. Pretransplant acquired hepatitis C virus (HCV) infection was detected in 12 patients (16.7%). Urinary tract infections (UTIs) were seen in 31 (43.1%) recipients. The 1, 5 and 10 yr graft survival rates were 91, 84 and 77%, respectively, and corresponding patient survival rates were 97, 84 and 77%, respectively by Kaplan-Meier method. The graft and overall survival was not correlated with sex, donor type, treatment modality, acute rejection episodes, hypertension, UTIs, CMV and HCV infection. PMID- 15667614 TI - Acute neurological complications after hematopoietic stem cell transplantation in children. AB - Children undergoing hematopoietic stem cell transplantation (HSCT) may develop toxicity-related neurological complications (NC). Known risk factors include total body irradiation (TBI) and the use of busulfan or cyclosporine A, but other risk factors might also be of importance. The medical records of 144 children (0 18 yr) who underwent their first HSCT at Karolinska University Hospital (Huddinge) between 1995 and 2002 were reviewed retrospectively concerning pretransplantation parameters and clinical course during the first 3 months after HSCT. Sibling donors were used in 49 transplantations, unrelated donors in 88 and haploidentical donors in seven cases. Nineteen patients (13%) developed NC within the first 3 months after HSCT. A significant association was seen between pretransplant viral status, defined as a higher number of positive herpes group viral serologies in the recipient before transplantation, and NC (p = 0.04). A significant association was also seen for CMV-positive recipients and NC (p = 0.01) as well as for disturbances in serum levels of sodium, potassium and calcium and NC. No association was found between sex, age at HSCT, underlying disease, previous neurological symptoms, the conditioning regimen, GVHD, donor type and NC. Number of positive herpes group viral serologies in the recipient before transplantation and certain electrolyte disturbances may contribute to neurological complications after HSCT. PMID- 15667615 TI - Extrahepatic bile duct atresia and viral involvement. AB - Several etiological factors have been suggested in the pathogenesis of extrahepatic biliary atresia (EHBA); congenital, metabolic, infectious, and multifactorial. Herein we present a study of 10 children with EHBA, the aim being to explore viral infection as a possible cause of their condition. During a period of 2 yr, all infants with EHBA were included in a study and examined on viral disease on admittance for Kasai operation. In eight of the 10 children and in one parental couple the laboratory results suggested recent or persistent viral infection. Four infections were caused by cytomegalovirus (CMV) and another five by Epstein-Barr virus (EBV). CMV-DNA was detected in two liver biopsies, EBV DNA in one liver biopsy. In a control group of 10 patients matched by age and tested by PCR in serum and viral antibodies, no sign of viral infections were detected. CMV or EBV infection were present in an unexpectedly high proportion of infants with EHBA, justifying exhaustive examination on viral disease in these children. PMID- 15667616 TI - Ascites after orthotopic liver transplantation in children. AB - Ascites is a poorly understood postoperative complication of orthotopic liver transplantation (OLT). It is associated with additional morbidity and can prolong hospitalization considerably. The incidence, the factors predictive of occurrence and the etiology of this complication are not known. The charts of 118 patients with 138 OLT were analyzed according to the following criteria: ascites lasting longer than the first 10 postoperative days, assessed by loss of ascitic fluid through drainage tubes, surgical wounds or paracentesis, with a peak volume of > or =10 mL/kg/day. Patients were divided into three groups: Group 1, no ascites; Group 2, ascites associated with postoperative complications, including chylus ascites; and Group 3, ascites not associated with postoperative complications. Postoperative ascites occurred in 43 of 138 OLT (31.2%). Patients with biliary atresia, preoperative portal hypertension, postoperative pleural effusion or at retransplantation had ascites significantly more often. In 32 of 138 (23.2%) OLT, ascites was associated with postoperative complications, including thrombosis, abdominal infections, intestinal perforation, biliary leak, pancreatitis, and chylus ascites. In 11 of 138 (7.9%) OLT, ascites was the only postoperative complication (group 3). Group three patients were significantly older, and had lower preoperative platelet counts and preoperative ascites more often than group 1 patients. The primary liver diseases were mainly cystic fibrosis of the pancreas, congenital hepatic fibrosis, and North American Indian childhood cirrhosis. The serum-ascites albumin gradient suggested a hepatic origin of ascites. Postoperative ascites is associated with the duration and degree of preoperative portal hypertension. We speculate that the mechanism involved includes a disproportion between venous blood volume and liver uptake capacity of the donor organ. PMID- 15667617 TI - Mycophenolate mofetil in pediatric renal transplantation: non-induction vs. induction with basiliximab. AB - North American Pediatric Renal Transplant Cooperative Study (NAPRTCS) reports have shown anti-T cell antibody, OKT3, to be deleterious in pediatric renal transplant recipients treated with mycophenolate mofetil (MMF). Unlike OKT3, basiliximab is a chimeric monoclonal antibody to the alpha subunit of the interleukin-2 receptor on activated T-lymphocytes. We sought to examine the outcome of MMF with or without basiliximab induction therapy in pediatric renal transplantation. Between January 1998, and June 2001, 49 pediatric renal transplants were performed at our center and 41 met the criteria for this study. We retrospectively analyzed the records of 25 patients who received MMF, Prednisone, CSA or TAC, alone (group I) and 16 patients who received MMF, CSA or TAC, and Prednisone in combination with basiliximab (group II). The two groups were similar with respect to recipient or donor age, gender, ethnicity, donor source (LD vs. CAD), cold ischemia time, and primary diagnosis. The basiliximab group had a shorter follow up period because of its more recent addition to our pediatric immunosuppression protocol, 12.9 +/- 5.9 months vs. 35.5 +/- 7.2 months for group I (p < 0.0001). At 6 months, the acute rejection rate was 16% (group I) compared with 25% (group II) (p = 0.689). The patient and graft survival at 6 and 12 months were 100% respectively for both groups. Basiliximab was well tolerated without significant adverse events. At 6 months, there was no significant difference between the groups in the incidence of urinary tract infection or cytomegalovirus infection. These data suggest that in the short-term, MMF with or without basiliximab induction therapy appears to yield excellent and statistically similar outcomes. However, further controlled studies are necessary to verify these findings as well as to define the role of basiliximab in MMF treated pediatric renal transplant recipients. PMID- 15667618 TI - Immune mechanisms in the pathogenesis of bronchiolitis obliterans syndrome after lung transplantation. AB - Lung transplantation is recognized as the only viable treatment option in a variety of end-stage pulmonary diseases. However, the long-term survival after lung transplantation is limited by the development of obliterative bronchiolitis, and its clinical correlate bronchiolitis obliterans syndrome (BOS), which is considered to represent chronic lung allograft rejection. Histopathologically, BOS is an inflammatory process that leads to fibrous scarring of the terminal and respiratory bronchioles and subsequent total occlusion of the airways. The specific etiology and pathogenesis of BOS are not well understood. The current premise is that BOS represents a common lesion in which different inflammatory insults such as ischemia-reperfusion, rejection, and infection can lead to a similar histological and clinical outcome. However, the low incidence of BOS in non-transplanted individuals and the observation that early development of BOS is predicted by the frequency and severity of acute rejection episodes indicate that alloimmune-dependent mechanisms play a crucial role in the pathogenesis of BOS. The evidence presented in this review indicates that BOS is the result of humoral and cellular immune responses developed against major histocompatibility complex molecules expressed by airway epithelial cells of the lung allograft. This process is aggravated by alloimmune-independent mechanisms such as ischemia reperfusion and infection. Currently, treatment of BOS is frequently unsuccessful. Therefore, a better understanding of the immunopathogenesis of BOS is of paramount importance toward improving long-term patient and graft survival after lung transplantation. PMID- 15667619 TI - Severe neutropenia and thrombocytopenia following cardiac transplantation in a child. AB - Hematological complications have been occasionally described after cardiac transplantation. We are reporting a 5-yr-old child who developed sequential severe neutropenia and thrombocytopenia following cardiac transplantation while on tacrolimus-based immune suppression therapy. There was no improvement in blood counts following a change in immune suppression to cyclosporine A. The neutropenia was associated with a maturation arrest in the bone marrow. The occurrence of thrombocytopenia coincided with rising anti-herpes virus 6 IgG titers suggesting a possible contributory role. Neutropenia resolved following treatment with rituximab, and the thrombocytopenia responded to Dapsone therapy eventually. This case points out the potential multifactorial pathogenesis of cytopenias following cardiac transplantation with differing response to various immune suppressive therapies. PMID- 15667620 TI - Gastrointestinal leukocytoclastic vasculitis: an adverse effect of sirolimus. AB - An 18-yr-old Hispanic female with end-stage renal disease secondary to chronic glomerulonephritis of unknown etiology underwent cadaveric renal transplantation. She was placed on a steroid-free protocol with tacrolimus and mycophenolate mofetil (MMF) for maintenance immunosuppression. Approximately 8 months post transplantation, MMF was replaced by sirolimus (SRL) because of persistent leukopenia. Four months after the initiation of SRL, the patient began to experience chronic, constant periumbilical abdominal pain in the absence of vomiting, diarrhea or melena. Esophagogastroduodenoscopy and CT scans revealed significant diffuse mucosal thickening of the antrum, duodenum, and jejunum; leukocytoclastic vasculitis was identified on antral biopsy. A repeat biopsy after reduction of sirolimus dose by 50% over 6 months showed mild chronic inflammation of stomach and duodenum with some improvement in abdominal pain. Discontinuation of SRL and replacement with low dose MMF resulted in complete resolution of pain and normalization of gastrointestinal anatomy by imaging studies within 2 months. In light of this report, drug-induced leukocytoclastic vasculitis caused by SRL should be considered in the differential diagnosis of chronic abdominal pain in a patient with organ transplantation. PMID- 15667621 TI - Successful bone marrow transplantation in a patient with c-mpl-mutated congenital amegakaryocytic thrombocytopenia from a carrier donor. AB - Congenital amegakaryocytic thrombocytopenia (CAMT) is characterized by severe thrombocytopenia and the absence of megakaryocytes in bone marrow. Furthermore, mutation of the c-mpl gene has been identified as a cause of this disorder. The only curative treatment is allogeneic stem cell transplantation (SCT). The current report describes a patient exhibiting c-mpl mutation in both alleles who underwent transplantation of allogeneic bone marrow donated by her brother, a c mpl mutated carrier, employing a fludarabine-based conditioning regimen. Engraftment and reconstitution of hematopoietic cells was rapid and without complications. These findings suggest that the carrier donor displaying the c-mpl mutation can serve as a donor source for SCT. PMID- 15667622 TI - Improved outcome of pediatric kidney transplantations in the Netherlands -- effect of the introduction of mycophenolate mofetil? AB - Collaboration of the Dutch centers for kidney transplantation in children started in 1997 with a shared immunosuppressive protocol, aimed at improving graft survival by diminishing the incidence of acute rejections. This study compares the results of transplantations in these patients to those in a historical reference group. Ninety-six consecutive patients receiving a first kidney transplant were treated with an immunosuppressive regimen consisting of mycophenolate mofetil, cyclosporine and corticosteroids. The results were compared with those of historic controls (first transplants between 1985 and 1995, n = 207), treated with different combinations of corticosteroids, cyclosporine A and/or azathioprine. Cytomegalovirus (CMV) prophylaxis was prescribed to high-risk patients in the study group, and only a small proportion of the reference group. The graft survival at 1 yr improved significantly: 92% in the study group, vs. 73% in the reference group (p < 0.001). In the study group 63% of patients remained rejection-free during the first year; in the reference group 28% (p < 0.001). After statistical adjustment of differences in baseline data, as cold ischemia time, the proportion of LRD, preemptive transplantation, and young donors, the difference between study and reference group in graft survival (RR 0.33, p = 0.003) and incidence of acute rejection (RR 0.37, p < 0.001), as the only factor, remained statistically significant, indicating the effect of the immunosuppressive therapy. In the first year one case of malignancy occurred in each group. CMV disease occurred less frequently in the study group (11%) than in the reference group (26%, p = 0.02). As a new complication in 4 patients bronchiectasis was diagnosed. A new consensus protocol, including the introduction of mycophenolate mofetil, considerably improved the outcome of pediatric kidney transplantation in the Netherlands, measured as reduction of the incidence of acute rejection and improved graft survival. PMID- 15667623 TI - Fulminant hepatic failure: Wilson's disease or autoimmune hepatitis? Implications for transplantation. AB - Fulminant hepatic failure (FHF) accounts for 10-15% of pediatric liver transplants in the USA annually. Because the onset of FHF may be the first presentation of Wilson's disease (WD) and autoimmune hepatitis (AIH) in previously asymptomatic adolescents, determination of the etiology of FHF is critical as treatment and prognosis differ between these two entities. Patients with AIH may be salvaged by medical treatment. On the contrary, liver transplantation is currently the only life saving therapeutic option available for patients with WD who present with fulminant liver failure. To establish the diagnosis of WD and AIH in the setting of FHF remains challenging for diagnosticians and decisions regarding liver transplantation may be necessary before a diagnosis is firmly established. We report a previously asymptomatic patient who presented with FHF and clinical and laboratory features suggestive of both WD and AIH and who underwent successful therapeutic liver transplantation before the diagnosis of WD could be confirmed. PMID- 15667624 TI - Reversal of tacrolimus-related hypertrophic obstructive cardiomyopathy 5 years after kidney transplant in a 6-year-old recipient. AB - Hypertrophic obstructive cardiomyopathy (HOCM) is an unusual but serious side effect of tacrolimus (TAC) based immunosuppression primarily affecting pediatric patients after solid organ transplant. TAC-induced HOCM has already been described in patients after liver, bowel and heart transplant shortly after the procedure. Herein, we present the first case report of TAC-induced HOCM in a young renal transplant recipient 5 yr after renal transplant. The condition was diagnosed by ECHO and EKG and successfully treated by discontinuation of TAC followed by conversion to cyclosporine (CsA). PMID- 15667625 TI - Low toxicity and efficacy of (153)samarium-EDTMP and melphalan as a conditioning regimen for secondary acute myelogenous leukemia. AB - We report the case of a 15-yr-old girl who developed secondary acute myelogenous leukemia (AML) 4 yr after completion of therapy for metastatic Ewing sarcoma (primary right acetabulum with metastatic disease to the lungs). Peripheral blood stem cells were collected after the second cycle of chemotherapy with the plan for future consolidation with high-dose chemotherapy and autologous stem cell rescue; however, because of the patient's excellent response to chemotherapy and surgery, therapy was completed without the need for high-dose chemotherapy. No human leukocyte antigen (HLA)-matched related donor was available for a bone marrow transplant. Because of previous lung radiation, high-dose samarium [30 mCi/kg of samarium-153 ethylenediaminetetramethylenephosphonate ((153)Sm-EDTMP) day -14] and melphalan (140 mg/m(2) day -2) were chosen as the conditioning regimen to avoid potential lung complications. The patient received an infusion of 6.1 x 10(8)/kg mononuclear autologous cells on day 0. She achieved engraftment on day +23. Three years after transplantation, she continues to have complete remission. Samarium and melphalan constitute a well-tolerated regimen with potential antileukemic activity. PMID- 15667626 TI - Development of hepatopulmonary syndrome and portopulmonary hypertension in a paediatric liver transplant patient. AB - BACKGROUND: Hepatopulmonary syndrome (HPS) and portopulmonary hypertension (PPH) are pulmonary vascular disorders which occur in patients with severe liver disease and/or portal hypertension. Although these syndromes are frequently diagnosed in patients undergoing assessment for liver transplantation, they seldom occur in the same patient. METHOD: This report describes a female paediatric patient, born with extra-hepatic biliary atresia, who required liver transplantation, at the age of 15, for secondary biliary cirrhosis. She had severe HPS prior to her first liver transplant, which resolved rapidly following surgery, as well as indirect evidence for PPH. She required a second liver transplant 1 yr later for chronic rejection. Whilst evaluating the patient for a third liver transplant, 4 yr later, severe PPH was discovered. The patient died 3 months later from right heart failure. CONCLUSION: HPS and PPH may coexist however they may show differing responses to liver transplantation with progression of PPH despite the resolution of HPS. PMID- 15667627 TI - Re: Trial of metronidazole vs. azithromycin for treatment of cyclosporine-induced gingival overgrowth. PMID- 15667628 TI - The effect of fibre insertion on fracture resistance of root filled molar teeth with MOD preparations restored with composite. AB - AIM: To evaluate the effect of using flowable composite with or without leno woven ultra high modulus polyethylene fibre reinforcement on fracture resistance of root filled mandibular molars with mesio-occlusodistal (MOD) preparations. METHODOLOGY: Sixty sound extracted human mandibular molars were randomly assigned to five groups (n = 12). Group 1 did not receive any preparation. From groups 2 to 5, the teeth were root filled and MOD preparations were created. Group 2 remained unrestored. Group 3 was restored with a dentine bonding system (DBS; SE Bond, Kuraray, Japan) and composite resin (CR) (AP-X; Kuraray). In group 4, flowable composite resin (Protect Liner F; Kuraray) was used before restoring teeth with CR. In group 5, leno woven ultra high modulus polyethylene ribbon fibre (Ribbond, Seattle, WA, USA) was inserted into the cavities in a buccal to lingual direction and the teeth were then restored with DBS and CR. After finishing and polishing, the specimens were stored in 100% humidity at 37 degrees C for 1 day. Compressive loading of the teeth was performed using a universal testing machine at a crosshead speed of 0.5 mm min(-1). The mean load necessary to fracture the samples were recorded in newtons (N) and were subjected to analysis of variance (ANOVA) and Tukey post-hoc test. RESULTS: The mean load necessary to fracture the samples in each group were (in N): group 1: 1676.75 +/ 154.63(a), group 2: 376.51 +/- 37.36(b), group 3: 733.23 +/- 133.33(c), group 4: 786.48 +/- 145.34(c), group 5: 943.63 +/- 121.15(d). There were statistically significant differences between the groups annotated with different letters. CONCLUSIONS: (i) Use of flowable composite resin under composite restorations had no effect on fracture resistance of root filled molar teeth with MOD preparations, (ii) use of polyethylene ribbon fibre under composite restorations in root filled teeth with MOD preparations significantly increased fracture strength. PMID- 15667629 TI - Periapical status and quality of endodontic treatment in an adult Irish population. AB - AIM: To determine the prevalence of apical periodontitis and the quality of root fillings in an adult Irish population using a retrospective analysis of orthopantomograms (OPGs). METHODOLOGY: A systematic sample of clinical records and OPGs of 302 adult patients attending the Dublin Dental Hospital, Ireland, were screened by two examiners to determine the quality of root canal treatment and the prevalence of apical periodontitis. The operators who carried out the treatment were unknown. Two examiners inspected OPGs after inter-examiner correlation. European Society of Endodontology (ESE) guidelines were used to determine adequacy of root treatment. RESULTS: Of the 7427 teeth examined 2% had root fillings. Apical periodontitis was evident in 1.6% of all nonroot filled teeth whilst 33.1% of the subjects had at least one tooth with apical periodontitis. Of the root filled teeth, 25% had apical periodontitis and 52.6% were considered technically inadequate by ESE guidelines. There was a statistically significant (P < 0.05) negative correlation between the quality of the root fillings and the prevalence of apical periodontitis. Posterior root filled teeth (premolars and molars) had a greater prevalence of apical periodontitis than anterior root filled teeth. CONCLUSIONS: The technical quality of root fillings in an adult Irish population was poor and was consistent with a high prevalence of apical periodontitis. PMID- 15667630 TI - An evaluation of .06 tapered gutta-percha cones for filling of .06 taper prepared curved root canals. AB - AIM: To compare the area occupied by gutta-percha, sealer, or void in standardized .06 tapered prepared simulated curved canals and in mesio-buccal canals of extracted maxillary first molars filled with a single .06 gutta-percha point and sealer or lateral condensation of multiple .02 gutta-percha points and sealer. METHODOLOGY: Simulated canals in resin blocks with either a 30 degrees curve and radius of 10.5 mm (n = 20) or a 58 degrees curve and 4.7 mm radius (n = 20) and curved mesio-buccal canals of extracted maxillary first molars (n = 20) were prepared using .06 ProFiles in a variable tip crown-down sequence to an apical size 35 at 0.5 mm from the canal terminus or apical foramen. Ten 30 degrees and 58 degrees curved resin canals and 10 canals in the extracted teeth group were obturated with .02 taper gutta-percha cones and AH 26 sealer using lateral condensation. The time required to obturate was recorded. The remaining canals were obturated with a single .06 taper gutta-percha cone and AH 26 sealer. Excess gutta-percha was removed from the specimens using heat and the warm mass vertically condensed. Horizontal sections were cut at 0.5, 1.5, 2.5, 4.5, 7.5 and 11.5 mm from the canal terminus or apical foramen. Colour photographs were taken using an Olympus 35 mm camera attached to a stereomicroscope set at x40 magnification, and then digitized using a flatbed scanner. The cross-sectional area of the canal contents was analysed using Adobe PhotoShop. The percentage of gutta-percha, sealer or voids to the total root canal area were derived and data analysed using unpaired Student's t-test and the Mann-Whitney U-test. RESULTS: In the 30 degrees curved canals the levels had between 94 and 100% of the area filled with gutta-percha with no significant difference (P > 0.05) between the lateral condensation and single cone techniques. In the 58 degrees curved canals the levels had 92-99% of the area filled with gutta-percha, with the single cone technique having significantly (P < 0.05) more gutta-percha fill at the 2.5 mm level only. In the mesio-buccal canals of the teeth the levels had between 72 and 96% of the area filled with gutta-percha with no significant difference (P > 0.05) between the lateral condensation and single cone technique. The time for obturation was significantly (P < 0.05) greater for lateral condensation compared with the single cone technique in all groups. CONCLUSIONS: The .06 taper single cone technique was comparable with lateral condensation in the amount of gutta percha occupying a prepared .06 tapered canal. The .06 single cone technique was faster than lateral condensation. PMID- 15667631 TI - Influence of irrigant needle depth in removing bioluminescent bacteria inoculated into instrumented root canals using real-time imaging in vitro. AB - AIM: To test the hypothesis that the mechanical efficacy of irrigation in reducing bacteria in the root canal is dependent on depth of placement of the irrigation needle. METHODOLOGY: The root canals of 30 permanent cuspids were instrumented to apical size 60 using a crown-down technique. A suspension of the bioluminescent reporter strain Pseudomonas fluorescens 5RL was inoculated into each canal of sterilized teeth. Emission of bioluminescence (photons s(-1)) from each tooth was quantified on four sequential occasions using luminometry and bioluminescence imaging: (i) background, (ii) after inoculation, (iii) after irrigating the inoculated teeth with 3 mL of a nonantimicrobial irrigant delivered either 1 mm (group 1, n = 15) or 5 mm (group 2, n = 15) from working length (WL) using a 28G safety-ended irrigating needle, (iv) after an additional 3 mL irrigation (total 6 mL). Intragroup and intergroup comparisons were made using Wilcoxon matched pairs and Mann-Whitney tests, respectively. RESULTS: In group 1, there was a mean log10 decrease in bacteria of 0.68 +/- 0.26 after 3 mL of irrigant compared with 1.19 +/- 0.48 after 6 mL (P < 0.001); in group 2 the mean log10 decrease was 0.58 +/- 0.28 after 3 mL of irrigant compared with 0.69 +/- 0.35 after 6 mL (P < 0.02) (Wilcoxon matched pairs). Using 3 mL of irrigant, needle depth did not have a significant effect on reduction of intracanal bacteria (P = 0.407), but the effect became significant when 6 mL of irrigant was used (P < 0.002) (Mann-Whitney tests). CONCLUSIONS: The mechanical efficacy of 6 mL of irrigant in reducing intracanal bacteria was significantly greater when delivered 1 mm compared with 5 mm from WL. PMID- 15667632 TI - Frequency and distribution of root filled teeth and apical periodontitis in a Greek population. AB - AIM: To investigate the prevalence of root filled teeth and apical periodontitis (AP) in a Greek population. METHODOLOGY: A random sample of 320 patients who required full mouth periapical radiographic examination as a part of diagnostic and planning procedures were included. The age of the patients ranged from 16 to 77 years. A total of 7664 teeth were assessed and the frequency of root filled teeth and periapical status was recorded. Two observers evaluated the radiographs under standardized conditions. AP was defined as distinct periapical radiolucency or widening of the periodontal ligament space exceeding two times the normal width. Statistical evaluation of differences in proportions between groups was performed using random effects logistic regression models. RESULTS: The periapical status of 286 (3.7%) teeth was impossible to evaluate because of radiographic faults; these teeth were excluded from further analysis. A total of 1040 (13.6%) teeth had radiographic signs of AP and 680 (9.2%) teeth had been root filled. Of the root filled teeth, 408 (60.0%) had AP. There was no difference in the number of root filled teeth between males and females; the prevalence of root filled teeth increased with age. Significantly more molars (13.1%) and premolars (11.9%) than anterior teeth (5.8%) had been root filled (P < 0.001). The prevalence of AP was significantly higher (P < 0.001) in molars (23.9%) and premolars (14.0%) than anterior teeth (9.4%). CONCLUSIONS: The prevalence of AP and the frequency of root filled teeth with AP in this Greek population were higher than those found in many other European countries. The frequency of root filled teeth was comparable with findings in other epidemiological studies. PMID- 15667633 TI - Probability of removing fractured instruments from root canals. AB - AIM: To evaluate in a clinical case series the location of fractured instruments, how many of them could be removed and to compare these findings with the results of a similar study. METHODOLOGY: Within an 18-month period all referred endodontic cases involving fractured instruments within root canals were analysed. The protocol for removal of fractured instruments was: create straight line access to the coronal portion of the fractured instrument, attempt to create a ditched groove around the coronal aspect of the instrument using ultrasonic files and/or to bypass it with K-Files. Subsequently, the fractured instrument was vibrated ultrasonically and flushed out of the root canal or an attempt was made to remove the instrument with the Tube-and-Hedstrom file-Method or similar techniques. The location of the fractured instrument and the time required for removal were recorded. Successful removal was defined as complete removal from the root canal without creating a clinically detectable perforation. RESULTS: In total, 97 consecutive cases of instrument fracture were included in the time period. In all, 84 instruments (87%) were removed successfully. There was a significant correlation between the time needed to remove fractured instruments and a decrease in success rate. Curved canals had significantly more fractured instruments than straight canals (P < 0.05). Rotary instruments fractured significantly more often in curved canals (P < 0.05) compared with other instruments. Half of all instrument fractures occurred in mesial roots of lower molars and most often when using rotating instruments. There was no statistically significant difference in the success rate with respect to the location of the fractured instrument (tooth/root type), the type of fractured instrument or the different methods of instrument removal. CONCLUSIONS: Curved canals are a higher risk for instrument fracture than straight canals. In curved canals rotary instruments (including lentulo spirals) fractured more often than other instruments. In all, 87% of the fractured instruments were removed successfully. A decrease in success rate was evident with increasing treatment time. The use of an operating microscope was a prerequisite for the techniques used to remove the fractured instruments. PMID- 15667634 TI - Detection of the second mesiobuccal canal in mesiobuccal roots of maxillary molar teeth ex vivo. AB - AIM: To assess the effectiveness of magnification and dentine removal (troughing) when locating the second mesiobuccal canal in mesiobuccal roots of maxillary molars. METHODOLOGY: A total of 208 extracted human maxillary molars were examined. After crown and pulp removal, the MB1 and 2 canals in the mesiobuccal root were located in three stages that were performed by two undergraduate dental students. Stage 1: canals were located with an endodontic explorer; stage 2: additional canals in the same teeth were located under magnification with a digital microscope (VH-8000, Keyence, Japan); stage 3: additional canals in the same teeth were located by removing dentine (troughing) from the pulp chamber floor within 3 mm from MB1 canal towards the palatal canal with an Enac ultrasonic tip (ST 21, Osada, Japan). In each group, the canals were prepared with Gates Glidden burs and K-files. The distal and palatal roots were then removed, and Indian ink was injected into the canal system within the mesio buccal root. The root surfaces were washed with 6% NaOCl, and then rendered transparent to observe canal morphology. The root canal configurations were classified into five categories following the modified Weine's classification. RESULTS: More than one canal in the mesio-buccal root was observed in 48% of specimens. Detection rates of multiple canals were 7, 18 and 42% following stages 1, 2 and 3, respectively. There was a significant difference between the stages for detecting the MB2 canal (P < 0.05, Friedman test). CONCLUSIONS: Both magnification (stage 2) and dentine removal under magnification (stage 3) were effective in detecting the presence of the MB2 canal. However, MB2 canals could not be detected in 13% of the teeth because of canal calcification or branching located more apically. PMID- 15667635 TI - Longitudinal study on microleakage of three root-end filling materials by the fluid transport method and by capillary flow porometry. AB - AIM: (i) To compare the root-end sealing ability of IRM Caps (IRM), Fuji IX Capsules (Fuji IX) and Pro Root MTA Tooth-Coloured Formula (MTA) in teeth obtained from cadavers. (ii) Further research on leakage study methodology by means of comparison of the fluid transport method (FTM) and capillary flow porometry (CFP). METHODOLOGY: Root canal treatment was performed on 33 cadaver teeth in situ 2 weeks prior to root resection and ultrasonic retropreparation (S12/90 degrees D-tip on Suni-Max), after which the teeth were retrieved from the cadavers. Two teeth were kept as positive and negative controls. The other teeth were divided in three different groups at random, with each group receiving one of the retrofill materials. Retrofills were exposed to water 5 min after placement. The teeth were stored at 37 degrees C for 12 h after which the root filling was removed. Microleakage (L in microL day(-1)) was measured for 24 h under a pressure of 1.2 atm using FTM and recorded as L = 0, 0 < L < or = 10, L > 10. The measurements were repeated after 1 and 6 months. After 6 months, leakage was also assessed by CFP in order to measure through pores and their diameters. Results were analysed statistically using nonparametric Kruskal-Wallis and Mann Whitney U-tests, and Spearman correlation coefficients between the results of both methods were calculated. The level of significance was set at 0.05. RESULTS: (i) A statistically significant difference could be demonstrated between Fuji IX and IRM at 1 month with FTM. FTM revealed a significant difference between Fuji IX and the other materials at 6 months, whereas CFP did not. However, using both methods, Fuji IX showed the best result. (ii) When comparing both techniques, CFP demonstrated through pores in all teeth, whereas with FTM in only 14 of the 31 teeth could through pores be demonstrated. A positive correlation between both methods was demonstrated. CONCLUSIONS: Under the conditions of this study (i) the conventionally setting glass-ionomer cement Fuji IX showed the best results when used as a root-end material and (ii) CFP appeared to be a useful method for leakage evaluation of through pores in endodontics. PMID- 15667636 TI - Effect of ProRoot MTA mixed with chlorhexidine on apoptosis and cell cycle of fibroblasts and macrophages in vitro. AB - AIM: To compare the percentage of apoptotic cells and the cell cycle profile of fibroblasts and macrophages exposed to either ProRoot mineral trioxide aggregate (MTA) mixed with chlorhexidine (CHX), or exposed to ProRoot MTA mixed with sterile water. METHODOLOGY: Mouse gingival fibroblasts or mouse macrophages were seeded in six-well plates and allowed to attach overnight. Freshly mixed or set (allowed to dry for 24 h) specimens of tooth-coloured (white) ProRoot MTA were prepared with 0.12% CHX gluconate (MTA/CHX) or with sterile water (MTA/H2O). The cells were exposed for 24 h to the MTA specimens, which were placed over permeable membrane inserts to avoid direct contact with the cells. Untreated cells served as controls. Propidium iodide staining followed by flow cytometry was used to evaluate the effects of ProRoot MTA on cell apoptosis and cell cycle. Statistical analyses were performed by one-way anova followed by post-hoc tests with the use of the SigmaStat 2.0 software, and significance was determined at P < or = 0.05. RESULTS: MTA specimens containing CHX induced apoptosis of macrophages and fibroblasts (P < 0.05). In contrast, no change in the proportion of apoptotic cells was observed when sterile water was used to prepare the specimens (P > 0.05). Cell cycle analysis showed that exposure to MTA/CHX decreased the percentage of fibroblasts and macrophages in S phase (DNA synthesis) as compared with exposure to MTA/H2O (P < 0.05). CONCLUSION: This in vitro study demonstrated that the substitution of CHX for sterile water in MTA increases its cytotoxicity. This suggests that the potentially beneficial antimicrobial effect of CHX may be accompanied by an increase in the cytotoxicity of the resulting MTA-based material. PMID- 15667638 TI - What price education? PMID- 15667639 TI - Skeletal effects of bite jumping therapy on the mandible - removable vs. fixed functional appliances. AB - AUTHORS: Shen G, Hagg U, Darendeliler MA. OBJECTIVE: Based on an extensive review of the literature, the aim of this study was to explore the mainstream consensus on the controversial topic of whether the bite jumping treatment could enhance mandibular growth. DESIGN: The data for removable and fixed functional appliances were respectively comprehended and analyzed with regard to their attributes in mandibular growth modification. Furthermore, numerous reported findings were assessed by relating them to some important factors influencing the effects of bite jumping, such as treatment timing, treatment duration and post-treatment follow-up, to allow for a more objective and accurate evaluation. RESULTS: The key differences between removable and fixed appliances are working hours (intermittent vs. continuous), length of treatment time (long vs. short), optimal treatment timing (before puberty growth vs. at or after puberty spurt), and mode of bite-jumping (considerable vertical opening vs. limited vertical opening). These different features lead to different treatment effects on mandibular and TMJ growth, such as the intensity of possibly increased growth (clinically less significant vs. significant), the direction of enhanced growth (vertical vs. horizontal), and the stability of treatment changes (unstable vs. stable). The short-term or long-term post-treatment relapse mainly relates to the rebound of dental position. CONCLUSION: The immediate effects of bite jumping functional appliances on the mandibular growth enhancement are convincing during actual treatment. This extra gain of growth might be sustainable during the short-term and long-term post-treatment period. PMID- 15667640 TI - The role of type X collagen in facilitating and regulating endochondral ossification of articular cartilage. AB - AUTHOR: Shen G Objective -This review was compiled to explore the role of type X collagen in growth, development and remodeling of articular cartilage by elucidating the linkage between the synthesis of this protein and the phenotypic changes in chondrogenesis and the onset of endochondral ossification. DESIGN: The current studies closely dedicated to elucidating the role of type X collagen incorporating into chondrogenesis and endochondral ossification of articular cartilage were assessed and analyzed to allow for obtaining the mainstream consensus on the bio-molecular mechanism with which type X collagen functions in articular cartilage. RESULTS: There are spatial and temporal correlations between synthesis of type X collagen and occurrence of endochondral ossification. The expression of type X collagen is confined within hypertrophic condrocytes and precedes the embark of endochondral bone formation. Type X collagen facilitates endochondral ossification by regulating matrix mineralization and compartmentalizing matrix components. CONCLUSION: Type X collagen is a reliable marker for new bone formation in articular cartilage. The future clinical application of this collagen in inducing or mediating endochondral ossification is perceived, e.g. the fracture healing of synovial joints and adaptive remodeling of madibular condyle. PMID- 15667641 TI - Gatekeepers of scholarship. PMID- 15667642 TI - Transverse maxillary deficiency in Class II and Class III malocclusions: a cephalometric and morphometric study on postero-anterior films. AB - AUTHORS: Franchi L, Baccetti T. OBJECTIVES: The aim of the present study is to evaluate the dentoskeletal features of subjects with either Class II or Class III malocclusions in the mixed dentition using both conventional cephalometric analysis and TPS morphometric analysis applied to posteroanterior (PA) cephalograms. DESIGN: TPS analyses of PA cephalograms on 49 Cl-II, and 20 Cl-III subjects. Tracings were done by hand. SETTING AND SAMPLE POPULATION: The Department of Orthodontics, University of Florence. OUTCOME MEASURE: Size and shape differences between Cl-II and Cl-III malocclusions. RESULTS: Maxillary width was smaller in both Cl-II and Cl-III subjects compared with normal as measured conventionally. The TPS analysis revealed transverse plane compression and extension in the vertical plane. CONCLUSION: In Cl-II and Cl-III subjects the maxillary width was smaller 2.5 and 4 mm, respectively. TPS analyses corroborate these findings. PMID- 15667643 TI - Bilaterally impacted maxillary canines and multiple missing teeth: a challenging adult case. AB - AUTHORS: Mesotten K, Naert I, van Steenberghe D, Willems G. OBJECTIVES: Orthodontic correction of bilateral maxillary canine impaction with agenesis of the lower second premolars and extraction of the lower first molars in an adult patient. METHODS: After surgical exposure, the canines were relocated in the arch by means of immediate orthodontic traction. Endosseous Branemark system implants were inserted in the lower jaw with a double purpose: stabilization of orthodontic anchorage and prosthetic tooth replacement. RESULTS: Closure of the edentulous space in the lower arch was accomplished by a multidisciplinary approach while orthodontic correction of the initial malocclusion was achieved. CONCLUSION: Implants provided anchorage control for tooth movement and created the possibility of prosthetic rehabilitation through a multidisciplinary treatment approach. PMID- 15667645 TI - Myelomeningocele in young adults. PMID- 15667644 TI - Apoptosis in the regenerate produced by mandibular osteodistraction in the mature rat. AB - AUTHORS: Liu ZJ, Anderson MW, Gu GM, King GJ. OBJECTIVE: Little is known about the contribution of apoptosis to the formation of the regenerate in distraction osteogenesis. This study investigated the role of apoptosis in the regulation of osteogenesis in relation to mandibular distraction rate and recovery time. METHODS: One hundred and twenty-nine 3-month rats received unilateral mandibular ramus osteotomy and distraction device placement. After 3-day latency, these were distracted for 5 days. The slow group was distracted a total of 1 mm (0.2 mm/day), the moderate 2 mm (0.4 mm/day), the rapid 3 mm (0.6 mm/day), and the sham was not distracted (0.0 mm/day). Eight to nine rats from each group were killed at 6 (mid-distraction), 10 (early-consolidation), 24 (mid-consolidation) and 38 (late-consolidation) days following device placement. Baseline data were obtained from an additional eight rats killed at day 3 (end of latency). Sagittal sections (7 mum) of the harvested hemi-mandibles were embedded in paraffin, double-stained with the DeadEnd(TM) Colorimetric TUNEL system for apoptotic cells and tartrate-resistant acid phosphatase for osteoclasts. Cell counts of apoptotic cells and osteoclasts (apoptotic or otherwise) were preformed at 40x magnification using a calibrated grid at the middle regions of the caudal, central and rostral parts of the distraction gap. Counting reproducibility was verified as <13%. RESULTS: Counts from all three regions were combined because no significant regional difference was found. The majority of the fibrous matrix regenerate was seen at days 6 and 10 while osseous regenerate occurred mainly at days 24 and 38. Significantly higher levels of apoptotic activity were seen at day 24. Apoptotic osteoclasts also peaked at day 24. Apoptotic cell numbers in the slow and moderate groups most closely followed the pattern of the sham in which the apoptosis activity peaked at days 10 or 24. However, those in the rapid group showed delayed peaks at days 24 or 38. CONCLUSIONS: Chalmers Street, Surry Hill, NSW 2010, Australia. gshe6437@mail.usyd.edu.au The transition from fibrous to osseous regenerate during mandibular distraction osteogenesis is accompanied by an increase in cell clearance via apoptosis. A slow to moderate distraction rate allows for the most typical pattern of bone healing while a rapid rate prolongs the healing process. PMID- 15667646 TI - Prevalence and characteristics of screen-detected prostate carcinomas at low prostate-specific antigen levels: aggressive or insignificant? AB - Screening for prostate cancer at low prostate-specific antigen (PSA) levels (